PMID- 15267476 TI - Calculating potentials of mean force from steered molecular dynamics simulations. AB - Steered molecular dynamics (SMD) permits efficient investigations of molecular processes by focusing on selected degrees of freedom. We explain how one can, in the framework of SMD, employ Jarzynski's equality (also known as the nonequilibrium work relation) to calculate potentials of mean force (PMF). We outline the theory that serves this purpose and connects nonequilibrium processes (such as SMD simulations) with equilibrium properties (such as the PMF). We review the derivation of Jarzynski's equality, generalize it to isobaric- isothermal processes, and discuss its implications in relation to the second law of thermodynamics and computer simulations. In the relevant regime of steering by means of stiff springs, we demonstrate that the work on the system is Gaussian distributed regardless of the speed of the process simulated. In this case, the cumulant expansion of Jarzynski's equality can be safely terminated at second order. We illustrate the PMF calculation method for an exemplary simulation and demonstrate the Gaussian nature of the resulting work distribution. PMID- 15267477 TI - Spectral difference Lanczos method for efficient time propagation in quantum control theory. AB - Spectral difference methods represent the real-space Hamiltonian of a quantum system as a banded matrix which possesses the accuracy of the discrete variable representation (DVR) and the efficiency of finite differences. When applied to time-dependent quantum mechanics, spectral differences enhance the efficiency of propagation methods for evolving the Schrodinger equation. We develop a spectral difference Lanczos method which is computationally more economical than the sinc DVR Lanczos method, the split-operator technique, and even the fast-Fourier Transform Lanczos method. Application of fast propagation is made to quantum control theory where chirped laser pulses are designed to dissociate both diatomic and polyatomic molecules. The specificity of the chirped laser fields is also tested as a possible method for molecular identification and discrimination. PMID- 15267478 TI - A state-specific approach to multireference coupled electron-pair approximation like methods: development and applications. AB - The traditional multireference (MR) coupled-cluster (CC) methods based on the effective Hamiltonian are often beset by the problem of intruder states, and are not suitable for studying potential energy surface (PES) involving real or avoided curve crossing. State-specific MR-based approaches obviate this limitation. The state-specific MRCC (SS-MRCC) method developed some years ago can handle quasidegeneracy of varying degrees over a wide range of PES, including regions of real or avoided curve-crossing. Motivated by its success, we have suggested and explored in this paper a suite of physically motivated coupled electron-pair approximations (SS-MRCEPA) like methods, which are designed to capture the essential strength of the parent SS-MRCC method without significant sacrificing its accuracy. These SS-MRCEPA theories, like their CC counterparts, are based on complete active space, treat all the reference functions on the same footing and provide a description of potentially uniform precision of PES of states with varying MR character. The combining coefficients of the reference functions are self-consistently determined along with the cluster amplitudes themselves. The newly developed SS-MRCEPA methods are size-extensive, and are also size-consistent with localized orbitals. Among the various versions, there are two which are invariant with respect to the restricted rotations among doubly occupied and active orbitals separately. Similarity of performance of this latter and the noninvariant versions at the crossing points of the degenerate orbitals imply that the all the methods presented are rather robust with respect to the rotations among degenerate orbitals. Illustrative numerical applications are presented for PES of the ground state of a number of difficult test cases such as the model H4, H(8) problems, the insertion of Be into H(2), and Li(2), where intruders exist and for a state of a molecule such as CH(2), with pronounced MR character. Results obtained with SS-MRCEPA methods are found to be comparable in accuracy to the parent SS-MRCC and FCI/large scale CI results throughout the PES, which indicates the efficacy of our SS-MRCEPA methods over a wide range of geometries, despite their neglect of a host of complicated nonlinear terms, even when the traditional MR-based methods based on effective Hamiltonians fail due to intruders. PMID- 15267479 TI - Phase dilemma in density matrix functional theory. AB - For closed-shell systems, a particular parametrization of coefficients in a configuration interaction (CI) expansion provides a convenient formulation for the search over electronic wave functions constrained by a set of natural orbitals (NOs) and the corresponding occupation numbers that are invoked in every variational construction of the density matrix functional (DMF) V(ee)(Gamma) for the electron--electron repulsion energy. It produces an explicit expression for V(ee) in terms of the Coulomb and exchange integrals over NOs, and an idempotent matrix omega, diagonal elements of which equal the occupation numbers. At the same time, it reveals a very serious bottleneck affecting any rigorous approach to the DMF theory, namely the phase dilemma that stems from the necessity to carry out minimization over a large number of possible combinations of CI coefficient signs. While underscoring its lack of variational nature, a simple approximation for the phase factor products provides a strict derivation for the recently proposed Kollmar-Hess functional. PMID- 15267480 TI - An ab initio study of tunneling splittings in the water dimer. AB - Tunneling splittings in the water dimer have been determined by the semiclassical WKB method, based on pathways characterized at the computational level of second order Moller-Plesset (MP2) theory with basis sets of aug-cc-pVTZ quality. This calculation takes into account all three acceptor tunneling, donor-acceptor interchange, and bifurcation tunneling rearrangements of the water dimer. The tunneling splittings were evaluated as 7.73 cm(-1) (large splitting) and 0.42 cm( 1) (small splitting), which are in good agreement with the corresponding experimental values of 11.18 cm(-1) and 0.70 cm(-1), respectively. PMID- 15267481 TI - The investigation of spin-orbit effect for the F((2)P)+HD reaction. AB - In this paper, we employ the time-dependent quantum wave packet method to study the reaction of F((2)P(3/2), (2)P(1/2)) with HD on the Alexander-Stark-Werner potential energy surface. The reaction probabilities and total integral cross sections of the spin-orbit ground and excited states for the two possible products of the system are calculated. Because the reaction channel of the excited spin-orbit state is closed at the resonance energy, the resonance feature does not appear in the reaction probabilities and cross section for the F((2)P(1/2))+HD(v=j=0)-->HF+D reaction, in contrast with that found for the ground spin--orbit state. We also compare the average cross sections of the two possible products with the experimental measurement. The resonance peak in the present average cross section for the HF+D product is slightly larger than the experimental result, but much smaller than that of the single-state calculations on the potential energy surface of Stark and Werner. It seems that the spin- orbit coupling would play a relatively important role in this reaction. Moreover, the isotope effects of the ground and excited spin--orbit states and the reactivity of the two product channels from the excited spin--orbit state are presented. PMID- 15267482 TI - Double photoionization of C(60) and C(70) in the valence region. AB - Photoion yields from gaseous fullerenes, C(60) and C(70), for production of singly and doubly charged ions are measured by mass spectrometry combined with tunable synchrotron radiation at hnu=25-150 eV. Since the signal of triply or highly charged ions is very weak, the total photoionization yield curve can be estimated from the sum of the yields of the singly and doubly charged ions. A distinct feature appears in the resultant curve of C(60) which is absent in the calculated total photoabsorption cross section previously reported. This difference is attributed to C(60) (2+) ions chiefly produced by spectator Auger ionization of the shape resonance states followed by tunneling of the trapped electron or by cascade Auger ionization. Ratios between the yields of doubly and singly charged ions for C(60) and C(70) are larger than unity at hnu>50 eV. These ratios are quite different from those reported in the experiments using electron impact ionization. PMID- 15267483 TI - Near threshold photoionization of the ground and first excited states of C(2). AB - Calculations using the multichannel Schwinger configuration-interaction method are presented for the photoionization from the ground and the first excited states of the C(2) molecule. Both single channel and multichannel calculations are presented in a photon energy range from the threshold to about 50 eV of photon energy. For the ground state, inclusion of both intrinsic and dynamical correlation effects is seen to strongly alter the picture of the photoionization process inferred from single-channel frozen-core Hartree-Fock calculations. Furthermore, the photoionization study of the first excited state of molecular carbon has revealed the presence of strong interchannel coupling between the 3sigma(g)-->ksigma(u) channel and the photoionization channels leading to the A (4)Pi(g) and f (2)Pi(g) ionic states in the near threshold region. PMID- 15267484 TI - Theoretical studies on dynamics and thermochemistry of the reactions CF(3)CHCl(2)+Cl-->CF(3)CCl(2)+HCl and CF(3)CHFCl+Cl-->CF(3)CFCl+HCl. AB - A dual-level direct dynamics study has been carried out for the two hydrogen abstraction reactions CF(3)CHCl(2)+Cl and CF(3)CHFCl+Cl. The geometries and frequencies of the stationary points are optimized at the BHLYP/6-311G(d,p), B3LYP/6-311G(d,p), and MP2/6-31G(d) levels, respectively, with single-point calculations for energy at the BHLYP/6-311++G(3df,2p), G3(MP2), and QCISD(T)/6 311G(d,p) levels. The enthalpies of formation for the species CF(3)CHCl(2), CF(3)CHFCl, CF(3)CCl(2), and CF(3)CFCl are evaluated at higher levels. With the information of the potential energy surface at BHLYP/6-311++G(3df,2p)//6 311G(d,p) level, we employ canonical variational transition-state theory with small-curvature tunneling correction to calculate the rate constants. The agreement between theoretical and experimental rate constants is good in the measured temperature range 276-382 K. The effect of fluorine substitution on reactivity of the C-H bond is discussed. PMID- 15267485 TI - Ultraviolet spectroscopy of pyrene in a supersonic jet and in liquid helium droplets. AB - In a series of experiments devoted to the study of polycyclic aromatic hydrocarbons for astrophysical applications, the S(2)<--S(0) transition of jet cooled pyrene (C(16)H(10)) at 321 nm has been studied by an absorption technique for the first time. The spectra observed by cavity ring-down spectroscopy closely resemble the excitation spectra obtained earlier by laser-induced fluorescence (LIF) and show the same band clusters arising from the vibronic interaction of S(2) with S(1). We have also investigated pyrene when it was incorporated into 380 mK cold helium droplets. These spectra which were recorded employing LIF and molecular beam depletion spectroscopy are broadened and redshifted by 0.94 nm but retain the essential features of the gas phase spectra. PMID- 15267486 TI - Theoretical study of Ng--NiN(2) (Ng=Ar,Ne,He). AB - It is shown that NiN(2) and noble gas atoms, Ar, Ne, and He, combine with the binding energy of 11.52, 4.06, and 7.37 kcal/mol, respectively, by the multireference perturbational (CASPT2) method. By the density functional theory calculations using MPWPW91 functionals, the Ni-N-N bending frequency in NiN(2) and Ar-NiN(2) is estimated as 310.7 and 358.7 cm(-1), respectively, the latter of which is in good agreement with the corresponding experimental frequency, 357.0 cm(-1), determined for NiN(2) isolated in solid argon. PMID- 15267487 TI - Quantum study of the N+N(2) exchange reaction: state-to-state reaction probabilities, initial state selected probabilities, Feshbach resonances, and product distributions. AB - We report a detailed three-dimensional time-dependent quantum dynamics study of the state-to-state N+N(2) exchange scattering in the 2.1-3.2 eV range using a recently developed ab initio potential energy surface (PES). The reactive flux arrives at the dividing surface in the asymptotic product region in a series of six packets, instead of a single packet. Further study shows that these features arise from the "Lake Eyring" region of the PES, a region with a shallow well between two transition states. Trappings due to Feshbach resonances are found to be the major cause of the time delay. A detailed analysis of the Feshbach resonance features is carried out using an L(2) calculation of the metastable states in the "Lake Eyring" region. Strong resonance features are found in the state-to-state and initial state selected reaction probabilities. The metastable states with bending motions and/or bending coupled with stretching motions are found to be the predominant source of the resonance structure. Initial state selected reaction probabilities further indicate that the lifetimes of the metastable states with bending motions in the "Lake Eyring" region are longer than those of states with stretching motions and thus dominate the reactive resonances. Resonance structures are also visible in some of the integral cross sections and should provide a means for future experimental observation of the resonance behavior. A study of the final rotational distributions shows that, for the energy range studied here, the final products are distributed toward high rotational states. Final vibrational distributions at the temperatures 2000 and 10,000 K are also reported. PMID- 15267488 TI - Non-Born-Oppenheimer study of positronic molecular systems: e(+)LiH. AB - Very accurate non-Born-Oppenheimer variational calculations of the ground state of e(+)LiH have been performed using explicitly correlated Gaussian functions with preexponential factors dependent on powers of the internuclear distance. In order to determine the positron detachment energy of e(+)LiH and the dissociation energy corresponding to the e(+)LiH fragmentation into HPs and Li(+) we also calculated non-BO energies of HPs, LiH, and Li(+). For all the systems the calculations provided the lowest ever-reported variational upper-bounds to the ground state energies. Annihilation rates of HPs and e(+)LiH were also computed. The dissociation energy of e(+)LiH into HPs and Li(+) was determined to be 0.036 548 hartree. PMID- 15267489 TI - Theoretical investigation of ground and excited states of the methylene amidogene radical (H(2)CN). AB - The excited states and the absorption spectrum of the methylene amidogene radical are studied by high-level ab initio calculations. The multireference configuration interaction method was used in combination with different basis sets and basis set extrapolation to compute equilibrium geometries, harmonic frequencies, and excitation energies of the four lowest doublet electronic states of the title species. Potential curves and transition dipole moment functions were determined along the normal mode coordinates of the electronic ground state. These functions were employed to determine vibronic absorption spectra. The intensities of dipole forbidden but vibronically allowed transitions were calculated by explicitly evaluating integrals over the vibrational wave functions and the transition dipole functions of the involved electronic states. By this method the oscillator strengths of the dipole allowed (2)A(1)<--(2)B(2) and the dipole forbidden (2)B(1)<--(2)B(2) bands were computed. It turns out that the dipole forbidden transition is two orders of magnitude weaker than the dipole allowed one. The 0-0 excitation energies are found to be 30 256 cm(-1) for the (2)B(1) state and 34,646 cm(-1) for the (2)A(1) state. From the combined results of the excitation energies and oscillator strengths it is concluded that the experimentally observed peaks must be due to the (2)A(1) state, in contradiction to earlier assignments. PMID- 15267490 TI - Barrier-free intermolecular proton transfer induced by excess electron attachment to the complex of alanine with uracil. AB - The photoelectron spectrum of the uracil-alanine anionic complex (UA)(-) has been recorded with 2.540 eV photons. This spectrum reveals a broad feature with a maximum between 1.6 and 2.1 eV. The vertical electron detachment energy is too large to be attributed to an (UA)(-) anionic complex in which an intact uracil anion is solvated by alanine, or vice versa. The neutral and anionic complexes of uracil and alanine were studied at the B3LYP and second-order Moller-Plesset level of theory with 6-31++G(*) (*) basis sets. The neutral complexes form cyclic hydrogen bonds and the three most stable neutral complexes are bound by 0.72, 0.61, and 0.57 eV. The electron hole in complexes of uracil with alanine is localized on uracil, but the formation of a complex with alanine strongly modulates the vertical ionization energy of uracil. The theoretical results indicate that the excess electron in (UA)(-) occupies a pi(*) orbital localized on uracil. The excess electron attachment to the complex can induce a barrier free proton transfer (BFPT) from the carboxylic group of alanine to the O8 atom of uracil. As a result, the four most stable structures of the uracil-alanine anionic complex can be characterized as a neutral radical of hydrogenated uracil solvated by a deprotonated alanine. Our current results for the anionic complex of uracil with alanine are similar to our previous results for the anion of uracil with glycine, and together they indicate that the BFPT process is not very sensitive to the nature of the amino acid's hydrophobic residual group. The BFPT to the O8 atom of uracil may be relevant to the damage suffered by nucleic acid bases due to exposure to low energy electrons. PMID- 15267491 TI - Time-dependent wave packet study on trans-cis isomerization of HONO. AB - Using a full six-dimensional ab initio potential energy surface and nuclear motion Hamiltonian, time-dependent computations were performed for the cis-trans isomerization of HONO. The multiconfiguration time-dependent Hartree method was used to propagate the six-dimensional wave packets. The initial excitations were chosen to be excitations of the local stretch modes and the HON local bend mode. The energy redistribution within 2 to 5 ps in the energy region of the OH stretching modes in both isomers was analyzed. The Fourier transformed frequency domain spectra were attributed to the eigenstates calculated previously by the time-independent variational approach. The results are also compared with classical trajectory computations of Thomson et al. on empirical surfaces. In agreement with matrix experiments, the cis-->trans isomerization was found to be much faster than the opposite interconversion. The intramolecular dynamics were found to be very complex involving numerous weakly excited delocalized eigenstates and anharmonic resonances. Particularly in the cis-isomer, the excitation of the HON bending local mode leads to fast energy redistribution in cis-trans delocalized modes. Neither the excitation of the OH stretching local mode in the cis nor in the trans form produces a fast isomerization, in agreement with the strongly localized characters of the corresponding eigenstates calculated variationally by Richter et al. and the gas phase spectra of HONO. PMID- 15267492 TI - A reoptimization of the five-site water potential (TIP5P) for use with Ewald sums. AB - The five-site transferable interaction potential (TIP5P) for water is most accurate at reproducing experimental data when used with a simple spherical cutoff for the long-ranged electrostatic interactions. When used with other methods for treating long-ranged interactions, the model is considerably less accurate. With small modifications, a new TIP5P-like potential can be made which is very accurate for liquid water when used with Ewald sums, a more physical and increasingly more commonly used method for treating long-ranged electrostatic interactions. The new model demonstrates a density maximum near 4 degrees C, like the TIP5P model, and otherwise is similar to the TIP5P model for thermodynamic, dielectric, and dynamical properties of liquid water over a range of temperatures and densities. An analysis of this and other commonly used water models reveals how the quadrupole moment of a model can influence the dielectric response of liquid water. PMID- 15267493 TI - Domain catalyzed chemical reactions: a molecular dynamics simulation of isomerization kinetics. AB - The model for domain catalyzed isomerization kinetics in condensed fluids is applied for a diluted mixture of a chiral solute with a consolute temperature. The solution is quench to phase separation at temperatures below the consolute temperature. The droplet coalescence enhances the isomerization kinetics due to the substantial excess pressure inside the small droplets given by the Laplace equation. The domain catalyzed isomerization kinetics breaks the symmetry, and the droplets end with only one dominating species. We argue that D-glyceraldehyde which is only moderately solvable in water and which has played a crucial role in the evolution is a candidate for the stereo specific ordering in bio-organic matter. PMID- 15267494 TI - Acetone hydration in supercritical water: (13)C-NMR spectroscopy and Monte Carlo simulation. AB - The (13)C-NMR chemical shift of acetone delta((13)C[Double Bond]O) was measured in aqueous solution at high temperatures up to 400 degrees C and water densities of 0.10-0.60 g/cm(3) for the study of hydration structure in the supercritical conditions. The average number N(HB) of hydrogen bonds (HBs) between an acetone and solvent waters and the energy change DeltaE upon the HB formation were evaluated from the delta and its temperature dependence, respectively. At 400 degrees C, N(HB) is an increasing function of the water density, the increase being slower at higher water densities. The acetone-water HB formation is exothermic in supercritical water with larger negative DeltaE at lower water densities (-3.3 kcal/mol at 0.10 g/cm(3) and -0.3 kcal/mol at 0.60 g/cm(3)), in contrast to the positive DeltaE in ambient water (+0.078 kcal/mol at 4 degrees C). The corresponding Monte Carlo simulations were performed to calculate the radial and orientational distribution functions of waters around the acetone molecule. The density dependence of N(HB) calculated at 400 degrees C is in a qualitative agreement with the experimental results. In the supercritical conditions, the HB angle in a neighboring acetone-water pair is weakly influenced by the water density, because of the absence of collective HB structure. This is in sharp contrast to the hydration structure in ambient water, where the acetone water HB formation is orientationally disturbed by the tetrahedral HB network formation among the surrounding waters. PMID- 15267495 TI - Influence of diffusion on the kinetics of excited-state association--dissociation reactions: comparison of theory and simulation. AB - Several recent theories of the kinetics of diffusion influenced excited-state association--dissociation reactions are tested against accurate Brownian dynamics simulation results for a wide range of parameters. The theories include the relaxation time approximation (RTA), multiparticle kernel decoupling approximations and the so-called kinetic theory. In the irreversible limit, none of these theories reduce to the Smoluchowski result. For the pseudo-first-order target problem, we show how the RTA can be modified so that the resulting formalism does reduce correctly in the irreversible limit. We call this the unified Smoluchowski approximation, because it unites modern theories of reversible reactions with Smoluchowski's theory of irreversible reactions. PMID- 15267496 TI - Non-Newtonian behavior in simple fluids. AB - Using nonequilibrium molecular dynamics simulations, we study the non-Newtonian rheology of a microscopic sample of simple fluid. The calculations were performed using a configurational thermostat which unlike previous nonequilibrium molecular dynamics or nonequilibrium Brownian dynamics methods does not exert any additional constraint on the flow profile. Our findings are in agreement with experimental results on concentrated "hard sphere"-like colloidal suspensions. We observe: (i) a shear thickening regime under steady shear; (ii) a strain thickening regime under oscillatory shear at low frequencies; and (iii) shear induced ordering under oscillatory shear at higher frequencies. These results significantly differ from previous simulation results which showed systematically a strong ordering for all frequencies. They also indicate that shear thickening can occur even in the absence of a solvent. PMID- 15267497 TI - Ab initio molecular dynamics simulations of local structure of supercooled Ni. AB - We report results of first-principles molecular dynamics simulations for stable and undercooled nickel liquids. The calculated structure factors as a function of temperature are discussed with respect to recent experimental measurements. In addition, structural analysis using bonding orientational order and three dimensional pair analysis techniques have been performed in detail and the effect of undercooling on the microstructure has been analyzed. More particularly, we show the importance of fivefold symmetry local structures. PMID- 15267498 TI - Liquid stability in a model for ortho-terphenyl. AB - We report an extensive study of the phase diagram of a simple model for ortho terphenyl, focusing on the limits of stability of the liquid state. Reported data extend previous studies of the same model to both lower and higher densities and to higher temperatures. We estimate the location of the homogeneous liquid-gas nucleation line and of the spinodal locus. Within the potential energy landscape formalism, we calculate the distributions of depth, number, and shape of the potential energy minima and show that the statistical properties of the landscape are consistent with a Gaussian distribution of minima over a wide range of volumes. We report the volume dependence of the parameters entering in the Gaussian distribution (amplitude, average energy, variance). We finally evaluate the locus where the configurational entropy vanishes, the so-called Kauzmann line, and discuss the relative location of the spinodal and Kauzmann loci. PMID- 15267499 TI - Disentangling density and temperature effects in the viscous slowing down of glassforming liquids. AB - We present a consistent picture of the respective role of density (rho) and temperature (T) in the viscous slowing down of glassforming liquids and polymers. Specifically, based in part upon a new analysis of simulation and experimental data on liquid ortho-terphenyl, we conclude that a zeroth-order description of the approach to the glass transition (in the range of experimentally accessible pressures) should be formulated in terms of a temperature-driven super-Arrhenius activated behavior rather than a density-driven congestion or jamming phenomenon. The density plays a role at a quantitative level, but its effect on the viscosity and the alpha-relaxation time can be simply described via a single parameter, an effective interaction energy that is characteristic of the high-T liquid regime; as a result, rho does not affect the "fragility" of the glassforming system. PMID- 15267500 TI - First principles calculations of electronic structures and metal mobility of Na(x)Si(46) and Na(x)Si(34) clathrates. AB - Energetics, geometry, electronic band structures, and charge transfer for Na(x)Si(46) and Na(x)Si(34) clathrates with different degrees of cavity filling by sodium, and the mobility of the Na atom inside the different cavities are studied using first principles density functional calculations within the generalized gradient approximation. The stabilization of the clathrate lattice and the cell volume variation upon the inclusion of Na (which appears to move easily in the larger cavities of Na(x)Si(34), thus justifying the experimental observations) are discussed in connection with the onset of the repulsion between Na and Si for distances shorter than approximately 3.4 A. For all degrees of filling of the different cavities examined we find that the electron population of the s orbitals in the partially ionized Na atoms increases with a decrease in the size of the cavity, and that the Na states contribute significantly to the density of states at the Fermi level and thus influence the properties of these compounds. PMID- 15267501 TI - Radiative decay of nonstationary system. AB - When a finite quantum system, say a fluorescent molecule is attached to a bulk surface and excited by a short laser pulse, the decay dynamics of the system is modulated by the surface and the signal is enhanced due to the bulk surface. We have considered the decay dynamics of a model of displaced distorted molecule whose excited potential surface is coupled to a continuum and then this first continuum is in turn coupled to a second continuum. In the short time scale there is a coherent exchange of energy between the system molecule and the first continuum states. In the long time scale the energy of the whole system plus first continuum drains out to the final continuum states. A dendrimer nanocomposite with the gold surface shows an enhanced light emission. This can be qualitatively understood from the model we proposed here. We have numerically studied the various potential parameters of the molecule which can affect the signal. When the potential surfaces are flat, the band structure of the first continuum states along with its initial excitation has some nontrivial effect on the profile of the radiative decay. PMID- 15267502 TI - A theoretical model of the photoabsorption spectra of carbon buckyonions. AB - A theoretical model has been developed to predict the photoabsorption spectra of spherical carbon buckyonions in the region dominated by the pi-plasmon feature. This model makes use of the microscopic electronic structure of the system, which is provided by an effective Huckel one-electron model. The important screening effects are treated within the random phase approximation, whose form is an extension to the dynamic case of the one derived in a previous work for the static polarizabilities of these species. A systematic analysis as a function of the buckyonion size is performed. We compare the spectra obtained in this way with those derived from a different representation of the electron motion, namely a two-dimensional spherical electron gas, and from a classical dielectric model. PMID- 15267503 TI - Triple point of Lennard-Jones fluid in slit nanopore: solidification of critical condensate. AB - We report the results of a molecular dynamics simulation that looked for the triple point of Lennard-Jones fluid in slit-shaped nanopores. The simulation method employed for this purpose is able to maintain vapor-liquid coexistence in a nanopore at a specific equilibrium bulk-phase pressure. The triple point is the freezing point of the critical condensate. The triple-point temperature could be higher or lower than the bulk triple point, depending on the pore size. This is thought to be due to two opposing factors: the elevating effect of the pore-wall potential energy, and the depressing effect of the capillary condensate's tensile condition. Because of the cancellation, the deviation of the triple-point temperature from the bulk triple-point temperature was not considered significant. The pressure of the triple point, however, was significantly different from that of the bulk triple point. A simple model to describe the triple point is developed and shown to agree well with the results of the simulation. The importance of the two factors in nanoscale pores, which cannot be described by the classic Gibbs-Thomson equation, is emphasized. PMID- 15267504 TI - Dopant distribution in a Tm(3+)-Yb(3+) codoped silica based glass ceramic: an infrared-laser induced upconversion study. AB - The optically active dopant distribution in a Tm(3+)-Yb(3+) doped silica based glass ceramic sample has been investigated. A systematic analysis of the upconversion fluorescence of the Tm(3+)-Yb(3+) codoped glass and glass ceramic has been performed at room temperature. Tm(3+) and Yb(3+) single doped glass and glass ceramics have also been included in the study. Upon infrared excitation at 790 nm into the (3)H(4) level of the Tm(3+) ions a blue upconversion emission is observed, which is drastically increased in the Yb(3+) codoped samples. A rate equation model confirmed the energy transfer upconversion mechanism. Based on these results, the temporal dynamic curves of the levels involved in the upconversion process, (3)H(4), (2)F(5/2), and (1)G(4) were interpreted in the glass ceramic samples. The contribution of the optically active Tm(3+) and Yb(3+) ions in the crystalline and in the vitreous phase of the glass ceramic was distinguished and the ratio of Tm(3+) ions in the crystalline phase could be quantified for the 1 mol % Tm(3+)-2.5 mol % Yb(3+) glass ceramic. A surprising result was obtained for that concentration: the main contribution to the upconversion emission of the glass ceramic is due to Tm(3+)-Yb(3+) ions in the vitreous phase. PMID- 15267505 TI - A rotational study of laser ablated thiourea. AB - A laser ablation device in combination with a molecular beam Fourier-transform microwave spectrometer has allowed the observation of the rotational spectrum of solid thiourea for the first time. The sensitivity reached in the experiment allowed the observation of the isotopomers (34)S, (13)C, and (15)N in their natural abundance. The spectrum of D(4)-thiourea was also analyzed from an enriched sample. The complicated hyperfine structure arising from the presence of two (14)N quadrupolar nuclei has been fully resolved and analyzed. The substitution r(s) structure has been derived from the experimental moments of inertia. Thiourea in gas phase presents a planar heavy atom skeleton. Experimental inertial defect values and high-level ab initio calculations reveal that the amino groups hydrogen atoms lie out-of-plane with a C(2) symmetry configuration and are involved in large amplitude inversion motions. PMID- 15267506 TI - Small-angle neutron scattering study of structural changes in temperature sensitive microgel colloids. AB - The structure of temperature-sensitive poly(N-isopropylacrylamide) microgels in dilute suspension was investigated by means of small-angle neutron scattering. A direct modeling expression for the scattering intensity distribution was derived which describes very well the experimental data at all temperatures over an extensive q range. The overall particle form as well as the internal structure of the microgel network is described by the model. The influence of temperature, cross-linking density, and particle size on the structure was revealed by radial density profiles and clearly showed that the segment density in the swollen state is not homogeneous, but gradually decays at the surface. The density profile reveals a box profile only when the particles are collapsed at elevated temperatures. An increase of the cross-linking density resulted in both an increase of the polymer volume fraction in the inner region of the particle and a reduction of the smearing of the surface. The polymer volume fraction inside the colloid decreased with increasing particle size. The structural changes are in good agreement with the kinetics of the emulsion copolymerization used to prepare the microgel colloids. PMID- 15267507 TI - Water's polyamorphic transitions and amorphization of ice under pressure. AB - Transformations of water's high density amorph (HDA) to low density amorph (LDA) and of LDA's to cubic ice (Ic) have been studied by in situ thermal conductivity kappa measurements at high pressures. The HDA to LDA transformation is unobservable at p of 0.07 GPa, indicating that, for a fixed heating rate, an increase in pressure increases the temperature of HDA to LDA transformation and decreases that of LDA to ice Ic, causing thereby the two transformations to merge, and HDA appears to convert directly to ice Ic. Thus either LDA forms but converts extremely rapidly to ice Ic, or LDA does not form. At a fixed p and T, in the range of pressure amorphization of hexagonal ice, kappa continues to decrease with time. Therefore, the amorphization of ice Ih is kinetically controlled. When HDA at 1 GPa was heated from 130 to 157 K and densified to very HDA, its kappa increased by 3%. Our findings and a scrutiny of earlier reports show that a reversible transition between HDA and LDA does not occur at approximately 135 K and approximately 0.2 GPa. Since there is no unique HDA, it is difficult to justify the conjecture for a second critical point for water. PMID- 15267508 TI - Surface/interface electronic structure in C(60) anchored aminothiolate self assembled monolayer: an approach to molecular electronics. AB - Electronic structure in self-assembled monolayers (SAMs) of C(60) anchored 11 amino-1-undecane thiol (C(60)-11-AUT) on Au(111) was studied by means of ultraviolet photoelectron spectroscopy and hybrid density functional theory calculations. Valence band features of the molecular conformation revealed the interface electronic structure to be dominated by sigma(S-Au), localized at the thiolate anchor to Au. Formation of a localized covalent bond as a result of hybridization between N P(z) orbital of -NH(2) group of the thiolate SAM and the pi level of C(60) resulted in a symmetry change from I(h) in C(60) to C1 in C(60) 11-AUT SAM. Appearance of low, but finite amplitude surface electronic states of bonded C(60), much beyond the Fermi level, ruled out Au-C(60) end group contact. The band gap E(g) of the SAM, determined to be 2.7 eV, was drastically reduced from the insulating alkanethiol SAMs ( approximately 8.0 eV) and fell intermediate between the C(60) ground state (N electrons, 1.6 eV) and C(60) solid (N+/-1 electrons, 3.7 eV). PMID- 15267509 TI - Theoretical study of the H(2) reaction with a Pt(4) (111) cluster. AB - The C(s) symmetry reaction of the H(2) molecule on a Pt(4) (111) clusters, has been studied using ab initio multiconfiguration self-consistent field plus extensive multireference configuration interaction variational and perturbative calculations. The H(2) interaction by the vertex and by the base of a tetrahedral Pt(4) cluster were studied in ground and excited triplet and singlet states (closed and open shells), where the reaction curves are obtained through many avoided crossings. The Pt(4) cluster captures and activates the hydrogen molecule; it shows a similar behavior compared with other Pt(n) (n=1,2,3) systems. The Pt(4) cluster in their lowest five open and closed shell electronic states: (3)B(2), (1)B(2), (1)A(1) (3)A(1), (1)A(1), respectively, may capture and dissociate the H(2) molecule without activation barriers for the hydrogen molecule vertex approach. For the threefolded site reaction, i.e., by the base, the situation is different, the hydrogen adsorption presents some barriers. The potential energy minima occur outside and inside the cluster, with strong activation of the H-H bond. In all cases studied, the Pt(4) cluster does not absorb the hydrogen molecule. PMID- 15267510 TI - The role of polymer spacers in specific adhesion. AB - We study the role of flexible spacers in specific adhesion from the point of view of polymer reaction--diffusion theory. By assuming that the interactions between complementary adhesion moieties occur on a length scale much smaller than the size of the polymer spacer, we describe in detail binding and rupture between two opposing surfaces. Predictions are given for the physical properties of interest such as the time evolution of bond density and the ranges of attraction and unbinding. We also discuss the dynamic crossover between reversible and irreversible bridging. PMID- 15267511 TI - Electric field effects on photoinduced electron transfer processes of methylene linked compounds of pyrene and N,N-dimethylaniline in a polymer film. AB - Time-resolved measurements of the electric-field-induced change in fluorescence intensity have been made for methylene-linked compounds of pyrene and N,N dimethylaniline (DMA) doped in a polymer film. The lifetime of the fluorescence emitted from the locally excited state of pyrene chromophore becomes shorter in the presence of electric field (F), when the dopant concentration is high. The lifetime of the excipelx fluorescence resulting from the photoinduced electron transfer (PIET) from DMA to the excited state of pyrene chromophore between different molecules also becomes shorter in the presence of F. Based on the simulation of the electric field effect on fluorescence decay, the mechanism of intermolecular PIET between DMA and pyrene chromophore in a polymer film is discussed. PMID- 15267512 TI - Influence of sequence correlations on the adsorption of random heteropolymers onto homogeneous planar surfaces. AB - Using a reference system approach, we develop an analytical theory for the adsorption of random heteropolymers with exponentially decaying and/or oscillating sequence correlations on planar homogeneous surfaces. We obtain a simple equation for the adsorption--desorption transition line. This result as well as the validity of the reference system approach is tested by a comparison with numerical lattice calculations. PMID- 15267513 TI - Size and persistence length of molecular bottle-brushes by Monte Carlo simulations. AB - Single-chain simulations of densely branched comb polymers, or "molecular bottle brushes" with side-chains attached to every (or every second) backbone monomer, were carried out by off-lattice Monte Carlo technique. A coarse-grained model, described by hard spheres connected by harmonic springs, was employed. Backbone lengths of up to 100 units were considered, and compared with the corresponding linear chains. The backbone molecular size was investigated as a function of its length at fixed arm size, and as a function of the arm size at fixed backbone length. The apparent swelling exponents obtained by a power-law fit were found to be larger than those for the corresponding linear polymers, indicative of stiffening of the comb backbone. The probability distribution function for the backbone end-to-end distance was also investigated for different backbone lengths and arm sizes. Analysis of this function yielded the critical exponents, which revealed an increase in the swelling exponent consistent with values found from the molecular size. The apparent persistence length of the backbone was also determined, and was found to increase with increasing branching density. Finally, the static structure factors of the whole bottle-brushes and of their backbones are discussed, which provides another consistent estimate of the swelling exponents. PMID- 15267514 TI - Transport coefficients of aqueous dodecyltrimethylammonium bromide solutions: comparison between experiments, analytical calculations and numerical simulations. AB - We study dynamical properties of ionic species in aqueous solutions of dodecyltrimethylammonium bromide, for several concentrations below and above the critical micellar concentration (cmc). New experimental determinations of the electrical conductivity are given which are compared to results obtained from an analytical transport theory; transport coefficients of ions in these solutions above the cmc are also computed from Brownian dynamics simulations. Analytical calculations as well as the simulation treat the solution within the framework of the continuous solvent model. Above the cmc, three ionic species are considered: the monomer surfactant, the micelle and the counterion. The analytical transport theory describes the structural properties of the electrolyte solution within the mean spherical approximation and assumes that the dominant forces which determine the deviations of transport processes from the ideal behavior (i.e., without any interactions between ions) are hydrodynamic interactions and electrostatic relaxation forces. In the simulations, both direct interactions and hydrodynamic interactions between solutes are taken into account. The interaction potential is modeled by pairwise repulsive 1/r(12) interactions and Coulomb interactions. The input parameters of the simulation (radii and self-diffusion coefficients of ions at infinite dilution) are partially obtained from the analytical transport theory which fits the experimental determinations of the electrical conductivity. Both the electrical conductivity of the solution and the self-diffusion coefficients of each species computed from Brownian dynamics are compared to available experimental data. In every case, the influence of hydrodynamic interactions (HIs) on the transport coefficients is investigated. It is shown that HIs are crucial to obtain agreement with experiments. In particular, the self-diffusion coefficient of the micelle, which is the largest and most charged species in the present system, is enhanced when HIs are included whereas the diffusion coefficients of the monomer and the counterion are roughly not influenced by HIs. PMID- 15267515 TI - Thermodynamic stability and kinetic foldability of a lattice protein model. AB - By using serial mutations, i.e., a residue replaced by 19 kinds of naturally occurring residues, the stability of native conformation and folding behavior of mutated sequences are studied. The 3 x 3 x 3 lattice protein model with two kinds of interaction potentials between the residues, namely the original Miyazawa and Jernigan (MJ) potentials and the modified MJ potentials (MMJ), is used. Effects of various sites in the mutated sequences on the stability and foldability are characterized through the Z-score and the folding time. It is found that the sites can be divided into three types, namely the hydrophobic-type (H-type), the hydrophilic-type (P-type) and the neutral-type (N-type). These three types of sites relate to the hydrophobic core, the hydrophilic surface and the parts between them. The stability of the native conformation for the serial mutated sequences increases (or decreases) as the increasing in the hydrophobicity of the mutated residues for the H-type sites (or the P-type sites), while varies randomly for the N-type sites. However, the foldability of the mutated sequences is not always consistent with the thermodynamic stability, and their relationship depends on the site types. Since the hydrophobic tendency of the MJ potentials is strong, the ratio between the number of the H-type sites and the number of the P type sites is found to be 1:2. Differently, for the MJJ potentials it is found that such a ratio is about 1:1 which is relevant to that of real proteins. This suggests that the modification of the MJ potentials is rational in the aspect of thermodynamic stability. The folding of model proteins with the MMJ potentials is fast. However, the relationship between the foldability and the thermodynamic stability of the mutated sequences is complex. PMID- 15267516 TI - Mixtures of lattice polymers with structured monomers. AB - The influence of monomer structure on the thermodynamic properties of lattice model polymer blends is investigated through Monte Carlo computations. The model of lattice polymers with monomer structure has been used extensively in the context of the lattice cluster theory (LCT), a thermodynamic theory for polymer mixtures in the liquid state. The Monte Carlo computations provide the first unequivocal test of the accuracy of the LCT predictions for binary mixtures of polymers with structured monomers. Four types of monomer structures are analyzed, corresponding to to the monomers of polyethylene, polypropylene, polyethylethylene, and polyisobutylene (PIB). Most computations use chains with M=12 and 24 beads and the total volume fraction of the beads is phi=0.6. Both structurally symmetric and asymmetric blends are investigated. For the symmetric case, the predictions of the LCT for the energies of mixing and the liquid-liquid coexistence curves are in qualitative agreement with the Monte Carlo computations, except for the PIB/PIB symmetric blend. For structurally asymmetric blends, the LCT does not capture contributions to the energy of mixing arising solely from structural differences between the components. Computational estimates of the nonideal entropy of mixing indicate that the LCT also underestimates the entropic cost of mixing chains with different structures, thus explaining some discrepancies between the theoretical and the Monte Carlo liquid- liquid coexistence curves. PMID- 15267517 TI - Emergence of multiple tori structures in a single polyelectrolyte chain. AB - We investigated the collapsed structure of a weakly charged wormlike chain under a moderate concentration of 1:1 electrolyte solution. By assuming a torus as a grand state, we found that the size of a torus is determined by the balance between surface energy and electrostatic energy, which leads to a finite torus thickness almost independent of the chain contour length. Owing to this unique characteristic, a long charged wormlike chain forms multiple tori structure as a collapsed product, which is never seen with a neutral wormlike chain. These features were confirmed by a Monte Carlo simulation. PMID- 15267518 TI - Simulation of polymer--polymer interdiffusion using the dynamic lattice liquid model. AB - In this paper, we present computer simulation results concerning interdiffusion of fully compatible components in symmetric binary (AB) polymer mixtures in solutions. The simulation is performed in two dimensions using the algorithm based on the dynamic lattice liquid model. The solvent molecules are taken into account explicitly. The evolution of the concentration profiles in time at an interface is studied for chain lengths N=2,4,8,16 for three polymer concentrations phi=0.1,0.5,0.9. The tracer diffusion coefficients for polymer chains and for the solvent are obtained by monitoring the mean square displacements of their center of mass. The relationships between coefficients of interdiffusion and self-diffusion are tested. PMID- 15267519 TI - Electron attachment to C(2)Cl(4) and Trojan horse ionization. AB - In a recent study of tetrachloroethylene, the anion yield curves were analyzed using three published negative-ion Morse potentials. Unexpected ions at zero electron energy were explained by the "Trojan horse" mechanism. This communication also attributes formation of Cl(2)(-) at higher energies to a Trojan horse mechanism. Six new Morse potentials are calculated to account for the observed anion states. These combine all extant electron impact and attachment data. The electron affinity of the C(2)Cl(3) radical, 3.1(1) eV, and the C-Cl bond dissociation energy 4.0(1) eV are reported. PMID- 15267520 TI - Comment on "The nucleation behavior of supercooled water vapor in helium" [J. Chem. Phys. 117, 5647 (2002)]. AB - In a recent paper Peeters et al. published new experimental data on nucleation rates of water in the temperature range of 200-235 K. They reported about a drastic change in the nucleation rate at 207 K. An error in their experimental procedure has been found. The data of Peeters et al. have been reinterpreted. The jump in nucleation rate disappears and the corrected nucleation rate data are in good agreement with data found by Wolk and Strey with a different experimental facility. PMID- 15267522 TI - Kinetic theory and hydrodynamics of dense, reacting fluids far from equilibrium. AB - The kinetic theory for a fluid of hard spheres which undergo endothermic and/or exothermic reactions with mass transfer is developed. The exact balance equations for concentration, density, velocity, and temperature are derived. The Enskog approximation is discussed and used as the basis for the derivation, via the Chapman-Enskog procedure, of the Navier-Stokes reaction equations under various assumptions about the speed of the chemical reactions. It is shown that the phenomenological description consisting of a reaction-diffusion equation with a convective coupling to the Navier-Stokes equations is of limited applicability. PMID- 15267523 TI - Basis set and electron correlation effects on initial convergence for vibrational nonlinear optical properties of conjugated organic molecules. AB - Using three typical pi-conjugated molecules (1,3,5-hexatriene, 1-formyl-6 hydroxyhexa-1,3,5- triene, and 1,1-diamino-6,6-dinitrohexa-1,3,5-triene) we investigate the level of ab initio theory necessary to produce reliable values for linear and nonlinear optical properties, with emphasis on the vibrational contributions that are known to be important or potentially important. These calculations are made feasible by employing field-induced coordinates in combination with a finite field procedure. For many, but not all, purposes the MP2/6-31+G(d) level is adequate. Based on our results the convergence of the usual perturbation treatment for vibrational anharmonicity was examined. Although this treatment is initially convergent in most circumstances, a problematic situation has been identified. PMID- 15267524 TI - Test of the quantum instanton approximation for thermal rate constants for some collinear reactions. AB - Two variants of the recently developed quantum instanton (QI) model for calculating thermal rate constants of chemical reactions are applied to several collinear atom-diatom reactions with various skew angles. The results show that the original QI version of the model is consistently more accurate than the "simplest" quantum instanton version (both being applied here with one "dividing surface") and thus to be preferred. Also, for these examples (as with other earlier applications) the QI results agree well with the correct quantum rates (to within approximately 20% or better) for all temperatures >200 K, except for situations where dynamical corrections to transition state theory (i.e., "re crossing" dynamics) are evident. (Since re-crossing effects are substantially reduced in higher dimensionality, this is not a cause for serious concern.) A procedure is also described which facilitates use of the METROPOLIS algorithm for evaluating all quantities that appear in the QI rate expression by Monte Carlo path integral methods. PMID- 15267525 TI - Synchronization of trajectories in canonical molecular-dynamics simulations: observation, explanation, and exploitation. AB - For two methods commonly used to achieve canonical-ensemble sampling in a molecular-dynamics simulation, the Langevin thermostat and the Andersen [H. C. Andersen, J. Chem. Phys. 72, 2384 (1980)] thermostat, we observe, as have others, synchronization of initially independent trajectories in the same potential basin when the same random number sequence is employed. For the first time, we derive the time dependence of this synchronization for a harmonic well and show that the rate of synchronization is proportional to the thermostat coupling strength at weak coupling and inversely proportional at strong coupling with a peak in between. Explanations for the synchronization and the coupling dependence are given for both thermostats. Observation of the effect for a realistic 97-atom system indicates that this phenomenon is quite general. We discuss some of the implications of this effect and propose that it can be exploited to develop new simulation techniques. We give three examples: efficient thermalization (a concept which was also noted by Fahy and Hamann [S. Fahy and D. R. Hamann, Phys. Rev. Lett. 69, 761 (1992)]), time-parallelization of a trajectory in an infrequent-event system, and detecting transitions in an infrequent-event system. PMID- 15267526 TI - A local second-order Moller-Plesset method with localized orbitals: a parallelized efficient electron correlation method. AB - Using orthogonal localized occupied orbitals we have developed and implemented a parallelized local second-order Moller-Plesset (MP2) method based on the idea developed by Head-Gordon and co-workers. A subset of nonorthogonal correlation functions (the orbital domain) was assigned to each of the localized occupied orbitals using a distance criterion and excitations from localized occupied orbitals that were arranged into subsets. The correlation energy was estimated using a partial diagonalization and an iterative efficient method for solving large-scale linear equations. Some illustrative calculations are provided for molecules with up to 1484 Cartesian basis sets. The orbital domain sizes were found to be independent of the molecular size, and the present local MP2 method covered about 98%-99% of the correlation energy of the conventional canonical MP2 method. PMID- 15267527 TI - A constrained variational approach for energy derivatives in Fock-space multireference coupled-cluster theory. AB - In this paper, we present a formulation based on constrained variational approach to enable efficient computation of energy derivatives using Fock-space multireference coupled-cluster theory. Adopting conventional normal ordered exponential with Bloch projection approach, we present a method of deriving equations when general incomplete model spaces are used. Essential simplifications arise when effective Hamiltonian definition becomes explicit as in the case of complete model spaces or some special quasicomplete model spaces. We apply the method to derive explicit generic expressions upto third-order energy derivatives for [0,1], [1,0], and [1,1] Fock-space sectors. Specific diagrammatic expressions for zeroth-order Lagrange multiplier equations for [0,1], [1,0], and [1,1] sectors are presented. PMID- 15267528 TI - An electron in a finite-dipole potential. AB - Some structural properties of the energy eigenfunctions of an electron in a finite-dipole field are analyzed, in particular the asymptotic behavior when the electron is far away, and the coalescence and cusp properties when it is close to the dipole charges. Model wave functions incorporating these properties are developed, which give accurate values for the energies and some other quantities, and a useful insight into the physical structure of the system. Critical radius for the existence of the bound states is obtained from the Wentzel-Kramers Brillouin approach. These considerations are extended to the description of the system in two dimensions. PMID- 15267529 TI - Relativistic correlating basis sets for the sixth-period d-block atoms from Lu to Hg. AB - Contracted Gaussian-type function sets to describe valence correlation are developed for the sixth-period d-block atoms Lu through Hg. A segmented contraction scheme is employed for their compactness and efficiency. Contraction coefficients and exponents are determined by minimizing the deviation from accurate natural orbitals generated from configuration interaction calculations, in which relativistic effects are incorporated through the third-order Douglas Kroll approximation. The present basis sets yield more than 99% of atomic correlation energies predicted by accurate natural orbital sets of the same size. Relativistic model core potential calculations with the present correlating sets give the spectroscopic constants of the AuH molecule in excellent agreement with experimental results. PMID- 15267530 TI - Improving the accuracy of interpolated potential energy surfaces by using an analytical zeroth-order potential function. AB - We present a method for improving the accuracy and efficiency of interpolation methods, in which an analytical zeroth-order potential-energy surface is employed as a reference surface. To investigate and test the method, we apply it to hydrogen peroxide where there exists an accurate analytical surface which we take as the "exact" surface for obtaining the energies and derivatives for fitting and assessing the accuracy. Examples are given for four-dimensional and six dimensional surfaces interpolated by using either the modified Shepard or second degree interpolating moving least-squares approach, with comparisons for cases with and without using the zeroth-order potential. PMID- 15267531 TI - Kolmogorov-Sinai entropy for the A+B-->P reaction in transitional flows. AB - We study global decay of the bimolecular reaction A+B-->P as c(t) approximately t(-alpha) in a nonlinear transitional flow. A relationship is established between the decay exponent alpha, and a modified Kolmogorov-Sinai entropy, hr. We find that for dynamic conditions which induce relatively strong mixing, the decay exponent is alpha is proportional to ln psi(-Bhr) with B being a characteristic reactive mix-down time for the system, and psi is a space-time scaling parameter. Dynamic conditions which imply weak mixing lead to a degenerate dependence of alpha on hr. The proposed relationship between alpha on hr should be a useful link between the dynamical evolution of the flow field and reaction kinetics in vortex dominated flows. PMID- 15267532 TI - Exit interaction effect on nascent product state distribution of O(1D)+N2O- >NO+NO. AB - We have determined the rotational state distributions of NO(v'=0,1,2) products produced from the reaction O(1D)+N2O. This is the first full characterization of the product rotational distribution of this reaction. The main part of each rotational distribution (up to j' approximately 80) has rotational temperature approximately 20,000 K and all these distributions are quite near to those predicted by the phase space theory (PST). This observation and previously reported vibrational distribution indicate that the most part of the energy partitioning of the reaction products is at least apparently statistical although the intermediate of this reaction is not so stable as to ensure the long lifetime. On the other hand, the distributions in the high rotational levels (j'=80-100) are found to decrease more sharply as j' increases than the PST predictions. The origin of the observed decrease of the distribution is discussed with quasiclassical trajectory (QCT) calculations on a five-dimensional ab initio potential energy surface (PES). The observed near-statistical distribution and the sharp decrease in the high-j' levels are well reproduced by a "half collision" QCT calculation, where statistical distribution at the reaction intermediate is assumed. This agreement shows the rotation-translation interaction in the exit region has an effect of yielding small high-j' populations. However, a little bias of the calculated distribution toward lower rotational excitation than the observed one indicates that the combination of the statistical intermediate and the exit interaction on the current PES does not completely describe the real system. It is suggested that the reaction intermediate is generated with the distribution which is close to statistical but a little biased toward yielding high-j' products, and that the interaction in the exit region of the PES results in the sharp decrease in the high-j' levels. PMID- 15267533 TI - Sub-Doppler rotationally resolved spectroscopy of lower vibronic bands of benzene with Zeeman effects. AB - Sub-Doppler high-resolution excitation spectra and the Zeeman effects of the 6(0)(1), 1(0)(1)6(0)(1), and 1(0)(2)6(0)(1) bands of the S1(1)B2u<--S(0)(1)A1g transition of benzene were measured by crossing laser beam perpendicular to a collimated molecular beam. 1593 rotational lines of the 1(0) (1)6(0) (1) band and 928 lines of the 1(0)(2)6(0)(1) band were assigned, and the molecular constants of the excited states were determined. Energy shifts were observed for the S1(1)B2u(v1=1,v6=1,J,Kl=-11) levels, and those were identified as originating from a perpendicular Coriolis interaction. Many energy shifts were observed for the S1(1)B2u(v1=2,v6=1,J,Kl) levels. The Zeeman splitting of a given J level was observed to increase with K and reach the maximum at K=J, which demonstrates that the magnetic moment lies perpendicular to the molecular plane. The Zeeman splittings of the K=J levels were observed to increase linearly with J. From the analysis, the magnetic moment is shown to be originating mostly from mixing of the S1(1)B2u and S2(1)B1u states by the J-L coupling (electronic Coriolis interaction). The number of perturbations was observed to increase as the excess energy increases, and all the perturbing levels were found to be a singlet state from the Zeeman spectra. PMID- 15267534 TI - A theoretical and experimental study of the SO2(2+) dication. AB - The double photoionization spectrum of SO2 has been measured using the TOF-PEPECO technique and contains one resolved band. Detailed electronic structure calculations and experimental comparisons allow the resolved band to be identified as the A 1A2 state of the SO2(2+) dication, with its adiabatic ionization energy at 35.284+/-0.02 eV. According to the most accurate calculations, the ground state level of SO2(2+) must be located near 33.48 eV, well below the range accessed by vertical transitions from neutral SO2. Transient SO2 (2+) molecules detected by mass spectrometry may be identified either as the sharp levels of the A 1A2 state or as ground state levels populated by nonvertical ionization pathways. PMID- 15267535 TI - Gas phase infrared spectroscopy of mono- and divanadium oxide cluster cations. AB - The vibrational spectroscopy of the mono- and divanadium oxide cluster cations VO(1-3)+ and V2O(2-6)+ is studied in the region from 600 to 1600 wave numbers by infrared photodissociation of the corresponding cluster cation-helium atom complexes. The comparison of the experimental depletion spectra with the results of density functional calculations on bare vanadium oxide cluster cations allows for an unambiguous identification of the cluster geometry in most cases and, for VO(1-3)+ and V2O(5,6)+, also of the electronic ground state. A common structural motif of all the studied divanadium cluster cations is a four-membered V-O-V-O ring, with three characteristic absorption bands in the 550-900 wave number region. For the V-O-V and V=O stretch modes the relationship between vibrational frequencies and V-O bond distances follows the Badger rule. PMID- 15267536 TI - Potential energy surface and rovibrational states of the ground Ar-HI complex. AB - A potential energy surface for the ground electronic state of the Ar-HI van der Waals complex is calculated at the coupled-cluster with single and double excitations and a noniterative perturbation treatment of triple excitations [CCSD(T)] level of theory. Calculations are performed using for the iodine atom a correlation consistent triple-zeta valence basis set in conjunction with large core Stuttgart-Dresden-Bonn relativistic pseudopotential, whereas specific augmented correlation consistent basis sets are employed for the H and Ar atoms supplemented with an additional set of bond functions. In agreement with previous studies, the equilibrium structure is found to be linear Ar-I-H, with a well depth of 205.38 cm(-1). Another two secondary minima are also predicted at a linear and bent Ar-H-I configurations with well depths of 153.57 and 151.57 cm( 1), respectively. The parametrized CCSD(T) potential is used to calculate rovibrational bound states of Ar-HI/Ar-DI complexes, and the vibrationally averaged structures of the different isomers are determined. Spectroscopic constants are also computed from the CCSD(T) surface and their comparison with available experimental data demonstrates the quality of the present surface in the corresponding configuration regions. PMID- 15267537 TI - An application of error reduction and harmonic inversion schemes to the semiclassical calculation of molecular vibrational energy levels. AB - A singular value decomposition based harmonic inversion signal processing scheme is applied to the semiclassical initial value representation (IVR) calculation of molecular vibrational states. Relative to usual IVR procedure of Fourier analysis of a signal made from the Monte Carlo evaluation of the phase space integral in which many trajectories are needed, the new procedure obtains acceptable results with many fewer trajectories. Calculations are carried out for vibrational energy levels of H2O to illustrate the overall procedure. PMID- 15267538 TI - Static dipole polarizability and binding energy of sodium clusters Nan (n=1-10): A critical assessment of all-electron based post Hartree-Fock and density functional methods. AB - A systematic all electron post Hartree-Fock as well as density functional theory (DFT) based calculations for the polarizability and binding energy of sodium metal clusters have been performed and an in-depth analysis of the discrepancy between the experimental and theoretical results is presented. A systematic investigation for the assessment of different DFT exchange-correlation functionals in predicting the polarizability values has also been reported. All the pure DFT functionals have been found to considerably underestimate the calculated polarizability values as compared to the MP2 results. DFT calculations using the full Hartree-Fock exchange along with one-parameter progressive correlation functional have, however, been shown to yield results in good agreement with the MP2 and experimental results. The possible sources of error present in the experimental measurements as well as in the different theoretical methods have also been analyzed. One of the most important conclusions of the present study is that the effect of electron correlation plays a significant role in determining the polarizability of the clusters and the MP2 method can be considered to be one of the most reliable methods for their prediction. It has also been noted that the polarizability value of the lower member clusters (Na2 and Na4) calculated by highly sophisticated methods such as, CCSD and CCSD(T) are found to be very close to the corresponding MP2 values. The polarizability and the binding energy of the clusters are found to be inversely related to each other and their correlation is rationalized by invoking the minimum polarizability principle. A good linear correlation between the polarizability and volume of the cluster has also been found to exist. PMID- 15267539 TI - Trans-1-chloro-2-fluoroethylene: microwave spectra and anharmonic force field. AB - For the first time the millimeter-wave spectra of the trans-35ClHC=CHF and trans 37ClHC=CHF isotopomers have been observed in natural abundance. Many DeltaJ=0, +/ 1 DeltaK(-1)=+1 transitions for 35ClHC=CHF and DeltaJ=0 DeltaK(-1)=+1 transitions for 37ClHC=CHF have been detected and assigned. This allowed us to accurately determine the vibrational ground-state rotational constants, quartic and some sextic centrifugal distortion constants, and nuclear quadrupole coupling constants for both 35Cl and 37Cl. The experimental investigation has been supported by highly accurate theoretical predictions. As far as ab initio computations are concerned, the complete set of cubic and quartic force constants have been evaluated by numerical differentiation of the analytic second-order Moller-Plesset many-body perturbation theory/correlation consistent polarized valence triple zeta second derivatives. The anharmonic part of the force field completes the theoretical study on the equilibrium structure, dipole moment, chlorine quadrupolar tensor, and harmonic force field previously carried out by the same authors. PMID- 15267540 TI - Photodissociation of bromobenzene, dibromobenzene, and 1,3,5-tribromobenzene. AB - Quantum chemical calculations have been performed on the ground state and several low-lying excited states of bromobenzene, ortho-, meta-, and para-dibromobenzene, and 1,3,5-tribromobenzene using high-level ab initio and hybrid density functional methods. Experimental observations of ultrafast predissociation in these molecules are clarified from extensive theoretical information about all low-energy potential-energy curves together with symmetry arguments. The intriguing observation that o- and m-dibromobenzene have two ultrafast predissociation channels while bromobenzene, p-dibromobenzene, and 1,3,5 tribromobenzene only have one such channel is explained from the calculated potential-energy curves. These show that the lowering of point-group symmetry from C2v to Cs along the main photodissociation reaction coordinate, which only occurs in o- and m-dibromobenzene, opens up a new predissociation channel. Dynamical quantum simulations based on the calculated potential-energy curves are used to estimate the coupling strength at the intersystem crossing point in bromobenzene. PMID- 15267541 TI - Interactions of Au cluster anions with oxygen. AB - Experimental and theoretical evidence is presented for the nondissociative chemisorption of O2 on free Au cluster anions (Aun-, n=number of atoms) with n=2, 4, 6 at room temperature, indicating that the stabilization of the activated di oxygen species is the key for the unusual catalytic activities of Au-based catalysts. In contrast to Aun- with n=2, 4, 6, O2 adsorbs atomically on Au monomer anions. For the Au monomer neutral, calculations based on density functional theory reveal that oxygen should be molecularly bound. On Au dimer and tetramer neutrals, oxygen is molecularly bound with the O-O bond being less activated with respect to their anionic counterparts, suggesting that the excess electron in the anionic state plays a crucial role for the O-O activation. We demonstrate that interplay between experiments on gas phase clusters and theoretical approach can be a promising strategy to unveil mechanisms of elementary steps in nanocatalysis. PMID- 15267542 TI - Topology of the distribution of zeros of the Husimi function in the LiNC/LiCN molecular system. AB - Phase space representations of quantum mechanics constitute useful tools to study vibrations in molecular systems. Among all possibilities, the Husimi function or coherent state representation is very widely used, its maxima indicating which regions of phase space are relevant in the dynamics of the system. The corresponding zeros are also a good indicator to investigate the characteristics of the eigenstates, and it has been shown how the corresponding distributions can discriminate between regular, irregular, and scarred wave functions. In this paper, we discuss how this result can be understood in terms of the overlap between coherent states and system eigenfunctions. PMID- 15267543 TI - Competitive C-H and O-D bond fission channels in the UV photodissociation of the deuterated hydroxymethyl radical CH2OD. AB - Photodissociation studies of the CH2OD radical in the region 28,000-41,000 cm(-1) (357-244 nm), which includes excitation to the 3s, 3p(x), and 3p(z) states, are reported. H and D photofragments are monitored by using resonance-enhanced multiphoton ionization (REMPI) from the onset of H formation at approximately 30,500 cm(-1) to the origin band region of the 3pz(2A")<--1 2A" transition at 41,050 cm(-1). Kinetic energy distributions P(ET) and recoil anisotropy parameters as a function of kinetic energy, beta(eff)(ET), are determined by the core sampling technique for the channels producing H and D fragments. Two dissociation channels are identified: (I) D+CH2O and (II) H+CHOD. The contribution of channel II increases monotonically as the excitation energy is increased. Based on the calculations of Hoffmann and Yarkony [J. Chem. Phys. 116, 8300 (2002)], it is concluded that conical intersections between 3s and the ground state determine the final branching ratio even when initial excitation accesses the 3px) and 3pz states. The different beta(eff) values obtained for channels I and II (-0.7 and approximately 0.0, respectively) are attributed to the different extents of out-of-plane nuclear motions in the specific couplings between 3s and the ground state (of A' and A'' symmetry, respectively) that lead to each channel. The upper limit to the dissociation energy of the C-H bond, determined from P(ET), is D0(C-H)=3.4+/-0.1 eV (79+/-2 kcal/mol). Combining this value with the known heats of formation of H and CH2OD, the heat of formation of CHOD is estimated at DeltaHf(0)(CHOD)=24+/-2 kcal/mol. PMID- 15267544 TI - High resolution photoabsorption and photofragment fluorescence spectroscopy of water between 10.9 and 12 eV. AB - This work presents absorption and photofragment fluorescence spectra of water (H2O and D2O) simultaneously recorded at rotational resolution and at room temperature, by means of a synchrotron radiation source in the range 10.9-12 eV, covering the nd intense series from n=3 to 8. The Rydberg states observed are assigned in the light of the most advanced theoretical work available [M. S. Child, Philos. Trans. R. Soc. London, Ser. A 355, 1623 (1997)], and by reference to the stretching and bending mode progressions. Comparison between absorption and fluorescence spectra is shown to reveal a fast predissociation mechanism involving the linear 3pb2 1B2 state, and permits the identification of its (0,14,0) vibrational level observed in the absorption spectra. PMID- 15267545 TI - Current density maps, magnetizability, and nuclear magnetic shielding tensors of bis-heteropentalenes. II. Furo-furan isomers. AB - Magnetic susceptibility and nuclear magnetic shielding at the nuclei of bis heteropentalenes formed by two furan units ([2,3-b], [3,2-b], [3,4-b], and [3,4 c] isomers) have been computed by several approximated techniques and a large Gaussian basis set to achieve near Hartree-Fock estimates. Ab initio models of the ring currents induced by a magnetic field normal to the molecular plane were obtained for the three isomeric systems of higher symmetry, showing that the pi electrons give rise to intense diamagnetic circulation. The pi currents are responsible for enhanced magnetic anisotropy and strong out-of-plane proton deshielding. The theoretical findings are used to build up a "diatropicity matrix" for two fused five-membered heterocyclic systems. PMID- 15267546 TI - Observation of the 3(3)Sigma(+)-X1Sigma+ transition of KRb by resonance enhanced two-photon ionization in a pulsed molecular beam: Hyperfine structures of 39K85Rb and 39K87Rb isotopomers. AB - The 3(3)Sigma(+)-X1Sigma+ transition of KRb is observed by resonance enhanced two photon ionization in a pulsed molecular beam. Hyperfine splittings of 39K85Rb and 39K87Rb isotopomers are observed. From the magnitude of hyperfine splittings, we found that the main hyperfine structure was dominated by the Fermi contact interaction between the Rb nuclear spin and the unpaired electron spin. The Fermi contact interaction constants were determined to be 291 MHz for 39K85Rb and 665 MHz for 39K87Rb. In the KRb 3(3)Sigma+ state the electron spin couples more strongly with the Rb nuclear spin than with other angular momenta, and the energy level structure is well described by the hyperfine angular momentum coupling scheme of the b(betaS) case. The molecular constants and the Rydberg-Klein-Rees potential energy curve of the 3(3)Sigma+ state were determined. PMID- 15267547 TI - Electron attachment to gas-phase uracil. AB - We present results about dissociative electron attachment (DEA) to gas-phase uracil (U) for incident electron energies between 0 and 14 eV using a crossed electron/molecule beam apparatus. The most abundant negative ion formed via DEA is (U-H)-, where the resonance with the highest intensity appears at 1.01 eV. The anion yield of (U-H)- shows a number of peaks, which can be explained in part as being due to the formation of different (U-H)- isomers. Our results are compared with high level ab initio calculations using the G2MP2 method. There was no measurable amount of a parent ion U-. We also report the occurrence of 12 other fragments produced by dissociative electron attachment to uracil but with lower cross sections than (U-H)-. In addition we observed a parasitic contaminating process for conditions where uracil was introduced simultaneously with calibrant gases SF6 and CCl4 that leads to a sharp peak in the (U-H)- cross section close to 0 eV. For (U-H)- and all other fragments we determined rough measures for the absolute partial cross section yielding in the case of (U-H)- a peak value of sigma (at 1.01 eV)=3 x 10(-20) m2. PMID- 15267548 TI - Ab initio studies of alkyl radical reactions: Combination and disproportionation reactions of CH3 with C2H5, and the decomposition of chemically activated C3H8. AB - This paper reports the first quantitative ab initio prediction of the disproportionation/combination ratio of alkyl+alkyl reactions using CH3+C2H5 as an example. The reaction has been investigated by the modified Gaussian-2 method with variational transition state or Rice-Ramsperger-Kassel-Marcus calculations for several channels producing (1) CH4+CH2CH2, (2) C3H8, (3) CH4CH3CH, (4) H2+CH3CHCH2, (5) H2+CH3CCH3, and (6) C2H6+CH2 by H-abstraction and association/decomposition mechanisms through singlet and triplet potential energy paths. Significantly, the disproportionation reaction (1) producing CH4+C2H4 was found to occur primarily by the lowest energy path via a loose hydrogen-bonding singlet molecular complex, H3CHC2H4, with a 3.5 kcal/mol binding energy and a small decomposition barrier (1.9 kcal/mol), instead of a direct H-abstraction process. Bimolecular reaction rate constants for the formation of the above products have been calculated in the temperature range 300-3000 K. At 1 atm, formation of C3H8 is dominant below 1200 K. Over 1200 K, the disproportionation reaction becomes competitive. The sum of products (3)-(6) accounts for less than 0.3% below 1500 K and it reaches around 1%-4% above 2000 K. The predicted rate constant for the disproportionation reaction with multiple reflections above the complex well, k1=5.04 x T(0.41) exp(429/T) at 200-600 K and k1=1.96 x 10(-20) T(2.45) exp(1470/T) cm3 molecule(-1) s(-1) at 600-3000 K, agrees closely with experimental values. Similarly, the predicted high-pressure rate constants for the combination reaction forming C3H8 and its reverse dissociation reaction in the temperature range 300-3000 K, k2(infinity)=2.41 x 10(-10) T(-0.34) exp(259/T) cm3 molecule(-1) s(-1) and k(-2)(infinity)=8.89 x 10(22) T(-1.67)exp(-46 037/T) s(-1), respectively, are also in good agreement with available experimental data. PMID- 15267549 TI - Saturated adsorption of CO and coadsorption of CO and O2 on AuN- (N=2-7) clusters. AB - A first-principles quantum chemistry method, based on the Kohn-Sham density functional theory, is used to investigate the adsorption of CO and O2 on small gas-phase gold cluster anions. The saturated adsorption of carbon monoxide on gold cluster anions AuN- (N=2-7) is discussed. The adsorption ability of CO reduces with the increase of the number of CO molecules bound to gold cluster anions, resulting in saturated adsorption at a certain amount of absorbed CO molecules, which is determined by geometric and electronic properties of gold clusters cooperatively. The effect of CO preadsorption on the electronic properties of gold cluster anions depends on the cluster size and the number of adsorbed CO, and the vertical detachment energies of CO-adsorbed gold cluster anions show a few changes with respect to corresponding pure gold cluster anions. The results indicate that the impinging adsorption of CO molecules may lead to geometry structure transformation on Au3- cluster. For the coadsorption of CO and O2 on Au2-, Au3- isomers, Au4-, and Au6-, we describe the cooperative adsorption between CO and O2, and find that the O2 dissociation is difficult on gas-phase gold cluster anions even with the preadsorption of CO. PMID- 15267550 TI - Accurate theoretical near-equilibrium potential energy and dipole moment surfaces of HgClO and HgBrO. AB - The complexes HgBrO and HgClO have been previously determined by ab initio methods to be strongly bound and were suggested to be important intermediates during mercury depletions events observed in the polar troposphere. In the present work accurate near-equilibrium potential energy surfaces (PESs) of these species are reported. The PESs are determined using accurate coupled cluster methods and a series of correlation consistent basis sets with subsequent extrapolation to the complete basis set limit. Additive corrections for both core valence correlation energy and relativistic effects are also included. The anharmonic ro-vibrational spectra of HgBrO and HgClO have been calculated in variational calculations. Strong infrared band strengths are predicted for all fundamentals in these species. The spin-orbit splitting dominates over the vibronic coupling effect in both HgClO and HgBrO. The Renner-Teller vibronic energy levels corresponding to the bending mode of these molecules are calculated via perturbation theory. PMID- 15267551 TI - Thermal decomposition of ethanol. III. A computational study of the kinetics and mechanism for the CH3+C2H5OH reaction. AB - The mechanism for the CH3+C2H5OH reaction has been investigated by the modified Gaussian-2 method based on the geometric parameters of the stationary points optimized at the B3LYP/6-311+G(d,p) level of theory. Five transition states have been identified for the production of CH4+CH3CHOH (TS1), CH4+CH3CH2O (TS2), CH4+CH2CH2OH (TS3), CH3OH+CH3CH2 (TS4), and CH3CH2OCH3+H (TS5) with the corresponding barriers 12.0, 13.2, 16.0, 44.7, and 49.9 kcal/mol, respectively. The predicted rate constants and branching ratios for the three lower-energy H abstraction reactions were calculated using the conventional and variational transition state theory with quantum-mechanical tunneling corrections for the temperature range 300-3000 K. The predicted total rate constant, kt=8.36 x 10( 76) T(20.00) exp(5258/T) cm3 mol(-1) s(-1) (300-600 K) and 6.10 x 10(-25) T(4.10)exp(-4058/T) cm3 mol(-1) s(-1) (600-3000 K), agrees closely with existing experimental data in the temperature range 403-523 K. Similarly, the predicted rate constants for CH3+CH3CD2OH and CD3+C2H5OD are also in reasonable agreement with available low temperature kinetic data. PMID- 15267552 TI - Optimal control of quantum non-Markovian dissipation: reduced Liouville-space theory. AB - An optimal control theory for open quantum systems is constructed containing non Markovian dissipation manipulated by an external control field. The control theory is developed based on a novel quantum dissipation formulation that treats both the initial canonical ensemble and the subsequent reduced control dynamics. An associated scheme of backward propagation is presented, allowing the efficient evaluation of general optimal control problems. As an illustration, the control theory is applied to the vibration of the hydrogen fluoride molecule embedded in a non-Markovian dissipative medium. The importance of control-dissipation correlation is evident in the results. PMID- 15267553 TI - Raman spectra of ionic liquids: interpretation via computer simulation. AB - Theoretical Raman spectra of the complex-forming ionic liquids LaCl3 and ScCl3, derived from molecular dynamics computer simulations, are presented. These simulations, which use polarizable ion interaction models, have previously been shown to predict structural properties in excellent agreement with diffraction experiments. The dependence of the polarizability of the melt on the ionic positions, which determines the Raman spectrum through the time dependence of the polarizability correlation function, is modeled on the basis of ab initio electronic structure calculations carried out on alkali chlorides. New simulation techniques are introduced in order to allow the spectrum to be calculated with acceptable statistics. The calculated spectra are in semiquantitative agreement with experimental data. The distinctive bands which appear in the spectra of such complex melts are linked to the vibrations of the transient coordination complexes which form in these melts and new interpretations for the origin of several well-known features are proposed. The simulations thus enable a link between the structure of a melt as perceived through Raman spectroscopy and through diffraction experiments to be made. PMID- 15267554 TI - Quantum dynamics in simple fluids. AB - We use quantum-correction factors to calculate approximately the quantum velocity time-correlation function (TCF) of supercritical Lennard-Jones argon from the classical TCF. We find that for this quite classical system, several different quantum-correction schemes yield essentially identical results for the real and imaginary parts of the quantum TCF, and also agree well with the recent forward backward semiclassical dynamics (FBSD) results of Wright and Makri [J. Chem. Phys. 119, 1634 (2003)]. We also consider a more quantum-mechanical fluid of lighter atoms (neon) at a lower temperature. In this case different quantum correction schemes give different results. FBSD calculations show that the harmonic quantum correction factor works the best for this system PMID- 15267555 TI - Infrared spectroscopy of acetone-water liquid mixtures. II. Molecular model. AB - In aqueous acetone solutions, the strong bathochromic shifts observed on the OH and CO stretch infrared (IR) bands are due to hydrogen bonds between these groups. These shifts were evaluated by factor analysis (FA) that separated the band components from which five water and five acetone principal factors were retrieved [J. Chem. Phys. 119, 5632 (2003)]. However, these factors were abstract making them difficult to interpret. To render them real an organization model of molecules is here developed whose abundances are compared to the experimental ones. The model considers that the molecules are randomly organized limited by the hydrogen bond network formed between the water hydrogen atoms and the acetone or water oxygen atoms, indifferently. Because the oxygen of water has two covalent hydrogen atoms which are hydrogen-bonded and may receive up to two hydrogen atoms from neighbor molecules hydrogen-bonded to it, three types of water molecules are found: OH2, OH3, and OH4 (covalent and hydrogen bonds). In the OH stretch region these molecules generate three absorption regimes composed of nu3, nu1, and their satellites. The strength of the H-bond given increases with the number of H-bonds accepted by the oxygen atom of the water H-bond donor, producing nine water situations. Since FA cannot separate those species that evolve concomitantly the nine water situations are regrouped into five factors, the abundance of which compared exactly to that retrieved by FA. From the factors' real spectra the OH stretch absorption are simulated to, respectively, give for the nu3 and nu1 components the mean values for OH2, 3608, 3508; OH3, 3473, 3282 and OH4, 3391, 3223 cm(-1). The mean separations from the gas-phase position which are respectively about 150, 330, and 400 cm(-1) are related to the vacancy of the oxygen electron doublets: two, one, and zero, respectively. No acetone hydrate that sequesters water molecules is formed. Similarly, acetone produces ten species, two of which evolve concomitantly. Spectral similarities further reduce these to five principal IR factors, the abundance of which compared adequately to the experimental results obtained from FA. The band assignment of the five-acetone spectra is given. PMID- 15267556 TI - Mode-coupling theory for reaction dynamics in liquids. AB - A theory for chemical reaction dynamics in condensed phase systems based on the generalized Langevin formalism of Grote and Hynes [J. Chem. Phys. 73, 2715 (1980)] is presented. A microscopic approach to calculate the dynamic friction is developed within the framework of a combination of kinetic and mode-coupling theories. The approach provides a powerful analytic tool to study chemical reactions in realistic condensed phase environments. The accuracy of the approach is tested for a model isomerization reaction in a Lennard-Jones fluid. Good agreement is obtained for the transmission coefficient at different solvent densities, in comparison with numerical simulations based on the reactive-flux approach. PMID- 15267557 TI - Towards an understanding of the heat capacity of liquids. A simple two-state model for molecular association. AB - A model for the temperature dependence of the isobaric heat capacity of associated pure liquids C(p,m)(o)(T) is proposed. Taking the ideal gas as a reference state, the residual heat capacity is divided into nonspecific C(p) (res,ns) and associational C(p) (res,ass) contributions. Statistical mechanics is used to obtain C(p)(res,ass) by means of a two-state model. All the experimentally observed C(p,m)(o)(T) types of curves in the literature are qualitatively described from the combination of the ideal gas heat capacity C(p)(id)(T) and C(p)(res,ass)(T). The existence of C(p,m)(o)(T) curves with a maximum is predicted and experimentally observed, for the first time, through the measurement of C(p,m)(o)(T) for highly sterically hindered alcohols. A detailed quantitative analysis of C(p,m)(o)(T) for several series of substances (n alkanes, linear and branched alcohols, and thiols) is made. All the basic features of C(p,m)(o)(T) at atmospheric and high pressures are successfully described, the model parameters being physically meaningful. In particular, the molecular association energies and the C(p)(res,ns) values from the proposed model are found to be in agreement with those obtained through quantum mechanical ab initio calculations and the Flory model, respectively. It is concluded that C(p,m)(o)(T) is governed by the association energy between molecules, their self association capability and molecular size. PMID- 15267558 TI - Homogeneous nucleation of droplets from supersaturated vapor in a closed system. AB - Kinetic equations describing homogeneous nucleation kinetics within standard model are solved numerically under the condition of a constant number of molecules in the considered system. It has consequences to decrease the supersaturation of the supersaturated vapor during the process of the formation of small droplets of a new phase. The decrease of supersaturation occurs in a short time and reaches some value which remains unchanged for a relatively long time (quasistationary regime), especially at lower initial supersaturations. This time interval decreases with increasing value of the initial supersaturation. In the quasistationary regime the nucleation rate reaches its stationary value. At higher initial supersaturation, the rate of formation of nuclei goes to some maximum value corresponding to the stationary nucleation rate and then decreases with time due to the decrease of supersaturation. PMID- 15267559 TI - Heat capacity and electron spin echo evidence for low frequency vibrational modes and lattice disorder in L-alanine at cryogenic temperatures. AB - With the view of understanding the low frequency (40-50 cm(-1)) motional processes in L-alanine around 4 K, we have carried out heat capacity (CP) and electron spin echo (ESE) measurements on L-alanine and L-alanine-d7. The obtained CP data show the so-called boson peak (seen as a maximum in CP/T3 versus T plots) in the low temperature region (1.8-20 K). The phase memory time, T(M), and spin lattice relaxation time, T1, of the spin probe, the so-called first stable alanine radical (SAR1), *CHCH3COOH, have been measured between 4 and 105 K. The obtained relaxation rate 1/T1 shows an anomalous increase which coincides with the emergence of a boson peak in the low temperature region (4-20 K). Together, the ESE and the CP data confirm the existence of a thermally activated dynamic orientational disorder in the lattices of both compounds below 20 K. The results help explain the discrepancy between the CP data from powders and single crystals of alanine, as well as the proanomalous relaxation mechanisms for SAR1 in these lattices, and they also provide a mechanism for the spin-lattice relaxation process for SAR1 at cryogenic temperatures. PMID- 15267560 TI - Temperature dependence of the hydrophobic hydration and interaction of simple solutes: an examination of five popular water models. AB - We examine five different popular rigid water models (SPC, SPCE, TIP3P, TIP4P, and TIP5P) using molecular dynamics simulations in order to investigate the hydrophobic hydration and interaction of apolar Lennard-Jones solutes as a function of temperature in the range between 275 and 375 K along the 0.1 MPa isobar. For all investigated models and state points we calculate the excess chemical potential for the noble gases and methane employing the Widom particle insertion technique. All water models exhibit too small hydration entropies, but show a clear hierarchy. TIP3P shows poorest agreement with experiment, whereas TIP5P is closest to the experimental data at lower temperatures and SPCE is closest at higher temperatures. As a first approximation, this behavior can be rationalized as a temperature shift with respect to the solvation behavior found in real water. A rescaling procedure inspired by the information theory model of Hummer et al. [Chem. Phys. 258, 349 (2000)] suggests that the different solubility curves for the different models and real water can be largely explained on the basis of the different density curves at constant pressure. In addition, the models that give a good representation of the water structure at ambient conditions (TIP5P, SPCE, and TIP4P) show considerably better agreement with the experimental data than the ones which exhibit less structured O-O correlation functions (SPC and TIP3P). In the second part of the paper we calculate the hydrophobic interaction between xenon particles directly from a series of 60 ns simulation runs. We find that the temperature dependence of the association is to a large extent related to the strength of the solvation entropy. Nevertheless, differences between the models seem to require a more detailed molecular picture. The TIP5P model shows by far the strongest temperature dependence. The suggested density rescaling is also applied to the chemical potential in the xenon-xenon contact-pair configuration, indicating the presence of a temperature where the hydrophobic interaction turns into purely repulsive. The predicted association for xenon in real water suggests the presence of a strong variation with temperature, comparable to the behavior found for TIP5P water. Comparing different water models and experimental data we conclude that a proper description of density effects is an important requirement for a water model to account correctly for the correct description of the hydrophobic effects. A water model exhibiting a density maximum at the correct temperature is desirable. PMID- 15267561 TI - Aspects of prewetting at nonplanar surfaces. AB - We employ Monte Carlo simulations in the grand canonical ensemble (GCEMC) to investigate the impact of nonplanarity of a solid substrate on the locus of the prewetting phase transition. The substrate is modelled as a periodic sequence of furrows of depth D and periodicity sx in the x direction; the furrows are infinitely long in the y direction. Our results indicate that a necessary prerequisite for a prewetting transition is the formation of a(n approximately) planar interface between molecularly thin films and an adjacent (bulk) gas. Thus, in general the prewetting transition is shifted to larger chemical potentials because the formation of a planar film-gas interface is more difficult next to a nonplanar compared with a planar solid surface. However, this shift turns out to be nonmonotonic depending on D on account of subtle packing effects manifested in the deviation of the local density Deltarho(x,Deltaz;D) at the nonplanar solid surface from that at a planar substrate. If D becomes sufficiently large prewetting as a discontinuous phase transition is suppressed because inside the furrow a highly ordered film forms that prevents a planar film-gas interface from forming. PMID- 15267562 TI - Reduced photobleaching of chromophores close to a metal surface. AB - The photobleaching of chromophores in front of a metal film is measured by recording the emitted fluorescence intensity from an ensemble of chromophores as a function of time. A strong dependence of the photostability on the distance from the metal surface is found. The experimental data are well described in a classical electromagnetic model with the additional assumption that photobleaching occurs at a constant rate from the excited state. The metal interface influences the photostability of the chromophores in two ways, first by altering the excitation rate by local enhancement of the electromagnetic field and second by altering the electromagnetic decay rate. PMID- 15267563 TI - Density functional theory investigation of benzenethiol adsorption on Au(111). AB - We have studied the adsorption of benzenethiol molecules on the Au(111) surface by using first principles total energy calculations. A single thiolate molecule is adsorbed at the bridge site slightly shifted toward the fcc-hollow site, and is tilted by 61 degrees from the surface normal. As for the self-assembled monolayer (SAM) structures, the (2 square root of 3 x square root of 3)R30 degrees herringbone structure is stabilized against the (square root 3 x square root 3)R30 degrees structure by large steric relaxation. In the most stable (2 square root 3 x square root 3)R30 degrees SAM structure, the molecule is adsorbed at the bridge site with the tilting angle of 21 degrees, which is much smaller compared with the single molecule adsorption. The van der Waals interaction plays an important role in forming the SAM structure. The adsorption of benzenethiolates induces the repulsive interaction between surface Au atoms, which facilitates the formation of surface Au vacancy. PMID- 15267564 TI - Dielectric response of modified Hubbard models with neutral-ionic and Peierls transitions. AB - The dipole P(F) of systems with periodic boundary conditions in a static electric field F is applied to one-dimensional Peierls-Hubbard models for organic charge transfer (CT) salts. Exact results for P(F) are obtained for finite systems of N=14 and 16 sites that are almost converged to infinite chains in deformable lattices subject to a Peierls transition. The electronic polarizability per site, alpha(el)=(partial differential P/partial differential F)0, of rigid stacks with alternating transfer integrals t(1+/-delta) diverges at the neutral-ionic transition for delta=0 but remains finite for delta>0 in dimerized chains. The Peierls or dimerization mode couples to charge fluctuations along the stack and results in large vibrational contributions alpha(vib) that are related to partial differential P/ partial differential delta and that peak sharply at the Peierls transition. The extension of P(F) to correlated electronic states yields the dielectric response kappa of models with neutral-ionic or Peierls transitions, where kappa peaks >100 are found with parameters used previously for variable ionicity rho and vibrational spectra of CT salts. The calculated kappa accounts for the dielectric response of CT salts based on substituted TTF's (tetrathiafulvalene) and substituted CA's (chloranil). The role of lattice stiffness appears clearly in models: soft systems have a Peierls instability at small rho and continuous crossover to large rho, while stiff stacks such as TTF CA have a first-order transition with discontinuous rho that is both a neutral ionic and Peierls transition. The transitions are associated with tuning the electronic ground state of insulators via temperature or pressure in experiments, or via model parameters in calculations. PMID- 15267565 TI - Density-functional study of organic-inorganic hybrid single crystal ZnSe(C2H8N2)(1/2). AB - Unusual properties (i.e., strong band dispersion, high carrier mobility, wide absorption-energy window, and sharp band-edge transition) that are desirable for hybrid-material electronics and for solar electric energy conversion are predicted to exist in the organic-inorganic chalcogenide single crystal ZnSe(C2H8N2)(1/2) by using density-functional calculations. A simple mechanism, namely that the band-edge electronic states of the hybrid composite is predominantly determined by the inorganic constituent, is revealed to be responsible for governing these properties. Suggestions for further engineering hybrid semiconductors are also provided PMID- 15267566 TI - Effects of the Gaussian energy dispersion on the statistics of polarons and bipolarons in conducting polymers. AB - We discuss the interpretation of usually broad oxidation peaks observed in electronically conducting polymers, in terms of the statistical distributions functions of polarons and bipolarons. The analysis is based on examining the chemical capacitance, that relates the change of concentration to a modification of the chemical potential of a given species, for different statistical models. We first review the standard models for single energy species that provide a nernstian dependence, and the limitations of these models are discussed. A new model that assumes a Gaussian distribution of energies related to molecular geometry fluctuations is suggested, and this model shows excellent agreement with the results of electrochemical oxidation of polypyrrole in quasiequilibrium conditions. From a fit of the data, it is found that the density of conjugated chain segments in polypyrrole, Ns approximately 10(21) cm(-3), shows a Gaussian distribution of half width sigma approximately 170 meV, tentatively attributed to bipolaron formation energies. PMID- 15267567 TI - Integral equation study of a Stockmayer fluid adsorbed in polar disordered matrices. AB - Based on replica integral equations in the (reference) hypernetted chain approximation we investigate the structural features and phase properties of a dipolar Stockmayer fluid confined to a disordered dipolar matrix. The integral equations are applied to the homogeneous high-temperature phase where the system is globally isotropic. At low densities we find the influence of dipolar interactions between fluid (f) and matrix (m) particles to be surprisingly similar to the previously investigated effect of attractive isotropic (fm) interactions: the critical temperature of the vapor-liquid transition decreases with increasing (fm) coupling, while the critical density increases. The anisotropic nature of the dipolar (fm) interactions turns out to play a more dominant role at high fluid densities where we observe a pronounced sensitivity in the dielectric constant and a strong degree of local orientational ordering of the fluid particles along the local fields generated by the matrix. Moreover, an instability of the dielectric constant, which is a precursor of ferroelectric ordering occurring both in bulk Stockmayer fluids and in fluids in nonpolar matrices, is observed only for very small dipolar (fm) couplings. PMID- 15267568 TI - Hybrid atomistic-coarse-grained treatment of thin-film lubrication. I. AB - A technique that melds an atomistic description of the interfacial region with a coarse-grained description of the far regions of the solid substrates is presented and applied to a two-dimensional model contact consisting of planar solid substrates separated by a monolayer fluid film. The hybrid method yields results in excellent agreement with the "exact" (i.e., fully atomistic) results. The importance of a proper accounting for the elastic response of the substrates, which is reliably and efficiently accomplished through coarse-graining of the far regions, is demonstrated. PMID- 15267569 TI - A new cell model for equation of state of polyethylene melt and n-alkane. AB - The connectivity of successive carbon atoms in polymer decreases the degrees of freedom, and hence, the external degrees of freedom should correspond only to translational motion. We therefore, introduce a coarse-grained particle of a few successive monomers. A cell model (or the hard core model with attractive potential) for the particles can accordingly be used for the derivation of the equation of state of polymers. Modifying the classical cell model by Lennard Jones and Devonshire, we construct a new equation of state for polyethylene melt and for liquid n-alkane; the free volume term is modified by using the Sutherland potential instead of the Lennard-Jones potential. The characteristic quantities P*, V*0, and T* in the equation of state are almost independent of temperature; the principle of corresponding state holds well. Since our equation of state contains the external degrees of freedom explicitly, we can evaluate the external degrees of freedom, c, for CH2. The value of c for the coarse-grained particle is equal to 1, and hence the particle is composed of 1/c repeating units. The linear length of the particle evaluated, 4.09 A at 0 K, is consistent with that obtained by neutron and x-ray scatterings. PMID- 15267570 TI - Annealing contour Monte Carlo algorithm for structure optimization in an off lattice protein model. AB - We present a space annealing version for a contour Monte Carlo algorithm and show that it can be applied successfully to finding the ground states for an off lattice protein model. The comparison shows that the algorithm has made a significant improvement over the pruned-enriched-Rosenbluth method and the Metropolis Monte Carlo method in finding the ground states for AB models. For all sequences, the algorithm has renewed the putative ground energy values in the two dimensional AB model and set the putative ground energy values in the three dimensional AB model. PMID- 15267571 TI - Large effect of polydispersity on defect concentrations in colloidal crystals. AB - We compute the equilibrium concentration of stacking faults and point defects in polydisperse hard-sphere crystals. We find that, while the concentration of stacking faults remains similar to that of monodisperse hard-sphere crystals, the concentration of vacancies decreases by about a factor of 2. Most strikingly, the concentration of interstitials in the maximally polydisperse crystal may be some six orders of magnitude larger than in a monodisperse crystal. We show that this dramatic increase in interstitial concentration is due to the increased probability of finding small particles and that the small-particle tail of the particle size distribution is crucial for the interstitial concentration in a colloidal crystal. PMID- 15267572 TI - Folding probabilities: a novel approach to folding transitions and the two dimensional Ising-model. AB - The theoretical concept of folding probability, p(fold), has proven to be a useful means to characterize the kinetics of protein folding. Here, we illustrate the practical importance of p(fold) and demonstrate how it can be determined theoretically. We derive a general analytical expression for p(fold) and show how it can be estimated from simulations for systems where the transition rates between the relevant microstates are not known. By analyzing the Ising model we are able to determine the scaling behavior of the numerical error in the p(fold) estimate as function of the number of analyzed Monte Carlo runs. We apply our method to a simple, newly developed protein folding model for the formation of alpha helices. It is demonstrated that our technique highly parallelizes the calculation of p(fold) and that it is orders of magnitude more efficient than conventional approaches. PMID- 15267573 TI - Thermodynamics of lattice heteropolymers. AB - We calculate thermodynamic quantities of hydrophobic-polar (HP) lattice proteins by means of a multicanonical chain-growth algorithm that connects the new variants of the Pruned-Enriched Rosenbluth Method and flat histogram sampling of the entire energy space. Since our method directly simulates the density of states, we obtain results for thermodynamic quantities of the system for all temperatures. In particular, this algorithm enables us to accurately simulate the usually difficult accessible low-temperature region. Therefore, it becomes possible to perform detailed analyses of the low-temperature transition between ground states and compact globules. PMID- 15267574 TI - Helium nanodroplet isolation spectroscopy of perylene and its complexes with oxygen. PMID- 15267575 TI - Comment on "Unraveling the mysteries of metastable O4*" [J. Chem. Phys. 110, 6095 (1999)]. PMID- 15267578 TI - The surprising nonlinear optical properties of conjugated polyyne oligomers. AB - Polyynes represent a unique class of conjugated organic compounds. The third order nonlinear optical response of polyynes has been extensively modeled theoretically, and it is generally believed that the increase in molecular second hyperpolarizability (gamma) as a function of length for polyynes should be lower than that for polyenes. Experimental evidence to test this prediction, however, has been absent. We have synthesized conjugated polyynes that contain up to 20 consecutive sp-hybridized carbons, and we have determined their nonresonant gamma values as a function of the number of acetylene repeat units (n). These gamma values demonstrate a power-law behavior versus n(gamma approximately n(4.28+/ 0.13)), with an exponent that is both larger than theoretically predicted for polyynes and substantially higher than that observed for polyenes or polyenynes. Furthermore, no saturation of the linear or nonlinear optical properties is observed. PMID- 15267579 TI - The Huggins band of ozone: unambiguous electronic and vibrational assignment. AB - The Huggins band of ozone is investigated by means of exact dynamics calculations using a new (diabatic) potential energy surface for the (1)B(2) state. The remarkable agreement with the measured spectrum strongly suggests that the Huggins band is due to the two C(s) potential wells of the (1)B(2) state. The vibrational assignment, based on the nodal structure of wave functions, supports the most recent experimental assignment. PMID- 15267580 TI - Bohmian dynamics on subspaces using linearized quantum force. AB - In the de Broglie-Bohm formulation of quantum mechanics the time-dependent Schrodinger equation is solved in terms of quantum trajectories evolving under the influence of quantum and classical potentials. For a practical implementation that scales favorably with system size and is accurate for semiclassical systems, we use approximate quantum potentials. Recently, we have shown that optimization of the nonclassical component of the momentum operator in terms of fitting functions leads to the energy-conserving approximate quantum potential. In particular, linear fitting functions give the exact time evolution of a Gaussian wave packet in a locally quadratic potential and can describe the dominant quantum-mechanical effects in the semiclassical scattering problems of nuclear dynamics. In this paper we formulate the Bohmian dynamics on subspaces and define the energy-conserving approximate quantum potential in terms of optimized nonclassical momentum, extended to include the domain boundary functions. This generalization allows a better description of the non-Gaussian wave packets and general potentials in terms of simple fitting functions. The optimization is performed independently for each domain and each dimension. For linear fitting functions optimal parameters are expressed in terms of the first and second moments of the trajectory distribution. Examples are given for one-dimensional anharmonic systems and for the collinear hydrogen exchange reaction. PMID- 15267581 TI - Variational second-order Moller-Plesset theory based on the Luttinger-Ward functional. AB - In recent years there have been some rather successful applications of a new variational technique for calculating the total energies of electronic systems. The new method is based on many-body perturbation theory and uses the one electron Green function as the basic "variable" rather than the wave function of traditional variational calculations. It is the purpose of the present work to promote the new methods within the realm of traditional theoretical chemistry by demonstrating their utility for calculating the correlation energies of a number of atoms at a level corresponding to second-order Moller-Plesset perturbation theory. The generalization to any desired order of perturbation theory is not hard to accomplish. PMID- 15267582 TI - The importance of three-body terms in the fragment molecular orbital method. AB - A previously proposed two-body fragment molecular orbital method based on the restricted Hartree-Fock (RHF) method was extended to include explicit three-body terms. The accuracy of the method was tested on a set of representative molecules: (H(2)O)(n), n=16, 32, and 64, as well as alpha and beta n-mers of alanine, n=10, 20, and 40, using STO-3G, 3-21G, 6-31G, and 6-31++G(**) basis sets. Two- and three-body results are presented separately for assigning one and two molecules (or residues) per fragment. Total energies are found to differ from the regular RHF method by at most DeltaE(2/1)=0.06, DeltaE(2/2)=0.04, DeltaE(3/1)=0.02, and DeltaE(3/2)=0.003 (a.u.); rms energy gradients differ by at most DeltaG(2/1)=0.0015, DeltaG(2/2)=0.000 75, DeltaG(3/1)=0.000 20, and DeltaG(3/2)=0.000 10 (a.u./bohr), and rms dipole moments are reproduced with at most deltaD(2/1)=3.7, deltaD(2/2)=3.4, deltaD(3/1)=2.6, and deltaD(3/2)=3.1 (%) relative error, where the subscript notation n/m refers to the n-body method based on m molecules (residues) per fragment. A few of the largest three-body calculations were performed with a separated trimer approximation, which presumably somewhat lowered the accuracy of mostly dipole moments which are very sensitive to slight variations in the density distribution. The proposed method is capable of providing sufficient chemical accuracy while providing detailed information on many-body interactions. PMID- 15267583 TI - Analytic second derivatives for general coupled-cluster and configuration interaction models. AB - Analytic second derivatives of energy for general coupled-cluster (CC) and configuration-interaction (CI) methods have been implemented using string-based many-body algorithms. Wave functions truncated at an arbitrary excitation level are considered. The presented method is applied to the calculation of CC and CI harmonic frequencies and nuclear magnetic resonance chemical shifts up to the full CI level for some selected systems. The present benchmarks underline the importance of higher excitations in high-accuracy calculations. PMID- 15267584 TI - Unified treatment of chemical and van der Waals forces via symmetry-adapted perturbation expansion. AB - We propose a symmetry-adapted perturbation theory (SAPT) expansion of the intermolecular interaction energy which in a finite order provides the correct values of the constants determining the asymptotics of the interaction energy (the van der Waals constants) and is convergent when the energy of the interacting system is submerged in the continuum of Pauli-forbidden states-the situation common when at least one of the monomers has more than two electrons. These desirable features are achieved by splitting the intermolecular electron nucleus attraction terms of the Hamiltonian into regular (long-range) and singular (short-range) parts. In the perturbation theory development, the regular part is treated as in the conventional polarization theory, which guarantees the correct asymptotics of the interaction energy, while the singular part is weakened sufficiently by an application of permutational symmetry projectors so that a convergent perturbation series is obtained. The convergence is demonstrated numerically, for both the chemical and van der Waals minima, by performing high-order calculations of the interaction energy of the ground-state lithium and hydrogen atoms-the simplest system for which the physical ground state is submerged in the Pauli-forbidden continuum. The obtained expansion enables a systematic extension of SAPT calculations beyond second order with respect to the intermolecular interaction operator. PMID- 15267585 TI - Simulation of environmental effects on coherent quantum dynamics in many-body systems. AB - In this paper we describe an application of the trajectory-based semiclassical Liouville method for modeling coherent molecular dynamics on multiple electronic surfaces to the treatment of the evolution and decay of quantum electronic coherence in many-body systems. We consider a model representing the coherent evolution of quantum wave packets on two excited electronic surfaces of a diatomic molecule in the gas phase and in rare gas solvent environments, ranging from small clusters to a cryogenic solid. For the gas phase system, the semiclassical trajectory method is shown to reproduce the evolution of the electronic-nuclear coherence nearly quantitatively. The dynamics of decoherence are then investigated for the solvated systems using the semiclassical approach. It is found that, although solvation in general leads to more rapid and extensive loss of quantum coherence, the details of the coupled system-bath dynamics are important, and in some cases the environment can preserve or even enhance quantum coherence beyond that seen in the isolated system. PMID- 15267586 TI - Revealing the roles of Hamiltonian coupling in bound-state quantum systems. AB - A Hamiltonian coupling identification (HCI) technique is introduced to reveal the independent and cooperative roles of Hamiltonian matrix elements in determining the bound-state energies of quantum systems. The HCI technique operates by encoding each Hamiltonian matrix element with a unique modulation signal, producing a nonlinear signature in the energy eigenvalues that may be decoded to reveal the contributing coupling structure in the Hamiltonian. The HCI technique is capable of exploring the roles of Hamiltonian coupling structure within and beyond the convergence limits of standard perturbation theory expansions. The flexibility residing in the encoding and decoding processes may be exploited to tailor the analysis to meet the desired degree of sought-after information about the Hamiltonian coupling structure. HCI, based on a Fourier encoding and decoding scheme, is illustrated by extracting information on the role of coupling interactions in the potential matrix elements of several simple model systems. PMID- 15267587 TI - Computing minimal entropy production trajectories: an approach to model reduction in chemical kinetics. AB - Advanced experimental techniques in chemistry and physics provide increasing access to detailed deterministic mass action models for chemical reaction kinetics. Especially in complex technical or biochemical systems the huge amount of species and reaction pathways involved in a detailed modeling approach call for efficient methods of model reduction. These should be automatic and based on a firm mathematical analysis of the ordinary differential equations underlying the chemical kinetics in deterministic models. A main purpose of model reduction is to enable accurate numerical simulations of even high dimensional and spatially extended reaction systems. The latter include physical transport mechanisms and are modeled by partial differential equations. Their numerical solution for hundreds or thousands of species within a reasonable time will exceed computer capacities available now and in a foreseeable future. The central idea of model reduction is to replace the high dimensional dynamics by a low dimensional approximation with an appropriate degree of accuracy. Here I present a global approach to model reduction based on the concept of minimal entropy production and its numerical implementation. For given values of a single species concentration in a chemical system all other species concentrations are computed under the assumption that the system is as close as possible to its attractor, the thermodynamic equilibrium, in the sense that all modes of thermodynamic forces are maximally relaxed except the one, which drives the remaining system dynamics. This relaxation is expressed in terms of minimal entropy production for single reaction steps along phase space trajectories. PMID- 15267588 TI - Meta-generalized gradient approximation: explanation of a realistic nonempirical density functional. AB - Tao, Perdew, Staroverov, and Scuseria (TPSS) have constructed a nonempirical meta generalized gradient approximation (meta-GGA) [Phys. Rev. Lett. 91, 146401 (2003)] for the exchange-correlation energy, imposing exact constraints relevant to the paradigm densities of condensed matter physics and quantum chemistry. Results of their extensive tests on molecules, solids, and solid surfaces are encouraging, suggesting that this density functional achieves uniform accuracy for diverse properties and systems. In the present work, this functional is explained and details of its construction are presented. In particular, the functional is constructed to yield accurate energies under uniform coordinate scaling to the low-density or strong-interaction limit. Its nonlocality is displayed by plotting the factor F(xc) that gives the enhancement relative to the local density approximation for exchange. We also discuss an apparently harmless order-of-limits problem in the meta-GGA. The performance of this functional is investigated for exchange and correlation energies and shell-removal energies of atoms and ions. Non-self-consistent molecular atomization energies and bond lengths of the TPSS meta-GGA, calculated with GGA orbitals and densities, agree well with those calculated self-consistently. We suggest that satisfaction of additional exact constraints on higher rungs of a ladder of density functional approximations can lead to further progress. PMID- 15267589 TI - A complete nearest-neighbor force field model for C60. AB - A force field model is developed for C(60) that features 13 force constants representing all interactions between nearest-neighboring atoms. The model is compared with, and tested against, other force field models in the literature. Force constants for C(60) are then deduced by fitting the model to the 14 known optically accessible vibrational frequencies of the molecule. Finally, the model is fitted to two existing theoretical calculations of the complete vibrational spectrum of C(60). Fair agreement is obtained with the theoretical calculations, implying that interactions with atoms other than nearest neighbors are small. PMID- 15267590 TI - Spectroscopy of the OC-HF hydrogen-bonded complex at vHF=3. AB - The v(HF)=3 levels of the linear OC-HF complex are observed in the range of 10,800-11,500 cm(-1) using intracavity Ti-sapphire laser-induced fluorescence. The vibrational predissociation linewidths of both (30000) and (3001(1)0) states exceed 5 GHz; thus, the measured spectra are not rotationally resolvable. Under the assumption that these levels are not strongly perturbed, the rotational constants of the two levels are determined to be 0.1100(1) cm(-1) for (30000), 0.1081(1), and 0.1065(1) cm(-1) for f and e sublevels of (3001(1)0), respectively, through band contour fitting. The (30000)<--(00000) band origin is at 10,894.46(1) cm(-1), showing a HF wave number redshift of 478.3 cm(-1). The 4.07 redshift ratio of v(HF)=3 to that of v(HF)=1 indicates a significantly nonlinear increase of the intermolecular interaction energy through HF valence excitation. An ab initio interaction potential surface for HF valence coordinates varying from 0.8 to 1.25 A is used to examine vibrational dynamics. The HF valence vibration v(1) is treated perturbatively, showing that the vibrational redshifts are determined essentially in first order with only a very small second order contribution. The (3001(1)0)<--(00000) combination transition is observed with the band origin at 11,432.66(1) cm(-1), giving the HF intermolecular bending mode to be 538.2 cm(-1). The high frequency of this vibration, compared to that in similar HF complexes, shows the strong angular anisotropy of the intermolecular interaction potential of OC-HF with respect to the HF subunit. The lifetime of the (3001(1)0) level increases to 28 ps from 14 ps for (30000). PMID- 15267591 TI - High-level ab initio studies of the electronic excited states of the hydroxyl radical and water-hydroxyl complex. AB - The lowest-energy electronic transitions in the hydroxyl radical and the hydrogen bound complex H(2)O.HO have been studied using ab initio methods. We have used the complete active-space self-consistent field and multireference configuration interaction (MRCI) methods to calculate vertical excitation energies and oscillator strengths. At the MRCI level the lowest-lying (2)Sigma(+)<--(2)Pi electronic transition is redshifted by about 2500 cm(-1) upon formation of the H(2)O.HO complex. We propose that this transition could be used to identify the complex in the gas phase, which in turn could be used to examine the role of H(2)O.HO in atmospheric reactions. PMID- 15267592 TI - Fourier transform microwave spectroscopy of the Rg-SH(2Pi(i)) complexes (Rg:Ne, Kr): determination of the intermolecular potential energy surfaces. AB - Pure rotational spectra of Ne-SH and Kr-SH have been studied by Fourier transform microwave spectroscopy. R-branch transitions in the lower-spin component (Omega=3/2) corresponding to a linear (2)Pi(i) radical were observed for J(")=1.5 4.5 in the region 11-25 GHz for Ne-SH and for J(")=1.5-6.5 in the region 5-20 GHz for Kr-SH, respectively, with parity doublings and hyperfine splittings associated with the H nucleus. Although the spectral pattern of Kr-SH is relatively regular, that of Ne-SH is irregular with the J dependence of the parity doublings quite different from other Rg-SH or Ar-OH complexes. Two dimensional intermolecular potential energy surfaces (IPSs) for both of the species have been determined from the least-squares fittings of the observed rotational transitions utilizing results of high-level ab initio calculations. These IPSs reproduce the observed transition frequencies within the experimental error and provide accurate knowledge on the intermolecular interaction and internal dynamics. Systematic comparisons of Rg-SH complexes have clarified various features of this series of complexes. PMID- 15267593 TI - Vacuum ultraviolet pulsed-field ionization-photoelectron study of H2S in the energy range of 10-17 eV. AB - Vacuum ultraviolet pulsed-field ionization-photoelectron (PFI-PE) spectra of H(2)S have been recorded at PFI-PE resolutions of 0.6-1.0 meV in the energy range of 10-17 eV using high-resolution synchrotron radiation. The PFI-PE spectrum, which covers the formation of the valence electronic states H(2)S(+) (X (2)B(1), A (2)A(1), and B (2)B(2)), is compared to the recent high-resolution He I photoelectron spectra of H(2)S obtained by Baltzer et al. [Chem. Phys. 195, 403 (1995)]. In addition to the overwhelmingly dominated origin vibrational band, the PFI-PE spectrum for H(2)S(+)(X (2)B(1)) is found to exhibit weak vibrational progressions due to excitation of the combination bands in the nu(1) (+) symmetric stretching and nu(2) (+) bending modes. While the ionization energy (IE) for H(2)S(+)(X (2)B(1)) obtained here is in accord with values determined in previously laser PFI-PE measurements, the observation of a new PFI-PE band at 12.642+/-0.001 eV suggests that the IE for H(2)S(+)(A (2)A(1)) may be 0.12 eV lower than that reported in the He I study. The simulation of rotational structures resolved in PFI-PE bands shows that the formation of H(2)S(+)(X (2)B(1)) and H(2)S(+)(A (2)A(1)) from photoionization of H(2)S(X (1)A(1)) is dominated by type-C and type-B transitions, respectively. This observation is consistent with predictions of the multichannel quantum defect theory. The small changes in rotational angular momentum observed are consistent with the dominant atomiclike character of the 2b(1) and 5a(1) molecular orbitals of H(2)S. The PFI PE measurement has revealed perturbations of the (0, 6, 0) K(+)=3 and (0, 6, 0) K(+)=4 bands of H(2)S(+)(A (2)A(1)). Interpreting that these perturbations arise from Renner-Teller interactions at energies close to the common barriers to linearity of the H(2)S(+) (X (2)B(1) and A (2)A(1)) states, we have deduced a barrier of 23,209 cm(-1) for H(2)S(+)(X (2)B(1)) and 5668 cm(-1) for H(2)S(+)(A (2)A(1)). The barrier of 23 209 cm(-1) for H(2)S(+)(X (2)B(1)) is found to be in excellent agreement with the results of previous studies. The vibrational PFI-PE bands for H(2)S(+)(B (2)B(2)) are broad, indicative of the predissociative nature of this state. PMID- 15267594 TI - Photolysis of oxalyl chloride (ClCO)2 at 193 nm: emission of CO(vH2O+H3O(+). AB - We report quantum and quasiclassical calculations of proton transfer in the reaction H(3)O(+)+H(2)O in three degrees of freedom, the two OH(+) bond lengths and the OH(+)O angle. The reduced dimensional potential energy surface is obtained from the full dimensional OSS3(p) energy function of H(5)O(2) (+) [L. Ojamae, I. Shavitt, and S. J. Singer, J. Chem. Phys. 109, 5547 (1998)], with an additional long-range correction to reproduce the correct ion-molecule interaction. This surface is used to perform both quasiclassical trajectory and quantum reactive scattering calculations of the zero total angular momentum cumulative reaction probability and cross sections for initial rotational states 0, 1, and 2. Comparison of these quantities are made to assess the importance of quantum effects in this reduced dimensional reaction. Additional quasiclassical cross sections are calculated to obtain the thermal rate constant for the reaction. PMID- 15267603 TI - Electron attachment and detachment and the electron affinity of cyclo-C4F8. AB - New measurements have been made of rate constants for electron attachment to c C(4)F(8) (octafluorocyclobutane) and thermal electron detachment from the parent anion, c-C(4)F(8) (-), over the temperature range 298-400 K in 133 Pa of He gas in a flowing-afterglow Langmuir-probe apparatus. From these data the electron affinity for c-C(4)F(8) was determined, EA(c-C(4)F(8))=0.63+/-0.05 eV. The motivation was to resolve a discrepancy between our earlier EA estimate and a higher value (EA=1.05+/-0.10 eV) reported from a recent experiment of Hiraoka et al. [J. Chem. Phys. 116, 7574 (2002)]. The electron attachment rate constant is 9.3+/-3.0x10(-9) cm(3) s(-1) at 298 K. The electron detachment rate constant is negligible at room temperature but climbs to 1945+/-680 s(-1) at 400 K. G3(MP2) calculations were carried out for the neutral (D(2d), (1)A(1)) and anion (D(4h), (2)A(2u)) and yielded EA(c-C(4)F(8) (-))=0.595 eV. Bond energies were also calculated for loss of F from c-C(4)F(8) and loss of F or F(-) from c-C(4)F(8) ( ). From these, dissociative electron attachment is found to be endothermic by at least 1.55 eV. PMID- 15267604 TI - Excitation energy transfer (EET) between molecules in condensed matter: a novel application of the polarizable continuum model (PCM). AB - We present a quantum-mechanical theory to study excitation energy transfers between molecular systems in solution. The model is developed within the time dependent (TD) density-functional theory and the solvent effects are introduced in terms of the polarizable continuum model (PCM). Unique characteristic of this model is that both "reaction field" and screening effects are included in a coherent and self-consistent way. This is obtained by introducing proper solvent specific operators in the Kohn-Sham equations and in the corresponding TD scheme. The solvation model exploits the integral equation formalism (IEF) version of PCM and it defines the solvent operators on a molecular cavity modeled on the real three-dimensional (3D) structure of the solute systems. Applications to EET in dimers of ethylene and naphtalene are presented and discussed. PMID- 15267605 TI - Influence of strain on transport in dense Lennard-Jones systems. AB - We study the shear stress relaxation and temperature dependence of the diffusion coefficient, viscosity, and thermal conductivity along a high-density Lennard Jones isochore of the reduced density of 1.0, as it crosses the freezing and melting lines, in equilibrium and under constant strain. PMID- 15267606 TI - Reorganization energy associated with small polaron mobility in iron oxide. AB - The reorganization energy is an important quantity controlling electron transfer rates. The internal contribution arising from the energy to reorganize donor/acceptor bonds can be evaluated by the "direct" and "4-point" methods. We examine how spatial separation leading to the noninteracting character of the donor and acceptor affects the reorganization energy. We show that the direct method captures contributions from interaction of the donor and acceptor while the 4-point method does not, and the two methods converge at large separation. Comparing reorganization energies determined by the two methods yields a measure of the degree of interaction between the initial and final states. The analysis is illustrated in the characterization of small polarons in iron oxides. PMID- 15267607 TI - van der Waals-Tonks-type equations of state for hard-hypersphere fluids in four and five dimensions. AB - Recently, we developed accurate van der Waals-Tonks-type equations of state for hard-disk and hard-sphere fluids by using the known virial coefficients. In this paper, we derive the van der Waals-Tonks-type equations of state. We further apply these equations of state to hard-hypersphere fluids in four and five dimensions. In the low-density fluid regime, these equations of state are in good agreement with the simulation results and existing equations of state. PMID- 15267608 TI - Ab initio based force field and molecular dynamics simulations of crystalline TATB. AB - An all-atom force field for 1,3,5-triamino-2,4,6-trinitrobenzene (TATB) is presented. The classical intermolecular interaction potential for TATB is based on single-point energies determined from high-level ab initio calculations of TATB dimers. The newly developed potential function is used to examine bulk crystalline TATB via molecular dynamics simulations. The isobaric thermal expansion and isothermal compression under hydrostatic pressures obtained from the molecular dynamics simulations are in good agreement with experiment. The calculated volume-temperature expansion is almost one dimensional along the c crystallographic axis, whereas under compression, all three unit cell axes participate, albeit unequally. PMID- 15267609 TI - Periodic change of viscosity and density in an oscillating chemical reaction. AB - It was found that the periodic change of the solution viscosity and density was generated in the Belousov-Zhabotinsky (BZ) reaction. This rhythmic phenomenon was observed in both the iron catalyst [[Fe(Phen)(3)](2+)-[Fe(Phen)(3)](3+)] and the cerium catalyst [Ce(III)-Ce(IV)] system, where the solution viscosity and density were synchronized with the redox potential in the in-phase mode. However, the time delay existed between the redox potential and the solution viscosity and density. The behavior of the BZ reaction was also monitored in the presence of the nonionic surfactant. This experiment revealed that, beyond the critical micelle concentration, the phase between the redox potential and the solution viscosity and density was synchronized into the antiphase mode. We suggested that the variation of the catalyst drove the oscillation of the solution viscosity and density in the BZ reaction. PMID- 15267610 TI - Experimental verification of the Smoluchowski theory for a bimolecular diffusion controlled reaction in liquid phase. AB - We report experimental verification of the Smoluchowski theory for diffusion controlled reactions in solution at the steady-state limit. We have determined both the diffusion coefficients and the self-termination reaction rates of the diphenylmethyl radical simultaneously. Smoluchowski theory is insufficient to discuss the reaction rate for the self-termination reaction of the diphenylmethyl radical, so the reaction rate of an encounter complex based on the Collins Kimball treatment is estimated. PMID- 15267611 TI - Is styrene planar in liquid phases? AB - The proton NMR spectra of two (13)C-labeled isotopomers of styrene dissolved in two liquid crystalline solvents have been obtained and analyzed to yield four sets each of 24 dipolar couplings. These couplings were then used to investigate the structure of the ring and the ene fragments of the molecule, and the position of the maximum, phi(0), in the ring-ene bond rotational probability distribution. To do this, the effect on the dipolar couplings of small-amplitude vibrational motion was taken into account using vibrational wave functions calculated by molecular orbital and density functional methods. It is concluded that the NMR data are consistent with the ring fragment, averaged over the ring-ene rotation, planar, while the ene fragment is not. The value of phi(0) is found to be 18.0 degrees +/-0.2 degrees for the two solutions, compared with a value of 27 degrees calculated by the molecular method MP2/6-31G(*). PMID- 15267612 TI - On the influence of semirigid environments on proton transfer along molecular chains. AB - The dynamics of proton transfer along ammonia chains (chemical composition N(x)H(+)(3x+1), x=2, 4, and 6) in a constraining environment is investigated by ab initio molecular dynamics simulations. A carbon nanotube of defined length and diameter is used as an idealized constraining environment such that the ammonia chain is forced to maintain its quasilinear geometry. It is found that, although the energetics of proton transport shows considerable energetic barriers, proton translocation along the wire is possible at finite temperature for all chain lengths studied. The proton transport involves rotational reorientation of the proton-carrying ammonia molecule. High level ab initio calculations (MP2/aug-cc pVTZ) yield barriers for internal rotation of 9.1 kcal/mol for NH(4) (+)-NH(3) and 11.7 kcal/mol for OH(3) (+)-OH(2), respectively. The infrared spectrum calculated from the dipole-dipole autocorrelation function shows distinct spectral features in the regions (2000-3000 cm(-1)) where the NHN proton transfer mode is expected to absorb. Assigning moderate opposite total charges between 0.002 and 0.2e to the carbon atoms at the end caps of the nanotube leads to a considerable speedup of the proton transfer. PMID- 15267613 TI - Energy of charged states in the acetanilide crystal: trapping of charge-transfer states at vacancies as a possible mechanism for optical damage. AB - Calculations for the acetanilide crystal yield the effective polarizability (16.6 A(3)), local electric field tensor, effective dipole moment (5.41 D), and dipole dipole energy (-12.8 kJ/mol). Fourier-transform techniques are used to calculate the polarization energy P for a single charge in the perfect crystal (-1.16 eV); the charge-dipole energy W(D) is zero if the crystal carries no bulk dipole moment. Polarization energies for charge-transfer (CT) pairs combine with the Coulomb energy E(C) to give the screened Coulomb energy E(scr); screening is nearly isotropic, with E(scr) approximately E(C)/2.7. For CT pairs W(D) reduces to a term deltaW(D) arising from the interaction of the charge on each ion with the change in dipole moment on the other ion relative to the neutral molecule. The dipole moments calculated by density-functional theory methods with the B3LYP functional at the 6-311++G(**) level are 3.62 D for the neutral molecule, changing to 7.13 D and 4.38 D for the anion and cation, relative to the center of mass. Because of the large change in the anion, deltaW(D) reaches -0.9 eV and modifies the sequence of CT energies markedly from that of E(scr), giving the lowest two CT pairs at -1.98 eV and -1.41 eV. The changes in P and W(D) near a vacancy are calculated; W(D) changes for the individual charges because the vacancy removes a dipole moment and modifies the crystal dielectric response, but deltaW(D) and E(C) do not change. A vacancy yields a positive change DeltaP that scatters a charge or CT pair, but the change DeltaW(D) can be negative and large enough to outweigh DeltaP, yielding traps with depths that can exceed 150 meV for single charges and for CT pairs. Divacancies yield traps with depths nearly equal to the sum of those produced by the separate vacancies and so they can exceed 300 meV. These results are consistent with a mechanism of optical damage in which vacancies trap optically generated CT pairs that recombine and release energy; this can disrupt the lattice around the vacancy, thereby favoring trapping and recombination of CT pairs generated by subsequent photon absorption, leading to further lattice disruption. Revisions to previous calculations on trapping of CT pairs in anthracene are reported. PMID- 15267614 TI - Sum frequency vibrational spectroscopy of leucine molecules adsorbed at air-water interface. AB - Sum frequency vibrational spectroscopy was used to study adsorption of leucine molecules at air-water interface from solutions with different concentrations and pH values. The surface density and the orientation of the isopropyl head group of the adsorbed leucine molecules could be deduced from the measurements. It was found that the orientation depends on the surface density, but only weakly on bulk pH value at the saturated surface density. The vibrational spectra of the interfacial water molecules appeared to be strongly affected by the charge state of the adsorbed leucine molecules. Enhancement and inversion of polar orientation of interfacial water molecules by surface charges or field controllable by the bulk pH value were observed. PMID- 15267615 TI - Kinetics of the CO oxidation reaction on Pt(111) studied by in situ high resolution x-ray photoelectron spectroscopy. AB - High-resolution x-ray photoelectron spectroscopy has been used to study the kinetics of the CO oxidation reaction on a Pt(111) surface in situ. The study focuses on the interaction of a preadsorbed p(2x2) layer of atomic oxygen with CO dosed using a supersonic molecular beam. Measurements of O 1s and C 1s spectra at 120 K show that CO adsorbs on the oxygen precovered substrate, but no reaction occurs. A maximum CO coverage of 0.23 ML (monolayer) is observed, with CO exclusively bound on on-top sites. In accordance with the literature, bridge sites are blocked by the presence of atomic oxygen. The reaction of CO with preadsorbed O to CO(2) is studied isothermally in a temperature range between 275 and 305 K. The reaction rate initially increases with CO pressure, but saturates at 9x10(-7) mbar. The data indicate that a certain amount of disordered oxygen within the p(2x2) layer acts as a starting point of the reaction and for a given temperature reacts with a higher rate than O in the well-ordered oxygen p(2x2) phase. For the reaction of CO with this ordered phase, the results confirm the assumption of a reaction mechanism, which is restricted to the edges of compact oxygen islands. The activation energy of the reaction is determined to (0.53+/ 0.04) eV, with a prefactor of 4.7x10(6+/-0.7) s(-1). PMID- 15267616 TI - Electrofreezing of confined water. AB - We report results from molecular dynamics simulations of the freezing transition of TIP5P water molecules confined between two parallel plates under the influence of a homogeneous external electric field, with magnitude of 5 V/nm, along the lateral direction. For water confined to a thickness of a trilayer we find two different phases of ice at a temperature of T=280 K. The transformation between the two, proton-ordered, ice phases is found to be a strong first-order transition. The low-density ice phase is built from hexagonal rings parallel to the confining walls and corresponds to the structure of cubic ice. The high density ice phase has an in-plane rhombic symmetry of the oxygen atoms and larger distortion of hydrogen bond angles. The short-range order of the two ice phases is the same as the local structure of the two bilayer phases of liquid water found recently in the absence of an electric field [J. Chem. Phys. 119, 1694 (2003)]. These high- and low-density phases of water differ in local ordering at the level of the second shell of nearest neighbors. The results reported in this paper, show a close similarity between the local structure of the liquid phase and the short-range order of the corresponding solid phase. This similarity might be enhanced in water due to the deep attractive well characterizing hydrogen bond interactions. We also investigate the low-density ice phase confined to a thickness of 4, 5, and 8 molecular layers under the influence of an electric field at T=300 K. In general, we find that the degree of ordering decreases as the distance between the two confining walls increases. PMID- 15267617 TI - Nanometer-size cluster formation in alkali-metal-doped fullerene layers. AB - Kinetic Monte Carlo methods have been used to simulate structural transformations in fullerene layers during electrochemical intercalation with alkali-metal ions (A). Special attention is paid to the thermodynamic stability of the A(x)C(60) phases. The calculations point out a phase separation in the doped fullerene layer into alkali-metal-rich and alkali-metal-depleted areas at room temperature. The final state is represented by two phases which coexist as a stable fine mixture of nanoscale particles. The instability of homogeneous layers has potentially critical impact on their electrical properties and can explain the formation of nanostructures (20-50 nm) at the fullerene-electrolyte interface. Rb(3)C(60) clusters are predicted to be larger than K(3)C(60) ones for equal mean alkali-metal concentrations. Experimental data on electrochemical metal deposition on alkali-metal-doped fullerene substrates-in particular, atomic force microscopy measurements-are also consistent with the model proposed. PMID- 15267618 TI - Reflectivity and anisotropic optical functions of quaterthiophene single crystals. AB - Polarized reflectance spectra of quaterthiophene single crystals are reported with different angles and planes of incidence. The strong dependence of the spectral features on the experimental configuration is described by an orthorhombic model and the components of the complex diagonal dielectric tensor are given. PMID- 15267619 TI - Gold particle interaction in regular arrays probed by surface enhanced Raman scattering. AB - Lithographically designed two-dimensional arrays consisting of gold nanoparticles deposited on a smooth gold film are used as substrate to examine the SERS effect of the trans-1,2-bis (4-pyridyl) ethylene molecule. These arrays display two plasmon bands instead of the single one observed for the same arrays of particles but deposited on indium tin oxide coated glass. Laser excitation within the short wavelength band does not bring about any SERS spectrum, while excitation within the long wavelength band yields SERS spectra with a gain per molecule rising up to 10(8). The simultaneous investigation of extinction and Raman spectra of arrays exhibiting various topography parameters enables us to suggest an interpretation for both the occurrence of the two plasmon resonances and for the high Raman enhancement. We suggest to assign the short wavelength band to a plasmon wave propagating at the gold glass interface and the long wavelength one to an air/gold surface plasmon mode modified by particle-particle interaction. PMID- 15267620 TI - Ozone adsorption on carbon nanotubes: the role of Stone-Wales defects. AB - First-principles calculations within the density functional theory have been performed in order to investigate ozone adsorption on carbon nanotubes. Particular emphasis is placed on the effects of Stone-Wales-like defects on the structural and electronic properties of (i) ideal tubes and (ii) tubes in the presence of ozone. Our results show that structural deformations induced on the pure carbon nanotubes by Stone-Wales defects are similar, as expected, to those induced on graphite; for the (10,0) tube, the semiconducting character is kept, though with a small reduction of the band gap. As for the ozone adsorption, the process on ideal nanotubes is most likely physisorption, though slightly stronger if compared to other previously studied molecules and consistent with the strong oxydizing nature of O(3). However, when ozone adsorbs on Stone-Wales defects, a strong chemisorption occurs, leading to relevant structural relaxations and to the formation of a CO covalent bond; this is consistent with experimental observations of CO functional groups, as well as of the liberation of CO gas phase and of the formation of C vacancies, thus explaining the consumption of the nanotube film upon ozone exposure. PMID- 15267621 TI - Infrared spectroscopy of the solid phases of ammonia. AB - Thin films of solid ammonia (NH(3) and ND(3)) have been characterized using low temperature (25-110 K) Fourier-transform infrared (FTIR) spectroscopy, and the three solid phase (amorphous, metastable, and crystalline) spectra are reported. This work has been motivated by confusion in the literature about the metastable and crystalline phases as a result of an early erroneous report by Staats and Morgan [(J. Chem. Phys. 31, 553 (1959)]. Although the crystalline phase has subsequently been reported correctly, the metastable phase has not been described in the literature in detail. The unique characteristics of the metastable phase, reported here for the first time, include multiple peaks in the nu(2) and nu(3) regions and peak intensities that are dependent on the deposition temperature. This behavior may be the result of (a) preferential molecular orientations in the solid, or (b) exciton splitting due to different crystal shapes in the solid. The amorphous and metastable phases of deuterated ammonia are also reported for the first time. PMID- 15267622 TI - Broadband femtosecond sum-frequency spectroscopy of CO on Ru1010 in the frequency and time domains. AB - CO on Ru[1010] was investigated by broadband femtosecond sum-frequency spectroscopy at 200 K. Approximately half of the frequency shift of 71 cm(-1) over the coverage range from 0.15 to 1.22 monolayers is shown to originate from dipole-dipole coupling, with the remainder due to a chemical shift. Despite low adlayer-surface registration at the highest coverages, the linewidth of the C-O stretch is comparatively low, and is described by homogeneous broadening according to sum-frequency free-induction decay measurements in the time domain. This can be explained by the dominance of the CO dipole coupling strength over the static disorder present in a coincidence structure. As the coverage decreases below 0.3 monolayer, the linewidth increases considerably, indicative of inhomogeneous broadening. Supported by a concomitant frequency change we suggest that at low coverages CO molecules form chains of irregular length in the [0001] direction, as has been shown for other surfaces with similar symmetry. PMID- 15267623 TI - Lagrange multiplier based transport theory for quantum wires. AB - We discuss how the Lagrange multiplier method of nonequilibrium steady state statistical mechanics can be applied to describe the electronic transport in a quantum wire. We describe the theoretical scheme using a tight-binding model. The Hamiltonian of the wire is extended via a Lagrange multiplier to "open" the quantum system and to drive current through it. The diagonalization of the extended Hamiltonian yields the transport properties of wire. We show that the Lagrange multiplier method is equivalent to the Landauer approach within the considered model. PMID- 15267624 TI - Interaction of atomic hydrogen with single-walled carbon nanotubes: a density functional theory study. AB - We have studied the interaction of atomic hydrogen with (5,5) and (10,0) single walled carbon nanotubes (SWNT) using density functional theory. These calculations use Gaussian orbitals and periodic boundary conditions. We compare results from the local spin density approximation, generalized gradient approximation (GGA), and hybrid density functionals. We have first kept the SWNT geometric structure fixed while a single H atom approaches the tube on top of a carbon atom. In that case, a weakly bound state with binding energies from -0.8 to -0.4 eV was found. Full geometry relaxation leads to a strong SWNT deformation, weakening the nearest C-C bonds and increasing the binding energy by about 1 eV. Full hydrogen coverage of the (5,5) SWNT converts this metallic nanotube into an insulator with a band gap of 3.4 eV for the GGA functional and 4.8 eV for the hybrid functional. Hybrid functionals perform similar to pure density functional theory functionals for the calculation of binding energies while band gaps critically depend on the functional choice. PMID- 15267625 TI - Analytic density-functional self-consistent-field theory of diblock copolymers near patterned surfaces. AB - Analytical solutions are derived for the density profiles and the free energies of compressible diblock copolymer melts (or incompressible copolymer solutions) near patterned surfaces. The density-functional self-consistent-field theory is employed along with a Gaussian chain model for bonding constraints and a random mixing approximation for nonbonded interactions. An analytical solution is rendered possible by expanding the chain distribution function around an inhomogeneous reference state with a nontrivial analytical solution, by retaining the linear terms, and by requiring consistency with the homopolymer limit. The density profiles are determined by both real and complex roots of a sixth-degree polynomial that may easily be obtained by solving a generalized eigenvalue problem. This analytical formulation enables one to efficiently explore the large nine-dimensional parameter space and can serve as a first approximation to computationally intensive studies with more detailed models. Illustrative computations are provided for uniform and patterned surfaces above the order disorder transition. The results are consistent with the previous self-consistent field calculations in that lamellar ordering appears near the surface above the order-disorder transition and the lamella order perpendicular or parallel to the surface depending on the commensurability between the periods of the surface pattern and the density oscillations. PMID- 15267627 TI - Localization and freezing of a Gaussian chain in a quenched random potential. AB - The Gaussian chain in a quenched random potential (which is characterized by the disorder strength Delta) is investigated in the d-dimensional space by the replicated variational method. The general expression for the free energy within so-called one-step-replica symmetry breaking (1-RSB) scenario has been systematically derived. We have shown that the replica symmetrical (RS) limit of this expression can describe the chain center-of-mass localization and collapse. The critical disorder when the chain becomes localized scales as Delta(c) approximately b(d)N(-2+d/2) (where b is the length of the Kuhn segment length and N is the chain length) whereas the chain gyration radius R(g) approximately b(b(d)/Delta)(1/(4-d)). The freezing of the internal degrees of freedom follows to the 1-RSB-scenario and is characterized by the beads localization length D(2). It was demonstrated that the solution for D(2) appears as a metastable state at Delta=Delta(A) and behaves similarly to the corresponding frozen states in heteropolymers or in p-spin random spherical model. PMID- 15267626 TI - Membrane inclusions as coupled harmonic oscillators: effects due to anisotropic membrane slope relaxation. AB - Membrane-mediated interaction between membrane-spanning peptides or protein segments plays an important role in their function and stability. Our rigorous "coupled harmonic oscillators" representation is extended to account for the complex boundary conditions permitting anisotropic relaxation of the membrane slope along the contours of the inclusions. Using this representation and applying a highly efficient finite-difference algorithm, we have analyzed the membrane-mediated interaction triggered by deformation of the hydrophobic tails of lipid molecules to match the lipophilic exterior of the inserted peptide. We establish that anisotropic relaxation crucially affects the interaction energy, leading to a short-range attraction between two inclusions, while conventional isotropic boundary conditions result in their strong repulsion. In a multi inclusion cluster, this attraction is further enhanced and modified due to nonpairwise interactions. The results for dimyristoyl phosphatidylcholine and glyceryl monooleate membranes are compared, and the effects of the inclusion radius are considered. The possible role of slope relaxation in the reported stabilization of linked gramicidin channels and in proteins' functional cooperativity is outlined. PMID- 15267628 TI - Properties of branched confined polymers. AB - A model of star-branched polymer chains confined in a slit formed by two parallel surfaces was studied. The chains were embedded to a simple cubic lattice and consisted of f=3 branches of equal length. The macromolecules had the excluded volume and the confining surfaces were impenetrable for polymer segments. No attractive interactions between polymer segments and then between polymer segments and the surfaces were assumed and therefore the system was a thermal. Monte Carlo simulations were carried out employing the sampling algorithm based on chain's local changes of conformation. Lateral diffusion of star-branched chains was studied. Dynamic properties of star-branched chains between the walls with impenetrable rod-like obstacles were also studied and compared to the previous case. The density profiles of polymer segments on the slit were determined. The analysis of contacts between the polymer chain and the surfaces was also carried out. PMID- 15267629 TI - Microscopic theory of gelation and elasticity in polymer-particle suspensions. AB - A simplified mode-coupling theory (MCT) of ergodic-nonergodic transitions, in conjunction with an accurate two-component polymer reference interaction site model (PRISM) theory for equilibrium structural correlations, has been systematically applied to investigate gelation, localization, and elasticity of flexible polymer-hard particle suspensions. The particle volume fraction at the fluid-gel transition is predicted to depend exponentially on reduced polymer concentration and size asymmetry ratio at relatively high colloid concentrations. In contrast, at lower particle volume fractions, a power-law dependence on polymer concentration is found with effective exponents and prefactors that depend systematically on the polymer/particle size ratio. Remarkable power-law and near universal scaling behavior is found for the localization length and elastic shear modulus. Multiple experiments for gel boundaries and shear moduli are in good agreement with the no adjustable parameter theory. The one exception is the absolute magnitude of the shear modulus which is strongly overpredicted, apparently due to nonequilibrium dense cluster formation. The simplified MCT PRISM theory also captures the qualitative aspects of the weak depletion-driven "glass melting" phenomenon at high particle volume fractions. Calculations based on an effective one-component model of structure within a low particle volume fraction framework yield qualitatively different features than the two-component approach and are apparently all in disagreement with experiments. This suggests that volume fraction and size asymmetry dependent many-body screening of polymer mediated depletion attractions at finite particle concentrations are important. PMID- 15267630 TI - Density-functional theory of spherical electric double layers and zeta potentials of colloidal particles in restricted-primitive-model electrolyte solutions. AB - A density-functional theory is proposed to describe the density profiles of small ions around an isolated colloidal particle in the framework of the restricted primitive model where the small ions have uniform size and the solvent is represented by a dielectric continuum. The excess Helmholtz energy functional is derived from a modified fundamental measure theory for the hard-sphere repulsion and a quadratic functional Taylor expansion for the electrostatic interactions. The theoretical predictions are in good agreement with the results from Monte Carlo simulations and from previous investigations using integral-equation theory for the ionic density profiles and the zeta potentials of spherical particles at a variety of solution conditions. Like the integral-equation approaches, the density-functional theory is able to capture the oscillatory density profiles of small ions and the charge inversion (overcharging) phenomena for particles with elevated charge density. In particular, our density-functional theory predicts the formation of a second counterion layer near the surface of highly charged spherical particle. Conversely, the nonlinear Poisson-Boltzmann theory and its variations are unable to represent the oscillatory behavior of small ion distributions and charge inversion. Finally, our density-functional theory predicts charge inversion even in a 1:1 electrolyte solution as long as the salt concentration is sufficiently high. PMID- 15267632 TI - Collision complexes in dissociative single electron transfer between Ne2+ and N2. AB - Experiments involving the coincident detection of the two monocationic products (Ne+ and N+) from the dissociative electron transfer reaction between Ne2+ and N2 at 7.8 eV collision energy allow the nascent velocity vectors of the ionic and neutral (N) products to be determined. Examination of the correlations between these vectors shows that one pathway to the products involves the dissociation of a transitory collision complex (N2Ne2+). PMID- 15267633 TI - Inelastic tunneling spectroscopy using scanning tunneling microscopy on trans-2 butene molecule: spectroscopy and mapping of vibrational feature. AB - Inelastic tunneling spectroscopy (IETS) measurement using scanning tunneling microscopy (STM) with a commercially available STM set up is presented. The STM IETS spectrum measured on an isolated trans-2-butene molecule on the Pd(110) shows a clear vibrational feature in d2I/dV2 at the bias voltage of 360 mV and 363 mV, which corresponds to the nu(C-H) mode (d2I/dV2 approximately 10 nA/V2). In addition, we have obtained an image by mapping the vibrational feature of nu(C H) in d2I/dV2. The image is obtained by scanning the tip on the surface with the feedback loop activated while the modulation voltage is superimposed on the sample voltage. With the method that is readily performable with conventional software, we have clearly differentiated the molecules of trans-2-butene and butadiene through the mapping of the vibrational feature, demonstrating its capability of chemical identification in atomic scale. PMID- 15267634 TI - Density-functional generalized-gradient and hybrid calculations of electromagnetic properties using Slater basis sets. AB - In this paper we extend our density-functional theory calculations, with generalized gradient approximation and hybrid functionals, using Slater-type orbitals (STOs), to the determination of second-order molecular properties. The key to the entire methodology involves the fitting of all STO basis function products to an auxiliary STO basis, through the minimization of electron repulsion integrals. The selected properties are (i) dipole polarizabilities, (ii) nuclear magnetic shielding constants, and (iii) nuclear spin-spin coupling constants. In all cases the one-electron integrals involving STOs were evaluated by quadrature. The implementation for (ii) involved some complexity because we used gauge-including atomic orbitals. The presence of two-electron integrals on the right-hand side of the coupled equations meant that the fitting procedure had to be implemented. For (iii) in the hybrid case, fitting procedures were again required for the exchange contributions. For each property we studied a number of small molecules. We first obtained an estimate of the basis set limit using Gaussian-type orbitals (GTOs). We then showed how it is possible to reproduce these values using a STO basis set. For (ii) a regular TZ2P quality STO basis was adequate; for (i) the addition of one set of diffuse functions (determined by Slater's rules) gave the required accuracy; for (iii) it was necessary to add a set of 1s functions, including one very tight function, to give the desired result. In summary, we show that it is possible to predict second-order molecular properties using STO basis sets with an accuracy comparable with large GTO basis sets. We did not encounter any major difficulties with either the selection of the bases or the implementation of the procedures. Although the energy code (especially in the hybrid case) may not be competitive with a regular GTO code, for properties we find that STOs are more attractive. PMID- 15267635 TI - An empirical charge transfer potential with correct dissociation limits. AB - The empirical valence bond (EVB) method [J. Chem. Phys. 52, 1262 (1970)] has always embodied charge transfer processes. The mechanism of that behavior is examined here and recast for use as a new empirical potential energy surface for large-scale simulations. A two-state model is explored. The main features of the model are: (1) explicit decomposition of the total system electron density is invoked; (2) the charge is defined through the density decomposition into constituent contributions; (3) the charge transfer behavior is controlled through the resonance energy matrix elements which cannot be ignored; and (4) a reference state approach, similar in spirit to the EVB method, is used to define the resonance state energy contributions in terms of "knowable" quantities. With equal validity, the new potential energy can be expressed as a nonthermal ensemble average with a nonlinear but analytical charge dependence in the occupation number. Dissociation to neutral species for a gas-phase process is preserved. A variant of constrained search density functional theory is advocated as the preferred way to define an energy for a given charge. PMID- 15267636 TI - Assessment and validation of a screened Coulomb hybrid density functional. AB - This paper presents a revised and improved version of the Heyd-Scuseria-Ernzerhof screened Coulomb hybrid functional. The performance of this functional is assessed on a variety of molecules for the prediction of enthalpies of formation, geometries, and vibrational frequencies, yielding results as good as or better than the successful PBE0 hybrid functional. Results for ionization potentials and electron affinities are of slightly lower quality but are still acceptable. The comprehensive test results presented here validate our assumption that the screened, short-range Hartree-Fock (HF) exchange exhibits all physically relevant properties of the full HF exchange. Thus, hybrids can be constructed which neglect the computationally demanding long-range part of HF exchange while still retaining the superior accuracy of hybrid functionals, compared to pure density functionals. PMID- 15267637 TI - Dynamically weighted multiconfiguration self-consistent field: multistate calculations for F+H2O-->HF+OH reaction paths. AB - A novel method of dynamically adjusted weighting factors in state-averaged multiconfiguration self-consistent-field calculations (SA-MCSCF) is described that is applicable to systems of arbitrary dimensionality. The proposed dynamically weighted approach automatically weights the relevant electronic states in each region of the potential energy surface, smoothly adjusting between these regions with an energy dependent functional. This method is tested on the F(2P)+H2O-->HF+OH(2Pi) reaction, which otherwise proves challenging to describe with traditional SA-MCSCF methods due to (i) different asymptotic degeneracies of reactant (threefold) and product (twofold) channels, and (ii) presence of low lying charge transfer configurations near the transition state region. The smoothly varying wave functions obtained by dynamically weighted multiconfigurational self-consistent field represent excellent reference states for high-level multireference configuration interaction calculations and offer an ideal starting point for construction of multiple state potential energy surfaces. PMID- 15267638 TI - Atomic and molecular intracules for excited states. AB - Intracules in position space, momentum space and phase space have been calculated for low-lying excited states of the He atom, Be atom, formaldehyde and butadiene. The phase-space intracules (Wigner intracules) provide significantly more information than the position- and momentum-space intracules, particularly for the Be atom. Exchange effects are investigated through the differences between corresponding singlet and triplet states. PMID- 15267639 TI - Real versus artifactual symmetry-breaking effects in Hartree-Fock, density functional, and coupled-cluster methods. AB - We have examined the relative abilities of Hartree-Fock, density-functional theory (DFT), and coupled-cluster theory in describing second-order (pseudo) Jahn Teller (SOJT) effects, perhaps the most commonly encountered form of symmetry breaking in polyatomic molecules. As test cases, we have considered two prototypical systems: the 2Sigmau+ states of D( infinity h) BNB and C3+ for which interaction with a low-lying 2Sigmag+ excited state leads to symmetry breaking of the nuclear framework. We find that the Hartree-Fock and B3LYP methods correctly reproduce the pole structure of quadratic force constants expected from exact SOJT theory, but that both methods appear to underestimate the strength of the coupling between the electronic states. Although the Tamm-Dancoff (CIS) approximation gives excitation energies with no relationship to the SOJT interaction, the random-phase-approximation (RPA) approach to Hartree-Fock and time-dependent DFT excitation energies predicts state crossings coinciding nearly perfectly with the positions of the force constant poles. On the other hand, the RPA excited-state energies exhibit unphysical curvature near their crossings with the ground (reference) state, a problem arising directly from the mathematical structure of the RPA equations. Coupled-cluster methods appear to accurately predict the strength of the SOJT interactions between the 2Sigmau+ and 2Sigmag+ states, assuming that the inclusion of full triple excitations provides a suitable approximation to the exact wave function, and are the only methods examined here which predict symmetry breaking in BNB. However, coupled-cluster methods are plagued by artifactual force constant poles arising from the response of the underlying reference molecular orbitals to the geometric perturbation. Furthermore, the structure of the "true" SOJT force constant poles predicted by coupled-cluster methods, although correctly positioned, has the wrong structure. PMID- 15267640 TI - Relation between different variants of the generalized Douglas-Kroll transformation through sixth order. AB - Wolf et al. have recently investigated a generalized Douglas-Kroll transformation. From a general class of unitary transformations that can be used in the Douglas-Kroll transformation, they pick one which is supposed to give, at a given order, an optimal transformed Dirac Hamiltonian. Results were presented through the fifth order. However, no data were given to demonstrate to which extent the so-called "optimal" Douglas-Kroll transformation is superior to other choices. In this work, the Douglas-Kroll transformation is extended to the sixth order for the first time, using computer algebra algorithms to obtain the working equations. It is shown how, at a given order, different variants of the Douglas Kroll Hamiltonians are related. Various choices of the generalized transformation are examined numerically for the ground states of the one-electron atomic ions with nuclear charges Z=20, 40, 60, 80, 100, and 120. It is shown that compared to the improvement obtained by including the next order, the differences between various choices for the generalized Douglas-Kroll transformation are almost negligible. Results closest to the Dirac eigenvalues are not obtained with the optimal Douglas-Kroll transformation given by Wolf et al., but with the parametrization originally suggested by Douglas and Kroll. PMID- 15267641 TI - Mixing and reaction fronts in laminar flows. AB - Autocatalytic reaction fronts between unreacted and reacted mixtures in the absence of fluid flow propagate as solitary waves. In the presence of imposed flow, the interplay between diffusion and advection enhances the mixing, leading to Taylor hydrodynamic dispersion. We present asymptotic theories in the two limits of small and large Thiele modulus (slow and fast reaction kinetics, respectively) that incorporate flow, diffusion, and reaction. For the first case, we show that the problem can be handled to leading order by the introduction of the Taylor dispersion replacing the molecular diffusion coefficient by its Taylor counterpart. In the second case, the leading-order behavior satisfies the eikonal equation. Numerical simulations using a lattice gas model show good agreement with the theory. The Taylor model is relevant to microfluidics applications, whereas the eikonal model applies at larger length scales. PMID- 15267642 TI - Analytic evaluation of nonadiabatic coupling terms at the MR-CI level. I. Formalism. AB - An efficient and general method for the analytic computation of the nonandiabatic coupling vector at the multireference configuration interaction (MR-CI) level is presented. This method is based on a previously developed formalism for analytic MR-CI gradients adapted to the use for the computation of nonadiabatic coupling terms. As was the case for the analytic energy gradients, very general, separate choices of invariant orbital subspaces at the multiconfiguration self-consistent field and MR-CI levels are possible, allowing flexible selections of MR-CI wave functions. The computational cost for the calculation of the nonadiabatic coupling vector at the MR-CI level is far below the cost for the energy calculation. In this paper the formalism of the method is presented and in the following paper [Dallos et al., J. Chem. Phys. 120, 7330 (2004)] applications concerning the optimization of minima on the crossing seam are described. PMID- 15267643 TI - Analytic evaluation of nonadiabatic coupling terms at the MR-CI level. II. Minima on the crossing seam: formaldehyde and the photodimerization of ethylene. AB - The method for the analytic calculation of the nonadiabatic coupling vector at the multireference configuration-interaction (MR-CI) level and its program implementation into the COLUMBUS program system described in the preceding paper [Lischka et al., J. Chem. Phys. 120, 7322 (2004)] has been combined with automatic searches for minima on the crossing seam (MXS). Based on a perturbative description of the vicinity of a conical intersection, a Lagrange formalism for the determination of MXS has been derived. Geometry optimization by direct inversion in the iterative subspace extrapolation is used to improve the convergence properties of the corresponding Newton-Raphson procedure. Three examples have been investigated: the crossing between the 1(1)B1/2(1)A1 valence states in formaldehyde, the crossing between the 2(1)A1/3(1)A1 pi-pi* valence and ny-3py Rydberg states in formaldehyde, and three crossings in the case of the photodimerization of ethylene. The methods developed allow MXS searches of significantly larger systems at the MR-CI level than have been possible before and significantly more accurate calculations as compared to previous complete active space self-consistent field approaches. PMID- 15267644 TI - Irreducible Brillouin conditions and contracted Schrodinger equations for n electron systems. III. Systems of noninteracting electrons. AB - We analyze the structure and the solutions of the irreducible k-particle Brillouin conditions (IBCk) and the irreducible contracted Schrodinger equations (ICSEk) for an n-electron system without electron interaction. This exercise is very instructive in that it gives one both the perspective and the strategies to be followed in applying the IBC and ICSE to physically realistic systems with electron interaction. The IBC1 leads to a Liouville equation for the one-particle density matrix gamma1=gamma, consistent with our earlier analysis that the IBC1 holds both for a pure and an ensemble state. The IBC1 or the ICSE1 must be solved subject to the constraints imposed by the n-representability condition, which is particularly simple for gamma. For a closed-shell state gamma is idempotent, i.e., all natural spin orbitals (NSO's) have occupation numbers 0 or 1, and all cumulants lambdak with k> or =2 vanish. For open-shell states there are NSO's with fractional occupation number, and at the same time nonvanishing elements of lambda2, which are related to spin and symmetry coupling. It is often useful to describe an open-shell state by a totally symmetric ensemble state. If one wants to treat a one-particle perturbation by means of perturbation theory, this mainly as a run-up for the study of a two-particle perturbation, one is faced with the problem that the perturbation expansion of the Liouville equation gives information only on the nondiagonal elements (in a basis of the unperturbed states) of gamma. There are essentially three possibilities to construct the diagonal elements of gamma: (i) to consider the perturbation expansion of the characteristic polynomial of gamma, especially the idempotency for closed-shell states, (ii) to rely on the ICSE1, which (at variance with the IBC1) also gives information on the diagonal elements, though not in a very efficient manner, and (iii) to formulate the perturbation theory in terms of a unitary transformation in Fock space. The latter is particularly powerful, especially, when one wishes to study realistic Hamiltonians with a two-body interaction. PMID- 15267645 TI - Irreducible Brillouin conditions and contracted Schrodinger equations for n electron systems. IV. Perturbative analysis. AB - The k-particle irreducible Brillouin conditions IBCk and the k-particle irreducible contracted Schrodinger equations ICSEk for a closed-shell state are analyzed in terms of a Moller-Plesset-type perturbation expansion. The zeroth order is Hartree-Fock. From the IBC2(1), i.e., from the two-particle IBC to first order in the perturbation parameter mu, one gets the leading correction lambda2(1) to the two-particle cumulant lambda2 correctly. However, in order to construct the second-order energy E(2), one also needs the second-order diagonal correction gammaD(2) to the one-particle density matrix gamma. This can be obtained: (i) from the idempotency of the n-particle density matrix, i.e., essentially from the requirement of n-representability; (ii) from the ICSE1(2); or (iii) by means of perturbation theory via a unitary transformation in Fock space. Method (ii) is very unsatisfactory, because one must first solve the ICSE3(2) to get lambda3(2), which is needed in the ICSE2(2) to get lambda2(2), which, in turn, is needed in the ICSE1(2) to get gamma(2). Generally the (k+1) particle approximation is needed to obtain Ek correctly. One gains something, if one replaces the standard hierarchy, in which one solves the ICSEk, ignoring lambda(k+1) and lambda(k+2), by a renormalized hierarchy, in which only lambda(k+2) is ignored, and lambda(k+1) is expressed in terms of the lambdap of lower particle rank via the partial trace relation for lambda(k+2). Then the k particle approximation is needed to obtain E(k) correctly. This is still poorer than coupled-cluster theory, where the k-particle approximation yields E(k+1). We also study the possibility to use some simple necessary n-representability conditions, based on the non-negativity of gamma(2) and two related matrices, in order to get estimates for gammaD(2) in terms of lambda2(1). In general these estimates are rather weak, but they can become close to the best possible bounds in special situations characterized by a very sparse structure of lambda2 in terms of a localized representation. The perturbative analysis does not encourage the use of a k-particle hierarchy based on the ICSEk (or on their reducible counterparts, the CSEk), it rather favors the approach in terms of the unitary transformation, where the k-particle approximation yields the energy correct up to E(2k-1). The problems that arise are related to the unavoidable appearance of exclusion-principle violating cumulants. The good experience with perturbation theory in terms of a unitary transformation suggests that one should abandon a linearly convergent iteration scheme based on the ICSEk hierarchy, in favor of a quadratically convergent one based on successive unitary transformations. PMID- 15267646 TI - Structural properties of reciprocal form factor in neutral atoms and singly charged ions. AB - Structural characteristics of the spherically averaged internally folded density or reciprocal form factor Br are studied within the Hartree-Fock framework for 103 neutral atoms, 54 singly charged cations, and 43 anions in their ground state. The function Br is classified throughout the Periodic Table into three types: (i) monotonic decrease from the origin, (ii) maximum at r=0 and a negative minimum at r>0, and (iii) a local maximum at r=0 and a pair maximum-minimum out of the origin. A detailed study of the corresponding properties for individual subshells as well as their relative weight for the total Br is also carried out. For completeness, the analytical Br for hydrogenlike atoms in both ground and excited states is also analyzed. PMID- 15267647 TI - A local approach to delocalized electronic systems: semilocal evaluation of the cohesive energies of tight-binding Hamiltonians. AB - Starting from strongly localized N-electron functions built from either pure atomic orbitals or fully localized bond molecular orbitals, it is possible to evaluate the ground state energy of a periodic lattice ruled by a tight-binding Hamiltonian without explicitly introducing the monoelectronic crystal orbitals. The method consists of a self-consistent perturbation of the zeroth-order wave function which incorporates high order effects and offers reasonable convergence properties. Along this framework, a single variable per bond type is introduced, namely the amplitude of the charge transfer. The method leads to a set of coupled equations which can be numerically solved, if not analytically. Short-range delocalization effects under periodic conditions are explicitly taken into account and relatively accurate cohesive energies are estimated for regular homoatomic and heteroatomic one-dimensional chains as well as for honeycomb lattices. In addition, good agreement with experiment for the distortion amplitude in polyacetylene is obtained. This exploratory tool may be easily extended to more sophisticated Hamiltonians, for which the solutions are not accessible. Since our approach only introduces short-range delocalization effects, its performance questions the importance of the specifically collective delocalization effects. PMID- 15267648 TI - Globally uniform semiclassical surface-hopping wave function for nonadiabatic scattering. AB - A globally uniform time-independent semiclassical wave function for nonadiabatic scattering is presented. This wave function, which takes the form of a surface hopping expansion, is motivated by the globally uniform semiclassical wave function of Kay and co-workers for the single-surface case. The surface-hopping expansion is similar to a previously presented primitive semiclassical wave function for nonadiabatic problems. This earlier wave function has the important feature that it correctly incorporates all phase terms, allowing for an accurate treatment of quantum interference effects. The globally uniform expression has important numerical advantages over the primitive formulation. The globally uniform wave function does not have caustic singularities, and the globally uniform calculation avoids a root search for trajectories obeying double-ended boundary conditions that is required by the primitive semiclassical calculation. PMID- 15267649 TI - Dissociative recombination of NH4+ and ND4+ ions: storage ring experiments and ab initio molecular dynamics. AB - The dissociative recombination (DR) process of NH4+ and ND4+ molecular ions with free electrons has been studied at the heavy-ion storage ring CRYRING (Manne Siegbahn Laboratory, Stockholm University). The absolute cross sections for DR of NH4+ and ND4+ in the collision energy range 0.001-1 eV are reported, and thermal rate coefficients for the temperature interval from 10 to 2000 K are calculated from the experimental data. The absolute cross section for NH4+ agrees well with earlier work and is about a factor of 2 larger than the cross section for ND4+. The dissociative recombination of NH4+ is dominated by the product channels NH3+H (0.85+/-0.04) and NH2+2H (0.13+/-0.01), while the DR of ND4+ mainly results in ND3+D (0.94+/-0.03). Ab initio direct dynamics simulations, based on the assumption that the dissociation dynamics is governed by the neutral ground-state potential energy surface, suggest that the primary product formed in the DR process is NH3+H. The ejection of the H atom is direct and leaves the NH3 molecule highly vibrationally excited. A fraction of the excited ammonia molecules may subsequently undergo secondary fragmentation forming NH2+H. It is concluded that the model results are consistent with gross features of the experimental results, including the sensitivity of the branching ratio for the three-body channel NH2+2H to isotopic exchange. PMID- 15267650 TI - Picosecond IR-UV pump-probe spectroscopic study of the dynamics of the vibrational relaxation of jet-cooled phenol. I. Intramolecular vibrational energy redistribution of the OH and CH stretching vibrations of bare phenol. AB - The intramolecular vibrational energy redistribution (IVR) of the OH stretching vibration of jet-cooled phenol-h6 (C6H8OH) and phenol-d8 (C6D8OH) in the electronic ground state has been investigated by picosecond time-resolved IR-UV pump-probe spectroscopy. The OH stretching vibration of phenol was excited with a picosecond IR laser pulse, and the subsequent temporal evolutions of the initially excited level and the redistributed ones due to the IVR were observed by multiphoton ionization detection with a picosecond UV pulse. The IVR lifetime for the OH stretch vibration of phenol-h6 was determined to be 14 ps, while that of the OH stretch for phenol-d8 was found to be 80 ps. This remarkable change of the IVR rate constant upon the dueteration of the CH groups strongly suggests that the "doorway states" for the IVR from the OH level would be the vibrational states involving the CH stretching modes. We also investigated the IVR rate of the CH stretching vibration for phenol-h6. It was found that the IVR lifetime of the CH stretch is less than 5 ps. The fast IVR is described by the strong anharmonic resonance of the CH stretch with many other combinations or overtone bands. PMID- 15267651 TI - Picosecond IR-UV pump-probe spectroscopic study of the dynamics of the vibrational relaxation of jet-cooled phenol. II. Intracluster vibrational energy redistribution of the OH stretching vibration of hydrogen-bonded clusters. AB - A picosecond time-resolved IR-UV pump-probe spectroscopic study has been carried out for investigating the intracluster vibrational energy redistribution (IVR) and subsequent dissociation of hydrogen-bonded clusters of phenol (C6H5OH) and partially deuterated phenol (C6D5OH, phenol-d5) with various solvent molecules. The H-bonded OH stretching vibration was pumped by a picosecond IR pulse, and the transient S1-S0 UV spectra from the pumped level as well as the redistributed levels were observed with a picosecond UV laser. Two types of hydrogen-bonded clusters were investigated with respect to the effect of the H-bonding strength on the energy flow process: the first is of a strong "sigma-type H-bond" such as phenol-(dimethyl ether)(n=1) and phenol dimer, and the second is phenol (ethylene)(n=1) having a weak "pi-type H-bond." It was found that the population of the IR-pumped OH level exhibits a single-exponential decay, whose rate increases with the H-bond strength. On the other hand, the transient UV spectrum due to the redistributed levels showed a different time evolutions at different monitoring UV frequency. From an analysis of the time profiles of the transient UV spectra, the following three-step scheme has been proposed for describing the energy flow starting from the IVR of the initially excited H-bonded OH stretching level to the dissociation of the H bond. (1) The intramolecular vibrational energy redistribution takes place within the phenolic site, preparing a hot phenol. (2) The energy flows from the hot phenol to the intermolecular vibrational modes of the cluster. (3) Finally, the hydrogen bond dissociates. Among the three steps, the rate constant of the first step was strongly dependent on the H-bond strength, while the rate constants of the other two steps were almost independent of the H-bond strength. For the dissociation of the hydrogen bond, the observed rate constants were compared with those calculated by the Rice, Ramsperger, Kassel, and Marcus model. The result suggests that dissociation of the hydrogen bond takes place much faster than complete energy randomization within the clusters. PMID- 15267652 TI - Spectroscopic properties and potential energy curves of low-lying electronic states of RuC. AB - The RuC molecule has been a challenging species due to the open-shell nature of Ru resulting in a large number of low-lying electronic states. We have carried out state-of-the-art calculations using the complete active space multiconfiguration self-consistent field followed by multireference configuration interaction methods that included up to 18 million configurations, in conjunction with relativistic effects. We have computed 29 low-lying electronic states of RuC with different spin multiplicities and spatial symmetries with energy separations less than 38,000 cm(-1). We find two very closely low-lying electronic states for RuC, viz., 1Sigma+ and 3Delta with the 1Sigma+ being stabilized at higher levels of theory. Our computed spectroscopic constants and dipole moments are in good agreement with experiment although we have reported more electronic states than those that have been observed experimentally. Our computations reveal a strongly bound 1Sigma+ state with a large dipole moment which is most likely the experimentally observed ground state and an energetically close 3Delta state with a smaller dipole moment. Overall our computed spectroscopic constants of the excited states with energy separations less than 18,000 cm(-1) agree quite well with those of the corresponding observed states. PMID- 15267653 TI - Semiclassical dynamics of the van der Waals states in O3(X1A1). AB - We present the analysis and the semiclassical quantization of the van der Waals states of ozone in the ground electronic state X1A1. Progressions of these states dominate the spectrum of O3 at threshold. Periodic orbits are used to perform assignment and quantization of the vibrational states. Semiclassical quantization is numerically accurate despite the fact that the classical phase space is chaotic while the nodal patterns of the quantum mechanical wave functions are regular. The lifetimes of recombination of the van der Waals states into the "normal" ozone are also discussed. PMID- 15267654 TI - Alignment of molecules in pulsed resonant laser fields. AB - We investigate by numerical simulations the dynamics of alignment of linear molecules in resonant pulsed laser fields and its dependence on pulse length, field strength, and molecular parameters. We propose an analytical short-time approximation for the time-dependent wave packets. We provide a theoretical basis for the occurrence of saturation in the rotational pumping. We present a formula to predict the time at which the maximum alignment occurs. We discuss the magnitude of the laser-induced alignment and we relate it to a theoretical upper limit. PMID- 15267655 TI - Radiation damage of biosystems mediated by secondary electrons: resonant precursors for uracil molecules. AB - Calculations are presented for the energy locations and spatial structures of low energy resonant states describing transient negative ions (TNIs) of the uracil molecule in the gas phase. The resonant states are modeled using scattering calculations of low energy electrons interacting with isolated molecules in their equilibrium geometry. The interaction forces used in this model are described in detail. Examination of the spatial densities of the excess resonant electrons for the various TNIs found by the calculations allows one to associate the metastable anions with specific features of the experimentally observed fragmentation patterns. PMID- 15267656 TI - Potential energy surfaces for HenNe+ ions: ab initio and diatomics-in-molecule results. AB - The potential energy surface of He2Ne+ has been reinvestigated using a combination of ab initio and diatomics-in-molecule (DIM) calculations. In contrast to the reports of two recent studies the ion is found to have an asymmetric linear He-Ne-He structure, with no barrier to formation from the separated atoms on the ground-state surface. The He-Ne+ bond lengths at the potential minimum are 1.51 and 1.81 A, and the total bonding energy is 0.717 eV. Comparing the He2Ne+ energy to that of HeNe+, the bonding energy for the second helium atom is 0.06 eV, about 10% of that of the first He atom. The saddle point between the two equivalent minima is a symmetric structure, 0.0074 eV above the potential minimum. A symmetric geometry becomes the overall potential minimum if the 2s hole on the Ne is excluded from the reference states of a multireference configuration interaction calculation. A DIM potential was created for the HenNe+ family of ions. The DIM potential is consistent with the asymmetric He2Ne+ ion serving as a core; it predicts a slightly more asymmetric geometry than the ab initio results. Additional helium atoms form five-membered rings around the bonds of the core ion to fill the first shell and then add to the ends of the cluster. The asymmetric core ion and the highly compact structure help to account for the lack of apparent shell structure in the mass spectrometry of HenNe+ clusters. Finally, we recommend that the value De=0.63+/-0.04 eV be adopted for the ground state of HeNe+. PMID- 15267657 TI - A computational study of microsolvation effect on ethylene glycol by density functional method. AB - This study focuses on the conformational analysis of ethylene glycol-(water)n (n=1-3) complex by using density functional theory method and the basis set 6 311++G*. Different conformers are reported and the basis set superposition error corrected total energy is -306.767 5171, -383.221 3135, and -459.694 1528 for lowest energy conformer with 1, 2, and 3 water molecules, respectively, with corresponding binding energy -7.75, -15.43, and -36.28 kcal/mol. On applying many body analysis it has been found that relaxation energy, two-body, three-body energy have significant contribution to the binding energy for ethylene glycol (water)3 complex whereas four-body energies are negligible. The most stable conformers of ethylene glycol-(water)n complex are the cyclic structures in which water molecules bridge between the two hydroxyl group of ethylene glycol. PMID- 15267658 TI - Ground and valence excited states of BI: a MR-CISD+Q study. AB - Ab initio calculations on the valence electronic states of the BI molecule have been performed by using the entirely uncontracted all-electronic aug-cc-pVQZ (for the B atom) and Sadlej-pVTZ (for the I atom) basis sets and the internally contracted multireference singles and doubles configuration interaction method with Davidson size-extensively correction and Douglas-Kroll scalar relativistic correction. The potential energy curves of all valence states and the spectroscopic constants of bound states are fitted. It is the first time that the 12 Lambda-S states of BI molecule and all of the 23 Omega states generated from the former are studied in a theoretical way. Calculation results reproduce well most of the experimental data. The effects of the spin-orbit coupling and the avoided crossing rule between Omega states of the same symmetry are analyzed. The transition properties of the A3Pi0+, B3Pi1, and C1Pi1 states to the ground-state transitions are predicted, including the transition dipole moments, the Franck Condon factors, and the radiative lifetimes. The radiative lifetime of the C1Pi1 state of BI molecule is less than 1 micros, while that of the A3Pi0+ and B3Pi1 states are the order of 1 ms. PMID- 15267659 TI - Ab initio study of the BiSe and BiTe electronic spectra: what happens with X2-X1 emission in the heavier Bi chalcogenides? AB - A series of spin-orbit configuration interaction calculations has been carried out for the BiSe and BiTe molecules and analyzed in comparison with data obtained earlier for the isovalent BiO and BiS systems. An avoided crossing caused by the spin-orbit interaction between the X2Pi and A4Pi electronic states is shown to have a decisive effect on the lower-energy spectrum in each case. Irregularities in the X2 3/2 state vibrational manifold occur as a consequence of this nonadiabatic interaction, and the v vibrational number for the onset of these perturbations is found to gradually decrease in going from BiO to BiSe, in agreement with experiment. In BiTe the shape of the X2 potential curve is so altered by the avoided crossing that its minimum becomes shifted to a significantly larger distance than for the X1 state, unlike the case for BiSe or the lighter Bi chalcogenides. This characteristic appears to be the root cause for the fact that the X2 state has not yet been found experimentally in the BiTe spectrum, despite careful searches in the expected energy range. Radiative lifetimes have also been calculated for the low-lying states of both the BiSe and BiTe molecules, and these results are found to be consistent with experimental observations. PMID- 15267660 TI - State-to-state rotational relaxation rate constants for CO+Ne from IR-IR double resonance experiments: comparing theory to experiment. AB - IR-IR double-resonance experiments were used to study the state-to-state rotational relaxation of CO with Ne as a collision partner. Rotational levels in the range Ji=2-9 were excited and collisional energy transfer of population to the levels Jf=2-8 was monitored. The resulting data set was analyzed by fitting to numerical solutions of the master equation. State-to-state rate constant matrices were generated using fitting law functions. Fitting laws based on the modified exponential gap (MEG) and statistical power exponential gap (SPEG) models were used; the MEG model performed better than the SPEG model. A rate constant matrix was also generated from scattering calculations that employed the ab initio potential energy surface of McBane and Cybulski [J. Chem. Phys. 110, 11 734 (1999)]. This theoretical rate constant matrix yielded kinetic simulations that agreed with the data nearly as well as the fitted MEG model and was unique in its ability to reproduce both the rotational energy transfer and pressure broadening data for Ne-CO. The theoretical rate coefficients varied more slowly with the energy gap than coefficients from either of the fitting laws. PMID- 15267661 TI - A multimode analysis of the gas-phase photoelectron spectra in oligoacenes. AB - We present a multimode vibrational analysis of the gas-phase ultraviolet photoelectron spectra of the first ionization in anthracene, tetracene, and pentacene, using electron-vibration constants computed at the density functional theory level. The first ionization of each molecule exhibits a high-frequency vibronic structure; it is shown that this regularly spaced feature is actually the consequence of the collective action of several vibrational modes rather than the result of the interaction with a single mode. We interpret this feature in terms of the missing mode effect. We also discuss the vibronic coupling constants and relaxation energies obtained from the fit of the photoelectron spectra with the linear vibronic model. PMID- 15267662 TI - Resonantly enhanced two photon ionization and zero kinetic energy spectroscopy of jet-cooled 4-aminopyridine. AB - We report studies of supersonically cooled 4-aminopyridine (4-AP) using two-color resonantly enhanced multiphoton ionization (REMPI) and two-color zero kinetic energy (ZEKE) photoelectron spectroscopy. With the aid of ab initio and density functional calculations, vibrational modes of the first electronically excited state (S1) of the neutral species and those of the cation have been assigned, and the adiabatic ionization potential has been determined to be 62291+/-6 cm(-1). The REMPI spectrum of the S1 state is dominated by ring deformation modes and the inversion mode of the amino group, while the ZEKE spectra demonstrate a strong propensity of Deltav=0, where v is the vibrational quantum number of the intermediate vibronic state from S1. In addition, the ZEKE spectra obtained via different vibrational levels of the S1 state contain four common features, corresponding to the activation of four different vibrational modes of the cation. These observations are explained in terms of the structural changes from the ground state to S1 and further to the cation. The vibrational mode distributions in both the REMPI and the ZEKE spectra, the excitation energy of the S1 state, and the ionization potential of 4-AP, are remarkably similar to those of aniline, suggesting that the electronic activity is centered on the ring. PMID- 15267663 TI - Variationally stable calculations for molecular systems: polarizabilities and two photon ionization cross section for the hydrogen molecule. AB - The variationally stable method of Gao and Starace [B. Gao and A. F. Starace, Phys. Rev. Lett. 61, 404 (1988); Phys. Rev. A 39, 4550 (1989)] has been applied for the first time to the study of multiphoton processes in molecular systems. The generalization in theory is presented, as well as the calculation of properties such as the static and dynamic polarizabilities of the hydrogen molecule and the generalized two-photon ionization cross section. The Schwinger variational iterative method [R. R. Lucchese and V. McKoy, Phys. Rev. A 21, 112 (1980)] has been applied in the achievement of the photoelectron wave function, while a Hartree-Fock representation has been used for the target. This research has been motivated by the scarceness of ab initio calculations of molecular multiphoton ionization cross sections in the literature. PMID- 15267664 TI - Spectroscopic investigation of nonbonding interactions of group-14 atoms with rare gases: the SnAr van der Waals complex. AB - The laser fluorescence excitation spectra of the SnAr van der Waals complex, in the vicinity of the individual fine-structure lines of the Sn 5s25p6s3P0<-- 5s25p2(3)P atomic resonance transition in the spectral region 317-270 nm are reported. Excited-state (v',0) progressions of bands built upon the individual J'<-- J" fine-structure atomic lines were observed. Because the collisional spin orbit relaxation was slow, transitions were observed out of the lower SnAr states built upon all the J'' atomic asymptotes. The spectra were interpreted through model potential energy curves based on the isoelectronic SiAr system. Lower bounds to the dissociation energies of all lower SnAr states were determined. The binding energies of the group-13, and -14-atom-argon complexes and the effect of the spin-orbit interaction on moderating nonbonding interactions are discussed. PMID- 15267665 TI - Electronic and infrared absorption spectra of linear and cyclic C6+ in a neon matrix. AB - Electronic and infrared absorption spectra of mass-selected C6+, generated by dissociative electron impact ionization of C6Cl6 and C6Br6, have been recorded in 6 K neon matrices. Linear and cyclic forms of C6+ have been observed. The 2Pig<- Chi2Piu electronic transition of linear C6+ has its origin band at 646 nm whereas for the (2) 2B2<--Chi2A1 system of the cyclic isomer it lies at 570 nm. An infrared active fundamental mode in the ground electronic state of C6+ is observed at 2092 and 1972 cm(-1) for the linear and cyclic isomer, respectively. PMID- 15267666 TI - Transport of matter and energy in a mesoscopic thermo-hydrodynamic approach. AB - We derive a thermo-hydrodynamic theory for particles and energy flow, based on a nonequilibrium grand-canonical ensemble formalism. The time-dependent kinetic coefficients are explicitly given in terms of microscopic mechanical quantities. The time evolution equations describing the coupled flow of energy and particles are derived. The second-rank tensorial fluxes of current of energy and particles present in the nonequilibrium ensemble are nondiagonal. We obtain a generalized Fick's Law, which presents the effect of the energy flow on the particle diffusion equation. PMID- 15267667 TI - Solvent reorganization energy of electron-transfer reactions in polar solvents. AB - A microscopic theory of solvent reorganization energy in polar molecular solvents is developed. The theory represents the solvent response as a combination of the density and polarization fluctuations of the solvent given in terms of the density and polarization structure factors. A fully analytical formulation of the theory is provided for a solute of arbitrary shape with an arbitrary distribution of charge. A good agreement between the analytical procedure and the results of Monte Carlo simulations of model systems is achieved. The reorganization energy splits into the contributions from density fluctuations and polarization fluctuations. The polarization part is dominated by longitudinal polarization response. The density part is inversely proportional to temperature. The dependence of the solvent reorganization energy on the solvent dipole moment and refractive index is discussed. PMID- 15267668 TI - New approach to the first-order phase transition of Lennard-Jones fluids. AB - The multicanonical Monte Carlo method is applied to a bulk Lennard-Jones fluid system to investigate the liquid-solid phase transition. We take the example of a system of 108 argon particles. The multicanonical weight factor we determined turned out to be reliable for the energy range between -7.0 and -4.0 kJ/mol, which corresponds to the temperature range between 60 and 250 K. The expectation values of the thermodynamic quantities obtained from the multicanonical production run by the reweighting techniques exhibit the characteristics of first order phase transitions between liquid and solid states around 150 K. The present study reveals that the multicanonical algorithm is particularly suitable for analyzing the transition state of the first-order phase transition in detail. PMID- 15267669 TI - An efficient molecular dynamics simulation method for calculating the diffusion influenced reaction rates. AB - We present a molecular dynamics (MD) simulation method for calculating the diffusion-influenced reaction rates in the limit of low reactant concentrations. To calculate the reaction rate coefficient, we use MD trajectories of a nonreactive equilibrium system that are initiated with a pair of reactant molecules in reactive configuration. Hence reaction systems involving complicated reactant molecules with geometrically restricted reactivities can be treated with comparable efficiency as the simple hard-sphere reaction system. Compared to the similar MD method proposed by Van Beijeren, Dong, and Bocquet [J. Chem. Phys. 114, 6265 (2001)], the present method has a couple of advantages. First, reactions involving more general sink functions can be treated. Second, more accurate results can be obtained when the reaction probability upon collision is less than unity. As an application, we investigate the effects of nondiffusive dynamics and hydrodynamic interaction of reactants on the reaction rate. PMID- 15267670 TI - Dynamics and the breaking of a driven cage: I2 in solid Ar. AB - Pump-probe measurements of I2 in solid Ar are reported and analyzed to extract a description of cage response to impulsive excitation, from the gentle kick, up to the breaking point. The most informative data are obtained through wavepacket motion on cage-bound, but otherwise dissociative, potentials where the chromophore acts as a transducer to drive the cage and to report on the local dynamics. This general class of dynamics is identified and analyzed as a function of energy in Ar, Kr, and Xe. The overdriven cage rebounds with a characteristic period of 1.2 ps that shows little dependence on excitation amplitude in all hosts. After rebound, the cage rings as a local resonant mode in Ar, with a period of 1 ps and dephasing time of 3 ps. This mode remains at the Debye edge in Kr and Xe, with periods of 630 and 800 fs, and dephasing times of 8 and 6 ps, respectively. In the bound B-state, the cage fluctuates toward its dilated equilibrium structure on a time scale of 3 ps, which is extracted from the down chirp in the molecular vibrational frequency. When kicked with excess energy of 4 eV, the Ar cage breaks with 50% probability, and the molecule dissociates. The kinetics of polarization selective, multiphoton dissociation with Gaussian laser intensity profiles is delineated and the ballistics of cage breakout is described: The photodissociation proceeds by destruction of the local lattice, by creating interstitials and vacancies. During large amplitude motion on cage-bound potentials, sudden, nonadiabatic spin-flip transitions can be observed and quantified in space and time. The spin-flip occurs with unit probability in Ar when the I*-I bond is stretched beyond 6 A. PMID- 15267671 TI - Mode-coupling study on the dynamics of hydrophobic hydration. AB - The molecular motion of water in water-hydrophobic solute mixtures was investigated by the mode-coupling theory for molecular liquids based on the interaction-site description. When the model Lennard-Jones solute was mixed with water, both the translational and reorientational motions of solvent water become slower, in harmony with various experiments and molecular dynamics simulations. We compared the mechanism of the slowing down with that of the pressure dependence of the molecular motion of neat water [T. Yamaguchi, S.-H. Chong, and F. Hirata, J. Chem. Phys. 119, 1021 (2003)]. We found that the decrease in the solvent mobility caused by the solute can essentially be elucidated by the same mechanism: That is, the fluctuation of the number density of solvent due to the cavity formation by the solute strengthens the friction on the collective polarization through the dielectric friction mechanism: We also employed the solute molecule that is the same as solvent water except for the amount of partial charges, in order to alter the strength of the solute-solvent interaction continuously. The mobility of the solvent water was reduced both by the hydrophobic and strongly hydrophilic solutes, but it was enhanced in the intermediate case. Such a behavior was discussed in connection with the concept of positive and negative hydrations. PMID- 15267672 TI - Magnetic resonance imaging of spin-polarization transfer of polarized Xe atoms dissolving into ethanol. AB - We detect the free-induction signals of xenon atoms polarized by spin-exchange optical pumping. The temperature dependence of dissolution and spin-polarization transfer of xenon atoms to ethanol is measured by simultaneous detection of both xenon and proton signals. The polarization of proton is efficiently enhanced in the xenon-saturated solution at low magnetic fields. The large polarization and chemical shift enable us to obtain clearly the distribution image of xenon atoms near the gas-liquid and liquid-liquid boundaries. Therefore the localization of polarized xenon atoms is observed near the surface. By time-resolved magnetic resonance imaging of polarized xenon and polarization-enhanced proton, the spin dynamics is qualitatively studied for the nuclear spins interacting with each other in a dense solution. PMID- 15267673 TI - Molecular dynamics investigation of ferrous-ferric electron transfer in a hydrolyzing aqueous solution: calculation of the pH dependence of the diabatic transfer barrier and the potential of mean force. AB - We present a molecular model for ferrous-ferric electron transfer in an aqueous solution that accounts for electronic polarizability and exhibits spontaneous cation hydrolysis. An extended Lagrangian technique is introduced for carrying out calculations of electron-transfer barriers in polarizable systems. The model predicts that the diabatic barrier to electron transfer increases with increasing pH, due to stabilization of the Fe3+ by fluctuations in the number of hydroxide ions in its first coordination sphere, in much the same way as the barrier would increase with increasing dielectric constant in the Marcus theory. We have also calculated the effect of pH on the potential of mean force between two hydrolyzing ions in aqueous solution. As expected, increasing pH reduces the potential of mean force between the ferrous and ferric ions in the model system. The magnitudes of the predicted increase in diabatic transfer barrier and the predicted decrease in the potential of mean force nearly cancel each other at the canonical transfer distance of 0.55 nm. Even though hydrolysis is allowed in our calculations, the distribution of reorganization energies has only one maximum and is Gaussian to an excellent approximation, giving a harmonic free energy surface in the reorganization energy F(DeltaE) with a single minimum. There is thus a surprising amount of overlap in electron-transfer reorganization energies for Fe(2+)-Fe(H2O)6(3+), Fe(2+)-Fe(OH)(H2O)5(2+), and Fe(2+)-Fe(OH)2(H2O)+ couples, indicating that fluctuations in hydrolysis state can be viewed on a continuum with other solvent contributions to the reorganization energy. There appears to be little justification for thinking of the transfer rate as arising from the contributions of different hydrolysis states. Electronic structure calculations indicate that Fe(H2O)6(2+)-Fe(OH)n(H2O)(6-n)(3-n)+ complexes interacting through H3O2- bridges do not have large electronic couplings. PMID- 15267674 TI - Electron correlation effects in the adiabatic charge transfer reactions at the metal/polar liquid interface. AB - New simple expressions for average number of electrons in the valence orbital of a reacting ion and the charge susceptibility are obtained that allow one to calculate adiabatic free energy surfaces (AFES) and corresponding kinetic regime diagrams (KRD) for adiabatic processes of electron transfer from the ion, located in a polar liquid, to a metal within the framework of the exactly solvable (in the limit T-->0) model of the metal with the infinitely wide conduction band. This model represents one of limiting cases of the Anderson model that may be applied to s-p metals. Unlike previous studies of the adiabatic reactions in the model of the metal with the infinitely wide conduction band, the present work takes into account the electron-electron correlation effects in an exact manner. General results are illustrated with KRD which determine the regions of the physical parameters of the system corresponding to various types of electron transfer processes. AFES are calculated for some typical parameters sets. The exact AFES are compared with those calculated within the Hartree-Fock approximation. It is shown that the correlation effects are of importance and results not only in a considerable decrease of the activation free energy but also to qualitatively different shapes of AFES in some regions of the system parameters. PMID- 15267675 TI - A cell model of a liquid droplet. AB - An expression for the free energy of a droplet composed of attracting hard spheres is found using a simple cell model. The hard-sphere repulsion is assumed to act only between molecules in the same cell, whereas attraction extends over many cells. A maximum term analysis gives rise to a mean-field free energy which includes terms proportional to the first and second power of the droplet radius R with coefficients which can be related to the planar surface tension and Tolman length. Certain Gaussian fluctuations about the maximum term are also considered, corresponding to droplet translation and capillary wave fluctuations. Inclusion of these fluctuations is necessary to ensure that the nucleation rate is proportional to the system volume. They also reduce the planar surface tension and introduce a logarithmic term, -4/3 ln R, into the free energy. The inclusion of other fluctuations and the relationship between these equations and those arising in density-functional theories of nucleation is discussed. PMID- 15267676 TI - Temperature dependent exciton emission from herringbone aggregates of conjugated oligomers. AB - In this work, the effect of temperature, exciton bandwidth, and size on the photoluminescence spectra of defect-free two-dimensional herringbone aggregates of pi-conjugated oligomers such as oligophenylene vinylene and oligothiophene is investigated theoretically. The model is based on exciton-phonon coupling in two dimensional herringbone lattices with the exciton deriving from the lowest optical (1Ag-->1Bu) transition and the phonon from the most strongly coupled intramolecular vibrational mode with frequency omega0. Simple analytical expressions are obtained for the line strengths of the emission origin (0-0) and first replica (0-1) as a function of the number of molecules comprising the aggregate, N, the free exciton bandwidth, WD, and the temperature, T. At a given temperature, the 0-0 emission intensity initially scales as N/Nth, where Nth is the superradiant threshold number, but eventually converges to NT/Nth, where NT is the size independent thermal coherence number. NT is inversely proportional to temperature and proportional to the exciton band curvature (omegac) near the band bottom; NT=1+4piomegac/kbT. In striking contrast, the 0-1 line strength is relatively insensitive to temperature and size, but scales as the inverse square of WD+omega0. The insensitivity of the first replica to the exciton coherence number makes the ratio of the 0-0 to 0-1 line strengths a measure of the exciton coherence number. The ratio can be used to test for crystal purity. Comparison to experiments on thin films of quaterthiophene shows that the thermal coherence size is given by NT approximately 1+450/T (K) and that superradiance, which requires NT>Nth, can only be observed at temperatures less than 1 K. PMID- 15267677 TI - Inverse isotope effects and electron-phonon coupling in the positively charged deutero- and fluoroacenes. AB - Electron-phonon interactions in the monocations of deutero- and fluoroacenes are studied and compared with those in the monocations of acenes and those in the monoanions of fluoroacenes. Because of the significant phase pattern difference between the highest occupied molecular orbitals (HOMO) and the lowest unoccupied molecular orbitals (LUMO), the frequency modes lower than 500 cm(-1) and the high frequency modes around 1400 cm(-1) couple more strongly to the LUMO than to the HOMO, while the frequency modes around 500 cm(-1) and the frequency modes around 1600 cm(-1) couple more strongly to the HOMO than to the LUMO in fluoroacenes with D2h geometry. The total electron-phonon coupling constants for the monocations (l(HOMO)) are estimated and compared with those for the monoanions (l(LUMO)) in deutero- and fluoroacenes. The l(HOMO) values are estimated to be 0.418, 0.399, 0.301, 0.255, and 0.222 eV for C6F6 (1f), C10F8 (2f), C14F10 (3f), C18F12 (4f), and C22F14 (5f), respectively. The l(HOMO) values are smaller than the l(LUMO) values in small fluoroacenes. But the l(HOMO) value decreases with an increase in molecular size less rapidly than the l(LUMO) value in fluoroacenes, and the l(HOMO) value of 0.074 eV is much larger than the l(LUMO) value of 0.009 eV in polyfluoroacene. The logarithmically averaged phonon frequencies for the monocations (omega(ln,HOMO)) are estimated to be larger than those for the monoanions (omega(ln,LUMO)) in fluoroacenes. This is because the C-C stretching modes around 1600 cm(-1) couple most strongly to the HOMO, and those around 1400 cm(-1) couple the most strongly to the LUMO in fluoroacenes. The significant phase pattern difference between the HOMO and the LUMO is the main reason for the calculational results. The l(HOMO) values increase much more significantly by H-F substitution than by H-D substitution in acenes. The possible inverse isotope effects in the electron-phonon interactions as a consequence of deuteration in the monocations of nanosized molecules are suggested. PMID- 15267678 TI - Adsorption of O2 and oxidation of CO at Au nanoparticles supported by TiO2(110). AB - Density functional theory calculations are performed for the adsorption of O2, coadsorption of CO, and the CO+O2 reaction at the interfacial perimeter of nanoparticles supported by rutile TiO2(110). Both stoichiometric and reduced TiO2 surfaces are considered, with various relative arrangements of the supported Au particles with respect to the substrate vacancies. Rather stable binding configurations are found for the O2 adsorbed either at the trough Ti atoms or leaning against the Au particles. The presence of a supported Au particle strongly stabilizes the adsorption of O2. A sizable electronic charge transfer from the Au to the O2 is found together with a concomitant electronic polarization of the support meaning that the substrate is mediating the charge transfer. The O2 attains two different charge states, with either one or two surplus electrons depending on the precise O2 adsorption site at or in front of the Au particle. From the least charged state, the O2 can react with CO adsorbed at the edge sites of the Au particles leading to the formation of CO2 with very low (approximately 0.15 eV) energy barriers. PMID- 15267679 TI - Molecular-dynamics simulation of the effect of ions on a liquid-liquid interface for a partially miscible mixture. AB - Molecular-dynamics simulations were performed to model the effect of added salt ions on the liquid-liquid interface in a partially miscible system. Simulations of the interface between saturated phases of a model 1-hexanol+water system show a bilayer structure of 1-hexanol molecules at the interface with -OH heads of the first layer directed into the water phase and the opposite orientation for the second layer. The alignment of the polar -OH groups at the interface stabilizes a charge separation of sodium and chloride ions when salt is introduced into the aqueous phase, producing an electrical double layer. Chloride ions aggregate nearer the interface and sodium ions move toward the bulk water phase, consistent with the explanation that the -OH alignment presents a region of partial positive charges to which the hydrated chloride atoms are attracted. Ions near the interface were found to be less solvated than those in the bulk phase. An electric field was also applied to drive ions through the interface. Ions crossing the interface tended to shed water molecules as they entered the hexanol bilayer, leaving a trail of water molecules. Stabilization and facilitated transport of the ion by interactions with the second layer of hexanol molecules appeared to be an important step in the mechanism of sodium ion transport. PMID- 15267680 TI - Clustering and percolation in lithium borate glasses. AB - Molecular dynamics simulations are carried out in xLi2O-(1-x)B2O3 glasses (x=0.2 0.6) at T=1250 K, where cluster size distributions for Li cations and nonbridging oxygen (NBO) atoms are calculated. The existence of percolating clusters above x=0.3 places the percolation threshold between x=0.3 and 0.4 for the system under investigation, which is consistent with the abrupt increase of the diffusion coefficient of Li cations observed at x=0.4. It is also shown that the clusters of Li cations consist mainly of Li atoms found in the vicinity of NBO atoms. This result explains the higher mobility exhibited by this type of cations compared to the mobility of Li cations in the vicinity of bridging oxygen atoms. PMID- 15267681 TI - Theoretical studies of hyperthermal O(3P) collisions with hydrocarbon self assembled monolayers. AB - We present a dynamics study of inelastic and reactive scattering processes in collisions of hyperthermal (5 eV) O(3P) atoms with a hydrocarbon self-assembled monolayer (SAM). Molecular-dynamics simulations are carried out using a quantum mechanics/molecular mechanics (QM/MM) interaction potential that uses a high quality semiempirical Hamiltonian for the QM part and the MM3 force field for the MM part. A variety of products coming from reaction are identified, including H abstraction to generate OH, O atom addition to the SAM with subsequent elimination of H atoms, and direct C-C breakage. The C-C breakage mechanism provides a pathway for significant surface mass loss in single reactive events whereas the O addition-H elimination channel leads to surface oxidation. Reaction probabilities, product energy, and angular distributions are examined to gain insight on polymer erosion in low Earth orbit conditions and on fundamentals of inelastic and reactive hyperthermal gas-surface interactions. PMID- 15267682 TI - Pattern formation and fluctuation-induced transitions in protein crystallization. AB - A kinetic model of protein crystallization accounting for the nucleation stage, the growth and competition of solid particles and the formation of macroscopic patterns is developed. Different versions are considered corresponding successively, to a continuous one-dimensional crystallization reactor, a coarse grained two-box model and a model describing the evolution of the space averaged values of fluid and solid material. The analysis brings out the high multiplicity of the patterns. It provides information on their stability as well as on the kinetics of transitions between different states under the influence of the fluctuations. PMID- 15267683 TI - Determining the adsorptive and catalytic properties of strained metal surfaces using adsorption-induced stress. AB - We demonstrate a model for determining the adsorptive and catalytic properties of strained metal surfaces based on linear elastic theory, using first-principles calculations of CO adsorption on Au and K surfaces and CO dissociation on Ru surface. The model involves a single calculation of the adsorption-induced surface stress on the unstrained metal surface, which determines quantitatively how adsorption energy changes with external strain. The model is generally applicable to both transition- and non-transition-metal surfaces, as well as to different adsorption sites on the same surface. Extending the model to both the reactant and transition state of surface reactions should allow determination of the effect of strain on surface reactivity. PMID- 15267684 TI - Spectroscopic consideration of the surface potential built across phthalocyanine thin films on a metal electrode. AB - The nonlinear optical properties of tert-butyl phthalocyanine copper Langmuir Blodgett (CuttbPc LB) films and vacuum-evaporated phthalocyanine copper (CuPc) films deposited on a metal surface were investigated by second-harmonic generation (SHG) spectroscopy. At the organic/metal interface, a space charge field is formed due to the presence of excess charge injected from a metal electrode to the organic layer. Since the Pc molecule has D4h symmetry, an inversion center is present and the optical SH process is not allowed under the electric-dipole approximation. However, the space charge field at the interface directly influences the symmetric structure of the electrons in the Pc molecule. We investigated the contributions of the surface potential to the SHG using Pc LB and vacuum-evaporated films deposited on aluminum (Al) and gold (Au) metal electrodes, where a distinctive difference in the spectrum for the Pc films on the Al and Au surfaces was observed. The contribution of the surface potential was revealed based on the resonant conditions of the SH process, taking into account the electric-quadrupole transition and dc-field-induced electric-dipole transition. PMID- 15267685 TI - Electronic transport in Z-junction carbon nanotubes. AB - In this paper, the electronic transport in different Z-shape carbon nanotubes containing double knee junction structures on the same tube is studied. One consists of (5,5)-(9,0)-(5,5) double knee nano-metal-metal-metal junctions and another consists (6,6)-(10,0)-(6,6) double knee nano-metal-semiconductor-metal junctions. With the nearest-neighbor pi-orbital tight-binding model, quantum conductances of these double knee junctions are calculated using the Landauer formula. The interesting conductance curves are provided to exhibit a potential application in the arena of molecular electronics. PMID- 15267686 TI - Electronic states of linear Au clusters supported on metal surfaces: why are they like those of a particle in a box? AB - Scanning tunneling spectroscopy and microscopy show that the empty states of linear Au clusters supported on a metal surface behave as if they are the states of an electron in an empty one-dimensional box. We show here that certain difficulties of this description are removed by a particle-in-a-cylinder model. This interpretation is supported by density functional calculations. PMID- 15267687 TI - Atomic and electronic structure of unreduced and reduced CeO2 surfaces: a first principles study. AB - The atomic and electronic structure of (111), (110), and (100) surfaces of ceria (CeO2) were studied using density-functional theory within the generalized gradient approximation. Both stoichiometric surfaces and surfaces with oxygen vacancies (unreduced and reduced surfaces, respectively) have been examined. It is found that the (111) surface is the most stable among the considered surfaces, followed by (110) and (100) surfaces, in agreement with experimental observations and previous theoretical results. Different features of relaxation are found for the three surfaces. While the (111) surface undergoes very small relaxation, considerably larger relaxations are found for the (110) and (100) surfaces. The formation of an oxygen vacancy is closely related to the surface structure and occurs more easily for the (110) surface than for (111). The preferred vacancy location is in the surface layer for CeO2(110) and in the subsurface layer (the second O-atomic layer) for CeO2(111). For both surfaces, the O vacancy forms more readily than in the bulk. An interesting oscillatory behavior is found for the vacancy formation energy in the upper three layers of CeO2(111). Analysis of the reduced surfaces suggests that the additional charge resulting from the formation of the oxygen vacancies is localized in the first three layers of the surface. Furthermore, they are not only trapped in the 4f states of cerium. PMID- 15267688 TI - Infrared spectroscopy of physisorbed and chemisorbed O2 on Pt(111). AB - The adsorption of O2 on the Pt(111) surface, with particular emphasis on the influence of substrate temperature, has been studied by infrared reflection absorption spectroscopy (IRAS). In the temperature range 30-90 K the IRAS spectra reveal three different molecular adsorption states. A physisorbed state appears below 40 K while chemisorbed peroxo- and superoxo-like states are observed in the whole temperature range, the characteristic vibrational frequencies are at full coverages of 16O2, 1543 cm(-1) and around 700 cm(-1) and 870 cm(-1), respectively. Flash heating from 30 K to 45 K reveal that the physisorbed state acts as a precursor to the superoxo chemisorption. Theoretical calculations suggest that peroxo molecules may occupy both fcc and hcp threefold sites on the Pt(111) surface. However, within the high resolution of the IRAS measurements we only observe one peroxo state in the temperature range 45-90 K, assigned to occupy the fcc site. The peroxo adsorption probability is significantly lower at 45 K than at 90 K, presumably due to reduced thermal activation from the physisorbed precursor state. A longer lifetime in this precursor state at the low temperature results in formation of larger superoxo islands already at low oxygen coverage. PMID- 15267689 TI - Rate-promoting vibrations and coupled hydrogen-electron transfer reactions in the condensed phase: a model for enzymatic catalysis. AB - A model is presented for coupled hydrogen-electron transfer reactions in condensed phase in the presence of a rate promoting vibration. Large kinetic isotope effects (KIEs) are found when the hydrogen is substituted with deuterium. While these KIEs are essentially temperature independent, reaction rates do exhibit temperature dependence. These findings agree with recent experimental data for various enzyme-catalyzed reactions, such as the amine dehydrogenases and soybean lipoxygenase. Consistent with earlier results, turning off the promoting vibration results in an increased KIE. Increasing the barrier height increases the KIE, while increasing the rate of electron transfer decreases it. These results are discussed in light of other views of vibrationally enhanced tunneling in enzymes. PMID- 15267690 TI - Tunable effective interactions between dendritic macromolecules. AB - We employ extensive Monte Carlo and molecular-dynamics simulations to investigate the effective interactions between the centers of mass of dendritic macromolecules of variable flexibility and generation number. Two different models for the connectivity and steric interactions between the monomers are employed, the first one being purely entropic in nature and the second explicitly involving energetic interactions. We find that the effective potentials have a generic Gaussian shape, whose range and strength can be tuned via modifications in the generation number and flexibility of the spacers. We supplement our simulation analysis by a density-functional approach in which the connectivity between the monomers is approximated by an external confining potential that holds the monomer beads together. Using a simple density functional for the interactions between the monomers, we find semiquantitative agreement between theory and simulation. The implications of our findings for the interpretation of scattering data from concentrated dendrimer solutions are also discussed. PMID- 15267691 TI - Computer simulations of localized small polarons in amorphous polyethylene. AB - We use a simple mean field scheme to compute the polarization energy of an excess electron in amorphous polyethylene that allows us to study dynamical properties. Nonadiabatic simulations of an excess electron in amorphous polyethylene at room temperature show the spontaneous formation of localized small polaron states in which the electron is confined in a spherically shaped region with a typical dimension of 5 A. We compute the self-trapping energy to be -0.06+/-0.03 eV, with a lifetime on the time scale of a few tens of picoseconds. PMID- 15267692 TI - Microscopic calculation of the energetics of charged states in amorphous polyethylene. AB - Polarization energies are calculated for a single excess charge on a polyethylene chain in amorphous polyethylene using (i) local segment and nonlocal distributed molecular polarizabilities, (ii) material structures simulated by both general purpose and specialist Monte Carlo software, and (iii) uniform and Gaussian distributions of charge with different extents of charge delocalization. Local and distributed response lead to results that are essentially the same except that they correspond to different mean polarizabilities. With increasing delocalization of the charge along the chain, the polarization energies shift to higher values and the width of their distribution decreases, the differences being more pronounced for the uniform distribution. The polarization energies for charges delocalized over 10-20 methylene units form a distribution some 14 eV wide centered around 1 eV, narrowing significantly for more homogeneous polymer melts. The calculations are relevant to trapping of charge in polyethylene. They also yield the microscopic variation in the potential along the polymer chain caused by the polarization energy difference, and so may provide useful inputs to theories of electronic conduction in polymer materials. PMID- 15267693 TI - Polymer dynamics in repton model at large fields. AB - Polymer dynamics at large fields in Rubinstein-Duke repton model is investigated theoretically. Simple diagrammatic approach and analogy with asymmetric simple exclusion models are used to analyze the reptation dynamics of polymers. It is found that for polyelectrolytes the drift velocity decreases exponentially as a function of the external field with an exponent depending on polymer size and parity, while for polyampholytes the drift velocity is independent of polymer chain size. However, for polymers, consisting of charged and neutral blocks, the drift velocity approaches the constant limit which is determined by the size of the neutral block. The theoretical arguments are supported by extensive numerical calculations by means of density-matrix renormalization group techniques. PMID- 15267694 TI - Molecular dynamics simulation study on the phase behavior of the Gay-Berne model with a terminal dipole and a flexible tail. AB - To study the effect of the alkyl tail and the terminal dipole on the stability of the liquid crystalline phase of mesogens, we have carried out molecular dynamics simulations for 1CB(4-methyl-4'-cyanobiphenyl) and 5CB(4-n-pentyl-4' cyanobiphenyl) by using a coarse-grained model. In the coarse-grained model, a 5CB molecule is divided into the rigid part of 1CB moiety, which is represented by an ellipsoid, and the remaining flexible part which is represented by a chain of united atoms. The nonbonded potential between coarse-grained segments is represented by the generalized Gay-Berne (GB) potential and the potential parameters are determined by directly comparing the GB potential with the atomistic potentials averaged over the rotation of the mesogen around its axis. In addition, a dipole moment is placed at one end of the ellipsoid opposite to the flexible tail. The ordered state obtained in the polar 5CB model was assigned as the nematic phase, and the experimental static and dynamical properties were reproduced well by using this coarse-grained model. Both the dipole-dipole interactions and the thermal fluctuation of the flexible tail increase the positional disorder in the director direction, and stabilize the nematic phase. Thus, the nematic phase in the polar 5CB is induced by a cooperative effect of the flexible tail and the terminal dipole. It is noted that a local bilayer structure with head-to-head association is formed in the nematic phase, as experimentally observed by x-ray diffraction measurements. PMID- 15267695 TI - Scaling behavior of nonisothermal phase separation. AB - The phase separation process in a critical mixture of polydimethylsiloxane and polyethylmethylsiloxane (PDMS/PEMS, a system with an upper critical solution temperature) was investigated by time-resolved light scattering during continuous quenches from the one-phase into the two-phase region. Continuous quenches were realized by cooling ramps with different cooling rates kappa. Phase separation kinetics is studied by means of the temporal evolution of the scattering vector qm and the intensity Im at the scattering peak. The curves qm(t) for different cooling rates can be shifted onto a single mastercurve. The curves Im(t) show similar behavior. As shift factors, a characteristic length Lc and a characteristic time tc are introduced. Both characteristic quantities depend on the cooling rate through power laws: Lc approximately kappa(-delta) and tc approximately kappa(-rho). Scaling behavior in isothermal critical demixing is well known. There the temporal evolutions of qm and Im for different quench depths DeltaT can be scaled with the correlation length xi and the interdiffusion coefficient D, both depending on DeltaT through critical power laws. We show in this paper that the cooling rate scaling in nonisothermal demixing is a consequence of the quench depth scaling in the isothermal case. The exponents delta and rho are related to the critical exponents nu and nu* of xi and D, respectively. The structure growth during nonisothermal demixing can be described with a semiempirical model based on the hydrodynamic coarsening mechanism well known in the isothermal case. In very late stages of nonisothermal phase separation a secondary scattering maximum appears. This is due to secondary demixing. We explain the onset of secondary demixing by a competition between interdiffusion and coarsening. PMID- 15267696 TI - Electron transfer in proteins: nonorthogonal projections onto donor-acceptor subspace of the Hilbert space. AB - We develop nonorthogonal projectors, called Lowdin projectors, to construct an effective donor-acceptor system composed of localized donor (D) and acceptor (A) states of a long-distance electron transfer problem. When these states have a nonvanishing overlap with the bridge states these projectors are non-Hermitian and there are various possible effective two-level systems that can be built. We show how these can be constructed directly from the Schrodinger or Dyson equation projected onto the D-A subspace of the Hilbert space and explore these equations to determine the connection between Hamiltonian and Green function partitioning. We illustrate the use of these effective two-level systems in estimating the electron transfer rate in the context of a simple electron transfer model. PMID- 15267698 TI - Interelectronic angle densities of equivalent electrons in Hartree-Fock theory of atoms. AB - The interelectronic angle density A(theta12) is the probability density function that the angle thetaij (0 < or = thetaij < or = pi) subtended by the vectors ri and rj of any two electrons i and j becomes theta12. For equivalent electrons in atoms, it is shown that the density A(theta12) in the Hartree-Fock theory is given by a simple polynomial of cos theta12. Detailed expressions are reported for all LS terms arising from s2, pN (N = 2-6), dN (N = 2-10), and f(N) (N = 2,12) electron configurations. With no modifications, the present results apply as well to the interelectronic angle density A(theta12) in momentum space, where theta12 is the angle between two electron momenta. PMID- 15267699 TI - Monomer basis-set truncation effects in calculations of interaction energies: A model study. AB - Supermolecular interaction energies are analyzed in terms of the symmetry-adapted perturbation theory and operators defining the inaccuracy of the monomer wave functions. The basis set truncation effects are shown to be of first order in the monomer inaccuracy operators. On the contrary, the usual counterpoise correction schemes are of second order in these operators. Recognition of this difference is used to suggest an approach to corrections for basis-set truncation effects. Also earlier claims--that dimer-centered basis sets may lead to interaction energies free of basis-set superposition effects--are shown to be misleading. According to the present study the basis-set truncation contributions, evaluated by means of the symmetry-adapted perturbation theory with monomer-centered basis sets, provide physically and mathematically justified corrections to supermolecular results for interaction energies. PMID- 15267700 TI - A computational strategy for geometry optimization of ionic and covalent excited states, applied to butadiene and hexatriene. AB - We propose a computational strategy that enables ionic and covalent pipi* excited states to be described in a balanced way. This strategy depends upon (1) the restricted active space self-consistent field method, in which the dynamic correlation between core sigma and valence pi electrons can be described by adding single sigma excitations to all pi configurations and (2) the use of a new conventional one-electron basis set specifically designed for the description of valence ionic states. Together, these provide excitation energies comparable with more accurate and expensive ab initio methods--e.g., multiconfigurational second order perturbation theory and multireference configuration interaction. Moreover, our strategy also allows full optimization of excited-state geometries--including conical intersections between ionic and covalent excited states--to be routinely carried out, thanks to the availability of analytical energy gradients. The prototype systems studied are the cis and trans isomers of butadiene and hexatriene, for which the ground 1A(1/g), lower-lying dark (i.e., symmetry forbidden covalent) 2A(1/g) and spectroscopic 1B(2/u) (valence ionic) states were investigated. PMID- 15267701 TI - Body frames in the separation of collective angles in quantum N-body problems. AB - The application of the concept of body-fixed reference frames, proposed by C. Eckart [Phys. Rev. 47, 552 (1935)], to the problem of the separation of three collective angles in quantum N-body problems is analyzed based on the technique recently developed by Meremianin and Briggs [Phys. Rep. 384, 121 (2003)]. Special attention is paid to the body frame defined by the "second Eckart condition" which minimizes vibro-rotational couplings near the equilibrium position. The important case of the Eckart frame for three-body systems is considered in detail. The connection of the basis vectors of the Eckart frame with Jacobi vectors is derived. All results of this work are valid for an arbitrary choice of internal (body-frame) coordinates. PMID- 15267702 TI - A growing string method for determining transition states: comparison to the nudged elastic band and string methods. AB - Interpolation methods such as the nudged elastic band and string methods are widely used for calculating minimum energy pathways and transition states for chemical reactions. Both methods require an initial guess for the reaction pathway. A poorly chosen initial guess can cause slow convergence, convergence to an incorrect pathway, or even failed electronic structure force calculations along the guessed pathway. This paper presents a growing string method that can find minimum energy pathways and transition states without the requirement of an initial guess for the pathway. The growing string begins as two string fragments, one associated with the reactants and the other with the products. Each string fragment is grown separately until the fragments converge. Once the two fragments join, the full string moves toward the minimum energy pathway according to the algorithm for the string method. This paper compares the growing string method to the string method and to the nudged elastic band method using the alanine dipeptide rearrangement as an example. In this example, for which the linearly interpolated guess is far from the minimum energy pathway, the growing string method finds the saddle point with significantly fewer electronic structure force calculations than the string method or the nudged elastic band method. PMID- 15267703 TI - Extracting atoms from molecular electron densities via integral equations. AB - The observation that a molecular electron density is close to the superposition of its constituent atoms leads naturally to the idea of modeling a density by a sum of nuclear-centered, spherically symmetric functions. The functions that are optimal in a least-squares sense are known as Stewart atoms. Previous attempts to construct Stewart atoms by expanding them in an auxiliary basis have been thwarted by slow convergence with respect to the size of the auxiliary basis used. We present a method for constructing Stewart atoms via convolution integrals which bypasses the need for an auxiliary basis, and is able to produce highly accurate approximations to Stewart atoms. PMID- 15267704 TI - The role of symmetry and optical selection rules in revealing the molecular structure of the lowest Rydberg and ionic states of the 1,4 diazabicyclo[2.2.2]octane-Arn (n = 1,2,3) van der Waals complexes. AB - The 1,4-diazabicyclo[2.2.2]octane-Arn (n = 1,2,3) van der Waals complexes (DABCO Arn) have been investigated using a combination of (1 + 1') resonance enhanced multiphoton ionization (REMPI) and zero electron kinetic energy (ZEKE) spectroscopy. The additivity of the spectral shifts observed in both REMPI and ZEKE spectra, taken together with analysis of vibrational structure, suggest that in both DABCO-Ar and DABCO-Ar2 the argon atoms bind in equivalent equatorial (face) locations between two adjacent (CH2)2 bridges. However, the cumulative evidence from both REMPI and ZEKE spectra, together with ab initio results, suggests that the DABCO-Ar3 complex does not revert to D3h symmetry, but rather adopts a C2v structure in which all three argon atoms bind to one side of the DABCO framework. The exceptionally low wave-number vibrational structure observed in the REMPI spectra suggest that the van der Waals interaction in the excited state is extremely weak. However, ionization necessarily increases the strength of the interaction by virtue of the introduction of charge-induced dipole forces, as revealed by a consistent increase in vibrational wave numbers of the modes observed in the resultant ZEKE spectra. PMID- 15267705 TI - Two- and three-body photodissociation of gas phase I(3) (-). AB - The photodissociation dynamics of I3- from 390 to 290 nm (3.18 to 4.28 eV) have been investigated using fast beam photofragment translational spectroscopy in which the products are detected and analyzed with coincidence imaging. At photon energies < or = 3.87 eV, two-body dissociation that generates I- + I2(A 3Pi1) and vibrationally excited I2- (X 2Sigmau+) + I(2P(3/2)) is observed, while at energies > or = 3.87 eV, I*(2P(1/2)) + I2- (X 2Sigmau+) is the primary two-body dissociation channel. In addition, three-body dissociation yielding I- +2I(2P(3/2)) photofragments is seen throughout the energy range probed; this is the dominant channel at all but the lowest photon energy. Analysis of the three body dissociation events indicates that this channel results primarily from a synchronous concerted decay mechanism. PMID- 15267706 TI - Measurement of orientation and alignment moment relaxation by polarization spectroscopy: theory and experiment. AB - A diagrammatic perturbation theory description of one-color polarization spectroscopy (PS) is developed which emphasizes the significance of orientation and alignment tensor moments of the rotational angular momentum, and their collisional evolution. The influences of Doppler motion, velocity-changing collisions, decay of population, orientation and alignment, and nuclear hyperfine depolarization on the calculated PS signal are discussed. Illustrative simulations are presented of the evolution of the PS signal as a function of pump probe laser delay. These are generated by a Monte Carlo integration of the derived equations for the signal electric field over typical experimental pump and probe laser temporal profiles and velocity distributions for a commonly studied system, the OH A 2Sigma+ -X 2Pi (0,0) band. These predictions are compared with a preliminary set of results obtained in an experimental apparatus designed for one-color polarization spectroscopy using independent pump and probe lasers. Measurements are presented using linearly polarized pump light on the Q1(2.5) transition of the OH A 2Sigma+ -X 2Pi (0,0) band with He as the collision partner. The decay of the experimental PS pump-probe signal is discussed with reference to inelastic collisional population transfer rates in the literature. It is concluded that the collisional depolarization of rotational alignment is rapid, with a rate approximately twice that of population transfer. This is consistent with previous measurements in atmospheric pressure flames. PS is shown to be a viable novel spectroscopic method for determining rotational angular momentum orientation and alignment relaxation rates, which are valuable quantities because they are sensitive probes of the forces involved in inelastic collisions. PMID- 15267707 TI - Electronic properties of CrF and CrCl in the X 6Sigma+ state: observation of the halogen hyperfine structure by Fourier transform microwave spectroscopy. AB - The rotational spectra of the CrF and CrCl radicals in the X 6Sigma+ state were observed by employing a Fourier transform microwave spectrometer. The CrF and CrCl radicals were generated by the reaction of laser-ablated Cr with F2 and Cl2, respectively, diluted in Ar. A chromium rod made of chromium powder pasted with epoxy resin was ablated by a Nd:YAG laser. Rotational transitions were measured in the region between 8 and 26 GHz. Several hyperfine constants due to the halogen nuclei were determined by a least-squares analysis. The electronic properties of CrF and CrCl were derived from their hyperfine constants and were compared with those of other 3d transition metal monohalides: TiF, MnF, FeF, CoF, NiF, and FeCl. PMID- 15267708 TI - Electron momentum spectroscopy of CF2Cl2: experimental and theoretical momentum profiles for outer valence orbitals. AB - Electron momentum distributions for outer valence orbitals of CF2Cl2 have been obtained by (e,2e) electron momentum spectroscopy at an incident energy of 1200 eV + binding energy. The experimental electron momentum profiles are compared with Hartree-Fock and density functional theory (DFT) calculations using B3LYP hybrid functional with the 6-31G and 6-311+G* basis sets. Generally, the shapes of the experimental momentum profiles are well reproduced by DFT calculations using larger basis sets 6-311 + G*. An attempt has been made to clarify the ordering of the outer valence orbitals, which have been in controversy, by comparing experimental results with B3LYP/6-311 + G* calculations. PMID- 15267709 TI - Complete basis set extrapolated potential energy, dipole, and polarizability surfaces of alkali halide ion-neutral weakly avoided crossings with and without applied electric fields. AB - Complete basis set extrapolations of alkali halide (LiF, LiCl, NaF, NaCl) energy, dipole, and polarizability surfaces are performed with and without applied fields along the internuclear axis using state-averaged multireference configuration interaction. Comparison between properties (equilibrium separation, dissociation energy, crossing distance, diabatic coupling constant, dipole, and polarizability) derived from the extrapolated potential energy (or dipole) surfaces are made with those obtained from direct extrapolation from the basis set trends. The two extrapolation procedures are generally found to agree well for these systems. Crossing distances from this work are compared to those of previous work and values obtained from the Rittner potential. Complete basis set extrapolated crossing distances agree well with those derived from the Rittner potential for LiF, but were significantly larger for LiCl, NaF, and NaCl. The results presented here serve as an important set of benchmark data for the development of new-generation many-body force fields that are able to model charge transfer. PMID- 15267710 TI - Ground and excited states of NH4: Electron propagator and quantum defect analysis. AB - Vertical excitation energies of the Rydberg radical NH4 are inferred from ab initio electron propagator calculations on the electron affinities of NH4+. The adiabatic ionization energy of NH4 is evaluated with coupled-cluster calculations. These predictions provide optimal parameters for the molecular adapted quantum defect orbital method, which is used to determine Einstein emission coefficients and radiative lifetimes. Comparisons with spectroscopic data and previous calculations are discussed. PMID- 15267711 TI - Products of the addition of water molecules to Al3O3- clusters: structure, bonding, and electron binding energies in Al3O4H2-, Al3O5H4-, Al3O4H2, and Al3O5H4. AB - Two stable products of reactions of water molecules with the Al3O3- cluster, Al3O4H2- and Al3O5H4-, are studied with electronic structure calculations. There are several minima with similar energies for both anions and the corresponding molecules. Dissociative absorption of a water molecule to produce an anionic cluster with hydroxide ions is thermodynamically favored over the formation of Al3O3-(H2O)n complexes. Vertical electron detachment energies of Al3O4H2- and Al3O5H4- calculated with ab initio electron propagator methods provide a quantitative interpretation of recent anion photoelectron spectra. Contrasts and similarities in these spectra may be explained in terms of the Dyson orbitals associated with each transition energy. PMID- 15267712 TI - Theoretical prediction of electronic structures of fully pi-conjugated zinc oligoporphyrins with curved surface structures. AB - A theoretical prediction of the electronic structures of fully pi-conjugated zinc oligoporphyrins with curved surface, ring, tube, and ball-shaped structures was conducted as the objective for the future development of triply meso-meso-, beta beta-, and beta-beta-linked planar zinc oligoporphyrins. The excitation energies and oscillator strengths for the optimal ring and ball structures were calculated using the time-dependent density functional theory (DFT). Although there is an extremely small energy difference of < 0.1 eV between the highest occupied molecular orbital (HOMO) and the lowest unoccupied molecular orbital (LUMO) of the ring structure relative to the same-sized triply linked planar one, the Q and B bands of the former are smaller redshifted excitation energies and intensified oscillator strengths than those of the latter due to the structurally shortened effective pi-conjugated lengths for the electron transition. It is expected that the ball structure becomes an excellent electron acceptor and shows the highly redshifted Q' band in the near-IR region relative to the monomer. The minimum value of the HOMO-LUMO energy gaps of the infinite-length ring structures was estimated using periodic boundary conditions within the DFT, resulting in the metallic characters of both the tube structures with and without the spiral triply linked porphyrin array. The relation between the diameters and strain energies of the tube and ball structures was also examined. The present fused zinc porphyrins may become more colorful materials with new optelectronic properties including artificial photosynthesis than the carbon nanotubes and fullerenes when the axial coordinations of the central metal of porphyrins are functionally used. PMID- 15267713 TI - Quantum chemical density-functional theory calculations of the structures of defect C60 with four vacancies. AB - Quantum chemical density-functional theory (DFT) calculations have been carried out for the six isomers obtained by removing four adjacent atoms from C60. The most stable isomer consists of twelve 5-member and eighteen 6-member rings, indicating that the removal of some atoms from C60, which contains twelve 5 member rings and twenty 6-member rings, does not always generate larger holes. Each of the other five isomers contains at least one 4-member ring and one larger ring (7-, 8-, 9-, or 10-member ring) besides the 5- and 6-member rings. All isomers have similar structures for singlet and triplet spin multiplicities but with different stabilities. The ground states for two of the isomers are triplets, whereas the ground states for the other isomers are singlets. Furthermore, a comparison between the various isomers allowed one to examine the effect of the structure on the stability of fullerene cages. PMID- 15267714 TI - Exploring the dynamics of hydrogen atom release from the radical-radical reaction of O(3P) with C3H5. AB - The gas-phase radical-radical reaction dynamics of O(3P) + C3H5 --> H(2S) + C3H4O was studied at an average collision energy of 6.4 kcal/mol in a crossed beam configuration. The ground-state atomic oxygen [O(3P)] and allyl radicals (C3H5) were generated by the photolysis of NO2 and the supersonic flash pyrolysis of allyl iodide, respectively. Nascent hydrogen atom products were probed by the vacuum-ultraviolet-laser induced fluorescence spectroscopy in the Lyman-alpha region centered at 121.6 nm. With the aid of the CBS-QB3 level of ab initio theory, it has been found that the barrierless addition of O(3P) to C3H5 forms the energy-rich addition complexes on the lowest doublet potential energy surface, which are predicted to undergo a subsequent direct decomposition step leading to the reaction products H + C3H4O. The major counterpart C3H4O of the probed hydrogen atom is calculated to be acrolein after taking into account the factors of barrier height, reaction enthalpy, and the number of intermediates involved along the reaction pathway. The nascent H-atom Doppler profile analysis shows that the average center-of-mass translational energy of the H + C3H4O products and the fraction of the total available energy released as the translational energy were determined to be 3.83 kcal/mol and 0.054, respectively. On the basis of comparison with statistical calculations, the reaction proceeds through the formation of short-lived addition complexes rather than statistical, long-lived intermediates, and the polyatomic acrolein product is significantly internally excited at the moment of the decomposition. PMID- 15267715 TI - Formation of doubly positively charged diatomic ions of Mo2(2+) produced by Ar+ sputtering of an Mo metal surface. AB - Long-lived metastable doubly positively charged diatomic ions of Mo2(2+) have been produced by Ar+ bombardment of a molybdenum metal surface. These exotic molecular dications, such as for example 92,95Mo2(2+) at m/z 93.5, could be observed in positive ion mass spectra for ion flight times of approximately 17 micros in a Cameca IMS-3f secondary ion mass spectrometer, when the ion extraction field was adjusted for detection of ions that are formed in the gas phase several micrometers in front of the sputtered surface. Mo2(2+) was observed at high primary current densities for projectile ions of Ar+, but could not be detected under very similar bombarding conditions for projectile ions of Xe+. Such a dependence of ion production by inert gas sputtering on the primary ion species [ionization energies: IP1(Ar) = 15.76 eV and IP1(Xe) = 12.13 eV] is unusual. It is shown that formation of Mo2(2+) dications takes place by resonant charge transfer in grazing gas-phase collisions between incoming projectile ions of Ar+ and sputtered molecular ions of Mo2+. The efficiency for such a resonant electron capture (Mo2+ + Ar+ --> Mo2(2+) + Ar) is of the order of 10(-5) for the bombarding conditions in our mass spectrometer and corresponds to a cross section of a few 10(-15) cm2. PMID- 15267716 TI - Calculation of the transport properties of carbon dioxide. II. Thermal conductivity and thermomagnetic effects. AB - The transport properties of pure carbon dioxide have been calculated from the intermolecular potential using the classical trajectory method. Results are reported in the dilute-gas limit for thermal conductivity and thermomagnetic coefficients for temperatures ranging from 200 K to 1000 K. Three recent carbon dioxide potential energy hypersurfaces have been investigated. Since thermal conductivity is influenced by vibrational degrees of freedom, not included in the rigid-rotor classical trajectory calculation, a correction for vibration has also been employed. The calculations indicate that the second-order thermal conductivity corrections due to the angular momentum polarization (< 2%) and velocity polarization (< 1%) are both small. Thermal conductivity values calculated using the potential energy hypersurface by Bukowski et al. (1999) are in good agreement with the available experimental data. They underestimate the best experimental data at room temperature by 1% and in the range up to 470 K by 1%-3%, depending on the data source. Outside this range the calculated values, we believe, may be more reliable than the currently available experimental data. Our results are consistent with measurements of the thermomagnetic effect at 300 K only when the vibrational degrees of freedom are considered fully. This excellent agreement for these properties indicates that particularly the potential surface of Bukowski et al. provides a realistic description of the anisotropy of the surface. PMID- 15267717 TI - Positive electron affinity of fullerenes: Its effect and origin. AB - The universal variation pattern of the total energy of various fullerenes including single-walled carbon nanotubes with respect to their extra charge is revealed by the density-functional-theory calculations. The parabolic energy charge curve with its lowest energy value corresponding to a negatively charged fullerene indicates that these carbon materials have positive electron affinity and are not in the most stable state. The positive electron affinity seems to originate from the pi-electrons and is found to be related to the aggregation property of fullerenes. PMID- 15267718 TI - Quantum model simulations of symmetry breaking and control of bond selective dissociation of FHF- using IR+UV laser pulses. AB - Symmetry breaking and control of bond selective dissociation can be achieved by means of ultrashort few-cycle-infrared (IR) and ultraviolet (UV) laser pulses. The mechanism is demonstrated for the oriented model system, FHF-, by nuclear wave packets which are propagated on two-dimensional potential energy surfaces calculated at the QCISD/d-aug-cc-pVTZ level of theory. The IR laser pulse is optimized to drive the wave packet coherently along alternate bonds. Next, a well timed ultrashort UV laser pulse excites the wave packet, via photodetachment of the negative bihalide anion, to the bond selective domain of the neutral surface close to the transition state. The excited wave packet is then biased to evolve along the pre-excited bond toward the target product channel, rather than bifurcating in equal amounts. Comparison of the vibrational frequencies obtained within our model with harmonic and experimental frequencies indicates substantial anharmonicities and mode couplings which impose restrictions on the mechanism in the domain of ultrashort laser fields. Extended applications of the method to randomly oriented or to asymmetric systems XHY- are also discussed, implying the control of product directionality and competing bond-breaking. PMID- 15267719 TI - Mass analyzed threshold ionization spectroscopy of p-fluorostyrene. AB - Adiabatic ionization energy (AIE) and two-color threshold ion vibrational spectra of p-fluorostyrene have been measured by mass analyzed threshold ionization (MATI) method via three different intermediate levels in the first excited state, vibrationless S1 origin, 42(1)41(1), and 23(1) vibronic levels. Features of the ion vibrational spectra indicates that the geometry of the molecular ion including the conformation of the vinyl chain in the ionic ground state (D0) is almost identical to that of its neutral ground state (S0), and ionization has very little effect on the vibrational potentials of the aromatic ring modes. Comparison of the AIE with the reported value of styrene shows that fluorination at the para position of the aromatic ring has little effect on energy of the electron ejected in ionization process from the styrene chromophore. PMID- 15267720 TI - Ab initio study on structural and electronic properties of BanOm clusters. AB - Density-functional calculation within local density approximation, shows that the electronic property of a barium oxide cluster is strongly correlated with its equilibrium structure. The ground-state structures of BanOm (4 < or = n < or = 9,m < or = n) clusters can be classified into four categories: (a) compact, (b) dangling state, (c) F-center, and (d) stoichiometric. The compact cluster is metallic, almost no energy gap exists between the highest occupied and the lowest unoccupied molecular orbitals. The energy gap for the dangling state cluster is larger than that for the F-center cluster, while the stoichiometric cluster has the largest energy gap. PMID- 15267721 TI - Yield of electronically excited N2 molecules from the dissociative recombination of N2H+ with e-. AB - Quantitative spectroscopic observations of the N2 first positive band system (N2(B 3Pig-A 3Sigmau+))/electron in a recombining N2H+ flowing-afterglow plasma indicate that a substantial fraction of the product N2 molecules are formed in one or more of the low-lying triplet states, B 3Pig, A 3Sigmau+, and W 3Deltau. The total measured N2(B-A) emission intensity from N2(B,v' > or = 1) is equivalent to a yield of (19 +/- 8)%. The effect of rapid collision-induced transitions between states of the triplet manifold is discussed.. PMID- 15267722 TI - Mass transport following impulsive optical excitation. AB - Transition from reverse-saturable absorption to saturable absorption of the chloroaluminum phthalocyanine solution excited by a giant laser pulse is ascribed not just to the saturation of excited state absorption, but also to the outward migration of the solute molecules at the laser beam center. While the saturation of excited state absorption occurs within a single picosecond laser pulse, the beam center population decrease is sustained much longer than the pulse duration. We distinguish these two mechanisms with the Z-scan technique, utilizing picosecond pulses with pulse-to-pulse separations ranging from 0.1 to 5.0 s. PMID- 15267723 TI - Adapting the nudged elastic band method for determining minimum-energy paths of chemical reactions in enzymes. AB - Optimization of reaction paths for enzymatic systems is a challenging problem because such systems have a very large number of degrees of freedom and many of these degrees are flexible. To meet this challenge, an efficient, robust and general approach is presented based on the well-known nudged elastic band reaction path optimization method with the following extensions: (1) soft spectator degrees of freedom are excluded from path definitions by using only inter-atomic distances corresponding to forming/breaking bonds in a reaction; (2) a general transformation of the distances is defined to treat multistep reactions without knowing the partitioning of steps in advance; (3) a multistage strategy, in which path optimizations are carried out for reference systems with gradually decreasing rigidity, is developed to maximize the opportunity of obtaining continuously changing environments along the path. We demonstrate the applicability of the approach using the acylation reaction of type A beta lactamase as an example. The reaction mechanism investigated involves four elementary reaction steps, eight forming/breaking bonds. We obtained a continuous minimum energy path without any assumption on reaction coordinates, or on the possible sequence or the concertedness of chemical events. We expect our approach to have general applicability in the modeling of enzymatic reactions with quantum mechanical/molecular mechanical models. PMID- 15267724 TI - Fictive temperature, cooling rate, and viscosity of glasses. AB - The physical correlation between the fictive temperature dependence of the cooling rate of the melts and the temperature dependence of the equilibrium viscosity has been found by doing differential scanning calorimetric and viscometric measurements on a silicate melt, and by performing finite element simulations of the fiber drawing from that melt. This correlation is governed by a correlation factor Kc (in Pa K) which is constant and universal for silicate glasses. The factor Kc is obtained in the cooling rate range from 10(-2) to 10(6) K/s and is in good agreement with that theoretically predicted. The physical feature of the correlation is discussed in the paper. When the fictive temperature equals the actual temperature, a linear relation exists between the cooling rate and the Maxwell relaxation rate, the slope of which depends on the fragility of the glass melts. The Avramov equation is extended to describe the cooling rate dependence of the fictive temperature. The cooling rate equation contains only one adjusting parameter, i.e., the fragility parameter alpha. PMID- 15267725 TI - Equation of state and structural changes in diaminodinitroethylene under compression. AB - Structural changes in 1,1-diamino-2,2-dinitroethylene (DADNE, FOX-7) compressed to high pressure in diamond anvil cells were investigated using angle-dispersive x-ray diffraction analysis, Raman spectroscopy, and optical polarizing microscopy. The x-ray results show several changes above 1 GPa. When the x-ray data are indexed according to the ambient-pressure structure, DADNE shows anisotropic compression, with higher compression along the b axis than along the a or c axis. An ambient-temperature isothermal equation of state of DADNE was generated from these data. In addition, the experimentally obtained Raman spectra were matched with vibrational normal modes calculated using quantum chemistry calculations. The shifts in vibrational modes indicate changes in H-wagging vibrations with pressure. PMID- 15267726 TI - A pulse sequence for directly measuring the anharmonicities of coupled vibrations: Two-quantum two-dimensional infrared spectroscopy. AB - We have experimentally demonstrated a pulse sequence for the acquisition of heterodyned two-dimensional infrared (2D IR) spectra that correlates the overtone and combination bands to the fundamental frequencies. The spectra are generated by Fourier transforming the time domain signal that is allowed to evolve during one- and two-quantum coherence times. In this manner, the overtone and combination bands appear along the two-quantum axis, resulting in a direct determination of the diagonal and off-diagonal anharmonicities. To demonstrate this pulse sequence, we have collected two-quantum 2D IR spectra of a ruthenium dicarbonyl complex, extracted the diagonal and off-diagonal anharmonicities, and simulated the spectra using an exciton model. Several polarization conditions are presented that suppress the diagonal or cross peaks and we have used them to improve the accuracy of the measurement. PMID- 15267727 TI - Pattern formation in excitable media with concentration-dependent diffusivities. AB - We study a model of pattern formation in an excitable medium with concentration dependent diffusivities. The reaction terms correspond to a two-variable Gray Scott model in which the system has only one stable steady state. The diffusion coefficients of the two species are assumed to have a functional relationship with the concentration of the autocatalyst. A transition from self-replicating behavior to stationary spots is observed as the influence of the local autocatalyst concentration on the diffusion process increases. Notably, the transition occurs even though there is no change in the relative diffusivities of the activator and inhibitor. The observed time-independent patterns exhibit an unusual dependence on the size and geometry of an initial perturbation. Initial perturbations with a large spatial size, for example, sometimes revert to the homogeneous equilibrium state, whereas perturbations of smaller spatial extent develop into stable spots at the same parameter values. PMID- 15267728 TI - Collective dynamics in molten potassium: an inelastic x-ray scattering study. AB - The high-frequency collective dynamics of molten potassium has been investigated by inelastic x-ray scattering, disclosing an energy/momentum transfer region unreachable by previous inelastic neutron scattering (INS) experiments. We find that a two-step relaxation scenario, similar to that found in other liquid metals, applies to liquid potassium. In particular, we show how the sound velocity determined by INS experiments, exceeding the hydrodynamic value by approximately 30%, is the higher limit of a speedup, located in the momentum region 1 < Q < 3 nm(-1), which marks the departure from the isothermal value. We point out how this phenomenology is the consequence of a microscopic relaxation process that, in turn, can be traced back to the presence of "instantaneous" disorder, rather than to the crossover from a liquid to solidlike response. PMID- 15267729 TI - Activated radiationless decay of rhodamine-3B: nonequilibrium polarization effects in viscous solvents. AB - Nonequilibrium polarization effects that arise in high viscous polar solvents are discussed as regards to the rhodamine-3B-activated radiationless process. Rate constants are interpreted using dipole isomerization theories which enable the recovery of a barrier top region wave number identical to that previously obtained in less viscous solvents [J. Phys Chem. A 104, 11909 (2000)]. The Onsager-frequency-dependent reaction field can model the friction effect on the rate constants that in glycerol were estimated also from an adiabatic charge transfer model. The cusp barrier height is half the electronic coupling, as expected from the equality found for the frequencies of the reactant well and barrier top in this process. Coupling to solvent polarization modes can control the friction effect on the reactive mode. A two-dimensional reaction surface explains the photophysical features detected in the radiationless decay and a state energy diagram is proposed for rhodamine-3B. PMID- 15267730 TI - Combined electronic structure/molecular dynamics approach for ultrafast infrared spectroscopy of dilute HOD in liquid H2O and D2O. AB - We present a new approach that combines electronic structure methods and molecular dynamics simulations to investigate the infrared spectroscopy of condensed phase systems. This approach is applied to the OH stretch band of dilute HOD in liquid D2O and the OD stretch band of dilute HOD in liquid H2O for two commonly employed models of water, TIP4P and SPC/E. Ab initio OH and OD anharmonic transition frequencies are calculated for 100 HOD x (D2O)n and HOD x(H2O)n (n = 4-9) clusters randomly selected from liquid water simulations. A linear empirical relationship between the ab initio frequencies and the component of the electric field from the solvent along the bond of interest is developed. This relationship is used in a molecular dynamics simulation to compute frequency fluctuation time-correlation functions and infrared absorption line shapes. The normalized frequency fluctuation time-correlation functions are in good agreement with the results of previous theoretical approaches. Their long-time decay times are 0.5 ps for the TIP4P model and 0.9 ps for the SPC/E model, both of which appear to be somewhat too fast compared to recent experiments. The calculated line shapes are in good agreement with experiment, and improve upon the results of previous theoretical approaches. The methods presented are simple, and transferable to more complicated systems. PMID- 15267731 TI - Kinetics of Ca2+ complexation with some carbohydrates in aqueous solutions. AB - For solutions of four saccharides in water with alkaline-earth chlorides added ultrasonic attenuation spectra between 100 kHz and 2 GHz are reported and compared to those for carbohydrate solutions without salt. Calcium chloride does not alter the relaxation times in the spectra of D-glucose and D+-maltose solutions, reflecting the exocyclic hydroxymethyl group rotation, a saccharide saccharide association, and, with the disaccharide, also motions of both rings of a molecule relative to one another. The spectra of D-xylose and D-fructose solutions are substantially changed by the salts. With both saccharides an additional term with relaxation time around some nanoseconds exists which is assigned to a rearrangement of a carbohydrate-cation complex. Other relaxation terms of these saccharide solutions are also subject to noticeable changes by the salt, indicating specific carbohydrate-cation interactions. The ultrasonic spectra show that such interactions may exist also with carbohydrates which do not display the particular hydroxyl group sequences that are considered to promote complexation with cations. PMID- 15267732 TI - Simulations of time-dependent fluorescence in nano-confined solvents. AB - The time-dependent fluorescence of a model diatomic molecule with a charge transfer electronic transition in confined solvents has been simulated. The effect of confining the solvent is examined by comparing results for solutions contained within hydrophobic spherical cavities of varying size (radii of 10-20 angstroms). In previous work [J. Chem. Phys. 118, 6618 (2002)] it was found that the solute position in the cavity critically affects the absorption and fluorescence spectra and their dependence on cavity size. Here we examine the effect of cavity size on the time-dependent fluorescence, a common experimental probe of solvent dynamics. The present results confirm a prediction that motion of the solute in the cavity after excitation can be important in the time dependent fluorescence. The effects of solvent density are also considered. The results are discussed in the context of interpreting time-dependent fluorescence measurements of confined solvent systems. PMID- 15267733 TI - Comparison of kinetic Monte Carlo and molecular dynamics simulations of diffusion in a model glass former. AB - We present results from kinetic Monte Carlo (KMC) simulations of diffusion in a model glass former. We find that the diffusion constants obtained from KMC simulations have Arrhenius temperature dependence, while the correct behavior, obtained from molecular dynamics simulations, can be super-Arrhenius. We conclude that the discrepancy is due to undersampling of higher-lying local minima in the KMC runs. We suggest that the relevant connectivity of minima on the potential energy surface is proportional to the energy density of the local minima, which determines the "inherent structure entropy." The changing connectivity with potential energy may produce a correlation between dynamics and thermodynamics. PMID- 15267734 TI - Onset of decoherence: six-wave mixing measurements of vibrational decoherence on the excited electronic state of I2 in solid argon. AB - Pump-probe, four-wave, and six-wave mixing measurements of I2 isolated in solid argon are used to provide a clear experimental measure for the onset of vibrational quantum decoherence on the excited electronic state. The electronically resonant, six-wave mixing measurements bypass the rapid electronic dephasing, and measure the quantum cross-correlation between two packets launched on the B-state. The vibrational quantum coherence survives one period of motion, 400 fs, during which approximately 2000 cm(-1) of energy is transferred to the lattice. The decoherence occurs during the second cycle of motion, while classically coherent motion measured via pump-probe spectroscopy using the same electronic resonances continues for approximately 15 periods. This is contrasted with vibrational dephasing on the ground electronic surface, which lasts for 10(2) periods, as measured through time-resolved coherent anti-Stokes Raman scattering. The measurements and observables are discussed through time-circuit diagrams, and a mechanistic description of decoherence is derived through semiclassical analysis and simulations that reproduce the experiments. PMID- 15267735 TI - A path integral influence functional for excess electron in fluids: Density functional formulation. AB - In this paper, we propose a path integral influence functional from a solvent to determine a self-correlation function of a quantum particle in classical simple fluid. It is shown that the influence functional is related to a grand potential functional of the pure solvent under a three-dimensional external field arising from a classical isomorphic polymer, on which the quantum particle is mapped. The influence functional can be calculated from the self-correlation function, the solute-solvent and the solvent-solvent pair correlation function. The obtained equation of the self-correlation function is applied to an excess electron problem in fluid helium. The Fourier path-integral Monte Carlo method is employed to perform the path integral of the electron. The solute-solvent pair correlation function is estimated from a reference interaction site model integral equation. These results obtained form our proposed influence functional and from that proposed by Chandler, Singh, and Richardson are compared with those provided by a path integral Monte Carlo simulation with the explicit helium solvent. PMID- 15267736 TI - Rapid fluorescence quenching of S2-xanthione by 3,3-diethylpentane in perfluorohydrocarbons. AB - Rapid fluorescence quenching of S2-xanthione by 3,3-diethylpentane has been studied in three perfluorohydrocarbons of different viscosities. The donor fluorescence decay in the presence of a quencher was fitted using the Smoluchowski-Collins-Kimball (SCK) model. The molecular parameters, R (the sum of the molecular radii), D (the sum of diffusion coefficients), and the specific rate constant of the process, kappa, were determined. The values of parameter D for all systems studied differed from the sum of the macroscopic diffusion coefficients Dx measured independently. This behavior is explained by the dependence of the molecular diffusion coefficient (as determined from the SCK model) on the distance traveled by the donor and quencher molecules during the excited donor lifetime. PMID- 15267737 TI - Molecular dynamics simulations of transport and separation of carbon dioxide alkane mixtures in carbon nanopores. AB - The configurational-bias Monte Carlo method, which is used for efficient generation of molecular models of n-alkane chains, is combined for the first time with the dual control-volume grand-canonical molecular-dynamics simulation, which has been developed for studying transport of molecules in pores under an external potential gradient, to investigate transport and separation of binary mixtures of n-alkanes, as well as mixtures of CO2 and n-alkanes, in carbon nanopores. The effect of various factors, such as the temperature of the system, the composition of the mixture, and the pore size, on the separation of the mixtures is investigated. We also report the preliminary results of an experimental study of transport and separation of some of the same mixtures in a carbon molecular-sieve membrane with comparable pore sizes. The results indicate that, for the mixtures considered in this paper, even in very small carbon nanopores the energetic effects still play a dominant role in the transport and separation properties of the mixtures, whereas in a real membrane they are dominated by the membrane's morphological characteristics. As a result, for the mixtures considered, a single pore may be a grossly inadequate model of a real membrane, and hence one must resort to three-dimensional molecular pore network models of the membrane. PMID- 15267738 TI - Characterization of the molecular parameters determining charge transport in anthradithiophene. AB - The molecular parameters that govern charge transport in anthradithiophene (ADT) are studied by a joint experimental/theoretical approach involving high resolution gas-phase photoelectron spectroscopy and quantum-mechanical methods. The hole reorganization energy of ADT has been determined by an analysis of the vibrational structure of the lowest ionization band in the gas-phase photoelectron spectrum as well as by density-functional theory calculations. In addition, various dimers and clusters of ADT molecules have been considered in order to understand the effect of molecular packing on the hole and electron intermolecular transfer integrals. The results indicate that the intrinsic electronic structure, the relevant intramolecular vibrational modes, and the intermolecular interactions in ADT are very similar to those in pentacene. PMID- 15267739 TI - Dynamics of caged ions in glassy ionic conductors. AB - At sufficiently high frequency and low temperature, the dielectric responses of glassy, crystalline, and molten ionic conductors all invariably exhibit nearly constant loss. This ubiquitous characteristic occurs in the short-time regime when the ions are still caged, indicating that it could be a determining factor of the mobility of the ions in conduction at longer times. An improved understanding of its origin should benefit the research of ion conducting materials for portable energy source as well as the resolution of the fundamental problem of the dynamics of ions. We perform molecular dynamics simulations of glassy lithium metasilicate (Li2SiO3) and find that the length scales of the caged Li+ ions motions are distributed according to a Levy distribution that has a long tail. These results suggest that the nearly constant loss originates from "dynamic anharmonicity" experienced by the moving but caged Li+ ions and provided by the surrounding matrix atoms executing correlated movements. The results pave the way for rigorous treatments of caged ion dynamics by nonlinear Hamiltonian dynamics. PMID- 15267740 TI - Signatures of beta-sheet secondary structures in linear and two-dimensional infrared spectroscopy. AB - Using idealized models for parallel and antiparallel beta sheets, we calculate the linear and two-dimensional infrared spectra of the amide I vibration as a function of size and secondary structure. The model assumes transition-dipole coupling between the amide I oscillators in the sheet and accounts for the anharmonic nature of these oscillators. Using analytical and numerical methods, we show that the nature of the one-quantum vibrational eigenstates, which govern the linear spectrum, is, to a large extent, determined by the symmetry of the system and the relative magnitude of interstrand interactions. We also find that the eigenstates, in particular their trends with system size, depend sensitively on the secondary structure of the sheet. While in practice these differences may be difficult to distinguish in congested linear spectra, we demonstrate that they give rise to promising markers for secondary structure in the two-dimensional spectra. In particular, distinct differences occur between the spectra of parallel and antiparallel beta sheets and between beta hairpins and extended beta sheets. PMID- 15267741 TI - Atomic hydrogen interaction with Ru(1010). AB - The interaction of atomic hydrogen with clean and deuterium precovered Ru(1010) was studied by means of temperature-programmed desorption (TPD) spectroscopy. Compared to molecular hydrogen experiments, after exposure of the clean surface to gas-phase atomic hydrogen at 90 K, two additional peaks grow in the desorption spectra at 115 and 150 K. The surface saturation coverage, determined by equilibrium between abstraction and adsorption reactions, is 2.5 monolayers. Preadsorbed deuterium abstraction experiments with gas-phase atomic hydrogen show that a pure Eley-Rideal mechanism is not involved in the process, while a hot atom (HA) kinetics describes well the reaction. By least-squares fitting of the experimental data, a simplified HA kinetic model yields an abstraction cross section value of 0.5 +/- 0.2 angstroms2. The atomic hydrogen interaction with an oxygen precovered surface was also studied by means of both TPD and x-ray photoelectron spectroscopy: oxygen hydrogenation and water production take place already at very low temperature (90 K). PMID- 15267742 TI - Different adsorption structures of pyridine on Si(001) and Ge(001) surfaces. AB - The adsorption and reaction of pyridine on the Si(001) and Ge(001) surfaces are investigated by first-principles density-functional calculations within the generalized gradient approximation. On both surfaces the N atom of pyridine initially reacts with the down atom of the dimer, forming a single bond between the N atom and the down atom. On Ge(001) such an adsorption configuration is most favorable, but on Si(001) a further reaction with a neighboring dimer occurs, resulting in formation of a bridge-type configuration. Especially we find that on Ge(001) the bridge-type configuration is less stable than the gas phase. Our results provide an explanation for a subtle difference in the adsorption structures of pyridine on Si(001) and Ge(001), which was observed from recent scanning tunneling microscopy experiments. PMID- 15267743 TI - Dissociative photoionization of N(2)O in the region of the N(2)O(+)(B (2)Pi) state studied by ion-electron velocity vector correlation. AB - Dissociative direct photoionization of the N2O(X 1Sigma+) linear molecule via the N2O+(B 2Pi) ionic state induced by linearly polarized synchrotron radiation P in the 18-22 eV photon energy range is investigated using the (VA+,Ve,P) vector correlation method, where VA+ is the nascent velocity vector of the NO+, N2+, or O+ ionic fragment and Ve that of the photoelectron. The DPI processes are identified by the ion-electron kinetic energy correlation, and the IchiA+(thetae,phie) molecular frame photoelectron angular distributions (MFPADs) are reported for the dominant reaction leading to NO+ (X 1Sigma+,v) + N(2D)+ e. The measured MFPADs are found in satisfactory agreement with the reported multichannel Schwinger configuration interaction calculations, when bending of the N2O+(B 2Pi) molecular ion prior to dissociation is taken into account. A significant evolution of the electron scattering anisotropies is observed, in particular in the azimuthal dependence of the MFPADs, characteristic of a photoionization transition between a neutral state of Sigma symmetry and an ionic state of Pi symmetry. This interpretation is supported by a simple model describing the photoionization transition by the coherent superposition of two ssigma and ddelta partial waves and the associated Coulomb phases. PMID- 15267744 TI - Simplified crossover droplet model for adsorption of pure fluids in slit pores. AB - We present a generalized crossover (GC) model for the excess adsorption of pure fluids at a flat solid-liquid interface, which reproduces scaling behavior of the excess adsorption in the critical region and is reduced to the classical, van der Waals-type analytical model far away from the bulk critical point. In developing this model, we used the density-functional theory (DFT) approach for the order parameter profile calculations with a generalized corresponding states model for the local free-energy density. The GC DFT model well represents the available experimental adsorption data for Kr/graphite, C2H4/graphite, C3H8/graphite, CO2/silica, and SF6/graphite systems in the entire density range 0 < rho < or = 3rhoc and temperatures up to 1.7Tc. In the critical region 0.5 rhoc < r < or = 1.5rhoc and T < or = 1.15Tc, the GC DFT model is consistent with the predictions of the asymptotic renormalization-group crossover model for the critical adsorption in a semi-infinite system developed earlier. For the excess adsorption on the critical isochore, both theories predict a scaling-law behavior Gamma proportional tau(-nu+beta), but fail to reproduce a "critical depletion" of the excess adsorption along the critical isochore of the SF6/graphite system near Tc. We show that an anomalous decrease of adsorption observed in this system at tau = T/Tc - 1 < 10(-2) can be explained by finite-size effect and develop a simplified crossover droplet (SCD) model for the excess adsorption in a slit pore. With the effective size of the pore of L = 50 nm, the SCD model reproduces all available experimental data for SF6/graphite, including the critical isochore data where tau-->0, within experimental accuracy. At L >> xib (where xib is a bulk correlation length) the SCD model is transformed into the GC DFT model for semi infinite systems. Application of the SCD model to the excess adsorption of carbon dioxide on the silica gel is also discussed. PMID- 15267745 TI - Amplification of chirality in helical supramolecular polymers beyond the long chain limit. AB - The optical activity of helical homopolymers devoid of chiral centers increases drastically when a small amount of homochiral monomers is incorporated into them. We study this so-called sergeants-and-soldiers effect of chirality amplification in solutions of helical supramolecular polymers with a theoretical model that bears a strong resemblance to a one-dimensional, two-component Ising model. In the limit of very long self-assembled helical polymers, the strength of the sergeants-and-soldiers effect depends strongly on the free energy of a helix reversal and less so on the concentration of aggregating material. Outside the long-chain limit, we find the reverse--that is, a strong concentration dependence and a weak dependence on the helix-reversal energy. Our treatment gives an excellent agreement with recently published circular-dichroism measurements on mixed aggregates of discotic molecules in the solvents water and n-butanol, at two different overall concentrations. PMID- 15267746 TI - Mesoscopic model for diffusion-influenced reaction dynamics. AB - A hybrid mesoscopic multiparticle collision model is used to study diffusion influenced reaction kinetics. The mesoscopic particle dynamics conserves mass, momentum, and energy so that hydrodynamic effects are fully taken into account. Reactive and nonreactive interactions with catalytic solute particles are described by full molecular dynamics. Results are presented for large-scale, three-dimensional simulations to study the influence of diffusion on the rate constants of the A + C <==> B + C reaction. In the limit of a dilute solution of catalytic C particles, the simulation results are compared with diffusion equation approaches for both the irreversible and reversible reaction cases. Simulation results for systems where the volume fraction phi of catalytic spheres is high are also presented, and collective interactions among reactions on catalytic spheres that introduce volume fraction dependence in the rate constants are studied. PMID- 15267747 TI - Folding of small proteins using a single continuous potential. AB - Extensive Monte Carlo folding simulations for four proteins of various structural classes are carried out, using a single continuous potential (united-residue force field). In all cases, collapse occurs at a very early stage, and proteins fold into their nativelike conformations at appropriate temperatures. We also observe that glassy transitions occur at low temperatures. The simulation results demonstrate that the folding mechanism is controlled not only by thermodynamic factors but also by kinetic factors: The way a protein folds into its native structure is also determined by the convergence point of early folding trajectories, which cannot be obtained by the free energy surface. PMID- 15267748 TI - Relaxation processes in mixtures of liquid crystals and polymers near phase boundaries and during phase separation. AB - We present experimental studies of the relaxation of concentration fluctuations in a semidilute solution of polystyrene (PS) (30% by weight) in 4-cyano-4'-n octyl-biphenyl (8CB) (70% by weight) using the photon correlation spectroscopy (PCS). In the homogeneous phase there are two modes of relaxation. The slow one (typical time scale is taus = 0.001 s) is due to the diffusion of polymer chains (of molecular mass 65,000) in the LC matrix (of molecular mass 290), while the fast one has the time scale of the order of tauf approximately 0.00001 s. The amplitude of the fast mode is much weaker than the one for the slow mode. Moreover it does not depend on the scattering wave vector, q. The value of the diffusion coefficient, Dc = 1/(tausq2) for the slow mode decreases with temperature according to the Arhenius law until we reach the coexistence curve. Its value close to the coexistence is Dc = 4 x 10(5) nm2/s and the activation energy in the homogeneous mixture is Ec=127 kJ/mol. If we gradually undercool the mixture below the coexistence into the metastable two-phase region without inducing the phase separation we find unexpectedly that Dc does not change with temperature even 4 degrees below the coexistence curve. The characteristic time of the fast mode does not depend on the scattering wave vector indicating that it is related to the transient gel structure. We have shown that it is possible to measure the short time relaxation of concentration fluctuations during the phase separation in the mixture. At low temperature close to the isotropic-nematic phase transition we have observed that the relaxation is well separated in time from the typical time of the domain growth. This relaxation mode is characterized by the large diffusion coefficient D = 2 x 10(8) nm2/s. The mode probably comes from the coupling between the orientational dynamics of liquid crystals and the transient gel structure of polymers. PMID- 15267749 TI - Jamming concepts in cold polymeric glasses. AB - The purpose of the present paper is to propose an idea to construct the volume function W for polymer glasses and calculate the volume of polymer glasses as a function of the compactivity X utilizing a statistical-mechanical method developed for semiflexible polymers. We also discuss the analogue for glasses of "tapping" experiments which show the validity of statistical mechanics and the entropy concept in powders. PMID- 15267750 TI - A finite-size scaling study of a model of globular proteins. AB - Grand canonical Monte Carlo simulations are used to explore the metastable fluid fluid coexistence curve of the modified Lennard-Jones model of globular proteins of ten Wolde and Frenkel [Science, 277, 1975 (1997)]. Using both mixed-field finite-size scaling and histogram-reweighting methods, the joint distribution of density and energy fluctuations is analyzed at coexistence to accurately determine the critical-point parameters. The subcritical coexistence region is explored using the recently developed hyper parallel tempering Monte Carlo simulation method along with histogram reweighting to obtain the density distributions. The phase diagram for the metastable fluid-fluid coexistence curve is calculated in close proximity to the critical point, a region previously unattained by simulations. PMID- 15267751 TI - Bulk and interfacial properties of binary polymer mixtures. AB - A microscopic density functional theory is used to investigate a binary mixture of polymers, built of freely jointed tangent hard spheres. The difference in the chain length and in the segment diameter of polymers gives rise to a demixing transition. We evaluate the bulk fluid phase equilibria (binodal) and the limit of stability of a mixed state (spinodal) for selected systems, and analyze the decay of the critical packing fraction, critical mole fraction, and critical pressure with an increase of the chain length. The bulk results are subsequently used in the calculations of the density profiles across the fluid-fluid interface. The obtained profiles are smooth and do not exhibit any oscillations on the length scale of the segment diameter. Upon approaching the critical point the interfacial tension vanishes as (Deltarho)3, where Deltarho is the difference between bulk densities of one component in bulk phases rich and poor in that species. This indicates that the microscopic density functional theory applied here is of a mean-field type. PMID- 15267752 TI - Thermodynamics of beta-amyloid fibril formation. AB - Amyloid fibers are aggregates of proteins. They are built out of a peptide called beta-amyloid (Abeta) containing between 41 and 43 residues, produced by the action of an enzyme which cleaves a much larger protein known as the amyloid precursor protein (APP). X-ray diffraction experiments have shown that these fibrils are rich in beta-structures, whereas the shape of the peptide displays an alpha-helix structure within the APP in its biologically active conformation. A realistic model of fibril formation is developed based on the 17 residues Abeta12 28 amyloid peptide, which has been shown to form fibrils structurally similar to those of the whole Abeta peptide. With the help of physical arguments and in keeping with experimental findings, the Abeta12-28 monomer is assumed to be in four possible states (i.e., native helix conformation, beta-hairpin, globular low energy state, and unfolded state). Making use of these monomeric states, oligomers (dimers, tertramers, and octamers) were constructed. With the help of short, detailed molecular dynamics calculations of the three monomers and of a variety of oligomers, energies for these structures were obtained. Making use of these results within the framework of a simple yet realistic model to describe the entropic terms associated with the variety of amyloid conformations, a phase diagram can be calculated of the whole many-body system, leading to a thermodynamical picture in overall agreement with the experimental findings. In particular, the existence of micellar metastable states seem to be a key issue to determine the thermodynamical properties of the system. PMID- 15267753 TI - Crystal nucleation for a model of globular proteins. AB - A continuum model of globular proteins proposed by Talanquer and Oxtoby [J. Chem. Phys. 109, 223 (1998)] is investigated numerically, with particular emphasis on the region near the metastable fluid-fluid coexistence curve. Classical nucleation theory is shown to be invalid not only in the vicinity of the metastable critical point but also close to the liquidus line. An approximate analytic solution is also presented for the shape and properties of the nucleating crystal droplet. PMID- 15267754 TI - Density and energy relaxation in an open one-dimensional system. AB - A new master equation to mimic the dynamics of a collection of interacting random walkers in an open system is proposed and solved numerically. In this model, the random walkers interact through excluded volume interaction (single-file system); and the total number of walkers in the lattice can fluctuate because of exchange with a bath. In addition, the movement of the random walkers is biased by an external perturbation. Two models for the latter are considered: (1) an inverse potential (V proportional, variant 1/r), where r is the distance between the center of the perturbation and the random walker and (2) an inverse of sixth power potential (V proportional, 1/r6). The calculated density of the walkers and the total energy show interesting dynamics. When the size of the system is comparable to the range of the perturbing field, the energy relaxation is found to be highly nonexponential. In this range, the system can show stretched exponential (e-(t/taus)beta) and even logarithmic time dependence of energy relaxation over a limited range of time. Introduction of density exchange in the lattice markedly weakens this nonexponentiality of the relaxation function, irrespective of the nature of perturbation. PMID- 15267755 TI - Comment on "Group-theoretical analysis of the electronic structure data for molecular ions CN+/-60 (Ih) derived from multipole expansion of the Coulomb interelectronic interactions" [J. Chem. Phys. 119, 11429 (2003)]. PMID- 15267756 TI - The vibrational Stokes shift of water (HOD in D2O). AB - The vibrational Stokes shift of the OH stretching transition nu(OH) of water is the shift between the ground-state absorption and the excited-state (v=1) emission. A recent measurement on HOD in D(2)O solvent [S. Woutersen and H. J. Bakker, Phys. Rev. Lett. 83, 2077 (1999)] of a 70 cm(-1) redshift, and a subsequent calculation of a 57 cm(-1) redshift using equilibrium molecular dynamics simulations [C. P. Lawrence and J. L. Skinner, J. Chem. Phys. 117, 8847 (2002)] were in good agreement. We now report extensive measurements of the vibrational Stokes shift in HOD/D(2)O using an ultrafast IR pump, Raman probe method. The vibrational Stokes shift is seen to depend on the pump pulse frequency and on time delay; by varying these parameters it can be made to range from 112 to -32 cm(-1) (negative values indicate a blueshift in the excited state). The equilibrium vibrational Stokes shift is actually a negative rather than a positive quantity. Possible reasons for the disagreement between experiment and theory are briefly discussed. PMID- 15267757 TI - Broadband rotational resonance in solid state NMR spectroscopy. AB - A new technique for restoring nuclear magnetic dipole-dipole couplings under magic-angle spinning (MAS) in solid state nuclear magnetic resonance (NMR) spectroscopy is described and demonstrated. In this technique, called broadband rotational resonance (BroBaRR), the coupling between a pair of nuclear spins with NMR frequency difference close (but not necessarily equal) to the MAS frequency is restored by the application of a train of weak radio-frequency pulses at a carrier frequency close to the average of the two NMR frequencies. Phase or amplitude modulation of the pulse train at half the MAS frequency splits the carrier into sidebands close to the two NMR frequencies. The pulse train then removes offsets from the exact rotational resonance condition, leading to dipolar recoupling over a bandwidth controlled by the amplitude of the pulse train. (13)C NMR experiments on uniformly (15)N,(13)C-labeled L-valineHClH(2)O powder validate the theoretical analysis. BroBaRR will be useful in studies of molecular structures by solid state NMR, for example in the detection of long-range couplings between carbons in uniformly labeled organic and biological materials. PMID- 15267758 TI - Excitation energies for a benchmark set of molecules obtained within time dependent current-density functional theory using the Vignale-Kohn functional. AB - In this article we explain how the existing linear response theory of time dependent density-functional theory can be extended to obtain excitation energies in the framework of time-dependent current-density-functional theory. We use the Vignale-Kohn current-functional [G. Vignale and W. Kohn, Phys. Rev. Lett. 77, 2037 (1996)] which has proven to be successful for describing ultranonlocal exchange-correlation effects in the case of the axial polarizability of molecular chains [M. van Faassen, P. L. de Boeij, R. van Leeuwen, J. A. Berger, and J. G. Snijders, Phys. Rev. Lett. 88, 186401 (2002); J. Chem. Phys. 118, 1044 (2003)]. We study a variety of singlet excitations for a benchmark set of molecules. The pi(*)<--pi transitions obtained with the Vignale-Kohn functional are in good agreement with experiment and other theoretical results and they are in general an improvement upon the adiabatic local density approximation. In case of the pi(*)<--n transitions the Vignale-Kohn functional fails, giving results that strongly overestimate the experimental and other theoretical results. The benchmark set also contains some other types of excitations for which no clear failures or improvements are observed. PMID- 15267759 TI - The spin-unrestricted molecular Kohn-Sham solution and the analogue of Koopmans's theorem for open-shell molecules. AB - Spin-unrestricted Kohn-Sham (KS) solutions are constructed from accurate ab initio spin densities for the prototype doublet molecules NO(2), ClO(2), and NF(2) with the iterative local updating procedure of van Leeuwen and Baerends (LB). A qualitative justification of the LB procedure is given with a "strong" form of the Hohenberg-Kohn theorem. The calculated energies epsilon(isigma) of the occupied KS spin orbitals provide numerical support to the analogue of Koopmans' theorem in spin-density functional theory. In particular, the energies epsilon(ibeta) of the minor spin (beta) valence orbitals of the considered doublet molecules correspond fairly well to the experimental vertical ionization potentials (VIPs) I(i) (1) to the triplet cationic states. The energy epsilon(Halpha) of the highest occupied (spin-unpaired) alpha orbital is equal to the first VIP I(H) (0) to the singlet cationic state. In turn, the energies epsilon(ialpha) of the major spin (alpha) valence orbitals of the closed subshells correspond to a fifty-fifty average of the experimental VIPs I(i) (1) and I(i) (0) to the triplet and singlet states. For the Li atom we find that the exact spin densities are represented by a spin-polarized Kohn-Sham system which is not in its ground state, i.e., the orbital energy of the lowest unoccupied beta spin orbital is lower than that of the highest occupied alpha spin orbital ("a hole below the Fermi level"). The addition of a magnetic field in the -z direction will shift the beta levels up so as to restore the Aufbau principle. This is an example of the nonuniqueness of the mapping of the spin density on the KS spin-dependent potentials discussed recently in the literature. The KS potentials may no longer go to zero at infinity, and it is in general the differences nu(ssigma)( infinity )-epsilon(isigma) that can be interpreted as (averages of) ionization energies. In total, the present results suggest the spin unrestricted KS theory as a natural one-electron independent-particle model for interpretation and assignment of the experimental photoelectron spectra of open shell molecules. PMID- 15267760 TI - Disentangling multidimensional femtosecond spectra of excitons by pulse shaping with coherent control. AB - Sequences of carefully timed and shaped optical pulses provide femtosecond snapshots of molecular structure as well as electronic and vibrational dynamical processes, in analogy with multidimensional NMR. We apply a genetic learning algorithm towards the design of pulse sequences which simplify the multidimensional signals by controlling the relative intensities of various peaks. Numerical simulations demonstrate how poorly resolved weak features may be amplified and observed by using optimized optical pulses, specifically shaped to achieve a desired spectroscopic target. PMID- 15267761 TI - Technique for incorporating the density functional Hessian into the geometry optimization of biomolecules, solvated molecules, and large floppy molecules. AB - Traditional geometry optimization methods require the gradient of the potential surface, together with a Hessian which is often approximated. Approximation of the Hessian causes difficulties for large, floppy molecules, increasing the number of steps required to reach the minimum. In this article, the costly evaluation of the exact Hessian is avoided by expanding the density functional to second order in both the nuclear and electronic variables, and then searching for the minimum of the quadratic functional. The quadratic search involves the simultaneous determination of both the geometry step and the associated change in the electron density matrix. Trial calculations on Taxol indicate that the cost of the quadratic search is comparable to the cost of the density functional energy plus gradient. While this procedure circumvents the bottleneck coupled perturbed step in the evaluation of the full Hessian, the second derivatives of the electron-repulsion integrals are still required for atomic-orbital-based calculations, and they are presently more expensive than the energy plus gradient. Hence, we anticipate that the quadratic optimizer will initially find application in fields in which existing optimizers breakdown or are inefficient, particularly biochemistry and solvation chemistry. PMID- 15267762 TI - Complex dynamics in a periodically perturbed electro-chemical system. AB - Dynamical response of a passivation model subjected to parametric periodic and stochastic perturbations is studied numerically. In response to weak periodic modulation, the system exhibits a rich variety of resonance behavior and induced dynamics, including periodically induced oscillation, birhythmicity, switching between two bistable states, selection of one of the bistable states, mixed-mode and chaotic oscillations. These phenomena are discussed in terms of the stability of saddle focus and an incomplete homoclinic connection. Our numerical results are relevant for a wide class of electro-chemical oscillatory systems, where the re-injection of unstable trajectory on the neighborhood of a saddle focus is a typical feature in the phase space. PMID- 15267763 TI - Independent particle theory with electron correlation. AB - We formulate an effective independent particle model where the effective Hamiltonian is composed of the Fock operator and a correlation potential. Within the model the kinetic energy and the exchange energy can be expressed exactly leaving the correlation energy functional as the remaining unknown. Our efforts concentrate on finding a correlation potential such that exact ionization potentials and electron affinities can be reproduced as orbital energies. The equation-of-motion coupled-cluster approach enables us to define an effective Hamiltonian from which a correlation potential can be extracted. We also make the connection to electron propagator theory. The disadvantage of the latter is the inherit energy dependence of the potential resulting in a different Hamiltonian for each orbital. Alternatively, the Fock space coupled-cluster approach employs an effective Hamiltonian which is energy independent and universal for all orbitals. A correlation potential is extracted which yields the exact ionization potentials and electron affinities and a set of associated molecular orbitals. We also describe the close relationship to Brueckner theory. PMID- 15267764 TI - Full configuration interaction calculation of Be3. AB - The full configuration interaction (FCI) study of the ground state of the neutral beryllium trimer has been performed using an atomic natural orbitals [3s2p1d] basis set. Both triangular and linear structures have been considered for the Be(3) cluster. The optimal geometry for the equilateral triangle has been calculated. The potential energy cut sections along the normal a(1)(') mode and one of the components of the e(') mode have then been studied. The FCI symmetric atomization potential of the linear cluster is also reported. It shows a secondary van der Waals minimum at a long bond distance. All singular points in the potential energy curves are characterized. Other properties, like dissociation energies D(e) and vibrational frequencies, have been estimated from a fourth-order fitting of a large range of points around the minima. The calculated FCI wave number values for the nu(1) and nu(2) normal modes are (467.33+/-0.43) cm(-1) and (390.77+/-0.56) cm(-1). PMID- 15267765 TI - Finite-size scaling for critical conditions for stable quadrupole-bound anions. AB - We present finite-size scaling calculations of the critical parameters for binding an electron to a finite linear quadrupole field. This approach gives very accurate results for the critical parameters by using a systematic expansion in a finite basis set. The model Hamiltonian consists of a charge Q located at the origin of the coordinates and k charges -Q/k located at distances R(i), i=1, em leader,k. After proper scaling of distances and energies, the rescaled Hamiltonian depends only on one free parameter q=QR. Two different linear charge configurations with q>0 and q<0 are studied using basis sets in both spherical and prolate spheroidal coordinates. For the case with q>0, the finite size scaling calculations give an extrapolated critical value of q(c)=1.469 70+/-0.000 05 a.u. by using a basis set with prolate spheroidal coordinates. For the quadrupole case with q<0, we obtained an extrapolated critical value of mid R:q(c)mid R:=3.982 51+/-0.000 01 a.u. for stable quadrupole bound anions. The corresponding critical exponent for the ground state energy alpha=1.9964+/ 0.0005, with E approximately (q-q(c))(alpha). PMID- 15267766 TI - Derivation of the electronic nonadiabatic coupling field in molecular systems: an algebraic-vectorial approach. AB - In this Communication it is suggested that various elements of the nonadiabatic coupling matrix, tau(jk)(s) are created by the singular nonadiabatic coupling terms of the system. Moreover, given the spatial distribution of these coupling terms in the close vicinity of their singularity points yields, according to this approach, the integrated intensity of the field at every point in the region of interest. To support these statements we consider the conical intersections of the three lower states of the H+H(2) system: From an ab initio treatment we obtain the nonadiabatic coupling terms around each conical intersection separately (at its close vicinity) and having those, create the field at every desired point employing vector-algebra. This approach is also used to calculate the intensity of the Curl of those matrix elements that lack their own sources [tau(13)(s) in the present case]. The final results are compared with relevant ab initio calculations. PMID- 15267767 TI - A long-range-corrected time-dependent density functional theory. AB - We apply the long-range correction (LC) scheme for exchange functionals of density functional theory to time-dependent density functional theory (TDDFT) and examine its efficiency in dealing with the serious problems of TDDFT, i.e., the underestimations of Rydberg excitation energies, oscillator strengths, and charge transfer excitation energies. By calculating vertical excitation energies of typical molecules, it was found that LC-TDDFT gives accurate excitation energies, within an error of 0.5 eV, and reasonable oscillator strengths, while TDDFT employing a pure functional provides 1.5 eV lower excitation energies and two orders of magnitude lower oscillator strengths for the Rydberg excitations. It was also found that LC-TDDFT clearly reproduces the correct asymptotic behavior of the charge-transfer excitation energy of ethylene-tetrafluoroethylene dimer for the long intramolecular distance, unlike a conventional far-nucleus asymptotic correction scheme. It is, therefore, presumed that poor TDDFT results for pure functionals may be due to their lack of a long-range orbital-orbital interaction. PMID- 15267768 TI - Spectrally resolved femtosecond two-color three-pulse photon echoes: study of ground and excited state dynamics in molecules. AB - We report the use of spectrally resolved femtosecond two-color three-pulse photon echoes as a potentially powerful multidimensional technique for studying vibrational and electronic dynamics in complex molecules. The wavelengths of the pump and probe laser pulses are found to have a dramatic effect on the spectrum of the photon echo signal and can be chosen to select different sets of energy levels in the vibrational manifold, allowing a study of the dynamics and vibrational splitting in either the ground or the excited state. The technique is applied to studies of the dynamics of vibrational electronic states in the dye molecule Rhodamine 101 in methanol. PMID- 15267769 TI - Intracluster stereochemistry in van der Waals complexes: steric effects in ultraviolet photodissociation of state-selected Ar-HOD/H2O. AB - High-resolution IR-UV multiple resonance methods are employed to elucidate the photodissociation dynamics of quantum state-selected Ar-HOD and Ar-H(2)O van der Waals clusters. A single mode pulsed OPO operating in the region of the OH second overtone is used to prepare individual rovibrational states that are selectively photodissociated at specific excimer wavelengths. Subsequent fluorescence excitation of the resulting OH (OD) fragments yields dynamical information on the photofragmentation event and any resulting intracluster collisions. This technique is used to characterize spectroscopically the Pi(1(01)), nu(OH)=3<- Sigma(0(00)), v(OH)=0 overtone band of the Ar-HOD complex with an origin at 10648.27 cm(-1). The effects of Ar complexation on the dissociation dynamics are inferred by comparison of the OD photofragment quantum state distributions resulting from dissociation of single rovibrational states of the complex with those from isolated HOD photodissociation. The important role played by the initial internal state of the complex is demonstrated by comparison of the current Ar-HOD data with previously published results for the Ar-H(2)O Sigma(0(00))[03(-)> state. We interpret the dramatic differences in the dynamics of the two systems as manifestations of the nodal structure of the vibrational state in the parent complex and the way in which it governs the collision probability between the Ar atom and the escaping photofragments. PMID- 15267770 TI - Hydrogen bonds in 1,4-dioxane/ammonia binary clusters. AB - With synchrotron radiation, we have studied the photoionization and dissociation of 1,4-dioxane/ammonia clusters in a supersonic expansion. The observed major product ions are the 1,4-dioxane cation M(+) and protonated cluster ions M(NH(3))(n)H(+) (where M=1,4-dioxane), and the intensities of the unprotonated cluster ions M(NH(3))(n) (+) are much lower. Fully optimized geometries and energies of the neutral cluster M(NH(3))(2) and related cluster ions have been obtained using the ab initio molecular orbital method and density functional theory. The potential energy surface of the excited state of M(NH(3))(2) (+) was also calculated. With these results, the mechanisms of different photoionization dissociation channels have been suggested. The most probable channel is electron ejection from the highest occupied molecular orbital, followed by the dissociation into M(+) and (NH(3))(2). For another main channel, after removing an electron from the second highest occupied molecular orbital, the intracluster proton transfer process takes place to form the stable unprotonated cluster ion M(NH(3))H(+)-NH(2), which usually leads to the dissociated protonated cluster ion M(NH(3))H(+) and a radical NH(2). PMID- 15267771 TI - First-principles study of the electronic structures of icosahedral TiN (N=13,19,43,55) clusters. AB - We have studied the electronic structures of icosahedral Ti(N) clusters (N=13, 19, 43, and 55) by using a real-space first-principles cluster method with generalized gradient approximation for exchange-correlation potential. The hexagonal close-packed and fcc close-packed clusters have been studied additionally for comparisons. It is found that the icosahedral structures are the most stable ones except for Ti(43), where fcc close-packed structure is favorable in energy. We present and discuss the variation of bond length, the features of the highest occupied molecular orbitals and the lowest unoccupied molecular orbital, the evolution of density of states, and the magnetic moment in detail. The results are in good agreement with the predictions from the collision-induced dissociation and size-selected anion photoelectron spectroscopy experiments. PMID- 15267772 TI - Heuristic overlap-exchange model of noble gas chemical shifts. AB - It is now generally recognized that overlap-exchange interactions are the primary cause of the medium-dependent magnetic shielding (chemical shift) in all noble gases except helium, although the attractive electrostatic-dispersion (van der Waals) interactions play an indirect role in determining the penetration of the interacting species into the repulsive overlap-exchange region. The short-range nature of these overlap-exchange interactions, combined with the fact that they often can be approximated by simple functions of the overlap of the wave functions of the interacting species, suggests a useful semiempirical model of these chemical shifts. In it the total shielding is the sum of shieldings due to pairwise interactions of the noble gas atom with the individual atoms of the medium, with the "atomic" shielding terms either estimated by simple functions of the atomic overlap integrals averaged over their Boltzmann-weighted separations, or determined by fits to experimental data in systems whose complexity makes the former procedure impractical. Results for (129)Xe chemical shifts in the noble gases and in a variety of molecular and condensed systems, including families of n-alkanes, straight-chain alcohols, and the endohedral compounds Xe@C(60) and Xe@C(70) are encouraging for the applicability of the model to systems of technical and biomedical interest. PMID- 15267773 TI - Vacuum ultraviolet pulsed field ionization study of ND3: accurate thermochemistry for the ND2-ND2+ and ND3-ND3+ system. AB - The dissociation of energy-selected ND(3) (+) to form ND(2) (+)+D near its threshold has been investigated using the pulsed field ionization-photoelectron (PFI-PE)-photoion coincidence method. The breakdown curves for ND(3) (+) and ND(2) (+) give a value of 15.891+/-0.001 eV for the 0 K dissociation threshold or appearance energy (AE) for ND(2) (+) from ND(3). We have also measured the PFI-PE vibrational bands for ND(3) (+)(X;v(2) (+)=0, 1, 2, and 3), revealing partially resolved rotational structures. The simulation of these bands yields precise ionization energies (IEs) for ND(3) (+) X(0,v(2) (+)=0-3,0,0)<--ND(3) X(0,0,0,0). Using the 0 K AE (ND(2) (+)) and IE(ND(3))=10.200+/-0.001 eV determined in the present study, together with the known 0 K bond dissociation energy for ND(3) [D(0)(D-ND(2))=4.7126+/-0.0025 eV], we have determined the D(0)(ND(2) (+)-D), IE(ND(2)), and 0 K heat of formation for ND(2) (+) to be 5.691+/-0.001 eV, 11.1784+/-0.0025 eV, and 1261.82+/-0.4 kJ/mol, respectively. The PFI-PE spectrum is found to exhibit a steplike feature near the AE(ND(2) (+)), indicating that the dissociation of excited ND(3) (+) at energies slightly above the dissociation threshold is prompt, occurring in the time scale He+H2+ +e-. AB - Relative doubly differential cross sections for the Penning ionization of H(2) by spin-state-selected metastable He (1s2s) are reported at center-of-mass collision energies E of 3.1 and 4.2 kcal/mol in a crossed supersonic beam experiment employing a rotatable mass spectrometer detector. The measurements are sufficiently dense in velocity space as to avoid having to functionalize the differential cross sections in order to transform the intensities into the c.m. The H(2) (+) product is scattered sharply forward, c.m. Deltatheta<10 degrees half-width at half-maximum, with respect to the incident direction of H(2) at both energies for both spin states. On the average the products have lost energy upon recoil, mean recoil energy E(') C in solutions. AB - Non-Markovian kinetic equations of the reversible monomolecular-bimolecular reactions of the type A+B right arrow over left arrow C (at arbitrary ratio between A and B concentrations) derived earlier are used in the calculation of kinetics on macroscopic space-time scales. It is found that the kinetics of the systems with different structure of reactants is universal, and it is the direct generalization of the kinetic law of mass action of formal chemical kinetics. The analysis of the kinetics allows one to establish the time range of the applicability of the law of mass action. It is shown that beyond these limits the usual kinetic law of mass action becomes invalid, and correct description of the kinetics even in the most rough approximation calls for the non-Markovian corrections to usual kinetic laws. PMID- 15267796 TI - Polymorphism in simple liquids: a Gibbs ensemble Monte Carlo study. AB - We perform Gibbs ensemble Monte Carlo (GEMC) simulations of a one-component system of hard spheres with a repulsive shoulder and an attractive well. We show the existence of two distinct liquid-gas and liquid-liquid phase equilibria. The GEMC estimate of the critical parameters, as following from an interpolation of the binodal points, is only slightly influenced by finite size effects. The liquid-gas critical temperature and pressure are lower than those of the liquid liquid phase separation. A discussion of our findings in comparison with those of previous numerical studies is also presented. PMID- 15267797 TI - Thermal conductivity of molten alkali halides from equilibrium molecular dynamics simulations. AB - The thermal conductivity of molten sodium chloride and potassium chloride has been computed through equilibrium molecular dynamics Green-Kubo simulations in the microcanonical ensemble (N,V,E). In order to access the temperature dependence of the thermal conductivity coefficient of these materials, the simulations were performed at five different state points. The form of the microscopic energy flux for ionic systems whose Coulombic interactions are calculated through the Ewald method is discussed in detail and an efficient formula is used by analogy with the methods used to evaluate the stress tensor in Coulombic systems. The results show that the Born-Mayer-Huggins-Tosi-Fumi potential predicts a weak negative temperature dependence for the thermal conductivity of NaCl and KCl. The simulation results are in agreement with part of the experimental data available in the literature with simulation values generally overpredicting the thermal conductivity by 10%-20%. PMID- 15267798 TI - Electron solvation by highly polar molecules: density functional theory study of atomic sodium interaction with water, ammonia, and methanol. AB - This study further extends the scope of a previous paper [Y. Ferro and A. Allouche, J. Chem. Phys. 118, 10461 (2003)] on the reactivity of atomic Na with water to some other highly polar molecules known for their solvation properties connected to efficient hydrogen bonding. The solvation mechanisms of ammonia and methanol are compared to the hydration mechanism. It is shown that in the case of ammonia, the stability of the solvated system is only ensured by electrostatic interactions, whereas the methanol action is more similar to that of water. More specific attention is given to the solvation process of the valence 3s Na electron. The consequences on the chemical reactivity are analyzed: Whereas ammonia is nonreactive when interacting with atomic sodium, two chemical reactions are proposed for methanol. The first process is dehydrogenation and yields methoxy species and hydrogen. The other one is dehydration and the final products are methoxy species, but also methyl radical and water. The respective roles of electron solvation and hydrogen bonds network are analyzed in detail in view of the density of states of the reactive systems. PMID- 15267799 TI - Electron solvation by polar molecules: the interaction of Na atoms with solid methanol films studied with MIES and density functional theory calculations. AB - The interaction of Na atoms with CH(3)OH films was studied with metastable impact electron spectroscopy (MIES) under UHV conditions. The films were grown at 90(+/ 10) K on tungsten substrates and exposed to Na. Na-induced formation of methoxy (CH(3)O) species takes place, and Na atoms become ionized. At small Na exposures the outermost solvent layer remains largely intact as concluded from the absence of MIES signals caused by the reaction products. However, emission from CH(3)O, located at the film surface, occurs at larger exposures. In the same exposure range also Na species can be detected at the surface. The spectral feature from 3s Na ionization occurs at an energetic position different from that found for metals or semiconductors. The results are compared with density functional theory calculations [see Y. Ferro, A. Allouche, and V. Kempter, J. Chem. Phys. 120, 8683 (2004), preceding paper]. Experiment and theory agree in the energetic positions of the main spectral features from the methanol and sodium ionization. The calculations suggest that the 3s Na emission observed experimentally originates from solvated 3s electrons which are located far from the Na core and become stabilized by solvent molecules. The simultaneous emergence of emission from CH(3)O and from solvated 3s electrons suggests that the delocalization and, consequently, the solvation play an important role in the Na-induced formation of CH(3)O from CH(3)OH. PMID- 15267800 TI - Reactivity of Al3O3- cluster toward H2O studied by density functional theory. AB - Density functional theory calculations (Becke's three parameter hybrid functional) have been done on a wide range of possible structures for the complexes formed in the reaction between Al(3)O(3) (-) and one or two water molecules. Both energetically competitive structural isomers of Al(3)O(3) (-) (kitelike and distorted rectangle) were considered. The structures of neutral complexes accessed from detachment of the stable anion structures were also optimized. The calculations predict that hydroxide complexes are energetically favored over Lewis acid-base and charge-dipole complexes. For Al(3)O(3) (-)/H(2)O complexes, the kite-based hydroxide and rectangle-based hydroxide are predicted to be nearly isoenergetic, while for Al(3)O(3) (-)/(H(2)O)(2), the rectangle based dihydroxide emerges as being 0.5 eV more stable than the lowest energy kite based dihydroxide. The structures of these and their neutrals are used to analyze anion PE spectra of Al(3)O(4)H(2) (-) and Al(3)O(5)H(4) (-) obtained previously [F. A. Akin and C. C. Jarrold, J. Chem. Phys. 118, 5841 (2003)]. PMID- 15267801 TI - Surface tension of quantum fluids from molecular simulations. AB - We present the first molecular simulations of the vapor-liquid surface tension of quantum liquids. The path integral formalism of Feynman was used to account for the quantum mechanical behavior of both the liquid and the vapor. A replica-data parallel algorithm was implemented to achieve good parallel performance of the simulation code on at least 32 processors. We have computed the surface tension and the vapor-liquid phase diagram of pure hydrogen over the temperature range 18 30 K and pure deuterium from 19 to 34 K. The simulation results for surface tension and vapor-liquid orthobaric densities are in very good agreement with experimental data. We have computed the interfacial properties of hydrogen deuterium mixtures over the entire concentration range at 20.4 and 24 K. The calculated equilibrium compositions of the mixtures are in excellent agreement with experimental data. The computed mixture surface tension shows negative deviations from ideal solution behavior, in agreement with experimental data and predictions from Prigogine's theory. The magnitude of the deviations at 20.4 K are substantially larger from simulations and from theory than from experiments. We conclude that the experimentally measured mixture surface tension values are systematically too high. Analysis of the concentration profiles in the interfacial region shows that the nonideal behavior can be described entirely by segregation of H(2) to the interface, indicating that H(2) acts as a surfactant in H(2)-D(2) mixtures. PMID- 15267802 TI - Effect of the erbium dopant architecture on the femtosecond relaxation dynamics of silicon nanocrystals. AB - Femtosecond pump-probe absorption spectroscopy is used to investigate the role of Er(3+) dopants in the early relaxation pathways of photoexcited Si nanocrystals. The fate of photoexcited electrons in three different Si nanostructures was studied and correlated with the effect of Er-doping and the nature of the dopant architecture. In Si nanocrystals without Er(3+) dopant, a trapping component was identified to be a major electron relaxation mechanism. Addition of Er(3+) ions into the core or surface shell of the nanocrystals was found to open up additional nonradiative relaxation pathways, which is attributed to Er-induced trap states in the Si host. Analysis of the photodynamics of the Si nanocrystal samples reveals an electron trapping mechanism involving trap-to-trap hopping in the doped nanocrystals, whereby the density of deep traps seem to increase with the presence of erbium. To gain additional insights on the relative depths of the trapping sites on the investigated nanostructures, benzoquinone was used as a surface adsorbed electron acceptor to facilitate photoinduced electron transfer across the nanocrystal surface and subsequently assist in back electron transfer. The established reduction potential (-0.45 V versus SCE) of the electron acceptor helped reveal that the erbium-doped nanocrystal samples have deeper trapping sites than the undoped Si. Furthermore, the measurements indicate that internally Er-doped Si have relatively deeper trapping sites than the erbium surface enriched nanocrystals. The electron-shuttling experiment also reveals that the back electron transfer seems not to recover completely to the ground state in the doped Si nanocrystals, which is explained by a mechanism whereby the electrons are captured by deep trapping sites induced by erbium addition in the Si lattice. PMID- 15267803 TI - Effects of resolution and friction in the interpretation of QHAS measurements. AB - We use Langevin molecular dynamics (MD) simulations to improve the picture of the processes that contribute to QHAS broadening, as a function of momentum transfer at the crystal. We use a simulation of realistic damped vibrational motion in a harmonic well to establish the contributions to QHAS measurements due to both vibrational motion and intracell diffusion (usually referred to as vibrational dephasing). We demonstrate the experimental conditions under which such contributions are likely to be important. These results are compared with those from a simulation of thermal diffusion over a sinusoidally corrugated potential. We show that resolution and atom-surface "friction" play an important role in determining the form of QHAS measurements and we demonstrate that there is no simple relationship between the "activation energy" derived from an Arrhenius plot of QHAS data and the adiabatic potential barrier height. MD simulations are therefore necessary to perform a complete analysis of the data. Finally, we discuss the application of our results to more sophisticated potentials, particularly those with multiple adsorption sites. PMID- 15267804 TI - Ab initio many-body investigation of structure and stability of two-fold rings in silicates. AB - In this paper we present ab initio many-body calculations on the strain energy of W silica, taken as a model system for edge-sharing tetrahedral SiO(2) systems with respect to corner-sharing ones as in alpha quartz. The mean-field results were obtained using the restricted Hartree-Fock approach, while the many-body effects were taken into account by the second-order Moller-Plesset perturbation theory and the coupled-cluster approach. Correlation contributions are found to play an important role to determine the stability of edge-sharing units. The most sophisticated method used in our calculation, i.e., the coupled-cluster approach with single and double excitations, yields a strain energy of 0.0427 a.u. per Si(2)O(4) unit with respect to alpha quartz, which is even smaller than the value obtained by a previous density functional theory calculation. PMID- 15267805 TI - Molecular exchange through vesicle membranes: a pulsed field gradient nuclear magnetic resonance study. AB - The permeability of block copolymer vesicles is studied using pulsed field gradient nuclear magnetic resonance spectroscopy together with a numerical data analysis procedure. Polyethylene oxide molecules of various molecular masses are used to sample the permeability of the vesicle membrane by observing the trans membrane exchange process under equilibrium conditions. For shorter polyethylene oxide chains, the analysis yields a nearly linear dependence of the logarithmic trans-membrane exchange rate on the hydrodynamic radius of the sample molecules. PMID- 15267806 TI - Internal stochastic resonance under two-parameter modulation in intercellular calcium ion oscillations. AB - Internal stochastic resonance (ISR) in a model of intercellular calcium ion oscillations is investigated under the modulation of two parameters, viz., degree of extracellular stimulation (beta) and leak rate (k(f)). ISR can occur when either beta or k(f) is subjected to a noise. Internal stochastic biresonance (ISBR) can occur when noise is added to the two parameters simultaneously. The distance to the bifurcation point is found to be able to enhance or suppress the ISBR, and to affect the number of peaks of ISR. PMID- 15267807 TI - The adsorption and dissociation of Cl2 on the MgO (001) surface with vacancies: embedded cluster model study. AB - The adsorption of Cl(2) at a low-coordinated oxygen site (edge or corner site) and vacancy site (terrace, edge, corner F, F(+), or F(2+) center) has been studied by the density functional method, in conjunction with the embedded cluster models. First, we have studied the adsorption of Cl(2) at the edge and corner oxygen sites and the results show that Cl(2), energetically, is inclined to adsorb at the corner oxygen site. Moreover, similar to the most advantageous adsorption mode for Cl(2) on the MgO (001) perfect surface, the most favorable adsorption occurs when Cl(2) approaches the corner oxygen site along the normal direction. A small amount of electrons are transferred from the substrate to the antibonding orbital of the adsorbate, leading to the Cl-Cl bond strength weakened a little. Regarding Cl(2) adsorption at the oxygen vacancy site (F, F(+), or F(2+) center), both large adsorption energies and rather much elongation of the Cl-Cl bond length have been obtained, in particular at the corner oxygen vacancy site, with concurrently large amounts of electrons transferred from the substrate to the antibonding orbital of Cl(2). It suggests, at the oxygen vacancy site, that Cl(2) prefers to dissociate into Cl subspecies. And the potential energy surface indicates that the dissociation process of molecular Cl(2) to atomic Cl is virtually barrierless. PMID- 15267808 TI - Hot-atom versus Eley-Rideal dynamics in hydrogen recombination on Ni(100). I. The single-adsorbate case. AB - We compare the efficiency of the Eley-Rideal (ER) reaction with the formation of hot-atom (HA) species in the simplest case, i.e., the scattering of a projectile off a single adsorbate, considering the Hydrogen and Hydrogen-on-Ni(100) system. We use classical mechanics and the accurate embedded diatomics-in-molecules potential to study the collision system over a wide range of collision energies (0.10-1.50 eV), both with a rigid and a nonrigid Ni substrate and for impact on the occupied and neighboring empty cells. In the rigid model metastable and truly bound hot-atoms occur and we find that the cross section for the formation of bound hot-atoms is considerably higher than that for the ER reaction over the whole range of collision energies examined. Metastable hot-atoms form because of the inefficient energy transfer to the adsorbate and have lifetimes of the order 0.1-0.7 ps, depending on the collision energy. When considering the effects of lattice vibrations we find, on average, a consistent energy transfer to the substrate, say 0.1-0.2 eV, which forced us to devise a two-step dynamical model to get rid of the problems associated with the use of periodic boundary conditions. Results for long-lived HA formation due to scattering on the occupied cell at a surface temperature of 120 K agree well with those of the rigid model, suggesting that in the above process the substrate plays only a secondary role and further calculations at surface temperatures of 50 and 300 K are in line with these findings. However, considerably high cross sections for formation of long lived hot-atoms result also from scattering off the neighboring cells where the energy transfer to the lattice cannot be neglected. Metastable hot-atoms are reduced in number and have usually lifetimes shorter than those of the rigid model, say less than 0.3 ps. In addition, ER cross sections are only slightly affected by the lattice motion and show a little temperature dependence. Finally, we find also that absorption and reflection strongly depend on the correct consideration of lattice vibrations and the occurrence of trapping. PMID- 15267809 TI - Mechanism of linear and nonlinear optical effects of chalcopyrite AgGaX2 (X=S, Se, and Te) crystals. AB - The electronic band structures for AgGaX(2) (X=S, Se, Te) chalcopyrites have been calculated using a pseudopotential total energy method. First-principles calculations of the linear and nonlinear optical properties are presented for these crystals, with the electronic band structures obtained from pseudopotential method as input. The theoretical refractive indices and nonlinear optical coefficients are in good agreement with available experimental values. The origin of the nonlinear optical effects is explained through real-space atom-cutting analysis. The contribution of the GaX(2) group (X=S, Se, Te) for second harmonic generation (SHG) effect is dominant while that of the cation Ag is negligible. In addition, the percentage contribution to the SHG coefficients from the different bonds increase with increase of the bond order. PMID- 15267810 TI - Contact area between a viscoelastic solid and a hard, randomly rough, substrate. AB - We study the time-dependent contact area as a viscoelastic solid is squeezed against a randomly rough substrate. Using a recently developed contact mechanics theory we study how the contact area depends on time and on the magnification zeta. Numerical results are presented for self-affine fractal surfaces, and applications to tack, rubber friction, and sealing are given. PMID- 15267811 TI - Quantum trajectories in atom-surface scattering with single adsorbates: the role of quantum vortices. AB - In this work, a full quantum study of the scattering of He atoms off single CO molecules, adsorbed onto the Pt(111) surface, is presented within the formalism of quantum trajectories provided by Bohmian mechanics. By means of this theory, it is shown that the underlying dynamics is strongly dominated by the existence of a transient vortitial trapping with measurable effects on the whole diffraction pattern. This kind of trapping emphasizes the key role played by quantum vortices in this scattering. Moreover, an analysis of the surface rainbow effect caused by the local corrugation that the CO molecule induces on the surface, and its manifestation in the corresponding intensity pattern, is also presented and discussed. PMID- 15267812 TI - Electrolyte-induced collapse of a polyelectrolyte brush. AB - We have investigated the electrolyte-induced collapse of a polyelectrolyte brush covalently attached to a planar solid surface. Positively charged poly-4-vinyl [N methyl-pyridinium] (MePVP) brushes were prepared in situ at the surface by free radical chain polymerization using a surface-immobilized initiator monolayer ("grafting from" technique) and 4-vinylpyridine as the monomer, followed by a polymer-analogous quaternization reaction. The height of the brushes was measured as a function of the external salt concentration via multiple-angle null ellipsometry. As predicted by mean-field theory, the height of the MePVP brushes remains unaffected by the addition of low amounts of external salt. At higher salt concentrations the brush height decreases. The extent to which the brush shrinks strongly depends on the nature of the salt present in the environment. MePVP brushes collapse to almost the dry layer thickness upon the addition of potassium iodide to a contacting aqueous medium. In contrast, the collapse of MePVP brushes having bromide or chloride counterions is much less pronounced. These brushes remain in a highly swollen state even after large amounts of salt have been added to the solution. PMID- 15267813 TI - Molecular structure-dynamics relationships in glassy poly(isophthalamide)s as revealed by wide angle x-ray scattering, dielectric loss spectroscopy, and molecular modelling. AB - The effect of molecular structure on the gamma relaxation dynamics has been studied in a set of aromatic poly(isophthalamide)s. This polymer family differ in the bridge group between phenylene rings [hexafluoroisopropylidene (C(CF(3))(2)) or ether] and also in the presence of t-butyl groups (C(CH(3))(3)) as pendant substituent on the five position of isophthalic ring. The results obtained from wide angle x-ray scattering in the glassy state indicated that both (C(CF(3))(2)) and (C(CH(3))(3)) groups favor the separation between chains, which is reflected on different interchain average distances. Dielectric experiments showed that both bulky groups favor the mobility in the glassy state. Molecular modelling methods were used to know the kind of molecular motions associated to the dielectric relaxation observed below the glass transition temperature. PMID- 15267814 TI - Aging in short-ranged attractive colloids: a numerical study. AB - We study the aging dynamics in a model for dense simple liquids, in which particles interact through a hard-core repulsion complemented by a short-ranged attractive potential, of the kind found in colloidal suspensions. In this system, at large packing fractions, kinetically arrested disordered states can be created both on cooling (attractive glass) and on heating (repulsive glass). The possibility of having two distinct glasses, at the same packing fraction, with two different dynamics offers the unique possibility of comparing-within the same model-the differences in aging dynamics. We find that, while the aging dynamics of the repulsive glass is similar to the one observed in atomic and molecular systems, the aging dynamics of the attractive glass shows novel unexpected features. PMID- 15267815 TI - Static and dynamic properties of tethered chains at adsorbing surfaces: a Monte Carlo study. AB - We present extensive Monte Carlo simulations of tethered chains of length N on adsorbing surfaces, considering the dilute case in good solvents, and analyze our results using scaling arguments. We focus on the mean number M of chain contacts with the adsorbing wall, on the chain's extension (the radius of gyration) perpendicular and parallel to the adsorbing surface, on the probability distribution of the free end and on the density profile for all monomers. At the critical adsorption strength epsilon(c) one has M(c) approximately N(phi), and we find (using the above results) as best candidate phi to equal 0.59. However, slight changes in the estimation of epsilon(c) lead to large deviations in the resulting phi; this might be a possible reason for the difference in the phi values reported in the literature. We also investigate the dynamical scaling behavior at epsilon(c), by focusing on the end-to-end correlation function and on the correlation function of monomers adsorbed at the wall. We find that at epsilon(c) the dynamic scaling exponent a (which describes the relaxation time of the chain as a function of N) is the same as that of free chains. Furthermore, we find that for tethered chains the modes perpendicular to the surface relax quicker than those parallel to it, which may be seen as a splitting in the relaxation spectrum. PMID- 15267816 TI - An investigation of a sol-gel/melt transition: the poly(ethylene oxide)/methanol/LiClO4 system. AB - The crossover behavior of 50 000 molar mass poly(ethylene oxide)/methanol solutions from dilute solution to the melt/gel was examined. At first this behavior was investigated without LiClO(4) and then reexamined with LiClO(4). To better understand this behavior, the dependencies of dynamic light scattering (specifically, photon correlation spectroscopy) measurement results on polymer concentration, on the scattering wave vector and on temperature, and the dependence of static light scattering results on the scattering wave vector were studied. This study produced interesting and important results about network structure and behavior in poly(ethylene oxide) solutions and melts generally and about the effects of LiClO(4) on this structure and behavior more particularly. PMID- 15267817 TI - Relative importance of local and collective effects in the distortivity of one dimensional chains. AB - On the basis of an original coupled-cluster type formalism developed previously [J.-P. Malrieu and V. Robert, J. Chem. Phys. 120, 7374 (2004)], the cohesive energies and the occurrence of Peierls instability are investigated in half filled one-dimensional systems. Starting from various parametrizations of the Huckel Hamiltonian, this approach allows one to evaluate the relative contribution of local and collective interactions by comparison to exact tight binding crystal calculations. For an alternating (AA('))(n) chain, quantitative agreement with the exact solution is obtained starting from either an atom centered or a bond-centered reference function. The distortion takes place beyond a critical value of the electron-phonon/elastic strength ratio which is correctly predicted. Its amplitude and the corresponding stabilization energy are also accurately reproduced, suggesting that the driving force of the second-order Peierls distortion is essentially local. For homogeneous (A)(n) systems and the first-order Peierls distortion traditionally presented as resulting from a band gap opening (i.e., collective effects), our localized approaches are deficient only in the domain of weak electron-phonon/elastic ratio where the distortion amplitude is almost negligible. These results confirm that the short-range delocalization effects are the leading phenomenon responsible for the bond alternation in conjugated hydrocarbons. PMID- 15267818 TI - Moments and distribution function of polyelectrolyte chains. AB - It is demonstrated that the moments R(2k) of the end-to-end distance distribution function f(r) of charged wormlike chains with excluded volume effects in solution with added salt, in the Debye-Huckel approximation, obey a remarkably simple relation. It is shown that the R(2k) can be expressed as weighted sums of the corresponding moments of ideal wormlike chains. As an application of this method, we show that the Fourier transform of f(r) can also be represented by a superposition of distribution functions of ideal chains. The quantities so calculated are compared with the results of Monte Carlo simulations. Excellent agreement between them is observed. PMID- 15267819 TI - Pure intermolecular vibrational relaxation of the OH bending mode of water molecules. PMID- 15267820 TI - Comment on "Equations of state for fluids: the Dieterici approach revisited" [J. Chem. Phys. 115, 1460 (2001)]. PMID- 15267823 TI - Anomalous mixing behavior of polyisobutylene/polypropylene blends: molecular dynamics simulation study. AB - The unusual mixing behavior of polyisobutylene (PIB) with head-to-head (hhPP) and head-to-tail polypropylene (PP) is studied using large-scale molecular dynamics (MD). The heats of mixing and Flory chi parameters were computed from MD simulations of both blends using a united atom model. The chi parameters from the simulations were estimated from the structure factors using the random phase approximation in analogy with neutron scattering (SANS) experiments. MD simulations for syndiotactic hhPP/PIB predicted a lower critical solution temperature with a chi parameter in very good agreement with SANS experiments on the atactic hhPP/PIB blend. MD simulations also predicted that the isotactic PP/PIB blend was immiscible at high molecular weight in qualitative agreement with cloud point measurements on atactic PP/PIB. PMID- 15267824 TI - Polarizability and optical rotation calculated from the approximate coupled cluster singles and doubles CC2 linear response theory using Cholesky decompositions. AB - A new implementation of the approximate coupled cluster singles and doubles CC2 linear response model using Cholesky decomposition of the two-electron integrals is presented. Significantly reducing storage demands and computational effort without sacrificing accuracy compared to the conventional model, the algorithm is well suited for large-scale applications. Extensive basis set convergence studies are presented for the static and frequency-dependent electric dipole polarizability of benzene and C60, and for the optical rotation of CNOFH2 and (-) trans-cyclooctene (TCO). The origin-dependence of the optical rotation is calculated and shown to persist for CC2 even at basis set convergence. PMID- 15267825 TI - Fluctuation theorem for nonequilibrium reactions. AB - A fluctuation theorem is derived for stochastic nonequilibrium reactions ruled by the chemical master equation. The theorem is expressed in terms of the generating and large-deviation functions characterizing the fluctuations of a quantity which measures the loss of detailed balance out of thermodynamic equilibrium. The relationship to entropy production is established and discussed. The fluctuation theorem is verified in the Schlogl model of far-from-equilibrium bistability. PMID- 15267826 TI - The Fisher-Shannon information plane, an electron correlation tool. AB - A new correlation measure, the product of the Shannon entropy power and the Fisher information of the electron density, is introduced by analyzing the Fisher Shannon information plane of some two-electron systems (He-like ions, Hooke's atoms). The uncertainty and scaling properties of this information product are pointed out. In addition, the Fisher and Shannon measures of a finite many electron system are shown to be bounded by the corresponding single-electron measures and the number of electrons of the system. PMID- 15267827 TI - Fully adaptive propagation of the quantum-classical Liouville equation. AB - In mixed quantum-classical molecular dynamics few but important degrees of freedom of a dynamical system are modeled quantum-mechanically while the remaining ones are treated within the classical approximation. Rothe methods established in the theory of partial differential equations are used to control both temporal and spatial discretization errors on grounds of a global tolerance criterion. The TRAIL (trapezoidal rule for adaptive integration of Liouville dynamics) scheme [I. Horenko and M. Weiser, J. Comput. Chem. 24, 1921 (2003)] has been extended to account for nonadiabatic effects in molecular dynamics described by the quantum-classical Liouville equation. In the context of particle methods, the quality of the spatial approximation of the phase-space distributions is maximized while the numerical condition of the least-squares problem for the parameters of particles is minimized. The resulting dynamical scheme is based on a simultaneous propagation of moving particles (Gaussian and Dirac deltalike trajectories) in phase space employing a fully adaptive strategy to upgrade Dirac to Gaussian particles and, vice versa, downgrading Gaussians to Dirac-type trajectories. This allows for the combination of Monte-Carlo-based strategies for the sampling of densities and coherences in multidimensional problems with deterministic treatment of nonadiabatic effects. Numerical examples demonstrate the application of the method to spin-boson systems in different dimensionality. Nonadiabatic effects occurring at conical intersections are treated in the diabatic representation. By decreasing the global tolerance, the numerical solution obtained from the TRAIL scheme are shown to converge towards exact results. PMID- 15267828 TI - Kinematic effects associated with molecular frames in structural isomerization dynamics of clusters. AB - Kinematic effects associated with movements of molecular frames, which specify instantaneous orientation of molecules, is investigated in structural isomerization dynamics of a triatomic cluster whose total angular momentum is zero. The principal-axis frame is employed to introduce the so-called principal axis hyperspherical coordinates, with which the mechanism of structural isomerization dynamics of the cluster is systematically analyzed. A force called "democratic centrifugal force" is extracted from the associated kinematics. This force arises from an intrinsic non-Euclidean metric in the internal space and has an effect of distorting the triatomic cluster to a collapsed shape and of trapping the system around collinear transition states. The latter effect is particularly important in that the kinematics effectively makes a basin at the saddle (transition state) on the potential surface. Based on this framework, we study the effect of the gauge field associated with the Eckart frame in internal space, which has not been carefully examined in the conventional reaction rate theories. Numerical comparison between the dynamics with and without the gauge field has revealed that this field has an effect to suppress the rate of isomerization reaction to a considerable amount. Thus a theory neglecting this effect will significantly overestimate the rate of isomerization. We show the physical origin of this suppressing effect. PMID- 15267829 TI - Semiclassical theory of electronically nonadiabatic chemical dynamics: incorporation of the Zhu-Nakamura theory into the frozen Gaussian propagation method. AB - The title theory is developed by combining the Herman-Kluk semiclassical theory for adiabatic propagation on single potential-energy surface and the semiclassical Zhu-Nakamura theory for nonadiabatic transition. The formulation with use of natural mathematical principles leads to a quite simple expression for the propagator based on classical trajectories and simple formulas are derived for overall adiabatic and nonadiabatic processes. The theory is applied to electronically nonadiabatic photodissociation processes: a one-dimensional problem of H2+ in a cw (continuous wave) laser field and a two-dimensional model problem of H2O in a cw laser field. The theory is found to work well for the propagation duration of several molecular vibrational periods and wide energy range. Although the formulation is made for the case of laser induced nonadiabatic processes, it is straightforwardly applicable to ordinary electronically nonadiabatic chemical dynamics. PMID- 15267830 TI - Determination of noncovalent interaction energies from electronic densities. AB - Starting from the Hellmann-Feynman theorem, an expression is derived for the interaction energy in forming a noncovalently bound complex. It invokes only classical electrostatics, involving the charge distributions of the components as they are in the complex. We propose a method for obtaining these from the total charge distribution. Integration over the electronic densities is carried out by a numerical procedure slightly modified from that of Gavezzotti. We calculate the interaction energies for four molecular dimers at a variety of computational levels. The results are analyzed and compared to the best estimated values available in the literature. PMID- 15267831 TI - Solution of quantum Langevin equation: approximations, theoretical and numerical aspects. AB - Based on a coherent state representation of noise operator and an ensemble averaging procedure using Wigner canonical thermal distribution for harmonic oscillators, a generalized quantum Langevin equation has been recently developed [Phys. Rev. E 65, 021109 (2002); 66, 051106 (2002)] to derive the equations of motion for probability distribution functions in c-number phase-space. We extend the treatment to explore several systematic approximation schemes for the solutions of the Langevin equation for nonlinear potentials for a wide range of noise correlation, strength and temperature down to the vacuum limit. The method is exemplified by an analytic application to harmonic oscillator for arbitrary memory kernel and with the help of a numerical calculation of barrier crossing, in a cubic potential to demonstrate the quantum Kramers' turnover and the quantum Arrhenius plot. PMID- 15267832 TI - Lifetime and predissociation yield of 14N2 b 1Piu(v=1). AB - The lifetime of the b 1Piu(v=1) state in 14N2 has been determined experimentally using a laser-based pump-probe scheme and an exceptionally long lifetime of 2.61 ns was found. Semiempirical close-coupling calculations of the radiative lifetime, which include Rydberg-valence interactions in the singlet manifold, are consistent with this large value, giving a value of 3.61 ns and suggesting a predissociation yield of approximately 28% for this level of the b state. PMID- 15267833 TI - Carbon-carbon bond cleavage in the photoionization of ethanol and 1-propanol clusters. AB - Tunable VUV laser was used to initiate the ion-molecule reactions in the clusters of ethanol and 1-propanol by photoionization in the region between 10.49 to 10.08 eV. Ionic products were detected by the time-of-flight mass spectrometer. In addition to the protonated clusters from proton transfer reactions, the products corresponding to beta carbon-carbon bond cleavage were found to be one of the major products for small sizes of clusters. A comparison with photoionization of methanol clusters and the results of ab initio calculation has been made. PMID- 15267834 TI - Structures and stability of medium silicon clusters. II. Ab initio molecular orbital calculations of Si12-Si20. AB - Ab initio all-electron molecular-orbital calculations are carried out to study the structures and relative stability of low-energy silicon clusters (Si(n),n = 12-20). Selected geometric isomers include those predicted by Ho et al. [Nature (London) 392, 582 (1998)] based on an unbiased search with tight-binding/genetic algorithm, as well as those found by Rata et al. [Phys. Rev. Lett. 85, 546 (2000)] based on density-functional tight-binding/single-parent evolution algorithm. These geometric isomers are optimized at the Moller-Plesset (MP2) MP2/6-31G(d) level. The single-point energy at the coupled-cluster single and double substitutions (including triple excitations) [CCSD(T)] CCSD(T)/6-31G(d) level for several low-lying isomers are further computed. Harmonic vibrational frequency analysis at the MP2/6-31G(d) level of theory is also undertaken to assure that the optimized geometries are stable. For Si12-Si17 and Si19 the isomer with the lowest-energy at the CCSD(T)/6-31G(d) level is the same as that predicted by Ho et al., whereas for Si18 and Si20, the same as predicted by Rata et al. However, for Si14 and Si15, the vibrational frequency analysis indicates that the isomer with the lowest CCSD(T)/6-31G(d) single-point energy gives rise to imaginary frequencies. Small structural perturbation onto the Si14 and Si15 isomers can remove the imaginary frequencies and results in new isomers with slightly lower MP2/6-31G(d) energy; however the new isomers have a higher single point energy at the CCSD(T)/6-31G(d) level. For most Si(n) (n = 12-18,20) the low lying isomers are prolate in shape, whereas for Si19 a spherical-like isomer is slightly lower in energy at the CCSD(T)/6-31G(d) level than low-lying prolate isomers. PMID- 15267835 TI - Competition between linear and cyclic structures in monochromium carbide clusters CrCn- and CrCn (n = 2-8): a photoelectron spectroscopy and density functional study. AB - Photoelectron spectroscopy (PES) is combined with density functional theory (DFT) to study the monochromium carbide clusters CrCn- and CrCn (n = 2-8). Well resolved PES spectra were obtained, yielding structural, electronic, and vibrational information about both the anionic and neutral clusters. Experimental evidence was observed for the coexistence of two isomers for CrC2-, CrC3-, CrC4-, and CrC6-. Sharp and well-resolved PES spectra were observed for CrCn- (n = 4,6,8), whereas broad spectra were observed for CrC5- and CrC7-. Extensive DFT calculations using the generalized gradient approximation were carried out for the ground and low-lying excited states of all the CrCn- and CrCn species, as well as coupled-cluster calculations for CrC2- and CrC2. Theoretical electron affinities and vertical detachment energies were calculated and compared with the experimental data to help the assignment of the ground states and obtain structural information. We found that CrC2- and CrC3- each possess a close-lying cyclic and linear structure, which were both populated experimentally. For the larger CrCn- clusters with n = 4, 6, 8, linear structures are the overwhelming favorite, giving rise to the sharp PES spectral features. CrC7- was found to have a cyclic structure. The broad PES spectra of CrC5- suggested a cyclic structure, whereas the DFT results predicted a linear one. PMID- 15267836 TI - The influence of laser field noise on controlled quantum dynamics. AB - The influence of laser noise on the dynamics of simple quantum systems is analyzed. An anharmonic ladder is chosen for illustration and several pulses are obtained that optimize the yield of a quantum transition by constraining the laser parameters. The following models of laser noise are introduced: Amplitude white noise, phase white noise, frequency white noise and shot-to-shot static noise in the different pulse parameters. It is shown that the optimal pulses are robust to white amplitude noise, since the system acts as a dynamical filter. White phase noise affects the optimal pulses in a similar way by reducing the pulse area. This effect can be easily compensated for by pulse amplitude rescaling, up to a high level of noise. White frequency noise reduces the pulse area and induces spectral broadening, more strongly affecting the high frequency components. It can be partially compensated for by amplitude rescaling. The effects of static noise in the parameters cannot be easily corrected. It is shown that optimal pulses that drive n-photon transitions become more sensitive to noise in the amplitude and less sensitive to noise in the frequency as n increases. The effects of noise in the relative phase rapidly become constant for a large number of interfering pathways. PMID- 15267837 TI - Efficient dehalogenation of polyhalomethanes and production of strong acids in aqueous environments: water-catalyzed O--H-insertion and HI-elimination reactions of isodiiodomethane (CH2I-I) with water. AB - A combined experimental and theoretical study of the ultraviolet photolysis of CH2I2 in water is reported. Ultraviolet photolysis of low concentrations of CH2I2 in water was experimentally observed to lead to almost complete conversion into CH2(OH)2 and 2HI products. Picosecond time-resolved resonance Raman spectroscopy experiments in mixed water/acetonitrile solvents (25%-75% water) showed that appreciable amounts of isodiiodomethane (CH2I-I) were formed within several picoseconds and the decay of the CH2I-I species became substantially shorter with increasing water concentration, suggesting that CH2I-I may be reacting with water. Ab initio calculations demonstrate the CH2I-I species is able to react readily with water via a water-catalyzed O--H-insertion and HI-elimination reaction followed by its CH2I(OH) product undergoing a further water-catalyzed HI elimination reaction to make a H2C=O product. These HI-elimination reactions produce the two HI leaving groups observed experimentally and the H2C=O product further reacts with water to produce the other final CH2(OH)2 product observed in the photochemistry experiments. These results suggest that CH2I-I is the species that reacts with water to produce the CH2(OH)2 and 2HI products seen in the photochemistry experiments. The present study demonstrates that ultraviolet photolysis of CH2I2 at low concentration leads to efficient dehalogenation and release of multiple strong acid (HI) leaving groups. Some possible ramifications for the decomposition of polyhalomethanes and halomethanols in aqueous environments as well as the photochemistry of polyhalomethanes in the natural environment are briefly discussed. PMID- 15267838 TI - Infrared-induced conformational isomerization and vibrational relaxation dynamics in melatonin and 5-methoxy-N-acetyl tryptophan methyl amide. AB - The conformational isomerization dynamics of melatonin and 5-methoxy N acetyltryptophan methyl amide (5-methoxy NATMA) have been studied using the methods of IR-UV hole-filling spectroscopy and IR-induced population transfer spectroscopy. Using these techniques, single conformers of melatonin were excited via a well-defined NH stretch fundamental with an IR pump laser. This excess energy was used to drive conformational isomerization. By carrying out the infrared excitation early in a supersonic expansion, the excited molecules were re-cooled into their zero-point levels, partially re-filling the hole created in the ground state population of the excited conformer, and creating gains in population of the other conformers. These changes in population were detected using laser-induced fluorescence downstream in the expansion via an UV probe laser. The isomerization quantum yields for melatonin show some conformation specificity but no hint of vibrational mode specificity. In 5-methoxy NATMA, no isomerization was observed out of the single conformational well populated in the expansion in the absence of the infrared excitation. In order to study the dependence of the isomerization on the cooling rate, the experimental arrangement was modified so that faster cooling conditions could be studied. In this arrangement, the pump and probe lasers were overlapped in space in the high density region of the expansion, and the time dependence of the zero-point level populations of the conformers was probed following selective excitation of a single conformation. The analysis needed to extract isomerization quantum yields from the timing scans was developed and applied to the melatonin timing scans. Comparison between the frequency and time domain isomerization quantum yields under identical experimental conditions produced similar results. Under fast cooling conditions, the product quantum yields were shifted from their values under standard conditions. The results for melatonin are compared with those for N-acetyl tryptophan methyl amide. PMID- 15267839 TI - Microwave and ab initio studies of rare gas-methane van der Waals complexes. AB - Rotational spectra of the weakly bound Kr-methane van der Waals complex were recorded using a pulsed molecular beam Fourier transform microwave spectrometer in the range from 3.5 to 18 GHz. Spectra of 25 isotopomers of Kr-methane were assigned and analyzed. For isotopomers containing CH4, 13CH4, and CD4, two sets of transitions with K = 0 and one with K = 1 were recorded, correlating to the j = 0, 1, and 2 rotational levels of free methane, respectively (j is the rotational angular momentum quantum number of the methane monomer). For isotopomers containing CH3D and CHD3, two K = 0 components were recorded, correlating to the j(k) = 0(0) and 1(1) rotational levels of free methane (k corresponds to the projection of j onto the C3 axis of CH3D and CHD3). The obtained spectroscopic results were used to derive van der Waals bond distance R, van der Waals stretching frequency nu(s), and the corresponding stretching force constant k(s). Nuclear spin statistical weights of individual states were obtained from molecular symmetry group analyses and were compared with the observed relative transition intensities. The tentatively assigned j = 2 transitions were more intense than predicted from symmetry considerations. This is attributed to a relatively large effective dipole moment of this state, supported by ab initio dipole moment calculations. Ab initio potential energy calculations of Kr-CH4 and Ar-CH4 were done at the coupled cluster level of theory, with single and double excitations and perturbative inclusion of triple excitations, using the aug-cc-pVTZ basis set supplemented with bond functions. The theoretical results show that the angular dynamics of the dimer does not change significantly when the binding partner of methane changes from Ar to Kr. The dipole moment of Ar-CH4 was calculated at various configurations, providing a qualitative explanation for the unsuccessful spectral searches for rotational transitions of Ar-CH4. PMID- 15267840 TI - A new expression for the direct quantum mechanical evaluation of the thermal rate constant. AB - Based on the formalism of Miller, Schwartz, and Tromp [J. Chem. Phys. 79, 4889(1983)], we derive a new expression for the thermal rate constant for a chemical reaction. The expression involves an unperturbed, i.e., reactant or product channel Boltzmann operator for the imaginary time propagation, making it possible to compute efficiently the rate constant for a range of temperatures. We illustrate numerical aspects with an extensive study of the one-dimensional Eckart barrier problem, as well as a study of the three-dimensional (J = 0) D + H2 problem. PMID- 15267841 TI - Rotational dynamics of CO solvated in small He clusters: a quantum Monte Carlo study. AB - The rotational dynamics of CO single molecules solvated in small He clusters (CO @ HeN) has been studied using reptation quantum Monte Carlo simulations for cluster sizes up to N = 30. Our results are in good agreement with the rotovibrational features of the infrared spectrum recently determined for this system and provide a deep insight into the relation between the structure of the cluster and its dynamics. Simulations for large N also provide a prediction of the effective moment of inertia of CO in the He nanodroplet regime, which has not been measured so far. PMID- 15267842 TI - Are insertion compounds of CH2CHF and the rare gases stable? A computational study. AB - Ab initio calculations, using second-order Moller-Plesset perturbation theory with a 6-311+ +G(2d,2p) basis set, predict the stability of two novel compounds of monofluoroethene, CH2CHF, with the rare-gas atoms Ar and Kr. The dissociation energies to the lowest-energy fragmentation products, CH2CHF + Rg (Rg = Ar,Kr), were computed to be -528 and -449 kJ mol(-1), respectively, at the coupled cluster singles, doubles, and triples level of theory. Possible transition states (at second-order Moller-Plesset theory) via a C-Rg-F bending mode for these fragmentation reactions were also located with barrier heights of about 76 and 106 kJ mol(-1), for the Ar- and Kr-containing species, respectively. However, the Ar-containing species may not exist at all as it is less stable than the fragments CH2CH + F + Ar at the higher level of theory and may possibly dissociate via this route. PMID- 15267843 TI - Quantum capping potentials with point charges: a simple QM/MM approach for the calculation of large-molecule NMR shielding tensors. AB - A simple quantum-mechanics/molecular-mechanics (QM/MM) approach for calculating NMR shielding tensors (sigma) is presented. The method involves capping the QM region with quantum capping potentials (QCPs) and representing the MM region with point charges. Test calculations on simple systems without MM charges show that calculated sigma values improve relative to the full QM results with increasing distance between the capped bond and chromophore. Calculations on the histidine amino acid and cytosine monophosphate (CMP) nucleic acid show that the use of QCPs with point charges result in mean errors in the isotropic component of sigma that are less than 1.6 ppm. The results also reveal that, contrary to previous work, the explicit effect of point charges on sigma through coupling with gauge factors, as in the gauge including atomic orbital approach, is minimal for the CMP molecule. The present QM/MM approach for calculating sigma is easy to apply and requires no code modification. PMID- 15267844 TI - S(1S) production following electron impact on thiophosgene (Cl2CS). AB - A special xenon matrix detector has been used to study the production of S(1S) following controlled electron impact on thiophosgene (Cl2CS) targets over an electron energy range from threshold to 400 eV. Time-of-flight spectroscopy has been used to measure S(1S) fragment kinetic energies. Fragments with energies in excess of 1 eV have been observed. The absolute cross section for S(1S) production reaches a maximum of [1.05+/-0.35] x 10(-18) cm2 at approximately 125 eV impact energy. Two different fragmentation processes, involving triplet and singlet excited states of the parent Cl2CS molecule, have been identified. PMID- 15267845 TI - Kinetic theory of radio frequency quadrupole ion traps. I. Trapping of atomic ions in a pure atomic gas. AB - A kinetic theory based on the Boltzmann equation is developed for the trapping of atomic ions in a radio-frequency quadrupole ion trap containing enough neutral atoms that ion-neutral collisions cannot be ignored. The collisions are treated at the same level of sophistication and detail as is used to deal with the time- and space-dependent electric fields in the trap. As a result, microscopic definitions are obtained for the damping and stochastic forces that originate from such collisions. These definitions contrast with corresponding phenomenological terms added ad hoc in previous treatments to create damped Mathieu and Langevin equations, respectively. Furthermore, the theory indicates that either collisional cooling or heating of the ions is possible, depending upon details of the ion-neutral mass ratios and interaction potential. The kinetic theory is not dependent on any special assumptions about the electric field strengths, the ion-neutral interaction potentials, or the ion-neutral mass ratio. It also provides an ab initio way to describe the ion kinetic energies, temperatures, and other properties by a series of successive approximations. PMID- 15267846 TI - Accurate intermolecular ground state potential of the Ne-N2 van der Waals complex. AB - Ab initio ground state potential energy surfaces are obtained from interaction energies calculated with the coupled cluster singles and doubles model including connected triples corrections [CCSD(T)] and the aug-cc-pVXZ (X=5,Q,T,D) basis sets augmented with two different sets of midbond functions (denoted 33221 and 33211). The aug-cc-pV5Z-33221 surface is characterized by a T-shaped 49.5 cm(-1) minimum at Re=3.38 Angstroms and a linear saddle point at 3.95 Angstroms with De=36.6 cm(-1). These results agree well with the values provided by the accurate semiempirical potentials available. The rovibronic spectroscopic properties are determined and compared to the available experimental data and previous theoretical results. We study the basis set convergence of the intermolecular potentials and the rotational frequencies. The aug-cc-pVTZ basis sets provide reasonable binding parameters, but seem not to be converged enough for the evaluation of the microwave spectra. The aug-cc-pVQZ basis sets considerably improve the triple zeta results. The differences between the results obtained with the aug-cc-pVTZ-33221 basis set surface and those with the aug-cc-pVQZ-33221 are smaller than those of the corresponding bases with the set of 33211 midbond functions. The aug-cc-pVQZ surfaces are close to the aug-cc-pV5Z, that are expected to be close to convergence. With our best surfaces the errors in the frequencies with respect to the accurate experimental results go down to 0.6%. PMID- 15267847 TI - Equation of state of a seven-dimensional hard-sphere fluid. Percus-Yevick theory and molecular-dynamics simulations. AB - Following the work of Leutheusser [Physica A 127, 667 (1984)], the solution to the Percus-Yevick equation for a seven-dimensional hard-sphere fluid is explicitly found. This allows the derivation of the equation of state for the fluid taking both the virial and the compressibility routes. An analysis of the virial coefficients and the determination of the radius of convergence of the virial series are carried out. Molecular-dynamics simulations of the same system are also performed and a comparison between the simulation results for the compressibility factor and theoretical expressions for the same quantity is presented. PMID- 15267848 TI - Molecular alignment in a liquid induced by a nonresonant laser field: Molecular dynamics simulation. AB - We carried out molecular dynamics (MD) simulations for a dilute aqueous solution of pyrimidine in order to investigate the mechanisms of field-induced molecular alignment in a liquid phase. An anisotopically polarizable molecule can be aligned in a liquid phase by the interaction with a nonresonant intense laser field. We derived the effective forces induced by a nonresonant field on the basis of the concept of the average of the total potential over one optical cycle. The results of MD simulations show that a pyrimidine molecule is aligned in an aqueous solution by a linearly polarized field of light intensity I approximately 10(13) W/cm2 and wavelength lambda = 800 nm. The temporal behavior of field-induced alignment is adequately reproduced by the solution of the Fokker Planck equation for a model system in which environmental fluctuations are represented by Gaussian white noise. From this analysis, we have revealed that the time required for alignment in a liquid phase is in the order of the reciprocals of rotational diffusion coefficients of a solute molecule. The degree of alignment is determined by the anisotropy of the polarizability of a molecule, light intensity, and temperature. We also discuss differences between the mechanisms of optical alignment in a gas phase and a liquid phase. PMID- 15267849 TI - Multicomponent dynamical nucleation theory and sensitivity analysis. AB - Vapor to liquid multicomponent nucleation is a dynamical process governed by a delicate interplay between condensation and evaporation. Since the population of the vapor phase is dominated by monomers at reasonable supersaturations, the formation of clusters is governed by monomer association and dissociation reactions. Although there is no intrinsic barrier in the interaction potential along the minimum energy path for the association process, the formation of a cluster is impeded by a free energy barrier. Dynamical nucleation theory provides a framework in which equilibrium evaporation rate constants can be calculated and the corresponding condensation rate constants determined from detailed balance. The nucleation rate can then be obtained by solving the kinetic equations. The rate constants governing the multistep kinetics of multicomponent nucleation including sensitivity analysis and the potential influence of contaminants will be presented and discussed. PMID- 15267850 TI - Diffusion-influenced reversible geminate recombination in one dimension. III. Field effect on the excited-state reaction. AB - We obtain exact analytic solutions of the diffusion-influenced excited-state reversible geminate recombination reaction, A* + B<-->(AB)*, with two different lifetimes and quenching under the influence of a constant external field in one dimension. These fundamental solutions generalize two previous results [Kim et al., J. Chem. Phys. 111, 3791 (1999); 114, 3905 (2001)] and provide us with the insight necessary to analyze their specific relations and asymptotic kinetic transition behaviors. We find that the number of kinetic transitions can be changed due to interplay between the field strength and lifetimes. Unlike the previous works, the number of lifetime dependent transitions is found to be one or zero. On the other hand, the number of the field dependent transitions becomes two, one, or zero. We find a new pattern of kinetic transition e(t)-->t(-1/2)- >e(t) when there is only one field dependent transition. PMID- 15267851 TI - Molecular dynamics studies of melting and solid-state transitions of ammonium nitrate. AB - Molecular dynamics simulations are used to calculate the melting point and some aspects of high-temperature solid-state phase transitions of ammonium nitrate (AN). The force field used in the simulations is that developed by Sorescu and Thompson [J. Phys. Chem. A 105, 720 (2001)] to describe the solid-state properties of the low-temperature phase-V AN. Simulations at various temperatures were performed with this force field for a 4 x 4 x 5 supercell of phase-II AN. The melting point of AN was determined from calculations on this supercell with voids introduced in the solid structure to eliminate superheating effects. The melting temperature was determined by calculating the density and the nitrogen nitrogen radial distribution functions as functions of temperature. The melting point was predicted to be in the range 445 +/- 10 K, in excellent agreement with the experimental value of 442 K. The computed temperature dependences of the density, diffusion, and viscosity coefficient for the liquid are in good agreement with experiment. Structural changes in the perfect crystal at various temperatures were also investigated. The ammonium ions in the phase-II structure are rotationally disordered at 400 K. At higher temperatures, beginning at 530 K, the nitrate ions are essentially rotationally unhindered. The density and radial distribution functions in this temperature range show that the AN solid is superheated. The rotational disorder is qualitatively similar to that observed in the experimental phase-II to phase-I solid-state transition. PMID- 15267852 TI - Ab initio quantum dynamics with very weak van der Waals interactions: Structure and stability of small Li2(1Sigmag+)-(He)n clusters. AB - The potential energy surface (PES) for the interaction between Li2(1Sigmag+) and 4He has been computed using an accurate, post-Hartree-Fock quantum calculation for its ground electronic state. The orientational anisotropy of the forces and the interplay between repulsive and attractive effects within the PES are analyzed to extract information on the possible existence of bound states in the triatomic system. The structures of a few of the Li2(He)n small clusters are examined by comparing a classical approach with a full quantum one to generate bound configurations and to extract information on the possible spatial arrangements of the smaller clusters via a vis the location of the Li2 dopant. Some significant consequences on the Li2 behavior in larger clusters and droplets are drawn from the above findings. PMID- 15267853 TI - First-principle molecular dynamics of the Berry pseudorotation: insights on 19F NMR in SF4. AB - First-principles [density-functional theory (DFT)] molecular-dynamic simulations of the Berry pseudorotation mechanism in SF4 were performed using the atom centered density-matrix propagation method. The reaction was monitored by following the chemical shieldings of the fluorine atoms, computed on snapshots along the trajectories. In particular we compared the results obtained using a standard functional based on the generalized gradient approximation with those issuing from its hybrid Hartree-Fock-DFT counterpart using a number of basis sets. Our results show that both the basis set and the functional choice rule the quality of the molecular properties monitored as well as the trajectory over the potential-energy surface. PMID- 15267854 TI - On the structural and transport properties of the soft sticky dipole and related single-point water models. AB - The density maximum and temperature dependence of the self-diffusion constant were investigated for the soft sticky dipole (SSD) water model and two related reparametrizations of this single-point model. A combination of microcanonical and isobaric-isothermal molecular dynamics simulations was used to calculate these properties, both with and without the use of reaction field to handle long range electrostatics. The isobaric-isothermal simulations of the melting of both ice-Ih and ice-Ic showed a density maximum near 260 K. In most cases, the use of the reaction field resulted in calculated densities which were significantly lower than experimental densities. Analysis of self-diffusion constants shows that the original SSD model captures the transport properties of experimental water very well in both the normal and supercooled liquid regimes. We also present our reparametrized versions of SSD for use both with the reaction field or without any long-range electrostatic corrections. These are called the SSD/RF and SSD/E models, respectively. These modified models were shown to maintain or improve upon the experimental agreement with the structural and transport properties that can be obtained with either the original SSD or the density corrected version of the original model (SSD1). Additionally, a novel low-density ice structure is presented which appears to be the most stable ice structure for the entire SSD family. PMID- 15267855 TI - A density-functional theory study of the confined soft ellipsoid fluid. AB - A system of soft ellipsoid molecules confined between two planar walls is studied using classical density-functional theory. Both the isotropic and nematic phases are considered. The excess free energy is evaluated using two different Ansatze and the intermolecular interaction is incorporated using two different direct correlation functions (DCF's). The first is a numerical DCF obtained from simulations of bulk soft ellipsoid fluids and the second is taken from the Parsons-Lee theory. In both the isotropic and nematic phases the numerical DCF gives density and order parameter profiles in reasonable agreement with simulation. The Parsons-Lee DCF also gives reasonable agreement in the isotropic phase but poor agreement in the nematic phase. PMID- 15267856 TI - Critical fluctuations near the consolute point of n-pentanol-nitromethane. An ultrasonic spectrometry, dynamic light scattering, and shear viscosity study. AB - Ultrasonic attenuation spectra, the shear viscosity, and the mutual diffusion coefficient of the n-pentanol-nitromethane mixture of critical composition have been measured at different temperatures near the critical temperature. The noncritical background contribution, proportional to frequency, to the acoustical attenuation-per-wavelength spectra has been determined and subtracted from the total attenuation to yield the critical contribution. When plotted versus the reduced frequency, with the relaxation rate of order-parameter fluctuations from the shear viscosity and diffusion coefficient measurements, the critical part in the sonic attenuation coefficient displays a scaling function which nicely fits to the data for the critical system 3-methylpentane-nitromethane and also to the empirical scaling function of the Bhattacharjee-Ferrell dynamic scaling theory. The scaled half-attenuation frequency follows from the experimental data as Omega(1/2)emp= 1.8+/-0.1. The relaxation rate of order-parameter fluctuation shows power-law behavior with the theoretically predicted universal exponent and the extraordinary high amplitude Gammao= (187+/-2) x 10(9) s(-1). The amount of the adiabatic coupling constant /g/= 0.03, as estimated from the amplitude of the critical contribution to the acoustical spectra, is unusually small. PMID- 15267857 TI - Thermally activated escape rate for the Brownian motion of a fixed axis rotator in a double well potential for all values of the dissipation. AB - The extension of the Kramers theory of the escape rate of a Brownian particle from a potential well to the entire range of damping proposed by Mel'nikov and Meshkov [J. Chem, Phys. 85, 1018 (1986)] is applied to the rotational Brownian motion of fixed axis rotators in a double well cosine potential. The procedure yields an expression for the Kramers escape rate valid for all values of the dissipation including the very low damping (VLD), very high damping (VHD), and crossover regimes. This equation provides a good asymptotic estimate of the correlation time tau per pendicular of the longitudinal dipole moment correlation function calculated by solving the underlying Langevin equation using the matrix continued fraction method. Moreover, for low barriers, where the Mel'nikov and Meshkov approach is not applicable, analytic equations for tau in the VLD and VHD limits are derived and a simple extrapolating equation that is valid for all values of the damping is proposed. PMID- 15267858 TI - Quantum chemical calculation of infrared spectra of acidic groups in chabazite in the presence of water. AB - The changes in the spectra of the acidic group in chabazite are studied by quantum chemical calculations. The zeolite is modeled by two clusters consisting of eight tetrahedral atoms arranged in a ring and seven tetrahedral atoms coordinated around the zeolite OH group. The potential energy and dipole surfaces were constructed from the zeolite OH stretch, in-plane and out-of-plane bending coordinates, and the intermolecular stretch coordinate that corresponds to a movement of the water molecule as a whole. Both the anharmonicities of the potential energy and dipole were taken into account by calculation of the frequencies and intensities. The matrix elements of the vibrational Hamiltonian were calculated within the discrete variable representation basis set. We have assigned the experimentally observed frequencies at approximately 2900, approximately 2400, and approximately 1700 cm(-1) to the strongly perturbed zeolite OH vibrations caused by the hydrogen bonding with the water molecule. The ABC triplet is a Fermi resonance of the zeolite OH stretch mode with the overtone of the in-plane bending (the A band) and the overtone of the out-of-plane bending (the C band). In the B band the stretch is also coupled with the second overtone of the out-of-plane bending. The frequencies at approximately 3700 and approximately 3550 cm(-1) we have assigned to the OH stretch frequencies of a slightly perturbed water molecule. PMID- 15267859 TI - Ordered binary crystal phases of Lennard-Jones mixtures. AB - The lattice energies at zero temperature are calculated, using Lennard-Jones interactions, for a large number of crystal structures associated with ordered binary compounds. In units of the AA interaction length and strength (i.e., sigmaAA= epsilonAA= 1.0) we examine the lowest energy structures, including coexisting phases, across the space of cross-species interactions 0.6< or = sigmaAB< or = 1.1 and 1.0< or = epsilonAB< or = 2.0. The remaining parameters sigmaBB= 0.88 and epsilonBB= 0.5 are chosen so that the parameter space studied includes the space of binary glass-forming alloys. In addition to some large unit cell structures such as Ni3P and PuBr3 appearing among the lowest lattice energies, a number of low-energy structures based on close-packed lattices are found that do not correspond to any experimentally observed crystals. The prevalence and stability of metastable crystal phases at the compositions AB, A2B, and A3B is examined. PMID- 15267860 TI - Dynamical behavior of one-dimensional water molecule chains in zeolites: nanosecond time-scale molecular dynamics simulations of bikitaite. AB - Nanosecond scale molecular dynamics simulations of the behavior of the one dimensional water molecule chains adsorbed in the parallel nanochannels of bikitaite, a rare lithium containing zeolite, were performed at different temperatures and for the fully and partially hydrated material. New empirical potential functions have been developed for representing lithium-water interactions. The structure and the vibrational spectrum of bikitaite were in agreement both with experimental data and Car-Parrinello molecular dynamics results. Classical molecular dynamics simulations were extended to the nanosecond time scale in order to study the flip motion of water molecules around the hydrogen bonds connecting adjacent molecules in the chains, which has been observed by NMR experiments, and the dehydration mechanism at high temperature. Computed relaxation times of the flip motion follow the Arrhenius behavior found experimentally, but the activation energy of the simulated system is slightly underestimated. Based on the results of the simulations, it may be suggested that the dehydration proceeds by a defect-driven stepwise diffusion. The diffusive mechanism appears as a single-file motion: the molecules never pass one another, even at temperatures as high as about 1000 K, nor can they switch between different channels. However, the mean square displacement (MSD) of the molecules, computed with respect to the center of mass of the simulated system, shows an irregular trend from which the single-file diffusion cannot be clearly evidenced. If the MSDs are evaluated with respect to the center of mass of the molecules hosted in each channel, the expected dependence on the square root of time finally appears. PMID- 15267861 TI - Advantages of polarized two-beam second-harmonic generation in precise characterization of thin films. AB - Polarized second-harmonic generation using two fundamental beams, instead of one, offers significant advantages for characterizing nonlinear optical thin films. The technique is more precise and allows the internal consistency of the results to be verified. The superiority of the two-beam arrangement over the traditional single-beam arrangement is demonstrated by determining the susceptibility tensors of Langmuir-Blodgett films. We show that, for a well-understood reference sample, the results obtained using two fundamental beams agree qualitatively with those obtained with a single fundamental beam, but are more precise. In a more complicated situation, however, the single-beam technique appears to work well but yields results that are, in fact, incorrect. The two-beam technique, instead, yields clearly inconsistent results, thereby highlighting systematic errors in the experimental arrangement or in the theoretical model used to interpret the results. PMID- 15267862 TI - Energetics, transition states, and intrinsic reaction coordinates for reactions associated with O(3P) processing of hydrocarbon materials. AB - Electronic structure calculations based on multiconfiguration wave functions are used to investigate a set of archetypal reactions relevant to O(3P) processing of hydrocarbon molecules and surfaces. These include O(3P) reactions with methane and ethane to give OH plus methyl or ethyl radicals, O(3P) + ethane to give CH3O + CH3, and secondary reactions of the OH product radical with ethane and the ethyl radical. Geometry optimization is carried out with CASSCF/cc-pVTZ for all reactions, and with CASPT2/cc-pVTZ for O(3P) + methane/ethane. Single-point energy corrections are applied with CASPT2, CASPT3, and MRCI + Q with the cc-pVTZ and cc-pVQZ basis sets, and the energies extrapolated to the complete basis set limit (CBL). Where comparison of computed barriers and energies of reaction with experiment is possible, the agreement is good to excellent. The best agreement (within experimental error) is found for MRCI + Q/CBL applied to O(3P) + methane. For the other reactions, CASPT2/CBL and MRCI + Q/CBL predictions differ from experiment by 1-5 kcal/mol for 0 K enthalpies of reaction, and are within 1 kcal/mol of the best-estimate experimental range of 0 K barriers for O(3P) + ethane and OH + ethane. The accuracy of MRCI + Q/CBL is limited mainly by the quality of the active space. CASPT2/CBL barriers are consistently lower than MRCI + Q/CBL barriers with identical reference spaces. PMID- 15267863 TI - Affinity distribution functions in multicomponent heterogeneous adsorption. Analytical inversion of isotherms to obtain affinity spectra. AB - An analytical approach for the interpretation of multicomponent heterogeneous adsorption or complexation isotherms in terms of multidimensional affinity spectra is presented. Fourier transform, applied to analyze the corresponding integral equation, leads to an inversion formula which allows the computation of the multicomponent affinity spectrum underlying a given competitive isotherm. Although a different mathematical methodology is used, this procedure can be seen as the extension to multicomponent systems of the classical Sips's work devoted to monocomponent systems. Furthermore, a methodology which yields analytical expressions for the main statistical properties (mean free energies of binding and covariance matrix) of multidimensional affinity spectra is reported. Thus, the level of binding correlation between the different components can be quantified. It has to be highlighted that the reported methodology does not require the knowledge of the affinity spectrum to calculate the means, variances, and covariance of the binding energies of the different components. Nonideal competitive consistent adsorption isotherm, widely used in metal/proton competitive complexation to environmental macromolecules, and Frumkin competitive isotherms are selected to illustrate the application of the reported results. Explicit analytical expressions for the affinity spectrum as well as for the matrix correlation are obtained for the NICCA case. PMID- 15267864 TI - Two-dimensional Monte Carlo simulations of ionic and nonionic silane self assembly on hydrophilic surfaces. AB - Aqueous chemistries have recently been shown to be useful for the deposition of hydrophobic films of nonionic and cationic silanes on hydrophilic substrates for the prevention of stiction in MEMS. The Monte Carlo method is used to simulate in two dimensions the self-assembly of silane films on a hydrophilic surface. We investigate the impact of charged group in cationic silane on the overall structure of the films. We characterize the film structure with spatial pair correlations at each molecular layer of the deposited films. The simulations reveal long-range correlations for the film of cationic silanes. Based on our two dimensional simulations, we report an average "most probable" structure for the films of nonionic and cationic silanes. PMID- 15267865 TI - High density gradients in the (square root 3 x square root 3)R30 degrees-CO layer on Ru(0001). AB - The coverage regime just beyond 0.33 ML, representative of a perfectly ordered (square root 3 x square root 3)R30 degrees-CO layer on Ru(0001), has been investigated using infrared-absorption spectroscopy. Different isotopic mixtures of CO have been employed to derive a profound understanding of structural properties of such layers. It is found that extra CO molecules incorporated into the (square root 3 x square root 3)R30 degrees-CO layer affect their nearest neighbor molecules only, and the associated density gradient extends over no more than a few angstroms. Contrary to existing belief, the model system CO on Ru(0001) does not represent a case of an unusually shallow adsorption potential corrugation. Rather, CO experiences an exceptionally strong site preference when adsorbed on Ru(0001). Annealing causes the local distortion of the overlattice to propagate laterally, most probably in a density wave-like manner. Incipient motion on the atomic scale thereby has been detected by means of isotopic labeling of inequivalent molecules within the high density areas. All major conclusions are based on observations of (isotopically labeled) minority CO species which feature negligible dynamical lateral coupling. The majority CO species, on the other hand, provide laterally averaged, unspecific information on the status of the layer. PMID- 15267866 TI - Thermodynamic properties of the Cu-Au system using a face-centered-cubic lattice model with a renormalized potential. AB - A Monte Carlo simulation is carried out to study thermodynamic properties of Cu Au alloys using a face-centered-cubic (fcc) lattice-gas model. To obtain quantitatively accurate results, a Finnis-Sinclair-type potential, which has been widely used for molecular dynamics (MD) simulations, is employed. To overcome some shortcomings of lattice-gas models such as neglecting vibrational entropy, the potential is mapped onto the fcc lattice using the renormalization technique. The renormalized potential gives an improved Cu-Au phase diagram compared to the original MD potential applied directly on the lattice. PMID- 15267867 TI - Ultrafast chemical interface scattering as an additional decay channel for nascent nonthermal electrons in small metal nanoparticles. AB - The use of 4.2 nm gold nanoparticles wrapped in an adsorbates shell and embedded in a TiO2 metal oxide matrix gives the opportunity to investigate ultrafast electron-electron scattering dynamics in combination with electronic surface phenomena via the surface plasmon lifetimes. These gold nanoparticles (NPs) exhibit a large nonclassical broadening of the surface plasmon band, which is attributed to a chemical interface damping. The acceleration of the loss of surface plasmon phase coherence indicates that the energy and the momentum of the collective electrons can be dissipated into electronic affinity levels of adsorbates. As a result of the preparation process, gold NPs are wrapped in a shell of sulfate compounds that gives rise to a large density of interfacial molecules confined between Au and TiO2, as revealed by Fourier-transform-infrared spectroscopy. A detailed analysis of the transient absorption spectra obtained by broadband femtosecond transient absorption spectroscopy allows separating electron-electron and electron-phonon interaction. Internal thermalization times (electron-electron scattering) are determined by probing the decay of nascent nonthermal electrons (NNEs) and the build-up of the Fermi-Dirac electron distribution, giving time constants of 540 to 760 fs at 0.42 and 0.34 eV from the Fermi level, respectively. Comparison with literature data reveals that lifetimes of NNEs measured for these small gold NPs are more than four times longer than for silver NPs with similar sizes. The surprisingly long internal thermalization time is attributed to an additional decay mechanism (besides the classical e-e scattering) for the energy loss of NNEs, identified as the ultrafast chemical interface scattering process. NNEs experience an inelastic resonant scattering process into unoccupied electronic states of adsorbates, that directly act as an efficient heat bath, via the excitation of molecular vibrational modes. The two temperature model is no longer valid for this system because of (i) the temporal overlap between the internal and external thermalization process is very important; (ii) a part of the photonic energy is directly transferred toward the adsorbates (not among "cold" conduction band electrons). These findings have important consequence for femtochemistry on metal surfaces since they show that reactions can be initiated by nascent nonthermal electrons (as photoexcited, out of a Fermi-Dirac distribution) besides of the hot electron gas. PMID- 15267868 TI - "Triplet-excited region" in polyene oligomers revisited: Pariser-Parr-Pople model studied with the density matrix renormalization group method. AB - We have carried out density matrix renormalization group calculations on the T1 state of linear polyenes applying the Pariser-Parr-Pople (PPP) Model. The geometry optimization for the polyene oligomers C(2n)H(2n+2) (n = 4,5,6,...,15) shows that the S0 to T1 excitation region is composed of a soliton-antisoliton pair located symmetrically away from the center of the chain and leads to single- and double-bond interconversions in between. The distance between the soliton and antisoliton centers in T1 state changes with the length of the chain, contradictory to earlier conclusions obtained with PPP-SDCI or ab initio SCI methods. The inconsistency most possibly comes from the insufficient consideration of the electron correlations in small-scale CI methods. PMID- 15267869 TI - "Swiss-cheese" polyelectrolyte gels as media with extremely inhomogeneous distribution of charged species. AB - "Swiss-cheese" polyelectrolyte gels (i.e., gels containing a regular set of closed spherical pores) are considered as a suitable system for modeling of a medium with extremely inhomogeneous distribution of charged species. It is shown that the inhomogeneous distribution of ions in Swiss-cheese polyelectrolyte gels can be reached simply by immersion of the gels in an aqueous solution of charged species (e.g., low-molecular 1-1 salt or multivalent ions and macroions charged likely to the gel chains). If a polymer gel is kept in such a solution for a long time, the concentration of ions within relatively big voids becomes equal to that in external solution. On the other hand, due to the Donnan effect the ion's concentration in polymer matrix is always lower than that in external solution. As a result the multivalent ions distribute between water voids and polymer matrix. The extent of this distribution is characterized by partition coefficient kD (determined as ratio kD = n(s)(void)/n(s)(mat) of the concentrations n(s)(void) and n(s)(mat) of ions in water voids and in polymer matrix, correspondingly). It is shown that the partition coefficient kD can be larger than 10 for low-molecular salt, reaches 10(3) for bivalent ions, and is higher than 10(6) for tetravalent ions. In the case of polymer macroions the partition coefficient kD tends to infinity. Our calculations show that the lower limit of characteristic scales of heterogeneity (determined by water voids size starting from which the condition of total electroneutrality is fulfilled and effect of partition is the most pronounced) can be equal to tens of nanometers. PMID- 15267870 TI - A Langevin dynamics study of mobile filler particles in phase-separating binary systems. AB - The dynamics of phase separation in a simple binary mixture containing mobile filler particles that are preferentially wet by one of the two components is investigated systematically via Langevin simulations in two dimensions. We found that while the filler particles reduce the growth rate of spinodal decomposition, the domain growth remains essentially identical to that of the pure binary mixture. The growth rate diminishes as either the filler particles concentration is increased or their diffusivity is decreased. PMID- 15267871 TI - Polymer-particle mixtures: depletion and packing effects. AB - The structure of polymers in the vicinity of spherical colloids is investigated by Monte Carlo simulations and integral equation theory. Polymers are represented by a simple bead-spring model; only repulsive Lennard-Jones interactions are taken into account. Using advanced trial moves that alter chain connectivity, depletion and packing effects are analyzed as a function of chain length and density, both at the bond and the chain level. Chain ends segregate to the colloidal surface and polymer bonds orient parallel to it. In the dilute regime, the polymer chain length governs the range of depletion and has a negligible influence on monomer packing in dense polymer melts. Polymers adopt an ellipsoidal shape, with the larger axis parallel to the surface of the particle, as they approach larger colloids. The dimensions are perturbed within the range of the depletion layer. PMID- 15267872 TI - Theory of counter-ion condensation on flexible polyelectrolytes: adsorption mechanism. AB - A new model is presented for counterion distribution around flexible polyelectrolytes by considering (i) free energy of the polyelectrolyte chain, (ii) translational entropy of adsorbed counterions, (iii) adsorption energy, (iv) translational entropy of unadsorbed counterions, (v) fluctuations of dissociated ions, and (vi) correlation among ion-pairs formed by adsorbed counterions on the polymer. The effective charge and size of the polymer are calculated self consistently. The degree of ionization f of the polymer decreases continuously with 1/epsilonT (epsilon and T are the dielectric constant of the solvent and temperature, respectively), depending sensitively on local dielectric heterogeneity. Further, f decreases with an increase in salt concentration, monomer concentration, or chain flexibility. The polymer size, accompanying the changes in f, depends nonmonotonically on 1/epsilonT. The predictions of the model are consistent with all trends observed previously in simulations and are distinctly different from the Manning argument for rodlike chains. PMID- 15267873 TI - Diffusion and trapping in a suspension of spheres with simultaneous reaction in the continuous phase. AB - Much progress has been made in modeling the reaction of Brownian particles with spherical traps. Previously, work has focused on the effective reaction rate of systems of particles that diffuse freely until they are trapped by spheres in the dispersion. A particularly effective and efficient method to describe the reacting system is based on first-passage time distributions, from which an effective reaction rate coefficient of the suspension can be determined. The analysis presented here addresses reaction and diffusion in systems in which particles can undergo reaction in the continuous phase as well as reaction at the sphere surface. The first-passage method is extended to allow reaction or decay of the diffusing species in the continuous phase. The diffusion path is divided into a series of first-passage regions and is considered the probability of the particle being consumed in each of these regions. This allows the determination of the total reaction rate of the suspension (continuous phase reaction plus trapping) and the relative consumption rate in each phase. The extended method is applied to a model system of concentric spheres with a known continuum solution. It is shown to be accurate for consumption of reactant in the continuous phase from approximately 0 to approximately 100%. The method then is applied to a suspension of spheres. PMID- 15267874 TI - Microscopic theory of rubber elasticity. AB - A microscopic integral equation theory of elasticity in polymer liquids and networks is developed which addresses the nonclassical problem of the consequences of interchain repulsive interactions and packing correlations on mechanical response. The theory predicts strain induced softening, and a nonclassical intermolecular contribution to the linear modulus. The latter is of the same magnitude as the classical single chain entropy contribution at low polymer concentrations, but becomes much more important in the melt state, and dominant as the isotropic-nematic liquid crystal phase transition is approached. Comparison of the calculated stress-strain curve and induced nematic order parameter with computer simulations show good agreement. A nearly quadratic dependence of the linear elastic modulus on segmental concentration is found, as well as a novel fractional power law dependence on degree of polymerization. Quantitative comparison of the theory with experiments on polydimethylsiloxane networks are presented and good agreement is found. However, a nonzero modulus in the long chain limit is not predicted since quenched chemical crosslinks and trapped entanglements are not explicitly taken into account. The theory is generalizable to treat the structure, thermodynamics and mechanical response of nematic elastomers. PMID- 15267875 TI - Influence of confinement on the vibrational density of states and the Boson peak in a polymer glass. AB - We have performed a normal-mode analysis on a glass forming polymer system for bulk and free-standing film geometries prepared under identical conditions. It is found that for free-standing film glasses, the normal-mode spectrum exhibits significant differences from the bulk glass with the appearance of an additional low-frequency peak and a higher intensity at the Boson peak frequency. A detailed eigenvector analysis shows that the low-frequency peak corresponds to a shear horizontal mode which is predicted by continuum theory. The peak at higher frequency (Boson peak) corresponds to motions that are correlated over a length scale of approximately twice the interaction site diameter. These observations shed some light on the microscopic dynamics of glass formers, and help explain decreasing fragility that arises with decreasing thickness in thin films. PMID- 15267876 TI - Ab initio study of the electronic and structural properties of the crystalline polyethyleneimine polymer. AB - Polyethyleneimine is a very interesting polymer, not only for its extensive use in biological applications, but also for its crystal structure as a double stranded helix in the anhydrous state. In order to elucidate the electronic bulk properties of the crystalline (or linear) polyethyleneimine built from ethylenediamine molecules in anhydrous conditions, we performed ab initio density functional theory calculations on water-free molecular crystal structures. The resulting polymer is a semiconductor with a small band gap: Eg = 0.40 eV. PMID- 15267877 TI - Cubatic liquid-crystalline behavior in a system of hard cuboids. AB - The lyotropic phase behavior of cuboidal particles was investigated via Monte Carlo simulations. Hard cubes were approximated by suitably shaped clusters of hard spheres. Changes in concentration and structure of the system were monitored as a function of osmotic pressure P* (imposed in an isobaric ensemble). As expected, an isotropic phase prevailed at low concentrations (low P*) and a crystalline phase formed at high concentrations (high P*). A third distinct phase was also observed for an intermediate range of concentrations (approximately marked by breaks in the P* versus concentration curve). The structure of this mesophase was characterized both visually and analytically by calculating radial distribution functions and order parameters. It was found that such a mesophase exhibits orientational ordering along three axes (cubatic order) but significant translational disorder, thus having a structure clearly distinct from both isotropic and crystalline phases. PMID- 15267878 TI - Reappraisal of four different approaches for finding the mean reaction time in the multi-trap variant of the Adam-Delbruck problem. AB - Adam and Delbruck argued that the dimensionality of the diffusion space determines the average lifetime of a diffusing particle confined to a region with a central trap. Doubts have often been aired as to whether their calculation is relevant to real biological systems, where the number of traps is usually much larger than unity; or whether the rate enhancement is merely a manifestation of an increase in the concentration of the traps; or whether the diverse multi-trap versions of their expression for the mean lifetime in two dimensions are trustworthy. These issues are addressed, and the long-standing problem of finding the low-density limit of trapping time in two dimensions solved, by examining previous treatments of the problem, and by carrying out simulations of two dimensional systems in which the particles undergo a Pearsonian random walk, and the traps are distributed randomly or on a square lattice. The mean lifetimes are found to be different in the two situations, and it is concluded that the neglect of this aspect lies at the root of the conflict between some of the existing expressions for the mean lifetime. Relations expressing the mean lifetime as a function of the concentration of the traps are presented together with a discussion of their applicability. PMID- 15267879 TI - Complexation of semiflexible chains with oppositely charged cylinder. AB - We study the complexation of long thin semiflexible polymer chains with an oppositely charged cylinder. Starting from the linear Poisson-Boltzmann equation, we calculate the electrostatic potential and the energy of such a charge distribution. We find that sufficiently flexible chains prefer to wrap around the cylinder in a helical manner, when their charge density is smaller than that of the cylinder. The optimal value of the helical pitch is found by minimization of the sum of electrostatic and bending energies. The dependence of the pitch on the number of chains, their rigidity, and salt concentration in solution is analyzed. We discuss our results in the light of recent experiments on DNA complexation with cylindrical dendronized polymers. PMID- 15267880 TI - Second-order nonlinear optical coefficient of polyphosphazene-based materials: a theoretical study. AB - The second-order nonlinear optical coefficient of polyphosphazene oligomers of increasing size has been determined by using ab initio methods taking into account electron correlation and frequency dispersion effects. The calculated first hyperpolarizability per unit cell converges rapidly with respect to chain length. It attains an amplitude of about one-third of the one of classical push pull systems. This amplitude can be strongly increased by replacing the nitrogen of the backbone by silicon. The effects of the side groups (H, CH3, F, Cl, Br, and OH) on the first hyperpolarizability have been investigated as well. The different results have been rationalized in terms of alternations of bond lengths and atomic charges. PMID- 15267881 TI - On the role of block copolymer additives for calcium carbonate crystallization: small angle neutron scattering investigation by applying contrast variation. AB - The role of the double-hydrophilic block copolymer poly(ethylen glycol)-block poly(methacrylic acid) (PEG-b-PMAA) on the morphogenesis of calcium carbonate (CaCO3) was studied by applying the contrast variation small angle neutron scattering technique. The morphology and size of CaCO3 crystals is strongly affected by the addition of PEG-b-PMAA. In order to determine the partial scattering functions of the polymer and CaCO3 mineral, we developed both an experimental and theoretical approach with a sophisticated method of their determination from the scattering intensity. Partial scattering functions give detailed information for each component. In particular, the partial scattering function of the polymer, Spp, shows a monotonic slope with Q(-2 to -3) where the scattering vector Q is low (Q < 0.01 Angstrom(-1)), which is a clear evidence that the polymer within the CaCO3 mineral has a mass fractal dimension. The other partial scattering functions reflected the geometry of the CaCO3 particles or the "interaction" of polymer and CaCO3 on a microscopic scale, which leads to a coherent view with Spp. PMID- 15267882 TI - Electric field gradients from shifted-nucleus calculations: an alternative to the point charge nuclear quadrupole moment model. PMID- 15267883 TI - Comment on "Theoretical evaluation of hydrogen storage capacity in pure carbon nanostructures" [J. Chem. Phys. 119, 2376 (2003)]. PMID- 15267885 TI - Bias in the temperature of helium nanodroplets measured by an embedded rotor. AB - The rovibrational spectra of molecules dissolved in liquid 4He nanodroplets display rotational structure. Where resolved, this structure has been used to determine a temperature that has been assumed to equal that of the intrinsic excitations of the helium droplets containing the molecules. Consideration of the density of states as a function of energy and total angular momentum demonstrates that there is a small but significant bias of the rotor populations that make the temperature extracted from a fit to its rotational level populations slightly higher than the temperature of the ripplons of the droplet. This bias grows with both the total angular momentum of the droplet and with the moment of inertia of the solute molecule. PMID- 15267886 TI - From transition paths to transition states and rate coefficients. AB - Transition states are defined as points in configuration space with the highest probability that trajectories passing through them are reactive (i.e., form transition paths between reactants and products). In the high-friction (diffusive) limit of Langevin dynamics, the resulting ensemble of transition states is shown to coincide with the separatrix formed by points of equal commitment (or splitting) probabilities for reaching the product and reactant regions. Transition states according to the new criterion can be identified directly from equilibrium trajectories, or indirectly by calculating probability densities in the equilibrium and transition-path ensembles using umbrella and transition-path sampling, respectively. An algorithm is proposed to calculate rate coefficients from the transition-path and equilibrium ensembles by estimating the frequency of transitions between reactants and products. PMID- 15267887 TI - The impact of the self-interaction error on the density functional theory description of dissociating radical cations: ionic and covalent dissociation limits. AB - Self-interaction corrected density functional theory was used to determine the self-interaction error for dissociating one-electron bonds. The self-interaction error of the unpaired electron mimics nondynamic correlation effects that have no physical basis where these effects increase for increasing separation distance. For short distances the magnitude of the self-interaction error takes a minimum and increases then again for decreasing R. The position of the minimum of the magnitude of the self-interaction error influences the equilibrium properties of the one-electron bond in the radical cations H2+ (1), B2H4+ (2), and C2H6+ (3), which differ significantly. These differences are explained by hyperconjugative interactions in 2 and 3 that are directly reflected by the self-interaction error and its orbital contributions. The density functional theory description of the dissociating radical cations suffers not only from the self-interaction error but also from the simplified description of interelectronic exchange. The calculated differences between ionic and covalent dissociation for 1, 2, and 3 provide an excellent criterion for determining the basic failures of density functional theory, self-interaction corrected density functional theory, and other methods. Pure electronic, orbital relaxation, and geometric relaxation contributions to the self-interaction error are discussed. The relevance of these effects for the description of transition states and charge transfer complexes is shown. Suggestions for the construction of new exchange-correlation functionals are given. In this connection, the disadvantages of recently suggested self interaction error-free density functional theory methods are emphasized. PMID- 15267888 TI - A reinterpretation of the nature of the Fermi hole. AB - A reinterpretation of the Boyd-Coulson [R. J. Boyd and C. A. Coulson, J. Phys. B 7, 1805 (1974)] definition of the Fermi hole is presented. Through this reinterpretation, which makes no reference to the hypothetical Hartree level, we are able to show the essentially identical character of the Boyd-Coulson definition with the one based on a conditional probability analysis. The basis set dependence of the Fermi hole is emphasized and the effect of canonical, localized and delocalized Kohn-Sham and Hartree-Fock basis sets is examined for selected atoms and molecules. PMID- 15267889 TI - Mapped grid methods for long-range molecules and cold collisions. AB - The paper discusses ways of improving the accuracy of numerical calculations for vibrational levels of diatomic molecules close to the dissociation limit or for ultracold collisions, in the framework of a grid representation. In order to avoid the implementation of very large grids, Kokoouline et al. [J. Chem. Phys. 110, 9865 (1999)] have proposed a mapping procedure through introduction of an adaptive coordinate x subjected to the variation of the local de Broglie wavelength as a function of the internuclear distance R. Some unphysical levels ("ghosts") then appear in the vibrational series computed via a mapped Fourier grid representation. In the present work the choice of the basis set is reexamined, and two alternative expansions are discussed: Sine functions and Hardy functions. It is shown that use of a basis set with fixed nodes at both grid ends is efficient to eliminate "ghost" solutions. It is further shown that the Hamiltonian matrix in the sine basis can be calculated very accurately by using an auxiliary basis of cosine functions, overcoming the problems arising from numerical calculation of the Jacobian J(x) of the R-->x coordinate transformation. PMID- 15267890 TI - Fast vibrational self-consistent field calculations through a reduced mode-mode coupling scheme. AB - We present a new methodology to perform fast correlation-corrected vibrational self-consistent field (CC-VSCF) calculations using ab initio potential energy points calculated on the fly. Our method is based on the replacement of all electron basis sets with a pseudo-potential basis for heavy atoms, and on an efficient reduction of the number of pair-coupling elements used in the CC-VSCF procedure. The method is applied to several test systems: H2O, NH3, and CH4, where it proves to be efficient, providing a speedup factor of 2 compared to a standard CC-VSCF calculation. We also apply our technique to the simulation of the vibrational spectrum of ethane and show that very accurate results can be obtained with a substantial speedup for this system. PMID- 15267891 TI - Spectral differences in real-space electronic structure calculations. AB - Real-space grids for electronic structure calculations are efficient because the potential is diagonal while the second derivative in the kinetic energy may be sparsely evaluated with finite differences or finite elements. In applications to vibrational problems in chemical physics a family of methods known as spectral differences has improved finite differences by several orders of magnitude. In this paper the use of spectral differences for electronic structure is studied. Spectral differences are implemented in two electronic structure programs PARSEC and HARES which currently employ finite differences. Applications to silicon clusters and lattices indicate that spectral differences achieve the same accuracy as finite differences with less computational work. PMID- 15267892 TI - Semiclassical representations of electronic structure and dynamics. AB - We use a new formulation of the semiclassical coherent state propagator to derive and evaluate several different approximate representations of electron dynamics. For each representation we examine: (1) its ability to treat quantum effects and electron correlation, (2) its expected scaling with system size, and (3) the types of systems for which it can be used. We also apply two of the methods to a pair of model problems, namely the minimal basis electron dynamics in H2 and the magnetization dynamics in a cluster model of the Kagome lattice, in order to verify the feasibility of these approaches for realistic systems. Based on all these criteria, we find that the representation that takes the electron spins as the classical variables is particularly promising for the quantitative and qualitative description of large systems. PMID- 15267893 TI - Design and application of a multicoefficient correlation method for dispersion interactions. AB - A new multicoefficient correlation method (MCCM) is presented for the determination of accurate van der Waals interactions. The method utilizes a novel parametrization strategy that simultaneously fits to very high-level binding, Hartree-Fock and correlation energies of homo- and heteronuclear rare gas dimers of He, Ne, and Ar. The decomposition of the energy into Hartree-Fock and correlation components leads to a more transferable model. The method is applied to the krypton dimer system, rare gas-water interactions, and three-body interactions of rare gas trimers He3, Ne3, and Ar3. For the latter, a very high level method that corrects the rare-gas two-body interactions to the total binding energy is introduced. A comparison with high-level CCSD(T) calculations using large basis sets demonstrates the MCCM method is transferable to a variety of systems not considered in the parametrization. The method allows dispersion interactions of larger systems to be studied reliably at a fraction of the computational cost, and offers a new tool for applications to rare-gas clusters, and the development of dispersion parameters for molecular simulation force fields and new semiempirical quantum models. PMID- 15267894 TI - A new method for solving the quantum hydrodynamic equations of motion: application to two-dimensional reactive scattering. AB - The de Broglie-Bohm hydrodynamic equations of motion are solved using a meshless method based on a moving least squares approach and an arbitrary Lagrangian Eulerian frame of reference. A regridding algorithm adds and deletes computational points as needed in order to maintain a uniform interparticle spacing, and unitary time evolution is obtained by propagating the wave packet using averaged fields. The numerical instabilities associated with the formation of nodes in the reflected portion of the wave packet are avoided by adding artificial viscosity to the equations of motion. The methodology is applied to a two-dimensional model collinear reaction with an activation barrier. Reaction probabilities are computed as a function of both time and energy, and are in excellent agreement with those based on the quantum trajectory method. PMID- 15267895 TI - Mode-specific photoelectron scattering effects on CO2(+)(C2Sigmag+) vibrations. AB - Using high-resolution photoelectron spectroscopy, we have determined the energy dependent vibrational branching ratios for the symmetric stretch [v+ = (100)], bend [v+ = (010)], and antisymmetric stretch [v+ = (001)], as well as several overtones and combination bands in the 4sigmag(-1) photoionization of CO2. Data were acquired over the range from 20-110 eV, and this wide spectral coverage highlighted that alternative vibrational modes exhibit contrasting behavior, even over a range usually considered to be dominated by atomic effects. Alternative vibrational modes exhibit qualitatively distinct energy dependences, and this contrasting mode-specific behavior underscores the point that vibrationally resolved measurements reflect the sensitivity of the electron scattering dynamics to well-defined changes in molecular geometry. In particular, such energy dependent studies help to elucidate the mechanism(s) responsible for populating the symmetry forbidden vibrational levels [i.e., v+ =( 010), (001), (030), and (110)]. This is the first study in which vibrationally resolved data have been acquired as a function of energy for all of the vibrational modes of a polyatomic system. Theoretical Schwinger variational calculations are used to interpret the experimental data, and they indicate that a 4sigmag-->ksigmau shape resonance is responsible for most of the excursions observed for the vibrational branching ratios. Generally, the energy dependent trends are reproduced well by theory, but a notable exception is the symmetric stretch vibrational branching ratio. The calculated results display a strong peak in the vibrational branching ratio while the experimental data show a pronounced minimum. This suggests an interference mechanism that is not accounted for in the single-channel adiabatic-nuclei calculations. Electronic branching ratios were also measured and compared to the vibrational branching ratios to assess the relative contributions of interchannel (i.e., Herzberg-Teller) versus intrachannel (i.e., photoelectron-mediated) coupling. PMID- 15267896 TI - Variational transition state theory calculations for the rate constants of the hydrogen scrambling and the dissociation of BH5 using the multiconfiguration molecular mechanics algorithm. AB - The BH5 molecule contains a weak two-electron-three-center bond and it requires extremely high level of theories to calculate the energy and structure correctly. The potential energy of the hydrogen scrambling in BH5 has been generated by the multiconfiguration molecular mechanics algorithm with 15 high-level Shepard interpolation points, which would be practically impossible to obtain otherwise. The high-level interpolation points were obtained from the multicoefficient correlated quantum mechanical methods. The more high-level points are used, the better the shape of the potential energy surface. The rate constants are calculated using the variational transition state theory including multidimensional tunneling approximation. The potential energy curve for the BH5 dissociation has also been calculated, and the variational transition state was located to obtain the dissociation rate constants. Tunneling is very important in the scrambling, and there is large variational effect on the dissociation. The rate constants for the scrambling and the dissociation are 2.1 x 10(9) and 2.3 x 10(12) s(-1) at 300 K, respectively, which suggests that the dissociation is three orders of magnitude faster than the scrambling. PMID- 15267897 TI - The ground-state tunneling splitting of various carboxylic acid dimers. AB - Carboxylic acid dimers in gas phase reveal ground-state tunneling splittings due to a double proton transfer between the two subunits. In this study we apply a recently developed accurate semiclassical method to determine the ground-state tunneling splittings of eight different carboxylic acid derivative dimers (formic acid, benzoic acid, carbamic acid, fluoro formic acid, carbonic acid, glyoxylic acid, acrylic acid, and N,N-dimethyl carbamic acid) and their fully deuterated analogs. The calculated splittings range from 5.3e-4 to 0.13 cm(-1) (for the deuterated species from 2.8e-7 to 3.3e-4 cm(-1)), thus indicating a strong substituent dependence of the splitting, which varies by more than two orders of magnitude. One reason for differences in the splittings could be addressed to different barriers heights, which vary from 6.3 to 8.8 kcal/mol, due to different mesomeric stabilization of the various transition states. The calculated splittings were compared to available experimental data and good agreement was found. A correlation could be found between the tunneling splitting and the energy barrier of the double proton transfer, as the splitting increases with increased strength of the hydrogen bonds. From this correlation an empirical formula was derived, which allows the prediction of the ground-state tunneling splitting of carboxylic acid dimers at a very low cost and the tunneling splittings for parahalogen substituted benzoic acid dimers is predicted. PMID- 15267898 TI - Potential energy curves of diatomic molecular ions from high-resolution photoelectron spectroscopy. I. The first six electronic states of Ar2+. AB - High-resolution photoelectron spectroscopic data have been used to determine the potential energy curves of the first six electronic states of Ar2+. The potential energy functions properly include the effects of the long-range interactions and of the spin-orbit interaction and are of spectroscopic accuracy (1-2 cm(-1)) over a wide range of internuclear distances. The total number of adjustable parameters could be reduced to only 12 by truncating the long-range interaction series after the R(-6) term and assuming an R-independent spin-orbit coupling constant. This assumption was verified to be valid to an accuracy of +/-2 cm(-1) over the range of internuclear distances between 3.0 and 4.6 A. The interaction potential proposed by Siska [P. E. Siska, J. Chem. Phys. 85, 7497 (1986)] was generalized to a form that is expected to be sufficiently flexible to describe chemical bonding in other diatomic molecular ions. The potential energy curves are more accurate than the best available ab initio curves by two orders of magnitude and provide quantitative information on dissociation energies and equilibrium internuclear distances. The local maximum between the two potential wells of the I(1/2g) state was determined to lie 62 cm(-1) below the Ar(1S0)+Ar(+)(2P(3/2)) dissociation limit, and the II(1/2g) state is found to be significantly more bound (De=177 cm(-1)) than previously assumed. PMID- 15267899 TI - High-level ab initio computations of structures and interaction energies of naphthalene dimers: origin of attraction and its directionality. AB - The intermolecular interaction energies of naphthalene dimers have been calculated by using an aromatic intermolecular interaction model (a model chemistry for the evaluation of intermolecular interactions between aromatic molecules). The CCSD(T) (coupled cluster calculations with single and double substitutions with noniterative triple excitations) interaction energy at the basis set limit has been estimated from the second-order Moller-Plesset perturbation interaction energy near saturation and the CCSD(T) correction term obtained using a medium-size basis set. The estimated interaction energies of the set of geometries explored in this work show that two structures emerge as being the lowest energy, and may effectively be considered as isoenergetic on the basis of the errors inherent in out extrapolation procedure. These structures are the slipped-parallel (Ci) structure (-5.73 kcal/mol) and the cross (D2d) structure ( 5.28 kcal/mol). The T-shaped (C2v) and sandwich (D2h) dimers are substantially less stable (-4.34 and -3.78 kcal/mol, respectively). The dispersion interaction is found to be the major source of attraction in the naphthalene dimer. The electrostatic interaction is substantially smaller than the dispersion interaction. The large dispersion interaction is the cause of the large binding energies of the cross and slipped-parallel dimers. PMID- 15267900 TI - Photodissociation dynamics of the Kr-HBr cluster: the effect of the rare gas atom substitution. AB - The ultraviolet photolysis dynamics of Kr-HBr(v=0) is investigated by means of wave packet calculations, focusing on the fragmentation pathway Kr-HBr+ variant Planck's over 2pi omega-->H+Kr-Br. Photolysis is simulated by starting from two different cluster initial states, namely the ground van der Waals (vdW) and an excited vdW bending state, associated with the Kr-H-Br and Kr-Br-H isomers, respectively. The results show that, for the two initial states of the cluster, the Kr-Br product yield is lower than that of Ar-Br radicals found in previous studies on Ar-HBr photolysis. Despite this decrease, the Kr-Br yield is found to be still rather high, in particular for the initial excited vdW state of Kr HBr(v=0). In addition, the Kr-Br product state distributions exhibit a remarkably higher excitation (mainly rotational) than the corresponding Ar-Br distributions. The lower yield and higher excitation of Kr-Br as compared to Ar-Br, are attributed to a larger share of the energy available for the radical going to internal excitation in the case of the Kr-Br product. The different partition of the energy available for Kr-Br also causes significant deviations in the photolysis behavior of Kr-HBr when compared to that of Ar-HBr, in the case of the initial excited vdW state of both clusters. A common feature of the photodissociation of Kr-HBr and Ar-HBr is the manifestation of quantum interference effects in the Kr-Br and Ar-Br rotational state distributions, in the form of pronounced structures of supernumerary rotational rainbows. PMID- 15267901 TI - Many-body effects in molecular photoionization in intense laser fields; time dependent Hartree-Fock simulations. AB - The time evolution of the reduced single electron density matrix for eight electrons in a one-dimensional finite box potential driven by an intense laser field is calculated by numerically integrating the time-dependent Hartree-Fock equations. We study the effects of the Coulomb interaction, field intensity, and frequency on the time profile of the ionization process. Our computed saturation ionization intensity (Isat) is in good agreement with experimental results for decatetraene [Ivanov et al. J. Chem. Phys. 117, 1575 (2002)]. PMID- 15267902 TI - Structures of mixed gold-silver cluster cations (Ag(m)Au(n)+, m+n<6): ion mobility measurements and density-functional calculations. AB - The collision cross sections of Ag(m)Au(n)+ (m+n)<6 cluster ions were determined. For bimetallic clusters, we observe a significant intracluster charge transfer leaving most of the ions positive charge on the silver atoms. The mixed trimeric ions Ag2Au+ and AgAu2+ are triangular like the pure gold and silver trimers. Most of the tetrameric clusters are rhombus shaped, with the exception of Ag3Au+, which has a Y structure with the gold atom in the center. Among the pentamers we find distorted X structures for all systems. For Ag2Au3+ we find an additional isomer which is a trigonal bipyramid. These findings are in line with predictions based on density-functional theory calculations, i.e., all these structures either represent the global minima or are within less than 0.1 eV of the predicted global minimum. PMID- 15267903 TI - Dipole-bound anions of highly polar molecules: ethylene carbonate and vinylene carbonate. AB - Results of experimental and theoretical studies of dipole-bound negative ions of the highly polar molecules ethylene carbonate (EC, C3H4O3, mu=5.35 D) and vinylene carbonate (VC, C3H2O3, mu=4.55 D) are presented. These negative ions are prepared in Rydberg electron transfer (RET) reactions in which rubidium (Rb) atoms, excited to ns or nd Rydberg states, collide with EC or VC molecules to produce EC- or VC- ions. In both cases ions are produced only when the Rb atoms are excited to states described by a relatively narrow range of effective principal quantum numbers, n*; the greatest yields of EC- and VC- are obtained for n*(max)=9.0+/-0.5 and 11.6+/-0.5, respectively. Charge transfer from low lying Rydberg states of Rb is characteristic of a large excess electron binding energy (Eb) of the neutral parent; employing the previously derived empirical relationship Eb=23/n*(max)(2.8) eV, the electron binding energies are estimated to be 49+/-8 meV for EC and 24+/-3 meV for VC. Electron photodetachment studies of EC- show that the excess electron is bound by 49+/-5 meV, in excellent agreement with the RET results, lending credibility to the empirical relationship between Eb and n*(max). Vertical electron affinities for EC and VC are computed employing aug-cc-pVDZ atom-centered basis sets supplemented with a (5s5p) set of diffuse Gaussian primitives to support the dipole-bound electron; at the CCSD(T) level of theory the computed electron affinities are 40.9 and 20.1 meV for EC and VC, respectively. PMID- 15267904 TI - Spectroscopy of highly excited vibrational states of HCN in its ground electronic state. AB - An experimental technique based on a scheme of vibrationally mediated photodissociation has been developed and applied to the spectroscopic study of highly excited vibrational states in HCN, with energies between 29,000 and 30,000 cm(-1). The technique consists of four sequential steps: in the first one, a high power laser is used to vibrationally excite the sample to an intermediate state, typically (0,0,4), the nu3 mode being approximately equivalent to the C-H stretching vibration. Then a second laser is used to search for transitions between this intermediate state and highly vibrationally excited states. When one of these transitions is found, HCN molecules are transferred to a highly excited vibrational state. Third, a ultraviolet laser photodissociates the highly excited molecules to produce H and CN radicals in its A 2Pi electronic state. Finally, a fourth laser (probe) detects the presence of the CN(A) photofragments by means of an A-->B-->X laser induced fluorescence scheme. The spectra obtained with this technique, consisting of several rotationally resolved vibrational bands, have been analyzed. The positions and rotational parameters of the states observed are presented and compared with the results of a state-of-the-art variational calculation. PMID- 15267905 TI - Electron attachment to chlorouracil: a comparison between 6-ClU and 5-ClU. AB - Low energy electron impact to the isomers 6-chlorouracil (6-ClU) and 5 chlorouracil (5-ClU) yields a variety of negative ion fragments with surprisingly high cross sections. These ions are dominantly formed via sharply structured resonance features at energies below the threshold for electronic excitation and result from dissociative electron attachment (DEA). The most dominant DEA channel is formation of (M-HCl)-, i.e., ejection of a neutral HCl molecule with the negative charge remaining on the ring. The reaction cross section is 9 x 10(-18) m2 and 5 x 10(-18) m2 for 6-Cl and 5-ClU, respectively, and thus about two orders of magnitude higher than the geometrical cross section of the molecule. Further reactions also operative via low energy resonances (<2.5 eV) are Cl- abstraction, dehydrogenation [formation of (M-H)-, M=ClU], and DEA processes associated with a ring opening. Most of the ion yield curves exhibit remarkably sharp structures which have not been observed before in DEA to a polyatomic system. Although some possibilities on their origin are discussed, their interpretation remains a challenge for theory and further experiments. While electron attachment to both 6 ClU and 5-ClU generates fragments of the same stoichiometric composition, their ion yields and also their relative intensities show some very pronounced differences which can be explained by the different structure but also the different energetic situation in the two isomers. PMID- 15267906 TI - Intermolecular potential and second virial coefficient of the water-hydrogen complex. AB - We construct a rigid-body (five-dimensional) potential-energy surface for the water-hydrogen complex using scaled perturbation theory (SPT). An analytic fit of this surface is obtained, and, using this, two minima are found. The global minimum has C2v symmetry, with the hydrogen molecule acting as a proton donor to the oxygen atom on water. A local minimum with Cs symmetry has the hydrogen molecule acting as a proton acceptor to one of the hydrogen atoms on water, where the OH bond and H2 are in a T-shaped configuration. The SPT global minimum is bound by 1097 microEh (Eh approximately 4.359744 x 10(-18) J). Our best estimate of the binding energy, from a complete basis set extrapolation of coupled-cluster calculations, is 1076.1 microEh. The fitted surface is used to calculate the second cross virial coefficient over a wide temperature range (100-3000 K). Three complementary methods are used to quantify quantum statistical mechanical effects that become significant at low temperatures. We compare our results with experimental data, which are available over a smaller temperature range (230-700 K). Generally good agreement is found, but the experimental data are subject to larger uncertainties. PMID- 15267907 TI - Characterization of hydrated Na+(phenol) and K+(phenol) complexes using infrared spectroscopy. AB - Hydrated alkali metal ion-phenol complexes were studied to model these species in aqueous solution for M=Na and K. IR predissociation spectroscopy in the O-H stretch region was used to analyze the structures of M+(Phenol)(H2O)n cluster ions, for n = 1-4. The onset of hydrogen bonding was observed to occur at n=4. Ab initio calculations were used to qualitatively explore the types of hydrogen bonded structures of the M+(Phenol)(H2O)4 isomers. By combining the ab initio calculations and IR spectra, several different structures were identified for each metal ion. In contrast to benzene, detailed in a previous study of Na+(Benzene)n(H2O)m [J. Chem. Phys. 110, 8429 (1999)], phenol is able to bind directly to Na+ even in the presence of four waters. This is likely the result of the sigma-type interaction between the phenol oxygen and the ion. With K+, the dominant isomers are those in which the phenol O-H group is involved in a hydrogen bond with the water molecules, while with Na+, the dominant isomers are those in which the phenol O-H group is free and the water molecules are hydrogen bonded to each other. Spectra and ab initio calculations for the M+(Phenol)Ar cluster ions for M=Na and K are reported to characterize the free phenol O-H stretch in the M+(Phenol) complex. While pi-type configurations were observed for binary M+(Phenol) complexes, sigma-type configurations appear to dominate the hydrated cluster ions. PMID- 15267908 TI - Crossed beams and theoretical studies of the O(3P)+CH4-->H+OCH3 reaction excitation function. AB - The excitation function for the reaction, O(3P)+CH4-->H+OCH3, has been measured in a crossed molecular beams experiment and determined with direct dynamics calculations that use the quasiclassical trajectory method in conjunction with a recently developed semiempirical Hamiltonian. Good agreement is found between experiment and theory, enabling us to address two fundamental issues for the O(3P)+CH4 reaction that arise for all O(3P)+saturated hydrocarbon reactions: (1) the importance of triplet excited states that correlate adiabatically to ground state reactants and products and (2) the importance of intersystem crossing processes involving the lowest singlet surface [corresponding to reaction with O(1D)]. Our results indicate that the first excited triplet surface contributes substantially to the cross section when the collision energy exceeds the reaction barrier (approximately 2 eV) by more than 0.5 eV. Although triplet-singlet crossings may occur at all energies, we have found that their effect on the excitation function is negligible for the collision energies studied-up to 1.5 eV above threshold. PMID- 15267909 TI - Effects of pulse strength, width, and sample spinning speed on the spectral spin diffusion of multiquantum coherences of spin-3/2 quadrupolar nuclei. AB - The effects of radio-frequency pulse strength, width, and sample spinning speed on the spin-diffusion spectrum of half-integer quadrupolar spins in solids have been studied by theoretical, numerical, and experimental investigations. It is revealed that the line shape of the cross peaks changes nonmonotonically with respect to the change of pulse strength, pulse width, or sample spinning speed. It is also found that the sample spinning speed has much more pronounced influence on the spin diffusion spectral line shape. In many cases of practical importance, the effect of sample spinning must be included in spectral analysis, in contrast to the practice of previous studies. Moreover, this effect can be exploited to further improve the precision in the determination of relative orientation of the electric-field gradient tensors of the exchange partners. PMID- 15267910 TI - Molecular electrostatic potentials and electron densities in nitroazacubanes. AB - Successive introduction of nitrogen atoms in the cubyl corners instead of C-NO2 groups of octanitrocubane (CNO2)8, the most powerful explosives known to date, leads to a class of energy-rich compounds known as nitroazacubanes. In present work the ab initio Hartree-Fock and hybrid density functional calculations have been carried out on the possible conformers of (CNO2)(8-alpha)Nalpha (with alpha=0-8), nitroazacubanes. The charge distributions in these systems have been derived using the topography of the molecular electrostatic potential and electron density. Molecular electrostatic potential investigations reveal that of different nitroazacubane conformers, the electron-rich regions around nitro oxygens of the lowest energy conformer having face opposite nitrogen atoms within a cube are more delocalized. These conformers are predicted to have the largest difference of the energies of the highest occupied molecular orbital and lowest unoccupied molecular orbital relative to the other conformers. The dipole moments of nitroazacubanes are dependent on the nitrogen sites within a cube, caused by the resultant of C-N bond moments and nearly insensitive to position of the NO2 groups. The lowest frequency vibration (522 cm(-1)) suggests octa-azacubane having robust structure in the nitroazacubane series. Substitution of nitrogen atom instead of C-NO2 group leads to increase in electron density at the bond critical point of the X-N (X=C or N) bonds in a cube. The heats of formation of different nitroazacubanes were calculated by using the isodesmic reaction approach. The present calculation has shown that for the di- though hexanitroazacubanes the most destabilized conformer possess largest dipole moment and the heat of formation as well. A linear correlation of the electron density at the bond critical point of X-N bonds and the heat of formation has been obtained. PMID- 15267911 TI - Reaction of Cu+ with dimethoxyethane: competition between association and multiple dissociation channels. AB - The reaction of Cu+ with dimethoxyethane (DXE) is studied using kinetic-energy dependent guided ion beam mass spectrometry. The bimolecular reaction forms an associative Cu(+)(DXE) complex that is long-lived and dissociates into several competitive channels: C4H9O2(+)+CuH, Cu(+)(C3H6O)+CH3OH, back to reactants, and other minor channels. The kinetic-energy dependences of the cross sections for the three largest product channels are interpreted with several different models (including rigorous phase space theory) to yield 0 K bond energies after accounting for the effects of multiple ion-molecule collisions, internal energy of the reactant ions, Doppler broadening, and dissociation lifetimes. These values are compared with bond energies obtained from collision-induced dissociation (CID) studies of the Cu(+)(DXE) complex and found to be self consistent. Although all models provide reasonable thermochemistry, phase space theory reproduces the details of the cross sections most accurately. We also examine the dynamics of this reaction using time-of-flight methods and a retarding potential analysis. This provides additional insight into the unimolecular decay of the long-lived Cu(+)(DXE) association complex. Comparison of results from this study with those from the complementary CID study, thus forming the same energized Cu(+)(DXE) complex in two distinct ways, allows an assessment of the models used to interpret CID thresholds. PMID- 15267912 TI - Superexcited state reconstruction of HCl using photoelectron and photoion imaging. AB - The velocity-map imaging technique was used to record photoelectron and photofragment ion images of HCl following two-photon excitation of the E Sigma(+)(0+), V 1Sigma(+)(0+) (nu=9,10,11) states and subsequent ionization. The images allowed us to determine the branching ratios between autoionization and dissociation channels for the different intermediate states. These branching ratios can be explained on the basis of intermediate state electron configurations, since the configuration largely prohibits direct ionization in a one-electron process, and competition between autoionization and dissociation into H* (n=2)+Cl and H+Cl*(4s,4p,3d) is observed. From a fit to the vibrationally resolved photoelectron spectrum of HCl+ it is apparent that a single superexcited state acts as a gateway to autoionization and dissociation into H+Cl*(4s). Potential reconstruction of the superexcited state to autoionization was undertaken and from a comparison of different autoionization models it appears most likely that the gateway state is a purely repulsive and low-n Rydberg state with a (4Pi) ion core. PMID- 15267913 TI - Vibrational branching ratios in photoionization of CO and N2. AB - We report results of experimental and theoretical studies of the vibrational branching ratios for CO 4sigma(-1) photoionization from 20 to 185 eV. Comparison with results for the 2sigma(u)(-1) channel of the isoelectronic N2 molecule shows the branching ratios for these two systems to be qualitatively different due to the underlying scattering dynamics: CO has a shape resonance at low energy but lacks a Cooper minimum at higher energies whereas the situation is reversed for N2. PMID- 15267914 TI - An overlap expansion method for improving ab initio model potentials: anisotropic intermolecular potentials of N2, CO, and C2H2 with He*(2(3)S). AB - An overlap expansion method is proposed for improving ab initio model potentials. Correction terms are expanded in terms of overlap integrals between orbitals of the interacting system. The method is used to improve ab initio model potentials for N2+He*(2(3)S), CO+He*(2(3)S), and C2H2+He*(2(3)S). Physical meanings of the optimization are elucidated in terms of target orbitals. Correction terms are found to be dominated by the components of HOMO, LUMO, next-HOMO, and next-LUMO on the target molecule. The present overlap expansion method using a limited number of correction terms related to frontier orbitals provides an efficient and intuitive approach for construction of highly anisotropic intermolecular interaction potentials. PMID- 15267915 TI - Bond and mode selectivity in the reaction of atomic chlorine with vibrationally excited CH2D2. AB - The title reaction is investigated by co-expanding a mixture of Cl2 and CH2D2 into a vacuum chamber and initiating the reaction by photolyzing Cl2 with linearly polarized 355 nm light. Excitation of the first C-H overtone of CH2D2 leads to a preference for hydrogen abstraction over deuterium abstraction by at least a factor of 20, whereas excitation of the first C-D overtone of CH2D2 reverses this preference by at least a factor of 10. Reactions with CH2D2 prepared in a local mode containing two quanta in one C-H oscillator /2000>- or in a local mode containing one quantum each in two C-H oscillators /1100> lead to products with significantly different rotational, vibrational, and angular distributions, although the vibrational energy for each mode is nearly identical. The Cl+CH2D2/2000>- reaction yields methyl radical products primarily in their ground state, whereas the Cl+CH2D2/1100> reaction yields methyl radical products that are C-H stretch excited. The HCl(v=1) rotational distribution from the Cl+CH2D2/2000>- reaction is significantly hotter than the HCl(v=1) rotational distribution from the Cl+CH2D2/1100> reaction, and the HCl(v=1) differential cross-section (DCS) of the Cl+CH2D2/2000>- reaction is more broadly side scattered than the HCl(v=1) DCS of the Cl+CH2D2/1100> reaction. The results can be explained by a simple spectator model and by noting that the /2000>- mode leads to a wider cone of acceptance for the reaction than the /1100> mode. These measurements represent the first example of mode selectivity observed in a differential cross section, and they demonstrate that vibrational excitation can be used to direct the reaction pathway of the Cl+CH2D2 reaction. PMID- 15267916 TI - Simulation of the reactive scattering of F+D2 on a model family of potential energy surfaces with various topographies: the correlation approach. AB - The connection between the salient features of the potential energy surface (PES) and the dynamical characteristics of the elementary collision process is studied using a correlation approach based on quasiclassical trajectory simulations. The method is demonstrated for the reaction F + D2 --> D + DF for which the scattering characteristics were calculated on a model family of PES's based on a London-Eyring-Polanyi-Sato-type five-parameter equation. The correlations between the reactive cross section and the vibrational and rotational quantum numbers and the scattering angle of the DF product, and the various parameters of the collinear and noncollinear PES's, such as the location and height of the minimal barrier and the Sato coefficients, are reported. Although usually correlations between two variables suffice, in some cases coefficients of correlation among three variables are required. The role of nonlinear parameter dependencies in computing the correlation coefficients is also considered. The correlation approach makes it possible to examine a large set of potential surfaces without intermediate human control and obtain quantitative information. PMID- 15267917 TI - A different approach for calculating Franck-Condon factors including anharmonicity. AB - An efficient new procedure for calculating Franck-Condon factors, based on the direct solution of an appropriate set of simultaneous equations, is presented. Both Duschinsky rotations and anharmonicity are included, the latter by means of second-order perturbation theory. The critical truncation of basis set is accomplished by a build-up procedure that simultaneously removes negligible vibrational states. A successful test is carried out on ClO2 for which there are experimental data and other theoretical calculations. PMID- 15267918 TI - The binding energies of the D2d and S4 water octamer isomers: high-level electronic structure and empirical potential results. AB - The MP2 complete basis set (CBS) limit for the binding energy of the two low lying water octamer isomers of D2d and S4 symmetry is estimated at -72.7+/-0.4 kcal/mol using the family of augmented correlation-consistent orbital basis sets of double through quintuple zeta quality. The largest MP2 calculation with the augmented quintuple zeta (aug-cc-pV5Z) basis set produced binding energies of 73.70 (D2d) and -73.67 kcal/mol (S4). The effects of higher correlation, computed at the CCSD(T) level of theory, are estimated at <0.1 kcal/mol. The newly established MP2/CBS limit for the water octamer is reproduced quite accurately by the newly developed all atom polarizable, flexible interaction potential (TTM2 F). The TTM2-F binding energies of -73.21 (D2d) and -73.24 kcal/mol (S4) for the two isomers are just 0.5 kcal/mol (or 0.7%) larger than the MP2/CBS limit. PMID- 15267919 TI - Molecular dynamics simulation of liquid N2O4<-->2NO2 by orientation-sensitive pairwise potential. III. Reaction dynamics. AB - The dissociation and association dynamics of N2O4 [see text] 2NO2 in liquid state are studied by classical molecular dynamics simulations of reactive liquid NO2. An OSPP+LJ potential between NO2 molecules, which is a sum of an orientation sensitive pairwise potential (OSPP) between N-N atoms proposed in Paper I [J. Chem. Phys. 115, 10852 (2001)] and Lennard-Jones potentials between N-O and O-O atoms, has been used in the simulation. The reaction dynamics is studied as a function of well depth De and anisotropy factors of the OSPP potential: Atheta (0< or =Atheta< or =1) for the rocking angle and Atau (0< or =Atau< or =0.5) for the torsional angle of relative NO2-NO2 orientation. The lifetime tauD of initially prepared NO2 dimers is found to increase as De increases, Atheta increases, and Atau decreases. Dissociation and association dynamics are studied in detail around the extreme limit of pure NO2-dimer liquid: De=0.12 x 10(-18) J, Atheta=0.5, and Atau=0.1, which has been found to reproduce both the observed liquid phase equilibrium properties and Raman band shapes of the dissociation mode very well. The dissociation dynamics from microscopic reaction trajectories is compared with the potential of the mean force (PMF) as a function of the N-N distance R. The PMF of reactive liquid NO2 shows a transition state barrier at R=2.3-2.5 A, and NO2-trimer structure is found to be formed at the barrier. Two types of dissociation of the NO2 dimer-the dissociation by collisional activation of the reactive mode to cross the dissociation limit and the NO2-mediated dissociation via bond transfer-are studied. The latter needs less free energy and is found to be much more probable. The dissociation trajectories and PMF in reactive liquid NO2 are compared with those of a reactive NO2 pair in inert solvent N2O4. PMID- 15267920 TI - Fractionation of peptide with disulfide bond for quantum mechanical calculation of interaction energy with molecules. AB - We present a computational study of a recently developed molecular fractionation with conjugated caps (MFCC) method for application to peptide/protein that has disulfide bonds. Specifically, we employ the MFCC approach to generate peptide fragments in which a disulfide bond is cut and a pair of conjugated caps are inserted. The method is tested on two peptides interacting with a water molecule. The first is a dipeptide consisting of two cysteines (Cys-Cys) connected by a disulfide bond and the second is a seven amino acid peptide consisting of Gly-Cys Gly-Gly-Gly-Cys-Gly with a disulfide cross link. One-dimensional peptide-water potential curves are computed using the MFCC method at various ab initio levels for a number of interaction geometries. The calculated interaction energies are found to be in excellent agreement with the results obtained from the corresponding full system ab initio calculations for both peptide/water systems. The current study provides further numerical support for the accuracy of the MFCC method in full quantum mechanical calculation of protein/peptide that contains disulfide bonds. PMID- 15267921 TI - Measurement and dynamics of the spatial distribution of an electron localized at a metal-dielectric interface. AB - The ability of time- and angle-resolved two-photon photoemission to estimate the size distribution of electron localization in the plane of a metal-adsorbate interface is discussed. It is shown that the width of angular distribution of the photoelectric current is inversely proportional to the electron localization size within the most common approximations in the description of image potential states. The localization of the n=1 image potential state for two monolayers of butyronitrile on Ag(111) is used as an example. For the delocalized n=1 state, the shape of the signal amplitude as a function of momentum parallel to the surface changes rapidly with time, indicating efficient intraband relaxation on a 100 fs time scale. For the localized state, little change was observed. The latter is related to the constant size distribution of electron localization, which is estimated to be a Gaussian with a 15+/-4 A full width at half maximum in the plane of the interface. A simple model was used to study the effect of a weak localization potential on the overall width of the angular distribution of the photoemitted electrons, which exhibited little sensitivity to the details of the potential. This substantiates the validity of the localization size estimate. PMID- 15267922 TI - Classification of secondary relaxation in glass-formers based on dynamic properties. AB - Dynamic properties, derived from dielectric relaxation spectra of glass-formers at variable temperature and pressure, are used to characterize and classify any resolved or unresolved secondary relaxation based on their different behaviors. The dynamic properties of the secondary relaxation used include: (1) the pressure and temperature dependences; (2) the separation between its relaxation time taubeta and the primary relaxation time taualpha at any chosen taualpha; (3) whether taubeta is approximately equal to the independent (primitive) relaxation time tau0 of the coupling model; (4) whether both taubeta and tau0 have the same pressure and temperature dependences; (5) whether it is responsible for the "excess wing" of the primary relaxation observed in some glass-formers; (6) how the excess wing changes on aging, blending with another miscible glass-former, or increasing the molecular weight of the glass-former; (7) the change of temperature dependence of its dielectric strength Deltaepsilonbeta and taubeta across the glass transition temperature Tg; (8) the changes of Deltaepsilonbeta and taubeta with aging below Tg; (9) whether it arises in a glass-former composed of totally rigid molecules without any internal degree of freedom; (10) whether only a part of the molecule is involved; and (11) whether it tends to merge with the alpha-relaxation at temperatures above Tg. After the secondary relaxations in many glass-formers have been characterized and classified, we identify the class of secondary relaxations that bears a strong connection or correlation to the primary relaxation in all the dynamic properties. Secondary relaxations found in rigid molecular glass-formers belong to this class. The secondary relaxations in this class play the important role as a precursor or local step of the primary relaxation, and we propose that only they should be called the Johari-Goldstein beta-relaxation. PMID- 15267923 TI - The effects of solute-solvent electrostatic interactions on solvatochromic shifts in supercritical CO2. AB - Solvent clustering around attractive solutes is an important feature of supercritical solvation. We examine here the effects of the local density enhancement on solvatochromic shifts in electronic absorption and emission spectra in supercritical CO2. We use molecular dynamics (MD) simulation to study the spectral line shifts for model diatomic solutes that become more polar upon electronic excitation. The electronic transition is modeled as either a change from a quadrupolar to a dipolar solute charge distribution or as an increase in the magnitude of the solute dipole. Our main focus is on the density dependence of the line shifts at 320 K, which corresponds to about 1.05 times the solvent critical temperature, Tc, but results for higher temperatures are also obtained in order to determine the behavior of the line shifts in the absence of local density enhancement. We find that the extent of local density enhancement at 1.05Tc is strongly correlated with solute-solvent electrostatic attraction and that the density dependence of the emission line shifts resembles the behavior of the effective local densities, rho(eff), obtained from the first-shell coordination numbers. The differences that are seen are shown to be due to solute solvent orientational correlations which provide an additional source of enhancement for electrostatic solvation energies and spectral line shifts. PMID- 15267924 TI - Rotation-libration and rotor-rotor coupling in 4-methylpyridine. AB - The low temperature rotational dynamics of methyl groups in 4-methylpyridine is analyzed in terms of a model potential including rotation-libration and rotor rotor coupling. The parameters of the model potential are adjusted by comparison of calculated with published and newly recorded inelastic neutron scattering spectra. Initial evaluations of the potential parameters of the model are obtained from molecular mechanics calculations. Experimental spectra are calculated from these potentials by numerical solution of Schrodinger's equation for clusters of coupled rotors embedded in a bigger ensemble of rotors treated in the mean field approximation. Adjustment of the potential parameters leads to excellent agreement with the experimental spectra of protonated 4-methylpyridine, measured at well-defined spin temperatures. At higher levels of deuteration, agreement with experiment is qualitative, only. The observed deviations are attributed to the increasing frustration of the system of coupled methyl groups and mutual localization, effects leading to a phase transition around 5.5 K in isotopic mixtures, as shown in diffraction experiments. PMID- 15267925 TI - On combining molecular dynamics and stochastic dynamics simulations to compute reaction rates in liquids. AB - An approach that combines molecular dynamics and stochastic dynamics calculations for obtaining reaction rates in liquids is investigated by studying the cis- >trans isomerization of HONO in liquid krypton. The isomerization rates are computed for several liquid densities by employing full-dimensional molecular dynamics simulations. The rates are also computed by employing the stochastic dynamics method for a wide range of collision frequencies. Comparisons of the two sets of the computed rates show that for a wide range of liquid densities there is a simple linear relation between the liquid density rho and the collision frequency alpha, that is, alpha=crho. This suggests that once the constant c is determined from a molecular-dynamics calculation at a single density, the reaction rates can be obtained from stochastic dynamics calculations for the entire range of liquid densities where alpha=crho holds. The applicability of the combined molecular dynamics and stochastic dynamics approach provides a practical means for obtaining rate constants at considerable savings of computer time compared to that required by using full-dimensional molecular-dynamics simulations alone. PMID- 15267926 TI - Implicit solvation based on generalized Born theory in different dielectric environments. AB - In this paper we are investigating the effect of the dielectric environment on atomic Born radii used in generalized Born (GB) methods. Motivated by the Kirkwood expression for the reaction field of a single off-center charge in a spherical cavity, we are proposing extended formalisms for the calculation of Born radii as a function of external and internal dielectric constants. We demonstrate that reaction field energies calculated from environmentally dependent Born radii lead to much improved agreement with Poisson-Boltzmann solutions for low dielectric external environments, such as biological membranes or organic solvent, compared to previous methods where the calculation of Born radii does not depend on the environment. We also examine how this new approach can be applied for the calculation of transfer free energies from vacuum to a given external dielectric for a system with an internal dielectric larger than one. This has not been possible with standard GB theory but is relevant when scoring minimized or average structures with implicit solvent. PMID- 15267927 TI - Optical pump-terahertz probe spectroscopy of dyes in solutions: probing the dynamics of liquid solvent or solid precipitate? AB - The optical pump-terahertz probe spectroscopy was used together with ab initio calculations and molecular dynamics simulations to investigate ultrafast dynamics following electronic excitation of Coumarin 153 and TBNC (2,11,20,29-tetra-tert butyl-2,3-naphtalocyanine) dyes in polar solvents. By scanning the terahertz waveform for different pump-probe delays this experimental technique allows us to obtain two dimensional spectra directly reflecting the temporal response of the system. A distinct signal was obtained for TBNC in chloroform, 2-propanol, and n butanol, while no signal was recorded for Coumarin 153 in either of these solvents. We explain the nonequilibrium signal detected in TBNC solutions by the presence of a solid, polycrystalline phase of the dye resulting from irradiating the solution by intense optical pulses. PMID- 15267928 TI - Vibrational relaxation in methyl hydrocarbons at high temperatures: propane, isobutene, isobutane, neopentane, and toluene. AB - Vibrational relaxation has been seen in shock waves in propane, isobutene, isobutane, neopentane, and toluene dilute in krypton with the laser-schlieren technique. These experiments cover about 600-2200 K and post-shock pressures from 5 to 29 Torr. The process cannot be resolved in any for T<600 K, or in any for large molecule fraction. All the ultrasonic measurements of relaxation in these at room temperature show characteristic times in the 1-5 ns atm range, corresponding to fewer than five collisions, whereas the relaxation times in the shock waves range from 20 to 200 ns atm, with a clearly defined negative or "inverted" temperature dependence. It would seem the observed slowdown of relaxation with increasing T is simply a consequence of the much increased energy transfer required at high temperature in such large polyatomics when this is combined with a collision efficiency, here interpreted as down, already so large it cannot much increase. The simple method for the extraction of a down from relaxation data offered here by consideration of the energy relaxation equation for Evib=0 appears to be original and should prove quite useful in connecting thermal relaxation data to values obtained from spectroscopy and master-equation analyses. Here it is found that the derived down extrapolate well to room temperature ultrasonic measurements, showing a slight increase with temperature. PMID- 15267929 TI - Classical density functional theory of orientational order at interfaces: application to water. AB - A classical density functional formalism has been developed to predict the position-orientation number density of structured fluids. It is applied to the liquid-vapor interface of pure water, where it consists of a classical term, a gradient correction, and an anisotropic term that yields order through density gradients. The model is calibrated to predict that water molecules have their dipole moments almost parallel to a planar interface, while the molecular plane is parallel to it on the liquid side and perpendicular to it on the vapor side. For a planar interface, the surface tension obtained is twice its experimental value, while the surface potential is in qualitative agreement with that calculated by others. The model is also used to predict the orientation of water molecules near the surface of droplets, as well as the dependence of equilibrium vapor pressure around them on their size. PMID- 15267930 TI - Monte Carlo molecular simulation of the hydration of Na-montmorillonite at reservoir conditions. AB - The hydration of Na-saturated Wyoming-type montmorillonite is investigated by Monte Carlo simulations at constant stress in the NP(zz)T ensemble and at constant chemical potential in the microVT ensemble, at the sedimentary basin temperature of 353 K and pressure of 625 bar, equivalent to 2-4 km depth. The simulations use procedures established in Chavez-Paez et al. [J. Chem. Phys. 114, 1405 (2001)]. At these conditions, simulations predict a single stable form of 1,2-water layer Na-montmorillonite, containing 164.38 mg/g or 53.37 molecules/layer of adsorbed water and having a spacing of 12.72 A. The corresponding density is 0.32 g/ml. Sodium ions are coordinated with six molecules of water separated 2.30-2.33 A. Water molecules are closer to the central interlayer plane and the spacing is larger than that at 300 K and 1 bar. The interlayer configuration consists of two symmetrical layers of oriented water molecules 1.038 A from the central plane, with the hydrogen atoms in two outermost layers, 3.826 A apart, and the sodium ions on the central plane located between the water layers. The interlayer configuration can be considered to be a stable two-layer intermediate between the one- and two-layer hydrates. Our simulations do not predict formation of other hydrates of Na-montmorillonite at 353 K and 615 bar. PMID- 15267931 TI - Electrokinetic microfluidic phenomena by a lattice Boltzmann model using a modified Poisson-Boltzmann equation with an excluded volume effect. AB - A lattice Boltzmann model (LBM) for electrokinetic microfluidics recently proposed by us [J. Colloid Interface Sci. 263, 144 (2003); Langmuir 19, 3041 (2003)] is employed by consideration of a modified Poisson-Boltzmann equation including an excluded volume effect. In our study, pressure is considered as the only external force for liquid flow. As commonly used, the Poisson-Boltzmann equation assumes only point charge, the predicted microfluidic phenomena of KCl and LiCl electrolyte solutions are theoretically the same; these phenomena, however, have been found to be experimentally different. Our LBM in conjunction with the modified Poisson-Boltzmann equation are capable of explaining this discrepancy and the results are in good agreement with recent experimental data for KCl and LiCl electrolyte solutions in pressure-driven microchannel flow, suggesting that our proposed LBM can be employed to predict the more complex microfluidic systems that might be problematic using conventional methods and electrokinetic models. PMID- 15267932 TI - Ground state and phase transitions in a system of arg-cysteamines self-assembled on a Au(111) crystal surface. AB - The translational and orientation order of arg-cysteamine molecules chemiabsorbed on the Au(111) crystal surface is considered. Couplings between carbon, nitrogen, and hydrogen atoms of the n-alkanethiols are approximated by the Lennard-Jones potential. Moreover, hydrogen bonds between oxygen and nitrogen and dipole-dipole interactions of the dipole moments of different atomic groups are taken into account. It is found that molecules are arranged in a 2 x 2 lattice and have the total symmetry C6 x Z2. The critical temperature of the phase transition to the tilted state Tc1, which breaks the symmetry C6, is estimated to be extremely high. The spontaneous breakdown of the remaining symmetry Z2 leads to the twisted state of the molecules and has the critical temperature Tc2=340 K. PMID- 15267933 TI - Magnetic interactions in calcium and sodium ladder vanadates. AB - Magnetic interactions in ladder vanadates are determined with quantum chemical computational schemes using the embedded cluster model approach to represent the material. The available experimental data for calcium vanadate is accurately reproduced and the nature of the interladder interaction is established to be ferromagnetic. An analysis of the main contributions to the magnetic couplings is presented and the role of the covalently bonded apex oxygen is elucidated. In the sodium vanadate, the ground state configuration of the rungs is V-3d1-O-2p5-V 3d1. We show that with this configuration good intrachain coupling constants are obtained for the high-temperature phase. The interchain coupling in NaV2O5 is predicted to be approximately 34 K, ferromagnetic in nature. PMID- 15267934 TI - Negative ion formation in electron-stimulated desorption of CF2Cl2 coadsorbed with polar NH3 on Ru(0001). AB - Photon-induced dissociation of CF2Cl2 (freon-12) in the stratosphere contributes substantially to atmospheric ozone depletion. We report recent results on dissociation and negative ion formation in electron-stimulated desorption (ESD) of CF2Cl2 on Ru(0001), when CF2Cl2 is coadsorbed with a polar molecule (NH3), for electron energies ranging from 50 to 300 eV. Two different time-of-flight methods are used in this investigation: (a) an ESD ion angular distribution detector with wide collection angle and (b) a quadrupole mass spectrometer with narrow collection angle and high mass resolution. Many negative ESD fragments are seen (F-, Cl-, FCl-, CF-, F2-, and Cl2-), whose intensities depend on the surface preparation. Using both detectors we observe a giant enhancement of Cl- and F- yields for ESD of CF2Cl2 coadsorbed with approximately 1 ML of NH3; this enhancement (>10(3) for Cl-) is specific to certain ions, and is attributed to an increased probability of dissociative electron attachment due to "trapped" low energy secondary electrons, i.e., precursor states of the solvated electron in NH3. In further studies, the influence of polar NH3 spacer layers (1-10 ML) on ESD of top-layer CF2Cl2 is determined, and compared with thick films of condensed CF2Cl2. The magnitudes and energy dependences of the Cl- yields are different in these cases, due to several contributing factors. PMID- 15267935 TI - Surface reaction mechanisms of hydrazine on Si(100)-2 x 1 surface: NH3 desorption pathways. AB - Multireference as well as single-reference wave functions were adopted to study the surface reaction mechanisms of hydrazine. The initial surface mechanisms resemble those of ammonia and its methyl derivatives. MRMP2 values indicate that the lifetime of initial molecularly adsorbed species should be longer than previously suggested. High energy path as well as low energy path of subsequent surface reactions were found. The theoretical initial surface product of low energy path is consistent with the experimentally suggested structure. Both paths eventually lead to very stable surface products, which are also consistent with the experimentally suggested structures. The reaction channels of the experimentally observed NH3 desorptions were also revealed. It was shown that the high reactivity of hydrazine as compared to ammonia and its methyl derivatives is due to the high nucleophilic ability of the additional nitrogen atom of hydrazine. PMID- 15267936 TI - Adsorption, diffusion, and dissociation of molecular oxygen at defected TiO2(110): a density functional theory study. AB - The properties of reduced rutile TiO2(110) surfaces, as well as the adsorption, diffusion, and dissociation of molecular oxygen are investigated by means of density functional theory. The O2 molecule is found to bind strongly to bridging oxygen vacancies, attaining a molecular state with an expanded O-O bond of 1.44 A. The molecular oxygen also binds (with somewhat shortened bond lengths) to the fivefold coordinated Ti atoms in the troughs between the bridging oxygen rows, but only when vacancies are present somewhere in the surface. In all cases, the magnetic moment of O2 is lost upon adsorption. The expanded bond lengths reveal together with inspection of electron density and electronic density of state plots that charging of the adsorbed molecular oxygen is of key importance in forming the adsorption bond. The processes of O2 diffusion from a vacancy to a trough and O2 dissociation at a vacancy are both hindered by relative large barriers. However, we find that the presence of neighboring vacancies can strongly affect the ability of O2 to dissociate. The implications of this in connection with diffusion of the bridging oxygen vacancies are discussed. PMID- 15267937 TI - Solution stability and variability in a simple model of globular proteins. AB - It is well known among molecular biologists that proteins with a common ancestor and that perform the same function in similar organisms, can have rather different amino-acid sequences. Mutations have altered the amino-acid sequences without affecting the function. A simple model of a protein in which the interactions are encoded by sequences of bits is introduced, and used to study how mutations can change these bits, and hence the interactions, while maintaining the stability of the protein solution. This stability is a simple minimal requirement on our model proteins which mimics part of the requirement on a real protein to be functional. The properties of our model protein, such as its second virial coefficient, are found to vary significantly from one model protein to another. It is suggested that this may also be the case for real proteins in vivo. PMID- 15267938 TI - Electron-phonon interactions in charged cubic fluorocarbon cluster, (CF)8. AB - Electron-phonon interactions in the charged cubic fluorocarbon, (CF)8 are studied, and compared with those in charged (CH)8 and (CD)8. The A1g mode of 1470 cm(-1) much more strongly couples to the a1g lowest unoccupied molecular orbitals (LUMO) than the A1g mode of 554 cm(-1) in (CF)8. The T2g mode of 1030 cm(-1), the Eg mode of 980 cm(-1), and the A1g mode of 1470 cm(-1) strongly couple to the t2u highest occupied molecular orbitals (HOMO) in (CF)8. The total electron-phonon coupling constants for the monoanion (l(-1)) and monocation (l(+1)) of (CF)8 are estimated to be 0.932 and 0.585 eV, respectively. The logarithmically averaged phonon frequencies for the monoanion (omega(ln,-1)) and monocation (omega(ln,+1)) of (CF)8 are estimated to be 1365 and 998 cm(-1), respectively. The l(-1) and omega(ln,-1) values increase much more significantly by H-F substitution than by H-D substitution in cubane. The larger displacements of carbon atoms in the high frequency vibronic active mode in (CF)8 than those in (CD)8 due to larger atomic mass of fluorine than that of deuterium, and the unchanged electron distributions in the LUMO somewhat localized on carbon atoms as a consequence of H-F and H-D substitution in cubane, are the main reason why the l(-1) and omega(ln,-1) values increase much more significantly by H-F substitution than by H-D substitution. The l(+1) and omega(ln,+1) values less significantly change than the l(-1) and omega(ln,-1) values by H-F substitution as well as by H-D substitution in cubane. This is because the t2u HOMO in (CF)8 and the t2g HOMO in (CH)8 are somewhat localized on fluorine atoms, and thus, the high frequency vibronic active modes in which the displacements of carbon atoms are large cannot necessarily very strongly couple to the HOMO somewhat localized on fluorine atoms in (CF)8. PMID- 15267939 TI - The structure of fluids confined in crystalline slitlike nanoscopic pores: bilayers. AB - Grand canonical and canonical ensemble Monte Carlo simulation methods are used to study the structure and phase behavior of Lennard-Jones fluids confined between the parallel (100) planes of the face centered cubic crystal. Ultra thin slit pores of the width allowing for the formation of only two adsorbate layers are considered. It is demonstrated that the structure of adsorbed phases is very sensitive to the wall-wall separation and to the strength of the fluid-wall potential. It is also shown that the structure of low temperature (solid) phases strongly depends on the fluid density. In particular, when the surface field is sufficiently strong, then the high density phases may exhibit a domain wall structure, quite the same as found in monolayer films adsorbed at a single substrate wall. On the other hand, the weakening of the surface potential leads to the regime in which only the hexagonally ordered bilayer structure is stable. The phase diagrams for a series of systems are estimated. It is shown that, depending on the pore width and the temperature, the condensation leads to the formation of the commensurate or incommensurate phases. The incommensurate phases may have the domain-wall or the hexagonal structure depending on the pore width and the strength of the fluid-wall potential. PMID- 15267940 TI - A washboard with moment of inertia model of gas-surface scattering. AB - A washboard with moment of inertia (WBMI) model for gas atom scattering from a flexible surface is proposed and applied. This model is a direct extension of the washboard model [J. Chem. Phys. 92, 680 (1990)] proposed for gas atom scattering from relatively rigid, corrugated surfaces. In addition, a moment of inertia is incorporated in the original washboard model to describe the flexibility of softer, more highly corrugated surfaces such as polymer or liquid surfaces. The moment of inertia of the effective surface object introduces a dependence of the efficiency of energy transfer on the position and direction of impact, a feature that has been shown to be critical by molecular dynamics simulations. The WBMI model is solved numerically by Monte Carlo integration, which makes the implementation of multiple impacts between a colliding atom and the surface very efficient. The model is applied to Ne and Ar atoms scattering from an alkylthiolate self-assembled monolayer surface and reproduces the major results obtained by classical trajectory simulation of the same system, i.e., a bimodal translation energy distribution P(E(f)) with the low-energy component well-fit with a Boltzmann distribution, but with a temperature that may (Ar) or may not (Ne) be the same as the surface temperature. This indicates that the WBMI model, with well-motivated physical assumptions and simplified interaction, reveals many of the major aspects of the gas-surface collision dynamics, though it does not take into account the real-time dynamics explicitly. PMID- 15267941 TI - Hydrolysis of a two-membered silica ring on the amorphous silica surface. AB - We have combined density functional theory (DFT) with classical interatomic potential functions to model hydrolysis of amorphous silica surfaces. The water silica interaction is described by DFT with incorporation of a long-range elastic field described by classical interatomic potentials. Both physisorption and chemisorption of water on a surface site, known as the two-membered silica ring, are studied in detail. The hybrid quantum-mechanical and classical mechanical method enables more realistic treatment of chemical processes on an extended surface than previous methods. We have studied cooperative events in the hydrolytic reactions and discovered a new reaction pathway that involves a double proton transfer process. In addition, the evaluation of the total energy in a hybrid quantum-mechanical and classical mechanical system is discussed. PMID- 15267942 TI - Structure of carbon onions and nanotubes formed by arc in liquids. AB - Since carbon nanotubes and onions were discovered, many methods have been proposed for their production. For applications the main requirements are low capital cost, high purity of the produced material, simplicity of technique, and its potential for scale up. Recently a cathodic arc between two graphitic electrodes immersed in liquids has been demonstrated to be a simple method to produce carbon nanoparticles such as nanotubes and onions. In this paper high resolution transmission electron microscopy is employed to examine the shape of the nanoparticles and the purity of the final material produced under various conditions. In this study we have used an arc discharge in two different liquids- liquid nitrogen and distilled water--and we have changed the grade of the carbon electrodes. The variety in structure, shape, and size of the produced particles is discussed in line with a model proposed to describe the physical process. PMID- 15267943 TI - Solvent-free simulations of fluid membrane bilayers. AB - A molecular level model for lipid bilayers is presented. Lipids are represented by rigid, asymmetric, soft spherocylinders in implicit solvent. A simple three parameter potential between pairs of lipids gives rise to a rich assortment of phases including (but not limited to) micelles, fluid bilayers, and gel-like bilayers. Monte Carlo simulations have been carried out to verify self-assembly, characterize the phases corresponding to different potential parametrizations, and to quantify the physical properties associated with those parameter sets corresponding to fluid bilayer behavior. The studied fluid bilayers have compressibility moduli in agreement with experimental systems, but display bending moduli at least three times larger than typical biological membranes without cholesterol. PMID- 15267944 TI - Time series analysis of collective motions in proteins. AB - The dynamics of alpha-amylase inhibitor tendamistat around its native state is investigated using time series analysis of the principal components of the C(alpha) atomic displacements obtained from molecular dynamics trajectories. Collective motion along a principal component is modeled as a homogeneous nonstationary process, which is the result of the damped oscillations in local minima superimposed on a random walk. The motion in local minima is described by a stationary autoregressive moving average model, consisting of the frequency, damping factor, moving average parameters and random shock terms. Frequencies for the first 50 principal components are found to be in the 3-25 cm(-1) range, which are well correlated with the principal component indices and also with atomistic normal mode analysis results. Damping factors, though their correlation is less pronounced, decrease as principal component indices increase, indicating that low frequency motions are less affected by friction. The existence of a positive moving average parameter indicates that the stochastic force term is likely to disturb the mode in opposite directions for two successive sampling times, showing the modes tendency to stay close to minimum. All these four parameters affect the mean square fluctuations of a principal mode within a single minimum. The inter-minima transitions are described by a random walk model, which is driven by a random shock term considerably smaller than that for the intra minimum motion. The principal modes are classified into three subspaces based on their dynamics: essential, semiconstrained, and constrained, at least in partial consistency with previous studies. The Gaussian-type distributions of the intermediate modes, called "semiconstrained" modes, are explained by asserting that this random walk behavior is not completely free but between energy barriers. PMID- 15267945 TI - Temperature modeling of laser-irradiated azo-polymer thin films. AB - Azobenzene polymer thin films exhibit reversible surface mass transport when irradiated with a light intensity and/or polarization gradient, although the exact mechanism remains unknown. In order to address the role of thermal effects in the surface relief grating formation process peculiar to azo polymers, a cellular automaton simulation was developed to model heat flow in thin films undergoing laser irradiation. Typical irradiation intensities of 50 mW/cm2 resulted in film temperature rises on the order of 5 K, confirmed experimentally. The temperature gradient between the light maxima and minima was found, however, to stabilize at only 10(-4) K within 2 micros. These results indicate that thermal effects play a negligible role during inscription, for films of any thickness. Experiments monitoring surface relief grating formation on substrates of different thermal conductivity confirm that inscription is insensitive to film temperature. Further simulations suggest that high-intensity pulsed irradiation leads to destructive temperatures and sample ablation, not to reversible optical mass transport. PMID- 15267946 TI - Phase behavior of binary mixtures of sterically stabilized colloids with large size asymmetry. AB - Experimental phase diagrams of three types of mixtures of sterically stabilized colloids are presented. The size ratios are kept similar, 0.15< or =xi< or =0.17, while the thickness and the chemical nature of the steric layers are varied. For all particles their effective volume fractions are calculated from their hydrodynamic radii. When their phase behavior is expressed in this way, the experimental liquidus curves all lie slightly above recent computer simulation predictions for the fluid-solid binodal of additive hard sphere mixtures. No dramatic shift of the experimental liquidus curves due to nonadditive particle interactions is observed. The dense phase is in all cases solid, with crystallites of the large spheres visible in some samples. PMID- 15267947 TI - Phase behavior in thin films of cylinder-forming ABA block copolymers: experiments. AB - We experimentally establish a phase diagram of thin films of concentrated solutions of a cylinder forming polystyrene-block-polybutadiene-block-polystyrene triblock copolymer in chloroform. During annealing the film forms islands and holes with energetically favored values of film thickness. The thin film structure depends on the local thickness of the film and the polymer concentration. Typically, at a thickness close to a favored film thickness parallel orientation of cylinders is observed, while perpendicular orientation is formed at an intermediate film thickness. At high polymer concentration the cylindrical microdomains reconstruct to a perforated lamella structure. Deviations from the bulk structure, such as the perforated lamella and a wetting layer are stabilized in films thinner than approximately 1.5 domain spacings. PMID- 15267948 TI - Phase behavior in thin films of cylinder-forming ABA block copolymers: mesoscale modeling. AB - The phase behavior of cylinder-forming ABA block copolymers in thin films is modeled in detail using dynamic density functional theory and compared with recent experiments on polystyrene-block-polybutadiene-block-polystyrene triblock copolymers. Deviations from the bulk structure, such as wetting layer, perforated lamella, and lamella, are identified as surface reconstructions. Their stability regions are determined by an interplay between surface fields and confinement effects. Our results give evidence for a general mechanism governing the phase behavior in thin films of modulated phases. PMID- 15267949 TI - Role of dissimilar interfaces in thin films of cylinder-forming block copolymers. AB - We study the effect of dissimilar interfaces on the phase behavior of cylinder forming block copolymers in thin films by means of dynamic density-functional theory. In this article, we show that dissimilarity of the interfaces induces hybrid structures. These structures appear when the surface fields at the two interfaces stabilize different surface structures and/or reconstructions. We propose a general classification of hybrid structures and give an unifying description of phase behavior of cylinder forming block copolymer films. Our results are consistent with experimental observations. PMID- 15267950 TI - An estimate of random close packing density in monodisperse hard spheres. PMID- 15267951 TI - Ferromagnetic dimer interactions in Cu2Cl4(CH3CN)2. PMID- 15267954 TI - Optimal control of ultrafast selection. AB - Optimal laser control for ultrafast selection of closely lying excited states whose energy separation is smaller than the laser bandwidth is reported on the two-photon transition of atomic cesium; Cs(6S-->7D(J), J=5/2 and 3/2). Selective excitation was carried out by pulse shaping of ultrashort laser pulses which were adaptively modulated in a closed-loop learning system handling eight parameters representing the electric field. Two-color fluorescence from the respective excited states was monitored to measure the selectivity. The fitness used in the learning algorithm was evaluated from the ratio of the fluorescence yields. After fifty generations, a pair of nearly transform-limited pulses were obtained as an optimal pulse shape, proving the effectiveness of the "Ramsey fringes" mechanism. The contrast of the selection ratio was improved by approximately 30% from the simple "Ramsey fringes" experiment. PMID- 15267955 TI - Chaperoned alchemical free energy simulations: a general method for QM, MM, and QM/MM potentials. AB - A general method for alchemical free energy simulations using QM, MM, and QM/MM potential is developed by introducing "chaperones" to restrain the structures, particularly near the end points. A calculation of the free energy difference between two triazole tautomers in aqueous solution is used to illustrate the method. PMID- 15267956 TI - Unified treatment of multicenter integrals of integer and noninteger u Yukawa type screened Coulomb type potentials and their derivatives over Slater orbitals. AB - Multicenter integrals with integer and noninteger values of indices u of Yukawa type screened Coulomb-type potentials (SCTPs) f(u)(eta,r)=r(u-1)e(-etar) and their derivatives over Slater-type orbitals (STOs) are evaluated using series expansion formulas obtained from the expansions in terms of complete orthonormal sets of Psi(alpha)-exponential-type orbitals (Psi(alpha)-ETOs, alpha=1, 0, -1, 2,...). The final results obtained are valid for the arbitrary values of parameters of SCTPs and STOs. PMID- 15267957 TI - An efficient method for constructing nonorthogonal localized molecular orbitals. AB - A new method for constructing nonorthogonal localized molecular orbitals (NOLMOs) is presented. The set of highly localized NOLMOs is obtained by minimization of the spread functional starting from an initial set of canonical orthogonal molecular orbitals. To enhance the stability and efficiency, the centroids of the NOLMOs are constrained to be those of the corresponding orthogonal localized molecular orbitals (OLMOs), which are obtained with the Boys criterion in advance. In particular, these centroid constraints make the optimization for each NOLMO independent of the others, which is an attractive feature for application to large systems. The minimization with the constraints incorporated through the multiplier-penalty function method is stable and efficient in convergence. While exhibiting the classical bonding pattern in chemistry and sharing a spatial distribution similar to that of the corresponding OLMOs, the obtained NOLMOs are more compact than the corresponding OLMOs with about 10%-28% reduction in the value of the spread functional and devoid of the troublesome "orthogonalization tails." PMID- 15267958 TI - A diradical mechanism for the addition of F2 to ethene: a density functional theory study. AB - The long disputed mechanism for the F(2) addition to ethene is elucidated by density functional theory calculations. With hybrid functionals and a large basis set, DFT provides an excellent description for the dissociation of ground state F(2), and a reasonable account for the F(2)...C(2)H(4) van der Waals complex, which makes it possible for the modeling of reactions between elemental fluorine and organic molecules. The attack of F(2) on ethene first produces a diradical intermediate, which then dissociates into two radicals CH(2)F-CH(2) and F. The first step is exothermic with a low barrier around 1.8 kcal/mol, and the exothermic energy is more than enough to overcome the barrier in the second step for the homolysis of the dangling F-F bond in the diradical, although the presence of solvents and matrices environment could stabilize the diradical. Our calculations provide a coherent framework to understand this reaction not only in the gas and solution phases, but also in the matrices environment where mode specific enhancement is observed for the addition process induced by infrared radiation. PMID- 15267959 TI - Comparison of density functionals for energy and structural differences between the high- [5T2g: (t2g)4(eg)2] and low- [1A1g: (t2g)6(eg)0] spin states of the hexaquoferrous cation [Fe(H2O)6]2+. AB - A comparison of density functionals is made for the calculation of energy and geometry differences for the high- [(5)T(2g): (t(2g))(4)(e(g))(2)] and low- [(1)A(1g): (t(2g))(6)(e(g))(0)] spin states of the hexaquoferrous cation [Fe(H(2)O)(6)](2+). Since very little experimental results are available (except for crystal structures involving the cation in its high-spin state), the primary comparison is with our own complete active-space self-consistent field (CASSCF), second-order perturbation theory-corrected complete active-space self-consistent field (CASPT2), and spectroscopy-oriented configuration interaction (SORCI) calculations. We find that generalized gradient approximations (GGAs) and the B3LYP hybrid functional provide geometries in good agreement with experiment and with our CASSCF calculations provided sufficiently extended basis sets are used (i.e., polarization functions on the iron and polarization and diffuse functions on the water molecules). In contrast, CASPT2 calculations of the low-spin-high spin energy difference DeltaE(LH)=E(LS)-E(HS) appear to be significantly overestimated due to basis set limitations in the sense that the energy difference of the atomic asymptotes ((5)D-->(1)I excitation of Fe(2+)) are overestimated by about 3000 cm(-1). An empirical shift of the molecular DeltaE(LH) based upon atomic calculations provides a best estimate of 12 000-13 000 cm(-1). Our unshifted SORCI result is 13 300 cm(-1), consistent with previous comparisons between SORCI and experimental excitation energies which suggest that no such empirical shift is needed in conjunction with this method. In contrast, after estimation of incomplete basis set effects, GGAs with one exception underestimate this value by 3000-4000 cm(-1) while the B3LYP functional underestimates it by only about 1000 cm(-1). The exception is the GGA functional RPBE which appears to perform as well as or better than the B3LYP functional for the properties studied here. In order to obtain a best estimate of the molecular DeltaE(LH) within the context of density functional theory (DFT) calculations we have also performed atomic excitation energy calculations using the multiplet sum method. These atomic DFT calculations suggest that no empirical correction is needed for the DFT calculations. PMID- 15267960 TI - Vibrational predissociation in the HCl dimer. AB - We present results of a combined theoretical and experimental study on the vibrational predissociation of the HCl dimer. On the theoretical side, photodissociation linewidths and product-state distributions for monomer stretch excited states with total angular momentum J=0 were computed, using the Fermi golden rule approximation. The resonances investigated include excitation of the hydrogen bond donor and acceptor stretches, as well as combinations of one of these modes with the intermolecular stretch and geared bend modes, for both even and odd permutation symmetry. Line strengths for the transitions from the J=1, K=0 ground state to excited states with J=0 were computed using quasibound states. On the experimental side, the photofragment angular distribution method was employed to obtain complete final-state distributions for the monomer stretch excited states. Three different transitions were probed, all starting from the lower tunneling component of the ground state: the (R)Q(0)(1) transition for excitation of the acceptor stretch and the (Q)R(0)(0) transition and unresolved (R)Q(0) branch for the donor stretch excitation. We find that, in contrast to the HF dimer, the excited-state alignment of the HCl dimer, resulting from excitation using a polarized laser beam, is completely lost on the time scale of the dissociation. The agreement between theory and experiment for the product-state distributions and line strengths is reasonable. The computed lifetimes are 1-2 orders of magnitude too small, which is attributed to a deficiency in the potential energy surface. PMID- 15267961 TI - Use of the ultraviolet absorption spectrum of CF2 to determine the spatially resolved absolute CF2 density, rotational temperature, and vibrational distribution in a plasma etching reactor. AB - Broadband ultraviolet absorption spectroscopy has been used to determine CF(2) densities in a plasma etch reactor used for industrial wafer processing, using the CF(2) A (1)B(1)<--X (1)A(1) absorption spectrum. Attempts to fit the experimental spectra using previously published Franck-Condon factors gave poor results, and values for the higher vibrational levels of the A state [(0,v(2),0), with v(2) (')>6] from the ground state were missing; hence new values were calculated. These were computed for transitions between low-lying vibrational levels of CF(2) X (1)A(1) to vibrational levels of CF(2) A (1)B(1) (v(1) ('),v(2) ('),0) up to high values of the vibrational quantum numbers using high level ab initio calculations combined with an anharmonic Franck Condon factor method. The Franck Condon factors were used to determine the absorption cross sections of CF(2) at selected wavelengths, which in turn were used to calculate number densities from the experimental spectra. Number densities of CF(2) have been determined in different regions of the plasma, including the center of the plasma and outside the plasma volume, and CF(2) rotational temperatures and vibrational energy distributions were estimated. For absorption spectra obtained outside the confined plasma volume, the CF(2) density was determined as (0.39+/-0.08)x10(13) molecule cm(-3) and the vibrational and rotational temperatures were determined as 303 and 350 K, respectively. In the center of the plasma reactor, the CF(2) density is estimated as (3.0+/-0.6)x10(13) molecules cm(-3) with T(rot) approximately 500 K. The fitted vibrational distribution in the CF(2) ground state corresponds to two Boltzmann distributions with T(vib) approximately 300 and T(vib) approximately 1000 K, indicating that CF(2) molecules are initially produced highly vibrationally excited, but are partially relaxed in the plasma by collision. PMID- 15267962 TI - Product angular distributions in dissociative recombination. AB - The dependence of the dissociative recombination cross section upon the angle between the incoming electron beam and the ion internuclear axis is determined for diatomic molecules. Product angular distributions are derived for the component partial waves of the Coulomb wave function. In agreement with earlier results for dissociative attachment, it is shown that in the slow rotation approximation, if electron capture is dominated by a single partial wave, the product angular distribution is given by the square of the absolute value of the partial wave spherical harmonic describing the incoming electron. PMID- 15267963 TI - Structure of hydrated clusters of tetrahydroisoquinoline [THIQ-(H2O)(n=1,3)] investigated by jet spectroscopy. AB - The hydrated clusters of tetrahydroisoquinoline have been investigated by laser induced fluorescence (LIF), UV-UV hole burning, and IR-UV double-resonance spectroscopy in a seeded supersonic jet. Clusters of different sizes and isomeric structures have different 0-0 transitions (origins) in the LIF spectrum. UV-UV hole burning spectroscopy has been used to identify different cluster species and their vibrational modes. The structures of the clusters have been predicted by comparing the observed OH and NH frequencies in the IR-UV double-resonance spectra with the results calculated at different levels of sophistication. It is found that the water molecules form linear and six- and eight-membered cyclic H bonded structures at the nitrogen center of 1:1, 1:2, and 1:3 clusters, respectively. PMID- 15267964 TI - Dissociation chemistry of hydrogen halides in water. AB - To understand the mechanism of aqueous acid dissociation, which plays a fundamental role in aqueous chemistry, the ionic dissociation of HX acids (X=F, Cl, Br, and I) in water clusters up to hexamer is examined using density functional theory and Moller-Plesset second-order perturbation methods (MP2). Further accurate analysis based on the coupled clusters theory with singles and doubles excitations agrees with the MP2 results. The equilibrium structures, binding energies, electronic properties, stretching frequencies, and rotational constants of HX(H(2)O)(n) and X(-)(H(3)O)(+)(H(2)O)(n-1) are calculated. The dissociated structures of HF and HCl can be formed for n>/=4, while those of HBr and HI can be formed for n>/=3. Among these, the dissociated structures of HX (X=Cl, Br, and I) are more stable than the undissociated ones for n>/=4, while such cases for HF would require much more than six water molecules, in agreement with previous reports. The IR spectra of stable clusters including anharmonic frequencies are predicted to facilitate IR experimental studies. Undissociated systems have X-H stretching modes which are highly redshifted by hydration. Dissociated hydrogen halides show three characteristic OH stretching modes of hydronium moiety, which are redshifted from the OH stretching modes of water molecules. PMID- 15267965 TI - The diazocarbene (CNN) molecule: characterization of the X 3Sigma- and A 3Pi electronic states. AB - The ground (X (3)Sigma(-)) and first excited triplet (A (3)Pi) electronic states of diazocarbene (CNN) have been investigated systematically starting from the self-consistent-field theory and proceeding to the coupled cluster with single, double, and full triple excitations (CCSDT) method with a wide range of basis sets. While the linear X (3)Sigma(-) ground state of CNN has a real degenerate bending vibrational frequency, the A (3)Pi state of CNN is subject to the Renner Teller effect and presents two distinct real vibrational frequencies along the bending coordinate. The bending vibrational frequencies of the A (3)Pi state were evaluated via the equation-of-motion coupled cluster (EOM-CC) techniques. The significant sensitivity to level of theory in predicting the ground-state geometry, harmonic vibrational frequencies, and associated infrared intensities has been attributed to the fact that the reference wave function is strongly perturbed by the excitations of 1pi-->3pi followed by a spin flip. At the highest level of theory with the largest basis set, correlation-consistent polarized valence quadruple zeta (cc-pVQZ) CCSDT, the classical X-A splitting (T(e) value) was predicted to be 68.5 kcal/mol (2.97 eV, 24 000 cm(-1)) and the quantum mechanical splitting (T(0) value) to be 69.7 kcal/mol (3.02 eV, 24 400 cm(-1)), which are in excellent agreement with the experimental T(0) values, 67.5-68.2 kcal/mol (2.93-2.96 eV, 23 600-23 900 cm(-1)). With the EOM-CCSD method the Renner parameter (epsilon) and averaged bending vibrational frequency (omega(2)) for the A (3)Pi state were evaluated to be epsilon=-0.118 and omega(2)=615 cm( 1), respectively. They are in fair agreement with the experimental values of epsilon=-0.07 and nu(2)=525 cm(-1). PMID- 15267966 TI - Mass spectrometry study of the fragmentation of valence and core-shell (Cl 2p) excited CHCl3 and CDCl3 molecules. AB - The dissociative photoionization of the chloroform and chloroform-d molecules has been studied in the valence region and around the chlorine 2p edge. Time-of flight mass spectrometry in the coincidence mode-namely, photoelectron-photoion coincidence (PEPICO)-was employed. He I lamp and tunable synchrotron radiation were used as light sources. Total and partial ion yields have been recorded as a function of the photon energy. Singly, doubly, and triply ionized species have been observed below (195 eV), on (201 eV), and above (230 eV) the Cl 2p resonances. A definite degree of site-selective fragmentation was observed at the Cl 2p resonance as the relative contributions of several ionic species were seen to go through a maximum at 201 eV. At the same time all stable doubly charged ions were also observed at 198 eV (below the 2p resonances), resulting from direct ionization processes. Isotopic substitution is shown to provide a very efficient means of improving the mass resolution and assignment of unresolved peaks in spectra of CHCl(3), particularly for those fragments differing by a hydrogen atom. It is suggested that ultrafast fragmentation of the system following 2p excitation to a strongly antibonding state contributes to the large amount of Cl(+) observed in the PEPICO spectrum measured at 201 eV. Kinetic energy distributions were determined for the H(+), D(+), and Cl(+) fragments. PMID- 15267967 TI - Calculation of the fourth-rank molecular hypermagnetizability of some small molecules. AB - A computational scheme has been developed within the framework of Rayleigh Schrodinger perturbation theory to evaluate nonlinear interaction energy contributions for a molecule in the presence of an external spatially uniform, time-independent magnetic field. Terms connected with the fourth power of the perturbing field, representing the fourth-rank hypermagnetizabilities of five small molecules, have been evaluated at the coupled Hartree-Fock level of accuracy within the conventional common-origin approach. Gaugeless basis sets of increasing size and flexibility have been employed in a numerical test, adopting two different coordinate systems to estimate the degree of convergence of theoretical tensor components. PMID- 15267968 TI - Single-photon vacuum-ultraviolet laser-pulsed-field ionization-photoelectron studies of trans- and cis-1-bromopropenes. AB - The vacuum-ultraviolet (VUV) pulsed-field ionization-photoelectron (VUV-PFI-PE) spectra of trans-1-bromopropene (trans-CH(3)CH[Double Bond]CHBr) and cis-1 bromopropene (cis-CH(3)CH[Double Bond]CHBr) have been measured in the energy region of 74 720-76 840 cm(-1). The simulation of fine structures observed in the origin VUV-PFI-PE vibrational bands of these molecules has provided the ionization energies (IEs) of trans-1-bromopropene and cis-1-bromopropene to be 74 779.3+/-2.0 cm(-1) (9.2715+/-0.0002 eV) and 75 140.2+/-2.0 cm(-1) (9.3162+/ 0.0002 eV), respectively. The vibrational bands resolved in these VUV-PFI-PE spectra at energies 0-1700 cm(-1) above the IEs of trans-1-bromopropene and cis-1 bromopropene have been assigned based on theoretical vibrational frequencies and calculated Franck-Condon factors for the ionization transitions. PMID- 15267969 TI - Crossed beam studies of the reactions of atomic oxygen in the ground 3P and first electronically excited 1D states with hydrogen sulfide. AB - The reactions of both ground, (3)P, and electronically excited, (1)D, oxygen atoms with hydrogen sulfide, H(2)S, have been investigated by means of the crossed molecular beams method with mass spectrometric detection at different collision energies. Amongst the possible reaction channels those leading to HSO+H for the O((3)P) reaction and to HSO/HOS+H and SO+H(2) for the O((1)D) reaction have been identified and investigated. The dynamics of the channels leading to HSO/HOS+H are elucidated for the reactions of both states and the trend with increasing the collision energy analyzed. Noteworthily, the formation of SO+H(2) products appears to be an open channel for the O((1)D) reaction, at least for the highest collision energy investigated (11.8 kcal/mol). Finally, the recent experimental and theoretical estimates of the enthalpy of formation of the HSO radical have been critically analyzed to evaluate their conformity with the present experimental data. PMID- 15267970 TI - Converged quantum calculations of HO2 bound states and resonances for J=6 and 10. AB - Bound and resonance states of HO(2) are calculated quantum mechanically using both the Lanczos homogeneous filter diagonalization method and the real Chebyshev filter diagonalization method for nonzero total angular momentum J=6 and 10, using a parallel computing strategy. For bound states, agreement between the two methods is quite satisfactory; for resonances, while the energies are in good agreement, the widths are in general agreement. The quantum nonzero-J specific unimolecular dissociation rates for HO(2) are also calculated. PMID- 15267971 TI - Adsorption energies of molecular oxygen on Au clusters. AB - The adsorption properties of O(2) molecules on anionic, cationic, and neutral Au(n) clusters (n=1-6) are studied using the density functional theory (DFT) with the generalized gradient approximation (GGA), and with the hybrid functional. The results show that the GGA calculations with the PW91 functional systemically overestimate the adsorption energy by 0.2-0.4 eV than the DFT ones with the hybrid functional, resulting in the failure of GGA with the PW91 functional for predicting the adsorption behavior of molecular oxygen on Au clusters. Our DFT calculations with the hybrid functional give the same adsorption behavior of molecular oxygen on Au cluster anions and cations as the experimental measurements. For the neutral Au clusters, the hybrid DFT predicts that only Au(3) and Au(5) clusters can adsorb one O(2) molecule. PMID- 15267972 TI - Photoinduced electron transfer and geminate recombination for photoexcited acceptors in a pure donor solvent. AB - Photoinduced electron transfer and geminate recombination are studied for the systems rhodamine 3B (R3B(+)) and rhodamine 6G (R6G(+)), which are cations, in neat neutral N,N-dimethylaniline (DMA). Following photoexcitation of R3B(+) or R6G(+) (abbreviated as R(+)), an electron is transferred from DMA to give the neutral radical R and the cation DMA(+). Because the DMA hole acceptor is the neat solvent, the forward transfer rate is very large, approximately 5x10(12) s( 1). The forward transfer is followed by geminate recombination, which displays a long-lived component suggesting several percent of the radicals escape geminate recombination. Spectrally resolved pump-probe experiments are used in which the probe is a "white" light continuum, and the full time-dependent spectrum is recorded with a spectrometer/charge-coupled device. Observations of stimulated emission (excited state decay-forward electron transfer), the R neutral radical spectrum, and the DMA(+) radical cation spectrum as well as the ground-state bleach recovery (geminate recombination) make it possible to unambiguously follow the electron transfer kinetics. Theoretical modeling shows that the long-lived component can be explained without invoking hole hopping or spin-forbidden transitions. PMID- 15267973 TI - An ice phase of lowest thermal conductivity. AB - On pressurizing at temperatures near 130 K, hexagonal and cubic ices transform implosively at 0.8-1 GPa. The phase produced on transformation has the lowest thermal conductivity among the known crystalline ices and its value decreases on increase in temperature. An ice phase of similar thermal conductivity is produced also when high-density amorphous ice kept at 1 GPa transforms on slow heating when the temperature reaches approximately 155 K. These unusual formation conditions, the density and its distinguished thermal conductivity, all indicate that a distinct crystal phase of ice has been produced. PMID- 15267974 TI - Fluorescence quenching by reversible excimer formation: Kinetics and yield predictions for a classical potential association-dissociation model. AB - Fluorescence quenching by reversible excimer formation is studied on the assumption that excimer formation and dissociation can be modelled as entering and leaving the attractive region of an monomer excited-monomer interaction potential by diffusion. To get some general insight in the kinetic consequences of such a type of modelling, the simple case of an attractive square-well potential is investigated. It is shown that three different kinetic regimes have to be distinguished: Two "reversible" ones in case of slow excimer radiative decay, in which the quenching kinetics can be formulated by Markovian or non Markovian rate equations with both excimer formation and excimer dissociation terms, and an effectively "irreversible" regime if the excimer radiative decay is too rapid to allow the excimer equilibration. In the latter case a dissociation coefficient can no longer be defined and the quenching kinetics can only be predicted on the basis of generalized rate equations of a net-excimer-formation type. It is shown how the quenching constant formula must be generalized to be applicable in all kinetic situations. PMID- 15267975 TI - Selective excitation in spin systems with homogeneous broadening. AB - We have found that the application of a weak radio frequency (rf) pulse to a spin system with indirect spin-spin (J) couplings can produce a narrow inverted peak in spectral regions where there are many overlapping peaks. Examples of three compounds with (1)H-(1)H J couplings and one with (1)H-(13)C J couplings are given. The dependences of the signals on the frequency, duration, and amplitude of the weak rf irradiation have been studied. For an rf power of gammaB(1)/2pi=1.0 Hz, pulse widths longer than 0.25 s consistently produce inverted narrow peaks in spectral regions with unresolved peaks. An interpretation of the origin of this unusual result of selective excitation in spin systems with homogeneous broadening is given: the inverted signal observed can be considered as a cumulative effect of the weak rf irradiation acting on many uncorrelated transitions that are slightly off resonance on both sides of the irradiating frequency; the only role of the J couplings (or dipolar couplings in liquid crystals) is to produce a large number of closely spaced peaks that overlap with each other. Computer simulations of both coupled and noninteracting spin systems have been carried out, and the results support this interpretation. PMID- 15267976 TI - Low temperature extension of the generalized Zusman phase space equations for electron transfer. AB - In a previous paper [J. Chem. Phys. 119, 11864 (2003)], we derived a set of two coupled equations which describe electron transfer in the presence of dissipation at high temperature. Employing the low temperature extension of the Fokker-Planck operator, suggested by Haake and Reibold [Phys. Rev. A 32, 2462 (1985)] and Ankerhold [Europhys. Lett. 61, 301 (2003)], we show that one may extend the generalized Zusman equations in a similar manner to low temperature. Numerical simulation shows that addition of the temperature-dependent term which couples the coordinate and momentum causes an increase in the electron transfer rate as compared to the rate obtained from the previous high temperature equations. The increase in the rate comes from the increase in the equilibrium variances of the coordinate and momentum. The low temperature quantum theory allows for higher energy portions of phase space to contribute to the electron transfer rate where the rate is higher thus enhancing the overall rate. PMID- 15267977 TI - Glass structure and ion dynamics of lead-cadmium fluorgermanate glasses. AB - Glass structure and fluorine motion dynamics are investigated in lead-cadmium fluorgermanate glasses by means of differential scanning calorimetry, Raman scattering, x-ray absorption (EXAFS), electrical conductivity (EC), and (19)F nuclear magnetic resonance (NMR) techniques. Glasses with composition 60PbGeO(3) xPbF(2)-yCdF(2) (in mol %), with x+y=40 and x=10, 20, 30, 40, are studied. Addition of metal fluorides to the base PbGeO(3) glass leads to a decrease of the glass transition temperature (T(g)) and to an enhancement of the ionic conductivity properties. Raman and EXAFS data analysis suggest that metagermanate chains form the basic structural feature of these glasses. The NMR study leads to the conclusion that the F-F distances are similar to those found in pure crystalline phases. Experimental results suggest the existence of a heterogeneous glass structure at the molecular scale, which can be described by fluorine rich regions permeating the metagermanate chains. The temperature dependence of the NMR line shapes and relaxation times exhibits the qualitative and quantitative features associated with the high fluorine mobility in these systems. PMID- 15267978 TI - Relaxation of the electrical double layer after an electron transfer approached by Brownian dynamics simulation. AB - In this paper, the dynamical properties of the electrochemical double layer following an electron transfer are investigated by using Brownian dynamics simulations. This work is motivated by recent developments in ultrafast cyclic voltammetry which allow nanosecond time scales to be reached. A simple model of an electrochemical cell is developed by considering a 1:1 supporting electrolyte between two parallel walls carrying opposite surface charges, representing the electrodes; the solution also contains two neutral solutes representing the electroactive species. Equilibrium Brownian dynamics simulations of this system are performed. To mimic electron transfer processes at the electrode, the charge of the electroactive species are suddenly changed, and the subsequent relaxation of the surrounding ionic atmosphere are followed, using nonequilibrium Brownian dynamics. The electrostatic potential created in the center of the electroactive species by other ions is found to have an exponential decay which allows the evaluation of a characteristic relaxation time. The influence of the surface charge and of the electrolyte concentration on this time is discussed, for several conditions that mirror the ones of classical electrochemical experiments. The computed relaxation time of the double layer in aqueous solutions is found in the range 0.1 to 0.4 ns for electrolyte concentrations between 0.1 and 1 mol L( 1) and surface charges between 0.032 and 0.128 C m(-2). PMID- 15267979 TI - Conformational kinetics of disaccharides in aqueous solutions. AB - Between 100 kHz and 2 GHz ultrasonic attenuation spectra of disaccharides in water have been measured at 25 degrees C. Some additional measurements have been performed at different temperatures between 10 and 35 degrees C and as a function of saccharide concentration c (0.5 mol/l/=0.5. The predictions of the superposition approximation, where the solvation force response for the rough pores is deduced from the solvation force response of the slit-shaped pores, was in excellent agreement with simulation results for the structured pores and for lambda/sigma(ff)>/=7 in the case of smooth walled pores. Grand potential computations illustrate that interactions between the walls of the pore can alter the pore width corresponding to the thermodynamically stable state, with wall-wall interactions playing an important role at smaller pore widths and higher amplitudes of roughness. PMID- 15267986 TI - The characteristic curve and site-selective laser excitation of local relaxation in glass. AB - The so-called characteristic curve describing photosensitivity change is elaborated and shown to be a powerful tool for understanding and characterizing photosensitive growth both at a fundamental and practical level. It has been used successfully to diagnose when optimal hypersensitization has been achieved and the physical basis for this is explained. By way of example, previous results using 355 nm hypersensitization are re-examined. Evidence of single site selective glass relaxation through direct laser excitation offers a new approach to accessing and studying induced relaxations. PMID- 15267987 TI - Thickness-dependent ordering of water layers at the NaCl(100) surface. AB - We have determined the ordering properties of water adsorbed at room temperature on the rock salt (100) surface under four different conditions: ultrahigh vacuum, dry nitrogen atmosphere, 45% and 75% relative humidity. Details of the atomic structure are determined for both sides of the solid-liquid interface. The top most layer of NaCl shows a small relaxation that changes from an expansion to a contraction with increasing humidity. Under all measured conditions water monolayers with different ordering properties are present at the interface. Surprisingly, we find that the amount of ordering in the first layer is increasing with increasing thickness of the water film. At a solid-liquid interface, the ordering appears to be correlated with the solubility. PMID- 15267988 TI - A first-principles study on quasi-1D alkali metal chains within zeolite channels. AB - We report first-principles studies on systems formed by alkali metal (Na, K, or Rb) added to zeolite ITQ-4. Geometric and electronic structures of the quasi-1D chains of intercalated alkali metal atoms at experimental loading (4 metal atoms per 32 Si) are studied. Clear differences between different kinds of alkali metal are found, with a general trend of decreased ionization and less metallic character for the lighter alkali metals. Within the zeolite channels, it is possible to form insulated and metallic alkali metal chains by doping Na or Rb. Agreeing with experiments, only Rb here is found to be a good candidate to generate inorganic electride. We also predict that a large quantity of Na can be doped into the zeolite channel, while no more than 4 Rb per 16 Si can be doped. PMID- 15267989 TI - The hydrophobic effect: molecular dynamics simulations of water confined between extended hydrophobic and hydrophilic surfaces. AB - Structural and dynamic properties of water confined between two parallel, extended, either hydrophobic or hydrophilic crystalline surfaces of n-alkane C(36)H(74) or n-alcohol C(35)H(71)OH, are studied by molecular dynamics simulations. Electron density profiles, directly compared with corresponding experimental data from x-ray reflectivity measurements, reveal a uniform weak de wetting characteristic for the extended hydrophobic surface, while the hydrophilic surface is weakly wetted. These microscopic data are consistent with macroscopic contact angle measurements. Specific water orientation is present at both surfaces. The ordering is characteristically different between the surfaces and of longer range at the hydrophilic surface. Furthermore, the dynamic properties of water are different at the two surfaces and different from the bulk behavior. In particular, at the hydrophobic surface, time-correlation functions reveal that water molecules have characteristic diffusive behavior and orientational ordering due to the lack of hydrogen bonding interactions with the surface. These observations suggest that the altered dynamical properties of water in contact with extended hydrophobic surfaces together with a partial drying of the surfaces are more indicative of the hydrophobic effect than structural ordering, which we suggest to be independent of surface topology. PMID- 15267990 TI - Adsorption of 3-pyrroline on Si(100) from first principles. AB - The chemisorption of 3-pyrroline (C(4)H(7)N) on Si(100) is studied from first principles. Three different structures can be realized for which, depending on the temperature, the chemisorption process is facile (for two of them it is essentially barrierless); among these configurations the most favored one, from a thermodynamical point of view, is a dissociated structure obtained through an exothermic reaction characterized by the formation of a N-Si bond and a H-Si bond in which the H atom is detached from the molecule. Several other chemisorption structures are possible which, however, require overcoming a significant energy barrier and often breaking multiple bonds. A number of reaction paths going from one stable structure to another have been investigated. We have also generated, for the two basic adsorption structures, theoretical scanning tunneling microscopy images which could facilitate the interpretation of experimental measurements, and we propose a possible reaction mechanism for nitrogen incorporation. PMID- 15267991 TI - On the closure conjectures for the Gibbsian approximation model of a binary droplet. AB - Within the framework of Gibbsian thermodynamics, a binary droplet is regarded to consist of a uniform interior and dividing surface. The properties of the droplet interior are those of the bulk liquid solution, but the dividing surface is a fictitious phase whose chemical potentials cannot be rigorously determined. The state of the nucleus interior and free energy of nucleus formation can be found without knowing the surface chemical potentials, but the latter are still needed to determine the state of the whole nucleus (including the dividing surface) and develop the kinetics of nucleation. Thus it is necessary to recur to additional conjectures in order to build a complete, thermodynamic, and kinetic theory of nucleation within the framework of the Gibbsian approximation. Here we consider and analyze the problem of closing the Gibbsian approximation droplet model. We identify micro- and Gamma-closure conjectures concerning the surface chemical potentials and excess surface coverages, respectively, for the droplet surface of tension. With these two closure conjectures, the Gibbsian approximation model of a binary droplet becomes complete so that one can determine both the surface and internal characteristics of the whole nucleus and develop the kinetic theory, based on this model. Theoretical results are illustrated by numerical evaluations for binary nucleation in a water-methanol vapor mixture at T=298.15 K. Numerical results show a striking increase in the droplet surface tension with decreasing droplet size at constant overall droplet composition. A comparison of the Gibbsian approximation with density functional calculations for a model surfactant system indicate that the excess surface coverages from the Gibbsian approximation are accurate enough for large droplets and droplets that are not too concentrated with respect to the solute. PMID- 15267992 TI - Photoluminescence studies of oligothiophene self-assembled monolayers at low excitation energy. AB - Photoluminescence spectroscopy studies have been performed on self-assembled monolayers (SAMs) on Au(111) of thiophene oligomers with the number of thiophen rings N=3 and N=4. Photoluminescence spectra of SAMs reveal excitonic behavior with different band resolution and temperature dependence. These differences are attributed to different SAMs structure (degree of ordering). PMID- 15267993 TI - Application of density functional theory to capillary phenomena in cylindrical mesopores with radial and longitudinal density distributions. AB - In this paper, we applied a version of the nonlocal density functional theory (NLDFT) accounting radial and longitudinal density distributions to study the adsorption and desorption of argon in finite as well as infinite cylindrical nanopores at 87.3 K. Features that have not been observed before with one dimensional NLDFT are observed in the analysis of an inhomogeneous fluid along the axis of a finite cylindrical pore using the two-dimensional version of the NLDFT. The phase transition in pore is not strictly vapor-liquid transition as assumed and observed in the conventional version, but rather it exhibits a much elaborated feature with phase transition being complicated by the formation of solid phase. Depending on the pore size, there are more than one phase transition in the adsorption-desorption isotherm. The solid formation in finite pore has been found to be initiated by the presence of the meniscus. Details of the analysis of the extended version of NLDFT will be discussed in the paper. PMID- 15267994 TI - A new correlation effect in the Helmholtz and surface potentials of the electrical double layer. AB - The restricted primitive model of an electrical double layer around a spherical macroparticle is studied by using integral equation theories and Monte Carlo simulations. The resulting theoretical curves for the Helmholtz and surface potentials versus the macroparticle charge show an unexpected positive curvature when the ionic size of uni- and divalent electrolyte species is increased. This is a novel effect that is confirmed here by computer experiments. An explanation of this phenomenon is advanced in terms of the adsorption and layering of the electrolytic species and of the compactness of the diffuse double layer. It is claimed that the interplay between electrostatic and ionic size correlation effects, absent in the classical Poisson-Boltzmann view, is responsible for this singularity. PMID- 15267995 TI - Density-functional study of the cycloaddition of acrylonitrile on the Si(100) surface. AB - Using a density functional approach, we have explored the cycloaddition of acrylonitrile on the Si(100) surface. The buckling of the surface dimers characteristic for the (2x1) reconstructed surface is shown to favor structures with a dipolar moment such as the resonant form of acrylonitrile with cumulative double bonds. The bond of acrylonitrile via a single C atom is a possible intermediate leading to the nitrile structure of the adsorbed molecule. PMID- 15267996 TI - Kinetics of phase transformation on a Bethe lattice in the presence of spin exchange. AB - Kinetics of phase transformation on a Bethe lattice governed by single-spin-flip Glauber and spin-exchange Kawasaki dynamics is examined. For a general Glauber dynamics for which all processes (splitting and coagulation, growth and decay of clusters, as well as creation and annihilation of single-spin clusters) take place, the addition of the Kawasaki dynamics accelerates the transformation process without changing the qualitative behavior. In the growth-decay regime of the Glauber dynamics, regime in which the splitting and coagulation, and creation and annihilation processes due to single-spin flips are negligible, the Kawasaki dynamics strongly increases the fraction of transformed phase because of the splitting and coagulation of clusters induced by the spin-exchange processes. Acting alone, the Kawasaki dynamics leads to the growth of the clusters of each of the phases after the quenching of the temperature to a lower value. When the final temperature T(f) is smaller than a certain temperature T(f0), the average cluster radius grows linearly with time during both the initial and intermediate stages of the kinetic process, and diverges as log(2)(t(d)-t)(-1) when the time t approaches the value t(d) at which infinite clusters arise. It is shown that, among the various spin-exchange processes involved in Kawasaki dynamics, the main contribution is provided by those which decrease or increase the number of clusters by unity. PMID- 15267997 TI - Nuclear magnetic resonance study of the vapor contribution to diffusion in silica glasses with micrometer pores partially filled with liquid cyclohexane or water. AB - The contribution of the vapor phase to molecular diffusion in porous silica glass (Vitrapor#5; mean pore diameter 1 micrometer) partially filled with cyclohexane (nonpolar) or water (polar) was investigated with the aid of field-gradient NMR diffusometry. Due to the vapor phase, the effective diffusion coefficient of cyclohexane increased up to ten times relative to the value in bulk liquid upon reduction of the pore space filling factor. On the other hand, the effective diffusion coefficient of water first decreases and then increases when the liquid content is reduced. A two-phase exchange theory is presented accounting well for all experimental diffusion features. The diffusion behavior in the samples with micrometer pores under investigation here is in contrast to previous findings for the same solvents in a material with nanometer pores (Vycor; mean pore diameter 4 nm) where the fast-exchange limit had to be assumed [Ardelean et al., J. Chem. Phys. 119, 10358 (2003)]. It is concluded that the pore size plays a crucial role for the relevance of molecular exchange limits relative to the experimental diffusion/exchange time. PMID- 15267998 TI - Flexible polyelectrolyte simulations at the Poisson-Boltzmann level: a comparison of the kink-jump and multigrid configurational-bias Monte Carlo methods. AB - We present a new approach for simulating the motions of flexible polyelectrolyte chains based on the continuous kink-jump Monte Carlo technique coupled to a lattice field theory based calculation of the Poisson-Boltzmann (PB) electrostatic free energy "on the fly." This approach is compared to the configurational-bias Monte Carlo technique, in which the chains are grown on a lattice and the PB equation is solved for each configuration with a linear scaling multigrid method to obtain the many-body free energy. The two approaches are used to calculate end-to-end distances of charged polymer chains in solutions with varying ionic strengths and give similar numerical results. The configurational-bias Monte Carlo/multigrid PB method is found to be more efficient, while the kink-jump Monte Carlo method shows potential utility for simulating nonequilibrium polyelectrolyte dynamics. PMID- 15267999 TI - Thermodynamic properties and aggregate formation of surfactant-like molecules from theory and simulation. AB - The goal of this work is twofold: to predict the phase equilibria behavior of simplified surfactant models and to predict the population of aggregates as a function of pressure. We compare Monte Carlo simulation results of these systems with predictions from a modified version of the statistical associating fluid theory (soft-SAFT). Surfactant-like molecules are modeled as Lennard-Jones chains of tangent segments with one or two association sites. We study the influence of the number and location of the association sites on the thermodynamic properties and fraction of nonbonded molecules in all cases. The influence of the chain length is also investigated for a particular location of the sites. Results are compared with NPT Monte Carlo simulations to test the accuracy of the theory, and to study the molecular configurations of the system. Soft-SAFT is able to quantitatively predict the MC PVT results, independently of the location of the association sites. The theory is also able to capture the qualitative trend of the population of aggregates with pressure. Quantitative agreement is only obtained for specific locations of the sites. PMID- 15268000 TI - Concentration fluctuation effects on the phase behavior of compressible diblock copolymers. AB - A Hartree analysis has been performed for compressible diblock copolymers of incompatible pairs to investigate the concentration fluctuation effects on their microphase separation behavior. The free energy in the Hartree analysis is obtained from the self-consistent correction to its mean-field cousin, which was recently formulated for such copolymer systems. The mean-field phase diagram is shown to be significantly affected by the fluctuation effects as the copolymer chain size N is lowered. An effective interaction chi(cRPA), which carries not only the change in contact interactions but also the compressibility difference between block components, plays a key role in understanding of the phase behavior and the pressure responses of various thermodynamic transitions for the copolymers with finite sizes. In particular, a symmetric copolymer at disorder-to lamella transition is found to satisfy Nchi(cRPA)(q*)=10.495+41.022N(-1/3) when evaluated at a characteristic wave number q* for ordered microphases. PMID- 15268001 TI - Combined Monte Carlo and molecular dynamics simulation of hydrated 18:0 sphingomyelin-cholesterol lipid bilayers. AB - We have carried out atomic level molecular dynamics and Monte Carlo simulations of hydrated 18:0 sphingomyelin (SM)-cholesterol (CHOL) bilayers at temperatures of 20 and 50 degrees C. The simulated systems each contained 266 SM, 122 CHOL, and 11861 water molecules. Each simulation was run for 10 ns under semi-isotropic pressure boundary conditions. The particle-mesh Ewald method was used for long range electrostatic interactions. Properties of the systems were calculated over the final 3 ns. We compare the properties of 20 and 50 degrees C bilayer systems with each other, with experimental data, and with experimental and simulated properties of pure SM bilayers and dipalmitoyl phospatidyl choline (DPPC)-CHOL bilayers. The simulations reveal an overall similarity of both systems, despite the 30 degrees C temperature difference which brackets the pure SM main phase transition. The area per molecule, lipid chain order parameter profiles, atom distributions, and electron density profiles are all very similar for the two simulated systems. Consistent with simulations from our lab and others, we find strong intramolecular hydrogen bonding in SM molecules between the phosphate ester oxygen and the hydroxyl hydrogen atoms. We also find that cholesterol hydroxyl groups tend to form hydrogen bonds primarily with SM carbonyl, methyl, and amide moieties and to a lesser extent methyl and hydroxyl oxygens. PMID- 15268002 TI - Brownian dynamics simulations of needle chain and nugget chain polymer models- rigid constraint conditions versus infinitely stiff springs. AB - It is well known that orientational correlations appear in polymer chain models when the subunits are linked by ball-socket joints implemented as rigid constraint conditions. These correlations do not appear when the subunits are connected by springlike potential forces, even in the limit of infinitely stiff springs. In a widely used class of algorithms for Brownian dynamics simulations, inertia effects are ignored. However, in the recently introduced needle chain and nugget chain algorithms, the rigid constraint correlations depend on the mass and moment of inertia. This inconsistency does not appear in the bead-rod (Kramers) polymer chain model, which also has orientational correlations introduced by rigid constraint conditions. Explicit expressions for the correlation functions are given for thermodynamic equilibrium states. Analytical expressions for the associated forces ("metric forces") and simulation results showing how the rigid constraint correlations influence dynamical properties, are also presented. Further we discuss the physical relevance of these correlations and show via simulations that their influence on stationary and dynamical properties depend significantly on chain length. We further show that if the metric forces are removed, algorithms designed with rigid constraint conditions describe a chain of segments connected by infinitely stiff springs. Finally we show that the results presented here for needle chains are relevant also for the bead-rod (Kramers) chain model, making it possible to simulate a bead-spring chain with infinitely stiff springs. PMID- 15268003 TI - Phase behavior of aqueous solutions containing dipolar proteins from second-order perturbation theory. AB - Due to the interplay of Coulombic repulsion and attractive dipolar and van der Waals interactions, solutions of globular proteins display a rich variety of phase behavior featuring fluid-fluid and fluid-solid transitions that strongly depend on solution pH and salt concentration. Using a simple model for charge, dispersion and dipole-related contributions to the interprotein potential, we calculate phase diagrams for protein solutions within the framework of second order perturbation theory. For each phase, we determine the Helmholtz energy as the sum of a hard-sphere reference term and a perturbation term that reflects both the electrostatic and dispersion interactions. Dipolar effects can induce fluid-fluid phase separation or crystallization even in the absence of any significant dispersion attraction. Because dissolved electrolytes screen the charge-charge repulsion more strongly than the dipolar attraction, the ionic strength dependence of the potential of mean force can feature a minimum at intermediate ionic strengths offering an explanation for the observed nonmonotonic dependence of the phase behavior on salt concentration. Inclusion of correlations between charge-dipole and dipole-dipole interactions is essential for a reliable calculation of phase diagrams for systems containing charged dipolar proteins and colloids. PMID- 15268004 TI - Correlation of femtosecond wave packets and fluorescence interference in a conjugated polymer: Towards the measurement of site homogeneous dephasing. AB - Probing electronic femtosecond (fs) coherence among segmental sites that are congested by static and dynamic site disorder and subject to structural relaxation is a big, experimental challenge in the study of photophysics of poly(p-phenylenevinylene). In this work, fs-wave-packet fluorescence interferometry experiments are presented that measure macroscopic coherent kernels and their phase-relaxation in the low-temperature, bottom-state regime of the density-of-states below the migrational threshold energy where downhill site to-site transfer is marginal. By using freely propagating and tunable 70 fs excitation/probing pulses and employing narrow-band spectral filtering of wave packets, fluorescence interferograms with strongly damped beatings can be observed. The coherences formally follow the in-phase superpositions of two site optical free-induction-decays and originate from distinct pairs of coherent doorway-states, different in energy and space, each of them being targeted, by two discrete quantum-arrival-states 1(alpha) and 1(beta), via independent, isoenergetic 0-->1 fluorescence transitions. The coherent transients are explained as site-to-site polarization beatings, caused by the interference of two fluorescence correlation signals. The numerical analysis of the damping regime, based upon second-order perturbational solutions, reveals the lower limit value of homogeneous dephasing in the range from T(2) approximately 100 fs to T(2) approximately 200 fs depending on the site-excitation energy of the bottom states. The experiments enable to look into the formation of the relaxed state as a special molecular process of electron-phonon coupling and hence open-up a quite new perspective in the puzzle of multichromophore optical dynamics and structural relaxation in conjugated polymers. PMID- 15268005 TI - Comment on "An exact quantum Monte Carlo calculation of the helium-helium intermolecular potential" [J. Chem. Phys. 115, 4546 (2001)]. PMID- 15268006 TI - Preparation and photoelectron spectrum of the glycine molecular anion: assignment to a dipole-bound electron species with a high-dipole moment, non-zwitterionic form of the neutral core. AB - We report the gas-phase preparation of negatively charged glycine as well as the Gly(H(2)O)(1,2) (-) complexes by entrainment of the neutral precursor into an ionized supersonic expansion tuned to optimize the (H(2)O)(n) (-)Ar(m) clusters. The photoelectron spectrum of Gly(-) displays the signature of a dipole-bound species, with sufficient vibrational fine structure to characterize the core neutral as a higher energy, non-zwitterionic isomer of the amino acid. PMID- 15268007 TI - Many-body force field models based solely on pairwise Coulomb screening do not simultaneously reproduce correct gas-phase and condensed-phase polarizability limits. AB - It is demonstrated that many-body force field models based solely on pairwise Coulomb screening cannot simultaneously reproduce both gas-phase and condensed phase polarizability limits. Several many-body force field model forms are tested and compared with basis set-corrected ab initio results for a series of bifurcated water chains. Models are parameterized to reproduce the ab initio polarizability of an isolated water molecule, and pairwise damping functions are set to reproduce the polarizability of a water dimer as a function of dimer separation. When these models are applied to extended water chains, the polarization is over-predicted, and this over-polarization increased as a function of the overlap of molecular orbitals as the chains are compressed. This suggests that polarizable models based solely on pairwise Coulomb screening have some limitations, and that coupling with non-classical many-body effects, in particular exchange terms, may be important. PMID- 15268008 TI - A wave packet based statistical approach to complex-forming reactions. AB - A wave packet based statistical model is suggested for complex-forming reactions. This model assumes statistical formation and decay of the long-lived reaction complex and computes reaction cross sections and their energy dependence from capture probabilities. This model is very efficient and reasonably accurate for reactions dominated by long-lived resonances, as confirmed by its application to the C((1)D)+H(2) reaction. PMID- 15268009 TI - Minima hopping: an efficient search method for the global minimum of the potential energy surface of complex molecular systems. AB - A method is presented that can find the global minimum of very complex condensed matter systems. It is based on the simple principle of exploring the configurational space as fast as possible and of avoiding revisiting known parts of this space. Even though it is not a genetic algorithm, it is not based on thermodynamics. The efficiency of the method depends strongly on the type of moves that are used to hop into new local minima. Moves that find low-barrier escape-paths out of the present minimum generally lead into low energy minima. PMID- 15268010 TI - Accurate reaction paths using a Hessian based predictor-corrector integrator. AB - Central to the theoretical description of a chemical reaction is the reaction pathway. The intrinsic reaction coordinate is defined as the steepest descent path in mass weighted Cartesian coordinates that connects the transition state to reactants and products. In this work, a new integrator for the steepest descent pathway is presented. This method is a Hessian based predictor-corrector algorithm that affords pathways comparable to our previous fourth order method at the cost of a second order approach. The proposed integrator is tested on an analytic surface, four moderately sized chemical reactions, and one larger organometallic system. PMID- 15268011 TI - Coarse-grained controllability of wavepackets by free evolution and phase shifts. AB - We describe an approach to controlling wavepacket dynamics and a criterion of wavepacket controllability based on discretized properties of the wavepacket's localization on the orbit. The notion of "coarse-grained control" and the coarse grained description of the controllability in infinite-dimensional Hilbert spaces are introduced and studied using the mathematical apparatus of loop groups. We prove that 2D rotational wavepackets are controllable by only free evolution and phase kicks by AC Stark shift implemented at fractional revivals. This scheme works even if the AC Stark shifts can have only a smooth coordinate dependence, correspondent to the action of a linearly polarized laser field. PMID- 15268012 TI - Electric fields with ultrasoft pseudo-potentials: applications to benzene and anthracene. AB - We present density-functional perturbation theory for electric field perturbations and ultra-soft pseudopotentials. Applications to benzene and anthracene molecules and surfaces are reported as examples. We point out several issues concerning the evaluation of the polarizability of molecules and slabs with periodic boundary conditions. PMID- 15268013 TI - Efficient chemical kinetic modeling through neural network maps. AB - An approach to modeling nonlinear chemical kinetics using neural networks is introduced. It is found that neural networks based on a simple multivariate polynomial architecture are useful in approximating a wide variety of chemical kinetic systems. The accuracy and efficiency of these ridge polynomial networks (RPNs) are demonstrated by modeling the kinetics of H(2) bromination, formaldehyde oxidation, and H(2)+O(2) combustion. RPN kinetic modeling has a broad range of applications, including kinetic parameter inversion, simulation of reactor dynamics, and atmospheric modeling. PMID- 15268014 TI - Decomposition of nuclear magnetic resonance spin-spin coupling constants into active and passive orbital contributions. AB - The theory of the J-OC-PSP (decomposition of J into orbital contributions using orbital currents and partial spin polarization) method is derived to distinguish between the role of active, passive, and frozen orbitals on the nuclear magnetic resonance (NMR) spin-spin coupling mechanism. Application of J-OC-PSP to the NMR spin-spin coupling constants of ethylene, which are calculated using coupled perturbed density functional theory in connection with the B3LYP hybrid functional and a [7s,6p,2d/4s,2p] basis set, reveal that the well-known pi mechanism for Fermi contact (FC) spin coupling is based on passive pi orbital contributions. The pi orbitals contribute to the spin polarization of the sigma orbitals at the coupling nuclei by mediating spin information between sigma orbitals (spin-transport mechanism) or by increasing the spin information of a sigma orbital by an echo effect. The calculated FC(pi) value of the SSCC (1)J(CC) of ethylene is 4.5 Hz and by this clearly smaller than previously assumed. PMID- 15268015 TI - On the importance of the "density per particle" (shape function) in the density functional theory. AB - The central role of the shape function sigma(r) from the density functional theory (DFT), the ratio of the electron density rho(r) and the number of electrons N of the system (density per particle), is investigated. Moreover, its relationship with DFT based reactivity indices is established. In the first part, it is shown that an estimate for the chemical hardness can be obtained from the long range behavior of the shape function and its derivative with respect to the number of electrons at a fixed external potential. Next, the energy of the system is minimized with the constraint that the shape function should integrate to unity; the associated Lagrange multiplier is shown to be related to the electronic chemical potential micro of the system. Finally, the importance of the shape function for both molecular structure, reactivity, and similarity is outlined. PMID- 15268016 TI - Highly accurate evaluation of atomic three-electron integrals of lowest orders. AB - Calculations of three-electron atomic systems in Hylleraas coordinates require integrals involving all the interparticle distances r(ij), which have usually been evaluated by introducing series expansions. For integrals with the smallest powers of r(ij) these expansions do not converge at a satisfactory rate, leading some investigators to introduce convergence-acceleration procedures. This paper recommends the alternative of evaluating these integrals in closed form and presents stable explicit formulas for so doing. Some of the formulas are more compact versions of those in the literature; others have not been previously reported. It is also shown that finite-difference methods can be used with advantage to obtain additional low-order integrals. Sample integral values have been provided for test purposes. PMID- 15268017 TI - Measurement-assisted coherent control. AB - Two advantageous roles of the influence of measurement on a system subject to coherent control are exposed using a five-level model system. In particular, a continuous measurement of the population in a branch state in the Kobrak-Rice extended stimulated Raman adiabatic passage scheme is shown to provide a powerful means for controlling the population transfer branching ratio between two degenerate target states. It is demonstrated that a measurement with a large strength may be used to completely shut off the yield of one target state and that the same measurement with a weak strength can dramatically enhance the robustness of the controlled branching ratio against dephasing. PMID- 15268018 TI - Quantum scattering and adiabatic channel treatment of the low-energy and low temperature capture of a rotating quadrupolar molecule by an ion. AB - The capture rate coefficients of homonuclear diatomic molecules (H(2) and N(2)) in the rotational state j=1 interacting with ions (Ar+ and He+) are calculated for low collision energies assuming a long-range anisotropic ion-induced dipole and ion-quadrupole interaction. A comparison of accurate quantum rates with quantum and state-specific classical adiabatic channel approximations shows that the former becomes inappropriate in the case when the cross section is dominated by few partial contributions, while the latter performs better. This unexpected result is related to the fact that the classical adiabatic channel approximation artificially simulates the quantum effects of tunneling and overbarrier reflection as well as the Coriolis coupling and it suppresses too high values of the centrifugal barriers predicted by a quantum adiabatic channel approach. For H2(j=1)+Ar+ and N(2)(j=1)+He+ capture, the rate constants at T-->0 K are about 3 and 6 times higher than the corresponding values for H2(j=0)+Ar+ and N(2)(j=0)+He+ capture. PMID- 15268019 TI - The gas-phase chemiionization reaction between samarium and oxygen atoms: a theoretical study. AB - The Sm+O chemiionization reaction has been investigated theoretically using a method that allows for correlation and relativistic effects. Potential energy curves have been calculated for several electronic states of SmO and SmO+. Comparison with available spectroscopic and thermodynamic values for these species is reported and a mechanism for the chemiionization reaction Sm+O is proposed. The importance of spin-orbit coupling in the excited states of SmO, in allowing this chemiionization reaction to take place, has been revealed by these calculations. This paper shows the metal-plus-oxidant chemiionization reaction. PMID- 15268020 TI - Fourier transform infrared emission spectra of MgH and MgD. AB - High resolution Fourier transform infrared emission spectra of MgH and MgD have been recorded. The molecules were generated in an emission source that combines an electrical discharge with a high temperature furnace. Several vibration rotation bands were observed for all six isotopomers in the X (2)Sigma(+) ground electronic state: v=1-->0 to 4-->3 for (24)MgH, v=1-->0 to 3-->2 for (25)MgH and (26)MgH, v=1-->0 to 5-->4 for (24)MgD, v=1-->0 to 4-->3 for (25)MgD and (26)MgD. The new data were combined with the previous ground state data, obtained from diode laser vibration-rotation measurements and pure rotation spectra, and spectroscopic constants were determined for the v=0 to 4 levels of (24)MgH and the v=0 to 5 levels of (24)MgD. In addition, Dunham constants and Born Oppenheimer breakdown correction parameters were obtained in a combined fit of the six isotopomers. The equilibrium vibrational constants (omega(e)) for (24)MgH and (24)MgD were found to be 1492.776(7) cm(-1) and 1077.298(5) cm(-1), respectively, while the equilibrium rotational constants (B(e)) are 5.825 523(8) cm(-1) and 3.034 344(4) cm(-1). The associated equilibrium bond distances (r(e)) were determined to be 1.729 721(1) A for (24)MgH and 1.729 157(1) A for (24)MgD. PMID- 15268021 TI - The outer valance orbital electron densities of cyclopentane by binary (e,2e) spectroscopy. AB - The binding energy spectra and electron distributions in momentum space of the valence orbitals of cyclopentane (C(5)H(10)) are studied by Electron Momentum Spectroscopy (EMS) in a noncoplanar symmetric geometry. The impact energy was 1200 eV plus binding energy and energy resolution of the EMS spectrometer was 1.2 eV. The experimental momentum profiles of the outer valence orbitals are compared with the theoretical momentum distributions calculated using Hartree-Fock and density functional theory (DFT) methods. The shapes of the experimental momentum distributions are generally quite well described by both the Hartree-Fock and DFT calculations when the large and diffuse basis sets are used. PMID- 15268022 TI - Intra- and intermolecular energy transfer in highly excited ozone complexes. AB - The energy transfer of highly excited ozone molecules is investigated by means of classical trajectories. Both intramolecular energy redistribution and the intermolecular energy transfer in collisions with argon atoms are considered. The sign and magnitude of the intramolecular energy flow between the vibrational and the rotational degrees of freedom crucially depend on the projection K(a) of the total angular momentum of ozone on the body-fixed a axis. The intermolecular energy transfer in single collisions between O(3) and Ar is dominated by transfer of the rotational energy. In accordance with previous theoretical predictions, the direct vibrational de-excitation is exceedingly small. Vibration-rotation relaxation in multiple Ar+O(3) collisions is also studied. It is found that the relaxation proceeds in two clearly distinguishable steps: (1) During the time between collisions, the vibrational degrees of freedom are "cooled" by transfer of energy to rotation; even at low pressure equilibration of the internal energy is slow compared to the time between collisions. (2) In collisions, mainly the rotational modes are "cool" by energy transfer to argon. PMID- 15268023 TI - Investigation of photoinduced electron transfer in model system of vitamin E duroquinone by time-dependent density functional theory. AB - Photoinduced electron transfer of the model system composed of vitamin E and duroquinone has been investigated using time-dependent density functional theory. Calculations for the excited states tell that the photoexcitation of the model system can directly yield the charge transfer states in which the vitamin E moiety is positively charged but the duroquinone moiety is negatively charged. Our theoretical investigations indicate that the second charge transfer state of the model system can also be produced through the decay of higher locally excited state S(4). Since S(4) state in the model system corresponds to S(1) state of the isolated duroquinone used as a model for peroxyl radical, and S(2) state has the character of electron transfer from the tertiary amine group of the vitamin E moiety to the duroquinone moiety, the decay from S(4) to S(2) corresponds to the dynamic process following the photoexcitation of the duroquinone moiety of the model system, i.e., the initial stage of antioxidant reaction of vitamin E. Calculations of the kinetic parameters for the electron transfer have been carried out in the framework of the Marcus-Jortner-Levich formalism. Our calculations confirm that the electron transfer from S(4) to S(2) possesses the character of the inverted regime and the barrier is negligibly small. PMID- 15268024 TI - Regularities in molecular properties of ground state stable diatomics. AB - A simple relationship is reported between vibrational frequencies, bond lengths, and reduced masses for many families of stable, ground state diatomics: the frequency is proportional to the reciprocal of the product of the bond length and the square root of the reduced mass. This is demonstrated with each of the following related families: the alkali metal diatomics, the group 15 diatomics, the group 16 diatomics, the halogen diatomics, the alkali metal hydrides, the alkaline earth oxides, the group 14 oxides and their sulfides, the diatomics of carbon, of silicon and of germanium with group 16 elements, the hydrogen halides, the halides of lithium, of sodium, of potassium, of rubidium and of cesium, the chlorides of the alkali metals and of silver, and the polyatomic hydrides of groups 14 and 15. Although correlation coefficients of 0.99 or greater in each of the 21 families examined demonstrate the validity of the correlation, the deviations found are significantly larger than can be attributed to experimental uncertainties. PMID- 15268025 TI - Infrared photodissociation spectroscopy of V+(CO2)n and V+(CO2)nAr complexes. AB - V+(CO2)n and V+(CO2)nAr complexes are generated by laser vaporization in a pulsed supersonic expansion. The complexes are mass-selected within a reflectron time-of flight mass spectrometer and studied by infrared resonance-enhanced (IR-REPD) photodissociation spectroscopy. Photofragmentation proceeds exclusively through loss of intact CO2 molecules from V+(CO2)n complexes or by elimination of Ar from V+(CO2)nAr mixed complexes. Vibrational resonances are identified and assigned in the region of the asymmetric stretch of free CO2 at 2349 cm(-1). A linear geometry is confirmed for V+(CO2). Small complexes have resonances that are blueshifted from the asymmetric stretch of free CO2, consistent with structures in which all ligands are bound directly to the metal ion. Fragmentation of the larger clusters terminates at the size of n=4, and a new vibrational band at 2350 cm(-1) assigned to external ligands is observed for V+(CO2)5 and larger cluster sizes. These combined observations indicate that the coordination number for CO2 molecules around V+ is exactly four. Fourfold coordination contrasts with that seen in condensed phase complexes, where a coordination number of six is typical for V+. The spectra of larger complexes provide evidence for an intracluster insertion reaction that produces a metal oxide-carbonyl species. PMID- 15268026 TI - A discrete time-dependent method for metastable atoms and molecules in intense fields. AB - The full-dimensional time-dependent Schrodinger equation for the electronic dynamics of single-electron systems in intense external fields is solved directly using a discrete method. Our approach combines the finite-difference and Lagrange mesh methods. The method is applied to calculate the quasienergies and ionization probabilities of atomic and molecular systems in intense static and dynamic electric fields. The gauge invariance and accuracy of the method is established. Applications to multiphoton ionization of positronium, the hydrogen atom and the hydrogen molecular ion are presented. At very high laser intensity, above the saturation threshold, we extend the method using a scaling technique to estimate the quasienergies of metastable states of the hydrogen molecular ion. The results are in good agreement with recent experiments. PMID- 15268027 TI - Infrared spectra of X-.CO(2).Ar cluster anions (X=Cl,Br,I). AB - Ion-molecule clusters of the heavier halide anions X-.CO(2) (X=Cl-,Br-,I-) with CO2 have been studied by gas phase infrared photodissociation spectroscopy, using Ar evaporation from the complexes X-.CO2.Ar upon infrared excitation. We observe that the asymmetric stretch vibrational mode of the CO(2) molecule is red-shifted from the frequency of free CO2, with the red-shift increasing toward the lighter halide ions. A similar trend is repeated in the region of the Fermi resonance of the combination bands of the asymmetric stretch vibration with two quanta of the bending vibration and the symmetric stretch vibration. We discuss our findings in the framework of ab initio and density functional theory calculations. PMID- 15268028 TI - Interaction of benzene thiol and thiolate with small gold clusters. AB - We studied the interaction between benzene thiol and thiolate molecules, and gold clusters made of 1 to 3 atoms, by means of ab initio density functional theory in the local density approximation. We find that the thiolate is energetically more stable than the thiol, however the process of detachment of H from the thiol appears to be possibly mediated by the intermediate step of H chemisorption on Au. Cleavage of the S-H bond is accompanied by a 90 degrees rotation of the molecule around the S-Au bond, showing a strong steric specificity. Such a rotation is induced by the relative energy shift of the S atom p orbitals with respect to the benzene pi ring and the Au d orbitals. By analyzing the correlation of the bond energy, bond lengths, and HOMO-LUMO gap with the number of S-Au bonds, we find that the thiolate S atom appears to prefer a low coordination condition on Au clusters. PMID- 15268029 TI - Jahn-Teller effect in van der Waals complexes; Ar-C6H6 + and Ar-C6D6 +. AB - The two asymptotically degenerate potential energy surfaces of argon interacting with the X (2)E(1g) ground state benzene(+) cation were calculated ab initio from the interaction energy of the neutral Ar-benzene complex given by Koch et al. [J. Chem. Phys. 111, 198 (1999)] and the difference of the geometry-dependent ionization energies of the complex and the benzene monomer computed by the outer valence Green's function method. Coinciding minima in the two potential surfaces of the ionic complex occur for Ar on the C(6v) symmetry axis of benzene(+) (the z axis) at z(e)=3.506 A. The binding energy D(e) of 520 cm(-1) is only 34% larger than the value for the neutral Ar-benzene complex. The higher one of the two surfaces is similar in shape to the neutral Ar-benzene potential, the lower potential is much flatter in the (x,y) bend direction. Nonadiabatic (Jahn-Teller) coupling was taken into account by transformation of the two adiabatic potentials to a two-by-two matrix of diabatic potentials. This transformation is based on the assumption that the adiabatic states of the Ar-benzene(+) complex geometrically follow the Ar atom. Ab initio calculations of the nonadiabatic coupling matrix element between the adiabatic states with the two-state-averaged CAS-SCF(5,6) method confirmed the validity of this assumption. The bound vibronic states of both Ar-C(6)H(6) (+) and Ar-C(6)D(6) (+) were computed with this two state diabatic model in a basis of three-dimensional harmonic oscillator functions for the van der Waals modes. The binding energy D(0)=480 cm(-1) of the perdeuterated complex agrees well with the experimental upper bound of 485 cm( 1). The ground and excited vibronic levels and wave functions were used, with a simple model dipole function, to generate a theoretical far-infrared spectrum. Strong absorption lines were found at 10.1 cm(-1) (bend) and 47.9 cm(-1) (stretch) that agree well with measurements. The unusually low bend frequency is related to the flatness of the lower adiabatic potential in the (x,y) direction. The van der Waals bend mode of e(1) symmetry is quadratically Jahn-Teller active and shows a large splitting, with vibronic levels of A(1), E(2), and A(2) symmetry at 1.3, 10.1, and 50.2 cm(-1). The level at 1.3 cm(-1) leads to a strong absorption line as well, which could not be measured because it is too close to the monomer line. The level at 50.2 cm(-1) gives rise to weaker absorption. Several other weak lines in the frequency range of 10 to 60 cm(-1) were found. PMID- 15268030 TI - Systematic ab initio calculations on the energetics and stability of covalent O4. AB - Ab initio calculations with highly correlated methods together with extensive basis sets have been used to obtain the most accurate heat of formation and stability with respect to dissociation (into molecular oxygen) for the chemically bound tetraoxygen molecule. Our calculations show that the heat of formation is significantly smaller and that the barrier to dissociation is larger than previously assumed. In particular, we have shown that the previous theoretical estimate for the heat of formation of tetraoxygen was in error by a significant amount (18%-24%) owing to lack of accuracy in the theoretical method then used. Our best estimates places that value in the range 93-95 kcal/mol and this should be taken into consideration when discussing the possible relevance of tetraoxygen in a variety of experiments, as well as in the fundamental atmospheric chemical processes where oxygen species participate. PMID- 15268031 TI - A theoretical study of the fine and hyperfine interactions in the NCO and CNO radicals. AB - The geometries, the harmonic vibrational frequencies, and the Renner-Teller parameter have been reported for the NCO(+)(X (3)Sigma(-)), NCO(X (2)Pi,A (2)Sigma(+),B (2)Pi,2 (2)Sigma(+)), NCO(-)(X (1)Sigma(+)), CNO(+)(X), CNO(X (2)Pi,A (2)Sigma(+),B (2)Pi,2 (2)Sigma(+)), and CNO(-)(X (1)Sigma(+)) systems at the full valence-complete active space self-consistent-field (fv-CASSCF) level of theory. The (2)Pi electronic states of the NCO and CNO radicals have two distinct real vibrational frequencies for the bending modes and these states are subject to the type A Renner-Teller effect. The total energy of CNO(+) without zero point energy correction of the linear geometry is approximately 31 cm(-1) higher than the bent geometry at the fv-CASSCF level and the inversion barrier vanishes after the zero point energy correction; therefore, the ground state of the CNO(+) may possess a quasilinear geometry. The spin-orbit coupling constants estimated using atomic mean field Hamiltonian at the fv-CASSCF level of theory are in better agreement with the experimental values. The excitation energies, the electron affinity, and the ionization potential have been computed at the complete active space second order perturbation theory (CASPT2) and the multireference singles and doubles configuration (MRSD-CI) levels of theory. The computed values of the electric hyperfine coupling constants for the (14)N atom in the ground state of the NCO radical agree well with the experimental data. The magnetic hyperfine coupling constants (HFCC's) have been estimated employing the configuration selected MRSD-CI and the multireference singles configuration interaction (MRS CI) methods using iterative natural orbitals (ino) as one particle basis. Sufficiently accurate value of the isotropic contribution to the HFCC's can be obtained using an MRS-CI-ino procedure. PMID- 15268032 TI - Hydrogen transfer in excited pyrrole-ammonia clusters. AB - The excited state hydrogen atom transfer reaction (ESHT) has been studied in pyrrole-ammonia clusters [PyH-(NH(3))(n)+hnu-->Py.+.NH(4)(NH(3))(n-1)]. The reaction is clearly evidenced through two-color R2P1 experiments using delayed ionization and presents a threshold around 235 nm (5.3 eV). The cluster dynamics has also been explored by picosecond time scale experiments. The clusters decay in the 10-30 ps range with lifetimes increasing with the cluster size. The appearance times for the reaction products are similar to the decay times of the parent clusters. Evaporation processes are also observed in competition with the reaction, and the cluster lifetime after evaporation is estimated to be around 10 ns. The kinetic energy of the reaction products is fairly large and the energy distribution seems quasi mono kinetic. These experimental results rule out the hypothesis that the reaction proceeds through a direct N-H bond rupture but rather imply the existence of a fairly long-lived intermediate state. Calculations performed at the CASSCF/CASMP2 level confirm the experimental observations, and provide some hints regarding the reaction mechanism. PMID- 15268033 TI - Effect of anisotropic diffusion and external electric field on the rate of diffusion-controlled reactions. AB - In this paper we investigate theoretically the effect of an external electric field on the rate constant of steady-state bulk diffusion-controlled reactions. We generalize previously derived results for isotropic diffusion in the absence of interparticle interaction [J. Chem. Phys. 87, 4622 (1987)] to the case where translational diffusion is anisotropic. A frequently occurring situation of transverse isotropy where D(x)=D(y) not equal to D(z) is considered in detail. We derive the first-order expansion for the reaction rate constant in terms of the electric field strength E, k(E)=k(0) (1+1/2epsilongamma), where gamma=k(0)/4piRD( perpendicular ), epsilon=qER/k(B)T, q is the charge, R is the contact distance, and D( perpendicular ) is the transverse diffusion coefficient. Numerical calculations show that this first-order expansion works well in the whole range of applicability of the Nernst-Einstein relation, i.e., for epsilon<1. PMID- 15268034 TI - Sum-frequency vibrational spectroscopy of chiral liquids off and close to electronic resonance and the antisymmetric Raman tensor. AB - The strength of the chiral vibrational peaks in infrared-visible sum-frequency (SF) vibrational spectra from isotropic chiral liquids is proportional to the square of the corresponding antisymmetric Raman element. Under the Born Oppenheimer adiabatic approximation with nonadiabatic corrections, the antisymmetric Raman tensor is much weaker than the symmetric counterpart, but becomes significantly stronger as the input frequency (or the sum-frequency in SF generation) approaches electronic resonance. We verify the theory with experimental results obtained from infrared-visible doubly resonant sum-frequency generation from an isotropic solution of chiral molecules. PMID- 15268035 TI - Transition state dynamics of N2O4 <==> 2NO2 in liquid state. AB - Transition state dynamics of dissociation and association reactions N2O4 <==> 2NO2 in liquid state are studied by classical molecular dynamics simulations of reactive liquid NO(2) at 298 K. An OSPP+LJ potential between NO(2) molecules proposed in paper I [J. Chem. Phys. 115, 10852 (2001)], which takes into account the orientational sensitivity of the chemical bond, has been used in the simulation. The trajectory and energy evolution of various reactions are studied in the OSPP+LJ liquid, which reproduces both the observed liquid phase equilibrium constant and Raman band shape of the dissociation mode. It is found that a NO(2) pair in reactive liquid NO(2) is bound when E(T)<0 and dissociates when E(T)>0, and the dissociation of a reactant pair occurs when the transition state (TS) surface of E(T)=0 is crossed from negative to positive, where E(T) is the sum of the potential and kinetic energies of intermolecular motion of the pair. Two types of dissociation are found depending on the source of energy for dissociation; the first type D is the dissociation via collisional activation of the reactive mode by solvent molecules, and the second type T is the dissociation via bond transfer from a dimer to a monomer NO(2) through the TS of NO(2) trimer. It is concluded that the type T dissociation is found to be much more probable than the type D dissociation because of easy energy conservation. The reactant experiences the TS of NO(2) trimer for a long time (1-10 ps) in NO(2) mediated bond transfer reactions, and crossing and recrossing trajectories and dynamics in the TS neighborhood are studied. PMID- 15268036 TI - Phase coexistence in polydisperse charged hard-sphere fluids: mean spherical approximation. AB - Taking advantage of the availability of the analytic solution of the mean spherical approximation for a mixture of charged hard spheres with an arbitrary number of components we show that the polydisperse fluid mixture of charged hard spheres belongs to the class of truncatable free energy models, i.e., to those systems where the thermodynamic properties can be represented by a finite number of (generalized) moments of the distribution function that characterizes the mixture. Thus, the formally infinitely many equations that determine the parameters of the two coexisting phases can be mapped onto a system of coupled nonlinear equations in these moments. We present the formalism and demonstrate the power of this approach for two systems; we calculate the full phase diagram in terms of cloud and shadow curves as well as binodals and discuss the distribution functions of the coexisting daughter phases and their charge distributions. PMID- 15268037 TI - Early chemistry in hot and dense nitromethane: molecular dynamics simulations. AB - We report density functional molecular dynamic simulations to determine the early chemical events of hot (T=3000 K) and dense (rho=1.97 g/cm(3), V/V(0)=0.68) nitromethane (CH(3)NO(2)). The first step in the decomposition process is an intermolecular proton abstraction mechanism that leads to the formation of CH(3)NO(2)H(+) and the aci ion H(2)CNO(2) (-). This event is also confirmed to occur in a fast annealing simulation to a final temperature of 4000 K at rho=2.20 g/cm(3). An intramolecular hydrogen transfer that transforms nitromethane into the aci acid form, CH(2)NO(2)H, accompanies this event. To our knowledge, this is the first confirmation of chemical reactivity with bond selectivity for an energetic material near the Chapman-Jouget state of the fully reacted material. We also report the decomposition mechanism followed up to the formation of H(2)O as the first stable product. We note that similarities in the global features of reactants, intermediates, and products of the reacting fluid seem to indicate a threshold for similar chemistry in the range of high densities and temperatures reported herein. PMID- 15268038 TI - Ions in water: the microscopic structure of concentrated NaOH solutions. AB - A neutron diffraction experiment with isotopic H/D substitution on four concentrated NaOH/H(2)O solutions is presented. The full set of partial structure factors is extracted, by combining the diffraction data with a Monte Carlo simulation. These allow to investigate both the changes of the water structure in the presence of ions and their solvation shells. It is found that the interaction with the solute affects the tetrahedral network of hydrogen bonded water molecules in a manner similar to the application of high pressure to pure water. The solvation shell of the OH(-) ions has an almost concentration independent structure, although with concentration dependent coordination numbers. The hydrogen site coordinates a water molecule through a weak bond, while the oxygen site forms strong hydrogen bonds with a number of molecules that is on the average very close to four at the higher water concentrations and decreases to about three at the lowest one. The competition between hydrogen bond interaction and Coulomb forces in determining the orientation of water molecules within the cation solvation shell is visible in the behavior of the g(NaHw)(r) function PMID- 15268039 TI - The structure of methane hydrate under geological conditions a combined Rietveld and maximum entropy analysis. AB - We present a study of the structure of a fully deuterated methane hydrate under the geological conditions found in the world's oceans. In situ high-resolution neutron diffraction experiments have been performed at temperatures of 220, 275, and 280 K and a pressure of 100 bar, corresponding to the conditions at 1000 m water depth. The data were analyzed with a combination of Rietveld refinement and maximum entropy methods. From the Rietveld refinement, precise atomic parameters of the host lattice could be determined, indicating increasing distortions of the structure of the cages at elevated temperatures and pressures. Debye-Waller factors of the encaged CD(4) molecules have been found to exceed the values of the Debye-Waller factors of the D(2)O molecules considerably. In the large cage of structure type I the thermal center-of-mass displacements of the guests are 5 10 times larger than those of the water molecules. From the maximum entropy analysis maps of the scattering length density have been obtained, showing details of the vibrational amplitudes of the atoms in methane hydrate. The Debye Waller factors of all molecules have been found to deviate considerably from a simple spherical geometry. PMID- 15268040 TI - Structural properties of a calcium aluminosilicate glass from molecular-dynamics simulations: a finite size effects study. AB - We study a calcium aluminosilicate glass of composition (SiO(2))(0.67) (Al(2)O(3))(0.12)-(CaO)(0.21) by means of molecular-dynamics simulations, using a potential made of two-body and three-body interactions. In order to prepare small samples that can subsequently be studied by first principles, the finite size effects on the liquid dynamics and on the glass structural properties are investigated. We find that finite size effects affect the Si-O-Si and Si-O-Al angular distributions, the first peaks of the Si-O, Al-O, and Ca-O pair correlation functions, the Ca coordination, and the oxygen atoms' environment in the smallest system (100 atoms). We give evidence that these finite size effects can be directly attributed to the use of three-body interactions. PMID- 15268041 TI - Spinodal and ideal glass limits for metastable liquids. AB - A spinodal line provides a high-temperature limit to the stability of a superheated liquid and a line of ideal glass transitions may provide a low temperature limit for a supercooled liquid. For models in which the thermodynamic properties depend on only one independent external variable the line of ideal glass transition can be viewed as a second-order transition between two equilibrated phases and it meets the spinodal with the same slope at the maximum tension a stretched liquid can sustain. For real materials there are two independent external variables, temperature and pressure; the line of ideal glass transitions cannot be viewed as a second-order phase transition and the two lines need not have the same slope where they meet. PMID- 15268042 TI - Shear stress relaxation in liquids. AB - We show that at high densities, as the system size decreases, liquid becomes able to permanently sustain increasing internal shear stress after a constant deformation, although the other characteristic liquid properties, such as the pair distribution function and diffusion coefficient do not change under strain. The system size necessary for observation of this effect increases with the decrease in temperature, and it is stronger in pair potentials with steeper repulsive part. We relate this result to the size of the "cooperatively rearranging regions" of the Adam-Gibbs theory of glass transition. PMID- 15268043 TI - Negative contributions in the velocity correlation function of supercooled liquid water. AB - The translational dynamics of supercooled and normal liquid water is investigated via a specific correlation function DeltaB with the aim of explaining the behavior of the centers of mass velocity correlation function (VCF). DeltaB is divided into diffusive and nondiffusive parts that yield separated contributions to the VCF, namely an Enskog-type diffusive one, modeled by an exponential function, and a nondiffusive one, made up by damped oscillations of a vanishing time integral. In the translational density of states (DOS), the oscillations yield the bands at omega(1) congruent with 50 cm(-1), omega(3) congruent with 240 cm(-1) (the two well-known experimental bands of the Raman spectra) and omega(2) congruent with 160 cm(-1) (the Einstein frequency of the liquid). It is shown that the chief negative lobe of the VCF is mainly due to the DOS component at the lowest frequency omega(1). The study of the relative pair dynamics shows that this lobe is due to the transverse dynamics, while the longitudinal one determines the fast DOS component at omega(3). The presence of a negative tail is highlighted. Its contribution extends beyond the region of the fast dynamics (t<0.7 ps) up to about 1.5 ps and is due to a low-frequency oscillating mode that produces a low-frequency DOS band centered at about omega(0)=20 cm(-1). PMID- 15268044 TI - The nuclear magnetic resonance line shapes of Xe in the cages of clathrate hydrates. AB - We report, for the first time, a prediction of the line shapes that would be observed in the (129)Xe nuclear magnetic resonance (NMR) spectrum of xenon in the cages of clathrate hydrates. We use the dimer tensor model to represent pairwise contributions to the intermolecular magnetic shielding tensor for Xe at a specific location in a clathrate cage. The individual tensor components from quantum mechanical calculations in clathrate hydrate structure I are represented by contributions from parallel and perpendicular tensor components of Xe-O and Xe H dimers. Subsequently these dimer tensor components are used to reconstruct the full magnetic shielding tensor for Xe at an arbitrary location in a clathrate cage. The reconstructed tensors are employed in canonical Monte Carlo simulations to find the Xe shielding tensor component along a particular magnetic field direction. The shielding tensor component weighted according to the probability of finding a crystal fragment oriented along this direction in a polycrystalline sample leads to a predicted line shape. Using the same set of Xe-O and Xe-H shielding functions and the same Xe-O and Xe-H potential functions we calculate the Xe NMR spectra of Xe atom in 12 distinct cage types in clathrate hydrates structures I, II, H, and bromine hydrate. Agreement with experimental spectra in terms of the number of unique tensor components and their relative magnitudes is excellent. Agreement with absolute magnitudes of chemical shifts relative to free Xe atom is very good. We predict the Xe line shapes in two cages in which Xe has not yet been observed. PMID- 15268045 TI - Site-specific vibrational dynamics of the CD3zeta membrane peptide using heterodyned two-dimensional infrared photon echo spectroscopy. AB - Heterodyned two-dimensional infrared (2D IR) spectroscopy has been used to study the amide I vibrational dynamics of a 27-residue peptide in lipid vesicles that encompasses the transmembrane domain of the T-cell receptor CD3zeta. Using 1 (13)C[Double Bond](18)O isotope labeling, the amide I mode of the 49-Leucine residue was spectroscopically isolated and the homogeneous and inhomogeneous linewidths of this mode were measured by fitting the 2D IR spectrum collected with a photon echo pulse sequence. The pure dephasing and inhomogeneous linewidths are 2 and 32 cm(-1), respectively. The population relaxation time of the amide I band was measured with a transient grating, and it contributes 9 cm( 1) to the linewidth. Comparison of the 49-Leucine amide I mode and the amide I band of the entire CD3zeta peptide reveals that the vibrational dynamics are not uniform along the length of the peptide. Possible origins for the large amount of inhomogeneity present at the 49-Leucine site are discussed. PMID- 15268046 TI - Physisorption of molecular oxygen on C60 thin films. AB - The interaction of oxygen molecules with a fullerene surface has been studied using high resolution electron energy loss spectroscopy and temperature programmed desorption. Vibrational excitation of the adsorbed oxygen is observed at 190 meV, an energy value comparable with that for molecular oxygen in the gas phase. We take this to indicate physisorption of molecular oxygen on the C(60) surface. Thermal desorption results also show that the bonding of oxygen molecules to the C(60) overlayer is comparable to that on a graphite surface. A detailed study of the energy dependence of the vibrational excitation reveals an inelastic electron resonance scattering process. The angular dependence of the resonant vibrational excitation exhibits features distinctively different from those for molecular oxygen physisorbed on the related graphite surface, at a comparable coverage. One possible reason is that the corrugated surface potential, due to the curvature of the C(60) molecules, promotes the preferential ordering of the physisorbed oxygen molecules perpendicular to the surface plane of the C(60) overlayer. PMID- 15268047 TI - A molecular-dynamics study of structural and physical properties of nitromethane nanoparticles. AB - The structural and physical properties of nanoparticles of nitromethane are studied by using molecular dynamics methods with a previously developed force field. [Agrawal et al., J. Chem. Phys. 119, 9617 (2003).] This force field accurately predicts solid- and liquid-state properties as well as melting of bulk nitromethane. Molecular dynamics simulations of nanoparticles with 480, 240, 144, 96, 48, and 32 nitromethane molecules have been carried out at various temperatures. The carbon-carbon radial distribution function, dipole-dipole correlation function, core density, internal enthalpy, and atomic diffusion coefficients of the nanoparticles were calculated at each temperature. These properties were used to characterize the physical phases and thus determine the melting transitions of the nanoparticles. The melting temperatures predicted by the various properties are consistent with one another and show that the melting temperature increases with particle size, approaching the bulk limit for the largest particle. A size dependence of melting points has been observed in experimental and theoretical studies of atomic nanoparticles, and this is a further demonstration of the effect for large nanoparticles of complex molecular materials. PMID- 15268048 TI - Modification of the surface electronic and chemical properties of Pt(111) by subsurface 3d transition metals. AB - The modification of the electronic and chemical properties of Pt(111) surfaces by subsurface 3d transition metals was studied using density-functional theory. In each case investigated, the Pt surface d-band was broadened and lowered in energy by interactions with the subsurface 3d metals, resulting in weaker dissociative adsorption energies of hydrogen and oxygen on these surfaces. The magnitude of the decrease in adsorption energy was largest for the early 3d transition metals and smallest for the late 3d transition metals. In some cases, dissociative adsorption was calculated to be endothermic. The surfaces investigated in this study had no lateral strain in them, demonstrating that strain is not a necessary factor in the modification of bimetallic surface properties. The implications of these findings are discussed in the context of catalyst design, particularly for fuel cell electrocatalysts. PMID- 15268049 TI - Theoretical studies of the kinetics of methane hydrate crystallization in external electromagnetic fields. AB - Nonequilibrium molecular-dynamics (MD) simulations have been performed for the growth and dissolution of a spherical methane hydrate crystallite, surrounded by a saturated water-methane liquid phase, in both the absence and presence of external electromagnetic (e/m) fields in the microwave to far infrared range (5 7500 GHz) at root-mean square (rms) electric field intensities of up to 0.2 V/A. A rigid/polarizable potential was used to model water and a rigid/nonpolarizable model was utilized for methane. In the absence of a field, it was found that the average growth rate of the crystallite was approximately 0.32 water and 0.045 methane molecules per picosecond, evaluated over a 500 ps NPT simulation for three different initial geometries. Upon the application of an e/m field, it was found that no significant deviations from the zero-field crystal growth patterns were observed for rms electric field intensities of less than about 0.1 V/A, regardless of the field frequency. At, and above, this "threshold" intensity, it was found that dissolution took place. The mobility of the molecules in the system was enhanced by the e/m field, to the greatest extent for frequencies of 50-100 GHz. Furthermore, it was observed that there was a systematic frequency variation in the pattern of dipole alignment with the external field and this led to marked differences in the rate of dissolution. PMID- 15268050 TI - Prototype reflectivity analyses of hydrogen storage levels in single-walled carbon nanotubes. AB - A prototype case study is presented that examines the level of hydrogen content in H-SWNTs using the Surface Plasmon Resonance technique. The damping effect and the angular shift in the resonance minimum of an SWNT-gold interface due to the presence of hydrogen is analyzed using a parametric model, which is based on the concept of an effective permittivity. The new approach provides for a non invasive analysis of the level of hydrogen content in H-SWNTs and is potentially extendable to other carbon-based hydrogen storage materials. PMID- 15268051 TI - Adsorption and vibrational spectroscopy of ammonia at mordenite: ab initio study. AB - The adsorption of ammonia at various active centers at the outer and inner surfaces of mordenite, involving Bronsted acid (BA) sites, terminal silanol groups, and Lewis sites has been investigated using periodic ab initio density functional theory. It is shown that ammonia forms an ammonium ion when adsorbed at strong BA sites. The calculated adsorption energies for different BA sites vary in the interval from 111.5 to 174.7 kJ/mol depending on the local environment of the adduct. The lowest adsorption energy is found for a monodentate complex in the main channel, the highest for a tetradentate configuration in the side pocket. At weak BA sites such as terminal silanol groups or a defect with a BA site in a two-membered ring ammonia is H bonded via the N atom. Additional weak H bonds are formed between H atoms of ammonia and O atoms of neighboring terminal silanol groups. The calculated adsorption energies for such adducts range between 61.7 and 70.9 kJ/mol. The interaction of ammonia with different Lewis sites is shown to range between weak (DeltaE(ads)=17.8 kJ/mol) and very strong (DeltaE(ads)=161.7 kJ/mol), the strongest Lewis site being a tricoordinated Al atom at the outer surface. Our results are in very good agreement with the distribution of desorption energies estimated from temperature programmed desorption (TPD) and microcalorimetry experiments, the multipeaked structure of the TPD spectra is shown to arise from strong and weak Bronsted and Lewis sites. The vibrational properties of the adsorption complexes are investigated using a force-constant approach. The stretching and bending modes of NH(4) (+) adsorbed to the zeolite are strongly influenced by the local environment. The strongest redshift is calculated for the asymmetric stretching mode involving the NH group hydrogen bonded to the bridging O atom of the BA site, the shift is largest for a monodentate and smallest for a tetradentate adsorption complex. The reduced symmetry of the adsorbate also leads to a substantial splitting of the stretching and bending modes. In agreement with experiment we show that the main vibrational feature which differentiates coordinatively bonded ammonia from a hydrogen-bonded ammonium ion is the absence of bending modes above 1630 cm(-1) and in the region between 1260 and 1600 cm( 1), and a low-frequency bending band in the range from 1130 to 1260 cm(-1). The calculated distribution of vibrational frequencies agrees very well with the measured infrared adsorption spectra. From the comparison of the adsorption data and the vibrational spectra we conclude that due to the complex adsorption geometry the redshift of the asymmetric stretching is a better measure of the acidity of an active sites than the adsorption energy. PMID- 15268052 TI - Relation between pressure shift and electric-field shift of single-molecule lines in a polymer glass. AB - The pressure shifts and the electric-field shifts of individual chromophores in an amporphous matrix are--due to strong disorder--subject to broad distributions. By means of single-molecule spectroscopy we measured both the pressure and the electric-field shift of about 800 tetra-tert-butylterrylene molecules in polyisobutylene. We found a significant correlation of 0.52 (Kendall's correlation coefficient) between the two observables. Analytical calculations and Monte Carlo simulations based on a model by Laird and Skinner predict a nonzero, yet, distinctly smaller correlation. The Monte Carlo simulations showed that the usual assumptions of a spherical shape and isotropic polarizability of the chromophores in glassy systems is an oversimplification of the complex nanoscopic structure and cannot reproduce our experimental results. By taking the molecular anisotropy into account, we obtain agreement of the simulated and the measured correlation between pressure shift and electric-field shift parameter. PMID- 15268053 TI - Molecular-dynamics analysis of the diffusion of molecular hydrogen in all-silica sodalite. AB - In order to investigate the technical feasibility of crystalline porous silicates as hydrogen storage materials, the self-diffusion of molecular hydrogen in all silica sodalite is modeled using large-scale classical molecular-dynamics simulations employing full lattice flexibility. In the temperature range of 700 1200 K, the diffusion coefficient is found to range from 1.610(-10) to 1.810(-9) m(2)/s. The energy barrier for hydrogen diffusion is determined from the simulations allowing the application of transition state theory, which, together with the finding that the pre-exponential factor in the Arrhenius-type equation for the hopping rate is temperature-independent, enables extrapolation of our results to lower temperatures. Estimates based on mass penetration theory calculations indicate a promising hydrogen uptake rate at 573 K. PMID- 15268054 TI - Monte Carlo simulation of dendrimers in variable solvent quality. AB - We study via lattice Monte Carlo simulation and Flory theory the properties of g=1-6 dendrimers in variable solvent quality. For all the generations studied, we find that the radius of gyration R(g) collapses significantly (factor of 2) going from athermal to extreme poor solvent conditions, indicating that varying solvent quality is an effective means of controlling dendrimer size. We also find that in athermal, theta, and extreme poor solvent conditions, the radius of gyration of dendrimers scales with the total number of monomers roughly as R(g) approximately N(1/3). However, a more careful analysis shows that in athermal and theta solvents, there is, in fact, a small but systematic deviation of R(g) from R(g) approximately N(1/3) scaling and the simulation data is described better by the Flory theory prediction of R(g) approximately N(1/5)[(g+1)m](2/5) in athermal solvents and R(g) approximately N(1/4)[(g+1)m](1/4) in theta solvents. We also find for our simulation data that stronger deviations from constant density scaling are possible, with scaling behavior as shallow as R(g) approximately N(0.26) possible for solvent conditions in between theta and the completely collapsed state. It is evident therefore that dendrimers do not obey (or even approximately obey) R(g) approximately N(1/3) scaling under all solvent conditions. Under all solvent conditions, we find that the intramolecular density is dense corelike (i.e., the density maximum is in the interior of the dendrimer) and terminal groups are delocalized throughout the dendrimer. PMID- 15268055 TI - Modification of statistical threading in two-component pseudorotaxane melts using the amphiphilic approach and variations in the confinement geometry. AB - Recently we described a coarse-grained model of poly(ethylene oxide) and then employed that model to study the amount of spontaneous threading of cyclic molecules by linear chains in the melt [C. A. Helfer, G. Xu, W. L. Mattice, and C. Pugh, Macromolecules 36, 10071 (2003)]. Since the amount of statistical threading at equilibrium is small, there is interest in identifying physical changes in the system that will increase the threading. We now use that coarse grained model to investigate the effect on threading of various hypothetical (but feasible) modifications to the two-component system of macrocycles and linear chains in the melt, and different confinement geometries, that can bring about correlations in the arrangement of the rings. Our work follows on the concept of an amphiphilic approach [C. Pugh, J.-Y. Bae, J. R. Scott, and C. L. Wilkins, Macromolecules 30, 8139 (1997)] for increasing the statistical threading in homopolyrotaxane melts. We investigate whether introducing such correlations in the macrocycles can increase the spontaneous threading. This paper shows that some of our modifications can yield more than double the amount of threading seen in purely statistical mixing. PMID- 15268056 TI - The role of attractive interactions in rod-sphere mixtures. AB - We present a computer simulation study of binary mixtures of prolate Gay-Berne particles and Lennard-Jones spheres. Results are presented for three such rod sphere systems which differ from each other only in the interaction between unlike particles. Both the mixing-demixing behavior and the transitions between the isotropic and any liquid crystalline phases are studied for each system, as a function of temperature and concentration ratio. For systems which show macroscopic demixing, the rod-sphere interaction is shown to give direct control over interfacial anchoring properties, giving rise to the possibility of micellar phase formation in the case of homeotropic anchoring. Additionally, it is shown that on incorporating high concentrations of spheres into a system of rods with weak demixing properties, microphase-separated structures can be induced, including bicontinuous and lamellar arrangements. PMID- 15268057 TI - Theoretical study of the longitudinal first hyperpolarizability of polysilaacetylene. AB - With the help of ab initio tools taking into account dynamic electron correlation effects, we study the longitudinal electronic first hyperpolarizability of carbon silicon analogues to polyacetylene. It turns out that the MP2/6-31G(d)//HF/6 31G(d) scheme is suitable to obtain a semiquantitative accuracy for the first hyperpolarizability of long polysilaacetylene oligomers. The conformation of the chain has a crucial impact on its second-order nonlinear optical properties. We also show that, for some chain lengths, the frequency dispersion effects may have a huge impact, even when far away from resonance. These phenomena are rationalized in terms of delocalization and asymmetry. PMID- 15268058 TI - Monte Carlo simulation of the self-assembly and phase behavior of semiflexible equilibrium polymers. AB - Grand canonical Monte Carlo simulations of a simple model semiflexible equilibrium polymer system, consisting of hard sphere monomers reversibly self assembling into chains of arbitrary length, have been performed using a novel sampling method to add or remove multiple monomers during a single MC move. Systems with two different persistence lengths and a range of bond association constants have been studied. We find first-order lyotropic phase transitions between isotropic and nematic phases near the concentrations predicted by a statistical thermodynamic theory, but with significantly narrower coexistence regions. A possible contribution to the discrepancy between theory and simulation is that the length distribution of chains in the nematic phase is bi-exponential, differing from the simple exponential distribution found in the isotropic phase and predicted from a mean-field treatment of the nematic. The additional short length-scale characterizing the distribution appears to arise from the lower orientational order of short chains. The dependence of this length-scale on chemical potential, bond association constant, and total monomer concentration has been examined. PMID- 15268059 TI - Variation of the secondary relaxation strength in a binary glass former. PMID- 15268061 TI - Spin-orbit coupling in O2(v)+O2 collisions: a new energy transfer mechanism. AB - A reduced dimensionality model is used to study the relaxation of highly vibrationally excited O(2)(X (3)Sigma(g) (-),v>/=20) in collisions with O(2)(X (3)Sigma(g) (-),v=0). Spin-orbit coupled potential energy surfaces are employed to incorporate the vibrational-to-electronic energy transfer mechanism involving the O(2)(a (1)Delta(g)) and O(2)(b (1)Sigma(g) (+)) excited states. The transition probabilities obtained show a sharp increase for v>/=26 providing the first direct evidence of the important role played by the electronic energy transfer processes in the depletion of O(2)(X (3)Sigma(g) (-),v>/=26). PMID- 15268062 TI - Basis set effects on frontier molecular orbital energies and energy gaps: a comparative study between plane waves and localized basis functions in molecular systems. AB - In order to study the Kohn-Sham frontier molecular orbital energies in the complete basis limit, a comparative study between localized functions and plane waves, obtained with the local density approximation exchange-correlation functional is made. The analyzed systems are ethylene and butadiene, since they are theoretical and experimentally well characterized. The localized basis sets used are those developed by Dunning. For the plane-waves method, the pseudopotential approximation is employed. The results obtained by the localized basis sets suggest that it is possible to get an estimation of the orbital energies in the limit of the complete basis set, when the basis set size is large. It is shown that the frontier molecular orbital energies and the energy gaps obtained with plane waves are similar to those obtained with a large localized basis set, when the size of the supercell and the plane-wave expansion have been appropriately calibrated. PMID- 15268063 TI - Size versus volume extensivity of a new class of density matrix functionals. AB - Despite being size-extensive, the "second-generation" 1-matrix functionals for the electron-electron repulsion energy V(ee) yield vanishing correlation energy for the homogeneous electron gas. This failure is directly related to the idempotency condition imposed upon an auxiliary matrix that enters the expression for V(ee). In particular, the recently proposed Kollmar-Hess functional is not volume-extensive and thus is incapable of properly describing any delocalized system at its thermodynamic limit. PMID- 15268064 TI - Atomic dipole moments calculated using analytical molecular second-moment gradients. AB - We have implemented analytical second-moment gradients for Hartree-Fock and multiconfigurational self-consistent-field wave functions. The code is used to calculate atomic dipole moments based on the generalized atomic polar tensor (GAPT) formalism [Phys. Rev. Lett. 62, 1469 (1989)], and the proposal of Dinur and Hagler (DH) for the calculation of atomic multipoles [J. Chem. Phys. 91, 2949 (1989)]. Both approaches display smooth basis-set convergence toward a well defined basis-set limit and give reasonable electron correlation effects on the calculated atomic properties. However, the atomic charges and atomic dipole moments obtained from the GAPT partitioning scheme are unable to provide even qualitatively meaningful molecular quadrupole moments for some molecules, and thus the atomic multipole moments calculated in this scheme cannot be considered well suited for analyzing the electron density in molecules and for calculating intermolecular interaction energies. In contrast, the DH approach gives atomic charges and dipole moments that by definition exactly reproduce the molecular quadrupole moments. The approach of DH is, however, restricted to planar molecules and thus suffers from not being applicable to molecules of arbitrary shape. Both the GAPT and DH approaches give rather poor results for octupole and hexadecapole moments, indicating that at least atomic quadrupole moments are required for an accurate representation of the molecular charge distribution in terms of atomic electric moments. PMID- 15268065 TI - An exact reformulation of the diagonalization step in electronic structure calculations as a set of second order nonlinear equations. AB - A new formulation of the diagonalization step in self-consistent-field (SCF) electronic structure calculations is presented. It exactly replaces the diagonalization of the effective Hamiltonian with the solution of a set of second order nonlinear equations. The density matrix and/or the new set of occupied orbitals can be directly obtained from the resulting solution. This formulation may offer interesting possibilities for new approaches to efficient SCF calculations. The working equations can be derived either from energy minimization with respect to a Cayley-type parametrization of a unitary matrix, or from a similarity transformation approach. PMID- 15268066 TI - Geminal model chemistry II. Perturbative corrections. AB - We introduce and investigate a chemical model based on perturbative corrections to the product of singlet-type strongly orthogonal geminals wave function. Two specific points are addressed (i) Overall chemical accuracy of such a model with perturbative corrections at a leading order; (ii) Quality of strong orthogonality approximation of geminals in diverse chemical systems. We use the Epstein-Nesbet form of perturbation theory and show that its known shortcomings disappear when it is used with the reference Hamiltonian based on strongly orthogonal geminals. Application of this model to various chemical systems reveals that strongly orthogonal geminals are well suited for chemical models, with dispersion interactions between the geminals being the dominant effect missing in the reference wave functions. PMID- 15268067 TI - The effect of spin-orbit coupling on fast neutral chemical reaction O(3P)+CH3- >CH3O. AB - The effect of nonadiabatic transitions through the spin-orbit couplings has been investigated on the fast neutral reaction, O((3)P)+CH(3)-->CH(3)O. Adiabatic potential energies and the spin-orbit coupling terms have been evaluated for the four electronic states of CH(3)O ((2)E, (2)A(2), (4)E, and (4)A(2)) that correlate with the O((3)P)+CH(3) asymptote, as a function of CO distance and OCH angle under the C(3v) symmetry, by ab initio electronic structure calculations using multireference internally contracted single and double excitation configuration interaction method with the 6-311G(2df,2pd) basis sets. Multistate quantum reactive scattering calculations have been carried out with the use of thus obtained potential energies and spin-orbit coupling matrices, based on the generalized R-matrix propagation method. The calculated thermal rate constants show a slight positive dependence on temperature in a range between 50 and 2000 K, supporting the previous experimental results. It is shown that the spin-orbit coupled excited states give rise to reflections over the centrifugal barrier due to the quantum interference. Classical capture calculations yield larger rate constants due to the neglect of quantum reflections. It is concluded that the effect of nonadiabatic transitions is of minor importance on the overall reactivity in this reaction. PMID- 15268068 TI - A systematic ab initio study of the equilibrium geometry and vibrational wave numbers of bismuthine. AB - The equilibrium structure and the harmonic and anharmonic force fields of BiH(3) are determined by high-level ab initio calculations using a variety of correlation treatments, basis sets, and pseudopotentials, partly in combination with core polarization potentials. Spin-orbit effects are included by a configuration interaction treatment. This systematic study serves to establish a reliable computational protocol for such calculations and, in particular, to minimize basis set superposition errors through an improved new basis set and/or counterpoise corrections. Using the recommended procedures, the best ab initio results for the equilibrium geometry and the fundamental vibrational wave numbers are in good agreement with the available experimental data, which further supports the recent spectroscopic identification of BiH(3). The ground-state total atomization energy of BiH(3) is predicted to be 153.1 kcal/mol. PMID- 15268069 TI - Geometry and electronic structure of Vn(Bz)m complexes. AB - First-principles calculations based on the generalized gradient approximation to the density functional theory are performed to explore the global geometries, ground-state spin multiplicities, relative stabilities, and energetics of neutral and anionic V(n)(Bz)(m) (n=1-3, m=1-4, with nH+O(2)(nu(")), for the bimolecular exchange process, H+O(2)(nu)-->OH(nu('))+O, and for the dissociative process, H+O(2)(nu)-->H+O+O. The dissociative process appears to be a mere extension of the process of transferring large amounts of energy. State-to-state rate constants are given for the exchange reaction, and they are in reasonable agreement with previous results, while the energy transfer and dissociative rate constants have never been reported previously. The lifetime distributions of the HO(2) complex, calculated as a function of v and temperature, were used as a basis for determining the relative contributions of various vibrational states of O(2) to the thermal rate coefficients for recombination at various pressures. This novel approach, based on the complex's ability to survive until it collides in a secondary process with an inert gas, is used here for the first time. Complete falloff curves for the recombination of H+O(2) are also calculated over a wide range of temperatures and pressures. The combination of the two separate studies results in pressure- and temperature dependent rate constants for H+O(2)(nu)(+Ar) right arrow over left arrow HO(2)(+Ar). It is found that, unlike the exchange reaction, vibrational and rotational-translational energy are liabilities in promoting recombination. PMID- 15268077 TI - Spectroscopic properties of novel aromatic metal clusters: NaM4 (M=Al,Ga,In) and their cations and anions. AB - The ground- and several excited states of metal aromatic clusters, namely NaM(4) and NaM(4) (+/-) (M=Al,Ga,In) clusters have been investigated by employing complete active-space self-consistent-field followed by multireference singles and doubles configuration interaction computations that included up to 10 million configurations and other methods. The ground states NaM(4) (-) of aromatic anions are found to be symmetric C(4nu) ((1)A(1)) electronic states with ideal square pyramid geometries. While the ground state of NaIn(4) is also predicted to be a symmetric C(4nu) ((2)A(1)) square pyramid, the ground state of the NaAl(4) cluster is found to have a C(2nu) ((2)A(1)) pyramid with a rhombus base, and the ground state of NaGa(4) possesses a C(2nu) ((2)A(1)) pyramid with a rectangle base. In general, these structures exhibit two competing geometries, viz., an ideal C(4nu) structure and a distorted rhomboidal or rectangular pyramid structure (C(2nu)). All of the ground states of the NaM(4) (+) (M=Al,Ga,In) cations are computed to be C(2nu) ((3)A(2)) pyramids with rhombus bases. The equilibrium geometries, vibrational frequencies, dissociation energies, adiabatic ionization potentials, adiabatic electron affinities for the electronic states of NaM(4) (M=Al,Ga,In), and their ions are computed and compared with experimental results and other theoretical calculations. On the basis of our computed excited states energy separations, we have tentatively suggested assignments to the observed X and A states in the anion photoelectron spectra of Al(4)Na(-) reported by Li et al. [X. Li, A. E. Kuznetov, H. F. Zheng, A. I. Boldyrev, and L. S. Wang, Science 291, 859 (2001)]. The X state can be assigned to a C(2nu) ((2)A(1)) rhomboidal pyramid. The A state observed in the anion spectrum is assigned to the first excited state ((2)B(1)) of the neutral NaAl(4) with the C(4nu) symmetry. The assignments of the excited states are consistent with the experimental excitation energies and the previous Green's function-based methods for the vertical transition energy separations between the X and A bands. PMID- 15268078 TI - Mass-analyzed threshold ionization spectroscopy of p-methylphenol and p ethylphenol cations and the alkyl substitution effect. AB - The mass-analyzed threshold ionization (MATI) spectra of p-methylphenol and p ethylphenol have been recorded by ionizing via various vibronic levels. The adiabatic ionization energies (IEs) of p-methylphenol and p-ethylphenol are determined to be 65918+/-5 and 65628+/-5 cm(-1), which are less than that of phenol by 2707 and 2997 cm(-1), respectively. This redshift indicates that the interaction between the alkyl group and the ring of alkylphenols in the cationic D(0) state is greater than that in the neutral S(0) state. Moreover, a longer alkyl group gives rise to a greater redshift in the IE. Analysis of the MATI spectra shows that most of the active modes are related to the in-plane ring vibrations of these two cations. However, the length of the alkyl group has an insignificant effect on the frequency of the observed ring vibrations. No band with frequency less than 350 cm(-1) is observed for the p-methylphenol cation. In contrast, many low-frequency bands resulting from the characteristic motions (e.g., the C-C(2)H(5) torsion and C-C(2)H(5) and C-OH bending vibrations) appear in the MATI spectra of p-ethylphenol. The present results show that the ethyl group enhances the substituent-sensitive and many large-amplitude vibrations of the cation. PMID- 15268079 TI - Broadening and line mixing in the 20 (0)0<--01 (1)0, 11 (1)0<--00 (0)0 and 12 (2)0<--01 (1)0 Q branches of carbon dioxide: experimental results and energy corrected sudden modeling. AB - Using both a difference frequency spectrometer and a Fourier transform spectrometer, we have measured transitions in the 12 (2)0<--01 (1)0 band of carbon dioxide at room temperature and pressures up to 19 atm. The low-pressure spectra were analyzed using a variety of standard spectral profiles, all with an asymmetric component to account for weak line mixing. For this band, we have been able to retrieve experimental line strengths and the broadening and weak mixing parameters. In this paper we also compare the suitability of the energy-corrected sudden model to predict mixing in the two previously measured Q branches 20 (0)0< -01 (1)0, the 11 (1)0<--00 (0)0, and the present Q branch of pure CO(2), all at room temperature. PMID- 15268080 TI - Rotational state-dependent mixings between resonance states of vibrationally highly excited DCO (X2A'). AB - Rotational state-dependent mixings between highly excited resonance states of DCO (X (2)A(')) were investigated by stimulated emission pumping spectroscopy via a series of intermediate rotational levels in the B (2)A(') electronic state of the radical. Two examples for such interactions, between pairs of accidentally nearly degenerate vibrational states at energies of E(v) approximately 6450 and E(v) approximately 10 060 cm(-1), respectively, were analyzed in detail. Deperturbations of the measured spectra provided the zeroth-order vibration rotation term energies, widths, and rotational constants of the states and the absolute values of the vibrational coupling matrix elements. The coupled states turned out to have very different A rotational constants so that their mixings switch on or off as they are tuned relative to each other as function of the K(a) rotational quantum number. The respective zeroth-order states could be assigned to different interlaced vibrational polyads. Thus, when two states belonging to different polyads are accidentally nearly isoenergetic, even very weak interpolyad interactions may start to play important roles. The derived interpolyad coupling elements are small compared to the typical intrapolyad coupling terms so that their influences on the vibrational term energies are small. However, large effects on the widths (i.e., decay rates) of the states can be observed even from weak coupling terms when a narrow, long-lived state is perturbed by a broad, highly dissociative state. This influence contributes to the previously observed strong state-to-state fluctuations of the unimolecular decay rates of the DCO radical as function of vibrational excitation. Similar mechanisms are likely to promote the transition to "statistical" rates in many larger molecules. PMID- 15268081 TI - Relative vibrational overtone intensity of cis-cis and trans-perp peroxynitrous acid. AB - The vibrational overtone spectrum of HOONO is examined in the region of the 2 nu(OH) and 3 nu(OH) bands using action spectroscopy in conjunction with ab initio intensity calculations. The present measurements indicate that the oscillator strength associated with the higher energy trans-perp conformer of HOONO is stronger relative to the lower energy cis-cis conformer for both these vibrational overtone levels. Ab initio intensity calculations carried out at the QCISD level of theory suggest that this disparity in oscillator strength apparently arises from differences in the second derivative of the transition dipole moment function of the two isomers. The calculations indicate that the oscillator strength for the trans-perp isomer is approximately 5.4 times larger than that of the cis-cis isomer for the 2 nu(OH) band and approximately 2 times larger for 3 nu(OH) band. The band positions and intensities predicted by the calculations are used to aid in the assignment of features in the experimental action spectra associated with the OH stretching overtones of HOONO. The observed relative intensities in the experimental action spectra when normalized to the calculated oscillator strengths appears to suggest that the concentration of the higher energy trans-perp isomer is comparable to the concentration of the cis-cis isomer in these room temperature experiments. PMID- 15268082 TI - Polar isomer of formic acid dimers formed in helium nanodroplets. AB - The infrared spectrum of formic acid dimers in helium nanodroplets has been observed corresponding to excitation of the "free" OH and CH stretches. The experimental results are consistent with a polar acyclic structure for the dimer. The formation of this structure in helium, as opposed to the much more stable cyclic isomer with two O-H...O hydrogen bonds, is attributed to the unique growth conditions that exist in helium droplets, at a temperature of 0.37 K. Theoretical calculations are also reported to aid in the interpretation of the experimental results. At long range the intermolecular interaction between the two monomers is dominated by the dipole-dipole interaction, which favors the formation of a polar dimer. By following the minimum-energy path, the calculations predict the formation of an acyclic dimer having one O-H...O and one C-H...O contact. This structure corresponds to a local minimum on the potential energy surface and differs significantly from the structure observed in the gas phase. PMID- 15268083 TI - Confirmation of the "long-lived" tetra-nitrogen (N4) molecule using neutralization-reionization mass spectrometry and ab initio calculations. AB - Tetra-nitrogen (N(4)), which has been the subject of recent controversy [Cacace, d. Petris, and Troiani, Science 295, 480 (2002); Cacace, Chem. Eur. J. 8, 3839 (2002); Nguyen et al., J. Phys. Chem. A 107, 5452 (2003); Nguyen, Coord. Chem. Rev. 244, 93 (2003)] as well as of great theoretical interest, has been prepared from the N(4) (+) cation and then detected as a reionized gaseous metastable molecule with a lifetime exceeding 0.8 micros in experiments based on neutralization-reionization mass spectrometry. Moreover, we have used the nature of the charge-transfer reaction which occurs between a beam of fast N(4) (+) ions (8 keV translational energy) and various stationary gas targets to identify the vertical neutralization energy of the N(4) (+) ion. The measured value, 10.3+/ 0.5, most closely matches that of the lowest energy azidonitrene (4)N(4) (+)C(s)((4)A(')) ion, resulting in the formation of the neutral bound azidonitrene (3)N(4)C(s)((3)A(")). Neutralization of the global minimum (2)N(4) (+)D( infinity h)((2)Sigma(u) (+)) ion leads to a structure 166 kJ mol(-1) above the dissociation products [N(2)((1)Sigma(g) (+))+N(2)((1)Sigma(g) (+))]; moreover, it was not possible to find a minimum on the (1)N(4) neutral potential energy surface for a covalently bonded structure. Ab initio calculations at the G3, QCISD/6-31G(d), and MP2/AUG-cc-pVTZ levels of theory have been used to determine geometries and both vertical neutralization energies of ions (doublet and quartet) and ionization energies of neutrals (singlet and triplet). In addition, we have also described in detail the EI ion source for the Ottawa VG ZAB mass spectrometer [Holmes and Mayer, J. Phys. Chem. A 99, 1366 (1995)] which was modified for high-pressure use, i.e., for the production of dimer and higher number cluster ions. PMID- 15268084 TI - Jet-cooled laser spectroscopy of the cyclohexoxy radical. AB - The laser-induced fluorescence and laser-excited dispersed fluorescence spectra of the cyclohexoxy radical has been observed under two sets of free-jet-cooling conditions, characterized by rotational temperatures of approximately 1 and 100 K. Although five conformers of cyclohexoxy are possible, it appears that all presently observed spectral bands can be accounted for by a single one. All cold spectral bands are assigned to the B-X electronic transition of the cyclohexoxy radical. Transitions to both a' and a" B state vibrational levels are observed and allowed due to a substantial pseudo-Jahn-Teller effect in the X state. Hot bands are also observed, which we attributed to transitions to the B state from the low-lying A electronic state. Analysis of the spectra yields vibrational frequencies for the X, A, and B states as well as the energy separations of their vibrationless levels. PMID- 15268085 TI - Analysis of the vibronic fine structure in circularly polarized emission spectra from chiral molecular aggregates. AB - Using a Frenkel-exciton model, the degree of circular polarization of the luminescence (g(lum)) from one-dimensional, helical aggregates of chromophoric molecules is investigated theoretically. The coupling between the electronic excitation and a local, intramolecular vibrational mode is taken into account. Analytical expressions for the fluorescence band shape and g(lum) are presented for the case of strong and weak electronic coupling between the chromophoric units. Results are compared to those from numerical calculations obtained using the three particle approximation. g(lum) for the 0-0 vibronic band is found to be independent of the relative strength of electronic coupling between chromophores and excitation-vibration coupling. It depends solely on the number of coherently coupled molecules. In contrast, for the higher vibronic transitions[g(lum)] decreases with decreasing strength of the electronic coupling. In the limit of strong electronic coupling, [g(lum)] is almost constant throughout the series of vibronic transitions but for weak coupling [g(lum)] becomes vanishingly small for all vibronic transitions except for the 0-0 transition. The results are interpreted in terms of dynamic localization of the excitation during the zero point vibrational motion in the excited state of the aggregate. It is concluded that circular polarization measurements provide an independent way to determine the coherence size and bandwidth of the lowest exciton state for chiral aggregates. PMID- 15268086 TI - Heat capacity effects associated with the hydrophobic hydration and interaction of simple solutes: a detailed structural and energetical analysis based on molecular dynamics simulations. AB - We examine the SPCE [H. J. C. Berendsen et al., J. Chem. Phys. 91, 6269 (1987)] and TIP5P [M. W. Mahoney and W. L. Jorgensen, J. Chem. Phys 112, 8910 (2000)] water models using a temperature series of molecular-dynamics simulations in order to study heat-capacity effects associated with the hydrophobic hydration and interaction of xenon particles. The temperature interval between 275 and 375 K along the 0.1-MPa isobar is studied. For all investigated models and state points we calculate the excess chemical potential for xenon employing the Widom particle insertion technique. The solvation enthalpy and excess heat capacity is obtained from the temperature dependence of the chemical potentials and, alternatively, directly by Ewald summation, as well as a reaction field based method. All three methods provide consistent results. In addition, the reaction field technique allows a separation of the solvation enthalpy into solute/solvent and solvent/solvent parts. We find that the solvent/solvent contribution to the excess heat capacity is dominating, being about one order of magnitude larger than the solute/solvent part. This observation is attributed to the enlarged heat capacity of the water molecules in the hydration shell. A detailed spatial analysis of the heat capacity of the water molecules around a pair of xenon particles at different separations reveals that even more enhanced heat capacity of the water located in the bisector plane between two adjacent xenon atoms is responsible for the maximum of the heat capacity found for the desolvation barrier distance, recently reported by Shimizu and Chan [J. Am. Chem. Soc. 123, 2083 (2001)]. The about 60% enlarged heat capacity of water in the concave part of the joint xenon-xenon hydration shell is the result of a counterplay of strengthened hydrogen bonds and an enhanced breaking of hydrogen bonds with increasing temperature. Differences between the two models with respect to the heat capacity in the xenon-xenon contact state are attributed to the different water model bulk heat capacities, and to the different spatial extension of the structure effect introduced by the hydrophobic particles. Similarities between the different states of water in the joint xenon-xenon hydration shell and the properties of stretched water are discussed. PMID- 15268087 TI - Polymerization of nitrogen in sodium azide. AB - The high-pressure behavior of nitrogen in NaN(3) was studied to 160 GPa at 120 3300 K using Raman spectroscopy, electrical conductivity, laser heating, and shear deformation methods. Nitrogen in sodium azide is in a molecularlike form; azide ions N(3-) are straight chains of three atoms linked with covalent bonds and weakly interact with each other. By application of high pressures we strongly increased interaction between ions. We found that at pressures above 19 GPa a new phase appeared, indicating a strong coupling between the azide ions. Another transformation occurs at about 50 GPa, accompanied by the appearance of new Raman peaks and a darkening of the sample. With increasing pressure, the sample becomes completely opaque above 120 GPa, and the azide molecular vibron disappears, evidencing completion of the transformation to a nonmolecular nitrogen state with amorphouslike structure which crystallizes after laser heating up to 3300 K. Laser heating and the application of shear stress accelerates the transformation and causes the transformations to occur at lower pressures. These changes can be interpreted in terms of a transformation of the azide ions to larger nitrogen clusters and then polymeric nitrogen net. The polymeric forms can be preserved on decompression in the diamond anvil cell but transform back to the starting azide and other new phases under ambient conditions. PMID- 15268088 TI - Centroid molecular dynamics approach to the transport properties of liquid para hydrogen over the wide temperature range. AB - Fundamental transport properties of liquid para-hydrogen (p-H(2)), i.e., diffusion coefficients, thermal conductivity, shear viscosity, and bulk viscosity, have been evaluated by means of the path integral centroid molecular dynamics (CMD) calculations. These transport properties have been obtained over the wide temperature range, 14-32 K. Calculated values of the diffusion coefficients and the shear viscosity are in good agreement with the experimental values at all the investigated temperatures. Although a relatively large deviation is found for the thermal conductivity, the calculated values are less than three times the amount of the experimental values at any temperature. On the other hand, the classical molecular dynamics has led all the transport properties to much larger deviation. For the bulk viscosity of liquid p-H(2), which was never known from experiments, the present CMD has given a clear temperature dependence. In addition, from the comparison based on the principle of corresponding states, it has been shown that the marked deviation of the transport properties of liquid p-H(2) from the feature which is expected from the molecular parameters is due to the quantum effect. PMID- 15268089 TI - Density functional theory of fluids in the isothermal-isobaric ensemble. AB - We present a density functional theory for inhomogeneous fluids at constant external pressure. The theory is formulated for a volume-dependent density, n(r,V), defined as the conjugate variable of a generalized external potential, nu(r,V), that conveys the information on the pressure. An exact expression for the isothermal-isobaric free-energy density functional is obtained in terms of the corresponding canonical ensemble functional. As an application we consider a hard-sphere system in a spherical pore with fluctuating radius. In general we obtain very good agreement with simulation. However, in some situations a peak develops in the center of the cavity and the agreement between theory and simulation becomes worse. This happens for systems where the number of particles is close to the magic numbers N=13, 55, and 147. PMID- 15268090 TI - Adam-Gibbs model for the supercooled dynamics in the ortho-terphenyl ortho phenylphenol mixture. AB - Dielectric measurements of the alpha-relaxation time were carried out on a mixture of ortho-terphenyl (OTP) with ortho-phenylphenol, over a range of temperatures at two pressures, 0.1 and 28.8 MPa. These are the same conditions for which heat capacity, thermal expansivity, and compressibility measurements were reported by Takahara et al. [S. Takahara, M. Ishikawa, O. Yamamuro, and T. Matsuo, J. Phys. Chem. B 103, 3288 (1999)] for the same mixture. From the combined dynamic and thermodynamic data, we determine that density and temperature govern to an equivalent degree the variation of the relaxation times with temperature. Over the measured range, the dependence of the relaxation times on configurational entropy is in accord with the Adam-Gibbs model, and this dependence is invariant to pressure. Consistent with the implied connection between relaxation and thermodynamic properties, the kinetic and thermodynamic fragilities are found to have the same pressure independence. In comparing the relaxation properties of the mixture to those of neat OTP, density effects are stronger in the former, perhaps suggestive of less efficient packing. PMID- 15268091 TI - A semiclassical generalized quantum master equation for an arbitrary system-bath coupling. AB - The Nakajima-Zwanzig generalized quantum master equation (GQME) provides a general, and formally exact, prescription for simulating the reduced dynamics of a quantum system coupled to a, possibly anharmonic, quantum bath. In this equation, a memory kernel superoperator accounts for the influence of the bath on the dynamics of the system. In a previous paper [Q. Shi and E. Geva, J. Chem. Phys. 119, 12045 (2003)] we proposed a new approach to calculating the memory kernel, in the case of arbitrary system-bath coupling. Within this approach, the memory kernel is obtained by solving a set of two integral equations, which requires a new type of two-time system-dependent bath correlation functions as input. In the present paper, we consider the application of the linearized semiclassical (LSC) approximation for calculating those correlation functions, and subsequently the memory kernel. The new approach is tested on a benchmark spin-boson model. Application of the LSC approximation for calculating the relatively short-lived memory kernel, followed by a numerically exact solution of the GQME, is found to provide an accurate description of the relaxation dynamics. The success of the proposed LSC-GQME methodology is contrasted with the failure of both the direct application of the LSC approximation and the weak coupling treatment to provide an accurate description of the dynamics, for the same model, except at very short times. The feasibility of the new methodology to anharmonic systems is also demonstrated in the case of a two level system coupled to a chain of Lennard-Jones atoms. PMID- 15268092 TI - Intermolecular double-quantum coherences in two-dimensional spectra of binary mixtures in solution. The role of diffusion. AB - Proton NMR two-dimensional 2-D spectra of binary mixtures, obtained with the correlation spectroscopy revamped by asymmetric Z gradient echo detection pulse sequence, were employed to test various assumptions usually adopted to describe the role of diffusion in intermolecular double quantum coherences. When two molecular species, with significantly different diffusivities, are considered, the relative amplitudes of the peaks, and their widths, furnish a stringent test that unveils some inadequacies in standard approximations. PMID- 15268093 TI - Landscapes and fragilities. AB - The concept of fragility provides a possibility to rank different supercooled liquids on the basis of the temperature dependence of dynamic and/or thermodynamic quantities. We recall here the definitions of kinetic and thermodynamic fragility proposed in the last years and discuss their interrelations. At the same time we analyze some recently introduced models for the statistical properties of the potential energy landscape. Building on the Adam-Gibbs relation, which connects structural relaxation times to configurational entropy, we analyze the relation between statistical properties of the landscape and fragility. We call attention to the fact that the knowledge of number, energy depth, and shape of the basins of the potential energy landscape may not be sufficient for predicting fragility. Finally, we discuss two different possibilities for generating strong behavior. PMID- 15268094 TI - Computer simulation study of the closure relations in hard sphere fluids. AB - We study, using Monte Carlo simulations, the cavity and the bridge functions of various hard sphere fluids: one component system, equimolar additive, and nonadditive binary mixtures. In particular, we numerically check the assumption of local dependency of the bridge functions from the indirect correlation functions, on which most of the existing integral equation theories hinge. We find that this condition can be violated either in the region around the first and second neighbors shell, or inside the hard core, for the systems here considered. The violations manifest themselves clearly in the so-called Duh Haymet plots of the bridge functions versus the indirect correlation functions and become amplified as the coupling of the system increases. PMID- 15268095 TI - A vibrational spectroscopic study of structure evolution of water dissolved in supercritical carbon dioxide under isobaric heating. AB - A combination of Raman scattering spectroscopy and infrared absorption was applied to investigate the structural evolution of water dissolved in supercritical carbon dioxide under isobaric heating (T=40-340 degrees C, P=250 bar). Quantitative analysis of experimental spectra allowed us to determine that at relatively moderate temperatures water dissolved in CO(2)-rich phase exists only under monomeric form (solitary water surrounding by CO(2) molecules), but hydrogen-bonded species, namely, dimers, begin to appear upon heating. At the same time, the ratio of dimers to monomers concentration increases with further temperature increase and at temperatures close to the temperature of total miscibility of the mixture (T=366 degrees C, P=250 bar), water dimers only are present in the CO(2)-rich phase. PMID- 15268096 TI - Bridging continuum and statistical thermodynamics via equations of state and the density of states. AB - The connection between molecular force fields and equations of state (EoS) is typically established at the level of predicted quantities, e.g., by comparing simulation data and EoS data. In this paper we show how an EoS can be used to extract the density of states (Omega) of a system thus establishing a deeper connection between EoSs and statistical thermodynamics. We also show how such an EoS Omega can be used to aid molecular simulation methods designed to map out Omega (like the multicanonical approach). Central to the implementation of these ideas is the fact that the configurational Omega is related to thermodynamic properties accessible by an EoS via Boltzmann's equation. Sample calculations are presented for the Omega relevant to isothermal-isobaric and grand canonical ensemble simulations using the hard-sphere system and the Lennard-Jones system as model fluids, and the Carnahan-Starling EoS and a cubic EoS, respectively, as thermodynamic models. PMID- 15268097 TI - Thermodynamic forces in highly curved fluid interfaces. AB - The identification of the force distribution in curved interfaces as a thermodynamic force [Baus and Lovett, J. Chem. Phys. 101, 377 (1995)] can be interpreted as a relation between the force distribution and the grand canonical free energy difference between two distinct systems. Using this interpretation, molecular expressions are developed for the force distribution in cylindrical and spherical interfaces that remain valid for very highly curved interfaces. PMID- 15268098 TI - Density inhomogeneity and diffusion behavior of fluids in micropores by molecular dynamics simulation. AB - The density profiles and the diffusion behavior of fluid argon confined in micropores were studied by molecular-dynamics simulations. The effects of pore size (width), temperature and number density on the density profiles and the self diffusion coefficients in micropores were simulated with pore widths from 0.6 to 4.0 nm. The density profiles are greatly affected by the pore size. Strong inhomogeneities in the channel direction and vapor-liquid phase separation in the micropores were observed when initial conditions were chosen in the coexistence region of the fluid. The self-diffusion coefficient in the channel direction in the pores was found to be much lower than in the bulk, and decreasing with decreasing pore size, decreasing temperature, and increasing density. PMID- 15268099 TI - New theory of equation of state for surface monolayer. AB - A novel statistical-thermodynamic approach to deriving an equation of state for a surface monolayer has been elaborated on the basis of excluded area. A master differential equation relating surface (two-dimensional) pressure to excluded area has been derived to generate equations of state for a surface monolayer. The crudest solution (the zero approximation) of the master equation reproduces the known van Laar and Frumkin equations of state. The first approximation yields the two-dimensional van der Waals equation. The second, third, and fourth approximations lead to new and more accurate equations of state. The particular result of the fourth approximation is a precise equation of state for hard disks with deviation not more than 0.46% from data obtained by Monte Carlo and molecular-dynamics simulations within the whole range of surface density. The role of the third dimension for surface equations of state is discussed. An orientation equation of state has been proposed for monolayers containing anisometric particles. It follows from the orientation equation obtained that the orientation effect creates possibility for a two-dimensional phase transition. PMID- 15268100 TI - Wavelength dependence of the hyper Rayleigh scattering response from gold nanoparticles. AB - The wavelength dependence of the quadratic hyperpolarizability of 11 nm diam gold nanoparticles, is reported as measured by hyper Rayleigh scattering. An important photoluminescence background underlying the hyper Rayleigh signal is observed, a contribution attributed to radiative electron-hole recombinations following multiphoton excitation favored by adsorbed organic compound like citrate on the surface of the nanoparticles. The absolute value of the quadratic hyperpolarizability of gold spherical nanoparticles is determined and a strong enhancement is observed for harmonic frequencies in resonance with the dipolar surface plasmon excitation. No contribution of the interband transition is observed. The absolute values reported, beta(C)=5.1x10(-26) esu at the second harmonic energy 2.39 eV, have been measured with femtosecond long laser pulse, and are 1 order of magnitude weaker that the one previously reported with nanosecond long pulses. This difference can be related to similar measurements performed on the second order hyperpolarizability of gold nanoparticles and may be attributed to different electronic relaxation regimes. Finally, the spectrum of the quadratic hyperpolarizability is compared to the theoretically expected one. PMID- 15268101 TI - Simulation study of angle-resolved photoemission spectra and intramolecular energy-band dispersion of a poly(tetrafluoroethylene) oligomer film. AB - Theoretical simulations of the angle-resolved ultraviolet photoemission spectra (ARUPS) for the oligomer of poly(tetrafluoroethylene) [(CF(2))(n); PTFE] were performed using the independent-atomic-center approximation combined with ab initio molecular orbital calculations. Previously observed normal-emission spectra for the end-on oriented sample (with long-chain axis perpendicular to the surface) showed the incident photon-energy (hnu) dependence due to the intramolecular energy-band dispersion along the one-dimensional chain, and the present simulations successfully reproduced this hnu dependence of the observed spectra. We employed the experimentally observed helical structure for PTFE oligomers for the simulations. We also calculated the density of states (DOS) for the planar zigzag structure, and examined the changes in the electronic structure due to the difference in the molecular structure by comparing the DOS for the helical and planar zigzag structures. Only a small change in the DOS was found between these structures, showing little change of the electronic structure between these conformations. We also evaluated the inner potential V(0), which is the parameter defining the energy origin of the free-electron-like final state, and checked the validity of the value of -10 eV estimated in our previous study using the experimentally observed hnu dependence of the peak intensity. The estimation of V(0) was performed by pursuing the best agreement between the energy-band dispersion [E=E(k)] relation along the chain direction obtained from the simulated spectra and the experimentally deduced one. An excellent agreement in the topmost band was achieved when the assumed inner potential V(0) was set at about zero. This value of V(0) is much different from the value of V(0)=-10 eV in the previous study, suggesting the invalidity of the previous assumption at the estimation of V(0) from the peak intensity variation with hnu. Using the presently obtained V(0), we could derive more reliable E=E(k) dispersion relation from the observed ARUPS spectra. The comparison of this newly derived relation gave good agreement with theoretically calculated E=E(k) relations, in contrast to the poor agreement for the previous results with V(0)=-10 eV. PMID- 15268102 TI - Coarse-grained nonequilibrium approach to the molecular modeling of permeation through microporous membranes. AB - We present a modeling technique that combines a statistical-mechanical coarse graining scheme with a nonequilibrium molecular simulation algorithm to provide an efficient simulation of steady-state permeation across a microporous material. The coarse-graining scheme is based on the mapping of an atomistic model to a lattice using multidimensional free-energy and transition-state calculations. The nonequilibrium simulation algorithm is a stochastic, lattice version of the recently developed atomistic dual-control-volume grand canonical molecular dynamics. We demonstrate the approach on a model of methane permeating through a bulk portion of siliceous zeolite ZK4 at 300 K under imposed fugacity differences. We predict the coarse-grained (cage-level) density profiles and observe the development of nonlinearities as the magnitude of the fugacity difference is increased. From the net flux of methane we also predict a mean permeability coefficient under the various conditions. The simulation results are obtained over time scales on the order of microseconds and length scales on the order of dozens of nanometers. PMID- 15268103 TI - Hamiltonian theory for vibrational line shapes of atoms adsorbed on surfaces. AB - The vibrational motions of atomic adsorbates on surfaces can be probed by helium atom scattering. The experimental observable is the dynamic structure factor, which shows an inelastic peak around the vibrational frequency of the isolated adsorbates known as the frustrated translational or T-mode peak. In this paper we develop a theory for the line shape of this peak, as well as for its temperature dependent shift and broadening, based on a Hamiltonian equivalent of the generalized Langevin equation. The theory can be used to infer physical parameters of the adatom-surface interaction, such as the friction coefficient, the barrier height to diffusion, and the anharmonicity parameter. Numerical simulations are used to ascertain the range of validity of the theory, which is also generalized to describe multidimensional systems and to include quantum corrections. We compare the theoretical predictions for the shift and broadening with experimental results for the Na/Cu(001) system, showing quantitative agreement within experimental resolution. PMID- 15268104 TI - Application of the exact exchange potential method for half metallic intermediate band alloy semiconductor. AB - In this paper we present an analysis of the convergence of the band structure properties, particularly the influence on the modification of the bandgap and bandwidth values in half metallic compounds by the use of the exact exchange formalism. This formalism for general solids has been implemented using a localized basis set of numerical functions to represent the exchange density. The implementation has been carried out using a code which uses a linear combination of confined numerical pseudoatomic functions to represent the Kohn-Sham orbitals. The application of this exact exchange scheme to a half-metallic semiconductor compound, in particular to Ga(4)P(3)Ti, a promising material in the field of high efficiency solar cells, confirms the existence of the isolated intermediate band in this compound. PMID- 15268105 TI - Imaging water on Ag(111): field induced reorientation and contrast inversion. AB - Water adsorbed on Ag(111) at 70 K forms circular clusters that consist of six molecules. In scanning tunneling microscopy, this cyclic hexamer is imaged as a protrusion for voltages below V(SS)=-93 meV and as a depression for voltages above V(SS). The electronic density of states, however, increases around V(SS). We explain this counterintuitive result with the aid of calculated images by a change from constructive to destructive interference between different tunneling channels due to a field induced reorientation of the molecule under the tunneling tip. PMID- 15268106 TI - Dipole-induced structure in aromatic-terminated self-assembled monolayers: a study by near edge x-ray absorption fine structure spectroscopy. AB - The structure of self-assembled monolayers presenting aromatic rings at a surface is studied by near edge x-ray absorption fine structure spectroscopy (NEXAFS). Fluorine substitution at asymmetric positions in the aromatic rings is used to generate a layer of dipoles at the surface of the monolayer. We find that fluorine substituted aromatic rings are more ordered than unsubstituted aromatic rings by a factor of two based on the polarization dependence of the lowest C 1s to pi* transition, which is associated with transitions from phenyl carbons attached to hydrogens. This result is consistent with the influence of dipole dipole interactions and quadrupolar interactions between the aromatic groups due to the substitution of fluorine atoms. The work also serves to illustrate how subtle variations in the orientation of an end group of a self-assembled monolayer can be determined by using NEXAFS. PMID- 15268107 TI - Trends in the band structures of the group-I and -II oxides. AB - Measured and calculated band structures for the six lightest group-I and -II oxides are reported. Band structures have been measured using electron momentum spectroscopy, a technique that maps the ground-state occupied orbitals resolved both in energy and momentum. Measurements are compared with first-principles calculations carried out within the linear combination of atomic orbitals approximation using both Hartree-Fock (HF) and density functional (DFT) methods. Three DFT functionals are used representative of the local density approximation, the generalized gradient approximation, and a hybrid method incorporating exact exchange. The calculated O 2p bandwidths and O 2p-2s band gaps generally scale linearly with the inverse of the oxygen-oxygen separation squared, but consistently show an anomaly at Li(2)O. These trends, including the anomaly, are also observed in the experimental data. HF calculations consistently overestimate the oxygen 2p-2s band gap by almost a factor of two. Measured band gaps lie within the range of the three DFT functionals employed, with evidence that the description of exchange becomes more important as the cation size increases. Both HF and DFT calculations overestimate the oxygen valence bandwidths, with DFT giving more accurate predictions. Both observed and calculated bandwidths converge as the cation size increases, indicating that exchange-correlation effects become less important as the metallic ion becomes larger. PMID- 15268108 TI - Optical properties of passivated silicon nanoclusters: the role of synthesis. AB - The effect of preparation conditions on the structural and optical properties of silicon nanoparticles is investigated. Nanoscale reconstructions, unique to curved nanosurfaces, are presented for silicon nanocrystals and shown to have lower energy and larger optical gaps than bulk-derived structures. We find that high-temperature synthesis processes can produce metastable noncrystalline nanostructures with different core structures than bulk-derived crystalline clusters. The type of core structure that forms from a given synthesis process may depend on the passivation mechanism and time scale. The effect of oxygen on the optical of different types of silicon structures is calculated. In contrast to the behavior of bulklike nanostructures, for noncrystalline and reconstructed crystalline structures surface oxygen atoms do not decrease the gap. In some cases, the presence of oxygen atoms at the nanocluster surface can significantly increase the optical absorption gap, due to decreased angular distortion of the silicon bonds. The relationship between strain and the optical gap in silicon nanoclusters is discussed. PMID- 15268109 TI - Density-induced coupling effects on the dispersivity of a flexible chain particle. AB - A model is introduced to investigate the transport properties of an inhomogeneously dense flexible chain particle. The specific model used is a sedimenting non-neutrally buoyant inhomogenously weighted flexible Brownian dumbbell, and it is shown that density inhomogeneity gives rise to a novel coupling effect between the "shape-fluctuation" and "size-fluctuation" dispersion mechanisms. The previously reported shape-fluctuation dispersion term stems from the dumbbell's nonspherical shape and the ensuing anisotropic mobility tensor, while the already investigated size fluctuation term is the result of the dependence of the overall dumbbell translational mobility on the separation distance between the constitutive spheres. Because the density of the constitutive spheres is unequal, the external force simultaneously reorients and deforms the flexible dumbbell, and it is this mutual dependence between dumbbell orientation and size that induces the coupling. Numerical results are presented for the case of a tethered dumbbell composed of two spheres, identical in size but differing in density. The "weak-field" limit is addressed, where the externally applied torque and particle deformation forces are dominated by the thermal fluctuations associated with rotational and deformation Brownian motion. This numerical solution, obtained by including a large number of higher order hydrodynamic interactions (120 terms), describes the Brownian particle's long time transport without resorting to ad hoc approximations, such as preaveraging the hydrodynamic force or incorporating only first-order hydrodynamic interaction effects (such as employing the Burgers-Oseen tensor). Separate analytical solutions, based on these respective approximations, are also presented and it is concluded that in the limit of "long tethers," where the ratio of tether length to sphere size is greater than seven, no more than 15% error is introduced by neglecting higher-order hydrodynamic interactions. Similarly, the preaveraging approximation introduces no more than a few percent error in the limit of "almost rigid" dumbbells, where the ratio of tether length to sphere size is less than three. For tethers of "intermediate" length, the full numerical solution must be employed. PMID- 15268110 TI - Orientational and interaction induced dynamics in the isotropic phase of a liquid crystal: polarization resolved ultrafast optical Kerr effect spectroscopy. AB - The ultrafast dynamics of the isotropic phase of a liquid crystal 4'-pentyl-4-p biphenylcarbonitrile (5CB) have been investigated using polarization resolved optical Kerr effect spectroscopy. Measurements were made as a function of both temperature and dilution in nonpolar solvents. To separate single molecule and interaction induced components to the relaxation of the induced birefringence, measurements of both the anisotropic and isotropic response were made. The isotropic response was found to be dominated by a damped low-frequency mode of intramolecular origin. There is a minor additional component assigned to an interaction induced contribution. There is at most an extremely weak isotropic signal beyond 1 ps, showing that the picosecond time scale dynamics of 5CB are dominated by orientational relaxation. The isotropic response is independent of temperature in the range studied (0.2-50 K above the nematic to isotropic phase transition temperature). The anisotropic response exhibits relaxation dynamics on time scales spanning subpicosecond to several hundred picoseconds and beyond. The fastest components are dominated by a librational response, but there are smaller contributions from three low-frequency intramolecular modes, and a contribution from interaction induced effects. The low-frequency spectral density extracted from these data are independent of temperature in the range studied, 0.2-30 K above the phase-transition temperature, but shift to lower frequency on dilution in alkane solvents. In neat 5CB the picosecond time scale orientational dynamics are dominated by temperature-independent reorientation within the pseudonematic domains, while in solution these are disrupted, and the orientational response becomes faster and temperature dependent. PMID- 15268111 TI - Prediction of membrane protein structures by replica-exchange Monte Carlo simulations: case of two helices. AB - We test our prediction method of membrane protein structures with glycophorin A transmembrane dimer and analyze the predicted structures in detail. Our method consists of two parts. In the first part, we obtain the amino-acid sequences of the transmembrane helix regions from one of existing WWW servers and use them as an input for the second part of our method. In the second part, we perform a replica-exchange Monte Carlo simulation of these transmembrane helices with some constraints that indirectly represent surrounding lipid and water effects and identify the predicted structure as the global-minimum-energy state. The structure obtained in the case for the dielectric constant epsilon=1.0 is very close to that from the nuclear magnetic resonance experiments, while that for epsilon=4.0 is more packed than the native one. Our results imply that the helix helix interaction is the main driving force for the native structure formation and that the stability of the native structure is determined by the balance of the electrostatic term, van der Waals term, and torsion term, and the contribution of electrostatic energy is indeed important for correct predictions. The inclusion of atomistic details of side chains is essential for estimating this balance accurately because helices are tightly packed. PMID- 15268112 TI - Nematic ordering in dilute solutions of rodlike polyelectrolytes. AB - Quantitative theory of orientational behavior of rodlike polyelectrolytes in dilute solution is developed. We find that in salt-free solutions many-body Coulomb interactions between macro- and counterions favor nematic ordering. It is shown that the orientationally isotropic phase of the solution becomes unstable toward nematic ordering at polymer concentration smaller than the overlap concentration. Our predictions are consistent with experimental observations for synthetic polyelectrolytes poly(p-phenylene)sulfonates in aqueous solutions. PMID- 15268115 TI - Three-dimensional orientational colocalization of individual donor--acceptor pairs. AB - We report on the determination of the three-dimensional orientation of the donor and acceptor transition dipoles in individual fluorescence resonance energy transfer (FRET) pairs by means of scanning optical microscopy with annular illumination. Knowledge of the mutual orientation of the donor and acceptor dipole is mandatory for reliable distance determination based on FRET efficiency measurements. In our model system perylenediimide as the donor and terryelenediimide as the acceptor are coupled via a stiff p-terphenyl linker. The absorption dipoles of the donor and acceptor are selectively addressed by the 488 nm and 647 line of an Ar/Kr mixed gas laser, respectively. A clear deviation from collinearity is observed with a distribution of misalignment angles peaked around 22 degrees. PMID- 15268116 TI - Silver nanoparticle array structures that produce remarkably narrow plasmon lineshapes. AB - Using electrodynamics calculations, we have discovered one dimensional array structures built from spherical silver nanoparticles that produce remarkably narrow ( approximately meV or less) plasmon resonance spectra upon irradiation with light that is polarized perpendicular to the array axis. The narrow lines require a minimum particle radius of about 30 nm to achieve. Variations of the plasmon resonance wavelength, extinction efficiency and width with particle size, array structure, interparticle distance and polarization direction are examined, and conditions which lead to the smallest widths are demonstrated. A simple analytical expression valid for infinite lattices shows that the sharp resonance arises from cancellation between the single particle width and the imaginary part of the radiative dipolar interaction. PMID- 15268117 TI - Single-ensemble nonequilibrium path-sampling estimates of free energy differences. AB - We introduce a straightforward, single-ensemble, path sampling approach to calculate free energy differences based on Jarzynski's relation. For a two dimensional "toy" test system, the new (minimally optimized) method performs roughly one hundred times faster than either optimized "traditional" Jarzynski calculations or conventional thermodynamic integration. The simplicity of the underlying formalism suggests the approach will find broad applicability in molecular systems. PMID- 15268118 TI - Computing time scales from reaction coordinates by milestoning. AB - An algorithm is presented to compute time scales of complex processes following predetermined milestones along a reaction coordinate. A non-Markovian hopping mechanism is assumed and constructed from underlying microscopic dynamics. General analytical analysis, a pedagogical example, and numerical solutions of the non-Markovian model are presented. No assumption is made in the theoretical derivation on the type of microscopic dynamics along the reaction coordinate. However, the detailed calculations are for Brownian dynamics in which the velocities are uncorrelated in time (but spatial memory remains). PMID- 15268119 TI - A hybrid scheme for the resolution-of-the-identity approximation in second-order Moller-Plesset linear-r(12) perturbation theory. AB - In the framework of second-order Moller-Plesset linear-r(12) (MP2-R12) perturbation theory, a method is developed and implemented that uses an auxiliary basis set for the resolution-of-the-identity (RI) approximation for the three- and four-electron integrals. In contrast to previous work, the two-electron integrals that must be evaluated never involve more than one auxiliary basis function. The new method therefore scales linearly with the number of auxiliary basis functions and is much more efficient than the previous one, which scaled quadratically. A general formulation of MP2-R12 theory is presented for various ansatze, approximations, and orbitals (canonical or localized). The new method is assessed by computations of the valence-shell second-order Moller-Plesset correlation energy of a few small closed-shell systems. The preliminary calculations indicate that the difference between the new and previous methods is about one order of magnitude smaller than the errors that occur due to basis-set truncations and RI approximations and under the assumptions of generalized and extended Brillouin conditions. PMID- 15268120 TI - Effective force fields for condensed phase systems from ab initio molecular dynamics simulation: a new method for force-matching. AB - A novel least-squares fitting approach is presented to obtain classical force fields from trajectory and force databases produced by ab initio (e.g., Car Parrinello) molecular dynamics (MD) simulations. The method was applied to derive effective nonpolarizable three-site force fields for liquid water at ambient conditions from Car-Parrinello MD simulations in the Becke-Lee-Yang-Parr approximation to the electronic density functional theory. The force-matching procedure includes a fit of short-ranged nonbonded forces, bonded forces, and atomic partial charges. The various parameterizations of the water force field differ by an enforced smooth cut-off applied to the short-ranged interaction term. These were obtained by fitting to the trajectory and force data produced by Car-Parrinello MD simulations of systems of 32 and 64 H(2)O molecules. The new water force fields were developed assuming both flexible or rigid molecular geometry. The simulated structural and self-diffusion properties of liquid water using the fitted force fields are in close agreement with those observed in the underlying Car-Parrinello MD simulations. The resulting empirical models compare to experiment much better than many conventional simple point charge (SPC) models. The fitted potential is also shown to combine well with more sophisticated intramolecular potentials. Importantly, the computational cost of the new models is comparable to that for SPC-like potentials. PMID- 15268121 TI - The hardness profile as a tool to detect spurious stationary points in the potential energy surface. AB - In the present work, we have computed the energy and hardness profiles for a series of inter and intramolecular conformational changes at several levels of calculation. All processes studied have in common the fact that the choice of a weak methodology or a poor basis set results in the presence of spurious stationary points in the energy profile. At variance with the energy profiles, the hardness profiles calculated as the difference between the vertical ionization potential and electron affinity always show the correct number of stationary points independently of the basis set and methodology used. For this reason, we have concluded that hardness profiles can be used to check the reliability of the energy profiles for those chemical systems that, because of their size, cannot be treated with high level ab initio methods. PMID- 15268122 TI - Calculation of free energy through successive umbrella sampling. AB - We consider an implementation of umbrella sampling in which the pertinent range of states is subdivided into small windows that are sampled consecutively and linked together. This allows us to simulate without a weight function or to extrapolate the results to the neighboring window in order to estimate a weight function. Additionally, we present a detailed error analysis in which we demonstrate that the error in umbrella sampling is controlled and, in the absence of sampling difficulties, independent of the window sizes. In this case, the efficiency of our implementation is comparable to a multicanonical simulation with a very good weight function, which in our scheme does not need to be known ahead of time. The analysis also allows us to detect sampling difficulties such as correlations between adjacent windows and provides a test of equilibration. We exemplify the scheme by simulating the liquid-vapor coexistence in a Lennard Jones system. PMID- 15268123 TI - Optimized Jastrow-Slater wave functions for ground and excited states: application to the lowest states of ethene. AB - A quantum Monte Carlo method is presented for determining multideterminantal Jastrow-Slater wave functions for which the energy is stationary with respect to the simultaneous optimization of orbitals and configuration interaction coefficients. The approach is within the framework of the so-called energy fluctuation potential method which minimizes the energy in an iterative fashion based on Monte Carlo sampling and a fitting of the local energy fluctuations. The optimization of the orbitals is combined with the optimization of the configuration interaction coefficients through the use of additional single excitations to a set of external orbitals. A new set of orbitals is then obtained from the natural orbitals of this enlarged configuration interaction expansion. For excited states, the approach is extended to treat the average of several states within the same irreducible representation of the pointgroup of the molecule. The relationship of our optimization method with the stochastic reconfiguration technique by Sorella et al. is examined. Finally, the performance of our approach is illustrated with the lowest states of ethene, in particular with the difficult case of the 1(1)B(1u) state. PMID- 15268124 TI - Calculation of the A term of magnetic circular dichroism based on time dependent density functional theory I. Formulation and implementation. AB - A procedure for calculating the A term and the A/D ratio of magnetic circular dichroism (MCD) within time-dependent density functional theory (TD-DFT) is described. Utilizing an implementation of the MCD theory within the Amsterdam Density Functional program, the A term contributions to the MCD spectra of MnO(4) (-), CrO(4) (2-), VO(4) (3-), MoO(4) (2-), VO(4) (3-), MoS(4) (2-), Se(4) (2+), Te(4) (2+), Fe(CN)(6) (4-), Ni(CN)(4) (2-), trichlorobenzene, hexachlorobenzene, tribromobenzene, and hexabromobenzene are calculated. For the most part, agreement between theory and experiment for A/D ratios and the relative magnitude of A terms is found to be good, leading to simulated spectra that are similar in appearance to those derived from measurements. The A terms are found to be too small whenever comparison with experiment was possible, probably due to the neglect of environment effects on the incident radiation and the relative low accuracy of dipole strengths calculated within TD-DFT. PMID- 15268125 TI - Implementing quantum gates on oriented optical isomers. AB - Optical enantiomers are proposed to encode molecular two-qubit information processing. Using sequences of pairs of nonresonant optimally polarized pulses, different schemes to implement quantum gates, and to prepare entangled states, are described. We discuss the role of the entanglement phase and the robustness of the pulse sequences which depend on the area theorem. Finally, possible scenarios to generalize the schemes to n-qubit systems are suggested. PMID- 15268126 TI - A Bohmian total potential view to quantum effects. I. Methodology and simple model systems. AB - The coherent-state wave packet dynamics of several model systems is analyzed in terms of Bohm's total potential. The quantum dynamics has been obtained by solving the time-dependent Schrodinger equation, and a method for obtaining the total potential from it, involving just matrix algebra, has been proposed. Contrary to what one may expect, it is shown that the time- and state-dependent features of the total potential admit a rationale, classical-like description of quantum effects, leading to a unified picture of them, which is not critically dependent, as for the key features, on the classical potential. An outstanding feature is found to be the relation of the state system's density amplitude and sharpness (in its dependence with position) with quantum effects. Sharp density profiles and low densities cause the total potential to strongly depart from the classical value, in both time regimes and position ranges, which provide a clearer, more deterministic view to quantum dynamics. Free motion as well as scattering processes by square and Eckart barriers have been analyzed by means of careful inspection of several time dependent snapshots. The result is an insightful picture of processes involving tunneling and antitunneling, including their dynamical variants, as well as resonances and quantization. PMID- 15268127 TI - On the valence shell electronic spectroscopy of 2-vinyl furan. AB - The vacuum ultraviolet (VUV) absorption spectrum (3.50-10.33 eV, 350-120 nm) of gaseous 2-vinyl furan has been measured for the first time using both synchrotron radiation source and electron energy loss spectroscopies with absolute cross section determinations. The He I photoelectron spectrum obtained at higher resolution than previously has been interpreted with the aid of semiempirical molecular orbital calculations. Three excited states of type (1)pipi(*) are found responsible for an intense and structured first band observed between 4.2 and 5.8 eV (295-214 nm). Three triplet states were detected for the first time at about 2.46, 3.35, and 3.8 eV (477, 370, and 328 nm) which are, from the calculations, assigned as (3)pipi(*). Some partial Rydberg series, linked to IE(1) and IE(2) are identified. The VUV absorption spectrum bears little resemblance to that of the parent compound, furan. The electronically excited molecule is found akin to a linear polyene. PMID- 15268128 TI - Dynamics of photodissociation of ethylene and its isotopomers at 157 nm: branching ratios and kinetic-energy distributions. AB - We investigated the photodissociation of ethylene and its isotopomers at 157 nm in a molecular-beam apparatus using photofragment translational spectroscopy combined with synchrotron-based photoionization. The time-of-flight (TOF) spectra of all photofragments H, H(2), C(2)H(2), C(2)H(3), and their deuterium isotopic variants were recorded, from which kinetic-energy distributions P(E(t)) and branching ratios were obtained. Most C(2)H(3) spontaneously dissociates to C(2)H(2)+H and only C(2)H(3) with small internal energy survives. The C(2)H(2) fragment due to H(2) elimination is observed leading the C(2)H(2) fragment due to 2H elimination in TOF distribution because the former process has more kinetic energy release. An analogous result is observed for C(2)D(4) photolysis. That elimination of molecular hydrogen is site-specific and is revealed from photolysis of three dideuterated ethylene isotopomers, in which an isotopic effect plays a significant role. Observations of C(2)D(2)+2H and C(2)H(2)+2D product channels in the photolysis of 1,1-CH(2)CD(2) provide evidence for migrations of H and D atoms. A comparison with previous experimental and theoretical results is made. PMID- 15268129 TI - Dynamics of photodissociation of 3,3,3-d(3)-propene at 157 nm: site effect and hydrogen migration. AB - In a preceding paper [Lee et al., J. Chem. Phys. 119, 827 (2003)], we measured the kinetic-energy distributions P(E(t)) and branching ratios of products from photolysis of propene at 157 nm using time-of-flight spectroscopy combined with photoionization. In the present work, hydrogen migration before fragmentation and a site effect on P(E(t)) and branching ratios were revealed from the photodissociation of CD(3)CHCH(2). Labeling of the methyl group with deuterium enabled us to differentiate between elimination of atomic and molecular hydrogen from the vinyl moiety and from the methyl moiety; the P(E(t)) and relative yields for the formation of H, D, H(2), HD, and D(2) were measured. Deuterium labeling allowed us to also differentiate the fragmentation after hydrogen transfer from that before hydrogen migration. The observation of isotopic variants of CD(3) and C(2)H(3) radicals in the C-C bond cleavage provides evidence for hydrogen transfer of propene because of site specificity. The fraction of fragmentation after hydrogen transfer is estimated to be 25%. The isotope-specific branching ratios for five dissociation pathways of CD(3)CHCH(2) were evaluated. PMID- 15268130 TI - Photoinduced dynamics of ethene in the N, V, and Z valence states: a six dimensional nonadiabatic quantum dynamics investigation. AB - The photoinduced dynamics of ethene following pi-->pi(*) excitation is investigated by quantum wave-packet dynamics on three coupled six-dimensional diabatic potential-energy surfaces representing the N, V, and Z valence states, which have been developed previously. The C-C stretching and torsion, as well as the pyramidalization and scissoring of both CH(2) groups are included in this description. The wave-packet calculations have been performed using the multiconfigurational time-dependent Hartree method for a time period up to 100 fs. While a small amount of population transfer to the electronic ground state is found within this period, the overall population decay time of the V state is found to exceed the 100 fs range significantly. The autocorrelation function of the wave packet and the stationary absorption spectrum of the V state also have been calculated. It is found that both the torsional mode as well as the C-C stretching mode contribute to the very extended vibrational structure of the absorption spectrum, and that both modes are strongly coupled. At least on the present ab initio surface of limited dimensionality, the speed of pyramidalization of 90 degrees twisted ethene appears as the bottleneck for the ultrafast radiationless decay of the V state. PMID- 15268131 TI - Combined perturbative-variational investigation of the vibrations of CHBr(3) and CDBr(3). AB - A full dimensional vibrational treatment of CHBr(3) and CDBr(3) using Van Vleck perturbation theory followed by a variational calculation is presented. The calculation of a force field, and its adjustment for better match with experiment, is discussed. The computed eigenstates and spectral features are compared to experiment. Changes in intensities of the nu(1) and 2nu(4) bands upon simple alterations of the dipole moment expansion are described. PMID- 15268132 TI - Above the surface multifragmentation of surface scattered fullerenes. AB - C(60) (-) ions were scattered from a gold surface at impact energies of 80-900 eV. The C(n) (-) fragments abundance distribution (odd and even) and the sharp fragmentation threshold observed, point at a prompt shattering event. The measured angle and energy distributions of the C(n) (-) fragments (n=2-12) provide clear evidence for a multifragmentation process where the superheated fullerenes leave the surface "intact" and disintegrate away from the surface. PMID- 15268133 TI - A high-resolution pulsed field ionization-photoelectron-photoion coincidence study of vinyl bromide. AB - By employing the high-resolution pulsed field ionization-photoelectron (PFI-PE) photoion coincidence method, we have examined the unimolecular dissociation reaction of energy-selected C(2)H(3)Br(+) to form C(2)H(3) (+)+Br near its threshold. The analysis of the breakdown curves for C(2)H(3)Br(+) and C(2)H(3) (+) yields a value of 11.9010+/-0.0015 eV for the 0 K dissociative photoionization threshold or appearance energy (AE) for C(2)H(3) (+) from C(2)H(3)Br. This AE(C(2)H(3) (+)) value, together with the ionization energy (IE) for C(2)H(3)Br (9.8200+/-0.0015 eV) obtained by PFI-PE and threshold photoelectron (TPE) measurements, has allowed the determination of the 0 K dissociation energy (D(0)) for the C(2)H(3) (+)-Br bond to be 2.081+/-0.002 eV. The 0 K AE(C(2)H(3) (+)) from C(2)H(3)Br obtained in this study corresponds to DeltaH(f0) ( composite function )(C(2)H(3) (+))=1123.7+/-1.9 kJ/mol. Combining the latter value and the known DeltaH(f0) ( composite function )(C(2)H(3))=306.7+/-2.1 kJ/mol, we calculated a value of 8.468+/-0.029 eV for the IE(C(2)H(3)), which is in accord with the result obtained in the previous photoionization efficiency study. We have also carried out high-level ab initio calculations for the IE(C(2)H(3)) at the Gaussian-3 and the CCSD(T,full)/CBS level of theory. The CCSD(T,full)/CBS prediction of 8.487 eV for the IE(C(2)H(3)- >bridged-C(2)H(3) (+)) is in good agreement with the IE(C(2)H(3)) value derived in the present experiment. Combining the 0 K AE(C(2)H(3) (+))=11.9010+/-0.0015 eV and the IE(C(2)H(3))=8.468+/-0.029 eV yields the value of 3.433+/-0.029 eV for D(0)(C(2)H(3)-Br). We have also recorded the TPE spectrum of C(2)H(3)Br in the energy range of 9.80-12.20 eV. Members (n=5-14) of four autoionizing Rydberg series converging to the C(2)H(3)Br(+)(A (2)A(')) state are observed in the TPE spectrum. The analysis of the converging limit of these Rydberg series and the vibrational TPE bands for C(2)H(3)Br(+)(A (2)A(')) has provided more precise values for the nu(6) (+) (1217+/-10 cm(-1)) and nu(8) (+) (478+/-8 cm(-1)) modes and the IE (10.9156+/-0.0010 eV) for the formation of C(2)H(3)Br(+)(A (2)A(')) from C(2)H(3)Br. PMID- 15268134 TI - Photodissociation and multiphoton dissociative ionization processes in CH(3)S(2)CH(3) at 193 nm studied using velocity-map imaging. AB - Dissociation and ionization processes in dimethyl disulfide, CH(3)S(2)CH(3), induced by one- or two-photon absorption of 193 nm light, have been studied using velocity-map ion imaging. The analysis of the ion images of the CH(3)S(2) (+), CH(3)S(+), S(2) (+), and S(+) fragments has allowed the characterization of the scattering dynamics of some of the main photolysis and dissociative-ionization processes. In particular, the experiments corroborate the formation of electronically excited SCH(3)((2)A(1)) products in the 193 nm photodissociation of dimethyl disulfide seen in earlier studies, and show that laser ionization provides a very sensitive method for their detection. The data have also allowed determination of the recoil energy and angular distributions of the CH(3)S(2) (+) and CH(3)S(+) products of the two-photon dissociative-ionization of the CH(3)S(2)CH(3) molecule. The measured distributions for these products are consistent with the formation of a transient parent ion which dissociates after a substantial intramolecular rearrangement, possibly yielding the most stable isomeric forms of the fragments, namely CH(2)S(2)H(+) and CH(2)SH(+). PMID- 15268135 TI - Infrared line collisional parameters of HCl in argon, beyond the impact approximation: measurements and classical path calculations. AB - Measurement of room temperature absorption by HCl-Ar mixtures in the 1-0 and 2-0 bands have been made for pressures between 10 and 50 atm. Fits of these spectra are made for the determination of the width, spectral shift, asymmetry, and intensity of individual lines. The broadening and shifting parameters are in satisfactory agreement with previous determinations but provide the first complete and self-consistent sets covering P(15)-R(14) and P(7)-R(8) in the 1-0 and 2-0 bands, respectively. The asymmetries of the profiles, which have been studied for the first time, are smaller than typically 10(-3) atm(-1) and cannot be determined experimentally. On the other hand, the intensities of the low j lines show a significant linear decrease with increasing Ar pressure. Calculations of all measured quantities are made with a classical path approach and an accurate vibrational-dependent HCl-Ar potential energy surface (PES). Comparisons with experimental values show that widths and shifts are well predicted, confirming the quality of the PES and of the theoretical model, and the calculations confirm that asymmetries are small. The damping factors of the intensities are analyzed by considering three contributions: The first is due to the formation of van der Waals complexes, the second results from the finite duration of collisions, and the last comes from initial correlations. Calculations indicate that the last process has negligible consequences but that the first two processes lead to effects of the same order and explain most of the observed decrease of the intensities, even if some discrepancies persist for the mid R:mmid R:=1 rotational components. PMID- 15268136 TI - Collision-energy-resolved Penning ionization electron spectroscopy of p benzoquinone: study of electronic structure and anisotropic interaction with He(*)(2 (3)S) metastable atoms. AB - Collision energy dependence of partial ionization cross sections (CEDPICS) of p benzoquinone with He(*)(2 (3)S) metastable atoms indicates that interaction potentials between p-benzoquinone and He(*)(2 (3)S) are highly anisotropic in the studied collision energy range (100-250 meV). Attractive interactions were found around the C==O groups for in-plane and out-of-plane directions, while repulsive interactions were found around CH bonds and the benzenoid ring. Assignment of the first four ionic states of p-benzoquinone and an analogous methyl-substituted compound was examined with CEDPICS and anisotropic distributions of the corresponding two nonbonding oxygen orbitals (n(O) (+),n(O) (-)) and two pi(CC) orbitals (pi(CC) (+),pi(CC) (-)). An extra band that shows negative CEDPICS was observed at ca. 7.2 eV in Penning ionization electron spectrum. PMID- 15268137 TI - Ab initio molecular orbital study of structures and energetics of Si(3)H(2), Si(3)H(2) (+), and Si(3)H(2) (-). AB - The geometric structures, isomeric stabilities, and potential energy profiles of various isomers and transition states in Si(3)H(2) neutral, cation and anion are investigated at the coupled-cluster singles, doubles (triples) level of theory. For the geometrical survey, the basis sets used are of the Dunning's correlation consistent basis sets of triple-zeta quality (cc-pVTZ) for the neutral and cation and the Dunning's correlation consistent basis sets of double-zeta quality with diffuse functions (aug-cc-pVDZ) for the anion. For the final energy calculations, the aug-cc-pVTZ: Dunning's correlation consistent basis sets of triple-zeta quality with diffuse functions and cc-pVQZ: Dunning's correlation consistent basis sets of quadruple-zeta quality basis sets are used for the neutral and the aug-cc-pVTZ ones for the cation and anion. The global minimum neutral (I-1: (1)A(1)) has the same framework as that (cyclopropenylidene) of the C(3)H(2) molecule. Other low-lying three isomers (I-2, I-3, and I-4) are also predicted to be within 20 kJ/mol. Five transition states are optimized and their energy relationships with the isomers are clarified. The geometric structure of the global minimum cation (C-1: (2)A(1)) has the same framework as that of the neutral, but that of the anion (A-1: (2)A(')) differs very much from those of the neutral and cation. The calculated vertical and adiabatic ionization potentials from the global minimum neutral (I-1) are 7.85 and 7.77 eV, respectively. The adiabatic electron affinity of the neutral I-1 and the electron detachment energy of the global minimum anion (A-1) are predicted to be 1.21 and 1.92 eV, respectively. The two-electron three-centered bond is widely observed in the present Si(3)H(2) neutral, cation, and anion. The contour plots of their localized molecular orbitals clearly show the existence of such nonclassical chemical bonds. PMID- 15268138 TI - Exponential probe rotation in glass-forming liquids. AB - Using time resolved optical depolarization, we have studied the rotational behavior of molecular probes in supercooled liquids near the glass transition temperature T(g). Simultaneously, the dynamics of the liquid immediately surrounding these rigid probes is measured by triplet state solvation experiments. This direct comparison of solute and solvent dynamics is particularly suited for assessing the origin of exponential orientational correlation functions of probe molecules embedded in liquids which exhibit highly nonexponential structural relaxation. Polarization angle dependent Stokes shift correlation functions demonstrate that probe rotation time and solvent response time are locally correlated quantities in the case of smaller probe molecules. Varying the size of both guest and host molecules shows that the size ratio determines the rotational behavior of the probes. The results are indicative of time averaging being at the origin of exponential rotation of probes whose rotational time constant is slower than solvent relaxation by a factor of 20 or more. PMID- 15268139 TI - New results for phase transitions from catastrophe theory. AB - Catastrophe theory predicts that in certain limits universal relations should exist between barrier heights, curvatures and the positions of local maxima and minima on a potential or free energy surface. In the present work we investigate these relations for both first- and second-order phase transitions, revealing that the ideal ratios often hold quite well over a wide range of conditions. This elementary catastrophe theory is illustrated for the melting transition of an atomic cluster, the isotropic-to-nematic transition in a liquid crystal, and the ferromagnetic-to-paramagnetic phase transition in the two-dimensional Ising model. PMID- 15268140 TI - Excess entropy scaling for the diffusion coefficient in expanded liquid metals. AB - Molecular-dynamics simulation is used to compute the pair correlation function and the velocity autocorrelation function of Cs and Rb along the liquid-vapor coexistence curve, from which the excess entropy S(ex) and the diffusion coefficient D are deduced. The numerical results of both physical properties are correlated and a scaling law between the excess entropy and the reduced diffusion coefficient D(*)(=D/D(0)) is investigated for different expressions of the reduction parameter D(0). The choice of thermodynamic states along the liquid- vapor coexistence curve gives us the possibility to extend the investigation of the relation between the reduced diffusion coefficient and the excess entropy over a wide area and to test the adequacy of the scaling law confidently. PMID- 15268141 TI - Isotope effects associated with tunneling and double proton transfer in the hydrogen bonds of benzoic acid. AB - The isotope effects associated with double proton transfer in the hydrogen bonds of benzoic acid (BA) dimers have been measured using field-cycling (1)H NMR relaxometry and quasielastic neutron scattering. By studying mixed isotope (hydrogen and deuterium) samples, the dynamics of three isotopologues, BA-HH, BA HD, and BA-DD, have been investigated. Low temperature measurements provide accurate measurements of the incoherent tunneling rate, k(0). This parameter scales accurately with the mass number, m, according to the formula k(0)=(E/m)e( Fm) providing conclusive evidence that the proton transfer process is a strongly correlated motion of two hydrons. Furthermore, we conclude that the tunneling pathway is the same for the three isotopologue species. Measurements at higher temperatures illuminate the through barrier processes that are mediated via intermediate or excited vibrational states. In parallel with the investigation of proton transfer dynamics, the theoretical and experimental aspects of studying spin-lattice relaxation in single crystals of mixed isotope samples are investigated in depth. Heteronuclear dipolar interactions between (1)H and (2)H isotopes contribute significantly to the overall proton spin-lattice relaxation and it is shown that these must be modeled correctly to obtain accurate values for the proton transfer rates. Since the sample used in the NMR measurements was a single crystal, full account of the orientation dependence of the spin-lattice relaxation with respect to the applied B field was incorporated into the data analysis. PMID- 15268142 TI - Femtosecond primary events in bacteriorhodopsin and its retinal modified analogs: revision of commonly accepted interpretation of electronic spectra of transient intermediates in the bacteriorhodopsin photocycle. AB - Femtosecond primary events in bacteriorhodopsin (BR) and its retinal modified analogs are discussed. Ultrafast time resolved electronic spectra of the primary intermediates induced in the BR photocycle are discussed along with spectral and kinetic inconsistencies of the previous models proposed in the literature. The theoretical model proposed in this paper based on vibrational coupling between the electronic transition of the chromophore and intramolecular vibrational modes allows us to calculate the equilibrium electronic absorption band shape and the hole burning profiles. The model is able to rationalize the complex pattern of behavior for the primary events in BR and explain the origin of the apparent inconsistencies between the experiment and the previous theoretical models. The model presented in the paper is based on the anharmonic coupling assumption in the adiabatic approximation using the canonical transformation method for diagonalization of the vibrational Hamiltonian instead of the commonly used perturbation theory. The electronic transition occurs between the Born Oppenheimer potential energy surfaces with the electron involved in the transition being coupled to the intramolecular vibrational modes of the molecule (chromophore). The relaxation of the excited state occurs by indirect damping (dephasing) mechanisms. The indirect dephasing is governed by the time evolution of the anharmonic coupling constant driven by the resonance energy exchange between the intramolecular vibrational mode and the bath. The coupling with the intramolecular vibrational modes results in the Franck-Condon progression of bands that are broadened due to the vibrational dephasing mechanisms. The electronic absorption line shape has been calculated based on the linear response theory whereas the third order nonlinear response functions have been used to analyze the hole burning profiles obtained from the pump-probe time-resolved measurements. The theoretical treatment proposed in this paper provides a basis for a substantial revision of the commonly accepted interpretation of the primary events in the BR photocycle that exists in the literature. PMID- 15268143 TI - An application of flexible constraints in Monte Carlo simulations of the isobaric -isothermal ensemble of liquid water and ice Ih with the polarizable and flexible mobile charge densities in harmonic oscillators model. AB - The method of flexible constraints was implemented in a Monte Carlo code to perform numerical simulations of liquid water and ice Ih in the constant number of molecules, volume, and temperature and constant pressure, instead of volume ensembles, using the polarizable and flexible mobile charge densities in harmonic oscillators (MCDHO) model. The structural and energetic results for the liquid at T=298 K and rho=997 kg m(-3) were in good agreement with those obtained from molecular dynamics. The density obtained at P=1 atm with flexible constraints, rho=1008 kg m(-3), was slightly lower than with the classical sampling of the intramolecular vibrations, rho=1010 kg m(-3). The comparison of the structures and energies found for water hexamers and for ice Ih with six standard empirical models to those obtained with MCDHO, show this latter to perform better in describing water far from ambient conditions: the MCDHO minimum lattice energy, density, and lattice constants were in good agreement with experiment. The average angle HOH of the water molecule in ice was predicted to be slightly larger than in the liquid, yet 1.2% smaller than the experimental value. PMID- 15268144 TI - Photon-stimulated desorption of F(-) ions from CF(3)Cl adsorbed on Si(111)-7x7. AB - We report the photon-stimulated desorption of negative ions induced by direct dipolar dissociation and dissociative electron attachment. The photon-stimulated desorption of F(-) ions from CF(3)Cl physisorbed on a Si(111)-7x7 surface at 30 K in the photon energy range 12-35 eV was studied. The F(-) ion yield exhibits four resonances, at 12.8, 16.2, 19.5, and 22.3 eV, quite unlike the gas phase photodissociation cross section. The intensities of these resonances depend strongly on the CF(3)Cl coverage in a manner which varies from peak to peak. The resonances at 19.5 and 22.3 eV, which have a significant enhancement in the monolayer regime, are due to electron mediated dipolar dissociation of adsorbed CF(3)Cl molecules. The enhancement is attributed to surface electron attachment following molecular excitation. A significant enhancement in the monolayer regime has also been observed for the resonances at 12.8 and 16.2 eV. These two resonances are ascribable to a combination of electron mediated dipolar dissociation and dissociative electron attachment driven by photoelectrons generated in the neighboring molecules. PMID- 15268145 TI - Irreversible adsorption of particles at random-site surfaces. AB - Irreversible adsorption of negatively charged polystyrene latex particles (averaged diameter 0.9 microm) at heterogeneous surfaces was studied experimentally. The substrate bearing a controlled number of adsorption sites was produced by precovering mica sheets by positively charged polystyrene latex (averaged diameter of 0.45 microm). Positive latex (site) deposition was carried out under diffusion-controlled transport conditions and its coverage was determined by direct particle counting using the optical microscopy. Deposition kinetics of larger latex particles (averaged diameter 0.9 microm) at heterogeneous surfaces produced in this way was studied by direct optical microscope observations in the diffusion cell (under no-convection transport conditions). It was demonstrated that the structure of larger particle monolayers, characterized in terms of the pair correlation function, showed much more short-range ordering than it was predicted for homogeneous surface monolayers at the same coverage. This was found in agreement with theoretical predictions derived from the Monte Carlo simulations. On the other hand, particle adsorption kinetics was quantitatively interpreted in terms of numerical solutions of the governing diffusion equation with the nonlinear boundary condition derived from Monte Carlo simulations. From these kinetic measurements maximum (jamming) coverage of particles was determined in an accurate way by extrapolation. It was concluded that both the monolayer structure and jamming coverage were strongly influenced by the site multiplicity (coordination) effect. PMID- 15268146 TI - Solvent-induced microphase separation in diblock copolymer thin films with reversibly switchable morphology. AB - We have studied the surface morphology of symmetric poly(styrene)-block poly(methyl methacrylate) diblock copolymer thin films after solvent vapor treatment selective for poly(methyl methacrylate). Highly ordered nanoscale depressions or striped morphologies are obtained by varying the solvent annealing time. The resulting nanostructured films turn out to be sensitive to the surrounding medium, that is, their morphologies and surface properties can be reversibly switchable upon exposure to different block-selective solvents. PMID- 15268147 TI - Oscillatory wetting instability induced by liquid-liquid decomposition in a Ga- Pb alloy. AB - We present the first experimental investigation and pertinent theoretical modeling of an interfacial oscillatory instability in a binary fluid alloy, the Ga-Pb system. It is characterized by spinodal decomposition at elevated temperatures and by a complete wetting transition at liquid-liquid coexistence. For the alloy Ga(0.95)Pb(0.05) the fluid interface has been probed by second harmonic generation (SHG) under UHV conditions at temperatures between 740 and 550 K. At conditions inside the miscibility gap clear oscillations of the SHG intensity with a period of approximately 30 min are found for different cooling cycles and also at constant temperatures. These interfacial oscillatory instabilities simultaneously induce temperature oscillations in the bulk fluid with the same period. This phenomenon can be explained by a periodic variation of the fluid interfacial emissivity. A model has been developed which describes the wetting-dewetting dynamics by hydrodynamic equations within the Reynolds approximation. It is found that the interfacial oscillatory instability is determined by capillary-gravitation instability. The model quantitatively describes the time evolution of the interfacial and temperature oscillations and gives the correct value of the oscillation period. A detailed comparison of the experimental and model results is given. PMID- 15268148 TI - Excitation migration in trimeric cyanobacterial photosystem I. AB - A structure-based description of excitation migration in multireaction center light harvesting systems is introduced. The description is an extension of the sojourn expansion, which decomposes excitation migration in terms of repeated detrapping and recapture events. The approach is applied to light harvesting in the trimeric form of cyanobacterial photosystem I (PSI). Excitation is found to be shared between PSI monomers and the chlorophylls providing the strongest respective links are identified. Excitation sharing is investigated by computing cross-monomer excitation trapping probabilities. It is seen that on the average there is a nearly 40% chance of excitation cross transfer and trapping, indicating efficient coupling between monomers. The robustness and optimality of the chlorophyll network of trimeric PSI is examined. PMID- 15268149 TI - Determination of low-pressure crystalline--liquid phase boundary of SnI(4). AB - The location of the liquidus in the low-pressure crystalline phase of SnI(4) was determined utilizing in situ x-ray diffraction measurements under pressures up to approximately 3.5 GPa. The liquidus is not well fitted to a monotonically increasing curve such as Simon's equation, but breaks near 1.5 GPa and then becomes almost flat. The results are compared to those from molecular dynamics simulations. Ways to improve the model potential adopted in the simulations are discussed. PMID- 15268150 TI - Monte Carlo simulation for the formation of a mixed crystal from two solids in contact. AB - The study focuses on nucleation and growth of a binary mixed crystal phase from two pure crystals in contact. Monte Carlo simulations of this process are conducted, with the dynamics proceeding via activated atom-vacancy exchanges. Intermolecular interactions, ranging up to next-nearest neighbors, are of size typical of hydrogen bonded systems. The process is driven by the formation of strong AB bonds at the expense of weaker AA and BB bonds. In the resulting model, the material is channeled and transported through the mixed phase crust along antiphase boundaries. The flow of molecules through the channels is directed, due to molecular energy lowering via gradual acquisition of an increasing number of nearest neighbors of the second species. On the other hand, defect motion is quasirandom. The model accounts partially for the t(1/alpha) (alpha>3) time dependence observed for conversion of nanoparticles of HBr dihydrate to monohydrate, by exposure to acid adsorbate. PMID- 15268151 TI - Electron-nuclear correlations for photo-induced dynamics in molecular dimers. AB - Ultrafast photoinduced dynamics of electronic excitation in molecular dimers is drastically affected by the dynamic reorganization of inter- and intra- molecular nuclear configuration modeled by a quantized nuclear degree of freedom. The dynamics of the electronic population and nuclear coherence is analyzed by solving the chain of coupled differential equations for population inversion, electron-vibrational correlation, etc. Intriguing results are obtained in the approximation of a small change of the nuclear equilibrium upon photoexcitation. In the limiting case of resonance between the electronic energy gap and the frequency of the nuclear mode these results are justified by comparison to the exactly solvable Jaynes-Cummings model. It is found that the photoinduced processes in the model dimer are arranged according to their time scales: (i) Fast scale of nuclear motion, (ii) intermediate scale of dynamical redistribution of electronic population between excited states as well as growth and dynamics of electron-nuclear correlation, (iii) slow scale of electronic population approach to the quasi-equilibrium distribution, decay of electron-nuclear correlation, and decrease of the amplitude of mean coordinate oscillation. The latter processes are accompanied by a noticeable growth of the nuclear coordinate dispersion associated with the overall nuclear wave packet width. The demonstrated quantum relaxation features of the photoinduced vibronic dynamics in molecular dimers are obtained by a simple method, applicable to systems with many degrees of freedom. PMID- 15268152 TI - Interfacial properties of the nanostructured dye-sensitized solid heterojunction TiO(2)/RuL(2)(NCS)(2)/CuI. AB - The interfaces of the nanostructured dye-sensitized solid heterojunction TiO(2)/Ru-dye/CuI have been studied using photoelectron spectroscopy of core and valence levels, x-ray absorption spectroscopy and atomic force microscopy. A nanostructured anatase TiO(2) film sensitized with RuL(2)(NCS)(2) [cis-bis(4,4(') dicarboxy-2,2(')-bipyridine)-bis(isothio-cyanato)-ruthenium(II)] was prepared in a controlled way using a novel combined in-situ and ex-situ (Ar atmosphere) method. Onto this film CuI was deposited in-situ. The formation of the dye-CuI interface and the changes brought upon the dye-TiO(2) interface could be monitored in a stepwise fashion. A direct interaction between the dye NCS groups and the CuI is evident in the core level photoelectron spectra. Concerning the energy matching of the valence electronic levels, the photoelectron spectra indicate that the dye HOMO overlaps in energy with the Cu 3d-I 5p hydrid states. The CuI grow in the form of particles, which at the initial stages displace the dye molecules causing dye-TiO(2) bond breaking. Consequently, the very efficient charge injection channel provided by the dye-TiO(2) carboxylic bonding is directly affected for a substantial part of the dye molecules. This may be of importance for the functional properties of such a heterojunction. PMID- 15268153 TI - Glassy phases in random heteropolymers with correlated sequences. AB - We develop an analytic approach for the study of lattice heteropolymers and apply it to copolymers with correlated Markovian sequences. According to our analysis, heteropolymers present three different dense phases depending upon the temperature, the nature of the monomer interactions, and the sequence correlations: (i) a liquid phase, (ii) a "soft glass" phase, and (iii) a "frozen glass" phase. The presence of the intermediate "soft glass" phase is predicted, for instance, in the case of polyampholytes with sequences that favor the alternation of monomers. Our approach is based on the cavity method, a refined Bethe-Peierls approximation adapted to frustrated systems. It amounts to a mean field treatment in which the nearest-neighbor correlations, which are crucial in the dense phases of heteropolymers, are handled exactly. This approach is powerful and versatile; it can be improved systematically and generalized to other polymeric systems. PMID- 15268154 TI - In search of temporal power laws in the orientational relaxation near isotropic nematic phase transition in model nematogens. AB - Recent Kerr relaxation experiments by Gottke et al. have revealed the existence of a pronounced temporal power law decay in the orientational relaxation near the isotropic-nematic phase transition (INPT) of nematogens of rather small aspect ratio, kappa (kappa approximately 3-4). We have carried out very long (50 ns) molecular dynamics simulations of model (Gay-Berne) prolate ellipsoids with aspect ratio 3 in order to investigate the origin of this power law. The model chosen is known to undergo an isotropic to nematic phase transition for a range of density and temperature. The distance dependence of the calculated angular pair correlation function correctly shows the emergence of a long range correlation as the INPT is approached along the density axis. In the vicinity of INPT, the single particle second rank orientational time correlation function exhibits power law decay, (t(-alpha)) with exponent alpha approximately 2/3. More importantly, we find the sudden appearance of a pronounced power-law decay in the collective part of the second rank orientational time correlation function at short times when the density is very close to the transition density. The power law has an exponent close to unity, that is, the correlation function decays almost linearly with time. At long times, the decay is exponential-like, as predicted by Landau-de Gennes mean field theory. Since Kerr relaxation experiments measure the time derivative of the collective second rank orientational pair correlation function, the simulations recover the near independence of the signal on time observed in experiments. In order to capture the microscopic essence of the dynamics of pseudonematic domains inside the isotropic phase, we introduce and calculate a dynamic orientational pair correlation function (DOPCF) obtained from the coefficients in the expansion of the distinct part of orientational van Hove time correlation function in terms of spherical harmonics. The DOPCF exhibits power law relaxation when the pair separation length is below certain critical length. The orientational relaxation of a local director, defined in terms of the sum of unit vectors of all the ellipsoidal molecules, is also found to show slow power law relaxation over a long time scale. These results have been interpreted in terms of a newly developed mode coupling theory of orientational dynamics near the INPT. In the present case, the difference between the single particle and the collective orientational relaxation is huge which can be explained by the frequency dependence of the memory kernel, calculated from the mode coupling theory. The relationship of this power law with the one observed in a supercooled liquid near its glass transition temperature is explored. PMID- 15268155 TI - Curvature elasticity of mixed amphiphilic bilayers. AB - Elastic bending constants of mixed amphiphilic bilayers are calculated using a molecular approach. The free energy is expanded up to quadratic order in curvatures and compositions, choosing a flat symmetrical bilayer as the reference state. Bending constants are then calculated from the derivatives of the free energy evaluated at this reference state. Two-component bilayers are considered. As a novelty, the local compositions are allowed to fully relax upon bending so that the 2 monolayers are at chemical equilibrium with each other at every curvature. The compositional degree of freedom is shown to affect the bending constant k, but not the saddle-splay constant k. The influence on the membrane elastic properties of various chain structural features, such as length, volume, and stiffness, is investigated. This may prove useful to model mixed bilayers composed of hydrocarbon/hydrocarbon and hydrocarbon/fluorocarbon chains. PMID- 15268156 TI - Freezing and folding behavior in simple off-lattice heteropolymers. AB - We have performed parallel tempering Monte Carlo simulations using a simple continuum heteropolymer model for proteins. All 10 heteropolymer sequences which we have studied have shown first-order transitions at low temperature to ordered states dominated by single chain conformations. These results are in contrast with the theoretical predictions of the random energy model for heteropolymers, from which we would expect continuous transitions to glassy behavior at low temperatures. PMID- 15268157 TI - Folding behavior of model proteins with weak energetic frustration. AB - The native structure of fast-folding proteins, albeit a deep local free-energy minimum, may involve a relatively small energetic penalty due to nonoptimal, though favorable, contacts between amino acid residues. The weak energetic frustration that such contacts represent varies among different proteins and may account for folding behavior not seen in unfrustrated models. Minimalist model proteins with heterogeneous contacts--as represented by lattice heteropolymers consisting of three types of monomers--also give rise to weak energetic frustration in their corresponding native structures, and the present study of their equilibrium and nonequilibrium properties reveals some of the breadth in their behavior. In order to capture this range within a detailed study of only a few proteins, four candidate protein structures (with their cognate sequences) have been selected according to a figure of merit called the winding index--a characteristic of the number of turns the protein winds about an axis. The temperature-dependent heat capacities reveal a high-temperature collapse transition, and an infrequently observed low-temperature rearrangement transition that arises because of the presence of weak energetic frustration. Simulation results motivate the definition of a new measure of folding affinity as a sequence-dependent free energy--a function of both a reduced stability gap and high accessibility to non-native structures--that correlates strongly with folding rates. PMID- 15268158 TI - Simulation of the effects of chain architecture on the sorption of ethylene in polyethylene. AB - An osmotic ensemble hyperparallel tempering technique has been developed to study the solubility of ethylene in amorphous linear low-density polyethylene of different chain architectures. The NERD united-atom force field (Nath, Escobedo, and de Pablo revised united-atom force field) is used in all simulations. We have investigated the effect of polyethylene chain length and branching on ethylene solubility. In this study, we have considered short-chain branching of amorphous linear low-density ethylene-1-hexene copolymers under typical polymerization reactor conditions. It is observed that, in the polymer, ethylene prefers to reside in the vicinity of polymer chain ends. This clustering causes a decrease in ethylene solubility with polymer chain length. When short-chain branches are introduced to a linear polymer chain, however, the chain-end clustering effect is counteracted by a higher density, thereby leading to an ethylene solubility almost identical to that in the linear polymer. PMID- 15268159 TI - Phase and orientational ordering of low molecular weight rod molecules in a quenched liquid crystalline polymer matrix with mobile side chains. AB - We study the phase diagram and orientational ordering of guest liquid crystalline (LC) rods immersed in a quenched host made of a liquid crystalline polymer (LCP) matrix with mobile side chains. The LCP matrix lies below the glass transition of the polymer backbone. The side chains are mobile and can align to the guest rod molecules in a plane normal to the local LCP chain contour. A field theoretic formulation for this system is proposed and the effects of the LCP matrix on LC ordering are determined numerically. We obtain simple analytical equations for the nematic/isotropic phase diagram boundaries. Our calculation show a nematic nematic (N/N) first order transition from a guest stabilized to a guest-host stabilized region and the possibility of a reentrant transition from a guest stabilized nematic region to a host only stabilized regime separated by an isotropic phase. A detailed study of thermodynamic variables and interactions on orientational ordering and phases is carried out and the relevance of our predictions to experiments and computer simulations is presented. PMID- 15268160 TI - Nuclear magnetic resonance study of self-diffusion in liquid crystals. AB - A simple and accurate pulsed-gradient nuclear magnetic resonance technique for measuring coefficients of self-diffusion in liquid crystals is described. It is based on exciting sharp response signals with long weak pulses. The method uses an extremely weak radio-frequency field, which eliminates the problem of radio frequency heating of the sample. The temperature dependencies of coefficients of self-diffusion for two liquid crystals, 5CB (4-pentyl-4(')-cyanobiphenyl) and EBBA (N-(4-ethoxybenzylidene)-4-butylaniline), are presented. PMID- 15268161 TI - Nonadiabatic transition in the dissociation process from inner valence states of O(2) (+). AB - In a previous paper we reported a study of the electronic structures of inner valence states of O(2) (+) and the dissociation process, where there remained some questions as to the origins of the dissociation fragment formation of the O+((2)D)+O((3)P) limit in observed spectra. In this paper, we present the results of calculations of the nonadiabatic transition probabilities of the multichannel dissociation process from the inner valence states of O(2) (+) and reproduce the general features of observed spectra previously reported, including fragment formation, using the Zhu-Nakamura theory. PMID- 15268163 TI - Low-energy electron induced restructuring of water monolayers on NaCl(100). AB - The influence of electron irradiation on the controversially discussed monolayer structure of H(2)O on NaCl(100) is investigated with helium atom diffraction before and after a low-damage low-energy electron diffraction (LEED) experiment. The ordered (1x1) structure observed initially with He atoms is found to be transformed to a stable c(4x2) structure after a 90 eV electron dosage of only 10(15) electrons cm(-2) or about 2 incident electrons per adsorbate molecule. Based on previously reported structure models for the two phases, the transition is attributed to a reorientation, and a possible compression of the water film induced by the electrons. PMID- 15268164 TI - Proton transfer dynamics via high resolution spectroscopy in the gas phase and instanton calculations. AB - Tunneling splittings have been observed in the eigenstate-resolved electronic spectrum of the 2-hydroxypyridine/2-pyridone dimer in the gas phase. Deuterium substitution experiments show that these splittings are caused by a concerted double proton transfer reaction along the O-H...O and N...H-N hydrogen bonds that hold the dimer together, substitution of the weaker and longer N...H-N bond having the larger effect. Tunneling splittings calculated by the instanton method for the zero-point level of the ground state are in good agreement with experiment for all observed isotopomers, showing that the dynamics occurs in this state, rather than in the electronically excited state. PMID- 15268165 TI - Liquids confined in wedge shaped pores: nonuniform pressure induced by pore geometry. AB - Lennard-Jones liquids confined in wedge shaped nanopores are investigated using molecular dynamics computer simulations. We show that small deviations from the parallel slit geometry result in nonuniform pressures and density profiles along the pore. In conditions of high confinement and thermodynamic states close to the triple point, wedge shaped pores can induce the formation of solid phases in specific regions within the nanopore. PMID- 15268166 TI - Fragmentation dynamics of ionized neon trimer inside helium nanodroplets: a theoretical study. AB - We report a theoretical study of the fragmentation dynamics of Ne(3) (+) inside helium nanodroplets, following vertical ionization of the neutral neon trimer. The motion of the neon atoms is treated classically, while transitions between the electronic states of the ionic cluster are treated quantum mechanically. A diatomics-in-molecules description of the potential energy surfaces is used, in a minimal basis set consisting of three effective p orbitals on each neon atom for the missing electron. The helium environment is modeled by a friction force acting on the neon atoms when their speed exceeds the Landau velocity. A reasonable range of values for the corresponding friction coefficient is obtained by comparison with existing experimental measurements. PMID- 15268167 TI - Magic clusters MAu4 (M=Ti and Zr) and their dimers: how magic are they? AB - The stability of closed shell bimetallic magic clusters MAu(4) (M=Ti and Zr) is investigated theoretically through ab initio molecular orbital calculations. Both these clusters have tetrahedral structures and are found to be associated with large values of the ionization potential, HOMO-LUMO gap as well as the binding energies, which are characteristic of the magic clusters. However, the cluster cluster interaction energy corresponding to a dimer formation is found to be unusually high ( approximately 5-7 eV) in contradiction to the usual properties of a magic cluster and is attributed to a 3-center-2-electron M-Au-M type bridge bonding as well as aurophilic attraction. Gross geometrical features of the individual clusters are, however, mostly retained in the dimer, thus satisfying the basic requirements for the cluster-assembled materials. This work would have important implications in the design of novel cluster-based nanomaterials for various nanoscale applications. PMID- 15268168 TI - Mesoscopic chiral reshaping of the Ag(110) surface induced by the organic molecule PVBA. AB - We report scanning tunneling microscopy observations on the restructuring of a Ag(110) surface induced by the molecule 4-[trans-2-(pyrid-4-yl-vinyl)]benzoic acid (PVBA). Our data reveal that the surface undergoes a mesoscopic step faceting following exposure to submonolayer coverages and thermal activation. A sawtooth arrangement evolves implying long-range mass transport of substrate atoms and forming a regular arrangement of kink sites. Its formation is associated with the molecules' functional headgroups forming carboxylates with [100] Ag microfacets at step edges, and eventually operating to reshape the surface morphology. Interestingly, the resulting microfacets act as chiral templates for the growth of supramolecular PVBA structures. Theoretical modeling based on ab initio results indicates that chiral recognition processes discriminating between the two enantiomers of adsorbed PVBA molecules occur in this process. PMID- 15268169 TI - Vibronic states in single molecules: C60 and C70 on ultrathin Al2O3 films. AB - Vibronic states are observed in single C(60) and C(70) molecules by scanning tunneling microscopy. When single fullerene molecules are adsorbed on a thin layer of Al(2)O(3) grown on a NiAl(110) substrate, equally spaced features are observed in the differential conductance (dI/dV), which are clearly resolved in d(2)I/dV(2) spectra. These features are attributed to the vibronic states of the molecule. The vibronic progressions are sensitive to the molecular orientations and can have different spacings in different electronic bands of the same molecule. For C(60,) these vibronic states are associated with the intramolecular A(g) and H(g) vibrational modes. Vibronic states are not resolved in molecules adsorbed on the metal surface. However, inelastic electron tunneling spectroscopy exhibits a vibrational mode at 64 meV for C(60) and 61 meV for C(70) adsorbed on NiAl(110). PMID- 15268170 TI - No evidence for large-scale proton ordering in Antarctic ice from powder neutron diffraction. AB - We have examined a sample of 3000 year old Antarctic ice, collected at the Kohnen Station, by time-of-flight powder neutron diffraction to test the hypothesis of Fukazawa et al. [e.g., Ann. Glaciol. 31, 247 (2000)] that such ice may be partially proton ordered. Great care was taken to keep our sample below the proposed ordering temperature (237 K) at all times, but we did not observe any evidence of proton ordering. PMID- 15268171 TI - An analysis of electronic dephasing in the spin-boson model. AB - In order to develop a more complete understanding of the limitations of mixed quantum-classical simulation methods, the origins of electronic dephasing are analyzed in a simple model of the condensed phase, namely, the spin-boson model with an ohmic spectral density. We focus on the decay of the thermally averaged nuclear overlap/phase function (NOPF). Considering the strong coupling/high temperature limit, a relationship is obtained at short time between the rate of electronic coherence loss and the electronic dephasing rate characteristic of a classical bath. Using this relationship, we clarify the origin of the decay of the NOPF. In the same limit, we also reproduce an earlier relationship between the electronic decoherence time and a solvation relaxation time. Finally, we point out that, for the spin-boson model, the exact quantum mechanical description of electronic dephasing is reproduced by mixed quantum/classical methods if a Gaussian distribution of quantum fluctuations around each classical phase space point is introduced. That spatial distribution of quantum fluctuations is functionally the same as that appearing in the Feynman-Kleinert variational local harmonic approximation, and also that implemented in existing classical trajectory-based estimates of coherence dissipation times. PMID- 15268172 TI - Theoretical method for full ab initio calculation of DNA/RNA-ligand interaction energy. AB - In this paper, we further develop the molecular fractionation with conjugate caps (MFCC) scheme for quantum mechanical computation of DNA-ligand interaction energy. We study three oligonuclear acid interaction systems: dinucleotide dCG/water, trinucleotide dCGT/water, and a Watson-Crick paired DNA segment, dCGT/dGCA. Using the basic MFCC approach, the nucleotide chains are cut at each phosphate group and a pair of conjugate caps (concaps) are inserted. Five cap molecules have been tested among which the dimethyl phosphate anion is proposed to be the standard concap for application. For each system, one-dimensional interaction potential curves are computed using the MFCC method and the calculated interaction energies are found to be in excellent agreement with corresponding results obtained from the full system ab initio calculations. The current study extends the application of the MFCC method to ab initio calculations for DNA- or RNA-ligand interaction energies. PMID- 15268173 TI - Fast semiempirical calculations for nuclear magnetic resonance chemical shifts: a divide-and-conquer approach. AB - A new approach to calculate nuclear magnetic resonance chemical shifts has been implemented at the semiempirical modified neglect of diatomic overlap level using gauge-including atomic orbitals. The perturbed density matrix with respect to the magnetic field is obtained by the diagonalization of the complex Fock matrix using the divide and conquer (DC) method, instead of by solving the computationally expensive coupled perturbed Hartree-Fock equations. Adopting the Patchkovskii and Thiel parameters [S. Patchkovskii and W. Thiel J. Comput. Chem. 20, 1220 (1999)], we were able to reproduce their results for small organic molecules. The errors introduced by DC method are negligible, as shown by the calculations on a series of polyalaine structures. Test calculations on proteins have demonstrated that our approach makes it possible to calculate chemical shifts routinely on systems with hundreds of atoms with good accuracy. PMID- 15268174 TI - An adaptive immune optimization algorithm for energy minimization problems. AB - Based on the immune theory of biology, a novel evolutionary algorithm, adaptive immune optimization algorithm (AIOA), is proposed. In AIOA, density regulation and immune selection is adopted to control the individual diversity and the convergence adaptively. By an application of the algorithm to the optimization of test functions, it is shown that the algorithm is a highly efficient optimization method compared with other stochastic optimization methods. The algorithm was also applied to the optimization of Lennard-Jones clusters, and the results show that the method can find the optimal structure of NHNC isomerization dynamics induced by sub-one cycle and few-cycle IR pulses, which we represent as Gaussian pulses with 0.25-2 optical cycles in the pulse width. Starting from vibrationally pre-excited states, isomerization probabilities of up to 50% are obtained for optimized pulses. With decreasing number of optical cycles a strong dependence on the carrier-envelope phase (CEP) emerges. Although the optimized pulse parameters change significantly with the number of optical cycles, the distortion by the Gaussian envelope produces nearly equal fields, with a positive lobe followed by a negative one. The positions and areas of the lobes are also almost unchanged, irrespective of the number of cycles in the half-width. Isomerization proceeds via a pump-dumplike mechanism induced by the sequential lobes. The first lobe prepares a wave packet incorporating many delocalized states above the barrier. It is the motion of this wave packet across the barrier, which determines the timing of the pump and dump lobes. The role of the pulse parameters, and in particular of the CEP, is to produce the correct lobe sequence, size and timing within a continuous pulse. PMID- 15268188 TI - Molecular isomerization induced by ultrashort infrared pulses. II. Pump-dump isomerization using pairs of time-delayed half-cycle pulses. AB - We investigate population transfer across the barrier in a double-well potential, induced by a pair of time-delayed single-lobe half-cycle pulses. We apply this setup both to a one-dimensional (1D) quartic model potential and to a three dimensional potential representing HCN-->HNC isomerization. Overall the results for the two systems are similar, although in the 3D system some additional features appear not seen in the 1D case. The generic mechanism of population transfer is the preparation by the pump pulse of a wave packet involving delocalized states above the barrier, followed by the essentially 1D motion of the delocalized part of wave packet across the barrier, and the eventual de excitation by the dump pulse to localized states in the other well. The correct timing is given by the well-to-well passage time of the wave packet and its recurrence properties, and by the signs of the field lobes which determine the direction and acceleration or deceleration of the wave packet motion. In the 3D system an additional pump-pump-dump mechanism linked to wave packet motion in the reagent well can mediate isomerization. Since the transfer time and the pulse durations are of the same order of magnitude, there is also a marked dependence of the dynamics and the transfer yield on the pulse duration. Our analysis also sheds light on the pronounced carrier envelope phase dependence previously observed for isomerization and molecular dissociation with one-cycle and sub-one cycle pulses. PMID- 15268189 TI - Theoretical study of the UV photodissociation of Cl2: potentials, transition moments, extinction coefficients, and Cl*/Cl branching ratio. AB - Potential energy curves for the X (1)Sigma(g) (+) ground state and Omega=0(u) (+), 1(u) valence states and dipole moments for the 0(u) (+), 1(u)-X transitions are obtained in an ab initio configuration interaction study of Cl(2) including spin-orbit coupling. In contrast to common assumptions, it is found that the B (3)Pi(0(+)u)-X transition moment strongly depends on internuclear distance, which has an important influence on the Cl(2) photodissociation. Computed energy curves and transition moments are employed to calculate the A, B, C<--X extinction coefficients, the total spectrum for the first absorption band, and the Cl(*)((2)P(1/2))/Cl((2)P(3/2)) branching ratio as a function of excitation wavelength. The calculated data are shown to be in good agreement with available experimental results. PMID- 15268190 TI - Theory of the two step enantiomeric purification of 1,3 dimethylallene. AB - An application of a recently proposed [P. Kral et al., Phys. Rev. Lett. 90, 033001 (2003)] two step optical control scenario to the purification of a racemic mixture of 1,3 dimethylallene is presented. Both steps combine adiabatic and diabatic passage phenomena. In the first step, three laser pulses of mutually perpendicular linear polarizations, applied in a "cyclic adiabatic passage" scheme, are shown to be able to distinguish between the L and D enantiomers due to their difference in matter-radiation phase. In the second step, which immediately follows the first, a sequence of pulses is used to convert one enantiomer to its mirror-imaged form. This scenario, which only negligibly populates the first excited electronic state, proves extremely useful for systems such as dimethylallene, which can suffer losses from dissociation and internal conversion upon electronic excitation. We computationally observe conversion of a racemic mixture of dimethylallene to a sample containing approximately 95% of the enantiomer of choice. PMID- 15268191 TI - Resonance Raman spectra of uracil based on Kramers-Kronig relations using time dependent density functional calculations and multireference perturbation theory. AB - The use of time-dependent density functional calculations for the optimization of excited-state structures and the subsequent calculation of resonance Raman intensities within the transform-theory framework is compared to calculations of Hartree-Fock/configuration interaction singles-type (CIS). The transform theory of resonance Raman scattering is based on Kramers-Kronig relations between polarizability tensor components and the optical absorption. Stationary points for the two lowest excited singlet states of uracil are optimized and characterized by means of numerical differentiation of analytical excited-state gradients. It is shown that the effect of electron correlation leads to substantial modifications of the relative intensities. Calculations of vibrational frequencies for ground and excited states are carried out, which show that the neglect of Duschinsky mixing and the assumption of equal wave numbers for ground and excited state are not in all cases good approximations. We also compare the transform-theory resonance Raman intensities with those obtained within a simple approximation from excited-state gradients at the ground-state equilibrium position, and find that they are in qualitative agreement in the case of CIS, but show some important differences in calculations based on density functional theory. Since the results from CIS calculations are in better agreement with experiment, we also present approximate resonance Raman spectra obtained using excited-state gradients from multireference perturbation theory calculations, which confirm the CIS gradients. PMID- 15268192 TI - Vacuum ultraviolet mass-analyzed threshold ionization spectroscopy of hexafluorobenzene: the Jahn-Teller effect and vibrational analysis. AB - One-photon mass-analyzed threshold ionization (MATI) spectrum of hexafluorobenzene was obtained by using vacuum ultraviolet radiation generated by four-wave difference frequency mixing in Kr. The ionization energy of hexafluorobenzene determined from the position of the 0-0 band was 9.9108+/ 0.0006 eV. To aid the spectral analysis, the Jahn-Teller coupling parameters for four e(2g) modes of C(6)F(6) (+) in the ground electronic state were calculated from the topographical data of the potential energy surface obtained at the density functional theory (DFT) level. These were used in the initial calculation of the energies of the Jahn-Teller states and upgraded through the multimode fit to the experimental data. Excellent agreement between the experimental and calculated frequencies was achieved. The vibrations which are not linear Jahn Teller active were observed and could be assigned by referring to the frequencies obtained at the DFT level. PMID- 15268193 TI - Toward subchemical accuracy in computational thermochemistry: focal point analysis of the heat of formation of NCO and [H,N,C,O] isomers. AB - In continuing pursuit of thermochemical accuracy to the level of 0.1 kcal mol( 1), the heats of formation of NCO, HNCO, HOCN, HCNO, and HONC have been rigorously determined using state-of-the-art ab initio electronic structure theory, including conventional coupled cluster methods [coupled cluster singles and doubles (CCSD), CCSD with perturbative triples (CCSD(T)), and full coupled cluster through triple excitations (CCSDT)] with large basis sets, conjoined in cases with explicitly correlated MP2-R12/A computations. Limits of valence and all-electron correlation energies were extrapolated via focal point analysis using correlation consistent basis sets of the form cc-pVXZ (X=2-6) and cc-pCVXZ (X=2-5), respectively. In order to reach subchemical accuracy targets, core correlation, spin-orbit coupling, special relativity, the diagonal Born Oppenheimer correction, and anharmonicity in zero-point vibrational energies were accounted for. Various coupled cluster schemes for partially including connected quadruple excitations were also explored, although none of these approaches gave reliable improvements over CCSDT theory. Based on numerous, independent thermochemical paths, each designed to balance residual ab initio errors, our final proposals are DeltaH(f,0) ( composite function )(NCO)=+30.5, DeltaH(f,0) ( composite function )(HNCO)=-27.6, DeltaH(f,0) ( composite function )(HOCN)=-3.1, DeltaH(f,0) ( composite function )(HCNO)=+40.9, and DeltaH(f,0) ( composite function )(HONC)=+56.3 kcal mol(-1). The internal consistency and convergence behavior of the data suggests accuracies of +/-0.2 kcal mol(-1) in these predictions, except perhaps in the HCNO case. However, the possibility of somewhat larger systematic errors cannot be excluded, and the need for CCSDTQ [full coupled cluster through quadruple excitations] computations to eliminate remaining uncertainties is apparent. PMID- 15268195 TI - Equilibrium geometries of low-lying isomers of some Li clusters, within Hartree Fock theory plus bond order or MP2 correlation corrections. AB - In a recent study by Kornath et al. [J. Chem. Phys. 118, 6957 (2003)], the Li(n) clusters with n=2, 4, and 8 have been isolated in argon matrices at 15 K and characterized by Raman spectroscopy. This has prompted us to carry out a theoretical study on such clusters up to n=10, using Hartree-Fock theory, plus low-order Moller-Plesset perturbation corrections. To check against the above study of Kornath et al., as a by-product we have made the same approximations for n=6 and 8 as we have for n=10. This has led us to emphasize trends with n through the Li(n) clusters for (i) ground-state energy, (ii) HOMO-LUMO energy gap, (iii) dissociation energy, and (iv) Hartree-Fock eigenvalue sum. The role of electron correlation in distinguishing between low-lying isomers is plainly crucial, and will need a combination of experiment and theory to obtain decisive results such as that of Kornath et al. for Li(8). In particular, it is shown that Hartree-Fock theory plus bond order correlations does account for the experimentally observed symmetry T(d) symmetry for Li(8). PMID- 15268194 TI - Fragmentation and deformation mechanism of glycine isomers in gas phase: Investigations of charge effect. AB - The structural parameters, relative stability, proton transfer energy barriers of four typical and life related isomers and conformers of different charged (n=0,+/ 1,+/-2) glycine species have been investigated using B3LYP, BHLYP, and CCSD(T) methods. Results indicate that those neutral and (+/-1)-charged species are stable. For the (+2)-charged cases, all four triplet-state glycine species and only the singlet-state zwitterionic one are stable. On the other hand, only the singlet-state zwtterionic glycine ((1)GlyZW(-2)) and the corresponding neutral form counterpart ((1)Gly(-2)) are stable for the (-2)-charged cases. Either of the two stable structures holds a proton lying in the position (2-3 A) of being separated from its corresponding parental species. Those unstable divalent glycine species are dissociated into different smaller species spontaneously according to the characters of their different structures and electron spins. The presented fragmentation and deformation mechanisms can effectively predict and satisfactorily explain some experimental phenomena, which had been puzzling the mass spectrometry chemists. Also, the mechanisms should be suitable for any other similar molecule systems. Comparisons of the relative energies of the four (+1) charged glycine species show that doublet-state glycine III ((2)GlyIII1) is more stable in energy by 12.1 kcal/mol than the (+1)-charged glycine Gly ((2)Gly1). This is consistent with the energy ordering of their corresponding mono-valence metal ion-bound derivatives. In addition, calculations show that an intramolecular proton transfer of (2)Gly(-1) to become its zwitterionic counterpart is preferred due to its least activation energy barrier (5.8 kcal/mol) among four discussed processes. PMID- 15268196 TI - Homogeneous nucleation rates of higher n-alcohols measured in a laminar flow diffusion chamber. AB - Nucleation rate isotherms of n-butanol, n-pentanol, n-hexanol, n-heptanol, and n octanol were measured in a laminar flow diffusion chamber using helium as carrier gas. The measurements were made at 250-310 K, corresponding to reduced temperatures of 0.43-0.50, and at atmospheric pressure. Experimental nucleation rate range was from 10(3) to 10(7) cm(-3) s(-1). The expression and accuracy of thermodynamic parameters, in particular equilibrium vapor pressure, were found to have a significant effect on calculated nucleation rates. The results were compared to the classical nucleation theory (CNT), the self-consistency corrected classical theory (SCC) and the Hale's scaled model of the CNT. The average ratio between the experimental and theoretical nucleation rates for all alcohols used was 1.5x10(3) when the CNT was used, and 0.2x10(-1) when the SCC was used and 0.7x10(-1) when the Hale's scaled theory was used. The average values represent all the alcohols used at the same reduced temperatures. The average ratio was about the same throughout the temperature range, although J(exp)/J(the) calculated with the Hale's scaled theory increased slightly with increasing temperature. The saturation ratio dependency was predicted closest to experiment with the classical nucleation theory. The nucleation rates were compared to those found in the literature. The measurements were in reasonable agreement with each other. The molecular content of critical alcohol clusters was between 35 and 80 molecules. At a fixed reduced temperature, the number of molecules in a critical cluster decreased as a function of alcohol carbon chain length. The number of molecules in critical clusters was compared to those predicted by the Kelvin equation. The theory predicted the critical cluster sizes well. PMID- 15268197 TI - Real time observation of the photo-Fries rearrangement. AB - The photo-Fries rearrangement of 4-tert-butylphenyl acetate dissolved in cyclohexane is investigated by two-color femtosecond pump probe spectroscopy. The spectral transmission changes are characterized in the visible and ultraviolet spectral region and allow for the first time to temporally resolve the primary reaction steps. We find that the photoinduced homolytic cleavage of the CO bond occurs within 2 ps and that the geminate recombination of the generated radical pair to the intermediate substituted cyclohexadienone takes 13 ps. The experimental results support a model in which the initial reaction proceeds from the originally excited pipi(*) state via a barrier to a dissociative pisigma(*) state. PMID- 15268198 TI - Nonequilibrium melting and crystallization of a model Lennard-Jones system. AB - Nonequilibrium melting and crystallization of a model Lennard-Jones system were investigated with molecular dynamics simulations to quantify the maximum superheating/supercooling at fixed pressure, and over-pressurization/over depressurization at fixed temperature. The temperature and pressure hystereses were found to be equivalent with regard to the Gibbs free energy barrier for nucleation of liquid or solid. These results place upper bounds on hysteretic effects of solidification and melting in high heating- and strain-rate experiments such as shock wave loading and release. The authors also demonstrate that the equilibrium melting temperature at a given pressure can be obtained directly from temperatures at the maximum superheating and supercooling on the temperature hysteresis; this approach, called the hysteresis method, is a conceptually simple and computationally inexpensive alternative to solid-liquid coexistence simulation and thermodynamic integration methods, and should be regarded as a general method. We also found that the extent of maximum superheating/supercooling is weakly pressure dependent, and the solid-liquid interfacial energy increases with pressure. The Lindemann fractional root-mean squared displacement of solid and liquid at equilibrium and extreme metastable states is quantified, and is predicted to remain constant (0.14) at high pressures for solid at the equilibrium melting temperature. PMID- 15268199 TI - Double hydrogen tunneling revisited: the breakdown of experimental tunneling criteria. AB - Formic acid dimer was chosen as a model system to investigate synchronous double proton transfer by means of variational transition state theory (VTST) for various isotopically modified hydrogen species. The electronic barrier for the double proton transfer was evaluated to be 7.9 kcal/mol, thus being significantly lower than it was determined in previous studies. The tunneling probabilities were evaluated at temperatures from 100 up to 400 K and typical Arrhenius behavior with enhancement by tunneling is observed. When comparing the transmission factors kappa in dependence of the mass of the tunneling hydrogen, it was found that there are two maxima, one at very low masses (e.g., 0.114 amu, corresponding to the muonium entity) and one maximum at around 2 amu (corresponding to deuterium). With the knowledge of the VTST-hydrogen transfer rates and the corresponding tunneling corrections, various tunneling criteria were tested (e.g., Swain-Schaad exponents) and were shown to fail in this reaction in predicting the extent of tunneling. This finding adds another aspect in the ongoing "Tunneling-Enhancement by Enzymes" discussion, as the used tunneling criteria based on experimental reaction rates may fail to predict tunneling behavior correctly. PMID- 15268200 TI - Critical cavities and the kinetic spinodal for superheated liquids. AB - We present density-functional theory (DFT) calculations for critical cavities inside model superheated liquids with varying intermolecular potentials. Our calculations show that the radius of the critical cavity and the ratio of the work of formation of the critical cavity to the work of formation of the critical bubble as predicted by the classical nucleation theory exhibit universal scaling across similar intermolecular potentials. We then utilize this observed scaling behavior by proposing two new criteria for the kinetic spinodal of superheated liquids. These criteria are based on various properties of the critical cavity as obtained from our DFT studies of the superheated Lenanrd-Jones liquid. Our predictions of the kinetic spinodal compare favorably with experimental data of the limits of superheating for various organic liquids. PMID- 15268201 TI - Spontaneous transformation of water's high-density amorph and a two-stage crystallization to ice VI at 1 GPa: a dielectric study. AB - Dielectric relaxation spectra of a metastable crystal phase formed on implosive and exothermic transformation of pressure-amorphized hexagonal ice have been measured in situ at 0.97 GPa pressure over a range of temperature. The metastable phase showed no relaxation peak at 130 K and 0.97 GPa. When heated at a fixed pressure of 0.97 GPa, it began to transform at approximately 145 K exothermally to a phase whose relaxation rate and equilibrium dielectric permittivity increased. A second, but slower exothermic transformation also occurred at approximately 175 K. After keeping at 213 K, the relaxation rate and equilibrium permittivity reached the known values of these two quantities for ice VI. Thus the metastable phase transformed to ice VI in two stages. It is conjectured that the intermediate phase in this transformation could be ice XII. The rate of transformation is not determined by the reorientational relaxation rate of water molecules in the ices. PMID- 15268202 TI - Transitions between disordered phases in supercooled liquid silicon. AB - We have investigated the transitions between disordered phases in supercooled liquid silicon using computer simulations. The thermodynamic properties were directly obtained from the free energy, which was computed using the recently proposed reversible scaling method. The calculated free energies of the crystalline and liquid phases of silicon at zero pressure, obtained using the environment dependent interatomic potential, are in excellent agreement with the available experimental data. The results show that, at zero pressure, a weak first-order liquid-liquid transition occurs at 1135 K and a continuous liquid amorphous transition takes place at 843 K. These results are consistent with the existence of a second critical point for the liquid-liquid transition at a negative pressure. PMID- 15268203 TI - Nonmonotonic temperature dependence of heat capacity through the glass transition within a kinetic model. AB - The heat capacity of a supercooled liquid subjected to a temperature cycle through its glass transition is studied within a kinetic model. In this model, the beta process is assumed to be thermally activated and described by a two level system. The alpha process is described as a beta relaxation mediated cooperative transition in a double well. The overshoot of the heat capacity during the heating scan is well reproduced and is shown to be directly related to delayed energy relaxation in the double well. In addition, the calculated scan rate dependencies of the glass transition temperature T(g) and the limiting fictive temperature T(f) (L) show qualitative agreement with the known results. Heterogeneity is found to significantly reduce the overshoot of heat capacity. Furthermore, the frequency dependent heat capacity has been calculated within the present framework and found to be rather similar to the experimentally observed behavior of supercooled liquids. PMID- 15268204 TI - New phase for one-component hard spheres. AB - A completely new phase for one-component hard spheres is reported in an unexpected region of the phase diagram. The new phase is observed at compressibility factors intermediate between the solid and the metastable branches. It can be obtained from either Monte Carlo simulations alone or a combination of Monte Carlo and molecular dynamics calculations. An analysis of the intermediate scattering function data shows that the new phase is in a stable equilibrium. Radial distribution function data, configurational snapshots, bond order parameters, and translational order parameters obtained from molecular simulations indicate that the new phase is significantly different from the isotropic liquid, metastable, or crystalline phases traditionally observed in hard sphere systems. This result significantly changes our previous understanding of the behavior of hard spheres. PMID- 15268205 TI - Collective contributions to the dielectric relaxation of hydrogen-bonded liquids. AB - Dielectric relaxation times are often interpreted in terms of the reorientation of dipolar species or aggregates. The relevant time correlation function contains, however, cross terms between dipole moments of different particles. In the static case, these cross terms are accounted for by the Kirkwood factor g(K). Theories and molecular dynamics simulations suggest that such cross correlations may also affect the time-dependent properties, as reflected in the dielectric spectra. We present an experimental method for detecting effects of such cross correlations in dielectric spectra by a comparative analysis of dielectric and magnetic relaxation data. We demonstrate that such collective contributions can substantially affect dielectric relaxation. Experiments for n-pentanol (g(K)=3.06 at 298 K) and 2,2-dimethyl-3-ethyl-pentane-3-ol (g(K)=0.59) and their solutions in carbon tetrachloride show that in systems with g(K)>1, the cross correlations slow down dielectric relaxation. In systems with g(K)<1, dielectric relaxation is enhanced. The results conform to theoretical predictions by Madden and Kivelson [Adv. Chem. Phys. 56, 467 (1984)] and to results of molecular dynamics simulations. The relaxation enhancement by cross terms in the case of g(K)<1 is difficult to rationalize by conventional models of dielectric relaxation. PMID- 15268206 TI - Self-consistent Ornstein-Zernike approximation for the Sogami-Ise fluid. AB - We generalize the self-consistent Ornstein-Zernike approximation (SCOZA) to a fluid of particles with a pair potential consisting of a hard-core repulsion and a linear combination of Sogami-Ise tails, w(r)=-epsilonsigma summation operator (nu)(K(nu)/r+L(nu)z(nu))e(-z(nu)(r-sigma)). The formulation and implementation of the SCOZA takes advantage of the availability of semianalytic results for such systems within the mean-spherical approximation. The predictions for the thermodynamics, the phase behavior and the critical point are compared with optimized random phase approximation results; further, the effect of thermodynamic consistency is investigated. PMID- 15268207 TI - Ultrafast dynamics for electron photodetachment from aqueous hydroxide. AB - Charge-transfer-to-solvent reactions of hydroxide induced by 200 nm monophotonic or 337 and 389 nm biphotonic excitation of this anion in aqueous solution have been studied by means of pump-probe ultrafast laser spectroscopy. Transient absorption kinetics of the hydrated electron, e(aq) (-), have been observed, from a few hundred femtoseconds out to 600 ps, and studied as function of hydroxide concentration and temperature. The geminate decay kinetics are bimodal, with a fast exponential component ( approximately 13 ps) and a slower power "tail" due to the diffusional escape of the electrons. For the biphotonic excitation, the extrapolated fraction of escaped electrons is 1.8 times higher than for the monophotonic 200 nm excitation (31% versus 17.5% at 25 degrees C, respectively), due to the broadening of the electron distribution. The biphotonic electron detachment is very inefficient; the corresponding absorption coefficient at 400 nm is <4 cm TW(-1) M(-1) (assuming unity quantum efficiency for the photodetachment). For [OH(-)] between 10 mM and 10 M, almost no concentration dependence of the time profiles of solvated electron kinetics was observed. At higher temperature, the escape fraction of the electrons increases with a slope of 3x10(-3) K(-1) and the recombination and diffusion-controlled dissociation of the close pairs become faster. Activation energies of 8.3 and 22.3 kJ/mol for these two processes were obtained. The semianalytical theory of Shushin for diffusion controlled reactions in the central force field was used to model the geminate dynamics. The implications of these results for photoionization of water are discussed. PMID- 15268208 TI - Second order average Hamiltonian theory of symmetry-based pulse schemes in the nuclear magnetic resonance of rotating solids: application to triple-quantum dipolar recoupling. AB - The average Hamiltonian theory (AHT) of several classes of symmetry-based radio frequency pulse sequences is developed to second order, allowing quantitative analyses of a wide range of recoupling and decoupling applications in magic-angle spinning solid state nuclear magnetic resonance. General closed analytical expressions are presented for a cross term between any two interactions recoupled to second order AHT. We classify them into different categories and show that some properties of the recoupling pulse sequence may be predicted directly from this classification. These results are applied to examine a novel homonuclear recoupling strategy, effecting a second order average dipolar Hamiltonian comprising trilinear triple quantum (3Q) spin operators. We discuss general features and design principles of such 3Q recoupling sequences and demonstrate by numerical simulations and experiments that they provide more efficient excitation of (13)C 3Q coherences compared to previous techniques. We passed up to 15% of the signal through a state of 3Q coherence in rotating powders of uniformly (13)C labeled alanine and tyrosine. Second order recoupling-based (13)C homonuclear 3Q correlation spectroscopy is introduced and demonstrated on tyrosine. PMID- 15268209 TI - Stokes/anti-Stokes anomalies under surface enhanced Raman scattering conditions. AB - The possibility of achieving anti-Stokes stimulation and/or pumping under surface enhanced Raman scattering conditions has been the source of intense controversies in the literature. With the aim of clarifying some of the aspects of this problem, we study theoretically and experimentally the situation in a model system which highlights some of the difficulties in the interpretation of the data. We show that many of the assumptions often presumed in the literature need to be assessed with care in each case. Through careful experiments we show, in particular, that the anti-Stokes/Stokes ratio for a specific mode in the same sample can depend on the chosen laser wavelength. This latter effect is a manifestation of the influence of the internal plasmon resonances (hot spots) in the result. Different possibilities and further research directions are highlighted and discussed. PMID- 15268210 TI - Liquid-vapor interface of square-well fluids of variable interaction range. AB - Properties of the liquid-vapor interface of square-well fluids with ranges of interaction lambda=1.5, 2.0, and 3.0 are obtained by Monte Carlo simulations and from square-gradient theories that combine the Carnahan-Starling equation of state for hard spheres with the second and third virial coefficients. The predicted surface tensions show good agreement with the simulation results for lambda=2 and for lambda=3 in a temperature range reasonably close to the critical point, 0.8/=20 kJ mol(-1). This is much lower than the threshold incidence energy obtained for penetration of ice by HCl (E(i) approximately 96.5 kJ mol(-1)); the calculated average barrier to penetration of ice by HF is 16.0 kJ mol(-1) and is much lower than that previously reported for HCl. As was the case for HCl, penetration of ice by HF decreases with decreasing incidence energy and increasing incidence angle. Though in general, the penetration probability is independent of the molecule's initial rotational energy, penetration beyond the second bilayer (deep penetration) is suppressed by initial rotation. This suggests that, like was found for HCl, the steering operative in deep penetration is inhibited by initial rotation. Finally, because HF is a weak acid experimental observation of HF penetrated into ice may well be possible using infrared spectroscopy, and we suggest experiments along this line. PMID- 15268215 TI - Concentration-dependent self-diffusion of liquids in nanopores: a nuclear magnetic resonance study. AB - Nuclear magnetic resonance has been applied to study the details of molecular motion of low-molecular-weight polar and nonpolar organic liquids in nanoporous silicon crystals of straight cylindrical pore morphology at different pore loadings. Effective self-diffusion coefficients as obtained using the pulsed field gradient nuclear magnetic resonance method were found to pass through a maximum with increasing concentration for all liquids under study. Taking account of a concentration-dependent coexistence of capillary condensed, adsorbed and gaseous phases a generalized model for the effective self-diffusion coefficient was developed and shown to satisfactorily explain the experimental results. An explicit use of the adsorption isotherm properties within the model extends its applicability to the mesoporous range and highlights the role of surface interaction for the transport of molecules in small pores. The problem of surface diffusion and diffusion of multilayered molecules is also addressed. PMID- 15268216 TI - Phase diagram of subphtalocyanine ordering on Ag(111). AB - A model of subphtalocyanine molecules ordering on Ag(111) is proposed. We have demonstrated that the driving force of the ordering into honeycomb and hexagonal close packed patterns is a balance of intermolecular and subphtalocyanine-Ag interactions which can be generally expressed by a potential with infinite exclusion in a sufficiently large number of nearest coordination spheres of Ag(111) lattice and oscillatingly decaying behavior outside the sphere of exclusion. To cope with computational problems due to large size of the molecules compared to the substrate lattice period, we introduce the rescaling of Ag(111) lattice, and took into account an infinite exclusion of first, second, and third neighbors, attraction-of fourth and fifth, and repulsion-of sixth and seventh. The phase diagram is obtained by the lattice gas model using Monte Carlo simulations. Very strong first order phase transitions, causing the two-phase coexistence, were found between disordered and honeycomb as well as between disordered and hexagonal closed packed phases. PMID- 15268217 TI - Molecular ordering and phase transitions in alkanol monolayers at the water hexane interface. AB - The interface between bulk water and bulk hexane solutions of n-alkanols (H(CH(2))(m)OH, where m=20, 22, 24, or 30) is studied with x-ray reflectivity, x ray off-specular diffuse scattering, and interfacial tension measurements. The alkanols adsorb to the interface to form a monolayer. The highest density, lowest temperature monolayers contain alkanol molecules with progressive disordering of the chain from the -CH(2)OH to the -CH(3) group. In the terminal half of the chain that includes the -CH(3) group the chain density is similar to that observed in bulk liquid alkanes just above their freezing temperature. The density in the alkanol headgroup region is 10% greater than either bulk water or the ordered headgroup region found in alkanol monolayers at the water-vapor interface. We conjecture that this higher density is a result of water penetration into the headgroup region of the disordered monolayer. A ratio of 1:3 water to alkanol molecules is consistent with our data. We also place an upper limit of one hexane to five or six alkanol molecules mixed into the alkyl chain region of the monolayer. In contrast, H(CH(2))(30)OH at the water-vapor interface forms a close-packed, ordered phase of nearly rigid rods. Interfacial tension measurements as a function of temperature reveal a phase transition at the water hexane interface with a significant change in interfacial excess entropy. This transition is between a low temperature interface that is nearly fully covered with alkanols to a higher temperature interface with a much lower density of alkanols. The transition for the shorter alkanols appears to be first order whereas the transition for the longer alkanols appears to be weakly first order or second order. The x-ray data are consistent with the presence of monolayer domains at the interface and determine the domain coverage (fraction of interface covered by alkanol domains) as a function of temperature. This temperature dependence is consistent with a theoretical model for a second order phase transition that accounts for the domain stabilization as a balance between line tension and long range dipole forces. Several aspects of our measurements indicate that the presence of domains represents the appearance of a spatially inhomogeneous phase rather than the coexistence of two homogeneous phases. PMID- 15268218 TI - Adsorption of 1-octanol at the free water surface as studied by Monte Carlo simulation. AB - The adsorption of 1-octanol at the free water surface has been investigated by Monte Carlo computer simulation. Six different systems, built up by an aqueous and a vapor phase, the latter also containing various number of octanol molecules, have been simulated. The number of the octanol molecules has been chosen in such a way that the octanol surface density varies in a broad range, between 0.27 and 7.83 micromol/m(2) in the six systems simulated. For reference, the interfacial system containing bulk liquid octanol in the apolar phase has also been simulated. The results have shown that the formation of hydrogen bonds between the interfacial water and adsorbed octanol molecules is of key importance in determining the properties of the adsorbed layer. At low octanol surface concentration values all the octanol molecules are strongly (i.e., by hydrogen bonds) bound to the aqueous phase, whereas their interaction with each other is negligibly small. Hence, they are preferentially oriented in such a way that their own binding energy (and thus their own free energy) is minimized. In this preferred orientation the O-H bond of the octanol molecule points flatly toward the aqueous phase, declining by about 30 degrees from the interfacial plane, irrespectively from whether the octanol molecule is the H-donor or the H-acceptor partner in the hydrogen bond. Hence, in its preferred orientation the octanol molecule can form at least two low energy hydrogen bonds with water: one as a H donor and another one as a H-acceptor. Moreover, the preferred orientation of the hydrogen bonded water partners is close to one of the two preferred interfacial water alignments, in which the plane of the water molecule is parallel with the interface. When increasing the octanol surface density, the water surface gets saturated with hydrogen bonded octanols, and hence any further octanol molecule can just simply condense to the layer of the adsorbed octanols. The surface density value at which this saturation occurs is estimated to be about 1.7 micromol/m(2). Above this surface density value the hydrogen bonded octanols and their water partners are oriented in such a way that the number of the water octanol hydrogen bonds is maximized. Hence, the preferred alignment of the O...O axes of these hydrogen bonds is perpendicular to the interface. This orientation is far from the optimal alignment of the individual octanol molecules, which is also reflected in the observed fact that, unlike in the case of many other adsorbents, the average molecular binding energy of the adsorbed octanol molecules increases (i.e., becomes less negative) with increasing octanol surface density. PMID- 15268219 TI - Adsorption and diffusion on a stepped surface: atomic hydrogen on Pt(211). AB - We present density functional theory calculations for atomic hydrogen interacting with a stepped surface, the Pt(211) surface. The calculations have been performed at the generalized gradient approximation level, using a slab representation of the surface. This is the state-of-the-art method for calculating the interaction of atoms or molecules with metal surfaces, nevertheless only few studies have used it to study atoms or molecules interacting with stepped surfaces, and none, to the best of our knowledge, have considered hydrogen interacting with stepped platinum surfaces. Our goal has been to initiate a systematic study of this topic. We have calculated the full three-dimensional potential energy surface (PES) for the H/Pt(211) system together with the vibrational band structure and vibrational eigenfunctions of H. A deep global minimum of the PES is found for bridge-bonded hydrogen on the step edge, in agreement with experimental results for the similar H/Pt(533) system. All the local vibrational excitations at the global minimum have been identified, and this will serve as a helpful guide to the interpretation of future experiments on this (or similar) system(s). Furthermore, from the calculated PES and vibrational band structure, we identify a number of consequences for the interpretation or modelling of diffusion experiments studying the coverage and directional dependence of atomic hydrogen diffusion on stepped platinum surfaces. PMID- 15268220 TI - Confinement effect on the adsorption from a binary liquid system near liquid/liquid phase separation. AB - The preferential adsorption of one component of a binary system at the inner surfaces of mesoporous silica glasses was studied in a wide composition range at temperatures close to liquid/liquid phase separation. Confinement effects on the adsorption were investigated by using three controlled-pore glass (CPG-10) materials of different mean pore size (10 to 50 nm). For the experimental system (2-butoxyethanol+water), which exhibits an upper miscibility gap, strong preferential adsorption of water occurs, as the coexistence curve is approached at bulk compositions, at which water is the minority component. In this strong adsorption regime the area-related surface excess amount of adsorbed water decreases with decreasing pore width, while the shift in the volume-related mean composition of the pore liquid shows an opposite trend, i.e., greatest deviation from bulk composition occurring in the most narrow pores. A simple mean-field lattice model of a liquid mixture confined by parallel walls is adopted to rationalize these experimental findings. This model reproduces the main findings of the confinement effect on the adsorption near liquid/liquid phase separation. PMID- 15268221 TI - Exploration of the electronic structure of dendrimerlike acetylene-bridged oligothiophenes by correlating Raman spectroscopy, electrochemistry, and theory. AB - A series of radial thiophene-based structures consisting of a central benzene or thiophene ring surrounded by acetylene-bridged terthienyl arms has been investigated by physical and theoretical methods. Fourier transform Raman spectroscopy of the neutral solids shows that the terthiophene arms are weakly coupled across the core (benzene plus acetylene groups) likely due to cross conjugation or meta-conjugation effects that may prevent full delocalization. By increasing the number of arms around the central ring, the electronic structure of the molecules seems to be affected only at the core, whereas the outer terthiophene arms remain almost unaltered. Raman spectroelectrochemistry and quantum chemical calculations provide further insight into the charge delocalization of the oxidized species. There is no evidence to suggest that these oxidized forms, obtained upon electrochemical doping of the molecules, show charge delocalization across the core. PMID- 15268222 TI - Adsorption, diffusion, and recombination of hydrogen on pure and boron-doped graphite surfaces. AB - Boron inserted as impurity by substitution of carbon atoms in graphite is known to modify the reactivity of the surface in interaction with hydrogen. Boron induces a better H retention capability in graphite while it makes easier the recombination into molecular hydrogen under heating in thermal-desorption experimental conditions. It has already been calculated that boron modifies the electronic structure of the surface, which results in an increase of the adsorption energy for H. This result seems in good agreement with the better retention for H in doped graphite, but contradictory with the easier recombination observed. The aim of this work is to dismiss this contradiction by elucidating the modifications induced by boron in the recombination mechanism. We studied the diffusion of H on pure and boron-doped graphite in the density functional theory framework. We determined a diffusionlike mechanism leading to molecular hydrogen formation. Finally, we have shown the fundamental modifications induced by boron on the [0001] graphite surface reactivity. From these calculations it stands out that recombination is the result of desorption on pure graphite and diffusion on B-doped surfaces, while the activation energy for the rate limiting step is half reduced by boron. The results are compared to experimental observations. The connection between the cluster and periodic quantum modes for graphite is also discussed. PMID- 15268223 TI - Resonance Raman contribution to the D band of carbon materials: modeling defects with quantum chemistry. AB - Polycyclic aromatic hydrocarbons (PAHs) are employed to model the Raman features that are generally associated with sp(2) nanostructures in carbon materials or with disorder and defects in graphitic materials. To this end molecular parameters (geometry changes upon electronic excitation, vibrational normal modes, and displacement parameters) are computed with semiempirical quantum chemical methods for a series of PAHs ranging from 6 to 384 carbon atoms, and Raman intensities are evaluated according to Albrecht's formalism restricted to the A term. The computed preresonance and resonance Raman intensities are compared with available experimental data for hexa-peri-hexabenzocoronene and for pyrene. For the latter compound, simulations carried out at semiempirical and ab initio levels of theory are shown to be of comparable quality. Finally, the collection of displacement parameters computed for the sample of conjugated molecules is used to model the effect of disorder and defects in the Raman response of a carbon material containing sp(2) islands. It is shown that the computed D-band frequency dispersion, with respect to excitation wavelength, reproduces closely the experimental data measured for sp(2) hybridized carbon materials. PMID- 15268224 TI - Magnetic enhancement and magnetic reduction in binary clusters of transition metal atoms. AB - Electronic and magnetic properties of small binary clusters containing one or two transition metal atoms are investigated using ab initio calculations with a view to explain the experimentally observed magnetic enhancement/reduction in these systems. As the present investigations do not rely on spin-orbit effects, our results reveal the enhancement or reduction in the magnetic moment to depend on two main factors; namely geometry and, most importantly, the d-band filling. The results can be used as a guide in the experimental synthesis of high density magnetic grains. PMID- 15268225 TI - Radical initiated polymerization in a bifunctional mixture via computer simulation. AB - Computer simulations are performed to study the polymerization behavior in a mixture of bifunctional groups such as olefins (A) and acrylates (B) in an effective solvent (a coarse description for vegetable oil derived macromonomers (VOMMs) in solution) on a cubic lattice. A set of interactions between these units and solvent (S) constituents and their relative concentrations (p(A), p(B), and p(S)) are considered. Samples are equilibrated with Metropolis algorithm to model the perceived behavior of VOMMs. The covalent bonding between monomeric units is then implemented via reaction pathways initiated by stochastic motion of free radicals (a very small fraction). The rate of reaction shows decay patterns with the time steps (t) with power laws (i.e., R(ab)alphat(-r), r congruent with 0.4-0.8), exponential decays (i.e., R(ab)alphae(-0.001t)), and their combination. Growth of A-B bonding is studied as a function of polymer concentration p=p(A)+p(B) for four different model systems appropriate for VOMMs. The data from the free radical initiated simulations are compared to the original simulations with homopolymerization. While most of the data are consistent with experimental observations, the variations are found to be model dependent. PMID- 15268226 TI - Quantum chemical ab initio calculations of correlation effects in complex polymers: poly(para-phenylene). AB - Different quantum chemical approaches to the ground state correlation energy per unit cell of infinite poly(para-phenylene) (PPP) chains are presented. PPP is an organic polymer with interesting optical properties, due to its conjugated, aromatic pi system. The inclusion of correlation effects is crucial for a sound quantum chemical description of such a system. The correlation calculations were performed on the coupled cluster with single and double excitations (CCSD) level of theory using Dunning's spd correlation consistent polarized valence double zeta basis sets. The correlation energy per unit cell is determined by means of the incremental method, which comprises series of CCSD calculations with partial excitation spaces. The resulting correlation energy per unit cell of PPP is 21.797 eV and compares well with that obtained by a simple but much more demanding cluster convergence approach (-21.775 eV). In addition, the accuracy and performance of the incremental scheme is discussed with respect to full CCSD benchmark calculations on PPP oligomers. Two variants are considered, the conventional one based on bond-type local units, and an extended one based on natural chemical subunits. Whereas it is difficult to reach "chemical" accuracy with the first variant, the second variant allows an accurate and efficient treatment with only a few individual CCSD calculations for a polymer with an aromatic pi system such as PPP. PMID- 15268227 TI - Accelerated molecular dynamics: a promising and efficient simulation method for biomolecules. AB - Many interesting dynamic properties of biological molecules cannot be simulated directly using molecular dynamics because of nanosecond time scale limitations. These systems are trapped in potential energy minima with high free energy barriers for large numbers of computational steps. The dynamic evolution of many molecular systems occurs through a series of rare events as the system moves from one potential energy basin to another. Therefore, we have proposed a robust bias potential function that can be used in an efficient accelerated molecular dynamics approach to simulate the transition of high energy barriers without any advance knowledge of the location of either the potential energy wells or saddle points. In this method, the potential energy landscape is altered by adding a bias potential to the true potential such that the escape rates from potential wells are enhanced, which accelerates and extends the time scale in molecular dynamics simulations. Our definition of the bias potential echoes the underlying shape of the potential energy landscape on the modified surface, thus allowing for the potential energy minima to be well defined, and hence properly sampled during the simulation. We have shown that our approach, which can be extended to biomolecules, samples the conformational space more efficiently than normal molecular dynamics simulations, and converges to the correct canonical distribution. PMID- 15268228 TI - Thermoreversible crosslinking of polyelectrolyte chains. AB - Thermoreversible crosslinking of polyelectrolyte chains via short-range attractions such as hydrogen bonding induced by uncharged or charged particles is studied within the Flory model of ideal association. Electrostatic interactions between the charges at different linking fractions are taken into account by using a generalized random phase approximation approach which includes the network connectivity. We find that at certain concentration of linking agents an infinitely large polymer network is formed. We calculate the structural gelation lines for linkers of different charges and functionalities. PMID- 15268229 TI - Electrostatic interactions of charged dipolar proteins in reverse micelles. AB - The electrostatic interactions in a reverse micelle containing a small-ionized protein are studied by Monte Carlo simulation. The electrostatic contribution to the potential of mean force of the protein in the reverse micelle is determined for a neutral protein, a uniformly charged protein, and a uniformly charged protein with a dipole moment. The effect of addition of a simple electrolyte is studied. While symmetrically distributed micellar charge exerts no force on enclosed ionic species, the protein is driven to the micellar wall due to interactions with simple ions. Protein binding to the inner wall of the micelle can be regulated by added salt. The presence of a dipole drives the protein further to the wall. These effects are studied for several proteins characterized by different charges and dipole moments. For a weakly charged protein with a strong dipole moment the contribution of dipolar interaction to the free energy can represent a major driving force for protein solubilization in the microemulsion. PMID- 15268230 TI - The role of pressure in rubber elasticity. AB - We describe a series of molecular dynamics computations that reveal an intimate connection at the atomic scale between difference stress (which resists stretches) and pressure (which resists volume changes) in an idealized elastomer, in contrast to the classical theory of rubber elasticity. Our simulations idealize the elastomer as a "pearl necklace," in which the covalent bonds are stiff linear springs, while nonbonded atoms interact through a Lennard-Jones potential with energy epsilon(LJ) and radius sigma(LJ). We calculate the difference stress t(11)-(t(22)+t(33))/2 and mean stress (t(11)+t(22)+t(33))/3 induced by a constant volume extension in the x(1) direction, as a function of temperature T and reduced density rho(*)=Nsigma(IJ) (3)/nu. Here, N is the number of atoms in the simulation cell and nu is the cell volume. Results show that for rho(*)<1, the difference stress is purely entropic and is in good agreement with the classical affine network model of rubber elasticity, which neglects nonbonded interactions. However, data presented by van Krevelen [Properties of Polymers, 3rd ed. (Elsevier, Amsterdam, 1990), p. 79] indicate that rubber at standard conditions corresponds to rho(*)=1.2. For rho(*)>1, the system is entropic for kT/epsilon(LJ)>2, but at lower temperatures the difference stress contains an additional energy component, which increases as rho(*) increases and temperature decreases. Finally, the model exhibits a glass transition for rho(*)=1.2 and kT/epsilon(LJ) approximately 2. The atomic-scale processes responsible for generating stress are explored in detail. Simulations demonstrate that the repulsive portion of the Lennard-Jones potential provides a contribution sigma(nbr)>0 to the difference stress, the attractive portion provides sigma(nba) approximately 0, while the covalent bonds provide sigma(b)<0. In contrast, their respective contributions to the mean stress satisfy Pi(nbr)<0, Pi(nba)>0, and Pi(b)<0. Analytical calculations, together with simulations, demonstrate that mean and difference stresses are related by sigma(nbr)=-APi(nbr)P(2)(theta(b)), sigma(b)=BPi(b)P(2)(theta(b)), where P(2)(theta(b)) is a measure of the anisotropy of the orientation of the covalent bonds, and A and B are coefficients that depend weakly on rho(*) and temperature. For high values of rho(*), we find that [sigma(nbr)]>>[sigma(b)], and in this regime our model predicts behavior that is in good agreement with experimental data of D.L. Quested et al. [J. Appl. Phys. 52, 5977 (1981)] for the influence of pressure on the difference stress induced by stretching solithane. PMID- 15268233 TI - Quantum mechanical map for protein-ligand binding with application to beta trypsin/benzamidine complex. AB - We report full ab initio Hartree-Fock calculation to compute quantum mechanical interaction energies for beta-trypsin/benzamidine binding complex. In this study, the full quantum mechanical ab initio energy calculation for the entire protein complex with 3238 atoms is made possible by using a recently developed MFCC (molecular fractionation with conjugate caps) approach in which the protein molecule is decomposed into amino acid-based fragments that are properly capped. The present MFCC ab initio calculation enables us to obtain an "interaction spectrum" that provides detailed quantitative information on protein-ligand binding at the amino acid levels. These detailed information on individual residue-ligand interaction gives a quantitative molecular insight into our understanding of protein-ligand binding and provides a guidance to rational design of potential inhibitors of protein targets. PMID- 15268234 TI - Isotropic-liquid crystalline phase diagram of a CdSe nanorod solution. AB - We report the isotropic-liquid crystalline phase diagram of 3.0 nm x 60 nm CdSe nanorods dispersed in anhydrous cyclohexane. The coexistence concentrations of both phases are found to be lower and the biphasic region wider than the results predicted by the hard rod model, indicating that the attractive interaction between the nanorods may be important in the formation of the liquid crystalline phase in this system. PMID- 15268235 TI - Rules for minimal atomic multipole expansion of molecular fields. AB - A nonempirical minimal atomic multipole expansion (MAME) defines atomic charges or higher multipoles that reproduce electrostatic potential outside molecules. MAME eliminates problems associated with redundancy and with statistical sampling, and produces atomic multipoles in line with chemical intuition. PMID- 15268236 TI - Capillary force in atomic force microscopy. AB - Under ambient conditions, a water meniscus generally forms between a nanoscale atomic force microscope tip and a hydrophilic surface. Using a lattice gas model for water and thermodynamic integration methods, we calculate the capillary force due to the water meniscus for both hydrophobic and hydrophilic tips at various humidities. As humidity rises, the pull-off force rapidly reaches a plateau value for a hydrophobic tip but monotonically increases for a weakly hydrophilic tip. For a strongly hydrophilic tip, the force increases at low humidities (<30%) and then decreases. We show that mean-field density functional theory reproduces the simulated pull-off force very well. PMID- 15268237 TI - Full S matrix calculation via a single real-symmetric Lanczos recursion: the Lanczos artificial boundary inhomogeneity method. AB - We present an efficient and robust method for the calculation of all S matrix elements (elastic, inelastic, and reactive) over an arbitrary energy range from a single real-symmetric Lanczos recursion. Our new method transforms the fundamental equations associated with Light's artificial boundary inhomogeneity approach from the primary representation (original grid or basis representation of the Hamiltonian or its function) into a single tridiagonal Lanczos representation, thereby affording an iterative version of the original algorithm with greatly superior scaling properties. The method has important advantages over existing iterative quantum dynamical scattering methods: (a) the numerically intensive matrix propagation proceeds with real symmetric algebra, which is inherently more stable than its complex symmetric counterpart; (b) no complex absorbing potential or real damping operator is required, saving much of the exterior grid space which is commonly needed to support these operators and also removing the associated parameter dependence. Test calculations are presented for the collinear H+H(2) reaction, revealing excellent performance characteristics. PMID- 15268238 TI - Vibrational mode selectivity in hyperfine interactions: polarization quantum beat spectroscopy of HCF(A1A"). AB - We report on the vibrational mode dependence of the 19F and 1H hyperfine interaction constants in the A1A" state of HCF, determined using polarization quantum beat spectroscopy. The nuclear spin/overall rotation coupling constants display a pronounced energy dependence and mode selectivity which can be traced to variations in both the A rotational constant and nuclear spin/electron orbital coupling constant a. In particular, modes containing C-F stretching excitation display significantly larger 19F spin-rotation constants, which is explained in terms of a decrease in back donation of electron density into the C 2p(pi) orbitals. PMID- 15268239 TI - Infrared matrix-isolation spectroscopy using pulsed deposition of p-H2. AB - We employed pulsed deposition of p-H2 onto a cold target to form a matrix sample suitable for measurements of infrared absorption. Unlike the method of rapid vapor deposition at approximately 2.5 K, developed by Fajardo et al., this method can be performed at a temperature as high as 5.5 K, achievable with a closed cycle refrigerator; pumping on liquid helium in a cryostat is eliminated. Compared with the enclosed-cell method developed by Oka, Shida, Momose, and co workers, this method is more versatile in sample preparation, especially for samples at a greater concentration or with high reactivity. Two experiments were tested: the pulse-deposited sample of CH4/p-H2 yields an infrared absorption spectrum nearly identical to that recorded with rapid vapor deposition, and a sample of vinyl chloride (C2H3Cl) in solid p-H2 irradiated with laser emission at 193 nm yields C2H5, in contrast to formation of HCl, C2H2, and a complex of HClC2H2 observed upon photolysis of C2H3Cl in an Ar matrix. These experiments are also compared with those with n-H2 or Ne as the matrix host. PMID- 15268240 TI - Theory of time-resolved photoelectron imaging. Comparison of a density functional with a time-dependent density functional approach. AB - Time-resolved photoelectron differential cross sections are computed within a quantum dynamical theory that combines a formally exact solution of the nuclear dynamics with density functional theory (DFT)-based approximations of the electronic dynamics. Various observables of time-resolved photoelectron imaging techniques are computed at the Kohn-Sham and at the time-dependent DFT levels. Comparison of the results serves to assess the reliability of the former method and hence its usefulness as an economic approach for time-domain photoelectron cross section calculations, that is applicable to complex polyatomic systems. Analysis of the matrix elements that contain the electronic dynamics provides insight into a previously unexplored aspect of femtosecond-resolved photoelectron imaging. PMID- 15268241 TI - Energy conserving approximations to the quantum potential: dynamics with linearized quantum force. AB - Solution of the Schrodinger equation within the de Broglie-Bohm formulation is based on propagation of trajectories in the presence of a nonlocal quantum potential. We present a new strategy for defining approximate quantum potentials within a restricted trial function by performing the optimal fit to the log derivatives of the wave function density. This procedure results in the energy conserving dynamics for a closed system. For one particular form of the trial function leading to the linear quantum force, the optimization problem is solved analytically in terms of the first and second moments of the weighted trajectory distribution. This approach gives exact time-evolution of a correlated Gaussian wave function in a locally quadratic potential. The method is computationally cheap in many dimensions, conserves total energy and satisfies the criterion on the average quantum force. Expectation values are readily found by summing over trajectory weights. Efficient extraction of the phase-dependent quantities is discussed. We illustrate the efficiency and accuracy of the linear quantum force approximation by examining a one-dimensional scattering problem and by computing the wavepacket reaction probability for the hydrogen exchange reaction and the photodissociation spectrum of ICN in two dimensions. PMID- 15268242 TI - Wavelet treatment of structure and thermodynamics of simple liquids. AB - A new algorithm is developed to solve integral equations for simple liquids. The algorithm is based on the discrete wavelet transform of radial distribution functions. The Coifman 2 basis set is employed for the wavelet treatment. To solve integral equations we have applied the combined scheme in which the coarse part of the solution is calculated by wavelets, while the fine part by the direct iterations. Tests on the PY and HNC approximations have indicated that the proposed procedure is more effective than the conventional method based on the hybrid algorithm. Possibilities for application of the method to molecular liquids and mixed quantum-classical systems are discussed. PMID- 15268243 TI - Predicting shielding constants in solution using gauge invariant atomic orbital theory and the effective fragment potential method. AB - A method to approximate ab initio shielding constants is presented, in which the ab initio density matrix is replaced in the gauge invariant atomic orbital formalism with the density matrix resulting from an effective fragment potential calculation. The resulting first-order density matrix is then iterated to self consistency. The method is compared with fully ab initio gauge invariant atomic orbital restricted Hartree-Fock calculations on hydrogen chloride, water, and ammonia solutes with up to nine solvent water molecules using the 6-31G, 6 31G(d,p), and 6-31+G(d,p) basis sets. Using the 6-31G(d,p) basis sets, the average of the average absolute deviations for the three environments tested is 0.34 ppm. This is sufficiently accurate to allow for the identification of specific (1)H nuclei in a solvated molecule when the chemical shift between nuclei is not less than 1 ppm. The success of the method at this level of approximation is due to a cancellation of errors between the paramagnetic and diamagnetic terms of the shielding constant: the diamagnetic term is underestimated by roughly the same amount that the paramagnetic term is overestimated. PMID- 15268244 TI - Ab initio torsional potential and transition frequencies of acetaldehyde. AB - High-level ab initio electronic structure calculations, including extrapolations to the complete basis set limit as well as relativistic and diagonal Born Oppenheimer corrections, resulted in a torsional potential of acetaldehyde in its electronic ground state. This benchmark-quality potential fully reflects the symmetry and internal rotation dynamics of this molecule in the energy range probed by spectroscopic experiments in the infrared and microwave regions. The torsional transition frequencies calculated from this potential and the ab initio torsional inverse effective mass function are within 2 cm(-1) of the available experimental values. Furthermore, the computed contortional parameter rho of the rho-axis system Hamiltonian is also in excellent agreement with that obtained from spectral analyses of acetaldehyde. PMID- 15268245 TI - A theory of nonvertical triplet energy transfer in terms of accurate potential energy surfaces: the transfer reaction from pi,pi* triplet donors to 1,3,5,7 cyclooctatetraene. AB - Triplet energy transfer (TET) from aromatic donors to 1,3,5,7-cyclooctatetraene (COT) is an extreme case of "nonvertical" behavior, where the transfer rate for low-energy donors is considerably faster than that predicted for a thermally activated (Arrhenius) process. To explain the anomalous TET of COT and other molecules, a new theoretical model based on transition state theory for nonadiabatic processes is proposed here, which makes use of the adiabatic potential energy surfaces (PES) of reactants and products, as computed from high level quantum mechanical methods, and a nonadiabatic transfer rate constant. It is shown that the rate of transfer depends on a geometrical distortion parameter gamma=(2g(2)/kappa(1))(1/2) in which g stands for the norm of the energy gradient in the PES of the acceptor triplet state and kappa(1) is a combination of vibrational force constants of the ground-state acceptor in the gradient direction. The application of the model to existing experimental data for the triplet energy transfer reaction to COT from a series of pi,pi(*) triplet donors, provides a detailed interpretation of the parameters that determine the transfer rate constant. In addition, the model shows that the observed decrease of the acceptor electronic excitation energy is due to thermal activation of C=C bond stretchings and C-C bond torsions, which collectively change the ground-state COT bent conformation (D(2d)) toward a planar triplet state (D(8h)). PMID- 15268246 TI - DFT:B3LYP ab initio molecular dynamics study of the Zundel and Eigen proton complexes, H5O2+ and H9O4+, in the triplet state in gas phase and solution. AB - DFT:B3LYP ab initio molecular dynamics (MD) approach is used to elucidate the properties of the Zundel and Eigen, H5O2+ and H9O4+, proton complexes in the triplet state. The simulation considers the complexes in the gas phase (isolated complexes) and inside the clusters composed of 32, 64, and 128 water molecules, mimicking the behavior of aqueous solutions. MD simulations reveal three distinct periods. For the complex in solutions, the periods are smoothed out. The H5O2+ and H9O4+ complexes in the triplet state undergo structural rearrangements, which eventually result in hydrogen elimination. For the H5O2+, the hydrogen is eliminated from the center of the water cluster, whereas for the H9O4+ it is removed from a near-surface water molecule. The rate of hydrogen elimination decreases with increasing the number of water molecules surrounding the complex. PMID- 15268247 TI - A concerted three-body formation X+Y+C2H4 in the photodissociation of CH2XCH2Y (X,Y=Br,Cl) at 193 nm. AB - The photodissociation of CH2XCH2Y (X,Y=Br,Cl) through absorption of 193 nm photons was investigated using product translational spectroscopy. No stable CH2BrCH2 or CH2ClCH2 was detected. The recorded time-of-flight spectra indicate that these internally excited radicals dissociated into Y+C2H4 in a concerted reaction with the first C-X bond rupture. Product anisotropy implies that the overall reaction time for three-body formation is in a fraction of rotational period. According to an asynchronous concerted reaction model, the measured spectra were simulated with product translational energy distributions coupled by asymmetric angular distributions. For the mixed halide, CH2BrCH2Cl, triple products Br+Cl+C2H4 can be originated from the cleavage of either the C-Br bond or the C-Cl bond. The results are discussed and where appropriate, comparisons with previous investigations of the related molecules are included. PMID- 15268248 TI - Quantum wave packet and quasiclassical trajectory studies of OH+CO: influence of the reactant channel well on thermal rate constants. AB - We study the OH+CO-->H+CO2 reaction with both six-dimensional quantum wave packets (QM) and quasiclassical trajectories (QCT), determining reaction probabilities and thermal rate constants (or coefficients), and studying the influence of the reactant channel hydrogen-bonded complex well on the reaction dynamics. The calculations use the recently developed Lakin-Troya-Schatz-Harding (LTSH) ground electronic state potential energy surface, along with a modified surface developed for this study (mod-LTSH), in which the reactant channel well is removed. Our results show that there can be significant differences between the QM and QCT descriptions of the reaction for ground-state reactants and for energies important to the thermal rate constants. Zero-point energy violation plays an important role in the QCT results, and as a result, the QCT reaction probability (for ground-state reactants and zero impact parameter) is much higher than its QM counterpart at moderate to low reagent translational energies. The influence of the reactant channel well in the QCT results is to enhance reactivity at moderate energies and to suppress reactivity at the very lowest collision energies. The QM results also show the enhancement at moderate energies but, while the very lowest translational energies cannot be adequately converged, they do not indicate any tendency toward suppression as energy is reduced. QCT calculations for excited rotational states of the reactants show that the suppression of reactivity associated with the reactant channel well is less important when the reactants are rotating, and as a result, the influence of the reactant channel well on the thermal rate coefficients is relatively small, being important below 200 K. Our results indicate that there still remain important discrepancies between experiment and theory in this low temperature regime and that further improvements of the potential are needed. PMID- 15268249 TI - Laser-induced fluorescence spectroscopy of 3,4,9,10-perylenetetracarboxylic dianhydrid in helium nanodroplets. AB - Electronic spectra of the S1<--S0 transition of the 3,4,9,10 perylenetetracarboxylic-dianhydrid (PTCDA) monomer isolated in superfluid helium nanodroplets have been measured by means of laser-induced fluorescence. The 0(0)(0) transition appears at 20,988 cm(-1) as the dominant line. We obtain clearly resolved the vibrational structure of the molecule. A comparison to Raman spectra of PTCDA films on metallic substrates and PTCDA crystals as well as with calculated frequencies provides the identification of the different modes. The enhanced resolution in the low temperature helium environment and the obtained line positions provide new information about structural properties of perylene derivatives. PMID- 15268250 TI - CASSCF and CASPT2 studies on the structures, transition energies, and dipole moments of ground and excited states for azulene. AB - The electronic structure of azulene molecule has been studied. We have obtained the optimized structures of ground and singlet excited states by using the complete active space self-consistent-field (CASSCF) method, and calculated vertical and 0-0 transition energies between the ground and excited states with second-order Moller-Plesset perturbation theory (CASPT2). The CASPT2 calculations indicate that the bond-equalized C(2v) structure is more stable than the bond alternating C(s) structure in the ground state. For a physical understanding of electronic structure change from C(2v) to C(s), we have performed the CASSCF calculations of Duschinsky matrix describing mixing of the b(2) vibrational mode between the ground (1A(1)) and the first excited (1B(2)) states based on the Kekule-crossing model. The CASPT2 0-0 transition energies are in fairly good agreement with experimental results within 0.1-0.3 eV. The CASSCF oscillator strengths between the ground and excited states are calculated and compared with experimental data. Furthermore, we have calculated the CASPT2 dipole moments of ground and excited states, which show good agreement with experimental values. PMID- 15268251 TI - Activation barriers and rate constants for hydration of platinum and palladium square-planar complexes: an ab initio study. AB - In the present work, an ab initio study on hydration (a metal-ligand replacement by water molecule or OH- group) of cis- and transplatin and their palladium analogs was performed within a neutral pseudomolecule approach (e.g., metal complex+water as reactant complex). Subsequent replacement of the second ligand was considered. Optimizations were performed at the MP2/6-31+G(d) level with single-point energy evaluation using the CCSD(T)/6-31++G(d,p) approach. For the obtained structures of reactants, transition states (TS's), and products, both thermodynamic (reaction energies and Gibbs energies) and kinetic (rate constants) characteristics were estimated. It was found that all the hydration processes are mildly endothermic reactions-in the first step they require 8.7 and 10.2 kcal/mol for ammonium and chloride replacement in cisplatin and 13.8 and 17.8 kcal/mol in the transplatin case, respectively. Corresponding energies for cispalladium amount to 5.2 and 9.8 kcal/mol, and 11.0 and 17.7 kcal/mol for transpalladium. Based on vibrational analyses at MP2/6-31+G(d) level, transition state theory rate constants were computed for all the hydration reactions. A qualitative agreement between the predicted and known experimental data was achieved. It was also found that the close similarities in reaction thermodynamics of both Pd(II) and Pt(II) complexes (average difference for all the hydration reactions are approximately 1.8 kcal/mol) do not correspond to the TS characteristics. The TS energies for examined Pd(II) complexes are about 9.7 kcal/mol lower in comparison with the Pt analogs. This leads to 10(6) times faster reaction course in the Pd cases. This is by 1 or 2 orders of magnitude more than the results based on experimental measurements. PMID- 15268252 TI - Zero kinetic energy photoelectron study of SO2+(X2A1) using coherent extreme ultraviolet radiation. AB - Using our newly built extreme ultraviolet (XUV) photoelectron and photoion spectrometer, we have obtained the pulsed field ionization zero kinetic energy (ZEKE) photoelectron spectra of SO2+(X2A1)<--SO2(X1A1) by coherent XUV radiation in the energy range of 12.29-12.82 eV. The adiabatic ionization potential (IP) of SO2 is 12.3458+/-0.0002 (eV), which was determined by comparing the partially resolved rotational branch contour with the simulated one. Besides the bending vibrational mode (upsilon2) which was found to be exclusive in the photoelectron spectra (PE) reported previously, we also observed the other two modes: the symmetric stretching (upsilon1) and the antisymmetric stretching (upsilon3) vibrations. The fundamental of the symmetric stretching (upsilon(1)) is 1057 cm( 1) and the overtone of the antisymmetric stretching (2upsilon(3)) is 2494 cm(-1). The new vibrational progressions (upsilon(1)00)+, (1upsilon(2)0)+, (2upsilon(2)0)+, and (0upsilon(2)2)+ have also been observed, and these new observations suggested that the irregular structure of (0upsilon(2)0)+ assigned to the previous PE spectra should be reconsidered. The comparison of the intensities of these vibrational bands with the calculated Franck-Condon factors with harmonic approximation was also made. PMID- 15268253 TI - Nonadiabatic interactions in wave packet dynamics of the bromoacetyl chloride photodissociation. AB - The competitive photodissociation of bromoacetyl chloride BrCH2COCl in the first 1A" state (S1) by 248 nm photons is investigated by nonadiabatic wave packet simulations. We show that the preferential breaking of the stronger C-Cl bond (alpha to the excited carbonyl) over the weaker C-Br bond (beta) could be explained by a diabatic trapping or nonadiabatic recrossing as previously proposed. Our energy resolved flux analysis agrees fairly well with the experimental branching ratio (C-Cl:C-Br=1.0:0.4). Even if this does not prove the mechanism, this at least prevents to discard it. A reduced dimensionality approach based on constrained Hamiltonian is used. The nonadiabatic dissociation is studied in the two C-O/C-X (X=Br, Cl) subspaces to emphasize the role of the C O vibration upon [nO-->piCO*] excitation. The internal torsion and wagging dihedral angles are frozen at their Franck-Condon value, according to preliminary dynamical tests. The other inactive coordinates are optimized at the trans and Cs constrained geometry in the first excited state. Corresponding 2D cuts in the potential energy surfaces have been computed at the CASSCF level. The nonadiabatic kinetic couplings are highly peaked along an avoided crossing seam in both cases. A two-state diabatic model with a constant potential coupling is proposed in the two C-O/C-X subspaces. The inclusion of the C-O stretching in the active coordinates improves the value of the branching ratio over our previous 1D computation. PMID- 15268254 TI - The dissociation adiabaticity parameter and the strong field dissociation of HCl+. AB - In earlier work on H2+, we showed how a dissociation adiabaticity parameter, gammaDv identical with (Dv/2Upm)(1/2) (Dv is the dissociation energy from vibrational state v and U(pm) is the molecular ion system's ponderomotive energy), proposed by Walsh et al., can be modified and be a useful indicator of the strong field dissociation regime for a homonuclear diatomic. In the case of H2+, the new adiabaticity parameter, gamma(mol), indicates when a dissociation process can be most easily described as multiphoton above-threshold dissociation (gamma(mol)>1) and when it is better described using barrier-suppressed dissociation (gamma(mol)<1). In the case of a heteronuclear diatomic like HCl+, different electronic states can lead to different dissociation product channels to which are ascribed different gamma(mol) values. We show for a wide range of laser wavelengths and intensities that this adiabaticity parameter successfully predicts the type of dissociation dynamics (multiphoton above-threshold dissociation versus barrier-suppressed dissociation) on each electronic potential curve. We also discover that the dynamics in one electronic state can influence the dynamics in another at the same laser wavelengths and intensities, overriding the predictive capability of an adiabaticity parameter defined for a particular electronic state. Reasonable physical explanations are provided for these overriding cases. PMID- 15268255 TI - Franck-Condon simulation of the single vibronic level emission spectra of HSiF and DSiF including anharmonicity. AB - Potential energy functions (PEFs) of the X (1)A(') and A (1)A(") states of HSiF have been computed using the coupled-cluster single-double plus perturbative triple excitations and complete-active-space self-consistent-field multireference internally contracted configuration interaction methods, respectively, employing augmented correlation-consistent polarized-valence quadruple-zeta basis sets. For both electronic states of HSiF and DSiF, anharmonic vibrational wavefunctions and energies of all three modes have been calculated variationally with the ab initio PEFs and using Watson's Hamiltonian for nonlinear molecules. Franck-Condon factors between the two electronic states, allowing for Duschinsky rotation, were computed using the calculated anharmonic vibrational wavefunctions. These Franck Condon factors were used to simulate the single vibronic level (SVL) emission spectra recently reported by Hostutler et al. in J. Chem. Phys. 114, 10728 (2001). Excellent agreement between the simulated and observed spectra was obtained for the A (1)A(")(1,0,0)-->X (1)A(') SVL emission of HSiF. Discrepancies between the simulated and observed spectra of the A (1)A(")(0,1,0) and (1,1,0) SVL emissions of HSiF have been found. These are most likely, partly due to experimental deficiencies and, partly to inadequacies in the ab initio levels of theory employed in the calculation of the PEFs. Based on the computed Franck Condon factors, minor revisions of previous vibrational assignments are suggested. The calculated anharmonic wave functions of higher vibrational levels of the X (1)A(') state show strong mixings between the three vibrational modes of HSi stretching, bending, and SiF stretching. PMID- 15268256 TI - A study of the mode-selective trans--cis isomerization in HONO using ab initio methodology. AB - Ab initio calculations on the six-dimensional cis--trans double minimum potential energy surface of the electronic ground state of the HONO molecule were performed using a coupled cluster approach. An analytic fit to the data points was established. The interconversion barrier was calculated to be 4105 cm(-1). The nuclear motion problem was solved variationally using a full six-dimensional Hamiltonian in internal coordinates. The eigenstates up to about 3650 cm(-1) were tentatively assigned by harmonic quantum numbers. The assignment was based on the mean values of the internal coordinates of the six-dimensional eigenfunctions and on a comparison of the eigenenergies with those calculated by second-order perturbation theory from a full quartic force field in dimensionless normal coordinates. In cold matrices the trans- and the cis-OH nu(1) stretching modes and the first trans- and cis-NO 2nu(2) stretching overtones lead to isomerization. In the isolated molecule these modes (J=0) were found to be entirely localized. However, several overtones of the nu(4) ONO bending and nu(5) N-O stretching, which are close in energy to the OH stretch and combined with the torsional mode, were found to be strongly cis-trans delocalized. PMID- 15268257 TI - Improved LeRoy-Bernstein near-dissociation expansion formula, and prospect for photoassociation spectroscopy. AB - NDE (Near-dissociation expansion) including LeRoy-Bernstein formulas are improved by taking into account the multipole expansion coefficients and the nonasymptotic part of the potential curve. The theory is tested with the Rydberg-Klein-Rees (RKR) potential curve of the Cs(2)(0(g) (-)6s+6p(3/2)) state. Results indicate that the formula could be used to improve the determination of the asymptotic coefficient (within a 1% accuracy) and to extract relativistic correction from photoassociation spectra of long-range potential curve of diatomic molecules. PMID- 15268258 TI - An ab initio theoretical prediction: an antiaromatic ring pi-dihydrogen bond accompanied by two secondary interactions in a "wheel with a pair of pedals" shaped complex FH . . . C4H4 . . . HF. AB - By the counterpoise-correlated potential energy surface method (interaction energy optimization), the structure of the pi H-bond complex FH cdots, three dots, centered FH . . . C4H4 . . . HF has been obtained at the second-order Moller-Plesset perturbation theory (MP2/aug-cc-pVDZ) level. Intermolecular interaction energy of the complex is calculated to be -7.8 kcal/mol at the coupled-cluster theory with single, double substitutions and perturbatively linked triple excitations CCSD (T)/aug-cc-pVDZ level. The optimized structure is a "wheel with a pair of pedals" shaped (1mid R:1) structure in which both HF molecules almost lie on either vertical line passing through the middle-point of the C[Double Bond]C bond on either side of the horizontal plane of the C4 ring for cyclobutadiene. In the structure, an antiaromatic ring pi-dihydrogen bond is found, in which the proton acceptor is antiaromatic 4 electron and 4 center pi bond and the donors are both acidic H atoms of HF molecules. In accompanying with the pi-dihydrogen bond, two secondary interactions are exposed. The first is a repulsive interaction between an H atom of HF and a near pair of H atoms of C4H4 ring. The second is the double pi-type H bond between two lone pairs on a F atom and a far pair of H atoms. PMID- 15268259 TI - Electron and nuclear dynamics of molecular clusters in ultraintense laser fields. I. Extreme multielectron ionization. AB - In this paper we present a theoretical and computational study of extreme multielectron ionization (involving the stripping of all the electrons from light, first-row atoms, and the production of heavily charged ions, e.g., Xe(+q) (q< or =36) from heavy atoms) in elemental and molecular clusters of Xe(n),(D(2))(n), and (CD(4))(n) (n=55-1061) in ultraintense (intensity I=10(15) 10(19) W cm(-2)) laser fields. Single atom or molecule multielectron ionization can be adequately described by the semiclassical barrier suppression ionization (BSI) mechanism. Extreme cluster multielectron ionization is distinct from that of a single atomic or molecular species in terms of the mechanisms, the ionization level and the time scales for electron dynamics and for nuclear motion. The novel compound mechanism of cluster multielectron ionization, which applies when the cluster size (radius R(0)) considerably exceeds the barrier distance for the BSI of a single constituent, involves a sequential-parallel, inner-outer ionization. The cluster inner ionization driven by the BSI for the constituents is induced by a composite field consisting of the laser field and inner fields. The energetics and dynamics of the system consisting of high energy (< or =3 keV) electrons and of less, similar 100 keV ions in the laser field was treated by molecular dynamics simulations, which incorporate electron-electron, electron-ion, ion-ion, and charge-laser interactions. High-energy electron dynamics also incorporates relativistic effects and includes magnetic field effects. We treat inner ionization considering inner field ignition, screening and fluctuation contributions as well as small [(< or =13%)] impact ionization contributions. Subsequent to inner ionization a charged nanoplasma is contained within the cluster, whose response to the composite (laser+inner) field results in outer ionization, which can be approximately described by an entire cluster barrier suppression ionization mechanism. PMID- 15268260 TI - Electron and nuclear dynamics of molecular clusters in ultraintense laser fields. II. Electron dynamics and outer ionization of the nanoplasma. AB - We explore electron dynamics in molecular (CD4)(1061) clusters and elemental Xen (n=249-2171) clusters, responding to ultraintense (intensity I=10(16)-10(19) W cm(-2)) laser fields. Molecular dynamics simulations (including magnetic field and relativistic effects) and analyses of high-energy electron dynamics and nuclear ion dynamics in a cluster interacting with a Gaussian shaped laser field (frequency 0.35 fs(-1), photon energy 1.44 eV, phase 0, temporal width 25 fs) elucidated the time dependence of inner ionization, the formation of a nanoplasma of unbound electrons within the cluster or its vicinity, and of outer ionization. We determined the cluster size and the laser intensity dependence of these three sequential-parallel electronic processes. The characteristic times for cluster inner ionization (tau(ii)) and for outer ionization (tau(oi)) fall in the femtosecond time domain, i.e., tau(ii)=2-9 fs and tau(oi)=4-15 fs for (CD4)(1061), tau(ii)=7-30 fs and tau(oi)=5-13 fs for Xe(n) (n=479,1061), with both tau(ii) and tau(oi) decreasing with increasing I, in accord with the barrier suppression ionization mechanism for inner ionization of the constituents and the cluster barrier suppression ionization mechanism for outer ionization. The positive delay times Deltatau(OI) between outer and inner ionization (e.g., Deltatau(OI)=6.5 fs for Xen at I=10(16) W cm(-2) and Deltatau(OI)=0.2 fs for (CD4)(1061) at I=10(19) W cm(-2)) demonstrate that the outer/inner ionization processes are sequential. For (CD4)(1061), tau(ii)tau(oi), reflecting on the energetic hierarchy in the ionization of the Xe atoms. Quasiresonance contributions to the outer ionization of the nanoplasma were established, as manifested in the temporal oscillations in the inner/outer ionization levels, and in the center of mass of the nanoplasma electrons. The formation characteristics, dynamics, and response of the nanoplasma in molecular or elemental clusters were addressed. The nanoplasma is positively charged, with a high-average electron density [rho(P)=(2-3)10(22) cm(-3)], being characterized by high-average electron energies epsilon(av) (e.g., in Xe(1061) clusters epsilon(av)=54 eV at I=10(16) W cm(-2) and epsilon(av)=0.56-0.37 keV at I=10(18) W cm(-2), with epsilon(av) proportional, variant I(1/2)). Beyond the cluster boundary the average electron energy markedly increases, reaching electron energies in the range of 1.2-40 keV for outer ionization of Xe(n) (n=249-2171) clusters. The nanoplasma exhibits spatial inhomogeneity and angular anisotropy induced by the laser field. Femtosecond time scales are predicted for the nanoplasma production (rise times 7-3 fs), for the decay (decay times approximately 5 fs), and for the persistence time (30-10 fs) of a transient nanoplasma at I=10(17)-10(18) W cm(-2). At lower intensities of I=10(16) W cm(-2) a persistent nanoplasma with a "long" lifetime of > 50 fs will prevail. PMID- 15268261 TI - Applications of pi-photon-induced transparency in two-frequency pulse electron paramagnetic resonance experiments. AB - An approach to pulse electron paramagnetic resonance (EPR) experiments which are based on two different resonance fields is introduced. Instead of using two microwave (mw) sources or a magnetic field jump, bichromatic pulses consisting of a transverse microwave field with frequency omega(mw) and a longitudinal radio frequency field with frequency omega(rf) are employed. Such bichromatic pulses excite a number of multiple photon transitions at frequencies omega(mw)+komega(rf) (k in Z). The pi-photon-induced transparency phenomenon is used to select the required transitions. This approach is used in the stimulated soft electron spin echo envelope modulation and the four-pulse double electron electron resonance experiments. The results obtained using the bichromatic pulse approach are in agreement with those obtained with the standard pulse EPR techniques. It is shown that applying bichromatic pulses is straightforward and advantageous in several respects. PMID- 15268262 TI - Solvatochromic and thermochromic shifts of electronic spectra of polar solute molecules in a mixture of polar and nonpolar solvent; the role of solvent-solvent interactions. AB - A theoretical model is proposed to describe the influence of the concentration of a polar solvent and the temperature of a solution on the electronic spectra of a polar solute in a binary solvent mixture. It is shown that the interaction between molecules of the polar solvent in the first solvation shell makes the significant contribution to the formation of absorption and fluorescence bands of the solute. An experimental study of solvatochromic and thermochromic shifts of steady-state fluorescence spectra of 3-amino-N-methylphthalimide in decalin- propanol mixture for different values of propanol mole fraction is carried out. Good qualitative agreement between the experimental data and calculation results is observed. PMID- 15268263 TI - Dipole solvation in dielectrics. AB - This paper presents an exact solution for the free energy of linear solvation of a dipolar solute in an arbitrary dielectric material with a microscopic spectrum of polarization fluctuations. The solution is given in terms of wave vector dependent longitudinal and transverse structure factors of the polarization fluctuations in the pure dielectric. Good agreement with computer simulations of dipole solvation in dipolar and dipolar--quadrupolar liquids is achieved. PMID- 15268264 TI - Environmental swap energy and role of configurational entropy in transfer of small molecules from water into alkanes. AB - We studied the effect of segmented solvent molecules on the free energy of transfer of small molecules from water into alkanes (hexane, heptane, octane, decane, dodecane, tetradecane, and hexadecane). For these alkanes we measured partition coefficients of benzene, 3-methylindole (3MI), 2,3,4,6 tetrachlorophenol (TeCP), and 2,4,6-tribromophenol (TriBP) at 3, 11, 20, 33 [corrected], and 47 degrees C. For 3MI, TeCP, and TriBP the dependence of free energy of transfer on length of alkane chains was found to be very different from that for benzene. In contrast to benzene, the energy of transfer for 3MI, TeCP, and TriBP was independent of the number of carbons in alkanes. To interpret data, we used the classic Flory-Huggins (FH) theory of concentrated polymer solutions for the alkane phase. For benzene, the measured dependence of energy of transfer on the number of carbons in alkanes agreed well with predictions based on FH model in which the size of alkane segments was obtained from the ratio of molar volumes of alkanes and the solute. We show that for benzene, the energy of transfer can be divided into two components, one called environmental swap energy (ESE), and one representing the contribution of configurational entropy of alkane chains. For 3MI, TeCP, and TriBP the contribution of configurational entropy was not measurable even though the magnitude of the effect predicted from the FH model for short chain alkanes was as much as 20 times greater than experimental uncertainties. From the temperature dependence of ESE we obtained enthalpy and entropy of transfer for benzene, 3MI, TeCP, and TriBP. Experimental results are discussed in terms of a thermodynamic cycle considering creation of cavity, insertion of solute, and activation of solute-medium attractive interactions. Our results suggest that correcting experimental free energy of transfer by Flory Huggins configurational entropy term is not generally appropriate and cannot be applied indiscriminately. PMID- 15268265 TI - Molecular-dynamics studies of surface of ice Ih. AB - We performed molecular dynamics calculations of surface of ice Ih in order to investigate formation mechanism of melting layer on the surface. The results showed that the vibrational amplitude of the atoms in the surface layer greatly depends on the crystal orientation, whereas that in the ice bulk is isotropic. The anisotropy of the vibration is due to a dangling motion of the free O-H bonds exist at the surface layer. The dangling motion enhances the rotational motion of the water molecules. The vibrational density of state showed a coupling between the rotational vibration and the lattice vibration of the water molecules in the surface layer. The coupling of the vibrations causes a distortion of ice lattice. Through the hydrogen-bonding network, the distortion transmits to the interior of the crystal. We conclude that the dangling motion of the free O-H bonds exist at the surface layer is one of the dominant factors governing the surface melting of ice crystal. PMID- 15268266 TI - Ion mobilities and microscopic dynamics in liquid (Li,K)Cl. AB - The dynamical properties of ionic melts formed from mixtures of LiCl and KCl have been studied across the full composition range in computer simulations of sufficient length to enable reliable values for such collective transport coefficients as the viscosity, conductivity, and internal mobilities to be determined reliably. Interest centers on the nontrivial concentration dependence exhibited by these transport coefficients, which agrees well with that observed experimentally, and in relating this to the strength of the association between an ion and its first coordination shell. The relationships between the various transport coefficients, such as those between the diffusion coefficient and the viscosity (Stokes-Einstein) and the conductivity (Nernst-Einstein) also exhibit composition dependences that reflect this association. The connection between the internal mobility and two measures of the coordination shell dynamics (the cage relaxation time and the self-exchange velocity) is explored; it is shown that the self-exchange velocity follows the composition and temperature dependence of the internal mobility very well. Finally, it is shown that allowing for anion polarization in the interaction model increases the mobility of all species without changing the structure of the melt discernibly, with the largest effect being found for the Li(+) ion. PMID- 15268267 TI - Photochemistry in the charge transfer and neutral excited states of HCl in Xe and Kr matrices. AB - HCl-doped Xe and Kr films are irradiated with wavelength dispersed synchrotron radiation in the wavelength range from 200 to 130 nm. The growth of H, Cl, Xe2H+, XeH2, HXeCl, Kr2H+, and HKrCl as well as the decomposition of HCl are recorded by a combination of UV, VIS, and IR spectroscopy. A turnover in the formation of Xe2H+ and Kr2H+ by a predominant two-step reaction on neutral surfaces at low energies to a one-step formation on ionic surfaces is determined at 172 and 155 nm in Xe and Kr, respectively. A potential energy diagram for neutral and ionic states is derived that is consistent with a DIIS calculation, with new UV fluorescence bands from Xe+HCl- centers, with the turnover energies and with a deconvolution of the absorption spectra in neutral and ionic contributions. The cage exit of charged as well as of neutral H, the latter via a harpoon reaction, is discussed for the ionic surfaces. The self-limitation of HCl decomposition on the neutral surfaces due to absorption by H and Cl fragments is treated quantatively. Dissociation efficiencies phi(e), together with absolute absorption cross sections sigma(H) and sigma(Cl) of the fragments, are derived. sigma(H) and sigma(Cl) are of the order of 10(-16) cm(2) compared to 10(-18) cm(2) for sigma(HCl). Dissociation is accompanied by many excitation cycles of the fragments, which leads to light-induced migration of H and recombination. phi(e) therefore represents a product of the cage exit probability phi that was treated theoretically and the survival probability concerning geminate and nongeminate recombination. PMID- 15268268 TI - Dynamic light scattering in liquid and supercooled diphenylmethane. AB - We use a dynamic light scattering technique to measure both polarized (VV) and depolarized (VH) spectra of liquid diphenylmethane (DPM) between 288 and 362 K, covering both normal and supercooled liquid ranges. Our results allow extracting information on structural relaxation processes, rotational motions, rotation translation couplings, and molecular reorientation phenomena in liquid DPM. The VV spectra are modeled according to the microscopic theory of Wang, which assumes that a structural relaxation process dominates the spectrum. We find that the relaxation time of the structural relaxation in DPM follows an Arrhenius behavior. The Rayleigh dip was observed in the VH spectra, which are described using the Andersen-Pecora theory. Our results are discussed in terms of the rotation-translation coupling parameter, which we find independent of temperature over the experimental range. The collective reorientation time also follows an Arrhenius behavior with temperature. Finally, we calculate the hydrodynamic volumes for the reorientation process from geometric molecular models in two hydrodynamic limits: slip and stick boundary conditions. Our results suggest that the DMP molecule reorientates in quasi-slipping conditions in the bulk liquid. PMID- 15268269 TI - Low temperature electron transfer in strongly condensed phase. AB - Electron transfer coupled to a collective vibronic degree of freedom is studied in strongly condensed phase and at lower temperatures where quantum fluctuations are essential. Based on an exact representation of the reduced density matrix of the electronic + reaction coordinate compound in terms of path integrals, recent findings on the overdamped limit in quantum dissipative systems are employed. This allows us to give a consistent generalization of the well-known Zusman equations to the quantum domain. Detailed conditions for the range of validity are specified. Using the Wigner transform these results are also extended to the quantum dynamics in full phase space. As an important application electronic transfer rates are derived that comprise adiabatic and nonadiabatic processes in the low temperature regime including nuclear tunneling. Accurate agreement with precise quantum Monte Carlo data is observed. PMID- 15268270 TI - Experimental CC stretching phonon dispersion curves and electron phonon coupling in polyene derivatives. AB - We analyze the infrared and Raman spectra (both experimentally and with the aid of quantum chemical calculations) of a series of polyenals which provide us with the fortunate case of a set of polyene chains with one of the end groups consisting of a C=O group which not only does take part in the conjugation but also pulls electrons from the chain making the whole system highly polar, thus affecting the vibrational transition moments. In the following we show, for the first time, that it is possible to derive experimental phonon dispersion curves and these prove to be different for each chain length. We support our experimental findings with Density Functional Theory quantum chemical calculations which reproduce with sufficient accuracy the IR and Raman spectral pattern and at the same time help in disentangling the assignment of the fine structure observed in the experimental spectra. PMID- 15268271 TI - A fully self-consistent treatment of collective fluctuations in quantum liquids. AB - The problem of calculating collective density fluctuations in quantum liquids is revisited. A fully quantum mechanical self-consistent treatment based on a quantum mode-coupling theory is presented. The theory is compared with the maximum entropy analytic continuation approach and with available experimental results. The quantum mode-coupling theory provides semiquantitative results for both short and long time dynamics. The proper description of long time phenomena is important in future study of problems related to the physics of glassy quantum systems, and to the study of collective fluctuations in Bose fluids. PMID- 15268272 TI - Nonresonant holeburning in the Terahertz range: Brownian oscillator model. AB - The response to the field sequence of nonresonant hole burning, a pump-wait-probe experiment originally designed to investigate slow relaxation in complex systems, is calculated for a model of Brownian oscillators, thus including inertial effects. In the overdamped regime the model predictions are very similar to those of the purely dissipative stochastic models investigated earlier, including the possibility to discriminate between dynamic homogeneous and heterogeneous relaxation. The case of underdamped oscillations is of particular interest when low-frequency excitations in glassy systems are considered. We show that also in this situation a frequency selective modification of the response should be feasable. This means that it is possible to specifically address various parts of the spectrum. An experimental realization of nonresonant holeburning in the Terahertz regime therefore is expected to shed further light on the nature of the vibrations around the so-called boson peak. PMID- 15268273 TI - Non-Gaussian statistics of amide I mode frequency fluctuation of N methylacetamide in methanol solution: linear and nonlinear vibrational spectra. AB - By carrying out molecular dynamics simulations of an N-methylacetamide (NMA) in methanol solution, the amide I mode frequency fluctuation and hydrogen bonding dynamics were theoretically investigated. Combining an extrapolation formula developed from systematic ab initio calculation studies of NMA-(CH3OH)n clusters with a classical molecular dynamics simulation method, we were able to quantitatively describe the solvatochromic vibrational frequency shift induced by the hydrogen-bonding interaction between NMA and solvent methanol. It was found that the fluctuating amide I mode frequency distribution is notably non-Gaussian and it can be decomposed into two Gaussian peaks that are associated with two distinctively different solvation structures. The ensemble-average-calculated linear response function associated with the IR absorption is found to be oscillating, which is in turn related to the doublet amide I band shape. Numerically calculated infrared absorption spectra are directly compared with experiment and the agreement was found to be excellent. By using the Onsager's regression hypothesis, the rate constants of the interconversion process between the two solvation structures were obtained. Then, the nonlinear response functions associated with two-dimensional infrared pump-probe spectroscopy were simulated. The physics behind the two-dimensional line shape and origin of the cross peaks in the time-resolved pump-probe spectra is explained and the result is compared with 2D spectra experimentally measured recently by Woutersen et al. PMID- 15268274 TI - Semiclassical calculation of the vibrational echo. AB - The infrared echo measurement probes the time scales of the molecular motions that couple to a vibrational transition. Computation of the echo observable within rigorous quantum mechanics is problematic for systems with many degrees of freedom, motivating the development of semiclassical approximations to the nonlinear optical response. We present a semiclassical approximation to the echo observable, based on the Herman-Kluk propagator. This calculation requires averaging over a quantity generated by two pairs of classical trajectories and associated stability matrices, connected by a pair of phase-space jumps. Quantum, classical, and semiclassical echo calculations are compared for a thermal ensemble of noninteracting anharmonic oscillators. The semiclassical approach uses input from classical mechanics to reproduce the significant features of a complete, quantum mechanical calculation of the nonlinear response. PMID- 15268275 TI - Charge transfer mechanism in hybrid bulk heterojunction composites. AB - Charge transfer mechanisms of several conducting polymers and polymer/CdSe nanocrystal composites (hybrid bulk heterojunction composites) were investigated by means of cyclic voltammetry. Potential application of these composites in hybrid light-emitting diodes was discussed. It was found that charge transfer is observed in most of the composites, used so far, but was relatively slow or incomplete. The PVPy/CdSe nanocrystal composite showed promising results, and is favorable for use in electroluminescent devices. PMID- 15268276 TI - Attraction-driven disorder in a hard-core colloidal monolayer. AB - Monte Carlo simulation techniques were employed to explore the effect of short range attraction on the orientational ordering in a two-dimensional assembly of monodisperse spherical particles. We find that if the range of square-well attraction is approximately 15% of the particle diameter, the dense attractive fluid shows the same ordering behavior as the same density fluid composed of purely repulsive hard spheres. Fluids with an attraction range larger than 15% show an enhanced tendency to crystallization, while disorder occurs for fluids with an attractive range shorter than 15% of the particle diameter. A possible link with the existence of "repulsive" and "attractive" states in dense colloidal systems is discussed. PMID- 15268277 TI - Surface nuclear spin relaxation of 199Hg. AB - We report the results of a detailed study of surface relaxation of 199Hg nuclear spins in paraffin coated cells. From measurements of the magnetic field and temperature dependence of the spin relaxation rates we determine the correlation time for magnetic fluctuations and the surface adsorption energy. The data indicate that surface relaxation is caused by dipolar coupling to paramagnetic sites on the surface. We also observe changes in the spin relaxation rate caused by ultraviolet radiation resonant with the 254 nm transition in Hg. PMID- 15268278 TI - Adsorption, desorption, and clustering of H2O on Pt111. AB - The adsorption, desorption, and clustering behavior of H2O on Pt111 has been investigated by specular He scattering. The data show that water adsorbed on a clean Pt111 surface undergoes a structural transition from a random distribution to clustered islands near 60 K. The initial helium scattering cross sections as a function of temperature are found to be insensitive to the incident H2O flux over a range of 0.005 monolayers (ML)/s-0.55 ML/s indicating that the clustering process is more complex than simple surface diffusion. The coarsening process of an initially random distribution of water deposited at 25 K is found to occur over a broad temperature range, 60X+ transition in ethylene and ethylene-d4 have been recorded at a resolution of 0.09 cm(-1). The spectra provide new information on the large amplitude torsional motion in the cationic ground state. An effective one dimensional torsional potential was determined from the experimental data. Both C2H4+ and C2D4+ exhibit a twisted geometry, and the lowest two levels of the torsional potential form a tunneling pair with a tunneling splitting of 83.7(5) cm(-1) in C2H4+ and of 37.1(5) cm(-1) in C2D4+. A model was developed to quantitatively analyze the rotational structure of the photoelectron spectra by generalizing the model of Buckingham, Orr, and Sichel [Philos. Trans. R. Soc. London, Ser. A 268, 147 (1970)] to treat asymmetric top molecules. The quantitative analysis of the rotational intensity distributions of allowed as well as forbidden vibrational bands enabled the identification of strong vibronic mixing between the X+ and A+ states mediated by the torsional mode nu(4) and a weaker mixing between the X+ and B+ states mediated by the symmetric CH2 out-of plane bending mode nu7. The vibrational intensities could be accounted for quantitatively using a Herzberg-Teller-type model for vibronic intensity borrowing. The adiabatic ionization energies of C2H4 and C2D4 were determined to be 84 790.42(23) cm(-1) and 84 913.3(14) cm(-1), respectively. PMID- 15268307 TI - Double ionization of fluorinated benzenes: hole localization and delocalization effects. AB - The dense double ionization spectra of all the twelve fluoro-substituted benzene molecules are investigated in great detail by Green's function ADC2 calculations and a two-hole density mapping. Double ionization is shown to provide an extremely sensitive tool of electronic structure analysis. The calculations evidence and measure quantitatively how the charge distribution is dictated by the complex interplay between the resilience of the aromatic ring electronic structure and the disruptive effect of the electronegative halogen substituents. Successive substitutions are found not to have any synergic effect, but affect the spectra in very identifiable ways. The Auger spectra of the fluorobenzenes are interpreted in the light of the charge distribution results, using the foreign-imaging model. The double charge transfer spectra are also analyzed and discussed. PMID- 15268308 TI - Reaction dynamics of Cl+CH3SH: rotational and vibrational distributions of HCl probed with time-resolved Fourier-transform spectroscopy. AB - Rotationally resolved infrared emission spectra of HCl(v=1-3) in the reaction of Cl+CH3SH, initiated with radiation from a laser at 308 nm, are detected with a step-scan Fourier-transform spectrometer. Observed rotational temperature of HCl(v=1-3) decreases with duration of reaction due to collisional quenching; a short extrapolation to time zero based on data in the range 0.25-4.25 micros yields a nascent rotational temperature of 1150+/-80 K. The rotational energy averaged for HCl(v=1-3) is 8.2+/-0.9 kJ mol(-1), yielding a fraction of available energy going into rotation of HCl, fr=0.10+/-0.01, nearly identical to that of the reaction Cl+H(2)S. Observed temporal profiles of the vibrational population of HCl(v=1-3) are fitted with a kinetic model of formation and quenching of HCl(v=1-3) to yield a branching ratio (68+/-5):(25+/-4):(7+/-1) for formation of HCl(v=1):(v=2):(v=3) from the title reaction and its thermal rate coefficient k(2a)=(2.9+/-0.7)x10(-10) cm(3) molecule(-1) s(-1). Considering possible estimates of the vibrational population of HCl(v=0) based on various surprisal analyses, we report an average vibrational energy 36+/-6 kJ mol(-1) for HCl. The fraction of available energy going into vibration of HCl is f(v)=0.45+/-0.08, significantly greater than a value fv=0.33+/-0.06 determined previously for Cl+H2S. Reaction dynamics of Cl+H(2)S and Cl+CH3SH are compared; the adduct CH3S(Cl)H is likely more transitory than the adduct H(2)SCl. PMID- 15268309 TI - Isotopic effect on the cage-induced quenching of OH(A)/OD(A) inside small argon clusters. AB - In this paper we report on the isotopic effect on the cage-induced excited-state quenching inside small Ar(m) clusters (m<10(2)) solvated in large Ne(N) clusters (N approximately 7.5x10(3)). Excited OH(A)/OD(A) fragments are produced by photodissociation of H2O and D2O molecules and the quenching agents are correspondingly H or D atoms. The decrease of the fluorescence yield with the size of the cluster m>m0 is observed in both cases and it is attributed to the formation of the cage of argon atoms around the doped molecule. Interestingly, more atoms are needed to induce the fluorescence quenching of OD*(A) fragments, m0=21+/-3, compared to the electronically excited state quenching of OH*(A) molecules, 11+/-2. A diffusion model containing two free parameters, the quenching cross section sigmaq and the number of argon atoms forming the cage m0, explains the effect in terms of the residence time of the hydrogen atom inside the cage. We suggest that the melting of the doped rare gas clusters is responsible for the different predissociation dynamics. The quenching cross section obtained from the experimental data is in good agreement with former experiments. PMID- 15268310 TI - Electron-spin polarization of photoions produced through photoionization from the laser-excited triplet state of Sr. AB - We report the detailed experimental study on the production of electron-spin polarized Sr+ ions through one-photon resonant two-photon ionization via laser excited 5s5p 3P1 (MJ = +1) of Sr atoms produced by laser-ablation. We have experimentally confirmed that the use of laser-ablation for the production of Sr atoms prior to photoionization does not affect the electron-spin polarization. We have found that the degree of electron-spin polarization is 64+/-9%, which is in good agreement with our recent theoretical prediction. As we discuss in detail, we infer, from a simple analysis, that photoelectrons, being the counterpart of electron-spin-polarized Sr+ ions, have approximately the same degree of electron spin polarization. Our experimental results demonstrate that the combined use of laser-ablation technique and pulsed lasers for photoionization would be a compact and effective way to realize a pulsed source for spin-polarized ions and electrons for the studies of various spin-dependent dynamics in chemical physics. PMID- 15268311 TI - On symmetry breaking in BNB: real or artifactual? AB - The ground state of the linear BNB molecule has been examined with multireference based ab initio methods coupled with quantitative basis sets. Previous computational studies on BNB, even those using highly correlated single reference based methods, e.g., the CCSD(T) and BDT methods, suggested that the two BN bond lengths were unequal. In this paper, the BN(X 3Pi) + B(2Pu) potential energy curve is constructed using a state-averaged multireference-based correlated method (SA-CASSCF+PT2). The four lowest states of BN were included in the state averaging procedure. These calculations reveal no symmetry breaking along the antisymmetric stretching mode of the molecule. PMID- 15268312 TI - The mutual diffusion coefficient for (meth)acrylate monomers as determined with a nuclear microprobe. AB - The value of the mutual diffusion coefficient DV of two acrylic monomers is determined with nuclear microprobe measurements on a set of polymer films. These films have been prepared by allowing the monomers to diffuse into each other for a certain time and subsequently applying fast ultraviolet photo-polymerization, which freezes the concentration profile. The monomer diffusion profiles are studied with a scanning 2.1 MeV proton microprobe. Each monomer contains a marker element, e.g., Cl and Si, which are easily detected with proton induced x-ray emission. From the diffusion profiles, it is possible to determine the mutual diffusion coefficient. The mutual diffusion coefficient is dependent of concentration, which is concluded from the asymmetry in the Cl- and Si-profiles. A linear dependence of the mutual diffusion coefficient on the composition is used as a first order approximation. The best fits are obtained for a value of b=(0.38+/-0.15), which is the ratio of the diffusion coefficient of 1,3-bis(3 methacryloxypropyl)-1, 1,3,3-tetramethyldisiloxane in pure 2-chloroethyl acrylate and the diffusion coefficient of 2-chloroethyl acrylate in pure 1,3-bis(3 methacryloxypropyl)-1,1,3,3-tetramethyldisiloxane. Under the assumption of a linear dependence of the mutual diffusion coefficient DV on monomer composition, it follows that DV = (2.9+/-0.6)10(-10) m(2)/s at a 1:1 monomer ratio. With Flory Huggins expressions for the monomer chemical potentials, one can derive approximate values for the individual monomer diffusion coefficients. PMID- 15268313 TI - Molecular dynamics study of atomic transport properties in rapidly cooling liquid copper. AB - Based on Mei's embedded atom model molecular dynamics simulations have been performed to investigate the rapidly cooling processes of Cu. The atomic transport property, namely the self-diffusion coefficient, is computed in the liquid state, and the results near the melting point of Cu are in good agreement with experimental data and other computational values. The atom diffusion movements during the long period of relaxation have been also studied around the solidification temperature Tc. To describe the complex microstructural evolutions during the rapidly cooling processes and the long relaxation processes, the pair correlation function and the pair analysis technique are used. It is demonstrated that the crystallization of amorphous Cu is caused by the atomic diffusion. PMID- 15268314 TI - Orientational order in binary mixtures of hard Gaussian overlap molecules. AB - Based on a standard constant-pressure Monte Carlo molecular simulation, we have studied liquid crystal phases of binary mixtures of nonspherical molecules. The components of the mixtures are two types of hard Gaussian overlap (HGO) molecules. The first type of molecule has a small molecularelongation parameter (short HGO molecules) and cannot form stable liquid crystal phase in the bulk by themselves. The second type of molecule has a large elongation parameter (long HGO molecules) and can form a liquid crystal phase easily. In the mixtures, the short HGO molecules can form an orientationally ordered phase because the long HGO molecules form confining surfaces to induce the alignment of the short molecules. We also study the isotropic-nematic phase transition in different mixtures composed of short and long HGO molecules with different elongations and concentrations. The obtained result implies that small anisotropic molecules can show liquid crystal behavior. PMID- 15268315 TI - Precipitate pattern formation in fluctuating media. AB - Simulation of the Liesegang pattern formation in low concentration gradient is presented using concentration perturbation in a deterministic model. The precipitation process is based on ion-product supersaturation theory (Ostwald's model). In the classical experiments with high initial concentration gradients, appearance time and locations of the band formation are well reproducible. Decreasing initial concentration gradients results in a more stochastic pattern structure; this means that the reproducibility of the experiments becomes worse. The presented model and the results of the simulations exhibit the same trend, which were demonstrated and investigated experimentally by Kai et al. PMID- 15268316 TI - Conformation of p-terphenyl under hydrostatic pressure. AB - The conformation of p-terphenyl (C18H14) and deuterated p-terphenyl (C18D14) has been investigated, using high-pressure infrared spectroscopy at liquid-helium temperatures. First-principles calculations, together with the experimental results, were performed to determine the structure of p-terphenyl in the twisted conformation. At low temperatures and pressures, p-terphenyl belongs to the C2 point group of symmetry. In this configuration, the central ring is twisted with respect to the plane of the outer rings. The symmetry of the molecule is nearly C2h, consistent with previous x-ray diffraction measurements. PMID- 15268317 TI - Statistico-probabilistic approach to taking account of the vapor depletion in the kinetics of homogeneous nucleation: a free-molecular regime of droplet growth. AB - We propose a statistico-probabilistic approach to investigate the process of homogeneous formation of droplets in a vapor phase in the presence of an already formed and growing droplet under free-molecular regime of droplet growth after the instantaneous creation of initial vapor supersaturation. We find the probability density for the formation of a new, nearest (neighbor) droplet in the vicinity of an initially formed droplet. The mean distance between two neighboring droplets is also determined, as well as the average time lag for the formation of the nearest (neighbor) droplet; the latter quantity serves as an estimate for the duration of the nucleation stage. An estimate for the average number of droplets forming in unit volume by the end of the nucleation stage is also given. Our results are compared with the predictions of classical nucleation theory which assumes the density uniformity of a metastable phase. Where the proposed approach is applicable, there is observed qualitative agreement between the results. The underlying cause of this agreement is analyzed and the limits of applicability of the uniformity approximation are clarified. PMID- 15268318 TI - Structure of ionic liquids of 1-alkyl-3-methylimidazolium cations: a systematic computer simulation study. AB - Molecular dynamics simulations of room temperature molten salts (ionic liquids) containing imidazolium cations have been performed. Ten different systems were simulated at 323 K by using united atom force fields, in which the anion size (F , Cl-, Br-, and PF6-) and the length of the alkyl chain of 1-alkyl-3 methylimidazolium cations (1-methyl-, 1-ethyl-, 1-butyl-, and 1-octyl-) were systematically varied. It is shown that the resulting equilibrium structures account for the observed features of experimental static structure factors when available. A detailed analysis of the simultaneous effect of changing the anion and the alkyl chain on the preferential location of nearest-neighbor anions around the cations is provided. It is shown that regions above and below the imidazolium ring are the preferential ones in case of large anions. By increasing the length of the alkyl chain, nearest-neighbor anions are pushed away from the volume occupied by the flexible alkyl chain. Partial structure factors of 1-butyl and 1-octyl- derivatives display a peak at a wave vector smaller than the main peak, indicating the occurrence of an intermediate range order in these ionic liquids due to the presence of long alkyl chains. PMID- 15268319 TI - On the ordering of the first two excited electronic states in all-trans linear polyenes. AB - Reported experimental evidence of the relative position of the first two excited electronic states in linear polyenes was carefully examined and compared with that derived from time dependent density functional theory (TDDFT) theoretical calculations performed at the B3LYP level on optimized geometries. The energy values for the first two triplet states 3Bu and 3Ag, obtained from TDDFT calculations, were found to be highly strongly correlated with the experimental values. Also, the theoretical calculations for the electronic transition 1 1Ag - > 1 1Bu were also extremely well correlated with their experimental counterparts; even more important, the three reported experimental data for 1 1Ag --> 2 1Ag transitions in these systems conformed to the correlation for the TDDFT 1 1Ag --> 1 1Bu transition. The first excited electronic state in the linear polyenes studied (from ethene to the compound consisting of 40 ethene units, P40) was found to be 1Bu. The energy gap between the excited states 2 1Ag and 1 1Bu decreased with increasing length of the polyene chain, but not to the extent required to cause inversion, at least up to P40. In the all-trans linear polyenes studied, the widely analyzed energy gap from the ground electronic state to the first excited singlet state for infinitely long chains may be meaningless as, even in P40, it is uncertain whether the ground electronic state continues to be a singlet. PMID- 15268320 TI - Multipole induced splitting of metal-cage vibrations in crystalline endohedral D2d-M2@C84 dimetallofullerenes. AB - Metal-carbon cage vibrations of crystalline endohedral D2d-M2@C84 (M=Sc,Y,Dy) dimetallofullerenes were analyzed by temperature dependent Raman scattering and a dynamical force field model. Three groups of metal-carbon cage modes were found at energies of 35-200 cm(-1) and assigned to metal-cage stretching and deformation vibrations. They exhibit a textbook example for the splitting of molecular vibrations in a crystal field. Induced dipole-dipole and quadrupole quadrupole interactions account quantitatively for the observed mode splitting. Based on the metal-cage vibrational structure it is demonstrated that D2d-Y2@C84 dimetallofullerene retains a monoclinic crystal structure up to 550 K and undergoes a transition from a disordered to an ordered orientational state at a temperature of approximately 150 K. PMID- 15268321 TI - Probing lattice distortions in mixed CH3I1-cBrc by methyl rotational tunneling. AB - Methyl iodide alloyed with methyl bromide is studied for low methyl bromide concentrations c < or = 0.3 by rotational tunneling spectroscopy with neutrons. The appearance of three tunneling bands, their shift with respect to the pure materials and their broadening is explained semiquantitatively on the basis of the crystal structure and global and local changes of interatomic distances based on the r-dependence of intermolecular interactions. Besides the overall reduction of the lattice parameter local free volume around guest molecules is important. A local relaxation of the atomic position by 1.3% towards guest molecules is found. PMID- 15268322 TI - Near neighbor approximation in nuclear magnetic resonance. AB - A new way to deal with the excitation by multiple effective RF fields with interference is presented using the coherent averaging theory. It significantly simplifies the calculation of the effect of RF interference that occurs in the excitations by periodic pulses and phase-incremented pulses (PIPs). This approach shows that each neighboring RF field contributes to an excitation profile an offset shift, which is termed the Bloch-Siegert offset shift (BSOS). The BSOS depends not only on the strengths of both RF fields that interfere with each other but also on their relative phase between the two RF fields. Consequently, it can be positive, negative, and zero. In addition, the BSOS is also inversely proportional to the frequency separation of the two RF fields. Therefore, only a few near neighbors need to be taken into account in most cases, resulting in a near neighbor approximation (NNA). The BSOS for two multiband excitation profiles, one by a periodic pulse and the other by a PIP, are calculated using the NNA. The results are in good agreement with the computer simulated ones. PMID- 15268323 TI - Monte Carlo simulation methodology of the ghost interface theory for the planar surface tension. AB - A novel "ghost interface" expression for the surface tension of a planar liquid vapor interface is derived in detail from consideration of the free energy of the system, and a methodology for utilization of this new technique is given. An augmented Monte Carlo computer simulation procedure is developed specifically for the ghost interface, including derivation of long-range corrections resulting from potential truncation and a modified Gibbs ensemble technique for the simulation of adjacent coexisting phases. Results generated from the ghost interface theory for the surface tension are presented and found to be in good quantitative agreement with those resulting from the Kirkwood-Buff equation. Applications of this new approach to curved and to supersaturated systems are also discussed. PMID- 15268324 TI - Resonant Raman spectroscopy of PAH--Os self-assembled multilayers. AB - We present a resonant Raman scattering study of (PAH--Os/PVS)n and (PAH--Os/GOx)m self-assembled multilayers (n=1-11 and m=1-3). These Os polymer multilayers can be used in electrodes as efficient molecular wires for biomolecular recognition. The Raman intensity dependence on the number of self-assembly cycles provides information on the deposition process. The spectra are identical to that observed for PAH--Os in aqueous solution, indicating that the PAH--Os metal complex structure is conserved in the multilayers. We observe at approximately 500 nm incoming and outgoing Raman resonances of osmium and bipyridine vibrational modes. These resonances are associated to the metal-to-ligand charge transfer (MLCT) transition. We study the evolution of these Raman modes as a function of the Os oxidation state during in situ electrochemistry. During the oxidation process, Os(II)-->Os(III), the Raman resonance related to the MLCT disappears and the bipyridine related modes harden by approximately 10 cm(-1). These results are correlated with optical transmission measurements which show the disappearance of the visible region absorption when the Os complex is oxidized. We also find partial quenching of the Raman mode intensity after in situ voltamperometric cycles which demonstrates the existence of photo-electro-chemical processes. PMID- 15268325 TI - Anomalous dielectric relaxation of water molecules at the surface of an aqueous micelle. AB - Dielectric relaxation of aqueous solutions of micelles, proteins, and many complex systems shows an anomalous dispersion at frequencies intermediate between those corresponding to the rotational motion of bulk water and that of the organized assembly or macromolecule. The precise origin of this anomalous dispersion is not well-understood. In this work we employ large scale atomistic molecular dynamics simulations to investigate the dielectric relaxation (DR) of water molecules in an aqueous micellar solution of cesium pentadecafluorooctanoate. The simulations clearly show the presence of a slow component in the moment-moment time correlation function [PhiMW(t)] of water molecules, with a time constant of about 40 ps, in contrast to only 9 ps for bulk water. Interestingly, the orientational time correlation function [Cmu(t)] of individual water molecules at the surface exhibits a component with a time constant of about 19 ps. We show that these two time constants can be related by the well-known micro-macrorelations of statistical mechanics. In addition, the reorientation of surface water molecules exhibits a very slow component that decays with a time constant of about 500 ps. An analysis of hydrogen bond lifetime and of the rotational relaxation in the coordinate frame fixed on the micellar body seems to suggest that the 500 ps component owes its origin to the existence of an extended hydrogen bond network of water molecules at the surface. However, this ultraslow component is not found in the total moment-moment time correlation function of water molecules in the solution. The slow DR of hydration water is found to be well correlated with the slow solvation dynamics of cesium ions at the water-micelle interface. PMID- 15268326 TI - Solvation force for long-ranged wall--fluid potentials. AB - The solvation force of a simple fluid confined between identical planar walls is studied in two model systems with short ranged fluid-fluid interactions and long ranged wall-fluid potentials decaying as -Az(-p),z--> infinity, for various values of p. Results for the Ising spins system are obtained in two dimensions at vanishing bulk magnetic field h=0 by means of the density-matrix renormalization group method; results for the truncated Lennard-Jones (LJ) fluid are obtained within the nonlocal density functional theory. At low temperatures the solvation force f(solv) for the Ising film is repulsive and decays for large wall separations L in the same fashion as the boundary field f(solv) approximately L( p), whereas for temperatures larger than the bulk critical temperature f(solv) is attractive and the asymptotic decay is f(solv) approximately L(-(p+1)). For the LJ fluid system f(solv) is always repulsive away from the critical region and decays for large L with the the same power law as the wall-fluid potential. We discuss the influence of the critical Casimir effect and of capillary condensation on the behavior of the solvation force. PMID- 15268327 TI - Molecular dynamics simulation of the coalescence of nanometer-sized water droplets in n-heptane. AB - Molecular dynamics simulations using a modified Drieding 2.21 force field were carried out to study the coalescence behavior of nanometer-sized water droplets in vacuum and in n-heptane. The coalescence mechanisms of the water droplets in the above-noted environments are fairly similar in a sense that the water droplets form a bridge linking the droplets before they merge. However, in the latter situation, due to the presence of n-heptane molecules in between the water droplets, the coalescence was observed to be slowed down considerably, especially in the first 10 ps of the process. However, once the bridge is formed, the water droplets, in both situations, spend about the same amount of time to form a single droplet. The maximum distance between the droplets above which coalescence does not occur was found to be 10 A. In terms of the dynamics, the diffusion coefficient of n-heptane in the emulsion system was very close to its value in the pure liquid form. This may be because n-heptane is the continuous phase. Nonetheless, the dynamic behavior of water in n-heptane is different from that of pure water during and after the coalescence. In particular, the self-diffusion coefficient of water molecules in n-heptane is about 20% higher than the experimental value of pure water. Due to the lack of strong attraction forces between water and n-heptane molecules, the n-heptane molecules were observed to orient themselves perpendicularly to the water/n-heptane interfaces so that the contacting area is minimized. PMID- 15268328 TI - Molecular dynamics simulation of water in a contact with an iron pyrite FeS2 surface. AB - A strong adsorption of the water molecules to the pyrite surface is shown by a molecular dynamic simulation of the water-iron pyrite FeS2 interface. Water molecules closest to the pyrite surface are bound by an electrostatic interaction to the iron atoms in grooves running parallel to one of the crystal axes. The grooves are about two atoms wide and are directed along 010 for the (001) surface. The position of the water-surface potential minimum and the energy of adsorption were determined by optimization for a single water molecule at the interface. At room temperature and normal density there are altogether three distinguishable layers of water above the surface. One is associated with the groove: one with H bonding to the sulphur atoms comprising the ridges separating the grooves, and the third with the soft wall boundary between the absorbed water layers and bulk region of water. Simulations were also used to explore the effect of a temperature range significant for geophysical studies. PMID- 15268329 TI - Molecular dynamic simulation of the hydration and diffusion of chloride ions from bulk water to polypyrrole matrix. AB - A molecular dynamic simulation of wet polypyrrole film was carried out, in both oxidized and reduced state. The system was modeled by two layers of polypyrrole, water and chloride ions (as counterions required for charge balance in the oxidized state) in atomic detail to provide an insight into some dynamic and steady properties of the system. Our simulations pointed to a swelling of the polymer matrix after oxidation due to electrostatic repulsions between charged sites of the oxidized polypyrrole, followed by penetration of the polypyrrole by counterions to maintain the electroneutrality of the system. Associated with this penetration of counterions toward the core of the oxidized polypyrrole, dehydration of the counterions was observed. This dehydration was compensated (in part) by a strong coordination with the charged sites of the polymer. The remaining hydrophobicity inside the polymer also contributed to the dehydration of these counterions. The translational diffusion coefficient of chloride ions was also calculated at different positions of the polypyrrole/water interface, from bulk water to the inner polymer matrix. A value of 4.1 x 10(-5) cm(2) s(-1) was measured in the bulk water compared to 5 x 10(-7) cm(2) s(-1) inside the polymer, representing a diminution of two orders of magnitude for the translational diffusion coefficient from bulk water to the core of a oxidized polypyrrole matrix. These results were in good agreement with experimental data. PMID- 15268330 TI - Water in nanopores. I. Coexistence curves from Gibbs ensemble Monte Carlo simulations. AB - Coexistence curves of water in cylindrical and slitlike nanopores of different size and water-substrate interaction strength were simulated in the Gibbs ensemble. The two-phase coexistence regions cover a wide range of pore filling level and temperature, including ambient temperature. Five different kinds of two phase coexistence are observed. A single liquid-vapor coexistence is observed in hydrophobic and moderately hydrophilic pores. Surface transitions split from the main liquid-vapor coexistence region, when the water-substrate interaction becomes comparable or stronger than the water-water pair interaction. In this case prewetting, one and two layering transitions were observed. The critical temperature of the first layering transition decreases with strengthening water substrate interaction towards the critical temperature expected for two dimensional systems and is not sensitive to the variation of pore size and shape. Liquid-vapor phase transition in a pore with a wall which is already covered with two water layers is most typical for hydrophilic pores. The critical temperature of this transition is very sensitive to the pore size, in contrast to the liquid vapor critical temperature in hydrophobic pores. The observed rich phase behavior of water in pores evidences that the knowledge of coexistence curves is of crucial importance for the analysis of experimental results and a prerequiste of meaningful simulations. PMID- 15268331 TI - Interfacial tension and wetting in colloid-polymer mixtures. AB - We calculate the interfacial tension and the wetting behavior in phase separated colloid-polymer mixtures both for ideal and excluded volume interacting polymers. Within the recently developed extension of the free volume theory to include polymer interactions the interfacial tension of the free interface is calculated by adding a van der Waals squared gradient term. The wetting behavior at a hard wall is calculated following a Cahn-Fisher-Nakanishi approach taking the one- and two-body colloid-wall interactions into account. Comparing results for interacting polymers with those for ideal polymers we find that for interacting polymers the interfacial tension does not increase as steeply as a function of the gas-liquid colloid density difference. Furthermore, the wetting transition shifts to higher polymer concentrations, even to above the triple line. The predictions for both the interfacial tension and the wetting are compared to recent experiments. PMID- 15268332 TI - Modeling liquid crystal bilayer structures with minimal surfaces. AB - This paper describes a new convenient and accurate method of calculating x-ray diffraction integrated intensities from detailed cubic bilayer structures. The method is employed to investigate the structure of a particular surfactant system (didodecyldimethylammonium bromide in a solution of oil and heavy water), for which single-crystal experimental data have recently been collected. The diffracted peak intensities correlate well with theoretical structures based on mathematical minimal surfaces. Optimized electron density profiles of the bilayer are presented, providing new insight into key features of the bilayer structure. PMID- 15268333 TI - Optical control over photoconductivity in polyferrocenylsilane films. AB - We report the study and elucidate the origin of the photoconductivity of polyferrocenylsilanes achieved through photooxidation performed by ultraviolet irradiation in the presence of chloroform. The persistence over months of the changes in the optoelectronic properties allowed more detailed studies of the charge photogeneration process. The photocurrent spectrum mimics that of the absorption indicating that the photooxidized material is not a mechanical mixture of oxidized and unoxidized polymer units. Photomodulation spectroscopy revealed the existence of long-lived photoexcited states with a lifetime in the millisecond range. They have been interpreted as trapped excitons at the oxidized sites where the polymer is deformed due to the presence of the chloroform derived counter ions. Because of the relatively long lifetime of the trapped excitons they can dissociate and the formed charge carriers can be separated in an externally applied electric field. The effect of the polymer chain deformation around the oxidized unit extends over the neighboring polymer units. In light of the potential applications of this class of polymers in various electronic and photonic devices, the clarification of such a basic process as the photocurrent generation will be a key factor for further technological development. PMID- 15268334 TI - Squeezing wetting and nonwetting liquids. AB - We present molecular-dynamics results for the squeezing of octane (C8H18) between two approaching solid elastic walls with different wetting properties. The interaction energy between the octane bead units and the solid walls is varied from a very small value (1 meV), corresponding to a nonwetting surface with a very large contact angle (nearly 180 degrees), to a high value (18.6 meV) corresponding to complete wetting. When at least one of the solid walls is wetted by octane we observe well defined molecular layers develop in the lubricant film when the thickness of the film is of the order of a few atomic diameters. An external squeezing-pressure induces discontinuous, thermally activated changes in the number n of lubricant layers (n-->n-1 layering transitions). With increasing interaction energy between the octane bead units and the solid walls, the transitions from n to n-1 layers occur at higher average pressure. This results from the increasing activation barrier to nucleate the squeeze-out with increasing lubricant-wall binding energy (per unit surface area) in the contact zone. Thus, strongly wetting lubricant fluids are better boundary lubricants than the less wetting ones, and this should result in less wear. We analyze in detail the effect of capillary bridge formation (in the wetting case) and droplets formation (in the nonwetting case) on the forces exerted by the lubricant on the walls. For the latter case small liquid droplets may be trapped at the interface, resulting in a repulsive force between the walls during squeezing, until the solid walls come into direct contact, where the wall-wall interaction may be initially attractive. This effect is made use of in some practical applications, and we give one illustration involving conditioners for hair care application. PMID- 15268335 TI - The van der Waals interaction between protein molecules in an electrolyte solution. AB - In this report we present a general formulation to calculate the van der Waals interaction between two protein molecules in an electrolyte solution using boundary element method of solving linearized Poisson-Boltzmann equation. Our formulation is based upon an inhomogeneous dielectric model of proteins at the residue level. Our results for bovine pancreatic trypsin inhibitor at various relative orientations indicate that the anisotropy of the interaction can be tens of kBT. PMID- 15268336 TI - Thermodynamics of soft anisotropic interfaces. AB - The Gibbs-Duhem equation for interfaces between nematic liquid crystals and isotropic fluids is formulated and shown to be a generic equation for soft anisotropic surfaces. The one-to-one correspondence between the nematic and crystalline surface Gibbs-Duhem equations is established. Consistency between the surface Gibbs-Duhem equation and the classical equations of interfacial nematostatics is shown. Using a phase space that takes into account thermodynamics, liquid crystalline order, and geometric variables, the generalized nematic surface Gibbs-Duhem equation reveals the presence of couplings between shape, adsorption, temperature, and average molecular orientation. Merging the thermodynamic analysis with nematostatics results in a model for morphactancy, that is, adsorption-induced interfacial shape selection. The specific roles of gradient bulk Frank elasticity, interfacial tension, and anchoring energy are elucidated by analyzing particular paths in the thermodynamic-geometric phase space. PMID- 15268337 TI - Effect of pressure on dynamic heterogeneity in dendrimeric alkyd resin. AB - Broadband dielectric spectroscopy is employed to investigate the non-Debye relaxation behavior in a dendrimeric alkyd resin. From temperature-dependent measurements at ambient pressure, we found a very broad distribution of relaxation times. This is attributed to the complex geometrical topology of the molecule. However, compression significantly reduces the non-Debye character of the dielectric response; thus, pressure induces dynamic homogeneity in the dendrimeric alkyd resin. PMID- 15268338 TI - Coarse grained models for flexible liquid crystals: parameterization of the bond fluctuation model. AB - We extend the bond fluctuation model, originally devised to investigate polymer systems, to contain anisotropic interactions suitable for the simulation of large flexible molecules such as liquid crystalline polymers and dendrimers. This extended model coarse grains the interaction between the flexible chains at a similar level of detail to the mesogenic units. Suitable interaction parameters are obtained by performing trial simulations on a low molar mass liquid crystalline system. The phase diagram of this system is determined as a function of the molecular stiffness. The nematic to isotropic transition temperature is found to increase with increasing stiffness. PMID- 15268339 TI - Polymers confined between two parallel plane walls. AB - Single three-dimensional polymers confined to a slab, i.e., to the region between two parallel plane walls, are studied by Monte Carlo simulations. They are described by N-step walks on a simple cubic lattice confined to the region 1< or = z < or = D. The simulations cover both regions D<>RF (where RF approximately Nnu is the Flory radius, with nu approximately 0.587), as well as the cross-over region in between. Chain lengths are up to N=80 000, slab widths up to D=120. In order to test the analysis program and to check for finite size corrections, we actually studied three different models: (a) ordinary random walks (mimicking Theta polymers); (b) self-avoiding walks; and (c) Domb-Joyce walks with the self-repulsion tuned to the point where finite size corrections for free (unrestricted) chains are minimal. For the simulations we employ the pruned-enriched-Rosenbluth method with Markovian anticipation. In addition to the partition sum (which gives us a direct estimate of the forces exerted onto the walls), we measure the density profiles of monomers and of end points transverse to the slab, and the radial extent of the chain parallel to the walls. All scaling laws and some of the universal amplitude ratios are compared to theoretical predictions. PMID- 15268340 TI - Theoretical study of the second-order nonlinear optical properties of [N]helicenes and [N]phenylenes. AB - The second-order nonlinear optical properties of helicenes and phenylenes have been theoretically investigated at the time-dependent Hartree-Fock level using the Austin model 1 semiempirical Hamiltonian. Both the antisymmetric isotropic component of the first hyperpolarizability (beta) and its projection on the dipole moment (beta(parallel)) have been determined for increasingly large helical systems as well as for their analogs substituted by donor/acceptor pairs. It is found that (i) in nonsubstituted helicenes and phenylenes, beta increases monotonically with the size of the system and slightly depends on the nature of the helix; (ii) the corresponding beta(parallel) is mostly determined by the radial component of the first hyperpolarizability vector; (iii) in helicenes, beta(parallel) is positive and presents quasiperiodic oscillations with the helix; (iv) in phenylenes, beta(parallel) depends upon the size of the helix and it can be either positive or negative as a result of the differences in evolution with N of the radial components of the dipole moment and first hyperpolarizability. Substituting the helicenes and phenylenes by the prototypical NH2/NO2 donor/acceptor pair provides a diversity of effects on beta and beta(parallel) that encompasses decrease, increase, and change in sign. PMID- 15268341 TI - Box length search algorithm for molecular simulation of systems containing periodic structures. AB - We have developed a box length search algorithm to efficiently find the appropriate box dimensions for constant-volume molecular simulation of periodic structures. The algorithm works by finding the box lengths that equalize the pressure in each direction while maintaining constant total volume. Maintaining the volume at a fixed value ensures that quantitative comparisons can be made between simulation and experimental, theoretical or other simulation results for systems that are incompressible or nearly incompressible. We test the algorithm on a system of phase-separated block copolymers that has a preferred box length in one dimension. We also describe and test a Monte Carlo algorithm that allows the box lengths to change while maintaining constant volume. We find that the box length search algorithm converges at least two orders of magnitude more quickly than the variable box length Monte Carlo method. Although the box length search algorithm is not ergodic, it successfully finds the box length that minimizes the free energy of the system. We verify this by examining the free energy as determined by the Monte Carlo simulation. PMID- 15268342 TI - Continuum description of ionic and dielectric shielding for molecular-dynamics simulations of proteins in solution. AB - We extend our continuum description of solvent dielectrics in molecular-dynamics (MD) simulations, which has provided an efficient and accurate solution of the Poisson equation, to ionic solvents as described by the linearized Poisson Boltzmann (LPB) equation. We start with the formulation of a general theory for the electrostatics of an arbitrarily shaped molecular system, which consists of partially charged atoms and is embedded in a LPB continuum. This theory represents the reaction field induced by the continuum in terms of charge and dipole densities localized within the molecular system. Because these densities cannot be calculated analytically for systems of arbitrary shape, we introduce an atom-based discretization and a set of carefully designed approximations. This allows us to represent the densities by charges and dipoles located at the atoms. Coupled systems of linear equations determine these multipoles and can be rapidly solved by iteration during a MD simulation. The multipoles yield the reaction field forces and energies. Finally, we scrutinize the quality of our approach by comparisons with an analytical solution restricted to perfectly spherical systems and with results of a finite difference method. PMID- 15268343 TI - Geometric structure and electronic properties of neutral and anionic Fe2C3 and Fe2C4 clusters, as obtained by density-functional calculations. AB - We calculated the geometrical structures and electronic properties of neutral and anionic Fe2Cn clusters (n = 3,4) using a density-functional method that employs linear combinations of atomic orbitals as basis sets, standard nonlocal norm conserving pseudopotentials, and the generalized gradient approximation to exchange and correlation. We show that the ground-state structures of Fe2C3 and Fe2C4 are essentially the same in the neutral and anionic states, namely, planar rings that feature nonadjacent Fe atoms. For the anionic clusters, these findings contrast with previously published results. PMID- 15268344 TI - Binary liquid mixtures in porous solids. AB - The technique of nuclear magnetic resonance cryoporometry has been used to study the behavior of binary liquid mixtures of water and decane in porous sol-gel silicas. It was observed that the water preferentially adsorbed onto the silica surface and so was able to displace the decane from the pores. PMID- 15268345 TI - Strongly cluster size dependent reaction behavior of CO with O2 on free silver cluster anions. AB - Reactions of free silver anions Agn- (n = 1 - 13) with O2, CO, and their mixtures are investigated in a temperature controlled radio frequency ion trap setup. Cluster anions Agn- (n = 1 - 11) readily react with molecular oxygen to yield AgnOm- (m = 2, 4, or 6) oxide products. In contrast, no reaction of the silver cluster anions with carbon monoxide is detected. However, if silver cluster anions are exposed to the mixture of O2 and CO, new reaction products and a pronounced, discontinuous size dependence in the reaction behavior is observed. In particular, coadsorption complexes Agn(CO)O2- are detected for cluster sizes with n = 4 and 6 and, the most striking observation, in the case of the larger odd atom number clusters Ag7-, Ag9-, and Ag11-, the oxide product concentration decreases while a reappearance of the bare metal cluster signal is observed. This leads to the conclusion that carbon monoxide reacts with the activated oxygen on these silver clusters and indicates the prevalence of a catalytic reaction cycle. PMID- 15268346 TI - A doubly nudged elastic band method for finding transition states. AB - A modification of the nudged elastic band (NEB) method is presented that enables stable optimizations to be run using both the limited-memory Broyden-Fletcher Goldfarb-Shanno (L-BFGS) quasi-Newton and slow-response quenched velocity Verlet minimizers. The performance of this new "doubly nudged" DNEB method is analyzed in conjunction with both minimizers and compared with previous NEB formulations. We find that the fastest DNEB approach (DNEB/L-BFGS) can be quicker by up to 2 orders of magnitude. Applications to permutational rearrangements of the seven atom Lennard-Jones cluster (LJ7) and highly cooperative rearrangements of LJ38 and LJ75 are presented. We also outline an updated algorithm for constructing complicated multi-step pathways using successive DNEB runs. PMID- 15268347 TI - The reduced density matrix method for electronic structure calculations and the role of three-index representability conditions. AB - The variational approach for electronic structure based on the two-body reduced density matrix is studied, incorporating two representability conditions beyond the previously used P, Q, and G conditions. The additional conditions (called T1 and T2 here) are implicit in the work of Erdahl [Int. J. Quantum Chem. 13, 697 (1978)] and extend the well-known three-index diagonal conditions also known as the Weinhold-Wilson inequalities. The resulting optimization problem is a semidefinite program, a convex optimization problem for which computational methods have greatly advanced during the past decade. Formulating the reduced density matrix computation using the standard dual formulation of semidefinite programming, as opposed to the primal one, results in substantial computational savings and makes it possible to study larger systems than was done previously. Calculations of the ground state energy and the dipole moment are reported for 47 different systems, in each case using an STO-6G basis set and comparing with Hartree-Fock, singly and doubly substituted configuration interaction, Brueckner doubles (with triples), coupled cluster singles and doubles with perturbational treatment of triples, and full configuration interaction calculations. It is found that the use of the T1 and T2 conditions gives a significant improvement over just the P, Q, and G conditions, and provides in all cases that we have studied more accurate results than the other mentioned approximations. PMID- 15268348 TI - Current-dependent extension of the Perdew-Burke-Ernzerhof exchange-correlation functional. AB - The probability current density is used in addition to the electron density and its gradient as a variable in the construction of an exchange-correlation functional. Starting from the Perdew-Burke-Ernzerhof generalized gradient approximation, we employ exact conditions to build a nonempirical exchange functional. Matching the correlation functional to that for exchange yields a current-dependent approximation for correlation. The resulting functional is given in a simple closed form. Application of this approximation to open shell atoms eliminates the artificial level splitting of formally degenerate states observed with generalized gradient approximations. PMID- 15268349 TI - Stereographic projections path integral for inertia ellipsoids: applications to Arn-HF clusters. AB - The DeWitt formula for inertia ellipsoids mapped by stereographic projection coordinates is developed. We discover that by remapping the quaternion parameter space with stereographic projections, considerable simplification of the differential geometry for the inertia ellipsoid with spherical symmetry takes place. The metric tensor is diagonal and contains only one independent element in that case. We find no difficulties testing and implementing the DeWitt formula for the inertia ellipsoids of asymmetric tops mapped by stereographic projections. The path integral algorithm for the treatment of Rm x S2 manifolds based on a mixture of Cartesian and stereographic projection coordinates is tested for small Arn-HF clusters in the n = 2 to n = 5 range. In particular, we determine the quantum effects of the red shift and the isomerization patterns at finite temperatures. Our findings are consistent with previously reported computations and experimental data for small Arn-HF clusters. PMID- 15268350 TI - Constrained fluid lambda-integration: constructing a reversible thermodynamic path between the solid and liquid state. AB - A novel lambda-integration path is proposed for calculating the Gibbs free energy difference between any arbitrary solid and liquid state needed for the location of melting lines. This technique involves reversibly forcing a liquid state to a solid state across the phase transition along a nonphysical path, thermodynamically coupling the two states directly. The process eliminates the need for coupling to idealized reference states as is presently performed and hence simplifies the location of phase transitions for computer simulation systems. More specifically the path involves a three stage process, whereby, initially a liquid state is transformed to a weakly attractive fluid using linear lambda-integration scaling of the intermolecular potential. In the second stage, the resulting fluid is then constrained to the required solid configurational phase space via the insertion of a periodic lattice of 3D Gaussian wells. The final stage involves reversing to full strength the main intermolecular potential while gradually turning off the constraining 3D Gaussian lattice finally resulting in a stable (or metastable) solid state. Each stage was found to be completely reversible and the resulting change in free energy was thermodynamically integrable. The methodology is demonstrated and validated by calculating solid-liquid coexistence points using the new technique and comparing to those in present literature for the truncated and shifted Lennard-Jones system. The results are found to be in good agreement. The new method is not limited to melting phase transitions and is readily applicable to any simulation methodology, simulation cell size and/or intermolecular potential including ab initio methods. PMID- 15268351 TI - Relativistic spin-orbit effects on hyperfine coupling tensors by density functional theory. AB - A second-order perturbation theory treatment of spin-orbit corrections to hyperfine coupling tensors has been implemented within a density-functional framework. The method uses the all-electron atomic mean-field approximation and/or spin-orbit pseudopotentials in incorporating one- and two-electron spin orbit interaction within a first-principles framework. Validation of the approach on a set of main-group radicals and transition metal complexes indicates good agreement between all-electron and pseudopotential results for hyperfine coupling constants of the lighter nuclei in the system, except for cases in which scalar relativistic effects become important. The nonrelativistic Fermi contact part of the isotropic hyperfine coupling constants is not always accurately reproduced by the exchange-correlation functionals employed, particularly for the triplet and pi-type doublet radicals in the present work. For this reason, ab initio coupled cluster singles and doubles with perturbative triples results for the first-order contributions have been combined in the validation calculations with the density functional results for the second-order spin-orbit contributions. In the cases where spin-orbit corrections are of significant magnitude relative to the nonrelativistic first-order terms, they improve the agreement with experiment. Antisymmetric contributions to the hyperfine tensor arise from the spin-orbit contributions and are discussed for the IO2 radical, whereas rovibrational effects have been evaluated for RhC, NBr, and NI. PMID- 15268352 TI - A second quantization formulation of multimode dynamics. AB - A new formalism for calculating and analyzing many-mode quantum dynamics is presented. The formalism is similar in spirit to the second quantization formulation of electronic structure theory. The similarity means that similar techniques can be employed for calculating the many-mode nuclear wave function. As a consequence a new formulation of the vibrational self-consistent-field (VSCF) method can be developed. Another result is that the formalism opens up for the construction of new methods that go beyond the VSCF level. A vibrational coupled cluster (VCC) theory is constructed using the new formalism. The size extensivity concept is introduced in the context of multimode wave functions and the size extensivity of approximate VCC methods is illustrated in comparison with the non-size-extensive vibrational configuration interaction method. PMID- 15268353 TI - Vibrational coupled cluster theory. AB - The theory and first implementation of a vibrational coupled cluster (VCC) method for calculations of the vibrational structure of molecules is presented. Different methods for introducing approximate VCC methods are discussed including truncation according to a maximum number of simultaneous mode excitations as well as an interaction space order concept is introduced. The theory is tested on calculation of anharmonic frequencies for a three-mode model system and a formaldehyde quartic force field. The VCC method is compared to vibrational self consistent-field, vibrational Moller-Plesset perturbation theory, and vibrational configuration interaction (VCI). A VCC calculation typically gives higher accuracy than a corresponding VCI calculation with the same number of parameters and the same formal operation count. PMID- 15268354 TI - Path integral hybrid Monte Carlo algorithm for correlated Bose fluids. AB - Path integral hybrid Monte Carlo (PIHMC) algorithm for strongly correlated Bose fluids has been developed. This is an extended version of our previous method [S. Miura and S. Okazaki, Chem. Phys. Lett. 308, 115 (1999)] applied to a model system consisting of noninteracting bosons. Our PIHMC method for the correlated Bose fluids is constituted of two trial moves to sample path-variables describing system coordinates along imaginary time and a permutation of particle labels giving a boundary condition with respect to imaginary time. The path-variables for a given permutation are generated by a hybrid Monte Carlo method based on path integral molecular dynamics techniques. Equations of motion for the path variables are formulated on the basis of a collective coordinate representation of the path, staging variables, to enhance the sampling efficiency. The permutation sampling to satisfy Bose-Einstein statistics is performed using the multilevel Metropolis method developed by Ceperley and Pollock [Phys. Rev. Lett. 56, 351 (1986)]. Our PIHMC method has successfully been applied to liquid helium 4 at a state point where the system is in a superfluid phase. Parameters determining the sampling efficiency are optimized in such a way that correlation among successive PIHMC steps is minimized. PMID- 15268355 TI - On-the-fly localization of electronic orbitals in Car-Parrinello molecular dynamics. AB - The ab initio molecular-dynamics formalism of Car and Parrinello is extended to preserve the locality of the orbitals. The supplementary term in the Lagrangian does not affect the nuclear dynamics, but ensures "on the fly" localization of the electronic orbitals within a periodic supercell in the Gamma-point approximation. The relationship between the resulting equations of motion and the formation of a gauge-invariant Lagrangian combined with a gauge-fixing procedure is briefly discussed. The equations of motion can be used to generate a very stable and easy to implement numerical integration algorithm. It is demonstrated that this algorithm can be used to compute the trajectory of the maximally localized orbitals, known as Wannier orbitals, in ab initio molecular dynamics with only a modest increase in the overall computer time. In the present paper, the new method is implemented within the generalized gradient approximation to Kohn-Sham density-functional theory employing plane wave basis sets and atomic pseudopotentials. In the course of the presentation, we briefly discuss how the present approach can be combined with localized basis sets to design fast linear scaling ab initio molecular-dynamics methods. PMID- 15268356 TI - The open-shell interaction of He with the B 3Piu(0+) state of Br2: an ab initio study and its comparison with a diatomics-in-molecule perturbation model. AB - The interaction of He with Br2 in electronically excited B 3Piu state is investigated using spin-unrestricted single and double coupled-cluster approach with noniterative perturbative treatment of triple excitations. Internal electrons of the Br atom are described by effective core pseudopotentials. The validity of this approach is analyzed by comparing the lowest 2Sigma+ and 2Pi electronic states of the HeBr molecule with those obtained in all electron calculations [J. Chem. Phys. 115, 10438 (2001)]. In this context, we examine the performance of different basis sets and saturation with bond functions. The comparison of theoretical blue-shifts with the experiment provides confidence about the present ab initio calculations. In addition, He-Br results of ab initio calculations at the same level are used to obtain approximate He-Br2 (3Piu) interactions in the framework of the diatomics-in-molecule first order perturbation theory (IDIM-PT1) [J. Chem. Phys. 104, 9913 (1996)]. Overall, the IDIM-PT1 model results show a good agreement with the ab initio ones, being the main difference the sensitivity to the elongation of the Br-Br bond. PMID- 15268357 TI - Photodissociation dynamics of the methyl radical at 212.5 nm: effect of parent internal excitation. AB - Photodissociation dynamics of the CH3 radical at 212.5 nm has been investigated using the H atom Rydberg tagging time-of-flight method with a pure CH3 radical source generated by the photolysis of CH3I at 266 nm. Time-of-flight spectra of the H atom products from the photolysis of both cold and hot methyl radicals have been measured at different photolysis polarizations. Experimental results indicate that the photodissociation of the methyl radical in its ground vibrational state at 212.5 nm excitation occurs on a very fast time scale in comparison with its rotational period, indicating the CH3 dissociation at 212.5 nm occurs on the excited 3s Rydberg state surface. Experimental evidence also shows that the photodissociation of the methyl radical in the nu2 = 1 state of the umbrella mode at 212.5 nm excitation is characteristically different from that in the ground vibrational state. PMID- 15268358 TI - Electron localization-delocalization transitions in dissociation of the C4- anion: a large-D analysis. AB - We present a study, employing high level ab initio methods, of electron localization-delocalization transitions along the dissociation path of the C4- anion to C2 and C2-. We find that at the equilibrium geometry, the symmetrical and nonsymmetrical configurations of the linear C4- anion are almost isoenergetic. However, along a collinear dissociation path, the dipole moment drops abruptly to zero when the separation between the two middle carbon nuclei reaches about R = 2.15 angstroms. The dipole moment remains zero until about R = 2.78 angstroms, and then continuously increases as dissociation proceeds. This behavior is analogous to critical phenomena: The abrupt drop to zero of the dipole moment resembles a first-order phase transition, the later steady rise resembles a continuous phase transition. We show that a simple sub-Hamiltonian model, corresponding to the large-dimension limit for an electron in the field of four collinear carbon atoms, exhibits both kinds of phase transitions along the dissociation path. PMID- 15268359 TI - Hydrates of the most stable gas-phase mono- and di-protonated glycine derivatives: origin of no reservation energy bond in glycine-2H2+. AB - Series of hydrates of the most stable glycine-H+/2H2+ in the gas phase are presented at the B3LYP level. The results show that only the amino hydrogens and hydroxyl hydrogens can be monohydrated for the glycine-H+, and the amino hydrogens are preferred. The H6(O4) of glycine-2H2+ is the best site for a water molecule to attach, i.e., the corresponding hydrate is the most stable one among its isomers. Calculations reveal that the binding energies of hydrated hydrogens decrease relative to their counterparts in the isolated glycine-H+/2H2+ complexes and they are positive values and without proton transfer except those of monohydrated glycine-2H2+ complexes with the combination modes of H3O+...(glycine H+). The complex H3O+...(glycine-H+) is formed by the combination of a H2O molecule and one hydroxyl-site proton of glycine-2H2+, and with the proton transfer to H2O. Here the interaction between the proton of H3O+ and the glycine H+ mainly depends on an electronic one instead of an initial covalent one of the isolated glycine-2H2+. The generation of the bond between the H3O+ and the glycine-H+ makes the energy of the complex higher than the energy sum of its two separated species (or two reactants of the complex), just like the case of M+...(glycine-H+) bond (M = Li,Na). The observation can explain satisfactorily why the combinations of both a proton and an alkali ion or two alkali ions to a glycine molecule can make the corresponding complex hold reservation energy bond(s), while the combination of two protons and a glycine in our previous work cannot [H. Ai et al., J. Chem. Phys. 117, 7593 (2002)]. For the glycine-2H2+, monohydration at the any site of its amino hydrogens can make the binding strength of any other neighboring proton (hydrogens) stronger relative to its counterpart in the isolated glycine-2H2+. Further hydration, especially at the site of either of hydroxyl hydrogens, would disfavor the reservation energy of the system. PMID- 15268360 TI - A combined crossed beam and theoretical investigation of O(3P) + C3H3 --> C3H2 + OH. AB - The radical-radical reaction dynamics of ground-state atomic oxygen [O(3P)] with propargyl radicals (C3H3) has first been investigated in a crossed beam configuration. The radical reactants O(3P) and C3H3 were produced by the photodissociation of NO2 and the supersonic flash pyrolysis of precursor propargyl bromide, respectively. A new exothermic channel of O(3P) + C3H3 --> C3H2 + OH was identified and the nascent distributions of the product OH in the ground vibrational state (X 2Pi:nu" = 0) showed bimodal rotational excitations composed of the low- and high-N" components without spin-orbit propensities. The averaged ratios of Pi(A')/Pi(A") were determined to be 0.60 +/- 0.28. With the aid of ab initio theory it is predicted that on the lowest doublet potential energy surface, the reaction proceeds via the addition complexes formed through the barrierless addition of O(3P) to C3H3. The common direct abstraction pathway through a collinear geometry does not occur due to the high entrance barrier in our low collision energy regime. In addition, the major reaction channel is calculated to be the formation of propynal (CHCCHO) + H, and the counterpart C3H2 of the probed OH product in the title reaction is cyclopropenylidene (1c-C3H2) after considering the factors of barrier height, reaction enthalpy and structural features of the intermediates formed along the reaction coordinate. On the basis of the statistical prior and rotational surprisal analyses, the ratio of population partitioning for the low- and high-N" is found to be about 1:2, and the reaction is described in terms of two competing addition-complex mechanisms: a major short-lived dynamic complex and a minor long-lived statistical complex. The observed unusual reaction mechanism stands in sharp contrast with the reaction of O(3P) with allyl radical (C3H5), a second significant conjugated hydrocarbon radical, which shows totally dynamic processes [J. Chem. Phys. 117, 2017 (2002)], and should be understood based upon the characteristic electronic structures and reactivity of the intermediates on the potential energy surface. PMID- 15268361 TI - Kinetics of C2(a3Piu) radical reactions with alkanes by LIF. AB - The reactions of C2(a3Piu) radicals with a series of alkanes have been studied at room temperature and 6.5 torr total pressure using the pulsed laser photolysis/laser-induced fluorescence technique. C2(a3Piu) radicals were generated by photolysis of C2Cl4 with the focused output from the fourth harmonic of a Nd: YAG laser at 266 nm. The relative concentration of C2(a3Piu) radicals was monitored on the (0,0) band of the C2(d3Pig <-- a3Piu) transition at 516.5 nm by laser-induced fluorescence. From the analysis of the relative concentration time behavior of C2(a3Piu) under pseudofirst-order conditions, the rate constants for the reactions of C2(a3Piu) with alkanes (C1-C8) were determined. The rate constant increases linearly with the increasing of the number of CH2 groups in the alkanes. The experimental results indicate that the reaction of C2(a3Piu) with small alkanes (C1-C8) follows the typical hydrogen abstraction process. Based on the correlation of the experimental results with the bond dissociation energy of the alkanes, the reactions of C2(a3Piu) with small alkanes likely proceed via the mechanism of hydrogen abstraction. PMID- 15268362 TI - Imaging the quantum-state specific differential cross sections of HCl formed from reactions of chlorine atoms with methanol and dimethyl ether. AB - Center-of-mass frame scattering angle distributions obtained directly from crossed molecular beam velocity map images are reported for HCl formed in different rotational levels of its vibrational ground state by reaction of Cl atoms with CH3OH and CH3OCH3. Products are observed to scatter over all angles, with peaks in the distribution in the forward and backward directions (theta = 0 and 180 degrees with respect to the relative velocity vectors of the Cl atoms). Products of both reactions exhibit differential cross sections that vary with the rotational quantum number of the HCl, with a greater propensity for forward scatter for J = 2, shifting to more pronounced backward scatter for J = 5. This trend is, however, more evident for reaction of dimethyl ether than for methanol. The mean fractions of the available energy channeled into product kinetic energy vary with scattering angle, but the angle-averaged fractions are, respectively, 0.37 and 0.42 for the methanol and dimethyl ether reactions. On average, 46% or more of the available energy of the reactions becomes internal energy of the radical co-product. Results are interpreted with the aid of computed energies of transition states and molecular complexes along the reaction pathways, and comparisons are drawn with recent measurements of the scattering distributions and energy release for reactions of Cl atoms with small alkanes. PMID- 15268363 TI - Measurements and simulations of high energy O(3P) + Ar(1S) angular scattering: single and multi-collision regimes. AB - We present differential angular cross sections for O(3P) + Ar(1S) scattering at collision energies near 90 kcal mol(-1) (approximately 8 km s(-1) relative velocity) from molecular beam measurements and high-level theoretical calculations. Beams of hyperthermal O(3P) are now being used to investigate novel gas-phase and gas-surface chemistries, and the comparison of theory and measurements on this simple system will be a stringent test of the experimental methodology. Potential energy curves were generated for O(3P) + Ar(1S) using a large cc-pVQZ basis within a valence multi-configuration plus perturbation theory treatment. These curves were then used in quantum scattering calculations to generate differential cross sections. Agreement between experiment and theory is excellent. In addition to these comparisons, the cross sections were used in direct simulation Monte Carlo calculations to investigate effects of increasing the Ar flux above the "single-collision" regime. As the Ar flux increases, the observed differential angular cross sections change in two ways. In addition to the main "single-scatter" peak along the incident O-atom beam direction, a secondary O-atom peak appears in the direction of the incident Ar beam, and the multiple-scattered O-atom translational energy starts to reflect the energy of the relatively slow moving Ar beam. PMID- 15268364 TI - Quantum reactive scattering with a transmission-free absorbing potential. AB - A recently derived transmission-free absorbing potential is applied to the study of atom-diatom chemical reactions. This absorbing potential only depends on a single parameter--the width of the absorbing region--and its reflection properties are guaranteed to improve as this parameter is increased. Converged results can therefore be obtained very easily, as we illustrate with time dependent wave packet calculations on the H + H2,F + H2, and H + O2 reactions. PMID- 15268365 TI - Anomalous splittings of torsional sublevels induced by the aldehyde inversion motion in the S1 state of acetaldehyde. AB - The G6 group-theoretical high-barrier formalism developed previously for internally rotating and inverting CH3NHD is used to interpret the abnormal torsional splittings in the S1 state of acetaldehyde for levels 14(0-)15(0), 14(0 )15(1), and 14(0-)15(2), where 14(0-) denotes the upper inversion tunneling component of the aldehyde hydrogen and 15 denotes the methyl torsional vibration. This formalism, derived using an extended permutation-inversion group G6m, treats simultaneously methyl torsional tunneling, aldehyde-hydrogen inversion tunneling and overall rotation. Fits to the rotational states of the four pairs of inversion-torsion vibrational levels (14(0+)15(0A,E), 14(0-)15(0A,E)), (14(0+)15(1A,E), 14(0-)15(1A,E)), (14(0+)15(2A,E), 14(0-)15(2A,E)), and (14(0+)15(3A,E), 14(0-)15(3A,E)) are performed, giving root-mean-square deviations of 0.003, 0.004, 0.004, and 0.004 cm(-1), respectively, which are nearly equal to the experimental uncertainty of 0.003 cm(-1). For torsional levels lying near the top of the torsional barrier, this theoretical model, after including higher-order terms, provides satisfactory fits to the experimental data. The partially anomalous K-doublet structure of the S1 state, which deviates from that in a simple torsion-rotation molecule, is fitted using this formalism and is shown to arise from coupling of torsion and rotation motion with the aldehyde-hydrogen inversion. PMID- 15268366 TI - Full-dimensional quantum calculations of vibrational spectra of six-atom molecules. I. Theory and numerical results. AB - Two quantum mechanical Hamiltonians have been derived in orthogonal polyspherical coordinates, which can be formed by Jacobi and/or Radau vectors etc., for the study of the vibrational spectra of six-atom molecules. The Hamiltonians are expressed in an explicit Hermitian form in the spatial representation. Their matrix representations are described in both full discrete variable representation (DVR) and mixed DVR/nondirect product finite basis representation (FBR) bases. The two-layer Lanczos iteration algorithm [H.-G. Yu, J. Chem. Phys. 117, 8190 (2002)] is employed to solve the eigenvalue problem of the system. A strategy regarding how to carry out the Hamiltonian-vector products for a high dimensional problem is discussed. By exploiting the inversion symmetry of molecules, a unitary sequential 1D matrix-vector multiplication algorithm is proposed to perform the action of the Hamiltonian on the wavefunction in a symmetrically adapted DVR or FBR basis in the azimuthal angular variables. An application to the vibrational energy levels of the molecular hydrogen trimer (H2)3 in full dimension (12D) is presented. Results show that the rigid-H2 approximation can underestimate the binding energy of the trimer by 27%. Finally, it is demonstrated that the two-layer Lanczos algorithm is also capable of computing the eigenvectors of the system with minor effort. PMID- 15268367 TI - State-to-state rotational rate constants for CO + He: infrared double resonance measurements and simulation of the data using the SAPT theoretical potential energy surface. AB - An extensive data set of 54 time-resolved pump-probe measurements was used to examine CO + He rotational energy transfer within the CO v = 2 rotational manifold. Rotational levels in the range Ji = 2-9 were excited and collisional energy transfer of population to the levels Jf = 1-10 was monitored. The resulting data set was analyzed by fitting to numerical solutions of the master equation. State-to-state rate constant matrices were generated using fitting law functions and ab initio theoretical calculations that employed the SAPT potential energy surface of Heijmen et al. [J. Chem. Phys. 107, 9921 (1997)]. Fitting laws based on the modified exponential gap (MEG), statistical power exponential gap (SPEG), and energy corrected sudden with exponential power (ECS-EP) models all yielded acceptable simulations of the kinetic data, as did the theoretical rate constants. However, the latter were unique in their ability to reproduce both our kinetic data and the pressure broadening coefficients for CO + He. These results provide an impressive demonstration of the quality of the symmetry adapted perturbation theory (SAPT) potential energy surface. PMID- 15268368 TI - Quantum-mechanical theory of atom-molecule and molecular collisions in a magnetic field: spin depolarization. AB - A theory for quantum-mechanical calculations of cross sections for atom-molecule and molecular collisions in a magnetic field is presented. The formalism is based on the representation of the wave function as an expansion in a fully uncoupled space-fixed basis. The systems considered include 1S-atom-2Sigma-molecule, 1S atom-3Sigma-molecule, 2Sigma-molecule-2Sigma-molecule, and 3Sigma-molecule-3Sigma molecule. The theory is used to elucidate the mechanisms for collisionally induced spin depolarization. PMID- 15268369 TI - Ab initio rate constants from hyperspherical quantum scattering: application to H + CH4 --> H2 + CH3. AB - A general and practical procedure is described for calculating rate constants for chemical reactions using a minimal number of ab initio calculations and quantum dynamical computations. The method exploits a smooth interpolating functional developed in the hyperspherical representation. This functional is built from two Morse functions and depends on a relatively small number of parameters with respect to conventional functionals developed to date. Thus only a small number of ab initio points needs to be computed. The method is applied to the H + CH4 - > H2 + CH3 reaction. The quantum scattering calculations are performed treating explicitly the bonds being broken and formed. All the degrees of freedom except the breaking and forming bonds are optimized ab initio and harmonic vibrational frequencies and zero-point energies for them are calculated at the MP2(full) level with a cc-pVTZ basis set. Single point energies are calculated at a higher level of theory with the same basis set, namely CCSD(T, full). We report state-to state cross sections and thermal rate constants for the title reaction and make comparisons with previous results. The calculated rate constants are in good agreement with experiments. PMID- 15268370 TI - Ab initio study including spin-orbit effects on the B-X transition of AgI. AB - The lowest Omega = 0-,0+,1,2 fine-structure potential energy curves arising from the two lowest-lying singlet (X 1Sigma+ and 2 1Sigma+) and the first 3Pi electronic states of AgI were obtained through an effective Hamiltonian; the purely electronic LambdaSSigma energies were used as diagonal elements, which were calculated through extensive complete active space self-consistent field + averaged coupled pair functional calculations, with relativistic effective core potentials and optimized Gaussian basis sets for both atoms. The spin-orbit interactions were included using the Stuttgart effective spin-orbit potentials. For the excited Omega = 0+ states, very strong mixtures were found of the 2 1Sigma+ and 3Pi parents that lead to the fine-structure (0+) single B state (dominated by the 2 1Sigma+ parent at long distance), that explains the B <-- X transitions. The present results also explain the presence of a second long distance minimum for the B0+ state, experimentally Rydberg-Klein-Rees fitted. These calculations produced, as a byproduct, a new lower-lying Omega = 0+ yet unobserved fine-structure state predicted to exist around 22,000 cm(-1). Our theoretical results are compared and discussed in the light of the experimental data for the B-X transitions in silver halides [J. Chem. Phys. 109, 9831 (1998)]. PMID- 15268371 TI - Exciton exciton annihilation dynamics in chromophore complexes. II. Intensity dependent transient absorption of the LH2 antenna system. AB - Using the multiexciton density matrix theory of excitation energy transfer in chromophore complexes developed in a foregoing paper [J. Chem. Phys. 118, 746 (2003)], the computation of ultrafast transient absorption spectra is presented. Beside static disorder and standard mechanisms of excitation energy dissipation the theory incorporates exciton exciton annihilation (EEA) processes. To elucidate signatures of EEA in intensity dependent transient absorption data the approach is applied to the B850 ring of the LH2 found in rhodobacter sphaeroides. As main indications for two-exciton population and resulting EEA we found (i) a weakening of the dominant single-exciton bleaching structure in the transient absorption, and (ii) an intermediate suppression of long-wavelength and short wavelength shoulders around the bleaching structure. The suppression is caused by stimulated emission from the two-exciton to the one-exciton state and the return of the shoulders follows from a depletion of two-exciton population according to EEA. The EEA-signature survives as a short-wavelength shoulder in the transient absorption if orientational and energetic disorder are taken into account. Therefore, the observation of the EEA-signatures should be possible when doing frequency resolved transient absorption experiments with a sufficiently strongly varying pump-pulse intensity. PMID- 15268372 TI - Surface tension of associating fluids by Monte Carlo simulations. AB - Canonical Monte Carlo (NVT-MC) simulations were performed to obtain surface tension and coexistence densities at the liquid-vapor interface of one-site associating Lennard-Jones and hard-core Yukawa fluids, as functions of association strength and temperature. The method to obtain the components of the pressure tensor from NVT-MC simulations was validated by comparing the equation of state of the associative hard sphere system with that coming from isothermal isobaric Monte Carlo simulations. Surface tension of the associative Lennard Jones fluid determined from NVT-MC is compared with previously reported results obtained by molecular dynamics simulations of a pseudomixture model of monomers and dimers. A good agreement was found between both methods. Values of surface tension of associative hard-core Yukawa fluids are presented here for the first time. PMID- 15268373 TI - A self-consistent reaction field model of solvation using distributed multipoles. I. Energy and energy derivatives. AB - The self-consistent reaction field model developed previously by the authors in the case of single center multipole expansion of the electronic structure of the solute has been extended to the case of a distributed multipole expansion. Three different expansions have been tested and two of them have proved to be rapidly convergent. The performances of the code are illustrated by the density functional theory treatment of few test systems: guanine, cytosine, and cytosine hydrated with one and three water molecules. A robust fast computer code has been tested to get the electronic structure, the electrostatic contribution to the solute-solvent free energy of interaction, and the optimized molecular geometry in solution. PMID- 15268374 TI - Vibrational relaxation and coupling of two OH-stretch oscillators with an intramolecular hydrogen bond. AB - We studied the vibrational dynamics of the OH-stretch oscillators of an alcohol with two vicinal OH groups using femtosecond midinfrared pump-probe spectroscopy. The absorption spectrum of pinacol (2,3-dimethyl-2,3-butanediol) in CDCl3 shows two OH-stretch peaks belonging to hydrogen bonded and free OH groups. The anharmonicities of the hydrogen-bonded and free OH-stretch vibrations are 180 and 160 cm(-1), respectively. The lifetime T1 of the OH-stretch vibration is found to be 3.5 +/- 0.4 ps for the hydrogen bonded and 7.4 +/- 0.5 ps for the free OH group. We observed sidebands in the transient spectra after excitation of the bonded OH group, which we attribute to a progression in a low-frequency hydrogen bond mode. The sideband is redshifted 60 cm(-1) with respect to the 0 --> 1 transition. Due to the coupling between the two OH groups and the presence of the sidebands, simultaneous excitation of both OH-stretch vibrations leads to oscillations on the pump-probe signal with frequencies of 40 and 60 cm(-1). PMID- 15268375 TI - Exciton migration dynamics in a dendritic molecule: quantum master equation approach using ab initio molecular orbital configuration interaction method. AB - We investigate the exciton migration dynamics in a dendritic molecular model composed of pi-conjugation linear-leg units (acetylenes and diacetylene) and a benzene ring (branching point) using the quantum master equation approach with the ab initio molecular orbital (MO) configuration interaction (CI) method. The efficient migration of exciton from short-length linear legs (acetylenes) to long length linear leg (diacetylene) via a benzene ring is observed. As predicted in previous studies, the exciton (electron and hole) distributions are relatively well localized in each generation segmented by the meta-branching point (meta substituted benzene ring) though the electron and hole distributions are delocalized and are somewhat spatially different from each other within each generation. It is found that the excitons localized in the generation composed of short linear legs occupy in higher-lying exciton states, while those in the generation composed of long linear legs do in lower-lying ones. These features suggest the decoupling of pi-conjugation at the meta-branching point. On the other hand, the relaxation effect between exciton states is found to be caused by the exciton-phonon coupling, in which the existence of common configurations (electron-hole pairs) in CI wave functions between adjacent exciton states (having primary distributions on short and long linear-leg regions, respectively) is important for the relaxation between their exciton states. This feature indicates the importance of partial penetration of pi-conjugation through the meta-substituted benzene ring in excited states for such exciton migration. PMID- 15268376 TI - Molecular dynamics of oligofluorenes: a dielectric spectroscopy investigation. AB - The molecular dynamics were investigated in a series of "defect-free" oligofluorenes up to the polymer by dielectric spectroscopy (DS). The method is very sensitive to the presence of keto "defects" that when incorporated on the backbone give rise to poor optical and electronic properties. Two dielectrically active processes were found (beta and alpha process). The latter process (alpha) displays strongly temperature dependent relaxation times and temperature- and molecular weight-dependent spectral broadening associated with intramolecular correlations. The glass temperature (Tg) obeys the Fox-Flory equation and the polymer Tg is obtained by DS at 332 K. The effective dipole moment associated with the alpha process is 0.27 +/- 0.03 D. PMID- 15268377 TI - Structures of BaF2-CaF2 heterolayers and their influences on ionic conductivity. AB - Recently, artificial ion conductors have been prepared by growing epitaxial heterolayers consisting of BaF2-CaF2 using molecular beam epitaxy. The ionic conductivity of these heterolayers shows a strong dependence on the layer thickness [N. Sata, S. Eberman, K. Eberl, and J. Maier, Nature 408, 996 (2000)]. In this paper three such heterolayers with different spacings (sample A: 80 nm, sample B: 10 nm, sample C: 1 nm) are investigated by conventional transmission electron microscopy and high-resolution transmission electron microscopy. The spacings are chosen such that they fall into the three conductivity regimes observed in N. Sata et al. (l > 50 nm; 8 < l < 50 nm; l < 8 nm). In accordance with conductivity studies, the samples with spacings of 10 nm or greater (A,B) are epitaxial and continuous, whereas in the case of extremely small spacing (C) the continuity of the layers is destroyed by formation of a column-like structure. Analytical electron microscopy reveals that, instead of forming multilayers, Ca and Ba separate in different columns in sample C. The structure properties of sample A (large l) are quite ideal: Planar interfaces with regular arrays of misfit dislocations with their Burgers vectors on the interface are observed. In the case of sample B (medium l) the lattice misfit is accommodated, in addition, by wavy interfaces associated with dislocations characterized by a Burgers vector that makes a large angle to the interfaces. The (111) lattice spacing very close to the interfaces is markedly changed due to this novel relaxation mechanism in the multilayer. The influences of the crystallographic defects on the ionic conductivity are also discussed. PMID- 15268378 TI - On the variation of magnetic susceptibility of a molecular crystal with temperature: the 2,4,6-triphenylverdazyl system. AB - Magnetic susceptibilities of spin-1/2 systems of orthorhombic and higher crystal symmetries have been numerically investigated while taking possible anisotropy in the coupling constants along different crystal axes into account. The work relies on the magnon-based theory of ferromagnetic (FM) and antiferromagnetic (AFM) crystal systems of types FFF, AFF, AAF, and AAA [J. Chem. Phys. 111, 9009 (1999)]. The AAF crystal, in particular, shows interesting changes in the temperature dependence of magnetic susceptibility when the ferromagnetic exchange coupling constant is varied. We especially show that the susceptibility anomalies of molecular crystals fit naturally within the framework of the extended magnon theoretical formalism, and do not necessarily imply a FM --> AFM or a reverse phase transition. A real system, molecular crystal of 2,4,6-triphenylverdazyl (2,4,6-TPV), has been investigated here. It was previously interpreted as an AAF system from observed susceptibility data [Tomiyoshi et al., Phys. Rev. B 49, 16031 (1994)]. The trend of the temperature dependence of magnetic susceptibility studied in the present work also indicates that the crystal belongs to the AAF category with a less prominent FM exchange coupling constant. To reinforce our conclusions, we have adopted a two-pronged strategy. First, the geometry of the 2,4,6-TPV monomer has been optimized here by ab initio unrestricted Hartree-Fock (UHF) calculations using the STO-3G basis set. The optimized geometry is almost planar. A subsequent calculation has been carried out with the phenyl rings twisted out of the plane of the nitrogen atoms. The STO-3G optimized geometry, and the same geometry except for the twisted phenyl rings, have been used to perform ab initio coupled-cluster (UCCSD-T) calculations with the same basis, and UHF as well as density-functional (UB3LYP) calculations using the 6-31G basis set. The calculated data can easily rationalize the twists while the species remains in crystal. The magnetic category of the crystal has been unambiguously confirmed as AFA from ab initio UHF and UB3LYP calculations of the total energy in different spin states of dimers and trimers along the crystal axes. The computed energy values, however, fail to yield accurate estimates of the exchange coupling constants Ja, Jb, and Jc, because the latter are on the order of 1kBK corresponding to energy differences on the order of 10(-6) hartree between different spin states. In the second approach, the observed features of the susceptibility minimum and maximum have been used to determine the best values of the exchange coupling constants from the theoretical formulas for an anisotropic AFA crystal. The AFM (Ja and Jc) and FM (Jb) exchange coupling constants and the Neel temperature (TN) found from this analysis correspond to Ja + Jc = -1.05 kBK, Jb = 1.35 kBK, and TN = 1.75 K. The calculated J values significantly differ from those estimated from a linear Heisenberg chain model, but generate a susceptibility versus temperature graph that mimics the experimental plot. PMID- 15268379 TI - Application of the modified Shepard interpolation method to the determination of the potential energy surface for a molecule-surface reaction: H2 + Pt(111). AB - We have used a modified Shepard (MS) interpolation method, initially developed for gas phase reactions, to build a potential energy surface (PES) for studying the dissociative chemisorption of H2 on Pt(111). The aim was to study the efficiency and the accuracy of this interpolation method for an activated multidimensional molecule-surface reactive problem. The strategy used is based on previous applications of the MS method to gas phase reactions, but modified to take into account special features of molecule-surface reactions, like the presence of many similar reaction pathways which vary only slightly with surface site. The efficiency of the interpolation method was tested by using an already existing PES to provide the input data required for the construction of the new PES. The construction of the new PES required half as many ab initio data points as the construction of the old PES, and the comparison of the two PESs shows that the method is able to reproduce with good accuracy the most important features of the H2 + Pt(111) interaction potential. Finally, accuracy tests were done by comparing the results of dynamics simulations using the two different PESs. The good agreement obtained for reaction probabilities and probabilities for rotationally and diffractionally inelastic scattering shows clearly that the MS interpolation method can be used efficiently to yield accurate PESs for activated molecule-surface reactions. PMID- 15268380 TI - Improved interatomic potentials for silicon-fluorine and silicon-chlorine. AB - Improved sets of empirical interatomic potentials for silicon-fluorine and silicon-chlorine are presented. The Tersoff-Brenner potential form has been reparameterized using the density-functional theory (DFT) cluster calculations of Walch. Halogenated silicon cluster energetics computed with DFT are, on average, within several tenths of an eV of the energies of the corresponding clusters with the reparameterized empirical potential for both Si-F and Si-Cl. Using the reparameterized Tersoff-Brenner potentials, molecular-dynamics simulations of F and Cl atom exposure to undoped silicon surfaces are in excellent agreement with published data on etch probability, halogen coverage at steady state, and etch product distributions. PMID- 15268381 TI - Voronoi diagrams generated by regressing edges of precipitation fronts. AB - Reaction-diffusion systems where one of the reagents (outer electrolyte) penetrates into a gel by diffusion and forms a precipitate with the other reagent (inner electrolyte) homogenized in the gel, are able to produce various complex precipitation patterns. The previously studied NaOH + AgNO3 and recently discovered CuCl2 + K3[Fe(CN)6] processes, (where the first reagent is the outer electrolyte and the other is the inner electrolyte homogenized in the gel), when reacted using the above mentioned method, are able to generate tessellations of a plane by a mechanism dependant on the dynamics of so-called regressing edges of the reaction fronts. The spontaneous partitioning of the reacted phases results in the construction of a pattern analogous to a Voronoi diagram or one of their generalizations. PMID- 15268382 TI - Experimental and simulation study of neon collision dynamics with a 1-decanethiol monolayer. AB - A study of the energy accommodation of neon colliding with a crystalline self assembled 1-decanethiol monolayer adsorbed on Au(111) is presented. The intensity and velocity dependencies of the scattered neon as a function of incident angle and energy were experimentally measured. Scattering calculations show good agreement with these results, which allows us to examine the detailed dynamics of the energy and momentum exchange at the surface. Simulation results show that interaction times are, at most, a few picoseconds. Even for these short times, energy exchange with the surface, both normal and in-plane, is very rapid. An important factor in determining the efficiency of energy exchange is the location at which the neon collides with the highly corrugated and structurally dynamic unit cell. Moreover, our combined experimental and theoretical results confirm that these are truly surface collisions in that neon penetration into the organic boundary layer does not occur, even for the highest incident energies explored, 560 meV. PMID- 15268383 TI - Enhancements in dissociative electron attachment to CF4, chlorofluorocarbons and hydrochlorofluorocarbons adsorbed on H2O ice. AB - We report that the absolute cross sections for dissociative attachment of approximately 0 eV electrons to chlorofluorocarbons (CFCs) and hydrochlorofluorocarbons (HCFCs) are strongly enhanced by the presence of H2O ice. The absolute cross sections for CFCl3, CHF2Cl, and CH3CF2Cl on water ice are measured to be approximately 8.9 x 10(-14), approximately 5.1 x 10(-15), and approximately 4.9 x 10(-15) cm2 at approximately 0 eV, respectively. The former value is about 1 order of magnitude higher than that in the gas phase, while the latter two are 3-4 orders higher. In contrast, the resonances at electron energies > or = 2.0 eV are strongly suppressed either for CFCs and HCFCs or for CF4 adsorbed on H2O ice. The cross-section enhancement is interpreted to be due to electron transfer from precursor states of the solvated electron in ice to an unfilled molecular orbital of CFCs or HCFCs followed by its dissociation. This study indicates that electron-induced dissociation is a significant process leading to CFC and HCFC fragmentation on ice surfaces. PMID- 15268384 TI - Spectroscopy of the conformational disorder in molecular films: tetracosane and squalane on Pt(111). AB - The spectroscopic investigation of the molecular vibrations of adsorbed branched and unbranched alkane molecules using helium atom scattering (HAS) provides evidence for the thermal formation of gauche defects in tetracosane (C24H50) monolayers above 200 K. HAS results for the vibration of tetracosane molecules perpendicular to the Pt(111) surface reveal a strong frequency decrease and peak broadening above the transition temperature which can be related to a reduction of the force holding the molecules to the surface. This reduction of the force is interpreted as being due to the thermal formation of gauche defects within the tetracosane molecules. PMID- 15268385 TI - State-resolved dynamics of oxygen atom recombination on polycrystalline Ag. AB - Rotationally resolved, velocity distributions for desorbed O2 molecules formed by O-atom recombination on the surface of a polycrystalline Ag surface are reported. Surface O atoms are generated by oxygen permeation through a 0.25-mm-thick Ag foil heated to 1020 K. Desorbing O2 molecules are probed by (2 + 1) resonant multiphoton ionization via the C 3Pig (3ssigma), v' = 2 <-- <-- X 3Sigmag-, v" = 0 transition and time-of-flight mass spectrometry. Measured velocity distributions are near Maxwell-Boltzmann and yield average translational energies which are significantly lower than the surface temperature ([Et]/2kB approximately 515 K) and essentially independent of rotational excitation. Comparison of the observed C-X (2,0) resonantly enhanced multiphoton ionization spectrum with spectral simulations suggests that the v" = 0 rotational state distribution is more consistent with the surface temperature, but spectral congestion and apparent intensity perturbations prevent a more quantitative analysis. The calculated, sticking curves show a small barrier energy barrier (approximately 10 meV) beyond which sticking decreases. These observations are consistent with low energy desorption and adsorption pathways involving a weakly bound molecular O2 precursor. PMID- 15268386 TI - Laser control of product electronic state: desorption from alkali halides. AB - We demonstrate laser control of the electronic product state distribution of photodesorbed halogen atoms from alkali halide crystals. Our general model of surface exciton desorption dynamics is developed into a simple method for laser control of the relative halogen atom spin-orbit laser desorption yield. By tuning the excitation laser photon energy in a narrow region of the absorption threshold, the yield of excited state chorine atoms, Cl(2P(1/2)), can be made to vary from near 0 to 80% for KCl and from near 0 to 50% for NaCl relative to the total yield of Cl atoms. We describe the physical properties necessary to obtain a high degree of product state control and the limitation induced when these requirements are not met. These results demonstrate that laser control can be applied to solid state surface reactions and provide strong support for surface exciton-based desorption models. PMID- 15268387 TI - Molecular dynamics study of the n-hexane-water interface: towards a better understanding of the liquid-liquid interfacial broadening. AB - By molecular dynamics simulations, we have studied the hydrophilic-hydrophobic interface between water and n-hexane liquid phases. For all temperatures studied our computed interfacial tension agrees very well with the experimental value. However, the interfacial width calculated from capillary wave theory systematically overestimates the width obtained from fitting either the total density or composition profile. We rationalize the applicability of capillary wave theory for our system by reconsidering the usual value taken for the correlation length. This is motivated by the presence of order at the interface. Possible implications for recent experimental studies on the structure of model alkane-water interfaces are discussed, including the significance of the intrinsic width parameter. PMID- 15268388 TI - Phase diagram of mixtures of hard colloidal spheres and discs: a free-volume scaled-particle approach. AB - Phase diagrams of mixtures of colloidal hard spheres with hard discs are calculated by means of the free-volume theory. The free-volume fraction available to the discs is determined from scaled-particle theory. The calculations show that depletion induced phase separation should occur at low disc concentrations in systems now experimentally available. The gas-liquid equilibrium of the spheres becomes stable at comparable size ratios as with bimodal mixtures of spheres or mixtures of rods and spheres. Introducing finite thickness of the platelets gives rise to a significant lowering of the fluid branch of the binodal. PMID- 15268389 TI - Excluded volume entropic effects on protein unfolding times and intermediary stability. AB - The dynamics of protein folding result from both enthalpic and entropic contributions to the free energy. In this paper we focus on entropic volume exclusion effects. We carry out computer simulations using a model that allows us to independently change the size or biochemical properties of amino acid residues. To determine the importance of excluded volume effects, we investigate the effects of changing the size of side chains on the unfolding dynamics of a model four-helix bundle protein. In addition, we also investigate the effects of changing the thickness of the chain's backbone. This has relevance to the behavior of synthetic polymers where the size of the constituent units can be varied. We find that entropic excluded volume effects are crucially important for stabilizing the organized native state relative to the molten globule. PMID- 15268390 TI - Dynamics of chain closure: approximate treatment of nonlocal interactions. AB - The Wilemski-Fixman model of diffusion controlled-reactions [J. Chem. Phys. 58, 4009 (1973)] is combined with a generalized random walk description of chain conformations to predict the dependence of the closure time tau on the chain length N of polymers with reactive end groups and nonlocal interactions. The nonlocal interactions are modeled by a modification to the connectivity term in the Edwards continuum representation of the polymer. The modification involves a parameter h lying between 0 and 1 that is a measure of the extent of correlation between adjacent monomers on the chain backbone. Different choices of h correspond to chain conformations of different average radial dimensions. In particular, the values 1/3, 1/2 and 3/5 provide approximations to the statistics of polymers in poor, theta and good solvents, respectively. The closure time tau of such chains is calculated analytically for different N. In all cases, tau is found to vary as a power law in N, Nb, with b a function of h. For the special case h = 1/3, which models collapsed polymers and globular proteins, b is about 1.6-1.7. PMID- 15268391 TI - Excitonic coupling in polythiophenes: comparison of different calculation methods. AB - In conjugated polymers the optical excitation energy transfer is usually described as Forster-type hopping between so-called spectroscopic units. In the simplest approach using the point-dipole approximation the transfer rate is calculated based on the interaction between the transition dipoles of two spectroscopic units. In the present work we compare this approach with three others: The line-dipole approximation, the Coulomb integral between the transition densities, and a quantum-chemical calculation of the interacting dimer as entity. The latter two approaches are based on the semiempirical method ZINDO. The line-dipole approximation is an attractive compromise between computational effort and precision for calculations of the excitonic coupling in extended conjugated polymers. PMID- 15268392 TI - Lattice Monte Carlo simulations of three-dimensional charged polymer chains. AB - The configurational properties of strongly charged polyelectrolytes accompanied by neutralizing counterions in dilute solutions are simulated using the cooperative motion algorithm on the face-centered-cubic lattice. The full Coulomb potential and the excluded volume condition between different ions/beads are taken into account and the reduced temperature T* is considered the main, variable parameter. The calculations that have been carried out for solutions of both single and several chains indicate a few regions of their behavior: (1) for T*--> infinity, it corresponds to that of neutral, self-avoiding polymers under good solvent conditions; (2) for T* approximately 1, due to the electrostatic interactions being effectively stronger, the chains are more outstretched compared to their size at other temperatures; (3) for T* well below one, the counterion condensation becomes more and more dominant, which gradually leads to strongly collapsed chains; and (4) at the lowest temperatures the chains and counterions assume low-energy configurations in the form of neutral, compact aggregates. PMID- 15268393 TI - Lattice Monte Carlo simulations of three-dimensional charged polymer chains. II. Added salt. AB - The configurational properties of strongly charged polyelectrolytes accompanied by counterions and salt ions in dilute solutions are simulated using the cooperative motion algorithm on the face-centered-cubic lattice. The calculations indicate that both positive and negative ions condense on the chains at sufficiently low temperatures and their amount depends on the concentration of added salt. As the temperature decreases from high to low, the chains undergo conformational changes from neutral-like, self-avoiding polymers by more outstretched forms to compact globules. The observed extension of the chains at intermediate temperatures is also affected by the amount of salt. Furthermore, configurations with the lowest energies recorded at the lowest temperatures are aggregates of single or more entangled chains and ions of both types. PMID- 15268394 TI - Monte Carlo investigations of dense copolymer systems. III. Properties of triblock copolymers in good and theta solvent. AB - The present article gives an analysis of XYX triblock copolymers in a good solvent and in a theta solvent, the segments of type X and type Y being repulsive for each other. The results are compared to homopolymers as well as to copolymers in a selective solvent that is a good one for the outer blocks and a theta solvent for the inner one and vice versa, the strength of repulsion between blocks being the same as in the present types of copolymers. A lattice model is used for the investigations and the concentration ranges from a volume fraction phi = 0 up to phi = 0.8. In the limit phi --> 0 the triblocks in good solvent are slightly more expanded than homopolymers and in theta solvent mean square dimensions of triblocks are considerably increased compared to homopolymers due to the repulsion between blocks. With increasing concentration the dimensions decrease but then they increase again and for large concentrations they become similar for all types of copolymers studied, as the effect of the solvent levels off making the repulsive interaction between blocks the dominant interaction. This leads to an orientation effect and as a consequence to microphase separation which is demonstrated by the concentration dependence of various quantities as well as by visualization of snapshots. PMID- 15268395 TI - Shear-induced migration in flowing polymer solutions: simulation of long-chain DNA in microchannels [corrected]. AB - We simulate dilute solution dynamics of long flexible polymer molecules in pressure driven flow in channels with widths of roughly 0.1-10 times the polymer bulk radius of gyration. This is done using a self-consistent coarse-grained Langevin description of the polymer dynamics and a numerical simulation of the flow in the confined geometry that is generated by the motions of polymer segments. Results are presented for a model of DNA molecules of approximately 10 100 microm contour length in micron-scale channels. During flow, the chains migrate toward the channel centerline, in agreement with well-known experimental observations. The thickness of the resulting hydrodynamic depletion layer increases with molecular weight at constant flow strength; higher molecular weight chains therefore move with a higher average axial velocity than lower molecular weight chains. In contrast, if the hydrodynamic effects of the confining geometry are neglected, depletion of concentration is observed in the center of the channel rather than at the walls, contradicting experimental observations. The mechanisms for migration are illustrated using a simple kinetic theory dumbbell model of a confined flexible polymer. The simple theory correctly predicts the trends observed in the detailed simulations. We also examine the steady-state stretch of DNA chains as a function of channel width and flow strength. The flow strength needed to stretch a highly confined chain away from its equilibrium length is shown to increase with decreasing channel width, independent of molecular weight; this is fairly well explained using a simple blob picture. PMID- 15268396 TI - Photobleaching time distribution of a single tetramethylrhodamine molecule in agarose gel. PMID- 15268397 TI - Two-dimensional optical spectroscopy: two-color photon echoes of electronically coupled phthalocyanine dimers. AB - Two-color photon echo peak shift spectroscopy was used to study electronic coupling in a phthalocyanine homodimer. Two optical parametric amplifiers were used to produce pulses to excite the split lower states of LuPc2-. The existence of a two-color peak shift indicates the existence of correlation between these two dipole-allowed states. The nature of this correlation is discussed based on theoretical predictions of the interactions between exciton and charge resonance states. PMID- 15268398 TI - Atom-bond electronegativity equalization method fused into molecular mechanics. I. A seven-site fluctuating charge and flexible body water potential function for water clusters. AB - Recently, experimental and theoretical studies on the water system are very active and noticeable. A transferable intermolecular potential seven points approach including fluctuation charges and flexible body (ABEEM-7P) based on a combination of the atom-bond electronegativity equalization and molecular mechanics (ABEEM/MM), and its application to small water clusters are explored and tested in this paper. The consistent combination of ABEEM and molecular mechanics (MM) is to take the ABEEM charges of atoms, bonds, and lone-pair electrons into the intermolecular electrostatic interaction term in molecular mechanics. To examine the charge transfer we have used two models coming from the charge constraint types: one is a charge neutrality constraint on whole water system and the other is on each water molecule. Compared with previous water force fields, the ABEEM-7P model has two characters: (1) the ABEEM-7P model not only presents the electrostatic interaction of atoms, bonds and lone-pair electrons and their changing in respond to different ambient environment but also introduces "the hydrogen bond interaction region" in which a new parameter k(lp,H)(R(lp,H)) is used to describe the electrostatic interaction of the lone pair electron and the hydrogen atom which can form the hydrogen bond; (2) nonrigid but flexible water body permitting the vibration of the bond length and angle is allowed due to the combination of ABEEM and molecular mechanics, and for van der Waals interaction the ABEEM-7P model takes an all atom-atom interaction, i.e., oxygen-oxygen, hydrogen-hydrogen, oxygen-hydrogen interaction into account. The ABEEM-7P model based on ABEEM/MM gives quite accurate predictions for gas phase state properties of the small water clusters (H(2)O)(n) (n=2-6), such as optimized geometries, monomer dipole moments, vibrational frequencies, and cluster interaction energies. Due to its explicit description of charges and the hydrogen bond, the ABEEM-7P model will be applied to discuss properties of liquid water, ice, aqueous solutions, and biological systems. PMID- 15268399 TI - The role of fluctuations in both density functional and field theory of nanosystems. AB - The role of fluctuations in both the density functional theory (DFT) and the field theory (FT) of nanosystems is studied. It turns out that although fluctuations are rigorously incorporated into the general formalism of DFT, they are often omitted in the choice of an approximate free energy functional that must be constructed in order to solve the basic integral equation appearing in DFT. Aside from the analytical discussion, it is demonstrated, in connection with a particular system (fluid in a nanopore) that the effects of fluctuation are missing when one of the most common functionals for this system is used. The demonstration involves a comparison of the results of Monte Carlo simulation with the predictions of DFT when this free energy functional is used. The applicability of FT and DFT in the context of the theory of nucleation is also discussed. PMID- 15268400 TI - On the peculiarities of the diabatic framework: new insight. AB - In this article we consider the electronic diabatic presentation of a two-state system with the aim of earning insight regarding the distribution of conical intersections in a given region. In this process we revealed explicit relationship between the diabatic potentials and the locations of conical intersections. The study is accompanied with numerical examples as worked out for a model and ab initio potential energy surfaces of the Na+H2 system. PMID- 15268401 TI - Is there any group additive rules in the calculation of electron correlation energies of long straight chain alkane molecules? AB - According to the definition in the text, the correlation energy of 1s2C of carbon atoms, the primary and secondary C-H bonding electron pairs in some CH3, CH2 fragments and CH3(CH2)mCH3 (m=1-5) linear alkane molecules are calculated and analyzed. The transferability of the correlation energies of these electron pairs in the linear alkanes is investigated. The results indicate that the correlation energy of 1s2C is perfectly transferable in the respective methyl and methylene groups, while the correlation energies of the primary and secondary C-H bonding electron pairs are approximately transferable in methyl and methylene groups. The analysis of the results of group correlation energy shows that both of the correlation energies of methyl and methylene groups are transferable in these linear alkanes. The correlation energies of methylene group in CH3(CH2)mCH3 (m=1 5) molecules are slightly decreasing showing a converging trend to a "standard" methylene group in linear alkanes. The excellent fitting relationship between the total correlation energy and the number of methylene groups of the linear alkanes shows that the total correlation energy is a linear function of the number of methylene groups, which means that the total correlation energies of large linear alkanes can be reproduced and predicted by counting the numbers of methylene groups. In this way, total correlation energy of large linear alkane molecule can be approximately calculated using this simple group additive scheme with substantial saving in computational time. PMID- 15268402 TI - Coupled-cluster singles and doubles for extended systems. AB - Coupled-cluster theory with connected single and double excitation operators (CCSD) and related approximations, such as linearized CCSD, quadratic configuration interaction with single and double excitation operators, coupled cluster with connected double excitation operator (CCD), linearized CCD, approximate CCD, and second- and third-order many-body perturbation theories, are formulated and implemented for infinitely extended one-dimensional systems (polymers), on the basis of the periodic boundary conditions and distance-based screening of integrals, density matrix elements, and excitation amplitudes. The variation of correlation energies with the truncation radii of short- and long range lattice sums and with the number of wave vector sampling points in the first Brillouin zone is examined for polyethylene, polyacetylene, and polyyne, and is shown to be a function of the degree of pi-electron conjugation or the fundamental band gaps. The t2 and t1 amplitudes in the atomic orbital (AO) basis are obtained by first computing the t amplitudes in the Bloch-orbital basis and subsequently back-transforming them into the AO basis. The plot of these AO-based t amplitudes as a function of unit cells also indicates that the t2 amplitudes of polyacetylene and polyyne exhibit appreciably slower decay than those of polyethylene, although the asymptotic decay behavior is invariably 1/r3. The AO based t1 amplitudes appear to correlate strongly with the electronic structure, and they decay seemingly exponentially for polyethylene whereas they stay at a constant magnitude across the seventh nearest neighbors of polyacetylene and polyyne, which attests to far reaching effects of nondynamical electron correlation mediated by orbital rotation. Nonetheless, the unit cell contributions to the correlation energies taper below 10(-6) hartree after 15 A for all three polymers. The basis set dependence of the decay behavior of t2 amplitudes is also examined for linear hydrogen fluoride polymer (HF)infinity and linear beryllium polymer (Be)infinity employing the STO-3G, 6-31G, and 6-31G* basis sets, and proves to be rather small. PMID- 15268403 TI - Analytical energy gradient of the symmetry-adapted-cluster configuration interaction general-R method for singlet to septet ground and excited states. AB - A method of calculating analytical energy gradients of the singlet and triplet excited states, ionized states, electron-attached states, and high-spin states from quartet to septet states by the symmetry-adapted-cluster configuration interaction general-R method is developed and implemented. This method is a powerful tool in the studies of geometries, dynamics, and properties of the states of molecules in which not only one-electron processes but also two- and multielectron processes are involved. The performance of the present method was confirmed by calculating the geometries and the spectroscopic constants of the diatomic and polyatomic molecules in various electronic states involving the ground state and the one- to three-electron excited states. The accurate descriptions were obtained for the equilibrium geometries, vibrational frequencies, and adiabatic excitation energies, which show the potential usefulness of the present method. The particularly interesting applications were to the C' 1Ag state of acetylene, the A 2Deltau and B 2Sigmau+ states of CNC and the 4B1 and a 4Piu states of N3 radical. PMID- 15268404 TI - Semiclassical initial value treatment of correlation functions. AB - Two semiclassical, initial value representation (IVR) treatments are presented for the correlation function psi(f) e-iHt/h psi(i), where psi(i) and psi(f), are energy eigenfunctions of a "zero-order" Hamiltonian describing an arbitrary, integrable, vibrational system. These wave functions are treated semiclassically so that quantum calculations and numerical integrations over these states are unnecessary. While one of the new approximations describes the correlation function as an integral over all phase space variables of the system, in a manner similar to most existing IVR treatments, the second approximation describes the correlation function as an integral over only half of the phase space variables (i.e., the angle variables for the initial system). The relationship of these treatments to the conventional Herman-Kluk approximation for correlation functions is discussed. The accuracy and convergence of these treatments are tested by calculations of absorption spectra for model systems having up to 18 degrees of freedom, using Monte Carlo techniques to perform the multidimensional phase space integrations. Both treatments are found to be capable of producing spectra of excited, anharmonic states that agree well with quantum results. Although generally less accurate than full phase space or Herman-Kluk treatments, the half phase space method is found to require far fewer trajectories to achieve convergence. In addition, this number is observed to increase much more slowly with the system size than it does for the former methods, making the half-phase space technique a very promising method for the treatment of large systems. PMID- 15268405 TI - Free energy simulations: use of reverse cumulative averaging to determine the equilibrated region and the time required for convergence. AB - A method is proposed for improving the accuracy and efficiency of free energy simulations. The essential idea is that the convergence of the relevant measure (e.g., the free energy derivative in thermodynamic integration) is monitored in the reverse direction starting from the last frame of the trajectory, instead of the usual approach, which begins with the first frame and goes in the forward direction. This simple change in the use of the simulation data makes it straightforward to eliminate the contamination of the averages by contributions from the equilibrating region. A statistical criterion is introduced for distinguishing the equilibrated (production) region from the equilibrating region. The proposed method, called reverse cumulative averaging, is illustrated by its application to the well-studied case of the alchemical free energy simulation of ethane to methanol. PMID- 15268406 TI - Molecule intrinsic minimal basis sets. I. Exact resolution of ab initio optimized molecular orbitals in terms of deformed atomic minimal-basis orbitals. AB - A method is presented for expressing the occupied self-consistent-field (SCF) orbitals of a molecule exactly in terms of chemically deformed atomic minimal basis-set orbitals that deviate as little as possible from free-atom SCF minimal basis orbitals. The molecular orbitals referred to are the exact SCF orbitals, the free-atom orbitals referred to are the exact atomic SCF orbitals, and the formulation of the deformed "quasiatomic minimal-basis-sets" is independent of the calculational atomic orbital basis used. The resulting resolution of molecular orbitals in terms of quasiatomic minimal basis set orbitals is therefore intrinsic to the exact molecular wave functions. The deformations are analyzed in terms of interatomic contributions. The Mulliken population analysis is formulated in terms of the quasiatomic minimal-basis orbitals. In the virtual SCF orbital space the method leads to a quantitative ab initio formulation of the qualitative model of virtual valence orbitals, which are useful for calculating electron correlation and the interpretation of reactions. The method is applicable to Kohn-Sham density functional theory orbitals and is easily generalized to valence MCSCF orbitals. PMID- 15268407 TI - Molecule intrinsic minimal basis sets. II. Bonding analyses for Si4H6 and Si2 to Si10. AB - The method, introduced in the preceding paper, for recasting molecular self consistent field (SCF) or density functional theory (DFT) orbitals in terms of intrinsic minimal bases of quasiatomic orbitals, which differ only little from the optimal free-atom minimal-basis orbitals, is used to elucidate the bonding in several silicon clusters. The applications show that the quasiatomic orbitals deviate from the minimal-basis SCF orbitals of the free atoms by only very small deformations and that the latter arise mainly from bonded neighbor atoms. The Mulliken population analysis in terms of the quasiatomic minimal-basis orbitals leads to a quantum mechanical interpretation of small-ring strain in terms of antibonding encroachments of localized molecular-orbitals and identifies the origin of the bond-stretch isomerization in Si4H6. In the virtual SCF/DFT orbital space, the method places the qualitative notion of virtual valence orbitals on a firm basis and provides an unambiguous ab initio identification of the frontier orbitals. PMID- 15268408 TI - Estimating entropies from molecular dynamics simulations. AB - While the determination of free-energy differences by MD simulation has become a standard procedure for which many techniques have been developed, total entropies and entropy differences are still hardly ever computed. An overview of techniques to determine entropy differences is given, and the accuracy and convergence behavior of five methods based on thermodynamic integration and perturbation techniques was evaluated using liquid water as a test system. Reasonably accurate entropy differences are obtained through thermodynamic integration in which many copies of a solute are desolvated. When only one solute molecule is involved, only two methods seem to yield useful results, the calculation of solute-solvent entropy through thermodynamic integration, and the calculation of solvation entropy through the temperature derivative of the corresponding free-energy difference. One-step perturbation methods seem unsuitable to obtain entropy estimates. PMID- 15268409 TI - Laser-induced fluorescence spectroscopy of NC3S. AB - In a discharged supersonic jet of acetonitrile and carbon disulfide, we have for the first time observed an electronic transition of the NC(3)S radical using laser-induced fluorescence (LIF) spectroscopy. A progression originating from the C-S stretching mode of the upper electronic state appears in the excitation spectrum. Each band of the progression has a polyad structure due to anharmonic resonances with even overtones of bending modes. Rotationally resolved spectra have been observed by high-resolution laser scans, and the electronic transition is assigned to A 2Pii-X 2Pii. For the vibronic origin band, the position and the effective rotational constant of the upper level have been determined to be 21 553.874(1) and 0.046 689(4) cm(-1), respectively. The dispersed fluorescence spectrum from the zero vibrational level of A 2Pi3/2 has also been observed; its vibrational structure is similar to that of the LIF excitation spectrum, showing a prominent C-S stretching progression with polyad structures. The vibrational frequencies of the C-S stretching mode in the ground and excited electronic states are determined to be 550 and 520 cm(-1), respectively. Fluorescence decay profiles have been measured for several vibronic levels of the A state. PMID- 15268410 TI - Mode dependent vibrational autoionization of Rydberg states of NO2. II. Comparing the symmetric stretching and bending vibrations. AB - Triple-resonance excitation and high-resolution photoelectron spectroscopy are combined to characterize the mode selectivity of vibrational autoionization of the high Rydberg states of NO2. Photoelectron spectra and vibrational branching fractions are reported for autoionizing Rydberg states converging to the NO2+ X 1Sigmag +(110) state, that is, with one quantum in the symmetric stretch, nu1, and one quantum in the bending vibration, nu2. These results indicate that autoionization proceeds most efficiently through the loss of one quantum from the symmetric stretch rather than from the bending vibration. The implications of this result are discussed in terms of the autoionization mechanism. PMID- 15268411 TI - Combined experimental/theoretical investigation of the He+ICl interactions. I. Rovibronic spectrum of He...ICl complexes in the ICl B-X, 3-0 region. AB - Transitions of two different stereoisomers of the He...ICl(X,v" = 0) weakly bound complex, one with a T-shaped orientation and another that is most likely linear, have been observed in laser-induced fluorescence experiments performed in the ICl B-X region. Here we present experimental and theoretical results aimed at confirming the previous assignments and at gaining additional insights into the He+ICl interactions. High resolution action spectra were recorded in the same region to identify those features that could be attributed to transitions of the He...I35Cl(X,v" = 0) isomers and not to higher-order complexes, Hen...I35Cl, where n > or = 2, or I37Cl containing species. Calculations of the rovibronic spectra of the He...I35Cl complexes in the ICl B-X, 2-0 and 3-0 regions were performed using an ab initio potential energy surface for the He+ICl(X,v" = 0) ground state and two different pairwise additive potentials for the He+ICl(B,v' = 2,3) excited states. The rotation-vibration energies and wave functions for the He cdots, three dots, centered I35Cl complexes were obtained for all bound states with total angular momentum J < 10 using both of these potentials. Electronic spectra were generated using these results, assuming that the transition moment lies along the ICl bond and is not perturbed by the presence of the helium atom. The calculations qualitatively reproduce the He cdots, three dots, centered I35Cl action spectrum and strongly support the previous assignments. The calculations also indicate that some of the spectral congestion observed near the linear band may be attributed to transitions of the linear isomer to multiple intermolecular levels in the excited state. Coriolis coupling strongly mixes He cdots, three dots, centered ICl(B,v') states with rotational excitation, making simulations and assignments of the linear band observed in the experimental spectrum difficult. PMID- 15268412 TI - High resolution measurements of kinetic energy release distributions of neon, argon, and krypton cluster ions using a three sector field mass spectrometer. AB - Using a newly constructed three sector field mass spectrometer (resulting in a BE1E2 field configuration) we have measured the kinetic energy release distributions of neon, argon, and krypton cluster ions. In the present study we used the first two sectors, B and E1, constituting a high resolution mass spectrometer, to select the parent ions in terms of mass, charge, and energy, and studied the decay of those ions in the third field free region. Due to the improved mass resolution we were able to extend earlier studies carried out with a two sector field machine, where an upper size limit arose from the fact that several isotopomers contribute to a decaying parent ion beam when the cluster size exceeds a certain value. Furthermore we developed a new data analysis. It allows us to model also fragment ion peaks that are a superposition of different decay reactions and thus we can determine the average kinetic energy release for all decay reactions of a given cluster ion. In a further step we used these results to determine the binding energies of cluster ions Rg(n) (n> or =10) by applying finite heat bath theory. The smaller sizes have not been included in this analysis, because the validity of finite heat bath theory becomes questionable below n approximately 10. The present average kinetic energy releases and binding energies are compared with other experiments and various calculations. PMID- 15268413 TI - Dispersion corrections to density functionals for water aromatic interactions. AB - We investigate recently published methods for extending density functional theory to the description of long-range dispersive interactions. In all schemes an empirical correction consisting of a C6r(-6) term is introduced that is damped at short range. The coefficient C6 is calculated either from average molecular or atomic polarizabilities. We calculate geometry-dependent interaction energy profiles for the water benzene cluster and compare the results with second-order Moller-Plesset calculations. Our results indicate that the use of the B3LYP functional in combination with an appropriate mixing rule and damping function is recommended for the interaction of water with aromatics. PMID- 15268414 TI - Quantum-mechanical calculations on pressure and temperature dependence of three body recombination reactions: application to ozone formation rates. AB - A quantum-mechanical model is designed for the calculation of termolecular association reaction rate coefficients in the low-pressure fall-off regime. The dynamics is set up within the energy transfer mechanism and the kinetic scheme is the steady-state approximation. We applied this model to the formation of ozone O + O2 + M --> O3 + M for M = Ar, making use of semiquantitative potential energy surfaces. The stabilization process is treated by means of the vibrational close coupling infinite order sudden scattering theory. Major approximations include the neglect of the O3 vibrational bending mode and rovibrational couplings. We calculated individual isotope-specific rate constants and rate constant ratios over the temperature range 10-1000 K and the pressure fall-off region 10(-7) 10(2) bar. The present results show a qualitative and semiquantitative agreement with available experiments, particularly in the temperature region of atmospheric interest. PMID- 15268415 TI - Raman under nitrogen. The high-resolution Raman spectroscopy of crystalline uranocene, thorocene, and ferrocene. AB - The utility of recording Raman spectroscopy under liquid nitrogen, a technique we call Raman under nitrogen (RUN), is demonstrated for ferrocene, uranocene, and thorocene. Using RUN, low-temperature (liquid nitrogen cooled) Raman spectra for these compounds exhibit higher resolution than previous studies, and new vibrational features are reported. The first Raman spectra of crystalline uranocene at 77 K are reported using excitation from argon (5145 A) and krypton (6764 A) ion lasers. The spectra obtained showed bands corresponding to vibrational transitions at 212, 236, 259, 379, 753, 897, 1500, and 3042 cm(-1), assigned to ring-metal-ring stretching, ring-metal tilting, out-of-plane CCC bending, in-plane CCC bending, ring-breathing, C-H bending, CC stretching and CH stretching, respectively. The assigned vibrational bands are compared to those of uranocene in THF, (COT)2-, and thorocene. All vibrational frequencies of the ligands, except the 259 cm(-1) out-of-plane CCC bending mode, were found to increase upon coordination. A broad, polarizable band centered about approximately 460 cm(-1) was also observed. The 460 cm(-1) band is greatly enhanced relative to the vibrational Raman transitions with excitations from the krypton ion laser, which is indicative of an electronic resonance Raman process as has been shown previously. The electronic resonance Raman band is observed to split into three distinct bands at 450, 461, and 474 cm(-1) with 6764 A excitation. Relativistic density functional theory is used to provide theoretical interpretations of the measured spectra. PMID- 15268416 TI - Spin-locking of half-integer quadrupolar nuclei in nuclear magnetic resonance of solids: creation and evolution of coherences. AB - Spin-locking of half-integer quadrupolar nuclei, such as 23Na (I=3/2) and 27Al (I=5/2), is of renewed interest owing to the development of variants of the multiple-quantum and satellite-transition magic angle spinning (MAS) nuclear magnetic resonance experiments that either utilize spin-locking directly or offer the possibility that spin-locked states may arise. However, the large magnitude and, under MAS, the time dependence of the quadrupolar interaction often result in complex spin-locking phenomena that are not widely understood. Here we show that, following the application of a spin-locking pulse, a variety of coherence transfer processes occur on a time scale of approximately 1/omegaQ before the spin system settles down into a spin-locked state which may itself be time dependent if MAS is performed. We show theoretically for both spin I=3/2 and 5/2 nuclei that the spin-locked state created by this initial rapid dephasing typically consists of a variety of single- and multiple-quantum coherences and nonequilibrium population states and we discuss the subsequent evolution of these under MAS. In contrast to previous work, we consider spin-locking using a wide range of radio frequency field strengths, i.e., a range that covers both the "strong-field" (omega1 >> omegaQPAS and "weak-field" (omega1 << omegaQPAS limits. Single- and multiple-quantum filtered spin-locking experiments on NaNO2, NaNO3, and Al(acac)3, under both static and MAS conditions, are used to illustrate and confirm the results of the theoretical discussion. PMID- 15268417 TI - Can ortho-para transitions for water be observed? AB - The spectrum of water can be considered as the juxtaposition of the spectra of two molecules, with different total nuclear spin: ortho-H2O, and para-H2O. No transitions have ever been observed between the two different nuclear-spin isotopomers. The interconversion time is unknown and it is widely assumed that interconversion is forbidden without some other intervention. However, weak nuclear spin-rotation interaction occurs and can drive ortho to para transitions. Ab initio calculations show that the hyperfine nuclear spin-rotational coupling constants are about 30 kHz. These constants are used to explore the whole vibration-rotation spectrum with special emphasis on the coupling between nearby levels. Predictions are made for different spectral regions where the strongest transitions between ortho and para levels of water could be experimentally observed. PMID- 15268418 TI - Electric-field-induced g/u mixing of the E0g+(3P2) and D0u+(3P2) ion-pair states of jet-cooled I2 observed using optical triple resonance. AB - Electric-field-induced electronic state g/u mixing of nearly isoenergetic rovibrational levels of the E0g+(3P2) and D0u+(3P2) ion-pair states of I2 has been observed using optical triple resonance combined with resonance ionization. Detectable mixing with applied fields of 1 kV/cm occurs over a range of energy level separations of < or = 0.3 cm(-1). PMID- 15268419 TI - A theoretical study of small copper oxide clusters: Cu2Ox (x = 1 - 4). AB - Density functional theory (DFT) calculations are performed to study Cu2Ox (x = 1 4) clusters in their neutral, anionic and cationic states. The ground state structures are obtained and found to exhibit linear or near linear structures, which are different from the two- or three-dimensional ones suggested by the previous theoretical calculations. The calculated electron affinities of the clusters are in good agreement with the experimental ones. The low-lying excited states for the clusters are calculated using time-dependent DFT and used to assign the features in the photoelectron spectra. Our results compare well with the available experimental data. PMID- 15268420 TI - The structure of the phenol-nitrogen cluster: a joint experimental and ab initio study. AB - The rotationally resolved LIF spectra of four different isotopomers of the phenol -nitrogen cluster have been measured to elucidate the structural parameters of the cluster in ground and electronically excited (S1) state. The fit of the rotational constants has been performed by a genetic algorithm and by an assigned fit to the line frequencies. The results of both methods are compared. The intermolecular structures are fit to the inertial parameters and are compared to the results of ab initio calculations for both states. This fit was performed under the restriction that the geometry of the monomer moieties do not change upon complexation. Of the remaining five intermolecular parameters two dihedral angles were fixed due to the planarity of the complex, which was inferred from the inertial defects of all isotopomers. The distance of the nearest nitrogen atom to the hydrogen atom of the phenolic hydroxy group is found to decrease upon electronic excitation of the chromophore considerably more than predicted from ab initio calculations. This deviation between theory and experiment can be traced back to the absence of electron-electron correlation in the performed complete active space self-consistent field calculations. The shortening of the OH...NN "hydrogen" bond upon electronic excitation is in agreement with the increased dipole moment of phenol in the S1-state. PMID- 15268421 TI - Comparative studies of the photoinduced reactions in the Mg+-SCNC2H5 and Mg+ NCSC2H5 complexes. AB - The photoinduced reactions of the complexes Mg+-SCNC2H5 and Mg+-NCSC2H5 are studied comparatively in the spectral range of 230-440 nm. One-photon excitation of the complexes through the Mg+ chromophore (3 2P <-- 3 2S) gives rise to the evaporative fragment as well as the molecular activation and charge transfer products. The action spectra of the complexes consist of three broad peaks for Mg+-SCNC2H5 and two for Mg+-NCSC2H5, which accord with the structures obtained from quantum mechanics calculations. These calculations reveal two association isomers for Mg+-SCNC2H5: one is with Mg+ being linked to the S atom and the other to the N atom. The former is more stable than the latter by only 0.23 eV. Both of the isomers have been shown to exist in the complex source employed in our experiments. On the other hand, only one stable structure is found for the complex Mg+-NCSC2H5 characterized by the Mg+-N linkage. In general, the photofragments are dominated by Mg+ at lambda > 400 nm, which decreases with decreasing wavelength accompanied by the increase in other photoproducts. In addition, the branching ratios of Mg+ to other photoproducts are nearly constant in the short wavelength region but decrease with decreasing wavelength. The observed photoreactions have been reasonably explained. PMID- 15268422 TI - Electron affinity of the sodium atom within the coupled-channel hyperspherical approach. AB - We present a nonadiabatic calculation, within the hyperspherical adiabatic approach, for the ground state energy of the alkali-metal negative ions. An application to the sodium negative ion (Na-) is considered. This system is treated as a two-electron problem in which a model potential is used for the interaction between the Na+ core and the valence electrons. Potential curves and nonadiabatic couplings are obtained by a direct numerical calculation, as well as the channel functions. An analysis of convergence is made and comparisons of the electron affinity with results of prior work of other authors are given. PMID- 15268423 TI - Reaction pathway and potential barrier for the CaH product in the reaction of Ca(4s4p1P1) + H2 --> CaH(X2Sigma+) + H. AB - The nascent CaH product in the reaction Ca(4s4p1P1) + H2 --> CaH(X2Sigma+) + H is obtained using a pump-probe technique. The CaH(v = 0,1) distributions, with a population ratio of CaH(v = 0)/CaH(v = 1) = 2.7+/-0.2, may be characterized by low Boltzmann rotational temperature. According to Arrhenius theory, the temperature dependence measurement yields a potential barrier of 3820+/-480 cm( 1) for the current reaction. As a result of the potential energy surfaces (PES) calculations, the reaction pathway favors a Ca insertion into the H2 bond along a (near) C2v geometric approach. As the H2 bond is elongated, the configurational mixing between the orbital components of the 4p and nearby low-lying 3d state with the same symmetry makes significant the nonadiabatic transition between the 5A' and 2A' surface in the repulsive limbs. Therefore, the collision species are anticipated to track along the 5A' surface, then undergo nonadiabatic transition to the inner limb of the 2A' surface, and finally cross to the reactive 1A' surface. The observed energy barrier probably accounts for the energy requirement to surmount the repulsive hill in the entrance. The findings of the nascent CaH product distributions may be reasonably interpreted from the nature of the intermediate structure and lifetime after the 2A'-1A' surface transition. The distinct product distributions between the Ca(4 1P1) and Mg(3 1P1) reactions with H2 may also be realized with the aid of the PES calculations. PMID- 15268424 TI - Statistical evaporation of rotating clusters. III. Molecular clusters. AB - Unimolecular evaporation of weakly bound clusters made of rigid molecules is considered from the points of view of statistical theories and molecular dynamics simulations. We explicitly work out expressions for the kinetic energy released and product angular momentum distributions within the sphere+sphere and sphere+linear rigid body assumptions of phase space theory (PST). Various approximations are investigated, including the shape of the interaction potential between the two fragments and the anharmonicity of the vibrational density of states. The comparison between phase space theory and simulation for nitrogen and methane clusters shows a quantitative agreement, thereby suggesting that PST is accurate in predicting statistical observables in a wide range of systems under various physical conditions. PMID- 15268425 TI - A study of the static yield stress in a binary Lennard-Jones glass. AB - The stress-strain relations and the yield behavior of a model glass (a 80:20 binary Lennard-Jones mixture) is studied by means of molecular dynamics simulations. In a previous paper it was shown that, at temperatures below the glass transition temperature, Tg, the model exhibits shear banding under imposed shear. It was also suggested that this behavior is closely related to the existence of a (static) yield stress (under applied stress, the system does not flow until the stress sigma exceeds a threshold value sigmay). A thorough analysis of the static yield stress is presented via simulations under imposed stress. Furthermore, using steady shear simulations, the effect of physical aging, shear rate and temperature on the stress-strain relation is investigated. In particular, we find that the stress at the yield point (the "peak"-value of the stress-strain curve) exhibits a logarithmic dependence both on the imposed shear rate and on the "age" of the system in qualitative agreement with experiments on amorphous polymers, and on metallic glasses. In addition to the very observation of the yield stress which is an important feature seen in experiments on complex systems like pastes, dense colloidal suspensions and foams, further links between our model and soft glassy materials are found. An example is the existence of hysteresis loops in the system response to a varying imposed stress. Finally, we measure the static yield stress for our model and study its dependence on temperature. We find that for temperatures far below the mode coupling critical temperature of the model (Tc = 0.435 in Lennard-Jones units), sigmay decreases slowly upon heating followed by a stronger decrease as Tc is approached. We discuss the reliability of results on the static yield stress and give a criterion for its validity in terms of the time scales relevant to the problem. PMID- 15268426 TI - Intermolecular interactions in solution: elucidating the influence of the solvent. AB - A new approach for the analysis of intermolecular interactions in a solution is proposed. The changes in the interaction energy components due to the solvent effects are estimated on the basis of the interaction energy calculated in the presence of the electric field induced in a polarizable medium, or in the field of the effective fragment potentials. Obtained results indicate a significant increase in stabilization resulting from electrostatic interactions as a result of the cooperative interactions between interacting subsystems and solvent molecules. PMID- 15268427 TI - Thermodynamic manifestations of structure fluctuations in liquids. AB - Fluctuations of pressure and internal energy in a low-temperature liquid, generated by structure fluctuations, are related in a general and straightforward way to the differences between the long- and short-time thermodynamic susceptibilities. The relations are similar to, but not identical with, those between thermodynamic fluctuations and susceptibilities in classical statistical mechanics. The derivation is general and straightforward, and based on the assumption that the local structure is a slow-relaxing mode in low-temperature liquids and may be characterized by an order parameter. PMID- 15268428 TI - Local quasi-equilibrium description of slow relaxation systems. AB - We present a dynamical description of slow relaxation processes based on the extension of Onsager's fluctuation theory to systems in local quasi-equilibrium. A non-Markovian Fokker-Planck equation for the conditional probability density is derived, and from it we obtain the relaxation equation for the moments. We show that the fluctuation-dissipation theorem can be formulated in terms of the temperature of the system at local quasi-equilibrium which is related to that of the bath by means of a scaling factor revealing lack of thermal equilibrium. Our theory may be applied to a wide variety of systems undergoing slow relaxation. We discuss in particular slow dynamics in glassy systems and Brownian motion in a granular gas. PMID- 15268429 TI - Intermolecular electron transfer between coumarin dyes and aromatic amines in Triton-X-100 micellar solutions: evidence for Marcus inverted region. AB - Photoinduced electron transfer (ET) between coumarin dyes and aromatic amines has been investigated in Triton-X-100 micellar solutions and the results have been compared with those observed earlier in homogeneous medium. Significant static quenching of the coumarin fluorescence due to the presence of high concentration of amines around the coumarin fluorophore in the micelles has been observed in steady-state fluorescence studies. Time-resolved studies with nanosecond resolutions mostly show the dynamic part of the quenching for the excited coumarin dyes by the amine quenchers. A correlation of the quenching rate constants, estimated from the time-resolved measurements, with the free energy changes (DeltaG0) of the ET reactions shows the typical bell shaped curve as predicted by Marcus outer-sphere ET theory. The inversion in the ET rates for the present systems occurs at an exergonicity (-DeltaG0) of approximately 0.7-0.8 eV, which is unusually low considering the polarity of the Palisade layer of the micelles where the reactants reside. Present results have been rationalized on the basis of the two dimensional ET model assuming that the solvent relaxation in micellar media is much slower than the rate of the ET process. Detailed analysis of the experimental data shows that the diffusional model of the bimolecular quenching kinetics is not applicable for the ET reactions in the micellar solutions. In the present systems, the reactions can be better visualized as equivalent to intramolecular electron transfer processes, with statistical distribution of the donors and acceptors in the micelles. A low electron coupling (Vel) parameter is estimated from the correlation of the experimentally observed and the theoretically calculated ET rates, which indicates that the average donor -acceptor separation in the micellar ET reactions is substantially larger than for the donor--acceptor contact distance. Comparison of the Vel values in the micellar solution and in the donor--acceptor close contact suggests that there is an intervention of a surfactant chain between the interacting donor and acceptor in the micellar ET reaction. PMID- 15268430 TI - Double-quantum homonuclear correlation magic angle sample spinning nuclear magnetic resonance spectroscopy of dipolar-coupled quadrupolar nuclei. AB - A double-quantum homonuclear correlation nuclear magnetic resonance experiment for dipolar-coupled half-integer quadrupolar nuclei in solids is presented. The experiment is based on rotary resonance dipolar recoupling and uses bracketed spin-lock pulses to excite double-quantum coherence and later to convert it to the zero-quantum one. A central-transition-selective pi pulse at the beginning of the t1 evolution period differentiates coherence transfer pathways of double quantum coherences arising from coupled spins and from a single spin, so that the latter can be efficiently filtered out by phase cycling. The experiment was tested on an aluminophosphate molecular sieve AlPO4-14, a material with a variety of aluminum quadrupolar coupling constants, isotropic chemical shifts and homonuclear distances. In a two-dimensional spectrum aluminum dipolar couplings with internuclear distances between 2.9 and 5.5 A were resolved. Although the experiment requires an application of weak radio-frequency fields, frequency offsets did not affect its performance crucially. PMID- 15268431 TI - Phase separation in mixtures of Yukawa and charged Yukawa particles from Gibbs ensemble Monte Carlo simulations and the mean spherical approximation. AB - The phase equilibrium of mixtures of Yukawa and charged Yukawa particles is studied by means of Gibbs ensemble Monte Carlo (GEMC) simulation method and the mean spherical approximation (MSA). The strength of the Coulomb energy compared to that of the Yukawa attraction is characterized by a coupling constant. For low coupling constants a classical vapor--liquid phase separation appears with a good agreement between GEMC and the MSA. For high coupling constant, a phase separation between a salt poor and a salt rich phase occurs that resembles the phase equilibrium behavior of the solvent primitive model. PMID- 15268432 TI - Phase transformation in a lattice system in the presence of spin-exchange dynamics. AB - A joint action of the Glauber single-spin-flip and the Kawasaki spin-exchange mechanisms upon the processes of phase transformation is examined in the framework of the one-dimensional kinetic Ising model. It is shown that the addition of the Kawasaki dynamics to that of Glauber accelerates the process of phase transformation in the initial stage, but slows it down in later stages. For the truncated form of Glauber dynamics, which excludes the processes of splitting and coagulation of clusters, the addition of the Kawasaki dynamics always accelerates the phase transformation process. Acting alone, the Kawasaki mechanism provides a cluster growth proportional to t(1/2) (where t is the time) in the initial stage and proportional to t(1/3) (Lifshitz-Slyozov-Wagner law) in the intermediate stage. In the final stage, a cluster size approaches exponentially its equilibrium value. PMID- 15268433 TI - The Enskog-type theory of the velocity autocorrelations in the two-dimensional nematic of hard needles. AB - As it was shown from molecular dynamics of two-dimensional hard needles, the uniaxial velocity autocorrelation function (VACF) of this system exhibits a two time scale character. This corresponds to the symmetry of the particles. In this paper we provide a theory of the Enskog type that corroborates the idea that the VACF can be successfully described as a sum of two single decays. From the comparison between molecular dynamics and theoretical results, we show that the Enskog approach is a satisfactory kinetic theory, which functions as well in the nematic as in the isotropic phase. Different properties of VACFs have been investigated, in particular, the dependence on the orientational order, temperature, and particles' inertness. PMID- 15268434 TI - Ab initio melting curve of copper by the phase coexistence approach. AB - Ab initio calculations of the melting properties of copper in the pressure range 0-100 GPa are reported. The ab initio total energies and ionic forces of systems representing solid and liquid copper are calculated using the projector augmented wave implementation of density functional theory with the generalized gradient approximation for exchange-correlation energy. An initial approximation to the melting curve is obtained using an empirical reference system based on the embedded-atom model, points on the curve being determined by simulations in which solid and liquid coexist. The approximate melting curve so obtained is corrected using calculated free energy differences between the reference and ab initio system. It is shown that for system-size errors to be rendered negligible in this scheme, careful tuning of the reference system to reproduce ab initio energies is essential. The final melting curve is in satisfactory agreement with extrapolated experimental data available up to 20 GPa, and supports the validity of previous calculations of the melting curve up to 100 GPa. PMID- 15268435 TI - Absolute surface coverage measurement using a vibrational overtone. AB - Determination of absolute surface coverage with sub-monolayer sensitivity is demonstrated using evanescent-wave cavity ring-down spectroscopy (EW-CRDS) and conventional CRDS by employing conservation of the absolute integrated absorption intensity between gas and adsorbed phases. The first C-H stretching overtones of trichloroethylene (TCE), cis-dichloroethylene, and trans-dichloroethylene are probed using the idler of a seeded optical parametric amplifier having a 0.075 cm(-1) line width. Polarized absolute adsorbate spectra are obtained by EW-CRDS using a fused-silica monolithic folded resonator having a finesse of 28 500 at 6050 cm(-1), while absolute absorption cross sections for the gas-phase species are determined by conventional CRDS. A measure of the average transition moment orientation on the surface, which is utilized for the coverage determination, is derived from the polarization anisotropy of the surface spectra. Coverage measurement by EW-CRDS is compared to a mass-spectrometer-based surface-uptake technique, which we also employ for coverage measurements of TCE on thermally grown SiO(2) surfaces. To assess the potential for environmental sensing, we also compare EW-CRDS to optical waveguide techniques developed previously for TCE detection. PMID- 15268436 TI - Self-affine roughness influence on the friction coefficient for rubbers onto solid surfaces. AB - In this paper we investigate the influence of self-affine roughness on the friction coefficient mu(f) of a rubber body under incomplete contact onto a solid surface. The roughness is characterized by the rms amplitude w, the correlation length xi, and the roughness exponent H. It is shown that with increasing surface roughening at short and/or long length scales (decreasing H and/or increasing ratio w/xi, respectively), the maximum of the friction coefficient mu(f) shifts to lower sliding velocities. The latter occurs only for conditions of incomplete contact for small contact length scales lambda (>xi). PMID- 15268437 TI - Molecular dynamics study of the surface tension of a binary immiscible fluid. AB - The planar interface between two liquids having two degrees of affinity to mix has been studied by molecular dynamics simulations. The surface tension is calculated from the normal, PN, and transverse, PT, components of the pressure tensor P for a wide range of temperatures. An unusual increase in surface tension with increasing temperature is attributed to a pressure induced void transfer mechanism that is justified by basic thermodynamic arguments. This effect is diminished on the addition of a modest attractive potential between the two species, and there is a turnover point at higher temperatures beyond which the surface tension decreases with increasing temperature. An order parameter is identified as the gradient of the mole fraction distribution through the interfacial region. An additional effect is the dramatic inversion of the kinetic and potential contributions to the PN profile as the temperature is varied. It is found that a commonly used approximation for P, the Irving-Kirkwood 1 or IK1 method, results in a relatively modest unphysical variability in PN that weakly violates the condition of local mechanical stability. However, this artifact does not prevent the IK1 method from producing an interfacial tension which is nearly identical to that derived from the complete IK formula with no additional approximations. PMID- 15268438 TI - Wetting of nanopatterned surfaces: the hexagonal disk surface. AB - Metropolis Monte Carlo simulations are used to investigate the wetting of chemically nanopatterned surfaces, for the case of hexagonal disk patterns where liquid wishes to wet high-energy circular patches but not wet the background surface. We calculate the density profiles of saturated liquid adsorbed on a variety of such substrates, spanning the nanoscale to atomic scale patterns. In addition, statistical mechanical sum rules are used to obtain interfacial order parameters and interfacial free energies. We observe that Cassie's law is typically obeyed, together with an associated breakdown of the mechanical interpretation of Young's equation, for pattern wavelengths greater than 15 molecular diameters. Here, the adsorbed fluid exists as an array of hemi-drops. At about half this wavelength, the breakdown of Cassie's law lies within realistic energy scales and is associated with the unbending of the outer surface of adsorbed films. For atomic scale patterns, the usual interpretation of Young's equation is restored for films thicker than one monolayer. At high chemical contrast, when the monolayer in contact with high-energy regions would prefer to be crystalline, we observe a variety of exotic interfacial phenomena that may have technological significance. PMID- 15268439 TI - Grand canonical Monte Carlo simulation of argon adsorption at the surface of silica nanopores: effect of pore size, pore morphology, and surface roughness. AB - Argon adsorption (77 K) in atomistic silica nanopores of various sizes and shapes has been studied by means of grand canonical Monte Carlo simulations (GCMC). We discuss the effects of confinement (pore size), pore morphology (ellipsoidal, hexagonal, constricted pore), and surface texture (rough/smooth) on the thickness variation of the adsorbed film with pressure onto the disordered inner surface of porous materials (usually called t-plot or t-curve). We show that no confinement effect occurs when the diameter of the regular cylindrical pore is larger than 10 nm. For pores smaller than 6 nm, we find that the film thickness increases as the pore size decreases. We show that the adsorption isotherm in the rough pore can be described as the sum of an adsorbed amount similar to that found for a smooth pore (of the same radius) and a constant contribution due to atoms "trapped" in the infractuosities of the rough surface which act as a microporous texture. Simulation snapshots for Ar adsorption in hexagonal and ellipsoidal smooth pores indicate that at low pressures the gas/adsorbate interface retains memory of the pore shape and becomes cylindrical prior to the capillary condensation of the fluid in the pore. The film thickness in the hexagonal pore is close to that obtained for a cylindrical pore having a similar dimension. By contrast, we find that the film thickness for an ellipsoidal pore is always larger than that for an equivalent cylindrical pore (having the same length and volume but a circular section). We show that this effect strengthens as the pore size decreases and/or the pore asymmetry increases. Ar adsorption in a cylindrical constricted pore shows that the presence of the narrower part considerably modifies the adsorption mechanism. Finally, we report GCMC simulations of Ar adsorption (77 K) on a plane silica reference substrate for different intermolecular potentials. We discuss the effect of the interaction on the shape of the adsorption isotherm and compare our results with experiments. PMID- 15268440 TI - The surface temperature dependence of the inelastic scattering and dissociation of hydrogen molecules from metal surfaces. AB - High-dimensional, wave packet calculations have been carried out to model the surface temperature dependence of rovibrationally inelastic scattering and dissociation of hydrogen molecules from the Cu(111) surface. Both the molecule and the vibrating surface are treated fully quantum-mechanically. It is found, in agreement with experimental data, that the surface temperature dependence of a variety of dynamical processes has an Arrhenius form with an activation energy dependent on molecular translational energy and on the initial and final molecular states. The activation energy increases linearly with decreasing translational energy below the threshold energy. Above threshold the behavior is more complex. A quasianalytical model is proposed that faithfully reproduces the Arrhenius law and the translational energy dependence of the activation energy. In this model, it is essential to include quantized energy transfer between the surface and the molecule. It further predicts that for any process characterized by a large energy barrier and multiphonon excitation, the linear change in activation energy up to threshold has slope-1. This explains successfully the universal nature of the unit slope found experimentally for H2 and D2 dissociation on Cu. PMID- 15268441 TI - Statistical mechanics of bilayer membrane with a fixed projected area. AB - The equilibrium and fluctuation methods for determining the surface tension, sigma, and bending modulus, kappa, of a bilayer membrane with a fixed projected area are discussed. In the fluctuation method the elastic coefficients sigma and kappa are measured from the amplitude of thermal fluctuations of the planar membrane, while in the equilibrium method the free energy required to deform the membrane is considered. The latter approach is used to derive new expressions for sigma and kappa (as well as for the saddle-splay modulus), which relate them to the pair-interactions between the amphiphiles forming the membrane. We use linear response theory to argue that the two routes lead to similar values for sigma and kappa. This argument is confirmed by Monte Carlo simulations of a model membrane whose elastic coefficients are calculated using both methods. PMID- 15268442 TI - Dipole interactions and electrical polarity in nanosystems: the Clausius-Mossotti and related models. AB - Point polarizable molecules at fixed spatial positions have solvable electrostatic properties in classical approximation, the most familiar being the Clausius-Mossotti (CM) formula. This paper generalizes the model and imagines various applications to nanosystems. The behavior is worked out for a sequence of octahedral fragments of simple cubic crystals, and the crossover to the bulk CM law is found. Some relations to fixed moment systems are discussed and exploited. The one-dimensional dipole stack is introduced as an important model system. The energy of interaction of parallel stacks is worked out, and clarifies the diverse behavior found in different crystal structures. It also suggests patterns of self organization which polar molecules in solution might adopt. A sum rule on the stack interaction is found and tested. Stability of polarized states under thermal fluctuations is discussed, using the one-dimensional domain wall as an example. Possible structures for polar hard ellipsoids are considered. An idea is formulated for enhancing polarity of nanosystems by intentionally adding metallic coatings. PMID- 15268443 TI - Alkali halide nanocrystal growth and etching studied by AFM and modeled by MD simulations. AB - NaCl hillocks have been grown on the NaCl(100) monocrystalline surface by simply making the tip of an atomic force microscope cantilever interact with the surface. A quantitative discussion about the hillock dissolution process as well as physical stability of these ionic surfaces has been made. Molecular dynamics simulations helped us to interpret the experimental data. An explanation for the hillock formation and dissolution phenomena is also discussed. PMID- 15268444 TI - Swelling and deswelling kinetics of poly(N-isopropylacrylamide) gels. AB - Swelling and deswelling kinetics was investigated for three types of cylindrical poly(N-isopropylacrylamide) (PNIPA) gels differing in crosslink density. The temperature dependence curves of the volume of the gel specimens were different from one another. One of the gel specimens was considered as a critical gel showing the continuous volume phase transition. The volume change process of the specimens after a temperature jump was examined. In the deswelling processes with temperature jumps to temperatures higher than 35 degrees C, a phase separation was observed in the gel specimens and the volume change slowed down due to the homogenization after the phase separation. The value of the diffusion constant obtained without the phase separation decreased rapidly as temperature approaches the transition temperature. The rapid decrease for the critical gel indicates the emergence of the critical slowing-down. The value of the critical exponent for the correlation length suggests that the universality class for the volume phase transition of the critical PNIPA gel belongs to the class for the classical theory. PMID- 15268445 TI - Solid-fluid and solid-solid phase equilibrium in a model of n-alkane mixtures. AB - Solid-fluid and solid-solid phase equilibrium for binary mixtures of hard sphere chains modeling n-hexane, n-heptane, and n-octane has been calculated using Monte Carlo computer simulations. Thermodynamic integration was used to calculate the Gibbs free energy and chemical potentials in the solid and fluid phases from pure component reference values. A multiple stage free energy perturbation method was used to calculate the composition derivative of the Gibbs free energy. Equation of state and free energy data for the fluid phase indicate ideal solution behavior. Nonideality is much more significant in the solid phase with only partial solubility of shorter chains in the longer chains and essentially no solubility at the other end of the composition range. The miscibility decreases with increasing chain length difference between the components. For the model of n-hexane/n-octane mixtures solid--solid phase separation has been observed directly in some of the simulations, with the components segregating between the layers of the solid structure. The behavior is similar to that seen in some binary n-alkane mixtures with longer chain lengths but comparable chain length ratios between the components. Such phase separation, although indicated thermodynamically, is not seen directly in the simulations of the n-heptane/n octane mixture due to the difference in the pure component crystal structures. PMID- 15268446 TI - Molecular dynamics simulation of solvent-polymer interdiffusion: Fickian diffusion. AB - The interdiffusion of a solvent into a polymer melt has been studied using large scale molecular dynamics and Monte Carlo simulation techniques. The solvent concentration profile and weight gain by the polymer have been measured as a function of time. The weight gain is found to scale as t(1/2), which is expected for Fickian diffusion. The concentration profiles are fit very well assuming Fick's second law with a constant diffusivity. The diffusivity found from fitting Fick's second law is found to be independent of time and equal to the self diffusion constant in the dilute solvent limit. We separately calculated the diffusivity as a function of concentration using the Darken equation and found that the diffusivity is essentially constant for the concentration range relevant for interdiffusion. PMID- 15268447 TI - A time-integrated estimate of the entanglement mass in polymer melts in agreement with the one determined by time-resolved measurements. AB - We make a critical examination of how the entanglement molecular mass Me is determined from various measurable quantities. We are guided by reptation theory, where it is assumed that characteristic relaxations abruptly change and become equal to those of a chain moving in a Gaussian tube, as soon as the corresponding length scales surpass the tube diameter d or similarly as soon as the corresponding mass surpasses a critical value. Taking this critical mass as a definition of the "reptational" entanglement mass, we observe that all methods based on time-resolved quantities, such as the single-chain dynamic structure factor S(q,t) and the zero-shear relaxation modulus G(t), give the same result. We observe that such a value differs, beyond error bars, from that obtained from the plateau modulus, which is a time-integrated quantity. We have investigated an alternative definition of entanglement mass in terms of time-integrated quantities and observe that the value of this specific entanglement mass is consistent with that obtained from the time-resolved observables. We comment on possible reasons for the plateau modulus discrepancy. PMID- 15268448 TI - Determination of fluid--solid transitions in model protein solutions using the histogram reweighting method and expanded ensemble simulations. AB - Protein crystallization conditions are usually identified by empirical screening methods because of the complexity of the process, such as the existence of nonequilibrium phases and the different crystal forms that may result from changes in solution conditions. Here the crystallization of a model protein is studied using computer simulation. The model consists of spheres that have both an isotropic interaction of short range and anisotropic interactions between patch-antipatch pairs. The free energy of a protein crystal is calculated using expanded ensemble simulations of the Einstein crystal, and NpT-Monte Carlo simulations with histogram reweighting are used to determine the fluid-solid coexistence. The histogram reweighting method is also used to trace out the complete coexistence curve, including multiple crystal phases, with varying reduced temperature, which corresponds to changing solution conditions. At a patch-antipatch interaction strength five times that of the isotropic interaction, the protein molecules form a stable simple cubic structure near room temperature, whereas an orientationally disordered face-centered-cubic structure is favored at higher temperatures. The anisotropic attractions also lead to a weak first-order transition between orientationally disordered and ordered face centered-cubic structures at low temperature, although this transition is metastable. A complete phase diagram, including a fluid phase, three solid phases, and two triple points, is found for the six-patch protein model. A 12 patch protein model, consistent with the face-centered-cubic structure, leads to greater thermodynamic stability of the ordered phase. Metastable liquid-liquid phase equilibria for isotropic models with varying attraction tails are also predicted from Gibbs ensemble simulations. PMID- 15268449 TI - Numerical prediction of absolute crystallization rates in hard-sphere colloids. AB - Special computational techniques are required to compute absolute crystal nucleation rates of colloidal suspensions. Using crystal nucleation of hard sphere colloids as an example, we describe in some detail the novel computational tools that are needed to perform such calculations. In particular, we focus on the definition of appropriate order parameters that distinguish liquid from crystal, and on techniques to compute the kinetic prefactor that enters in the expression for the nucleation rate. In addition, we discuss the relation between simulation results and theoretical predictions based on classical nucleation theory. PMID- 15268450 TI - Discrete and heterogeneous rotational dynamics of single membrane probe dyes in gel phase supported lipid bilayer. AB - In order to probe the local dynamics of lipid bilayers in the gel phase, we measured the rotational time trajectories of a membrane probe, diI(3), in supported bilayers of DPPC (1,2-dipalmitoyl-sn-glycero-3-phosphocholine) using single molecule fluorescence polarization imaging. diI(3) has two hydrocarbon tails that mimic phospholipid tails and has its transition dipole moment lying mostly on the plane of the membrane; hence it is an excellent probe for rotational dynamics in membranes. Above the transition temperature, the probes are laterally mobile and do not display polarized emission. In the gel phase below the transition temperature, lateral mobility is severely reduced and the emission becomes polarized with its polarization direction changing in the milliseconds time scale. Molecule by molecule analysis of the rotational time scales revealed significant heterogeneities among molecules, much larger than would be due to statistical noise. Control experiments using small unilamellar vesicles suggest that the heterogeneities are not caused by surface interactions and are intrinsic to the gel phase membrane. The rotational dynamics is strongly temperature dependent and the thermally activated state for the rotational motion has a large entropic barrier (> 30kB), indicating that relatively large local disorder is required for the rotational motion to occur. Rotational hopping between discrete angles has been observed at the lowest temperatures (approximately 10 degrees C). Our results suggest that the gel phase membrane is not uniform at the microscopic level but is highly dynamic with the rigidity of local environments constantly changing. PMID- 15268451 TI - Comment on "Analysis of some integrals arising in the atomic four-electron problem" [J. Chem. Phys. 99, 3622 (1993)]. AB - A paper by F. W. King, "Analysis of some integrals arising in the atomic four electron problem", contains expressions which it is shown can be further simplified to an extent making the formulation significantly more efficient. Two errors in one of the equations are also identified. PMID- 15268455 TI - Quantum Monte Carlo for electronic excitations of free-base porphyrin. AB - Accurate calculations of allowed and nonallowed transitions in porphyrin are reported. Using the quantum Monte Carlo method in the diffusion Monte Carlo variant, the vertical transition between the ground state singlet and the second excited state singlet as well as the adiabatic transition between the ground state and the lowest triplet state have been computed for this 162-electron system. The present theoretical results are compared to experiment and to results of other theoretical methods. The diffusion Monte Carlo energy differences are found to be in excellent agreement with experiment. PMID- 15268456 TI - Reinvestigation of CS2 dissociation at 193 nm by means of product state-selective vacuum ultraviolet laser ionization and velocity imaging. AB - A branching ratio of 1.6 +/- 0.3 for S(3P)/S(1D) is obtained for the dissociation of CS2 with very low fluence 193 nm laser (less than 2 mJ/cm2), in which the S(3P) and S(1D) have been state-selectively ionized using VUV lasers at different wavelengths. The anisotropy parameters betamax(3P) = 0.8 and betamax(1D) = 1.9 indicate that these channels are preferentially populated at different geometries and the lifetime is very short. PMID- 15268457 TI - On the dynamics of coupled Bohmian and phase-space variables: a new hybrid quantum-classical approach. AB - A new approach to the coupling of quantum and classical dynamics is developed, by combining a hydrodynamic, Bohmian description for the quantum subsystem with a Liouville-space description for the classical subsystem. To this end, partial hydrodynamic moments are introduced, the dynamics of which is determined by a hierarchy of equations derived from the quantum Liouville equation. We focus on pure states (wave functions) and introduce a trajectory representation in a hybrid hydrodynamic-Liouvillian phase space. The interleaved trajectory dynamics is guided by a new type of quantum force. For illustration, we consider a pair of bilinearly coupled harmonic oscillators, for which the method is exact. PMID- 15268458 TI - Vibrational zero-point energies and thermodynamic functions beyond the harmonic approximation. AB - This paper compares harmonic and anharmonic zero-point energies and thermodynamic functions for a number of molecules of small and medium size. Anharmonic corrections cannot be neglected for quantitative studies, but can be obtained quite effectively by a perturbative treatment including cubic force constants to the second order and semidiagonal quartic constants to the first order. Simple finite difference equations provide all the necessary terms by at most 6N-11 Hessian evaluations, where N is the number of atoms in the system. Accurate values are obtained by this method using the Becke three parameter Lee-Yang-Parr functional, medium size basis sets, and, when needed, proper treatment of internal rotations. The whole model has been completely automated in the Gaussian package. PMID- 15268459 TI - On the use of Bennett's acceptance ratio method in multi-canonical-type simulations. AB - A common strategy for mapping coexistence curves is to employ multi-canonical (MUCA) sampling to simulate along a macrostate path connecting two phases. Central to this approach is the task of accurately calculating the importance weights used in the MUCA procedure, which are needed for both effective sampling and accurate determination of phase boundaries. The purpose of this study is to develop a strategy for determining the importance weights that is built upon Bennett's optimized acceptance ratio method. This approach is shown to be closely related to transition matrix schemes, and is used to compute the vapor-liquid equilibrium of a Lennard-Jones fluid and the liquid-liquid equilibrium of a n hexane/n-perfluorohexane mixture. For the Lennard-Jones system, the importance weights as a function of the number of particles "N" (at fixed temperature and volume) are obtained by using Bennett's method to estimate free energy differences between N and N+1 particle systems over the desired range of N values. In this application, the method is found to perform slightly better than a related transition matrix scheme. For the n-hexane/n-perfluorohexane liquid mixture, the method is designed to obtain weights as a function of composition (for fixed temperature, pressure, and total number of particles); in this case, the method is found to outperform the Gibbs ensemble approach. PMID- 15268460 TI - Slow manifold for a bimolecular association mechanism. AB - Finding the slow manifold for two-variable ordinary differential equation (ODE) models of chemical reactions with a single equilibrium is generally simple. In such planar ODEs the slow manifold is the unique trajectory corresponding to the slow relaxation of the system as it moves towards the equilibrium point. One method of finding the slow manifold is to use direct iteration of a functional equation; another method is to obtain a series solution of the trajectory differential equation of the system. In some cases these two methods agree order by-order in the singular perturbation parameter controlling the fast relaxation of the intermediate (complex). However, de la Llave has found a model ODE where the series method always diverges. Bimolecular association is an example of a chemical reaction where the series method for finding the slow manifold diverges but the iterative method converges. In this mechanism a complex is formed which can then undergo unimolecular decay, i.e., [reaction: see text]. The kinetics of this reaction are investigated and its properties compared with two other two step mechanisms where series expansion and iteration methods are equivalent: the Michaelis-Menten mechanism for enzyme kinetics, and the Lindemann-Christiansen mechanism of unimolecular decay in gas kinetics. PMID- 15268461 TI - On the efficient path integral evaluation of thermal rate constants within the quantum instanton approximation. AB - We present an efficient path integral approach for evaluating thermal rate constants within the quantum instanton (QI) approximation that was recently introduced to overcome the quantitative deficiencies of the earlier semiclassical instanton approach [Miller, Zhao, Ceotto, and Yang, J. Chem. Phys. 119, 1329 (2003)]. Since the QI rate constant is determined solely by properties of the (quantum) Boltzmann operator (specifically, by the zero time properties of the flux-flux and delta-delta correlation functions), it can be evaluated by well established techniques of imaginary time path integrals even for quite complex chemical reactions. Here we present a series of statistical estimators for relevant quantities which can be evaluated straightforwardly with any nonlinear reaction coordinates and general Hamiltonians in Cartesian space. To facilitate the search for the optimal dividing surfaces required by the QI approximation, we introduce a two-dimensional quantum free energy surface associated with the delta delta correlation function and describe how an adaptive umbrella sampling can be used effectively to construct such a free energy surface. The overall computational procedure is illustrated by the application to a hydrogen exchange reaction in gas phase, which shows excellent agreement of the QI rates with those obtained from quantum scattering calculations. PMID- 15268462 TI - Path integral calculation of thermal rate constants within the quantum instanton approximation: application to the H + CH4 --> H2 + CH3 hydrogen abstraction reaction in full Cartesian space. AB - The quantum instanton approximation for thermal rate constants of chemical reactions [Miller, Zhao, Ceotto, and Yang, J. Chem. Phys. 119, 1329 (2003)], which is modeled after the earlier semiclassical instanton approach, is applied to the hydrogen abstraction reaction from methane by a hydrogen atom, H + CH4 --> H2 + CH3, using a modified and recalibrated version of the Jordan-Gilbert potential surface. The quantum instanton rate is evaluated using path integral Monte Carlo approaches based on the recently proposed implementation schemes [Yamamoto and Miller, J. Chem. Phys. 120, 3086 (2004)]. The calculations were carried out using the Cartesian coordinates of all the atoms (thus involving 18 degrees of freedom), thereby taking explicit account of rotational effects of the whole system and also allowing the equivalent treatment of the four methane hydrogens. To achieve such a treatment, we present extended forms of the path integral estimators for relevant quantities that may be used for general N-atom systems with any generalized reaction coordinates. The quantum instanton rates thus obtained for the temperature range T = 200-2000 K show good agreement with available experimental data, which gives support to the accuracy of the underlying potential surface used. PMID- 15268463 TI - Auxiliary field Monte Carlo for charged particles. AB - This article describes Monte Carlo algorithms for charged systems using constrained updates for the electric field. The method is generalized to treat inhomogeneous dielectric media, electrolytes via the Poisson-Boltzmann equation and considers the problem of charge and current interpolation for off lattice models. We emphasize the differences between this algorithm and methods based on the electrostatic potential, calculated from the Poisson equation. PMID- 15268464 TI - A continuum, O(N) Monte Carlo algorithm for charged particles. AB - We introduce a Monte Carlo algorithm for the simulation of charged particles moving in the continuum. Electrostatic interactions are not instantaneous as in conventional approaches, but are mediated by a constrained, diffusing electric field on an interpolating lattice. We discuss the theoretical justifications of the algorithm and show that it efficiently equilibrates model electrolytes and polar fluids. In order to reduce lattice artifacts that arise from the interpolation of charges to the grid we implement a local, dynamic subtraction algorithm. This dynamic scheme is completely general and can also be used with other Coulomb codes, such as multigrid based methods. PMID- 15268465 TI - Solid-liquid phase coexistence of the Lennard-Jones system through phase-switch Monte Carlo simulation. AB - The phase-switch Monte Carlo method of Wilding and Bruce [Phys. Rev. Lett. 85, 5138 (2000)] is extended to enable calculation of solid-liquid phase coexistence for soft potentials. The method directly accesses coexistence information about a system while avoiding simulation of the interfacial region. Order parameters are introduced that allow one to define a path that connects liquid and crystalline phases. Transition matrix methods are employed to bias the sampling such that both phases are sampled in a rapid and efficient manner. Coexistence properties are determined through an analysis of specific volume probability distributions, which are generated naturally during a biased simulation. The approach is demonstrated with the Lennard-Jones system. Finite-size effects are examined and compared to those for the hard sphere system. In addition, two techniques are considered for accounting for long-range interactions. The methodology presented here is general and therefore provides a basis for its application to other soft systems. PMID- 15268466 TI - Calculation of electric dipole hypershieldings at the nuclei in the Hellmann Feynman approximation. AB - The third-rank electric hypershieldings at the nuclei of four small molecules have been evaluated at the Hartree-Fock level of theory in the Hellmann-Feynman approximation. The nuclear electric hypershieldings are closely related to molecular vibrational absorption intensities and a generalization of the atomic polar tensors (expanded in powers of the electric field strength) is proposed to rationalize these intensities. It is shown that the sum rules for rototranslational invariance and the constraints imposed by the virial theorem provide useful criteria for basis-set completeness and for near Hartree-Fock quality of nuclear shieldings and hypershieldings evaluated in the Hellmann Feynman approximation. Twelve basis sets of different size and quality have been employed for the water molecule in an extended numerical test on the practicality of the proposed scheme. The best results are obtained with the R12 and R12+ basis sets, designed for the calculation of electronic energies by the explicitly correlated R12 method. The R12 basis set is subsequently used to investigate three other molecules, CO, N2, and NH3, verifying that the R12 basis consistently performs very well. PMID- 15268467 TI - Calculation of electrostatic and polarization energies from electron densities. AB - We investigate procedures for calculating the electrostatic and polarization energies, Ees and Epol, associated with noncovalent interactions. The starting points are the electron densities of the isolated components and the complex; these could be obtained either computationally or experimentally. A slightly modified version of a scheme proposed by Gavezzotti is used to carry out numerical integrations over these electron densities. Our approach to estimating Epol is based upon partitioning the charge distributions of the components into overlapping and nonoverlapping regions. The effects of varying the integration parameters, computational techniques and basis sets are examined in detail for several noncovalently bound molecular dimers. Our results are in good agreement with the values of Ees and Epol produced by other methods, which require analytical integrations over interaction Hamiltonian matrix elements. PMID- 15268468 TI - The structure of the second-order reduced density matrix in density matrix functional theory and its construction from formal criteria. AB - Some formal requirements for the second-order reduced density matrix are discussed in the context of density matrix functional theory. They serve as a basis for the ad hoc construction of the second-order reduced density matrix in terms of the first-order reduced density matrix and lead to implicit functionals where the occupation numbers of the natural orbitals are obtained as diagonal elements of an idempotent matrix the elements of which represent the variational parameters to be optimized. The numerical results obtained from a first realization of such an implicit density matrix functional give excellent agreement with the results of full configuration interaction calculations for four-electron systems like LiH and Be. Results for H2O taken as an example for a somewhat larger molecule are numerically less satisfactory but still give reasonable occupation numbers of the natural orbitals and indicate the capability of density matrix functional theory to cope with static electron correlation. PMID- 15268469 TI - An algorithm for large scale density matrix renormalization group calculations. AB - We describe in detail our high-performance density matrix renormalization group (DMRG) algorithm for solving the electronic Schrodinger equation. We illustrate the linear scalability of our algorithm with calculations on up to 64 processors. The use of massively parallel machines in conjunction with our algorithm considerably extends the range of applicability of the DMRG in quantum chemistry. PMID- 15268470 TI - Detection of ClSO with time-resolved Fourier-transform infrared absorption spectroscopy. AB - ClSO was produced as an intermediate upon irradiating a flowing mixture of Cl2SO and Ar with a KrF excimer laser at 248 nm. A step-scan Fourier-transform infrared spectrometer coupled with a small multipass absorption cell was employed to detect time-resolved absorption spectrum of ClSO. A transient spectrum in the region 1120-1200 cm(-1), which diminished on prolonged reaction, is assigned to the S-O stretching (nu1) mode of ClSO. A spectrum with a resolution of 0.3 cm(-1) partially reveals rotational structure with the Q-branch at 1162.9 cm(-1). Calculations with density-functional theory (B3LYP/aug-cc-pVTZ) predict the geometry, vibrational, and rotational parameters of ClSO. An IR absorption spectrum of ClSO simulated based on predicted rotational parameters agrees satisfactorily with experimental results. ClSO produced from photolysis of Cl2SO at 248 nm is internally hot. PMID- 15268471 TI - Ornstein-Uhlenbeck diffusion quantum Monte Carlo study on the bond lengths and harmonic frequencies of some first-row diatomic molecules. AB - This article accesses the performance of the Ornstein-Uhlenbeck diffusion quantum Monte Carlo with regard to the calculation of molecular geometries and harmonic frequencies of H2, LiH, HF, Li2, LiF, CO, N2, and F2 molecules. A comparison of the results for the eight first-row diatomic molecules from experiments, CCSD(T)/6-311G(3df,3pd) and CCSD(T)/cc-pV5Z levels of theory as well as our work is given. The results presented show that quantum Monte Carlo is becoming powerful tools for ab initio electronic structure calculations. PMID- 15268472 TI - The C3-bending levels of the C3-Ar complex studied by optical spectroscopy and ab initio calculation. AB - The A-X electronic transition of C3-Ar, near 405 nm, has been studied by both laser-induced fluorescence and wavelength-resolved emission techniques. Emission spectra have been recorded from 14 vibrational levels of the A state of C3-Ar; these spectra consist of progressions in the ground state v2 and v4 vibrations (the in- and out-of-plane C3-bending motions, respectively). With increasing bending excitation, these ground state levels shift progressively downwards compared to those of free C3, indicating that the van der Waals complexes are becoming more tightly bound. The level structure of the two vibrations of C3-Ar has been fitted to a perturbed harmonic oscillator model, where the potential function has the form V = V1r cos theta + V2r2 cos 2theta (r is the amplitude of the C3-bending motion and theta gives the orientation of the rare gas atom relative to the plane of the bent C3 molecule). Ab initio calculations have been carried out for C3-Ar at the coupled-cluster singles, doubles (and triples)/correlation consistent polarization valence quadruple-zeta level. They predict that the C3-Ar complex is nearly T shaped at equilibrium, and that as the C3 molecule bends away from the linear configuration, the preferred orientation is "arrow" shaped. From the results of the best fit to the model and the emission spectral intensities, the relative orientation of the out-of-plane pi electron of the A-state complex and the Ar atom has been estimated. No bands of the Ar complex were found near the C3, A-X, (0,0) band, consistent with the fact that the A 1Piu, upsilon = 0 level of free C3 is strongly perturbed by triplet levels. In the excitation spectra of the Ar complex, the bands with upsilonb' > 0 show redshifts of about 16-36 cm(-1) compared to those of free C3, indicating that the A-state complex in these levels is more tightly bonded than the X-state complex. PMID- 15268473 TI - Intersections of potential energy surfaces of short-lived states: the complex analogue of conical intersections. AB - Whereas conical intersections between potential energy surfaces of bound states are well known, the interaction of short-lived states has been investigated only rarely. Here, we present several systematically constructed model Hamiltonians to study the topology of intersecting complex potential energy surfaces describing short-lived states: We find the general phenomenon of doubly intersecting complex energy surfaces, i.e., there are two points instead of one as in the case of bound states where the potential energy surfaces coalesce. In addition, seams of intersections of the respective real and imaginary parts of the potential energy surfaces emanate from these two points. Using the Sigma* and Pi* resonance states of the chloroethene anion as a practical example, we demonstrate that our complete linear model Hamiltonian is able to reproduce all phenomena found in explicitly calculated ab initio complex potential energy surfaces. PMID- 15268474 TI - Four-color hole burning spectra of phenol/ammonia 1:3 and 1:4 clusters. AB - The hole burning spectra of phenol/ammonia (1:3 and 1:4) clusters were measured by a newly developed four-color (UV-near-IR-UV-UV) hole burning spectroscopy, which is a kind of population labeling spectroscopy. From the hole burning spectra, it was found that single species is observed in an n = 3 cluster, while three isomers are observed simultaneously for n = 4. A possibility was suggested that the reaction efficiency of the hydrogen transfer from the electronically excited phenol/ammonia clusters, which was measured by a comparison with the action spectra of the corresponding cluster, depends on the initial vibronic levels. PMID- 15268475 TI - Ab initio study on the spectroscopy of CuCl2. I. Benchmark calculations on the X2Pig - 2Sigmag+ transition. AB - The modern theoretical predictions on the LambdaSSigma nature of the ground state of CuCl2 have led to different answers, depending on the type (DFT-based or ab initio) and the quality of the electronic correlation treatment; for this reason the X2Pig - 2Sigmag+ transition energy has been predicted to range from -1856 to +5887 cm(-1). The physical problem at hand lies in the difficulty of accurately describing the orientation of the 3d hole on the central Cu2+(3d9)/Cu+(3d94s1) ion (in the field of both chlorine ions), which implies the need of the most sophisticated nondynamic and dynamic electronic correlation treatments. We report here ab initio benchmark calculations using especially developed basis sets to study, at the CASSCF + CASPT2 and CASSCF + ACPF levels, the transition energy as well as the corresponding equilibrium geometries. The spin-orbit (SO) effects of both atoms were included in a second step through the effective Hamiltonian formalism, using the calibrated SO effective potentials developed by the Stuttgart group. Without SO at the CASSCF + ACPF level, the ground state is X2Pig but the vertical transition energy to the 2Sigmag+ is only 99 cm(-1) at 3.95 a.u. The inclusion of the SO effects leads to a Omega = 1/2 (59% 2Pig,41% 2Sigmag+) ground state, in contradiction with the Omega experimental value of 3/2. In a last step we show that the SO effects (and therefore the final Omega ordering) are critically dependent on the LambdaSSigma electronic energies, so that it is not impossible that the Omega ordering is actually changed. For theoreticians interest in this matter is not purely academic, since many properties of organometallic complexes are linked to such delicate physical effects. PMID- 15268476 TI - Spin and charge distribution in iron porphyrin models: a coupled cluster and density-functional study. AB - We recently performed detailed analyses of the electronic structure of low-spin iron porphyrins using density-functional theory (DFT). Both the spin-density distributions of the oxidized, ferric forms, as well as the changes in total charge density upon reduction to the ferrous forms have been explored. Here, we compare the DFT results with wave-function theory, more specifically, with the approximate singles and doubles coupled-cluster method (CC2). Different spin states are considered by studying representative models of low spin, intermediate spin, and high spin species. The CC2 calculations corroborate the DFT results; the spin density exhibits the same amount of molecular spin polarization, and the charge delocalization is of comparable magnitude. Slight differences in the descriptions are noted and discussed. PMID- 15268477 TI - 19F-19F spin-spin coupling constant surfaces for (HF)2 clusters: the orientation and distance dependence of the sign and magnitude of J(F-F). AB - Ab initio calculations using the equation-of-motion coupled cluster method have been carried out to investigate 19F-19F spin-spin coupling constants for a pair of HF molecules. The overall features of the J(F-F) coupling surface with respect to the F-F distance and the orientation of the pair of HF molecules reflect those of the Fermi-contact (FC) surface, although the FC term may not be a good quantitative estimate of J(F-F). The hydrogen-bonded HF dimer exhibits unusual behavior compared to other hydrogen-bonded complexes, since both the FC term and 2hJ(F-F) exhibit variations in sign and magnitude as the F-F distance changes and the linearity of the hydrogen bond is destroyed. The FC term for F-F coupling is relative small and negative for the equilibrium dimer. At the dimer F-F distance, the maximum negative value for the FC term is found for the linear arrangement F H...H-F, while the maximum positive value is found for the linear H-F...F-H arrangement, despite the fact that neither of these structures is bound. Changes in the sign and magnitude of the FC term are analyzed using the nuclear magnetic resonance triplet wave function model, which relates the orientation of magnetic nuclei to the phases of the wave functions for excited triplet states that couple to the ground state. The FC term for a particular orientation is a result of competing positive and negative contributions from different triplet states, the sign of each contribution being determined by the alignment of the nuclear magnetic moments in that state. Factors are identified which must play a role in determining which types of wave functions dominate. PMID- 15268478 TI - Disagreement between theory and experiment in the simplest chemical reaction: collision energy dependent rotational distributions for H + D2 --> HD(nu' = 3,j') + D. AB - We present experimental rotational distributions for the reaction H + D2 --> HD(nu' = 3,j') + D at eight different collision energies between 1.49 and 1.85 eV. We combine a previous measurement of the state-resolved excitation function for this reaction [Ayers et al., J. Chem. Phys. 119, 4662 (2003)] with the current data to produce a map of the relative reactive cross section as a function of both collision energy and rotational quantum number (an E-j' plot). To compare with the experimental data, we also present E-j' plots resulting from both time-dependent and time-independent quantum mechanical calculations carried out on the BKMP2 surface. The two calculations agree well with each other, but they produce rotational distributions significantly colder than the experiment, with the difference being more pronounced at higher collision energies. Disagreement between theory and experiment might be regarded as surprising considering the simplicity of this system; potential causes of this discrepancy are discussed. PMID- 15268479 TI - Collision energy dependence of the HD(nu' = 2) product rotational distribution of the H + D2 reaction in the range 1.30-1.89 eV. AB - An experimental and theoretical investigation of the collision energy dependence of the HD(nu' = 2,j') rotational product state distribution for the H + D2 reaction in the collision energy range of Ecol = 1.30-1.89 eV has been carried out. Theoretical results based on time-dependent and time-independent quantum mechanical methods agree nearly perfectly with each other, and the agreement with the experiment is good at low collision energies and very good at high collision energies. This behavior is in marked contrast to a previous report on the HD(nu' = 3,j') product state rotational distribution [Pomerantz et al., J. Chem. Phys. 120, 3244 (2004)] where a systematic difference between experiment and theory was observed, especially at the highest collision energies. The reason for this different behavior is not yet understood. In addition, this study employs Doppler free spectroscopy to resolve an ambiguity in the E, F-X resonantly enhanced multiphoton ionization transition originating from the HD(nu' = 2,j' = 1) state, which is found to be caused by an accidental blending with the transition coming from the HD(nu' = 1,j' = 14) state. PMID- 15268480 TI - Fourier transform millimeter-wave spectroscopy of the ethyl radical in the electronic ground state. AB - The pure rotational spectrum of the ethyl radical (C2H5) has been detected for the first time with the Fourier transform millimeter-wave spectrometer. The ethyl radical is produced by discharging the C2H5I gas diluted in Ar. The 1(01)-0(00) rotational transition of the ethyl radical is observed in the frequency range from 43,680 to 43,780 MHz. The observed spectrum shows a very complicated pattern of the fine and hyperfine structures of a doublet radical with the nuclear spins of five protons. The fine and hyperfine components are assigned with the aid of measurements of the Zeeman splittings. As a result, the 22 lines are ascribed to the transitions in the ground vibronic state (A2"). The rotational constant, the spin-rotation interaction constant, and hyperfine interaction constants are determined by the least-squares fit. The Fermi contact term of the alpha-proton is determined to be -64.1654 MHz in the gas phase, indicating that the structure of the -CH2 is essentially planar. The present rotational spectroscopic study further supports that the methyl group of the ethyl radical can be regarded as a nearly free internal rotor with a low energy barrier. A few unassigned lines still remain, which may be vibrational satellites of the internal rotation mode. PMID- 15268481 TI - Experimental and quantum-chemical studies on photoionization and dissociative photoionization of CH2Br2. AB - The dissociative photoionization of CH2Br2 in a region approximately 10-24 eV was investigated with photoionization mass spectroscopy using a synchrotron radiation source. An adiabatic ionization energy of 10.25 eV determined for CH2Br2 agrees satisfactorily with predictions of 10.26 and 10.25 eV with G2 and G3 methods, respectively. Observed major fragment ions CH2Br+, CHBr+, and CBr+ show appearance energies at 11.22, 12.59, and 15.42 eV, respectively; minor fragment ions CHBr2+, Br+, and CH2+ appear at 12.64, 15.31, and 16.80 eV, respectively. Energies for formation of observed fragment ions and their neutral counterparts upon ionization of CH2Br2 are computed with G2 and G3 methods. Dissociative photoionization channels associated with six observed fragment ions are proposed based on comparison of determined appearance energies and predicted energies. An upper limit of DeltaH0f,298(CHBr+) < or = 300.7 +/- 1.5 kcal mol(-1) is derived experimentally; the adiabatic ionization energy of CHBr is thus derived to be < or = 9.17 +/- 0.23 eV. Literature values for DeltaH0f,298(CBr+) = 362.5 kcal mol( 1) and ionization energy of 10.43 eV for CBr are revised to be less than 332 kcal mol(-1) and 9.11 eV, respectively. Also based on a new experimental ionization energy, DeltaH0f,298(CH2Br2+) is revised to be 236.4 +/- 1.5 kcal mol(-1). PMID- 15268482 TI - Nuclear magnetic shielding and chirality IV. The odd and even character of the shielding response to a chiral potential. AB - We investigate the odd and even character of the shielding response in a chiral molecule (modeled by a Ne8 helix) when subjected to a chiral potential. We establish that the diastereomeric splittings are a measure of odd powers of Vodd. Implications for diastereomeric, splittings of Xe in handed cages with handed tethers are discussed. PMID- 15268483 TI - Density functional study on the circular dichroism of photoelectron angular distribution from chiral derivatives of oxirane. AB - The linear combination of atomic orbitals B-spline density functional method has been successfully applied to a series of four chiral derivatives of oxirane, to calculate the photoionization dynamical parameters, the circular dichroism in the angular distribution effect, and to identify trends along the series. The computational algorithm has proven numerically stable and computationally competitive. The photoionization cross section, asymmetry, and dichroic parameter profiles relative to valence orbitals have been systematically studied for the states which retain their nature along the series: the identified trends have been ascribed to the different electronic properties of the substituents. A rather unexpected sensitivity of the dichroic parameter to changes in the electronic structure has been found in many instances, making this dynamical property suitable to investigate the electronic structure of chiral compounds. The magnitude of the circular dichroism in the angular distribution effect does not seem to be associated with the initial state chirality, but rather to be governed by the ability of the delocalized photoelectron wave function to probe the asymmetry of the molecular effective potential. PMID- 15268484 TI - Third-order Douglas-Kroll relativistic coupled-cluster theory through connected single, double, triple, and quadruple substitutions: applications to diatomic and triatomic hydrides. AB - Coupled-cluster methods including through and up to the connected single, double, triple, and quadruple substitutions have been derived and implemented automatically for sequential and parallel executions by an algebraic and symbolic manipulation program TCE (TENSOR CONTRACTION ENGINE) for use in conjunction with a one-component third-order Douglas-Kroll approximation for relativistic corrections. A combination of the converging electron-correlation methods, the accurate relativistic reference wave functions, and the use of systematic basis sets tailored to the relativistic approximation has been shown to predict the experimental singlet-triplet separations within 0.02 eV (0.5 kcal/mol) for five triatomic hydrides (CH2, NH2+, SiH2, PH2+, and AsH2+), the experimental bond lengths (re or r0) within 0.002 angstroms, rotational constants (Be or B0) within 0.02 cm(-1), vibration-rotation constants (alphae) within 0.01 cm(-1), centrifugal distortion constants (De) within 2%, harmonic vibration frequencies (omegae) within 8 cm(-1) (0.4%), anharmonic vibrational constants (xomegae) within 2 cm(-1), and dissociation energies (D0(0)) within 0.02 eV (0.4 kcal/mol) for twenty diatomic hydrides (BH, CH, NH, OH, FH, AlH, SiH, PH, SH, ClH, GaH, GeH, AsH, SeH, BrH, InH, SnH, SbH, TeH, and IH) containing main-group elements across the second through fifth rows of the periodic table. In these calculations, spin-orbit effects on dissociation energies, which were assumed to be additive, were estimated from the measured spin-orbit coupling constants of atoms and diatomic molecules, and an electronic energy in the complete-basis-set, complete-electron-correlation limit has been extrapolated in two ways to verify the robustness of the results: One assuming Gaussian-exponential dependence of total energies on double through quadruple zeta basis sets and the other assuming n(-3) dependence of correlation energies on double through quintuple zeta basis sets. PMID- 15268485 TI - Electron-intramolecular-vibration interactions in positively charged phenanthrene edge-type hydrocarbons. AB - Electron-phonon interactions in positively charged phenanthrene-edge-type hydrocarbons such as phenanthrene, chrysene, and picene are studied. The C-C stretching modes around 1500 cm(-1) and the low-frequency modes around 500 cm(-1) strongly couple to the highest occupied molecular orbitals (HOMO) in phenanthrene edge-type hydrocarbons. The total electron-phonon coupling constants for the monocations (lHOMO) of 0.251, 0.135, and 0.149 eV for phenanthrene, chrysene, and picene, respectively, are estimated to be larger than those of 0.130, 0.107, and 0.094 eV for anthracene, tetracene, and pentacene, respectively. The phase patterns difference between the HOMO localized on carbon atoms which are located at the molecular edge in acene-edge-type hydrocarbons and the delocalized HOMO in phenanthrene-edge-type hydrocarbons is the main reason for the result. Strengths of orbital interactions between two neighboring carbon atoms in the HOMO become weaker with an increase in molecular size because the electron density on each carbon atom in the HOMO becomes smaller with an increase in molecular size in phenanthrene-edge-type hydrocarbons. On the other hand, the frontier orbitals of acene-edge-type hydrocarbons have somewhat nonbonding characters and thus cannot strongly couple to the totally symmetric vibrational modes compared with the frontier orbitals of phenanthrene-edge-type hydrocarbons. This is the reason why the lHOMO value for phenanthrene-edge-type hydrocarbons decreases with an increase in molecular size more significantly than that for acene-edge-type hydrocarbons, and the reason why the lHOMO value for polyphenanthrene with C2v geometry (0.033 eV) is estimated to be similar to that for polyacene (0.036 eV). The reorganization energies between the neutral molecules and the corresponding monocations for phenanthrene-edge-type hydrocarbons with large molecular size are estimated to be larger than those for acene-edge-type hydrocarbons with large molecular size. PMID- 15268486 TI - Direct observation of an isopolyhalomethane O-H insertion reaction with water: picosecond time-resolved resonance Raman (ps-TR3) study of the isobromoform reaction with water to produce a CHBr2OH product. AB - Picosecond time-resolved resonance Raman (ps-TR3) spectroscopy was used to obtain the first definitive spectroscopic observation of an isopolyhalomethane O-H insertion reaction with water. The ps-TR3 spectra show that isobromoform is produced within several picoseconds after photolysis of CHBr3 and then reacts on the hundreds of picosecond time scale with water to produce a CHBr2OH reaction product. Photolysis of low concentrations of bromoform in aqueous solution resulted in noticeable formation of HBr strong acid. Ab initio calculations show that isobromoform can react with water to produce a CHBr2(OH) O-H insertion reaction product and a HBr leaving group. This is consistent with both the ps-TR3 experiments that observe the reaction of isobromoform with water to form a CHBr2(OH) product and photolysis experiments that show HBr acid formation. We briefly discuss the implications of these results for the phase dependent behavior of polyhalomethane photochemistry in the gas phase versus water solvated environments. PMID- 15268487 TI - A molecular dynamics study of the dielectric properties of aqueous solutions of alanine and alanine dipeptide. AB - Molecular dynamics simulations were used to compute the frequency-dependent dielectric susceptibility of aqueous solutions of alanine and alanine dipeptide. We studied four alanine solutions, ranging in concentration from 0.13-0.55 mol/liter, and two solutions of alanine dipeptide (0.13 and 0.27 mol/liter). In accord with experiment we find a strong dielectric increment for both solutes, whose molecular origin is shown to be the zwitterionic nature of the solutes. The dynamic properties were analyzed based on a dielectric component analysis into solute, a first hydration shell, and all remaining (bulk) waters. The results of this three component decomposition were interpreted directly, as well as by uniting the solute and hydration shell component to a "suprasolute" component. In both approaches three contributions to the frequency-dependent dielectric properties can be discerned. The quantitatively largest and fastest component arises from bulk water [i.e., water not influenced by the solute(s)]. The interaction between waters surrounding the solute(s) (the hydration shell) and bulk water molecules leads to a relaxation process occurring on an intermediate time scale. The slowest relaxation process originates from the solute(s) and the interaction of the solute(s) with the first hydration shell and bulk water. The primary importance of the hydration shell is the exchange of shell and bulk waters; the self-contribution from bound water molecules is comparatively small. While in the alanine solutions the solute-water cross-terms are more important than the solute self-term, the solute contribution is larger in the dipeptide solutions. In the latter systems a much clearer separation of time scales between water and alanine dipeptide related properties is observed. The similarities and differences of the dielectric properties of the amino acid/peptide solutions studied in this work and of solutions of mono- and disaccharides and of the protein ubiquitin are discussed. PMID- 15268488 TI - Transient response of a Brownian particle with general damping. AB - We study the transient response of a Brownian particle with general damping in a system of metastable potential well. The escape rate is evaluated as a function of time after an infinite wall is removed from the potential barrier. It takes a relaxation time for the rate to reach its limit value and this rate relaxation time differs from the relaxation time of the majority of the probability around the bottom of the potential well. The rate relaxation time is found to depend on the temperature as well as the damping constant. It involves the diffusion time and the instanton time, in general agreement with recent studies of the overdamped case by Bier et al. [Phys. Rev. E 59, 6422 (1999)]. PMID- 15268489 TI - Formation of HArF in solid Ar revisited: are mobile vacancies involved in the matrix-site conversion at 30 K? AB - The HArF molecule can occupy in solid Ar thermally unstable and stable configurations, and their microscopic structure is not understood at the moment. We present additional experimental results on the formation of two HArF configurations and analyze them with emphasis on possible reactions of the unstable configuration with matrix vacancies to form the stable configuration. We conclude that the existing computational scenarios do not describe fully the present experimental data. In order to explain qualitatively the experimental results, two tentative models are discussed. The first model is based on local mobility of matrix vacancies produced during photolysis and the second model considers isomerization of the HArF at Arn supermolecule. More importantly, the present results constitute the experimental basis for future theoretical studies. PMID- 15268490 TI - Sticking of CO to crystalline and amorphous ice surfaces. AB - We present results of classical trajectory calculations on the sticking of hyperthermal CO to the basal plane (0001) face of crystalline ice Ih and to the surface of amorphous ice Ia. The calculations were performed for normal incidence at a surface temperature Ts = 90 K for ice Ia, and at Ts = 90 and 150 K for ice Ih. For both surfaces, the sticking probability can be fitted to a simple exponentially decaying function of the incidence energy, Ei: Ps = 1.0e( Ei(kJ/mol)/90(kJ/mol)) at Ts = 90 K. The energy transfer from the impinging molecule to the crystalline and the amorphous surface is found to be quite efficient, in agreement with the results of molecular beam experiments on the scattering of the similar molecule, N2, from crystalline and amorphous ice. However, the energy transfer is less efficient for amorphous than for crystalline ice. Our calculations predict that the sticking probability decreases with Ts for CO scattering from crystalline ice, as the energy transfer from the impinging molecule to the warmer surfaces becomes less efficient. At high Ei (up to 193 kJ/mol), no surface penetration occurs in the case of crystalline ice. However, for CO colliding with the amorphous surface, a penetrating trajectory was observed to occur into a large water pore. The molecular dynamics calculations predict that the average potential energy of CO adsorbed to ice Ih is -10.1 +/- 0.2 and -8.4 +/- 0.2 kJ/mol for CO adsorbed to ice Ia. These values are in agreement with previous experimental and theoretical data. The distribution of the potential energy of CO adsorbed to ice Ia was found to be wider (with a standard deviation sigma of 2.4 kJ/mol) than that of CO interacting with ice Ih (sigma = 2.0 kJ/mol). In collisions with ice Ia, the CO molecules scatter at larger angles and over a wider distribution of angles than in collisions with ice Ih. PMID- 15268491 TI - Ultrafast carrier dynamics in single-walled carbon nanotubes probed by femtosecond spectroscopy. AB - Ultrafast carrier dynamics in individual semiconducting single-walled carbon nanotubes was studied by femtosecond transient absorption and fluorescence measurements. After photoexcitation of the second van Hove singularity of a specific tube structure, the relaxation of electrons and holes to the fundamental band edge occurs to within 100 fs. The fluorescence decay from this band is dependent on the excitation density and can be rationalized by exciton annihilation theory. In contrast to fluorescence, the transient absorption has a distinctly different time and intensity dependence for different tube structures, suggesting a branching to emissive and trap states following photoexcitation. PMID- 15268492 TI - Cooperative radiative and nonradiative effects in K2NaScF6 codoped with V3+ and Er3+. AB - K2NaScF6 crystals codoped with V3+ and Er3+ exhibit some novel cooperative near IR to visible upconversion processes at cryogenic temperatures. V3+ mainly acts as a broadband sensitizer. The V3+ 3T1g --> 3T2g excitation between 13,500 and 15,500 cm(-1), after fast relaxation to V3+ 1T2g, can be transferred to Er3+ 4I(11/2), and then upconversion takes place. Four upconversion mechanisms are identified and characterized. For narrow-band laser excitation the overall efficiency of the upconversion processes is low. However, at 12 K for broadband excitation, such as in a lamp, between 12,000 and 14,500 cm(-1) the number of emitted visible photons is roughly doubled by codoping V3+ in addition to Er3+. PMID- 15268493 TI - The efficiency of photolyase and indole complexes to repair DNA containing dimers of pyrimidine: a theoretical analysis of the electron transfer reactions. AB - We analyze the effects of competing reactions to the efficiency of enzymatic splitting of pyrimidine dimers formed in DNA by the incidence of ultraviolet radiation. This is accomplished with the aid of a formula that expresses the efficiency of the repair in terms of parameters that regulate the reaction rates for primary and for back long-range electron transfers taking place in the process. Comparison of experimental data with estimations on account of this formula supports early conjectures in the literature that attribute the relative high performance of the enzymatic complexes of photolyase to its ability to suppress the back reaction. PMID- 15268494 TI - Ab initio study of the alternating current impedance of a molecular junction. AB - The small-bias conductance of the C6 molecule, stretched between two metallic leads, is studied using time-dependent density functional theory within the adiabatic local density approximation. The leads are modeled by jellium slabs, the electronic density and the current density are described on a grid, whereas the core electrons and the highly oscillating valence orbitals are approximated using standard norm-conserving pseudopotentials. The jellium leads are supplemented by a complex absorbing potential that serves to absorb charge reaching the edge of the electrodes and hence mimic irreversible flow into the macroscopic metal. The system is rapidly exposed to a ramp potential directed along the C6 axis, which gives rise to the onset of charge and current oscillations. As time progresses, a fast redistribution of the molecular charge is observed, which translates into a direct current response. Accompanying the dc signal, alternating current fluctuations of charge and currents within the molecule and the metallic leads are observed. These form the complex impedance of the molecule and are especially strong at the plasmon frequency of the leads and the lowest excitation peak of C6. We study the molecular conductance in two limits: the strong coupling limit, where the edge atoms of the chain are submerged in the jellium and the weak coupling case, where the carbon atoms and the leads do not overlap spatially. PMID- 15268495 TI - Ca at C82 isomers: computed temperature dependency of relative concentrations. AB - Relative concentrations of nine isomers of Ca at C82 derived from the C82 isolated-pentagon-rule satisfying cages are computed in a wide temperature interval. The computations are based on the Gibbs energy constructed from partition functions supplied with molecular parameters from density functional theory calculations. Five structures show significant populations at higher temperatures: C2v > Cs > C2 > C3v > Cs. The computed relative stabilities agree well with available observations. PMID- 15268496 TI - van der Waals interactions across stratified media. AB - Working at the Lifshitz level, we investigate the van der Waals interactions across a series of layers with a periodic motif. We derive the complete form of the van der Waals interaction as an explicit function of the number of periodic layers. We then compare our result with an approximation based on an anisotropic continuum representation of the stratified medium. Satisfactory agreement between discrete-layer and continuum models is reached only for thicknesses of ten or more layers. PMID- 15268497 TI - Time-resolved spectroscopic behavior of Fe2O3 and ZnFe2O4 nanocrystals. AB - Using nanosecond (ns) and femtosecond (fs) time-resolved absorption spectroscopies (pump-probe technique), the carrier dynamics in transition metal oxide nanocrystals of alpha-Fe2O3 and ZnFe2O4 was studied during the photolysis process. For Fe2O3 and ZnFe2O4 nanocrystals, the fs measurements detect similar profiles of a positive nonlinear absorption in their capped nanocrystals, whereas much weak signals in the naked particles. In the nanosecond measurements Fe2O3 and ZnFe2O4 nanocrystals show obvious excitation-power dependent absorption properties and at the low pump power they show weak photobleaching, but at high pump power they produce positive nonlinear absorptions. For Fe2O3 nanocrystals, the threshold power of negative absorption (bleach) to positive absorption increases with reducing size, whereas for the ZnFe2O4 samples, the threshold powers reach minimum at a critical size of 11 nm, grow for both the bigger and the smaller nanocrystals. These results reflect the influences of their microscopic magnetic couplings and carrier correlation on biexciton absorption in Fe2O3 and ZnFe2O4 nanocrystals. All the results indicate that the time resolved photoabsorption techniques are useful to study the microscopic spin interactions and carrier correlations in transition metal oxide nanocrystals. PMID- 15268498 TI - Effect of exchangeable cation on the swelling property of 2:1 dioctahedral smectite--a periodic first principle study. AB - We used both localized and periodic calculations on a series of monovalent (Li+, Na+, K+, Rb+, Cs+) and divalent (Mg2+, Ca2+, Sr2+, Ba2+) cations to monitor their effect on the swelling of clays. The activity order obtained for the exchangeable cations among all the monovalent and divalent series studied: Ca2+ > Sr2+ > Mg2+ > Rb+ > Ba2+ > Na+ > Li+ > Cs+ > K+. We have shown that, in case of dioctahedral smectite, the hydroxyl groups play a major role in their interaction with water and other polar molecules in the presence of an interlayer cation. We studied both type of clays, with a different surface structure and with/without water using a periodic calculation. Interlayer cations and charged 2:1 clay surfaces interact strongly with polar solvents; when it is in an aqueous medium, clay expands and the phenomenon is known as crystalline swelling. The extent of swelling is controlled by a balance between relatively strong swelling forces and electrostatic forces of attraction between the negatively charged phyllosilicate layer and the positively charged interlayer cation. We have calculated the solvation energy at the first hydration shell of an exchangeable cation, but the results do not correspond directly to the experimental d-spacing values. A novel quantitative scale is proposed with the numbers generated by the relative nucleophilicity of the active cation sites in their hydrated state through Fukui functions within the helm of the hard soft acid base principle. The solvation effect thus measured show a perfect match with experiment, which proposes that the reactivity index calculation with a first hydration shell could rationalize the swelling mechanism for exchangeable cations. The conformers after electron donation or acceptance propose the swelling mechanism for monovalent and divalent cations. PMID- 15268499 TI - "Exact" surface free energies of iron surfaces using a modified embedded atom method potential and lambda integration. AB - Previously a new universal lambda-integration path and associated methodology was developed for the calculation of "exact" surface and interfacial free energies of solids. Such a method is in principle applicable to any intermolecular potential function, including those based on ab initio methods, but in previous work the method was only tested using a relatively simple embedded atom method iron potential. In this present work we apply the new methodology to the more sophisticated and more accurate modified embedded atom method (MEAM) iron potential, where application of other free- energy methods would be extremely difficult due to the complex many-body nature of the potential. We demonstrate that the new technique simplifies the process of obtaining "exact" surface free energies by calculating the complete set of these properties for the low index surface faces of bcc and fcc solid iron structures. By combining these data with further calculations of liquid surface tensions we obtain the first complete set of exact surface free energies for the solid and liquid phases of a realistic MEAM model system. We compare these predictions to various experimental and theoretical results. PMID- 15268500 TI - Melting behavior of one-dimensional zirconium nanowire. AB - In this paper, we analyze the melting behavior of zirconium nanowire using the results of a series of molecular dynamics simulations. Our calculation employs a well-fitted, tight-binding many-body potential for zirconium atoms. The melting point of the nanowire is predicted by the root-mean-square displacements for inner and outer shells. Our simulations predict two melting behaviors: one is the inner melting and the other is the outer melting. Our results reveal that the melting of nanowire starts from the inner shell atoms. The melting point of zirconium nanowire is lower than the bulk value (2125 K). Moreover, the melting point of the inner shell is lower than that of the outer shell. A coexistence of crystal and liquid units is observed in the melting process of nanowire. An investigation of local clusters is carried out to further analyze the melting mechanism of the nanowire. The presence of the local clusters 1331, 1321, 1211, etc. is an indication of disordered structures. The pair and angular correlation functions are also presented for the analysis of the melting behavior. It is not only the diffusion of single atom but the diffusion of clusters result in the occurrence of the melting. PMID- 15268501 TI - Fluorescence intensities of chromophores in front of a thin metal film. AB - The fluorescence intensity from a planar multilayered system with a chromophore separated from a gold film by a dielectric spacer is measured quantitatively. The direction of excitation and the spacer thickness are varied and the angular distribution of the emission is recorded as well as its polarization. The experimental data are compared to the predictions obtained from classical electromagnetic theory, taking into account the refractive indices of the layer system as well as the nonradiative decay rate and the relative orientation of absorption and emission dipole moments of the dye. Excellent agreement is found for a spacer thickness above 15 nm if proper values for these parameters are used. Samples with thinner spacer layers show significant deviations from classical theory. PMID- 15268502 TI - Monolayer/bilayer transition in Langmuir films of derivatized gold nanoparticles at the gas/water interface: an x-ray scattering study. AB - The microscopic structure of Langmuir films of derivatized gold nanoparticles has been studied as a function of area/particle on the water surface. The molecules (AuSHDA) consist of gold particles of mean core diameter D approximately 22 angstroms that have been stabilized by attachment of carboxylic acid terminated alkylthiols, HS-(CH2)15-COOH. Compression of the film results in a broad plateau of finite pressure in the surface pressure versus area/particle isotherm that is consistent with a first-order monolayer/bilayer transition. X-ray specular reflectivity (XR) and grazing incidence diffraction show that when first spread at large area/particle, AuSHDA particles aggregate two dimensionally to form hexagonally packed monolayer domains at a nearest-neighbor distance of a = 34 angstroms. The lateral positional correlations associated with the two dimensional (2D) hexagonal order are of short range and extend over only a few interparticle distances; this appears to be a result of the polydispersity in particle size. Subsequent compression of the film increases the surface coverage by the monolayer but has little effect on the interparticle distance in the close packed domains. The XR and off-specular diffuse scattering (XOSDS) results near the onset of the monolayer/bilayer coexistence plateau are consistent with complete surface coverage by a laterally homogeneous monolayer of AuSHDA particles. On the high-density side of the plateau, the electron-density profile extracted from XR clearly shows the formation of a bilayer in which the newly formed second layer on top is slightly less dense than the first layer. In contrast to the case of the homogeneous monolayer, the XOSDS intensities observed from the bilayer are higher than the prediction based on the capillary wave model and the assumption of homogeneity, indicating the presence of lateral density inhomogeneities in the bilayer. According to the results of Bragg rod measurements, the 2D hexagonal order in the two layers of the bilayer are only partially correlated. PMID- 15268503 TI - Polymer translocation through a nanopore. II. Excluded volume effect. AB - Following our previous study of a Gaussian chain translocation, we have investigated the transport of a self-avoiding chain from one sphere to another sphere through a narrow pore, using the self-consistent field theory formalism. The free energy landscape for polymer translocation is significantly modified by excluded volume interactions among monomers. The free energy barrier for the placement of one of the chain ends at the pore depends on the chain length N nonmonotonically, in contrast to the N-independence for Gaussian chains. This results in a nonmonotonic dependence of the average arrival time [tau0] on N for self-avoiding chains. When the polymer chain is partitioned between the donor and recipient spheres, a local free energy minimum develops, depending on the strength w of the excluded volume interaction and the relative sizes of the donor and recipient spheres. If the sizes of spheres are comparable, the average translocation time tau (the average time taken by the polymer, after the arrival at the pore, to convert from the donor to the recipient) increases with an increase in w for a fixed N value. On the other hand, for the highly asymmetric sizes of the donor and recipient spheres, tau decreases with an increase in w. As in the case of Gaussian chains, tau depends nonmonotonically on the pore length. PMID- 15268504 TI - A coarse-grained model and associated lattice Monte Carlo simulation of the coil helix transition of a homopolypeptide. AB - A new coarse-grained lattice model neglecting atomic details is proposed for the coil-helix transition and a new physical parameter is suggested to characterize a helical structure. In our model, each residue is represented by eight lattice sites, and side groups are not considered explicitly. Chirality and hydrogen bonding are taken into consideration in addition to chain connectivity and the excluded volume effect. Through a dynamic Monte Carlo simulation, the physical properties of the coil-helix transition of a single homopolypeptide have been produced successfully within a short computing time on the PC. We also examined the effects of the variation of chain configurations including chain size and chain shape, etc. A spatial correlation function has been introduced in order to characterize periodicity of a helical chain in a simple way. A propagation parameter and a nucleation parameter have also been calculated, which compares favorably with the results of the Zimm-Bragg theory for the coil-helix transition. PMID- 15268505 TI - Monte Carlo simulations of flexible polyanions complexing with whey proteins at their isoelectric point. AB - Electrostatic complexation of flexible polyanions with the whey proteins alpha lactalbumin and beta-lactoglobulin is studied using Monte Carlo simulations. The proteins are considered at their respective isoelectric points. Discrete charges on the model polyelectrolytes and proteins interact through Debye-Huckel potentials. Protein excluded volume is taken into account through a coarse grained model of the protein shape. Consistent with experimental results, it is found that alpha-lactalbumin complexes much more strongly than beta lactoglobulin. For alpha-lactalbumin, strong complexation is due to localized binding to a single large positive "charge patch," whereas for beta lactoglobulin, weak complexation is due to diffuse binding to multiple smaller charge patches. PMID- 15268506 TI - Nonlinear rheological behavior associated with structural transitions in block copolymer solutions via nonequilibrium molecular dynamics. AB - The nonequilibrium molecular dynamics computer simulation method was used to study microsegregated block copolymer systems in a selective solvent under a shear flow field. Two polymer concentrations were considered, 0.3 and 0.4, corresponding to the body centered cubic spherical and hexagonal cylindrical zero shear phases, respectively. As the shear rate increased, both systems exhibited two-stage shear thinning, a peak in the scalar pressure, and normal stress differences. Microscopic connections were investigated by calculating the gyration and bond orientation tensors and the interaction energies per particle. At high shear rates, polymer chains elongate and orient along the direction of shear, and this is accompanied by the breaking-up of domains. The structure rheology relation was discussed with regard to the morphological changes reported in our last study for the same systems. In particular, the structurally relevant critical values of the shear rate were found to delimit different behaviors of the shear rate-dependencies obtained in this work. PMID- 15268507 TI - Pressure-induced formation of diblock copolymer "micelles" in supercritical fluids. A combined study by small angle scattering experiments and mean-field theory. I. The critical micellization density concept. AB - We developed a simple mean-field theory to describe polymer and AB diblock copolymer phase separation in supercritical (SC) fluids. The highly compressible SC fluid has been described by using a phenomenological hole theory, properly extended to consider the solvent/polymer/vacancy pseudoternary mixture. The model has been applied to describe the phase behavior of AB-diblock copolymers under the assumption of a strong solvent selectivity for just one copolymer chain. In our model the solvent selectivity is a strong function of the external pressure because in compressible fluids vacancies reduce the number of favorable solvent polymer contacts. The combined effect of the pressure on the average solvent quality and selectivity for a single polymer chain makes the phase behavior of a diblock copolymer in SC fluids quite complex. Small angle neutron and x-ray scattering (SANS and SAXS) measurements have been performed on SC-CO2 solutions of different AB-diblock copolymers containing a perfluorinated chain. The data obtained over a wide range of pressure and temperature confirm our theoretical predictions. PMID- 15268508 TI - Pressure-induced formation of diblock copolymer "micelles" in supercritical fluids. A combined study by small angle scattering experiments and mean-field theory. II. Kinetics of the unimer-aggregate transition. AB - We developed a simple time-dependent mean-field theory to describe the phase separation kinetics of either homopolymers or AB-diblock copolymers in supercritical (SC) fluids. The model, previously used to describe the phase behavior of AB-block copolymers under the assumption of strong solvent selectivity for just one copolymer chain, has been extended to study the kinetics of the phase separation process. Time resolved small angle x-ray scattering (TR SAXS) measurements have been performed on different AB-diblock copolymers containing a perfluorinated chain and dissolved in SC-CO2. The data obtained over a wide range of pressure and temperature confirm our theoretical predictions. Particularly interesting is the presence of two relaxation frequencies for the homogeneous solution --> spherical aggregate transition, where the two relaxation processes depend on the depth of the pressure jump and on temperature. The whole phenomenon could be explained as an initial SC solvent/polymer phase separation followed by a slow reorientation process to form spherical aggregates driven by the copolymer solvophilic moiety. PMID- 15268509 TI - Correlations in simulated model bilayers. AB - Molecular dynamics simulations of a model bilayer in a solvent are used to determine the structure factors S(q) and other correlations at vanishing lateral tension of the system and at positive and negative tensions as well. The role played by curvature in the absence of surface tension, metastability, and phase transitions, and the dependence on the specific area, are discussed. Possible height-height correlation functions S(q) are examined and tested. PMID- 15268510 TI - Reassignment of the CH stretching frequency of CHF in the A electronic state. PMID- 15268513 TI - Molecular dynamics of glucose in solution: a quasielastic neutron scattering study. AB - The molecular dynamics of glucose dissolved in heavy water have been investigated at 280 K by the technique of quasielastic neutron scattering. The scattering was described by a dynamic structure factor that accounts for decoupled diffusive jumps and free rotational motions of the glucose molecules. With increasing glucose concentration, the diffusion constant decreases by a factor five and the time between jumps increases considerably. Our observations validate theoretical predictions concerning the impact of concentration on the environment of a glucose molecule and the formation of cages made by neighboring glucose molecules at higher concentrations. PMID- 15268514 TI - Gas-phase reactions of organic radicals and diradicals with ions. AB - Reactions of polyatomic organic radicals with gas phase ions have been studied at thermal energy using a flowing afterglow-selected ion flow tube (FA-SIFT) instrument. A supersonic pyrolysis nozzle produces allyl radical (CH2CHCH2) and ortho-benzyne diradical (o-C6H4) for reaction with ions. We have observed: [CH2CHCH2 + H3O+ --> C3H6+ + H2O], [CH2CHCH2 + HO- --> no ion products], [o-C6H4 + H3O+ --> C6H5+ + H2O], and [o-C6H4 + HO- --> C6H3- + H2O]. The proton transfer reactions with H3O+ occur at nearly every collision (kII approximately with 10( 9) cm3 s(-1)). The exothermic proton abstraction for o-C6H4 + HO- is unexpectedly slow (kII approximately with 10(-10) cm3 s(-1)). This has been rationalized by competing associative detachment: o-C6H4 + HO- --> C6H5O + e-. The allyl + HO- reaction proceeds presumably via similar detachment pathways. PMID- 15268515 TI - Direct determination of multipole moments of Cartesian Gaussian functions in spherical polar coordinates. AB - A new way of generating the multipole moments of Cartesian Gaussian functions in spherical polar coordinates has been established, bypassing the intermediary of Cartesian moment tensors. A new set of recurrence relations have also been derived for the resulting analytic integral values. The new method furnishes a conceptually simple and numerically efficient evaluation procedure for the multipole moments. The advantages over existing methods are documented. The results are relevant for the linear scaling quantum theories based on the fast multipole method. PMID- 15268516 TI - Density functional calculations of the vibronic structure of electronic absorption spectra. AB - Calculations of the vibronic structure in electronic spectra of large organic molecules based on density functional methods are presented. The geometries of the excited states are obtained from time-dependent density functional (TDDFT) calculations employing the B3LYP hybrid functional. The vibrational functions and transition dipole moment derivatives are calculated within the harmonic approximation by finite difference of analytical gradients and the transition dipole moment, respectively. Normal mode mixing is taken into account by the Duschinsky transformation. The vibronic structure of strongly dipole-allowed transitions is calculated within the Franck-Condon approximation. Weakly dipole allowed and dipole-forbidden transitions are treated within the Franck-Condon Herzberg-Teller and Herzberg-Teller approximation, respectively. The absorption spectra of several organic pi systems (anthracene, pentacene, pyrene, octatetraene, styrene, azulene, phenoxyl) are calculated and compared with experimental data. For dipole-allowed transitions in general a very good agreement between theory and experiment is obtained. This indicates the good quality of the optimized geometries and harmonic force fields. Larger errors are found for the weakly dipole-allowed S0 --> S1 transition of pyrene which can tentatively be assigned to TDDFT errors for the relative energies of excited states close to the target state. The weak bands of azulene and phenoxyl are very well described within the Franck-Condon approximation which can be explained by the large energy gap (>1.2 eV) to higher-lying excited states leading to small vibronic couplings. Once corrections are made for the errors in the theoretical 0 0 transition energies, the TDDFT approach to calculate vibronic structure seems to outperform both widely used ab initio methods based on configuration interaction singles or complete active space self-consistent field wave functions and semiempirical treatments regarding accuracy, applicability, and computational effort. Together with the parallel computer implementations employed, the present approach appears to be a valuable tool for a quantitative description and detailed understanding of electronic excitation processes in large molecules. PMID- 15268517 TI - Perturbation expansion theory corrected from basis set superposition error. I. Locally projected excited orbitals and single excitations. AB - The locally projected self-consistent field molecular orbital method for molecular interaction (LP SCF MI) is reformulated for multifragment systems. For the perturbation expansion, two types of the local excited orbitals are defined; one is fully local in the basis set on a fragment, and the other has to be partially delocalized to the basis sets on the other fragments. The perturbation expansion calculations only within single excitations (LP SE MP2) are tested for water dimer, hydrogen fluoride dimer, and colinear symmetric ArM+ Ar (M = Na and K). The calculated binding energies of LP SE MP2 are all close to the corresponding counterpoise corrected SCF binding energy. By adding the single excitations, the deficiency in LP SCF MI is thus removed. The results suggest that the exclusion of the charge-transfer effects in LP SCF MI might indeed be the cause of the underestimation for the binding energy. PMID- 15268518 TI - Assessing the efficiency of free energy calculation methods. AB - The efficiencies of two recently developed methods for calculating free energy changes along a generalized coordinate in a system are discussed in the context of other, related approaches. One method is based on Jarzynski's identity [Phys. Rev. Lett. 78, 2690 (1997)]. The second method relies on thermodynamic integration of the average force and is called the adaptive biasing force method [Darve and Pohorille, J. Chem. Phys. 115, 9169 (2001)]. Both methods are designed such that the system evolves along the chosen coordinate(s) without experiencing free energy barriers and they require calculating the instantaneous, unconstrained force acting on this coordinate using the formula derived by Darve and Pohorille. Efficiencies are analyzed by comparing analytical estimates of statistical errors and by considering two numerical examples-internal rotation of hydrated 1,2-dichloroethane and transfer of fluoromethane across a water-hexane interface. The efficiencies of both methods are approximately equal in the first but not in the second case. During transfer of fluoromethane the system is easily driven away from equilibrium and, therefore, the performance of the method based on Jarzynski's identity is poor. PMID- 15268519 TI - "Spectral implementation" for creating a labeled pseudo-pure state and the Bernstein-Vazirani algorithm in a four-qubit nuclear magnetic resonance quantum processor. AB - A quantum circuit is introduced to describe the preparation of a labeled pseudo pure state by multiplet-component excitation scheme which has been experimentally implemented on a 4-qubit nuclear magnetic resonance quantum processor. Meanwhile, we theoretically analyze and numerically investigate the low-power selective single-pulse implementation of a controlled-rotation gate, which manifests its validity in our experiment. Based on the labeled pseudo-pure state prepared, a 3 qubit Bernstein-Vazirani algorithm has been experimentally demonstrated by spectral implementation. The "answers" of the computations are identified from the split peak positions in the spectra of the observer spin, which are equivalent to projective measurements required by the algorithms. PMID- 15268520 TI - Army ants algorithm for rare event sampling of delocalized nonadiabatic transitions by trajectory surface hopping and the estimation of sampling errors by the bootstrap method. AB - The most widely used algorithm for Monte Carlo sampling of electronic transitions in trajectory surface hopping (TSH) calculations is the so-called anteater algorithm, which is inefficient for sampling low-probability nonadiabatic events. We present a new sampling scheme (called the army ants algorithm) for carrying out TSH calculations that is applicable to systems with any strength of coupling. The army ants algorithm is a form of rare event sampling whose efficiency is controlled by an input parameter. By choosing a suitable value of the input parameter the army ants algorithm can be reduced to the anteater algorithm (which is efficient for strongly coupled cases), and by optimizing the parameter the army ants algorithm may be efficiently applied to systems with low-probability events. To demonstrate the efficiency of the army ants algorithm, we performed atom-diatom scattering calculations on a model system involving weakly coupled electronic states. Fully converged quantum mechanical calculations were performed, and the probabilities for nonadiabatic reaction and nonreactive deexcitation (quenching) were found to be on the order of 10(-8). For such low probability events the anteater sampling scheme requires a large number of trajectories ( approximately 10(10)) to obtain good statistics and converged semiclassical results. In contrast by using the new army ants algorithm converged results were obtained by running 10(5) trajectories. Furthermore, the results were found to be in excellent agreement with the quantum mechanical results. Sampling errors were estimated using the bootstrap method, which is validated for use with the army ants algorithm. PMID- 15268521 TI - Relativistic hydrodynamics of magnetic and dielectric fluids in interaction with the electromagnetic field. AB - The semirelativistic hydrodynamic equations of motion of de Groot and Mazur [Non Equilibrium Thermodynamics (North-Holland, Amsterdam, 1962)] for a fluid with polarization and magnetization are derived from the relativistic energy-momentum conservation equation, mass conservation, and Maxwell's equations on the basis of a systematic expansion in inverse powers of the velocity of light. A modification of the de Groot-Suttorp field energy-momentum tensor [de Groot and Suttorp, Foundations of Electrodynamics (North-Holland, Amsterdam, 1972)] based on a shift of terms to the internal energy density of the fluid yields relatively simple expressions for energy density, energy current density, momentum density, and stress tensor. PMID- 15268522 TI - Determination of the proton tunneling splitting of the vinyl radical in the ground state by millimeter-wave spectroscopy combined with supersonic jet expansion and ultraviolet photolysis. AB - The vinyl radical in the ground vibronic state produced in a supersonic jet expansion by 193 nm excimer laser photolysis of vinyl bromide was investigated by millimeter-wave spectroscopy. Due to the proton tunneling, the ground state is split into two components, of which the lower and higher ones are denoted as 0+ and 0-, respectively. Eight pure rotational transitions with Ka = 0 and 1 obeying a-type selection rules were observed for each of the 0+ and 0- states in the frequency region of 60-250 GHz. Tunneling-rotation transitions connecting the lower (0+) and upper (0-) components of the tunneling doublet, obeying b-type selection rules, were also observed in the frequency region of 190-310 GHz, including three R- and six Q-branch transitions. The observed frequencies of the pure rotational and tunneling-rotation transitions were analyzed by using an effective Hamiltonian in which the coupling between the 0+ and 0- states was taken into account. A set of precise molecular constants was obtained. Among others, the proton tunneling splitting in the ground state was determined to be DeltaE0 = 16,272(2) MHz. The potential barrier height was estimated to be 1580 cm(-1) from the proton tunneling splitting, by an analysis using a detailed one dimensional model. The spin-rotation and hyperfine interaction constants were also determined for the 0+ and 0- states together with the off-diagonal interaction constants connecting the 0+ and 0- states, epsilonab + epsilonba for the spin-rotation interaction and Tab for the hyperfine interaction of the alpha (CH) proton. The hyperfine interaction constants, due to the alpha proton and the beta (CH2) protons, are consistent with those derived from electron spin resonance studies. PMID- 15268523 TI - Theoretical study of the hydrogen atom transfer in the heterodimer indole-ammonia and comparison with experimental results. AB - Ab initio calculations on the heterodimer C8H6NH...NH3 are carried out for its ground, the excited pisigma*, and the ground cationic electronic states, enabling the description of hydrogen or proton transfer, respectively. Two-dimensional quantum-dynamical computations on the pisigma* potential surface help one to understand the mechanism and the time scale of the hydrogen transfer. Subsequent decay processes are discussed depending on the vibrational excitation of the ammonium constituent. Finally, the theoretical results obtained are used for the interpretation of the time-dependent signals observed in femtosecond pump-probe experiments. PMID- 15268524 TI - Intermolecular complexes of HArF and HKrF with CO. AB - Stable linear weakly bound hydrogen-bonded complexes of HArF and HKrF with the CO molecule have been predicted by ab initio computations at the MP2/6- 311+ +G(2d,2p) level of theory. The complexes, having stabilities in the order, FArH...CO>FKrH...CO>FArH...OC>FKrH...OC are compared. They exhibit unusual blueshifts of the Ar-H (Kr-H) stretching frequency, as well as contraction of the Ar-H (Kr-H) bond, and these effects are rationalized mainly in terms of shifts in the electron density of HArF (HKrF) on complexation, caused mainly by a combination of the intermolecular electrostatic interaction, electron-electron (Pauli) and nuclear-nuclear repulsion and charge density transferred from the CO molecule to the rare-gas-containing molecule. PMID- 15268525 TI - Probe of bending motion following the 1s(-1)pi* excitation of N2O. AB - The doubly degenerate core-excited Pi state of N2O splits into two due to the static Renner-Teller effect. The lower state, A1, has a bent stable geometry and the molecule excited to this state starts to deform itself toward this bent geometry. To probe the effect of the potential energy surfaces of the core excited A1 states on the nuclear motion, we measure the momenta of the three atomic ions in coincidence by means of the ion momentum imaging technique. We find that the potential energy surface affects the molecular deformation significantly. N2O in the terminal N 1s(-1)3piA1 excited state is observed to be bent more than that in the central N 1s(-1)3piA1 excited state. This means that N2O in the terminal N 1s(-1)3piA1 excited state bends faster than that in the central N 1s(-1)3piA1 excited state. When the excitation energy is decreased within the 1s(-1)3pi resonances, the nuclear motion in the A1 states becomes faster. This is interpreted by the notion that the excitation occurs onto the steeper slope part of the potential energy surface of the excited state for the lower excitation energy. The branching ratio of the A1 excitation increases with the decrease in the excitation energy. PMID- 15268526 TI - Properties of selected diatomics using variational Monte Carlo methods. AB - Using variational Monte Carlo and highly accurate trial wave functions optimized by Filippi and Umrigar, we calculate a number of molecular properties for the ground state of Li2, Be2, B2, C2, N2, O2, and F2. This is the first time that many of these properties have been computed. PMID- 15268527 TI - Modeling benzene with single-site potentials from ab initio calculations: a step toward hybrid models of complex molecules. AB - Extensive ab initio calculations at the MP2/6-31G* level have been carried out to sample the energy surface for the interactions of the benzene dimers. This database has been used to parameterize two anisotropic single-site models, meant to be used as building blocks in hybrid models of complex, liquid crystal forming molecules. A quadrupolar Gay-Berne (GBQIII) and an S-function (SF) Corner potentials have been obtained in this way. Their ability to reproduce, qualitatively at least, the phase diagram as well as energetic and structural properties of benzene has been tested with Monte Carlo simulations and compared with previous literature potentials, GBQI [S. Gupta et al., Mol. Phys. 65, 961 (1988)] and GBQII [T. R. Walsh, Mol. Phys. 100, 2867 (2002)]. It turned out that GBQI showed no melting transition in the temperature range explored (100-400 K), while GBQII underwent a phase transition from solid to gas, with no liquid phase. Conversely, both models parameterized on our database of ab initio interaction energies (GBQIII and SF) gave rise to a stable liquid phase. Melting has been observed between 100 and 150 K (GBQIII) and in the range 300-350 K (SF), i.e., substantially below and slightly above the experimental value at ambient pressure, 278 K. The description of the crystal structure of benzene at atmospheric pressure is also in better agreement with experimental data if the SF model is used, while positional correlations in the liquid are better described by the GBQIII potential. The S-function potential is also computationally more convenient. These results could be useful in the semirealistic modeling of more complex molecules. PMID- 15268528 TI - Ultrafast nonadiabatic dynamics: quasiclassical calculation of the transient photoelectron spectrum of I2(-).(CO2)8. AB - In this paper we investigate the transient photoelectron spectrum of I2(-) in CO2 clusters recently measured by Neumark and co-workers. This work reveals a rich excited state dynamics with various competing electronic output channels. We find good agreement with experiments and we are able to relate the transient signal to different dynamical events that occur during the evolution of the cluster and its fragmentation products. PMID- 15268529 TI - Statisticodynamical approach of state distributions in the products of four-atom planar unimolecular reactions. I. Formal developments for conserved vibrations. AB - Traditional statistical approaches, entirely based on transition state theory (TST), do not allow the description of rotational state distributions in the products of indirect reactions governed by short-range forces. Owing to the interpretative power of TST, this limitation has long been acting as a brake upon a deep understanding of determining attributes of indirect reaction dynamics. Recently, however, we developed a statisticodynamical approach (SDA) of final state distributions for triatomic unimolecular reactions [P. Larregaray, L. Bonnet, and J. C. Rayez, J. Chem. Phys. 114, 3349 (2001); Phys. Chem. Chem. Phys. 4, 1577 (2002); 4, 1781 (2002)]. The approach combines TST for the description of state distributions at the transition state (TS) and the linear transformation model for the description of their alteration on the way from the TS to the products. The whole description is mostly analytical, thus keeping the rationalizing spirit of TST. The goal of the present series is to extend SDA to the case of four-atom planar unimolecular reactions, assuming that internal vibrations of the nascent products are conserved from the TS on. This first part is concerned with formal developments while the remaining parts deal with their validation and application, in particular to the fragmentation of isocyanic acid. PMID- 15268530 TI - Statisticodynamical approach of state distributions in the products of four-atom planar unimolecular reactions. II. Validation and distribution shape analysis in the barrier case. AB - In the first part of this series, we proposed a statisticodynamical approach of state distributions in the products of four-atom planar unimolecular reactions governed by short-range forces. In this second part, the approach is tested against quasiclassical trajectory calculations on an ab initio potential energy surface. The process considered is the fragmentation of isocyanic acid in the first excited singlet electronic state. The study leads to a very good agreement between both methods. In addition to that, we pinpoint in the barrier case the main mechanical parameters governing the shape of rotational state distributions. It appears that these parameters are related to two distinct physical effects. The first one is of the impulsive type. The second, already observed in triatomic processes, is the so-called bending effect. PMID- 15268531 TI - Critical evaluation of approximate quantum decoherence rates for an electronic transition in methanol solution. AB - We present a quantum molecular dynamics calculation of a semiclassical decoherence function to evaluate the accuracy of alternative short-time approximations for coherence loss in the dynamics of condensed phase electronically nonadiabatic processes. The semiclassical function from mixed quantum-classical molecular dynamics simulations and frozen Gaussian wave packets is computed for the electronic transition of an excited state excess electron to the ground state in liquid methanol. The decoherence function decays on a 10 fs time scale that is qualitatively similar to the aqueous case. We demonstrate that it is the motion of the hydrogen atom, and, in particular, the hydrogen rotation around the oxygen-methyl bond which is predominantly responsible for destroying the quantum correlations between alternative states. Multiple time scales due to the slower diffusive nuclear modes, which dominate the solvation response of methanol, do not contribute to the coherence loss. The choice of the coordinate representation is investigated in detail and concluded to be irrelevant to the decay. Changes in both nuclear momenta and positions on the two alternative potential surfaces are found to contribute to decoherence, the former dominating at short times (t < 5 fs), the latter controlling the decay at longer times. Various short-time approximations to the full dynamics for the decoherence function are tested for the first time. The present treatment rigorously develops the short-time description and establishes its range of validity. Whereas the lowest-order short-time approximation proves to be a very good approximation up to about 5 fs, we also find that it bounds the decay of the decoherence function. After 5 fs, the coherence decay in fact becomes faster than the single Gaussian predicted in the lowest-order short-time limit. This decay is well reflected by an enhanced low-order approximation, which is also easily computed from equilibrium classical forces. PMID- 15268532 TI - Searching for nanostructures of the cubic mesophase of liquid crystal molecules, BABH8. AB - Nanostructures of a thermotropic cubic phase forming liquid crystal compound, 1,2 bis-[4-n-octyloxy-benzoyl]-hydrazine was studied by molecular dynamics (MD) simulations. A model for its cubic phase structure was proposed, which was constructed from the building unit of a locally orientational ordered "bundle" consistent with the cell parameter and the space group (Ia3d) from the recent x ray results. Stability of the model structure was studied by multinanosecond MD simulations. A periodic nanostructure with 2.6 nm periodicity [coincides with the Ia3d (211) reflection] was kept up to 60 ns in the reduced pressure simulation which realizes the experimental value of density. However, the calculated fourth rank order parameter shows that the simulated final state above does not have cubic orientational symmetry but rather isotropic symmetry. PMID- 15268533 TI - Tunneling chemical reactions in solid parahydrogen: direct measurement of the rate constants of R + H2 --> RH + H (R = CD3,CD2H,CDH2,CH3) at 5 K. AB - Tunneling chemical reactions between deuterated methyl radicals and the hydrogen molecule in a parahydrogen crystal have been studied by Fourier transform infrared spectroscopy. The tunneling rates of the reactions R + H2 --> RH + H (R = CD3,CD2H,CDH2) in the vibrational ground state were determined directly from the temporal change in the intensity of the rovibrational absorption bands of the reactants and products in each reaction in solid parahydrogen observed at 5 K. The tunneling rate of each reaction was found to differ definitely depending upon the degree of deuteration in the methyl radicals. The tunneling rates were determined to be 3.3 x 10(-6) s(-1), 2.0 x 10(-6) s(-1), and 1.0 x 10(-6) s(-1) for the systems of CD3, CD2H, and CDH2, respectively. Conversely, the tunneling reaction between a CH3 radical and the hydrogen molecule did not proceed within a week's time. The upper limit of the tunneling rate of the reaction of the CH3 radical was estimated to be 8 x 10(-8) s(-1). PMID- 15268534 TI - Momentum representation of the solute/bath interaction in the dynamic theory of chemical processes in condensed phase. AB - For the system consisting of the chemically reactive solute immersed in the oscillator bath, we consider an approach based on the solute/medium interaction expressed in terms of momenta rather than coordinates. In the adiabatic representation the medium reorganization effects are suppressed, being hidden in the solute renormalized potential and new spectral density function. The advantage proposed by the bilinear interaction in momentum representation is its spatial uniformity important for approximate dynamical treatments. The procedure of explicit transforming a standard spectral density (coordinate representation of interaction) into the spectral density in adiabatic representation (momentum representation of interaction) is the main new result of the present study. Illustrative calculations for several types of spectral functions are performed. Special discussion is devoted to clarifying the nature of the slow diffusion coordinate, to which the present approach is mainly addressed. PMID- 15268535 TI - Thermal relaxation of glycerol and propylene glycol studied by photothermal spectroscopy. AB - In this paper we report on experimental data for the frequency and temperature dependence of the thermal properties of supercooled glycerol and propylene glycol. By using a photopyroelectric method the specific heat capacity and thermal conductivity were separately determined in a bandwidth of several decades. We have recently shown that the thermal conductivity has no relaxation behavior, which simplifies the analysis of our results. The relaxation behavior of the specific heat capacity is compared with literature results for other physical quantities and a detailed analysis of the temperature dependence of the relaxation parameters is presented. PMID- 15268536 TI - Hybrid diatomics-in-molecules-based quantum mechanical/molecular mechanical approach applied to the modeling of structures and spectra of mixed molecular clusters Arn(HCl)m and Arn(HF)m. AB - A new hybrid QM/DIM approach aimed at describing equilibrium structures and spectroscopic properties of medium size mixed molecular clusters is developed. This methodology is applied to vibrational spectra of hydrogen chloride and hydrogen fluoride clusters with up to four monomer molecules embedded in argon shells Arn(H(Cl/F))m (n = 1-62, m = 1-4). The hydrogen halide complexes (QM part) are treated at the MP2/aug-cc-pVTZ level, while the interaction between HX molecules and Ar atoms (MM part) is described in terms of the semiempirical DIM methodology, based on the proper mixing between neutral and ionic states of the system [Grigorenko et al., J. Chem. Phys. 104, 5510 (1996)]. A detailed analysis of the resulting topology of the QM/DIM potential energy (hyper-)surface in the triatomic subsystem Ar-HX reveals more pronounced nonadditive atomic induction and dispersion contributions to the total interaction energy in the case of the Ar-HCl system. An extension of the original analytical DIM-based potential in the frame of the present model as well as the current limitations of the method are discussed. A modified algorithm for the gradient geometry optimization, along with partly analytical force constant matrix evaluation, is developed to treat large cages of argon atoms around molecular clusters. Calculated frequency redshifts of HX stretching vibrations in the mixed clusters relative to the isolated hydrogen-bonded complexes are in good agreement with experimental findings. PMID- 15268537 TI - Nonequilibrium thermodynamic description of the three-phase contact line. AB - In order to give a solid foundation of the description of the motion of the three phase contact line, we develop the nonequilibrium thermodynamic description of the contact line. It is postulated that during its motion the contact line is featured as a separate thermodynamic system. Conservation laws are given for the excess densities of the components, the momentum and the energy along the line. The Gibbs law is formulated for the contact line and using this law the excess entropy production density along the line is constructed. This identifies the conjugate thermodynamic forces and fluxes for the contact line. Linear laws relating these quantities can then be given. The special case considered by Shikmurzaev, who gave the first satisfactory description of the motion of the contact line, is considered in more detail. A new boundary condition is found, which was not used by Shikhmurzaev. The need for such an additional boundary condition for this case was discussed in recent work by Billingham. PMID- 15268538 TI - Multicomponent nucleation: thermodynamically consistent description of the nucleation work. AB - A thermodynamically consistent formula is derived for the nucleation work in multicomponent homogeneous nucleation. The derivation relies on the conservative dividing surface which defines the nucleus as having specific surface energy equal to the specific surface energy sigma0 of the interface between the macroscopically large new and old phases at coexistence. Expressions are given for the radius of the nucleus defined by the conservative dividing surface and by the surface of tension. As a side result, the curvature dependence of the surface tension sigmaT of the nucleus defined by the surface of tension is also determined. The analysis is valid for nuclei of any size, i.e., for nucleation in the whole range of conditions between the binodal and the spinodal of the metastable old phase provided the inequality sigmaT < or = sigma0 is satisfied. It is found that under the conditions of validity of the analysis the nucleation rate is higher than the nucleation rate given by the classical nucleation theory. The general results are applied to nucleation of unary liquids or solids in binary gaseous, liquid or solid mixtures. PMID- 15268539 TI - Connection between the observable and centroid structural properties of a quantum fluid: application to liquid para-hydrogen. AB - It is shown that the discrepancy between path integral Monte Carlo [M. Zoppi et al., Phys. Rev. B 65, 092204 (2002)] and path integral centroid molecular dynamics [F. J. Bermejo et al., Phys. Rev. Lett. 84, 5359 (2000)] calculations of the static structure factor of liquid para-hydrogen can be explained based on a deconvolution equation connecting centroid and physical radial distribution functions. An explicit expression for the kernel of the deconvolution equation has been obtained using functional derivative techniques. In the superposition approximation, this kernel is given by the functional derivative of the effective potential with respect to the pairwise classical potential. Results of path integral Monte Carlo calculations for the radial distribution function and the static structure factor of liquid para-hydrogen are presented. PMID- 15268540 TI - Thermal conductivity of solid argon from molecular dynamics simulations. AB - The thermal conductivity of solid argon in the classical limit has been calculated by equilibrium molecular dynamic simulations using the Green-Kubo formalism and a Lennard-Jones interatomic potential. Contrary to previous theoretical reports, we find that the computed thermal conductivities are in good agreement with experimental data. The computed values are also in agreement with the high-temperature limit of the three-phonon scattering contribution to the thermal conductivity. We find that finite-size effects are negligible and that phonon lifetimes have two characteristic time scales, so that agreement with kinetic theory is obtained only after appropriate averaging of the calculated phonon lifetimes. PMID- 15268541 TI - Determination of solvation free energies by adaptive expanded ensemble molecular dynamics. AB - A new method of calculating absolute free energies is presented. It was developed as an extension to the expanded ensemble molecular dynamics scheme and uses probability density estimation to continuously optimize the expanded ensemble parameters. The new method is much faster as it removes the time-consuming and expertise-requiring step of determining balancing factors. Its efficiency and accuracy are demonstrated for the dissolution of three qualitatively very different chemical species in water: methane, ionic salts, and benzylamine. A recently suggested optimization scheme by Wang and Landau [Phys. Rev. Lett. 86, 2050 (2001)] was also implemented and found to be computationally less efficient than the proposed adaptive expanded ensemble method. PMID- 15268542 TI - Adiabatic population transfer in a liquid: taking advantage of a decaying target state. AB - The feasibility of efficient population transfer between an initial state and a decaying target state of the same parity without populating an intermediate state, in the presence of large-amplitude stochastic energy level fluctuations that mimic the dephasing in a solute molecule due to the influence of a solvent, is demonstrated theoretically. In particular, it is shown that a decaying target state, whose decay rate constant is large compared with the band width of picosecond laser pulses but small compared with the associated peak Rabi frequencies, can dramatically suppress the dephasing-induced nonadiabaticity associated with the dynamics of population transfer, resulting in, irrespective of the correlation time of stochastic energy level fluctuations, negligible population in the intermediate state and complete population transfer to the decaying target state. These results should further motivate experimental studies of optical control of molecular dynamics in a liquid. An interesting connection between our results and the quantum Zeno and anti-Zeno effects is also discussed. PMID- 15268543 TI - Second harmonic generation second hyperpolarizability of water calculated using the combined coupled cluster dielectric continuum or different molecular mechanics methods. AB - In this article we report the first calculations of second harmonic generation second hyperpolarizability of liquid water using coupled cluster/molecular mechanics (CC/MM) methods or coupled cluster/dielectric continuum (CC/DC) methods. The latter approach treats the solvent as an isotropic homogeneous fluid while the former accounts for the discrete nature of the solvent molecules. The CC/MM approach may include or exclude polarization effects explicitly. Alternatively, polarization effects may be included using perturbation theory. The CC descriptions implemented are the coupled cluster second-order approximate singles and doubles (CC2) and coupled cluster singles and doubles models. The second harmonic generation second hyperpolarizabilities are, depending on the model, obtained using either an analytical implementation of the cubic response function or using an analytical implementation of the quadratic response function combined with the finite field technique. The CC/MM results for the second harmonic generation second hyperpolarizability compare excellently with experimental data while a significant overestimation is found when using the CC/DC model. Particular, the cavity radius in the CC/DC calculations have an enormous effects on this fourth-order property. PMID- 15268544 TI - Atomic oxygen reactions with semifluorinated and n-alkanethiolate self-assembled monolayers. AB - The interaction of atomic oxygen (O(3P)) with semifluorinated self-assembled monolayers (CF-SAMs), two different n-alkanethiolate self-assembled monolayers, and a carbonaceous overlayer derived from an x-ray modified n-alkanethiolate SAM have been studied using in situ x-ray photoelectron spectroscopy. For short atomic oxygen exposures, CF-SAMs remain intact, an effect ascribed to the inertness of C-F and C-C bonds toward atomic oxygen and the well-ordered structure of the CF-SAMs. Following this initial induction period, atomic oxygen permeates through the CF3(CF2)7 overlayer and initiates reactions at the film/substrate interface, evidenced by the formation of sulfonate (RSO3) species and Au2O3. These reactions lead to the desorption of intact adsorbate chains, evidenced by the loss of carbon and fluorine from the film while the C(1s) spectral envelope and the C(1s)/F(1s) ratio remain virtually constant. In contrast, the reactivity of atomic oxygen with alkanethiolate SAMs is initiated at the vacuum/film interface, producing oxygen-containing carbon functional groups. Subsequent reactions of these new species with atomic oxygen lead to erosion of the hydrocarbon film. Experiments on the different hydrocarbon-based films reveal that the atomic oxygen-induced kinetics are influenced by the thickness as well as the structural and chemical characteristics of the hydrocarbon overlayer. Results from this investigation are also discussed in the context of material erosion by AO in low Earth orbit. PMID- 15268545 TI - Performance of the tau-dependent functionals in predicting the magnetic coupling of ionic antiferromagnetic insulators. AB - The performance of some kinetic energy density (tau) dependent functionals in predicting the effective Heisenberg exchange has been explored using the KNiF3 and K2NiF4 insulators as case examples. Our results show that this new generation of functionals represents an important improvement with respect to the current local and gradient corrected functionals yielding a semi-quantitative description of the antiferromagnetic coupling without the need of hybrid approaches thus avoiding the calculation of exact, Hartree-Fock exchange. This feature opens a wide field of application especially in solid state. PMID- 15268546 TI - Phase stability of nanocarbon in one dimension: nanotubes versus diamond nanowires. AB - Since their discovery in 1990, the study of sp2 bonded carbon nanotubes has grown into a field of research in it's own right; however the development of the sp3 analog, diamond nanowires, has been slow. A number of theoretical models have been proposed to compare the relative stability of diamond and graphite at the nanoscale; and more recently, to compare nanodiamonds and fullerenes. Presented here is a study of the phase stability of nanocarbon in one-dimension. The structural energies of carbon nanotubes and diamond nanowires have been calculated using density functional theory within the generalized gradient approximation, and used to determine the atomic heat of formation as a function of size. PMID- 15268547 TI - Elasticity of polyelectrolyte multilayer microcapsules. AB - We present a novel approach to probe elastic properties of polyelectrolyte multilayer microcapsules. The method is based on measurements of the capsule load deformation curves with the atomic force microscope. The experiment suggests that at low applied load deformations of the capsule shell are elastic. Using elastic theory of membranes we relate force, deformation, elastic moduli, and characteristic sizes of the capsule. Fitting to the prediction of the model yields the lower limit for Young's modulus of the polyelectrolyte multilayers of the order of 1-100 MPa, depending on the template and solvent used for its dissolution. These values correspond to Young's modulus of an elastomer. PMID- 15268548 TI - Low-temperature dynamics of weakly localized Frenkel excitons in disordered linear chains. AB - We calculate the temperature dependence of the fluorescence Stokes shift and the fluorescence decay time in linear Frenkel exciton systems resulting from the thermal redistribution of exciton population over the band states. The following factors, relevant to common experimental conditions, are accounted for in our kinetic model: (weak) localization of the exciton states by static disorder, coupling of the localized excitons to vibrations in the host medium, a possible nonequilibrium of the subsystem of localized Frenkel excitons on the time scale of the emission process, and different excitation conditions (resonant or nonresonant). A Pauli master equation, with microscopically calculated transition rates, is used to describe the redistribution of the exciton population over the manifold of localized exciton states. We find a counterintuitive nonmonotonic temperature dependence of the Stokes shift. In addition, we show that depending on experimental conditions, the observed fluorescence decay time may be determined by vibration-induced intraband relaxation, rather than radiative relaxation to the ground state. The model considered has relevance to a wide variety of materials, such as linear molecular aggregates, conjugated polymers, and polysilanes. PMID- 15268549 TI - Thermal expansion and impurity effects on lattice thermal conductivity of solid argon. AB - Thermal expansion and impurity effects on the lattice thermal conductivity of solid argon have been investigated with equilibrium molecular dynamics simulation. Thermal conductivity is simulated over the temperature range of 20-80 K. Thermal expansion effects, which strongly reduce thermal conductivity, are incorporated into the simulations using experimentally measured lattice constants of solid argon at different temperatures. It is found that the experimentally measured deviations from a T(-1) high-temperature dependence in thermal conductivity can be quantitatively attributed to thermal expansion effects. Phonon scattering on defects also contributes to the deviations. Comparison of simulation results on argon lattices with vacancy and impurity defects to those predicted from the theoretical models of Klemens and Ashegi et al. demonstrates that phonon scattering on impurities due to lattice strain is stronger than that due to differences in mass between the defect and the surrounding matrix. In addition, the results indicate the utility of molecular dynamics simulation for determining parameters in theoretical impurity scattering models under a wide range of conditions. It is also confirmed from the simulation results that thermal conductivity is not sensitive to the impurity concentration at high temperatures. PMID- 15268550 TI - Analysis of the phase transitions in alkyl-mica by density and pressure profiles. AB - In a previous work [Heinz, Castelijns, and Suter, J. Am. Chem. Soc. 115, 9500 (2003)], we developed an accurate force field and simulated the phase transitions in C18-mica (octadecyltrimethylammonium-mica) as well as the absence of such transitions in 2C18-mica (dioctadecyldimethylammonium-mica) between room temperature and 100 degrees C. Here we analyze (i) average z coordinates of the carbon atoms and interdigitation of the hydrocarbon bilayers, (ii) density profiles, and (iii) pressure profiles of the structures along all Cartesian axes. In C18-mica, the standard deviation in the z coordinate for the chain atoms is high and more than doubles in the disordered phase. The order-disorder transition is accompanied by a change in the orientation of the ammonium head group, as well as decreasing tensile and shear stress in the disordered phase. In 2C18-mica, the standard deviation in the z coordinate for the chain atoms is low and does not increase remarkably on heating. The backbones display a highly regular structure, which is slightly obscured by rotations in the C18 backbones and minor head group displacements at 100 degrees C. Close contacts between the bulky head groups with sidearms cause significant local pressure which is in part not relieved at 100 degrees C. An increase of the basal-plane spacing at higher temperature is found in both systems due to larger separation between the two hydrocarbon layers and an increased z spacing between adjacent chain atoms (=decreased tilt of the chains relative to the surface normal), and, in C18-mica only, a stronger upward orientation of the C18 chain at the ammonium head group. The likelihood for chain interdigitation between the two hydrocarbon layers is 24%-30% for C18-mica, and 65%-26% for 2C18-mica (for 20-100 degrees C). PMID- 15268551 TI - Transport properties of nitrogen in single walled carbon nanotubes. AB - Transport properties including collective and tracer diffusivities of nitrogen, modeled as a diatomic molecule, in single walled carbon nanotubes have been studied by equilibrium molecular dynamics at different temperatures and as a function of pressure. It is shown that while the asymptotic decay of the translational and rotational velocity autocorrelation function is algebraic, the collective velocity decays exponentially with the relaxation time related to the interfacial friction. The tracer diffusivity in the nanochannel, which is comparable in magnitude with diffusivity in the equilibrium bulk phase, depends only weakly on the conditions at the fluid-solid interface, whereas the collective diffusivity is a strong function of the hydrodynamic boundary conditions and is found to be three orders of magnitude higher than self diffusivity in carbon nanotubes and for the comparatively rough surface of the rare-gas tube it is one order of magnitude greater. A relationship between the collective diffusivity and the Maxwell coefficient describing wall collisions is obtained. The transport coefficients appear to be insensitive to the long-range details of the potential function. PMID- 15268552 TI - Water formation on Pd(111) by reaction of oxygen with atomic and molecular hydrogen. AB - In this work we have studied the steady-state reaction of molecular and atomic hydrogen with oxygen on a Pd(111) surface at a low total pressure (<10(-7) mbar) and at sample temperatures ranging from 100 to 1100 K. Characteristic features of the water formation rate Phi(pH2; pO2; TPd) are presented and discussed, including effects that are due to the use of gas-phase atomic hydrogen for exposure. Optimum impingement ratios (OIR) for hydrogen and oxygen for water formation and their dependence on the sample temperature have been determined. The occurring shift in the OIR could be ascribed to the temperature dependence of the sticking coefficients for hydrogen (SH2) and oxygen (SO2) on Pd(111). Using gas-phase atomic hydrogen for water formation leads to an increase of the OIR, suggesting that hydrogen abstraction via hot-atom reactions competes with H2O formation. The velocity distributions of the desorbing water molecules formed on the Pd(111) surface have been measured by time-of-flight spectroscopy under various conditions, using either gas-phase H atoms or molecular H2 as reactants. In all cases, the desorbing water flux could be represented by a Maxwellian distribution corresponding to the surface temperature, thus giving direct evidence for a Langmuir-Hinshelwood mechanism for water formation on Pd(111). PMID- 15268553 TI - Oxide-free oxygen incorporation into Ru(0001). AB - A smooth Ru(0001) surface prepared under ultra-high vacuum conditions has been loaded with oxygen under high-pressure (p approximately 1 bar) and low temperature (T < 600 K) conditions. Oxygen phases created in this way have been investigated by means of thermal desorption spectroscopy, low-energy electron diffraction, and ultraviolet photoelectron spectroscopy. The exposure procedures applied lead to oxygen incorporation into the subsurface region without creation of RuO2 domains. For oxygen exposures ranging from 10(11) to 10(14) L oxygen contents up to about 4 monolayer equivalent could be achieved. The oxygen incorporation is thermally activated. The CO oxidation reaction conducted at mild temperatures (T < 500 K) at a sample loaded with subsurface oxygen reaches CO --> CO2 conversion probabilities of 10(-3). PMID- 15268554 TI - Surface vibrations in alkanethiol self-assembled monolayers of varying chain length. AB - The effect of chain length on the low-energy vibrations of alkanethiol striped phase self-assembled monolayers on Au(111) was studied. We have examined the low energy vibrational structure of well-ordered, low-density 1-decanethiol (C10), 1 octanethiol (C8), and 1-hexanethiol (C6) to further understand the interaction between adsorbate and substrate. Dispersionless Einstein mode phonons, polarized perpendicularly to the surface, were observed for the striped phases of C10, C8, and C6 at 8.0, 7.3, and 7.3 meV, respectively. An overtone at 12.3 meV was also observed for C6/Au(111). These results, in concert with molecular dynamics simulations, indicate that the forces between the adsorbate and substrate can be described using simple van der Waals forces between the hydrocarbon chains and the Au substrate with the sulfur chemisorbed in the threefold hollow site. PMID- 15268555 TI - A test of the method of images at the surface of molecular materials. AB - The method of images is tested by comparing two ways of calculating the polarization energy in crystalline fullerene C60 and in bulk amorphous polyethylene (PE): (i) treating the whole molecular material microscopically, and (ii) replacing part of the material by a uniform dielectric continuum of the same relative permittivity. The method of images is accurate to within 5% once the distance of the charge from the surface of the dielectric continuum exceeds about twice the average spacing between the polarizable units in the molecular material. For C60 crystals the method of images always overestimates the magnitude of the polarization energy, partly because it ignores the reduction in the relative permittivity of the dielectric continuum near its surface. For amorphous PE the method of images can overestimate or underestimate the true result, depending on the local density around the charge. PMID- 15268556 TI - A polarizable continuum model for molecules at diffuse interfaces. AB - In this work we illustrate an extension of the polarizable continuum model to describe solvation effects on molecules at the interface between two fluid phases (liquid/liquid, liquid/vapor). This extension goes beyond the naive picture of the interface as a plane dividing two distinct dielectrics, commonly employed in continuum models. The main feature of the model is the use of a diffuse interface with an electric permittivity depending on the position. This characteristic clearly allows the study of simple interfaces as well as more complex membrane or multilayer structures. Moreover the smooth variation of the permittivity in the diffuse interface, in contrast to the sharp boundary between two regions, overcomes the numerical divergences due to charges placed at the boundary. The implementation of the model relies on the integral equation formalism, which allows one to calculate the reaction field acting on a molecule immersed in a dielectric environment once the proper Green's function is known. In the present case, such a Green's function is obtained numerically, allowing a large flexibility in the choice of the dielectric permittivity profile. The applications have been selected with the aim of illustrating the capabilities of the model; its present limitations are also discussed. PMID- 15268557 TI - Picosecond acoustic transmission measurements. I. Transient grating generation and detection of acoustic responses in thin metal films. AB - The technique of impulsive stimulated thermal scattering is extended to backside measurement of acoustic wave packets that have propagated through thin metal films following their generation by pulsed optical excitation, heating, and thermal expansion at the front side. The acoustic transmission measurement at the backside substantially isolates the acoustic responses from thermal and electronic responses of the metal film that often dominate acoustic reflection signals measured from the front side, and permits straightforward measurement of the acoustic response generated by optical excitation at a substrate-thin film interface. It can thus better distinguish among different factors that limit the bandwidth of the acoustic wave packet, an issue of concern in the measurement of high frequency responses. The paper that follows demonstrates the application of the backside measurement to a study of high frequency structural relaxation in the glass-forming liquid glycerol. PMID- 15268558 TI - Picosecond acoustic transmission measurements. II. Probing high frequency structural relaxation in supercooled glycerol. AB - The high frequency acoustic response of liquids is measured in a manner directly analogous to conventional ultrasonic measurements. Two thin metal films act as acoustic transducer and receiver for a liquid layer between them. Pulsed optical excitation generates high bandwidth wave packets in the transducer, and these are detected in the receiver after damping and dispersion by the liquid. This initial measurement probes structural relaxation dynamics of glycerol in the frequency range 2-20 GHz, for temperatures between 235 and 291 K. The analysis presented here demonstrates the presence of excess relaxation, not accounted for by either the alpha or beta relaxation of the mode-coupling theory, and suggests the presence of constant loss in the susceptibility spectrum of supercooled glycerol. PMID- 15268559 TI - Abstractive dissociation of oxygen over Al(111): a nonadiabatic quantum model. AB - The dissociation of oxygen on a clean aluminum surface is studied theoretically. A nonadiabatic quantum dynamical model is used, based on four electronically distinct potential energy surfaces characterized by the extent of charge transfer from the metal to the adsorbate. A flat surface approximation is used to reduce the computation complexity. The conservation of the helicopter angular momentum allows Boltzmann averaging of the outcome of the propagation of a three degrees of freedom wave function. The dissociation event is simulated by solving the time dependent Schrodinger equation for a period of 30 femtoseconds. As a function of incident kinetic energy, the dissociation yield follows the experimental trend. An attempt at simulation employing only the lowest adiabatic surface failed, qualitatively disagreeing with both experiment and nonadiabatic calculations. The final products, adsorptive dissociation and abstractive dissociation, are obtained by carrying out a semiclassical molecular dynamics simulation with surface hopping which describes the back charge transfer from an oxygen atom negative ion to the surface. The final adsorbed oxygen pair distribution compares well with experiment. By running the dynamical events backward in time, a correlation is established between the products and the initial conditions which lead to their production. Qualitative agreement is thus obtained with recent experiments that show suppression of abstraction by rotational excitation. PMID- 15268560 TI - Morphology of microphase separated domains in rod-coil copolymer melts. AB - We investigate the morphology of microphase separated domains in diblock copolymers where each chain consists of a stiff rod block and a flexible coil block. A simplified phenomenological model system is introduced, which is coarse grained in terms of the local concentration difference between the two blocks and the local director field of the rod part. Computer simulations of this set of time-evolution equations in two dimensions show in the weak segregation regime that the elastic energy in the rod-block rich domains affects drastically the structures of microphase separated domains. A coil-to-rod transition is incorporated into the model system to examine the elastic and anisotropic effects. The effects of the external electric field are also investigated to control the domain morphology. PMID- 15268561 TI - Computer simulation study of the global phase behavior of linear rigid Lennard Jones chain molecules: comparison with flexible models. AB - The global phase behavior (i.e., vapor-liquid and fluid-solid equilibria) of rigid linear Lennard-Jones (LJ) chain molecules is studied. The phase diagrams for three-center and five-center rigid model molecules are obtained by computer simulation. The segment-segment bond lengths are L = sigma, so that models of tangent monomers are considered in this study. The vapor-liquid equilibrium conditions are obtained using the Gibbs ensemble Monte Carlo method and by performing isobaric-isothermal NPT calculations at zero pressure. The phase envelopes and critical conditions are compared with those of flexible LJ molecules of tangent segments. An increase in the critical temperature of linear rigid chains with respect to their flexible counterparts is observed. In the limit of infinitely long chains the critical temperature of linear rigid LJ chains of tangent segments seems to be higher than that of flexible LJ chains. The solid-fluid equilibrium is obtained by Gibbs-Duhem integration, and by performing NPT simulations at zero pressure. A stabilization of the solid phase, an increase in the triple-point temperature, and a widening of the transition region are observed for linear rigid chains when compared to flexible chains with the same number of segments. The triple-point temperature of linear rigid LJ chains increases dramatically with chain length. The results of this work suggest that the fluid-vapor transition could be metastable with respect to the fluid solid transition for chains with more than six LJ monomer units. PMID- 15268562 TI - Interaction between charged anisotropic macromolecules: application to rod-like polyelectrolytes. AB - In this paper we propose a framework allowing one to compute the effective interactions between two anisotropic macromolecules, thereby generalizing the Derjaguin, Landau, Verwey, and Overbeek theory [E. J. W. Verwey and J. T. G. Overbeek, Theory of the Stability of Lyophobic Colloids (Elsevier, Amsterdam, 1948)] to nonspherical finite size colloids. We show in particular that the effective interaction potential remains anisotropic at all distances and provide an expression for the anisotropy factor. We then apply this framework to the case of finite rod-like polyelectrolytes. The calculation of the interaction energy requires the numerical computation of the surface charge profiles, which result here from a constant surface potential on the rod-like colloids. However, a simplified analytical description is proposed, leading to an excellent agreement with the full numerical solution. Conclusions on the phase properties of rod-like colloids are proposed in this context. PMID- 15268563 TI - Coupled folding-binding versus docking: a lattice model study. AB - Using a simple hydrophobic/polar protein model, we perform a Monte Carlo study of the thermodynamics and kinetics of binding to a target structure for two closely related sequences, one of which has a unique folded state while the other is unstructured. We obtain significant differences in their binding behavior. The stable sequence has rigid docking as its preferred binding mode, while the unstructured chain tends to first attach to the target and then fold. The free energy profiles associated with these two binding modes are compared. PMID- 15268564 TI - Elastic moduli of multiblock copolymers in the lamellar phase. AB - We study the linear elastic response of multiblock copolymer melts in the lamellar phase, where the molecules are composed of tethered symmetric AB diblock copolymers. We use a self-consistent field theory method, and introduce a real space approach to calculate the tensile and shear moduli as a function of block number. The former is found to be in qualitative agreement with experiment. We find that the increase in bridging fraction with block number, that follows the increase in modulus, is not responsible for the increase in modulus. It is demonstrated that the change in modulus is due to an increase in mixing of repulsive A and B monomers. Under extension, this increase originates from a widening of the interface, and more molecules pulled free of the interface. Under compression, only the second of these two processes acts to increase the modulus. PMID- 15268565 TI - Patch size effect on diffusion and incorporation in dilute suspension of partially active spheres. AB - The normalized overall rate constant kp/kf for diffuse-and-incorporate processes in dilute suspension of spheres partially covered with randomly distributed nonoverlapping active patches is studied with a sped-up Brownian dynamic simulation scheme. The normalized overall rate constant is found to increase with decreasing characteristic size of the active patch under the condition of fixed fcover, the surface area fraction covered by the active patches. A scaling relation is proposed as (kp/kf)NDL/[(1 + P) - (1 + P/fcover)(kp/kf)NDL] approximately Np(1/2) with Np being the number of active patches on the sphere, P a parameter characterizing the relative dominance of surface incorporation over diffusion, and NDL signifying the nondiffusion-limited condition. This scaling relation is verified with rate constant data from the sped-up Brownian dynamic simulation. From this scaling relation, the maximum achievable kp/kf is derived to be (1 + P)/(1 + P/fcover). This result implies that kp/kf can approach unity under the diffusion-limited situation even for a partially active sphere by reducing the size of active patch, while it is not possible to achieve unit kp/kf for nondiffusion-limited systems. PMID- 15268566 TI - Competition between compaction of single chains and bundling of multiple chains in giant DNA molecules. AB - It has been established that in a dilute solution individual giant DNA molecules undergo a large discrete transition between an elongated coil state and a folded compact state. On the other hand, in concentrated solutions, DNA molecules assemble into various characteristic states, including multichain aggregate, liquid crystalline, ionic crystal, etc. In this study, we compared single-chain and multiple-chain events by observing individual chains using fluorescence microscopy. We used spermidine, SPD(3+), as a condensing agent for giant DNA. When the concentration of DNA is below 1 microM in base-pair units, individual DNA molecules exhibit a transition from an elongated state to a compact state. When the concentration of DNA is increased to 10 microM, a thick fiberlike assembly of multiple chains appears. AFM measurements of this thick fiber revealed that more than tens of DNA molecules form a bundle structure with parallel ordering of the chains. The transition between single-chain compaction and bundle formation with multiple-chain assemblies was reproduced by a theoretical calculation. PMID- 15268567 TI - Static properties of end-tethered polymers in good solution: a comparison between different models. AB - We present a comparison between results, obtained from different simulation models, for the static properties of end-tethered polymer layers in good solvent. Our analysis includes data from two previous studies--the bond fluctuation model of Wittmer et al. [J. Chem. Phys. 101, 4379 (1994)] and the off-lattice bead spring model of Grest and Murat [Macromolecules 26, 3108 (1993)]. Additionally, we explore the properties of a similar off-lattice model simulated close to the Theta temperature. We show that the data for the bond fluctuation and the Grest Murat model can be analyzed in terms of scaling theory because chains are swollen inside the Pincus blob. In the vicinity of the Theta point the structure of the chains is essentially Gaussian in the Pincus blob. Therefore, the data for the second off-lattice model can be compared quantitatively to the self-consistent field theory. Different ways to determine the parameters of the self-consistent field theory are discussed. PMID- 15268568 TI - Growth dynamics of isotactic polypropylene single crystals during isothermal crystallization from a miscible polymeric solvent. AB - The present article presents a spatiotemporal growth of isotactic polypropylene (iPP) single crystals, melt crystallized from a polymeric solvent, i.e., poly (ethylene octene) copolymer that is known to be miscible with iPP. Optical and atomic force microscopic investigations reveal that the melt grown single crystals of iPP develop in the form of two parallel rows of crystal lamellae, but these crystals merge at the tips. To elucidate the mechanism of these emerging parallel rows of iPP crystals, a phase field model pertaining to solidification phenomena has been employed that involves a nonconserved crystal order parameter and a chain-tilting angle. This phase field model is based on the free energy of crystallization, having an asymmetric double well, and a tensorial surface free energy of the crystal interface coupled with a curvature elastic free energy that is possessed by the solid-liquid interface. The spatiotemporal simulation of iPP single crystal growth has been carried out on a square lattice based on the finite difference method for spatial steps and an explicit method for temporal steps with a periodic boundary condition. The appearance of the seemingly twin crystal is captured in the simulation, which may be attributed to the sector demarcation that is taking place in the anisotropically growing single crystal of iPP. PMID- 15268569 TI - Diffusion-diffusion correlation and exchange as a signature for local order and dynamics. AB - We demonstrate the use of new two-dimensional nuclear magnetic resonance experiments in the examination of local diffusional anisotropy under conditions of global isotropy. The methods, known as diffusion-diffusion correlation spectroscopy and diffusion exchange spectroscopy, employ successive pairs of magnetic field gradient pulses, with signal analysis using two-dimensional inverse Laplace transformation. Diffusional anisotropy is measured for water molecules in a polydomain lamellar phase lyotropic liquid crystal, 40 wt % nonionic surfactant C10E3 (C10H21O(CH2CH2O)6H) in H2O. PMID- 15268570 TI - Linear viscoelastic analysis of formation and relaxation of azobenzene polymer gratings. AB - Surface relief gratings on azobenzene containing polymer films were prepared under irradiation by actinic light. Finite element modeling of the inscription process was carried out using linear viscoelastic analysis. It was assumed that under illumination the polymer film undergoes considerable plastification, which reduces its original Young's modulus by at least three orders of magnitude. Force densities of about 10(11) N/m3 were necessary to reproduce the growth of the surface relief grating. It was shown that at large deformations the force of surface tension becomes comparable to the inscription force and therefore plays an essential role in the retardation of the inscription process. In addition to surface profiling the gradual development of an accompanying density grating was predicted for the regime of continuous exposure. Surface grating development under pulselike exposure cannot be explained in the frame of an incompressible fluid model. However, it was easily reproduced using the viscoelastic model with finite compressibility. PMID- 15268571 TI - Tunneling-induced spin alignment at low and zero field. AB - The transfer of rotational to spin angular momentum of CH3 groups according to the Haupt effect is shown to be independent of magnetic field strength, including zero field. Haupt enhanced pulsed nuclear resonance signals of gamma-picoline have been observed at fields below 50 mT with a sensitivity enhancement of more than 3 orders of magnitude over thermally polarized experiments. PMID- 15268572 TI - Rate constants for diffusive processes by partial path sampling. AB - We introduce a path sampling method for the computation of rate constants for complex systems with a highly diffusive character. Based on the recently developed transition interface sampling (TIS) algorithm this procedure increases the efficiency by sampling only parts of complete transition trajectories. The algorithm assumes the loss of memory for diffusive progression along the reaction coordinate. We compare the new partial path technique to the TIS method for a simple diatomic system and show that the computational effort of the new method scales linearly, instead of quadratically, with the width of the diffusive barrier. The validity of the memory loss assumption is also discussed. PMID- 15268573 TI - Three-body problem in quantum mechanics: hyperspherical elliptic coordinates and harmonic basis sets. AB - Elliptic coordinates within the hyperspherical formalism for three-body problems were proposed some time ago [V. Aquilanti, S. Cavalli, and G. Grossi, J. Chem. Phys. 85, 1362 (1986)] and recently have also found application, for example, in chemical reaction theory [see O. I. Tolstikhin and H. Nakamura, J. Chem. Phys. 108, 8899 (1998)]. Here we consider their role in providing a smooth transition between the known "symmetric" and "asymmetric" parametrizations, and focus on the corresponding hyperspherical harmonics. These harmonics, which will be called hyperspherical elliptic, involve products of two associated Lame polynomials. We will provide an expansion of these new sets in a finite series of standard hyperspherical harmonics, producing a powerful tool for future applications in the field of scattering and bound-state quantum-mechanical three-body problems. PMID- 15268575 TI - Trajectory approach to dissipative quantum phase space dynamics: Application to barrier scattering. AB - The Caldeira-Leggett master equation, expressed in Lindblad form, has been used in the numerical study of the effect of a thermal environment on the dynamics of the scattering of a wave packet from a repulsive Eckart barrier. The dynamics are studied in terms of phase space trajectories associated with the distribution function, W(q,p,t). The equations of motion for the trajectories include quantum terms that introduce nonlocality into the motion, which imply that an ensemble of correlated trajectories needs to be propagated. However, use of the derivative propagation method (DPM) allows each trajectory to be propagated individually. This is achieved by deriving equations of motion for the partial derivatives of W(q,p,t) that appear in the master equation. The effects of dissipation on the trajectories are studied and results are shown for the transmission probability. On short time scales, decoherence is demonstrated by a swelling of trajectories into momentum space. For a nondissipative system, a comparison is made of the DPM with the "exact" transmission probability calculated from a fixed grid calculation. PMID- 15268574 TI - A new perspective on the coarse-grained dynamics of fluids. AB - A computational methodology is presented that is designed to model, at a coarse grained level, the mesoscale dynamics of fluids and potentially other forms of soft matter. Within a molecular dynamics simulation, "ghost" particles of a specific size, corresponding to the fundamental length-scale of coarse-graining, are used as micro-probes designed to respond to local mesoscale fluid flows and stress gradients. A subsequent coarse-grained model is then developed that incorporates both the coarse-grained mesoscale dynamics and isothermal compressibility of the original microscopic system. The method is applied to water and methanol. A contrast with dissipative particle dynamics (DPD) is also presented. PMID- 15268576 TI - The one-particle Green's function method in the Dirac-Hartree-Fock framework. I. Second-order valence ionization energies of Ne through Xe. AB - The one-particle Green's function theory in its various implementations is a well established many-body approach for the calculation of electron ionization and attachment energies in atoms and molecules. In order to describe not only scalar relativistic effects but also spin-orbit splitting on an equal footing an embedding of this theory in the four-component framework was carried out and fully relativistic ionization energies of the noble gas atoms Ne through Xe were calculated using the second-order algebraic diagrammatic construction [ADC2] approximation scheme. Comparison with nonrelativistic ADC2 results and experimental data was made. PMID- 15268577 TI - Semiclassical tunneling splittings from short time dynamics: Herman-Kluk propagation and harmonic inversion. AB - We investigate a recently proposed method [J. Chem. Phys. 108, 9206 (1998)] to obtain tunneling splittings from short time cross-correlation matrices that were propagated according to the semiclassical propagator of Herman and Kluk. The energy levels were extracted by harmonic inversion of the cross-correlation matrix using the filter diagonalization technique. The aim of this study is twofold: First, the short time behavior of the Herman-Kluk-propagator and the meaning of using cross-correlation matrices rather than autocorrelation functions is addressed. Numerical examples are given for one- and two-dimensional model potentials. Second, the performance of the method is investigated for a system with considerable anharmonicity and coupling. Here the proton transfer in 3,7 dichlorotropolone is considered using an ab initio reaction surface Hamiltonian approach. For this example also the extension to more dimensions is critically discussed. PMID- 15268578 TI - The incomplete beta function law for parallel tempering sampling of classical canonical systems. AB - We show that the acceptance probability for swaps in the parallel tempering Monte Carlo method for classical canonical systems is given by a universal function that depends on the average statistical fluctuations of the potential and on the ratio of the temperatures. The law, called the incomplete beta function law, is valid in the limit that the two temperatures involved in swaps are close to one another. An empirical version of the law, which involves the heat capacity of the system, is developed and tested on a Lennard-Jones cluster. We argue that the best initial guess for the distribution of intermediate temperatures for parallel tempering is a geometric progression and we also propose a technique for the computation of optimal temperature schedules. Finally, we demonstrate that the swap efficiency of the parallel tempering method for condensed-phase systems decreases naturally to zero at least as fast as the inverse square root of the dimensionality of the physical system. PMID- 15268579 TI - W3 theory: robust computational thermochemistry in the kJ/mol accuracy range. AB - We are proposing a new computational thermochemistry protocol denoted W3 theory, as a successor to W1 and W2 theory proposed earlier [Martin and De Oliveira, J. Chem. Phys. 111, 1843 (1999)]. The new method is both more accurate overall (error statistics for total atomization energies approximately cut in half) and more robust (particularly towards systems exhibiting significant nondynamical correlation) than W2 theory. The cardinal improvement rests in an approximate account for post-CCSD(T) correlation effects. Iterative T3 (connected triple excitations) effects exhibit a basis set convergence behavior similar to the T3 contribution overall. They almost universally decrease molecular binding energies. Their inclusion in isolation yields less accurate results than CCSD(T) nearly across the board: It is only when T4 (connected quadruple excitations) effects are included that superior performance is achieved. T4 effects systematically increase molecular binding energies. Their basis set convergence is quite rapid, and even CCSDTQ/cc-pVDZ scaled by an empirical factor of 1.2532 will yield a quite passable quadruples contribution. The effect of still higher order excitations was gauged for a subset of molecules (notably the eight-valence electron systems): T5 (connected quintuple excitations) contributions reach 0.3 kcal/mol for the pathologically multireference X 1Sigmag+ state of C2 but are quite small for other systems. A variety of avenues for achieving accuracy beyond that of W3 theory were explored, to no significant avail. W3 thus appears to represent a good compromise between accuracy and computational cost for those seeking a robust method for computational thermochemistry in the kJ/mol accuracy range on small systems. PMID- 15268580 TI - On the zirconium oxide neutral cluster distribution in the gas phase: detection through 118 nm single photon, and 193 and 355 nm multiphoton, ionization. AB - Zirconium oxide clusters are generated in the gas phase by laser ablation of the metal into a flow of ca. 5% O2/95% He at 100 psig and supersonic expansion into a vacuum chamber. Mass spectra of neutral gas phase zirconium oxide clusters are obtained through photoionization at three different laser wavelengths: 118, 193, and 355 nm. Ionization of the clusters with 118 nm laser radiation is through a single photon ionization mechanism, while ionization by 193 and 355 nm laser radiation is through a multiphoton (three or more photon) mechanism. Fragment ion features are observed in the mass spectra of ZrmOn+ for only the 193 nm and 355 nm ionization schemes. The true neutral ZrmOn cluster distribution is obtained only through 118 nm single photon ionization, as verified by mass spectral peak linewidths and calculations of the cluster binding energies, ionization energies, and fragmentation rates. The neutral cluster distribution consists mainly of the series ZrmO2m and ZrmO(2m+1) for m = 1,..., approximately 30. PMID- 15268581 TI - On the iron oxide neutral cluster distribution in the gas phase. I. Detection through 193 nm multiphoton ionization. AB - Iron oxide (FemOn) neutral clusters are generated in the gas phase through laser ablation of the metal and reaction with various concentrations of O2 in He. The mixture of expansion gas and neutral FemOn cluster species is expanded through a supersonic nozzle into a vacuum system, in which the clusters are ionized by an ArF excimer laser at 193 nm, and the ions are detected and identified in a time of-flight mass spectrometer. In this report, the experimental parameters that influence the observed cluster distributions, such as ablation laser power, expansion pressure, vacuum system pressure, and 193 nm ArF ionization laser power, are explored. In the second paper in this series, the effect of the ionization laser wavelength (355 nm, 193 nm, 118 nm) on the observed cluster ion distribution is explored. The cluster ion distribution observed employing 193 nm laser ionization, is sensitive to the neutral cluster distribution as evidenced by the change in the observed time-of-flight mass spectra with changes in laser power, growth conditions, and expansion conditions. The thermodynamically stable neutral clusters for saturated O2 growth conditions are suggested to be of the forms FemOm, FemO(m+1), and FemO(m+2); which one of these series of neutral clusters is most stable depends on the size of the cluster. For m < 10, FemOm is the most stable neutral cluster series, for 10 < or = m < or = 20, FemO(m+1) is the most stable neutral cluster series, and for 21 < or = m < = 30, FemO(m+2) is the most stable neutral cluster series. Some neutral cluster fragmentation is clearly present for 193 nm ionization due to multiphoton absorption in both the neutral and ionic cluster species. PMID- 15268582 TI - On the iron oxide neutral cluster distribution in the gas phase. II. Detection through 118 nm single photon ionization. AB - Neutral clusters of iron oxide are created by laser ablation of iron metal and subsequent reaction of the gas phase metal atoms, ions, clusters, etc., with an O2/He mixture. The FemOn clusters are cooled in a supersonic expansion and detected and identified in a time-of-flight mass spectrometer following laser ionization at 118 nm (10.5 eV), 193 nm (6.4 eV), or 355 nm (3.53 eV) photons. With 118 nm radiation, the neutral clusters do not fragment because single photon absorption is sufficient to ionize all the clusters and the energy/pulse is approximately 1 microJ. Comparison of the mass spectra obtained at 118 nm ionization (single photon) with those obtained at 193 nm and 355 nm ionization (through multiphoton processes), with regard to intensities and linewidths, leads to an understanding of the multiphoton neutral cluster fragmentation pathways. The multiphoton fragmentation mechanism for neutral iron oxide clusters during the ionization process that seems most consistent with all the data is the loss of one or two oxygen atoms. In all instances of ionization by laser photons, the most intense features are of the forms FemOm+, FemO(m+1)+, and FemO(m+2)+, and this strongly suggests that, for a given m, the most prevalent neutral clusters are of the forms FemOm, FemO(m+1), and FemO(m+2). As the value of m increases, the more oxygen rich neutral clusters appear to increase in stability. PMID- 15268583 TI - On the copper oxide neutral cluster distribution in the gas phase: detection through 355 nm and 193 nm multiphoton and 118 nm single photon ionization. AB - The distribution of neutral copper oxide clusters in the gas phase created by laser ablation is detected and characterized through time-of-flight mass spectroscopy (TOFMS). The neutral copper oxide clusters are ionized by two different approaches: Multiphoton absorption of 355 and 193 nm radiation; and single photon absorption of 118 nm radiation. Based on the observed cluster patterns as a function of experimental conditions (e.g., copper oxide or metal sample, ablation laser power, expansion gas, etc.) and on the width of the TOFMS features, one can uncover the true neutral cluster distribution of CumOn species following laser ablation of the sample. Ablation of a metal sample generates only small neutral CumOn clusters for m less, similar 4 and n approximately 1, 2. Ablation of copper oxide samples generates neutral clusters of the form CumOm (m < or = 4) and CumO(m-1) (m > 4). These clusters are directly detected without fragmentation using single photon, photoionization with 118 nm laser radiation. Using 355 and 193 nm multiphoton ionization, the observed cluster ions are mostly of the form Cu2mOm+ for 4 < or = m < or = 10 (193 nm ionization) and CumO1,2 (355 nm ionization) for copper oxide samples. Neutral cluster fragmentation due to multiphoton processes seems mainly to be of the form CumO(m,m-1) --> CumO(m/2,m/2+1). Neutral cluster growth mechanisms are discussed based on the cluster yield from different samples (e.g., Cu metal, CuO powder, and Cu2O powder). PMID- 15268584 TI - Low-energy electron scattering by cubane: resonant states and Ramsauer-Townsend features from quantum calculations in the gas phase. AB - Calculations are carried out, using a nonempirical modeling of the interaction potential and solving the quantum scattering coupled channel equations, for low energy electron scattering from cubane (C8H8) molecules in the gas phase. Total integral cross sections are obtained and partial contributions are analyzed for the most important irreducible representations that describe the continuum electron in the Oh molecular symmetry. Several trapping resonances are found and analyzed in terms of the molecular-type features of the resonant electron states associated with them. A Ramsauer-Townsend minimum is also found and its possible behavior related to features of the scattering length as k --> 0. PMID- 15268585 TI - Neutral and zwitterionic glycine.H(2)O complexes: A theoretical and matrix isolation Fourier transform infrared study. AB - The H-bond interaction between glycine and H2O has been studied by a combined theoretical (DFT(B3LYP)/6-31++G(**)) and experimental (matrix-isolation FT-IR) methodology. The 1:1 and 1:2 complexes of the most stable conformation (I) of glycine appear to be neutral complexes which have been vibrationally characterized in detail. The higher stoichiometry complexes (glycine).(H2O)n with n larger than 3 are demonstrated to be zwitterionic H-bonded complexes. A set of characteristic IR absorption bands for this zwitterionic structure has been observed in low-temperature Ar matrices. This would be the first experimental IR evidence for proton transfer occurring between the NH2 and COOH groups of amino acids by a H-bonded water molecular channel in isolated conditions. PMID- 15268586 TI - Spectroscopic interpretation: the high vibrations of CDBrClF. AB - We extract the dynamics implicit in an algebraic fitted model Hamiltonian for the deuterium chromophore's vibrational motion in the molecule CDBrClF. The original model has four degrees of freedom, three positions and one representing interbond couplings. A conserved polyad allows in a semiclassical approach the reduction to three degrees of freedom. For most quantum states we can identify the underlying motion that when quantized gives the said state. Most of the classifications, identifications, and assignments are done by visual inspection of the already available wave function semiclassically transformed from the number representation to a representation on the reduced dimension toroidal configuration space corresponding to the classical action and angle variables. The concentration of the wave function density to lower dimensional subsets centered on idealized simple lower dimensional organizing structures and the behavior of the phase along such organizing centers already reveals the atomic motion. Extremely little computational work is needed. PMID- 15268587 TI - Electronic ground states of the V2O4+/0/- species from multireference correlation and density functional studies. AB - The molecular and electronic structures of the V2O4+/0/- species are examined by multireference averaged coupled-pair functional (MR-ACPF) and density functional B3LYP calculations. For all three species, new conformers have been found. Shallow potential energy curves imply high mobility of the oxygen atoms in the neutral and anionic species for which antiferromagnetic coupling of the weakly interacting 3dV electrons is found. Good agreement between the MR-ACPF and B3LYP results for the molecular structures and the relative energies of states with different spin multiplicity, as well as for the ionization energy and electron affinity, is observed. For the computation of the height of the transition barriers between different conformers elaborated MR-ACPF calculations are required. PMID- 15268588 TI - Ab initio configuration interaction study of the low-lying 1Sigma+ electronic states of LiCl. AB - Ab initio configuration interaction calculations have been performed for the X 1Sigma+ and B 1Sigma+ electronic states of LiCl. Potential energy curves, dipole moment functions, and dipole transition moments have been computed for internuclear distances between R = 2.5a0 and 50a0. Single- and double-excitation configuration interaction wave functions were constructed using molecular orbitals obtained from a two-state averaged multiconfiguration self-consistent field calculation. This procedure yielded an accurate energy splitting between the covalent and ionic separated-atom limits. The calculated avoided crossing of the X and B state curves occurs at R = 16.2a0, in close agreement with previous calculations using a semiempirical covalent-ionic resonance model. X 1Sigma+ state spectroscopic constants are in excellent agreement with experimental values. PMID- 15268589 TI - 193-nm photodissociation of acryloyl chloride to probe the unimolecular dissociation of CH2CHCO radicals and CH2CCO. AB - The work presented here uses photofragment translational spectroscopy to investigate the primary and secondary dissociation channels of acryloyl chloride (CH2==CHCOCl) excited at 193 nm. Three primary channels were observed. Two C-Cl fission channels occur, one producing fragments with high kinetic recoil energies and the other producing fragments with low translational energies. These channels produced nascent CH2CHCO radicals with internal energies ranging from 23 to 66 kcal/mol for the high-translational-energy channel and from 50 to 68 kcal/mol for the low-translational-energy channel. We found that all nascent CH2CHCO radicals were unstable to CH2CH + CO formation, in agreement with the G3//B3LYP barrier height of 22.4 kcal/mol to within experimental and computational uncertainties. The third primary channel is HCl elimination. All of the nascent CH2CCO coproducts were found to have enough internal energy to dissociate, producing CH2C: + CO, in qualitative agreement with the G3//B3LYP barrier of 39.5 kcal/mol. We derive from the experimental results an upper limit of 23 +/- 3 kcal/mol for the zero-point-corrected barrier to the unimolecular dissociation of the CH2CHCO radical to form CH2CH + CO. PMID- 15268590 TI - State-to-state dynamics of the Cl + CH3OH --> HCl + CH2OH reaction. AB - Molecular chlorine, methanol, and helium are co-expanded into a vacuum chamber using a custom designed "late-mixing" nozzle. The title reaction is initiated by photolysis of Cl2 at 355 nm, which generates monoenergetic Cl atoms that react with CH3OH at a collision energy of 1960 +/- 170 cm(-1) (0.24 +/- 0.02 eV). Rovibrational state distributions of the nascent HCl products are obtained via 2 + 1 resonance enhanced multiphoton ionization, center-of-mass scattering distributions are measured by the core-extraction technique, and the average internal energy of the CH3OH co-products is deduced by measuring the spatial anisotropy of the HCl products. The majority (84 +/- 7%) of the HCl reaction products are formed in HCl(v = 0) with an average rotational energy of [Erot] = 390 +/- 70 cm(-1). The remaining 16 +/- 7% are formed in HCl(v = 1) and have an average rotational energy of [Erot] = 190 +/- 30 cm(-1). The HCl(v = 1) products are primarily forward scattered, and they are formed in coincidence with CH2OH products that have little internal energy. In contrast, the HCl(v = 0) products are formed in coincidence with CH2OH products that have significant internal energy. These results indicate that two or more different mechanisms are responsible for the dynamics in the Cl + CH3OH reaction. We suggest that (1) the HCl(v = 1) products are formed primarily from collisions at high impact parameter via a stripping mechanism in which the CH2OH co-products act as spectators, and (2) the HCl(v = 0) products are formed from collisions over a wide range of impact parameters, resulting in both a stripping mechanism and a rebound mechanism in which the CH2OH co-products are active participants. In all cases, the reaction of fast Cl atoms with CH3OH is with the hydrogen atoms on the methyl group, not the hydrogen on the hydroxyl group. PMID- 15268591 TI - Ab initio study of the reactions of Ga(2P, 2S, and 2P) with methane. AB - The interactions of Ga(2P:4s(2)4p1, 2S:4s(2)5s1, and 2P:4s(2)5p1) with CH4 is studied by means of Hartree-Fock self-consistent-field (SCF) calculations using relativistic effective core potentials and multiconfigurational-SCF plus multireference variational and perturbational on second-order Moller-Plesset configuration interaction calculations. The Ga atom 2P(4s(2)5p1) state can spontaneously insert into the CH4. In this interaction the 4 2A potential energy surface is initially attractive and becomes repulsive only after meeting with the 3 2A surface, adiabatically linked with the Ga(2S:4s(2)5s1) + CH4 fragments. The Ga atom 2S(4s(2)5s1) excited state inserts in the C-H bond. In this interaction the 3 2A potential energy surface initially attractive, becomes repulsive after meet the 2 2A' surface linked with the Ga(2P:4s(2)4p1) + CH4 fragments. The two 2A curves (2 2A and X 2A) derived from the interaction of Ga(2P:4s(2)4p1) atoms with methane molecules are initially repulsive. The 2 2A curve after an avoided crossing with the 3 2A curve goes smoothly down and reaches a minimum: after this point, it shows an energy barrier. The top of this barrier is located below the energy value of the Ga(2S:4s(2)5s1) + CH4 fragments. After this energy top the 2 2A curve goes down to meet the X 2A curve. The 2 2A curve becomes repulsive after the avoided crossing with the X 2A curve. The X 2A curve becomes attractive only after its avoided crossing with the 2 2A curve. The lowest-lying X 2A potential leads to the HGaCH3 X 2A intermediate molecule. This intermediate molecule, diabatically correlated with the Ga(2S:4s(2)5s1) + CH4 fragments, which lie 6 kcal/mol, above the ground-state reactants, the dissociation channels of this intermediate molecule leading to the GaH + CH3 and H + GaCH3 products. These products are reached from the HGaCH3 intermediate without activation barriers. The work results suggest that Ga atom in the first excited state in gas-phase methane molecules could produce better quality a-C:H thin films through CH3 radicals, as well as gallium carbide materials. PMID- 15268592 TI - The microwave and infrared spectroscopy of benzaldehyde: conflict between theory and experimental deductions. AB - Recently, it has been proposed that ab initio calculations cannot accurately treat molecules comprised of a benzene ring with a pi-conjugated substituent, for example, benzaldehyde. Theoretical predictions of the benzaldehyde barrier to internal rotation are typically a factor of 2 too high in comparison to the experimental values of 4.67 (infared) and 4.90 (microwave) kcal mol(-1). However, both experiments use Pitzer's 1946 model to compute the reduced moment of inertia and employ the experimentally observed torsional frequency to deduce benzaldehyde's rotational barrier. When Pitzer's model is applied to a system with a nonconjugated functional group, such as phenol, the model and theoretical values are in close agreement. Therefore, we conclude the model may not account for conjugation between the substituent and the pi-system of benzene. The experimental values of the benzaldehyde rotational barrier are therefore misleading. The true rotational barrier lies closer to the theoretically extrapolated limit of 7.7 kcal mol(-1), based on coupled cluster theory. PMID- 15268593 TI - A density-functional study of Al-doped Ti clusters: TinAl (n = 1-13). AB - Equilibrium geometries, stabilities, and electronic properties of TinAl (n = 1 13) clusters have been studied by using density-functional theory with local spin density approximation and generalized gradient approximation. The ground-state structures of TinAl clusters have been obtained. The resulting geometries show that the aluminum atom remains on the surface of clusters for n < 9, but is slowly getting trapped beyond n = 9, meanwhile, the Al atom exhibits a valent transition from monovalent to trivalent. The geometric effects and electronic effects clearly demonstrate the Ti4Al cluster to be endowed with special stability. The studies on the bonds indicate the change from ionic to metalliclike. PMID- 15268594 TI - Persistence rewarded: successful observation of the B 2Sigmau+-X 2Pig electronic transition of jet-cooled BS2. AB - The B-X electronic transition of jet-cooled BS2 has been observed using laser induced fluorescence techniques. The boron disulfide radical was produced in a pulsed electric discharge jet using a mixture of BCl3 and CS2 in high-pressure argon as the precursor. The spectrum consists of a strong 0(0)(0) band with the 2Sigma-2Pi(3/2) component at 24,393.2 cm(-1) and short progressions in the symmetric stretching (nu1' = 506.7 cm(-1)) and bending (nu2' = 303.2 cm(-1)) modes. A rotational analysis of both spin-orbit components of the 0(0)(0) band gave an upper state B value of 0.0932779(19) cm(-1) and a ground-state spin-orbit coupling constant of A = -405.163(4) cm(-1). The ground-state bond length of 1.66492 angstroms increases to 1.6812(1) angstroms on sigmau --> pig electronic excitation. The B-X data have been used to further refine the Renner-Teller analysis, which is in good agreement with our previous work [J. Chem. Phys. 119, 2047 (2003)]. PMID- 15268595 TI - New results for the OH (nu = 0,j = 0) + CO (nu = 0,j = 0) --> H + CO2 reaction: Five- and full-dimensional quantum dynamical study on several potential energy surfaces. AB - Full- [six-dimensional (6-D)] and reduced-dimensional [five-dimensional (5-D)] quantum wave packet calculations have been performed for the title reaction to obtain reaction probabilities deriving from the ground rovibrational states of OH and CO with total angular momentum J = 0. Three potential energy surfaces (PES) are studied, namely, those of Bradley and Schatz (BS), Yu, Muckerman, and Sears (YMS), and Lakin, Troya, Schatz, and Harding (LTSH). 6-D calculations are performed only for the BS PES, while 5-D results are reported for all three PES'. The 6-D results obtained in the present work improve on those previously reported, since a larger vibrational basis and a better representation of the OH and CO bonds has been introduced. In particular, we now employ a generalized Lanczos-Morse discrete variable representation for both the OH and CO vibrations. In a further improvement, the generalized discrete variable representation of the CO vibration is based on different CO intramolecular potentials for the asymptotic and product grids employed in our projection formalism. This new treatment of the vibrational bases allows for a large reduction in computation time with respect to our previous implementation of the wave packet method, for a given level of accuracy. As a result, we have been able to extend the range of collision energies for which we can obtain converged 6-D results to a higher energy (0.8 eV) than was possible before (0.5 eV). The comparison of the new 6-D and previous 5-D results for the BS PES shows good agreement of the general trend in the reaction probabilities over all collision energies considered (0.1-0.8 eV), while our previous 6-D calculation showed reaction probabilities that differed from the 5-D results by up to 10% between 0.5 and 0.8 eV. The 5-D reaction probabilities reveal interesting trends for the different PES'. In particular, at low energies (< 0.2 eV) the LTSH PES gives rise to much larger reactivity than the other PES', while at high energies (> 0.3 eV) its reaction probability decreases with respect to the BS and YMS PES', being more than a factor of 2 smaller at 0.8 eV. A 5-D calculation on a modified version of the LTSH surface shows that the van der Waals interaction in the entrance channel, which is not correctly described in the other PES' is largely responsible for its larger reactivity at low energies. The large difference between the 5-D reaction probabilities for the YMS and LTSH PES' serves to emphasize the importance of the van der Waals interaction for the reactivity at low energies, because most of the stationary point energies on the YMS and LTSH PES are rather similar, being in line with high-level ab initio information. PMID- 15268596 TI - An ab initio potential energy surface and predissociative resonances of HArF. AB - A three-dimensional potential energy surface of the ground electronic state HArF is constructed from more than 2000 ab initio points at the multireference averaged quadratic coupled-cluster level employing an augmented large basis set. The calculations indicate that the linear HArF molecule is metastable with a barrier of 0.643 eV in the atomization (HArF --> H + Ar + F) channel and a barrier of 1.017 eV in the dissociation (HArF --> Ar + HF) channel. Variational calculations of low-lying predissociative resonances of both HArF and DArF are performed on the three-dimensional potential energy surface using a complex symmetric Lanczos propagation method, which yields both positions and widths of the resonance states. The resonance lifetime generally decreases with energy, but strong mode selectivity exists. Reasonably good agreement with experiment confirms the accuracy of our potential. These calculations provide valuable information on the stability and dynamics of HArF/DArF in its ground electronic state. PMID- 15268597 TI - Energy distributions in multiple photon absorption experiments. AB - Photofragmentation experiments on molecules and clusters often involve multiple photon absorption. The distributions of the absorbed number of photons are frequently approximated by Poisson distributions. For realistic laser beam profiles, this approximation fails seriously due to the spatial variation of the mean number of absorbed photons across the laser beam. We calculate the distribution of absorbed energy for various laser and molecular-beam parameters. For a Gaussian laser beam, the spatially averaged distributions have a power-law behavior for low energy with a cutoff at an energy which is proportional to fluence. The power varies between -1 for an almost parallel laser beam and -5/2 for a divergent beam (on the scale of the molecular beam). We show that the experimental abundance spectra of fullerenes and small carbon clusters can be used to reconstruct the distribution of internal energy in the excited C60 molecule prior to fragmentation and find good agreement with the calculated curves. PMID- 15268598 TI - An ab initio study of the ground and valence excited states of GaF. AB - Ab initio calculations on the ground and valence excited states of the GaF molecule have been performed by using the internally contracted multireference electronic correlation methods (MR-CISD, MR-CISD + Q, and MR-AQCC) with entirely uncontracted all-electronic basis sets and Douglas-Kroll scalar relativistic correction. The potential energy curves of all valence states and the spectroscopic constants of bound states are fitted. It is the first time that the 12 valence Lambda-S states of GaF molecule and all of the 23 Omega states generated from the former are studied in a theoretical way. Calculation results well reproduce most of the experimental data. The effects of the size-extensivity correction and the avoided crossing rule between Omega states of the same symmetry are analyzed. The transition properties of the A 3Pi0+, B 3Pi1, C 1Pi1, and 3Sigma1+ states are predicted, including the transition dipole moments, the Franck-Condon factors and the radiative lifetimes. The radiative lifetime of the C 1Pi1 state of GaF molecule is of the order of nanosecond, implying that it is a rather short-live state. The lifetimes of the B 3Pi1 and 3Sigma1+ states are of the order of microsecond, while the lifetime of the A 3Pi0+ state are the order of millisecond. PMID- 15268599 TI - Infrared laser spectroscopy of jet cooled cobalt tricarbonyl nitrosyl. AB - Rotationally resolved infrared absorption spectra for the 1(0)(1) band of jet cooled cobalt tricarbonyl nitrosyl have been observed and analyzed. Several longitudinal modes of a Pb-salt diode laser were utilized to measure 105 rovibrational transitions for this particular vibrational band centered near 2112 cm(-1). Spectra were optimized using both argon and helium carrier gases and these experiments eventually led to rovibrational transitions being assigned to four different K subbands, specifically the K = 0, 3, 6, and 9 subbands. An iterative least-squares analysis of the spectroscopic data yielded the following molecular parameters nu0 = 2111.7457(9) cm(-1), B0 = 0.034747(12) cm(-1), B1 = 0.034695(15) cm(-1), C1 = 0.03380(9) cm(-1), and D1K = 6.3(9) x 10(-6) cm(-1) (where 3sigma uncertainties are listed in parenthesis). PMID- 15268600 TI - Experimental and theoretical study of proton spin-lattice relaxation in H2-Ar gas mixtures: critical examination of the XC(fit) potential energy surface. AB - Proton nuclear magnetic resonance spin-lattice relaxation time measurements have been carried out at 500 MHz proton Larmor frequency on two hydrogen-argon gas mixtures with 1.90% and 3.93% hydrogen at four different temperatures in the range 225 K < T < 337 K and at two different number densities. The results for different hydrogen mole percentages have been extrapolated to infinite dilution to obtain the contributions to the overall relaxation times arising from the hydrogen-argon interaction. The extrapolated relaxation times fall in the reciprocal regime in which relaxation times are inversely proportional to the density. Relaxation times have also been calculated using quantum mechanical close-coupled computations based on the H2-Ar XC(fit) potential energy surface obtained by Bissonnette et al. [J. Chem. Phys. 105, 2639 (1996)]. Significant differences found between the experimental and theoretical results indicate that the short-range anisotropy of the XC(fit) potential surface is too weak. The reciprocal regime is shown to have a much higher sensitivity to changes in the anisotropic component of the intermolecular potential energy surface. PMID- 15268601 TI - Quantum and classical studies of the O(3P) + H2(v = 0-3,j = 0) --> OH + H reaction using benchmark potential surfaces. AB - We present results of time-dependent quantum mechanics (TDQM) and quasiclassical trajectory (QCT) studies of the excitation function for O(3P) + H2(v = 0-3,j = 0) --> OH + H from threshold to 30 kcal/mol collision energy using benchmark potential energy surfaces [Rogers et al., J. Phys. Chem. A 104, 2308 (2000)]. For H2(v = 0) there is excellent agreement between quantum and classical results. The TDQM results show that the reactive threshold drops from 10 kcal/mol for v = 0 to 6 for v = 1, 5 for v = 2 and 4 for v = 3, suggesting a much slower increase in rate constant with vibrational excitation above v = 1 than below. For H2(v > 0), the classical results are larger than the quantum results by a factor approximately 2 near threshold, but the agreement monotonically improves until they are within approximately 10% near 30 kcal/mol collision energy. We believe these differences arise from stronger vibrational adiabaticity in the quantum dynamics, an effect examined before for this system at lower energies. We have also computed QCT OH(v',j') state-resolved cross sections and angular distributions. The QCT state-resolved OH(v') cross sections peak at the same vibrational quantum number as the H2 reagent. The OH rotational distributions are also quite hot and tend to cluster around high rotational quantum numbers. However, the dynamics seem to dictate a cutoff in the energy going into OH rotation indicating an angular momentum constraint. The state-resolved OH distributions were fit to probability functions based on conventional information theory extended to include an energy gap law for product vibrations. PMID- 15268602 TI - Microsolvation of N2H): the nature of interactions in N2H+-(H2)n (n = 1-14) complexes. AB - Experimental studies of the consecutive growth of N2H + (H2)n clusters led to the discovery of an unusual bonding pattern for species with n = 2-4. Theoretical studies revealed that the ligands are located within five well-separated solvation shells that are visible in structures, values of successive enthalpies and entropies of clustering reactions, vibrational motions, the distribution of atomic charges, and interaction energy decomposition components. The pattern of consecutive enthalpy changes for the second shell (n = 2-5) is complicated. This pattern shows anomalous behavior, although its interpretation is not univocal. A large part of consecutive enthalpies for the clustering reactions is a contribution due to the rotational and vibrational properties of clusters which are difficult for adequate modeling in large systems. The structures of clusters are rationalized based on interaction energy contributions of a different nature. Geometries of complexes are determined by prevailing covalent forces. PMID- 15268603 TI - Investigating bonding in small silicon-carbon clusters: exploration of the potential energy surfaces of Si3C4, Si4C3, and Si4C4 using ab initio molecular dynamics. AB - A theoretical investigation of the properties of the Si3C4, Si4C3, and Si4C4 clusters is reported. Systematic explorations of the potential energy surfaces of the three clusters are performed using a combination of ab initio molecular dynamics and local energy minimizations using density functional theory. A large number of isomers with a large variety of geometries has been found. The geometries, energies, and vibrational frequencies yielded are discussed. Furthermore, a quantitative analysis of the interatomic distances, angles, and coordination numbers observed, as well as the conclusions on the bonding properties, are presented. The cluster properties are then compared to those of solid SiC and of the smaller Si-C clusters (with size up to 6) obtained in a previous study. Analysis of our results and comparison with bulk properties show that even clusters as small as Si3C4, Si4C3, and Si4C4 exhibit properties similar to those of the amorphous bulk, in particular as for the structures and bonds formed by C atoms. PMID- 15268605 TI - Electric conductivities of 1:1 electrolytes in liquid methanol along the liquid vapor coexistence curve up to the critical temperature. I. NaCl, KCl, and CsCl solutions. AB - The molar conductivities Lambda of NaCl, KCl, and CsCl in liquid methanol were measured in the concentration range of (0.3-2.0) x 10(-3) mol dm(-3) and the temperature range of 60-240 degrees C along the liquid-vapor coexistence curve. The temperature range corresponds to the solvent density range of (2.78 1.55)rhoc, where rhoc = 0.2756 g cm(-3) is the critical density of methanol. The concentration dependence of Lambda at each temperature and density (pressure) has been analyzed by the Fuoss-Chen-Justice equation to obtain the limiting molar conductivity Lambda0 and the molar association constant KA. For all the electrolytes studied, Lambda0 increased almost linearly with decreasing density at densities above 2.0rhoc, while the opposite tendency was observed at lower densities. The relative contribution of the nonhydrodynamic effect on the translational friction coefficient zeta was estimated in terms of Deltazeta/zeta, where the residual friction coefficient Deltazeta is the difference between zeta and the Stokes friction coefficient zetaS. At densities above 2.0rhoc, Deltazeta/zeta increased with decreasing density though zeta and Deltazeta decrease, and the tendencies are common for all the ions studied. The density dependences of zeta and Deltazeta/zeta were explained well by the Hubbard-Onsager (HO) dielectric friction theory based on the sphere-in-continuum model. At densities below 2.0rhoc, however, the experimental results cannot be explained by the HO theory. PMID- 15268604 TI - Inertial solvent dynamics and the analysis of spectral line shapes: Temperature dependent absorption spectrum of beta-carotene in nonpolar solvent. AB - The influence of solvent dynamics on optical spectra is often described by a stochastic model which assumes exponential relaxation of the time-correlation function for solvent-induced frequency fluctuations. In contrast, theory and experiment suggest that the initial (subpicosecond) phase of solvent relaxation, resulting from inertial motion of the solvent, is a Gaussian function of time. In this work, we employ numerical and analytical calculations to compare the predicted absorption line shapes and the derived solvent reorganization energies obtained from exponential (Brownian oscillator) versus Gaussian (inertial) solvent dynamics. Both models predict motional narrowing as the ratio kappa = Lambda/Delta is increased, where Lambda and Delta are the frequency and variance, respectively, of the solvent-induced frequency fluctuations. However, the motional narrowing limit is achieved at lower values of kappa for the Brownian oscillator model compared to the inertial model. For a given line shape, the derived value of the solvent reorganization energy lambdasolv is only weakly dependent on the solvent relaxation model employed, though different solvent parameters Lambda and Delta are obtained. The two models are applied to the analysis of the temperature-dependent absorption spectrum of beta-carotene in isopentane and CS2. The derived values of lambdasolv using the Gaussian model are found to be in better agreement with the high temperature limit of Delta2/2kBT than are the values obtained using the Brownian oscillator model. In either approach, the solvent reorganization energy is found to increase slightly with temperature as a result of an increase in the variance Delta of the solvent induced frequency fluctuations. PMID- 15268606 TI - Investigation of vapor-deposited amorphous ice and irradiated ice by molecular dynamics simulation. AB - With the purpose of clarifying a number of points raised in the experimental literature, we investigate by molecular dynamics simulation the thermodynamics, the structure and the vibrational properties of vapor-deposited amorphous ice (ASW) as well as the phase transformations experienced by crystalline and vitreous ice under ion bombardment. Concerning ASW, we have shown that by changing the conditions of the deposition process, it is possible to form either a nonmicroporous amorphous deposit whose density (approximately 1.0 g/cm3) is essentially invariant with the temperature of deposition, or a microporous sample whose density varies drastically upon temperature annealing. We find that ASW is energetically different from glassy water except at the glass transition temperature and above. Moreover, the molecular dynamics simulation shows no evidence for the formation of a high-density phase when depositing water molecules at very low temperature. In order to model the processing of interstellar ices by cosmic ray protons and heavy ions coming from the magnetospheric radiation environment around the giant planets, we bombarded samples of vitreous ice and cubic ice with 35 eV water molecules. After irradiation the recovered samples were found to be densified, the lower the temperature, the higher the density of the recovered sample. The analysis of the structure and vibrational properties of this new high-density phase of amorphous ice shows a close relationship with those of high-density amorphous ice obtained by pressure-induced amorphization. PMID- 15268607 TI - Amide I vibrational circular dichroism of dipeptide: Conformation dependence and fragment analysis. AB - The amide I vibrational circular dichroic response of alanine dipeptide analog (ADA) was theoretically investigated and the density functional theory calculation and fragment analysis results are presented. A variety of vibrational spectroscopic properties, local and normal mode frequencies, coupling constant, dipole, and rotational strengths, are calculated by varying two dihedral angles determining the three-dimensional ADA conformation. Considering two monopeptide fragments separately, we show that the amide I vibrational circular dichroism of the ADA can be quantitatively predicted. For several representative conformations of the model ADA, vibrational circular dichroism spectra are calculated by using both the density functional theory calculation and fragment analysis methods. PMID- 15268608 TI - Angular resolution and range of dipole-dipole correlations in water. AB - We investigate the dipolar correlations in liquid water at angular resolution by molecular-dynamics simulations of a large periodic simulation system containing about 40,000 molecules. Because we are particularly interested in the long-range ordering, we use a simple three-point model for these molecules. The electrostatics is treated both by Ewald summation and by minimum image truncation combined with a reaction field approach. To gain insight into the angular dependence of the simulated dipolar ordering we introduce a suitable expansion of the molecular pair distribution function into a set of two-dimensional correlation functions. We show that these functions enable detailed insights into the shell structure of the dipolar ordering around a given water molecule. For these functions we derive analytical expressions in the particular case in which liquid water is conceived as a dielectric continuum. Comparisons of these continuum models with the correlation functions derived from the simulations yield the key result that liquid water behaves like a continuum dielectric beyond distances of about 15 angstroms from a given water molecule. We argue that this should be a generic property of water independent of our modeling. By comparison of the results of the two different electrostatics treatments with the continuum description we show that the boundary artifacts occurring in both methods are isotropically distributed and are locally small in the respective boundary regions. PMID- 15268609 TI - Particle rearrangements during transitions between local minima of the potential energy landscape of a binary Lennard-Jones liquid. AB - The potential energy landscape (PEL) of binary Lennard-Jones (BLJ) mixtures exhibits local minima, or inherent structures (IS), which are organized into metabasins (MBs). We study the particle rearrangements related to transitions between both successive IS and successive MB for a small 80:20 BLJ system near the mode-coupling temperature TMCT. The analysis includes the displacements of individual particles, the localization of the rearrangements, and the relevance of string-like motion. We find that the particle rearrangements during IS and MB transitions do not change significantly at TMCT. In particular, an onset of single particle hopping on the length scale of the interparticle distance is not observed. Further, it is demonstrated that IS and MB dynamics are spatially heterogeneous and facilitated by string-like motion. To investigate the mechanism of string-like motion, we follow the particle rearrangements during suitable sequences of IS transitions. We find that most strings observed after a series of transitions do not move coherently during a single transition, but subunits of different sizes are active at different times. Several findings suggest that, though string-like motion is of comparable relevance when the system explores a MB and when it moves from one MB to another, the occurrence of a successful string enables the system to exit a MB. Moreover, we show that the particle rearrangements during two consecutive MB transitions are basically uncorrelated. In particular, different groups of particles are highly mobile. We further find the positions of strings during successive MB transitions weakly but positively correlated, supporting the idea of dynamic facilitation. Finally, the relation between the features of the potential energy landscape and the relaxation processes in supercooled liquids is discussed. PMID- 15268610 TI - Particle dynamics and the development of string-like motion in a simulated monoatomic supercooled liquid. AB - The microscopic details of local particle dynamics is studied in a glass-forming one component supercooled liquid modeled by a Dzugutov potential developed for simple metallic glass formers. Our main goal is to investigate particle motion in the supercooled liquid state, and to ascertain the extent to which this motion is cooperative and occurring in quasi-one-dimesional, string-like paths. To this end we investigate in detail the mechanism by which particles move along these paths. In particular, we show that the degree of coherence--that is, simultaneous motion by consecutive particles along a string--depends on the length of the string. For short strings, the motion is highly coherent. For longer strings, the motion is highly coherent only within shorter segments of the string, which we call "microstrings." Very large strings may contain several microstrings within which particles move simultaneously, but individual microstrings within a given string are temporally uncorrelated with each other. We discuss possible underlying mechanism for this complex dynamical behavior, and examine our results in the context of recent work by Garrahan and Chandler [Phys. Rev. Lett. 89, 035704 (2002)] in which dynamic facilitation plays a central role in the glass transition. PMID- 15268611 TI - Molecular cluster decay viewed as escape from a potential of mean force. AB - We show that evaporation from a quasistable molecular cluster may be treated as a kinetic problem involving the stochastically driven escape of a molecule from a potential of mean force. We derive expressions for the decay rate, and a relationship between the depth of the potential and the change in system free energy upon loss of a molecule from the cluster. This establishes a connection between kinetic and thermodynamic treatments of evaporation, but also reveals differences in the prefactor in the rate expression. We perform constant energy molecular dynamics simulations of cluster dynamics to calculate potentials of mean force, friction coefficients and effective temperatures for use in the kinetic analysis, and to compare the results with the directly observed escape rates. We also use the simulations to estimate the escape rates by a probabilistic analysis. It is much more efficient to calculate the decay rate by the methods we have developed than it is to monitor escape directly, making these approaches potentially useful for the assessment of molecular cluster stability. PMID- 15268612 TI - Bridge mediated two-electron transfer reactions: analysis of stepwise and concerted pathways. AB - A theory of nonadiabatic donor (D)-acceptor (A) two-electron transfer (TET) mediated by a single regular bridge (B) is developed. The presence of different intermediate two-electron states connecting the reactant state D-(-)BA with the product state DBA-(-) results in complex multiexponential kinetics. The conditions are discussed at which a reduction to two-exponential as well as single-exponential kinetics becomes possible. For the latter case the rate KTET is calculated, which describes the bridge-mediated reaction as an effective two electron D-A transfer. In the limit of small populations of the intermediate TET states D-B-A, DB-(-)A, D-BA-, and DB-A-, KTET is obtained as a sum of the rates KTET(step) and KTET(sup). The first rate describes stepwise TET originated by transitions of a single electron. It starts at D-(-)BA and reaches DBA-(-) via the intermediate state D-BA-. These transitions cover contributions from sequential as well as superexchange reactions all including reduced bridge states. In contrast, a specific two-electron superexchange mechanism from D-(-)BA to DBA-(-) defines KTET(sup). An analytic dependence of KTET(step) and KTET(sup) on the number of bridging units is presented and different regimes of D-A TET are studied. PMID- 15268613 TI - Size dependent ion hydration, its asymmetry, and convergence to macroscopic behavior. AB - The packing and orientation of water molecules in the vicinity of solutes strongly influence the solute hydration thermodynamics in aqueous solutions. Here we study the charge density dependent hydration of a broad range of spherical monovalent ionic solutes (with solute diameters from approximately 0.4 nm to 1.7 nm) through molecular dynamics simulations in the simple point charge model of water. Consistent with previous experimental and theoretical studies, we observe a distinct asymmetry in the structure and thermodynamics of hydration of ions. In particular, the free energy of hydration of negative ions is more favorable than that of positive ions of the same size. This asymmetry persists over the entire range of solute sizes and cannot be captured by a continuum description of the solvent. The favorable hydration of negative ions arises primarily from the asymmetric charge distribution in the water molecule itself, and is reflected in (i) a small positive electrostatic potential at the center of a neutral solute, and (ii) clear structural (packing and orientation) differences in the hydration shell of positive and negative ions. While the asymmetry arising from the positive potential can be quantified in a straightforward manner, that arising from the structural differences in the fully charged states is difficult to quantify. The structural differences are highest for the small ions and diminish with increasing ion size, converging to hydrophobiclike hydration structure for the largest ions studied here. We discuss semiempirical measures following Latimer, Pitzer, and Slansky [J. Chem. Phys. 7, 108 (1939)] that account for these structural differences through a shift in the ion radius. We find that these two contributions account completely for the asymmetry of hydration of positive and negative ions over the entire range of ion sizes studied here. We also present preliminary calculations of the dependence of ion hydration asymmetry on the choice of water model that demonstrate its sensitivity to the details of ion-water interactions. PMID- 15268614 TI - Case study of enthalpy-entropy noncompensation. AB - Enthalpy-entropy noncompensation characterizes the relative changes in the hydration thermodynamic functions upon "transforming" ethane into fluoromethane, chloromethane, bromomethane, and iodomethane. An analysis grounded on a simple statistical mechanical theory of hydration allows a plausible rationalization of such enthalpy-entropy noncompensation. It is shown that increasing the strength of solute-water attractive interactions modifying the chemical nature of a part of the solute molecule, but not its size, is a largely noncompensating process for the hydration of noncharged and nonhydrogen bonding species, and dominates the compensating contribution coming from the reorganization of water H bonds. PMID- 15268615 TI - Tractable molecular theory of transport of Lennard-Jones fluids in nanopores. AB - We present here a tractable theory of transport of simple fluids in cylindrical nanopores, which is applicable over a wide range of densities and pore sizes. In the Henry law low-density region the theory considers the trajectories of molecules oscillating between diffuse wall collisions, while at higher densities beyond this region the contribution from viscous flow becomes significant and is included through our recent approach utilizing a local average density model. The model is validated by means of equilibrium as well nonequilibrium molecular dynamics simulations of supercritical methane transport in cylindrical silica pores over a wide range of temperature, density, and pore size. The model for the Henry law region is exact and found to yield an excellent match with simulations at all conditions, including the single-file region of very small pore size where it is shown to provide the density-independent collective transport coefficient. It is also shown that in the absence of dispersive interactions the model reduces to the classical Knudsen result, but in the presence of such interactions the latter model drastically overpredicts the transport coefficient. For larger micropores beyond the single-file region the transport coefficient is reduced at high density because of intermolecular interactions and hindrance to particle crossings leading to a large decrease in surface slip that is not well represented by the model. However, for mesopores the transport coefficient increases monotonically with density, over the range studied, and is very well predicted by the theory, though at very high density the contribution from surface slip is slightly overpredicted. It is also seen that the concept of activated diffusion, commonly associated with diffusion in small pores, is fundamentally invalid for smooth pores, and the apparent activation energy is not simply related to the minimum pore potential or the adsorption energy as generally assumed. PMID- 15268616 TI - Dissociation mechanism of 2-propanol on a Si(111)-(7x7) surface studied by scanning tunneling microscopy. AB - Adsorption of 2-propanol, (CH3)2CHOH, on a Si(111)-7x7 surface was studied by scanning tunneling microscopy. (CH3)2CHOH adsorbs equally on the faulted and unfaulted half unit cells by forming Si-OCH(CH3)2 and Si-H on an adatom and rest atom pair. Si-OCH(CH3)2 is consecutively increased in each half unit cell, and the adsorption is saturated when every half unit cell has three Si-OCH(CH3)2, which corresponds to 0.5 of the adatom coverage. The sticking probability for the dissociation of (CH3)2CHOH is independent of the adatom coverage from 0 to 0.4, but it depends on coverage at higher than 0.4. By counting the darkened adatoms, Si-OCH(CH3)2 on the center adatom (m) and that on the corner adatom (n), it was found the m/n ratio is ca. 4 for the first dissociation of (CH3)2CHOH in virgin half unit cell, but it becomes ca. 1.9 and 1.8 when two and three Si-OCH(CH3)2 are contained in a half unit cell. This result reveals that the dissociation probability of (CH3)2CHOH at the adatom-rest atom pair site is influenced by the nearest Si-OCH(CH3)2 in the half unit cell. PMID- 15268617 TI - Electrorheology in nanopores via lattice Boltzmann simulation. AB - The conductance of ionic species through a cylindrical nanochannel is explored by means of a mesoscopic lattice Boltzmann numerical method. It is shown that in the absence of any external voltage, the ionic profiles develop a considerable amount of structure within the channel. The details of this structure depend on the ionic density, channel length, and Debye length. In the presence of a trans membrane electrostatic potential, less structure is observed and Ohm's law is found to hold. PMID- 15268618 TI - Photodissociation of hydrogen iodide on the surface of large argon clusters: the orientation of the librational wave function and the scattering from the cluster cage. AB - A set of photodissociation experiments and simulations of hydrogen iodide (HI) on Arn clusters, with an average size n = 139, has been carried out for different laser polarizations. The doped clusters are prepared by a pick-up process. The HI molecule is then photodissociated by a UV laser pulse and the outgoing H fragment is ionized by resonance enhanced multiphoton ionization in a (2 + 1) excitation scheme within the same laser pulse at the wavelength of 243 nm. The measured time of-flight spectra are transformed into hydrogen kinetic energy distributions. They exhibit a strong fraction of caged H atoms at zero-kinetic energy and peaks at the unperturbed cage exit for both spin-orbit channels nearly independent of the polarization. At this dissociation wavelength, the bare HI molecule exhibits a strict state separation, with a parallel transition to the spin-orbit excited state and perpendicular transitions to the ground state. The experimental results have been reproduced using molecular simulation techniques. Classical molecular dynamics was used to estimate the HI dopant distribution after the pick-up procedure. Subsequently, quasi-classical molecular dynamics (Wigner trajectories approach) has been applied for the photodissociation dynamics. The following main results have been obtained: (i) The HI dopant lands on the surface of the argon cluster during the pick-up process, (ii) zero-point energy plays a dominant role for the hydrogen orientation in the ground state of HI-Arn surface clusters, qualitatively changing the result of the photodissociation experiment upon increasing the number of argon atoms, and, finally, (iii) the scattering of hydrogen atoms from the cage which originate from different dissociation states seriously affects the experimentally measured kinetic energy distributions. PMID- 15268619 TI - First principles study of CO oxidation on TiO2(110): the role of surface oxygen vacancies. AB - The reactivities of the stoichiometric and partially reduced rutile TiO2(110) surfaces towards oxygen adsorption and carbon monoxide oxidation have been studied by means of periodic density functional theory calculations within the Car-Parrinello approach. O2 adsorption as well as CO oxidation are found to take place only in the presence of surface oxygen vacancies (partially reduced surface). The oxidation of CO by molecularly adsorbed O2 at the O-vacancy site is found to have an activation energy of about 0.4 eV. When the adsorbed O2 is dissociated, the resulting adatoms can oxidize incoming gas-phase CO molecules with no barrier. In all studied cases, once CO is oxidized to form CO2, the resulting surface is defect-free and no catalytic cycle can be established. PMID- 15268620 TI - Rotational dynamics of colloidal spheres probed with fluorescence recovery after photobleaching. AB - We report a polarized fluorescence recovery after photobleaching (pFRAP) method to measure the rotational dynamics of fluorescent colloids over a wide dynamic range. The method is based on the polarization anisotropy in the fluorescence intensity, generated by bleaching of fluorescently labeled particles with an intense pulse of linearly polarized laser light. The rotational mobilities of the fluorescent particles can be extracted from the relaxation kinetics of the postbleach fluorescence polarization anisotropy. Our pFRAP setup has access to correlation times over a range of time scales from tens of microseconds to tens of seconds, and is highly sensitive, so very low concentrations of labeled particles can be probed. We present a detailed description of the theoretical background of pFRAP. The performance of the equipment is demonstrated for fluorescent colloidal silica spheres, dispersed in pure solvents as well as in fd virus suspensions. PMID- 15268621 TI - Ab initio calculation of electrostatic multipoles with Wannier functions for large-scale biomolecular simulations. AB - It has long been known that accurate electrostatics is a key issue for improving current force fields for large-scale biomolecular simulations. Typically, this calls for an improved and more accurate description of the molecular electrostatic potential, which eliminates the artifacts associated with current point charge-based descriptions. In turn, this involves the partitioning of the extended molecular charge distribution, so that charges and multipole moments can be assigned to different atoms. As an alternate to current approaches, we have investigated a charge partitioning scheme that is based on the maximally localized Wannier functions. This has the advantage of partitioning the charge, and placing it around the molecule in a chemically meaningful manner. Moreover, higher order multipoles may all be calculated without any undue numerical difficulties. Tests on isolated molecules and water dimers, show that the molecular electrostatic potentials generated by such a Wannier-function based approach are in excellent agreement with the density functional-based calculations. PMID- 15268623 TI - New anti-inflammatory therapies and targets for asthma and chronic obstructive pulmonary disease. AB - Asthma and chronic obstructive pulmonary disease (COPD) are diseases of the airways with an underlying inflammatory component. The prevalence and healthcare burden of asthma and COPD is still rising and is predicted to continue to rise in the foreseeable future. Beta-agonists and corticosteroids form the basis of the therapies available to treat asthma. However, the treatments available for COPD, corticosteroids and anticholinergics, reduce the number and severity of exacerbations, but have a limited effect on slowing the progression of the disease. The inflammatory processes underlying the pathology of asthma have received a great deal of attention and more recently, those underlying COPD have begun to be elucidated. This has resulted in the identification of new targets that will allow the development of novel approaches by the pharmaceutical industry, which will be able to focus its efforts in an attempt to provide new and improved therapies to treat these debilitating diseases. The resultant therapies should impinge on the underlying development of these diseases rather than providing symptomatic relief or palliative treatment alone. This review will outline new targets and novel approaches currently under investigation, which may provide opportunities for novel anti-inflammatory therapeutic interventions that slow or halt disease progression in asthma and COPD. PMID- 15268624 TI - Novel therapy for Crohn's disease targeting IL-6 signalling. AB - The aetiology of Crohn's disease (CD) remains unknown, however, the importance of pro-inflammatory cytokines including IL-6 has been shown. IL-6 can transduce its signal into the cells lacking membrane-bound receptors by forming a complex with soluble IL-6R (sIL-6R). To pursue the therapeutic potential of IL-6 signalling blockade for CD, anti-IL-6R monoclonal antibody (mAb) was introduced to experimental colitis, and successfully prevented and treated intestinal inflammation by suppressing vascular adhesion molecules and by inducing lamina propria T cell apoptosis. Based on these results, the clinical trial of humanised anti-IL-6R mAb, MRA, was carried out. Eighty per cent of the patients given twice weekly MRA had a significantly better clinical response rate as compared to 31% of the placebo-treated patients. The response rate of the every-4-weeks regimen was 42%. The incidence of adverse events was similar in all groups. These data strongly suggest a therapeutic potential of MRA for CD. PMID- 15268625 TI - Recent developments in target identification against hepatitis C virus. AB - Chronic hepatitis C is a leading cause of liver cirrhosis and hepatocellular carcinoma worldwide. Recent progress in the understanding of the molecular virology of hepatitis C has allowed the identification of novel antiviral targets. Moreover, in vitro and in vivo model systems have been developed that allow the systematic evaluation of new therapeutic strategies. Exciting results from proof-of-concept clinical studies have now been reported for a specific hepatitis C virus serine protease inhibitor. These and other novel antiviral strategies may complement existing therapeutic modalities in the future. PMID- 15268626 TI - The complexity of tissue-type plasminogen activator: can serine protease inhibitors help in stroke management? AB - Stroke, the third leading cause of death in industrialised countries, represents a major burden on healthcare authorities. The elucidation of molecular events sustaining infarct evolution in experimental models has allowed the development of putative therapeutic agents. However, despite marked benefits in animals, most of them have failed in clinical trials. At present, the only approved therapy for stroke is early reperfusion by intravenous injection of the thrombolytic agent, tissue-type plasminogen activator (tPA). tPA-dependent thrombolysis sometimes promotes haemorrhage, but improves neurological outcome in a great proportion of patients, provided it is performed within the recommended therapeutic window. In addition to the benefit of tPA injection in the vascular compartment, this endogenously produced serine protease could also promote excitotoxic processes within the cerebral parenchyma. This article reviews the various aspects of tPA during stroke, and discusses potential improvements to current clinical management, with a particular emphasis on targeting the deleterious actions of tPA through endogenous serine protease inhibitors (serpins). PMID- 15268627 TI - Newly emerging Ca2+ entry channel molecules that regulate the vascular tone. AB - Local blood flow is critically determined by the arterial tone in which sustained Ca(2+) influx, activated by a variety of mechanisms, plays a central regulatory role. Recent progress in molecular biological research has disclosed unexpectedly diverse and complex facets of Ca(2+) entry channel molecules involved in this Ca(2+) influx. Candidates include several transient receptor potential (TRP) superfamily members such as TRPC1, TRPC4, TRPC6, TRPV2, TRPV4 and TRPM4, none of which exhibit simple properties attributable to a single particular role. Rather, they appear to be multimodally activated or modulated by receptor stimulation, temperature, mechanical stress or lipid second messengers generated from various sources, and may be involved in both acute vasomotor control and long-term vascular remodelling. This paper provides an overview of existing knowledge of TRP proteins, and their possible relationships with principal factors regulating the arterial tone (i.e., autonomic nerves, various autocrine and paracrine factors, and intravascular pressure). PMID- 15268628 TI - Development of ligand-targeted liposomes for cancer therapy. AB - The continued evolution of targeted liposomal therapeutics has resulted in new agents with remarkable antitumour efficacy and relatively mild toxicity profiles. A careful selection of the ligand is necessary to reduce immunogenicity, retain extended circulation lifetimes, target tumour-specific cell surface epitopes, and induce internalisation and subsequent release of the therapeutic substance from the liposome. Methods for assembling targeted liposomes, including a novel micellar insertion technology, for incorporation of targeting molecules that efficiently transforms a non-targeted liposomal therapeutic to a targeted one, greatly assist the translation of targeted liposome technology into the clinic. Targeting strategies with liposomes directed at solid tumours and vascular targets are discussed. The authors believe the development of ligand-targeted liposomes is now in the advanced stage and offers unique and important advantages among other targeted therapies. Anti-HER2 immunoliposomal doxorubicin is awaiting Phase I clinical trials, the results of which should provide new insights into the promise of ligand-targeted liposomal therapies. PMID- 15268629 TI - Is RGS-2 a new drug development target in cardiovascular disease? AB - Hypertension is the most common cardiovascular disease. The regulator of G protein signalling (RGS) proteins modify the activity of G proteins, and mice deficient in RGS-2 are hypertensive. On vascular smooth muscle, RGS-2 is involved in cross-talk between the nitric oxide (NO)-relaxation pathway and thrombin contraction pathway. RGS-2 binds to the cGMP-dependent protein kinase I-alpha from the NO relaxation pathway to terminate protease-activated receptor-1 signalling. It has been suggested that RGS-2 is a new drug development target for hypertension. Mice deficient in RGS-2 also have impaired antiviral immunity. It is difficult to envisage how RGS-2 could be targeted to have effects on the cardiovascular system without affecting immunity. Also, it is not clear whether or not targeting RGS-2 will have advantages over targeting receptors. Only an increased understanding of the physiological and pathological role of RGS-2 will help us resolve these issues. PMID- 15268630 TI - 5th Annual International Conference on HDL Cholesterol: metabolic pathways and drug developments. AB - Cardiovascular diseases (CVD) are the number one killer in the USA and are likely to be a significant cause of mortality in developing countries in the near future. High levels of high-density lipoprotein (HDL) are known to be inversely related to CVD incidence. However, whether or not HDL plays a direct preventative role or is simply a coincident marker for other protective processes has been controversial. Fortunately, recent discoveries have dramatically increased understanding of HDL function and recent clinical studies suggest that raising HDL levels may indeed be an effective strategy for preventing CVD. This meeting was organised to highlight state-of-the art studies focusing on the mechanisms of HDL function. Important work concerning small-molecule and immunological approaches for raising HDL were presented, including the targeting of regulatory nuclear receptors, plasma enzymes/transfer proteins that remodel HDL, and cell surface proteins that influence HDL formation and function. A detailed understanding of the molecular basis for the protective effect of HDL will hopefully lead to the development of new therapeutics that exploit this pathway. PMID- 15268631 TI - Efflux pumps: an answer to Gram-negative bacterial resistance? PMID- 15268632 TI - Novel modes of antifungal drug administration. AB - Administration of antifungals by routes other than that for which the agent was designed or approved have been utilised in attempts to provide directed therapy, reduce adverse effects and improve drug penetration into selected infection sites, such as the central nervous system, lungs and peritoneum. The most widely investigated agent utilising a novel method of drug delivery is amphotericin B. Dose forms for this agent include topicals (aerosol, nasal spray, irrigations, pastes, absorbable sponges, impregnated bone cement and gelatin), oral dosage forms (solutions, suspensions, tablets and so on) and ophthalmic preparations (drops, ointments and injections). Amphotericin B has been administered by routes such as oral, endobronchial, intrathecal, intracisternal, intra-articular, intraperitoneal, ophthalmic and as an antibiotic 'line lock'. Nystatin has been administered as an aerosol, percutaneous paste and bladder washes. Azoles, such as miconazole, fluconazole, ketoconazole and posaconazole, have been administered by novel methods but to a lesser degree. Most of these reports involve miconazole. The dose forms and routes of administration for azoles have included irrigants (bladder, joint), ophthalmic preparations (eye drops, intraocular injections, ointments), impregnated bone cement, endobronchial and intrathecal administration. Finally, both methylene blue (bladder washes) and flucytosine (peritoneal lavage, ophthalmic eye drops) have also been employed. Adequate evaluations of both the safety and efficacy of these therapies are most often hindered by prior or concomitant antifungal therapies, comorbidities and the lack of controlled clinical trials. In addition, the availability of newer treatment options, which demonstrate significant improvement in drug distribution and treatment-related adverse effects make many such novel modes of administration less practical or necessary. In contrast, the inhalation of antifungal aerosols, such as amphotericin B, is rapidly becoming a viable prophylactic option. PMID- 15268633 TI - Understanding HIV resistance, fitness, replication capacity and compensation: targeting viral fitness as a therapeutic strategy. AB - The increasingly prevalent emergence of drug-resistant virus strains in patients being treated with highly active antiretroviral regimens and the increasing rates of transmission of drug-resistant virus strains have focused attention on the critical need for additional antiretroviral agents with novel mechanisms of action and enhanced potency. Furthermore, novel means of employing highly active antiretroviral therapy are needed to reduce or eliminate the virological treatment failures that currently occur. Over the past several years, evidence has mounted supporting the fact that the emergence of resistant strains is associated with reductions in viral fitness, yielding decreases in plasma virus load in treated patients harbouring resistant populations of the virus. Additional mutations that serve to modify fitness (compensatory mutations) and mutations that impact the viral replication capacity also emerge under the selective pressure of drug treatment, and have both negative and positive effects on virus growth. Fitness is generally accepted to refer to the ability of HIV to replicate in a defined environment and thus is used to describe the viral replication potential in the absence of the drug. Although viral fitness and replication capacity are related in some ways, it is important to recognise that viral fitness is not the same as viral replication capacity. This review will assess the recent literature on antiviral drug resistance, viral fitness and viral replication capacity, and discuss means by which the adaptability of HIV to respond rapidly to antiviral treatment through mutation may be used against it. This would be done by treating patients with an aim to lock the deleterious mutations into the resistant virus genome, resulting in a positive therapeutic outcome despite the presence of resistance to the selecting agents. The review will specifically discuss the literature on nucleoside and non-nucleoside reverse transcriptase inhibitors, protease inhibitors, integrase inhibitors, fusion inhibitors, as well as other biological factors involved in viral fitness. PMID- 15268634 TI - Recent developments in human papillomavirus vaccines. AB - The association of certain high-risk human papillomaviruses with the development of anogenital cancer in humans is well-established. Numerous preclinical studies have underwritten the development of both prophylactic and therapeutic vaccine candidates for clinical evaluation. Prophylactic strategies are utilising virus like particles composed of the L1 viral capsid protein to induce neutralising antibodies while therapeutic approaches are aimed at generating specific T cells targeted at the viral E6 and/or E7 oncogene products. Thus far, human papillomavirus virus-like particle vaccines have proven to be clinically efficacious in the early trials looking at the prevention of infection. Important future milestones will be showing the prevention of high-grade cervical intraepithelial neoplasia and sufficient longevity for such protection. Different types of therapeutic vaccines including peptide, protein, DNA or viral vector based vaccines have proven to be safe and immunogenic in patients, although there is often no correlation with clinical outcome. The possibility of combined prophylactic and therapeutic vaccines may offer the best chance for a significant reduction in the incidence of death from cervical cancer worldwide. PMID- 15268635 TI - Cephalosporins in clinical development. AB - Bacterial resistance to established classes of antibiotics in clinical use is continuing to increase, making the need for new agents that can be used to treat the newly multi-drug resistant organisms steadily more urgent. Cephalosporins have been a successful group of antibiotics since they were first introduced to combat drug-resistant organisms, including staphylococci. The history of cephalosporins has emphasised an improvement of their stability towards beta lactamases, thus expanding their spectrum of activity against important Gram negative pathogens. New cephalosporins that have potent activity against multi resistant Gram-positive bacteria, including methicillin-resistant staphylococci and penicillin-resistant pneumococci have recently entered clinical development. At least two of these, BAL-5788 and S-3578, also have Gram-negative activity, which is comparable to that of the third-and fourth-generation cephalosporins, making them broad-spectrum agents that could be used in hospital infections where methicillin-resistant staphylococci is likely to be present. PMID- 15268636 TI - Novel antihypertensive agents. AB - Coronary heart disease and cerebrovascular disease continue to be the leading causes of illness and death among adults from developed countries. Their prevalence is strongly related to the effects of many different risk factors, including high blood pressure, cigarette smoking, dyslipidaemia and diabetes. The management of cardiovascular (CV) risk factors should be viewed as an integrated strategy of intervention aimed at correcting as many of the underlying causes of CV disease as possible. Blocking the renin-angiotensin-aldosterone system with new, well-tolerated antihypertensive drugs, and blood pressure regulation by means of new drugs that also reduce plasma cholesterol levels or improve insulin resistance and glucose tolerance, could lead to a reduction of CV diseases. PMID- 15268637 TI - New developments in the use of beta-blockers for the management of heart failure. AB - Chronic heart failure (HF) has become a significant healthcare problem in the US. The number of new cases per year continues to grow steadily due to an ageing population and improved survival from acute coronary syndromes. As a consequence, the management of HF patients is of great importance. Effective management of HF includes stabilising the patient and improving the clinical symptoms associated with HF. Patients with HF have increased sympathetic nervous system activity that contributes to impaired cardiovascular function over time and subsequently results in death. beta-blockers prevent such impairment through inhibition of the sympathetic nervous system neurohormonal pathway. Numerous clinical trials conducted over the past decade have demonstrated that beta-blockers, in conjunction with angiotensin-converting enzyme inhibitors, are not only effective but are superior to other medical interventions for the treatment of HF. The standard of care for patients with HF now includes beta-blockers as well as ACE inhibitors. PMID- 15268638 TI - Rheumatoid arthritis: developing pharmacological therapies. AB - Rheumatoid arthritis (RA) is a chronic inflammatory disease that often results in significant morbidity, mortality and disability. Over the past 20 years a better understanding of the pathogenesis of RA has led to the development of new approaches to disease treatment. The recent introduction of biological agents has changed the treatment paradigm for RA. The success of early biological therapies including TNF-alpha and IL-1 antagonists has spurred interest in the development of additional novel targets in the treatment of RA. Biological therapies approved for other indications, such as rituximab, are now being evaluated for the treatment of rheumatic diseases such as RA. A co-stimulatory blocker, abatacept, is also in pivotal Phase III trials. This article reviews evolving pharmacological therapies in RA with an emphasis on the newer approaches to treatment including inhibition of cognate signalling and T- and B-cell targets. PMID- 15268639 TI - Treatment of autoimmune mucocutaneous blistering diseases with intravenous immunoglobulin therapy. AB - Autoimmune mucocutaneous blistering diseases (AMBDs) are a group of rare diseases that affect the skin and mucous membranes and are potentially fatal. They have variable clinical presentation, course and prognosis. Their immunopathology is well-characterised and target antigens have been studied. Many patients respond to conventional therapy, which consists of high-dose long-term systemic corticosteroids with an immunosuppressive agent, but side effects develop that can produce a poor quality of life. Many patients develop significant sequelae, such as blindness, loss of voice, vaginal and anal stenosis. In most patients cause of death is opportunistic infections secondary to immune suppression. To date, intravenous immunoglobulin (IVIg) has been reported to have benefited 156 patients with AMBDs. Its most important features include the ability to reduce or eliminate conventional therapy, the enabling of clinical control, the ability to induce and maintain long-term clinical remission, the capacity for usage based on a defined protocol with a described end point and a resulting increase in quality of life. IVIg produces the best clinical outcome when combined with aggressive topical therapy, sublesional injections of triamcinolone and rapid detection, and early treatment of cutaneous and mucosal infection. Successful therapy requires a physician to spend significant time with each patient. This manuscript provides the opinion of the author on the current use of IVIg to treat AMBDs. PMID- 15268640 TI - COX-2 inhibitors as adjunctive therapy in schizophrenia. AB - Cyclooxygenase-2 (COX-2) is constitutively expressed in the central nervous system, and is thought to have an important functional role therein. COX-2 interacts with neurotransmitters such as acetylcholine, 5-hydroxytryptamine and glutamate but is also involved in the regulation of the central nervous system immune system and in inflammation via the effects of prostaglandins, in particular prostaglandin E2. A general therapeutic effect of the COX-2 inhibitor celecoxib on symptoms of schizophrenia was observed during a prospective, randomised, double-blind study of celecoxib add-on treatment to the atypical antipsychotic risperidone. The results from this trial of adjunctive therapy with a COX-2 inhibitor in schizophrenia are encouraging, and the findings support the view that an immunological/inflammatory process is involved in the pathogenesis of the disease. The add-on to an antipsychotic design of the study was chosen due to ethical reasons; in less acute schizophrenic states a monotherapy with COX-2 inhibitors would be interesting. From a theoretical point of view, other psychiatric indications for selective COX-2 inhibitors are discussed. COX-2 inhibitors have failed to show therapeutic effects in Alzheimer's disease but studies from basic research and a clinical perspective suggest it has an effect on disturbed cognition. In depression, however, signs of inflammation have been described for many years. Although results of clinical studies with COX-2 inhibitors in depression are still lacking, clinical improvement of a depressive syndrome has been observed in patients who have been treated with the COX-2 inhibitor rofecoxib due to other indications. These preliminary clinical data are encouraging for clinical therapeutic effects of the selective COX-2 inhibitors in psychiatric disorders, although these effects have to be confirmed in larger clinical studies. PMID- 15268641 TI - Preclinical evaluation of novel antibacterial agents by microbiological and molecular techniques. AB - The defining property of an antibacterial agent is its ability to selectively interfere with bacterial growth and/or survival. Consequently, a considerable and crucial part of the preclinical evaluation of any novel antibacterial drug involves judging and characterising its effects on bacteria in vitro. These critical stages in drug development are sometimes made to appear somewhat trivial, sandwiched as they are between the highly demanding antibacterial discovery process and the formidable task of demonstrating safety and efficacy in vivo. However, careful biological evaluation in vitro is key to quantifying and understanding the basis of the antibacterial activity, providing preliminary indications and evaluations of therapeutic potential, assessing the likelihood for the development of bacterial resistance, guiding chemical refinement and assisting subsequent stages of the appraisal of any new antibacterial drug. This review covers concepts in, and strategies for, the in vitro microbiological and molecular evaluation of antibacterial drug candidates. PMID- 15268642 TI - 5th Antiviral Drug Discovery and Development Summit. AB - The 5th Antiviral Drug Discovery and Development Summit provided an up-to-date snapshot of the ongoing developments in the area. The topics covered ranged from updates on recently launched drugs (Kaletra), Fuzeon) and new investigational inhibitors (T-1249, Reverset, UK-427857, L-870810, PA-457, remofovir, VX-950), to the discovery of new antiviral targets and advances in technologies that may provide the substrate for the next generation of therapeutics. It is apparent from the range of presentations that much of today's efforts are focused on developing new classes of HIV inhibitors (gp41, integrase), while there is also considerable progress in hepatitis C, where a number of inhibitors have or should reach proof-of-concept studies in the coming months. Here we provide the highlights of this meeting, with particular emphasis on the new developments in HIV and hepatitis C virus. PMID- 15268643 TI - Strategic Research Institute G-Protein-Coupled Receptors Drug Discovery World Summit. AB - The Strategic Research Institute provided a well-organised 2-day summit that offered presentations and posters on new assay technology, structure-based small molecule discovery and examples of clinical candidates targeted to G-protein coupled receptor (GPCR) targets. A wide variety of topics were presented providing recent advances in GPCR target selection, bioassay-enabling technology and medicinal chemistry targeted to GPCR-relevant chemical libraries. GPCRs continue to be an attractive platform for drug discovery. PMID- 15268644 TI - Gastrointestinal safety of paracetamol: is there any cause for concern? AB - Paracetamol is classically considered as a very safe analgesic/antipyretic compound and, more specifically, as being virtually devoid of any gastrointestinal (GI) ulcerogenic potential. Accordingly, it is widely stated that paracetamol is particularly suitable for patients at high risk of developing GI ulcers or bleeds. This view has been challenged by recent epidemiological studies using computerised prescription data, which indicated that paracetamol exhibits dose-dependent GI toxicity. However, the results of these studies are most likely incorrect for reasons of inherent biases and confounding. Furthermore, their findings conflict with those of clinical trials and case control studies in which information about drug exposure was obtained by direct questioning of both cases and controls. Finally, paracetamol, especially at high doses, may induce upper GI symptoms such as abdominal pain/discomfort, heartburn, nausea or vomiting. Conversely, the risk for ulcers and ulcer complications due to paracetamol is not supported by available data. PMID- 15268645 TI - Risk:benefit ratio of nonsteroidal anti-inflammatory drugs in systemic lupus erythematosus. AB - Nearly 80% of patients with systemic lupus erythematosus (SLE) are treated with NSAIDs for fever, arthritis, serositis and headaches. This article reviews currently available literature on non-selective and selective inhibitors of cyclooxygenases, with an emphasis on the efficacy and safety profile reported in SLE patients. All NSAIDs, regardless of their cyclooxygenase selectivity, induced renal side effects including sodium retention and reduction in glomerular filtration rate. In addition, lupus nephritis is a risk factor for NSAID-induced acute renal failure. NSAID-induced hepatotoxicity is increased in SLE patients in addition to cutaneous and allergic reactions. Finally, aseptic meningitis has been reported more frequently in NSAID-treated SLE patients. Nevertheless, NSAIDs can safely be prescribed to most lupus patients provided that their administration is re-evaluated on a regular basis and the patient is closely monitored. PMID- 15268646 TI - Safety of hydroxyurea in the treatment of HIV infection. AB - Lifelong adherence to very complex anti-HIV therapy presents drawbacks such as drug resistance and chronic drug-related toxicity, underscoring the need for innovative therapeutic options. As it is becoming increasingly evident that immune activation may be responsible for immune pathology, novel approaches to limit immune activation are under investigation. Hydroxyurea is the prototype of a new family of anti-HIV drugs called virostatics, acting both as an antiviral (directly suppressing HIV) and cytostatic (preventing immune system overactivation). Data from in vitro and clinical studies have proven that hydroxyurea-based regimens are effective options for patients with HIV. However, concerns over hydroxyurea toxicity have limited its use. This review critically examines the role of hydroxyurea for HIV-infected patients, focusing on past and recent clinical trials including the RIGHT 702 study, which identified the safest and most efficacious hydroxyurea dose. PMID- 15268647 TI - Benefit-risk assessment of interferon-beta therapy for relapsing multiple sclerosis. AB - IFN-beta therapy has a central place in the management of relapsing multiple sclerosis, as demonstrated by the pivotal studies of three IFN-beta treatment regimens. However, questions remain concerning the optimal choice of preparation and dose regimen. The benefit-risk ratio for a given preparation is an important consideration in optimising treatment for an individual patient. Of the three IFN beta preparations currently available, all have shown benefit on activity measures (relapses and active lesions apparent on magnetic resonance imaging), while benefit on progression measures (disability and total lesion burden) has been less consistent. Available data across studies suggest that dose and/or dose frequency are important determinants of efficacy, a finding supported by direct comparative data on different IFN-beta preparations. The added benefit of high dose, high-frequency IFN-beta therapy is not achieved at the cost of compromised safety or tolerability, indicating that three-times-weekly treatment offers a superior benefit-risk ratio to once-weekly treatment. The potential advantages of IFN-beta therapy may be enhanced by regular monitoring of efficacy and safety in order to maintain patients on therapy beyond the first few months when side effects are most apparent. PMID- 15268648 TI - Safety of growth hormone replacement therapy in adults. AB - Growth hormone (GH) is a powerful metabolic hormone that regulates fuel homeostasis through its protein anabolic and lipolytic actions. The introduction of recombinant human GH has expanded the narrow indication of treating children with severe GH deficiency (GHD) to include a broader target population of children with growth retardation and short stature and adults with hypopituitarism and severe GHD. Furthermore, because children continue to receive GH replacement therapy into adult life, the duration of treatment exposure has increased and the safety of long-term GH treatment has become increasingly important. This is of particular concern given that GH-deficient children and adults may be more vulnerable to the mitogenic stimuli of GH and insulin-like growth factor-1, both because of the underlying cause of GHD and also because of previous treatment such as radiotherapy and chemotherapy. This review focuses on the safety of treating adults with severe GHD, with specific emphasis on dose regimens, carbohydrate metabolism, neoplasia, and morbidity and mortality. Available experience from long-term replacement therapy, studies using supraphysiological doses of GH in adults and lessons learned from patients with acromegaly who have high endogenous GH levels over many years, is considered. PMID- 15268649 TI - Efficacy and safety of trastuzumab. AB - The human epidermal growth factor receptor (HER) 2 is overexpressed in approximately 20-25% of human breast cancers and is an independent adverse prognostic factor. Targeted therapy directed against this receptor has been developed in the form of a humanised monoclonal antibody, trastuzumab. This antibody has shown activity as a single agent in metastatic breast cancer both prior to chemotherapy and in heavily pretreated patients. A pivotal Phase III trial has demonstrated that its use in combination with an anthracycline or paclitaxel results in a significant improvement in survival, time to progression, and response. This has recently been reinforced by another trial with docetaxel. The HER2 status of a tumour is a critical determinant of response to trastuzumab based treatment. Those that express HER2 at the highest level on immunohistochemistry (IHC), 3+, derive more benefit from treatment with trastuzumab than those with overexpression at the 2+ level. Benefit correlates best with tumours that are positive on fluorescence in situ hybridisation for HER2, regardless of IHC status. Treatment with trastuzumab is generally well tolerated with a low incidence of adverse events. Some patients may experience fever, chills, dyspnoea and pain, particularly with the first administration. Unexpectedly, cardiac toxicity has developed in some patients treated with trastuzumab, and this has a higher incidence in those treated in combination with an anthracycline. 'Cross-talk' between the oestrogen receptor and HER2 pathway has stimulated interest in using trastuzumab in combination with endocrine therapy. Current clinical trials are investigating the role of this agent in the adjuvant setting. PMID- 15268650 TI - Causality assessment in drug-induced hepatotoxicity. AB - Drugs are currently an important cause of liver disease, ranked as the most frequent reason for acute liver failure. Despite recent advances in knowledge of the mechanisms implicated in drug-induced hepatocellular damage and cholestasis, as well as the identification of several risk factors, the diagnosis of hepatotoxicity remains a difficult task because specific tests are not available. In a step-by-step approach, the incrimination of a drug in liver symptoms requires a high degree of suspicion on the part of the physician, temporal eligibility, awareness of the drug's hepatotoxic potential, the exclusion of alternative causes of liver damage, and the ability to detect the presence of subtle data that favour a toxic aetiology. Ultimately, the use of diagnostic algorithms may add consistency to the diagnostic process either by translating the suspicion into a quantitative score or by providing a framework that emphasises the features that merit attention in cases of suspected hepatic adverse reactions. PMID- 15268651 TI - Safety of verteporfin for treatment of subfoveal choroidal neovascular membranes associated with age-related macular degeneration. AB - Photodynamic therapy (PDT) is a novel treatment entity that exploits the photophysical properties of various photosensitive chemical entities which, upon light activation, results in targeted photooxidation and subsequent tissue destruction. The antiangiogenic properties of PDT have been adapted for treatment of subfoveal choroidal neovascular membranes due to disease states such as age related macular degeneration (AMD). Historically, PDT has been limited by a lack of suitable photosensitive dyes. However, agents such as verteporfin, a second generation benzoporphyrin derivative, appear to be free from the extensive phototoxicity that limited the success of previous agents. Verteporfin has a high affinity for choroidal neovascular membranes, typically found with exudative AMD, and upon photoactivation results in targeted microvascular damage and thrombus formation with resultant vessel occlusion. Scrutiny of diagnostic indicators for verteporfin administration, including critical angiographic evaluation of lesion size and visual acuity, is essential to treatment success. Large lesions with relatively good visual acuity (20/50 or better) may be at particular risk for marked vision loss following verteporfin administration. Lesion composition also appears to influence visual outcome with verteporfin use. The safety of verteporfin is directly dependent upon the appropriate integration of dosage, infusion and light activation required for a suitable pharmacotherapeutic outcome. When used appropriately, and with adequate patient education regarding photosensitivity, the risk-benefit of verteporfin for the medical treatment of neovascular AMD is favourable. PMID- 15268652 TI - Patients' role in reporting adverse drug reactions. AB - This review discusses the involvement of patients in the reporting of adverse drug reactions (ADRs). Patients benefit from drugs but also experience their adverse effects. Since concerns about the safety of drugs are also patients' concerns, the patient could also play a part in decreasing the risks of drug therapy. Patient interest in the safety aspects of drugs is evident. At an international level, the merits of patient reports are being considered. To date, the literature does not yet provide any actual results in relation to the detection of ADRs by patients. Different considerations regarding ADR reporting by patients are discussed. The authors conclude that we should positively value patients' involvement in drug therapy and their concern regarding possible adverse effects. As a consequence, patients' reports on ADRs should be accepted. PMID- 15268653 TI - Safety of droperidol in behavioural emergencies. AB - By the year 2000, droperidol had become a standard drug for the treatment of behavioural emergencies in both psychiatric and medical settings. In 2001, the US FDA issued a 'black box' warning, citing cases of QT prolongation and/or torsades de pointes. As a result, the use of droperidol has been sharply circumscribed. The authors will review the literature on antipsychotic medications in general, focusing on droperidol in particular, with regard to QT interval prolongation, dysrhythmia, and sudden death. In addition, the mechanism of drug-induced QT interval prolongation will be discussed. The authors will then review their extensive experience with droperidol. The authors conclude that, while in theory droperidol may prolong the QT interval to an extent similar to thioridazine, its long history of clinical use has shown no pattern of sudden deaths analogous to those that provoked the FDA warning. Although the numbers presented by the FDA initially appear alarming, after further evaluation it is clear that more definitive studies should have been carried out. Droperidol is safe, extremely effective, and now underused as a treatment for severely agitated or violent patients. PMID- 15268654 TI - Strategies for more effective and reliable ADME. 30-31 March 2004, London, UK. AB - This conference was organised by Vision in Business, in order to address the issue that 'more than 40% of drug preclinical failure is due to ADME-related issues and many drugs are later withdrawn because of unanticipated drug-drug interactions (C Masimirembwa, AstraZeneca Molndal, Sweden). Academic and industrial scientists were brought together to discuss and present current knowledge/theories on subjects of importance in preclinical and clinical drug development. These included in vivo/in vitro correlations, approaches and experimental models used, in silico models and methods, metabolic databases and their application, as well as safety and financial outcomes. Participants were challenged by the issues covered and encouraged to share their own experiences and opinions during extensive discussions with other scientists. PMID- 15268655 TI - The choroid plexus: function, pathology and therapeutic potential of its transplantation. AB - The choroid plexus (CP) produces cerebrospinal fluid (CSF) and forms the blood CSF barrier. However, the CP may have additional functions in the CNS beyond these traditional roles. Preclinical and clinical studies in ageing and neurodegeneration demonstrate anatomical and physiological changes in CP, suggesting roles in normal and pathological conditions and potentially endogenous repair processes following trauma. One of the broadest functions of the CP is establishing and maintaining the extracellular milieu throughout the brain and spinal cord, in part by secreting numerous growth factors into the CSF. The endogenous secretion of growth factors raises the possibility that transplantable CP might enable delivery of these molecules to the brain, while avoiding the conventional molecular and genetic alterations associated with modifying cells to secrete selected products. This review describes some of the anatomical and functional changes of CP in ageing and neurodegeneration, and recent demonstrations of the therapeutic potential of transplanted CP for neural trauma. PMID- 15268656 TI - Protein and peptide parenteral controlled delivery. AB - Protein and peptide delivery has been a challenge due to their limited stability during preparation of formulation, storage and in vitro and in vivo release. These biopolymers have traditionally been administered via intramuscular or subcutaneous routes. Recent efforts have been made to develop formulations for non-invasive routes of administration, including oral, intranasal, transdermal and transmucosal delivery. Despite these efforts, invasive delivery remains the main method of administering peptide and protein drugs. This review focuses on recent developments in injectable, polymeric controlled-release formulations, with an emphasis on hydrogels and particulate systems. PMID- 15268657 TI - Nanoparticle-mediated gene delivery: state of the art. AB - With the development of genomic and proteomic technologies, the prospect for gene therapy has progressed rapidly. This has been partly possible due to the emergence of a diverse array of polymeric and non-polymeric nanoparticles that are being investigated for their ability to deliver genes and drugs. In this review, particles have been pragmatically divided as chitosan-related and chitosan-unrelated nanomaterials. The state of the art in terms of the development, characterisation and evaluation of their in vitro and/or in vivo potential is discussed for each of these various particles. Although substantial progress has been made, the potential of these particles in the clinical arena and human responses remain to be evaluated. It is hoped that this review will provide an impetus for further studies of these particles, with the ultimate intent that one or more of these diverse nanoparticle-based non-viral approaches for gene transfer will translate from 'bench to bedside' in the future. PMID- 15268658 TI - Gene therapy for thyroid cancer. AB - Thyroid carcinomas are suitable targets for gene therapy because they can be highly lethal on one hand, while being susceptible to specific tumour targeting on the other hand. Several gene therapy modalities have been evaluated so far in experimental models of thyroid cancer, including tumour suppressor gene replacement, oncogene inhibition, suicide gene therapy, immunotherapy, antiangiogenesis, and viral oncolysis. All of these strategies have shown promising results, but clinical studies are lacking. Based on the clinical experience achieved in a pilot study in patients with advanced thyroid cancer and on clinical results in other types of solid cancer, it is suggested that combined gene therapy approaches, as well as multimodality therapeutic regimens, including gene therapy and conventional treatments, should be pursued to achieve clinically significant results. PMID- 15268659 TI - Conditionally replicative adenovirus for gastrointestinal cancers. AB - The clinical outcome of advanced gastrointestinal (GI) cancers (especially pancreatic and oesophageal cancers) is dismal, despite the advance of conventional therapeutic strategies. Cancer gene therapy is a category of new therapeutics, among which conditionally replicative adenovirus (CRAd) is one promising strategy to overcome existing obstacles of cancer gene therapy. Various CRAds have been developed for GI cancer treatment by taking advantage of the replication biology of adenovirus. Some CRAds have already been tested in clinical trials, but have fallen short of initial expectations. Concerns for clinical applicability include therapeutic potency, replication selectivity and interval end points in clinical trials. In addition, improvement of experimental animal models is needed for a deeper understanding of CRAd biology. Despite these obstacles, CRAds continue to be an exciting area of investigation with great potential for clinical utility. Further virological and oncological research will eventually lead to full realisation of the therapeutic potential of CRAds in the field of GI cancers. PMID- 15268660 TI - Advances in the development of therapeutic nucleic acids against cervical cancer. AB - Cervical cancer is the second most common neoplastic disease affecting women worldwide. Basic, clinical and epidemiological analyses indicate that expression of high-risk human papillomaviruses (HPVs) E6/E7 genes is the primary cause of cervical cancer and represent ideal targets for the application of therapeutic nucleic acids (TNAs). Antisense oligodeoxyribonucleotides (AS-ODNs) and ribozymes (RZs) are the most effective TNAs able to inhibit in vivo tumour growth by eliminating HPV-16 and HPV-18 E6/E7 transcripts. Expression of multiple RZs directed against alternative target sites by triplex expression systems may result in the abrogation of highly variable HPVs. More recently, RNA interference (RNAi) gene knockdown phenomenon, induced by small interfering RNA (siRNA), has demonstrated its potential value as an effective TNA for cervical cancer. siRNA and aptamers as TNAs will have a place in the armament for cervical cancer. TNAs against cervical cancer is in a dynamic state, and clinical trials will define the TNAs in preventive and therapeutic roles to control tumour growth, debulk tumour mass, prevent metastasis and facilitate immune interaction. PMID- 15268661 TI - Using antibodies in tumour immunotherapy. AB - The role of antibodies as therapeutic cancer vaccines includes two distinct approaches, which are summarised in this review, namely anti-idiotypic vaccines and antigen-antibody complex therapies. Bispecific antibodies directed against T cells or antigen-presenting cells are also referenced. The report focuses on theoretical issues, laboratory data on the mechanism of action, examples of humoral and cellular immune induction, and novel therapeutic advances in vaccine development. The biology of antigen processing and recent advances in the field of dendritic cell biology are critical to understanding the potent immune response induction. Future directions include combination therapies to manipulate immune regulatory mechanisms and to enhance clinical effects. Additional applications of antibodies targeting costimulatory or regulatory receptors on antigen-presenting cells and T cells, neutralising immune suppressive cytokines, and depleting T regulatory cells hold promise for future mono- and particularly combination therapies. PMID- 15268662 TI - Messenger RNA-based vaccines. AB - RNA is the only molecule known to recapitulate all biochemical functions of life: definition, control and transmission of genetic information, creation of defined three-dimensional structures, enzymatic activities and storage of energy. Because of its versatility and thanks to several recent scientific breakthroughs, RNA became the focus of intense research in molecular medicine at the beginning of the millennium. In particular, mRNA can be seen as a safe and efficient alternative to protein-, recombinant virus- or DNA-based therapies in the field of vaccination. This review summarises the most remarkable advances in this area and presents the advantages and limits of the five different mRNA-based vaccination methods. The paper will present the official, industrial and financial aspects of mRNA-based vaccination that are paving the way for therapeutic and prophylactic drugs with mRNA as the active component. PMID- 15268663 TI - West Nile virus vaccines. AB - West Nile virus (WNV) is a mosquito-borne flavivirus that is emerging as a global pathogen. In the last decade, virulent strains of the virus have been associated with significant outbreaks of human and animal disease in Europe, the Middle East and North America. Efforts to develop human and veterinary vaccines have taken both traditional and novel approaches. A formalin-inactivated whole virus vaccine has been approved for use in horses. DNA vaccines coding for the structural WNV proteins have also been assessed for veterinary use and have been found to be protective in mice, horses and birds. Live attenuated yellow fever WNV chimeric vaccines have also been successful in animals and are currently undergoing human trials. Additional studies have shown that immunisation with a relatively benign Australian variant of WNV, the Kunjin virus, also provides protective immunity against the virulent North American strain. Levels of efficacy and safety, as well as logistical, economic and environmental issues, must all be carefully considered before vaccine candidates are approved and selected for large-scale manufacture and distribution. PMID- 15268664 TI - Future directions for the field of oncolytic virotherapy: a perspective on the use of vaccinia virus. AB - Oncolytic virotherapy is an emerging biotherapeutic platform based on genetic engineering of viruses capable of selectively infecting and replicating within cancer cells. Such viruses have been found to be both safe and to produce antitumour effects in a number of Phase I and II clinical trials. Early work in this field has been pioneered with strains of adenovirus which, although well suited to gene therapy approaches, have displayed certain limitations in their ability to directly destroy and spread through tumour tissues, particularly after systemic administration. Investigators have subsequently been examining the feasibility of using a variety of different viruses as oncolytic agents. Vaccinia virus is perhaps the most widely administered and successful medical product in history; it displays many of the qualities thought necessary for an effective antitumour agent and is particularly well characterised in people due to its role in the eradication of smallpox. Vaccinia has a short life cycle and rapid spread, strong lytic ability, inherent systemic tumour targeting, a large cloning capacity and well-defined molecular biology. In addition, the virus produces no known disease in humans, has been delivered safely to millions of people and has already demonstrated antitumoural efficacy in trials with vaccine strains. These qualities, along with strategies for further improving the safety and antitumour effectiveness of vaccinia, will be discussed in relation to the broad spectrum of clinical experience already achieved with this virus in cancer therapy. PMID- 15268665 TI - 90Yttrium-ibritumomab tiuxetan: a novel treatment for non-Hodgkin's lymphoma. AB - Indolent non-Hodgkin's lymphomas (NHLs) comprise 35-40% of all adult NHLs. No standard curative therapy exists for these malignancies. Newer therapies, including monoclonal antibodies and radioimmunotherapy, have had notable activity in chemotherapy-resistant patients. 90Yttrium-ibritumomab tiuxetan is a radioimmunoconjugate approved by the Food and Drug Administration in February 2002 for the treatment of relapsed and refractory, low-grade, follicular or transformed NHL. Reported response rates are 67-80%, with a median remission duration of approximately 12 months. Toxicities are primarily neutropenia and thrombocytopenia, with few infectious complications. 90Yttrium-ibritumomab tiuxetan is an effective therapeutic alternative for the treatment of NHL. PMID- 15268666 TI - Anakinra: an inhibitor of IL-1 for the treatment of rheumatoid arthritis. AB - Anakinra (Amgen, Inc.) is a specific receptor antagonist of IL-1 that differs from naturally occurring IL-1 receptor antagonist by the presence of a methionine group. Anakinra has been shown to be of benefit in patients with active rheumatoid arthritis, either when given alone or in combination with methotrexate, as assessed by improvement in clinical signs and symptoms, decreased radiographic progression and improvement in patient function, pain and fatigue, although it appears to be effective in fewer patients than anti-TNF agents. It has a favourable safety profile as demonstrated in clinical trials. The physician and patient must be cognizant of serious infectious episodes. Many of the rare side effects seen with TNF blockers, such as tuberculosis, other opportunistic infections, worsening of congestive heart failure and the development of demyelinating disease, have not been seen in patients treated with anakinra. Anakinra should not be given in combination with anti-TNF agents. PMID- 15268667 TI - Small molecules for small minds? The case for biologic pharmaceuticals. AB - Biologic pharmaceuticals are gaining in both market share and clinical utility compared with small molecule therapeutics. This market growth is, in part, reflective of a field of science entering its toddlerhood, where with increased maturity, both development timelines and costs of manufacturing for these complex molecules will decrease, further enhancing the profitability side of the equation. Although a firm understanding of the rules governing toxicity (especially antibody responses to therapeutic proteins) remains to be defined, it is clear that proteins are less prone to much of the idiosyncratic toxicity associated with small molecule drug candidates. Proteins are disadvantaged in that they are unlikely to find much use in targeting intercellular processes; however, they have clear strengths over small molecules in targeting protein protein interactions and the specific targeting of surface features of particular cells (e.g., in oncology). As each aspect of protein pharmaceutical technology advances, it is clear that this will be the major area for growth in the industry over the next decade. PMID- 15268668 TI - Eleventh Annual Conference of the American Society for Neural Transplantation and Repair. AB - The Eleventh Annual Conference of the American Society for Neural Transplantation and Repair was held at the Sheraton Sand Key Resort in Clearwater Florida over a typically warm and sunny 4 days. The scientific programme was organised by Dr T Collier of Rush Presbyterian Medical Center and Dr P Bickford of the University of South Florida. The 2004 conference was attended by a large portion of the approximately 300 society members. The purpose of the meeting was to bring together clinicians and basic researchers focused on areas ranging from understanding the biology of degenerative CNS diseases to utilising a variety of cell-based therapies to prevent neuronal cell death or repair and reconstitute damaged brain circuits. PMID- 15268669 TI - Induction of tolerance to self-antigens using genetically modified bone marrow cells. AB - The challenge of finding a lasting cure for autoimmune disease(s) has not been met. Although the use of systemic anti-inflammatory agents still dominates the treatment of these diseases, there is a push towards developing novel and more specific strategies. In addressing autoimmunity, there is the intrinsic need to understand the mechanisms that lead to the development and maintenance of immunological tolerance to self-antigens. Experimental evidence has shown that directed antigen expression in the thymus can induce immunological tolerance to that antigen. This forms the cornerstone of one strategy directed towards the cure of autoimmunity. In this strategy, individuals with autoimmune disease are transplanted with bone marrow stem cells that have been genetically modified and in this way allow expression of the self-antigen in the thymus. PMID- 15268670 TI - Bone engineering by controlled delivery of osteoinductive molecules and cells. AB - Bone regeneration can be enhanced or accelerated by the delivery of osteogenic signalling factors or bone forming cells. These factors have commonly provided benefit when retained at the defect site with a delivery vehicle formed from natural or synthetic materials. Growth factors can be directly delivered as recombinant proteins or expressed by genetically modified cells to induce bone formation. Furthermore, bone regeneration has been achieved with the transplantation of various cell types that can participate in bone healing. Carriers utilised for the delivery of osteoinductive material allow for a prolonged presentation at the repair site and the timing of presentation can be readily adjusted to correspond to the extent necessary for bone regeneration. This review examines some of the recent developments in delivery systems used to manage the presentation of these factors at the desired site. Moreover, the authors provide suggestions for continued progress in bone regeneration. PMID- 15268671 TI - Controlled-release and local delivery of therapeutic antibodies. AB - Human and humanised antibodies are now poised to become a major new class of protein-based therapeutic agents. A significant fraction of new drugs in clinical testing (approximately 20% in 2002) are antibody classes. Monoclonal antibodies (mAbs) with high affinities against newly discovered disease targets, both cellularly and extracellularly, are now clinically proven to elicit high bioactivities against numerous diseases, including tumours, infections, asthma, inflammation, arthritis and osteoporosis. Clinical humanised antibody delivery is typically intravenous, with large multiple doses (grams) required for systemic volumes of distribution. Due to the relatively high costs of both this drug type, and its common mode of administration, alternatives are sought where doses might be reduced and the bioavailability and efficacy enhanced. Local, controlled release methods that deliver antibodies locally to site of disease, offer new possibilities with these potential advantages. However, protein drugs frequently exhibit formulation challenges when packaged in delivery vehicles, and as globular proteins, antibodies are no exception. Several examples of mAb controlled-release and local delivery strategies against several disease targets are reviewed. Importantly, several antibody delivery methods work in tandem with existing clinically-accepted therapeutics, sometimes exhibiting potentiating or synergistic effects in animal models with small molecule, systemically administered drugs. PMID- 15268672 TI - Enhancement of vaccine potency through improved delivery. AB - The need for more potent, safe and well-characterised vaccines has necessitated the discovery and development of new vaccine technologies. These include adjuvants to target the innate immune system to provide a stimulus that potentiates the development of an antigen-specific immune response, and delivery systems to ensure that the antigen and adjuvant are localised to the appropriate immune compartments. Several such technologies are being tested in human clinical trials and a few have been licensed for limited use in human vaccines. This review will highlight some of the promising technologies that may have an impact on how vaccines are administered. PMID- 15268673 TI - Gene therapy for proliferative ocular diseases. AB - Proliferative ocular diseases encompass a wide variety of pathological processes with adverse cellular differentiation, proliferation and migration as common features. Pathologies may involve neovascular responses associated with diabetic retinopathy, retinopathy of prematurity or age-related macular degeneration. These diseases are quite prevalent and account for substantial visual impairment and blindness worldwide. Although treatment strategies are largely surgical, advances in our understanding of the proteins crucial to cell transdifferentiation, proliferation and migration, along with better gene transfer techniques, have greatly increased the potential for biological treatment options. In this report, the most common proliferative ocular vascular diseases and existing therapeutic modalities will be reviewed and an overview of possible gene therapy options will be discussed, along with potential candidate genes. PMID- 15268674 TI - Gene therapy approaches to prolonging corneal allograft survival. AB - Irreversible immunological rejection is the major cause of human corneal allograft failure and occurs despite the use of topical glucocorticoid immunosuppression. Systemic pharmacological interventions have not found widespread favour in corneal transplantation because of associated morbidities and inadequate demonstration of efficacy. Gene therapy offers tantalising prospects for improving corneal allograft survival, especially in those recipients at high risk of graft rejection. Donor corneas can be gene-modified ex vivo, while in storage prior to implantation, and the relative isolation of the transplanted cornea from the circulation decreases the risk of potential systemic complications. A wide variety of vectors have been found suitable for gene transfer to the cornea. The mechanisms involved in corneal graft rejection have been placed on a relatively secure footing over the past decade and in consequence a number of transgenes with promise for modulating rejection have been identified. However, relatively few studies have thus far demonstrated significant prolongation of corneal allograft survival after gene transfer to the donor cornea. In these instances, the therapeutic protein almost certainly acted at a proximal level in the afferent immune response, within the ocular environs. PMID- 15268675 TI - Gene therapy of liver diseases. AB - Many liver diseases lack satisfactory treatment and alternative therapeutic options are urgently needed. Gene therapy is a new mode of treatment for both inherited and acquired diseases, based on the transfer of genetic material to the tissues. Genes are incorporated into appropriate vectors in order to facilitate their entrance and function inside the target cells. Gene therapy vectors can be constructed on the basis of viral or non-viral molecular structures. Viral vectors are frequently used, due to their higher transduction efficiency. Both the type of vector and the expression cassette determine the duration, specificity and inducibility of gene expression. A considerable number of preclinical studies indicate that a great variety of liver diseases, including inherited metabolic defects, chronic viral hepatitis, liver cirrhosis and primary and metastatic liver cancer, are amenable to gene therapy. Gene transfer to the liver can also be used to convert this organ into a factory of secreted proteins needed to treat conditions that do not affect the liver itself. Clinical trials of gene therapy for the treatment of inherited diseases and liver cancer have been initiated but human gene therapy is still in its infancy. Recent progress in vector technology and imaging techniques, allowing in vivo assessment of gene expression, will facilitate the development of clinical applications of gene therapy. PMID- 15268676 TI - Recombinant adeno-associated virus vectors for gene therapy. AB - Recombinant adeno-associated virus (rAAV) vectors are based on a non-pathogenic human parvovirus (AAV) that is unique in its ability to persist in human cells without causing any pathologic effects. Studies of the potential barriers to rAAV mediated transduction of relatively resistant cells has led to an understanding of the mechanisms of cell attachment and entry, cytoplasmic translocation, nuclear entry, conversion to active double-stranded DNA, activation of transcription and establishment of persistent molecular forms. Each of these areas is individually discussed, as are recent applications in vivo in preclinical models and clinical trials. PMID- 15268677 TI - Intravenous, non-viral RNAi gene therapy of brain cancer. AB - RNA interference (RNAi) has the potential to knock down oncogenes in cancer, including brain cancer. However, the therapeutic potential of RNAi will not be realised until the rate-limiting step of delivery is solved. The development of RNA-based therapeutics is not practical, due to the instability of RNA in vivo. However, plasmid DNA can be engineered to express short hairpin RNA (shRNA), similar to endogenous microRNAs. Intravenous, non-viral RNAi-based gene therapy is enabled with a new gene-targeting technology, which encapsulates the plasmid DNA inside receptor-specific pegylated immunoliposomes (PILs). The feasibility of this RNAi approach was evaluated by showing it was possible to achieve a 90% knockdown of brain tumour-specific gene expression with a single intravenous injection in adult rats or mice with intracranial brain cancer. The survival of mice with intracranial human brain cancer was extended by nearly 90% with weekly intravenous injections of PILs carrying plasmid DNA expressing a shRNA directed against the human epidermal growth factor receptor. RNAi-based gene therapy can be coupled with gene therapy that replaces mutated tumour suppressor genes to build a polygenic approach to the gene therapy of cancer. PMID- 15268678 TI - Recombinant immunotoxins for the treatment of haematological malignancies. AB - Recombinant immunotoxins are fusion proteins which contain a ligand derived from the immune system fused to a toxin. The protein toxin is truncated to delete its binding domain, allowing selective ligand-directed binding. Growth factor fusion toxins are often considered immunotoxins. One of these molecules, containing the truncated diphtheria toxin and human IL-2 (Ontak), Ligand Pharmaceuticals), has been approved for the treatment of cutaneous T-cell lymphoma. Recombinant immunotoxins have also been produced containing the variable domains (Fv fragment) of monoclonal antibodies fused to toxins. These agents are relatively versatile with respect to the range of antigens possible. Several of these recombinant immunotoxins have showed clinical effectiveness in Phase I testing against haematological malignancies. One of these molecules, BL22, targets CD22 on hairy-cell leukaemia and has enabled patients to achieve complete remissions despite previous treatment and resistance to chemotherapy. PMID- 15268679 TI - Taking a Toll on human disease: Toll-like receptor 4 agonists as vaccine adjuvants and monotherapeutic agents. AB - Toll-like receptor (TLR) agonists are being developed for use as vaccine adjuvants and as stand-alone immunomodulators because of their ability to stimulate innate and adaptive immune responses. Among the most thoroughly studied TLR agonists are the lipid A molecules that target the TLR4 complex. One promising candidate, monophosphoryl lipid A, which is a derivative of lipid A from Salmonella minnesota, has proven to be safe and effective as a vaccine adjuvant in > 120,000 human doses. A new class of synthetic lipid A mimetics, the aminoalkyl glucosaminide 4-phosphates (AGPs), have been engineered specifically to target human TLR4 and are showing promise as vaccine adjuvants and as monotherapeutic agents capable of eliciting nonspecific protection against a wide range of infectious pathogens. In this review, the authors provide an update of the preclinical and clinical experiences with the TLR4 agonists, MPL (Corixa Corporation) adjuvant and the AGPs. PMID- 15268680 TI - Influenza virosomes as an efficient system for adjuvanted vaccine delivery. AB - Immunopotentiating reconstituted influenza virosomes possess several characteristics defining them as vaccine adjuvants. Virosomes have been shown to provide vaccine components with protection from extracellular degradation; a regular, repetitive antigen structure aiding presentation to B lymphocytes and fully functional, fusion-active, influenza haemagglutinin envelope proteins that enables receptor-mediated uptake and intracellular processing of the antigen. In addition, virosomes, as vaccine delivery systems, have been shown to be safe and not to engender any antibodies against the phospholipid components. Through the use of virosomes as a delivery vehicle, a number of vaccines have been developed. In humans, virosome-based vaccines containing inactivated hepatitis A and influenza antigens have been found to be efficacious and well-tolerated and have been on the market for several years. Hepatitis B, nucleic acids, cytotoxic drugs, and tetanus and diphtheria toxoids have also been incorporated into virosomes. Further investigations are ongoing in order to define the full potential of virosomes in both prophylactic and immunotherapeutic applications. PMID- 15268681 TI - Oxygen-carrying blood substitutes: a microvascular perspective. AB - Development of a viable blood substitute began by focusing on recreating the oxygen-carrying capacity of blood, leading to the recognition that haemoglobin (Hb) is presently unequalled for this function. However, as human Hb is the only realistic source of this protein, the production of a blood substitute that solves transfusional blood availability problems and shortages must introduce a multiplying factor between supply of natural blood and blood substitute, while maintaining equivalency of function/efficacy. In other words, a unit of blood should produce several units of equivalent blood substitute. This expansion is now possible because of new understanding of how blood delivers oxygen in the microcirculation and the consequences of reducing oxygen-carrying capacity in haemorrhage. This information is used to provide improved resuscitation capacity and maintenance of tissue metabolism by tailoring the properties of a blood substitute to the task of maintaining microvascular function, rather than oxygen delivery capacity. Resuscitation in an organism that is haemorrhaging requires maintenance perfusion, a process directly linked to the maintenance of adequate levels of shear stress on the endothelium, induced by either increased blood/plasma viscosity or increased blood flow velocity in the microcirculation. This process must also be intimately coupled with the requirement that no portion of the tissue is anoxic. This disparate set of requirements can be satisfied with high viscosity Hb solutions that have high affinity for oxygen, a combination of properties that causes the microcirculation to remain functional, and a requirement that supersedes restoration of oxygen-carrying capacity in the treatment of haemorrhage. PMID- 15268682 TI - Immunotherapy of ovarian cancer with antibodies: a focus on oregovomab. AB - Recent advances in the molecular and cellular biology of malignancy and tumour immunology have stimulated significant progress in the application of immunotherapies as adjuvant treatments in cancer. Oregovomab (OvaRex, AltaRex) is a murine monoclonal antibody with high affinity to the ovarian cancer associated antigen CA125. Infusion of low-dose antibody results in formation of circulating immune complexes which can trigger a cellular immune response targeting CA125 and the ovarian cancer. Oregovomab has activity following initial chemotherapy and in recurrent disease settings and is in Phase III trials to establish its efficacy to prolong time to relapse in patients with advanced ovarian cancer and favourable outcomes to their front-line treatment. Additional studies of antigen processing and combination chemo-immunotherapy are ongoing. The treatment shows promise as a potential new addition to the standard care of ovarian cancer. PMID- 15268683 TI - Enzyme replacement therapy for Fabry disease: lessons from two alpha galactosidase A orphan products and one FDA approval. AB - Two virtually identical products have been developed for enzyme replacement therapy for the treatment of Fabry disease, which is a rare and debilitating genetic disease caused by decreased activity of the lysosomal enzyme alpha galactosidase A. Lack of this enzyme results in progressive tissue accumulation of globotriaosylceramide (GL-3), resulting in life-threatening renal, cardiac and cerebrovascular complications. Both enzyme replacement products, agalsidase alfa (Replagal; Transkaryotic Therapies, Cambridge, MA, USA) and agalsidase beta (Fabrazyme; Genzyme Corporation, Cambridge, MA, USA), were approved by the European Agency for the Evaluation of Medicinal Products in 2001; agalsidase alfa at a recommended dose of 0.2 mg/kg and agalsidase beta at a recommended dose of 1 mg/kg. In the US, however, orphan drug laws dictated that only one of these products could be approved. In April 2003, after a rigorous evaluation of both products by the US FDA, this approval was granted to agalsidase beta. This decision reflected clinical trial design, how dosages were determined, antibody effects and the ability of each product to demonstrate either clinical efficacy or reduction of tissue storage of GL-3 in major organs of pathology when administered at the recommended dosage. The process also highlighted important issues in the evaluation of drugs to treat life-threatening genetic diseases for which the pathological basis is well-defined. PMID- 15268686 TI - Attitudes and concerns of French veterinarians towards pain and analgesia in dogs and cats. AB - OBJECTIVE: To compare the attitudes of French veterinarians to pain, and their provision of analgesia to animals, with that reported from other countries. STUDY DESIGN: Epidemiological study. METHODS: In June 1999, 379 French veterinarians were surveyed to ascertain their views on pain evaluation and control in dogs and cats, and their use of analgesics in daily practice. Survey results are expressed as a percentage of responses. RESULTS: The questionnaire was returned by 189 veterinarians (49.9%). The response rate was influenced by age (younger veterinarians were more likely to respond) but not gender. A majority (99.5%) expressed moderate to extreme concern over pain in their patients. Pain evaluation was based on the animal's attitude (88.3% dogs, 82.5% cats), interaction with the caregiver, response to palpation of the painful area (66.5% dogs, 62.7% cats) and inappetence (29.3% dogs, 46.3% cats, p < 0.001). Only 14.3% of respondents considered their knowledge of pain recognition to be inadequate. Many (58.8%) considered their methods of pain quantification and control (47% dogs, 59% cats) to be inadequate. Difficulties in recognizing pain (58.3%), a lack of knowledge in the appropriate use of analgesics (41.7%) and fear of drug side effects (30%) were used to explain inadequate provision of analgesia. Only 16.1 and 8.1% used opioids in dogs and cats, respectively. This low level of use resulted from the imposition of French narcotic legislation (79.9%) and lack of knowledge of opioid pharmacology (73.7%). Nonsteroidal anti-inflammatory drugs (NSAIDs) and glucocorticoids were the most commonly used analgesics in both species (100% (dogs) and 96.7% (cats)). The most popular NSAID used in France was tolfenamic acid, followed by meloxicam (dogs), ketoprofen, nimesulide (cats) and carprofen (dogs). The type of surgery performed influenced the use of analgesics, from 17.2% for castration to 83.7% for orthopaedic procedures. Nonsurgical conditions believed to warrant analgesia included osteoarthritis (97.8%), trauma (97.3%) and bone neoplasia (93.4%). Female veterinarians were more likely than males to evaluate pain and provide analgesia. CONCLUSION: French practitioners demonstrate a level of interest in analgesia, which appears to be at least equivalent to that reported from English-speaking countries. The signs used to indicate the presence of pain do not, in general, appear to differ. Excessive confidence in their ability to recognize pain (despite a general ignorance of the subject), the minor role of animal health technicians in pain management and misconceptions about analgesics (mainly opioids) are French pecularities. PMID- 15268687 TI - Comparison of preoperative carprofen and postoperative butorphanol as postsurgical analgesics in cats undergoing ovariohysterectomy. AB - OBJECTIVE: To compare carprofen to butorphanol, with regard to postsurgical analgesic effects, duration of analgesia, and adverse side effects. STUDY DESIGN: Blinded, randomized clinical study. ANIMALS: Seventy-one cats, 0.5-5 years of age, weighing 3.24 +/- 0.61 kg, undergoing ovariohysterectomy (OHE). METHODS: Cats were premedicated with subcutaneous atropine (0.04 mg kg(-1)), acepromazine (0.02 mg kg(-1)), and ketamine (5 mg kg(-1)). Anesthesia was induced with ketamine (5 mg kg(-1)) and diazepam (0.25 mg kg(-1)) given intravenously, and maintained with isoflurane. There were three treatment groups: group C (4 mg kg( 1) carprofen SC at induction), group B (0.4 mg kg(-1) butorphanol SC at end of surgery), and group S (0.08 mL kg(-1) of sterile saline SC at induction and end of surgery). Behavioral data were collected using a composite pain scale (CPS), prior to surgery (baseline) and 1, 2, 3, 4, 8, 12, 16, 20, and 24 hours post surgery. Interaction scores were analyzed separately. Cats with CPS scores >12 received rescue analgesia (meperidine, 4 mg kg(-1), intramuscular). RESULTS: Sixty cats completed the study. The CPS scores did not differ significantly between groups C and B at any time period. CPS scores for groups B and C were significantly increased for 12 hours post-surgery, and in group S for 20 hours. Both group C and B CPS scores were significantly lower than group S in this 20 hour postoperative period, except at 4 hours (B and C) and at 3 and 8 hours (B alone). Interaction scores for group C returned to preoperative baseline 4 hours after surgery, while both groups B and S remained increased for at least 24 hours post-surgery. Nine cats required meperidine. CONCLUSION: In this study, carprofen provided better postsurgical analgesia than butorphanol. Clinical relevance Neither drug completely abolished pain, however preoperative carprofen provided better pain control compared with postoperative butorphanol in the 24-hour period following OHE surgery in cats. PMID- 15268688 TI - Comparison of perioperative racemic methadone, levo-methadone and dextromoramide in cats using indicators of post-operative pain. AB - OBJECTIVE: To compare three opioid agonist drugs for perioperative analgesia in cats. STUDY DESIGN: Prospective, blind, controlled, randomised trial. ANIMALS: Ninety client-owned cats, weighing 3.1 (2.1-4.5) kg, aged 14.6 (6.0-84.0) months, were studied. METHODS: Seventy-six cats, scheduled for ovariectomy, received either 0.6 mg kg(-1) racemic methadone, 0.3 mg kg(-1) levo-methadone, 0.05 mg kg( 1) dextromoramide or a saline placebo IM. Behaviour and body position were assessed and scored 20 minutes later by a single 'blinded' observer. Anaesthesia was induced with propofol and maintained with halothane. Heart rate (HR), respiratory rate (RR), Fe'CO2 and SpO2 were recorded during anaesthesia. Post operatively, pain was categorised as absent, moderate or severe, on the basis of appearance, behaviour and response to palpation of the surgical wound (pain score). Appearance, pain scores and physiological variables were monitored every 30 minutes, for a duration of 4 hours. Differences between time-dependent continuous variables were analysed using mixed models for repeated measurements. Differences in categorical, time-dependent variables were analysed using chi2 tests. Significance was set at p < or = 0.05. RESULTS: There were no significant changes in appearance after pre-anaesthetic medication. After surgery, there was no association between appearance and pain score with HR or RR. The assessment of pain depended on comparison with the placebo group, by comparing animals' reactions to wound palpation. Sixteen of the 18 cats in the placebo group and 14 of the 19 cats in the dextromoramide group showed signs of moderate-to-severe pain after surgery. In the levo-methadone group (n = 20), one animal showed pain after 60 minutes and two after 120 minutes. One cat in the racemic methadone group (n = 19) showed pain signs and behavioural changes at 60 minutes. Compared to the two methadone groups, 'rescue' analgesia was required more often in cats treated with dextromoramide or saline. CONCLUSION AND CLINICAL RELEVANCE: Dextromoramide (0.05 mg kg(-1)) was ineffective, while racemic methadone (0.6 mg kg(-1)) and levo-methadone (0.3 mg kg(-1)) provided effective analgesia in cats following ovariectomy, without behavioural, respiratory or cardiovascular side effects. PMID- 15268689 TI - Effect of transdermally administered fentanyl on the minimum alveolar concentration of isoflurane in cats. AB - OBJECTIVE: To determine the effect of two doses of fentanyl, administered transdermally, on the minimum alveolar concentration (MAC) of isoflurane in cats. STUDY DESIGN: Prospective, randomized study. ANIMALS: Five healthy, spayed, female cats. METHODS: Each cat was studied thrice with at least 2 weeks between each study. In study 1, the baseline isoflurane MAC was determined in triplicate for each cat. In studies 2 and 3, isoflurane MAC was determined 24 hours after placement of either a 25 or 50 microg hour(-1) fentanyl patch. In each MAC study, cats were instrumented to allow collection of arterial blood and measurement of arterial blood pressure. Twenty-four hours prior to studies 2 and 3, a catheter was placed and secured in the jugular vein and either a 25 or 50 microg hour(-1) fentanyl patch was placed in random order on the left thorax. Blood samples for plasma fentanyl determination were collected prior to patch placement and at regular intervals up to 144 hours. After determination of MAC in studies 2 and 3, naloxone was administered as a bolus dose (0.1 mg kg(-1)) followed by an infusion (1 mg kg(-1) hour(-1)) and MAC redetermined. RESULTS: The baseline isoflurane MAC was 1.51 +/- 0.21% (mean +/- SD). Fentanyl (25 and 50 micro g hour(-1)) administered transdermally significantly reduced MAC to 1.25 +/- 0.26 and 1.22 +/ 0.16%, respectively. These MAC reductions were not significantly different from each other. Isoflurane MAC determined during administration of fentanyl 25 micro g hour(-1) and naloxone (1.44 +/- 0.16%) and fentanyl 50 micro g hour(-1) and naloxone (1.51 +/- 0.19%) was not significantly different from baseline MAC (1.51 +/- 0.21%). CONCLUSIONS AND CLINICAL RELEVANCE: Fentanyl patches are placed to provide long-lasting analgesia. In order to be effective postoperatively, fentanyl patches must be placed prior to surgery. Plasma fentanyl concentrations achieved intraoperatively decrease the need for potent inhalant anesthetics in cats. PMID- 15268690 TI - Distribution of new methylene blue injected into the lumbosacral epidural space in cats. AB - OBJECTIVE: To determine the effects of injection volume and vertebral anatomy on the spread of new methylene blue (NMB) injected into the lumbosacral epidural space in cats. STUDY DESIGN: Prospective experimental study. SAMPLE POPULATION: Sixteen cats. METHODS: Cats were randomly assigned to four groups and received from 0.1 to 0.4 mL kg(-1) of 0.12% NMB in 0.9% saline. Injection was made into the lumbosacral epidural space using a dorsal approach with the cats in sternal recumbency. The extent of cranial migration of the dye as indicated by the staining of epidural fat and dura mater was measured. RESULTS: The mean +/- SD (range) number of stained vertebrae in the 0.3 and 0.4 mL kg(-1) groups, were 11.5 +/- 1.5 (T7-T11) and 12.4 +/- 1.8 (T6-T10), respectively. This was significantly greater than the number in the 0.1 and 0.2 mL kg(-1) groups, 4.3 +/ 0.6 (L3-L4) and 6.0 +/- 0.7 (L1-L2) vertebrae, respectively (p < 0.001). Linear regression analysis showed that the volume injected correlated significantly with the number of stained vertebrae (R2 = 0.83, p < 0.001). In the dorsal and lateral aspect of the spinal cord, NMB solution distributed between epidural fat and dura mater. Migration under the spinal cord occurred along the two longitudinal epidural veins. CONCLUSIONS AND CLINICAL RELEVANCE: The larger the volume of solution injected into the lumbosacral epidural space in cats, the greater the spread. PMID- 15268691 TI - Sedative, analgesic and cardiorespiratory effects of romifidine in cats. AB - OBJECTIVE: To evaluate the sedative, analgesic, and cardiorespiratory effects of intramascular (IM) romifidine in cats. STUDY DESIGN: Prospective, randomized experimental trial. ANIMALS: Ten healthy adult cats. METHODS: Romifidine (100, 200, and 400 microg kg(-1)) or xylazine (1 mg kg(-1)) was given IM in a cross over study design. Heart rate (HR), respiratory rate (RR), rectal temperature (RT), hemoglobin saturation, oscillometric arterial pressure, and scores for sedation, muscle relaxation, position, auditory response, and analgesia were determined before and after drug administration. Time to recumbency, duration of recumbency, and time to recover from sedation were determined. Subjective evaluation and cardiorespiratory variables were recorded before and at regular intervals for 60 minutes after drug administration. RESULTS: Bradycardia developed in all cats that were given romifidine or xylazine. No other significant differences in physiologic parameters were observed from baseline values or between treatments. Increasing the dose of romifidine did not result in increased sedation or muscle relaxation. Cats given xylazine showed higher sedation and muscle relaxation scores over time. Analgesia scores were significantly higher after administration of romifidine (400 microg kg(-1)) and xylazine (1 mg kg(-1)) than after romifidine at 100 or 200 microg kg(-1). Duration of lateral recumbency was not significantly different between treatments; however, cats took longer to recover after administration of 400 micro g kg(-1) romifidine. CONCLUSIONS AND CLINICAL RELEVANCE: Bradycardia is the most important adverse effect after IM administration of romifidine at doses ranging from 100 to 400 microg kg(-1) or 1 mg kg(-1) of xylazine in cats. The sedative effects of romifidine at 200 microg kg(-1) are comparable to those of 1 mg kg(-1) of xylazine, although muscle relaxation and analgesia were significantly less with romifidine than with xylazine. PMID- 15268692 TI - A clinical evaluation of the 'mini parallel Lack' breathing system in cats and comparison with a modified Ayre's T-piece. AB - OBJECTIVE: To evaluate the suitability of a 'mini parallel Lack' (MPL) breathing system for use in spontaneously breathing cats and to compare the fresh gas flow requirement with that of a modified Ayre's T-piece (MATP). ANIMALS: Twenty client owned cats, ASA I and II, presented for elective procedures requiring anaesthesia. MATERIALS AND METHODS: Pre-anaesthetic medication and induction of anaesthesia were carried out using several techniques commonly used in our teaching hospital. Anaesthesia was maintained with halothane or isoflurane vaporized in either oxygen or with a mixture of oxygen and nitrous oxide. Both breathing systems were evaluated in each cat, with the order of use randomized. Initial fresh gas flows were 300 mL kg(-1) minute(-1) for the MPL and 500 mL kg(- 1) minute(-1) for the MATP. After a 20-minute stabilization period, fresh gas flow was reduced by 200 mL minute(-1) every 5 minutes until re-breathing--defined as an increase in the inspired partial pressure of carbon dioxide to 0.3 kPa (2 mm Hg)--was detected. The fresh gas flow was then increased in 100 mL minute(-1) increments until re-breathing was no longer detectable, and this value was recorded as the minimum fresh gas flow requirement for the breathing system in use. The procedure was then repeated for the second breathing system. Minimum fresh gas flow requirements were compared using a paired Students t-test. Cardiopulmonary variables were compared using anova. Valve opening pressure was measured in the MPL using a manometer. RESULTS: The mean (+/-SD) fresh gas flow that prevented re-breathing with the MPL (510 +/- 170 mL minute(-1); equivalent to 142 +/- 47 mL kg(-1) minute(-1)) was significantly lower than that required for the MATP (1430 +/- 560 mL minute(-1); equivalent to 397 +/-155 mL kg(-1) minute(-1)). There were no significant differences in cardiopulmonary variables attributable to the use of the two breathing systems. The MPL valve opening pressure was 1.1 cm H2O. CONCLUSIONS: The MPL breathing system used lower gas flows than the MATP without affecting cardiovascular or respiratory function. Clinical relevance In spontaneously breathing cats, the MPL offers the advantages of a reduction in cost and atmospheric pollution because less volatile agent is vaporized. PMID- 15268693 TI - Evaluation of laryngeal mask as an alternative to endotracheal intubation in cats anesthetized under spontaneous or controlled ventilation. AB - OBJECTIVE: To compare the cardiorespiratory effects and incidence of gastroesophageal reflux with the use of a laryngeal mask airway (LMA) or endotracheal tube (ET) in anesthetized cats during spontaneous (SV) or controlled ventilation (CV). STUDY DESIGN: Prospective randomized experimental trial. ANIMALS: Thirty-two adult crossbred cats, weighing 2.7 +/- 0.4 kg. METHODS: The cats were sedated with intramuscular (IM) methotrimeprazine (0.5 mg kg(-1)) and buprenorphine (0.005 mg kg(-1)), followed 30 minutes later by induction of anesthesia with intravenous (IV) thiopental (12.5-20 mg kg(-1)). An ET was used in 16 cats and an LMA in the remaining 16 animals. Anesthesia was maintained with 0.5 minimum alveolar concentration (0.6%) of halothane in oxygen using a Mapleson D breathing system. Cats in both groups were further divided into two equal groups (n = 8), undergoing either SV or CV. Neuromuscular blockade with pancuronium (0.06 mg kg(-1)) was used to facilitate CV. Heart and respiratory rates, direct arterial blood pressure, capnometry (PE'CO2) and arterial blood gases were measured. Gastric reflux and possible aspiration was investigated by intragastric administration of 5 mL of radiographic contrast immediately after induction of anesthesia. Cervical and thoracic radiographs were taken at the end of anesthesia. Data were analyzed using anova followed by Student-Newman-Keuls, Kruskal-Wallis or Friedman test where appropriate. RESULTS: Values for PaCO2 and PE'CO2 were higher in spontaneously breathing cats with the LMA when compared with other groups. Values of PaO2 and hemoglobin oxygen saturation did not differ between groups. Gastroesophageal reflux occurred in four of eight and two of eight cats undergoing CV with ET or LMA, respectively. There was no tracheal or pulmonary aspiration in any cases. CONCLUSIONS AND CLINICAL RELEVANCE: The use of an LMA may be used as an alternative to endotracheal intubation in anesthetized cats. Although aspiration was not observed, gastric reflux may occur in mechanically ventilated animals. PMID- 15268694 TI - Effects of tiletamine/zolazepam-romifidine-atropine in ocelots (Leopardus pardalis). AB - OBJECTIVES: To evaluate the effects of a combination of tiletamine-zolazepam romifidine-atropine in ocelots. DESIGN: Prospective experimental trial. ANIMALS: Eight captive adult ocelots (three females and five males). METHODS: Calculated doses of tiletamine-zolazepam (3.75 mg kg(-1)), romifidine (50 microg kg(-1)) and atropine (0.04 mg kg(-1)) were administered intramuscularly. After immobilization, animals were weighed and the real doses determined. Heart rate, respiratory frequency, noninvasive systolic, diastolic, and mean arterial pressure, arterial oxygen hemoglobin saturation, and rectal temperature were measured. Data were analyzed by means of anova for repeated measures, followed by the Tukey test to compare values over time. RESULTS: Doses administered were 3.4 +/- 0.6 mg kg(-1) of tiletamine-zolazepam, 0.04 +/- 7.0 mg kg(-1) of romifidine, and 0.03 +/- 0.007 mg kg(-1) of atropine. The mean time to recumbency and duration of immobilization were 7.0 +/- 4.5 and 109.2 +/- 27.9 minutes, respectively. The median times to standing and walking were 52.3 [0-90] and 2.3 [0-69.3] minutes, respectively. A decrease in heart rate was observed 45 minutes following drug administration. Arterial blood pressure was maintained during the study. CONCLUSIONS AND CLINICAL RELEVANCE: This protocol produced good immobilization in ocelots with minimal changes over time in cardiovascular parameters. PMID- 15268695 TI - Unstable endobronchial intubation in a cat undergoing tracheal laceration repair. AB - A peri-carinal tracheal laceration was produced in a 11-year-old cat during tracheal intubation. Before reconstructive surgery began, the leak was bypassed with an endobronchial tube positioned using endoscopy and direct vision. However, single-lung ventilation could not be sustained because the tube became dislodged and could not be repositioned. Consequently, surgery was completed with periods of intermittent apnoea interspersed with manually controlled hyperventilation. Cardiovascular variables were stable during anaesthesia and no signs of hypoxia were detected. The difficulties in maintaining endobronchial tube position resulted from the animal's small size relative to the dimensions of the endotracheal tube. PMID- 15268696 TI - Anesthetic equipment fault leading to hypercapnia in a cat. AB - Spontaneous ventilation during positive pressure ventilation was observed in a 4 year-old DSH cat maintained under general anesthesia with isoflurane delivered with a nonrebreathing system. This was accompanied by an increase in heart rate and blood pressure. On investigation, neither an inadequate plane of anesthesia, nor hypoxemia, nor hyperthermia was present. The nonrebreathing system was replaced and the hypercapnia resolved. A defect in the inner fresh gas delivery tube of the Bain system was identified. A simple and quick test is described which can be performed to verify the integrity of the inner tube of the Bain breathing circuit. PMID- 15268697 TI - Effects of propofol on the electrocardiogram and systolic blood pressure of healthy cats pre-medicated with acepromazine. AB - OBJECTIVE: To obtain and analyze the electrocardiogram and systolic blood pressure of cats before, during, and after a continuous infusion of propofol. STUDY DESIGN: Prospective, uncontrolled experimental trial. ANIMALS: Twenty healthy adult crossbred male and female cats aged between 3 and 5 years, weighing 2.8-5.0 kg (mean 3.9 kg). METHODS: Cats were pre-medicated with acepromazine 0.1 mg kg(-1) subcutaneously and anesthesia was induced with intravenous (IV) propofol 6 mg kg(-1) and maintained with a continuous infusion of propofol at 0.5 mg kg(-1) minute(-1) for 60 minutes. Electrocardiographic parameters and systolic blood pressure obtained by Doppler ultrasound were recorded before pre-medication (T0), 30 (T30), and 60 (T60) minutes after beginning the continuous infusion, and 30 minutes after its cessation (T90). Repeated measures anova was used to perform statistical analysis. RESULTS: A significant decrease in heart rate was observed at all time points when compared with T0 values. The PR interval increased significantly at T60 and T90. Systolic blood pressures during anesthesia were significantly lower than at T0 and T90. CONCLUSION AND CLINICAL RELEVANCE: The changes seen were not clinically important in normal cats but given the reduction in heart rate and systolic blood pressure, careful consideration should be given before using this technique in patients in which hypotension or a reduction in heart rate would be poorly tolerated. PMID- 15268698 TI - Inclusive intake screening: shaping medical problems into specialist-appropriate cases. AB - This paper examines medical intake screening through the process of making appointments with medical specialists. By employing a multi-method, qualitative approach, it shows how decisions to schedule doctors' appointments are based on medical knowledge about physicians' specialties and specific organisational practices. It draws on insights from first-contact interactions between clients and institutional gatekeepers to enrich our understanding of intake screening. In relation to gatekeeping, rationing commonly gets framed as restrictive screening practices, with a preference for denying or limiting access to treatment. Restrictive screening practices are typically organised to elicit a narrow range of information ('facts') relevant to specific eligibility criteria; whereas inclusive intake screening tends to involve less scripted, more complex and open ended interactional exchanges between workers and clients, wherein workers help clients frame their claims in ways that will increase their chances of getting accepted. Front-office workers hold a preference for inclusive intake screening, a preference that is undergirded by the referral-driven nature of this stage of patient processing, and by a work environment that favours inclusive screening. This finding builds on the literature within medical sociology, but also extends our understanding of frontline decision-making and the distribution of resources within a variety of people-processing institutions. PMID- 15268699 TI - Accommodating health and social care needs: routine resource allocation in stroke rehabilitation. AB - This paper explores routine resource allocation processes in health and social care. While there has been a small body of work which has drawn on Lipsky's (1980) insights into street level bureaucracy, few have taken seriously the opportunity offered by ethnography to explore in detail the work of front-line staff as a way of observing policy processes in action. Utilising ethnographic data from research into the continuing care of adults who had suffered a first acute stroke, we analyse how staff accommodated patient need and consider the implications that this had for the quality, equality and equity of service provision. PMID- 15268700 TI - Contextualising experiences of depression in women from South Asian communities: a discursive approach. AB - The aim of this paper is to present an interpretation of the accounts of depression provided by women from South Asian communities. The paper presents the findings from a qualitative study, conducted in the UK, which explored women from South Asian communities and their experiences of depression. It is argued here, through examples of women's accounts of their experiences, that depression is 'embodied', that is, grounded in the materiality of the body which is also immersed in subjective experiences and in the social context of women's lives. Qualitative data were collected from four focus groups and ten individual interviews with women. The analysis involved a discursive approach. Analysis revealed how women made strategic choices in how they presented their symptoms as legitimate and for gaining access to what they perceived to be appropriate healthcare. This is not to argue that this is a culturally specific phenomenon but one which is a feature of all healthcare negotiations. PMID- 15268701 TI - Beyond medicalization-healthicization? A rejoinder to Hislop and Arber. PMID- 15268703 TI - Body Worlds: clinical detachment and anatomical awe. AB - If studying anatomy in medical school promotes clinical detachment, how do lay people respond to the crash course in anatomy they receive on visiting the Korperwelten / Body Worlds exhibition? If late modernity's celebration of the living body makes the dead body problematic, how do visitors respond to the aestheticised dead bodies on display? Through examining the written comments of visitors, the article identifies a number of responses. The chief is an elementary scientific gaze in which obvious interest is shown in anatomical details. But because the exhibits are dry, odourless and anonymous, this does not generate the defence of emotional detachment; indeed, among several emotional responses, are fascination and, for some, awe. Body Worlds is less a popularised anatomy lab than a shrine to the human body, a shrine in which medically untrained people can look at the body in new ways. PMID- 15268705 TI - What to call the LEJC-BFV nevus? PMID- 15268706 TI - Variability in nomenclature used for nevi with architectural disorder and cytologic atypia (microscopically dysplastic nevi) by dermatologists and dermatopathologists. AB - BACKGROUND: Although a nevus with the microscopic features of a "dysplastic nevus" is commonly seen, the nomenclature used to describe such a lesion has been thought to be inconsistent. A 1992 National Institutes of Health (NIH) Consensus Conference sought to unify nomenclature and suggested that the term "nevus with architectural disorder" be used along with a comment on melanocytic atypia. METHODS: We performed a cross-sectional mail survey to determine preferred terminology as well as the level of adherence to the NIH-recommended nomenclature. All 856 active members of the American Society of Dermatopathology (ASDP) and 1100 (13.0%) of the 8471 active members of the American Academy of Dermatology (AAD) were surveyed. RESULTS: Five hundred and thirty-three ASDP members and 483 AAD members who fulfilled eligibility criteria completed the questionnaire. The term "dysplastic nevus" was favored by the largest number of responders (favored by 39.1% of ASDP members and 62.3% of AAD members), while the 1992 NIH Consensus Conference-recommended terminology was the second most popular term (25.3% of ASDP and 15.1% of AAD members). Dermatopathologists (OR = 1.9, p = 0.0001) and those who had dual training in dermatology and dermatopathology (OR = 1.6, p = 0.02 for ASDP members; OR = 2.3, p = 0.02 for AAD members) were more likely to adhere to the 1992 NIH Consensus Conference nomenclature. CONCLUSIONS: Despite attempts to unify nomenclature for microscopically dysplastic nevi through the NIH Consensus Conference, wide variation in terminology persists. PMID- 15268707 TI - Pityriasis lichenoides: a cytotoxic T-cell-mediated skin disorder. Evidence of human parvovirus B19 DNA in nine cases. AB - BACKGROUND: Pityriasis lichenoides et varioliformis acuta (PLEVA) and pityriasis lichenoides chronica (PLC) probably represent the polar ends of the same pathologic process, i.e. pityriasis lichenoides (PL), with intermediate forms in between. Previous studies have demonstrated that the inflammatory infiltrate in PLEVA is composed of cytotoxic suppressor T cells, whereas in PLC the helper/inducer T-cell population drives the immunological answer. Furthermore, monoclonal rearrangement of the T-cell receptor-gamma (TCR-gamma) genes was repeatedly found both in PLEVA and PLC. METHODS: Forty-one formalin-fixed, paraffin-embedded tissue specimens of 40 cases of PL were retrieved from the files of the authors. Immunophenotyping for cytotoxic granular proteins (Tia 1/GMP-17 and Granzyme B) and T-cell-related antigens (n = 41), TCR-gamma chain gene analysis (n = 30) and molecular investigations for parvovirus B19 (PVB19) DNA (n = 30) were performed. RESULTS: Overlapping immunophenotypes were observed in PLEVA and PLC. The dermal and epidermal T cells predominantly expressed CD2, CD3, CD8, and Tia-1 with a variable positivity for CD45RA, CD45RO, and Granzyme B. A monoclonal rearrangement pattern of the TCR-gamma genes was detected in three cases (10%). PVB19 DNA was found in nine cases (30%). T-cell monoclonality in conjunction with genomic PVB19 DNA was present in one case. CONCLUSIONS: Our results demonstrate that PL is a skin disorder mediated by the effector cytotoxic T-cell population. The identification of PVB19 DNA in nine cases may be interpreted ambiguously: PVB19 as a true pathogen or as an innocent bystander. PMID- 15268708 TI - Cutaneous horns of the eyelid: a clinicopathological study of 48 cases. AB - BACKGROUND: Cutaneous horn (cornu cutaneum) is a morphological designation for a protuberant mass of keratin that resembles the horn of an animal. It results from unusual cohesiveness of keratinized material from the superficial layers of the skin or implanted deeply in the cutis. This lesion may be associated with a benign, premalignant, or malignant lesion at the base, masking numerous conditions. METHODS: A retrospective analysis of 48 cases of cutaneous horns of the eyelid treated between 1992 and 2002 has been performed. RESULTS: Twenty-four men and 19 women, with a mean age of 62 years (range 16-90), were treated by surgery. Histologically, 77.1% were associated with benign specimens at the base pathology, 14.6% were premalignant, and finally, 8.3% were caused by malignant skin tumors. The most common lesion was seborrheic keratosis among the benign lesions, actinic keratosis among the premalignant ones, and basal cell carcinoma and squamous cell carcinoma among the malignant ones. CONCLUSION: Cutaneous horns usually appear on exposed skin areas in elderly men. The important issue in this condition is not the horn itself, which is just dead keratin, but rather the nature of the underlying disease, although the horns are usually benign. PMID- 15268709 TI - Expression profiles of p63, p53, survivin, and hTERT in skin tumors. AB - BACKGROUND: p63 is a p53 homolog and a marker expressed in replicating keratinocytes. Survivin is a recently characterized inhibitor of apoptosis protein that is abundantly expressed in most solid and hematologic malignancies. Telomerase reverse transcriptase (TERT) is the major determinant of human telomerase activity, and its expression is indicative of unlimited replication. We herein evaluated the expression profiles of p63, p53, survivin, and hTERT in usual skin cancers, including squamous cell carcinoma (SCC) and basal cell carcinoma (BCC) and putative preneoplastic epidermal lesions, including actinic keratosis (AK), Bowen's disease, and porokeratosis. METHODS: Immunohistochemistry using antibodies against p63, p53, survivin, and hTERT was performed. Semi quantitative evaluation (-, +, 2+, 3+) was carried out. RESULTS: BCCs showed diffuse p63 expression and SCCs heterogeneous p63 expression with negativity in terminally differentiated squamous cells. All preneoplastic epidermal lesions showed p63 expression in all cell layers. p53 was found in seven of 10 cases of BCCs, all 10 cases of SCCs, and nine of 10 cases of Bowen's disease. AK and porokeratosis revealed focal to moderate p53 expression. Survivin was found in eight of 10 cases of SCCs and eight of 10 cases of Bowen's disease. Six of 10 cases of BCCs revealed weak survivin positivity. AK and porokeratosis showed survivin expression confined to the basal layer. hTERT expression was found in most cases of skin cancers and preneoplastic lesions. CONCLUSIONS: p63 expression may be a marker of basal/progenitor cells and a diagnostic marker in skin tumors. p63 expression is not related to p53 expression in these tumors. This study points to a putative role of survivin and hTERT in the development of certain skin cancers. In addition, our data support the concept of porokeratosis being a premalignant condition. PMID- 15268710 TI - Proliferative characterization of basal-cell carcinoma and trichoepithelioma in small biopsy specimens. AB - We examined the proliferative characteristics of 20 basal-cell carcinomas (BCCs) and 16 trichoepitheliomas (TEps) in an effort to understand and explore possible differences in their tumorigenic cell-cycle properties. These tumors were first compared for their expression of the nuclear proliferative protein Ki-67 and the tumor suppressor protein p53. We also compared the p53 downstream effector, p21(waf-1/cip-1), an inhibitor of cyclin-dependent kinases. The other p53 dependent, cyclin-dependent kinase inhibitor, p27(kip-1), has shown to be increased in TEps, which is consistent with this benign neoplasm's better differentiated state. In our findings, we confirmed through immunohistochemical staining for Ki-67 that BCCs qualitatively showed a greater proliferative fraction compared to TEps (50.0 vs. 13.0%, p < 0.00001) as well as over expression of p53 (2+ vs. 1+, p < 0.0008). BCCs marked by p21 demonstrated scattered nuclear positivity compared to the virtual absence of staining in the TEps (p < 0.019). In studying their cell-cycle properties, our findings suggest that abnormalities in the p53 pathway allow BCCs to obtain a growth advantage. We show that Ki-67 and p53 staining both appear useful in resolving challenging differential diagnoses and thereby help in directing appropriate treatment strategies. PMID- 15268711 TI - Extramedullary hematopoiesis in a pyogenic granuloma: a case report and review. AB - BACKGROUND: Extramedullary hematopoiesis in the adult life is usually associated with hematological disorders. It can often occur in several organs under certain pathological conditions, including the spleen, liver, and skin, which are normal sites during the embryogenesis, the skin being very uncommon. CASE REPORT: A case of extramedullary hematopoiesis in a granuloma pyogenicum at the anterior aspect of the right thigh of a 31-year-old Japanese man is reported. RESULTS: Histologic examination revealed an ulcerated polypoid lesion composed of numerous newly formed capillaries and a myxoid edematous stroma. In addition, three islands of hematopoietic precursors were found. CONCLUSIONS: This is, to our knowledge, the second case of extramedullary hematopoiesis in a granuloma pyogenicum. We believe that vascular lesions have potential to generate hematopoietic precursor cells; however, hematopoiesis will only occur in a setting of adequate microenvironment, as it occurs in the bone marrow. PMID- 15268712 TI - Pilomatricoma with a bullous appearance. AB - BACKGROUND: Pilomatricoma is a benign, cutaneous neoplasm with differentiation toward hair matrix. The tumor is usually a deep-seated, solitary, firm nodule with overlying normal epidermis. Pilomatricoma with a bullous appearance is very rare. METHODS: A 16-year-old Chinese girl with a soft, purplish, translucent bulla on the left shoulder for 6 months and a nodule underlying the bulla is described. RESULTS: The histopathologic findings were consistent with pilomatricoma. There was extraordinary dilation of lymphatic vessels in the overlying dermis, which explains the clinical bullous appearance. The surrounding dermis had an edematous appearance. A Verhoeff-van Gieson stain disclosed the marked reduction of the elastic fibers, and an alcian blue stain was negative. CONCLUSIONS: The patient is diagnosed as bullous variant of pilomatricoma. PMID- 15268713 TI - Hidradenoma papilliferum with mixed histopathologic features of syringocystadenoma papilliferum and anogenital mammary-like glands. AB - A case of hidradenoma papilliferum with mixed features of syringocystadenoma papilliferum (SCAP) and anogenital mammary-like glands is reported. A single, fresh red-colored nodule developed in the sulcus between the labia majora and minora of a 49-year-old Japanese woman. Histopathologically, the tumor showed epithelial lining with apocrine secretion and slight connective tissues characteristics. Our case was unique because, like SCAP, the tumor was connected to the epidermis and cystic invaginations extended downward into the deep dermis. In addition, beneath the tumor, tubular structures that resembled normal mammary tissue were present in the subcutaneous fatty tissue. In this study, it has been suggested that this tumor might have been developed from these mammary-like glands. PMID- 15268714 TI - Pseudotumoral encapsulated fat necrosis with diffuse pseudomembranous degeneration. AB - An extraordinary case of encapsulated fat necrosis characterized by its large size, diffuse formation of pseudomembranes, and tendency to recur after excision is reported. A 67-year-old Caucasian woman suffering from morbid obesity was admitted for diagnosis and surgical treatment of a soft tissue mass showing a longest diameter of 14 cm and lying adjacently to the scar from previous appendicectomy. Histopathologic features were consistent with a nodular-cystic encapsulated fat necrosis with diffuse pseudomembranous transformation. Eight months after surgery, a new larger mass (longest diameter of 18 cm) sharing identical histopathologic features appeared in the same location. Encapsulated fat necrosis is a well-defined entity even though several names have been proposed for this condition, including mobile encapsulated lipoma, encapsulated necrosis, or nodular-cystic fat necrosis. Its pathogenesis seems to be related to ischemic changes secondary to previous trauma. It may occasionally show degenerative changes, including dystrophic calcifications and presence of pseudomembranes. To our knowledge, these are the first reported cases of encapsulated fat necrosis presenting as lesions of such size and showing diffuse formation of pseudomembranes; these particular features made diagnosis difficult and led to consideration of a wide range of potential diagnostic possibilities. This case expands the clinico-pathologic spectrum of membranocystic fat necrosis, including the potential ability of this subcutaneous fatty tissue abnormality to recur after surgical excision. Felipo F, Vaquero M, del Agua C. Pseudotumoral encapsulated fat necrosis with diffuse pseudomembranous degeneration. PMID- 15268715 TI - Cellular neurothekeoma with histiocytic differentiation. AB - BACKGROUND: It is generally accepted that the two types of neurothekeoma (myxoid type and cellular type) represent the two poles of a spectrum. This concept, however, has recently been challenged, and cellular neurothekeomas have been suggested as a separate classification and are included in the "fibrohistiocytic" category by some authors. Cellular neurothekeomas have been reported to show negative immunohistochemical staining for histiocytic markers, and PG-M1 is now considered to be the most reliable histiocytic marker. CASE REPORT: We report a case of cellular neurothekeoma. The histopathological features in this case were typical for cellular neurothekeoma. Immunohistochemically, the neoplastic cells were diffusely positive for S-100A6 protein, PGP9.5, CD10, CD68 (KP1), PG-M1, and Vimentin, and negative for other antibodies including S-100 protein and factor XIIIa. CONCLUSIONS: Cellular neurothekeoma expressing both KP-1 and PG-M1 is considered to show histiocytic differentiation, and may be interpreted as a neoplasm with immature nerve sheath differentiation, incidentally expressing histiocytic markers, or as an undifferentiated neoplasm derived from the neural crest cells of nerve sheath/fibrohistiocyte lineage. These results, such as the concomitant expressions of PGP9.5/S-100A6 and PG-M1/CD68 (KP-1), support the theory of multiple differentiation in cellular neurothekeomas. The significance of the expression of CD10 in this cellular neurothekeoma is unclear. PMID- 15268716 TI - Cutaneous lobular capillary hemangioma induced by pregnancy. PMID- 15268719 TI - Living lobar lung transplantation: is it a necessary option? PMID- 15268720 TI - Long-term renal allograft survival: a cup both half-full and half-empty. PMID- 15268721 TI - Screening for West Nile virus: more uncertainty. PMID- 15268722 TI - Cytomegalovirus and lung transplantation. AB - Cytomegalovirus (CMV) infection remains a serious problem in lung transplant recipients. Development of potent oral antiviral agents, molecular techniques for the detection of infection and its response to therapy and the emergence of isolates with antiviral resistance have had significant impacts on the approach to CMV in these patients. This article discusses the following issues as part of a comprehensive CMV management strategy in lung transplant recipients: (1) Prevention strategies in the era of potent oral antiviral agents, (2) the role of new diagnostic techniques in the management of CMV, (3) treatment regimens for established CMV infection or disease, (4) the potential impact of treatment of CMV on the indirect effects on long-term allograft function, and (5) the incidence, risk factors for and impact of ganciclovir resistance following lung transplantation. PMID- 15268723 TI - RNA interference: a potent tool for gene-specific therapeutics. AB - RNA interference (RNAi) is a process through which double-stranded RNA induces the activation of cellular pathways, leading to potent and selective silencing of genes with homology to the double strand. Much excitement surrounding small interfering RNA (siRNA)-mediated therapeutics arises from the fact that this approach overcomes many of the shortcomings previously experienced with approaches such as antibodies, antisense oligonucleotides and pharmacological inhibitors. Induction of RNAi through administration of siRNA has been successfully used in treatment of hepatitis, viral infections, and cancer. In this review we will present a brief history of RNAi, methods of inducing RNAi, application of RNAi in the therapeutic setting, and the possibilities of using this highly promising approach in the context of transplantation. PMID- 15268724 TI - Alloantibody production is regulated by CD4+ T cells' alloreactive pathway, rather than precursor frequency or Th1/Th2 differentiation. AB - Although CD4(+) T cells play an important role in the regulation of allograft rejection, the exact mechanisms by which they operate and the actual contribution of direct and indirect alloreactivity pathways remain to be fully characterized. Previous studies have established a possible relationship between the indirect alloreactivity pathway and antibody production, but interpretation of these results have been complicated by shortcomings inherent to the models used in these studies. To address this issue, we have developed a model based on TCR transgenic mice derived from a CD4(+) T-cell clone which recognize specific alloantigens by both alloreactivity pathways. Skin allografts on alphabeta T-cell deficient mice adoptively transferred with transgenic CD4(+) T cells were rejected without significant delay between the two alloreactivity pathways. No IgG alloantibody was produced following allograft rejection by the direct alloreactivity pathway alone. Importantly, production of antibodies against alloantigens of the direct pathway was shown to require help from CD4(+) T cells activated by the indirect pathway. These results indicate that the events leading to the initiation of immune responses responsible for graft rejection are clearly dependent on the population of antigen-presenting cells involved in T- and B lymphocyte activation. PMID- 15268725 TI - Caspase inhibition prevents the increase in caspase-3, -2, -8 and -9 activity and apoptosis in the cold ischemic mouse kidney. AB - Prolonged cold ischemic time is a risk factor for the development of delayed graft function. The adverse impact of cold ischemia may be associated with tubular cell death in the kidney. Caspase-3 is a major mediator of apoptotic cell death. We hypothesized that caspase inhibition would reduce apoptosis and other features of cold ischemia. Kidneys of C57BL/6 mice were perfused with cold University of Wisconsin solution containing a pancaspase inhibitor or vehicle via the left ventricle. The contralateral right kidney was used as a control. The left kidney was stored for 48 h at 4 degrees C to produce cold ischemia. Caspase 3 activity was massively (100-fold) increased in cold ischemic kidneys compared with controls. On immunoblot analysis, the processed form of caspase-3 was increased in cold ischemic kidneys compared with controls. The increase in caspase-3 was associated with significantly more renal tubular apoptosis and brush-border injury. In addition, caspase-2, -8 and -9 activities were increased in cold ischemic kidneys. The pancaspase inhibitor prevented the formation of the processed form of caspase-3 and the increase in caspase activity, and reduced apoptosis and brush-border injury. Caspase inhibition may prove useful in kidney preservation. PMID- 15268726 TI - Novel Duchenne muscular dystrophy treatment through myoblast transplantation tolerance with anti-CD45RB, anti-CD154 and mixed chimerism. AB - Duchenne muscular dystrophy (DMD) is a fatal disease caused by a defect in the skeletal muscle protein, dystrophin. One potential therapy for DMD involves transplantation of myoblasts from normal individuals. Unfortunately, myoblast allografts are particularly immunogenic and transplant tolerance in dystrophic (mdx/mdx) mice has not yet been achieved despite using strategies successful in other allograft models. Here, we attempted to induce 'central tolerance' using either haplo- or fully allogeneic bone marrow after conditioning with low-dose (3 Gy) whole body irradiation and anti-CD154 or anti-CD45RB mAbs. With one exception, these mice lacked persistent chimerism, long-term survival of myoblast allografts, or tolerance. In contrast, the addition of anti-CD45RB to anti-CD154 uniformly resulted in long-lived high-level mixed chimerism, long-term (>100 days) engraftment of allogeneic myoblasts and deletion of donor-reactive cells. Moreover, all recipients exhibited tolerance to second myoblast allografts or donor-specific tolerance to skin transplants performed >80 days after the initial graft. Thus, we now report that anti-CD45RB synergizes with anti-CD40L to promote stable mixed chimerism and robust tolerance to myoblast allografts for the first time. This novel protocol may be applicable to future clinical trials in myoblast transplantation for treatment of DMD and for transplantation of other immunogenic allografts. PMID- 15268727 TI - Prolonged hypothermia causes primary nonfunction in preserved canine renal allografts due to humoral rejection. AB - Canine kidney preservation models have historically used autotransplants to avoid the complications of rejection, although clinically all transplants are allografts. This study investigated the effects of preservation time and method on early kidney function in a canine allograft vs. autograft model. Kidneys were harvested from beagles, preserved by cold storage (CS) in UW solution for 0, 24 or 72 h, or by machine perfusion (MP) with Belzer MPS for 72 h. In some experiments 45 min of warm ischemia (WI) was performed in situ before harvest. Allograft recipients received steroid immunosuppression. Kidney function was assessed by serum creatinine and survival for 7 days. Allografts preserved for 0 and 24 h performed as well as autografts. Allografts preserved for 72 h by either CS or MP had a higher incidence of primary nonfunction (PNF) compared with autografts, as determined by survival (50% vs. 100%, p < 0.003). Primary nonfunction kidneys had thrombotic microangiopathy, vascular and peritubular capillary binding of IgM and complement C4d, and evidence of circulating donor specific antibodies; all consistent with humoral rejection. These responses were dependent on hypothermia time and were not attributable to ischemia, immunosuppression, preservation solution, or cellular rejection. In conclusion, prolonged hypothermia can cause PNF in allografts owing to acute humoral rejection. PMID- 15268728 TI - Risk factors for the development of post-transplant lymphoproliferative disorder in a large animal model. AB - A high incidence of a post-transplant lymphoproliferative disorder (PTLD) is observed in miniature swine conditioned for allogeneic hematopoietic cell transplantation using a protocol involving T-cell depletion and cyclosporine therapy. This study was designed to assess contributing factors to disease development. Forty-six animals were studied including 12 (26%) that developed PTLD. A number of risk factors for PTLD were examined, including degree of immunosuppression, degree of MHC mismatch and infection by a porcine lymphotrophic herpesvirus (PLHV-1). Flow cytometry was used to measure host and donor T- and B-cell levels in the peripheral blood. Porcine lymphotrophic herpesvirus viral load was determined by quantitative PCR. Animals developing PTLD had significantly lower levels of T cells on the day of transplant. Cyclosporine levels did not differ significantly between animals with and without PTLD. Animals receiving transplants across a two-haplotype mismatch barrier showed an increased incidence of PTLD. All animals with PTLD had significant increases in PLHV-1 viral loads. Porcine lymphotrophic herpesvirus viral copy numbers remained at low levels in the absence of disease. The availability of a preclinical large-animal model with similarities to PTLD of humans may allow studies of the pathogenesis and treatment of that disorder. PMID- 15268729 TI - A decade of living lobar lung transplantation: perioperative complications after 253 donor lobectomies. AB - Living lobar lung transplantation places two donors at risk for each recipient. We examined the perioperative outcomes associated with the 253 donor lobectomies performed at our institution during our first decade of living lobar lung transplantation. There have been no perioperative or long-term deaths. 80.2% of donors (n = 203) had no perioperative complications, while fifty (19.8%) had one or more complication. The incidence of intraoperative complications was 3.6%. Complications requiring reoperation occurred in 3.2% of donors. 15.0% of donors had other perioperative complications; the most serious were two donors who developed pulmonary artery thrombosis, while the most common was the need for an additional thoracostomy tube or a thoracostomy tube for >/=14 d for persistent air leaks and/or drainage. Right-sided donors were more likely to have a perioperative complication than left-sided donors (odd ratio 2.02, p = 0.04), probably secondary to right lower and middle lobe anatomy. This experience has shown donor lobectomy to be associated with a relatively low morbidity and no mortality, and is important if this procedure is to be considered an option at more pulmonary transplant centers, given continued organ shortages and differences in philosophical and ethical acceptance of live PMID- 15268730 TI - Long-term renal allograft survival: have we made significant progress or is it time to rethink our analytic and therapeutic strategies? AB - Impressive renal allograft survival improvement between 1988 and 1995 has been described using projections of half-lives based on limited actual follow up. We aimed, now with sufficient follow up available to calculate real half-lives. Real half-lives calculated from Kaplan-Meier curves for the overall population as well as subsets of repeat transplants and African Americans recipients were examined. Real half-lives were substantially shorter than projected half-lives. As a whole, half-lives have improved by about 2 years between 1988 and 1995 as compared to the earlier projected 6 years of improvement. The improvement seems to be driven primarily by the improvement in graft survival of re-transplants. First transplants showed a cumulative increase in graft survival of less than 6 months. Projected half-lives are a risky estimation of long-term survival especially when based on short actual follow up. First transplant survival has only marginally improved during the early years of post transplant follow up while no significant improvement in long-term survival could be detected between 1988 and 1995. Redirection of attention from early endpoints towards the process of long-term graft loss may be necessary to sustain early gains in the long term. PMID- 15268731 TI - Screening for West Nile virus in organ transplantation: a medical decision analysis. AB - The Organ Procurement and Transplant Network (OPTN) has recently announced that screening for West Nile Virus (WNV) in deceased organ donors is not recommended at this time. The purpose of this report was to examine the impact of this recommendation by using medical decision analysis. Without screening the rate of disease transmission was assumed to be the same as in donated blood with a case fatality rate of 25%. With screening we assumed the baseline screening test specificity and sensitivity to be 99.5% and 95%, respectively. The analysis was confined to heart, liver and kidney recipients. Survival probabilities and transplant rates were taken from UNOS. Annual screening could result in the loss of potentially 452.4 life years (113.8 for heart, 272.6 for liver and 66.0 for kidney). Most positive test results would be false-positive. Screening would be preferable for kidney donors in areas of high disease prevalence and high test specificity. However, for heart and liver most scenarios were associated with a net loss of life with screening, except if patients were stable on the wait list with particularly high case fatality rates from WNV. Current recommendations by OPTN that screening is not mandatory seem appropriate until further data are available. PMID- 15268732 TI - Allograft survival following adult-to-adult living donor liver transplantation. AB - Adult-to-adult living donor liver transplantation (AALDLT) is emerging as a method to treat patients with end-stage liver disease. The aims of this study were to identify donor and recipient characteristics of AALDLT, to determine variables that affect allograft survival, and to examine outcomes compared with those achieved following cadaveric transplantation. Cox proportional hazards models were fit to examine characteristics associated with the survival of AALDLT. Survival of AALDLT was then compared with cadaveric allografts in multivariable Cox models. Older donor age (>44 years), female-to-male donor to recipient relationship, recipient race, and the recipient medical condition before transplant were factors related to allograft failure among 731 AALDLT. Despite favorable donor and recipient characteristics, the rate of allograft failure, specifically the need for retransplantation, was increased among AALDLT (hazard ratio 1.66, 95% C.I. = 1.30-2.11) compared with cadaveric recipients. In conclusion, among AALDLT recipients, selecting younger donors, placing the allografts in recipients who have not had a prior transplant and are not in the ICU, may enhance allograft survival. Analysis of this early experience with AALDLT suggests that allograft failure may be higher than among recipients of a cadaveric liver. PMID- 15268733 TI - Pilot randomized study of early tacrolimus withdrawal from a regimen with sirolimus plus tacrolimus in kidney transplantation. AB - We performed a randomized trial to compare two regimens of low-risk kidney allograft recipients in the first year after transplantation. Both regimens initially included sirolimus, tacrolimus and steroids; one with long-term maintenance with these drugs vs. tacrolimus withdrawal. Group I: sirolimus levels of 4-8 ng/mL, plus tacrolimus 8-12 ng/mL for 3 months, and 5-10 ng/mL after month 3. Group II: sirolimus concentration of 8-16 ng/mL, plus tacrolimus 3-8 ng/mL with tacrolimus elimination from month 3 onwards. Owing to difficulties in achieving target levels, the protocol was amended to increase the doses. Eighty seven patients were recruited. In the intention-to-treat analysis, glomerular filtration rate (GFR) at 12 months, adjusted to zero for graft loss, was similar in both groups (58.8 and 59.9 mL/min). Analysis of patients remaining on protocol showed that GFR was higher in group II only in the patients postamendment (58.4 and 72.9 mL/min, p = 0.03). Rates of biopsy-confirmed rejection (BCAR) were 9.3% and 22.7% in groups I and II, respectively (p = NS). After amendment, BCAR rates were 10.3% and 11.1% (p = NS). Diastolic blood pressure was significantly lower in patients who eliminated tacrolimus (80.4 vs. 75.6 mmHg) (p = 0.03). Combining sirolimus and tacrolimus with adequate loading doses was associated with a low incidence of BCAR, and allowed tacrolimus elimination in a high proportion of patients, which may be followed by amelioration in renal function and blood pressure. PMID- 15268734 TI - Plasmapheresis, CMV hyperimmune globulin, and anti-CD20 allow ABO-incompatible renal transplantation without splenectomy. AB - The majority of preconditioning protocols developed to allow ABO-incompatible (ABOi) renal transplantation include concurrent splenectomy as a prerequisite to successful engraftment. Our center has developed a preconditioning protocol that includes plasmapheresis (PP), low-dose CMV hyperimmune globulin (CMVIg), and anti CD20 monoclonal antibody (rituximab) to allow ABOi renal transplantation without splenectomy. Our initial experience has included treatment of six recipients and successful transplantation from blood group A(1), A(2), and group B living donors. Mean (+/- SD) serum creatinine was 1.3 +/- 0.1 mg/dL among the six recipients and no episodes of antibody-mediated rejection (AMR) occurred at a median follow-up of 12 months. ABO antibody titers have remained below pretreatment levels. The absence of AMR and stable allograft function in this series show the potential of this preconditioning protocol to increase ABOi renal transplantation. The use of rituximab, allowing avoidance of splenectomy, may further remove one of the significant disincentives to ABOi transplantation, and eliminate the risk of post-splenectomy infections. PMID- 15268735 TI - Acute humoral rejection of human lung allografts and elevation of C4d in bronchoalveolar lavage fluid. AB - Antibody-mediated rejection is well established for renal allografts but remains controversial for lung allografts. Cardinal features of antibody-mediated rejection in renal allografts include antibodies to donor human leukocyte antigen (HLA) and evidence for antibody action, such as complement activation demonstrated by C4d deposition. We report a lung allograft recipient with circulating antibodies to donor HLA who failed treatment for acute cellular rejection but responded to therapy for humoral rejection. To address the second criteria for antibody-mediated rejection, we determined whether complement activation could be detected by measuring C4d in bronchoalveolar lavage fluid (BALF) by ELISA. Airway allergen challenge of asthmatics activates the complement pathway; therefore, we used BALF from asthmatics pre- and post-allergen challenge to measure C4d. These controls demonstrated that ELISA could detect increases in C4d after allergen challenge. BALF from the index patient had elevated C4d concomitant with graft dysfunction and anti-donor HLA in the absence of infection. Analysis of BALF from 25 additional lung allograft recipients showed that C4d concentrations >100 ng/mL were correlated with anti-HLA antibodies (p = 0.006), but were also observed with infection and in asyptomatic patients. The findings support the occurrence of anti-HLA-mediated lung allograft rejection and suggest that C4d measurement in BALF may be useful in diagnosis. PMID- 15268736 TI - Prospective evaluation of the clinical utility of different methods for the detection of human cytomegalovirus disease after liver transplantation. AB - Standardized human cytomegalovirus (HCMV) assays were prospectively evaluated to predict HCMV disease. In 135 consecutive adult liver transplantations, pp65 antigenemia, quantitative HCMV-DNA and qualitative pp67-messenger-RNA were determined weekly. No ganciclovir prophylaxis or preemptive treatment was used. One hundred and ten (81.5%) patients showed no HCMV-infection, 25 patients were positive in at least one of the HCMV-tests (18.5%). Four suffered from HCMV viral syndrome (3.0%) and another four from tissue invasive disease. In total, pp65 antigenemia was detected in 18, HCMV-DNA in 22 and pp67-mRNA in 18 patients. The sensitivity and negative predictive value (NPV) for HCMV-disease was 100% for all tests. The PPV for symptomatic HCMV-infection was 47% for pp67 mRNA. In contrast, the PPV of pp65-antigenemia (using a threshold of > 2/200 000 cells) and quantitative PCR (using a cutoff of > 5000 copies/mL) were 80% and 89%, respectively. A cost analysis revealed symptom-triggered or preemptive treatment was less expensive than general ganciclovir prophylaxis, if the incidence of CMV disease was low (<30%). Quantitative human cytomegalovirus (HCMV)-DNA and pp65 antigen assays have a comparable sensitivity and can therefore predict the onset of HCMV symptoms at an early stage. Compared with general prophylaxis, symptom triggered or preemptive treatment based on one of these assays might reduce the costs and also the danger of ganciclovir resistance. PMID- 15268737 TI - Calcium oxalate deposition in renal allografts: morphologic spectrum and clinical implications. AB - Many aspects of calcium oxalate (CaOx) deposition in renal transplant biopsies are not known. Review of all renal transplant biopsies performed in a 7-year period showed that CaOx deposition could be classified into three groups. Group I: Seven biopsies within a month post-transplant displayed rare CaOx foci against a background of acute tubular necrosis or acute cell-mediated rejection. At follow-up, five grafts functioned well and two failed due to chronic allograft nephropathy. CaOx in this context was an incidental finding secondary to a sudden excretion of an end-stage renal disease-induced increased body burden of CaOx. Group II: Two biopsies performed 2 and 10 months post-transplant showed rare CaOx foci against a background of chronic allograft nephropathy, leading to graft loss. CaOx in this context reflected nonspecific parenchymal deposition due to chronic renal failure regardless of causes. Group III: One biopsy with recurrent PH1 characterized by marked CaOx deposition associated with severe tubulointerstitial injury and graft loss 6 months post-transplant. There were two previously reported cases in which CaOx deposition in the renal allografts was due the antihypertensive drug naftidrofuryl oxalate or increased intestinal absorption of CaOx. CaOx deposition in renal allografts can be classified in different categories with distinctive morphologic features and clinical implications. PMID- 15268738 TI - Therapeutic drug monitoring of sirolimus: effect of concomitant immunosuppressive therapy and optimization of drug dosing. AB - Sirolimus (SRL) is a new immunosuppressant which shares a common metabolic pathway with several other immunosuppressive agents. This leads to potential pharmacokinetic interactions that might affect SRL blood levels with relevant clinical consequences. As a validated laboratory, 2658 SRL trough samples (corresponding to 495 kidney transplant recipients treated with different immunosuppressive regimens) from more than 40 Italian Transplant Units were analyzed. We found that dose-normalized SRL trough levels were significantly higher in patients treated with cyclosporine (CsA) and SRL (4.15 +/- 2.23 ng/mL/mg SRL), compared with patients treated with mycophenolate mofetil (MMF) and SRL (3.26 +/- 1.86 ng/mL/mg SRL; p < 0.01) or with MMF, steroids and SRL (2.52 +/- 1.73 ng/mL/mg SRL; p < 0.01). Mean intra- and interpatient variabilities were 19% and 47%, respectively. Both parameters are significantly affected by the time postsurgery, with the first week post transplantation being associated with the greatest variability. As additional analysis, a simple dose adjustment formula has been proposed as a useful tool to guide SRL dose changes. The proposed equation has been able to predict SRL concentration after a dose change in 73% of the tested samples. These findings suggest that different immunosuppressants significantly interfere with SRL bioavailability. Strategies aimed at reducing variability in SRL exposure may have a positive clinical impact. PMID- 15268739 TI - Interferon-alpha therapy in liver transplant recipients: lack of association with increased production of anti-HLA antibodies. AB - Interferon-alpha (IFN) is a useful treatment for active HCV infection. In kidney transplantation, IFN has been shown to trigger acute rejection with de novo anti HLA antibodies. Interferon-alpha has not been reported to enhance the risk of acute rejection in HCV-positive liver transplant recipients (LTRs). Sera were collected from 44 LTRs greater than 6 months post-transplant. Sera were tested with ELISA for the presence and the specificity of anti-HLA antibodies. The prevalence of anti-HLA antibodies was 11% and was not significantly different in 13 HCV-positive recipients who received IFN, compared with 10 who did not receive IFN (8% vs. 20%), or with 21 HCV-negative recipients (10%). None of the patients had an acute rejection after starting IFN. In this study, LTRs receiving IFN did not have an increased frequency of anti-HLA antibodies. This may partially explain the safety of IFN previously reported in LTRs requiring antiviral therapy. PMID- 15268740 TI - Rituximab, anti-CD20, induces in vivo cytokine release but does not impair ex vivo T-cell responses. AB - Pre-formed HLA antibodies (Ab), reported as panel-reactive antibody (PRA), prolong transplant waiting time. We hypothesized that rituximab (RIT) could reduce PRA via B-cell depletion. As part of a Phase I study of single RIT dose, we studied in vivo and ex vivo effects on T-cell immune responses. Nine subjects (n = 3) were treated at 50, 150, and 375 mg/m(2). Serum interleukin-1alpha (IL 1alpha), IL-6, IL-12, tumor necrosis factor beta (TNF-beta), and interferon-gamma (IFN-gamma) were measured by enzyme-linked immunosorbent assay (ELISA). T-cell function was monitored with T-cell proliferation assays. IL-6 levels rose in eight patients (7.15 +/- 4.38 pg/mL to 86.22 +/- 77.08, p = 0.021). The high-dose group had detectable TNF-betapost rituximab infusion (874.7 +/- 1466.5 pg/mL). There was no decline in T-cell proliferation in response to phytohemagglutinin or allogeneic lymphocyte stimuli. Stimulation indices in the presence of both concentrations of tetanus toxoid rose significantly at 4 weeks. PMID- 15268741 TI - Intravenous infusion of apoptotic cells simultaneously with allogeneic hematopoietic grafts alters anti-donor humoral immune responses. AB - Intravenous infusion of apoptotic donor or third-party leukocytes simultaneously with an allogeneic donor bone marrow (BM) graft favors engraftment across major histocompatibility barriers. While verifying that such apoptotic cell infusion might not also be associated with antibody (Ab)-mediated allo-immune responses, we found, rather strikingly, that apoptotic cell infusion could in fact successfully prevent a humoral allo-immunization against a BM graft in mice. Indeed, among recipients having rejected their BM graft, prior apoptotic cell infusion was associated with a near absence of Ab-mediated allo-responses, while such an immunization was frequently observed in the absence of apoptotic cell infusion. This was also observed when infusing host apoptotic cells, thus showing that the prevention of immunization was linked to the apoptotic state of the cells rather than mediated by residual anti-recipient activity. In vivo anti transforming growth factor-beta (TGF-beta) treatment resulted in the loss of this apoptotic cell infusion-associated protective effect on humoral allo-responses. Further studies will determine whether apoptotic cell infusion, in addition to hematopoietic graft facilitation might also contribute to preventing deleterious Ab-mediated allo-responses in various transplantation settings. PMID- 15268742 TI - Invasive pneumococcal infections in adult lung transplant recipients. AB - An increased risk of invasive pneumococcal infection (IPI) has been described among kidney or heart transplant recipients, but the epidemiology of IPI among lung transplant recipients has not been previously reported. We undertook a single center, retrospective cohort study to define the incidence, timing, clinical, and microbiologic features of IPI in lung transplant patients. Fourteen out of 220 recipients (6.4%) developed IPI at a median of 1.3 years after transplantation (incidence rate: 22.7 cases per 1000 person-years). All patients were receiving trimethoprim-sulfamethoxazole (TMP-SMX) prophylaxis at the time of diagnosis, and 10 (71%) had TMP-SMX-resistant isolates. All isolates were from the 23 valent polysaccharide vaccine-associated serogroups. The high incidence of IPI in lung transplant recipients is similar to that reported in kidney and heart recipients. Alternative prevention strategies, including use of the conjugated pneumococcal vaccine, should be explored in future studies. PMID- 15268743 TI - Steroid- and ATG-resistant rejection after double forearm transplantation responds to Campath-1H. AB - We herein report on immunological and histological observations in the first bilateral forearm transplant recipient. The last of three rejection episodes occurring on day 95 after transplantation was resistant to steroid and antithymocyte globulin (ATG) treatment. Histology demonstrated lymphocytic infiltrates, apoptotic and necrotic keratinocytes and focal desquamation of the epidermis. Therapy with Campath-1H, however, resulted in complete restitution of normal skin. This is the first report on a successful rescue therapy with Campath 1H in a severe, steroid- and ATG-resistant rejection. Hence, Campath-1H treatment might be a novel and powerful therapeutic option for multiresistant allograft rejection. PMID- 15268744 TI - Severe hepatitis C infection in a renal transplant recipient following hepatitis C genotype mismatch transplant. AB - We report a case of a 32-year-old female with histologically and clinically inactive chronic hepatic C infection, who received a cadaveric renal transplant from a hepatitis C-positive donor with a different genotype. The genotype mismatch (genotype 1 to genotype 2) and change to tacrolimus-based immunosuppression resulted in severe hepatitis C infection characterized by a 10 fold increase in transaminase levels and grade 3 inflammation histologically. Our report highlights the risk of genotype-mismatch transplants in solid organ transplantation. PMID- 15268746 TI - Payment for living kidney donors (vendors) is not an abstract ethical discussion occurring in a vacuum. PMID- 15268747 TI - Signalling shutdown strategies in aging immune cells. AB - It is important for the resolution of inflammation that the number and activity of immune cells are reduced. Clearance of immune cells may be achieved by apoptosis and phagocytosis of cell fragments by macrophages. However, signalling shutdown by immune cells committed to apoptosis occurs early in the progression of these cells towards fragmentation and, it could be argued, is a key feature of apoptosis. There is surprisingly little known about the mechanisms that underlie this signalling shutdown, in particular the shutdown of Ca(2+) influx. The consequences and the potential mechanisms by which Ca(2+) influx shutdown is achieved are discussed. In addition, the potential consequences for cell signalling of cytochrome c release from mitochondria and of phosphatidyl-serine externalization are discussed. The aim of the review is therefore to highlight the evidence for various signalling shutdown strategies in immune cells that may limit their activity during progression towards apoptosis. PMID- 15268748 TI - Stress responses and innate immunity: aging as a contributory factor. AB - Evolutionary pressure has selected individuals with traits that allow them to survive to reproduction, without consideration of the consequences for the post child rearing years and old age. In the 21st century, society is populated increasingly by the elderly and with the falling birth rate and improved health care this trend is set to continue for the foreseeable future. To minimize the potential burden on health services one would hope that 'growing old gracefully' should also mean 'growing old healthily'. However, for too many the aging process is accompanied by increasing physical and mental frailty producing an elevated risk of physical and psychological stress in old age. Stress is a potent modulator of immune function, which in youth can be compensated for by the presence of an optimal immune response. In the elderly the immune response is blunted as a result of the decline in several components of the immune system (immune senescence) and a shifting to a chronic pro-inflammatory status (the so called 'inflamm-aging' effect). We discuss here what is known of the effects of both stress and aging upon the innate immune system, focusing in particular upon the age-related alterations in the hypopituitary-adrenal axis. We propose a double hit model for age and stress in which the age-related increase in the cortisol/sulphated dehydroepiandrosterone ratio synergizes with elevated cortisol during stress to reduce immunity in the elderly significantly. PMID- 15268749 TI - Innate immunity in aging: impact on macrophage function. AB - Innate and adaptive immune functions decline with age, leading to increased susceptibility to infectious diseases and cancer, and reduced responses to preventive vaccination in the elderly population. Macrophages function as 'pathogen sensors' and play an important role in the initiation of inflammatory responses, elimination of pathogens, manipulation of the adaptive immune response and reparation of damaged tissue. In this paper, we review the literature addressing the impact of aging on the macrophage population. PMID- 15268750 TI - How chronic inflammation can affect the brain and support the development of Alzheimer's disease in old age: the role of microglia and astrocytes. AB - A huge amount of evidence has implicated amyloid beta (A beta) peptides and other derivatives of the amyloid precursor protein (beta APP) as central to the pathogenesis of Alzheimer's disease (AD). It is also widely recognized that age is the most important risk factor for AD and that the innate immune system plays a role in the development of neurodegeneration. Little is known, however, about the molecular mechanisms that underlie age-related changes of innate immunity and how they affect brain pathology. Aging is characteristically accompanied by a shift within innate immunity towards a pro-inflammatory status. Pro-inflammatory mediators such as tumour necrosis factor-alpha or interleukin-1 beta can then in combination with interferon-gamma be toxic on neurons and affect the metabolism of beta APP such that increased concentrations of amyloidogenic peptides are produced by neuronal cells as well as by astrocytes. A disturbed balance between the production and the degradation of A beta can trigger chronic inflammatory processes in microglial cells and astrocytes and thus initiate a vicious circle. This leads to a perpetuation of the disease. PMID- 15268751 TI - NK and NKT cell functions in immunosenescence. AB - Immunosenescence is defined as the state of dysregulated immune function that contributes to the increased susceptibility to infection, cancer and autoimmune diseases observed in old organisms, including humans. However, dysregulations in the immune functions are normally counterbalanced by continuous adaptation of the body to the deteriorations that occur over time. These adaptive changes are likely to occur in healthy human centenarians. Both innate (natural) and adaptive (acquired) immune responses decline with advancing age. Natural killer (NK) and natural killer T (NKT) cells represent the best model to describe innate and adaptive immune response in aging. NK and NKT cell cytotoxicity decreases in aging as well as interferon-gamma (IFN-gamma) production by both activated cell types. Their innate and acquired immune responses are preserved in very old age. However, NKT cells bearing T-cell receptor (TCR) gammadelta also display an increased cytotoxicity and IFN-gamma production in very old age. This fact suggests that NKT cells bearing TCRgammadelta are more involved in maintaining innate and adaptive immune response in aging leading to successful aging. The role played by the neuroendocrine-immune network and by nutritional factors, such as zinc, in maintaining NK and NKT cell functions in aging is discussed. PMID- 15268752 TI - Regulation of aging and innate immunity in C. elegans. AB - The free-living soil nematode Caenorhabditis elegans is a versatile model for the study of the genetic regulation of aging and of host-pathogen interactions. Many genes affecting multiple processes, such as neuroendocrine signalling, nutritional sensing and mitochondrial functions, have been shown to play important roles in determining the lifespan of C. elegans. The DAF-2-mediated insulin signalling pathway is the major pathway that regulates aging in this nematode and this role appears universal; neuroendrocrine signalling also affects aging in Drosophila and mice. Recent studies have shown that the innate immune function in C. elegans is modulated by signalling from the TGF-beta-like, the p38 MAPK and the DAF-2 insulin pathways. The requirement for the DAF-2 pathway in modulating aging and immunity suggests that these processes may be linked at the molecular level. It is well known that as humans age, immunosenescence occurs in which there is a general degradation of immune efficiency. However, the molecular mechanisms involved in this process remain unclear. In this review, we discuss the molecular mechanisms that modulate aging and immune response and attempt to suggest molecular links between these two processes. PMID- 15268753 TI - Immunity and aging: the enemy within? AB - Functional analyses of changes in the immune response indicate that aging is associated with a decline of adaptive immunity whereas innate immunity is ramped up. Gene expression studies also support age-dependent changes in immunity. Studies using a large panel of methodologies and multiple species show that some of the most dramatic transcriptional changes that occur during aging are associated with immunity. This observation leads to two fundamental questions: (1) Why is the immune response altered with age? (2) Is this a consequence of aging or does it contribute to it? The origin of these changes and the mechanistic relationship among them as well as with aging must be identified. In mammals, this task is complicated by the interdependence of the innate and adaptive immune systems. The value of invertebrates as model organisms to help answer these questions is presented. This includes a description of the immune response in invertebrate models and how it compares with vertebrates, focusing on conserved pathways. Finally, these questions are explored in light of recent reports and data from our laboratory. Experimental alterations of longevity indicate that the differential expression of immunity-related genes during aging is linked to the rate of aging. Long-lived nematodes are more resistant to pathogens and blocking the expression of immune-related genes can prevent lifespan extension. These observations suggest that the immune response has a positive effect on longevity, possibly by increasing fitness. By contrast, it has been reported that activation of the immune system can reduce longevity upon starvation. We also observed that deregulation of the immune response has drastic effects on viability and longevity in Drosophila. These data suggest that the immune response results in a trade-off between beneficial and detrimental effects that might profoundly affect the aging process. Given this, immunity may be an ally early in life, but turns out to be an enemy as we age. PMID- 15268754 TI - Hormones and immune function: implications of aging. AB - Aging is associated with a decline in immunity described as immunosenescence. This is paralleled by a decline in the production of several hormones, as typically illustrated by the menopausal loss of ovarian oestrogen production. However, other hormonal changes that occur with aging and that potentially impact on immune function include the release of the pineal gland hormone melatonin and pituitary growth hormone, adrenal production of dehydroepiandrosterone and tissue specific availability of active vitamin D. It remains to be established whether hormonal changes with aging actually contribute to immunosenescence and this area is at the interface of fact and fiction, clearly inviting systematic research efforts. As a step in this direction, the present review summarizes established facts on the physiology of secretion and function of hormones that, in most cases, decline with aging and that are likely to affect the immune system. PMID- 15268755 TI - Signal transduction and functional changes in neutrophils with aging. AB - It is well known that the immune response decreases during aging, leading to a higher susceptibility to infections, cancers and autoimmune disorders. Most widely studied have been alterations in the adaptive immune response. Recently, the role of the innate immune response as a first-line defence against bacterial invasion and as a modulator of the adaptive immune response has become more widely recognized. One of the most important cell components of the innate response is neutrophils and it is therefore important to elucidate their function during aging. With aging there is an alteration of the receptor-driven functions of human neutrophils, such as superoxide anion production, chemotaxis and apoptosis. One of the alterations underlying these functional changes is a decrease in signalling elicited by specific receptors. Alterations were also found in the neutrophil membrane lipid rafts. These alterations in neutrophil functions and signal transduction that occur during aging might contribute to the significant increase in infections in old age. PMID- 15268756 TI - Neurogenesis in a rat model of age-related cognitive decline. AB - Age-related decrements in hippocampal neurogenesis have been suggested as a basis for learning impairment during aging. In the current study, a rodent model of age related cognitive decline was used to evaluate neurogenesis in relation to hippocampal function. New hippocampal cell survival was assessed approximately 1 month after a series of intraperitoneal injections of 5-bromo-2'-deoxyuridine (BrdU). Correlational analyses between individual measures of BrdU-positive cells and performance on the Morris water maze task provided no indication that this measure of neurogenesis was more preserved in aged rats with intact cognitive abilities. On the contrary, among aged rats, higher numbers of BrdU-positive cells in the granule cell layer were associated with a greater degree of impairment on the learning task. Double-labelling studies confirmed that the majority of the BrdU+ cells were of the neuronal phenotype; the proportion of differentiated neurons was not different across a broad range of cognitive abilities. These data demonstrate that aged rats that maintain cognitive function do so despite pronounced reductions in hippocampal neurogenesis. In addition, these findings suggest the interesting possibility that impaired hippocampal function is associated with greater survival of newly generated hippocampal neurons at advanced ages. PMID- 15268757 TI - Genome-scale expression profiling of Hutchinson-Gilford progeria syndrome reveals widespread transcriptional misregulation leading to mesodermal/mesenchymal defects and accelerated atherosclerosis. AB - Hutchinson-Gilford progeria syndrome (HGPS) is a rare genetic disease with widespread phenotypic features resembling premature aging. HGPS was recently shown to be caused by dominant mutations in the LMNA gene, resulting in the in frame deletion of 50 amino acids near the carboxyl terminus of the encoded lamin A protein. Children with this disease typically succumb to myocardial infarction or stroke caused by severe atherosclerosis at an average age of 13 years. To elucidate further the molecular pathogenesis of this disease, we compared the gene expression patterns of three HGPS fibroblast cell strains heterozygous for the LMNA mutation with three normal, age-matched cell strains. We defined a set of 361 genes (1.1% of the approximately 33,000 genes analysed) that showed at least a 2-fold, statistically significant change. The most prominent categories encode transcription factors and extracellular matrix proteins, many of which are known to function in the tissues severely affected in HGPS. The most affected gene, MEOX2/GAX, is a homeobox transcription factor implicated as a negative regulator of mesodermal tissue proliferation. Thus, at the gene expression level, HGPS shows the hallmarks of a developmental disorder affecting mesodermal and mesenchymal cell lineages. The identification of a large number of genes implicated in atherosclerosis is especially valuable, because it provides clues to pathological processes that can now be investigated in HGPS patients or animal models. PMID- 15268759 TI - Transient spleen enlargement in peripheral blood progenitor cell donors given G CSF. AB - The administration of granulocyte colony-stimulating factor (G-CSF) to peripheral blood progenitor cell (PBPC) donors causes spleen length to increase, but the duration of enlargement is not known. Eighteen healthy subjects were given 10 microg/kg of G-CSF for 5 days and a PBSC concentrate was collected by apheresis. Ultrasound scans were used to assess craniocaudal spleen length before and after G-CSF administration. Mean spleen length increased from a baseline length of 10.7 +/- 1.3 cm to 12.1 +/- 1.2 cm on the apheresis day (p < 0.001). Ten days after apheresis, spleen length fell to 10.5 +/- 1.2 cm and did not differ from baseline levels (p = 0.57), but in 3 subjects remained 0.5 cm greater than baseline length. Increases in spleen length in PBPC donors are transient and reversible. PMID- 15268761 TI - Echocardiographic assessment and percutaneous closure of multiple atrial septal defects. AB - Atrial septal defect closure is now routinely performed using a percutaneous approach under echocardiographic guidance. Centrally located, secundum defects are ideal for device closure but there is considerable morphological variation in size and location of the defects. A small proportion of atrial septal defects may have multiple fenestrations and these are often considered unsuitable for device closure. We report three cases of multiple atrial septal defects successfully closed with two Amplatzer septal occluders. PMID- 15268760 TI - Cancer of the endometrium: current aspects of diagnostics and treatment. AB - BACKGROUND: Endometrial cancer represents a tumor entity with a great variation in its incidence throughout the world (range 1 to 25). This suggests enormous possibilities of cancer prevention due to the fact that the incidence is very much endocrine-related, chiefly with obesity, and thus most frequent in the developed world. As far as treatment is concerned, it is generally accepted that surgery represents the first choice of treatment. However, several recommendations seem reasonable especially with lymphadenectomy, even though they are not based on evidence. All high-risk cases are generally recommended for radiotherapy. METHODS: A literature search of the Medline was carried out for all articles on endometrial carcinoma related to diagnosis and treatment. The articles were systematically reviewed and were categorized into incidence, etiology, precancerosis, early diagnosis, classification, staging, prevention, and treatment. The article is organized into several similar subheadings. CONCLUSIONS: In spite of the overall good prognosis during the early stages of the disease, the survival is poor in advanced stages or recurrences. Diagnostic measures are very well able to detect asymptomatic recurrences. These only seem justified if patients' chances are likely to improve, otherwise such measures increases costs as well as decrease the patients' quality of life. To date neither current nor improved concepts of endocrine treatment or chemotherapy have been able to substantially increase patients' chances of survival. Therefore, newer concepts into the use of antibodies e.g. trastuzumab in HER2-overexpressing tumors and the newer endocrine compounds will need to be investigated. Furthermore, it would seem highly desirable if future studies were to identify valid criteria for an individualized management, thereby maximizing the benefits and minimizing the risks. PMID- 15268762 TI - Insights into the pathogenesis of vein graft disease: lessons from intravascular ultrasound. AB - The success of coronary artery bypass grafting (CABG) is limited by poor long term graft patency. Saphenous vein is used in the vast majority of CABG operations, although 15% are occluded at one year with as many as 50% occluded at 10 years due to progressive graft atherosclerosis. Intravascular ultrasound (IVUS) has greatly increased our understanding of this process. IVUS studies have shown that early wall thickening and adaptive remodeling of vein grafts occurs within the first few weeks post implantation, with these changes stabilising in angiographically normal vein grafts after six months. Early changes predispose to later atherosclerosis with occlusive plaque detectable in vein grafts within the first year. Both expansive and constrictive remodelling is present in diseased vein grafts, where the latter contributes significantly to occlusive disease. These findings correlate closely with experimental and clinicopathological studies and help define the windows for prevention, intervention or plaque stabilisation strategies. IVUS is also the natural tool for evaluating the effectiveness of pharmacological and other treatments that may prevent or slow the progression of vein graft disease in clinical trials. PMID- 15268763 TI - Early endothelial dysfunction in cholesterol-fed rabbits: a non-invasive in vivo ultrasound study. AB - BACKGROUND: Endothelial function in hypercholesterolemic rabbits is usually evaluated ex vivo on isolated aortic rings. In vivo evaluation requires invasive imaging procedures that cannot be repeated serially. AIM: We evaluated a non invasive ultrasound technique to assess early endothelial function in rabbits and compare data with ex vivo measurements. METHODS: Twenty-four rabbits (fed with a cholesterol diet (0.5%) for 2 to 8 weeks) were given progressive infusions of acetylcholine (0.05-0.5 microg/kg/min) and their endothelial function was assessed in vivo by transcutaneous vascular ultrasound of the abdominal aorta. Ex vivo endothelial function was evaluated on isolated aortic rings and compared to in vivo data. RESULTS: Significant endothelial dysfunction was demonstrated in hypercholesterolemic animals as early as 2 weeks after beginning the cholesterol diet (aortic cross-sectional area variation: -2.9% vs. +4% for controls, p < 0.05). Unexpectedly, response to acetylcholine at 8 weeks was more variable. Endothelial function improved in 5 rabbits while 2 rabbits regained a normal endothelial function. These data corroborated well with ex vivo results. CONCLUSION: Endothelial function can be evaluated non-invasively in vivo by transcutaneous vascular ultrasound of the abdominal aorta in the rabbit and results correlate well with ex vivo data. PMID- 15268764 TI - Patients' perspectives on taking warfarin: qualitative study in family practice. AB - BACKGROUND: Despite the well-documented benefits of using warfarin to prevent stroke, physicians remain reluctant to initiate therapy, and especially so with the elderly owing to the higher risk of hemorrhage. Prior research suggests that patients are more accepting of the risk of bleeding than are physicians, although there have been few qualitative studies. The aim of this study was to employ qualitative methods to investigate the experience and perspective of individuals taking warfarin. METHODS: We conducted face-to-face interviews with 21 older patients (12 male, 9 female) who had been taking warfarin for a minimum of six months. Participants were patients at a family practice clinic situated in a large, tertiary care teaching hospital. We used a semistructured interview guide with four main thematic areas: decision-making, knowledge/education, impact, and satisfaction. Data were analysed according to the principles of content analysis. RESULTS AND DISCUSSION: Participants tended to have minimal input into the decision to initiate warfarin therapy, instead relying in great part on physicians' expertise. There appeared to be low retention of information received regarding the therapy; half the patients in our sample possessed only a superficial level of understanding of the risks and benefits. This notwithstanding, participants reported a high level of satisfaction with the care provided and a low level of impact on their day-to-day lives. CONCLUSIONS: Minimal patient involvement in the initial decision and modest knowledge did not appear to diminish satisfaction with warfarin management. At the same time, care providers exert a tremendous influence on the initiation of warfarin therapy and should strive to incorporate patient preferences and expectations into the decision-making process. PMID- 15268765 TI - Attentional influences on functional mapping of speech sounds in human auditory cortex. AB - BACKGROUND: The speech signal contains both information about phonological features such as place of articulation and non-phonological features such as speaker identity. These are different aspects of the 'what'-processing stream (speaker vs. speech content), and here we show that they can be further segregated as they may occur in parallel but within different neural substrates. Subjects listened to two different vowels, each spoken by two different speakers. During one block, they were asked to identify a given vowel irrespectively of the speaker (phonological categorization), while during the other block the speaker had to be identified irrespectively of the vowel (speaker categorization). Auditory evoked fields were recorded using 148-channel magnetoencephalography (MEG), and magnetic source imaging was obtained for 17 subjects. RESULTS: During phonological categorization, a vowel-dependent difference of N100m source location perpendicular to the main tonotopic gradient replicated previous findings. In speaker categorization, the relative mapping of vowels remained unchanged but sources were shifted towards more posterior and more superior locations. CONCLUSIONS: These results imply that the N100m reflects the extraction of abstract invariants from the speech signal. This part of the processing is accomplished in auditory areas anterior to AI, which are part of the auditory 'what' system. This network seems to include spatially separable modules for identifying the phonological information and for associating it with a particular speaker that are activated in synchrony but within different regions, suggesting that the 'what' processing can be more adequately modeled by a stream of parallel stages. The relative activation of the parallel processing stages can be modulated by attentional or task demands. PMID- 15268766 TI - The FU gene and its possible protein isoforms. AB - BACKGROUND: FU is the human homologue of the Drosophila gene fused whose product fused is a positive regulator of the transcription factor Cubitus interruptus (Ci). Thus, FU may act as a regulator of the human counterparts of Ci, the GLI transcription factors. Since Ci and GLI are targets of Hedgehog signaling in development and morphogenesis, it is expected that FU plays an important role in Sonic, Desert and/or Indian Hedgehog induced cellular signaling. RESULTS: The FU gene was identified on chromosome 2q35 at 217.56 Mb and its exon-intron organization determined. The human developmental disorder Syndactyly type 1 (SD1) maps to this region on chromosome 2 and the FU coding region was sequenced using genomic DNA from an affected individual in a linked family. While no FU mutations were found, three single nucleotide polymorphisms were identified. The expression pattern of FU was thoroughly investigated and all examined tissues express FU. It is also clear that different tissues express transcripts of different sizes and some tissues express more than one transcript. By means of nested PCR of specific regions in RT/PCR generated cDNA, it was possible to verify two alternative splicing events. This also suggests the existence of at least two additional protein isoforms besides the FU protein that has previously been described. This long FU and a much shorter isoform were compared for the ability to regulate GLI1 and GLI2. None of the FU isoforms showed any effects on GLI1 induced transcription but the long form can enhance GLI2 activity. Apparently FU did not have any effect on SUFU induced inhibition of GLI. CONCLUSIONS: The FU gene and its genomic structure was identified. FU is a candidate gene for SD1, but we have not identified a pathogenic mutation in the FU coding region in a family with SD1. The sequence information and expression analyses show that transcripts of different sizes are expressed and subjected to alternative splicing. Thus, mRNAs may contain different 5'UTRs and encode different protein isoforms. Furthermore, FU is able to enhance the activity of GLI2 but not of GLI1, implicating FU in some aspects of Hedgehog signaling. PMID- 15268767 TI - Two-stage normalization using background intensities in cDNA microarray data. AB - BACKGROUND: In the microarray experiment, many undesirable systematic variations are commonly observed. Normalization is the process of removing such variation that affects the measured gene expression levels. Normalization plays an important role in the earlier stage of microarray data analysis. The subsequent analysis results are highly dependent on normalization. One major source of variation is the background intensities. Recently, some methods have been employed for correcting the background intensities. However, all these methods focus on defining signal intensities appropriately from foreground and background intensities in the image analysis. Although a number of normalization methods have been proposed, no systematic methods have been proposed using the background intensities in the normalization process. RESULTS: In this paper, we propose a two-stage method adjusting for the effect of background intensities in the normalization process. The first stage fits a regression model to adjust for the effect of background intensities and the second stage applies the usual normalization method such as a nonlinear LOWESS method to the background-adjusted intensities. In order to carry out the two-stage normalization method, we consider nine different background measures and investigate their performances in normalization. The performance of two-stage normalization is compared to those of global median normalization as well as intensity dependent nonlinear LOWESS normalization. We use the variability among the replicated slides to compare performance of normalization methods. CONCLUSIONS: For the selected background measures, the proposed two-stage normalization method performs better than global or intensity dependent nonlinear LOWESS normalization method. Especially, when there is a strong relationship between the background intensity and the signal intensity, the proposed method performs much better. Regardless of background correction methods used in the image analysis, the proposed two-stage normalization method can be applicable as long as both signal intensity and background intensity are available. PMID- 15268769 TI - The construction of meaning in computational integrative biology. AB - It is the contention of this paper that current data mining work in bioinformatics tends to emphasize data representation to the neglect of another essential aspect of biological systems, namely dynamics. This results in a divorce of both the enterprise and the teaching of bioinformatics from its central aim of meaning-construction. The paper argues that this neglect of dynamics is rooted in an information-processing view of cognitive psychology, and needs to be complemented by a more narrative perspective which emphasizes the explanation, rather than the mere description, of observed patterns in data. This increased emphasis on explanatory narrative in the form of dynamical modeling leads to both a deeper understanding of biological information and a more invigorating approach to the teaching of bioinformatics. The paper presents a cross-curricular teaching framework for a first-year undergraduate course in bioinformatic dynamical modeling which is based around the use of narrative plots. PMID- 15268770 TI - Integrated analysis of microarray data and gene function information. AB - Microarray data should be interpreted in the context of existing biological knowledge. Here we present integrated analysis of microarray data and gene function classification data using homogeneity analysis. Homogeneity analysis is a graphical multivariate statistical method for analyzing categorical data. It converts categorical data into graphical display. By simultaneously quantifying the microarray-derived gene groups and gene function categories, it captures the complex relations between biological information derived from microarray data and the existing knowledge about the gene function. Thus, homogeneity analysis provides a mathematical framework for integrating the analysis of microarray data and the existing biological knowledge. PMID- 15268771 TI - Discrimination models using variance-stabilizing transformation of metabolomic NMR data. AB - After the extensive work that is being done in the areas of genomics, proteomics, and metabolomics, the study of metabolites has come of interest in its own right. Metabolites in biological systems give an understanding of the state of the system and provide a powerful tool for the study of disease and other maladies. Several analytical techniques such as mass spectrometry and high-resolution NMR spectroscopy have been used to study metabolites. The data, however, from these techniques remains quite complex. Traditionally, multivariate analyses have been used for such data. These methods however have an underlying assumption that the data is multivariate normal with a constant variance. This is not necessarily the case. It has been shown that a generalized log transformation renders the variance of the data constant effectively making the data more suitable for multivariate analysis. We demonstrate the effectiveness of these transformations on NMR data taken on a set of 18 abalone that were categorized as either being healthy, stunted, or diseased. We show how the transformation makes multivariate classification of the abalone into the healthy, stunted and diseased categories much more effective and gives a tool for identifying potential metabolic biomarkers for disease. PMID- 15268772 TI - Finding kinetic parameters using text mining. AB - The mathematical modeling and description of complex biological processes has become more and more important over the last years. Systems biology aims at the computational simulation of complex systems, up to whole cell simulations. An essential part focuses on solving a large number of parameterized differential equations. However, measuring those parameters is an expensive task, and finding them in the literature is very laborious. We developed a text mining system that supports researchers in their search for experimentally obtained parameters for kinetic models. Our system classifies full text documents regarding the question whether or not they contain appropriate data using a support vector machine. We evaluated our approach on a manually tagged corpus of 800 documents and found that it outperforms keyword searches in abstracts by a factor of five in terms of precision. PMID- 15268773 TI - Protein folding and unfolding simulations: a new challenge for data mining. AB - One of the unsolved paradigms in molecular biology is the protein folding problem. In recent years, with the identification of several diseases as protein folding disorders and with the explosion of genome information and the need for efficient ways to predict protein structure, protein folding became a central issue in molecular sciences research. Using molecular dynamics unfolding simulations of an amyloidogenic protein--transthyretin--as an example, we put forward a series of ideas on how simulations of this type may be used to infer rules and unfolding behavior in amyloidogenic proteins, and to extrapolate rules for protein folding in different structural classes of proteins. These, in turn, could help in the development of protein structure prediction methods. The need to analyse different proteins and to run multiple simulations creates a huge amount of data which has to be stored, managed, analyzed and shared (database and Grid technology; data mining). Once the data is captured, the next challenge is to find meaningful patterns (associations, correlations, clusters, rules, relationships) among molecular properties, or their relative importance at different stages of the folding or unfolding processes. This clearly puts new and interesting challenges to the bioinformatics community. PMID- 15268774 TI - Inferring a system model of subcellular topoisomerase II beta localization dynamics. AB - Measuring the mobility of proteins in living cells has become critical to many studies in cell biology and forms the basis for discussion on sub-cellular dynamics. Increasingly localization networks are being put together into compartment models to represent the exchange of molecules, represented mathematically as ordinary differential equations (ODE). The set-up is based on published literature, the "knowledge" of the investigator and 3D visualization of the data. Here we demonstrate this method on the example of a simple distribution model of the molecule Topoisomerase II beta (Topo II beta), nuclear protein that modifies DNA topology. It is found in high concentration in the nucleolus and diffuse in the nucleoplasm, demonstrating a non-membranous inhomogeneity in its distribution. We expand on the simple model by adding additional components to fit fluorescence recovery after photobleaching (FRAP) experiments for protein (GFP) labeled Topo II beta to measure its mobility. This model is then validated by comparing it with alternative scenarios and shown to have predictive power. PMID- 15268775 TI - Mining functional modules in genetic networks with decomposable graphical models. AB - In recent years, graphical models have become an increasingly important tool for the structural analysis of genome-wide expression profiles at the systems level. Here we present a new graphical modelling technique, which is based on decomposable graphical models, and apply it to a set of gene expression profiles from acute lymphoblastic leukemia (ALL). The new method explains probabilistic dependencies of expression levels in terms of the concerted action of underlying genetic functional modules, which are represented as so-called "cliques" in the graph. In addition, the method uses continuous-valued (instead of discretized) expression levels, and makes no particular assumption about their probability distribution. We show that the method successfully groups members of known functional modules to cliques. Our method allows the evaluation of the importance of genes for global cellular functions based on both link count and the clique membership count. PMID- 15268776 TI - Initial observations on the effect of medium composition on the differentiation of murine embryonic stem cells to alveolar type II cells. AB - The pluripotency and high proliferative index of embryonic stem (ES) cells make them a good potential source of cells for tissue engineering purposes. We have shown that ES cells can be induced to differentiate in vitro into pulmonary epithelial cells (type II pneumocytes) using a serum-free medium designed for the maintenance of mature distal lung epithelial cells in culture (SAGM). However, the resulting cell cultures were heterogeneous. Our aim in this study was to attempt to increase pneumocyte yield and differentiation state by determining which medium components enhance the differentiation of pneumocytes and modifying the medium accordingly. Quantitative RT-PCR was used to measure changes in the expression of a type II pneumocyte-specific gene, surfactant protein C (SPC), in response to alterations in the cell culture medium. Results suggested that most individual SAGM growth factors were inhibitory for type II pneumocyte differentiation, with the largest increases in SPC expression (approximately threefold) being observed upon removal of retinoic acid and triiodothryonine. However, large standard deviations occurred between replicates, illustrating the highly variable nature of ES cell differentiation. Nevertheless, these observations represent an initial step towards achieving directed differentiation of pneumocytes from stem cells that could lead to their purification for tissue engineering purposes. PMID- 15268778 TI - Synchronization of goat fibroblast cells at quiescent stage and determination of their transition from G0 to G1 by detection of cyclin D1 mRNA. AB - A number of studies have reported that donor cells consisting of serum starved cells, which are assumed to be at quiescence (G0), or non-starved confluent cells or mitotic cells obtained by shake-off, both of which are assumed to be at G1 phase, give better results in nuclear transfer (NT) than cells at other phases of the cell cycle. Whether G0 or G1 cells function better as donor cells is yet to be determined by detailed studies. The aims of this study were to analyze the cell cycle of goat transfected fibroblasts and determine the timing of transition from G0 to G1 by detecting G1-specific marker, cyclin D1 mRNA. Fluorescent activated cell sorting (FACS) analyses of cells after 4 days of serum starvation showed that more that 90% of cells were in G0/G1. Additionally, detection of cyclin D1 mRNA by northern blot analysis showed that 4-day serum starved quiescent cells started entering G1 a few hours after addition of 10% serum to the medium. Taken together, the data indicated that serum starved transfected primary fibroblasts of adult goats experienced the G0 to G1 transition within 5 h of serum stimulation and were at the mid-G1 stage within 10 h of serum stimulation. PMID- 15268779 TI - Comparison of in vitro development of porcine nuclear-transferred oocytes receiving fetal somatic cells by injection and fusion methods. AB - The present study compared the potential of nuclear-transferred porcine oocytes receiving fetal somatic cells by direct injection and cell fusion procedures to develop into blastocysts. After brief treatment of in vitro matured oocytes with demecolcine and sucrose, the protrusion containing the condensed chromosome mass was mechanically removed. Single donor cells were fused with enucleated oocytes following electric pulses or injected into oocytes by piezo-actuated microinjection. The reconstruction rate by direct injection was significantly higher than that following cell fusion (89 vs. 48%). The potential of nuclear transferred oocytes to develop into blastocysts, however, was not different between injection and fusion methods (13% vs. 18%). Total cell number, inner cell mass, and trophectoderm cell numbers of cloned blastocysts were also not different between the two groups. PMID- 15268780 TI - The Assessment of Food Quality from Cloned Animals. Proceedings of a symposium, November 21-23, 2003, Jouy-en-Josas, France. PMID- 15268781 TI - Food consumption risks associated with animal clones: what should be investigated? AB - Somatic Cell Nuclear Transfer (SCNT), or cloning, is likely to be used for the expansion of elite breeding stock of agronomically important livestock used for food. The Center for Veterinary Medicine at the US Food and Drug Administration has been developing a risk assessment to identify hazards and characterize food consumption risks that may result from cloning. The risk assessment is comprised of two prongs. The first evaluates the health of animal clones, and is referred to as the Critical Biological Systems Approach. The second considers the composition of meat and milk from animal clones. Assessing the safety of food products from animal clones and their progeny, at least during these early stages of the development of the technology, is best accomplished by using both approaches: prospectively drawing on our knowledge of biological systems in development and maturation, and in retrograde, from an analysis of food products. Subtle hazards and potential risks that may be posed by animal clones must, however, be considered in the context of other mutations and epigenetic changes that occur in all food animal populations. PMID- 15268782 TI - Health status of cloned cattle at different ages. AB - The procedure of somatic cloning is associated with important losses during pregnancy and in the perinatal period, reducing the overall efficacy to less than 5% in most cases. A mean of 30% of the cloned calves die before reaching 6 months of age with a wide range of pathologies, including, for the most common, respiratory failure, abnormal kidney development, liver steatosis. Heart and liver weight in relation to body weight are also increased. Surviving animals, although mostly clinically normal, differ from controls obtained by artificial insemination (AI) within the first 1-2 months, to become undistinguishable from them thereafter. Hemoglobin concentrations, for instance, are lower, and leptin concentrations are elevated. In response to the lack of prospective studies addressing the health of adult clones, a long-term, 3-4-year study is currently being conducted to assess the health of mature bovine clones at INRA. Preliminary results over 1 year of study do not show any statistical difference between groups for hematological parameters. PMID- 15268783 TI - The health of somatic cell cloned cattle and their offspring. AB - The cloning syndrome is a continuum with the consequences of abnormal reprogramming manifest throughout gestation, the neo-natal period, and into adulthood in the cloned generation, but it does not appear to be transmitted to subsequent offspring following sexual reproduction. Most in vivo studies on bovine somatic cell cloning have focused on development during pregnancy and the neo-natal period. In this paper, we report on the viability and health of cloned cattle in adulthood. From our studies at AgResearch, we find that between weaning and 4 years of age, the annual mortality rate in cattle cloned from somatic cells is at least 8%. Although the reasons for death are variable and some potentially preventable, the main mortality factor in this period is euthanasia due to musculoskeletal abnormalities. This includes animals with severely contracted flexor tendons and those displaying chronic lameness, particularly in milking cows. In contrast, no deaths beyond weaning have so far been encountered with the offspring of clones where the oldest animals are 3 years of age. In surviving cloned cattle, blood profiles and other indicators of general physiological function such as growth rate, reproduction, rearing of offspring, and milk production are all within the normal phenotypic ranges. PMID- 15268784 TI - Zootechnical performance of cloned cattle and offspring: preliminary results. AB - This paper presents information on the evolution of sets of cloned heifers of Holstein breed in comparison to that of control heifers derived from artificial insemination (AI) in the same farm, as well as data on a set of cloned bulls and their semen characteristics. Preliminary observations on a group of calves sired by a cloned bull and offspring of cloned females are reported. Mean birth weight in the clone group (50 females) was statistically higher than that of 68 contemporary female controls obtained by AI (49.27 +/- 10.98 vs. 40.57 +/- 5.55 kg, respectively, p < 0.05). Growth rate was within normal values for Holstein heifers (from 0.7 to 0.8 kg/day) and daily gain was not influenced by the high or low birth weight of clones. Within animals of the same clone, variability of daily gain was reduced compared to their control counterparts. Semen production from three cloned bulls was within the parameters expected for young bull of the same age. A direct comparison of morphological analysis was made between the frozen thawed semen of the donor bull and of his three clones collected at the same age. The overall semen picture appeared within acceptable limits and the clones presented similar percentages of sperm abnormalities (80% of morphologically normal spermatozoa) as the donor. These preliminary results suggest no deleterious effect of cloning on the semen picture of cloned sires. Frozen semen from one clone bull was used for an AI trial, resulting in 65% pregnancies, 25 live calves were naturally delivered. Concerning the offspring of both female and male clones, the phenotypical and clinical observation of the calves in the first week of age did not reveal any clinical abnormality, suggesting that the deviations observed in clones are not transmitted to the progeny. PMID- 15268785 TI - Immune status: normal expression of MHC class I in the placenta and what is expected in clones. AB - Major histocompatibility complex (MHC) class I genes are highly polymorphic, widely expressed genes that play a vital role in immune responses. Because of their involvement in tissue transplant rejection there has been much interest in their expression during pregnancy. This review examines the evidence for their role in livestock reproduction, and speculates that the process of creating clones may modify their expression with deleterious results. PMID- 15268786 TI - Genetic and epigenetic aspects of cloning and potential effects on offspring of cloned mammals. AB - Although the biological mechanisms by which host cytoplasm and donor nuclei interact to produce a developmentally competent reconstructed embryo remain largely unknown, some advances have been made to our understanding of the genetic and epigenetic factors involved in the of reprogramming of the donor nucleus. Genetic alterations, which comprise changes to the genetic information in both the nuclear and cytoplasm compartments, are passed on to subsequent generations at fertilization and are a potential source of variation among cloned animals and their offspring. Apart from the major chromosomal anomalies found in developmentally arrested embryos and fetuses, less detrimental rearrangements and/or mutations are likely to go unnoticed in most donor cell karyotypes, suggesting that such problems could lead to inheritable anomalies among clones and their offspring. Mitochondrial DNA is also relevant to cloning because most animals inherit most or all of their mitochondria from the host oocyte. Epigenetic alterations to the DNA or to the histone packaging proteins are independent of gene sequences. Aberrant epigenetic events may lead to variable gene expression or mitosis and consequent effects on development and phenotype. Although much of the epigenetic marking is reset during embryogenesis and development, the impact of epimutations on progeny remains unexplored. PMID- 15268787 TI - Genetic identity of clones and methods to explore DNA. AB - Cloning by nuclear transfer has made it possible to produce genetically identical animals in terms of nuclear DNA content. Recent molecular biology tools are offering scientific ways to get an insight into the identity issues, by exploring and comparing genomes of cloned animals in order to test their genetic identity and methylation differences. We have initiated a study to compare genomic DNA of bovine adult clones, of normal phenotype. We have used, in parallel, the AFLP technique (amplification fragment length polymorphism) and one of its variant, MSAP (methylation-sensitive amplification polymorphism). We are also investigating other techniques leading to the detection of sequence polymorphisms between two genomes based on genomes hybridisation. We chose the representational difference analysis (RDA) methods that can be combined with mismatch-specific recognition or mismatch binding property of some proteins (CEL I, MutS). We plan to use these RDA methods for genome-wide detection of subtle mutations, then to focus on changes affecting the methylation status of promoting genomic regions in abnormal clones. This will be achieved using MSAP with NotI and applying, in parallel, the RLGS (restriction landmark genome scanning) technique. This study will hopefully improve the molecular and functional characterizations of these "new animals." PMID- 15268788 TI - Gene activation and gene silencing: a subtle equilibrium. AB - The genetic make-up of a cell resides entirely in its DNA. Now that the nucleotide sequence of several genomes has been determined, the major challenging problem is to understand how cell differentiation, proliferation or death are controlled. Major steps include analysis of the determinants of the cell cycle, the unravelling of RNAs and proteins involved in the control of gene expression and the dissection of the protein-destruction machinery. The successive steps to be considered are transcription of RNA on the DNA template, mRNA stabilization or degradation, and mRNA translation and protein localization in the right cell compartment. Gene expression or gene silencing is the result of many DNA-RNA protein interactions and chromatin is among the key regulators of gene expression. Open chromatin (euchromatin) allows expression of the DNA message. This chromatin structure is generally characterized by the presence on the gene promoters of transcription complexes associated with histone acetyltransferases (HATs). On the contrary, closed chromatin (heterochromatin) is poorly acetylated and more condensed. It contains histone deacetylases (HDACs), potentially associated with DNA methyltransferases (DNMTs). DNMT activity leads to methylation and silencing of the DNA. Thus, a major problem in the field of gene regulation resides in understanding chromatin structure at each promoter, a formidable task for the years to come. PMID- 15268789 TI - Maternal-fetal transplacental leakage of mitochondrial DNA in bovine nuclear transfer pregnancies: potential implications for offspring and recipients. AB - The synepitheliochorial placenta of ruminants is constructed of multiple tissue layers that separate maternal and fetal blood. In nuclear transfer cloned ruminants, however, placental anomalies such as abnormal vascular development and hemorrhagic cotyledons have been reported. We have investigated the possible exchange of genetic material between somatic cell nuclear transfer cloned (SCNT) bovine fetuses and recipients at day 80 of gestation using mitochondrial DNA (mtDNA) as a marker. Twenty-three recovered SCNT-fetuses and their recipients were screened for divergent and thus informative mtDNA combinations. Twenty-one fetuses generated by in vitro fertilization (IVF) or multiple ovulation embryo transfer (MOET) and the corresponding recipients served as controls. A search for recipient mtDNA haplotype in DNA extracts from fetal blood by PCR-RFLP analysis revealed three cases of chimerism (two SCNT, one IVF) among a total of 19 informative fetus-recipient pairs (eight SCNT, seven IVF, four MOET). Placental anomalies have also been observed in some IVF fetuses and the present data therefore suggests transplacental leakage of cell components or cells from the recipient into some fetuses generated by in vitro techniques. Further studies are necessary to determine (i) the nature of leaked material, (ii) whether there is bi-directional leakage, and (iii) whether leaked material is present in recipients and calves after parturition, i.e. whether leakage takes place in vivo. If recipients were chimeric for DNA or cells derived from genetically modified SCNT (or IVF) embryos, their subsequent utilization might be affected. PMID- 15268790 TI - Performance of dairy cattle clones and evaluation of their milk composition. AB - Genetic and phenotypic performance of U.S. Holstein embryo-split and nuclear transfer clones was documented for yield and fitness traits. For cows, mean genetic superiority based on pedigree was 186 kg of milk, 9 kg of fat, and 7 kg of protein for embryo-split clones and 165, 10, and 8 kg, respectively, for nuclear-transfer clones compared with the population for the same birth year; pedigree advantage for male clones generally was slightly greater. Estimates of genetic merit that considered a clone's own performance as well as pedigree merit were slightly lower for embryo-split cows than for their full siblings for yield but not for milk composition (fat and protein percentages), mastitis resistance (somatic cell score), longevity (productive life), or cow fertility (daughter pregnancy rate); no corresponding genetic differences were found for nuclear transfer cows or for cloned bulls regardless of clone type. For bulls, estimated genetic merit based on daughter yield was more similar for clone pairs with apparent identical genotype than for pairs from the same biotechnology but nonidentical as confirmed by blood typing. Yield deviations were lower for clones than for their full siblings. Milk composition (total solids, fat, fatty acid profile, lactose, and protein) also was compared for nuclear-transfer clones (Brown Swiss, Holstein, and Holstein-Jersey cross) with non-cloned cows and literature values; no differences were found for gross chemical composition of milk. No obvious differences were evident between cloned and non-cloned animals or for the milk that they produced. PMID- 15268791 TI - Evaluation of meat products from cloned cattle: biological and biochemical properties. AB - Agricultural utilization of cloned livestock produced by nuclear transfer and their products for food will require public and governmental acceptance. A series of studies of properties of meat derived from cloned cattle was carried out to collect data for the safety assessment of cloned cattle products. Meat samples obtained from embryonic cloned, somatic cloned and non-cloned cattle were analyzed for chemical composition, as well as amino acids and fatty acids. Digestibility, allergenicity, and mutagenicity of meat were also examined. There were no significant differences in these properties among embryonic cloned, somatic cloned and non-cloned cattle. The analyses and tests revealed that there were no significant biological differences in meat from a non-cloned, an embryonic cloned, or a somatic cloned animal. A 14-week feeding trial in rats showed there were no abnormalities in body growth, general condition, locomotor activity, reflexes, sexual cycle, urinalysis, hematology, blood biochemistry, and histology. This study showed for the first time that the biological/biochemical properties of meat of cloned cattle are similar to those of non-cloned cattle. PMID- 15268792 TI - Nutritional value of milk and meat products derived from cloning. AB - The development and use of milk and meat products derived from cloning depends on their safety and on the nutritional advantages they can confer to the products as perceived by consumers. The development of such products thus implies (i) to demonstrate their safety and security, (ii) to show that their nutritional value is equivalent to the traditional products, and (iii) to identify the conditions under which cloning could allow additional nutritional and health benefit in comparison to traditional products for the consumers. Both milk and meat products are a source of high quality protein as determined from their protein content and essential amino acid profile. Milk is a source of calcium, phosphorus, zinc, magnesium and vitamin B2 and B12. Meat is a source of iron, zinc and vitamin B12. An important issue regarding the nutritional quality of meat and milk is the level and quality of fat which usually present a high content in saturated fat and some modification of the fat fraction could improve the nutritional quality of the products. The role of the dietary proteins as potential allergens has to be taken into account and an important aspect regarding this question is to evaluate whether the cloning does not produce the appearance of novel allergenic structures. The presence of bio-activities associated to specific components of milk (lactoferrin, immunoglobulins, growth factors, anti-microbial components) also represents a promising development. Preliminary results obtained in rats fed cow's milk or meat-based diets prepared from control animals or from animals derived from cloning did not show any difference between control and cloning derived products. PMID- 15268793 TI - Ethical, legal, and social aspects of farm animal cloning in the 6th Framework Programme for Research. AB - Cloned livestock have potential importance in the provision of improved medicine as well as in the development of livestock production. The public is, however, increasingly concerned about the social and ethical consequences of these advances in knowledge and techniques. There is unevenness throughout Europe in different Member States' attitudes to research into livestock cloning. Although there is EU legislation controlling the use of animals for research purposes, there is no legislation specifically governing cloning in livestock production. The main EU reference is the 9th Opinion of the European Group on Ethics, which states "Cloning of farm animals may prove to be of medical and agricultural as well as economic benefit. It is acceptable only when the aims and methods are ethically justified and when carried out under ethical conditions." The ethical justification includes the avoidance of suffering, the use of the 3Rs principle and a lack of better alternatives. The Commission addresses these issues in the 6th Framework Programme by promoting the integration of ethical, legal and social aspects in all proposals where they are relevant, by fostering ethical awareness and foresight in the proposals, by encouraging public dialogue, and by supporting specific actions to promote the debate. Research must respect fundamental ethical principles, including animal welfare requirements. PMID- 15268794 TI - Public perceptions of reproductive biotechnologies: the case of farm animal breeding and reproduction in France and the United Kingdom. AB - Results of previous qualitative and quantitative stages of the research project demonstrated that, although consumers had poor knowledge about breeding and reproduction procedures, they were concerned about the impact of breeding practices on their food items. They acknowledged breeding and reproduction to be at the very core of animal-based food chain process. Since however modern breeding programmers beg so much for genetics, their practices increasingly raised consumer concerns. This paper presents results of a research addressing this issue and based on interviews of livestock breeders and specialized scientists. This research was undertaken within the frame of an EU funded project (Sustainable Farm Animal Breeding and Reproduction Project, 2000-2003). Interviews were performed according to the methodology of focus groups and results were used to prepare a discussion guide, including definitions of breeding techniques such as artificial insemination, embryo transfer, in vitro fertilization, and molecular genetics. Farm animal breeding and reproduction methods raised high level of concerns in conventional terms like safety, healthiness and quality of food, factory farming and related consequences on environment, international issues, and cost. Several propositions were presented that deal with modern farm animal breeding and reproduction, EU regulation of breeding procedures, education of consumers on breeding methods, and labelling of products on breeding and reproduction grounds. PMID- 15268795 TI - Potential uses of cloning in breeding schemes: dairy cattle. AB - Cloning by nuclear transfer has many potential applications in a dairy cattle breeding program. It can be used to increase the accuracy of selection and therefore the rate of genetic progress, to speed up the dissemination of the genes from animals of exceptionally high genetic merit to the commercial population, and to reproduce transgenic animals. Today, however, the main limitation of the use of cloning besides governmental regulations is its low success rate and consequently the high cost to produce an animal ready for reproduction. As a result cloning is mostly limited to the reproduction of animals of very high genetic merit or that carry genes of specific interest. Examples of this are top-ranked bulls which do not produce enough semen for the demand due to various reasons. A strategy that could be used by artificial insemination (AI) centers would be to create a bank of somatic cells for every bull entering AI facilities long before they are placed on the young sire proving program. The other use of cloning is to assist in the selection and reproduction of bull dams. Marker assisted selection (MAS) can substantially enhance the accuracy of selection for embryos or young animals without comprehensive performance records, and therefore can greatly increase the value of cloning such embryos or young animals. PMID- 15268796 TI - Agro-economic impact of cattle cloning. AB - The purpose of this paper is to review the economic and social implications of cloned cattle, their products, and their offspring as related to production agriculture. Cloning technology in cattle has several applications outside of traditional production agriculture. These applications can include bio-medical applications, such as the production of pharmaceuticals in the blood or milk of transgenic cattle. Cloning may also be useful in the production of research models. These models may or may not include genetic modifications. Uses in agriculture include many applications of the technology. These include making genetic copies of elite seed stock and prize winning show cattle. Other purposes may range from "insurance" to making copies of cattle that have sentimental value, similar to cloning of pets. Increased selection opportunities available with cloning may provide for improvement in genetic gain. The ultimate goal of cloning has often been envisioned as a system for producing quantity and uniformity of the perfect dairy cow. However, only if heritability were 100%, would clone mates have complete uniformity. Changes in the environment may have significant impact on the productivity and longevity of the resulting clones. Changes in consumer preferences and economic input costs may all change the definition of the perfect cow. The cost of producing such animals via cloning must be economically feasible to meet the intended applications. Present inefficiencies limit cloning opportunities to highly valued animals. Improvements are necessary to move the applications toward commercial application. Cloning has additional obstacles to conquer. Social and regulatory acceptance of cloning is paramount to its utilization in production agriculture. Regulatory acceptance will need to address the animal, its products, and its offspring. In summary, cloning is another tool in the animal biotechnology toolbox, which includes artificial insemination, sexing of semen, embryo sexing and in vitro fertilization. While it will not replace any of the above mentioned, its degree of utilization will depend on both improvement in efficiency as well as social and regulatory acceptance. PMID- 15268797 TI - [Hfgl2/fibroleukin expression in liver and peripheral blood mononuclear cells (PBMC) and its correlation with disease severity]. AB - OBJECTIVE: Viral hepatitis remains a major public health problem and the most common type of liver disease worldwide. There are an increasing number of patients with chronic hepatitis B who develop acute hepatitis on chronic condition (AOC) and die of acute hepatic failure both as a result of lack of understanding of the pathogenesis of the disease and lack of effective treatment. The hallmark of AOC is the extreme rapidity of the necromicroinflammatory process resulting in widespread or total hepatocellular necrosis in weeks or even days. Our previous studies have shown in an experimental animal model of fulminant viral hepatitis caused by murine hepatitis virus strain 3, the importance of macrophage activation, and expression of a unique gene mfgl 2 which encodes a serine protease capable of directly cleaving prothrombin to thrombin, resulting in widespread fibrin deposition within the liver and hepatocyte necrosis. The undergoing study in this report is designed to identify the role of hfgl 2 (human fibrinogen like protein 2) /fibroleukin in patients with viral hepatitis. METHODS: Liver tissues were obtained from 23 patients with AOC hepatitis B, and from 13 patients with inactive chronic hepatitis B (CHB) and 14 patients with chronic hepatitis B with cirrhosis during the year of 1995 to the end of 2001. Liver biopsies were performed within 30 min after the patients were diagnosed with death as a result of acute hepatic failure. Liver samples were also obtained from 4 liver donors as normal controls. In addition, peripheral blood mononuclear cells (PBMC) were isolated from 30 patients (unpaired) with AOC hepatitis B and 10 patients with CHB during the May of 2001 to March of 2002 and 10 healthy volunteer as negative control. PBMCs were freshly isolated and smeared on slides and kept at -80 degree C for further use. Histological sections were stained with hemotoxylin and eosin. A 169 bp of hfgl 2 cDNA probe and a polyclonal or monoclonal antibody against hfgl 2 were used to detect the expression of hfgl 2 mRNA and protein in liver samples as well as PBMC by immune histochemistry separately. RESULTS: Liver tissues from the patients with acute on chronic hepatitis had classical pathological features of acute necroinflammation. Hfgl 2 was detected by immune histochemistry in 21 of 23 patients (91.3%) in liver sections from patients with acute on CHB, while only 1 of 13 patients (7.7%) with CHB and cirrhosis and no evidence of active disease had hfgl 2 mRNA or protein expression. 28 of 30 patients (93.3%) with acute on CHB and 1 of 10 with CHB were detected with hfgl 2 expression in PBMC. There was no hfgl 2 expression in either the liver tissue or the PBMC from the normal donors. There was positive correlation of hfgl 2 expression and the severity of the disease displayed by the value of bilirubin and PT. CONCLUSION: The molecular and cellular results reported here in patients with acute on chronic hepatitis and who died of acute hepatic failure correlates with previous report in 8 patients with fulminant hepatic failure (FHF) and mimic closely the changes observed in the murine model of fulminant viral hepatitis in which the pathogenesis of the disease has been studied in a stepwise fashion. This study further suggests that virus induced hfgl 2 prothrombinase/fibroleukin expression and the potent function of the protein it encodes plays a pivotal role in initiating acute severe hepatitis on the baseline of chronic hepatitis. The measurement of hfgl 2/fibroleukin expression in PBMC may serve as a useful marker to monitor the severity of disease in patients with the AOC hepatitis B and a target for therapeutic intervention. PMID- 15268798 TI - [Clinical study on the severe hepatitis with nosocomial fungal infections and risk factors]. AB - OBJECTIVE: To study the nosocomial fungal infections in the patient with severe hepatitis and analyze of risk factor. METHODS: All 115 severe hepatitis with fungal infections inpatients was studied prospectively. RESULTS: We identified 115 cases with fungal infections, the mean age of patients was 37.2+/-21.5 years, male: 49 cases, female 66 cases. Infection of abdominal cavity accounted for 40.9%, infectious rate in respiratory tract and digestive tract were 26.9%, 21.8%, respectively. Candida albicans accounted for 67.6%. Use of broad-spectrum antibiotic and corticosteroids, neutropenia, severity of liver disease, improper medical manipulations as significant risk factors for fungal infection. Death rate of study group and control group was 59.1%, 34.8%, respectively (x2=36.0). In multivariate analysis, neutropenia, disseminated infection and severity of liver diseases were independent prognostic factors. CONCLUSION: Identification of risk factors and predictors of a poor outcome in patients with severe hepatitis with fungal infections, it suggested that implications in prophylaxis of fungal infection, early diagnosis and appropriate therapy would be important for these patients. PMID- 15268799 TI - [The inhibitory effects on hepatitis B virus replication by stable expression of DN mutants of hepatitis B virus X gene pRev X-GFP]. AB - OBJECTIVE: Persistent replication of hepatitis B virus (HBV) is one of the major obstacles in HBV infection treatment. Reduction or clearance of HBV propagation would be one of the aims of HBV therapy. The drugs approved in clinical used such as nucleotide analogs or interferon, were limited effects on HBV replication. The newly developing gene therapy method, dominant negative mutants, were be used as new promising HBV therapy strategy, and a dominant negative mutant of HBVX gene pRev X-GFP which we have reported in our previous study has some effects both on HBV replication and expression in transient expression, but the effects were interfered by persistent secretion of HBV in HepG2 2.2.15 cells without transfection pRev X-GFP in the experiment. To make sure the effects of dominant negative mutant of pRev X-GFP, we established a HBX DN stable express cell clone, and evaluated the effects of HBX dominant negative mutant on HBV replication. METHODS: The X gene mutant, in which a specific point mutation of 3'-end ATG to AAG and fused with human green fluorescence protein (GFP) were cloned into pRev TRE vector, assigned to pRev HBX-GFP dominant mutant (pRev X-GFP). And the plasmid contains the wild type X gene or GFP gene was cloned into the same vector to construct pRev Xwt, pRev GFP constructs. All the constructs then transfected into HepG2 2.2.15 cells by liposome. After 7 days resistance selection of hygromycin (300 microg/ml), and cell clones which stable expression HBX-GFP, HBXwt, GFP were obtained. After reseeding of 106 cells of each clones in 12 wells with a 12 well cell plate and another 12 wells 2.2.15 cell were serve as blank control. The cells and media were harvested after cultured in DMEM with 10% FBS for 3 days. HBV-related DNA was assayed by dot blot and Southern blot. RESULTS: The 100% expression of pRev HBX-GFP, GFP and wild type X constructs were obtained. The stable expressed HBX-GFP can significantly reduce HBV DNA level both in cell media and cells by dot blot and Southern blot analysis, but not for pRev Xwt and pRev GFP. CONCLUSION: The dominant negative mutant pRev HBX-GFP can significantly inhibit the HBV gene expression. It also suggested that X gene might be one of promising target for HBV gene therapy. PMID- 15268800 TI - [Detection and analysis of HAV-HEV, HGV infection in patients with viral hepatitis]. AB - OBJECTIVE: To study the simple infection and super/co-infection of HAV-HEV, HGV in patients with viral hepatitis. METHODS: Using EIA method to detect anti-HAV IgM, HBV serum markers, anti-HCV IgM, anti-HDV IgM, anti-HEV IgM, anti-HGV IgM in viral hepatitis patients with different clinical types. RESULTS: Seventy-three percent patients (154/210) had HBV infection markers, twenty-nine percent patients (61/210) had HAV infection marker, eight percent patients (17/210) had HCV, HDV infection markers, ten percent patients (21/210) had HEV infection and seven percent patients (15/210) had HGV infection. Only nine percent patients (20/210) had viral hepatitis serum markers negative. In all clinical types, sixty one percent patients had only one type hepatitis virus infection, thirty-two percent patients had two types of hepatitis virus super/co-infection, six percent patients had three types of hepatitis virus super/co-infection. Super/co infection often occurred in patients who had cirrhosis or hepatic failure. CONCLUSION: HBV and HAV infection is very common in viral hepatitis patients, whereas HCV, HDV, HEV and HGV infection is relatively low; double super/co infection of HAV-HEV, HGV frequently occurs in severe patients with viral hepatitis. PMID- 15268801 TI - [A preliminary investigation of nuclear factor kappa B in peripheral blood lymphocytes of patients with hepatitis B]. AB - OBJECTIVE: To explore the activity of nuclear factor kappa B (NF-kappaB) in peripheral blood lymphocytes (PBL) of patients with hepatitis B. METHODS: The PBL of patients with different types of hepatitis B and healthy individuals were isolated and then the nuclear extract was prepared. Assessment of NF-kappaB DNA binding activity was performed by electrophoretic mobility shift assay (EMSA) using digoxin labeled double-stranded oligonucleotide containing kappa B consensus sequence. RESULTS: Densitometric scanning of the EMSA bands showed that the mean optical densities (A) from the groups of normal control, acute hepatitis B, chronic hepatitis B and chronic severe hepatitis B were 20.18+/-2.16, 27.75+/ 4.11, 13.90+/-3.20 and 8.02+/-2.65 respectively. Analysis of variance showed the F value was 26.112 and the difference among the groups was significant. The difference between two groups with Dunnett T3 analysis showed there are statistically difference between the groups of the normal group and the chronic severe hepatitis B group, the acute hepatitis B group and the chronic hepatitis B group, and the acute hepatitis B group and the chronic severe hepatitis B. CONCLUSION: The activity of NF-kappaB in PBL of patients with hepatitis B is related with the different outcomes of hepatitis B virus (HBV) infection. Decreased activity of NF-kappaB may be an important cause for the dysfunction of PBL in chronic HBV-infected patients. PMID- 15268802 TI - [The plasma levels of transforming growth factor beta1 and the protein expressions of alpha-SMA, urokinase plasminogen activator and plasminogen activator inhibitor-1 in liver of patients with different grades of hepatic fibrosis]. AB - OBJECTIVE: To measure the plasma levels of transforming growth factor beta1 (TGFbeta1), the protein expression of alpha-SMA in hepatic stellate cells and urokinase plasminogen activator (uPA) and plasminogen activator inhibitor-1 (PAI 1), and study on the relationships between the plasma levels of TGFbeta1, the protein expression and the serum hyaluronic acid (HA) in patients with different grades of hepatic fibrosis. METHODS: Thirty seven cases with hepatic fibrosis of different grades were classified according to HE and VG staining categories from 0 to 4, in which there were 8 cases in grade 1, 9 cases in grade 2, 7 cases in grade 3, 13 cases in grade 4. The plasma levels of TGFbeta1 and the serum levels of HA were detected by ELISA. The protein expressions of a-SMA, uPA and PAI-1 in fibrotic liver tissue were observed by immunohistochemistry. RESULTS: With the progression of hepatic fibrosis, the plasma levels of TGFbeta1 and the protein expression of a-SMA, uPA and PAI-1 in fibrotic liver tissue were increased. In grade 3 and 4, the plasma levels of TGFbeta and the protein expression of a-SMA and PAI-1 in fibrotic liver tissue were significantly increased, but the protein expression of uPA in cirrhosis liver tissue did not increased. CONCLUSION: TGFbeta1, a-SMA, uPA and PAI-1 play an important role in the progression of hepatic fibrosis. Inhibiting the early activation of latent TGFbeta1 or increasing uPA and inhibiting PAI-1 over express may contribute to matrix degradation and retard the progression of hepatic fibrosis. PMID- 15268803 TI - [Interaction effect of p53 with HBV in hepatoma cell line SMMC-7721]. AB - OBJECTIVES: To study the interaction of hepatitis virus B (HBV) and tumor suppressor p53. METHODS: Plasmid pCMVp53 was transfected or cotransfected with pCMVHBVa (wild type HBV) or PCMVHBVb (mutant type HBV) into the hepatoma cell line SMMC-7721 by lipofectamine. Apoptosis cells were labeled by annexin V-FITC and confirmed by flow cytometry. Reporter plasmids PG13-CAT or p21-luc were cotransfected respectively in each group to indicate transactivation activity of p53 and it's effect on p21 promoter. Western blot was performed to observe p53 expression in each group. RESULTS: The group transfected by pCMVp53 alone exhibit higher luciferase activity and higher apoptosis rate, otherwise, p53 expression, enzyme activity of PG13-CAT or p21- luc and cell apoptosis rate were much higher in the group cotransfected by pCMVp53 and pCMVHBVa, but not in the other cotransfected group; HBV replication was enhanced in p53 cotransfected group. CONCLUSION: p53 expression and effects could be enhanced by HBV and p53 had positive regulation effect on HBV replication. PMID- 15268804 TI - [Effects of growth factors and extracellular matrix on proliferation and differentiation of fetal liver progenitor cell in vitro]. AB - OBJECTIVE: Liver development needs a number of growth factors and components of the extracellular matrix. The study is to explore how growth factors and extracellular matrix regulate proliferation and differentiation of fetal liver progenitor cell. METHODS: We demonstrate isolation of hepatic progenitor/stem cells from ED 14.5 SD rat liver, which contains a large number of hepatoblasts. Proliferation assay-3H thymidine incorporation was used to detect the effect of growth factors on proliferation of hepatic progenitor cell. Growth factor and extracellular matrix were added and stem cell clone formation was counted. Mark of bile duct and hepatocyte were detected with double-marker immunocytochemistry. RESULTS: Progenitor liver cells displayed clonogenic capacity, expressed markers of hepatocytes and bile duct cells and G-6-P. HGF, EGF can accelerate DNA synthesis and stem cell clone formation of hepatic progenitor cell. Extracellular matrix collagen I, collagen IV or laminin were essential for formation of stem cell clone. Single cell culture needed HGF, EGF, extracellular matrix and supernatant of mix cell (which contained fetal parenchymal cells, mesenchymal cells and hematopoietic cells) culture. CONCLUSION: Growth factors especially HGF and EGF play crucial role in proliferation and differentiation of liver progenitor cell. Some factors secreted from mesenchymal cell and hematopoietic cells may be involved. PMID- 15268805 TI - [Serum from partial hepatectomy rat and hepatocyte growth factor stimulate bone marrow cell expressing albumin and alpha fetoprotein]. AB - OBJECTIVE: To explore the effect of serum from partial hepatectomy (PH) rat and hepatocyte growth factor (HGF) on expression of albumin and AFP of bone marrow cells. METHODS: The bone marrow mono-nucleated cells were separated from SD rats and cultured in three groups: (1) The medium only group as control was added normal fetal bovine serum; (2) Rat hepatic injury serum group (was added 15% rat serum from 2-AAF+PH model); (3) HGF group (HGF 20 ng/ml). The role of these factors was determined by RT-PCR, immunohistochemistry (IHC) and Western blot, using AFP and albumin as special hepatocytic markers. RESULTS: By immunohistochemical staining and Western blot, the fresh bone marrow cells were AFP-negative, same as the cells cultured with medium only group. While bone marrow cells, co-cultured with rat hepatic injury serum or HGF at day 10 and 20, expressed AFP protein. AFP mRNA expression could be found in bone marrow cells after 10 and 20 days cultured with rat hepatic injury serum or HGF, but not in fresh bone marrow cells and bone marrow cells cultured with medium only. Albumin mRNA expression was weak in fresh bone marrow cell and increased in groups 2 and 3. CONCLUSION: The rat hepatic injury serum or HGF could stimulate the expression of AFP protein and it's mRNA of bone marrow cells. Also they can stimulate albumin mRNA expression. It seems that, in bone marrow, there is a kind of cells so called bone marrow derived liver stem cell which can express albumin mRNA in a weak style. PMID- 15268806 TI - [Associated risk factors of fatty liver in the patients with type 2 diabetes mellitus]. AB - OBJECTIVE: To analyze the associated risk factors, clinical characteristics and laboratory abnormalities of type 2 diabetes patients with fatty liver. METHODS: The data of type 2 diabetes cases with fatty liver were collected in our hospital. 63 cases of type 2 diabetes without fatty liver were selected randomly as control during the same period. The associated variables were analyzed by using logistic regression model. The clinical data and liver function were compared between two groups. RESULTS: The proportion of obesity and hyperlipidemia was higher in type 2 diabetes patients with fatty liver than without fatty liver. Body mass index (BMI) (OR: 4.392) was positive correlation to fatty liver in the patients with type 2 diabetes. In contrast, insulin sensitivity index (ISI) (OR: 0.000) and regular insulin treatment (OR: 0.058) were negative correlation to it. The abnormal frequencies of aspartate aminotransferase (AST, 16.0%), alanine aminotransferase (ALT, 25.2%), the ratio of AST/ALT less than 1 (52.8%) and gamma-glutamyltransferase (GGT, 31.9%) of type 2 diabetes patients with fatty liver were significantly higher than those without fatty liver (3.2%, 6.4%, 36.5% and 11.1% respectively). CONCLUSION: Obesity and insulin resistance might increase the risk of fatty liver in the patients with type 2 diabetes. Patients of type 2 diabetes with fatty liver show higher serum lipid level and more obvious damages of liver function than those without fatty liver PMID- 15268807 TI - [Hypoxia inducible factor 1-alpha mRNA expression in alcoholic liver disease]. AB - OBJECTIVE: To investigate the effect of hypoxia on chronic alcoholic liver disease. METHODS: Twenty four male Sprague-Dawley rats were randomly into two groups. The alcohol group (n=12) was fed 56% (v/v) of ethanol once per day by gastric infusion at 8 g/kg body weight for 24 weeks. The control group (n=12) was gastrically infused with normal saline with the same dose. At the end of 24 weeks, a blood sample was collected for determination of hepatic enzymes and then the rat was killed. Liver specimens were obtained for immunohistochemical staining and frozen at -80 degrees C used for RT-PCR. RESULTS: Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activity increased significantly compared to the control group. A significant elevation in the expression of HIF1-alpha in liver of alcohol group was found compared to the control group. CONCLUSION: Hypoxia inducible factor 1-alpha expression was activated by ethanol-induced injury. This information suggested that hypoxia was involved in mechanism of alcoholic liver disease. PMID- 15268808 TI - [Tolerance enhancement on the liver ischemia-reperfusion injuries in rats by ethanol pretreatment]. AB - OBJECTIVE: To investigate the feasibility and efficacy of the ethanol pretreatment. Study was designed to confirm the proper range of the ethanol according to the toxicity and mortality, and then evaluate the possibility of application of the ethanol pretreatment. METHODS: (1) Thirty six male adult wistar rats pretreated with 40% ethanol were divided randomizely into six groups by different dosage: group A (8 g/kg), group B (7 g/kg), group C (6 g/kg), group D (5 g/kg), group E (4 g/kg), normal control group (0 g/kg). The safe dosage range of ethanol in rats was predicted by the observation of the symptoms after ethanol administration and pathological changes after 24 h. (2) Based on the results of experiment (1), this experiment were set as follows: 78 wistar rats were divided randomizely into 4 groups: normal control group, ethanol group, ischemia/reperfusion group (IR), ethanol pretreatment group (EP), in each group, the specimen were harvested from the rats at 3, 6, 12, 24 h after reperfusion and then were determined by different methods. (3) Based on the three variant factors (concentration, dosage and proper time for ethanol pretreatment), a orthogonal test were designed to optimize the ethanol pretreatment. 54 wistar rats used in this step were all subjected to hepatic schema procedure for 90 min and the specimens were harvested at 24 h after reperfusion. RESULTS: Less than 5 g/kg ethanol is safe to the rat, and it can reduce the 90 minutes IR injuries to the liver. Under the mode of A1B1C3, the more protection can be got for hepatic ischemia/reperfusion injuries. CONCLUSION: Proper dose of ethanol gavages to the rat is a safe pretreatment method, it maybe enhance the tolerance of rat liver to the I/R injuries. PMID- 15268809 TI - [The influence of HCV core protein and apoptosis on cellular telomerase activities]. PMID- 15268810 TI - [Efficacy and safety in chronic hepatitis B adolescent patients with lamivudine therapy]. AB - OBJECTIVE: To analysis the efficacy and safety of lamivudine (made in China) therapy for 52 weeks in adolescent patients with chronic hepatitis B (CHB). METHODS: One hundred and five teenage CHB patients were treated with lamivudine 100 mg once daily for 52 weeks. Patients with elevated ALT at baseline were in group 1 and those with normal ALT were in group 2. The changes of HBV DNA, HBV seromarkers and ALT at the end of 12, 24 and 52 weeks after lamivudine therapy were compared with those at baseline. Adverse events were recorded and evaluated. RESULTS: At the end of 52 weeks of lamivudine therapy, HBV DNA-ve, HBeAg loss and anti-HBe seroconversion were observed in 92.0%, 24.4% and 22.0% in group 1 patients and 76.1%, 14.2% and 14.2% in group 2 patients respectively. No significant differences were found between two groups. At 12, 24 and 52 weeks, normalization rates of ALT were 59.0%, 66.7% and 76.0%, normal ALT with undetectable HBV DNA were 44.9%, 64.1% and 70.7% at the same time. During 52 weeks lamivudine treatment 26 mild adverse events were observed in 18 patients. CONCLUSION: Lamivudine can inhibit HBV replication rapidly and normalize ALT in majority adolescent CHB patients. HBeAg loss or seroconversion of anti-HBe was observed in some of these patients. All patients in this study were safety and well tolerated. PMID- 15268811 TI - [Obesity and liver fibrosis]. PMID- 15268812 TI - [Common biochemical changes in obesity related liver diseases]. PMID- 15268813 TI - [Clinical and histological features of the patients with hepatitis B recurrence after allo-genetic hemopoietic stem cell transplantation]. PMID- 15268814 TI - [Relationship between alleles of HLA-DRB and HLA-DQB1 and Chinese patients with primary biliary cirrhosis]. PMID- 15268815 TI - [The expression of PPAR gamma in experimental fatty liver disease]. PMID- 15268816 TI - [Identification of HBV infection-associated genes in first trimester human fetal hepatocytes with suppression subtractive hybridization]. PMID- 15268817 TI - [The dynamic change of liver injury in patients with severe acute respiratory syndrome]. PMID- 15268818 TI - [HBx can facilitate the expression of vascular endothelial growth factor in human hepatoma tissue of nude mice]. PMID- 15268819 TI - [Changes of vascular endothelial growth factor and endothelin-1 and nitric oxide in patients with primary hepatocellular carcinoma]. PMID- 15268820 TI - [Analysis of dangerous factors for alcoholic liver disease]. PMID- 15268821 TI - [Study on the detection of duck hepatitis B virus with SYBR based quantitative real-time PCR]. PMID- 15268822 TI - [Classification and diagnostic strategies of drug-induced hepatic disease]. PMID- 15268823 TI - [SEN virus, a recently discovered hepatitis viruses]. PMID- 15268825 TI - [Present status and developmental trend of retinoblastoma research in China]. PMID- 15268826 TI - [Clinical and pathological features of 273 cases of lacrimal epithelial tumors]. AB - OBJECTIVE: A retrospective study of histopathological and clinical aspects of 261 cases of lacrimal tumors (273 paraffin specimens) from Ophthalmic Pathology Laboratory, Beijing Tongren Hospital (Dec. 1961 to Jun. 2002) was performed to investigate the pathological classification and clinical features of these tumors. METHODS: Clinical features were analyzed from 261 patients of lacrimal tumors, including patient's history, age, gender, laterality, visual acuity, ultrasound B scan, X-ray, CT, MRI, pre- and post-operation condition, follow-up data and others. These data were compared with the pathological features. RESULTS: Pleomorphic adenoma was the most common tumor in epithelial tumors of lacrimal gland, followed by adenoid cystic carcinoma, pleomorphic adenocarcinoma and adenocarcinoma. Recurrence was observed in 12 cases. The recurrence rate is 4.6%. Four cases was dead, the mortality was 1.5%. CONCLUSION: Pleomorphic adenoma shows a low mortality and a relatively high recurrence rate. The main treatment for pleomorphic adenoma is surgical treatment. It is important to have a correct clinical diagnosis, to select the proper surgical method, drug treatment and postoperative management based on the pathological diagnosis, which can result in a decrease of recurrence rate. PMID- 15268827 TI - [A new orthotopic retinoblastoma model expressing green fluorescent protein]. AB - OBJECTIVE: In order to observe the growth and metastasis of the tumor directly, a new orthotopic retinoblastoma model was established with human RB cells expressing green fluorescent protein (GFP). METHODS: pEGFP-N(1), the eukaryotic expressive plasmid of GFP, was transferred into human RB cell line HXO-RB(44) by liposome Dosper. Then, the cell clones expressing GFP steadily were selected by means of neomycin, fluorescence microscope, and flow cytometer. Two microliters of RB cells (density at 4.5 x 10(8) - 5.5 x 10(8) cells per ml) were injected into the subretinal space of 30 nude mice (60 eyes) under binocular operating microscope. The growth of transplanted RB was observed in vivo using fluorescence stereomicroscope. The nude mice were killed at different times post-operatively to investigate the metastasis process of the tumor to the optic nerve, brain, and other organs including lung, liver and kidney. RESULTS: The spread process of the tumor in the subretinal space was successfully observed under stereomicroscope. Transplanted RB developed into extra-ocular stage phase at 34 - 37 days after the operation. And metastasis to the cranium along the optic nerve was observed, with green RB cells distributing along the optic nerve sheath and long posterior ciliary artery. The histopathological characteristics of the transplanted tumor were similar to the human RB. Immunohistochemical staining showed positive expression of GFP in the tumor cells. CONCLUSION: The established orthotopic RB model expressing GFP via injection of human RB cells into the subretinal space of nude mice provides a new approach to exploring the growth and metastasis processes of RB in natural situations. PMID- 15268828 TI - [An experiment study of in vitro induced antitumor immune responses by vaccination of tumor lysate-pulsed dendritic cells to kill SO-RB(50)]. AB - OBJECTIVE: To investigate induced antitumor immune responses by vaccination of tumor lysate-pulsed dendritic cells (DC) to kill retinoblastoma cells SO-RB(50). We hope to offer new approach for the treatment of patients with retinoblastoma. METHODS: DC was pulsed with RB tumor lysates in vitro and incubated with autologous lymphocytes to induce antigen specific CTL (cytotoxic T lymphocyte, CTL). SO-RB(50) cells were used as target cells and Raji cells were used as control target cells. Cytotoxicity of CTL was evaluated by MTT method (methyl thiazolyl letrazolium). The specific cytotoxicity of CTL to SO-RB(50) and Raji cells was compared. The cytotoxicity of CTL from RB and normal subjects was compared between these two groups. RESULTS: Antigen specific CTL showed greater cytotoxicity to SO-RB(50) than Raji cells, the difference was statistically significant, P < 0.01. The cytotoxicity was dose-dependent to the ratio of CTL/target cell. The nonspecific cytotoxicity to Raji cells was the same in CTL from RB patients and normal subjects, P > 0.05. The specific cytotoxicity of CTL from RB patients to SO-RB(50) was weaker than that from the healthy subjects, P < 0.01. CONCLUSION: DC pulsed with RB tumor lysate in vitro can induce antigen specific CTL which can kill the SO-RB(50) target cells specifically. This method may have potential value of therapy for the RB patients. PMID- 15268829 TI - [The study on the correlation between telomerase and histopathologic features of retinoblastoma]. AB - OBJECTIVE: To investigate the correlation between the expression of human telomerase RNA (hTR), human telomerase reverse transcriptase (hTRT) and the proliferation and differentiation of retinoblastoma (RB). METHODS: In 37 RB specimens, the expression of hTR and hTRT was investigated with the methods of in situ hybridization (ISH). Immunohistochemical method was performed to detect the expression of proliferating cell nuclear antigen (PCNA) in RB. All data were analyzed statistically with SPSS. RESULTS: Of 37 RB specimens, positive expression of hTR and hTRT was 83.8% and 89.2%, respectively, while negative results were observed in 2 normal retina tissues. The expressions of hTR and hTRT showed a close correlation to the proliferation and differentiation of RB. Furthermore, there was a close correlation among the expression of hTR and hTRT. CONCLUSIONS: These results suggest that higher expression of hTRT and hTR may be used as new parameters for evaluating the degree of malignancy of RB. The close correlation between the expression of hTR and hTRT and proliferation and differentiation degrees of RB indicats that the expression of telomerase can influence the proliferation and differentiation of RB. Because the telomerase is presented in the Rb, therefore, modulating the telomerase activity may be used as a novel treatment for the RB. PMID- 15268830 TI - [Phacoemulsification using burst mode]. AB - OBJECTIVE: To observe the effect of burst mode combined with high vacuum on phacoemulsification. METHODS: Nine hundreds and fifty-four eyes (819 patients) were randomly divided into three groups (continue group, pulse group and burst group). Each group consisted of 318 eyes. A temporal clear corneal incision was performed in each group. Various phaco modes were used. In continue group, the vacuum was 160 mm Hg, max phaco power was 60%. In pulse group, vacuum was 160 mm Hg and the power was 60%. In burst group, vacuum was 250 - 300 mm Hg and the power was 60%. Actual phaco power, phaco time, visual acuity, corneal endothelial cell loss and other complications were recorded. RESULTS: The actual phaco power in these 3 groups was 25.36% +/- 6.65%, 19.54% +/- 3.50% and 6.27% +/- 1.27%, respectively. Phaco time in these 3 groups was (222 +/- 30), (186 +/- 41) and (36 +/- 6) seconds, respectively. In comparison of the power and time among these 3 groups, the difference between continue group and pulse or burst group was significant (P = 0.005 and 0.0001). Visual acuity was significant improved in all 3 groups postoperatively. The difference between the continue group and the pulse group was not significant. The difference between the continue group and burst group was statistically significant. There was no statistically significant difference in mean number of corneal endothelial cell between these 3 groups preoperatively. Endothelial cell loss postoperatively was lower in the pulse and burst groups than that in the continue group. The difference was significant between the continue group and pulse or burst group (P = 0.005 and 0.001). Serious corneal edema occurred more frequently in the continues group than that in the pulse and burst groups. CONCLUSION: High vacuum combined with pulse or burst phaco mode reduced phaco time and actual phaco power consumed during phacoemulsification, especially in the burst mode, which enhanced the effectiveness of phaco chop and reduced the loss of corneal endothelial cell and corneal edema. It is safe to use the burst phacoemulsification clinically. PMID- 15268831 TI - [The effect of weakening inferior oblique muscles on the status of ocular torsion]. AB - OBJECTIVE: To understand the clinical significance of subjective and objective examination of cyclodeviations and investigate the effect of weakening inferior oblique muscles on the status of ocular torsion. METHODS: Twenty patients (40 eyes) with overacting inferior oblique muscles underwent bilateral myotomy or partial myectomy of inferior oblique muscles. Subjective cyclodeviations were measured before surgery as well as one week, two months after surgery by double maddox rod test (DMRT). Objective cyclodeviations were assessed by fundus photography before and 7 days after surgery in 15 cases. The photograph was transferred to a computer, and the fovea-disc angle was measured by means of drawing picture software. RESULTS: Nine patients with primary overaction of inferior oblique muscles were negative with DMRT before surgery, and only one case revealed incyclotropia 5.0 degrees tested two months after surgery. Four out of 11 patients with secondary overaction of inferior oblique muscles showed excyclotropia 2.5 degrees - 5.0 degrees with DMRT before surgery, and all patients indicated no subjective cyclotropia with DMRT two months after surgery. Fundus photography determination of the right eye revealed extorsion 16.83 degrees +/- 6.39 degrees, the left eye 14.92 degrees +/- 4.51 degrees before surgery. The reduction of the cyclodeviations by weakening inferior oblique muscle for the right and left eye was 13.07 degrees +/- 3.38 degrees and 10.54 degrees +/- 3.75 degrees respectively. The comparison of objective ocular torsion for both eyes showed high significant differences (P < 0.01) pre- and post operatively. CONCLUSIONS: The results of subjective and objective examination of cyclodeviations in patients with primary or secondary overacting inferior oblique muscle early after birth were not consistent. Weakening inferior oblique muscle could correct excyclodeviation. There exist complicated compensatory mechanisms for subjective change of ocular torsion after surgery. The changes of subjective and objective cyclodeviations are still inconsistent. PMID- 15268832 TI - [The results and analysis of making corneal flaps with Moria M(2) microkeratome in LASIK]. AB - OBJECTIVE: To investigate the results and technology of making corneal flaps with Moria M(2) microkeratome. METHODS: Eight hundred and six corneal flaps (in 409 cases) were made using Moria M(2) microkeratome in LASIK. The group consisted of 205 males (405 eyes), whose mean age was (23.61 +/- 5.44) years, mean refractive spherical equivalent was (-6.32 +/- 3.61) D, and mean corneal thickness was (542.72 +/- 29.54) micro m. The 130-micron head option was chosen, the pressure was less than 250 mm Hg (1 mm Hg = 0.133 kPa), the stop-ring was adjusted to the 8.0 mm position and the suction ring was chosen according to corneal curvatures. RESULTS: All 806 corneal flaps were made successfully at one time with suitable hinge size, and without ruptured or thin or incomplete flaps. Flaps could be turned and reattached easily. There was no breaking down during the surgical procedure. The flaps showed smooth edges with smooth keratectomy beds. COMPLICATIONS: (1) Free flaps occurred in 3 eyes (0.37%). (2) Island phenomenon occurred in 2 eyes (0.25%). (3) Failure to make the flaps occurred in 3 eyes (0.37%). CONCLUSION: All 806 corneal flaps were made successfully at one time with no serious complications. Choosing the suction ring according to corneal curvature, checking blades preoperatively and inserting blades correctly may help to avoid complications when making corneal flaps with Moria M(2) microkeratome. PMID- 15268833 TI - [A comparison study of pulsitile ocular blood flow in normal eyes and primary open angle glaucoma]. AB - OBJECTIVE: To compare pulsatile ocular blood flow (POBF) and intraocular pressure (IOP) in primary open angle glaucoma (POAG) and normal control group matched for age, sex and refraction error, to investigate the rule of hem-dynamics in POAG and to determine the sensitivity and specificity of POBF measurement as a diagnostic test for glaucoma. METHODS: Prior to the test a questionnaire was completed to determine age, sex, refractive error, family history of glaucoma, history of eye diseases, ocular medication, medical history and using of systemic beta-blockers. Patients of POAG were determined by following diagnostics standards: (1) Three IOP >25 mm Hg in different times of one day. (2) The fluctuate of IOP > 8 mm Hg during 24 hours. (3) Typical glaucoma changes in the visual field. (4) Typical glaucoma changes in optic disc. There were 100 POAG subjects with single eye observed (50 male and 50 female). We picked up 100 eyes randomly (50 male and 50 female) in 534 normal persons who matched for following conditions: (1) Sex. (2) Discrepancy of age less than 5 years. (3) Discrepancy of the refraction error less than +/- 2.00 DS. as the normal comparison group. The tonometer used was the POBF Tonometry. Pulse amplitude of IOP (PA), pulsatile ocular blood flow (POBF), pulse/heart rate (PR), maximum-IOP (max-IOP), minimum IOP (min-IOP) and average IOP (aver-IOP) were obtained before the medical therapy and 1 or 2 weeks after the operation. The correlation between the POBF & mean value of the perimeter was analyzed. POBF was analyzed to determine the sensitivity and specificity of POBF measurement as a diagnostic test. RESULTS: The value of POBF in POAG and normal control was (9.72 +/- 3.47) microl/s and (12.04 +/- 4.68) microl/s, respectively. POAG patients' POBF, PV, PA, and AVE-IOP were less than those in the normal control, and the difference was statistically significant. There was no statistically significant correlation between the changes of visual field and POBF (r = 0.224, P = 0.219). The sensitivity and specificity of using the POBF for diagnosis of POAG (less than 10.75 microl/s being abnormal) were 0.422 and 0.623, respectively. CONCLUSION: Abnormal of vas autoregulation and blood supply play a role in the pathogenesis of POA. Due to the lower sensitivity and specificity, it is not suitable to use POBF as a diagnostic test to distinguish the POAG and normal. But the POBF reflects the change of blood flow in a pulsing cycle and provide more information than the measurement of IOP only. PMID- 15268834 TI - [The inhibitory effect of tranilast on transforming growth factor-beta(2) expression in cultured human trabecular meshwork cells]. AB - OBJECTIVE: To investigate the effect of tranilast on transforming growth factor beta(2) (TGF-beta(2)) expression in cultured human trabecular meshwork cells. METHODS: TGF-beta(2) expression in cultured 3-5 passage human trabecular meshwork cells was measured by semi-quantitative RT-PCR after treated with 0.0 micro g/ml (control), 12.5, 25.0 and 50.0 mg/L tranilast for 48 h. RESULTS: The value of TGF beta(2)/G3PDH of cells treated with 12.5, 25.0 and 50.0 mg/L tranilast was 1.85 +/- 0.35, 1.66 +/- 0.42, 1.16 +/- 0.24, respectively. The difference between these treated groups and that of the control group (3.82 +/- 0.56) was statistically significant (q' = 10.77, 11.80, 14.54, P < 0.01), respectively. The value of TGF-beta(2)/G3PDH in the tranilast treated trabecular meshwork cells decreased in a dose-dependent manner. CONCLUSION: Tranilast could inhibit TGF beta(2) expression in cultured human trabecular meshwork cells. It is worth to study the using of tranilast in the treatment of primary open-angle glaucoma. PMID- 15268835 TI - [The study of photorefractive keratectomy induced defocus on emmetropization in infant monkeys]. AB - OBJECTIVE: To investigate the effect of photorefractive keratectomy (PRK) induced defocus on emmetropization in infant rhesus monkeys and to determine the role of visual feedback on emmetropization. METHODS: PRK was performed in 8 healthy rhesus monkeys age ranging from 2 to 3 months. Either 3.00 D of relative hyperopic or myopic defocus was produced in one eye and the fellow eye as controls. The eyes were examined periodically with corneal topography, cycloplegic retinoscopy, A-scan ultrasonography, and slit-lamp biomicroscopy. RESULTS: All operated eyes demonstrated re-epithelialization within 3 days post surgically and maintained transparent. Compensating axial length growth occurred throughout the observation period. The rate of vitreous chamber elongation varied with the post-PRK refractive status of the eye. The hyperopia-induced eyes exhibited a vitreous chamber elongation faster than the fellow eyes (t = 3.656, P = 0.0354), whereas myopia-induced eyes demonstrated a vitreous chamber elongation slower than the fellow eyes (t = 3.576, P = 0.0374). CONCLUSIONS: PRK-induced defocus altered axial growth rates and emmetropization in infant monkeys predictably. These results indicate that primate emmetropization is regulated by visual feedback and correction of refractive error on children should be careful. PRK-induced-defocus is a promising method for myopic animal model and for the investigation of vision science. PMID- 15268836 TI - [Dynamic changes of tissue inhibitors of metalloproteinases in the rabbit iris and aqueous humor after lens mechanical injury]. AB - OBJECTIVE: To investigate dynamic changes of expression of tissue inhibitors of metalloproteinases (TIMPs) in the rabbit iris and aqueous humor after the lens mechanical injury. METHODS: Rabbit lens was mechanically injured by a needle through the limbus. Eyes were enucleated on the 1st, 3rd, 7th and 15th day after injury, the aqueous humor was collected and the iris was isolated. The activity of two main TIMPs (TIMP-1 and TIMP-2) in the aqueous humor and the iris was assessed by reverse zymography assay while the activity of matrix metalloproteinase-2 (MMP-2) was analyzed by zymography assay. RESULTS: In the healthy and uninjured rabbit eye, neither TIMP-1 nor TIMP-2 was detected in the aqueous humor or in the iris. In the iris of the injured eye, the activity of TIMP-1 and TIMP-2 was increased significantly 1 day after the injury (P < 0.05), the activity of MMP-2 was inhibited. In the aqueous humor of the injured eye, the activity of TIMP-1 and -2 increased significantly (P < 0.05) 1 day after the injury and the activity of MMP-2 was not detectable. The activity of TIMP-1 and 2 decreased afterwards and no significant difference was detected between the injured eyes and the control eyes 7 days after the injury (P value was 0.097 and 0.777, respectively), while the activity of MMP-2 was increased gradually. CONCLUSION: TIMPs are involved in the acute inflammation process after the lens injury, which may play an important role in the inhibition of inflammation and in the wound healing. PMID- 15268837 TI - [Experimental choroidal neovascularization is inhibited by subretinal administration of Endostatin]. AB - OBJECTIVE: Endostatin is an endogeneous angiogenesis inhibitor. The purpose of this study was to investigated the effect of Endostatin on the eyes of rats with experimental choroidal neovascularization (CNV). METHODS: Experimental CNV was induced by laser photocoagulation. Animals were given subretinal injections of recombinant human Endostatin 20 microl (5 g/L) or 0.9% chlorine sodium. The intensity of fluorescein leakage from the photocoagulated lesions was studied 13 days after photocoagulation. The area of CNV at each rupture site was measured using high molecular weight FITC-dextran (MW 2 x 10(6)) for high resolution angiography in RPE-choroid-sclera flat mounts. In addition, 8 eyes in each group were removed and fixed 14 days after photocoagulation, cut into thin sections. The sections were examined by light microscopy. Immunolocalization of Endoglin (CD105) and factor VIII on sections of CNV lesions was studied by immunohistochemical evaluation. RESULTS: After Endostatin injection, fluorescein leakage from the CNV lesions decreased significantly compared with the control eyes. The average area of CNV at sites of the Bruch's membrane rupture showed significant difference in eyes injected with Endostatin compared with control eyes. Endothelial cells demonstrated strong immunoreactivity of CD105 and factor VIII in CNV lesions of control eyes. CD105-positive cell were not detected in normal chorioretinal tissues. CONCLUSIONS: The development of CNV can be inhibited by injection of Endostatin, which suggest that Endostatin may be beneficial in treating CNV and that further studies can be considered to evaluate this possibility. PMID- 15268838 TI - [Experimental study of Retinoblastoma inhibition using all trans Retinoic Acid ]. PMID- 15268839 TI - [Successful treatment in one case of vision loss due to postoperative complications ]. PMID- 15268840 TI - [Pseudoxanthoma elasticum one case]. PMID- 15268841 TI - [intraocular Lincomycin causes retinal toxic retinochoroiditis]. PMID- 15268842 TI - [How to develop evidence-based ophthalmology]. PMID- 15268843 TI - [Present status of the method of microwave thermotherapy for the tumor in eye]. PMID- 15268844 TI - [Summary for fourth national young ophthalmologist conference in China]. PMID- 15268845 TI - [A military nurse on a mission]. AB - In this article some feelings and experiences of a military nurse are described. In the actual war scenario many people are suffering and nurses offer their professional care in cultures and environments very often different from their own. PMID- 15268846 TI - [Saint Antony's healing rituals]. AB - Sant Antonio Abate rouses a particular charm for Italians in general but we focus the attention on Abruzzo people there are many stories and legends about his life and his miracles. This work is a result of a study conducted in the popular culture of Abruzzo Region about the rituals of Sant Antonio. Whom principal function was to protect animals. In the middle of the cult we found his capacities to recover people and animals. The association between the Saint and the fire is connected of his thaumaturgy capacities. The fire is often associated to doctors and to cure sickness, and with him we have therapeutics effects. The hagiographical criticism says that the imagine of Saint has to be referred to a historical fact of IX century in France. Here there was a laic hospital community to recover sick people of cancer egotism that presented hurters blistered skin, fever, eruptions and hallucinations. The pig grass was used to cure egotism and due to this reason sickness was names the evil of Saint Antony or Saint Antonio fire. Due to the multitude of sick people occurring, a hospital run by Augustinians canonists order has been built. PMID- 15268847 TI - [The role of voluntary associations in healthcare politics]. AB - In the recent years an increasing interest has been observed on volunteering; among social scientist, politicians, and economists. A different approach to the management of volunteering has been observed both in the political and social set. Furthermore a remarkable development of the phenomenon is present worldwide as within Italy. The contribute to social development, universally attributed to the volunteer action has been acknowledged by the General Assembly of United Nations that declared the 2001 The International Years of Volunteering. Different aims was attributed to this event: the promotion of volunteer action, as expression of cohesiveness and democratic participation of civic society progress; stimulate the improvement of the volunteering status and role; increasing facilities to the activities realization; facilitate the communication opportunities among volunteers, associations and governments. Nevertheless, the phenomenon relevance has to be viewed in social, cultural, political and economical changes taking place in the last years and still in action in the country that on the one hand have produced new needs and an increasing socio assistance demand and the other hand the needs of changing the responding traditional style and way from Institutions. The major aims of this paper consist in the analysis of motivation and relevance of socio-sanitary role of volunteer in contemporary society. PMID- 15268848 TI - [Anxiety and depression of cancer patients hospitalized and at home]. AB - The aim of this study was to investigate the anxiety and depression of cancer patients hospitalized and at home. Using a descriptive, correlational and comparative design and the Roy Adaptation Model, a sample of 80 oncologic patients was studied. Several instruments were used to measure anxiety and depression (HADS), quality of life and symptoms (RSCL), sociodemographic factors, variables connected to the hospitalization, quality of the relationship with health practitioners, family members and friends and the degree of satisfaction for the received information and support. The examined variables were measured on the same patients at hospital and at home. About the 30% of the patients were anxious and depressed. Statistical analysis showed that while anxiety did not change from the hospital to home, depression increased soon after the discharge and decreased over time and after the increasing of the number of hospital access. Anxiety and depression were positively correlated to boredom during the hospitalization, physical symptoms, number of the patients children, and previous anxious and depressive problems. Anxiety and depression were negatively correlated to the ward comfort, the support of health practitioners, family members and friends and the satisfaction for the received information. Differences between this study and the international literature are discussed. Recommendations for the future research and nursing practice are given. PMID- 15268849 TI - [The perception of anxiety and stress in day surgery: a comparison among patients, family members and nurses]. AB - The purpose of this study was to assess anxiety and stress in day surgery patients, and to compare anxiety and stress as perceived by patients, family members and nurses. A descriptive design involving a convenience sample of 40 patients was used. Patients were waiting for ambulatory procedures of varicectomy, inguinal hernia and breast biopsy. Researchers conducted 120 interviews (40 on patients; 40 on family members and 40 on nurses) with a Questionnaire based on 25 stress factors. The mean intensity of patients' anxiety, measured with a Numerical Rating Scale, was 5,15 (median 5, SD 2,63). Family members and nurses overestimated patients anxiety. When nurses were caring for the same patient more times measured anxiety and stress more accurately. The following stressing factors emerged from this study: a) Fear due to the consequences of the surgery; b) losing one self autonomy in ADL; c) pain; d) anaesthesia. The majority of these factors could be reduce through adequate education and information strategies. PMID- 15268850 TI - [Nursing and clinical tutoring. An experience at University of Chieti]. AB - The national and international experiences of tutoring certainly show today more conformity with the necessities of a formation centred on the student,based on the needs of a society in continuous change, able to form professional people, who can assure and keep adequate levels of competences. Adults' learning active models especially, support the tutoring of teaching and furnish tutors with methods ad instruments which permit to effect efficaciously and efficiently their specific function. The clinical tutoring course for nurses in Chieti University consented to clinical tutors to acquire and/or reinforce psychological, pedagogical, clinical, ethical and managerial competences. PMID- 15268851 TI - Requirement for Abl kinases in T cell receptor signaling. AB - BACKGROUND: The c-Abl and Arg proteins comprise a unique family of nonreceptor tyrosine kinases that have been implicated in the regulation of cell proliferation and survival, cytoskeletal reorganization, cell migration, and the response to oxidative stress and DNA damage. Targeted deletion or mutation of c Abl in mice results in a variety of immune system phenotypes, including splenic and thymic atrophy, lymphopenia, and an increased susceptibility to infection. However, despite the generation of these mice over a decade ago, little is known regarding the mechanisms responsible for these phenotypes or the immune-related consequences of ablation of both the c-Abl and Arg kinases, which are coexpressed in lymphoid tissues. RESULTS: Here, we report that T cell receptor (TCR) stimulation results in activation of the endogenous Abl kinases. We demonstrate that Zap70 and the transmembrane adaptor linker for activation of T cells (LAT) are targets of the Abl kinases, and that loss of Abl kinase activity reduces TCR induced Zap70 phosphorylation at tyrosine 319. This correlates with diminished LAT tyrosine phosphorylation, as well as reduced tyrosine phosphorylation and recruitment of phospholipase Cgamma1 to LAT. Significantly, we show that Abl kinase activity is required for maximal signaling leading to transcription of the IL-2 promoter, as well as TCR-induced IL-2 production and proliferation of primary T cells. CONCLUSIONS: We conclude that the Abl kinases have a role in the regulation of TCR-mediated signal transduction leading to IL-2 production and cell proliferation. PMID- 15268852 TI - MAX3/CCD7 is a carotenoid cleavage dioxygenase required for the synthesis of a novel plant signaling molecule. AB - BACKGROUND: Plant development is exquisitely environmentally sensitive, with plant hormones acting as long-range signals that integrate developmental, genetic, and environmental inputs to regulate development. A good example of this is in the control of shoot branching, where wide variation in plant form can be generated in a single genotype in response to environmental and developmental cues. RESULTS: Here we present evidence for a novel plant signaling molecule involved in the regulation of shoot branching. We show that the MAX3 gene of Arabidopsis is required for the production of a graft-transmissible, highly active branch inhibitor that is distinct from any of the previously characterized branch-inhibiting hormones. Consistent with its proposed function in the synthesis of a novel signaling molecule, we show that MAX3 encodes a plastidic dioxygenase that can cleave multiple carotenoids. CONCLUSIONS: We conclude that MAX3 is required for the synthesis of a novel carotenoid-derived long-range signal that regulates shoot branching. PMID- 15268853 TI - Pigeon homing along highways and exits. AB - BACKGROUND: Anecdotal observations and early airplane and helicopter tracking studies suggest that pigeons sometimes follow large roads and use landmarks as turning points during their homeward journey. However, technical limitations in tracking pigeon routes have prevented proof. RESULTS: Here, we present experimental and statistical evidence for this strategy from the analysis of 216 GPS-recorded pigeon tracks over distances up to 50 km. Experienced pigeons released from familiar sites during 3 years around Rome, Italy, were significantly attracted to highways and a railway track running toward home, in many cases without anything forcing them to follow such guide-rails. Birds often broke off from the highways when these veered away from home, but many continued their flight along the highway until a major junction, even when the detour added substantially to their journey. The degree of road following increased with repeated releases but not flight length. Significant road following (in 40%-50% of the tracks) was mainly observed from release sites along northwest-southeast axis. CONCLUSIONS: Our data demonstrate the existence of a learned road-following homing strategy of pigeons and the use of particular topographical points for final navigation to the loft. Apparently, the better-directed early stages of the flight compensated the added final detour. During early and middle stages of the flight, following large and distinct roads is likely to reflect stabilization of a compass course rather than the presence of a mental roadmap. A cognitive (roadmap) component manifested by repeated crossing of preferred topographical points, including highway exits, is more likely when pigeons approach the loft area. However, it might only be expected in pigeons raised in an area characterized by navigationally relevant highway systems. PMID- 15268854 TI - A double segment periodicity underlies segment generation in centipede development. AB - The number of leg-bearing segments in centipedes varies extensively, between 15 and 191, and yet it is always odd. This suggests that segment generation in centipedes involves a stage with double segment periodicity and that evolutionary variation in segment number reflects the generation of these double segmental units. However, previous studies have revealed no trace of this. Here we report the expression of two genes, an odd-skipped related gene (odr1) and a caudal homolog, that serve as markers for early steps of segment formation in the geophilomorph centipede, Strigamia maritima. Dynamic expression of odr1 around the proctodaeum resolves into a series of concentric rings, revealing a pattern of double segment periodicity in overtly unsegmented tissue. Initially, the expression of the caudal homolog mirrors this double segment periodicity, but shortly before engrailed expression and overt segmentation, the intercalation of additional stripes generates a repeat with single segment periodicity. Our results provide the first clues about the causality of the unique and fascinating "all-odd" pattern of variation in centipede segment numbers and have implications for the evolution of the mechanisms of arthropod segmentation. PMID- 15268855 TI - The ERK MAP kinase cascade mediates tail swelling and a protective response to rectal infection in C. elegans. AB - The nematode Caenorhabditis elegans is proving to be an attractive model organism for investigating innate immune responses to infection. Among the known pathogens of C. elegans is the bacterium Microbacterium nematophilum, which adheres to the nematode rectum and postanal cuticle, inducing swelling of the underlying hypodermal tissue and causing mild constipation. We find that on infection by M. nematophilum, an extracellular signal-regulated kinase (ERK) mitogen-activated protein (MAP) kinase cascade mediates tail swelling and protects C. elegans from severe constipation, which would otherwise arrest development and cause sterility. Involvement in pathogen defense represents a new role for ERK MAP kinase signaling in this organism. PMID- 15268856 TI - Compensatory proliferation induced by cell death in the Drosophila wing disc requires activity of the apical cell death caspase Dronc in a nonapoptotic role. AB - Achieving proper organ size requires a balance between proliferation and cell death. For example, at least 40%-60% of cells in the Drosophila wing disc can be lost, yet these discs go on to give rise to normal-looking adult wings as a result of compensatory proliferation. The signals that drive this proliferation are unknown. One intriguing possibility is that they derive, at least in part, from the dying cells. To explore this hypothesis, we activated cell death signaling in specific populations of cells in the developing wing but prevented these cells from dying through expression of the baculovirus p35 protein, which inhibits the activity of effector caspases that mediate apoptosis. This allowed us to uncouple the activation steps of apoptosis from death itself. Here we report that stimulation of cell death signaling in the wing disc-in the absence of cell death-results in increased proliferation and ectopic expression of Wingless, a known mitogen in the wing. Activation of the apical cell death caspase Dronc is necessary and sufficient to drive both of these processes. Our results demonstrate an unanticipated function, the nonautonomous induction of proliferation, of an apical cell death caspase. This activity is likely to contribute to tissue homeostasis by promoting local compensatory proliferation in response to cell death. We speculate that dying cells may communicate cell fate or behavior instructions to their neighbors in other contexts as well. PMID- 15268857 TI - Experience in early infancy is indispensable for color perception. AB - Early visual experience is indispensable to shape the maturation of cortical circuits during development. Monocular deprivation in infancy, for instance, leads to an irreversible reduction of visually driven activity in the visual cortex through the deprived eye and a loss of binocular depth perception. It was tested whether or not early experience is also necessary for color perception. Infant monkeys were reared for nearly a year in a separate room where the illumination came from only monochromatic lights. After extensive training, they were able to perform color matching. But, their judgment of color similarity was quite different from that of normal animals. Furthermore, they had severe deficits in color constancy; their color vision was very much wavelength dominated, so they could not compensate for the changes in wavelength composition. These results indicate that early visual experience is also indispensable for normal color perception. PMID- 15268858 TI - Nerve growth factor signaling regulates motility and docking of axonal mitochondria. AB - Axonal transport is thought to distribute mitochondria to regions of the neuron where their functions are required. In cultured neurons, mitochondrial transport responds to growth cone activity, and this involves both a transition between motile and stationary states of mitochondria and modulation of their anterograde transport activity. Although the exact cellular signals responsible for this regulation remain unknown, we recently showed that mitochondria accumulate in sensory neurons at regions of focal stimulation with NGF and suggested that this involves downstream kinase signaling. Here, we demonstrate that NGF regulation of axonal organelle transport is specific to mitochondria. Quantitative analyses of motility show that the accumulation of axonal mitochondria near a focus of NGF stimulation is due to increased movement into bead regions followed by inhibition of movement out of these regions and that anterograde and retrograde movement are differentially affected. In axons made devoid of F-actin by latrunculin B treatment, bidirectional transport of mitochondria continues, but they can no longer accumulate in the region of NGF stimulation. These results indicate that intracellular signaling can specifically regulate mitochondrial transport in neurons, and they suggest that axonal mitochondria can respond to signals by locally altering their transport behavior and by undergoing docking interactions with the actin cytoskeleton. PMID- 15268859 TI - Control of centromere localization of the MEI-S332 cohesion protection protein. AB - In mitosis and meiosis, cohesion is maintained at the centromere until sister chromatid separation. Drosophila MEI-S332 is essential for centromeric cohesion in meiosis and contributes to, though is not absolutely required for, cohesion in mitosis. It localizes specifically to centromeres in prometaphase and delocalizes at the metaphase-anaphase transition. In mei-S332 mutants, centromeric sister chromatid cohesion is lost at anaphase I, giving meiosis II missegregation. MEI S332 is the founding member of a family of proteins important for chromosome segregation. One likely activity of these proteins is to protect the cohesin subunit Rec8 from cleavage at the metaphase I-anaphase I transition. Although the family members do not show high sequence identity, there are two short stretches of homology, and mutations in conserved residues affect protein function. Here we analyze the cis- and trans-acting factors required for MEI-S332 localization. We find a striking correlation between domains necessary for MEI-S332 centromere localization and conserved regions within the protein family. Drosophila MEI-S332 expressed in human cells localizes to mitotic centromeres, further highlighting this functional conservation. MEI-S332 can localize independently of cohesin, assembling even onto unreplicated chromatids. However, the separase pathway that regulates cohesin dissociation is needed for MEI-S332 delocalization at anaphase. PMID- 15268860 TI - Clustered organization of reproductive genes in the C. elegans genome. AB - Defining the forces that sculpt genome organization is fundamental for understanding the origin, persistence, and diversification of species. The genomic sequences of the nematodes Caenorhabditis elegans and Caenorhabditis briggsae provide an excellent opportunity to explore the dynamics of chromosome evolution. Extensive chromosomal rearrangement has accompanied divergence from their common ancestor, an event occurring roughly 100 million years ago (Mya); yet, morphologically, these species are nearly indistinguishable and both reproduce primarily by self-fertilization. Here, we show that genes expressed during spermatogenesis (sperm genes) are nonrandomly distributed across the C. elegans genome into three large clusters located on two autosomes. In addition to sperm genes, these chromosomal regions are enriched for genes involved in the hermaphrodite sperm/oocyte switch and in the reception of sperm signals that control fertilization. Most loci are present in single copy, suggesting that cluster formation is largely due to gene aggregation and not to tandem duplication. Comparative mapping indicates that the C. briggsae genome differs dramatically from the C. elegans genome in clustering. Because clustered genes have a direct role in reproduction and thus fitness, their aggregated pattern might have been shaped by natural selection, perhaps as hermaphroditism evolved. PMID- 15268861 TI - The C. elegans thermosensory neuron AFD responds to warming. AB - The mechanism of temperature sensation is far less understood than the sensory response to other environmental stimuli such as light, odor, and taste. Thermotaxis behavior in C. elegans requires the ability to discriminate temperature differences as small as approximately 0.05 degrees C and to memorize the previously cultivated temperature. The AFD neuron is the only major thermosensory neuron required for the thermotaxis behavior. Genetic analyses have revealed several signal transduction molecules that are required for the sensation and/or memory of temperature information in the AFD neuron, but its physiological properties, such as its ability to sense absolute temperature or temperature change, have been unclear. We show here that the AFD neuron responds to warming. Calcium concentration in the cell body of AFD neuron is increased transiently in response to warming, but not to absolute temperature or to cooling. The transient response requires the activity of the TAX-4 cGMP-gated cation channel, which plays an essential role in the function of the AFD neuron. Interestingly, the AFD neuron further responds to step-like warming above a threshold that is set by temperature memory. We suggest that C. elegans provides an ideal model to genetically and physiologically reveal the molecular mechanism for sensation and memory of temperature information. PMID- 15268862 TI - Rictor, a novel binding partner of mTOR, defines a rapamycin-insensitive and raptor-independent pathway that regulates the cytoskeleton. AB - The mammalian TOR (mTOR) pathway integrates nutrient- and growth factor-derived signals to regulate growth, the process whereby cells accumulate mass and increase in size. mTOR is a large protein kinase and the target of rapamycin, an immunosuppressant that also blocks vessel restenosis and has potential anticancer applications. mTOR interacts with the raptor and GbetaL proteins to form a complex that is the target of rapamycin. Here, we demonstrate that mTOR is also part of a distinct complex defined by the novel protein rictor (rapamycin insensitive companion of mTOR). Rictor shares homology with the previously described pianissimo from D. discoidieum, STE20p from S. pombe, and AVO3p from S. cerevisiae. Interestingly, AVO3p is part of a rapamycin-insensitive TOR complex that does not contain the yeast homolog of raptor and signals to the actin cytoskeleton through PKC1. Consistent with this finding, the rictor-containing mTOR complex contains GbetaL but not raptor and it neither regulates the mTOR effector S6K1 nor is it bound by FKBP12-rapamycin. We find that the rictor-mTOR complex modulates the phosphorylation of Protein Kinase C alpha (PKCalpha) and the actin cytoskeleton, suggesting that this aspect of TOR signaling is conserved between yeast and mammals. PMID- 15268864 TI - Europe seeks to double research budget. PMID- 15268863 TI - Contextual taste cues modulate olfactory learning in C. elegans by an occasion setting mechanism. AB - Manipulations of context can affect learning and memory performance across species in many associative learning paradigms. Using taste cues to create distinct contexts for olfactory adaptation assays in the nematode Caenorhabditis elegans, we now show that performance in this associative learning paradigm is sensitive to context manipulations, and we investigate the mechanism(s) used for the integration of context cues in learning. One possibility is that the taste and olfactory stimuli are perceived as a combined, blended cue that the animals then associate with the unconditioned stimulus (US) in the same manner as with any other unitary conditioned stimuli (CS). Alternatively, an occasion-setting model suggests that the taste cues only define the appropriate context for olfactory memory retrieval without directly entering into the primary association. Analysis of genetic mutants demonstrated that the olfactory and context cues are sensed by distinct primary sensory neurons and that the animals' ability to use taste cues to modulate olfactory learning is independent from their ability to utilize these same taste cues for adaptation. We interpret these results as evidence for the occasion-setting mechanism in which context cues modulate primary Pavlovian association by functioning in a hierarchical manner to define the appropriate setting for memory recall. PMID- 15268865 TI - One long argument. PMID- 15268866 TI - Karl Sigmund. PMID- 15268867 TI - Shady deals. PMID- 15268869 TI - Schistosoma. PMID- 15268870 TI - Histones and histone modifications. PMID- 15268871 TI - Insecticide resistance: a silent base prediction. PMID- 15268872 TI - Chromosome condensation: DNA compaction in real time. AB - Mitotic chromosomes must be organised into a highly ordered and compacted form to allow proper segregation of DNA during each round of cell division. Two new studies report observations of DNA compaction by eukaryotic and bacterial condensin molecules in real time using magnetic and optical trapping micromanipulation techniques. PMID- 15268873 TI - Arthropod segmentation: why centipedes are odd. AB - Recent work has revealed a double segmental periodicity of gene expression in the centipede, a potential molecular explanation for the observation that this arthropod always has an odd number of trunk segments. Is this an oddity of centipedes, or might it mean that double segmental pair-rule patterning dates back to the Ur-arthropod? PMID- 15268874 TI - Taste perception: Drosophila - a model of good taste. AB - Recent studies of taste receptors in Drosophila show remarkable parallels with the mammalian gustatory system, although the pathways are anatomically distinct. These parallels may reflect crucial constraints in the design of taste detection systems. PMID- 15268875 TI - TCR signaling: another Abl-bodied kinase joins the cascade. AB - Protein tyrosine kinases have long been recognized as the most proximal actors in T-cell antigen receptor (TCR) signaling. Three non-receptor tyrosine kinase families (Src, ZAP-70 and Tec) are known to be critical, but a new study now shows that room needs to be made in this pathway for yet another protein tyrosine kinase family - Abl/Arg. PMID- 15268876 TI - Telomerase RNA: a flexible RNA scaffold for telomerase biosynthesis. AB - Determination of the structure of the yeast telomerase RNA component TLC1 has been hampered by its large size and high rate of evolutionary divergence. But detailed phylogenetic comparisons have now revealed the unusually flexible and modular architecture of this important RNA molecule. PMID- 15268877 TI - Sex determination: balancing selection in the honey bee. AB - Sequences of alleles of the honey bee's primary sex-determining gene have extremely high diversity, with many amino acid variants, suggesting that different alleles of this gene have been maintained in populations for very long evolutionary times. PMID- 15268878 TI - Developmental biology: Notching the hindbrain. AB - One of the best-characterized lineage restrictions in developing vertebrates occurs between adjacent -rhombomeres of the hindbrain. It was recently shown that cells at the boundaries of zebrafish rhombomeres also differ from non-boundary cells in their migratory abilities, a difference driven by Notch signaling. PMID- 15268879 TI - Working memory: imaging the magic number four. AB - Many brain regions have been implicated in memory performance, but the relationship between memory capacity and neural activity has not been clear. Recent studies show that activity in the posterior parietal cortex increases with working memory load, implicating this region in the storage of representations in visual memory. PMID- 15268880 TI - Comparative genomics: prediction of the ciliary and basal body proteome. AB - Defects in mammalian cilia lead to a range of diseases, but our understanding of the composition of these organelles and of the basal bodies from which they arise is limited. Two recent studies used comparative genomics to predict the ciliary and basal body proteomes, providing datasets that are rich sources of human disease gene candidates. PMID- 15268881 TI - Germ cell specification and migration in Drosophila and beyond. AB - The passage of an individual's genome to future generations is essential for the maintenance of species and is mediated by highly specialized cells, the germ cells. Genetic studies in a number of model organisms have provided insight into the molecular mechanisms that control specification, migration and survival of early germ cells. Focusing on Drosophila, we will discuss the mechanisms by which germ cells initially form and remain transcriptionally silent while somatic cells are transcriptionally active. We will further discuss three separate attractive and repellent guidance pathways, mediated by a G-protein coupled receptor, two lipid phosphate phosphohydrolases, and isoprenylation. We will compare and contrast these findings with those obtained in other organisms, in particular zebrafish and mice. While aspects of germ cell specification are strikingly different between these species, germ cell specific gene functions have been conserved. In particular, mechanisms that sense directional cues during germ cell migration seem to be shared between invertebrates and vertebrates. PMID- 15268882 TI - Misconceptions IV--the hypophosphatemia of primary hyperparathyroidism is the result of renal phosphate wasting. PMID- 15268883 TI - The varying distribution of intra- and inter-vertebral height ratios determines the prevalence of vertebral fractures. AB - Credible inferences regarding the burden of vertebral fractures (VFs) cannot be made without a globally accepted quantitative definition of 'fracture'. Currently, differences in anterior, middle, or posterior vertebral heights (VHs) within a vertebra, or between adjacent vertebrae, are used to define 'fracture'. However, VH differences are essential for the construction of thoracolumbar curves, evolutionary adaptations that provide stability in bipedal stance and gait. As there is no reference standard to distinguish anatomical variation from fracture, approaches to defining a VF use a reference range of VH ratios derived in premenopausal women or derived by trimming, a method that iteratively removes the tails of a distribution of VH ratios to produce a normal distribution. From this, reference ranges of VH ratio means and standard deviations (SDs) are obtained and a nominal deviation of 15% or more, or 3 SD or more is regarded as a 'fracture'. We measured VHs by quantitative vertebral morphometry (QVM) and bone mineral density (BMD) by dual energy X-ray absorptiometry in 697 Lebanese women (age 20-89 years) to compare the prevalence of VF ascertained by published methods and a new method that uses the premenopausal range (without trimming) and requires two VH abnormalities. VF prevalence using published methods reached 60% to 70% in pre- and post-menopausal women, and in women with normal or high BMD because VH ratios were not normally distributed and cut-offs used to define VF fracture fell within the observed distribution of the data. The new method resulted in a VF prevalence of 3.3% in younger and 14% in older women, 7% (high), 10% (middle), and 20% (low) BMD tertiles consistent with the notion that the method detected VF due to bone fragility. We suggest that using a fixed trimming method to define reference range and cut-offs or applying fixed cut-offs to identify VFs in populations, where these ratios are not normally distributed, may result in the capture of anatomical variation, not structural failure. Thus, group differences in the VF prevalence may reflect differences in methodology, not bone fragility. Improved criteria to define VF are needed before credible inferences can be made regarding the burden of VFs in women and men, and between sexes, races, countries, decades, and placebo arms of clinical trials. PMID- 15268884 TI - Does follow-up duration influence the ultrasound and DXA prediction of hip fracture? The EPIDOS prospective study. AB - While the potential of quantitative ultrasound (QUS) in the management of osteoporosis has been accepted, its interaction with follow-up time has never been investigated. The aim of our study is to prospectively evaluate the influence of follow-up time on the prediction of hip fracture by ultrasound parameters in the elderly as compared to bone mineral density (BMD) and to establish a long-term fracture prediction model. In the multicenter prospective study EPIDOS, 5898 Caucasian healthy women, aged 75 and over, had femoral dual energy X-ray absorptiometry (DXA) and heel ultrasound measurements at baseline. A survey of fracture occurrence was conducted every 4 months. Statistical analyses were performed for three different average lengths of follow-up, namely, 1.5, 2.5 and 3.5 years. Relative risks per standard deviation decrease (RR) and the area under the receiver operating characteristic (AUC) curves were given. Estimates of the long-term hip fracture prediction by DXA and QUS were extrapolated. During an average of 3.5 years follow-up, 227 women sustained their first non-traumatic hip fracture. For the three categories of follow-up, low values of both calcaneal ultrasound and hip BMD were associated with a significant increased risk of hip fracture [e.g. ultrasound Stiffness index RR = 2.8 (2.1-3.8), 2.1 (1.7-2.6) and 1.9 (1.7-2.3) for 1.5, 2.5 and 3.5 years of follow-up, respectively]. The combination of femoral neck BMD with the Stiffness showed an improvement of the hip fracture prediction model. Using extrapolation, the prediction of hip fracture by the Stiffness remained significant up to 7.5 years [RR = 1.2 (1.03 1.41)], whereas the limit of significance was reached at 10 years for the femoral neck BMD [RR = 1.25 (1.04-1.52)]. Our results indicate that the Stiffness tends to be the best short- and long-term predictor of hip fracture among ultrasound parameters. This paper provides additional information on the long-term prediction of hip fracture, which has always been an important issue in routine clinical practice as it influences the management of the disease. Our model should give a relatively good estimation of the fracture risk prediction at 5 years with the ultrasound and 10 years for the femoral neck BMD. PMID- 15268885 TI - Determinants and heterogeneity of mechanical competence throughout the thoracolumbar spine of elderly women and men. AB - Vertebral fractures represent the hallmark of osteoporosis. Here, we test the hypotheses that (sub)cortical bone strength and density predict failure better than trabecular core strength and density, and that elderly women display lower failure stress of thoracic vertebrae than men. We examined the vertebral bodies T3 to L5 in 39 spines from elderly donors (23 women; 16 men; age 79 +/- 11 years). Peripheral quantitative computed tomography was used to measure total, trabecular, and (sub)cortical bone density. Mechanical tests were performed in functional spinal units, planoparallel sections of vertebrae, trabecular cores, and (sub)cortical ring specimens. The failure stress decreased with descending vertebral level. Failure stress was highest for the (sub)cortical rings and planoparallel sections and lowest for the trabecular core. The failure stress did not differ significantly between men and women. Mechanical strength of the functional unit was more strongly correlated with the strength of the (sub)cortical ring (r = 0.78) than with that of the trabecular core (r = 0.62). However, total density was more highly correlated with mechanical strength of the same and remote vertebrae (r = 0.63) than trabecular (r = 0.50) or (sub)cortical density (r = 0.36), respectively. The results show that vertebral strength is similar in elderly women and men. Strength of (sub)cortical bone provides significantly better prediction of strength of functional spinal units than that of the trabecular core. However, total density predicts functional segment failure stress with higher accuracy than (sub)cortical or trabecular density and is thus recommended for predicting fracture strength clinically. PMID- 15268886 TI - A meta-analysis of previous fracture and subsequent fracture risk. AB - Previous fracture is a well-documented risk factor for future fracture. The aim of this study was to quantify this risk on an international basis and to explore the relationship of this risk with age, sex, and bone mineral density (BMD). We studied 15259 men and 44902 women from 11 cohorts comprising EVOS/EPOS, OFELY, CaMos, Rochester, Sheffield, Rotterdam, Kuopio, DOES, Hiroshima, and two cohorts from Gothenburg. Cohorts were followed for a total of 250000 person-years. The effect of a prior history of fracture on the risk of any fracture, any osteoporotic fracture, and hip fracture alone was examined using a Poisson model for each sex from each cohort. Covariates examined were age, sex, and BMD. The results of the different studies were merged by using the weighted beta coefficients. A previous fracture history was associated with a significantly increased risk of any fracture compared with individuals without a prior fracture (RR = 1.86; 95% CI = 1.75-1.98). The risk ratio was similar for the outcome of osteoporotic fracture or for hip fracture. There was no significant difference in risk ratio between men and women. Risk ratio (RR) was marginally downward adjusted when account was taken of BMD. Low BMD explained a minority of the risk for any fracture (8%) and for hip fracture (22%). The risk ratio was stable with age except in the case of hip fracture outcome where the risk ratio decreased significantly with age. We conclude that previous history of fracture confers an increased risk of fracture of substantial importance beyond that explained by measurement of BMD. Its validation on an international basis permits the use of this risk factor in case finding strategies. PMID- 15268887 TI - Health care costs of women with symptomatic vertebral fractures. AB - BACKGROUND: An important aspect of the economics of fracture prevention is averted fracture costs. However, while vertebral fractures represent a significant burden to society, quantifying their cost is difficult for several reasons. In this paper, we examine the health care costs of symptomatic vertebral fractures occurring in women aged 50 years and above in the UK. METHODS: We used a variety of data sources. The prevalence of pharmaceutical treatment for fracture prevention and number of general practitioner consultations, referrals, and hospital admissions associated with a diagnosis of vertebral fracture were identified from a case control study. For the unit cost of a general practitioner consultation, referral, and cost per inpatient day, we used 2002 data produced by the Personal Social Services Research Unit. Hospital Episode Statistics (HES) for 2001-2002 were used to estimate the median length of stay in hospital for women aged 50 years and above, and the Monthly Index of Medical Specialities (MIMS) was used to identify the costs of pharmaceutical treatments. Costs were discounted at 6%. RESULTS: From these data, we estimated that for the year prior and post diagnosis the average additional health care costs for those diagnosed with vertebral fracture were pounds 165, pounds 134, and pounds 2314 for general practitioner consultations, referrals, and hospital admissions, respectively (i.e., pounds 2613). The cost of pharmaceutical treatments prescribed for fracture prevention in the year following diagnosis was pound 97. DISCUSSION: Vertebral fractures are associated with significantly increased health care costs. These costs need to be set against the costs of fracture prevention. PMID- 15268888 TI - Assessment of the skeletal health of healthy Nigerian men and women using quantitative ultrasound. AB - The dietary intake of calcium by African populations, particularly in sub-Saharan Africa, is relatively low compared to the recommended intake for US adults. However, the rate of osteoporotic fractures in West African women is reported to be less than that for Caucasian populations. Because there is little published data regarding the skeletal status of African men and women, we used quantitative ultrasound (QUS) to assess the bone density of 435 Nigerian women and 321 Nigerian men between 16 and 89 years of age. A progressive decline in bone quality was observed beginning at about 40 years of age for both men and women. The mean stiffness index (SI) for the women between 20 and 35 years of age (n = 186) in this study was 102 +/- 17. The equation that best described the age versus SI relationship for women was SI = 79.7 + 1.887 (age) + -0.043 (age)2 + 0.00020 (age)3. For Nigerian men, the peak SI of 115 +/- 17 was seen in the 20- to 29-year-old age group. For men, the SI values remained above 100 until about age 60 years when a significant decline in SI was then observed. The best-fit curve of SI versus age for men was SI = 134.9 - 1.27 (age) + 0.019 (age)2 - 0.00014 (age)3. The broadband ultrasound attenuation (BUA), speed of sound (SOS), and SI values for the Nigerian men and women were comparable to or higher than those reported for Caucasian and Asian populations. These data should serve as reference values for adult men and women in sub-Saharan Africa. PMID- 15268889 TI - Test of linkage and/or association between the estrogen receptor alpha gene with bone mineral density in Caucasian nuclear families. AB - Extensive studies have been performed on the association between the estrogen receptor alpha (ER-alpha) gene and bone mineral density (BMD). Despite considerable efforts, the studies using limited markers and relatively small sample size have yielded largely inconsistent results. In this study, 1873 Caucasian subjects from 405 nuclear families containing 1512 sib pairs were recruited. BMD at the lumbar spine (LS) and femoral neck (FN) was measured by dual-energy X-ray absorptiometry (DXA). Seven single-nucleotide polymorphisms (SNPs) spanning from exon 1 to 8 in the ER-alpha gene were genotyped. The program QTDT (quantitative transmission disequilibrium test) was applied to test linkage and/or association of the ER-alpha gene and BMD variation using individual SNP markers and reconstructed haplotypes. Linkage disequilibrium (LD) was generally detected for SNPs in the ER-a gene (P < 0.05). Associations were observed between SNP rs932477 and FN BMD (P = 0.028), and between the most predominant three marker haplotype (GCG) containing SNP rs932477 and FN BMD (P = 0.010). Within family association (present only with both linkage and association) between SNP rs2228480 (G2014A) and FN BMD (P = 0.015) was observed. The most predominant seven-SNP haplotype (TCGCGGG) was associated with higher LS BMD (P = 0.015). However, after correction for multiple testing, these associations did not reach statistical significance. Denser markers may be necessary to better define the relationship between the ER-alpha gene and BMD variation in our sample. PMID- 15268890 TI - Fourier and wavelet analyses of dental radiographs detect trabecular changes in osteoporosis. AB - OBJECTIVES: Osteoporosis results in loss of bone mass and microarchitectural deterioration. Dental radiographs potentially offer a means of screening for osteoporosis as they are commonly made on adults. Spatial frequency analyses are well suited to detect subtle changes in image patterns. We hypothesize that individuals with osteoporosis exhibit an altered radiographic trabecular pattern that can be detected by spatial frequency and strut analysis. STUDY DESIGN: Maxillary and mandibular periapical radiographs of 26 women with osteoporosis and 23 controls were examined using one-dimensional discrete Fourier and wavelet analyses in both jaws to measure the spatial frequency distributions of trabecular structures. A strut analysis was also performed. RESULTS: Individuals with osteoporosis revealed an altered trabecular pattern compared to controls. Using Fourier and strut variables allows classification of subjects with 92% sensitivity, 96% specificity, and a 22% cross-validation error rate. Wavelet analysis was also useful but did not perform better than Fourier analysis for subject classification. CONCLUSIONS: Spatial frequency analysis of digitized dental radiographs, especially Fourier analysis, and strut analysis provide value for identifying individuals with osteoporosis. PMID- 15268891 TI - Vitamin D deficiency and secondary hyperparathyroidism and the association with bone mineral density in persons with Pakistani and Norwegian background living in Oslo, Norway, The Oslo Health Study. AB - We studied the prevalence of poor vitamin D status and the association with bone density in men and women born in Norway (quoted as Norwegians, n = 869) and Pakistan (quoted as Pakistanis, n = 177) in the population-based Oslo Health Study, 2000-2001. We measured 25-hydroxyvitamin D, iPTH and ionized calcium in serum and bone mineral density at the forearm site with single energy X-ray absorptiometry. Mean 25-hydroxyvitamin D was 74.8 +/- 23.7 nmol/l in the Norwegians and 25.0 +/- 13.6 nmol/l in the Pakistanis (P = 0.000). The prevalence of secondary hyperparathyroidism (iPTH > or = 8.5 pmol/l, 25-hydroxyvitamin D < 50 nmol/l and Ca2+ < or = 1.35 mmol/l) was four times higher in Pakistani compared to Norwegian women. Also in Pakistani men, serious vitamin D deficiency defined as secondary hyperparathyroidism was prevalent, and five times as frequent as in Norwegian men. However, whereas BMD was significantly lower in Norwegian women with, compared to Norwegian women without, secondary hyperparathyroidism, there was no difference in BMD between Pakistani women with and without secondary hyperparathyroidism. In conclusion, vitamin D deficiency was prevalent among Pakistani immigrants, and in great contrast to the vitamin D replete Norwegians. Serious vitamin D deficiency was interestingly not associated with reduced forearm bone density among Pakistani women. PMID- 15268892 TI - The effect of prophylactic treatment with risedronate on stress fracture incidence among infantry recruits. AB - When subjected to strains or strain rates higher than usual, the bone remodels to repair microdamage and to strengthen itself. During the initial resorption phase of remodeling, the bone is transitorily weakened and microdamage can accumulate leading to stress fracture. To determine whether short-term suppression of bone turnover using bisphosphonates can prevent the initial loss of bone during the remodeling response to high bone strain and strain rates and potentially prevent stress fracture, we conducted a randomized, double-blind, placebo-controlled trial of 324 new infantry recruits known to be at high risk for stress fracture. Recruits were given a loading dose of 30 mg of risedronate or placebo daily for 10 doses during the first 2 weeks of basic training and then a once a week maintenance dose for the following 12 weeks. Recruits were monitored by biweekly orthopedic examinations during 15 weeks of basic training for stress fractures. Bone scans for suspected tibial and femoral stress fractures and radiographs for suspected metatarsal stress fractures were used to verify stress fracture occurrence. By the intention-to-treat analysis and per-protocol analysis, there was no statistically significant difference in the tibial, femoral, metatarsal, or total stress fracture incidence between the treatment group and the placebo. We conclude that prophylactic treatment with risedronate in a training population at high risk for stress fracture using a maintenance dosage for the treatment of osteoporosis does not lower stress fracture risk. PMID- 15268893 TI - Morphological and mechanical properties of caudal vertebrae in the SAMP6 mouse model of senile osteoporosis. AB - The senescence-accelerated mouse, strain P6 (SAMP6), is a model of senile osteoporosis with relatively low bone mineral density (BMD), low rates of bone formation and reduced long-bone bending strength. Seeking to extend previous descriptions of the SAMP6 skeletal phenotype, we assessed the morphological and mechanical properties of vertebrae from SAM mice at 4 and 12 months of age. We hypothesized that, relative to SAMR1 controls, vertebrae from SAMP6 mice have: (1) less trabecular bone, (2) increased endosteal and periosteal bone size and (3) decreased whole-bone strength. Caudal vertebrae from adult female mice (SAMR1 and SAMP6; 4 and 12 months; n = 10-11 per group) were evaluated by micro-computed tomography and mechanical compression testing. SAMP6 vertebrae had 33% less trabecular bone volume in cephalad and caudal end regions than SAMR1 (P < 0.001) due to significant reductions in both trabecular number (P = 0.002) and thickness (P < 0.001). In contrast to previous findings in SAMP6 long bones, SAMP6 vertebrae showed no evidence of increased overall bone dimensions compared to SAMR1, and in fact had smaller total bone area (TA) and endosteal area (EA) at 12 months of age. Whereas SAMR1 vertebrae showed signs of age-related expansion from 4 to 12 months, SAMP6 vertebral dimensions did not change with age. Mechanical properties of SAMP6 vertebrae were not significantly different than SAMR1 vertebrae (stiffness, yield force, ultimate force, displacement at ultimate force, energy to ultimate force; P > 0.05), another finding in contrast to previous results in SAMP6 long bones. In summary, reduced vertebral trabecular bone volume is another feature of the SAMP6 mice with relevance to senile osteoporosis. However, age-related bone expansion and reduced whole-bone strength were not evident in SAMP6 vertebrae, indicating that while the SAMP6 mouse has many features relevant to senile osteoporosis in humans, not all features are observed or detectable at all skeletal sites. PMID- 15268894 TI - Analysis of three-dimensional microarchitecture and degree of mineralization in bone metastases from prostate cancer using synchrotron microcomputed tomography. AB - Bone architecture and mineralization are generally considered to be important components of bone quality, and determine bone strength in conjunction with bone mineral density. Although the features of bone quality have recently been studied under conditions in which bone density decreases, such as osteoporosis, little is known in osteosclerotic diseases. In this study, we compared the trabecular bone microarchitecture and degree of mineralization between osteoblastic bone metastasis and degenerative osteosclerosis using synchrotron radiation microcomputed tomography (SR-microCT). Small cubes of lumbar vertebrae were excised postmortem from the sites of osteoblastic metastasis, degenerative osteosclerosis, and comparative sites of normal subjects without skeletal lesions. The samples were imaged at high spatial resolution (voxel size = 6 microm) using the SR-microCT system developed at the synchrotron facility (SPring 8), Hyogo, Japan. The three-dimensional (3D) image data were then analyzed for the morphological parameters and the degree of mineralization of bone (DMB). Trabecular bone in metastatic lesions showed a highly connected and isotropic network pattern compared with the normal samples. Although the trabecular surface was markedly irregular in osteoblastic metastases, no significant difference was found in the mean trabecular thickness (Tb.Th) between osteoblastic metastases and normal tissue. The DMB of trabeculae in metastatic lesions had a broader range and lower mean than that of the normal tissue. In contrast, trabecular bone in degenerative osteosclerotic lesions showed a similar degree of anisotropy (DA) and connectivity to the normal tissue, whereas the trabecular thickness was greater in the degenerative osteosclerotic lesions. No significant difference in DBM between degenerative osteosclerosis and normal tissue was detected. These results characterize the difference in bone quality between osteoblastic bone metastasis and degenerative osteosclerosis. Further study on the relationship between bone quality and bone strength in these osteosclerotic lesions would improve our understanding of the pathogenesis of bone fragility. PMID- 15268895 TI - The association of bone metabolism with bone mineral density, serum sex hormone concentrations, and regular exercise in middle-aged men. AB - Physical activity is an important factor in attaining bone mass. Our aim was to investigate if low to moderate intensity exercise affects bone resorption [serum tartrate-resistant acid phosphatase (TRAP) 5b activity] and formation (serum osteocalcin concentration) in a randomized controlled exercise intervention trial in Finnish middle-aged men. In addition, the relations of these bone turnover markers with bone mineral density (BMD) and serum sex hormone concentrations [circulating testosterone (T), estradiol (E2), and sex hormone-binding globulin (SHBG) concentrations] were evaluated. Serum TRAP 5b activity and osteocalcin concentration were measured at randomization and after 1 and 4 years of the exercise intervention. BMDs of the lumbar spine (L2-L4), femoral neck, and total proximal femur were measured with a dual-energy X-ray absorptiometry (DXA). At randomization, TRAP 5b activity was strongly correlated with the osteocalcin concentration (Spearman r = 0.541, P < 0.0001). In addition, TRAP 5b activity was significantly correlated with proximal femur BMD values (r = -0.201, P = 0.018) and osteocalcin concentration with femoral neck and proximal femur BMD values (r = -0.187, P = 0.028; r = -0.240, P = 0.005, respectively). Serum E2, free E2, and free T concentrations were inversely correlated with both bone turnover markers. After 1 year of exercise intervention, TRAP 5b activity was significantly lower in the exercise than reference group (P = 0.006). However, after 4 years of exercise intervention, the difference was no longer statistically significant. There were no differences in the osteocalcin concentrations between the study groups during the intervention. Our results show a connection between serum TRAP 5b activity and osteocalcin concentration. Furthermore, our results suggest that low to moderate exercise intervention and serum sex hormone concentrations may induce changes in bone metabolism in middle-aged men. However, exercise-induced effects on bone metabolism should be confirmed in other randomized controlled exercise trials taking into account exercise intensity and dose-response issues. PMID- 15268896 TI - Unique regulation of SOST, the sclerosteosis gene, by BMPs and steroid hormones in human osteoblasts. AB - SOST, a novel bone morphogenetic protein (BMP) antagonist and negative regulator of bone formation, is expressed in osteogenic cells. Null mutations in the SOST gene are associated with the sclerosteosis phenotype typified by high bone mass. We sought to delineate the pathways involved in the regulation of SOST expression in human osteoblastic cells. We evaluated the effects of bone growth factors and hormones on the RNA levels of SOST and the BMP antagonists, noggin and gremlin. Parathyroid hormone (PTH), transforming growth factor-beta1 (TGF-beta1), fibroblast growth factors 1 and 2 (FGF1, FGF2), and insulin-like growth factor-1 (IGF-1) had negligible effects on SOST expression in human osteoblasts. In comparison, BMPs-2, 4, and 6 induced the message levels of SOST in a time- and dose-dependent manner. The levels of noggin and, to a lesser extent, gremlin were also increased by BMPs. BMP's stimulatory effects on SOST were further enhanced by retinoic acid or 1,25-dihydroxyvitamin D3. In contrast, dexamethasone (DEX) blocked the effects of the BMPs on SOST and gremlin, but not on noggin. Retinoic acid and 1,25-dihydroxyvitamin D3 did not affect the BMP-enhanced expression of gremlin or noggin. The steroids did not affect the endogenous levels of the BMP antagonists. These findings show that the levels of SOST are modulated by BMPs and the interactions of the BMPs with steroid hormones in human osteoblasts. These effects differed markedly from that of noggin or gremlin, suggesting that there is an exquisite regulation of the expressions of BMP antagonists in cells of the osteoblast lineage. PMID- 15268897 TI - FGF23 is processed by proprotein convertases but not by PHEX. AB - X-linked hypophosphatemia (XLH) and autosomal dominant hypophosphatemic rickets (ADHR) are characterized by renal phosphate wasting, rickets, and osteomalacia. ADHR is caused by gain of function mutations in the fibroblast growth factor 23 gene (FGF23). During secretion, FGF23 is processed at the C-terminus between amino acids 179 and 180. The cleavage site is mutated in ADHR, preventing processing of FGF23. Here, we show that FGF23 is likely to be cleaved by subtilisin-like proprotein convertases (SPC) as cleavage can be inhibited by a specific SPC inhibitor in HEK293 cells. SPCs, which are widely expressed, were demonstrated to be also present in HEK293 cells as well as in osteoblasts. XLH is caused by loss of function mutations in the putative endopeptidase PHEX. It was tempting to speculate that FGF23 is a substrate of PHEX, but studies have been inconclusive so far. Here, we used a secreted form of PHEX (secPHEX) and tagged and untagged FGF23 constructs for co-incubation experiments. These experiments provided evidence against cleavage of intact FGF23(25-251) as well as of N terminal (FGF23(25-179)) and C-terminal (FGF23(180-251)) fragments by the endopeptidase PHEX. PMID- 15268898 TI - Expression of ICAM-1 by osteoblasts in healthy individuals and in patients suffering from osteoarthritis and osteoporosis. AB - OBJECTIVES: To describe the pattern of expression of intercellular adhesion molecule-1 (ICAM-1) at the surface of human osteoblasts (Ob) recovered from normal (control), osteoporotic (OP), and osteoarthritic (OA) bone. To relate ICAM 1 expression in OA Ob with interleukin-6 (IL-6) and prostaglandin E2 (PGE2) production. MATERIALS AND METHODS: Trabecular bone specimens were taken from patients suffering from OA of the hip (n = 19) or knee (n = 19) or from hip fracture caused by osteoporosis (n = 10). Control bone specimens came from the posterosuperior iliac crest (n = 5) and from the femoral condyle of organ donors (n = 6). Bone explants were digested with collagenase and cultured. Ob were obtained after 6 weeks. ICAM-1 expression was studied by immunocytology. IL-6 and PGE2 were evaluated by standard ELISA. RESULTS: Average ICAM-1 expression was different between control and OP bone (P < 0.02). Separation of specimens into high and low ICAM-1 expression showed a significant difference between high and low ICAM-1 expressors. The distribution of specimens after subclassification into high or low ICAM-1 expression groups revealed only 18.2% of patients in the high expression group for the controls, compared to 70% for OP bone (P < 0.03), 52.6% for hip OA and 47.4% for knee OA. IL-6 and PGE2 levels in OA Ob from both groups were found to be significantly elevated with high ICAM-1 expression compared to low ICAM-1 expression. CONCLUSION: The results show that ICAM-1 expression in human bone seems to be pathology-dependent and correlates with IL-6 and PGE2 production, at least in OA individuals. This implies that ICAM-1 could discriminate functionally different populations of Ob and possibly alter the clinical evolution of the disease. PMID- 15268899 TI - Chemically modified tetracyclines act through multiple mechanisms directly on osteoclast precursors. AB - Chemically modified tetracyclines (CMTs) are thought to inhibit bone resorption primarily through their ability to inhibit matrix metalloproteinases (MMPs). We have previously demonstrated that some tetracycline compounds (TCs) induce apoptosis in mature rabbit osteoclasts and inhibit osteoclastic resorption in mouse osteoblast/marrow co-cultures in vitro. In this report, we now show that non-antibiotic analogues of doxycycline (CMT-3) and minocycline (CMT-8) are potent inhibitors of osteoclastogenesis in vitro from human peripheral blood mononuclear cells (PBMC) stimulated with macrophage colony stimulating factor (MCSF) and receptor activator of NF-kappaB ligand (RANKL), through an action that is independent of osteoblast-osteoclast interactions. Osteoclast formation over 20 days was completely abrogated when CMT-3 or CMT-8 were included in PBMC cultures at a concentration of 250 ng/ml, although doxycycline at this concentration reduced osteoclast formation to ca. 50% of control. CMT-3 and CMT-8 also significantly induced apoptosis over 24 h in mature osteoclasts generated over 20 days when added to cultures at 5 microg/ml or more. In a time-course experiment, apoptosis was evident after a delay of 1-2 h following treatment of mature osteoclasts with CMT-3 at 20 microg/ml. The broad-spectrum MMP inhibitor BB94 (Batimastat) did not recapitulate the apoptosis induced by CMT-3, even at a concentration where MMP-13 activity was completely inhibited. There was no evidence for an anabolic effect of any of the TCs on osteoblast lineage cells in a calcifying fibroblastic colony (CFU-f) formation assay, where CMT-3 partially inhibited CFU-f formation at 5 microg/ml. Our data indicate that inhibition of osteoclast formation and induction of osteoclast apoptosis are pharmacologically significant actions of CMTs in inhibiting bone resorption, and that osteoclast apoptosis cannot be attributed to the ability of CMTs to inhibit MMPs or to actions mediated by osteoblastic lineage cells. PMID- 15268900 TI - Extracellular calcium downregulates estrogen receptor alpha and increases its transcriptional activity through calcium-sensing receptor in breast cancer cells. AB - Skeleton is the most common organ targeted by breast cancer cells, especially from estrogen receptor alpha (ER)-positive neoplasms. Metastatic cells can stimulate directly or indirectly osteoclast-mediated bone resorption. Tumor induced osteolysis is often extensive and leads to the release of large quantities of calcium. Metastatic cancer cells can be thus exposed to high calcium concentrations (40 mM has been reported at the resorption site). However, the effects of Ca2+ on breast cancer cells have been minimally examined. We showed that 20-mM extracellular Ca2+ induced a downregulation of ER protein in MCF-7 cells and caused ER-mediated transactivation of a reporter gene by 55 +/- 10% (mean +/- SD) in MVLN cells (MCF-7 cells stably transfected with ERE and luciferase reporter gene). Moreover, 3 mM Ca2+ increased progesterone receptor (PgR) expression by 45 +/- 8%. Mg2+ tested at up to 20 mM did not exert any effects, while 17beta-estradiol downregulated ER, transactivated the reporter gene, and enhanced PgR expression. The pure antiestrogen ICI 182,780 was able to abrogate the transactivation of the reporter gene and the increase in PgR levels induced by Ca2+, indicating that Ca2+ may exert a weak and specific estrogenic effect in MCF-7 cells. Ca2+ effects on ER probably start at the cell membrane level since a large Ca2+ influx caused by the ionophore A23187 failed to activate ER. We have thus studied the involvement of the membrane calcium-sensing receptor (CaR) that is known to be expressed notably in MCF-7 cells. We first tested the effects of a specific activator of CaR. Exposure to 10(-4) M calcimimetic NPS R 467 mirrored the changes observed with extracellular Ca2+ by inducing a marked decrease in ER protein levels, increasing the transcriptional activity of ER (67 +/- 12%) and stimulating PgR expression (41 +/- 4%). As expected, the NPS S-467 isomer was less effective. Furthermore, a highly selective CaR antagonist partly suppressed the downregulation of ER as well as transactivation of the reporter gene induced by Ca(2+). Our results suggest that the effects of extracellular Ca2+ on ER expression and activity are mediated, at least in part, by the CaR. In summary, calcium released during the process of metastatic bone destruction could modulate the functions of the estrogen receptor, a key receptor involved in breast cancer cells growth and function, and thus participate in the pathogenesis of tumor-induced osteolysis. PMID- 15268901 TI - Daidzein together with high calcium preserve bone mass and biomechanical strength at multiple sites in ovariectomized mice. AB - As the prevalence of osteoporosis is increasing, and the adverse effects of hormone replacement therapy are evident, women are searching for natural alternatives such as soy isoflavones to help prevent postmenopausal osteoporosis. Daidzein is one of the most abundant isoflavones present in soy and it is unique as it can be further metabolized to equol, a compound with greater estrogenic activity than other isoflavones. The objective of this study was to determine the effects of purified daidzein in combination with high calcium (Ca) on preserving femur and lumbar vertebrae (LV1-LV4) bone mineral density (BMD) and biomechanical bone strength at three different sites (femur midpoint, femur neck and LV3) in ovariectomized mice. Sham (SH) mice (n = 12) received control diet (AIN93G) containing 2 g Ca/kg diet and ovariectomized mice were randomized to 1 of 6 groups (n = 12/group): OVX (2 g Ca/kg diet), HCa (25 g Ca/kg diet), HD (2 g Ca + 200 mg daidzein/kg diet), HDCa (25 g Ca + 200 mg daidzein/kg diet), LD (2 g Ca + 100 mg daidzein/kg diet) or LDCa (25 g Ca + 100 mg daidzein/kg diet) for 12 weeks. HDCa preserved femur and vertebrae BMD and biomechanical bone strength (at all three sites) compared to the OVX group, however, only femur yield load (at midpoint) was preserved to a level that was greater (P < 0.05) than HCa alone. Mice fed HD diet had greater (P < 0.05) femur BMD than OVX group, however, daidzein alone (HD) did not appear to preserve trabecular bone (i.e., vertebrae BMD and vertebra peak load). All mice fed daidzein produced equol and there were no uterotrophic effects of daidzein at either dose. Both daidzein and Ca attenuated the increase in serum IL-1beta observed in the OVX group. The results from this study suggest that the combination of daidzein and high Ca favorably affect cortical and trabecular bone as indicated by femur and lumbar vertebrae BMD and biomechanical strength but much of this effect is mediated by the high Ca diet. Further investigation is required to determine optimal dietary levels of daidzein and Ca with the long-term goal of developing a dietary strategy to prevent postmenopausal osteoporosis and related fragility fractures. PMID- 15268902 TI - Alterations in mineral composition observed in osteoarthritic joints of cynomolgus monkeys. AB - Osteoarthritis (OA) is a prevalent joint disease that affects more than 40 million Americans and is characterized by degeneration of the articular cartilage and thickening of the underlying subchondral bone. Although subchondral bone thickening has been implicated in articular cartilage degeneration, very little is known about the composition of subchondral bone in OA. In the present study, infrared microspectroscopy (IRMS) was used to determine the chemical composition of the calcified cartilage-subchondral bone plate in a monkey model of OA. Specifically, the levels of mineralization (mineral/protein ratio), carbonate accumulation (carbonate/protein ratio), crystallinity, and collagen structure were determined as a function of animal age and OA severity. OA severity was assessed using a grading scheme that included scores or measurements for several histomorphometric parameters including articular cartilage fibrillation or clefting, subchondral bone thickness, and numbers of tidemarks and chondrocyte clones. Individual scores and measurements were summarized using principal components (factor) analysis. Results demonstrated that the level of mineralization and carbonate content increased as a function of animal age. In addition, bone mineralization level increased as subchondral bone thickness increased. Dramatic increases in the mineralization level and carbonate accumulation were also observed as a function of the number of tidemarks. The presence of multiple tidemarks indicates the occurrence of one or more additional phases of cartilage calcification, suggesting that the observed compositional changes are due to cartilage mineralization. Our results support a reactivation of endochondral ossification that occurs with age, which is more pronounced in OA. No relationships were observed between mineral crystallinity and collagen cross-linking as a function of age or OA severity. In summary, compositional analysis of the mineralized plate beneath the articular cartilage in OA is characterized by thickened, overmineralized calcified cartilage or subchondral bone, which likely puts added mechanical stress on the joint, contributing to the progression of OA. PMID- 15268903 TI - Dual roles for NF-kappaB activation in osteoblastic cells by serum deprivation: osteoblastic apoptosis and cell-cycle arrest. AB - To clarify the mechanisms of osteoblastic cell death, we examined whether serum deprivation would cause activation of the apoptotic signal cascade and arrest of the cell cycle in mouse osteoblastic MC3T3-E1 cells. Serum withdrawal from osteoblastic cell cultures resulted in growth arrest and cell-cycle arrest at G0/G1, which actions were accompanied by transient and potent activation of NF kappaB, caspase-8, caspase-2, caspase-3, and caspase-9 in this order. Apoptosis, but not necrosis, in serum-deprived cells could be detected by FACS using Annexin V/propidium iodine double staining. Serum deprivation also resulted in transient activation of the 20S proteasome, which is an important component for regulation of the cell cycle by the ubiquitin-proteasome system. The 20S proteasome inhibitor (PSI) but not NF-kappaB inhibitor SN50 suppressed the activation of proteasomes in serum-deprived cells. Although caspase inhibitors could not prevent the G0/G1 arrest in the serum-deprived cells, SN50 and the 20S proteasome inhibitor could block it. Since SN50, 20S proteasome inhibitor and caspase inhibitor could rescue cells from serum deprivation-induced apoptosis, the pathway for NF-kappaB/caspase activation is independent of the NF-kappaB/cell cycle pathway, and the events downstream of the NF-kappaB/caspase-9 cascade lead to apoptosis. Taken together, our present results identify a novel role for NF kappaB in cell-cycle and apoptosis regulation and underscore the significance of each independent signal cascade in serum-deprived osteoblastic cells. PMID- 15268904 TI - In vitro growth and osteoblastic differentiation of human bone marrow stromal cells supported by autologous plasma. AB - Autologous bone marrow stromal cells have been proposed as an adjuvant in the treatment of bone nonunion. This cell therapy requires the establishment of culture conditions that permit the rapid expansion of these cells ex vivo while retaining their potential for further differentiation. Several culture models have been proposed, all of them using fetal calf serum (FCS) as a source of growth factors. This is problematic for subsequent autologous implantation because of possible disease transmission. Here we report the establishment and characterization of a cell culture system in which standard FCS has been replaced by autologous plasma recovered from bone marrow (APM). Short-term cultures of human bone marrow stromal (HBMS) cells grown in mineralizing conditions with APM exhibited a significantly higher number of ALP-positive colonies than those grown with FCS, indicating an enhanced ability of APM to recruit osteoprogenitor cells for culture. Analyses of long-term cultures showed that the use of APM did not affect cell proliferation as cell number at confluence and proliferation rate were similar whether primary cultures had been maintained with APM or FCS. In first-passage cultures, an osteoblastic differentiation was observed in both cases as the cells expressed ALP and formed mineralized bone-like nodules. We noted that the age of donor had a negative effect on the number of osteoprogenitor cells recruited for culture. This effect had an impact on proliferation rate in primary cultures performed with APM, although the cell number obtained after expansion remained independent of age. Our study shows that proliferative capacity and osteoblastic differentiation potential of HBMS cells are maintained when cultured with APM. Thus, this cell culture system could provide a new and safer tool to elaborate an autologous cell therapy designed to enhance osteogenesis. PMID- 15268905 TI - Extracellular signal-regulated kinase-1 and -2 are both essential for the shear stress-induced human osteoblast proliferation. AB - Extracellular signal-regulated kinases (Erk)-1 and -2 are key mediators of various mitogenic signaling pathways, including mechanical stress-induced osteoblast proliferation. Mechanical stimuli, such as flow shear stress, simultaneously activate both Erk-1 and -2 in osteoblasts, resulting in stimulation of osteoblast proliferation. This study sought to test whether Erk-1, -2, or both are essential for the fluid flow shear stress-induced osteoblast proliferation. Moloney leukemia virus (MLV)-based vectors expressing wild-type (wt)- or kinase-deficient (kd) Erk-1 and Erk-2, respectively, were constructed and used to transduce human TE85 osteosarcoma cells with an MOI of 30. An MLV-red fluorescent protein (RFP) vector was included as a control. Effects of Erk-1 and 2 overexpression on cell proliferation in response to a 30-min constant fluid flow shear stress at 20 dynes/cm2 were determined with [3H]thymidine incorporation 24 h after the shear stress. The MLV-Erk vector-transduced TE85 cells showed a >10- and approximately 2-fold overexpression of Erk-1 and -2 protein, respectively. The RFP expressing control cells and the parental TE85 cells each showed an approximately twofold increase (P < 0.01) in [3H]thymidine incorporation in response to the shear stress. Cells overexpressing wt-Erk-1 or 2 showed small enhancing effects on the response to the shear stress in the increases in [3H]thymidine incorporation and cell number. Conversely, overexpression of kd-Erk-1 or -2 each alone completely abolished the shear stress induced osteoblast proliferation. Overexpression of either kd-Erk-1 or kd-Erk-2 alone did not have a significant effect on basal osteoblast proliferation, suggesting that the Erk signaling pathway may not be essential for basal cell proliferation. In summary, this study demonstrates for the first time that Erk-1 and -2 are both required for the mitogenic response to fluid flow shear stress in human osteoblasts and that blocking Erk-1 or -2 each alone is sufficient to completely block the mitogenic response to shear stress-induced proliferation. PMID- 15268906 TI - Microvascular invasion during endochondral ossification in experimental fractures in rats. AB - In this study morphologic techniques have been used to detail the angiogenic response that accompanies endochondral fracture healing in a clinically relevant, reproducible rat model. In this displaced fracture, the gap fills with cartilage that later is replaced by bone, via endochondral ossification. A transient periosteal circulation, followed by a permanent medullary circulation accompany this progression. From 2 to 6 weeks, vessels grow out from the periosteal tissue and give rise to vascular buds, which abut directly onto the avascular zone corresponding to the fracture defect. From 3 weeks onwards, a second wave of vessels grows out from the marrow to the cartilage-filled fracture defect, terminating as vascular buds and loops lined by endothelial and perivascular cells. The loops and buds stain strongly for laminin but transmission electron microscopy does not demonstrate an identifiable basement membrane, pointing to a region of active extracellular matrix turnover. These vessels are intimately associated with osteoblasts and newly formed woven bone forming finger-like composite structures that protrude into the mineralized cartilage matrix with which they form a clearly demarcated interface. Invading vessels and woven bone successively replace the cartilage matrix to mediate repair. Both the vascular structures and progression of endochondral ossification observed, closely resemble those described in the normal epiphyseal growth plate, indicating that the fundamental processes are similar. However, there is a difference in the spatial orientation of cells such that the healing front in the fracture model is relatively disorganized, compared to the orderly linear array of cells at the epiphyseal growth plate. PMID- 15268907 TI - VEGF expression in adult permanent thyroid cartilage: implications for lack of cartilage ossification. AB - Vascular endothelial growth factor (VEGF) has been shown to play an important role during endochondral bone formation in hypertrophic cartilage remodeling, ossification, and angiogenesis, but it is not expressed in normal adult articular cartilage. Thyroid cartilage undergoes only partial ossification beginning at the age of about 20. Because it never completely ossifies, we investigated a possible role of VEGF and its receptors (VEGFRs) as well as the angiogenetic inhibitor endostatin in this permanent cartilage. In analysis of cartilage samples from all specimens evaluated, VEGF121 and VEGF165 were identified as the only VEGF splice forms expressed. In addition to VEGF, VEGFR-2 (kinase domain region/fetal liver kinase 1), but not VEGFR-1 (fms-like tyrosine kinase 1), was detectable by RT-PCR in cartilage. However, VEGFR-2 expression was only detectable up to the age of 19 years. Deposition of VEGF and VEGFR was confirmed by immunohistochemistry. VEGF concentrations measured by ELISA in thyroid cartilage increased with age in males but decreased in females. Endostatin concentrations measured by ELISA in thyroid cartilage were three times lower than in articular cartilage and showed no change with age, either in females or males. VEGF was immunostained within the intra- and pericellular matrices of some but not all chondrocytes. Thus, apart from its production in hypertrophic chondrocytes of growth plates, VEGF is also produced in single chondrocytes of thyroid cartilage. The data allow us to speculate that thyroid cartilage persists in an embryological state until it has reached its final size. After reaching its final size at the end of the second decade, VEGFR 2 is downregulated and ossification starts in the posterior part of the thyroid cartilage, proceeding ventrally. Both proteins, VEGF121 and VEGF165, should contribute to this process. VEGF concentration is high and changes in an age related and sex-specific manner. Therefore, we postulate that VEGF is at least one of the key factors that is important for the lifelong ossification in thyroid cartilage. PMID- 15268908 TI - Expression of dentin matrix protein 1 (DMP1) during fracture healing. AB - Dentin matrix protein 1 (DMP1) is one of the acidic phosphorylated extracellular matrix proteins called the SIBLING (small integrin-binding ligand, N-linked glycoproteins) family. Recent studies showed that DMP1 is expressed in the mineralized tissues and suggested that DMP1 is involved in the mineralization. We investigated the precise localization of DMP1 messenger RNA (mRNA) and protein during fracture healing. In situ hybridization demonstrated that DMP1 mRNA was strongly expressed in preosteocytes and osteocytes in the bony callus during intramembranous and endochondral ossification while DMP1 mRNA was not detected in osteoblasts and chondrocytes. During endochondral ossification, however, a low number of DMP1-expressing cells were identified in the cluster of hypertrophic chondrocytes. However, these DMP1-expressing cells were not hypertrophic and were likely to be osteoblast-lineage cells, which were embedded in the matrix of bone or cartilage, because type I collagen-expressing cells and invasion of capillary vessels were observed in the same area. Northern blot, in situ hybridization, and immunohistochemical analyses showed that DMP1 mRNA and protein expressions were increased until day 14 postfracture, when bony callus was formed, and then declined to a lower level during remodeling of the bony callus. Therefore, DMP1 is likely to play an important role in the mineralization of the bony callus. PMID- 15268909 TI - Dual growth factor delivery and controlled scaffold degradation enhance in vivo bone formation by transplanted bone marrow stromal cells. AB - Supraphysiological concentrations of exogenous growth factors are typically required to obtain bone regeneration, and it is unclear why lower levels are not effective. We hypothesized that delivery of bone progenitor cells along with appropriate combinations of growth factors and scaffold characteristics would allow physiological doses of proteins to be used for therapeutic bone regeneration. We tested this hypothesis by measuring bone formation by rat bone marrow stromal cells (BMSCs) transplanted ectopically in SCID mice using alginate hydrogels. The alginate was gamma-irradiated to vary the degradation rate and then covalently modified with RGD-containing peptides to control cell behavior. In the same delivery vehicle, we incorporated bone morphogenetic protein-2 (BMP2) and transforming growth factor-beta3 (TGF-beta3), either individually or in combination. Individual delivery of BMP2 or TGF-beta3 resulted in negligible bone tissue formation up to 22 weeks, regardless of the implant degradation rate. In contrast, when growth factors were delivered together from readily degradable hydrogels, there was significant bone formation by the transplanted BMSCs as early as 6 weeks after implantation. Furthermore, bone formation, which appeared to occur by endochondral ossification, was achieved with the dual growth factor condition at protein concentrations that were more than an order of magnitude less than those reported previously to be necessary for bone formation. These data demonstrate that appropriate combinations of soluble and biomaterial mediated regulatory signals in cell-based tissue engineering systems can result in both more efficient and more effective tissue regeneration. PMID- 15268910 TI - Evidence of downregulation of matrix extracellular phosphoglycoprotein during terminal differentiation in human osteoblasts. AB - Matrix extracellular phosphoglycoprotein (MEPE) is an extracellular matrix protein that was first detected in tumor-induced osteomalacia (TIO). Investigations in mice revealed that MEPE is expressed in bone and teeth in a maturation-dependent manner, reaching its maximum during mineralization. However, from knockout experiments, although it has become clear that MEPE might function as a mineralization inhibitor, the exact mechanism of action is still unclear. Even less is known about the regulation of MEPE in men. Therefore, we have studied the time- and maturation-dependent expression of MEPE in two human osteoblast culture systems, the osteosarcoma cell line HOS 58 and primary trabecular osteoblasts. Cells were cultured for up to 29 days, and the influence of beta-glycerophosphate (bGP), ascorbate, transforming growth factor beta (TGF beta), BMP-2, and dexamethasone was studied. HOS 58 cells showed no significant effect on MEPE gene expression up to 5.0 mM, but a significant inhibition was revealed at 10 and 20 mM, when osteocalcin (OC) expression was maximal. Under the same conditions, primary human osteoblasts showed no effect on MEPE gene expression. However, when cultured in the presence of 5 mM beta-glycerophosphate, ascorbate, and dexamethasone for 29 days, which are similar conditions to those described by Owen in his differentiation model in rat osteoblasts, a progressive inhibition of MEPE gene expression to 20% of the maximum was observed. Increasing osteocalcin expression indicated advancing differentiation. In conclusion, in contrast to the results in mice, when MEPE was maximally expressed during mineralization, in the human system, this factor seems to be maximally active in the proliferation and early matrix maturation phase. It was, however, strongly suppressed, associated with the mineralization phase. PMID- 15268911 TI - Perspectives on calcium homeostasis. PMID- 15268912 TI - Frontal and posterior sources of event-related potentials in semantic comprehension. AB - An important question in brain and language research is how activity in multiple brain networks is coordinated over time during semantic comprehension. To address this question, we applied spatiotemporal source analysis to event-related potentials (ERPs) recorded as subjects read words that were meaningful or incongruous in the context of a sentence (N400 paradigm). The incongruous word was placed either early in the sentence or at the end. Source analysis showed activity in language areas of the left hemisphere, right temporal cortex and medial limbic cortex. The initial detection of semantic incongruity (approximately 250 ms) engaged the left prefrontal cortex and left anterior cingulate. In the critical (300-500 ms) interval, regional sources in left and right lateral prefrontal cortex, right temporal cortex, and both anterior and posterior cingulate were responsive to the semantic manipulation. Left hemisphere activity preceded right hemisphere activity, and semantic effects in frontal regions began earlier and were more sustained than the transient effects within posterior cortical regions. Findings are discussed with respect to recent theories of corticothalamic and corticolimbic networks in attention and semantic processing. PMID- 15268913 TI - Contribution of tactile and interoceptive cues to the perception of the direction of gravity. AB - Without relevant visual cues, the Subjective Visual Vertical (SVV) is biased in roll tilted subjects toward the body axis (Aubert or A-effect). This effect is generally ascribed to changes in the vestibular and somatosensory inputs following a body tilt. This study focused on the contribution of interoception and tactile cues in the SVV. The body-cast technology and gastric fullness were used to obtain a diffuse tactile stimulation and an overload stomach stimulation, respectively. Fifteen subjects placed in a tilt-chair were rolled sideways from 0 degrees to 90 degrees. They were asked to adjust a luminous line to the vertical under two body restriction conditions (strapped vs. body-cast) and two stomach load conditions (empty vs. full). Results showed (1) an improvement in the SVV judgment when somaesthetic cues were available in the full stomach condition (p < 0.001), (2) an increased A-effect for the higher body tilt values in the body cast condition (beyond 45 degrees, p < 0.001), and (3) a smaller disrupting effect of the body-cast in the SVV judgment in the full stomach condition (p < 0.05). Since the vestibular system produced the same gravity response in all conditions, it can be stated that somaesthetic cues are involved in the SVV. Tactile mechanoreceptors may have contributed by detecting the changing pattern of pressures generated on the skin that results from changes in body orientation. The stomach load may act through the inertial forces exerted against the gravity load when the stomach is full by the mechanoreceptors in the fundus. Thus, the somaesthetic system can indicate the direction of gravity via patterns of pressure within and at the surface of the body. PMID- 15268914 TI - The song system of the human brain. AB - Although sophisticated insights have been gained into the neurobiology of singing in songbirds, little comparable knowledge exists for humans, the most complex singers in nature. Human song complexity is evidenced by the capacity to generate both richly structured melodies and coordinated multi-part harmonizations. The present study aimed to elucidate this multi-faceted vocal system by using 15O water positron emission tomography to scan "listen and respond" performances of amateur musicians either singing repetitions of novel melodies, singing harmonizations with novel melodies, or vocalizing monotonically. Overall, major blood flow increases were seen in the primary and secondary auditory cortices, primary motor cortex, frontal operculum, supplementary motor area, insula, posterior cerebellum, and basal ganglia. Melody repetition and harmonization produced highly similar patterns of activation. However, whereas all three tasks activated secondary auditory cortex (posterior Brodmann Area 22), only melody repetition and harmonization activated the planum polare (BA 38). This result implies that BA 38 is responsible for an even higher level of musical processing than BA 22. Finally, all three of these "listen and respond" tasks activated the frontal operculum (Broca's area), a region involved in cognitive/motor sequence production and imitation, thereby implicating it in musical imitation and vocal learning. PMID- 15268915 TI - Alpha rhythm of the EEG modulates visual detection performance in humans. AB - The effects of the changes in the frequency spectrum of the electroencephalogram (EEG) on the perception of near-threshold visual stimuli and on the event-related potentials (ERPs) produced by these stimuli were investigated on 12 healthy volunteers. The stimulus intensity, at which each subject could detect 50% of the presented stimuli, was defined as the sensory threshold for that subject. Single ERP trials were separated into two groups: trials with detected and undetected stimuli. The ERPs and the average power spectra of the 1 s prestimulus periods were computed for both conditions. P300 amplitudes of the ERPs, and total power and relative band powers of the delta (0.5-4 Hz), theta (4-7.5 Hz), alpha (7.5-13 Hz), beta (13-30 Hz), and gamma (30-70 Hz) frequency bands of the prestimulus power spectra were measured. Between the two conditions, a specific difference was observed in the relative power of the alpha band, which was significantly lower before detected stimuli (p < 0.01) in line with significantly higher amplitudes of the ERPs (p < 0.001). These results show that short-lasting changes in brain's excitability state are reflected the relative alpha power of the EEG, which may explain significant variability in perceptual processes and ERP generation especially at boundary conditions such as sensory threshold. PMID- 15268916 TI - Regional brain response to faces of humans and dogs. AB - The extent to which the brain regions associated with face processing are selective for that specific function remains controversial. In addition, little is known regarding the extent to which face-responsive brain regions are selective for human faces. To study regional selectivity of face processing, we used functional magnetic resonance imaging to examine whole brain activation in response to human faces, dog faces, and houses. Fourteen healthy right-handed volunteers participated in a passive viewing, blocked experiment. Results indicate that the lateral fusiform gyrus (Brodmann's area 37) responds maximally to both dog and human faces when compared with other sites, followed by the middle/inferior occipital gyrus (BA 18/19). Sites that were activated by houses versus dog and human faces included the medial fusiform gyrus (BA 19/37), the posterior cingulate (BA 30), and the superior occipital gyrus (BA 19). The only site that displayed significant differences in activation between dog and human faces was the lingual/medial fusiform gyrus. In this site, houses elicited the strongest activation, followed by dog faces, while the response to human faces was negligible and did not differ from fixation. The parahippocampal gyrus/amygdala was the sole site that displayed significant activation to human faces, but not to dog faces or houses. PMID- 15268917 TI - Neural correlates of metaphor processing. AB - Metaphoric language is used to express meaning that is otherwise difficult to conceptualize elegantly. Beyond semantic analysis, understanding the figurative meaning of a metaphor requires mental linkage of different category domains normally not related to each other. We investigated processing of metaphoric sentences using event-related functional magnetic resonance imaging (fMRI). Stimuli consisted of 60 novel short German sentence pairs with either metaphoric or literal meaning. The pairs differed only in their last one to three words and were matched for syntax structure, word frequency, connotation and tense. Fifteen healthy subjects (six female, nine male, 19-51 years) read these sentences silently and judged by pressing one of two buttons whether they had a positive or negative connotation. Reading metaphors in contrast to literal sentences revealed signal changes in the left lateral inferior frontal (BA 45/47), inferior temporal (BA 20) and posterior middle/inferior temporal (BA 37) gyri. The activation in the left inferior frontal gyrus may reflect semantic inferencing processes during the understanding of a metaphor. This is in line with the results from other functional imaging studies showing an involvement of the left inferior frontal gyrus in integrating word and sentence meanings. Previous results of a right hemispheric involvement in metaphor processing might reflect understanding of complex sentences. PMID- 15268918 TI - Behavioural and physiological impairments of sustained attention after traumatic brain injury. AB - Sustaining attention under conditions of low external demand taxes our ability to stay on task and to avoid more appealing trains of thought or environmental distractions. By contrast, a stimulating, novel environment engages attention far more freely without the subjective feeling of having to override monotony. Our ability to maintain a goal-directed focus without support from the environment requires the endogenous control of behaviour. This control can be modulated by fronto-parietal circuits and this ability is compromised following traumatic brain injury (TBI) leading to increased lapses of attention. In this paper, we further explore a laboratory paradigm that we argue is particularly sensitive to sustained attention as opposed to other aspects of attentional control involving the selection and management of goals in working memory. The paradigm (fixed sequence Sustained Attention to Response Task--SARTfixed) involves withholding a key press to an infrequent no-go target embedded within a predictable sequence of numbers. We demonstrate that TBI patients in this study make disproportionately more errors than controls on this task. An analysis of response times (RTs) and EEG alpha power across the task demonstrates group differences preceding the critical no-go trial. Controls demonstrate a lengthening of RTs accompanied by desynchronization of power within the alpha band (approximately 10 Hz) preceding the no-go trial. Conversely, the TBI group showed a shortening of RTs during this period with no evidence of alpha desynchronization. These findings suggest that TBI patients may have dysfunctional alpha generators as a consequence of their injury that impairs endogenous control during the task. PMID- 15268919 TI - Electrophysiological correlates of object categorization: back to basics. AB - The time course of visual object categorization as a function of electrophysiological activity in the brain was investigated using a variant of the "oddball" design. Category level was manipulated by sequentially presenting subordinate, basic or superordinate target objects among a variety of non-target objects. It was found that superordinate categorizations were performed more quickly and differentiated from basic level categorizations in amplitude early in visual processing (320-420 ms). In contrast, subordinate categorizations took longer to perform and differentiated from basic level categorizations in amplitude and latency at later stages (450-550 ms). Notably, these effects were observed using the same objects categorized at different levels suggesting that visually categorizing objects at varying levels of abstraction engaged specific cognitive processes. These results are consistent with research on rapid visual categorization that challenges the generality of basic category level superiority effects. PMID- 15268920 TI - Processing of affective prosody and lexical-semantics in spoken utterances as differentiated by event-related potentials. AB - In the current study, event-related potentials (ERPs) were utilized to assess whether ERP correlates would distinguish between prosodic and lexical-semantic information processed during the comprehension of a spoken affective message. To this end, we employed a standard oddball paradigm with stimuli varying in lexical semantic or prosodic characteristics. An N400 component was obtained in response to all stimuli and conditions (non-targets and targets). Greater negativity in the N400 amplitude was observed in response to semantic as compared to prosodic stimuli. An anterior (P3a) positive component was increased for prosodic as compared to semantic targets. We also investigated whether an N400 and/or P3a component would be present when a stimulus carried affective semantic and affective prosodic information. The ERP structure observed in response to targets of this condition showed a reduction in the amplitude of the N400 component and an explicit anterior P3a component, significantly greater than the P3a component in response to prosodic or semantic targets. Finally, a P3b component was evoked in response to targets, regardless of communicative dimension. PMID- 15268921 TI - Dopaminergic modulation of response inhibition: an fMRI study. AB - Dopamine has been hypothesized to modulate response inhibition. To test this hypothesis, we used functional magnetic resonance imaging (fMRI) to measure the effects of the dopamine prodrug levodopa on the brain responses to a well validated response inhibition task (go/no-go, or GNG). Since abnormalities of response inhibition and dopamine have been thought to underlie tics and other symptoms of Tourette syndrome, we studied 8 neuroleptic-naive adults with tic disorders as well as 10 well-matched healthy controls. Subjects were pretreated with the peripheral decarboxylase inhibitor carbidopa, then scanned during GNG and control blocks, both before and during i.v. levodopa infusion. Both groups had similar task performance and task-related regional brain activity before and during levodopa infusion. Levodopa did not affect reaction times or accuracy, so fMRI findings can be interpreted without concern that they simply reflect a performance difference between conditions. Levodopa did affect the magnitude of GNG-related fMRI responses in the right cerebellum and right parietal cortex, significantly reducing both. Pre-levodopa activity in the right cerebellum correlated with reaction times (higher magnitudes associated with faster reaction times), and pre-levodopa activity in the right parietal cortex correlated with false alarm rate (higher magnitudes associated with higher error). In summary, right parietal and cerebellar regions important in mediating specific aspects of the GNG task were modulated by levodopa, suggesting a region-specific role for dopamine in response inhibition. PMID- 15268922 TI - Residual behavioral and neuroanatomical effects of short-term chronic ethanol consumption in rats. AB - The residual effects of short-term chronic ethanol consumption were investigated in rats maintained on an ethanol liquid diet for 26 consecutive days (mean intake = 16.1 g/kg/day). Animals were assessed for spontaneous motor activity (12 days post-ethanol), spatial working memory (17 days post-ethanol), spatial reference memory (184 days post-ethanol), and retention of passive avoidance (201 days post ethanol). Measurements of brain weights and cortical thickness vertices within the dorsomedial and ventrolateral cortex of eight coronal planes were determined 260 days post-ethanol. Two-dimensional cell profile densities within six coronal planes and within CA1 region of the hippocampus were also obtained, along with the total volumetric measurement of the hippocampus proper. Results indicated between group differences when subjects were assessed on working memory with ethanol-treated animals exhibiting longer escape latencies in a Morris water maze, an effect partially attributed to the perseverance of ethanol-treated animals in exhibiting thigmotaxicity. No other ethanol-related behavioral impairment was noted. Neuroanatomically, ethanol-treated rats had thinner cortical mantles (6.3% and 6.6% reductions) within the frontoparietal cortex and had lower two-dimensional cell profile densities within the most caudal cortical region studied. Interestingly, control animals with thicker cortical mantles tended to perform better on the working memory task, whereas the opposite was true for ethanol-treated subjects. These data led to the conclusion that chronic ethanol consumption of a relatively short duration produces working memory impairments, albeit mild, that are partially related to an inability to abandon ineffectual behavioral strategies, and also produces neuroanatomical alterations within the cortex. PMID- 15268923 TI - The impact of problem structure on planning: insights from the Tower of London task. AB - Despite the large number of behavioral and functional neuroimaging studies employing the Tower of London (ToL), the task's structural parameters and particularly their impact on planning have not been addressed so far. In this paper, we highlight the structural properties of ToL problems and provide evidence for their systematic and substantial effects on the cognitive processes involved in planning. In a problem set with three-move problems, the following structural parameters were experimentally manipulated: the ambiguity of goal hierarchy, the demand for subgoal generation, and the existence of suboptimal alternatives. Analysis of preplanning time as an indicator for the planning process revealed highly significant effects for all three parameters which seems to reflect differences in cognitive processing due to structural task properties. Therefore, we suggest that the common consideration of ToL problem difficulty solely in terms of the minimum number of moves is not sufficient. Moreover, the applied problem sets should be more carefully selected and their structural parameters explicitly noted. PMID- 15268924 TI - Pre-attentive categorization of vowel formant structure in complex tones. AB - It has been demonstrated that vowel information can be extracted from speech sounds without attention focused on them, despite widely varying non-speech acoustic information in the input. The present study tested whether even complex tones that were constructed based on F0, F1 and F2 vowel frequencies to resemble the defining features of speech sounds, but were not speech, are categorized pre attentively according to vowel space information. The Mismatch Negativity brain response was elicited by infrequent tokens of the complex tones, showing that the auditory system can pre-attentively categorize speech information on the basis of the minimal, defining auditory features. The human mind extracts the language relevant information from complex tones despite the non-relevant variation in the sound input. PMID- 15268925 TI - Decrease in prefrontal hemoglobin oxygenation during reaching tasks with delayed visual feedback: a near-infrared spectroscopy study. AB - Visual feedback of hand movement is crucial to accurate reaching. Although previous studies have extensively examined spatial alteration of visual feedback (e.g., prism adaptation), temporal delay of visual feedback has been less explored. In the present study, we investigated the effect of delayed visual feedback of the moving hand in a reaching task. The prefrontal cortical activity was measured by near-infrared spectroscopy (NIRS). Twelve subjects performed reaching tasks under two conditions where visual feedback of their own hand was delayed by 200 ms (delay condition) or 0 ms (normal condition). Introducing the visual feedback delay significantly disrupted the reaching performance, although the subjects gradually adapted to the delay during the experiment. There was a clear tendency to overreach the target in the delay condition, even after the reaching movement had been practiced sufficiently in the normal condition. We observed marked oxy- and total-Hb decreases in the dorsal prefrontal area in the delay conditions. The decrease began shortly after task onset and diminished during the rest period, indicating that the decrease was task-induced. Furthermore, the oxy- and total-Hb decreases were significantly correlated with task performance--the degree of decrease was larger as the subject made more errors. We suggest that the decreases in oxy- and total-Hb at the dorsal prefrontal area are related with the visuomotor recalibration process. PMID- 15268926 TI - Attentional modulation of the human somatosensory evoked potential in a trial-by trial spatial cueing and sustained spatial attention task measured with high density 128 channels EEG. AB - We investigated the modulation of the somatosensory evoked potential (SEP) elicited by mechanical stimuli in a spatial sustained attention and a spatial trial-by-trial cueing design by means of high density electrode array EEG recordings. Subjects were instructed to detect rare tactile target stimuli at the to-be-attended hand while ignoring stimuli at the other hand. Analysis of the SEP revealed a highly complex pattern of results. The P50 component was significantly increased for attended stimuli in the sustained attention as opposed to the trial by-trial cueing condition. However, no difference in amplitude was found for attended as opposed to unattended stimuli. High density electrode array recordings revealed a centero-frontal N140 component (N140c), which preceded the parietal N140 (N140p) by about 20 ms. The N140c exhibited an attention effect in particular in the trial-by-trial spatial cueing condition. The N140p was significantly enlarged with attention across both experimental conditions, but a closer inspection demonstrated that this was mainly due to the great attention effect in the trial-by-trial spatial cueing condition. The late positive component (190-380 ms after stimulus onset) exhibited a significant attention effect in both experimental conditions. The present experiment provides evidence that the attentional modulation of the SEP is different when tactile as opposed to electrical stimuli were used and when only somatosensory stimuli are presented with no further sensory stimulation in other modalities. Furthermore, transient as opposed to sustained spatial attention affected various components of the SEP in a different way. PMID- 15268927 TI - Learned audio-visual cross-modal associations in observed piano playing activate the left planum temporale. An fMRI study. AB - Lip reading is known to activate the planum temporale (PT), a brain region which may integrate visual and auditory information. To find out whether other types of learned audio-visual integration occur in the PT, we investigated "key-touch reading" using functional magnetic resonance imaging (fMRI). As well-trained pianists are able to identify pieces of music by watching the key-touching movements of the hands, we hypothesised that the visual information of observed sequential finger movements is transformed into the auditory modality during "key touch reading" as is the case during lip reading. We therefore predicted activation of the PT during key-touch reading. Twenty-six healthy right-handed volunteers were recruited for fMRI. Of these, 7 subjects had never experienced piano training (naive group), 10 had a little experience of piano playing (less trained group), and the remaining 9 had been trained for more than 8 years (well trained group). During task periods, subjects were required to view the bimanual hand movements of a piano player making key presses. During control periods, subjects viewed the same hands sliding from side to side without tapping movements of the fingers. No sound was provided. Sequences of key presses during task periods consisted of pieces of familiar music, unfamiliar music, or random sequences. Well-trained subjects were able to identify the familiar music, whereas less-trained subjects were not. The left PT of the well-trained subjects was equally activated by observation of familiar music, unfamiliar music, and random sequences. The naive and less trained groups did not show activation of the left PT during any of the tasks. These results suggest that PT activation reflects a learned process. As the activation was elicited by viewing key pressing actions regardless of whether they constituted a piece of music, the PT may be involved in processes that occur prior to the identification of a piece of music, that is, mapping the complex sequence structure of hand movements onto the sequence of sounds. PMID- 15268928 TI - Hemispheric asymmetry for spatially filtered stimuli belonging to different semantic categories. AB - The hemispheric specialization for processing visual stimuli as a function of spatial-frequency content and semantic category was investigated. Spatially filtered pictures of animals and tools were displayed to both hemifields at nine levels of spatial-frequency filtering following a coarse-to-fine design. Results showed a differential hemispheric specialization in relation to the semantic category of stimuli. Animals might be identified at low spatial frequencies (coarse information) in a similar extent in both hemifield conditions, while a hemispheric dissociation was found for tools (the increasing of levels of high spatial frequency information was especially needed when such stimuli were presented to the left visual field /right hemisphere (RH) relative to that for right visual field/left hemisphere presentations). PMID- 15268929 TI - Object structure and saccade planning. AB - When orienting to a newly appearing display, evidence shows that two saccadic eye movements are often prepared together. By looking the relation between the landing positions of the first and the second saccade, we examine the frame of reference used for the preparation of the second saccade aiming for a new object or exploring within the same object. We demonstrate that the action to be performed on the object affects the coding of the second saccade. A second saccade directed to a new object is coded to aim for a target position on it and is adjusted to the landing position of the first saccade, whereas a second saccade within the same object is coded as a fixed motor vector applied irrespective of the initial landing position on the object. PMID- 15268930 TI - When does the brain register deviances from standard word spellings?--An ERP study. AB - Event-related potentials (ERPs) in response to the standard form of words (e.g., taxi) were compared with ERPs in response to letter-altered (e.g., taksi) or case altered forms (e.g., taXi). The altered forms always resulted in the same reading as the standard forms. First divergences between ERPs were found at around 160 ms. At occipital sites, the peak amplitude of the N160 was higher for standard than letter-altered strings. At frontal and central sites, the standard strings diverged from the altered strings persistently by higher positivity from about 160 ms onwards. These early ERP differences between standard and altered visual word forms speak for early contact between the letter input and stored visual orthographic representations of words. PMID- 15268931 TI - Regulation of polyhydroxyalkanoate biosynthesis in Pseudomonas putida and Pseudomonas aeruginosa. AB - Regulation of medium-chain-length polyhydroxyalkanoate biosynthesis in Pseudomonas aeruginosa and Pseudomonas putida was studied conducting PHA accumulation experiments and transcriptional analysis of PHA biosynthesis genes with wild type strains and rpoN-negative mutants. In P. putida PHA accumulation was RpoN-independent, whereas in P. aeruginosa PHA accumulation was RpoN dependent. Transcriptional analysis applying reverse transcriptase-polymerase chain reaction showed strong induction of phaG, encoding the transacylase, under nitrogen starvation in P. putida KT2440 and the respective rpoN-negative mutant, indicating an RpoN-independent regulation of phaG. No transcription of phaG and no PHA accumulation was detected in the rpoN-negative mutant of P. aeruginosa neither from gluconate nor from octanoate as carbon source. Alginate overproducing mutant P. aeruginosa FRD1 showed strongly decreased PHA accumulation from gluconate but no difference in phaC1 (encoding the PHA synthase) transcription, indicating that alginate biosynthesis competes with PHA biosynthesis regarding acetyl-CoA as precursor for both biopolymers. Transcription of phaF and phaI-F was nitrogen independent. PMID- 15268932 TI - Genome diversity among regional populations of Francisella tularensis subspecies tularensis and Francisella tularensis subspecies holarctica isolated from the US. AB - Francisella tularensis is a highly infectious facultative intracellular pathogen that is considered a potential agent of bioterrorism. Four different F. tularensis subspecies have been identified and they appear to display different ecological and virulence characteristics as well as differences in geographical distribution. One simple explanation for the variation in ecological and virulence characteristics is that they are conferred by differences in genome content. To characterize genome content among stains isolated from United States, we have used a DNA microarray designed from a shotgun library of a reference strain. Polymorphisms distributed among polyphyletic sets of strains was the most common pattern of genome alteration observed, indicating that strain-specific genome variability is significant. Nonetheless, 13 different contiguous segments of the genome were found to be missing exclusively in each of the subsp. holarctica strains tested. All 13 are associated with repeat sequences or transposases that could promote insertion/deletion events. Comparison of the live vaccine strain to other holarctica strains also identified three regions that are absent exclusively in the live vaccine strain derived from holarctica. PMID- 15268933 TI - Detailed analysis of the insertion site of the mobile elements R997, pMERPH, R392, R705 and R391 in E. coli K12. AB - The IncJ group of mobile elements have not been extensively studied until recently, due to the inability to isolate extrachromosomal DNA from IncJ-strains. Sequence analysis of the prototype IncJ element, R391, revealed it to be a mosaic structure, integrated into the prfC gene in E. coli. Using inverse PCR (iPCR), we localised the other available IncJ elements (R392, R705, R997 and pMERPH) site of insertion to a 17-bp sequence, within the 5' end of prfC at 99.31 min on the E. coli chromosome, and confirmed this for R391. Despite disrupting prfC, the IncJ's encode novel promoter and 5' sequences, restoring function of the disrupted prfC. Sequence analysis of the elements ends revealed that they contain integrase genes, which share extensive homologies among the group, despite being isolated from broad geographic locations. The elements excise from the host chromosome by recombination between their attL and attR sites, with subsequent recombination between the attP sites on the circular forms and the attB sites in the host genomes. The attB site is highly conserved and found in many different bacteria, suggesting a possible broad host range. PMID- 15268934 TI - Mobile DNA elements in the gas vesicle gene cluster of the planktonic cyanobacteria Microcystis aeruginosa. AB - Insertion sequences (IS) have been characterized in Microcystis aeruginosa gas vesicle-deficient mutants. ISMae4, a homolog of the cyanobacterial IS702, belongs to the IS5 family, subgroup ISL2. ISMae2 and ISMae3 display typical IS features and express a transposase of the IS4 and IS1 family, respectively. ISMae1 exhibits a more complex genetic structure and harbours a degenerated transposase gene distantly related to IS1 elements. Hybridizations with IS-specific DNA probes suggest that transposition of ISMae2 and ISMae3 occurred by a replicative type mechanism. To our knowledge this is the first report showing that IS1 elements can be mobile in cyanobacteria. PMID- 15268935 TI - Isolation and characterization of the Rhodococcus opacus thiostrepton-inducible genes tipAL and tipAS: application for recombinant protein expression in Rhodococcus. AB - We cloned the Rhodococcus opacus (strain DSM 44193) tipA gene, which encodes two translation products, TipAL and TipAS. The gene products are homologous to the Streptomyces spp. TipAL and TipAS proteins, respectively. The tipA promoter is highly active and TipAS protein is predominantly accumulated in R. opacus cells when the inducer of transcription, thiostrepton, was presented in culture medium. We found that thiostrepton is also induced the expression of an endogenous TipA family protein in Rhodococcus erythropolis (strain JCM3201). The minimal tipA promoter region was defined (57 bp) and the conserved nucleotide sequence of the putative TipAL protein binding site (TipA-box) was identified in that region. The tipA gene is presumed to be transcribed into a leaderless mRNA. We applied the tipA promoter successfully for recombinant protein expression in R. erythropolis cells. PMID- 15268936 TI - Phenazine-1-carboxylic acid is negatively regulated and pyoluteorin positively regulated by gacA in Pseudomonas sp. M18. AB - The biosynthesis of antimicrobial metabolites is controlled by the GacS/GacA two component regulatory system in Pseudomonas species. The production of phenazine-1 carboxylic acid and pyoluteorin is differentially regulated by GacA in Pseudomonas sp. M18. Pyoluteorin was reduced to nondetectable level in culture of the gacA insertional mutant strain M18G grown in King's medium B broth, whereas phenazine-1-carboxylic acid production was increased 30-fold over that of the wild-type strain. Production of both antibiotics was restored to wild-type levels after complementation in trans with the wild-type gacA gene. Expression of the translational fusions phzA'-'lacZ and pltA'-'lacZ confirmed the effect of GacA on both biosynthetic operons. PMID- 15268937 TI - Fluorescence measurements of free [Mg2+] by use of mag-fura 2 in Salmonella enterica. AB - The Mg2+ fluorescent dye mag-fura 2, entrapped in cells or organelles, has frequently been used for dual excitation ratio-metric determinations of free ionic Mg2+ concentrations in eukaryotic, mostly mammalian cells. Here we report its successful application to measure free Mg2+ concentrations ([Mg2+]i) in Salmonella enterica cells. When kept in nominally Mg2+ free buffer (resting conditions), the [Mg2+]i of wild-type cells has been determined to be 0.9 mM. An increase in the external Mg2+ concentration ([Mg2+]e) resulted in a rapid increase of [Mg2+]i, saturating within a few seconds at about 1.5 mM with [Mg2+]e of 20 mM. In contrast, cells lacking the Mg2+ transport proteins CorA, MgtA, MgtB failed to show this rapid increase. Instead, their [Mg2+]i increased steadily over extended periods of time and saturated at concentrations below those of wild type cells. Mg2+ uptake rates increased more than 15-fold when corA was overexpressed in these mutant cells. Uptake of Mg2+ into corA expressing cells was strongly stimulated by nigericin, which increased the membrane potential DeltaPsi at the expense of DeltapH, and drastically reduced by valinomycin, which decreased the membrane potential DeltaPsi. These results reveal mag-fura 2 as a useful indicator to measure steady-state [Mg2+]i values in resting bacterial cells and to determine Mg2+ uptake rates. They confirm the role of CorA as the major Mg2+ transport protein and reveal the membrane potential as driving force for Mg2+ uptake into S. enterica cells. PMID- 15268938 TI - Simple and rapid PCR method for identification of streptococcal species relevant to animal infections based on 23S rDNA sequence. AB - A PCR identification system targeting 23S rDNA sequences for the identification of eight streptococcal species relevant to animal infections (Streptococcus agalactiae, S. bovis, S. canis, S. dysgalactiae, S. equi, S. porcinus, S. suis and S. uberis) was developed. This system consists of two PCR reactions, A and B, in which seven and eight primers, respectively, are used simultaneously, and was designed so that each amplification product indicates a species by its size. A total of 111 cultures, including the type strain of eight species, could be successfully identified and differentiated as individual species, except for the cross reactivity between S. bovis and S. equinus. The developed PCR system can complete the identification procedure for eight streptococcal species through two tube reactions per isolate, and, therefore, might provide a rapid, simple and accurate diagnostic tool for veterinary laboratories. PMID- 15268939 TI - Metabolism of octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine by Clostridium bifermentans strain HAW-1 and several other H2-producing fermentative anaerobic bacteria. AB - Several H2-producing fermentative anaerobic bacteria including Clostridium, Klebsiella and Fusobacteria degraded octahydro-1,3,5,7-tetranitro-1,3,5,7 tetrazocine (HMX) (36 microM) to formaldehyde (HCHO) and nitrous oxide (N2O) with rates ranging from 5 to 190 nmol h(-1)g [dry weight] of cells(-1). Among these strains, C. bifermentans strain HAW-1 grew and transformed HMX rapidly with the detection of the two key intermediates the mononitroso product and methylenedinitramine. Its cellular extract alone did not seem to degrade HMX appreciably, but degraded much faster in the presence of H2, NADH or NADPH. The disappearance of HMX was concurrent with the release of nitrite without the formation of the nitroso derivative(s). Results suggest that two types of enzymes were involved in HMX metabolism: one for denitration and the second for reduction to the nitroso derivative(s). PMID- 15268940 TI - The LIV-I/LS system as a determinant of azaserine sensitivity of Escherichia coli K-12. AB - The growth of Escherichia coli is inhibited by an antibiotic compound, azaserine (O-diazoacetyl-L-serine). Previous studies revealed the biochemical properties of azaserine, which involves inhibition of various enzymatic reactions as well as introduction of DNA breakage. However, genetically, nothing has been elucidated except that all the azaserine-resistant strains isolated so far carry lesions in the aroP gene as a primary determinant. Here, we demonstrate that, in addition to AroP, the LIV-I/LS system, an ATP-binding cassette type transporter, is involved in azaserine sensitivity of E. coli, by genetic analysis and transport studies, in which Ki value for azaserine was determined to be approximately 10(-3) M. PMID- 15268941 TI - Extensive set of mitochondrial LSU rDNA-based oligonucleotide probes for the detection of common airborne fungi. AB - Fungi exist in every indoor and outdoor environment. Many fungi are toxigenic or pathogens that may cause various public health concerns. Rapid and accurate detection and identification of fungi require specific markers. In this study, partial mitochondrial large subunit rDNA was amplified and sequenced from 32 fungal strains representing 31 species from 14 genera. Based on the sequence variation pattern, 26 oligonucleotide probes were designed for their discrimination. The specificity of the probes was evaluated through homology search against GenBank database and hybridization examination on 38 fungal strains. The 26 probes were verified as highly specific to 20 fungal species. A two-step detection procedure through PCR followed by probe hybridization gave ten fold increase in detection sensitivity than single-step PCR assay and would be a practical approach for environmental sample screening. The probes developed in this study can be applied in clinical diagnosis and environmental monitoring of fungal agents. PMID- 15268942 TI - Multigene phylogenies of Ophiostoma clavigerum and closely related species from bark beetle-attacked Pinus in North America. AB - Leptographium pyrinum, Leptographium terebrantis, Ophiostoma aureum, Ophiostoma clavigerum, and Ophiostoma robustum are very similar in morphology, host trees choice, and the way they are disseminated by bark beetles. Their phylogenetic relationships were clarified using rDNA and protein coding genes including actin, beta-tubulin, and translation elongation factor-1alpha. Protein coding gene trees showed better resolution than the rDNA tree, which generated three clades: O. clavigerum, L. terebrantis/L. pyrinum, and O. robustum/O. aureum. A combined gene phylogenetic tree, which was supported by high bootstrap values, showed that O. aureum, L. pyrinum, O. robustum, and O. clavigerum each formed distinct clades while L. terebrantis was paraphyletic to O. clavigerum. The higher variability of the protein coding genes and the congruity in their phylogenetic results suggested that these genes may be better markers for identifying closely related species. These gene trees have also facilitated the description of the evolutionary relationships among these species. PMID- 15268943 TI - In vivo induced antigenic determinants of Actinobacillus actinomycetemcomitans. AB - Actinobacillus actinomycetemcomitans is a Gram-negative capnophilic rod and the etiological agent of localized aggressive periodontitis. The genome-wide survey of A. actinomycetemcomitans using in vivo induced antigen technology (IVIAT) has previously resulted in the discovery of antigenic determinants expressed specifically in diseased patients. The present study evaluated the potential of these antigens as putative disease markers, and investigating their contribution to the pathogenesis of the microorganism. Sera from patients had a significantly greater antibody titer than sera from healthy controls against six antigens, which supports the in vivo expression of these antigens, and suggests their usefulness as disease markers. A. actinomycetemcomitans invasion of epithelium derived HeLa cells resulted in the induction of all three genes tested, as evidenced by real-time PCR. Isogenic mutants of these three genes were constructed and the adhesion and intracellular survival of the mutants was assayed in a competition assay with the wild-type strain. A significant defect in the intracellular survival of two of these mutant strains (orf1402 and orf859) was found. This defect could not be attributed to an adhesion defect. In contrast, a mutation in vapA, a homologue of a novel putative transcriptional regulator, out-competed the wild-type strain in the same assay. The virulent phenotype was restored for a mutant strain in orf859 upon complementation. This data provided new insight into the pathogenic personality of A. actinomycetemcomitans in vivo and supported the use of HeLa cells as a valid in vitro host-pathogen interactions model for that microorganism. IVIAT is applicable to most pathogens and will undoubtedly lead to the discovery of novel therapies, antibiotics and diagnostic tools. PMID- 15268944 TI - The rapid identification of European Armillaria species from soil samples by nested PCR. AB - New specific primers AR1 and AR2 were successfully used for the amplification of a specific part of internal transcribed spacer (ITS) of rDNA of Armillaria isolated from soil samples. DNA was isolated from 0.5 g of forest soil and ITS region was amplified by nested PCR reaction with external primers ITS1 and ITS4 and internal primers AR1 and AR2. The individual species were distinguished by restriction fragment length polymorphisms (RFLPs) analysis with restriction endonuclease HinfI. The fragments were analysed by ion-exchange HPLC that is more sensible and more rapid than electrophoresis. The amplicons were sequenced to improve the discrimination between the species. The method enables the identification of Armillaria species within one day directly from soil samples without the need for previous isolation and cultivation of mycelium of Armillaria. PMID- 15268945 TI - Mutational analyses of the thermostable NAD+-dependent DNA ligase from Thermus filiformis. AB - The crystal structure of NAD+-dependent DNA ligase from Thermus filiformis (Tfi) revealed that the protein comprised four structural domains. In order to investigate the biochemical activities of these domains, seven deletion mutants were constructed from the Tfi DNA ligase. The mutants Tfi-M1 (residues 1-581), Tfi-M2 (residues 1-448), Tfi-M3 (residues 1-403) and Tfi-M4 (residues 1-314) showed the same adenylation activity as that of wild-type. This result indicates that only the adenylation domain (domain 1) is essential for the formation of enzyme-AMP complex. It was found that the zinc finger and helix-hairpin-helix (HhH) motif domain (domain 3) and the oligomer binding (OB)-fold domain (domain 2) are important for the formation of enzyme-DNA complex. The mutant Tfi-M1 alone showed the activities for in vitro nick-closing and in vivo complementation in Escherichia coli as those of wild-type. These results indicate that the BRCT domain (domain 4) of Tfi DNA ligase is not essential for the enzyme activity. The enzymatic properties of Tfi-M1 mutant (deleted the BRCT domain) were slightly different from those of wild-type and the nick-closing activity of Tfi-M1 mutant was approximately 50% compared with that of wild-type. PMID- 15268946 TI - A molecular beacon-based real-time NASBA assay for detection of Mycobacterium avium subsp. paratuberculosis in water and milk. AB - A molecular beacon-based real-time NASBA assay for detection and identification of Mycobacterium avium subsp. paratuberculosis has been developed. It targets and amplifies sequences from the dnaA gene which are specific for this bacterium. The assay includes an internal amplification control, to allow identification of inhibited reactions. The assay was tested against 18 isolates of M. avium subsp. paratuberculosis, 17 other mycobacterial strains and 25 non-mycobacterial strains, and was fully selective in that it detected all the targets but none of the non-targets. The lowest number of cells which the assay can detect with 99% probability is 150-200 cells per reaction (as determined using pure culture suspensions). Using centrifugation and nucleic acid extraction as sample treatment, the assay was able to consistently detect 10(3) M. avium subsp. paratuberculosis cells in 20 ml artificially contaminated drinking water. With a simple detergent and enzymatic sample pretreatment before centrifugation and nucleic acid extraction, the assay was able to consistently detect 10(4) M. avium subsp. paratuberculosis cells in 20 ml artificially contaminated semi-skimmed milk. The assay will be a useful addition to the range of diagnostic tools available for the study of M. avium subsp. paratuberculosis. PMID- 15268947 TI - Purification and characterization of a novel metalloprotease isolated from Aeromonas caviae. AB - A novel protease produced by Aeromonas caviae was purified and characterized. The molecular weight of the protease (AP19) was estimated as 19 kDa on SDS polyacrylamide gel electrophoresis. The protease activity was not inhibited completely by heating at 100 degrees C for 15 min. The proteolytic activities were inhibited by metalloprotease inhibitor. The N-terminal amino acid sequence of AP19 was VTASVSFSGRCTN. AP19 did not activate Aeromonas proaerolysin, and did not show fluid accumulation in the rabbit intestinal loop test. A high concentration of the protease showed cytotoxic activity against Vero cells. PMID- 15268948 TI - Monitoring of the bacterial composition of dairy starter cultures by RAPD-PCR. AB - Randomly amplified polymorphic DNA (RAPD)-PCR was used to verify the species composition of commercial dairy starters and to detect possible shifts in strain composition of these cultures. After RAPD-PCR analysis, not all the strains isolated in the years 2001 and 2002 fell within the same dendrogram cluster of the strains isolated in the year 2000 and used as reference strains. Changes in composition of the microbial population and/or voluntary immission of new biotypes with respect to the original strain formulation had occurred in the starters. The microbial composition of modern dairy starters represents a key point because the complex relationships among microorganisms can easily be altered. Little variations in the microbial composition could have unexpected effects on cheese quality. PMID- 15268949 TI - Effect of uncouplers on endogenous respiration and ferrous iron oxidation in a chemolithoautotrophic bacterium Acidithiobacillus (Thiobacillus) ferrooxidans. AB - Oxidation of ferrous iron (Fe2+) to ferric iron (Fe3+) with oxygen (O2) by Acidithiobacillus (Thiobacillus) ferrooxidans is considered to be inhibited by uncouplers. Oxidation of the endogenous substrates (presumably NADH) with O2 or Fe3+, on the other hand, was stimulated by uncouplers, 2,4-dinitrophenol (DNP) and carbonylcyanide-m-chlorophenyl-hydrazone (CCCP), as expected in respiratorily controlled mitochondria or heterotrophic bacteria. Amytal and rotenone were inhibitory. Fe3+ reduction by endogenous substrates was studied extensively and was found to be stimulated by a permeable anion, SCN- and weak acids, as well as the above uncouplers. Proton translocating properties of some of these stimulators were shown by following a pH change in the cell suspension. It was concluded that any compounds that destroy proton electrochemical gradient, Deltap, stimulated endogenous respiration. Stimulation of Fe2+ or ascorbate oxidation by lower concentrations of uncouplers was successfully demonstrated by shortening the reaction time, but only to a small extent. Uncouplers at concentrations stimulatory to endogenous respiration inhibited Fe2+ oxidation if present before Fe2+ addition. The inhibition by 10 microM CCCP was reversed by washing the cells in a buffer. Complex I inhibitors, atabrine, rotenone and amytal inhibited Fe2+ oxidation, more strongly in the presence of 0.1 mM DNP. It is proposed that Fe2+ oxidation required Deltap perhaps to climb an energetically uphill reaction or to reduce NAD+ to NADH by reversed electron flow for CO2 fixation. The latter interpretation implies some obligatory coupling between Fe2+ oxidation and NAD+ reduction. PMID- 15268950 TI - Occurrence of tetracycline resistance genes tet(M) and tet(S) in bacteria from marine aquaculture sites. AB - Occurrence of tetracycline resistance genes encoding ribosomal protection proteins was examined in 151 tetracycline-resistant bacterial isolates from fish and seawater at coastal aquaculture sites in Japan and Korea. The tet(M) gene was detected in 34 Japanese and Korean isolates, which included Vibrio sp., Lactococcus garvieae, Photobacterium damsela subsp. piscicida, and unidentified Gram-positive bacteria. The majority of these bacterial isolates displayed high level resistance with a minimum inhibitory concentrations (MICs) equal to or greater than 250 microg/ml of oxytetracycline and only four isolates had MICs less than 31.3 microg/ml. 16S rDNA RFLP typing of tet(M)-positive Vibrio isolates suggests that these are clonal populations of the same phylotype specific to a particular location. One Vibrio clone (phylotype III), however, is widely disseminated, being detected during different sampling years, at different locations, and in different fish species in both Japan and Korea. The tet(S) gene was detected in L. garvieae from yellowtail in Japan and in Vibrio sp. from seawater in Korea. This is the first report of tet(S) occurrence in Gram-negative facultative anaerobes. These results suggest that tet(M) and tet(S) genes are present in fish intestinal and seawater bacteria at aquaculture sites and could be an important reservoir of tetracycline resistance genes in the marine environment. PMID- 15268951 TI - Cloning of an agr homologue of Staphylococcus saprophyticus. AB - An agr homologue of Staphylococcus saprophyticus was identified, cloned and sequenced. The gene locus shows homologies to other staphylococcal agr systems, especially to those of S. epidermidis and S. lugdunensis. A putative RNAIII was identified and found to be differentially expressed during the growth phases. In contrast to the RNAIII molecules of S. epidermidis and S. aureus it does not contain an open reading frame that codes for a protein with homologies to the delta-toxin. Using PCR, the agr was found to be present in clinical isolates of S. saprophyticus. PMID- 15268952 TI - Brefeldin A enhances Helicobacter pylori vacuolating cytotoxin-induced vacuolation of epithelial cells. AB - Intracellular VacA localises to the vacuolar (late endosome/lysosome) membrane, but little is known about the trafficking of the toxin beyond this region. We show that the Golgi-disturbing agent brefeldin A (BFA) enhances VacA-induced vacuolation of epithelial cells by Helicobacter pylori co-culture and, importantly, BFA treatment induces vacuolation by less toxic forms of VacA. The effect is BFA dose-dependent and occurs within 2.5 h. These data suggest that VacA may be routed deeper within the cell than the vacuole, and that vacuolation is minimised when this occurs efficiently. This may explain why some forms of VacA do not cause vacuolation and why vacuolation is minimal at the low bacteria:cell ratios observed in vivo. PMID- 15268953 TI - Emission inventory and sources of polycyclic aromatic hydrocarbons in the atmosphere at a suburban area in Taiwan. AB - The application of a chemical mass balance air pollution model to ambient measurements of polycyclic aromatic hydrocarbons (PAHs) is presented. Sixteen air samples were collected at seven sites in a suburban area in Taiwan and analyzed for the concentration of 21 compounds between July 2001 and September 2001. Each ambient sample was evaluated for the PAH contribution from six sources (heavy oil combustion, natural gas combustion, coal combustion, diesel combustion, vehicles and municipal solid waste incinerator). Average predictions agree well with the emission inventory. By this method, the average contributions are 49%, 14%, 22%, 12%, and 2% from vehicles, heavy oil combustion, natural gas combustion, coal combustion and diesel combustion at these seven receptors. By far, vehicles are the major PAH emission sources and municipal solid waste incinerator is a minor contributor. The calculated result of particulate PAHs is compared with that of total (gaseous and particulate) PAHs. The estimate based on total PAHs is better than the estimate based on particulate PAHs only. Contributions of eight low reactive PAHs for the same emission sources and receptors were calculated. Atmospheric reactivity seems not a problem for source apportionment in this study. PMID- 15268954 TI - Treatment of refectory oily wastewater by electro-coagulation process. AB - Electro-coagulation was used to treat refectory wastewater with high oil and grease contents. Different operational conditions were examined, including pH, current density, reaction time, conductivity, electrode distance and inlet concentration. The optimum current density was 10-14 A m(-2) within 30 min depending on the wastewater properties tested. Conductivity had little effect on the treatment efficiency. Although the addition of extra salts (e.g., sodium chloride) to the wastewater did not help increase the pollutant removal efficiency, it could save the power consumption significantly. The COD(Cr) and oil removal efficiency descended with increasing electrode distance. The optimal electrode distance was determined to be 10 mm for this equipment in consideration of the treatment cost and efficiency together. The pH effect on the performance of the electro-coagulation process was not very significant in the range of 3-10. The removal efficiency of oil and COD(Cr) under normal condition exceeded 95% and 75%, respectively. PMID- 15268955 TI - Critical effect of hydrogen peroxide concentration in photochemical oxidative degradation of C.I. Acid Red 27 (AR27). AB - The photochemical decolorization of C.I. Acid Red 27 (AR27), an anionic monoazo dye, was studied in the UV/H2O2 process by using a batch reactor with a UV-C lamp emitting at 254 nm (30 W). The decolorization rate follows pseudo-first order kinetic with respect to the AR27 concentration. The results indicate that the apparent reaction rate constant in the UV/H2O2 process is a function of H2O2 concentration. In this work, a mathematical relation between the apparent reaction rate constant of the AR27 removal and used H2O2 was established. The applied amount of H2O2 was performed in two forms: (i) the light fraction absorbed by H2O2 in 254 nm, (ii) the initial concentration ratio of H2O2 to AR27. The results obtained from this mathematical model are in good agreement with experimental results. PMID- 15268956 TI - Engineering and environmental properties of thermally treated mixtures containing MSWI fly ash and low-cost additives. AB - An experimental work was carried out to investigate the feasibility of application of a sintering process to mixtures composed of Municipal Solid Waste Incinerator (MSWI) fly ash and low-cost additives (waste from feldspar production and cullet). The proportions of the three constituents were varied to adjust the mixture compositions to within the optimal range for sintering. The material was compacted in cylindrical specimens and treated at 1100 and 1150 degrees C for 30 and 60 min. Engineering and environmental characteristics including weight loss, dimensional changes, density, open porosity, mechanical strength, chemical stability and leaching behavior were determined for the treated material, allowing the relationship between the degree of sintering and both mixture composition and treatment conditions to be singled out. Mineralogical analyses detected the presence of neo-formation minerals from the pyroxene group. Estimation of the extent of metal loss from the samples indicated that the potential for volatilization of species of Pb, Cd and Zn is still a matter of major concern when dealing with thermal treatment of incinerator ash. PMID- 15268957 TI - Relationship between pressure fluctuations and generation of organic pollutants with different particle size distributions in a fluidized bed incinerator. AB - The hydrodynamic behaviors of fluidization perhaps significantly influence the uniformity of fluidization in fluidized bed incinerator. Good uniformity of fluidization expressed the air across uniformly through the bed and the particles being distributed well in the fluid stream. The aggregates, flocs and channels of particles do not happen during fluidization. The Good uniformity will maintain high heat and mass distribution to improve reaction efficiency. These parameters include the height of static bed, gas velocity, mixing and distribution of bed particle, which have rarely been studied in previous investigations. Consequently, this study examines how the hydrodynamic parameters affect the generation of organic pollutants (BTEXs and PAHs) during incineration. The statistical and power spectral analysis of the measured pressure fluctuation during incineration are used to elucidate the relationship between behaviors of fluidization and generation of pollutants during incineration. Experimental results show the organic concentration does not increase with uniformity of fluidization decreasing. The reason may be the explosion of the gas and the consequent thermal shock destroy the coalescent bubbles to form small bubbles again and enhance the efficiency of transfer of oxygen to increase combustion efficiency. Additionally, the mean amplitude and fluidized index of pressure fluctuation similarly vary with the concentration of organic pollutants. These two indices can be used to assess the efficiency of combustion. The four particle size distributions could be divided into two groups by statistical analysis. The Gaussian and narrow distributions belong to one group and the binary and flat the other. The organic concentration of the Gaussian and narrow distributions are lower than that of the other distributions. Consequently, the bed materials should maintain narrow or Gaussian distributions to maintain a good combustion efficiency during incineration. PMID- 15268958 TI - Temperature dependence of hydroxyl radical formation in the hv/Fe3+/H2O2 and Fe3+/H2O2 systems. AB - The thermal enhancement of the formation of *OH by the hv/Fe(III)/H2O2 system (including the Fe(III)/H2O2 system) was quantitatively investigated with reaction temperatures ranging from 25 to 50 degrees C. A temperature dependent kinetic model for the hv/Fe(III)/H2O2 system, incorporating 12 major reactions with no fitted rate constants or activation energies, was developed, and successfully explained the experimental measurements. Particularly, the thermal enhancement of Fe(OH)2+ photolysis which is the most significant step in the hv/Fe(III)/H2O2 system was effectively explained by two factors; (1) the variation of the Fe(OH)2+ concentration with temperature, and (2) the temperature dependence of the quantum yield for Fe(OH)2+ photolysis (measured activation energy=11.4 kJ mol(-1)). Although in both the hv/Fe(III)/H2O2 and Fe(III)/H2O2 systems, elevated temperatures enhanced the formation of *OH, the thermal enhancement was much higher in the dark Fe(III)/H2O2 system than the hv/Fe(III)/H2O2 system. Furthermore, it was found that the relative thermal enhancement of the formation of *OH in the presence of *OH scavengers (tert-butyl alcohol) was magnified in the Fe(III)/H2O2 system but was not in the hv/Fe(III)/H2O2 system. PMID- 15268960 TI - Pressurised liquid-liquid extraction. An approach to the removal of inorganic non metal species from used industrial oils. AB - Modified pressurised hot water is used for the development of a high pressure liquid-liquid extraction method for the decontamination of used industrial oils from inorganic non-metal species (chlorine, fluorine and sulphur). The oils were subjected to dynamic extraction with water modified with 5% v/v HNO3 at 200 degrees C as extractant. Under these working conditions the analytes were transferred to the aqueous phase. Spontaneous separation of the two immiscible liquid phases (the used oil and extract) takes place in the collection flask after extraction. The treated and untreated oil samples were oxidised and the chloride, fluoride and sulphate thus formed were determined by ion chromatography. The method was applied to four oil samples from different locations in Spain. A residence time of approximately =10 min provided oil samples from which 88.3%, 89.4% and 89.4% of chloride, fluoride and sulphate, respectively, have been removed with respect to the initial concentration of each analyte in the oil. The repeatability, expressed as relative standard deviation (RSD), was of 11.9%, 13.7% and 7.2% for Cl(-), F(-) and SO4(2-), respectively; whilst the within-laboratory reproducibility yielded RSDs of 6.2%, 7.9% and 6.2% for the same analytes. The proposed approach has proved to be efficient, simple, easily transferable to industrial scale, cheap, fast and environmentally friendly. PMID- 15268959 TI - Dechlorination ability of municipal waste incineration fly ash for polychlorinated phenols. AB - Pathways of pentachlorophenol dechlorination have been investigated on municipal waste incineration fly ash at 200 degrees C under nitrogen atmosphere. Thermodynamic calculations have been carried out for these dechlorination conditions using the method of total Gibbs energy minimization for the whole system consisting of gaseous components, i.e., chlorinated phenols, phenol, hydrogen chloride and the Cu3Cl3 trimer and of solid Cu2O and CuCl2 components. The effects of water, temperature and of the amounts of the reaction components on the thermodynamic equilibrium have been discussed and the experimental results compared with the calculated thermodynamic data. PMID- 15268961 TI - The influence of pH on the degradation of phenol and chlorophenols by potassium ferrate. AB - This paper presents information concerning the influence of solution pH on the aqueous reaction between potassium ferrate and phenol and three chlorinated phenols: 4-chlorophenol (CP), 2,4-dichlorophenol (DCP), 2,4,6-trichlorophenol (TCP). The redox potential and aqueous stability of the ferrate ion, and the reactivity of dissociating compounds, are known to be pH dependent. Laboratory tests have been undertaken over a wide range of pH (5.8-11) and reactant concentrations (ferrate:compound molar ratios of 1:1 to 8:1). The reactivity of trichloroethylene was also investigated as a reference compound owing to its non dissociating nature. The extent of compound degradation by ferrate was found to be highly pH dependent, and the optimal pH (maximum degradation) decreased in the order: phenol/CP, DCP, TCP; at the optimal pH the degree of degradation of these compounds was similar. The results indicate that for the group of phenol and chlorophenols studied, the presence of an increasing number of chlorine substituent atoms corresponds to an increasing reactivity of the undissociated compound, and a decreasing reactivity of the dissociated compound. PMID- 15268962 TI - Implications of rhizospheric heavy metals and nutrients for the growth of alfalfa in sludge amended soil. AB - Rhizospheric distribution of nutrients and heavy metals in sludge amended soil was investigated using the rhizobag technique to give an indication of the release of metals from wastewater sludge. DTPA-extractable Zn, Cd, Ni and Mn, and available P, K and NH4+-N in the rhizosphere were markedly depleted when soil was amended with sludge. There was no conspicuous depletion or accumulation of DTPA extractable Cu in the rhizosphere when the soil was amended with sewage sludge but DTPA-extractable Fe accumulated in the rhizosphere when the soil was amended with increasing amounts of sludge. The pH value in the rhizosphere increased with distance from the roots when soil was amended with larger amounts of sludge. The exchangeable fraction of Cu in the rhizosphere was depleted whether or not the soil was treated with sludge. Carbonate, oxide, organic and residual fractions of Cu and Zn were depleted in the rhizosphere at a distance of 0-2 mm from the roots when soil was amended with 50% sludge. Application of sewage sludge had a positive effect on alfalfa growth. With an increase in sludge amounts, the concentrations of Fe, Cu and Zn in alfalfa shoots did not change. Soil amendments with less than 25% sludge did not increase the availability or mobility of heavy metals. The depletion in rhizospheric DTPA-extractable Zn, Cd and Ni indicates that with the sole exception of Cu, release of metals from sludge amended soil was very limited. PMID- 15268963 TI - Effects of humic substances on photodegradation of bensulfuron-methyl on dry soil surfaces. AB - The photodegradation of the herbicide bensulfuron-methyl on dry soil surfaces in the presence and absence of humic substances was investigated under Xe lamp irradiation. A rapid rate of disappearance occurs in the humus-removed soil. The presence of humic acid (HA) and fulvic acid (FA) reduces degradation rates and has a quenching effect on the photodecomposition of bensulfuron-methyl. The quenching effect increases with increasing HA and FA concentration in soil. HA has a slightly greater ability to quenching the photolysis compared to FA. In addition, co-effects of HA and FA on quenching the photolysis are stronger than single effects of HA or FA. PMID- 15268964 TI - Feed-forward dose control of wastewater chlorination using on-line pH and ORP titration. AB - In this study, a simple automatic pH-ORP titration device was developed for identifying the various ammonia concentrations and chlorine dose requirements for wastewater chlorination by identifying the peaks in the ORP-slope profiles and knees/valleys in the pH profiles. In addition, breakpoint chlorination experiments have shown that the ORP values at the monochloramine humps and breakpoints are linearly correlated with the ammonia concentrations. Therefore, a feed-forward control strategy, based on the chlorine/ammonia weight ratio (Cl/N), is proposed in this paper, to control the chlorine doses for a continuous wastewater chlorination experiment in a laboratory-scale reactor. The result of this continuous wastewater chlorination experiment has shown that the pH-ORP titration was able to precisely determine the variations of ammonia concentrations in the chlorination influent. Under this control strategy, effective and stable disinfection efficiencies in terms of total coliform count were obtained. PMID- 15268965 TI - Sonochemical decomposition of natural polyphenolic compound (condensed tannin). AB - Sonochemical treatment of grape condensed tannin was studied with the aim to destroy phenolic constituents, using ultrasonic frequency of 20 kHz in the presence and in the absence of hydrogen peroxide (H2O2), as an oxidative agent. Various pH, H2O2 concentration and temperature combinations were used in order to investigate the role of these parameters in the studied process. In order to estimate the effect of ultrasound, similar experiments were conducted in terms of pH, H2O2 concentration and temperature conditions, but without sonication. In all cases the presence of H2O2 was resulted in considerably higher removal of total phenolics (TP). It was observed that pH and temperature present also a significant effect on TP removal in the presence of H2O2, both with and without sonication. Under similar experimental conditions higher reaction rates, concerning TP removal, were obtained in the presence of ultrasonic irradiation, than in its absence. PMID- 15268966 TI - Degradation of 2-chlorophenol via a hydrogenotrophic biofilm under different reductive conditions. AB - This research studies the 2-chlorophenol (2-CP) degradation by the hydrogenotrophic biofilm cultivated in three silicone-tube membrane bioreactors under the conditions of denitrification (DN), sulfate-reduction (DS) and dechlorination (DC). Experimental results showed that after acclimation for more than four months with 2-CP, the respective 2-CP removal efficiency was 95% in DN, 94% in DS and 95% in DC reactors, under the condition of influent 2-CP 25 mg/l with hydraulic retention time (HRT) of 15 h. The metabolic pathway of 2-CP was different in different reactors. The 2-CP was thought to be utilized as carbon and energy source in DN and DS reactors, while the dechlorination occurred in the DC reactor in lack of nitrate and sulfate. The pH dramatically affected the 2-CP degradation in all reactors. Experimental results showed that the optimal pH range was around 6+/-0.2 in DN, 7+/-0.2 in DS, and 5.8-7.2 in DC reactors. Both nitrate and sulfate inhibited the 2-CP dechlorination, but the inhibition levels were different. Nitrate completely inhibited the dechlorination at once, while sulfate took a longer time to reach complete inhibition, only after the bacteria were adapted to the sulfate-reducing condition. Both inhibitions were accomplished by taking the place of 2-CP as electron acceptors. H2 served as an electron donor for dechlorination of 2-CP. The dechlorination was apparently stopped when lacking H2 and another pathway was responsible for the 2-CP degradation. PMID- 15268967 TI - Abiotic reductive dechlorination of chlorinated ethylenes by iron-bearing phyllosilicates. AB - Abiotic reductive dechlorination of chlorinated ethylenes (tetrachloroethylene (PCE), trichloroethylene (TCE), cis-dichloroethylene (c-DCE), and vinylchloride (VC)) by iron-bearing phyllosilicates (biotite, vermiculite, and montmorillonite) was characterized to obtain better understanding of the behavior of these contaminants in systems undergoing remediation by natural attenuation and redox manipulation. Batch experiments were conducted to evaluate dechlorination kinetics and some experiments were conducted with addition of Fe(II) to simulate impact of microbial iron reduction. A modified Langmuir-Hinshelwood kinetic model adequately described reductive dechlorination kinetics of target organics by the iron-bearing phyllosilicates. The rate constants stayed between 0.08 (+/-10.4%) and 0.401 (+/-8.1%) day(-1) and the specific initial reductive capacity of iron bearing phyllosilicates for chlorinated ethylenes stayed between 0.177 (+/-6.1%) and 1.06 (+/-7.1%) microM g(-1). The rate constants for the reductive dechlorination of TCE at reactive biotite surface increased as pH (5.5-8.5) and concentration of sorbed Fe(II) (0-0.15 mM g(-1)) increased. The appropriateness of the model is supported by the fact that the rate constants were independent of solid concentration (0.0085-0.17 g g(-1)) and initial TCE concentration (0.15 0.60 mM). Biotite had the greatest rate constant among the phyllosilicates both with and without Fe(II) addition. The rate constants were increased by a factor of 1.4-2.5 by Fe(II) addition. Between 1.8% and 36% of chlorinated ethylenes removed were partitioned to the phyllosilicates. Chloride was produced as a product of degradation and no chlorinated intermediates were observed throughout the experiment. PMID- 15268968 TI - Decolorizing of lignin wastewater using the photochemical UV/TiO2 process. AB - Studies on applying the photochemical UV/TiO2 oxidation process to treat the lignin-containing wastewater for dissolved organic carbon (DOC), color and reducing A254 (the absorption at the wavelength of 254 nm) have been carried out. The data obtained in this study demonstrate that the UV/TiO2 process is effective in oxidizing the lignin thus reducing the color and DOC of the wastewater treated. The combined UV/TiO2 treatment can achieve better removal of DOC and color than the UV treatment alone. Color removal, based on American Dye Manufacture Index (ADMI) measurement, is greater than 99% if the pH is maintained at 3.0 with the addition of 1 g l(-1) TiO2. When 10 g l(-1) TiO2 is applied, the oxidation reduction potential (ORP) value is reached to result in an 88% removal of both DOC and color. A model was developed based on the variation of ORP during the photochemical reaction to simulate the decoloring process. The proposed model can be used to predict the color removal efficiency of the UV/TiO2 process. PMID- 15268969 TI - Therapeutic effects of herbal extracts and constituents in animal models of psychiatric disorders. AB - A search for novel pharmacotherapy from medicinal plants for psychiatric illnesses has progressed significantly in the past decade. This is reflected in the large number of herbal preparations for which psychotherapeutic potential has been evaluated in a variety of animal models. The objective of this review is to provide an overview of herbal extracts and constituents that have significant therapeutic effects in animal models of psychiatric illnesses. Eighty five individual herbs reviewed were classified as anxiolytic, antidepressant, neuroleptic, antidementia, or anti-substance abuse herbs. The full scientific name of each herb, herbal part used, active constituent, extract, dose range and route, animal model, possible mechanisms of action, and pertinent references are presented via synoptic tables. The herbal mixtures were also mentioned. A considerable number of herbal constituents whose behavioral effects and pharmacological actions have been well characterized may be good candidates for further investigations that may ultimately result in clinical use. The investigation of a large portion of the herbal extracts and herbal mixtures is in its infancy. Herbal remedies that have demonstrable psychotherapeutic activities have provided a potential to psychiatric pharmaceuticals and deserve increased attention in future studies. PMID- 15268970 TI - Protective effects of Curcuma longa on ischemia-reperfusion induced myocardial injuries and their mechanisms. AB - The present study was undertaken to evaluate the cardioprotective potential of Curcuma longa (Turmeric) in the ischemia-reperfusion (I/R) model of myocardial infarction (MI). Wistar rats were divided into three groups and received saline orally (sham, control I/R group) and Curcuma longa 100 mg/kg (CL-100 treated group) respectively for one month. On the 31st day, rats of the control I/R and Cl treated groups were subjected to 45 min of occlusion of the LAD coronary artery and were thereafter reperfused for 1 h. I/R resulted in significant cardiac necrosis, depression in left ventricular function, decline in antioxidant status and elevation in lipid perodixation in the control I/R group as compared to sham control. Myocardial infarction produced after I/R was significantly reduced in the Cl treated group. Cl treatment resulted in restoration of the myocardial antioxidant status and altered hemodynamic parameters as compared to control I/R. Furthermore, I/R-induced lipid peroxidation was significantly inhibited by Cl treatment. The beneficial cardioprotective effects also translated into the functional recovery of the heart. Cardioprotective effect of Cl likely results from the suppression of oxidative stress and correlates with the improved ventricular function. Histopathological examination further confirmed the protective effects of Cl on the heart. PMID- 15268971 TI - Indole-3-acetic acid increases glutamine utilization by high peroxidase activity presenting leukocytes. AB - Indole-3-acetic acid (IAA) is toxic for human tumor cells and in association with horseradish peroxidase (HRP) can be used as a new prodrug/enzyme combination for targeted cancer therapy. The toxic effect of IAA on neutrophils, macrophages and lymphocytes is associated with cell peroxidase activity, which is high in neutrophils and low in lymphocytes. The effect of IAA on glucose and glutamine metabolism in leukocytes presenting different peroxidase activities: neutrophils, thioglycollate-elicited macrophages and lymphocytes was investigated. A time course effect (from 6 to 48 h in culture) of IAA on glucose and glutamine metabolism of neutrophils, thioglycollate-elicited macrophages, and lymphocytes was then carried out. Addition of IAA (0.25 mM) did not have a marked effect on glucose utilization and lactate formation by the three cell types but it raised glutamine consumption and glutamate production by neutrophils and macrophages. IAA had no effect on glutamine consumption and glutamate production by lymphocytes. A strong relationship was found between glutamine utilization (0.999) and glutamate production (0.999) and peroxidase activity. IAA did not change the activities of hexokinase, glucose-6-phosphate dehydrogenase, citrate synthase, lactate dehydrogenase, and phosphate-dependent glutaminase of 24 h cultured neutrophils and lymphocytes. The effect of IAA (1 mM) on glucose and glutamine metabolism was also investigated by 1 h incubated leukocytes in PBS. IAA did not affect glucose and glutamine metabolism of lymphocytes but enhanced glucose and glutamine metabolism by 1 h incubated neutrophils and thioglycollate elicited macrophages. IAA caused a marked increase on oxygen consumption by neutrophils, which was more pronounced in the presence of the glutamine as compared to glucose. The stimulation of oxygen consumption leads to a reduction in NADH/NAD+ ratio that activates the flux of substrates through the Krebs cycle. Since glutamine is mainly metabolized through the left hand side of the Krebs cycle, a reduction in the redox state of the cells may accelerate the flux of substrates through glutaminolysis. The toxic results presented here show that the affect of IAA in association with peroxidase involves activation of glutamine metabolism. PMID- 15268972 TI - Effects of Lactobacillus helveticus fermented milk on bone cells in vitro. AB - Milk fermented with Lactobacillus helveticus (L. helveticus) contains small peptides such as isoleucyl-prolyl-proline (IPP) and valyl-prolyl-proline (VPP), which inhibit the angiotensin converting enzyme (ACE). We investigated the effects of L. helveticus fermented milk whey (Lh-whey) and its components, sour milk whey, calcium and IPP and VPP peptides, on bone cells in vitro. An osteoblast assay was performed by determining the amount of deposited calcium as an index of bone formation in cultures of mouse osteoblasts formed from bone marrow-derived osteoblast precursor cells. An osteoclast assay was performed by determining the activity of tartrate-resistant acid phosphatase released into the culture medium in cultures of mouse osteoclasts formed from bone marrow-derived osteoclast precursor cells. The Lh-whey increased bone formation 1.3-1.4 times with the 1 x 10(-5), 1 x 10(-4) and 1 x 10(-3) solutions. The IPP and VPP peptides also demonstrated a significant 5-fold activation of bone formation in in vitro osteoblast cultures, whereas the sour milk whey and calcium had no effect. No significant effects were observed on osteoclasts in vitro with any of the study products. L. helveticus fermented milk whey contains bioactive components that increase osteoblastic bone formation in vitro. The effect may be due to the ACE-inhibitory IPP and VPP peptides, which showed a similar effect to that of the L. helveticus fermented milk whey. PMID- 15268973 TI - Salacia oblonga improves cardiac fibrosis and inhibits postprandial hyperglycemia in obese Zucker rats. AB - Diabetes has a markedly greater incidence of cardiovascular disease than the non diabetic population. The heart shows a slowly developing increase in fibrosis in diabetes. Extended cardiac fibrosis results in increased myocardial stiffness, causing ventricular dysfunction and, ultimately, heart failure. Reversal of fibrosis may improve organ function survival. Postprandial hyperglycemia plays an important role in the development of type 2 diabetes and cardiovascular complications, and has been proposed as an independent risk factor for cardiovascular diseases. Salacia oblonga (S.O.) is traditionally used in the prevention and treatment of diabetes. We investigated the effects of its water extract on cardiac fibrosis and hyperglycemia in a genetic model of type 2 diabetes, the obese Zucker rat (OZR). Chronic administration of the extract markedly improved interstitial and perivascular fibrosis in the hearts of the OZR. It also reduced plasma glucose levels in non-fasted OZR, whereas it had little effect in the fasted animals, suggesting inhibition of postprandial hyperglycemia in type 2 diabetic animals, which might play a role in improvement of the cardiac complications of OZR. Furthermore, S.O. markedly suppressed the overexpression of mRNAs encoding transforming growth factor betas 1 and 3 in the OZR heart, which may be an important part of the overall molecular mechanisms. S.O. dose-dependently inhibited the increase of plasma glucose in sucrose-, but not in glucose-loaded mice. S.O. demonstrated a strong inhibition of alpha glucosidase activity in vitro, which is suggested to contribute to the improvement of postprandial hyperglycemia. PMID- 15268974 TI - Effects of norepinephrine or prostaglandin E2 on extracellular acidification rate of MCG 101, or K1735-M2 tumor cells. AB - We studied by microphysiometry functional effects of two different signalling molecules in the murine tumor cell lines, MCG 101 and K1735-M2, namely norepinephrine (NE) and prostaglandin E2 (PGE2). This methodology implies estimation of intracellular metabolism by measurements of extracellular acidification rate (ECAR). MCG 101 (an undifferentiated, epithelial-like tumor), in contrast to K1735-M2 (a melanoma), has been found to produce great amounts of PGE2. Challenge of MCG 101 cells with PGE2 (0.284 and 2.84 microM for 9 min) elicited an increase in ECAR by about 10 and 41% above basal level, respectively. Pretreatment with indomethacin (0.5 microM) reduced the response to the two PGE2 concentrations by about 70 and 25%, respectively. In contrast, PGE2 caused virtually no response in K1735-M2 cells. Moreover, NE caused increases in ECAR in both cell types, possibly via beta3-adrenoceptors, as investigated pharmacologically in MCG 101, and by immunocytochemistry in both cell lines. The results obtained strongly suggest functional receptors for PGE2 in MCG 101, but not K1735-M2 tumor cells. Functional receptors for NE were demonstrated in both cell lines. There is possibly an autocrine loop in the MCG 101 cells, in which PGE2 activates cyclooxygenase. PMID- 15268975 TI - Effects of carbachol on rat Sertoli cell proliferation and muscarinic acetylcholine receptors regulation: an in vitro study. AB - The aim of the present work was to study the effect of muscarinic agonist on cell proliferation and muscarinic acetylcholine receptors (mAChRs) regulation in rat Sertoli cells. Primary cultures of Sertoli cells were obtained from 8-day and 15 day old male Wistar rats. In proliferation assays, [methyl-3H]thymidine incorporation in Sertoli cells from 8-day and 15-day old rats reached a plateau after 60 min of carbachol incubation and decreased after 120 min of agonist incubation. Binding studies with [N-Methyl-3H]scopolamine ([3H]NMS) indicated a rapid loss of cell surface mAChRs when Sertoli cells from 15-day old rats were incubated with carbachol at 35 degrees C for 2 min. This effect was temperature dependent. When the incubation of the cells was prolonged at 35 degrees C or at 4 degrees C, after the agonist had been washed away, 94% of mAChRs were present in the cell surface after 120 min incubation at 35 degrees C. At 4 degrees C, however, a low percentage of mAChRs was detected in the cell surface. In the presence of cycloheximide, the recycling of mAChRs to the cell surface was not changed, suggesting that the appearance of mAChRs on cell surface was not dependent on de novo receptor synthesis. In conclusion, our studies indicate that the activation of mAChRs may play a role in rat Sertoli cell proliferation. These receptors may be under regulation (internalization and recycling) when cells are exposed to muscarinic cholinergic agonist. PMID- 15268976 TI - Protective effect of Ligusticum chuanxiong and Angelica sinensis on endothelial cell damage induced by hydrogen peroxide. AB - Ligusticum chuanxiong and Angelica sinensis have been widely used in traditional Chinese medicine to treat some pathological settings such as atherosclerosis and hypertension. We determined the protective effect of the extract of Ligusticum chuanxiong and Angelica sinensis (ELCAS) on human umbilical vein endothelial cells (ECV304) damage induced by hydrogen peroxide. ECV304 cells were pre-treated with ELCAS and exposed to 5 mM hydrogen peroxide. The results show that ELCAS dose- and time-dependently protected ECV304 cells against hydrogen peroxide damage and suppressed the production of reactive oxygen species (ROS). The decrement of ROS may be associated with increased activities of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPX). Western blot analysis revealed that ELCAS significantly increased the phosphorylation of ERK and promoted eNOS expression. These observations indicate that ELCAS protected ECV304 cells against hydrogen peroxide damage by enhancing the antioxidative ability, activating ERK and eNOS signaling pathway. Our data also provide new evidence of Ligusticum chuanxiong and Angelica sinensis in preventing both cardiovascular and cerebrovascular diseases. PMID- 15268977 TI - Transport, transformation and distribution space of propofol in the rat liver studied by means of the indicator-dilution technique. AB - 1. The transport, transformation and distribution space of the endovenous anaesthetic propofol in the isolated perfused rat liver were investigated by using the multiple-indicator dilution technique with constant infusion (step input). 2. The behaviour of propofol in the liver was described by a space distributed variable transit-time model. The drug permeated the cell membrane at very high rates and its distribution into the cellular space was flow-limited. The apparent distribution space of propofol varied between 284 and 125 times the water space, and was inversely related to the tested portal concentrations (33 250 microM). 3. The corresponding ratios of intra- to extracellular concentration varied between 319 and 187, revealing a very high affinity of the liver for propofol. They most probably reflect binding to several cellular structures, including membranes and proteins. 3. The single-pass rate coefficients for biotransformation decreased with increases in the portal concentration of propofol. The liver released significant amounts of 4-hydroxypropofol, reaching 41.7 % of the total single pass of 67.2 microM propofol biotransformation. These results disprove previous notions that hydroxylation is rate limiting for conjugation and suggest that the liver might function as a 4-hydroxypropofol source for conjugation to glucuronic acid or sulfate in other tissues. PMID- 15268978 TI - Role of individual human cytochrome P450 enzymes in the in vitro metabolism of hydromorphone. AB - 1. The aim was to identify the individual human cytochrome P450 (CYP) enzymes responsible for the in vitro N-demethylation of hydromorphone and to determine the potential effect of the inhibition of this metabolic pathway on the formation of other hydromorphone metabolites. 2. Hydromorphone was metabolized to norhydromorphone (apparent Km = 206 - 822 microM, Vmax = 104 - 834 pmol min(-1) mg(-1) protein) and dihydroisomorphine (apparent Km = 62 - 557 microM, Vmax = 17 122 pmol min(-1) mg(-1) protein) by human liver microsomes. 5. In pooled human liver microsomes, troleandomycin, ketoconazole and sulfaphenazole reduced norhydromorphone formation by an average of 45, 50 and 25%, respectively, whereas furafylline, quinidine and omeprazole had no effect. In an individual liver microsome sample with a high CYP3A protein content, troleandomycin and ketoconazole inhibited norhydromorphone formation by 80%. 5. The reduction in norhydromorphone formation by troleandomycin and ketoconazole was accompanied by a stimulation in dihydroisomorphine production. Recombinant CYP3A4, CYP3A5, CYP2C9 and CYP2D6, but not CYP1A2, catalysed norhydromorphone formation, whereas none of these enzymes was active in dihydroisomorphine formation. 6. In summary, CYP3A and, to a lesser extent, CYP2C9 catalysed hydromorphone N-demethylation in human liver microsomes. The inhibition of norhydromorphone formation by troleandomycin and ketoconazole resulted in a stimulation of microsomal dihydroisomorphine formation. PMID- 15268979 TI - Identification of 2,3-diaminophenazine and of o-benzoquinone dioxime as the major in vitro metabolites of benzofuroxan. AB - 1. The results of an in vitro study of the metabolism of benzofuroxan using either cytosolic or microsomal fractions obtained from rat liver are reported. 2. Benzofuroxan was incubated with an appropriate volume of cytosol or microsomal suspension; control incubations were performed without the beta-nicotinamide adenine dinucleotide phosphate-generating system or, alternatively, by using the subcellular fractions inactivated by heating. Incubation mixtures were analysed by high-performance liquid chromatography. Two principal metabolites (M1, M2) were identified in the cytosolic fraction only. The dependence of M2 formation on thiol cofactors, incubation time and protein concentration was examined. 3. The two metabolites were isolated and characterized by their 1H-, 13C-nuclear magnetic resonance, infrared and mass spectra. The structures of o benzoquinonedioxime (2) and 2,3-diaminopleuozuc (3), were arranged to M1 and M2 respectively. The proposed structures were confirmed by the identity of the metabolites with authentic samples obtained by synthesis. X-ray analysis showed that the dioxime metabolite had an amphy configuration. 4. A metabolic scheme for the formation of the two products is proposed. PMID- 15268980 TI - Apparent absolute oral bioavailability in excess of 100% for a vitronectin receptor antagonist (SB-265123) in rat. I. Investigation of potential experimental and mechanistic explanations. AB - 1. SB-265123 is a novel alphavbeta3 (the vitronectin receptor) antagonist. Previous rat studies with it revealed an apparent absolute oral bioavailability (Fapp) of greater than 100%. The present studies were conducted to investigate the potential causes for this observation. 2. Of 49 SB-265123 analogues evaluated in rat using an identical experimental design, Fapp > 100% was observed for 22 of them, suggesting that the observed Fapp >100% with SB-265123 was not anomalous. All 22 compounds had clearances < 15 ml min(-1) kg(-1). However, Fapp>100% were not recorded for all low-clearance analogues. 3. Using SB-265123 as a model to investigate potential artefacts, it was demonstrated that using a chiral assay did not decrease Fapp. Additionally, qualitative sample analysis demonstrated that no metabolites were present in the plasma that could interfere with the liquid chromatography coupled with tandem mass spectrometry detection assay. The high Fapp was also dose-order-, delivery system- and vehicle-independent, and was not affected by the feeding status of the animals. Furthermore, a linearity experiment and an absorption study indicated that oral administration of SB 265123 does not result in hepatic portal vein concentrations that exceed the pharmacokinetic linearity of SB-265123. 4. These observations suggest that the observed Fapp > 100% for SB-265123 is not due to an experimental artefact or an obvious pharmacokinetic non-linearity. The mechanism(s) for this phenomenon is explored further in the second part of the present paper. PMID- 15268981 TI - Apparent absolute oral bioavailability in excess of 100% for a vitronectin receptor antagonist (SB-265123) in rat. II. Studies implicating transporter mediated intestinal secretion. AB - 1. Transporters have been increasingly identified as a factor in limiting the oral bioavailability of certain drugs. Previously, the present authors investigated a compound (SB-265123) with an apparent absolute oral bioavailability (Fapp) consistently > 100%, and excluded likely artefactual causes for this observation, as well as standard considerations of non-stationary or non-linear pharmacokinetics. The data led the authors to believe that SB 265123 might be a transporter substrate in the rat, and it was hypothesized that transporter interactions might be responsible for the observed Fapp > 100%. 2. In the present study, a model was proposed incorporating rapid and complete absorption and elimination by a saturable intestinal secretory pathway. Intestinal secretion was demonstrated for SB-265123 using a rat single-pass intestinal perfusion technique. In addition, in a study employing both independent and simultaneous intravenous and oral administration of SB-265123, exposure to SB-265123 was greater than additive on joint intravenous and oral administration, lending further support to the hypothesis of a saturable transporter. Furthermore, in a study with co-administration of GF120918A, a transporter inhibitor, the observed Fapp for SB-265123 was only 84 +/- 17%, providing additional evidence for transporter involvement in the >100% Fapp phenomenon. 3. Experience with SB-265123 illustrates a counterintuitive impact of transporters on oral bioavailability and highlights the importance of considering transporter interactions in the systemic disposition of xenobiotics, even those not demonstrating low oral bioavailability. PMID- 15268982 TI - Comparative disposition of [14C]ertapenem, a novel carbapenem antibiotic, in rat, monkey and man. AB - 1. The disposition and metabolism of ertapenem, a carbapenem antibiotic, was examined in rat, monkey and man. Sprague-Dawley rats and Rhesus monkeys were given, by intravenous administration, radiolabelled doses of ertapenem (60 and 30 mg kg(-1), respectively), and healthy normal volunteers received a single fixed dose of 1000 mg. Urine and faeces were collected for determination of total radioactivity. 2. In healthy volunteers, [14C]ertapenem was eliminated by a combination of hydrolytic metabolism to a beta-lactam ring-opened derivative and renal excretion of unchanged drug. Approximately equal amounts were excreted as a beta-lactam ring-opened metabolite and unchanged drug (36.7 and 37.5% of dose, respectively). A secondary amide hydrolysis product accounted for about 1% of the dose in man. About 10% of the administered radioactivity was recovered in faeces, which suggested that a minor fraction underwent biliary and/or intestinal excretion. 3. In animals, a greater fraction of the dose was eliminated via metabolism; excretion of unchanged drug accounted for 17 and 5% of dose in rats and monkeys, respectively. In monkeys, the beta-lactam ring-opened and amide hydrolysis metabolites accounted for 74.8 and 7.59% of the dose, respectively, whereas in rats, these metabolites accounted for 31.9 and 20% of dose, respectively. 4. In vitro studies with fresh rat tissue homogenates indicated that lung and kidney were the primary organs involved in mediating formation of the beta-lactam ring-opened metabolite. The specific inhibitor of dehydropeptidase-I, cilastatin, inhibited the in vivo and in vitro metabolism of ertapenem in rats, which suggested strongly that the hydrolysis of ertapenem in lung and kidney was mediated by this enzyme. PMID- 15268997 TI - Postural control in children: implications for pediatric practice. AB - Based on a systems theory of motor control, reactive postural control (RPA) and anticipatory postural control (APA) in children are reviewed from several perspectives in order to develop an evidence-based intervention strategy for improving postural control in children with limitations in motor function. Research on development of postural control, postural control in children with specific motor disabilities, and interventions to improve postural control is analyzed. A strategy for intervention to improve postural control systems at the impairment and functional activity levels based on a systems theoretical perspective is presented. Suggestions for research to improve evidence for best practice are provided. PMID- 15268998 TI - Muscle force and range of motion as predictors of standing balance in children with cerebral palsy. AB - Children with cerebral palsy frequently receive therapeutic intervention to remediate standing balance deficits. Evaluation of the impairments associated with poor balance could facilitate more effective treatment programs. This study evaluated the relationship between lower extremity force production, range of motion and standing balance in thirty-five children between the ages of 6 and 14 years of age with spastic cerebral palsy. Standing balance was evaluated using the Pediatric Clinical Test of Sensory Interaction (P-CTSIB). Hand-held dynamometry was used to assess force production and goniometry was used to assess range of motion. The results indicated that force production and range of motion are highly related to standing balance. Blocked, hierarchical multiple regression analysis revealed that force production explained 41% of the variance in P-CTSIB scores in this sample, while range of motion explained an additional 13%. Therefore, the total variance explained by these variables was 54%. Results of this study suggest that impairment level testing may allow the development of more effective individualized intervention programs to remediate balance deficits. Clinical suggestions are provided. PMID- 15268999 TI - Activity-focused motor interventions for children with neurological conditions. AB - This article presents a model to guide activity-focused physical therapy and occupational therapy interventions for children with neurological conditions. Activity-focused interventions involve structured practice and repetition of functional actions and are directed toward the learning of motor tasks that will increase independence and participation in daily routines. According to this model, the pediatric therapist: (1) develops activity-related goals in collaboration with the child and the family; (2) plans activity-focused interventions by adapting knowledge of motor learning to the child's individual learning strengths and needs; and (3) integrates impairment-focused intervention with activity-focused intervention. PMID- 15269000 TI - Changes in mobility of children with cerebral palsy over time and across environmental settings. AB - This study examined changes in mobility methods of children with cerebral palsy (CP) over time and across environmental settings. Sixty-two children with CP, ages 6-14 years and classified as levels II-IV on the Gross Motor Function Classification System, were randomly selected from a larger data base and followed for three to four years. On each of several assessments, parents completed a questionnaire on their child's usual mobility methods in the home, school, and outdoors/community settings. During the first assessment interval, mobility methods increased to methods requiring more gross motor control. During the second assessment interval, mobility methods were unchanged or decreased to methods requiring less gross motor control. Changes within the child and within the environment are hypothesized to occur and to impact changes in mobility methods. Screening at regular intervals is recommended to monitor changes in mobility. Interventions to enhance mobility may be indicated during periods of change in the child or exposure to new environments. PMID- 15269001 TI - Enhancing prehension in infants and children: fostering neuromotor strategies. AB - Learning to reach for and manipulate objects requires considerable neuromotor control and flexibility. Through environmental and object exploration individual neuromotor strategies expand, and prehensile skills improve, as infants and children overcome constraints. Infants and children with prehensile deficits often have difficulty exploring objects and the environment, thus, may not sufficiently develop the strategies needed to expand their prehensile skills. This article reviews neuromotor factors that influence prehension development, discusses limitations to prehensile function and provides guidelines that can be used to examine and enhance prehensile behaviors in infants and young children based on a task-oriented approach addressing impairments, motor strategies and function. PMID- 15269002 TI - Reliability of a measure of muscle extensibility in fullterm and preterm newborns. AB - PURPOSE: The purpose of this study was to examine the test-retest and inter-rater reliability of a measure of muscle extensibility developed by Tardieu, de la Tour, Bret, and Tardieu (1982) in fullterm and preterm newborns. METHOD: Twenty one fullterm infants and twenty preterm infants were examined by two physical therapists. Each physical therapist measured AO (shortened position of the muscle belly and lengthened tendon) and AMax (maximum muscle belly and tendon length) of the gastrocnemius/ soleus muscle twice in succession. Reliability was assessed using intraclass correlation coefficients (2,2) and (3,2). RESULTS: Inter-rater reliability between the two examiners ranged from .86 to .97 and test-retest reliability on the two measures ranged from .91 to .98. CONCLUSIONS: The results suggest that this measure of muscle extensibility is reliable in the gastrocnemius/soleus muscle with fullterm and preterm newborns. Further research is needed to investigate if differences in muscle extensibility are present between fullterm and preterm infants and the relationship between muscle extensibility and active movement. PMID- 15269003 TI - Functional genomics of lung disease. Second annual Pittsburgh International Lung Conference, October 2003. PMID- 15269004 TI - Cholinergic receptor-mediated differential cytoskeletal recruitment of actin- and integrin-binding proteins in intact airway smooth muscle. AB - We tested the hypothesis that cholinergic receptor stimulation recruits actin- and integrin-binding proteins from the cytoplasm to the cytoskeleton-membrane complex in intact airway smooth muscle. We stimulated bovine tracheal smooth muscle with carbachol and fractionated the tissue homogenate into pellet (P) and supernatant (S) by ultracentrifugation. In unstimulated tissues, calponin exhibited the highest basal P-to-S ratio (P/S; 2.74 +/- 0.47), whereas vinculin exhibited the lowest P/S (0.52 +/- 0.09). Cholinergic receptor stimulation increased P/S of the following proteins in descending order of sensitivity: alpha actinin > talin approximately metavinculin > alpha-smooth muscle actin > vinculin approximately calponin. Carbachol induced ERK1/2 phosphorylation by 300% of basal value. U0126 (10 microM) completely inhibited carbachol-induced ERK1/2 phosphorylation but did not significantly affect the correlation between alpha actinin P/S and carbachol concentration. This observation indicates that cytoskeletal/membrane recruitment of alpha-actinin is independent of ERK1/2 mitogen-activated protein kinase activation. Metavinculin and vinculin are splice variants of a single gene, but metavinculin P/S was significantly higher than vinculin P/S. Furthermore, the P/S of metavinculin but not vinculin increased significantly in response to cholinergic receptor stimulation. Calponin and alpha actinin both belong to the family of calponin homology (CH) domain proteins. However, unlike alpha-actinin, the calponin P/S did not change significantly in response to cholinergic receptor stimulation. These findings indicate differential cytoskeletal/membrane recruitment of actin- and integrin-binding proteins in response to cholinergic receptor stimulation in intact airway smooth muscle. alpha-Actinin, talin, and metavinculin appear to be key cytoskeletal proteins involved in the recruitment process. PMID- 15269005 TI - Signals regulating trafficking of Menkes (MNK; ATP7A) copper-translocating P-type ATPase in polarized MDCK cells. AB - The Menkes protein (MNK; ATP7A) functions as a transmembrane copper-translocating P-type ATPase and plays a vital role in systemic copper absorption in the gut and copper reabsorption in the kidney. Polarized epithelial cells such as Madin-Darby canine kidney (MDCK) cells are a physiologically relevant model for systemic copper absorption and reabsorption in vivo. In this study, cultured MDCK cells were used to characterize MNK trafficking and enabled the identification of signaling motifs required to target the protein to specific membranes. Using confocal laser scanning microscopy and surface biotinylation we demonstrate that MNK relocalizes from the Golgi to the basolateral (BL) membrane under elevated copper conditions. As previously shown in nonpolarized cells, the metal binding sites in the NH2-terminal domain of MNK were found to be required for copper regulated trafficking from the Golgi to the plasma membrane. These data provide molecular evidence that is consistent with the presumed role of this protein in systemic copper absorption in the gut and reabsorption in the kidney. Using site directed mutagenesis, we identified a dileucine motif proximal to the COOH terminus of MNK that was critical for correctly targeting the protein to the BL membrane and a putative PDZ target motif that was required for localization at the BL membrane in elevated copper. PMID- 15269006 TI - P2X7 receptor-mediated apoptosis of human cervical epithelial cells. AB - Normal human ectocervical epithelial (hECE) cells undergo apoptosis in culture. Baseline apoptosis could be increased by shifting cells to serum-free medium and blocked by lowering extracellular calcium. Treatment with the ATPase apyrase attenuated baseline apoptosis, suggesting that extracellular ATP and purinergic mechanisms control the apoptosis. Treatment with ATP and the P2X7 receptor analog 2'-3'-O-(4-benzoylbenzoyl)adenosine 5'-triphosphate (BzATP) increased apoptosis significantly, in a time- and dose-related manner. The threshold of ATP effect was 0.5 microM in hECE cells and approximately 1 microM in CaSki cancer cells. The apoptotic effect of BzATP was additive in part to that of tumor necrosis factor (TNF)-alpha, and it could be attenuated by lowering extracellular calcium and by treatment with the caspase-9 inhibitor Leu-Glu-His-Asp-O-methyl fluoromethylketone (LEHD-FMK). Treatment with BzATP activated caspase-9, and, in contrast to TNF-alpha, it had only a mild effect on caspase-8. Both BzATP and TNF alpha activated caspase-3, suggesting that BzATP activates predominantly the mitochondrial apoptotic pathway. Both hECE and CaSki cells secrete ATP into the extracellular fluid, and mean ATP activity in conditioned medium was approximately 0.5 microM, which is in the range of values that suffice to activate the P2X7 receptor. On the basis of these findings we propose a novel autocrine-paracrine mechanism of cervical cell apoptosis that operates by P2X7 receptor control of cytosolic calcium and utilizes the mitochondrial apoptotic pathway. PMID- 15269007 TI - Decreased affinity for oxygen of cytochrome-c oxidase in Leigh syndrome caused by SURF1 mutations. AB - Mutations in the gene SURF1 prevent synthesis of cytochrome-c oxidase (COX) specific assembly protein and result in a fatal neurological disorder, Leigh syndrome. Because this severe COX deficiency presents with barely detectable changes of cellular respiratory rates under normoxic conditions, we analyzed the respiratory response to low oxygen in cultured fibroblasts harboring SURF1 mutations with high-resolution respirometry. The oxygen kinetics was quantified by the partial pressure of oxygen (PO2) at half-maximal respiration rate (P50) in intact coupled cells and in digitonin-permeabilized uncoupled cells. In both cases, the P50 in patients was elevated 2.1- and 3.3-fold, respectively, indicating decreased affinity of COX for oxygen. These results suggest that at physiologically low intracellular PO2, the depressed oxygen affinity may lead in vivo to limitations of respiration, resulting in impaired energy provision in Leigh syndrome patients. PMID- 15269009 TI - Fear of cancer. PMID- 15269010 TI - Enhancing research in academic radiology departments: recommendations of the 2003 Consensus Conference. PMID- 15269011 TI - Adenoid cystic carcinoma of the airways: helical CT and histopathologic correlation. PMID- 15269012 TI - Mucinous cystadenoma of the lung. PMID- 15269013 TI - Does CT of thymic epithelial tumors enable us to differentiate histologic subtypes and predict prognosis? AB - OBJECTIVE: The aims of our study were to describe the CT findings of thymic epithelial tumors and to correlate these findings with the histopathologic subtypes and prognosis. MATERIALS AND METHODS: The CT findings of thymic epithelial tumors were analyzed in 91 patients who had undergone surgery between May 1995 and June 2002. Two observers, who were unaware of the histopathologic classification made in accordance with World Health Organization (WHO) recommendations and the prognosis of the tumors, retrospectively reviewed the initial CT findings in terms of the contours and shapes of the tumors and the presence of necrosis, calcification, mediastinal fat or great vessel invasion, pleural seeding, contrast enhancement, and lymph node enlargement. These findings were compared with the simplified subgroups of WHO histologic classification (low risk thymomas [types A, AB, and B1], high-risk thymomas [types B2 and B3], and thymic carcinomas [type C]) and with postoperative recurrence. RESULTS: The study found 31 low-risk thymomas (eight type A, 16 type AB, and seven type B1 tumors), 45 high-risk thymomas (25 type B2 and 20 type B3), and 15 thymic carcinomas (type C). Lobulated contour was more often seen in high-risk thymomas (26/45, 58%; p = 0.0456) and thymic carcinomas (10/15, 67%; p = 0.033) than in low-risk thymomas (9/31, 29%). Mediastinal fat invasion was more often seen in thymic carcinomas (5/15, 33%; p = 0.0133) than in low-risk thymomas (1/31, 3%). Great vessel invasion was seen only in thymic carcinomas (2/15, 13%; p = 0.0244). Tumors with a lobulated or irregular contour, an oval shape, mediastinal fat or great vessel invasion, and pleural seeding showed significantly more frequent recurrence and metastasis (all, p < 0.05). CONCLUSION: Although CT is of limited value in differentiating histologic subtypes according to the WHO classification, CT findings may serve as predictors of postoperative recurrence or metastasis for the thymic epithelial tumors. PMID- 15269014 TI - Partial tracheal duplication: MDCT bronchoscopic diagnosis. PMID- 15269015 TI - Pulmonary tumorlets: CT findings. AB - OBJECTIVE: Pulmonary tumorlets are defined in pathologic terms as benign localized neuroendocrine cell proliferations a few millimeters in size that are usually associated with damaged and ectatic small airways. The purpose of this study was to determine the frequency with which pulmonary tumorlets can be seen on CT and to describe their CT appearance. CONCLUSION: In 33 patients with proven tumorlets, a nodule was visible on CT in the same region as that of the resected specimen. Despite its ominous-sounding name, a pulmonary tumorlet represents benign tissue that may manifest as a subcentimeter pulmonary nodule and should be considered in the differential diagnosis of small pulmonary nodules visible on CT. PMID- 15269016 TI - Application of an artificial neural network to high-resolution CT: usefulness in differential diagnosis of diffuse lung disease. AB - OBJECTIVE: The purpose of our study was to evaluate the diagnostic performance of an artificial neural network (ANN) in differentiating among certain diffuse lung diseases using high-resolution CT (HRCT) and the effect of ANN output on radiologists' diagnostic performance. MATERIALS AND METHODS: We selected 130 clinical cases of diffuse lung disease. We used a single three-layer, feed forward ANN with a back-propagation algorithm. The ANN was designed to differentiate among 11 diffuse lung diseases by using 10 clinical parameters and 23 HRCT features. Therefore, the ANN consisted of 33 input units and 11 output units. Subjective ratings for 23 HRCT features were provided independently by eight radiologists. All clinical cases were used for training and testing of the ANN by implementing a round-robin technique. In the observer test, a subset of 45 cases was selected from the database of 130 cases. HRCT images were viewed by eight radiologists first without and then with ANN output. The radiologists' performance was evaluated with receiver operating characteristic (ROC) analysis with a continuous rating scale. RESULTS: The average area under the ROC curve for ANN performance obtained with all clinical parameters and HRCT features was 0.956. The diagnostic performance of four chest radiologists and four general radiologists was increased from 0.986 to 0.992 (p = 0.071) and 0.958 and 0.971 (p < 0.001), respectively, when they used the ANN output based on their own feature ratings. CONCLUSION: The ANN can provide a useful output as a second opinion to improve general radiologists' diagnostic performance in the differential diagnosis of certain diffuse lung diseases using HRCT. PMID- 15269017 TI - Imaging of hereditary hemorrhagic telangiectasia. PMID- 15269018 TI - Size and morphology of the trachea before and after lung volume reduction surgery. AB - OBJECTIVE: The purpose of this investigation was to determine the effect of lung volume reduction surgery on measured tracheal features. MATERIALS AND METHODS: Twenty-four male and 19 female patients with emphysema underwent lung volume reduction surgery, pulmonary function testing, and repeated CT. The tracheal air column was segmented from axial images. The sagittal and coronal dimensions of the intrathoracic trachea were determined. Tracheal morphology was quantified using the tracheal (coronal and sagittal dimensions) and circularity indexes. The results were compared with pulmonary function test results. RESULTS: Morphologic appearance of the intrathoracic trachea was consistent before and 3 months after surgery. The group means of the tracheal length, mean area, and volume were 78.60 mm (+/- 16.88 mm), 283.84 mm(2) (+/- 61.47 mm(2)), and 22.59 cm(3) (+/- 7.69 cm(3)), respectively, before surgery and 67.53 mm (+/- 15.78 mm), 309.12 mm(2) (+/- 79.83 mm(2)), and 20.99 cm(3) (+/- 7.27 cm(3)), respectively, after surgery (p < 0.05). Mean tracheal indexes were 0.85 (+/- 0.11) before surgery and 0.82 (+/- 0.04) after surgery (p < 0.01). Mean circularity indexes were 0.91 (+/- 0.03) before surgery and 0.90 (+/- 0.04) after surgery (p < 0.05). The size of the trachea was significantly correlated with lung volume before and after surgery (p < 0.05). The changes in tracheal features and changes in pulmonary function were not correlated (p > 0.05), except for tracheal area (p < 0.05). CONCLUSION: Our data suggest that tracheal dimensions reflect the severity of emphysema as reflected by increased lung volumes. Tracheal features were poor predictors of changes in postsurgical pulmonary function parameters evaluated in this preliminary study. PMID- 15269019 TI - MRI of seemingly isolated greater trochanteric fractures. AB - OBJECTIVE: The objective of this article is to show that greater trochanteric fractures commonly perceived on routine radiographs as isolated are often neither isolated nor minor and that MR images can serve as a basis for more informed treatment by revealing the actual extent of such fractures in acute posttraumatic settings. CONCLUSION: A pitfall in diagnosing seemingly isolated greater trochanteric fractures on routinely used imaging techniques lies in the fact that the injuries usually involve a large anatomic area. In our experience, MRI more accurately defines the true geographic extent of greater trochanteric fractures sustained through acute trauma than do radiography and bone scintigraphy and thus could provide a more reliable basis for anticipating complications and for planning appropriate treatment. PMID- 15269020 TI - Imaging of ancient schwannoma. AB - OBJECTIVE: We surveyed the clinical symptoms and radiologic features of ancient schwannoma, a rare variant of schwannoma characterized by degenerative changes. MATERIALS AND METHODS: We present the clinical, radiologic, and pathologic features in seven patients with ancient schwannoma (mean age, 62 years; range, 45 80 years) treated at our department between 1998 and 2003. RESULTS: The most characteristic clinical features were a sign like Tinel's sign and a long interval between the onset of symptoms and surgery (mean interval, 8.3 years). Ancient schwannomas can grow large; the biggest tumor seen in our study was 14 cm long. The highly accurate radiologic assessment made possible with contrast enhanced MRI and CT scanning showed enhancement at a peridegenerative area and sometimes at a capsule. These findings differ from those of the typical schwannoma and neurofibroma patterns reported to date. Furthermore, bone scintigraphy showed uptake in the tumor, but no accumulation was seen on gallium 67 citrate scintigraphy. CONCLUSION: The characteristic clinical and radiologic findings of ancient schwannoma should make it possible to differentiate it from malignant tumors. PMID- 15269021 TI - Giant cell tumors of the tendon sheath: analysis of sonographic findings. AB - OBJECTIVE: The purpose of this study was to analyze the sonographic characteristics of giant cell tumors of the tendon sheath. CONCLUSION: Giant cell tumors of the hand typically appear as solid, homogeneous hypoechoic masses with detectable internal vascularity that are associated with the flexor tendons of the fingers. PMID- 15269022 TI - Osteoid osteoma of the tarsal cuboid mimicking osteomyelitis. PMID- 15269023 TI - Musculoskeletal MRI at 3.0 T: relaxation times and image contrast. AB - OBJECTIVE: The purpose of our study was to measure relaxation times in musculoskeletal tissues at 1.5 and 3.0 T to optimize musculoskeletal MRI methods at 3.0 T. MATERIALS AND METHODS: In the knees of five healthy volunteers, we measured the T1 and T2 relaxation times of cartilage, synovial fluid, muscle, marrow, and fat at 1.5 and 3.0 T. The T1 relaxation times were measured using a spiral Look-Locker sequence with eight samples along the T1 recovery curve. The T2 relaxation times were measured using a spiral T2 preparation sequence with six echoes. Accuracy and repeatability of the T1 and T2 measurement sequences were verified in phantoms. RESULTS: T1 relaxation times in cartilage, muscle, synovial fluid, marrow, and subcutaneous fat at 3.0 T were consistently higher than those measured at 1.5 T. Measured T2 relaxation times were reduced at 3.0 T compared with 1.5 T. Relaxation time measurements in vivo were verified using calculated and measured signal-to-noise results. Relaxation times were used to develop a high-resolution protocol for T2-weighted imaging of the knee at 3.0 T. CONCLUSION: MRI at 3.0 T can improve resolution and speed in musculoskeletal imaging; however, interactions between field strength and relaxation times need to be considered for optimal image contrast and signal-to-noise ratio. Scanning can be performed in shorter times at 3.0 T using single-average acquisitions. Efficient higher-resolution imaging at 3.0 T can be done by increasing the TR to account for increased T1 relaxation times and acquiring thinner slices than at 1.5 T. PMID- 15269024 TI - Fabellar snapping as a cause of knee pain after total knee replacement: assessment using dynamic sonography. PMID- 15269025 TI - Humeral avulsion of the posterior band of the inferior glenohumeral ligament: MR arthrography and clinical correlation in 17 patients. AB - OBJECTIVE: The purpose of this article is to describe the MR arthrography findings of humeral avulsion of the posterior band of the inferior glenohumeral ligament in 17 patients. To elucidate the clinical importance of this abnormality, we also correlate our imaging findings with the presence of coexisting structural abnormalities; clinical presentation; and, when available, arthroscopic evaluation (n = 8). CONCLUSION: Humeral avulsion or insufficiency of the posterior band of the inferior glenohumeral ligament can be easily detected using MR arthrography. This ligamentous abnormality may be seen in isolation, or it may occur in conjunction with posterior or, less often, anteroinferior capsulolabral abnormalities. The presence of this lesion in a subgroup of patients with the clinical diagnosis of multidirectional instability may offer insight into the causes and pathogenesis of this complex entity. PMID- 15269026 TI - Radiofrequency ablation of 40 lung neoplasms: preliminary results. AB - OBJECTIVE: Radiofrequency thermal ablation is a minimally invasive treatment widely used for treatment of liver neoplasms and has also been tested on other types of tumor. Few studies have been published regarding the use of radiofrequency thermal ablation in the treatment of lung neoplasms. This study was performed to evaluate the technical feasibility, the safety, and the efficacy of lung radiofrequency thermal ablation. SUBJECTS AND METHODS: Between February 2002 and March 2003, 18 subjects with unresectable lung neoplasms, four of whom had primary neoplasms and 14 of whom had metastatic neoplasms, underwent lung radiofrequency ablation. The technique was performed percutaneously using a monopolar cooled-tip electrode needle under CT guidance with the patient under general anesthesia. Patients had no more than three nodules with a total diameter of 10 cm and no evidence of extrathoracic disease. A total of 40 nodules were treated in 24 therapeutic sessions. After treatment, patients underwent follow-up every 3 months by CT and nuclear MRI with gadolinium for a median time of 8 months (range, 2-14 months). RESULTS: No evidence of local relapse was discovered in 94.4% of subjects. The treatment was safe and well tolerated. Complications encountered included massive pneumothorax, which occurred in one subject, requiring pleural drainage. Other complications were moderate pneumothorax (also requiring pleural drainage), cough, fever, slight dyspnea, and pain, but these complications were short in duration and successfully treated. CONCLUSION: Radiofrequency thermal ablation is a promising technique in the treatment of patients with lung neoplasms and has been found to be both safe and technically feasible. PMID- 15269027 TI - TIPS versus transcatheter sclerotherapy for gastric varices. AB - OBJECTIVE: The purpose of our study was to compare the efficacy and long-term results of transjugular intrahepatic portosystemic shunt (TIPS) with those of transcatheter sclerotherapy for the treatment of gastric varices. MATERIALS AND METHODS: A total of 139 cirrhotic patients with gastric varices underwent endovascular treatment. Of the 139 patients, 104 without hepatocellular carcinoma were enrolled; 27 patients were treated with TIPS, and 77 patients with transcatheter sclerotherapy. Bleeding of gastric varices and survival rates were compared between the TIPS and transcatheter sclerotherapy groups. Multivariate analysis was used to identify the prognostic factors for gastric variceal bleeding and survival. Changes in liver function were evaluated in each group. RESULTS: The cumulative gastric variceal bleeding rate at 1 year was 20% in the TIPS group and 2% in the transcatheter sclerotherapy group (p < 0.01). The prognostic factor associated with gastric variceal bleeding was the treatment method. The cumulative survival rates at 1, 3, and 5 years were, respectively, 81%, 64%, and 40% in the TIPS group and 96%, 83%, and 76% in the transcatheter sclerotherapy group (p < 0.01). The prognostic factors for survival were the treatment method and the Child-Pugh classification of liver disease. For patients categorized in Child-Pugh class A, the survival rate was higher in the transcatheter sclerotherapy group than in the TIPS group (p < 0.01). For patients in Child-Pugh classes B and C, no significant difference was seen between the two groups. Liver function tended to improve in the transcatheter sclerotherapy group. CONCLUSION: Transcatheter sclerotherapy may provide better control of gastric variceal bleeding than TIPS. Transcatheter sclerotherapy may contribute to a higher survival rate than TIPS in patients with Child-Pugh class A disease. PMID- 15269028 TI - Risks of outpatient angiography and interventional procedures: a prospective study. AB - OBJECTIVE: The purpose of this study was to evaluate complications in diagnostic and interventional angiographic procedures performed on outpatients. MATERIALS AND METHODS: Data were collected prospectively for 2,683 procedures performed on an outpatient basis in 2,248 patients from the period March 1997 to March 2002. Patients were assessed by nursing or medical staff within 2-4 hr of the procedure and again via telephone 24-48 hr after the procedure. The collected data were summarized on the basis of procedure type into four main groupings: aortofemoral studies, cerebral studies, interventional procedures, and other studies. Complication frequency distribution was determined for each procedure type. An interim summary of complication rates was prepared for the period March 1997 to June 1999. Statistical analysis using a two-tailed z-test for the comparison of two proportions was performed to determine if a significant difference existed in the rates of complications from data collected before and after the June 1999 summary. RESULTS: Ninety-one percent of cases completed follow-up. In total, 561 complications were identified in 2,436 cases (23%). Most complications consisted of either local pain or puncture site hematoma and bruising. No deaths occurred. In the 1,128 diagnostic aortofemoral studies performed, 211 complications (19%) occurred. In the 359 cerebral studies, 87 complications (24%) occurred. The 441 interventional procedures resulted in 146 complications (33%). In the remaining 508 procedures, 117 complications (23%) occurred. Major complications in each group are presented. CONCLUSION: We observed a low incidence of complications requiring further treatment or resulting in a permanent deficit. The rates are comparable to published data from similar studies and practice standards guidelines. A statistically significant improvement was seen in the total complication rate between the periods March 1997-June 1999 and July 1999-March 2002 (p = 0.01). PMID- 15269029 TI - Does percutaneous liver biopsy of hepatocellular carcinoma cause hematogenous dissemination? An in vivo study with quantitative assay of circulating tumor DNA using methylation-specific real-time polymerase chain reaction. AB - OBJECTIVE: Our purpose was to find out whether percutaneous biopsy of hepatocellular carcinoma will cause significant dissemination of tumor into the circulation by quantitative analysis of circulating tumor DNA. SUBJECTS AND METHODS: In this prospective study of 32 patients with suspected hepatocellular carcinoma who underwent sonographically guided liver biopsy, a peripheral venous blood sample was obtained before and 5 min after the procedure. Biopsy was performed using an 18-gauge biopsy gun. DNA was extracted from the plasma of the blood samples for methylation-specific polymerase chain reaction. Quantitative measures of the plasma tumor DNA were determined with real-time quantitative polymerase chain reaction, and the amount was expressed as a methylation index (%) in plasma. RESULTS: Nineteen (59.4%) of 32 patients did not have detectable p16 tumor suppressor gene marker (p16M) in plasma before biopsy, and they showed no detectable plasma p16M after biopsy. Thirteen (65%) of 20 patients had p16M identified in the plasma before liver biopsy. Quantitative analysis of the plasma tumor DNA in these 13 patients showed no statistically significant difference in the methylation index before and after biopsy (p = 0.345, Wilcoxon's signed rank test). CONCLUSION: No evidence exists that percutaneous liver biopsy results in hematogenous dissemination of hepatocellular carcinoma as shown by quantitative analysis of circulating tumor DNA (p16M) using methylation-specific real-time polymerase chain reaction. PMID- 15269030 TI - Enteric feeding with gastric decompression: management with separate gastric accesses. AB - OBJECTIVE: Our purpose is to describe our experience with combined enteral feeding and gastric decompression or drainage in debilitated patients with persistent gastroesophageal reflux using two separate catheters. CONCLUSION: The placement of two percutaneous catheters through separate skin sites is a feasible and successful approach to providing enteral feeding and gastric decompression or drainage in debilitated patients with persistent gastroesophageal reflux and aspiration pneumonia. PMID- 15269031 TI - The catheter-driven MRI scanner: a new approach to intravascular catheter tracking and imaging-parameter adjustment for interventional MRI. AB - OBJECTIVE: Our aim was to test the feasibility of a hands-free approach to MRI that allows the interventionalist to track an angiographic catheter in real time throughout the procedure and to automatically change imaging parameters by catheter manipulation. MATERIALS AND METHODS: A tracking method that is based on an active device localization was implemented on a 1.5-T MRI scanner. The system determines the current position and orientation of a catheter in 3D space in an endless feedback loop. Automatic scanning plane-adjustment procedures written in the software of the MRI system ensure image acquisition at the location of the catheter tip. The system calculates the device velocity to automatically adjust parameters such as field of view (FOV) and resolution. To evaluate the feasibility and performance in vivo and ex vivo, we performed experiments in two vessel phantoms and on six pigs. RESULTS: The system collected the tracking data within 40 msec; an additional 10-20 msec was then required to perform the localization and velocity calculations and to update the image parameters. The system could localize a motionless catheter in the aorta in 100% and a moving catheter in 98% of measured attempts. The system responded in real time to changes in device velocity by dynamically adjusting spatial resolution and FOV in both phantom and porcine trials. Using this technique, we successfully catheterized the renal artery in two pigs. CONCLUSION: Active tracking, combined with automatic scanning plane and imaging parameter adjustment, provides an intuitive MRI scanner interface for the guidance of the vascular procedure. PMID- 15269032 TI - Internal abdominal herniations. PMID- 15269033 TI - Imaging of an accessory spleen presenting as a slow-growing mass in the transplanted pancreas. PMID- 15269034 TI - Relationship between diffuse esophageal spasm and lower esophageal sphincter dysfunction on barium studies and manometry in 14 patients. AB - OBJECTIVE: We sought to reassess the function and clinical characteristics of the lower esophageal sphincter in a series of patients with radiographically defined diffuse esophageal spasm. MATERIALS AND METHODS: In reviewing records in the radiology database at our hospital, we identified 14 patients with diffuse esophageal spasm confirmed on barium studies who also underwent esophageal manometry. The radiographic findings were reviewed and correlated with the manometric findings. Medical records were also reviewed to determine the clinical presentation, treatment, and patient course. RESULTS: All 14 patients were symptomatic, presenting with dysphagia, chest pain, or both. All the barium studies revealed intermittently absent or weakened peristalsis, with nonperistaltic contractions that were moderate in six patients (43%) and marked in eight patients (57%) (contractions nearly obliterating the lumen in six and completely obliterating the lumen in two). Nine patients (64%) had impaired opening of the lower esophageal sphincter, manifested by beaklike narrowing of the distal esophagus, and five (36%) had normal opening of the lower esophageal sphincter. Manometry revealed abnormal peristalsis in all 14 patients, with repetitive simultaneous contractions in eight (57%) and lower esophageal sphincter dysfunction in 12 (86%). All eight patients with lower esophageal sphincter dysfunction or incomplete relaxation of the lower esophageal sphincter on barium studies or manometry who were treated with the Clostridium botulinum toxin or endoscopic balloon dilatation had a positive response. CONCLUSION: Our preliminary data show that diffuse esophageal spasm is characterized on barium studies by frequent lower esophageal sphincter dysfunction rather than a classic corkscrew appearance. Barium and manometric studies may have complementary roles in the evaluation of patients with diffuse esophageal spasm. PMID- 15269035 TI - Gastrointestinal stromal tumors of the duodenum: CT and barium study findings. AB - OBJECTIVE: The purpose of this study was to describe the CT and radiographic features of gastrointestinal stromal tumors of the duodenum. CONCLUSION: Gastrointestinal stromal tumors of the duodenum appear on barium studies as extrinsically compressing or submucosal masses with or without ulceration. These tumors usually appear on contrast-enhanced CT as well-defined masses with an exoenteric growth pattern and relatively good heterogeneous enhancement. PMID- 15269036 TI - Staging of rectal cancer: diagnostic potential of multiplanar reconstructions with MDCT. AB - OBJECTIVE: The purpose of this study was to evaluate whether multiplanar reconstructions (MPRs) of MDCT could improve local staging of rectal cancer. CONCLUSION: Adding MPRs, on the basis of the MDCT data sets, provides definite improvements in the accurate local staging of rectal cancer compared with standard axial reconstructions alone. PMID- 15269037 TI - Translucency rendering in 3D endoluminal CT colonography: a useful tool for increasing polyp specificity and decreasing interpretation time. PMID- 15269038 TI - Relationship between various patterns of transient increased hepatic attenuation on CT and portal vein thrombosis related to acute cholecystitis. AB - OBJECTIVE: We sought to investigate the prevalence of portal vein thrombosis in patients with acute cholecystitis and the relationship between portal vein thrombosis and the various patterns of transient increased hepatic attenuation on CT. MATERIALS AND METHODS: We studied 72 of 107 patients with acute cholecystitis who, during a 3-year period, underwent dual-phase contrast-enhanced CT before percutaneous cholecystostomy or cholecystectomy. CT scans were retrospectively reviewed for the presence of portal vein thrombosis and location of the thrombi and for patterns of transient increased hepatic attenuation, which were classified as either pericholecystic, segmental, or mixed. RESULTS: Portal vein thrombi (two in hepatic segment IV, three in the left portal vein, and one in the right posterior portal vein) were found in six (8.3%) of 72 patients, and in those patients, transient increased attenuation with a segmental (five patients) or mixed (one patient) pattern was seen on CT. The pattern of transient increased attenuation in the 54 patients without portal vein thrombosis was pericholecystic in 41 (75.9%) and mixed in 13 (24.1%). Nineteen patients had segmental distribution (segmental or mixed pattern) that in 31.6% (6/19) of the patients was associated with portal vein thrombosis. Segmental distribution was more frequently found in those patients who had acute cholecystitis with portal vein thrombosis than in those who had only acute cholecystitis (p = 0.001). CONCLUSION: In patients with acute cholecystitis, portal vein thrombosis is not uncommon. Patterns of transient increased hepatic attenuation were found to vary, depending on the presence or absence of portal vein thrombosis. PMID- 15269039 TI - Comprehensive analysis of hypervascular liver lesions using 16-MDCT and advanced image processing. PMID- 15269040 TI - False-negative rate of abdominal sonography for detecting hepatocellular carcinoma in patients with hepatitis B and elevated serum alpha-fetoprotein levels. AB - OBJECTIVE: Routine screening for hepatocellular carcinoma among chronic carriers of hepatitis B virus using a combination of abdominal sonography and serum alpha fetoprotein levels is widely practiced. Negative results on an abdominal sonogram generally indicate the absence of hepatocellular carcinoma despite the elevation of alpha-fetoprotein levels, but the false-negative rate of abdominal sonography has not been established prospectively. SUBJECTS AND METHODS: In our screening program, we routinely investigated patients with Lipiodol (iodized oil) CT when they presented with alpha-fetoprotein levels above 20 ng/mL or a focal lesion as depicted on abdominal sonography. Lipiodol CT comprised a hepatic angiogram with injection of Lipiodol selectively in the hepatic arteries, followed by an unenhanced CT scan 10 days later. Positive findings on Lipiodol CT were confirmed histologically by biopsy or surgical resection. We defined false-negative as histologic diagnosis of hepatocellular carcinoma within 3 months of normal findings on screening abdominal sonography. RESULTS: One hundred three patients with elevated alpha-fetoprotein levels were investigated with Lipiodol CT within 2 months of abdominal sonography. Of these, three of 70 patients with negative abdominal sonography had histologically confirmed hepatocellular carcinoma. Thus, abdominal sonography has a false-negative rate of 4.3%. Lipiodol CT is associated with a significant false-positive rate of 43.7%. The sensitivity, specificity, and positive predictive value of abdominal sonography for early detection of hepatocellular carcinoma among hepatitis B virus carriers with elevated alpha fetoprotein levels was 85.7%, 81.7%, and 54.5%, respectively. CONCLUSION: Negative results on a screening abdominal sonogram among hepatitis B virus carriers with elevated alpha-fetoprotein levels does not rule out the presence of small hepatocellular carcinoma. Routine use of Lipiodol CT as a supplementary screening tool is not recommended. PMID- 15269041 TI - Transient hepatic attenuation differences. PMID- 15269042 TI - Effect of different pharmacologic and chemical agents on the integrity of hydatid cyst membranes. AB - OBJECTIVE: We performed an in vitro investigation of the effects of widely used scolicidal and sclerosing agents, as well as some pharmacologic products, on the integrity of the membrane of hydatid cysts. MATERIALS AND METHODS: Two milliliters each of 22 agents, 2 mL of clear fluid, and one piece of hydatid cyst membrane were put into bottles. The hydatid cyst membranes were evaluated by visual observation and manual palpation. Visual examination of the bottles was performed daily for 7 days, and observations of membrane changes, including translucency, destruction, swelling, and melting, were recorded. Manual evaluation was done on the seventh day by finger examination, and membrane fragility was scored. RESULTS: The hydatid cyst membrane was completely melted in a few minutes in a 2.5% solution of sodium hypochlorite and in 1 hr by a 0.1% sodium hypochlorite solution. The integrity of the hydatid cyst membrane was preserved in alcohol, acetone, glutaraldehyde, albendazole, acetylsalicylic acid, formaldehyde, lidocaine, hydrochloric acid, ammonia, pancreatin, Betadine, methylene blue, and isotonic saline samples. The membranes in the metronidazole and hypertonic saline solutions were not damaged but showed significantly increased fragility. The membranes in levamisole and piperazine hexahydrate became translucent and showed moderate fragility. CONCLUSION: None of the agents that are used in clinical practice had important effects on the dissolution of hydatid cyst membranes. However, sodium hypochlorite solutions completely melted the hydatid cyst membranes. Because the use of this agent on living tissue is limited, further study is needed to investigate its clinical use. PMID- 15269043 TI - CT angiography for evaluation of living renal donors: comparison of four reconstruction methods. AB - OBJECTIVE: We sought to compare various reconstruction methods for CT angiographic images in evaluating living renal donors. MATERIALS AND METHODS: In 76 patients who underwent donor nephrectomy, vascular phase CT data were obtained using an MDCT scanner (detector array, 1.25 mm x 4; beam pitch, 1.5). Two radiologists independently reconstructed CT angiographic images using thick-slab volume rendering, thick-slab maximum intensity projection (MIP), sliding thin slab volume rendering, and sliding thin-slab MIP. The radiologists counted the number of renal arteries, early branching arteries, and renal veins. We compared the accuracy rates for the detection of vessels achieved with the four types of reconstructed images, using the surgical findings as the gold standard. Agreement between the two observers and between the surgical and CT angiographic findings was evaluated. RESULTS: The sensitivity for detecting the supernumerary artery was significantly greater with sliding thin-slab volume rendering and sliding thin-slab MIP (97%) than with thick-slab volume rendering (59%) (p = 0.039). No significant difference between the other comparison pairs of reconstruction methods was found. The interobserver agreement for detecting supernumerary and early branching arteries with sliding thin-slab volume rendering and MIP was excellent (kappa = 0.820-0.859) and good for renal veins (kappa = 0.698-0.724), whereas the agreement of thick-slab volume rendering and MIP was good for arteries (kappa = 0.706-0.791) and moderate for veins (kappa = 0.443-0.579). The agreement between CT angiographic reconstructed images and surgical findings for detection of vessels was better with sliding thin-slab volume rendering and MIP (kappa = 0.793-1.000) than in thick-slab volume rendering and MIP (kappa = 0.306 0.613). CONCLUSION: For CT angiographic evaluation of living renal donors, sliding thin-slab reconstruction is superior to thick-slab reconstruction. PMID- 15269044 TI - Impact of FDG PET on defining the extent of disease and on the treatment of patients with recurrent or metastatic breast cancer. AB - OBJECTIVE: Our aim was to evaluate the impact of FDG PET on defining the extent of disease and on the treatment of patients with advanced breast cancer. MATERIALS AND METHODS: The medical records of 125 consecutive patients with recurrent or metastatic breast cancer referred for FDG PET from January 1998 through May 2002 were retrospectively reviewed. The rationale for FDG PET referral and the impact of FDG PET on subsequent treatment decisions for patients were determined by chart review. The impact of FDG PET on defining the extent of disease was determined by comparing the FDG PET interpretation at the time of the examination with findings from conventional imaging (CI) performed before FDG PET. FDG PET results were confirmed in nearly half (n = 61) of the patients by histopathology (n = 23) or follow-up imaging (n = 38; mean follow-up interval, 21.3 months). RESULTS: Patients were referred for FDG PET for the following reasons: evaluation of disease response or viability after therapy (n = 43 [35%]), local recurrence, with intent of aggressive local treatment (n = 39 [31%]), equivocal findings on CI (n = 25 [20%]), evaluation of disease extent in patients with known metastases (n = 13 [10%]), and elevated tumor markers with unknown disease site (n = 5 [4%]). Compared with CI findings, the extent of disease increased in 54 (43%), did not change in 41 (33%), and decreased in 30 (24%) of 125 patients using FDG PET. Results of FDG PET altered the therapeutic plan in 40 (32%), directly helped to support the therapeutic plan in 34 (27%), and did not change the plan devised before FDG PET in 51 (41%) of 125 patients. FDG PET altered therapy most frequently in the patients suspected of having locoregional recurrence and in those being evaluated for treatment response versus other referral categories (p = 0.04). For patients with confirmation of FDG PET findings, the sensitivity, specificity, and accuracy of FDG PET were 94%, 91%, and 92%, respectively. CONCLUSION: FDG PET contributes significantly to defining the extent of disease and deciding on treatment of patients with advanced breast cancer. PMID- 15269046 TI - Oropharyngeal teratoma: prenatal diagnosis and assessment using sonography, MRI, and CT with management by ex utero intrapartum treatment procedure. PMID- 15269045 TI - Optimization of fetal weight estimates using MRI: comparison of acquisitions. AB - OBJECTIVE: The purpose of this study was to determine whether differences are seen in calculation of fetal weight using 5-mm sagittal, 3-mm coronal, and 8-mm axial MRI acquisitions compared with term birth weight and contemporaneous sonography. MATERIALS AND METHODS: Fetal volume measurements were obtained from MRI acquisitions as follows: 5-mm sagittal (2 acquisitions), 3-mm coronal (2 acquisitions), and 8-mm axial (1 acquisition). A 90-sec single-shot fast spin echo sequence was used. MRI and sonographic studies for fetal weight estimates were performed within 3 hr of term delivery. MRI calculation was based on the equation 0.12 + 1.031 x fetal volume (fetal area x slice thickness) (mL) = MRI fetal weight (kg). The sonographic fetal weight estimate was calculated using the Hadlock formula. MRI and sonographic calculations were compared with birth weight. Concordance coefficient analysis was performed. RESULTS: Thirty-five retrospective fetal calculations were performed. Concordance coefficients, gram weight means and standard deviations (mean +/- SD) between birth weight and MRI acquisitions were as follows: 8-mm axial, 0.91 (3,554 +/- 431 g); 3-mm coronal, 0.84 (3,752 +/- 578 g); and 5-mm sagittal, 0.83 (3,685 +/- 567 g), compared with 0.78 (3,518 +/- 332 g) for sonography. The MRI axial concordance coefficient was significantly different from that of the sonographic estimates (p = 0.05). MRI axial concordance coefficient was not statistically different from that of the MRI coronal concordance coefficient (p = 0.22) or the MRI sagittal concordance coefficient (p = 0.19). CONCLUSION: Calculated weights from a 90-sec single-shot fast spin-echo sequence MR acquisition with 8-mm-thick slices in the axial plane at term are better than sonographic estimates. PMID- 15269047 TI - Redefinition of multiple sclerosis plaque size using diffusion tensor MRI. AB - OBJECTIVE: We used diffusion tensor MRI to redefine the size of multiple sclerosis (MS) plaques on fractional anisotropy (FA) maps. MATERIALS AND METHODS: Thirty-six white matter (WM) plaques were identified in 20 patients with MS. Plaque FA was measured by placing regions of interest (ROIs) on plaques on diffusion tensor images. We compared FA values in identical mirror-image ROIs placed on normal-appearing WM in the contralateral hemisphere. This comparison showed a mean decrease in FA of 41% in plaques, serving as the threshold for outlining abnormal regions in normal-appearing WM surrounding plaques. ROIs were placed around each plaque and FA values were compared with those in the mirror image ROIs. Combined areas of perilesional normal-appearing WM with 40% or more FA reduction plus plaque were compared with the areas of abnormality on T2 weighted images using a paired Student's t test. A p value of 0.05 or less was considered significant. RESULTS: Mean plaque area was 60 mm(2) (range, 15-103 mm(2)), mean plaque FA was 0.251 (range, 0.133-0.436), and mean FA of contralateral normal-appearing WM was 0.429 (range, 0.204-0.712). Applying a threshold of 40% FA reduction, mean combined area of abnormal WM (including plaque seen on T2-weighted sequences) was 87 mm(2) (range, 30-251 mm(2)) or 145% of the mean plaque area that was seen on T2-weighted images (p < 0.001). CONCLUSION: Using an operator-defined threshold of abnormal FA values based on plaque anisotropy characteristics, we saw a statistically significant increase in plaque size. PMID- 15269048 TI - Trilateral retinoblastoma: clinical and radiologic progression. AB - OBJECTIVE: The purpose of this study was to assess the clinical and radiologic features of tumor progression in children with trilateral retinoblastoma. MATERIALS AND METHODS: Clinical records of eight children with trilateral retinoblastoma were reviewed for the patient's age at the time of diagnosis of the ocular tumor, time interval from diagnosis of ocular retinoblastoma to discovery of the intracranial tumor, time interval from diagnosis of retinoblastoma to death, and time interval from diagnosis of the intracranial tumor to death. CT or MRI studies were reviewed for the appearance of the primary intracranial neoplasm, intracranial metastases, and spinal metastases. RESULTS: The mean age of the patients at diagnosis of bilateral retinoblastoma was 4.5 months, and the mean age at diagnosis of the intracranial midline tumor was 26 months. The mean interval from the time of diagnosis of retinoblastoma to discovery of the intracranial tumor was 21.5 months. Two children had spinal leptomeningeal metastases at the time of discovery of the midline intracranial mass although no intracranial metastases were seen on imaging. In the other children, intracranial and spinal leptomeningeal metastases frequently developed within months of the diagnosis of retinoblastoma despite lack of progression in the midline intracranial lesion. Six children died of leptomeningeal spread of tumor. The mean interval from diagnosis of the ocular tumor to death was 46 months and from diagnosis of the intracranial tumor to death was 17 months. One child developed metastatic retinoblastoma in the ulna 10 years after the diagnosis of the intracranial tumor. CONCLUSION: Children typically died of leptomeningeal tumor dissemination despite lack of progression in the midline intracranial mass. Effective treatment of trilateral retinoblastoma may require close evaluation of these children for leptomeningeal dissemination. PMID- 15269049 TI - Model to quantify lymph node enhancement on indirect sonographic lymphography. AB - OBJECTIVE: Our goal was to develop a reliable technique that has minimal operator dependence for quantifying lymph node enhancement to test and optimize new sonography contrast formulations. MATERIALS AND METHODS: Twenty healthy rabbits were studied using five agents, labeled A-G. Agents D and E were the same agent and agents F and G were Imagent, studied blindly to test reproducibility. One milliliter of contrast agent was injected into each hind footpad. A 13-MHz transducer was fixed over the popliteal node, which was imaged at a 4.8-MHz central transmit frequency using phase-inversion technology at 100% power and one frame per second. Immediately after each injection, the footpad was massaged 12 times for 30 sec each time and then imaged after each massage to assess the number of times the node could be refilled from each injection. Lymph node video intensity was measured, and the degree of enhancement was evaluated using analysis of variance with the massage number and the agent used as independent variables. RESULTS: Lymph node enhancement was observed after the first massage with all agents. Degree of enhancement was least with agents A and B, intermediate with agents D and F, and greatest with agent C. Agent A was effective after the first two massages, agent B after the first four, agent C after all 12, agent D after the first eight, and agent F after the first nine. Performance of agents D and F was similar to that of their duplicates, E and G. CONCLUSION: We established a reproducible technique to quantify lymph node enhancement that can distinguish between different agents. The differences in performance suggest that it is possible to optimize agent formulation for indirect sonographic lymphography. PMID- 15269050 TI - Impact of fusion of indium-111 capromab pendetide volume data sets with those from MRI or CT in patients with recurrent prostate cancer. AB - OBJECTIVE: Our goal was to evaluate the impact of image fusion on the interpretation of indium-111 Prosta-Scint SPECT scans. MATERIALS AND METHODS: Sixty-seven consecutive patients referred for rising prostate-specific antigen (PSA) levels after initial therapy for primary prostate cancer underwent SPECT 96 hr after infusion of (111)In Prosta-Scint, with simultaneous technetium-99m blood pool imaging. Volume data sets from the SPECT scans were then fused with those from CT and MR images of the pelvis using a 3D landmark-based warping program. The SPECT scans were initially interpreted without benefit of MRI or CT fusion. The fused Prosta-Scint MRI-CT volumes were reevaluated by a nuclear radiologist and an MRI radiologist. Independent reviews before and after fusion were available in these patients. Validation of results after fusion was performed through correlation with PSA changes after radiation therapy. RESULTS: Six patients with sites that could not be evaluated and three without their original Prosta-Scint scanning reports were excluded; thus, 58 patients were studied clinically. Seventy-four of 161 prefusion-positive sites were found to be negative after fusion. These 74 sites subsequently were identified primarily as showing bowel, vessel, or marrow uptake after fusion. In two patients, nodal disease was identified although the review before perfusion indicated none. Twenty-five patients previously thought to have nodal disease appeared to have only local disease after fusion. After local radiation therapy, PSA levels decreased in 12 of 25 patients, increased in five, and were unavailable in eight. CONCLUSION: Although Prosta-Scint SPECT alone can help in the proper management of recurrent prostate cancer, fusion with MRI-CT of the pelvis can improve the specificity of the examination. PMID- 15269051 TI - Diffuse splenic metastases from seminoma visualized on FDG PET. PMID- 15269052 TI - A simple method for extracting DICOM images from a magnetooptic disk. AB - OBJECTIVE: Our objective was to develop a simple and easy-to-use method to extract DICOM images from magnetooptic (MO) disks to the computer desktop for research purposes. CONCLUSION: The method we developed allows users to extract DICOM images directly from MO disks to a PC desktop. The hardware component that we used is commercially available and is plug-and-play. The system is lower in cost than a clinical workstation. Users do not need to have special computer skills to use our method. DICOM images can be transferred directly from the MO disks to computer desktop folders using drag-and-drop. In our implementation, we store the DICOM files in a shared folder in our hospital network, so users can access the data from their office or research computers. PMID- 15269053 TI - MyFreePACS: a free web-based radiology image storage and viewing tool. AB - OBJECTIVE: We developed an easy-to-use method for central storage and subsequent viewing of radiology images for use on any PC equipped with Internet Explorer. CONCLUSION: We developed MyFreePACS, a program that uses a DICOM server to receive and store images and transmit them over the Web to the MyFreePACS Web client. The MyFreePACS Web client is a Web page that uses an ActiveX control for viewing and manipulating images. The client contains many of the tools found in modern image viewing stations including 3D localization and multiplanar reformation. The system is built entirely with free components and is freely available for download and installation from the Web at www.myfreepacs.com. PMID- 15269054 TI - A comment on CT-guided radiofrequency thermal ablation of osteoid osteoma. PMID- 15269055 TI - Adjunct sonography and not screening in cancer detection. PMID- 15269056 TI - Percutaneous treatment of biliary stones: cost-effective or just effective? PMID- 15269057 TI - Tattoos and MRI. PMID- 15269058 TI - Open low-field-strength MRI of the shoulder is not so bad. PMID- 15269059 TI - Community child health in crisis. PMID- 15269060 TI - Community paediatrics in crisis. PMID- 15269063 TI - Crossed fused ectopic left kidney. PMID- 15269061 TI - Is nutritional rickets returning? PMID- 15269062 TI - Substance abuse by children and young people. PMID- 15269065 TI - Epilepsy related mortality. PMID- 15269066 TI - Are sleep problems under-recognised in general practice? AB - AIMS: To evaluate the frequency of sleep problems in Australian children aged 4.5 16.5 years, and to determine whether the frequency of sleep problems on questionnaire predicts the reporting of sleep problems at consultation. METHODS: Parents of 361 children (aged 4.5-16.5 years) attending their general practitioner for "sick" visits were asked to assess their child's sleep over the previous six months using the Sleep Disturbance Scale for Children, from which six sleep "disorder" factors and a total sleep problem score were obtained. RESULTS: The percentage of children with a total sleep problem score indicative of clinical significance (T score >70 or >95th centile) was 24.6% (89/361). Despite this high frequency, parents only addressed sleep problems in 4.1% (13/317) of cases and reported that GPs discussed sleep problems in 7.9% (25/317) of cases. Of the 79 children who reported total sleep problem T scores in the clinical range, only 13.9% (11/79) discussed sleep with their general practitioner within the previous 12 months. Regression analyses revealed an age related decrease in problems with sleep-wake transition and sleep related obstructive breathing; sleep hyperhydrosis, initiating and maintaining sleep, and excessive daytime sleepiness did not significantly decrease with age. No significant gender differences were observed. CONCLUSIONS: Results suggest that chronic sleep problems in Australian children are significantly under-reported by parents during general practice consultations despite a relatively high frequency across all age groups. Given the impact on children and families, there is a need for increased awareness of children's sleep problems in the community and for these to be more actively addressed at consultation. PMID- 15269067 TI - Monitoring growth in asthmatic children treated with high dose inhaled glucocorticoids does not predict adrenal suppression. AB - AIMS: To determine whether routine outpatient monitoring of growth predicts adrenal suppression in prepubertal children treated with high dose inhaled glucocorticoid. METHODS: Observational study of 35 prepubertal children (aged 4 10 years) treated with at least 1000 microg/day of inhaled budesonide or equivalent potency glucocorticoid for at least six months. Main outcome measures were: changes in HtSDS over 6 and 12 month periods preceding adrenal function testing, and increment and peak cortisol after stimulation by low dose tetracosactrin test. Adrenal suppression was defined as a peak cortisol < or =500 nmol/l. RESULTS: The areas under the receiver operator characteristic curves for a decrease in HtSDS as a predictor of adrenal insufficiency 6 and 12 months prior to adrenal testing were 0.50 (SE 0.10) and 0.59 (SE 0.10). Prediction values of an HtSDS change of -0.5 for adrenal insufficiency at 12 months prior to testing were: sensitivity 13%, specificity 95%, and positive likelihood ratio of 2.4. Peak cortisol reached correlated poorly with change in HtSDS (rho = 0.23, p = 0.19 at 6 months; rho = 0.33, p = 0.06 at 12 months). CONCLUSIONS: Monitoring growth does not enable prediction of which children treated with high dose inhaled glucocorticoids are at risk of potentially serious adrenal suppression. Both growth and adrenal function should be monitored in patients on high dose inhaled glucocorticoids. Further research is required to determine the optimal frequency of monitoring adrenal function. PMID- 15269068 TI - Nurse management of intractable functional constipation: a randomised controlled trial. AB - AIMS: To evaluate the effectiveness of a nurse led clinic (NLC) compared with a consultant led paediatric gastroenterology clinic (PGC) in the management of chronic constipation. METHODS: Children (age 1-15 years) with functional constipation were randomised following a detailed medical assessment to follow up in either the NLC or PGC. An escalating algorithm of treatment was used as the basis of management in both the NLC and PGC. Main outcome measures were: time to cure at last visit or later confirmed by telephone; time to cure at last visit; and time to prematurely leaving the study. RESULTS: A total of 102 children were recruited, of whom 52 were randomly assigned to NLC and 50 to PGC. Outcome assessment showed that 34 children in the NLC and 25 children in the PGC were confirmed cured at their last visit or later confirmed by telephone. The median time to cure was 18.0 months in the NLC and 23.2 months in the PGC. The probability of being cured was estimated as 33% higher in the NLC compared to PGC (hazard ratio 1.33). Attending the NLC hastened time to cure by an estimated 18.4%. CONCLUSION: Children who attend an NLC are equally as, if not more likely to be cured of intractable constipation, than those attending a PGC and on average their cure will occur sooner. Results suggest that an NLC can significantly improve follow up for children with intractable constipation and highlight the important role for clinic nurse specialists in management of children with gastrointestinal disease. PMID- 15269069 TI - Prognosis of constipation: clinical factors and colonic transit time. AB - BACKGROUND: Measurement of colonic transit time (CTT) is sometimes used in the evaluation of patients with chronic constipation. AIM: To investigate the relation between symptoms and CTT, and to assess the importance of symptoms and CTT in predicting outcome. METHODS: Between 1995 and 2000, 169 consecutive patients (median age 8.4 years, 65% boys) fulfilling the criteria for constipation were enrolled. During the intervention and follow up period, all kept a diary to record symptoms. CTT was measured at entry to the study. RESULTS: At entry, defecation frequency was lower in girls than in boys, while the frequency of encopresis episodes was higher in boys. CTT values were significantly higher in those with a low defecation frequency (< or =1/week) and a high frequency of encopresis (> or =2/day). However, 50% had CTT values within the normal range. Successful outcome occurred more often in those with a rectal impaction. CTT results <100 hours were not predictive of outcome. However, those with CTT >100 hours were less likely to have had a successful outcome. CONCLUSION: The presence of a rectal impaction at presentation is associated with a better outcome at one year. A CTT >100 hours is associated with a poor outcome at one year. PMID- 15269070 TI - Does weaning influence growth and health up to 18 months? AB - BACKGROUND: National and international recommendations for the age of introducing solid foods (weaning) are founded on insufficient evidence and little is known about the short and medium term consequences associated with early or late weaning. AIMS AND METHODS: Data from over 1600 infants from five prospective randomised trials conducted in the UK between 1993 and 1997 were used to determine the influence of weaning < or =12 weeks or >12 weeks on growth and health outcomes (diarrhoea and vomiting, lower respiratory chest infections, atopy, sleep patterns) up to 18 months post-term, in term appropriate for gestational age (AGA), term small for gestational age (SGA), and preterm infants. RESULTS: Term infants weaned < or =12 weeks were heavier at 12 weeks of age than those weaned after 12 weeks, but showed slower gain in weight, length, and head circumference between 12 weeks and 18 months than those weaned after 12 weeks; by 18 months, there were no significant differences in size between the two groups. A similar pattern was seen in preterm infants. Breast fed term infants were more likely to be sleeping though the night at 9 months if they were weaned before 12 weeks. No weaning effects or interactions were observed for health outcomes. CONCLUSIONS: We found little evidence that weaning before or after 12 weeks influences health outcomes up to 18 months. Early weaned infants were larger at 12 weeks than later weaned infants but the growth trajectories of the two groups "converged" by 18 months. These findings do not exclude the later emergence of programmed effects of weaning practices. PMID- 15269071 TI - Effectiveness of influenza vaccine for the prevention of asthma exacerbations. AB - BACKGROUND: There is a lack of clinical evidence that annual vaccination against influenza prevents asthma exacerbations in children. METHODS: Retrospective cohort study of 800 children with asthma, where one half did, and the other half did not receive the influenza vaccine. The two groups were compared with respect to clinic visits, emergency department (ED) visits, and hospitalisations for asthma. In multivariable analyses, adjustment was made for baseline asthma severity, prior utilisation of health services, receipt of vaccine in the previous year, and demographic variables. RESULTS: After adjusting for other variables, the vaccine group had a significantly increased risk of asthma related clinic visits and ED visits (odds ratios 3.4 and 1.9, respectively). CONCLUSION: This study failed to provide evidence that the influenza vaccine prevents paediatric asthma exacerbations. PMID- 15269072 TI - Fertility preservation in adolescent males with cancer in the United Kingdom: a survey of practice. PMID- 15269073 TI - Radionuclide bone scintigraphy in Engelmann-Camurati disease. PMID- 15269074 TI - Mandatory temperature monitoring in schools during SARS. AB - During the SARS outbreak, temperature monitoring was mandatory for all Singapore schoolchildren. None of the Singapore children with SARS were detected through school temperature screening. However, temperature monitoring procedures have a powerful psychological effect of reassuring parents and the public that schools are safe during a SARS outbreak. PMID- 15269075 TI - Should recombinant human growth hormone therapy be used in short small for gestational age children? AB - Short small for gestational age (SGA) children represent 20% of all children with short stature and therefore constitute a significant portion of the caseload in a growth clinic. The recent approval of recombinant human growth hormone (GH) for the treatment of short stature in SGA children by the European Union's Committee on Proprietary Medicinal Products offers a new licensed therapeutic option. This article examines the role of GH therapy in short SGA children with particular reference to selection of patients, effectiveness, safety, and its potential metabolic implications. PMID- 15269076 TI - Juvenile thyrotoxicosis; can we do better? AB - Thyrotoxicosis remains a frustrating condition for the young person, family, and health professionals involved. The associated symptoms do not always suggest thyroid disease and patients can be unwell for many months before the diagnosis is made. The antithyroid drug regimen used to treat children and adolescents with thyrotoxicosis varies from one unit to another and yet the potentially life threatening side effects and remission rates post-treatment may be related to the regimen used. Most patients with thyrotoxicosis will need many years of drug therapy if the thyroid gland is not removed surgically or destroyed by radioiodine. Even "definitive" treatment will typically necessitate thyroxine replacement for life. PMID- 15269077 TI - Febrile seizures: an update. AB - This review focuses on the latest knowledge and understanding of febrile seizures and outlines the more important issues in the management of children who present with an apparent "febrile seizure". It is not the remit of this paper to discuss the detailed management of febrile seizures. Throughout this review, the words "partial" and "focal" will be used interchangeably and the term "febrile seizure" (FS) will be used, reflecting the proposed changes in the terminology of seizures and epilepsies.1 PMID- 15269078 TI - Twenty year surveillance of invasive pneumococcal disease in Nottingham: serogroups responsible and implications for immunisation. AB - AIMS: To evaluate the incidence, spectrum of clinical manifestations, and outcome of invasive pneumococcal disease (IPD) in children. To determine the major serogroups of Streptococcus pneumoniae responsible for invasive disease and the potential coverage by the new pneumococcal conjugate vaccines. METHODS: Analysis of prospectively recorded information of all children admitted to two teaching hospitals in Nottingham with IPD between January 1980 and December 1999. RESULTS: A total of 266 episodes of IPD in children were identified; 103 (39%) were aged <1 year and 160 (60%) <2 years. Major clinical presentations were meningitis in 86 (32%), pneumonia in 82 (31%), and bacteraemia without an obvious focus in 80 (30%). The age specific mean annual incidence rates of IPD overall among children aged <1, <2, and <5 years were 47.1, 37.8, and 20 per 100 000 population, respectively. Mortality rates for children with meningitis and non-meningitic infection were 20% and 7%, respectively. Neurological sequelae following meningitis were documented in 16 (26%) of the 61 survivors assessed. The potential coverage rates in children between the ages of 6 months and 5 years are 84% by the 7-valent, 91% by the 9-valent, and 95% by the 11-valent conjugate vaccines. CONCLUSION: This study indicates that inclusion of a pneumococcal conjugate vaccine in the primary immunisation programme in the UK would have a considerable effect on the mortality and morbidity associated with IPD. PMID- 15269079 TI - The implications of NICE guidelines on the management of children presenting with head injury. AB - BACKGROUND: NICE guidelines for the management of head injury were published in June 2003. Their recommendations differ markedly from previous guidelines published by the Royal College of Surgeons (RCS). In place of skull radiography and admission, computed tomography (CT) is advocated. The impact of these guidelines on service provision in the UK is unknown. METHODS: Data on all clinical correlates of children presenting with any severity of head injury was collected in three hospitals in the northwest of England. The current skull radiograph (SXR), CT scan, and admission rates were determined. The rates of SXR, CT scan, and admission that should have occurred when following either the RCS or NICE guidelines were then determined. RESULTS: Data from 10 965 patients who attended three hospitals between February 2000 and August 2002 was studied. Twenty five per cent of patients received a SXR, 0.9% a CT scan, and 3.7% were admitted. Strict adherence to the RCS guidelines would have resulted in a 50% SXR rate, a 1.6% CT scan rate, and a 7.1% admission rate. Adherence to NICE guidelines would result in a 0.3% SXR rate, an 8.7% CT scan rate, and a 1.4% admission rate, although the CT rate would drop to 6.3% if vomiting three or more times in the under 12s was used instead of more than one vomit. CONCLUSIONS: The new NICE guidelines do not increase the workload caused by patients attending with head injury but they move their management from the observation ward to the radiology department. PMID- 15269080 TI - Hospitalisation for gastroenteritis in Western Australia. AB - AIMS: To document gastroenteritis hospitalisations of the 1995-96 cohort of infants born in Western Australia to mid-2002, and to assess factors associated with their hospitalisations and readmissions. METHODS: Retrospective analysis of the State's hospitalisation data, Midwives' Notification of Births data, the Australian Bureau of Statistics mortality data and clinical and demographic information. RESULTS: Aboriginal infants were hospitalised for gastroenteritis eight times more frequently than their non-Aboriginal peers, and were readmitted more frequently and sooner for diarrhoeal illnesses than the other group. They also stayed in hospital for twice as long and many Aboriginal patients were hospitalised on numerous occasions. Hospitalisation rates were higher in remote areas and were significantly associated with co-morbidities such as undernutrition, anaemia, co-existing infections, and intestinal carbohydrate intolerance. CONCLUSIONS: Gastroenteritis is very prevalent in Australian Aboriginal infants and children and is a major cause of their hospitalisation in Western Australia. It is often associated with undernutrition, anaemia, intestinal parasitic infestations, other infections, intestinal carbohydrate intolerance, and, in some instances, with low birth weight. This is often due to unhygienic living conditions and behaviours and presents major challenges to public health, health promotion, and clinical personnel, particularly paediatric services. Childhood diarrhoeal diseases occur commonly in other indigenous groups but have not received the attention that they deserve. PMID- 15269081 TI - An improved urine collection pad method: a randomised clinical trial. AB - AIM: To evaluate a modified urine collection pad (UCP) method for its ability to reduce heavy mixed growth bacterial contamination of UCP samples in young children with suspected urinary tract infection (UTI). METHOD: Febrile children under 2 years of age were randomised to two UCP METHODS: the same UCP kept in the nappy until urine was passed (single UCP group), or the UCP replaced with a fresh one every 30 minutes until urine was passed (replaced UCP group). In both groups a moisture sensitive audio alarm was used to signal passage of urine. RESULTS: Eighty children were enrolled and a satisfactory sample was obtained in 68 (37 in the single UCP group and 31 in the replaced UCP group). In 12 children (15%), collection failed, mainly because of faecal soiling of the pad. UTI occurred in three children (4%). In the remaining 65 samples, heavy mixed growth (> 10(5) organisms/ml) occurred in 1/31 (3%) in the replaced UCP group compared with 10/35 (29%) in the single UCP group (p = 0.008). There were no adverse effects from the use of the moisture sensitive audio alarm. CONCLUSION: Changing the UCP every 30 minutes almost eliminates heavy mixed growth contamination of UCP samples and substantially increases the proportion of UCP results that confidently exclude UTI. This represents a simple and clinically important improvement to the UCP method which is reliable for diagnosing and excluding UTI in young children still in nappies. It has potential for use in outpatient clinics, in the primary healthcare setting, or at home. PMID- 15269082 TI - Coronary risk factors in Kawasaki disease treated with additional gammaglobulin. AB - AIMS: To assess the hypothesis that an additional intravenous gammaglobulin (IVGG) infusion, if administered early, may prevent coronary artery lesions (CAL) in patients with Kawasaki disease (KD) who do not respond to initial IVGG therapy. METHODS: Forty four KD patients (17 with CAL and 27 without CAL), treated with additional IVGG because of persistent or recrudescent fever after initial IVGG therapy, were studied. Main outcome measures were the presence of CAL by echocardiography and the number of febrile days before and after start of additional IVGG infusion (pre- and post-additional IVGG). RESULTS: In univariate analyses, risk factors for CAL were the number of febrile days pre-additional IVGG, the number of febrile days post-additional IVGG, the number of days that initial IVGG was divided over, the white blood cell count pre- and post additional IVGG, and the C reactive protein concentration pre-additional IVGG. In a multivariate analysis, the only independent risk factor was the number of febrile days pre-additional IVGG (> or =10 days; odds ratio 7.86; 95% CI 1.44 to 42.8; p = 0.02). CONCLUSIONS: Among KD patients with persistent or recrudescent fever after initial IVGG therapy, administration of additional IVGG before the first 10 febrile days was associated with a decreased prevalence of CAL, when compared with the prevalence in those who were retreated later. An additional IVGG infusion, if administered early, may prevent CAL in initial IVGG non responders. PMID- 15269084 TI - Angioinvasive aspergillosis. PMID- 15269083 TI - Presentation of vitamin D deficiency. AB - AIMS: To describe the various ways in which vitamin D deficiency presents in children in selected districts of London and to identify which factors, if any, determine the mode of presentation. METHODS: Retrospective review of patients presenting to Newham General, Royal London, Great Ormond Street, and King's College Hospitals between 1996 and 2001 with either hypocalcaemia or rickets caused by vitamin D deficiency. Children with plasma 25-hydroxyvitamin D levels <25 nmol/l (10 ng/ml) were considered to have vitamin D deficiency. RESULTS: Sixty five children, mostly from Black or Asian ethnic minority groups, were identified, 29 of whom had hypocalcaemic symptoms. Seventeen of these had no radiological evidence of rickets. The remainder (48 children) had radiological evidence of rickets with or without clinical signs. Symptoms and signs reverted to normal in all cases with vitamin D supplementation. All children who presented with symptomatic hypocalcaemia were aged either <3 or >10 years. There was a strong correlation between age at presentation and population growth velocity reference data. CONCLUSIONS: Rickets remains a problem in the UK especially in "at risk" ethnic minority groups. Symptomatic hypocalcaemia is an important, but under-recognised presenting feature. Growth rate is likely to be an important factor in determining the mode of presentation. Unexplained hypocalcaemia should be attributed to vitamin D deficiency in "at risk" ethnic minority groups until proved otherwise. PMID- 15269086 TI - The use of interferon beta in relapsing-remitting multiple sclerosis. AB - The use of interferon beta-1a to treat multiple sclerosis in a child of 7 years of age is discussed. To date, there is only one other published report of the use of interferon beta in a child as young as this. One year after commencing treatment she had shown significant clinical improvement, with a marked reduction in number of relapses. In her second year of treatment she suffered a major relapse from which she slowly recovered. PMID- 15269085 TI - Clinical and laboratory evaluation of compliance in acute lymphoblastic leukaemia. AB - AIM: To evaluate compliance in children with acute lymphoblastic leukaemia (ALL). METHODS: Compliance was assessed through specific interviews, annotations from medical charts, and erythrocytic determination of 6-mercaptopurine metabolites. RESULTS: A total of 39 patients who had concluded maintenance phase of chemotherapy were included in the study. Mothers were responsible for delivering 6-MP in 87% of cases. Thirty five interviewees said that medical prescription was well understood and that the main reason for non-compliance was forgetfulness. Non-compliance was detected through interviews (33.3% of the cases), reports from medical charts (30.7%), and drug determination (16.6%); 53.8% of children were found to be non-compliant. Non-compliance was significantly associated with chronic undernourishment. Although not statistically significant, there was a trend for the group of non-compliant children to be associated with low per capita family income. No significant associations of non-compliance with age at diagnosis, gender, parents' schooling level, number of family members, power consumption, and medians of absolute leucocyte or neutrophil blood counts were detected. A short follow up period precluded valid analysis on outcome. In the non-compliant group (n = 21), seven children relapsed, contrasting with three relapses in the compliant group (n = 18). CONCLUSIONS: Results suggest that non compliance is one of the mechanisms which underlies the adverse influence of socioeconomic factors on the outcome of children with ALL. Additional studies are necessary to confirm this hypothesis. Comprehensive approaches to the problem of non-compliance are urgently needed. PMID- 15269087 TI - Paediatric craniopharyngioma. PMID- 15269088 TI - Euroaspirations. PMID- 15269089 TI - Thrombocytopenia is predictive of lethality in severe childhood falciparum malaria. PMID- 15269090 TI - Judged by our legacy. PMID- 15269091 TI - Response to salbutamol by wheezy infants. PMID- 15269092 TI - The use of insulin pumps improves the metabolic control in children and adolescents with type 1 diabetes. PMID- 15269093 TI - Read the label carefully. PMID- 15269094 TI - Board senseless. PMID- 15269095 TI - Malignancy and mortality in people with coeliac disease: population based cohort study. AB - OBJECTIVE: To quantify the risks of malignancy and mortality in people with coeliac disease compared with the general population. DESIGN: Population based cohort study. SETTING: General practice research database. PARTICIPANTS: 4732 people with coeliac disease and 23,620 matched controls. MAIN OUTCOME MEASURES: Hazard ratios for malignancy and mortality. RESULTS: Of the 4732 people with coeliac disease, 134 (2.8%) had at least one malignancy and 237 (5.0%) died. The overall hazard ratios were: for any malignancy 1.29 (95% confidence interval 1.06 to 1.55), for mortality 1.31 (1.13 to 1.51), for gastrointestinal cancer 1.85 (1.22 to 2.81), for breast cancer 0.35 (0.17 to 0.72), for lung cancer 0.34 (0.13 to 0.95), and for lymphoproliferative disease 4.80 (2.71 to 8.50). The increased risk was primarily in the first year after diagnosis, with the risk for only lymphoproliferative disease remaining significantly raised thereafter. After excluding events in the year after diagnosis, the hazard ratio for malignancy was 1.10 (0.87 to 1.39) and for mortality was 1.17 (0.98 to 1.38), giving absolute excess rates of 6 and 17 per 10,000 person years, respectively. CONCLUSIONS: People with coeliac disease have modest increases in overall risks of malignancy and mortality. Most of this excess risk occurs in the year of follow up after diagnosis. People with coeliac disease also have a noticeably reduced risk of breast cancer. The mechanism of this merits further attention as it may provide insights into the cause of this common malignancy. PMID- 15269096 TI - Estrogen actions on follicle formation and early follicle development. AB - Estradiol-17beta (E(2)) affects late follicular development, whereas primordial follicle differentiation and early activation are believed to be independent of E(2). To test this hypothesis we compared numbers of primordial and primary follicles in wild-type and E(2)-deficient, aromatase knockout (ArKO) mice, and the immunohistochemical staining or mRNA expression of Mullerian inhibiting substance (MIS), Wilms tumor 1 (WT-1), and growth differentiation factor (GDF9), which are known to effect early follicular differentiation. Proliferating cell nuclear antigen (PCNA) staining was a marker of proliferative index. The effects of E(2) replacement for 3 wk in 7-wk-old ArKO and wild-type mice on these parameters were also tested. ArKO mice had reduced numbers of primordial and primary follicles compared with wild-type mice (63%, P < 0.001 and 60%, P = 0.062, respectively). This reduction was not corrected by E(2) treatment, suggesting that E(2) affects the initial formation or activation of primordial follicles. There was a significant increase in the diameters of the oocytes in primordial follicles of ArKO mice compared with mice of the wild type. There were no differences in the immunostaining of MIS, WT-1, and PCNA in primordial and primary follicles between wild-type and ArKO mice. The only difference was as a consequence of Sertoli and Leydig cells that develop in ovaries of ArKO mice. GDF9 mRNA expression was markedly increased in ArKO ovaries. E(2) treatment restored the ovarian follicular morphology in ArKO mice, and consequently the immunostaining patterns, but had no effect on early follicle numbers. In conclusion, E(2) has a role in controlling the size of the oocyte and primordial follicle pool in mice. PMID- 15269097 TI - Nuclear origin of aging-associated meiotic defects in senescence-accelerated mice. AB - Factors of both cytoplasmic and nuclear origin regulate metaphase chromosome alignment and spindle checkpoint during mitosis. Most aneuploidies associated with maternal aging are believed to derive from nondisjunction and meiotic errors, such as aberrations in spindle formation and chromosome alignment at meiosis I. Senescence-accelerated mice (SAM) exhibit aging-associated meiotic defects, specifically chromosome misalignments at meiosis I and II that resemble those found in human female aging. How maternal aging disrupts meiosis remains largely unexplained. Using germinal vesicle nuclear transfer, we found that aging associated misalignment of metaphase chromosomes is predominately associated with the nuclear factors in the SAM model. Cytoplasm of young hybrid B6C3F1 mouse oocytes could partly rescue aging-associated meiotic chromosome misalignment, whereas cytoplasm of young SAM was ineffective in preventing the meiotic defects of old SAM oocytes, which is indicative of a deficiency of SAM oocyte cytoplasm. Our results demonstrate that both nuclear and cytoplasmic factors contribute to the meiotic defects of the old SAM oocytes and that the nuclear compartment plays the predominant role in the etiology of aging-related meiotic defects. PMID- 15269098 TI - Real-time relationships in intraluteal release among Prostaglandin F2alpha, endothelin-1, and angiotensin II during spontaneous luteolysis in the cow. AB - It is well known that prostaglandin F(2alpha) (PGF(2alpha)) is a physiological luteolysine, and that its pulsatile release from the endometrium is a luteolytic signal in many species. There is now clear evidence that the vasoactive peptides endothelin-1 (ET-1) and angiotensin II (Ang II) interact with PGF(2alpha) in the luteolytic cascade during PGF(2alpha)-induced luteolysis in the cow. Thus, we investigated the local secretion of PGF(2alpha), ET-1, and Ang II in the corpus luteum (CL) and their real-time relationships during spontaneous luteolysis in the cow. For this purpose, an in vivo microdialysis system (MDS) implanted in the CL was utilized to observe local secretion changes within the CL microenvironment. Each CL of cyclic Holstein cows (n = 6) was surgically implanted with MDS capillary membranes (18 lines/6 cows) on Day 15 (estrus = Day 0) of the estrous cycle. The concentrations of PGF(2alpha), ET-1, Ang II, and progesterone (P) in the MDS samples were determined by enzyme immunoassays. The intraluteal PGF(2alpha) secretion slightly increased from 12 h after the onset of luteolysis (0 h) and drastically increased (by about 300%) from 24 h. Intraluteal ET-1 secretion increased from 12 h. Intraluteal Ang II secretion was elevated from 0 h and was maintained at high levels (about 180%) toward estrus. In each MDS lines (in the same microenvironment) within the regressing CL, the local releasing profiles of PGF(2alpha), ET-1, and Ang II CL positively correlated with each other (P < 0.05) at high proportions in 18 MDS lines (PGF(2alpha) vs. ET-1, 44.4%; PGF(2alpha) vs. Ang II, 55.6%; ET-1 vs. Ang II, 38.9%). In contrast, there was no clear relationship among these substances released into different MDS lines implanted in the same CL (with different microenvironments). In conclusion, we propose that the increase of PGF(2alpha), ET-1, and Ang II within the CL during luteolysis is a common phenomenon for both PGF(2alpha)-induced and spontaneous luteolysis. Moreover, this study illustrated the in vivo relationships in intraluteal release among PGF(2alpha), ET-1, and Ang II during spontaneous luteolysis in the cow. The data suggest that these vasoactive substances may interact with each other in a local positive feedback manner to activate their secretion in the regressing CL, thus accelerating and completing luteolysis. PMID- 15269099 TI - Urinary progesterone-induced blocking factor concentration is related to pregnancy outcome. AB - Peripheral lymphocytes from healthy pregnant women secrete a mediator protein named the progesterone-induced blocking factor (PIBF) that exerts an immunomodulatory function and contributes to the maintenance of pregnancy in mice. The gene coding for PIBF mRNA has been cloned and sequenced, and now the recombinant human protein is available. The aim of this study was to develop an ELISA test for determining PIBF concentrations in biological samples of pregnant women. We determined urinary PIBF concentrations of 86 healthy nonpregnant individuals and from almost 500 pregnant women by ELISA. During normal pregnancy, the concentration of PIBF continuously increased until the 37th gestational week and was followed by a sharp decrease after the 41st week of gestation. In pathological pregnancies, urinary PIBF levels failed to increase. The onset of labor was predictable on the basis of this test, whether it was term or preterm delivery. In urine of patients with preeclampsia, PIBF concentrations were significantly lower than in normal pregnancy and showed a correlation with the number of symptoms presented. These data, in line with previous in vivo findings, suggest that PIBF production is a characteristic feature of normal pregnancy, and determination of PIBF concentration in urine might be of use for the diagnosis of threatened premature pregnancy termination. PMID- 15269100 TI - Granzyme N, a novel granzyme, is expressed in spermatocytes and spermatids of the mouse testis. AB - We cloned a cDNA for a novel granzyme, granzyme N (Gzmn), from a mouse testes cDNA library. The testes contained two distinct species of Gzmn mRNA, one of which codes for a complete protein of 248 amino acids with three essential residues required for catalytic activity. The Gzmn mRNA was specifically expressed in the testes of adult mice. The Gzmn expression was found to initiate in the testes at 3 wk of age and to become more prominent as the animal reached sexual maturity. In situ hybridization analysis revealed that both spermatocytes and spermatids of the adult mouse testes express Gzmn mRNA. Consistent with these findings, the protein was immunohistochemically detected in the spermatocytes and spermatids, although some of the germ cells showed no positive staining. Gzmn was demonstrated to be a secretory and N-glycosylated protein that exists in two protein forms in the testes extract. In the cryptorchid testes, the expression of Gzmn transcript was drastically reduced on Postoperative Day 10, whereas the protein level was gradually decreased starting on Day 6. The local heating (43 degrees C, 20 min) of the testes did not change the Gzmn expression level at either 8 or 16 h after treatment. These results suggest that Gzmn is not involved in the process of germ cell apoptosis induced by heat shock, but that it may be involved in spermatogenesis in the mouse testes. PMID- 15269101 TI - Acute temporal regulation of placental vascular endothelial growth/permeability factor expression in baboons by estrogen. AB - Vascular endothelial growth/permeability factor (VEG/PF) has an established role in angiogenesis, however, the regulation of placental VEG/PF expression during primate pregnancy is incompletely understood. A temporal study was conducted in baboons to determine the effect of acute administration of estradiol on the expression of VEG/PF by cells of the villous placenta. VEG/PF mRNA levels were determined by reverse transcription-polymerase chain reaction in isolated placental cell fractions of baboons after acute i.v. and i.m. administration of estradiol. Within 2 h of estradiol treatment, VEG/PF mRNA (attomoles/ micrograms total RNA) increased within villous cytotrophoblasts to a level (mean +/- SEM, 12,612 +/- 2419) that was almost 2-fold greater (P < 0.05) than in untreated controls (6810 +/- 1368). Cytotrophoblast VEG/PF mRNA levels remained elevated (P < 0.01) 6 h after estradiol treatment (15,006 +/- 506), but were not different from controls 18 h after estradiol administration. VEG/ PF mRNA levels in whole villous tissue also were greater 6 h (12,667 +/- 2284, P < 0.05) and 18 h (16,080 +/- 3816, P < 0.01) after estradiol treatment than in untreated animals (3380 +/- 594). In contrast, VEG/PF mRNA levels in cells of the inner villous core were not altered by estradiol treatment. Expression of both the VEG/PF(121) and VEG/PF(165) mRNA species appeared to increase in the placenta 6 h after estradiol treatment of baboons. We propose that estrogen regulates VEG/PF expression within the placenta in a cell-specific manner, providing a paracrine system to promote vascularization of the villous placenta during the first half of primate pregnancy. PMID- 15269102 TI - Two isoforms of the leptin receptor are enhanced in pregnancy-specific tissues and soluble leptin receptor is enhanced in maternal serum with advancing gestation in the baboon. AB - Leptin is a polypeptide hormone produced by adipose and other endocrine tissues. Although it has been linked to receptor-mediated pathways that directly influence human conceptus development, mechanisms that regulate the leptin receptor in pregnancy-specific tissues remain unclear. Therefore, we assessed leptin-receptor ontogeny and regulation in the baboon (Papio sp.), a primate model for human pregnancy. Placentae, decidua, and amniochorion were collected from baboons in early (Days 54-63, n = 4), mid (Days 98-103, n = 4), and late (Days 159-165, n = 4) gestation. Regulation by estrogen was assessed by elimination of androgen precursors via removal of the fetus (fetectomy) at midgestation and collection of tissues in late gestation (n = 4; term, approximately 184 days). Maternal serum was sampled with advancing gestation, and the abundance of soluble leptin receptor (solLepR), a potential mediator of gestational hyperleptinemia, was determined. Two placental leptin-receptor isoforms (130 and 150 kDa) increased (P < 0.04 and P < 0.02, respectively) in abundance with advancing gestation. Similarly, the 130-kDa isoform increased approximately fourfold (P < 0.0025) in decidua and approximately 10-fold (P < 0.015) in amniochorion between early and late gestation. Following fetectomy, maternal serum estradiol levels declined approximately 85% (P < 0.03), and the 150-kDa placental leptin-receptor isoform was reduced by more than half (P < 0.002). Maternal serum solLepR concentrations were correlated with gestational age (r = 0.52, P < 0.01) and were unaffected by fetectomy. The presence of leptin-receptor isoforms in pregnancy-specific tissues further denoted leptin's potential to directly influence conceptus development, whereas the 130-kDa solLepR identified in maternal serum suggested a means to facilitate the hyperleptinemia typical of primate pregnancy. Although estrogen did not appear to be the principal regulator of solLepR, it and other factors linked to advancing gestation may be implicated in the regulation of leptin receptor synthesis. PMID- 15269103 TI - Meiosis-activating sterol synthesis in rat preovulatory follicle: is it involved in resumption of meiosis? AB - Meiosis-activating sterol (MAS) was shown to overcome the inhibitory effect of hypoxanthine on spontaneous maturation of mouse oocytes and was suggested to mediate the stimulation of meiosis by gonadotropins. Follicular fluid (FF)-MAS is synthesized by cytochrome P450 lanosterol 14alpha-demethylase (LDM). Follicular LDM was preferentially localized in oocytes by immunohistochemistry. Using [3H]acetate or R-[5-3H]mevalonate as precursors as well as high-performance liquid chromatographic and thin-layer chromatographic separation, we have measured the concentrations of de novo-synthesized lanosterol, FF-MAS, and cholesterol in rat graafian follicles, cumulus-oocyte complexes (COCs), and denuded oocytes (DOs) treated with LH, AY-9944 (an inhibitor of Delta14 reductase, which was anticipated to increase FF-MAS levels by inhibiting its metabolism), or both after 8 h of culture. In follicles, both LH and AY-9944 increased the accumulation of FF-MAS as compared to controls. In COCs, AY-9944 caused a marked increase in FF-MAS, but we were unable to detect accumulation of FF-MAS in DOs. Neither the endogenous increases in FF-MAS accumulation nor the addition of FF-MAS to the culture medium could overcome the inhibition on resumption of meiosis by phosphodiesterase inhibitors. Compared to LH-induced resumption of meiosis in follicles, that induced by AY-9944 was much delayed. These results call into question any role of FF-MAS as an obligatory mediator of LH activity on germinal vesicle breakdown. The discrepancy between the positive staining for LDM in oocytes and our inability to detect de novo synthesized FF MAS in DOs may relate to the sensitivity of the methodology employed and either the number of oocytes used or a deficiency in LDM synthetic activity in such oocytes. Further studies are required to confirm any of these alternatives. PMID- 15269104 TI - Cellular mechanisms and modulation of activin A- and transforming growth factor beta-mediated differentiation in cultured hen granulosa cells. AB - Undifferentiated granulosa cells from prehierarchal (6- to 8-mm-diameter) hen follicles express very low to undetectable levels of LH receptor (LH-R) mRNA, P450 cholesterol side chain cleavage (P450scc) enzyme activity, and steroidogenic acute regulatory (StAR) protein, and produce negligible progesterone, in vitro, following an acute (3-h) challenge with either FSH or LH. It has previously been established that culturing such cells with FSH for 18-20 h induces LH-R, P450scc, and StAR expression, which enables the initiation of progesterone production. The present studies were conducted to characterize the ability of activin and transforming growth factor (TGF) beta, both alone and in combination with FSH, to promote hen granulosa cell differentiation, in vitro. A 20-h culture of prehierarchal follicle granulosa cells with activin A or transforming growth factor beta (TGFbeta)1 increased LH-R mRNA levels compared with control cultured cells. Activin A and TGFbeta1 also promoted FSH-receptor (FSH-R) mRNA expression when combined with FSH treatment. Neither activin A nor TGFbeta1 alone stimulated progesterone production after 20 h culture. However, preculture with either factor for 20 h (to induce gonadotropin receptor mRNA expression) followed by a 3 h challenge with FSH or LH potentiated StAR expression and progesterone production compared with cells challenged with gonadotropin in the absence of activin A or TGFbeta1 preculture. Significantly, activation of the mitogen activated protein (MAP) kinase pathway with transforming growth factor alpha (TGFalpha) (monitored by Erk phosphorylation) blocked TGFbeta1-induced LH-R expression, and this effect was associated with the inhibition of Smad2 phosphorylation. We conclude that a primary differentiation-inducing action of activin A and TGFbeta1 on hen granulosa cells from prehierarchal follicles is directed toward LH-R expression. Enhanced LH-R levels subsequently sensitize granulosa cells to LH, which in turn promotes StAR plus P450scc expression and subsequently an increase in P4 production. Significantly, the finding that TGFbeta signaling is negatively regulated by MAP kinase signaling is proposed to represent a mechanism that prevents premature differentiation of granulosa cells. PMID- 15269105 TI - Functional characterization of the placental fusogenic membrane protein syncytin. AB - Syncytin is an envelope protein of the human endogenous retrovirus family W (HERV W). Syncytin is specifically expressed in the human placenta and mediates trophoblast cell fusion into the multinucleated syncytiotrophoblast layer. It is a polypeptide of 538 amino acids and is predicted to be posttranslationally cleaved into a surface (SU) subunit and a transmembrane (TM) subunit. Functional characterization of syncytin protein can aid understanding of the molecular mechanism underlying syncytin-mediated cell fusion. In this report, we studied the structure-function relationship of syncytin in 293T and HeLa cells transiently expressing wild-type syncytin or syncytin mutants generated by linker scanning and deletion mutagenesis. Of the 22 linker-inserted mutants, mutants InS51, InV139, InE156, InS493, InA506, and InL529 were fusogenic, suggesting that regions around amino acids S51, V139, and E156 in the SU subunit and S493, A506, and L529 in the cytoplasmic domain (CTM) of syncytin are flexible in conformation. Of the 17 deletion mutants, nine mutants with deletions in the region from amino acids 479 to 538 were fusogenic. The deletion mutant DelI480, containing only the first four amino acid residues in the cytoplasmic domain, had enhanced fusogenic activity in comparison with the wild-type. In addition, two heptad repeat regions (HRA and B) were defined in the TM subunit of syncytin. A peptide inhibitor derived from the C-terminal heptad repeat region (HRB) was shown to potently inhibit syncytin-mediated cell fusion. Our results suggest that the cytoplasmic domain of syncytin is not essential for syncytin-mediated fusion but may play a regulatory role, and an intramolecular interaction between HRA and B is involved in the fusion process. PMID- 15269106 TI - Impaired filtering of distracter stimuli by TE neurons following V4 and TEO lesions in macaques. AB - Directing attention to a behaviorally relevant visual stimulus can overcome the distracting effects of other nearby stimuli. Correspondingly, physiological studies indicate that attention serves to filter distracting stimuli from receptive fields (RFs) in several extrastriate areas. Moreover, a recent study demonstrated that lesions of extrastriate areas V4 and TEO produce impairments in attentional filtering. A critical remaining question concerns why lesions of ventral stream areas cause attentional filtering impairments. To address this question, we tested the effects of restricted area V4 and TEO lesions on both behavioral performance and the responses of downstream neurons in area TE. The lesions impaired behavioral discrimination thresholds and altered neuronal selectivity for target stimuli in the presence of distracters. With attention to the target, but in the absence of V4 and/or TEO inputs, TE neurons responded as though attentional inputs could no longer be used to filter distracters from their RFs. This presumably occurred because top-down attentional signals were no longer able to filter distracters from the RFs of the cells that provide TE with major input. Consistent with this interpretation, increasing the spatial separation between targets and distracters, such that they no longer fell within a typical V4 RF dimension, restored both behavioral performance and neuronal selectivity in the portion of TE RFs affected by the V4 lesion. PMID- 15269107 TI - The functional neuroanatomy of metrical stress evaluation of perceived and imagined spoken words. AB - We hypothesized that areas in the temporal lobe that have been implicated in the phonological processing of spoken words would also be activated during the generation and phonological processing of imagined speech. We tested this hypothesis using functional magnetic resonance imaging during a behaviorally controlled task of metrical stress evaluation. Subjects were presented with bisyllabic words and had to determine the alternation of strong and weak syllables. Thus, they were required to discriminate between weak-initial words and strong-initial words. In one condition, the stimuli were presented auditorily to the subjects (by headphones). In the other condition the stimuli were presented visually on a screen and subjects were asked to imagine hearing the word. Results showed activation of the supplementary motor area, inferior frontal gyrus (Broca's area) and insula in both conditions. In the superior temporal gyrus (STG) and in the superior temporal sulcus (STS) strong activation was observed during the auditory (perceptual) condition. However, a region located in the posterior part of the STS/STG also responded during the imagery condition. No activation of this same region of the STS was observed during a control condition which also involved processing of visually presented words, but which required a semantic decision from the subject. We suggest that processing of metrical stress, with or without auditory input, relies in part on cortical interface systems located in the posterior part of STS/STG. These results corroborate behavioral evidence regarding phonological loop involvement in auditory-verbal imagery. PMID- 15269108 TI - BDNF-induced white matter neuroprotection and stage-dependent neuronal survival following a neonatal excitotoxic challenge. AB - Excitotoxicity may be critical in the formation of brain lesions associated with cerebral palsy. When injected into the murine neopallium at postnatal day (P) 5, ibotenate (activating NMDA and metabotropic glutamate receptors) produces neuronal death and white matter cysts. Such white matter cysts resemble those seen in periventricular leukomalacia, a lesion evident in numerous human premature newborns. The goal of this study was to assess BDNF neuroprotection against neonatal excitotoxic lesions. Cortical and white matter lesions induced by ibotenate at P5 were reduced by BDNF by up to 36 and 60%, respectively. BDNF neuroprotection involved TrkB receptors, MAPK pathway and reduced apoptosis. Although BDNF did not prevent the initial appearance of white matter lesions, it promoted secondary decrease of the lesion size. BDNF neuroprotection at P5 was maximal against lesions induced by NMDA or ibotenate but was moderate against lesions produced by an AMPA-kainate agonist. Finally, BDNF exacerbated neuronal death produced by ibotenate at P0 through increased apoptosis and p75(NTR) receptors, while BDNF had no detectable effect on lesions induced at P10. Altogether, these data showed that BDNF neuroprotection against neonatal excitotoxicity is dependent upon the type of activated glutamate receptors, the lesion localization and the developmental stage. PMID- 15269109 TI - A contralateral preference in the lateral occipital area: sensory and attentional mechanisms. AB - Here we examined the level of the lateral occipital (LO) area within the processing stream of the ventral visual cortex. An important determinant of an area's level of processing is whether it codes visual elements on both sides of the visual field, as do higher visual areas, or prefers those in the contralateral visual field, as do early visual areas. The former would suggest that LO, on one side, combines bilateral visual elements into a whole, while the latter suggests that it codes only the parts of forms. We showed that LO has a relative preference for visual objects in the contralateral visual field. LO responses were influenced by attention. However, relative changes in LO activity caused by changes in object location were preserved even when attention was shifted away from the objects to moving random dot patterns on the opposite side. Our data offer a new view on LO as an intermediate, but not a high-level, visual area in which neurons are driven by visual input and spatial attention in a multiplicative fashion. PMID- 15269110 TI - Layer-specific production of nitric oxide during cortical circuit formation in postnatal mouse brain. AB - In the developing cerebral cortex, neuronal nitric oxide synthase (nNOS) is expressed abundantly, but temporarily. During the early postnatal stage, cortical neurons located in the multi-layered structure of the cortical plate start forming well-organized cortical circuits, but little is known about the molecular machinery for layer-specific circuit formation. To address the involvement of nitric oxide (NO), we utilized a new NO indicator (DAR-4M) and developed a protocol for the real-time imaging of NO produced in fresh cortical slices upon N methyl-D-aspartic acid stimulation. At postnatal day 0 (P0), NO production was restricted to the deep layers (layers V and VI) of the somatosensory cortex where transient synapses are formed. At P10, the production of NO was expanded to layer IV where large numbers of thalamocortical axons form synapses. The pattern of NO production could correspond to active sites for synaptic formation. This study is the first clear demonstration of NO production in the postnatal mouse neocortex. The findings presented may reflect a function of NO in relation to the layer specific development of neural circuits in the neocortex. PMID- 15269111 TI - Opposite effects of amphetamine self-administration experience on dendritic spines in the medial and orbital prefrontal cortex. AB - We studied the long-term effects of amphetamine self-administration experience (or sucrose reward training) on dendritic morphology (spine density) in nucleus accumbens (Nacc), medial (MPC) and orbital prefrontal cortex (OFC), and hippocampus (CA1 and dentate). Independent groups of rats were trained under a continuous schedule of reinforcement to nose-poke for infusions of amphetamine (0.125 mg/kg/inf) or to receive sucrose pellets during 2 h daily test sessions for 14-20 days. One month after the last training session, the brains were collected and processed for Golgi-Cox staining. We found that: (i) amphetamine self-administration experience selectively increased spine density on medium spiny neurons in the Nacc and on pyramidal neurons in the MPC; (ii) in contrast, amphetamine self-administration decreased spine density in the OFC, whereas sucrose-reward training increased spine density; and (iii) both amphetamine self administration and sucrose-reward experience increased spines in the CA1, but had no effect in the dentate gyrus. Thus, amphetamine self-administration experience produces long-lasting and regionally-selective morphological alterations in rat forebrain--alterations that may underlie some of the persistent psychomotor, cognitive and motivational consequences of chronic drug exposure. PMID- 15269112 TI - Neonatal NMDA receptor blockade disturbs neuronal migration in rat somatosensory cortex in vivo. AB - Glutamate plays an important role in the control of neuronal migration in the developing cerebral cortex. The present study describes changes in the structure and function of the cerebral cortex after transient blockade of N-methyl-D aspartate (NMDA) receptors during the late period of neuronal migration. Elvax slices containing the NMDA antagonist MK801 were placed over the somatosensory cortex of newborn rats and the drug was released over a period of 2-3 days. After survival times of 1 or 2 weeks, neuroanatomical and in vitro electrophysiological analyses revealed prominent structural and functional alterations in the cortical region underlying the implant. Cortical lamination was disturbed and heterotopic cell clusters were found in layer I of MK801-treated animals. Morphologically identified pyramidal neurons recorded in MK801-treated cortex revealed late NMDA receptor-mediated synaptic inputs and fragile monosynaptic responses at stimulation frequencies >0.2 Hz. Our data indicate that impairment of NMDA receptors during early corticogenesis induces neuronal migration disorders and delays the functional maturation of the developing cortical network. PMID- 15269116 TI - Functional connectivity in the human language system: a cortico-cortical evoked potential study. AB - A better understanding of the mechanisms involved in human higher cortical functions requires a detailed knowledge of neuronal connectivity between functional cortical regions. Currently no good method for tracking in vivo neuronal connectivity exists. We investigated the inter-areal connections in vivo in the human language system using a new method, which we termed 'cortico cortical evoked potentials' (CCEPs). Eight patients with epilepsy (age 13-42 years) underwent invasive monitoring with subdural electrodes for epilepsy surgery. Six patients had language dominance on the side of grid implantation and two had bilateral language representation by the intracarotid amobarbital test. Conventional cortical electrical stimulation was performed to identify the anterior and posterior language areas. Single pulse electrical stimuli were delivered to the anterior language (eight patients), posterior language (four patients) or face motor (two patients) area, and CCEPs were obtained by averaging electrocorticograms (ECoGs) recorded from the perisylvian and extrasylvian basal temporal language areas time-locked to the stimulus. The subjects were not asked to perform any tasks during the study. Stimulation at the anterior language area elicited CCEPs in the lateral temporo-parietal area (seven of eight patients) in the middle and posterior part of the superior temporal gyrus, the adjacent part of the middle temporal gyrus and the supramarginal gyrus. CCEPs were recorded in 3-21 electrodes per patient. CCEPs occurred at or around the particular electrodes in the posterior language area which, when stimulated, produced speech arrest. Similar early and late CCEPs were obtained from the basal temporal area by stimulating the anterior language area (three of three patients). In contrast, stimulation of the adjacent face motor area did not elicit CCEPs in language areas but rather in the postcentral gyrus. Stimulation of the posterior language area produced CCEPs in the anterior language (three of four patients) as well as in the basal temporal area (one of two patients). These CCEPs were less well defined. These findings suggest that perisylvian and extrasylvian language areas participate in the language system as components of a network by means of feed forward and feed-back projections. Different from the classical Wernicke Geschwind model, the present study revealed a bidirectional connection between Broca's and Wernicke's areas probably through the arcuate fasciculus and/or the cortico-subcortico-cortical pathway. CCEPs were recorded from a larger area than the posterior language area identified by electrical stimulation. This suggests the existence of a rather broad neuronal network surrounding the previously recognized core region of this area. PMID- 15269117 TI - Degradation of neuronal function following a spinal cord injury: mechanisms and countermeasures. AB - The aim of this study was to evaluate the course of spinal neuronal activity following spinal cord injury (SCI). In patients with a complete SCI, the leg muscle EMG activity early and up to 33 years after an SCI was analysed during locomotor movements induced and assisted by a driven gait orthosis (DGO). Only in chronic SCI patients did a premature exhaustion of neuronal activity occur. This was reflected in a reduced density and fading of leg muscle EMG activity. The early exhaustion of EMG activity was more pronounced in the leg flexor (e.g. biceps femoris) than extensor (e.g. gastrocnemius) muscles. The timing of the leg muscle pattern remained unchanged in the chronic patients. A preserved amplitude of motor action potentials following repetitive peripheral nerve stimulation and during spasms indicated an interneuronal site of impairment. In patients who participated in a locomotor training programme lasting up to 13 weeks, no positive effect on the slope of exhaustion was seen. It is concluded that a degradation of spinal neuronal activity takes place following an SCI. If in the future regeneration of spinal tract fibres becomes feasible in patients with complete SCI, such an approach can only become functionally successful if neuronal activity below the level of the lesion is maintained. This might be achieved by a continuous training approach starting early after injury. PMID- 15269118 TI - Rat gustatory neurons in the geniculate ganglion express glutamate receptor subunits. AB - Taste receptor cells are innervated by primary gustatory neurons that relay sensory information to the central nervous system. The transmitter(s) at synapses between taste receptor cells and primary afferent fibers is (are) not yet known. By analogy with other sensory organs, glutamate might a transmitter in taste buds. We examined the presence of AMPA and NMDA receptor subunits in rat gustatory primary neurons in the ganglion that innervates the anterior tongue (geniculate ganglion). AMPA and NMDA type subunits were immunohistochemically detected with antibodies against GluR1, GluR2, GluR2/3, GluR4 and NR1 subunits. Gustatory neurons were specifically identified by retrograde tracing with fluorogold from injections made into the anterior portion of the tongue. Most gustatory neurons in the geniculate ganglion were strongly immunoreactive for GluR2/3 (68%), GluR4 (78%) or NR1 (71%). GluR1 was seen in few cells (16%). We further examined if glutamate receptors were present in the peripheral terminals of primary gustatory neurons in taste buds. Many axonal varicosities in fungiform and vallate taste buds were immunoreactive for GluR2/3 but not for NR1. We conclude that gustatory neurons express glutamate receptors and that glutamate receptors of the AMPA type are likely targeted to synapses within taste buds. PMID- 15269119 TI - Responses of the rat chorda tympani nerve to glutamate-sucrose mixtures. AB - Monosodium glutamate (MSG) has a multifaceted, unusual taste to humans. Rats and other rodents also detect a complex taste to MSG. Responses of the chorda tympani nerve (CT) to glutamate applied to the front of the tongue were recorded in 13 anesthetized rats. Whole-nerve responses to 30 mM, 100 mM and 300 mM MSG mixed with 300 mM sucrose were recorded before and after adding 30 micro M amiloride to the rinse and stimulus solutions. Responses of CT single fibers were also recorded. Predictions from models of whole-nerve responses to binary mixtures were compared to the observed data. Results indicated that MSG-elicited CT responses have multiple sources, even in an amiloride-inhibited environment in rats. Those sources include responses of sucrose-sensitive CT neural units, which may provide the substrate for a sucrose-glutamate perceptual similarity, and responses of sucrose-insensitive CT neural units, which may respond synergistically to MSG-sucrose mixtures. PMID- 15269120 TI - Olfactory receptor neuron profiling using sandalwood odorants. AB - The mammalian olfactory system can discriminate between volatile molecules with subtle differences in their molecular structures. Efforts in synthetic chemistry have delivered a myriad of smelling compounds of different qualities as well as many molecules with very similar olfactive properties. One important class of molecules in the fragrance industry are sandalwood odorants. Sandalwood oil and four synthetic sandalwood molecules were selected to study the activation profile of endogenous olfactory receptors when exposed to compounds from the same odorant family. Dissociated rat olfactory receptor neurons were exposed to the sandalwood molecules and the receptor activation studied by monitoring fluxes in the internal calcium concentration. Olfactory receptor neurons were identified that were specifically stimulated by sandalwood compounds. These neurons expressed olfactory receptors that can discriminate between sandalwood odorants with slight differences in their molecular structures. This is the first study in which an important class of perfume compounds was analyzed for its ability to activate endogenous olfactory receptors in olfactory receptor neurons. PMID- 15269121 TI - The relative affective potency of glycine, L-serine and sucrose as assessed by a brief-access taste test in inbred strains of mice. AB - In general, rodents prefer both sucrose and L-serine relative to water and treat both compounds as possessing a similar taste quality (e.g. 'sweetness') despite that they are believed to bind with different T1R heterodimeric receptors in taste bud cells. We assessed the affective potency of these compounds along with glycine, which is thought to bind with both T1R receptor complexes, using a brief access taste test in a gustometer. Unconditioned licking responses of two 'taster' strains (C57BL/6J and SWR/J), which display high preference for low concentrations of sucrose, and two 'non-taster' (129P3/J and DBA/2J) strains, which display blunted preference for low concentrations of sucrose, were measured during 5 s trials of varying concentrations of a single compound when mice (n=10/strain/stimulus) were non-deprived and when access to home-cage water was restricted. In non-deprived mice, sucrose generated monotonically increasing concentration-response curves regardless of strain, whereas glycine was only marginally effective at stimulating licking and L-serine produced relatively flat functions. The profile of responsiveness across strains was more complex than expected. For example, when tested with sucrose in the non-deprived condition, the 129P3/J non-taster strain surpassed the responsiveness of taster mice at mid range to high concentrations. Under water-restricted conditions, these mice also were significantly more responsive to high concentrations of both sucrose and glycine compared with the other strains when stimulus licking was standardized relative to water. Thus, the affective potency of the stimuli tested here seems to be related to the ability of the compounds to bind with the T1R2+3 receptor complex. However, the profile of strain responsiveness to these tastants in the brief-access test does not appear to be simply explained by the sweetener 'taster' status of the strain. PMID- 15269122 TI - The role of heliothine hairpencil compounds in female Heliothis virescens (Lepidoptera: Noctuidae) behavior and mate acceptance. AB - Studies on numerous insect species suggest that male-produced sex pheromones play a role in attracting females; as aphrodisiacs, making females more quiescent; or as a means of inhibiting competing males. Male heliothine moths display abdominal hairpencils during courtship, but the specific effects of the odors released on female behavior have not yet been elucidated. This study investigates the role of male hairpencil compounds in female Heliothis virescens mating behavior. Female H. virescens were exposed to filter paper loaded with hairpencil extracts of male H. virescens, Heliothis subflexa and Helicoverpa zea, and observed for behavioral responses to odors. Single synthetic compounds found in the H. virescens hairpencil blend were also tested. In mating assays between single male and female H. virescens it was found that: (i) antennectomized females mated less frequently than sham-operated females; (ii) females mated less frequently with males whose hairpencils had been surgically removed; (iii) females mated with males with ablated hairpencils if a filter paper loaded with one male equivalent of H. virescens hairpencil extract was presented simultaneously; and (iv) this effect was species-specific, as presentation of H. subflexa or H. zea hairpencil extracts did not restore mate acceptance. This study suggests that odors released by male hairpencils are important in mate acceptance by female H. virescens, and may play a role in mate choice and species isolation. PMID- 15269123 TI - Oral zinc sulfate solutions inhibit sweet taste perception. AB - We investigated the ability of zinc sulfate (5, 25, 50 mM) to inhibit the sweetness of 12 chemically diverse sweeteners, which were all intensity matched to 300 mM sucrose [800 mM glucose, 475 mM fructose, 3.25 mM aspartame, 3.5 mM saccharin, 12 mM sodium cyclamate, 14 mM acesulfame-K, 1.04 M sorbitol, 0.629 mM sucralose, 0.375 mM neohesperidin dihydrochalcone (NHDC), 1.5 mM stevioside and 0.0163 mM thaumatin]. Zinc sulfate inhibited the sweetness of most compounds in a concentration dependent manner, peaking with 80% inhibition by 50 mM. Curiously, zinc sulfate never inhibited the sweetness of Na-cyclamate. This suggests that Na cyclamate may access a sweet taste mechanism that is different from the other sweeteners, which were inhibited uniformly (except thaumatin) at every concentration of zinc sulfate. We hypothesize that this set of compounds either accesses a single receptor or multiple receptors that are inhibited equally by zinc sulfate at each concentration. PMID- 15269124 TI - Female rats show a bimodal preference response to the artificial sweetener sucralose. AB - The preference of female Sprague-Dawley rats for sucralose, a non-nutritive sweetener derived from sucrose, was evaluated in 23 h two-bottle tests with water or saccharin. Overall, the rats displayed weak or no preferences for sucralose (0.25-4 g/l) over water but strong preferences for saccharin (0.5-8 g/l) over water and saccharin (1 g/l) over sucralose (0.5 g/l). The rats also preferred a saccharin + sucrose mixture to sucrose, but sucrose to a sucralose + sucrose mixture. There were marked individual differences in sucralose preferences: about half the rats preferred sucralose to water at some concentrations while most remaining rats avoided sucralose. Both subgroups preferred saccharin to sucralose. Sucralose appears to have an aversive off-taste that reduces its palatability to rats. PMID- 15269125 TI - Dose-response relationships in an olfactory flux detector model revisited. AB - A simple model of an odorant flux detector including odorant uptake, activation of odorant receptor molecules and enzymatic odorant deactivation can produce different types of static dose-response relationships. Depending on the binding characteristics of the odorant to the receptor molecule and to the deactivating enzyme, the receptor occupation by the odorant as related to the odorant uptake is quasi-hyperbolic, linear or, close to saturation, steeper than linear. In Rospars et al. (2003, Chem. Senses, 28: 509-522) a note contributed by both of us stated erroneously that an equation describing these relationships given previously (Kaissling, 1998, Chem. Senses, 23; 99-111; Kaissling, 2001, Chem. Senses, 26: 125-150) was incorrect. We show here that the difference in equations was due to a simplifying assumption in Rospars et al. (2003) about the deactivating enzyme, we summarize briefly the properties of the correct equation of Kaissling (1998, 2001) and we discuss the relation with the model studied in Rospars et al. (2003). PMID- 15269126 TI - Olfactory sensitivity of subjects working in odorous environments. AB - The aim of the present study was to investigate whether people with a professional interest in odors also exhibit higher olfactory sensitivity. To this end, we investigated 58 subjects (age 33.6 +/- 11.0 years, mean +/- SD; 55 women) employed in perfume retail outlets and compared their olfactory sensitivity to 58 controls (age 34.6 +/- 9.9 years; 53 women) matched for age, gender and professional activities who did not work in such odorous environments. Olfactory function was assessed using the 'Sniffin' Sticks' test kit which includes tests for n-butanol odor threshold, odor discrimination and odor identification. Subjects working in perfume retail outlets scored higher in odor discrimination tests compared to controls. Working in an odorous environment for a full day had no major effect on general olfactory abilities, as indicated by measures performed at the beginning and end of a working day. Taken together, results from the present study do not support the idea that odorous environments are deleterious to general olfactory function. PMID- 15269127 TI - Neurochemistry of the gustatory subgemmal plexus. AB - Nerve fibers present in the basal plexus of the vallate papilla of the rat tongue were analyzed using cytochemical, immunocytochemical and ultrastructural methods to investigate whether the subgemmal plexus is subdivided into neurochemical compartments and to provide a clear definition of the reciprocal spatial relationships between nitrergic, peptidergic and acetylesterase positive structures. Several neuronal fibers were detected under the chemoreceptorial epithelium. Some of these fibers were in contact with the taste buds and in some cases neuronal projections were also present between the buds or inside them; some others fibers were present below this layer but in a more peripheral area. Antibodies against CGRP, SP and CCK stained fibers just below the chemoreceptorial epithelium, whereas fibers more distally located were immunolabeled by anti VIP, NOS-1 and NF-200 antibodies. Some double staining experiments were conducted using confocal microscopy. Other sections were processed cytochemically for AChE and subsequently for NADPH-d in colocalization experiments. All the data obtained using these techniques confirmed the results obtained with single immunostaining, as did the ultrastructural results. In conclusion, the present work demonstrates that the subgemmal plexus is a bilayered structure, suggesting that the complex relationship between the two layers plays a pivotal role in taste and in the control of processes ancillary to taste, such as control of vascular or secretory mechanisms. PMID- 15269128 TI - Demographic and cognitive predictors of cued odor identification: evidence from a population-based study. AB - This study investigated demographic and cognitive correlates of cued odor identification in a population-based sample from the Betula project: 1906 healthy adults varying in age from 45 to 90 years were assessed in a number of tasks tapping various cognitive domains, including cognitive speed, semantic memory and executive functioning. The results revealed a gradual and linear deterioration in cued odor identification across the adult life span. Overall, females identified more odors than men, although men and women performed at the same level in the oldest age cohort (85-90 years). Hierarchical regression analyses revealed that age, sex, education, cognitive speed and vocabulary were reliable correlates of performance in the odor identification task. In addition, age-related deficits in the included demographic and cognitive variables could not fully account for the observed age-related impairment in identification, suggesting that additional factors are underlying the observed deterioration. Likely candidates here are sensory abilities such as olfactory detection and discrimination. PMID- 15269129 TI - The clinical application of targeting cancer through histone acetylation and hypomethylation. AB - Methods of gene inactivation include genetic events such as mutations or deletions. Epigenetic changes, heritable traits that are mediated by changes in DNA other than nucleotide sequences, play an important role in gene expression. Two epigenetic events that have been associated with transcriptional silencing include methylation of CpG islands located in gene promoter regions of cancer cells and changes in chromatin conformation involving histone acetylation. Recent evidence demonstrates that these processes form layers of epigenetic silencing. Reversal of these epigenetic processes and up-regulation of genes important to prevent or reverse the malignant phenotype has therefore become a new therapeutic target in cancer treatment. PMID- 15269130 TI - DNA array-based gene profiling in tumor immunology. AB - Recent advances in tumor immunology have fostered the clinical implementation of different immunotherapy modalities. However, the alternate success of such regimens underscores the fact that the molecular mechanisms underlying tumor immune rejection are still poorly understood. Given the complexity of the immune system network and the multidimensionality of tumor-host interactions, the comprehension of tumor immunology might greatly benefit from high-throughput DNA array analysis, which can portray the molecular kinetics of immune response on a genome-wide scale, thus accelerating the accumulation of knowledge and ultimately catalyzing the development of new hypotheses in cell biology. Although in its infancy, the implementation of DNA array technology in tumor immunology studies has already provided investigators with novel data and intriguing hypotheses on the cascade of molecular events leading to an effective immune response against cancer. Although the principles of DNA array-based gene profiling techniques have become common knowledge, the need for mastering this technique to produce meaningful data and correctly interpret this enormous output of information is critical and represents a tremendous challenge for investigators. In the present work, we summarize the main technical features and critical issues characterizing this powerful laboratory tool and review its applications in the fascinating field of cancer immunogenomics. PMID- 15269131 TI - Development of the novel biologically targeted anticancer agent gefitinib: determining the optimum dose for clinical efficacy. AB - The emergence of novel, biologically targeted anticancer agents such as gefitinib ('Iressa', ZD1839) has raised the question of how the dose for later-stage clinical development and clinical use is best determined. For cytotoxic drugs, because toxic effects and antitumor activity often fall within the same dose range and are dose dependent, the clinically used dose will depend on the therapeutic window. Therefore, the maximum tolerated dose identified in Phase I trials is typically used to determine the dose level for Phase II and III trials. However, because biologically targeted agents are expected to provide clinical benefits that are not predicted by surrogate end points of toxicity to normal replicating tissue, new Phase I trials have been designed to determine the optimum biological dose for use in further studies. A large, multifaceted Phase I program was designed to evaluate the pharmacokinetics, safety, efficacy, and targeted biological activity of a once-daily oral dose of gefitinib. The maximum tolerated dose was >or=700 mg/day, although doses as low as 150 mg/day provided (a). plasma concentrations sufficient for pharmacological activity, (b). evidence of targeted biological effect, and (c). antitumor activity. From these observations, two large Phase II trials ('Iressa' Dose Evaluation in Advanced Lung Cancer 1 and 2) evaluated 250- and 500-mg/day doses of gefitinib in patients with advanced non-small cell lung cancer (NSCLC). As predicted from the Phase I trials, doses >250 mg/day provided no additional efficacy benefit, whereas adverse effects increased in a dose-dependent manner. Consequently, the recommended dose of gefitinib in NSCLC is 250 mg/day. The early clinical trial development of gefitinib provides a model for the development of novel, noncytotoxic anticancer agents. PMID- 15269132 TI - The cooperative prostate cancer tissue resource: a specimen and data resource for cancer researchers. AB - PURPOSE: The Cooperative Prostate Cancer Tissue Resource (CPCTR) is a National Cancer Institute-supported tissue bank that provides large numbers of clinically annotated prostate cancer specimens to investigators. This communication describes the CPCTR to investigators interested in obtaining prostate cancer tissue samples. EXPERIMENTAL DESIGN: The CPCTR, through its four participating institutions, has collected specimens and clinical data for prostate cancer cases diagnosed from 1989 onward. These specimens include paraffin blocks and frozen tissue from radical prostatectomy specimens and paraffin blocks from prostate needle biopsies. Standardized histopathological characterization and clinical data extraction are performed for all cases. Information on histopathology, demography (including ethnicity), laboratory data (prostate-specific antigen values), and clinical outcome related to prostate cancer are entered into the CPCTR database for all cases. Materials in the CPCTR are available in multiple tissue formats, including tissue microarray sections, paraffin-embedded tissue sections, serum, and frozen tissue specimens. These are available for research purposes following an application process that is described on the CPCTR web site (www.prostatetissues.org). RESULTS: The CPCTR currently (as of October 2003) contains 5135 prostate cancer cases including 4723 radical prostatectomy cases. Frozen tissues, in some instances including patient serum samples, are available for 1226 cases. Biochemical recurrence data allow identification of cases with residual disease, cases with recurrence, and recurrence-free cases. CONCLUSIONS: The CPCTR offers large numbers of highly characterized prostate cancer tissue specimens, including tissue microarrays, with associated clinical data for biomarker studies. Interested investigators are encouraged to apply for use of this material (www.prostatetissues.org). PMID- 15269133 TI - State of the translational science: summary of Baltimore workshop on gene re expression as a therapeutic target in cancer January 2003. AB - A workshop was held in Baltimore, Maryland in January 2003 to discuss translational aspects of cancer therapies targeted at impacting aberrant gene transcription due to epigenetic changes. The mission of the meeting was the development of strategies for scientifically sound, clinically feasible applications targeting epigenetics in cancer therapy. Sessions included preclinical discussions of DNA methylation, the histone code, chromatin remodeling, and transcriptional control. Data on the histone deacetylase and DNA methyltransferase inhibitors under preclinical and clinical investigation were presented and discussed. The optimal correlative laboratory studies for monitoring clinical trials with these agents remain controversial. DNA methyltransferase and histone deacetylase inhibitors will be combined with each other to maximally re-express genes silenced through promoter methylation. Other classes of agents that may be rationally combined with these classes of drugs include retinoids, steroid hormones, and cytotoxic drugs. PMID- 15269134 TI - Hypoxia and Photofrin uptake in the intraperitoneal carcinomatosis and sarcomatosis of photodynamic therapy patients. AB - PURPOSE: Response to photodynamic therapy depends on adequate tumor oxygenation as well as sufficient accumulation of photosensitizer in the tumor. The goal of this study was to investigate the presence of hypoxia and retention of the photosensitizer Photofrin in the tumors of patients with intra-abdominal carcinomatosis or sarcomatosis. EXPERIMENTAL DESIGN: Tumor nodules from 10 patients were studied. In nine of these patients, hypoxia was identified in histological sections of biopsied tumor after administration of the hypoxia marker 2-(2-nitroimidazol-1[H]-yl)-N-(2,2,3,3,3-pentafluoropropyl)acetamide (EF5). In separate tumor nodules from 10 patients, Photofrin uptake was measured by fluorescence after tissue solubilization. RESULTS: Hypoxia existed in the tumors of five patients, with three of these patients demonstrating at least one severely hypoxic nodule. Physiological levels of oxygen were present in the tumors of four patients. An association between tumor size and hypoxia was not evident because some tumor nodules as small as approximately 2 mm in diameter were severely hypoxic. However, even these tumor nodules contained vascular networks. Three patients with severely hypoxic tumor nodules exhibited moderate levels of Photofrin uptake of 3.9 +/- 0.4 to 3.9 +/- 0.5 ng/mg (mean +/- SE). The four patients with tumors of physiological oxygenation did not consistently exhibit high tumor concentrations of Photofrin: mean +/- SE drug uptake among these patients ranged from 0.6 +/- 0.8 to 5.8 +/- 0.5 ng/mg. CONCLUSIONS: Carcinomatosis or sarcomatosis of the i.p. cavity may exhibit severe tumor hypoxia. Photofrin accumulation in tumors varied by a factor of approximately 10x among all patients, and, on average, those with severe hypoxia in at least one nodule did not demonstrate poor Photofrin uptake in separate tumor samples. These data emphasize the need for reconsideration of the generally accepted paradigm of small tumor size, good oxygenation, and good drug delivery because this may vary on an individual tumor basis. PMID- 15269135 TI - Her2/neu expression predicts the response to antiaromatase neoadjuvant therapy in primary breast cancer: subgroup analysis from celecoxib antiaromatase neoadjuvant trial. AB - PURPOSE: Many studies suggest that Her2/neu play an important role in neoadjuvant endocrine therapy. This study aimed to determine whether the level of Her2/neu expression in advanced breast cancer changes after antiaromatase neoadjuvant treatment, as well as to identify the relationship between Her2/neu expression and response to this kind of therapy. EXPERIMENTAL DESIGN: Thirty-six postmenopausal patients with hormonal receptor-positive primary breast cancer were included in a study of three monthly cycles of neoadjuvant endocrine therapy with either Aromasin (25 mg daily) or Femara (2.5 mg daily). Immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) for Her2/neu were conducted both on pretreatment biopsies and surgical tumors. RESULTS: Using IHC, 5 of 36 (13.9%) of the patients had a Her2/neu overexpression after treatment, as compared with 16 of 36 (44.4%) before. Meanwhile, there was no change in 21 (58.3%) patients, and through FISH, there was a change from amplification to no amplification in 15 (41.7%) patients. The response rate to the treatment was 75% for Her2/neu (+) tumors and 35% for Her2/neu (-) tumors (P = 0.017) while FISH was performed. The response rate was also significantly affected by the decrease in Her2/neu status after the treatment, with 73% of the tumors showing decreased Her2/neu expression and with 38% of the tumors showing no change of Her2/neu expression (P = 0.037). CONCLUSIONS: Using both IHC and FISH, advanced breast cancers show statistical evidence of decreasing incidence of Her2/neu expression after antiaromatase neoadjuvant treatment. Our data also suggest that Her2/neu expression and its change during the treatment might be predictive markers for this kind of therapy. PMID- 15269136 TI - Patient characteristics compete with dose as predictors of acute treatment toxicity in early phase clinical trials. AB - PURPOSE: The purpose of this study was to identify patient characteristics that may be risk factors or markers of susceptibility to adverse treatment effects in cancer Phase I and II clinical trials. PATIENTS AND METHODS: A total of 459 patients enrolled in 23 therapeutic Phase I and II studies at the Fox Chase Cancer Center were included in the analysis. Patient-specific characteristics, medical and treatment history, doses of experimental agents, and graded toxicities were extracted from case report forms. We developed a novel summary measure, the toxicity index (TI), to better discriminate patients on the basis of their overall toxicity experiences. Mixed model ANOVA was used to model TI on the basis of data from all trials using a specific agent. Generalized estimating equations in the context of binary logistic regression were used to model dose limiting toxicity. RESULTS: Seventeen pretreatment factors, including performance status, alkaline phosphatase, total bilirubin, serum creatinine, and tobacco use, emerged as significant predictors of toxicity as defined by dose-limiting toxicity or TI. Unexpectedly, dose was not always a predictor of toxicity. Even for values within the normal range, the TI identified serum bilirubin and alkaline phosphatase as predictors of toxicity after treatment with docetaxel and alkaline phosphatase as a predictor for toxicity after treatment with irinotecan. CONCLUSIONS: Independent of dose, certain pretreatment characteristics, including measures of organ function that are in the normal range, were found to be predictors of treatment toxicity. Because of its sensitivity to differences in overall toxicity, the TI should prove to be a useful tool for identifying predictors of chemotherapy-related toxicity. PMID- 15269137 TI - Alternative splicing of the multidrug resistance protein 1/ATP binding cassette transporter subfamily gene in ovarian cancer creates functional splice variants and is associated with increased expression of the splicing factors PTB and SRp20. AB - PURPOSE: Overexpression of multidrug resistance protein 1 (MRP1) confers resistance to a range of chemotherapeutic agents in cell lines and could be involved in clinical drug resistance of some tumor types also. We examined MRP1 expression in a small series of untreated human ovarian tumors and matched normal tissues. EXPERIMENTAL DESIGN: We analyzed ten pairs of snap-frozen ovarian tumor and matched normal total ovarian tissues from the same patients for expression of MRP1 by reverse transcription-PCR. Amplified PCR products were sequenced to reveal splicing events of MRP1. MRP1 splice variants were expressed as enhanced green fluorescent fusion proteins in HEK293T cells to demonstrate their localization in the cell and their activity in conferring resistance to doxorubicin. The expression of splicing factors PTB and SRp20 was examined by Western blot. RESULTS: MRP1 was expressed in all 10 of the pairs of specimens. Multiple MRP1 cDNA fragments of various sizes were amplified between exons 10 and 19. Of interest, more MRP1 cDNA fragments were detected in ovarian tumors than in matched normal tissues in 9 of 10 pairs. We identified 10 splicing forms between exons 10 and 19 of the MRP1 gene with exon skipping ranging from 1 to 7. Amplification of the entire coding region of MRP1 from 1 ovarian tumor revealed >20 splice variants. We found whole and partial exon skipping and partial intron inclusion in these splice variants. We expressed 3 of these MRP1 splice variants in HEK293T cells and found that they appeared to localize to the plasma membrane and were functional in conferring resistance to doxorubicin. In addition, we identified a few nucleotide variations in this gene. To understand the basis for increased splice variants in the tumors, we examined splicing factor expression in these tissues. Western blot analysis revealed that two splicing factors, PTB and SRp20, were overexpressed in most ovarian tumors compared with their matched normal ovarian tissues. Importantly, overexpression of both of these splicing factors was associated with the increased number of MRP1 splicing forms in the ovarian tissues. CONCLUSION: The MRP1 gene undergoes alternative splicing at a higher frequency in ovarian tumors than in matched normal tissues. Some of these splice variants confer resistance to doxorubicin. Expression of splicing factors PTB and SRp20 is strongly associated with the alternative splicing of the MRP1 gene. PMID- 15269138 TI - Increased expression of the copper efflux transporter ATP7A mediates resistance to cisplatin, carboplatin, and oxaliplatin in ovarian cancer cells. AB - PURPOSE: The goal of this study was to determine the effect of small changes in ATP7A expression on the pharmacodynamics of cisplatin, carboplatin, and oxaliplatin in human ovarian carcinoma cells. EXPERIMENTAL DESIGN: Drug sensitivity and cellular pharmacology parameters were determined in human 2008 ovarian carcinoma cells and a subline transfected with an ATP7A-expression vector ATP7A (2008/MNK). Drug sensitivity was determined by clonogenic assay, platinum (Pt) levels were measured by inductively coupled plasma mass spectroscopy, copper (Cu) accumulation was quantified with (64)Cu, and the subcellular distribution of ATP7A was assessed by confocal digital microscopy. RESULTS: The 1.5-fold higher expression of ATP7A in the 2008/MNK cells was sufficient to alter Cu cellular pharmacokinetics but not confer Cu resistance. In contrast, it was sufficient to render the 2008/MNK cells resistant to cisplatin, carboplatin, and oxaliplatin. Resistance was associated with increased rather than decreased whole-cell Pt drug accumulation and increased sequestration of Pt into the vesicular fraction. Cu triggered relocalization of ATP7A away from the perinuclear region, whereas at equitoxic concentrations the Pt drugs did not. CONCLUSIONS: A small increase in ATP7A expression produced resistance to all three of the clinically available Pt drugs. Whereas increased expression of ATP7A reduced Cu accumulation, it did not reduce accumulation of the Pt drugs. Under conditions where Cu triggered ATP7A relocalization, the Pt drugs did not. Thus, although ATP7A is an important determinant of sensitivity to the Pt drugs, there are substantial differences between Cu and the Pt drugs with respect to how they interact with ATP7A and the mechanism by which ATP7A protects the cell. PMID- 15269139 TI - Inhibition of cyclooxygenase-1 and -2 expression by targeting the endothelin a receptor in human ovarian carcinoma cells. AB - PURPOSE AND EXPERIMENTAL DESIGN: New therapies against cancer are based on targeting cyclooxygenase (COX)-2. Activation of the endothelin A receptor (ET(A)R) by endothelin (ET)-1 is biologically relevant in several malignancies, including ovarian carcinoma. In this tumor, the ET-1/ET(A)R autocrine pathway promotes mitogenesis, apoptosis protection, invasion, and neoangiogenesis. Because COX-1 and COX-2 are involved in ovarian carcinoma progression, we investigated whether ET-1 induced COX-1 and COX-2 expression through the ET(A)R at the mRNA and protein level in HEY and OVCA 433 ovarian carcinoma cell lines by Northern blot, reverse transcription-PCR, Western blot, and immunohistochemistry; we also investigated the activity of the COX-2 promoter by luciferase assay and the release of prostaglandin (PG) E(2) by ELISA. RESULTS: ET-1 significantly increases the expression of COX-1 and COX-2, COX-2 promoter activity, and PGE(2) production. These effects depend on ET(A)R activation and involve multiple mitogen-activated protein kinase (MAPK) signaling pathways, including p42/44 MAPK, p38 MAPK, and transactivation of the epidermal growth factor receptor. COX 2 inhibitors and, in part, COX-1 inhibitor blocked ET-1-induced PGE(2) and vascular endothelial growth factor release, indicating that both enzymes participate in PGE(2) production to a different extent. Moreover, inhibition of human ovarian tumor growth in nude mice after treatment with the potent ET(A)R selective antagonist ABT-627 is associated with reduced COX-2 and vascular endothelial growth factor expression. CONCLUSIONS: These results indicate that impairing COX-1 and COX-2 and their downstream effect by targeting ET(A)R can be therapeutically advantageous in ovarian carcinoma treatment. Pharmacological blockade of the ET(A)R is an attractive strategy to control COX-2 induction, which has been associated with ovarian carcinoma progression and chemoresistance. PMID- 15269140 TI - A phase II clinical and pharmacodynamic study of E7070 in patients with metastatic, recurrent, or refractory squamous cell carcinoma of the head and neck: modulation of retinoblastoma protein phosphorylation by a novel chloroindolyl sulfonamide cell cycle inhibitor. AB - PURPOSE: E7070 is a synthetic sulfonamide cell cycle inhibitor that induces hypophosphorylation of the retinoblastoma (Rb) protein and G(1) arrest in vitro. This Phase II study was conducted to explore the efficacy, safety, and pharmacodynamics of E7070 in squamous cell carcinoma of the head and neck (SCCHN). EXPERIMENTAL DESIGN: Patients with metastatic, recurrent, or refractory SCCHN, treated with no more than one prior therapy for recurrent disease, received E7070 at 700 mg/m(2) over 1 h every 3 weeks. Pre- and posttreatment tumor fine needle aspirates were subjected to immunohistochemistry with a panel of phospho-specific anti-Rb antibodies. End points included progression-free survival, response rate and duration, overall survival, toxicity profile, and inhibition of Rb phosphorylation. RESULTS: Because none of the first 15 patients achieved progression-free survival > 4 months, the early stopping rule was invoked. Eleven patients had oropharyngeal cancer and 12 were male. Median age was 59 years (range, 49-73 years). Thirty-nine cycles of E7070 were delivered (median, 2.6 cycles/patient; range, 1-5 cycles). Six patients had stable disease after 2 cycles and 2 patients each subsequently received 1, 2, and 3 additional cycles, respectively, before experiencing progression. Immunohistochemistry of tumor cell aspirates from 3 patients demonstrated reduced Rb phosphorylation posttreatment. CONCLUSIONS: At this dose and schedule, E7070 is unlikely to be superior over single-agent chemotherapy in SCCHN. However, the data suggest that cdk activity can be inhibited in tumor cells, resulting in posttreatment modulation of Rb phosphorylation. In the absence of cytotoxicity, more frequent administration of E7070 may be required to sustain Rb hypophosphorylation and cytostatic growth arrest. PMID- 15269141 TI - Immunization of patients with the hTERT:540-548 peptide induces peptide-reactive T lymphocytes that do not recognize tumors endogenously expressing telomerase. AB - PURPOSE: Telomerase is an attractive target antigen for cancer immunotherapies because it is expressed in >85% of human tumors but is rarely found in normal tissues. A HLA-A*0201-restricted T-cell epitope was previously identified within telomerase reverse transcriptase hTERT:540-548. This peptide was reported to induce CTL that recognized tumor cells and transfectants that endogenously expressed telomerase. Therefore, we initiated a clinical protocol to evaluate the therapeutic and immunological efficacy of this peptide. EXPERIMENTAL DESIGN: Fourteen patients with metastatic cancers were vaccinated with hTERT:540-548 emulsified in incomplete Freund's adjuvant. RESULTS: In 7 patients, peripheral blood mononuclear cells collected after immunization recognized hTERT:540-548, whereas those collected before vaccination did not. However, none of these CTLs recognized tumors that endogenously expressed telomerase, and none of the patients had an objective clinical response. Several highly avid T-cell clones were generated that recognized T2 cells pulsed with 75% of tumor samples. The expression of TIMP-1, -2, and -3, on the other hand, was found to depend on the histological subtype: TIMP 3 was often found in classical MB, whereas TIMP-2 was often expressed in desmoplastic MB (P = 0.007-0.001). In addition, both TIMP-3 and -2 correlated significantly with the expression of all studied metalloproteinases except MMP-2. TIMP-1, detected only in classical MB in a low percentage, was the only TIMP that correlated with the expression of MMP-2. Kaplan-Meier estimation revealed significantly reduced long-term survival of patients with strong MMP expression in tumor samples. In multivariate logistic regression analysis, however, the prognosis was significantly determined only by clinical parameters. CONCLUSIONS: TIMP-3 and -2 expression is highly correlated with histological subtypes of MBs and strongly associated with the expression of certain MMPs. The expression of TIMPs and MMPs, however, does not determine prognosis independently of clinical parameters. PMID- 15269149 TI - On the role of melanoma-specific CD8+ T-cell immunity in disease progression of advanced-stage melanoma patients. AB - Cytotoxic T-cell immunity directed against melanosomal differentiation antigens is arguably the best-studied and most prevalent form of tumor-specific T-cell immunity in humans. Despite this, the role of T-cell responses directed against melanosomal antigens in disease progression has not been elucidated. To address this issue, we have related the presence of circulating melanoma-specific T cells with disease progression and survival in a large cohort of patients with advanced stage melanoma who had not received prior treatment. In 42 (68%) of 62 patients, melanoma-specific T cells were detected, sometimes in surprisingly large numbers. Disease progression during treatment was more frequent in patients with circulating melanoma-specific T cells, and mean survival of patients with circulating melanoma-specific T cells was equal to the survival of patients without melanoma-specific T cells. These data suggest that the induction of melanosomal differentiation antigen-specific T-cell reactivity in advanced stage melanoma is a late event most likely due to antigen load and spreading and is not accompanied by a clinically significant antitumor effect. These melanoma-specific T cells may be functionally distinct from T cells raised during spontaneous regression or up vaccination. PMID- 15269150 TI - Expression of trypsinogen-1, trypsinogen-2, and tumor-associated trypsin inhibitor in ovarian cancer: prognostic study on tissue and serum. AB - PURPOSE: The purpose is to study the prognostic significance of tissue expression of trypsinogen-1, trypsinogen-2, and tumor-associated trypsin inhibitor (TATI) and serum concentration of trypsinogen-2, trypsin-2-API (complex of trypsin-2 with alpha-1-proteinase inhibitor), and TATI in epithelial ovarian cancer. EXPERIMENTAL DESIGN: Expression of trypsinogen-1, trypsinogen-2, and TATI was determined by immunohistochemistry with monoclonal antibodies in tissue sections of tumors from 119 patients with untreated primary epithelial ovarian cancer. Preoperative serum concentrations of trypsinogen-2, trypsin-2-API and TATI were analyzed using specific immunoassays. RESULTS: Fifty-four percent of the tumors expressed trypsinogen-1, 45% expressed trypsinogen-2, and 30% expressed TATI. In patients with stage III and IV disease, TATI tissue expression (P = 0.002) and elevated TATI concentration in serum (P = 0.048) were associated with adverse cancer-specific and progression-free survival in univariate analysis. In multivariate analysis, TATI tissue expression (P = 0.005), tumor grade (P = 0.0001), histological type (P = 0.02), and stage (P = 0.0005) were independent prognostic factors for adverse cancer-specific survival and TATI tissue expression (P = 0.006) and grade (P = 0.0003) for progression-free survival. In multivariate analysis of all patients and those with advanced disease, serum trypsin-2-API concentration was an adverse prognostic factor for cancer-specific and progression-free survival, and it was independent of stage and histological type of the tumor (P A, T481K; 1714C-->T, R571X]. The 1422C-->A allele was inherited from the mother, whereas the 1714C-->T, allele paternally inherited, was apparently null as a result of nonsense-mediated mRNA decay (NMD). Interestingly, the 1714C-->T mutation is the same as previously identified in an unrelated English ATLD family (probands ATLD3 and ATLD4), suggesting an important role for NMD in saving potentially lethal mutations. Lymphoblastoid cell lines (LCLs) derived from ATLD5 and ATLD6 were normal for ATM, but defective for Mre11, Rad50 and Nbs1 (the MRN complex) protein expression. Their response to gamma-radiation was abnormal, as evidenced by the enhanced radiosensitivity, attenuated autophosphorylation of ATM S1981 and phosphorylation of the ATM targets p53-S15 and Smc1-S966, failure to form Mre11 nuclear foci and defective G1 checkpoint arrest. The fibroblasts, but not LCLs, from ATLD5 and ATLD6 showed an impaired ATM-dependent Chk2 phosphorylation. These findings further underscore the interconnection between ATM activity and MRN function, which rationalizes the clinical similarity between ataxia-telangiectasia (A-T) and ATLD. PMID- 15269181 TI - Characterization of Ighmbp2 in motor neurons and implications for the pathomechanism in a mouse model of human spinal muscular atrophy with respiratory distress type 1 (SMARD1). AB - Spinal muscular atrophy with respiratory distress type 1 (SMARD1) is caused by recessive mutations of the IGHMBP2 gene. The role of IGHMBP2 (immunoglobulin mu binding protein 2) in the pathomechanism of motor neuron disease is unknown. We have generated antibodies against Ighmbp2 and showed that low levels of Ighmbp2 immunoreactivity are present in the nucleus of spinal motor neurons and high levels in cell bodies, axons and growth cones. Ighmbp2 protein levels are strongly reduced in neuromuscular degeneration (nmd) mice, the mouse model of SMARD1. Mutant mice show severe motor neuron degeneration before first clinical symptoms become apparent. The loss of motor neuron cell bodies in lumbar spinal cord is followed by axonal degeneration in corresponding nerves such as the femoral quadriceps and sciatic nerve and loss of axon terminals at motor endplates. Motor neuron degeneration and clinical symptoms then slowly progress until the mice die at the age of 3-4 months. In addition, myopathic changes seem to contribute to muscle weakness and especially to respiratory failure, which is characteristic of the disorder in humans. Cultured motor neurons from embryonic nmd mice did not show any abnormality with respect to survival, axonal growth or growth cone size, thus differing from motor neurons derived from, e.g. Smn (survival motor neuron) deficient mice, the model of spinal muscular atrophy (SMA). Our data suggest that the pathomechanism in SMARD1 is clearly distinct from other motor neuron diseases such as classic SMA. PMID- 15269182 TI - Spastin interacts with the centrosomal protein NA14, and is enriched in the spindle pole, the midbody and the distal axon. AB - Hereditary spastic paraplegia (HSP) is characterized by the specific retrograde degeneration of the longest axons in the central nervous system, the corticospinal tracts. The gene most frequently involved in autosomal dominant cases of this disease, SPG4, encodes spastin, an ATPase belonging to the AAA family. AAA proteins are thought to exert their function by the energy-dependent rearrangement of protein complexes. The composite function of these proteins is directed by their binding to regulatory factors and adaptor proteins that target their activity into specific pathways in vivo. We previously found that overexpressed spastin interacts dynamically with microtubules and displays microtubule-severing activity. Here, we demonstrate that spastin is enriched in cell regions containing dynamic microtubules. During cell division spastin is found in the spindle pole, the central spindle and the midbody, whereas in immortalized motoneurons it is enriched in the distal axon and the branching points. Furthermore, spastin interacts with the centrosomal protein NA14, and co fractionates with gamma-tubulin, a centrosomal marker. Deletion of the region required for binding to NA14 disrupts spastin interaction with microtubules, suggesting that NA14 may be an important adaptor to target spastin activity at the centrosome. These data strongly argue that spastin plays a role in cytoskeletal rearrangements and dynamics, and provide an attractive explanation for the degeneration of motor axons in HSP. PMID- 15269183 TI - Inflammation-induced transcriptional regulation of Golgi transporters required for the synthesis of sulfo sLex glycan epitopes. AB - The de novo synthesis and expression of sulfo sLex glycan on vascular endothelial glycoproteins has a central role in the initiation of inflammatory reactions, serving as a putative ZIP code for organ-specific trafficking of leukocytes into sites of inflammation. The synthesis of sulfo sLex requires energy carrying donors, CMP-sialic acid (CMP-SA), GDP-fucose (GDP-Fuc), and adenosine 3' phosphate 5'-phosphosulphate (PAPS) for donation of SA, Fuc, and sulfate, respectively. These donors are synthesized in the nucleus or cytosol and translocated into Golgi by specific transporters where corresponding transferase and proteins as well as enzymatic activities increase on inflammatory stimuli. Here we analyze the transcriptional coregulation of CMP-SA, GDP-Fuc, and PAPS transporters with in situ hybridization and real-time PCR in acute inflammation using kidney and heart allografts as model systems. Our results indicate that these three transporters display coordinated transcriptional regulation during the induction of the sulfo sLex glycan biosynthesis. With in silico analysis, the data generated with 230 human Affymetrix U133A gene chips indicated that the coregulated expression of CMP-SA and GDP-Fuc transporters was not common. Taken together our results suggest that inflammation-induced transcriptional regulation exists for Golgi membrane transporters required for the synthesis of the inflammation-inducible ZIP code sulfo sLex glycans. PMID- 15269184 TI - Two major grapefruit juice components differ in intestinal CYP3A4 inhibition kinetic and binding properties. AB - Bergamottin (BG) and 6',7'-dihydroxybergamottin (DHB) are the most abundant furanocoumarins present in grapefruit juice and have been proposed as major intestinal CYP3A4 inhibitors contributing to grapefruit juice-drug interactions. The relative contribution of BG versus DHB to the interaction potential is unclear, in part due to inconsistencies in the literature regarding inhibitory potency. To resolve these inconsistencies, the inhibitory kinetics of each furanocoumarin toward CYP3A4 catalytic activity were systematically characterized using representative probes from two distinct CYP3A4 substrate subgroups (testosterone and midazolam). With human intestinal microsomes, DHB was a substrate-independent reversible (Ki, approximately 0.8 microM) and mechanism based (KI, approximately 3 microM; kinact, 0.3-0.4 min(-1)) inhibitor of CYP3A4. In contrast, BG was a substrate-dependent reversible inhibitor, with a Ki (13 microM) using midazolam that was 8-fold greater than that using testosterone, but a substrate-independent mechanism-based inhibitor (KI, approximately 25 microM; kinact, approximately 0.35 min(-1)). Similar trends resulted with cDNA-expressed CYP3A4, only the KI values for BG were approximately 10-fold lower than with microsomes. This seemed to reflect a much greater degree of microsomal protein binding by BG compared with DHB. Differential inhibition kinetics and binding properties between BG and DHB could account in part for the apparent in vitro inconsistencies in the literature. Results also emphasize the importance of appropriate substrate selection when designing inhibition studies involving dietary constituents. PMID- 15269185 TI - Triclosan as a substrate and inhibitor of 3'-phosphoadenosine 5'-phosphosulfate sulfotransferase and UDP-glucuronosyl transferase in human liver fractions. AB - Triclosan is a broad spectrum antibacterial agent used in many household products. Due to its structural similarity to polychlorobiphenylols, which are potent inhibitors of the sulfonation and glucuronidation of 3-hydroxy benzo[a]pyrene, it was hypothesized that triclosan would inhibit these phase II enzymes. This study was designed to assess the interactions of triclosan as a substrate and inhibitor of 3'-phosphoadenosine 5'-phosphosulfate sulfotransferases and UDP-glucuronosyltransferases in human liver cytosol and microsomes. Triclosan was sulfonated and glucuronidated in human liver. The apparent Km and Vmax values for triclosan sulfonation were 8.5 microM and 0.096 nmol/min/mg protein, whereas Km and Vmax values for glucuronidation were 107 microM and 0.739 nmol/min/mg protein. Triclosan inhibited the hepatic cytosolic sulfonation of 3-hydroxybenzo(a)pyrene (3-OH-BaP), bisphenol A, p-nitrophenol, and acetaminophen with IC50 concentrations of 2.87, 2.96, 6.45, and 17.8 microM, respectively. Studies of 3-OH-BaP sulfonation by expressed human SULT1A1*1, SULT1A1*2, SULT1B1, and SULT1E1 showed that triclosan inhibited the activities of each of these purified enzymes with IC50 concentrations between 2.09 and 7.5 microM. Triclosan was generally a less potent inhibitor of microsomal glucuronidation. IC50 concentrations for triclosan with 3-OH-BaP, acetaminophen, and bisphenol A as substrates were 4.55, 297, and >200 microM, respectively. Morphine glucuronidation was not inhibited by 50 microM triclosan. The kinetics of 3-OH-BaP sulfonation and glucuronidation were examined in the presence of varying concentrations of triclosan: the inhibition of sulfonation was noncompetitive, whereas that of glucuronidation was competitive. These findings reveal that the commonly used bactericide triclosan is a selective inhibitor of the glucuronidation and sulfonation of phenolic xenobiotics. PMID- 15269186 TI - Tocotrienols activate the steroid and xenobiotic receptor, SXR, and selectively regulate expression of its target genes. AB - Vitamin E is an essential nutrient with antioxidant activity. Vitamin E is comprised of eight members, alpha-, beta-, gamma-, and delta-tocopherols and alpha-, beta-, gamma-, and delta-tocotrienols. All forms of vitamin E are initially metabolized by omega-oxidation, which is catalyzed by cytochrome P450 enzymes. The steroid and xenobiotic receptor (SXR) is a nuclear receptor that regulates drug clearance in the liver and intestine via induction of genes involved in drug and xenobiotic metabolism. We show here that all four tocotrienols specifically bind to and activate SXR, whereas tocopherols neither bind nor activate. Surprisingly, tocotrienols show tissue-specific induction of SXR target genes, particularly CYP3A4. Tocotrienols up-regulate expression of CYP3A4 but not UDP-glucuronosyltransferase 1A1 (UGT1A1) or multidrug resistance protein-1 (MDR1) in primary hepatocytes. In contrast, tocotrienols induce MDR1 and UGT1A1 but not CYP3A4 expression in intestinal LS180 cells. We found that nuclear receptor corepressor (NCoR) is expressed at relatively high levels in intestinal LS180 cells compared with primary hepatocytes. The unliganded SXR interacts with NCoR, and this interaction is only partially disrupted by tocotrienols. Expression of a dominant-negative NCoR enhanced the ability of tocotrienols to induce CYP3A4 in LS180 cells, suggesting that NCoR plays an important role in tissue-specific gene regulation by SXR. Our findings provide a molecular mechanism explaining how vitamin supplements affect the absorption and effectiveness of drugs. Knowledge of drug-nutrient interactions may help reduce the incidence of decreased drug efficacy. PMID- 15269187 TI - Quantitative structure-metabolism relationship modeling of metabolic N dealkylation reaction rates. AB - It is widely recognized that preclinical drug discovery can be improved via the parallel assessment of bioactivity, absorption, distribution, metabolism, excretion, and toxicity properties of molecules. High-throughput computational methods may enable such assessment at the earliest, least expensive discovery stages, such as during screening compound libraries and the hit-to-lead process. As an attempt to predict drug metabolism and toxicity, we have developed an approach for evaluation of the rate of N-dealkylation mediated by two of the most important human cytochrome P450s (P450), namely CYP3A4 and CYP2D6. We have taken a novel approach by using descriptors generated for the whole molecule, the reaction centroid, and the leaving group, and then applying neural network computations and sensitivity analysis to generate quantitative structure metabolism relationship models. The quality of these models was assessed by using the cross-validated correlation coefficients of 0.82 for CYP3A4 and 0.79 for CYP2D6 as well as external test molecules for each enzyme. The relative performance of different neural networks was also compared, and modular neural networks with two hidden layers provided the best predictive ability. Functional dependencies between the neural network input and output variables, generalization ability, and limitations of the described approach are also discussed. These models represent an initial approach to predicting the rate of P450-mediated metabolism and may be applied and integrated with other models for P450 binding to produce a systems-based approach for predicting drug metabolism. PMID- 15269188 TI - Identification of the cytosolic carboxylesterase catalyzing the 5'-deoxy-5 fluorocytidine formation from capecitabine in human liver. AB - Capecitabine, a prodrug of 5-fluorouracil, is first metabolized to 5'-deoxy-5 fluorocytidine (5'-DFCR) by carboxylesterase (CES), which is mainly expressed in microsomes. Recently, we clarified that 5'-DFCR formation was catalyzed by the enzyme in cytosol as well as microsomes in human liver. In the present study, the cytosolic enzyme involved in 5'-DFCR formation from capecitabine was identified. This enzyme was purified in the cytosolic preparation by ammonium sulfate precipitation, Sephacryl S-300 gel filtration, Mono P chromatofocusing, and Superdex 200 gel filtration. The purified enzyme was identified by the amino acid sequence analysis to be CES1A1 or a CES1A1 precursor. Based on the result of the N-terminal amino acid sequence analysis, the purified enzyme has no putative signal peptide, indicating that it was CES1A1. The apparent Km and Vmax values of 5'-DFCR formation were 19.2 mM and 88.3 nmol/min/mg protein, respectively. The 5' DFCR formation catalyzed by the purified enzyme was inhibited by both diisopropylfluorophosphate and bis(p-nitrophenyl)phosphate in a concentration dependent manner. 7-Ethyl-10-hydroxycamptothecin (SN-38) formation from irinotecan also occurred in the purified enzyme, cytosol, and microsomes. In conclusion, the cytosolic enzyme involved in 5'-DFCR formation from capecitabine would be CES1A1. It is suggested that the cytosolic CES has significant hydrolysis activity and plays an important role as the microsomal CES in drug metabolism. It is worthy to investigate the metabolic enzyme in cytosol involved in the activation of ester-type prodrugs such as capecitabine. PMID- 15269189 TI - Identification of di-(2-ethylhexyl) phthalate-induced carboxylesterase 1 in C57BL/6 mouse liver microsomes: purification, cDNA cloning, and baculovirus mediated expression. AB - Several mouse carboxylesterase (CES) isozymes have been identified, but information about their roles in drug metabolism is limited. In this study, we purified and characterized a mouse CES1 isozyme that was induced by di-(2 ethylhexyl) phthalate. Purified mouse CES1 shared some biological characteristics with other CES isozymes, such as molecular weight of a subunit and isoelectronic point. In addition, purified mouse CES1 behaved as a trimer, a specific characteristic of CES1A subfamily isozymes. The purified enzyme possessed temocapril hydrolase activity, and it was found to contribute significantly to temocapril hydrolase activity in mouse liver microsomes. To identify the nucleotide sequences coding mouse CES1, antibody screening of a cDNA library was performed. The deduced amino acid sequence of the obtained cDNA, mCES1, exhibited striking similarity to those of CES1A isozymes. When expressed in Sf9 cells, recombinant mCES1 showed hydrolytic activity toward temocapril, as did purified mouse CES1. Based on these results, together with the findings that recombinant mouse CES1 had the same molecular weight of a subunit, the same isoelectronic point, and the same native protein mass as those of purified mouse CES1, it was concluded that mCES1 encoded mouse CES1. Furthermore, tissue expression profiles of mCES1 were found to be very similar to those of the human CES1 isozyme. This finding, together with our other results, suggests that mCES1 shares many biological properties with the human CES1 isozyme. The present study has provided useful information for study of metabolism and disposition of ester-prodrugs as well as ester-drugs. PMID- 15269190 TI - Plasma pharmacokinetics and metabolism of the histone deacetylase inhibitor trichostatin a after intraperitoneal administration to mice. AB - Trichostatin A is a potent and specific histone deacetylase inhibitor with promising antitumor activity in preclinical models. Plasma pharmacokinetics of trichostatin A were studied following single-dose intraperitoneal administration of 80 mg/kg (high dose) or 0.5 mg/kg (low dose) to female BALB/c mice. Plasma trichostatin A concentrations were quantified by high performance liquid chromatography (HPLC)-UV assay (high dose) or by HPLC-multiple reaction monitoring assay (low dose). Trichostatin A was rapidly absorbed from the peritoneum and detectable in plasma within 2 min. Cmax of 40 microg/ml and 8 ng/ml occurred within 5 min, followed by rapid exponential decay in plasma trichostatin A concentration with t1/2 of 6.3 min and 9.6 min (high and low doses, respectively). Phase I metabolites at the high dose were identified by simultaneous UV and positive ion electrospray mass spectrometry. Trichostatin A underwent extensive metabolism: primary metabolic pathways were N-demethylation, reduction of the hydroxamic acid to the corresponding trichostatin A amide, and oxidative deamination to trichostatic acid. N-Monomethyl trichostatin A amide was the major plasma metabolite. No didemethylated compounds were identified. Trichostatic acid underwent further biotransformation: reduction and beta oxidation of the carboxylic acid, with or without N-demethylation, resulted in formation of dihydro trichostatic acid and dinor dihydro trichostatic acids. HPLC fractions corresponding to trichostatin A and N-demethylated trichostatin A exhibited histone deacetylase-inhibitory activity; no other fractions were biologically active. We conclude that trichostatin A is rapidly and extensively metabolized in vivo following intraperitoneal administration to mice, and N demethylation does not compromise histone deacetylase-inhibitory activity. PMID- 15269191 TI - Clinical and bacteriological efficacy and safety of 5 and 7 day regimens of telithromycin once daily compared with a 10 day regimen of clarithromycin twice daily in patients with mild to moderate community-acquired pneumonia. AB - OBJECTIVES: This study was conducted to investigate the potential equivalence in clinical efficacy and assess safety of a 5 or 7 day regimen of oral telithromycin (800 mg once daily) and a 10 day regimen of oral clarithromycin (500 mg twice daily) in treating community-acquired pneumonia (CAP). Bacteriological efficacy was also compared. METHODS: This was a multicentre, randomized, double-blind, active-controlled study. Patients with mild to moderate CAP received telithromycin 800 mg once a day for 5 (n=193) or 7 (n=195) days or clarithromycin 500 mg twice a day for 10 days (n=187). In these groups, 159, 161 and 146 patients, respectively, completed the study. RESULTS: At the post-therapy/test-of cure evaluation, clinical cure rates (per-protocol clinical population) were 89.3% (5 days) and 88.8% (7 days) for telithromycin, and 91.8% for clarithromycin 10 days. Satisfactory bacteriological outcome rates (per-protocol bacteriological population) were 87.7% and 80.0% for 5 and 7 days of telithromycin, respectively, and 83.3% for 10 days of clarithromycin. Bacteriological eradication rates in the respective treatment groups were, for Streptococcus pneumoniae, 95.8% (23/24), 96.7% (29/30) and 88.5% (23/26); for Haemophilus influenzae, 88.0% (22/25), 84.0% (21/25) and 88.2% (15/17) and for Moraxella catarrhalis, 1/1, 4/5 and 3/4. Both telithromycin regimens demonstrated clinical efficacy against pneumococcal bacteraemia (19/19), atypical pathogens (9/9) and erythromycin-resistant S. pneumoniae isolates (5/5). Most treatment-emergent adverse events were mild to moderate in intensity with most commonly reported adverse events involving the gastrointestinal system. CONCLUSIONS: Telithromycin 800 mg administered once a day for 5 or 7 days was as effective and safe as clarithromycin 500 mg administered twice a day for 10 days in treating patients with CAP caused by common respiratory pathogens, including macrolide-resistant isolates, and pneumococcal bacteraemia. PMID- 15269192 TI - Development of a prescribing indicator for objective quantification of antibiotic usage in secondary care. AB - OBJECTIVES: To compare the recognized defined daily dose per 100 bed-days (DDD/100 bed-days) measure with the defined daily dose per finished consultant episode (DDD/FCE) in a group of hospitals with a variety of medicines management strategies. To compare antibiotic usage using the above indicators in hospitals with and without electronic prescribing systems. METHODS: Twelve hospitals were used in the study. Nine hospitals were selected and split into three cohorts (three high-scoring, three medium-scoring and three low-scoring) by their 2001 medicines management self-assessment scores (MMAS). An additional cohort of three electronic prescribing hospitals was included for comparison. MMAS were compared to antibiotic management scores (AMS) developed from a questionnaire relating specifically to control of antibiotics. FCEs and occupied bed-days were obtained from published statistics and statistical analyses of the DDD/100 bed-days and DDD/FCE were carried out using SPSS. RESULTS: The DDD/100 bed-days varied from 81.33 to 189.37 whilst the DDD/FCE varied from 2.88 to 7.43. The two indicators showed a high degree of correlation with r=0.74. MMAS were from 9 to 22 (possible range 0-23) and the AMS from 2 to 13 (possible range 0-22). The two scores showed a high degree of correlation with r=0.74. No correlation was established between either indicator and either score. CONCLUSIONS: The WHO indicator for medicines utilization, DDD/100 bed-days, exhibited the same level of conformity as that exhibited from the use of the DDD/FCE indicating that the DDD/FCE is a useful additional indicator for identifying hospitals which require further study. The MMAS can be assumed to be an accurate guide to antibiotic medicines management controls. No relationship has been found between a high degree of medicines management control and the quantity of antibiotic prescribed. PMID- 15269193 TI - Epidemiology and antibiotic resistance of group A streptococci isolated from healthy schoolchildren in Korea. AB - OBJECTIVES: To assess the epidemiology, antibiotic resistance and genotypic characterization of group A streptococci (GAS) in Jinju, Korea. METHODS: Isolates were characterized in terms of their antibiotic resistance, the phenotypes of erythromycin resistance, the frequencies of erm(B), erm(A) and mef(A) genes, and by emm genotyping or M typing. The data were compared with those from 85 GAS strains collected during 1995 in the same area. RESULTS: A total of 98 (16.9%) of 581 healthy schoolchildren yielded GAS from throat swab culture during 2002. The most frequent emm types were emm12 (34.4%), followed by emm75 (10.4%), emm18 (9.4%), emm22 (8.3%) and emm1 (7.3%) in 2002, whereas M12 (21.2%) and M22 (14.1%) were common in 1995. The resistance rates to erythromycin and clindamycin in 2002 were 51.0% and 33.7%, respectively, compared with 29.4% and 10.1% in 1995. Among the erythromycin-resistant strains, constitutive resistance, inducible resistance, and the M phenotype were observed in 61.2%, 2.0% and 36.7% in 2002, compared with 64.0%, 0% and 36.0% in 1995, respectively, which correlated with the presence of resistance genes. Most of the emm12 strains showed constitutive resistance, whereas emm18 and emm75 showed the M phenotype. The organisms with other emm genotypes were susceptible to both erythromycin and clindamycin. CONCLUSIONS: Erythromycin and clindamycin resistance increased markedly during the period 1995-2002 in Korea. Constitutive resistance is more common than the M phenotype, with inducible resistance occurring rarely. The phenotypes of erythromycin resistance seem to be associated with certain emm genotypes. PMID- 15269194 TI - Detection and assay of beta-lactamases in clinical and non-clinical strains of Yersinia enterocolitica biovar 1A. AB - OBJECTIVES: To detect beta-lactamases (A & B) and extended-spectrum beta lactamases (ESBLs) in clinical and non-clinical isolates of Yersinia enterocolitica biovar 1A, and to determine their activity in the presence of specific lactamase inhibitors. METHODS: The presence of beta-lactamases and ESBLs was detected by disc diffusion in 219 (36 clinical, 183 non-clinical) isolates. beta-Lactamase activity was assayed spectrophotometrically in all 36 clinical and 10 representative non-clinical isolates using nitrocefin as the substrate. Inhibition of beta-lactamases was studied by clavulanic acid, aztreonam and cloxacillin. RESULTS: Of the 219 isolates, all except two non-clinical isolates indicated the presence of beta-lactamase A (Bla-A) based on the smaller (2-8 mm) radius of the inhibition zone around the ticarcillin disc. Synergy between ticarcillin and co-amoxiclav discs was, however, observed in only 34% of isolates of non-clinical origin. beta-Lactamase B (Bla-B) was found to be consistently positive among all the clinical and non-clinical isolates, as indicated by its characteristic appearance of flattening of the zone of inhibition around the cefotaxime disc adjacent to an imipenem disc. Bla-B was induced more strongly in clinical than in non-clinical isolates. Inhibition of enzyme A by clavulanic acid, aztreonam and cloxacillin was found to be similar, whereas enzyme B was inhibited more strongly by aztreonam and cloxacillin. None of the isolates showed the unequivocal presence of ESBL. CONCLUSION: This is the first report on beta lactamases of Yersinia enterocolitica biovar 1A from Asia. Y. enterocolitica biovar 1A expressed both Bla-A and Bla-B. Heterogeneity was, however, discerned in the expression of Bla-A and by induction of Bla-B among clinical and non clinical isolates of Y. enterocolitica biovar 1A. PMID- 15269195 TI - Combination therapy with polymyxin B for the treatment of multidrug-resistant Gram-negative respiratory tract infections. AB - BACKGROUND: The treatment of infections caused by multidrug-resistant (MDR) Gram negative organisms poses a therapeutic challenge. The use of polymyxin B has been resurrected specifically for this purpose. PATIENTS AND METHODS: We retrospectively reviewed the clinical and microbiological efficacy, and safety profile of polymyxin B in the treatment of MDR Gram-negative bacterial infections of the respiratory tract. Twenty-five critically ill patients received a total of 29 courses of polymyxin B administered in combination with another antimicrobial agent. RESULTS: Patients were treated with intravenous, and/or aerosolized polymyxin B. Mean duration of polymyxin B therapy was 19 days (range 2-57 days). End of treatment mortality was 21%, and overall mortality at discharge was 48%. Nephrotoxicity was observed in three patients (10%) and did not result in discontinuation of therapy. CONCLUSIONS: Polymyxin B in combination with other antimicrobials can be considered a reasonable and safe treatment option for MDR Gram-negative respiratory tract infections in the setting of limited therapeutic options. PMID- 15269196 TI - Effect of gum arabic on the absorption of a single oral dose of amoxicillin in healthy Sudanese volunteers. PMID- 15269197 TI - Survey, characterization and susceptibility to fusidic acid of Staphylococcus aureus in the Carmarthen area. AB - OBJECTIVE: This retrospective study was designed to investigate the possible reasons for an apparent increase in fusidic acid resistance among Staphylococcus aureus. MATERIALS AND METHODS: The Datastore records of the Communicable Disease Surveillance Centre, Wales, UK were reviewed in conjunction with information concerning the prescribing of fusidic acid. RESULTS: During the 5 year study period (1997-2001), a rise in the incidence of fusidic acid resistance was noted, particularly among paediatric patients presenting with infected eczema and impetigo, which may be related to the observed increase in prescriptions of topical fusidic acid. Extended phenotypic and genotypic characterization of a limited number (n=31) of isolates from 2002 showed that fusidic acid-resistant strains of S. aureus were typically from patients with impetigo and isolates fell into a single clonal group. Conversely, isolates from other skin disease (eczema, dermatitis and abscesses) were usually susceptible to fusidic acid and proved a diverse group. CONCLUSION: This study provides valuable data on the prevalence of fusidic acid-resistant S. aureus, the genetic background of the strains, and their association with clinical disease in both the healthcare environment and community setting in the catchment area served by the Laboratory. PMID- 15269198 TI - Effect of social and climatological factors on antimicrobial use and Streptococcus pneumoniae resistance in different provinces in Spain. AB - OBJECTIVES: To investigate the association between geographical differences in antibiotic consumption and resistance of Streptococcus pneumoniae to penicillin and erythromycin in 15 provinces of Spain, taking into account the potential influence of a series of social and climatological factors. METHODS: Possible correlations between prevalence of resistance to penicillin and erythromycin of S. pneumoniae, as determined in the national reference laboratory, and antibiotic consumption, and socio-economic and climatological variables were investigated. Partial correlations and multivariate linear regression were performed to assess the relative importance of variables predicting resistance and to investigate explicative factors for antibiotic consumption, respectively. RESULTS: A correlation was found between resistance and educational level, the proportion of young people in the population and climate, but was explained by their effects on differences in antibiotic use, which appeared to be the basic and only force behind resistance patterns in different geographical areas. Antibiotic use was found to be determined by the interplay of adult illiteracy, rainfall and GDP per capita. CONCLUSIONS: Interventions aimed at improving educational level and economic growth might therefore be followed by a noticeable reduction in overall antibiotic consumption, which might in turn be followed by a reduction in penicillin and erythromycin resistance in clinical isolates of S. pneumoniae. PMID- 15269199 TI - Effect of triclosan or a phenolic farm disinfectant on the selection of antibiotic-resistant Salmonella enterica. AB - OBJECTIVE: To determine the effect of growth of five strains of Salmonella enterica and their isogenic multiply antibiotic-resistant (MAR) derivatives with a phenolic farm disinfectant or triclosan (biocides) upon the frequency of mutation to resistance to antibiotics or cyclohexane. METHODS: Strains were grown in broth with or without the biocides and then spread on to agar containing ampicillin, ciprofloxacin or tetracycline each at 4x MIC or agar overlaid with cyclohexane. Incubation was for 24 and 48 h and the frequency of mutation to resistance was calculated for strains with and without prior growth with the biocides. MICs were determined and the presence of mutations in the acrR and marR regions was determined by sequencing and the presence of mutations in gyrA by light-cycler analysis, for a selection of the mutants that arose. RESULTS: The mean frequency of mutation to antibiotic or cyclohexane resistance was increased approximately 10- to 100-fold by prior growth with the phenolic disinfectant or triclosan. The increases were statistically significant for all antibiotics and cyclohexane following exposure to the phenolic disinfectant (P 1 mg/L ciprofloxacin arose only from strains that were MAR. Reduced susceptibility to ciprofloxacin (at 4x MIC for parent strains) alone was associated with mutations in gyrA. MAR mutants did not contain mutations in the acrR or marR region. CONCLUSIONS: These data renew fears that the use of biocides may lead to an increased selective pressure towards antibiotic resistance. PMID- 15269200 TI - Protease inhibitor-sparing simplified maintenance therapy: a need for perspective. AB - Body fat changes and metabolic abnormalities such as hyperlipidaemia and diabetes have been increasingly reported following the successful introduction of highly active antiretroviral therapy (HAART). These side effects were attributed initially to the use of protease inhibitors (PIs). As a consequence, a series of trials were conducted where patients with well-controlled HIV viraemia either continued on PIs or were switched to a simplified maintenance therapy (SMT) without PIs. Evidence from these trials is still insufficient to show that switching from PIs to either abacavir, nevirapine or efavirenz is safe. However, patients with suboptimal pre-HAART treatment are at increased risk of virological failure if switched to an SMT. Patients switched from PI regimens tend to stay longer on an SMT and those switched to abacavir show a reduction in total cholesterol, but there is no evidence of any additional benefit from non-PI-based SMT. There is a clear need for a better understanding of HAART-related lipodystrophy and metabolic toxicity, and pharmacogenetic tests to identify those patients most at risk. The advent of simpler formulations for all drug classes, and new PIs with less metabolic toxicity, is likely to reshape completely the role of SMT. PMID- 15269201 TI - Demethylation of 3-methylthymine in DNA by bacterial and human DNA dioxygenases. AB - Rare DNA lesions that are chemically stable and refractory to repair may add disproportionately to the accumulation of mutations in long lived cells. 3 Methylthymine is a minor lesion that is induced by DNA-methylating agents and for which no repair process has been described previously. Here we demonstrate that this lesion can be directly demethylated in vitro by bacterial and human DNA dioxygenases. The Escherichia coli AlkB and human ABH3 proteins repaired 3 methylthymine in both single-stranded and double-stranded polydeoxynucleotides, whereas the human ABH2 protein preferred a duplex substrate. Thus, the known substrates of these enzymes now include 3-methylthymine in DNA, as well as 1 methyladenine and 3-methylcytosine, which all have structurally similar sites of alkylation. Repair of 3-methylthymine by AlkB and ABH3 was optimal at pH 6, but inefficient. At physiological pH, 3-methylthymine, which is a minor methylated lesion, was more slowly repaired than the major lesion generated in single stranded DNA, 3-methylcytosine. Our data suggest that 3-methylthymine residues in DNA will be repaired inefficiently in vivo and therefore may occur at a low steady-state level, but the residues should not gradually accumulate to high levels in long lived cells. PMID- 15269202 TI - Role of MAPK phosphatase-1 (MKP-1) in adipocyte differentiation. AB - Both time-dependent modulation of intracellular signaling molecules and sequential induction of transcriptional regulators are essential for the differentiation of preadipocytes into adipocytes. We have now shown that the activity, but not the abundance, of p42/p44 mitogen-activated protein kinase (MAPK) is down-regulated during adipocyte differentiation. This decrease in p42/p44 MAPK activity does not appear to be a direct effect of hormonal inducers of differentiation but rather represents a characteristic event of adipocyte differentiation that is achieved through a persistent change in intracellular signaling. Although the phosphorylation or abundance of MEK, an upstream kinase for p42/p44 MAPK, was not altered during differentiation, the abundance of MAPK phosphatase-1 (MKP-1), a negative regulator of p42/p44 MAPK, was increased with a time course similar to that of the down-regulation of p42/p44 MAPK activity. Ectopic expression of MKP-1 in preadipocytes reduced and depletion of endogenous MKP-1 in mature adipocytes increased the activity of p42/p44 MAPK. Prevention of the up-regulation of MKP-1 abundance in preadipocytes by expression of Mkp-1 antisense RNA resulted in persistence of p42/p44 MAPK activation and blocked differentiation, effects that were reversed by the MEK inhibitor PD98059. These results suggest that MKP-1 plays an essential role in adipocyte differentiation through down-regulation of p42/p44 MAPK activity. PMID- 15269203 TI - ATM-dependent CHK2 activation induced by anticancer agent, irofulven. AB - Irofulven (6-hydroxymethylacylfulvene, HMAF, MGI 114) is one of a new class of anticancer agents that are semisynthetic derivatives of the mushroom toxin illudin S. Preclinical studies and clinical trials have demonstrated that irofulven is effective against several tumor types. Mechanisms of action studies indicate that irofulven induces DNA damage, MAPK activation, and apoptosis. In this study we found that in ovarian cancer cells, CHK2 kinase is activated by irofulven while CHK1 kinase is not activated even when treated at higher concentrations of the drug. By using GM00847 human fibroblast expressing tetracycline-controlled, FLAG-tagged kinase-dead ATR (ATR.kd), it was demonstrated that ATR kinase does not play a major role in irofulven-induced CHK2 activation. Results from human fibroblasts proficient or deficient in ATM function (GM00637 and GM05849) indicated that CHK2 activation by irofulven is mediated by the upstream ATM kinase. Phosphorylation of ATM on Ser(1981), which is critical for kinase activation, was observed in ovarian cancer cell lines treated with irofulven. RNA interference results confirmed that CHK2 activation was inhibited after introducing siRNA for ATM. Finally, experiments done with human colon cancer cell line HCT116 and its isogenic CHK2 knockout derivative; and experiments done by expressing kinase-dead CHK2 in an ovarian cancer cell line demonstrated that CHK2 activation contributes to irofulven-induced S phase arrest. In addition, it was shown that NBS1, SMC1, and p53 were phosphorylated in an ATM-dependent manner, and p53 phosphorylation on serine 20 is dependent on CHK2 after irofulven treatment. In summary, we found that the anticancer agent, irofulven, activates the ATM-CHK2 DNA damage-signaling pathway, and CHK2 activation contributes to S phase cell cycle arrest induced by irofulven. PMID- 15269204 TI - Three-dimensional structure of the vacuolar ATPase. Localization of subunit H by difference imaging and chemical cross-linking. AB - The structure of the proton-pumping vacuolar ATPase (V-ATPase) from bovine brain clathrin coated vesicles was analyzed by electron microscopy and single molecule image analysis. A three-dimensional structural model of the complex was calculated by the angular reconstitution method at a resolution of 27 A. Overall, the appearance of the V(0) and V(1) domains in the three-dimensional model of the intact bovine V-ATPase resembles the models of the isolated bovine V(0) and yeast V(1) domains determined previously. To determine the binding position of subunit H in the V-ATPase, electron microscopy and cysteine-mediated photochemical cross linking were used. Difference maps calculated from projection images of intact bovine V-ATPase and a V-ATPase preparation in which the two H subunit isoforms were removed by treatment with cystine revealed less protein density at the bottom of the V(1) in the subunit H-depleted enzyme, suggesting that subunit H isoforms bind at the interface of the V(1) and V(0) domains. A comparison of three-dimensional models calculated for intact and subunit H-depleted enzyme indicated that at least one of the subunit H isoforms, although poorly resolved in the three-dimensional electron density, binds near the putative N-terminal domain of the a subunit of the V(0). For photochemical cross-linking, unique cysteine residues were introduced into the yeast V-ATPase B subunit at sites that were localized based on molecular modeling using the crystal structure of the mitochondrial F(1) domain. Cross-linking was performed using the photoactivatable sulfhydryl reagent 4-(N-maleimido)benzophenone. Cross-linking to subunit H was observed from two sites on subunit B (E494 and T501) predicted to be located on the outer surface of the subunit closest to the membrane. Results from both electron microscopy and cross-linking analysis thus place subunit H near the interface of the V(1) and V(0) domains and suggest a close structural similarity between the V-ATPases of yeast and mammals. PMID- 15269205 TI - The crystal structure of Escherichia coli MoaB suggests a probable role in molybdenum cofactor synthesis. AB - The crystal structure of Escherichia coli MoaB was determined by multiwavelength anomalous diffraction phasing and refined at 1.6-A resolution. The molecule displayed a modified Rossman fold. MoaB is assembled into a hexamer composed of two trimers. The monomers have high structural similarity with two proteins, MogA and MoeA, from the molybdenum cofactor synthesis pathway in E. coli, as well as with domains of mammalian gephyrin and plant Cnx1, which are also involved in molybdopterin synthesis. Structural comparison between these proteins and the amino acid conservation patterns revealed a putative active site in MoaB. The structural analysis of this site allowed to advance several hypothesis that can be tested in further studies. PMID- 15269206 TI - The histone octamer is invisible when NF-kappaB binds to the nucleosome. AB - The transcription factor NF-kappaB is involved in the transcriptional control of more than 150 genes, but the way it acts at the level of nucleosomal templates is not known. Here we report on a study examining the interaction of NF-kappaB p50 with its DNA recognition sequence in a positioned nucleosome. We demonstrate that NF-kappaB p50 was able to bind to the nucleosome with an apparent association constant close to that for free DNA. In agreement with this, the affinity of NF kappaB p50 binding does not depend on the localization of its recognition sequence relative to the nucleosome dyad axis. In addition, the binding of NF kappaB p50 does not induce eviction of histones and does not perturb the overall structure of the nucleosome. The NF-kappaB p50-nucleosome complex exhibits, however, local structural alterations within the NF-kappaB p50 recognition site. Importantly, these alterations were very similar to those found in the NF-kappaB p50-DNA complex. Our data suggest that NF-kappaB p50 can accommodate the distorted, bent DNA within the nucleosome. This peculiar property of NF-kappaB p50 might have evolved to meet the requirements for its function as a central switch for stress responses. PMID- 15269207 TI - Cryptosporidium parvum IMP dehydrogenase: identification of functional, structural, and dynamic properties that can be exploited for drug design. AB - The protozoan parasite Cryptosporidium parvum causes severe enteritis with substantial morbidity and mortality among AIDS patients and young children. No fully effective treatment is available. C. parvum relies on inosine 5' monophosphate dehydrogenase (IMPDH) to produce guanine nucleotides and is highly susceptible to IMPDH inhibition. Furthermore, C. parvum obtained its IMPDH gene by lateral transfer from an epsilon-proteobacterium, suggesting that the parasite enzyme might have very different characteristics than the human counterpart. Here we describe the expression of recombinant C. parvum IMPDH in an Escherichia coli strain lacking the bacterial homolog. Expression of the parasite gene restores growth of this mutant on minimal medium, confirming that the protein has IMPDH activity. The recombinant protein was purified to homogeneity and used to probe the enzyme's mechanism, structure, and inhibition profile in a series of kinetic experiments. The mechanism of the C. parvum enzyme involves the random addition of substrates and ordered release of products with rate-limiting hydrolysis of a covalent enzyme intermediate. The pronounced resistance of C. parvum IMPDH to mycophenolic acid inhibition is in strong agreement with its bacterial origin. The values of Km for NAD and Ki for mycophenolic acid as well as the synergistic interaction between tiazofurin and ADP differ significantly from those of the human enzymes. These data suggest that the structure and dynamic properties of the NAD binding site of C. parvum IMPDH can be exploited to develop parasite specific inhibitors. PMID- 15269208 TI - Crystal structure and biological implications of a bacterial albumin binding module in complex with human serum albumin. AB - Many bactericide species express surface proteins that interact with human serum albumin (HSA). Protein PAB from the anaerobic bacterium Finegoldia magna (formerly Peptostreptococcus magnus) represents one of these proteins. Protein PAB contains a domain of 53 amino acid residues known as the GA module. GA homologs are also found in protein G of group C and G streptococci. Here we report the crystal structure of HSA in complex with the GA module of protein PAB. The model of the complex was refined to a resolution of 2.7 A and reveals a novel binding epitope located in domain II of the albumin molecule. The GA module is composed of a left-handed three-helix bundle, and residues from the second helix and the loops surrounding it were found to be involved in HSA binding. Furthermore, the presence of HSA-bound fatty acids seems to influence HSA-GA complex formation. F. magna has a much more restricted host specificity compared with C and G streptococci, which is also reflected in the binding of different animal albumins by proteins PAB and G. The structure of the HSA-GA complex offers a molecular explanation to this unusually clear example of bacterial adaptation. PMID- 15269209 TI - Naturally occurring proteolytic antibodies: selective immunoglobulin M-catalyzed hydrolysis of HIV gp120. AB - We report the selective catalytic cleavage of the HIV coat protein gp120, a B cell superantigen, by IgM antibodies (Abs) from uninfected humans and mice that had not been previously exposed to gp120. The rate of IgM-catalyzed gp120 cleavage was greater than of other polypeptide substrates, including the bacterial superantigen protein A. The kinetic parameters of gp120 cleavage varied over a broad range depending on the source of the IgMs, and turnover numbers as great as 2.1/min were observed, suggesting that different Abs possess distinct gp120 recognition properties. IgG Abs failed to cleave gp120 detectably. The Fab fragment of a monoclonal IgM cleaved gp120, suggesting that the catalytic activity belongs to the antibody combining site. The electrophoretic profile of gp120 incubated with a monoclonal human IgM suggested hydrolysis at several sites. One of the cleavage sites was identified as the Lys(432)-Ala(433) peptide bond, located within the region thought to be the Ab-recognizable superantigenic determinant. A covalently reactive peptide analog (CRA) corresponding to gp120 residues 421-431 with a C-terminal amidino phosphonate diester mimetic of the Lys(432)-Ala(433) bond was employed to probe IgM nucleophilic reactivity. The peptidyl CRA inhibited the IgM-catalyzed cleavage of gp120 and formed covalent IgM adducts at levels exceeding a control hapten CRA devoid of the peptide sequence. These observations suggest that IgMs can selectively cleave gp120 by a nucleophilic mechanism and raise the possibility of their role as defense enzymes. PMID- 15269210 TI - Crystal structure of the cytochrome p450cam mutant that exhibits the same spectral perturbations induced by putidaredoxin binding. AB - The cytochrome P450cam active site is known to be perturbed by binding to its redox partner, putidaredoxin (Pdx). Pdx binding also enhances the camphor monooxygenation reaction (Nagano, S., Shimada, H., Tarumi, A., Hishiki, T., Kimata-Ariga, Y., Egawa, T., Suematsu, M., Park, S.-Y., Adachi, S., Shiro, Y., and Ishimura, Y. (2003) Biochemistry 42, 14507-14514). These effects are unique to Pdx because nonphysiological electron donors are unable to support camphor monooxygenation. The accompanying 1H NMR paper (Tosha, T., Yoshioka, S., Ishimori, K., and Morishima, I. (2004) J. Biol. Chem. 279, 42836-42843) shows that the conformation of active site residues, Thr-252 and Cys-357, and the substrate in the ferrous (Fe(II)) CO complex of the L358P mutant mimics that of the wild-type enzyme complexed to Pdx. To explore how these changes are transmitted from the Pdx-binding site to the active site, we have solved the crystal structures of the ferrous and ferrous-CO complex of wild-type and the L358P mutant. Comparison of these structures shows that the L358P mutation results in the movement of Arg-112, a residue known to be important for putidaredoxin binding, toward the heme. This change could optimize the Pdx binding site leading to a higher affinity for Pdx. The mutation also pushes the heme toward the substrate and ligand binding pocket, which relocates the substrate to a position favorable for regio-selective hydroxylation. The camphor is held more firmly in place as indicated by a lower average temperature factor. Residues involved in the catalytically important proton shuttle system in the I helix are also altered by the mutation. Such conformational alterations and the enhanced reactivity of the mutant oxy complex with non-physiological electron donors suggest that Pdx binding optimizes the distal pocket for monooxygenation of camphor. PMID- 15269211 TI - L358P mutation on cytochrome P450cam simulates structural changes upon putidaredoxin binding: the structural changes trigger electron transfer to oxy P450cam from electron donors. AB - To investigate the functional and structural characterization of a crucial cytochrome P450cam (P450cam)-putidaredoxin (Pdx) complex, we utilized a mutant whose spectroscopic property corresponds to the properties of the wild type P450cam in the presence of Pdx. The 1H NMR spectrum of the carbonmonoxy adduct of the mutant, the Leu-358 --> Pro mutant (L358P), in the absence of Pdx showed that the ring current-shifted signals arising from d-camphor were upfield-shifted and observed as resolved signals, which are typical for the wild type enzyme in the presence of Pdx. Signals from the beta-proton of the axial cysteine and the gamma methyl group of Thr-252 were also shifted upfield and down-field, respectively, in the L358P mutant as observed for Pdx-bound wild type P450cam. The close similarity in the NMR spectra suggests that the heme environment of the L358P mutant mimics that of the Pdx-bound enzyme. The functional analysis of the L358P mutant has revealed that the oxygen adduct of the L358P mutant can promote the oxygenation reaction for d-camphor with nonphysiological electron donors such as dithionite and ascorbic acid, showing that oxygenated L358P is "activated" to receive electron from the donor. Based on the structural and functional characterization of the L358P mutant, we conclude that the Pdx-induced structural changes in P450cam would facilitate the electron transfer from the electron donor, and the Pdx binding to P450cam would be a trigger for the electron transfer to oxygenated P450cam. PMID- 15269212 TI - HOXB6 protein is bound to CREB-binding protein and represses globin expression in a DNA binding-dependent, PBX interaction-independent process. AB - Although HOXB6 and other HOX genes have previously been associated with hematopoiesis and leukemias, the precise mechanism of action of their protein products remains unclear. Here we use a biological model in which HOXB6 represses alpha- and gamma-globin mRNA levels to perform a structure/function analysis for this homeodomain protein. HOXB6 protein represses globin transcript levels in stably transfected K562 cells in a DNA-binding dependent fashion. However, the capacity to form cooperative DNA-binding complexes with the PBX co-factor protein is not required for HOXB6 biological activity. Neither the conserved extreme N terminal region, a polyglutamic acid region at the protein C terminus, nor the Ser(214) CKII phosphorylation site was required for DNA binding or activity in this model. We have previously reported that HOX proteins can inhibit CREB binding protein (CBP)-histone acetyltransferase-mediated potentiation of reporter gene transcription. We now show that endogenous CBP is co-precipitated with exogenous HOXB6 from nuclear and cytoplasmic compartments of transfected K562 cells. Furthermore, endogenous CBP co-precipitates with endogenous HOXB6 in day 14.5 murine fetal liver cells during active globin gene expression in this tissue. The CBP interaction motif was localized to the homeodomain but does not require the highly conserved helix 3. Our data suggest that the homeodomain contains most or all of the important structures required for HOXB6 activity in blood cells. PMID- 15269213 TI - Mitochondrial oxidative phosphorylation is defective in the long-lived mutant clk 1. AB - The long-lived mutant of Caenorhabditis elegans, clk-1, is unable to synthesize ubiquinone, CoQ(9). Instead, the mutant accumulates demethoxyubiquinone(9) and small amounts of rhodoquinone(9) as well as dietary CoQ(8). We found a profound defect in oxidative phosphorylation, a test of integrated mitochondrial function, in clk-1 mitochondria fueled by NADH-linked electron donors, i.e. complex I dependent substrates. Electron transfer from complex I to complex III, which requires quinones, is severely depressed, whereas the individual complexes are fully active. In contrast, oxidative phosphorylation initiated through complex II, which also requires quinones, is completely normal. Here we show that complexes I and II differ in their ability to use the quinone pool in clk-1. This is the first direct demonstration of a differential interaction of complex I and complex II with the endogenous quinone pool. This study uses the combined power of molecular genetics and biochemistry to highlight the role of quinones in mitochondrial function and aging. PMID- 15269214 TI - A 10-amino acid domain within human T-cell leukemia virus type 1 and type 2 tax protein sequences is responsible for their divergent subcellular distribution. AB - Human T-cell leukemia virus type 1 and type 2 (HTLV-1/2) are related retroviruses that infect T-lymphocytes. Whereas HTLV-1 infection can cause leukemia, HTLV-2 has not been demonstrated to be the agent of a hematological malignant disease. Nevertheless, the virally encoded Tax-1 and Tax-2 transactivators display a high percentage of similarity. Tax-1 is a shuttling protein that contains a noncanonical nuclear localization signal as well as a nuclear export signal. The presence of the nuclear localization signal and the nuclear export signal domains in the Tax-2 sequence has not been determined. The distribution of Tax-2 in infected cells is not known but has been assumed to be similar to that of Tax-1. By using a Tax-2-specific antibody, we report here that Tax-2 is located predominantly in the cytoplasm of the HTLV-2 immortalized or transformed infected T-cells. These results were confirmed after transient transfection of untagged Tax-1 and Tax-2 constructs, histidine tag Tax1/Tax2, GFP-Tax, and Tax-GFP fusion constructs in several cell lines. We show that this unanticipated localization is not due to a default in the Tax-2 nuclear localization signal functions nor to major differences in Tax-2 versus Tax-1 binding to the IKKgamma/NEMO protein. In addition, we demonstrate that inhibiting the proteasome results in a relocalization of Tax-1 in the cytoplasm, similar to that of Tax-2. By using a series of Tax-1/Tax-2 chimeras, we determined that the minimal domain that is necessary for Tax-2 peculiar distribution encompasses amino acids 90-100. Finally, we show a high correlation between intracellular localization of Tax and their NF-kappaB or CREB transactivating ability. PMID- 15269215 TI - gAd-globular head domain of adiponectin increases fatty acid oxidation in newborn rabbit hearts. AB - Adiponectin is an adipocyte-derived hormone that has a number of metabolic effects in the body, including the control of both glucose and fatty acid metabolism. The globular head domain of adiponectin, gAd, has also been shown to increase fatty acid oxidation in skeletal muscle. Within days after birth, a rapid increase in fatty acid oxidation occurs in the heart. We examined whether adiponectin or gAd plays a role in this maturation of cardiac fatty acid oxidation. Plasma adiponectin increased in newborn rabbits following birth: 1.2 +/- 0.3 microg/ml in 1-day-old, 6.8 +/- 1.8 microg/ml in 7-day-old, and 45 +/- 5 microg/ml in 6-week-old rabbits. Because plasma insulin levels decrease and remain low throughout the suckling period, and because this decrease may contribute to the maturation of fatty acid oxidation, we examined the effects of adiponectin and gAd on fatty acid oxidation in isolated perfused 1-day-old rabbit hearts in the presence or absence of 100 microunits/ml insulin. Adiponectin (10 microg/ml) did not alter fatty acid oxidation in the presence of insulin. In the absence of insulin, the addition of recombinant gAd (1.5 microg/ml) increased fatty acid oxidation compared with control (129 +/- 18 versus 66 +/- 11 nmol.g dry weight(-1).min(-1), respectively (p < 0.05). In 7-day-old hearts, where fatty acid oxidation rates were 5-fold higher than 1-day-old hearts, gAd did not alter fatty acid oxidation rates. The increase in fatty acid oxidation in 1-day-old hearts occurred independently of changes in 5'-AMP-activated protein kinase, acetyl-CoA carboxylase, or malonyl-CoA. The effect of gAd on fatty acid oxidation was reversed in the presence of 100 microunits/ml insulin. These results suggest that a decrease in plasma insulin and increase in gAd are involved in the increase of cardiac fatty acid oxidation in the immediate newborn period. PMID- 15269216 TI - Inhibition of mitochondrial Na+-Ca2+ exchange restores agonist-induced ATP production and Ca2+ handling in human complex I deficiency. AB - Human mitochondrial complex I (NADH:ubiquinone oxidoreductase) of the oxidative phosphorylation system is a multiprotein assembly comprising both nuclear and mitochondrially encoded subunits. Deficiency of this complex is associated with numerous clinical syndromes ranging from highly progressive, often early lethal encephalopathies, of which Leigh disease is the most frequent, to neurodegenerative disorders in adult life, including Leber's hereditary optic neuropathy and Parkinson disease. We show here that the cytosolic Ca2+ signal in response to hormonal stimulation with bradykinin was impaired in skin fibroblasts from children between the ages of 0 and 5 years with an isolated complex I deficiency caused by mutations in nuclear encoded structural subunits of the complex. Inhibition of mitochondrial Na+-Ca2+ exchange by the benzothiazepine CGP37157 completely restored the aberrant cytosolic Ca2+ signal. This effect of the inhibitor was paralleled by complete restoration of the bradykinin-induced increases in mitochondrial Ca2+ concentration and ensuing ATP production. Thus, impaired mitochondrial Ca2+ accumulation during agonist stimulation is a major consequence of human complex I deficiency, a finding that may provide the basis for the development of new therapeutic approaches to this disorder. PMID- 15269217 TI - ABCA1 is required for normal central nervous system ApoE levels and for lipidation of astrocyte-secreted apoE. AB - ABCA1 is an ATP-binding cassette protein that transports cellular cholesterol and phospholipids onto high density lipoproteins (HDL) in plasma. Lack of ABCA1 in humans and mice causes abnormal lipidation and increased catabolism of HDL, resulting in very low plasma apoA-I, apoA-II, and HDL. Herein, we have used Abca1 /- mice to ask whether ABCA1 is involved in lipidation of HDL in the central nervous system (CNS). ApoE is the most abundant CNS apolipoprotein and is present in HDL-like lipoproteins in CSF. We found that Abca1-/- mice have greatly decreased apoE levels in both the cortex (80% reduction) and the CSF (98% reduction). CSF from Abca1-/- mice had significantly reduced cholesterol as well as small apoE-containing lipoproteins, suggesting abnormal lipidation of apoE. Astrocytes, the primary producer of CNS apoE, were cultured from Abca1+/+, +/-, and -/- mice, and nascent lipoprotein particles were collected. Abca1-/- astrocytes secreted lipoprotein particles that had markedly decreased cholesterol and apoE and had smaller apoE-containing particles than particles from Abca1+/+ astrocytes. These findings demonstrate that ABCA1 plays a critical role in CNS apoE metabolism. Since apoE isoforms and levels strongly influence Alzheimer's disease pathology and risk, these data suggest that ABCA1 may be a novel therapeutic target. PMID- 15269218 TI - Deficiency of ABCA1 impairs apolipoprotein E metabolism in brain. AB - ABCA1 is a cholesterol transporter that is widely expressed throughout the body. Outside the central nervous system (CNS), ABCA1 functions in the biogenesis of high-density lipoprotein (HDL), where it mediates the efflux of cholesterol and phospholipids to apolipoprotein (apo) A-I. Deficiency of ABCA1 results in lack of circulating HDL and greatly reduced levels of apoA-I. ABCA1 is also expressed in cells within the CNS, but its roles in brain lipid metabolism are not yet fully understood. In the brain, glia synthesize the apolipoproteins involved in CNS lipid metabolism. Here we demonstrate that glial ABCA1 is required for cholesterol efflux to apoA-I and plays a key role in facilitating cholesterol efflux to apoE, which is the major apolipoprotein in the brain. In both astrocytes and microglia, ABCA1 deficiency reduces lipid efflux to exogenous apoE. The impaired ability to efflux lipids in ABCA1-/- glia results in lipid accumulation in both astrocytes and microglia under normal culture conditions. Additionally, apoE secretion is compromised in ABCA1-/- astrocytes and microglia. In vivo, deficiency of ABCA1 results in a 65% decrease in apoE levels in whole brain, and a 75-80% decrease in apoE levels in hippocampus and striatum. Additionally, the effect of ABCA1 on apoE is selective, as apoJ levels are unchanged in brains of ABCA1-/- mice. Taken together, these results show that glial ABCA1 is a key influence on apoE metabolism in the CNS. PMID- 15269219 TI - Coenzyme specificity of Sir2 protein deacetylases: implications for physiological regulation. AB - Sir2 (silent information regulator 2) enzymes catalyze a unique protein deacetylation reaction that requires the coenzyme NAD(+) and produces nicotinamide and a newly discovered metabolite, O-acetyl-ADP-ribose (OAADPr). Conserved from bacteria to humans, these proteins are implicated in the control of gene silencing, metabolism, apoptosis, and aging. Here we examine the role of NAD(+) metabolites/derivatives and salvage pathway intermediates as activators, inhibitors, or coenzyme substrates of Sir2 enzymes in vitro. Also, we probe the coenzyme binding site using inhibitor binding studies and alternative coenzyme derivatives as substrates. Sir2 enzymes showed an exquisite selectivity for the nicotinamide base coenzyme, with the most dramatic losses in binding affinity/reactivity resulting from relatively minor changes in the nicotinamide ring, either by reduction, as in NADH, or by converting the amide to its acid analogue. Both ends of the dinucleotide NAD(+) are shown to be critical for high selectivity and high affinity. Among the NAD(+) metabolites tested none were able to allosterically activate, although all led to various extents of inhibition, consistent with competition at the coenzyme binding site. Nicotinamide was the most potent inhibitor examined, suggesting that cellular nicotinamide levels would provide an effective small molecule regulator of protein deacetylation and generation of OAADPr. The presented findings also suggest that changes in the physiological NAD(+):NADH ratio, without a change in NAD(+), would yield little alteration in Sir2 activity. That is, NADH is an extremely ineffective inhibitor of Sir2 enzymes (average IC(50) of 17 mm). We propose that changes in both free nicotinamide and free NAD(+) afford the greatest contribution to cellular activity of Sir2 enzymes but with nicotinamide having a more dramatic effect during smaller fluctuations in concentration. PMID- 15269220 TI - Both nuclear-cytoplasmic shuttling of the dual specificity phosphatase MKP-3 and its ability to anchor MAP kinase in the cytoplasm are mediated by a conserved nuclear export signal. AB - MAP kinase phosphatase (MKP)-3 is a cytoplasmic dual specificity protein phosphatase that specifically binds to and inactivates the ERK1/2 MAP kinases in mammalian cells. However, the molecular basis of the cytoplasmic localization of MKP-3 or its physiological significance is unknown. We have used MKP-3-green fluorescent protein fusions in conjunction with leptomycin B to show that the cytoplasmic localization of MKP-3 is mediated by a chromosome region maintenance 1 (CRM1)-dependent nuclear export pathway. Furthermore, the nuclear translocation of MKP-3 seen in the presence of leptomycin B is mediated by an active process, indicating that MKP-3 shuttles between the nucleus and cytoplasm. The amino terminal noncatalytic domain of MKP-3 is both necessary and sufficient for nuclear export of the phosphatase and contains a single functional leucine-rich nuclear export signal (NES). Even though this domain of the protein also mediates the binding of MKP-3 to MAP kinase, we show that mutations of the kinase interaction motif which abrogate ERK2 binding do not affect MKP-3 localization. Conversely, mutation of the NES does not affect either the binding or phosphatase activity of MKP-3 toward ERK2, indicating that the kinase interaction motif and NES function independently. Finally, we demonstrate that the ability of MKP-3 to cause the cytoplasmic retention of ERK2 requires both a functional kinase interaction motif and NES. We conclude that in addition to its established function in the regulated dephosphorylation and inactivation of MAP kinase, MKP-3 may also play a role in determining the subcellular localization of its substrate. Our results reinforce the idea that regulatory proteins such as MKP-3 may play a key role in the spatio-temporal regulation of MAP kinase activity. PMID- 15269221 TI - A novel fold revealed by Mycobacterium tuberculosis NAD kinase, a key allosteric enzyme in NADP biosynthesis. AB - NAD kinase catalyzes the magnesium-dependent phosphorylation of NAD, representing the sole source of freshly synthesized NADP in all organisms. The enzyme is essential for the growth of the deadly multidrug-resistant pathogen Mycobacterium tuberculosis and is an attractive target for novel antitubercular agents. The crystal structure of NAD kinase has been solved by multiwavelength anomalous dispersion at a resolution of 2.3 A in its T state. Two crystal forms have been obtained revealing either a dimer or a tetramer. The enzyme architecture discloses a novel molecular arrangement, with each subunit consisting of an alpha/beta N-terminal domain and a C-terminal 12-stranded beta sandwich domain, connected by swapped beta strands. The C-terminal domain shows a striking internal approximate 222 symmetry and an unprecedented topology, revealing a novel fold within the family of all beta structures. The catalytic site is located in the long crevice that defines the interface between the domains. The conserved GGDG structural fingerprint of the catalytic site is reminiscent of the related region in 6-phosphofructokinase, supporting the hypothesis that NAD kinase belongs to a newly reported superfamily of kinases. PMID- 15269222 TI - Differential regulation of matrix metalloproteinase-2 and -9 expression and activity in adult rat cardiac fibroblasts in response to interleukin-1beta. AB - Matrix metalloproteinases (MMPs), a family of endoproteinases, are implicated in cardiac remodeling. Interleukin-1beta (IL-1beta), which is increased in the heart following myocardial infarction, increases expression and activity of MMP-2 (gelatinase A) and -9 (gelatinase B) in cardiac fibroblasts. Previously, we have shown that IL-1beta activates ERK1/2, JNKs, and protein kinase C (PKC). However, signaling pathways involved in the regulation of MMP-2 and -9 expression and activity are not yet well understood. Using adult rat cardiac fibroblasts, we show that inhibition of ERK1/2 and JNKs inhibits IL-1beta-stimulated increases in MMP-9, not MMP-2, expression and activity. Chelerythrine, an inhibitor of PKC, inhibited activation of ERK1/2 and JNKs and expression and activity of both MMPs. Selective inhibition of PKC-alpha/beta1 using Go6976 inhibited JNKs activation and the expression and activity of MMP-9, not MMP-2. Inhibition of PKC-theta and PKC-zeta using pseudosubstrates inhibited IL-1beta-stimulated activation of ERK1/2 and JNKs and the expression and activity of MMP-2 and -9. Inhibition of PKC-epsilon had no effect. IL-1beta activated NF-kappaB pathway as measured by increased phosphorylation of IKKalpha/beta and IkappaB-alpha. Inhibition of ERK1/2, JNKs, and PKC-alpha/beta1 had no effect on NF-kappaB activation, whereas inhibition of PKC-theta and PKC-zeta inhibited IL-1beta-stimulated activation of NF-kappaB. SN50, NF-kappaB inhibitor peptide, inhibited IL-1beta-stimulated increases in MMP-2 and -9 expression and activity. These observations suggest that 1) activation of ERK1/2 and JNKs plays a critical role in the regulation of MMP-9, not MMP-2, expression and activity; 2) PKC-alpha/beta1 act upstream of JNKs, not ERK1/2; 3) PKC-zeta and -theta, not PKC-epsilon, act upstream of JNKs, ERK1/2, and NF-kappaB; and 4) activation of NF-kappaB stimulates expression and activity of MMP-2 and -9. PMID- 15269223 TI - Phosphatidylcholine and N-methylated phospholipids are nonessential in Saccharomyces cerevisiae. AB - Phosphatidylcholine (PtdCho) is the most abundant phospholipid in numerous eukaryotes and is generally thought to be essential for membrane structure and cellular function. We designed a specific test of this idea by using genetic and biochemical manipulation of yeast. Yeast mutants (pem1 pem2Delta) lacking the phosphatidylethanolamine (PtdEtn) methyltransferase enzymes require choline for growth and cannot make N-methylated phospholipids. When these strains are grown on a glucose carbon source supplemented with 20 mm propanolamine (Prn), the PtdCho level declines precipitously to the limits of detection (<0.6%), and the hexagonal phase-forming, primary amine-containing lipids, PtdEtn and PtdPrn, constitute approximately 60% of the total phospholipid content of the cell. When the lipids were analyzed by mass spectrometry, there was no compensatory shift in unsaturation of the PtdEtn and PtdPrn toward more bilayer-forming species. Thus the majority of the cellular amino phospholipids remained hexagonal phase forming. The pem1 pem2Delta cells will also grow without choline, in the presence of Prn, on nonfermentable carbon sources (requiring functional mitochondria) and accumulate nearly 70% of their phospholipid as hexagonal phase-forming types. These data provide compelling evidence that the functions of PtdCho and N methylated lipids in membranes are nonessential in Saccharomyces cerevisiae. PMID- 15269224 TI - Regulation of SHP-1 tyrosine phosphatase in human platelets by serine phosphorylation at its C terminus. AB - SHP-1 is a Src homology 2 (SH2) domain-containing tyrosine phosphatase that plays an essential role in negative regulation of immune cell activity. We describe here a new model for regulation of SHP-1 involving phosphorylation of its C terminal Ser591 by associated protein kinase Calpha. In human platelets, SHP-1 was found to constitutively associate with its substrate Vav1 and, through its SH2 domains, with protein kinase Calpha. Upon activation of either PAR1 or PAR4 thrombin receptors, the association between the three proteins was retained, and Vav1 became phosphorylated on tyrosine and SHP-1 became phosphorylated on Ser591. Phosphorylation of SHP-1 was mediated by protein kinase C and negatively regulated the activity of SHP-1 as demonstrated by a decrease in the in vitro ability of SHP-1 to dephosphorylate Vav1 on tyrosine. Protein kinase Calpha therefore critically and negatively regulates SHP-1 function, forming part of a mechanism to retain SHP-1 in a basal active state through interaction with its SH2 domains, and phosphorylating its C-terminal Ser591 upon cellular activation leading to inhibition of SHP-1 activity and an increase in the tyrosine phosphorylation status of its substrates. PMID- 15269225 TI - Diffusion of epidermal growth factor in rat brain extracellular space measured by integrative optical imaging. AB - Epidermal growth factor (EGF) stimulates proliferation, process outgrowth, and survival in the CNS. Understanding the actions of EGF necessitates characterizing its distribution in brain tissue following drug delivery or release from cellular sources. We used the integrative optical imaging (IOI) method to measure diffusion of fluorescently labeled EGF (6,600 Mr; 4 microg/ml) in the presence of excess unlabeled EGF (90 microg/ml) to compete off specific receptor binding and reveal the "true" EGF diffusion coefficient following injection in rat brain slices (400 microm). The effective diffusion coefficient was 5.18 +/- 0.16 x 10( 7) (SE) cm2/s (n = 22) in rat somatosensory cortex and the free diffusion coefficient, determined in dilute agarose gel, was 16.6 +/- 0.12 x 10(-7) cm2/s (n = 27). Tortuosity (lambda), a parameter representing the hindrance imposed on EGF by the convoluted brain extracellular space (ECS), was 1.8, the lowest yet measured by IOI for a protein in brain. Control experiments with fluorescent dextran of similar molecular weight and tetramethylammonium confirmed EGF did not affect local ECS structure. We conclude that transport of smaller growth factors such as EGF through brain ECS is less hindered than that of larger proteins (>10,000 Mr, e.g., nerve growth factor) where typically lambda > 2.1. Modeling was used to predict that low lambda will allow EGF sources in the brain to be further from target cells and still elicit a biological response. High lambda values for larger growth factors imply more constrained local biological effects than with smaller proteins such as EGF. PMID- 15269226 TI - Parietal representation of hand velocity in a copy task. AB - We recorded neural activity from ensembles of neurons in areas 5 and 2 of parietal cortex, while two monkeys copied triangles, squares, trapezoids, and inverted triangles and used both linear and nonlinear models to predict the hand velocity from the neural activity of the ensembles. The linear model generally outperformed the nonlinear model, suggesting a reasonably linear relation between the neural activity and the hand velocity. We also found that the average transfer function of the linear model fit to individual cells was a low-pass filter because the neural response had considerable high-frequency power, whereas the hand velocity only had power at frequencies below approximately 5 Hz. Increasing the width of the transfer function, up to a width of 700-800 ms, improved the fit of the model. Furthermore, the Rsqr of the linear model improved monotonically with the number of cells in the ensemble, saturating at 60-80% for a filter width of 700 ms. Finally, it was found that including an interaction term, which allowed the transfer function to shift with the eye position, did not improve the fit of the model. Thus ensemble neural responses in superior parietal cortex provide a high-fidelity, linear representation of hand kinematics within our task. PMID- 15269227 TI - Differential roles of neuronal activity in the supplementary and presupplementary motor areas: from information retrieval to motor planning and execution. AB - We explored functional differences between the supplementary and presupplementary motor areas (SMA and pre-SMA, respectively) systematically with respect to multiple behavioral factors, ranging from the retrieval and processing of associative visual signals to the planning and execution of target-reaching movement. We analyzed neuronal activity while monkeys performed a behavioral task in which two visual instruction cues were given successively with a delay: one cue instructed the location of the reach target, and the other instructed arm use (right or left). After a second delay, the monkey received a motor-set cue to be prepared to make the reaching movement as instructed. Finally, after a GO signal, it reached for the instructed target with the instructed arm. We found the following apparent differences in activity: 1) neuronal activity preceding the appearance of visual cues was more frequent in the pre-SMA; 2) a majority of pre SMA neurons, but many fewer SMA neurons, responded to the first or second cue, reflecting what was shown or instructed; 3) in addition, pre-SMA neurons often reflected information combining the instructions in the first and second cues; 4) during the motor-set period, pre-SMA neurons preferentially reflected the location of the target, while SMA neurons mainly reflected which arm to use; and 5) when executing the movement, a majority of SMA neurons increased their activity and were largely selective for the use of either the ipsilateral or contralateral arm. In contrast, the activity of pre-SMA neurons tended to be suppressed. These findings point to the functional specialization of the two areas, with respect to receiving associative cues, information processing, motor behavior planning, and movement execution. PMID- 15269228 TI - Contributions of mossy fiber and CA1 pyramidal cell sprouting to dentate granule cell hyperexcitability in kainic acid-treated hippocampal slice cultures. AB - Axonal sprouting like that of the mossy fibers is commonly associated with temporal lobe epilepsy, but its significance remains uncertain. To investigate the functional consequences of sprouting of mossy fibers and alternative pathways, kainic acid (KA) was used to induce robust mossy fiber sprouting in hippocampal slice cultures. Physiological comparisons documented many similarities in granule cell responses between KA- and vehicle-treated cultures, including: seizures, epileptiform bursts, and spontaneous excitatory postsynaptic currents (sEPSCs) >600 pA. GABAergic control and contribution of glutamatergic synaptic transmission were similar. Analyses of neurobiotin-filled CA1 pyramidal cells revealed robust axonal sprouting in both vehicle- and KA-treated cultures, which was significantly greater in KA-treated cultures. Hilar stimulation evoked an antidromic population spike followed by variable numbers of postsynaptic potentials (PSPs) and population spikes in both vehicle- and KA-treated cultures. Despite robust mossy fiber sprouting, knife cuts separating CA1 from dentate gyrus virtually abolished EPSPs evoked by hilar stimulation in KA-treated but not vehicle-treated cultures, suggesting a pivotal role of functional afferents from CA1 to dentate gyrus in KA-treated cultures. Together, these findings demonstrate striking hyperexcitability of dentate granule cells in long-term hippocampal slice cultures after treatment with either vehicle or KA. The contribution to hilar-evoked hyperexcitability of granule cells by the unexpected axonal projection from CA1 to dentate in KA-treated cultures reinforces the idea that axonal sprouting may contribute to pathologic hyperexcitability of granule cells. PMID- 15269229 TI - Orexin (hypocretin) effects on constitutively active inward rectifier K+ channels in cultured nucleus basalis neurons. AB - Orexins are excitatory transmitters implicated in sleep disorders. Because orexins were discovered only recently, their ionic and signal transduction mechanisms have not been well clarified. We recently reported that orexin A (OXA) inhibits G protein-coupled inward rectifier K+ (GIRK) channels in cultured locus coeruleus and nucleus tuberomammillaris neurons. Other work in our laboratory revealed the existence of a novel inward rectifier K+ channel (KirNB), which is located in cholinergic neurons of the nucleus basalis (NB) and possesses unique single-channel characteristics. The mean open time is considerably shorter in KirNB than in Kir2.0 channels. Constitutive activity and a smaller unitary conductance set KirNB apart from cloned Kir3.0 channels. Previously, we found that substance P excites NB neurons by inhibiting KirNB channels. Here we show that orexins suppress KirNB channel activity, likely leading to neuronal excitation. Electrophysiological studies were performed on cultured NB neurons from the basal forebrain. OXA application decreased whole cell conductance through a pertussis toxin (PTX)-insensitive G protein. The OXA-suppressed current was inwardly rectifying with a reversal potential around E(K). Single-channel recordings of NB neurons revealed that constitutively active KirNB channels were transiently inhibited by OXA. Okadaic acid pretreatment abolished the recovery. The results suggest that OXA inhibition of KirNB is mediated by a PTX-insensitive G protein (i.e., G(q/11)), which eventually results in channel phosphorylation. Recovery from this inhibition is by dephosphorylation. These results, taken together with our previous study, suggest that orexin receptors can elicit neuronal excitation through at least two families of inward rectifier K+ channels: GIRK and KirNB channels. PMID- 15269230 TI - Intracellular measurements of spatial integration and the MAX operation in complex cells of the cat primary visual cortex. AB - We have examined the spatial integration properties of complex cells to determine whether some of their responses can be described by a maximum operation (MAX) like computation, as suggested by Riesenhuber and Poggio's model of object recognition. Membrane potential was recorded from anesthetized cats while optimally oriented bars were presented, either alone or in pairs, in different parts of the cells' receptive field. In most cells, the membrane potential response to two bars presented simultaneously could not be predicted by the sum of the responses to individual bars. In many cells, however, the responses closely approximated a MAX-like model. That is, the response of the cell to two bars was similar to the larger of the two individual responses ("soft-MAX"). The degree of nonlinear summation varied from cell to cell and varied within single cells from one stimulus configuration to another but on average fit most closely to the MAX model. The firing response of the cells was also well predicted by the MAX-like model. The MAX-like behavior was independent of the distance between the bars (orthogonal to the preferred orientation), independent of the relative amplitude of the responses, and slightly less pronounced at low levels of contrast. This MAX-like behavior of a subset of complex cells may play an important role in invariant object recognition in clutter. PMID- 15269231 TI - Quantifying the ontogeny of optokinetic and vestibuloocular behaviors in zebrafish, medaka, and goldfish. AB - We quantitatively studied the ontogeny of oculomotor behavior in larval fish as a foundation for studies linking oculomotor structure and function with genetics. Horizontal optokinetic and vestibuloocular reflexes (OKR and VOR, respectively) were measured in three different species (goldfish, zebrafish, and medaka) during the first month after hatching. For all sizes of medaka, and most zebrafish, Bode plots of OKR (0.065-3.0 Hz, +/-10 degrees/s) revealed that eye velocity closely followed stimulus velocity (gain > 0.8) at low frequency but dropped sharply above 1 Hz (gain < 0.3 at 3 Hz). Goldfish showed increased gain proportional to size across frequencies. Linearity testing with steps and sinusoids showed excellent visual performance (gain > 0.8) in medaka almost from hatching; but zebrafish and goldfish exhibited progressive improvement, with only the largest equaling medaka performance. Monocular visual stimulation in zebrafish and goldfish produced gains of 0.5 versus <0.1 for the eye viewing a moving versus stationary stimulus pattern but 0.25 versus <0.1 in medaka. Angular VOR appeared much later than OKR, initially at only high accelerations (>200 degrees /s at 0.5 Hz), first in medaka followed by larger (8.11 mm) zebrafish; but it was virtually nonexistent in goldfish. Velocity storage was not observed except for an eye velocity build-up in the largest medaka. In summary, a robust OKR was achieved shortly after hatching in all three species. In contrast, larval fish seem to be unique among vertebrates tested in their lack of significant angular VOR at stages where active movement is required for feeding and survival. PMID- 15269232 TI - Motor-unit coherence and its relation with synchrony are influenced by training. AB - The purpose of the study was to quantify the strength of motor-unit coherence from the left and right first dorsal interosseous muscles in untrained, skill trained (musicians), and strength-trained (weightlifters) individuals who had long-term specialized use of their hand muscles. The strength of motor-unit coherence was quantified from a total of 394 motor-unit pairs in 13 subjects using data from a previous study in which differences were found in the strength of motor-unit synchronization depending on training status. In the present study, we found that the strength of motor-unit coherence was significantly greater in the left compared with the right hand of untrained right-handed subjects with the largest differences observed between 21 and 24 Hz. The strength of motor-unit coherence was lower in both hands of skill-trained subjects (21-27 Hz) and the right (skilled) hand of untrained subjects (21-24 Hz), whereas the largest motor unit coherence was observed in both hands of strength-trained subjects (3-9 and 21-27 Hz). A strong curvilinear association was observed between motor-unit synchronization and the integral of coherence at 10-30 Hz in all motor-unit pairs (r2 = 0.77), and was most pronounced in strength-trained subjects (r2 = 0.90). Furthermore, this association was accentuated when using synchronization data with broad peaks (>11 ms), suggesting that the 10- to 30-Hz coherence is due to oscillatory activity in indirect branched common inputs. The altered coherence with training may be due to an interaction between cortical inhibition and the number of direct common inputs to motor neurons in skill- or strength-trained hands. PMID- 15269233 TI - Regulatory roles of JNK in programmed cell death. AB - Programmed cell death or apoptosis is the regulatory mechanism for removing unneeded cells during animal development and in tissue homeostasis. Perturbation of the cell death mechanisms leads to various disorders, including neurodegenerative diseases, immunodeficiency diseases, and tumors. c-Jun N terminal kinase (JNK) has crucial roles in the regulation of cell death in response to many stimuli. Since JNK is highly conserved from yeast to mammals, genetic studies using model animals are helpful in understanding the principal cell death mechanisms regulated by JNK. For example, loss-of-function studies using the targeted disruption of murine genes have established the genetic framework of the mechanisms of the cell death induced by UV radiation. Also, in Drosophila, many cell death-related genes have been identified by genetics. Genetic studies of JNK-dependent cell death mechanisms should shed light on the regulation of both physiological and pathological cell death. PMID- 15269234 TI - Roles of MAP kinase cascades in Caenorhabditis elegans. AB - Mitogen-activated protein kinases (MAPKs) are serine/threonine protein kinases that are activated by diverse stimuli such as growth factors, cytokines, neurotransmitters and various cellular stresses. MAPK cascades are generally present as three-component modules, consisting of MAPKKK, MAPKK and MAPK. The precise molecular mechanisms by which these MAPK cascades transmit signals is an area of intense research, and our evolving understanding of these signal cascades has been facilitated in great part by genetic analyses in model organisms. One organism that has been commonly used for genetic manipulation and physiological characterization is the nematode Caenorhabditis elegans. Genes sequenced in the C. elegans genome project have furthered the identification of components involved in several MAPK pathways. Genetic and biochemical studies on these components have shed light on the physiological roles of MAPK cascades in the control of cell fate decision, neuronal function and immunity in C. elegans. PMID- 15269235 TI - Mechanisms for removal of developmentally abnormal cells: cell competition and morphogenetic apoptosis. AB - Various cell differentiation signals are tightly linked with apoptotic signals. For example, as a result of somatic mutations, cells within a developing field occasionally receive an altered level of morphogenetic signaling that gives rise to an abnormal cell type. However, these developmentally abnormal cells are frequently removed by activating apoptotic signals. Although such phenomena are crucial for assuring normal development and maintaining a healthy state of various organs, the molecular mechanisms that sense aberrant signals and activate the apoptotic pathway(s) have not fully been investigated. In this review, we discuss recent progress in this area. Cell competition and morphogenetic apoptosis are two kinds of cell death, both of which are mediated by abnormal signaling of Dpp, a member of the TGF-beta superfamily that functions in Drosophila as a morphogen, mitogen and survival factor. Cell competition results in autonomous apoptosis induced by reduced reception of the extracellular survival factor Dpp, while morphogenetic apoptosis is nonautonomous, and is induced by contact of cells receiving different levels of Dpp signaling. PMID- 15269236 TI - A new approach for rapidly reshaping single-chain antibody in vitro by combining DNA shuffling with ribosome display. AB - Antibody reshaping is an effective way to reduce the immunogenicity while maintaining or improving the affinity of murine antibodies. This paper describe a new in vitro approach for rapidly reshaping murine antibodies by combining DNA shuffling with ribosome display. With the new method, a reshaping anti-4-1BB single-chain antibody (scFv), Re-4B4-1 scFv, which bound to its antigen (4-1BB) specifically and strongly, was selected from a reshaping library. These results proved definitely the feasibility of the new designed approach for antibody reshaping. PMID- 15269237 TI - Side chain-side chain interactions of arginine with tyrosine and aspartic acid in Arg/Gly/Tyr-rich domains within plant glycine-rich RNA binding proteins. AB - Plant glycine-rich RNA-binding proteins (GRRBPs) contain a glycine-rich region at the C-terminus whose structure is quite unknown. The C-terminal glycine-rich part is interposed with arginine and tyrosine (arginine/glycine/tyrosine (RGY)-rich domain). Comparative sequence analysis of forty-one GRRBPs revealed that the RGY rich domain contains multiple repeats of Tyr-(Xaa)h-(Arg)k-(Xaa)l, where Xaa is mainly Gly, "k" is 1 or 2, and "h" and "l" range from 0 to 10. Two peptides, 1 (G1G2Y3G4G5G6R7R8D9G10) and 2 (G1G2R3R4D5G6G7Y8G9G10), corresponding to sections of the RGY-rich domain in Zea mays RAB15, were selected for CD and NMR experiments. The CD spectra indicate a unique, positive band near 228 nm in both peptides that has been ascribed to tyrosine residues in ordered structures. The pH titration by NMR revealed that a side chain-side chain interaction, presumably an H-Nepsilon...O=Cgamma hydrogen bonding interaction in the salt bridge, occurs between Arg (i) and Asp (i + 2). 1D GOESY experiments indicated the presence of NOE between the aromatic side chain proton and the arginine side chain proton in the two peptides suggesting strongly that the Arg (i) aromatic side chain interacts directly with the Tyr (i +/- 4 or i +/- 5) side chain. PMID- 15269238 TI - Fluorescence resonance energy transfer between points on actin and the C-terminal region of tropomyosin in skeletal muscle thin filaments. AB - Fluorescence resonance energy transfer between points on tropomyosin (positions 87 and 190) and actin (Gln-41, Lys-61, Cys-374, and the ATP-binding site) showed no positional change of tropomyosin relative to actin on the thin filament in response to changes in Ca2+ concentration (Miki et al. (1998) J. Biochem. 123, 1104-1111). This is consistent with recent electron cryo-microscopy analysis, which showed that the C-terminal one-third of tropomyosin shifted significantly towards the outer domain of actin, while the N-terminal half of tropomyosin shifted only a little (Narita et al. (2001) J. Mol. Biol. 308, 241-261). In order to detect any significant positional change of the C-terminal region of tropomyosin relative to actin, we generated mutant tropomyosin molecules with a unique cysteine residue at position 237, 245, 247, or 252 in the C-terminal region. The energy donor probe was attached to these positions on tropomyosin and the acceptor probe was attached to Cys-374 or Gln-41 of actin. These probe labeled mutant tropomyosin molecules retain the ability to regulate the acto-S1 ATPase activity in conjunction with troponin and Ca2+. Fluorescence resonance energy transfer between these points of tropomyosin and actin showed a high transfer efficiency, which should be very sensitive to changes in distance between probes attached to actin and tropomyosin. However, the transfer efficiency did not change appreciably upon removal of Ca2+ ions, suggesting that the C-terminal region of tropomyosin did not shift significantly relative to actin on the reconstituted thin filament in response to the change of Ca2+ concentration. PMID- 15269239 TI - Intermediates in the inactivation and unfolding of dimeric arginine kinase induced by GdnHCl. AB - Equilibrium studies of guanidine hydrochloride (GdnHCl)-induced unfolding of dimeric arginine kinase (AK) from sea cucumber have been performed by monitoring by enzyme activity, intrinsic protein fluorescence, circular dichroism (CD), 1 anilinonaphthalene-8sulfonate (ANS) binding, size-exclusion chromatography and glutaraldehyde cross-linking. The unfolding is a multiphasic process involving at least two dimeric intermediates. The first intermediate, I1, which exists at 0 0.4 M GdnHCl, is a compact inactive dimer lacking partial global structure, while the second dimeric intermediate, I2, formed at 0.5-2.0 M GdnHCl, possesses characteristics similar to the globular folding intermediates described in the literature. The whole unfolding process can be described as follows: (1) inactivation and the appearance of the dimeric intermediate I1; (2) sudden unwinding of I1 to another dimeric intermediate, I2; (3) dissociation of dimeric intermediate I2 to monomers U. The refolding processes initiated by rapid dilution in renaturation buffers indicate that denaturation at low GdnHCl concentrations (below 0.4 M GdnHCl) is reversible and that there seems to be an energy barrier between the two intermediates (0.4-0.5 M GdnHCl), which makes it difficult for AK denatured at high GdnHCl concentrations (above 0.5 M) to reconstitute and regain its catalytic activity completely. PMID- 15269240 TI - Interaction of myosin.ADP.fluorometal complexes with fluorescent probes and direct observation using quick-freeze deep-etch electron microscopy. AB - Myosin forms stable ternary complexes with ADP and phosphate analogues of fluorometals that mimic different ATPase reaction intermediates corresponding to each step of the cross-bridge cycle. In the present study, we monitored the formation of ternary complexes of myosin.ADP.fluorometal using the fluorescence probe prodan. It has been reported that the fluorescence changes of the probe reflect the formation of intermediates in the ATPase reaction [Hiratsuka (1998) Biochemistry 37, 7167-7176]. Prodan bound to skeletal muscle heavy-mero-myosin (HMM).ADP.fluorometal, with each complex showing different fluorescence spectra. Prodan bound to the HMM.ADP.BeFn complex showed a slightly smaller red-shift than other complexes in the presence of ATP, suggesting a difference in the localized conformation or a difference in the population of BeFn species of global shape. We also examined directly the global structure of the HMM.ADP.fluorometal complexes using quick-freeze deep-etch replica electron microscopy. The HMM heads in the absence of nucleotides were mostly straight and elongated. In contrast, the HMM heads of ternary complexes showed sharply kinked or rounded configurations as seen in the presence of ATP. This is the first report of the direct observation of myosin-ADP-fluorometal ternary complexes, and the results suggest that these complexes indeed mimic the shape of the myosin head during ATP hydrolysis. PMID- 15269241 TI - Lysosomal acid lipase as a preproprotein. AB - Lysosomal acid lipase (LAL; EC 3.1.1.13) hydrolyzes intracellular triglycerides and cholesterol esters taken up by various cell-types. Previously, LAL purified from human liver tissue was described as a preproprotein with a 27 amino acid signal peptide and a 49 amino acid propeptide. Three mutants of the putative proregion of LAL were produced and expressed in Spodoptera frugiperda insect cells. Pulse-chase experiments demonstrated that LAL undergoes proteolytical processing. The deletion of the 49 amino acids led to a complete loss of the LAL activity. The two other mutants were produced at the C-terminus of the pro region, at positions 49 and 50, by site-directed mutagenesis. Mutant K49R showed wild-type LAL activity, but mutant G50A showed significantly reduced enzyme activity compared to wild-type LAL and a greater reduction in culture medium than in detergent cell extracts. Kinetic data suggest that mutant G50A is less stable than wild-type LAL and mutant K49R. In contrast to K49, the highly conserved amino acid residue G50 seems to be in a very important position and its mutation influences both secretion and enzyme activity of LAL. A three-dimensional model of LAL shows that K49 and G50 are localized in the loop-region between two beta sheets, highly accessible for proteolytic enzymes. These data together indicate that LAL is indeed a preproprotein, in which the pro-region is essential for its folding and stability, secretion, and enzyme activity. PMID- 15269242 TI - Construction and function of two Cys146-mutants with high activity, derived from recombinant human soluble B lymphocyte stimulator. AB - B lymphocyte stimulator (BLyS) is a novel member of tumor necrosis factor (TNF) ligand family that is important in B cell maturation and survival. Previous studies were almost related to the function or mechanism of its wild type. Here, we constructed two site-directed mutants of the recombinant human soluble BLyS, the BY-A and BY-V, and found that BY-V ranked the highest whenever in the process of promoting proliferation of B lymphocytes in vitro or stimulating total serum IgG and IgM secretion in vivo. Besides, assays for the biological responses of human leukemic cell lines to BLyS, BY-A and BY-V demonstrated that they could suppress the proliferation of Raji cells but promote the growth of THP-1. The discovery of BY-V with high activity will help come to a conclusion that the mutation of Cys146 to Val146 might improve the biological activity of BLyS. PMID- 15269243 TI - Protective properties of neoechinulin A against SIN-1-induced neuronal cell death. AB - Peroxynitrite (ONOO-) is thought to be involved in the neurodegenerative process. To screen for neuroprotective compounds against ONOO- -induced cell death, we developed 96-well based assay procedures for measuring surviving cell numbers under oxidative stress caused by 3-(4-morpholinyl) sydnonimine hydrochloride (SIN 1), a generator of ONOO-, and sodium N,N-dietyldithiocarbamate trihydrate (DDC), an inhibitor of Cu/Zn superoxide (O2-) dismutase. Using these procedures, we obtained a microbial metabolite that rescued primary neuronal cells from SIN-1 induced damage, but not from DDC-induced damage. By NMR analysis, the compound was identified as neoechinulin A, an antioxidant compound that suppresses lipid oxidation. We found that the compound rescues neuronal cells such as primary neuronal cells and differentiated PC12 cells from damage induced by extracellular ONOO-. However, non-neuronal cells, undifferentiated PC12 cells and cells of the fibroblast cell line 3Y1 were not rescued. Neoechinulin A has scavenging, neurotrophic factor-like and anti-apoptotic activities. This compound specifically scavenges ONOO-, but not O2- or nitric oxide (NO). Similar to known neuroprotective substances such as nerve growth factor and extracts of Gingko biloba leaves, neoechinulin A inhibits the SIN-1-induced activation of caspase-3 like proteases and increases NADH-dehydrogenase activity. These results suggest that neoechinulin A might be useful for protecting against neuronal cell death in neurodegenerative diseases. PMID- 15269244 TI - Protein-tyrosine phosphatase 1B associates with insulin receptor and negatively regulates insulin signaling without receptor internalization. AB - Phosphorylated platelet-derived growth factor (PDGF) receptor becomes internalized and then is dephosphorylated by protein-tyrosine phosphatase (PTP) 1B at the endoplasmic reticulum (ER). However, it remains unclear where PTP1B dephosphorylates insulin receptor and inhibits its activity. To clarify how and where PTP1B could interact with insulin receptor, we overexpressed a phosphatase inactive mutant, PTP1BC/S, in 3T3-L1 adipocytes. Although PDGF receptor was maximally associated with PTP1BC/S at 30 min after PDGF stimulation, the maximal association of insulin receptor with PTP1BC/S was attained at 5 min after insulin stimulation. Furthermore, dansylcadaverine, a blocker of receptor internalization, inhibited this PDGF-induced association of PTP1BC/S with its receptor. However, dansylcadaverine did not affect the insulin-stimulated association of PTP1BC/S with insulin receptor, as well as dephosphorylation of insulin receptor by PTP1B. These results indicate that PTP1B might interact with insulin receptor and deactivate it without internalization. Finally, we overexpressed the wild-type and cytosolic-form of PTP1B to determine the role of ER-anchoring of PTP1B, and found that both inhibited insulin signaling equally. Thus, our data indicate that localization of PTP1B at the ER is not needed for insulin receptor dephosphorylation by PTP1B. PMID- 15269245 TI - Massspectrometric analyses of transmembrane proteins in human erythrocyte membrane. AB - It is difficult to understand the functional mechanisms of integral membrane proteins without having protein chemical information on these proteins. Although there have been many attempts to identify functionally important amino acids in membrane proteins, chemically and enzymatically cleaved peptides of integral membrane proteins have been difficult to handle because of their hydrophobic properties. In the present study, we have applied an analytical method to transmembrane proteins combining amino acid sequencing, matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry, and liquid chromatography with electrospray ionization (LC/ESI) mass spectrometry. We could analyze most (97%) of the tryptic fragments of the transmembrane domains of band 3 as well as other minor membrane proteins. The peptide mapping of the transmembrane domain of band 3 was completed and the peptide mapping information allowed us to identify the fragments containing lysine residues susceptible to 4 acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic acid (SITS) and to 2,4 dinitrofluorobenzene (DNFB). This method should be applicable to membrane proteins not only in erythrocyte membranes but also in other membranes. PMID- 15269246 TI - N-terminal modification and its effect on the biochemical characteristics of Akazara scallop tropomyosins expressed in Escherichia coli. AB - Akazara scallop striated muscle tropomyosin mutants without a fused amino acid (nf-Tm), and with Ala- (A-Tm) or Asp-Ala- (DA-Tm) fused at the N-terminus were expressed in Escherichia coli cells. Among them, nf-Tm alone has an initial methionine. The native Akazara scallop tropomyosin and DA-Tm showed similar alpha helix contents and intrinsic viscosity, but nf-Tm and A-Tm exhibited lower values than those of the native tropomyosin. According to the relative viscosity, all the expressed tropomyosins appear to have lost head-to-tail polymerization ability. Though nf-Tm has extremely low actin-binding ability, the ability was almost completely recovered with a two amino acid fusion but incompletely with a one amino acid fusion. On the other hand, an amino acid fusion, irrespective of the number, seemed to inhibit the Mg-ATPase activity of actomyosin. However, the bacterially expressed tropomyosins together with Akazara scallop troponin do not confer the full Ca(2+)-regulation ability of Mg-ATPase activity of actomyosin. These results support that N-terminal blocking probably by an acetyl group of Akazara scallop tropomyosin plays an important role not only in head-to-tail polymerization and actin-binding, as known for vertebrate tropomyosin, but also in maintaining the secondary or higher structure and Ca(2+)-regulation together with troponin. PMID- 15269247 TI - Thermal equilibrium of two conformations in photosensitive nitrile hydratase probed by the FTIR band of nitric oxide bound to the non-heme iron center. AB - Nitrile hydratase (NHase) from Rhodococcus N-771 is a novel enzyme that is inactive in the dark due to an enodogenous nitric oxide (NO) molecule bound to the non-heme iron center, and is activated by its photodissociation. FTIR spectra in the NO stretching region of the dark-inactive NHase were recorded in the temperature range of 270-80 K. Two NO peaks were observed at 1854 and 1846 cm-1 at 270 K, and both frequencies upshifted as the temperature was lowered, retaining the peak separation of 8-9 cm-1. The relative intensity of the lower frequency peak increased with decreasing temperature up to ~120 K, whereas it was mostly unchanged below this temperature. This observation indicates that two distinct conformations with slightly different NO structures are thermally equilibrated in the dark-inactive NHase above ~120 K, and the interconversion is frozen-in at lower temperatures. The intensity ratio of the NO bands changed gradually upon increasing the pH from 5.5 to 11.0, but no specific pKa value was found. This result, together with the comparison of the light-induced FTIR difference spectra measured at pH 6.5 and 9.0, suggests that the protonation/deprotonation of a specific amino acid group in the active site of NHase is not a direct cause of the occurrence of the two conformations, although several protonatable groups in the protein may influence the energetics of the two conformers. From the previous observation that the isolated alpha subunit of NHase exhibited a single broad NO peak, it is suggested that interaction of the beta subunit forming the reactive cavity is essential for the double-minimum potential of the active-site structure. The frequencies and widths of the two NO bands changed upon addition of propionamide, 1,4-dioxane, and cyclohexyl isocyanide, indicating that these compounds are bound to the active pocket and change the interactions of the iron center or the dielectric environments around the NO molecule. Thus, the NO bands of NHase can also be a useful probe to monitor the binding of substrates and their analogues to the active pocket. PMID- 15269248 TI - A dataset of human liver proteins identified by protein profiling via isotope coded affinity tag (ICAT) and tandem mass spectrometry. AB - Proteins from human liver carcinoma Huh7 cells, representing transformed liver cells, and cultured primary human fetal hepatocytes (HFH) and human HH4 hepatocytes, representing nontransformed liver cells, were extracted and processed for proteome analysis. Proteins from stimulated cells (interferon-alpha treatment for the Huh7 and HFH cells and induction of hepatitis C virus [HCV] proteins for the HH4 cells) and corresponding control cells were labeled with light and heavy cleavable ICAT reagents, respectively. The labeled samples were combined, trypsinized, and subject to cation-exchange and avidin-affinity chromatographies. The resulting cysteine-containing peptides were analyzed by microcapillary LC-MS/MS. The MS/MS spectra were initially analyzed by searching the human International Protein Index database using the SEQUEST software (1). Subsequently, new statistical algorithms were applied to the collective SEQUEST search results of each experiment. First, the PeptideProphet software (2) was applied to discriminate true assignments of MS/MS spectra to peptide sequences from false assignments, to assign a probability value for each identified peptide, and to compute the sensitivity and error rate for the assignment of spectra to sequences in each experiment. Second, the ProteinProphet software (3) was used to infer the protein identifications and to compute probabilities that a protein had been correctly identified, based on the available peptide sequence evidence. The resulting protein lists were filtered by a ProteinProphet probability score p > or = 0.5, which corresponded to an error rate of less than 5%. A total of 1,296, 1,430, and 1,476 proteins or related protein groups were identified in three subdatasets from the Huh7, HFH, and HH4 cells, respectively. In total, these subdatasets contained 2,486 unique protein identifications from human liver cells. An increase of the threshold to p > or = 0.9 (corresponding to an error rate of less than 1%) resulted in 2,159 unique protein identifications (1,146, 1,235, and 1,318 for the Huh7, HFH, and HH4 cells, respectively). PMID- 15269249 TI - Informatics platform for global proteomic profiling and biomarker discovery using liquid chromatography-tandem mass spectrometry. AB - We have developed an integrated suite of algorithms, statistical methods, and computer applications to support large-scale LC-MS-based gel-free shotgun profiling of complex protein mixtures using basic experimental procedures. The programs automatically detect and quantify large numbers of peptide peaks in feature-rich ion mass chromatograms, compensate for spurious fluctuations in peptide signal intensities and retention times, and reliably match related peaks across many different datasets. Application of this toolkit markedly facilitates pattern recognition and biomarker discovery in global comparative proteomic studies, simplifying mechanistic investigation of physiological responses and the detection of proteomic signatures of disease. PMID- 15269250 TI - Transfection and functional expression of CYP4A1 and CYP4A2 using bicistronic vectors in vascular cells and tissues. AB - 20-hydroxyeicosatetraenoic acid (20-HETE), a CYP4A-derived arachidonic acid metabolite, is a potent vasoconstrictor and a modulator of vascular reactivity. We have shown that CYP4A1 and CYP4A2 are the major CYP4A isoforms expressed in the rat renal microcirculation. In the present study, we constructed two bicistronic vectors, pIRES2-EGFP-4A1 and pIRES2-EGFP-4A2, and examined their functional efficacy in COS-1 and vascular smooth muscle (A7r5) cells and in microdissected rat interlobar arteries. Immunocytochemistry coupled with fluorescence microscopy of pIRES2-EGFP-4A1- or pIRES2-EGFP-4A2-transfected COS-1 and A7r5 cells indicated that both enhanced green fluorescence protein (EGFP) and CYP4A1/4A2 were expressed in 80 to 90% of the cells. Western blot analysis showed a 3- to 5-fold increase of CYP4A1 and CYP4A2 proteins in pIRES2-EGFP-4A1- and pIRES2-EGFP-4A2-transfected cells as compared with control pIRES2-transfected cells. Cells transfected with pIRES2-EGFP-4A1 and pIRES2-EGFP-4A2 catalyzed arachidonic acid omega-hydroxylation to 20-HETE at rates of 0.85 +/- 0.29 and 0.27 +/- 0.04 nmol/10(7) cells/h, respectively. Transfection of interlobar arteries with either plasmid yielded EGFP immunofluorescence that was localized to the intima, media, and adventitia. Arteries transfected with pIRES2-EGFP-4A1 and pIRES2-EGFP-4A2 showed increased vasoreactivity displaying EC50 to phenylephrine of 0.24 +/- 0.07 and 0.11 +/- 0.03 microM, respectively, as compared with arteries transfected with pIRES2-EGFP (1.11 +/- 0.21 microM; n=6, p <0.05). The increased vasoreactivity to phenylephrine was inhibited by N methylsulfonyl-12,12-dibromododec-11-enamide, an inhibitor of CYP4A-catalyzed reactions, suggesting that a product of CYP4A1 and CYP4A2 catalytic activity contributed to the increased constrictor responsiveness. Removal of the endothelium did not prevent the sensitization to phenylephrine in vessels transfected with the plasmid containing the CYP4A1 cDNA, suggesting that the CYP4A product responsible for the sensitizing effect, presumably 20-HETE, is not of endothelial cell origin. PMID- 15269251 TI - Role of external pallidal segment in primate parkinsonism: comparison of the effects of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced parkinsonism and lesions of the external pallidal segment. AB - These experiments re-examined the notion that reduced activity in the external pallidal segment (GPe) results in the abnormalities of neuronal discharge in the subthalamic nucleus (STN) and the internal pallidal segment (GPi) and in the development of parkinsonian motor signs. Extracellular recording in two rhesus monkeys, which had been rendered parkinsonian by treatment with 1-methyl-4-phenyl 1,2,3,6-tetrahydropyridine (MPTP), revealed that the average neuronal discharge rate decreased in GPe but increased in STN and GPi. After MPTP, neurons in all three nuclei tended to discharge in oscillatory bursts. In addition, GABA release in STN (measured with microdialysis) was reduced, indicative of reduced activity along the GPe-STN pathway. Finally, the concentration of glutamic acid dehydrogenase (GAD; measured with autoimmunoradiography) was increased in GPe and GPi, likely reflecting increased striatal input and increased activity of local axon collaterals, respectively. Surprisingly, GAD protein in STN remained unchanged, indicating that the usual assumption that GAD levels are determined primarily by the overall activity of GABAergic elements may be too simplistic. The results from the MPTP-treated animals were compared with results obtained in a second group of three animals with ibotenic acid lesions of GPe. GPe lesions resulted in increased discharge in STN and GPi, comparable with the changes seen after MPTP but did not induce oscillatory bursting and had no behavioral effects. The results indicate that a mere reduction of GPe activity does not produce parkinsonism. Other changes, such as altered discharge patterns in STN and GPi, may play an important role in the generation of parkinsonism. PMID- 15269253 TI - Overexpression of glial cell line-derived neurotrophic factor using a lentiviral vector induces time- and dose-dependent downregulation of tyrosine hydroxylase in the intact nigrostriatal dopamine system. AB - The effects of continuous glial cell line-derived neurotrophic factor (GDNF) overexpression in the intact nigrostriatal dopamine (DA) system was studied using recombinant lentiviral (rLV) vector delivery of GDNF to the striatum or substantia nigra (SN) in the rat. Intrastriatal delivery of rLV-GDNF resulted in significant overexpression of GDNF in the striatum (2-4 ng/mg tissue) and anterograde transport of GDNF protein to the SN. Striatal rLV-GDNF delivery initially induced an increase in DA turnover (1-6 weeks), accompanied by significant contralateral turning in response to amphetamine, suggesting an enhancement of the DA system on the injected side. Starting 6 weeks after continuous GDNF delivery, we observed a selective downregulation of tyrosine hydroxylase (TH) protein (approximately 70%) that was maintained until the end of the experiment (24 weeks). A similar effect was observed when rLV-GDNF was injected into the SN. The magnitude of TH downregulation was related to the level of GDNF expression and was most pronounced in animals in which the striatal GDNF level exceeded 0.7 ng/mg tissue. The decreased TH protein levels were associated with similar reductions in the in vitro TH enzyme activity (approximately 70%); however, in vivo L-3,4-dihydroxyphenylalanine production rate and DA tissue levels were maintained at normal levels. The results indicate that downregulation of TH protein reflects a compensatory effect in response to continuous GDNF stimulation of the DA neurons mediated by a combination of overactivity at the DA synapse and a direct GDNF-induced action on TH gene expression. This compensatory mechanism is proposed to maintain long-term DA neuron function within the normal range. PMID- 15269252 TI - The retinitis pigmentosa 1 protein is a photoreceptor microtubule-associated protein. AB - The outer segments of rod and cone photoreceptor cells are highly specialized sensory cilia made up of hundreds of membrane discs stacked into an orderly array along the photoreceptor axoneme. It is not known how the alignment of the outer segment discs is controlled, although it has been suggested that the axoneme may play a role in this process. Mutations in the retinitis pigmentosa 1 (RP1) gene are a common cause of retinitis pigmentosa (RP). Disruption of the Rp1 gene in mice causes misorientation of outer segment discs, suggesting a role for RP1 in outer segment organization. Here, we show that the RP1 protein is part of the photoreceptor axoneme. Amino acids 28-228 of RP1, which share limited homology with the microtubule-binding domains of the neuronal microtubule-associated protein (MAP) doublecortin, mediate the interaction between RP1 and microtubules, indicating that the putative doublecortin (DCX) domains in RP1 are functional. The N-terminal portion of RP1 stimulates the formation of microtubules in vitro and stabilizes cytoplasmic microtubules in heterologous cells. Evaluation of photoreceptor axonemes from mice with targeted disruptions of the Rp1 gene shows that Rp1 proteins that contain the DCX domains also help control axoneme length and stability in vivo. These results demonstrate that RP1 is a MAP. Given the specific expression of RP1 in photoreceptors, RP1 is thus the first photoreceptor specific MAP to be identified. Furthermore, these findings indicate that the RP1 form of inherited retinal degeneration is part of the larger class of neurodegenerative diseases caused by MAP dysfunction. PMID- 15269254 TI - The ventral striatum in off-line processing: ensemble reactivation during sleep and modulation by hippocampal ripples. AB - Previously it has been shown that the hippocampus and neocortex can spontaneously reactivate ensemble activity patterns during post-behavioral sleep and rest periods. Here we examined whether such reactivation also occurs in a subcortical structure, the ventral striatum, which receives a direct input from the hippocampal formation and has been implicated in guidance of consummatory and conditioned behaviors. During a reward-searching task on a T-maze, flanked by sleep and rest periods, parallel recordings were made from ventral striatal ensembles while EEG signals were derived from the hippocampus. Statistical measures indicated a significant amount of reactivation in the ventral striatum. In line with hippocampal data, reactivation was especially prominent during post behavioral slow-wave sleep, but unlike the hippocampus, no decay in pattern recurrence was visible in the ventral striatum across the first 40 min of post behavioral rest. We next studied the relationship between ensemble firing patterns in ventral striatum and hippocampal ripples-sharp waves, which have been implicated in pattern replay. Firing rates were significantly modulated in close temporal association with hippocampal ripples in 25% of the units, showing a marked transient enhancement in the average response profile. Strikingly, ripple modulated neurons in ventral striatum showed a clear reactivation, whereas nonmodulated cells did not. These data suggest, first, the occurrence of pattern replay in a subcortical structure implied in the processing and prediction of reward and, second, a functional linkage between ventral striatal reactivation and a specific type of high-frequency population activity associated with hippocampal replay. PMID- 15269255 TI - Absence of C1q leads to less neuropathology in transgenic mouse models of Alzheimer's disease. AB - C1q, the recognition component of the classical complement activation pathway, is a multifunctional protein known to be expressed in brain of Alzheimer's disease (AD) patients. To experimentally address the role of C1q in AD, a mouse model lacking C1q (APPQ-/-) was generated by crossing Tg2576 animals (APP) with C1q deficient mice. The pathology of APPQ-/- was compared with that of APP mice and B6SJL controls at 3-16 months of age by immunohistochemistry and Western blot analysis. At younger ages (3-6 months), when no plaque pathology was present, no significant differences were seen in any of the neuronal or glial markers tested. At older ages (9-16 months), the APP and APPQ-/- mice developed comparable total amyloid and fibrillar beta-amyloid in frontal cortex and hippocampus; however, the level of activated glia surrounding the plaques was significantly lower in the APPQ-/- mice at 12 and 16 months. In addition, although Tg2576 mice showed a progressive decrease in synaptophysin and MAP2 in the CA3 area of hippocampus compared with control B6SJL at 9, 12, and 16 months, the APPQ-/- mice had significantly less of a decrease in these markers at 12 and 16 months. In a second murine model for AD containing transgenes for both APP and mutant presenilin 1 (APP/PS1), a similar reduction of pathology was seen in the APPPS1Q /- mice. These data suggest that at ages when the fibrillar plaque pathology is present, C1q exerts a detrimental effect on neuronal integrity, most likely through the activation of the classical complement cascade and the enhancement of inflammation. PMID- 15269256 TI - Alteration of neuronal firing properties after in vivo experience in a FosGFP transgenic mouse. AB - Identifying the cells and circuits that underlie perception, behavior, and learning is a central goal of contemporary neuroscience. Although techniques such as lesion analysis, functional magnetic resonance imaging, 2-deoxyglucose studies, and induction of gene expression have been helpful in determining the brain areas responsible for particular functions, these methods are technically limited. Currently, there is no method that allows for the identification and electrophysiological characterization of individual neurons that are associated with a particular function in living tissue. We developed a strain of transgenic mice in which the expression of the green fluorescent protein (GFP) is controlled by the promoter of the activity-dependent gene c-fos. These mice enable an in vivo or ex vivo characterization of the cells and synapses that are activated by particular pharmacological and behavioral manipulations. Cortical and subcortical fosGFP expression could be induced in a regionally restricted manner after specific activation of neuronal ensembles. Using the fosGFP mice to identify discrete cortical areas, we found that neurons in sensory-spared areas rapidly regulate action potential threshold and spike frequency to decrease excitability. This method will enhance our ability to study the way neuronal networks are activated and changed by both experience and pharmacological manipulations. In addition, because activated neurons can be functionally characterized, this tool may enable the development of better pharmaceuticals that directly affect the neurons involved in disease states. PMID- 15269257 TI - Effects of PKA-mediated phosphorylation of Snapin on synaptic transmission in cultured hippocampal neurons. AB - Use-dependent activation of protein kinase A (PKA) modulates transmitter release, contributing to synaptic plasticity. Snapin, a PKA substrate in neurons, associates with the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex, and its phosphorylation leads to increased binding of synaptotagmin to the SNARE complex. We investigated the role of PKA-dependent phosphorylation of Snapin in hippocampal neurons. Overexpression of Snapin S50D, a mutant mimicking the phosphorylated state, resulted in a decreased number of readily releasable vesicles. In addition, both the release probability of individual vesicles and the depression rate during high-frequency stimulation were increased. Overexpression of Snapin S50A, a mutant that cannot be phosphorylated, did not alter the size of the pool or the probability of release. Furthermore, dialysis of Sp-cAMPS, a nonhydrolyzable analog of cAMP that will promote phosphorylation by PKA, also led to increased synaptic depression in cells overexpressing wild-type Snapin. These results establish Snapin as an important target of PKA in CNS synapses and indicate a role for Snapin in the plasticity of transmitter release. PMID- 15269258 TI - Cell type-specific interleukin-1beta signaling in the CNS. AB - Interleukin-1beta (IL-1beta) is a potent and pleiotropic inflammatory cytokine that is highly produced in the CNS under conditions of damage, disease, or stress. This cytokine acts on CNS glia to effect inflammatory responses, mediated in part via activation of the nuclear factor-kappaB (NF-kappaB) transcription factor, and consequent induction of numerous cytokines. Neurons as well as astrocytes in the hippocampus also express the type 1 IL-1 receptor, indicating that this cytokine can influence neuronal function directly, yet IL-1beta does not induce production of cytokines in neurons as it does in glia. In contrast, IL 1beta regulates synaptic function of hippocampal neurons. Here we demonstrate that different signaling pathways mediate IL-1beta actions in neurons as compared with astrocytes. IL-1beta activates the p38 mitogen-activated protein kinase (MAPK) signaling pathway and induces the activation of CREB in hippocampal neurons, in contrast to the activation of NF-kappaB in hippocampal astrocytes, demonstrating cell type-specific signaling responses to IL-1 in the brain and yielding distinct functional responses. PMID- 15269259 TI - Differences in hippocampal neuronal population responses to modifications of an environmental context: evidence for distinct, yet complementary, functions of CA3 and CA1 ensembles. AB - Understanding how the hippocampus processes information critical for establishing spatial and declarative memories will benefit greatly from determining not only what kind of information the hippocampus registers, but also how this information is processed across the different hippocampal subfields. We addressed this question using a novel immediate-early gene-based brain-imaging method (Arc/H1a catFISH) that allows comparisons of neuronal ensembles activated by two experiences separated by approximately 30 min. Rats exposed to the same environment twice activated CA3 and CA1 ensembles with a similarly high degree of overlap. Changing the identity or configuration of local cues, or changing distal cues, activated CA3 and CA1 ensembles with reduced overlap. Yet, the overlap was greater in CA3 than in CA1. In contrast, rats exposed to two completely different environments activated CA3 and CA1 ensembles with low overlap, and this overlap was even lower in CA3 compared with CA1. Thus, CA3 has a discontinuous, whereas CA1 has a graded, population response to alterations of an environment. Additionally, as indicated by the percentage of active neurons, the context representation was more sparse in CA3 (approximately 18%) than in CA1 (approximately 35%). Finally, CA3 and CA1 activity levels were not correlated within a session, arguing against a simple coactivation of these regions. Instead, the within-rat ratio of CA3/CA1 cell activity was correlated across sessions, suggesting that the balance of CA3/CA1 activity is individual specific. Taken together, these findings suggest that CA3 and CA1 neuronal ensembles perform distinct, yet complementary, functions in the processing of spatial and contextual information. PMID- 15269260 TI - Novel environments enhance the induction and maintenance of long-term potentiation in the dentate gyrus. AB - The induction of long-term potentiation (LTP) in the hippocampal formation can be modulated by different behavioral states. However, few studies have addressed modulation of LTP during behavioral states in which the animal is likely acquiring new information. Here, we demonstrate that both the induction and the longevity of LTP in the dentate gyrus are enhanced when LTP is induced during the initial exploration of a novel environment. These effects are independent from locomotor activity, changes in brain temperature, and theta rhythm. Previous exposure to the novel environment attenuated this enhancement, suggesting that the effects of novelty habituate with familiarity. LTP longevity also was enhanced when induced in familiar environments containing novel objects. Together, these data indicate that both LTP induction and maintenance are enhanced when LTP is induced while rats investigate novel stimuli. We suggest that novelty initiates a transition of the hippocampal formation to a mode that is particularly conducive to synaptic plasticity, a process that could allow for new learning while preserving the stability of previously stored information. In addition, LTP induced in novel environments elicited a sustained late LTP. This suggests that a single synaptic population can display distinct profiles of LTP maintenance and that this depends on the animal's behavioral state during its induction. Furthermore, the duration of LTP enhanced by novelty parallels the time period during which the hippocampal formation is thought necessary for memory, consistent with the view that dentate LTP is of a duration sufficient to sustain memory in the hippocampal formation. PMID- 15269261 TI - Stereotyped odor-evoked activity in the mushroom body of Drosophila revealed by green fluorescent protein-based Ca2+ imaging. AB - To study the representation of olfactory information in higher brain centers, we expressed a green fluorescent protein-based Ca2+ sensor, G-CaMP, in the Drosophila mushroom body (MB). Using two-photon microscopy, we imaged odor-evoked G-CaMP fluorescence transients in MB neurons [Kenyon cells (KCs)] with single cell resolution. Odors produced large fluorescence transients in a subset of KC somata and in restricted regions of the calyx, the neuropil of the MB. In different KCs, odor-evoked fluorescence transients showed diverse changes with odor concentration: in some KCs, fluorescence transients were evoked by an odor at concentrations spanning several orders of magnitude, whereas in others only at a narrow concentration range. Different odors produced fluorescence transients in different subsets of KCs. The spatial distributions of KCs showing fluorescence transients evoked by a given odor were similar across individuals. For some odors, individual KCs with fluorescence transients evoked by a particular odor could be found in similar locations in different flies with spatial precisions on the order of the size of KC somata. These results indicate that odor-evoked activity can have remarkable spatial specificity in the MB. PMID- 15269262 TI - Motor skill learning depends on protein synthesis in motor cortex after training. AB - The role of protein synthesis in memory consolidation is well established for hippocampus-dependent learning and synaptic plasticity. Whether protein synthesis is required for motor skill learning is unknown. We hypothesized that skill learning is interrupted by protein synthesis inhibition (PSI). We intended to test whether local protein synthesis in motor cortex or cerebellum is required during skill acquisition and consolidation. Anisomycin (ANI; 100 microg/microl in 1 microl of PBS) injected into motor cortex, posterior parietal cortex, or cerebellum produced 84.0 +/- 1.44% (mean +/- SEM), 85.9 +/- 2.31%, and 87.3 +/- 0.17% of PSI 60 min after administration, respectively. In motor cortex, protein synthesis was still reduced at 24 hr (72.0 +/- 4.68% PSI) but normalized at 48 hr after a second injection given 24 hr after the first. To test for the effects of PSI on learning of a skilled reaching task, ANI was injected into motor cortex contralateral to the trained limb or into ipsilateral cerebellum immediately after daily training sessions 1 and 2. Two control groups received motor cortex injections of vehicle or ANI injections into contralateral parietal cortex. Control and cerebellar animals showed a sigmoid learning curve, which plateaued after day 4. PSI in motor cortex significantly reduced learning during days 1-4. Thereafter, when protein synthesis normalized, learning was reinitiated. ANI injections into motor cortex did not induce a motor deficit, because animals injected during the performance plateau did not deteriorate. This demonstrates that motor skill learning depends on de novo synthesis of proteins in motor cortex after training. PMID- 15269263 TI - Brain steroidogenesis mediates ethanol modulation of GABAA receptor activity in rat hippocampus. AB - An interaction with the GABA type A (GABA(A)) receptor has long been recognized as one of the main neurochemical mechanisms underlying many of the pharmacological actions of ethanol. However, more recent data have suggested that certain behavioral and electrophysiological actions of ethanol are mediated by an increase in brain concentration of neuroactive steroids that results from stimulation of the hypothalamic-pituitary-adrenal (HPA) axis. Neuroactive steroids such as 3alpha-hydroxy-5alpha-pregnan-20-one (3alpha,5alpha-THP) are, in fact, potent and efficacious endogenous positive modulators of GABA(A) receptor function. Because neurosteroids can be synthesized de novo in the brain, we have investigated whether ethanol might affect both neurosteroid synthesis and GABA(A) receptor function in isolated rat hippocampal tissue. Here, we show that ethanol increases the concentration of 3alpha,5alpha-THP as well as the amplitude of GABA(A) receptor-mediated IPSCs recorded from CA1 pyramidal neurons in isolated hippocampal slices. These effects are shared by the neurosteroid precursor progesterone, the peripheral benzodiazepine receptor-selective agonist CB34, and gamma-hydroxybutyrate, all of which are known to increase the formation of neuroactive steroids in plasma and in the brain. The action of ethanol on GABA(A) receptor-mediated IPSC amplitude is biphasic, consisting of a rapid, direct effect on GABA(A) receptor activity and an indirect effect that appears to be mediated by neurosteroid synthesis. Furthermore, ethanol affects GABA(A) receptor activity through a presynaptic action, an effect that is not dependent on neurosteroid formation. These observations suggest that ethanol may modulate GABA(A) receptor function through an increase in de novo neurosteroid synthesis in the brain that is independent of the HPA axis. This novel mechanism may have a crucial role in mediating specific central effects of ethanol. PMID- 15269264 TI - Studies on the development and behavior of the dystrophic growth cone, the hallmark of regeneration failure, in an in vitro model of the glial scar and after spinal cord injury. AB - We have developed a novel in vitro model of the glial scar that mimics the gradient of proteoglycan found in vivo after spinal cord injury. In this model, regenerated axons from adult sensory neurons that extended deeply into the gradient developed bulbous, vacuolated endings that looked remarkably similar to dystrophic endings formed in vivo. We demonstrate that despite their highly abnormal appearance and stalled forward progress, dystrophic endings are extremely dynamic both in vitro and in vivo after spinal cord injury. Time-lapse movies demonstrated that dystrophic endings continually send out membrane veils and endocytose large membrane vesicles at the leading edge, which were then retrogradely transported to the rear of the "growth cone." This direction of movement is contrary to membrane dynamics that occur during normal neurite outgrowth. As further evidence of this motility, dystrophic endings endocytosed large amounts of dextran both in vitro and in vivo. We now have an in vitro model of the glial scar that may serve as a potent tool for developing and screening potential treatments to help promote regeneration past the lesion in vivo. PMID- 15269265 TI - Pain encoding in the human forebrain: binary and analog exteroceptive channels. AB - The neuronal system signaling pain has often been characterized as a labeled line consisting of neurons in the pain-signaling pathway to the brain [spinothalamic tract (STT)] that respond only to painful stimuli. It has been proposed recently that the STT contains a series of analog labeled lines, each signaling a different aspect of the internal state of the body (interoception) (e.g., visceral-cold-itch sensations). In this view, pain is the unpleasant emotion produced by disequilibrium of the internal state. We now show that stimulation of an STT receiving zone in awake humans (66 patients) produces two different responses. The first is a binary response signaling the presence of painful stimuli. The second is an analog response in which nonpainful and painful sensations are graded with intensity of the stimulus. Compared with the second pathway, the first was characterized by higher pain ratings and stimulus-evoked sensations covering more of the body surface (projected fields). Both painful responses to stimulation were described in terms usually applied to external stimuli (exteroception) rather than to internal or emotional phenomena, which were infrequently evoked by stimulation of either pathway. These results are consistent with those of functional imaging studies that have identified brain regions activated in a binary manner by the application of a specific, painful stimulus while increases in stimulus intensity do not produce increased activation. Such binary pain functions could be involved in pain-related alarm alerting functions, which are independent of stimulus amplitude. PMID- 15269266 TI - Corticotropin-releasing factor receptors CRF1 and CRF2 exert both additive and opposing influences on defensive startle behavior. AB - The corticotropin-releasing factor (CRF) receptors (CRF1 and CRF2) are crucial mediators of physiological and behavioral responses to stress. In animals, CRF1 appears to primarily mediate CRF-induced anxiety-like responses, but the role of CRF2 during stress is still unclear. Here we report the effects of CRF1 and CRF2 on the magnitude and plasticity of defensive startle responses in mice. Startle plasticity is measured by inhibition of startle by sensory stimuli, i.e., prepulse inhibition (PPI), and is disrupted in patients with panic or posttraumatic stress disorders in which CRF neurotransmission may be overactive. Pharmacological blockade of CRF1 reversed both CRF-induced increases in startle and CRF-induced deficits in PPI. CRF2 blockade attenuated high-dose but not low dose CRF-induced increases in startle and reduced PPI. Conversely, activation of CRF2 enhanced PPI. CRF had no effect on startle and increased PPI in CRF1 knock out mice. These data indicate that CRF receptors act in concert to increase the magnitude of defensive startle yet in opposition to regulate the flexibility of startle. These data support a new model of respective CRF receptor roles in stress-related behavior such that, although both receptors enhance the magnitude of defensive responses, CRF1 receptors contravene, whereas CRF2 receptors enhance, the impact of sensory information on defensive behavior. We hypothesize that excessive CRF1 activation combined with reduced CRF2 signaling may contribute to information processing deficits seen in panic and posttraumatic stress disorder patients and support CRF1-specific pharmacotherapy. PMID- 15269267 TI - Neural activity protects hypothalamic magnocellular neurons against axotomy induced programmed cell death. AB - Axotomy typically leads to retrograde neuronal degeneration in the CNS. Studies in the hypothalamo-neurohypophysial system (HNS) have suggested that neural activity is supportive of magnocellular neuronal (MCN) survival after axotomy. In this study, we directly test this hypothesis by inhibiting neural activity in the HNS, both in vivo and in vitro, by the use of tetrodotoxin (TTX). After median eminence compression to produce axonal injury, unilateral superfusion of 3 microM TTX into the rat supraoptic nucleus (SON), delivered with the use of a miniature osmotic pump for 2 weeks in vivo, produced a decrease in the number of surviving MCNs in the TTX-treated SON, compared with the contralateral untreated side of the SON. In vitro application of 2.5 microM TTX for 2 weeks to the SON in organotypic culture produced a 73% decrease in the surviving MCNs, compared with untreated control cultures. Raising the extracellular KCl in the culture medium to 25 mM rescued the MCNs from the axotomy- and TTX-induced cell death. These data support the proposal that after axotomy, neural activity is neuroprotective in the HNS. PMID- 15269268 TI - Somatic localization of a specific large-conductance calcium-activated potassium channel subtype controls compartmentalized ethanol sensitivity in the nucleus accumbens. AB - Alcohol is an addictive drug that targets a variety of ion channels and receptors. To address whether the effects of alcohol are compartment specific (soma vs dendrite), we examined the effects of ethanol (EtOH) on large conductance calcium-activated potassium channels (BK) in cell bodies and dendrites of freshly isolated neurons from the rat nucleus accumbens (NAcc), a region known to be critical for the development of addiction. Compartment specific drug action was indeed observed. Clinically relevant concentrations of EtOH increased somatic but not dendritic BK channel open probability. Electrophysiological single-channel recordings and pharmacological analysis of the BK channel in excised patches from each region indicated a number of differences, suggestive of a compartment-specific expression of the beta4 subunit of the BK channel, that might explain the differential alcohol sensitivity. These parameters included activation kinetics, calcium dependency, and toxin blockade. Reverse transcription-PCR showed that both BK channel beta1 and beta4 subunit mRNAs are found in the NAcc, although the signal for beta1 is significantly weaker. Immunohistochemistry revealed that beta1 subunits were found in both soma and dendrites, whereas beta4 appeared restricted to the soma. These findings suggest that the beta4 subunit may confer EtOH sensitivity to somatic BK channels, whereas the absence of beta4 in the dendrite results in insensitivity to the drug. Consistent with this idea, acute EtOH potentiated alphabeta4 BK currents in transfected human embryonic kidney cells, whereas it failed to alter alphabeta1 BK channel-mediated currents. Finally, an EtOH concentration (50 mm) that increased BK channel open probability strongly decreased the duration of somatic-generated action potential in NAcc neurons. PMID- 15269269 TI - Shaggy, the homolog of glycogen synthase kinase 3, controls neuromuscular junction growth in Drosophila. AB - A protein-trap screen using the Drosophila neuromuscular junction (NMJ) as a model synapse was performed to identify genes that control synaptic structure or plasticity. We found that Shaggy (Sgg), the Drosophila homolog of the mammalian glycogen synthase kinases 3 alpha and beta, two serine-threonine kinases, was concentrated at this synapse. Using various combinations of mutant alleles of shaggy, we found that Shaggy negatively controlled the NMJ growth. Moreover, tissue-specific expression of a dominant-negative Sgg indicated that this kinase is required in the motoneuron, but not in the muscle, to control NMJ growth. Finally, we show that Sgg controlled the microtubule cytoskeleton dynamics in the motoneuron and that Futsch, a microtubule-associated protein, was required for Shaggy function on synaptic growth. PMID- 15269270 TI - Modulation of synaptic plasticity by antimanic agents: the role of AMPA glutamate receptor subunit 1 synaptic expression. AB - Increasing data suggest that impairments of cellular plasticity underlie the pathophysiology of bipolar disorder. In this context, it is noteworthy that AMPA glutamate receptor trafficking regulates synaptic plasticity, effects mediated by signaling cascades, which are targets for antimanic agents. The present studies were undertaken to determine whether two clinically effective, but structurally highly dissimilar, antimanic agents lithium and valproate regulate synaptic expression of AMPA receptor subunit glutamate receptor 1 (GluR1). Chronic (but not acute) treatment of rats with therapeutically relevant concentrations of lithium or valproate reduced hippocampal synaptosomal GluR1 levels. The reduction in synaptic GluR1 by lithium and valproate was attributable to a reduction of surface GluR1 distribution onto the neuronal membrane as demonstrated by three independent assays in cultured hippocampal neurons. Furthermore, these agents induced a decrease in GluR1 phosphorylation at a specific PKA site (GluR1p845), which is known to be critical for AMPA receptor insertion. Sp-cAMP treatment reversed the attenuation of phosphorylation by lithium and valproate and also brought GluR1 back to the surface, suggesting that phosphorylation of GluR1p845 is involved in the mechanism of GluR1 surface attenuation. In addition, GluR1p845 phosphorylation also was attenuated in hippocampus from lithium- or valproate treated animals in vivo. In contrast, imipramine, an antidepressant that can trigger manic episodes, increased synaptic expression of GluR1 in hippocampus in vivo. These studies suggest that regulation of glutamatergically mediated synaptic plasticity may play a role in the treatment of bipolar disorder and raise the possibility that agents more directly affecting synaptic GluR1 may represent novel therapies for this devastating illness. PMID- 15269271 TI - Mood stabilizer valproate promotes ERK pathway-dependent cortical neuronal growth and neurogenesis. AB - Manic-depressive illness has been conceptualized as a neurochemical illness. However, brain imaging and postmortem studies reveal gray-matter reductions, as well as neuronal and glial atrophy and loss in discrete brain regions of manic depressive patients. The roles of such cerebral morphological deficits in the neuropathophysiology and therapeutic mechanisms of manic-depressive illness are unknown. Valproate (2-propylpentanoate) is a commonly used mood stabilizer. The ERK (extracellular signal-regulated kinase) pathway is used by neurotrophic factors to regulate neurogenesis, neurite outgrowth, and neuronal survival. We found that chronic treatment of rats with valproate increased levels of activated phospho-ERK44/42 in neurons of the anterior cingulate, a region in which we found valproate-induced increases in expression of an ERK pathway-regulated gene, bcl 2. Valproate time and concentration dependently increased activated phospho ERK44/42 and phospho-RSK1 (ribosomal S6 kinase 1) levels in cultured cortical cells. These increases were attenuated by Raf and MEK (mitogen-activated protein kinase/ERK kinase) inhibitors. Although valproate affects the functions of GSK-3 (glycogen synthase kinase-3) and histone deacetylase (HDAC), its effects on the ERK pathway were not fully mimicked by selective inhibitors of GSK-3 or HDAC. Similar to neurotrophic factors, valproate enhanced ERK pathway-dependent cortical neuronal growth. Valproate also promoted neural stem cell proliferation maturation (neurogenesis), demonstrated by bromodeoxyuridine (BrdU) incorporation and double staining of BrdU with nestin, Tuj1, or the neuronal nuclei marker NeuN (neuronal-specific nuclear protein). Chronic treatment with valproate enhanced neurogenesis in the dentate gyrus of the hippocampus. Together, these data demonstrate that valproate activates the ERK pathway and induces ERK pathway mediated neurotrophic actions. This cascade of events provides a potential mechanism whereby mood stabilizers alleviate cerebral morphometric deficits associated with manic-depressive illness. PMID- 15269272 TI - Differential involvement of orbitofrontal cortex subregions in conditioned cue induced and cocaine-primed reinstatement of cocaine seeking in rats. AB - Orbitofrontal cortex (OFC) damage elicits impulsivity and perseveration, and impairments in OFC function may underlie compulsive drug seeking in cocaine users. To test this hypothesis, we assessed the effects of fiber-sparing lesions or functional inactivation of OFC subregions on cocaine seeking in rats. Rats were trained to lever press for intravenous cocaine (0.20 mg/infusion) paired with the presentations of light plus tone stimuli. Responding was then allowed to extinguish. Rats received bilateral NMDA (0.1 M) or sham lesions of the lateral OFC (lOFC) or medial OFC (mOFC) before self-administration training (experiment 1) or muscimol plus baclofen (0.1 and 1.0 mM) or vehicle infusions into the lOFC or mOFC before reinstatement testing (experiment 2). The effects of these manipulations on reinstatement of cocaine seeking (i.e., responding on the previously cocaine-paired lever) were assessed in the presence of the light plus tone stimuli or after a cocaine priming injection (10 mg/kg, i.p.). Post-training lOFC inactivation impaired conditioned cue-induced reinstatement, whereas other manipulations failed to alter this behavior. This suggests that the lOFC plays a critical role in assessing the current motivational significance of cocaine conditioned stimuli or in using this information to guide cocaine-seeking behavior if stimulus-reward learning takes place before lOFC damage. OFC inactivation failed to alter cocaine-primed reinstatement. However, lOFC lesions augmented cocaine-primed reinstatement in a perseverative manner, whereas mOFC lesions attenuated cocaine-primed reinstatement, suggesting that prolonged cell loss in OFC subregions may modulate the propensity for cocaine seeking in a subregion-specific manner. PMID- 15269273 TI - Processing of odor mixtures in the zebrafish olfactory bulb. AB - Components of odor mixtures often are not perceived individually, suggesting that neural representations of mixtures are not simple combinations of the representations of the components. We studied odor responses to binary mixtures of amino acids and food extracts at different processing stages in the olfactory bulb (OB) of zebrafish. Odor-evoked input to the OB was measured by imaging Ca2+ signals in afferents to olfactory glomeruli. Activity patterns evoked by mixtures were predictable within narrow limits from the component patterns, indicating that mixture interactions in the peripheral olfactory system are weak. OB output neurons, the mitral cells (MCs), were recorded extra- and intracellularly and responded to odors with stimulus-dependent temporal firing rate modulations. Responses to mixtures of amino acids often were dominated by one of the component responses. Responses to mixtures of food extracts, in contrast, were more distinct from both component responses. These results show that mixture interactions can result from processing in the OB. Moreover, our data indicate that mixture interactions in the OB become more pronounced with increasing overlap of input activity patterns evoked by the components. Emerging from these results are rules of mixture interactions that may explain behavioral data and provide a basis for understanding the processing of natural odor stimuli in the OB. PMID- 15269274 TI - cGMP-dependent kinase regulates response sensitivity of the mouse on bipolar cell. AB - The visual system can adjust its sensitivity over a wide range of light intensities. Photoreceptors account for some of this adjustment, but there is evidence that postreceptoral processes also exist. To investigate the latter, we pharmacologically mimicked the effects of light stimulation on mouse On bipolar cells, thus avoiding confounding effects of receptoral mechanisms. Here, we report that cGMP selectively enhances responses to dim, but not bright, stimuli through a purely postsynaptic mechanism. This action of cGMP was completely blocked by inhibitors of cGMP-dependent kinase. We propose that cGMP-dependent kinase decreases coupling of the On bipolar cell glutamate receptor to the downstream cascade, thus amplifying small decreases in photoreceptor transmitter levels that would otherwise go undetected by the visual system. PMID- 15269275 TI - A complete genetic analysis of neuronal Rab3 function. AB - Rab3A, Rab3B, Rab3C, and Rab3D are closely related GTP-binding proteins of synaptic vesicles that may function in neurotransmitter release. We have produced knock-out (KO) mice for Rab3B and Rab3C and crossed them with previously generated Rab3A and 3D knock-out mice to generate double, triple, and quadruple Rab3 knock-out mice. We have found that all single and double Rab3 knock-out mice are viable and fertile. Most triple Rab3 knock-out mice perish whenever Rab3A is one of the three deleted proteins, whereas all triple knock-out mice that express Rab3A are viable and fertile. Finally, all quadruple knock-out mice die shortly after birth. Quadruple Rab3 KO mice initially develop normally and are born alive but succumb to respiratory failure. Rab3-deficient mice display no apparent changes in synapse structure or brain composition except for a loss of rabphilin, a Rab3-binding protein. Analysis of cultured hippocampal neurons from quadruple knock-out mice uncovered no significant change in spontaneous or sucrose-evoked release but an approximately 30% decrease in evoked responses. This decrease was caused by a decline in the synaptic and the vesicular release probabilities, suggesting that Rab3 proteins are essential for the normal regulation of Ca2+ triggered synaptic vesicle exocytosis but not for synaptic vesicle exocytosis as such. Our data show that Rab3 is required for survival in mice and that the four Rab3 proteins are functionally redundant in this role. Furthermore, our data demonstrate that Rab3 is not in itself essential for synaptic membrane traffic but functions to modulate the basic release machinery. PMID- 15269276 TI - Rhesus macaque class I duplicon structures, organization, and evolution within the alpha block of the major histocompatibility complex. AB - The alpha block of the human and chimpanzee major histocompatibility complex (MHC) class I genomic region contains 10 to 11 duplicated MHC class I genes, including the HLA/Patr-A, -G, and -F genes. In comparison, the alpha block of the rhesus macaque (Macaca mulatta, Mamu) has an additional 20 MHC class I genes within this orthologous region. The present study describes the identification and analysis of the duplicated segmental genomic structures (duplicons) and genomic markers within the alpha block of the rhesus macaque and their use to reconstruct the duplication history of the genes within this region. A variety of MHC class I genes, pseudogenes, transposons, and retrotransposons, such as Alu and ERV16, were used to categorize the 28 duplicons into four distinct structural categories. The phylogenetic relationship of MHC class I genes, Alu, and LTR16B sequences within the duplicons was examined by use of the Neighbor-Joining (NJ) method. Two single-duplicon tandem duplications, two polyduplicon tandem duplications with an accompanying inversion product per duplication, eight polyduplicon tandem duplications steps, 12 deletions, and at least two recombinations were reconstructed to explain the highly complex organization and evolution of the 28 duplicons (nine inversions) within the Mamu alpha block. On the basis of the phylogenetic evidence and the reconstructed tandem duplication history of the 28 duplicons, the Mamu/Patr/HLA-F ortholog was the first MHC class I gene to have been fixed without further duplication within the alpha block of primates. Assuming that the rhesus macaque and the chimpanzee/human lineages had started with the same number of MHC class I duplicons at the time of their divergence approximately 24 to 31 MYA, then the number of genes within the alpha block have been duplicated at an approximately three times greater rate in the rhesus macaque than in either the human or chimpanzee. PMID- 15269277 TI - The molecular basis of adaptive evolution of squirrelfish rhodopsins. AB - The wavelengths of maximal absorption (lambdamax) of the rhodopsins of nine squirrelfishes (N. sammara, N. argenteus, S. punctatissimum, S. microstoma, S. diadema, S. xantherythrum, S. spiniferum, N. aurolineatus, and S. tiere) and two soldierfishes (M. violacea and M. berndti) vary between 481 and 502 nm. Phylogenetic and mutagenesis analyses suggest that the common ancestor of these pigments had a lambdamax value of approximately 493 nm, and the contemporary lambdamax values were generated mostly by amino acid replacements E122M, F261Y, and A292S. The probability of observing all these amino acid replacements at specific branches of the phylogenetic tree is only 2.5 x 10(-9); it is highly unlikely that these changes have occurred by neutral evolution. Because of a close association between the lambdamax values of these pigments and the wavelengths of light available to the corresponding species, the excess number of amino acid changes at specific branches in the phylogenetic tree strongly suggests that the rhodopsins have undergone adaptive changes at various stages of the holocentrid evolution. PMID- 15269278 TI - Transcription profiling of cyclic AMP signaling in Candida albicans. AB - We used transcription profiling in Candida albicans to investigate cellular regulation involving cAMP. We found that many genes require the adenylyl cyclase Cdc35p for proper expression. These include genes encoding ribosomal subunit proteins and RNA polymerase subunit proteins, suggesting that growth could be controlled in part by cAMP-mediated modulation of gene expression. Other genes influenced by loss of adenylyl cyclase are involved in metabolism, the cell wall, and stress response and include a group of genes of unknown function that are unique to C. albicans. The profiles generated by loss of the adenylyl cyclase regulator Ras1p and a downstream effector Efg1p were also examined. The loss of Ras1p function disturbs the expression of a subset of the genes regulated by adenylyl cyclase, suggesting both that the primary role of Ras1p in transcriptional regulation involves its influence on the function of Cdc35p and that there are Ras1p independent roles for Cdc35p. The transcription factor Efg1p is also needed for the expression of many genes; however, these genes are distinct from those modulated by Cdc35p with the exception of a class of hyphal specific genes. Therefore transcription profiling establishes that cAMP plays a key role in the overall regulation of gene expression in C. albicans, and enhances our detailed understanding of the circuitry controlling this regulation. PMID- 15269279 TI - Autoantigen Golgin-97, an effector of Arl1 GTPase, participates in traffic from the endosome to the trans-golgi network. AB - The precise cellular function of Arl1 and its effectors, the GRIP domain Golgins, is not resolved, despite our recent understanding that Arl1 regulates the membrane recruitment of these Golgins. In this report, we describe our functional study of Golgin-97. Using a Shiga toxin B fragment (STxB)-based in vitro transport assay, we demonstrated that Golgin-97 plays a role in transport from the endosome to the trans-Golgi network (TGN). The recombinant GRIP domain of Golgin-97 as well as antibodies against Golgin-97 inhibited the transport of STxB in vitro. Membrane-associated Golgin-97, but not its cytosolic pool, was required in the in vitro transport assay. The kinetic characterization of inhibition by anti-Golgin-97 antibody in comparison with anti-Syntaxin 16 antibody established that Golgin-97 acts before Syntaxin 16 in endosome-to-TGN transport. Knock down of Golgin-97 or Arl1 by their respective small interference RNAs (siRNAs) also significantly inhibited the transport of STxB to the Golgi in vivo. In siRNA treated cells with reduced levels of Arl1, internalized STxB was instead distributed peripherally. Microinjection of Golgin-97 antibody led to the fragmentation of Golgi apparatus and the arrested transport to the Golgi of internalized Cholera toxin B fragment. We suggest that Golgin-97 may function as a tethering molecule in endosome-to-TGN retrograde traffic. PMID- 15269280 TI - Kinetochore localization of spindle checkpoint proteins: who controls whom? AB - The spindle checkpoint prevents anaphase onset until all the chromosomes have successfully attached to the spindle microtubules. The mechanisms by which unattached kinetochores trigger and transmit a primary signal are poorly understood, although it seems to be dependent at least in part, on the kinetochore localization of the different checkpoint components. By using protein immunodepletion and mRNA translation in Xenopus egg extracts, we have studied the hierarchic sequence and the interdependent network that governs protein recruitment at the kinetochore in the spindle checkpoint pathway. Our results show that the first regulatory step of this cascade is defined by Aurora B/INCENP complex. Aurora B/INCENP controls the activation of a second regulatory level by inducing at the kinetochore the localization of Mps1, Bub1, Bub3, and CENP-E. This localization, in turn, promotes the recruitment to the kinetochore of Mad1/Mad2, Cdc20, and the anaphase promoting complex (APC). Unlike Aurora B/INCENP, Mps1, Bub1, and CENP-E, the downstream checkpoint protein Mad1 does not regulate the kinetochore localization of either Cdc20 or APC. Similarly, Cdc20 and APC do not require each other to be localized at these chromosome structures. Thus, at the last step of the spindle checkpoint cascade, Mad1/Mad2, Cdc20, and APC are recruited at the kinetochores independently from each other. PMID- 15269281 TI - Cell-specific nucleolar localization of TBP-related factor 2. AB - TATA-binding protein (TBP)-related factor 2 (TRF2) is one of four closely related RNA polymerase II transcription factors. We compared the intracellular localizations of TBP and TRF2 during the cell cycle and mitosis in HeLa cells. We show that during interphase, endogenous or exogenously expressed TRF2 is located almost exclusively in the nucleolus in HeLa or Cos cells. TRF2 localization is not affected by stress or mitotic stimuli, but TRF2 is rapidly released from the nucleolus upon inhibition of pol I transcription or treatment by RNase. These results suggest that localization of HeLa TRF2 requires a nucleolar-associated RNA species. In contrast, in 3T3 fibroblast cells, exogenously expressed TRF2 localizes to the nucleoplasm. Constitutive expression of ectopic TRF2 in 3T3 cells leads to a prolonged S phase of the cell cycle and reduced proliferation. Together with previous data, our results highlight the cell-specific localization and functions of TRF2. Furthermore, we show that during cell division, HeLa TRF2 and TBP are localized in the mitotic cytoplasm and TRF2 relocalizes into the nascent nucleoli immediately after mitosis, whereas TBP reassociates with the chromatin. Although partially contradictory results have been reported, our data are consistent with a model where only small proportion of the cellular TBP remains associated with specific promoter loci during mitosis. PMID- 15269282 TI - The essential role of PP1beta in Drosophila is to regulate nonmuscle myosin. AB - Reversible phosphorylation of myosin regulatory light chain (MRLC) is a key regulatory mechanism controlling myosin activity and thus regulating the actin/myosin cytoskeleton. We show that Drosophila PP1beta, a specific isoform of serine/threonine protein phosphatase 1 (PP1), regulates nonmuscle myosin and that this is the essential role of PP1beta. Loss of PP1beta leads to increased levels of phosphorylated nonmuscle MRLC (Sqh) and actin disorganisation; these phenotypes can be suppressed by reducing the amount of active myosin. Drosophila has two nonmuscle myosin targeting subunits, one of which (MYPT-75D) resembles MYPT3, binds specifically to PP1beta, and activates PP1beta's Sqh phosphatase activity. Expression of a mutant form of MYPT-75D that is unable to bind PP1 results in elevation of Sqh phosphorylation in vivo and leads to phenotypes that can also be suppressed by reducing the amount of active myosin. The similarity between fly and human PP1beta and MYPT genes suggests this role may be conserved. PMID- 15269283 TI - Glucose mediates transcriptional repression of the human angiotensin type-1 receptor gene: role for a novel cis-acting element. AB - Human angiotensin type 1 receptor (hAT1R) gene is regulated by hormones, second messengers, and both pathophysiological and developmental states. The focus of the present study was to determine the role of glucose in the trans-repression of hAT1R gene transcription and to identify the functional cis-acting response element(s). Serial deletions of the hAT1R promoter region indicated that an area between -1717 and -1543 base pairs upstream of the 5' end of the cDNA sequence has a glucose responsive regulatory element (GluRE) to down-regulate the gene expression. Further analysis revealed a putative 29-bp (5' AACTGATTTTTGTATATTGATCTTGTATT-3') repressor element located between -1582 and 1610 bp was necessary for transcriptional repression. Removal of this region from promoter construct abolished repression of the hAT1R gene transcription in human proximal tubule epithelial cells (hPTECs). Using mobility shift assays, we demonstrated DNA binding activity to the labeled repressor element in hPTEC nuclear extracts. Additional studies demonstrated increased DNA binding activity to the labeled repressor element in nuclear extracts treated with high glucose (25 mM). Southwestern analysis identified two GluRE binding proteins of 34 and 36 kDa in glucose-treated extracts. Glucose-induced activity of the repressor trans acting factor(s) reached a maximum at 4 h, which correlated with decreased transcriptional activity of the hAT1R gene, suggesting that glucose can down regulate the transcription of the hAT1R gene through the repressor element. Furthermore, insertion of the glucose response element into heterologous SV40 promoter (SV40) chloramphenicol acetyl transferase (CAT) vector showed orientation/distance-independent repression of SV40 promoter-mediated CAT activity in hPTECs. Our results show that the glucose response factor(s) acts as trans-acting factor(s) binding to the cis-acting repressor element in the hAT1R promoter, which may participate in the control of basal transcription as well as glucose-mediated transcriptional inhibition of the hAT1R gene. PMID- 15269284 TI - TetraThymosinbeta is required for actin dynamics in Caenorhabditis elegans and acts via functionally different actin-binding repeats. AB - Generating specific actin structures via controlled actin polymerization is a prerequisite for eukaryote development and reproduction. We here report on an essential Caenorhabditis elegans protein tetraThymosinbeta expressed in developing neurons and crucial during oocyte maturation in adults. TetraThymosinbeta has four repeats, each related to the actin monomer sequestering protein thymosinbeta 4 and assists in actin filament elongation. For homologues with similar multirepeat structures, a profilin-like mechanism of ushering actin onto filament barbed ends, based on the formation of a 1:1 complex, is proposed to underlie this activity. We, however, demonstrate that tetraThymosinbeta binds multiple actin monomers via different repeats and in addition also interacts with filamentous actin. All repeats need to be functional for attaining full activity in various in vitro assays. The activities on actin are thus a direct consequence of the repeated structure. In containing both G- and F-actin interaction sites, tetraThymosinbeta may be reminiscent of nonhomologous multimodular actin regulatory proteins implicated in actin filament dynamics. A mutation that suppresses expression of tetraThymosinbeta is homozygous lethal. Mutant organisms develop into adults but display a dumpy phenotype and fail to reproduce as their oocytes lack essential actin structures. This strongly suggests that the activity of tetraThymosinbeta is of crucial importance at specific developmental stages requiring actin polymerization. PMID- 15269285 TI - A novel mechanism for mitogen-activated protein kinase localization. AB - Mitogen-activated protein kinases/extracellular signal regulated kinases (MAPKs/ERKs) are typically thought to be soluble cytoplasmic enzymes that translocate to the nucleus subsequent to their phosphorylation by their activating kinases or mitogen-activated protein/extracellular signal regulated kinase kinase. We report here the first example of nuclear translocation of a MAPK that occurs via temporally regulated exit from a membranous organelle. Confocal microscopy examining the subcellular localization of ERK3 in several cell lines indicated that this enzyme was targeted to the Golgi/endoplasmic reticulum Golgi intermediate compartment. Deletion analysis of green fluorescent protein (GFP)-ERK3 uncovered a nuclear form that was carboxy-terminally truncated and established a Golgi targeting motif at the carboxy terminus. Immunoblot analysis of cells treated with the proteasome inhibitor MG132 further revealed two cleavage products, suggesting that in vivo, carboxy-terminal cleavage of the full-length protein controls its subcellular localization. In support of this hypothesis, we found that deletion of a small region rich in acidic residues within the carboxy terminus eliminated both the cleavage and nuclear translocation of GFP-ERK3. Finally, cell cycle synchronization studies revealed that the subcellular localization of ERK3 is temporally regulated. These data suggest a novel mechanism for the localization of an MAPK family member, ERK3, in which cell cycle-regulated, site-specific proteolysis generates the nuclear form of the protein. PMID- 15269287 TI - Sulfasalazine down-regulates the expression of the angiogenic factors platelet derived endothelial cell growth factor/thymidine phosphorylase and interleukin-8 in human monocytic-macrophage THP1 and U937 cells. AB - Platelet-derived endothelial cell growth factor/thymidine phosphorylase (PD ECGF/TP) and interleukin-8 (IL-8) are angiogenic factors produced by tumor infiltrating macrophages. Here, we show that prolonged exposure of human monocytic/macrophage THP1 and U937 cells to sulfasalazine, an anti-inflammatory drug and inhibitor of nuclear factor-kappaB (NF-kappaB), resulted in down regulation of PD-ECGF/TP and IL-8 (mRNA, protein and activity) along with elimination of their induction by tumor necrosis factor-alpha and interferon gamma. Concomitantly, sulfasalazine-exposed cells were markedly resistant to 5' deoxyfluorouridine, the last intermediate of capecitabine requiring activation by PD-ECGF/TP. This is the first report suggesting that disruption of NF-kappaB dependent signaling pathways can provoke a marked and sustained down-regulation of macrophage-related angiogenic factors. However, this may also negatively affect capecitabine efficacy. PMID- 15269288 TI - Stannin, a protein that localizes to the mitochondria and sensitizes NIH-3T3 cells to trimethyltin and dimethyltin toxicity. AB - Stannin (Snn) is a highly conserved, 88-amino acid protein that may mediate the selective toxicity of organotins. Snn is localized in tissues with known sensitivity to trimethyltin (TMT), including the central nervous system, immune system, spleen, kidney and lung. Cells in culture that do not express Snn show considerable resistance to TMT toxicity. In vitro, Snn peptide can bind TMT in a 1:1 ratio and can de-alkylate TMT to dimethyltin (DMT). We now show that transfection with Snn sensitized TMT-resistant NIH-3T3 mouse fibroblasts to both TMT and DMT cytotoxicity. Triple label confocal microscopy of Snn-transfected cells and Percoll gradient purification of mitochondria showed Snn localized to the mitochondria and other membrane structures. The mitochondrial localization of Snn, coupled with its ability to bind and dealkylate organotin compounds, indicates a possible mechanism by which selective alkyltin toxicity might be mediated. PMID- 15269286 TI - A dynein light intermediate chain, D1bLIC, is required for retrograde intraflagellar transport. AB - Intraflagellar transport (IFT), the bidirectional movement of particles along flagella, is essential for flagellar assembly. The motor for retrograde IFT in Chlamydomonas is cytoplasmic dynein 1b, which contains the dynein heavy chain DHC1b and the light intermediate chain (LIC) D1bLIC. To investigate a possible role for the LIC in IFT, we identified a d1blic mutant. DHC1b is reduced in the mutant, indicating that D1bLIC is important for stabilizing dynein 1b. The mutant has variable length flagella that accumulate IFT-particle proteins, indicative of a defect in retrograde IFT. Interestingly, the remaining DHC1b is normally distributed in the mutant flagella, strongly suggesting that the defect is in binding of cargo to the retrograde motor rather than in motor activity per se. Cell growth and Golgi apparatus localization and morphology are normal in the mutant, indicating that D1bLIC is involved mainly in retrograde IFT. Like mammalian LICs, D1bLIC has a phosphate-binding domain (P-loop) at its N-terminus. To investigate the function of this conserved domain, d1blic mutant cells were transformed with constructs designed to express D1bLIC proteins with mutated P loops. The constructs rescued the mutant cells to a wild-type phenotype, indicating that the function of D1bLIC in IFT is independent of its P-loop. PMID- 15269289 TI - Bilirubin: an endogenous product of heme degradation with both cytotoxic and cytoprotective properties. PMID- 15269290 TI - Ca2+ channels as integrators of G protein-mediated signaling in neurons. AB - The observations from Dunlap and Fischbach that transmitter-mediated shortening of the duration of action potentials could be caused by a decrease in calcium conductance led to numerous studies of the mechanisms of modulation of voltage dependent calcium channels. Calcium channels are well known targets for inhibition by receptor-G protein pathways, and multiple forms of inhibition have been described. Inhibition of Ca(2+) channels can be mediated by G protein betagamma-subunits or by kinases, such as protein kinase C and tyrosine kinases. In the last few years, it has been shown that integration of G protein signaling can take place at the level of the calcium channel by regulation of the interaction of the channel pore-forming subunit with different cellular proteins. PMID- 15269291 TI - Analytical validation of telomerase activity for cancer early detection: TRAP/PCR CE and hTERT mRNA quantification assay for high-throughput screening of tumor cells. AB - Activation of telomerase plays a critical role in unlimited proliferation and immortalization of cells. Telomerase activity has been shown to correlate with tumor progression, indicating that tumors expressing this enzyme possess aggressive clinical behavior and that telomerase activity may be a useful biomarker for early detection of cancer. However, measurements of telomerase activity by current methods such as telomeric repeat amplification protocol (TRAP)/polymerase chain reaction (PCR) or antibody-based radioimmunoassay (RIA) are low-throughput and not robust enough to easily accommodate the required statistical analysis to determine whether telomerase activity is a practical biomarker. As part of the National Cancer Institute Early Detection Research Network of analytical validation, we have developed a robot assisted TRAP assay (RApidTRAP) of telomerase, a potential biomarker for cancer early detection. Measurements of human telomerase reverse transcriptase catalytic subunit (hTERT) mRNA were performed in concert with measurement of telomerase activity. For this purpose we determined hTERT mRNA concentration and telomerase activity in human normal (RPE-28) and cancer (A549) cell lines as well as in human serum (SRM 1951A). Telomerase activity measurements were made using the TRAP/PCR capillary electrophoresis (CE) method on (50 to 1000) cells/reaction isolated from cell extracts. Measurement of hTERT mRNA was made using specific primers and probes on a LightCycler in the range of (10 to 7000) cells/reaction. Comparison of high throughput telomerase activity measurements using the robot and those performed manually were consistent in sensitivity and reproducibility. Using this combination of telomerase activity and hTERT mRNA measurements, the automated system improved efficiency over traditional TRAP/PCR methods. PMID- 15269292 TI - Comparative genomic hybridization analysis of astrocytomas: prognostic and diagnostic implications. AB - Astrocytoma is comprised of a group of common intracranial neoplasms that are classified into four grades based on the World Health Organization histological criteria and patient survival. To date, histological grade, patient age, and clinical performance, as reflected in the Karnofsky score, are the most reliable prognostic predictors. Recently, there has been a significant effort to identify additional prognostic markers using objective molecular genetic techniques. We believe that the identification of such markers will characterize new chromosomal loci important in astrocytoma progression and aid clinical diagnosis and prognosis. To this end, our laboratory used comparative genomic hybridization to identify DNA sequence copy number changes in 102 astrocytomas. Novel losses of 19p loci were detected in low-grade pilocytic astrocytomas and losses of loci on 9p, 10, and 22 along with gains on 7, 19, and 20 were detected in a significant proportion of high-grade astrocytomas. The Cox proportional hazards statistical modeling showed that the presence of +7q and -10q comparative genomic hybridization alterations significantly increased a patient's risk of dying, independent of histological grade. This investigation demonstrates the efficacy of comparative genomic hybridization for identifying tumor suppressor and oncogene loci in different astrocytic grades. The cumulative effect of these loci is an important consideration in their diagnostic and prognostic implications. PMID- 15269293 TI - Gene expression screening of salivary gland neoplasms: molecular markers of potential histogenetic and clinical significance. AB - Salivary gland neoplasms comprise phenotypically and biologically diverse lesions of uncertain histogenesis. The molecular events associated with their development and clinicopathological heterogeneity remain unknown. To reveal these events, we performed microarray expression analysis using a nylon-filter membrane platform on 18 primary lesions representing the most common benign and malignant types. Our study identified a small set of genes that are differentially altered between normal salivary gland tissues and benign and malignant tumors. Of the 5000 genes arrayed, 136 genes were differentially expressed by normal tissue, benign tumors, and various malignant neoplasms. Hierarchical clustering analysis differentiated between adenoid cystic carcinomas (ACCs) and other malignant subtypes. Non-ACC specimens manifested overlapping patterns of gene expression within and between tumors. Most of the differentially expressed genes share functional similarities with members of the adhesion, proliferation, and signal transduction pathways. Our study identified: 1) a set of genes that differentiate normal tissue from tumor specimens, 2) genes that differentiate pleomorphic adenoma and ACCs from other malignant salivary gland neoplasms, and 3) different patterns of expression between ACCs arising from major and minor salivary gland sites. The differentially expressed genes provide new information on potential genetic events of biological significance in future studies of salivary gland tumorigenesis. PMID- 15269294 TI - Direct detection of Staphylococcus aureus from adult and neonate nasal swab specimens using real-time polymerase chain reaction. AB - Nasal carriage of Staphylococcus aureus is considered a source of subsequent infection in health care settings. Utilization of real-time polymerase chain reaction (PCR) for detection of S. aureus has the potential to dramatically affect infection control practice by rapidly identifying S. aureus-colonized patients. We developed and validated the use of real-time PCR for detection of S. aureus colonization in two patient populations. Paired nasal swabs were collected from 299 neonates and from 151 adult patients at Evanston Hospital. One swab was used for culture and the other placed into a bacterial lysis solution containing achromopeptidase. The DNA liberated was used as the template for real-time PCR with primers for the femA gene. SYBR Green was used for amplicon detection. In the neonatal population the sensitivity, specificity, predictive value positive and predictive value negative for culture and PCR was 92% versus 96%, 100% versus 100%, 100% versus 100%, and 98% versus 99%, respectively. In the adults the results were 90% versus 100%, 100% versus 98%, 100% versus 96%, and 95% versus 100%, respectively. Real-time PCR was able to detect S. aureus in 2 hours compared to 1 to 4 days for culture and provided sensitivity equal to or greater than culture. PMID- 15269295 TI - Association of platelet-derived growth factor receptor alpha mutations with gastric primary site and epithelioid or mixed cell morphology in gastrointestinal stromal tumors. AB - Most gastrointestinal stromal tumors (GISTs) carry activating mutations of the KIT gene encoding the receptor tyrosine kinase KIT. In a previous study we were able to show an association between the lack of KIT mutations (wild-type GISTs) and the presence of a significant epithelioid tumor component. A very recent study described the occurrence of PDGFRalpha mutations in KIT wt GISTS. Therefore, we studied a panel of 87 GISTs for mutations in the hot spot regions of the PDGFRalpha gene with single strand conformation polymorphism analysis and sequencing and correlated the PDGFRalpha status with pathomorphological data. We detected 20 cases with exon 18 mutations but none with exon 12 mutations. The mutations were located in the second kinase domain of PDGFRalpha with 16 point mutations, and four larger deletions of 9 to 12 bp. All cases with mutations in the PDGFRalpha gene revealed wild-type KIT in common regions of mutation, ie, exons 9 and 11. Most interestingly, the occurrence of PDGFRalpha mutations was significantly associated with a higher frequency of epithelioid or mixed morphology (18 of 20 cases, P < 0.0001) and gastric location (all cases, P = 0.0008). Our data indicate that GISTs represent distinctive entities, differing in genetic, biological, and morphological features. PMID- 15269296 TI - A study of MECT1-MAML2 in mucoepidermoid carcinoma and Warthin's tumor of salivary glands. AB - The t(11;19)(q21;p13) chromosomal translocation has been described in two distinct types of salivary gland neoplasms: mucoepidermoid carcinoma (MEC) and Warthin's tumor (WT). Since this translocation has been recently shown to generate a MECT1-MAML2 fusion gene, we evaluated 10 primary MEC and seven primary WT to further define the molecular association of these two entities using cytogenetic, as well as in situ hybridization (ISH) and reverse transcriptase polymerase chain reaction (RT-PCR) analyses directed against the fusion gene. A karyotype was established in all neoplasms except for two MEC cases. Of the eight karyotyped MECs, five showed the t(11;19)(q21;p13), two had a normal karyotype, and one case presented a -Y and +X. Three of the WT revealed a normal karyotype and four had several abnormalities which did not involve chromosomes 11 and 19. ISH analysis performed in cytogenetic suspension and/or in tumor paraffin sections demonstrated MAML2 rearrangement in 7 of 10 cases of MEC: all five cases with t(11;19), one case with normal karyotype, and one unkaryotyped case. RT-PCR analysis confirmed the expression of the MECT1-MAML2 gene in all MEC cases that were positive by ISH analysis. Neither the t(11;19) nor MECT1-MAML2 was detected in any case of WT, nor in control samples from polymorphous low-grade adenocarcinoma, acinic cell carcinoma, or normal parotid gland tissue. We have demonstrated that ISH and RT-PCR are sensitive methods for detecting MECT1-MAML2 in MEC. In contrast, we did not detect the t(11;19) nor MECT1-MAML2 expression in seven cases of WT. PMID- 15269297 TI - High-resolution melting analysis for detection of internal tandem duplications. AB - High-resolution melting analysis (HRMA) is a recently introduced closed-tube fluorescence-based method for rapid mutation screening and detection. However, all of the targets by which this technique has been validated thus far have had single-base substitutions, deletions, or similarly small mutational deviations from the wild-type sequence. In the current study, we sought to determine the feasibility of utilization of HRMA for the detection of larger sequence aberrations, using internal tandem duplications (ITD) in the juxtamembrane domain of the FLT3 gene as a model system. This gene is important in the growth and differentiation of hematopoietic progenitors and ITDs in this gene have been identified in a subset of poor-prognosis acute myelogenous leukemias (AML). DNA extracted from 62 AML samples was analyzed on a prototype high-resolution melting instrument. The samples interrogated for the FLT3 ITDs were subjected to post amplification denaturation with frequent and regular fluorescence acquisition. The fluorescence versus temperature melting graphs generated were analyzed for deviation from the profiles reproducibly obtained for the wild-type samples. Results by HRMA were compared to results obtained using capillary electrophoresis based fragment analysis, temperature gradient capillary electrophoresis detection, and sequencing of ITDs. FLT3 ITDs were detected in 13 of 62 AML samples with 100% concordance between the detection methods. This study demonstrates the utility of HRMA to rapidly and accurately screen samples for the presence of large sequence aberrations including FLT3 ITDs. PMID- 15269298 TI - Establishment of real-time polymerase chain reaction method for quantitative analysis of asparagine synthetase expression. AB - We established a real-time quantitative PCR (RQ-PCR) with which to measure abundance of the asparagine synthetase (AS) mRNA. The level of AS mRNA paralleled AS enzyme activity, as well as the AS protein level detected by Western blotting and by in situ immunostaining. Cytotoxicity tests in vitro showed that the AS mRNA level also synchronized with cellular resistance to L-asparaginase in cell lines. Cellular levels of AS enzyme activity correlated with resistance to L asparaginase. These results indicate that the AS mRNA level is an index of resistance to L-asparaginase. RQ-PCR is superior to enzyme assays, Western blotting, and immunostaining in the following ways: less labor and time, accurate and reproducible quantitativity, and broad dynamic range. In addition, RQ-PCR could evaluate differences in L-asparaginase sensitivity although immunostaining could not. And in clinical samples, we analyzed eight pediatric leukemia cases by this RQ-PCR to evaluate whether this method was applicable to clinical laboratories and the expression level of AS mRNA in each case were predictable for the effectiveness of L-asparaginase treatment. Consequently, this method was useful enough in defining candidates for selective therapy that targets an AS deficiency. PMID- 15269299 TI - Increased sensitivity of the Roche COBAS AMPLICOR HCV test, version 2.0, using modified extraction techniques. AB - Processing modifications were made to the COBAS AMPLICOR HCV version 2.0 assay to enhance sensitivity. Two methods of specimen concentration, centrifugation ("ultraspin") and cationic detergent plus silica membrane ("ultracolumn"), were compared to the standard method. The effect of these changes on assay sensitivity and specificity was examined using commercial hepatitis C virus (HCV) preparations. The limits of detection (LOD, defined as detection of HCV RNA in >/= 95% of replicates) of genotype 1a were 50, 12, and 6 by standard method, ultraspin and ultracolumn, respectively. For genotype 1b, the LOD was 25 IU/ml, 12 IU/ml, and 3 IU/ml; for 2b, it was 50, 12, and 3; for 3a, it was 25, 12, and 1.5; for 4 it was 18, 4, and 2; for 5a, it was 38, 9, and 2; and for 6a it was 47, 6, and 3. No false positives were detected after ultraspin when controls containing high or low HCV concentrations were alternated with normal human plasma. Plasmas in which HCV RNA was not detected by the standard assay were re tested with modified methods to assess the effect of altered processing in clinical specimens. Three of 152 specimens with no detectable HCV RNA by the standard method were positive by ultraspin and 2 of 109 were positive by ultracolumn, suggesting that these methods may increase assay sensitivity in clinical specimens. PMID- 15269300 TI - Detection of Mycobacterium tuberculosis complex in formalin-fixed, paraffin embedded tissue specimens with necrotizing granulomatous inflammation by strand displacement amplification. AB - Rapid, reliable diagnosis of tuberculosis is essential to initiate correct treatment, avoid severe complications, and prevent transmission. Conventional microbiological methods may not be an option if samples are formalin-fixed and paraffin-embedded (FFPE) for histopathological examination. With the demonstration of necrotizing granulomatous inflammation, tuberculosis becomes an important differential diagnosis, although it was not initially suspected. Following paraffin extraction, BDProbeTec ET strand displacement amplification for detection of Mycobacterium tuberculosis complex (MTC) was applied to 47 prospectively and 19 retrospectively collected FFPE samples from various sources with granulomatous inflammation and results were compared to tuberculosis notification. Of the prospective samples, 20 were from patients who were notified as having tuberculosis and the assay was positive in 18 (90%). Specificity was 100%. For 27 of the patients with prospectively collected FFPE specimens, culture was performed on a specimen collected at a later date from the same location. Culture revealed MTC in 14 and nontuberculous mycobacteria in four. BDProbeTec ET was positive in 13 (92.8%) of the patients with positive MTC culture and negative in the remaining. The sensitivity and specificity in 19 archival samples was 40% and 100%, respectively, compared to notification data. The assay provided rapid, correct diagnosis on different sources of FFPE samples collected prospectively and therefore offers an important supplementary method for patients where tuberculosis was not initially suspected. PMID- 15269301 TI - Isothermal multiple displacement amplification: a highly reliable approach for generating unlimited high molecular weight genomic DNA from clinical specimens. AB - Isothermal multiple strand displacement amplification (IMDA) of the whole human genome is a promising method for procuring abundant DNA from valuable and often limited clinical specimens. However, whether DNA generated by this method is of high quality and a faithful replication of the DNA in the original specimen, allowing for subsequent molecular diagnostic testing, requires verification. In this study, we evaluated the suitability of IMDA-generated DNA (IMDA-DNA) for detecting antigen receptor gene rearrangements, chromosomal translocations, and gene mutations using Southern blot analysis, polymerase chain reaction (PCR) methods, or sequencing methods in 28 lymphoma and leukemia clinical specimens. Molecular testing before and after whole genome amplification of these specimens using the IMDA technique showed concordance in 27 of 28 (96%) specimens. Analysis of IMDA-DNA by Southern blot analysis detected restriction fragments >12 kilobases long. No amplification bias was observed at all loci tested demonstrating that this method can be useful in generating large amounts of unbiased, high molecular weight DNA from limited clinical specimens. PMID- 15269302 TI - Allelic imbalance of 8p indicates poor survival in gastric cancer. AB - Gastric cancer is a common tumor worldwide and a tremendous health burden. However, the underlying mechanisms of tumorigenesis in this cancer's development are primarily undefined. Allelic imbalance (AI) of 8p has been reported in many cancers, yet, the target(s) of alteration and the importance of allelic imbalance on this chromosomal arm in gastric carcinoma development remained to be characterized. Our findings confirmed a high rate of AI on 8p in gastric cancers. Moreover, we demonstrated that AI on 8p, either overall or at marker D8S560, was associated with poorer survival in patients with gastric cancer. Finally, gastric cancers with a high rate of microsatellite instability were significantly associated with noncardia tumors and with female gender. PMID- 15269303 TI - Quantification of melanoma mRNA markers in sentinel nodes: pre-clinical evaluation of a single-step real-time reverse transcriptase-polymerase chain reaction assay. AB - Although reverse transcriptase-polymerase chain reaction (RT-PCR) analysis of melanocyte-associated mRNA can detect sentinel node melanoma metastases, most published assays are semi-quantitative methods of unknown sensitivity and precision, unsuitable for clinical use. We describe a single-step real-time quantitative RT-PCR assay for MART-1 and tyrosinase mRNAs, suitable for sentinel node analysis in a clinical setting. Using serial dilutions of melanoma cell line SK-MEL-28 RNA in water as a calibrator, we obtained linear calibration curves covering the range 0.5 to 10,000 arbitrary units (SK-MEL-28 melanoma cell equivalents). The sensitivity limit was 0.32 (MART-1) and 5 (tyrosinase) arbitrary units. Analytical imprecision was between 11% and 34%. MART-1 PCR efficiency was unaffected when samples were diluted with negative lymph node RNA rather than water, whereas tyrosinase PCR efficiency was halved. To evaluate the clinical suitability of our assay, we quantified melanocyte mRNAs in sentinel nodes with histologically verified micrometastases (n = 10) and benign nevus inclusions (n = 10), and in sentinel nodes without evidence of intranodal melanocytes (n = 10). We found significant differences in median melanocyte derived mRNA levels comparing the three types of lymph nodes, suggesting that this quantitative molecular protocol may increase assay precision and be useful for the clinical evaluation of sentinel nodes. PMID- 15269304 TI - Homogeneous polymerase chain reaction nucleobase quenching assay to detect the 1 kbp deletion in CLN3 that causes Batten disease. AB - Batten disease is an autosomal recessive disorder also known as juvenile neuronal ceroid lipofuscinosis. The most common mutation for this disease is an approximately 1-kbp deletion in the CLN3 gene, which accounts for about 80 to 85% of the mutation load. We developed a rapid assay for this mutation using the PCR to produce amplicons that are detected by nucleobase quenching of the fluorescent signal from a probe labeled with a fluorescent dye. The probe overlaps the deletion breakpoint and is completely base paired to the mutant amplicon. However, three bases at the 5' end of the probe do not base pair with the wild type amplicon. The alleles are distinguished by the different melting temperatures of the probe amplicon hybrids. Comparison of this new method with an allele-specific PCR and gel electrophoresis-based method showed 100% concordance in determination of the genotype for 30 specimens (11 homozygous mutant, 8 heterozygotes, and 11 homozygous normal). PCR followed by allele-specific melting curve analysis using nucleobase quenching has utility as a rapid method for detection of the most common mutation that causes Batten disease. PMID- 15269305 TI - Long-range (17.7 kb) allele-specific polymerase chain reaction method for direct haplotyping of R117H and IVS-8 mutations of the cystic fibrosis transmembrane regulator gene. AB - Genotyping of genetic polymorphisms is widely used in clinical molecular laboratories to confirm or predict diseases due to single locus mutations. In contrast, very few molecular methods determine the phase or haplotype of two or more mutations that are kilobases apart. In this report, we describe a new method for haplotyping based on long-range allele-specific PCR. Reaction conditions were established to circumvent the incompatibility of using allele-specific primers and a polymerase with proofreading activity. Haplotypes are determined by post PCR analysis using different detection methods. The clinical application presented here directly determines the phase of two mutations separated by 17.7 kilobases in the cystic fibrosis transmembrane conductance regulator gene. Each mutation, the missense mutation R117H in exon 4 and the 5T polymorphism in intron 8 (IVS-8), have mild phenotypic effect unless they are present on the same chromosome (in cis). If an individual is heterozygous for both R117H and the IVS 8 5T variant, cis/trans testing is required to completely interpret results. The molecular method presented here bypasses the need to perform family studies to establish haplotypes. We propose use of this assay as a reflex clinical test for R117H- 5T-positive samples. PMID- 15269306 TI - Fluorescence in situ hybridization identifies cryptic t(16;16)(p13;q22) masked by del(16)(q22) in a case of AML-M4 Eo. AB - We report a patient presenting with acute myeloid leukemia (AML)-M4 Eo, in whom conventional cytogenetic analysis revealed a 46, XY, del(16)(q22) karyotype. Molecular analysis of the bone marrow cells using reverse transcriptase polymerase chain reaction (RT-PCR) identified a CBFbeta-MYH11, "type A" fusion transcript. However, despite a thorough reevaluation, a balanced chromosome 16 abnormality could not be definitively identified by cytogenetics. Since there exists a small possibility of obtaining a false-positive PCR result, fluorescence in situ hybridization (FISH) analysis using dual-color, break-apart probes for CBFbeta was performed to elucidate the mechanism of fusion gene formation and thus confirm the RT-PCR results. FISH analysis clearly revealed a cryptic t(16;16), which was probably masked by the del(16)(q22). FISH is the preferred diagnostic procedure to elucidate the CBFbeta-MYH11 fusion in this situation, and resolves the possibility of both false-positive and false-negative results with RT-PCR technique. Due to the improved prognosis of AML associated with the CBFbeta-MYH11 fusion compared to AML generally, we recommend the use of FISH for detection of inv(16)/t(16;16)/CBFbeta-MYH11 in patients with failed, complex, or apparently normal cytogenetics, and in whom the cell morphology indicates the strong possibility of this gene fusion. PMID- 15269307 TI - Public registration of clinical trials. PMID- 15269308 TI - The price tag on progress--chemotherapy for colorectal cancer. PMID- 15269309 TI - Age-related macular degeneration and the extracellular matrix. PMID- 15269310 TI - Vestibular neuritis, or driving dizzily through Donegal. PMID- 15269311 TI - Translational implications of the parathyroid calcium receptor. PMID- 15269312 TI - Higher versus lower positive end-expiratory pressures in patients with the acute respiratory distress syndrome. AB - BACKGROUND: Most patients requiring mechanical ventilation for acute lung injury and the acute respiratory distress syndrome (ARDS) receive positive end expiratory pressure (PEEP) of 5 to 12 cm of water. Higher PEEP levels may improve oxygenation and reduce ventilator-induced lung injury but may also cause circulatory depression and lung injury from overdistention. We conducted this trial to compare the effects of higher and lower PEEP levels on clinical outcomes in these patients. METHODS: We randomly assigned 549 patients with acute lung injury and ARDS to receive mechanical ventilation with either lower or higher PEEP levels, which were set according to different tables of predetermined combinations of PEEP and fraction of inspired oxygen. RESULTS: Mean (+/-SD) PEEP values on days 1 through 4 were 8.3+/-3.2 cm of water in the lower-PEEP group and 13.2+/-3.5 cm of water in the higher-PEEP group (P<0.001). The rates of death before hospital discharge were 24.9 percent and 27.5 percent, respectively (P=0.48; 95 percent confidence interval for the difference between groups, -10.0 to 4.7 percent). From day 1 to day 28, breathing was unassisted for a mean of 14.5+/-10.4 days in the lower-PEEP group and 13.8+/-10.6 days in the higher-PEEP group (P=0.50). CONCLUSIONS: These results suggest that in patients with acute lung injury and ARDS who receive mechanical ventilation with a tidal-volume goal of 6 ml per kilogram of predicted body weight and an end-inspiratory plateau pressure limit of 30 cm of water, clinical outcomes are similar whether lower or higher PEEP levels are used. PMID- 15269313 TI - Cetuximab monotherapy and cetuximab plus irinotecan in irinotecan-refractory metastatic colorectal cancer. AB - BACKGROUND: The epidermal growth factor receptor (EGFR), which participates in signaling pathways that are deregulated in cancer cells, commonly appears on colorectal-cancer cells. Cetuximab is a monoclonal antibody that specifically blocks the EGFR. We compared the efficacy of cetuximab in combination with irinotecan with that of cetuximab alone in metastatic colorectal cancer that was refractory to treatment with irinotecan. METHODS: We randomly assigned 329 patients whose disease had progressed during or within three months after treatment with an irinotecan-based regimen to receive either cetuximab and irinotecan (at the same dose and schedule as in a prestudy regimen [218 patients]) or cetuximab monotherapy (111 patients). In cases of disease progression, the addition of irinotecan to cetuximab monotherapy was permitted. The patients were evaluated radiologically for tumor response and were also evaluated for the time to tumor progression, survival, and side effects of treatment. RESULTS: The rate of response in the combination-therapy group was significantly higher than that in the monotherapy group (22.9 percent [95 percent confidence interval, 17.5 to 29.1 percent] vs. 10.8 percent [95 percent confidence interval, 5.7 to 18.1 percent], P=0.007). The median time to progression was significantly greater in the combination-therapy group (4.1 vs. 1.5 months, P<0.001 by the log-rank test). The median survival time was 8.6 months in the combination-therapy group and 6.9 months in the monotherapy group (P=0.48). Toxic effects were more frequent in the combination-therapy group, but their severity and incidence were similar to those that would be expected with irinotecan alone. CONCLUSIONS: Cetuximab has clinically significant activity when given alone or in combination with irinotecan in patients with irinotecan refractory colorectal cancer. PMID- 15269314 TI - Missense variations in the fibulin 5 gene and age-related macular degeneration. AB - BACKGROUND: Age-related macular degeneration (AMD) is the most common cause of irreversible vision loss in the developed world. The study of a rare mendelian form of macular degeneration implicated fibulin genes in the pathogenesis of more common forms of this disease. We evaluated five fibulin genes in a large series of patients with AMD. METHODS: We studied 402 patients with AMD and 429 control subjects from the same clinic population. Patients were examined by means of indirect ophthalmoscopy, slit-lamp microscopy, and fundus photography to establish the presence and phenotypic pattern of AMD. DNA samples were screened for sequence variations in five members of the fibulin gene family. RESULTS: Amino acid-altering sequence variations were found in all five fibulin genes, many of which were observed only in patients with AMD. Several of the altered residues have been conserved during evolution. Seven of the 402 patients with AMD had amino acid-altering sequence variations in the fibulin 5 gene, whereas none were observed among 429 control subjects (P<0.01). In addition, these seven patients all had small, circular drusen, which are commonly referred to as basal laminar or cuticular drusen. CONCLUSIONS: Missense mutations in the fibulin 5 gene were found in 1.7 percent of patients with AMD. Many variations in other fibulin genes were also found in these patients, and the evolutionary conservation of the affected residues suggests that several of these variations may also be involved in AMD. PMID- 15269315 TI - Methylprednisolone, valacyclovir, or the combination for vestibular neuritis. AB - BACKGROUND: Vestibular neuritis is the second most common cause of peripheral vestibular vertigo. Its assumed cause is a reactivation of herpes simplex virus type 1 infection. Therefore, corticosteroids, antiviral agents, or a combination of the two might improve the outcome in patients with vestibular neuritis. METHODS: We performed a prospective, randomized, double-blind, two-by-two factorial trial in which patients with acute vestibular neuritis were randomly assigned to treatment with placebo, methylprednisolone, valacyclovir, or methylprednisolone plus valacyclovir. Vestibular function was determined by caloric irrigation, with the use of the vestibular paresis formula (to measure the extent of unilateral caloric paresis) within 3 days after the onset of symptoms and 12 months afterward. RESULTS: Of a total of 141 patients who underwent randomization, 38 received placebo, 35 methylprednisolone, 33 valacyclovir, and 35 methylprednisolone plus valacyclovir. At the onset of symptoms there was no difference among the groups in the severity of vestibular paresis. The mean (+/-SD) improvement in peripheral vestibular function at the 12 month follow-up was 39.6+/-28.1 percentage points in the placebo group, 62.4+/ 16.9 percentage points in the methylprednisolone group, 36.0+/-26.7 percentage points in the valacyclovir group, and 59.2+/-24.1 percentage points in the methylprednisolone-plus-valacyclovir group. Analysis of variance showed a significant effect of methylprednisolone (P<0.001) but not of valacyclovir (P=0.43). The combination of methylprednisolone and valacyclovir was not superior to corticosteroid monotherapy. CONCLUSIONS: Methylprednisolone significantly improves the recovery of peripheral vestibular function in patients with vestibular neuritis, whereas valacyclovir does not. PMID- 15269316 TI - Acquired hypocalciuric hypercalcemia due to autoantibodies against the calcium sensing receptor. PMID- 15269317 TI - Flavivirus encephalitis. PMID- 15269318 TI - Images in clinical medicine. Intussusception of the small bowel. PMID- 15269319 TI - Case records of the Massachusetts General Hospital. Weekly clinicopathological exercises. Case 23-2004. A 50-year-old woman with low oxygen saturation. PMID- 15269320 TI - PEEP in ARDS--how much is enough? PMID- 15269321 TI - New treatment options for colorectal cancer. PMID- 15269322 TI - Death, destruction, and the proteasome. PMID- 15269323 TI - Free cortisol and critically ill patients. PMID- 15269324 TI - Trends in assisted reproductive technology. PMID- 15269325 TI - Ductal carcinoma in situ of the breast. PMID- 15269326 TI - Irradiation of food. PMID- 15269327 TI - Five years after the "Pocket Monster" seizures. PMID- 15269328 TI - Images in clinical medicine. A cystic mass in the liver. PMID- 15269330 TI - Hydrogen photoproduction is attenuated by disruption of an isoamylase gene in Chlamydomonas reinhardtii. AB - DNA insertional transformants of Chlamydomonas reinhardtii were screened chemochromically for attenuated H(2) production. One mutant, displaying low H(2) gas photoproduction, has a nonfunctional copy of a gene that shows high homology to the family of isoamylase genes found in several photosynthetic organisms. DNA gel blotting and gene complementation were used to link this isoamylase gene to previously characterized nontagged sta7 mutants. This mutant is therefore denoted sta7-10. In C. reinhardtii, the STA7 isoamylase gene is important for the accumulation of crystalline starch, and the sta7-10 mutant reported here contains <3% of the glucose found in insoluble starch when compared with wild-type control cells. Hydrogen photoproduction rates, induced after several hours of dark, anaerobic treatment, are attenuated in sta7 mutants. RNA gel blot analysis indicates that the mRNA transcripts for both the HydA1 and HydA2 [Fe]-hydrogenase genes are expressed in the sta7-10 mutant at greater than wild-type levels 0.5 h after anaerobic induction. However, after 1.5 h, transcript levels of both HydA1 and HydA2 begin to decline rapidly and reach nearly undetectable levels after 7 h. In wild-type cells, the hydrogenase transcripts accumulate more slowly, reach a plateau after 4 h of anaerobic treatment, and maintain the same level of expression for >7 h under anaerobic incubation. Complementation of mutant cells with genomic DNA corresponding to the STA7 gene restores both the starch accumulation and H(2) production phenotypes. The results indicate that STA7 and starch metabolism play an important role in C. reinhardtii H(2) photoproduction. Moreover, the results indicate that mere anaerobiosis is not sufficient to maintain hydrogenase gene expression without the underlying physiology, an important aspect of which is starch metabolism. PMID- 15269331 TI - Vascular associated death1, a novel GRAM domain-containing protein, is a regulator of cell death and defense responses in vascular tissues. AB - The hypersensitive response (HR) is a programmed cell death that is commonly associated with plant disease resistance. A novel lesion mimic mutant, vad1 (for vascular associated death1), that exhibits light conditional appearance of propagative HR-like lesions along the vascular system was identified. Lesion formation is associated with expression of defense genes, production of high levels of salicylic acid (SA), and increased resistance to virulent and avirulent strains of Pseudomonas syringae pv tomato. Analyses of the progeny from crosses between vad1 plants and either nahG transgenic plants, sid1, nonexpressor of PR1 (npr1), enhanced disease susceptibility1 (eds1), or non-race specific disease resistance1 (ndr1) mutants, revealed the vad1 cell death phenotype to be dependent on SA biosynthesis but NPR1 independent; in addition, both EDS1 and NDR1 are necessary for the proper timing and amplification of cell death as well as for increased resistance to Pseudomonas strains. VAD1 encodes a novel putative membrane-associated protein containing a GRAM domain, a lipid or protein binding signaling domain, and is expressed in response to pathogen infection at the vicinity of the hypersensitive lesions. VAD1 might thus represent a new potential function in cell death control associated with cells in the vicinity of vascular bundles. PMID- 15269332 TI - Genome-wide analysis of Arabidopsis pentatricopeptide repeat proteins reveals their essential role in organelle biogenesis. AB - The complete sequence of the Arabidopsis thaliana genome revealed thousands of previously unsuspected genes, many of which cannot be ascribed even putative functions. One of the largest and most enigmatic gene families discovered in this way is characterized by tandem arrays of pentatricopeptide repeats (PPRs). We describe a detailed bioinformatic analysis of 441 members of the Arabidopsis PPR family plus genomic and genetic data on the expression (microarray data), localization (green fluorescent protein and red fluorescent protein fusions), and general function (insertion mutants and RNA binding assays) of many family members. The basic picture that arises from these studies is that PPR proteins play constitutive, often essential roles in mitochondria and chloroplasts, probably via binding to organellar transcripts. These results confirm, but massively extend, the very sparse observations previously obtained from detailed characterization of individual mutants in other organisms. PMID- 15269333 TI - Increased glutathione biosynthesis plays a role in nickel tolerance in thlaspi nickel hyperaccumulators. AB - Worldwide more than 400 plant species are now known that hyperaccumulate various trace metals (Cd, Co, Cu, Mn, Ni, and Zn), metalloids (As) and nonmetals (Se) in their shoots. Of these, almost one-quarter are Brassicaceae family members, including numerous Thlaspi species that hyperaccumulate Ni up to 3% of there shoot dry weight. We observed that concentrations of glutathione, Cys, and O acetyl-l-serine (OAS), in shoot tissue, are strongly correlated with the ability to hyperaccumulate Ni in various Thlaspi hyperaccumulators collected from serpentine soils, including Thlaspi goesingense, T. oxyceras, and T. rosulare, and nonaccumulator relatives, including T. perfoliatum, T. arvense, and Arabidopsis thaliana. Further analysis of the Austrian Ni hyperaccumulator T. goesingense revealed that the high concentrations of OAS, Cys, and GSH observed in this hyperaccumulator coincide with constitutively high activity of both serine acetyltransferase (SAT) and glutathione reductase. SAT catalyzes the acetylation of l-Ser to produce OAS, which acts as both a key positive regulator of sulfur assimilation and forms the carbon skeleton for Cys biosynthesis. These changes in Cys and GSH metabolism also coincide with the ability of T. goesingense to both hyperaccumulate Ni and resist its damaging oxidative effects. Overproduction of T. goesingense SAT in the nonaccumulator Brassicaceae family member Arabidopsis was found to cause accumulation of OAS, Cys, and glutathione, mimicking the biochemical changes observed in the Ni hyperaccumulators. In these transgenic Arabidopsis, glutathione concentrations strongly correlate with increased resistance to both the growth inhibitory and oxidative stress induced effects of Ni. Taken together, such evidence supports our conclusion that elevated GSH concentrations, driven by constitutively elevated SAT activity, are involved in conferring tolerance to Ni-induced oxidative stress in Thlaspi Ni hyperaccumulators. PMID- 15269334 TI - Phosphoinositides in constitutive membrane traffic. AB - Proteins that make, consume, and bind to phosphoinositides are important for constitutive membrane traffic. Different phosphoinositides are concentrated in different parts of the central vacuolar pathway, with phosphatidylinositol 4 phosphate predominate on Golgi, phosphatidylinositol 4,5-bisphosphate predominate at the plasma membrane, phosphatidylinositol 3-phosphate the major phosphoinositide on early endosomes, and phosphatidylinositol 3,5-bisphosphate found on late endocytic organelles. This spatial segregation may be the mechanism by which the direction of membrane traffic is controlled. Phosphoinositides increase the affinity of membranes for peripheral membrane proteins that function for sorting protein cargo or for the docking and fusion of transport vesicles. This implies that constitutive membrane traffic may be regulated by the mechanisms that control the activity of the enzymes that produce and consume phosphoinositides. Although the lipid kinases and phosphatases that function in constitutive membrane traffic are beginning to be identified, their regulation is poorly understood. PMID- 15269335 TI - Nitric oxide in health and disease of the respiratory system. AB - During the past decade a plethora of studies have unravelled the multiple roles of nitric oxide (NO) in airway physiology and pathophysiology. In the respiratory tract, NO is produced by a wide variety of cell types and is generated via oxidation of l-arginine that is catalyzed by the enzyme NO synthase (NOS). NOS exists in three distinct isoforms: neuronal NOS (nNOS), inducible NOS (iNOS), and endothelial NOS (eNOS). NO derived from the constitutive isoforms of NOS (nNOS and eNOS) and other NO-adduct molecules (nitrosothiols) have been shown to be modulators of bronchomotor tone. On the other hand, NO derived from iNOS seems to be a proinflammatory mediator with immunomodulatory effects. The concentration of this molecule in exhaled air is abnormal in activated states of different inflammatory airway diseases, and its monitoring is potentially a major advance in the management of, e.g., asthma. Finally, the production of NO under oxidative stress conditions secondarily generates strong oxidizing agents (reactive nitrogen species) that may modulate the development of chronic inflammatory airway diseases and/or amplify the inflammatory response. The fundamental mechanisms driving the altered NO bioactivity under pathological conditions still need to be fully clarified, because their regulation provides a novel target in the prevention and treatment of chronic inflammatory diseases of the airways. PMID- 15269336 TI - Molecular regulation of vascular smooth muscle cell differentiation in development and disease. AB - The focus of this review is to provide an overview of the current state of knowledge of molecular mechanisms/processes that control differentiation of vascular smooth muscle cells (SMC) during normal development and maturation of the vasculature, as well as how these mechanisms/processes are altered in vascular injury or disease. A major challenge in understanding differentiation of the vascular SMC is that this cell can exhibit a wide range of different phenotypes at different stages of development, and even in adult organisms the cell is not terminally differentiated. Indeed, the SMC is capable of major changes in its phenotype in response to changes in local environmental cues including growth factors/inhibitors, mechanical influences, cell-cell and cell matrix interactions, and various inflammatory mediators. There has been much progress in recent years to identify mechanisms that control expression of the repertoire of genes that are specific or selective for the vascular SMC and required for its differentiated function. One of the most exciting recent discoveries was the identification of the serum response factor (SRF) coactivator gene myocardin that appears to be required for expression of many SMC differentiation marker genes, and for initial differentiation of SMC during development. However, it is critical to recognize that overall control of SMC differentiation/maturation, and regulation of its responses to changing environmental cues, is extremely complex and involves the cooperative interaction of many factors and signaling pathways that are just beginning to be understood. There is also relatively recent evidence that circulating stem cell populations can give rise to smooth muscle-like cells in association with vascular injury and atherosclerotic lesion development, although the exact role and properties of these cells remain to be clearly elucidated. The goal of this review is to summarize the current state of our knowledge in this area and to attempt to identify some of the key unresolved challenges and questions that require further study. PMID- 15269337 TI - Structure and function of Kv4-family transient potassium channels. AB - Shal-type (Kv4.x) K(+) channels are expressed in a variety of tissue, with particularly high levels in the brain and heart. These channels are the primary subunits that contribute to transient, voltage-dependent K(+) currents in the nervous system (A currents) and the heart (transient outward current). Recent studies have revealed an enormous degree of complexity in the regulation of these channels. In this review, we describe the surprisingly large number of ancillary subunits and scaffolding proteins that can interact with the primary subunits, resulting in alterations in channel trafficking and kinetic properties. Furthermore, we discuss posttranslational modification of Kv4.x channel function with an emphasis on the role of kinase modulation of these channels in regulating membrane properties. This concept is especially intriguing as Kv4.2 channels may integrate a variety of intracellular signaling cascades into a coordinated output that dynamically modulates membrane excitability. Finally, the pathophysiology that may arise from dysregulation of these channels is also reviewed. PMID- 15269338 TI - Molecular structure and physiological functions of GABA(B) receptors. AB - GABA(B) receptors are broadly expressed in the nervous system and have been implicated in a wide variety of neurological and psychiatric disorders. The cloning of the first GABA(B) receptor cDNAs in 1997 revived interest in these receptors and their potential as therapeutic targets. With the availability of molecular tools, rapid progress was made in our understanding of the GABA(B) system. This led to the surprising discovery that GABA(B) receptors need to assemble from distinct subunits to function and provided exciting new insights into the structure of G protein-coupled receptors (GPCRs) in general. As a consequence of this discovery, it is now widely accepted that GPCRs can exist as heterodimers. The cloning of GABA(B) receptors allowed some important questions in the field to be answered. It is now clear that molecular studies do not support the existence of pharmacologically distinct GABA(B) receptors, as predicted by work on native receptors. Advances were also made in clarifying the relationship between GABA(B) receptors and the receptors for gamma hydroxybutyrate, an emerging drug of abuse. There are now the first indications linking GABA(B) receptor polymorphisms to epilepsy. Significantly, the cloning of GABA(B) receptors enabled identification of the first allosteric GABA(B) receptor compounds, which is expected to broaden the spectrum of therapeutic applications. Here we review current concepts on the molecular composition and function of GABA(B) receptors and discuss ongoing drug-discovery efforts. PMID- 15269339 TI - Endothelial cell-to-cell junctions: molecular organization and role in vascular homeostasis. AB - Intercellular junctions mediate adhesion and communication between adjoining endothelial and epithelial cells. In the endothelium, junctional complexes comprise tight junctions, adherens junctions, and gap junctions. The expression and organization of these complexes depend on the type of vessels and the permeability requirements of perfused organs. Gap junctions are communication structures, which allow the passage of small molecular weight solutes between neighboring cells. Tight junctions serve the major functional purpose of providing a "barrier" and a "fence" within the membrane, by regulating paracellular permeability and maintaining cell polarity. Adherens junctions play an important role in contact inhibition of endothelial cell growth, paracellular permeability to circulating leukocytes and solutes. In addition, they are required for a correct organization of new vessels in angiogenesis. Extensive research in the past decade has identified several molecular components of the tight and adherens junctions, including integral membrane and intracellular proteins. These proteins interact both among themselves and with other molecules. Here, we review the individual molecules of junctions and their complex network of interactions. We also emphasize how the molecular architectures and interactions may represent a mechanistic basis for the function and regulation of junctions, focusing on junction assembly and permeability regulation. Finally, we analyze in vivo studies and highlight information that specifically relates to the role of junctions in vascular endothelial cells. PMID- 15269340 TI - Vascular actions of calcitonin gene-related peptide and adrenomedullin. AB - This review summarizes the receptor-mediated vascular activities of calcitonin gene-related peptide (CGRP) and the structurally related peptide adrenomedullin (AM). CGRP is a 37-amino acid neuropeptide, primarily released from sensory nerves, whilst AM is produced by stimulated vascular cells, and amylin is secreted from the pancreas. They share vasodilator activity, albeit to varying extents depending on species and tissue. In particular, CGRP has potent activity in the cerebral circulation, which is possibly relevant to the pathology of migraine, whilst vascular sources of AM contribute to dysfunction in cardiovascular disease. Both peptides exhibit potent activity in microvascular beds. All three peptides can act on a family of CGRP receptors that consist of calcitonin receptor-like receptor (CL) linked to one of three receptor activity modifying proteins (RAMPs) that are essential for functional activity. The association of CL with RAMP1 produces a CGRP receptor, with RAMP2 an AM receptor and with RAMP3 a CGRP/AM receptor. Evidence for the selective activity of the first nonpeptide CGRP antagonist BIBN4096BS for the CGRP receptor is presented. The cardiovascular activity of these peptides in a range of species and in human clinical conditions is detailed, and potential therapeutic applications based on use of antagonists and gene targeting of agonists are discussed. PMID- 15269341 TI - Urinary bladder contraction and relaxation: physiology and pathophysiology. AB - The detrusor smooth muscle is the main muscle component of the urinary bladder wall. Its ability to contract over a large length interval and to relax determines the bladder function during filling and micturition. These processes are regulated by several external nervous and hormonal control systems, and the detrusor contains multiple receptors and signaling pathways. Functional changes of the detrusor can be found in several clinically important conditions, e.g., lower urinary tract symptoms (LUTS) and bladder outlet obstruction. The aim of this review is to summarize and synthesize basic information and recent advances in the understanding of the properties of the detrusor smooth muscle, its contractile system, cellular signaling, membrane properties, and cellular receptors. Alterations in these systems in pathological conditions of the bladder wall are described, and some areas for future research are suggested. PMID- 15269342 TI - Molecular and cellular physiology of renal organic cation and anion transport. AB - Organic cations and anions (OCs and OAs, respectively) constitute an extraordinarily diverse array of compounds of physiological, pharmacological, and toxicological importance. Renal secretion of these compounds, which occurs principally along the proximal portion of the nephron, plays a critical role in regulating their plasma concentrations and in clearing the body of potentially toxic xenobiotics agents. The transepithelial transport involves separate entry and exit steps at the basolateral and luminal aspects of renal tubular cells. It is increasingly apparent that basolateral and luminal OC and OA transport reflects the concerted activity of a suite of separate transport processes arranged in parallel in each pole of proximal tubule cells. The cloning of multiple members of several distinct transport families, the subsequent characterization of their activity, and their subcellular localization within distinct regions of the kidney now allows the development of models describing the molecular basis of the renal secretion of OCs and OAs. This review examines recent work on this issue, with particular emphasis on attempts to integrate information concerning the activity of cloned transporters in heterologous expression systems to that observed in studies of physiologically intact renal systems. PMID- 15269343 TI - A novel signaling mechanism in brain development. PMID- 15269344 TI - CCAAT displacement protein/cut homolog recruits G9a histone lysine methyltransferase to repress transcription. AB - CCAAT displacement protein/cut homolog (CDP/cut) is a highly conserved homeodomain protein that contains three cut repeat sequences. CDP/cut is a transcriptional factor for many diverse cellular and viral genes that are involved in most cellular processes, including differentiation, development, and proliferation. Here, we report that CDP/cut interacts with a histone lysine methyltransferase (HKMT), G9a, in vivo and in vitro. The deletion of the cut repeats within CDP/cut abrogates the interaction with G9a. The transcriptional repressor function of CDP/cut is mediated through HKMT activity of G9a associated with CDP/cut. We show that the recruitment of G9a to the human p21(waf1/cdi1) promoter is contingent on the interaction with CDP/cut, and CDP/cut is directly associated with an increase in the methylation in vivo of Lys-9 in histone H3 within the CDP/cut-regulatory region of the p21(waf1/cdi1) promoter. The endogenous level of p21(waf1/cdi1) expression is repressed through CDP/cut and mediated by HKMT activity of G9a. Furthermore, we report the identification of G9a as a component of CDP/cut complex. G9a colocalizes with CDP/cut in the nucleus. These results indicate that G9a functions as a transcriptional corepressor in association with a CDP/cut complex. These studies now reveal the interaction of G9a with a sequence-specific transcription factor that regulates gene repression through CDP/cut. PMID- 15269345 TI - Anchor residues in protein-protein interactions. AB - We show that the mechanism for molecular recognition requires one of the interacting proteins, usually the smaller of the two, to anchor a specific side chain in a structurally constrained binding groove of the other protein, providing a steric constraint that helps to stabilize a native-like bound intermediate. We identify the anchor residues in 39 protein-protein complexes and verify that, even in the absence of their interacting partners, the anchor side chains are found in conformations similar to those observed in the bound complex. These ready-made recognition motifs correspond to surface side chains that bury the largest solvent-accessible surface area after forming the complex (> or =100 A2). The existence of such anchors implies that binding pathways can avoid kinetically costly structural rearrangements at the core of the binding interface, allowing for a relatively smooth recognition process. Once anchors are docked, an induced fit process further contributes to forming the final high affinity complex. This later stage involves flexible (solvent-exposed) side chains that latch to the encounter complex in the periphery of the binding pocket. Our results suggest that the evolutionary conservation of anchor side chains applies to the actual structure that these residues assume before the encounter complex and not just to their loci. Implications for protein docking are also discussed. PMID- 15269346 TI - Molecular structure of double-minute chromosomes bearing amplified copies of the epidermal growth factor receptor gene in gliomas. AB - Amplification of the epidermal growth factor receptor gene on double minutes is recurrently observed in cells of advanced gliomas, but the structure of these extrachromosomal circular DNA molecules and the mechanisms responsible for their formation are still poorly understood. By using quantitative PCR and chromosome walking, we investigated the genetic content and the organization of the repeats in the double minutes of seven gliomas. It was established that all of the amplicons of a given tumor derive from a single founding extrachromosomal DNA molecule. In each of these gliomas, the founding molecule was generated by a simple event that circularizes a chromosome fragment overlapping the epidermal growth factor receptor gene. In all cases, the fusion of the two ends of this initial amplicon resulted from microhomology-based nonhomologous end-joining. Furthermore, the corresponding chromosomal loci were not rearranged, which strongly suggests that a postreplicative event was responsible for the formation of each of these initial amplicons. PMID- 15269347 TI - A specific gene expression program triggered by Gram-positive bacteria in the cytosol. AB - Innate and adaptive immunity depends critically on host recognition of pathogen associated molecules. Toll-like receptors (TLRs) are key mediators of pathogen surveillance at the cell or phagocytic vacuole surface. However, mechanisms underlying recognition of pathogens in other cellular compartments remain unclear, and responses elicited by cytosolic challenge are poorly characterized. We therefore used mouse cDNA microarrays to investigate gene expression triggered by infection of bone marrow-derived macrophages with cytosol- and vacuole localized Listeria monocytogenes (Lm), a model cytosolic pathogen. The resulting gene expression program included two basic categories of induced genes: an "early/persistent" cluster consistent with NF-kappaB-dependent responses downstream of TLRs, and a subsequent "late response" cluster largely composed of IFN-responsive genes (IRGs). The early/persistent cluster was observed upon infection with WT, heat-killed, or mutant Lm lacking listeriolysin O, the pore forming hemolysin that promotes escape from phagocytic vacuoles. However, the IRG cluster depended on entry of WT Lm into the cytosol. Infection with listeriolysin O-expressing, cytosolic Bacillus subtilis (Bs) strikingly recapitulated the expression profile associated with WT Lm, including IRG induction. IRG up regulation was associated with MyD88-independent induction of IFN-beta transcription and activity. Whereas Staphylococcus aureus (Sa) lipoteichoic acid treatment confirmed that many late-response genes could also be stimulated through TLRs, our study identified a cytosol-specific transcriptional program independent of TLR signaling through MyD88. Further characterization of cytosolic surveillance pathway(s) and their points of convergence with TLR- and IFN dependent pathways will enhance our understanding of the means by which mammals detect and respond to pathogens. PMID- 15269348 TI - Mpl Baltimore: a thrombopoietin receptor polymorphism associated with thrombocytosis. AB - The chronic myeloproliferative disorders (MPD) are clonal hematopoietic stem cell disorders of unknown etiology. We have reported defective thrombopoietin receptor (Mpl) protein expression in MPD patients. To determine whether the basis of abnormal Mpl protein expression was due to mutations in the Mpl gene, we sequenced Mpl cDNA from MPD patients. We found a single nucleotide substitution (G1238T) that results in a change from lysine to asparagine at amino acid 39 (K39N) in three African-American women referred for an evaluation of an MPD. We subsequently screened more than 400 patients and controls and found that the K39N substitution is a polymorphism restricted to African Americans and that approximately 7% of African Americans are heterozygous for K39N. African Americans with the K39N polymorphism had a significantly higher platelet count than controls without the polymorphism (P < 0.001) and reduced platelet protein Mpl expression. Expression of an Mpl cDNA containing the K39N substitution in cell lines was associated with incomplete processing and a reduction in Mpl protein, recapitulating the Mpl protein defect observed in platelets from individuals with K39N. K39N represents an identified functional Mpl polymorphism and is associated with altered protein expression of Mpl and a clinical phenotype of thrombocytosis. PMID- 15269350 TI - Negative regulation of herpes simplex virus type 1 ICP4 promoter by IE180 protein of pseudorabies virus. AB - Recombinant pseudorabies viruses (PRVs) gIS8 and N1aHTK were constructed by the insertion of a chimeric gene (alpha4-TK) from herpes simplex virus type 1 (HSV-1) into wild-type PRV. HSV-1 TK expression by these recombinant viruses resulted in enhanced sensitivity to ganciclovir, compared to that of the wild-type PRV, and was similar to the sensitivity shown by HSV-1. Infection with gIS8 or N1aHTK recombinant viruses led to expression of HSV-1 TK mRNA as an immediate-early (IE) gene, observed by downregulation of the HSV-1 alpha4 promoter. This negative regulation was due to a PRV IE protein, IE180. IE180, however, does not have all the regulatory functions of the infected-cell protein ICP4, as it does not restore the growth of ICP4-deficient HSV-1 mutants. PMID- 15269349 TI - Beta-adrenergic modulation of emotional memory-evoked human amygdala and hippocampal responses. AB - Human emotional experience is typically associated with enhanced episodic memory. We have used functional magnetic resonance imaging to demonstrate that successful encoding of emotional, compared to neutral, verbal stimuli evokes increased human amygdala responses. Items that evoke amygdala activation at encoding evoke greater hippocampal responses at retrieval compared to neutral items. Administration of the beta-adrenergic antagonist propranolol at encoding abolishes the enhanced amygdala encoding and hippocampal retrieval effects, despite propranolol being no longer present at retrieval. Thus, memory-related amygdala responses at encoding and hippocampal responses at recognition for emotional items depend on beta-adrenergic engagement at encoding. Our results suggest that human emotional memory is associated with a beta-adrenergic dependent modulation of amygdala-hippocampal interactions. PMID- 15269351 TI - A synthetic peptide from a heptad repeat region of herpesvirus glycoprotein B inhibits virus replication. AB - Glycoprotein B (gB) is the most conserved glycoprotein of herpesviruses and plays important roles in virus infectivity. Two intervening heptad repeat (HR) sequences were found in the C-terminal half of all herpesvirus gBs analysed. A synthetic peptide derived from the HR region (aa 477-510) of bovine herpesvirus type 1 (BoHV-1) gB was studied for its ability to inhibit virus replication. The peptide interfered with cell-to-cell spread and consistently inhibited replication of BoHV-1, with a 50 % effective concentration value (EC(50)) of 5 microM. Inhibition of replication was obtained not only with herpesviruses including pseudorabies virus and herpes simplex virus type 1 but also partly with Newcastle disease virus. Possible mechanisms of membrane fusion inhibition by the peptide are discussed. PMID- 15269352 TI - CD4(+) T-cell responses to herpes simplex virus type 2 (HSV-2) glycoprotein G are type specific and differ in symptomatic and asymptomatic HSV-2-infected individuals. AB - T-cell recognition of the secreted and membrane-bound portions of the herpes simplex virus type 2 (HSV-2) glycoprotein G (sgG-2 and mgG-2, respectively) was compared in symptomatic and asymptomatic HSV-2-infected individuals and in HSV-2 seronegative controls and the responses with HSV-1 glycoproteins C and E (gC-1 and gE-1) were compared. CD4(+) T cells from HSV-2-infected individuals specifically recognized both sgG-2 and mgG-2, whereas HSV-1-infected and HSV seronegative controls did not respond to these glycoproteins. The responses to gC 1 and gE-1, on the other hand, were not type specific, as blood mononuclear cells from both HSV-1- and HSV-2-infected individuals responded in vitro. There was an association between the status of the infection (symptomatic versus asymptomatic) and the CD4(+) T-cell responsiveness. Symptomatic HSV-2-seropositive individuals responded with significantly lower Th1 cytokine production to sgG-2 and mgG-2 than did asymptomatic HSV-2-infected carriers, especially within the HSV-1 negative cohort. No differences in T-cell proliferation were observed between asymptomatic and symptomatic individuals. The results have implications for studies of HSV-2-specific CD4(+) T-cell reactivity in general and for analysis of immunological differences between asymptomatic and symptomatic individuals in particular. PMID- 15269353 TI - Sumoylation of the major immediate-early IE2 protein of human cytomegalovirus Towne strain is not required for virus growth in cultured human fibroblasts. AB - Sumoylation of the major immediate-early IE2 protein of human cytomegalovirus has been shown to increase transactivation activity in target reporter gene assays. This study examined the role of IE2 sumoylation in viral infection. A Towne strain-based bacterial artificial chromosome clone was generated encoding a mutated form of the IE2 protein with Lys-->Arg substitutions at positions 175 and 180, the two major sumoylation sites. When human fibroblast (HF) cells were infected with the reconstituted mutant virus, (i) viral growth kinetics, (ii) the accumulation of IE1 (UL123), IE2 (UL122), p52 (UL44) and pp65 (UL83) proteins and (iii) the relocalization of the cellular small ubiquitin-like modifier (SUMO)-1, p53 and proliferating cell nuclear antigen proteins into viral DNA replication compartments were comparable with those of the wild-type and the revertant virus. The data demonstrate that sumoylation of IE2 is not essential for virus growth in cultured HF cells. PMID- 15269354 TI - Functional co-operation between the Kaposi's sarcoma-associated herpesvirus ORF57 and ORF50 regulatory proteins. AB - Kaposi's sarcoma (KS)-associated herpesvirus (KSHV) proteins ORF57 (also known as MTA) and ORF50 (also known as RTA) act post-transcriptionally and transcriptionally to regulate viral lytic gene expression and synergistically activate certain early and late KSHV promoters. When ORF57 and ORF50 were co expressed, they co-operatively stimulated expression from the promoter of the immediate-early ORF50 gene itself. Co-immunoprecipitations with extracts of KSHV infected cells showed that ORF57 and ORF50 proteins were present in the same complex. Using the pull-down assay with extracts of KSHV-infected cells, ORF50 protein was shown to interact with a glutathione S-transferase-ORF57 fusion protein. A chromatin immunoprecipitation assay showed that ORF50 promoter sequences were preferentially associated with immunoprecipitated chromatin using both anti-ORF50 and anti-ORF57 antibodies consistent with both an in vivo physical association between ORF57 and ORF50 and a potential role for ORF57 at the transcriptional level. This is the first demonstration of an interaction between these two lytic regulatory proteins in a gammaherpesvirus. Expression of ORF50 protein is sufficient to induce lytic replication in latently infected cells and may determine viral host range, spread and KS pathogenesis in vivo. A new insight into the co-ordinated activities of these two key regulatory proteins is provided in which upregulation of the ORF50 promoter with augmentation of ORF50 activity by ORF57 protein, and vice versa, would facilitate the cascade of lytic viral gene expression, thereby breaking latency. A functional and physical interaction between these two gammaherpesvirus regulatory protein counterparts could be a general feature of the herpesviruses. PMID- 15269355 TI - Modified vaccinia virus Ankara induces moderate activation of human dendritic cells. AB - Modified vaccinia virus Ankara (MVA) is a highly attenuated strain known to be an effective vaccine vector. Here it is demonstrated that MVA, unlike standard vaccinia virus (VACV) strains, activates monocyte-derived human dendritic cells (DCs) as testified by an increase in surface co-stimulatory molecules and the secretion of pro-inflammatory cytokines. Inhibition of virus gene expression by subjecting MVA to UV light or heat treatment did not alter its ability to activate DCs. On the other hand, standard VACV strains activated DCs if virus gene expression was prevented by prior UV light or heat treatment. These results suggest that MVA or standard VACV particles are responsible for DC activation but, in the case of standard VACV strains, virus gene expression prevents activation. Additional experiments showed that DCs were activated by MVA-infected HeLa cells and, under these conditions, could induce secretion of gamma interferon from T lymphocytes more efficiently than if a replication-competent VACV strain was employed. These data provide one explanation for the remarkable immune-stimulating capacity of MVA in the absence of virus multiplication. PMID- 15269356 TI - Characterization of pathogenic and non-pathogenic African swine fever virus isolates from Ornithodoros erraticus inhabiting pig premises in Portugal. AB - Ten African swine fever virus isolates from the soft tick Ornithodoros erraticus collected on three farms in the province of Alentejo in Portugal were characterized by their ability to cause haemadsorption (HAD) of red blood cells to infected pig macrophages, using restriction enzyme site mapping of the virus genomes and by experimental infection of pigs. Six virus isolates induced haemadsorption and four were non-haemadsorbing (non-HAD) in pig macrophage cell cultures. The restriction enzyme site maps of two non-HAD viruses, when compared with a virulent HAD isolate, showed a deletion of 9.6 kbp in the fragment adjacent to the left terminal fragment and of 1.6 kbp in the right terminal fragment and an insertion of 0.2 kbp in the central region. The six HAD viruses isolated were pathogenic and produced typical acute African swine fever in pigs and the four non-HAD isolates were non-pathogenic. Pigs that were infected with non-HAD viruses were fully resistant or had a delay of up to 14 days in the onset of disease, after challenge with pathogenic Portuguese viruses. Non-HAD viruses could be transmitted by contact but with a lower efficiency (42-50 %) compared with HAD viruses (100 %). The clinical differences found between the virus isolates from the ticks could have implications for the long-term persistence of virus in the field because of the cross-protection produced by the non-pathogenic isolates. This may also explain the presence of seropositive pigs in herds in Alentejo where no clinical disease had been reported. PMID- 15269357 TI - Human papillomavirus genotypes in cervical cancers in Mozambique. AB - The distribution of human papillomavirus (HPV) types in cervical cancers is essential for design and evaluation of HPV type-specific vaccines. To follow up on a previous report that HPV types 35 and 58 were the dominant HPV types in cervical neoplasia in Mozambique, the HPV types in a consecutive case series of 74 invasive cervical cancers in Mozambique were determined. The most common worldwide major oncogenic HPV types 16 and 18 were present in 69 % of cervical cancers, suggesting that a vaccine targeting HPV-16 and -18 would have a substantial impact on cervical cancer also in Mozambique. PMID- 15269358 TI - Broad-spectrum detection of papillomaviruses in bovine teat papillomas and healthy teat skin. AB - To investigate the prevalence of bovine papillomavirus (BPV) in bovine papilloma and healthy skin, DNA extracted from teat papillomas and healthy teat skin swabs was analysed by PCR using the primer pairs FAP59/FAP64 and MY09/MY11. Papillomavirus (PV) DNA was detected in all 15 papilloma specimens using FAP59/FAP64 and in 8 of the 15 papilloma specimens using MY09/MY11. In swab samples, 21 and 8 of the 122 samples were PV DNA positive using FAP59/FAP64 and MY09/MY11, respectively. Four BPV types (BPV-1, -3, -5 and -6), two previously identified putative BPV types (BAA1 and -5) and 11 putative new PV types (designated BAPV1 to -10 and BAPV11MY) were found in the 39 PV DNA-positive samples. Amino acid sequence alignments of the putative new PV types with reported BPVs and phylogenetic analyses of the putative new PV types with human and animal PV types showed that BAPV1 to -10 and BAPV11MY are putative new BPV types. These results also showed the genomic diversity and extent of subclinical infection of BPV. PMID- 15269359 TI - Attachment of bovine parvovirus to sialic acids on bovine cell membranes. AB - Although it has previously been shown that bovine parvovirus (BPV) attaches to the sialated glycoprotein glycophorin A on erythrocytes, the nature of virus binding moieties on mammalian nucleated cells is less clear. Buffalo lung fibroblasts (Bu), primary bovine embryonic kidney cells, Madin-Darby bovine kidney cells and bovine embryonic trachea (EBTr) cells were assessed for molecules capable of binding BPV. Competition studies were carried out on both erythrocyte and nucleated cell targets using a variety of sialated compounds and sialic acid-negative compounds. Glycophorin A was found to inhibit BPV binding, while mucin exhibited low-level inhibition. These two sialated compounds also blocked attachment of BPV-modified microsphere carriers to the Bu cell membrane. Influenza A virus was used as a sialic acid competitor and interfered with BPV attachment to erythrocytes and replication in Bu cells. Significantly, the enzyme sialidase removed BPV-binding sites from Bu and EBTr cells. The binding sites could be reconstituted on sialidase-treated cells by the enzymes alpha-2,3-O sialyltransferase and alpha-2,3-N-sialyltransferase. These results indicated that BPV can attach to sialic acid on cell membranes and that the sialylglycoproteins available for virus attachment appear to contain both N- and O-linked carbohydrate moieties, but that not all members of the sialic acid family can bind BPV. Moreover, there may be other moieties that can bind BPV, which may act as either primary or secondary receptors. PMID- 15269360 TI - The adenovirus E1A and E1B19K genes provide a helper function for transfection based adeno-associated virus vector production. AB - Although the adenoviral E1, E2A, E4 and VA RNA regions are required for efficient adeno-associated virus (AAV) vector production, the role that the individual E1 genes (E1A, E1B19K, E1B55K and protein IX) play in AAV vector production has not been clearly determined. E1 mutants were analysed for their ability to mediate AAV vector production in HeLa or KB cells, when cotransfected with plasmids encoding all other packaging functions. Disruption of E1A and E1B19K genes resulted in vector yield reduction by up to 10- and 100-fold, respectively, relative to the wild-type E1. Interruption of the E1B55K and protein IX genes had a modest effect on vector production. Interestingly, expression of anti-apoptotic E1B19K cellular homologues such as Bcl-2 or Bcl-x(L) fully complemented E1B19K mutants for AAV vector production. These findings may be valuable for the future development of packaging cell lines for AAV vector production. PMID- 15269361 TI - Phylogenetic evidence of widespread distribution of genotype 3 JC virus in Africa and identification of a type 7 isolate in an African AIDS patient. AB - JC virus (JCV) is the cause of progressive multifocal leukoencephalophathy (PML) in immunocompromised patients. The paucity of reports from Africa has led to the hypothesis that PML is rare because of an absence of virus genotypes associated with the condition. Genotypes 3 and 6 have been identified in East and West Africa but the distribution of types across the rest of Africa is unknown. Full length sequences of five JCV cerebrospinal fluid samples from PML patients in South Africa are reported here. Three isolates from African AIDS patients grouped with type 3A or 3B, and one with type 7, while one from a Caucasian leukaemia patient grouped with type 2D. Widespread distribution of type 3 on the continent may reflect migration patterns in antiquity, but this is the first report of type 7 in an African individual. Type 2D has only been isolated previously in South Asia, although transmission of this genotype to Europeans who later settled in South Africa is not unlikely. PMID- 15269362 TI - VP1 of infectious bursal disease virus is an RNA-dependent RNA polymerase. AB - Segment B of the bisegmented, double-stranded RNA genome of infectious bursal disease virus (IBDV) encodes the viral protein VP1. This has been presumed to represent the RNA-dependent RNA polymerase (RdRp) as it contains motifs that are typical for the RdRp of plus-strand RNA viruses. Here it is demonstrated that baculovirus-expressed wild-type but not motif A mutated VP1 acts as an RdRp on IBDV-specific RNA templates. Thus, on a plus-strand IBDV segment A cRNA template, minus-strand synthesis occurred in such a way that a covalently linked double stranded RNA product was generated (by a 'copy-back' mechanism). Importantly, enzyme activity was observed only with templates that comprised the 3' non-coding region of plus-strand RNAs transcribed from IBDV segments A and B, indicating template specificity. RdRp activity was shown to have a temperature optimum of 37 degrees C and required magnesium ions for enzyme activity. Thus, it has been demonstrated unequivocally that VP1 represents the RdRp of IBDV. PMID- 15269363 TI - Peptides resulting from the pVP2 C-terminal processing are present in infectious pancreatic necrosis virus particles. AB - The capsid of birnaviruses contains two proteins, VP2 and VP3, which derive from the processing of a large polyprotein, NH(2)-pVP2-VP4-VP3-COOH. The proteolytic cascade involved in processing the polyprotein, and in the final maturation of pVP2 (the precursor of VP2), has recently been shown to generate VP2 and four structural peptides in infectious bursal disease virus and blotched snakehead virus. The presence of peptides in infectious pancreatic necrosis virus particles was investigated using mass spectrometry and N-terminal sequencing of virus particles. Three peptides deriving from the C terminus of pVP2 (residues 443-486, 487-495 and 496-508 of the polyprotein) and 14 additional peptides produced by further processing of peptides [443-486] and [496-508] were identified. These results indicate that the presence of several virus-encoded peptides in the virions is a hallmark of birnaviruses. PMID- 15269364 TI - Termination and read-through proteins encoded by genome segment 9 of Colorado tick fever virus. AB - Genome segment 9 (Seg-9) of Colorado tick fever virus (CTFV) is 1884 bp long and contains a large open reading frame (ORF; 1845 nt in length overall), although a single in-frame stop codon (at nt 1052-1054) reduces the ORF coding capacity by approximately 40 %. However, analyses of highly conserved RNA sequences in the vicinity of the stop codon indicate that it belongs to a class of 'leaky terminators'. The third nucleotide positions in codons situated both before and after the stop codon, shows the highest variability, suggesting that both regions are translated during virus replication. This also suggests that the stop signal is functionally leaky, allowing read-through translation to occur. Indeed, both the truncated 'termination' protein and the full-length 'read-through' protein (VP9 and VP9', respectively) were detected in CTFV-infected cells, in cells transfected with a plasmid expressing only Seg-9 protein products, and in the in vitro translation products from undenatured Seg-9 ssRNA. The ratios of full length and truncated proteins generated suggest that read-through may be down regulated by other viral proteins. Western blot analysis of infected cells and purified CTFV showed that VP9 is a structural component of the virion, while VP9' is a non-structural protein. PMID- 15269365 TI - A comparison of the effects of oral inoculation with Rotashield and pentavalent reassortant rotavirus vaccine (WC3-PV) on suckling CB17scid mice. AB - The effects of oral inoculation into infant CB17(scid) mice of two reassortant rotavirus vaccines were compared. The vaccines were Rotashield and WC3-PV, a mixture of five reassortants (G1, G2, G3, G4 and P1; pentavalent reassortant vaccine). Control mice were inoculated with a placebo. At 6 days post-inoculation (p.i.), 8 of 13 (62 %; P<0.005) Rotashield-inoculated mice developed hepatitis and/or bile-duct obstruction compared with none of 11 mice given WC3-PV and none of 14 given placebo. In the Rotashield-inoculated mice, only serotype G3 rhesus rotavirus (RRV) was isolated from multiple sites, including intestine, liver, pancreas, spleen, blood and mesenteric lymph nodes. Recovery of RRV from Rotashield-inoculated mice followed a biphasic pattern. The two peaks of RRV recovery appeared to coincide firstly with replication in the intestine during days 1-3 p.i., and secondly with virus infection of the liver from days 10 to 15 p.i. WC3 reassortants of four different serotypes were detected only at day 1 p.i. in the intestine, liver, pancreas and blood cells from three WC3-PV inoculated mouse pups. However, WC3-PV did not produce any hepatopathology. Rotashield and WC3-PV appeared to exhibit different biological activity in infant CB17(scid) mouse pups. PMID- 15269366 TI - Potent virucidal activity in larval Heliothis virescens plasma against Helicoverpa zea single capsid nucleopolyhedrovirus. AB - Lepidopteran larvae resist baculovirus infection by selective apoptosis of infected midgut epithelial cells and by sloughing off infected cells from the midgut. Once the infection breaches the midgut epithelial barrier and propagates from infective foci to the haemocoel, however, there are few mechanisms known to account for the resistance and clearance of infection observed in some virus-host combinations. The hypothesis that factors present in the plasma of infected pest larvae act to limit the spread of virus from initial infective foci within the haemocoel was tested. An in vitro bioassay was developed in which Helicoverpa zea single capsid nucleopolyhedrovirus (HzSNPV) was incubated with plasma collected from uninfected Heliothis virescens larvae. Infectious HzSNPV particles were then titrated on HzAM1 cells. Diluted plasma from larval Heliothis virescens exhibited a virucidal effect against HzSNPV in vitro, reducing the TCID(50) ml(-1) by more than 64-fold (from 4.3+/-3.6x10(5) to 6.7+/-0.6x10(3)). The antiviral activity was heat-labile but was unaffected by freezing. In addition, protease inhibitors and specific chemical inhibitors of phenol oxidase or prophenol oxidase activation added to diluted plasma eliminated the virucidal activity. Thus, in the plasma of larval lepidopterans, the enzyme phenol oxidase may act as a constitutive, humoral innate antiviral immune response. PMID- 15269367 TI - Complete nucleotide sequence of Kashmir bee virus and comparison with acute bee paralysis virus. AB - The complete nucleotide sequence of a novel virus is presented here together with serological evidence that it belongs to Kashmir bee virus (KBV). Analysis reveals that KBV is a cricket paralysis-like virus (family Dicistroviridae: genus Cripavirus), with a non-structural polyprotein open reading frame in the 5' portion of the genome separated by an intergenic region from a structural polyprotein open reading frame in the 3' part of the genome. The genome also has a polyadenylated tail at the 3' terminus. KBV is one of several related viruses that also includes acute bee paralysis virus (ABPV). Although KBV and ABPV are about 70 % identical over the entire genome, there are considerable differences between them in significant areas of the genome, such as the 5' non-translated region (42 % nucleotide identity), between the helicase and 3C-protease domains of the non-structural polyprotein (57 % amino acid identity) and in a 90 aa stretch of the structural polyprotein (33 % amino acid identity). Phylogenetic analyses show that KBV and ABPV isolates fall into clearly separated clades with moderate evolutionary distance between them. Whether these genomic and evolutionary differences are sufficient to classify KBV and ABPV as separate species remains to be determined. PMID- 15269368 TI - Sequence analysis of human rhinoviruses in the RNA-dependent RNA polymerase coding region reveals large within-species variation. AB - Human rhinoviruses (HRVs; family Picornaviridae), the most frequent causative agents of respiratory infections, comprise more than 100 distinct serotypes. According to previous phylogenetic analysis of the VP4/VP2-coding sequences, all but one of the HRV prototype strains distribute between the two established species, Human rhinovirus A (HRV-A) and Human rhinovirus B (HRV-B). Here, partial sequences of the RNA-dependent RNA polymerase (3D polymerase)-coding gene of 48 HRV prototype strains and 12 field isolates were analysed. The designated division of the HRV strains into the species HRV-A and HRV-B was also seen in the 3D-coding region. Phylogenetically, HRV-B clustered closer to human enterovirus (HEV) species HEV-B, HEV-C and poliovirus than to HRV-A. Intraspecies variation within both HRV-A and HRV-B was greater in the 3D-coding region than in the VP4/VP2-coding region, with the difference maxima reaching 48 % at the nucleotide level and 36 % at the amino acid level in HRV-A and 53 and 35 %, respectively, in HRV-B. Within both species, a few strains formed a separate cluster differing from the majority of strains as much as HEV-B from HEV-C. Furthermore, the tree topology within HRV-A differed from that for VP4/VP2, suggesting possible recombination events in the evolutionary history of the strains. However, all 12 field isolates clustered similarly, as in the capsid region. These results showed that the within-species variation in the 3D region is greater in HRV than in HEV. Furthermore, HRV variation in the 3D region exceeds that in the capsid-coding region. PMID- 15269369 TI - All five cold-shock domains of unr (upstream of N-ras) are required for stimulation of human rhinovirus RNA translation. AB - Efficient translation of human rhinovirus-2 (HRV-2) RNA from its internal ribosome entry site (IRES) depends on the presence of cellular trans-acting factors upstream of N-ras (unr) and polypyrimidine-tract-binding protein. unr contains five cold-shock domains (CSDs) and is predicted to act as an RNA chaperone, allowing the HRV-2 IRES to attain the correct conformation for ribosome binding. To investigate the role of each of the CSDs in IRES-dependent translation, five unr mutants, each harbouring a point mutation in a different CSD, were generated. All five mutants were severely impaired in their ability to bind to the IRES and to stimulate translation from it. This showed that the ability of unr to function as an activator of HRV-2 RNA translation requires the RNA-binding activity of all five CSDs. PMID- 15269370 TI - Expansion of host-cell tropism of foot-and-mouth disease virus despite replication in a constant environment. AB - Foot-and-mouth disease virus (FMDV) variants adapted to BHK-21 cells showed an expanded host-cell tropism that extended to primate and human cell lines. Virus replication in human HeLa and Jurkat cells has been documented by titration of virus infectivity, quantification of virus RNA, expression of a virus-specific non-structural antigen, and serial passage of virus in the cells. Parallel serial infections of human Jurkat cells with the same variant FMDVs indicates a strong stochastic component in the progression of infection. Chimeric viruses identified the capsid as a genomic region involved in tropism expansion. These results indicate that, contrary to theoretical predictions, replication of an RNA virus in a constant cellular environment may lead to expansion of cellular tropism, rather than to a more specialized infection of the cellular type to which the virus has been adapted. PMID- 15269371 TI - Functional properties of a 16 kDa protein translated from an alternative open reading frame of the core-encoding genomic region of hepatitis C virus. AB - Hepatitis C virus (HCV) often causes persistent infection in humans. This could be due in part to the effect of viral proteins on cellular gene expression. Earlier observations suggest that the HCV core protein expressed from genotype 1a modulates important cellular genes at the transcriptional level, affects programmed cell death (apoptosis) and promotes cell growth. Recently, different groups of investigators have reported the translation of an approximately 16 kDa protein (named F/ARFP/core+1 ORF) from an alternate open reading frame of the HCV core-encoding genomic region. The functional significance of this F protein is presently unknown. Thus, whether the F and core proteins have both shared and distinct functions was investigated here. The experimental observations suggested that the F protein does not significantly modulate c-myc, hTERT and p53 promoter activities, unlike the HCV core protein. Interestingly, the F protein repressed p21 expression. Further studies indicated that the F protein does not inhibit tumour necrosis factor alpha-mediated apoptosis of HepG2 cells or promote rat embryo fibroblast growth. Taken together, these results suggest that the F protein does not share major properties identified previously for the HCV core protein, other than regulating p21 expression. PMID- 15269372 TI - Identification of the homotypic interaction domain of the core protein of dengue virus type 2. AB - Dengue virus causes dengue haemorrhagic fever or dengue shock syndrome with a high mortality rate. The genome of dengue virus is a positive-sense, single stranded RNA encoding three structural and seven non-structural proteins. The core protein is one of the three structural proteins and is the building block of the nucleocapsid of dengue virus. The core protein of dengue virus type 2 (DEN2) is composed of 100 aa with four alpha-helix domains. An internal hydrophobic domain located at aa 44-60 was identified. The DEN2 core protein was shown to form homodimers. Deletion of aa 1-36 or 73-100 decreased but did not completely abolish the core-to-core homotypic interaction, whereas deletion of a portion (aa 44-60) within aa 37-72 completely abolished the ability of the DEN2 core proteins to interact with each other. A recombinant DEN2 core protein corresponding to aa 37-72 was able to undergo homotypic interaction and bound to a native DEN2 core protein. The results of this study indicated that the homotypic interaction domain of the DEN2 core protein is located at aa 37-72 and that the internal hydrophobic domain located at aa 44-60 plays a pivotal role in core-to-core homotypic interaction. PMID- 15269373 TI - Mx1 GTPase accumulates in distinct nuclear domains and inhibits influenza A virus in cells that lack promyelocytic leukaemia protein nuclear bodies. AB - The interferon-induced murine Mx1 GTPase is a nuclear protein. It specifically inhibits influenza A viruses at the step of primary transcription, a process known to occur in the nucleus of infected cells. However, the exact mechanism of inhibition is still poorly understood. The Mx1 GTPase has previously been shown to accumulate in distinct nuclear dots that are spatially associated with promyelocytic leukaemia protein (PML) nuclear bodies (NBs), but the significance of this association is not known. Here it is reported that, in cells lacking PML and, as a consequence, PML NBs, Mx1 still formed nuclear dots. These dots were indistinguishable from the dots observed in wild-type cells, indicating that intact PML NBs are not required for Mx1 dot formation. Furthermore, Mx1 retained its antiviral activity against influenza A virus in these PML-deficient cells, which were fully permissive for influenza A virus. Nuclear Mx proteins from other species showed a similar subnuclear distribution. This was also the case for the human MxA GTPase when this otherwise cytoplasmic protein was translocated into the nucleus by virtue of a foreign nuclear localization signal. Human MxA and mouse Mx1 do not interact or form heterooligomers. Yet, they co-localized to a large degree when co-expressed in the nucleus. Taken together, these findings suggest that Mx1 dots represent distinct nuclear domains ('Mx nuclear domains') that are frequently associated with, but functionally independent of, PML NBs. PMID- 15269374 TI - Influenza A viruses in feral Canadian ducks: extensive reassortment in nature. AB - The current dogma of influenza accepts that feral aquatic birds are the reservoir for influenza A viruses. Although the genomic information of human influenza A viruses is increasing, little of this type of data is available for viruses circulating in feral waterfowl. This study presents the genetic characterization of 35 viruses isolated from wild Canadian ducks from 1983 to 2000, as the first attempt at a comprehensive genotypic analysis of influenza viruses isolated from feral ducks. This study demonstrates that influenza virus genes circulating in Canadian ducks have achieved evolutionary stasis. The majority of these duck virus genes are clustered in distinct North American clades; however, some H6 and H9 genes are clustered with those from Eurasian viruses. Genes appeared to reassort in a random fashion. None of the genotypes identified remained present throughout all of the years examined and most PA and PB2 genes that crossed over into swine were clustered in one phylogenetic grouping. Additionally, matrix genes were identified that branch very early in the evolutionary tree. These findings demonstrate the diversity of the influenza virus gene pool in Canadian ducks, and suggest that genes which cluster in specific phylogenetic groupings in the PB2 and PA genes can be used for markers of viruses with the potential for crossing the species barrier. A more comprehensive study of this important reservoir is needed to provide further insight into the genomic composition of viruses that crossover the species barrier, which would be a useful component to pandemic planning. PMID- 15269375 TI - Total viral genome copies and virus-Ig complexes after infection with influenza virus in the nasal secretions of immunized mice. AB - The kinetics of infectious virus (p.f.u.), total virus and virus-Ig complex formation following influenza A/PR8 (H1N1) viral infection was examined in the nasal secretions of naive mice and mice immunized with A/PR8, A/Yamagata (H1N1), A/Guizhou (H3N2) and B/Ibaraki influenza viruses. The total number of virus particles and the number within virus-Ig complexes, captured in advance using an anti-mouse Ig-coated plate, were determined on the basis of viral genome copy number using quantitative RT-PCR. The kinetics of infectious and total virus particle formation, the latter of which increased by 10(3)-10(4)-fold above infectious virus numbers, showed that virus elimination from the nasal area was earlier in A/PR8, A/Yamagata and A/Guizhou-X virus-immunized mice, in decreasing order, compared with naive mice. Early virus elimination correlated with the level of A/PR8 virus-reactive antibodies in immunized mice. Virus elimination coincided with the appearance of virus-Ig complexes shortly after infection. This result suggested that antibodies led to the formation of immune complexes in a dose-dependent manner together with a reduction in number of infectious virus particles. The fact that a large number of virus particles was observed in immune complexes for a wide range antibody levels made it difficult to detect slight differences in virus number within the immune complexes, depending on antibody level. These results suggested that the formation of virus-Ig complexes in virus immunized mice shortly after infection is involved in early virus elimination, which is determined by the strength of protective immunity against challenge viruses. PMID- 15269376 TI - Active NF-kappaB signalling is a prerequisite for influenza virus infection. AB - Influenza virus still poses a major threat to human health. Despite widespread vaccination programmes and the development of drugs targeting essential viral proteins, the extremely high mutation rate of influenza virus still leads to the emergence of new pathogenic virus strains. Therefore, it has been suggested that cellular cofactors that are essential for influenza virus infection might be better targets for antiviral therapy. It has previously been reported that influenza virus efficiently infects Epstein-Barr virus-immortalized B cells, whereas Burkitt's lymphoma cells are virtually resistant to infection. Using this cellular system, it has been shown here that an active NF-kappaB signalling pathway is a general prerequisite for influenza virus infection of human cells. Cells with low NF-kappaB activity were resistant to influenza virus infection, but became susceptible upon activation of NF-kappaB. In addition, blocking of NF kappaB activation severely impaired influenza virus infection of otherwise highly susceptible cells, including the human lung carcinoma cell lines A549 and U1752 and primary human cells. On the other hand, infection with vaccinia virus was not dependent on an active NF-kappaB signalling pathway, demonstrating the specificity of this pathway for influenza virus infection. These results might be of major importance for both the development of new antiviral therapies and the understanding of influenza virus biology. PMID- 15269377 TI - Cytokine and contact-dependent activation of natural killer cells by influenza A or Sendai virus-infected macrophages. AB - NK cells participate in innate immune responses by secreting gamma interferon (IFN-gamma) and by destroying virus-infected cells. Here the interaction between influenza A or Sendai virus-infected macrophages and NK cells has been studied. A rapid, cell-cell contact-dependent production of IFN-gamma from NK cells cultured with virus-infected macrophages was observed. Expression of the MHC class I related chain B (MICB) gene, a ligand for NK cell-activating receptor NKG2D, was upregulated in virus-infected macrophages suggesting a role for MICB in the activation of the IFN-gamma gene in NK cells. IL12Rbeta2, IL18R and T-bet mRNA synthesis was enhanced in NK cells cultured with virus-infected macrophages. Upregulation of these genes was dependent on macrophage-derived IFN-alpha. In contrast to IL12Rbeta2, expression of WSX-1/TCCR, a receptor for IL27, was reduced in NK cells in response to virus-induced IFN-alpha. In conclusion, these results show that virus-infected macrophages activate NK cells via cytokines and direct cellular interactions and further emphasize the role of IFN-alpha in the activation of innate immunity. PMID- 15269378 TI - Human CD8(+) T cell responses against five newly identified respiratory syncytial virus-derived epitopes. AB - CD8(+) T lymphocytes play a major role in the clearance of respiratory syncytial virus (RSV) infections. To be able to study the primary CTL response in RSV infected children, epitopes presented by a set of commonly used HLA alleles (HLA A1, -A3, -B44 and -B51) were searched for. Five epitopes were characterized derived from the matrix (M), non-structural (NS2) and second matrix (M2) proteins of RSV. All epitopes were shown to be processed and presented by RSV-infected antigen-presenting cells. HLA-A1 tetramers for one of these epitopes derived from the M protein were constructed and used to quantify and phenotype the memory CD8(+) T cell pool in a panel of healthy adult donors. In about 60 % of the donors, CD8(+) T cells specific for the M protein could be identified. These cells belonged to the memory T cell subset characterized by expression of CD27 and CD28, and down-regulation of CCR7 and CD45RA. The frequency of tetramer positive cells varied between 0.4 and 3 per 10(4) CD8(+) T cells in PBMC of healthy asymptomatic adult donors. PMID- 15269379 TI - The P gene of Newcastle disease virus does not encode an accessory X protein. AB - Many paramyxoviruses encode non-essential accessory proteins that are involved in the regulation of virus replication and inhibition of cellular antiviral responses. It has been suggested that the P gene mRNA of Newcastle disease virus (NDV) encodes an accessory protein - the so-called X protein - by translation initiation at a conserved in-frame AUG codon at position 120. Using a monoclonal antibody that specifically detected the P and X proteins, it was shown that an accessory X protein was not expressed in NDV-infected cells. Recombinant NDV strains in which the AUG was changed into a GCC (Ala) or GUC (Val) codon were viable but showed a reduction in virulence, probably because the amino acid change affected the function of the P and/or V protein. PMID- 15269380 TI - Immunization with dendritic cells can break immunological ignorance toward a persisting virus in the central nervous system and induce partial protection against intracerebral viral challenge. AB - Dendritic cells (DCs) have been used successfully to induce CD8 T cells that control virus infections and growth of tumours. The efficacy of DC-mediated immunization for the control of neurotropic Borna disease virus (BDV) in mice was evaluated. Certain strains of mice only rarely develop spontaneous neurological disease, despite massive BDV replication in the brain. Resistance to disease is due to immunological ignorance toward BDV antigen in the central nervous system. Ignorance in mice can be broken by immunization with DCs coated with TELEISSI, a peptide derived from the N protein of BDV, which represents the immunodominant cytotoxic T lymphocyte epitope in H-2(k) mice. Immunization with TELEISSI-coated DCs further induced solid protective immunity against intravenous challenge with a recombinant vaccinia virus expressing BDV-N. Interestingly, however, this immunization scheme induced only moderate protection against intracerebral challenge with BDV, suggesting that immune memory raised against a shared antigen may be sufficient to control a peripherally replicating virus, but not a highly neurotropic virus that is able to avoid activation of T cells. This difference might be due to the lack of BDV-specific CD4 T cells and/or inefficient reactivation of DC-primed, BDV-specific CD8 T cells by the locally restricted BDV infection. Thus, a successful vaccine against persistent viruses with strong neurotropism should probably induce antiviral CD8 (as well as CD4) T-cell responses and should favour the accumulation of virus-specific memory T cells in cervical lymph nodes. PMID- 15269381 TI - Host RNA polymerase II makes minimal contributions to retroviral frame-shift mutations. AB - The rate of mutation during retrovirus replication is high. Mutations can occur during transcription of the viral genomic RNA from the integrated provirus or during reverse transcription from viral RNA to form viral DNA or during replication of the proviral DNA as the host cell is dividing. Therefore, three polymerases may all contribute to retroviral evolution: host RNA polymerase II, viral reverse transcriptases and host DNA polymerases, respectively. Since the rate of mutation for host DNA polymerase is very low, mutations are more likely to be caused by the host RNA polymerase II and/or the viral reverse transcriptase. A system was established to detect the frequency of frame-shift mutations caused by cellular RNA polymerase II, as well as the rate of retroviral mutation during a single cycle of replication in vivo. In this study, it was determined that RNA polymerase II contributes less than 3 % to frame-shift mutations that occur during retrovirus replication. Therefore, the majority of frame-shift mutations detected within the viral genome are the result of errors during reverse transcription. PMID- 15269382 TI - Human T-cell leukaemia virus type I is highly sensitive to UV-C light. AB - The biological characteristics of human T-cell leukaemia virus type I (HTLV-I) are not yet well understood. UV light C (UV-C) sensitivity of HTLV-I was studied using a newly established infectivity assay: infection with cell-free HTLV-I dose dependently induced syncytial plaques in cat cells transduced with the tax1 gene of HTLV-I. HTLV-I was inactivated by a much lower UV dose than bovine leukaemia virus (BLV). The D(10) (10 % survival dose) of HTLV-I was about 20 J m(-2), while that of BLV was about 180 J m(-2), which was similar to the reported D(10) of BLV. The UV sensitivity of HTLV-I and BLV was also examined by detecting viral DNA synthesis 24 h after infection. The D(10) values determined by PCR using the gag primers for HTLV-I and BLV were close to those determined by the infectivity assays. Further PCR analyses were then performed to determine D(10) values using several different primers located between the 5'-long terminal repeat (5'-LTR) and the tax1 gene. The difference in UV sensitivity between HTLV-I and BLV was detected very early during replication, even during reverse transcription of the 5'-LTR of irradiated viruses, and became more prominent as reverse transcription proceeded towards the tax1 gene. Chimeric mouse retroviruses that contain the LTR tax1 fragments of HTLV-I and BLV were made and hardly any difference in UV sensitivity was detected between them, suggesting that the difference was not determined by the linear RNA sequences of HTLV-I and BLV. HTLV-I was found to be much more sensitive than other retroviruses to UV. PMID- 15269383 TI - Enhanced cellular immunity and systemic control of SHIV infection by combined parenteral and mucosal administration of a DNA prime MVA boost vaccine regimen. AB - The immunogenicity and protective efficacy of a DNA and recombinant modified vaccinia Ankara (MVA) vaccine administered by two different routes were investigated. DNA expressing HIV-1 IIIB env, gag, RT, rev, tat and nef, and MVA expressing HIV-1 IIIB nef, tat and rev and simian immunodeficiency virus (SIV) macJ5 gag/pol and vaccinia HIV-1 env, were used as immunogens. Four cynomolgus macaques received DNA intramuscularly (i.m.) at month 0 and intrarectally (i.r.) and intra-orally (i.o.) at 2 months, followed by MVA i.m. at 4 months and i.r. and i.o. at 8 months. Another group of four monkeys received the same immunogens but only i.m. Overall, stronger cellular immune responses measured by ELISPOT and T-cell proliferation assay were detected in the group primed i.m. and boosted mucosally. Following homologous intravenous simian-human immunodeficiency virus (SHIV) challenge, one of eight vaccinated animals was completely protected. This monkey, immunized i.m. and i.r.+i.o., exhibited the highest levels of HIV Env, Nef and Tat antibodies, high HIV Tat cytotoxic T-lymphocyte activity and T lymphocyte proliferative responses to HIV Env. Four weeks post-challenge none of the monkeys immunized i.m. and i.r.+i.o., and only two out of four animals immunized i.m., demonstrated detectable plasma viral RNA levels. In contrast, all eight control animals had demonstrable plasma viral RNA levels 4 weeks post challenge. Thus, stronger cellular immune responses and reduction of challenge virus burden were demonstrated in animals immunized i.m. as well as mucosally, compared with animals immunized i.m. only. The breadth and magnitude of the induced immune responses correlated with protective efficacy. PMID- 15269384 TI - Characterization of germline porcine endogenous retroviruses from Large White pig. AB - Porcine endogenous retroviruses (PERV) are of concern when the microbiological safety aspects of xenotransplantation are considered. Four unique isolates of PERV B have been identified previously from a lambda library constructed from genomic DNA from a Large White pig. This study shows that none of these isolates are replication competent when transfected into permissive human or pig cells in vitro, and the removal of flanking genomic sequences does not confer a human tropic replication competent (HTRC) phenotype on these PERV proviruses. Analysis of the envelope sequences revealed that PERV B demonstrated high similarity to the envelope sequences derived from replication-competent PERV, indicating that lack of replication competence does not appear to be attributable to this region of the provirus. These data complement recent findings that HTRC PERV are recombinants between the PERV A and PERV C subgroups, and that these recombinants are not present in the germline of miniature swine. Together, these results indicate that these individual PERV B proviruses are unlikely to give rise to HTRC PERV. PMID- 15269385 TI - The p36 and p95 replicase proteins of Carnation Italian ringspot virus cooperate in stabilizing defective interfering RNA. AB - The p36 and p95 proteins of Carnation Italian ringspot virus (CIRV), when expressed in Saccharomyces cerevisiae, supported the replication of defective interfering (DI) RNA. Double-label confocal immunofluorescence showed that both proteins localized to mitochondria, independently of each other. DI RNA progeny was localized by in situ hybridization both to mitochondria and to their proximity. Fractionation of cell extracts showed that replicase proteins associated with membranes with a consistent portion of DI RNA. DI RNA transcripts were stabilized more efficiently when co-expressed with both p36 and p95 than with either protein alone. By using the copper-inducible CUP1 promoter, p36 was shown to have an effect on DI RNA stability only above a threshold concentration, suggesting an 'all-or-none' behaviour. Conversely, the stabilizing activity of p95 was proportional to protein concentration in the range examined. Similarly, DI RNA replication level was proportional to p95 concentration and depended on a threshold concentration of p36. PMID- 15269386 TI - Population structure and genetic variability within isolates of Grapevine fanleaf virus from a naturally infected vineyard in France: evidence for mixed infection and recombination. AB - The nematode-borne Grapevine fanleaf virus, from the genus Nepovirus in the family Comoviridae, causes severe degeneration of grapevines in most vineyards worldwide. We characterized 347 isolates from transgenic and conventional grapevines from two vineyard sites in the Champagne region of France for their molecular variant composition. The population structure and genetic diversity were examined in the coat protein gene by IC-RT-PCR-RFLP analysis with EcoRI and StyI, and nucleotide sequencing, respectively. RFLP data suggested that 55 % (191 of 347) of the isolates had a population structure consisting of one predominant variant. Sequencing data of 51 isolates representing the different restrictotypes confirmed the existence of mixed infection with a frequency of 33 % (17 of 51) and showed two major predominant haplotypes representing 71 % (60 of 85) of the sequence variants. Comparative nucleotide diversity among population subsets implied a lack of genetic differentiation according to host (transgenic vs conventional) or field site for most restrictotypes (17 of 18 and 13 of 18) and for haplotypes in most phylogenetic groups (seven of eight and six of eight), respectively. Interestingly, five of the 85 haplotypes sequenced had an intermediate divergence (0.036-0.066) between the lower (0.005-0.028) and upper range (0.083-0.138) of nucleotide variability, suggesting the occurrence of homologous RNA recombination. Sequence alignments clearly indicated a mosaic structure for four of these five variants, for which recombination sites were identified and parental lineages proposed. This is the first in-depth characterization of the population structure and genetic diversity in a nepovirus. PMID- 15269387 TI - Mutation of Phe50 to Ser50 in the 126/183-kDa proteins of Odontoglossum ringspot virus abolishes virus replication but can be complemented and restored by exact reversion. AB - Sequence comparison of a non-biologically active full-length cDNA clone of Odontoglossum ringspot virus (ORSV) pOT1 with a biologically active ORSV cDNA clone pOT2 revealed a single nucleotide change of T-->C at position 211. This resulted in the change of Phe50 in OT2 to Ser50 in OT1. It was not the nucleotide but the amino acid change of Phe50 that was responsible for the inability of OT1 to replicate. Time-course experiments showed that no minus-strand RNA synthesis was detected in mutants with a Phe50 substitution. Corresponding mutants in Tobacco mosaic virus (TMV) showed identical results, suggesting that Phe50 may play an important role in replication in all tobamoviruses. Complementation of a full-length mutant OT1 was demonstrated in a co-infected local-lesion host, a systemic host and protoplasts by replication-competent mutants tORSV.GFP or tORSV.GFPm, and further confirmed by co-inoculation using tOT1.GFP+tORSV (TTC), suggesting that ORSV contains no RNA sequence inhibitory to replication in trans. Surprisingly, a small number of exact revertants were detected in plants inoculated with tOT1+tORSV.GFPm or tOT1.GFP+tORSV (TTC). No recombination was detected after screening of silent markers in virus progeny extracted from total RNA or viral RNA from inoculated and upper non-inoculated leaves as well as from transfected protoplasts. Exact reversion from TCT (OT1) to TTT (OT2), rather than recombination, restored its replication function in co-inoculated leaves of Nicotiana benthamiana. PMID- 15269388 TI - Nucleo-cytoplasmic shuttling of the beet necrotic yellow vein virus RNA-3-encoded p25 protein. AB - The protein p25 encoded by beet necrotic yellow vein virus (BNYVV) RNA-3 is involved in symptom expression of infected plants. Confocal microscopy analysis of wild-type and mutated p25 fused to GFP and transiently expressed in BY-2 tobacco suspension cells identified a nuclear localization signal (NLS) in the N terminal part of the protein. Functionality of the NLS was confirmed by pull-down assays using rice and pepper importin-alpha. Furthermore, it was demonstrated that p25 contains a nuclear export sequence sensitive to leptomycin B. The nuclear export signal (NES) was characterized by mutagenesis. A GFP-p25 fusion protein expressed during a BNYVV infection of Chenopodium quinoa leaves had the same subcellular localization as observed during transient expression in BY-2 cells. The symptom phenotype induced by expression of GFP-p25 during infection was similar to that induced by wild-type virus. Studies with mutated derivatives of GFP-p25 revealed that symptom phenotype was altered when the subcellular localization of GFP-p25 was modified. PMID- 15269389 TI - Characterization of two distinct prion strains derived from bovine spongiform encephalopathy transmissions to inbred mice. AB - Distinct prion strains can be distinguished by differences in incubation period, neuropathology and biochemical properties of disease-associated prion protein (PrP(Sc)) in inoculated mice. Reliable comparisons of mouse prion strain properties can only be achieved after passage in genetically identical mice, as host prion protein sequence and genetic background are known to modulate prion disease phenotypes. While multiple prion strains have been identified in sheep scrapie and Creutzfeldt-Jakob disease, bovine spongiform encephalopathy (BSE) is thought to be caused by a single prion strain. Primary passage of BSE prions to different lines of inbred mice resulted in the propagation of two distinct PrP(Sc) types, suggesting that two prion strains may have been isolated. To investigate this further, these isolates were subpassaged in a single line of inbred mice (SJL) and it was confirmed that two distinct prion strains had been identified. MRC1 was characterized by a short incubation time (110+/-3 days), a mono-glycosylated-dominant PrP(Sc) type and a generalized diffuse pattern of PrP immunoreactive deposits, while MRC2 displayed a much longer incubation time (155+/-1 days), a di-glycosylated-dominant PrP(Sc) type and a distinct pattern of PrP-immunoreactive deposits and neuronal loss. These data indicate a crucial involvement of the host genome in modulating prion strain selection and propagation in mice. It is possible that multiple disease phenotypes may also be possible in BSE prion infection in humans and other animals. PMID- 15269390 TI - Evaluation of new cell culture inhibitors of protease-resistant prion protein against scrapie infection in mice. AB - In vitro inhibitors of the accumulation of abnormal (protease-resistant) prion protein (PrP-res) can sometimes prolong the lives of scrapie-infected rodents. Here, transgenic mice were used to test the in vivo anti-scrapie activities of new PrP-res inhibitors, which, because they are approved drugs or edible natural products, might be considered for clinical trials in humans or livestock with transmissible spongiform encephalopathies (TSEs). These inhibitors were amodiaquine, thioridazine, thiothixene, trifluoperazine, tetrandrine, tannic acid and polyphenolic extracts of tea, grape seed and pine bark. Test compounds were administered for several weeks beginning 1-2 weeks prior to, or 2 weeks after, intracerebral or intraperitoneal 263K scrapie challenge. Tannic acid was also tested by direct preincubation with inoculum. None of the compounds significantly prolonged the scrapie incubation periods. These results highlight the need to assess TSE inhibitors active in cell culture against TSE infections in vivo prior to testing these compounds in humans and livestock. PMID- 15269391 TI - The biology of cyclins and cyclin-dependent protein kinases: an introduction. PMID- 15269392 TI - In situ immunofluorescence analysis: immunofluorescence microscopy. PMID- 15269393 TI - In situ immunofluorescence analysis: analyzing RNA synthesis by 5-bromouridine-5' triphosphate labeling. PMID- 15269394 TI - Immunofluorescence analysis using epitope-tagged proteins: in vitro system. PMID- 15269395 TI - Analysis of in vivo gene expression using epitope-tagged proteins. PMID- 15269396 TI - Chromatin immunoprecipitation. PMID- 15269397 TI - Protein-deoxyribonucleic acid interactions linked to gene expression: electrophoretic mobility shift assay. PMID- 15269398 TI - Protein-deoxyribonucleic acid interactions linked to gene expression: DNase I digestion. PMID- 15269399 TI - Protein-deoxyribonucleic acid interactions linked to gene expression: ligation mediated polymerase chain reaction. PMID- 15269400 TI - Assays for cyclin-dependent kinase inhibitors. PMID- 15269401 TI - Protein degradation via the proteosome. PMID- 15269402 TI - The transformed phenotype. PMID- 15269403 TI - A morphologic approach to detect apoptosis based on electron microscopy. PMID- 15269404 TI - Detection of apoptotic deoxyribonucleic acid break by in situ nick translation. PMID- 15269405 TI - Induction of deoxyribonucleic acid damage by alkylating agents. PMID- 15269406 TI - Induction of deoxyribonucleic acid damage by gamma irradiation. PMID- 15269407 TI - Ultraviolet irradiation of cells. PMID- 15269408 TI - Transient production of retroviral- and lentiviral-based vectors for the transduction of Mammalian cells. PMID- 15269409 TI - Determination of functional viral titer by drug-resistance colony assay, expression of green fluorescent protein, and beta-galactoside staining. PMID- 15269410 TI - Retroviral and lentiviral vector titration by the analysis of the activity of viral reverse transcriptase. PMID- 15269411 TI - Single and double colloidal gold labeling in postembedding immunoelectron microscopy. PMID- 15269412 TI - Multifluorescence labeling and colocalization analyses. PMID- 15269413 TI - Host cell compatibility in protein expression. AB - The expression of cloned genes in prokaryotic or eukaryotic host cells provides the means not only for the study of gene function but also for the production of substantial amounts of protein and nonprotein molecules for commercial and investigational use. In the case of proteins, strategies for determining the most appropriate vector-host combination for the expression of an exogenous gene depend on a diverse range of factors that relate ultimately to the properties of the gene and its product. The approach used in the downstream purification of the product is another factor that impinges on this selection. However, among the most important considerations in the choice of vector and host in ensuring the maximal amount of expression is the compatibility of the host cells to translate the RNA transcript, to ensure the proper folding of the product, and to sustain the protein in the intact and functional state. PMID- 15269414 TI - Production of recombinant proteins: challenges and solutions. AB - Efficient strategies for the production of recombinant proteins are gaining increasing importance, as more applications that require high amounts of high quality proteins reach the market. Higher production efficiencies and, consequently, lower costs of the final product are needed for obtaining a commercially viable process. In this chapter, common problems in recombinant protein production are reviewed and strategies for their solution are discussed. Such strategies include molecular biology techniques, as well as manipulation of the culture environment. Finally, specific problems relevant to different hosts are discussed. PMID- 15269415 TI - Folding-promoting agents in recombinant protein production. AB - Recombinant protein production has become an essential tool for providing the necessary amounts of a protein of interest to either research or therapy. The target proteins are not in every case soluble and/or correctly folded. That is why different production parameters, such as host, cultivation conditions, and co expression of chaperones and foldases, are applied in order to gain functional recombinant proteins. Furthermore, the addition of folding-promoting agents during the cultivation is increasingly performed. The impact of all these strategies cannot be predicted and must be analyzed and optimized for the corresponding target protein. In this chapter recent cases of using folding promoting agents in recombinant protein production are reviewed and discussed with respect to their in vivo applicability. Their effects in the cells are mostly not known in detail but at least partially comparable with the in vitro mode of action. The corresponding in vitro effects are also included in the chapter in order to facilitate a decision about their potential in vivo use. PMID- 15269416 TI - Back to basics: pBR322 and protein expression systems in E. coli. AB - The extensive variety of plasmid-based expression systems in E. coli resulted from the fact that there is no single strategy for achieving maximal expression of every cloned gene. Although a number of strategies have been implemented to deal with problems associated to gene transcription and translation, protein folding, secretion, location, posttranslational modifications, particularities of different strains, and the like and more integrated processes have been developed, the basic plasmid-borne elements and their interaction with the particular host strain will influence the overall expression system and final productivity. Plasmid vector pBR322 is a well-established multipurpose cloning vector in laboratories worldwide, and a large number of derivatives have been created for specific applications and research purposes, including gene expression in its natural host, E. coli, and few other bacteria. The early characterization of the molecule, including its nucleotide sequence, replication and maintenance mechanisms, and determination of its coding regions, accounted for its success, not only as a universal cloning vector, but also as a provider of genes and an origin of replication for other intraspecies vectors. Since the publication of the aforementioned reviews, novel discoveries pertaining to these issues have appeared in the literature that deepen the understanding of the plasmid's features, behavior, and impact in gene expression systems, as well as some important strain characteristics that affect plasmid replication and stability. The objectives of this review include updating and discussing the new information about (1) the replication and maintenance of pBR322; (2) the host related modulation mechanisms of plasmid replication; (3) the effects of growth rate on replication control, stability, and recombinant gene expression; (4) ways for plasmid amplification and elimination. Finally, (5) a summary of novel ancillary studies about pBR322 is presented. PMID- 15269417 TI - alpha-complementation-enabled T7 expression vectors and their use for the expression of recombinant polypeptides for protein transduction experiments. AB - Over the past few years protein transduction has emerged as a powerful means for the delivery of proteins into cultured cells and into whole mice. This method is based on the ability of proteins containing protein transduction domains (PTDs), short stretches of 9-16 predominantly basic amino acids, to traverse the cytoplasmic membrane and accumulate inside cells in a time- and dose-dependent fashion. The number of PTDs, both natural and synthetic, is constantly expanding, as is the need to test newly discovered PTDs for their ability to mediate the internalization of the corresponding fusion proteins. Here we describe a strategy and methodology that can be used for the construction of vectors for the T7 RNA polymerase-driven expression of PTD fusions. The cloning in these vectors is facilitated by alpha-complementation. Also, these vectors are small in size (less than 3 kbp) and express influenza virus hemagglutinin tag as well as His tag as part of the fusion for immunological identification and purification respectively of expressed proteins. PMID- 15269418 TI - Expression of recombinant alkaline phosphatase conjugates in Escherichia coli. AB - The methods described in this article are relative to the use of a positive cloning/screening recombinant system for the generation in Escherichia coli of foreign proteins fused to a highly active bacterial alkaline phosphatase (PhoA) variant as reporter enzyme. Appropriate insertion of the DNA encoding the foreign peptides, proteic domains, or proteins between codons +6 and +7 of the phoa gene restores the initial frame of the phoa gene in the vector. Consequently, only recombinant clones appear as blue colonies when plating onto an agar medium containing a chromogenic substrate for PhoA. The presence of an intact PhoA signal peptide yields to a systematic secretion of the fusion proteins into the periplasm where the PhoA dimerises to its active form, and disulfides can be formed if necessary. The resultant PhoA-tagged proteins are particularly convenient novel tools that can be used in a wide range of applications, including expression, epitope mapping, histochemistry, immunoblotting, mutant analysis, and competition or sandwich ELISAs. Expression of an scFv antibody fragment derived from an IgG2a/kappa immunoglobulin specific for curaremimetic toxins from snake (named M-alpha2-3), will be used to illustrate the methods utilized for its cloning, expression in E.coli, extraction, and functional characterization. PMID- 15269419 TI - Overexpression of chromosomal genes in Escherichia coli. AB - Conversion of some carbon sources into desired compounds by a biological system is the goal of many biotechnologists. The understanding of the mechanisms by which an organism does these conversions permits the improved production of specific metabolites. Pathway engineering involves the strategies to modify cells to overproduce desired molecules. We describe here the methodology to modify chromosomal genes by replacing their native regulatory regions with promoter cassettes to increase or deregulate expression of chromosomal genes. PMID- 15269420 TI - Chromosomal expression of foreign and native genes from regulatable promoters in Escherichia coli. AB - A two-step cloning system for expression of foreign and native genes from heterologous promoters in single copy from the E. coli chromosome is described. The system is based on the conditional-replication integration and modular CRIM plasmid technology and new CRIM plasmids described herein. The gene of interest is first synthesized by Polymerase chain reaction (PCR) by using primers with specially designed SapI site extensions and cloned into the SapI CRIM cloning plasmid pKZ20. The gene is then subcloned with SapI into a CRIM expression plasmid, such as pKZ14, pLZ41, or pLZ42, which carry the regulatable promoter araBp8, rhaBp3, or lacUV5, respectively. The system is described for gfp, which encodes green fluorescence protein, as an example. The resulting CRIM expression plasmids are then integrated in single copy into a chromosomal phage attachment site by supplying integrase from a helper plasmid. Such integrants can be used for conditional expression of any target gene in single copy on the chromosome, especially in gene-structure-function studies where it is important to avoid copy number artifacts. PMID- 15269421 TI - Plasmid vectors for marker-free chromosomal insertion of genetic material in Escherichia coli. AB - A method to achieve the insertion of genetic material into the chromosome of Escherichia coli is described. The method is based on the use of integration vectors from the pBRINTs-rAnbR family. These vectors offer the choice of using the antibiotics chloramphenicol, gentamycin, or kanamycin to select for chromosomal integration events. In addition, it is possible to eliminate these chromosomal antibiotic resistance markers, after integration has taken place. The overall insertion strategy is as follows: a fragment containing the gene(s) to be integrated in the chromosome is inserted into the multiple cloning site of a pBRINTs-rAnbR vector and the resulting plasmid is used to transform E. coli cells. The plasmid is first allowed to replicate in the cell at the permissive temperature of 30 degrees C. Next, the temperature of the culture is raised to 44 degrees C to inhibit plasmid replication and to select for the integrants in the presence of the appropriate antibiotic. Chromosomal excision of the AnbR gene can then be catalyzed by the Cre recombinase that is transiently expressed in the cell from the temperature-sensitive pJW168 plasmid. This plasmid is finally eliminated from the cells by increasing the temperature of the culture to 44 degrees C. PMID- 15269422 TI - Copy-control pBAC/oriV vectors for genomic cloning. AB - The use of the improved BAC system for cloning genomic DNA and library constructions is described. This system retains all the advantages of the original BACs but, in addition, permits, on command, amplification of the BAC plasmids and cloned DNA. This system consists of (1) plasmid pBAC/oriV containing an additional replication origin, oriV, and (2) a host carrying the up-mutants of the trfA replicator gene expressed from the l-arabinose-inducible Para promoter. The pBAC/oriV clones are always maintained in the single-copy state, but if more DNA is required, they could be amplified up to 100-fold, depending on the size of the cloned insert. PMID- 15269423 TI - Copy-control tightly regulated expression vectors based on pBAC/oriV. AB - A novel type of expression vectors with a dual regulation of both the plasmid copy number and gene expression, is described. The most important and beneficial feature of these vectors is that when they are not induced, they are maintained as a single-copy plasmid, and therefore, any residual expression is much more tightly regulated than for the conventional multicopy expression vectors. The simplest version of these copy-control expression vectors is based on the pBAC/oriV plasmid that carries the trfA up-mutant gene under control of the l arabinose-inducible Para promoter (araC-PBAD). The same promoter controls expression of a gene cloned into MCS. Thus, addition of the inducer (l-arabinose) simultaneously turns on amplification of the plasmid and expression of the cloned gene. Net result is about a 50,000-fold increase in the cloned gene expression. However, when not induced, background expression level is very low, which is important for the maintenance of any "toxic" genes. This vector could be used in most E. coli hosts. Similar versions of the described vector employ the rhamnose inducible Prha promoter (rhaS-Prha). Other expression systems allow independent regulation of the plasmid amplification and of the cloned gene expression, and some also use the PLtetO-1 promoter. Copy-control expression vector pETcoco, based on the pT7lacO promoter, is commercially available. PMID- 15269424 TI - Cell-free protein synthesis with prokaryotic combined transcription-translation. AB - Cell-free biology exploits and studies complex biological processes in a controlled environment without intact cells. One model system is prokaryotic cell free protein synthesis. This technology offers an attractive and convenient approach to produce properly folded recombinant DNA (rDNA) proteins on a laboratory scale, screen PCR fragment libraries in a high-throughput format, express pharmaceutical proteins, incorporate labeled or unnatural amino acids into proteins, and activate microbial physiology to allow for investigation of biological systems. We describe the preparation of materials necessary for the expression, quantification, and purification of rDNA proteins from active Escherichia coli extracts. PMID- 15269425 TI - Genetic tools for the manipulation of moderately halophilic bacteria of the family Halomonadaceae. AB - Moderately halophilic bacteria of the family Halomonadaceae (Halomonas, Chromohalobacter, and Zymobacter) have promising applications in biotechnology as a source of compatible solutes (stabilizers of biomolecules and cells), salt tolerant enzymes, biosurfactants, and extracellular polysaccharides, among other products. In addition, they offer a number of advantages to be used as cell factories, alternative to conventional prokaryotic hosts like E. coli or Bacillus, for the production of recombinant proteins: (1) their high salt tolerance decreases to a minimum the necessity for aseptic conditions, resulting in cost-reducing conditions; (2) they are very easy to grow and maintain in the laboratory, and their nutritional requirements are simple; and (3) the majority can use a large range of compounds as a sole carbon and energy source. In this decade, the efforts of our group and others have made possible the genetic manipulation of this bacterial group. In this review, the most relevant tools are described, with emphasis given to cloning vectors, genetic exchange mechanisms, mutagenesis approaches, and reporter genes. Due to its relevance for genetic studies, complementary sections describing the influence of salinity on the susceptibility of moderately halophilic bacteria to antimicrobials, as well as the growth media most routinely used, culture conditions, and nucleic acid isolation procedures for these microorganisms, are included. PMID- 15269426 TI - Gene transfer and expression of recombinant proteins in moderately halophilic bacteria. AB - Moderately halophilic bacteria (MHB) of the genera Halomonas and Chromohalobacter have been used as hosts for the expression of heterologous proteins of biotechnological interest, thus expanding their potential to be used as cell factories for various applications. This chapter deals with the methodology for the construction of recombinant plasmids, their transfer to a number of MHB, and the assaying of the corresponding heterologous proteins activity. The transferred genes include (1) inaZ, encoding the ice nucleation protein of the plant pathogen Pseudomonas syringae, (2) gfp, encoding a green fluorescent protein from the marine bioluminescent jellyfish Aequorea victoria, and (3) the alpha-amylase gene from the hyperthermophilic archeon Pyrococcus woesei. Vector pHS15, which was designed for expression of heterologous proteins in both E. coli and MHB, was used for the subcloning and transfer of the above genes. The recombinant constructs were introduced to MHB by assisted conjugal transfer from E. coli donors. The expression and function of the recombinant proteins in the MHB transconjugants is described. PMID- 15269427 TI - Recombinant protein production in Antarctic Gram-negative bacteria. AB - This review reports some results from our laboratory on the setting up of a psychrophilic expression system for the homologous/heterologous protein production in cold-adapted bacteria by using natural plasmids as cloning vectors. By screening some Antarctic bacteria for the presence of extrachromosomal elements, we identified three new plasmids, pMtBL from Pseudoalteromonas haloplanktis TAC125, and pTAUp and pTADw, from Psychrobacter sp. TA144. The latter autoreplicating elements were isolated, cloned, and fully sequenced and their molecular characterisation was carried out; however, we focused our attention on the small multicopy plasmid, pMtBL, from the Gram-negative P. haloplanktis TAC125 strain. This episome turned out to be an interesting extrachromosomal element, since it displays unique molecular features as its transcriptional inactivity. Being cryptic, the inheritance of pMtBL totally relied on the efficiency of its replication function. This function was bound to a region of about 850 bp, identified by an in vivo assay based on the possibility to efficiently mobilize plasmidic DNA from a mesophilic donor (Escherichia coli) to psychrophilic recipient by intergeneric conjugation. This information was instrumental in the construction of a shuttle vector, able to replicate either in E. coli or in several cold-adapted hosts (clone Q). Since the conversion of a cloning system into an expression vector requires the insertion of transcription and translation regulative sequences, the corresponding signals from the aspartate aminotransferase gene isolated from P. haloplanktis TAC125 were inserted, generating the pFF vector. To investigate the possibility of obtaining recombinant proteins in this cold-adapted host, we used the psychrophilic alpha amylase from the Antarctic bacterium P. haloplanktis TAB23 (previously known as Alteromonas haloplanktis A23) as a model enzyme to be produced. Our results demonstrate that the cold-adapted enzyme was not only produced but also efficiently secreted by the recombinant PhTAC125 cells. The described expression system represents the first example of heterologous protein production based on a true cold-adapted replicon. PMID- 15269428 TI - Recombinant protein production in yeasts. AB - Recombinant DNA (rDNA) technologies (genetic, protein, and metabolic engineering) allow the production of a wide range of peptides, proteins, and biochemicals from naturally nonproducing cells. This technology, now approx 25 yr old, is becoming one of the most important technologies developed in the 20th century. Pharmaceutical products and industrial enzymes were the first biotech products on the world market made by means of rDNA. Despite important advances regarding rDNA applications in mammalian cells, yeasts still represent attractive hosts for the production of heterologous proteins. In this review we summarize the advantages and limitations of the main and promising yeast hosts. PMID- 15269429 TI - Controlled expression of homologous genes by genomic promoter replacement in the yeast Saccharomyces cerevisiae. AB - Exchange of the promoter of a gene in the genome for another promoter whose expression can be controlled easily can overcome problems associated with the expression of the same gene from a promoter on a plasmid. Some genes are difficult or impossible to clone in plasmid-based vectors and often a stable expression and maintenance of the gene during cell proliferation is desirable. We present a method by which any genomic promoter can be replaced by a promoter of choice to achieve controlled (or constitutive and strong) expression of the gene concerned. The new promoter and a marker gene of choice are amplified by PCR using primers with a tail homologous to the regions adjacent to the site of integration in the genome and primers with a restriction site allowing ligation of the promoter and marker PCR products. After ligation of these PCR products, the ligated construct is transformed into yeast cells and allowed to exchange for the original promoter by homologous recombination. The transformants are selected based on the presence of the marker gene and proper exchange of the original promoter for the new promoter is checked by means of PCR amplification using primers in the new promoter and in the gene under its control. PMID- 15269430 TI - High-throughput expression in microplate format in Saccharomyces cerevisiae. AB - We have developed a high-throughput technology that allows parallel expression, purification, and analysis of large numbers of cloned cDNAs in the yeast Saccharomyces cerevisiae. The technology is based on a vector for intracellular protein expression under control of the inducible CUP1 promoter, where the gene products are fused to specific peptide sequences. These N-terminal and C-terminal epitope tags allow the immunological identification and purification of the gene products independent of the protein produced. By introducing the method of recombinational cloning we avoid time-consuming re-cloning steps and enable the easy switching between different expression vectors and host systems. PMID- 15269431 TI - High-throughput expression in microplate format in Pichia pastoris. AB - The methylotrophic yeast Pichia pastoris has become a powerful host for the heterologous expression of proteins. To serve the increasing demand for clones expressing different cDNAs, we developed a cultivation and induction protocol amenable to automation to increase the number of clones screened for protein expression. Therefore cDNAs are cloned for intracellular expression. The resulting fusion proteins carry affinity tags (6*HIS and StrepII, respectively) at the N- and C-terminus for the immunological detection and chromatographic purification of full-length proteins. Expression is controlled by the tightly regulated and highly inducible alcohol oxidase 1 (AOX1) promoter. The screening procedure is based on a culture volume of 2 mL in a 24-well format. Lysis of the cells occurs via chemical lysis without mechanical disruption. Using the optimized feeding and induction protocol, we are now able to screen for and identify expression clones that produce heterologous protein with a yield of 2 mg per L culture volume or higher. PMID- 15269432 TI - Multiple gene expression by chromosomal integration and CRE-loxP-mediated marker recycling in Saccharomyces cerevisiae. AB - Multiple gene expression can be introduced in a yeast strain with using only two markers by means of the two new vectors described, the expression vector pB3 PGK and the CRE recombinase vector pCRE3. The pB3 PGK has a zeocin-selectable marker flanked by loxP sequences and an expression cassette consisting of the strong PGK1 promoter and the GCY1 terminator. The gene of interest (YFG1) is cloned between the promoter and terminator of pB3 PGK. The pB3 PGK-YFG1 is integrated into the genome by a single restriction cut within the YFG1 gene and integrated in the YFG1 locus. The strain is further transformed with the pCRE3 vector. The CRE recombinase expressed from this vector removes the zeocin marker and makes it possible to use the pB3 PGK vector over again in the same strain after curing of the pCRE3 vector. The 2 micro -based pCRE3 carries the aureobasidin A, zeocin and URA3 markers. pCRE3 is easily cured by growth in nonselective medium without active counterselection. The screening for loss of the chromosomal zeocin marker, as well as curing of the pCRE3 vector, is done in one step, by scoring zeocin sensitivity. This can be done because the zeocin marker is present in both the pB3 PGK and pCRE3. The S. cerevisiae pentose phosphate pathway genes RK11, RPE1, TAL1, and TKL1 were cloned in pB3 PGK and integrated in the locus of the respective gene, resulting in simultaneous overexpression of the genes in the xylose-fermenting S. cerevisiae strain TMB3001. PMID- 15269433 TI - Three decades of fungal transformation: key concepts and applications. AB - Filamentous fungi include a large, heterogeneous group of heterotrophic organisms with profound influence in human activities. In spite of their economic and scientific relevance, little information is available at the molecular level about their biology. The development of genetic transformation protocols has contributed greatly to the molecular dissection of fungal behavior. The use of this approach in combination with large-scale genome sequencing projects has now provided the basis for gaining insight into the function of fungal genes. This chapter reviews the technology of the transformation of filamentous fungi. The protocols for gene transference by protoplasting/PEG, LiAc, electroporation, biolistics and A. tumefaciens are described, and possible mechanisms for transformation are discussed. A brief description of previously reported selection systems is also included. The application of transforming protocols concerning the study of gene function and manipulation are discussed in relation to some fungal species. PMID- 15269434 TI - Three decades of fungal transformation: novel technologies. AB - Fungi are lower eukaryotes that play important roles in many human activities, including biotechnological processes, phytopathology, and biomedical research. In addition, they are excellent models for molecular and genetic studies. An important key in the advancement of genetics and molecular biology of a given organism is the development of genetic transformation systems. This technology makes possible the analysis and manipulation of the genome of the organism of interest. Thirty years from the first report of transformation of a fungus, transformation of many other fungi has been achieved. However, the development of gene tagging systems generally applicable to a wide range of filamentous fungi has remained elusive. A widely used gene tagging strategy for filamentous fungi is restriction enzyme mediated integration (REMI). In recent years numerous reports have been published describing the effective application of REMI. However, REMI shows certain disadvantages for some fungi. Recently a very promising alternative strategy has been reported based on the use of the soil bacterium Agrobacterium tumefaciens. Using this system a well-defined DNA segment (T-DNA) is transferred, which integrates by illegitimate recombination and is 100 1000 times more efficient than conventional methods. The T-DNA can be used as an efficient tool to generate recombinant strains where DNA is integrated as a single copy, allowing the generation of collections of gene-tagged mutants of the fungus of interest. PMID- 15269435 TI - Gene transfer and expression in plants. AB - Until recently, agriculture and plant breeding relied solely on the accumulated experience of generations of farmers and breeders that is, on sexual transfer of genes between plant species. However, recent developments in plant molecular biology and genomics now give us access to knowledge and understanding of plant genomes and the possibility of modifying them. This chapter presents an updated overview of the two most powerful technologies for transferring genetic material (DNA) into plants: Agrobacterium-mediated transformation and microparticle bombardment (biolistics). Some of the topics that are discussed in detail are the main variables controlling the transformation efficiency that can be achieved using each one of these approaches; the advantages and limitations of each methodology; transient versus stable transformation approaches; the potential of some in planta transformation systems; alternatives to developing transgenic plants without selection markers; the availability of diverse genetic tools generated as part of the genome sequencing of different plant species; transgene expression, gene silencing, and their association with regulatory elements; and prospects and ways to possibly overcome some transgene expression difficulties, in particular the use of matrix-attachment regions (MARs). PMID- 15269436 TI - Production of recombinant proteins by hairy roots cultured in plastic sleeve bioreactors. AB - Plant-based expression of recombinant proteins offers significant advantages over transgenic animal-and cell-based systems. Unlike bacteria, plants perform the complex protein-processing steps required to produce eukaryotic proteins in active form. In order to facilitate protein production and purification we used hairy root cultures as a secretion-based in vitro plant system. We utilized the green fluorescent protein (GFP) as our model protein and expressed it for secretion in tobacco hairy root cultures. For large scale production of GFP, we adapted the Life-Reactor trade mark plastic sleeve bioreactor for growth of hairy roots in cultures containing up to 5 L of medium. Yields higher than 800 micro g of GFP per liter of culture were obtained after 21 d of incubation, representing almost 20% of the total secreted protein. The use of the plastic sleeve bioreactor system for expression of proteins in hairy roots allows for continuous or inducible production and recovery, while maintaining absolute containment, of the recombinant product. PMID- 15269437 TI - Engineering the chloroplast genome for hyperexpression of human therapeutic proteins and vaccine antigens. AB - The chloroplast genome is ideal for engineering because it offers a number of attractive advantages, including high-level gene expression, the feasibility of expressing multiple genes or pathways in a single transformation event, and transgene containment due to lack of pollen transmission. The chloroplast-based expression system is suitable for hyperexpression of foreign proteins, oral delivery of vaccine antigens and therapeutic proteins, via both leaves and fruits. Through the refinement of expression vectors and use of chaperones, chloroplasts produce up to 47% of foreign protein in the total cellular protein in transgenic tissues. This chapter describes various techniques for creating chloroplast transgenic plants and their biochemical and molecular characterization. Suitable examples for application of chloroplast genetic engineering in human medicine are provided. PMID- 15269438 TI - New selection marker for plant transformation. AB - A number of systems to insert foreign DNA into a plant genome have been developed so far. However, only a small percentage of transgenic plants are obtained using any of these methods. Stable transgenic plants are selected by co-introduction of a selectable marker gene, which in most cases are genes that confer resistance against antibiotics or herbicides. In this chapter we describe a new method for selection of transgenic plants after transformation. The selection agent used is the nontoxic and common sugar glucose. Wild-type Arabidopsis thaliana plantlets that have been germinated on glucose have small white cotyledons and remain petite because the external sugar switches off the photosynthetic mechanism. The selectable marker gene encodes the essential trehalose-6-phophate synthase, AtTPS1, that catalyzes the first reaction of the two-step trehalose synthesis. Upon ectopic expression of AtTPS1 driven by the 35S promoter, transformed Arabidopsis thaliana plants became insensitive to glucose in comparison to wild type plants. After transformation using AtTPS1 as a selection marker and 6% glucose as selection agent it is possible to single out the green and normal sized transgenic plants amid the nontransformed plantlets. PMID- 15269439 TI - Enhancer detection and gene trapping as tools for functional genomics in plants. AB - Although more than 25,000 genes of Arabidopsis thaliana have been sequenced and mapped, adequate expression or functional information is available for less than 15% of them. In the case of Oryza sativa (rice), about half of more than 55,000 predicted genes have been assigned to a vague functional category on the basis of their sequence, but fewer than 100 have been ascribed a precise, verified function after the identification of a mutant phenotype caused by the molecular disruption of the corresponding gene. Enhancer detection and gene trapping represent insertional mutagenesis strategies that report random expression of many genes and often generate loss-of-function mutations. Several trapping vectors have been designed in a limited number of species, and large-scale enhancer detection and gene trap screens that aim to generate a wide range of spatially and temporally restricted expression patterns have been initiated in both Arabidopsis and rice. These strategies are proving to be essential to the functional annotation of completely sequenced genomes, enabling the analysis of gene function in the context of the entire plant life cycle and substantially expanding our understanding of plant growth and development. PMID- 15269440 TI - Gene transfer and expression in mammalian cell lines and transgenic animals. AB - Manipulation of the eukaryotic genome not only has contributed to the progress in our knowledge of multicellular organisms but has also ameliorated our experimental strategies. Biological questions can now be addressed with more efficiency and reproducibility. There are new and varied strategies for gene transfer with improved methodologies that facilitate the acquisition of results. Cellular systems and transgenic animals have demonstrated their invaluable benefits. This chapter presents an overview of the methods of gene transfer, with particular attention to cultured cell lines. Alternative strategies of gene transfer are also shown and the applications of such methods are discussed. Finally, several comments are made about the influence of chromatin structure on gene expression. Recent experimental data have shown that for convenient stable transgene expression, the influence of chromatin structure should be seriously taken into account. Novel chromatin regulatory and structural elements are proposed as an alternative for proper and sustained gene expression. These chromatin elements are facing a new era in transgenesis and we are probably beginning a new generation of gene and cancer therapy vectors. PMID- 15269441 TI - Sustained heterologous transgene expression in mammalian and avian cell lines. AB - Chromosomal position effects are the cause of variegation of transgene expression relating to the eukaryotic genome environment. These effects modify heterologous transgene expression, presumably due to dominant epigenetic marks that cause variegated or gradual silencing of transgene expression. As a solution, chromosomal insulators have arisen as a way to protect transgene expression against repressive marks (position effects), allowing homogeneous and consistent expression. Sustainable expression enables us to gain insight into in vivo protein function and protein stability, or to study position effects. Here, we show that the chicken cHS4 beta-globin insulator is able to protect transgene expression in cell lines, demonstrating its potential as a tool to improve the heterologous expression of any desired transgene. In addition, we show that green fluorescent protein (GFP) can be used as a reporter protein in the study of chromosomal position effects. PMID- 15269442 TI - Inducible gene expression in mammalian cells and mice. AB - Inducible expression of desired transgenes in mammalian cells and animals is a current priority in basic and applied research, biopharmaceutical manufacturing, gene therapy, and tissue engineering, as well as in drug discovery. Among the most prominent human-compatible transgene control technologies are engineered promoter/transactivator configurations that adjust heterologous target gene transcription in response to clinically licensed antibiotics (tetracyclines, streptogramins, macrolides). In this chapter we provide a detailed case study on macrolide-inducible expression of the human model glycoprotein SEAP (human placental secreted alkaline phosphatase) in transgenic Chinese hamster ovary (CHO) cell cultures or following implantation of microencapsulated CHO cells into mice. PMID- 15269443 TI - Flp-mediated integration of expression cassettes into FRT-tagged chromosomal loci in mammalian cells. AB - The creation of recombinant mammalian cells with defined expression levels requires extensive screening. This is mainly due to the unpredictable site and copy number of the recombinant DNA in the hosts' chromosomal DNA. The method presented here is based on the exchange of expression cassettes in a previously tagged site. Since the site of chromosomal integration remains the same in all targeted cells and since the copy number is reduced to one, the level of expression is highly predictable. The use of this method includes two steps. The first one, the tagging step, makes use of retroviral reporter constructs, which ensure single-copy integration of the tagging vector. Upon screening for cell clones with appropriate expression strength these cells will be targeted. The targeting construct harboring the gene of interest will precisely replace the tagging reporter cassette making use of the Flp recombinase. PMID- 15269444 TI - Generation of high-recombinant- protein-producing Chinese hamster ovary (CHO) cells. AB - The purpose of this chapter is to propose a practical procedure for the generation and selection of Chinese hamster ovary cells producing high levels of recombinant protein by combining in vitro and in vivo amplification of the foreign gene. A detailed description of the expression and amplification plasmids utilized for the in vitro generation of long DNA concatenamers, as well as the cell transfection and selection protocols are given. The procedure required for in vivo gene amplification using the dihydrofolate reductase/methotrexate system is also described. PMID- 15269445 TI - Preparation of recombinant proteins in milk. AB - Using transgenic animals as the source of recombinant proteins has several specific advantages. Large amounts of proteins can be obtained, essentially from milk. These proteins are often properly processed. They are in a number of cases correctly folded, assembled, cleaved, glycosylated, gamma-carboxylated, and so on. Purification of recombinant proteins from milk is not a particularly difficult task. The level of expression of foreign genes in milk cannot be predicted in all cases and appropriate vectors must be used. Generation of transgenic mice is popular but their production is quite limited. Transgenesis in larger animals, rabbits and farm animals, is achieved essentially by a few companies. Some recombinant proteins may be found in blood circulation and alter animal health. Milk from transgenic animals has become a quite attractive alternative to other sources of recombinant proteins. PMID- 15269446 TI - [Alcohol and women: clinical aspects]. AB - Alcohol-related pathologies lead to most serious expressions, both at clinical and social level. The diffused social acceptance of consumption and abuse behavior and the lack of alcohol education for professionals (physicians, psychologists, social workers etc.) make difficult to put in the right frame this issue. Just a multidimensional approach can make the problem understandable. The history of alcohol consumption during the time gives us an exhaustive picture of the negative consequences of alcohol consumption. Alcohol consumption during pregnancy is a problem still underestimated and represents a serious risk for the health of the newborns: children alcohol-exposed in uterus are at risk to develop many pathologies and even the fetal-alcohol syndrome (FAS) that leads to facial anomalies, growth deficiencies and neurological damages. Therefore interventions coping with this kind of issues are needed in order to enhance people's health. PMID- 15269447 TI - [Psychological aspects involved in alcohol consumption]. AB - Cultural traditions, social surroundings, life cycle phases and psychological reasons are considered the factors that determine the use and abuse of alcohol among women. The psychological mechanisms underlying alcohol consumption are viewed from a psychoanalytic point of view. According to the psychoanalytic theory the different meanings that the consumption of alcohol assumes depend on the unconscious conflicts and the personality structure. The hypothesis is that alcohol represents a "transitional object" unconsciously utilised in a paradoxical way since it expresses, contemporarily, a desire and a defence from the transition, be it between oneself and the others, be it between inner aspects of oneself. PMID- 15269448 TI - [Women and alcohol: biological vulnerability]. AB - Alcohol abuse is the major cause of morbidity and mortality in western countries. In the last decade, the rate of women involved in alcohol abuse is increasing. This is particularly dangerous since alcohol abuse causes heavier damages in women than in men. This is due to physiological differences about body composition, enzymatic and hormonal patterns. At now gender studies are not enough and are to be deepened. Obtained results are synthetically described in this paper. Emerging problems are new habits of alcohol abuse, the decreasing age of first use in women and the teratogenic effect of alcohol on fetal development. PMID- 15269449 TI - [Epidemiology of alcohol consumption among women]. AB - In the past 20 years, the lifestyles of the Italian population have changed radically, especially those of women, who have adopted behavioural models traditionally associated with men. One of the most important changes has been the increasing number of women who smoke, and more recently, those who drink, habits that can be considered to reflect the changing role of women in society yet which obviously have an important impact on the health status of the female population. PMID- 15269450 TI - [Women and cross-dependence]. AB - Most scientific researches on drug abuse and dependence suffer from "gender bias" being male dominated with the assumption that men are the main users of drugs. In reality, women use mind-altering drugs in the same way as men. This paper focus on alcohol, drugs use/abuse and dependence amongst females on public treatment Services, on recreational settings as well as at school. Problem related to alcohol and cocaine are discussed the most. Findings on aggressive behavior, psychological aspects, younger age of abuse, traumatic events, patterns of use, subset of gender specific symptoms manifestation (depression, mood swings, paranoia) the vicious circle of anxiety and alcoholism, the cross-dependence on legal and illegal drugs, comorbidity between drug abuse and psychiatric syndromes are treated in relation to academic literature. PMID- 15269451 TI - [Female alcoholic consumption between search for equality and increase of risk: which prevention?]. AB - During the latest 5 years, Italy noticed an increase in the number of consumers, mainly among women and young people, together with a general drop in the consumption of alcohol. Among women, and particularly among young women, the trend toward higher and extra meals consumption is growing, mainly in bars or discos, in accordance with a pattern imported by North Europe countries. We suppose that the success of such a model of consumption among women is linked, for its peculiarities, customs and contexts involved, to the attainment of a status of a greater emancipation and equality with men, which is considered an important value in this historical period. Nowadays women seem to be more exposed to the risk of alcohol-related problems just because of their search for emancipation and equality. In that situation, we need measures of prevention specifically aimed at the female population, informing women about the specific risks and responsibilities connected with the female role in the field of alcohol consumption. In the mean time these measures of prevention must be able to respect the search of equality and emancipation that has been motivating the evolution of female behaviours for the latest years. PMID- 15269452 TI - [Primary prevention for alcohol misuse in young people: a Cochrane Systematic Review]. AB - The Cochrane Collaboration is an international no-profit organization established in 1992 in UK. The aim of the Collaboration is the conduction, update and dissemination of systematic reviews about health care. Systematic reviews are electronic documents systematically updated which synthesise the results of randomized controlled studies about treatments. The Cochrane Group on Drugs and Alcohol has the editorial base in Rome (Department of Epidemiology ASL RME) where the Coordinator, the Coordinating Editor and the Trial Search Coordinator, coordinate the work of seven editors based in several countries. As of April 2003 we published 17 reviews and 11 protocols of review. The systematic reviews on primary prevention for alcohol misuse in young people, was conducted by David Foxcroft and published by the group in 2002. The objectives of the systematic review were the identification and synthesis of the studies on psychosocial and educational programs for prevention of alcohol abuse and the assessment of long term interventions (over three years). PMID- 15269453 TI - Genetic analysis of the diversity in Toxoplasma gondii. AB - More than 50 different genetic markers are now available for Toxoplasma gondii isolate typing. For most of them, allelic polymorphism is low (2 to 4 alleles). A few markers, more polymorphic, can be used to fingerprint isolates for epidemiological studies. Phylogenetic analysis detects 2 main groups in Toxoplasma population. Group 2 is genetically heterogeneous. From a practical point of view, 3 main types (types I, II and III) are usually considered. Multilocus genotyping is necessary to reveal atypical or recombinant genotypes, more frequent among isolates collected in remote areas or in wild game. Toxoplasma isolates from human toxoplasmosis are mainly from type II (whatever the severity of clinical disease), but type I and atypical genotypes are found only in severe cases of human toxoplasmosis. PMID- 15269454 TI - Impact of stage differentiation on diagnosis of toxoplasmosis. AB - Within its intermediate host, such as man, the protozoan parasite Toxoplasma gondii interconverts between tachyzoites and bradyzoites. The replicative tachyzoite stage is thought to be responsible for acute/active infection and expresses immunodominant antigens thereby inducing a strong cellular immune response, which vice versa triggers the differentiation process into dormant and immunologically weak cyst stages. The immunodominance of tachyzoites is also responsible for the induction of a strong humoral immune response leading to the formation of antibodies specifically directed against tachyzoite antigens. In contrast, the bradyzoite stage which is associated with inactive/chronic infection, seems not to be a strong inducer of specific antibodies. However, since the humoral antibody response is also directed against antigens that are expressed in both stages, serodiagnosis cannot always adequately discriminate between active and inactive/chronic infection. This short review focuses on the impact of stage differentiation and discusses the potential of stage-specifically expressed antigens that might be useful in a recombinant form in order to improve future serodiagnostical approaches. PMID- 15269455 TI - Immune response to Toxoplasma gondii. AB - Oral-route infection with Toxoplasma gondii sporozoites or tachyzoites leads to the rapid spread of quick-replicating cytolytic tachyzoites throughout the whole body. Toxoplasma easily crosses the blood-retina, encephalic and placental barriers. The acute phase of this infection lasts for less than around ten days. The parasite causes a very strong type-1 response focused on the interferon-gamma secreted by the T lymphocytes. This immune response limits the tissue extension of the parasite, ensuring the survival of the host, but, paradoxically, also aiding the survival of the parasite by converting it into a bradyzoite, an intracellular quiescent resistant form persisting in the muscle and brain tissues. PMID- 15269456 TI - Serodiagnosis of toxoplasmosis. The impact of measurement of IgG avidity. AB - The development of IgG avidity assays has revolutionised serological diagnosis of Toxoplasma infections. The measurement of IgG avidity has shown its power in various clinical settings, especially in situations where timing and differentiation of primary and secondary infections is crucial. However, no laboratory test performed alone is self-sustained, whereby the diagnostic strategy of choice is sequential (or combinatorial) use of high-quality IgG, IgM, IgA and IgG-avidity assays. The impact of IgG avidity measurement will be discussed in five clinical scenarios: acute acquired infection, primary infection during pregnancy, congenital toxoplasmosis, ocular toxoplasmosis and Toxoplasma infection in immunocompromised patients. All in all in toxoplasmosis, superior diagnostic performance is achieved by appropriate combinations of serological, culture-based and PCR techniques. PMID- 15269457 TI - [Nerve growth factor and multiple sclerosis: studies on animal models and in humans]. AB - We first reported that the level of nerve growth factor (NGF), a pleiotrophic factor produced in central nervous system (CNS) implicated in growth, differentiation and repair of brain neurones, undergoes through significant changes in brain of patients with multiple sclerosis (MS) and of its animal model experimental autoimmune encephalomyelitis (EAE). Recently, much attention has been evolved around these studies, reinforcing the hypothesis about the role of NGF in this disorder. Indeed, current studies indicate that NGF stimulates growth and differentiation of stem cell in EAE, exerts anti-inflammatory action and reduces the severity of EAE in non-human primate, prospecting the clinical potentiality of NGF for MS. However, despite these findings, crucial evidences, such as, the identification and characterisation of the mechanism(s) implicated in tissue repair and in inflammatory responses, needs to be done to evaluate the role of NGF to identify potential therapeutic strategies for this demyelinating disorder. PMID- 15269458 TI - Health effects of exposure to waste incinerator emissions:a review of epidemiological studies. AB - This review evaluates the epidemiological literature on health effects in relation to incineration facilities. Several adverse health effects have been reported. Significant exposure-disease associations are reported by two thirds of the papers focusing on cancer (lung and larynx cancer, non-Hodgkin's lymphoma). Positive associations were found for congenital malformations and residence near incinerators. Exposure to PCB and heavy metals were associated with several health outcomes and in particular with reduction of thyroid hormones. Findings on non-carcinogen pathologies are inconclusive. Effect of biases and confounding factors must be considered in the explanation of findings. Methodological problems and insufficient exposure information generate difficulties on study results. Research needs include a better definition of exposure in qualitative and quantitative terms in particular by developing the use of biomarkers and by implementing environmental measurements. PMID- 15269459 TI - [Microbiological risk associated with consumption of drinking water in developed countries]. AB - This article provides an overview of the waterborne disease (WBD) issue in developed countries (USA, Canada, UK and other European countries). The factors involved in the epidemiology of waterborne infection are analysed (microbial, social, environmental, etc.) and the main WBD etiological agents are described with particular interest to emerging pathogens (i.e. Cryptosporidium parvum, Legionella and Calicivirus). Microorganism characteristics related to water contamination risk are described and examples of waterborne outbreak are reported. A short account about the detection methods of these microorganisms in the water is given. In conclusion, some possible strategies to control waterborne microbiological risk in industrialised countries are discussed considering the application of the risk analysis and the HACCP system in the water production and the need of a WBD surveillance system. PMID- 15269460 TI - Tuberculous meningitis and hydrocephalus. PMID- 15269461 TI - Surgical management of complex intracranial aneurysms. AB - Despite the improvements in increasing popularity of endovascular therapy for intracranial aneurysms, there remain a large number of these lesions that currently are not amenable to endovascular therapy. As endovascular therapy becomes more popular, those aneurysms requiring surgical intervention will become increasingly complex. To manage these challenging lesions, neurosurgeons must use all available innovations and advances, including diagnostic, technical and perioperative adjuncts. In this review article, we discuss limitations of endovascular therapy, the populations of aneurysms that continue to require surgical treatment, the factors that make an aneurysm complex and the multiple adjuncts utilized to successfully treat these challenging lesions. PMID- 15269462 TI - Anesthesia and intracranial arteriovenous malformation. AB - Anesthetic management of intracranial arteriovenous malformation (AVM) poses multiple challenges to the anesthesiologist in view of its complex and poorly understood pathophysiology and multiple modalities for its treatment involving different sub-specialties. The diagnosis of AVM is based on clinical presentation as well as radiological investigation. Pregnant patients with intracranial AVM and neonates with vein of Galen malformation may also pose a special challenge and require close attention. Despite technological advancement, reported morbidity or mortality after AVM treatment remains high and largely depends on age of the patient, recruitment of perforating vessels, its size, location in the brain, history of previous bleed and post-treatment hyperemic complication. Anesthetic management includes a thorough preoperative visit with meticulous planning based on different modalities of treatment including anesthesia for radiological investigation. Proper attention should be directed while transporting the patient for the procedure. Protection of the airway, adequate monitoring, and maintaining neurological and cardiovascular stability, and the patient's immobility during the radiological procedures, appreciation and management of various complications that can occur during and after the procedure and meticulous ICU management is essential. PMID- 15269463 TI - Diagnostic criteria for neurocysticercosis: some modifications are needed for Indian patients. AB - In India and other less developed countries the diagnosis of neurocysticercosis is frequently difficult because several other prevalent neurological disorders can present with a similar clinical and neuroimaging picture. Currently available international criteria seem to be helpful for the diagnosis of neurocysticercosis, however, these criteria have been criticized for not being effective in differentiating several other infective and neoplastic diseases of central nervous system (CNS), like CNS tuberculosis, from neurocysticercosis. In this article, modifications in the recent diagnostic criteria given by Del Brutto et al (2001) are being suggested, so, it can become more suitable for Indian patients. In India the overwhelming majority of patients with neurocysticercosis have either single enhancing or less frequently multiple enhancing CT lesions. Imaging and clinical features of various infective conditions, like tuberculoma, fungal granuloma, and parasitic granuloma, and of neoplastic conditions like cerebral metastasis, are remarkably similar. Keeping this in mind, the modification suggested in this article is to replace epidemiological criteria with the section diagnosis of neurocysticercosis with caution in certain situations. These situations are middle or old age, evidence of pre-existing tuberculosis or malignancy, pre-existing HIV infection and in patients with grossly abnormal neurological examination. In these situations, in the absence of one of the absolute criteria, it should be essential to consider and exclude all other likely possibilities before making a diagnosis of neurocysticercosis. However, because of the high prevalence of several disorders with similar features it is difficult to make reliable diagnostic criteria for neurocysticercosis, which are easy to use, and have a high specificity and sensitivity. PMID- 15269464 TI - Vertebral artery in relationship to C1-C2 vertebrae: an anatomical study. AB - BACKGROUND: Ten randomly selected adult cadaveric specimens were dissected to analyse the anatomy of the vertebral artery during its course from the C3 transverse process to its entry into the spinal dural canal at the level of C1. In addition, 10 dry cadaveric C1-C2 bones were studied. The course of the artery and the parameters relevant during surgery in the region are evaluated. METHODS: Ten adult cadaveric specimens and 10 adult dry cadaveric C1 and C2 bones were studied. In five cadaveric specimens, the arteries and veins were injected with coloured silicon. The artery during its course from the transverse process of C3 to the transverse process of C2 was labelled as V1 segment, the artery during its course from the C2 transverse process to the C1 transverse process was labelled as V2 segment and the segment of the artery after its exit from the transverse foramen of C1 to the point of its dural entry was labelled as V3 segment. The relationship of the artery to the C1-2 joint and facets, distance of the location of the artery from the midline, from the C2 ganglion and from the other surgery related landmarks were evaluated. The extent of occupancy of the artery into the vertebral artery groove on the inferior surface of the superior facet of the C2 vertebra, and over the posterior arch of the atlas was studied. RESULTS: The V1 segment of the vertebral artery takes a varying degree of loop inside the vertebral artery foramen on the inferior aspect of the superior facet of the C2 vertebra. The loop extends towards the midline and was at an average distance of 14.6 mm from the midline of the vertebral body. The V2 segment of the artery takes an initial lateral loop after its exit from the transverse process of the C2 vertebra. The average distance of the artery from the lateral end of the C2 ganglion was 7.2 mm and from the dural tube was 15.3 mm. The vertebral artery groove in the superior facet of C2 and the groove over the posterior arch of the atlas were completely occupied by the vertebral artery only in six sides and in none respectively, suggesting the possibility of the dynamic nature of the relationship of the artery to the bone. CONCLUSIONS: The vertebral artery adopts a serpentine course in relationship to the C2 vertebra, making it susceptible to injury during the surgical procedures in the region. The multiple loops of the artery and a buffer space within the vertebral artery groove on the inferior surface of the superior facet of the C2 vertebra and over the posterior arch of atlas provide the artery an extra length and space, probably essential to avoid any stretch during neck movements. PMID- 15269465 TI - Head up tilt test in the diagnosis of neurocardiogenic syncope in childhood and adolescence. AB - BACKGROUND: Neurocardiogenic syncope (NCS) is a common paroxysmal disorder that is often misdiagnosed as a seizure disorder. Head up tilt test (HUTT) has been used to confirm this diagnosis. There is no data available of its use in children / adolescents from India. AIM: To study the usefulness of the HUTT in children and adolescents with suspected NCS. SETTINGS AND DESIGN: This was a part retrospective and later prospective study set in a tertiary child neurology outpatient department (OPD). MATERIAL AND METHODS: Patients with a strong clinical suspicion of syncope were recruited for the study. Clinical and treatment details were either retrieved from the chart or prospectively recorded in later patients. The HUTT was then carried out at baseline and after provocation and the results correlated with the clinical diagnosis. RESULTS: Eighteen children with a mean age of 10.8 years were studied. Eight had precipitating factors. Thirteen had premonitory symptoms. Pallor, temperature change, diaphoresis, headache, tonic / clonic movements, post-ictal confusion and peri-ictal headache were symptoms noticed. Sixteen had a positive HUTT. Seven were on long-term anti-epileptic drugs (AEDs). Two had epileptiform abnormalities on their electroencephalogram (EEG). CONCLUSION: The diagnosis of syncope is often confused with epilepsy. Head up tilt test has a high sensitivity in the diagnosis of NCS in children / adolescents. It is fairly safe and easy to perform. PMID- 15269466 TI - Lumboperitoneal shunts: review of 409 cases. AB - BACKGROUND AND AIMS: A prospective study was carried out to evaluate the lumboperitoneal shunt procedure. MATERIAL AND METHODS: Four hundred and nine patients having communicating hydrocephalus were selected for the procedure during a 10-year period from March 1992 to February 2002. The average follow-up was 45.34 months. RESULTS: Tubercular meningitis (TBM)-related hydrocephalus was detected in 285 patients. Forty per cent of the patients were less than 15 years of age. Glasgow Coma Scale (GCS) of less than 8 was seen in 40% patients and 14.9% patients were in GCS 13-15. At the time of discharge 56.7% patients improved in their GCS to 13 -15 and 14.9% were in GCS 8 or less. The overall mortality was 5.13% and shunt-related mortality was seen in 2% patients. Shunt malfunction requiring revision was seen in 32 patients (7.8%) and the total number of shunt revisions was 44 (11%). Shunt infection was noted in 3.4% patients. CSF leak at the lumbar end occurred in 12 patients. Four patients required conversion of LP shunt to VP shunt. CONCLUSIONS: Lumboperitoneal shunt is an effective shunting procedure in communicating hydrocephalus. PMID- 15269467 TI - Prescribing pattern of antiedema therapy in stroke by neurologists and general physicians. AB - BACKGROUND: In acute stroke, a number of drugs are used to reduce the raised intracranial pressure (ICP) although their scientific basis has not been established or shown in randomized controlled trials. AIMS: In this communication, we report the pattern of use of antiedema therapy in acute stroke by general physicians (GPs) and neurophysicians (NPs) in India. MATERIAL AND METHODS: A questionnaire was developed regarding the use of various antiedema measures in stroke and responses were collected either through post or when the responders were attending a national conference. The use of antiedema therapy by NPs and GPs was analyzed employing the Chi-square test. RESULTS: We could collect responses from 102 physicians, of whom 48 were NPs and 54 GPs. More than two thirds of the physicians managed more than three strokes per week and all used antiedema therapy at some time or the other. Thirteen used it in all the patients and the remaining used it in patients with large and moderate strokes or in patients with herniation. Twelve used only one drug, while the remaining physicians used various combinations in different doses and frequency. The prescribing pattern was significantly different between GPs and NPs with respect to the frequency of the antiedema drugs used, type of stroke where these were used, combination of drugs, timing and dose of mannitol. CONCLUSION: This study highlights that antiedema therapy in acute stroke is practiced without any uniformity. PMID- 15269468 TI - Tuberculous meningitis with pulmonary miliary tuberculosis: a clinicoradiological study. AB - BACKGROUND AND AIMS: This study aims at evaluating the clinical and radiological outcome of tuberculous meningitis (TBM) patients with pulmonary miliary tuberculosis. MATERIAL AND METHODS: Diagnosis of TBM was based on clinical, CT scan or MRI and CSF criteria, and that of miliary tuberculosis on chest radiograph. Detailed clinical evaluation was done in all. Severity of meningitis was graded into Grades I, II and III. Complete hemogram, serum chemistry and Montaux tests were performed. The recovery was defined on the basis of 6 months Barthel index score as poor, partial or complete. RESULTS: 20 out of 165 patients with TBM had pulmonary miliary tuberculosis. Their mean age was 30 years; there was one child and 13 patients were females. The mean duration of symptoms was 6.3 months. Montoux test was negative in 9 patients. Six patients were in stage I, 3 in stage II, and 11 in stage III meningitis. Hemoglobin was below 12 gm% in 13 and liver dysfunction and hypocalcaemia was present in 8 and 18 patients respectively. CT scan was abnormal in 16 patients and revealed hydrocephalus (10), granuloma (7), exudates (3) and infarction (1). MRI was abnormal in 7 out of 8 patients and 3 of these patients had normal CT scan. MRI revealed multiple granuloma in 7 patients and exudates in 2. At 6 months, 2 patients died, 10 had complete, 2 had partial and 4 had poor recovery. CONCLUSION: TBM with pulmonary miliary tuberculosis was commoner amongst females who were anemic and hypocalcaemic. MRI revealed multiple granuloma and the majority of the patients improved. PMID- 15269469 TI - Epilepsy with focal cerebral calcification: role of magnetization transfer MR imaging. AB - BACKGROUND: Some patients with focal cerebral calcification (FCC) have no seizure or a benign course of epilepsy, whilst others with a similar lesion have uncontrolled epilepsy. AIMS: To look for perilesional hyperintensity, presumed to be indicative of gliosis, around FCC on magnetization transfer (MT) MRI and to correlate seizure outcome with its presence. SETTING AND DESIGN: Case control study. MATERIAL AND METHODS: Fifty-one patients with epilepsy and 30 controls with single calcified cerebral lesion on CT were studied. Clinical and treatment details were noted. EEG and T1, T2, MT and contrast enhanced MRI were done. STATISTICAL ANALYSIS USED: Student's t test. RESULTS: On MT MRI, perilesional gliosis was seen around the focal calcified lesion in 17 (33.3%) patients. None of the controls had perilesional gliosis. The mean monthly seizure frequency was significantly higher in the 17 patients having perilesional gliosis (2.63+1.15) as compared to the 34 patients without it (0.59+0.63; P= 0.0014). Perilesional gliosis was seen in 8 out of 11 (72.7%) patients who were on 2 AEDs and in all 5 (100%) patients who were on 3 or more AEDs. It was present only in 4 (11.4%) out of 35 patients who were on one AED. CONCLUSION: Gliosis around a cerebral calcified lesion as seen on T1 weighted MT MRI indicates poor seizure control. PMID- 15269470 TI - A clinical study of non-parkinsonian and non-cerebellar tremor at a specialty movement disorders clinic. AB - BACKGROUND: No Indian studies have focused on the clinical aspects of tremor. AIMS: To study the distribution of various etiological types of tremor disorders at a Movement Disorders clinic of a large, tertiary care hospital in India and to study the clinical characteristics of essential tremor [ET]. SETTING AND DESIGN: Prospective cross-sectional study at a tertiary care specialty clinic. MATERIAL AND METHODS: Patients presenting with tremor as the chief complaint, with no features suggestive of parkinsonism, cerebellar disorder or acute central nervous system disorder, were included. Patients were classified into different etiological categories from detailed history. All patients diagnosed as ET, were further interrogated for a detailed family history and examined for characteristics of tremor. These patients were then classified into 'definite', 'probable' and 'possible' ET. RESULTS: One hundred and six patients (mean age 44.4 + 15.1 years) were examined during the study period. ET (59.4%) and dystonic tremor (21.7%) were the commonest types. Only 43% patients of ET reported progression; response to alcohol was seen in only a single patient, a positive family history was present in 52.4% and in 36.4% the inheritance was of an autosomal dominant pattern. CONCLUSION: ET and dystonic tremor are the commonest causes of tremor presenting to a specialty Movement Disorders clinic. Most patients with ET have high-frequency tremor, with mild asymmetry in 40% cases. Alcohol responsiveness may not be a useful tool in the diagnosis of ET. PMID- 15269471 TI - Valley sign in Becker muscular dystrophy and outliers of Duchenne and Becker muscular dystrophy. AB - Valley sign has been described in patients with Duchenne muscular dystrophy (DMD). As there are genetic and clinical similarities between DMD and Becker muscular dystrophy (BMD), this clinical sign is evaluated in this study in BMD and DMD/BMD outliers. To evaluate the sign, 28 patients with Becker muscular dystrophy (BMD), 8 DMD/BMD outliers and 44 age-matched male controls with other neuromuscular diseases were studied. The sign was examined after asking patients to abduct their arms to about 90 cent with hands directed upwards; the muscle bulk over the back of the shoulders was observed. The sign was considered positive if the infraspinatus and deltoid muscles were enlarged and between these two muscles, the muscles forming the posterior axillary fold were wasted as if there were a valley between the two mounts. Twenty-five BMD patients and 7 DMD/BMD outliers had positive valley sign. However, it was less remarkable in comparison to DMD. It was absent in all the 44 controls. It was concluded that the presence of valley sign may help in differentiating BMD from other progressive neuromuscular disorders of that age group. PMID- 15269472 TI - Possible relationship between phenylthiocarbamide taste sensitivity and epilepsy. AB - The study was based on the data of a sample of 400 epileptic patients (200 idiopathic and 200 symptomatic) and 100 normal healthy individuals serving as controls. The PTC threshold distribution was bimodal. The number of non-tasters among idiopathic epileptics (35.5%) and symptomatic epileptics (32.5%) was significantly higher than controls (20%). The relative incidence of non-tasters in idiopathic and symptomatic epilepsies was 2.20 and 1.93 respectively. There is evidence that non-tasters tend to ingest a greater quantity of bitter tasting goitrogenic substances present naturally in edible plants which in turn exert greater thyroid stress in non-tasters or less sensitive tasters. Such a stress during intrauterine or early childhood growth and development might have affected neurological maturation which in turn made them more susceptible to epilepsy than tasters, who faced lesser stress. PMID- 15269473 TI - Spectrum of epilepsy in tuberous sclerosis. AB - Tuberous sclerosis (TS) is an autosomal dominant disease that affects the brain, skin, eye, heart and kidney. The diagnostic criteria for tuberous sclerosis complex (TSC) have recently been revised. There are relatively few Indian studies on this disorder. Twenty-six patients diagnosed as having TS over a period of 18 years are being reported. The onset of seizures ranged from infancy to adolescence. The patterns of epilepsy encountered were generalized tonic clonic seizures (13), complex partial seizures (10), simple partial seizures (9) and myoclonic jerks (4) including infantile spasms (3). Patients often had more than one seizure type. Nineteen patients were mentally subnormal. Cutaneous manifestations were facial angiofibroma i.e. adenoma sebaceum (20), shagreen patches (7), hypopigmented macules (6), ash leaf spots (4), cafe-au-lait spots (2), facial hypoplasia (2) and periungual fibromas (1). One patient each had retinal phakoma and renal angiomyolipoma. CT scan revealed sub-ependymal calcifications (12), parenchymal tubers (3), cerebral edema (3) and cortical atrophy (1). One patient had enhancement of peri-ventricular sub-ependymal lesions on MRI. Anticonvulsants prescribed were phenobarbitone (20), diphenyl hydantoin (14), carbamazepine (8), sodium valproate (4), benzodiazepines (4), ACTH (2), prednisone (1), mysoline (1) and vigabatrin (1). Most patients were on combinations of anti-convulsants and response to therapy was usually not very satisfactory. However, the child treated with vigabatrin did well. PMID- 15269474 TI - Some observations on the spectrum of dementia. AB - A study was designed to generate epidemiological and clinical data on dementia, in a teaching hospital in India. It was conducted on 124 (94 male and 30 female) elderly patients (aged more than 60 years) presenting with clinical syndrome of dementia (DSM-3). Their age range was 64-78 (mean 65.7 4.1) years. Detailed clinical, biochemical, radiological and electrophysiological evaluation was done to establish etiology. Patients with psychiatric ailments, cranial trauma and tumors were excluded. The study period was 4.2 years. Multi-infarct dementia (MID) was observed to be commonest cause of dementia and was present in 59 (47.6%) cases. There were 10 (8%) patients each of tuberculosis (TB) and neurocysticercosis (NCC). Alcohol-related dementia was present in 13 (10.5%), while malnutrition (Vitamin B12 deficiency) was present in 9 (7.2%). Alzheimer's Disease (AD) was present (NINCDS-ADRDA criteria) in 6 patients (4.8%). There were 3 (2.4%) cases 1 each of Huntington's disease, Parkinson's and Normal Pressure Hydrocephalus and 2 each of diabetes, hypothyroidism, hyperthyroidism and Creutzfeldt' Jakob Disease. We conclude that AD, which is irreversible and common in the west, is relatively uncommon in India as compared to MID, infections and malnutrition, which are potentially treatable. PMID- 15269475 TI - C3-4 level cervical spondylotic myelopathy. AB - Cervical spondylotic myelopathy (CSM) is uncommon at the C3-4 level. Fourteen patients with C3-4 CSM were treated over a period of 3 years. The radiological factors contributing to CSM at the C3-4 level were studied. These factors included the assessment of static and dynamic canal diameters, retrolisthesis, posterior osteophytes and degenerative spinal segmental fusion on plain X-rays; and, the antero-posterior cord compression ratio (APCR) on magnetic resonance imaging (MRI). The clinical status of the patients was assessed by the modified Japanese orthopedic association scale (mJOAS). The mean difference between the static and dynamic canal diameters was statistically significant at C3-4 (p < 0.01). The APCR obtained at different levels showed a significant compression at the C3-4 level in comparison to the lower level. There was a correlation between the APCR and the preop mJOAS, r=0.6 (p< 0.05). The mean mJOAS improved from 9.35 to 14.35 at follo-up. The recovery rate calculated using the modified Hirabayashi rate was 66.9%. Degenerative changes at lower cervical segments predispose to increased mobility and spondylotic changes at the C3-4 level. The patients in this study were young as compared to those reported in the international literature. PMID- 15269476 TI - Predictive value of routine hematological and biochemical parameters on 30-day fatality in acute stroke. AB - OBJECTIVE: This prospective study was planned to study the prognostic value of routine clinical, hematological and biochemical parameters, including platelet aggregation in patients of acute stroke, on fatality occurring during the first 30 days. MATERIAL AND METHODS: In this study 116 consecutive patients (77 males and 39 females) of stroke (within 72 hours of onset) were included. After clinical evaluation and neuroimaging, blood investigations, hemoglobin, total leukocyte count, platelet count, platelet aggregation, erythrocyte sedimentation rate (ESR), blood sugar, urea, creatinine, sodium, potassium, serum cholesterol, serum bilirubin, aspartate aminotransferase (SGOT), alanine aminotransferase (SGPT), albumin, and globulin estimations were performed. The patients were followed up for a maximum period of 30 days from the onset of stroke, and patients who expired were grouped as 'expired' and the rest as 'survivors'. Logistic regression analysis was carried out among the significant parameters to identify independent predictors of 30-day fatality. RESULTS: Univariate analysis demonstrated that among hematological parameters, high total leukocyte count and ESR, at admission, correlated significantly with an undesirable outcome during the initial 30 days. Among biochemical parameters, elevated urea, creatinine, serum transaminases (SGOT and SGPT) and globulin levels correlated significantly with death. Logistic regression analysis demonstrated that a low Glasgow Coma Scale (GCS) score along with biochemical parameters such as high serum creatinine, SGPT, ESR and total leukocyte count correlated with death. CONCLUSION: Impaired consciousness, high total leukocyte count, raised ESR, elevated creatinine and SGPT levels, estimated within 24 hours of hospitalization, are the most important indicators of 30-day mortality in patients with first-time ischemic stroke. PMID- 15269477 TI - Increased depressant effect of phenytoin sodium as compared to carbamazepine on cortical excitability: a transcranial magnetic evaluation. AB - To evaluate the effect of monotherapy (phenytoin sodium (DPH) and carbamazepine (CBZ) on the threshold intensity (TI), cortical latency (CL), central conduction time (CCT), using transcranial magnetic stimulation (TMS). A single pulse transcranial magnetic stimulation was used for recording the motor-evoked potentials (MEP) from the thenar muscles of both hands, in 36 patients with well controlled epilepsy on monotherapy, with normal EEG and imaging studies. The TI, CL, CCT and the MEP amplitude were recorded and compared with 20 healthy controls. The threshold intensity was significantly higher in patients on DPH, (P< 0.05) with a significant decrease in the MEP amplitude when compared with controls (P< 0.05). Anticonvulsants alter the excitability of human motor pathways in epileptic subjects. This effect differs among the drugs used; DPH had a greater depressant effect on the excitability than CBZ in the present study. PMID- 15269478 TI - p53 protein alterations in adult astrocytic tumors and oligodendrogliomas. AB - BACKGROUND: p53 is a tumor suppressor gene implicated in the genesis of a variety of malignancies including brain tumors. Overexpression of the p53 protein is often used as a surrogate indicator of alterations in the p53 gene. AIMS: In this study, data is presented on p53 protein expression in adult cases (>15 years of age) of astrocytic (n=152) and oligodendroglial (n=28) tumors of all grades. Of the astrocytic tumors, 86% were supratentorial in location while remaining 14% were located infratentorially - 8 in the the cerebellum and 13 in the brainstem. All the oligodendrogliomas were supratentorial. MATERIALS AND METHODS: p53 protein expression was evaluated on formalin-fixed paraffin-embedded sections using streptavidin biotin immunoperoxidase technique after high temperature antigen retrieval. RESULTS: Overall 52% of supratentorial astrocytic tumors showed p53 immunopositivity with no correlation to the histological grade. Thus, 58.8% of diffuse astrocytomas (WHO Grade II), 53.8% of anaplastic astrocytomas (WHO Grade III) and 50% of glioblastomas (WHO Grade IV) were p53 protein positive. In contrast, all the infratentorial tumors were p53 negative except for one brainstem glioblastoma. Similarly, pilocytic astrocytomas were uniformly p53 negative irrespective of the location. Among oligodendroglial tumors, the overall frequency of p53 immunopositivity was lower (only 28%), though a trend of positive correlation with the tumor grade was noted - 25% in Grade II and 31.5% in grade III (anaplastic oligodendroglioma). Interestingly, p53 labeling index (p53 LI) did not correlate with the histopathological grade in both astrocytic and oligodendroglial tumors. CONCLUSIONS: Thus, this study gives an insight into the genetic and hence biological heterogeneity of gliomas, not only between astrocytic tumors vs. oligodendrogliomas but also within astrocytic tumors with regard to their grade and location. With p53 gene therapy trials in progress, this will possibly have future therapeutic implications. PMID- 15269479 TI - Challenging epilepsy with antiepileptic pharmacotherapy in a tertiary teaching hospital in Sri Lanka. AB - The goal of antiepileptic drug (AED) therapy is to achieve a seizure-free state and eliminate the medical and psychosocial risks of recurrent seizures. Burden of epilepsy on the economy of a country may be largely due to the expenditure on AEDs. The adverse effects may influence the compliance to AEDs and effective control of epilepsy. We determined the pattern of AED use, the degree of epileptic control achieved and the adverse effects experienced by the epileptics in a Tertiary Teaching Hospital in Sri Lanka. Carbamazepine was found to be the most frequently used AED. Monotherapy was used on 70.8% of subjects. 86.27% of the study sample had achieved effective control of epilepsy with a 50% or more reduction in seizure frequency. Of them 72.64% were on monotherapy and they were either on carbamazepine, sodium valproate, phenytoin sodium or phenobarbitone. None of the new AEDs were prescribed to these patients. 50.9% on monotherapy and 51.5% on polytherapy reported adverse effects. Somnolence followed by headache was found to be the most frequently reported adverse effects by those on monotherapy and polytherapy both. This study shows that most epileptics can be effectively managed with the conventional AEDs with clinical monitoring. PMID- 15269480 TI - Role of magnetic resonance perfusion studies in moyamoya disease. AB - Moyamoya disease, Japanese for 'puff of smoke', is a rare disease that presents most commonly with recurrent TIAs (transient ischemic attacks) / stroke in childhood. Ischemic symptoms in patients with moyamoya disease are usually due to hemodynamically-mediated perfusion failure. Identification of abnormal tissue perfusion is an important aspect of the evaluation of these patients. We present the radiological features including the Magnetic Resonance (MR) Perfusion findings illustrating the hemodynamic changes of cerebral ischemia in a case of moyamoya disease. PMID- 15269481 TI - Amyloidoma of the craniovertebral junction. AB - We report a rare case of localized amyloidoma of the craniovertebral junction causing severe myelopathy and respiratory distress and death. The clinical features and the natural history of this rare condition are discussed. PMID- 15269482 TI - Cauda equina paraganglioma presenting with intracranial hypertension: case report and review of the literature. AB - An unusual case of intradural paragangliomas in the cauda equina region in a 29 year-old male is presented. The patient presented with signs and symptoms of raised intracranial pressure. The symptoms resolved after tumor resection. PMID- 15269483 TI - Hashimoto's encephalopathy: response to plasma exchange. AB - We report a case of 52-year-old female with steroid-unresponsive Hashimoto's encephalopathy. She underwent plasma exchange that resulted in marked clinical improvement. PMID- 15269484 TI - Intracranial subfrontal schwannoma treated with surgery and 3D conformal radiotherapy. AB - Subfrontal schwannoma not arising from the cranial nerves are rare tumors. A 19 year-old man presented with a large subfrontal extra-axial enhancing mass with a preoperative diagnosis of skull base meningioma. A subtotal resection of the tumor mass was carried out. Microscopic examination revealed it to be a schwannoma. The residual tumor was treated with fractionated three-dimensional conformal radiotherapy (3D CRT). The rationale of treating with radiotherapy in such cases is analyzed. PMID- 15269485 TI - Subarachnoid spread of germinoma mimicking tuberculous meningitis. AB - A case of pineal germinaoma spreading along the basal subarachnoid space, clinically and macroscopically at autopsy, resembling tuberculous basal arachnoiditis is reported. Need to carry out CSF cytology of even hemorrhagic CSF is stressed, to diagnose the condition. PMID- 15269486 TI - Congenital fiber type disproportion: a rare type of congenital myopathy: a report of four cases. AB - Congenital fiber type disproportion is a rare type of congenital myopathy which presents as hypotonia, delayed motor milestones and dysmorphic facies. During the past 2 years we received 449 muscle biopsies, of which 4 cases were diagnosed as congenital fiber type disproportion (CFTD). In addition to CFTD, one case also had centronuclear features. Three of them were females and one was a male child. Although rare, it should be considered in the differential diagnosis of childhood muscle diseases. Histochemical staining is necessary for the diagnosis of this entity. PMID- 15269487 TI - Alveolar soft-part sarcoma presenting with multiple intracranial metastases. AB - A 28-year-old man presented with history of raised intracranial pressure and one episode of generalized tonic clonic seizures. Computed Tomogram revealed multiple contrast enhancing intracranial lesions. Biopsy of one of the lesions was reported as metastatic alveolar soft part sarcoma. He was advised whole brain radiotherapy. PMID- 15269488 TI - Giant intraparenchymal neurocysticercosis: unusual MRI findings. AB - We report a case of surgically proven giant neurocysticercosis (NCC). MR imaging revealed an unusually large solitary parenchymal cystic lesion showing signal intensity similar to CSF on all pulse sequences, with internal septations and a small nodule in the anterior aspect of this lesion compatible with this diagnosis. Identification of a scolex in a cystic lesion with CSF intensity plays a key role in the diagnosis of NCC. The presence of internal septations is an atypical feature. PMID- 15269489 TI - Cystic olfactory groove schwannoma. AB - Intracranial schwannoma not related to cranial nerves are unusual and rarely found in the subfrontal region. We report a case of cystic olfactory groove schwannoma in a 55-year- old male, who presented with late onset seizure without raised intracranial pressure. The tumor was excised completely. PMID- 15269490 TI - Intracranial plasma cell granuloma. AB - We report two rare cases of primary intracranial plasma cell granuloma. The tumors probably arose from the dura and involved the cerebral parenchyma. These patients presented with clinical features of raised intracranial pressure and there was focal neurological deficit. The management issues are discussed. PMID- 15269491 TI - Intramedullary cysticercosis. AB - A 42-year-old soldier, a known case of cerebral parenchymal neurocysticercosis presented with insidious onset gradually progressive weakness of both lower limbs for six months. Investigations revealed an intramedullary cyst in the cervicodorsal region. Following surgical excision of an intramedullary cysticercus cyst, the patient showed recovery in his neurological deficits. PMID- 15269492 TI - New solitary cysticercus granulomas causing recurrent symptoms in patients with resolved solitary granulomas. AB - Recurrence of symptoms in a patient with a resolved solitary cerebral cysticercus granuloma (SCCG) is uncommon. Recurrent seizures in these patients are generally attributed to an epileptogenic scar or calcific residue of the granuloma. We report two patients with recurrent seizures and one patient with headache; all three patients were previously diagnosed to have SCCG and had complete resolution of the granuloma on follow-up imaging. Computed tomography (CT) at the time of recurrent symptoms showed a SCCG at a site different from the initial lesion, but in the same cerebral hemisphere in all the three patients. Since a new lesion can cause recurrent symptoms in patients with a resolved SCCG, repeat imaging should be performed in all these patients. We also postulate that recurrent cysticercal lesions in patients who have previously had a SCCG, tend to be solitary. PMID- 15269493 TI - Fenofibrate-induced myopathy. AB - Fenofibrate induced myopathy is a rare adverse event. We present a case of muscle pain and quadriparesis following administration of 200mg of fenofibrate for 35 days. Patient gradually improved after stopping the drug. As per our knowledge, this is probably the first case report of fenofibrate induced myopathy from India. PMID- 15269494 TI - Differentiating paralytic rabies from post antirabies vaccine polyradiculoneuropathy. PMID- 15269496 TI - Satoyoshi syndrome: comments. PMID- 15269498 TI - Neurosurgical training in India at the crossroads? PMID- 15269499 TI - Non-traumatic atlantoaxial rotatory subluxation. PMID- 15269500 TI - Nadroparin in acute ischemic stroke. PMID- 15269501 TI - Olanzapine for delirium in parkinsonism: therapeutic benefits in lieu of adverse consequences. PMID- 15269502 TI - Recurrent cerebral venous and peripheral arterial thrombosis. PMID- 15269503 TI - Cervical dystonia responsive to levodopa. PMID- 15269504 TI - Calvarial tuberculosis. PMID- 15269505 TI - Vertebrobasilar dolichoectasia presenting as lower cranial nerve palsy. PMID- 15269506 TI - Uncommon manifestations of neurosarcoidosis. PMID- 15269507 TI - Trigger autoimmunity -development of multiple plexopathy in a patient with chronic idiopathic thrombocytopenic purpura. PMID- 15269508 TI - Multiple sclerosis in a patient with chronic ulcerative colitis. PMID- 15269509 TI - Acute inflammatory demyelinating polyneuropathy in patients with pregnancy. PMID- 15269510 TI - Death following ventricular cerebrospinal fluid shunting in supratentorial malignant tumor associated with hydrocephalus. PMID- 15269511 TI - Isolated foot weakness caused by a parasagittal metastatic parotid adenocarcinoma. PMID- 15269512 TI - Scimitar sacrum. PMID- 15269514 TI - Health in relation to unemployment and sick leave among immigrants in Sweden from a gender perspective. AB - The purpose of this study was to analyze health in relation to unemployment and sick leave among immigrants from a gender perspective. Questionnaire, observations, and group discussions were used for data collection. The study group consisted of 60 unemployed persons with immigrant or refugee background, 30 women and 30 men. Slightly more than half of the participants considered their health to be poor and experienced physical and/or mental disorders. The female participants in comparison to male participants experienced poorer health. The results show that there is a reciprocal influence between health, work, and migration. Immigration may cause poor health, which as a selection effect leads to unemployment and/or sick leave. Immigration may also bring about an inferior position in the labor market, which leads to poor health due to exposure effects. The influence on health is more marked for immigrant women than for immigrant men. PMID- 15269515 TI - Predictors of clinical breast examination among South Asian immigrant women. AB - To determine predictors of clinical breast examination (CBE) among South Asian immigrant women residing in Toronto, Canada. A cross-sectional self-administered survey with women patients visiting family physician group practices. Fifty-four women participated in the study (response rate 77%). Twenty women (38.5%) "ever had" CBE. Compared to women who never had CBE, women who had CBE were statistically older, had lived more years in Canada, had better knowledge of breast cancer, had lower perceived barriers to CBE, and were more likely to have ever had a periodic health exam. No significant differences were found between the two groups for education, employment, English language abilities, perceived health, and perceived benefits of CBE. A direct logistic regression with five predictor variables, significant at a univariate level, was statistically reliable, chi(2) (5, n=51) = 34.7, p < 0.001 and explained 67% of the variance in the CBE status. Age and perceived barriers to CBE remained significant over and above other predictor variables. The odds of "ever had" CBE increased with age and decreased with more perceived barriers. The study highlights the need for education interventions on breast cancer and screening among SA recent immigrant women. PMID- 15269516 TI - Development and testing of interview questions to determine last menstrual period in Mexican immigrant populations. AB - The last menstrual period is used to estimate gestational age. This paper examines sources of measurement error related to the recall of last menstrual period among Mexican immigrant women living in the United States. Qualitative analyses (focus groups and cognitive interviews) suggest that last menstrual period recall does not seem to be a large source of measurement error in the calculation of gestational age and the impact of this type of error on the misclassification of preterm births appears to be minimal. Questions for querying about last menstrual period in this population are offered. PMID- 15269518 TI - Expression of a human lactoferrin N-lobe in Nicotiana benthmiana with potato virus X-based agroinfection. AB - A DNA fragment encoding the N-terminal half (N-lobe) of the human lactoferrin (hLfN) gene has now been cloned into recombinant Potexvirus potato virus X (PVX) vector and expressed in Nicotiana benthmiana using agroinfection. Western blot analysis showed the recombinant protein with an apparent molecular mass on electrophoresis of ca. 40 kDa, corresponding to the predicted size of the hLfN. The yield of hLfN reached a maximum (up to 0.6% of total soluble proteins) when recombinant PVX systemically infected an entire plant. Protein extracts from infected plants had antibacterial activity. PMID- 15269517 TI - The health of the California region bordering Mexico. AB - Healthy Border (HB) 2010 is the health promotion and disease prevention agenda through the year 2010 of the United States-Mexico Border Health Commission (BHC). On the United States side, it draws from the Healthy People (HP) 2010 objectives, identifying those most important and relevant for the border. The BHC has harmonized the list of objectives from both countries into a set of 19 that will be monitored and addressed in a collaborative manner. HB provides a framework for describing the border region's health and comparing with others. For this report, available data were collected for the HB indicators for San Diego and Imperial counties, and for California. Data on Latino populations were considered a proxy for Mexican-Americans and people of Mexican origin in California, because more specific data are not available. Results are presented on the 14 indicators for which the data were most complete. Those of most concern include access to health care and tuberculosis in both counties, plus motor vehicle crash injury deaths and asthma hospitalizations in Imperial. These issues should be given priority attention. Conversely, the region's and Latinos' experience with breast cancer mortality and infant mortality is favorable. Recommendations include binational collaborations in assessing and improving the health of our border communities. PMID- 15269519 TI - Cloning, characterization and expression of the polypeptide release factor gene, eRF1, of Blepharisma japonicum. AB - The polypeptide release factor gene, eRF1, of Blepharisma japonicum (Bj-eRF1) was cloned and sequenced. Its coding region was 1314 base pairs and encodes a protein of 437 amino acids. The cloned gene was expressed in Escherichia coli and the recombinant Bj-eRF1 polypeptide was purified by Ni2+-nitrilotriacetic acid agarose and Superose12 chromatography. Pull-down analysis showed that the recombinant Bj-eRF1 interacts with the heterologously-expressed release factor, eRF3C, of Euplotes octocarinatus. PMID- 15269520 TI - Enantioselective hydrolysis of insoluble (R,S)-ketoprofen ethyl ester in dispersed aqueous reaction system induced by chiral cyclodextrin. AB - The enantioselective hydrolysis of insoluble (R,S)-ketoprofen ethyl ester to the optically active (S)-ketoprofen was carried out in a dispersed aqueous lipase reaction system induced by the inclusion of chiral cyclodextrins for complexation of the substrate. Hydroxypropyl-beta-cyclodextrin was the most effective chiral selector and disperser giving an enantiomeric excess and conversion yield of 0.99 and 0.49, respectively. PMID- 15269521 TI - Self-rotation of red blood cells in optical tweezers: prospects for high throughput malaria diagnosis. AB - A simple and sensitive approach for detection of malarial parasite in blood samples is demonstrated. The approach exploits our finding that, in hypertonic buffer, a normal red blood cell (RBC) rotates by itself when trapped by an optical tweezers. The rotational speed increases linearly at lower trap-beam powers and more rapidly at higher powers. In contrast, under the same experimental conditions, RBC having a malarial parasite does not rotate. The rotational speeds of other RBCs from malaria-infected sample are of an order of magnitude less than that for normal RBC and also increase much more slowly with an increase in trap beam power than that for normal RBC. The difference in rotational speeds could be exploited for the diagnosis of malaria. PMID- 15269522 TI - Cloning and over-expression of an alkaline protease from Bacillus licheniformis. AB - The alkaline protease gene, apr, from Bacillus licheniformis 2709 was cloned into a Bacillus shuttle expression vector, pHL, to yield the recombinant plasmid pHL apr. The pHL-apr was expressed in Bacillus subtilis WB600, yielding a high expression strain BW-016. The amount of alkaline protease produced in the recombinant increased by 65% relative to the original strain. SDS-PAGE analysis indicated a Mr of 30.5 kDa. The amino acid sequence deduced from the DNA sequence analysis revealed a 98% identity to that of Bacillus licheniformis 6816. PMID- 15269523 TI - High-level production of soluble tumor necrosis factor-related apoptosis-inducing ligand (Apo2L/TRAIL) in high-density cultivation of recombinant Escherichia coli using a combined feeding strategy. AB - Recombinant Escherichia coli strain C600/pBV-TRAIL (encoding for 114-281 amino acids of TRAIL's soluble fragment) produced a recombinant human tumor necrosis factor-related apoptosis-inducing ligand (Apo2L/TRAIL). Using a combined strategy of exponential feeding and pH-stat feeding, high concentrations of biomass (65 g dry wt l(-1)) and active soluble TRAIL (1.4 g l(-1)) were obtained within 30 h. The accumulation of acetate, which usually occurs during the process of high density culture of Escherichia coli and especially in the induction stage of protein synthesis, was avoided. PMID- 15269524 TI - Reverse transcription quantitative-PCR of three genes with high homology encoding 3-hydroxy-methylglutaryl-CoA reductase in rice. AB - Reverse transcription followed by RT Q-PCR is useful for the systematic measurement of changes in gene expression. RT Q-PCR with two pairs of primers for each gene was used for relative expression of three genes with high homology encoding 3-hydroxy-methylglutaryl-CoA reductase (HMGR) in rice. At various growth stages of etiolated seedling and various times after UV-irradiation treatment, RT Q-PCR of each HMGR gene showed a consistent pattern of relative expression with the RT Q-PCR data, using two pairs of primers, giving a high degree of accuracy. Furthermore, the different expression levels of three HMGR genes in a sample were determined by diluting the cDNA concentration. These results indicate that RT Q PCR with only one pair of primers for a gene can quantify the relative expression and that the high expression level of HMGR2 could be quantified in comparison to the low level of HMGR1 expression. PMID- 15269525 TI - Quantification of Saccharomyces cerevisiae viability using BacLight. AB - Yeast viability can be accurately quantified using BacLight, a kit which so far has been used only for bacterial analysis. Upon staining, viable cells can be differentiated from non-viable ones by either confocal laser scanning microscopy (CLSM), epifluorescence microscopy, or flow cytometry. Using Saccharomyces cerevisiae as a model, viabilities quantified by CLSM deviated an average of 1.7% from the actual data, and those determined by flow-cytometry by 1.4%. PMID- 15269526 TI - Development of an optical immunosensor based on the fluorescence of Cyanine-5 for veterinarian diagnostics. AB - A model optical immunosensor was developed to quantify an antibody present in a sample by measuring the fluorescence of Cyanine-5 conjugated with the antibody, using a competitive and a sandwich immunoreaction configuration, with the antigen immobilised in controlled pore glass beads. At pH 2, 94% of the antigen-antibody complex was dissociated, allowing reutilisation. Photobleaching had no effect on the fluorescence. This model system was used to detect Brucella sp. infection and could quantify anti-Brucella sp. antibodies in ovine serum samples in the range from 0.005 to 0.11 mg ml(-1). PMID- 15269527 TI - A quantitative, non-radioactive single-nucleotide insertion assay for analysis of DNA replication fidelity by using an automated DNA sequencer. AB - A method to determine the steady-state kinetic parameters of single-nucleotide insertion in replication was developed using an automated DNA sequencer. The insertion of nucleoside 5'-triphosphates into a 6-carboxyfluorescein-labeled primer by DNA polymerase was quantified from the band pattern on a gel using GeneScan software. The parameters determined by this method were consistent with those obtained by the conventional radioisotope-labeling method. This non radioactive, fluorescent-based method is rapid and can handle a large number of samples to assess cognate or non-cognate base pair formation between natural or unnatural bases in replication. PMID- 15269528 TI - Toxicity and degradation of metal-complexed cyanide by a bacterial consortium under sulfate-reducing conditions. AB - Free cyanide at 1 mm decreased the initial sulfate reduction rate of a batch culture of granular sludge from 0.3 to 0.14 mmol d(-1) g(-1) SS (suspended solid), whereas 0.5 mm cyanide had a minimal effect (0.25 mmol d(-1) g(-1) SS). The order of toxicity of metal-complexed cyanides to the sludge was as follows: zinc-complexed cyanide (most toxic) > free cyanide = nickel-complexed cyanide > copper-complexed cyanide (least toxic), which also corresponds well with the order of the stability (dissociation) constants of the metal-cyanide complexes. A consortium degrading cyanide was enriched using nickel cyanide as the sole nitrogen source. This consortium completely removed 0.5 mm of nickel-complexed cyanide under sulfate-reducing conditions in 11 d. Analysis of clone library of 16S rRNA genes shows that the consortium was composed of three major phylotypes including Desulfovibrio. PMID- 15269529 TI - Improvement of recombinant hirudin production by controlling NH4+ concentration in Pichia pastoris fermentation. AB - In recombinant Pichia pastoris fermentation for hirudin production in a 5 l fermenter, a new strategy was explored to match the short fermentation time at low NH4+ concentration with decreased hirudin degradation at high NH4+ concentration. A combination of a defined medium containing initial 0.025 m NH4+ with NH4+ addition up to 0.6 m in the growth phase was achieved in both the improvement of hirudin production and the repression of hirudin degradation. Intact and total hirudin reached 2.63 g l(-1) and 4.25 g l(-1), respectively. PMID- 15269530 TI - Controllable regioselective enzymatic synthesis of polymerizable 5'-O-vinyl- and 3'-O-vinyl-nucleoside analogues in acetone. AB - An efficient synthesis of polymerizable 3'- and 5'-O-acyl-nucleoside derivatives has been developed from inosine and 2'-deoxyuridine by enzyme-catalyzed regioselective acylation with divinyl dicarboxylates. In acetone, Lipozyme (immobilized lipase from Mucor miehei) gave 5'-O-acyl-nucleoside products, and PPL (lipase from porcine pancreas) provided 3'-O-acyl-nucleoside products. PMID- 15269531 TI - Fermentation of sugar cane bagasse hemicellulose hydrolysate to L(+)-lactic acid by a thermotolerant acidophilic Bacillus sp. AB - Sugar cane bagasse hemicellulose, hydrolyzed by dilute H2SO4, supplemented with mineral salts and 0.5% corn steep liquor, was fermented to L(+)-lactic acid using a newly isolated strain of Bacillus sp. In batch fermentations at 50 degrees C and pH 5, over 5.5% (w/v) L(+)-lactic acid was produced (89% theoretical yield; 0.9 g lactate per g sugar) with an optical purity of 99.5%. PMID- 15269532 TI - Method for purifying porcine mature interleukin-18 from silkworm haemolymph. AB - Recombinant porcine mature interleukin-18 (rPomIL-18) has been purified from the haemolymph of silkworms. After co-infection of two recombinant baculoviruses (BmAcpVL1392-IL-18-His and BmAcpVL1392-casp-1) into the silkworm, rPomIL-18 was produced and secreted into the haemolymph. Polyethylene glycol (PEG) 6000 was added to the haemolymph at 8% (v/w) to precipitate storage proteins which would otherwise bind non-specifically to the metal chelating column and the supernatant then was applied to Sepharose bonded with Ni2+. rPomIL-18 was eluted from the column using 100 mM imidazole buffer at pH 8 with a purity of 93.6%. Approximately 5.3 mg purified rPomIL-18 was obtained from 22 ml haemolymph. It could induce interferon-gamma formation from porcine peripheral blood mononuclear cells. PMID- 15269533 TI - Improved cell growth and Taxol production of suspension-cultured Taxus chinensis var. mairei in alternating and direct current magnetic fields. AB - The growth of suspension cultures of Taxus chinensis var. mairei and Taxol production were promoted both by a sinusoidal alternating current magnetic field (50 Hz, 3.5 mT) and by a direct current magnetic field (3.5 mT). Taxol production increased rapidly from the 4th d with the direct current magnetic field but most slowly with the alternating current magnetic field. The maximal yield of Taxol was 490 microg l(-1) with the direct current magnetic field and 425 microg l(-1) with the alternating current magnetic field after 8 d of culture, which were, respectively, 1.4-fold and 1.2-fold of that without exposure to a magnetic field. PMID- 15269534 TI - Chitosan/gelatin composite microcarrier for hepatocyte culture. AB - Solid and porous chitosan/gelatin (CG) composite microcarriers were prepared by a water-in-oil emulsion process with additional freezing and lyophilization. Adult rat hepatocytes (10(6) cells ml(-1)) attached on CG microcarriers maintained at least 15 d of viability and differentiated functions. Over 15 d, unimmobilized hepatocytes released 1.34-fold less lactate dehydrogenase (LDH), and retained 1.63-, 1.51- and 1.28-fold higher albumin secretion, urea synthesis and 7 ethoxycoumarin deethylation activities, respectively, than those on collagen coated microcarriers. The CG matrix is therefore a promising microcarrier for hepatocyte culture. PMID- 15269535 TI - Establishment of a xylose metabolic pathway in an industrial strain of Saccharomyces cerevisiae. AB - To produce an industrial strain of Saccharomyces cerevisiae that metabolizes xylose, we constructed a rDNA integration vector and YIp integration vector, containing the xylose-utilizing genes, XYL1 and XYL2, which encode xylose reductase (XR) and xylitol dehydrogenase (XDH) from Pichia stipitis, and XKS1, which encodes xylulokinase (XK) from S. cerevisiae, with the G418 resistance gene KanMX as a dominant selectable marker. The rDNA results in integration of multiple copies of the target genes. The industrial stain of S. cerevisiae NAN-27 was transformed with the two integration vectors to produce two recombinant strains, S. cerevisiae NAN-127 and NAN-123. Upon transformation, multiple copies of the xylose-utilizing genes were integrated into the genome rDNA locus of S. cerevisiae. Strain NAN-127 consumed twice as much xylose and produced 39% more ethanol than the parent strain, while NAN-123 consumed 10% more xylose and produced 10% more ethanol than the parent strain over 94 h. PMID- 15269536 TI - Real-time quantitative assay of telomerase activity using the duplex scorpion primer. AB - A novel real-time quantitative method for measuring telomerase activity is described in which the duplex scorpion primer is used to provide an intramolecular probing mechanism for specific detection of telomerase activity. Using this method, linearity from 10 to 10(4) cells expressing telomerase activity could be obtained (R2 = 0.994). The requirement of post-PCR steps is thus obviated. PMID- 15269537 TI - Rapid detection of 'rare' actinomycetes in environmental samples. AB - Natural products continue to be a useful source of new leads for the pharmaceutical industry. Actinomycetes are prolific producers of natural products and one strategy to increase the possibility of discovering novel chemical entities is to screen actinomycetes considered 'rare' in the environment and previously under-represented in natural product screening collections. We describe a method using bacteriophage as a marker to detect these actinomycetes in environmental samples. This method allows samples to be pre-screened for the presence of target actinomycetes before lengthy isolation programmes are undertaken. PMID- 15269538 TI - Inhibition of matrix metalloproteinase-1 activity by the soybean Bowman-Birk inhibitor. AB - Inductively coupled plasma analysis of soybean Bowman-Birk inhibitor (BBI) indicated that BBI was a metalloprotein which contained magnesium, calcium, and zinc at 0.40, 0.43 and 0.008 atom/mol BBI, respectively. Heparin-enhanced gelatin zymography, quenched fluorescence substrate hydrolysis analysis, and the Biotrak assay of the interaction of BBI with the matrix metalloproteinase-1 (MMP-1) demonstrated that demineralized BBI at 30 nM inhibited MMP-1 activity whereas mineralized BBI was inhibitory at 115 nM. PMID- 15269539 TI - Production, purification, and characterization of a novel thermostable serine protease from soil isolate, Streptomyces tendae. AB - An isolate of Streptomyces tendae produced a extracellular protease which was purified to apparent homogeneity giving a single band on SDS-PAGE with a molecular mass of 21 kDa. Optimum activity was at 70 degrees C and pH 6. It was stable at 55 degrees C for 30 min and between pH 4 and 9. It was resistant to neutral detergents and organic solvents such as Triton X-100, Tween 80, methanol, ethanol, acetone, and 2-propanol at 5% (v/v). The enzyme was completely inhibited by 5 mM PMSF, indicating it to be a serine protease. N-terminal amino acid sequence did not show any homology with other known proteolytic enzymes. The protease may therefore be a novel neutral serine protease, which is stable at high temperature and over a broad range of pH. PMID- 15269540 TI - 1,3-Propanediol production by Klebsiella pneumoniae under different aeration strategies. AB - 1,3-Propanediol production by Klebsiella pneumoniae was studied in batch cultures under N2 flow and four airflow systems. Different byproducts were formed under different aeration conditions. An anaerobic/aerobic combined fed-batch culture was developed giving 70 g 1,3-propanediol l(-1) and 16 g 2,3-butanediol l(-1) with total diol yield of 0.6 mol(-1) glycerol. PMID- 15269541 TI - Transesterification activity of lipases immobilized in a phyllosilicate sol-gel matrix. AB - Lipases from Pseudomonas cepacia (P.c.) and Thermomyces lanuginosa (T.l.) were immobilized in a phyllosilicate sol-gel matrix and studied for their ability to catalyze the alcoholysis of fats and oils to simple alkyl esters. At 50 degrees C and 48 h reaction immobilized T.l. lipase gave higher alkyl ester yields (70 to 100%) from fats and oils regardless of chain length or degree of unsaturation of the acyl groups in the triacylglycerols than did immobilized P.c. lipase (20 90%), which preferred unsaturated oils. Both immobilized lipases catalyzed ester formation (80-90%) from greases containing a range of free fatty acids (2.6 to 36%). Molecular sieves had no effect on ester yields in the immobilized T.l. lipase-catalyzed alcoholysis of greases but did improve yields (5-10%) in the immobilized P.c. lipase-catalyzed reactions. PMID- 15269542 TI - Production of an anti-complement exo-polymer produced by Auricularia auricula judae in submerged culture. AB - Exo-polymer (EP) was produced at 1.2 g l(-1) in submerged culture of Auricular auricula-judae. Crude EP (AJ-0) has 70% anti-complementary activity (inhibition of total complementary hemolysis 50%; ITCH50). The activating pathway of the complement system occurred through both the classical and alternative pathways, though the major pathway was the classical one. Fractionation of AJ-0 using Sepharose CL-6B gel chromatography gave three major fractions (AJ-Fr-I, II and III) of which the first was the most active. The mycelial growth and EP production of A. auricula-judae were optimal at pH 6, 25 degrees C and pH 5, 25 degrees C, respectively. PMID- 15269543 TI - BpaI and BpnI: novel type II restriction endonucleases from Bacillus pasteurii and Bacillus pantothenticus. AB - Two novel type II restriction endonucleases, designated as BpaI and BpnI, were isolated from Bacillus pasteurii strain1761 and Bacillus pantothenticus strain1639, respectively. They were partially purified and SDS-PAGE indicated Mr values of 28 and 67 kDa for BpaI, 28 and 48 kDa for BpnI. The partially purified endonucleases hydrolyzed DNA into discrete fragments: pUC18 (2.6 kb for BpaI; 1.8 and 0.8 kb for BpnI), pBR322 (2.5 and 1.8 kb for BpaI; 2.6 and 1.7 kb for BpnI) and phix174 DNA (3.2 and 2.1 kb for BpaI; 4 and 1.3 kb for BpnI). PMID- 15269544 TI - Flow process for electroextraction of intracellular enzymes from the fission yeast, Schizosaccharomyces pombe. AB - Flow treatment of the yeast, Schizosaccharomyces pombe, with high intensity electric field pulses released intracellular enzymes such as glyceraldehyde-3 phosphate dehydrogenase and phosphoglycerate kinase. Over 70% of the total activity was liberated within 4 h after pulse application. The optimal field intensities were considerably higher than that needed for irreversible plasma membrane permeabilization. PMID- 15269545 TI - Construction and application of fusion proteins of D-amino acid oxidase and glutaryl-7-aminocephalosporanic acid acylase for direct bioconversion of cephalosporin C to 7-aminocephalosporanic acid. AB - Two fusion proteins of D-amino acid oxidase (DAAO) and glutaryl-7 aminocephalosporanic acid acylase (GLA) were designed to simplify the bioconversion process of cephalosporin C to 7-aminocephalosporanic acid (7-ACA), which is conventionally produced in a two-step enzymatic process. Two recombinant plasmids, pET-DLA and pET-ALD, were constructed to express fusion proteins of DAAO-linker-GLA (DLA) and GLA-linker-DAAO (ALD), respectively. When the recombinant plasmids were expressed in E. coli, the fusion protein DLA was not correctly folded and only DAAO activity could be detected. ALD, however, possessed activities of both DAAO and GLA, which directly catalyze the conversion of cephalosporin C into 7-ACA. PMID- 15269546 TI - Optimization of methanol biosynthesis from methane using Methylosinus trichosporium OB3b. AB - Methylosinus trichosporium OB3b oxidized methane to methanol in the presence of a high concentration of Cu2+. Further oxidation of methanol to formaldehyde was prevented by adding 200 mM NaCl which acted as a methanol dehydrogenase H inhibitor. The bacterium, 0.6 mg dry cell ml(-1), in methane/air (1:4, v/v) at 25 degrees C in 12.9 mM phosphate buffer (pH 7) containing 20 mM sodium formate and 200 mM NaCl accumulated 7.7 mM methanol over 36 h. PMID- 15269547 TI - Mathematical modeling for prediction of endo-xylanase activity and arabinoxylans concentration during mashing of barley malts for brewing. AB - A mathematical model describing the degradation of arabinoxylans by endo-xylanase during mashing process was developed. Endo-xylanase activities and arabinoxylans concentrations in laboratory scale mashing process at different temperature profiles were measured and then used for identifying the model parameters for Harrington barley malt. The modeling errors range for the final concentration of arabinoxylans in wort was -4% to +11.9%. The model developed was also used for predicting the other three different malts mashing processes in laboratory scale, and the prediction errors ranged from -9.5% to +13.6%. The model prediction accuracy for industrial scale mashing process was lower than that in laboratory scale. The simulation results showed that, a lower concentration of arabinoxylans could be achieved when maintaining the mashing-in at 45 degrees C and prolonging the mashing-in time. PMID- 15269548 TI - Expression of immunogenic VP2 protein of infectious bursal disease virus in Arabidopsis thaliana. AB - VP2 protein is the major host-protective immunogen of infectious bursal disease virus (IBDV) of chickens. Transgenic lines of Arabidopsis thaliana expressing recombinant VP2 were developed. The VP2 gene of an IBDV antigenic variant E strain was isolated, amplified by RT-PCR and introduced into a plant expression vector, pE1857, having a strong promoter for plant expression. A resulting construct with a Bar gene cassette for bialaphos selection in plant (rpE-VP2) was introduced into Agrobacterium tumefaciens by electroporation. Agrobacterium containing the rpE-VP2 construct was used to transform Ar. thaliana and transgenic plants were selected using bialaphos. The presence of VP2 transgene in plants was confirmed by PCR and Southern blot analysis and its expression was confirmed by RT-PCR. Western blot analysis and antigen-capture ELISA assay using monoclonal anti-VP2 were used to determine the expression of VP2 protein in transgenic plants. The level of VP2 protein in the leaf extracts of selected transgenic plants varied from 0.5% to 4.8% of the total soluble protein. Recombinant VP2 protein produced in plants induced antibody response against IBDV in orally-fed chickens. PMID- 15269549 TI - Alkaloid accumulation in Catharanthus roseus cell suspension cultures fed with stemmadenine. AB - Feeding stemmadenine to Catharanthus roseus cell suspension culture resulted in the accumulation of catharanthine, tabersonine and condylocarpine. Condylocarpine is not an intermediate in the pathway to catharanthine or tabersonine when it is fed to the cultures. The results support the hypothesis that stemmadenine is an intermediate in the pathway to catharanthine and tabersonine. PMID- 15269550 TI - Biosurfactant production by Serratia marcescens SS-1 and its isogenic strain SMdeltaR defective in SpnR, a quorum-sensing LuxR family protein. AB - Serratia marcescens SS-1 and its SpnR-defective isogenic mutant, SMdeltaR, produced an extracellular surfactant able to decrease surface tension of water from 72 to 37 dyne cm(-1) (SMdeltaR strain) and to 45 dyne cm(-1) (SS-1 strain). The biosurfactant also emulsified kerosene and diesel with a maximum emulsion index of 77% (diesel and kerosene) for the SMdeltaR strain, and 72% (kerosene) and 40% (diesel) for the SS-1 strain. Deletion of spnR gene appeared to enhance biosurfactant production. Model simulations suggest that biosurfactant production by the two strains was growth-associated. The SMdeltaR strain had a yield coefficient of 22-32% g dry cell(-1), which is 32-50% higher than that of the SS 1 strain. PMID- 15269551 TI - Biodegradation of polycyclic aromatic hydrocarbons by Pichia anomala. AB - Pichia anomala 2.2540, isolated from soil contaminated by crude oil, degraded naphthalene, dibenzothiophene, phenanthrene and chrysene, both singly and in combination. The yeast degraded 4.5 mg naphthalene l(-1) within 24 h. Phenanthrene was degraded after a lag of 24 h. When a mixture of all four polycyclic aromatic hydrocarbons was treated at either 0.1-1.6 mg l(-1) or 3.1 5.3 mg l(-1), naphthalene was completely degraded first within 24 h, followed by phenanthrene and dibenzothiophene after 48 h. Chrysene, which remained in the mixture even after 96 h, could be degraded along with naphthalene. Chrysene at 0.7 and 1 mg l(-1), in the presence of 4.3 and 65 mg naphthalene l(-1), respectively, was removed within 96 h. PMID- 15269552 TI - Development of a varicella virus vaccine stabilizer containing no animal-derived component. AB - Various stabilizers containing no animal-derived components were assessed for their efficacy in stabilizing live attenuated varicella virus under various storage temperatures. Stabilizers containing carrageenan, soy protein hydrolysates, or sucrose had a satisfactory performance to maintain infectivity of the cell-free virus when compared to control stabilizer containing animal derived gelatin/gelatin hydrolysate. PMID- 15269553 TI - Cloning of two carotenoid ketolase genes from Nostoc punctiforme for the heterologous production of canthaxanthin and astaxanthin. AB - For the heterologous synthesis of keto-carotenoids such as astaxanthin, two carotenoid ketolase genes crtW38 and crtW148, were cloned from the cyanobacterium, Nostoc punctiforme PCC 73102 and functionally characterized. Upon expression in Escherichia coli, both genes mediated the conversion of beta carotene to canthaxanthin. However in a zeaxanthin-producing E. coli, only the gene product of crtW148 introduced 4-keto groups into the 3,3'-dihydroxy carotenoid zeaxanthin yielding astaxanthin. The gene product of crtW38 was unable to catalyze this reaction. Both ketolases differ in their interaction with a hydroxylase in the biosynthetic pathway from beta-carotene to astaxanthin. PMID- 15269554 TI - Odorous swine wastewater treatment by purple non-sulfur bacteria, Rhodopseudomonas palustris, isolated from eutrophicated ponds. AB - Three strains of phototrophic, purple, non-sulfur bacteria, isolated from eutrophic ponds, were used to treat odorous swine wastewater. One isolate, Rhodopseudomonas palustris, when cultured in swine wastewater without supplementation for 7 d, removed odorous organic acids (170 mg l(-1)), COD (10,000 mg l(-1)) and phosphate (180 mg l(-1)). PMID- 15269555 TI - Starch hydrolysing Bacillus halodurans isolates from a Kenyan soda lake. AB - Fourteen obligate alkaliphilic and halotolerant bacterial isolates, exhibiting extracellular amylase activity at 55 degrees C and pH 10, were isolated from hot springs around Lake Bogoria, Kenya. From 16S rDNA sequence analysis, nine isolates shared 100% identity with Bacillus halodurans strain DSM 497T, while the rest shared 99% identity with alkaliphilic Bacillus species A-59. PCR of the intergenic spacer region between 16S and 23S rRNA genes (ISR-PCR) divided the isolates into two groups, while tDNA-PCR divided them into three groups. Bacillus halodurans DSM 497T had a different ISR pattern from the isolates, while it had a tDNA-PCR profile similar to the group that shared 99% identity with alkaliphilic Bacillus species A-59. All isolates hydrolysed soluble starch as well as amylose, amylopectin and pullulan. The amylase activity (1.2-1.8 U ml(-1)) in the culture broths had an optimum temperature of 55-65 degrees C, was stimulated by 1 mm Ca2+, and was either partially (16-30%) or completely inhibited by 1 mM EDTA. Activity staining of the cell-free culture supernatant from the isolates revealed five alkaline active amylase bands. PMID- 15269556 TI - Optimization of cryoprotectants for cryopreservation of rat hepatocyte. AB - Rat hepatocytes were cryopreserved in hormonally-defined medium (HDM) containing either fetal bovine serum (FBS), glycerol, dimethyl sulfoxide (DMSO), sucrose or a mixture of these as a cryoprotectant. The best survival was with 10% (v/v) DMSO containing 30% (v/v) FBS using 5 x 10(5) hepatocytes ml(-1) at -70 degrees C for 5 d on type I collagen-coated dishes. After thawing, the cell viability was 81% determined by the MTT-test. The cryopreserved hepatocytes had the capacity of albumin synthesis similar to hepatocytes without cryopreservation. This result shows that cryopreservation of rat hepatocyte can be used for the evaluation of hepatic functions. PMID- 15269557 TI - Fluorometric assay for determining epoxide hydrolase activity. AB - We have developed a rapid screening procedure that enables the screening of hundreds of enzyme samples or variants for epoxide hydrolase activity towards any substrate. The procedure detects the products of the enzymatic reaction via periodate cleavage and remaining fluorescence of carboxyfluorescein. PMID- 15269558 TI - Enhancement of mycelial growth and polysaccharide production in Ganoderma lucidum (the Chinese medicinal fungus, 'Lingzhi') by the addition of ethanol. AB - Methanol, ethanol, 1-propanol and 2-propanol, at 1.5% (v/v), enhanced the growth and polysaccharide production of Ganoderma lucidum. Ethanol was the most effective at 1.5% (v/v) for increasing the biomass production, however, the maximal polysaccharide concentration was produced with 2% (v/v) ethanol in the medium. There was no new polysaccharide component produced by the addition of ethanol. PMID- 15269559 TI - Use of Zymomonas mobilis and Saccharomyces cerevisiae mixed with Kluyveromyces fragilis for improved ethanol production from Jerusalem artichoke tubers. AB - Jerusalem artichoke mashed tubers were fermented using single yeasts and a bacterium as well as mixed culture of microorganisms. Kluyveromyces fragilis, a yeast with an active inulinase, was used together with either a commercial distillery yeast, Saccharomyces cerevisiae, or the bacterium Zymomonas mobilis. After batch fermentation the best ethanol concentration of 0.48 g g(-1) for the mixed population and 0.46 g g(-1) for the single population can be obtained. The theoretical yield of the mixed cultures was 2-12% higher than for the single microorganism. PMID- 15269560 TI - Preparation of dodecanol-tolerant strains of Yarrowia lipolytica. AB - Dodecanol (1% v/v) and dodecanoic acid (1% w/v) inhibited growth of Yarrowia lipolytica in complex media supplemented with glucose but dodecanedioic acid (1% w/v) was not toxic. Dodecanol-tolerant strains were prepared from the wild type strain H222 as well as the acyl-CoA oxidase deleted (deltaPOX2, POX3, POX5) strain MTLY35. These strains grew in rich media containing up to 10% (v/v) dodecanol. Dodecanol-tolerant strains remained dodecanol tolerant after they had been cultured in rich media without dodecanol. No significant amount of dodecanedioic acid was accumulated by the dodecanol-tolerant strains when grown on glucose in the presence of dodecanol. PMID- 15269561 TI - Rosmarinic acid production by Lavandula vera MM cell suspension: the effect of temperature. AB - Lavandula vera MM cell suspension, grown at 28 degrees C in a 3-l bioreactor, produced rosmarinic acid maximally at 3 g l(-1)) though most biomass (33.2 g dry wt l(-1)) was at 30 degrees C. PMID- 15269562 TI - Microbial monitoring by molecular tools of a two-phase anaerobic bioreactor treating fruit and vegetable wastes. AB - Microbial consortia in a two-phase, anaerobic bioreactor using a mixture of fruit and vegetable wastes were established. Bacterial and archaeal communities obtained by a culture-independent approach based on single strand conformation polymorphism analysis of total 16S rDNA showed the adaptation of the microflora to the process parameters. Throughout the 90 d of the study, the species composition of the bacterial community changed significantly. Bacterial 16S rDNA showed at least 7 different major species with a very prominent one corresponding to a Megasphaera elsdenii whereas bacterial 16S rDNA of a methanization bioreactor showed 10 different major species. After two weeks, Prevotella ruminicola became major and its dominance increased continuously until day 50. After an acid shock at pH 5, the 16S rDNA archaeal patterns in the acidogenic reactor showed two major prominent species corresponding to Methanosphaera stadtmanii and Methanobrevibacter wolinii, a hydrogenotrophic bacterium. PMID- 15269563 TI - The elderly and cardiovascular disease: some differences, but many similarities to their younger counterparts. PMID- 15269564 TI - Cardiopulmonary resuscitation programs revisited: results of a community study among older African Americans. AB - Early cardiopulmonary resuscitation (CPR) performed by a layperson and prompt defibrillation in the field are critical links in the chain of survival of out-of hospital sudden cardiac arrest. It has been suggested that minorities, women, and elderly persons are often left out of CPR training programs. To examine knowledge and attitudes toward CPR and automatic external defibrillation among African Americans, the author and colleagues conducted home interviews in a population sample of 425 older men and women in Miami-Dade County, FL. It was found that 25% of the participants did not know what CPR was. Only 18% of men and 28% of women had ever taken CPR classes. Mean age the time of CPR training was for men 36 years and for women 46 years. About 74% of all subjects did not know whom to contact for CPR training, and fewer than 5% knew about the American Heart Association Heartsaver Program (including automatic external defibrillation performed by laypersons). The majority of participants suggested churches or community organizations as organizers of CPR training. This study shows that there is a major need for improving knowledge and intensifying CPR training programs among older African Americans. Community organizations and churches may play a critical role in reaching this goal. PMID- 15269565 TI - Presenting symptoms, admission electrocardiogram, management, and prognosis in acute coronary syndromes: differences by age. AB - In a nationwide survey conducted in all 26 hospitals in Israel during February and March 2000, data were collected on 2133 consecutive acute coronary syndrome patients. The patients were divided into three age subgroups: <65 years (n=974), 65-74 years (n=500), and > or =75 years (n=639). The frequency of no anginal pain/atypical symptoms on presentation increased with age for all acute coronary syndrome patients (14%, 21%, and 32%, in the three age subgroups, respectively; p for trend <0.0001). The frequency of ST-elevation on admission electrocardiogram decreased with advancing age (59%, 46%, and 42%, in the three age subgroups, respectively; p for trend <0.0001), whereas ST-depression gradually increased (14%, 24%, and 28%, respectively; p for trend <0.0001). In multivariate analysis, variables associated with no anginal pain/atypical symptoms on presentation (in decreasing order) were: history of heart failure, age, lack of past angina, diabetes, and nonsmoking. ST-elevation was inversely associated with no anginal pain/atypical symptoms on admission (odds ratio, 0.48; 95% confidence interval, 0.37-0.63). The use of acute reperfusion therapy significantly declined with advancing age. Seven-day, 30-day, and 1-year mortality increased with advancing age. No anginal pain/atypical symptoms on presentation were associated with an increased early and late mortality in all three age subgroups. PMID- 15269566 TI - Angiotensin II receptor blockers in older patients. AB - Older patients with hypertension are often inadequately treated due to misconceptions regarding reasonable goal blood pressures or concerns about treatment side effects. Adequately treating hypertension can yield impressive benefits in terms of improved morbidity and enhanced quality of life in persons of any age. Antagonists of the renin-angiotensin-aldosterone system are especially effective in older persons, many of whom have concomitant conditions such as diabetes mellitus, renal dysfunction, and other cardiovascular risk factors. Treatment with angiotensin-converting enzyme inhibitors and angiotensin II receptor blockers has been shown to improve many of the complications of hypertension, including left ventricular hypertrophy and renal disease. Results of recent key studies such as Losartan Intervention For Endpoint Reduction in Hypertension (LIFE), Valsartan in Heart Failure Trial (Val-HeFT), Candesartan in Heart Failure Assessment of Reduction in Mortality and Morbidity (CHARM), and Valsartan in Acute Myocardial Infarction (VALIANT) add to the evidence that angiotensin II receptor blockers are well suited for the treatment of hypertension in older patients. These trials also indicate that they are appropriate therapy for heart failure patients and for patients who have experienced acute myocardial infarction, particularly those who are unable to tolerate an angiotensin-converting enzyme inhibitor. PMID- 15269567 TI - Cardiopulmonary resuscitation in the African-American community. PMID- 15269568 TI - Guidelines of the Cardiogeriatrics Department of the Brazilian Cardiology Society: commentary by Editorial Board Members of The American Journal of Geriatric Cardiology. PMID- 15269569 TI - Aggressive lowering of low-density lipoprotein cholesterol: do the studies apply to the elderly? PMID- 15269570 TI - Ethical issues in the management of geriatric cardiac patients. PMID- 15269571 TI - Ximelagatran: light at the end of the tunnel or the next tunnel? PMID- 15269572 TI - Electrocardiology teacher analysis and review: hyperkalemia. PMID- 15269574 TI - Gynecomastia: an outcome analysis. AB - BACKGROUND: Gynecomastia refers to the presence of femalelike mammary glands in a male. This disorder can lead to significant psychologic stress and self consciousness. This study consists of a chart review of 174 patients treated surgically between July 1, 1976, and February 27, 2001. RESULTS: Operative procedures were excision, excision with suction-assisted lipectomy (SAL), SAL, skin excision (skin) and skin excision with SAL. Overall complication rate was 20%. No significant difference in complication rates was found between grades or procedures. Revision rates between grades were I = 10.3%, II = 14.5% and III = 34.8% (P < 0.001). In grade III, gynecomastia revision rates for excision +/- SAL was 29% and skin +/- SAL was 38.1% (P = 0.644). Of the 8 revisions in the skin sparing procedures, 6 were revised with a scar-forming procedure. Therefore, 77% of patients with grade III gynecomastia were adequately treated with a skin sparing procedure. CONCLUSION: Skin-sparing operations should be the initial procedure chosen for most grade III gynecomastia patients. PMID- 15269575 TI - Primary cleft nasal repair: the composite V-Y flap with extended mucosal tab. AB - A method of primary cleft lip nasal repair utilizing a medially based composite alar flap with a mucosal tab extension is presented. The procedure modifies, with a 5- to 6-mm mucosal tab extension, a previously described chondromucosal flap technique. Most cases were done concurrent with a modified Tennison lip repair. The flap consists of the lateral crus of the alar cartilage, together with its vestibular lining. The flap is advanced medially so the dome provides the tip support for the affected side of the nose. The goal is to restore symmetry, obviating the need for future major nasal surgery. Experience with this technique in 32 patients over 4 years is reported. Although encouraged by our results, it is anticipated significant percentage of patients will still benefit from secondary nasal surgery when their nasal growth is complete. PMID- 15269577 TI - Management of maxillofacial problems in self-inflicted rifle wounds. AB - Severe gunshot wounds to the face, produced by high-velocity rifles or shotgun blasts, present a formidable challenge to reconstructive surgeons. In this study, the results of 14 cases with gunshot wounded faces caused by fire from rifles are presented, and the principles of the management of those victims were determined. These patients had attempted to commit suicide and placed the muzzles of the rifles beneath their chins. The ages of the patients ranged from 20 to 24 years, with a mean age of 22 years. These wounds were caused by close-range gunshots (<10 cm), and the missiles had high velocity (more than 800 m/second). All patients had wounds in their submental triangle areas. The exit sites of the missiles differed among patients. All exit wounds were in the angle limited by the deviation from the gun-barrel axis. After clinical and radiologic evaluation and conservative debridement of all devitalized tissues, the fractures were reduced and stabilized appropriately. Large bony defects were treated by bone grafting, and all soft tissue lesions were closed in layers. The entrance and exit sites were covered primarily after thorough debridement except one case whose defect was reconstructed with bilateral sternocleidomastoid (SCM) flaps, one for submental skin and the other for the mouth floor. Intraoral soft tissues were then repaired by primary closure, tongue flaps, or SCM flaps in case they were necessary. Free tissue transfers were not required for treatment of secondary soft-tissue problems. Resolution of tissue edema, softening of scars in time, and insertion of bone graft may improve the deformity significantly. The initial anatomic reconstruction of the existing bone skeleton and the maximal use of regional tissue for cutaneous reconstruction provide an esthetic appearance that can never be duplicated by secondary reconstruction. PMID- 15269578 TI - Subtotal reconstruction of the nasal septum using a conchal reshaped graft. AB - The caudalmost section of the cartilaginous nasal septum performs the important function of supporting the middle and lower third of the nose. Its absence leads inevitably to deformation of the nasal pyramid and collapse of the internal nasal valve. The most frequent causes of its loss are iatrogenic and traumatic, and the mucoperichondrial lining remains intact in most cases. A graft of conchal cartilage constitutes the preferred method of reconstruction due to the capacity of the transplanted tissue to acquire characteristics of shape, elasticity, and strength closely resembling those of the original tissue to be replaced. The auricular concha differs in anatomic shape from the nasal septum and tends, when deformed, to return to its initial appearance due to cartilaginous memory. The auricular cartilage is also less robust than the quadrangular. The paper describes a surgical technique for reshaping of the conchal cartilage that makes it possible to obtain a practically straight conchal graft that will retain its stability over time with no risk of subsequent modification. The technique involves a double figure-of-8 suture together with incisions on the concave side of the graft for straightening purposes. The simultaneous use of 2 spreader grafts taken from and attached to the concha itself helps to maintain straightness and reinforce the structure. The graft is then placed in position via open access and secured between the 2 mucoperichondrial flaps after these have been carefully detached. The tissue and technique used make it possible to restore the original condition in anatomic and physiological terms, eliminating the esthetic impairment and regaining respiratory functionality. PMID- 15269579 TI - Usefulness of a palmar crease template for the treatment of complicated syndactyly. AB - The treatment of complicated syndactyly has been a difficult problem because it involves not only cutaneous syndactyly but also abnormal arrangement of the finger rays. This means that this anomaly is characterized by abnormal patterns of the palmar creases. The authors prepared a template for tracing the proximal, middle, and distal palmar creases of the nonaffected hand on a clear film and used it as a mirror image to create the preoperative design, to measure the distance between the fingers, and to estimate the location and size of the skin graft intraoperatively. The authors think that the recreation of the normal palmar crease pattern in the affected hand can lead to normal arrangement of the fingers and it is very useful for the reconstruction of multiple finger webbing as part of the treatment of complicated syndactyly. PMID- 15269580 TI - Localization of the arcuate line from surface anatomic landmarks: a cadaveric study. AB - The arcuate line is a relevant structure when reconstructing the abdominal wall after rectus abdominis musculocutaneous flap harvest. Its location is classically taught to be half the distance from the pubic symphysis to the umbilicus, but recent anatomic literature provides evidence to the contrary. Better understanding of the relationship between the arcuate line and surface anatomic landmarks could facilitate better preoperative planning when harvesting a rectus abdominis musculocutaneous flap. A total of 32 arcuate lines were dissected in 18 cadavers, and the location was correlated to various surface anatomic landmarks. The arcuate line was found to lie at 74.6% of the distance from the pubic symphysis to the umbilicus, and 32.7% of the distance from the pubic symphysis to the xiphoid. This location was 1.8 +/- 1.7 cm superior to the level of the anterior superior iliac spines (ASIS). This study provides further support for the finding in the anatomic literature that the arcuate line is substantially more superior than classically described. This knowledge may prove useful in preoperative planning of rectus abdominis musculocutaneous flap harvest. PMID- 15269581 TI - Clinical comparative study of aquacel and paraffin gauze dressing for split-skin donor site treatment. AB - The management of split-thickness skin graft donor sites is targeted towards promoting the healing process, while minimizing adverse effects and complications. The aim of this study was to compare donor site treatment outcome between Aquacel, a carboxymethylcellulose-based hydrofiber dressing, and the standard mesh paraffin gauze dressing. The study included 23 adult patients. Half of the skin graft donor site in the proximal thigh was dressed with paraffin gauze and the rest with Aquacel. The results indicated that patients treated with Aquacel experienced significantly less pain and a more rapid rate of epithelialization compared with patients treated with mesh paraffin gauze dressing. Final scarring (ie, after the 1-year follow-up) was significantly better with the Aquacel dressing. We conclude that Aquacel dressing is superior to the standard mesh paraffin gauze dressing for split-thickness donor site area in pain relief, ease of treatment, promotion of epithelialization, and the quality of scarring. PMID- 15269582 TI - In vivo visualization of platelet/endothelium cell interaction in muscle flaps. AB - Increasing evidence underlines the substantial pathophysiological impact of platelets on the development of ischemia/reperfusion injury (I/R) in flaps. Methods for studying dynamic platelet mechanisms in flaps in vivo are not available. The aim of this study was to develop a model enabling quantitative analysis of platelet kinetics and platelet-endothelium cell interaction within the microcirculation of muscle flaps in vivo. Balb/c mice (n = 16) were anesthetized, and an epigastric muscle flap was prepared. Autologous platelets were separated from blood donor animals (n = 16) and labeled ex vivo by means of rhodamine-6-G. After I/R (90 minutes' clamping, 10 minutes' reperfusion), the platelets were administered intra-arterially (i.a.). Microhemodynamics and kinetics of platelets were investigated by intravital fluorescence microscopy. I/R of muscle flaps induced disturbances in microcirculation. The number of rolling platelets, as well as platelets adhering to the inner vessel wall of venules, was increased in the ischemia group. Using intravital fluorescence microscopy, platelet kinetics were analyzed directly in flap microcirculation in vivo for the first time. Since platelet/endothelial cell interaction is a key event in the pathophysiology after microsurgical procedures, this model will help to understand basic molecular mechanisms of platelet behavior during I/R. PMID- 15269583 TI - The effect of hyperbaric oxygen therapy on composite graft survival. AB - Auricular composite grafts are a useful reconstructive option, particularly for nasal reconstruction. This study evaluates the effect of hyperbaric oxygen (HBO) therapy on auricular composite graft survival in rabbits. Circular chondrocutaneous composite grafts of 0.5, 1, or 2 cm in diameter were resected from the ears of rabbits. The grafts were sutured back into position. Half the rabbits in each group received HBO postoperatively, consisting of 90 minutes at 2.4 atm. Rabbits received 7 treatments in 5 days. Control rabbits did not receive HBO. On day 21 the percentage area of graft survival was calculated from gross and histologic examination. Two-centimeter grafts treated with HBO (n = 8) had a mean graft survival rate of 85.8 +/- 15.7% compared with a survival rate of 51.31 +/- 38.5% for the control group (n = 8; P = 0.0478). There was no such benefit in smaller grafts. HBO could prove clinically useful for larger composite grafts. PMID- 15269584 TI - Neurocutaneous island flap:: an experimental model using the rabbit ear. AB - The rabbit ear has often been used as an experimental model; however, a neurocutaneous flap has never been described before. The authors examined the reliability of a skin cartilage flap with a pedicle composed only of a sensory nerve with its vascular network in a rabbit model: A 2 x 2-cm cutaneous cartilage flap was harvested on the dorsal side of the ear bilaterally. Vessels were tied and cut on the 4 sides of the flap, including the central auricular artery and vein. The nerve was cut at the distal side of the flap and was "skeletonized" to the extent of 1 cm on the proximal side, meticulously preserving its vascular network. Subsequently, the flap was elevated with the cartilage as a composite flap and was sutured back to its natural site. The authors elevated 28 skin cartilage flaps on the dorsal side of both the ears of 14 New Zealand White rabbits, centered on the central neurovascular axis. All 28 flaps survived. In the control group of 7 rabbits, the artery, vein, and nerve (pedicle) were severed, and the skin cartilage island was sutured back as a composite graft. None of these 14 grafts survived. In 4 additional rabbits, the authors performed a histologic examination 1 day, 3 days, 7 days, and 3 weeks postoperatively of the neurovascular axis after the same skin cartilage flap was harvested. They compared these results with the histologic examination of the nerve of the contralateral nonoperated ear, and noted a marked dilatation and multiplication of blood vessels in the operated ear beginning on day 1 postoperatively. The presence of this neurocutaneous vascularization should be considered when other kinds of flaps (venous flaps or others) are used as experimental models using the rabbit ear. PMID- 15269585 TI - Immunolocalization of vascular endothelial growth factor during heterotopic bone formation induced from grafted periosteum. AB - Vessel invasion is an important step in cartilage replacement that leads to bone formation, and vascular endothelial growth factor (VEGF) has been implicated as a key player in this process. Although grafted periosteum undergoes endochondral ossification, little is known about the role of VEGF in this process. In the current study the authors investigated by immunohistochemical, histochemical, and ultrastructural techniques the localization of VEGF during bone formation in periosteal grafts. At day 14 after grafting the tibias of Japanese white rabbits, periosteal cells in the grafted tissue had differentiated into chondrocytes to form cartilage. Some chondrocytes were immunopositive for VEGF expression, and subsequent vessel invasion occurred predominantly in these VEGF-positive areas. At day 45, the cartilage invaded by blood vessels had been replaced by newly formed bone. These findings suggest that VEGF is associated with the process of blood vessel invasion into cartilage before bone replacement in endochondral ossification from grafted periosteum. PMID- 15269586 TI - Alpha V integrin prolongs collagenase production through Jun activation binding protein 1. AB - Robust expression of alphav integrin and matrix metalloproteinase 1 (MMP1) plays an important role in cancer metastasis and wound healing. A patient with an abnormal scar that appeared stretched and thinned out was found to have fibroblasts that overexpressed alphav integrin; therefore, a relationship between alphav integrin expression and MMP1 production was sought. A yeast 2 hybrid screen revealed alphav integrin interacts with jun activation binding domain-1 (JAB1). Mesenchymal-derived cells were transfected with the alphav integrin gene and incorporated into collagen lattices. Transfected cells maximally contracted collagen lattices beginning on day 5, whereas control transfected cells did not contract lattices. Late-phase collagen lattice contraction was inhibited by a pan MMP inhibitor, BB4. Overexpression of alphav correlated with enhanced MMP1 transcription, as determined by a luciferase assay (P < or = 0.05). Diminution of JAB1 with JAB1 antisense abolished alphav integrin up-regulation of MMP1. We conclude alphav integrin signals through JAB1 to prolong MMP1 production and that this signaling pathway in fibroblasts may lead to abnormal scarring. PMID- 15269587 TI - Tetanus and the plastic surgeon. AB - Tetanus in the United States is decidedly rare, and most of us will not see a case of it during our careers. Given its lethality, it is a disease about which one must be aware. Be willing to consider it as a diagnosis, no matter the immunization status of the patient, if clinical signs and symptoms warrant. To emphasize this point, the authors present a case of an otherwise healthy 41-year old man who sustained electrical burns when he fell from a ladder and struck a power line on his way to the ground. He developed a compartment syndrome of his left leg at the exit site and subsequently underwent fasciotomies. When he later began to exhibit signs and symptoms of sepsis, his wound was debrided, and most of his anterior compartment was resected. Despite this, his condition worsened, and his clinical picture was suggestive of tetanus, including the classic findings of trismus, risus sardonicus, and opisthotonus. Using mechanical ventilation, paralysis, narcotics, and muscle-relaxing sedatives, the authors supported him until his tetany subsided. He survived and was discharged to home when complete coverage of his burns and left leg anterior compartment was obtained. The authors discuss the presentation, diagnosis, and treatment of tetanus, as well as its incidence in the general population and in the previously immunized patient. PMID- 15269588 TI - TRAM flap breast reconstruction after abdominal liposuction. AB - The transverse rectus abdominis musculocutaneous (TRAM) flap has become the gold standard of autologous breast reconstruction. It is typically a low-risk procedure with few surgical contraindications. A relative contraindication, however, is prior liposuction of the abdomen. The contention has been that the trauma of the liposuction procedure can damage or destroy the musculocutaneous perforators that supply circulation to the TRAM flap skin paddle. The authors present 2 patients who previously underwent suction-assisted abdominal lipectomy and, after mastectomies, successfully underwent unilateral breast reconstruction using single-pedicle TRAM flaps. They also examine the literature that supports the feasibility of this procedure. PMID- 15269589 TI - The preexpanded anterolateral thigh free flap. AB - The anterolateral thigh flap has many of the attributes of the ideal soft-tissue flap. However, a major detriment is the potential conspicuous donor site deformity, especially if skin grafted. In elective situations, preexpansion of the lateral thigh with subsequent transfer of even a wide anterolateral thigh flap can permit primary donor site closure and avoidance of a skin graft. This has been achieved successfully in 2 compliant patients, and is a reasonable solution to minimize the morbidity of this otherwise important donor site. PMID- 15269590 TI - Heel reconstruction with an iliac osteocutaneous free flap: 10-year follow-up. AB - During the Homeland War in Croatia, many civilians suffered from war wounds of the extremities. Explosive war wounds create composite and devastating injuries mainly by high-energy transfer to the tissue. We present an early reconstruction of explosive heel wound with an iliac osteocutaneous free flap with late follow up result. PMID- 15269591 TI - Combination of short-pulsed CO2 laser resurfacing and cultured epidermal sheet autografting in the treatment of vitiligo: a preliminary report. AB - Cultured epidermal autografting has been employed in a variety of clinical treatments including vitiligo management. In this study, we successfully treated 2 patients with vitiligo using a short-pulsed CO2 laser and by grafting the autologous cultured epidermis. Small pieces of uninvolved skin (2 x 1 cm) were taken for cultivation from a pudendal or axillary area and were expanded into 2 pieces of epidermal sheets 100 cm. Before grafting, the lesions were abraded superficially using a short-pulsed CO2 laser with a computerized pattern generator. After successful grafting, repigmentation was visible within 1 to 2 months. One year after grafting, the skin color was almost the same as that of the surrounding normal skin. Thus, the combination of short-pulsed CO2 laser resurfacing and cultured epidermal grafting is a powerful option for treating an asymmetric and wide vitiliginous lesion. PMID- 15269592 TI - Microsurgical revascularization of almost totally amputated alar wing of the nose. AB - A case of nearly complete amputation of the alar wing is presented whereby a successful arterial revascularization was accomplished using an arterial rerouting technique. Venous stasis was overcome by means of stab-wound wiping. An excellent result was obtained following complete survival of the revascularized segment. The authors conclude that microvascular revascularization should always be attempted whenever possible, even if a skin bridge is preserved in nearly complete amputations of the nose. PMID- 15269593 TI - A-a type, arterialized, venous, flow-through, free flap for simultaneous digital revascularization and soft tissue reconstruction-revisited. AB - Significant soft tissue injuries to palmar surfaces are frequently associated with digital vessel damage. Flap coverage might have to be combined with microsurgical revascularization using vein grafts if the digit is to be salvaged. Two cases of simultaneous digital revascularization and soft tissue reconstruction using an arterialized, venous flow-through flap are presented in detail. These flaps initially "pinked up" for 24 to 48 hours. This was followed by a period of venous congestion lasting approximately 1 week, after which flap perfusion gradually returned to normal. Good long-term functional and cosmetic results were achieved. Distal finger perfusion was maintained in both cases. This technique, although previously described, has not been popularized. It should be considered early in reconstruction of ischemic digits requiring simultaneous vascular and soft tissue reconstruction. PMID- 15269595 TI - Excellence in surgery: lessons learned from the world of sports. PMID- 15269594 TI - Re: factors determining shape and symmetry in immediate breast reconstruction. PMID- 15269596 TI - Hsp90: an emerging target for breast cancer therapy. AB - Rapidly evolving insights into the specific molecular genetic abnormalities that drive the growth and metastasis of breast cancer have led to the development of targeted therapeutics that do not rely on the generalized disruption of DNA metabolism and cell division for activity. Of particular interest are inhibitors of cellular signal transduction pathways involving tyrosine kinases as well as selective modulators of steroid hormone signaling, histone acetylation, angiogenesis and tumor cell apoptosis. Unique within this array of promising new agents, however, are compounds that target heat shock protein 90 (Hsp90). This molecular chaperone associates with a distinct, but surprisingly diverse, set of proteins that are referred to as Hsp90 client proteins. Hsp90 binds to these clients, and plays a key role in regulating their stability and function. Many of the proteins chaperoned by Hsp90 are involved in breast cancer progression and resistance to therapy, including the estrogen receptor, receptor tyrosine kinases of the erbB family, Akt, and mutant p53. Several small molecule inhibitors of Hsp90 have been identified that can deplete cellular levels of multiple oncogenic client proteins simultaneously by enhancing their ubiquitination and proteasome mediated degradation. The activity of Hsp90 inhibitors has been well validated in preclinical breast cancer models, both in single-agent studies and in combination with conventional chemotherapy. One of these inhibitors, 17-allylamino, 17 demethoxygeldanamycin (17-AAG, NSC 330507) has recently completed phase I testing. The agent was well tolerated at drug exposures that were shown to cause modulation of Hsp90 client protein levels. Given the redundancy and complexity of the molecular abnormalities present in most breast cancers, the ability of Hsp90 inhibitors to alter the activity of multiple oncogenic targets may prove of unique therapeutic benefit. PMID- 15269597 TI - Metastasis suppressor genes: a role for raf kinase inhibitor protein (RKIP). AB - The metastatic cascade is a complicated process that involves many steps from gain of the metastatic phenotype in the primary tumor cells through establishment of macroscopic tumor at the distant target organ. A group of genes, termed metastasis suppressor genes (MSG), encode for proteins that inhibit various steps of the metastatic cascade. Accordingly, loss of MSG promotes the metastatic phenotype. Although several MSG have been identified, the mechanisms through which they enhance metastasis are not clearly defined. Gene array analysis of a low metastatic LNCaP prostate cancer cell line compared to its highly metastatic derivative C4-2B prostate cancer cell line revealed decreased expression of raf kinase inhibitor protein (RKIP) in the C4-2B cell line. RKIP blocks the activation of several signaling pathways including MEK, G-proteins and NFkappaB. Immunohistochemical analysis of prostate cancer primary tumors and metastases revealed that RKIP protein expression was decreased in metastases. Restoration of RKIP expression in the C4-2B cell line diminished metastasis in a murine model. These results demonstrate that RKIP is a MSG. Loss of RKIP enhanced both angiogenesis and vascular invasion, and protected against apoptosis. These findings suggest that targeting the RKIP pathway may diminish the metastatic cascade. However, challenges exist as to the best method to target RKIP expression. Restoration of RKIP expression in all cancer cells in vivo is challenging. A plausible strategy is to use small molecules that target proteins in signaling pathways that are dysregulated due to loss of RKIP. PMID- 15269598 TI - Telomere dynamics determine episodes of anticancer drug resistance in rat hepatoma cells. AB - Clinical and experimental observations indicate that resistance to anticancer drugs may be spontaneously reversible over time, but the mechanisms of this reversal are unknown. The resistance of cultured hepatoma cells to methotrexate (MTX) and cisplatin was followed for 9 months. Cells were exposed to three treatments: MTX 200 nM for 24 h or 15 nM continuously and cisplatin 50 microM for 2 h. We investigated the relation between the temporal pattern of cell resistance and the previously reported fluctuations in cell proliferation rate, telomere length and telomerase activity. Spontaneous major peaks in resistance to each drug fell in time windows of 2-3 months (60-70 population doublings) and were at different times for each drug. The frequency of the fluctuations in drug resistance was the same as that of variations in cell growth rate, but amplitudes were unrelated. By contrast, resistance was directly related to telomere length dynamics in the same cells. MTX resistance occurred when telomeres shortened and cisplatin resistance when they were elongated. Furthermore, peaks of resistance to the continuous treatment with MTX were observed at 350-bp intervals of mean telomere length (9.06, 9.41, and 9.76 kbp) during the two 2-month phases of telomere shortening. Statistical analysis demonstrates the sinusoidal relationship between intermittent MTX resistance and telomere length. Possibly, erosion of telomeres encroaches on periodically spaced nucleosomal proteins, defining the onset of resistance phases. This evidence that resistance of tumoral cells to anticancer drugs may be intermittent and that onset of resistance is dictated by telomere length has major implications for clinical practice. PMID- 15269599 TI - No anti-apoptotic effects of single copies of mutant p53 genes in drug-treated tumor cells. AB - Some mutant forms of the p53 tumor suppressor have been documented to exert novel oncogenic functions including the increase of tumorigenicity, metastatic potential, genomic instability and therapy resistance of tumor cells. The latter has been suggested to be caused, primarily, by inhibition of apoptosis and, in part, through the activation of genes by mutant p53 whose products can counteract drug activities. Recently described in this context was the dUTPase, which may confer resistance to fluoropyrimidine drugs such as 5-fluorouracil (5-FU). We report here findings that call in question the existence of a direct anti apoptotic effect of mutant p53. Wild-type p53-negative human fibroblasts, and Saos-2, H1299 and HCT116 tumor cells, treated with adriamycin, etoposide, cisplatin or 5-FU, failed to show apoptosis resistance when retrovirally bulk infected to express the p53 mutants 175H or 273H at levels observed in naturally mutant p53-producing tumor cells. Furthermore, dUTPase gene expression was not stimulated by mutant p53, but instead by cellular events that involve DNA synthesis. We interpret the combined available data to suggest that much of the anti-apoptotic effect of mutant p53 is indirect and secondary to DNA-damaging and/or repair-interfering effects of these proteins. PMID- 15269600 TI - Cellular response and molecular mechanism of antitumor activity by leinamycin in MiaPaCa human pancreatic cancer cells. AB - Previous in vitro biochemical studies have revealed that the antitumor drug leinamycin causes oxidative DNA damage and DNA alkylation. However, it is still not clear whether the same mechanism(s) of action operate in cultured human tumor cells. Here, we evaluated the effects of leinamycin in the human pancreatic carcinoma cell line MiaPaCa. Leinamycin was highly toxic to MiaPaCa cells in vitro, with an IC50 value of 50 nM, and extensive DNA fragmentation was observed in leinamycin-treated MiaPaCa cells. Flow cytometric experiments showed that leinamycin was able to disrupt normal cell cycle progression, resulting in an initial arrest of the cells in S phase. With increased time or at higher concentrations of leinamycin, the population of cells in the sub-G1 phase gradually increased, indicative of apoptotic cell death due to DNA damage. Mammalian Chk2, but not Chk1 kinase, was found to be activated in MiaPaCa cells treated with leinamycin, indicating that cellular responses to leinamycin could be attributed to DNA strand break formation rather than DNA adduct formation. Like other DNA-damaging anticancer drugs, the downregulation of telomerase activity was also observed in MiaPaCa cells at cytotoxic concentrations. However, leinamycin failed to induce DNA ligase I expression in MiaPaCa cells, unlike other DNA-damaging agents, which are known to inhibit DNA replication by arresting DNA replication forks. Taken together, the results from our study indicate that the DNA strand breakage caused by the oxidative DNA-damaging property of leinamycin is directly related to the cellular responses of this drug in MiaPaCa cells over the DNA alkylation property in a dose-responsive manner. PMID- 15269601 TI - In vitro and in vivo growth inhibition and G1 arrest in human cancer cell lines by diaminophenyladamantane derivatives. AB - We describe the discovery of a novel series of anticancer adamantane derivatives which induce G1 arrest in Colo 205 and HT 29 colon cancer cells. Seven adamantane derivatives were screened for their activity in vitro against 60 human cancer cell lines in the National Cancer Institute (NCI)'s Anticancer Drug Screen system. The relationships between structure and in vitro anticancer activity are discussed. 1,3-Bis(4-(4-amino-3-hydroxyphenoxy)phenyl)adamantane (1,3-DPA/OH/NH2) and 2,2-bis(4-(4-amino-3-hydroxyphenoxy)phenyl)adamantane (DPA) exhibited strong growth inhibitory on anticancer activities in vitro. The IC50s of 1,3-DPA/OH/NH2 (NSC-706835) and DPA (NSC-706832) were found to be < 3 microM against 45 (85%) and 48 (91%) cell lines, respectively. 2,2-Substituted adamantane derivatives exhibited stronger growth inhibition on anticancer activities in vitro than the corresponding 1,3-substituted analogs. Very strong growth inhibition of 2,2-bis(4 aminophenyl)adamantane (NSC-711117) was observed against two colon cancer lines (HT-29 and KM-12), one CNS cancer line (SF-295) and one breast cancer line (NCI/ADR-RES) with IC50 < 1.0 microM, i.e. 0.1, 0.01, 0.059 and 0.079 microM, respectively. In addition, we also examined the in vitro and in vivo effects of DPA on three human colon cancer cells. DPA-treated Colo 205 and HT-29 cells were arrested at G0/G1 as analyzed by flow cytometric analysis. The DPA-induced cell growth inhibition was irreversible after removal of DPA. The in vivo effect of tumor growth suppression by DPA was also observed on colon Colo 205 xenografts. No acute toxicity was observed after an i.p. challenge of DPA in ICR nude mice weekly. These results suggest that DPA appears to be a new potentially less toxic modality of cancer therapy. PMID- 15269602 TI - Reduction of tamoxifen resistance in human breast carcinomas by tamoxifen containing liposomes in vivo. AB - We investigated whether it is possible to reduce anti-estrogen resistance using liposomally encapsulated tamoxifen in vivo. Small liposomal vesicles containing up to 5.1 mg tamoxifen/ml liposomal suspension, together with an alkylphospholipid to enhance the cellular uptake, were prepared and characterized. Mice transplanted with different tumor models were treated with tamoxifen liposomes administered i.p. or orally as a bolus dose of 50 mg/kg once a week or as a daily dose of 10 mg/kg/day, both during a 4-week period. After orally administered tamoxifen liposomes, tumor growth was significantly reduced for the 3366/tamoxifen (acquired resistance) and for the MCF-7 (inherent resistance) models to 47 and 16%, respectively (treated to control value of relative tumor volume). Intraperitoneal treatment with tamoxifen liposomes revealed similar results. Investigation of biodistribution revealed especially an accumulation of liposomal tamoxifen in MCF-7 tumors and livers of the treated mice. These liposomes had uterotrophic properties comparable to the dissolved compound. This study demonstrates for the first time that a liposomal formulation of tamoxifen was able to induce pharmacological effects and to improve the therapeutic efficacy in several anti-estrogen-resistant xenografts. PMID- 15269603 TI - Limited cerebrospinal fluid penetration of docetaxel. AB - Our purpose was to investigate the cerebrospinal fluid (CSF) penetration of docetaxel in cancer patients. Docetaxel was administered as a 1-h infusion at a dose of 75 mg/m2 to two patients with metastatic breast cancer and leptomeningeal carcinomatosis. CSF samples were obtained using a lumbar puncture up to a 72-h time period. Total and unbound docetaxel concentrations in plasma and CSF were determined by liquid chromatography (lower limit of quantitation: 0.5 nM for plasma and 0.050 nM for CSF) and equilibrium dialysis, respectively. The pharmacokinetics of docetaxel in plasma are in line with data of previous studies. The concentrations of docetaxel in CSF did not follow the general pattern in plasma, with relatively stable concentrations over the 72-h time period. The fraction of unbound docetaxel in plasma ranged from 6 to 13%, while that in CSF ranged from 67 to 103%. For total and unbound docetaxel, the CSF to plasma concentration ratio progressively increased in 72 h from 0.01 to 0.6% and from 0.1 to 9%, respectively. These data suggest that measurement of unbound docetaxel is required to accurately assess the extent of drug penetration into CSF and that the drug can produce distribution to CSF at levels associated with significant antitumor activity in experimental models. PMID- 15269604 TI - Mitomycin C, 5-fluorouracil and folinic acid (Mi-Fu-Fo) as salvage chemotherapy in breast cancer patients with liver metastases and impaired hepatic function: a phase II study. AB - Patients with measurable liver metastases due to breast cancer and elevated liver enzymes were enrolled into the study. The planned schedule was mitomycin C 8 mg/m2 on day 1, 5-fluorouracil 750 mg/m2 and folinic acid 300 mg/m2 on day 1 and 2 every 4 weeks (Mi-Fu-Fo). Between May 1998 and December 2002, 30 patients with a median age of 51 years (range 33-74) were enrolled. All of them suffered from extensive metastases of the liver resulting in liver dysfunction. Myelosuppression was the most frequent toxicity. Six patients had a partial remission, 12 patients had stable disease and 12 patients progressed during treatment. The median time to progression was 4.5 months in all patients and 7.0 months in patients who responded to the therapy. The median overall survival for the total population was 6.0 months and in the group of responding patients 12.0 months. Mi-Fu-Fo, therefore, provides a valid option for breast cancer patients with liver dysfunction due to liver metastases. PMID- 15269605 TI - Treatment of advanced digestive non-colon cancer with a weekly 24-h infusion of high-dose 5-fluorouracil modulated by folinic acid and cisplatin: an easy-to-use and well-tolerated combination. AB - The combination of 5-fluorouracil (5-FU) modulated by folinic acid (FA) and cisplatin is commonly used in advanced digestive non-colon cancers (ADNCC). In order to simplify treatment administration by avoiding cisplatin-related hydration, we investigated a weekly regimen of 5-FU/FA/cisplatin. Patients with ADNCC were treated with 5-FU 2.0 g/m2, FA 500 mg/m2 and cisplatin 25 mg/m2 day 1, for 6 weeks with a 2-week rest, and were assessed for toxicity, tumor response and disease-free survival. Forty-three patients with measurable ADNCC were treated with this weekly regimen. Primary tumor sites were mainly esophagus (n = 17), stomach (n = 12) and pancreas (n = 9). Results were as follows. Toxicity was mostly hematological, with 16% grade 3/4 neutropenia (seven of 43) and 4% febrile neutropenia (two of 43). Objective response (OR) was observed in 19 of 43 (44%) patients including four complete responses (9%) and 15 partial responses (35%). Another 18 patients (42%) experienced stable disease. Time to progression was 6.5 months. The median response and stable disease durations were 4.3 (range 3-34) and 5 (range 2-16) months, respectively. We conclude that weekly administration of 5-FU/FA/cisplatin is an active and well-tolerated regimen. Toxicity is manageable and allows chemotherapy on an outpatient basis without hydration program as required when cisplatin is used at the dose of 50 mg/m2. PMID- 15269606 TI - Long-term cytogenetic remission with ubenimex monotherapy in a case of chronic myeloid leukemia. AB - A 65-year-old man was diagnosed with chronic myeloid leukemia (CML) in February 1990, and was treated with busulfan and ubenimex. Cytogenetic analysis of the bone marrow revealed the Philadelphia (Ph) chromosome in 100% of cells of analyzed at diagnosis. Treatment with busulfan was stopped in March 1993 due to bone marrow suppression. The Ph chromosome was seen in 80% of cells in June 1993. He received ubenimex monotherapy after cessation of busulfan. Complete disappearance of the Ph chromosome was confirmed in May 1995 and has continued to date. This suggests that ubenimex might specifically affect the Ph chromosome and be useful as maintenance therapy for CML. PMID- 15269607 TI - Severe disabling sensory-motor polyneuropathy during oxaliplatin-based chemotherapy. AB - Oxaliplatin-based combination chemotherapy is an option for first-line therapy of metastatic colorectal cancer. It is associated with acute hyperexcitability of motor and sensory nerves, and a cumulative sensory axonal neuropathy. We describe a 56-year-old male with metastatic colorectal cancer treated with oxaliplatin and capecitabine who developed a rapidly ascending motor and sensory neuropathy, which rendered him wheelchair-bound. Heightened clinical suspicion for possible oxaliplatin-induced motor neuropathies may be warranted. PMID- 15269609 TI - Dynamic contrast-enhanced MR imaging. AB - Dynamic contrast-enhanced magnetic resonance imaging is a useful clinical tool in evaluation of soft tissue neoplasm and lymph nodes in head and neck. It is thought to be a useful predictor of response to radiotherapy for head and neck carcinoma and used to monitor the treatment and distinguish post-therapeutic changes from recurrent mass with greater confidence. It can be used to distinguish between normal and malignant tissue and to differentiate a malignant lymphoma from other lymph nodal enlargements. The technique utilizes relative differences in microvasculature and microcirculation between malignant and non malignant tissue to achieve greater contrast in signal imaging following bolus contrast administration. This article explains the underlying principles and imaging techniques for this new diagnostic tool. The clinical applications and technical challenges are discussed. The future challenges and some contradictions in results are also outlined. PMID- 15269610 TI - Magnetic resonance imaging of perineural spread of head and neck malignancies. AB - Perineural invasion is a common mechanism of spread of head and neck cancers. Imaging plays an important role in detection of this condition because a large number of patients with perineural spread (PNS) are clinically asymptomatic. Accurate detection of PNS requires an understanding of anatomy of commonly involved neural pathways. High level of suspicion on the part of the radiologist, awareness of common imaging signs of PNS and careful attention to imaging technique can aid in earlier detection of this condition. PMID- 15269611 TI - Magnetic resonance spectroscopy of head and neck neoplasms. AB - Magnetic resonance spectroscopy (MRS) is a validated noninvasive method for evaluation of possible malignant tumor and lymph nodes of the head and neck. From its roots as a budding research application, it has made the critical transition to a widespread clinical tool. MRS analyzes the tissue at a molecular level and searches for the presence of specific metabolites, which are markers for malignancy. Differentiation of benign from malignant neoplasm, detection of recurrence of malignant tumor and noninvasive treatment monitoring of treated or untreated tumor are some of the important utilities of MRS. One dimensional 1H MRS is the most popular and promising technique for spectroscopic analysis while P-31 MRA and two-dimensional correlated spectroscopy (2D COSY) have also showed some promise. This article describes the application of magnetic resonance spectroscopy for evaluation of malignant tumors of the neck. PMID- 15269612 TI - Pediatric head and neck masses. AB - Most neck masses in the pediatric head and neck region are benign. Congenital, developmental, and inflammatory lesions make up most of the masses in the pediatric head and neck. For example, neck masses due to inflammatory lymphadenitis are common in children because of the frequency of upper respiratory tract infections. Although many of the malignant tumors in children are found in the head and neck, they account for only a small portion of the neck masses. The choice of the imaging modality is based on a number of factors, several of which are unique to the pediatric population. Although the bulk of disease entities are adequately evaluated by CT, MRI can provide additional vital information in many cases. MRI provides better soft tissue characterization than CT, has multiplanar capabilities. In this article, we will attempt to provide an overview of conditions that present as neck masses. PMID- 15269613 TI - Superparamagnetic iron oxide nanoparticles: nodal metastases and beyond. AB - Superparamagnetic iron oxide (SPIO) nanoparticles are unique MR contrast agents and are of great interest for their multiple potentials. SPIO nanoparticles have a higher diagnostic accuracy for detecting metastatic lymph nodes than conventional MR studies, particularly in head and neck. The impact of this unique MR contrast agent on treatment decision of patients with head and neck cancer needs to be investigated in comparison with contrast-enhanced CT. As MR technology advances, the accuracy of SPIO nanoparticles for detection of metastasis certainly improves; thus, 1 day we may be able to reliably detect metastases in stage N0 patients, so that treatment strategy is established for each individual patient. This article presents physiologic properties of SPIO, technical considerations and diagnostic accuracy for imaging with SPIO, and other potential applications of SPIO agents. PMID- 15269614 TI - MR imaging of brachial plexus. AB - The brachial plexus is a complex anatomic component originating from ventral rami of the lower cervical nerve roots from C5 to C8 and upper thoracic spinal nerve roots from T1, providing sensory and motor innervation to the upper extremities. As it is inaccessible to palpation, clinical evaluation of the brachial plexus is very challenging and localizing lesions along its course is very difficult. The gamut of pathologic conditions involving the brachial plexus includes primary tumor, direct extension of adjacent tumor, metastasis, trauma, or an inflammatory condition. MR imaging provides superior diagnostic ability due to its ability of multiplanar imaging and greater soft tissue contrast. This article discusses MR imaging findings in a variety of pathologic conditions, with special emphasis on neoplastic process. PMID- 15269615 TI - Guideline for the evaluation of cholestatic jaundice in infants: recommendations of the North American Society for Pediatric Gastroenterology, Hepatology and Nutrition. AB - For the primary care provider, cholestatic jaundice in infancy, defined as jaundice caused by an elevated conjugated bilirubin, is an uncommon but potentially serious problem that indicates hepatobiliary dysfunction. Early detection of cholestatic jaundice by the primary care physician and timely, accurate diagnosis by the pediatric gastroenterologist are important for successful treatment and a favorable prognosis. The Cholestasis Guideline Committee of the North American Society for Pediatric Gastroenterology, Hepatology and Nutrition has formulated a clinical practice guideline for the diagnostic evaluation of cholestatic jaundice in the infant. The Cholestasis Guideline Committee, consisting of a primary care pediatrician, a clinical epidemiologist (who also practices primary care pediatrics), and five pediatric gastroenterologists, based its recommendations on a comprehensive and systematic review of the medical literature integrated with expert opinion. Consensus was achieved through the Nominal Group Technique, a structured quantitative method. The Committee examined the value of diagnostic tests commonly used for the evaluation of cholestatic jaundice and how those interventions can be applied to clinical situations in the infant. The guideline provides recommendations for management by the primary care provider, indications for consultation by a pediatric gastroenterologist, and recommendations for management by the pediatric gastroenterologist. The Cholestasis Guideline Committee recommends that any infant noted to be jaundiced at 2 weeks of age be evaluated for cholestasis with measurement of total and direct serum bilirubin. However, breast-fed infants who can be reliably monitored and who have an otherwise normal history (no dark urine or light stools) and physical examination may be asked to return at 3 weeks of age and, if jaundice persists, have measurement of total and direct serum bilirubin at that time. This document represents the official recommendations of the North American Society for Pediatric Gastroenterology, Hepatology and Nutrition on the evaluation of cholestatic jaundice in infants. The American Academy of Pediatrics has also endorsed these recommendations. These recommendations are a general guideline and are not intended as a substitute for clinical judgment or as a protocol for the care of all patients with this problem. PMID- 15269616 TI - Bone mass and bone metabolism in pediatric gastrointestinal disorders. PMID- 15269617 TI - Home-based treatment of malnourished Malawian children with locally produced or imported ready-to-use food. AB - OBJECTIVES: To determine the efficacy of home-based therapy with ready-to-use food (RTUF) in producing catch-up growth in malnourished children and to compare locally produced RTUF with imported RTUF for this purpose. METHODS: After a brief inpatient stabilization, 260 children with severe malnutrition were enrolled and systematically allocated to receive home therapy with either imported, commercially produced RTUF or locally produced RTUF. Each child received 730 kJ/kg/day and was followed up fortnightly. Children completed the study when they reached a weight-for-height Z score > -0.5 (WHZ), relapsed, died, or failed to achieve WHZ > -0.5 after 16 weeks. Analyses were stratified by human immunodeficiency virus (HIV) status. RESULTS: 78% of all children reached WHZ > 0.5, 95% of those with HIV-negative status and 59% of those with HIV-positive status. Eighty percent of those receiving locally produced RTUF and 75% of those receiving imported RTUF reached WHZ > -0.5. The difference between recovery rates was 5% (95% confidence interval [CI], -5-15%). The rate of weight gain was 0.4 g/kg/day (95% CI, -0.6, 1.4) greater among children receiving locally produced RTUF. The prevalence of diarrhea reported by mothers was 3.7% for locally produced RTUF and 4.3% for imported RTUF. After completion of home therapy and resumption of habitual diet for 6 months, 91% of all children maintained a normal WHZ. CONCLUSIONS: Home-based therapy with RTUF was successful in affecting complete catch-up growth. In this study, locally produced and imported RTUF were similar in efficacy in treating of severe childhood malnutrition. PMID- 15269618 TI - Effects of long-term consumption of a fermented infant formula (with Bifidobacterium breve c50 and Streptococcus thermophilus 065) on acute diarrhea in healthy infants. AB - OBJECTIVE: To determine whether long-term consumption of a fermented infant formula could influence the incidence of acute diarrhea and its severity in healthy infants. METHOD: Nine hundred seventy-one infants, ranging in age from 4 to 6 months, were included in a randomized, double-blind, placebo-controlled trial during a period of 5 months. They consumed daily either a fermented infant formula (FF) (fermentation with Bifidobacterium breve C50 and Streptococcus thermophilus 065) or a standard infant formula (SF) of the same nutritional composition. EVALUATION CRITERIA: Number and duration of acute diarrhea episodes were evaluated. Severity of the episodes was determined by the number of hospital admissions, incidence of dehydration, number of medical consultations, number of oral rehydration solution prescriptions, and number of formula switches. RESULTS: Growth of the infants and acceptability of the formulas were identical in the two groups. Incidence, duration of diarrhea episodes, and number of hospital admissions did not differ significantly between groups. Episodes were less severe in the FF (fermented formula) group. There were fewer cases of dehydration 2.5%versus 6.1% (P = 0.01), fewer medical consultations (46%v 56.6%, P = 0.003), fewer ORS prescriptions 41.9%v 51.9% (P = 0.003) and fewer switches to other formulas (59.5%v 74.9%, P = 0.0001) in FF infants compared to SF. CONCLUSION: A fermented formula may reduce the severity of acute diarrhea among healthy young infants. This outcome may be linked to the bifidogenic effects of fermentation products and their interactions with the intestinal immune system. PMID- 15269619 TI - Intestinal inflammation measured by fecal neopterin in Gambian children with enteropathy: association with growth failure, Giardia lamblia, and intestinal permeability. AB - OBJECTIVES: Investigate whether fecal neopterin concentration (a potential marker of gut inflammation) in Gambian children with enteropathy was associated with growth failure. Secondary outcome measures tested the associations between Giardia lamblia infestation, fecal neopterin and lactulose mannitol absorption ratio(L:M), a measure of intestinal permeability. METHODS: Seventy-two children had height and weight measured every 6 to 8 weeks until 15 months of age in a rural Gambian village. L:M ratio, a measure of small intestinal permeability and fecal neopterin were measured at these times. Stool was examined by immunofluorescence and light microscope for Giardia cysts. RESULTS: Long-term height and weight gains were negatively associated with mean subject fecal neopterin concentration (r = -0.29 and -0.36, respectively; P < 0.001). There was no correlation between fecal neopterin and intestinal permeability or history of diarrhea. Of 72 children studied, 19 had Giardia cysts in stool and 38 had negative stool examinations. Infected children had a mean of 0.7 days of diarrhea/week (95% confidence interval [CI], 0.31-1.03) versus 0.8 days/week (95% CI, 0.71-0.85) in uninfected children. No difference in growth was detected between those with positive or negative fecal smears. Mean L:M ratio was the same in both groups (0.31; 95% CI, 0.26-0.34). CONCLUSIONS: Consistent with the theory that intestinal inflammation in tropical infants may impair growth, fecal neopterin concentrations were inversely associated with growth. Factors other than Giardia are causing enteropathy and growth failure in Gambian infants. PMID- 15269620 TI - Portal hypertension and duodenal ulcer in children. AB - OBJECTIVES: The prevalence of duodenal ulcer (DU) in adult patients with portal hypertension is higher than in patients without portal hypertension. This study investigates the prevalence and characteristics of DU in children with portal hypertension. PATIENTS AND METHODS: From January 1997 to December 2001, 80 children with portal hypertension who had undergone upper intestinal endoscopic examinations were enrolled. Possible factors contributing to the development of DU including severity of liver disease, portal hypertension, H. pylori, and serum gastrin level were studied. The control group consisted of 80 age-and sex-matched children with gastrointestinal symptoms but no liver disease and who underwent endoscopic examination during the same period. RESULTS: The prevalence of DU was significantly higher in children with portal hypertension than in children with digestive symptoms only (22.5%v 8.8%; P =0.017). DU was more common and appeared earlier in children with a history of variceal bleeding. The presence of DU was independent of the severity of liver disease, H. pylori infection and serum gastrin level. CONCLUSION: DU occurs commonly in children with portal hypertension, especially in those who have had variceal bleeding. It is mandatory to screen a patient with gastrointestinal bleeding for DU even in the presence of esophageal varices. Elevated portal pressure might be a factor contributing to the development of DU. PMID- 15269621 TI - Glucoamylase activity in infants and children: normal values and relationship to symptoms and histological findings. AB - BACKGROUND: Starch digestion is dependent on a combination of pancreatic and salivary amylase and the intestinal brush border enzymes glucoamylase and sucrase isomaltase. Glucoamylase splits successive glucose molecules from the nonreducing end of starch molecules and is particularly important in infants who are relatively deficient in pancreatic amylase. METHODS: The authors measured glucoamylase activity in endoscopic mucosal biopsies submitted for measurement of disaccharidase activity from 214 patients aged 1 month to 20 years. Glucoamylase activity was measured using glucose polymers (polycose) as the substrate. The authors also related enzyme activity to histologic appearance and clinical indication for endoscopy. RESULTS: The most common reasons for biopsy were abdominal pain, gastroesophageal reflux, and vomiting. Disaccharidase activity by age was similar to previous reports. Glucoamylase activity did not differ with age, but was 2 to 3 times the activity reported previously. Glucoamylase activity was significantly depressed in children with the most severe histologic abnormalities. Normal glucoamylase activity (+/-2 SD) was 80.6 +/- 54.8 micromoles of glucose produced per minute per gram of protein. CONCLUSIONS: Glucoamylase activity is 2 to 3 times higher when glucose polymers are used as substrate than when glycogen is used. Severe mucosal disease is associated with reduced glucoamylase activity. Quantitation of glucoamylase activity with glucose polymers is more appropriate in evaluating children since these polymers are commonly used as carbohydrate source in the diet. PMID- 15269622 TI - Infliximab is effective in acute but not chronic childhood ulcerative colitis. AB - PURPOSE: The authors report their experience with infliximab in pediatric patients with ulcerative colitis (UC). METHODS: Fourteen patients were reviewed. Group 1 included five patients with newly diagnosed, fulminant colitis refractory to 7 to 10 days of intravenous steroids. Group 2 included four patients with ulcerative colitis in remission off steroid therapy who experienced relapse and were hospitalized with fulminant colitis refractory to intravenous steroids for 7 to 10 days. Group 3 included five patients chronically dependent on steroids with colitis refractory to medical management. All patients were treated on an open label basis with infliximab infusions of 5 mg/kg/dose at 0, 2, and 6 weeks and every 6 to 8 weeks thereafter. Follow-up was maintained for at least 6 weeks. Clinical status was scored with the Lichtiger Colitis Activity Index (LCAI) at each visit. LCAI >or=10 was considered treatment failure. We defined success as LCAI or=11 before infliximab treatment. All group 1 patients experienced response to infliximab. All but one (75%) patient in group 2 had a response. Only one (20%) group 3 patient had a response to infliximab. CONCLUSION: Infliximab was an effective agent in the treatment of acute UC in our patients. Long-term steroid use and emergency colectomy were avoided. Infliximab was less effective in patients who were dependent on steroids. PMID- 15269623 TI - Partial protection against dextran sodium sulphate induced colitis in histamine deficient, histidine decarboxylase knockout mice. AB - OBJECTIVES: Chemically induced mucosal inflammation in animal models is a suitable tool for studying factors in the pathogenesis of inflammatory bowel disease. The aim of this study was to determine whether absence of histamine has an effect on the development of experimental colitis. METHODS: Histamine deficient, histidine decarboxylase (HDC) knockout Balb/c mice and genetically identical control animals with intact HDC were studied. Colitis was induced by the administration of 2% dextran sodium sulphate in drinking water. Mice were killed after 5 days and disease activity assessed by clinical, histologic, and immunohistologic parameters. Bacterial components of stool were examined. RESULTS: Clinical disease activity was higher in the mice with intact HDC (disease activity index, 2.21) than in the histamine-deficient knock-out mice (1.88). Histologic findings were similar in the two groups. On day 5, the inflammation score of the HDC sufficient group was 5.25 (+/-1.055) and the crypt score was 5.00 (+/-1.128). The scores in the HDC knock-out group were 4.667 (+/- 0.707) and 4.667 (+/- 0.86), respectively. There was a significant difference in the number of interleukin (IL-10)-producing lymphocytes in colon mucosa. Large numbers of IL-10-positive lymphocytes were observed in wild type mice both those with DSS induced colitis and untreated controls. Only sporadic IL-10 positivity was found in histamine-deficient mice. Significant differences were found in the composition of the fecal bacterial flora between the two groups. CONCLUSION: The reduced number of IL-10-positive lymphocytes in the intestinal mucosa of histamine-deficient, histidine decarboxylase knockout mice and the altered fecal bacterial flora in these animals suggest that histamine may play a role in the pathophysiology of inflammation in the colon of normal animals by upregulating local IL-10 production and stimulating a local shift to Th2 response. PMID- 15269624 TI - Menetrier disease of early infancy: a separate entity? PMID- 15269625 TI - Comparison of longstanding pediatric-onset and adult-onset Crohn's disease. AB - INTRODUCTION: Limited information is available on the characteristics of longstanding Crohn's disease with onset in childhood or adolescence. MATERIALS AND METHODS: In this retrospective study, 224 patients with pediatric-onset Crohn's disease were compared to a group of patients with disease diagnosed as adults. RESULTS: More than 50% of the patients in the pediatric-onset group were followed for more than a decade, with a mean follow-up of 12.2 years. There were 96 male (42.9%) and 128 female (57.1%) patients. Most had disease diagnosed in adolescence. The female predominance, similar to adult-onset disease, was noted only among patients with disease diagnosed in adolescence (13-19 years), not in childhood. Disease most often involved both ileum and colon (128 of 224; 57.1%). Isolated ileal or colonic disease was less common. Upper gastrointestinal tract disease was present in 42 of 224 (18.8%) patients. The incidence of strictures (28.6%) and penetrating complications (46.4%) was similar to that of adults followed for the same period of time. CONCLUSION: In pediatric-onset Crohn's disease, female-predominance emerges during adolescence. Pediatric-onset disease is more extensive than adult-onset disease. When followed for an extended period of time, it has high rates of disease complexity, with strictures and penetrating complications similar to adult-onset disease. PMID- 15269626 TI - Gallbladder motility in children with Down syndrome. AB - OBJECTIVE: The aim of this study was to investigate gallbladder motility in children with Down syndrome by measuring gall-bladder volume and contraction index. METHODS: This study, performed between January 2001 and December 2002 at the Ondokuz Mayis University, School of Medicine, Department of Paediatric Neurology, Samsun, Turkey, included 21 patients with Down syndrome (study group) and 22 healthy children (control group). After an 8-hour fast, gallbladder diameters in both groups were measured in length, width, and height by ultrasonography before and 30 minutes after a test meal. The volume of gallbladder before and after a test meal was determined, and the contraction index was calculated. Blood triglyceride and cholesterol levels were measured, and 5-hydroxyindoleacetic acid (5-HIAA) levels in urine were determined. RESULTS: Mean gallbladder volume before test meal in the study group and controls was 8,412.4 +/- 5,174 mm and 16,516.8 +/- 6,796.1 mm (P < 0.001), respectively. The mean contraction index of the study group was 41.2% +/- 19.4% and of controls, 75.0% +/- 12.3% (P < 0.001). The mean triglyceride level of the study group was significantly higher than controls (P < 0.05). The mean urine 5-HIAA level of the study group was lower than controls (P < 0.05). CONCLUSION: CI was lower in patients with Down syndrome, suggesting gallbladder hypomotility. Hypomotility may be a feature associated with the high prevalence of gallstones in Down syndrome. PMID- 15269627 TI - Treatment of functional abdominal pain in childhood with cognitive behavioral strategies. AB - OBJECTIVE: The aim of this study was to assess the efficacy of a cognitive behavioral approach to the treatment of recurrent abdominal pain caused by childhood functional gastrointestinal disorders (FGIDs). METHODS: From September 2001 to December 2002, 18 patients (12 male; mean age, 12.1 +/- 4.9 years) with chronic abdominal pain (mean duration, 11.8 +/- 13.3 months) caused by FGIDs were referred to our facility's mind-body institute (MBI). Treatment included guided imagery and progressive relaxation techniques. The mean number of sessions per patient was 4.3 +/- 3.4. Outcomes included change in abdominal pain and quality of life, evaluated by the Pediatric Quality of Life Scale (PedsQL). Follow-up was 10.6 +/- 2.3 months after the last MBI session. RESULTS: Abdominal pain improved in 89% of patients; weekly pain episodes decreased from 5.5 +/- 0.9 to 2.0 +/- 2.7 (P < 0.05); pain intensity (0 to 3 scale) decreased from 2.7 +/- 0.6 to 0.6 +/- 0.7 (P < 0.04); missed school days/month decreased from 4.6 +/- 1.7 to 1.4 +/ 3.2 (P < 0.05); social activities/week increased from 0.3 +/- 0.6 to 1.3 +/- 0.6 (P < 0.05); physician office contacts/year decreased from 24 +/- 10.2 to 8.7 +/- 13.1 (P = 0.07). PedsQL scores (0 to 100 scale) improved from 55.3 +/- 11.9 to 80.0 +/- 10.7 (P < 0.03). CONCLUSIONS: Guided imagery and progressive relaxation can safely and effectively reduce chronic abdominal pain in children with FGIDs. This treatment also improved social functioning and school attendance. PMID- 15269628 TI - Polyethylene glycol for constipation in children younger than eighteen months old. AB - BACKGROUND: Polyethylene glycol (PEG) is a safe and effective treatment for constipation in children older than 18 months. Data on its safety and efficacy in infants are lacking. The goal of this study was to determine safety, efficacy, and optimal dose of polyethylene glycol powder for treatment of constipation in patients younger than 18 months. METHODS: The authors reviewed the charts of patients younger than 18 months treated with PEG 3350 for constipation. The initial dose, effective maintenance dose, response to therapy, duration of therapy, and side effects were recorded. RESULTS: Twenty-eight patients younger than 18 months of age treated with PEG were identified (3, age 0-5 months; 9, age 6-11 months; 16, age 12-17 months). Mean duration of therapy was 6.2 +/- 5 months (range, 3 weeks-21 months). Mean initial dose was 0.88 g/kg/day (range, 0.26-2.14 g/kg/day). Mean effective maintenance dose was 0.78 g/kg/day (range, 0.26-1.26 g/kg/day). PEG relieved constipation in 97.6% of patients. One infant experienced increased gas per rectum and four others experienced transient diarrhea that resolved after adjusting the dose. CONCLUSION: Oral powdered polyethylene glycol at a maintenance dose of 0.78 g/kg/day is safe and effective for patients younger than 18 months. Dose and safety profiles are similar for those reported in older children. PMID- 15269629 TI - Acute hepatic crisis in children with sickle cell disease. PMID- 15269630 TI - Development of Crohn disease during anti-TNF-alpha therapy in a child with juvenile idiopathic arthritis. PMID- 15269631 TI - Lack of benefit of gluten-free diet on autoimmune hepatitis in a boy with celiac disease. PMID- 15269632 TI - Leiomyomatosis of the oesophagus. PMID- 15269633 TI - Vegetarian diet and exocrine pancreatic function using fecal tests. PMID- 15269635 TI - Sweat test results in children with primary protein energy malnutrition. PMID- 15269636 TI - High prevalence of unrecognized celiac disease in an unselected hospital population in north-eastern Brasil (Recife, Pernambuco). PMID- 15269637 TI - Serum homocysteine concentrations in children with inflammatory bowel disease. PMID- 15269638 TI - CARD 15 / NOD 2 in paediatric onset Crohn's disease. PMID- 15269639 TI - Patchy duodenal atrophy or proximal duodenal involvement by celiac disease? PMID- 15269641 TI - Complications of childhood Peutz-Jeghers syndrome: implications for pediatric screening. PMID- 15269642 TI - Neonatally acquired hepatitis C--not so risky? PMID- 15269643 TI - Tackling obesity: we need a hand from Nanny. PMID- 15269644 TI - Cross-boundary working: a generic worker for older people in the community. AB - The care of older people often crosses the boundaries of health and social care. The new role of a health and social care trained generic worker was developed to provide comprehensive care for older people living at home. The role is a cross between a nursing auxiliary, health care assistant and a community support worker. The evaluation of the one-year pilot project demonstrated that clients were very satisfied with the care they received, particularly the emotional aspects of care. A high proportion of the generic workers time was spent listening and responding to their clients' mental health needs, and providing comfort and emotional support. Having been trained by local health professionals, the generic workers felt valued and respected, better able to communicate with their health colleagues, and therefore able to provide holistic care to their clients. PMID- 15269645 TI - Family health nursing: the education programme for the WHO Europe Scottish Pilot. AB - This article outlines the development of the family health nurse (FNH) programme, which was delivered by the University of Stirling in the highlands and islands of Scotland as part of a World Health Organization European pilot project. An outline of the structure of the programme and its key features is described. The concept of the FHN emerged from the WHO's initiative to develop a practitioner who has the family as the organizing focus of their practice (WHO, 2000). An insight is provided into the experience of the first students to undergo this programme, along with a brief summary of the main findings of the external evaluation of both the education programme and the implementation of the role in the remote and rural communities of the highlands and islands of Scotland. Suggestions are made that will hopefully influence the second phase of this project that the Scottish Executive are supporting in an urban setting, which is due to begin in September 2004. PMID- 15269646 TI - Essence of Care: Implementing continence benchmarks in primary care. AB - 'Essence of Care' is a national programme which was launched in 2001. The document provides a set of national benchmarks which are evidence based. The benchmarks focus on basic care as identified in a First Class Service (DH, 1998). The programme has been implemented widely by acute services, however with community and primary care services, the take up appeared to have been spasmodic. A conference in 2003 'Implementing Essence of Care in Primary Care' described the implementation of Essence of Care in Primary Care as a 'challenge'. This article identifies the implementation of the continence benchmark within a PCT. The article details the process and the difficulties encountered within primary care, but also details the benefits of utilising the model despite a labour intensive process. In particular the article suggests the importance of incorporating the benchmarks into current systems within the PCT in order to avoid duplication and extra layers of groups and meetings. Finally the article highlights some of the benefits which the PCT consider the process has achieved to the continence service. PMID- 15269647 TI - The law of negligence and community nursing: a case study. PMID- 15269648 TI - The new NICE guidelines on COPD. AB - The National Institute of Clinical Excellence guidelines on chronic obstructive pulmonary disease, published in March this year, provide a new way of thinking about the management of the disease. This article highlights the key areas and recommendations made by the guidelines. PMID- 15269649 TI - Time for a radical rethink of nurse education. PMID- 15269650 TI - Reflection is the way to improve practice. PMID- 15269651 TI - Priorities in nursing management of fistulas in a community setting. AB - The aim of this article is to discuss care in the community of patients with a fistula. Areas considered include definition and classification of fistulae, potential causal factors and associated diseases. The article concentrates on enterocutaneous fistulation and the care of patients, but also includes a summary about caring for a patient with an anal fistula. The essential elements of nursing in a community setting are described, and information given on skin care, nutrition, fluids and appliances. PMID- 15269652 TI - The Lindsay Leg Club Model: a model for evidence-based leg ulcer management. AB - Leg Club is a unique model of community-based leg ulcer care. By providing nursing care in a non-medical, social environment, the model has several benefits: it removes the stigma associated with leg ulcers and helps isolated older people reintegrate into their communities, which in turn improves concordance and has a positive impact on healing and recurrence rates. In an atmosphere of de-stigmatisation, empathy and peer support, positive health beliefs are promoted and patients take ownership of their treatment. The Leg Club model creates a framework in which nurses, patients and local community can collaborate as partners in the provision of holistic care. The model also provides an environment for appropriate supportive education, advice and information. PMID- 15269653 TI - Evaluation of a honey-impregnated tulle dressing in primary care. AB - Honey has been used for its healing properties for centuries and has been used to dress wounds with favourable results. The emergence of antibiotic resistance and growing interest in "natural" or "complementary" therapies has led to an interest in honey dressings. Much of the research to date has been related to honey's antibacterial properties. However, the healing properties claimed for honey also include stimulating new tissue growth, moist wound healing, fluid handling and promoting epithelialization. Until recently, honey had not been developed as a wound management product and was not a certified pharmaceutical device. Activon Tulle is a sterile, non-adherent dressing impregnated with Leptospermum scoparium hone. The claimed properties of honey dressings would make this a valuable addition to the dressing currently available in the primary care setting. An evaluation was undertaken involving 20 patients with a variety of wounds. A conclusion is drawn that while further research is needed, medical grade honey does appear to be a valuable addition to the wound management formulary. PMID- 15269655 TI - [Genetic diversity of voltage-gated calcium channels]. AB - Understanding of the properties of normal and diseased voltage-dependent calcium channels has greatly improved these last Years after the extensive development of the patch-clamp and molecular biology studies and the functional expression strategies. The calcium channel diversity is based on the expression of numerous genes that encode pore channel subunits (10 genes) and auxiliary/regulatory subunits (16 genes). In addition, most of these genes are subject to alternative splicing. The study of calcium channels has also benefited from the discovery of genetic diseases linked to calcium channel mutations: the calcium channelopathies. The review describes the recent data and working hypothesis that address the challenging question of how the calcium channel diversity occurs and how alterations in channel function lead to selective cellular dysfunction. PMID- 15269656 TI - [Ion channels and demyelination: basis of a treatment of experimental autoimmune encephalomyelitis (EAE) by potassium channel blockers]. AB - Voltage-gated potassium channels (Kv channels) are ion channels, openings of which provide an outward flow of potassium ions repolarising the cell. In neurons, Kv channels play a crucial role in action potential repolarisation and in shaping neuronal excitability. In non-excitable cells, such as T lymphocytes, Kv channels and calcium-activated K+ channels (KCa channels) determine the driving force for Ca2+ entry. During T cell activation the calcium entry depolarises the cell and increases the cytosolic calcium concentration, which in return activates Kv and KCa channels. K+ channel opening repolarises the cell and drives the membrane potential to a negative voltage. The roles of Kv channels in nervous and immune systems have been investigated here by means of a rat experimental autoimmune disease of the central nervous system, the experimental autoimmune encephalomyelitis (EAE). EAE is characterised clinically by paralysis, and pathologically by inflammatory cell infiltrations into the brain and the spinal cord. Among the inflammatory cells, T lymphocytes play a major role. Hence, EAE can be adoptively transferred into syngenic animals by the injection of T cells reactive to myelin antigens. During adoptive-EAE, somato-sensory evoked potentials recorded along the spinal tracts decrease in amplitude and axonal propagation is disrupted. We have analysed the consequences of Kv channels blockade by peptidyl toxins on central nerve conduction, on T cell activation and on the time course of EAE. In rat optic nerves, Kv channels have been identified up from postnatal day 1. Their blockade by kaliotoxin (a scorpion toxin) or by dendrotoxin-I (a snake toxin) enlarges the compound action potentials, demonstrating the participation of Kv channels to spike repolarisation. This effect disappears at adult age due to the sequestration of Kv channels under the myelin, in the paranodal regions. During acute demyelination by lysophosphatidyl choline, the surface area of compound action potential decreased probably because conduction block occurred. Demyelination unmasked Kv channels, which are again accessible to toxins. Their blockade by dendrotoxin-I or kaliotoxin favoured a slow delayed conduction suggesting that those Kv channel blockers exert a neurological benefit during demyelinating diseases. In a T-cell line reactive to myelin basic protein antigen, which is used to adoptively transfer experimental autoimmune encephalomyelitis, Kv1.3 channels are constitutively expressed. Their blockade leads to a pronounced reduction of the T cell proliferative response, cytokine production and Ca2+ influx. In the rat, blockade of Kv1.3 inhibits the delayed type hypersensitivity response to myelin basic protein prevents and treats adoptive experimental autoimmune encephalomyelitis. Blockade of Kv channels alone or in combination with KCa channels improves the symptoms of the disease. These results demonstrate that K+ channel blockers displaying high selectivity are potent immunosuppressive agents with beneficial symptomatic effects in experimental autoimmune encephalomyelitis. PMID- 15269657 TI - [Dosage and specificity of anti-calcium channel antibodies in Lambert-Eaton myasthenic syndrome]. AB - Lambert-Eaton myasthenic syndrome (LEMS) is an autoimmune channelopathy in which patients produce autoantibodies directed against voltage-gated calcium channels. Autoantibodies down-regulate calcium channels resulting in reduced transmitter release, which in turn leads to muscular weakness and autonomic dysfunction. LEMS is paraneoplastic in 60-70% of patients, most frequently associated with small cell lung carcinoma (SCLC). SCLC lines express many neuronal and neuroendocrine proteins including neuronal calcium channels of the Cav2 family (P/Q and N-type channels). It is thus likely that the paraneoplastic form of LEMS is the consequence of an anti-tumoral immune response and the production of antibodies that cross-react with identical or homologous antigens in nerve terminals. Neurological symptoms generally appear several Months before detection of the tumor. Consequently correct diagnosis of LEMS is crucial as it can allow early treatment of a particularly aggressive carcinoma. Based on published studies, our laboratory has set-up serological assays for LEMS autoantibodies as an aid to diagnosis. Calcium channels in detergent extracts of rat brain or cerebellum membranes were labeled with radioligands specific for N-type (125I-omega conotoxin GVIA) or P/Q-type (125I-omega conotoxin MVIIC) calcium channels. Autoantibodies that immunoprecipitate the ligand/channel complex can thus be titrated. Analysis of 31 LEMS sera revealed the presence of anti-N type channel antibodies in 58% and anti-P/Q type channel antibodies in 74% of patients with titres ranging from 90 to 2950 pM. Only 5 patients were seronegative in both tests, thus a combination of the two assays reliably detected autoantibodies in 26/31 (84%) patients. PMID- 15269658 TI - [Muscular genetic disorders caused by abnormal membrane excitability]. AB - Ion channels are transmembrane proteins which enable ion exchanges between the inner and the outer part of the cell. During evolution, the property of ligand- or voltage-gating conferred cell excitability which permitted intercellular communication. The study of muscle diseases, periodic paralysis and myotonia, has led to the discovery of mutations in the genes encoding ion channels. The analysis of the functional consequences on muscle membrane gave insight into pathophysiology. A loss of function of sodium or calcium channels leads to hypokalaemic periodic paralysis. A gain of function of sodium channel results in hyperkalaemic periodic paralysis or paramyotonia, depending of its level. It is also known that mutations in other genes can cause membrane excitatibility such as the gene encoding perlecan (Schwartz-Jampel syndrome). The study of muscle channelopathies has opened a new field in neurological disorders. Molecular diagnosis is now possible and the efficacy of treatments is better understood. PMID- 15269659 TI - [Andersen syndrome: a particular form of paralysis with cardiac dysrhythmia]. AB - Andersen syndrome includes a clinical triad with periodic paralysis, cardiac arrhythmia and dysmorphic features most often mild but relevant. It is a potassium channelopathy due to mutation of KCJN2 gene coding for Kir 2.1 protein. We report a familial case with mutation R218W of Kir 2.1 and discuss the main phenotypic and genetic aspects of Andersen syndrome. Muscle manifestations are essentially a periodic paralysis most often of hypokaliemic type. Muscle biopsy reveals tubular aggregates but can be normal as it is shown in the same patient in our kindred. Our proband complained of paralytic attacks since childhood and at adult age she demonstrated a mild permanent deficit of pelvic girdle muscles as it has been described in other types of periodic paralysis after a long duration course. Cardiac manifestations may include in a variable manner a long QT syndrome, premature ventricular contractions, complex ventricular ectopy, polymorphic or bidirectional ventricular tachycardia. Imipramine had a positive effect on arrhythmia in our case. Dysmorphic features are often mild and have to be cautiously looked for as a clue to the diagnosis of Andersen syndrome. They can be easily overlooked if not systematically looked for. Clinical expressivity is variable including in the same family. In our observation, the daughter showed a complete triad, early expressed, which allowed the diagnosis. Her father was late diagnosed on ventricular dysrhytmia but without muscle manifestations and dysmorphic features. Since KCJN2 gene mutation identification, locus heterogeneity of Andersen syndrome was shown. Andersen syndrome kindreds without mutations in KCNJ2 were clinically indistinguishable from KCNJ2-associated subjects. KCNJ2 gene encodes the inward rectifier K+ channel Kir2.1 which plays an important role in maintaining membrane potential and during the terminal phase of cardiac action potential repolarization. Several studies showed a dominant negative effect of the mutation on Kir 2.1 channel function. PMID- 15269660 TI - Electrophysiology and molecular pharmacology of muscle channelopathies. AB - As voltage-gated ion channels are essential for membrane excitation, it is not surprising that mutations in the respective channel genes cause diseases characterised by altered cell excitability. Skeletal muscle was the first tIssue in which such diseases, namely the myotonias and periodic paralyses, were recognised as ion channelopathies. The detection of the functional defect that is brought about by the disease-causing mutation is essential for the understanding of the pathology. Much progress on the road to this aim was achieved by the combination of molecular biology and electrophysiological patch clamp techniques. The functional expression of the mutations in expression systems allows to study the functional alterations of mutant channels and to develop new strategies for the therapy of ion channelopathies, e.g. by designing drugs that specifically suppress the effects of malfunctioning channels. PMID- 15269661 TI - [Electrophysiological testing in muscle channelopathies]. AB - Electrodiagnostic testing should always be tailored to the clinical setting; in the muscle channelopathies, provocative tests are essential to demonstrate the modifications of the muscle excitability. When a permanent muscle weakness is present, a post-exercise (or post-tetanic) potentiation should be search to demonstrate a presynaptic neuromuscular channelopathy; when the weakness is fluctuating it is useful to perform repetitive nerve stimulation to see if a decrement is present or if the jitter is prolonged, indicative of a postsynaptic neuromuscular channelopathy; when the weakness is episodic, the prolonged exercise test is the only test to demonstrate the appearance of a late post exercise decrement (sodium and calcium channelopathies), and when a myotonic reaction is seen to search the myotonic discharges by needle emg. The concomitant presence of myotonic discharges and abnormal decrement of the motor responses during repetitive nerve stimulation train depend on the type of mutations in the chloride gene and the amount of CTG repeats in myotonic dystrophy type 1. In paramyotonia congenita there a prolonged decrement when the tests are carried out with cooling the recorded muscle. Electrophysiological testings appear therefore useful to diagnose muscle channelopathies but also give information about the prognosis of the disorder; they could also be viewed as useful tests to predict response to treatments. PMID- 15269662 TI - Treatment in myotonia and periodic paralysis. AB - The myotonic disorders, including the myotonic dystrophies (myotonic dystrophy type 1, DM1; myotonic dystrophy type 2, DM2/PROMM/PDM), the muscle channelopathies or non-dystrophic myotonias (chloride, sodium, calcium and potassium channelopathies) are all characterized by myotonia and muscle weakness despite different pathophysiology involved in these disorders. Myotonia may affect the eye, facial and jaw muscles as well as the hands and legs. It may be painful and disabling. Muscle weakness may be episodical as in the paralytic attacks of the sodium and calcium channelopathies or culminate in permanent muscle weakness as in the calcium channelopathies and some sodium channelopathies associated to specific point mutations. The severity of myotonia may fluctuate in the myotonic dystrophies, but weakness is usually fixed, affecting neck flexors, facial and jaw muscles as well as proximal and distal muscles of the limbs. Despite the recent progress in molecular genetics the precise mechanisms responsible for myotonia and weakness are not fully understood and there is no standardized treatment strategy. We present a review of selected treatment trials in the myotonic disorders and the muscle channelopathies, and discuss our experience in the treatment of myotonia and muscle weakness, with reference to the limits and advantages of treatment trials in this field. PMID- 15269663 TI - [Genetic of diseases by abnormal functioning of the skeletal muscle-calcium releasing complex]. AB - Myoplasmic calcium homeostasis is an essential feature of skeletal muscle contraction. The calcium mobilisation complex (CMC) located at the level of the triadic junction plays a major role for the regulation of calcium fluxes between extra-cellular, cytoplasmic and intra-cellular compartments. The ryanodine receptor type I (RYR1), which is located at the level of the terminal cisternae of the sarcoplasmic reticulum is a key component of the CMC. RYR1 allow the release into the myoplasm of the intralumenal stores of calcium. RYR1 interacts with other proteins: DiHydroPyridine Receptor, triadin, calsequestrin, FKBP12, calmodulin. Malignant hyperthermia (MHS) and congenital core myopathies have been associated with a dysfunction of the CMC. MHS is an autosomic dominant pharmacogenetic disease. The MH crisis is induced by exposure of the predisposed patients to halogenated volatile anaesthetics. MHS is characterised by a genetic heterogeneity and two genes, RYR1 and CACNA1S, have been associated so far with the disease. Mutations in the RYR1 gene have been recently associated with heat stroke, a related syndrome. Central Core Disease (CCD) and Multi minicore Disease (MmD) are congenital myopathies presenting with clinical variability and characterized by the presence of specific although heterogeneous muscle histological features: the cores. Clinical boundaries between the two diseases may overlap and the specific diagnosis is often based on the nature of the cores. These diseases show genetic heterogeneity with both autosomic dominant and recessive mode of inheritance and mutations in the SEPN1, RYR1, ACTA1, TPM3 genes have been reported. Mutations associated with MHS were mainly identified into 2 regions of the N-terminal part of RYR1. Functional role of these two domains is still unclear. Mutations responsible for congenital myopathies mainly mapped to the C terminal region of RYR1 that form the transmembrane calcium channel. Functional studies of the RYR1 mutations have shown that MHS mutations were mainly associated with an alteration of the calcium fluxes in response to caffeine or halothane while CCD mutations would result in a leaky RYR1 channel or would alter the Excitation-Contraction coupling at the level of the CMC. PMID- 15269664 TI - [Congenital myasthenic syndromes due to mutations in the rapsyn gene]. AB - Congenital myasthenic syndromes (CMS) are genetic diseases characterized by dysfunctional neuromuscular transmission and usually start during the neonatal period. Most are due to postsynaptic abnormalities, specifically to mutations in the acetylcholine receptor (AChR) genes. In 2002, the group of A Engel reported the first cases of CMS with mutations in the gene coding rapsyn, a postsynaptic molecule which stabilizes AChR aggregates at the neuromuscular junction. Since this first publication, more than 30 other cases, including six in France, have been reported. Study of these published cases allows us to distinguish three classes of phenotypes: 1) severe neonatal cases; 2) more benign cases, starting during infancy; 3) cases with facial malformations, involving Jewish patients originating from the Near-East. Comparison of the observations of other groups with our own has led us to the following conclusions: the N88K mutation is frequent (homozygous in 50% of cases); besides the N88K mutation, the second mutation varies considerably; heterozygous allelic cases (N88K + another mutation) are severe; there is probably a founder effect in the European population. There is phenotypic variability in the homozygous N88K cases, with benign cases and severe cases of early expression. A Engel and colleagues report that the seven cases of benign CMS with facial malformation, previously described in the Jewish population of Iraq and Iran, were caused by mutation in the promoter region of the rapsyn gene. PMID- 15269665 TI - Neuromyotonia. AB - There is increasing evidence that autoimmunity is implicated in the pathogenesis of peripheral nerve hyperexcitability (neuromyotonia, NMT and Cramp-fasciculation syndrome C-FS ) and in Maladie de Morvan in which CNS features are also present. All three conditions can associate with thymoma, myasthenia gravis and other autoimmune disorders, and can often respond to plasma exchange. In NMT, patient's plasma or IgG can transfer the electrophysiological features to mice, and can reduce voltage-gated potassium channel currents in vitro. Antibodies to voltage gated potassium channels can be detected in the serum of many patients who have peripheral nerve hyperexcitability, and also in those with Maladie de Morvan. These latter patients have clinical features similar to limbic encephalitis in which VGKC antibodies can also occur. Thus neuromyotonia, cramp-fasciculation syndrome and Maladie de Morvan can occur as antibody-mediated autoimmune ion channelopathies like myasthenia gravis and the Lambert-Eaton myasthenic syndrome. These discoveries should aid diagnosis and offer new approaches to treatment. PMID- 15269666 TI - [Recent insights into the implication of ion channels in familial forms of epilepsies associated or non associated to febrile convulsions]. AB - Major advances have recently been made in the understanding of the genetic bases of monogenic inherited epilepsies. For several idiopathic epilepsies, mutations in genes encoding subunits of ion channels or ligand receptors have been demonstrated. This is the case for some generalized idiopathic epilepsies and generalized epilepsies associated with febrile seizures. In this Article, we review the recent clinical and genetic data of these forms of epilepsy. PMID- 15269667 TI - [Indications for how and when to treat low grade gliomas]. PMID- 15269668 TI - [Deep brain stimulation]. AB - The present renewal of the surgical treatment of Parkinson's disease, almost abandoned for twenty Years, arises from two main reasons. The first is the better understanding of the functional organization of the basal ganglia. It was demonstrated in animal models of Parkinson's disease that the loss of dopaminergic neurons within the substantia nigra, at the origin of the striatal dopaminergic defect, induces an overactivity of the excitatory glutamatergic subthalamo-internal pallidum pathway. The decrease in this hyperactivity might lead to an improvement in the pakinsonian symptoms. The second reason is the improvement in stereotactic neurosurgery in relation with the progress in neuroimaging techniques and with intraoperative electrophysiological microrecordings and stimulations, which help determine the location of the deep brain targets. In the 1970s chronic deep brain stimulation in humans was applied to the sensory nucleus of the thalamus for the treatment of intractable pain. In 1987, Benabid and colleagues suggested high frequency stimulation of the ventral intermediate nucleus of the thalamus in order to treat drug-resistant tremors and to avoid the adverse effects of thalamotomies. How deep brain stimulation works is not well known but it has been hypothetized that it could change the neuronal activities and thus avoid disease-related abnormal neuronal discharges. Potential candidates for deep brain stimulation are selected according to exclusion and inclusion criteria. Surgery can be applied to patients in good general and mental health, neither depressive nor demented and who are severely disabled despite all available drug therapies but still responsive to levodopa. The first session of surgery consists in the location of the target by ventriculography and/or brain MRI. The electrodes are implanted during the second session. The last session consists in the implantation of the neurostimulator. The ventral intermediate nucleus of the thalamus was the first target in which chronic deep brain stimulation electrodes were implanted in order to alleviate tremor. This technique can be applied bilaterally without the adverse effects of bilateral thalamotomies. Like pallidotomy, internal globus pallidum stimulation has a dramatic beneficial effect on levodopa-induced dyskinesia but its effects on the parkinsonian triad are less constant and opposite motor effects are sometimes observed in relation with the stimulated contact. The inconstant results, perhaps related to the complexity of the structure led to the development of subthalamic nucleus stimulation. The alleviation of motor fluctuations and the improvement in all motor symptoms allows a significant decrease in levodopa daily dose and in levodopa-induced dyskinesia. Presently, deep brain stimulation is a fashionable neurosurgical technique to treat Parkinson's disease. Subthalamic nucleus stimulation seems to be the most suitable target to control the parkinsonian triad and the motor fluctuations. Because of the possible adverse effects it must be reserved for disabled parkinsonian patients. No large randomized study comparing different targets and different neurosurgical techniques has been performed yet. Such studies, including cost benefit studies would be useful to assess the respective value of these different techniques. PMID- 15269669 TI - [The blood-brain barrier should not be underestimated in neuro-oncology]. AB - INTRODUCTION: Limited therapeutic success in the treatment of CNS neoplasia with chemotherapy is generally attributed to two factors: natural or acquired resistance to chemotherapy expressed by tumor cells, and delivery impediment related to the blood-brain barrier. STATE OF ART: The anatomic and physiological properties of the normal blood-brain barrier prevent passage of ionized water soluble compounds with a molecular weight greater than 500 Daltons. Although complex, the blood-brain barrier basically functions at the level of the tight junctions of the cerebral vascular endothelial cells. Different approaches have been advocated to improve delivery across the blood-brain barrier. One such approach, transient osmotic permeabilization of the blood-brain barrier, is an invasive procedure offering the potential of global delivery. This strategy involves cerebral intravascular infusion of a hypertonic solution to produce, in a given cerebral distribution (carotid or vertebral), producing a transient increase in blood-brain barrier permeability. Two parameters are paramount in the ability to mediate a hyperosmolar modification of the barrier: the osmolality of the solution, and the infusion time. The procedure has been found to produce a marked increase (10- to 100-fold) in brain and cerebrospinal fluid concentrations of methotrexate and of other markers. PERSPECTIVES: Pre-clinical studies are underway to assess the use of this procedure to improve delivery of different molecules, including standard chemotherapy, monoclonal antibodies and gene therapy molecules. This approach has been standardized for clinical use. It has been extensively used in patients. Using a standard protocol of osmotic blood brain barrier disruption to enhance chemotherapy delivery with three different chemotherapy regimens, more than 3000 procedures have been performed in more than 300 patients across the blood-brain barrier distribution consortium, an entity which includes six university centers coordinated by the Oregon Health Sciences University. The procedure has been found to be safe, with very limited toxicity. As part of this consortium, the Sherbrooke University center has been offering this treatment modality since November 1999. We have performed more than 500 procedures in 122 patients with various histologies (malignant gliomas, primary central nervous system lymphoma and metastasis) with low toxicity. CONCLUSIONS: In our view, the median survival rate of 138 weeks obtained with glioblastoma multiforme patients is promising; further research to improve these results is needed. PMID- 15269670 TI - [Neoadjuvant chemotherapy for symptomatic non operable grade II fibrillary astrocytoma in adults]. AB - We collected 6 case-reports of symptomatric non removable low grade fibrillary astrocytoma of adults treated with a procarbazine-CCNU-vincristine chemotherapy regimen. All patients had drug-resistant epilepsy but brain imaging was stable. Total gross resection was rejected because of Volume or tumor location. After 4 to 7 cycles of chemotherapy, 2 patients had partial response and one minor response on brain MRI. All of them were seizure-free. Progression free survival was not reached at 5 Years. Up-front chemotherapy for low-grade astrocytomas may be useful and has to be prospectively evaluated. PMID- 15269671 TI - [Results of salvage stereotactic radiosurgery in 14 patients with grade III or IV gliomas]. AB - AIMS: To determine local control and overall survival rates of 14 patients treated for a grade III or IV glioma relapsing in a previously irradiated area and re-irradiated by stereotactic radiosurgery. PATIENTS AND METHODS: From January 1997 to October 2001, 14 patients (median age 52 Years, age range 49-58 Years, Karnofski performance score 80 to 100) received radiosurgery for a relapse of grade III (3 patients) and or grade IV (10 patients) malignant gliomas. Before relapse, all patients had undergone surgery and had been given with a classical radiation protocol. Median maximum diameter and Volume of the tumors were 38.5mm (24-86mm) and 7cm3 (2-35cm3), respectively. RESULTS: Median maximal dose at the isocenter and median minimal dose at the periphery of the lesion were 21Gy (16 38Gy) and 13Gy (9-17Gy), respectively. Mean follow-up was 8.5 Months (1-29). Median overall survival was 11.6 Months; 6-Month, 1- and 2-Year overall survival rates were 85p.100, 36p.100 and 12p.100, respectively. At univariate analysis, only histological grade was a significant prognostic factor of overall survival (p=0.03). Median disease-free survival was 8.2 Months while 6-Month and 1-Year disease-free survival rates were 69p.100 and 14p.100, respectively. According to univariate analysis, histological grade (p=0.033) and minimal dose delivered at the margin of the target Volume (p=0.02) were prognostic factors for disease-free survival. Two patients developed a symptomatic radionecrosis. CONCLUSION: Radiosurgery of relapsed primitive high-grade brain tumors is efficient and overall survival rates were encouraging. PMID- 15269672 TI - [Usefulness and limitations of polymygraphic recordings in dystonia]. AB - INTRODUCTION: Diagnosis of dystonia is based on well-known clinical findings, but electrophysiological explorations can help to establish the diagnosis. METHODS: We report 179 patients presenting primary or secondary dystonia. Polymyographic recordings were available for all. We studied activation patterns of agonist and antagonists muscles (prolonged bursts, co-contraction), presence of antagonistic gestures and other associated movement disorders such as tremor or myoclonus. RESULTS: 83.9 percent of patients presented abnormalities which evoked dystonia (prolonged bursts: 75.3 percent, co-contraction: 48.7 percent, successful antagonistic gesture successful with reduction or suppression of EMG activity, 8.7 percent). For the patients who presented no EMG abnormality, diagnosis was maintained in 22 cases because of the limitations of the exploration: intermittent symptoms, dystonia involving deep muscles which were not recorded by the surface electromyogram, or difficulties to reproduce the conditions of apparition of the dystonic symptoms during examination or to recognize a pathologic EMG pattern in hand cramps for instance. Another diagnosis was retained because of the polymyographic findings in 16.2 p.cent of these patients, particularly those who had tremor associated with dystonia (essentially diagnosis of primary writing tremor). CONCLUSION: Polymyography is an effective tool for confirming clinical findings. PMID- 15269673 TI - [A principal component analysis of the AGGIR scale in demented elderly patients]. AB - AGGIR grid is the national standardized instrument determining aimed at the dependency of old people in France living in institutions as well as in the community. Attribution of the governmental financial assistance APA (Allocation Personnalisee d'Autonomie) depends essentially on the classification of frail old people in 6 degrees of dependency (GIR1 to GIR6). The aim of the present study was to test the reliability of this grid to evaluate the degree of dependency in demented elderly people. Mild, moderate or severe demented patients were included in the study (n= 120). A factorial validation of the A GGIR grid was performed by principal components analysis (PCA). This analysis showed a 5-factor solution: factor 1 named the property factor (27 percent of the variance), factor 2 named the dynamic factor (21 percent),factor 3 named the cognitive factor (20 percent), factor 4 named the external mobility factor (11 percent) and factor 5 named the communication factor (11 percent). The result showed that the AGGIR grid takes physical dependency more into account than psychological and behavioral dependency. This result suggests a need for readjustment of the AGGIR grid for demented patients by adding new variables taking into account psychosocial and behavioral disorders. PMID- 15269674 TI - [Guillain-Barre syndrome in HIV-infected patients at Bobo-Dioulasso Hospital (Burkina Faso)]. AB - Neurological manifestations of human immunodeficiency virus (HIV) infection are frequent and several associated peripheral neuropathies have been recognized. Among them, Guillain-Barre syndrome (GBS) may occur either early or during the course of the illness. We present a prospective study of 32 consecutive cases of GBS managed over a 5-Year period at Bobo-Dioulasso Hospital where HIV prevalence reaches 20.1p.cent. Male gender predominated (24/32). GBS occurred during the dry season for 65.7p.cent of the patients. Prior infections were found in 84.4p.cent. The motor deficit was paraplegia or tetraplegia. Clinically, paraplegia was associated with transient urinary sphincteric involvement in 24 HIV-infected patients and 3 HIV negative patients. Facial nerve paralysis was found in 3 patients. Among the 32 patients with GBS, 27 were tested positive for HIV. Two patients were infected by HIV1 and HIV2. Cerebrospinal fluid examination showed albumin-cell dissociation and elevated albumin level in 75p.cent of the samples. Autonomic neuropathies were seen in 9 HIV-infected patients. The CD4 counts were above 200/mm3 in 10 among 18 HIV-infected patients. The clinical presentations were more severe in HIV-positive patients with a longer duration of symptoms. HIV infected patients walked unaided within 51.1 days of peak paralysis. No fatal event occurred. This study indicates clearly that GBS in young adults is strongly associated with HIV infection and should be considered as an indicator of HIV infection in Black Africans. In the tropical context GBS should lead to HIV screening. PMID- 15269675 TI - [Alien hand syndrome due to a corpus callosum infarct: a case report]. AB - A 64-year-old man with alien hand syndrome presented with abnormal feelings in the left upper limb, associated with an intermanual conflict. There were no clinical signs of frontal or parietal apraxia. This syndrome was induced by an infarct localized in the right posterior area of the splenium, subsequent to a cardiogenic embolus. This observation enlightens the rising syndrome of callosal type alien syndrome due to a posterior callosal infarct. PMID- 15269676 TI - [The presentation of central pontine myelinolysis with a left upper monoplegia unassociated with hyponatremia or malnutrition]. AB - Arising via an unidentified pathogenic mechanism, central pontine myelinolysis affects the central portion of the base of the pons. Rapid correction of hyponatremia is a frequent cause. The main symptoms are spastic tetraparesis, pseudobulbar paralysis and locked-in syndrome. We report a case of central pontine myelinolysis without hyponatremia, revealed by left upper monoplegia in a 40-Year-old female alcoholic patient without malnutrition symptom. Central pontine myelinolysis occurred consecutive to a suicide attempt with multiple drugs and alcohol. The diagnosis was confirmed by brain magnetic resonance imaging. This case illustrates the paucisymptomatic clinical characteristics of central ponine myelinolysis in patients without electrolyte and nutritional disorders. PMID- 15269677 TI - [Idiopathic medullary decompression sickness: myth or reality?]. AB - Severe decompression sickness occurs unfrequently, with, generally an identifying cause (error in decompression protocols, promoting factors.). We report a case of severe spinal cord damage; onset after a common dive, neither deep nor long, without any promoting factor, absence of responsiveness to recompression, three hours post-dive, importance of MRI signal abnormalities, make us to point out the confounding variability of onset and evolution of such illness. PMID- 15269678 TI - [Diffusion-weighted magnetic resonance imaging in hypoglycemic coma]. AB - Hypoglycemia is a classic cause of coma and can result in irreversible neuronal loss. Until now, the main prognostic factors were depth of hypoglycemia and duration of coma. We report the case of a 55-Year-old woman who suffered severe hypoglycemic coma with abnormal cortico- subcortical diffusion weighted MR images. These MRI abnormalities preceded severe atrophy of these cerebral areas. This findings suggests that diffusion abnormalities in hypoglycemic coma may be related to neuronal loss and may thus have prognostic value. PMID- 15269679 TI - [Ischemic stroke and herpes simplex virus type-1 associated meningoencephalitis]. AB - The etiology of stroke in young patients is often unknown. Although systemic infections as well as specific infection agents, like herpes zoster virus or cysticercus, are often considered as risk factors, there are no indications that herpes simplex type 1 plays a role in the pathogenesis of stroke. We present the case of a young patient who suffered a stroke during a meningoencephalitis due to herpes simplex 1 and we review the relevant literature for a possible relation between the two entities. PMID- 15269680 TI - [Cerebral infraction by calcified embolism: a spontaneous complication of calcified aortic stenosis]. AB - Calcified aortic stenosis (CAS) is an unusual cause of cerebral infarct. The presence of cerebral intra-vascular or intra-parenchymatous calcifications, symptomatic or not, is suggestive of the diagnosis of CAS. We report two patients who experienced stroke induced by spontaneous calcic emboli from a calcified aortic valve and underline the importance of brain CT scan. PMID- 15269681 TI - [Cerebral Erdheim-Chester disease]. AB - We report the case of a 26-old-year man hospitalized for first partial complex epileptic seizure. Brain MRI showed an asymptomatic pseudo-tumor lesion in the brainstem. Diabetes insipidus, hypophyseal gonadotropic deficiency and osteosclerosis of long bones strongly suggested Erdheim-Chester disease, a rare histiocytosis, confirmed after tibial biopsy. Six months later, the patient remained stable. A persistent, and even increased, enhancement with Gd-DTPA on brain MR images was noted as previously described. The review of the literature collected 64 cases, and only 7 cases of cerebral "tumor". PMID- 15269682 TI - [A case of spongiform encephalopathy mimicking cortico-basal degeneration]. AB - INTRODUCTION: The presentation of subacute spongiform encephalopathies is varied and includes various movement disorders such as parkinsonism, myoclonus, or dystonia. These signs, especially when asymmetrical, can lead to the diagnostic of corticobasal degeneration. CASE REPORT: We report the case of a 75-Year-old woman who developed clinical signs suggestive of corticobasal degeneration: asymmetric rigidity and apraxia, limb dystonia, and postural instability. The final diagnosis of spongiform encephalopathy was suspected because of rapid decline and confirmed by post-mortem examination. CONCLUSION: This case highlights the importance of considering subacute spongiform encephalopathy in patients with a clinical presentation compatible with corticobasal degeneration. PMID- 15269683 TI - [The diagnosis of orthostatic tremor]. PMID- 15269684 TI - [Cerebral aspergillosis]. AB - INTRODUCTION: The brain is almost always a localization of invasive aspergillosis, after hematogenous spread from pulmonary aspergillosis. Brain aspergilosis is not rare and is one of the worst prognosis factors of invasive aspergillosis. STATE OF ART: The incidence of this severe mycosis is currently on the rise due to the development of major immunosuppressive treatments. Brain aspergillosis is noteworthy for its vascular tropism, leading to infectious cerebral vasculitis, mainly involving thalamoperforating and lenticulostriate arteries, with a high frequency of thalamic or basal nuclei lesions. Extra neurologic features that suggest this diagnosis are: i) risk factors for invasive aspergillosis (major or prolonged neutropenia, hematologic malignancies, prolonged corticosteroid treatment, bone marrow or solid organ transplant, AIDS); ii) persistent fever not responding to presumptive antibacterial treatment; iii) respiratory signs (brain aspergillosis is associated with pulmonary aspergillosis in 80 to 95 p. 100 of cases). Perspectives. Two recent major improvements in brain aspergillosis management must be outlined: i) for diagnostic purposes, the development of testing for Aspergillus antigenemia (a non-invasive procedure with good diagnostic value for invasive aspergillosis); ii) for therapeutic purposes, the demonstration that voriconazole is better than amphotericin B in terms of clinical response, tolerance and survival, for all types of invasive aspergillosis, the benefit being probably even greater in case of brain aspergillosis because of the good diffusion of voriconazole into the central nervous system. CONCLUSIONS: Brain aspergillosis is a severe emerging opportunistic infection for which diagnostic and therapeutic tools have recently improved. Thus, this diagnostic must be suspected early, especially in the immunocompromised patient, in the event of respiratory symptoms and when the brain lesions are localized in the central nuclei and the thalamus. PMID- 15269685 TI - [Report from the 56th annual meeting of the American Academy of Neurology]. PMID- 15269686 TI - Thorax: new millennium vascular imaging. AB - In recent years the technological development of computed tomography (CT) and magnetic resonance (MR) has promoted new improvements in diagnosis by means of imaging. In particular the introduction of multislice CT and MR angiography (MRA) has broadened the non-invasive diagnostic possibilities in the vascular study of the thorax. The new technological developments of CT and MR enable functional studies as well. Recent studies have demonstrated that CT and MR are as accurate in finding the vascular anomalies as digital subtraction angiography, while they are more precise in recognising possible associated pathologies which modify therapeutic treatment (for example of the trachea, bronchi, oesophagus etc.). There are many vascular structures in the thorax which need to be considered (aorta, pulmonary and coronary arteries, pulmonary veins, vena cava). The field of associated pathologies is also broad, and includes congenital vascular anomalies, vascular malformations, aorta dissection, vascular compression syndromes, atherosclerotic stenosis or occlusions, and pulmonary embolism. In pulmonary embolism some authors have demonstrated the utility of CT, in showing pulmonary segmental perfusion defects, and MRA, in identifying sub-segment pulmonary embolism. In this paper we analyse the most important CT and MR applications for the study of vascular thoracic diseases and compare them with other diagnostic techniques. We also evaluate the morpho-functional capabilities of CT and MR in this field. PMID- 15269687 TI - CT and functional respiratory tests. Evaluation of efficacy of bronchodilator therapy in chronic obstructive pulmonary disease (COPD). AB - PURPOSE: To evaluate changes in qualitative and quantitative parameters at high resolution CT (HRCT) and in respiratory function indexes (RFI) after bronchodilator administration in COPD patients. MATERIALS AND METHODS: Fifteen former smokers (9 male and 6 female, mean age 65 years), with COPD were studied. Patients with asthmatic or pulmonary diseases were excluded. After informed consent, the patients underwent blood gas analysis, baseline RFI (respiratory function index), inspiratory/expiratory HRCT, administration of salbutamol and bromide (by metered-dose inhaler), RFI after 45 minutes, and inspiratory/expiratory HRCT. First, we performed a visual qualitative analysis (by means of a preformed questionnaire), and then a quantitative analysis by "density mask" programme at three levels: aortic arch, carina, and supradiaphragmatic level. By using this diagnostic approach we were able to determined the global area of each lung, dependent and independent of density, bronchus-artery ratio, emphysema extension, air trapping, morphologic distortion of the airways. RESULTS: Data correlation analysed through linear regression statistical test. Outstanding correlations were found between the RFI (Raw, SVC, IC, RV, FEV1, VP70) and, respectively, density, global lung areas, and bronchus artery ratio changes before and after bronchodilators administration. CONCLUSIONS: Our study confirmed the usefulness of bronchodilators and the need for a multiparametric and comparative (radiological and functional) approach to COPD, as well as the usefulness of HRCT in evaluating the response to visible airway physiologic stimulus and extension, and reversibility of air trapping. PMID- 15269688 TI - Virtual bronchoscopy in patients with central endobronchial stenosing lesions. Technique optimisation with single slice spiral CT. AB - PURPOSE: To describe an original protocol for single slice spiral Computed Tomography (CT) virtual bronchoscopy in the evaluation of patients with central airway stenoses and compare the results with fibreoptic bronchoscopy. MATERIALS AND METHODS: Ten patients (4 female and 6 male; age range 22-60 years; mean age 44 years) with endobronchial disease diagnosed by fibreoptic bronchoscopy (8 malignant tumours, 1 benign tumour and 1 fibroid stenosis) underwent virtual bronchoscopy with single slice spiral CT. A panoramic spiral CT scan of the whole chest was first obtained. Once the area of interest had been identified, a new contrast enhanced scan was performed, from bottom to top, with the following parameters: 2 mm slice thickness, 1 mm reconstruction index, 1.3 pitch, 120 Kvp, 80 mAs. Virtual bronchoscopy was generated with an upper threshold of -500 HU from the cross-sectional images of the second scan on a dedicated workstation. Axial, multiplanar reformations (MPR), and virtual endoscopy simulation were simultaneously visualised. Virtual CT bronchoscopy findings were compared with those of fibreoptic bronchoscopy. RESULTS: The protocol we used to perform single slice spiral CT virtual bronchoscopy enabled us to obtain virtual bronchoscopy images that correlated well with fibreoptic bronchoscopy findings in all cases, as well as allowing the visualization of the airways beyond the stenoses. Information about tissues surrounding the tracheobronchial tree was also available from axial and MPR images. Only in 1 case were motion artefacts observed. CONCLUSIONS: The set of the most appropriate parameters for performing virtual bronchoscopy by single slice spiral CT has not yet been standardized. In our opinion the appropriate selection of the protocol to adequately realize virtual bronchoscopic images is crucial when using CT devices such as the above, so as to achieve the correct balance between the quality of image definition and exposure dose. PMID- 15269689 TI - Diagnostic work up of arrhythmogenic right ventricular cardiomyopathy by cardiovascular magnetic resonance (CMR). Consensus statement. AB - Cardiovascular Magnetic Resonance (CMR) has become a widespread diagnostic tool. Since its introduction CMR has been used to image patients with known or suspected arrhythmogenic right ventricular cardiomyopathy (ARVC). Several abnormalities have been found and described by CMR and at present this diagnostic tool is considered very important for the diagnosis. However, the diagnosis of ARVC relies upon the fulfillment of both clinical and functional criteria and CMR can provide several but not all the information useful for the diagnosis. Furthermore, some findings such as evidence of right ventricular epicardial fat once considered a peculiar marker of ARVC, have been shown to possess a low specificity. This document was prepared by representatives of the three Italian official Organizations involved in CMR. Its main scope is to highlight the problems encountered when studying patients with suspected ARVC by CMR, to indicate the basic technical equipment needed, to recommend a proper imaging protocol and to offer a consensus on the main diagnostic features relevant for the diagnosis. PMID- 15269690 TI - Serial plain abdominal film findings in the assessment of acute abdomen: spastic ileus, hypotonic ileus, mechanical ileus and paralytic ileus. AB - In recent years the increasing use of ultrasonography and computed tomography in the assessment of diseases causing acute abdomen and the diagnostic possibilities of magnetic resonance have decreased the role of conventional radiology techniques, especially of plain abdominal film in the diagnosis of acute abdomen. However, serial plain abdominal film is still the first diagnostic procedure used in the assessment of patients with acute abdominal pain, providing important diagnostic information if correctly performed and carefully observed. In this paper serial plain abdominal film findings related to the different types of ileus (spastic ileus, hypotonic ileus, mechanical ileus and paralytic ileus) are presented. PMID- 15269691 TI - Comparison of diagnostic performance of unenhanced vs SonoVue - enhanced ultrasonography in focal liver lesions characterization. The experience of three Italian centers. AB - PURPOSE: To evaluate the diagnostic performance of contrast-enhanced US with SonoVue (Bracco, Milan, Italy) compared to baseline US in focal hepatic lesions characterization. MATERIALS AND METHODS: A comprehensive number of four operators from 3 hospitals evaluated 57 consecutive patients with 60 focal hepatic lesions (28 hepatocellular carcinomas, 11 metastases, 13 hemangiomas, 1 hepatocellular adenoma and 7 focal nodular hyperplasias) by baseline gray-scale ultrasound (US) and color Doppler US. The same lesions were subsequently scanned by contrast enhanced US after intravenous bolus administration of 2,4-4,8 ml of SonoVue by employing intermittent high or continuous low transmit power imaging. The diagnosis of lesions nature (benign or malignant) and histotype proposed by the on-site operator was finally compared to the definite diagnosis reached by reference procedures (multiphasic contrast-material enhanced helical-computed tomography or magnetic resonance in 24 lesions and fine needle US guided biopsy in 36 lesions). Diagnostic performance (sensitivity, specificity and overall accuracy expressed by the agreement with the reference procedures) of baseline and contrast enhanced US were compared. RESULTS: Differences in sensitivity (baseline vs contrast-enhanced US: 13/39 [33%] vs 32/39 [82%]), specificity (baseline vs contrast-enhanced US: 12/21 [57%] vs 16/21 [76%]) and overall accuracy (baseline vs contrast-enhanced US: 25/60 [41%] vs 47/60 [78%]) were significant (p<0.05; McNemar test). CONCLUSIONS: SonoVue-enhanced US determined a significant improvement in diagnostic performance in the characterization of focal liver lesions if compared to baseline US. PMID- 15269692 TI - Contrast enhanced ultrasound with second generation contrast agent in traumatic liver lesions. AB - PURPOSE: To evaluate the accuracy of second generation contrast enhanced US (Sonovue) in the diagnosis and staging of traumatic hepatic lesions, compared with conventional US and spiral CT. MATERIAL AND METHODS: A total of 203 patients (127 males, 76 females, mean age 36 years) with isolated abdominal trauma were examined with conventional and contrast enhanced US (Sonovue, Bracco, Italy) between March 2002 and February 2003. The sonographic contrast agent was administered at a dose of 2 ml/10-15", repeated twice. CT examinations were performed with single- (Rhota, Esaote Biomedica, Italy) or multislice spiral CT with administration of contrast agent. The presence and number of lesions, hepatic capsular involvement, size and sonographic pattern were evaluated. RESULTS: Conventional US demonstrated hepatic lesions in 27 patients, in 3 cases it identified 2 foci (30 lesions, size 2-8 cm). Contrast enhanced US (CEUS) revealed another 2 lesions and in 4 patients it identified lesions not shown at conventional US (size 2-5 cm). Capsular involvement was detected in 14 cases (11 with conventional US). The sonographic pattern of the lesions at conventional US was hypo-anechoic in 19 cases, and hyperechoic in 11. In CEUS all the lesions appeared strongly hypoechoic against a strongly hyperechoic parenchyma, with clear borders and larger size as compared with conventional US. CONCLUSIONS: In isolated blunt abdominal trauma CEUS is more accurate than conventional US in determining the number and size of lesions and detecting capsular involvement. This has a strong impact on diagnosis as the number of false negatives is reduced and on prognosis as the lesions are more accurately graded, and there is close correlation with spiral CT. CEUS can be used as a first approach in mild isolated abdominal trauma, in paediatric patients and in the follow-up, whereas CT is the method of choice in severe trauma and in multiple traumas. PMID- 15269693 TI - Alterations in hepatic uptake and distribution of organ-specific superparamagnetic MRI contrast media: clinical findings and classification according to pathogenesis. AB - PURPOSE: The aim of this study was to classify the alterations in the liver uptake and distribution of superparamagnetic contrast media that could potentially lead to diagnostic errors. These alterations, referred to as SPIO LUDA for convenience, may be caused by a variety of disorders, such as cirrhosis, vascular thrombosis, hepatitis and liver steatosis, that interfere with the normal uptake of the contrast material. These conditions can give rise to focal or diffuse areas of hyperintensity or hypointensity which may mimic the presence of neoplastic lesions or hinder the detection or characterisation of neoplasms. MATERIALS AND METHODS: We retrospectively reviewed the hepatic MR examinations of 412 patients performed to detect hepatocellular carcinoma in 349 cases and metastases in 63 cases, with the aim of identifying conditions corresponding to the definition of SPIO-LUDA. All the examinations were performed using a 1.5 Tesla MR unit with SE and GE sequences, T1 and T2 weighted images, and SPIO as a contrast agent, in 402 cases ferumoxide (Endorem, Guerbet) and in 10 cases SHU 555-A (Resovist, Schering). The SPIO-LUDA were classed into two groups: due to reduced uptake (cell replacement, reduced vascular flow and cellular inhibition) and due to increased uptake (Kupffer cell hyperactivity or increased vascularity). From a quantitative point of view we evaluated the percentage of signal loss (PSIL) of the SPIO-LUDA relative to the surrounding healthy parenchyma, as an expression of increase or reduced uptake of contrast material. RESULTS: In 54 patients we identified potentially misleading SPIO-LUDA: 41 were due to reduced uptake and 13 to increased uptake. In 16 cases, all of which cases of reduced uptake, the alteration significantly limited the diagnostic effectiveness of the MR examination. The reduced-uptake SPIO-LUDA were caused by fibrosis in 31 cases, by portal vein thrombosis in 3, by suprahepatic vein thrombosis in 2, by peritumoural vascular shunts in 3 and by hepatitis in 2. The increased-uptake SPIO-LUDA were caused by focal steatosis in 2 cases and by dysplastic nodules in cirrhosis in 11. The reduced-uptake SPIO-LUDA exhibited 30% lower PSIL values than the normal liver (range 15-45%); the increased-uptake SPIO LUDA displayed 19% higher PSIL values than the surrounding liver (range 15-23%). Out of a total of 412 patients, the alteration of SPIO uptake was so severe as to prevent detection or exclusion of focal lesions in 4% of cases. CONCLUSIONS: SPIO LUDA constitute a diagnostic challenge. The recognition and correct interpretation of these alterations are fundamental for avoiding confusion with other diseases and to obtain further clues for the interpretation of abnormal patterns detected at MRI or other imaging modalities. PMID- 15269694 TI - Imaging of early postoperative complications after polypropylene mesh repair of inguinal hernia. AB - PURPOSE: We report our experience with the use of US and CT in postoperative complications of inguinal hernioplasty using a prosthetic polypropylene mesh. MATERIALS AND METHODS: This study was divided into two parts. In the first we evaluated the in-vitro sonographic and CT appearance of a fragment of prosthetic mesh. In the second, we retrospectively reviewed the imaging findings in 31 patients (aged 42 to 75 years) examined after inguinal hernia repair between December 2000 and December 2002. Seventeen hernias had been repaired with a laparoscopic approach, and the others with the anterior tension-free technique proposed by Lichtenstein (12 cases) and Trabucco (2 cases). Sonography was performed to assess suspected complications between the second and the fourth postoperative day. Both high-resolution 7.5-10 MHz linear transducers and a 3.5 MHz convex probe were employed to ensure complete evaluation of superficial and deep structures. Eight obese patients also underwent CT for confirmation of the US results. RESULTS: At sonography the prosthetic mesh appeared as a linear hyperechoic image measuring about 2 mm in thickness, with posterior acoustic shadow and a finely irregular surface. Only one of the 17 patients examined after laparoscopic inguinal hernioplasty had a seroma; in the other 14 repaired with the anterior tension-free technique we identified 2 abscesses, 3 seromas, 2 "foldings" of the prosthetic mesh, and 2 mesh displacements with associated recurrence of hernia. CT confirmed the US results as to the presence of fluid collections, and visualised the prosthetic mesh in only 2/8 cases. CONCLUSIONS: Sonography is a useful means of assessing postoperative changes in laparoscopic and in anterior tension-free hernia repair. It can differentiate these complications from recurrences of hernia. Colour-Doppler US can also correctly detect normal blood flow of the testes. Sonography is the only technique that can easily demonstrate the prosthetic mesh in the abdominal wall. PMID- 15269695 TI - CT angiography in the assessment of carotid atherosclerotic disease: results of more than two years' experience. AB - PURPOSE: To assess the role of CT Angiography (CTA) in patients with carotid atherosclerotic disease as compared to echo-colour Doppler (CDUS) ultrasound of the supra-aortic trunks (SAT) and surgery. MATERIALS AND METHODS: Eighty-two patients with suspected carotid atherosclerotic disease were submitted to CDUS and CTA of the supra-aortic trunks. Agreement between CDUS and CTA was first evaluated with regard to the following parameters: degree of stenosis according to NASCET criteria, plaque morphology, presence of ulcerations, tandem lesions and vessel abnormalities. Secondly, data provided by the two methodologies were compared with the surgical specimens (35 patients); in 12 cases, the stenosis was measured on the cast of the carotid plaque made of for biologic use silicone. RESULTS: The correlation between CDUS and CTA in evaluating the degree of stenosis was 75.6%; poor agreement was found for mild (61.1%) and severe (69.1%) stenoses; agreement in the evaluation of vessel abnormalities, plaque morphology and ulcerations was 81.7%, 89.0% and 96.3%, respectively. CTA demonstrated 11 tandem lesions not detected CDUS. Compared to surgery, CTA correctly classified the degree of stenosis according to NASCET criteria in 31/35 cases (88.6%) - as opposed to 29/35 by CDUS (82.9%) - and never overestimated the stenosis. CTA proved superior to CDUS in detecting plaque ulcerations (75% vs 25% sensitivity) and vessel abnormalities (100% vs 44.4% sensitivity). CONCLUSIONS: CTA is recommended as a second-level examination in patients with carotid atherosclerotic steno-obstructive disease who are surgical candidates. PMID- 15269696 TI - Noninvasive evaluation of coronary artery stents patency after PTCA: role of Multislice Computed Tomography. AB - PURPOSE: Restenosis of a coronary artery treated with stent implantation is a well-known process that can compromise over time the success of a coronary angioplasty and, accordingly, treated patients must undergo periodic controls. We have recently witnessed a shift towards a greater use of Multi-slice CT (msCT) in the study of coronary disease without its precise indications and limits having yet been underlined. The purpose of our study is to assess the role of msCT in the follow-up of patients treated with coronary angioplasty. MATERIALS AND METHODS: Forty-eight patients, for a total of 72 lesions, who underwent treatment with a slotted tube stent implant, had an msCT examination 1 week before scheduled coronary angiography, and the results were compared. 34 stents/72 (47.2%) were inserted on the left anterior descending; 21/72 (29.2%) on the right coronary; 17/72 (23.6%) on the circumflex artery or obtuse marginal branches. RESULTS: The observation of the opacification of the vessel located distally to treated segments allowed us to assess the patency of all stents. Coronary angiography identified a significant intrastent restenosis or a stent occlusion in 12 of the 72 stents analysed (16.7%). msCT enabled easier visualization of the lumen of the treated artery and its differentiation from the stent struts in the ones located on the left anterior descending artery than those on the circumflex (28 stents out of 34 [82.4%] vs 13/17 [76.5%]; p<0.05), and on the right coronary artery, which were difficult to evaluate (11/21 [52.4%]). We were also able to visualize the lumen of 14/15 stents with a calibre over 3.5 mm [93.3%] vs 35/45 stent with dimensions between 3.1 e 3.4 mm [77.8%], and only 4 stents <3 mm/12 [33.3%]. On multivariate analysis, the characteristics that were significantly and independently associated with accurate visualization of the lumen of a stented vessel were location on the proximal anterior descending artery (OR 4.03 [IC 95%: from 2.34 to 8.05]; p<0.0001) and stent size of >3.5 mm (OR 2.97 [IC 95%: from 1.67 to 4.86]; p<0.01). CONCLUSIONS: The msCT technology available at present makes the study of smaller stents and those positioned on the right coronary artery and circumflex rather complex; on the other hand msCT appears a promising study method for stents greater then 3.5 mm and for those positioned on the proximal segment of the left anterior descending artery. PMID- 15269697 TI - Surveillance for certain health behaviors among selected local areas--United States, Behavioral Risk Factor Surveillance System, 2002. AB - PROBLEM: Monitoring risk behaviors for chronic diseases and participation in preventive practices are important for developing effective health education and intervention programs to prevent morbidity and mortality. Therefore, continual monitoring of these behaviors and practices at the state, city, and county levels can assist public health programs in evaluating and monitoring progress toward improving their community's health. REPORTING PERIOD COVERED: Data collected in 2002 are presented for states, selected metropolitan, and micropolitan statistical areas (MMSA), and their counties. DESCRIPTION OF THE SYSTEM: The Behavioral Risk Factor Surveillance System (BRFSS) is an on-going, state-based, telephone survey of the civilian, noninstitutionalized population aged >18 years. All 50 states, the District of Columbia (DC), Guam, the Virgin Islands, and the Commonwealth of Puerto Rico participated in BRFSS during 2002. Metropolitan and MMSA and their counties with >500 respondents or a minimum sample size of 19 per weighting class were included in the analyses for a total of 98 MMSA and 146 counties. RESULTS: Prevalence of high-risk behaviors for chronic diseases, awareness of certain medical conditions, and use of preventive health-care services varied substantially by state, county, and MMSA. Obesity ranged from 27.6% in West Virginia, 29.4% in Charleston, West Virginia, and 32.0% in Florence County, South Carolina, to 16.5% in Colorado, 12.8% in Bethesda-Frederick Gaithersburg, Maryland, and 11.8% in Washington County, Rhode Island. No leisuretime physical activity ranged from 33.6% in Tennessee, 36.8% in Miami Miami Beach-Kendall, Florida, and 36.8% in Miami-Dade County, Florida to 15.0% in Washington, 13.8% in Seattle-Bellevue-Everett Washington, and 11.4% in King County, Washington. Cigarette smoking ranged from 32.6% in Kentucky, 32.8% in Youngstown-Warren- Boardman, Ohio-Pennsylvania, and 31.1% in Jefferson County, Kentucky to 16.4% in California, 13.8% in Ogden- Clearfield, Utah, and 10.9% in Davis County, Utah. Binge drinking ranged from 24.9% in Wisconsin, 26.1% in Fargo, North Dakota-Minnesota, and 25.1% Cass County, North Dakota, to 7.9% in Kentucky, 8.2% in Greensboro- High Point, North Carolina, and 6.6% in Henderson County, North Carolina. At risk for heavy drinking ranged from 8.7% in Arizona, 9.5% in Lebanon, New Hampshire-Vermont, and 11.3% in Richland County, South Carolina, to 2.8% in Utah, 1.9% in Ogden-Clearfield, Utah, and 1.7% in King County, New York. Adults who were told they had diabetes ranged from 10.2% in West Virginia, 11.1% in Charleston, West Virginia, and 11.1% in Richland, South Carolina, to 3.5% in Alaska, 2.7% in Anchorage, Alaska, and 2.4% in Weber County, Utah. Percentage of adults aged>50 years who were ever screened for colorectal cancer ranged from 64.8% in Minnesota, 67.9% in Minneapolis-St. Paul-Bloomington Minnesota-Wisconsin, and 73.6% in Ramsey County, Minnesota, to 39.2% in Hawaii, 30.7% in Kahului-Wailuku, Hawaii, and 30.7% in Maui County, Hawaii. Persons aged >65 years who had received pneumococcal vaccine ranged from 72.5% in North Dakota, 74.8% in Minneapolis-St. Paul-Bloomington, Minnesota-Wisconsin, and 73.1% in Milwaukee County, Wisconsin, to 47.9% in DC, 47.5% in New York-Wayne-White Plains, New York, New Jersey, and 47.9% in DC County, DC. Older adults who had received influenza vaccine ranged from 76.6% in Minnesota, 80.0% in Minneapolis St. Paul-Bloomington, Minnesota-Wisconsin, and 76.3% in Middlesex County, Massachusetts, to 57.0% in Florida, 55.8% in Houston-Baytown-Sugar Land, Texas, and 56.2% in Cook County, Illinois. INTERPRETATION: BRFSS data indicate substantial variation in high-risk behaviors, participation in preventive healthcare services, and screening among U.S. adults at states and selected local areas, indicating a need for continued efforts to evaluate public health programs or policies designed to reduce morbidity and mortality. PUBLIC HEALTH ACTIONS: Data from BRFSS are useful in developing and guiding public health programs and policies. Therefore, states, selected MMSA, and their counties can use BRFSS data as a tool to prevent premature morbidity and mortality among adult population and to assess progress toward national health objectives. The data indicate a continued need to develop and implement health promotion programs for targeting specific behaviors and practices and serve as a baseline for future surveillance at the local level in the United States. PMID- 15269698 TI - Changing patterns of pneumoconiosis mortality--United States, 1968-2000. AB - Pneumoconioses are caused by the inhalation and deposition of mineral dusts in the lungs, resulting in pulmonary fibrosis and other parenchymal changes. Many persons with early pneumoconiosis are asymptomatic, but advanced disease often is accompanied by disability and premature death. Known pneumoconioses include coal workers' pneumoconiosis (CWP), silicosis, asbestosis, mixed dust pneumoconiosis, graphitosis, and talcosis. No effective treatment for these diseases is available. This report describes the temporal patterns of pneumoconiosis mortality during 1968-2000, which indicates an overall decrease in pneumoconiosis mortality. However, asbestosis increased steadily and is now the most frequently recorded pneumoconiosis on death certificates. Increased awareness of this trend is needed among health-care providers, employers, workers, and public health agencies. PMID- 15269699 TI - Acute hemorrhagic conjunctivitis outbreak caused by Coxsackievirus A24--Puerto Rico, 2003. AB - Acute hemorrhagic conjunctivitis (AHC) is an epidemic form of highly contagious conjunctivitis and is characterized by sudden onset of painful, swollen, red eyes, with conjunctival hemorrhaging and excessive tearing. Since 1981, when AHC was first detected in the Western Hemisphere, three major epidemics had occurred until 2003, all affecting the Caribbean. During August-October 2003, a fourth epidemic occurred in Puerto Rico (2000 population: 3.8 million). This report summarizes the outbreak investigation conducted by the Puerto Rico Department of Health (PRDOH), which documented an estimated 490,000 persons with illness, including >51,000 cases reported by physicians; demonstrated laboratory evidence of Coxsackievirus A24 (CA24); and determined that school-aged children (i.e., aged 5-18 years) and those living in crowded urban areas were at highest risk. To control outbreaks of AHC, prevention methods (e.g., frequent hand washing and avoidance of sharing towels and bedding) should be targeted to groups at highest risk, and information should be disseminated after the first report of AHC in the area. PMID- 15269700 TI - Progress toward poliomyelitis eradication--Afghanistan and Pakistan, January 2003 May 2004. AB - Since the 1988 World Health Assembly resolution to eradicate poliomyelitis, the number of countries where polio is endemic decreased from approximately 125 to six by the end of 2003. In 2003, poliovirus importations were reported in 10 countries, including eight in West and Central Africa, one in Southern Africa (Botswana), and one in the Middle East (Lebanon). Two countries where poliovirus remains endemic are Afghanistan and Pakistan, which are analyzed together because of their geographic proximity, frequent cross-border population movements, and genetically similar wild poliovirus (WPV) lineages. This report describes intensified polio eradication activities in Afghanistan and Pakistan during January 2003-May 2004, summarizes progress made toward eradication, and highlights the remaining challenges to interrupting poliovirus transmission. PMID- 15269701 TI - West Nile virus activity--United States, July 14-20, 2004. AB - During the week of July 14-20, a total of 74 cases of human West Nile virus (WNV) illness were reported from seven states (Arizona, California, Florida, New Mexico, New York, South Dakota, and Texas). PMID- 15269702 TI - How bereaved multiple-birth parents cope with hospitalization, homecoming, disposition for deceased, and attachment to survivors. AB - OBJECTIVE: To elicit bereaved multiple-birth parents' perceptions regarding support, disposition decisions, attachment to surviving multiples, discharge, and later coping. STUDY DESIGN: Narrative email survey of 70 bereaved parents with quantitative and qualitative analysis. RESULTS: Bereaved parents of multiples find neonatal hospitalization stressful. Not all caregivers acknowledged loss, although most parents would welcome brief loss discussions during survivors' hospitalization. Half of respondents felt social workers could help coordinate support or mental health care. Most participants recalled hospital support for loss, but only 43% recalled support for neonatal hospitalization. Respondents praised peer support and written materials. Parents felt ambivalent about disposition for deceased babies while comultiples were ill; most made prompt arrangements. Attachment to survivors was difficult for half. Breastfeeding and discharge planning were important, and 31% had difficulty coping at home. CONCLUSIONS: Caregivers must communicate better with bereaved multiple-birth parents, whose desires for support, loss discussions, disposition, and discharge vary considerably. PMID- 15269703 TI - Epinephrine delivery during neonatal resuscitation: comparison of direct endotracheal tube vs catheter inserted into endotracheal tube administration. AB - OBJECTIVES: The optimal method for epinephrine administration during neonatal resuscitation is not known. We hypothesized that epinephrine will be delivered more efficiently when administered via a feeding catheter inserted into the endotracheal tube (C-ETT) vs when administered directly into the ETT (D-ETT). Our objectives were to (1) compare the delivery of epinephrine to the distal end of the ETT when administered via D-ETT vs C-ETT; (2) measure the retention of epinephrine within the ETT vs the feeding catheter used for the drug delivery; and (3) compare the delivery of the drug with and without an air flush after administration via C-ETT. METHODS: All experiments were performed in vitro, simulating epinephrine administration during neonatal resuscitation, according to the standard guidelines. Radiolabeled epinephrine, diluted to 1 microCi/ml, was used and experiments were repeated at least 4 times. Epinephrine administration via D-ETT was followed by one manual breath via a self-inflating bag attached to the ETT. Epinephrine delivery via C-ETT was followed by 1 ml saline flush, and in some experiments, this was also followed by a 1 cm(3) air flush. Epinephrine delivery and retention were assessed by measuring the radioactive content of the effluent fluid and that of the ETT or of the feeding catheter used for drug delivery. RESULTS: Significantly higher dosage of the drug was delivered when administered via D-ETT vs C-ETT, if air flush following C-ETT method was not used. However, with an air flush following the saline flush after the drug instillation, there was no difference in the amount of epinephrine delivered between the two methods. Retention in the ETT wall or the catheter was <7.5% of the administered dose with either method. CONCLUSIONS: Without an air flush following C-ETT method of epinephrine delivery, higher dosage of the drug is delivered via D-ETT vs C-ETT method. An air flush following the saline flush during C-ETT method improves drug delivery. Given that the C-ETT method is more cumbersome and time consuming, and does not improve drug delivery, D-ETT administration should be the method of choice for epinephrine delivery during neonatal resuscitation. PMID- 15269704 TI - Prevalence, awareness, treatment and control of hypertension in North America, North Africa and Asia. AB - Results from national surveys of prevalence, awareness, treatment and control provide the most meaningful basis for assessing the burden of hypertension in the community. National surveys conducted in a variety of countries in North America, Europe, Australia, Asia and Africa have identified a strikingly similar relationship between age and blood pressure (BP), with a progressive and steep increase in systolic BP throughout adult life and a less steep increase in diastolic BP from adolescence until the fifth or sixth decade. In most countries surveyed, there was a high prevalence of hypertension. Approximately, one quarter of all adults in the United States and Egypt had hypertension (systolic BP>/=140 mmHg or diastolic BP>/=90 mmHg or use of antihypertensive medication) in national surveys conducted in 1988-1991 and 1991-1993, respectively. The corresponding percentage was somewhat lower (14.4%) for adults surveyed in China during 1991, but temporal trends indicate that the prevalence of hypertension is increasing rapidly in that country. In the 1988-1991 national survey, more than 25% of US adults were unaware of their diagnosis, only 55% were being treated with antihypertensive medication and only 29% were on antihypertensive medication with a systolic/diastolic BP >140/90 mmHg. The situation was much worse in Egypt and China, with only 8% and <5% of adults with hypertension, respectively, being treated with antihypertensive medication and having a systolic/diastolic BP <140/90 mmHg. These survey results underscore the fact that hypertension is highly prevalent, poorly treated and controlled, and an escalating health challenge in economically developing countries. PMID- 15269705 TI - Moxonidine in the treatment of overweight and obese patients with the metabolic syndrome: a postmarketing surveillance study. AB - Moxonidine is a centrally active imidazoline receptor agonist that effectively lowers blood pressure and has been shown to have beneficial effects on lipid and carbohydrate metabolism. We assessed the efficacy of moxonidine in a postmarketing surveillance study (CAMUS) conducted in 772 practices in Germany, documenting 4005 patients with hypertension, who were overweight and/or suffered from metabolic syndrome. Patients were treated with moxonidine (Cynt) for the first time following the baseline visit for 8 weeks. Mean blood pressure decreased from 168/97 to 141/83 mmHg for all patients and from 168/96 to 141/83 mmHg for patients with metabolic syndrome. Blood pressure reduction was particularly pronounced in patients with severe hypertension at baseline. The response rate (DBP< or =90 mmHg or reduction > or =10 mmHg) of antihypertensive treatment with moxonidine was 94.0% for all patients and 93.8% for patients with metabolic syndrome. The recommended targets for antihypertensive treatment of the German Diabetes Society/German Hypertension Society were reached by 30.5% of nondiabetics (goal: <140/90 mmHg) and by 3.6% of diabetics (goal: <130/80 mmHg) observed. After 8 weeks of treatment, patients achieved a mean weight loss of 1.4 kg, which was particularly pronounced in obese patients. The rate of patients receiving antihypertensive combination therapy was 81.1% for those with metabolic syndrome, and 63.3% for all other patients. Patients with metabolic syndrome were preferentially treated with ACE inhibitors and diuretics. We conclude that moxonidine effectively reduces blood pressure in patients with metabolic syndrome while simultaneously reducing body weight in obese patients. PMID- 15269706 TI - Blood pressure responses to whole-body cold exposure: effect of metoprolol. PMID- 15269707 TI - Baseline Na-Li countertransport and risk of hypertension in children: a 10-year prospective study in Hanzhong children. AB - Sodium-lithium countertransport (Na-Li CT) is associated with blood pressure (BP) and in many cross-sectional investigations and some longitudinal studies, essential hypertension has been proposed as a biochemical marker or predictor of hypertension risk in adults. The present study investigated prospectively whether baseline Na-Li CT rate was an index of increased risk of future development of hypertension in children. At baseline visit in 1987 of the Hanzhong Children Hypertension Study comprising 4000 school children aged 5-6 years old, 310 samples were randomly selected for measurement of baseline Na-Li CT rate; we made a 10-year follow-up of them in the same season in 1997. This cohort of children is the sample for analysis in the present report. Baseline Na-Li CT rate was positively correlated to systolic BP (SBP) both in baseline and follow-up (baseline, gamma=0.21, P<0.05; follow-up, gamma=0.32, P<0.01), and positively correlated to diastolic BP (DBP) (gamma=0.20, P<0.05) and body mass index (gamma=0.18, P<0.05) in follow-up examination. Longitudinal analysis of 10-year BP evolution, children in higher baseline Na-Li CT (ie, >260 micromoll RBC/h) had greater BP change than children in lower baseline Na-Li CT (ie, 260 mumol/l RBC/h) were associated with approximately 1.5 times greater risk of high BP) in comparison to placement in lower Na-Li CT (2 years) when all control mice had died. In addition to retarding lesion progression, treatment caused lesion remodeling from a vulnerable-looking to a more stable-appearing phenotype. In conclusion, HD-Ad-mediated LDLR gene therapy is effective in conferring long-term protection against atherosclerosis in a mouse model of familial hypercholesterolemia. PMID- 15269712 TI - Improved chitosan-mediated gene delivery based on easily dissociated chitosan polyplexes of highly defined chitosan oligomers. AB - Nonviral gene delivery systems based on conventional high-molecular-weight chitosans are efficient after lung administration in vivo, but have poor physical properties such as aggregated shapes, low solubility at neutral pH, high viscosity at concentrations used for in vivo delivery and a slow dissociation and release of plasmid DNA, resulting in a slow onset of action. We therefore developed highly effective nonviral gene delivery systems with improved physical properties from a series of chitosan oligomers, ranging in molecular weight from 1.2 to 10 kDa. First, we established structure-property relationships with regard to polyplex formation and in vivo efficiency after lung administration to mice. In a second step, we isolated chitosan oligomers from a preferred oligomer fraction to obtain fractions, ranging from 10 to 50-mers, of more homogeneous size distributions with polydispersities ranging from 1.01 to 1.09. Polyplexes based on chitosan oligomers dissociated more easily than those of a high molecular-weight ultrapure chitosan (UPC, approximately a 1000-mer), and released pDNA in the presence of anionic heparin. The more easily dissociated polyplexes mediated a faster onset of action and gave a higher gene expression both in 293 cells in vitro and after lung administration in vivo as compared to the more stable UPC polyplexes. Already 24 h after intratracheal administration, a 120- to 260-fold higher luciferase gene expression was observed compared to UPC in the mouse lung in vivo. The gene expression in the lung was comparable to that of PEI (respective AUCs of 2756+/-710 and 3320+/-871 pg luciferase x days/mg of total lung protein). In conclusion, a major improvement of chitosan-mediated nonviral gene delivery to the lung was obtained by using polyplexes of well-defined chitosan oligomers. Polyplexes of oligomer fractions also had superior physicochemical properties to commonly used high-molecular-weight UPC. PMID- 15269713 TI - CTLA4Ig delivered by high-capacity adenoviral vector induces stable expression of dystrophin in mdx mouse muscle. AB - Adenoviral (Ad) vector-mediated gene delivery of normal, full-length dystrophin to skeletal muscle provides a promising strategy for the treatment of Duchenne muscular dystrophy (DMD), an X-linked recessive, dystrophin-deficient muscle disease. Studies in animal models suggest that successful DMD gene therapy by Ad vector-mediated gene transfer would be precluded by cellular and humoral immune responses induced by vector capsid and transgene proteins. To address the immunity induced by Ad vector-mediated dystrophin gene delivery to dystrophic muscle, we developed high-capacity adenoviral (HC-Ad) vectors expressing mouse dystrophin driven by the muscle creatine kinase promoter (AdmDys) and mCTLA4Ig (AdmCTLA4Ig) individually, or together from one vector (AdmCTLA4Ig/mDys). We found stable expression of dystrophin protein in the tibialis anterior muscles of mdx mice, coinjected with AdmCTLA4Ig and AdmDys, or injected alone with AdmCTLA4Ig/mDys, whereas the expression of dystrophin protein in the control group coinjected with AdmDys and an empty vector decreased by at least 50% between 2 and 8 weeks after administration. Additionally, we observed reductions in Ad vector-induced Th1 and Th2 cytokines, Ad vector-specific cytotoxic T lymphocyte activation and neutralizing anti-Ad antibodies in both experimental groups that received a mCTLA4Ig-expressing vector as compared to the control group. This study demonstrates that the coexpression of mCTLA4Ig and dystrophin in skeletal muscle provided by HC-Ad vector-mediated gene transfer can provide stable expression of dystrophin in immunocompetent, adult mdx mouse muscle and applies a potentially powerful strategy to overcome adaptive immunity induced by Ad vector-mediated dystrophin gene delivery toward the ultimate goal of treatment for DMD. PMID- 15269714 TI - Adenovirus hexon T-cell epitope is recognized by most adults and is restricted by HLA DP4, the most common class II allele. AB - The immunogenicity of adenovirus (Ad) vectors is enhanced by virus-specific memory immune responses present in most individuals as a result of past exposure to these ubiquitous pathogens. We previously identified the first human T-cell epitope from the major capsid protein hexon, H910-924, and found that it is highly conserved among different Ad serotypes. Memory/effector T-cell responses to H910-924 were detected in 14 of 18 (78%) healthy adults by an interferon-gamma ELISPOT assay. Hexon peptide-specific CD4 T-cell lines were generated from three HLA-typed donors and analyzed using a panel of HLA homozygous B-cell lines and monoclonal antibodies to HLA class II loci. These studies reveal that the hexon epitope is restricted by HLA DP4, a class II allele present in 75% of the population. Analysis of overlapping peptides and peptides with single residue mutations identified a HLA DP4-binding motif. Additionally, antibodies to the hexon peptide were detected in all donor sera by dot blot assay and ELISA. Therefore, most individuals exhibit both memory B- and T-cell responses to this highly conserved epitope on hexon, an obligate component of all Ad vectors, including 'gutted' vectors. These data suggest that current strategies for the use of Ad gene therapy vectors will not evade memory immune responses to Ad. PMID- 15269715 TI - Salivary glands and gene therapy: the mouth waters. PMID- 15269716 TI - Development of efficient plasmid DNA transfer into adult rat central nervous system using microbubble-enhanced ultrasound. AB - Although gene therapy might become a promising approach for central nervous system diseases, the safety issue is a serious consideration in human gene therapy. To overcome this problem, we developed an efficient gene transfer method into the adult rat brain based on plasmid DNA using a microbubble-enhanced ultrasound method, since microbubble-enhanced ultrasound has shown promise for transfecting genes into other tissues such as blood vessels. Using the microbubble-enhanced ultrasound method, luciferase expression was increased approximately 10-fold as compared to injection of naked plasmid DNA alone. Interestingly, the site of gene expression was limited to the site of insonation with intracisternal injection, in contrast to previous studies using viruses. Expression of the reporter gene, Venus, was readily detected in the central nervous system. The transfected cells were mainly detected in meningeal cells with intracisternal injection, and in glial cells with intrastriatal injection. There was no obvious evidence of tissue damage by microbubble-enhanced ultrasound. Overall, the present study demonstrated the feasibility of efficient plasmid DNA transfer into the central nervous system, providing a new option for treating various diseases such as tumors. PMID- 15269717 TI - Adenovirus expressing Fas ligand gene decreases airway hyper-responsiveness and eosinophilia in a murine model of asthma. AB - Allergic asthma is characterized by airway hyper-responsiveness (AHR) and cellular infiltration of the airway with predominantly eosinophils and Th2 cells. The normal resolution of inflammation in the lung occurs through the regulated removal of unneeded cells by Fas-Fas ligand-mediated apoptosis. Fas ligand (FasL) is a member of the tumor necrosis factor family, and when bound to Fas, it induces apoptosis of the cells. To examine the effect of the FasL gene on airway inflammation and immune effector cells in allergic asthma, recombinant adenovirus expressing murine FasL (Ad-FasL) was delivered intratracheally into ovalbumin (OVA)-immunized mice. We found that a single administration of Ad-FasL in OVA immunized mice significantly alleviated AHR and eosinophilia by inducing the apoptosis of eosinophils and/or reducing eosinophil attractant factors, such as IL-5 and eotaxin levels. The number of infiltrated lymphocytes and Th2 cytokines, including IL-5 and IL-13, decreased in OVA-immunized mice by administration of Ad FasL. KC and TNF-alpha production also decreased in Ad-FasL-treated OVA-immunized mice. These findings indicated that the administration of Ad-FasL to OVA sensitized mice significantly suppressed pulmonary allergic responses. Although more studies are needed, these results suggested that Ad-FasL might be applied as an alternative therapy for allergic asthma. PMID- 15269718 TI - The size of sinusoidal fenestrae is a critical determinant of hepatocyte transduction after adenoviral gene transfer. AB - The hepatotropism and intrahepatic distribution of adenoviral vectors may be species dependent. Hepatocyte transduction was evaluated in three rabbit strains after transfer with E1E3E4-deleted adenoviral vectors containing a hepatocyte specific alpha1-antitrypsin promoter-driven expression cassette (AdAT4). Intravenous administration of 4 x 10(12) particles/kg of AdAT4 induced human apo A-I levels above 40 mg/dl in Dutch Belt, but below 1 mg/dl in New Zealand White and Fauve de Bourgogne rabbits. Diameters of sinusoidal fenestrae were significantly (P=0.0014) larger in Dutch Belt (124+/-3.4 nm) than in New Zealand White (108+/-1.3 nm) and Fauve de Bourgogne (105+/-2.6 nm) rabbits, suggesting that a smaller size constitutes a barrier for hepatocyte transduction. Indeed, intraportal transfer preceded by intraportal injection of sodium decanoate, which increases the diameter of sinusoidal fenestrae to 123+/-3.4 nm (P<0.01) in New Zealand White rabbits, increased human apo A-I levels 32- and 120-fold in New Zealand White and Fauve de Bourgogne rabbits, respectively, but did not affect expression in Dutch Belt rabbits. In conclusion, size of sinusoidal fenestrae appears to be a critical determinant of hepatocyte transduction after adenoviral transfer. PMID- 15269719 TI - Genomic profiling of interpopulation diversity guides prioritization of candidate genes for autoimmunity. AB - Autoimmune diseases seem to have strong genetic attributes, and are affected to some extent by shared susceptibility loci. The latter potentially amount to hundreds of candidate genes (CG), creating the need for a prioritization strategy in genetic association studies. To form such a strategy, 26 autoimmune-related CG were genotyped for a total of 72 single nucleotide polymorphisms (SNPs) in three distinct Israeli ethnic populations: Ashkenazi Jews, Sephardic Jews and Arabs. Four quantitative criteria reflecting population stratification were analyzed: allele frequencies, haplotype frequencies, the Fst statistic for homozygotes distribution and linkage disequilibrium extents. According to the consequent interpopulation genomic diversity profiles, the genes were classified into conserved, intermediate and diversified gene groups. Our results demonstrate a correlation between the biological role of autoimmune-related CG and their interpopulation diversity profiles as classified by the different analyses. Annotation analysis suggests that genes more readily influenced by environmental conditions, such as immunological mediators, are 'population specific'. Conversely, genes showing genetic conservation across all populations are characterized by apoptotic and cleaving functions. We suggest a research strategy by which CG association studies should focus first on likely conserved gene categories, to increase the likelihood of attaining significant results and promote the development of gene-based therapies. PMID- 15269720 TI - Lipoxygenases at the heart of atherosclerosis susceptibility. PMID- 15269725 TI - Ignorance is not bliss. PMID- 15269727 TI - Los Alamos grinds to a halt after classified information vanishes. PMID- 15269726 TI - Drug research abused. PMID- 15269728 TI - Biologists seek to revamp biowarfare register. PMID- 15269729 TI - Budget delays threaten to leave US science in limbo. PMID- 15269730 TI - Health minister ignites row over drugs for HIV mothers. PMID- 15269731 TI - Stem-cell specialists split over proposal for US repository. PMID- 15269732 TI - Britain decides 'open access' is still an open issue. PMID- 15269733 TI - Space group touts 'next steps' for astronauts. PMID- 15269734 TI - Native Americans voice fears for relics in land-transfer deal. PMID- 15269735 TI - Project probes impact of waste carbon dioxide on marine life. PMID- 15269737 TI - Science and the war on drugs: a hard habit to break. PMID- 15269738 TI - Biodiversity: a tragedy with many players. PMID- 15269739 TI - Keeping a clear head on effects of illicit drugs. PMID- 15269740 TI - The animal-care regulatory system is a sham. PMID- 15269741 TI - Label of 'autism' could hold back gifted children. PMID- 15269742 TI - A question of balance. PMID- 15269748 TI - A radical reorientation. PMID- 15269749 TI - Earth science: deeper understanding. PMID- 15269750 TI - Language: children think before they speak. PMID- 15269751 TI - Palaeontology: echinoderm roots. PMID- 15269753 TI - Liquid crystals: a missing phase found at last? PMID- 15269754 TI - Animal behaviour: a social call. PMID- 15269755 TI - Obituary: Thomas Gold (1920-2004). PMID- 15269757 TI - Animal behaviour: coalition among male fiddler crabs. AB - Until now, no compelling evidence has emerged from studies of animal territoriality to indicate that a resident will strategically help a neighbour to defend its territory against an intruder. We show here that territory-owning Australian fiddler crabs will judiciously assist other crabs in defending their neighbouring territories. This cooperation supports the prediction that it is sometimes less costly to assist a familiar neighbour than to renegotiate boundaries with a new, and possibly stronger, neighbour. PMID- 15269759 TI - Developmental plasticity and human health. AB - Many plants and animals are capable of developing in a variety of ways, forming characteristics that are well adapted to the environments in which they are likely to live. In adverse circumstances, for example, small size and slow metabolism can facilitate survival, whereas larger size and more rapid metabolism have advantages for reproductive success when resources are more abundant. Often these characteristics are induced in early life or are even set by cues to which their parents or grandparents were exposed. Individuals developmentally adapted to one environment may, however, be at risk when exposed to another when they are older. The biological evidence may be relevant to the understanding of human development and susceptibility to disease. As the nutritional state of many human mothers has improved around the world, the characteristics of their offspring- such as body size and metabolism--have also changed. Responsiveness to their mothers' condition before birth may generally prepare individuals so that they are best suited to the environment forecast by cues available in early life. Paradoxically, however, rapid improvements in nutrition and other environmental conditions may have damaging effects on the health of those people whose parents and grandparents lived in impoverished conditions. A fuller understanding of patterns of human plasticity in response to early nutrition and other environmental factors will have implications for the administration of public health. PMID- 15269760 TI - Ancestral echinoderms from the Chengjiang deposits of China. AB - Deuterostomes are a remarkably diverse super-phylum, including not only the chordates (to which we belong) but groups as disparate as the echinoderms and the hemichordates. The phylogeny of deuterostomes is now achieving some degree of stability, especially on account of new molecular data, but this leaves as conjectural the appearance of extinct intermediate forms that would throw light on the sequence of evolutionary events leading to the extant groups. Such data can be supplied from the fossil record, notably those deposits with exceptional soft-part preservation. Excavations near Kunming in southwestern China have revealed a variety of remarkable early deuterostomes, including the vetulicolians and yunnanozoans. Here we describe a new group, the vetulocystids. They appear to have similarities not only to the vetulicolians but also to the homalozoans, a bizarre group of primitive echinoderms whose phylogenetic position has been highly controversial. PMID- 15269761 TI - An X-ray outburst from the rapidly accreting young star that illuminates McNeil's nebula. AB - Young, low-mass stars are luminous X-ray sources whose powerful X-ray flares may exert a profound influence over the process of planet formation. The origin of the X-ray emission is uncertain. Although many (or perhaps most) recently formed, low-mass stars emit X-rays as a consequence of solar-like coronal activity, it has also been suggested that X-ray emission may be a direct result of mass accretion onto the forming star. Here we report X-ray imaging spectroscopy observations which reveal a factor approximately 50 increase in the X-ray flux from a young star that is at present undergoing a spectacular optical/infrared outburst (this star illuminates McNeil's nebula). The outburst seems to be due to the sudden onset of a phase of rapid accretion. The coincidence of a surge in X ray brightness with the optical/infrared eruption demonstrates that strongly enhanced high-energy emission from young stars can occur as a consequence of high accretion rates. We suggest that such accretion-enhanced X-ray emission from erupting young stars may be short-lived, because intense star-disk magnetospheric interactions are quenched rapidly by the subsequent flood of new material onto the star. PMID- 15269762 TI - Single-shot read-out of an individual electron spin in a quantum dot. AB - Spin is a fundamental property of all elementary particles. Classically it can be viewed as a tiny magnetic moment, but a measurement of an electron spin along the direction of an external magnetic field can have only two outcomes: parallel or anti-parallel to the field. This discreteness reflects the quantum mechanical nature of spin. Ensembles of many spins have found diverse applications ranging from magnetic resonance imaging to magneto-electronic devices, while individual spins are considered as carriers for quantum information. Read-out of single spin states has been achieved using optical techniques, and is within reach of magnetic resonance force microscopy. However, electrical read-out of single spins has so far remained elusive. Here we demonstrate electrical single-shot measurement of the state of an individual electron spin in a semiconductor quantum dot. We use spin-to-charge conversion of a single electron confined in the dot, and detect the single-electron charge using a quantum point contact; the spin measurement visibility is approximately 65%. Furthermore, we observe very long single-spin energy relaxation times (up to approximately 0.85 ms at a magnetic field of 8 T), which are encouraging for the use of electron spins as carriers of quantum information. PMID- 15269763 TI - Electrical detection of the spin resonance of a single electron in a silicon field-effect transistor. AB - The ability to manipulate and monitor a single-electron spin using electron spin resonance is a long-sought goal. Such control would be invaluable for nanoscopic spin electronics, quantum information processing using individual electron spin qubits and magnetic resonance imaging of single molecules. There have been several examples of magnetic resonance detection of a single-electron spin in solids. Spin resonance of a nitrogen-vacancy defect centre in diamond has been detected optically, and spin precession of a localized electron spin on a surface was detected using scanning tunnelling microscopy. Spins in semiconductors are particularly attractive for study because of their very long decoherence times. Here we demonstrate electrical sensing of the magnetic resonance spin-flips of a single electron paramagnetic spin centre, formed by a defect in the gate oxide of a standard silicon transistor. The spin orientation is converted to electric charge, which we measure as a change in the source/drain channel current. Our set up may facilitate the direct study of the physics of spin decoherence, and has the practical advantage of being composed of test transistors in a conventional, commercial, silicon integrated circuit. It is well known from the rich literature of magnetic resonance studies that there sometimes exist structural paramagnetic defects near the Si/SiO2 interface. For a small transistor, there might be only one isolated trap state that is within a tunnelling distance of the channel, and that has a charging energy close to the Fermi level. PMID- 15269764 TI - Surface transfer doping of diamond. AB - The electronic properties of many materials can be controlled by introducing appropriate impurities into the bulk crystal lattice in a process known as doping. In this way, diamond (a well-known insulator) can be transformed into a semiconductor, and recent progress in thin-film diamond synthesis has sparked interest in the potential applications of semiconducting diamond. However, the high dopant activation energies (in excess of 0.36 eV) and the limitation of donor incorporation to (111) growth facets only have hampered the development of diamond-based devices. Here we report a doping mechanism for diamond, using a method that does not require the introduction of foreign atoms into the diamond lattice. Instead, C60 molecules are evaporated onto the hydrogen-terminated diamond surface, where they induce a subsurface hole accumulation and a significant rise in two-dimensional conductivity. Our observations bear a resemblance to the so-called surface conductivity of diamond seen when hydrogenated diamond surfaces are exposed to air, and support an electrochemical model in which the reduction of hydrated protons in an aqueous surface layer gives rise to a hole accumulation layer. We expect that transfer doping by C60 will open a broad vista of possible semiconductor applications for diamond. PMID- 15269765 TI - The elasticity of the MgSiO3 post-perovskite phase in the Earth's lowermost mantle. AB - MgSiO3 perovskite has been assumed to be the dominant component of the Earth's lower mantle, although this phase alone cannot explain the discontinuity in seismic velocities observed 200-300 km above the core-mantle boundary (the D" discontinuity) or the polarization anisotropy observed in the lowermost mantle. Experimental and theoretical studies that have attempted to attribute these phenomena to a phase transition in the perovskite phase have tended to simply confirm the stability of the perovskite phase. However, recent in situ X-ray diffraction measurements have revealed a transition to a 'post-perovskite' phase above 125 GPa and 2,500 K--conditions close to those at the D" discontinuity. Here we show the results of first-principles calculations of the structure, stability and elasticity of both phases at zero temperature. We find that the post-perovskite phase becomes the stable phase above 98 GPa, and may be responsible for the observed seismic discontinuity and anisotropy in the lowermost mantle. Although our ground-state calculations of the unit cell do not include the effects of temperature and minor elements, they do provide a consistent explanation for a number of properties of the D" layer. PMID- 15269766 TI - Theoretical and experimental evidence for a post-perovskite phase of MgSiO3 in Earth's D" layer. AB - The Earth's lower mantle is believed to be composed mainly of (Mg,Fe)SiO3 perovskite, with lesser amounts of (Mg,Fe)O and CaSiO3 (ref. 1). But it has not been possible to explain many unusual properties of the lowermost approximately 150 km of the mantle (the D" layer) with this mineralogy. Here, using ab initio simulations and high-pressure experiments, we show that at pressures and temperatures of the D" layer, MgSiO3 transforms from perovskite into a layered CaIrO3-type post-perovskite phase. The elastic properties of the post-perovskite phase and its stability field explain several observed puzzling properties of the D" layer: its seismic anisotropy, the strongly undulating shear-wave discontinuity at its top and possibly the anticorrelation between shear and bulk sound velocities. PMID- 15269767 TI - Audience drives male songbird response to partner's voice. AB - According to the social intelligence hypothesis, social context represents an important force driving the selection of animal cognitive abilities such as the capacity to estimate the nature of the social relationships between other individuals. Despite this importance, the influence of this force has been assessed only in primates and never in other animals showing social interactions. In this way, avian communication generally takes place in a network of signallers and receivers, which represents an audience altering individual signalling behaviours. Indeed, vocal amplitude and repertoire are known to be socially regulated and the attitude towards the opposite sex may change depending on the audience. This 'audience effect' provides support for the reality of social awareness in some bird species. However no evidence has yet been found to suggest that birds are able to estimate the characteristics of the social relationships between group-mates. Here we show that the male of a gregarious songbird species- the zebra finch (Taeniopygia guttata)--pays attention to the mating status of conspecific pairs, and uses this information to control its behaviour towards its female partner. PMID- 15269768 TI - Anthrax kills wild chimpanzees in a tropical rainforest. AB - Infectious disease has joined habitat loss and hunting as threats to the survival of the remaining wild populations of great apes. Nevertheless, relatively little is known about the causative agents. We investigated an unusually high number of sudden deaths observed over nine months in three communities of wild chimpanzees (Pan troglodytes verus) in the Tai National Park, Ivory Coast. Here we report combined pathological, cytological and molecular investigations that identified Bacillus anthracis as the cause of death for at least six individuals. We show that anthrax can be found in wild non-human primates living in a tropical rainforest, a habitat not previously known to harbour B. anthracis. Anthrax is an acute disease that infects ruminants, but other mammals, including humans, can be infected through contacting or inhaling high doses of spores or by consuming meat from infected animals. Respiratory and gastrointestinal anthrax are characterized by rapid onset, fever, septicaemia and a high fatality rate without early antibiotic treatment. Our results suggest that epidemic diseases represent substantial threats to wild ape populations, and through bushmeat consumption also pose a hazard to human health. PMID- 15269769 TI - Conceptual precursors to language. AB - Because human languages vary in sound and meaning, children must learn which distinctions their language uses. For speech perception, this learning is selective: initially infants are sensitive to most acoustic distinctions used in any language, and this sensitivity reflects basic properties of the auditory system rather than mechanisms specific to language; however, infants' sensitivity to non-native sound distinctions declines over the course of the first year. Here we ask whether a similar process governs learning of word meanings. We investigated the sensitivity of 5-month-old infants in an English-speaking environment to a conceptual distinction that is marked in Korean but not English; that is, the distinction between 'tight' and 'loose' fit of one object to another. Like adult Korean speakers but unlike adult English speakers, these infants detected this distinction and divided a continuum of motion-into-contact actions into tight- and loose-fit categories. Infants' sensitivity to this distinction is linked to representations of object mechanics that are shared by non-human animals. Language learning therefore seems to develop by linking linguistic forms to universal, pre-existing representations of sound and meaning. PMID- 15269770 TI - Drosophila long-term memory formation involves regulation of cathepsin activity. AB - Whereas short-term memory lasts from minutes to hours, long-term memory (LTM) can last for days or even an entire lifetime. LTM generally forms after spaced repeated training sessions and involves the regulation of gene expression, thereby implicating transcription factors in the initial steps of LTM establishment. However, the direct participation of effector genes in memory formation has been rarely documented, and many of the mechanisms involved in LTM formation remain to be understood. Here we describe a Drosophila melanogaster mutant, crammer (cer), which shows a specific LTM defect. The cer gene encodes an inhibitor of a subfamily of cysteine proteinases, named cathepsins, some of which might be involved in human Alzheimer's disease. The Cer peptide was found in the mushroom bodies (MBs), the Drosophila olfactory memory centre and in glial cells around the MBs. The overexpression of cer in glial cells but not in MB neurons induces a decrease in LTM, suggesting that Cer might have a role in glia and that the concentration of the Cer peptide is critical for LTM. In wild-type flies, cer expression transiently decreases after LTM conditioning, indicating that cysteine proteinases are activated early in LTM formation. PMID- 15269771 TI - Transmission of cutaneous leishmaniasis by sand flies is enhanced by regurgitation of fPPG. AB - Sand flies are the exclusive vectors of the protozoan parasite Leishmania, but the mechanism of transmission by fly bite has not been determined nor incorporated into experimental models of infection. In sand flies with mature Leishmania infections the anterior midgut is blocked by a gel of parasite origin, the promastigote secretory gel. Here we analyse the inocula from Leishmania mexicana-infected Lutzomyia longipalpis sand flies. Analysis revealed the size of the infectious dose, the underlying mechanism of parasite delivery by regurgitation, and the novel contribution made to infection by filamentous proteophosphoglycan (fPPG), a component of promastigote secretory gel found to accompany the parasites during transmission. Collectively these results have important implications for understanding the relationship between the parasite and its vector, the pathology of cutaneous leishmaniasis in humans and also the development of effective vaccines and drugs. These findings emphasize that to fully understand transmission of vector-borne diseases the interaction between the parasite, its vector and the mammalian host must be considered together. PMID- 15269772 TI - Reciprocal regulation of haem biosynthesis and the circadian clock in mammals. AB - The circadian clock is the central timing system that controls numerous physiological processes. In mammals, one such process is haem biosynthesis, which the clock controls through regulation of the rate-limiting enzyme aminolevulinate synthase 1 (Alas1). Several members of the core clock mechanism are PAS domain proteins, one of which, neuronal PAS 2 (NPAS2), has a haem-binding motif. Indeed, haem controls activity of the BMAL1-NPAS2 transcription complex in vitro by inhibiting DNA binding in response to carbon monoxide. Here we show that haem differentially modulates expression of the mammalian Period genes mPer1 and mPer2 in vivo by a mechanism involving NPAS2 and mPER2. Further experiments show that mPER2 positively stimulates activity of the BMAL1-NPAS2 transcription complex and, in turn, NPAS2 transcriptionally regulates Alas1. Vitamin B12 and haem compete for binding to NPAS2 and mPER2, but they have opposite effects on mPer2 and mPer1 expression in vivo. Our data show that the circadian clock and haem biosynthesis are reciprocally regulated and suggest that porphyrin-containing molecules are potential targets for therapy of circadian disorders. PMID- 15269773 TI - Role of transposable elements in heterochromatin and epigenetic control. AB - Heterochromatin has been defined as deeply staining chromosomal material that remains condensed in interphase, whereas euchromatin undergoes de-condensation. Heterochromatin is found near centromeres and telomeres, but interstitial sites of heterochromatin (knobs) are common in plant genomes and were first described in maize. These regions are repetitive and late-replicating. In Drosophila, heterochromatin influences gene expression, a heterochromatin phenomenon called position effect variegation. Similarities between position effect variegation in Drosophila and gene silencing in maize mediated by "controlling elements" (that is, transposable elements) led in part to the proposal that heterochromatin is composed of transposable elements, and that such elements scattered throughout the genome might regulate development. Using microarray analysis, we show that heterochromatin in Arabidopsis is determined by transposable elements and related tandem repeats, under the control of the chromatin remodelling ATPase DDM1 (Decrease in DNA Methylation 1). Small interfering RNAs (siRNAs) correspond to these sequences, suggesting a role in guiding DDM1. We also show that transposable elements can regulate genes epigenetically, but only when inserted within or very close to them. This probably accounts for the regulation by DDM1 and the DNA methyltransferase MET1 of the euchromatic, imprinted gene FWA, as its promoter is provided by transposable-element-derived tandem repeats that are associated with siRNAs. PMID- 15269774 TI - Periodic cycles of RNA unwinding and pausing by hepatitis C virus NS3 helicase. AB - The NS3 helicase is essential for cytoplasmic RNA replication by the hepatitis C virus, and it is a representative member of helicase superfamily 2 (SF2). NS3 is an important model system for understanding unwinding activities of DExH/D proteins, and it has been the subject of extensive structural and mutational analyses. Despite intense interest in NS3, the molecular and kinetic mechanisms for RNA unwinding by this helicase have remained obscure. We have developed a combinatorial, time-resolved approach for monitoring the microscopic behaviour of a helicase at each nucleotide of a duplex substrate. By applying this analysis to NS3, we have independently established the 'physical' and 'kinetic' step size for unwinding of RNA (18 base pairs, in each case), which we relate to the stoichiometry of the functional, translocating species. Having obtained microscopic unwinding rate constants at each position along the duplex, we demonstrate that NS3 unwinds RNA through a highly coordinated cycle of fast ripping and local pausing that occurs with regular spacing along the duplex substrate, much like the stepping behaviour of cytoskeletal motor proteins. PMID- 15269775 TI - Detection of an intermediate of photosynthetic water oxidation. AB - The oxygen that we breathe is produced by photosystem II of cyanobacteria and plants. The catalytic centre, a Mn4Ca cluster, accumulates four oxidizing equivalents before oxygen is formed, seemingly in a single reaction step 2H2O<==>O2 + 4H+ + 4e-. The energy and cycling of this reaction derives solely from light. No intermediate oxidation product of water has been detected so far. Here, we shifted the equilibrium of the terminal reaction backward by increasing the oxygen pressure and monitoring (by absorption transients in the near ultraviolet spectrum) the electron transfer from bound water into the catalytic centre. A tenfold increase of ambient oxygen pressure (2.3 bar) half-suppressed the full progression to oxygen. The remaining electron transfer at saturating pressure (30 bar) was compatible with the formation of a stabilized intermediate. The abstraction of four electrons from water was probably split into at least two electron transfers: mildly endergonic from the centre's highest oxidation state to an intermediate, and exergonic from the intermediate to oxygen. There is little leeway for photosynthetic organisms to push the atmospheric oxygen concentration much above the present level. PMID- 15269777 TI - Alternative paths. PMID- 15269778 TI - Making the move into drug sales. PMID- 15269780 TI - Recruiters and industry. Dual competencies. PMID- 15269782 TI - Genetic analysis of genome-wide variation in human gene expression. AB - Natural variation in gene expression is extensive in humans and other organisms, and variation in the baseline expression level of many genes has a heritable component. To localize the genetic determinants of these quantitative traits (expression phenotypes) in humans, we used microarrays to measure gene expression levels and performed genome-wide linkage analysis for expression levels of 3,554 genes in 14 large families. For approximately 1,000 expression phenotypes, there was significant evidence of linkage to specific chromosomal regions. Both cis- and trans-acting loci regulate variation in the expression levels of genes, although most act in trans. Many gene expression phenotypes are influenced by several genetic determinants. Furthermore, we found hotspots of transcriptional regulation where significant evidence of linkage for several expression phenotypes (up to 31) coincides, and expression levels of many genes that share the same regulatory region are significantly correlated. The combination of microarray techniques for phenotyping and linkage analysis for quantitative traits allows the genetic mapping of determinants that contribute to variation in human gene expression. PMID- 15269783 TI - The synthetic peptide PFWT disrupts AF4-AF9 protein complexes and induces apoptosis in t(4;11) leukemia cells. AB - The MLL gene at chromosome band 11q23 is commonly involved in reciprocal translocations detected in acute leukemias. A number of experiments show that the resulting MLL fusion genes directly contribute to leukemogenesis. Among the many known MLL fusion partners, AF4 is relatively common, particularly in acute lymphoblastic leukemia in infants. The AF4 protein interacts with the product of another gene, AF9, which is also fused to MLL in acute leukemias. Based on mapping studies of the AF9-binding domain of AF4, we have developed a peptide, designated PFWT, which disrupts the AF4-AF9 interaction in vitro and in vivo. We provide evidence that this peptide is able to inhibit the proliferation of leukemia cells with t(4;11) chromosomal translocations expressing MLL-AF4 fusion genes. Further, we show that this inhibition is mediated through apoptosis. Importantly, the peptide does not affect the proliferative capacity of hematopoietic progenitor cells. Our findings indicate that the AF4-AF9 protein complex is a promising new target for leukemia therapy and that the PFWT peptide may serve as a lead compound for drug development. PMID- 15269784 TI - Arsenic trioxide: expanding roles for an ancient drug? PMID- 15269785 TI - A clinical and pharmacological study of arsenic trioxide in advanced multiple myeloma patients. AB - We previously showed that arsenic trioxide (ATO) and melarsoprol may inhibit the growth of multiple myeloma (MM) cells in vitro and in vivo. We report here the administration of arsenic derivatives in 12 relapsing or refractory secretory MM patients. A total of 10 patients received ATO (eight in a continuous schedule, two discontinuously) and two received melarsoprol. The melarsoprol arm was prematurely closed due to toxicity. In the ATO arm, median duration of treatment was 38 days (9-54). Hepatic toxicity was grade 3 and 2 in one and eight patients, respectively. Other toxicities included neuropathy (n=2, grade 2), encephalitis (n=1, grade 3) and leuconeutropenia (n=4, grade 3). At 2 weeks after treatment initiation, mean serum concentration of arsenic was 1.11+/-0.16 micromol/l. No complete or partial remission was observed. A minor response (25-49% reduction of M protein in serum) and a stabilization of the M-protein level were observed in three and four patients, respectively. After ATO discontinuation, these responses were of short duration in all cases. ATO as a single agent did not produce any significant response in advanced MM patients despite sufficient arsenic exposure. Strategies to improve biodistribution, pharmacokinetic and efficacy of the drug as well as treatment combinations are needed. PMID- 15269786 TI - Ancient adaptive evolution of the primate antiviral DNA-editing enzyme APOBEC3G. AB - Host genomes have adopted several strategies to curb the proliferation of transposable elements and viruses. A recently discovered novel primate defense against retroviral infection involves a single-stranded DNA-editing enzyme, APOBEC3G, that causes hypermutation of HIV. The HIV-encoded virion infectivity factor (Vif) protein targets APOBEC3G for destruction, setting up a genetic conflict between the APOBEC3G and Vif genes. This kind of conflict leads to rapid fixation of mutations that alter amino acids at the protein-protein interface, referred to as positive selection. We show that the APOBEC3G gene has been subject to strong positive selection throughout the history of primate evolution. Unexpectedly, this selection appears more ancient than, and is likely only partially caused by, modern lentiviruses. Furthermore, five additional APOBEC genes in the human genome appear to be engaged in similar genetic conflicts, displaying some of the highest signals for positive selection in the human genome. Despite being only recently discovered, editing of RNA and DNA may thus represent an ancient form of host defense in primate genomes. PMID- 15269787 TI - An integrin-dependent role of pouch endoderm in hyoid cartilage development. AB - Pharyngeal endoderm is essential for and can reprogram development of the head skeleton. Here we investigate the roles of specific endodermal structures in regulating craniofacial development. We have isolated an integrinalpha5 mutant in zebrafish that has region-specific losses of facial cartilages derived from hyoid neural crest cells. In addition, the cranial muscles that normally attach to the affected cartilage region and their associated nerve are secondarily reduced in integrinalpha5- animals. Earlier in development, integrinalpha5 mutants also have specific defects in the formation of the first pouch, an outpocketing of the pharyngeal endoderm. By fate mapping, we show that the cartilage regions that are lost in integrinalpha5 mutants develop from neural crest cells directly adjacent to the first pouch in wild-type animals. Furthermore, we demonstrate that Integrinalpha5 functions in the endoderm to control pouch formation and cartilage development. Time-lapse recordings suggest that the first pouch promotes region specific cartilage development by regulating the local compaction and survival of skeletogenic neural crest cells. Thus, our results reveal a hierarchy of tissue interactions, at the top of which is the first endodermal pouch, which locally coordinates the development of multiple tissues in a specific region of the vertebrate face. Lastly, we discuss the implications of a mosaic assembly of the facial skeleton for the evolution of ray-finned fish. PMID- 15269788 TI - Cellular and genetic analysis of wound healing in Drosophila larvae. AB - To establish a genetic system to study postembryonic wound healing, we characterized epidermal wound healing in Drosophila larvae. Following puncture wounding, larvae begin to bleed but within an hour a plug forms in the wound gap. Over the next couple of hours the outer part of the plug melanizes to form a scab, and epidermal cells surrounding the plug orient toward it and then fuse to form a syncytium. Subsequently, more-peripheral cells orient toward and fuse with the central syncytium. During this time, the Jun N-terminal kinase (JNK) pathway is activated in a gradient emanating out from the wound, and the epidermal cells spread along or through the wound plug to reestablish a continuous epithelium and its basal lamina and apical cuticle lining. Inactivation of the JNK pathway inhibits epidermal spreading and reepithelialization but does not affect scab formation or other wound healing responses. Conversely, mutations that block scab formation, and a scabless wounding procedure, provide evidence that the scab stabilizes the wound site but is not required to initiate other wound responses. However, in the absence of a scab, the JNK pathway is hyperinduced, reepithelialization initiates but is not always completed, and a chronic wound ensues. The results demonstrate that the cellular responses of wound healing are under separate genetic control, and that the responses are coordinated by multiple signals emanating from the wound site, including a negative feedback signal between scab formation and the JNK pathway. Cell biological and molecular parallels to vertebrate wound healing lead us to speculate that wound healing is an ancient response that has diversified during evolution. PMID- 15269790 TI - Theory and numerical simulation of droplet dynamics in complex flows--a review. AB - We review theoretical and numerical studies and methods for droplet deformation, breakup and coalescence in flows relevant to the design of micro channels for droplet generation and manipulation. PMID- 15269791 TI - Principles of droplet electrohydrodynamics for lab-on-a-chip. AB - Electrically controlled droplet-based labs-on-a-chip operate under the principles of electro-capillarity and dielectrophoresis. The microfluidic mechanics of manipulating electrified droplets are complex and not entirely understood. In this article, we analyse these operating principles, especially electrowetting on dielectric (a form of electro-capillarity) and dielectrophoresis, under a unified framework of droplet electrohydrodynamics. We differentiate them by their electric origins and their energy transduction mechanisms. Our study shows that both electrowetting on dielectric and dielectrophoresis are effective for droplet generation and manipulation. In addition, our study demonstrates: (1) the presence of a wetting contribution to dielectrophoresis; and (2) contact angle reduction is merely an observable consequence of, not a condition for, the occurrence of electrowetting on dielectric. Simulations are used extensively in this article to illustrate device operation, to expose underlying physics, and to validate our conclusions. Simulations of electrically driven droplet generation, droplet translocation, droplet fusion, and droplet fission are presented. PMID- 15269792 TI - Transport and reaction in microscale segmented gas-liquid flow. AB - We use micro particle image velocimetry (microPIV) and fluorescence microscopy techniques to characterize microscale segmented gas-liquid flow at low superficial velocities relevant for chemical reactions with residence times of up to several minutes. Different gas-liquid microfluidic channel networks of rectangular cross section are fabricated in poly(dimethylsiloxane) (PDMS) using soft lithography techniques. The recirculation motion in the liquid segments associated with gas-liquid flows as well as the symmetry characteristics of the recirculations are quantified for straight and meandering channel networks. Even minor surface roughness effects and the compressibility of the gas phase induce loss of symmetry and enhance mixing across the centerline in straight channels. Mixing is further accelerated in meandering channels by the periodic switching of recirculation patterns across the channel center. We demonstrate a new, piezoelectrically activated flow injection technique for determining residence time distributions (RTDs) of fluid elements in multiphase microfluidic systems. The results confirm a narrowed liquid phase RTD in segmented flows in comparison to their single-phase counterparts. The enhanced mixing and narrow RTD characteristics of segmented gas-liquid flows are applied to liquid mixing and in sol-gel synthesis of colloidal nanoparticles. PMID- 15269793 TI - Laser techniques in acoustically levitated micro droplets. AB - Laser techniques were applied to an acoustically levitated droplet for remote investigation of the diameter, species concentration and temperature of the suspended droplet. To this end, the third and the fourth harmonic of a Nd:YAG laser were used for investigation of elastic, fluorescence and phosphorescence signals from the droplet. The droplet was seeded with thermographic phosphors and acetone for the phosphorescence and fluorescence measurements, respectively. The techniques were applied simultaneously using an imaging stereoscope. The imaging device allowed for an identical visualization of incoming signal through separate optical filters. Temperature measurements in droplets is important in the study of e.g. exothermic chemical reactions, spray processes, combustion, and in bioanalytical applications where the biological material is temperature sensitive or dependent on optimal temperature for function. Results from these investigations showed that temperature measurements in acoustically levitated droplets using laser-induced phosphorescence are feasible. The results also show the potential of simultaneous laser based measurements on levitated droplets. Diameter variation (surface area), mixture concentration and temperature were measured simultaneously. PMID- 15269794 TI - Design of microfluidic channel geometries for the control of droplet volume, chemical concentration, and sorting. AB - Passive microfluidic channel geometries for control of droplet fission, fusion and sorting are designed, fabricated, and tested. In droplet fission, the inlet width of the bifurcating junction is used to control the range of breakable droplet sizes and the relative resistances of the daughter channels were used to control the volume of the daughter droplets. Droplet fission is shown to produce concentration differences in the daughter droplets generated from a primary drop with an incompletely mixed chemical gradient, and for droplets in each of the bifurcated channels, droplets were found to be monodispersed with a less than 2% variation in size. Droplet fusion is demonstrated using a flow rectifying design that can fuse multiple droplets of same or different sizes generated at various frequencies. Droplet sorting is achieved using a bifurcating flow design that allows droplets to be separated base on their sizes by controlling the widths of the daughter channels. Using this sorting design, submicron satellite droplets are separated from the larger droplets. PMID- 15269795 TI - Dielectrophoresis-based programmable fluidic processors. AB - Droplet-based programmable processors promise to offer solutions to a wide range of applications in which chemical and biological analysis and/or small-scale synthesis are required, suggesting they will become the microfluidic equivalents of microprocessors by offering off-the-shelf solutions for almost any fluid based analysis or small scale synthesis problem. A general purpose droplet processor should be able to manipulate droplets of different compositions (including those that are electrically conductive or insulating and those of polar or non-polar nature), to control reagent titrations accurately, and to remain free of contamination and carry over on its reaction surfaces. In this article we discuss the application of dielectrophoresis to droplet based processors and demonstrate that it can provide the means for accurately titrating, moving and mixing polar or non-polar droplets whether they are electrically conductive or not. DEP does not require contact with control surfaces and several strategies for minimizing surface contact are presented. As an example of a DEP actuated general purpose droplet processor, we show an embodiment based on a scaleable CMOS architecture that uses DEP manipulation on a 32 x 32 electrode array having built-in control and switching circuitry. Lastly, we demonstrate the concept of a general-purpose programming environment that facilitates droplet software development for any type of droplet processor. PMID- 15269796 TI - An integrated digital microfluidic lab-on-a-chip for clinical diagnostics on human physiological fluids. AB - Clinical diagnostics is one of the most promising applications for microfluidic lab-on-a-chip systems, especially in a point-of-care setting. Conventional microfluidic devices are usually based on continuous-flow in microchannels, and offer little flexibility in terms of reconfigurability and scalability. Handling of real physiological samples has also been a major challenge in these devices. We present an alternative paradigm--a fully integrated and reconfigurable droplet based "digital" microfluidic lab-on-a-chip for clinical diagnostics on human physiological fluids. The microdroplets, which act as solution-phase reaction chambers, are manipulated using the electrowetting effect. Reliable and repeatable high-speed transport of microdroplets of human whole blood, serum, plasma, urine, saliva, sweat and tear, is demonstrated to establish the basic compatibility of these physiological fluids with the electrowetting platform. We further performed a colorimetric enzymatic glucose assay on serum, plasma, urine, and saliva, to show the feasibility of performing bioassays on real samples in our system. The concentrations obtained compare well with those obtained using a reference method, except for urine, where there is a significant difference due to interference by uric acid. A lab-on-a-chip architecture, integrating previously developed digital microfluidic components, is proposed for integrated and automated analysis of multiple analytes on a monolithic device. The lab-on-a chip integrates sample injection, on-chip reservoirs, droplet formation structures, fluidic pathways, mixing areas and optical detection sites, on the same substrate. The pipelined operation of two glucose assays is shown on a prototype digital microfluidic lab-on-chip, as a proof-of-concept. PMID- 15269797 TI - Multi-step synthesis of nanoparticles performed on millisecond time scale in a microfluidic droplet-based system. AB - This paper reports a plug-based, microfluidic method for performing multi-step chemical reactions with millisecond time-control. It builds upon a previously reported method where aqueous reagents were injected into a flow of immiscible fluid (fluorocarbons)(H. Song et al., Angew. Chem. Int. Ed., 2003, 42, 768). The aqueous reagents formed plugs--droplets surrounded and transported by the immiscible fluid. Winding channels rapidly mixed the reagents in droplets. This paper shows that further stages of the reaction could be initiated by flowing additional reagent streams directly into the droplets of initial reaction mixture. The conditions necessary for an aqueous stream to merge with aqueous droplets were characterized. The Capillary number could be used to predict the behavior of the two-phase flow at the merging junction. By transporting solid reaction products in droplets, the products were kept from aggregating on the walls of the microchannels. To demonstrate the utility of this microfluidic method it was used to synthesize colloidal CdS and CdS/CdSe core-shell nanoparticles. PMID- 15269798 TI - Poly(dimethylsiloxane) microchip: microchannel with integrated open electrospray tip. AB - A polymer microchip with an open tip for electrospray mass spectrometry is presented. The tip consists of a groove with parallel walls where a droplet can form at the end surface. A lid covers the whole chip except at the microchannel tip, which is left open. Poly(dimethylsiloxane) (PDMS) microchips were cast using a nickel mould which in turn was replicated from a dry etched silicon wafer. Tips with microchannel widths of around 50 microm could easily be replicated. Since the tip had no cover, the assembly of microchip and cover was simplified. A total ion current variation of 5% during 300 s was achieved for a 1 microM myoglobin solution. The non-complex design of the cover makes it suitable for versatile tests of chip prototypes. The nickel mould was found to be useful for PDMS microstructure fabrication. Also, such a robust mould allows casting electrospray tips in more rigid thermoset materials. PMID- 15269799 TI - Micro wet analysis system using multi-phase laminar flows in three-dimensional microchannel network. AB - A three-dimensional microchannel network with two-level crossings of channels was constructed in a glass microchip by sandwiching an insulating glass plate between two glass plates with microchannels followed by thermal bonding. Pressure-driven stable multi-phase laminar flows inside the three-dimensional channel network were realized by balancing flow rates of syringe pumps. Micro unit operations for mixing, reaction, solvent extraction, and detection were properly arranged in the multi-phase laminar flows, so that four parallel analyses, comprising twenty unit operations in total, could be integrated onto a single chip. Two chelating reagents and two sample solutions containing heavy metal ions (Fe(ii) or Co(ii)) were mixed and reacted in four different combinations using the three-dimensional channel network. After chelating reactions were completed, post processing (solvent extraction or addition of acid) was applied to each solution stream to remove the interferences of coexisting metal ions. Finally, target metal complexes were detected using a thermal lens microscope (TLM). Integrity of the micro system was confirmed by qualitative analysis of Fe(ii) and Co(ii). This is the first example of continuous flow chemical processing utilizing multi-phase laminar flow realized in a three-dimensional channel network. PMID- 15269800 TI - Micro patterning of active proteins with perforated PDMS sheets (PDMS sieve). AB - We propose a novel technique for patterning active proteins on a glass substrate using a perforated polydimethylsiloxane (PDMS) sheet-sieve. The sieve, which has tapering holes, is fabricated by spin-coating PDMS on a pyramidal-shaped mold. By means of this sieve, FITC (fluorescent isothiocyanate, bovine)-albumin was successfully spotted in a 5 x 5 microm(2) area in an array. The patterned spots were perfectly isolated, which eliminates the problem of non-specific binding of proteins to undesired areas. To show that proteins maintained their activity after the patterning, we used F(1)-ATPase biomolecular motors; their activity can easily be verified by observing their rotary motion after patterning. Selective patterning with three kinds of fluorescent micro beads indicated the possibility of patterning of different proteins on the same substrate by using the sieve. PMID- 15269801 TI - Filter-based microfluidic device as a platform for immunofluorescent assay of microbial cells. AB - A filter-based microfluidic device was combined with immunofluorescent labeling as a platform to rapidly detect microbial cells. The coin-sized device consisted of micro-chambers, micro-channels and filter weirs (gap = 1-2 microm), and was demonstrated to effectively trap and concentrate microbial cells (i.e., Cryptosporidium parvum and Giardia lamblia), which were larger in size than the weir gap. After sample injection, a staining solution containing fluorescently labeled antibodies was continuously provided into the device (flow rate = 20 microl min(-1)) to flush the microbial cells toward the weirs and to accelerate the fluorescent labeling reaction. Using a staining solution that was 10 to 100 times more dilute than the recommended concentration used in a conventional glass method, those target cells with a fluorescent signal-to-noise ratio of 12 could be microscopically observed at single-cell level within 2 to 5 min prior to secondary washing. PMID- 15269802 TI - Combinatorial mixing of microfluidic streams. AB - We have devised a microfluidic mixer design that produces all the mixture combinations of a given number of dilutions of the input compounds. As proof of the concept, we present a device that generates four titrations of two dye solutions, blue and yellow, and combinatorially mixes the blue titrations with the yellow titrations to deliver the sixteen mixture combinations in separate outlet microchannels. Our device features four different flow levels made by stacking nine laser-cut Mylar laminates. The fluidic network has a symmetric design that guarantees that the flow rates are the same at all the outlets, with deviations attributable to imperfections in the fabrication, assembly, or perfusion processes. Design rules for scaling up the number of compounds and/or dilutions are presented. The mixing scheme has broad applicability in high throughput combinatorial testing applications such as drug screening, cell-based biochemical assays, lab-on-a-chip devices, and biosensors. PMID- 15269803 TI - Injection and flow control system for microchannels. AB - In spite of considerable efforts, flow control in micro-channels remains a challenge owing to the very small ratio of channel/supply-system volumes, as well as the induction of spurious flows by extremely small pressure or geometry changes. We present here an inexpensive and robust system for flow control in a microchannel system, based on a dynamic control of reservoir pressures at the end of each channel. This system allows flow equilibration with a time constant smaller than one second, and is also able to maintain stable flux from stopped flow to many microl min(-1) range over several hours. It is robust to changes in ambient pressure and temperature. This system further includes a feature for sub microliter sample injection during the experiment. We quantify flow control in elastomer and thermoplastic channels, and demonstrate the impact on one application of the system, namely the reproducible, automated separation of large DNA by electrophoresis in a self-organized magnetic bead matrix in a microchannel. PMID- 15269804 TI - A microfluidic device to confine a single cardiac myocyte in a sub-nanoliter volume on planar microelectrodes for extracellular potential recordings. AB - A hybrid chip is described which combines a microfluidic network fabricated in a silicone elastomer (PDMS) with planar microelectrodes. It was used to measure extracellular potentials from single adult murine cardiac myocytes in a restricted extracellular space. The recorded variations in the extracellular potentials were caused by transmembrane currents associated with spontaneously initiated intracellular calcium waves. Single cells were trapped inside the 100 pl microchamber by pressure gradients and maintained for several hours by continuous perfusion. In addition, the localized delivery of drugs to a portion of the cell was demonstrated. The impedance of the electrodes was reduced by a factor of 10 to 20 after the electrodeposition of platinum black. Biopotentials recorded from single cells with platinum black electrodes showed a three-fold decrease in the noise, resulting in a maximum signal-to-noise ratio of 15:1. Characteristic variations in the frequency and shape of the extracellular potentials were observed among different cells which are most likely due to differences in the sarcoplasmic reticulum Ca(2+) load. Our device architecture permits the integration of electrochemical and optical sensors for multiparameter recordings. PMID- 15269805 TI - Efficient electrospray ionization from polymer microchannels using integrated hydrophobic membranes. AB - A simple process for realizing stable and reliable electrospray ionization (ESI) tips in polymer microfluidic systems is described. The process is based on the addition of a thin hydrophobic membrane at the microchannel exit to constrain lateral dispersion of the Taylor cone formed during ESI. Using this approach, ESI chips are shown to exhibit well-defined Taylor cones at flow rates as low as 80 nL min(-1) through optical imaging. Furthermore, stable electrospray current has been measured for flow rates as low as 10 nL min(-1) over several hours of continuous operation. Characterization of the electrospray process by optical and electrical monitoring of fabricated ESI chips is reported, together with mass spectrometry validation using myoglobin as a model protein. The novel process offers the potential for low-cost, direct interfacing of disposable polymer microfluidic separation platforms to mass spectrometry. PMID- 15269806 TI - Characterization of molecular transport in poly(dimethylsiloxane) microchannels for electrophoresis fabricated with synchrotron radiation-lithography and UV photolithography. AB - In the present paper, a study was undertaken of molecular transport in ploy(dimethylsiloxane) microchannels that were fabricated by ultraviolet (UV) photolithography and synchrotron radiation (SR)-lithography characterized and compared for microchip capillary electrophoresis by evaluating in-channel molecular dispersion. A fluorescent tag, sulforhodamine B was used as the probing molecule. It was found that microchannels made by SR-lithography fabrication were superior to those made by UV-photolithography fabrication in terms of molecular transport performance. A deep insight into surface conditions characterized by scanning electron microscopy suggested it was related to the difference in surface roughness. Chromatographic retention in electropherograms further supported such a conclusion, which depended on the phase ratio of the channel surface. The results revealed for PDMS microchannels in this work were in good agreement with the phenomenon found for glass microchannels in the literature. PMID- 15269807 TI - Measurements of scattered light on a microchip flow cytometer with integrated polymer based optical elements. AB - Flow cytometry is widely used for analyzing microparticles, such as cells and bacteria. In this paper, we report an innovative microsystem, in which several different optical elements (waveguides, lens and fiber-to-waveguide couplers) are integrated with microfluidic channels to form a complete microchip flow cytometer. All the optical elements, the microfluidic system, and the fiber-to waveguide couplers were defined in one layer of polymer (SU-8, negative photoresist) by standard photolithography. With only a single mask procedure required, all the fabrication and packaging processes can be finished in one day. Polystyrene beads were measured in the microchip flow cytometer, and three signals (forward scattering, large angle scattering and extinction) were measured simultaneously for each bead. To our knowledge this is the first time forward scattered light and incident light extinction were measured in a microsystem using integrated optics. The microsystem can be applied for analyzing different kinds of particles and cells, and can easily be integrated with other microfluidic components. PMID- 15269808 TI - Ceramic microreactors for heterogeneously catalysed gas-phase reactions. AB - The high surface to volume ratio of microchannel components offers many advantages in micro chemical engineering. It is obvious, however, that the reactor material and corrosion phenomena play an important role when applying these components. For chemical reactions at very high temperatures or/and with corrosive reactants involved, microchannel components made of metals or polymers are not suited. Hence, a modular microreactor system made of alumina was developed and fabricated using a rapid prototyping process chain. With exchangeable inserts the system can be adapted to the requirements of various reactions. Two heterogeneously catalysed gas-phase reactions (oxidative coupling of methane, isoprene selective oxidation to citraconic anhydride) were investigated to check the suitability of the system at temperatures of up to 1000 degrees C. Apart from the high thermal and chemical resistance, the lack of any blind activity was found to be another advantage of ceramic components. PMID- 15269809 TI - Development of a micro-fluidic manifold for copper monitoring utilising chemiluminescence detection. AB - The progressive development of a micro-fluidic manifold for the chemiluminescent detection of copper in water samples, based on the measurement of light emitted from the Cu(ii) catalysed oxidation of 1,10-phenanthroline by hydrogen peroxide, is reported. Micro-fluidic manifolds were designed and manufactured from polymethylmethacrylate (PMMA) using three micro-fabrication techniques, namely hot embossing, laser ablation and direct micro-milling. The final laser ablated design incorporated a reagent mixing channel of dimensions 7.3 cm in length and 250 x 250 microm in width and depth (triangular cross section), and a detection channel of 2.1 cm in length and 250 x 250 microm in width and depth (total approx. volume of between 16 to 22 microL). Optimised reagents conditions were found to be 0.07 mM 1,10-phenanthroline, containing 0.10 M cetyltrimethylammonium bromide and 0.075 M sodium hydroxide (reagent 1 delivered at 0.025 mL min(-1)) and 5% hydrogen peroxide (reagent 2 delivered at 0.025 mL min(-1)). The sample stream was mixed with reagent 1 in the mixing channel and subsequently mixed with reagent 2 at the start of the detection channel. The laser ablated manifold was found to give a linear response (R(2) = 0.998) over the concentration ranges 0 150 microg L(-1) and be reproducible (% RSD = 3.4 for five repeat injections of a 75 microg L(-1) std). Detection limits for Cu(ii) were found to be 20 microg L( 1). Selectivity was investigated using a copper selective mini-chelating column, which showed common cations found in drinking waters did not cause interference with the detection of Cu(ii). Finally the optimised system was successfully used for trace Cu(ii) determinations in a standard reference freshwater sample (SRM 1640). PMID- 15269810 TI - Rapid prototyping of polymer microsystems via excimer laser ablation of polymeric moulds. AB - This study presents a novel method for rapid prototyping of polymer microsystems. The method is based on excimer laser ablation of a thermally and mechanically stable polymer, such as PEEK (poly-ether-ether-ketone). A negative of the desired microsystem is laser machined in PEEK, which can then be used directly for hot embossing or injection moulding of a series of prototypes. This approach is very rapid and considerably cheaper than more traditional approaches to toolmaking, while still performing well in terms of reproduction of tool dimensions. The reduction in time and cost for a master tool using this method opens up new possibilities for testing small series in the R&D phase of a microsystem. Finally, two particular applications of the technique are presented. PMID- 15269811 TI - An AC electroosmotic micropump for circular chromatographic applications. AB - Flow rates of up to 50 microm s(-1) have been successfully achieved in a closed loop channel using an AC electroosmotic pump. The AC electroosmotic pump is made of an interdigitated array of unequal width electrodes located at the bottom of a channel, with an AC voltage applied between the small and the large electrodes. The flow rate was found to increase linearly with the applied voltage and to decrease linearly with the applied frequency. The pump is expected to be suitable for circular chromatography for the following reasons: the driving forces are distributed over the channel length and the pumping direction is set by the direction of the interdigitated electrodes. Pumping in a closed-loop channel can be achieved by arranging the electrode pattern in a circle. In addition the inherent working principle of AC electroosmotic pumping enables the independent optimisation of the channel height or the flow velocity. PMID- 15269812 TI - Bulk-micromachined submicroliter-volume PCR chip with very rapid thermal response and low power consumption. AB - The current paper describes the design, fabrication, and testing of a micromachined submicroliter-volume polymerase chain reaction (PCR) chip with a fast thermal response and very low power consumption. The chip consists of a bulk micromachined Si component and hot-embossed poly(methyl methacrylate)(PMMA) component. The Si component contains an integral microheater and temperature sensor on a thermally well-isolated membrane, while the PMMA component contains a submicroliter-volume PCR chamber, valves, and channels. The micro hot membrane under the submicroliter-volume chamber is a silicon oxide/silicon nitride/silicon oxide (O/N/O) diaphragm with a thickness of 1.9 microm, resulting in a very low thermal mass. In experiments, the proposed chip only required 45 mW to heat the reaction chamber to 92 degrees C, the denaturation temperature of DNA, plus the heating and cooling rates are about 80 degrees C s(-1) and 60 degrees C s(-1), respectively. We validated, from the fluorescence results from DNA stained with SYBR Green I, that the proposed chip amplified the DNA from vector clone, containing tumor suppressor gene BRCA 1 (127 base pairs at 11th exon), after 30 thermal cycles of 3 s, 5 s, and 5 s at 92 degrees C, 55 degrees C, and 72 degrees C, respectively, in a 200 nL-volume chamber. As for specificity of DNA products, owing to difficulty in analyzing the very small volume PCR results from the micro chip, we vicariously employed the larger volume PCR products after cycling with the same sustaining temperatures as with the micro chip but with much slower ramping rates (3.3 degrees C s(-1) when rising, 2.5 degrees C s(-1) when cooling) within circa 20 minutes on a commercial PCR machine and confirmed the specificity to BRCA 1 (127 base pairs) with agarose gel electrophoresis. Accordingly, the fabricated micro chip demonstrated a very low power consumption and rapid thermal response, both of which are crucial to the development of a fully integrated and battery-powered instrument for a lab-on-a-chip DNA analysis. PMID- 15269813 TI - Compact, high voltage power supply for the lab-on-a-chip. AB - The design, construction and operation of a simple, inexpensive and compact high voltage power supply (HVPS) for use in conjunction with a simple cross capillary electrophoresis microchip is presented. The microchip HVPS utilizes a single high voltage power supply (15 kV), a voltage-divider network, to give the voltages necessary to operate a gated injection valve, and two high voltage relays for switching between the open and closed gate sequences of the injection. In order to accommodate the application of different simple cross microchip dimensions, a set of equations for defining the resistor network and ensuring proper gate performance are presented. PMID- 15269814 TI - "Smart" mobile affinity matrix for microfluidic immunoassays. AB - There is a current need for simple methods for immobilizing biomolecules within microfluidic channels. Here, a technique is reported for reversibly immobilizing immunoassay components in a channel zone that can be simply controlled by integrated heating elements. Latex beads were modified with the temperature responsive polymer poly(N-isopropylacrylamide)(PNIPAAm) and co-modified with biotinylated poly(ethylene glycol)(PEG). PNIPAAm undergoes a hydrophilic-to hydrophobic transition when the temperature is raised above the lower critical solution temperature (LCST)( approximately 28 degrees C in the solutions used here). This reversible transition drives the aggregation and dis-aggregation of the modified beads in heated zones within poly(ethylene terephthalate)(PET) microchannels. Biotinylated monoclonal antibodies for the drug digoxin were bound via streptavidin to the biotin-PEG-coated beads. These antibody-functionalized beads were then reversibly immobilized by aggregation and hydrophobic adhesion to the surface of PET microfluidic channels in response to a thermal stimulus. The antibodies on the beads immobilized in the channel were shown to bind digoxin and a competitor fluorescent ligand from a flow stream in a quantitative competitive assay format that reported the digoxin concentration. The antibodies could be replenished for each immunoassay trial, using the reversible, temperature controlled immobilization process. This technique allows reagent immobilization immediately prior to an analytical procedure, following the removal of previously utilized beads, guaranteeing fresh and active immobilized biomolecules. Furthermore, it provides a simple approach to multiplexing through the simultaneous or sequential injection of different antibody-coated bead species, potentially at multiple sites in the integrated device channels. PMID- 15269815 TI - Shiga toxin-induced thrombotic microangiopathy: a thrombin-dependent process? PMID- 15269816 TI - Clopidogrel "resistance". PMID- 15269817 TI - Dr. Jekyll and Mr. Hyde: the many faces of wound healing regulators. PMID- 15269818 TI - Regulated de novo biosynthesis of fibrinogen in extrahepatic epithelial cells in response to inflammation. AB - Hepatic fibrinogen (FBG) is upregulated during an acute phase response (APR) induced by glucocorticoids and interleukin (IL)-6. Furthermore, intestine and lung epithelium synthesize FBG after exposure to inflammatory mediators, and both plasma and lung cell-derived FBG, along with fibronectin, assemble in detergent insoluble extracellular matrices (ECM) of pneumocytes and fibroblasts independent of thrombin or plasmin cleavage. An epitope cryptic in soluble FBG (beta(15-21)) but exposed in matrix-FBG and fibrin induces cell proliferation and actin cytoskeleton reorganization during wound repair and angiogenesis. Although fibrin(ogen) is involved in hemostasis and homeostasis, mechanisms regulating extrahepatic FBG expression remain unexplored. Herein we examined FBG production by lung compared to liver epithelial cell lines in response to dexamethasone (DEX)+IL-6. Regulated synthesis of HepG2-FBG follows the pathway shown for constitutive synthesis by liver epithelium. Constitutive A549-FBG expression was not detectable, however, intracellular FBG precursors in DEX+IL-6-treated A549 lung cells were similar to HepG2 cells with two notable exceptions. The relative rate of chain synthesis in HepG2 cells was unequal, whereas nascent synthesis of all three chains occurred at equivalent rates in stimulated A549 cells. Unlike HepG2 cells, which rapidly secreted intact FBG, nascent dimeric FBG accumulated in the A549 cell-associated fraction prior to release into medium. Furthermore, soluble A549-FBG was susceptible to thrombin and plasmin cleavage. Interestingly, many functionally diverse proteins possess FBG-related domains that direct cell fate determination during development or wound repair, suggesting that extrahepatic FBG biosynthesis evoked only during inflammation plays such a role during localized injury and repair to restore tissue homeostasis. PMID- 15269819 TI - Periodontal disease, but not edentulism, is independently associated with increased plasma fibrinogen levels. Results from a population-based study. AB - The systemic response to periodontal disease was analyzed in the cross-sectional Study of Health in Pomerania (SHIP). The completed data of 2,738 subjects aged 20 to 59 years were used for logistic regression analysis with an increased plasma fibrinogen level (> or =3.25 g/L according to Clauss) as the dependent variable. Participants were divided into four groups according to the number of periodontal pockets > or =4 mm (0, 1-7, 8-14, > or =15 pocketing). An additional group comprised the 52 edentulous subjects. The adjusted odds ratio (OR) of > or =15 periodontal pockets for increased plasma fibrinogen levels was 1.88 (95% CI: 1.25 2.83). Edentulism per se was not associated with increased plasma fibrinogen levels but was contained in a two-way interaction with the number of cigarettes/day in current smokers (p = 0.031). For edentulous nonsmokers the adjusted OR was 1.10 (95% CI: 0.51-2.39). Furthermore, body mass index, the interaction between gender and body mass index, serum LDL cholesterol, medication, the interaction between LDL cholesterol and medication, aspirin, smoking, school education, chronic bronchitis, and the interaction between alcohol consumption and chronic gastritis were associated with plasma fibrinogen levels. Our results show that periodontal disease but not edentulism per se is associated with an increased plasma fibrinogen level. PMID- 15269820 TI - Extracellular regulation of TGF-beta activity in wound repair: growth factor latency as a sensor mechanism for injury. AB - The transforming growth factor-beta (TGFbeta) family of growth factors are major regulators of wound repair, scar formation, and fibrosis. One of the prominent features of TGFbeta biology is the fact that this growth factor is secreted as a latent precursor, which may be directed to and stored at specific sites in the cellular microenvironment. Targeting and mobilization, and particularly extracellular activation of latent TGF-beta control the biological action of this growth factor. This review will focus on mechanisms of extracellular TGF-beta regulation relevant to and potentially operating in wound repair and scarring. PMID- 15269821 TI - Dissecting the roles of endothelin, TGF-beta and GM-CSF on myofibroblast differentiation by keratinocytes. AB - Myofibroblasts are specialized fibroblasts that contribute to wound healing by producing extracellular matrix and by contracting the granulation tissue. They appear in a phase of wound healing when the dermis strongly interacts with activated epidermal keratinocytes. Direct co-culture with keratinocytes upregulates TGFbeta activity and also induces fibroblast to differentiate into alpha-smooth muscle actin (alphaSMA)-positive myofibroblasts. TGF-beta activity alone cannot completely account for alphaSMA induction in these co-cultures, and here we analyze mechanical force generation, another potent inducer of myofibroblast differentiation in this model. Using deformable silicone substrates, we show that contractile activity of fibroblasts is already induced after 1-2-days of co-culture, when fibroblasts are generally alphaSMA negative. Endothelin-1 (ET-1), the most potent inducer of smooth muscle cell contraction, was up-regulated in co-cultures, while blocking ET-1 with the ET receptor inhibitor PD156252 inhibited contraction in these early co-cultures. In 4-5 days of co-culture, however, fibroblast contractile activity correlated with an increased expression of alphaSMA expression. Stimulation of fibroblast mono cultures with ET-1 in a low serum medium did not induce alphaSMA expression; however, ET-1 did synergize with TGF-beta. Surprisingly, GM-CSF, another mediatorstimulating myofibroblast differentiation in granulation tissue, inhibited alphaSMA expression in fibroblasts, costimulated with TGF-beta and ET 1. GM-CSF activated NFkappaB, thus interfering with TGF-beta signaling. Blocking TGFbeta and ET-1 largely impaired alphaSMA induction in co-cultures at day 7 and, in combination, almost completely prevented alphaSMA induction. Our results dissect the roles of TGF-beta and ET-1 on mechanical force generation in keratinocyte-fibroblast co-cultures, and identify GM-CSF as an inducer of myofibroblasts acting indirectly. PMID- 15269822 TI - Neutrophil function in the healing wound: adding insult to injury? AB - Cells of the innate immune system, including neutrophils and macrophages, are a highly visible component of normal wound healing in adult mammals. The role of inflammatory cells in the healing wound has been widely investigated, and evidence for both positive and negative influences exists. Several recent investigations support the emerging paradigm that robust inflammation is detrimental to wound closure. This developing information suggests that the functional role of inflammatory cells in wound healing must be reevaluated. PMID- 15269823 TI - Heparin binding protein is increased in chronic leg ulcer fluid and released from granulocytes by secreted products of Pseudomonas aeruginosa. AB - Recently it was demonstrated by Gautam, et al. that release of neutrophil-borne heparin-binding protein (HBP), also known as CAP37/azurocidin, induced endothelial hyperpermeability and neutrophil efflux. Here, we show that chronic leg ulcer fluid, in contrast to wound fluid from acute wounds, contains highly increased levels of HBP. Immunohistochemistry demonstrated the presence of HBP in chronic ulcer tissues. Furthermore, secreted products of Pseudomonas aeruginosa were found to induce release of HBP from human neutrophils. Our data suggest a possible link between bacterial presence and HBP-release in chronic ulcers. Thus, targeting HBP offers an interesting option for reduction of endothelial barrier dysfunction in chronic ulcers. PMID- 15269824 TI - Aberrant mucosal wound repair in the absence of secretory leukocyte protease inhibitor. AB - Secretory leukocyte protease inhibitor (SLPI) is a cationic serine protease inhibitor with anti-microbial and anti-inflammatory properties found in large quantities in mucosal fluids, including saliva. SLPI is expressed during cutaneous wound healing, however, its role in oral wound repair is unknown. We have used a novel approach involving a murine buccal mucosal acute wound model to investigate the role of SLPI in oral healing. In parallel to the observed cutaneous healing phenotype, an absence of SLPI results in markedly impaired oral wound healing associated with increased inflammation and raised elastase activity. Moreover, matrix deposition was decreased, while MMP activity was enhanced in the oral SLPI null wounds suggesting deregulated proteolysis. Intriguingly, regardless of genotype, reduced collagen deposition was observed in oral compared to dermal wounds, associated with reduced TGF-beta expression and decreased fibroblast collagen expression in vitro. We propose that SLPI is a pivotal endogenous factor necessary for optimal tissue repair including intra oral wound healing. In addition, our model provides a unique opportunity to delineate the cellular and molecular mechanisms underlying the differences between dermal scarring and oral scar-free healing. PMID- 15269825 TI - Thrombotic complications in patients with hereditary bleeding disorders. AB - Thromboses in patients with hereditary bleeding disorders are uncommon. However, in some cases, the co-existence of prothrombotic risk factors may increase the likelihood of developing thrombotic complications in such patients. This review summarizes the cases of thrombosis reported in the literature and analyzes the most important risk factors for thrombosis in patients with a congenital bleeding tendency. In particular we focus on central venous catheter (CVC)-associated thrombosis, on the thrombotic complications of coagulation factor concentrate therapy and on the presence of prothrombotic gene mutations. Data were identified by searches of the published literature, including PubMed, references from reviews and abstracts from the most important meetings on this topic. In conclusion, there is increasing evidence that thrombotic complications in patients with hereditary bleeding disorders have a multifactorial pathogenesis, depending on exogenous (coagulation factor replacement therapy, CVC, HIV infection) and/or endogenous (prothrombotic gene mutations) risk factors. PMID- 15269826 TI - Do prothrombotic factors influence clinical phenotype of severe haemophilia? A review of the literature. AB - There is considerable variability in bleeding patterns of severe haemophilia (<1% factor VIII). Knowledge of the contribution of thrombophilic factors in these patterns may improve individually tailored treatment strategies. We reviewed the literature regarding the relation between prothrombotic factors and clinical phenotype of severe haemophilia. Medline and EMBASE were searched for relevant articles. 9369 articles published between 1963 and September 2003 were screened and seven relevant papers were retrieved. Each of these reported on a different combination of thrombophilic factors. Presence of the factor V Leiden mutation appears to decrease the severity of severe haemophilia most consistently. Findings on other thrombophilic factors were inconclusive. There is a clear need for additional research on potential determinants of phenotypes of severe haemophilia before such knowledge can be translated into individual care for severe haemophilia patients with confidence. PMID- 15269827 TI - Pharmacokinetics of clopidogrel after administration of a high loading dose. AB - The adenosine diphosphate (ADP) receptor P2Y12 blocking agent clopidogrel is clinically proven to be efficient in preventing thrombotic events. However, its therapeutic value is limited by an, as yet poorly explained, interindividual heterogeneity in platelet inhibition. To evaluate possible pharmacokinetic determinants of this response variability, we developed a sensitive and specific liquid chromatography tandem mass spectrometry (LC-MS/MS) assay for quantification of unmodified inactive clopidogrel, its inactive carboxyl metabolite, and its active thiol metabolite in plasma. Analyte concentrations and platelet aggregation were assessed in ten healthy volunteers receiving an oral load of 600 mg clopidogrel. Subjects showed marked inter-individual differences in maximal platelet inhibition and in plasma pharmacokinetics. Univariate regression revealed linear correlations between maximal antiplatelet effect and peak plasma concentrations (cmax) of unchanged clopidogrel (r=0.76; p=0.01), of the carboxyl metabolite (r=0.70; p=0.03), and of the thiol metabolite (r=0.73; p=0.02), as well as linear correlations between cmax values of clopidogrel and its metabolites. This indicates that the response variability is predominantly caused by individual differences in clopidogrel absorption and that other factors, such as ADP receptor reactivity or differences in bioactivation of clopidogrel, do not play a major role. PMID- 15269828 TI - Expression of therapeutic levels of factor VIII in hemophilia A mice using a novel adeno/adeno-associated hybrid virus. AB - We have generated an E1a/E1b/E3-deleted adeno/adeno-associated (Ad/AAV) hybrid virus driven by a small nuclear RNA (pHU1-1) promoter for expression of a B domain-deleted (Thr761-Asn1639) factor VIII transgene (FVIIIDelta761-1639). Productive replication of Ad/AAV/FVIIIDelta761-1639 in AAV rep-expressing cells resulted in generation of monomeric and dimeric mini-adenoviral (mAd) replicative forms that retained the AAV integration elements (mAd/FVIIIDelta761-1639). In vitro studies using Ad/AAV/FVIIIDelta761-1639 generated approximately 2-logs greater FVIII activity than mAd/FVIIIDelta761-1639. To determine its capacity for in vivo excision and/or genomic integration, Ad/AAV/FVIIIDelta761-1639 was injected by tail vein into three groups of hemophilia A mice (2 x 10(11) vp [n = 3]; 4 x 10(11) vp [n = 3]; 8 x 10(11) vp [n = 3]), with clear concentration dependent increase in FVIII activity (range 160-510 mU/ml; plasma activity 16% 51% of normal). Peak activity was seen by Day (D) 5, with slow return to baseline by D28 (0.1-0.9% activity); in only 3/9 mice was loss of FVIII activity associated with development of anti-FVIII antibodies. Quantitative-PCR using genomic DNA isolated from D28 liver, spleen, heart, lungs, and kidney demonstrated the highest concentration in liver (approximately 10 genomes/cell), with little to no organ toxicity at early (D5 or 6) or late (D28) post-infusion time points. There was no evidence for spontaneous transgene excision or genomic integration in vivo as evaluated by quantitative PCR and genomic blotting. These data establish (i) the feasibility and applicability of developing high-titer Ad/AAV hybrid viruses for FVIII delivery using a small cellular promoter, (ii) the potential utility of this virus for generation of "gutted" monomeric and dimeric mAD/FVIII retaining AAV integration elements, and (iii) that the development of strategies for regulated Rep68/78 co-expression may provide a novel approach for excision, integration, and long-term FVIII transgene expression. PMID- 15269829 TI - Incidence of post-thrombotic syndrome and its association with various risk factors in a cohort of Spanish patients after one year of follow-up following acute deep venous thrombosis. AB - Post-thrombotic syndrome (PTS) is a frequent complication of deep venous thrombosis (DVT). However, neither the incidence nor the moment of PTS appearance are known. The main reason are the criteria used to define PTS, the characteristics of the patients, the study design and the time of follow-up. Our aims were to estimate the early incidence of PTS and its associated factors in a cohort of carefully defined DVT patients. 135 patients with a previous episode of acute idiopathic, phlebographically confirmed DVT, in the lower limbs, were followed up over 12 months. Phlebography was then repeated to determine the appearance of PTS. In addition, we used a validated clinical scale in order to assess the correlation between the clinical and phlebographical diagnosis of the PTS. This scale was applied at 6 and 12 months. The incidence of phlebographically confirmed PTS within the first year was 56.3% for the isolated PTS and 5.9% for PTS plus recurrent DVT, regardless of age, sex, platelet count, INR, or anticoagulation. None of these patients could be diagnosed as having PTS using the clinical validated scale. However, those patients with phlebographically diagnosed PTS had a higher clinical score than those without (P=0.012). The only factor related to a higher risk of developing a PTS was the localization of the DVT, subjects with both proximal and distal DVT having the highest incidence (P=0.001). In conclusion, although patients had appropriate anticoagulation, early incidence of PTS was very high, thus making it necessary to develop better diagnostic methods in order to evaluate the PTS impact. PMID- 15269830 TI - Postmenopausal hormone replacement and venous thromboembolism following hip and knee arthroplasty. AB - Hormone replacement therapy has been associated with venous thromboembolism. Controversy exists regarding the appropriate management of hormone replacement in the perioperative setting and in other situations--such as acute illness- predisposing to acute venous thromboembolism. We performed a case-control study to determine whether perioperative hormone replacement is associated with venous thromboembolism following hip and knee arthroplasty. 108 patients with postoperative venous thrombosis were matched by age, date and type of surgery, and surgeon with 210 controls without thrombosis. Perioperative hormone replacement use was no more prevalent in patients with postoperative thrombosis than those without. Eighteen (16.7%) women with post-operative thrombosis had taken perioperative hormone replacement compared to 49 (23.3%) of controls: odds ratio = 0.66; (95% CI 0.35-1.18; p=0.17). After multivariate analysis, the adjusted odds ratios were similar. Variables predicting post-operative thrombosis included: prior venous thromboembolism (OR = 2.3; p = 0.02), rheumatologic disease (OR = 2.2; p = 0.03), and absence of pharmacologic antithrombotic prophylaxis (OR = 13.4; p = 0.005). Cases and controls were otherwise similar. Users of hormone replacement were similar to non-users except that they were less likely to have coronary disease (OR 0.34; p = 0.03) or prior thrombosis (OR = 0.28; p = 0.04), and were younger (median age 67 versus 74 years; p <0.0001). We found no association between perioperative hormone replacement and post-operative thrombosis in patients undergoing major orthopaedic surgery. Routine discontinuation of these medications preoperatively--and possibly in other situations predisposing to thrombosis, such as acute medical illness--may be unnecessary in patients receiving appropriate pharmacologic antithrombotic prophylaxis. PMID- 15269831 TI - Genetic and environmental influences on the fibrinolytic system: a twin study. AB - The determination of heritability is a key issue to assess the predictive power of polymorphisms for disease in clinical studies. The aim of this study was to determine the heritability of proteins and activation markers of the fibrinolytic system in a large cohort of healthy twins. Heritability was calculated as 0.76 for thrombin activatable fibrinolysis inhibitor (TAFI), 0.44 for plasminogen activator inhibitor-1 (PAI-1), and 0.43 for tissue plasminogen activator. No significant genetic influence was observed for alpha2-antiplasmin-plasmin-complex and D-dimer. Heritability explained by single gene polymorphisms was 25.2% for TAFI 505G>A, 31.5% for 1542C>G, and 50.0% for combination of both. The influence on TAFI levels of 1542C>G (CC-->GG, median: -80.5%) was considerably stronger than that of 505G>A (GG-->AA, median: +49.3%) and in both cases there seems to be a dose-response relationship. Significant environmental influences on TAFI levels were observed for combined interaction terms (age*sex and bmi*sex). The PAI-1 4G/5G polymorphism explained 56.4% of the calculated heritability. The genetic variables accounting for the 43% heritability of tPA remain unknown. Our data show that the production of several key components of the fibrinolytic system is strongly genetically determined. This genetic influence is accounted for in large part but not completely by a limited number of polymorphisms within the respective genes associated with plasma levels of the gene products. PMID- 15269832 TI - Characterization of plasminogen variants in healthy subjects and plasminogen mutants in patients with inherited plasminogen deficiency by isoelectric focusing gel electrophoresis. AB - Plasmin(ogen) plays an important role in fibrinolysis and wound healing. Severe hypoplasminogenemia has recently been linked to ligneous conjunctivitis. Plasminogen (plg) is known as a polymorphic protein and most of these variants have been identified using isoelectric focusing (IEF) gel electrophoresis. Here, we studied common plg variants from healthy subjects and plg mutants from three patients with hypoplasminogenemia and three subjects with dysplasminogenemia by molecular genetic analysis and IEF. Analysis of 24 healthy subjects showed that subjects with the most common IEF plg phenotype A (n=12) were homozygous for aspartate at position 453 (453D), while both subjects with IEF plg phenotype B were homozygous for asparagine at this position (453N). Subjects with IEF plg phenotype AB (n=10) were compound-heterozygous for 453D/453N. Three patients with severe hypoplasminogenemia and different plg gene mutations exhibited characteristic "abnormal" IEF band patterns when compared with IEF plg phenotypes A and B. In all heterozygous family members the observed IEF plg phenotype was derived from the wild type plg molecule only, probably due to low concentration of the mutant plg molecule in plasma. In contrast, in three unrelated subjects with heterozygous dysplasminogenemia an equal "mixture" of wild type and mutant plg was found by IEF analysis. In conclusion, plg phenotyping by IEF in combination with molecular analysis of the plg gene seems to be a useful method for characterization of plg variants and mutants. PMID- 15269833 TI - Tissue plasminogen activator and neuroserpin are widely expressed in the human central nervous system. AB - Tissue plasminogen activator (tPA) is increasingly recognized to play important roles in various physiological and pathological processes in the central nervous system (CNS). Much of the data on the involvement of plasminogen activators in neurophysiology and -pathology have been derived from studies on experimental animals. We have now performed a systematic characterization of the expression of tPA and its inhibitor, neuroserpin, in normal human CNS. Brain and spinal cord samples from 30-36 anatomic locations covering all major brain regions were collected at 9 autopsies of donors with no neurological disease. Tissues were embedded in paraffin and tissue arrays were constructed. In two cases parallel samples were snap-frozen for biochemical analysis. Expression and activity profiling of tPA and neuroserpin were performed by immunohistochemistry, in situ hybridization, immunocapture and zymography assays. In the adult CNS, tPA was expressed at the mRNA and protein levels in many types of neurons, in particular in thalamus, cortex of cerebellum, pontine nuclei, neocortex, limbic system, and medulla oblongata. Interestingly, tPA was often co-expressed with its CNS inhibitor, neuroserpin. Despite overlapping expression of tPA and neuroserpin, zymography and immunocapture assays demonstrated that human neural tissue is a rich source of active tPA. Our analysis documents a detailed map of expression of tPA and its inhibitor in the human CNS and is compatible with the view that tPA is a key player in CNS physiology and pathology. PMID- 15269834 TI - Platelet dysfunction during Bothrops jararaca snake envenomation in rabbits. AB - Despite being well established that snake envenomation causes blood coagulation and fibrinolysis disturbances, scant information is available about blood platelet disorders. Herein we show that experimentally Bothrops jararaca envenomed rabbits presented thrombocytopenia, hypofibrinogenemia, elevation of von Willebrand factor plasma levels, platelet hypoaggregation in platelet rich plasma and whole blood, normoaggregation in washed platelet suspensions, decreased platelet ATP secretion, normal plasma levels of platelet factor 4, and constant intraplatelet serotonin levels. Furthermore, by flow cytometric analyses, platelets displayed a significant decrease in the expression of a GPIIb IIIa epitope recognized by P2 monoclonal antibody (p< 0.05) and an increased expression of a ligand-induced binding site (LIBS-1) of GPIIIa (p< 0.05), but total GPIIb-IIIa expression, evaluated with specific polyclonal antibodies, was normal. Fibrinogen and fibrin(ogen) degradation product (FfDP) expression on platelet surface showed no significant alteration. Nonetheless, significant elevations of platelet P-selectin were noticed on circulating platelets. The percentage of circulating reticulated platelets and the survival time of biotinylated platelets of envenomed rabbits were not statistically different from control animals. We suggest that thrombin engendered by procoagulating components of B. jararaca venom has an essential role in the pathogenesis of platelet and coagulation disorders in this experimental model. Increased expression of P selectin in the experimental group demonstrates that platelets of envenomed rabbits are indeed activated in circulation, and that decreased fibrinogen or increased FfDP levels are not the primary cause of platelet dysfunction. These results imply the existence of an inhibitor in plasma that interferes with platelet aggregation in bothropic envenomation. PMID- 15269835 TI - Platelet glycoprotein Ibalpha polymorphisms modulate the risk for myocardial infarction. AB - Platelet glycoprotein Iba (GPIba) gene polymorphisms have been reported to affect the risk of developing coronary heart disease. Here, within the GPIba gene, we determine the association between the variable number of tandem repeats (VNTR), the -5C/T Kozak sequence dimorphism, and the human platelet antigen (HPA)-2 polymorphisms with occurrence of myocardial infarction (MI). Patients (n=180) presenting survivors of MI were compared to 180 controls matched by age, gender, and race. Carriers of VNTR-CD genotype had a 2-fold higher risk for MI compared to controls. The prevalence of VNTR-BC was lower among patients than among controls (P=.007). These data are in agreement with recent reports of increased plug formation by human platelets containing VNTRCD but no other VNTR genotypes. Among patients, the number of vessels severely occluded was greater among carriers of the D-allele (P=.019) or VNTR-CD (P=.026) and lower among carriers of the C-allele (P=.003) or VNTR-CC (P=.0009) compared to non-carriers of these alleles. No influence was seen with the Kozak or HPA-2 polymorphisms. Determination of VNTR of the GPIba gene may prove useful for identifying high risk individuals for MI. PMID- 15269836 TI - Lepirudin prevents lethal effects of Shiga toxin in a canine model. AB - Microvascular thrombosis is a major cause of organ damage in Shiga toxin-mediated hemolytic uremic syndrome (Stx-HUS). In vitro and clinical studies implicate thrombin-mediated mechanisms in the pathogenesis of Stx microvascular thrombosis. In a greyhound model, administration of 0.03 microg/kg to 0.05 microg/kg Stx1 or Stx2 causes severe bloody diarrhea and HUS with microvascular thrombosis requiring humane euthanasia within 65 hours. Using a greyhound model of Stx-HUS we analyzed early hemostatic changes, and tested the hypothesis that thrombin blockade with lepirudin would prevent lethal Stx effects. Two Stx1-exposed greyhounds were analyzed for hemostatic changes prior to onset of clinical manifestations. Serial hemostasis studies after Stx1 challenge revealed trends of increased aPTT, fibrinogen levels, and prothrombin fragment 1+2, and appearance of abnormally large von Willebrand factor multimers. Three greyhounds were anticoagulated with lepirudin to maintain activated partial thromboplastin times (aPTT) >2.5-fold normal, followed by administration of Stx2 and observation of clinical responses. Among the 3 lepirudin-treated, Stx2-challenged greyhounds, one developed severe illness requiring euthanasia. Remarkably, 2 of the 3 greyhounds developed only hypersalivation and restlessness that resolved (P <.03 compared to 14 historical controls). These two greyhounds were clinically, hematologically and biochemically normal 74 hours after Stx administration, well beyond the time of euthanasia of any previous greyhound. This study suggests that greyhounds exposed to Stx develop procoagulant changes similar to humans, and that thrombin may be a critical factor in the pathogenesis and treatment of Stx HUS. PMID- 15269837 TI - The functional role of blood platelet components in angiogenesis. AB - The process of neovascularization greatly depends on the induction of the angiogenic phenotype of endothelial cells that is strictly controlled by humoral factors as well as by cellular communications in the vascular system. Although blood platelets contain several secretable pro- and antiangiogenic components, their overall role in angiogenesis remains poorly understood. In a mouse model of hypoxia-induced retinal angiogenesis, the situation of thrombocytopenia as well as inhibition of platelet aggregation by a highly specific alphaIIbbeta3-integrin antagonist or acetyl salicylic acid (Aspirin) administration, respectively, resulted in about 35-50% reduction of retinal neovascularization, compatible with a significant contribution of blood platelets in angiogenesis. Platelet remnants and microvesicles were found at sites of angiogenic sprouts. In vitro isolated platelets incorporated in a fibrin gel induced capillary sprouting of microvascular endothelial cells. Similarly, platelet releasate elevated the permeability of confluent endothelial cell monolayers to the same extent as hypoxia did. Platelet-derived VEGF as well as butanol-extractable lipid mediators were identified as predominant activators of angiogenesis, particularly of microvascular endothelial cell proliferation and migration. In addition, a synergistic effect between platelet-derived VEGF and bFGF in capillary sprouting and endothelial cell proliferation was found. Based on this proangiogenic role of platelets in neovascularization, anti-platelet substances can be considered as potent inhibitors of angiogenesis. PMID- 15269838 TI - Histidine-proline rich glycoprotein (HPRG) binds and transduces anti-angiogenic signals through cell surface tropomyosin on endothelial cells. AB - The anti-angiogenic properties of the histidine-proline-rich (H/P) domain of HPRG have recently been described (Juarez JC, et al. Cancer Research 2002; 62: 5344 50). However, the binding site that mediates these properties is unknown. HPRG is evolutionarily, functionally and structurally related to cleaved high molecular weight kininogen (HKa), an anti-angiogenic polypeptide that stimulates apoptosis of proliferating endothelial cells through binding to cell-surface tropomyosin (Zhang J-C, et al. Proc Natl Acad Sci USA 2002; 99: 12224-9). In this study, we demonstrate that HPRG binds with high affinity to FGF-2-stimulated human umbilical vein endothelial cells (HUVEC) and immobilized tropomyosin in a Zn2+ or pH-dependent manner, and that this interaction is mediated by the H/P domain of HPRG. At least two binding sites for HPRG, tropomyosin and heparan sulfate proteoglycans (HSPs), were identified on the surface of FGF-2-activated endothelial cells. Translocation of tropomyosin to the surface of HUVEC occurred in response to FGF-2, and the anti-angiogenic activity of HPRG in a Matrigel plug model was partially inhibited by soluble tropomyosin. These results suggest that HPRG binds to endothelial cell surface tropomyosin which at least partially mediates the antiangiogenic effects of HPRG. PMID- 15269839 TI - Association of NOS3 gene with metabolic syndrome in hypertensive patients. AB - Recent data from animal models indicate that the eNOS null mice present a phenotype that resemble the human metabolic syndrome (hypertension, insulin resistance and hypertriglyceridemia). In this work, we have studied whether NOS3 gene, previously related to endothelial dysfunction, might have a role in metabolic syndrome susceptibility in hypertensive patients. To carry out the study, we genotyped 105 hypertensive patients < or = 60 years old with two polymorphisms of NOS3 gene: 1132 T>C and 7164 G>T (GeneBank:AF519768.1). To check the allelic frequency of these polymorphisms in our geographical area, we also genotyped 94 unselected healthy controls (control group). To perform sample genotyping, we designed a novel FRET system coupled to real time PCR. There were no differences in genotypic distribution or allelic frequency between hypertensive patients and the control group. However, we observed that 786CC genotype was significantly more frequent in hypertensive patients with metabolic syndrome than in those without the syndrome (p=0.0022). When both polymorphisms were analyzed, we identified the 786C894G as the risk haplotype for metabolic syndrome susceptibility (p=0.011). These data suggest a role of the NOS3 gene in the pathogenesis of metabolic syndrome in hypertensive patients. PMID- 15269840 TI - CD14+CD16+ monocytes in coronary artery disease and their relationship to serum TNF-alpha levels. AB - Monocytes play a central role in the inflammatory disease atherosclerosis. CD14+CD16+ monocytes are considered proinflammatory monocytes, as they have an increased capacity to produce proinflammatory cytokines, such as TNF-alpha, and are elevated in various inflammatory diseases. We hypothesized that patients with coronary artery disease (CAD) have increased levels of CD14+CD16+ monocytes, and that CD14+CD16+ monocytes are associated with inflammation markers. We investigated CD14+CD16+ monocytes in 247 patients with CAD and 61 control subjects using flow cytometry. In addition serum concentrations of TNF-alpha, IL 6, and Hs-CRP were assessed. Patients with CAD had higher levels of CD14+CD16+ monocytes than controls (13.6% versus 11.4%; p<0.001). Logistic regression analysis including quartiles of CD14+CD16+ monocytes showed that CD14+CD16+ monocytes were associated with prevalence of CAD (OR 4.9, 95% CI 2.5-19.1, for subjects in the fourth quartile in comparison to subjects in the first quartile). The association between CD14+CD16+ monocytes and CAD remained independently significant after adjustment for most potential confounders (OR 5.0, 95% CI 1.2 20.0). Serum concentrations of TNF-alpha were elevated in subjects within the highest quartiles of CD14+CD16+ monocytes (p=0.018). Our study showed that increased numbers of CD14+CD16+ monocytes are associated with coronary atherosclerosis and TNF-alpha. In accordance, recent animal studies suggest a possibly important role of these monocytes in the development of atherosclerosis. PMID- 15269841 TI - Acute myocardial infarction during substitution with recombinant factor VIII concentrate in a patient with mild haemophilia A. PMID- 15269842 TI - Charge analysis of N-glycans from human recombinant coagulation factor VIII and human FVIII standards. PMID- 15269843 TI - Significantly lower need for blood transfusions associated with post-operatively initiated subcutaneous melagatran/oral ximelagatran compared with enoxaparin. PMID- 15269844 TI - Evaluation of a Russell's viper venom-based protein C assay. PMID- 15269845 TI - The role of angiotensin-converting enzyme gene polymorphism in pulmonary thromboembolism. PMID- 15269846 TI - Addendum to: The association between adverse pregnancy outcomes and maternal factor V Leiden genotype. A meta-analysis. PMID- 15269847 TI - [The measurement parameters and its significance of diagnostic tests]. AB - Diagnostic tests play an important part in both clinical and research fields. With a great number of statistical methods, different measurement parameters have been applied to different diagnostic tests. Selecting the appropriate measurement parameters is of the utmost importance for clinicians and researchers in their studies. This paper is aimed to help the clinicians and researchers to have a comprehensive understanding of the right measurement parameters, interpretation of the statistical outcomes accurately in their researches, and evaluation of the diagnostic tests objectively, with an example of the study on magnetic resonance arthrography applied to the diagnosis of intraarticular adhesions of TMJ. The measurement parameters include sensitivity, specificity, accuracy, predictive value, prevalence, likelihood ratio, and receiver operator characteristic curve. PMID- 15269848 TI - [CT imaging analysis in 138 cases with orbital fractures]. AB - PURPOSE: To investigate CT findings of orbital fractures in maxillofacial injuries in order to improve clinical diagnostic accuracy. METHODS: CT findings of 138 cases (145 orbits) with orbital fractures in maxillofacial injuries were analyzed. RESULTS: In maxillofacial injuries, orbital fractures of medial and posterolateral walls were most common,which often showed multiple walls fractures. In 145 orbital fractures, there were single wall fractures in 51 cases (35.17%), two-wall fractures in 48 cases (33.10%), three-wall fractures in 31 cases (21.38%) and four-wall fractures in 15 cases (10.35%). CONCLUSIONS: There were complex orbital fracture in maxillofacial injures, CT examination is a very important tool for diagnosis,which should be applied routinely. PMID- 15269849 TI - [The study on expression of bFGF and quantity of mast cell in infant hemangiomas of grandulae parotid gland]. AB - The qualitative and quantitative study on mast cells and bFGF in 40 cases of infant hemangiomas of the grandulae parotid and 5 cases of normal grandulae parotid tissues were investigated by special mast cell staining methods, immunohistochemistry and image analysis technique. The quantity, area and average density of MC remarkably increased, there were significant differences. The mast cells mainly distributed around premature blood vessels and in parotid gland leaves; bFGF was highly expressed in infant hemangiomas of grandulae parotid. Its immuno-reactivity was present in the cytoplasm of endotheliocytes,mainly in extracellular matrix, distributed in light reticulation. It is concluded that there was a close relationship between bFGF and mast cell in proliferation of infant hemangiomas of grandulae parotid. PMID- 15269850 TI - [Magnetic resonance arthrography for the diagnosis of intraarticular adhesions of TMJ adhesions]. AB - PURPOSE: To evaluate the value of magnetic resonance arthrography (MRAr) on the diagnosis of intraarticular adhesions of the temporomandibular joint(TMJ). METHODS: 30 consecutive patients (31 TMJs) diagnosed as internal derangement of TMJ were examined by MRAr and arthroscopy. The findings of interpreting MRAr were recorded as positive, suspicious and negative according to the MRAr radiographic criteria. After comparing the findings of MRAr with those of arthroscopy, the numbers of true positive, true negative,false positive and false negative were obtained. Through SPSS11.0, receiver operator characteristic curve(ROC curve) was made, and the area under the ROC curve was calculated. According to the area, the diagnostic accuracy of MRAr was evaluated. RESULTS: Surgery results confirmed that 22 TMJs in 31 TMJs had adhesions. According to the above criteria, we found 20 TMJs were positive, 3 TMJs were suspicious, and 8 TMJs were negative. After comparing the findings of MRI with those of arthroscopy, true positive in 19 TMJs and false positive in 1 TMJs were detected among 20 positive TMJs, adhesion in 2 TMJs and no adhesion in 1 TMJs were detected among 3 suspicious TMJs, and true negative in 7 TMJs and false negative in 1 TMJs were detected among 8 negative TMJs. The area under ROC curve was 0.91(0.86,0.95), P<0.05. CONCLUSION: It is concluded that: (1)The diagnostic accuracy of MRAr is excellent, much better than that of PFA and MRI. (2) MRAr is an ideal method to diagnose intraarticular adhesions of TMJ. PMID- 15269851 TI - [Three-dimensional reconstruction on facial soft tissue using three-dimensional grating projection technique]. AB - PURPOSE: The purpose of this study is to establish a method to reconstruct the facial soft tissue three-dimensionally. METHODS: By ATOS three-dimensional opticalmetry, which used in industry, 3D data on facial profile of a patient with mandibular hyperplasia was achieved quickly. RESULTS: Three-dimensional data of facial profile was accurate and the images were reconstructed with verisimilitude. CONCLUSIONS: Traditional device of grating projection was modified by using two CCD cameras to obtain grating images.3D data on facial profile were achieved quickly and accurately. PMID- 15269852 TI - [The investigation and analysis of children with dental fear]. AB - PURPOSE: The purpose of this study was to investigate and analyze the clinical behavior and factors causing dental fear (DF) in children of different ages to provide some valuable information for preventing and reducing the incidence of DF in clinical practice. METHODS: 571 cases of children patients receiving treatment in the pedodontics were evaluated and analyzed according to the evaluation standards of DF. RESULTS: The results suggested that the incidence of DF differed in different age groups. The ratio was higher in younger groups while it was lower in elder groups. The ratio also varied with living conditions,education level of mothers,past experience of dental treatment and other children's response at the clinic. CONCLUSION: The DF caused by various factors can be reduced by the combined effort of children, parents and the medical practitioners. PMID- 15269853 TI - [The effect of coronal preflaring on the working length measurement of root canals in molars]. AB - PURPOSE: To study the influence of coronal preflaring of root canal on the working length measurement in molars. METHODS: 48 molars from 47 adult patients with pulpitis were divided into two groups randomly. 74 root canals of 24 molars were examined in Group 1. After preparing an access cavity, the root canal orifices were located. A 15# K-type file was inserted into root canal to detect resistance at the apical region. 75 root canals of 24 molars were examined in Group 2. Before testing the apical resistance,a size 25# and 20# ProFile.06 taper NiTi rotary instrument was used to enlarge the canal orifice and flare the coronal portion of the root canal. After placing 15# K-type files, a radiograph was taken with the bisecting-angle technique. The distance between the tip of the file and the radiographic apex was measured on the radiographs. Statistical analysis of univariate and multiple logistic regression were carried out for analysis. RESULTS: There was a significant difference between the two groups (P<0.05), there were more accurate measurements in Group 2 than in Group 1. No statistical correlation was found between X-ray measurements and age, sex and the position of molars,while there was significant correlation between maxillary molars and mandibular molars (P<0.05). CONCLUSION: Preflaring the coronal portion of the root canals could significantly increase the accurate measurements of the working length of root canals in molars. PMID- 15269854 TI - [Association of HLA-DRB1*1501 polymorphism with the susceptibility to severe chronic periodontitis in Chinese Han nationality]. AB - PURPOSE: To investigate the association of HLA-DRB1*1501 polymorphism with the susceptibility to severe chronic periodontitis (CP) in Chinese Han nationality. METHODS: DNA samples were collected with buccal swabs from 134 subjects with severe chronic periodontitis and 81 periodontal healthy control. HLA-DRB1*1501 genotype was determined using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). The genotype distribution was statistically analyzed by chi2 test. RESULTS: The allele frequencies of DRB1*1501 was detected more frequently in severe CP patients than in the controls [OR=3.874 (2.401 6.252), P<0.05]. The 1501/1501 genotype frequencies were significantly increased in patients compared with the reference population [OR=20.896 (4.866-89.726), P<0.05]. CONCLUSION: The results suggest that HLA-DRB1*1501 allele may be a risk indicator for the susceptibility to severe CP in Han nationality (P<0.05). The HLA-DRB1*1501 homozygote genotype was associated significantly with severe CP (P<0.05). PMID- 15269855 TI - [The test of metal-ceramic bonding strength among three ceramic alloies]. AB - PURPOSE: To compare the metal-ceramic bonding strength of different ceramic alloies. METHODS: 30 wax sheets were divided into 6 groups at random. Each group included 5 sheets. After being invested conventionally with Bellaves SH,the samples were casted and fused with porcelain.The metal-ceramic bonding strength was evaluated. RESULTS: The highest bonding strength was found in Bio Herador N alloy, and then in TILITE alloy and Heraenium S alloy respectively. Bio Herador N alloy and TILITE alloy all had significant differences of the bonding strength before and after treatment (P<0.05). There was no significant differences in the bonding strength of the Heraenium S alloy after treatment (P>0.05). CONCLUSION: The bonding strength of precious metal-ceramic was higher than that of non precious metal-ceramic.Pre-oxygen treatment can improve the metal-ceramic bonding strength of Bio Herador N alloy and TILITE alloy,but it had no effect on the metal-ceramic bonding strength of the Heraenium S alloy. PMID- 15269856 TI - [Metal elements from intraoral metal restorations]. AB - PURPOSE: To detect the metal element from intraoral metal restorations. METHODS: A small sample was taken from each intraoral metal restorations. Energy dispersive X ray fluorescence spectrometry (EDXRF) was used to detect the metal elements from them. RESULTS: From 14 suspected patients of dental metal allergy, 120 intraoral metal restorations were detected. The ten most frequent elements detected,in decreasing order of frequency, were Ag, Cu, Au, Pd, Zn, Sn, Co, Cr, In and Mo. The types and percentages of alloys containing these elements were Au Ag-Pd alloys (65%), Ag-Sn-Hg amalgams (2.5%), Ag alloys (5.4%) , Au alloys (14.6%), Ni-Cr alloys and Co-Cr alloys (10.1%), other alloys (2.4%). In these, Ag Sn-Hg amalgams (2.5%) are in lower using rate than 10 years before (16%). Ni, Co, Cr alloys are in same using rate as 10 years before. There are 12 patients with two to five kinds of different alloys present at an oral environment. In one of 12 patients,five kinds of different alloys were present within an oral environment. CONCLUSIONS: In patients suspicious of dental metal allergy, co existence of different metals alloys was often detected, which indicated that in order to prevent dental metal allergy, heterogeneous metal and metal alloys containing Hg and Ni should be avoided as much as possible. PMID- 15269857 TI - [Specification and application of cephalometric analysis and image database]. AB - PURPOSE: To summarize the basic requirement of cephalometric analysis and database product concluded from practice, and apply related products to improve the levels of medical treatment, teaching and scientific research. METHODS: By the use of digital X ray machine, digital camera, oral endoscope,scanner to import the image into system database, to edit, measure, make diagram, analyze and store the images, to catalog diagnosis according to the different need of patient to make convenient for retrieving by group, to design multi-page electronic case report including oral general diagram, the result of diagnosis cataloging, the result diagram of analysis, picture and text report, all of the above mentioned constitute a complete patient database system. RESULTS: About 400 medical case were accumulated, and these improved evidently the effect to present the production of medical treatment to potential patients, were convenient to the professional communication between doctors and conformation of treatment plan, provided a solid foundation to multimedia teaching, and promoted the development of the task of graduate student and the task of scientific research. CONCLUSION: Using related product with perfect functions to develop standardized cephalometric analysis and build standardized patient's database including image information, when such information is accumulated to certain extent, will become the invaluable treasure of relevant medical institution, will bring the obvious effect to improve the level of medical treatment, teaching and scientific research. PMID- 15269858 TI - [A review of statistical analysis methods in measurement data]. AB - Enumeration data is consisted of categorical variable. It is obtained by counting the frequency of the inhomogeneous data. In clinical research, the collected data cover quite of categorical variables. Composed of those categorical variables, the enumeration data had to be correspondingly analyzed with special statistical methods on the ground of their design methods. The statistical methods of measurement data include descriptive statistics and inferential statistics. Descriptive statistics is mainly used to measure the relative number such as rate, proportion and ratio. Inferential statistics is mainly employed to estimate the confidence limit such as estimating the 95% confidence interval of rate and to have the hypothesis testing in the example of having chi-square test etc. By highlighting the specific statistical methods of enumeration data, this review intends to help the clinicians and researchers select correct statistical methods in accordance with enumeration data. PMID- 15269859 TI - [A review of the statistical methods used in the journal of prosthetic dentistry]. AB - PURPOSE: To understand the present condition of statistical methods used in the Journal of Prosthetic Dentistry. METHODS: The statistical methods used in 448 treatises (from 2001 to 2003) in the journals were reviewed, classified and analysed. RESULTS: The utilization rates of statistical methods were 28.06%, 48.34% and 51.02% from 2001 to 2003,respectively. The utilization rate of statistical methods was increasing significantly with year (chi2=29.453,P<0.001). The main statistical methods were analysis of variance (ANOVA), multiple comparison, nonparametric test, t-test, linear regression and correlation and chi2 test. CONCLUSION: The utilization rates of sophisticated statistical methods and the proportion of using more than two kinds of statistical tests were all higher than that in the medical journals published in China. PMID- 15269860 TI - [The basic characters and research advancements of odontoblasts]. AB - As the main components of dental pulp cells,the odontoblasts were responsible for the formation and maintenance of dentin during the development and mature age of teeth. Presently the studies were mostly based on the two-dimensional vitro cultured technique,but with the developing research of pulp tissue, three dimensional vitro cultured technique will be the study hotspot in future. The author retrospected the investigation about the odontoblasts these years and mostly expatiated the morphology,ultrastructure,function and characters of protein expression about the odontoblasts,including the research advancements and research aspects in future. PMID- 15269861 TI - [The effects of PL-derived growth factor on osteogenesis and skeletal reconstruction]. AB - Platelet-derived growth factor (PDGF) plays an important role in osteogenesis and skeletal reconstruction. PDGF participates in the process of osteogenesis and bone absorption simultaneously. It stimulates the marrow stromal mesenchymal cell to proliferate and differentiate, accelerates the calcium aggradation into extracellular matrix, and stimulates chondrocytes proliferation and differentiation. Furthermore, PDGF can act on osteoclasts directly or indirectly to promote bone absorption. The dual-effects of PDGF facilitate osteogenesis and skeletal reconstruction. PMID- 15269862 TI - [Management of major cervical vessel injury: clinical experience]. AB - PURPOSE: To review the diagnosis and management of injury of the major cervical vessels. METHODS: The clinical data of 12 cases with injury of the major cervical vessels were analyzed retrospectively. The causes of injury, emergency management, outcomes and complications were investigated. RESULTS: Among 12 cases, no death and no severe complications developed through appropriate management. CONCLUSIONS: For patients with cervical injury, the possibility of vascular injury should be considered. The key to salvage on spot is hemostasis by local compression with rapid filling. Complete exposure of the vessels is the key to success of surgical salvage. Anti-shock measures, endotracheal intubation general anesthesia and immediate operation are mandatory to improvement of treatment outcomes. PMID- 15269863 TI - [Oral-facial-digits syndrome I: case report]. AB - To report a case of oral-facial-digits syndrome I who underwent plastic surgery of tongue, palate and digital abnormalities. The tongue of the patient restored normal mobility approximately but the speech function could not recover completely. The function recovery of the patient depended on the degree of mental retardation, its possible mechanism was the mutation of OFD I gene. PMID- 15269864 TI - [Use of cortical (lag) screws in internal fixation of mental fractures]. AB - PURPOSE: To evaluate the clinical results of cortical (lag) screws in internal fixation of mental fractures. METHODS: 12 patients with single linear mental fracture underwent internal fixation using cortical screws. The patients were followed up for 6 to 9 months after fixation. RESULTS: Of 12 cases, 1 cortical screw was used in 10 cases, and 2 cortical screws were used in 2 cases. The wound healed primarily postoperatively. The patients had normal occlusion and mouth opening. CONCLUSION: Fixation of mental fracture using cortical screw can achieve good stability and appropriate compression. The technique is simple and easily performed, which avoids the bending of the reconstruction plate to adapt to the complex configuration of the mandible, reduce the surgical time, and promote the healing process by producing stress in the fracture lines. PMID- 15269865 TI - [Effect of ultrasonic in instrumentation on curved and obstructed root canal preparation]. AB - PURPOSE: To investigate and evaluate the effect of ultrasonic instrumentation on curved and obstructed root canal preparation. METHODS: Ultrasonic preparation was used on 117 root canals of 92 teeth due to resinifying therapy, pulp mummification and aging which could not be reopened and treated with traditional methods. RESULTS: 108 canals were successfully reopened with a successful rate of 92.3%. Failure occurred in 9 cases. 2 of them were lateral perforation. 2 were obstructed on curving position. 2 of them were due to pulp calcification in older patients. 3 were failing after resinifying therapy. CONCLUSION: The ultrasonic treatment of root canal was an effective and safe method. It was still limited in a cases with pulp calcification and some fine root canals treated by resinifying therapy. PMID- 15269866 TI - [Treatment of zygomaxillary complex multiple fractures: an analysis of 25 consecutive cases]. AB - PURPOSE: This retrospective study was performed to analyze the causes, 3D spiral CT examination and the treatment of ZMC fractures. METHODS: Rigid internal fixation was used in 25 patients with ZMC fractures after 3D spiral CT examination and anatomical reduction. The patients were followed up for 3 to 12 months. RESULTS: In the 25 cases, 13 cases had traffic accident injury, 5 had industrial accident injury. Local collapsed teratism were found in 21 cases,malocclusion was noted in 19 cases, limitation of mouth opening was found in 15 cases, diplopia was in 17 cases, infraorbital numbness in 11 cases, 5 cases had sagittal maxillary fracture. Spiral 3D CT images showed the steric anatomical images and relationship to the adjacent structures clearly in 25 patients. All patients were treated via rigid internal fixation with titanium plates and pins. 22 patients had satisfied maxillofacial appearance. CONCLUSION: Using topical, coronal and vestibular incisions accordingly, and rigid internal fixation with titanium plates, satisfied outcomes can be achieved in patients with ZMC fractures. PMID- 15269867 TI - [Effect of tinidazole-dexamethasone-iodoform paste on controlling endodontic interappointment emergencies]. AB - PURPOSE: To compare the clinical effect of tinidazole-dexamethasone-iodoform paste on intracanal dressing medication for teeth with chronic periapical periodontitis with that of formocresol. METHODS: 520 permanent teeth with chronic periapical periodontitis were selected and divided randomly into tinidazole dexamethasone-iodoform paste group (A group) and formocresol group (B group). The periapical signs and symptoms were recorded, radiographs were taken. After root canal preparation, the tinidazole-dexamethasone-iodoform paste was used as an intracanal dressing medication for one week in A group and the formocresol paper point was used in B group. During the course of canal treatment, the clinical findings were assessed with clinical periapical index (CPI), the cases were followed up for two years. RESULTS: There was significant difference between A and B group (P<0.01), group A was better than group B. There was no significant difference between two groups of the success rate after following up two years (P>0.05). CONCLUSION: As an intra-canal sterilization medication,tinidazole dexamethasone-iodoform can reduce the occurrence of endodontic interappointment emergencies in teeth with chronic periapical periodontitis and has a good long term result. PMID- 15269868 TI - [Analysis of preoperative misdiagnosis causes in 26 cases of neurolemmoma]. AB - PURPOSE: To improve the understanding of the diagnosis and treatment of neurolemmoma in maxillofacial region by summarizing its clinical features. METHODS: The preoperative misdiagnosis causes and treatment in 26 cases of neurolemmoma were reviewed. RESULTS: Among all cases treated surgically, 4 cases were pleomorphic adenoma of the parotid gland; 3 cases were malignant tumor of the parotid gland; 1 case was zygomatic osteoma; 4 cases were carotid body tumor; 1 case was branchial cleft cysts; 3 cases were cervical lymph node metastasis; 2 cases were pharyngeal malignant tumor; 4 cases were fibromatosis of the tongue; 2 cases were dermoid cyst; 2 cases were sublingual gland tumor. CONCLUSION: It is difficult to diagnose neurolemmoma before operation, however, it is important to carefully protect the function of the nerve in order to avoid severe complications. PMID- 15269869 TI - [Comparison of the results in 59 patients with mandibular fractures treated by internal rigid fixation and inter maxillary ligation]. AB - PURPOSE: The purpose of this study is to explore the clinical effect of the miniplate osteosynthesis on the treatment of mandibular fractures. METHODS: 59 patients with mandibular fractures were randomly divided into two groups. In the first group,29 patients underwent arch bar splints fixation and intermaxillary ligation. In the second group, miniplate osteosynthesis and intermaxillary ligation were performed in 30 patients. RESULTS: In the two groups,it was found that there was no significant difference in the postoperative infection and bone healing whereas the difference of occlusal relation,mouth opening and weight change was statistically significant. CONCLUSIONS: From this study,we conclude that miniplate osteosynthesis may be an appropriate and effective method for the treatment of mandibular fractures. As to rehabilitation of occlusal relation and mouth opening,it is much better than intermaxillary fixation. PMID- 15269870 TI - [Clinical using and observation of occlusion splint of RPD]. AB - PURPOSE: This article describes the design and restoration for 180 cases diagnosed with dentition defect accompanied with medium to severe abrasion and attrition. The purpose of this study is to discuss the rehabilitation of vertical dimension, the balance of occlusion and the whole occlusion system. METHODS: Before clinical preparation of the 180 clinical cases, diagnostic casts were made. Entire restoration treatment plan was adopted to make removable partial dentures with full crown restorations along with occlusion splints. The clinical effect were evaluated at 2nd week, 4th week and 9th week. RESULTS: Of the 180 cases, 148 patients (82.2%) adapted to the denture in 2-3 weeks, 24 patients (13.3%) in 4-5 weeks, 8 patients (14.5%) in 5-6 weeks. The further observation were evaluated over 10-15 years with clinical satisfaction. CONCLUSION: This method not only reserves the tooth structure, but also restores configuration and function of the lost tissues. It's really a constantly effective remedy. PMID- 15269871 TI - [Experience of repairing alveolar bone defects by collagen membrane and hydroxyapatite and its long-term result]. AB - PURPOSE: To observe the clinical effect of using the domestic BME-10X medical collagen membrane and hydroxyapatite (HA) in guide tissue regeneration of the bone defects caused by periodontitis and periapical cyst. METHODS: 18 cases with 9 points in 9 teeth with II degree furation involvement, 16 points in 11 teeth with 2 or 3 bone bottom pockets and 6 cases with periapical cyst were chosen in our study. The defect region of alveolar bone was stuffed with HA and covered with collagen membrane. The pathological changes after over one year were observed and recorded. RESULTS: The pathological changes of alveolar bone defects were significant in 7 cases with furcation involvements (77.8%),12 cases with bone bottom pockets (75%),and 6 cases of periapical cyst (100%). CONCLUSION: The combination of collagen membrane and HA can be applied in repairing the alveolar bone defects resulting form periodontitis and periapical cysts. PMID- 15269872 TI - [Treatment results of 216 cracked teeth: clinical analysis]. AB - PURPOSE: To evaluate the treatment results of 216 cracked teeth. METHODS: Between May 1991 and May 2003, 208 patients with 216 cracked teeth were treated according to the stages. The patients were followed up and the final outcomes were analyzed, retrospectively. RESULTS: The majority of the patients were between 40 and 60 years, with a predominance in males. The cracked lines were mainly located in the mesial grooves. The dominant treatment modality was binding of band, adjustment of occlusion, endodontic therapy and restoration with artificial crowns, with a success rate of 92%. CONCLUSION: Cracked teeth are commonly seen clinically with appropriate treatment, these teeth can be still useful. Routine extraction of these teeth are not advocated. PMID- 15269873 TI - [The effects of non-surgical periodontal treatment on the formation of root dentin hypersensitivity]. AB - PURPOSE: To study the effects of non-surgical periodontal treatment on the occurrence of root-dentin hypersensitivity and explore its mechanisms through clinical observations. METHODS: 1453 teeth of 52 patients with chronic periodontitis were given scaling and root-planing procedures. 3 months follow-up of these teeth were taken to find root-dentin hypersensitivity after treatment. RESULTS: 432 teeth of 39 patients had different degree of root-dentin hypersensitivity. However,the symptoms disappeared in most patients after active desensitization treatment. CONCLUSION: Subgingival scaling and root-planing can induce the occurrence of root dentin hypersensitivity,therefore, desensitization treatment should be considered as a part of periodontal treatment plan. PMID- 15269875 TI - [Salvage operations and their differential indication for the distal radioulnar joint]. AB - The most common cause of an arthritically damaged distal radioulnar joint is a malunion of a distal radius fracture. Therapeutically, ulnar head resection, hemiresection-interposition-technique, Kapandji-Sauve procedure and implantation of an ulnar head prosthesis have been described. None of these procedures is able to restore the complete function of the joint. Therefore, anatomical reconstruction of the joint in acute or secondary correction osteotomy for malunited fractures of the distal radius should be performed to avoid the development of the arthrosis. Numerous clinical studies have demonstrated a similar reduction of the clinical symptoms for all procedures. Therefore, classification of the different procedures has to consider the number of complications. Biomechanically, partial resection of the distal ulna will destabilize the distal radioulnar context and clinically may lead to painful radioulnar and/or dorsopalmar instability of the distal ulnar stump. Biomechanically and clinically, this complication, next to secondary extensor tendon ruptures, has to be expected far more often following complete resection of the ulnar head than in the alternative procedures. We do not see any remaining indication for complete resection of the ulnar head. Clinical results and the occurrence of painful instability of the distal ulnar stump have been reported almost identically for the hemiresection-interposition technique and the Kapandji Sauve procedure. Therefore, both procedures appear to be equally suitable for the treatment of painful arthrosis of the distal radioulnar joint. In patients with a preexisting instability of the distal radioulnar joint, or a major deformity of the radius or the ulna, we prefer to perform the hemiresection-interposition technique. In these conditions we consider the remaining contact of the triangular fibrocartilage complex with the distal end of the ulna a biomechanical advantage to reduce the risk of secondary instability. Biomechanically as well as clinically, replacement of the ulnar head using a prosthesis has been shown to either avoid or solve the problem of instability. We therefore consider ulnar head replacement the treatment of choice in secondary painful instability following resection procedures at the distal end of the ulna. Primary ulnar head replacement should be considered in special indications until long-term follow-up results are available. PMID- 15269876 TI - [Factors influencing perioperative morbidity and mortality in primary hip arthroplasty]. AB - INTRODUCTION: In the present study we examined preoperative parameters that may identify patients at high risk for postoperative complications after endoprosthetic joint replacement. MATERIALS AND METHODS: The incidence of risk factors and perioperative complications in 628 primary hip arthroplasties (THA) (549 patients) was investigated in an unselected, retrospective study. Concomitant illnesses were found in 426 cases. Intra- and postoperative complications (93 specifically orthopedic and 42 common ones) were observed in 104 cases. RESULTS: High risk scores based on Lutz and Klose criteria, a prolonged operation time, and the number of previous operations were significantly correlated to the incidence of postoperative complications. In contrast, obese patients had a significantly lower rate of intra- and postoperative complications and a diminished perioperative blood loss. THAs performed under intubation anesthesia led to a higher blood transfusion volume. The patient's age and the kind and quantity of concomitant illnesses did not influence the perioperative complication rate. CONCLUSION: The complication rate of elective primary THAs is not dependent on risk factors suspected up to now such as advanced patient age or the kind and quantity of concomitant illnesses. High-risk patients can only be determined by complex scores, not by single parameters. Adiposity becomes a relevant economic factor only by dint of the prolonged operation time. PMID- 15269877 TI - Exciting new developments in chronic lymphocytic leukemia: an introduction and an editorial. PMID- 15269878 TI - Patient management decisions in chronic lymphocytic leukemia: an introduction and an editorial. PMID- 15269879 TI - Clonal lymphocytes in persons without known chronic lymphocytic leukemia (CLL): implications of recent findings in family members of CLL patients. AB - Several genetic abnormalities have been characterized in chronic lymphocytic leukemia (CLL) but these are predominantly secondary events and the initiating phenomena in the etiology of the disease are yet to be established. Studies of inherited susceptibility have identified the early oncogenic events in both familial and "sporadic" forms of several malignant disorders, and this may also be possible in CLL. However, the utility of linkage analysis in identifying a predisposition locus for the disease is limited because large multigenerational families segregating CLL are rare, while the more frequent small nuclear CLL families contain insufficient numbers of affected individuals. The power to detect predisposition gene(s) could be greatly increased by extending the number of affected individuals within a particular family, for example, by identifying family members with subclinical levels of disease. High-sensitivity flow cytometry techniques, developed to monitor disease in CLL patients undergoing treatment, have allowed accurate enumeration of subclinical levels of CLL cells in healthy individuals from the general population and CLL families. Emerging evidence confirms the phenotypic, genotypic, and clinical associations between the aberrant cells in healthy individuals and those in CLL patients. The data suggest that inherited factors increase the susceptibility to both indolent and aggressive CLL, and they provide unbiased demonstration that the age of onset in CLL families is younger than in the general population. PMID- 15269880 TI - Perspectives on familial chronic lymphocytic leukemia: genes and the environment. AB - Chronic lymphocytic leukemia (CLL) comprises a substantial proportion of leukemias in adults in the western hemisphere. Male gender, increasing age, ethnicity (high in Caucasians, lowest in Asians), and family history are risk factors. Although no specific extrinsic etiologic factors have been established, farming and pesticide exposure are associated with increased risk. Migration studies confirm that ethnic groups retain the risk associated with their origin rather than their new location, favoring a role for heredity. Kindreds with multiple cases of CLL have been well described in the literature and studies in large populations confirm that lymphoproliferative malignancies and especially CLL occur together at a rate that cannot be attributed to chance. Since environmental factors cannot readily explain the familial aggregations, a hereditary factor that affects susceptibility to CLL is likely. The identification of clones that are immunophenotypically identical to CLL in healthy individuals from CLL kindreds (14% to 18%) as well as in the general population (3.5% in age bracket >65 years) suggests a possible precursor condition, but longitudinal studies will be necessary to establish significance in the general population. Family (linkage) and population (candidate gene) studies to date have been too small to identify the specific genes that account for increased susceptibility; larger studies including planned consortia to identify additional high-risk kindreds for genetic studies, as well as the application of advanced technologies such as genomics, cytogenetic, expression, and proteomics, are widely expected to advance understanding over the next few years. PMID- 15269881 TI - Recent advances in the molecular biology and immunobiology of chronic lymphocytic leukemia. AB - B-cell chronic lymphocytic leukemia (B-CLL) has long been viewed as a relatively homogeneous disease caused by the accumulation of monoclonal immature, immunoincompetent B cells with faulty apoptotic capacities. However, recent evidence, reviewed here, demonstrates that at least two different B-CLL subgroups exist with different clinical courses and outcomes. The malignant cells from both B-CLL subgroups are antigen-experienced cells that have a normal apoptotic apparatus and turnover continually. The leukemic cells of the two B-CLL subgroups have engaged antigen before transformation, although primarily the cells of patients in the poor outcome subgroup can respond to antigens following transformation. The difference in the ability to respond to antigen as a full fledged B-CLL probably accounts for the different biological features and clinical outcomes of the patients in these subgroups. PMID- 15269882 TI - Advances in chemotherapy for chronic lymphocytic leukemia. AB - While chemotherapy based on alkylating agents has been the standard treatment of chronic lymphocytic leukemia (CLL) for decades, purine analogues and their combinations have emerged as effective new therapies for previously untreated and pretreated patients. As single agents, fludarabine and cladribine are the most promising, showing higher remission rates compared to chlorambucil. For younger and physically fit patients, the combination of fludarabine and cyclophosphamide has shown benefit. Fludarabine plus epirubicin appears equally potent. The addition of monoclonal antibodies, such as rituximab and alemtuzumab, to purine analogues alone or in combination seems to be even more effective for chemotherapy-naive and pretreated CLL patients. Another promising agent in the armamentarium of therapies for CLL is bendamustine, which has properties of both an alkylating agent and a purine analogue. Clinical trials are ongoing with novel drugs that interfere with cell cycle regulation and signaling molecules in CLL, including flavopiridol, UCN-01, bryostatin 1, depsipeptide, and oblimersen. It remains to be seen whether these chemotherapeutic approaches offer real benefit for patients by prolonging survival with an improved quality of life. PMID- 15269883 TI - Advances in the use of monoclonal antibodies in the therapy of chronic lymphocytic leukemia. AB - Monoclonal antibody (moAb)-based therapies are evolving as an integrated component in the treatment of chronic lymphocytic leukemia (CLL). Advantages such as different mechanisms of action (compared with those of chemotherapy), no or minimal stem cell toxicity, as well as the absence of hair loss and delayed nausea may result in a rapidly increasing usage of these agents in different phases of the disease. The combination of moAbs with chemotheraputic agents has shown promising results in early studies as well as their role in the eradication of minimal residual disease (MRD). The availability of an increasing number of new moAbs together with a better understanding of their effector function will hopefully lead to improved therapeutic outcomes for patients with CLL and related disorders. PMID- 15269884 TI - The role of stem cell transplantation in chronic lymphocytic leukemia. AB - Chronic lymphocytic leukemia (CLL) generally follows an indolent clinical course and usually occurs in the elderly. However, the disease is heterogeneous with some patients having a more aggressive clinical course and short survival. Although the role of fludarabine in combination with other chemotherapy drugs and/or monoclonal antibody therapy appears promising, to date chemotherapy has not been curative in this disease. At present, the only potential cure for CLL appears to be stem cell transplantation (SCT), but its role in the management of CLL has not been established. In particular, patient selection for consideration of SCT, timing of SCT in the clinical course of CLL, selection of autologous versus allogeneic SCT, use of nonmyeloablative regimens, and exploitation of the graft-versus-leukemia (GVL) effect are currently under investigation. PMID- 15269886 TI - Molecular mechanisms of neural crest induction. AB - The neural crest is an embryonic cell population that originates at the border between the neural plate and the prospective epidermis. Around the time of neural tube closure, neural crest cells emigrate from the neural tube, migrate along defined paths in the embryo and differentiate into a wealth of derivatives. Most of the craniofacial skeleton, the peripheral nervous system, and the pigment cells of the body originate from neural crest cells. This cell type has important clinical relevance, since many of the most common craniofacial birth defects are a consequence of abnormal neural crest development. Whereas the migration and differentiation of the neural crest have been extensively studied, we are just beginning to understand how this tissue originates. The formation of the neural crest has been described as a classic example of embryonic induction, in which specific tissue interactions and the concerted action of signaling pathways converge to induce a multipotent population of neural crest precursor cells. In this review, we summarize the current status of knowledge on neural crest induction. We place particular emphasis on the signaling molecules and tissue interactions involved, and the relationship between neural crest induction, the formation of the neural plate and neural plate border, and the genes that are upregulated as a consequence of the inductive events. PMID- 15269887 TI - A slug, a fox, a pair of sox: transcriptional responses to neural crest inducing signals. AB - The neural crest, a cell type found only in vertebrate embryos, gives rise to the structures of the skull and face and most of the peripheral nervous system, as well as other cell types characteristic of vertebrates. These cells are of great clinical significance and a wide variety of congenital defects are due to aberrant neural crest development. Increasing numbers of studies are contributing to our understanding of how this group of cells form and differentiate during normal development. Wnt, FGF, BMP, and Notch-mediated signals all have essential roles in this process, and several of these signals appear to play multiple temporally distinct roles. Changes in the response of neural crest cells to the same signal over time may be mediated, in part, by an ever-changing cocktail of transcription factors expressed within these cells. Neural crest development is thus a complex multistep process, and elucidating the molecular mechanisms that mediate distinct aspects of this process will require that we determine the role of each of these factors alone and in combination. Here, we review some recent advances in our understanding of the signals and downstream transcription factors involved in neural crest cell formation. PMID- 15269888 TI - Cell surface molecules and truncal neural crest ontogeny: a perspective. AB - The neural crest cell is synonymous with vertebrates and can be viewed as a transitory, mobile vector that conveys neuroepithelial stem cells to a diverse number of remote locations in the embryo. Neural crest cells have been studied intensively over the past 30 years, and it is increasingly apparent that their fate is, at least in part, directed extrinsically by the environment to which they are exposed in vivo. The interface between the cell surface and the opposing environment is clearly an important compartment for the correct deployment of the neural crest. Here, we review some of the molecules present in this location and how they influence the fate of the neural crest and generate disease. PMID- 15269890 TI - The adult hair follicle: cradle for pluripotent neural crest stem cells. AB - This review focuses on the recent identification of two novel neural crest derived cells in the adult mammalian hair follicle, pluripotent stem cells, and Merkel cells. Wnt1-cre/R26R compound transgenic mice, which in the periphery express beta-galactosidase in a neural crest-specific manner, were used to trace neural crest cells. Neural crest cells invade the facial epidermis as early as embryonic day 9.5. Neural crest-derived cells are present along the entire extent of the whisker follicle. This includes the bulge area, an epidermal niche for keratinocyte stem cells, as well as the matrix at the base of the hair follicle. We have determined by in vitro clonal analysis that the bulge area of the adult whisker follicle contains pluripotent neural crest stem cells. In culture, beta galactosidase-positive cells emigrate from bulge explants, identifying them as neural crest-derived cells. When these cells are resuspended and grown in clonal culture, they give rise to colonies that contain multiple differentiated cell types, including neurons, Schwann cells, smooth muscle cells, pigment cells, chondrocytes, and possibly other types of cells. This result provides evidence for the pluripotentiality of the clone-forming cell. Serial cloning showed that bulge-derived neural crest cells undergo self-renewal, which identifies them as stem cells. Pluripotent neural crest cells are also localized in the back skin hair of adult mice. The bulge area of the whisker follicle is surrounded by numerous Merkel cells, which together with innervating nerve endings form slowly adapting mechanoreceptors that transduce steady skin indentation. Merkel cells express beta-galactosidase in double transgenic mice, which confirms their neural crest origin. Taken together, our data indicate that the epidermis of the adult hair follicle contains pluripotent neural crest stem cells, termed epidermal neural crest stem cells (eNCSCs), and one newly identified neural crest derivative, the Merkel cell. The intrinsic high degree of plasticity of eNCSCs and the fact that they are easily accessible in the skin make them attractive candidates for diverse autologous cell therapy strategies. PMID- 15269889 TI - Combinatorial transcriptional interaction within the cardiac neural crest: a pair of HANDs in heart formation. AB - The cardiac neural crest cells migrate from the rostral dorsal neural folds and populate the branchial arches, which contribute directly to the cardiac-outflow structures. Although neural crest cell specification is associated with a number of morphogenic factors, little is understood about the mechanisms by which transcription factors actually implement the transcriptional programs that dictate cell migration and later the differentiation into the proper cell types within the great vessels and the heart. It is clear from genetic evidence that members of the paired box family and basic helix-loop-helix (bHLH) transcription factors from the twist family of proteins are expressed in and play an important function in cardiac neural crest specification and differentiation. Interestingly, both paired box and bHLH factors can function as dimers and, in the case of twist family bHLH factors, partner choice can clearly dictate a change in transcriptional program. The focus of this review is to consider what role the protein-protein interactions of these transcription factors may play in determining cardiac neural crest specification and differentiation, and how genetic alteration of transcription factor stoichiometry within the cell may reflect more than a simple null event. PMID- 15269891 TI - The neural crest: basic biology and clinical relationships in the craniofacial and enteric nervous systems. AB - The highly migratory, mesenchymal neural crest cell population was discovered over 100 years ago. Proposals of these cells' origin within the neuroepithelium, and of the tissues they gave rise to, initiated decades-long heated debates, since these proposals challenged the powerful germ-layer theory. Having survived this storm, the neural crest is now regarded as a pluripotent stem cell population that makes vital contributions to an astounding array of both neural and non-neural organ systems. The earliest model systems for studying the neural crest were amphibian, and these pioneering contributions have been ably refined and extended by studies in the chick, mouse, and more recently the fish to provide detailed understanding of the cellular and molecular mechanisms regulating and regulated by the neural crest. The key questions regarding control of craniofacial morphogenesis and innervation of the gut illustrate the wide range of developmental contexts in which the neural crest plays an important role. These questions also focus attention on common issues such as the role of growth factor signaling in neural crest cell development and highlight the central role of the neural crest in human congenital disease. PMID- 15269892 TI - Understanding endothelin-1 function during craniofacial development in the mouse and zebrafish. AB - Morphogenesis of the face and neck is driven by an intricate relay of signaling molecules and transcription factors organized into hierarchical pathways. The coordinated action of these pathways regulates the development of neural crest cells within the pharyngeal arches, resulting in proper spatiotemporal formation of bone, cartilage, and connective tissue. While the functions of many genes involved in these processes were initially elucidated through the use of knockout technology in the mouse, increasing numbers of zebrafish craniofacial mutants have led to a rapid expansion in the identification of genes involved in craniofacial development. A comparative analysis of signaling pathways involved in these processes between mouse and zebrafish holds the potential not only to pinpoint conserved and therefore crucial gene functions in craniofacial development, but also to rapidly identify and study downstream effectors. These complementary approaches will also allow rapid identification of candidate genes and gene functions disrupted in human craniofacial dysmorphologies. In this brief review, we present a comparative analysis of one molecule involved in craniofacial development, endothelin-1, a small, secreted protein that is crucial for patterning the neural crest cells that give rise to lower jaw and throat structures. PMID- 15269893 TI - Neural crest contribution to mammalian tooth formation. AB - The cranial neural crest cells, which are specialized cells of neural origin, are central to the process of mammalian tooth development. They are the only source of mesenchyme able to sustain tooth development, and give rise not only to most of the dental tissues, but also to the periodontium, the surrounding tissues that hold teeth in position. Tooth organogenesis is regulated by a series of interactions between cranial neural crest cells and the oral epithelium. In the development of a tooth, the epithelium covering the inside of the developing oral cavity provides the first instructive signals. Signaling molecules secreted by the oral epithelium 1) establish large cellular fields competent to form a specific tooth shape (mono- or multicuspid) along a proximodistal axis; 2) define an oral (capable of forming teeth) and non-oral mesenchyme along a rostrocaudal axis; and 3) position the sites of future tooth development. The critical information to model tooth shape resides later in the neural crest-derived mesenchyme. Cranial neural crest cells ultimately differentiate into highly specialized cell types to produce mature dental organs. Some cranial neural crest cells located in the dental pulp, however, maintain plasticity in their developmental potential up to postnatal life, offering new prospects for regeneration of dental tissues. PMID- 15269895 TI - Toxicity assessment of oil-contaminated freshwater sediments. AB - The performance of four microscale toxicity bioassays conducted on whole sediments was evaluated during a bioremediation project undertaken in 1999-2000 on a crude oil-contaminated freshwater shoreline of the St. Lawrence River, Quebec, Canada. The toxicity tests assessed included: (1) the Microtox solid phase assay (MSPT), (2) the Biotox Flash solid-phase test (Flash), (3) the algal solid-phase assay (ASPA), and 4) the Ostracodtoxkit solid-phase assay. Data generated with these assays were compared with those obtained using the standard endobenthic amphipod (Hyalella azteca) bioassay. Bioanalytical comparisons indicated that all five solid-phase tests were useful in detecting the toxicity of oiled sediments; however, statistical analyses distinguished a difference in response between the invertebrate (amphipod and Ostracodtoxkit) and bacterial luminescence tests (MSPT and Flash). Based on these results, it is recommended that careful selection of biotests be made in the design of the test battery for assessment of residual oil sediment toxicity. Time-series toxicity data generated with ASPA indicated that oiled sediments in the freshwater wetlands of the St. Lawrence River remained toxic to phytoplankton for at least 65 weeks and that remediation treatment was able to accelerate detoxification by 16 weeks. PMID- 15269896 TI - Application of Toxkit microbiotests for toxicity assessment in soil and compost. AB - The potential of Toxkit microbiotests to detect and analyze pollution in agricultural soil and the quality of compost was studied. The toxicity tests used included seed germination biotests using cress salad (Lepidum sativum L.), tomato (Lycopersicum esculentum L.), and cucumber (Cucumis sativus L.), and the Toxkit microbiotests included those with microalgae (Selenastrum capricornutum), protozoa (Tetrahymena thermophila), crustaceans (Daphnia magna, Thamnocephalus platyurus, and Heterocypris incongruens), and rotifers (Brachionus calyciflorus). Experiments on compost were undertaken in a modified solid-state fermentation system (SSF) and under field conditions (in a windrow). To promote the composting process, two strains of Trichoderma (Trichoderma lignorum and Trichoderma viride), as well as a nitrification association that regulated the nitrogen ammonification and nitrification processes were applied. PMID- 15269897 TI - Were volatile organic compounds the inducing factors for subjective symptoms of employees working in newly constructed hospitals? AB - This study demonstrated possible relationships between environmental, personal, and occupational factors and changes in the subjective health symptoms of 214 employees after the relocation of a hospital in a region of Japan. Eight indoor volatile organic compounds (VOCs) were detected in at least one of the 19 rooms investigated, and total VOC (TVOC) concentrations in 8 rooms exceeded the advisable value (400 microg/m(3)) established by the Ministry of Health, Labour and Welfare of Japan. Formaldehyde was detected in all the investigated rooms, but none of the results exceeded the guideline value (100 microg/m(3)). Multiple logistic regression analysis was applied to select variables significantly associated with the subjective symptoms that can be induced by sick building syndrome. The results showed that subjective symptoms of deterioration in the skin, eye, ear, throat, chest, central nervous system, autonomic system, musculoskeletal system, and digestive system among employees were associated mainly with gender difference and high TVOC concentrations (>1200 microg/m(3)). Long work hours (>50 h per week) in females and smoking in males were to be blamed for the deterioration of their symptoms. The present findings suggest that to protect employees from indoor environment-related adverse health effects, it is necessary to reduce the concentration of indoor chemicals in new buildings, to decrease work hours, and to forbid smoking. PMID- 15269898 TI - Toxicity of pesticides to predatory mites and insects in apple-tree site under field conditions. AB - Various applications of active ingredients of six fungicides and three insecticides and acaricides at normally recommended dosages were tested on two predatory mite species (Amblyseius andersoni Chant and Anthoseius bakeri Garman) from the family Phytoseiidae and on two predatory insect species (Coccinella septempunctata L. Chrysopa perla L.) dominantly present on apple trees. Small differences were found between fungicide treatments. On the trees treated with six fungicide applications the predatory mites and insects survived and increased to a high level, often 20-40 phytoseiids per 100 leaves and 4-8 predatory insects per sample unit. Only the active ingredients tolylfluanid and myclobutanil resulted in lower densities of predatory mites (10-20 phytoseiids per 100 leaves). One application of insecticides-acaricides (active ingredients: clofentezine, phosalone) showed no toxic effect on predatory mites and insects. Two applications of phosalone and one of alpha-cypermethrine were slightly or moderately toxic. Two applications of alpha-cypermethrine and eight routine sprays of various insecticides-acaricides and fungicides were very toxic and resulted in the lowest maximum number of predatory mites and insects, approximately 0-10 phytoseiids per 100 leaves and 1-4 predatory insects per sample unit. The toxicity of pesticides to predatory mites and insects is based on the toxicity of the pesticide' active ingredient and the spray frequency. The active ingredients of fungicides and only one or two applications of insecticides and acaricides were not or slightly toxic and could be used in integrated pest management. PMID- 15269899 TI - Effect of deicing salts on urban soils and health status of roadside trees in the Opole region. AB - This article reports on a study whose aim was to evaluate the impact of snow removal salts on urban soil properties and the health of roadside trees. The evaluation was done by chemical analyses of soil samples and plant matter combined with toxicity testing, performed with a Protoxkit F, a protozoan microbiotest. Samples were collected at 45 locations on three main roads in the town of Opole (Poland). The roads differed in the snow removal technology and amount of chemical substances (mostly NaCl) used on them during the winter. The study showed that when soil was exposed to a high level of NaCl, it tended to be more alkaline and also exhibited increased content of Na(+) and Cl(-). The toxic effects of the soil extract on protozoa appeared at 26.0 mg Na(+)/100 g soil dry mass (s.d.m.) and 12.0 mg Cl(-)/100 g s.d.m., whereas salt injury symptoms (chlorosis and necrosis of the edge of leaf blades) appeared at 13.2 mg Na(+)/100 g s.d.m. and 3.9 mg Cl(-)/100 g s.d.m., becoming more severe at 26.0 mg Na(+)/100 g s.d.m. and 12.0 mg Cl(-)/100 g s.d.m. because of extensive necrosis and defoliation. The lysimetric experiment, which was used to test soil samples collected from the city park area, indicated that salt plays a significant role in the pollution of soil in urban areas, with the least toxic salt being CaCl(2). PMID- 15269900 TI - Toxicity assessment of wastewaters, river waters, and sediments in Austria using cost-effective microbiotests. AB - The toxicity and chemical quality of surface water and sediment in the River Traun in Austria were studied because of recurrent fish mortality in some alpine rivers over the last few years. The analyses were carried out on samples collected during winter and summer upstream and downstream of two municipal wastewater treatment plants (WWTPs) and on effluents taken at the points of discharge of these two plants. Toxicity tests were performed on 20 samples of surface water, effluent, and sediment pore water. The test battery was composed of microbiotests with protozoans (Protoxkit F), microalgae (Algaltoxkit F), crustaceans (Daphtoxkit F magna and Thamnotoxkit F), and a higher plant (seed germination and root elongation assay on cress). Direct contact tests were performed on whole sediment with crustaceans (Ostracodtoxkit F). The physical chemical characteristics of the surface water, effluent, and sediment pore water samples analyzed were conductivity, total hardness, pH, O(2), BOD(5), TOC, DOC, AOX, NH(4), NH(3), NO(2), PO(4)--P, Cd, Pb, Cu, and Zn. The toxicity data were expressed as percentage mortality or percentage inhibition, depending on the effect criterion of the respective assay. None of the water samples collected upstream and downstream of the WWTPs showed any significant (short-term) toxicity in either winter or in summer, but the effluents of the first municipal wastewater treatment plant were toxic to some of the test biota. All the sediment pore water samples induced serious inhibition of root growth of cress, and several pore waters were toxic to other test biota as well, particularly at the outlets of the WWTPs. The toxic character of some sediments was confirmed by direct contact tests with the ostracod crustacean. The chemical analyses did not reveal particularly high concentrations of any chemical that is very toxic. As a result no direct causal relationship could be established between the detected toxic effects and the chemical composition of the surface waters or sediment pore waters. The outcome of this preliminary study again highlights the need to complement chemical analyses with toxicity tests to determine the toxic hazard to aquatic environments that may be threatened by contamination. Furthermore, the investigations also confirmed the need to apply a battery of tests for an ecologically meaningful evaluation of the hazards of waters, sediments, and wastewaters. Finally, the results of the 360 bioassays performed show that culture-independent microbiotests are practical and reliable tools for low-cost toxicity monitoring of aquatic environments. PMID- 15269901 TI - Comparison of 17 biotests for detection of cyanobacterial toxicity. AB - The aim of the present study was to compare the sensitivity of 17 acute bioassays of cyanobacterial toxicity by assessment of crude extracts of three cyanobacterial samples (all dominated by Microcystis sp. but substantially differing in microcystin-LR content). Toxicity of the fractions prepared by solid phase extraction (SPE) for microcystins was also determined. The most sensitive bioassay was the 24-h test with crustacean Thamnocephalus platyurus, which elicited high lethality in the samples and also in fractions without microcystins. The fruit fly Drosophila melanogaster, protozoans Spirostomum ambiguum and Tetrahymena termophyla, and the crustacean Daphnia pulex formed the second group of sensitive bioassays. Good selective toxicity response to microcystins also was observed in the weakly sensitive biotests with the oligochaete Tubifex tubifex and the rotifer Brachionus calyciflorus. Preconcentration of microcystins by SPE substantially decreased variation of the results in bioassays and improved the discriminating potential of most assays employed. PMID- 15269902 TI - Physical and chemical aspects of long-term biodeterioration of some polymers and composites. AB - A biodeterioration study was performed on synthetic polymeric materials including homogenous film made from poly(tetrafluorine ethylene), copolymer film made from tetrafluorine ethylene and perfluoromethyl vinyl ether, vulcanized rubber containing natural caoutchouc, and vulcanized rubber, the main component of which was synthetic butadiene nitrile caoutchouc. The materials were exposed for 12 years to the open air, in mycological containers, and in a cellar in maritime climate conditions: air humidity 72%-90% and seasonal average temperature of 17 degrees C in summer and -2.5 degrees C in winter. The studies of optical and electron microscopy revealed that microorganisms were able to develop not only on the surface of the materials but also to penetrate inside into deeper layers. The fungi that produced the most intensive deterioration in the fluorine polymers and vulcanized rubbers belonged to the Alternaria, Aspergillus, Aureobasidium, Cladosporium, Penicillium, Oidiodendron and Trichoderma genera. The fungi Aspergillus fumigatus, A. niger, Aureobasidium pullulans, and Trichoderma viride produced the most intensive deterioration in the fluorine films, whereas Alternaria tenuissima, Cladosporium herbarum, C. sphaerospermum, and fungi of the Oidiodendron genus were widespread on vulcanized rubbers. Fungi of the Aspergillus and Penicillium genera prevailed on both fluorine films and rubbers exposed in a cellar. Infrared spectroscopy indicated that the structures of poly(tetrafluorine ethylene) and the copolymer of tetrafluorine ethylene and perfluoromethyl vinyl ether did not change after the 12-year exposure; only insignificant changes in surface morphology were observed by optical microscopy. Vulcanized rubber made both from natural and from synthetic caoutchouc exposed for the same length of time showed rather evident changes in appearance and structure. X-ray graphical analysis revealed that new crystallization of the caoutchouc and a possible change in chemical composition of the fillers had occurred. PMID- 15269903 TI - Hazard assessment of a simulated oil spill on intertidal areas of the St. Lawrence River with SPMD-TOX. AB - Phytoremediation in a simulated crude oil spill was studied with a "minimalistic" approach. The SPMD-TOX paradigm-a miniature passive sorptive device to collect and concentrate chemicals and microscale tests to detect toxicity-was used to monitor over time the bioavailability and potential toxicity of an oil spill. A simulated crude oil spill was initiated on an intertidal freshwater grass-wetland along the St. Lawrence River southwest of Quebec City, Quebec, Canada. Several phytoremediation treatments were investigated; to dissipate and ameliorate the spill, treatments included nutrient amendments with inorganic nitrogen sources (ammonium nitrate and sodium nitrate) and phosphate (super triple phosphate) with and without cut plants, with natural attenuation (no phytoremedial treatment) as a control. Sequestered oil residues were bioavailable in all oil-treated plots in Weeks 1 and 2. Interestingly, the samples were colored and fluoresced under ultraviolet light. In addition, microscale tests showed that sequestered residues were acutely toxic and genotoxic, as well as that they induced hepatic P(450) enzymes. Analysis of these data suggested that polycyclic aromatic hydrocarbons were among the bioavailable residues sequestered. In addition, these findings suggested that the toxic bioavailable fractions of the oil spill and degradation products dissipated rapidly over time because after the second week the water column contained no oil or detectable degradation products in this riverine intertidal wetland. SPMD-TOX revealed no evidence of bioavailable oil products in Weeks 4, 6, 8, and 12. All phytoremediation efforts appeared to be ineffective in changing either the dissipation rate or the ability to ameliorate the oil toxicity. SPMD-TOX analysis of the water columns from these riverine experimental plots profiled the occurrence, dissipation, and influence of phytoremediation on the bioavailability and toxicity of oil products (parent or degradation products). PMID- 15269904 TI - The medicinal leech as a convenient tool for water toxicity assessment. AB - Medicinal leeches previously were used in various toxicological and pharmacological studies because they are sensitive and easy to keep under laboratory conditions. Toxicological studies using leeches became restricted when their natural sources decreased dramatically. We breed medicinal leeches under laboratory conditions and have the possibility of using them for various investigations. The aim of the current study was to investigate changes in behavioral and physiological responses of leeches exposed to a heavy metal model mixture (HMMM). The composition of the HMMM was determined on the basis of the average annual amounts of representative metals (Zn, Mn, Cu, Cr, Ni, Cd, and Pb) in wastewater discharging from the Ignalina Nuclear Power Plant. An annual average was considered a concentration of 1%. Animals were exposed to 0.13%, 0.25%, 0.5%, 1%, and 2% concentrations of the HMMM. The avoidance responses, changes in body weight, feeding behavior, and excretion rates of the leeches were recorded. Avoidance response was observed in animals treated with 1% and 2% concentrations of the HMMM during the first 2 h of exposure. A decrease in body weight was recorded after a 4-week exposure to all tested concentrations. Disturbances in feeding behavior were observed after a 4-week exposure to 0.13% 2% concentrations. An increase in the defecation rate was observed during the 5 week exposure to 0.25%-2% concentrations. The excretion rate via nephridia was decreased during postfeeding period after exposure to 0.25%-2% concentrations. The avoidance response of medicinal leeches can be used as an express method for water quality assessment, whereas physiological responses may be used for the assessment of chronic toxicity of polluted environments. PMID- 15269905 TI - Development of a biosensor for the detection of tributyltin. AB - A biosensor (LUMISENS I), based on the inducible bioluminescence of the Escherichia coli strain TBT3 (Ec::luxAB TBT3), was developed for the detection of the biocide tributyltin. LUMISENS I was set up with a minibioreactor and additional equipment for growth monitoring and light acquisition. The 100-mL minibioreactor has allowed us to establish a stable and reproducible environment for the bacteria (regulation of the growth rate, temperature, pH, and oxygenation), as well as for in situ contact with the xenobiotic. The optical components of the transducer were chosen according to the spectral emission of the strain being studied using a highly sensitive spectrophotometer that was initially devoted to Raman scattering. LUMISENS I was patented according to the in situ, automatic, and simultaneous measurement of the cell density and bioluminescence in the bioreactor. The first results showed that cells cultivated in a synthetic glucose medium provided a better detection limit than did those cultivated in a complex Luria-Bertani (LB) medium (0.02 and 1.5 microM of tributyltin, respectively). Cells maintained at a high growth rate (0.9 h(-1)) led to maximum bioluminescence. Moreover, air bubbling was efficient enough to provide suitable quantities of oxygen for both growth and light emission. When the TBT3 strain used the luxAB genes on its own, decanal, a long-chain aldehyde, had to be added to obtain the bioluminescence reaction. We found that the continuous addition of decanal was the most effective means of obtaining this reaction. The monitoring of the bioluminescence after tributyltin induction showed that the aldehyde was not toxic up to 300 microM during a 7-day experiment. Measurement of tributyltin with LUMISENS I was performed, which showed significant response up to 0.125 microM without any effect on optical density. Even though optimization of the performance of LUMISENS I is still under development, because of its original design, this biosensor is already in use as a warning system for the online monitoring of tributyltin. PMID- 15269906 TI - Toxicity to Tradescantia of technogenic radionuclides and their mixture with heavy metals. AB - The genotoxic effects on Tradescantia of (137)Cs, (90)Sr, and (236, 242)Pu, a heavy metal mixture [Cd, Cr(VI), Cu, Mn(II), Ni, Pb, Zn] and of a complex mixture of these toxicants were determined. The impact of radionuclides on plants subjected to ionizing radiation exposure was estimated. The number of somatic mutations and the quantity of nonviable stamen hairs were used as end points in the testing. An increase in the quantity of nonviable stamen hairs was observed with increasing internal exposure to (137)Cs, (90)Sr, and (236, 242)Pu; however, the number of somatic mutations was not observed to be dependent on ionizing radiation. The internal dose of individual radionuclides necessary to decrease the quantity of viable stamen hairs in Tradescantia by 50% can be arranged in the following sequence: (236, 242)Pu > (137)Cs > (90)Sr. Tradescantia died in the mixture of the radionuclides (90)Sr, (137)Cs, and (236)Pu (5 x 10(-2), 7 x 10( 5), and 4 x 10(-10) Gy, respectively) after 14 days, whereas the heavy metal mixture caused somatic mutations in 3% of the Tradescantia and nonviable stamen hairs in 7% but no mortality. However, the Tradescantia died in a combined mixture of these heavy metals and the radionuclides after 14 days. On the basis of all these observations, it can be concluded that the toxic effect of radionuclides was more significant than that of heavy metals. PMID- 15269907 TI - Assessment of toxic interactions of heavy metals in a multicomponent mixture using Lepidium sativum and Spirodela polyrrhiza. AB - The toxicities of copper, chromium, cadmium, nickel, manganese, zinc, and lead ions and various concentrations of mixtures of them were studied using the aquatic plant Spirodela polyrrhiza and the terrestrial plant Lepidium sativum. The composition of the model mixture was based on average analytical data of the annual amounts of representative heavy metals (HM) in wastewater discharged from the Ignalina Nuclear Power Plant (Lithuania) during 1996. The observed and predicted effects of the HM mixture on tested plants were evaluated and compared with the prediction models used in describing the toxic interactions of heavy metals in the mixture. The type of toxic interaction at each tested concentration of the mixture was assessed by a statistical approach that tested the null hypothesis of additive toxicity (Ince et al., 1999) and the mixture toxicity index (MTI; Konemann, 1981). For both plant organisms the effect of the HM mixture calculated using the MTI was synergistic. However, assessment of the HM interaction type at 50% effect concentrations using the hypothesis of additive toxicity showed a synergistic effect for Spirodela polyrrhiza and an additive effect for Lepidium sativum. Though the results obtained using both prediction models for assessing the HM mixture's toxicity were similar, in our opinion, the additive toxicity model is more suitable than the MTI model because the former allows evaluation of the impact of various mixture concentrations, not only those with a 50% effect. PMID- 15269908 TI - Spirotox-Spirostomum ambiguum acute toxicity test-10 years of experience. AB - Spirotox is a short-term acute toxicity test with a large ciliated protozoan Spirostomum ambiguum. The test was created in the Department of Environmental Health Sciences, Medical University of Warsaw, Poland, during the early 1990s. It was presented for the first time during the 6th International Symposium on Toxicity Assessment, in Berlin in 1993. S. ambiguum was very resistant to a wide range of environmental conditions, especially to pH and dissolved oxygen. Over the last 10 years the sensitivity of S. ambiguum to many classes of toxicants was evaluated. Spirotox was found to be very sensitive to heavy metals, fungicides, and pharmaceuticals used to cure diseases of the human nervous system. On the other hand, it was generally less sensitive than standard bioassays to simple organics. Spirotox was also used for analysis of cyanobacterial blooms. Though it was moderately sensitive to hepatotoxins, the test seems to be a good tool for evaluation of the entire toxicity of blooms. The last but not the least of the applications of the Spirotox test was evaluation of the toxicity of extracts from medical devices. Protozoa, which are the simplest eukaryotes, seem to be good screening bioassays for monitoring the technology of medical device production. PMID- 15269909 TI - Analysis of micronuclei in blue mussels and fish from the Baltic and North Seas. AB - Micronuclei (MN) were analyzed in erythrocytes of flounder (Platichthys flesus) and wrasse (Symphodus melops) and in gill cells of blue mussels (Mytilus edulis). The organisms were collected from three study stations in the Baltic Sea and from seven stations in the North Sea (Karmsund area, Norway) 4 times. The statistically significant differences obtained were related to the season, sex of the fish, and sampling locality. Higher MN frequencies were found in fish and mussels collected from the most polluted study stations in the North Sea. The same tendency could be described in the Baltic Sea; however, it was masked by the recent oil spill from the Butinge oil terminal. Our results showing higher MN frequencies in presumably what were the most polluted study locations suggest that MN tests in fish and mussels may be used for the detection of genotoxic effects in a marine environment. The endpoint is well characterized and can be easily recognized, and the technique is convenient to use in field samplings following standard procedures and protocols. PMID- 15269910 TI - Elemental sulfur: toxicity in vivo and in vitro to bacterial luciferase, in vitro yeast alcohol dehydrogenase, and bovine liver catalase. AB - The aim of this research was to analyze the effects and the modes of action of elemental sulfur (S(0)) in bioluminescence and respiration of Vibrio fischeri cells and the enzymes crude luciferase, pure catalase, and alcohol dehydrogenase (ADH). Metallic copper removed sulfur and reduced the toxicity of acetone extracts of sediment samples analyzed in the bioluminescence test. The sulfur inhibition of cell bioluminescence was noncompetitive with decanal, the luciferase substrate; reversible, with maximum toxicity after 15 min (EC(50) = 11.8 microg/L); and almost totally recovered after 2 h. In vitro preincubation of crude luciferase extract with sulfur (0.28 ppm) weakly inhibited bioluminescence at 5 min, but at 30 min the inhibition reached 60%. Increasing the concentration of sulfur in the parts per million concentration range in vitro decreased bioluminescence, which was not constant, but depended on exposure time, and no dead-end/total inhibition was observed. The redox state of enzymes in the in vitro system significantly affected inhibition. Hydrogen peroxide restored fully and the reducing agent dithiothreitol, itself toxic, restored only partially luciferase activity in the presence of sulfur. Sulfur (5.5 ppm) slightly inhibited ADH and catalase, and dithiothreitol enhanced sulfur inhibition. High sulfur concentrations (2.2 ppm) inhibited the bioluminescence and enhanced the respiration rate of V. fischeri cells. Elemental sulfur data were interpreted to show that sulfur acted on at least a few V. fischeri cell sites: reversibly modifying luciferase at sites sensitive to/protected by oxidative and reducing agents and by affecting electron transport processes, resulting in enhanced oxygen consumption. Sulfur together with an enzyme reducing agent inhibited the oxidoreductive enzymes ADH and catalase, which have --SH groups, metal ion cofactors, or heme, respectively, in their active centers. PMID- 15269911 TI - Genotoxicity related to transfer of oil spill pollutants from mussels to mammals via food. AB - Heavy fuel oils containing high levels of polycyclic aromatic hydrocarbons (PAHs) were released into the marine environment after the Erika oil spill on the Atlantic coast. As highly condensed PAH pollutants can bioaccumulate in invertebrates, their transfer to vertebrates through the food chain was of concern. This study aimed to estimate potential genotoxic effects in rats fed for 2 or 4 weeks with the marine mussel Mytilus edulis contaminated by oil pollutants. Two levels of PAH contamination were studied, around 100 and 500 microg of total PAHs/kg dry weight (d.w.) in mussels. Genotoxic damage in rats was investigated by single-cell gel electrophoresis (Comet assay) and micronucleus assays in liver, bone marrow, and peripheral blood. DNA damage was observed in the liver of rats fed with the most contaminated mussels (500 microg PAHs/kg d.w.).DNA damage also was observed in the bone marrow but less than that in the liver. A small increase in micronuclei frequency was registered as well. This work underlines the bioavailability of pollutants in fuel-oil-contaminated mussels to consumers and the usefulness of the Comet assay as a sensitive tool in biomonitoring to analyze responses to PAH transfer in food. The occurrence of substituted PAHs and related compounds such as benzothiophenes in addition to nonsubstituted PAHs in fuel oils and mussels raised the question of whether they were implicated in the genotoxic effects registered in rats. PMID- 15269912 TI - Toxicity testing of heavy-metal-polluted soils with algae Selenastrum capricornutum: a soil suspension assay. AB - A small-scale Selenastrum capricornutum (Rhapidocelis subcapitata) growth inhibition assay was applied to the toxicity testing of suspensions of heavy metal-polluted soils. The OECD 201 standard test procedure was followed, and algal biomass was measured by the fluorescence of extracted chlorophyll. The soils, which contained up to (per kilogram) 1390 mg of Zn, 20 mg of Cd, and 1050 mg of Pb were sampled around lead and zinc smelters in northern France. The water extractability of the metals in suspensions (1 part soil/99 parts water w/v) was not proportional to the pollution level, as extractability was lower for soil samples that were more polluted. Thus, the same amount of metals could be leached out of soils of different levels of pollution, showing that total concentrations of heavy metals in soil (currently used for risk assessment purposes) are poor predictors of the real environmental risk via the soil-water path. Despite high concentrations of water-extracted zinc (0.6-1.4 mg/L of Zn in the test), exceeding by approximately 10-fold the EC(50) value for S. capricornutum (0.1 mg Zn/L), 72-h algal growth in the soil extracts was comparable or better than growth in the standard control OECD mineral medium. The soil suspension stimulated the growth of algae up to eightfold greater than growth using the OECD control medium. Growth stimulation of algae was observed even when soil suspensions contained up to 12.5 mg Zn/L and could not be explained by supplementary nitrogen, phosphorous, and carbonate leached from the soil. However, if the growth of algae in suspensions of clean and polluted soils was compared, a dose-dependent inhibitory effect of metals on algal growth was demonstrated. Thus, as soil contains nutrients/supplements that mask the adverse effect of heavy metals, a clean soil that has properties similar to the polluted soils should be used instead of mineral salt solution as a control for analysis of the ecotoxicity of soils. PMID- 15269913 TI - Leachate toxicity assessment by responses of algae Nitellopsis obtusa membrane ATPase and cell resting potential, and with Daphtoxkit F magna test. AB - A microscale bioassay based on 50% inhibition of K(+), Mg(2+)-ATPase activity in a microsomal fraction isolated from Nitellopsis obtusa cells was developed. Compared to that for a plasma membrane fraction purified in a sucrose gradient, the preparation procedure for a microsomal fraction is less time consuming and the yield is substantially higher. Characteristics of the microsomal preparation proved to be similar to those of the highly purified plasma membrane preparation (Manusadzianas et al., 2002), at least for heavy metals. Sensitivity to CuSO(4) of the frozen (-8 degrees C) microsomal fraction [49 +/- 17 (SD) microM; n = 8] did not significantly differ from that of the freshly isolated one (52 +/- 30, n = 8), at least for 40 days. Toxicity of leachate water from Kairiai (northern Lithuania) solid waste landfill was assessed by taking samples from various points including temporary reservoirs and analyzing them immediately after spillage (summer 2002) and after storage for almost 2 years at 4 degrees C-6 degrees C. Two tests with the macrophytic alga Nitellopsis obtusa (Charatox, 45 min EC(50) of resting potential depolarization, and ATPase assay, IC(50) of membrane ATPase activity) and one test with the crustacean Daphnia magna (Daphtoxkit F, 48-h 50% immobilization) tests were used. In general, all three tests showed successively decreasing values of landfill leachate toxicity with an increasing degree of dilution with surface waters. The possibility of employing preserved algal preparations on demand in test batteries seems to be promising, especially in emergencies. PMID- 15269914 TI - Hormonelike effects of humic substances on fish, amphibians, and invertebrates. AB - Humic substances comprise the majority of organic matter in freshwater ecosystems and were thought to be inert or refractory, except for photolytic degradation. However, evidence is increasing that humic substances interact with aquatic organisms similarly to weak anthropogenic chemicals with nonspecific and specific effects. One specific effect is a hormonelike effect, namely, modulation of the number of offspring, which was first described with the nematode Caenorhabditis elegans. Yet a hormonelike effect is not restricted to only the nematode. With the ornamental swordtail fish, Xiphophorus helleri, and the South African clawed frog, Xenopus laevis, we present phenomenological evidence that slight feminization occurred when these vertebrate species were exposed to a synthetic humic substance, a condensation product of polyphenols. The slight feminization was dose dependent. PMID- 15269915 TI - Effect of lead and chromium on reproductive success of Japanese quail. AB - An experimental model based on application of Japanese quail (Coturnix coturnix japonica) lines with high (H) and low (L) concentrations of plasma lactate was designed in order to investigate the effects of lead and chromium on embryonic development and reproductive success of experimental groups representing different genotypes. During the first stage of the trial, mature males from quail lines H and L (marked as generation T) were fed subchronically for 12 weeks with lead as Pb(NO(3))(2) and chromium as K(2)Cr(2)O(7) (dosage 0.8 and 0.142 g/kg, respectively, in the feed). In addition, a binary mixture of both lead and chromium was administered to the third group of males. The control group consisted of untreated birds. After mating with untreated females (8 female x 5 male in each group), progeny with four genotypes were obtained. The chicks were marked as generation F1 and used in the second stage of the trial. The data obtained show that heavy metals decreased the hatchability of sexually mature quail males: chromium (14%), lead (19%), and binary lead-chromium mixture (28%). Early embryonic mortality increased as much as 2-3 times. Single lead and binary lead-chromium additions decreased fertility after treating Japanese quail during the development of sexual maturity: lead increased the number of unfertile eggs up to 30% (twofold), binary lead and chromium, up to 50% (more than threefold) relative to untreated controls. There was no effect of chromium alone on fertility. In general, hatchability, fertility, embryonic mortality and reproductive success of Japanese quail were dependent on age, sex, genotypes, and on single or binary treatment with the inorganic salts of lead and chromium. PMID- 15269916 TI - Increase of crustacean sensitivity to purified hepatotoxic cyanobacterial extracts by manipulation of experimental conditions. AB - Toxic cyanobacterial blooms are one of the most common consequences of water eutrophication. Microbiotests with crustaceans are not expensive and are easy to prepare for screening tests. They can be applied in the determination of bioactivity and interaction between toxic substances in water, including hepatotoxins. The principal aim of this study was to modify the standard conditions in the Thamnotoxkit F trade mark and Artoxkit M in order to increase crustacean sensitivity to purified cyanobacterial extracts containing microcystins. The results reported show that exposure time, higher temperature, and presence of DMSO can increase the sensitivity of microbiotests to microcystins. The best sensitivity with the Artemia salina test was achieved after a 48-h exposure at 25 degrees C. The tests using a 24-h exposure at 27 degrees C were the most sensitive for Thamnocephalus platyurus. The test without preincubation with DMSO provided the best correlation of microcystin concentration and LC(50) for Thamnocephalus platyurus and is recommended. PMID- 15269917 TI - Impact of crude oil on bacteriocenosis of the digestive tract of mollusks. AB - In this study the effect of crude oil on intestinal bacterial populations of the mollusk Viviparus contactus was investigated. The addition of crude oil into an environment of mollusks induced no clear changes in the saprophytic, amylolytic, and total coliform bacterial counts in the digestive tract of the mollusk. After 10 days of contamination, the saprophytic, amylolytic, and total coliform bacterial numbers were of the same order of magnitude as the initial numbers. Significant numbers of indigenous hydrocarbon-degrading bacteria were observed in the intestinal tracts of mollusks before contamination with crude oil. Introduction of crude oil into the mollusk environment resulted in an increase of 2 orders of magnitude in the number of hydrocarbon-degrading bacteria in the digestive tract. Therefore, measuring the hydrocarbon-degrading bacterial populations in the digestive tracts of hydrobionts can be considered an important component of contaminated-site assessment studies. PMID- 15269918 TI - Use of freshwater algae and duckweeds for phytotoxicity testing. AB - The toxicity of contaminated water of different origins and chemicals [Cr(III), Pb(II), Cu(II), Cd(II), pyrene] were tested using four test species: the alga Selenastrum capricornutum (new name Raphidocelis subcapitata), the duckweed Lemna minor, and the crustaceans Thamnocephalus platyurus and Daphnia magna. On the basis of the results obtained, the sensitivity of plant species and problems concerning the interpretation of the phytotoxicity data are discussed. The data indicated that the sensitivities of crustaceans and plant species both to individual contaminants and to mixtures are unpredictable and that there is no reason to consider plant species less sensitive than animal species. Lemna minor is more sensitive than Selenastrum capricornutum. With colored samples, duckweed is preferable for toxicity testing. To raise the predictive utility of the phytotoxicity data, it is recommended that natural water be used in the test procedure. PMID- 15269919 TI - A novel approach for phytotoxicity assessment by CCD fluorescence imaging. AB - Rapidity, cost effectiveness, ecological reliability, and the possibility of the direct interpretation of bioassay results are the main motivations for the development of new approaches in ecotoxicity testing. Color, turbidity, and nutrient content are factors of great importance in phytotoxicity testing of natural samples. Some algal bioassay end points are markedly influenced by such factors or are impossible to estimate in their presence. An algal toxicity test applicable as an early-warning system has to be able to give a signal in the shortest time possible (hours). We used CCD fluorescence imaging to evaluate toxicity effects in algae, cyanobacteria, and vascular plants, and the data were compared with standard end points. Plant physiologists use this device mainly for photosynthesis research, but common photosynthetic parameters used to characterize chlorophyll fluorescence (F(v)/F(m), F(0), and F(m)) or its quenching (NPQ) have only limited ecotoxicological applicability. Previously published estimations based on the geometrical complement to measured data (complementary area) in the fast kinetics (Kautsky effect) respond especially to pollutants affecting electron transport in the PSII. We recommend using a definite integral value combined with relative fluorescence decay (Rfd) to obtain a sensitive and fast toxicity response. Our approach integrates more mechanisms of toxicity (effects on membranes, proton pump, ATP synthesis, etc.) and improves the toxicity signal, which is more ecotoxicologically relevant. This method can give results after 2-6 h of exposure and is especially useful as an early-warning system and for the toxicity assessment of environmental samples with unknown nutrient status. Results with our approach after 4-6 h are comparable with those obtained with a 96-h standard algal assay. A similar methodical approach can be applied for toxicity evaluation of plants or lichens, or for in situ ecotoxicological studies of microphytobenthos communities. PMID- 15269920 TI - Defense systems of benthic invertebrates in response to environmental stressors. AB - Environmental stress factors may be responsible for biological changes in living species that are able to overcome deleterious effects depending on their detoxifying capacities. Defense systems present in every living species are involved in elimination of reactive chemical species of endogenous or exogenous origin, neutralization of their effects, repair of initial lesions, and compensation of deficient metabolic pathways. Consequently, the performance of defense systems and their inducibility will explain adaptation to environmental disturbances, whereas their alteration will augur toxicity in the exposed species. Several field studies have illustrated the relationships between antioxidants and toxicity in benthic invertebrates in rivers. They highlighted that defense systems may be useful biomarkers in mechanistic studies of ecotoxicity and in the biomonitoring of living species in polluted environments. PMID- 15269921 TI - Field study on changes in viability of airborne fungal propagules exposed to UV radiation. AB - The responses of airborne fungi to UV-B under natural conditions were investigated at the coastal station in Preila, Lithuania. Results of this investigation demonstrated that solar radiation has a marked lethal effect on outdoor airborne fungi. Sensitivity to solar radiation was the highest for the fungal propagules collected late in the evening (relative recovery 6.2%) and early morning (25.3%). The lowest sensitivity to solar radiation was observed for fungal propagules collected at midday (50.0%) and in the afternoon (53.0%). The reason for the lethal effect is thought to be elimination of the sensitive fraction of the night-time fungal populations as solar radiation gradually increases beginning at dawn. Among 356 fungus strains isolated during the investigations, 128 can be characterized as potential pathogens, and 21 strains among those most common in fungal populations belonging to plant, animal, and human pathogen groups. The collected fungal populations were exposed to solar UV B irradiation for 2 h (from 12 p.m. to 2 p.m.). Fungal communities in the air samples were composed of saprotrophs, some of which are regarded as potential phytopathogens (Alternaria, Cladosporium, and Fusarium) or as entomopathogens (Beauveria, Paecilomyces, and Metarhizium). The airborne fungal species identified after exposure to solar radiation were predominantly: Aspergillus niger, Alternaria alternata, Cladosporium cladosporioides, Arthrinium phaerosporum, and dematiaceous sterile mycelium. PMID- 15269922 TI - Evaluation of solar UV damage to Crepis capillaris by chromosome aberration test. AB - Ultraviolet (UV) radiation comprises only a small portion of the electromagnetic spectrum of solar light, but it exerts a disproportionally greater genotoxic effect on all organisms, including water plants. However, genotoxicity evaluation of solar UV is complicated because of the simultaneous actions of UVB, UVA, and photoreactivating light (PHL). The latter very effectively repairs the main type of DNA lesions, pyrimidine dimers (PD), which are induced specifically only by UV. However, other types of DNA lesions are induced by UV; they are unrepairable by PHL and present a real danger to the plant genome. To evaluate this part of DNA lesions, the frequency of chromosome aberrations (CA) was determined after solar UVB and UVB+UVA irradiation with or without PHL. Meristematic cells of Crepis capillaris were irradiated in special chambers with filters. The 4-year investigation showed that only about half of CA had been repaired with PHL. Both findings of the study, of the part of CA that remained after PHL and of the stronger genotoxicity of UVB+UVA, are discussed. PMID- 15269923 TI - Acute toxicity investigations of ester-based lubricants by using biotests with algae and bacteria. AB - Although ester-based lubricants are ecologically acceptable due to their good biodegradability, there are still some environmental ecotoxicological impacts that have to be considered. Information on the acute ecotoxicological behavior of lubricants is obtained in this work using several single species bioassays. In previous studies it was observed that lubricating fluids containing additives for the enhancement of their technical performance were more problematic than base fluids especially with respect to algae growth inhibition. In order to clarify the influence of additives, the anti-wear additive tri-n-butyl phosphate was tested. It was very toxic to algae though not to bacteria. Additionally, a mixture of this additive with a base fluid is characterized. Despite the high toxicity of the single additive, the water extract of the mixture of tri-n-butyl phosphate with hydraulic base fluid caused almost no toxicity. Therefore, tri-n butyl phosphate cannot explain the effect observed for the toxicity of water extracts of the commercially available lubricants. PMID- 15269924 TI - Comparative study on sensitivity of higher plants and fish to heavy fuel oil. AB - Laboratory tests were conducted on higher plants [garden cress (Lepidium sativum), great duckweed (Spirodela polyrrhiza), and Tradescantia clone BNL 02] and fish [rainbow trout (Oncorhynchus mykiss) at all stages of development: eggs, larvae and adults] to estimate their sensitivity to heavy fuel oil (HFO). A number of biological indices (survival, growth, and physiological and morphological parameters) as well as the genotoxic impact (Tradescantia) of HFO was evaluated by acute and chronic toxicity tests. Fish were found to be more sensitive to the toxic effect of HFO than were higher plants. EC(50) values obtained for higher plants ranged from 8.7 g/L (L. sativum) to 19.8 g/L (Tradescantia), and maximum-acceptable-toxicant concentration (MATC) values ranged from 0.1 to 1.0 g/L of total HFO for L. sativum and Tradescantia, respectively. The 96-h LC(50) values ranged from 0.33 g/L, for larvae, to 2.97 g/L, for adult fish, and the MATC value for fish was found to be equal to 0.0042 g/L of total HFO. To evaluate and predict the ecological risk of the overall effects of oil spills, studies should be performed using a set of acute and chronic bioassays that include test species of different phylogenetic levels with the most sensitive morphological, physiological, and genotoxic indices. PMID- 15269925 TI - Rhabdomyolysis with HMG-CoA reductase inhibitors and gemfibrozil combination therapy. AB - CONTEXT: Elevated total cholesterol (total-C) and low-density lipoprotein cholesterol (LDL-C) levels are established risk factors for cardiovascular disease (CVD). HMG-CoA reductase inhibitors (statins) are effective cholesterol lowering drugs that are commonly prescribed to treat this condition. These drugs are often combined with another class of drugs, fibric acid derivatives, to lower both cholesterol and triglyceride levels. Rhabdomyolysis is a known, rare serious side effect of statin monotherapy and of statin-fibrate combination therapy. OBJECTIVE: To examine Food and Drug Administration's (FDA's) postmarketing database for cases of rhabdomyolysis in relation to monotherapy and combination use and calculate reporting rates for this event. DESIGN: Domestic cases of statin- and statin/gemfibrozil-associated rhabdomyolysis were culled from FDA's database and reviewed. Rhabdomyolysis was defined by CPK > or = 10,000 IU/L, myopathic signs and symptoms and clinical diagnosis of rhabdomyolysis. Reporting rates, consisting of number of reported cases/number of prescriptions for each drug, were then calculated to determine whether the reporting of rhabdomyolysis cases was commensurate with extent of use of each statin in the population. SETTING: Cases were obtained from the FDA adverse event reporting system (AERS) database. PATIENTS: NA. MAIN OUTCOME MEASURES: Number of rhabdomyolysis cases were evaluated, along with outcomes, such as renal failure, dialysis and death. RESULTS: Of 866 total reported cases, 482 (56%) were associated with monotherapy and 384 (44%) related to combination therapy. More than 80% of reported cases for each drug resulted in hospitalization for renal failure and dialysis. 80 patients expired from events related directly to rhabdomyolysis. Reporting rates for all statins, except for cerivastatin, were similar and much lower than 1 per 100,000 prescriptions. The cerivastatin-reporting rate was much higher at 4.24/100,000 prescriptions. CONCLUSIONS: Rhabdomyolysis is a rare, serious side effect of statin monotherapy and of statin-fibrate combination therapy. Clinicians need to remain cognizant of this potential adverse event and discuss signs and symptoms of muscle toxicity with patients in order improve the benefits-to-risks of treating dyslipidemia with statins. PMID- 15269926 TI - Venous thromboembolism associated with cyproterone acetate in combination with ethinyloestradiol (Dianette): observational studies using the UK General Practice Research Database. AB - PURPOSE: To derive risk estimates for venous thromboembolism (VTE) in women prescribed cyproterone acetate combined with ethinyloestradiol (CPA/EE), a drug licensed in the UK for the treatment of women with acne or hirsutism. CPA/EE provides a treatment option for women with polycystic ovary syndrome (PCOS). CPA/EE has been associated with an increased risk of VTE. METHODS: Using the General Practice Research Database, we conducted cohort and case-control analyses in all women aged 15-39 and then nested in a population of women of the same age with acne, hirsutism or PCOS. RESULTS: The incidence rate ratio (IRR) for VTE in women exposed to CPA/EE versus conventional combined oral contraceptives (COCs) was significantly raised (all women: 1.92; 95% CI: 1.22,2.88; nested: 2.51; 95% CI: 1.07,5.75). Using exposure to conventional COCs as the reference, the adjusted odds ratio (ORadj) for VTE associated with CPA/EE was 1.45 (95% CI: 0.80,2.64) in all women and 1.71 (95% CI: 0.31,9.49) in women with acne, hirsutism or PCOS. CONCLUSIONS: The risk of VTE associated with CPA/EE use does not differ significantly from that associated with the use of conventional COCs. These data are reassuring and together with knowledge of the risks associated with other treatments for acne, in particular, should influence prescribing practice. PMID- 15269927 TI - Agreement between GPRD smoking data: a survey of general practitioners and a population-based survey. AB - BACKGROUND: Cigarette smoking is a common habit that is associated with many diseases. Smoking is often an important confounding variable in pharmacoepidemiological studies. The General Practice Research Database (GPRD) is widely used in pharmacoepidemiological research. In this study, we compare data recorded in the GPRD with the smoking history obtained from direct query of general practitioners (GPs) and from a population-based survey. METHODS: We completed a mailed survey of GPs caring for a random sample of 150 patients with inflammatory bowel disease. The survey asked the GP to categorize the patients smoking status on a specified date. These results were then compared to the data recorded in the GPRD. Smoking status of 225,308 randomly selected GPRD patients without inflammatory bowel disease was compared to the results of a population based household survey. RESULTS: Completed surveys with usable data were received from GPs on 136 of the 150 patients (91%). The sensitivity and positive predictive value of the database for current smoking were 78% (95% CI: 52-94) and 70% (95% CI: 46-88) respectively. The sensitivity and positive predictive value of former smoking were 53% (95% CI: 28-77) and 60% (95% CI: 32-84) respectively. Current and former smoking rates in the GPRD were 79% and 29% respectively of expected rates according to the population-based survey. CONCLUSIONS: Current smoking is more completely recorded in the GPRD than former smoking. These data need to be considered when planning GPRD studies where smoking is an important exposure variable. PMID- 15269928 TI - The use of drugs in mothers of offspring with neural-tube defects. AB - OBJECTIVE: To study the risk of maternal drugs use during pregnancy in the origin of isolated neural-tube defects (NTD). MATERIALS AND METHODS: 1202 cases with NTD, 38,151 population controls without any defects and 22,475 patient controls with other defects were compared in the population-based data set of the Hungarian Case-Control Surveillance of Congenital Abnormalities (HCCSCA), 1980 1996. The HCCSCA contains 542 drugs, however only those drugs were evaluated which included five or more mothers in the NTD group. Drugs with the same chemical structures were combined. In addition, only drug use in the second month of pregnancy was evaluated because it is the critical period for NTD. Of course, it is necessary to exclude different biases, mainly recall bias at the evaluation of these drugs. Of 121 chemicals, only oxytetracycline, carbamazepine and valproic acid had some association with NTD. High doses of exogenous oestrogens, clomiphene, chorionic gonadotropin, lynesterol and ergotamine also seemed to have some indirect association with NTD because their exposures occurred more frequently before the critical period of NTD due to maternal infertility. CONCLUSION: Our findings suggest that drugs used during pregnancy do not appear to substantially contribute to the occurrence of isolated NTD but some drugs have a role in the origin of these defects. PMID- 15269929 TI - Pharmacists' role in reporting adverse drug reactions in an international perspective. AB - INTRODUCTION: The participation of the pharmacist in national spontaneous reporting systems for adverse drug reactions (ADRs) has not always been a matter of course. Even today, there are a number of countries, in particular the Scandinavian countries, where pharmacists are not authorised to report ADRs. In those countries in which they are allowed to report, they do not always use this opportunity. METHODS: We have conducted a review of the literature to investigate the involvement of pharmacists in ADR reporting. In addition, we evaluated the pharmacists' actual contributions in 2001 by means of an international questionnaire-based survey among the countries participating in the WHO Drug Monitoring Programme in September 2002. Apart from the numbers of pharmacists' reports, respondents were asked to indicate their assessment of both the quality and the significance of the contribution. Of the 68 participating countries, 41 responded by returning the questionnaire. RESULTS AND CONCLUSIONS: The appreciation of pharmacists' ADR reports is high in those countries that have more experience with greater numbers of pharmacists' reports. The countries that received fewer reports from pharmacists gave lower scores to their contribution. If the specific contribution pharmacists can make to the quantity and quality of ADR reports were to be exploited to a greater extent, this could lead to a substantial improvement of the international adverse drug reactions reporting system. PMID- 15269930 TI - Use of antibiotics at hospitals in Stockholm: a benchmarking project using internet. AB - PURPOSE: To evaluate the internet way of feedback to prescribing physicians, providing data on antimicrobial use and to assess the applicability of the DU90% (i.e. Drug Utilisation 90%--the number of drugs accounting for 90% of the volume of usage in defined daily doses, (DDDs)) methodology in the hospital setting. METHODS: Antimicrobial drug use was evaluated in all major departments in seven hospitals in Stockholm in the year 2000. All data were presented anonymously on www.janusinfo.org. Aggregate data on antimicrobial drug use were expressed as the number of DDDs and costs per 100 bed-days. We focused on the number of drugs accounting for 90% of the volume (DDD), including the level of adherence to guidelines. The chief physicians assessed this feedback by a questionnaire. RESULTS: The number of DDDs/100 bed-days varied among different clinics and ranged from 39 to 57 (internal medicine) to 102 to 161 (infectious disease). The cost per 100 bed-days varied more than two-fold. The number of different antibiotics within the DU90% segment ranged from 9 to 13 (orthopaedic clinics) to 16 to 23 (infectious disease). According to the questionnaire, data were considered to be clearly presented and physicians would like to receive this kind of reports in the future, not only for antibiotics but also for other drugs. CONCLUSION: Presenting physicians with aggregate data on drug use via internet could provide a stimulus for prescribing improvement. PMID- 15269931 TI - Assessment of doctor-shopping for high dosage buprenorphine maintenance treatment in a French region: development of a new method for prescription database. AB - PURPOSE: To assess the extent of doctor-shopping for buprenorphine maintenance therapy in a French region with a specific indicator. METHODS: Use of a quasi exhaustive prescription database in a French region (information system of the French General Health Insurance Scheme). Extraction of all buprenorphine prescriptions between September 1999 and December 2000. Definition and calculation of three quantities for each patient: delivered, prescribed and doctor-shopping quantity. The calculation of these three quantities is done by an automated and reproducible method determining the overlaps in prescription periods of different physicians for a given patient. Calculation of the corresponding daily dose was done for each quantity. RESULTS: A total of 64 326 prescriptions of buprenorphine by 1313 physicians to 3259 patients were extracted. Quantities and doses were calculated for 2587 patients. The total doctor-shopping quantity represented 18.6% of the delivered quantity. Doctor shopping involved a minority of patients and was highly concentrated: 87 patients with doctor-shopping doses superior to 16 mg/day were responsible for 45.4% of the total doctor-shopping quantity. CONCLUSIONS: Doctor-shopping appears to be an important problem for buprenorphine maintenance treatment in France but may be resolved by regulatory interventions. The use of adequate indicators on prescription databases may help to limit the effects of such interventions on legitimate care. The method presented here may be used with slight adaptations for other medications to assess their abuse potential. PMID- 15269932 TI - Under-reporting of serious adverse drug reactions in Sweden. AB - INTRODUCTION: Adverse drug reactions (ADR) constitute a major problem, both from a medical point of view and as an economical burden. Spontaneous reporting of ADRs is one of the methods for post marketing surveillance of drug safety. Under reporting can also provide an important obstacle to rapid and relevant signal detection. AIM: To investigate the rate of under-reporting serious ADRs of selected ICD 10 diagnoses. METHOD: In order to investigate the under-reporting rate we investigated at five hospitals within the county of Norrbotten in Sweden the total number of diagnosed cases during a period of 5 years (1996-2000) with the following diagnoses: cerebral haemorrhage (I 61.0-I 61.9), pulmonary embolism (I 26.0 and I 26.9), embolism or thrombosis (I 74.0-I 74.9), phlebititis, thrombophlebitits or venous thrombosis (I 80.0-I 80.3, I 80.8 and I 80.9) and portal vein thrombosis and other thrombosis or emboli (I 82.0-I 82.3, I 82.8 and I 82.9). The identity of these patients was obtained through a database search. The patients' case records were then scrutinized by a specially trained nurse and the drugs used at the time of the event were noted. An assessment of the possibility of an ADR was performed using standard WHO causality criteria. Later, database search in the Swedish ADR registry was performed in order to investigate whether these suspected ADRs had been reported to the national authority in Sweden or not. RESULTS: In total 1349 case records were found and scrutinized. Of these, 107 patients had received drugs that could have been a probable or possible cause to the diagnoses. Of these 92 cases had not been reported and only 15 patients were found in the database, giving an overall under-reporting rate of all ADRs of 86%. The most commonly occurring diagnoses were cerebral haemorrhage followed by venous thrombosis, 545 and 468 respectively. Among those cases that should have been reported according to the existing rules for spontaneous reporting of suspected ADRs the most frequently occurring diagnosis was cerebral haemorrhage (I 61.0) in connection to treatment with anticoagulants. CONCLUSION: The rate of spontaneous ADR reporting is very low, also for serious and fatal reactions. PMID- 15269933 TI - Predictors of herbal medicine use in a Swedish health practice. AB - OBJECTIVE: The aim of this study was to characterise users of herbal medicines and assess the effect of socio-demographic characteristics, perceived health and chronic disease on the use of herbal medicine in a multi-ethnic Swedish health practice population. METHODS: A questionnaire was completed by 1433 (out of a total of 1776) patients aged 16 years and above who visited the Jordbro Health Centre (JHC) in Stockholm, Sweden, between 14 January and 30 June 2002. The results were linked to computerised medical records. RESULTS: Altogether 320 (22.3%) of this patient population used some form of herbal medicine. The bivariate analysis showed that the use of herbal medicine were more common among patients aged 45-64 years, females, high educated, patients born in Nordic countries or Europe compared to other age groups, males, low educated, patients born outside Europe and without chronic disease. In the logistic regression analysis when the effects of confounders were taken into account, females, high educated patients and patients with chronic disease had higher odds for use of herbal medicine than males, low educated and patients without chronic disease. The odds ratios (ORs) and 95% confidence interval (95% CI) were 1.95 (1.40-2.76) for female as compared with male patients; 2.10 (1.49-2.97) for subjects with a high level of education compared with subjects with a low educational level and 1.62 (1.15-2.29) for subjects with chronic disease compared with subjects without chronic disease. The common diagnoses were musculo-skeletal, respiratory and circulatory disorders, signs and symptoms and external causes according to the International Classification of Diseases (ICD10). There were no significant differences between users and non-users of herbal medicine regarding the number of consultations with any physician or the general practitioner (GP), contacts with the health care centre, use of prescribed medicines or number of days of sick leave during the past year. CONCLUSION: Females, well-educated patients and patients with chronic disease had higher odds for use of herbal medicine than others irrespective of other socio-demographic characteristics, and herbal medicine was seen to be used independently of conventional medicine. PMID- 15269934 TI - Does uncollected medication reduce the validity of pharmacy dispensing data? AB - PURPOSE: When using pharmacy data as collected in the InterAction database (IADB) for pharmacoepidemiology studies, we tend to ignore the fact that filled prescriptions are not always collected by the patient. This study investigated whether unclaimed prescriptions pose a validity threat for pharmacy data, by estimating the percentage of filled prescriptions that are not collected, the percentage of patients who do not collect their filled prescriptions and describing the items remaining unclaimed. METHODS: Prospective study in three independent pharmacies in the region that is covered by the IADB. All prescriptions that entered these pharmacies on 3 days in 1 week in October 2002 were monitored for a month with respect to whether and when they were claimed. RESULTS: A total of 3946 prescriptions concerning 3082 patients were filled. The majority of prescriptions were collected the day they were filled. In total 18 prescription items (0.46%) were not collected within 1 month; excluding health products and homeopathic drugs 13 remained. These 13 covered a variety of drug groups; 0.45% of the patients did not claim their medication within 1 month. CONCLUSIONS: Primary non-compliance due to not claiming medication has little impact on the validity of pharmacy dispensing data in the region under study. PMID- 15269935 TI - The 2003 Nobel prize for MRI: significance and impact. PMID- 15269938 TI - Nobel Prizes for nuclear magnetic resonance: 2003 and historical perspectives. PMID- 15269940 TI - The historical documentation of scientific developments: scientists should participate. PMID- 15269941 TI - Significant events in the development of MRI. PMID- 15269942 TI - Differentiation of metastases from high-grade gliomas using short echo time 1H spectroscopy. AB - PURPOSE: To determine if short echo time (TE) (1)H magnetic resonance spectroscopy (MRS) can distinguish between intracranial metastases and glioblastomas. MATERIALS AND METHODS: TE 30-msec spectra were acquired (1.5 T) from voxels entirely within tumors from 23 glioblastoma patients and 24 metastases patients (3 breast carcinomas, 1 bladder carcinoma, 8 lung carcinomas, 3 probable lung carcinomas, 6 melanomas, 1 stomach carcinoma, and 2 undetermined). Spectra were analyzed quantitatively (LCModel) to determine metabolite, lipid, and macromolecule concentrations. All tumors were previously untreated and classified histopathologically. RESULTS: The lipid peak area (LPA) ratio (total peak area at ca. delta1.3 to that at ca. delta0.9) was 2.6 +/- 0.6 (N = 25) for glioblastomas and 3.8 +/- 1.4 (N = 34) for metastases (P < 0.0001). There were no significant differences in metabolite or lipid concentrations between the tumor groups. The LPA ratio provided 80% sensitivity and 80% specificity for discriminating metastases from glioblastomas. CONCLUSION: Lipid and macromolecule (LM) signals can dominate (1)H spectra of high-grade tumors and have characteristics that allow significant discrimination of metastases from glioblastomas. Work is now needed to determine the source and biophysical characteristics of these LM signals to further improve differentiation by optimizing the data acquisition and analysis protocol. PMID- 15269943 TI - A comparison of images generated from diffusion-weighted and diffusion-tensor imaging data in hyper-acute stroke. AB - PURPOSE: To compare isotropic (combined diffusion-weighted image [CMB], apparent diffusion coefficient [ADC], TRACE, exponential ADC [eADC], and isotropically weighted diffusion image [isoDWI]) and anisotropic (relative anisotropy [RA], fractional anisotropy [FA], and volume ratio [VR]) diffusion images collected with fast magnetic resonance (MR) diffusion-weighted (DWI) and diffusion-tensor (DTI) acquisition strategies (each less than one minute) in hyper-acute stroke. MATERIALS AND METHODS: Twenty-one patients suffering from ischemic stroke-imaged within six hours of symptom onset using both DWI and DTI-were analyzed. Regions of interest were placed in the ischemic lesion and in normal contralateral tissue and the percent difference in image intensity was calculated for all nine generated images. RESULTS: The average absolute percent changes for the isotropic strategies were all > 38%, with isoDWI found to have a difference of 50.7% +/- 7.9% (mean +/- standard error, P < 0.001). The ADC maps had the most significant difference (-42.4% +/- 2.0%, P < 0.001, coefficient of variation = 0.22). No anisotropic images had significant differences. CONCLUSION: Anisotropic maps do not consistently show changes in the first six hours of ischemic stroke; therefore, isotropic maps, such as those obtained using DWI, are more appropriate for detecting hyper-acute stroke. Anisotropic images, however, may be useful to differentiate hyper-acute stroke from acute and sub-acute stroke. PMID- 15269944 TI - Development of brain infarct volume as assessed by magnetic resonance imaging (MRI): follow-up of diffusion-weighted MRI lesions. AB - PURPOSE: To investigate the development of ischemic brain lesions, as present in the acute stroke phase, by diffusion-weighted magnetic resonance imaging (DWI), and in the subacute and chronic phases until up to four months after stroke, in fluid-attenuated inversion recovery (FLAIR)- and T2-weighted (T2W) magnetic resonance (MR) images. MATERIALS AND METHODS: Twelve consecutive patients with their first middle cerebral artery (MCA) infarction were included. Lesion volumes were assessed on T2W images recorded with a turbo spin echo (TSE) and on images recorded with the FLAIR sequence on average on day 8 and after about four months. They were compared with acute lesion volumes in perfusion and DWI images taken within 24 hours of stroke onset. RESULTS: On day 8, lesion volumes in images obtained with FLAIR exceeded the acute infarct volumes in DWI. The chronic lesion volumes were almost identical in T2W and FLAIR images but significantly reduced compared with the acute DWI lesions. The lesion volumes assessed on DWI images correlated highly with the lesions in the images obtained with TSE or FLAIR, as did the lesions in the images obtained with FLAIR and TSE. The secondary lesion shrinkage was accompanied by ventricular enlargement and perilesional sulcal widening, as most clearly visible in the images obtained with FLAIR. CONCLUSION: Our results show that the acute DWI lesions are highly predictive for the infarct lesion in the chronic stage after stroke despite a dynamic lesion evolution most evident in MR images obtained with FLAIR. PMID- 15269945 TI - Correlative MR imaging and histopathology in porcine neurocysticercosis. AB - PURPOSE: To examine whether all the histopathologically seen features of cysticercus cysts excised from brain of swine naturally infected with neurocysticercosis during its evolution are actually visible on magnetic resonance imaging (MRI). MATERIALS AND METHODS: Five swine naturally infected with cerebral cysticercosis were subjected to fast spin-echo (SE) T2, SE T1, fluid attenuated inversion recovery imaging, T1-weighted magnetization transfer (MT), and postcontrast T1-weighted MT sequences on MRI. These animals were sacrificed after imaging and ex vivo imaging of the intact excised brain using the same imaging protocol was also performed. Grossing of these brains was done similar to the ex vivo imaging planes. Numeral density and external appearance of each cyst and scolex were evaluated on each pulse sequence. Amount of pericystic edema, if present, was also assessed. On histopathology, cellular characteristics, inflammatory response, and the extent of edema, if present, in the brain parenchyma around the cysts were graded. Cysts were categorized into viable, early, and late degenerated on histopathology. The MRI features of each cyst were correlated with their histopathologic findings. RESULTS: Out of 31 cysts, eight were found to be viable, 13 early degenerated, and 10 late degenerated on histopathology. T2-weighted imaging demonstrated all the cysts while T1-weighted imaging showed 97% of the cysts. Scolex was seen in 90.3% and 93.5% of the cysts on T2- and T1-weighted images, respectively. Minimal edema (grade I) and inflammation in degenerating cysts present on histopathology was not visible on MRI. All but one of eight degenerated cysts, which showed enhancement on postcontrast MRI, had edema on imaging as well as on histopathology. CONCLUSION: T2-weighted MRI demonstrated all the cysts that were visible on histopathology. Non-enhancement of some of the degenerated cysts along with absence of edema on MRI is likely to underestimate the staging of neurocysticercosis evolution, and these early degenerating cysts may be misdiagnosed as in viable stage. PMID- 15269946 TI - Diffusion anisotropy in subcortical white matter and cortical gray matter: changes with aging and the role of CSF-suppression. AB - PURPOSE: To determine the relevance of cerebrospinal fluid (CSF)-suppression for the measurement of diffusion anisotropy in well-localized areas of the brain, particularly the subcortical white matter (WM) within the gyri and cortical gray matter (GM), in young and elderly subjects, and to assess the changes of water diffusivity in the brain with normal aging. MATERIALS AND METHODS: Quantitative measures of anisotropy in 26 regions, including subcortical WM (i.e., in the gyri), cortical GM, major deep WM, and deep GM regions of young (21-25 years, N = 8) and elderly (61-74 years, N = 10) normal volunteers, were assessed with CSF suppressed diffusion tensor imaging (DTI) relative to standard DTI. RESULTS: CSF suppressed DTI demonstrated significant increases in fractional anisotropy (FA) of 3-12% in the young and 2-14% in the elderly groups with the largest changes being in the subcortical WM of the gyri. Furthermore, FA decreased by 10-19% in the subcortical WM of the gyri of the elderly subjects relative to the young, primarily due to increases in the perpendicular diffusivity, lambda(3), with age. CONCLUSION: CSF-suppressed DTI yields more accurate measures of quantitative anisotropy in cortical and subcortical brain regions. Reductions of anisotropy with aging were predominantly observed in subcortical WM of the gyri. PMID- 15269947 TI - Request form history, clinical indication, and yield of brain magnetic resonance studies. AB - PURPOSE: To investigate whether improved clinical history allows the radiologist to better predict the pretest probability of obtaining a positive or negative result from a magnetic resonance (MR) examination. MATERIALS AND METHODS: Six neuroradiologists prospectively reviewed 100 consecutive requests for brain MR examinations and sequentially assessed 1) quality of written history, 2) degree of indication for requested study, and 3) any pertinent new information found during chart review that may have altered the degree of indication. MR yield was correlated with the degree of indication assessed before and after chart review. RESULTS: Most request form histories were judged as poor (63%), and chart review reduced the overall indications for MR examinations, as there was a tendency for high-indication requests to migrate to the low-indication category. Based on request form history alone, the yields for low- and high-indication studies were 13% and 37%, respectively. Correlations between MR yield and indication after chart review improved significantly (P < 0.05) with 2% and 61% for low and high indications, respectively. Sensitivity and specificity for a positive MR yield were 71% and 62%, respectively, for the indication judged by the request history alone, and 96% and 80%, respectively, after chart review. Positive and negative prediction rates were 37% and 87%, respectively, for the indication judged by the request history alone, and 61% and 98%, respectively, for the indication judged after chart review. CONCLUSION: Based on our limited data, most request form histories were inadequate, and essential information available in the chart before MR examinations was frequently missing from the request forms. When adequate information was provided, the indication for the studies as judged by the radiologists predicted the MR yield more accurately, particularly for those requests with low indication. Therefore, our study suggests that MR imaging (MRI) may be used more effectively when pertinent clinical history is available. However, our study is limited and further studies are needed to confirm our results. PMID- 15269948 TI - Relationship of apparent myocardial T2 and oxygenation: towards quantification of myocardial oxygen extraction fraction. AB - PURPOSE: To explore the relationship of myocardial T(2) and oxygenation for the quantification of myocardial oxygen extraction fraction (OEF). MATERIALS AND METHODS: A proposed myocardial T(2)-OEF relationship was evaluated by computer simulation and in nine normal dogs in vivo. The relationship was based on a simplified two-compartment T(2) model. In the dogs, dipyridamole was infused intravenously to increase blood flow and change in myocardial oxygen content. The accuracy of the measurement in myocardial OEF in vivo by magnetic resonance imaging (MRI) was determined by arterial and coronary sinus blood sampling. RESULTS: Global myocardial T(2) increased 16.1% from rest to the peak of dipyridamole-induced vasodilation (44.6 +/- 2.1 msec vs. 51.4 +/- 2.1 msec, P < 0.001). Corresponding OEF measured by arterial and venous (AV) sampling decreased from 0.64 +/- 0.15 at rest to 0.18 +/- 0.08 during the dipyridamole vasodilation, whereas OEF calculated by MRI at the peak effect of dipyridamole was 20 +/- 4%. Global myocardial OEF measured dynamically by MRI showed a strong correlation with OEF measured by blood sampling (correlation coefficient (CC) = 0.83) during pharmacologic vasodilation. CONCLUSION: When combined with vasodilator stress, assessment of OEF may provide a putative measure of myocardial flow reserve, allowing consecutive monitoring of myocardial dose-responses to a variety of interventions and offering a new tool for the detection of coronary artery disease. PMID- 15269949 TI - Early heterogeneous enhancement of the liver: magnetic resonance imaging findings and clinical significance. AB - PURPOSE: To investigate and assess the radiologic, serological, and histopathologic findings in patients who presented with early heterogeneous enhancement (EHE) on gadolinium-enhanced early-phase magnetic resonance imaging (MRI) of the liver. MATERIALS AND METHODS: We searched our radiologic records of MRI of the liver from July 1999 to April 2002 to identify patients with EHE. Three investigators retrospectively evaluated in consensus the MR images in each patient for intensity and characteristic of EHE blinded to clinical information. Serological laboratory values and clinical information were obtained in all patients, and histologic findings were available in 19. RESULTS: We identified 67 patients with EHE. Of them, 62 patients (93%) had underlying chronic liver disease. Twenty-seven patients had viral hepatitis, 13 had alcohol abuse, 6 had primary sclerosing cholangitis, and the others had miscellaneous etiologies. The five patients without chronic liver disease had the following clinical histories: concurrent chemotherapy for extrahepatic malignancy (two patients), concurrent intraabdominal infection (one), and no known associated disease (two). Intensity of EHE was intense in 6 (9%), moderate in 22 (33%), and mild in 39 (58%). Pattern of EHE was geographic in 15 patients (22%), patchy in 37 (55%), and miliary in 15 (22%). All EHE showed rapid fading on postcontrast late-phase images. EHE showed mild to moderate hyperintensity on T2-weighted images in 30 patients (45%). In the 19 patients with histological correlation, 19 (100%) had hepatocellular necrosis, 19 (100%) had fibrosis, 18 (95%) had inflammatory cell infiltration, 17 (89%) had capillary-size vessels within fibrous septa, and 16 (84%) had ductal proliferation. No statistical correlation was found between the intensity or pattern of EHE on MR images and the extent of elevation of serological laboratory values or severity of histologic findings. CONCLUSION: Several different types of underlying chronic liver disease were observed in most of the patients with EHE. Hepatocyte necrosis, fibrosis, or inflammatory processes were found in all patients with EHE who had histopathological correlation. PMID- 15269950 TI - Nodule-in-nodule appearance of hepatocellular carcinomas: comparison of gadolinium-enhanced and ferumoxides-enhanced magnetic resonance imaging. AB - PURPOSE: To perform comparison of gadolinium-enhanced and ferumoxides-enhanced magnetic resonance imaging (MRI) in the detection of nodule-in-nodule appearance of hepatocellular carcinomas (HCCs). MATERIALS AND METHODS: During a recent 45 month period, we had eight patients (five men and three women; age range, 63-84 years; mean, 71 years) with HCCs with nodule-in-nodule appearance who underwent gadolinium-enhanced MRI, ferumoxides-enhanced MRI, and computed tomography during arterial portography (CTAP) and computed tomography during hepatic arteriography (CTHA), combined and separately, within an interval of two weeks. Two blinded radiologists in consensus retrospectively evaluated three sets of sequences: unenhanced T1- and T2-weighted MR, gadolinium-enhanced MR, and ferumoxides enhanced MR images in random order of patients and imaging sequences. The depiction degree of nodule-in-nodule appearance of HCC was evaluated in a semiquantitative fashion. The sensitivities of unenhanced T1- and T2-weighted, gadolinium-enhanced, and ferumoxides-enhanced MR images were compared with McNemar's test. RESULTS: The eight HCCs with nodule-in-nodule appearance ranged in size from 16-26 mm (mean, 20.0 +/- 4.0 mm), and there existed nine internal HCC foci ranging in size from 5-14 mm (mean, 7.9 +/- 3.5 mm). On gadolinium enhanced MR images, the nodule-in-nodule appearance of HCC was typically seen as hypervascular foci in an iso- or hypovascular area: the depiction degree of nodule-in-nodule appearance was distinct in two lesions, equivocal in three, and absent in three. On ferumoxides-enhanced MR images, it was typically seen as hyperintense foci in a hypointense area: the depiction degree was distinct in four, moderate in one, and absent in three. The sensitivities for detection of nodule-in-nodule appearance were 25%, 25%, and 63% on T1- and T2-weighted, gadolinium-enhanced, and ferumoxides-enhanced MR images, respectively, but there was no significant difference in sensitivity. CONCLUSION: Nodule-in-nodule appearance of HCCs can be seen on ferumoxides-enhanced MR images, in some cases more clearly than on gadolinium-enhanced MR images, particularly when the background nodule shows hyperintensity on precontrast T1-weighted images. Ferumoxides-enhanced MRI may be considered when development of malignant foci is suspected during routine examinations. PMID- 15269951 TI - Contrast-enhanced magnetic resonance cholangiography using mangafodipir compared with standard T2W MRC sequences: a pictorial essay. AB - Mangafodipir, a manganese-containing hepatobiliary contrast agent, is excreted in bile. We review the principles and practice of a contrast-enhanced MRC technique using mangafodipir and compare it with standard T2-weighted magnetic resonance cholangiography (MRC) sequences. Potential applications include the evaluation of leaks and strictures; the assessment of drainage in normal, surgically by-passed, stented and obstructed biliary systems; the diagnosis of cholecystitis; and the evaluation of normal and variant biliary anatomy. PMID- 15269952 TI - Combined magnetic resonance urography and targeted helical CT in patients with renal colic: a new approach to reduce delivered dose. AB - PURPOSE: To determine whether magnetic resonance urography (MRU), obtained before helical computed tomography (CT) in patients with acute renal colic, can help delimit the obstructed area to be subsequently examined by a targeted CT scan, thus reducing the dose of radiation. MATERIALS AND METHODS: Patients (51) with symptoms of acute renal colic underwent MRU and a total urinary tract helical CT. CT images from the 5 cm below the level of ureteral obstruction as demonstrated by MRU were selected out. Combined interpretation of MRU and selected CT images constituted protocol A. Protocol B consisted of the entire unenhanced helical CT of the urinary tract. The two protocols were compared regarding the following points: 1) sensitivity in diagnosing the presence of obstructing urinary stones, and 2) the delivered radiation dose. RESULTS: Protocol A and protocol B had, respectively, 98% and 100% sensitivity in demonstrating ureteral stone as a cause of renal colic. Estimated average dose calculated from phantom study was 0.52 mSv for protocol A and 2.83 mSv for protocol B. Therefore, the effective radiation dose was 5.4 times lower in protocol A compared to protocol B. CONCLUSION: Combined MRU and short helical CT has a high sensitivity in detecting ureteral calculi with a reduced radiation dose. PMID- 15269953 TI - Impact of diet on stool signal in dark lumen magnetic resonance colonography. AB - PURPOSE: To examine the magnetic resonance (MR) properties of different foods and their effect on the colonic stool signal to potentially support fecal tagging strategies for dark lumen MR colonography (MRC). MATERIALS AND METHODS: T1 relaxation times of 120 different foods (partially diluted with sufficient water) were determined by use of a multi-flip-angle two-dimensional gradient echo (GRE) sequence and correlated to the foods' signal in a three-dimensional GRE volumetric interpolated breath-hold examination (VIBE) sequence. Different dilutions of six foods were examined. VIBE stool signal was determined in six volunteers under two different conditions: after a three-day diet of short T1 food and of long T1 food, respectively. RESULTS: Most foods exhibit short to very short T1 relaxation times. T1 correlates well with the fat-saturated VIBE signal except for fatty products. Diluted food exhibits T1 times similar to water; concentrated food strongly varies according to their T1 values. No significant difference in stool signal could be found in the in vivo examination comparing the two diets. CONCLUSION: According to our results, a restricted diet strategy to reduce fecal signal for dark lumen MRC is unlikely to be successful. Moreover, the stool signal reduction found in the other fecal tagging studies can be explained at least to a great extent by the relative content of other material with long T1 relaxation times, such as water or oral barium. PMID- 15269954 TI - Combined quantitative dynamic contrast-enhanced MR imaging and (1)H MR spectroscopic imaging of human prostate cancer. AB - PURPOSE: To differentiate prostate carcinoma from healthy peripheral zone and central gland using quantitative dynamic contrast-enhanced (DCE) magnetic resonance (MR) imaging and two-dimensional (1)H MR spectroscopic imaging (MRSI) combined into one clinical protocol. MATERIALS AND METHODS: Twenty-three prostate cancer patients were studied with a combined DCE-MRI and MRSI protocol. Cancer regions were localized by histopathology of whole mount sections after radical prostatectomy. Pharmacokinetic modeling parameters, K(trans) and k(ep), as well as the relative levels of the prostate metabolites citrate, choline, and creatine, were determined in cancer, healthy peripheral zone (PZ), and in central gland (CG). RESULTS: K(trans) and k(ep) were higher (P < 0.05) in cancer and in CG than in normal PZ. The (choline + creatine)/citrate ratio was elevated in cancer compared to the PZ and CG (P < 0.05). While a (choline + creatine)/citrate ratio above 0.68 was found to be a reliable indicator of cancer, elevated K(trans) was only a reliable cancer indicator in the diagnosis of individual patients. K(trans) and (choline + creatine)/citrate ratios in cancer were poorly correlated (Pearson r(2) = 0.07), and thus microvascular and metabolic abnormalities may have complementary value in cancer diagnosis. CONCLUSION: The combination of high-resolution spatio-vascular information from dynamic MRI and metabolic information from MRSI has excellent potential for improved localization and characterization of prostate cancer in a clinical setting. J. Magn. Reson. Imaging 2004;20:279-287. PMID- 15269955 TI - Improved three-dimensional free-breathing coronary magnetic resonance angiography using gadocoletic acid (B-22956) for intravascular contrast enhancement. AB - PURPOSE: To evaluate gadocoletic acid (B-22956), a gadolinium-based paramagnetic blood pool agent, for contrast-enhanced coronary magnetic resonance angiography (MRA) in a Phase I clinical trial, and to compare the findings with those obtained using a standard noncontrast T2 preparation sequence. MATERIALS AND METHODS: The left coronary system was imaged in 12 healthy volunteers before B 22956 application and 5 (N = 11) and 45 (N = 7) minutes after application of 0.075 mmol/kg of body weight (BW) of B-22956. Additionally, imaging of the right coronary system was performed 23 minutes after B-22956 application (N = 6). A three-dimensional gradient echo sequence with T2 preparation (precontrast) or inversion recovery (IR) pulse (postcontrast) with real-time navigator correction was used. Assessment of the left and right coronary systems was performed qualitatively (a 4-point visual score for image quality) and quantitatively in terms of signal-to-noise ratio (SNR), contrast-to-noise ratio (CNR), vessel sharpness, visible vessel length, maximal luminal diameter, and the number of visible side branches. RESULTS: Significant (P < 0.01) increases in SNR (+42%) and CNR (+86%) were noted five minutes after B-22956 application, compared to precontrast T2 preparation values. A significant increase in CNR (+40%, P < 0.05) was also noted 45 minutes postcontrast. Vessels (left anterior descending artery (LAD), left coronary circumflex (LCx), and right coronary artery (RCA)) were also significantly (P < 0.05) sharper on postcontrast images. Significant increases in vessel length were noted for the LAD (P < 0.05) and LCx and RCA (both P < 0.01), while significantly more side branches were noted for the LAD and RCA (both P < 0.05) when compared to precontrast T2 preparation values. CONCLUSION: The use of the intravascular contrast agent B-22956 substantially improves both objective and subjective parameters of image quality on high-resolution three-dimensional coronary MRA. The increase in SNR, CNR, and vessel sharpness minimizes current limitations of coronary artery visualization with high-resolution coronary MRA. PMID- 15269956 TI - Tumor enhancement using Mn-metalloporphyrin in mice: magnetic resonance imaging and histopathologic correlation. AB - PURPOSE: To determine the signal enhancement characteristics of tumors after administration of a metalloporphyrin derivative, HOP-9P (13, 17-bis (1 carboxypropionyl) carbamoylethyl-3, 8-bis (1-phenylpropyloxyethyl)-2, 7, 12, 18 tetramethyl-porphyrinato manganese (III)) and to determine whether HOP-9P is tumor-necrosis specific. MATERIALS AND METHODS: Ten C3H/He mice bearing a SCC VII tumor in the right flank were examined using T1-weighted conventional spin echo magnetic resonance (MR) imaging before contrast injection, and five minutes, one hour, and 24 hours after intravenous administration of 0.1 mmol/kg of HOP-9P. Following the imaging schedule, the mice were sacrificed, and sectioned in the same axial planes as the MR images. Based on an MR imaging-histopathologic correlation, mean signal intensities were measured, and signal-to-noise ratios (SNR) were calculated for both pure viable component and admixture of necrotic and viable component of the tumor. RESULTS: Mean SNR of the pure viable component peaked at one hour (35.0 +/- 3.8) and maintained that level until 24 hours (34.6 +/- 3.6). Mean SNR of the admixture of necrotic and viable component peaked at 24 hours (44.3 +/- 12.1). CONCLUSION: Although different enhancement patterns were seen between the pure viable component and the admixture of necrotic and viable component, HOP-9P enhanced both of the two components. PMID- 15269957 TI - Sensitivity of femoral orientation estimates to condylar surface and MR image plane location. AB - PURPOSE: To define the femoral anatomic region that provides the most reliable reference for measuring femoral orientation, from which patellofemoral and tibiofemoral orientation can be measured. MATERIALS AND METHODS: After a three dimensional image-based osteo-alignment procedure, two independent estimates of distal femoral orientation, the anterior (AFA) and posterior femoral angles (PFA), were acquired. These sets were comprised of 31 axial femoral orientation estimates each (obtained across a region 10 mm above to 20 mm below the patellar center) from 42 normal axial magnetic resonance (MR) image sets. An analysis of variance was conducted to determine the influence of image plane location, sex, and side on femoral orientation measures. RESULTS: The AFA presented greater variability across subjects (average SD = 3.13 degrees) as compared to the PFA (average SD = 1.78 degrees ). There were significant differences in the AFA based on image plane location and sex. In contrast, no differences were found between sides, sexes, or image plane locations for the PFA. CONCLUSION: In total, the posterior femoral condylar surface is more reliable than the anterior condylar surface for measuring femoral orientation and is best measured at least 8 mm below the patellar center. PMID- 15269958 TI - Image construction methods for phased array magnetic resonance imaging. AB - PURPOSE: To study image construction in phased array magnetic resonance imaging (MRI) systems from a statistical signal processing point of view. MATERIALS AND METHODS: Three new approaches for image combination with multiple coils are proposed: 1) one based on the singular value decomposition of the measurement matrix, which is asymptotically optimal in the signal-to-noise ratio sense; 2) one based on a maximum-likelihood formulation, incorporating a priori information on the coil sensitivities in a Bayesian manner; and 3) one based on a least squares formulation, which incorporates a smoothness constraint on the coil sensitivities. RESULTS: Numerical examples using synthetic and real data are presented to illustrate the performance of these new approaches. Results on the synthetic data show improvement in signal-to-error ratio, while results on the real data (a 4.7 T four-coil image of a cat spinal cord) show that the proposed methods can improve the SNR in the final image by up to 3 dB in the regions of interest compared to conventional sum-of-squares processing. CONCLUSION: It is demonstrated that phased array MRI reconstruction performance can be improved by the use of more elaborate statistical signal processing algorithms. PMID- 15269959 TI - Evaluation of specific absorption rate as a dosimeter of MRI-related implant heating. AB - PURPOSE: To compare the magnetic resonance imaging (MRI)-related heating per unit of whole body averaged specific absorption rate (SAR) of a conductive implant exposed to two different 1.5-Tesla/64 MHz MR systems. MATERIALS AND METHODS: Temperature changes at the electrode contacts of a deep brain stimulation lead were measured using fluoroptic thermometry. The leads were placed in a typical surgical implant configuration within a gel-filled phantom of the human head and torso. MRI was performed using two different transmit/receive body coils on two different generation 1.5-Tesla MR systems from the same manufacturer. Temperature changes were normalized to whole body averaged SAR values and compared between the two scanners. RESULTS: Depending on the landmark location, the normalized temperature change for the implant was significantly higher on one MR system compared to the other (P < 0.001). CONCLUSION: The findings revealed marked differences across two MR systems in the level of radiofrequency (RF)-induced temperature changes per unit of whole body SAR for a conductive implant. Thus, these data suggest that using SAR to guide MR safety recommendations for neurostimulation systems or other similar implants across different MR systems is unreliable and, therefore, potentially dangerous. Better, more universal, measures are required in order to ensure patient safety. PMID- 15269960 TI - Functional MRI using multiple receiver coils: BOLD signal changes and signal-to noise ratio for three-dimensional-PRESTO vs. single shot EPI in comparison to a standard quadrature head coil. AB - PURPOSE: To compare the performance of single shot echo planar imaging (SSEPI) with three-dimensional-multishot echo-planar imaging (EPI) based on principles-of echo-shifting-with-a-train-of-observations (PRESTO) in combination with a standard quadrature head coil and, as an alternative, a multiple receiver coil in intraoperative functional magnetic resonance imaging (fMRI). MATERIALS AND METHODS: Six healthy subjects underwent fMRI with visual stimulation using a SSEPI and a PRESTO-sequence with both coil systems. Statistical evaluation was done with a scanner-based post-processing software and SPM 99. The number of activated voxels in the visual cortex, the percent signal change between rest and activation, and finally the signal-to-noise ratio (SNR) during time course were measured and compared for both coil systems and both sequences, used in four different combinations. RESULTS: Blood oxygen level dependent (BOLD) signal changes were the lowest with PRESTO and standard head coil and the highest for SSEPI and phased array coil. For the latter combination, a significantly higher signal change and larger activation size was observed together with a better SNR. SSEPI yielded similar performance using both coils. CONCLUSION: SSEPI was superior due to its better SNR and a higher BOLD signal change in the defined settings, irrespective of the coil used. In a stereotactical setup the phased array coil can be used to generate fMRI data without loss of image quality. PMID- 15269961 TI - Myocardial delayed enhancement imaging using inversion recovery single-shot steady-state free precession: initial experience. AB - PURPOSE: To evaluate the feasibility of using an inversion recovery single-shot steady-state free precession (SS_SSFP) sequence for myocardial delayed enhancement (MDE) imaging, and to compare SS_SSFP with the conventional inversion recovery segmented fast gradient echo (IR_FGRE) technique. MATERIALS AND METHODS: Ten subjects (four volunteers and six patients with suspected or known coronary disease) were included in this study. All subjects were scanned with both IR_FGRE and SS_SSFP sequences 15-25 minutes after gadopentetate dimeglumine injection. Overall image quality, signal-to-noise ratios (SNRs), and contrast-to-noise ratios (CNRs) between the two techniques were compared. RESULTS: Compared to IR_FGRE, SS_SSFP exhibited adequate image quality (average scores = 3.8 for IR_FGRE and 3.9 for SS_SSFP) with much shorter acquisition time (14.4 seconds for IR_FGRE and 1.3 seconds for SS_SSFP). SS_SSFP images showed higher SNRs (P < 0.05) and less motion artifact from breathing. Enhanced myocardium was detected by both techniques in three patients, but the image sharpness is compromised in SS_SSFP images. CONCLUSION: SS_SSFP provides adequate image quality compared to IR_FGRE, while requiring a much shorter acquisition time. It is feasible to use SS_SSFP as an alternative method for MDE imaging, especially in patients who have difficulty with holding their breath. PMID- 15269962 TI - MRI of helium-3 gas in healthy lungs: posture related variations of alveolar size. AB - PURPOSE: To probe the variation of alveolar size in healthy lung tissue as a function of posture using diffusion-weighted helium-3 hyperpolarized gas imaging. MATERIALS AND METHODS: Measurements of the helium-3 apparent diffusion coefficient (ADC) were made on six healthy subjects. These were used to show the variation of alveolar size between the lowermost dependent regions of the lung compared to the uppermost regions of the lung in four postures: supine, prone, left-lateral decubitus, and right-lateral decubitus. RESULTS: The distribution of acinar size in the lungs was found to be heterogeneous, and influenced by lung orientation. In nearly all postures, the ADC was significantly higher in the non dependent uppermost regions of the lung compared to the dependent lowermost regions of the lung; the greatest variation was found in the left-lateral decubitus position. The difference in ADC between uppermost and lowermost regions was on average 0.012 cm(2)second(-1), which represents 20% of the average ADC value for the whole lung. A systematic decrease in ADC from the apex of the lung to the base was also found, which corresponds to an inherent gradient in alveolar size. CONCLUSION: The posture dependent variations in ADC were attributed to compression of the parenchyma under its own weight and the mass of the heart. PMID- 15269963 TI - Single-shot half-Fourier RARE sequence with ultra-short inter-echo spacing for lung imaging. AB - PURPOSE: To improve the image quality of pulmonary magnetic resonance (MR) imaging using an ultra-short inter-echo spacing half-Fourier single shot rapid acquisition with relaxation enhancement (USHA-RARE) sequence. MATERIALS AND METHODS: Pulmonary MR images were acquired by USHA-RARE sequence with various inter-echo spacings. The sequence parameters were as follows: repetition time (TR)/effective TE: infinite/39-41 msec; section thickness: 10 mm; acquisition matrix: 128 x 128; field of view: 450 x 450 mm. Inter-echo spacing varied (2.5 msec, 3.0 msec, 3.5 msec, 4.0 msec, 4.5 msec, 5.0 msec), and the respective phase encoding steps were 80, 77, 75, 74, 73, and 72. Signal-to-noise ratios (SNRs), the signal ratios between lung and fat (lung-to-fat ratio: LFRs), and the signal ratios between the lung and the serratus anterior muscle (lung-to-muscle ratio: LMRs) of each inter-echo spacing were calculated, and statistically evaluated. RESULTS: The SNRs at inter-echo spacings of < or = 3.0 msec were significantly higher than those > or = 4.0 msec (P < 0.05). The LFRs and LMRs at inter-echo spacing < or = 3.0 msec were significantly higher than those > or = 4.0 msec (P < 0.05). CONCLUSION: USHA-RARE sequence does improve signal intensity from the lung. PMID- 15269964 TI - Statistics-based approach for aneurysm volume measurements. AB - PURPOSE: To evaluate the ability of high-resolution MRA to monitor changes in intracranial aneurysm volume, and devise a highly reliable technique for obtaining these measurements. MATERIALS AND METHODS: To obtain a baseline estimate of the repeatability of MRA scans and validate the statistics-based technique for aneurysm volume measurement, multiple scans were obtained on individual subjects over a period of up to 1 year. These 3D MRA data sets were coregistered and then analyzed using the volumetric analysis of segmented data and the proposed statistical method. RESULTS: It was shown that high-resolution MRA provides highly repeatable data sets. Both methods used for the aneurysm volume measurements showed consistent results. However, the proposed statistical method had lower error and was much less sensitive to the choice of segmentation parameter than the volumetric analysis of segmented data. A change of 1 mm in the average radius of the aneurysm was detectable with the statistics-based technique. CONCLUSIONS: This study demonstrates that the statistical method of aneurysm volume measurement in high-resolution MRA allows reliable and accurate assessments of aneurysm volume changes. PMID- 15269965 TI - Infragenual cuff-compression reduces venous contamination in contrast-enhanced MR angiography of the calf. AB - PURPOSE: To reduce venous contamination at the calf level in three-dimensional contrast-enhanced MR angiography (CE-MRA) by applying continuous infragenual cuff compression. MATERIALS AND METHODS: Ten patients with clinically relevant peripheral arterial occlusive disease (PAOD) underwent dynamic three-dimensional CE-MRA of the calf. Six consecutive measurements were acquired with the first measurement serving as mask. Cuff-compression of 50 mmHg was attached below the knee. To allow intra-individual comparison, compression was applied unilaterally. The cuff was inflated three minutes before scanning and was continued throughout the MRA session. Venous contamination and arterial visualization scores were ranked using a five-point rating scale. Contrast-to-noise ratios (CNRs) of superficial enhancing calf-veins on the uncompressed and compressed calf sides were evaluated. An asymmetry index (AI) defined by CNR(mean) (uncompressed)/CNR(mean) (compressed) was introduced to describe the ratio in venous contrast agent supply between both sides quantitatively. RESULTS: Three dimensional CE-MRA of the calves demonstrated significantly lower superficial venous contamination scores (P < 0.004) and clearly improved arterial visualization (P < 0.009) on the compressed side. Additionally, AI values were larger than 1 (P < 0.02), indicating a higher contrast agent supply in the superficial veins on the uncompressed side. CONCLUSION: Infragenual cuff compression minimizes venous overlay in three-dimensional CE-MRA at calf level by reduction of contrast agent supply in the superficial veins. PMID- 15269966 TI - Recurrent excitation of granule cells with basal dendrites and low interneuron density and inhibitory postsynaptic current frequency in the dentate gyrus of macaque monkeys. AB - Temporal lobe epilepsy is often associated with pathological changes in the dentate gyrus, and such changes may be more common in humans than in some nonprimate species. To examine species-specific characteristics that might predispose the dentate gyrus to epileptogenic damage, we evaluated recurrent excitation of granule cells with and without basal dendrites in macaque monkeys, measured miniature inhibitory postsynaptic currents (mIPSCs) of granule cells in macaque monkeys and compared them to rats, and estimated the granule cell-to interneuron ratio in macaque monkeys and rats. In hippocampal slices from monkeys, whole-cell patch recording revealed antidromically evoked excitatory PSCs that were four times larger and inhibitory PSCs that were over two times larger in granule cells with basal dendrites than without. These findings suggest that granule cells with basal dendrites receive more recurrent excitation and, to a lesser degree, more recurrent inhibition. Miniature IPSC amplitude was slightly larger in monkey granule cells with basal dendrites than in those without, but mIPSC frequency was similar and only 26% of that reported for rats. In situ hybridization for glutamic acid decarboxylase and immunocytochemistry for somatostatin, parvalbumin, and neuronal nuclei revealed interneuron proportions and distributions in monkeys that were similar to those reported for rats. However, the interneuron-to-granule cell ratio was lower in monkeys (1:28) than in rats (1:11). These findings suggest that in the primate dentate gyrus, recurrent excitation is enhanced and inhibition is reduced compared with rodents. These primate characteristics may contribute to the susceptibility of the human dentate gyrus to epileptogenic injuries. PMID- 15269967 TI - Morphogenesis and regionalization of the medaka embryonic brain. AB - We examined the morphogenesis and regionalization of the embryonic brain of an acanthopterygian teleost, medaka (Oryzias latipes), by in situ hybridization using 14 gene probes. We compared our results with previous studies in other vertebrates, particularly zebrafish, an ostariophysan teleost. During the early development of the medaka neural rod, three initial brain vesicles arose: the anterior brain vesicle, which later developed into the telencephalon and rostral diencephalon; the intermediate brain vesicle, which later developed into the caudal diencephalon, mesencephalon, and metencephalon; and the posterior brain vesicle, which later developed into the myelencephalon. In the late neural rod, the rostral brain bent ventrally and the axis of the brain had a marked curvature at the diencephalon. In the final stage of the neural rod, ventricles began to develop, transforming the neural rod into the neural tube. In situ hybridization revealed that the brain can be divided into three longitudinal zones (dorsal, intermediate, and ventral) and many transverse subdivisions, on the basis of molecular expression patterns. The telencephalon was subdivided into two transverse domains. Our results support the basic concept of neuromeric models, including the prosomeric model, which suggests the existence of a conserved organization of all vertebrate neural tubes. Our results also show that brain development in medaka differs from that reported in other vertebrates, including zebrafish, in gene-expression patterns in the telencephalon, in brain vesicle formation, and in developmental speed. Developmental and genetic programs for brain development may be somewhat different even among teleosts. PMID- 15269968 TI - Axonogenesis in the medaka embryonic brain. AB - In order to know the general pattern of axonogenesis in vertebrates, we examined axonogenesis in the embryonic brain of a teleost fish, medaka (Oryzias latipes), and the results were compared with previous studies in zebrafish and mouse. The axons and somata were stained immunocytochemically using antibodies to a cell surface marker (HNK-1) and acetylated tubulin and visualized by retrograde and anterograde labeling with a lipophilic dye. The fiber systems developed correlating with the organization of the longitudinal and transverse subdivisions of the embryonic brain. The first axons extended from the synencephalic tegmentum, forming the first fiber tract (fasciculus longitudinalis medialis) in the ventral longitudinal zone of the neural rod, 38 hours after fertilization. In the neural tube, throughout the entire brain two pairs of longitudinal fiber systems, one ventral series and one dorsal or intermediate series, and four pairs of transverse fiber tracts in the rostral brain were formed sequentially during the first 16 hours of axon production. In one of the dorsal longitudinal tracts, its branch retracted and disappeared at later stages. One of the transverse tracts was found to course in the telencephalon and hypothalamus. The overall pattern of the longitudinal fiber systems in medaka brain is similar to that in mouse, but apparently different from that in zebrafish. We propose that a ventral tract reported in zebrafish partially belongs to the dorsal fiber system, and that the longitudinal fiber systems in all vertebrate brains pass through a common layout defined by conserved genetic and developmental programs. PMID- 15269969 TI - Bipolar cells of the mouse retina: a gene gun, morphological study. AB - One of the key elements concerning our understanding of the organization of the mouse retina is the complete classification of the various types of bipolar cells. With the present study, we tried to contribute to this important issue. Unfortunately, most of the antibodies that stain specifically bipolar cells in the retina of other mammals hardly work for the retina of the mouse. We succeeded in overcoming this limitation by using a relatively novel technique based on the gene gun transfer of fluorescent dyes to cells. Hence, we were able to stain a considerable number of bipolar cells that could be characterized according to morphological and comparative criteria. We also performed a complete morphometric analysis of a subset of bipolar cells stained by anti-neurokinin-3 receptor antibodies. We found nine types of cone bipolar cells and one type of rod bipolar cell; these data are consistent with the findings of previous studies on the retinas of other mammals, such as rabbits, rats, and monkeys and with a recent study based on the mouse retina (Ghosh et al. [2004] J Comp Neurol 469:70-82). Our results also confirm the existence of a common structural similarity among mammalian retinas. It remains to be elucidated what is exactly the functional role of the various types of cone bipolar cells and what is the specific contribution they provide to the perception of a given visual stimulus. Most probably, each bipolar cell type constitutes a specialized channel for the computation of a selected component of the visual stimulus. More complex signal coding, involving the coordinated activity of various types of bipolar cells, could also be postulated, as it has been shown for ganglion cells (Meister [1996] Proc Natl Acad Sci U S A 93:609-614). PMID- 15269970 TI - Development of the human motor-related thalamic nuclei during the first half of gestation, with special emphasis on GABAergic circuits. AB - This study analyzed the expression of differentiation markers (Calbindin D28K: CaBP; parvalbumin: PARV; calretinin: CalR), gamma-aminobutyric acid (GABA) markers (GABA, glutamic acid decarboxylases: GAD65, GAD67; and GABA transporters: GAT1, GAT3), and other markers (neurotensin: NT, and neurofilament-specific protein: SMI32) in the human thalamus at 8-23 gestation weeks (g.w.), focusing on the motor-related nuclei. From 8-13 g.w. mainly CaBP was expressed in the cells while fiber bundles traversing the thalamus in addition to CaBP expressed all GABA markers except GAD67. CaBP and PARV expression patterns in different nuclei changed over the time course studied, whereas NT was expressed consistently along the anterior-lateral curvature of the thalamus. CalR and SMI were detectable at 23 g.w. in the ventral parts of the dorsal thalamus. Most remarkably, punctate GAD65 immunoreactivity in the neuropil was confined to the nigro- and pallidothalamic afferent receiving nuclei from 16 to about 21 g.w., overlapping with that of CaBP in some of these nuclei (subdivisions of the ventral anterior and mediodorsal nuclei) and with PARV in others (centromedian nucleus). During this period, GAD65 immunoreactivity can be considered a marker of the basal ganglia afferent receiving territory in the motor thalamus. GAD67-positive local circuit neurons were first detected at 12-13 g.w. in the thalamic nuclei outside the basal ganglia afferent receiving territory. In the ventral anterior and centromedian nuclei, GAD-containing local circuit neurons were not conspicuous even at 22-23 g.w. The cells of the reticular nucleus expressed GAD67 and PARV from 12 g.w. on starting in the lateral-posterior regions. By 23 g.w., both markers were expressed in about two-thirds of the nucleus except for its most medial-anterior part. The results imply spatially and temporally differential expression of GABA and differentiation markers in the developing human thalamus. PMID- 15269971 TI - Subventricular zone-derived neuronal progenitors migrate into the subcortical forebrain of postnatal mice. AB - The presence of a germinal layer and the capacity to generate neurons, once thought restricted to the embryonic brain, persists in the forebrain of both postnatal and adult mammals. The two regions in which this phenomenon has been extensively demonstrated are the hippocampal dentate gyrus and the lateral ventricle subventricular zone (SVZ). SVZ-derived cells migrate along the rostral migratory stream into the olfactory bulb, where they differentiate into local interneurons. In this study, using tracer injections into the SVZ at different postnatal ages, we investigated the occurrence of secondary migratory pathways in the mouse subcortical forebrain. During the course of the first week postnatal, in addition to the well-characterized rostral migratory stream, SVZ-derived progenitors migrate in a ventral migratory mass across the nucleus accumbens into the basal forebrain and along a ventrocaudal migratory stream originating at the elbow between the vertical and horizontal limbs of the rostral migratory stream. These cells give rise to granule neurons in the Islands of Calleja and olfactory tubercle pyramidal layer, respectively. In adult, a very small number of cells continue to migrate along the ventrocaudal migratory stream, whereas no migration was observed across the nucleus accumbens. These data demonstrate that in early postnatal and, to a minor extent in adult mice, SVZ-derived cells contribute new neurons to the subcortical forebrain. PMID- 15269972 TI - Differentiation of the midbrain dopaminergic pathways during mouse development. AB - Dopaminergic (DA) neurons in the substantia nigra (SN) and ventral tegmental area (VTA) of the midbrain project to the dorsolateral caudate/putamen and to the ventromedially located nucleus accumbens, respectively, establishing the mesostriatal and the mesolimbic pathways. Disruptions in this system have been implicated in Parkinson's disease, drug addiction, schizophrenia, and attention deficit hyperactivity disorder. However, progress in our understanding has been hindered by a lack of knowledge of how these pathways develop. In this study, different retrograde tracers, placed into the dorsolateral caudate/putamen and the nucleus accumbens, were used to analyze the development of the dopaminergic pathways. In embryonic day 15 mouse embryos, both SN and VTA neurons, as well as their fibers, were doubly labeled by striatal injections into the dorsolateral and ventromedial striatum. However, by birth, the SN DA neurons were labeled exclusively by DiA placed in the dorsolateral striatum, and the VTA DA neurons were labeled only by DiI injected into the ventromedial striatum. These data suggest that initial projections from midbrain DA neurons target nonspecifically to both the dorsolateral striatum and the nucleus accumbens. Later during development, the separate mesostriatal and mesolimbic pathways differentiate through the selective elimination of mistargeted collaterals. PMID- 15269974 TI - [Recent progress in hair follicle research]. PMID- 15269975 TI - [Skin-like structure generated from implantation of hair follicle bulb cells into collagen/chitosan porous scaffolds in vitro]. AB - OBJECTIVE: To observe the skin regeneration after hair follicle bulb cells were implanted into collagen/chitosan porous scaffolds in vitro. METHODS: The cultured dorsal hair follicle bulb cells of 4d-old C57BL/6J mice were implanted into collagen/chitosan porous scaffolds in vitro. The skin regeneration was observed. RESULT: The skin-like structure was formed on the collagen/chitosan porous scaffolds where were cultured the hair follicle bulb cells before 4th passages. CONCLUSION: The skin-like structure is generated in vitro when early passages of cultured hair bulb cells are implanted into collagen/chitosan porous scaffolds. PMID- 15269976 TI - [Hair follicle regeneration after implantation of hair follicle cells into subcutis of nude mice]. AB - OBJECTIVE: To observe the hair follicle regeneration after implantation of hair follicle cells into the subcutis of nude mice. METHODS: The cultured hair papilla cells,dermal sheath cells and fibroblast of human scalp were mixed with the cells of hair follicle epithelium in different ratio, and then implanted into the subcutis of nude mice. The regeneration of hair follicle was observed. RESULT: The hair follicle-like structure was formed in cluster where the cultured hair follicle epithelium cells were mixed with hair papilla cells. But no hair follicles were formed where the hair follicle epithelium was implanted with dermal sheath cells or fibroblasts. CONCLUSION: The hair follicle-like structure is generated in vivo when the mixed cells of early passages cultured hair papilla cells with hair follicle epithelium are implanted into the subcutis of nude mice. PMID- 15269977 TI - [Protective effects of minocycline against hair follicle damage induced by cytosine arabinoside in vitro]. AB - OBJECTIVE: To investigate the protective effects of minocycline against hair follicle damage induced by cytosine arabinoside (Ara-c). METHODS: An in vitro organ culture of mouse vibrissa follicles was used and different concentrations of Ara-c and minocycline were added in the culture media. The total growth length, growth speed and growth period of hair were observed with invert microscopy and the survival of hair bulb cells was measured by MTT method. RESULT: Minocycline (0.3 x 10(-6) approximately 10(-5) mol/L) improved hair follicle total growth length, growth speed and hair growth period and also improved survival of hair bulb cells in vitro organ culture, which were inhibited by Ara-c. CONCLUSION: Minocycline can protect hair follicle directly from damage induced by Ara-c. PMID- 15269978 TI - [Expressions of bFGF, ET-1 and SCF in dermal papilla cells and the relation to their biological properties]. AB - OBJECTIVE: To investigate the expression of bFGF, ET-1 and SCF in different passages of cultured dermal papilla cells (DPC), and their possible effect on biological behaviour of DPC. METHODS: The expression of bFGF, ET-1 and SCF in different passages of cultured DPC was detected by immunocytochemistry and in situ hybridization. RESULT: The expression of ET-1 and SCF in early passages of cultured DPC was stronger, but became negative in late passages (>6 passages). The stronger the expression of ET-1 and SCF in DPC, the higher ability of DPC to induce hair follicle regeneration. CONCLUSION: The expression strength of ET-1 and SCF is related to the ability of DPC inducing hair follicle regeneration. PMID- 15269979 TI - [Development of a system for quick screening of efficient HBx-siRNA]. AB - OBJECTIVE: To develop a system for quick screening of efficient siRNA targeted HBx mRNA. METHODS: Using recombination DNA technique, the fusion expression plasmid of HBx and EGFP was constructed, and siRNA expression cassettes (SECs) containing U6+1, H1 or tRNA(Val )promoter were prepared via one-step overlapping extension PCR. By co-transfection with recombinant plasmid and SECs into AD293 cell, the inhibition effects on the transient expression of HBx-EGFP fusion protein were analyzed by FACS and semi-quantitated RT-PCR analysis. RESULT: (1)HBx-EGFP fusion protein expression plasmid pHBx-EGFP was constructed successfully, which expressed green fluorescence in cell mainly located at plasma or the periphery of nucleus in granules. (2) Co-transfection with recombinant plasmid and SECs into AD293 cells resulted in inhibition of HBx-EGFP expression. SEC-siHBx388 showed significant inhibition effect on HBx-EGFP expression compared with SEC-siHBx271, indicating that siHBx388 is effective siRNA site and could be screened out with our screening system. In addition,the results of that U6+1-, tRNA(Val) and H1-siHBx388 reduced HBx-EGFP expression by 21.7%, 12.9% and 12.4% of control respectively indicated that both tRNAVal and H1 promoter was high efficient in driving effect of siHBx388. CONCLUSION: Combination of the HBx expression carrying reporter gene and PCR-based multi promoter SECs may develop a useful system to be applied in identification of optimal HBx- siRNA and its matching promoter. PMID- 15269980 TI - [Effects of physical and chemical properties of polymeric nerve conduit on peripheral neuranagenesis]. AB - OBJECTIVE: To evaluate the effects of degradation time and permeability of polymeric conduits on nerve regeneration. METHODS: After establishment of rat models with over 10 mm gap of sciatic nerve in right hind legs,four kinds of poly (ester, carbonate) nerve conduits was used to bridge the gaps and one group without conduit in gaps was used as control. The nerve regeneration and conduit degradation were examined both macroscopically and microscopically. The contraction of the muscle controlled by regenerated nerve was measured electrophysiologically at 4, 12 and 20 weeks after the operation. RESULT: Biodegradation time of nerve conduits in vivo was as fellows: 12 weeks in group A,4 weeks in group B and group C,and 20 weeks in group D,respectively. The histological quality of regenerative sciatic neurofibra in group A was the best among all groups (Mean rank 53.17, 38.83, 26. 60, 49.17 and 20.23,P<0.005), but the inflammatory reaction in group A was only less than that in group D and more than that in the other groups (Mean rank 45.87, 36.27, 34. 83, 51.63 and 21.4,P=0.001). The responsive rates of tibialis anterior muscle for electric stimulation in group A, B, C and D were 93.33%, 60%,20% and 73. 33%, respectively (P<0.005). CONCLUSION: Absorbable conduits with relatively good permeability and appropriate middle degrade time improve nerve regeneration and renovate function. PMID- 15269981 TI - [Impairment of IRS-2 signaling in rat insulinoma INS-1 cells by nelfinavir]. AB - OBJECTIVE: To investigate whether HIV-1 protease inhibitor nelfinavir alters the insulin stimulated phosphorylation of insulin signaling parameters in rat insulinoma INS-1 cells. METHODS: INS-1 cells were incubated with nelfinavir for 48 h and stimulated with 100 nmol/L insulin for 2 min. Immunoprecipitation and Western blot analysis of the insulin stimulated insulin receptor substrate (IRS) 1,-2 and Akt-Thr(308) phosphorylation were performed on cell lysates. Cytotoxic effects of nelfinavir were measured by cell count with trypan blue and MTT reduction test. RESULT: Nelfinavir decreased insulin stimulated phosphorylation of IRS-2 and Akt-Thr(308) in a dose-dependent manner; for 10 micromol/L of nelfinavir, the decrease was 52% and 55%, respectively. CONCLUSION: Treatment with nelfinavir might impair IRS-2-mediated signaling in pancreatic beta cells. PMID- 15269982 TI - [Interaction between polysaccharides and interferon-gamma using an improved ELISA approach]. AB - OBJECTIVE: To establish an ELISA approach to study the interaction of polysaccharides with cytokine in vitro. METHODS: The heparin BSA complexes (HBC) were synthesized with a chemistry method and separated using a 1 X 90 cm column of Separose 4B. After identification of the complex via SDS-PAGE,the wells of ELISA plates were coated with HBC and the interaction of HBC with interferon gamma (IFN-gamma) was detected. The effects of heparin, low molecular weight heparin (LMW heparin), chondroitin sulfate (CS), hyaluronic acid (HA) and carrageenans on the binding of HBC to IFN-gamma were tested in this system. RESULT: Human recombinant IFN-gamma bound to heparin in a concentration dependent manner, the binding of IFN-gamma to HBC was detected at the concentration of 0.25 ng, and saturated at around 2 ng. Free heparin, LMW heparin, CS,HA and carrageenans competed for the binding of IFN-gamma to HBC with significant different ability. The IC(50)concentrations of heparin and LMW heparin were 2.40 microg/ml and 18.60 microg/ml respectively. CONCLUSION: IFN gamma is a cytokine with high binding affinity to heparin and carrageenans family but poor to CS-A and CS-C. ELISA is a simple, sensitive approach to detect the interaction of polysaccharides with cytokine in vitro. PMID- 15269983 TI - [Role of area postrema of medulla in regulation of rat cardiovascular activity]. AB - OBJECTIVE: To explore the role of area postrema (AP) of medulla in control of cardiovascular functions in rat. METHODS: (1) Sprague Dawley rats were anaesthetized with urethane and pentobarbital and the AP was stimulated by electrical stimulus with intensity of 0.1 mA and frequencies ranged 10 approximate, equals 80 Hz. (2) Excitatory amino acid L-glutamate (L- Glu, 0.1 approximate, equals 0.5 mol/L) was microinjected into AP in urethane anaesthetized rats and the changes of mean arterial pressure (MAP) and heart rate (HR) were recorded. RESULT: (1) When the frequencies of 10 Hz, 20 Hz and 40 Hz were used, the electrical stimulation of AP caused decrease of MAP and HR (P<0.001),while the electrical stimulation with the frequencies of 60 Hz and 80 Hz caused an increase of MAP (P<0.05) but a decrease of HR (P<0.001). (2) Microinjection of L-Glu at 0.1 mol/L had no effect on MAP and HR (P>0.05), but it decreased MAP and HR at 0.15 mol/L (P<0.001, P<0.05). The MAP was increased (P<0.001) but HR (P<0.05) was decreased at the concentrations of 0.2 mol/L and 0.5 mol/L, respectively. CONCLUSION: Alterations of MAP and HR induced by electrical or chemical stimulation on AP of medulla are related to the frequency of electrical stimulation or concentration of L-Glu. PMID- 15269984 TI - [Expression of sialylated carbohydrate antigens and nm23-H1 gene in prognosis of breast cancer]. AB - OBJECTIVE: To assess the significance of expression of sialylated carbohydrate antigens and nm23-H1 gene in metastasis and prognosis of breast cancer. METHODS: Tissue specimens from 102 cases of primary breast cancer were stained with antibodies against sialyl Lewis A (SleA) and salyl Lewis X (SleX), and nm23-H1 proteins by immunohistochemical methods. RESULT: Of the 102 cases, the positive cases of SleA and SleX were 24.5% (25/102) and 59.89% (61/102),respectively; the reduced expression of nm23-H1 was showed in 37.3% (38/102) of the cases. The positive expression of SleX and the reduced expression of nm23-H1 gene were significantly associated with lymph node involvement. Among the 100 patients who underwent curative surgery, the disease-free survival rate was significantly correlated with nm23-H1 and SleX expression, respectively,but not with SleA expression. In multivariate analysis using Cox regression model, combination assay of nm23 H1 and SleX expression emerged as independent prognostic factors. CONCLUSION: These results suggest that nm23-H1 gene and SleX may be involved in the metastatic process in human breast cancer, and immunohistochemical detection of SleX and nm23-H1 may be used as a biologic marker of prognosis. PMID- 15269985 TI - [Effect of STAT3 phosphorylation and p53 expression on human epidermal non melanoma cutaneous tumors]. AB - OBJECTIVE: To investigate the effect of stat3 phosphorylation and p53 expression on human epidermal non-melanoma cutaneous tumours. METHODS: Immunohistochemistry technique was employed to measure the expression of p-stat3 and p53 protein in skin tissue from 30 cases of skin squamous cell carcinoma (SCC), 20 cases of basal cell carcinoma (BCC), 20 cases of seborrhoeic keratosis (SK) and 20 normal subjects. RESULT: (1) p-stat3 protein was abnormally increased in SCC and BCC as compared with normal skin and SK. Expression of p-stat3 in SCC was also significantly higher than that in BCC. (2) Expression of p-stat3 was higher in poorly-differentiated cancers than that in well-differentiated cancers in SCC. The positive rate of p-stat3 expression was correlated with the depth of tumor invasion, but not with tumor size. (3) There was no p53 protein expression on normal skin and SK, it was significantly upregulated in SCC and BCC. In SCC, the intensity of p53 expression was associated with tumor differentiation. There was no correlation between the positive rate of p53 expression and the depth of tumor invasion, whereas the positive rate of p53 expression was correlated with the sun exposure area. (4) There existed positive correlation between the expression intensity of p-stat3 and p53 in SCC (r=0.641, P<0.05). CONCLUSION: (1) The overexpression of p-stat3 may play an important role in the development of epidermal tumors. (2) The abnormal activation of stat3 may be related to metastatic potentials in SCC. (3) Both p53 gene and stat3 may contribute to the pathogenesis of skin SCC. PMID- 15269986 TI - [Experimental induction of posterior vitreous detachment in rabbits with hyaluronidase and perfluoroethane (C2F6)]. AB - OBJECTIVE: To evaluate the experimental induction of posterior vitreous detachment (PVD) by intravitreous injection of hyaluronidase and perfluoroethane (C(2)F(6)). METHODS: Fifteen rabbits (30 eyes) were divided into 3 experimental groups,the contralateral eyes in same animals served as the controls. Eyes in group A and B were received two vitreous injections of 15 IU of hyaluronidase at an interval of 5 d. The eyes in group C and all control eyes were injected with balanced salt solution (BSS). Seven days after injection, the experimental eyes in group A and C were received 0.5 ml of Fifteen rabbits (30 eyes) were divided into 3 experimental groups, the contralateral eyes in same animals served as the controls. Eyes in group A and B were received two vitreous injections of 15 IU of hyaluronidase at an interval of 5 d. The eyes in group C and all control eyes were injected with balanced salt solution (BSS). Seven days after injection,the experimental eyes in group A and C were received 0.5 ml of C(2)F(6) injection. The ocular and retinal signs were examined for 8 following weeks and then killed for histological examination. RESULT: Five eyes in group A (100.0%) showed complete separation of the vitreous cortex from the retina (PVD), three eyes in group B(60.0%) showed partial PVD, and no PVD was detected in group C and all control eyes. On electroretinogram no significant difference was found in amplitude and latency of a-(or b-) wave in both experimental and control eyes, between before and after experiments. No evidence of ocular or retinal toxicity was revealed by light or scanning electronic microscopy in all eyes. CONCLUSION: Vitreous injection of hyaluronidase combined with perfluoroethane, as a safety method, can induce posterior vitreous detachment without mechanical vitrectomy. PMID- 15269987 TI - [Relationship of serum levels of HGF and MMP-9 with disease activity of patients with systemic lupus erythematosus]. AB - OBJECTIVE: To determine the relationship of serum hepatocyte growth factor (HGF) and matrix metalloproteinase-9 (MMP-9) levels with the disease activity of systemic lupus erythematosus (SLE). METHODS: Serum levels of HGF and MMP-9 were measured by ELISA in 36 patients with SLE and 30 healthy subjects as controls. RESULT: (1)Significantly increased serum level of HGF was found in SLE patients as compared with that in healthy controls (P<0.001), but serum level of MMP-9 in SLE patients decreased (P<0.001). Serum level of HGF was significantly decreased after treatment in SLE patients (P<0.05), but serum level of MMP-9 was increased (P<0.05). (2)Serum level of HGF was markedly higher in patients with active disease (24 cases) than those with inactive disease (P<0.05), but serum level of MMP-9 was lower (P<0.05). (3)Significantly increased serum level of HGF was found in patients with renal damage (16 cases) than those without (P<0.001), but serum level of MMP-9 was lower in patients with renal damage than those without (P<0.005). (4)Serum level of HGF was higher in patients with arthritis (23 cases) than those without (P<0.01), but serum level of MMP-9 had no significant difference in two groups (P>0.05). (5)The area of ROC curve was 0.707 and the sensitivity was 66.7% if serum level of HGF was served as diagnostic standard. The area of ROC curve was 0.984 and the sensitivity was 97.2% if serum level of MMP-9 was served as diagnostic standard. The sensitivity was 66.7% (24/36) if the two markers were examined simultaneously. CONCLUSION: The data suggest that HGF and MMP-9 could be involved in the pathogenesis of SLE, and serum levels of HGF and MMP-9 be used as markers to monitor disease activity, renal damage, disease progression and amelioration in SLE. PMID- 15269988 TI - [Study of muscle pressure exerted on deciduous normal occlusion]. AB - OBJECTIVE: To measure the muscle pressure exerted on the deciduous normal occlusion and to explore the relationship between the denture,occlusion, skeleton and muscle pressure. METHODS: Thirty volunteers of deciduous normal occlusion were included in the study. The muscle pressure of natural head posture (NHP) was measured at rest and during swallowing by a PC real-time measuring system and the data were analyzed by statistical software SAS. RESULT: The forces from lips, cheeks and tongue at rest were 0 approximate, equals 1.47 g/cm(2), while the pressure were increased to 9.60 approximate, equals 20.13 g/cm(2) during swallowing. The lip pressure was higher than the lingual side but there was no statistical difference at rest. The boys had higher pressure than girls,but there was also no statistical difference at rest. The pressure of normal occlusion was related to sex at the position of the upper labial incisor and the side of the dental arch during swallowing. The pressure of both sides of the dental arch increased significantly during swallowing. CONCLUSION: The deciduous dentition is in a state of dynamic equilibrium. This equilibrium may result in a special facial morphology. PMID- 15269989 TI - [Chromosome subtelomeric analysis by FISH in patients with mental retardation]. AB - OBJECTIVE: To assess subtelomeric chromosome anomalies in patients with idiopathic mental retardation (MR). METHODS: Subtelomeric screening was performed in 46 patients with undiagnosed mental retardation. The patients were selected based on the following criteria: (1) MR with two or more of the following conditions: dysmorphic features, prenatal growth retardation, postnatal growth abnormalities, a suggestive family history; (2) chromosome karyotype at the level >450 bands being normal; (3) exclusion of other identified genetic or environmental diagnosis. Fluorescence in situ hybridization (FISH) was performed using ToTelVysion DNA probes. Abnormal findings were confirmed by FISH with a specific subtelomeric probes and family studies were carried out to determine its inheritance. RESULT: Clinically significant aberrations were detected in two cases with 6q and 2q terminal microdeletion. The deletion in one case was inherited from a similarly affected father. Subtle chromosomal subtelomeric abnormalities occurred with a frequency of 7.6% in children with moderate to severe mental retardation and of 3.0% in the children with mild retardation. CONCLUSION: The results suggest that cryptic abnormalities of the ends of chromosomes might represent a significant cause of mental retardation, and screening for subtelomeric rearrangements might be warranted in children with unexplained mental retardation. PMID- 15269990 TI - [Prostaglandin E1 reduces reperfusion injury of myocardium by inhibiting cytokines production during cardiac surgery]. AB - OBJECTIVE: To investigate the effects of prostaglandin E1 (PGE1) on the cytokines (IL-6, IL-8, IL-10) levels and ischemic -reperfusion injury of myocardium during cardiac surgery. METHODS: A total of 30 patients undergoing cardiac surgery (mitral valve replacement) under extracorporeal circulation were randomized into two groups: PGE1 group (receiving 0.04 microg.kg(-1). min(-1) of Lipo-PGE1 from the beginning of surgery to the end of study, n=15) and control group (no PGE1 given, n=15). Levels of serum IL-6, IL-8 and IL-10 were measured by enzyme-linked immunosorbent assays (ELISA). Isoenzyme of creatine kinase with muscle and brain subunits(CKMB) and troponin-T (cTn-T) were measured by ultraviolet absorption spectrophotometry method and enzyme immunoassay on 5 time-points during the study. RESULT: In both groups serum IL-6, IL-8, CK-MB and cTn-T levels increased significantly after aorta declamping (especially from 2 h after aorta declamping) compared with preoperative levels (P<0.05).However,the elevations of these cytokines and enzymes were more prominent in the control group than in the PGE1 group (P<0.05). Serum IL-10 concentration increased significantly from 2 h after aorta declamping compared with preoperative value (P<0.05); but there were no differences between the two groups. IL-6 and IL-8 levels were correlated with CK MB and cTn-T concentrations (r2=0.40, r2=0.38 P>0.05 and r2=0.56, r2=0.14; P>0.05, respectively). CONCLUSION: During cardiac surgery (mitral valve replacement) PGE1 may suppressed the production of IL-6, IL-8 but not IL-10, which may be related to its myocardial protection effect. PMID- 15269991 TI - [Minilaparotomy approach for curative resection of colorectal cancer]. AB - OBJECTIVE: To evaluate the feasibility and safety of a minilaparotomy approach for curative resection of colorectal cancer in comparison with the conventional laparotomy. METHODS: Seventy-eight patients underwent radical resection for rectal cancer with minilaparotomy during April 2001 to December 2002. The minilaparotomy involved complete resection and a skin incision 2 cm above the link line of left anterior superior iliac spine to pubic symphysis and was about 7-10 cm in length. Another 86 patients who served as control group underwent a similar resection with a conventional laparotomy during the same period. RESULT: The minilaparotomy approach was successful in all 78 patients. The general status of patients, operative types and histopathological features of tumor were similar in the two groups (P>0.05). Operative blood loss in control group was greater (P<0.001), whereas incision length in minilaparotomy group was significantly shorter than that in conventional laparotomy (9.38 cm compared with 17.32 cm). The operative time, analgesia requirement, first passing flatus,first oral fluids and postoperative hospital stay were significantly shorter in the minilaparotomy group (P<0. 001). In an average 25.4-month follow-up, there were no tumor recurrences in the minilaparotomy group. CONCLUSION: A minilaparotomy approach for curative resection of rectal cancer may be an ideal alternative approach to conventional laparotomy. PMID- 15269992 TI - [Effects of calcium on ability of learning and memory in rats exposed to low level lead before and after birth]. AB - OBJECTIVE: To investigate the effects of calcium on learning and memory ability of rats exposed to low level lead before and after birth. METHODS: Wistar dam rats were randomly divided into normal group, lead-contaminated group and lead with Ca group. Corresponding food and water were given to pregnant rats from d 15 of gestation and to young rats till 7 w after birth. The weight of brain and hippocampus, blood lead content, serum calcium content, learning and memory ability of young rats were tested. RESULT: The blood lead concentrations: lead contaminated group was the highest, lead with Ca group the second and normal group the lowest. Serum calcium concentrations: normal group and lead with Ca group were both higher than lead contaminated group. Ability of learning and memory: lead with Ca group was better than lead-contaminated group, but poorer than normal group. No differences were found upon the weight of brain and hippocampus in all groups. CONCLUSION: A minilaparotomy approach for curative resection of rectal cancer may be an ideal alternative approach to conventional laparotomy. PMID- 15269993 TI - [Role of transthoracic echocardiography in transcatheter closure for atrial septal defect in children]. PMID- 15269994 TI - [Overbite correction with edgewise technique in adult patient]. PMID- 15269995 TI - [Research progress of bone morphogenetic proteins expressed in hair follicles]. PMID- 15269996 TI - Effects of pioglitazone versus rosiglitazone on lipoprotein subclasses. PMID- 15269997 TI - The question is what is the best retrograde root filling material not whether we should move from amalgam. PMID- 15269998 TI - Training our future rural medical workforce. PMID- 15269999 TI - Cancer in adolescents and young adults: treatment and outcome in Victoria. PMID- 15270000 TI - [Paroxystic alcohol intoxications]. AB - Paroxystic alcohol intoxications are classically defined by the alternation of acute and massive intoxications and abstinence periods but occur mostly in chronically drinking patients. Loss of control, sensation seeking and impulsivity are common features of these disorders that are often associated with multiple drug use. They also seem to be characterized by underlying biological and genetic risk factors and antisocial personality disorder. PMID- 15270001 TI - [Etoricoxib (Arcoxia)]. AB - Etoricoxib (Arcoxia) is a novel non steroidal anti-inflammatory drug (NSAID) that selectively inhibits the inducible form of cyclo-oxygenase (COX), COX-2. Etoricoxib has a higher COX-1/COX-2 selectivity ratio than the other COX-2 selective NSAIDs as rofecoxib, valdecoxib or celecoxib. Tablets of 60, 90 and 120 mg are available. The recommended dosage of etoricoxib is 60 mg/day for osteoarthritis, 90 mg/day for rheumatoid arthritis and 120 mg/day for acute gouty arthritis. Etoricoxib's efficacy has been widely studied in comparative studies, showing the same efficacy as non-COX-2 selective NSAID, with fewer gastro intestinal adverse effects. PMID- 15270002 TI - Diversity and species composition of sand flies (Diptera: Psychodidae) in a Venezuelan urban focus of cutaneous leishmaniasis. AB - The present study examined the spatial and temporal abundance and diversity of phlebotomine sand flies in an area of Venezuela that is an ancient focus of leishmaniasis. The study was conducted in 6 stations in urban localities in Trujillo City, located in northwestern Venezuela (9 degrees 22' 24" N, 70 degrees 26' 08" W), which is located in a mountain range in the Andean ecoregion (altitude = 600-1,010 m). During 1995-99, entomological surveys were conducted after and before the rainy season. Shannon light traps were operated from 1800 to 2000 h in peridomestic site trap locations. Twelve species were captured, and Lutzomyia youngi, L. ovallesi, L. scorzai, L. gomezi, L. lichyi, and L. shannoni occurred at all localities in each year. The abundance of these species showed low variation over time but high variation between localities. The Sorensen similarity index, used to compare diversity between years within each locality, ranged from 0.60 at Carmona to 0.84 at La Hacienda. Sand fly communities exhibited annual variation in species richness and diversity. Variations were affected more by changes in species abundance than by changes in species composition. Lutzomyia ovallesi, L. lichyi, and L. scorzai had the highest coefficient of variation between years (63, 38, and 23%, respectively). PMID- 15270003 TI - [Hemorrhagic infarction of the testicle in the newborn infant. A case presentation]. AB - Hemorrhagic infarction of the testicle is an unusual occurrence in the newborn infant. It usually develops as a consequence of torsion of the spermatic cord. We report a case of global testicular infarction in a newborn associated with a tense hydrocele. PMID- 15270004 TI - Asian Federation of Medicinal Chemistry (AFMC) - Fifth International Medicinal Chemistry Symposium. 14-17 October 2003, Kyoto, Japan. PMID- 15270005 TI - Asian Federation of Medicinal Chemistry (AFMC) - Fifth International Medicinal Chemistry Symposium. New drug highlights. 14-17 October 2003, Kyoto, Japan. PMID- 15270007 TI - Bibliography. Current world literature. Diseases of the pleura. PMID- 15270006 TI - European College of Neuropsychopharmacology - 16th Congress. 20-24 September 2003, Prague, Czech Republic. PMID- 15270009 TI - Current world literature. Nutrition in wasting disease. PMID- 15270008 TI - High-dose tizanidine abuse. PMID- 15270010 TI - Current world literature. Carbohydrates. PMID- 15270011 TI - Abstracts of the 6th European Congress on Epileptology. Vienna, Austria, 30 May-3 June, 2004. PMID- 15270012 TI - Abstracts of the 12th Alpe Adria Cardiology Meeting. April 29-May 1, 2004, Cividale del Friuli, Italy. PMID- 15270013 TI - Abstracts of the 39th Congress of the European Society for Surgical Research (ESSR). Athens, Greece, May 12-15, 2004. PMID- 15270014 TI - DEGRO 2004. Abstracts of the 10th Annual Congress of the German Society of Radio Oncology. Erfurt, 10-13 June 2004. PMID- 15270015 TI - Abstracts of the 8th Annual Meeting of the Scientific Association of Swiss Radiation Oncology (SASRO). Lucerne, 11-13 March 2004. PMID- 15270016 TI - Abstracts of the 3rd General Brachytherapy Conference, SASRO/DEGRO/OEGRO. Bern, 4 6 September 2003. PMID- 15270017 TI - Abstracts of the Barany Society XXIII International Congress. Paris, France, July 7-9, 2004. PMID- 15270018 TI - Abstracts of the 12th International Congress of Histochemistry and Cytochemistry: Cellular Interactions in Development and Disease. San Diego, California, USA, 24 29 July 2004. PMID- 15270019 TI - [Abstracts of the XXIV Cancer Forum. 29 June-1 July 2004, Paris, France]. PMID- 15270021 TI - Joint Brachytherapy Meeting, GEC/ESTRO-ABS-GLAC. Barcelona, Spain, 13-15 May 2004. Abstracts. PMID- 15270020 TI - Abstracts of the 2nd ESTRO Meeting on Radiotherapy for Non-Malignant Diseases. France, Nice, 1-3 April 2004. PMID- 15270022 TI - European patients find a voice. PMID- 15270023 TI - From certain death to benign lumps: paediatric cancer transformed. PMID- 15270024 TI - Addendum to "Regulation of the water channel aquaporin-1: isolation and reconstitution of the regulatory complex" [Cell Biol. Int. 2004(1):7-17]. PMID- 15270025 TI - Differential cAMP levels and serotonin effects in blood peripheral mononuclear cells and lymphocytes from major depression patients. AB - cAMP regulates immune responses, and modifications in cAMP signaling are involved in the pathophysiology and treatment of depression. In the present report, basal and forskolin-stimulated levels of cAMP were determined in mononuclear cells and lymphocytes from control individuals and major depression patients. Twenty-eight patients between 24 and 59 years old were diagnosed for a major depression episode according to the criteria of the Structural Clinical Interview for Disorders of Axis I of the American Psychiatric Association. These patients presented a score of 25 for severity as measured by Hamilton Rating Scale of Depression (HAM-D), and 23 for Beck Inventory of Depression (BID). Control and patient mononuclear cells were isolated by Ficoll/Hypaque gradients and their lymphocytes were separated from the total mononuclear population by differential adhesion to plastic surface. The basal concentration of cAMP was 50% lower in mononuclear cells and lymphocytes from the depressed patients compared with the control subjects. The response to forskolin was significantly smaller in lymphocytes of major depression patients than in the controls, but no difference was evident in the mononuclear cell preparations. There was a significant increase in cAMP produced by 5HT in mononuclear cells from the control group, but not in their lymphocytes. This effect on mononuclear cells was reduced by the antagonist of 5HT1A receptors, WAY-100,135. However, the simultaneous addition of a specific agonist of 5HT1A receptors, 8-hydroxy-(dipropylamino)tetralin (DPAT) and WAY-100,135 resulted in higher levels of cAMP than with the agonist alone. This effect probably indicates the blockade of 5HT1A receptors and action of 5HT1A agonist on the other subtypes of serotonin receptors expressed on human lymphocytes. This response was not observed in the patient's lymphocytes. In lymphocytes from major depression patients, serotonin and 8-hydroxy (dipropylamino)tetralin significantly increased cAmp levels, which was slightly reduced by WAY-100,135. The present report indicates: (1) differential responses of immune cells from control individuals and depressed patients, with lower apparent adenylate cyclase activity in patient's cells; (2) variation in the population of cells, with responses to serotonergic agonists being lower in mononuclear cells and higher in lymphocytes from major depression patients; (3) increases of cAMP levels by serotonin and 5HT1A agonist in the patient's cells; and (4) evidence of impairment in serotonergic transduction systems in immune cells during depression. PMID- 15270026 TI - Preventing infant death and injury during delivery. PMID- 15270027 TI - Hot drugs 2004. PMID- 15270028 TI - ConVir 2004. Abstracts of the 3rd European Conference on Viral Diseases. May 14 16, 2004, Regensburg, Germany. PMID- 15270029 TI - Future Pain Drugs - Europe 2003. 15-16 September 2003, London, UK. PMID- 15270030 TI - Mechanisms and Treatment of Neuropathic Pain - Sixth International Conference. 18 20 September 2003, San Francisco, CA, USA. PMID- 15270031 TI - Interscience Conference of Antimicrobial Agents and Chemotherapy - 43rd Annual Meeting. HIV and vaccines. 13-17 September 2003, Chicago, Il, USA. PMID- 15270032 TI - Interscience Conference on Antimicrobial Agents and Chemotherapy - 43rd Annual Meeting. Herpes viruses. 13-17 September 2003, Chicago, IL, USA. PMID- 15270033 TI - Thrombosis and Hemostasis Issues in Cancer - Second International Conference. 19 21 September, 2003, Bergamo, Italy. PMID- 15270034 TI - Protein & Peptide Drug Delivery - Third International Conference, 22-23 September 2003, Philadelphia, PA, USA. PMID- 15270035 TI - Protein & Peptide Drug Delivery - Third International Conference. Minimally invasive delivery methods. 22-23 September 2003, Philadelphia, PA, USA. PMID- 15270036 TI - AIDS Vaccine 2003. 18-21 September 2003, New York, NY, USA. PMID- 15270037 TI - Current awareness in human psychopharmacology. PMID- 15270038 TI - Inflammatory Processes in Drug Discovery - SRI Conference. 22-23 September 2003, Philadelphia, PA, USA. PMID- 15270039 TI - Cell & Gene Therapies - SMi Conference. 24-25 September 2003, London, UK. PMID- 15270040 TI - BioPartnering Europe - 11th Annual Conference. Investing in global partnerships. 12-14 October 2003, London, UK. PMID- 15270041 TI - BioBusiness Network 2003. 17-18 September 2003, Geneva, Switzerland. PMID- 15270042 TI - Essentials of a Magnetic work environment: part 2. PMID- 15270043 TI - Caring for a patient after CABG surgery. PMID- 15270044 TI - Taking the heat off: how to manage heat injuries. PMID- 15270045 TI - Patient education series. Gastroesophageal reflux disease (GERD). PMID- 15270047 TI - How herbs can affect your patient's health. PMID- 15270050 TI - [Imaging of the renal arteries: when, how and why?]. AB - The objectives of this course are both: to describe acquisition, injection and reconstruction parameters of volumic images for renal arteries examination and specific signs; to discuss the role of the different images in the diagnosis and in the therapeutic management. Ultrasound is one of the best imaging for the analysis of renal arteries in the detection of stenosis even if the sensitivity is less (around 85%)compared to CT Angiography (95%) and MR Angiography (90%). Because of this advantage and of 3D evaluation, CTA and MRA are sometimes in the first line for renal artery evaluation and can assess morphology before angioplasty. Renal scintigraphy with Captopril test and renin dosage are only used for small kidney evaluation. Arteriogram is systematically followed by angioplasty if possible. With the new endovascular materials, complications decrease (less than 5% with a major reduction in cholesterol emboli) and indications of endoprosthesis increase (71% of stenting with half of it in direct stenting technique). This course will give practical tools for imaging acquisition, specifically 3D imaging, for indications and management of lesion in accordance to symptoms and morphology. PMID- 15270051 TI - Translating research into practice: speeding the adoption of innovative health care programs. AB - For this study, the authors conducted case studies of four varied clinical programs to learn key factors influencing the diffusion and adoption of evidence based innovations in health care. They found that the success and speed of the adoption/diffusion process depend on: the roles of senior management and clinical leadership; the generation of credible supportive data; an infrastructure dedicated to translating the innovation from research into practice; the extent to which changes in organizational culture are required; and the amount of coordination needed across departments or disciplines. The translation process also depends on the characteristics and resources of the adopting organization, and on the degree to which people believe that the innovation responds to immediate and significant pressures in their environment. PMID- 15270052 TI - [The copyright. What happens in Germany with the intellectual property of authors?]. PMID- 15270053 TI - [Abstracts of the 30th Annual Meeting of the German Society of Neonatology and Pediatric Intensive Medicine. 24-26 June 2004]. PMID- 15270054 TI - Abstracts of the 13th International Symposium on Infections in the Immunocompromised Host. June 27-30, 2004, Granada, Spain. PMID- 15270055 TI - XXV Endocrinology Meeting of Pisa. Pisa, Italy, June 3-5, 2004. Abstracts. PMID- 15270056 TI - Safety lapses in kids' hospital care cost $1 billion, according to estimate. PMID- 15270057 TI - [Abstracts from the 45th National Congress of the Italian Society of Nephrology. Torino, Italy, 27-30 June 2004]. PMID- 15270058 TI - New guidelines urge early intervention for high blood pressure in children. PMID- 15270059 TI - Migrant Health in Europe. Abstracts of the International Conference on Differences in Health and in Health Care Provision. 23-25 June 2004, Rotterdam, The Netherlands. PMID- 15270060 TI - ICU physicians can save money, study says. PMID- 15270061 TI - Specialized nurse intervention improves post-hospitalization outcomes in elderly heart failure patients. PMID- 15270062 TI - Nucleic acid of infectious laryngotracheitis virus. PMID- 15270063 TI - Towards an embodied understanding of the structure/agency relationship. AB - Theoretical conceptualizations of the structure/agency relationship have been central to the development of the discipline, yet tend to exhibit two major limitations. First, they share a relatively disembodied view of the agent which overemphasizes cognition and marginalizes the significance of the emotional dimensions of interaction for human action and social structure. Second, most have difficulty maintaining the causal significance of both the 'people' and the 'parts' of the social system and are, therefore, unable to examine adequately their interplay. This paper suggests these problems are related, and examines the contribution recent formulations of the 'interaction order' can make toward overcoming the difficulties characteristic of this key sociological debate. The 'interaction order' identifies the embodied dimensions of interaction as consequential for, yet irreducible to, structures and agents, enables us to investigate the 'loose coupling' of interaction to individuals and social systems, but is underdeveloped in important respects. This paper addresses these limitations. It also highlights the utility for the structure/agency debate of identifying a sector of embodied interactions concerned with the maintenance of social selves, and suggests how this somatic sector of social life might be developed analytically. PMID- 15270064 TI - Weber's alleged emotivism. AB - This paper seeks to refute Alasdair MacIntyre's contention that the sociology of Max Weber is emotivist. MacIntyre understands emotivism to involve the collapse of all moral judgment into statements of personal preference. It is shown that Weber's sociology analyses this condition and seeks to repudiate it. In no way does Weber embrace emotivism. MacIntyre misses Weber's repudiation because he misreads Weber's sociological project. The paper shows that MacIntyre's reading of Weber can be refuted if attention is paid to the 'Politics as a Vocation' lecture. PMID- 15270065 TI - Is Bauman's bureau Weber's bureau?: a comment. AB - In his highly regarded and influential Modernity and the Holocaust Zygmunt Bauman launched one of the most passionate and sustained critiques of 'bureaucratic rationality' seen within social theory for some time. In so doing he drew heavily upon the work of Max Weber for support. In this brief paper I am interested in exploring the extent to which Weber really is the anti-bureaucratic ally Bauman claims him to be. In the first part of the paper I outline the main elements of Bauman's critique of bureaucratic rationality, drawing particular attention to its reliance upon a self-consciously Weberian theoretical lexicon. In the second part of the paper I seek to indicate that, despite his claims to be following in Weber's tracks, Bauman's conclusions regarding the moral vacuity of bureaucratic conduct are the very antithesis of Weber's own. PMID- 15270066 TI - Globalization and the policing of protest: the case of APEC 1997. AB - The policing of protest at international events conflicts with the political and policing culture of the host nation. Previous research shows a trend toward softer, more tolerant styles of policing protest within various Western democracies. We present a case study of an exception: the repression of protest at an international event in which one Western democracy hosted rulers of less democratic regimes in a ritual celebration of economic globalization. We explore reasons why, in the face of protests about undemocratic regimes elsewhere, the Canadian government and police were willing to use blatantly undemocratic tactics popularly believed to be more characteristic of those other regimes. Implications are discussed concerning protest policing, economic globalization, the nation state and social movements. PMID- 15270067 TI - Class voting, social change, and the left in Australia, 1943-96. AB - The relationship between class and voting choices has been the subject of controversy in recent years, especially in connection with the apparent decline of the traditional left. This paper examines class voting in Australia, focusing on three major issues: (1) changes in the overall strength of class voting (2) realignment, or changes in the relative political positions of the classes (3) the connection between the strength of class voting and support for Labor. It finds that (1) there is a decline in 'general' class voting (2) much of this decline involves a realignment of certain middle class groups, but there is no support for the popular idea that class alignments have become more complex (3) there is no connection between the strength of class voting and Labor performance. Our results cast doubt on accounts that regard the electoral difficulties of left parties as a symptom of the decline of class. PMID- 15270068 TI - Boredom and social meaning. AB - Meaning is necessary in social processes. An absence of meaning in an activity or circumstance leads to an experience of boredom. This is a restless, irritable feeling that the subject's current activity or situation holds no appeal, and that there is a need to get on with something interesting. Thus boredom emotionally registers an absence of meaning and leads the actor in question towards meaning. Boredom, then, is central to key social processes centered on questions of meaningfulness. Given the pervasive preconditions for boredom, release from boredom is a factor that explains characteristic social practices, including risk taking and intergroup conflict. PMID- 15270069 TI - Fish, field, habitus and madness: the first wave mental health users movement in Great Britain. AB - This paper traces and explains the emergence of the mental health users movement in Great Britain, focusing specifically upon the formation of the Mental Patients Union in the early 1970s. The analysis presented in the paper draws, to some extent, from conventional movement theory. In addition, however, it draws from the work of Pierre Bourdieu. This represents an innovation in movement analysis and the necessity of this innovation is argued for in an early section of the paper. PMID- 15270070 TI - Rational Choice theory and prison chaplaincy: the chaplain's dilemma. AB - Critical responses to the application of Rational Choice theories to the study of religious phenomena tend to be polarized between outright denial that the theories have any relevance to religion and equally outspoken claims that the theories are the only hope for progress in the sociology of religion. This article aims to avoid both of these extreme positions by raising a question, instead, about one of Rational Choice's central propositions about religion. This proposition holds that levels of religious vitality vary positively with the degree to which agencies of the state are prevented from regulating religious activity. The findings of recent research into prison chaplaincy in the UK and the USA will be used to test this claim. The main argument will be that the existence of an established church has facilitated a higher level of religious activity, especially for minority faiths, in prisons in England and Wales than is possible in American prisons. This difference in religious vitality is explained in terms of the Church of England's privileged position as a 'broker' between the state and minority faith communities. There is greater equality of opportunity for religious activity in American prisons, but the level of the activity is necessarily lower. In neither country is there anything truly resembling a 'free' market for religion in prisons, but the established Church of England is able to use its quasi-monopoly powers to broker advantageous conditions for minority faith communities. This brokerage function may be advantageous to all religious organizations in the highly regulated 'economy' of prisons. PMID- 15270071 TI - New oral drugs for erectile dysfunction. AB - In 1998, we concluded that sildenafil (Viagra--fizer Ltd), a selective phosphodiesterase type 5 inhibitor, appeared to offer advantages over other medical approaches for erectile dysfunction in terms of ease of administration and cost. Oral drug treatment is now widely advocated as first-line therapy for erectile dysfunction, except where the cause is clearly psychological. In the past 4 years, three more oral preparations have been licensed in the UK for the treatment of men with erectile dysfunction. A sublingual preparation of the dopaminergic agonist apomorphine (Uprima--Abbott Laboratories Ltd) is the first centrally acting drug to be licensed. Tadalafil (Cialis--Eli-Lilly) and vardenafil (Levitra--Bayer PLC) are phosphodiesterase type 5 inhibitors. Here we review the place of these preparations for men with erectile dysfunction. PMID- 15270072 TI - Managing excessive daytime sleepiness in adults. AB - Estimates suggest that up to 1 in 8 adults experiences excessive sleepiness that is severe enough to interfere with daytime activities. This problem can affect alertness, work, education and relationships and lead to accidents. Here we review the management of adults with excessive daytime sleepiness. PMID- 15270073 TI - Cooperation of heat shock protein 90 and p23 in aryl hydrocarbon receptor signaling. AB - Aryl hydrocarbon receptor (AhR) is a transcription factor that is activated by the binding of xenobiotic and endogenous ligands. AhR interacts with heat shock protein (Hsp) 90 complexes and can be used as a functional substrate to detect chaperone-dependent processes. Yeast Hsp90 (hsp82) mutants that variably affected AhR signaling were identified using reporter gene assays. Some mutated alleles resided in the p23/adenosine triphosphate (ATP)-binding pocket of Hsp90, so the relationship between the cochaperone Sba1 (yeast p23) and adenosine triphosphatase (ATPase) activity was investigated. Deletion of the p23 gene in the hsp82G170D mutant background had a greater effect on AhR signaling than the individual mutations, suggesting that these 2 mutations have separate actions on AhR signaling. In contrast, p23 overexpression suppressed temperature sensitivity and AhR signaling defects in the hsp82G170D mutant strain, suggesting that there is a relationship between these 2 proteins. The mutated hsp82G170D protein lacked detectable ATPase activity and p23 binding in vitro, which may relate to the weakened AhR signaling observed in mutant cells. Sba1 (p23) suppressed Hsp82 ATPase activity in vitro. These studies implicate the p23 protein and the G170 region of Hsp90 as being important, but not essential, for AhR signaling. Our results are consistent with a model in which p23 inhibits Hsp90 ATPase activity, thereby stabilizing ATP-Hsp90-client protein complexes. PMID- 15270074 TI - Transcriptional regulation and binding of heat shock factor 1 and heat shock factor 2 to 32 human heat shock genes during thermal stress and differentiation. AB - Transcription of mammalian heat shock genes can be regulated by heat shock factors (HSF) 1 and 2. Although it has been shown previously that these factors respond to distinct stimuli, a broad analysis of the induction and function of these factors in living cells has not been performed. In our study, we assayed binding of human HSF1 and HSF2 at the promoters of 32 genes identified through LocusLink as heat shock genes in response to elevated temperature and hemin induced differentiation in human K562 erythroleukemic cells using the chromatin immunoprecipitation technique. We also measured the induced expression of these genes under these 2 conditions. We found that 17 of the 32 genes were transcriptionally induced during heat shock, and HSF1 binding was detected at 15 of the 17 promoters. Nearly all the genes induced by heat shock were also induced to a lesser degree during hemin treatment. However, some genes were induced significantly more during hemin treatment than during heat shock. A new finding is that HSF1 and HSF2 bind to the same targets, but HSF1 binding is activated more by heat than by hemin treatment, and HSF2 binding is only activated by hemin treatment and not by heat. This technology also identified previously unknown HSF1 binding sites near genes that were previously shown to be heat inducible that may contribute to gene-specific regulation. PMID- 15270075 TI - HSP27 regulates fibroblast adhesion, motility, and matrix contraction. AB - Heat shock protein 27 (HSP27) modulates actin-dependent cell functions in several systems. We hypothesized that HSP27 modulates wound contraction. Stably transfected fibroblast cell lines that overexpress HSP27 (SS12) or underexpress HSP27 (AS10) were established, and cell behaviors related to wound contraction were examined. First, fibroblast-populated collagen lattice (FPCL) contraction was examined because it has been studied as a wound-healing model. In floating FPCL contraction assays, SS12 cells caused increased contraction, whereas AS10 cells caused reduced contraction. Because floating matrix contraction is thought to be mediated by the tractional force of the cells, cell behaviors related to tractional force were examined. In collagen matrix, SS12 cells elongated faster and to a greater extent and contained longer stress fibers than control cells, whereas AS10 cells were slower to elongate than control cells. SS12 cells attached to the dishes more efficiently than the control, whereas AS10 cells attached less efficiently. Migration of SS12 cells on collagen-coated dishes was also enhanced, although AS10 cells did not differ from the control cells. In summary, HSP27 regulates fibroblast adhesion, elongation, and migration and the contraction of the floating matrix in a manner dependent on the level of its expression. PMID- 15270076 TI - The mitochondrial 60-kDa heat shock protein in marine invertebrates: biochemical purification and molecular characterization. AB - Sessile marine invertebrates undergo constant direct exposure to the surrounding environmental conditions, including local and global environmental fluctuations that may lead to fatal protein damage. Induction of heat shock proteins (Hsps) constitutes an important defense mechanism that protects these organisms from deleterious stress conditions. In a previous study, we reported the immunological detection of a 60-kDa Hsp (Hsp60) in the sea anemone Anemonia viridis (formerly called Anemonia sulcata) and studied its expression under a variety of stress conditions. In the present study, we show that the sponge Tetilla sp. from tidal habitats with a highly variable temperature regime is characterized by an increased level of Hsp60. Moreover, we show the expression of Hsp60 in various species among Porifera and Cnidaria, suggesting a general importance of this protein among marine invertebrates. We further cloned the hsp60 gene from A viridis, using a combination of conventional protein isolation methods and screening of a complementary deoxyribonucleic acid library by polymerase chain reaction. The cloned sequence (1764 bp) encodes for a protein of 62.8 kDa (588 amino acids). The 62.8-kDa protein, which contains an amino terminal extension that may serve as a mitochondrial targeting signal, shares a significant identity with mitochondrial Hsp60s from several animals but less identity with Hsp60s from either bacteria or plants. PMID- 15270077 TI - Mild heat stress stimulates 20S proteasome and its 11S activator in human fibroblasts undergoing aging in vitro. AB - Repeated mild heat shock (RMHS) has been shown to have several beneficial hormetic effects on human skin fibroblast undergoing aging in vitro. Because an age-related decline in proteasome activity is 1 of the reasons for the accumulation of abnormal proteins during aging, we have investigated the effects of RMHS on the 20S proteasome, which is the major proteolytic system involved in the removal of abnormal and oxidatively damaged proteins. Serially passaged human skin fibroblasts exposed to RMHS at 41 degrees C for 60 minutes twice a week had increased 3 proteasomal activities by 40% to 95% in early- and midpassage cultures. RMHS-treated cells also contained a 2-fold higher amount of the proteasome activator 11S, and the extent of the bound activator was double in early- and midpassage cells only. Furthermore, there was no difference in the content of the 19S proteasome regulator in the stressed and the unstressed cells. Therefore, RMHS-induced proteasome stimulation in early- and midpassage fibroblasts appears to be due to an induction and enhanced binding of 11S proteasome activators. In contrast to this, the proteasomal system in late passage senescent cells appears to be less responsive to the stimulatory effects of mild heat shock. PMID- 15270078 TI - Induction of molecular chaperones in carbon tetrachloride-treated rat liver: implications in protection against liver damage. AB - Carbon tetrachloride (CCl4) induces liver damage, apparently through the formation of free-radical metabolites. Molecular chaperones such as heat shock protein (Hsp) of 70 kDa have been found to protect cells from various stresses. We previously found that cytosolic chaperone pairs of the Hsp70 family and their DnaJ homolog cochaperones prevent nitric oxide-mediated apoptosis and heat induced cell death. Expression of cytosolic chaperones, including Hsp70; heat shock cognate (Hsc) 70; and DnaJ homologs dj1 (DjB1/Hsp40/hdj-1), dj2 (DjA1/HSDJ/hdj-2), dj3 (DjA2), and dj4 (DjA4), in the liver of CCl4-treated rats was analyzed. Messenger ribonucleic acids for all these chaperones were markedly induced 3-12 hours after CCl4 treatment with a maximum at 6 hours. Hsp70 and dj1 proteins were markedly induced at 6-24 hours with a maximum at 12 hours, whereas dj2 and dj4 were moderately induced at around 12 hours. Hsc70 was weakly induced after treatment, and dj3 was little induced. To better understand the significance of the induction of chaperones, the effect of preinduction of chaperones on CCl4-induced liver damage was analyzed. When chaperones were preinduced in the liver by heat treatment, increase in serum alanine aminotransferase activity after CCl4 treatment was significantly attenuated. Hsp90, another major cytosolic chaperone, also was induced by heat treatment. On the other hand, Mn- and Cu/Zn-superoxide dismutase were not induced by heat treatment or by CCl4 treatment. These results suggest that cytosolic chaperones of Hsp70 and DnaJ families or Hsp90 (or both) are induced in CCl4-treated rat liver to protect the hepatocytes from the damage being inflicted. PMID- 15270079 TI - Serum and lymphocyte levels of heat shock protein 70 in aging: a study in the normal Chinese population. AB - Heat shock proteins (Hsps) have been reported to play an important role in both physiological and pathological processes. Hsps also may serve as biomarkers for evaluating disease states and exposure to environmental stresses. Whether Hsp levels in serum and lymphocytes are correlated with age and sex is largely unknown. In this study, we analyzed serum Hsp70 (the most abundant mammalian Hsp) levels by using Western dot blot in 327 healthy male donors aged between 15 and 50 years. We also investigated the association between Hsp70 levels and age in lymphocytes of 80 normal individuals aged between 40 and 77 years because various chronic diseases increase after the age of 40 years. Our data showed that serum Hsp70 levels were positively correlated with age in subjects aged between 15 and 30 years (P < 0.05) but negatively correlated with age in subjects aged between 30 and 50 years (P < 0.05). Serum Hsp70 levels were the highest in individuals aged between 25 and 30 years among all age groups. In the lymphocyte study there also was a significant age-related decrease in Hsp70 levels in lymphocytes of individuals older than 40 years. The Hsp70 levels were negatively correlated with age (r = -3.708, P < 0.0001) but not with sex (r = -10.536, P = 0.452). This suggests that both serum and lymphocyte Hsp70 levels are age-related and that these may be linked to age-related stress. Thus, age is an important factor in using serum and lymphocyte Hsp70 as biomarkers to evaluate the disease states or exposure to environmental stresses (or both). PMID- 15270080 TI - Endoplasmic reticulum stress as a correlate of cytotoxicity in human tumor cells exposed to diindolylmethane in vitro. AB - The dietary phytochemical indole-3-carbinol (I3C) protects against cervical cancer in animal model studies and in human clinical trials. I3C and its physiologic condensation product diindolylmethane (DIM) also induce apoptosis of tumor cells in vitro and in vivo, suggesting that these phytochemicals might be useful as therapeutic agents as well as for cancer prevention. Deoxyribonucleic acid microarray studies on transformed keratinocytes and tumor cell lines exposed to pharmacologic concentrations of DIM in vitro are consistent with a cellular response to nutritional deprivation or disruptions in protein homeostasis such as endoplasmic reticulum (ER) stress. In this report we investigate whether specific stress response pathways are activated in tumor cells exposed to DIM and whether the ER stress response might contribute to DIM's cytotoxicity. Induction of the stress response genes GADD153, GADD34 and GADD45A, XBP-1, GRP78, GRP94, and asparagine synthase was documented by Western blot and real-time reverse transcriptase-polymerase chain reaction in C33A cervical cancer cells, and induction of a subset of these was also observed in cancer cell lines from breast (MCF-7) and prostate (DU145). The results are consistent with activation of more than 1 stress response pathway in C33A cells exposed to 75 microM DIM. Phosphorylation elF2alpha was rapidly and transiently increased, followed by elevated levels of ATF4 protein. Activation of IRE1alpha was indicated by a rapid increase in the stress-specific spliced form of XBP-1 messenger ribonucleic acid and a rapid and persistent phosphorylation of JNK1 and JNK2. Transcriptional activation dependent on an ATF6-XBP-1 binding site was detected by transient expression in MCF-7, C33A, and a transformed epithelial cell line (HaCaT); induction of the GADD153 (CHOP) promoter was also confirmed by transient expression. Cleavage of caspase 12 was observed in both DIM-treated and untreated C33A cells but did not correlate with cytotoxicity, whereas caspase 7 was cleaved at later times, coinciding with the onset of apoptosis. The results support the hypothesis that cytotoxic concentrations of DIM can activate cellular stress response pathways in vitro, including the ER stress response. Conversely, DIM was especially cytotoxic to stressed cells. Thapsigargin and tunicamycin, agents that induce ER stress, sensitized cells to the cytotoxic effects of DIM to differing degrees; nutrient limitation had a similar, but even more pronounced, effect. Because DIM toxicity in vitro is enhanced in cells undergoing nutritional deprivation and ER stress, it is possible that stressed cells in vivo, such as those within developing solid tumors, also have increased sensitivity to killing by DIM. PMID- 15270081 TI - Administration of Hsp70 in vivo inhibits motor and sensory neuron degeneration. AB - The induction of heat shock proteins (Hsps) serves not only as a marker for cellular stress but also as a promoter of cell survival, which is especially important in the nervous system. We examined the regulation of the constitutive and stress-induced 70-kD Hsps (Hsc70 and Hsp70, respectively) after sciatic nerve (SN) axotomy in the neonatal mouse. Additionally, the prevention of axotomy induced SN cell death by administration of several preparations of exogenous Hsc70 and Hsp70 was tested. Immunohistochemistry and Western blot analyses showed that endogenous levels of Hsc70 and Hsp70 did not increase significantly in lumbar motor neurons or dorsal root ganglion sensory neurons up to 24 hours after axotomy. When a variety of Hsc70 and Hsp70 preparations at doses ranging from 5 to 75 microg were applied to the SN stump after axotomy, the survival of both motor and sensory neurons was significantly improved. Thus, it appears that motor and sensory neurons in the neonatal mouse do not initiate a typical Hsp70 response after traumatic injury and that administration of exogenous Hsc/Hsp70 can remedy that deficit and reduce the subsequent loss of neurons by apoptosis. PMID- 15270082 TI - Heat shock treatment protects against angiotensin II-induced hypertension and inflammation in aorta. AB - Angiotensin II (Ang II) is a potent vasoconstrictor and induces inflammation and end-organ injury through its activation of the proinflammatory transcription factor, nuclear factor-kappaB (NF-kappaB). Heat shock (HS) treatment with subsequent expression of heat shock proteins (Hsps) is an effective strategy for tissue protection against oxidative injuries. Recently, HS and Hsps have been shown to interact with NF-kappaB in tissue injury. In this study, we investigated whether HS could protect against Ang II-induced hypertension and inflammation by inhibiting NF-kappaB. Sprague-Dawley rats were divided into control and HS groups. Control and 24-hour post-heat shocked rats were treated with Ang II. At days 1, 3, 5, 7, 11, and 14 after Ang II administration, systolic blood pressures were measured by tail-cuff plethysmography, and aorta tissues were collected. Aorta NF-kappaB deoxyribonucleic acid-binding activity was measured by electrophoretic mobility shift assay, and NF-kappaB p65 subunit, Hsp70, Hsp27, and interleukin-6 (IL-6) expressions were measured by Western analysis. HS treatment significantly decreased Ang II-induced hypertension. The activation of NF-kappaB in aorta by Ang II was suppressed by HS treatment. The elevated expression of IL-6 induced by Ang II treatment was also decreased by HS treatment. Although Ang II treatment induced an increase in Hsp70 and Hsp27, HS treatment induced a greater elevation of Hsp70 and Hsp27 expression. HS treatment protects against Ang II-induced hypertension and inflammation. This protection may relate to the interaction of Hsps and the NF-kappaB pathway. PMID- 15270083 TI - Effects of Microphallus turgidus (Trematoda: Microphallidae) on the predation, behavior, and swimming stamina of the grass shrimp Palaemonetes pugio. AB - The effect of the trematode Microphallus turgidus on its second intermediate host, the grass shrimp, Palaemonetes pugio, was tested. To do so, we measured the susceptibility of infected and uninfected shrimp to predation by the mummichog, Fundulus heteroclitus. Shrimp behavior was compared in the presence and absence of a fish predator, and the swimming stamina and backthrust escape responses of infected and uninfected shrimp were measured. Infected shrimp were more likely to be eaten by a predator than uninfected shrimp, had lower swimming stamina, and spent more time swimming and less time motionless in the presence of a predator. There was no difference between backthrust distances traveled in response to a stimulus by either infected or uninfected shrimp. Thus, M. turgidus may increase the predation of P. pugio in the wild, possibly by affecting the swimming stamina and predator avoidance responses of the shrimp. PMID- 15270084 TI - Flow cytometric analysis of lectin binding to in vitro-cultured Perkinsus marinus surface carbohydrates. AB - Parasite surface glycoconjugates are frequently involved in cellular recognition and colonization of the host. This study reports on the identification of Perkinsus marinus surface carbohydrates by flow cytometric analyses of fluorescein isothiocyanate-conjugated lectin binding. Lectin-binding specificity was confirmed by sugar inhibition and Kolmogorov-Smirnov statistics. Clear, measurable fluorescence peaks were discriminated, and no parasite autofluorescence was observed. Parasites (GTLA-5 and Perkinsus-1 strains) harvested during log and stationary phases of growth in a protein-free medium reacted strongly with concanavalin A and wheat germ agglutinin, which bind to glucose-mannose and N-acetyl-D-glucosamine (GlcNAc) moieties, respectively. Both P. marinus strains bound with lower intensity to Maclura pomifera agglutinin, Bauhinia purpurea agglutinin, soybean agglutinin (N-acetyl-D-galactosamine specific lectins), peanut agglutinin (PNA) (terminal galactose specific), and Griffonia simplicifolia II (GlcNAc specific). Only background fluorescence levels were detected with Ulex europaeus agglutinin I (L-fucose specific) and Limulus polyphemus agglutinin (sialic acid specific). The lectin-binding profiles were similar for the 2 strains except for a greater relative binding intensity of PNA for Perkinsus-1 and an overall greater lectin-binding capacity of Perkinsus-1 compared with GTLA-5. Growth stage comparisons revealed increased lectin-binding intensities during stationary phase compared with log phase of growth. This is the first report of the identification of surface glycoconjugates on a Perkinsus spp. by flow cytometry and the first to demonstrate that differential surface sugar expression is growth phase and strain dependent. PMID- 15270085 TI - Endogenous polyamine levels in macrophages is sufficient to support growth of Toxoplasma gondii. AB - Cytotoxic-activated macrophages control Toxoplasma gondii growth by producing nitric oxide (NO). However, the parasite can partially inhibit NO production. NO is generated from arginine within the polyamine biosynthetic pathway. Two enzymes of this pathway are ornithine, decarboxylase (ODC) and arginine decarboxylase (ADC). The aim of the present work was to investigate whether T. gondii is able to modulate polyamine metabolism in macrophages. Toxoplasma gondii infection did not affect basal ODC or ADC activity. However, lipopolysaccharide induced an increase in ODC activity. Polyamine-treated macrophages exhibited a T. gondii infection index similar to controls but a higher adhesion index; the parasite did not grow in methyl-ornithine (ODC inhibitor)-treated macrophages. The parasites were able to take up putrescine with a Km of 0.92 microM, indicating the presence of a high-affinity putrescine-transporter system. Putrescine-treated T. gondii actively penetrated macrophages and Vero cells. However, NO production and lysosomal parasitophorous vacuole fusion were not inhibited. Considered together, these results demonstrate that T. gondii requires polyamines for multiplication. However, as opposed to Trypanosoma cruzi and because of a relatively high affinity putrescine-transporter system in the parasite, constitutive macrophage levels of putrescine seem sufficient to support T. gondii survival and multiplication. PMID- 15270086 TI - Comparative analysis of stress agents in a simplified in vitro system of Neospora caninum bradyzoite production. AB - Neospora caninum is an apicomplexan parasite identified as a major cause of abortion in cattle and neurological disease in various animal species. It is closely related to Toxoplasma gondii, sharing the ability to persist indefinitely in latent stage within the host as a tissue cyst containing slow-dividing bradyzoites. In this study, we compared different stress methods to induce in vitro bradyzoite conversion, using MARC-145 cells infected with Nc-Liverpool isolate. The tachyzoite-to-bradyzoite conversion rate was monitored at days 3, 5, and 7 after stress in a double-immunofluorescence assay using a monoclonal antibody against the tachyzoite antigen SAG1 (alphaSAG1) and a rabbit serum directed to the intracytoplasmic bradyzoite antigen BAG1 (alphaBAG1). Seven days of treatment with 70 microM sodium nitroprusside offered the highest bradyzoite transformation rate and the best yield of total parasitophorous vacuoles observed. In the present work, we introduce an alternative, simplified, and more advantageous method for bradyzoite production of N. caninum, using a reliable cell culture system easy to handle and with promising capacity of parasite purification. PMID- 15270087 TI - Helminth assemblages of whitefish (Coregonus lavaretus) in interconnected lakes: similarity as a function of species specific parasites and geographical separation. AB - This article examined the composition of parasite assemblages of whitefish (Coregonus lavaretus) in 8 interconnected lakes in northeastern Finland and evaluated the role of coregonid specific parasites and the geographical distance between populations in determining the similarity of the assemblages. Parasite assemblages were compared using the Jaccard qualitative similarity index and a quantitative similarity index and by incorporating the allogenic-autogenic species concept and the effects of 2 corresponding measures of geographical distance between the lakes. The majority of the parasite species found (10 of 14) were specific to salmonids. Similarity of assemblages of autogenic parasites between the lakes was negatively correlated with geographical distance. The dominance of 2 parasite species, the whitefish specialist Ichthyocotylurus erraticus and the generalist Ergasilus sieboldi, was also demonstrated. We concluded that the high proportion of widespread parasite species specific to coregonids is an important determinant of similarity in these assemblages. However, ecological factors were likely to contribute to qualitative (presence of species) and quantitative (difference in abundances) differences between lakes, in the case of autogenic parasites, their importance increasing with geographical separation. PMID- 15270088 TI - Supplemental diagnosis of Kudoa funduli (Myxozoa) parasitizing Fundulus heteroclitus (Cyprinodontidae) from coastal northeastern North America. AB - The diagnosis of Kudoa funduli (Hahn, 1915) Meglitsch, 1948 (Myxozoa), is supplemented through study of new material collected from Fundulus heteroclitus (Cyprinodontidae) in coastal waters of Nova Scotia, Canada, and Connecticut. Plasmodia normally develop intracellularly in striated muscle of the flank and head, eventually rupturing and releasing spores. Spores disperse along adjacent epimysium, sometimes as far as the skin surface. Some plasmodia develop extracellularly within the bony cavities of vertebrae. Formalin-fixed spores viewed with a light microscope possess rounded edges, an inconspicuous apical region, thin sutural ridges, measure 6.6-7.4 microm wide, 4.3-5 microm thick, and 5.1-5.4 microm long, and have 4 equally sized polar capsules, 1.7-2.3 microm length by 1.4-1.7 microm width. Scanning electron microscopy revealed that spores are almost stellate, with inconspicuous uplifted tips, and that, within intracellular plasmodia, are embedded in an extensive honeycomb-like matrix. Prevalence of infection of K. funduli was 100% in host populations sampled in both Nova Scotia and Connecticut. Molecular sequence data of the 18S ribosomal DNA (737 base pairs) reveal that K. funduli is a valid species and a member of a clade that includes Kudoa dianae Dykova, Avila, and Fiala, 2002, Kudoa miniauriculata Whitaker, Kent, and Sakanari, 1996, and Kudoa paniformis Kabata and Whitaker, 1981. PMID- 15270089 TI - Schistobrachia jordaanae n. sp. (Copepoda: Siphonostomatoida: Lernaeopodidae) from gill filaments of a diamond ray (Gymnura natalensis) captured in the Indian Ocean and a key to species of Schistobrachia, Dendrapta, and Brianella. AB - Schistobrachia jordaanae n. sp. (Copepoda: Siphonostomatoida: Lernaeopodidae) is described from adult female specimens collected from gill filaments of a diamond ray Gymnura natalensis (Gilchrist and Thompson, 1911) captured in the Indian Ocean off the South African coast. Schistobrachia jordaanae is best distinguished from its congeners by 2 unique characteristics: it possesses a necklike region between the origin of its maxillae and maxillipeds and the tips of its maxillae bifurcate repeatedly to form a rootlike anchor. PMID- 15270090 TI - Ectoparasites of gray squirrels in two different habitats and screening of selected ectoparasites for bartonellae. AB - Gray squirrels, Sciurus carolinensis, were livetrapped in 2 different habitat types, woodland (67 squirrels) and parkland (53 squirrels), in southeastern Georgia. Ectoparasites were recovered from anesthetized squirrels and compared between hosts from the 2 habitats. Because of the absence of low vegetation in parkland habitats, it was hypothesized that the ectoparasite fauna, especially ticks and chiggers, would be more diverse on woodland squirrels. The results were generally in agreement with this hypothesis. Seventeen species of ectoparasites were recovered from woodland squirrels, compared with 6 species from parkland squirrels. Five species of ticks and 3 species of chiggers parasitized the woodland squirrels compared with no ticks or chiggers on the parkland squirrels. Significantly higher infestation prevalences were recorded on woodland compared with parkland squirrels for the flea Orchopeas howardi, the tick Amblyomma americanum, and the mesostigmatid mite Androlaelaps fahrenholzi. The mean intensity for O. howardi also was significantly higher on woodland than on parkland squirrels. Because a new strain of Bartonella sp. was isolated recently from S. carolinensis in Georgia, selected ectoparasites from this study were screened for bartonellae by polymerase chain reaction (PCR). Some of the fleas and lice, but none of the mites tested, were PCR positive, suggesting that fleas, or lice, or both, might be vectors of bartonellae between squirrels. Six distinct strains of Bartonella sp. were detected, 2 in fleas and 4 in lice. PMID- 15270091 TI - Three new species of Antricola (Acari: Argasidae) from Brazil, with a key to the known species in the genus. AB - Three new species of Antricola (Acari: Argasidae) are described from adult specimens collected on bat guano in different caves in Brazil. The female of Antricola guglielmonei n. sp. is easily determined by the presence of 2 smooth, depressed areas lacking setae in the posterolateral portions of the dorsum of the idiosoma, together with the partial fusion of the tubercles in the anteromedian portion of the idiosoma. The male of this species has a small spiracular plate surrounded by a pattern of tubercles disposed concentrically in its dorsal portion. Both sexes have cervical grooves very slightly marked. Antricola delacruzi n. sp. represents the only Antricola species with the dorsum of the idiosoma devoid of tubercles in both sexes and with scarce and minute setae placed over the smooth cuticle. The female of A. inexpectata n. sp. is known only from a few specimens. In this species, lines of smooth cuticle lacking setae separate the tubercles of the dorsum. In addition, there are 3 clumps of plumose setae close to the spiracular plate, in the ventrolateral portions of the idiosoma, in 3 well-delimited regions over cuticular thickenings. These 3 species share the peculiarity of a Haller's organ with the anterior pit bearing only 7 + 2 setae. The collection of these new species in Brazilian caves greatly expands the known range of the genus. A key to the adults of all known species of the genus is provided. PMID- 15270092 TI - Phasmid ultrastructure of an ascaridoid nematode Hysterothylacium auctum. AB - Here, for the first time in an ascaridoid (Hysterothylacium auctum), we present structural features of the phasmids, paired sense organs, positioned in a bilateral manner close to the point of the tail; the features were obtained using scanning and transmission electron microscopy. We found that each phasmid consists of a single ciliated dendritic process situated in a phasmidial canal surrounded by 2 supporting cells, a socket and a sheath cell. The socket cell contains clusters of electron-dense fibrous material in its apical region and covers the phasmidial canal along its whole length. The sheath cell is characterized by a well-developed endoplasmic reticulum. The phasmidial canal is lined with a thin layer of cuticle that becomes incomplete at the base of the ciliated dendritic process. In this region, the dendritic process consists primarily of a high number of microtubule singlets and some peripheral microtubule doublets. The base of the dendritic process, containing numerous striated rootlets, gives off a large number of fingerlike offshoots, villi, invading the surrounding sheath cell. The systematic significance and functional implication of the phasmid in nematodes are also discussed. PMID- 15270093 TI - Test of pangamy by genetic analysis of Schistosoma mansoni pairs within its natural murine host in Guadeloupe. AB - Mating system plays a determinant role in the maintenance and distribution of genetic variations. It can be assessed indirectly by analyzing the distribution of the genetic variability within populations or directly by considering how mating pairs are formed. In the present study, 71 pairs of adult Schistosoma mansoni worms sampled from naturally infected rats were genotyped to investigate how male and female schistosomes paired according to their genetic relatedness. Among all samples, pangamy, the random association between males and females, could not be rejected. Whereas the schistosome mating system has been intensively studied under experimental conditions, to the best of our knowledge, our study is the first to attempt to understand the way in which males and females pair in natural conditions. PMID- 15270094 TI - Effect of hypoxia on macrophage infection by Leishmania amazonensis. AB - In the present study, we compared the effect of 5% oxygen tension (hypoxia) with a normal tension of 21% oxygen (normoxia) on macrophage infection by the protozoan parasite Leishmania amazonensis. Macrophages from different sources (human cell line U937, murine cell line J774, and murine peritoneal macrophages) exposed to hypoxia showed a reduction of the percentage of infected cells and the number of intracellular parasites per cell. Observations on the kinetics of infection indicated that hypoxia did not depress L. amazonensis phagocytosis but induced macrophages to reduce intracellular parasitism. Furthermore, hypoxia did not act synergistically with gamma-interferon and bacterial lipopolysaccharides in macrophages to induce killing of parasites. Experiments also indicated no correlation between nitric oxide production and control of infection in macrophages under hypoxic condition. Thus, we have provided the first evidence that hypoxia, which occurs in various pathological conditions, can alter macrophage susceptibility to a parasitic infection. PMID- 15270095 TI - Challenge of Trypanosoma cruzi chronically infected mice with trypomastigotes activates the immune system and reduces subpatent parasitemia levels. AB - Challenge of 1-yr Trypanosoma cruzi chronically infected mice with trypomastigotes results in a consistent reduction of parasite dissemination that correlates with spleen activation and increase in the anti-T. cruzi effector immune mechanisms. That is, parasite challenge results not only in elimination of the inoculum but also in a drastic decrease in basal subpatent parasitemia levels as revealed by transferring blood samples to immunosuppressed mice. Parasite elimination correlated with (1) a brief and intense burst in the ability of spleen cells to produce interferon-gamma, (2) an increase in total IgG2a producing spleen cells, (3) higher parasite-specific IgG2a serum levels, and (4) an accumulation of non-B, non-T class II+ cells in the spleen. Furthermore, challenged, chronically infected mice had increased numbers of B, CD4+, and CD8+ large spleen cells. Besides reinforcing the activation of protective Th1 effector mechanisms, challenge with T. cruzi also induced Th2 effector molecules, such as interleukin (IL)-10 and IL-4, and IL-4-dependent IgG1. Our results are the first evidence that the immune system of T. cruzi chronically infected mice can be optimized in its ability to restrict parasite dissemination, opening the possibility that therapeutic vaccination could be used to reduce the parasite load and pathology of patients with chronic Chagas' disease. PMID- 15270096 TI - Discovery and characterization of an antibody, anti-egressin, that is able to inhibit Trypanosoma cruzi egress in vitro. AB - Trypanosoma cruzi, a protozoan parasite, is the etiologic agent of Chagas disease. The disease is characterized by acute and chronic phases, with high and low parasitemia, respectively. A strong immune activation is necessary for the host to enter the chronic phase; however, immune mechanisms participating in the reduction of parasites between the acute and chronic phases of the disease have been very difficult to elucidate. We report here the discovery of anti-egressin, an antibody present in serum from chronically infected BALB/c mice that is able to inhibit parasite egress from infected BALB/c fibroblast cultures in vitro. The antibody is very concentrated in serum from these mice; chronic serum may be diluted 1:20 while still maintaining functional activity. Isotype analysis of anti-egressin has suggested it to be IgG2a. Further analysis revealed that anti egressin was specific for a component expressed on the surface of infected host cells. The specificity of anti-egressin toward the extracellular portion of infected host cells was demonstrated both by using a quantitative assay measuring released trypomastigotes and through immunocytochemical staining. The novel role of anti-egressin in the inhibition of parasite egress from infected host cells has not been described in the literature to date. We believe that anti-egressin plays an important role in achieving the low parasitemia characteristic of chronic Chagas disease. PMID- 15270097 TI - Host gender in parasitic infections of mammals: an evaluation of the female host supremacy paradigm. AB - A review of current literature on mammalian hosts' sexual dimorphism (SD) in parasitic infections revealed that (1) it is a scarcely and superficially studied biological phenomenon of considerable significance for individual health, behavior, and lifestyles and for the evolution of species; (2) there are many notable exceptions to the rule of a favorable female bias in susceptibility to infection; (3) a complex network of molecular and cellular reactions connecting the host's immuno-neuroendocrine systems with those of the parasite is responsible for the host-parasite relationship rather than just an adaptive immune response and sex hormones; (4) a lack of gender-specific immune profiles in response to different infections; (5) the direct effects of the host hormones on parasite physiology may significantly contribute to SD in parasitism; and (6) the need to enrich the reductionist approach to complex biological issues, like SD, with more penetrating approaches to the study of cause-effect relationships, i.e., network theory. The review concludes by advising against generalization regarding SD and parasitism and by pointing to some of the most promising lines of research. PMID- 15270098 TI - Dynamics of the cytokine messenger RNA expression pattern in the liver of baboons infected with Schistosoma mansoni. AB - Periovular granulomas are the major lesions in baboons infected with Schistosoma mansoni. Temporal Northern blot analysis of cytokine messenger RNA (mRNA) expression in granulomatous baboon livers demonstrated tissue-specific expression. Interleukin 1 beta (IL-1beta, IL-6, tumor necrosis factor alpha, and migration inhibitory factor (MIF) mRNAs were expressed strongly at week 6 of infection and decayed thereafter, whereas interferon gamma (IFN-gamma), IL-2, IL 10, and IL-12 mRNAs were first expressed at week 12, with IFN-gamma and IL-12 mRNA expression persisting until week 17. IL-4 and IL-5 mRNAs first appeared at week 12, with IL-4 persisting unchanged and IL-5 increasing by week 17. Thus, egg deposition induced strong hepatic expression of proinflammatory and downregulatory cytokines. The cooccurrence of IL-2, IFN-gamma, IL-4, and IL-5 mRNAs at week 12 confirms that baboons, like humans, show a mixed type 1-type 2 cytokine response. When granulomas had become smaller at 17 wk, IFN-gamma, IL-4, and IL-5 were the only cytokine mRNAs that were expressed strongly, implicating them in granuloma modulation. The early expression of MIF mRNA and MIF's role as the main counterregulator of glucocorticoid immunosuppression ties in with our earlier demonstrations of circulating adrenal steroids changing with the progression of schistosomiasis in baboons and of proinflammatory cytokine mRNA expression in the hypothalamic-pituitary-adrenal axis tissues of infected baboons. Together, these data imply neuroendocrinological influences on disease progression in schistosomiasis. PMID- 15270099 TI - The human cytokine response to Leishmania major early after exposure to the parasite in vitro. AB - Leishmaniasis is caused by the protozoan parasite Leishmania spp. In murine leishmaniasis, a T helper cell type-I (Th1) response, characterized by the secretion of interferon (IFN)-gamma is necessary for clearing the infection. whereas a Th2 response, accompanied by the production of interleukin (IL)-5, can exacerbate the disease. Moreover, the early cytokine milieu is thought to play an important role in determining the outcome of infection. In human leishmaniasis little is known about this early cytokine response. Because of this, we cocultured human peripheral blood mononuclear cells (PBMC) with Leishmania major in vitro and measured the production of IFN-gamma, IL-5, and IL-10. We also treated PBMC cultures with various cytokines and neutralizing anticytokines. We found that the principal cytokine produced was IFN-gamma and that its production was regulated by IL-10 and IL-12. In contrast, only low levels of Th2 cytokines such as IL-5 were produced. Therefore, the Th1-Th2 dichotomy that exists in inbred strains of mice does not appear to apply to the response of humans to L. major. Rather, Th2 cytokines may play a role in regulating IFN-gamma production. PMID- 15270100 TI - Temporal changes in the prevalence of parasites in two Oregon estuary-dwelling fishes. AB - The parasite faunas of juvenile English sole (Parophrys vetulus) in 1971-1972 and staghorn sculpin (Leptocottus armatus) in 1971 from Yaquina Bay, Oregon, were compared with faunas found in the same estuary in 1997-2000 (English sole) and 1999-2000 (staghorn sculpin). The 7 most commonly occurring parasites in 1971 were compared with the same species observed during the same month and sampling sites in 1997-2000. Multivariate community analysis of juvenile English sole parasites supported the suggestion that the 1971 parasite data were representative of the early-1970s time period. Four of the parasite species infecting English sole and 6 of those infecting staghorn sculpins had significantly lower prevalences in 1997-2000. Parasite species with significantly lower prevalences also had reduced intensity levels. One parasite (Glugea stephani) of English sole increased in prevalence in the 1997-2000 samples in association with the warm estuarine temperatures during the 1997 El Nino year. Although the causes for the changes in occurrence of other parasites were not determined, ecological changes in Yaquina Bay that may have influenced parasite ecology include apparent changes in the estuary ichthyofauna that occurred between the sampling periods. Such changes could be associated with increases in the number of California sea lions (Zalophus californianus) subsequent to establishment of the Marine Mammal Protection Act in 1972. PMID- 15270101 TI - Helminth associations in white-toothed shrews Crocidura russula (Insectivora: Soricidae) from the Albufera Natural Park, Spain. AB - The helminths of 218 white-toothed shrews from 29 sites in 2 biotopes in the Albufera Natural Park (Valencia, Spain) were examined from July 1990 to August 1991. An association analysis of helminths occurring at a prevalence of more than 4% was carried out for 4 species of cestodes located in the intestine (Hymenolepis pistillum, H. scalaris, H. tiara, and Pseudhymenolepis redonica) and 3 species of nematodes (Pseudophysaloptera sp. located in the stomach, Stammerinema rhopocephala larvae in the intestine and abdominal cavity, and Porrocaecum sp. in the thoracic and abdominal cavities). Bivariate (species pairs) versus multivariate analyses (associations within the entire set of species) were performed of presence-absence and of quantitative records (influence of intensity on associations). The associations were evaluated with respect to the sex and age of the host and to the sampling date and sites. The host and environment played a limited role, and the major determinant of species assemblage was phylogenetic. Positive associations were found among both the cestodes and the nematodes, whereas negative associations were found between cestodes and nematodes. The type of life cycle was probably the second greatest determinant of species associations. Nematodes using shrews as a paratenic host or as their definitive host were both positively associated. PMID- 15270102 TI - Characterization of pathology and parasite load in outbred and inbred mouse models of chronic Neospora caninum infection. AB - Here, we analyzed histological findings and parasite burden in chronic Neospora caninum infection in BALB/c and ICR mice and studied the correlation between lesion severity and parasite load in brain. To obtain a better understanding of the infection, we examined the influence of various host pathogen factors. Groups of outbred (ICR) and inbred (BALB/c) mice were inoculated using several NC-1 parasite doses (4 x 10(5), 10(6), and 5 x 10(6) tachyzoites), inoculation routes (intraperitoneal and subcutaneous), and 3 immunosuppressive treatments (methylprednisolone, cyclophosphamide, and vinblastine). Lesion severity was analyzed in the liver, lung, heart, and brain tissues, and parasite load was measured by real-time polymerase chain reaction in brain tissue. The results indicated more severe cerebral lesions and higher brain parasite burdens in inbred than in outbred mice. Hepatic tissue was the primary lesion site in immunosuppressed ICR mice. We also observed that increased inoculum size was reflected in greater lesion severity and a higher cerebral parasite load. No difference was observed with respect to inoculation route. The study also showed an association between brain parasite burden and severity of cerebral lesions in BALB/c mice. PMID- 15270103 TI - Kudoa hypoepicardialis n. sp. (Myxozoa: Kudoidae) and associated lesions from the heart of seven perciform fishes in the northern Gulf of Mexico. AB - Kudoa hypoepicardialis n. sp. infects the space between the epicardium and the compact myocardium and, in intense infections, the pericardial chamber of man-of war fish (Nomeus gronovii) (Nomeidae) (the type host), blue runner (Caranx crysos) (Carangidae), Warsaw grouper (Epinephelus nigritus) (Serranidae), Atlantic tripletail (Lobotes surinamensis) (Lobotidae), northern red snapper (Lutjanus campechanus) (Lutjanidae), black drum (Pogonias cromis) (Sciaenidae), and bluefish (Pomatomus saltatrix) (Pomatomidae) in the northern Gulf of Mexico. This is the first report of a Kudoa sp. from the heart of a fish in the Gulf of Mexico, and of these hosts, only the bluefish was previously identified as a host for a species of Kudoa. Spores of the new species varied slightly in size among these hosts but were regarded as conspecific based on their nearly identical (99.9%) small-subunit (SSU) ribosomal DNA (rDNA) sequence. The new species differs both from the 4 nominal species of Kudoa reported from fishes in the Gulf of Mexico and from K. pericardialis, an allopatric species that infects the pericardial cavity, by the combination of having a large spore, a small polar capsule, and a polar filament with a single coil. The new species is morphologically and genetically most similar to K. shiomitsui, an allopatric species that infects the heart and pericardial cavity, but is distinguished from it based on a 4.2% difference in the SSU rDNA sequence. Heart lesions primarily were restricted to the vicinity of plasmodia and included a layer of fibrinous inflammation characterized by lymphocytes, macrophages, and granulomas as well as epithelioid encapsulations around plasmodia. Heavily infected hosts had melanin like deposits and adipose cells beneath the epicardium. and the epicardium was discontinuous and apparently breached by plasmodia in some regions. Cardiac muscle, gill, liver, spleen, intestine, and kidney were normal. PMID- 15270104 TI - Comments on the gonotyl of Proctocaecum macroclemidis (Tkach and Snyder, 2003) n. comb. (Digenea: Acanthostomidae: Acanthostominae), with a key to the genera of acanthostominae and new phylogenetic tree for Proctocaecum Baugh, 1957. AB - The species recently described as Acanthostomum macroclemidis possesses the gonotyl in the form of a solid muscular pad uniquely diagnostic for species of Proctocaecum and is accordingly transferred to that genus. An artificial key to the 5 acanthostomine genera, as well as an updated phylogenetic hypothesis for the 10 known species of Proctocaecum, based on 11 characters and including 2 species described since the last phylogenetic analysis, are presented. The single most parsimonious phylogenetic tree with a consistency index of 87.5% suggests that Proctocaecum originated in Africa and spread to North America and South America before the breakup of Pangaea. As a result, the 2 North American and 1 South American species are most closely related to different African members of the genus. African and Indo-Pacific species inhabit crocodylids; hence, the occurrence of North American species in alligatorids and chelonians and a South American species in alligatorids are the result of host switches. PMID- 15270105 TI - Two new species of Falcaustra and comments on helminths of Norops tropidolepis (Sauria: Polychrotidae) from Costa Rica. AB - Falcaustra costaricae n. sp. from the intestines of the lizard Norops tropidolepis and F. heosemydis n. sp. from the large intestine of the turtle Heosemys depressa are described and illustrated. Falcaustra costaricae represents the 10th Neotropical species assigned to this genus and is distinguished from other Neotropical species by the distribution pattern of caudal papillae (10 preanal, 0 adanal, 12 postanal, and 1 median), length and width of spicules (510 561 microm long, 18-24 microm wide), and absence of pseudosucker. Falcaustra heosemydis represents the 29th Oriental species and is distinguished from other Oriental species by the distribution pattern of caudal papillae (10 preanal, 0 adanal, 12 postanal, and 1 median), length of spicules (790-890 microm), and absence of pseudosucker. Norops tropidolepis was found to harbor 3 species of Nematoda, F. costaricae, Rhabdias anolis, and acuariid larvae, and 2 species of Acanthocephala (centrorhynchid cystacanths and oligacanthorhynchid cystacanths). PMID- 15270106 TI - Saccocoelium megasacculum n. sp., a new haploporid trematode from Liza carinatus in the Taiwan strait. AB - Saccocoelium megasacculum n. sp. (Digenea: Haploporidae) was collected from the intestine of the mugilid fish. Liza carinatus (Cuvier and Valenciennes), in the Taiwan Strait. It is the first record of Saccocoelium in China. The parasite most closely resembles Saccocoelium obesum Looss, 1902 and Saccocoelium tensum Looss, 1902 in general morphology and body size, but it is easily distinguished from them in having a larger hermaphroditic sac in relation to body size; larger eggs; smaller pharynx, testis, ovary, and vitellaria; and a uterine seminal receptacle instead of a true seminal receptacle. PMID- 15270107 TI - Observations on a cucullanid nematode of marine fishes from Taiwan Strait, Dichelyne (Cucullanellus) jialaris n. sp. AB - During a helminthological examination of marine fishes from south of the Minnan Taiwan Bank Fishing Ground, Taiwan Strait, Fujian, China, a new cucullanid nematode, Dichelyne (Cucullanellus) jialaris n. sp., was removed from the intestine of the red seabream, Pagrus major (Temminck & Schlegel, 1834). The new species differs from its congeners mainly in the following characters: body size medium but with relative long spicules of 1.01 mm (0.97-1.06) in length or 20.0% (18.21-21.8%) of the body length; proximal end of spicules somewhat expanded and distal end rounded; gubernaculum I-shaped, slightly narrow in the middle part, both ends rounded; both anterior and posterior cloaca lips round or oval, prominent and unequal in size. The anterior cloaca lip is at least 2 times larger than the posterior one. There is a conspicuous papilliform structure within the central of anterior and posterior cloacal lip. Vulva of female is not prominent, slightly postequatorial; distance from vulva to anterior end of body is 4.3 (3.0 5.5) mm or 58.0% (54.0-62.0%) of the body length. Considering the result of comparing the structure of so-called unpaired median papilla with the 10 pairs of caudal petiolated papillae in the body of the same individual. the papilliform structures are just a backstop for the cloacal lips, this new species represents the first record of a nematode of the Dichelyne, subgenus Cucullanellus in marine fishes of China Sea. PMID- 15270108 TI - A paraphyly of the genus Bothriocephalus Rudolphi, 1808 (Cestoda: Pseudophyllidea) inferred from internal transcribed spacer-2 and 18S ribosomal DNA sequences. AB - Phylogenetic relationships between Bothriocephalus species from freshwater and marine teleosts from different geographical regions were studied using internal transcribed spacer-2 and partial 18S ribosomal DNA. The analyses revealed a paraphyly of Bothriocephalus with respect to the genera Polyonchobothrium, Anantrum, and Clestobothrium. The freshwater species Bothriocephalus claviceps, B. acheilognathi, and Bothriocephalus sp. from Dorosoma petenense formed a well supported monophyletic cluster, with Polyonchobothrium at its base. In contrast, the type species, B. scorpii, clustered within a distinct lineage formed by a heterogeneous assemblage of marine species, Clestobothrium crassiceps and Anantrum tortum, and the freshwater species B. cf. japonicus. This shows that the current morphology-based classification is unlikely to reflect the phylogenetic relationships within this group and will require a thorough revision. PMID- 15270109 TI - Phylogeny of the multivalvulidae (Myxozoa: Myxosporea) based on comparative ribosomal DNA sequence analysis. AB - Fish parasites of the Multivalvulida (Myxozoa, Myxosporea) are widespread and can be associated with mortality or poor flesh quality in their commercially important marine hosts. Traditional classifications divide members of this order into families based on spore valve and polar capsule numbers. Analyses of the small-subunit (SSU) ribosomal DNA (rDNA) sequences from all representative families in the order (Trilosporidae, Kudoidae, Pentacapsulidae, Hexacapsulidae, and Septemcapsulidae) indicate that a revision of the taxonomy and nomenclature is warranted. In our phylogenetic analysis of (SSU and large subunit) rDNA sequences, members of Pentacapsula, Hexacapsula, and Septemcapsula root within a clade of Kudoa species with Unicapsula (Trilosporidae) as an outlier to these genera. Therefore, we propose to synonymize Pentacapsulidae, Hexacapsulidae, and Septemcapsulidae with Kudoidae alter the diagnosis of Kudoidae and Kudoa to accommodate all marine myxozoan parasites having 4 or more shell valves and polar capsules. PMID- 15270110 TI - Parabothriocephalus psenopsis n. sp. (Eucestoda: Pseudophyllidea) in Psenopsis anomala from the Taiwan Strait, China. AB - Palabothriocephalus psenopsis n. sp. (Eucestoda: Pseudophyllidea) is described from the gastrointestinal tract of Psenopsis anomala caught off the coast of Xiamen, China. The new species most closely resembles, but differs from, Parabothriocephalus segmentatus in its possession of muscular globular appendages on the posterior margin of the proglottids, a limited proglottid number (9-13), and the shorter strobila (7.6-13.2 mm vs. 165 mm). In addition, the uterus of P. psenopsis is strongly coiled, whereas that of P. segmentatus is S shaped; P. segmentatus has a spherical expansion in the middle of the vagina, whereas that of P. psenopsis does not. Finally, P. psenopsis differs from Parabothriocephalus gracilis. Parabothriocephalus sagitticeps, and Parabothriocephalus macruri by the posterolateral expansion of the proglottids. PMID- 15270111 TI - Torquatoides trogoni n. sp. and Excisa ramphastina n. sp. (Nematoda: Habronematoidea: Habronematidae) in birds from the area de Conservacion Guanacaste, Costa Rica. AB - Two new species of habronematid nematodes are described in birds from the Area de Conservacion Guanacaste, Costa Rica. Torquatoides trogoni n. sp., in Trogon massena, can be distinguished from T. torquata, T. bengalensis, and T. crotophaga in lacking lateral alae. Among species lacking lateral alae, the new species differs from T. balanocephala in having 14 versus 8-10 cephalic cuticular plaques, 21-22 versus 13-17 pairs of preanal papillae, and a beak-shaped versus U shaped gubernaculum. The new species differs from T. singhi in body length, in having 21-22 versus 10 pairs of precloacal papillae, longer spicules, and larger eggs. The new species differs from T. crotophaga, the only other species known from Central America, in lacking lateral alae, and having 14 versus 6 cephalic cuticular plaques, 21-22 versus 17 pairs of precloacal and 3 versus 2 pairs of postcloacal papillae, and a gubernaculum. Excisa ramphastina n. sp., in Ramphastos sulfuratus, can be distinguished from E. excisa, E. biloba, E. buckleyi, E. dentifera, and E. khalili in having 1 lateral ala versus none, cervical papillae anterior versus posterior to the nerve ring, and asymmetrical caudal alae. Excisa ramphastina is similar to E. curvata in having cervical papillae anterior to the nerve ring but differs in having 1 lateral ala versus none, asymmetrical caudal alae, an average spicule ratio of 1:4.4 versus 1:3.3, and 4 versus 2 pairs of sessile papillae. The new species differs from E. columbi in having 1 versus 2 lateral alae, in the length of the spicules, in having a different spicule ratio, and in the numbers of sessile papillae. PMID- 15270112 TI - Characterization, cloning, and expression of two diagnostic antigens for Taenia solium tapeworm infection. AB - Adult and larval stages of Taenia solium cause 2 diseases in humans, i.e., taeniasis and cysticercosis, respectively. Diagnosis and treatment of taeniasis are the ultimate means to eliminate cysticercosis. A serological taeniasis diagnostic test has been developed for laboratory use. However, recombinant forms of the taeniasis diagnostic proteins are required to overcome the limited supply of native proteins and allow the development of a low-cost and field-applicable test with high sensitivity and specificity. Using 2-dimensional electrophoresis of T. solium excretory and secretory (TSES) products from hamster adult tape-worm in vitro cultures, we have identified 5 T. solium-specific protein spots, with molecular weights of 33 kDa (protein isoelectrofocusing point [pI]: 5.6, 5.3, 5.1) and 38 kDa (pI: 4.6, 4.5). Protein sequencing and molecular cloning of these proteins showed that although endowed with different pls, the proteins with the same molecular weights shared the same protein backbone, named TSES33 and TSES38. Their full-length complementary DNAs encode proteins with 267 and 278 amino acids, respectively. TSES33 and TSES38 were expressed in a baculovirus system. Both recombinant proteins were recognized by a panel of taeniasis, but not cysticercosis patient serum samples, indicating that they can potentially replace the native proteins in the development of a more efficacious taeniasis diagnostic test. PMID- 15270113 TI - Efficacy of ponazuril in vitro and in preventing and treating Toxoplasma gondii infections in mice. AB - Toxoplasma gondii is an important apicomplexan parasite of humans and other warm blooded animals. Ponazuril is a triazine anticoccidial recently approved for use in horses in the United States. We determined that ponazuril significantly inhibited T. gondii tachyzoite production (P < 0.05) at 5.0, 1.0, or 0.1 microg/ml in African green monkey kidney cells. We used outbred female CD-1 mice to determine the efficacy of ponazuril in preventing and treating acute toxoplasmosis. Each mouse was subcutaneously infected with 1,000 tachyzoites of the RH strain of T. gondii. Mice were weighed daily, and ponazuril was administered orally in a suspension. Mice given 10 or 20 mg/kg body weight ponazuril 1 day before infection and then daily for 10 days were completely protected against acute toxoplasmosis. Relapse did not occur after prophylactic treatments were stopped. Toxoplasma gondii DNA could not be detected in the brains of these mice using polymerase chain reaction (PCR). One hundred percent of mice treated with 10 or 20 mg/kg ponazuril at 3 days after infection and then daily for 10 days were protected from fatal toxoplasmosis. Sixty percent of mice treated with 10 mg/kg ponazuril at 6 days after infection and 100% of mice treated with 20 mg/kg or 50 mg ponazuril 6 days after infection and then daily for 10 days were protected from fatal toxoplasmosis. Relapse did not occur after treatments were stopped. Toxoplasma gondii DNA was detected in the brains of some, but not all, of these mice using PCR. The results demonstrate that ponazuril is effective in preventing and treating toxoplasmosis in mice. It should be further investigated as a safe and effective treatment for this disease in animals. PMID- 15270114 TI - Seroprevalence of Toxoplasma gondii antibodies in wild dolphins from the Spanish Mediterranean coast. AB - Although Toxoplasma gondii infection has been found occasionally in cetaceans, little is known of the prevalence of antibodies to T. gondii in wild dolphins. Antibodies to T. gondii were determined in serum samples from 58 dolphins stranded in the Spanish Mediterranean coast. Modified agglutination test was used to determine T. gondii antibodies, and a titer of 1:25 was considered indicative of T. gondii infection. Antibodies to T. gondii were found in 4 of 36 striped dolphins (Stenella coeruleoalba), in 2 of 4 common dolphins (Delphinus delphis), in 4 of 7 bottlenose dolphins (Tursiops truncatus), and in 1 harbour porpoise (Phocoena phocoena). Antibodies were not found in 9 Risso's dolphins (Grampus griseus) and in 1 long-finned pilot whale (Globicephala melas) surveyed. The results indicate that T. gondii infection is frequent in at least 3 dolphin species from the Mediterranean Sea. PMID- 15270115 TI - Learedius learedi infection in black turtles (Chelonia mydas agassizii), Baja California Sur, Mexico. AB - Black turtle (Chelonia mydas agassizii) carcasses, recovered as a result of incidental capture in Magdalena Bay, Mexico, revealed invasion by spirorchiid trematode eggs in liver, kidney, intestines, muscle, heart, pancreas, and duodenum. Seventy-five adult Learedius learedi Price, 1934, were recovered from the heart of 1 turtle. Most of the organs showed a mild or absent inflammatory response in histological sections, with the exception of a pancreatic-duodenal section that revealed severe lymphocyte and phagocyte infiltration associated with an infestation of more than 200 eggs. A linear formation of 35 eggs from the pancreas toward the intestinal lumen is described as resembling migration. This is among the first reports of a parasitic infection of L. learedi Price 1934, in C. m. agassizii in Mexico. PMID- 15270116 TI - Determination of Trichuris skrjabini by sequencing of the ITS1-5.8S-ITS2 segment of the ribosomal DNA: comparative molecular study of different species of trichurids. AB - Adults of Trichuris skrjahini have been isolated from the cecum of caprine hosts (Capra hircus), Trichuris ovis and Trichuris globulosa from Ovis aries (sheep) and C. hircus (goats), and Trichuris leporis from Lepus europaeus (rabbits) in Spain. Genomic DNA was isolated and the ITS1-5.8S-ITS2 segment from the ribosomal DNA (rDNA) was amplified and sequenced by polymerase chain reaction (PCR) techniques. The ITS1 of T. skrjabini, T. ovis, T. globulosa, and T. leporis was 495, 757, 757, and 536 nucleotides in length, respectively, and had G + C contents of 59.6, 58.7, 58.7, and 60.8%, respectively. Intraindividual variation was detected in the ITSI sequences of the 4 species. Furthermore, the 5.8S sequences of T. skrjabini, T. ovis, T. globulosa, and T. leporis were compared. A total of 157, 152, 153, and 157 nucleotides in length was observed in the 5.8S sequences of these 4 species, respectively. There were no sequence differences of ITS1 and 5.8S products between T. ovis and T. globulosa. Nevertheless, clear differences were detected between the ITS1 sequences of T. skrjabini, T. ovis, T. leporis, Trichuris muris, and T. arvicolae. The ITS2 fragment from the rDNA of T. skrjabini was sequenced. A comparative study of the ITS2 sequence of T. skrjabini with the previously published ITS2 sequence data of T. ovis, T. leporis, T. muris, and T. arvicolae suggested that the combined use of sequence data from both spacers would be useful in the molecular characterization of trichurid parasites. PMID- 15270117 TI - Serological survey of Toxoplasma gondii infection among slaughtered pigs in northwestern Taiwan. AB - A serological survey of Toxoplasma gondii infection among slaughtered pigs in the largest slaughterhouse located in Taoyuan County of northwestern Taiwan was conducted using the latex agglutination (LA) test during 1998. The overall seroprevalence of T. gondii infection was 28.8% (32/111) with LA titers of 1:32 (6, 18.8%), 1:64 (10, 31.2%), 1:128 (9, 28.1%), 1:256 (6, 18.8%), and 1:512 (1, 3.1%). No significant difference (P > 0.05) in seroprevalence between male (28.6%, 20/70) and female (29.7%, 12/41) slaughtered pigs was observed. A decreasing trend in the seroprevalence among slaughtered pigs examined in the same slaughterhouse was observed because of a lower seroprevalence (P < 0.05) than that (44.4%, 128/288) previously reported about 10 yr ago using the LA test. Nevertheless, it is important to avoid eating raw or undercooked pork in order to prevent the acquisition of T. gondii infection among people in Taiwan. PMID- 15270118 TI - Infectivity of microsporidia spores stored in seawater at environmental temperatures. AB - To determine how long spores of Encephalitozoon cuniculi, E. hellem, and E. intestinalis remain viable in seawater at environmental temperatures, culture derived spores were stored in 10, 20, and 30 ppt artificial seawater at 10 and 20 C. At intervals of 1, 2, 4, 8, and 12 wk, spores were tested for infectivity in monolayer cultures of Madin Darby bovine kidney cells. Spores of E. hellem appeared the most robust, some remaining infectious in 30 ppt seawater at 10 C for 12 wk and in 30 ppt seawater at 20 C for 2 wk. Those of E. intestinalis were slightly less robust, remaining infectious in 30 ppt seawater at 10 and 20 C for 1 and 2 wk, respectively. Spores of E. cuniculi remained infectious in 10 ppt seawater at 10 and 20 C for 2 wk but not at higher salinities. These findings indicate that the spores of the 3 species of Encephalitozoon vary in their ability to remain viable when exposed to a conservative range of salinities and temperatures found in nature but, based strictly on salinity and temperature, can potentially remain infectious long enough to become widely dispersed in estuarine and coastal waters. PMID- 15270119 TI - Ticks (Acari: Ixodidae) parasitizing wild carnivores in Phu Khieo Wildlife Sanctuary, Thailand. AB - Ixodid ticks were collected and identified from 8 wild carnivore species in Phu Khieo Wildlife Sanctuary, northeastern Thailand. Six tick species belonging to 4 genera were recovered and identified from 132 individuals. These included Amblyomma testudinarium (n = 36), Haemaphysalis asiatica (n = 58), H. hystricis (n = 31), H. semermis (n = 3), Rhipicephalus haemaphysaloides (n = 3), and Ixodes granulatus (n = 1). Leopard cats (Prionailurus bengalensis) (n = 19) were infested with 4 tick species, whereas yellow-throated marten (Martes flavigula) (n = 4), clouded leopard (Neofelis nebulosa) (n = 2), and dhole (Cuon alpinus) (n = 1) were infested with 3 tick species, Asiatic golden cat (Catopuma temmincki) (n = 2) with 2 species, and marbled cat (Pardofelis marmorata), binturong (Arctictis binturong), and large Indian civet (Viverra zibetha) each infested with 1 species. This information contributes to the knowledge available on the ectoparasites of wild carnivores in Southeast Asia. PMID- 15270120 TI - First report of the giant kidney worm (Dioctophyme renale) in a harbor seal (Phoca vitulina). AB - A male harbor seal (Phoca vitulina) was found moribund on the coast of New Jersey in January of 2003 and died a few hours later in the Marine Mammal Stranding Center. On necropsy, a single female Dioctophyme renale was recovered from the peritoneal cavity, and a tissue mass was found adjacent to the pelvic urethra and urinary bladder. Within this tissue mass were found D. renale ova. This is the first report of this nematode in the harbor seal and in a North American marine mammal. PMID- 15270121 TI - Localization of a 56-kDa antigen that is present in multiple developmental stages of Neospora caninum. AB - The purpose of the present study was to characterize the intracellular distribution of a native Neospora caninum 56-kDa protein that is recognized by sera from N. caninum-infected dairy cattle. The complementary DNA coding for this protein was expressed in Escherichia coli as a polyHis fusion protein to which antiserum was prepared and used to localize the antigen in N. caninum tachyzoites and bradyzoites. By sodium dodecylsulfate-polyacrylamide gel electrophoresis and immunoblotting, antirecombinant Nc56 serum recognized a major 56-kDa protein and 2 minor (43 and 39 kDa) proteins of N. caninum tachyzoites. Antiserum to recombinant 56-kDa protein showed this antigen to be present in both N. caninum tachyzoites and bradyzoites/cysts as detected by immunofluorescence staining. Immunoelectron microscopy revealed the 56-kDa antigen to be present in the apical end of both tachyzoites and bradyzoites and possibly extracellularly secreted by tachyzoites. PMID- 15270122 TI - Occurrence of acanthocephalans in largemouth bass and smallmouth bass (Centrarchidae) from Gull Lake, Michigan. AB - A total of 65 largemouth bass, Micropterus salmoides, and 27 smallmouth bass, M. dolomieu, collected in April-September 2000 and April-July 2001 from Gull Lake, Michigan, were examined for acanthocephalans. Leptorhynchoides thecatus and Neoechinorhynchus cylindratus infected all the bass examined. Leptorhynchoides thecatus had the highest mean intensity (258.2 +/- 185.4 in 2000 and 145.0 +/- 61.0 in 2001) of the species infecting smallmouth bass. Although N. cylindratus had higher mean intensities (42.1 +/- 37.9 in 2000 and 68.9 +/- 70.5 in 2001) than did L. thecatus in largemouth bass, the values were not significantly different between bass species. The prevalence, mean intensity, and mean abundance of Pomphorhynchus bulbocolli in the bass species were below the values for the other acanthocephalan species. Leptorhynchoides thecatus and N. cylindratus are the most abundant intestinal helminths in bass from Gull Lake. PMID- 15270123 TI - Free-pool amino acids in Biomphalaria glabrata infected with Echinostoma caproni as determined by thin-layer chromatography. AB - Thin-layer chromatography was used to analyze the free-pool amino acids of the digestive gland-gonad complex (DGG) of Biomphalaria glabrata infected with Echinostoma caproni and uninfected (control) snails. Qualitative analysis revealed the presence of histidine, lysine, serine, alanine, valine, and isoleucine or leucine in all samples. Quantitative analysis of lysine and valine gave mean weight percentages of 0.00699 +/- 0.0022 and 0.00174 +/- 0.00056, respectively, in the DGG of uninfected snails, and 0.00504 +/- 0.0014 and 0.00254 +/- 0.00033, respectively, in the DGG of infected snails. The differences in values between infected and uninfected snails were not statistically significant (Student's t-test, P > 0.05). PMID- 15270124 TI - The epidemiological investigation of Trichinella infection in brown rats (Rattus norvegicus) and domestic pigs in Croatia suggests that rats are not a reservoir at the farm level. AB - Whether the brown rat (Rattus norvegicus) is a reservoir of Trichinella spp. infection or merely an accidental host, which may be vector of Trichinella spp., continues to be debated. We estimated the prevalence of Trichinella sp. infection in brown rat populations and in domestic pigs in 2 villages in Croatia, where Trichinella sp. infection in pigs has been endemic in the past 10 yr. Trichinella spiralis larvae, identified by a multiplex polymerase chain reaction analyses, were the only species detected in both rats and pigs. In 2001 and 2002, 2,287 rats were collected on 60 farms with different levels of sanitation and with, or without, T. spiralis-infected pigs. The prevalence of infection in rats ranged from 0.2 to 10.7%. Infected rats were detected only on farms with T. spiralis positive pigs and low sanitation or formerly with low sanitation (P = 0.007, Fisher's exact test), yet no infected rat was detected on farms with T. spiralis negative pigs. The finding that no infected rat was found on farms with T. spiralis-negative pigs suggests that, in the investigated area, the brown rat is not a reservoir but only a victim of improper pig slaughtering. PMID- 15270125 TI - High prevalence of Hepatozoon spp. (Apicomplexa, Hepatozoidae) infection in water pythons (Liasis fuscus) from tropical Australia. AB - Molecular methods were used to identify blood parasites frequently observed in blood smears of water pythons (Liasis fuscus) captured in our study area in the Northern Territory of Australia. A nested polymerase chain reaction (PCR) using primers amplifying the 18s ribosomal RNA (rRNA) nuclear gene resulted in a short PCR product (180 bp) matching this region in the genus Hepatozoon. However, because of the short sequence obtained. 2 new primers were designed based on 18s rRNA sequences of 3 Hepatozoon taxa available in GenBank. Using these primers, approximately 600 bp of the parasite's 18s rRNA gene was amplified successfully and sequenced from 2 water python samples. The new primers were used to investigate the prevalence of blood parasites in 100 pythons. In 25 of these samples we did not observe any blood parasites when examining stained slides. All the samples revealed a 600-bp PCR product, demonstrating that pythons in which we did not visually observe any parasites were infected by Hepatozoon spp. We also analyzed the nucleotide sequences of blood parasites in 4 other reptile taxa commonly encountered in our study area. The sequences obtained from water pythons and from 1 of these taxa were identical, suggesting that the parasite is capable of infecting hosts at different taxonomic levels. PMID- 15270126 TI - Glutamate-induced cytoplasmic Ca2+ transients in neurones isolated from the rat dorsal cochlear nucleus. AB - Extracellular application of glutamate elicited cytoplasmic Ca2+ transients in freshly dissociated rat neurones of the dorsal cochlear nucleus (DCN) (identified as pyramidal cells) with half-maximal concentration of 513 micromol/l while saturating doses (5 mmol/l) of this neurotransmitter caused transients of 46.1 +/ 3.0 nmol/l on an average. The genesis of these glutamate-evoked Ca2+ transients required extracellular Ca2+. When [Mg2+]o was 1 mmol/l, the NMDA receptor antagonist AP5 (100 micromol/l) had no effects while 100 micromol/l CNQX and 10 micromol/l NBQX, inhibitors of the AMPA receptors, greatly decreased the glutamate-induced Ca2+ transients (a decrease of 92 and 57%, respectively). When facilitating the activation of the NMDA receptors (50 micromol/l glycine, 20 micromol/l [Mg2+]o) in the presence of 100 micromol/l CNQX, Ca2+ transients of 55.4 +/- 13.1 nmol/l could be produced. Block of the voltage-gated Ca2+ channels (200 micromol/l Cd2+) decreased the Ca2+ transients to approx. 50%. The data indicate that under our control experimental circumstances the glutamate-induced Ca2+ transients of the isolated DCN neurones are produced mainly by Ca2+ entry through voltage-gated Ca2+ channels and AMPA receptors. However, when the activation of the NMDA receptors may take place, these receptors also contribute significantly to the genesis of the glutamate-evoked cytoplasmic [Ca2+] elevations. PMID- 15270127 TI - Different types of noise in leaky integrate-and-fire model of neuronal dynamics with discrete periodical input. AB - Different variants of stochastic leaky integrate-and-fire model for the membrane depolarisation of neurons are investigated. The model is driven by a constant input and equidistant pulses of fixed amplitude. These two types of signal are considered under the influence of three types of noise: white noise, jitter on interpulse distance, and noise in the amplitude of pulses. The results of computational experiments demonstrate the enhancement of the signal by noise in subthreshold regime and deterioration of the signal if it is sufficiently strong to carry the information in absence of noise. Our study holds mainly to central neurons that process discrete pulses although an application in sensory system is also available. PMID- 15270128 TI - The effect of Pycnogenol on the erythrocyte membrane fluidity. AB - In the present study, the in vitro effect of polyphenol rich plant extract, flavonoid--Pycnogenol (Pyc), on erythrocyte membrane fluidity was studied. Membrane fluidity was determined using 1-[4-trimethyl-aminophenyl]-6-phenyl-1,3,5 hexatriene (TMA-DPH), 1,6-diphenyl-1,3,5-hexatriene (DPH) and 12-(9-anthroyloxy) stearic acid (12-AS) fluorescence anisotropy. After Pyc action (50 microg/ml to 300 microg/ml), we observed decreases in the anisotropy values of TMA-DPH and DPH in a dose-dependent manner compared with the untreated erythrocyte membranes. Pyc significantly increased the membrane fluidity predominantly at the membrane surface. Further, we observed the protective effect of Pyc against lipid peroxidation, TBARP generation and oxidative hemolysis induced by H2O2. Pyc can reduce the lipid peroxidation and oxidative hemolysis either by quenching free radicals or by chelating metal ions, or by both. The exact mechanism(s) of the positive effect of Pyc is not known. We assume that Pyc efficacy to modify effectively some membrane dependent processes is related not only to the chemical action of Pyc but also to its ability to interact directly with cell membranes and/or penetrate the membrane thus inducing modification of the lipid bilayer and lipid-protein interactions. PMID- 15270129 TI - Application of oscillating potentials to the Shaker potassium channel. AB - Nonequilibrium response spectroscopy (NRS) has been proposed recently to complement standard electrophysiological techniques used to investigate ion channels. It involves application of rapidly oscillating potentials that drive the ion channel ensemble far from equilibrium. It is argued that new, so far undiscovered features of ion channel gating kinetics may become apparent under such nonequilibrium conditions. In this paper we explore the possibility of using regular, sinusoidal voltages with the NRS protocols to facilitate Markov model selection for ion channels. As a test case we consider the Shaker potassium channel for which various Markov models have been proposed recently. We concentrate on certain classes of such models and show that while some models might be virtually indistinguishable using standard methods, they show marked differences when driven with an oscillating voltage. Model currents are compared to experimental data obtained for the Shaker K+ channel expressed in mammalian cells (tsA 201). PMID- 15270130 TI - Binding of imipramine to phospholipid bilayers using radioligand binding assay. AB - Binding of the tricyclic antidepressant imipramine (IMI) to neutral and negatively charged lipid membranes was investigated using a radioligand binding assay combined with centrifugation or filtration. Lipid bilayers were composed of brain phosphatidylcholine (PC) and phosphatidylserine (PS). IMI binding isotherms were measured up to IMI concentration of 0.5 mmol/l. Due to electrostatic attraction, binding between the positively charged IMI and the negatively charged surfaces of PS membranes was augmented compared to binding to neutral PC membranes. After correction for electrostatic effects by means of the Gouy Chapman theory, the binding isotherms were described both by surface partition coefficients and by binding parameters (association constants and binding capacities). It was confirmed that binding of IMI to model membranes is strongly affected by negatively charged phospholipids and that the binding is heterogeneous; in fact, weak surface adsorption and incorporation of the drug into the hydrophobic core of lipid bilayer can be seen and characterized. These results support the hypothesis suggesting that the lipid part of biological membranes plays a role in the mechanism of antidepressant action. PMID- 15270131 TI - Ultrastructural morphometry of precompacted bovine embryos produced in vivo and in vitro after activation by electric pulse AC/DC. AB - Early bovine precompacted embryos (at 1- to 8-blastomere stage) were analyzed by electron microscopy. The volume density of cellular components was determined by morphometric analysis to quantify the ultrastructure of early bovine embryos produced either in vivo or parthenogenetically after stimulation of oocytes by electric pulse AC/DC. In embryos obtained in vivo, most of cellular volume was occupied by cytoplasm (82.93%). The relative volume of lipids, vacuoles, mitochondria was relatively low (5.46; 5.07; 3.78%, respectively), and the relative volume of Golgi apparatus and cell inclusions was the lowest (1.51%). AC/DC-derived parthenogenotes had a relative high area occupied by vacuoles and lipids (18.68 vs. 14.33%) and a significantly lower relative volume was occupied by cytoplasm (60.63%) when compared with the control in vivo embryos. These observations demonstrated that parthenogenetic embryos had significantly altered ultrastructure, indicating extensive subcellular damages. These findings are discussed from the physiological and functional point of view. PMID- 15270132 TI - Effect of cholesterol on the bilayer thickness in unilamellar extruded DLPC and DOPC liposomes: SANS contrast variation study. AB - Small-angle neutron scattering on extruded unilamellar vesicles in water was used to study bilayer thickness when cholesterol (CHOL) was added to dilauroylphosphatidylcholine (DLPC) and dioleoylphosphatidylcholine (DOPC) bilayers in molar fraction 0.44. Using the H2O/2H2O contrast variation and the small-angle form of Kratky-Porod approximation, the bilayer gyration radius at infinite contrast R(g,infinity) and the bilayer thickness parameter d(g,infinity) = 12(0.5)R(g,infinity) were obtained at 25 degrees C. Addition of CHOL to DLPC increased the d(g,infinity) from 4.058 +/- 0.028 nm to 4.62 +/- 0.114 nm, while in case of DOPC the d(g,infinity) values were the same in the absence (4.618 +/- 0.148 nm) and in the presence (4.577 +/- 0.144 nm) of CHOL within experimental errors. The role of CHOL-induced changes of bilayer thickness in the protein insertion, orientation and function in membranes is discussed. PMID- 15270133 TI - Tort reform under attack--AMS and AMA to intervene. PMID- 15270134 TI - Emotional, uninformed juries are dangerous. PMID- 15270135 TI - The effect of retrograde femoral nailing on residual femoral growth in a sheep model. PMID- 15270136 TI - School entry immunization: 15% of students lack complete vaccination protection. PMID- 15270137 TI - Oral health of low-income school children: self assessment. PMID- 15270138 TI - Drug-induced long QT syndrome and Torsade de Pointes. PMID- 15270139 TI - Are antidepressants safe to use in children and adolescents? AB - The Food and Drug Administration is investigating 10 antidepressants to determine possible increase of depression or suicidality in children and adolescents while on the medication. Results of the data analysis should be available in the summer of 2004. PMID- 15270140 TI - A clearing in the forest: the wondrous genome. PMID- 15270141 TI - (Sub)standard of care. PMID- 15270142 TI - Message questioned. PMID- 15270143 TI - Salient points. PMID- 15270144 TI - Blatant bias. PMID- 15270145 TI - Lasers in dentistry. PMID- 15270146 TI - Laser article 'balanced'. PMID- 15270147 TI - Laser article 'a disservice'. PMID- 15270148 TI - Laser article supported. PMID- 15270149 TI - Portfolios for initial licensure. PMID- 15270150 TI - 'Oversimplifying' licensure? PMID- 15270151 TI - More about licensure. PMID- 15270152 TI - Drawing conclusions. PMID- 15270153 TI - Laboratory technology. PMID- 15270154 TI - Do you see any conflict between the desire to provide high-quality dental care and the desire to run an efficient, profitable business? PMID- 15270155 TI - Motivating parents to prevent caries in their young children: one-year findings. AB - BACKGROUND: The authors conducted a study to compare the effect of a motivational interviewing counseling treatment with that of traditional health education on parents of young children at high risk of developing dental caries. OVERVIEW: The authors enrolled in the study parents of 240 infants aged 6 to 18 months and randomly assigned them to either a motivational interviewing, or MI, group or a traditional health education (control) group. Parents in the control group received a pamphlet and watched a video. Parents in the MI group also received the pamphlet and watched the video; in addition, they received a personalized MI counseling session and six follow-up telephone calls. RESULTS: After one year, children in the MI group had .71 new carious lesions (standard deviation, or SD, = 2.8), while those in the control group had 1.91 (SD = 4.8) new carious lesions (t[238] = 2.37, one-tailed, P < .01). CONCLUSIONS: MI is a promising approach that should receive further attention. CLINICAL IMPLICATIONS: MI may lead parents and others to better accept dental recommendations about preventing caries in their children. PMID- 15270156 TI - Abutment tooth loss in patients with overdentures. AB - BACKGROUND: Since the 1960s, the use of natural teeth as overdenture abutments has become part of accepted clinical practice. Several longitudinal studies have been conducted, but tooth loss has not been reported to be a significant problem. The aim of this study was to identify the incidence and causes of tooth loss in a prospective cohort study of subjects wearing overdentures. METHODS: The study, conducted between 1973 and 1994, evaluated 273 subjects (62.3 percent male) with a mean age of 59.6 years. RESULTS: Of the 273 subjects with 666 abutments, 74 lost 133 abutments. The most common cause of tooth loss was periodontal disease (29.3 percent) followed by periapical lesions (18.8 percent) and caries (16.5 percent). Through logistic regression, the authors found that subjects who lost teeth were more likely to have medical problems that could cause soft-tissue lesions of the oral mucosa, were less likely to use fluoride daily and were less likely to return for yearly recall visits. The authors found 22 vertical fractures in 17 subjects. Chi2 analysis revealed that overdenture teeth in the maxillary arch that were opposed by natural teeth were more likely to experience vertical fractures. CONCLUSIONS: In a study that followed up some patients for as long as 22 years, the rate of tooth loss was 20.0 percent. Many of these failures could have been prevented if patients had practiced better oral hygiene. CLINICAL IMPLICATIONS: The findings suggest that if a dentist recommends overdenture therapy, the patient needs to be examined regularly to reduce the risk of experiencing caries and periodontal disease. Also, if the abutments are in the maxilla and are opposed by natural teeth, the dentist should consider using thimble crowns to reduce the risk of vertical fractures. PMID- 15270157 TI - Conducting systematic reviews and creating clinical practice guidelines in dentistry: lessons learned. AB - BACKGROUND: High-quality systematic reviews are the basis of valid, reliable clinical practice guidelines, or CPGs. In 1999, a Canadian collaboration of dentists embarked on the process of developing guidelines. METHODS: The Canadian Collaboration on Clinical Practice Guidelines in Dentistry, or CCCD, is a coalition of multiple stakeholders from organized dentistry and academia whose mandate is to develop CPGs for practicing dentists. In the development of the first CPG based on a systematic review of the literature, the CCCD Methodology Resource Group (of which the authors were co-chairs) gained some valuable insights. The authors wrote this article to share their experiences and lessons learned and to offer practical advice to others who may undertake similar projects. RESULTS: The authors identify a number of methodological issues and logistical problems and make suggestions for effective management of the review and guideline development processes. CONCLUSIONS AND PRACTICE IMPLICATIONS: Systematic reviews and the development of CPGs require rigorous methodology, as well as input from content experts and clinicians, to ensure validity and relevance. The processes are costly and time-intensive, but the anticipated outcome is enhanced clinical decision making and improved oral health. PMID- 15270158 TI - Squamous cell carcinoma arising in an oral lichenoid lesion. AB - BACKGROUND: Oral lichen planus, or OLP, is a chronic inflammatory mucocutaneous disease that frequently involves the oral mucosa. Lichenoid dysplasia, or LD, refers to lesions that could be mistaken clinically for OLP but have histologic features of dysplasia and a true malignant predisposition. Published case reports of OLP conversion to squamous cell carcinoma, or SCC, have created a great deal of controversy about the true nature of OLP, highlighting the need to verify its clinical diagnosis histologically. CASE DESCRIPTION: The authors document the development of SCC in a 58-year-old woman with an oral lesion diagnosed clinically as OLP and described histologically as having lichenoid features with dysplastic changes. The time from the initial diagnosis of oral lichenoid lesions to the patient's return visit to the medical center with clinically evident cancer was three years and eight months. The SCC developed in the labial mucobuccal fold and left mandibular edentulous ridge, which had undergone multiple biopsy procedures. CLINICAL IMPLICATIONS: This case does not provide answers to the ongoing controversy about the innate propensity of OLP to become malignant. However, in view of both the common occurrence of OLP and unresolved issues regarding its premalignant potential, this case report illustrates the need for histologic confirmation and close follow-up of patients with clinical lesions that have lichenoid features. PMID- 15270159 TI - Conservative treatment of the Class I lesion: a new paradigm for dentistry. AB - BACKGROUND: A shift is occurring in dentistry that involves a change from reliance on gross mechanical instrumentation of dental caries to early diagnosis and treatment of the bacterial infection that causes caries. TYPES OF STUDIES REVIEWED: The author explores the topic of minimally invasive dentistry, and cites several studies that offer scientific evidence of the effectiveness of this approach. The author also examines the role of third-party payers, who are reluctant to provide reimbursement for sealants or treatment of incipient caries. CONCLUSIONS: As dentists embrace a new paradigm in the treatment of the Class I lesion, they are beginning to acknowledge their role as clinical cariologists with the means to accurately assess the extent and threat of existing disease, determine the appropriate clinical response, provide minimally invasive treatment and unambiguously describe services rendered. CLINICAL IMPLICATIONS: As evidence based protocols become more widely accepted, dentistry will have the necessary tools to interact with third parties, which also are struggling to cope with and adapt to an emerging standard of care. PMID- 15270160 TI - Using a time-saving form to document emergency assessments. PMID- 15270161 TI - Is occlusion becoming more confusing? A plea for simplicity. AB - It is not difficult to observe and record patient occlusal characteristics before starting simple or complex occlusal rehabilitations. If this is done, and if the subsequently placed crowns and fixed prostheses are constructed in observation of similar characteristics, clinical success usually is the result. Deviations from the suggestion to duplicate the "normal" occlusion should be made when the original natural occlusion had caused overt pathosis, or when all teeth or one arch of the teeth is being restored at one time. If this is the case, centric relation occlusion is more reproducible and easier to develop than occlusion with a shift from centric relation to centric occlusion. Peculiar requests of patients relative to occlusal positioning, or routine dependence on various devices to predetermine occlusal characteristics for rehabilitation (as is currently popular in some groups), should be considered, but they should be tempered with careful observation of preoperative occlusal characteristics. PMID- 15270162 TI - Panic disorder: psychopathology, medical management and dental implications. AB - BACKGROUND: This article reviews the clinical features, epidemiology, pathophysiology, dental findings, and dental and medical management of the care of patients with panic disorder, or PD. TYPES OF STUDIES REVIEWED: The authors conducted a MEDLINE search for the period 1998 through 2003, using the key term "panic disorder" to define the pathophysiology of the disorder, its epidemiology and dental implications. The articles they selected for further review included those published in peer-reviewed journals. RESULTS: PD is a common and debilitating psychiatric disease in which a person experiences sudden and unpredictable panic attacks, or PAs, with symptoms of overwhelming anxiety, chest pain, palpitations and shortness of breath. Persistent concern about having another attack and worry that it may indicate a heart attack or "going crazy" impairs the person's social, family and working lives. Frequently accompanying the disorder is agoraphobia, depression and mitral valve prolapse, or MVP. CLINICAL IMPLICATIONS: In patients with PD, the prevalence of dental disease may be extensive because of the xerostomic effects of psychiatric medications used to treat it. Dental treatment consists of preventive dental education and prescribing saliva substitutes and anticaries agents. Precautions must be taken when prescribing or administering analgesics, antibiotics or sedative agents that may have an adverse interaction with the psychiatric medications. Because there is a connection between PAs and MVP, the dentist needs to consult with the patient's physician to determine the presence of MVP and whether there is associated mitral valve regurgitation. Patients with MVP and accompanying mitral valve regurgitation require prophylactic antibiotics when undergoing dental procedures known to cause a bacteremia and heightened risk of endocarditis. PMID- 15270163 TI - Access to dental care: the triad of essential factors in access-to-care programs. AB - BACKGROUND: Solutions to the problem of limited access to dental care for some segments of the population are being sought. Certain factors in access-to-dental care program design are critical to the success of these programs. OVERVIEW: The author discusses the triad of factors that are essential in the successful design and operation of programs aimed at improving access to dental care for underserved populations. CONCLUSIONS: The author found that attention must be paid to creating an effective demand for dental care, an adequate dental work force to respond to that demand and an economic environment that enables patients and dentists to participate in any programs that are aimed at improving access to care. PRACTICE IMPLICATIONS: Failure to adequately address demand, work force and economic factors will condemn any program designed to increase access to dental care to failure at worst, or to limited success at best. PMID- 15270164 TI - Inventory management. AB - As dentistry continues to evolve, the best management systems of the business world need to be incorporated into each practice. As always, my goal in these columns is to bring and modify the best business principles available to readers of The Journal of the American Dental Association. Just in Time ordering and inventory control is one of the best, as evidenced by the fact that top performing companies worldwide have adopted it. PMID- 15270165 TI - Stress, burnout, anxiety and depression among dentists. AB - BACKGROUND: Dentists encounter numerous sources of professional stress, beginning in dental school. This stress can have a negative impact on their personal and professional lives. CONCLUSIONS: Dentists are prone to professional burnout, anxiety disorders and clinical depression, owing to the nature of clinical practice and the personality traits common among those who decide to pursue careers in dentistry. Fortunately, treatment modalities and prevention strategies can help dentists conquer and avoid these disorders. PRACTICE IMPLICATIONS: To enjoy satisfying professional and personal lives, dentists must be aware of the importance of maintaining good physical and mental health. A large part of effective practice management is understanding the implications of stress. PMID- 15270166 TI - A simulated-use evaluation of a strategy for preventing biofilm formation in dental unit waterlines. AB - BACKGROUND: Prevention of biofilm formation is important in the maintenance of dental unit waterline systems. Without effective control measures, the waterlines will become contaminated with routine use. METHODS: The authors used a simulated use dental unit waterline system to evaluate the ability of a test product, A-dec ICX (A-dec, Newburg, Ore.), to prevent biofilm formation. They evaluated buffered distilled water and hard water models versus mixed-challenge suspensions of Staphylococcus aureus, Klebsiella pneumoniae and Pseudomonas aeruginosa. RESULTS: The authors documented development of significant biofilm in untreated test units, while treated test units showed no indication of biofilm formation throughout the 16-week study. Student t tests and 95 percent confidence intervals performed on the plate count data confirmed that untreated test units had significantly greater bacterial populations than did treated test units (P < .05). Qualitative images by scanning electron microscopy verified these findings. CONCLUSION: In this simulated clinical-use study, the test product effectively reduced bacterial counts in incoming water and produced water quality exceeding stated recommendations of the American Dental Association. CLINICAL IMPLICATIONS: The test product provides an approach to dental unit waterline maintenance that is simple to use and that, by continuously preventing biofilm formation, reduces concerns about bacterial contamination in dental unit water. PMID- 15270167 TI - Court upholds final version of HIPAA privacy regulations. PMID- 15270168 TI - For the dental patient... Have a checkup before you travel. PMID- 15270169 TI - Smash and grab. PMID- 15270171 TI - The HSJ interview: Sir Jeremy Beecham. One chair for democracy. Interview by Nick Edwards. PMID- 15270172 TI - Clinical management. Where medicine meets management. Reduced circumstances. AB - The development of effective clinical teams is vital to the delivery of quality clinical care and reducing clinical errors. Implementation of the European working-time directive will disrupt team dynamics and reduce the training opportunities for junior doctors. Funding is needed to increase further the number of doctors if the directive is not to be at the expense of patient care. PMID- 15270173 TI - Finance. Acute little idea. PMID- 15270174 TI - Sterilisation services. Knife point. AB - Sterile services departments are in such a poor state that new centres are needed. A super-centre, shared between local trusts, is to be tried in Bradford and Leeds. Stocks and movement of instruments will need close monitoring under a centralised system. PMID- 15270175 TI - My brilliant career--estates and facilities management. Estate of mind. PMID- 15270176 TI - HSJ people. Planner in the works. PMID- 15270177 TI - Health care expenses in retirement and the use of health savings accounts. PMID- 15270178 TI - Emergency medicine: a relatively new specialty. PMID- 15270179 TI - Emergency section and overcrowding in a university hospital of Karachi, Pakistan. AB - OBJECTIVE: To quantify the extent of Emergency Section overcrowding at a tertiary care hospital and to identify possible solutions. METHODS: This was a retrospective chart review of all the patients presented to the Aga Khan University Hospital's (AKUH) Emergency Section (ES). The ES information system has an automatic patient log and generates daily report for patients who stay longer than 6 hours. The charts of all patients who stayed longer than 6 hours were reviewed. RESULTS: Among 9630 patients, 1999 (20.8%) were held in the ES for more than 6 hours. Of those 134 (6.7%) were discharged from the ES, while 1535 (76.8) were admitted to the hospital. About two-third of all delays were due to unavailability of bed, followed by financial constraints, involvement of multiple specialty, and because the admitting residents wanted to investigate the patients more thoroughly. CONCLUSION: Significant overcrowding exists in Emergency Section of the hospital. Four solutions were proposed: (1) early discharges of in patients, (2) creation of a holding unit, (3) flexible ward assignment, (4) active inter-facility transfer. These efforts will lead to an optimal care in the Emergency Section in rising patient demand. PMID- 15270180 TI - Bacterial infections in paediatric patients with chemotherapy induced neutropenia. AB - OBJECTIVE: To determine the pattern of bacterial infections, isolate and identify the pathogenic bacteria and their sensitivity to different antibiotics during febrile episodes in paediatric patients with chemotherapy induced neutropenia from January to June 2000 at the Paediatric OncologyUunit of Combined Military Hospital, Rawalpindi (CMH RWP). PATIENTS AND METHODS: The study material comprised of 62 febrile episodes occurring in 50 neutropenic children aged less than 12 years with various malignancies. All the episodes were worked up in detail including history, physical examination and relevant investigations. RESULTS: Total 29 bacteria were cultured in 62 febrile episodes. Fifty five percent organisms were isolated from blood and 45% from other sites, 15 (51.7%) were Gram-positive and 14 (48.3%) were Gram-negative. S. aureus was the most frequent Gram positive isolate and E. coli was the most common Gram negative isolate. The standard empiric antibiotic regimens for (combination of amikacin and ceftazidime) showed an overall response rate of 61.3%. The infection related mortality in this series was 22%. CONCLUSION: Fever is the commonest symptom of infection in neutropenic children with malignancy and demands an urgent empirical antibiotic therapy after the onset of fever. Based on this study we recommend a combination of ceftazidime and amikacin for use as empiric antibiotic therapy in these children. PMID- 15270181 TI - Absorption of non heme iron in typical and standard meals using extrinsically labeled iron 59Fe. AB - OBJECTIVE: To study the bioavailibility of nonheme iron from standard and test meals in Pakistani adults and to determine iron absorption using extrinsically tagged iron 59Fe. SUBJECTS AND METHODS: Nonheme iron absorption was measured from standard and test meals in ten healthy individuals. Total calories, carbohydrates, proteins, fats, ascorbic acid, total iron, phytate and ascorbic acid content were determined in both meals. Retention of iron was detected by whole body counting using gamma counter before and after administration of standard, test meals and reference dose. RESULTS: Iron absorption with test meal was 13% and after adjustment with serum ferritin and reference dose was 16.5 % and 14% respectively. The absorption of standard meal was 6.7% which after adjustment with serum ferritin and reference dose was 8.8% and 6.9% respectively. The iron and ascorbic acid content of test meal was 6.5 mg and 5.7 mg respectively while phytate phosphorus content was 114 mg. The iron and ascorbic acid content of standard meal was 1.3 mg and 2.4 mg respectively while phytate phosphorus content was 137 mg. CONCLUSION: This evaluated absorption from one of the typical Pakistani diet compared to the standard meal was better. This shows that there are some other physiological factors that lead to iron deficiency anemia in Pakistan. PMID- 15270182 TI - Polycythemia vera and idiopathic erythrocytosis: comparison of clinical and laboratory parameters. AB - OBJECTIVE: To evaluate the various clinical and laboratory parameters of Polycythemia vera and idiopathic erythrocytosis in order to differentiate between two entities at the Aga Khan University Hospital. METHODS: Twenty six patients of polycythemia vera and 34 patients of idiopathic erythrocytosis were analyzed with respect to clinical features and laboratory findings. RESULTS: Patients with idiopathic erythrocytosis were males with a mean age of 41 years and no splenomegaly. Patients with polycythemia were older males and females with splenomegaly, red cell count of mor than 6.5 million/cmm, haematocrit 55%, leucocytosis, thrombocytosis and low erythropoietin level. CONCLUSION: Based on the above-mentioned findings, we suggest that polycythemia vera and idiopathic erythrocytosis are separate entities and the diagnosis of these can be made on the basis of clinical and laboratory parameters. PMID- 15270183 TI - Increased expression of HLA DR2 in acquired aplastic anemia and its impact on response to immunosuppressive therapy. AB - OBJECTIVE: To study the frequency of HLA DR2 status of patients with aplastic anemia and their response to immunosuppressive therapy at a tertiary care hospital. METHODS: Thirty eight consecutive patients of acquired aplastic anemia were evaluated with respect to demographic features, severity of HLA DR2 status and response outcome to immunosuppressive therapy. RESULTS: The mean age of the patients was 24.6 years + 16.4 with a male to female ratio of 2.8:1. Positivity of HLA DR2 was markedly high in acquired aplastic anemia patients. Twenty four (65%) out of 38 patients as compared to 45 (15%) of 300 healthy controls (p<0.0001) were positive for HLA DR2. Response to immunosuppressive therapy, which included antilymphocyte globulin, cyclosporin and methylprednisolone, was available in sixteen HLA DR2 positive patients and was found satisfactory in 12/16 (75%) patients. CONCLUSION: HLA DR2 was significantly higher in patients with acquired aplastic anemia and favourable response to immunosuppressive therapy was also associated with HLA DR2 positivity. PMID- 15270184 TI - Increased levels of erythrocyte glutathione in acute myocardial infarction: an antioxidant defence. AB - OBJECTIVE: Glutathione (GSH) has a central role in the defence against oxidative damage. This study was carried out to investigate any change in erythrocyte GSH levels in a population of patients with acute myocardial infarction (AMI) and compare them with levels in normal healthy subjects. METHOD: GSH levels were determined in erythrocytes of one hundred and seventy six patients with AMI (age: 30-70 years; 131 males and 45 females) admitted to the National Institute of Cardiovascular Diseases, Karachi. These levels were compared with eryrocyte GSH levels obtained from 95 normal healthy subjects (controls). RESULTS: Mean +/- SD erythrocyte GSH levels in AMI patients and controls were found to be 2.34 +/- 0.62 micromol/ml of packed cells and 2.08+/- 0.62 micromol/ml of packed cells, respectively. The two values when compared with one way ANOVA were found to be significantly different (p=0.001). Age had little effect on erythrocyte GSH levels in both AMI patients and normal healthy subjects. CONCLUSION: Increased production of reactive oxygen species is a feature of cardiovascular disease, such as AMI and cells can respond to mild oxidative stress by upregulating antioxidant defence in terms of increased production of GSH. PMID- 15270185 TI - Coronary artery diameter in a cohort of adult Pakistani population. AB - OBJECTIVE: To study the diameter of coronary arteries in a cohort of adult Pakistani population and to compare these with the diameters of Caucasians mentioned in the literature. METHODS: A study of 220 adult patients referred to National Institute of Cardiovascular Disease (NICVD) for diagnostic coronary angiography between May 2000 to December 2000. RESULTS: The mean diameter of Right Coronary Artery (RCA) was found to be 3.08 + 0.78 mm with a 95% CI of 2.9 - 3.2, and of Left Main Coronary Artery (LMCA), 4.28 +/- 0.82 with a 95% CI of 4.2 4.4. While the mean diameter of Left Anterior Descending Artery (LAD) was 3.22 +/- 0.74 with a 95% CI of 3.1 - 3.33, and that of the Circumflex Artery (CX) 3.02 + 0.75 with a 95% CI 2.9 - 3.1. The total coronary area (TCA), diameter of three vessels was 9.32 + 1.68 with a 95% CI of 9.1 - 9.5, while the sum of 4 vessels diameter was 13.6 + 2.26 with a 95% CI of 13.3 - 13.9. CONCLUSION: The diameters of coronary arteries of Pakistani population are not significantly different from that of Caucasians and the cause of increased mortality in the people of South Asians origin seems to be other than the diameter of coronary arteries. PMID- 15270186 TI - Risk factors and behaviours for coronary artery disease (CAD) among ambulatory Pakistanis. AB - OBJECTIVE: To determine the frequency and distribution of various risk factors and behaviours for coronary artery disease (CAD) among ambulatory Pakistanis. METHODS: It is a cross-sectional descriptive study carried out at the Aga Khan University Hospital, a teaching hospital in Karachi. All the subjects were adults (18-60 years) presenting at the general checkup clinic with no history or evidence of CAD by convenient sampling method. Demographic variables included risk factors and behaviors including diabetes, hypertension, dyslipidemia, family history of heart disease, obesity, smoking and sedentary lifestyle. RESULTS: Among 370 ambulatory Pakistanis, the proportions of major risk factors for CAD were: sedentary life style 72%, family history 42%, dyslipidemia 31%, obesity 24%, hypertension 19% and diabetes mellitus 15%. Diabetes, hypertension and dyslipidemia were poorly controlled in the study population. Proportions of the three major risk factors (smoking, hypertension and dyslipidemia) occurring singly, doubly and all three together in the study population were found to be 39%, 11% and 1%, respectively. Data were also analyzed for risk factors by comparing those with and without family history of CAD to eliminate any bias. The results were not statistically significant except for the sedentary life style (P=0.016). CONCLUSION: There is a high prevalence of CAD risk factors in this study population. Modifiable risk factors like diabetes, hypertension and high cholesterol need better control. Preventive screening programs and healthy lifestyle behaviours need to be emphasized upon in the community. PMID- 15270187 TI - Fine needle aspiration of unilocular ovarian cysts--a cytohistological correlation. AB - OBJECTIVE: To assess the diagnostic efficacy of Fine Needle Aspiration Cytology in differentiating neoplastic and non-neoplastic dysfunctional ovarian cysts- cytological findings to be verified with histology of excised cyst. METHODS: In this prospective study fifty-three cases with unilocular nonseptate ovarian cystic masses,detected on ultrasound examination were subjected to ultrasound guided fine needle aspiration of the cyst contents at Department of Pathology, Allama Iqbal Medical College Lahore, from January 1999 - January 2000. Aspirated fluid was examined cytologicaly using Giemsa stain on the smears prepared from centrifuged deposit. The same cyst removed surgically was examined histologicaly and cytohistological correlation was carried out. RESULTS: In this series of 53 cases, histologically confirmed break up of various types of cysts was follicular cysts (n=25), leuteal cysts (n=6), serous cysts (n=15), mucous cysts (N=4) and Endometriotic cysts (n=3). Non-diagnostic fine needle aspirate was obtained in 19/53 cases, majority being the follicular cysts. The cytohistological correlation revealed no false positive but 40% false negative results for follicular cysts on cytological examination of the aspirate. Hence the specificity and sensitivity for cytological diagnosis of follicular cyst was 100% and 60% respectively. For leuteal cysts, false positive and false negative results on cytological examination were 0% and 16.6% respectively with a specificity and sensitivity value of 100% and 83% respectively. For neoplastic serous cysts cytologically false positive and false negative diagnosis was 0% and 46.6% respectively with specificity and sensitivity of 100% and 53%. For mucinous cystadenomas sensitivity and specificity of cytological diagnosis was 100%. For endometriotic cysts a sensitivity of 67% and specificity of 100% was procured with cytological evaluation. CONCLUSION: Guided fine needle aspiration cytology may prove to be one of the most valuable and acceptable tools in the differential diagnosis of ovarian cystic lesions. PMID- 15270188 TI - Self-expense among outpatient department patients at a public hospital in Karachi, Pakistan. PMID- 15270189 TI - Neurology in the 21st century: contemporary state of diagnostics and therapeutics. PMID- 15270190 TI - Emergency use of Eschlmann Stylet (tracheal tube introducer) in acute tracheo bronchial obstruction during general anesthesia. PMID- 15270191 TI - Gastrointestinal stromal tumours: a case report and recent concepts. PMID- 15270192 TI - Orbital cellulitis masquerading as cavernous sinus thrombosis--a case report. PMID- 15270193 TI - Mammary echinococcosis: two cases and literature review. PMID- 15270194 TI - Pulmonary infiltrates during chemotherapy-induced febrile neutropenia: incidence, patterns and outcomes. AB - OBJECTIVE: To analyze the incidence, etiologies, radiographic patterns, and clinical outcomes of adult leukemics with prolonged febrile neutropenia and pneumonia. METHODS: A retrospective study was conducted at a tertiary care hospital. The medical records of adult patients with acute myeloid leukemia diagnosed between January 1989 and June 2000 and undergoing induction chemotherapy were included. Only the patients who presented with a pulmonary infiltrate, secondary leukemia (e.g., transformed chronic myeloid leukemia underlying myelodysplastic syndrome, or disease following alkylating agent therapy) were included and those developing infiltrates following consolidation chemotherapy were excluded. RESULTS: A total of 124 patients were admitted to the hospital with a diagnosis of AML during the study period. Thirty-one patients were excluded; 93 patients received induction chemotherapy and were included in the study analysis. The median age was 36 years (15 - 70 years); 58 males and 35 females. Sixty two percent patients received Cytosine Arabinoside (Ara-C), 17% received Etoposide, 11% received Ara-C and Mitoxantrone, and 6% received All trans-retinoic Acid. The mean onset and duration of neutropenia were 5 and 15 days, respectively. Pulmonary infiltrates were identified during 45% of neutropenic episodes. A presumptive causative organism was isolated from 50% of patients with an infiltrate: Gram-positive bacteria were most common (47%) followed by Gram-negative bacilli (33%) and fungi (20%). Survival data were available for 88 patients; median disease free survival for the entire cohort was 7 months. Male sex (p=0.015), onset of neutropenia (p=0.02) and bilateral distribution of an infiltrate (p=0.03) were statistically significant predictors of early mortality. For patients with and without pneumonia, the median disease free interval and overall survival were 2.5 and 4.6 months and 9 and 13 months (p=0.038 and p=0.095) respectively. CONCLUSION: Neutropenia occurred at a mean of 5.0 after initiation of induction chemotherapy. The majority of patients had bilateral pulmonary infiltrates. Male sex, onset of neutropenia and bilateral distribution of an infiltrate were found to be statistically significant predictors of early mortality. PMID- 15270195 TI - Statins in type 2 diabetics having macrovascular disease, still a long way to go. PMID- 15270196 TI - Inhibition of neurofibrillary degeneration: a promising approach to Alzheimer's disease and other tauopathies. AB - Neurofibrillary degeneration (ND) is both a pivotal and a primary lesion of Alzheimer disease (AD) and related tauopathies. To date in all known tauopathics including AD, the neurofibrillary changes, whether of paired helical filaments (PHF), twisted ribbons or straight filaments (SF) are made up of abnormally hyperphosphorylated tau, and the number of these lesions directly correlates to the degree of dementia in the affected individuals. Unlike normal tau which promotes assembly and maintains structure of microtubules, the abnormal tau not only lacks these functions but also sequesters normal tau, MAPI and MAP2, and causes disassembly of microtubules. This toxic behavior of the abnormal tau is solely due to its hyperphosphorylation because dephosphorylation restores it into a normal-like protein. The abnormal hyperphosphorylation also promotes the self assembly of tau into PHF/SF. Missense mutations in tau that cosegregate with the disease in inherited cases of frontotemporal dementia make it a more favorable substrate for hyperphosphorylation. A cause of the abnormal hyperphosphorylation in AD brain is a decrease in the activity of protein phosphatase (PP)-2A, a major regulator of the phosphorylation of tau. The abnormal hyperphosphorylation of tau and neurofibrillary degeneration may be inhibited by increasing the activity of PP-2A, inhibiting the activity of one or more tau kinases or by the sequestration of normal tau by the abnormally hyperphosphorylated tau. A great advantage of developing therapeutic drugs to inhibit neurofibrillary degeneration is that the efficacy of these drugs can be monitored by assaying the CSF levels of phosphotau and total tau, both of which are elevated in AD. Thus, the development of drugs that inhibit neurofibrillary degeneration is a very promising and feasible therapeutic approach to AD and related tauopathies. PMID- 15270197 TI - Posttranslational modifications of tau--role in human tauopathies and modeling in transgenic animals. AB - Alzheimer's disease (AD) is characterized histopathologically by beta-amyloid containing plaques, neurofibrillary tangles (NFT), reduced synaptic density, and neuronal loss in selected brain areas. Plaques consist of aggregates of a small peptide termed Abeta which is derived by proteolysis of the larger amyloid precursor protein APP, whereas NFT are composed of hyperphosphorylated forms of the microtubule-associated protein tau. Tau pathology in the presence of scant or no beta-amyloid plaques characterizes additional neurodegenerative disorders collectively called tauopathies. In the course of plaque and NFT formation, the major proteinaceous components of these lesions undergo post-translational modifications. In the case of tau, these include phosphorylation of mainly serine and threonine, but also tyrosine residues. In addition, tau is subject to ubiquitination, nitration, truncation, prolyl isomerization, association with heparan sulfate proteoglycan, glycosylation, glycation and modification by advanced glycation end-products (AGEs). This review aims to provide insight into the complexity of tau modifications in human tauopathies such as AD and frontotemporal dementia with parkinsonism linked to chromosome 17 (FTDP-17). Selected aspects of the post-translational modification of tau have been reproduced in transgenic animal models. Most of this work has been done in mice, but insight has also been gained from studies in the sea lamprey, the nematode C. elegans and Drosophila. Attempts have been made to link specific post translational modifications with tau aggregation and nerve cell dysfunction. PMID- 15270198 TI - Cholesterol and Alzheimer's disease: clinical and experimental models suggest interactions of different genetic, dietary and environmental risk factors. AB - Alzheimer's disease (AD) is a progressive senile dementia characterized by deposition of a 4 kDa peptide of 39-42 residues known as amyloid beta-peptide (Abeta) in the form of senile plaques and the microtubule associated protein tau as paired helical filaments. Genetic studies have identified mutations in the Abeta precursor protein (APP) as the key triggers for the pathogenesis of AD. Other genes such as presenilins 1 and 2 (PS1/2) and apolipoprotein E (APOE) also play a critical role in increased Abeta deposition. Several biochemical and molecular studies using transfected cultured cells and transgenic animals point to mechanisms by which Abeta is generated and aggregated to trigger the neurodegeneration that may cause AD. Three important enzymes collectively known as 'secretases' participate in APP processing leading to the generation of either Abeta or non-amyloid proteins. However, the mechanisms of neurotoxicity of Abeta and the role of APP function in AD remain important unanswered questions. Although early studies recognized the loss of cholesterol and other lipids in the brain, these findings have been poorly connected with AD pathogenesis, despite the identification of the epsilon4 allele of APOE as a major risk factor in AD. The recent finding that cholesterol can modulate the yield of potentially toxic Abeta has boosted research on its role in AD. Consequently, several cholesterol reducing drugs are currently being evaluated for the treatment of AD. The present review summarizes our current understanding of the relationship of AD pathogenesis with cholesterol, lipids and other genetic and environmental risk factors. PMID- 15270199 TI - Cytokines in neuroinflammation and Alzheimer's disease. AB - Inflammation has been reported in numerous neurodegenerative disorders such as Parkinson's disease, stroke and Alzheimer's disease (AD). In AD, the inflammatory response is mainly located to the vicinity of amyloid plaques. Cytokines, such as Interleukin-1 (IL-1), Interleukin-6 (IL-6), Tumor Necrosis Factor alpha (TNF alpha) and Transforminng Growth Factor beta (TGF-beta) have been clearly involved in this inflammatory process. Although their expression is induced by the presence of amyloid-beta peptide, these cytokines are also able to promote the accumulation of amyloid-beta peptide. Altogether, IL-1, IL-6, TNF-alpha and TGF beta should be considered as key players of a vicious circle leading to the progression of the disease. PMID- 15270200 TI - Metal and inflammatory targets for Alzheimer's disease. AB - Alzheimer's disease (AD) has become linked to inflammation and metal biology. Metals (copper, zinc and iron) and inflammatory cytokines are significant factors that increase the onset of sporadic late onset forms of the dementia. The genetic discovery that alleles in the hemochromatosis gene accelerate the onset of disease by five years has certainly validated interest in the metallobiology of AD as originally described by biochemical criteria. Also the presence of an Iron Responsive Element (IRE) in the 5'UTR of the Amyloid Precursor Protein transcript (APP 5'UTR) provided the first molecular biological support for the current model that APP of AD is a metaloprotein. At the biochemical level, copper, zinc and iron were shown to accelerate the aggregation of the Abeta peptide and enhance metal catalyzed oxidative stress associated with amyloid plaque formation. These amyloid associated events remain the central pathological hallmark of AD in the brain cortex region of AD patients. The involvement of metals in the plaque of AD patients and the demonstration of metal dependent translation of APP mRNA have encouraged the development of chelators as a major new therapeutic strategy for the treatment of AD, running parallel to the development of a vaccine. The other notable pathological feature of AD discussed here is inflammation. The presence of neuro-inflammatory events during AD was supported by clinical trials wherein use of non steroidal anti-inflammatory drugs (NSAIDs) was shown to reduce the risk of developing AD. Drug targets that address inflammation include the use of small molecules that prevent Abeta peptide from activating microglia, the use of cytokine suppressive anti-inflammatory drugs (CSAIDS), and the continued search for a vaccine directed to Abeta sub-fragments (even though the full-length Abeta immunogen generated brain-inflammation and encephalitis in some patients). Our laboratory currently uses a transfection-based assay to screen for small molecule drugs that selectively suppress the capacity of the APP 5'UTR to confer expression to a downstream reporter gene. Based on the presence of both an Interleukin-1 (IL-1) responsive acute box domain and an IRE in the APP 5'UTR, we predict that our APP 5'UTR directed drug screens will identify both novel metal chelators and novel NSAIDS. These lead drugs are readily testable to measure APP holoprotein expression in a cell based secondary assay, and by use of an APP transgenic mouse model to test potential beneficial effects of lead drug treatments on amyloid burden. PMID- 15270201 TI - Beta-sheet breakers for Alzheimer's disease therapy. AB - A growing wealth of evidence indicates that the key pathological event in Alzheimer's disease is the conversion of the normal soluble amyloid-beta peptide into beta-sheet-rich oligomeric structures which have a neurotoxic activity and ability to form insoluble amyloid deposits that accumulate in the brain. Beta sheet breakers constitute a new class of drugs that are designed to specifically bind amyloid-beta peptide and block and/or reverse this abnormal conformational change. In this article we review this approach, describe diverse compounds reported to have this activity and summarize the data supporting the view that beta-sheet breakers could be serious candidates to combat this devastating disease. PMID- 15270202 TI - Mitotic and gender parallels in Alzheimer disease: therapeutic opportunities. AB - In this review, we discuss the role of cell cycle dysfunction in the pathogenesis of Alzheimer disease and propose that such mitotic catastrophe, as one of the earliest events in neuronal degeneration, may, in fact, be sufficient to initiate the neurodegenerative cascade. The question as to what molecule initiates cell cycle dysfunction is now beginning to become understood and, in this regard, the gender-predication, age-related penetrance and regional susceptibility of specific neuronal populations led us to consider luteinizing hormone as a key mediator of the abnormal mitotic process. As such, agents targeted toward luteinizing hormone or downstream sequelae may be of great therapeutic value in the treatment of Alzheimer disease. PMID- 15270203 TI - A new Alzheimer's disease interventive strategy: GLP-1. AB - Glucagon-like peptide-1 (7-36)-amide (GLP-1) is an endogenous 30-amino acid gut peptide, which binds at the GLP-1 receptor coupled to the cyclic AMP second messenger pathway. GLP-1 receptor stimulation enhances pancreatic islet beta-cell proliferation, glucose-dependent insulin secretion and lowers blood glucose and food intake in patients with type 2 diabetes mellitus. Not limited to the pancreas, the chemoarchitecture of GLP-1 receptor distribution in the brain of rodents and humans correlates with a central role for GLP-1 in the regulation of food intake. However emerging evidence suggests that stimulation of neuronal GLP 1 receptors plays an important role in regulating neuronal plasticity and cell survival. GLP-1 has been documented to induce neurite outgrowth and to protect against excitotoxic cell death and oxidative injury in cultured neuronal cells. Moreover, GLP-1 and exendin-4, a naturally occurring more stable analogue of GLP 1 that likewise binds at the GLP-1 receptor, were shown to reduce endogenous levels of amyloid-beta peptide (Abeta) in mouse brain and to reduce levels of beta-amyloid precursor protein (betaAPP) in neurons. Collectively these data suggest that treatment with GLP-1 or a related peptide beneficially affects a number of the therapeutic targets associated with Alzheimer's disease (AD). Although much remains to be elucidated with regards to the downstream signaling pathways involved in the pro-survival properties of GLP-1, modulation of calcium homeostasis may be critical. This review will consider the potential therapeutic relevance of GLP-1 to CNS disorders, such as AD. PMID- 15270204 TI - The role of cytochrome P450 enzymes in the metabolism of risperidone and its clinical relevance for drug interactions. AB - In the recent years it has been increasingly recognized that pharmacogenetical factors play an important role in the drug treatment. These factors may influence the appearance of side-effects and drug interactions due to interindividual differences in the activity of metabolizing enzymes. Risperidone in humans is mainly metabolized to 9-hydroxyrisperidone by the polymorphic cytochrome enzyme P450 2D6 (CYP2D6). Plasma concentrations of risperidone and 9-hydroxyrisperidone show large interindividual variability, which may be partly related to the activity of the CYP2D6 enzyme. Around seven percent of Caucasians have a genetically inherited impaired activity of the CYP2D6 enzyme. Debrisoquine metabolic ratio (a marker of CYP2D6 activity) and the number of CYP2D6 active genes have been related to risperidone plasma concentrations among patients during steady-state conditions. A large number drugs have been described to be metabolized by CYP2D6, and it is therefore important to evaluate the clinical significance of the impaired metabolism and possible drug interactions on the enzyme. Since risperidone/9-hydroxyrisperidone ratio strongly correlates with CYP2D6 enzyme activity and the number of CYP2D6 active genes, thus it might be a useful tool in clinical practice to estimate the possible risk of drug interactions due to impaired CYP2D6 enzyme activity. CYP3A4 is the most abundant drug metabolizing enzyme in humans, and in vitro and in vivo results suggest also a role for the enzyme in risperidone metabolism. The consideration of the implication of cytochrome P450 enzymes in risperidone metabolism may help to individualize dose schemes in order to avoid interactions and potentially dangerous side-effects, such us QTc interval lengthening among patients with cardiac risk factors. PMID- 15270205 TI - Molecular targets for modulating lung inflammation and injury. AB - The inflammatory response of the lung and airways is one of the main targets for tile development of new therapies for variety of disorders including the acute respiratory distress syndrome, cystic fibrosis, idiopathic pulmonary fibrosis, and chronic obstructive pulmonary disease. Over the last decade our understanding of the molecular biology of the inflammatory response has advanced considerably and has opened up new avenues for therapeutic intervention. Furthermore, the mechanism of action of many of the existing anti-inflammatory agents has been revealed by this burgeoning information. Here, we discuss the functions and therapeutic potential of molecules that might prove promising as targets for treatment of inflammatory lung diseases. These possible molecular targets include cell surface proteins/receptors [toll like receptors (TLRs), triggering receptors expressed on myeloid cells (TREMs), and syndecans)], transcription factors [NF kappaB, AP-1, PU.1, and high mobility group box 1 (HMGB1)], and regulatory proteins [macrophage migration inhibitory factor (MIF), granulocyte macrophage colony stimulating factor (GM-CSF), cyclooxygenase 2 (COX-2), heme oxygenase 1 (HO-1)]. PMID- 15270206 TI - To know or not to know the identity of gametes donors? The UK and European legal context. PMID- 15270207 TI - Human cervical mucus can act in vitro as a selective barrier against spermatozoa carrying fragmented DNA and chromatin structural abnormalities. AB - PURPOSE: We have carried out experiments to determine if human cervical mucus can act as an in vitro selective barrier against spermatozoa morphologically normal that carry genetic structural abnormalities. METHODS: Sperm chromatin abnormalities have been evaluated by Chromomycin A3 and "endogenous" nick translation. RESULTS: The data obtained have shown that spermatozoa possessing higher levels of DNA protamination are more proficient in crossing the cervical mucus barrier. Moreover, the levels of positivity to endogenous nick translation treatment was practically zero in such spermatozoa. CONCLUSIONS: We suggest that sperm penetration of cervical mucus could be used to select sperm preparations free of fragmented DNA or chromatin structural abnormalities for assisted reproduction. PMID- 15270208 TI - Monozygotic twins and triplets in association with blastocyst transfer. AB - PURPOSE: To compare the incidence of monozygotic twins following blastocyst versus day-3 embryo transfer (ET). METHODS: A retrospective analysis of the outcome of assisted reproductive technology (ART) cycles utilizing blastocyst ET during 1999-2000 was compared to a similar group of patients undergoing day-3 ET during 1997-1998. RESULTS: Blastocyst ET was used in 75 cycles with 2.0 +/- 2 embryos transferred. The comparison group consisted of 90 cycles with day-3 ET and 3.0 +/- 2 embryos transferred. CONCLUSIONS: High pregnancy rates are maintained with blastocyst ET even though fewer embryos are transferred. The rate of monozygotic twins is higher with blastocyst ET than with day-3 ET. This increase may partially negate the benefit of reduced high-order multiple gestations attributed to blastocyst ET. PMID- 15270209 TI - Parameters affecting the results in a program of artificial insemination with donor sperm. A 12-year retrospective review of more than 1800 cycles. AB - PURPOSE: We aimed to establish the influence of the parameters affecting artificial insemination (AI) results in order to describe the ideal situations to achieve the best results as well as to adequately counsel the patients undergoing these treatments about their pregnancy chances. METHODS: We performed a controlled retrospective clinical study over more than one decade in a total of 1858 cycles in 710 patients. Clinical histories and computer registers were systematically reviewed between January 1990 and June 2002. We analyzed the influence of diverse factors affecting AI results such as patient's age, ovarian stimulation, and seminal characteristics to offer a detailed description of the technique. RESULTS: Less than 35-years-old, smooth ovarian stimulation and 5 million of progressive motile sperm inseminated two consecutive days are the optimum conditions for achieving good results. Also, period of time that sperm remained frozen do not affect the result. Furthermore, we present the likely or expected outcomes of these treatments depending on the male and female etiologies. CONCLUSIONS: We discourage AI in aged patients, and strongly recommend undergoing ovarian stimulation. Nonetheless, we must reach an adequate amount of sperm with good motility in order to inseminate with maximum guaranties of success. PMID- 15270210 TI - DNA fragmentation, mitochondrial dysfunction and chromosomal aneuploidy in the spermatozoa of oligoasthenoteratozoospermic males. AB - PURPOSE: This study determined the incidence of sperm nuclear DNA fragmentation, mitochondrial dysfunction, and chromosomal aneuploidy. The results were correlated with the semen analysis parameters and fertilization rates. METHODS: Semen samples from 10 men showing oligoasthenoteratozoospermia (OAT) and undergoing ICSI treatment were analyzed. Another semen samples from 10 men showing normozoospermia and undergoing IVF treatment were analyzed for comparison. The samples were prepared using a two-step discontinuous Percoll gradient (80%-50%) and analyzed using a Hamilton-Thorne Integrated Visual Optical System (IVOS) Sperm Analyzer. DNA fragmentation was detected with a terminal deoxynucleotidyl transferase-mediated dUTP nick end label (TUNEL) assay. Functional integrity of mitochondria was detected using an Apoalert Mitochondrial Membrane Sensor Kit. Chromosomal aneuploidy was assayed by fluorescence in situ hybridization. RESULTS: Higher sperm DNA fragmentation rate (18.8% vs. 2.8%), mitochondrial dysfunction rate (24.9% vs. 5.7%), and chromosomal aneuploidy rate (0.12% vs. 0.06%) were found in the oligoasthenoteratozoospermic patients in comparison with the normozoospermic patients. CONCLUSIONS: The result indicates that spermatozoa from oligoasthenoteratozoospermic patients contain greater DNA fragmentation, mitochondrial dysfunction, and chromosomal aneuploidy. Because extremely poor semen samples are the indication for ICSI treatment, the result indicates the importance of selecting good quality sperm for oocyte injection. PMID- 15270211 TI - A proposal to change the method for determining the outcome according to the SART statistic--pregnancy rate per retrieval. PMID- 15270212 TI - Isolation and expression analysis of the testis-specific gene, human OPPO1. AB - PURPOSE: To investigate human spermatogenesis, we isolated human testis-specific genes. METHODS: Using mouse amino acid sequences, we found the region including homology in amino acid level in the human genome sequences. The primers encompassing introns were made and RT-PCR and RACE were carried out. The resultant PCR products were sequenced. RESULTS: The full-length cDNA of human OPPO1 was isolated. It encodes 257 amino acid residues. The expression of the human OPPO1 was predominantly in the testis. On the other hand, partial cDNAs of ZNF8, GR194, GR219, GR093, GR046, GR163, and GR200 were expressed in the various tissues. CONCLUSIONS: Our data suggests that the human OPPO1 may play important roles in human spermatogenesis. PMID- 15270213 TI - Neuromodulatory octopaminergic neurons and their functions during insect motor behaviour. The Ernst Florey memory lecture. AB - In this article we describe recent advances in functional studies on the role of octopamine released in the periphery by efferent dorsal or ventral unpaired median neurons. In addition to the previously described modulatory effects on the neuromuscular junction, we describe a metabolic regulatory role for these neurons. Due to their activity glycolytic rates in target tissues, such as muscles, are increased. In flight muscles that use carbohydrate catabolism only at take-off but have to switch to lipid oxidation during prolonged flight, these neurons are only active at rest but are inhibited as soon as flight motor patterns are selected. PMID- 15270214 TI - Properties of descending dorsal unpaired median (DUM) neurons of the locust suboesophageal ganglion. AB - A group of six dorsal unpaired median (DUM) neurons of the suboesophageal ganglion (SOG) of locusts was studied with neuroanatomical and electrophysiological techniques. The neurons are located posteriorly in the SOG and have axons that descend into the ganglia of the ventral nerve cord, some as far as the terminal abdominal ganglion. Within thoracic ganglia the neurons have profuse dendritic ramifications in many neuropiles, including ventral sensory neuropiles. Based on their projection patterns three different morphological types of neurons can be distinguished. These neurons receive excitatory inputs through sensory pathways that ascend from the thoracic ganglia and are activated by limb movements. They may be involved in the modulation of synaptic transmission in thoracic ganglia. PMID- 15270215 TI - Separate distribution of deutocerebral projection neurons in the mushroom bodies of the cricket brain. AB - Deutocerebral projection neurones in the brain of the cricket (Gryllus bimaculatus) have been investigated by experimental dextran staining, viewed by light and electron microscopy. These neurones of two separate somata clusters innervate two separate primary glomerular neuropils of the deutocerebral segment, either the antennal lobe receiving only antennal nerve sensory input, or the glomerular lobe, receiving input from sensory neurones of lower segmental origin, including chemosensory fibres from mouth parts. Projection neurones of the antennal lobe only invade the anterior calyx of the mushroom body neuropil via the inner antenno glomerular tract, while glomerular relay neurones of the glomerular lobe innervate only the posterior calyx via the tritocerebral tract. All types of projection neurones give rise to presynaptic boutons. forming the central core of microglomeruli with patterned distribution. These projection neurons are cholinergic. The results are discussed in view of maintained segregated modal information, first processed in the separated primary deutocerebral neuropiles and further on in the second order input neuropils of the mushroom bodies. The large posterior calyces are proposed as a compartment for gustatory information. PMID- 15270216 TI - Multisensory convergence in the mushroom bodies of ants and bees. AB - The mushroom bodies, central neuropils in the arthropod brain, are involved in learning and memory and in the control of complex behavior. In most insects, the mushroom bodies receive direct olfactory input in their calyx region. In Hymenoptera, olfactory input is layered in the calyx. In ants, several layers can be discriminated that correspond to different clusters of glomeruli in the antennal lobes, perhaps corresponding to different classes of odors. Only in Hymenoptera, the mushroom body calyx also receives direct visual input from the optic lobes. In bees, six calycal layers receive input from different classes of visual interneurons, probably representing different parts of the visual field and different visual properties. Taken together, the mushroom bodies receive distinct multisensory information in many segregated input layers. PMID- 15270217 TI - Comparative morphology of central neuropils in the brain of arthropods and its evolutionary and functional implications. AB - Most insects and decapod crustaceans possess an assemblage of midline neuropils, the central complex. Recent phylogenetic studies show a sister-group relationship between hexapods and decapods, suggesting that central complexes in both groups are homologous structures derived from a basal ancestral neuropil. This ancestral archetype of the central complex (lacking the protocerebral bridge) might be represented in the chilopods. Until recently, diplopods were regarded as closely related to chilopods and united within the taxon "Myriapoda". The entire lack of a midline neuropil in diplopods, however, renders the monophyletic origin of the class Myriapoda unlikely. In this study we used a palette of immunocytochemical and neuroanatomical methods to investigate mid-line neuropils in hitherto poorly examined arthropod groups. Of special interest for resolving arthropod phylogeny are onychophorans, who are believed to be an evolutionary ancient group that resembles the ancestors of modern arthropods. Striking similarities in central brain neuroarchitecture of the onychophoran Euperipatoides rowellii and of a chelicerate species, however, suggest a close phylogenetic relationship between these two groups. Our findings imply that onychophorans either represent the oldest form of the chelicerates or that extant onychophorans have developed from chelicerate-like ancestors by neoteny. PMID- 15270218 TI - Learning channels. Cellular physiology of odor processing neurons within the honeybee brain. AB - To understand the cellular mechanisms of olfactory learning in the honeybee brain we study the physiology of identified neurons within the olfactory pathway. Here, we review data on the voltage-sensitive and ligand-gated ionic currents of mushroom body Kenyon cells and antennal lobe neurons in vitro and in situ. Both cell types generate action potentials in vitro, but have different voltage sensitive K+ currents. They express nicotinic acetylcholine receptors and ionotropic GABA receptors, representing the major transmitter systems in the insect olfactory system. Our data are interpreted with respect to learning dependent plasticity in the honeybee brain. PMID- 15270219 TI - Nitric oxide regulates the levels of cGMP accumulation in the cricket brain. AB - Cricket brains were incubated in a saline containing nitric oxide (NO)-donor and phosphodiesterase inhibitor IBMX, which could activate soluble guanylate cyclase (sGC) to increase cGMP levels in the targets of NO. The increase of cGMP was detected by immunohistochemistry and enzyme linked immunosorbent assay. NO induced cGMP immunohistochemistry revealed that many cell bodies of cricket brain showed cGMP immunoreactivity when preparations were treated with a saline containing 10 mM NO-donor SNP and phosphodiesterase inhibitor IBMX, but only a few cell bodies showed immunoreactivity when preparations were incubated without NO-donor. The concentration of cGMP in cricket brains were then measured by using cGMP-specific enzyme linked immunosorbent assay. Cricket brains were treated with a saline containing 1 microM of NO-donor NOR3 and 1 mM IBMX. The cGMP levels in the brain were increased about 75% compared to control preparations that was treated with a cricket saline containing IBMX. The level of cGMP decreased about 40% when preparations were incubated NOR3 saline containing sGC inhibitor ODQ. These results indicate that NO activates sGC and increases the levels of cGMP in particular neurons of the cricket brain and that the level of cGMP would be kept a particular level, which might regulate synaptic efficacy in the neurotransmission. PMID- 15270220 TI - Action spectrum of foraging behavior of the Japanese yellow swallowtail butterfly, Papilio xuthus. AB - This paper describes the action spectrum of foraging behavior of a butterfly, Papilio xuthus. We first established an experimental protocol to evaluate learning and discrimination of monochromatic light by the butterflies. We trained butterflies to feed on sucrose solution at the window illuminated with certain monochromatic light produced through a monochromator. After confirming that they learned the monochromatic light, after 10 days of training, we tested the butterflies one by one. We presented training wavelengths for each individual at different intensities, and recorded whether they perform foraging behavior under freely-flying as well as tethered conditions. Freely-flying butterflies responded to light by visiting the window and searching for nectar around it, whereas tethered butterflies responded by extending their proboscides towards the window. The light intensity required to elicit 50% response for each tested monochromatic light was plotted. The resulting action spectrum for the visit was rather flat with the maximum sensitivity a 420 nm, whereas the spectrum for the proboscis extension had prominent peaks at 380, 500 and 600 nm. The difference in action spectra indicates that the visit and the proboscis extension are controlled by two independent mechanisms at least in part. PMID- 15270221 TI - Neurobiology of polarization vision in the locust Schistocerca gregaria. AB - The polarization pattern of the blue sky serves as an important reference for spatial orientation in insects. To understand the neural mechanisms involved in sky compass orientation we have analyzed the polarization vision system in the locust Schistocerca gregaria. As in other insects, photoreceptors adapted for the detection of sky polarization are concentrated in a dorsal rim area (DRA) of the compound eye. Stationary flying locusts show polarotactic yaw-torque responses when illuminated through a rotating polarizer from above. This response is abolished after painting the DRAs. Central stages of the polarization vision system, revealed through tracing studies, include dorsal areas in the lamina and medulla, the anterior lobe of the lobula, the anterior optic tubercle, the lateral accessory lobe and the central complex. Physiological analysis of polarization-sensitive (POL) neurons has focussed on the optic tubercle and on the central complex. Each POL neuron was maximally excited at a certain e-vector (phimax) and was maximally inhibited at an e-vector perpendicular to phimax. The neurons had large visual fields, and many neurons received input from both eyes. The neuronal organization of the central complex suggests a role as a spatial compass within the locust brain. PMID- 15270222 TI - From bioassays to Drosophila genetics: strategies for characterizing an essential insect neurohormone, bursicon. AB - We describe the molecular analysis and cellular expression of the insect peptide neurohormone, bursicon. Bursicon triggers the sclerotization of the soft insect cuticle after ecdysis. Using protein elution analyses from SDS gels, we determined the molecular weight of bursicon from different insects to be approximately 30 kDa. Four partial peptide sequences of Periplaneta americana bursicon were obtained from purified nerve cord homogenates separated on two dimensional gels. Antibodies produced against one of the sequences identified the cellular location of bursicon in different insects and showed that bursicon is co produced with crustacean cardioactive peptide (CCAP) in the same neurons in all insects tested so far. Additionally, using the partial peptide sequences, we successfully searched the Drosophila genome project for the gene encoding bursicon. With Drosophila as a tool, we can now verify the function of the sequence using transgenic flies. Sequence comparisons also allowed us to verify that bursicon is conserved, corroborating the older data from bioassays and immunohistochemical analyses. The sequence of bursicon will enable further analysis of its function, release, and evolution. PMID- 15270223 TI - Wasp manipulates cockroach behavior by injecting venom cocktail into prey central nervous system. AB - The parasitoid wasp Ampulex compressa induces behavioral changes in the cockroach prey by injecting venom into its central nervous system. In contrast to most other venomous predators, the wasp's sting does not induce paralysis. Rather, the two consecutive stings in the thoracic and head ganglia induce three stereotypic behavioral effects. The prey behavior is manipulated in a way beneficial to the wasp and its offspring by providing a living meal for its newborn larva. The first sting in the thorax causes a transient front leg paralysis lasting a few minutes. This paralysis prevents the cockroach from fighting with its front legs, thereby facilitating the second sting in the head. A postsynaptic block of central synaptic transmission mediates this leg paralysis. Following the head sting, dopamine identified in the venom induces 30 minutes of intense grooming that appears to prevent the cockroach from straying until the last and third behavioral effect of hypokinesia commences. In this lethargic state that lasts about three weeks, the cockroach does not respond to various stimuli nor does it initiates movement. However, other specific behaviors of the prey are unaffected. We propose that the venom represses the activity of head ganglia neurons thereby removing the descending excitatory drive to specific thoracic neurons. PMID- 15270224 TI - Auto-spermatophore extrusion reveals that the reproductive timer functions in the separated terminal abdominal ganglion in the male cricket. AB - Auto-spermatophore extrusion is a kind of spermatophore extrusion without genital coupling in the male cricket. It rarely occurred in intact males paired with a female, while it frequently occurred in all the males with the connectives cut under restraint and dissection. The time interval (SPaSE) between spermatophore preparation and auto-spermatophore extrusion was found to be comparable to that (RS2) of the time-fixed sexually refractory stage measured by the calling song. According to extracellular spike recording, the dorsal pouch motoneuron (mDP), which singly innervates the dorsal pouch muscles and is responsible for spermatophore extrusion, showed a burst discharge in association with auto spermatophore extrusion with an interval similar to RS2 in males with the connectives transected between the 6th abdominal ganglion and the terminal abdominal ganglion (TAG) after spermatophore preparation. These results strengthened our previous conclusion that the reproductive timer for RS2 is located in the TAG, and demonstrated that it functions normally even in the TAG separated from the rest of the central nervous system. PMID- 15270225 TI - Thee effect of 5-HTP on the reproductive timer in the male cricket. AB - The post-copulatory sexually refractory stage in the male cricket Gryllus bimaculatus consists of the two substages: the first refractory stage (RS1, time variable) between copulation and spermatophore preparation, and the second refractory stage (RS2, time-constant) between spermatophore preparation and the recommencement of courtship. To understand the mechanism of the timer for RS2, subcuticular or intraganglionic injection of biogenic amines (10(-2) mol l(-1)) was performed immediately after spermatophore preparation. RS2 was shortened by octopamine, 5-HT, 5-HTP and NA-5-HT. Among these, 5-HTP was most potent. It shortened RS2 to maximally about 38% of the control. The shortening effect continued for 4.5 h after subcuticular injection even when the hemolymph was washed out with saline at 1 hour after injection. Simultaneous injection of 5-HTP with the inhibitor (NSD-1015) of 5-HT synthesis enzyme nullified the effect of 5 HTP, indicating that the shortening effect was caused by 5-HT synthesized from extrinsic 5-HTP. Injection of the inhibitor (CHX) of protein synthesis had no effect of on RS2. These results suggest that the reproductive timer in the TAG may be controlled by 5-HT or a second messenger mediated by 5-HT. PMID- 15270226 TI - The locust frontal ganglion: a multi-tasked central pattern generator. AB - The locust frontal ganglion (FG) constitutes a major source of innervation to the foregut dilator muscles and thus plays a key role in control of foregut movements. This paper reviews our recent studies on the generation and characteristics of FG motor outputs in two distinct and fundamental locust behaviors: feeding and molting. In an in vitro preparation, isolated from all descending and sensory inputs, the FG was spontaneously active and generated rhythmic multi-unit bursts of action potentials, which could be recorded from all efferent nerves. Thus the FG motor pattern is generated by a central pattern generator within the ganglion. Intracellular recordings suggest that only a small fraction (10-20%) of the FG 100 neurons demonstrate rhythmic activity. The FG motor output in vivo was relatively complex, and strongly dependent on the locust's physiological and behavioral state. Rhythmic activity of the foregut was found to depend on the amount of food present in the crop; animals with full crop demonstrated higher FG burst frequency than those with empty crop. At the molt, the FG generates a distinct motor pattern that could be related to air-swallowing behavior. PMID- 15270227 TI - Neurophysiological studies of flight-related density-dependent phase characteristics in locusts. AB - Locusts show an extreme example of density-dependent phase polymorphism, demonstrating within the species differences in morphology as well as biology, dependent on the population density. Behavior is the primary density-dependent change which facilitates the appearance of various morphological and physiological phase characteristics. We have studied density dependent differences in flight related sensory and central neural elements in the desert locust Schistocerca gregaria. Wind generated high frequency spiking activity in the tritocerebral commissure giant (TCG, an identified interneuron that relay inputs from head hair receptors to thoracic motor centers) that was much less intense in solitary locusts, compared to gregarious ones. In addition the solitary locusts' TCG demonstrated much stronger adaptation of its response. In cases when flight was initiated high frequency TCG activity was independent of the locust phase. The tritocerebral commissure dwarf (TCD) is a GABAergic flight related interneuron that is sensitive to ambient illumination intensity. An increase in the TCD spontaneous activity under dark vs. light conditions was significantly higher in gregarious locusts then in solitary ones, implying a flight-related inhibitory mechanism that is far more active in gregarious locusts under dark conditions. Thus, density-dependent phase differences in interneuron activity pattern and properties well reflect and may be at least partially responsible to behavioral flight-related characteristics. PMID- 15270228 TI - Estradiol-stimulated nitric oxide release in nervous tissue, vasculature, and gonads of the giant cockroach Blaberus craniifer. AB - The vertebrate system of steroid hormones appears to have been conserved widely throughout the animal kingdom. The sex hormone estrogen, 17-beta-estradiol (E2), long considered to be exclusively a vertebrate hormone, is found also in invertebrates related to reproductive and developmental processes such as spawning, vitellogenesis and molting. These processes are affected by estrogen induced changes at the genomic level and take place at a large time scale. The discovery of surface membrane receptors for E2 has opened new possibilities for the involvement of estrogen in biological functions other than reproductive. These processes take place within a few seconds to minutes and involve sudden cytosolic calcium transients, activation of adenylate cyclase or activation of phospholipase C (PLC). E2 can modulate the production of nitric oxide (NO) in endotheliar and other cells. A similar mechanism linking estrogen to cNOS catalized nitric oxide (NO) release is reported herein for the first time in several tissues of the giant cockroach Blaberus craniifer. This process has been identified in the brain, nerve cord, vasculature and ovaries. This effect is concentration dependent and is inhibited by tamoxifen an estrogen receptor blocker. PMID- 15270229 TI - The early snail acquires the learning. Comparison of scores for conditioned taste aversion between morning and afternoon. AB - The pond snail Lymnaea stagnalis acquires conditioned taste aversion (CTA) and maintains its memory for more than a month. Snails in our laboratory were cultured at 20 degrees C on a 12:12 light-dark cycle (light from 7 am to 7 pm). To examine the hours during which snails acquire CTA effectively, we trained some snails in the morning and others in the afternoon, and then compared their scores. CTA developed in both cases, but scores were significantly better in the morning than in the afternoon. To elucidate the cause of this difference in scores, we observed the voluntary activity of snails and found the circadian rhythm reflected in the snails' free-movement distances; distances at the circadian time 0-12 (daytime) were significantly longer than those at the circadian time 12-24 (nighttime). This rhythm was kept up for at least 3 days, even in constant darkness. In conclusion, L. stagnalis should be trained in the morning to acquire associative learning, possibly because of its greater propensity to roam about at that time as opposed to the afternoon. PMID- 15270230 TI - Real-time quantitative RT-PCR method for estimation of mRNA level of CCAAT/enhancer binding protein in the central nervous system of Lymnaea stagnalis. AB - The fluorescence-based real-time reverse transcription polymerase chain reaction (RT-PCR) is becoming widely used to quantify mRNA level in cells and tissues and is now a crucial tool for basic biological researches and biotechnology. In the present study, on the basis of the real-time quantitative RT-RCR, we detected and quantified mRNA copies of the transcription factor, CCAAT/enhancer binding protein (C/EBP: an immediate-early gene that is involved in synaptic plasticity and learning and memory) in the central nervous system of the pond snail Lymnaea stagnalis. We designed the primer set and the probe in the specific insert for the detection of Lymnaea C/EBP (LymC/EBP) clone 1. This insert is not contained in LymC/EBP clone 2 by alternative splicing. The copy number of LymC/EBP clone 1 was linearly decreased relative to the dilution of cDNA, and it was estimated 30 copies/microl in test sample. The availability of the present study showed that the real-time quantitative RT-PCR technique is more accurate and more specific for the detection and quantification of the mRNA level of genes in L. stagnalis than the other PCR methods. PMID- 15270231 TI - The expression pattern of CREB genes in the central nervous system of the pond snail Lymnaea stagnalis. AB - To analyze the expression pattern of genes of cAMP responsive element binding protein (CREB), we performed in situ hybridization for the whole central nervous system (CNS) of the pond snail Lymnaea stagnalis. The CREB1 (activator) and CREB2 (repressor) homologues have already been cloned in L. stagnalis, and they are referred to as LymCREB1 and LymCREB2. Using the frozen sections and the whole mount preparations of the CNS, we mapped the distribution of LymCREB1 and LymCREB2 mRNA containing neurons. The present findings showed that the LymCREB1 mRNA containing neurons are a relatively few, whereas LymCREB2 mRNA is contained ubiquitously in the whole CNS of L. stagnalis. PMID- 15270232 TI - Octopaminergic modulation of the membrane currents in the central feeding system of the pond snail Lymnaea stagnalis. AB - Octopamine is released by the intrinsic OC interneurons in the paired buccal ganglia and serves both as a neurotransmitter and a neuromodulator in the central feeding network of the pond snail Lymnaea stagnalis. The identified B1 buccal motoneuron receives excitatory inputs from the OC interneurons and is more excitable in the presence of 10 microM octopamine in the bath. This modulatory effect of octopamine on the B1 motoneuron was studied using the two electrode voltage clamp method. In normal physiological saline depolarising voltage steps from the holding potential of -80 mV evoke a transient inward current, presumably carried by Na(+) ions. The peak values of this inward current are increased in the presence of 10 microM octopamine in the bath. In contrast, both the transient (IA) and delayed (IK) outward currents are unaffected by octopamine application. Replacing the normal saline with a Na(+)-free bathing solution containing K(+) channel blockers (50 mM TEACl, 4 mM 4AP) revealed the presence of an additional inward current of the B1 neurons, carried by Ca(2+). Octopamine (10 microM) in the bath decreased the amplitudes of this current. These results suggest that the membrane mechanisms which underlie the modulatory effect of octopamine on the B1 motoneuron include selective changes of the Na(+)- and Ca(2+)-channels. PMID- 15270233 TI - Second messengers of octopamine receptors in the snail Lymnaea. AB - We describe octopamine responses of 3 large buccal neurons of Lymnaea and test the hypothesis that these are cAMP-dependent. The B1 neuron is excited by octopamine and the depolarisation is significantly enlarged (P < 0.05) by application of the blocker of cAMP breakdown, 3-isobutyl-1-methylxanthine (IBMX). The B1 neuron is also depolarised by forskolin, an activator of adenylyl cyclase. The B2 and B3 neurons are inhibited by octopamine, and the response is not affected by IBMX. Both cells are excited by forskolin. We conclude that the B1 neuron response to octopamine is likely to be mediated by cAMP, while the B2 and B3 responses are cAMP-independent. PMID- 15270234 TI - Thee effect of food intake on the central monoaminergic system in the snail, Lymnaea stagnalis. AB - We investigated the effect of food intake on the serotonin and dopamine levels of the CNS as well as on the spontaneous firing activity of the CGC in isolated preparations from starved, feeding and satiated animals. Furthermore we investigated the effects of 1 microM serotonin and/or dopamine and their mixture on the firing activity of the CGC. The HPLC assay of serotonin and dopamine showed that during food intake both the serotonin and dopamine levels of the CNS increased whereas in satiated animals their levels were not significantly more than the control levels. Recording from the CGC in isolated CNS preparation from starved, feeding or satiated animals showed that feeding increased the firing frequency of the CGC compared to the starved control. The application of 1 microM dopamine decreased the firing frequency whereas the application of 1 microM serotonin increased the firing frequency of the CGC. We conclude that during food intake the external and internal food stimuli increase the activity of the central monoaminergic system and also increase the levels of monoamines in the CNS. Furthermore, we also suggest that the increased dopamine and serotonin levels both affect the activity of the serotonergic neurons during the different phases of feeding. PMID- 15270235 TI - Hyperpolarization by glucose of feeding-related neurons in snail. AB - In the pond snail Lymnaea stagnalis, D-glucose action was investigated on electrical activity of identified central neurons. In the CNS preparations isolated from specimens that starved for 24-96 h, D-glucose added to a standard or HiDi saline at 500-700 microg/ml effectively hyperpolarized ca. 90% of feeding related neurons B1, SO and CGC. However, not all feeding-related neurons examined were responsive to glucose. Experiments on cells of the serotonergic Pedal A cluster have shown that hyperpolarizing action of D-glucose is retained following complete isolation of "hunger" neurons. Threshold concentration producing 1-3 mV hyperpolarization was ca. 50 microg/ml. The results suggest a direct glucose involvement in the mechanisms that control feeding behavior in Lymnaea. PMID- 15270236 TI - Serotonin, nitric oxide and histamine enhance the excitability of neuron MCC by diverse mechanisms. AB - Serotonin, nitric oxide (NO) and histamine are neuromodulators used in molluscan nervous systems. We have found that each of them depolarizes and increases the excitability of the serotonergic feeding neural circuit modulator neuron, MCC, of Aplysia, but each induces different changes in background ionic currents and uses a different second messenger. Stimulation of neuron C2 in the cerebral ganglion induces a vsEPSP in MCC using NO and histamine. When these neurons are isolated in culture they form synapses that mediate the vsEPSP. The ionic currents induced by these neuromodulators were investigated in isolated cultured MCCs. Histamine reduced a background outward current between -70 and -30 mV that was blocked by cobalt treatment, indicating that it is a calcium activated potassium current. Serotonin reduced a background outward current from -65 mV to -30 mV and enhanced a potassium inward current more negative than -70 mV that was blocked by cesium and barium. This response was mimicked by 8-Br-cAMP. NO donors reduced a cobalt insensitive background outward current between -70 and -30 mV. This response was mimicked by 8-Br-cGMP. These responses show that MCC can produce complex time and state-dependent activity during its modulation of the feeding neural circuit. PMID- 15270237 TI - Regulation of afferent transmission in the feeding circuitry of Aplysia. AB - Although feeding in Aplysia is mediated by a central pattern generator (CPG), the activity of this CPG is modified by afferent input. To determine how afferent activity produces the widespread changes in motor programs that are necessary if behavior is to be modified, we have studied two classes of feeding sensory neurons. We have shown that afferent-induced changes in activity are widespread because sensory neurons make a number of synaptic connections. For example, sensory neurons make monosynaptic excitatory connections with feeding motor neurons. Sensori-motor transmission is, however, regulated so that changes in the periphery do not disrupt ongoing activity. This results from the fact that sensory neurons are also electrically coupled to feeding interneurons. During motor programs sensory neurons are, therefore, rhythmically depolarized via central input. These changes in membrane potential profoundly affect sensori motor transmission. For example, changes in membrane potential alter spike propagation in sensory neurons so that spikes are only actively transmitted to particular output regions when it is behaviorally appropriate. To summarize, afferent activity alters motor output because sensory neurons make direct contact with motor neurons. Sensori-motor transmission is, however, centrally regulated so that changes in the periphery alter motor programs in a phase-dependent manner. PMID- 15270238 TI - Functional morphology of the salivary gland of the snail, Helix pomatia: a histochemical and immunocytochemical study. AB - Functional morphology of Helix pomatia salivary gland cells was studied at light microscopic level by using different histochemical methods. Three cell types could be demonstrated in the salivary gland: mucocytes, granular and vacuolated cells. The distribution and the number of the different cell types were different in active and inactive snails. In active feeding animals, dilatated interlobular salivary ducts were observed, which were never present in inactive ones. In active animals an additional cell type, the cystic cell could also be observed. Periodic acid Schiff staining revealed both mucuos and serous elements in the salivary gland. Furthermore, hematoxyline-eosin staining indicated the occurrence of a cell layer with high mitotic activity in the acini. Applying immunohistochemical methods with monoclonal mouse anti-human Ki-67 clone, B56 and polyclonal rabbit anti-human Ki-67 antibodies, we also were able to demonstrate the occurrence of dividing cells in the salivary gland. Analysis of 1-2 microm semi-thin Araldite sections stained with toluidine-blue showed that the saliva can be released, in addition to possible exocytosis, by the lysis of cystic cells. Using an apoptosis kit, we could also establish that this process was due to rather an apoptotic than a necrotic mechanism. In the salivary gland of active snails, where an intensive salivation takes place, significantly more apoptotic cells occurred, if compared to that of inactive animals. It is suggested that programmed cell death may also be involved in the saliva release. PMID- 15270239 TI - Phasic coordination between locomotor and respiratory rhythms in Lymnaea. Real behavior and computer simulation. AB - In the pond snail Lymnaea stagnalis, a firm phase-locked coupling of pneumostome movements to the locomotor cycle was observed during terrestrial locomotion, thus demonstrating that the coordination between locomotor and respiratory rhythms is a natural behavioral event in this animal. The results of computational modelling suggest a possible scheme of coordination between these motor rhythms which is based on inhibitory projection from the central pattern generator for locomotion to that for respiration. These findings allow the neuronal mechanisms underlying coordination of two rhythmic behaviors to be investigated. PMID- 15270240 TI - Why the ovotestis of Helix aspersa is innervated. AB - Although Schmalz described the innervation of the ovotestis in pulmonate snails as early as 1914, no functions have been attributed to it. In H. aspersa, the intestinal nerve branches profusely within the ovotestis and terminates in the walls of the acini and in the sheath surrounding the early portion of the hermaphroditic duct. We found both sensory and motor functions for this innervation. Significantly, there is a tonic sensory discharge generated by the mechanical pressure of growing oocytes, and the level of tonic afferent activity is strongly correlated with the number of ripe oocytes; this is probably a permissive signal that gates ovulation. Tactile stimulation of the ovotestis causes a phasic sensory discharge and a pronounced cardio activation. Also, an efferent discharge is elicited in the ovotestis branch of the intestinal nerve. To study the motor consequences of efferent activity, the ovotestis branch was electrically stimulated. We found that such stimulation evokes peristaltic contractions of the initial portion of the hermaphroditic duct and increases beat frequencies of the cilia that line the interior of the duct. These effects could facilitate the transport of oocytes down the duct. Still other functions of afferent activity are implied by changes in the spontaneous activity of mesocerebral cells following nerve stimulation. Putative sensory neurons and putative motoneurons have been identified in the visceral and right parietal ganglia. PMID- 15270241 TI - Survival of egg-laying controlling neuroendocrine cells during reproductive senescence of a mollusc. AB - During brain aging neuronal degradation occurs. In some neurons this may result in degeneration and cell death, still other neurons may survive and maintain their basic properties. The present study deals with survival of the egg-laying controlling neuroendocrine caudodorsal cells (CDCs) during reproductive senescence of the pond snail Lymnaea stagnalis. In senescent animals CDCs exhibited reduced branching patterns but still maintained their electrophysiological characteristics. In the isolated CNS the cells could still respond with an afterdischarge upon electrical stimulation. After an extended period of no egg laying of Lymnaea CDCs failed to exhibit an afterdischarge. In senescent CDCs that failed an afterdischarge, discharge activity could be restored by exposure to peptides released by CDCs from reproductive animals. Moreover, raising the intracellular cAMP level could induce discharge activity in CDCs with afterdischarge failure. Discharge activity also occurred during depolarization of senescent CDCs by exposure of the cells to saline with a high potassium concentration. These results indicate that in senescent CDCs the pacemaking mechanism of the afterdischarge is still intact but that the initial activation fails. Chemical (auto)transmission of CDCs in such animals was indeed reduced as indicated by the small amplitude of the depolarizing afterpotential (DAP) induced by electrical stimulation. Interestingly, CDCs of senescent animals contained a relative large amount of a particular small peptide. The artificially synthesized peptide appeared to suppress DAP induction in CDCs. Possibly, release of the peptide contributes to the prevention of afterdischarge induction in senescent CDCs. The results so far indicate that in senescent Lymnaea neurons electrophysiological functions persist even after long periods of inactivity and severe morphological reduction. PMID- 15270242 TI - Pacemaker potentials are the physiologic basis of epileptiform activity in the buccal ganglia of Helix pomatia. AB - Mechanisms of epileptic activity in nervous systems were studied using the identified neurons B1 through B4 in the buccal ganglia of the snail Helix pomatia as a model system. Activities were recorded with intracellular microelectrodes. Epileptiform activity was induced by bath application of an epileptogenic drug (pentylenetetrazol: 1 mM to 40 mM, or etomidate: 0.1 mM to 1.0 mM). Epileptiform potentials recorded from the somata of neurons consisted of paroxysmal depolarization shifts (PDSs). With increasing concentration of an epileptogenic drug, pacemaker potentials in neuron B3 developed into PDS. Simultaneously several types of chemical post-synaptic potentials were suppressed in amplitude. Since on the one hand epileptic seizures only appear when PDS are synchronized in many neurons and since on the other hand synaptic potentials were found to be suppressed during epileptic conditions, mechanisms underlying neuronal synchronization were studied. Evidence was found that, under epileptogenic conditions only, neurons were synchronized by an non-synaptic release of substances. Strong depolarizations accompanied by an increase in intracellular calcium concentration are known to induce an unspecific exocytosis. Thus, an unspecific exocytosis from the dendrites of PDS-generating neurons probably appears under epileptic conditions and synchronizes neighbouring neurons. PMID- 15270243 TI - Continuous increase of epileptogenic effects following application of proteolytic enzymes (buccal ganglia of Helix pomatia). AB - Epileptic activity of neurons consists of paroxysmal depolarization shifts (PDS) which can be induced presumably in any nervous system by application of an epileptogenic drug. The spontaneous appearance of epileptic activity, however, is based on a largely unknown process which increases susceptibility to epileptic activity (seizure susceptibility in man). It is presently shown that the treatment of ganglia with proteolytic enzymes (Pronase) decreases the effective concentration of epileptogenic drugs, i.e. increases seizure susceptibility. Since proteolytic enzymes are known to primarily affect glial cells a contribution of glia to seizure susceptibility is discussed. PMID- 15270244 TI - Neuron-microglia communication in the CNS of the freshwater snail Planorbarius corneus. AB - The aim of the present study was to identify molecules that may be involved in neuron-microglia communication in the CNS of freshwater snail Planorbarius corneus. Messenger molecules are exchanged in normal and pathological conditions and we tried to identify some of them by immunocytochemistry on whole ganglia and cell cultures. In particular, we examined neurons and microglia for the expression of some cytokines, IL-1alpha, IL-1beta, IL-6 and TNF-alpha and the neurotransmitter glutamate. These substances may be released by suffering or injured neurons and communicate to microglia the damaging event. Even microglia, on own turn, once activated, express and released the same or other substances in order to reestablish the system homeostasis, depending on modalities and times of activation. We discuss the possibility that hyperactivated microglia can shift from neuroprotective to neurodegenerative. Moreover, we examined in neuron microglia co-coltures the direct interaction effects in terms of neuronal survival and improved neurite regeneration. PMID- 15270245 TI - Microglia proliferation as a response to activation in the freshwater snail Planorbarius corneus: a BrdU incorporation study. AB - Invertebrate microglia constitute a class of cells resident in the ganglionic nervous system which are activated after tissue injury or by the presence of pathogens. The microglia activation response includes graduated morpho-functional and biochemical changes and cell proliferation. In this study we verified in the freshwater snail Planorbarius corneus that an activation caused by a traumatic event may induce microglia division. Cell proliferation was assessed immunocytochemically using BrdU incorporation technique and documented on both ganglionic sections and microglia cultured cells at different experimental conditions and times after activation. In addition, we studied the possibility of increasing microglia proliferation by adding to the cultured medium the Macrophage-Colony Stimulating Factor (M-CSF) that has been shown to stimulate specifically this process in vertebrates. PMID- 15270246 TI - Induction of metamorphosis in the marine gastropod Ilyanassa obsoleta: 5HT, NO and programmed cell death. AB - The central nervous system (CNS) of a metamorphically competent larva of the caenogastropod Ilyanassa obsoleta contains a medial, unpaired apical ganglion (AG) of approximately 25 neurons that lies above the commissure connecting the paired cerebral ganglia. The AG, also known as the cephalic or apical sensory organ (ASO), contains numerous sensory neurons and innervates the ciliated velar lobes, the larval swimming and feeding structures. Before metamorphosis, the AG contains 5 serotonergic neurons and exogenous serotonin can induce metamorphosis in competent larvae. The AG appears to be a purely larval structure as it disappears within 3 days of metamorphic induction. In competent larvae, most neurons of the AG display nitric oxide synthase (NOS)-like immunoreactivity and inhibition of NOS activity can induce larval metamorphose. Because nitric oxide (NO) can prevent cells from undergoing apoptosis, a form of programmed cell death (PCD), we hypothesize that inhibition of NOS activity triggers the loss of the AG at the beginning of the metamorphic process. Within 24 hours of metamorphic induction, cellular changes that are typical of the early stages of PCD are visible in histological sections and results of a terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay in metamorphosing larvae show AG nuclei containing fragmented DNA, supporting our hypothesis. PMID- 15270247 TI - Embryogenesis of the histaminergic system in the pond snail, Lymnaea stagnalis L.: an immunocytochemical and biochemical study. AB - Embryogenesis of the histaminergic system in the pond snail, Lymnaea stagnalis, was investigated by means of immunocytochemistry and HPLC assay. From the earliest onset of the of histamine-immunoreactive (HA-IR) elements, the labelled neurons were confined to the pedal, cerebral and buccal ganglia, whereas no IR cells within the pleural, parietal and visceral ganglia were detectable during the embryogenesis. Peripheral projections of the embryonic HA-IR neurons were missing. No transient HA-IR neurons could be found either inside or outside the CNS. The first HA-IR elements appeared at about E55% of embryonic development, at the beginning of metamorphosis, and were represented by three pairs of neurons located in the cerebral ganglia. Following metamorphosis, four pairs of HA-IR neurons were added; two of them occurred in the pedal (E65% stage of development) and two in the buccal (E90% stage of development) ganglia. During embryogenesis, HA-IR fibers were present in the cerebro-pedal connectives and in the cerebral, pedal and buccal commissures, whereas only little arborization could be observed in the neuropil of the ganglia. HPLC measurements revealed a gradual increase of HA content in the embryos during development, corresponding well to the course of the appearance of immunolabeled elements. It is suggested that the developing HAergic system plays a specific role in the process of gangliogenesis and CNS plasticity of embryonic Lymnaea. PMID- 15270249 TI - Embryogenesis of GABAergic elements in the nervous system of Eisenia fetida (Annelida, Oligochaeta). AB - The appearance and development of the GABA-immunoreactive nervous elements in the central nervous system of Eisenia fetida were studied by immunocytochemistry. The nervous system originates from the neuroectoderm situated on the ventral side of the embryo. The organization of the circumpharyngeal ring starts earlier than that of the ventral cord. In the elementary ring the first GABA-immunopositive neurons can be observed (E1 stage) around the mouth. Later the cell number gradually increases and parallel to this process the elementary ring is separeted into a superficial and a deeper portion. The brain and the subesophageal ganglion will be organized from the superficial ring, while the nervous elements of the deeper ring will give rise for the first GABA-immunoreactive elements of the stomatogastric nervous system. In the early stages of the embryogenesis the immunoreactive cells of the developing brain appear solitary, while from the stage E4 they gradually are observed in groups. According to their position, these cell groups are similar to those observed in the brain of the adult earthworms. During embryogenesis the level of the ventral cord ganglia depends on their position in the ectodermal germ bands. It means, that the more organized ganglia are near the circumpharyngeal ring, mean while less developed ganglia are situated caudally from them. By the end of the embryogenesis all ganglia of the ventral cord will be equally well organized. The nerve tracts of the ganglia are built up from contra- and ipsilateral by projected fibres. From E3 stage the medial tracts, mean while from the E4 stage the lateral tracts begin to be formed. During the next stages, more and more fibres connect to the both tracts. At hatching, the development of the central nervous system of Eisenia fetida is not completed, the process is continued during the postembryonic development. PMID- 15270248 TI - Serotonergic and dopaminergic influence of the duration of embryogenesis and intracapsular locomotion of Lymnaea stagnalis L. AB - The role of the dopaminergic and serotonergic system was studied during the embryonic development of the pond snail Lymnaea stagnalis, with special attention to the effect of dopamine and serotonin as well as their agonists and antagonists on the rotation of the veliger larvae, and to the effect of precursors and inhibitors of the synthetizing enzymes on the duration of the embryonic life. Serotonin, D-lysergic acid diethylamide and N,N-dimethyltryptamine increased at a concentration of 1 microM the rotation by 50%, 90% and 87% respectively, and among them D-Lysergic acid diethylamide was found to be the most potent agonist. Other serotonergic agonists and antagonists enhanced the frequency of the rotation (from 165% to 355%) at higher threshold concentrations in the following rank order: methysergid > tryptamine > 2,5-dimethoxy-4-iodoamphetamine > 5 carboxyamidotryptamine > bromo-lysergic acid diethylamide > 7-methyltryptamine. Application of 1-(2-methoxyphenyl) piperazine decreased the rotation by 76%. The reuptake inhibitor desipramine completely blocked the rotation and killed the embryos. Dopaminergic agonists accelerated the rotation by 62% to 233%, and their effect was ranged as follows: dopamine > apomorphine > m-tyramine approximately equal to p-tyramine. Chlorpromazine at 100 microM concentration killed the embryos. At a concentration of 100 microg/ml, tyrosine, the precursor of DA, slowed down the embryonic development by increasing the duration of the embryonic life from 8 to 10 days. Decarboxylase inhibitors, alpha-methyl-3,4 dihydroxyphenyl-alanine (25 microg/ml) and m-hydroxybenzylhydrazin (5 microg/ml), killed 50% of the embryos, meanwhile the rest hatched ten days later, compared to the control animals. The development was partially blocked by the serotonin precusor L-tryptophane (50 microg/ml). Trytophan hydroxylase blocker, p chlorphenylalanine (50 microg/ml) resulted in a distortion of the body pattern of the embryos, and prevented the hatching of most (95%) of the animals. PMID- 15270250 TI - RFamide neuropeptide actions on the molluscan heart. AB - FMRFamide and the related tetrapeptide FLRFamide are highly excitatory in molluscan non-cardiac smooth muscle. They are also exceptionally excitatory in the atrium and internally perfused ventricle of Busycon canaliculatum. These two peptides, usually thought of as classic molluscan cardio-acceleratory agents are in fact simply two members of a large and ever growing superfamily, the RFamide family, whose phylogenetic distribution has been so elegantly mapped by Walker. Members of this family, often with extended peptide chains (e.g. penta, hepta and decapeptides), stretch in their known distribution from the cnidaria to the chordates. The effects of some of the members of this superfamily (FMRFamide. FLRFamide, YMRFamide, TNRNFLRFamide, SDPFLRFamide, LMS) were examined. The neuropeptides were found to be very potent at very low concentrations (10(-9) M) in the ventricle of both Buccinium and Busycon. Other neuropeptides (HFMRdFamide, SCPb, NLERFamide and pEGRFamide) were found to be without any effect. The Ca2+ dependency of these neuropeptides was also tested. The peptides appear to induce contraction of the ventricles by release of Ca2+ from internal pools. The neuropeptides appear to stimulate contraction in these cardiac muscles through a completely different pathway to Serotonin (the main excitatory neurotransmitter for the cardiac muscle). When the peptides were applied together with Serotonin an additive effect was observed clearly indicating the release of Ca2+ through different pathways. The nature of the RFamide receptor was also tested. It appears that the RFamide neuropeptides mobilize the 2nd messenger IP3 (Inositol trisphosphate), since the IP3 blocker Neomycin Sulphate inhibited the response of the neuropeptides. PMID- 15270251 TI - A structure-activity study of the neuropeptide PF1, SDPNFLRFamide, using the dorsal body wall muscle of the chicken nematode, Ascaridia galli. AB - The action of a range of N terminally modified peptides structurally related to the nematode peptide PF1, SDPNFLRFamide, has been investigated using a dorsal muscle strip preparation from the chicken nematode, Ascaridia galli. Acetylcholine contracts this muscle preparation in a concentration-dependent manner when applied in the range 1-100 microM with an EC50 value of 9 microM. These contractions are reduced in the presence of PF1 and its analogues, with a threshold effect of PF1 of around 1 nM and an IC50 value of 470 nM against 10 microM acetylcholine. All the PF1 analogues tested were less potent than PF1 in reducing the acetylcholine contractions, indicating the importance of the N terminal amino acids in the action of PF1 in this preparation. PMID- 15270252 TI - Opiate alkaloids in Ascaris suum. AB - The parasitic worm Ascaris suum contains the opiate alkaloids morphine and morphine-6-glucuronide as determined by HPLC coupled to electrochemical detection and by gas chromatography/mass spectrometry. The level of morphine in muscle tissue of female and male is 252 +/- 32.68, 1168 +/- 278 and 180 +/- 23.47 (ng/g of wet tissue), respectively. The level of M6G in muscle tissue of female and male is 167 +/- 28.37 and 92 +/- 11.45 (ng/g of wet tissue), respectively. Furthermore, Ascaris maintained for 5 days contained a significant amount of morphine, as did their medium, demonstrating their ability to synthesize the opiate alkaloid. The anatomic distribution of morphine was examined by indirect immunofluorescent staining and HPLC of various tissues dissected from male and female adult worms. Immunofluorescence revealed morphine in the subcuticle layers, in the animals' nerve chords and in the female reproductive organs. Morphine was found to be most prevalent in the muscle tissue and there is significantly more morphine in females than males, probably due to the large amounts in the female uterus. Morphine (10(-9) M) and morphine-6-glucuronide (10( 9) M) stimulated the release of NO from Ascaris muscle tissue. Naloxone (10(-7) M), and L-NAME (10(-6) M) blocked (P < 0.005) morphine-stimulated NO release from A. suum muscle. CTOP (10(-7) M) did not block morphine's NO release. However, naloxone could not block M6G stimulated NO release by muscle tissue, whereas CTOP (10(-7) M) blocked its release. These findings were in seeming contradiction to our inability to isolate a mu opiate receptor messenger RNA by RT-PCR using a human mu primer. This suggests that a novel mu opiate receptor was present and selective toward M6G. PMID- 15270253 TI - Clinical study of qingluo tongbi granules in treating 63 patients with rheumatoid arthritis of the type of yin-deficiency and heat in collaterals. AB - The study is to observe the therapeutic effects of qingluo tongbi granules (QTG) in patients with rheumatoid arthritis (RA) and the changes of immune indexes. In this series there are 63 patients with RA of the type of yin-deficiency and heat in collaterals treated with QTG as the treated group and 55 patients of the same type treated with Tripterygium glycosides as the control group. As a result, in the treated group, the curative rate is 9.52% and markedly effective rate 38.10%, with a total effective rate of 90.48%, while the corresponding rates in the control group are 0, 20.00% and 83.64%, respectively. The curative effect in the treated group is better than that in the control group (P<0.05). Besides, no obvious adverse reactions are found in the treated group. Therefore it is concluded that as a new medicinal preparation QTG is safe and effective in the treatment of RA. PMID- 15270254 TI - Theories and practice in prevention and treatment principles in relation to Chinese herbal medicine and bone loss. AB - Osteoporosis is a world wide problem that is increasing in significance as the global population both increases and ages. While osteoporosis has been extensively studied in recent years, the utilization of Traditional Chinese Herbal Medicine for the prevention and treatment of this condition have seldom been examined in the Western world. This paper reviews the theories and the literature that relate to prevention and treatment of bone loss at the time of menopause according to the principles of Traditional Chinese Herbal Medicine. Practical developments in these areas are also illustrated in this paper based on the authors' research findings in recent studies. PMID- 15270255 TI - Clinical application of the yang-activating method in acupuncture treatment. PMID- 15270256 TI - Dr. Li Yueqing's experience in treating female urethral syndrome. PMID- 15270257 TI - Jian shu wen qing tang used in the treatment for 60 cases of irritable bowel syndrome. PMID- 15270258 TI - Treatment of postmenopausal osteoporosis with yang huo san zi tang--a report of 30 cases. PMID- 15270259 TI - Clinical experience in the use of shaoyao gouteng muer tang. PMID- 15270260 TI - A combined TCM treatment for 72 cases of bony gonitis. PMID- 15270261 TI - Clinical application of the empirical prescriptions for chronic pelvic inflammation. PMID- 15270262 TI - Dr. Xuan Guowei's experience in treating dermatosis. PMID- 15270263 TI - A combined therapy for cervical spondylopathy. PMID- 15270264 TI - Clinical application of point Diji. PMID- 15270265 TI - Electroacupuncture treatment for 30 cases of anxiety neurosis. PMID- 15270266 TI - Clinical observation on the therapeutic effects of heavy moxibustion plus point injection in treatment of impotence. PMID- 15270267 TI - Moving flash-fire cupping along the channels--a new method for treating urticaria. PMID- 15270268 TI - Electroacupuncture treatment for 45 cases of postapoplectic dysphagia. PMID- 15270269 TI - Clinical study on acupuncture treatment of pseudobulbar paralysis. PMID- 15270270 TI - General situation on the study of inspection of sublingual veins. PMID- 15270271 TI - How to treat allergic rhinitis. PMID- 15270272 TI - Topography of acupoint Jianjing (GB 21). PMID- 15270273 TI - Effects of electroacupuncture on learning, memory and formation system of free radicals in brain tissues of vascular dementia model rats. AB - In order to observe the regulative effect of electro-acupuncture on the formation system of free radicals in the brain tissues and learning and memory in vascular dementia (VD) model rats, the Morris's water labyrinth was used for testing the learning ability and memory in VD model rats made by 4-vessel occlusion method, and the activities or contents of nitric oxide (NO), NO synthase (NOS), superoxide dismutase (SOD), malondialdehyde (MDA), glutathione peroxidase (GSH Px) were determined. Results showed that the mean escape latency in the electro acupuncture group was markedly reduced in place test, and the times swam the place of the plate-form in the original plate-form quadrant were significantly more than those in the rest three quadrants in spatial probe test as compared with the model group. In the electro-acupuncture group and the nimodipine group the contents of NO and MDA and the activity of NOS were decreased, while the activities of SOD and GSH-Px were increased. It is indicated that electro acupuncture can modulate the production and clearance of free radicals, and improve the ability of learning and memory of the VD model rats. PMID- 15270274 TI - Effects of acupuncture on myelogenic osteoclastogenesis and IL-6 mRNA expression. AB - The effect of acupuncture and moxibustion on myelogenic osteoclastogenesis and IL 6 mRNA expression is investigated. The result turns to be that the number of myelogenic osteoclasts in the model group is obviously bigger than in the sham operation group (P<0.01), and that in the acupuncture group is markedly smaller than in the model group (P<0.01). The IL-6 mRNA expression in the marrow cells of the model group is significantly elevated, as compared with that in the sham operation group (P<0.01), while the its elevation is lower in the acupuncture group than in the model group (P<0.01). It is therefore concluded that 1) acupuncture effectively modulates the secretion of and then to reduces IL-6 mRNA expression and 2) acupuncture decreases the number of myelogenic osteoclasts. PMID- 15270275 TI - Effects of bushen huoxue decoction on nitric oxide (NO) in serum, articular cartilage and synovium in rabbits of knee osteoarthritis. AB - Forty-eight New Zealand rabbits were divided into normal group (n=18), control group (n=18) and Chinese herbs treatment group (n=12) randomly. The rabbits in the normal group received sham-operation, and the OA model was established by Hulth's method. All the rabbits in the treatment group were given bushen huoxue decoction from the 6th week after the operation. At 6th, 8th and 12th week after the operation, the NO concentrations of the serum, joint cartilage and synovium were examined. RESULTS: Indicated that the NO concentrations of the serum, joint cartilage and synovium in the control group were all significantly higher than those in the normal group, with the joint cartilage more obvious (P<0.05). In the Chinese herbs treatment group the NO concentrations in all the parts obviously decreased as compared with the control group (P<0.05). It is suggested that bushen huoxue decoction decrease the levels of NO in the serum, synovium and joint cartilage in the OA rabbit. PMID- 15270276 TI - Progress in using combination of Chinese drug with chemotherapy to treat cancer. PMID- 15270277 TI - [Guidelines for the evaluation and treatment of patients with heart arrhythmia]. PMID- 15270278 TI - [Fuzzy logic in urology. How to reason in inaccurate terms]. AB - The Occidental thinking is basically binary, based on opposites. The classic logic constitutes a systematization of these thinking. The methods of pure sciences such as physics are based on systematic measurement, analysis and synthesis. Nature is described by deterministic differential equations this way. Medical knowledge does not adjust well to deterministic equations of physics so that probability methods are employed. However, this method is not free of problems, both theoretical and practical, so that it is not often possible even to know with certainty the probabilities of most events. On the other hand, the application of binary logic to medicine in general, and to urology particularly, finds serious difficulties such as the imprecise character of the definition of most diseases and the uncertainty associated with most medical acts. These are responsible for the fact that many medical recommendations are made using a literary language which is inaccurate, inconsistent and incoherent. The blurred logic is a way of reasoning coherently using inaccurate concepts. This logic was proposed by Lofti Zadeh in 1965 and it is based in two principles: the theory of blurred conjuncts and the use of blurred rules. A blurred conjunct is one the elements of which have a degree of belonging between 0 and 1. Each blurred conjunct is associated with an inaccurate property or linguistic variable. Blurred rules use the principles of classic logic adapted to blurred conjuncts taking the degree of belonging of each element to the blurred conjunct of reference as the value of truth. Blurred logic allows to do coherent urologic recommendations (i.e. what patient is the performance of PSA indicated in?, what to do in the face of an elevated PSA?), or to perform diagnosis adapted to the uncertainty of diagnostic tests (e.g. data obtained from pressure flow studies in females). PMID- 15270279 TI - [Dr. Victor Molla Frambuena (1901-1972): biography and analysis of his urologic work]. AB - OBJECTIVES/METHODS: Dr. Victor Molla Frambuena developed his practice in Valencia during his whole professional career, following the steps of his father (the prestigious Professor Rafael Molla Rodrigo), in the disappeared Red Cross Hospital where he became the first chairman of the department of urology. He published all his scientific work, which we analyze in this article, in this city, mainly in the journal "Cronica Medica". RESULTS/CONCLUSIONS: After an exhaustive search, we analyze all the data about his medical biography and his scientific publications, this latter mainly in the journal Cronica Medico from 1928 to 1938, the year in which the last number was published. PMID- 15270280 TI - [Female urethra diverticula]. AB - OBJECTIVES: To remind the most relevant features on the presenting clinical picture, diagnosis and treatment of this disease, which is not uncommon although many times is not suspected. METHODS: We review five cases of female urethra diverticula diagnosed in our department over the last five years. We describe the clinical picture, physical examination, diagnostic tests, as well as treatment undertaken in each, comparing them with current bibliography up to date. RESULTS: 1-Three out of five patients presented a tumor in the anterior vaginal wall; one had stress urinary incontinence and the other recurrent urinary tract infection. 2-The diagnostic methods employed were urethroscopy, retrograde and voiding urethrography, transvaginal ultrasound, and pelvic MRI. The lost two cases were diagnosed by MRI as the only diagnostic test. 3-Surgical treatment was chosen in all cases, being transvaginal diverticulectomy the chosen operation. One patient underwent transurethral diverticulectomy with the Sachse urethrotome for a post operative recurrence. CONCLUSIONS: Female urethra diverticulum is a clinical entity many times underdiagnosed that should be suspected in every patient with chronic lower urinary tract symptoms. We have several available imaging tests which can confirm the working diagnosis, either alone or in combination, being MRI the newest. Surgical treatment has demonstrated to be curative, with the transvaginal technique as the most effective and therefore the one of choice. PMID- 15270281 TI - [Cystocele repair with a polypropylene mesh: our experience]. AB - OBJECTIVES: To evaluate the efficacy of the polypropilene mesh shaped in a T, with a circular area (to repair the cystocele) and an anterior extension (to function as a tension free sling), for the combined treatment of cystocele and urinary incontinence. METHODS: Retrospective study including 31 female patients with cystocele, with or without urinary incontinence, undergoing mesh repair. Mean age was 62.3 yr. (range 55-72). All patients were multiparous. Number of childbirths varied between 1 and 4. Mean follow-up was 23.5 months (range 12 to 29 months). 80% had grade III cystocele and 20% grade IV. 28 patients (90.3%) presented with urinary incontinence and 3 (9.67%) urgency without incontinence. 16 patients had previously undergone hysterectomy and another 6 surgery for urinary incontinence (2 Raz and 4 Burch operations). 11 patients (33.6%) needed a combination technique in the same operation: associated vaginal hysterectomy in 4 patients and posterior mesh colporrhaphy for grade III symptomatic rectocele in 7 patients. RESULTS: No patient had prolapse recurrence, 1 patient needed clean intermittent catheterizations for 3 months, and 3 suffered de novo urgency (with good response to anticolinergic drugs). The worsening of stress urinary incontinence (SUI) in one patient was solved by suburethral sling insertion; another patient who had prolonged postoperative vaginal bleeding requiring blood transfusions and subsequent mesh erosion of the vaginal wall (she underwent re operation to cut off the mesh). 3 patients complained of intercourse discomfort which disappeared after an average of 3 months. No other remarkable intra or postoperative complications appeared. CONCLUSIONS: The polypropilene mesh associated with a sling is an effective treatment to repair cystocele with or without SUI, although long-term studies with a greater number of patients are required to validate the technique. PMID- 15270282 TI - [Perineal radical prostatectomy as monotherapy: ten-year experience]. AB - OBJECTIVES: To evaluate the clinical and oncological results of perineal radical prostatectomy (PRP) as monotherapy. METHODS: We include our initial series of 115 consecutive patients with prostate cancer without neoadjuvant hormonal therapy undergoing perineal radical prostatectomy as monotherapy from November 92, when we decide to abandon laparoscopic lymphadenectomy in patients deemed at low risk of N+ (PSA = 10 and Gleason score = 7 and organ confined or suspicion of minimal extra capsular extension on ultrasound). Functional results are compared with the first 115 consecutive patients in our own series of retropubic radical prostatectomy (RRP). No patient received adjuvant therapy and deferred intermittent androgen blockade was only applied when patients with biochemical progression reached a PSA of 4 ng/ml. RESULTS: After a mean follow-up of 57.7 months (3-130) we obtained: global survival 98.3%, disease specific 100%; biochemical progression 13.9% (161, pT 2 (5.2%), 32pT3a (21.8%), 7pT3b-pT4a (71.4%)). Incidence of positive margins 41.7% (52.1% unifocal with a progression rate of only 12%). 87% of the patients did not need transfusion during or after surgery. Urinary continence was 96.5% and the probability of potency preservation was 34.7%. Among 16 patients with biochemical progression, only 4 required deferred intermittent androgen blockade (1 cycle in 3 patients and two in the remainder). CONCLUSIONS: PRP without lymphadenectomy as monotherapy offers excellent oncological and functional results equivalent to RRP, which favour the perineal approach: short-term operative times, lower transfusion rate, possibility of regional anesthesia, better tolerance with minimal analgesia requirements, and shorter hospital stay. Watchful waiting in the follow-up of patients in biochemical progression with a deferred intermittent androgen blockade regimen offers an excellent quality of life, with survivals similar to any other therapeutic option after this follow-up time. PMID- 15270283 TI - [Descriptive epidemiology of bladder exstrophy in an area of Madrid]. AB - OBJECTIVE: To describe the procedures and complications of bladder exstrophy closure, epispadias repair and bladder neck surgery to achieve urinary control. METHODS: Retrospective chart review of 11 patients with the exstrophy-epispadias complex. RESULTS: Early bladder closure is performed after birth with or without concomitant pelvic osteotomy. The preferred procedures are: the Cantwell-Ransley technique for epispadias repair, the Young-Dees-Leadbetter procedure for bladder neck reconstruction and ileocystoplasty with Mitrofanoff for bladder augmentation. CONCLUSIONS: Augmentation and continent diversion procedures can increase the functional capacity of the exstrophic bladder, and allow the vast majority of patients to achieve continence and preserve renal function. Bladder lithiasis is the most significant complication in these patients. PMID- 15270284 TI - [Intraoperative complications and morbidity of laparoscopic radical prostatectomy (LRP) during the learning curve]. AB - OBJECTIVES: To analyze the complications and morbidity during our learning curve of laparoscopic radical prostatectomy (LRP) and compare them with other published series. METHODS: We review the 25 first laparoscopic radical prostatectomies performed in our department, evaluating the operative technique and other features such as surgical time, blood loss, complications and conversion to open surgery. We also evaluate morbidity, postoperative hospital stay, and functional features such as potency and continence. RESULTS: LRP was completed in 22 patients. Overall intraoperative complication rate was 32%. 3 cases were converted to open surgery due to technical difficulties or intraoperative complications. We had complications in 4 patients, that were not severe (bladder injury 2 cases, and epigastric artery injury another 2) and where solved without difficulties during the operation. The most severe intraoperative complication was related to the anesthesia procedure at the time of extubation of a patient who required tracheotomy. There were no severe postoperative complications, being leakage from the anastomosis the most common (7 cases). All of them were managed conservatively, although this resulted in a mean hospital stay of 10.8 days. 2 patients required endoscopic procedures in the immediate postoperative time for bladder catheter repositioning. All patients suffered erectile dysfunction and the continence rate at 3 months was 77.2%. CONCLUSIONS: Although LRP is a long operation and difficult during the learning curve, its complication rate is acceptable because they are not severe and can be managed in a relatively easy way. PMID- 15270285 TI - [Fatal evolution of a renal angiosarcoma]. AB - OBJECTIVES: Report a new case of renal angiosarcoma treated by surgery and adjuvant chemotherapy with bad results. METHODS: 72-year-old male undergoing right nephrectomy for renal tumor. Pathology reports renal angiosarcoma. RESULTS: Three months after surgery patient refers lumbar pain and hemoptysis and CT scan reveals the existence of multiple bone and lung metastasis; a regimen of systemic chemotherapy with Doxorrubicine+ Ifosfamide was started without response; he died two months later. CONCLUSIONS: Primary renal angiosarcoma is very rare, with less than 10 cases in the literature before 1998, and it is always associated with bad prognosis. Diagnosis is based on immunohistochemical studies (antibodies against CD31, CD34 and factor VIII related antigen) to define the endothelial differentiation of the tumor. There is no experience to define the best therapeutic strategy against this entity. PMID- 15270286 TI - [Leydig cell tumor presenting as precocious pseudopuberty in a 4-year-old boy]. AB - OBJECTIVES: To report one case of isosexual precocious pseudopuberty in a 4-year old boy caused by an interstitial cell testicular tumor. METHODS: Physical exam, blood tests, hormonal determinations, adrenal suppression tests, bone age, orchiectomy and pathologic study of the specimen were performed. RESULTS: Physical examination showed a boy with muscle development, acne; body, sexual and face hair corresponding to an older boy; with increased volume of the left testicle and infantile contralateral testicle. Urinary 17-ketosteroids were elevated and did not decrease after dexametasone. Bone age corresponded to an 11 year-old standard. Pathologic study showed an interstitial cell tumor. Puberty changes disappeared after orchiectomy. CONCLUSIONS: This diagnosis should be taken into consideration in every case of accelerated sexual development in a boy with testicular tumor and without maturation of the contralateral testicle. PMID- 15270287 TI - [Atypical or bizarre leiomyoma of the scrotum. Report of one case and bibliographic review]. AB - OBJECTIVES: To describe the clinicopathological characteristics of the scrotal bizarre leiomyoma in order to increase our understanding, and avoid the possibility of erroneous diagnosis and treatment. RESULTS: We report the case of a 43 year-old patient with a pendulous nodular scrotal tumour, which after microscopic study could be seen to be of smooth muscular origin, corresponding, because of its peculiar histological characteristics, to the rare variety called atypical or symplasmic bizarre leiomyoma. CONCLUSION: Although it is a form of leiomyoma which is extremely infrequently found in the scrotum, it is necessary to know it can occur and to know that we are dealing with a benign tumour that should be treated as such. PMID- 15270288 TI - [Multilocular cystic nephroma. A diagnostic and therapeutic challenge. Report of two cases]. AB - OBJECTIVES: Multilocular cystic nephroma is a rare benign entity grouped among the cystic non genetic diseases, which usually presents with a clinical picture and radiological features indistinguishable from malignant neoplasias, making impossible to rule out malignancy preoperatively. METHODS: We report two cases of multilocular cystic nephroma which were treated with different surgical attitudes despite their clinical and radiological similarities, because on the second case intraoperative pathologic study of the specimen was performed. RESULTS: The diagnosis of multilocular cystic nephroma was confirmed in both cases, with the difference that radical nephrectomy was performed in the first case whereas the intraoperative study of the tumor in the second allowed to perform tumorectomy with renal parenchyma preservation. CONCLUSIONS: We consider that in the presence of a multiloculated renal mass of complex appearance and benign clinical behavior, the intraoperative study of the tumor will avoid performance of radical surgery. PMID- 15270289 TI - [Extrarenal retroperitoneal angiomyolipoma: bibliography review and report of a new case]. AB - OBJECTIVES: To report the 7th case of pararenal angiomyolipoma published in the world literature and to review the international bibliography. METHODS: We report the case of a 46-year-old female with history of renal colic and a complex mass on radiological tests. RESULTS/CONCLUSIONS: Extrarenal retroperitoneal angiomyolipoma is a rare pathology with no more than 7 published cases. The diagnostic difficulty and radiological similarities with liposarcoma make surgery the treatment of choice. PMID- 15270290 TI - [Conservative treatment of a seminal vesicle abscess. Report of one case]. AB - OBJECTIVE: To describe the clinical conditions, the radiological findings and the management of a case of seminal vesicle abscess. METHODS: A 47-year old man presented with irritative voiding symptoms, fever, diminished ejaculated volume, hematuria and testicular pain. Diagnosis was made with digital rectal examination, ultrasound and CT. RESULTS: The patient was managed with antibiotic therapy alone for 4 weeks. Clinical and radiological resolution was achieved. CONCLUSIONS: Seminal vesicle abscess is a rare condition. Diagnosis is based on clinical data and radiological findings. Conservative treatment could be effective in selected cases. PMID- 15270291 TI - Long-term study to assess the efficacy of tamsulosin in the control of symptoms and complications developed in patients with symptomatic benign prostatic hyperplasia (OMNICONTROL study): first-year follow-up report. AB - OBJECTIVE: To evaluate: i. long-term efficacy of tamsulosin in the control of lower urinary tract symptoms (LUTS) suggestive of benign prostatic obstruction (BPO) using the I-PSS questionnaire, ii. the frequency of complications related to the disease, and iii. short and long-term tolerability of tamsulosin. METHODS: A total of 2.921 patients with LUTS suggestive of BPO for more than 6 months and total IPSS > 7 treated with tamsulosin (Omnic) in real life practice conditions in Spain entered an observational prospective multicentre clinical study. Efficacy was primarily assessed by changes from baseline to endpoint in I-PSS symptoms score (total, irritative and obstructive), and secondarily by the appearance of disease complications, and urinary flow measurements. Safety was assessed recording every suspected adverse reaction, blood pressure changes and laboratory data on months 6 and 12. Evolution in time of free flow and sonographical evaluation of the prostate were also obtained in 663 (22.7%) and 1346 (46.1%) cases, respectively, and the use of previous and concomitant medication was also analysed. RESULTS: After 6 and 12 months total I-PSS, irritative, and obstructive symptoms were significantly reduced with the use of tamsulosin 0,4 mg once daily. At 1 year follow-up total I-PSS score, irritative symptoms, and obstructive symptoms were reduced in 8.2, 3.5 and 4.8 points 146%, 45% and 48% improvement), respectively (p < 0.0001). The proportion of patients seriously symptomatic (total I-PSS score 20-35) was reduced from 34.8% at the start of the study to 8% at 6 months and 2.9% at 12 months. Mean QoL also significantly improved after 6 and 12 months of treatment. Average score QoL index was reduced from 4.1 to 1.86 after 12 months (2.24 points, 55% improvement) (p < 0.0001). Qmax also significantly improved after 6 and 12 months of treatment (p < 0.0001). The good tolerability profile of tamsulosin has been confirmed after 6 and 12 months of treatment. CONCLUSIONS: Therapeutic intervention with tamsulosin 0,4 mg once daily is effective in all parameters analysed (I-PSS questionnaire and flow study), very well tolerated and safe in the short-term (6 and 12 months) in patients with LUTS suggestive of BPO. Long-term data specifically regarding the decrease in prostate volume and the evolution of the BPH condition will be welcome. PMID- 15270292 TI - Experimental methods for "in vivo" study of detruso-sphincteric pharmacological response: a critical review. AB - OBJECTIVE: We review "in vivo" methods most commonly used for the investigation of detruso-sphincteric pharmacological response. We compare this information with the procedures used in our Institution for these purposes. METHODS: The medical databases MEDLINE, EMBASE and Pascal Biomed were searched to identify articles on this subject. The methods used have been critically analyzed and compared with the methods used in our experiments. RESULTS: The "in vivo" dynamic investigation of vesico-sphincteric function began at our Institution in the late 70s. Methods for the study of vesical or urethral dynamic behaviour have been devised to be applied independently or simultaneously. Great difficulties have been encountered, both in our experience and in the work of revised authors, in the integrated investigation of Lower Urinary Tract function. Methods to overcome these drawbacks have been proposed. CONCLUSIONS: Methods for "in vivo" studies of physiological and pharmacological detruso-sphincteric function are presented and compared with current procedures found in the literature. It is highlighted that an integrated method for the simultaneous study of vesico-sphincteric function is a difficult challenge yet to be taken up. PMID- 15270293 TI - [Multiple sclerosis in children and adolescents: history and current experience of immunomodulating treatment]. AB - An agreement accepted worldwide for treatment of multiple sclerosis (MS) in adult patients has not been achieved so far for children and adolescents, mostly due to the insufficient information on the features of MS with early age-at-onset. New methods of immunomodifying treatment and the conception of early MS treatment have changed the attitude to prognosis of MS with early onset. In such cases, a course of the disease is less aggressive and characterized by more intensive remyelination as well as brain plasticity and less neurological deficit that may predict highest efficacy of the disease modifying treatment timely started in these patients. Such drugs, as beta-interferons, glatiramer acetate and high dosage intravenous immunoglobulines G, which are able to decrease MS relapse frequency and disability progression in adult patients, are poorly studied in children and adolescents. Preliminary positive results of international and Russian investigations on the efficacy and tolerability of immunomodulating therapy of MS with early age-at-onset require further investigations in this field. PMID- 15270294 TI - [Clinical and neurophysiologic aspects of surgical treatment of pharmacoresistant forms of epilepsy]. AB - Surgical treatment of pharmacoresistant forms of epilepsy under neurophysiological monitoring is a key problem studied in A.L. Polenov Russian Neurosurgical Institute (Saint-Petersburg). A summary of long-term studies and main stages of surgical treatment development are presented. The indications and contra-indications, along with basic neurophysiologic strategic and tactic arguments of open and stereotaxic treatment of focal and generalized epilepsy based on neurophysiologic model which determines a role of epileptic focus, epileptic and inhibiting brain systems in spreading and arresting of seizure discharge at each disease stage are formulated. A program of clinico neurophysiologic monitoring of temporal epilepsy in pre- and intraoperative periods is considered. PMID- 15270295 TI - [Cerebral vascular pathology in systemic sclerosis]. AB - Thirty patients with systemic sclerosis (SS), aged 20-55 years, illness duration 1-10 years, were observed. Vasospastic syndrome of different expression predominated in the clinical picture of the disease. Among neurological signs, prevalent were peripheral nervous system lesion--isolated (20% of the cases) or in combination with chronic insufficiency of brain circulation (80%). Vascular pathology was distinctly determined by ophthalmoscopy: angiopathy was found in 42% of the patients, angiospasm--in 25%, angiosclerosis--in 17%. The following types of disorders characterized brain blood flow: distonic (42%), dyscirculatory (33%), normotonic (17%), hypotonic (8%). Besides, 83% of the patients had hemispheric asymmetry of blood flow and difficulties with venous outflow. In some cases, the signs of inner, external and combined hydrocephalia were determined by CT and MRI; more than a half of the patients had dilatation or deformation of brain ventricular system with intracranial hypertension. The data obtained suggest that extra- and intracranial blood flow pathology in SS may, to a large extent, determine both organic brain lesion development in general and the genesis of many symptoms (headaches, vestibular disorders, etc). PMID- 15270296 TI - [A differentiated approach to Parkinson's disease treatment in early stage of the disease]. AB - A conception of the approach to Parkinson's disease treatment at early stage, which could be realized in practice is suggested. The conception is based on the results of dopaminergic drugs treatment of 149 patients, aged 33-87 years. The patient's state was assessed by neurological examination and clinical scales with regard to different disease types, tremor and rigid, and levodopa doses--from moderate (up to 500 mg) to high (above 500 mg). A general treatment policy, including therapy directed to activation of regulatory and compensatory influences and differentiated usage of levodopa and combined therapy with levodopa and dopamine agonists, is presented. PMID- 15270297 TI - [Nonconvulsion epileptic encepholopathies and their treatment]. PMID- 15270298 TI - [A change of immune profile of patients with schizophrenia during treatment]. AB - Cellular and humoral immunological parameters have been studied in 59 schizophrenic patients and 38 healthy controls. Immunological indices (CIC, autoantibodies to cardiolipin) were found to be significantly elevated in patients in the acute disease stage before the treatment. After olanzapin therapy, a level of these parameters decreased and did not differ from that of controls. In patients, irrespective of clinical condition and treatment, functional activity of immunocompetent cells (phagocyte activity of neutrophils and monocytes, cytotoxic activity of lymphocytes natural killers, interleukin-2, interleukin-10 and gamma-interferon production), was significantly lower both before the treatment as after therapy, i.e. did not change during the whole study (28 weeks). In responders, a level of IL-1B production was higher than in controls before and during the treatment. In non-responders, it was similar to that in controls before the therapy, and increased during the treatment to a higher level. PMID- 15270299 TI - [Association study of NcoI and TaqI A dopamine receptor D2 gene polymorphism in opium addicts]. AB - The N2/N2 (NcoI) genotype of DRD2 gene is shown to be a marker of resistance to opium addiction, the N1/N1 genotype being considered as a risk marker for the disorder. The N1 allele was associated with opium addiction in Russian patients aged 16 years and younger at drug using onset. In Tatars, an association was found between the N1 allele and a risk of opium addiction development over 16 years of age. The N2/N2 genotype proved to be a marker of resistance to opium addiction in Tatars over 16 years of age at drug using onset. No association was found between TaqI A DRD2 polymorphism and opium addiction in the Russians and the Tatars. PMID- 15270300 TI - [A case of Canavan-Van Bogaert-Bertrand leukodystrophy]. PMID- 15270301 TI - [Congenital myasthenic syndrome related to homozygous missense mutation in promoter region of acetylcholine receptor epsilon-subunit gene]. PMID- 15270302 TI - [Follow-up study of patients with couvades syndrome]. PMID- 15270303 TI - [Vertebroneurological signs of cervical whiplash injury]. PMID- 15270304 TI - [Usage of a chondroxide ointment in the treatment of patients with spinal osteochondrosis]. PMID- 15270305 TI - [Modern concepts of cerebrospinal fluid filtration and sorption in nervous diseases]. PMID- 15270306 TI - [Pathogenetic bases of cognitive and psychotic disorders in Parkinson's disease]. PMID- 15270307 TI - Employ the law wisely. PMID- 15270308 TI - Endodontic techniques defined by principles. AB - Traditional endodontics, limited to the use of K-files, has proven so inefficient in achieving predictable superior results that its deficiencies led to the development and adoption of rotary NiTi systems that produce far superior results, often with much less physical effort. However, in tight curved canals, the inherent weakness of rotary NiTi to both torsion and cyclic fatigue has led to a troubling number of fractured instruments lodged in the canals. The introduction of the SafeSider instrumentation system eliminates the problems of both traditional K-file instrumentation and the fractured instruments associated with rotary NiTi, while delivering superior results in a time-efficient manner. PMID- 15270309 TI - Head and neck diagnostic challenge. AB - An interesting and unusual clinical case is presented to challenge the dentist's diagnostic acumen. Diagnostic and management consideration will be discussed, and a list of suggested readings is provided. PMID- 15270310 TI - Making esthetic provisionals. AB - The problems that arise from using traditional methyl methacrylate material to construct temporary restorations are discussed. The need for exceptional, esthetic and well-fitting temporaries in a contemporary dental practice is stressed. A practical method for constructing temporary restorations is described in detail. The advantages of the author's method are described. PMID- 15270311 TI - Radiographic oddities: Unusual calcifications in the dental pulp. AB - The author describes two examples of a "smiley-face tooth." PMID- 15270312 TI - Dentists' use, misuse, abuse or dependence on mood-altering substances. PMID- 15270313 TI - Adding flexibility to employee benefit programs. PMID- 15270314 TI - Institute of Medicine report calls for national effort to improve health literacy. PMID- 15270315 TI - A letter to Mrs. Jones. PMID- 15270316 TI - The Joint Commission is watching: is your disaster response plan in order? AB - With fall elections approaching, it's time to revisit your major incident response plan. Multiple practice drills will hone your responses to incidents of different levels of severity. Ensure staff that they will be safe and able to deliver quality care. A hospital incident command system is now a major area of interest for surveyors with the Joint Commission on Accreditation of Healthcare Organizations. PMID- 15270317 TI - Poor communication: root of most patient safety ills. AB - Poor communication in the ED can dramatically affect patient safety and satisfaction. Insist on open communication to ensure your team leader has the information he or she needs to make decisions. Don't assume the leader has all the information you have. Circumstance can project any team member into a leadership position, so everyone must think like a potential leader. With multiple critical patients in the ED simultaneously, a nurse or tech may have to assume temporary leadership for a patient. Review key information after every shift change. PMID- 15270318 TI - Three strategies to reduce overcrowding and gridlock. AB - ED overcrowding leads to lost revenue and service opportunities. Hoag Memorial Hospital Presbyterian in Newport Beach, CA, dropped diversion hours to zero with Code Emergency Saturation Triage (EST). Mobilize all staff to take extraordinary measures to move patients, i.e., immediate assessment of patients who should be moved upstairs and prioritizing bed cleaning. Pre-emptive bed requests can identify patient needs much sooner. Creative staffing, such as creating the position of emergency care admit nurse, can ease the paperwork burden of floor nurses. PMID- 15270319 TI - Make your ED part of a law enforcement team. AB - Your ED can become an integral part of law enforcement response teams. Health care providers and law enforcement personnel must have special training. Preventive medicine is a critical component of Tactical Emergency Medicine Support (TEMS). TEMS can be a valued community service and improve the public's perception of your facility. PMID- 15270320 TI - POC tests cut screening time down to 20 minutes. AB - Point-of-care testing can significantly reduce your door-to-diagnosis times. Choose device(s) that mirror your most common diagnoses. Training techs as well as RNs helps meet needs of the patient. Be prepared to justify cost by demonstrating improved turnaround. PMID- 15270321 TI - 8 questions shed light on best practices. PMID- 15270322 TI - Giving a second chance: expanding the pool of home health aides. PMID- 15270323 TI - Why you should stop physician hospital discharges. PMID- 15270324 TI - Palliative care in home care: a model for practice. PMID- 15270325 TI - Collaborative pharmacy practice enters hospice care. PMID- 15270326 TI - Health care for seniors: are our nation's physicians ready? PMID- 15270327 TI - Chess and gardening: the Rx for Alzheimer's? PMID- 15270328 TI - Know your clients, know yourself. PMID- 15270329 TI - Home care marketing best practices. PMID- 15270330 TI - New telemedicine solution targets diabetic retinopathy. PMID- 15270331 TI - HHS experiments with "cash & counseling". PMID- 15270332 TI - The healing power of hope. PMID- 15270333 TI - Medicare/prescription drug benefit in the crossfire. PMID- 15270334 TI - Lung cancer, chronic disease epidemiology, and medicine, 1948-1964. AB - Beginning in the early 1950s, a series of epidemiological, biochemical, pathological, and animal studies demonstrated a link between cigarette smoking and lung cancer. A number of reputable scientists challenged these findings, but for a variety of reasons, including the behavior of the tobacco industry, historians have assumed that these objections were insubstantial and disingenuous. Viewing these objections in scientific and medical perspective, however, suggests that there was a legitimate and reasonable scientific controversy over cigarette smoking and lung cancer in the 1950s and early 1960s. That controversy had important consequences. A new chronic disease epidemiology emerged, transforming the role and importance of epidemiology to medical research. This new epidemiology supplemented Koch's postulates, establishing a statistical method that allowed for linking environmental factors to the etiology of chronic diseases. The 1964 report to the surgeon general, Smoking and Health, represented the denouement and codification of these developments. This reexamination of the scientific controversy over smoking in the 1950s and early 1960s provides an important context for understanding the subsequent public relations battles between the tobacco industry and public health after 1964. PMID- 15270335 TI - A history of anatomy theaters in sixteenth-century Padua. AB - The history of anatomy includes not only professors and the support of their institutions but also medical students. Because medical students were quick to assess a teacher's pedagogy, their complaints tell us a great deal about the transition from Galenic to Aristotelian projects of anatomy. When Fabricius of Aquapendente instituted a new style of anatomical inquiry, one based on Aristotle and the search for universal principles, students repeatedly complained that his demonstrations did not provide technical education in structural anatomy (as demonstrations employing a hands-on, Galenic pedagogy did). Within the new anatomy theater (the second of its kind in Padua), however, students were persuaded to accept Fabricius's demonstrations. Fabricius's philosophical orientation combined with the formal atmosphere and aesthetic features of the new theater to create anatomy demonstrations that relied on orations and music for their structure (rather than on the progressive stages of human dissection). A place that emphasized a discourse of anatomy as the study of the "secrets of nature," the new theater so effectively publicized a new style of anatomy that a larger, more diverse group of spectators attended subsequent demonstrations and participated in the celebration of leading academic figures as well as the institution of the university. PMID- 15270336 TI - The reins of the soul: the centrality of the intercostal nerves to the neurology of Thomas Willis and to Samuel Parker's theology. AB - Thomas Willis's description of the intercostal nerves has not received much attention by historians of medicine. Yet the intercostal nerves are of paramount importance for his neurology. Willis explained that via these nerves, which connect the brain to the heart and lower viscera, the brain controls the passions and instincts of the lower body. In other words, Willis believed that the intercostal nerves mediate a kind of rationality and that therefore they make a human a rational being. Willis's theory, I argue, must be seen in the context of the early modern mind-body problem. In the second part of the article I discuss how Oxford theologian Samuel Parker took up Willis's argument while stating that the intercostal nerves are the most important instruments (reins) of the soul. They control the bodily passions so that humans can transform into more virtuous beings. The explanation of the intercostal nerves offered by Willis and Parker fits the Anglican optimism about the abilities of human reason as well as about the moral potential of humankind. PMID- 15270337 TI - Streptomycin, Schatz v. Waksman, and the balance of credit for discovery. AB - A recent article in Nature, arguing that "the misallocation of credit is endemic in science," used Selman Waksman as an illustration, claiming that the true discoverer of streptomycin was one of his graduate students. The article received wide publicity and seriously damaged Waksman's great reputation. What actually happened was that the success of penicillin stimulated Merck to fund research by Waksman, a soil scientist, into the collection of actinomycetes that he had assembled over thirty years. He applied the systematic, uncreative testing techniques that had made the German pharmaceutical industry so successful to these, and streptomycin was discovered within a matter of months. Work in the Mayo Institute then showed that it was marvelously effective against tuberculosis, and Waksman received the Nobel Prize for it in 1952. The test that turned out to be the crucial one could have been carried out by any of several students, but the lucky one was Albert Schatz. He then sued the university for a share of the royalties payable by Merck and also petitioned the Nobel committee to include him in the award. Although he obtained a very substantial out-of-court settlement, this probably damaged his subsequent academic career, and he has never ceased to argue his case for recognition, of which the Nature article is a reflection. To claim that Waksman took credit properly due to Schatz is to fail to understand that once pharmaceutical research had become primarily a matter of large-scale, routine testing, little individual creativity was left in this work. Credit for any successful results must therefore be given to whoever is the originator or director of a particular program. Nature refused to publish evidence that this case could not be used as an example of misallocation of credit for discovery. This in itself illustrates that editors of scientific journals should be every bit as mindful of scientists' reputations as they are of scientific facts. PMID- 15270338 TI - Plague in San Francisco: an essay review. PMID- 15270339 TI - Physico-chemical characteristics of river Ami in relation to discharge of paper mill effluent. AB - The indiscriminate discharge of large volumes of highly putrescible liquid waste from the Sanjai Paper & Chemical Industries Ltd. created serious pollution problem in river Ami near Maghar town area in the district Sant Kabir Nagar of U.P., India. The river is originated from Sikahra Tal near Hallaur (Tehsil - Dumariaganj) of Siddharthnagar district U.P. and disappears into Rapti river near Kouriram of Gorakhpur district U.P. In present investigation an attempt has been made to ascertain the present water quality condition of river Ami in relation to paper mill effluent discharge. The samples were collected from the upstream and downstream of the flow-path of the river from point source of pollution by the mill. The high degree of water quality degradation is reflected by the changes in values of BOD, COD, DO, nitrogen contents and chlorides etc in downstream. PMID- 15270340 TI - Sewage treatment by anaerobic hybrid reactor. AB - Attempts were made to utilize anaerobic hybrid reactor for sewage treatment. The reactor was seeded with digested sewage sludge and a HRT of 24 hrs. was kept in the start up. The HRT was subsequently decreased to 20, 16, 12, 8 and 4 hrs. A maximum COD removal efficiency of 74% was achieved at minimum HRT. The pH, alkalinity, solids and VFA of the effluent were within the permissible limits. PMID- 15270341 TI - Comparative studies of global migration levels of recycled vs virgin polyethylene for packing and storage of milk. AB - The use of Low-density polyethylene (LDPE) is increased for packaging and storage of food, milk etc. Virgin polyethylene is safe due to non-toxic, inertness and insoluble in milk. It is a replacement of conventional bottle packing of milk. Detection of contamination of milk contact of LDPE film in which milk was packed is being done by global migration method. LDPE films developed by well known technique i.e. Blown film extrusion technique, containing different ratio of virgin-recycled polyethylene materials as well as additives such antioxidants, filler, colourant etc. The impact of above LDPE films on the quality of milk was studied by global migration testing. After analyzing the observed results, it is found that the values of global migration increase with increase in percentage of recycled granules and percentage and type of additives in the film. PMID- 15270342 TI - Heavy metals contamination in surface and groundwater supply of an urban city. AB - There is a continuous increase in the demand of water supply in cities due to the industrialization and growing population. This extra supply is generally met by groundwaters or nearby available surface waters. It may lead into incomplete treatment and substandard supply of drinking water. To ensure that the intake water derived from surface and groundwater is clear, palatable, neither corrosive nor scale forming, free from undesirable taste, odor and acceptable from aesthetic and health point of view, the final water quality at Delhi have been evaluated. The final water supply of four treatment plants and 80 tubewells at Delhi were surveyed in 2000-2001 for cadmium, chromium, copper, iron, lead, manganese, nickel, selenium and zinc. The levels of manganese, copper, selenium and cadmium were found marginally above the Indian Standards (IS) specification regulated for drinking water. The data was used to assess the final water quality supplied at Delhi. PMID- 15270343 TI - Treatment of textile dyeing wastewater using UV/solar photofenton oxidation processes. AB - Colour removal of effluent from textile dyeing and finishing industry is becoming important because of aesthetic as well as environmental concerns. Conventional treatment methods have several limitations. Hence emerging technologies like advanced oxidation processes which were based on generation of hydroxyl free radicals (OH) were investigated. In the present work, photofenton oxidation process was used to treat textile dyeing wastewater and the study was carried out at different Fenton molar ratio's (H2O2/Fe2+) like 25:1, 50:1, 75:1, 100:1. It was found that maximum decolourisation occurred at a fenton molar ration of 50:1 and pH 3. A maximum colour removal of 97% was achieved after a contact time of 30 minutes and 70% COD reduction was observed after a contact time of 60 minutes in UV photofenton oxidation process. Whereas 80% colour removal and 50-55% COD reduction was observed after a contact time of 2 hrs in solar photofenton oxidation process. PMID- 15270344 TI - Design of Upelow Anaerobic Sludge Blanket reactor for treatment of organic wastewaters. AB - The Upflow Anaerobic Sludge Blanket (UASB) Reactor is widely applied anaerobic wastewater treatment method all over the world. Uniform distribution of wastewater at reactor bottom is necessary to establish proper contact between sludge and wastewater. In addition, proper functioning of Gas-Liquid-Solid (GLS) separator is crucial to ensure maximum sludge retention in the reactor and to achieve maximum COD removal rate in the reactor. Hence, proper design of reactor is necessary for appropriate functioning of various components for a given wastewater flow rate and COD concentration. The design procedure for UASB reactor taking due consideration to the GLS design and design of inlet arrangement is discussed in this paper for various wastewater strength and flow rates. A software is developed to make economical design of UASB reactor for different type of wastewater by adopting maximum loading conditions, based on literature recommendations, and at the same time to satisfy all design recommendation, as far as possible. PMID- 15270345 TI - Acute toxicity of lake water to fishes. AB - The present study investigates the acute toxicity of Hussainsagar lake water to fishes. In this context, experiments were conducted on three species of fishes (Cyprinus carpio, Tilapia mozambica and Lebistus reticularis), using 5, 10 and 20% volume of Jeedimetla nallah wastewater and were observed for the signs of survival within the specified period of time. The TLm value for 96 hours test period was found to be 17%. Similarly Toxicity studies were carried out on Hussainsagar lake water with dilutions varying from 10-100%. Absence of mortality during 96 hours of test period indicates that there is no acute toxicity of Hussainsagar lake water to fishes. PMID- 15270346 TI - Investigation of GC detectors performance and validation. AB - Detectors in gas chromatographs are used in many different ways. The functions varies with the requirement of sensing the chemicals at trace levels to measuring quantitatively the relative amounts of compounds in ppb to sub ppb levels. The requirement of minimum detectable sensitivity for GC detectors and its actual estimation during the analysis helps in assessing the performance of detectors and a stable response of the GC system. The paper reports the study carried out with a GC system equipped with detectors like Flame Ionization Detector(FID) and Electron Capture Detector(ECD). Both packed and capillary column have been used for estimating the minimum detectability of detectors. The FID sensitivity obtained is 1.14 coulombs/g C. The FID could minimum detect 2.65 x 10(-12) gram carbon per sec and ECD could minimum detect pesticides (Lindane) less than 1 femtogram. Such a procedure is useful for all those engaged in analysis with GC or are in the process of procuring good reliable GC system. PMID- 15270347 TI - Arsenic reduction from aqueous environment by water lettuce (Pistia stratiotes L.). AB - Arsenate uptake by aquatic plant water lettuce (Pistia stratiotes L.) was studied in the laboratory condition to investigate a low cost natural aquatic treatment system for pollutant removal. The plants were harvested from a local pollution free pond in young condition and hydroponically cultured in the laboratory. Bioaccumulation was noticed to be both concentration and duration dependent. The results show that the plant could effectively abosrb arsenic between a range of 0.25 to 5.0 mg/l to the extent of 82.0 to 22.8% for a biomass of 20g/l at pH 7.0 after 144 hours. The results were also plotted in Langmuir and Freundlich isotherms and the data were well fitted. The sorption capacity was evaluated as 1.43 mg/g for Langmuir isotherm and 1.01 mg/g for Freundlich isotherm. The removal efficiency was, however, noted to be maximum (87.5%) at pH 6.5. The effect of biomass quantity has also been investigated along with some metabolic parameters. PMID- 15270348 TI - Equilibrium and kinetics studies on removal of arsenite by iron oxide coated activated alumina. AB - Arsenic in drinking water is causing much concern because of its toxicity. It occurs in water naturally as As(III) and As(V). Of these As (III) is more toxic. Adsorption on activated alumina has been one of the most commonly used methods for As (V) removal from drinking water. But it is not very effective for As (III). Activated alumina was modified by coating it with Iron oxide to make it suitable for As (III) adsorption. Iron oxide coated activated alumina was tested for its effectiveness as an adsorbent for As (III). The As (III) adsorption was strongly dependent on pH and a maximum removal of 98% was observed at a pH of 12. The adsorption process followed a first order kinetics. The equilibrium was attained after 8 hours. The kinetic study was carried out with different initial As (III) concentration. It was observed that time taken to attain equilibrium was independent of initial concentration but percentage removal decreased with increasing initial concentration. The adsorption isotherms were fitted well to both Langmuir and Freundlich equation. PMID- 15270349 TI - Nitrate and fluoride levels in ground waters of Davanagere Taluka in Karnataka. AB - The quality of ground water supplies in Davanagere Taluk, situated in central part of Karnataka has been investigated with respect to pH, dissolved solids, chlorides, nitrates and fluorides. The levels of pH, dissolved solids and chlorides were found within the safe limits as prescribed by BIS, for more than 95% of the samples. Out of the 61 different borewell samples analysed, selected from different areas of Davanagere taluk, 26% of the samples are found to contain fluorides less than 0.50 PPM (lower safe limit prescribed by BIS) and 11.5% of the samples are found to contain more than 1.5 PPM of fluorides (higher safe limit prescribed by BIS). Further, it was also found during study that, 16.00% of the borewell samples analyzed, were found to contain more than 100.00 PPM of nitrates (measured as NO3 mg/L, safe limit prescribed by BIS). The values of fluorides and nitrates observed in different samples were in the range of 0.19 - 2.06 PPM and 0.08 - 308 PPM, respectively. PMID- 15270350 TI - Disappearence of some toxic elements in the surface water of Hussainsagar Lake following Ganesh-idol immersion. PMID- 15270351 TI - Change ideas. Dare to be different. PMID- 15270353 TI - The HSJ interview: John Reid. No option but choice. Interview by Alastair McLellan. PMID- 15270354 TI - The HSJ barometer. PMID- 15270355 TI - Clinical management. Where medicine meets management. Muscle power. AB - Physiotherapy-led services can dramatically cut orthopaedic waiting times, improve patient choice and reduce GP workload. Waiting times are cut by reducing the number of unnecessary referrals. Providing more direct access to physiotherapy services is key. PMID- 15270356 TI - Primary care. The odd couple. AB - Acute and primary care trust relationships can be fraught with baggage, vagueness and stereotypes. Changing that is about overt behaviours as well as structural and strategic issues. Payment by results and attitudes to commissioning pose significant challenges. PMID- 15270357 TI - Communications. Zap it to 'em. AB - Communication is something all trusts do, whether they know it or not. Reputation management involves proactively controlling how your organisation is, and will be, perceived. Patient choice and a move away from a waiting-list focus is changing the messages trusts need to get across. PMID- 15270358 TI - International development. Something to write home about. PMID- 15270359 TI - Finance. Defender of the faith. PMID- 15270360 TI - HSJ people. What HR people want. PMID- 15270361 TI - Law. Judge ye not. PMID- 15270362 TI - Law. Parental guidance. PMID- 15270363 TI - Law. Fraternity leave. PMID- 15270364 TI - Law. Storm troopers. PMID- 15270365 TI - Influence of yoga on brain and behaviour: facts and speculations. PMID- 15270366 TI - Endocrine and paracrine correlates of endometrial receptivity to blastocyst implantation in the human. AB - Synchronous attainment of maternal endometrial receptivity allows implantation stage adhesive blastocyst to undertake apposition, attachment and invasion. In the present essay, we propose a model according to which luteal phase progesterone induces a basic drive in endometrium toward receptivity and as a result, adequately primed endometrium differentiates through certain steps in a fixed action pattern. The implantation-stage embryo senses such endometrial responsiveness circumstantially by the factors secreted by maternal endometrium and undertakes differentiation to implant by secreting factors which act on maternal endometrial cells to further potentiate them to implantation stage specific changes. Such a dynamic temporo-spatial manner of interaction involving a set of specific factors acting synchronously leads to the activation of innate releasing process in both compartments towards embryo attachment followed by successful intrusion and controlled invasion of trophoblast cells into maternal endometrium. In the present review we discuss the potential role of various endocrine and paracrine factors in the process of blastocyst implantation in the human. PMID- 15270367 TI - Heart rate and blood pressure responses of left-handers and right-handers to autonomic stressors. AB - We hypothesized that cerebral dominance may contribute to differences in cardio vascular responses of right-handers (RH) and left-handers (LH) to autonomic stressors. We tested this hypothesis by exposing 14 RH, and 14 LH males to category I tests in which the hand and cerebral cortex were involved in performing the test viz.--i) Cold pressor test (CPT), ii) Handgrip dynamometry (HGD) and; category II (no use of hand)--i) Orthostatic Tolerance Test (OTT), ii) Valsalva Manuever (VM), iii) Controlled Breathing Test for sinus arrhythmia (SA) in a random sequence, and measured their heart rate (HR/min) and blood pressure (MAP mmHg). All subjects had similar resting HR and MAP values, and responded to the category I interventions with increased HR and BP. The absolute HR values of LH and RH did not differ significantly during the interventions. However, the increase in HR from control induced by the CPT, and the HGD was greater for LH (P<0.05). Also, LH showed a greater decrease in HR and MAP in the recovery phase (P<0.05). The VAS scores for degree of discomfort during the CPT were similar for both the groups. During the OTT, the increase in HR was more in RH (P<0.05). The Valsalva ratios for LH and RH were similar. Our findings suggest that the autonomic control over the cardio-vascular system may be different in LH and RH, and that this imbalance could be attributable to a variation in cerebral dominance. PMID- 15270368 TI - Effect of co-administration of cassava (Manihot esculenta Crantz) rich diet and alcohol in rats. AB - The effects of co-administration of a cassava rich diet and alcohol in rats were investigated. The animals were divided into four groups (1) Control, (2) Alcohol, (3) Cassava and (4) Alcohol + Cassava. Consumption of alcohol along with cassava reduced the alcohol induced toxicity which was evidenced by the lower activities of GOT, GPT, GGT, acid phosphatase and alkaline phosphatase in the liver and serum of co-administered group. The pyruvate content in the blood increased while the lactate content, lactate/pyruvate ratio and the activity of LDH decreased in the blood due to co-administration. The blood cyanide content, serum thiocyanate content and the activities of rhodanase and beta-glucuronidase increased on co administration. The histopathological studies also revealed that co administration reduced the alcohol induced toxicity. PMID- 15270369 TI - Adrenergic involvement in the locus ceruleus and adjoining regions in the facilitation of predatory attack behavior as induced by hypothalamic stimulation in cats. AB - The present study was carried out in five cats which did not attack the rats spontaneously. Predatory attack on an anaesthetized rat was elicited by electrical stimulation of extreme lateral regions of hypothalamus. These sites were stimulated at a current strength from 300-700 microa to evoke a predatory attack on an anaesthetized rat. The attack was accompanied by minimal affective display such as alertness, pupillary dilatation, and culminated in beck biting at higher current strength. A scoring system allowed the construction of stimulus response curves, which remained fairly constant when repeated over a period of 3 4 weeks. Microinfusions of norepineprine and clonidine in 4.0 and 5.0 microg dose respectively in locus ceruleus and adjoining tegmental fields facilitated the predatory attack and there was a significant reduction in the threshold current strength for the elicitation of affective and somatomotor components. Microinfusions of yohimbine, an alpha-2 blocker, in 5 microg dose completely blocked the predatory attach response as indicated by an increase in the threshold current strength for the affective components. The somatomotor components were completely inhibited and could not be elicited even when the current strength was increased to 1000 microA. The predatory attack behavior remained completely inhibited for almost two hours following microinfusion of yohimbine. During this period, the animal was extremely drowsy and reacted very slowly even to a painful stimulus such as pinching of tail. Microinfusions of propranalol (beta-blocker), practalol (beta-1 blocker), prazosin (alpha-1 antagonist), propylene glycol as well as saline in similar volumes (0.5 microl) as control failed to produce any blocking effect, thus indicating the involvement of alpha-2 adrenoceptive mechanisms in the modulation of predatory attack in this region of midbrain. The facilitatory effects of norepinephrine and clonidine were significant at P<0.01 and P<0.05 respectively with Wilcoxon's signed rank test. The inhibitory effects of yohimbine were significant at P<0.05. The present study indicates the involvement of alpha-2 adrenoceptive mechanisms in the facilitation of hypothalamically elicited predatory attack. PMID- 15270370 TI - Modulation of stress induced by isometric handgrip test in hypertensive patients following yogic relaxation training. AB - 13 essential hypertensive patients aged 41 to 60 years were given yoga training for 60 min daily, Monday through Saturday, for a total duration of 4 weeks. Blood pressure and heart rate (HR) were measured with non-invasive semi-automatic blood pressure monitor. Measurements were recorded before the training and at weekly intervals during the 4 week training period. Results of our study show a significant (P<0.001) reduction in resting HR and rate-pressure-product (RPP) after 2 weeks of yoga training. Systolic pressure (SP), diastolic pressure (DP) (P<0.001) and mean pressure (MP) (P<0.05) showed a significant reduction at 3 weeks of training period. After 4 weeks of training, there was further fall in SP, DP, pulse pressure (PP) (P<0.05), MP (P<0.001), HR and RPP. Isometric handgrip test before yoga training produced a significant rise in SP and MP and insignificant rise in DP, HR and RPP. After yoga training, there was a significant rise in all these parameters. Our results show that yoga training optimises the sympathetic response to stressful stimuli like isometric handgrip test and restores the autonomic regulatory reflex mechanisms in hypertensive patients. PMID- 15270371 TI - Effect of occupational noise on the nocturnal sleep architecture of healthy subjects. AB - BACKGROUND & OBJECTIVES: Noise is considered to be a non-specific stressor which generally causes physiological and psychological effects in an individual. Many occupations involve workers being subjected to loud noise levels without adequate protective measures. The study was done to document the changes, if any, in the nocturnal sleep architecture of healthy persons exposed to loud occupational noise during daytime. METHODS: The study was a retrospective cohort design wherein three groups of eight subjects each, exposed to continuous occupational background noise levels of >75dB for 1-2 years, 5-10 years and >15 years were selected. Corresponding age and gender matched healthy controls (eight for each group) who worked in a quiet atmosphere was also recruited. All night sleep polysomnography was done on all subjects. In the morning, subjects rated their quality of sleep on a Visual Analogue Scale. RESULTS: There is a strong association between occupational exposure to loud noise and poor sleep efficiency (Relative Risk 2.49; Confidence Interval 1.12 to 5.57; P=0.01, Fisher's exact test). The group exposed to noise for 1-2 years had a decrease in Total Rapid Eye Movement Time, Non Rapid Eye Movement Time, Slow Wave Sleep Time, Sleep Onset Latency and Total Sleep Time. The other two groups showed lesser number of changes in sleep architecture. Subjectively there was a decrease for sleep continuity in Group I and an increase for sleep onset in Group II. There is no correlation between loudness of noise in the workplace and sleep efficiency. INTERPRETATION & CONCLUSION: It can be concluded that workers exposed to loud background occupational noise are at an increased risk of having poor quality sleep but adaptation to this effect probably takes place after a few years. PMID- 15270372 TI - Receiver operating characteristic curve analysis of BMI and percentage body fat in type 2 diabetics of Punjab. AB - The present study attempted to establish appropriate cut off levels of Body Mass Index (BMI) for defining overweight as a risk for the development of type 2 diabetes considering percentage body fat (BF) as standard. A total of 300 patients of known type 2 diabetes participated in the study (150 males and 150 females, all > or = 40 years of age). Clinical examination was done. Anthropometric measurements as BMI, Waist Circumference (WC) and Waist-hip ratio (WHR) were calculated. Percentage BF was calculated using skinfold thickness method from the equation of Durnin and Womersley. Mean BMI for males was 24.97 (SD 4.3) kg/m2 and for females was 27.56 (SD 5.14) kg/m2. Mean percentage BF for males was 28.19 (SD 0.74) and for females was 38.22 (SD 5.29). A comparison of BF and BMI data with various ethnic groups revealed conspicuous differences. Receiver operating characteristic (ROC) curve analysis showed a low sensitivity of conventional cut off value of BMI (25 kg/m2) in identifying subjects with overweight as compared to the cut off values based on percentage BF (males > 25, females > 30). This results in substantial misclassification. Based on the ROC curve, a lower cut off value of BMI 22.3 kg/m2, displayed the optimal sensitivity and specificity, and less misclassification in identification of type 2 diabetics with high percentage BF. BF: BMI was calculated and was found to be higher in females. PMID- 15270373 TI - Antidiabetic activity of Aegle marmelos and its relationship with its antioxidant properties. AB - Oxidative stress induced by alloxan has been shown to damage pancreatic beta-cell and produce hyperglycemia in rats. Aegle marmelos leaf extract is being used in Ayurveda as a medicine for diabetes. The present study examined the action of Aegle marmelos against experimental diabetes as well as the antioxidant potential of the drug. A methanolic extract of Aegle marmelos was found to reduce blood sugar in alloxan diabetic rats. Reduction in blood sugar could be seen from 6th day after continuous administration of the extract and on 12th day sugar levels were found to be reduced by 54%. Oxidative stress produced by alloxan was found to be significantly lowered by the administration of Aegle marmelos extract. This was evident from a significant decrease in lipid peroxidation, conjugated diene and hydroperoxide levels in serum as well as in liver induced by alloxan. Catalase and glutathione peroxidase activity in blood and liver were found to be increased from 9th day onwards after drug administration. Superoxide dismutase and glutathione levels were found to be increased only on 12th day. These results indicate that Aegle marmelos extract effectively reduced the oxidative stress induced by alloxan and produced a reduction in blood sugar. PMID- 15270374 TI - Evaluation of zinc against salinomycin toxicity in broilers. AB - Salinomycin was studied for its toxicity and zinc (80 mg/kg) was assessed for prophylactic and therapeutic management in broiler chicks. Male broiler chicks were randomly divided into 7 groups consisting of 6 chicks in each. Group 1, 2 and 3 were maintained as control, therapeutic dose control (60 mg/kg feed) and toxic dose control (120 mg/kg feed), respectively. Group 4 was fed on feed containing salinomycin therapeutic dose and zinc. Group 5 received feed containing toxic dose of salinomycin. Group 6 and 7 were fed on feed containing toxic dose of salinomycin for the first 4 weeks for induction of ionophore toxicity and for the subsequent 2 weeks, group 6 received zinc and group 7 was fed on feed containing toxic dose of salinomycin along with zinc. Weekly body weights revealed a significant (P<0.01) decrease in toxic controls as compared to group 1, 2, 4 and 5. The activity of glutathione peroxidase, glutathione reductase and catalase, and the values of aspartate aminotransferase (AST), alanine aminotransferase (ALT), total proteins, total cholesterol, triglycerides, low density lipoproteins (LDL), urea, creatinine and blood urea nitrogen (BUN) were significantly (P<0.01) elevated in toxic controls, whereas glutathione (GSH) and high density lipoproteins (HDL) were significantly (P<0.01) lowered as compared to group 1, 2, 4 and 5. Following toxicity, zinc supplementation in group 6 and 7, all serobiochemical parameters were revived to normal. Thus, it is enunciated that salinomycin toxicity is due to oxidative damage and use of zinc in feed tends to cure and avoid any accidental toxicity. PMID- 15270375 TI - Effect of protein malnutrition on the intestinal absorption of monosaccharides in rats in vivo. AB - The present study was planned to elucidate the role of protein malnutrition on the intestinal absorption of monosaccharides particularly--glucose and xylose, in inbred female albino rats. The experimental rats were fed with protein deficient diet containing 3% protein, whereas the control rats were given a diet containing 18% protein. The study on intestinal absorption of monosaccharides was conducted on both the groups of rats after the 7th and 15th day of receiving respective diets. The results indicated no significant impairment of glucose absorption of experimental rats fed 3% diet for 7 days as compared to the controls. However a 42% decrease in glucose absorption was observed when the animals were fed with the same diet for 15 days. The impairment was significant in all segments of intestine suggesting diminution in the absorption capacity of small intestine in malnutrition perhaps as a result of some permanent injury to mucosal cells of small intestine. Regarding xylose absorption, in experimental rats an increase of intestinal uptake was noticed in most of the segments of small intestine as compared to control rats. PMID- 15270376 TI - Self-reported drug use and urinalysis results. AB - The study examined the consistency between retrospective self-reported drug use and urinalysis data among 281 male opioid dependent subjects attending out patient clinic of National Drug Dependence Treatment Centre from January 2001 to December 2001 at All India Institute of Medical Sciences, New Delhi. Preliminary analysis indicated that there was moderate to high concordance between the two measures among different drug types. On an average 85% of urine test results matched with self-report. Subject's over-reported drug use as indicated by the low positive predictive value. In contrast, subjects were more accurate when they were reporting no drug use as suggested by the high negative predictive value. The study suggests that urine analysis is a critical variable in substance abuse treatment programs. Clinicians should be cautious while prescribing agonist drug due to frequent over-reporting of drug use by patients in our setting. This will make the substance abuse program more meaningful. PMID- 15270377 TI - Effect of age and nutritional status on heart rate responses to cough and maximum handgrip. AB - Autonomic nerve activity can be assessed using simple bed side tests such as cough and maximum hand grip (MHG). The alterations in these tests are, however, poorly documented in physiological states. The present study aimed to uncover the effect of nutritional status and age on these tests. 93 male adults were divided into normal body mass index (BMI) (BMI; 18.5 to 25 kg/m2; young 18-30 yrs, n=28; old >60 yrs, n=25) and low BMI (BMI; <18.5 kg/m2; young 18-30 yrs, n=19; old >60 yrs, n=14) groups. The younger subjects showed a significantly higher heart rate response to cough and MHG in both normal and low BMI groups as compared to the older subjects (P<0.01). However, there were no significant differences for the heart rate responses to cough and MHG between the low and normal BMI groups either in the young or in the elderly. The data suggest that while the heart rate response to cough and MHG are useful tests of vagal activity to the heart when expected differences are large, they may be of limited use in uncovering more subtle changes. PMID- 15270378 TI - Effect of nifedipine and amlodipine on wound healing in rats. AB - The wound healing effect of two calcium channel blockers, nifedipine and amlodipine was studied in rats using incision and excision wound models. In incision wound, two straight paravertebral skin thickness incision were made and on tenth day skin tensile strength was measured by using continuous water flow technique. In excision wound, circular piece of skin excised in its full thickness and wound contraction monitored by alternate day wound tracing and epithelisation period was monitored by noting the number of days required for escher to fall. Drugs enhanced the skin tensile strength in incision wound model. In excision wound model, wound contraction is increased on 4th and 16th day but epithlisation period was not significantly altered. In conclusion, calcium channel blockers can be used to enhance wound healing, especially if wound healing was suppressed by steroids. PMID- 15270379 TI - Oxidative stress and antioxidants status in peptic ulcer and gastric carcinoma. AB - Oxidative stress is believed to initiate and aggravate many diseases including peptic ulcers and gastric carcinoma. We observed an increase in rat gastric mucosal lipid peroxidation (LPO) and superoxide dismutase (SOD) and a decrease in catalase (CAT) levels in cold restraint stress-induced gastric ulceration while, in clinical peptic ulceration and gastric carcinoma patients, an increase in serum LPO and a tendency to decrease in SOD and CAT levels were observed. The result thus, indicated a positive correlation between free radical-induced oxidative stress both in gastric and duodenal ulcers and gastric carcinoma. PMID- 15270380 TI - Effect of the antioxidants alpha-tocopherol acetate and sodium selenite on hepatotoxicity induced by antitubercular drugs in rats. PMID- 15270381 TI - Nobel Prize in Physiology or Medicine--2003. PMID- 15270382 TI - The power of persistence in medicine. PMID- 15270383 TI - Access to health care: a pressing issue for this country. PMID- 15270384 TI - The search for meaning in a medical life. PMID- 15270385 TI - An offer they can't refuse. As mergers and acquisitions make a comeback, hospitals large and small are choosing consolidation over competition. AB - Hospital deals are heating up as summer settles in, fueled in part by Medicare reform and the "Tenet effect." One recent move by Christus Health made Alice, Texas, a one-hospital town again. Christus' Peter Maddox, left, denies that the consolidation means prices will rise. "It's not a retail world anymore," he says. PMID- 15270386 TI - Bedside battle. Study prompts renewed call for ban on forced OT. PMID- 15270387 TI - Adopting automation. IT leaders try to establish voluntary certification. PMID- 15270388 TI - Pluses and minuses. Edwards' background as trial lawyer raises concerns. PMID- 15270389 TI - Not satisfied. Democrats call for independent probe of Scully. PMID- 15270390 TI - Scruggs' hospital lawsuit grows. PMID- 15270391 TI - Medicare mayhem. Fur flies over wage index policies, use of new MSAs. PMID- 15270392 TI - Raining on competition. Florida's specialty hospital ban is a bad idea for patients. PMID- 15270393 TI - The process comes first. Before assembling all the pieces of a clinical IT system, know what the initiative needs to accomplish and how the technology fits in. PMID- 15270394 TI - It's more than just the purchase. Make clear the commitments that it will trigger. PMID- 15270395 TI - Large issues for smaller hospitals. Proper planning leads to IT system with the right fit. PMID- 15270396 TI - A new game of leapfrog? RFID is rapidly changing the product-tracking process. Some say the technology--once costs drop--could displace bar-coding. PMID- 15270397 TI - Turbulence ahead. HFMA speakers confident despite myriad challenges. PMID- 15270398 TI - [The book of Isaac Israeli's fever and his importance in the Latin Western World]. PMID- 15270399 TI - Male pseudohermaphroditism: long-term quality of life outcome in five 46,XY individuals reared female. AB - We assessed the adult quality of life of five medical chart-selected genetic males (ages 29-34 years) assigned and reared as females due to ambiguity of the external genitalia. All five were treated following the traditional method proposed by John Money and colleagues in 1955, commonly referred to as the "optimal gender policy". The adult follow-up assessment included physical and endocrinological evaluation, completion of self-report questionnaires, and a semi structured interview assessing gender identity, sexual experience and orientation. Quality of life domains assessed by questionnaire included health related issues, satisfaction with health-care management, emotional distress, and relationship satisfaction. Vaginoplasty in four out of five patients was initially unsuccessful. Four patients had periodic lapses in adherence to hormone replacement therapy. Gender role behavior across development was masculine relative to norms for women. All participants reported a female gender identity without a history of gender dysphoria. The majority of participants (four of five) reported being sexually active and in long-term relationships (three heterosexual, one homosexual). Current emotional adaptation and health-related quality of life are within the normal range for four participants. Sex assignment of 46,XY individuals with ambiguous genitalia as females is compatible with a positive quality of life. PMID- 15270400 TI - Genital ambiguity with a Y chromosome: does gender assignment matter? AB - Recommendations for sex of rearing in newborns with genital ambiguity, testicular differentiation and a Y chromosome continue to be challenging. Complaints from former patients have forced those providing the medical, surgical and psychological care for these individuals to reassess evaluation and treatment strategies. In this paper, the histories of six patients born with genital ambiguity and at least partial testicular differentiation with a karyotype containing a Y chromosome are presented. Three of these patients were assigned as males and three as females. The factors involved in these individuals' adaptation to the assigned gender and their subsequent quality of life are discussed. Factors needing further study, including the parents' ability to accept and support the sex of rearing, the child's temperament, associated psychological disorders, and other influences, such as masculinization of the central nervous system, are highlighted. PMID- 15270401 TI - Imaging in intersex disorders. AB - BACKGROUND: Evaluation of the gonads and internal genital structures is an essential component for evaluation of patients presenting with ambiguous genitalia. Ultrasonography (US) and magnetic resonance imaging (MRI) are the two preferred modalities. OBJECTIVE: To compare US and MRI in patients with intersex for localization of gonads and internal genitalia. PATIENTS AND METHODS: Ten patients with proven intersex disorders were included in the study. Findings from US and MRI were corroborated by those from surgery/laparoscopy. RESULTS: For evaluation of the gonads, MRI was found to be marginally more sensitive than US. For internal genital structures, both modalities were found to be equally sensitive and specific with no false positive results. CONCLUSION: US still remains the modality of choice for screening patients with intersex disorders. MRI is helpful in cases with equivocal US findings. PMID- 15270402 TI - Gestational diabetes mellitus--implications of different treatment protocols. AB - OBJECTIVE: To assess the impact of different management approaches to gestational diabetes mellitus (GDM) on perinatal outcome. PATIENTS AND METHODS: The study group consisted of 2,060 patients with GDM treated in our center from January 1980 through December 1999. Four time periods were defined on the basis of changes in treatment protocols. Perinatal complications were compared between the periods and with normal pregnancy controls. RESULTS: The last two periods (1993 1999) were characterized by lower mean glucose level, lower mean gestational age at delivery, and a decline in macrosomia, shoulder dystocia and perinatal mortality rates, but also by high rates of labor induction and Cesarean delivery. A significant difference was found between the GDM and normal control groups in rates of labor induction (38.6% vs 10.8%, p < 0.001) and Cesarean delivery (34% vs 20%, p < 0.001) for the last period. CONCLUSIONS: Perinatal complications are preventable with good glycemic control and early induction of labor, but at a cost of a higher Cesarean section rate. PMID- 15270403 TI - Postprandial hyperlipidemia after a fat loading test in minority adolescents with type 2 diabetes mellitus and obesity. AB - The continuing increase in the incidence of type 2 diabetes mellitus (DM2) and obesity in children and adolescents is attributable to excessive caloric intake. Abnormal lipid metabolism in the postprandial state leads to long exposure of the vasculature to hyperlipidemia. Most children and adolescents with DM2 are obese, and many have fasting hypertriglyceridemia. Clustering of hyperlipidemia, DM2 and obesity increases the risk for cardiovascular disease. We therefore studied lipids, insulin, C-peptide, and glucose in response to an oral fat load simulating the fat content of a high-fat, fast-food meal in 12 type 2 diabetic obese, 15 non-diabetic obese, and 12 non-diabetic non-obese (control) adolescents (aged 10-19 yr; 87% African-Americans). All three groups were age-, sex-, and sexual maturation-matched. Mean body mass indices were similar in the diabetes and obese groups (32.7 +/- 1.1 vs 35.8 +/- 1.6 kg/m2). All patients with DM2 had fasting C-peptide > 0.2 nmol/l (0.7 ng/ml) and negative diabetes-associated autoantibodies. Serum total cholesterol, triglyceride, high- and low-density lipoprotein cholesterol, insulin, C-peptide, and plasma glucose levels were measured at 0, 2, 4, and 6 h after the fat load. The area under the curve (AUC) was calculated by trapezoidal estimation. Triglyceride AUC was significantly greater in the diabetes group than in the other two groups (15.7 +/- 2.9 vs 9.2 +/- 0.7 and 7.5 +/- 0.7 mmol x h/l [1389 +/- 258 vs 819 +/- 60 and 663 +/- 62 mg x h/dl]; p < 0.02 and <0.004, respectively), as were insulin, C-peptide, and glucose AUCs. Incremental triglyceride response (delta triglyceride = peak - fasting) in the diabetes group was significantly higher than that in the control group (2.1 +/- 0.7 vs 0.8 +/- 0.1 mmol/l 189.7 +/- 58.4 vs 71.2 +/- 11.1 mg/dl]; p < 0.04). Insulin resistance was estimated using the homeostasis model assessment (HOMA), which was greater in the diabetes group than in the obese and control groups (14.4 +/- 2.8 vs 5.2 +/- 0.8 and 3.2 +/- 0.4; p < 0.001 and < 0.0001, respectively). The diabetes group was divided into subgroups of high and normal fasting triglycerides on the basis of triglyceride levels above and below the 95th percentile. The delta triglyceride in the subgroup with high fasting triglycerides was substantially greater than in the subgroup with normal fasting triglycerides (3.4 +/- 1.1 vs 0.8 +/- 0.2 mmol/l [305.2 +/- 96.8 vs 74.2 +/- 18.0 mg/dl]; p < 0.001). Total cholesterol and triglyceride AUCs were much greater in the high vs normal fasting triglycerides subgroup (33.0 +/- 2.9 vs 24.2 +/- 1.9 and 23.6 +/- 3.5 vs 7.8 +/- 0.6 mmol x h/l [1274 +/- 113 vs 934 +/- 72 and 2085 +/- 309 vs 692 +/- 49 mg x h/dl]; p < 0.02 and <0.0001, respectively), as were insulin and C-peptide AUCs. HOMA was greater in the high vs normal fasting triglycerides subgroup (20.8 +/- 4.0 vs 8.0 +/- 1.6; p < 0.0001). In addition to elevated plasma glucose levels, there were no significant differences in either insulin or lipid parameters among the diabetes subgroup with normal fasting triglycerides, the obese group, and controls. Our data suggest that postprandial hyperlipidemia in response to a fat loading test is present in adolescents with DM2 who already have fasting hypertriglyceridemia. The degree of insulin resistance as an underlying abnormality--not DM per se--determines the degree of postprandial lipemia. PMID- 15270404 TI - Preschoolers are not miniature adolescents: a comparison of insulin pump doses in two groups of children with type 1 diabetes mellitus. AB - We analyzed glycemic control and insulin usage patterns of 14 preschoolers and 14 adolescents on continuous subcutaneous insulin infusion (CSII) matched for sex to determine how CSII therapy for type 1 diabetes mellitus (DM1) differs in preschoolers as compared to adolescents. Average hemoglobin A(1c) was lower in the adolescents. The percent of insulin delivered as the basal rate was the same in both groups; however, during the hours after midnight the preschoolers needed a much lower basal rate per kg body weight. There were also significant differences in insulin sensitivity and insulin/carbohydrate ratios. The number of basal rates, number of boluses, and percent of insulin administered as the basal rate were not different between groups. This is the first report of discrete differences in insulin usage patterns for preschoolers and adolescents on insulin pumps. Reasons for these dissimilarities include differences in hormone production, insulin absorption, frequency of food intake, and glycemic targets. Recognizing these variations is essential for safe and efficacious use of CSII in preschoolers with DM1. PMID- 15270405 TI - Type 1 and type 2 diabetes mellitus in childhood in the United States: practice patterns by pediatric endocrinologists. AB - AIM: To determine the relative frequency of type 2 diabetes mellitus (DM) in the US, and to assess diabetes practice patterns in the US. METHOD: A questionnaire regarding pediatric diabetes practice patterns was distributed to the members of the Lawson Wilkins Pediatric Endocrine Society in 1999. Only one member of each practice group was requested to respond. Responses received through early 2000 were analyzed. RESULTS: One hundred and twenty-six practices representing 45% of the members of the Society responded. 11.9% of pediatric patients with DM were considered to have type 2 DM. On average 53 new patients with DM were seen each year. The average practice consisted of 2.5 physicians, 1.5 nurse educators, 1.3 dieticians, 1.0 social workers and 0.5 nurse practitioners. Management practices comply by and large with the recommendations of the American Diabetes Association and reflect a trend toward more intensive treatment and monitoring. CONCLUSION: Type 2 DM was seen in 11.9% of patients. Most diabetes practices in the US utilize a team approach to the management of youth with DM. PMID- 15270406 TI - Decreased serum inhibin B/FSH ratio as a marker of Sertoli cell function in male survivors after chemotherapy in childhood and adolescence. AB - OBJECTIVE: Inhibin B produced by Sertoli cells may be an important marker of seminiferous tubule function in patients treated with chemotherapy (CT). The aim of this study was to evaluate the inhibin B/FSH ratio to detect male gonadal dysfunction in cancer survivors treated in childhood and adolescence. PATIENTS: Twenty-one male patients (group A) treated with 6-10 courses of CT for Hodgkin's disease during childhood and adolescence were examined 3-11 years after the conclusion of treatment. Twenty healthy young men (18-23 years old) were used as controls (group B). METHODS: Serum samples for the determination of inhibin B, follicle-stimulating hormone (FSH), luteinizing hormone (LH), testosterone (T), sex hormone-binding globulin (SHBG) and semen for analysis were collected. RESULTS: The median testicular volume of patients of group A was lower than those of group B (p = 0.001) and a positive correlation was found between testicular size and sperm count (r = -0.5, p = 0.01). Semen analysis revealed azoospermia in 11 patients, severe oligospermia in four and normal sperm count in three. No significant difference was found in the median of T, LH, SHBG, inhibin B concentrations and T/LH ratio between the groups. Serum inhibin B was correlated with the serum FSH levels (r = -0.5, p = 0.02). Median FSH was significantly higher (p = 0.0001), and median inhibin B/FSH ratio was significantly lower in group A than in controls (p = 0.0002), but the inhibin B/FSH ratio was higher in the patients with normal sperm count than in those with oligospermia (p = 0.00004). CONCLUSIONS: These results show that the cytotoxic effects of CT cause severe damage to the germinal epithelium with subtle effects on Sertoli cells. To assess Sertoli cell function in men with primary testicular damage after treatment with CT in childhood and adolescence, the inhibin B level needs to be interpreted in the context of the circulating FSH, especially when normal FSH levels are observed. PMID- 15270407 TI - Quantitative computed tomography measurements of bone mineral density in prepubertal children with congenital hypothyroidism treated with L-thyroxine. AB - Low bone density (BD) has been reported in patients with hyperthyroidism. Whether or not levothyroxine (LT4) therapy in children with congenital hypothyroidism (CH) affects BD is unclear. Medical records of 45 patients with various etiologies of CH who had at least one BD measurement (32 female, mean age 7.6 +/- 2.6 years) were reviewed. The mean LT4 dose was 3.6 +/- 0.88 microg/kg/day. Cancellous bone density (CaBD) was measured by quantitative computed tomography (CT) in all 45 patients and 20 had measurements of cortical bone density (CoBD), cross-sectional area (CSA) and cortical bone area (CBA) of the femur. TSH levels were considered partially or completely suppressed when values were <1.0 or <0.5 microIU/ml, respectively. The control group consisted of age- and gender-matched healthy children. No significant differences were found in CaBD, CoBD, CSA, or CBA between patients with CH and controls. There were no significant differences between initial and subsequent BD measurements. No correlations were found between CaBD and etiology of CH, dose or duration of LT4 therapy, or serum TSH. In pre-pubertal children with CH, LT4 appears to have no significant effect on BD. Moreover, absence or hypoplasia of the thyroid parenchyma appears to have no significant impact on bone formation within the first 10 years of life. PMID- 15270408 TI - Alterations in serum growth hormone (GH)/GH dependent ternary complex components (IGF-I, IGFBP-3, ALS, IGF-I/IGFBP-3 molar ratio) and the influence of these alterations on growth pattern in female rhythmic gymnasts. AB - BACKGROUND AND AIM: Normal growth in children is regulated to a great extent through the actions of the GH/IGF-I axis, a system consisting of GH and its mediators (ternary complex) that modulate growth in many tissues. The ternary complex (IGF-I/IGFBP-3/ALS) provides an acute regulatory mechanism in which IGF-I may be mobilized from the circulating reservoir of 150 kDa complexes to the tissues. Acute exercise is known to be a stimulus for GH secretion. The beneficial effects of scheduled exercise on body composition are also well established. However, the impact of strenuous exercise on the pubertal development of child athletes is still not well understood. The first goal of this study was to assess the acute effects of high intensity exercise training on GH-dependent ternary complex components in female rhythmic gymnasts compared to age-matched healthy female controls with normal physical activity. The second goal was to explore the influence of these exercise-induced changes on skeletal and pubertal growth in the same group prospectively over a period of 4 years. SUBJECTS AND METHODS: Seventeen female rhythmic gymnasts, aged 11.4 +/- 0.9 years, who had 10 h per week intense exercise for at least 4 months volunteered to participate in this study. Anthropometric measurement of height (Height SDS for chronological age [HtSDS(CA)], parentally adjusted height, predicted adult height), bone age and weight (BMI) were made using standard techniques in gymnasts and controls (aged 12.5 +/- 3.0 years, n = 12). Gymnasts were followed up to 4 years to observe growth velocity and pubertal progression. In order to determine the acute impact of exercise on levels of GH and GH-dependent ternary complex component (IGF-I, IGFBP-3, ALS, IGF-I/IGFBP-3 molar ratio), blood samples were obtained from gymnasts after a routine 2-h high-intensity training program and then after a 2-day rest period. These results were compared with age-matched controls with no scheduled sports activity. RESULTS: Despite the significant increment in serum GH and GH-dependent components immediately following the exercise, serum GH/IGF-I levels showed a significant decrement (p < 0.01) after a 2-day rest in gymnasts, to a nadir as low as those of the control subjects' baseline levels (p < 0.01). There was no difference in anthropometric characteristics of gymnasts and controls except BMI; gymnasts were leaner than controls. During a 4-year follow up, there were no differences between the gymnasts and controls in regard to skeletal growth and reaching their predicted height. However, in gymnasts there was a delay in pubertal tempo but not in growth. CONCLUSION: Intense exercise induces an acute rise in GH levels, but this acute elevation rapidly normalizes after a 2-day rest in female rhythmic gymnasts. These fluctuations in serum GH and GH-dependent ternary complex components had no reflection on the skeletal growth patterns in gymnasts over the 4-year follow up but there was a delay in their pubertal progression. PMID- 15270409 TI - An unusal case of hermaphroditism--a 46,XX/69,XXY chimera. AB - A diploid/triploid karyotype is an uncommon but important cause of true hermaphroditism and ambiguous genitalia. Individuals have a recognisable phenotype and characteristic hydatidiform placental changes. We report a 46,XX/69,XXY chimeric hermaphrodite. This case highlights the typical features (large placenta, intrauterine growth retardation, asymmetric growth, cranio facial anomalies, syndactyly and pigmentary dysplasia). It illustrates the importance of obtaining skin and gonadal karyotypes in the case of genital ambiguity, as the venous lymphocytic karyotype is usually diploid. PMID- 15270410 TI - Bilateral adrenal cysts and ectopic pancreatic tissue in Beckwith-Wiedemann syndrome: is a conservative approach acceptable? AB - Beckwith-Wiedemann syndrome is a common overgrowth syndrome associated with an increased risk of neoplasias which might be explained by the nature and localization of the genetic defect. While malignant tumors are often associated with hemihypertrophy, benign tumors are also found. We report a patient with the typical features of Beckwith-Wiedemann syndrome with two histologically different abdominal tumors, bilateral cystic adrenals and ectopic pancreatic tissue present at birth. In both tumors no malignancy could be detected. Ectopic pancreatic tissue is rarely seen and has been described in Beckwith-Wiedemann syndrome only once. After extirpation of the ectopic pancreatic tissue the cystic adrenals were left in situ since macroscopically no normal adrenal tissue could be identified and separated. Regular ultrasound examinations revealed complete resolution of the cystic adrenals within 24 months. Thus it seems that a conservative approach in selected tumors associated with the Beckwith-Wiedemann syndrome might be acceptable. PMID- 15270411 TI - Bardet-Biedl syndrome with syndrome X: a patient report. AB - Bardet-Biedl syndrome (BBS) is an autosomal recessive condition with a wide spectrum of clinical features. The principal manifestations are rod-cone dystrophy (sometimes called atypical retinitis pigmentosa), postaxial polydactyly, central obesity, mental retardation, hypogonadism, and renal dysfunction. The clinical diagnosis of syndrome X defines a patient with abnormal glucose metabolism, hypertension, hyperlipidemia and obesity. We report here a 15 year-old girl with BBS presenting with syndrome X. PMID- 15270412 TI - Novel compound heterozygous AIRE mutations in a Japanese patient with APECED. AB - Autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED) is a rare autosomal recessive disorder defined by the presence of two of three conditions, namely, Addison's disease, hypoparathyroidism, and mucocutaneous candidiasis. APECED is caused by alteration in a single gene, named the autoimmune regulator (AIRE) gene. We report AIRE gene mutations in a Japanese female with APECED. The patient is a 22-year-old Japanese female who was diagnosed with Addison's disease, hypoparathyroidism, and mucocutaneous candidiasis at age 8 years. Sequence analysis of the AIRE gene revealed novel compound heterozygous mutations. One was 1471 delCinsTT in exon 11 (GenBank accession no. AB006682), which leads to a frameshift and premature truncation of a 502 amino acid protein. The other was a G-->A transition at IVS11+1. Her mother was heterozygous for 1471 delCinsTT and was normal homozygous for IVS11+1. We found novel compound heterozygous mutations in the AIRE gene of a Japanese female with APECED. PMID- 15270413 TI - Role of nephron sparing surgery in the treatment of centrally located renal tumors. AB - OBJECTIVES: In recent years many long-term data have suggested that nephron sparing surgery can be considered as an effective method of treatment also in patients with small, solitary, unilateral renal cell carcinoma and a normal contralateral kidney. Generally, partial nephrectomy is performed for peripheral tumors and usually is limited to imperative indication for central tumors to avoid hemodialysis. We retrospectively evaluate the value of tumor location on technical and oncological results, particularly in patients with elective indication. PATIENTS AND METHODS: Between 1993 and 2002, 112 patients underwent nephron sparing surgery at our institution. The tumor was centrally and peripherally located in 22 and in 90 cases, respectively. The tumor was discovered in 13 (56%) central and in 57 (63%) peripheral tumors incidentally. The indication was imperative in 12 and elective in 10 patients for central group, while it was imperative in 34 and elective in 56 patients for peripheral group. RESULTS: The mean renal ischemia time was longer in central group compared to peripheral group (20.81 versus 18.8 minutes p<0.05) and the collecting system was violated more frequently in central group (53% versus 28% p<0.05). Postoperative complications were higher for central tumors compared to peripheral tumors (18% versus 4% p<0.05) but the ultimate mean serum creatinine level was similar for central and peripheral tumors (1.36 versus 1.22 mg/dl). The mean tumor size was 39.69 mm in central group and 32.77 mm in peripheral group (p<0.05). The mean diameter of central tumors in imperative indication was 42 mm while in elective indication was 32 mm (p<0.05). Pathological tumor stage was T1 to T3 in 18 (82%), 1 (4%) and 3 (14%) cases in central group and in 81 (88%), 6 (7%) and 5 (5%) cases in peripheral group. Grades was 1 to 3 in 4 (18%), 15 (68%) and 3 (14%) cases in central group and in 21 (23%), 61 (66%) and 10 (11%) cases in peripheral group. There was no difference in 5-year cancer specific survival (91% versus 98%) or postoperative local tumor recurrences (9% versus 6%) in central tumors compared to peripheral tumors and there was no local recurrence in elective partial nephrectomy performed in central tumors. CONCLUSIONS: Nephron sparing surgery is technically more demanding in patients with central tumors. However there were no significant differences in cancer specific survival and local recurrence between centrally versus peripherally located tumors. Elective partial nephrectomy can be performed also in patients with central tumors as long as really less than 4 cm. PMID- 15270414 TI - The choice of urinary drainage in patients with ureteral calculi of solitary kidneys. AB - PURPOSE: To evaluate the safety and efficacy of the treatment of patients with ureteral calculi of solitary kidneys (UCSK) in relation to the modality of urinary drainage. MATERIALS AND METHODS: We have studied a total of 55 patients with UCSK during the period from 1999 to 2002, 15 of them (27.3%) with radio opaque calculi (ROC). The group included 13 female and 42 male patients, aged 36 to 62 years. In 24 (43.6%) patients the stone treated represented the first stone episode. The stones were mainly located in distal ureter (in 39 patients, 70.9%). Stone size was heterogeneous, with the majority of the patients (26 patients, 47.3%) having calculi no more than 5 mm wide. In 41 patients (74.5%) reasons for early intervention occurred such as urosepsis, persistent ureteral obstruction and acute pyelonephritis with anuria. The patients were divided in 2 groups of patients in accordance with the modality of urinary drainage: 30 (54.5%) patients with double-J stent (group A) and 25 (45.5%) with percutanous nephrostomy (group B). After carrying out conservative methods for stone passage in all patients, we performed extracorporeal lithotripsy (ESWL) (1-3 treatments) in 40 (72.7%) patients. After ineffective stone removal we used instrumental procedures, such as ureteroscopy (with electrohydraulic or ultrasonic lithotripsy). RESULTS: The spontaneous passage rate of UCSK after conservative treatment and ESWL was obtained in 23 (76.7%) patients of the group A and in 21 (84.0%) patients of the group B. In 7 (23.3%) cases with ROC we observed acute back pain and anuria when the double-J stent was removed after ESWL. Ureteroscopy was performed in all such patients. Percutanous nephrostomy was placed in 5 patients with ROC before endoscopic manipulation. In 2 cases with ROC we performed ureteroscopy without previous nephrostomy but urosepsis and intractable pain occurred. Electrohydraulic endoscopic lithotripsy was performed in 4 (16.0%) cases of group B after ineffective ESWL. Ineffective result of ESWL could be explained by the sacro-iliac location of the stone in such cases. CONCLUSIONS: Placement of percutaneous nephrostomy in patients with radio-opaque ureteral calculi in solitary kidney could give us a chance to make the lithotripsy easier. We showed the feasibility of endoscopic manipulation of the ureter in solitary kidneys preventing open surgical intervention. PMID- 15270415 TI - Proposed urodynamic pressure-flow nomogram to diagnose female bladder outlet obstruction. AB - OBJECTIVE: To investigate female bladder outlet obstruction by urodynamics and create a nomogram to propose in clinical applications. PATIENTS AND METHODS: We investigated by urodynamic studies 200 women referred for lower urinary tract symptoms. A total of 179 patients were available for analysis: 136 served as control and 43 as obstructed. The following urodynamic variables were studied and compared in both groups: Free peak urinary flow (Free Qmax), intubated peak urinary flow (Qmax P/F), Free voiding time, pressure-flow voiding time, Detrusor pressure at peak flow rate (PdetQmax), Maximum detrusor pressure (PdetMax), Free postvoiding residual, Pressure-flow postvoiding residual. All data are presented according to descriptive statistics as mean and standard deviation (SD). Comparisons between the control and between the obstructed group were performed by means of Student's t test for equality of means, p<0.01 were considered significant. By creating ROC curves we calculated sensitivity, specificity, and positive and negative predictive values in all the patients data to derive the optimal combination of Maximum flow together with detrusor pressure at maxflow, and Maximum flow together with Maximum detrusor pressure. RESULTS: Free peak urinary flow (Free Qmax) was 26.6+/-11 ml/seconds for the control and 10.9+/-3.6 ml/seconds for the obstructed group (p<0.001). Qmax P/F were 22+/-8.7 cm water in the control group and 10+/-3.9 cm water in the obstructed group (p<0.001). Free voiding time were in both 28.2+/-15 seconds and 48+/-24 seconds (p<0.001) respectively; P/F voiding time were 41.4+/-21 seconds and 78.2+/-52 seconds (p<0.001) respectively. PdetQmax in control and obstructed group were 17.2+/-11.3 and 27.6+/-12.5 cm water (p<0.001) respectively. PdetMax were 25.2+/-14.0 and 39.4+/-18.9 cm water (p<0.001) respectively. Free postvoiding residual 25.1+/ 37.4 and 74.6+/-79.3 cc (P<0.001). P/F postvoiding residual were respectively 24.9+/-44.7 and 96.0+/-102.6 cc (p<0.001). All the differences among the variables investigated in both groups were statistically significant. We calculated sensitivity, specificity, and positive and negative predictive values. According to receiver operating characteristics (ROC) curve analysis, the overall combining values of free max flow rate of 13 ml/sec or less and detrusor pressure at max flow rate of 22 cm water or greater we obtained a sensitivity of 55.8%, a specificity of 96.3%, a positive predictive value of 82.8% and a negative predictive value of 87.3%. Moreover we combined Max flow rate of 13 ml/sec and Maximum detrusor pressure of 38 cm water to obtain a sensibility of 48.8%, a specificity of 99.3%, a positive predictive value of 95.5% and a negative predictive values of 86%. CONCLUSIONS: Diagnosing female BOO is a challenge condition and an accepted pressure-flow nomograms are still missing. We propose our nomogram as a valid and reliable tool to investigate female bladder outlet obstruction. PMID- 15270416 TI - Our experience in the treatment of penile curvature. AB - OBJECTIVES: Evaluation of the authors' results achieved using corporoplasty in patients treated for penile curvature from 1/1/1993 to 31/12/2002. MATERIALS AND METHODS: 26 patients underwent corporoplasty to correct penile curvature: congenital curvature without hypospadias or epispadias in 22 patients (aged 17 to 42) and curvature due to La Peyronie's disease in 4 patients (aged 48 to 65). In the latter 4 cases corporoplasty was chosen, instead of plaque excision or incision, since the plaque was quite small-sized and well stabilized, causing an overall limited curvature in a proportionately long penis with perfectly preserved erectile functions. The Ebbehoj-Metz technique (plication of the tunica albuginea of the corpora cavernosa using introflecting double cross-over stitch of 2/0 Prolene, grasping deep into the tunica in 4 positions) was used in 24 patients and the Yachia procedure (longitudinal incision of the tunica albuginea of the corpora cavernosa followed by transverse suture using 2/0 PDS) in 2 patients, both affected by congenital curvature. RESULTS: Penile straightening was completely satisfactory in 24 patients, while 2 with congenital curvature, who had undergone the Ebbehoj-Metz technique corporoplasty, had recurring penile curvature. One of these last 2 patients, aged 17, required further corporoplasty, using the same procedure again, obtaining penile straightening. The other patient, aged 42, refused further operation. Although the 2 patients treated using the Yachia technique had a satisfactory curvature correction, for some months they reported feeling a bothersome bump on the corpus cavernosum in the cut and sutured area of the tunica albuginea, with moderate pain during erections. For this reason this technique was not used in other patients. The only complication reported by 1 patient was the formation of a redundant balanopreputial scar due to suture dehiscence, later removed under local anaesthesia. CONCLUSIONS: Considering their results, the authors consider corporoplasty a reliable and easy to perform procedure in the treatment of both congenital penile curvature and in well selected cases of penile curvature due to induratio penis plastica. PMID- 15270417 TI - Ileocecal with teniamyotomies and ileal detubularized neobladders: considerations about a videourodynamic study after a long-term follow-up. AB - OBJECTIVE: To evaluate the functionality of ileal detubularized reservoir and ileocecal neobladder with multiple teniamyotomies after a long term follow-up. MATERIAL AND METHODS: Eight patients with ileal detubularized reservoir (IR) and 10 with ileocecal neobladder with multiple teniamyotomies (ICUS) with an average follow-up of 95 months, were submitted to a videourodynamic digital fluorongiographic examination. The patients had the longest disease-free follow up of our series. RESULTS: Urodynamic data collected were almost good and comparable between IR and ICUS. Anyway the exams showed that the smooth intestinal muscles remain active both in non-detubularized and detubularized bladders even after years. Continence is mostly assured by the striated sphincter, which can withstand transient high pressure peaks but is less effective when facing prolonged pressure increases. While in some cases a valid micturition was achieved simply relaxing the perineal floor, in other cases micturition was obtained by an abdominal straining against the resistance of a contracted urethral sphincter/pelvic floor even after a long follow-up. CONCLUSIONS: Detubularization and teniamyotomies can equally help the striated sphincter function by increasing the neobladder compliance. Moreover we observed that a non-spherical neobladder was compatible with good clinical results as well, proving that neobladder shape was less important in achieving good functional performance. Furthermore, in some cases the optimal relaxation of the perineal floor made abdominal straining superfluous especially when neobladders had physiological capacity. Thus a perineal floor musculature training and its co ordination with abdominal muscles may improve the quality of micturition. PMID- 15270418 TI - Classification of cystic structures located at the midline of the prostate: our experience. AB - OBJECTIVES: Between January 1994 and February 2002, 9086 men underwent biplane TRUS at our institution for a variety of reasons. 781 of the 9086 men (8.6%) showed evidence of one or more intraprostatic cystic lesions. We propose a new classification of cystic structures located at the midline of the prostate. MATERIAL AND METHODS: We have designed a methodology that reclassifies cystic structures located at the prostate midline through the ultrasonically guided transrectal aspiration of the cystic structure, the analysis of the PSA level of the aspirated fluid and the presence of spermatozoa, radiological studies (cyst injection with contrast medium, vasography, retrograde and/or voiding cystourethrography and utricle injection with contrast medium) and endoscopic studies (cystourethroscopy). RESULTS: Upon completion of the methodology, we have classified and defined these structures as the following: simple prostatic cysts, cysts of the mullerian ducts, megautricle, megautricle with inclusion of the ejaculatory ducts, "pseudocystic" dilation of the ejaculatory ducts and utriculoceles. CONCLUSIONS: This new classification of cystic structures located at the prostate midline is simple, useful and steers one away from any possible confusion. PMID- 15270419 TI - Incidence and evolution of aortic aneurysm in patients with bladder cancer. AB - OBJECTIVE: The incidence of aortic aneurysm is increasing, due to age, hypertension, hyperlipemia and voluptuary abuse like smoking, the last one of the most important cause of bladder cancer. Our study analyzes the incidence of aortic aneurysm in a group of patients who underwent radical cystectomy for bladder cancer and its evolution during follow-up in relationship with surgical procedure and adjuvant therapy. MATERIALS AND METHOD: During pre-operative staging of 173 patients, all affected by bladder cancer and then treated with radical cystectomy, we studied aorta and iliac artery diameters, as a part of our ultrasound scan evaluation. All patients underwent post-operative measurement of normal and abnormal aorta and common iliac artery during follow-up. RESULTS: At the pre-operative staging 19 patients (10.9%) had aneurysms in the aortic-iliac axis (A.A.). During follow-up in 5 patients the A.A. did not develop, whereas in 14 cases it increased within 12 months after surgery and then with an increase <0.5 mm per year, with no relationship with type of surgical procedure, urinary diversion, adjuvant therapy. No cases required a vascular surgical approach during the follow-up. Only 1 patient of basal 154 normal ones developed an aneurysm of the common right iliac artery, treated with endoprosthesis. CONCLUSIONS: The natural development or risk of aneurysm rupture in patients with bladder cancer depends on its dimensions but also on radical surgery, urinary diversion or adjuvant therapies. In our experience all these factors seem not to influence aneurysms if present nor determine de-novo development. PMID- 15270420 TI - Twenty-years experience on genitourinary tuberculosis. AB - OBJECTIVE: We reviewed our surgical experience on genitourinary tuberculosis in the past 20 years in order to evaluate if any change in the incidence and management of this disease has occurred. PATIENTS AND METHODS: From 1980 to 1999, at our Institution, 102 patients underwent surgery for genitourinary tuberculosis. We recorded the data and the surgical procedure of these subjects and compared patients treated in period 1980-1989 to those submitted to surgery in the period 1990-1999. RESULTS: The overall incidence of surgical management of genitourinary tuberculosis in the past 20 years was 0.50% (102 cases on a total of 20,299 urological surgical procedures). In the decade 1980-1989 the incidence was 0.67% (70 cases out of 10,428 patients) and in the decade 1990-1999 it was 0.32% (32 cases out of 9,871 patients). Nephrectomy was the most prevalent surgical procedure performed in both decades. CONCLUSIONS: Despite the availability of effective antimycobacteric drugs, surgery continues to play a role in the management of genitourinary tuberculosis. This disease is very slow to progress with minimal and subtle symptoms, often resulting in irreversible damage of the organs involved by the time a diagnosis is established. PMID- 15270421 TI - Renal papillary adenocarcinoma with unusual metastases: case report and review of the literature. AB - OBJECTIVE: This study focuses on a case report on a patient with renal carcinoma who developed metastases at unusual sites. We also reviewed the literature, including the theories proposed by various authors on the possible etiology of these odd localizations. CASE REPORT: A 48-year-old patient underwent conservative surgery for renal carcinoma (papillary adenocarcinoma). Twenty-two months later, he developed mediastinal metastases and underwent immunotherapy; two years after that he had metastases to unusual sites such as the urethra and the prostate. The mechanisms responsible for metastases to these unusual sites are not entirely clear yet. We feel it is important to note the highly aggressive and multifocal nature of papillary adenocarcinoma, and the need to perform closer follow-up on these patients, particularly if nephron-sparing surgery has been performed. PMID- 15270422 TI - Ureteral endometriosis: an unusual case of a pelvic mass arising in the ureter and involving the rectum and uterine cervix. AB - The case of a 41-year-old woman with a pelvic mass arising in the left ureter is reported. The diagnosis of endometriosis was made on transperineal biopsy exclusively. After unsuccessful treatment with LH-RH analogues, the patient underwent ureteral resection and ureteroneocystostomy. At six months' follow-up, she is asymptomatic with no evidence of hydronephrosis. PMID- 15270423 TI - Renal sarcoma associated with adult polycystic kidney disease. A case report and literature review. AB - The development of renal cell carcinoma in Adult Polycystic Kidney Disease (APKD) has been reported in the literature; one of the features of renal malignancy in APKD is the difficulty to make a diagnosis, and the majority of cases reported are incidental findings at surgery or autopsy. We report a rare case of renal sarcoma in a patient with APKD. Sarcoma associated with APKD does not seem to have particular biological characteristics when compared with primary renal sarcoma; however the polycystic kidney represents an aggravating circumstance, because of the difficulty in making an early diagnosis of a disease with a poor prognosis. PMID- 15270424 TI - Persistent priapism and histological modifications of the erectile tissue. Two case reports. AB - Prolonged veno-occlusive priapism is associated with a high risk of fibrosis of the corpora and impotence. We present 2 cases of prolonged low-flow priapism who came under our observation more then 72 hours after the onset of priapism. The first case was a 51-years old man in which the aethiology of priapism was cauda equina compression by a L4-L5 discal haernia, not recovered after surgical decompression. The second case was a 23-years old man suffering of painful priapism lasting for more than 7 days due by abuse of cocaine, alcohol and psychopharmaceuticals. In both cases drainage and irrigation of the corpora followed by injection with an alpha-agonist agent has been insufficient. Detumescence has been obtained with shunt procedure and compressive bondage. The biopsy of the corpora cavernosa showed fibrosis. PMID- 15270425 TI - Post-traumatic ureteropelvic junction obstruction. AB - Ureteropelvic junction (UPJ) injuries secondary to blunt abdominal trauma are rare, have been traditionally described in children and consist in laceration or avulsion of the ureter. The diagnosis is typically delayed owing to associated severe lesions and absence of hematuria in many cases. Late sequelae of non penetrating ureteral injuries have only been anecdotally described in world literature. To the best of our knowledge we report on the first case of UPJ obstruction in an adult man diagnosed 10 years after a blunt abdominal trauma. PMID- 15270426 TI - Canal wall down tympanoplasty surgery with or without ossiculoplasty in cholesteatoma: hearing results. AB - Aim of the study was to evaluate the functional results, in two homogeneous groups, for severity of the disease, submitted to canal-wall-down tympanoplasty (TPL CWD), with and without ossiculoplasty. A total of 60 patients who underwent canal-wall-down tympanoplasty for cholesteatoma were evaluated: 31 underwent ossiculoplasty (group A) and 29 classic Wullstein type III and IV operation (group B). Hearing results were evaluated 2 years after surgery according to the AAO-HHS guidelines. Pre-operative audiometrics revealed an air conduction PTA (AC PTA) of 45.12 dB in group A, and 56.25 dB in group B. Bone conduction PTA (BC PTA) was 16.86 dB in group A and 26.06 in group B. Two years after surgery, AC PTA was 42.98 dB in group A and 58.65 dB in group B; BCPTA was 18.79 dB in group A and 25.13 dB in group B. The mean pre-operative ABG was 28.44 dB in group A and 30.14 dB in group B. Two years after surgery, group A showed a mean ABG of 24.06 and group B of 35.54 dB, the difference between the two groups was significant (p = 0.03). In conclusion, the functional results observed in the present study support the need to reconstruct the ossicular chain in canal-wall-down tympanoplasty, in fact, ossiculoplasty is associated with a better hearing gain than the classic Wullstein operations. PMID- 15270427 TI - Role of dynamic posturography (Equitest) in the identification of feigned balance disturbances. AB - Clinico-instrumental criteria to reliably detect simulated vertigo remain to be defined. Computed dynamic posturography (Equitest) has been used to identify additional factors to distinguish simulated, from real vertigo. The present study population comprised 23 normal subjects and 16 patients with documented vestibular impairment. Normal subjects were also studied during a state of simulated vertigo. In malingerers, the Equilibrium Score and the Composite Equilibrium Score showed a statistically significant reduction in all test conditions as compared to normal subjects, patients and "non-malingerers". Upon Sensory Analysis, statistically significant differences were found for the somatosensory component between malingerers and "non-malingerers". In 20/23 cases, Strategy Score values recorded in malingerers were 2 Standard Deviations lower than the mean obtained in "non-malingerers" in at least one of the six test conditions. By combining the latter observation with Goebel's 1st criterion it was possible to differentiate malingerers from non-malingerers with 86.9% sensitivity and 89.7% specificity. The Equitest, therefore, in combination with conventional methods, provides the clinician with an important tool, in the identification of a state of simulated vertigo. PMID- 15270428 TI - Dysphonia and laryngopharyngeal reflux. AB - The correlation between laryngo-pharyngeal reflux and dysphonia has been evaluated in patients without significant laryngoscopic findings and without vocal misuse. Studies were performed, using a validated questionnaire on typical reflux symptoms as well as instrumental means, e.g. videolaryngoscopy, multi electrode 24-hr oesophageal pH monitoring, vocal acoustic analysis, gastro oesophagoscopy, on 62 patients (51 male, 11 female) with dysphonia for > or = 3 months, selected from 350) consecutive patients presenting with voice disorders. Standard criteria were: absence of laryngeal neoformation (benign or malignant) and correct use of voice. Anti-reflux treatment was prescribed in all selected patients. A group of 62 selected patients without laryngo-pharyngeal disease were studied as controls. Mean values of the harmonic to noise ratio and maximum phonation time were pathological in all patients with dysphonia and significantly correlated (p = 0) with the entity of the larynx alteration. The 24-hour pH monitoring revealed gastro-oesophageal reflux in all cases with a clear prevalence of episodes in the upright, compared to supine, position. From a multiple regression analysis of pH-metric values, considered important in predicting maximum phonation time and harmonic to noise ratio alteration. the significant predictors (p < 0.01) were those parameters indicating the existence of a laryngo-pharyngeal reflux disease: in an upright position, the prevalence of the number of refluxes and of time of pH < 4. In conclusion, the association between electro-acoustic reliefs and laryngoscopic data, as well as an alteration in maximum phonation time and harmonic to noise ratio in patients with pH-metric indicative parameters of laryngo-pharyngeal reflux disease led to the hypothesis of a possible correlation between entity and duration of the reflux and dysfunction of the arytenoid muscles, upon which chronic vocal fatigue, with consequent laryngeal compensatory stress, depends. PMID- 15270429 TI - Fantoni's translaryngeal tracheotomy complications. Personal experience. AB - Tracheotomy is a surgical procedure which, in conditions of acute respiratory emergency, guarantees an adequate airway through the trachea whereas, in cases of chronic respiratory failure, it is used to improve ventilation through the reduction of the dead respiratory space. Over the last few years, surgical techniques used in tracheotomy have been considerably modified, not only to respond to the needs of clinical indications but also on account of problems related to management of the patient and tracheostomy tube, particularly in the home setting. Besides traditional surgical techniques, in fact, in the Intensive Care Unit, percutaneous dilatative procedures are being used with increasing frequency, in particular, translaryngeal tracheotomy according to Fantoni. The latter, however, according to reports in the literature, has been shown to be followed by a higher peri-operative complication rate (40%) which involves maintenance of good function of the tracheostomy, a condition which is particularly dangerous in the management of patients in the home setting. Personal experience is described in the management of 6 patients submitted to tracheotomy according to Fantoni and in combined home treatment, who, some time after the operation. presented 'embedding' of the tracheostomy tube in the tracheostomy opening. The six patients were treated at home with ventilatory support using automatic ventilation system and were submitted, in our Clinic, to a surgical review with preparation of a tracheotomy according to the conventional method. Our experience showed a particular feature of the difficulty in the management of patients presenting respiratory diseases, submitted to translaryngeal tracheotomy and, thereafter, maintained in combined home treatment: in these subjects, in fact, the presence of the tube, the difficulty in cleaning the peristomial skin, the reduced autonomy from the automatic ventilation system and the frequent coexistence of mucopurulent tracheo-bronchial inflammatory diseases, trigger micro-lesions of the stoma and, therefore, scar keloid, narrowing of the lumen and embedding of the tube itself. In conclusion, in our personal experience, we are of the opinion that translaryngeal tracheotomy, since it is easily carried out and is a slightly invasive procedure, plays a very important role in the management of the Intensive Care Unit patient but should be reserved for the few cases requiring tracheostomy for limited periods of time, in low risk patients and within the first 18 days after the acute damaging event. PMID- 15270430 TI - Giant cell granuloma of the maxilla: case report. AB - Giant cell granuloma is an uncommon bony lesion in the head and neck region, most commonly affecting the maxilla and mandible. Although it is a benign disease process, it can also be locally destructive. Surgery is the traditional and still the most accepted treatment for giant cell granuloma. The case described here involved the maxilla which was treated with surgical excision, followed by local injection of steroids. PMID- 15270431 TI - Burkitt-like lymphoma of the sphenoid sinus: case report. AB - Burkitt's lymphoma is a malignant endemic neoplasia with a mandibular localization, described for the first time in 1958, in African children. The World Health Organization classification recognises several variants of Burkitt's lymphoma; all are highly malignant B cell lymphomas. Besides Burkitt's sporadic, endemic lymphoma and Burkitt's lymphoma associated with AIDS, the World Health Organization classification includes an "atypical or pleomorphic" variant of Burkitt's lymphoma. This subtype includes those cases diagnosed as "Burkitt-like" lymphoma in the REAL (Revised European-American Classification of Lymphoid Neoplasm). The therapeutic protocol is similar to that used for classic Burkitt's lymphoma, with chemotherapy being standard treatment. Prognosis is extremely poor, with a mean survival of < 1 year. The case is described of a sinus-nasal "Burkitt-like lymphoma", originating within sphenoid sinus. The extremely rare localisation of this histological variant of Burkitt's lymphoma is stressed as well as the extremely aggressive nature of the neoplasm. PMID- 15270432 TI - Monophasic synovial sarcoma of hypopharynx: case report and review of the literature. AB - Synovial sarcoma (SS) is a malignant mesenchymal neoplasm usually involving the lower limbs of young adults. Localization in head-neck district is rare. Histologically, these are characterised by a biphasic or monophasic variant, the latter being more rare and difficult to identify. Immunohistochemistry plays a crucial role in the diagnosis. Cytogenetics also play an important role since both the monophasic and the biphasic forms are characterised by a reciprocal translocation (x;18) (p 11.2;q 11.2). Treatment options include an aggressive surgical approach and radiotherapy, whereas the role of chemotherapy remains to be defined. The case is described of monophasic synovial sarcoma located in the hypopharynx and a review is made of the literature concerning this rare neoplasm. PMID- 15270433 TI - Actinomycosis of submandibular gland: an unusual presentation. AB - An unusual presentation of oro-facial actinomycosis, mimicking the clinical appearance of a malignant lesion is reported. The patient, a 74-year-old female, presented with a right submandibular mass, which slowly grew in size over a period of about 2 months, and a modest dysphagia. A painless cervical mass was palpable over the submandibular region. The rhino-pharyngo-laryngeal region, explored by flexible fiberoptic examination, was normal. At ultrasonography, a 2x2 cm infiltrating dyshomogeneous mass, involving the right submandibular gland, was visible. No connection with adjacent organs was found. There was no associated cervical lymphoadenopathy. Ultrasound-guided fine-needle aspiration cytology, performed on lesion, revealed no evidence of malignancy. The presence of characteristic colonies of actinomyces infection was found. The patient was treated initially with tetracycline chloridrate 100 mg: 1 tablet every 12 hours for 7 weeks, but a repeat ultrasonography showed no resolution. A further fine needle aspiration cytology showed no actinomyces infection in the specimen. The patient was treated with methylprednisolone, 20 mg every 24 hours, for 5 days. After steroid treatment, the patient has been well and, upon repeat ultrasonography, total resolution of the submandibular lesion was confirmed. In conclusion, the clinical presentation of cervicofacial actinomycosis is variable and may mimic a malignant lesion or chronic granulomatous infections. Diagnostic and therapeutic findings are discussed. PMID- 15270434 TI - Mortality for pancreatic cancer among aluminium smelter workers in Sardinia, Italy. AB - To investigate the relationship between exposure to polycyclic aromatic hydrocarbons (PAHs) and mortality for specifc cancer sites, 1152 men, employed for at least 1 year at a prebake aluminium smelter, were followed-up from 1972 until 31 December 2001. Exposure to PAHs was estimated from a detailed reconstruction of the working history experienced in the plant by each cohort member and from several environmental and personal shift-sampling measurements available, by task and working department, since 1979. Furthermore, information on smoking habits, previous jobs before engagement in the smelter and main clinical findings observed during the follow-up were collected from the personal medical files. This study showed no increased mortality for lung cancer or bladder cancer associated to exposure to PAHs. Mortality for pancreatic cancer, based on 6 observed deaths, was significantly higher than expected in the whole cohort (SMR 2.4; 95%CI 1.1-5.2) and particularly among workers employed in the anodes factory of the plant (SMR 5.0, 95%CI 2.1-12.1), where a relatively consistent exposure to PAHs has been estimated. The nested case-control study planned for pancreatic cancer cases, confirmed that, also after controlling for cigarette smoking, PAH exposure experienced in the anodes factory was associated with a significant increased risk of pancreatic cancer. A pre-existent diabetes mellitus and a potential occupational exposure to pesticides experienced in previous agricultural jobs were found as concurrent significant covariates increasing the risk. In conclusion, the relatively high exposure to PAHs, experienced in the anodes factory and particularly in the green-mill department of this prebake aluminium reduction plant, cannot be ruled out as one of the main factors in the multifactorial aetiology of the pancreatic cancers observed in this study. PMID- 15270435 TI - [Return to work of a pacemaker bearing worker: the relationship between health problems and electromagnetic interferences]. AB - The potential effects of electromagnetic fields is a problem that interest the public opinion, as the modern society expose all people to electromagnetic non ionizing radiations. The problem has a particular and important meaning facing the return to normal life and work conditions of a cardiopatic subject bearing a pacemaker (PM) or implantable cardioverter defibrillator (ICD). Electromagnetic interferences can produce temporary or permanent malfunctions in these devices. Checking for the absence of electromagnetic interferences is necessary considering that correct functioning of these medical devices is essential for the life of the bearer. Precautions normally adopted by these subjects are generally adequate to ensure protection from interferences present in life environment; for occupational environment, there is often lack of adequate information, also due to late involving of the doctor specialist in occupational health. This work intends to study in depth a specific job, a carpentry-workshop with welding activities, starting with a case of a PM bearer who asked a doctor specialist in occupational health to evaluate the problems involved in his return to work. Electric and magnetic fields produced by equipments present in the workshop were measured and compared to data supplied by the literature to evaluate the possibility of interactions in the normally functioning of implanted electronic devices. On the basis of our experience, we have found some criterions for specific risk assessement to adopt for the definition of operative protocols for return to work of PM or ICD carriers, also considering the lack of specific procedures and indications for the doctor specialist in occupational health. The collected information and data from the literature suggest that welding can be a risk for a subject with PM; as observed in experimental conditions, electromagnetic radiations can alter particular sensitive devices and those with uncorrected settings. PMID- 15270436 TI - [Indoor allergens in office. Evaluation of the work station]. AB - House dust mite and other indoor allergens play a prominent role in the pathogenesis of asthma and other allergic diseases. Several studies have shown a close relationship between sensitisation and/or onset of asthmatic symptoms and levels of indoor allergen exposure. Aim of the study was to investigate the concentration of specific markers of the indoor allergenic pollution, such as Der p 1, Der f 1, Mite Group 2, Fel d 1 and Bla g 2. Dust samples were taken using a standard method by means of a 1200 W vacuum cleaner connected with a dust sampling device (MITEST). A standard A4 size area has been vacuumed four times during 2 min. The concentrations of Der p 1, Der f 1, Mite Group 2, Fel d 1 and Bla g 2 were determined in dust samples from 53 different sources (office chair and carpet) using a commercial kit (DUSTSCREEN). House dust mite allergens were not always detectable in the offices. Indoor allergen concentrations (Der p 1, Der f 1, Mite Group 2, Fel d 1) were significant higher in the work station (chair) than in the carpet (p < 0.0001). Der 1 exceeded the current threshold for sensitization in about 1/4 of the samples. Der f 1 was predominant over Der p 1 according to other studies. A good correlation between the results of Der p 1 and Der f 1 was observed both in carpet and work station. Cat allergen was ubiquitous and predominantly detected in the chairs because of the employees' clothes. No appreciable levels for Mite Gr 2 and Bla g 2 were detected. Such an exposure for 8 hours in every working day may be an important occupational risk for the development of sensitization/elicitation symptoms to house dust mite. To reduce mite allergen levels are necessary preventive measure by means of specific techniques and products as barriers for preventing the direct contact with allergens. PMID- 15270437 TI - [Estimate of uncertainty of measurements in clinical laboratories and in environmental, occupational, and preventive medicine]. AB - According to recently issued (UNI CEI EN ISO/IEC 17025, UNI CEI ENV 13005 and prEN ISO 15189) standards, to assure the quality of analytical results and their comparability in time and in different places, testing and clinical laboratories must demonstrate to use validated methods, guarantee the traceability of their measurements and state the measurement uncertainty associated with each result. For some SI quantities, such as the mass, the traceability to SI and the estimate of measurement uncertainty are warranted by established methods. In the clinical laboratory and especially in preventive, environmental and occupational laboratory medicine, specific difficulties arise to warrant the traceability to the mole. On one hand, the whole concept of measurement uncertainty is new in laboratory medicine, on the other hand, its application faces practical difficulties, because of the wideness and the complexity of the analytical repertory, the lack of officially validated methods and matrix-specific reference materials traceable to SI. In this paper we discuss briefly the concept of measurement uncertainty and its meaning in comparison with other parameters used to define the performance of analytical methods. In addition, we describe the procedures recommended by international organisations for estimating measurement uncertainty and interpreting analytical results with an associated measurement uncertainty in comparison with limit values. PMID- 15270438 TI - [Galvanic technology in the workplace: prevention of occupational tumors]. AB - We studied 10 firms having production processes that involve galvanic technologies, with the goal to verify the "Titolo VII of D.Lgs 626/94". Research results showed out that there are many critical issues that underline the need of an improved awareness about the worker's exposure to carcinigenons materials. We also think that would be useful a greater cooperation among managers, occupational physicians and even workers. Firms cooperated in the research allowing us to fix some items and allowing us to reach results that represent one step ahead in the path through the whole management of cancer hazards: primary healthcare. PMID- 15270439 TI - [Burnout syndrome among health workers caring for AIDS patients: predictive variables]. AB - Psychological stress and coping strategies in staff working with AIDS patients were assessed using self report methods. MEASURES: Maslach Burnout Inventory (MBI), Coping Orientations to Problems Experiences (COPE), State-Trait Anxiety Inventory (STAI) and Depression Questionnaire (DQ) were completed by staff from 20 hospitals of North-Center Italy, including 329 doctors and nurses working with people with AIDS. The results suggested important correlations among burnout, coping style, depression and anxiety. Inadequate strategies used as Focusing on and Venting of emotion, Behavioral Disengagement and depression predicted high level of Emotional Exhaustion and Depersonalization, while Personal Accomplishment were predicted by more adequate strategies (Planning, Restrain coping and Seeking social support) and low level of anxiety. PMID- 15270440 TI - [Occupational stress and assessment of human resources in health: from assessment to the changing process]. AB - Health professionals are at risk for occupational stress, as confirmed by diverse sources including the Italian legislative decree, D.L. 626/94, the Health Promoting Hospital of the World Health Organization, and the Ottawa Charter. The aim of this study was to analyze quantitatively and qualitatively the principal sources of stress in the work environment and the resources, both individual and organizational, that health professionals feel they possess to cope with it. The instruments utilized for the first quantitative phase were: the Maslach Burn out Inventory, the Job Content Questionnaire, Coping Inventory for Stressful Situation and Team Climate Inventory. The study population consisted of 224 subjects belonging to the nursing profession (nurses-in-charge of ward, specialized and general nurses), working in the community and in hospital. The results show medium levels of burn out, and coping strategies that are primarily oriented towards a direct solution of the stressful situation. Vision is a critical aspect, ie. a clear perception of institutional choices and goals is lacking. The administration and analysis of the questionnaires was followed by a second phase in which, by means of the focus group methodology, the results were qualitatively analyzed and the health professionals stimulated to an active and pro-positive approach in the search for solutions to the critical situations. PMID- 15270441 TI - [A model for psychological assessment of mobbing]. AB - The phenomenon "mobbing" has been studied scientifically for twenty years. In this period, researchers have elaborated progressively more complex explanations for it. Despite the attempts to examine mobbing closely, and to define it accurately, further investigation, aimed at quantifying the factors that characterize it, is still needed. This study proposes a method for assessing mobbing and the personality features linked to it. The model was tested on a sample of 29 outpatients (19 males, 10 females) asking medical assistance for work-related psychopathological problems. Focusing on the occupational history and work environment, a psychological diagnosis (DSM IV-axis I) was made for 27 of them: 14 cases of generalized anxiety disorder, 6 subjects with depressive disorder, 1 dysthymic disorder, 1 post-traumatic stress disorder, 2 cases of adaptation disorder, 3 patients with personality disorder. Although the number of the examined subjects is limited, the proposed model seems to be utilizable in the clinical practice. PMID- 15270442 TI - [Welcome to the monographic course "In depth study of allergy", Pavia, March 3 31, 2004]. PMID- 15270443 TI - [Individual eye protective devices]. PMID- 15270444 TI - [Individual protection devices: anti-vibration gloves]. PMID- 15270445 TI - [Individual head protection devices]. PMID- 15270446 TI - [Work fitness and pregnancy]. PMID- 15270447 TI - [Neuropsychological course of patients with craniocerebral injuries in rehabilitation]. AB - The aim of the present paper is to describe a neuropsychological assessment and intervention model in Traumatic Brain Injury (TBI) patients. The theoretical and methodological frameworks are described and the following diagnostic and rehabilitative flowchart is fully explained: 1. first visit with patient and his relatives; 2. clinical and testing assessment; 3. diagnostic balance and its communication to patient and his relatives; 4. neuropsychological rehabilitation and psychological counseling. Whenever necessary, patient's relatives are involved. Furthermore, TBI patients' health related quality of life is outlined as an important clinical and scientific issue deserving more attention, in spite of the objective methodological difficulties which its evaluation implies. PMID- 15270448 TI - [Assessment of function recovery in patients with total knee prosthesis]. AB - The aim of this study was to analyse the evolution of motor performance in a group of patients who underwent surgical knee replacement. We also assessed patient satisfaction at 3 and 9 months after the operation. Sixty-two patients (40 women, 22 men, mean age 72.4 years) underwent isokinetic evaluation associated with surface EMG. All the patients had undergone a total prosthetic knee joint replacement for arthrosis, and had followed a standardised 30-day rehabilitation program at our centre for recovery of knee joint function, strengthening of the flexor-extensor muscles and restoration of ambulation as best as possible. Each of these patients underwent isokinetic evaluations 3 months and 9-10 months after their operation. The isokinetic test consisted of carrying out 5 flexion-extensions of the knee at an angular velocity of 60 degrees/second, followed by an endurance test of 30 repetitions of flexion extension of the knee at 120 degrees/second. This isokinetic test provided data on extensor strength and flexor strength; furthermore, specifically designed software allowed simultaneous visualisation of the surface EMG tracing, torque and joint excursion. The first analysis showed a macroscopic decrease in the strength of the flexor-extensor muscles of the knee. This muscle weakness was clearly evident 3 months after the operation and was particularly pronounced for the extensor muscles of the operated limb compared with the muscles of contralateral, unoperated limb. Muscle weakness was still present 9 months after the operation although there had been a considerable improvement compared with 6 months previously; the imbalance in the flexor/extensor ratio, which differed from that in the contralateral, unoperated limb, was noted to be still present. The surface EMG demonstrated persistent myoelectrical activity in the flexors even when the extensor activity was predominant: this is an expression of imbalance between agonists-antagonists. A questionnaire administered to the patients about their satisfaction with the operation revealed that 9 months after the surgery 23 patients still complained of continuous joint pain with increased loads, 27 complained of frequent pain with load-bearing and only 12 no longer complained of any disturbance, manifesting full satisfaction with the operation. Total knee replacement is thus a valid treatment in those cases in which degenerative joint disease necessitates a radical solution. Nevertheless, dissatisfaction with the operation is common, and may be due to persistence of joint pain and incomplete joint recovery of joint function or muscle strength. In our series of patients, we found that although there was progressive recovery of strength of both the flexor and extensor muscles, a considerable imbalance remained between the operated and unoperated limbs; this finding was also confirmed by the surface EMG investigation. PMID- 15270449 TI - The Human Tissue Bill--an opportunity about to be missed? PMID- 15270450 TI - A market study of green spray paints by Fourier transform infrared (FTIR) and Raman spectroscopy. AB - A market study of 40 different green spray paints was carried out using infrared (FTIR) and Raman spectroscopy. The infrared technique distinguished between the 12 main groups based on their binder and extender composition. After visual comparison of the spectra 22 subgroups were observed. Raman spectroscopy was also carried out on the 40 reference paints in order to determine the pigment content. Analyses were undertaken using two different excitation sources: Argon ion (514.5 nm) and Helium-Neon (632.8 nm). The first generated strong fluorescence for most of the samples and created eight groups. Using the red laser, 15 classes were observed. Finally, using an analytical sequence starting with infrared spectroscopy followed by Raman Helium-Neon and then by Raman Argon laser, most of the paints were differentiated. In this study infrared and Raman spectroscopy complemented each other. FTIR supplied information about the binder and some extenders, and Raman provided information on the main organic pigments present. PMID- 15270451 TI - The Horizontal Gaze Nystagmus test: fraudulent science in the American courts. AB - The Horizontal Gaze Nystagmus (HGN) test was conceived, developed and promulgated as a simple procedure for the determination of the blood alcohol concentration of drivers suspected of driving while intoxicated (DWI). Bypassing the usual scientific review process and touted through the good offices of the federal agency responsible for traffic safety, it was rushed into use as a law enforcement procedure, and was soon adopted and protected from scientific criticism by courts throughout the United States. In fact, research findings, training manuals and other relevant documents were often held as secrets by the state. Still, the protective certification of its practitioners and the immunity afforded by judicial notice failed to silence all the critics of this deeply flawed procedure. Responding to criticism, the sponsors of the test traveled the path documented in this paper that led from mere (if that word can ever truly apply to a matter of such gravity) carelessness in research through self-serving puffery and finally into deliberate fraud--always at the expense of the citizen accused. PMID- 15270452 TI - A study in relation to the random distribution of four fibre types on clothing (incorporating a review of previous target fibre studies). AB - A new target fibre study was undertaken building upon eight previous published studies, which are reviewed here. Fifty-eight items of outer clothing, obtained from households across England, were taped and searched for three types of commonly occurring fibres and one less common fibre type. The tapes were examined using a combination of automatic and manual searching. Any fibres similar to the target fibres were removed and examined using microspectrophotometry, comparison microscopy, thin layer chromatography (TLC) and fourier transform infrared spectroscopy (FTIR) where applicable. One polyester fibre, four acrylic fibres and eleven wool fibres were found that matched the three commonly occurring target fibre samples. None of the less common polyester fibres were found. One item had six matching wool fibres and another had the polyester fibre and an acrylic fibre recovered from it. The remaining eight fibres were each recovered from individual items. The results suggest that finding a large number of fibres that match a control is unlikely to happen by chance, which supports the findings of the previous target fibre studies. PMID- 15270453 TI - Autobrewing revisited: endogenous concentrations of blood ethanol in residents of the United Arab Emirates. AB - Endogenous ethanol concentrations in blood were determined by sensitive headspace gas chromatography/mass spectrometry in 1557 residents of the United Arab Emirates. The subjects were from 13 nationalities, of both sexes and of different age groups. There was no significant difference in blood ethanol concentration between nationalities or between sexes within and between nationalities. The data was pooled and the overall median, minimum, maximum, 25% percentile and 75% percentile were 0.04, 0.00, 3.52, 0.01 and 0.09 mg/dl respectively. The values of blood ethanol concentration as reported in this study indicate that they are far too low to have any forensic significance. PMID- 15270454 TI - Establishing the most appropriate databases for addressing source level propositions. AB - Previous papers in Science & Justice have described the work of the Case Assessment and Interpretation (CAI) project that has been running for several years within the Forensic Science Service (FSS). The principles of the CAI model, which have developed through casework, are the foundation of a balanced, robust and logical approach to interpretation. The question arises frequently as to what is the most appropriate database that should be available to assist in assigning a value to a given probability. In this paper we present a set of guidelines in the form of flowcharts and explore them within the context of a range of case examples. PMID- 15270455 TI - Dynamic of the ageing of ballpoint pen inks: quantification of phenoxyethanol by GC-MS. AB - The ageing process of some inks were studied to evaluate whether it is possible to date them. We used gas chromatography coupled with mass spectrometry to measure the evaporation of volatile components. The selected approach thus follows the disappearance of one ink volatile component (phenoxyethanol) as a function of time. The ink ageing curve represents the ratio of an evaporating compound to a stable compound of ink according to time. The results obtained are thus independent of the quantity of ink sampled for analysis. We obtained for two pens, containing two different inks, a curve highlighting an exponential decrease of the evaporating compound. By fitting these curves we determined the limitations of dating a ballpoint pen ink. Two distinct behaviours were observed in two distinct modes, the first called 'fast mode' and the second called 'slow mode'. In order to try to explain the phenomenon, the studies were based on solvent diffusion theory in complex matrix (such as polymer on varnish). Calculations from certain parameters showed an extremely fast evaporation of ink solvents, as well as varying behaviour depending on the paper used. The results showed that it is not possible to date ballpoint pen inks with this method in document examination casework. PMID- 15270456 TI - On reiterative justice. PMID- 15270457 TI - Multi-technique comparison of source and primary transfer soil samples: an experimental investigation by D Croft and K Pye. A comment. AB - The Croft and Pye paper has a story to tell about the differentiation capabilities of four analytical techniques. It is not however, the whole story. This may be due in part to the lack of reproducibility in the generation of results, and also in the effort to match soil samples and thus infer that the results provide stronger conclusions than is possible to make with this sort of analysis. More research must be done to establish in what instances these techniques and others provide reliable, reproducible, representative and accurate results, and in what capacity such results can be presented. As they stand, the results from this paper should be treated with great caution and not extrapolated to any other forensic case work until greater reliability of these and other techniques has been established. PMID- 15270458 TI - Fibre mapping--a case study. AB - Fibre mapping, more commonly known as one-to-one taping, was developed in Germany approximately twenty years ago. The technique facilitates the distribution of fibres on a surface to be recorded. The impact of this technique on the investigation of serious crime has been reported in the European Fibre Group on several occasions. This paper represents a case study of the application of the technique. It is believed to be the first time that this technique has been successfully applied in the United Kingdom. PMID- 15270459 TI - Scene it--done it. New technologies and approaches used at scenes. PMID- 15270460 TI - Workers of the world, panic! PMID- 15270461 TI - Bush tries to out-liberal the liberals on Medicare. PMID- 15270462 TI - Get ready for a life-and-death battle over obesity. PMID- 15270463 TI - While you were out. I'm doing the Fatkins diet. PMID- 15270464 TI - Clinical trial insurance in a comparative law perspective. AB - This paper presents the results of a comparative legal study on liability and insurance of clinical trials, including Belgium, France, Germany, the Netherlands, Spain, Switzerland, Sweden and the United Kingdom. In most countries the right to compensation of the trial subject is safeguarded, but the existing regimes show much variety. Seen from the perspective of the trial subject there is no justification for linking the extent of compensation to the object of the trial (involving drugs or not), to the nature of the trial (therapeutic or non therapeutic) or to the status of the researcher (public entity or not). PMID- 15270465 TI - A reassessment of the right to health care. AB - This paper examines the right to health care in the Netherlands as provided for under the Constitution. The author discusses the relationships between human rights, economic forces and political choices in this connection and offers a view that these are due for reassessment. PMID- 15270466 TI - Legislation for Ukraine's public health: the current situation and paths to further development. AB - The paper contains the findings of an analytical study of the current national health care legislation of Ukraine and identification of priority areas in its further development. One of the key objectives of the above study was to identify the compliance of the national healthcare legal framework with the approaches to health policy formation, which are set out in the documents of global and European regional international organizations, and to assess whether the national legislation includes that spectrum of functions that are to be covered by the health care legislation. The analysis showed that Ukraine's national healthcare legislation is rather strong and well developed. Though the national healthcare legislation is mostly in line with international approaches to the state health care policy formulation, the issue of enforcing already adopted laws and by-laws and ensuring their compliance by all legally established bodies, including state authorities and self-governments, citizens, NGOs, etc, is problematic. PMID- 15270467 TI - Maternal brain death, pregnancy and the foetus: the medico-legal implications for Ireland. AB - This paper examines some of the medico-legal issues that arose as a result of a situation which occurred in May 2001 in Ireland when a woman who was a British citizen and who was fourteen weeks pregnant collapsed and suffered a brain haemorrhage. She was taken to hospital where she was placed on life support but declared brain-dead. As a result of the uncertainty regarding the hospital's obligation to the foetus, life-support was maintained until further opinion was sought. After two weeks the foetus died and life support was only then discontinued. In Ireland there currently exists neither medical guidelines nor legislation to regulate such areas of medical practice. Also, the courts have not had the opportunity to comment on this particular matter and thus there exists widespread concern as to how healthcare providers will act if such situation were to occur again in the future. This article examines the following difficult medico-legal implications that arise from the above situation and especially in light of the constitutional protection of the unborn child in Ireland. PMID- 15270468 TI - Prevention of patient injuries: the Finnish patient insurance scheme. AB - Injuries sustained by patients in connection with health care or medical treatment in Finland are compensated for as provided in the Patient Injuries Act, which came into force on 1 May 1987. Between 1987 and 2003, a total of 95,411 claims were made to the Finnish Patient Insurance Centre, the body in charge of the claims handling. Every third claim qualified as a patient injury. In my presentation I will focus on examining how the 17 years' claims statistics are used for the prevention of patient injuries and for the improvement of health care quality control. While the Finnish Patient Insurance Centre considers injury prevention as an important issue, the significance of this work has also been widely discussed in public. Injury prevention is a common cause and a goal sought by all providers of health care and medical treatment. This preventive work is clearly linked to the development and management of the policyholders' quality control systems. Successful prevention work requires the creation of feedback systems, cooperation networks and increased interaction to ensure that all parties will be able to participate and commit themselves to the targets. The Finnish Patient Insurance Centre has upgraded information services and increased training e.g. by launching a wide training programme tailored to individual policyholders. By using the experience gained over the years, patient injuries can be prevented. This in turn reduces human suffering and results in increased patient satisfaction in the long run. Taking up issues in the workplace with an open and thorough approach also helps foster a positive workplace climate. From an economical point of view, insurance premiums are thus kept under better control, because premiums charged on public sector policies are fully experience rated. PMID- 15270469 TI - Road traffic accidents and secondary victimisation: the role of law professionals. AB - AIMS: The purpose of this study is to contribute to the understanding of the secondary victimization process related to the compensation procedures for physical injuries with regard to the role of law professionals. METHODS: Through the completion of an unsigned questionnaire, 81 victims of road traffic accidents reported their experience with the compensation process. The data obtained was submitted for statistical analysis. RESULTS: Factors which trauma victims considered as aggravating to their sequelae and suffering were: the compensation procedures, especially in judicial proceedings; the length of the process--when it exceeds one year; and the attitude of law professionals towards them. DISCUSSION: An intervention in the compensation system, whereby procedures and law professionals' attitudes are reviewed, is required to circumvent the secondary victimization of road traffic accident victims. PMID- 15270470 TI - Association between civil procedure and medical malpractice litigation in Japan. AB - The effect of shortcomings in the system of civil procedure in Japan, such as excessive delay and possible mistaken judgment on the existence of negligence, on medical malpractice litigation and legal outcomes has not been examined. Using data on judgments and the decisions in medical malpractice litigation by the Tokyo and Osaka District Courts, we examined the association between civil procedure and medical malpractice litigation, and predictors of the decisions of medical malpractice litigation. The basis of the civil procedure to secure quick decisions was related to the amount of medical malpractice litigation, but not to the decisions in medical malpractice litigation. Negligence and a factor other than negligence were related to the rate of decisions in favor of the plaintiff. Although the study implies that shortcomings in civil procedure negatively influence medical malpractice litigation, it was not determined whether decisions were made based on mistaken judgment concerning the existence of negligence. Since there are methodological limitations to this study, further studies are necessary to verify these findings. PMID- 15270471 TI - Promotion and enforcement of patients' rights. AB - European health systems have experienced a "right-revolution" in the last 10 years'. The existence of a large number of policy trends and normative initiatives in European countries demonstrates a strong interest in patients' rights. The increasing interest and involvement of the general public is also decisive and can favour further development and involvement by policy-makers and the legislators. Numerous measures exist to promote and enforce patients' rights. It is now time for the anti-cancer associations to play an active role in this important event and to try to define a new mission for the benefit of the patients. PMID- 15270472 TI - Ethics in the care of people with epilepsy--the perspective of community service organisations. PMID- 15270473 TI - Informed consent when taking genetic decisions. AB - Developments in genetics with diagnostic, pre-symptomatic and predictive testing involve significant changes in the decision-making process, because of the complexity of genetic information and the difficulty related to understanding the causes and mechanism of genetic diseases, ethical, psychological and social implications (psychological stress, anxiety, discrimination in employment and assurance, difficulties in interpersonal relationship), and indirect involvement of third parties. When taking genetic decisions, the patient should receive all the information about the objective and the type of the test, the hypothetical risk, the possibility of obtaining unexpected results, possible psycho-physical repercussion, and means of support for the long time that might pass between the diagnostic predictions and the possible onset of the disease: genetic counseling is a complex but essential operation for acquiring the informed consent of the patient. The outlined peculiarities of the process for informed consent in genetics requires the adequate training of medical personnel to manage the relationship with the patient in these complex cases. PMID- 15270474 TI - Informed consent--or the physician's anxiety delegated to the patient. AB - Medical information, the precondition of informed consent, the author argues, is passed on to the patient not only on the concrete cognitive level; deeper unconscious and emotional levels pertaining to transferential and countertransferential aspects of the patient-physician relationship may have an important impact on the healing process. Two short examples illustrate how the physician's own anxieties may interfere with his informing of the patient. The article concludes with a plea for including psychotherapeutic teaching in medical training. PMID- 15270475 TI - Guardianship and consent. AB - It has been argued elsewhere that "consent is the hallmark of our health care system". If this is correct, then what is the position of those who are not capable of deciding whether (or not) to give consent to health care? This paper briefly examines the law and ethics of substitute decision-making. Its principal arguments are three. Firstly, that because exercising a choice (or exercising one's autonomy) presupposes the capacity to do so, there are an increasing number of people who are not capable of exercising their autonomy in health care settings. Second, that as they are not capable of making an autonomous choice, the law permits another fundamental bioethical principle, that of beneficence, to operate so as to ensure that such people are not denied treatment which they may need. This principle is reflected in the use of Guardianship tribunals. Finally, very brief comment is made on the leading difficulty in this field, which is ascribing a clear meaning to the term competency--the standard which separates the people who are permitted to exercise an autonomous choice, and those not permitted to do so. PMID- 15270476 TI - "Cognac alibi" as a drunk-driving defense and medico-legal challenge. AB - Some drivers with positive forensic ethanol analyses, offer an explanation that they consumed alcohol a short time before a traffic accident or after driving. In medico legal practice this is commonly known as hip-flask defense, but to us as "cognac alibi" defense. In these cases, the lawyers require the medico legal experts to offer as much information as possible so that the court may come to the most reliable conclusions about the driver's blood alcohol concentration at the moment of the traffic accident (BAC(Acc)). At the Institute of Forensic Medicine our own analytical approach was established to study this medico legal problem. It consists of three inter-related phases in which it combines the obtained BAC values, with testimonies of the drunk driving suspect andalso witnesses. A specific algorithm was designed for calculating absorption and elimination of consumed alcohol. All the above-mentioned elements and blood ethanol values calculated according to Widmark's method were inserted into appropriate cells of MS Excel software in order to calculate BAC in the function of time. The result is a relevant analysis of the drunk driving suspect's BAC in 5-minute intervals, as well as a graphic representation in chart form. PMID- 15270477 TI - Registration of pharmaceuticals in Israel and its exemptions. AB - The requirement to obtain a preliminary approval for every pharmaceutical product in order to allow its regular distribution is an accepted practice worldwide. In Israel, the preliminary approval is known as registration in the national drug registry. The mechanism of drug registration has been in place since 1964, and is now based on Article 47A(b) of the Pharmacists Ordinance [New Version], 1981. The procedure of pharmaceutical registration is designed to assure that all pharmaceuticals marketed in Israel would prove to be safe, efficacious and of proper quality. However, in certain cases, it may lead to prevention of the distribution of necessary products that could not be registered for various reasons. In the context of pharmaceuticals, the unavailability of certain products may leave patients without proper treatment and put them in serious danger. This problem requires the duty of registration to have some exemptions. These exemptions must be balanced between the desire to assure a high level of preliminary evaluation of pharmaceutical products on the one hand, and the healthcare system's need to use products which have not been fully examined, on the other hand. This article analyzes the Israeli model of the exemptions from the duty to register drugs. PMID- 15270478 TI - Liability for pharmaceutical products: a difficult attempt at harmonisation. AB - On the 25 July 1985, the European Community adopted a Directive on liability for defective products. Doctors are very affected by this law because an injured person will possibly be able to take action against them. This directive had several aims: to harmonise member states' legislation concerning the legal protection of victims and to subject producers to the same framework of liability and competition rules. However, this directive currently remains relatively ineffective. Several reasons can be put forward to explain this failure. First, by leaving it open to member states to decide whether to opt for one or more derogations, the Directive has made it easier for different interpretations to emerge. Secondly, several ideas proposed by the Directive have not been defined. As a result, the different legal terminology used in different states makes the interpretation of these concepts more difficult. Our paper concludes that the Directive was soundly based. However, Member States must enquire about the solutions adopted in neighbouring countries, and the ECJ (European Court of Justice) should try to impress on states the importance of a community vision. Without an adequate level of convergence, the differences between national laws will encourage 'forum shopping' on the part of a claimant and the situation will become more difficult for prescribers. PMID- 15270479 TI - Memory accuracy of a real-life simulated incident. AB - Due to an awareness that both laboratory and real-life research present limitations, the present study attempts to combine the two research approaches, making use of the strengths of both methodologies. A real-life situation was created in a classroom and combined with laboratory conditions to determine the influence of certain variables (occupation, age, gender, race and retention interval) on the accuracy of eyewitness reporting. A total of 295 participants consisting of 12- to 14-year-olds, University students, the public and Police College students were divided into a short- and long-term memory group. The short term group were assessed immediately after witnessing the incident, while the long-term group were only assessed five to six weeks later. It was found that the effect of passage of time was insubstantial; however, the performance of the short-term group had already been disconcertingly poor. The school children did not perform significantly less well than other occupational groups, and the members of the Police did not perform significantly better than the other groups. No significant differences were found for the different gender and racial groups. PMID- 15270480 TI - Provision of assisted reproductive technology for single women in China: a new challenge. AB - Following the enactment of the Jilin Regulation, single women, for the first time, are allowed to access assisted reproductive services in China. This paper is intended to analyze the arguments over whether single women are entitled to access assisted reproductive services, in relation to Chinese legal, ethical and social characteristics. PMID- 15270481 TI - Consideration for a donation--economic aspects. AB - This article considers the economic aspects involved in introducing and putting into operation the model proposed by the author of this article. It rests on the premise that it would be worthwhile to provide money or other economic consideration in order to encourage and hence increase donations of human organs for transplant, whether from living or deceased donors. The model is based on an official central body, which would coordinate receipt of organs from donors, payment to the donors and distribution of the organs to patients, who would not pay anything for receiving the organ and would not even have any contact with the seller-donor. The purpose of this article is to try to examine and prove, that increasing the supply of human organs, whether from living or deceased persons by providing money or some other economic incentive would not only save the lives of persons waiting for organs, but also bring about substantial financial saving to the social community in general and the health system in particular, and thereby make available additional resources for saving lives, shorten the queues and benefit the weaker levels of society. We will show this through examining the various financial costs. This covers both the expense of treating various patients who need transplants on the one hand and the estimated costs to the community, including the patients and their families, on the other hand--as well as the loss of work force to the economy, payment of pensions, care from welfare organizations, etc. PMID- 15270482 TI - Effects of acid stress on Vibrio parahaemolyticus survival and cytotoxicity. AB - For several foodborne bacterial pathogens, an acid tolerance response appears to be an important strategy for counteracting acid stress imposed either during food processing or by the human host. The acid tolerance response enhances bacterial survival of lethal acid challenge following prior exposure to sublethal acidic conditions. Previous studies have revealed relationships between a foodborne pathogen's ability to survive acid challenge and its infectious dose. Vibrio parahaemolyticus is capable of causing gastroenteritis when sufficient cells of pathogenic strains are consumed. This study was designed to characterize acid sensitivities and to compare the effects of sublethal acid exposure (adaptation) on survival capabilities and cytotoxicities of different V. parahaemolyticus strains. Survival of acid challenge by stationary-phase cells differed by up to 3 log CFU/ml among the 25 isolates tested. No differences in acid resistance were found between strains when they were grouped by source (clinical isolates versus those obtained from food). Survival at pH 3.6 for log-phase cells that had been previously exposed to sublethal acidic conditions (pH 5.5) was enhanced compared with that for cells not previously exposed to pH 5.5. However, for stationary phase cells, exposure to pH 5.5 impaired both subsequent survival at pH 3.6 and cytotoxicity to human epithelial cells. Relative cytotoxicities of nonadapted stationary-phase cells were 1.2- to 4.8-fold higher than those of adapted cells. Sublethal acid exposure appears to impose measurable growth phase-dependent effects on subsequent lethal acid challenge survival and cytotoxicity of V. parahaemolyticus. PMID- 15270483 TI - An improved PCR primer pair based on 16S rDNA for the specific detection of Salmonella serovars in food samples. AB - Salmonella serovars are some of the major bacterial pathogens that can cause sporadic cases and outbreaks of foodborne illness. Based on the sequence data in the V3 region of the 16S rRNA gene, two PCR primer pairs have been designed for the detection of all serovars of Salmonella. However, none of these primers were specific for Salmonella because complete sequence homology with certain non Salmonella strains has been found within each of them. Thus, the specificities of these two primer pairs could not rely on only one of the two primers. In this study, we modified our previous 16SFI primer by extending one base at the 5' end and three bases at the 3' end. The modified primer, 16S-Sal, was designed with one or more mismatched bases near the 3' end of the primer annealing to the corresponding sequences of non-Salmonella strains. Such modification eliminates interference from Citrobacter freundii and Enterobacter cloacae as occurs with the 16SFI primer. When 16S-Sal and a degenerate primer, 16S-CCR, were used as a primer pair, detection specificity of Salmonella serovars was achieved. Because this primer pair was used for PCR detection of the salmonellae in food samples, such as whole milk and chicken meat, as low as 1 to 9 CFU/g (ml) of the food sample could be detected when a 8-h preculture step was performed prior to the PCR. For chicken meat, the endogenous microflora did not interfere with the PCR results. PMID- 15270484 TI - Inactivation of Salmonella during drying and storage of Roma tomatoes exposed to predrying treatments including peeling, blanching, and dipping in organic acid solutions. AB - The objective of this study was to evaluate the influence of predrying treatments, i.e., peeling, blanching prior to inoculation, and dipping in organic acid solutions, on inactivation of Salmonella during drying (60 degrees C for 14 h) and aerobic storage (25 degrees C for 28 days) of inoculated (five-strain composite, 7.1 to 7.4 log CFU/g) Roma tomato halves. Four predrying treatments groups were established. One group received no treatment (C). In the other three groups, unpeeled-unblanched, unpeeled-blanched (steam blanched at 88 degrees C for 3 min), peeled-unblanched, and peeled-blanched tomato halves were immersed for 10 min in water (W), ascorbic acid solution (AA; 3.40%, pH 2.48), or citric acid solution (CA; 0.21%, pH 2.51). Appropriate dilutions of homogenized tomato samples were spread plated on tryptic soy agar with 0.1% pyruvate and XLT4 agar for bacterial enumeration during drying and storage. Ten minutes of immersion in W, AA, or CA reduced bacterial populations by 0.7 to 1.6 log CFU/g. After 14 h of dehydration, total log reductions in the populations of bacteria were 3.2 to 4.5 (C), 3.7 to 4.9 (W), > 5.6 to > 6.1 (AA), and 4.5 to 5.5 (CA) log CFU/g, depending on type of agar used and condition of tomato samples. During drying and storage, the order of pathogen inactivation for predrying dipping treatments was AA > CA > W > C, with AA and CA rendering bacterial populations below detectable levels ( < 1.3 log CFU/g) prior to storage and between 7 and 14 days of storage, respectively. The results also indicated that peeling and blanching of tomatoes prior to inoculation may not necessarily affect destruction of Salmonella during the drying process. Use of predrying acid dipping treatments of tomatoes, especially in AA, may improve destruction of Salmonella during the dehydration process. PMID- 15270485 TI - Internalization of bacterial pathogens in tomatoes and their control by selected chemicals. AB - The effect of different washing or sanitizing agents was compared for preventing or reducing surface and internal contamination of tomatoes by Salmonella Typhimurium and Escherichia coli O157:H7. The tomatoes were inoculated by dipping them in a bacterial suspension containing approximately 6.0 log CFU/ml of each pathogen and then rinsing them with tap water, hypochlorite solution (250 mg/liter), or lactic acid solution (2%, wt/vol). All treatments were applied by dipping or spraying, and solutions were applied at 5, 25, 35, and 55 degrees C. With the exception of the lactic acid dip at 5 degrees C, all treatments reduced both pathogens on the surfaces of the tomatoes by at least 2.9 cycles. No significantly different results were obtained (P > 0.05) with the dipping and spraying techniques. For internalized pathogens, the mean counts for tomatoes treated with water alone or with chlorine ranged from 0.8 to 2.1 log CFU/g. In contrast, after lactic acid spray treatment, all core samples of tomatoes tested negative for Salmonella Typhimurium and, except for one sample with a low but detectable count, all samples tested negative for E. coli O157:H7 with a plate count method. When the absence of pathogens was verified by an enrichment method, Salmonella was not recovered from any samples, whereas two of four samples tested positive for E. coli O157:H7 even though the counts were negative. Few cells of internalized pathogens were able to survive in the center of the tomato during storage at room temperature (25 to 28 degrees C). The average superficial pH of tomatoes treated with tap water, chlorine, or lactic acid was 4.9 to 5.2, 4.1 to 4.3, and 2.5, respectively (P < 0.05), whereas no differences were observed in the internal pH (3.6 to 3.7) of the tomatoes treated with different sanitizers. The general practice in the tomato industry is to wash the tomatoes in chlorinated water. However, chlorine is rapidly degraded by organic matter usually present in produce. Therefore, lactic acid sprays may be a more effective alternative for decontaminating tomato surfaces. The use of warm (55 degrees C) sprays could reduce pathogen internalization during washing. PMID- 15270486 TI - Attachment of Salmonella Poona to cantaloupe rind and stem scar tissues as affected by temperature of fruit and inoculum. AB - A negative temperature differential between fruits or vegetables and the water in which they are immersed theoretically enhances infiltration of water and any microorganisms it might contain into tissues. The effect of temperature differentials between cantaloupes and wash water, each at 4 and 30 degrees C, on changes in cantaloupe weight and populations of Salmonella enterica Poona recovered from rinds and stem scar tissues of Eastern and Western (shipper) types of cantaloupes was assessed. The percent weight increase in Western cantaloupes was significantly greater (P < or = 0.05) than that in Eastern cantaloupes for all cantaloupe and inoculum temperature combinations. Salmonella Poona attachment to or infiltration of Eastern but not Western cantaloupe rind is enhanced when the fruit is at 4 degrees C, compared with 30 degrees C, regardless of the temperature of the immersion suspension. The number of Salmonella Poona cells recovered from rind tissue of Western cantaloupes at 30 degrees C immersed in inoculum at 30 degrees C was significantly less (P < or = 0.05) than that recovered from rind tissues of cantaloupes at 4 or 30 degrees C that were immersed in inoculum at 4 degrees C. Salmonella Poona in immersion water can adhere to or infiltrate surface tissues of cantaloupes. The populations of Salmonella Poona recovered from stem scar tissues of Eastern and Western types of cantaloupes were not significantly (P > 0.05) affected by cantaloupe and inoculum temperature combinations. Populations of cells adhering to or infiltrating various cantaloupe tissues is not dictated entirely by temperature differentials between fruits and immersion suspensions: rather, it also apparently is influenced by structures unique to surface tissues. PMID- 15270487 TI - Persistence of enterohemorrhagic Escherichia coli O157:H7 in soil and on leaf lettuce and parsley grown in fields treated with contaminated manure composts or irrigation water. AB - Outbreaks of enterohemorrhagic Escherichia coli O157:H7 infections associated with lettuce and other leaf crops have occurred with increasing frequency in recent years. Contaminated manure and polluted irrigation water are probable vehicles for the pathogen in many outbreaks. In this study, the occurrence and persistence of E. coli O157:H7 in soil fertilized with contaminated poultry or bovine manure composts or treated with contaminated irrigation water and on lettuce and parsley grown on these soils under natural environmental conditions was determined. Twenty-five plots, each 1.8 by 4.6 m, were used for each crop, with five treatments (one without compost, three with each of the three composts, and one without compost but treated with contaminated water) and five replication plots for each treatment. Three different types of compost, PM-5 (poultry manure compost), 338 (dairy manure compost), and NVIRO-4 (alkaline-stabilized dairy manure compost), and irrigation water were inoculated with an avirulent strain of E. coli O157:H7. Pathogen concentrations were 10(7) CFU/g of compost and 10(5) CFU/ml of water. Contaminated compost was applied to soil in the field as a strip at 4.5 metric tons per hectare on the day before lettuce and parsley seedlings were transplanted in late October 2002. Contaminated irrigation water was applied only once on the plants as a treatment in five plots for each crop at the rate of 2 liters per plot 3 weeks after the seedlings were transplanted. E. coli O157:H7 persisted for 154 to 217 days in soils amended with contaminated composts and was detected on lettuce and parsley for up to 77 and 177 days, respectively, after seedlings were planted. Very little difference was observed in E. coli O157:H7 persistence based on compost type alone. E. coli O157:H7 persisted longer (by > 60 days) in soil covered with parsley plants than in soil from lettuce plots, which were bare after lettuce was harvested. In all cases, E. coli O157:H7 in soil, regardless of source or crop type, persisted for > 5 months after application of contaminated compost or irrigation water. PMID- 15270488 TI - Efficacy of chlorine dioxide gas as a sanitizer of lettuce leaves. AB - Aqueous solutions of sodium hypochlorite or hypochlorous acid are typically used to sanitize fresh fruits and vegetables. However, pathogenic organisms occasionally survive aqueous sanitization in sufficient numbers to cause disease outbreaks. Chlorine dioxide (ClO2) gas generated by a dry chemical sachet was tested against foodborne pathogens on lettuce leaves. Lettuce leaves were inoculated with cocktail of three strains each of Escherichia coli O157:H7, Listeria monocytogenes, and Salmonella Typhimurium and treated with CLO2 gas for 30 min, 1 h, and 3 h in a model gas cabinet at room temperature (22 +/- 2 degrees C). After treatment, surviving cells, including injured cells, were enumerated on appropriate selective agar or using the overlay agar method, respectively. Total ClO2, generated by the gas packs was 4.3, 6.7, and 8.7 mg after 30 min, 1 h, and 3 h of treatment, respectively. Inoculated lettuce leaves exposed to ClO2 gas for 30 min experienced a 3.4-log reduction in E. coli, a 4.3-log reduction in Salmonella Typhimurium, and a 5.0-log reduction in L. monocytogenes when compared with the control. After 1 h. the three pathogens were reduced in number of CFU by 4.4. 5.3, and 5.2 log, respectively. After 3 h, the reductions were 6.9, 5.4, and 5.4 log, respectively. A similar pattern emerged when injured cells were enumerated. The ClO2, gas sachet was effective at killing pathogens on lettuce without deteriorating visual quality. Therefore, this product can be used during storage and transport of lettuce to improve its microbial safety. PMID- 15270489 TI - Effects of water source, dilution, storage, and bacterial and fecal loads on the efficacy of electrolyzed oxidizing water for the control of Escherichia coli O157:H7. AB - To evaluate the potential of using electrolyzed oxidizing (EO) water for controlling Escherichia coli O157:H7 in water for livestock, the effects of water source, electrolyte concentration, dilution, storage conditions, and bacterial or fecal load on the oxidative reduction potential (ORP) and bactericidal activity of EO water were investigated. Anode and combined (7:3 anode:cathode, vol/vol) EO waters reduced the pH and increased the ORP of deionized water, whereas cathode EO water increased pH and lowered ORP. Minimum concentrations (vol/vol) of anode and combined EO waters required to kill 10(4) CFU/ml planktonic suspensions of E. coli O157:H7 strain H4420 were 0.5 and 2.0%, respectively. Cathode EO water did not inhibit H4420 at concentrations up to 16% (vol/vol). Higher concentrations of anode or combined EO water were required to elevate the ORP of irrigation or chlorinated tap water compared with that of deionized water. Addition of feces to EO water products (0.5% anode or 2.0% combined, vol/vol) significantly reduced (P < 0.001) their ORP values to < 700 mV in all water types. A relationship between ORP and bactericidal activity of EO water was observed. The dilute EO waters retained the capacity to eliminate a 10(4) CFU/ml inoculation of E. coli O157:H7 H4420 for at least 70 h regardless of exposure to UV light or storage temperature (4 versus 24 degrees C). At 95 h and beyond, UV exposure reduced ORP, significantly more so (P < 0.05) in open than in closed containers. Bactericidal activity of EO products (anode or combined) was lost in samples in which ORP value had fallen to < or = 848 mV. When stored in the dark, the diluted EO waters retained an ORP of > 848 mV and bactericidal efficacy for at least 125 h; with refrigeration (4 degrees C), these conditions were retained for at least 180 h. Results suggest that EO water may be an effective means by which to control E. coli O157:H7 in livestock water with low organic matter content. PMID- 15270490 TI - The association between cleaning and disinfection of lairage pens and the prevalence of Salmonella enterica in swine at harvest. AB - A series of four field trials were conducted to evaluate the ability of a cleaning and disinfection procedure in swine lairage pens to reduce the prevalence of Salmonella enterica in slaughtered pigs. A cleaning and disinfection procedure was applied to lairage pens at a large Midwest abattoir. Each trial consisted of a cleaned (alkaline chloride detergent) and disinfected (H2O2 plus peracetic acid sanitizer) pen (treated) and a control pen, each holding 90 to 95 pigs for 2 to 3 h before slaughter. Ileocecal lymph nodes, cecal contents, and rectal contents were collected from 45 pigs from each study pen at harvest and cultured for S. enterica. In all trials, cleaning and disinfection reduced the prevalence of S. enterica-positive floor swabs in the treated pen (P < 0.05). However, the postharvest prevalence of S. enterica-positive pigs varied between trials. In trial 1, there was no significant difference in the prevalence of S. enterica in pigs between treatment and control groups. In trials 2 and 3, the prevalence of S. enterica was higher in pigs from treated pens versus pigs from control pens (91% versus 40%, P < 0.0001, and 91% versus 24%, P < 0.0001, respectively). In trial 4, the prevalence of S. enterica was lower in pigs from treated pens compared with pigs from control pens (5% versus 42%, P < 0.0001). This study indicates that cleaning and disinfection effectively reduces the amount of culturable S. enterica in lairage pens, but the ability of cleaned and disinfected pens to reduce the prevalence of S. enterica in market-weight pigs remains inconclusive. PMID- 15270491 TI - Validation of time and temperature values as critical limits for Salmonella and background flora growth during the production of fresh ground and boneless pork products. AB - To provide pork processors with valuable data to validate the critical limits set for temperature during pork fabrication and grinding, a study was conducted to determine the growth of Salmonella serotypes and background flora at various temperatures. Growth of Salmonella Typhimurium and Salmonella Enteritidis and of background flora was monitored in ground pork and boneless pork chops held at various temperatures to determine growth patterns. Case-ready modified atmosphere packaged ground pork and fresh whole pork loins were obtained locally. Boneless chops and ground pork were inoculated with a cocktail mixture of streptomycin resistant Salmonella to facilitate recovery in the presence of background flora. Samples were held at 4.4, 7.2C, and 10 degrees C and at room temperature (22.2 to 23.3 degrees C) to mimic typical processing and holding temperatures observed in pork processing environments. Salmonella counts were determined at regular intervals over 12 and 72 h for both room and refrigeration temperatures. No significant growth of Salmonella (P < 0.05) was observed in boneless pork chops held at refrigeration temperatures. However, Salmonella in boneless pork chops held at room temperature had grown significantly by 8 h. Salmonella grew at faster rates in ground pork. Significant growth was observed at 6, 24. and 72 h when samples were held at room temperature, 10 degrees C, and 7.2 degrees C, respectively. No significant growth was observed at 4.4 degrees C. Background flora in ground pork samples increased significantly after 10 h at room temperature and after 12 h for samples held at 10 and 7.2 degrees C. Background flora in samples held at refrigeration temperatures did not increase until 72 h. Background flora in the boneless chops increased significantly after 6 h at room temperature and after 24 h when held at 10 and 4.4 degrees C. These results illustrate that meat processors can utilize a variety of time and temperature combinations as critical limits to minimize Salmonella growth during production and storage of raw pork products. PMID- 15270492 TI - Thermal process validation for Escherichia coli O157:H7, Salmonella, and Listeria monocytogenes in ground turkey and beef products. AB - At 55 to 70 degrees C, thermal inactivation D-values for Escherichia coli O157:H7, Salmonella, and Listeria monocytogenes were 19.05 to 0.038, 43.10 to 0.096, and 33.11 to 0.12 min, respectively, in ground turkey and 21.55 to 0.055, 37.04 to 0.066, and 36.90 to 0.063 min, respectively, in ground beef. The z values were 5.73, 5.54, and 6.13 degrees C, respectively, in ground turkey and 5.43, 5.74, and 6.01 degrees C, respectively, in ground beef. In both ground turkey and beef, significant (P < 0.05) differences were found in the D-values between E. coli O157:H7 and Salmonella or between E. coli O157:H7 and L. monocytogenes. At 65 to 70 degrees C, D-values for E. coli O157:H7, Salmonella, and L. monocytogenes were also significantly (P < 0.05) different between turkey and beef. The obtained D- and z-values were used in predicting process lethality of the pathogens in ground turkey and beef patties cooked in an air impingement oven and confirmed by inoculation studies for a 7-log (CFU/g) reduction of E. coli O157:H7, Salmonella, and L. monocytogenes. PMID- 15270493 TI - Effect of sodium lactate on thermal inactivation of Listeria monocytogenes and Salmonella in ground chicken thigh and leg meat. AB - The effect of sodium lactate on thermal inactivation D- and z-values of Listeria monocytogenes and Salmonella was determined for chicken thigh and leg meat. At 55 to 70 degrees C, the D-value of L. monocytogenes in ground chicken thigh and leg meat with the addition of 4.8% sodium lactate (4.8 g sodium lactate per 100 g of meat) was 53 to 75% higher than that in the meat without sodium lactate. No significant difference was found for the D-values of Salmonella at 55 to 70 degrees C between the meat with and that without sodium lactate (4.8%. wt/wt). The z-values of both L. monocytogenes and Salmonella were not affected by sodium lactate (4.8%). The results from this study are useful for predicting thermal process lethality of L. montocytogenes and Salmonella in formulated chicken thigh and leg meat products. PMID- 15270494 TI - Use of carvacrol and cymene to control growth and viability of Listeria monocytogenes cells and predictions of survivors using frequency distribution functions. AB - The antibacterial action of carvacrol and cymene on two Listeria monocytogenes strains (STCC4031 and NCTN4032) was studied. Carvacrol or cymene showed inhibitory effect on the growth of L. monocytogenes during lag and exponential growth phases and was more evident with increasing concentrations in brain heart infusion broth at 30 degrees C. Carvacrol or cymene also decreased the survival of mid-exponential-growth-phase L. monocytogenes STCC4031 cells in potassium-N-2 hydroxy-ethylpiperazine-N-ethanesulfo nic acid, at 30 degrees C. The combination of carvacrol and cymene resulted in an increased antibacterial effect on the growth and a synergistic effect on the viability of L. monocytogenes compared with the natural compounds applied separately. The analysis of survival curves by the Weibull frequency distribution function allowed an accurate prediction of the level of inactivation achieved. Interestingly, an important bactericidal effect (4.7-log reduction) of low concentrations of both antimicrobials combined (0.75 mM) was observed on L. monocytogenes in carrot juice. This study indicates the potential use of carvacrol and cymene applied simultaneously for preservation of minimally processed foods. PMID- 15270495 TI - Distribution of Listeria monocytogenes molecular subtypes among human and food isolates from New York State shows persistence of human disease--associated Listeria monocytogenes strains in retail environments. AB - While there is considerable information available regarding Listeria monocytogenes contamination patterns in food processing plants, our understanding of L. monocytogenes contamination and transmission in retail operations is limited. We characterized 125 food, 40 environmental, and 342 human clinical L. monocytogenes isolates collected in New York State from 1997 to 2002 using automated ribotyping and hly allelic variation. All environmental isolates were obtained from retail establishments and the majority of food isolates (98 isolates) were obtained from foods that were prepared or handled at retail. Overall, food and/or environmental isolates from 50 different retail establishments were characterized. The 125 food and 40 environmental isolates were differentiated into 29 and 10 ribotypes, respectively. For 16 retail establishments, we found evidence for persistence of one or more specific L. monocytogenes strains as indicated by isolation of the same EcoRI ribotype from food or environmental samples collected in a given establishment on different days. The human isolates were differentiated into 48 ribotypes. Statistical analyses showed that two ribotypes were significantly (P < 0.0001) more common among food isolates as compared with human isolates. However, a total of 17 ribotypes found among the human clinical isolates were also found among the food and environmental isolates. We conclude that L. monocytogenes, including subtypes that have been linked to human disease, can persist in retail environments. Implementation of Listeria control procedures in retail operations, which process and handle products that permit the growth of L. monocytogenes, are thus a critical component of a farm-to-table L. monocytogenes control program. PMID- 15270496 TI - Influence of calcium lactate on the fate of spoilage and pathogenic microorganisms in orange juice. AB - Calcium lactate is used by the beverage industry as a source of calcium to fortify fruit juice. The objective of this study was to evaluate the influence of various concentrations of calcium lactate on the fate of pathogenic and spoilage microorganisms in orange juice. Commercial nonfortified orange juice was supplemented with calcium lactate at a concentration equivalent to 0, 5, 10, 15, 20, 25, or 30% dietary reference intake. The pH of each fortified juice was adjusted to 3.6 or 4.1. The prepared juice samples were inoculated separately with a three-strain mixture of salmonellae, a three-strain mixture of spoilage yeasts, and three single strains of spoilage bacteria including Alicyclobacillus acidoterrestris, Lactobacillus plantarum, and Lactobacillus sake. The contaminated juice was stored at 4 and 10 degrees C, respectively, for 6 to 7 weeks and assayed once a week for populations of salmonellae, spoilage yeasts, or spoilage bacteria. The results indicated that A. acidoterrestris was inhibited in all juice stored at 4 degrees C and low-pH juice stored at 10 degrees C. The bacterium, however, was able to grow at 10 degrees C in the high-pH juice with calcium lactate concentrations equivalent to 0 and 5% dietary reference intake. The cells of L. sake declined and eventually died off in low-pH juice stored at 4 and 10 degrees C and in high pH stored at 4 degrees C. But the organism flourished at 10 degrees C in the high-pH juice containing 0, 10, and 20% dietary reference intake of calcium lactate. The populations of L. plantarum remained approximately stable in low- as well as in high-pH juice stored at both 4 and 10 degrees C. While inhibited at 4 degrees C, the spoilage yeasts grew at 10 degrees C. Salmonellae died off in all juice stored at 4 degrees C and in low-pH juice stored at 10 degrees C. However, they persisted in the high-pH juice stored at 10 degrees C except in the samples that contained 20 to 30% dietary reference intake of calcium lactate. PMID- 15270497 TI - Microbiological quality of ground beef from conventionally-reared cattle and "raised without antibiotics" label claims. AB - The contamination of the food supply with pathogens and antimicrobial-resistant bacteria has emerged as an important health concern. We compared the microbiological quality of 77 samples of ground beef from conventionally raised cattle with 73 samples of ground beef from cattle raised without antimicrobial agents. Contamination with coliforms (1.7 log CFU/g) and Escherichia coli (0.51 log CFU/g) and Shiga toxin 2-producing E. coli (6% prevalence) was similar in both sample groups. Neither Salmonella. E. coli O157, nor vancomycin-resistant enterococci were isolated from any sample. Prevalence of E. coli resistant to ampicillin (39%), amoxicillin/clavulanic acid (23%), ceftriaxone (5%), tetracycline (19%), streptomycin (19%), kanamycin (11%), sulfamethoxazole/trimethoprim (2%), and gentamicin (1%) was similar in both groups. E. coli resistant to ciprofloxacin was not identified. Resistance to ceftiofur and chloramphenicol was more prevalent in beef from conventionally raised cattle at 18 and 30%, respectively, compared to 5 and 12% prevalence in beef from cattle raised without antimicrobial agents. These results do not correlate with the frequency of subtherapeutic use of these two antibiotics in beef production. Other factors in addition to, or in lieu of, the subtherapeutic use of specific antimicrobial agents in the preharvest stages of beef production may contribute significantly to the occurrence of antimicrobial-resistant bacteria in ground beef. PMID- 15270498 TI - Development of a laboratory scale clean-in-place system to test the effectiveness of "natural" antimicrobials against dairy biofilms. AB - A laboratory scale system, partially reproducing dairy plant conditions, was developed to quantify the effectiveness of chlorine and alternative sanitizers in reducing the number of viable bacteria attached to stainless steel surfaces. Stainless steel tubes fouled in a continuous flow reactor were exposed to a standard clean-in-place regime (water rinse, 1% sodium hydroxide at 70 degrees C for 10 min, water rinse, 0.8% nitric acid at 70 degrees C for 10 min, water rinse) followed by exposure to either chlorine (200 ppm) or combinations of nisin (500 ppm), lauricidin (100 ppm), and the lactoperoxidase system (LPS) (200 ppm) for 10 min or 2, 4, 8, 18, or 24 h. There was significant variation in the effectiveness of the alkaline-acid wash steps in reducing cell numbers (log reduction between 0 and 2). Following a 10-min treatment, none of the sanitizers significantly reduced the number of attached cells. Two hours of exposure to chlorine, nisin + the LPS, or lauricidin + the LPS achieved 2.8, 2.2, and 1.6 log reductions, respectively. Exposure times > 2 h did not further decrease the number of viable bacteria attached to the stainless steel. The effectiveness of combinations of nisin, lauricidin, and the LPS was similar to that of chlorine (P > 0.05), and these sanitizers could be used to decontaminate the surfaces of small-volume or critical hard-to-clean milk processing equipment. PMID- 15270499 TI - Prevalence of high-risk egg-preparation practices in restaurants that prepare breakfast egg entrees: an EHS-Net study. AB - Salmonella enterica serotype Enteritidis (SE) is a common cause of foodborne illness in the United States. Foods prepared with raw shell eggs have often been associated with SE outbreaks. The federal government published the Egg Safety Action Plan in December 1999 that called for reduction of egg-preparation practices that may contribute to the survival and proliferation of SE. In seven states, an interview and brief site evaluation of 153 restaurants that prepare eggs during all hours of operation was conducted by the Environmental Health Specialists Network to determine the prevalence of such practices. Fifty-four percent (83 of 153) of restaurants pooled raw shell eggs not intended for immediate service. These pooled eggs were held a median of 4 h for scrambled eggs, 5.5 h for omelets, and 6 h for pancakes and French toast. Nearly 26% (39 of 152) of restaurants reported storing eggs at room temperature, and 5% (7 of 152) stored eggs on ice or in cold-water baths before cooking. Generally, eggs were cooked to 72 to 83 degrees C, which is above the recommended final cook temperature of 63 to 68 degrees C. Employees reported sanitizing utensils used to prepare eggs less than once every 4 h in 42% (57 of 136) of restaurants. Several areas were identified in which further emphasis might reduce egg-associated SE infections in accordance with Healthy People 2010 goals. PMID- 15270500 TI - Effect of temperature and sanitizers on the survival of feline calicivirus, Escherichia coli, and F-specific coliphage MS2 on leafy salad vegetables. AB - We conducted a series of experiments to compare the survival of Escherichia coli, feline calicivirus, and F-specific coliphage MS2 on lettuce and cabbage with and without disinfection. Inoculated produce was held at 4, 25, or 37 degrees C for 21 days or was treated with different concentrations of sodium bicarbonate, chlorine bleach, peroxyacetic acid, or hydrogen peroxide. Survival was measured by the decimal reduction value (time to 90% reduction in titer) and the change in log titers of the test organisms. A stronger correlation of survival measures was observed between feline calicivirus and MS2 than between E. coli and either of the viral agents at 25 and 37 degrees C. The maximum time to detection limit for MS2 at all temperatures was 9 days, whereas feline calicivirus was detected for a maximum of 14 days at 4 degrees C. In contrast, E. coli was detectable for 21 days at 4 and 25 degrees C and for 14 days at 37 degrees C. Significant increases in E. coli titer occurred within the first 5 days, but virus titers decreased steadily throughout the experiments. E. coli was also highly susceptible to all disinfectants except 1% sodium bicarbonate and 50 ppm chlorine bleach, whereas the viruses were resistant to all four disinfectants. PMID- 15270501 TI - Autoinducer-2-like activity associated with foods and its interaction with food additives. AB - The autoinducer-2 (AI-2) molecule produced by bacteria as part of quorum sensing is considered to be a universal inducer signal in bacteria because it reportedly influences gene expression in a variety of both gram-negative and gram-positive bacteria. The objective of this study was to determine whether selected fresh produce and processed foods have AI-2-like activity and whether specific food additives can act as AI-2 mimics and result in AI-2-like activity. The luminescence-based response of the reporter strain Vibrio harveyi BB170 was used as the basis for determining AI-2 activity in the selected foods and food ingredients. Maximum AI-2 activity was seen on the frozen fish sample (203-fold, compared with the negative control) followed by tomato, cantaloupe, carrots, tofu, and milk samples. Interestingly, some samples were capable of inhibiting AI 2 activity. Turkey patties showed the highest inhibition (99.8% compared with the positive control) followed by chicken breast (97.5%), homemade cheeses (93.7%), beef steak (90.6%), and beef patties (84.4%). AI-2 activity was almost totally inhibited by sodium propionate, whereas sodium benzoate caused 93.3% inhibition, compared with 75% inhibition by sodium acetate. Sodium nitrate did not have any appreciable effect, even at 200 ppm. Understanding the relationships that exist between AI-2 activity on foods and the ecology of pathogens and food spoilage bacteria on foods could yield clues about factors controlling food spoilage and pathogen virulence. PMID- 15270502 TI - Campylobacter colonization of sibling turkey flocks reared under different management conditions. AB - Uncertainty exists concerning the key factors contributing to Campylobacter colonization of poultry, especially the possible role of vertical transmission from breeder hens to young birds. A longitudinal study of Campylobacter colonization was performed in two sibling pairs of turkey flocks (four flocks total). Each pair of sibling flocks shared breeder hen populations and was obtained from the same hatchery. One flock of each pair was grown on a commercial farm, and the other was grown in an instructional demonstration unit (Teaching Animal Unit [TAU]). Flocks were located within a 60-mi (96.8-km) radius. The time of placement, feed formulations, stocking density, and general husbandry were the same for both flocks, and each flock was processed at a commercial processing plant following standard feed withdrawal and transport protocols. Both flocks grown on the commercial farms became colonized with Campylobacter between weeks 2 and 3 and remained colonized until processing. Between 80 and 90% of isolates were Campylobacter coli, and the remainder were Campylobacter jejuni. In contrast, neither C. coli nor C. jejuni were isolated from either of the TAU flocks at any time during the production cycle. None of the fla types of Campylobacter from the breeders that provided poults to one of the commercial flocks matched those from the progeny. These results failed to provide evidence for vertical transmission and indicate that this type of transmission either did not occur or was not sufficient to render the TAU turkey flocks Campylobacter positive. Management practices such as proper litter maintenance, controlled traffic between the TAU farm and other turkey flocks, and other less well-defined aspects of turkey production were likely responsible for the absence of Campylobacter in the TAU flocks before harvest. PMID- 15270503 TI - Prevalence of potentially pathogenic Vibrio species in the seafood marketed in Malaysia. AB - Seafood samples obtained in seafood markets and supermarkets at 11 sites selected from four states in Malaysia were examined for the presence of nine potentially pathogenic species from the genus Vibrio between July 1998 and June 1999. We examined 768 sample sets that included shrimp, squid, crab, cockles, and mussels. We extensively examined shrimp samples from Selangor State to determine seasonal variation of Vibrio populations. Eight potentially pathogenic Vibrio species were detected, with overall incidence in the samples at 4.6% for V. cholerae, 4.7% for V. parahaemolyticus, 6.0% for V. vulnificus, 11% for V. alginolyticus, 9.9% for V. metschnikovii, 1.3% for V. mimicus, 13% for V. damsela, 7.6% for V. fluvialis, and 52% for a combined population of all of the above. As many as eight Vibrio species were detected in shrimp and only four in squid and peel mussels. The overall percent incidence of any of the eight vibrios was highest (82%) in cockles (Anadara granosa) among the seafoods examined and was highest (100%) in Kuching, Sarawak State, and lowest (25%) in Penang, Pulau Penang State, among the sampling sites. Of 97 strains of V. cholerae isolated, one strain belonged to the O1 serotype and 14 to the O139 serotype. The results indicate that the various seafood markets in Malaysia are contaminated with potentially pathogenic Vibrio species regardless of the season and suggest that there is a need for adequate consumer protection measures. PMID- 15270504 TI - Campylobacter prevalence in lactating dairy cows in the United States. AB - The objective of the present study was to determine the prevalence of intestinal Campylobacter in lactating dairy cows from various regions of the United States. Participating commercial dairy farms were chosen at random and were part of a national survey to determine E. coli O157:H7 and Salmonella prevalence in dairy cows. Farms had no previous history of Campylobacter problems. Fecal samples were collected rectally from 720 cows on farms in the northeast (four farms), in the desert southwest (three farms), and in the Pacific west (two farms). A minimum of 60 fecal samples per visit were collected from each farm. Thirty isolates were analyzed using the RiboPrinter Microbial Characterization System to obtain ribosomal RNA patterns. Twenty isolates were tentatively identified as Campylobacter jejuni, two as Campylobacter coli, three as Campylobacter spp., and five as unknown. Individual single-visit farm prevalence ranged from 0 to 10%. The disk diffusion method, employing 11 antibiotics, was used to test the antibiotic sensitivities of 27 of the isolates. Eight isolates were resistant to two or more antibiotics, 13 isolates were resistant to one antibiotic, and 6 were totally susceptible. Under the conditions of this study, the authors conclude that Campylobacter prevalence in lactating dairy cows in the United States is low, there is no difference in prevalence on the basis of geographical location, the predominant species is C. jejuni, and that the majority of these isolates are sensitive to antibiotics. PMID- 15270505 TI - Critical control points for monitoring salmonellae reduction in thai commercial frozen broiler processing. AB - Since 1998, pathogen reduction regulations for poultry have been enforced through the Food Safety and Inspection Service of the U.S. Department of Agriculture and through hazard analysis critical control point evaluation. This enforcement has focused attention on pathogen control and sanitation in the United States and in other countries, including Thailand. The objective of this study was to evaluate reduction in salmonellae achieved by Thai commercial exporters of frozen broiler chickens. A total of 188 broiler samples and 56 water overflows from two chillers were collected from nine processing lines of frozen broiler exporters at four identified critical control points (CCPs): CCP1, washing; CCP2, chilling; CCP3, deboning; and CCP4, packing. Samples were screened for salmonellae by enzyme linked immunosorbent assay, and bacterial identification was confirmed through biochemical and serological patterns. The overall prevalence of Salmonella was 24.6% (60 of 244 samples), with 12 serovars identified. Salmonella Albany was predominant (33.3%, 20 of 60 samples). Salmonella prevalence was 20.0% (6 of 30 samples) prior to CCP1 and was 12.5% (4 of 32), 22.7% (15 of 66), 33.3% (10 of 30), and 23.3% (7 of 30) after CCP1, CCP2, CCP3, and CCP4, respectively. The critical limit was 20% positive samples, and three CCPs failed to meet standards. Three corrective interventions were used at CCP2: 30 mg/liter hydrogen peroxide, 0.5% peracetic acid, and 125 mg/liter ozone. After these interventions, 65 broiler samples were collected for analysis of Salmonella prevalence. Results were compared with those obtained after chlorine was applied individually as a control. The Salmonella prevalences after intervention treatments were 16.0% (4 of 25), 5.0% (1 of 20), and 15.0% (3 of 20) after hydrogen peroxide, peracetic acid, and ozone treatments, respectively. All values were below the 20% critical limit, and the application of 0.5% peracetic acid produced significantly lower prevalences (P < 0.05). Repeated sampling after 1 to 4 months indicated that sanitation at these three plants was inconsistent (P < 0.05). PMID- 15270506 TI - Acute infection of swine by various Salmonella serovars. AB - The objective of this study was to evaluate the ability of various serovars of Salmonella enterica subsp. enterica to infect alimentary and nonalimentary tissues of swine within 3 h of inoculation. Fourteen wild-type S. enterica serovars (4,12:imonophasic, 6,7 nonmotile, Agona, Brandenburg, Bredeney, Derby, Heidelberg, Infantis, Muenchen, Thompson, Typhimurium, Typhimurium variant Copenhagen, untypeable, and Worthington), two known virulent S. enterica serovars (Choleraesuis strain SC-38 and Typhimurium strain chi4232), and two avirulent S. enterica Choleraesuis vaccine strains (Argus and SC-54) were inoculated intranasally (approximately 5 x 10(9) cells) into swine (four animals per Salmonella isolate). Three hours after inoculation, animals were euthanized, and both alimentary tissues (tonsil, colon contents, and cecum contents) and nonalimentary tissues (mandibular lymph node, thymus, lung, liver, spleen, ileocecal lymph node, and blood) were collected for Salmonella isolation. All Salmonella serovars evaluated except Salmonella Choleraesuis SC-54 acutely infected both alimentary and nonalimentary tissues. These results indicate that Salmonella isolates commonly found in swine are capable of acutely infecting both alimentary and nonalimentary tissues in a time frame consistent with that in which animals are transported and held in lairage prior to slaughter. PMID- 15270507 TI - Effect of short-term lairage on the prevalence of Salmonella enterica in cull sows. AB - This study was designed to compare Salmonella enterica prevalence in sows held in a holding pen at the abattoir for approximately 2 h (hold sows) with sows slaughtered immediately after transport to the abattoir (no-hold sows). Cull sows (n = 160) were sampled from four sampling periods over 8 weeks (February to March 2002) at the abattoir. Sows originated from an integrated swine farm and were sent to a live-hog market and then to the slaughter facility. Before testing, sows entered the abattoir pen and four 100-cm2 four-ply gauze squares were placed randomly on the pen floor for S. enterica culture. Sows were alternatively assigned to the hold or no-hold group. Samples collected from sows during slaughter were ileocecal lymph node, cecal contents, transverse colon contents, subiliac lymph node, sponge swabs of the left and right carcass section (300 cm2), and chopped meat. Overall, S. enterica was isolated from 44% (35 of 80) of the no-hold sows, which was significantly less (P < 0.05) than 59% (47 of 80) of the held sows. Also, no-hold sows had a lower cecal content prevalence (39%, 31 of 80) compared with that (55%, 44 of 80) of held sows (P < 0.05). S. enterica serovars isolated from no-hold sows were Brandenburg (n = 16), Derby (n = 12), Hadar (n = 8), Infantis (n = 6), Johannesburg (n = 3), 6,7:z10-monophasic (n = 3), and Typhimurium (n = 1). S. enterica serovars isolated from held sows (n = 61 isolates) were Derby (n = 19), 6,7: z10-monophasic (n = 15), Brandenburg (n = 10), Infantis (n = 6), Hadar (n = 5), Johannesburg (n = 4), and Tennessee (n = 2). Serovars recovered from the pen were Reading (n = 6), Derby (n = 4), Uganda (n = 2), and Manhattan (n = 2). Results of this study suggest that holding pens contribute to increased S. enterica carriage in cull sows. Abattoir holding pens might be an important control point for S. enterica in the ground pork production chain. PMID- 15270508 TI - Determination of the principal points of product contamination during beef carcass dressing processes in Northern Ireland. AB - To determine the principal points of microbial contamination of carcasses during beef carcass dressing in Northern Ireland, 190 carcasses were sampled by swabbing 1,000 cm2 of the brisket. A detailed survey of one abattoir was initially conducted, with sampling of a total of 100 carcasses immediately after hide removal (H), after carcass splitting (S), and immediately after washing (W) before dispatch to the chiller. The total bacterial counts after incubation at both 22 and 37 degrees C indicated that there was no significant increase in the numbers of bacteria after the first sampling point, H (P > 0.05). To determine whether this was the case in the majority of Northern Ireland abattoirs, 15 carcasses were then sampled at each of an additional six abattoirs, at points H and W only. Total bacterial counts were significantly higher (P < 0.05) at H than at W, indicating that hide pulling was the major point of bacterial contamination of beef carcasses and hence a critical control point for the final microbiological quality of the carcasses. Mean counts of Enterobacteriaceae at both incubation temperatures were very low (< 10 CFU/cm2) but were higher at W than at H, probably indicating that washing was redistributing bacteria from the posterior to the anterior region. PMID- 15270509 TI - Survival of Escherichia coli O157:H7, Salmonella enteritidis, Staphylococcus aureus, and Listeria monocytogenes in Kimchi. AB - The survival of gram-positive and gram-negative foodborne pathogens in both commercial and laboratory-prepared kimchi (a traditional fermented food widely consumed in Japan) was investigated. It was found that Escherichia coli O157:H7, Salmonella Enteritidis, Staphylococcus aureus, and Listeria monocytogenes could survive in both commercial and laboratory-prepared kimchi inoculated with these pathogens and incubated at 10 degrees C for 7 days. However, when incubation was prolonged, the S. aureus level decreased rapidly from the initial inoculum level to the minimum detectable level within 12 days, whereas Salmonella Enteritidis and L. monocytogenes took 16 days to reach similar levels in commercial kimchi. On the other hand, E. coli O157:H7 remained at high levels throughout the incubation period. For laboratory-prepared kimchi, the S. aureus level decreased rapidly from the initial inoculum level to the minimum detectable level within 12 days, and L. monocytogenes took 20 days to reach a similar level. E. coli O157:H7 and Salmonella Enteritidis remained at high levels throughout the incubation period. The results of this study suggest that the contamination of kimchi with E. coli O157:H7, Salmonella Enteritidis, S. aureus, or L. monocytogenes at any stage of production or marketing could pose a potential risk. PMID- 15270510 TI - Control of Escherichia coli O157:H7 with sodium metasilicate. AB - Three intervention strategies-trisodium phosphate, lactic acid, and sodium metasilicate--were examined for their in vitro antimicrobial activities in water at room temperature against a three-strain cocktail of Escherichia coli O157:H7 and a three-strain cocktail of "generic" E. coli. Both initial inhibition and recovery of injured cells were monitored. When 3.0% (wt/wt) lactic acid, pH 2.4, was inoculated with E. coli O157:H7 (approximately 6 log CFU/ml), viable microorganisms were recovered after a 20-min exposure to the acid. After 20 min in 1.0% (wt/wt) trisodium phosphate, pH 12.0, no viable E. coli O157:H7 microorganisms were detected. Exposure of E. coli O157:H7 to sodium metasilicate (5 to 10 s) at concentrations as low as 0.6%, pH 12.1, resulted in 100% inhibition with no recoverable E. coli O157:H7. No difference in inhibition profiles was detected between the E. coli O157:H7 and generic strains, suggesting that nonpathogenic strains may be used for in-plant sodium metasilicate studies. PMID- 15270511 TI - The BAX PCR assay for screening Listeria monocytogenes targets a partial putative gene lmo2234. AB - The BAX PCR for screening Listeria monocytogenes is a commercial PCR assay for specifically targeting L. monocytogenes, a foodborne pathogen that can contaminate a variety of foods and cause a potentially fatal disease, listeriosis, among high-risk populations. The high specificity (> 98%) of this PCR assay is achieved by targeting a species-specific genomic region (approximately 400 bp) presumably found only in L. monocytogenes. In this study, the identity of the BAX PCR-targeted genomic region was determined by using PCR cloning, DNA sequencing, and basic local alignment search tool (BLAST) analysis of the amplicon sequences of an L. monocytogenes serotype 1/2a strain. BLAST analysis identified the BAX PCR amplicon (GenBank accession no. AY364605) as a 423-bp genomic region between nucleotides 224,409 and 224,831 in the genome of L. monocytogenes (serotype 1/2a strain EGD-e), including a 145-bp noncoding region and a 278-bp partial coding sequence of a putative gene, lmo2234. The translated amino acid sequence (92 amino acids) of this partial coding region is highly conserved between L. monocytogenes and Listeria innocua (93% homology). Reverse position-specific BLAST analysis identified a conserved domain in Lmo2234 that was similar (95.3% aligned, E value = 9E-18) to the consensus amino acid sequence of sugar phosphate isomerases/epimerases (National Center for Biotechnology Information conserved domain database accession no. COG 1082.1, IolE), indicating that Lmo2234 might be involved in bacterial carbohydrate transport and metabolism. PMID- 15270512 TI - Evaluation of Staphylococcus aureus growth potential in ham during a slow-cooking process: use of predictions derived from the U.S. Department of Agriculture Pathogen Modeling Program 6.1 predictive model and an inoculation study. AB - The U.S. Department of Agriculture has cautioned against slow cooking meat such that the interior temperature increases from 10 degrees C (50 degrees F) to 54.4 degrees C (130 degrees F) in > or = 6 h. During a commercial ham-smoking process, the ham cold point is typically between 10 and 54.4 degrees C for 13 h, but the ham is subsequently exposed to heating sufficient to eliminate vegetative pathogenic bacteria. Thus, production of heat-stable staphylococcal enterotoxin is the primary biological hazard. For this study, uncooked surface and uncooked ground interior ham were inoculated with a three-strain Staphylococcus aureus mixture, exposed to simulated surface and interior slow-cook conditions, respectively, and analyzed periodically using the Baird-Parker agar and 3M Petrifilm Staph Express count plate methods. For the surface and interior conditions, S. aureus numbers increased by no more than 0.1 and 0.7 log units, respectively. Predictions derived from actual time and temperature data and S. aureus growth values from a computer-generated model (Pathogen Modeling Program 6.1, U.S. Department of Agriculture) were for 2.7 (ham surface) and 9.9 to 10.5 (ham interior) generations of S. aureus growth, indicating that use of model derived growth values would not falsely indicate safe slow cooking of ham. The Baird-Parker method recovered significantly (P < 0.05) greater numbers of S. aureus than the Petrifilm Staph Express method. For hams pumped with brine to attain (i) 18% (wt/wt) weight gain, (ii) > or = 2.3% sodium lactate, (iii) > or = 0.8% sodium chloride, and (iv) 200 ppm ingoing sodium nitrite, slow-cooking critical limits of < or = 4 h between 10 and 34 degrees C, < or = 5 h between 34 and 46 degrees C, and < or = 5 h between 46 and 54.4 degrees C could be considered adequate to ensure safety. PMID- 15270513 TI - Biocontrol of psychrotrophic enterotoxigenic Bacillus cereus in a nonfat hard cheese by an enterococcal strain-producing enterocin AS-48. AB - Bacillus cereus is a food poisoning bacterium of great concern, especially in milk products. In this study, we describe the efficient control of the psychrotrophic and toxigenic strain B. cereus LWL1 in milk and in a nonfat hard cow's cheese by the enterocin AS-48 producer strain Enterococcus faecalis A-48-32 (Bac+). No viable B. cereus cells were detected after 72 h incubation in milk coinoculated with the AS-48-producing strain and B. cereus. Diarrheic toxin production was also markedly inhibited by the Bac+ strain to eightfold lower levels compared with control cultures of B. cereus. In cheeses manufactured by inoculation with a commercial starter (about 6.8 log CFU/ml) and B. cereus (about 4 log CFU/ml), the latter reached 6.27 log CFU/g after 5 days of maturation, and approximately 8 log CFU/g after 15 days. However, in cheeses made from milk inoculated with the starter along with a mixture of E. faecalis-B. cereus (2/1 ratio), counts of B. cereus decreased by approximately 1.0, 2.0, 4.32, and 5.6 log units with respect to control cheeses after 5, 10, 15, and 30 days of ripening, respectively. Growth of E. faecalis A-48-32 was associated with enterocin AS-48 production and persistence in cheese. Interestingly, growth of starter cultures was not affected by the Bac+ strain, and neither was lactic acid production. These results clearly indicate that E. faecalis A-48-32 produced satisfactory amounts of bacteriocin in cheese and support the potential use of AS 48-producing strains as culture adjuncts to inhibit B. cereus during cheese manufacture and ripening. PMID- 15270514 TI - Compounds of parasitic roundworm absorbing in the visible region: target molecules for detection of roundworm in Atlantic cod. AB - The objective of this study was to contribute to the development of technology that will be able to replace manual operations in processing of fish fillets. Removal of parasites, black lining, remnants of skin, and bloodstains are costly and time-consuming operations to the fish processing industry. The presence of parasites in fish products tends to spoil consumers' appetites. Recent reports questioning the safety of eating cod infected with parasites might lower consumer acceptance of seafood. Presently, parasites are detected and removed manually. An average efficiency of about 75% under commercial conditions has been reported. In this study, we focused on biochemical differences between cod muscle and the prevalent anisakine nematode species (Anisakis simplex and Pseudoterranova decipiens) infecting Atlantic cod (Gadus morhua). Using reversed phase high performance liquid chromatography equipped with a photodiode array detector, substances absorbing in the range 300 to 600 nm were identified in extracts from parasite material. These substances were not detected in extracts from cod tissue. Significant biochemical differences between cod muscle and parasite material have thus been demonstrated. PMID- 15270515 TI - Content of toxic and essential metals in canned mussels commonly consumed in Tenerife, Canary Islands, Spain. AB - Concentrations of three toxic heavy metals (Hg, Pb, Cd) and six essential heavy metals (Fe, Zn, Mn, Cr, Cu, Ni) were determined in mussel conserves (Mytilus galloprovincialis, Bivalvia, Mollusca) consumed habitually by individuals in Tenerife (Canary Islands, Spain). A total of 600 samples were analyzed, corresponding to six different commercial brands and four different processing types: pickled sauce (mixture of olive oil, vinegar, red pepper, laurel, and salt), coquille St. Jacques sauce (coquille St. Jacques broth), nature (water and salt), and bionature (water, salt, and soluble vegetal fiber). Samples were collected weekly from markets in Santa Cruz de Tenerife during a 12-month period. All values for toxic metals were lower than the permitted maximum for human consumption as proscribed in European Community Directive 2001/22/CE (1,000 microg/kg wet weight for Pb and Cd) and European Community Decision 93/351/EEC (500 microg/kg wet weight for Hg). For the six essential heavy metals, mussels are a very good source, contributing high percentages of the recommended daily allowance. PMID- 15270516 TI - Modifications and adaptations of the Charm II rapid antibody assay for chloramphenicol in honey. AB - The Charm II screening method for the presence of chloramphenicol in honey has a sensitivity of 0.3 ppb. This screening method is a simple, rapid antibody assay using [3H]chloramphenicol and a binding reagent. Analysis of different types of honey revealed considerable differences in results. Honey can be liquid, crystallized (creamed), or partially crystallized and is classified by the U.S. Department of Agriculture into seven color categories: water white, extra white, white, extra light amber, light amber, amber, and dark amber. Fortified and nonfortified liquid amber honey tested appropriately with the Charm II unit and the negative control provided with the unit after slight modifications were made. However, approximately 70% of creamed honey samples fortified at 0.6 ppb did not test positive for the presence of chloramphenicol using the provided negative control. Matrix quenching effects were evaluated, and these effects were accounted for by establishing different assay conditions for different honey types. PMID- 15270517 TI - Growth potential of Clostridium perfringens during cooling of cooked meats. AB - Many meat-based food products are cooked to temperatures sufficient to inactivate vegetative cells of Clostridium perfringens, but spores of this bacterium can survive, germinate, and grow in these products if sufficient time, temperature, and other variables exist. Because ingestion of large numbers of vegetative cells can lead to concomitant sporulation, enterotoxin release in the gastrointestinal tract, and diarrhea-like illness, a necessary food safety objective is to ensure that not more than acceptable levels of C. perfringens are in finished products. As cooked meat items cool they will pass through the growth temperature range of C. perfringens (50 to 15 degrees C). Therefore, an important step in determining the likely level of C. perfringens in the final product is the estimation of growth of the pathogen during cooling of the cooked product. Numerous studies exist dealing with just such estimations, yet consensual methodologies, results, and conclusions are lacking. There is a need to consider the bulk of C. perfringens work relating to cooling of cooked meat-based products and attempt to move toward a better understanding of the true growth potential of the organism. This review attempts to summarize observations made by researchers and highlight variations in experimental approach as possible explanations for different outcomes. An attempt is also made here to identify and justify optimal procedures for conducting C. perfringens growth estimation in meat-based cooked food products during cooling. PMID- 15270518 TI - [Surgical treatment of colorectal liver metastasis in the year 2000]. AB - There is significant recent progress in the surgical and oncological treatment of colorectal liver metastases. In our review we try to explain the variability and the multidisciplinarity approach we use to treat our patients more successful after the year of 2000. Beside the old rule: "Resect if possible"--thanks to the improvement of chemotherapy and new surgical instruments- there are new trends: "Make it resectable" or "resect it as many times as possible" thanks to the loco regional and systemic neoadjuvant chemotherapy and/or the repeated interventions in metastasis surgery. The most important prognostic factors influencing the overall survival rate are free surgical margins, synchronous or metachronous metastasis and the need for repeated interventions. The view about extra hepatic metastases of CRC is changed in the last decade as the resection of these metastases is also suggested. After the year 2000 colorectal liver metastases should be treated "a-la-carte", based on multidisciplinary planning and treatment in specialised centres providing the long survival rates. PMID- 15270519 TI - [Changing trends in breast surgery over a five year period (1998-2002)--a retrospective analysis]. AB - Today's development in breast cancer surgery are characterized by the principle of the smallest necessary intervention in contrast with the radicality of the past. To achieve this goal the primary and crucial task is the recognition of early (stage I or II) breast cancer. The National Screening Program started 2002 provides ideal conditions. The authors present a five year (1998-2002) retrospective analysis of breast preserving surgery: over the period 861 operations were performed on breast cancer patients with an average of 46.5% of them with breast conservation. Complications after unnecessary axillary lymph node dissection occurred with a high incidence rate (60%); the authors suggest sentinel node identification and detection performed to avoid these complications. In the "early years" (1998-2001) only 15 preoperative wire-loop markings were performed in patients with non-palpable malignant lesions, whereas from 2002--owing to the National Screening Program--68 such interventions were carried out providing the immense importance of nationwide screening. PMID- 15270520 TI - [The importance of assessing the "quality of life" in surgical interventions for critical lower limb ischaemia]. AB - 'Patency' and 'limb salvage' are not automatically valid parameters when the functional outcome of treatment for critical limb ischaemia is assessed. In a small number of patients the functional result is not favourable despite the anatomical patency and limb salvage. The considerable investment of human/financial resources in the treatment of these patients is retrospectively questionable in such cases. Quality of Life questionnaires give valuable information on the functional outcome of any means of treatment for critical ischaemia. The problem with the generic tools in one particular sub-group of patients is the reliability and validity of the tests. The first disease-specific test in critical limb ischaemia is the King's College Vascular Quality of Life (VascuQoL) Questionnaire. Its use is recommended in patients with critical lower limb ischaemia. It is very useful for scientific reporting and is able to show retrospectively that particular group of patients in whom the technical success of the treatment did not result in improvement in quality of life. In general practice the use of the questionnaire can decrease the factor of subjectivity in the assessment of the current status of a patient with newly diagnosed or previously treated critical ischaemia. PMID- 15270521 TI - [Alternative options for examination of the patency of peritoneo-venous shunts]. AB - For the treatment of refractory ascites we use the saphenoperitoneal shunt described by Pang in 1992 approximately 2 years. This procedure eliminates the most frequent complications of the former synthetic shunts: occlusion of the collector branches and infections as well. In addition, the use of autologous vein is cost-saving. The first Hungarian publications (K. Vincze and Z. Nagy et al.) reported good results, which are confirmed also by us, after we performed 21 operations. The publications until now usually describe the technique. This intervention is now a widely accepted one. On the other hand, just a small number of papers describe the options for the examination of patency and the follow-ups. We report about the algorithm used in our department after surgery to evaluate graft patency and surgical efficacy. A method to determine the volume of ascites developed by ourselves is described. We feel that the successful application of saphenoperitoneal shunts depends on very close follow-up. Considering that no objective method to check the patency does exist, we are sure that decisions about further operations can only be made if simultaneous diverse follow-up methods are available. PMID- 15270522 TI - [Limb salvage by excision of calcaneus in diabetic atherosclerotic gangrene]. AB - We treated a 49-year-old female patient, who developed diabetic, arteriosclerotic gangrene. Only after the extensive wet gangrene extended to the proximal half sole, she was finally consenting to surgery, and a femoropopliteal Dacron bypass graft was performed. The area of the osteomyelitic calcaneus was removed by necrectomy (slough cutting) but the calcaneus fractured spontaneously after two weeks. Therefore the calcaneus was excised and the half-sole defect was covered by the available skin of the posterior heel and forefoot region. The wound required meticulous local treatment for two months. She has been able to walk without a frame using an orthopedic shoe fitted with a total contact insole with proper lifting of the heel region. Our aim was limb saving and recovery favourable to crural amputation, which could have been justified in such extensive gangrene. We would like to highlight to the possibility of the excision of the calcaneus, as no data has been found about it in the Hungarian literature. Walking after the excision of the calcaneus is possible with the help of a specially developed orthopaedic shoe, properly fitted with a total contact insole. PMID- 15270523 TI - [In memory of Gyorgy Frank (1910-1959)]. PMID- 15270524 TI - [The moral dilemmas of palliative surgery]. PMID- 15270525 TI - [Lichtenstein's hernioplasty]. AB - In recent years in Hungary similarly to the worldwide trend the treatment of inguinal hernias has changed. New tension-free methods were developed and--after the introduction by the Department of Surgery, Medical Faculty, University of Pecs--Lichtenstein's method is widely used. Lichtenstein's method has become the gold standard at our department because of its highly favourable results: simple technique, minimal postoperative pain, recurrence rate below 1%, short hospital stay, very low complication rate and early return to physical activity. PMID- 15270526 TI - [Current issues in the surgical treatment of rectal cancer]. AB - INTRODUCTION: Three topics associated with surgical treatment of rectal cancer are discussed. The need for standard preoperative irradiation, based on interdisciplinary approach is considered. In the authors' opinion the most urgent task is to increase the number of sphincter saving resections carried out by total mesorectal excision. The principles of surgical therapy for hepatic metastases are discussed. METHODS: The authors' experience is based on 328 radical resections performed between 1992-2003. RESULTS: Three periods are compared: we managed to raise the ratio of surgical interventions preserving the rectal sphincter over 60%, and during the last period the results have further improved. Total mesorectal excision is the standard in the department. The morbidity rate--28.8%, 18.6% and 5.3% for suture leak is reduced. During the last eight years in the presence of hepatic metastasis in 20 liver resections were performed. CONCLUSIONS: In addition to preoperative irradiation considered we recommend treatment consisting of 5x5 Gy for early cancer, and of 60 Gy/6 weeks for a progressed stage. In our opinion, application of atraumatic surgical techniques and up to date circular stapling guns can increase the ratio of sphincter preserving interventions. We hope, the use of total mesorectal excision carried out adequately may reduce morbidity (suture leak, nerve damage--impaired innervation and sexual dysfunction) and the rate of local recurrence and it may improve survival. The only curative treatment of hepatic metastases in colorectal cancers is resection, therefore the authors suggest the possibility of widening the range of surgical indications--more than three metastases in one lobe, even in bilateral involvement. PMID- 15270527 TI - [Laparoscopic exploration of the common bile duct]. AB - Since the first laparoscopic common bile duct exploration in Hungary published in 1999 the authors use the technique themselves. We review and analyse our activity between 1 June 1999 and 31 August 2003. The minimally invasive approach was selected in twelve patients with obstructive jaundice for suspected bile duct stones. Eight of these patients underwent preoperative endoscopy, but either the number and/or size of stones or various complications prevented successful endoscopic stone extraction; five sphincterotomies--two followed by stenting- were performed. The four other patients did not consent to endoscopy. During surgery the biliary tract was visualised by choledochoscope (a bile duct endoscope with video connection) in four cases via the distended cystic duct and in eight cases via longitudinal choledochotomy. In one case there was no occlusion at all, in another one dilatation was performed because of a structure. In nine patients stones were removed either through the choledochotomy/cystic duct or by passing them into the duodenum. In one patient we converted to open procedure due to a stone impacted in the papilla of Vater. After choledochotomy intracorporal suturing and knot tying techniques were used to close the incision. On six occasions a cystic drain, three occasions a T-tube and on three occasions primary closure was used. Two bile leaks were treated by endoscopic stenting and in one patient a laparotomy was needed. There was no mortality. We believe that laparoscopic common bile duct exploration can be successful even in complicated situations. PMID- 15270528 TI - [Thymic neuroendocrine tumor: three cases in our practice]. AB - Authors present 3 cases of the thymic neuroendocrine tumor. They describe the diagnosis, the TNM classification, histology, therapy, and prognosis of the disease. In their opinion the TNM and histology classification have adequate prognostic value. PMID- 15270529 TI - [Difficulties in the treatment of polycystic liver]. AB - A 52-year-old female patient was admitted to our department suffering from discomfort and tension in right side subcostal region for five months. The anamnesis contains cholecystectomy, appendectomy and gynecology treatment. CT examination and X-ray examination showed cysts in both lobe of liver. The primary treatment was ultrasonography guided punction in another department. This treatment caused anaphylactoid shock. After this dangerous treatment the patient refused the next punction. Following required arrangement laparoscopic exploration and adhaesiolysis were done in our department. Different size cysts had laparoscopically fenestrated. After half year the patient's symptoms resumed. Repeated CT and US examinations showed cysts again. Cysts were laparoscopically fenestrated again. Since the operation the patient had no complaint. Histology showed fibrocystic liver. After a month control CT examination showed cysts again. After two months we made ultrasonography guide punction of remain cysts following radiological consultation. PMID- 15270530 TI - [Unusual laparoscopic surgical cases: cholelithiasis in situs inversus totalis, and gallbladder agenesis]. AB - Organ anomalies and organ system transposition may cause diagnostic and therapeutic difficulties. We report a patient with situs inversus totalis and symptomatic cholelithiasis successfully treated via laparoscopic cholecystectomy. We present a laparoscopic and MR cholangiographic pictures of our patient with gallbladder agenesis. PMID- 15270531 TI - [Vertebral metastasis from malignant degeneration of urethro-perineal fistula. A case report]. AB - Adenocarcinoma mucinusum of the anal canal is a rare tumour, which may appear as a fistula. The fistula has an effect on quality of life and besides it may lead to tumour formation. Early recognition and regular check-up are important, and early operation is recommended. Early surgical excision combined with chemo- and radiotherapy may be effective, with 5 year recurrence rate lower than 37%. The prognosis of late treatment is poor with 1 year average survival. Our case report presents evidence that the key to successful therapy and improved quality of life is early diagnosis and combined surgical-oncological treatment. PMID- 15270532 TI - [National vascular surgery statistics, Hungarian Vascular Surgery Institute, 2003]. PMID- 15270533 TI - [Imre Ullmann (1861-1937). The first organ transplantation...]. PMID- 15270534 TI - [Report from the 12th Congress of the European Society of Surgical Oncology (ESSO) and about the European examination in the specialty of oncologic surgery]. PMID- 15270535 TI - Safety net sagging under multiple stresses. PMID- 15270536 TI - Insecticide susceptibility level of Anopheles arabiensis in two agrodevelopment localities in eastern Ethiopia. AB - Anopheles arabiensis strains reared from larvae and pupae collected from two different localities, Metehara and Melka-Worer, eastern Ethiopia, were evaluated against three insecticides. Resistance states of adult females were determined using the WHO test kits under field condition. The insecticides used were WHO discriminating doses of 4% DDT, 0.75% permethrin and 0.1% propoxur. The study revealed that 42.5% of the An. arabiensis population was resistant to DDT in Melka-Worer while only 30% of the species was resistant in Metehara to the same insecticide. In Metehara, 25% of An. arabiensis was also resistant to permethrin while the population was highly susceptible to the insecticide in Melka-Worer. Propoxur, which was not evaluated in Melka-Worer, was highly toxic to An. arabiensis in Metehara. The knocked-down time (KD50) for permethrin was 14.5 and 12.5 minutes in Metehara and Melka-Worer, respectively. The implications of these findings are discussed. PMID- 15270537 TI - Clinical signs and household characteristics associated with human fascioliasis among rural population in Egypt: a case-control study. AB - The symptomatology associated with human fascioliasis has been studied mostly in hospitalised subjects. Very little is known about clinical signs and symptoms associated with infections in human endemic zones, as well as on possible new ways of transmission which might be responsible for the increased number of human cases reported in recent years. This information is of great importance to facilitate diagnosis and plan effective control measures. With the objective to identify clinical signs, blood tests, household characteristics and hygienic habits associated with human fascioliasis, a cross-sectional case-control study was implemented among the rural population of three endemic foci in the Nile Delta, Egypt. Clinical history was collected from 53 cases of fascioliasis and the same number of individually matched controls. They received a complete clinical examination and a range of blood tests was performed on them. Information on socio-economic conditions, dietary and hygienic habits was also collected. The most important complaints, associated with the infection, were right abdominal pain (Odds Ratio 20, P = 0.005), epigastric burning (o.r. 16, P = 0.007) and nausea (o.r. 8, P = 0.05). Blood analyses reported a marked increase in blood eosinophils (o.r 1.3, P = 0.001) among cases. The presence of cows (o.r 3.2), buffaloes (o.r 3.0) and goats (o.r 2.6) in the household was closely associated with the infection together with the habit to bring those animals to the canal for bathing and/or drinking (o.r. 3.2). Among dietary habits investigated, eating raw seeds was more common in cases than controls (o.r. 9, P = 0.03) and emerged as a possible new way of infection. PMID- 15270538 TI - dnaA gene sequences from Wolbachia pipientis support subdivision into supergroups and provide no evidence for recombination in the lineages infecting nematodes. AB - Wolbachia pipientis is an intracellular bacterial endosymbiont of arthropods and filarial nematodes. Six main supergroups of W. pipientis have been described: supergroups A, B, E, and F encompass arthropod wolbachiae; supergroups C and D encompass nematode wolbachiae. The description of these six supergroups has been based on the analysis of only two genes (ftsZ and 16S rDNA) and before decisions are taken on the taxonomic status of the six supergroups, analysis of further genes is required. In addition, the branching order of the six supergroups is still unresolved. Sequence information from other genes is also needed to allow phylogenesis to be addressed through the analysis of a higher number of characters. Here we report sequences from a portion of the gene coding for the DNAA protein of W pipientis, generated from the endosymbionts of 22 host species. Phylogenies based on dnaA gene sequences are congruent with the existence of at least six supergroups of W pipientis. In addition, subtrees generated for nematode wolbachiae in supergroups C and D were compared to the trees based on the already available gene sequences (ftsZ, 16S rDNA and wsp). The congruence observed among the trees based on the different genes agrees with the hypothesis that recombination does not occur in nematode wolbachiae. PMID- 15270539 TI - [Dientamoeba fragilis: is it really fragile? Approach to specimen handling and rapid microscopic diagnosis]. AB - Dientamoeba fragilis is a pathogenic protozoan parasite with a world-wide distribution. Interestingly, a resistant cyst stage has not been demonstrated and it is still an unsolved problem how this parasite can survive successfully outside the human host. D. fragilis was found in 2% of approximately 2500 individuals unselected who submitted stools for parasitological examination during 2001 in Padua (Italy). The goal of this study was to detect the protozoan stages and the duration of persistence of this protozoa in faeces stored in different environmental conditions. The trophozoites of D. fragilis were detected up to 60 days after the collection of the faeces stored at 4 degrees C and Giemsa stained. The laboratory detection rate of the organism is greatly enhanced by use of preservative to fix stool specimens immediately after passage. Alternatively, a microscopic observation of the collected stool has to be performed immediately after passage followed by examination of permanently-stained smears. Demonstration of the charateristic "golf-club" and "acanthopodia-like" structures in unstained fixed faecal material by direct microscopy (400x) are suitable for a rapid identification of D. fragilis. PMID- 15270540 TI - Ovicidal and larvicidal activity against Aedes aegypti and Anopheles gambiae complex mosquitoes of essential oils extracted from three spontaneous plants of Burkina Faso. AB - Essential oils extracted from dried leaves of three spontaneous plants naturally growing in Burkina Faso, i.e. Cymbopogon proximus, Lippia multiflora and Ocimum canum, exhibited larvicidal activity by the WHO standard protocol against 3rd and 4th instar F1-larvae of field-collected mosquitoes vectors of human disease, namely Aedes aegypti and members of the Anopheles gambiae complex, An. arabiensis and An. gambiae. The median lethal concentration (LC50) for Ae. aegypti and An. gambiae s.l. larvae ranged between 53.5-258.5 ppm and 61.9-301.6 ppm, respectively. The LC90 estimates ranged 74.8-334.8 ppm for Ae. aegypti, and 121.6 582.9 ppm for An. gambiae s.l. Ovicidal activity against eggs of An. gambiae s.l. was also demonstrated. The LC50 values for An. gambiae s.l. eggs ranged between 17.1-188.7 ppm, while LC90 values ranged between 33.5-488 ppm. Lippia multiflora showed the highest activity against An. gambiae s.l. eggs and Ae. aegypti larvae, whereas no difference was found among C. proximus and L. multiflora in their activity against An. gambiae s.l. larvae. Of the three plants, essential oils from O. canum had the lowest activity against both eggs and larvae. Eggs were more susceptible than larvae. Ae. aegypti larvae were more susceptible than larvae of An. gambiae s.l. PMID- 15270541 TI - Malaria: use of restriction endonuclease digestion and mutation-specific PCR for antifolate resistance isolate detection. AB - Antifolate resistance isolates of Plasmodium falciparum in the blood of 56 patients was investigated by using PCR technology. DNA was extracted with three different methods from parasite lysate by phenol-chloroform, or from whole blood and from blood collected onto dry filter paper, by chelex-100. The expected 727 bp PCR product was obtained in all samples extracted by chelex-100, while three samples prepared by phenol-chloroform failed to show any amplified product. The crucial point mutation within the dhfr gene leading to pyrimethamine and cycloguanil resistance is localised in an Alul recognition site. Thus, the 727-bp PCR product was submitted to endonuclease digestion. Fifty out of the 56 blood samples analysed yielded the two expected restriction fragments and an undigested 727-bp band. These 50 samples likely represent mixed infection as also confirmed the specific mutation PCR. The six undigested samples amplify a 339-bp fragment using a nested PCR-specific for pyrimethamine resistance mutation. Our results show that, the rapid DNA extraction from blood using chelex-100 and the PCR endonuclease assay can be efficiently used for accurate chemosensitivity analysis in the field. PMID- 15270542 TI - An annotated checklist of Hymenolepidid cestodes described from birds: 1983-2002. AB - A survey of the literature covering the period from January 1983 to December 2002 revealed that 17 genera and 106 species of cestodes belonging to the family Hymenolepididae have been described from birds since the publication of Schmidt's Handbook of Tapeworm Identification (his literature search terminated in March, 1983). Although the family has been revised recently, Schmidt's monograph provides the most recent list of species within the family. This work updates the list of species reported from avian hosts and includes data on the host, and collection locality along with the dimensions of the strobila, the number and length of the rostellar hooks, where applicable, and the dimensions of the cirrus sac as reported in original descriptions. PMID- 15270543 TI - Editorial introduction to the paper by A. Castellani "Researches on the Etiology of Sleeping Sickness" reprinted from the Journal of Tropical Medicine, June 1, 1903. AB - The paper by Professor Aldo Castellani represents the first report suggesting the causative role of Trypanosoma spp. in the pathogenesis of sleeping sickness and confirms his pivotal role in solving the etiologic dilemma of the disease. Professor Castellani also contributed to a large extent to many other important microbiological discoveries, mainly in the field of mycology, protozoology and bacteriology. PMID- 15270544 TI - Analysis and test of laws for backward (metacontrast) masking. AB - In backward visual masking, it is common to find that the mask has its biggest effect when it follows the target by several tens of milliseconds. Research in the 1960s and 1970s suggested that masking effects were best characterized by the stimulus onset asynchrony (SOA) between the target and mask. In particular, one claim has been that the SOA for which masking is optimal remains fixed, even as target and mask durations varied. Experimental evidence supported this claim, and it was accepted as an SOA law. However, recent modeling (Francis, 1997) and experimental studies (Macknik and Livingstone, 1998) argued for new ISI (interstimulus interval) and STA (stimulus termination asynchrony) laws, respectively. This paper reports a mathematical analysis and experimental tests of the laws. The mathematical analysis demonstrates unsuspected relationships between the laws. The experiments test the predictions of the SOA, ISI, and STA laws. The data favor the ISI law over the SOA and the STA laws. PMID- 15270545 TI - Configuration effects on texture transparency. AB - This study examined the factors producing the perception of transparency between overlaid regions composed of Gabor micro-patterns as functions of their spatial frequency, separation of overlaid regions, and types of orientation modulation. The results showed that the likelihood of perceiving transparency was high both when (1) the difference in Gabor spatial frequency between regions was large, and (2) the region boundary, which was formed by short-range orientation differences in the Gabor micro-patterns, clearly emerged. We conclude that texture transparency appears to result from an interaction between a boundary-detection mechanism defining the shape of each region and a surface-detection mechanism assigning the boundary. PMID- 15270546 TI - Factors influencing the ability to detect motion reversals in rotation simulations. AB - Five experiments probed the conditions under which observers fail to report instantaneous reversals in the direction of motion of pixels that define the rotation of a transparent sphere or plane. Our results showed that the extent to which rotation reversals were not reported depended upon whether observers used strict or lax criteria to make their judgments, the degree of perspective present in the rotation simulations, and the percentage of pixels that actually reversed direction. Furthermore, we found failures to report rotation reversals both with stimuli whose pixels were confined to smooth surfaces and scattered within volumes. Reversal detection with planar stimuli, unlike sphere stimuli, depended upon the orientation of the stimulus at the moment of reversal. Treue et al. (1995) postulated a surface-interpolation process as the explanation for the apparent insensitivity of observers to such reversals. However, we suggest that other stages of processing (e.g. a structure-from-motion process) are required to account for these results. PMID- 15270547 TI - When motion is not perceived: evidence from adaptation and dynamical stability. AB - Adaptation was used to probe the perceiver's activation state when either motion or nonmotion percepts are formed for bistable, single-element apparent motion stimuli. Although adaptation was not observed in every instance, when it was observed its effect was to increase the probability of both motion-to-nonmotion and nonmotion-to-motion switches, the time scale of adaptation corresponding to neurophysiological observations for directionally selective cortical cells (Giaschi et al. 1993). This susceptibility to de-stabilizing adaptation effects indicated that the nonmotion percept was not the result of inadequate stimulation producing subthreshold levels of motion detector activation; if that were the case, activation-dependent adaptation would have decreased the nonmotion-to motion switching rate by reducing activation further below threshold. Above threshold activation levels are therefore associated with both nonmotion and motion perceptual states, and the failure to perceive motion despite the presence of adequate motion detector stimulation can be attributed to inhibitory competition between detectors activated by motion-specifying stimulus information and detectors activated to similar levels by motion-independent stimulus information, consistent with the dynamical quality of single-element apparent motion. PMID- 15270548 TI - In vitro maturation rates of canine oocytes from anoestrous bitches in simple media. AB - The meiotic competence of canine oocytes collected from anoestrous bitch ovaries and cultured for 72 h in different media was studied. The base culture medium was TCM 199 enriched with 10% fetal bovine serum (TCM); the effect of supplementation with EGF (50 ng x mL(-1)) or ITS (insulin: 10 microg x mL(-1); transferrin: 5.5 microg x mL(-1); selenium: 5 microg x mL(-1)) was also studied. TCM was also compared to a Synthetic Oviductal Fluid (SOF). All the media contained FSH (0.1 UI x mL(-1)), LH (10 UI x mL(-1)), 17beta-oestradiol (4 microg x mL(-1)) and kanamycin. Despite the anoestrous stage of the donor bitches, resumption of meiosis occurred in a high proportion of the oocytes, (mean value 77.3%). The number of oocytes showing the 'germinal vesicle breakdown' nuclear stage was not influenced by the type of the culture medium used. ITS had a positive effect on nuclear progression to later stages (from metaphase I to metaphase II); however, this effect was not statistically significant. PMID- 15270549 TI - Crop-residue supplementation of pregnant does influences birth weight and weight gain of kids, daily milk yield but not the progesterone profile of Red Sokoto goats. AB - The parameters investigated in this study with the objective of evaluating growth, lactation and reproductive performances, included birth weight, litter size, 0-90 days gain and average daily gain of kids as well as the milk yield and progesterone profile of Red Sokoto does supplemented with crop-residue based rations during the long-dry period of the subhumid zone in Nigeria. A total of 7 treatments of 4 goats each was utilised. All treatment groups had a basal diet of Digitaria smutsii hay and natural pasture ad libitum. Ration A supplemented with the conventional concentrate was used as the positive control; rations B and C were supplemented with crop residues; and ration D without supplement was used as the negative control. Supplementation with concentrate and crop residues significantly increased (P < 0.05) the birth weight and liveweight gains of kids, but littersize was unaffected. The heaviest kids at birth (1.3-1.4 kg) were from does in treatments 1A, 2A and 2C, while does in treatments 1B, 2B, 1C and D had the lightest kids (1.07-1.18 kg). The highest gains of 53.9 g x day(-1) were recorded in treatment 2A and the least (32.4 g x day(-1)) in treatment 1B. Supplementation also significantly influenced (P < 0.01) the daily milk yield of dams over the 90-day period of the dry season. All the does had similar progesterone profiles from late gestation through parturition to early lactation irrespective of their treatment group. It was concluded that ration C fed at the 2% level is a good and affordable supplementary feed package for increased birth weight and preweaning gains in kids for meat production. PMID- 15270550 TI - In vivo oxidation of carboxyl-labelled cyclic fatty acids formed from linoleic and linolenic acids in the rat. AB - Cyclic fatty acid monomers (CFAM), which occur from the intramolecular cyclisation of linoleic and linolenic acids, are subsequently present in some edible oils and are suspected to induce metabolic disorders. One may suggest that the presence of a ring would alter the ability of the organism to oxidise these molecules. In order to test this hypothesis, we assessed the oxidative metabolism of CFAM in rats. For this purpose, rats were force-fed from 1.5 to 2.6 MBq of [1 (14)C]-linoleic acid, [1-(14)C]-linolenic acid, [1-(14)C]-CFAM-18:2 or [1-(14)C] CFAM-18:3, and 14CO2 production was monitored for 24 h. The animals were then sacrificed and the radioactivity was determined in different tissues. No significant differences in 14CO2 production were found 24 h after the administration of CFAM and their respective precursors. Our data clearly demonstrate that, at least for the first beta-oxidation cycle, CFAM are oxidised in a similar way as both essential fatty acids. PMID- 15270551 TI - Myristic acid increases delta6-desaturase activity in cultured rat hepatocytes. AB - In order to study the effects of saturated fatty acids on delta6-desaturase activity, rat hepatocytes in primary culture were incubated with lauric (C12:0), myristic (C14:0) or palmitic (C16:0) acids. After optimization, the standard in vitro conditions for the measurement of delta6-desaturase activity were as follows: 60 micromol x L(-1) alpha-linolenic acid (C18:3n-3), reaction time of 20 min and protein content of 0.4 mg. Data showed that cell treatment with 0.5 mmol x L(-1) myristic acid during 43 h specifically increased delta6-desaturase activity. This improvement, reproducible for three substrates of delta6 desaturase, i.e. oleic acid (C18:1n-9), linoleic acid (C18:2n-6) and alpha linoleic acid (C18:3n-3) was dose-dependent in the range 0.1-0.5 mmol x L(-1) myristic acid concentration. PMID- 15270552 TI - X-linked mental retardation (XLMR): from clinical conditions to cloned genes. AB - X-linked mental retardation (XLMR) is a heterogenous set of conditions responsible for a large proportion of inherited mental retardation. Approximately 200 XLMR conditions and 45 cloned genes are now listed in our catalogue on the Internet at http://xlmr.interfree.it/home.htm. Traditionally, XLMR conditions were subdivided into specific (MRXS) and nonspecific (MRX) forms, depending on their clinical presentation. Now that a growing number of candidate genes have become available for screening XLMR families and patients, this distinction is becoming less useful and similar conditions that had been previously listed as separate can now be grouped together because different mutations in the same gene have been identified. Furthermore, different mutations in the same XLMR gene may account for diseases of increasing severity, but can also cause different phenotypes. As the functions of proteins corresponding to these genes are characterized, biological networks involved in causing mental retardation and conversely in supporting normal intellectual functioning will be discovered. Molecular biologists and neurobiologists will need to cooperate in order to verify the effects of XLMR gene mutations in the context of neuronal circuitry. Eventually, DNA and protein microarray technologies will assist researchers and physicians in reaching a diagnosis even in small families or in individual patients with XLMR. PMID- 15270553 TI - Human red blood cells: rheological aspects, uptake, and release of cytotoxic drugs. AB - The shape of a normal human red blood cell (RBC) is well known: under resting conditions it is that of a biconcave discocyte. However, RBCs can easily undergo transformation to other shapes with stomatocytes and echinocytes as extremes. Various anticancer agents, generally reactive and labile substances, e.g., oxazaphosphorines and fluoropyrimidines, can induce severe deformation of shape. Shape changes in erythrocytes can induce rheological disturbances, which occasionally have pathophysiological consequences. It is difficult to estimate the impact of shape changes on the in vivo behavior of agents of biological interest. However, it has been demonstrated for various anticancer agents that erythrocytes fulfill an important role in their uptake, transport, and release. Moreover, some anticancer agents are capable of influencing important transporters such as MRP and GLUT-1. Monitoring of erythrocyte concentrations of certain cytotoxic agents is therefore of interest as the data generated can have a predictive outcome for therapeutic efficacy. This is true for cyclophosphamide, ifosfamide, lometrexol, and 6-mercaptopurine, as well as MRP and GLUT-1 mediated agents. PMID- 15270554 TI - MUC1 and the MUCs: a family of human mucins with impact in cancer biology. AB - Mucins represent a family of glycoproteins characterized by repeat domains and a dense O-glycosylation. During the last two decades, the gene and peptide structures of various mucins as well as their glycosylation states were partly elucidated. Characteristic tumor-associated alterations of the expression patterns and glycosylation profiles were observed in biochemical, immunochemical, and histological studies and are discussed in the light of efforts to use the most prominent member in this family, MUC1, as a tumor target in anti-tumor strategies. Within this context the present review, focusing on MUC1, describes recent work on the regulation of mucin biosynthesis by cytokines and hormones, the role of mucins in cell adhesion, and their interaction with the immune system. Important aspects of clinical diagnostics based on mucin antigens are discussed, including the application of tumor serum assays and the significance of numerous studies revealing correlations between the expression of peptide cores or mucin-associated carbohydrates and clinicopathological parameters like tumor progression and prognosis. PMID- 15270555 TI - RNA interference of IL-10 in leukemic B-1 cells. AB - RNA interference, or RNAi, is designed to work by Watson-Crick base pairing and to result in a posttranscriptional block in protein synthesis. Antiapoptotic proteins are a major focus of cancer therapy and make attractive targets for RNAi. An IL-10 RNAi sequence was designed in accordance with Tuschl rules and was modeled to a hairpin configuration. In chronic lymphocytic leukemia (CLL), the most common leukemia in the Western world, the failure to undergo apoptosis may be responsible for the accumulation of malignant B-1 cells. Interleukin-10, despite controversy, has been shown to have antiapoptotic properties, and increased endogenous IL-10 production has been found in CLL by several labs. A malignant B-1 cell line, LNC, derived from an NZB mouse (a murine model for CLL) was utilized as a target for IL-10 RNAi. Our earlier studies of antisense IL-10 resulted in antiproliferative and proapoptotic effects. The cytotoxic effects of IL-10 RNAi were dose- and time-dependent, with an optimal dose 10-fold lower than that of antisense IL-10. IL-10 RNAi lowered IL-10 protein as measured by ELISA. 2 micro M IL-10 RNAi initiated a G2/M block and a decrease in the message for cdc25C, the M-phase inducer phosphatase. IL-10 RNAi efficiently induced apoptosis. Bcl7C, a member of the antiapoptotic Bcl family, was significantly down-regulated. IL-10 modulating Bcl7C expression represents a novel mechanism in the evasion of apoptosis. This approach, by itself or in conjunction with current therapies, merits consideration in similar B-cell malignancies. PMID- 15270556 TI - Sesquiterpenes and flavonol glycosides from Zingiber aromaticum and their CYP3A4 and CYP2D6 inhibitory activities. AB - Three new sesquiterpenes, (2R,3S,5R)-2,3-epoxy-6,9-humuladien-5-ol-8-one (1), (2R,3R,5R)-2,3-epoxy-6,9-humuladien-5-ol-8-one (2), and (5R)-2,6,9-humulatrien-5 ol-8-one (3), and two new flavonol glycosides, kaempferol-3-O-(2,3-di-O-acetyl alpha-l-rhamnopyranoside) (4) and kaempferol-3-O-(2,3,4-tri-O-acetyl-alpha-l rhamnopyranoside) (5), were isolated from the EtOAc-soluble fraction of the water extract of Zingiber aromaticum, along with 13 known compounds (6-18). The structures of the isolated compounds were elucidated on the basis of spectroscopic and chemical analyses. The isolated compounds were tested for their inhibitory activity on the metabolism mediated by CYP3A4 or CYP2D6 using [N methyl-(14)C]erythromycin or [O-methyl-(14)C]dextromethorphan as a substrate, respectively. Kaempferol-3-O-(2,3,4-tri-O-acetyl-alpha-l-rhamnopyranoside) (5) showed the most potent inhibitory activity (IC(50), 14.4 microM) on the metabolism mediated by CYP3A4, and kaempferol-3-O-methyl ether (14) inhibited CYP2D6 most potently (IC(50), 4.63 microM). PMID- 15270557 TI - A new antifungal metabolite from Penicillium expansum. AB - A new antifungal compound, (3S)-4,6-dihydro-8-methoxy-3,5-dimethyl-6-oxo-3H-2 benzopyran (4), was isolated from Penicillium expansum. During the isolation procedure 4 was determined to be unstable and readily reacted with methanol, ethanol, and water, forming three new isochromans, (1S,3S)-6-hydroxy-1,8 dimethoxy-3,5-dimethylisochroman (1), 1-ethoxy-6-hydroxy-8-methoxy-3,5 dimethylisochroman (2), and 1,6-dihydroxy-8-methoxy-3,5-dimethylisochroman (3), respectively. (3S)-6-Hydroxy-8-methoxy-3,5-dimethylisochroman (5) was reisolated from P. expansum. In fungicide disk assays, compounds 1, 2, and 4 inhibited the mycelial growth of Lasiodiplodia theobromae at 100 microg/mL by 76%, 74%, and 69%, respectively. PMID- 15270558 TI - New cytotoxic oleanane-type triterpenoids from the cones of Liquidamber styraciflua. AB - Two new oleanane-type triterpenoids (1 and 2), together with two known compounds, 6beta-hydroxy-3-oxo-lup-20(29)-en-28-oic acid (3) and 3,11-dioxoolean-12-en-28 oic acid (4), were isolated from the stem bark of Liquidamber styraciflua. The structures of 1 and 2 were determined to be 25-acetoxy-3alpha-hydroxyolean-12-en 28-oic acid (1) and 3alpha,25-dihydroxyolean-12-en-28-oic acid (2) on the basis of spectroscopic methods and chemical conversion. Compound 1 showed strong cytotoxicity against a disease-oriented panel of 39 human cancer cell lines, although compounds 2, 3, and 4 showed weaker activity compared to 1. PMID- 15270559 TI - New alkaloids from Daphniphyllum calycinum. AB - Four new alkaloids, 17-hydroxyhomodaphniphyllic acid (1), daphcalycinosidine C (2, a new iridoid alkaloid), yuzurimine E (3), and yuzurimic acid B (4), were isolated from the seeds of Daphniphyllum calycinum. The structures of these Daphniphyllum alkaloids were determined by spectroscopic analysis including mass spectrometry and 2D NMR. PMID- 15270560 TI - Synthesis of A-seco derivatives of betulinic acid with cytotoxic activity. AB - In this study, the relationships between the chemical structure and cytotoxic activity of betulinic acid (1) derivatives were investigated. Eight lupane derivatives (1-8), one of them new (6), five diosphenols (9-13), four of them new (10-13), two new norderivatives (14 and 15), five seco derivatives (16-20), four of them new (16, 17, 19, and 20), and three new seco-anhydrides (21-23) were synthesized from 1, and their activities were compared with the activities of known compounds. The effects of substitution on the A-ring and esterification of the carboxyl group in position 28 on cytotoxicity were of special interest. Significant cytotoxic activity against the T-lymphoblastic leukemia cell line CEM was found in diosphenols 9 and 13 (TCS(50) 4 and 5 micromol/L) and seco anhydrides 22 and 23 (TCS(50) 7 and 6 micromol/L). All compounds were also tested on cancer cell lines HT 29, K562, K562 Tax, and PC-3, and these confirmed activity of diosphenols 9, 10, and 11 and anhydride 22. Diosphenols, as the most promising group of derivatives, were further tested on four more lines (A 549, DU 145, MCF 7, SK-Mel2). PMID- 15270561 TI - Structures and histamine release inhibitory effects of prenylated orcinol derivatives from Rhododendron dauricum. AB - Four new prenylated orcinol derivatives, daurichromenes A-D (1-4), along with three known compounds, confluentin (5), grifolin (6), and orcinol (7), have been isolated from the Chinese medicinal plant Rhododendron dauricum. Their structures were established as 2R-(7'-hydroxy-4',8'-dimethyl-3'E,8'-nonadienyl)-5-hydroxy 2,7-dimethyl-2H-chromene (1), 2R-(3'-hydroxy-8'-methyl-4'-methyliden-7'-nonaenyl) 5-hydroxy-2,7-dimethyl-2H-chromene (2), 2R-(8'-hydroxy-4',8'-dimethyl-3'E,6'Z nonadienyl)-5-hydroxy-2,7-dimethyl-2H-chromene (3), and 2R-(9'-hydroxy-4',8' dimethyl-3'E,7'E-nonadienyl)-5-hydroxy-2,7-dimethyl-2H-chromene (4) by analysis of spectral data. The absolute configuration of the asymmetric carbons at the chromene ring in 1-5 was determined as R from their circular dichroism spectra. Compounds 1-6 significantly inhibited compound 48/80-induced histamine release from rat peritoneal mast cells. PMID- 15270562 TI - New lanostane-type triterpenoids from Ganoderma applanatum. AB - Four new lanostane-type triterpenes were isolated from the MeOH extract of the fruiting bodies of Ganoderma applanatum. Their structures were established as 3beta,7beta,20,23xi-tetrahydroxy-11,15-dioxolanosta-8-en-26-oic acid (1), 7beta,20,23xi-trihydroxy-3,11,15-trioxolanosta-8-en-26-oic acid (2), 7beta,23xi dihydroxy-3,11,15-trioxolanosta-8,20E(22)-dien-26-oic acid (3), and 7beta-hydroxy 3,11,15,23-tetraoxolanosta-8,20E(22)-dien-26-oic acid methyl ester (4), respectively, by extensive spectroscopic analyses. PMID- 15270563 TI - Glandulosides A-D, triterpene saponins from Acanthophyllum glandulosum. AB - Four novel triterpenoid saponins, glandulosides A (1), B (2), C (3), and D (4), together with two known saponins (5 and 6) have been isolated from the roots of Acanthophyllum glandulosum. Their structures were elucidated using a combination of homo- and heteronuclear 2D NMR techniques (COSY, TOCSY, NOESY, HSQC, and HMBC) and by FABMS. The new compounds were characterized as 23-O-beta-D galactopyranosylgypsogenic acid-28-O-beta-D-glucopyranosyl-(1-->3)-[beta-d galactopyranosyl-(1-->6)]-beta-D-galactopyranoside (1), 3-O-beta-D galactopyranosyl-(1-->2)-[beta-D-xylopyranosyl-(1-->3)]-beta-D glucuronopyranosylgypsogenin-28-O-beta-D-xylopyranosyl-(1-->3)-beta-D xylopyranosyl-(1-->4)-alpha-l-rhamnopyranosyl-(1-->2)-3-O-acetyl-beta-D fucopyranoside (2), 3-O-beta-D-galactopyranosyl-(1-->2)-[beta-D-xylopyranosyl-(1- >3)]-beta-D-glucuronopyranosylgypsogenin-28-O-beta-D-xylopyranosyl-(1-->3)-beta-D xylopyranosyl-(1-->4)-alpha-L-rhamnopyranosyl-(1-->2)-3,4-di-O-acetyl-beta-D fucopyranoside (3), and 3-O-beta-D-galactopyranosyl-(1-->2)-[beta-D-xylopyranosyl (1-->3)]-beta-D-glucuronopyranosylgypsogenin-28-O-beta-D-xylopyranosyl-(1-->3) beta-D-xylopyranosyl-(1-->4)-alpha-l-rhamnopyranosyl-(1-->2)-[3-O-acetyl-beta-D quinovopyranosyl-(1-->4)]-beta-D-fucopyranoside (4). PMID- 15270564 TI - New crinine-type alkaloids with inhibitory effect on induction of inducible nitric oxide synthase from Crinum yemense. AB - The 80% aqueous methanolic extract from the bulbs of Crinum yemense showed a potent inhibitory effect on nitric oxide production in lipopolysaccharide activated macrophages. Three new crinine-type alkaloids, yemenines A (1), B (2), and C (3), were isolated from the herbal extract together with six known alkaloids. The absolute configurations of 1-3 were determined on the basis of chemical and physicochemical evidence. The effects of the isolated alkaloids on nitric oxide production in lipopolysaccharide-activated macrophages were examined, and several alkaloids, e.g. 1, (+)-bulbispermine (6), (+)-crinamine (7), (+)-6-hydroxycrinamine (8), and (-)-lycorine (9), showed inhibitory effects on nitric oxide production and induction of inducible nitric oxide synthase. PMID- 15270565 TI - Chemical constituents of Millettia taiwaniana: structure elucidation of five new isoflavonoids and their cancer chemopreventive activity. AB - We describe the isolation and identification of five new isoflavonoids, millewanins A (1), B (2), C (3), D (4), and E (5), together with six known isoflavonoids and three rotenoids, from the stems of Millettia taiwaniana collected in Japan. The major component, auriculasin (6), exhibited significant inhibitory effect on mouse skin tumor promotion in an in vivo two-stage carcinogenesis test. The results of the present investigation indicate that 6 might be a valuable antitumor promoter. PMID- 15270566 TI - Quinazolin-4-one derivatives from Streptomyces isolates. AB - From the ethyl acetate extract of the strain Streptomyces sp. isolate GW23/1540, besides 16 known products, several 1H-quinazolin-4-one derivatives were isolated. (SR)-2-(1-Hydroxyethyl)-2-methyl-2,3-dihydro-1H-quinazolin-4-one (4) and (RR)-2 (1-hydroxyethyl)-2-methyl-2,3-dihydro-1H-quinazolin-4-one (5) are new natural products. 2-Methyl-3H-quinazolin-4-one (2) and 1H-quinazoline-2,4-dione (3) are known from other bacteria and plants, respectively. From another Streptomyces sp., GW2/577, 5-methyl-1H-quinazoline-2,4-dione (6) was isolated and the structure proven by comparison with the isomeric 7. The new natural products showed no activity against the microalgae Chlorella vulgaris, Chlorella sorokiniana, and Scenedesmus subspicatus, the fungus Mucor miehei, the yeast Candida albicans, and the bacteria Staphylococcus aureus, Bacillus subtilis, Escherichia coli, and Streptomyces viridochromogenes. PMID- 15270567 TI - New lignans from the roots of Valeriana prionophylla with antioxidative and vasorelaxant activities. AB - Two new 7,9':7',9-diepoxylignan glycosides have been isolated from the roots of Valeriana prionophylla. Their structures have been established on the basis of 1D and 2D NMR experiments as prinsepiol-4-O-beta-D-glucopyranoside (1) and fraxiresinol-4'-O-beta-D-glucopyranoside (2). In addition, 8-hydroxypinoresinol 4'-O-beta-d-glucopyranoside (3), 8-hydroxypinoresinol (4), prinsepiol (5), and chlorogenic acid were isolated. Compounds 1, 3, 4, and 5 were evaluated for their antioxidative properties in Trolox equivalent antioxidant activity (TEAC) and chemiluminescence (CL) assays. The same compounds were tested for their vascular activity in rat aorta rings. The aglycones 4 and 5 displayed powerful antioxidant activity; in addition, aglycone 4 showed a higher vasorelaxant activity than compounds 1, 3, and 5. PMID- 15270568 TI - Molecular bilateral symmetry of natural products: prediction of selectivity of dimeric molecules by density functional theory and semiempirical calculations. AB - A literature survey and theoretical calculations have been applied to explore bilateral symmetry in natural product systems. Molecular bilateral symmetry is defined to include C(2) (sigma plane or axis), C(s)(), and C(2)(v)() point groups in molecules. Natural products that possess chirality in the form of C(2)-axes or sigma planes of symmetry are present in higher proportions (69%) compared to molecules bearing achiral C(s)() or C(2)(v)() point groups (14% and 16%, respectively). Density functional theoretical and semiempirical calculations indicate that the dimers 3,3'-dibromo-5,5'-[N-(2-(3-bromo-4-hydroxyphenyl)ethyl) 2-hydroxyiminoacetamide]biphenyl-2,2'-diol (1), (S,S)-1,2-bis(2-amino-3H-imidazol 4-yl)-(R,R)-3,4-bis(1H-pyrrole-2-amido)cyclobutane (2), 2-oxo-dimethyl-1,3 bis(3,4-dibromobenzene-1,2-diol) (11), 1,7-bis(4-hydroxy-3-methoxyphenyl)hepta 1,6-diene-3,5-dione (12), and bis(5-isopropyl-8-methylazulene)methane (13) evolve more energy per connecting bond than the corresponding trimers or tetramers would. This we propose is a guiding parameter that may adjust molecule growth. The corresponding trimers, tetramers, or higher oligomers of 1, 2, and 11-13 appear to represent "missing" compounds in nature. Natural products 1, 2, and 11 13, having 3-fold and higher levels of symmetry, would founder on the lack of a facile method of synthesis and on the prohibitively high-energy costs caused by steric crowding at their core. PMID- 15270569 TI - Five new Ocotillone-type saponins from Gynostemma pentaphyllum. AB - Five new ocotillone-type saponins, gynosides A-E (1-5), along with 10 known dammarane-type saponins, were isolated from the aerial parts of Gynostemma pentaphyllum. The structures of these new compounds were determined by NMR analysis and acid hydrolysis. The structure and stereochemistry of gynoside A (1) were confirmed by X-ray crystallography. PMID- 15270570 TI - New azaphilones from the inedible mushroom Hypoxylon rubiginosum. AB - Fractionation of the methanol extract of the fruit bodies of the xylariaceus ascomycete Hypoxylon rubiginosum resulted in the isolation of three new azaphilone derivatives named rubiginosins A-C (1-3) and a new fatty acid named rubiginosic acid (4), together with three known compounds, entonaemin A (5), daldinin C (6), and orsellinic acid. Their structures were elucidated by 2D NMR, MS, IR, and UV spectra and chemical reaction. The absolute configuration of compound 3 was established by CD spectroscopy. The structure of 4 was confirmed by X-ray crystallographic analysis. PMID- 15270571 TI - Cytotoxic and antimicrobial constituents of the bark of Diospyros maritima collected in two geographical locations in Indonesia. AB - Bioactivity-directed fractionation of extracts of two Diospyros maritima bark samples from Indonesia,one collected at sea level in a beach forest in Java and the other collected at a slight elevation away from the sea shore on the island of Lombok, yielded a diverse set of secondary metabolites. The naphthoquinone plumbagin (1), although found in extracts of both specimens, constituted a much larger percentage of the former sample, which also yielded a series of plumbagin dimers, maritinone (2), chitranone (3), and zeylanone (4). The latter sample yielded a new naphthoquinone derivative, (4S)-shinanolone (5), and a new natural product coumarin, 7,8-dimethoxy-6-hydroxycoumarin (6), along with three other analogues of plumbagin, 2-methoxy-7-methyljuglone (7), 3-methoxy-7-methyljuglone (8), and 7-methyljuglone (9). The structures of compounds 5 and 6 were elaborated by physical, spectral, and chemical methods. All of the isolates were evaluated in both cytotoxicity and antimicrobial assays, and structure-activity relationships of these naphthoquinones are proposed. Plumbagin (1) and maritinone (2) were evaluated also for in vivo antitumor activity in the hollow fiber assay, but both were found to be inactive. PMID- 15270572 TI - A DNA-damaging oxoaporphine alkaloid from Piper caninum. AB - Bioassay-guided fractionation of an active organic extract of Piper caninum, using a sensitive yeast assay to monitor putative double-strand DNA-damaging activity, resulted in the isolation of the 4,5-dioxoaporphine alkaloid cepharadione A (1). Compound 1 exhibited potent inhibitory activity in a yeast cytotoxicity assay with IC(50) values of 50.2 nM toward RS321NpRAD52 grown on glucose versus 293 nM toward the same yeast strain grown on galactose. PMID- 15270573 TI - Monanchorin, a bicyclic alkaloid from the sponge Monanchora ungiculata. AB - Monanchorin, a guanidine alkaloid with an unusual bicyclic skeleton, together with the known pentacyclic alkaloid crambescidin acid have been isolated from the aqueous extract of the sponge Monanchoraungiculata. PMID- 15270574 TI - New lamellarin alkaloids from the Indian ascidian Didemnum obscurum and their antioxidant properties. AB - Three new lamellarin alkaloids, lamellarins gamma (1), alpha (2), and epsilon (3), along with eight known lamellarin alkaloids, lamellarins M (4), K (5), K diacetate (6), K-triacetate (7), U (8), I (9), C-diacetate (10), and X-triacetate (11), have been isolated from the Indian ascidian Didemnum obscurum. The structures of 1-11 were established using standard spectroscopic techniques. The structure of lamellarin K-triacetate (7) was further confirmed by X-ray crystallographic analysis. The antioxidant properties of lamellarin gamma, lamellarin gamma-monoacetate, lamellarins K, U, and I, and lamellarin C-diacetate were evaluated. PMID- 15270575 TI - Further membranolide diterpenes from the antarctic sponge dendrilla membranosa. AB - Chemical investigation of the Antarctic sponge Dendrilla membranosa collected from the vicinity of Palmer Station on Anvers Island, Antarctica, yielded three new diterpenes, membranolides B-D (2-4), as well as three previously reported sponge metabolites. Membranolides C and D (3, 4), bearing carboxylic acid functional groups, display Gram-negative antibiotic and antifungal activities. PMID- 15270576 TI - A new anti-HIV alkaloid, drymaritin, and a new C-glycoside flavonoid, diandraflavone, from Drymaria diandra. AB - A novel anti-HIV alkaloid, drymaritin (1), and a new C-glycoside flavonoid, diandraflavone (2), along with eight known compounds, torosaflavone A, isovitexin, spinasterol beta-d-glycoside, p-hydroxybenzoic acid, p hydroxybenzaldehyde, cis-p-coumarate, methyl 5-hydroxy-4-oxopentanoate, and glycerol-alpha-lignocerate, were isolated from Drymaria diandra. Drymaritin (1) exhibited anti-HIV effects in H9 lymphocytes with an EC(50) value of 0.699 microg/mL and a TI of 20.6. Compound 2 showed significantly selective inhibition on superoxide anion generation from human neutrophils stimulated by fMLP/CB with an IC(50) value of 10.0 microg/mL. PMID- 15270577 TI - Cyclic peptides from a Ruegeria strain of bacteria associated with the sponge Suberites domuncula. AB - Two new cyclic peptides, cyclo-(glycyl-L-seryl-L-prolyl-L-glutamyl) and cyclo (glycyl-L-prolyl-L-glutamyl), have been isolated from the cell extract of a Ruegeria strain associated with cell cultures of Suberites domuncula. Three other cyclopeptides have been isolated for the first time from a natural source. Additionally, a new diastereoisomer of a known compound is reported. The structures of isolated compounds have been elucidated by means of spectroscopic data (1D-, 2D-NMR, HRESIMS) and chiral HPLC analysis. The new cyclopeptides exhibited moderate antimicrobial activity against Bacillus subtilis. PMID- 15270578 TI - New alkaloids and a tetraflavonoid from Cephalotaxus wilsoniana. AB - A new homoerythrina alkaloid, C-3-epi-wilsonione (1), a new tetraflavonoid, taiwanhomoflavone C (2), and a new stereoisomer of desmethylcephalotaxinone (3) have been isolated from the leaves and heartwood of Cephalotaxus wilsoniana, respectively. The structures were elucidated by spectroscopic methods. Compound 1 showed cytotoxic activity against a number of human cancer cell lines in vitro. PMID- 15270579 TI - Cytotoxic furanosesterterpenes from a marine sponge Psammocinia sp. AB - Three new (1-3) and seven known (4-10) cytotoxic furanosesterterpenes were isolated from a marine sponge Psammocinia sp. by bioactivity-guided fractionation. The structures were established on the basis of NMR and MS analyses. The geometry and absolute configuration were determined on the basis of optical rotation, NMR, and CD data. These compounds were evaluated for cytotoxicity against a small panel of five human tumor cell lines, and most of the compounds showed toxicity to SK-MEL-2. The mixture of compounds 7 and 8 displayed significant inhibition of DNA replication and moderate antioxidant profile. PMID- 15270580 TI - New cytotoxic cembranes from the sea pen Gyrophyllum sibogae. AB - Two new cembrane-type diterpenoids have been isolated from the 2-propanol extract of the sea pen Gyrophyllum sibogae collected in South Africa: 7,8 dihydroflabellatene A (1) and 7,8-dihydroflabellatene B (2). Their structures were determined on the basis of detailed spectroscopic analysis and by single crystal X-ray analysis of the major metabolite 1, which showed strong in vitro cytotoxicity against a panel of 13 tumor cell lines. PMID- 15270581 TI - Hemisynthetic secofriedelane triterpenes with inhibitory activity against the growth of human tumor cell lines in vitro. AB - Seco acids 7 and 9 and hydroxylated analogues 5 and 6 derived from friedelane triterpenes were synthesized stereoselectively in high yields. Compounds 5-9 were evaluated for their ability to inhibit in vitro the growth of three human tumor cell lines, MCF-7 (breast adenocarcinoma), NCI-H460 (non-small cell lung cancer), and SF-268 (CNS cancer). Only compounds 7 and 9 were found to possess significant growth inhibitory effects, exhibiting GI(50) values that range from 24.6 to 32.8 microM and 10.9 to 17.6 microM, respectively. PMID- 15270582 TI - Detectors for digital mammography. AB - Interest in digital radiography was stimulated by the enthusiastic acceptance of computed tomography in the early 1970s. It quickly became apparent to the medical community that images with improved information content, whose display characteristics could be manipulated by the viewer, provided many advantages. Subsequently, digital systems for subtraction angiography and later for conventional projection radiography and fluoroscopy were developed. The timing of the introduction of these systems was highly dependent on the readiness of certain key component technologies to meet the requirements of each of these applications. These components are the x-ray detectors, analog to digital converters, computers, data storage systems and high-resolution electronic displays and printers used in image acquisition, storage and display. Mammography represents one of the most demanding radiographic applications, simultaneously requiring excellent contrast sensitivity, high spatial resolution, and wide dynamic range at as low as radiation dose to the breast as is reasonably achievable while meeting the other requirements. For this reason, it is one of the last radiographic procedures to "go digital". Here, some of the considerations related to the detector technology for digital mammography will be discussed and systems currently available will be described. PMID- 15270583 TI - Combination of digital mammography with semi-automated 3D breast ultrasound. AB - This paper describes work aimed at combining 3D ultrasound with full-field digital mammography via a semi-automatic prototype ultrasound scanning mechanism attached to the digital mammography system gantry. Initial efforts to obtain high x-ray and ultrasound image quality through a compression paddle are proving successful. Registration between the x-ray mammogram and ultrasound image volumes is quite promising when the breast is stably compressed. This prototype system takes advantage of many synergies between the co-registered digital mammography and pulse-echo ultrasound image data used for breast cancer detection and diagnosis. In addition, innovative combinations of advanced US and X-ray applications are being implemented and tested along with the basic modes. The basic and advanced applications are those that should provide relatively independent information about the breast tissues. Advanced applications include x ray tomosynthesis, for 3D delineation of mammographic structures, and non-linear elasticity and 3D color flow imaging by ultrasound, for mechanical and physiological information unavailable from conventional, non-contrast x-ray and ultrasound imaging. PMID- 15270584 TI - The impact of technological advances on the evolution of 3D conformal brachytherapy for early prostate cancer. AB - Permanent implantation of I-125 and Pd-103 seeds is one of the widely used treatment options for the early stage prostate cancer with minimum normal tissue complications and long-term local control of the tumor. This is possible because of several technological advances made in the past decade to better understand the procedural aspects of implantations with the desired clinical outcome and with acceptable morbidities. In addition, with the widespread use of PSA testing, more widely disseminated information about prostate cancer and increased patient awareness, over 70% of patients are diagnosed early with localized disease and therefore are candidates for definitive local therapy. Delineation of soft tissue structures including the prostate, rectum, urethra and bladder has become more accurate with the use of imaging modalities including Ultrasound and MRI, with or without the CT. A re-evaluation of the dosimetric parameters of the radioactive sources has lead to a more precise estimate of the dose delivered to the prostate and the associated critical normal structures. Technological improvements in the post implant dosimetry have helped to understand the factors, which makes an implant a "good implant" or a "poor implant". Intraoperative treatment planning with on line dosimetry is emerging as one of the best approaches for prostate brachytherapy. In addition, better software is now available producing dose volume histograms with 3D target and normal tissue reconstruction. The combination of seed implant followed by IMRT would provide scope for differentially boosting the regions under-dosed because of uncontrollable and unexpected reasons during the implant and unsuspected micro extensions of the tumor. PMID- 15270585 TI - Radiation dosimetry of a conformal heat-brachytherapy applicator. AB - The purpose of this paper is to report the radiation dosimetric characteristics of a new combination applicator for delivering heat and radiation simultaneously to large area superficial disease <1.5 cm deep. The applicator combines an array of brachytherapy catheters (for radiation delivery) with a conformal printed circuit board microwave antenna array (for heat generation), and a body conforming 5-10 mm thick temperature-controlled water bolus. The rationale for applying both modalities simultaneously includes the potential for significantly higher response rate due to enhanced synergism of modalities, and lower peak toxicity due to temporal extension of heat and radiation induced toxicities. Treatment plans and radiation dosimetry are calculated with IPSA (an optimization tool developed at UCSF) for 15 x 15 cm(2) and 35 x 24 cm(2) applicators, lesion thicknesses of 5 to 15 mm, flat and curved surfaces, and catheter separation of 5 and 10 mm. The effect on skin dose of bolus thickness and presence of thin copper antenna structures between radiation source and tissue are also evaluated. Results demonstrate the ability of the applicator to provide conformal radiation dose coverage for up to 15 mm deep target volumes under the applicator. For clinically acceptable plans, tumor coverage is > 98%, homogeneity index > 0.95 and the percentage of normal tissue irradiated is < 20%. The dose gradient at the skin surface varies from 3 to 5 cGy/mm depending on bolus thickness and lesion depth. Attenuation of the photon beam by the printed circuit antenna array is of the order 0.25% and secondary electron emissions are absorbed completely within 5 mm of water bolus and plastic layers. Both phenomena can then be neglected in dose calculations allowing commercial software to be used for treatment planning. This novel applicator should prove useful for the treatment of diffuse chestwall disease located over contoured anatomy that may be difficult to treat with single field external beam therapy. By delivering heat and radiation simultaneously, increased synergism is expected with a TER in the range of 2-5. Lowering radiation dose by an equivalent factor may produce lower radiation toxicity with similar efficacy, while preserving the option of subsequent retreatment(s) with thermoradiotherapy in order to further extend patient survival. PMID- 15270586 TI - Fiducial markers implanted during prostate brachytherapy for guiding conformal external beam radiation therapy. AB - Prostate movement imposes limits on safe dose-escalation with external beam radiation therapy. If the precise daily location of the prostate is known, dose escalation becomes more feasible. We have developed an approach to dose escalation using a combination of prostate brachytherapy followed by external beam radiation therapy in which fiducial markers are placed along with (125)I seeds during transperineal interstitial permanent prostate brachytherapy. These markers serve to verify daily prostate location during the subsequent external beam radiotherapy. Prior to implementing this approach, preliminary studies were performed to test visibility of the markers. Three different (125)I seed models, as well as gold and silver marker seeds were placed within tissue-equivalent phantoms. Images were obtained with conventional x-rays (75-85 kV) and 6 MV photons from a linear accelerator. All (125)I seed models were clearly visible on conventional x-rays but none were seen with 6 MV photons. The gold markers were visible with both energies. The silver markers were visible with conventional x rays and 6 MV x-rays, but not as clearly as the gold seeds at 6 MV. Subsequently, conventional x-rays, CT scans, and 6 MV port films were obtained in 29 patients in whom fiducial gold marker seeds were implanted into the prostate during (125)I prostate brachytherapy. To address the possibility of "seed migration" within the prostate, CT scans were repeated 5 weeks apart in 14 patients and relative positions of the gold seeds were evaluated. The repeated CT scans showed no change in intraprostatic gold marker location, suggesting minimal migration. The gold seeds were visible with conventional x-rays, CT, and 6 MV port films in all patients. During the course of external beam radiation therapy, the gold markers were visible on routine 6 MV port films and were seen in different locations from film to film suggesting prostate motion. Mean daily displacement was 4-5 mm in the anterior-posterior, and 4-5 mm in superior-inferior dimensions. Left-right displacement appeared less, averaging 2-3 mm. We conclude that implantation of gold marker seeds during prostate brachytherapy represents an easily implemented and practical means of prostate localization during subsequent image-guided external beam radiotherapy. With such markers, conformality of the external beam component can be confidently improved without expensive new equipment. PMID- 15270587 TI - The male lumpectomy: rationale for a cancer targeted approach for prostate cryoablation. A review. AB - Lumpectomy to treat breast cancer has revolutionized the management of that disease. Lumpectomy showed that the quality of life of the individual patient can successfully be integrated into the equation of cancer treatment, without major loss of cancer treatment efficacy. Prostate cancer raises many of the same issues that breast cancer does in women. Impotence and incontinence, affects the male self image and psyche no less than the loss of a breast does a woman. Management of prostate cancer ranges from no treatment at all ("watchful waiting") to treatments in which the whole gland is destroyed (radiation therapy, cryosurgery) or removed (radical prostatectomy), with presently no treatment in between these extremes. Pathologic literature indicates, however, that 35% of prostate cancers are solitary and unilateral. In addition, long term studies have confirmed that cryoabltion for prostate cancer is an efficacious treatment. In this paper we will examine the rationale for a "male lumpectomy" using cryoablation and present preliminary data supporting it's role in prostate cancer management. PMID- 15270588 TI - Cryotherapy of musculoskeletal tumors--from basic science to clinical results. AB - Combined modality treatment of musculoskeletal tumors led to improved patient survival. As survival improves, more consideration is given to the functional outcome of treatment, and interest is focused on the development of less mutilating and extensive surgery. One modality that can reduce patient disability significantly is cryosurgery, as it allows minimally invasive surgery based on marginal resection and tumor interface sterilization instead of wide resection of certain neoplasms. Classical cryosurgery as developed by Marcove involves pouring of liquid nitrogen into the tumor bed. This approach revolutionized the treatment of some tumors such as giant cell tumor of bone, allowing intra-lesional resection to substitute the wide-resection method used up to that time. However, complications of this method of treatment are common, including nitrogen emboli, fractures of the bone due to extensive necrosis and damage to neurovascular elements. A recent development in the field of cryosurgery has been the argon based system allowing controlled formation of an ice-ball surrounding a metallic probe. The system is computer controlled and allows precise evaluation of the tumor bed interface as well as surrounding structures that need to be protected. Prior to application of this method in humans it is important to ensure that interface sterilization is indeed achieved using cryosurgery. To evaluate this question, a Swarm rat chondrosarcoma was used. Cell viability was assessed following ice-ball formation. Histological evaluation indicated that cell death occurs up to 5 millimeters from the ice-ball if temperatures of -40 degrees Celsius at the metallic probe are achieved. A further evaluation was performed on samples obtained from patients during surgery. A minimum of two freezing cycles was shown to be necessary to achieve tissue viability similar to that of boiled tissue. Twenty-seven patients were operated to date using an argon-based cryosurgery system. The patients included 7 cases of grade I chondrosarcoma, 5 cases of giant cell tumor of bone, 14 cases of a metastatic lytic bone lesion and a single case of osseous-fibrous dysplasia. None of the patients suffered nerve injury during the operation. After a minimal follow-up period of 2 years only two of the surviving patients had a recurrence (a giant cell tumor of the proximal fibula, and the patient with the osseous-fibrous dysplasia whose tumor recurred as a frankly malignant adamantimoma). There were no pathological fractures. This method appears practical and allows close monitoring of the surrounding tissue to reduce the chances of recurrence. PMID- 15270589 TI - Magnetic resonance imaging detects early changes in microvascular permeability in xenograft tumors after treatment with the matrix metalloprotease inhibitor Prinomastat. AB - Macromolecular contrast medium-enhanced magnetic resonance imaging was applied to monitor the effect of matrix metalloprotease (MMP) inhibition on microvascular characteristics of human breast cancers implanted in athymic rats. Twice-daily intraperitoneal administration of Prinomastat over 1.5 days induced significant declines in MRI-assayed microvascular permeabilities (p<0.05); but this leak suppression effect had extinguished by the 10(th) day of MMP treatment using the same dose and time schedule. Results demonstrate that Prinomastat produces a rapid but transient decrease in tumor vascular permeability. Contrast-enhanced MRI using macromolecular contrast medium may prove useful as a biomarker for the dynamic MMP biological effect in cancers. PMID- 15270590 TI - Modifications of cellular autofluorescence emission spectra under oxidative stress induced by 1 alpha,25dihydroxyvitamin D(3) and its analog EB1089. AB - We attempted to characterize the cellular autofluorescence phenomenon of living HL-60 cells and to appraise its modifications under oxidative stress conditions induced by 1 alpha,25(OH)(2)D(3) (VD(3)) and its analog EB1089. Autofluorescence emission spectra of human promyelocytic HL-60 leukemic cells were monitored using laser scanning confocal microspectrofluorometry under UV excitation. Evaluation of reactive oxygen species (ROS) release was performed using the 2',7' dichlorodihydrofluorescein diacetate (H(2)-DCFDA) staining and fluorescence emission measurement. VD(3) (1, 10, 100 nM) or EB1089 (0.1, 1 and 10 nM) induces a decrease in autofluorescence emission intensity that can be attributed to the oxidation of the coenzyme nicotinamide adenine dinucleotide (phosphate) NAD(P)H into NAD(P)(+). A dose-dependent increase (p<0.05) in ROS release is observed in VD(3)- and EB1089-treated cells. As compared with VD(3)- or EB1089-treated cells, doxorubicin-VD(3) or doxorubicin-EB1089 treatments strongly decrease the autofluorescence intensity and induce a higher release of ROS (p<0.05). The association of antioxidants (N-acetyl cysteine, superoxide dismutase, catalase) with VD(3) or EB1089 induce a more limited autofluorescence decrease and a weaker ROS generation, as compared with VD(3) and EB1089 treated cells. In conclusion, the free radicals release, generated by VD(3) and EB1089, was associated with the decrease in autofluorescence emission and can be modulated by doxorubicin and antioxidants. PMID- 15270591 TI - Lighting up tumors with receptor-specific optical molecular probes. AB - Accurate and rapid detection of tumors is of great importance for interrogating the molecular basis of cancer pathogenesis, preventing the onset of complications, and implementing a tailored therapeutic regimen. In this era of molecular medicine, molecular probes that respond to, or target molecular processes are indispensable. Although numerous imaging modalities have been developed for visualizing pathologic conditions, the high sensitivity and relatively innocuous low energy radiation of optical imaging method makes it attractive for molecular imaging. While many human diseases have been studied successfully by using intrinsic optical properties of normal and pathologic tissues, molecular imaging of the expression of aberrant genes, proteins, and other pathophysiologic processes would be enhanced by the use of highly specific exogenous molecular beacons. This review focuses on the development of receptor specific molecular probes for optical imaging of tumors. Particularly, bioconjugates of probes that absorb and fluoresce in the near infrared wavelengths between 750 and 900 nm will be reviewed. PMID- 15270592 TI - Prescribing antiepileptic drugs: should patients be switched on the basis of cost? AB - To assess the costs of switching from one antiepileptic drug (AED) to another, all associated direct and indirect costs, not only drug acquisition costs, must be considered. The perspective of the healthcare system evaluated in cost effectiveness analysis is of crucial importance. Multiple clinical factors can influence clinical decisions regarding switching AEDs. The economic cost of poorly controlled epilepsy is enormous and the most cost-effective intervention is an AED that provides total seizure control. Cost-minimisation studies have evaluated costs associated with various medications. If only efficacy and adverse events were considered, then the 'older' AEDs were generally more cost effective than the 'newer' AEDs. Most studies only examine very specific clinical situations and are not suitable for establishing general clinical recommendations. The pharmacoeconomics of AED choice is highly country specific. While switching to generic formulations is, in general, cost effective, some changes may be detrimental and more costly than remaining on the trade name preparation. For example, as a result of differences in bioavailability and possible loss of seizure control, changing patients to generic phenytoin and carbamazepine can be problematic. Fosphenytoin may only be cost effective in certain clinical situations compared with intravenous phenytoin. Seizure control should not be sacrificed on the basis of costs alone, as the major endpoint in treating epilepsy with AEDs is seizure control without adverse effects. Switching AEDs in clinical practice still depends on the individual clinical situation and choosing AED therapy solely on the basis of initial acquisition costs is unlikely to be cost effective in the long-term care of patients with epilepsy. PMID- 15270593 TI - Psychotropic drugs in the treatment of obesity: what promise? AB - Obesity is a chronic and highly prevalent medical condition associated with increased risk for the development of numerous and sometimes fatal diseases. Despite its severity, there are few anti-obesity agents available on the market. Although psychotropic agents are not approved for the treatment of obesity, they have been used by clinicians as a therapeutic tool in daily clinical practice. The purpose of this article is to review the rationale, as well as the evidence, for the potential use of these agents in obesity treatment. Evidence for the efficacy of psychotropic agents in obesity treatment comes from different sources. The first type of evidence is weight loss observed with treatment in clinical trials of patients with neuropsychiatric syndromes (e.g. mood disorders, epilepsy). A recent example of such findings is the weight reduction reported in clinical trials involving obese patients with binge eating disorder. While randomised, controlled trials specifically designed to investigate the weight loss properties of psychotropic agents in obese patients are the most appropriate source of evidence of anti-obesity action, such trials remain scarce. The most studied psychotropic agents in obesity trials are drugs used in the treatment of mood disorders, i.e. mainly antidepressants and antiepileptics. SSRIs (e.g. fluoxetine, sertraline and fluvoxamine) were amongst the first psychotropic agents investigated in the treatment of obesity. Additional data have also been published for other antidepressants (e.g. venlafaxine, citalopram and bupropion) and antiepileptics (e.g. topiramate and zonisamide). Based on the available data for the efficacy of psychotropic agents in obesity and other related conditions, SSRIs may be considered for the management of certain subgroups of obese individuals with comorbid conditions such as depression, binge eating disorder and type 2 diabetes mellitus. In addition, some newer agents, such as bupropion, topiramate and zonisamide, appear to be promising candidates for selective use in the treatment of obesity. However, further studies are needed to define their possible role as new pharmacological options in the treatment of obesity. PMID- 15270594 TI - Management of secondary-progressive multiple sclerosis. AB - The majority of patients with relapse-onset multiple sclerosis (MS) will go on to develop secondary-progressive MS (SPMS) disease, with approximately 50% developing SPMS after 10 years. It remains unknown whether the relapsing and progressive phases of MS differ qualitatively. The pathogenesis of SPMS is poorly understood. The specific role that inflammation plays in disease progression is not well defined. Immunosuppressive therapies, which are capable of reducing or stopping clinical relapses and suppressing MRI activity, generally do not stop disease progression. Recent natural history studies suggest that disease progression occurs regardless of the presence of superimposed relapses. However, poor recovery from clinical relapses does account for the acquisition of disability. Therefore, stopping relapses with appropriate therapy delays the acquisition of disability but does not necessarily delay or prevent the development of SPMS. At present, the only disease-modifying therapies licensed for use in SPMS are interferon-beta-1b in Europe and the US, and mitoxantrone in the US. These agents can only be recommended for patients who continue to have relapses. Symptomatic therapies remain the cornerstone of treatment for patients with SPMS. Delivering high-quality, effective symptomatic therapies requires a multidisciplinary approach. The aim of symptomatic therapies should not only be to reduce neurological impairments but also to decrease disability and handicap and to improve the emotional well-being and health-related quality of life of patients with SPMS. PMID- 15270595 TI - Intranasal medications for the treatment of migraine and cluster headache. AB - Intranasal medications for the treatment of headache have recently received increased attention. This paper reviews intranasal formulations of a variety of available medications (dihydroergotamine mesylate [dihydroergotamine mesilate], sumatriptan, zolmitriptan, butorphanol, capsaicin and lidocaine [lignocaine]) and one experimental medication (civamide, a cis-isomer of capsaicin) for the treatment of migraine and cluster headache. Although the efficacy of intranasal agents varies with the product used, intranasal delivery may be both convenient and more effective than other modes of drug delivery for a variety of reasons: (i) intranasal administration bypasses small bowel gastrointestinal tract absorption, which is often significantly delayed during the acute phase of a migraine attack; (ii) nauseated patients may prefer non-oral formulations as they decrease the chance of vomiting and are more rapidly effective; (iii) intranasal administration causes no pain or injection site reaction and is easier and more convenient to administer than injection or suppository and so may be used earlier in a migraine attack, resulting in better efficacy; (iv) intranasal medication produces the same number or fewer adverse events than injections; and (v) intranasal formulations offer a more rapid onset of action than oral medications, for some of the above reasons and, as such, may be more useful in patients with cluster headache, although this needs to be verified. However, it is important to emphasise that a preference study showed that most patients prefer oral tablets to an intranasal formulation. Also, some nasal preparations have significant adverse effects or are not well absorbed and therefore do not work consistently; others are more challenging to administer as a result of their delivery apparatus. Nevertheless, it is our opinion that nasal preparations increase therapeutic options and may result in faster response times and better efficacy than oral formulations and better patient satisfaction than injectable preparations. PMID- 15270596 TI - Challenge in diagnosis of CD56+ lymphoproliferative disorders: two cases of CD56+CD33+ lymphoma/leukemia. AB - Two cases of CD56+CD33+ leukemia/lymphoma are reported. The patient in case 1 presented with skin rash, diffuse lymphadenopathy, and hepatosplenomegaly. Blasts with monocytoid and lymphoid features were present in the peripheral blood. The tumor cells expressed HLA-DR, CD4, CD33, CD38, and CD56. Cytogenetic analysis revealed del(2)(p13),del(9)(q22),add(6)(q25),add(12)(p12),-13,-18, and -20. The clinicopathologic features were similar to those of blastic natural killer cell leukemia/lymphoma or type 2 dentritic cell leukemia. The patient in case 2 presented with generalized weakness and skin erythema not responding to antibiotics. Circulating blasts with monocytoid features were seen in the peripheral blood. The tumor cells expressed CD7, CD13, CD33, CD38, and CD56, and cytogenetic analysis revealed -5,add(7)(p22),-8, del(10)(p11.2),-12,der(13; 14)(p10;p10),+14,-16,-18,-19, and del(20)(q13.1). The clinicopathologic features were consistent with a myeloid/ natural killer cell precursor acute leukemia. Both disorders are aggressive hematopoietic malignancies that have similar clinical presentation and morphology but differ in immunophenotype and cytogenetic features. PMID- 15270597 TI - A flower-like parasite with bladed petals. PMID- 15270598 TI - Gastric lipid islands. PMID- 15270599 TI - Leishmania in the glomerulus. PMID- 15270600 TI - Pathologic quiz case: a 40-year-old woman with a large pelvic mass, ascites, massive right hydrothorax, and elevated CA 125. Uterine symplastic leiomyoma associated with pseudo-Meigs syndrome and elevated CA 125. PMID- 15270601 TI - Pathologic quiz case: supraclavicular subcutaneous nodule in a 24-year-old woman. Myxoid plexiform fibrohistiocytic tumor without multinucleated giant cells. PMID- 15270602 TI - Pathologic quiz case: renal mass in a 3-week-old infant. Congenital mesoblastic nephroma, mixed type. PMID- 15270603 TI - Pathologic quiz case: right renal mass found incidentally in a 44-year-old woman. Metanephric adenoma. PMID- 15270604 TI - Large esophageal liposarcoma: a case report and review of the literature. AB - Liposarcoma is one of the most common neoplasms of adulthood. However, it is exceedingly rare in the gastrointestinal tract. To our knowledge, only 12 cases occurring in the esophagus have been reported in the world literature to date. We report the case of a 42-year-old man with a pleomorphic liposarcoma arising in the esophageal wall. The morphologic, immunophenotypic, and ultrastructural characteristics are presented, as well as the results of literature review. PMID- 15270605 TI - Sarcomatoid carcinoma of the small intestine: a case report and review of the literature. AB - Sarcomatoid carcinoma of the small bowel is rare; to our knowledge, 19 cases have been reported to date in the English literature under several names. We report an additional case occurring in the jejunum of a 55-year-old man. The tumor was a polypoid 7.5-cm mass, which infiltrated the full thickness of the intestinal wall and the serosa of an adhesed loop of small bowel. On microscopic examination, the neoplasm was composed of sheets of spindle cells; focally, an anaplastic component was present, including tumor giant cells with bizarre nuclei. On immunohistochemical stains, tumor cells were positive for cytokeratin 7, cytokeratin AE1/AE3, vimentin, and focally, epithelial membrane antigen. No staining for cytokeratin 20 was found. Sarcomatoid carcinoma must be kept in mind in the differential diagnosis of malignant spindle cell tumors of the small bowel. As consensus regarding the terminology of these rare tumors is being reached, immunohistochemical stains are essential for accurate diagnosis. PMID- 15270606 TI - Russell body gastritis: an unusual, tumor-like lesion of the gastric mucosa. AB - The case of an 80-year-old woman who presented with epigastric symptoms is reported. Upper gastrointestinal endoscopy displayed Candida esophagitis and a localized swelling of the fundic mucosa. Histologic examination of the gastric biopsy showed a distinctive accumulation of numerous uniform plasma cells filled with so-called Russell bodies. On low-power view, the lesion resembled a neoplastic process due to the marked expansion of the lamina propria with distension of fundic glands. However, immunohistochemistry confirmed a polyclonal pattern of the plasma cells. This unusual reactive lesion of the gastric mucosa has only rarely been described and has been termed Russell body gastritis. PMID- 15270607 TI - Xanthogranulomatous funiculitis and orchiepididymitis: report of 2 cases with immunohistochemical study and literature review. AB - Two patients with xanthogranulomatous inflammation are described, one with involvement of the spermatic cord and the other with 1 testicle and epididymis affected. To our knowledge, only 12 cases of xanthogranulomatous orchiepididymitis have been reported previously, one of which also presented a xanthogranulomatous funiculitis. Clinically, our patients presented with spermatic cord enlargement (case 1) and chronic orchitis that did not respond to treatment with antibiotics (case 2). Histopathologically, both cases showed extensive xanthogranulomatous inflammation with numerous foamy macrophages that were associated with colonies of microorganisms suggestive of actinomyces in case 1. Additionally, Escherichia coli was cultured from the surgical specimen of case 2. The possible underlying pathology may be diabetes in case 1 and phlebitis associated with chronic orchitis in case 2. Differential diagnoses with other lesions that are rich in macrophages, such as malakoplakia, and those testicular neoplasms without serologic tumor markers are discussed. PMID- 15270608 TI - Paneth cell carcinoma of the ampulla of Vater. AB - We describe a Paneth cell carcinoma arising within the ampulla of Vater in a 64 year-old man. The phenotype of virtually all neoplastic cells was consistent with that of Paneth cells, based on routine morphology and their strong positive immunostaining for lysozyme. Additional widespread positive immunostaining for carcinoembryonic antigen and CA 19.9 supports a totipotential cell as the origin of such neoplastic cells. This case, therefore, represents a true Paneth cell carcinoma, as opposed to inclusion of occasional neoplastic Paneth cells into a poorly differentiated adenocarcinoma. This pattern of differentiation is rare, and predictions regarding its ultimate biological behavior and malignant potential must be guarded. PMID- 15270609 TI - A case of acute hemolysis after ceftriaxone: immune complex mechanism demonstrated by flow cytometry. AB - An immune complex mechanism for ceftriaxone sodium- induced severe autoimmune hemolytic anemia has previously been demonstrated using routine blood bank techniques. We describe herein a patient with severe hemolysis that subsided once the drug was discontinued. Serologic techniques demonstrated immune complex mediated ceftriaxone-dependent red cell antibodies. These findings were further supported by the results of flow cytometry, in which a change in basal red cell autofluorescence was seen in the presence of the antibody and the drug. Our case illustrates the adjunctive value of flow cytometry in the diagnosis of ceftriaxone-dependent red cell antibody. PMID- 15270610 TI - Sinusoidal dilatation and congestion in liver biopsy: is it always due to venous outflow impairment? AB - CONTEXT: Impairment of venous outflow manifests as zone 3 sinusoidal dilatation and congestion (SDC) in liver biopsy. However, the finding of SDC is not specific for venous outflow impairment.Objectives.-To determine the specificity of SDC in liver biopsies for venous outflow impairment and to seek an explanation for SDC in patients without clinical or radiologic features of venous outflow impairment. DESIGN: Liver biopsies from 51 patients with sinusoidal dilatation were reviewed. Biopsies from transplant recipients, patients with cirrhosis, and patients with hepatic neoplasms (primary or metastatic) were not included. Clinical records were reviewed for laboratory tests and final clinicopathologic diagnosis. RESULTS: Thirty-four patients (66.7%) had confirmed venous outflow impairment. Of the 17 cases (33.3%) without clinical and/or radiologic evidence of venous outflow impairment, vascular causes were present in 5 cases (9.8%; nodular regenerative hyperplasia in 2 cases and portal vein thrombosis, congenital absence of the portal vein, and sickle cell anemia in 1 case each). Systemic inflammatory disorders were identified in 6 patients (11.8%). These included 2 cases of Castleman disease and 1 each of sarcoidosis, Crohn disease, rheumatoid arthritis, and Still disease. Three patients (5.9%) had tumors without direct involvement of the liver (1 case each of Hodgkin lymphoma, renal cell carcinoma, and pancreatic serous pseudopapillary tumor). In the remaining 3 patients, SDC was identified in wedge liver biopsies performed at the time of surgery, including gastric bypass surgery, cholecystectomy, and splenectomy. No other disease association was apparent in these cases. CONCLUSION: Sinusoidal dilatation and congestion in liver biopsy is associated with venous outflow impairment in two thirds of the cases. In the absence of clinical and/or radiological evidence of venous outflow, diagnostic considerations include other vascular conditions, such as portal vein insufficiency and nodular regenerative hyperplasia. Sinusoidal dilatation and congestion can also occur in the setting of systemic inflammatory diseases, granulomatous disorders, and neoplasms, as well as in wedge biopsies obtained during abdominal surgery. PMID- 15270611 TI - Immunohistochemical characterization of p57Kip2 expression in tetraploid hydropic placentas. AB - CONTEXT: Because there are differences in the origin, morphology, and natural history of hydropic placental villous issues, it is important to identify and document rare specimens that deviate from the diploid complete hydatidiform mole (CM), triploid partial hydatidiform mole (PM), or diploid hydropic abortion (HA). Tetraploid hydropic placentas have rarely been studied. OBJECTIVES: To evaluate the frequency of p57Kip2 protein (p57) expression in tetraploid hydropic placentas and to determine its clinicopathologic significance. DESIGN: Forty hydropic DNA tetraploid placental specimens were evaluated by immunohistochemistry of formalin-fixed tissues, using a monoclonal antibody against p57, a putative paternally imprinted inhibitor gene. DNA ploidy in all cases was analyzed by flow cytometry. RESULTS: Thirty cases were histologically diagnosed as CMs, 10 were HAs, and none were PMs. In all HAs, nuclear p57 was strongly expressed in cytotrophoblasts, intermediate trophoblasts, and villous stromal cells. In contrast, in CMs, p57 expression in cytotrophoblasts and villous stromal cells was either absent (26 cases) or very low (4 cases). Assuming that the degree of molar change roughly correlates with the proportion of paternal chromosomes present, all chromosomes might be paternally derived in all tetraploid CMs and the 10 HAs, including 2 that were karyotyped as 92,XXYY or 90,XXYY,-13,-14, which were presumably due to 2 sets of chromosomes each from paternal and maternal origin. CONCLUSIONS: Expression of p57 is aberrant in tetraploid CMs. This finding is in line with the hypothesis that the loss of p57 is involved in the abnormal development of androgenetic CMs. For the evaluation of a patient with trophoblastic disease, p57 immunostaining is an ancillary diagnostic method that may be used in concert with flow cytometry. PMID- 15270612 TI - Noninvasive carcinoma of the breast: angiogenesis and cell proliferation. AB - CONTEXT: Angiogenesis and the cell proliferation index can predict the prognosis of invasive breast carcinoma; however, little is known of their roles in noninvasive tumor. OBJECTIVE: To investigate the correlation of microvessel density and cell proliferation index with other histologic parameters (histologic type, nuclear grade, and mitotic count) in 65 cases of noninvasive carcinoma of the breast. DESIGN: Formalin-fixed, paraffin-embedded tissues from 65 cases of carcinoma in situ of the breast were immunostained with antibody against factor VIII antigen and proliferation-associated nuclear antigen MIB-1. The microvessel density was measured by counting the total number of microvessels around the carcinoma in situ per 10 low-power microscopic fields. The cell proliferation index was calculated by counting MIB-1-positive nuclei in 100 tumor cells. A chi2 test and Spearman rank correlation test were used for statistical analysis. RESULTS: The microvessel density and cell proliferation index of comedo-type, high-nuclear-grade ductal carcinomas in situ are significantly higher than those of either noncomedo type ductal carcinomas in situ or lobular carcinoma in situ (P <.001). CONCLUSIONS: Angiogenesis and the cell proliferation index are active biological processes and may be considered as markers to separate low- and high risk patients with noninvasive breast carcinomas. PMID- 15270613 TI - Corrections of clinical chemistry test results in a laboratory information system. AB - CONTEXT: The recently released reports by the Institute of Medicine, To Err Is Human and Patient Safety, have received national attention because of their focus on the problem of medical errors. Although a small number of studies have reported on errors in general clinical laboratories, there are, to our knowledge, no reported studies that focus on errors in pediatric clinical laboratory testing. OBJECTIVE: To characterize the errors that have caused corrections to have to be made in pediatric clinical chemistry results in the laboratory information system, Misys. To provide initial data on the errors detected in pediatric clinical chemistry laboratories in order to improve patient safety in pediatric health care. DESIGN: All clinical chemistry staff members were informed of the study and were requested to report in writing when a correction was made in the laboratory information system, Misys. Errors were detected either by the clinicians (the results did not fit the patients' clinical conditions) or by the laboratory technologists (the results were double-checked, and the worksheets were carefully examined twice a day). No incident that was discovered before or during the final validation was included. On each Monday of the study, we generated a report from Misys that listed all of the corrections made during the previous week. We then categorized the corrections according to the types and stages of the incidents that led to the corrections. RESULTS: A total of 187 incidents were detected during the 10-month study, representing a 0.26% error detection rate per requisition. The distribution of the detected incidents included 31 (17%) preanalytic incidents, 46 (25%) analytic incidents, and 110 (59%) postanalytic incidents. The errors related to noninterfaced tests accounted for 50% of the total incidents and for 37% of the affected tests and orderable panels, while the noninterfaced tests and panels accounted for 17% of the total test volume in our laboratory. CONCLUSION: This pilot study provided the rate and categories of errors detected in a pediatric clinical chemistry laboratory based on the corrections of results in the laboratory information system. A direct interface of the instruments to the laboratory information system showed that it had favorable effects on reducing laboratory errors. PMID- 15270614 TI - Evaluation of add-on testing in the clinical chemistry laboratory of a large academic medical center: operational considerations. AB - CONTEXT: Physicians frequently request that additional tests be performed on an existing specimen (add-ons). In our institution, add-ons comprise approximately 1% of the specimen volume and require a disproportionate number of employees. Not only are add-on tests time-consuming and expensive, but storing routine specimens for 7 days in anticipation of add-ons consumes valuable laboratory space. DESIGN: One hundred sixty add-on tests during a 1-week period were reviewed. OBJECTIVES: To analyze the pattern of add-on testing and determine methods to improve laboratory operations. RESULTS: All add-on tests were ordered within 24 hours of receipt of the original specimen, even though specimens were retained for 7 days. At our institution, 1.5 full-time equivalents are required to complete add-on testing, which accounts for less than 1% of the specimen volume. The most common add-on tests recorded during the study period were hepatic and electrolyte/renal/glucose panels. The medicine service ordered more than 60% of the add-on tests. Five percent of add-on tests were caused by a lack of order communication, 64.7% of cardiac marker add-ons were not ordered according to the chest pain protocol, and certain ordering patterns were present. CONCLUSIONS: Routine specimens do not need to be retained for 7 days to accommodate add-on tests. Decreasing the storage time to 2 days would save space, while still maintaining regulatory compliance. Order communication with the laboratory, educating physicians about chest pain protocols, and instituting admission laboratory panels would decrease the number of add-ons in our hospital. This change would translate into a reduction in laboratory expenses and an improvement in operations. PMID- 15270615 TI - Loss of heterozygosity patterns of sclerosing hemangioma of the lung and bronchioloalveolar carcinoma indicate a similar molecular pathogenesis. AB - CONTEXT: The histogenesis and origin of sclerosing hemangioma (SH) of lung were uncertain for many years. Many immunohistochemical, ultrastructural, and recent molecular studies support the hypothesis that SH is a neoplasm originating from the cells of the terminal lobular unit, similar to the nonmucinous variant of bronchioloalveolar carcinoma (BAC). Most cases of SH are benign, but they can metastasize to the regional lymph nodes. OBJECTIVE: To compare the patterns of allelic loss of tumor suppressor genes in SH and BAC by microdissection-based genotypic analysis. DESIGN: Microdissection-based loss of heterozygosity analysis of 9 cases of SH and 14 cases of BAC, using a panel of 7 polymorphic microsatellite markers located on 1p, 5q, 9p, 10q, and 17p. Microsatellite marker and chromosomal arm-based fractional allelic loss (FAL) were calculated in each case. RESULTS: Our results showed similar patterns of allelic loss between the 2 groups of tumors on an individual case basis. Chromosomal arms 5q and 10q showed frequent allelic loss in SH (66.7% and 62.5%, respectively), whereas in BAC, chromosomal arm 17p (52.6%) was frequently affected. A statistically significant difference in allelic loss between SH and BAC was located only on chromosomal arm 5q (P =.04). Microsatellite marker D5S615 was significantly more frequently affected in SH than in BAC (66.7% vs 28.6%; P =.04). CONCLUSION: Our molecular data support the hypothesis of common origin of SH and BAC. A putative tumor suppressor gene that might play a role in tumorigenesis of SH may be located on the chromosomal arm 5q. PMID- 15270616 TI - Renal pathologic spectrum in an autopsy series of patients with plasma cell dyscrasia. AB - CONTEXT: Renal dysfunction in plasma cell dyscrasias is common. It is the second most common cause of death in patients with myeloma. OBJECTIVE: We evaluated 77 sequential autopsies performed on patients dying from complications of plasma cell dyscrasias during an 11-year period at the University of Arkansas for Medical Sciences. These consisted of 15% of all the autopsies performed during this time. DESIGN: The kidneys were evaluated by light microscopy using hematoxylin-eosin-stained sections as well as Congo red and thioflavin T stains when amyloidosis was in the differential diagnosis. Immunofluorescence was performed on selected cases. RESULTS: The most common lesion identified was cast nephropathy (30%). Other findings included acute tubulopathy, AL-amyloidosis, light chain deposition disease, tubulointerstitial nephritis associated with monotypic light chain deposits, thrombotic microangiopathy, renal infarction, fungal infection, and plasma cell tumor nodules. Autolysis, an expected finding in autopsy evaluations, was significant in 25 cases. CONCLUSIONS: Renal lesions are heterogeneous in these patients. In some cases, combined pathologic lesions were noted. Myeloma cast nephropathy predominated among all the renal lesions noted. PMID- 15270617 TI - Lipoprotein lipase gene polymorphism and lipid profile in coronary artery disease. AB - CONTEXT: Lipoprotein lipase (LPL) plays a central role in lipid metabolism, hydrolyzing triglyceride in chylomicrons and very-low-density lipoproteins. The PvuII polymorphic variant of LPL gene is common and might affect risk of coronary artery disease (CAD). OBJECTIVE: Our aim was to determine whether LPL- PvuII polymorphism can be considered to be an independent risk factor or a predictor for CAD in Turkish subjects. DESIGN: We used polymerase chain reaction and restriction enzyme digestion to determine the distribution of the previously described C-->T transition that causes a PvuII polymorphism in intron 6 among healthy blood donors of Turkish origin and among angiographically confirmed CAD patients with comparable ethnic backgrounds. RESULTS: For the PvuII genotypes, within the CAD group (n = 80), the +/- genotype was found in 39 individuals (48.8%), whereas 25 (31.3%) carried the +/+ genotype, and 14 (17.5%) carried the /- genotype. Within the control group (n = 49), the -/- genotype was found in 19 individuals (38.8%), 16 (32.7%) carried the +/- genotype, and 14 (28.6%) carried the +/+ genotype. The genotype frequency distribution was significantly different (P =.049) in the CAD and control study groups. The most frequent genotype among CAD patients was +/-; this genotype was more frequent in patients than in control subjects. However, the -/- genotype was more prevalent in the control group. Lipoprotein lipase-PvuII polymorphism was found to be associated with fasting total cholesterol and low-density lipoprotein cholesterol levels. The +/+ genotype was found to have higher levels of total cholesterol and low-density lipoprotein cholesterol in both the CAD and control groups. CONCLUSION: There was a difference in the distribution of LPL-PvuII genotypes between the healthy subjects and the patients with CAD. Lipoprotein lipase-PvuII polymorphisms were not detected as independent risk factors for CAD in this study group, but had associations with lipid levels. PMID- 15270618 TI - A clinicopathologic evaluation of follicular lymphoma grade 3A versus grade 3B reveals no survival differences. AB - CONTEXT: The World Health Organization classification recommends categorizing grade 3 follicular lymphomas based on the presence of centrocytes (grade 3A) or of sheets of centroblasts (grade 3B). The clinical significance of this practice is not known. OBJECTIVE: To determine whether grade 3 follicular lymphoma subtype is associated with prognosis. DESIGN: Multi-institutional retrospective case series. MAIN OUTCOME MEASURE: Overall survival. RESULTS: Forty-five cases of grade 3 follicular lymphoma without diffuse large B-cell lymphoma were studied (35 cases of grade 3A, 10 cases of grade 3B) from 21 men and 24 women (median age, 67 years; mean age, 63.8 years; range, 26-86 years). Follow-up information from the time of diagnosis was available in all patients, with a median follow-up time of 24 months (mean, 34 months; range, 2- 115 months). Treatment information was available in 40 patients. There was no difference in age (P =.45, Wilcoxon test) or stage (P =.76, Fisher exact test) between patients with follicular lymphoma of grade 3A or grade 3B. Furthermore, the Cochran-Armitage test for trends showed no evidence that the proportion of patients with follicular lymphoma grade 3A or 3B increased or decreased with increasing stage at presentation. Kaplan-Meier analysis showed a median overall survival of 44 months from the time grade 3 follicular lymphoma was diagnosed, with no significant difference between cases diagnosed as grade 3A or grade 3B (P =.14, log-rank test). Univariable Cox proportional hazards modeling showed no evidence that an anthracycline-containing chemotherapy regimen or history of lower grade follicular lymphoma affected overall survival. CONCLUSIONS: In this retrospective series, subclassification of grade 3 follicular lymphoma into type 3A and 3B categories had limited clinical and prognostic significance. However, the study was limited by lack of statistical power. Since morphology often provides clues for progress in defining biologic differences, subtyping may still be useful, particularly in the setting of prospective clinical studies. PMID- 15270619 TI - Primary small cell neuroendocrine carcinoma of the vagina: a clinicopathologic study. AB - CONTEXT: Primary small cell neuroendocrine carcinoma of the vagina is extremely rare, and its clinical behavior is aggressive. To our knowledge, 22 patients with this tumor have been reported in the English literature to date. OBJECTIVE: To investigate 3 patients with this tumor clinically and pathologically. DESIGN: The pathology database at the University of Texas Medical Branch at Galveston was searched, and 3 cases of primary small cell neuroendocrine carcinoma of the vagina were found. The histologic, immunohistochemical, and ultrastructural profiles of the tumors were investigated. The medical charts of the patients were reviewed, and the patients were followed up. PATIENTS: Women with the diagnosis of primary small cell neuroendocrine carcinoma of vagina. RESULTS: All 3 patients presented with advanced disease, and 2 patients died within 4 months of the initial diagnosis. One 38-year-old patient was newly diagnosed, and her clinical outcome had not yet been determined. The histologic features of all 3 tumors were similar to those of their pulmonary counterpart. All cases were positive for cytokeratin, chromogranin A, and synaptophysin. The expression pattern of thyroid transcription factor 1 was examined in all 3 patients, of whom 2 were negative and 1 was positive with negative clinical and radiologic thyroid or pulmonary findings. Ultrastructural evaluation showed scattered intracytoplasmic electron dense neurosecretory granules. CONCLUSION: Primary small cell neuroendocrine carcinoma of the vagina has histologic, immunohistochemical, and ultrastructural features similar to those of its pulmonary counterpart. Because thyroid transcription factor 1 can be positive, it should not be used to differentiate primary from metastatic disease. The current therapies have usually resulted in poor outcomes, and new therapeutic modalities should be explored. PMID- 15270621 TI - Reagent strips may not detect Staphylococcus epidermidis contamination of platelet concentrates. PMID- 15270623 TI - The challenge of using autopsy information for quality improvement. PMID- 15270626 TI - Pathologic quiz case: a patient with Down syndrome presenting with "idiopathic" pericarditis. Primary pericardial malignant mesothelioma. PMID- 15270625 TI - Posttransplant primary cutaneous Ki-1 (CD30)+/CD56+ anaplastic large cell lymphoma. AB - An anaplastic large cell lymphoma that was negative for Epstein-Barr virus and positive for Ki-1 (CD30) presented as a polypoid scalp mass in a 56-year-old man 16 years after renal transplantation. The lymphoma was of the CD4+ cytotoxic T cell lineage, and the tumor cells also expressed CD56. Despite reduction in the dose of immunosuppression and localized radiotherapy, the tumor had rapidly progressed to involve the soft tissue of the right hand. Systemic chemotherapy induced complete regression of the soft tissue lesion. This case illustrates that posttransplant primary cutaneous CD30+ anaplastic large cell lymphomas may assume an aggressive clinical course but can still be controlled by systemic chemotherapy. PMID- 15270627 TI - Pathologic quiz case: a 35-year-old man with hematuria. Paraganglioma involving the prostate. PMID- 15270628 TI - The challenge of new and emerging lyssaviruses. PMID- 15270630 TI - Developmental immunology and vaccines: cellular immune development and future vaccine strategies. AB - This section deals with how new knowledge of the development of cellular immunity in the neonatal period informs vaccine development and reviews the immune responses of the most vulnerable group of newborns, those born prematurely, to vaccines. It describes how the development of future vaccine strategies relies on a detailed understanding of immune ontogeny and the potential consequences of intervention on developing cellular immune responses. PMID- 15270631 TI - Developmental immunology and vaccines: immune responses to vaccines in premature infants. AB - A comparatively small number of studies have assessed the safety, immunogenicity, efficacy and duration of immune responses in preterm infants compared with term infants for routinely recommended childhood immunizations. PMID- 15270632 TI - Vaccines for neonatal viral infections: hepatitis B vaccine. AB - In many parts of the world, intervention in the neonatal period is required for prevention of hepatitis B virus infection and its consequences. PMID- 15270633 TI - Vaccines for neonatal viral infections: towards a live respiratory syncytial virus vaccine:a study in risk. AB - There is risk attached to the development of respiratory syncytial virus vaccines, live attenuated or otherwise, but without the acceptance of this risk by manufacturers, health providers and the public, the conclusion of successful Phase III trials may lie in the distant future. PMID- 15270634 TI - Vaccines for neonatal viral infections: rotavirus. AB - We had hoped by now to have a rotavirus vaccine for the National Center for Immunization Research and Surveillance of Vaccine Preventable Diseases (NCIRS) to concern itsel f with. One day in the near future, we hope that Margaret's 'retirement' will be interrupted by good news about the prevention of another miserable disease. PMID- 15270635 TI - Vaccines for neonatal viral infections: vaccines to prevent neonatal herpes simplex virus infection. AB - Margaret Burgess has made many contributions to the study of congenital and perinatal infections. Her mentorship of young researchers is her long-term legacy to the development of vaccines to prevent infections in utero and in the newborn period. PMID- 15270636 TI - Vaccines for other neonatal infections: neonatal BCG vaccination against tuberculosis. AB - This section focuses on current progress in the use of vaccines against a range of other infectious agents, both bacterial (pertussis, pneumococcus and group B streptococcus) and mycobacterial (bacille Calmette-Guerin) to prevent neonatal infection. PMID- 15270637 TI - Vaccines for other neonatal infections: are group B streptococcal infections vaccine-preventable? AB - Preliminary studies suggest that a pentavalent group B streptococcus conjugate vaccine, given in a single dose at 32-34 weeks gestation, would prevent approximately 90% of early and late onset neonatal and most postpartum maternal group B streptococcus infections. PMID- 15270638 TI - Vaccines for other neonatal infections: vaccination strategies for the prevention of neonatal pertussis. AB - Current progress in the use of vaccines against a range of infectious agents, both bacterial (pertussis, pneumococcus and group B streptococcus) and mycobacterial (bacille Calmette-Guerin), to prevent neonatal infection are reviewed by Professors Gilbert, Britton and McIntyre and Dr Peter Richmond. PMID- 15270639 TI - Vaccines for other neonatal infections: pneumococcal disease: a major global health problem of young children. AB - Provided the high vaccine costs for pneumococcal conjugate vaccines can be addressed for the poorest countries, neonatal PCV immunization offers the hope of decreasing the significant morbidity that occurs in these vulnerable populations. PMID- 15270640 TI - Trial watch: Xenova's TA-NIC vaccine shows promise. PMID- 15270641 TI - Trial watch: SARS vaccine enters Phase I trials. PMID- 15270642 TI - Phacilitate Vaccine Forum, Spring 2004. PMID- 15270643 TI - Vaccines and vaccination strategies. PMID- 15270644 TI - TRICOM: enhanced vaccines as anticancer therapy. AB - Renewed interest in immune therapies for the treatment of cancer has sparked a number of different immune stimulation approaches. These include antibody based, dendritic cell-based and vaccine-based therapies. The poxvirus-based vaccines incorporating a TRIad of COstimulatory Molecules (TRICOM) will be reviewed here. Through stimulation of cancer-specific T-cell responses, researchers have demonstrated interesting clinical efficacy in addition to the high safety profile in clinical trials. PMID- 15270645 TI - Heat shock protein-based cancer vaccines. AB - Heat shock proteins (HSPs) exist ubiquitously across all species and function as chaperones stabilizing and delivering peptides. Tumor-derived HSP-peptide complex has been known to induce immunity against the original tumor in preclinical studies. HSP-based vaccines work across tumor types and bypass the need for identifying the responsible peptide(s) for inducing immunity. These vaccines are tumor- and patient-specific in that they capture the tumor cells' fingerprints. HSP-based vaccines have been studied in early phase clinical trials, mostly using HSP glycoprotein 96, for various types of malignancies including melanoma, renal cell carcinoma, gastric cancer, pancreatic cancer, low-grade lymphoma, colorectal cancer and chronic myelogenous leukemia. All showed minimal toxicity and potential efficacy. Phase III studies for melanoma and renal cell carcinoma are ongoing. HSP-based vaccines are a novel vaccine preparation with a promising role in cancer management. Further studies to determine the administering strategy and specific indication are warranted. PMID- 15270646 TI - Current status of the Arilvax; yellow fever vaccine. AB - Yellow fever is a viral hemorrhagic fever that involves dysfunction of the liver and kidneys. There is no antiviral therapy available and vaccination is a major strategy in the control of this important public health problem. Yellow fever vaccine is a live attenuated vaccine, strain 17D, which is very safe and efficacious and induces long-term protective immunity in vaccinees. The vaccine is produced by six manufacturers worldwide, including Arilvax by Chiron vaccines (CA, USA) that is marketed in at least ten countries. Acambis (MA, USA) have acquired the rights to sell and market Arilvax in the USA, and have undertaken three clinical trials to obtain data for a Biological License Application that should proceed in 2005. PMID- 15270647 TI - New and emerging vaccination strategies for prevention and treatment of dermatological diseases. AB - Accelerated by the rapid advancements of our understanding of the molecular and cellular pathology of diseases and of the components and mechanisms of cellular and humoral immune responses, new vaccination strategies are being developed and explored for treatment and prevention of infectious diseases, cancer, autoimmune disorders and allergies. Many newly developed vaccination strategies are already in clinical trials, some with very promising results. Although most of these strategies are still at very early stages of their development, it is foreseeable that vaccination will evolve to play an important role in prevention, treatment and management of all the above classes of diseases. PMID- 15270648 TI - Cost-effectiveness studies of pneumococcal conjugate vaccines. AB - The 7-valent pneumococcal conjugate vaccine is licensed in many countries for the prevention of pediatric pneumococcal disease. The vaccine is known to be highly immunogenic in infants and young children, and has been shown to be efficacious not only in decreasing disease in pediatric age groups but also in adults through herd immunity. Cost-effectiveness analyses of this vaccine have been performed in a number of countries. The present review compiles, summarizes and critiques these analyses. The range of values for cost-effectiveness, as measured in cost per life-years gained, in the studies reviewed, ranges from 14,000 US dollars to 147,000 US dollars with one outlier at 504,000 US dollars. For cost per quality adjusted life years the range is 26,000 US dollars to 66,000 US dollars. Recommendations for the use of the vaccine will take account not only of these ratios but also of the absolute burden of disease. Performing cost-effectiveness analyses for healthcare interventions in infants and children is one means of redressing inequalities. PMID- 15270649 TI - Influenza: current evidence and informed predictions. AB - Every winter there are sharp rises in medical visits, hospitalizations and deaths from acute respiratory illness worldwide. Influenza is an important cause of these and is the only common viral respiratory pathogen with licensed vaccines available that are safe and effective in preventing disease. Over 50 countries have national vaccination programs focusing on the elderly population and those at high risk. In the USA, healthy children are also now offered the vaccine; routine immunization of the young, who are similarly vulnerable to hospitalization as the old, is likely to become universal. There remains a need for further improvement in vaccine effectiveness (both in the short- and long term), vaccine administration and compliance. Recent technological developments raise the prospect of using new vaccine types, such as live attenuated and Vero cell-cultured vaccines, that are easier to administer and may induce broader immunity. PMID- 15270651 TI - Bacterial viruses as human vaccines? AB - Bacteriophages (or phages) are viruses of bacteria, consisting of nucleic acid packaged within a protein coat. In eukaryotic hosts, phages are unable to replicate and in the absence of a suitable prokaryotic host, behave as inert particulate antigens. In recent years, work has shown that whole phage particles can be used to deliver vaccines in the form of immunogenic peptides attached to modified phage coat proteins or as delivery vehicles for DNA vaccines, by incorporating a eukaryotic promoter-driven vaccine gene within their genome. While both approaches are promising by themselves, in future there is also the exciting possibility of creating a hybrid phage combining both components to create phage that are cheap, easy and rapid to produce and that deliver both protein and DNA vaccines via the oral route in the same construct. PMID- 15270650 TI - Antigen delivery systems. AB - Many vaccine candidates are highly purified, sometimes monomeric antigens and as a result, not very immunogenic. Antigen delivery systems optimize the presentation of antigens. They also play a major role in solving the problem of there being an increasing number of vaccines but limited opportunities in which to include these vaccines in immunization programs. The number of injections is restricted and combining vaccines may lead to immunological and physicochemical incompatibility. In this review, the current status with respect to parenteral and mucosal delivery systems is discussed. These include lipid-based systems such as liposomes and immunostimulating complexes, as well as polymeric microspheres. In addition, developments in needle-free, dermal delivery devices such as jet injectors, microneedles and patches are presented. PMID- 15270652 TI - Progress toward development of an inhalation vaccine against botulinum toxin. AB - The looming threat of bioterrorism has enhanced interest in the development of vaccines against agents such as botulinum toxin. This in turn has stimulated efforts to create vaccines that are effective by the oral and inhalation routes. Recently, considerable progress has been made in creating an inhalation vaccine against botulism. This work stems from the discovery that a polypeptide that represents a third of the toxin molecule retains the ability to be adsorbed from the airway and to evoke an immune response but retains none of the adverse effects of the native toxin. Interestingly, this polypeptide can also serve as a carrier molecule in the creation of inhalation vaccines against other pathogens. PMID- 15270653 TI - Intratumoral delivery of genes: a new weapon against cancer? PMID- 15270655 TI - Demethylation of DNA by decitabine in cancer chemotherapy. AB - Genes involved in all aspects of tumor development and growth can become aberrantly methylated in tumor cells, including genes involved in apoptosis and cell cycle regulation. Decitabine, 2'-deoxy-5-azacytidine, can inhibit DNA methyltransferases and reverse epigenetic silencing of aberrantly methylated genes. Nucleoside DNA methyltransferase inhibitors, such as decitabine, have been reported to have antitumor activity, especially against hematologic malignancies. Such demethylating agents have been proposed to reactivate tumor suppressor genes aberrantly methylated in tumor cells, leading to inhibition of tumor growth. An important consequence of this is that, unlike conventional cytotoxic agents, it may be best to use such drugs at concentrations lower than the maximum tolerated dose and in a manner dependent on their demethylating activity. Furthermore, synergistic activity with other types of investigational epigenetic therapies and existing chemotherapies opens the possibility of rational combinations and scheduling of these agents based on their biologic activity. PMID- 15270656 TI - Pemetrexed: a novel antifolate agent enters clinical practice. AB - Pemetrexed (Alimta, Eli Lilly) is a multitargeted antifolate that inhibits at least three enzymes in the nucleic acid synthetic pathways. The US Food and Drug Administration recently approved pemetrexed, in combination with cisplatin, for the first-line treatment of advanced malignant pleural mesothelioma. Moreover, pemetrexed was recently shown to be as efficacious as docetaxel (Taxotere, Aventis) in the second-line treatment of non-small cell lung cancer, and its toxicity profile was preferable. The main toxicity seen with pemetrexed is myelosuppression, which is considerably reduced by coadministration of folic acid and vitamin B12. Multiple Phase II clinical trials have demonstrated that pemetrexed has promising single-agent activity in many other solid tumors, including head and neck, breast and colorectal cancers. Combination regimens consisting of pemetrexed and other chemotherapeutics or novel molecular-targeted agents are currently under investigation. Future studies will better define and likely expand the role of pemetrexed for the treatment of cancer. PMID- 15270657 TI - Raloxifene and its role in breast cancer prevention. AB - Raloxifene (Evista, Eli Lilly), a selective estrogen receptor modulator (SERM) and ligand for the estrogen receptor (ER), competes with endogenous estrogen for ER binding. Raloxifene is approved for the prevention and treatment of osteoporosis, and shows promise as a breast cancer prevention drug. Raloxifene may be a preferred agent over tamoxifen due to its side-effect profile; in particular, it does not stimulate the endometrium and is not associated with endometrial cancer. The mechanisms for the differential tissue effects of raloxifene compared with other SERMs are not completely understood; the roles of ERalpha and -beta, classic and alternative signaling pathways, and drug conformation are discussed in this review. The utility of raloxifene will depend on the outcome of trials that are now underway, as well as acceptance by high risk women and their healthcare practitioners. PMID- 15270658 TI - Anagrelide: an update on its mechanisms of action and therapeutic potential. AB - Thrombocytosis is an increasingly recognized clinical problem due to the widespread availability of automated cell counters. While reactive thrombocytosis does not require any therapeutic intervention, clonal thrombocytosis may require therapy to prevent thrombohemorrhagic complications. The clinician has a number of therapeutic options available when confronted with a patient having clonal thrombocytosis. One of these agents is anagrelide (Agrylin, Bristol-Myers Squibb). In this drug profile, a synopsis of the available data on this agent and its role in the control of thrombocytosis will be provided. The main side effects of the medication are discussed, as well as the potential future developments in the field. PMID- 15270659 TI - COX-2 and its inhibition as a molecular target in the prevention and treatment of lung cancer. AB - Lung cancer is the leading cause of cancer death in the USA. Conventional therapy using chemotherapy, radiation therapy, and a combination of the two, has yielded modest improvement in patient outcome. Dysfunction and dysregulation of many molecular processes and signaling pathways are involved in the development and growth of malignant lung tumors, and in conferring resistance to standard cancer treatments. Cyclooxygenase (COX)-2, an enzyme involved in prostaglandin production in pathologic states, is often overexpressed in premalignant and malignant lesions. Overexpression of COX-2 in lung cancer is associated with more aggressive biologic tumor behavior and adverse patient outcome. In preclinical studies, inhibition of this enzyme with selective COX-2 inhibitors enhances tumor response to radiation and chemotherapeutic agents. These findings quickly led to clinical studies. Phase I and II clinical trials of the combination of selective COX-2 inhibitors with radiotherapy, chemotherapy, or both in patients with lung cancer have been initiated and some preliminary results are available. In this review, the relationship between overexpression of COX-2 and lung cancer, the antitumor effect of selective COX-2 inhibitors, and the rationale for using selective COX-2 inhibitors combined with radiotherapy and chemotherapy, will be described. Current clinical protocols and preliminary findings will also be summarized. PMID- 15270660 TI - Role of FDG-PET in the diagnosis and management of lung cancer. AB - Positron emission tomography (PET) using [(18)F]-2-deoxy-2-fluoro-d-glucose (FDG) has emerged as a valuable diagnostic modality in patients with non-small cell lung cancer (NSCLC). Data in the literature show that the addition of FDG-PET definitely alters clinical management in patients with potentially resectable NSCLC by adequately staging the mediastinum and detecting previously unknown distant metastases. Thus, the number of noncurative thoracotomies and unnecessary mediastinoscopies is reduced. Furthermore, there is increasing evidence that FDG PET will change radiation treatment planning by defining a biologic treatment volume, incorporating unsuspected additional locoregional disease, and avoiding overtreatment by identifying computerized tomography abnormalities as benign. For follow-up during systemic therapy, early FDG-PET appears to be predictive for the response to therapy. However, before FDG-PET-induced changes in patient management can be incorporated into clinical practice both for radiation treatment planning and chemotherapy, technical issues must be resolved, validation studies should be performed and, most importantly, randomized trials are necessary to evaluate the effect of FDG-PET on patient outcome parameters. PMID- 15270661 TI - Combination epidermal growth factor receptor inhibition and radical radiotherapy for NSCLC. AB - Non-small cell lung cancer (NSCLC) remains the most common cause of cancer related death in the developed world. Despite advances in therapy with conventional modalities, over 85% of patients will die from their disease within 5 years of diagnosis. For patients with inoperable lung cancer, the addition of chemotherapy to radical radiotherapy yields a small but significant 10% survival benefit at 3 years. However, the systemic toxicity of chemotherapy is common and may be severe. Over the past 20 years, dramatic improvements in our understanding of the molecular etiology of cancer have enabled the development of novel targeted therapies. Overexpression of the epidermal growth factor receptor (EGFR) in lung cancer correlates with an aggressive disease course and poor tumor response to radiotherapy. Strategies to inhibit this molecular switch have become a focus for drug development. Preclinical efficacy has been repeatedly demonstrated with anti-EGFR monoclonal antibodies and small molecule tyrosine kinase inhibitors, and responses have been documented in the clinic with acceptable toxicity. Phase III trials combining EGFR tyrosine kinase inhibitors with radical chemoradiation are recruiting at present. This review addresses the current challenges of discovering how best to use these new anticancer therapies, with particular emphasis on the enhancement of existing therapeutic strategies such as radical radiotherapy, factors relating to patient selection and prediction of clinical response. PMID- 15270662 TI - Evolving role of chemoradiation in the adjuvant treatment of gastric cancer. AB - The recently reported Intergroup trial (INT0116) has established combined chemoradiation as an important component of adjuvant therapy that should be considered for high-risk, completely resected adenocarcinoma of the stomach. However, implementation of this treatment has posed several problems with respect to the toxicity of the treatment, optimal chemotherapy regimen and optimal mode of radiotherapy delivery. At the same time, it has also provided new opportunities to build on the Intergroup results by attempting to improve treatment efficacy and reduce treatment toxicity. Strategies currently being developed include the use of newer generation cytotoxic agents, employing modern conformal techniques of radiation delivery and the use of preoperative chemoradiation. PMID- 15270663 TI - Current clinical management of gastrointestinal stromal tumors. AB - Gastrointestinal stromal tumors (GIST) are defined as c-KIT-positive mesenchymal neoplasias located in the gastrointestinal tract and abdomen, most of which present an activating KIT mutation, a fundamental step in the development of disease. However, recent studies reported a small subgroup of KIT-negative GIST, in which platelet-derived growth factor receptor A, protein kinase C-tau, and FLJ10261 expression was detected. Imatinib (Gleevec, Novartis) is an orally administered competitive inhibitor of the tyrosine kinase domain of receptors such as KIT, ABL, and BCR-ABL fusion proteins, and the platelet-derived growth factor receptor. Phase I-III clinical trials have demonstrated the efficacy of imatinib in the treatment of metastatic GIST. However, the optimal dose and role of imatinib in an adjuvant or neoadjuvant setting have yet to be defined. Therefore, further studies investigating the mechanism of resistance to imatinib in patients with GIST are warranted. PMID- 15270664 TI - Role of FDG-PET in the diagnosis and treatment of colorectal liver metastases. AB - Positron emission tomography (PET) using [(18)F]-2-deoxy-2-fluoro-d-glucose (FDG) has emerged as a promising diagnostic modality in recurrent colorectal cancer. Data in the literature show that the addition of FDG-PET changes disease management in up to 30% of patients with potentially resectable liver metastases, mainly by detecting previously unknown extrahepatic disease. Furthermore, FDG-PET is useful in the follow-up of patients who underwent surgical procedures of the liver, since it is exquisitely sensitive in detecting residual or relapse malignancy in scarred liver tissue following both resection and local ablative techniques. For follow-up during systemic therapy, early FDG-PET appears predictive for response to therapy. However, at present, the available data are insufficient to justify the FDG-PET-driven management of patients treated with chemotherapy. FDG-PET and computerized tomography are complimentary techniques in staging and restaging patients with advanced colorectal cancer. The combination of these two modalities significantly impacts upon patient management. PMID- 15270665 TI - Computerized tomography colonography. AB - First introduced a decade ago, computerized tomography (CT) colonography (virtual colonoscopy) is emerging as an important radiologic investigation for colorectal neoplasia, with diagnostic performance likely exceeding barium enema and comparable with optical colonoscopy. Employing state-of-the-art multislice technology, CT colonography allows a complete examination of the colon and surrounding organs in less than 30 seconds. This article reviews current techniques, indications, comparison with existing technologies, and diagnostic performance. Although already widely disseminated, important future developments, such as prepless bowel cleansing (laxative free) and computer-aided diagnosis, may establish CT colonography as the preferred first-line, whole-colon investigation. PMID- 15270666 TI - Developments in combination chemotherapy for colorectal cancer. AB - In the last 5 years, major advances have occurred in the treatment of colorectal cancer. Following a period of over 30 years in which 5-fluorouracil was the only proven treatment for colorectal cancer, survival for patients with metastatic colorectal cancer has nearly doubled following the introduction of irinotecan (Campto, Aventis) and oxaliplatin (Eloxatin, Sanofi-Synthelabo). Furthermore, recent studies have demonstrated additional improvements in response and survival when combination chemotherapy drugs are administered with biologic agents that target angiogenesis and tumor growth pathways. The benefit of these newer drugs is now being realized in the adjuvant setting, where the addition of oxaliplatin to infusional 5-fluorouracil/leucovrin has led to improvements in disease-free survival. Further adjuvant studies are testing the benefit of the addition of biologic therapies to oxaliplatin and irinotecan-based combination chemotherapy. PMID- 15270667 TI - Neoadjuvant chemotherapy in ovarian cancer. AB - Primary radical tumor debulking followed by platinum/taxane-based chemotherapy is considered standard for advanced stage ovarian carcinomas. The extent of postoperative residual disease is the most important prognostic factor. However, complete tumor resection is achieved in only 40-50% of advanced ovarian cancers. For the remaining patients, who have an unfavorable prognosis, the concept of neoadjuvant or primary chemotherapy followed by interval laparotomy has emerged. Two different strategies are pursued. One is to administer several courses of neoadjuvant chemotherapy in order to downstage the tumor prior to primary debulking surgery. The other is to administer chemotherapy after suboptimal debulking surgery to optimize cytoreduction during interval laparotomy. Numerous retrospective studies demonstrated that neoadjuvant chemotherapy followed by primary debulking surgery is a feasible and safe approach. It is becoming increasingly evident that the selection of appropriate patients is crucial. Some studies demonstrated that the volume of ascites proved to be an easily measurable biomarker that allowed prediction of tumor resectability. However, further investigations are needed to better define patients who will benefit from neoadjuvant chemotherapy. Despite promising results, neoadjuvant chemotherapy must still be considered experimental. Therefore, the potential advantages of neoadjuvant chemotherapy need to be confirmed in prospective randomized studies. PMID- 15270668 TI - Role of TGF-beta in cancer and the potential for therapy and prevention. AB - Transforming growth factor (TGF)-beta is a naturally occurring potent inhibitor of cell growth. TGF-beta binds first to a Type II (TGFBR2), then a Type I receptor (TGFBR1). TGFBR1 activation results in the phosphorylation of intracellular messengers, the SMADs. Unrestricted cell growth due to decreased growth inhibitory activity is a paramount feature of a defect in TGF-beta function. There is growing evidence that common variants of the TGF-beta pathway ligand and receptors that alter TGF-beta signaling modify cancer risk. Approximately 14% of the general population carry TGFBR1*6A, a variant of the TGFBR1 gene that results in decreased TGF-beta-mediated growth inhibition. Recent studies show that overall cancer risk is increased by 70 and 19% among TGFBR1*6A homozygotes and heterozygotes, respectively. This suggests that TGFBR1*6A may contribute to the development of a large proportion of common forms of cancer and may become a target for cancer chemoprevention. While decreased TGF-beta signaling increases cancer risk, TGF-beta secretion and activated TGF-beta signaling enhances the aggressiveness of several types of tumors. The activated TGF-beta signaling pathway is emerging as an attractive target in cancer and the authors predict that assessment of functionally relevant variants of this pathway will lead to the identification of individuals with a higher cancer risk and account for some forms of familial cancer susceptibility. In addition, it is predicted that inhibitors of the TGF-beta signaling pathway will find their way into cancer clinical trials, leading to delays in tumor progression and improvements in overall survival. PMID- 15270669 TI - Dose-escalated 3D conformal radiotherapy in prostate cancer. AB - 3D conformal radiotherapy involves the delivery of radiation to a defined 3D tumor volume while minimizing doses to adjacent critical tissues. The use of sophisticated imaging tools and advanced treatment planning software have allowed for better target definition enabling the oncologist to conform or shape radiation volume more closely around the target while minimizing dose to the rectum and bladder. 3D conformal radiotherapy has resulted in dramatic reductions in acute and late toxicity of radiation treatment in prostate cancer. It has also allowed for safe escalation of radiation dose with improved tumor control compared with conventional dose radiotherapy. Long-term tumor control rates with 3D conformal radiotherapy are comparable with results using radical prostatectomy. PMID- 15270670 TI - Advances in the management of testicular cancer. AB - Although testicular cancer is currently a rare disease, the incidence is rising. The most important risk factor remains cryptorchism and there is a variable association with testicular microlithiasis. Serum tumor markers remain important for diagnosis, and they have prognostic value and can be used to monitor therapy and follow-up. Conventional imaging can only be improved in specific categories of patients with positron emission tomography scanning. The optimal therapy after orchiectomy should be individualized based on the histology of the primary specimen, the presence or absence of metastasis, and marker levels. An optimal definition of risk factors will likely spur the development of risk-adjusted treatment modalities and subsequently lead to better results and less toxicity. Extensive research into the molecular biology of testicular cancer is ongoing and will hopefully offer new targets for the diagnosis, staging, and treatment of testicular cancer in the future. PMID- 15270671 TI - Current treatment options for endometrial cancer. AB - In North America, endometrial cancer is the most prevalent cancer of the female genital tract. On the basis of clinical and histologic variables, two main types of endometrial cancer have been described: Type I tumors, which are usually well differentiated and endometrioid in histology and account for the majority of cases; and Type II, which are poorly differentiated tumors, often with serous papillary or clear cell histology. Due to the early declaration of the disease by vaginal bleeding, approximately 80% of endometrial cancers are diagnosed at an early stage. Total abdominal hysterectomy and bilateral salpingo-oophorectomy with or without lymph node dissection remains the cornerstone of treatment. Tumor stage, histologic grade and depth of myometrial invasion are the most important prognostic factors. If myometrial invasion to 50% or more of the myometrial width and/or grade 2 or 3 histology is present, pelvic radiotherapy is indicated to reduce the risk of pelvic recurrence. Postoperative radiation therapy may improve local control but does not affect survival for Stage I endometrial cancer patients. Systemic chemotherapy is typically reserved for women with disseminated primary disease or extrapelvic recurrence. Although the combination of cisplatin plus doxorubicin is commonly used, carboplatin plus paclitaxel represents an efficacious, low-toxicity regimen for managing advanced or recurrent endometrial cancer. Recently, a significant percentage of Type II uterine tumors have been found to overexpress the epidermal growth factor Type II receptor. Anti-HER-2/neu targeted therapy might be a novel and attractive therapeutic strategy in patients harboring this biologically aggressive variant of endometrial cancer. PMID- 15270672 TI - Targeting mTOR-mediated survival signals in anticancer therapeutic strategies. AB - An important component of tumor progression is the generation of survival signals that overcome default apoptotic programs. In principle, survival signals are ideal targets for anticancer therapeutic strategies because blocking these signals leads to the death of cells that are dependent upon them. A common target of survival signals is mTOR. Survival signals generated by both phosphatidylinositol-3-kinase and phospholipase D target mTOR. Suppression of these mTOR-mediated survival signals provides the opportunity to reactivate default apoptotic pathways in cancer cells and allow the cancer cells to die on their own. In this review, the potential for anticancer strategies that target mTOR-mediated survival signals is explored. PMID- 15270673 TI - A sensitivity analysis of the fed-batch animal-cell bioreactor with respect to some control parameters. AB - Animal cell culture is widely used in the manufacture of valuable products, and this process is nowadays seeing a rapid expansion. The growth of animal cells is a complex process, because the cells are very sensitive to environmental changes (in, for example, nutrients, pH, temperature, oxygen and osmolarity) during this phase and to the toxic compounds produced by the cell itself. Ammonia and lactate are the two major waste materials of cell culture. They can have inhibitory effects on cell growth and product (monoclonal antibodies among others) formation. In order to model the behaviour of a fed-batch animal cell bioreactor producing monoclonal antibodies, it is necessary to use a complex kinetic model with optimal operating patterns ensuring high productivities. Good knowledge of such domains of operating parameters, together with the understanding of the response of this rather complex system to small modifications in the working conditions, are essential for on-line control to improve the quality of product and the yield of an animal cell culture. The present study focuses on the sensitivity analysis of a fed-batch animal cell bioreactor with respect to some candidate control parameters (substrate set-point concentrations, feeding time step patterns and concentration of feeding solutions), emphasizing the influence of these on the overall performance of the system. PMID- 15270674 TI - Involvement of the nucleolus in plant virus systemic infection. AB - The nucleolus is a prominent subnuclear domain and is classically regarded as the site of transcription of rRNA, processing of the precursor rRNAs and biogenesis of pre-ribosomal particles. In addition to these traditionally recognized activities, the nucleolus also participates in many other aspects of cell function. The umbravirus-encoded ORF3 protein is a multifunctional RNA-binding protein involved in long-distance RNA movement, and protection of viral RNA from RNase attack, including possibly small interfering RNA-guided RNA silencing. In addition to its presence in cytoplasmic ribonucleoprotein particles containing viral RNA, the umbraviral ORF3 protein accumulates in nuclei, preferentially targeting nucleoli. The ORF3 protein domains involved in the localization of the protein to the nucleolus were identified. Functional analysis of the mutants revealed the correlation between the ORF3 protein nucleolar localization and its ability to form the cytoplasmic ribonucleoprotein particles and transport viral RNA long distances via the phloem. Possible mechanisms of the nucleolar involvement in systemic virus infection are discussed. PMID- 15270675 TI - A plethora of plant serine/arginine-rich proteins: redundancy or evolution of novel gene functions? AB - Precursor-mRNA (pre-mRNA) processing is an important step in gene expression and its regulation leads to the expansion of the gene product repertoire. SR (serine arginine)-rich proteins are key players in intron recognition and spliceosome assembly and significantly contribute to the alternative splicing process. Due to several duplication events, at least 19 SR proteins are present in the Arabidopsis genome, which is almost twice as many as in humans. They fall into seven different subfamilies, three of them homologous with metazoan splicing factors, whereas the other four seem to be specific for plants. The current results show that most of the duplicated genes have different spatiotemporal expression patterns indicating functional diversification. Interestingly, most of the SR protein genes are alternatively spliced and in some cases this process was shown to be under developmental and/or environmental control. This might greatly influence gene expression of target genes as also exemplified by ectopic expression studies of particular SR proteins. PMID- 15270676 TI - RNA processing and Arabidopsis flowering time control. AB - Plants control their flowering time in order to ensure that they reproduce under favourable conditions. The components involved in this complex process have been identified using a molecular genetic approach in Arabidopsis and classified into genetically separable pathways. The autonomous pathway controls the level of mRNA encoding a floral repressor, FLC, and comprises three RNA-binding proteins, FCA, FPA and FLK. FCA interacts with the 3'-end RNA-processing factor FY to autoregulate its own expression post-transcriptionally and to control FLC. Other components of the autonomous pathway, FVE and FLD, regulate FLC epigenetically. This combination of epigenetic and post-transcriptional control gives precision to the control of FLC expression and flowering time. PMID- 15270677 TI - Post-transcriptional steps involved in the assembly of photosystem I in Chlamydomonas. AB - Assembly of the PSI (photosystem I) complex in eukaryotic photosynthetic organisms depends on the concerted interactions of the nuclear and chloroplast genetic systems. We have identified several nucleus-encoded factors of Chlamydomonas reinhardtii that are specifically required for the synthesis of the two large chloroplast-encoded reaction-centre polypeptides, PsaA and PsaB, of photosystem I and that function at plastid post-transcriptional steps. Raa1, Raa2 and Raa3 are required for the splicing of the three discontinuous psaA precursor transcripts; they are part of large RNA-protein complexes that are reminiscent of spliceosomal particles. Tab1 and Tab2 are involved in the initiation of translation of the psaB mRNA and are localized in the membrane and stromal phases of the chloroplast, where they are associated with high-molecular-mass complexes. Moreover, two chloroplast-encoded proteins, Ycf3 and Ycf4, are required for the primary steps of assembling the photosystem I subunits into a functional complex. PMID- 15270678 TI - Chloroplast RNA-binding and pentatricopeptide repeat proteins. AB - Chloroplast gene expression is mainly regulated at the post-transcriptional level by numerous nuclear-encoded RNA-binding protein factors. In the present study, we focus on two RNA-binding proteins: cpRNP (chloroplast ribonucleoprotein) and PPR (pentatricopeptide repeat) protein. These are suggested to be major contributors to chloroplast RNA metabolism. Tobacco cpRNPs are composed of five different proteins containing two RNA-recognition motifs and an acidic N-terminal domain. The cpRNPs are abundant proteins and form heterogeneous complexes with most ribosome-free mRNAs and the precursors of tRNAs in the stroma. The complexes could function as platforms for various RNA-processing events in chloroplasts. It has been demonstrated that cpRNPs contribute to RNA stabilization, 3'-end formation and editing. The PPR proteins occur as a superfamily only in the higher plant species. They are predicted to be involved in RNA/DNA metabolism in chloroplasts or mitochondria. Nuclear-encoded HCF152 is a chloroplast-localized protein that usually has 12 PPR motifs. The null mutant of Arabidopsis, hcf152, is impaired in the 5'-end processing and splicing of petB transcripts. HCF152 binds the petB exon-intron junctions with high affinity. The number of PPR motifs controls its affinity and specificity for RNA. It has been suggested that each of the highly variable PPR proteins is a gene-specific regulator of plant organellar RNA metabolism. PMID- 15270679 TI - Genetics of the DST-mediated mRNA decay pathway using a transgene-based selection. AB - mRNA sequences that control abundance, localization and translation initiation have been identified, yet the factors that recognize these sequences are largely unknown. In this report, a transgene-based strategy designed to isolate mutants of Arabidopsis thaliana that fail to recognize these sequences is described. In this strategy, a selectable gene and a screenable marker gene are put under the control of the sequence element being analysed and mutants are selected with altered abundance of the corresponding marker RNAs. The selection of mutants deficient in recognition of the DST (downstream) mRNA degradation signal is used as a test-case to illustrate some of the technical aspects that have facilitated success. Using this strategy, we report the isolation of a new mutant, dst3, deficient in the DST-mediated mRNA decay pathway. The targeted genetic strategy described circumvents certain technical limitations of biochemical approaches. Hence, it provides a means to investigate a variety of other mechanisms responsible for post-transcriptional regulation. PMID- 15270680 TI - Characterization of a crucifer plant pre-rRNA processing complex. AB - In cruciferous plants, the primary pre-rRNA cleavage site (P site) is immediately downstream of four similar, highly conserved sequences (A(1), A(2), A(3) and B) located within the 5'-ETS (5'-external transcribed spacer). In the present study, we describe the characterization of a plant NF D (nuclear factor D) that binds and interacts specifically with this A(123)BP cluster in the rDNA sequence. NF D is a high-molecular-mass complex containing nucleolin, fibrillarin and U3 and U14 snoRNAs. Furthermore, we show that NF D binds and cleaves pre-rRNA specifically at the P site. Thus we conclude that NF D is a pre-rRNA processing complex that may first assemble on rDNA and then bind nascent pre-rRNA. PMID- 15270681 TI - Plant growth: the translational connection. AB - The TOR (target of rapamycin) pathway is a phylogenetically conserved transduction system in eukaryotes linking the energy status of the cell to the protein synthesis apparatus and to cell growth. The TOR protein is specifically inhibited by a rapamycin-FKBP12 complex (where FKBP stands for FK506-binding protein) in yeast and animal cells. Whereas plants appear insensitive to rapamycin, Arabidopsis thaliana harbours a single TOR gene, which is essential for embryonic development. It was found that the product of this gene was capable of binding to rapamycin and yeast FKBP12. In-frame fusion with a GUS reporter gene shows that the TOR protein is produced essentially in proliferating zones, whereas the TOR mRNA can be detected in all organs suggesting a translational regulation of TOR. Phenotypic analysis of Arabidopsis TOR mutants indicates that the plant TOR pathway fulfils the same role in controlling cell growth as its other eukaryotic counterparts. PMID- 15270682 TI - The role of the initiation surveillance complex in promoting efficient protein synthesis. AB - Initiation is most often the rate-limiting step of translation. Translation initiation requires the involvement of numerous factors that assist binding of the 40 S ribosomal subunit to an mRNA and the assembly of the 80 S ribosome at the correct initiation codon. Recruitment of an initiation surveillance complex is required for translation and serves to identify mRNAs that are structurally and functionally competent for translation. For most cellular mRNAs, recruitment of the surveillance complex requires the 5'-cap and 3'-poly(A) tail. However, some cellular and viral mRNAs that naturally lack either of these have evolved alternatives that serve to recruit the complex. The initiation surveillance complex functions to stabilize eIF4F (where eIF stands for eukaryotic initiation factor), the cap-binding complex, to the cap; promote eIF4A helicase activity to remove secondary structure in the 5'-leader that might otherwise reduce 40 S ribosomal subunit scanning; promote eIF4B binding to increase eIF4A/eIF4F function and stabilize binding of the poly(A)-binding protein to the poly(A) tail. The surveillance complex is regulated through changes in phosphorylation in response to environmental conditions or by developmental signals as a means to regulate globally protein synthesis. Thus the initiation surveillance complex ensures that only intact mRNAs are recruited for translation and serves to regulate protein synthesis. PMID- 15270683 TI - Plant translation initiation factors: it is not easy to be green. AB - Plants have significant differences in some of the 'parts' of the translational machinery. There are two forms of eukaryotic initiation factor (eIF) 4F, eIF3 has two novel subunits, eIF4B is poorly conserved, and eIF2 kinases and eIF4E binding proteins (4E-BP) are yet to be discovered. These differences suggest that plants may regulate their translation in unique ways. PMID- 15270684 TI - Control of translation reinitiation on the cauliflower mosaic virus (CaMV) polycistronic RNA. AB - Translation of the polycistronic 35S RNA of CaMV (cauliflower mosaic virus) occurs via a reinitiation mechanism, which requires TAV (transactivator/viroplasmin). To allow translation reinitiation of the major open reading frames on the polycistronic RNA, TAV interacts with the host translational machinery via eIF3 (eukaryotic initiation factor 3) and the 60S ribosome. Accumulation of TAV and eIF3 in the polysomal fraction isolated from CaMV-infected cells suggested that TAV prevents loss of eIF3 from the translating ribosomes during the first initiation event. The TAV-eIF3-80S complex could be detected in vitro by sucrose-gradient-sedimentation analysis. The question is whether TAV interacts directly with the 48S preinitiation complex or enters polysomes after the first initiation event. eIF4B, a component of the 48S initiation complex, can preclude formation of the TAV-eIF3 complex via competition with TAV for eIF3 binding; the eIF4B- and TAV-binding sites on eIF3g overlap. eIF4B out-competes TAV for binding to eIF3 and to the eIF3-40S complex. Transient overexpression of eIF4B in plant protoplasts specifically inhibits TAV mediated transactivation of polycistronic translation. Our results thus indicate that eIF4B precludes TAV-eIF3-40S complex formation during the first initiation event. Consequently, overexpression of TAV in plant protoplasts affects only the second and subsequent initiation events. We propose a model in which TAV enters the host translational machinery at the eIF4B-removal step to stabilize eIF3 within polysomes. PMID- 15270685 TI - Autoregulation of the gene for cystathionine gamma-synthase in Arabidopsis: post transcriptional regulation induced by S-adenosylmethionine. AB - Cystathionine gamma-synthase (CGS) catalyses the first committed step of methionine biosynthesis in higher plants. CGS is encoded by the CGS1 gene in Arabidopsis. Stability of CGS1 mRNA is down-regulated in response to methionine application and the exon 1-coding region of CGS1 itself is necessary and sufficient for this regulation. mto1 (for methionine overaccumulation) mutants of Arabidopsis, which carry single-amino-acid sequence alterations within CGS1 exon 1, are deficient in this regulation and overaccumulate methionine. Since CGS1 exon 1 acts in cis during this regulation, we have proposed a model that the regulation occurs during translation of CGS1 mRNA when the nascent polypeptide of CGS and its mRNA are in close proximity. In fact, application of the translation inhibitor cycloheximide abolished this regulation in vivo. This model predicts that the regulation can be reproduced in an in vitro translation system. Studies using the in vitro translation system of wheatgerm extract have indicated that S adenosylmethionine, a direct metabolite of methionine, is the effector of this regulation. A 5'-truncated RNA species, which is a probable degradation intermediate of CGS1 mRNA in vivo, was also detected in vitro, suggesting that the wheatgerm in vitro translation system reflects the in vivo regulation. PMID- 15270686 TI - Regulation of chloroplast translation: interactions of RNA elements, RNA-binding proteins and the plastid ribosome. AB - Chloroplast gene expression is primarily controlled during the translation of plastid mRNAs into proteins, and genetic studies have identified cis-acting RNA elements and trans-acting protein factors required for chloroplast translation. Biochemical analysis has identified both general and specific mRNA-binding proteins as components of the regulation of chloroplast translation, and has revealed that chloroplast translation is related to bacterial translation but is more complex. Utilizing proteomic and bioinformatic analyses, we have identified the proteins that function in chloroplast translation, including a complete set of chloroplast ribosomal proteins, and homologues of the 70 S initiation, elongation and termination factors. These analyses show that the translational apparatus of chloroplasts is related to that of bacteria, but has adopted a number of eukaryotic mechanisms to facilitate and regulate chloroplast translation. PMID- 15270687 TI - Translation inhibition during the induction of apoptosis: RNA or protein degradation? AB - The induction of apoptosis leads to a substantial inhibition of protein synthesis. During this process changes to the translation-initiation factors, the ribosome and the cellular level of mRNA have been documented. However, it is by no means clear which of these events are necessary to achieve translational shutdown. In this article, we discuss modifications to the translational apparatus that occur during apoptosis and examine the potential contributions that they make to the inhibition of protein synthesis. Moreover, we present evidence that suggests that a global increase in the rate of mRNA degradation occurs before the caspase-dependent cleavage of initiation factors. Increased mRNA decay is temporally correlated with the shutdown of translation and therefore plays a major role in the inhibition of protein synthesis in apoptotic cells. PMID- 15270688 TI - A key factor of translation reinitiation, ribosomal protein L24, is involved in gynoecium development in Arabidopsis. AB - In polycistronic genes, uORFs (upstream open reading frames) within the 5' transcript leader sequence of major ORFs may regulate the translation of these major ORFs. In this case, ribosome reinitiates translation at a start codon of downstream ORF after translation termination of uORF. The plant RPL24 (ribosomal protein L24) is a key factor for translation reinitiation of downstream ORFs on the polycistronic cauliflower mosaic virus 35S RNA transcription unit. In the RPL24-deficient mutant of Arabidopsis, short valve (stv), the basal region of the ovary is shortened, whereas the gynophore appears elongated. This phenotype is never seen in known mutants of other ribosomal protein genes, suggesting that RPL24 has a specific role in gynoecium development. Similar phenotypes were observed in the ett (ettin) and mp (monopteros) mutants. Both ETT and MP genes possess uORFs. We examined the hypothesis that these uORFs regulate their downstream major ORFs by a transient expression assay in Arabidopsis mesophyll protoplasts. Our results supported the idea, suggesting that the structural defects of the gynoecium in stv mutants were caused by decreased efficiency of translation reinitiation of ETT and/or MP. PMID- 15270689 TI - Choice of a start codon in a single transcript determines DNA ligase 1 isoform production and intracellular targeting in Arabidopsis thaliana. AB - DNA ligase 1 (AtLIG1) is the only essential DNA ligase activity in Arabidopsis and is implicated in the important processes of DNA replication, repair and recombination and in transgene insertion during Agrobacterium-mediated plant transformations. The mitochondrial and nuclear forms of DNA ligase 1 in Arabidopsis are translated from a single mRNA species through the control of translation initiation from either the first (M1) or second (M2) in-frame AUG codons respectively. Translation from a third in-frame AUG codon (M3) occurs on transcripts in which M1 and M2 are mutagenized to stop codons. Wild-type AtLIG1 GFP constructs (where GFP stands for green fluorescent protein) can be targeted in planta to both the nucleus and mitochondria. AtLIG1-GFP translation from M1 specifically targets the fusion protein only to mitochondria in planta, whereas translation from M2 or M3 targets the fusion protein only to the nucleus. Interestingly, the AtLIG1-GFP fusion protein in which translation is initiated from M1 contains both an N-terminal mtPS (mitochondrial targeting presequence) and a nuclear localization signal; nonetheless, this protein is only targeted to the mitochondria. This result raises intriguing questions on the translational control mechanisms that regulate how the protein products of a single transcript are targeted to more than one cellular compartment. PMID- 15270690 TI - Mechanism of substrate recognition by Hsp70 chaperones. AB - The role of Hsp70 (heat-shock protein 70) chaperones in assisting protein-folding processes relies on their ability to associate with short peptide stretches of protein substrates in a transient and ATP-controlled manner. In the present study, we review the molecular details of the mechanism behind substrate recognition by Hsp70 proteins. PMID- 15270691 TI - Peptides complexed with the protein HSP70 generate efficient human cytolytic T lymphocyte responses. AB - Microbial HSPs (heat-shock proteins) are implicated in the induction of the innate and adaptive arms of the immune response. We set out to determine whether peptides complexed with HSP70 generate efficient CTL (cytolytic T-lymphocyte) responses. Human dendritic cells pulsed with peptide-loaded microbial HSP70 complexes generate potent antigen-specific CTL responses. Using fluorescence anisotropy, we have calculated the peptide-binding affinity of mycobacterial HSP70 (K(D)=14 microM) and show that 120 pM HSP70-bound peptide is sufficient to generate a peptide-specific CTL response that is four orders of magnitude more efficient than the peptide alone. Through the generation of mycobacterial HSP70 truncations, we find that the minimal 136 amino acid, mycobacterial HSP70 peptide binding domain is sufficient to generate CTL responses. The design of an HSP70 mutant, in which the peptide-binding site of HSP70 is filled with a bulky hydrophobic residue, leads to a large decrease in the peptide-binding affinity. This mutant HSP70 retains stimulatory capacity but is unable to generate CTL and has separated antigen delivery from immunostimulation of dendritic cells. PMID- 15270692 TI - BCG (Bacille Calmette-Guerin) HspCs (heat-shock protein-peptide complexes) induce T-helper 1 responses and protect against live challenge in a murine aerosol challenge model of pulmonary tuberculosis. AB - The need for an effective TB (tuberculosis) vaccine remains acute, with tuberculosis still one of the major killers worldwide and 3 million new infections annually. We report here on the immune responses elicited by HspCs (heat-shock protein-peptide complexes) isolated from BCG (Bacille Calmette Guerin) vaccine. These HspCs elicit both the appropriate cellular and protective immune responses required to merit their further development as TB vaccine candidates. PMID- 15270693 TI - Functional domains of HSP70 stimulate generation of cytokines and chemokines, maturation of dendritic cells and adjuvanticity. AB - Microbial HSP70 (heat-shock protein 70) consists of three functionally distinct domains: an N-terminal 44 kDa ATPase portion (amino acids 1-358), followed by an 18 kDa peptide-binding domain (amino acids 359-494) and a C-terminal 10 kDa fragment (amino acids 495-609). Immunological functions of these three different domains in stimulating monocytes and dendritic cells have not been fully defined. However, the C-terminal portion (amino acids 359-610) stimulates the production of CC chemokines, IL-12 (interleukin-12), TNFalpha(tumour necrosis factor alpha), NO and maturation of dendritic cells and also functions as an adjuvant in the induction of immune responses. In contrast, the ATPase domain of microbial HSP70 mostly lacks these functions. Since the receptor for HSP70 is CD40, which with its CD40 ligand constitutes a major co-stimulatory pathway in the interaction between antigen-presenting cells and T-cells, HSP70 may function as an alternative ligand to CD40L. HSP70-CD40 interaction has been demonstrated in non human primates to play a role in HIV infection, in protection against Mycobacterium tuberculosis and in conversion of tolerance to immunity. PMID- 15270694 TI - Scavenger receptors and heat-shock protein-mediated antigen cross-presentation. AB - Heat-shock proteins (HSPs) induce protective cytotoxic immune responses against tumour antigens. This property is related to their ability to bind to and to be internalized by DC (dendritic cells) before gaining access to the MHC class I processing pathway, a process called antigen cross-presentation. This process requires internalization of the antigen by DC via endocytic receptors. Owing to their particular immune properties, several studies were focused on the identification of HSP-binding elements on DC. We and others have reported that scavenger receptors are the main HSP-binding structures on human DC and have identified LOX-1 as one of these molecules. The binding of human Hsp70 to DC and the in vitro Hsp70-mediated antigen cross-presentation are inhibited by an anti LOX-1 monoclonal antibody. In vivo, targeting LOX-1 with a tumour antigen using an anti-LOX-1 monoclonal antibody induces antitumour immunity. Thus scavenger receptors are certainly new promising targets for cancer immunotherapy. PMID- 15270695 TI - Heat-shock protein 70 and heat-shock protein 90 associate with Toll-like receptor 4 in response to bacterial lipopolysaccharide. AB - Mammalian responses to bacterial LPS (lipopolysaccharide) from the outer membrane of Gram-negative bacteria can lead to an uncontrolled inflammatory response that can be deadly for the host. It has been shown that the innate immune system employs at least three cell surface receptors, CD14, TLR4 (Toll-like receptor 4) and MD-2, in order to recognize bacterial LPS. In our previous work we have found that Hsps (heat-shock proteins) are also involved in the innate recognition of bacterial products. Their presence on the cell surface, as well as their involvement in the innate recognition process, are poorly understood. In the present study we have investigated the association of TLR4 with Hsp70 and Hsp90 following LPS stimulation, both on the cell surface and intracellularly. Our results show that Hsp70 and Hsp90 form a cluster with TLR4 within lipid microdomains following LPS stimulation. In addition, Hsp70 and Hsp90 seem to be involved in TLR4/LPS trafficking and targeting to the Golgi apparatus, since upon LPS stimulation we found that both Hsps are targeted to the Golgi along with TLR4. The present study sheds new light into the involvement of Hsps in the innate immune response. PMID- 15270696 TI - Neuronal DnaJ proteins HSJ1a and HSJ1b: a role in linking the Hsp70 chaperone machine to the ubiquitin-proteasome system? AB - The heat-shock protein 70 chaperone machine is functionally connected to the ubiquitin-proteasome system by the co-chaperone CHIP. In this article, we discuss evidence that the neuronal DnaJ proteins HSJ1a and HSJ1b may represent a further link between the cellular protein folding and degradation machineries. We have demonstrated that HSJ1 proteins contain putative ubiquitin interaction motifs and can modulate the cellular processing of rhodopsin, a protein that is targeted for degradation by the proteasome when it is misfolded. PMID- 15270697 TI - Role of AIP and its homologue the blindness-associated protein AIPL1 in regulating client protein nuclear translocation. AB - Mutations in the AIPL1 (aryl hydrocarbon receptor interacting protein-like 1) cause the blinding disease Leber's congenital amaurosis. AIPL1 is a homologue of the AIP. AIP functions as part of a chaperone heterocomplex to facilitate signalling by the AhR and plays an important role in regulating the nuclear translocation of the receptor. We review the evidence for the role of AIP in protein translocation and compare the potential functions of AIPL1 in the translocation of its interacting partner the NEDD8 ultimate buster protein 1. PMID- 15270698 TI - Identification and biophysical characterization of a very-long-chain-fatty-acid substituted phosphatidylinositol in yeast subcellular membranes. AB - Morphological analysis of a conditional yeast mutant in acetyl-CoA carboxylase acc1ts/mtr7, the rate-limiting enzyme of fatty acid synthesis, suggested that the synthesis of C26 VLCFAs (very-long-chain fatty acids) is important for maintaining the structure and function of the nuclear membrane. To characterize this C26-dependent pathway in more detail, we have now examined cells that are blocked in pathways that require C26. In yeast, ceramide synthesis and remodelling of GPI (glycosylphosphatidylinositol)-anchors are two pathways that incorporate C26 into lipids. Conditional mutants blocked in either ceramide synthesis or the synthesis of GPI anchors do not display the characteristic alterations of the nuclear envelope observed in acc1ts, indicating that the synthesis of another C26-containing lipid may be affected in acc1ts mutant cells. Lipid analysis of isolated nuclear membranes revealed the presence of a novel C26 substituted PI (phosphatidylinositol). This C26-PI accounts for approx. 1% of all the PI species, and is present in both the nuclear and the plasma membrane. Remarkably, this C26-PI is the only C26-containing glycerophospholipid that is detectable in wild-type yeast, and the C26-substitution is highly specific for the sn-1 position of the glycerol backbone. To characterize the biophysical properties of this lipid, it was chemically synthesized. In contrast to PIs with normal long-chain fatty acids (C16 or C18), the C26-PI greatly reduced the bilayer to hexagonal phase transition of liposomes composed of 1,2-dielaidoyl-sn glycero-3-phosphoethanolamine (DEPE). The biophysical properties of this lipid are thus consistent with a possible role in stabilizing highly curved membrane domains. PMID- 15270699 TI - Protein kinase function and glutathionylation. AB - Intracellular reactive oxygen species are generated as a by-product of normal metabolic processes and can both damage cellular constituents and function as important signalling species. This signalling often involves changes in the thiol redox balance. As an antioxidant, glutathione serves in maintaining the reduced state of cellular protein thiol groups. The paper by Cross and Templeton appearing in this issue of the Biochemical Journal describes a mechanism by which glutathionylation plays a key role in the regulation of the kinase activity of MEKK1 [MAP (mitogen-activated protein kinase)/ERK (extracellular-signal-regulated kinase) kinase kinase; MAP3K] in response to oxidative stresses. This type of post-translational-modification glutathionylation may represent a general mechanism by which protein kinase function can be regulated. PMID- 15270700 TI - That zincing feeling: the effects of EDTA on the behaviour of zinc-binding transcriptional regulators. AB - Zinc-binding proteins account for nearly half of the transcription regulatory proteins in the human genome and are the most abundant class of proteins in the human proteome. The zinc-binding transcriptional regulatory proteins utilize Zn2+ to fold structural domains that participate in intermolecular interactions. A study by Matt et al. in this issue of the Biochemical Journal has examined the transcription factor binding properties of the zinc-binding module C/H1 (cysteine/histidine-rich region 1) found in the transcriptional co-activator proteins CBP (CREB-binding protein) and p300. Their studies revealed that EDTA treatment of native C/H1 leads to irreversible denaturation and aggregation. Of particular concern is their finding that unfolded C/H1 participates in non specific protein-protein interactions. The implications of these results are significant. EDTA is a very potent zinc-chelating agent that is used ubiquitously in protein interaction studies and in molecular biology in general. The potentially detrimental effects of EDTA on the structure and interactions of zinc binding proteins should be taken into account in the interpretation of a sizeable number of published studies and must be considered in future experiments. PMID- 15270702 TI - The biology and engineering of stem-cell control. AB - There is significant interest in studying stem cells, both to elucidate their basic biological functions during development and adulthood as well as to learn how to utilize them as new sources of specialized cells for tissue repair. Whether the motivation is basic biology or biomedical application, however, progress will hinge upon learning how to better control stem-cell function at a quantitative and molecular level. There are several major challenges within the field, including the identification of new signals and conditions that regulate and influence cell function, and the application of this information towards the design of stem-cell bioprocesses and therapies. Both of these efforts can significantly benefit from the synthesis of biological data into quantitative and increasingly mechanistic models that not only describe, but also predict, how a stem cell's environment can control its fate. This review will briefly summarize the history and current state of the stem-cell biology field, but will then focus on the development of predictive models for stem-cell control. Early models formulated on the assumption that cell fate was decided by stochastic, cell intrinsic processes have gradually evolved into hybrid deterministic-stochastic models with increasingly finer molecular resolution that accounts for environmental regulation. As our understanding of cellular control mechanisms expands from the cell surface and towards the nucleus, these efforts may culminate in the development of a stem-cell culture programme, or a series of signals to provide to the cells as a function of time to guide them along a desired developmental trajectory. PMID- 15270703 TI - Stem-cell plasticity and therapy for injuries of the peripheral nervous system. AB - Numerous publications have investigated stem-cell biology and the possible therapeutic use of stem cells in a wide range of injuries and diseases. This interest has been fueled by recent reports suggesting that mesenchymal stem cells can show unorthodox plasticity, their being able to transdifferentiate into cells of different lineages, such as neuronal phenotypes. This capability has obvious implications for their potential application in tissue engineering and tissue regeneration. The peripheral nervous system has an inherent capacity for regeneration, but this is limited and not matched by the level of reinnervation of target organs, with a resulting loss of functional recovery. Several approaches have been attempted in order to overcome this deficiency, and transplant of cultured Schwann cells into bioengineered conduits has been shown to improve regeneration. An alternative may be the use of stem-cell technology, whereby cultured and differentiated stem cells can be transplanted to the site of injury in order to promote enhanced regeneration. The present review discusses the use of stem cells applied to the repair of peripheral nerve injury and their role in the regeneration process. PMID- 15270704 TI - Human adult craniofacial muscle-derived cells: neural-cell adhesion-molecule (NCAM; CD56)-expressing cells appear to contain multipotential stem cells. AB - Skeletal muscle has been well characterized as a reservoir of myogenic precursors or satellite cells with the potential to participate in cellular repopulation therapies for muscle dysfunction. Recent evidence, however, suggests that the postnatal muscle compartment can be considered an alternative to bone marrow as a source of multipotent cells or muscle-derived stem cells (MDSCs). MDSCs, when primed with appropriate environmental cues, can differentiate into a variety of non-muscle cells. The present study describes the application of a new technique for the isolation of adult human myoblasts and putative MDSCs, based on microbead immunomagnetic selection of CD56+ cells, derived from craniofacial skeletal muscle, and details changes in morphological/molecular phenotype of the purified cells when maintained in either a myogenic or a non-myogenic milieu. Multiple immunofluorescence microscopy and two-colour flow-cytometric analysis of proliferating CD56+ cultures revealed positive staining for myogenic markers (CD56, desmin and M-cadherin) as well as putative stem-cell markers [the antigens CD34, CD90 and CD106, and Flk-1 (fetal liver kinase-1)/VEGFR-2 (vascular endothelial-growth-factor receptor)]. Confluent cultures subjected to cycles of adipogenic or osteogenic induction contained either adipocytes or osteoblasts and myotubes. In conclusion, the CD56+ subpopulation within adult human skeletal muscle is heterogeneous and is composed of both lineage-committed myogenic cells and multipotent cells (the candidate MDSCs), which are able to form non-muscle tissue such as fat and bone. PMID- 15270705 TI - Regulatory and microbiological safety issues surrounding cell and tissue engineering products. AB - Cell therapies and tissue-engineered products that contain living cells are potentially some of the most exciting of the novel therapeutic products currently under development. These products, however, present a number of important safety issues, particularly with respect to the transmission of human viruses. In addition, the short shelf life of these products precludes the normally extensive characterization performed on other biotherapeutic products. Careful examination of the risks and extensive testing of the raw materials have been used in place of product testing to ensure safety. PMID- 15270706 TI - The effect of hyperosmotic pressure on antibody production and gene expression in the GS-NS0 cell line. AB - It has been widely reported that metabolism, cell growth, cell density, product secretion and specific antibody productivity in mammalian cells are strongly affected by osmotic conditions. Previous studies have shown that hyperosmotic pressure suppresses cell growth while enhancing the productivity of individual cells, but the effect of these two changes does not result in an increase in final product concentration in the culture. An improved understanding of the basic cellular processes of a GS-NS0 mammalian cell culture system would assist in the design of a more efficient mammalian cell culture system and in further optimization of production processes. In this study, various properties of mammalian culture systems, such as productivity, cell viability, metabolism, ion balance and the genes regulated during the culture of the GS-NS0 system under osmotic pressure of iso- (290 mOsm/kg) and hyper- (450 mOsm/kg) osmolarity have been investigated, and we demonstrate that there is a decrease in the growth rate and an increase in specific production rate of hyperosmotic cultures as compared with iso-osmotic cultures. Furthermore, differences between iso- and hyper osmotic cultures have been identified in calcium accumulation and metabolism of NH4+, glucose and lactate. Analysis of gene expression reveals regulation of over 600 genes that are implicated in processes known to be affected by changes in osmotic pressure, such as ion transport, accumulation of osmolytes, cell cycle distribution, proliferation, cytoskeletal organization and cell metabolism. PMID- 15270707 TI - Evaluation of selected control strategies for fed-batch cultures of a hybridoma cell line. AB - While fed-batch suspension culture of animal cells continues to be of industrial importance for the large-scale production of pharmaceutical products, existing control concepts are still insufficient. The present paper illustrates the advantages and disadvantages of different fed-batch strategies, including fixed feed trajectories, control via OUR (oxygen uptake rate) (stoichiometric feeding), a priori determination of feed trajectories based on a kinetic model and the model-based adaptive OLFO (open-loop-feedback-optimal) control strategy. A recommendation as to which control strategy should be used for a specific process has to consider the respective process. For an established process with a well characterized and stable production cell line, probably the application of a fixed feed trajectory should be recommended. An adaptive, model-based control strategy could be the method of choice during cell-line development or for rapid production of small amounts of product for clinical trials, owing to its universal character and because it does not require intensive process development. PMID- 15270708 TI - Bioprosthetic heart valves: the need for a quantum leap. AB - More than 250,000 bioprosthetic heart valves are being implanted annually. Although the majority of recipients are elderly developed-world patients, the most urgent need for tissue valves is in younger patients, where rapid degeneration of contemporary prostheses remains a serious obstacle. After decades of empirical and mostly futile attempts to extend the longevity of tissue valve prostheses, new insights and solutions are on the horizon. Aetiologically, a shift of focus from mineralization to immune responses and inflammation emerges. On the development side, new engineering approaches to both selective extraction of tissue components and cross-links are increasingly defining the new direction. In order to dramatically improve the performance of bioprosthetic heart valves, these new developments need to lead to a broad consensus for a paradigm shift in a hitherto rather stagnant field of medical research. PMID- 15270709 TI - Analytical techniques for the evaluation of liquid protein therapeutics. AB - A common problem in the manufacture of liquid protein therapeutics is the tendency for aggregation and particle formation on extended storage. Analytical techniques are required to study the propensity of solutions to form aggregates and particles and to allow the investigation of the effect of conditions encountered during manufacture and storage. A key challenge is to utilize appropriate specific and sensitive techniques to allow the early detection of initial aggregation events, thereby avoiding the need to resort to extended stability trials. The present review evaluates a range of techniques for the detection of changes in protein conformation and the formation of aggregates and particles. It is hoped that the availability of this information will encourage and facilitate studies to resolve stability issues associated with protein therapeutics. PMID- 15270710 TI - Evaluation of disruption methods for the release of intracellular recombinant protein from Escherichia coli for analytical purposes. AB - The aim of the present study was to find disruption methods that allow fast and reproducible measurement of intracellular recombinant proteins with potential for on-line application. Production of rhSOD (recombinant human superoxide dismutase) by Escherichia coli was used as a model. Three methods of cell disruption, sonication, osmotic shock and chemical treatment using a non-ionic surfactant, were critically compared with respect to efficiency and reproducibility of the release of rhSOD. The release of the recombinant protein was monitored by (i) measurement of the protein content in cell-culture extracts using an SPR (surface plasmon resonance) biosensor, and (ii) assaying the enzyme activity with a colorimetric reagent using a spectrophotometer. Disruption by the non-ionic surfactant showed the best performance in terms of simplicity, reproducibility and efficiency of sample treatment. The surfactant did not interfere with the rhSOD binding to the antibody immobilized on the SPR chip or with the rhSOD activity assay. When comparing the two detection methods during monitoring of an E. coli cultivation, comparable results were obtained. PMID- 15270711 TI - An improved, inexpensive procedure for the large-scale purification of recombinant human erythropoietin. AB - A rapid and simple chromatographic procedure has been developed for the large scale purification of therapeutic-grade rHuEPO (recombinant human erythropoietin) from medium-conditioned cell cultures, which includes ion-exchange, hydrophobic interaction and gel-filtration chromatography. A combination of these well connected steps results in highly purified rHuEPO (> 99%), as revealed by SDS/PAGE and HPLC analyses, with a total yield of 38%. The specific activity of purified rHuEPO was 160,104 i.u./mg. Immunoblotting studies revealed that the protein possesses native EPO immunity. N-terminal sequencing of rHuEPO shows that the first 15 amino acids coincide with those of native EPO reported previously. PMID- 15270712 TI - Behaviour of a recombinant cabbage (Brassica oleracea) phospholipase D immobilized on CNBr-activated and antibody supports. AB - Recombinant cabbage (Brassica oleracea) PLD2 (phospholipase D2) immobilized covalently on CNBr-activated Sepharose expressed low activity (approximately 10%), while that immobilized by binding on to anti-PLD2 IgG-Sepharose was more active (approximately 38%). Coupling of PLD2 to CNBr-activated Sepharose resulted in significant improvement in storage stability without affecting its thermostability, as compared with the soluble enzyme. Binding of PLD2 to the antibody support, however, rendered the enzyme remarkably labile to high temperatures and storage. PMID- 15270713 TI - Production of cytidine 5'-monophospho-N-acetyl-beta-D-neuraminic acid (CMP-sialic acid) using enzymes or whole cells entrapped in calcium pectate-silica-gel beads. AB - The present study focuses on the application of immobilization technology to enzymic sugar syntheses. The paper describes an improved silica-alginate matrix established for entrapment and encapsulation. The replacement of alginate with pectate provided enhanced chemical resistance of the matrix, which allows the use of 1% (w/v) polyphosphate in reaction mixtures. Polylysine, a reagent for silica condensation, was replaced by a much cheaper alternative, namely polyethyleneimine. The proposed design was applied in the production of cytidine 5'-monophospho-N-acetyl-beta-D-neuraminic acid (CMP-sialic acid) by immobilized recombinant enzymes or Escherichia coli cells containing overexpressed enzymes. A comparison between these two strategies was made. On the basis of the results we conceptualized a system to synthesize sialyloligosaccharides by using a biocatalyst entrapped in calcium pectate-silica gel beads. PMID- 15270714 TI - Effect of pH on the production of the antitumor antibiotic retamycin by Streptomyces olindensis. AB - The effect of pH on cell growth and retamycin production in batch bioreactor cultures of Streptomyces olindensis ICB20 was investigated. In fermentations pH controlled over the range 6.0-8.0, the highest retamycin production was achieved at pH 7.0, and the maximum concentration of retamycin, about 1.36 A (absorbance) units, was about 43, 58 and 232% higher than the values obtained at pH 7.5, 6.0 and 8.0 respectively. PMID- 15270715 TI - Nitric oxide and the hyperdynamic circulation in cirrhosis: is there a role for selective intestinal decontamination? AB - Abnormal vascular tone is responsible for many of the complications seen in cirrhosis making the identification of the pathophysiology of abnormal dilatation a major focus in hepatology research. The study of abnormal vascular tone is complicated by the multiple vascular beds involved (hepatic, splanchnic, peripheral, renal and pulmonary), the differences in the underlying cause of portal hypertension (hepatic versus pre-hepatic) and the slow evolution of the hyperdynamic state. The autonomic nervous system, circulating vasodilators and abnormalities in vascular smooth muscle cells (receptors, ion channels, signalling systems and contraction) have all been implicated. There is overwhelming evidence for an overproduction of NO (nitric oxide) contributing to the peripheral dilatation in both animal models of, and in humans with, cirrhosis and portal hypertension. This review focuses on the proposal that endotoxaemia, possibly from gut-derived bacterial translocation, causes induction of NOS (NO synthase) leading to increased vascular NO production, which is the primary stimulus for the development of vasodilatation in cirrhosis and its accompanying clinical manifestations. The current controversy lies not in whether NO production is elevated, but in which isoform of NOS is responsible. We review the evidence for endotoxaemia in cirrhosis and the factors contributing to gut derived bacterial translocation, including intestinal motility and permeability, and finally discuss the possible role of selective intestinal decontamination in the management of circulatory abnormalities in cirrhosis. PMID- 15270716 TI - Cell-penetrating peptides do not cross mitochondrial membranes even when conjugated to a lipophilic cation: evidence against direct passage through phospholipid bilayers. AB - CPPs (cell-penetrating peptides) facilitate the cellular uptake of covalently attached oligonucleotides, proteins and other macromolecules, but the mechanism of their uptake is disputed. Two models are proposed: direct movement through the phospholipid bilayer and endocytic uptake. Mitochondria are a good model system to distinguish between these possibilities, since they have no vesicular transport systems. Furthermore, CPP-mediated delivery of macromolecules to the mitochondrial matrix would be a significant breakthrough in the study of mitochondrial function and dysfunction, and could also lead to new therapies for diseases caused by mitochondrial damage. Therefore we investigated whether two CPPs, penetratin and Tat, could act as mitochondrial delivery vectors. We also determined whether conjugation of the lipophilic cation TPP (triphenylphosphonium) to penetratin or Tat facilitated their uptake into mitochondria, since TPP leads to uptake of attached molecules into mitochondria driven by the membrane potential. Neither penetratin nor Tat, nor their TPP conjugates, are internalized by isolated mitochondria, indicating that these CPPs cannot cross mitochondrial phospholipid bilayers. Tat and TPP-Tat are taken up by cells, but they accumulate in endosomes and do not reach mitochondria. We conclude that CPPs cannot cross mitochondrial phospholipid bilayers, and therefore cannot deliver macromolecules directly to mitochondria. Our findings shed light on the mechanism of uptake of CPPs by cells. The lack of direct movement of CPPs through mitochondrial phospholipid bilayers, along with the observed endosomal accumulation of Tat and TPP-Tat in cells, makes it unlikely that CPPs enter cells by direct membrane passage, and instead favours cellular uptake via an endocytic pathway. PMID- 15270717 TI - Distinct classes of glyoxalase I: metal specificity of the Yersinia pestis, Pseudomonas aeruginosa and Neisseria meningitidis enzymes. AB - The metalloisomerase glyoxalase I (GlxI) catalyses the conversion of methylglyoxal-glutathione hemithioacetal and related derivatives into the corresponding thioesters. In contrast with the previously characterized GlxI enzymes of Homo sapiens, Pseudomonas putida and Saccharomyces cerevisiae, we recently determined that Escherichia coli GlxI surprisingly did not display Zn2+ activation, but instead exhibited maximal activity with Ni2+. To investigate whether non-Zn2+ activation defines a distinct, previously undocumented class of GlxI enzymes, or whether the E. coli GlxI is an exception to the previously established Zn2+-activated GlxI, we have cloned and characterized the bacterial GlxI from Yersinia pestis, Pseudomonas aeruginosa and Neisseria meningitidis. The metal-activation profiles for these additional GlxIs firmly establish the existence of a non-Zn2+-dependent grouping within the general category of GlxI enzymes. This second, established class of metal activation was formerly unidentified for this metalloenzyme. Amino acid sequence comparisons indicate a more extended peptide chain in the Zn2+-dependent forms of GlxI (H. sapiens, P. putida and S. cerevisiae), compared with the GlxI enzymes of E. coli, Y. pestis, P. aeruginosa and N. meningitidis. The longer sequence is due in part to the presence of additional regions situated fairly close to the metal ligands in the Zn2+-dependent forms of the lyase. With respect to sequence alignments, these inserts may potentially contribute to defining the metal specificity of GlxI at a structural level. PMID- 15270718 TI - NMR characterization of the interaction between the C-terminal domain of interferon-gamma and heparin-derived oligosaccharides. AB - Interferons are cytokines that play a complex role in the resistance of mammalian hosts to pathogens. IFNgamma (interferon-gamma) is secreted by activated T-cells and natural killer cells. IFNgamma is involved in a wide range of physiological processes, including antiviral activity, immune response, cell proliferation and apoptosis, as well as the stimulation and repression of a variety of genes. IFNgamma activity is modulated by the binding of its C-terminal domain to HS (heparan sulphate), a glycosaminoglycan found in the extracellular matrix and at the cell surface. In the present study, we analysed the interaction of isolated heparin-derived oligosaccharides with the C-terminal peptide of IFNgamma by NMR, in aqueous solution. We observed marked changes in the chemical shifts of both peptide and oligosaccharide compared with the free state. Our results provide evidence of a binding through electrostatic interactions between the charged side chains of the protein and the sulphate groups of heparin that does not induce specific conformation of the C-terminal part of IFNgamma. Our data also indicate that an oligosaccharide size of at least eight residues displays the most efficient binding. PMID- 15270719 TI - The gene encoding human retinoic acid-receptor-related orphan receptor alpha is a target for hypoxia-inducible factor 1. AB - Retinoic acid-receptor-related orphan receptor (ROR) alpha is a nuclear receptor involved in many pathophysiological processes such as cerebellar ataxia, inflammation, atherosclerosis and angiogenesis. In the present study we first demonstrate that hypoxia increases the amount of Rora transcripts in a wide panel of cell lines derived from diverse tissues. In addition, we identified a functional promoter sequence upstream of the first exon of the human Rora gene, spanning -487 and -45 from the translation initiation site of RORalpha1. When cloned in a luciferase reporter vector, this sequence allowed the efficient transcription of the luciferase gene in several cell lines. Interestingly, the activity of the Rora promoter was enhanced by hypoxia in HepG2 human hepatoma cells, and this effect was dependent on an HRE (hypoxia response element) spanning from -229 to -225. Using electrophoretic-mobility-shift assays, we showed that HIF-1 (hypoxia-inducible factor 1), which plays a key role in the transcriptional response to hypoxia, bound to this HRE. Overexpression of HIF 1alpha increased the activity of the Rora promoter through the HRE. Overexpression of a dominant-negative form of HIF-1alpha producing transcriptionally inactive HIF-1alpha/HIF-1beta dimers abolished hypoxic activation of the Rora promoter. This indicated that HIF-1 is involved in the response of RORalpha to hypoxia. Taken together, our data reveal Rora as a new HIF-1 target gene. This illustrates, at the molecular level, the existence of cross-talk between signalling pathways mediated by HIF-1 and those mediated by nuclear receptors. PMID- 15270720 TI - A chitosanase from Paecilomyces lilacinus with binding affinity for specific chito-oligosaccharides. AB - A purple-spore, rhizosphere-inhabiting nematophagous fungus, further identified as Paecilomyces lilacinus, was found to grow on chitosanase-detecting plate. An induced endochitosanase having a molecular mass of 23 kDa was purified from the culture medium by a single cation-exchange column-chromatography step. Its optimum pH, optimum temperature and pI were found to be 6.0, 50 degrees C and 8.3 respectively. The N-terminal amino acid sequence of the purified enzyme was partially determined. On the basis of the partial sequence XQLPANLXXIYD and the BLAST results, the purified chitosanase was classified as a new member of the family 75 glycohydrolases. Complete hydrolysis of 95% deacetylated chitosan by the isolated chitosanase released chitotriose, chitotetraose and chitopentaose as the major hydrolytic products. Two oligosaccharides, which were further determined to be GlcN-GlcN-GlcNAc and GlcNAc-GlcN-GlcN-GlcNAc by chemical methylation followed by liquid chromatography-tandem MS analysis, were obtained after the denaturation of the purified chitosanase. This is the first documented finding that chitosanase can be produced in a Paecilomyces strain and that it has binding affinity for specific N-acetylated oligosaccharides. PMID- 15270721 TI - Divide and conquer: the importance of cell division in regulating B-cell responses. AB - Proliferation is an essential characteristic of clonal selection and is required for the expansion of antigen reactive clones leading to the development of antibody of different isotypes and memory cells. New data for mouse and human B cells point to an important role for division in regulating isotype class and in optimizing development of protective immunity by the regulated entry of cells to the plasma cell lineage. PMID- 15270722 TI - The potential for Toll-like receptors to collaborate with other innate immune receptors. AB - Cells of the innate immune system express a large repertoire of germ-line encoded cell-surface glycoprotein receptors including Toll-like receptors (TLRs). TLRs recognize conserved motifs on microbes and induce inflammatory signals. Evidence suggests that individual members of the TLR family or other non-TLR surface antigens either physically or functionally interact with each other and cumulative effects of these interactions instruct the nature and outcome of the immune response to a particular pathogen. PMID- 15270723 TI - Increased expression of the natural killer cell inhibitory receptor CD85j/ILT2 on antigen-specific effector CD8 T cells and its impact on CD8 T-cell function. AB - We investigated whether inhibitory natural killer cell receptor (iNKR) expression contributes to impaired antigen-specific cytotoxicity and interferon-gamma (IFN gamma) production by CD8 T cells during chronic infection. iNKR immunoglobulin like transcript-2 (ILT2/CD85j) is expressed on 40-55% of cytomegalovirus (CMV)-, Epstein-Barr virus (EBV)- and human immunodeficiency virus (HIV)-specific CD8 T cells in both healthy and HIV-infected donors. Other iNKRs (CD158a, b1, e1/e2, k, CD94/NKG2A) are expressed on only a small minority of CD8 T cells and are not preferentially expressed on tetramer-staining virus-specific cells. In normal donors, ILT2 is expressed largely on perforin(+) CD27(-) effector cells. However, in HIV-infected donors, only a third of ILT2(+) cells are also perforin(+). In both normal and HIV-infected donors, ILT2(+) cells are prone to spontaneous apoptosis. Therefore, ILT2 is normally expressed during effector cytotoxic T lymphocyte (CTL) differentiation, but can also be expressed when effector maturation is incomplete, as in HIV infection. The effect of ILT2 on CD8 cell function was assessed by preincubating effector cells with ILT2 antibody. While blocking ILT2 engagement has no appreciable effect on cytotoxicity, it increases antiviral IFN-gamma production by approximately threefold in both normal and HIV infected donors. Thus, ILT2 expression, increased on antiviral CD8 cells in chronic infection, may interfere with protective CD8 T-cell function by suppressing IFN-gamma production. PMID- 15270724 TI - Transferrin is required for early T-cell differentiation. AB - Transferrin, the major plasma iron carrier, mediates iron entry into cells through interaction with its receptor. Several in vitro studies have demonstrated that transferrin plays an essential role in lymphocyte division, a role attributed to its iron transport function. In the present study we used hypotransferrinaemic (Trf(hpx/hpx)) mice to investigate the possible involvement of transferrin in T lymphocyte differentiation in vivo. The absolute number of thymocytes was substantially reduced in Trf(hpx/hpx) mice, a result that could not be attributed to increased apoptosis. Moreover, the proportions of the four major thymic subpopulations were maintained and the percentage of dividing cells was not reduced. A leaky block in the differentiation of CD4(-) CD8(-) CD3(-) CD44(-) CD25(+) (TN3) into CD4(-) CD8(-) CD3(-) CD44(-) CD25(-) (TN4) cells was observed. In addition, a similar impairment of early thymocyte differentiation was observed in mice with reduced levels of transferrin receptor. The present study demonstrates, for the first time, that transferrin itself or a pathway triggered by the interaction of transferrin with its receptor is essential for normal early T-cell differentiation in vivo. PMID- 15270725 TI - An investigation of the ability of orally primed and tolerised T cells to help B cells upon mucosal challenge. AB - The oral delivery of soluble antigens induces unresponsiveness to systemic challenge that can be demonstrated as a reduced ability of tolerised T cells to support B-cell expansion and antibody production. However, it remains controversial whether previously induced oral tolerance results in suppression or priming, or has no effect on B-cell responses upon oral challenge. Using a double adoptive transfer system, we primed or tolerised T cells (independently of B cells) with a high dose of fed antigen, and examined the ability of these primed or tolerised T cells to support B-cell clonal expansion in response to orally delivered conjugated antigen. We demonstrated directly in vivo that, in contrast to orally primed T cells, transgenic T cells tolerised by feeding a high dose of antigen are incapable of providing cognate help to support B-cell clonal expansion and antibody production in response to oral challenge. This defect appears to be a result of a reduced ability of orally tolerised transgenic T cells to clonally expand and migrate to B-cell follicles after oral challenge. PMID- 15270726 TI - 4-1BB and OX40 stimulation enhance CD8 and CD4 T-cell responses to a DNA prime, poxvirus boost vaccine. AB - 4-1BB (CD137) is a tumour necrosis factor receptor (TNFR) family member, expressed primarily on CD8 T cells after activation. Signalling through 4-1BB has been reported to enhance CD8 T-cell expansion and to protect activated CD8 T cells from death, resulting in an enlarged memory population. Although stimulating 4-1BB has been shown to significantly improve the immune response to weak immunogens such as tumours, little is known about its effect on the CD8 T cell response to a powerful viral vector such as vaccinia. To test 4-1BB's ability to improve the murine CD8 T cell response to a DNA prime, poxvirus boost vaccine, similar to those used for human immunodeficiency virus and simian immunodeficiency virus vaccines, we administered 4-1BB agonist antibody at the time of the poxvirus boost. 4-1BB stimulation increased the number of functional memory CD8 T cells by two- to fourfold. However, we saw a similar enhancement at the peak of the response and in the memory phase, thus we found no evidence in the context of virus infection that 4-1BB stimulation could increase the percentage of CD8 T cells that survive the acute activation phase to become memory cells. OX40 (CD134) is an analogous TNFR family member expressed primarily on activated CD4 T cells. OX40 stimulation increased the number of antigen specific CD4 T cells approximately threefold. Stimulating both 4-1BB and OX40 enhanced the CD8 T-cell response more than 4-1BB alone. Thus stimulating these receptors can improve the response to a powerful virus vector, and may be useful in vaccine development. PMID- 15270727 TI - Ubiquitin-fusion degradation pathway plays an indispensable role in naked DNA vaccination with a chimeric gene encoding a syngeneic cytotoxic T lymphocyte epitope of melanocyte and green fluorescent protein. AB - Antitumour immunity against murine melanoma B16 was achieved by genetic immunization with a naked chimeric DNA encoding a fusion protein linking green fluorescent protein (GFP) to the N-terminus of a major CD8(+) cytotoxic T lymphocyte (CTL) epitope of tyrosinase-related protein 2 (TRP-2(181-188)) of murine melanoma, designated as pGFP-TRP-2. Tumour growth was profoundly suppressed in C57BL/6 mice immunized with pGFP-TRP-2, while mice vaccinated with pTRP-2 showed rapid tumour growth and died within 40 days after tumour challenge. Splenocytes of mice immunized with pGFP-TRP-2 showed high CTL activity specific for TRP-2(181-188). GFP-TRP-2 expressed in COS-7 cells was rapidly degradated in vitro and the degradation was almost completely prevented by adding a proteasome inhibitor, MG-132, in the culture. Furthermore, the antimelanoma immunity induced by genetic immunization with pGFP-TRP-2 was completely cancelled in mice deficient in proteasome activator PA28alpha/beta. Taken together, GFP-TRP-2 processed by cytosolic proteasome played a central role in breaking peripheral tolerance to a melanoma/melanocyte antigen, TRP-2(181-188), by activating CD8(+) CTL specific for TRP-2(181-188). TRP-2(181-188) fused to GFP may be readily cut off from GFP by the ubiquitin-fusion degradation (UFD) pathway and efficiently presented to major histocompatibility complex class I molecules, resulting in effective induction of CD8(+) T cells specific for the CTL epitope. Furthermore, CD4(+) T cells specific for GFP were shown to play a crucial role in the antimelanoma immunity, probably potentiating activity of TRP-2-specific CTL and/or the "ubiquitin-proteasome pathway". It is noteworthy to document that genetic immunization with pGFP plus pTRP-2(181-188) failed to exert the antitumour immunity. PMID- 15270728 TI - Deficiency of BLNK hampers PLC-gamma2 phosphorylation and Ca2+ influx induced by the pre-B-cell receptor in human pre-B cells. AB - B-cell linker protein (BLNK) is a component of the B-cell receptor (BCR) as well as of the pre-BCR signalling pathway, and BLNK(-/-) mice have a block in B lymphopoiesis at the pro-B/pre-B cell stage. A recent report described the complete loss or drastic reduction of BLNK expression in approximately 50% of human childhood pre-B acute lymphoblastic leukaemias (ALL), therefore we investigated BLNK expression in human pre-B ALL cell lines. One of the four cell lines tested, HPB-NULL cells, was found to lack BLNK expression, and we used these human pre-B ALL cell lines that express and do not express BLNK to investigate the intracellular signalling events following pre-BCR cross-linking. When pre-BCR was cross-linked with anti-micro heavy-chain antibodies, significant phosphorylation of intracellular molecules, including Syk, Shc, ERK MAP kinase, and AKT, and an activation of Ras were observed without regard to deficiency of BLNK expression, suggesting that BLNK is not required for pre-BCR-mediated activation of MAP kinase and phosphatidyl-inositol 3 (PI3) kinase signalling. By contrast, phospholipase C-gamma2 (PLC-gamma2) phosphorylation and an increase in intracellular Ca(2+) level mediated by pre-BCR cross-linking were observed only in the BLNK-expressing cells, indicating that BLNK is essential for PLC-gamma2 induced Ca(2+) influx. Human pre-B cell lines expressing and not expressing BLNK should provide an in vitro model for investigation of the role of BLNK in the pre BCR-mediated signalling mechanism. PMID- 15270729 TI - Multiple cleavage sites for polymeric immunoglobulin receptor. AB - Human polymeric immunoglobulin receptor (pIgR) was expressed in baby hamster kidney (BHK) cells using a recombinant vaccinia virus transfection system. Cleavage of pIgR on the cell surface was partially inhibited by the proteinase inhibitor, leupeptin. We addressed the question whether some particular regions of pIgR could affect the efficient cleavage of this molecule, with the following results: (1) a mutant lacking the entire cytoplasmic region resulted in release of secretory component (SC) into the culture supernatant much faster than wild type; (2) a pIgR mutant lacking the entire extracellular domain 6, the region containing the susceptible cleavage sites, could be cleaved and released as a mutant SC. The transport kinetics of this mutant between endoplasmic reticulum (ER) and Golgi or Golgi and the cell surface was equivalent to wild-type pIgR. Our results indicate that although the main cleavage site is in domain 6, at least one other cleavage site may exist. PMID- 15270730 TI - The use of membrane translocating peptides to identify sites of interaction between the C5a receptor and downstream effector proteins. AB - The complement fragment C5a is a potent leucocyte chemoattractant and activator, mediating its effects through a G-protein-coupled receptor. Whilst the C-terminal domain of this receptor has been shown to be essential for receptor desensitization and internalization, it is not known which domains couple to the receptor's heterotrimeric G proteins. In this report we have used a membrane translocating sequence (MTS) to examine the effects of the four intracellular domains of the human C5a receptor (C5aR) on the receptor's signalling via G(alphai) family heterotrimeric G proteins in intact RBL-2H3 cells. The results indicate that all of the intracellular domains couple to downstream signalling, with the proximal region of the C terminus being a major binding site and intracellular loop 3 playing a role in G protein activation or receptor desensitization. PMID- 15270731 TI - Expression of interleukin-13 receptor alpha 1-subunit on peripheral blood eosinophils is regulated by cytokines. AB - Interleukin-13 (IL-13) is critical for the development of allergic asthma and is involved in the activation of eosinophils within the airways. IL-13 exerts its activity on target cells via the dimeric IL-13 receptor (IL-13R), which comprises the IL-13 receptor alpha1-chain (IL-13Ralpha1) as a specific component. The aim of this study was to investigate the expression of the IL-13Ralpha1-chain on primary human eosinophilic granulocytes. Furthermore, it addresses the regulatory influence of cytokines on the level of surface abundance of this receptor subunit. Expression of IL-13- and IL-4-receptor subunits in purified primary human eosinophils was monitored at the messenger RNA level by reverse transcription polymerase chain reaction and at the protein level by flow cytometry. For the analysis of IL-13Ralpha1 surface expression, a new monoclonal antibody, which was generated using genetic immunization, was employed. Different cytokines with established activity on eosinophils were studied with regard to their influence on IL-13Ralpha1 in vitro by flow cytometry. Whereas IL-13 and IL 4 had inhibitory effects on IL-13Ralpha1 expression on eosinophils, interferon gamma, tumour necrosis factor-alpha, and, to the largest extent, transforming growth factor-beta, enhanced the expression of this receptor subunit. A positive regulatory response evoked by transforming growth factor-beta and interferon gamma does not prevent inhibitory effects caused by IL-13. These findings suggest a regulatory cytokine network influencing the reactivity of eosinophils to IL-13. PMID- 15270732 TI - Danger signals derived from stressed and necrotic epithelial cells activate human eosinophils. AB - Eosinophilic granulocytes are found in tissues with an interface with the external environment, such as the gastrointestinal, genitourinary and respiratory tracts. These leucocytes have been associated with tissue damage in a variety of diseases. The aim of this study was to evaluate whether necrotic epithelial cells can activate eosinophils. The danger theory postulates that cells of the innate immune system primarily recognize substances that signal danger to the host. We damaged epithelial cell lines derived from the genital (HeLa cells), respiratory (HEp-2 cells) and intestinal tracts (HT29 cells) and assessed their capacity to cause eosinophilic migration, release of putative tissue-damaging factors, such as eosinophil peroxidase (EPO) and eosinophil cationic protein (ECP), as well as secretion of tissue-healing factors, e.g. fibroblast growth factors (FGF)-1 and 2 and transforming growth factor (TGF)-beta1. We found that necrotic intestinal cells induced chemotaxis in human eosinophils. EPO release was elicited in eosinophils stimulated with necrotic cells derived from all cell lines, as well as from viable HEp-2 and HT29 cells. Release of ECP was only seen in eosinophils incubated with necrotic intestinal or genital cells, not viable ones. Both necrotic intestinal and genital cells elicited FGF-2 secretion from eosinophils. TGF-beta1 was released from eosinophils exposed to viable and necrotic HT29 cells. These findings indicate that eosinophils are able to recognize and be activated by danger signals released from damaged epithelial cells, which may be of importance in understanding the role of eosinophils in the various inflammatory conditions in which they are involved. PMID- 15270733 TI - Chronic intestinal nematode infection induces Stat6-independent interleukin-5 production and causes eosinophilic inflammatory responses in mice. AB - The role of Stat6 (signal transducers and activators of transcription) in the recruitment and activation of eosinophils has been studied in detail in asthma and other allergic diseases. In this study, we demonstrated that eosinophil responses occur in a Stat6-independent manner in mice infected with the intestinal nematode, Nippostrongylus brasiliensis. Stat6-deficient (Stat6(-/-)) mice cannot expel N. brasiliensis and establish chronic infections. Prominent blood and intestinal eosinophilia were induced after day 14 postinfection (p.i.) and maintained at this level in Stat6(-/-) mice, whereas in wild-type mice eosinophil responses reached a peak on day 10 p.i. and declined thereafter. The introduction of a secondary infection of N. brasiliensis into wild-type mice induced rapid and exaggerated eosinophilia, whereas secondary infection in Stat6( /-) mice resulted in almost the same eosinophil responses as those of the primary infection, suggesting a lack of memory responses. Blood eosinophilia was also induced in Stat6(-/-) mice implanted with N. brasiliensis in the small intestine, suggesting that intestinal exposure to parasitic antigen is sufficient to induce eosinophil responses. Furthermore, this prominent eosinophil response of Stat6(-/ ) mice after day 14 was closely associated with an increase of interleukin (IL)-5 production in serum and intestine. Neither IL-4 nor eotaxin were significantly induced in Stat6(-/-) mice after infection with N. brasiliensis. We also found that mRNA for IL-5, GATA-3 and eosinophil peroxidase (EPO) are induced in the intestine of Stat6(-/-) mice on day 14 p.i. Taken together, these results provide evidence for Stat6-independent IL-5 production and subsequent eosinophil responses after chronic infection with N. brasiliensis. PMID- 15270734 TI - Inhibition of murine allergic airway disease by Bordetella pertussis. AB - Whole-cell pertussis vaccines have been shown to selectively induce T helper 1 (Th1)-type responses in human and animals. In this study, we investigated whether whole-cell B. pertussis could inhibit allergic airway reactions in a murine model of asthma induced by ovalbumin (OVA). Systemic administration of whole-cell B. pertussis strongly inhibited allergic airway reactions such as eosinophil recruitment into the airway, lung inflammation, and airway hyperresponsiveness to methacholine. The inhibitory effect of whole-cell B. pertussis was mediated by chromosomal DNA and pretreatment of DNA with CpG methylase or DNase I resulted in a loss of the inhibitory effect. Treatment of animals with B. pertussis DNA significantly decreased the Th2 cytokine (interleukins IL-4 and IL-5) concentrations in the airways without increasing Th1 cytokines. These results suggest that B. pertussis DNA containing unmethylated CpG appears to be responsible for the inhibitory effect of whole cell B. pertussis on the allergic airway reactions through inhibition of the Th2 response. PMID- 15270735 TI - Reversal of established CD4+ type 2 T helper-mediated allergic airway inflammation and eosinophilia by therapeutic treatment with DNA vaccines limits progression towards chronic inflammation and remodelling. AB - Immunostimulatory DNA-based vaccines can prevent the induction of CD4(+) type 2 T helper (Th2) cell-mediated airway inflammation in experimental models, when administered before or at the time of allergen exposure. Here we demonstrate their efficacy in limiting the progression of an established response to chronic pulmonary inflammation and airway remodelling on subsequent allergen challenge. Mice exhibiting Th2-mediated airway inflammation induced following sensitization and challenge with group 1 allergen derived from Dermatophagoides pteronyssinus group species (Der p 1), a major allergen of house dust mite, were treated with pDNA vaccines. Their airways were rechallenged and the extent of inflammation assessed. In plasma DNA (pDNA)-vaccinated mice, infiltration of inflammatory cells, goblet cell hyperplasia and mucus production were reduced and subepithelial fibrosis attenuated. The reduction in eosinophil numbers correlated with a fall in levels of the profibrotic mediator transforming growth factor (TGF)-beta1 in bronchoalveolar lavage (BAL) and lung tissue. In addition to lung epithelial cells and resident alveolar macrophages, infiltrating eosinophils, the principle inflammatory cells recruited following allergen exposure, were a major source of TGF-beta1. Protection, conferred irrespective of the specificity of the pDNA construct, did not correlate with a sustained increase in systemic interferon (IFN)-gamma production but in a reduction in levels of the Th2 pro inflammatory cytokines. Notably, there was a reduction in levels of interleukin (IL)-5 and IL-13 produced by systemic Der p 1 reactive CD4(+) Th2 cells on in vitro stimulation as well as in IL-4 and IL-5 levels in BAL fluid. These data suggest that suppression of CD4(+) Th2-mediated inflammation and eosinophilia were sufficient to attenuate progression towards airway remodelling. Immunostimulatory DNA may therefore have a therapeutic application in treatment of established allergic asthma in patients. PMID- 15270736 TI - Dual biological functions of an interleukin-1 receptor antagonist-interleukin-10 fusion protein and its suppressive effects on joint inflammation. AB - The aim of this study was to construct and purify a novel interleukin-1 receptor antagonist (IL-1ra)-interleukin-10 (IL-10) fusion protein and determine its biological function and anti-inflammatory effects. The isolated cDNAs of two inhibitory cytokines (IL-1ra, IL-10) were used to construct a cDNA for the IL-1ra IL-10 fusion protein. The expressed recombinant cytokines and fusion product were purified and their biological properties analysed. The anti-IL-1 effect was evaluated by using a thymocyte-proliferation assay, and the IL-10 effect was investigated by the inhibition of interferon-gamma (IFN-gamma) production from splenocytes. The clinical response and histological analyses were studied in an adjuvant arthritic rat model. The fusion protein was 38 000 molecular weight in size. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting demonstrated that the purified protein was recognized by both IL 1ra and IL-10 antibodies. The fusion protein significantly inhibited IL-1 mediated thymocyte proliferation and concanavalin A (ConA)-primed IFN-gamma production from splenocytes. The fusion protein also suppressed joint swelling (paw circumference reduced from 5.0 +/- 0.2 to 4.1 +/- 0.1 cm; paw thickness approximately 2 mm in difference) and synovial inflammation in adjuvant arthritis of rats. Our investigations indicate that this fusion protein effectively suppresses inflammatory arthritis and may initiate a trend for future clinical application to target multiple molecules at the same time. PMID- 15270737 TI - Enteroaggregative Escherichia coli flagellin-induced interleukin-8 secretion requires Toll-like receptor 5-dependent p38 MAP kinase activation. AB - Enteroaggregative Escherichia coli (EAEC) is an emerging enteric pathogen that causes acute and chronic diarrhoea in a number of clinical settings. EAEC diarrhoea involves bacterial aggregation, adherence to intestinal epithelial cells and elaboration of several toxigenic bacterial mediators. Flagellin (FliC EAEC), a major bacterial surface protein of EAEC, causes interleukin (IL)-8 release from several epithelial cell lines. The host response to flagellins from E. coli and several other bacteria is mediated by Toll-like receptor 5 (TLR5), which signals through nuclear factor kappa B (NF-kappaB) to induce transcription of pro-inflammatory cytokines. p38 mitogen-activating protein (MAP) kinase (MAPK) is a member of a family of stress-related kinases that influences a diverse range of cellular functions including host inflammatory responses to microbial products. We studied the role of p38 MAPK in FliC-EAEC-induced IL-8 secretion from Caco-2 human intestinal epithelial cells and THP-1 human monocytic cells. We found that IL-8 secretion from both cell types is dependent on p38 MAPK, which is phospho-activated in response to FliC-EAEC. The role of TLR5 in p38 MAPK dependent IL-8 secretion was verified in HEp-2 cells transiently transfected with a TLR5 expression construct. Activation of interleukin-1 receptor-associated kinase (IRAK) was also observed in Caco-2 and TLR5-transfected HEp-2 cells after exposure to FliC-EAEC. Finally, we demonstrated that pharmacological inhibition of p38 MAPK reduced IL-8 transcription and mRNA levels, but did not affect NF kappaB activation. Collectively, our results suggest that TLR5 mediates p38 MAPK dependent IL-8 secretion from epithelial and monocytic cells incubated with FliC EAEC, and that this effect requires IL-8 promoter activation independent of NF kappaB nuclear migration. PMID- 15270738 TI - Airways infection with virulent Mycobacterium tuberculosis delays the influx of dendritic cells and the expression of costimulatory molecules in mediastinal lymph nodes. AB - Despite tuberculosis resurgence and extensive dendritic cell (DC) research, there are no in vivo studies evaluating DC within regional lymphoid tissue during airways infection with virulent Mycobacterium tuberculosis (Mtb) H37Rv. Using DC specific antibodies, immunocytochemistry, flow cytometry and Ziehl-Neelsen (ZN) for bacilli staining, we searched for Mtb and DC changes within mediastinal lymph nodes, after intratracheal (ITT) inoculation of virulent Mtb. ZN and immunocytochemistry in frozen and paraffin sections of mediastinal lymph nodes identified Mtb until day 14 after ITT inoculation, associated with CD11c(+) and Dec205(+) DC. Analysing CD11c, MHC-CII, and Dec205 combinations by flow cytometry in MLN suspensions revealed that CD11c(+)/MHC-CII(+) and CD11c(+)/Dec205(+) DC did not increase until day 14, peaked on day 21, and sharply declined by day 28. No changes were seen in control, saline-inoculated animals. The costimulatory molecules evaluated in CD11c(+) DCs followed a similar trend; the CD80 increase was negligible, slightly surpassed by CD40. CD86 increased earlier and the three markers peaked at day 21, declining by day 28. While antigen-specific proliferation was not evident for MLN CD4(+) T cells at 2 weeks postinfection, delayed-type hypersensitivity responses upon ITT inoculation revealed that, as early as day 3 and 7, both the priming and peripheral systemic immune responses were clearly established, persisting until days 14-21. While airways infection with virulent Mtb triggers an early, systemic peripheral response maintained for three weeks, this seems dissociated from regional events within mediastinal lymph nodes, such as antigen-specific T-cell reactivity and a delay in the influx and local activation of DC. PMID- 15270739 TI - Increased expression of mRNA encoding interleukin (IL)-4 and its splice variant IL-4delta2 in cells from contacts of Mycobacterium tuberculosis, in the absence of in vitro stimulation. AB - Expression of interleukin (IL)-4 is increased in tuberculosis and thought to be detrimental. We show here that in healthy contacts there is increased expression of its naturally occurring antagonist, IL-4delta2 (IL-4delta2). We identified contacts by showing that their peripheral blood mononuclear cells (PBMC) released interferon (IFN)-gamma in response to the Mycobacterium tuberculosis-specific antigen 6 kDa early secretory antigenic target (ESAT-6). Fresh unstimulated PBMC from these contacts contained higher levels of mRNA encoding IL-4delta2 (P=0.002) than did cells from ESAT-6 negative donors (noncontacts). These data indicate that contact with M. tuberculosis induces unusual, previously unrecognized, immunological events. We tentatively hypothesize that progression to active disease might depend upon the underlying ratio of IL-4 to IL-4delta2. PMID- 15270740 TI - Differential production of interleukin-10 and interleukin-12 in mononuclear cells from leprosy patients with a Toll-like receptor 2 mutation. AB - Toll-like receptor 2 (TLR2) is a key mediator of the immune response to mycobacterial infections, and mutations in TLR2 have been shown to confer susceptibility to infection with mycobacteria. This study investigated the profiles of cytokines, such as interferon (IFN)-gamma, interleukin (IL)-10, IL-12 and tumour necrosis factor (TNF)-alpha in response to Mycobacterium leprae in peripheral blood mononuclear cells (PBMC) with the TLR2 mutation Arg677Trp, a recently reported polymorphism that is associated with lepromatous leprosy. In leprosy patients with the TLR2 mutation, production of IL-2, IL-12, IFN-gamma, and TNF-alpha by M. leprae-stimulated PBMC were significantly decreased compared with that in groups with wild-type TLR2. However, the cells from patients with the TLR2 mutation showed significantly increased production of IL-10. There was no significant difference in IL-4 production between the mutant and wild-type during stimulation. Thus, these results suggest that the TLR2 signal pathway plays a critical role in the alteration of cytokine profiles in PBMC from leprosy patients and the TLR2 mutation Arg677Trp provides a mechanism for the poor cellular immune response associated with lepromatous leprosy. PMID- 15270741 TI - Total family unit Helicobacter pylori eradication and pediatric re-infection rates. AB - BACKGROUND: Re-infection with Helicobacter pylori is more common in children than adults, and it is generally accepted that the family unit plays a significant role in primary childhood infection. We investigated whether the family unit plays a significant role in pediatric re-infection and if eradication of H. pylori from the entire family reduces the risk of childhood re-infection. METHODS: Fifty families, each with an H. pylori-infected pediatric index case (mean age 9.48 years), were recruited. A 13carbon urea breath test was performed on all family members in the same house as the index case. Each family unit was randomized into a 'family unit treatment' group (all infected family members treated) or an 'index case treatment' group (index case only treated). RESULTS: At long-term follow-up (mean 62.2 months), there were three re-infected children in the 'index case treatment' group compared with one in the 'family unit treatment' group. The re-infection rate was 2.4% per patient per year in the 'index case treatment' group and 0.7% per patient per year in the 'family unit treatment' group (p = .31). CONCLUSIONS: This study is the first to evaluate the effect of total family unit H. pylori eradication on pediatric re-infection rates and reports the longest period of re-infection follow-up in children. In childhood, re-infection with H. pylori is not significantly reduced by family unit H. pylori eradication. PMID- 15270742 TI - IgG subclass responses in childhood Helicobacter pylori duodenal ulcer: evidence of T-helper cell type 2 responses. AB - BACKGROUND: Duodenal ulcer in adults chronically infected with Helicobacter pylori is associated with a polarized T-helper cell type 1 (Th1) mucosal immune response, with a predominantly immunoglobulin G2 (IgG2) systemic specific response. It has been suggested that children colonized by H. pylori also produce a mucosal Th1 response, but there are few studies that have measured IgG subclass responses in children with duodenal ulcer. MATERIALS AND METHODS: Seven children with endoscopically proven duodenal ulcer and H. pylori infection and 18 children with biopsy proven H. pylori infection but no duodenal ulcer had relative concentrations of IgG subclass responses (IgGsc) against H. pylori antigens measured by ELISA. Eighteen IgG seropositive adults acted as controls. The range of antigens recognised by IgG1 and IgG2 subclass responses were investigated by Western blots. RESULTS: There were no differences in mean IgGsc responses between children with or without duodenal ulcer. Adults produced an IgG2 predominant response. Western blots showed no qualitative differences in antigens recognised by IgG1 or IgG2. CONCLUSION: Children with duodenal ulcer, in contrast to adults, produce an IgGsc response consistent with a mucosal Th2 response to H. pylori regardless of the presence of duodenal ulceration. This suggests that disease causation amongst children with H. pylori associated duodenal ulceration may not be dependant upon a mucosal Th1 biased response. PMID- 15270743 TI - Dual vs. triple therapy for childhood Helicobacter pylori gastritis: a double blind randomized multicentre trial. AB - BACKGROUND: Data on the efficacy of eradication treatment for Helicobacter pylori gastritis in children are scarce. AIM: To evaluate the efficacy of triple therapy with lansoprazole plus amoxicillin and tinidazole vs. dual therapy with amoxicillin and tinidazole in a double-blind randomized multicentre trial, and the usefulness of eradication in terms of long-term symptom resolution. SUBJECTS: We enrolled 43 consecutive children undergoing endoscopy for upper gastrointestinal dyspepsia with H. pylori gastritis. They underwent a 13C-urea breath test, completed a 2-week symptom diary card, and were randomized. Treatment was given in a Redidose box (Redidose Company Ltd., Brighton, UK) containing either lansoprazole-amoxicillin-tinidazole (triple therapy) or placebo plus amoxicillin-tinidazole (dual therapy) for 1 week. The completion of a 2-week symptom diary card and the performance of a breath test were repeated 6 weeks and 6 months after the end of therapy. One to two years later, a structured telephone interview was conducted with 36 of the children. RESULTS: According to the breath test, 6 weeks after the end of therapy H. pylori was eradicated in 15 of 22 children on triple therapy [68.2%; 95% confidence interval (CI) = 45-88] and in 15 of 21 children on dual therapy (71%; 95% CI = 48-89; not significant), and 6 months after the end of therapy it was eradicated in 16 of 22 children on triple therapy (72.7%) and in 15 of 21 children on dual therapy. Six months after therapy, symptoms were analysed in 11 H. pylori-positive and 31 H. pylori negative children, and it was found that dyspeptic symptoms had disappeared or improved in both groups, with no difference between them. One to two years later, 36 children were interviewed. Epigastric pain had recurred in three of 26 H. pylori-negative and in seven of 10 H. pylori-positive children (p = .001); in three of the latter, pain was severe and required additional treatment. CONCLUSION: One-week triple or dual therapy with two antibiotics achieved similar eradication rates. Soon after treatment, symptoms disappeared or improved in most children irrespective of eradication, but epigastric pain recurred in the majority of the still-infected children within 2 years. PMID- 15270744 TI - A novel in vitro infection model of Helicobacter pylori using mucin-producing murine gastric surface mucous cells. AB - BACKGROUND: Helicobacter pylori is found within the gastric surface mucous gel layer and in the epithelial surface. Gastric cancer cells have been used in experimental H. pylori infection in vitro, although cancer cells have some abnormalities in cellular properties. The aim of this study was to develop an in vitro H. pylori infection model using normal gastric surface cells that produce gastric mucin. MATERIALS AND METHODS: Normal murine gastric surface mucous cells (GSM06) were cultured by the liquid interface method using a serum-free medium and a collagen gel containing a fibroblast cell line (L929) and infected with H. pylori. Infection by H. pylori was assessed by enumerating the colony-forming units (CFU) of H. pylori adhered to GSM06 cells and by transmission electron microscopy. The production of mucin was determined by a lectin binding assay, sugar analysis, and MUC5AC gene expression. RESULTS: GSM06 cells cultured under these conditions produced mucin containing N-acetylgalactosamine and MUC5AC as the core protein. Significantly higher numbers of H. pylori adhered to GSM06 cells under mucin-producing conditions than under nonproducing conditions. Microscopic observation showed a filamentous structure resembling a type IV secretion system apparatus formed between the surface of GSM06 cells and H. pylori. CONCLUSIONS: This study demonstrates a novel in vitro H. pylori infection model using mucin-producing murine GSM06 cells for early stages of infection. PMID- 15270745 TI - Comparative chemical and biological characterization of the lipopolysaccharides of gastric and enterohepatic helicobacters. AB - BACKGROUND: The lipopolysaccharide of Helicobacter pylori plays an important role in colonization and pathogenicity. The present study sought to compare structural and biological features of lipopolysaccharides from gastric and enterohepatic Helicobacter spp. not previously characterized. MATERIALS AND METHODS: Purified lipopolysaccharides from four gastric Helicobacter spp. (H. pylori, Helicobacter felis, Helicobacter bizzozeronii and Helicobacter mustelae) and four enterohepatic Helicobacter spp. (Helicobacter hepaticus, Helicobacter bilis, 'Helicobacter sp. flexispira' and Helicobacter pullorum) were structurally characterized using electrophoretic, serological and chemical methods. RESULTS: Structural insights into all three moieties of the lipopolysaccharides, i.e. lipid A, core and O-polysaccharide chains, were gained. All species expressed lipopolysaccharides bearing an O-polysaccharide chain, but H. mustelae and H. hepaticus produced truncated semirough lipopolysaccharides. However, in contrast to lipopolysaccharides of H. pylori and H. mustelae, no blood group mimicry was detected in the other Helicobacter spp. examined. Intra-species, but not interspecies, fatty acid profiles of lipopolysaccharides were identical within the genus. Although shared lipopolysaccharide-core epitopes with H. pylori occurred, differing structural characteristics were noted in this lipopolysaccharide region of some Helicobacter spp. The lipopolysaccharides of the gastric helicobacters, H. bizzozeronii and H. mustelae, had relative Limulus amoebocyte lysate activities which clustered around that of H. pylori lipopolysaccharide, whereas H. bilis, 'Helicobacter sp. flexispira' and H. hepaticus formed a cluster with approximately 1000-10,000-fold lower activities. H. pullorum lipopolysaccharide had the highest relative Limulus amoebocyte lysate activity of all the helicobacter lipopolysaccharides (10-fold higher than that of H. pylori lipopolysaccharide), and all the lipopolysaccharides of enterohepatic Helicobacter spp. were capable of inducing nuclear factor-Kappa B(NF-kappaB) activation. CONCLUSIONS: The collective results demonstrate the structural heterogeneity and pathogenic potential of lipopolysaccharides of the Helicobacter genus as a group and these differences in lipopolysaccharides may be indicative of adaptation of the bacteria to different ecological niches. PMID- 15270746 TI - Role of Lewis A and Lewis B blood group antigens in Helicobacter pylori infection. AB - BACKGROUND AND AIMS: We investigated the prevalence of Helicobacter pylori infection in a large group of women to determine whether there was an association of current infection status with Lewis blood group antigen A and B phenotype. METHODS: Between November 2000 and November 2001, mothers were recruited after delivery of their offspring at the Department of Gynecology and Obstetrics at the University of Ulm, Ulm, Germany. The H. pylori infection status of the women was determined by 13C urea breath test. Their Lewis A and Lewis B phenotype was determined using standard laboratory techniques. RESULTS: In total, 22.2% of the 712 women included in the study (mean age 30.7 years) had a current H. pylori infection. The prevalence of infection varied from 15.5% in women of German nationality to 75.0% in women of Turkish nationality (p < .001). Most women (68.1%) had a Le(a-b+) phenotype. The prevalence of H. pylori infection in women with Le(a-b+) phenotypes was lower than in other women (p = .02). In multivariate analysis, the odds ratio (OR) for a current H. pylori infection given Le(a-b+) was 0.56 [95% confidence interval (CI) 0.33-0.95] compared to women with Le(a-b ). CONCLUSION: Le(a-b+) blood group phenotype in combination with secretor status may hinder colonization of H. pylori in the population studied. PMID- 15270747 TI - Polymorphism of the Helicobacter pylori feoB gene in Korea: a possible relation with iron-deficiency anemia? AB - BACKGROUND: Helicobacter pylori is a causative agent of gastritis, and H. pylori infection is thought to be correlated with iron-deficiency anemia (IDA) at puberty. The H. pylori feoB gene product, a high-affinity ferrous iron transporter, plays a central role in iron acquisition and virulence. This study was undertaken to analyze H. pylori feoB status according to clinical data, including antral gastritis with or without IDA. METHODS: Fourteen H. pylori positive patients aged from 10 to 18 years were categorized into subgroups based on the presence or absence of IDA. Eight patients were diagnosed as having IDA; the other six showed normal hematological findings. Genomic DNA was isolated from H. pylori cultured from each gastric biopsy specimen. Five sets of primers were used for the PCR amplification of the feoB gene. Linking and sequencing of PCR products generated the feoB region, which was 1.93 kb in size. The feoB gene sequences of H. pylori J99 and 26695 were compared with the clinical strains, and the sequences of feoB regions in the IDA (+) and (-) groups were compared. RESULTS: Sequence analysis of the complete coding region of the feoB gene revealed 16 sites of polymorphism or mutation. Among these, three polymorphisms (E/T254A, I263V, and K511Q) were indigenous to the Korean clinical strains. Although statistically significant differences were observed at four sites (K127T, A273S/P, I438V and I441T) between IDA (+) and (-), the number of specimens was too low to assess the significance of the differences. CONCLUSION: The four polymorphisms of the feoB gene observed appear to be related to the clinical phenotype of IDA, but the relation is unclear because of the small number of strains studied. Further studies are required to confirm a correlation between IDA and H. pylori infection. PMID- 15270748 TI - Seroconversion and seroreversion of Helicobacter pylori antibodies over a 9-year period and related factors in Japanese adults. AB - BACKGROUND: There are still insufficient data on the frequency of seroconversion and seroreversion of Helicobacter pylori antibodies. The frequency of serochange and related factors were investigated in this study over 9 years. SUBJECTS AND METHODS: Using sera from 3104 workers who underwent health checks in 1989 and again in 1998, H. pylori antibodies were measured. Those with intermediate serology were excluded from the study. Information on past history was collected using a questionnaire. RESULTS: Of the 912 seronegative and 1286 seropositive subjects in 1989, seroconversion was observed in 57 and seroreversion in 91 subjects. Seroconversion and seroreversion rates over the 9-year period were 6.3% and 7.1%, respectively, giving rates per 1000 person-years (with 95% confidence interval) of 7.0 (5.2-8.7) and 7.9 (6.3-9.4), respectively. Subjects that reported abdominal symptoms or gastric fiberscope use showed significantly higher seroconversion rates than controls (8.7 vs. 4.5 and 9.2 vs. 1.6, respectively), which remained significant after adjustment for age and gender. Those with a history of duodenal ulcers, a smoking habit or a drinking habit showed significantly lower seroreversion rates than controls (3.5 vs. 8.9, 5.4 vs. 9.2 and 5.9 vs. 13.3, respectively). After adjustment, the association between seroreversion and smoking habit disappeared, while the associations with history of duodenal ulcers and drinking habit remained. CONCLUSIONS: Those with a history of nonspecific abdominal symptoms and those with a history of gastric fiberscope use showed higher seroconversion rates. Alcohol consumption and duodenal ulcers may inhibit the autoeradication of H. pylori, possibly through increased acid secretion. PMID- 15270749 TI - Helicobacter pylori infection and immune thrombocytopenic purpura: an update. AB - Data are accumulating on the association between Helicobacter pylori infection and idiopathic thrombocytopenic purpura (ITP) and the significant increase in platelet count after bacterial eradication. The aim of this review was to consider the studies so far published on H. pylori infection and ITP in order to evaluate a possible correlation between these two conditions. A review of the literature showed that 278 out of the 482 ITP patients investigated (58%) were positive for H. pylori infection and that the bacterium was eradicated in 88% of cases. Eradication therapy was accompanied by a complete or partial platelet response in approximately half the cases. Overall, these data show that H. pylori eradication in patients with ITP is effective in increasing platelet count. However, because the studies so far published are few, are sometimes controversial and involve small series of patients, further studies on larger numbers of patients with longer follow-up are needed to confirm these preliminary findings. PMID- 15270750 TI - Stool antigen test for the diagnosis of Helicobacter pylori infection: a systematic review. AB - Our aim was to review systematically the diagnostic accuracy of the Helicobacter pylori stool antigen test. Bibliographical searches were performed in several electronic databases and abstracts from congresses up to May 2003. Eighty-nine studies (10,858 patients) evaluated the stool antigen test in untreated patients. Mean sensitivity, specificity, positive predictive value and negative predictive value were 91%, 93%, 92% and 87%, respectively. Analysis of the eight studies (1399 patients) in which pretreatment evaluation of the monoclonal stool antigen test was performed showed better (p < .001) results (96%, 97%, 96% and 97%, respectively), with a clearer distinction between positive and negative results. Thirty-nine studies (3147 patients) evaluated the stool antigen test for the confirmation of H. pylori eradication 4-8 weeks after therapy, with accuracies of 86%, 92%, 76% and 93% for mean sensitivity, specificity, positive predictive value and negative predictive value, respectively. Results were similar when a gold standard based on at least two methods was used. Relatively low accuracy was reported in some posttreatment studies with the polyclonal stool antigen test. However, excellent results (p < .001) were achieved in all the six studies evaluating the monoclonal stool antigen test 4-8 weeks posttreatment. Results evaluating the stool antigen test < 4 weeks posttreatment are contradictory. Proton-pump inhibitors seem to affect the accuracy of the stool antigen test. Sensitivity and/or specificity in patients with gastrointestinal bleeding may be suboptimal. The stool antigen test performs well in children. Finally, the stool antigen test seems to be a cost-effective method. PMID- 15270751 TI - Helicobacter pylori infection in patients with Hashimoto's thyroiditis. PMID- 15270753 TI - Peer-reviewed publication: a view from inside. PMID- 15270754 TI - Preclinical profile of combinations of some second-generation antiepileptic drugs: an isobolographic analysis. AB - PURPOSE: The need for an efficacious treatment of patients with intractable seizures is urgent and pressing, because approximately 30% of epilepsy patients worldwide are still inadequately medicated with current frontline antiepileptic drugs (AEDs). This study sought to determine the interactions among some newer AEDs [topiramate (TPM), felbamate (FBM), oxcarbazepine (OXC), and lamotrigine (LTG)] in the maximal electroshock-induced seizures (MES) and chimney test (motor performance) in mice, by using the isobolographic analysis. METHODS: Evaluation of the anticonvulsant and acute adverse (neurotoxic) effects in mice produced by the AEDs in combinations at the fixed ratios of 1:3, 1:1, and 3:1 allowed the assessment of their preclinical profile and the determination of benefit indices (BIs) for all individual combinations. RESULTS: Combinations of TPM+FBM at the fixed ratios of 1:3, 1:1, and 3:1 offered supraadditive (synergistic) interactions against electroconvulsions and subadditivity (antagonism) in terms of acute neurotoxic effects in the chimney test (BIs ranged between 1.90 and 2.59, the best combinations from a preclinical point of view). The examined combinations of TPM+OXC also were advantageous due to synergistic interactions in the MES, and additivity in terms of acute neurotoxic effects produced by the AEDs (BIs ranged between 1.35 and 1.71). In contrast, OXC+FBM exerted subadditive (antagonistic) interactions in the MES test and additive interactions in terms of acute motor impairment of animals (BIs ranged between 0.53 and 0.71). The worst combination was observed for OXC+LTG, at the fixed ratio of 1:1, displaying subadditivity (antagonism) against electroconvulsions and supraadditivity (synergy) with respect to neurotoxicity (BIs, 0.43). The remaining combinations of OXC+LTG tested (i.e., 1:3 and 3:1) exerted additivity in the MES test and supraadditivity in the chimney test (BIs 0.54 and 0.49, respectively). None of the studied AEDs affected the brain concentrations of other AEDs, so the existence of any pharmacokinetic interactions to be responsible for the observed effects is improbable. CONCLUSIONS: Based on the current preclinical data, the pharmacological profile of combinations of TPM+FBM and TPM+OXC evaluated with isobolography was beneficial and might be worth recommendation to further clinical practice. In contrast, utmost caution is required during the use of OXC+FBM or OXC+LTG in clinical practice, because of the high risk of neurotoxic adverse effect appearance. PMID- 15270755 TI - Chromosomal mapping of genetic loci controlling absence epilepsy phenotypes in the WAG/Rij rat. AB - PURPOSE: The WAG/Rij rat is among the most appropriate models for the study of spontaneous childhood absence epilepsy, without complex neurologic disorders that are associated with some mouse models for absence epilepsy. Previous studies have allowed the identification of distinct types of spike-wave discharges (SWDs) characterizing seizures in this strain. The purpose of this study was to investigate the genetic basis of electroencephalographic (EEG) properties of SWDs. METHODS: An intercross was derived from WAG/Rij and ACI inbred strains that are known to differ substantially in the number of SWDs. Phenotypic analyses based on 23-h EEG recording in all progenies allowed the quantification of type I and type II SWD phenotypes. A genome-wide scan was performed with 145 microsatellite markers, which were used to test for evidence of genetic linkage to SWD quantitative phenotypes. RESULTS: We were able to map quantitative trait loci independently, controlling type I and type II SWD variables to rat chromosomes 5 and 9. Strongest linkages were obtained for D5Mgh15 and total duration of type II SWD (lod, 3.64) and for D9Rat103 and the average duration of type I SWD (lod, 3.91). These loci were denoted T2swd/wag and T1swd/wag, respectively. CONCLUSIONS: The independent genetic control of type I and type II SWDs underlines the complexity of the molecular mechanisms participating in SWDs. The identification of these genetic loci represents an important step in our fundamental knowledge of the architecture of SWDs and may provide new insights for resolving the genetic heterogeneity of absence epilepsy. PMID- 15270756 TI - Distribution of cortical interneurons in grey matter heterotopia in patients with epilepsy. AB - PURPOSE: Grey matter heterotopia are well-defined malformations of the cortex often associated with severe epilepsy. Defects have been identified in genes, including DCX and FLN1, that influence radial migration of postmitotic cells from the ventricular zone to the cortical plate. A proportion of cortical gamma aminobutyric acid (GABA)-containing interneurons may arise from the ganglionic eminence of the ventral telencephalon. We aimed to identify the subtypes and localisation of interneurons within grey matter heterotopia relative to cortex. METHODS: By using quantitative immunohistochemistry, we studied the density and distribution of interneurons within six cases of grey matter heterotopia in postmortem tissue from patients with epilepsy. RESULTS: In many cases, a suggestion of focal rudimentary laminar arrangement and small reelin-positive cells was identified within the heterotopia. Immunohistochemistry for glutamic acid decarboxylase(65/57), parvalbumin, calbindin, and calretinin showed inhibitory neurons of all subtypes represented within the heterotopia, and of normal morphology. The mean densities of interneurons were overall similar to those of the overlying cortex, but the interneurons showed less organisation and were more randomly orientated compared with cortex. CONCLUSIONS: Interneurons within heterotopia probably arise from the ventricular zone, but their abnormal local organization may influence the epileptogenicity of these lesions. PMID- 15270757 TI - Dialyzability of the antiepileptic drug zonisamide in patients undergoing hemodialysis. AB - PURPOSE: The influence of hemodialysis on plasma zonisamide (ZNS) concentration has not been clarified. In this study, the dialyzability of ZNS during hemodialysis was investigated in four ZNS-treated women with systemic lupus erythematosus complicated by seizures. METHODS: The total and unbound plasma concentrations of ZNS were measured before and after hemodialysis. The concentration of ZNS in the spent dialysate also was determined. RESULTS: The reduction in plasma ZNS concentration after a 4.5-h hemodialysis was 52.0 +/- 7.6%, and the dialyzer (BLF-16GW) clearance of ZNS was 55.1 +/- 7.0 ml/min. Dosage was gradually increased up to 200 to 500 mg/day, and the seizures were controlled satisfactorily. CONCLUSIONS: The plasma concentration of ZNS was reduced by approximately 50% during one session of dialysis. For patients undergoing daytime hemodialysis sessions every 2 or 3 days, the usual dosage of ZNS (4-8 mg/kg/day) may be prescribed once a day in the evening. If seizures occur after hemodialysis, a supplemental daily dose may be prescribed in the morning. PMID- 15270758 TI - Evaluating the utility of inpatient video-EEG monitoring. AB - PURPOSE: Inpatient video-EEG monitoring (VEM) is widely used for the diagnosis, seizure classification, and presurgical evaluation of patients with seizure disorders. It is resource intensive and relatively expensive, so its utility continues to be debated. Few studies have specifically evaluated the utility of inpatient VEM in altering diagnosis or management of patients with seizure disorders. We sought to assess the proportion of patients for whom the preadmission diagnosis and management were altered after inpatient VEM of patients admitted for diagnostic and presurgical evaluation of seizure disorders. METHODS: Data from a consecutive cohort of patients admitted over a 3-year period to an inpatient VEM unit in a tertiary referral hospital were retrospectively analyzed. The preadmission diagnosis and management by the referring neurologist was compared with the diagnosis and management after the VEM. RESULTS: Of 131 patients, 91 (70%) were admitted for diagnostic evaluation and 39 (30%) for a presurgical workup. Mean evaluative period was 5.6 days. Mean number of seizures recorded was 2.9. No seizures were recorded in 31% of patients. Interictal EEG showed epileptiform changes in 56 (43%). In 76 (58%), the diagnosis was altered as a result of the VEM, with the greatest change being an increase in the nonepileptic diagnosis group (7% to 31%) and the generalized diagnosis group (5% to 11%). Management was changed after the VEM in 95 (73%). CONCLUSIONS: The results of this study demonstrate that inpatient VEM has a high yield in changing diagnosis and management. Future long-term cost-benefit studies of the management changes resulting from VEM evaluation will aid in further reinforcing its role. PMID- 15270759 TI - Cardiovascular regulation and hippocampal sclerosis. AB - PURPOSE: Cardiovascular dysregulation has been detected in patients with temporal lobe epilepsy (TLE) by using cardiovascular reflex tests and analysis of heart rate variability (HRV). The two methods have not previously been used in the same study to compare them in the assessment of cardioregulatory function. Magnetic resonance imaging (MRI) is considered the best method to reveal structural changes such as hippocampal sclerosis associated with TLE. It is not known whether these structural changes modify cardioregulatory function in patients with TLE. METHODS: Standard cardiovascular reflex tests and analysis of spectral and dynamic measures from 24-h electrocardiogram (ECG) recordings were performed for eight patients with and 31 patients without hippocampal sclerosis and for 72 control subjects. MRI also was performed in each patient to reveal hippocampal sclerosis. RESULTS: Various measures of cardiovascular reflexes and HRV were diminished in patients with TLE compared with the control subjects. No significant differences were found in the measures obtained from the cardiovascular reflex tests or analysis of HRV between those with and without hippocampal sclerosis, although a nonsignificant trend toward reduced values was seen among those with hippocampal sclerosis. The values of cardiovascular reflexes and spectral analysis of HRV correlated with each other. CONCLUSIONS: These results suggest that functional rather than structural changes related to TLE are involved mainly as a mechanism of altered cardioregulatory function. The cardiovascular reflex test and analysis of HRV both appear to be useful in studying cardioregulation in patients with TLE. PMID- 15270760 TI - Subtle microscopic abnormalities in hippocampal sclerosis do not predict clinical features of temporal lobe epilepsy. AB - PURPOSE: Subtle microdysplastic features are found in some patients with hippocampal sclerosis (HS) and refractory temporal lobe epilepsy. The significance of these findings is unknown. We investigated their frequency, relation to the pattern of HS, and clinical associations. METHODS: One-hundred forty patients with histologically confirmed HS (mean age at operation, 35 years; 85 women) were analyzed. The presence of HS and subtle structural abnormalities (SSAs) in the mesial temporal lobe and in the lateral neocortical tissue was assessed in detail. Antecedents, seizure characteristics, two verbal memory tests, and outcome in HS patients with and without SSAs were determined. RESULTS: SSAs were found in 60 (43%) of the 140 HS patients, being mesial only in 32 of the 60 cases, and lateral only in nine cases; the remaining 19 cases had both mesial and lateral abnormalities. The frequency of SSA was not related to the pattern of HS or other tested variables. Prolonged febrile convulsions were present in 26 (44%) patients with SSAs, and in 26 (34%) patients (not significant) without SSAs. The outcome after surgery did not differ between patients with SSAs (incidence rate ratio for seizure recurrence, 0.9; 95% confidence interval, 0.5-1.6) compared with patients without SSAs (reference ratio, 1). CONCLUSIONS: Forty-three percent of HS patients have SSAs in their lobectomy specimens. The presence of SSAs does not predict clinical characteristics, such as presence of prolonged febrile convulsions, postsurgical outcome, or neuropsychological performance, nor does it correlate with the histologic pattern of HS. PMID- 15270761 TI - Dystonic posturing associated with putaminal hyperperfusion depicted on subtraction SPECT. AB - PURPOSE: Dystonic posturing (DP) is one of the most reliable lateralizing indicators for temporal lobe epilepsy (TLE). We evaluated the ictal hyperperfusional areas in patients with DP by using ictal-interictal subtraction single-photon emission computed tomography (SPECT). METHODS: Ninety-seven patients were treated surgically for intractable TLE, and 39 patients underwent ictal and interictal SPECT studies with the same isotope. These patients were divided into three groups: group I with DP of the contralateral side extended to the epileptogenic focus, group II with elevated muscle tonus but without DP, and group III without DP or alteration of muscle tonus. Ictal, interictal SPECT and thin-slice magnetic resonance imaging (MRI) were overlaid by using the automatic multimodality registration program to construct ictal-interictal subtracted images of SPECT on MRI. RESULTS: Thirteen patients belonged to group I; 14, to group II; and 12, to group III. A statistically significant difference in hyperperfusion rate was observed in the putamen (10 patients in group I, three in group II and two in group III; p < 0.01) and mesial temporal lobe (10 patients in group I, seven in group II, and two in group III; p < 0.05) on the ipsilateral side of the epileptogenic focus. No statistically significant difference was observed for other ictal symptoms except ipsilateral upper-limb automatism (eight patients in group I, three in group II, and none in group III; p < 0.01). CONCLUSIONS: A strong correlation between DP and hyperperfusion in the putamen and mesial temporal lobe was demonstrated. Some patients showed a wide hyperperfusion area extending from the mesial temporal lobe to putamen, which may correspond to the propagation of epileptic discharges. Our results suggest a correlation between hyperperfusion of putamen and contralateral dystonic posturing. PMID- 15270762 TI - High-resolution MRI enhances identification of lesions amenable to surgical therapy in children with intractable epilepsy. AB - PURPOSE: Many children with refractory epilepsy can achieve better seizure control with surgical therapy. An abnormality on magnetic resonance imaging (MRI), along with corroborating localization by other modalities, markedly increases chances of successful surgical outcome. We studied the impact of high resolution MRI on the surgical outcome of intractable epilepsy. METHODS: High resolution MRI using four-coil phased surface array was obtained as part of the comprehensive presurgical protocol for children with focal onset intractable seizures evaluated by our epilepsy center during the first half of 2002. RESULTS: Thirteen consecutive children, ages 5 to 18 years, entered this prospective study. For four patients with a lesion on a recent MRI examination with a standard head coil, management did not change with high-resolution MRI. Standard MRI in the other nine patients did not identify a lesion. However, high resolution MRI with the phased-array surface coil found previously undiagnosed focal abnormalities in five of nine patients. These abnormalities included hippocampal dysplasia, hippocampal atrophy, and dual pathology with frontal cortical dysplasia. In four of nine patients, no identifiable lesion was identified on the high-resolution MRI. All patients underwent invasive monitoring. In three of five patients, newly diagnosed lesions correlated with EEG abnormalities, and resection was performed. CONCLUSIONS: In our center, high resolution MRI identified lesions not detected by standard MRI in more than half the children (56%). Technical advances such as four-coil phased surface array MRI can help identify and better delineate lesions, improving the diagnosis of patients who are candidates for surgical treatment of refractory epilepsy. PMID- 15270763 TI - Memory outcome after selective amygdalohippocampectomy in patients with temporal lobe epilepsy: one-year follow-up. AB - PURPOSE: In a previous study we reported clinically significant memory declines 3 months after selective amygdalohippocampectomy (SAH) in 140 patients with mesial temporal lobe epilepsy, particularly if the resection was left-sided. We supposed that the observed postoperative impairments might have reflected acute effects of surgery. Therefore we evaluated in the present study whether a recovery can be found 1 year after surgery. METHODS: Verbal and nonverbal memory functions were assessed in 115 patients before and 3 and 12 months after unilateral SAH. RESULTS: No recovery of postoperative verbal memory declines was found in the left-SAH group. Clinically meaningful losses were still evident in 33 to 50% of patients. In right-SAH patients, a recovery of verbal memory was indicated, and effects of surgical complications were no longer evident. One year after surgery, the corresponding preoperative performance was the only significant predictor of a postoperative change in the left-SAH group. CONCLUSIONS: Verbal memory decline observed 3 months after left SAH is persistent 1 year after surgery. Declines in verbal memory, which were observed in some right-SAH patients at the short-term follow-up, seem to be temporary. PMID- 15270764 TI - Lateralization of temporal lobe epilepsy and learning disabilities, as defined by disability-related civil rights law. AB - PURPOSE: Epilepsy research has identified higher rates of learning disorders in patients with temporal lobe epilepsy (TLE). However, most studies have not adequately assessed complex functional adult learning skills, such as reading comprehension and written language. We designed this study to evaluate our predictions that higher rates of reading comprehension, written language, and calculation disabilities would be associated with left TLE versus right TLE. METHODS: Reading comprehension, written language, and calculation skills were assessed by using selected subtests from the Woodcock-Johnson Psycho-Educational Tests of Achievement-Revised in a consecutive series of 31 presurgical patients with TLE. Learning disabilities were defined by one essential criterion consistent with the Americans with Disabilities Act of 1990. Patients had left hemisphere language dominance based on Wada results, left or right TLE based on inpatient EEG monitoring, and negative magnetic resonance imaging (MRI), other than MRI correlates of mesial temporal sclerosis. RESULTS: Higher rates of reading comprehension, written language, and calculation disabilities were associated with left TLE, as compared with right TLE. Nearly 75% of patients with left TLE, whereas fewer than 10% of those with right TLE, had at least one learning disability. CONCLUSIONS: Seizure onset in the language-dominant hemisphere, as compared with the nondominant hemisphere, was associated with higher rates of specific learning disabilities and a history of poor literacy or career development or both. These results support the potential clinical benefits of using lateralization of seizure onset as a predictor of the risk of learning disabilities that, once evaluated, could be accommodated to increase the participation of patients with epilepsy in work and educational settings. PMID- 15270765 TI - Perceived impact of childhood-onset epilepsy on quality of life as an adult. AB - PURPOSE: Childhood-onset epilepsy is a common disorder. The long-term impact of having childhood epilepsy on quality of life (QOL) as an adult, whether or not seizures are in remission, has not been systematically studied. METHODS: A population-based cohort of 245 children younger than 16 years with active epilepsy between 1961 and 1964 residing in the catchment area of Turku University Hospital was followed up prospectively until 1997. Of the 99 surviving cases with uncomplicated epilepsy and 99 matched population controls, 91 subjects and controls completed questionnaires on QOL and psychosocial outcomes. RESULTS: Of the 91 subjects, 61 (67%) were in remission off medication, 13 (14%) in remission on medications, and 17 (19%) were not in remission. Subjects on medication, whether in remission or not, had worse scores on both general measures of QOL and epilepsy-specific measures than did either controls or subjects in remission off medications. They also had significantly higher rates of unemployment (p < 0.001) and lower socioeconomic status. These differences could not be accounted for by differences in education or seizure frequency. Subjects in remission off medication had rates of employment and socioeconomic status similar to those of controls. All subjects, regardless of remission status, had lower rates of marriage and of having children than did controls (p < 0.001). CONCLUSIONS: Childhood-onset epilepsy has a persistent long-term adverse impact on health related quality of life. The major impact is on those still on medications as adults, whether or not they are in remission. The impact on those in remission off medications is relatively modest. PMID- 15270766 TI - Increased prevalence of epilepsy associated with severe falciparum malaria in children. AB - PURPOSE: Multiple, prolonged, generalized, or focal seizures are common in children with severe malaria, with or without coma. In other contexts, such seizures have been associated with the development of epilepsy. The relation between falciparum malaria and epilepsy is undetermined; thus we measured the prevalence and characteristics of epilepsy in children with a history of severe malaria. METHODS: We took a detailed epilepsy history from the parents of 487 children (aged 6-9 years) to compare the prevalence of epilepsy between three exposure groups: children with a history of cerebral malaria (CM), malaria and complicated seizures (M/S), or those unexposed to either complication. Each child had an EEG and was classified as having active, inactive, or no epilepsy. RESULTS: An increased prevalence of epilepsy was seen in children previously admitted with CM [9.2%; OR, 4.4; 95% confidence interval (CI), 1.4-13.7] or M/S (11.5%; OR, 6.1; 95% CI, 2.0-18.3) compared with the unexposed group (2.2%). The most commonly reported seizure types were tonic-clonic (42%), focal becoming secondarily generalized (16%), and both (21%). Twenty-six percent of the active epilepsy group initially had EEG abnormalities. CONCLUSIONS: These results suggest that children exposed to CM or M/S have an increased propensity for epilepsy relative to children unexposed to these complications. The prevalence of epilepsy associated with CM is similar to that reported after other severe encephalopathies. The prevalence associated with M/S is more than twice that reported after complicated febrile seizures. PMID- 15270767 TI - Prescience as an aura of temporal lobe epilepsy. AB - PURPOSE: A patient with a distinct aura of prescience as a manifestation of temporal lobe epilepsy was encountered. The experience prompted a review of this ictal phenomenon among patients attending a tertiary care epilepsy outpatient clinic. METHODS: A computer epilepsy database was searched for patients with simple partial sensory seizures and complex partial seizures with auras. Identified patients had charts reviewed for details of the auras; patients were contacted and asked to provide written descriptions of their experiences. Literature searches (PubMed) were done by using the terms "precognition" or "prescience" and "seizures" or "epilepsy." Standard comprehensive epilepsy textbooks were reviewed. RESULTS: The charts of 218 patients were reviewed from 927 in the database; three had prescience as an ictal feature. The patients' descriptions were very similar in all cases (a profound sense of "knowing" what was going to happen in their environment in the immediate future). The experience was distinct from deja vu and other psychic experiences. All patients probably have temporal lobe epilepsy. Only one other description of prescience as an ictal feature was found in the literature. CONCLUSIONS: Prescience can occur as an ictal feature of temporal lobe epilepsy and represents a previously underreported psychic phenomenon. The potential lateralizing value of this symptom is yet to be determined. PMID- 15270768 TI - Todd, Faraday, and the electrical basis of epilepsy. AB - PURPOSE: To consider the origins of our understanding of the electrical basis of epilepsy in the light of the Lumleian lectures to the Royal College of Physicians in London for 1849, "On the pathology and treatment of convulsive diseases," by Robert Bentley Todd (1809-1860). METHODS: I have reviewed Todd's neglected Lumleian lectures and his observations and concepts of the electrical basis of epilepsy in relation to the influence of Michael Faraday (1791-1867), his contemporary in London, and in relation to later nineteenth century writings on the subject by Jackson, Ferrier, and Hitzig, all of whom overlooked Todd's lectures. RESULTS: Todd was a clinical scientist as well as Professor of Physiology and Morbid Anatomy, with a special interest in the nervous system, at King's College, where he came into contact with Michael Faraday, the greatest electrical scientist of all time, at the nearby Royal Institution. On the basis of his own clinical and experimental studies and his cutting-edge knowledge of neuroanatomy, neurophysiology, neuropathology, and electrical science, Todd brilliantly developed his concepts of the electrical basis of brain activity and of epilepsy in particular. With his microscope, he perceived each nerve vesicle and its related fibres (neurone in later terminology) as distinct entities for the generation of nervous polarity (force) and its transmission in the white nerve fibres throughout the nervous system by unknown molecular mechanisms. In epilepsy, an increase in electrical tension, especially in the grey matter of the hemispheres, led to periodic, sudden explosive discharges, based on Faraday's concept of disruptive discharges. CONCLUSIONS: Todd was the United Kingdom's first outstanding neurologist and neuroscientist before these disciplines existed. Influenced by Faraday, he proposed and confirmed the electrical basis of nervous discharges in epilepsy more than 20 years ahead of Jackson, Ferrier, and Hitzig, who did not refer to his priority, although Ferrier also worked at King's College, and Jackson also gave his own famous Lumleian lectures on the same subject in 1890. Todd deserves the credit for laying the foundations of our modern understanding of epilepsy. PMID- 15270769 TI - Oxygenation prevents sudden death in seizure-prone mice. AB - PURPOSE: Sudden unexpected death in epilepsy (SUDEP) is a significant health problem for some with epilepsy, but no preventive therapy has been developed for high-risk patients. Sudden death in mice, after sound-induced (audiogenic) seizures, can model the seizure-associated respiratory arrest that contributes to SUDEP in some humans and may be useful for evaluating potential SUDEP therapies. This study evaluated the effects of oxygenation on sudden fatal audiogenic seizures (AGSs) in three mouse strains, DBA/2J (D2), B6SAS, and primed C57BL/6J (B6) that differ in genetic background and seizure etiology (idiopathic or acquired). METHODS: The incidence of fatal AGSs was measured in (a) a normal air environment; (b), an oxygen-rich environment; (c) a normal air environment 24 h after a nonfatal AGS; (d) an oxygen-rich environment 24 h after a nonfatal AGS; and (e) a normal air environment 60 s after exposure to an oxygen-rich environment. RESULTS: Sudden, fatal AGS occurred in 100%, 100%, and 58% of the D2, B6SAS, and primed-B6 mice, respectively, in the air environment. Oxygenation completely prevented fatal AGSs in each strain, but had no effect on seizure incidence or severity. Furthermore, the protective effect of oxygenation against fatal seizures occurred only when the mice were in an oxygen-rich environment. CONCLUSIONS: These findings indicate that oxygenation prevents sudden death in seizure-prone mice and suggest that oxygenation may protect some seizure-prone humans at risk for SUDEP. PMID- 15270770 TI - Malignant refractory epilepsy in identical twins mosaic for a supernumerary ring chromosome 19. AB - We report identical twins with supernumerary ring chromosome 19 mosaicism, who had severe refractory epilepsy at an early age. The epilepsy was dominated largely by severe life-threatening tonic seizures. Both twins died, likely as a consequence of their severe epilepsy. They displayed no dysmorphic features. Eight cases of ring chromosome 19 have been reported in the literature, all to our knowledge without epilepsy. The clinical picture of these twins emphasizes the importance of carrying out a karyotype study on patients with early-onset epilepsy even in the absence of dysmorphic features. PMID- 15270771 TI - Treatment of refractory status epilepticus with hemispherectomy. AB - A 7-year-old boy with left hemiparesis secondary to right hemispheric cortical dysplasia was admitted to the hospital with increasing numbers of seizures. Magnetic resonance imaging showed a small dysplastic right hemisphere with abnormally thickened gyri and an apparently normal left hemisphere. Previous video-electroencephalogram (EEG) monitoring showed bilateral independent spikes and generalized slow spike-and-wave episodes on EEG and [18F]fluorodeoxyglucose (FDG) positron emission tomography scan demonstrated scattered areas of regional hypometabolism bilaterally; therefore hemispherectomy was not undertaken at that time. During this hospital stay, nonconvulsive status epilepticus developed and was refractory to multiple medical therapies including pentobarbital (PTB) coma. Burst-suppression pattern during PTB coma appeared to be generalized spike and wave, but when EEG was reviewed with increased time resolution spikes suggested a right hemisphere origin. The patient underwent bilateral intracarotid amobarbital spike-suppression test that showed only minimal suppression of epileptiform discharges with injection of the left carotid, but complete suppression of spike activity after right-sided carotid injection. A right hemispherectomy was performed with complete cessation of status epilepticus. Postoperative EEG showed no epileptiform discharges. Patient follow-up was limited to 12 months after surgery. The patient had regained the ability to walk unaided and was seizure free with a single antiepileptic medication. This case illustrates a potentially life-saving procedure for refractory status epilepticus and several techniques including a spike-suppression test to aid in prediction of cessation of seizures after hemispherectomy. PMID- 15270772 TI - Hippocampal sclerosis. PMID- 15270775 TI - AP-2 transcription factor family member expression, activity, and regulation in human epidermal keratinocytes in vitro. AB - The AP-2 transcription factor family is presumed to play an important role in the regulation of the keratinocyte squamous differentiation program; however, limited functional data are available to support this. In the present study, the activity and regulation of AP-2 were examined in differentiating human epidermal keratinocytes. We report that (1) AP-2 transcriptional activity decreases in differentiated keratinocytes but remains unchanged in differentiation-insensitive squamous cell carcinoma cell lines, (2) diminished AP-2 transcriptional activity is associated with a loss of specific DNA-bound AP-2 complexes, and (3) there is an increase in the ability of cytoplasmic extracts, derived from differentiated keratinocytes, to phosphorylate AP-2 alpha and AP-2 beta when cells differentiate. In contrast, extracts from differentiation-insensitive squamous cell carcinoma cells are unable to phosphorylate AP-2 proteins. Finally, the phosphorylation of recombinant AP-2 alpha by cytosolic extracts from differentiated keratinocytes is associated with decreased AP-2 DNA-binding activity. Combined, these data indicate that AP-2 trans-activation and DNA binding activity decrease as keratinocytes differentiate, and that this decreased activity is associated with an enhanced ability to phosphorylate AP-2 alpha and beta. PMID- 15270776 TI - Inhibition of 5-alpha-reductase activity induces stromal remodeling and smooth muscle de-differentiation in adult gerbil ventral prostate. AB - Prostatic differentiation during embryogenesis and its further homeostatic state maintenance during adult life depend on androgens. Dihydrotestosterone, which is synthesized from testosterone by 5 alpha-reductase (5 alpha-r), is the active molecule triggering androgen action within the prostate. In the present work, we examined the effects of 5 alpha-reductase inhibition by finasteride in the ventral prostate (VP) of the adult gerbil, employing histochemical and electron microscopy techniques to demonstrate the morphological and organizational changes of the organ. After 10 days of finasteride treatment at a dose of 100 mg/kg/day, the prostatic complex (VP and dorsolateral prostate) absolute weight was reduced to about 18%. The epithelial cells became short and cuboidal, with less secretory blebs and reduced acid phosphatase activity. The luminal sectional area diminished, suggestive of decreased secretory activity. The stromal/epithelial ratio increased, the stroma becoming thicker but less cellular. There was a striking accumulation of collagen fibrils, which was accompanied by an increase in deposits of amorphous granular material adjacent to the basal lamina and in the clefts between smooth muscle cells (SMC). Additionally, the periacinar smooth muscle became loosely packed. Some SMC were atrophic and showed a denser array of the cytoskeleton, whereas other SMC had a highly irregular outline with numerous spine-like projections. The present data indicate that 5 alpha-r inhibition causes epithelial and stromal changes by affecting intra-prostatic hormone levels. These alterations are probably the result of an imbalance of the homeostatic interaction between the epithelium and the underlying stroma. PMID- 15270777 TI - Differential regulation of osteogenic marker gene expression by Wnt-3a in embryonic mesenchymal multipotential progenitor cells. AB - The Wnt family of secreted glycoproteins plays an integral role in embryonic development and differentiation. To explore the role of Wnt's in one aspect of differentiation, namely osteogenesis, we employed a retroviral gene transfer approach to express Wnt-3a in the multipotent murine embryonic mesenchymal cell line C3H10T1/2. We found that expression of Wnt-3a in these cells had a significant, positive effect on cell growth in serum-containing medium, in that the cells grew to very high densities compared to the control cells. Additionally, apoptosis was markedly inhibited by Wnt-3a. However, when the cells were grown in serum-deficient medium, the Wnt-3a-expressing cells arrested efficiently in G1 phase, indicating that serum growth factors were needed in addition to Wnt-3a for enhanced proliferation. Wnt-3a-expressing cells exhibited high levels of alkaline phosphatase gene expression and enzymatic activity, but did not show any matrix mineralization. Unexpectedly, basal expression of bone sialoprotein, osteocalcin, and osteopontin were markedly inhibited by Wnt-3a, as were other known target genes of Wnt-3a, such as Brachyury, FGF-10, and Cdx1. When Wnt-3a-expressing cells were treated with osteogenic supplements in the presence of BMP-2, alkaline phosphatase gene expression and activity were further elevated. Additionally, BMP-2 was able to reverse the inhibitory effect of Wnt-3a on osteocalcin and osteopontin gene expression. These results indicate that while Wnt-3a represses basal expression of some osteogenic genes, this repression can be partially reversed by BMP-2. Finally, the enhanced gene expression of alkaline phosphatase induced by Wnt-3a could be effectively suppressed by the combined action of dexamethasone and 1,25-dihydroxyvitamin D(3). These data show for the first time that Wnt-3a has an unusual effect on multipotential embryonic cells, in that it enhances cellular proliferation and expression of alkaline phosphatase, while it represses most other marker genes of osteogenic differentiation. PMID- 15270778 TI - Establishment and in vitro differentiation of a new embryonic stem cell line from human blastocyst. AB - Embryonic stem cells have the ability to remain undifferentiated and proliferate indefinitely in vitro while maintaining the potential to differentiate into derivatives of all three embryonic germ layers. These cells have, therefore, potential for in vitro differentiation studies, gene function, and so on. The aim of this study was to produce a human embryonic stem cell line. An inner cell mass of a human blastocyst was separated and cultured on mouse embryonic fibroblasts in embryonic stem cell medium with related additives. The established line was evaluated by morphology; passaging; freezing and thawing; alkaline phosphatase; Oct-4 expression; anti-surface markers including Tra-1-60 and Tra-1-81; and karyotype and spontaneous differentiation. Differentiated cardiomyocytes and neurons were evaluated by transmission electron microscopy and immunocytochemistry. Here, we report the derivation of a new embryonic stem cell line (Royan H1) from a human blastocyst that remains undifferentiated in morphology during continuous passaging for more than 30 passages, maintains a normal XX karyotype, is viable after freezing and thawing, and expresses alkaline phosphatase, Oct-4, Tra-1-60, and Tra-1-81. These cells remain undifferentiated when grown on mouse embryonic fibroblast feeder layers in the presence or absence of recombinant human leukemia inhibitory factor. Royan H1 cells can differentiate in vitro in the absence of feeder cells and can produce embryoid bodies that can further differentiate into beating cardiomyocytes as well as neurons. These results define Royan H1 cells as a new human embryonic stem cell line. PMID- 15270779 TI - Differentiation and isolation of hepatic-like cells from human embryonic stem cells. AB - Human embryonic stem cells are pluripotent cells that can serve as a cell source for transplantation medicine, and as a tool to study human embryogenesis. We investigate here the potential of human embryonic stem cells to differentiate into hepatic cells. We have characterized the expression level of liver-enriched genes in undifferentiated and differentiated human embryonic stem cells by DNA microarrays. Our analysis revealed a subset of fetal hepatic enriched genes that are expressed in human embryonic stem cells upon differentiation into embryoid bodies. In order to isolate the hepatic-like cells, we introduced a reporter gene regulated by a hepatocyte-specific promoter into human embryonic stem cells. We isolated clones of human embryonic stem cells that express enhanced green fluorescent protein upon in vitro differentiation. Through immunostaining, we showed that most of these cells express albumin, while some cells still express the earlier expressed protein alpha-fetoprotein. Using fluorescence activated cell sorter, we were able to sort out the fluorescent differentiated cells and expand them for a few more weeks. This is the first report to demonstrate the possibility of purifying differentiated derivatives of human embryonic stem cells and culturing them further. Through confocal microscopy, we detected clusters of hepatic-like cells in 20-day-old embryoid bodies and in teratomas. As observed during embryonic development, we showed that in teratomas, the hepatic-like endodermal cells develop next to cardiac mesodermal cells. In order to examine the secreted factors involved in the induction of hepatic differentiation, human embryonic stem cells were grown in the presence of various growth factors, demonstrating the potential involvement of acidic fibroblast growth factor in the differentiation. In conclusion, given certain growth conditions and genetic manipulation, we can now differentiate and isolate hepatic-like cells from human embryonic stem cells. PMID- 15270780 TI - Early expression of thyroid hormone receptor beta and retinoid X receptor gamma in the Xenopus embryo. AB - The role of thyroid hormone in Xenopus metamorphosis is particularly well understood as it plays an essential role in that process. However, recent evidence suggests that thyroid hormone may play an earlier role in amphibian embryogenesis. We demonstrate that Xenopus thyroid hormone receptor beta (XTR beta) is expressed shortly after neural fold closure, and that its expression is localized to the developing retina. Retinoid X receptor gamma (RXR gamma), a potential dimerization partner for XTR beta, was also found to be expressed in the retina at early stages, and at later stages RXR gamma was also expressed in the liver diverticulum. Addition of either thyroid hormone or 9-cis retinoic acid, the ligands for XTR beta and RXR gamma, respectively, did not alter the expression of their receptors. However, the addition of thyroid hormone and 9-cis retinoic acid did alter rhodopsin mRNA expression. Addition of thyroid hormone generates a small expansion of the rhodopsin expression domain. When 9-cis retinoic acid or a combination of thyroid hormone and 9-cis retinoic acid was administered, there was a decrease in the expression domain of rhodopsin in the developing retina. These results provide evidence for an early role for XTR beta and RXR gamma in the developing Xenopus retina. PMID- 15270781 TI - The proper management of diabetic foot ulcers: time for a paradigm shift. PMID- 15270782 TI - C-reactive protein, its role in inflammation, Type 2 diabetes and cardiovascular disease, and the effects of insulin-sensitizing treatment with thiazolidinediones. AB - Increased concentrations of the marker of inflammation, C-reactive protein (CRP), are associated with insulin resistance, Type 2 diabetes and the development of cardiovascular disease. In particular, inflammation is closely associated with endothelial dysfunction and is recognized as one of the cardiovascular risk factors clustering in the Insulin Resistance Syndrome or Metabolic Syndrome. The exact mechanisms linking insulin resistance and inflammation remain unclear. However, the close association between insulin resistance and inflammation in atherogenesis suggests that therapies that address both parameters may have benefits in reducing diabetes-related macrovascular complications. The thiazolidinedione class of oral anti-diabetic agents are powerful insulin sensitizers that also have anti-inflammatory properties. Treatment with these agents has a range of anti-atherogenic effects, including reduced levels of CRP, plasminogen activator inhibitor-1 (PAI-1), TNF-alpha and reactive oxygen species. Additionally, the insulin-sensitizing effect of thiazolidinediones improves other factors of the Insulin Resistance Syndrome, including dyslipidaemia and hypertension. Outcome studies are underway to determine if the effects of improving insulin sensitivity and reducing inflammation will translate into clinical benefits and reduce the cardiovascular morbidity and mortality associated with insulin resistance and Type 2 diabetes. PMID- 15270783 TI - Transforming growth factor beta at clinical onset of Type 1 diabetes mellitus. A pilot study. AB - AIMS: The aims of the study were to determine whether transforming growth factor beta1 TGF-beta1 levels are raised at diagnosis of Type 1 diabetes mellitus and are related to blood glucose. SUBJECTS AND METHODS: Fourteen patients (mean age 24.3 +/- 4.9 years) admitted to hospital for onset of Type 1 diabetes were studied. On the first day of hospitalization, before insulin therapy, and at 1, 4 and 16 weeks, fasting blood glucose, HbA(1c), lipid profile and TGF-beta1 levels and TGF-beta1 levels in 24-h urine were determined. The control group included 14 non-diabetic subjects with similar characteristics to those of the diabetic group. RESULTS: Plasma and urinary TGF-beta1 levels were significantly lower in controls (4.7 (1.6-6.8) ng/ml P < 0.001; 5.7 (1.5-8.5) ng/mg urinary creatinine, P < 0.01) than in patients with Type 1 diabetes mellitus [10.5 (1.8-24.9) ng/ml; 10.1 (4.2-29.8) ng/mg urinary creatinine]. On study completion, HbA(1c) fell from 11.6 +/- 2.0 to 5.4 +/- 0.6% (P < 0.001). Improved metabolic control was not associated with changes in plasma (9.4 (2.6-19.5)/5.9 (1.6-21.5)/7.0 (2.3 30.2)/10.5 (1.8-24.9) ng/ml at baseline, 1, 4 and 16 weeks, respectively) or urinary (12.0 (4.7-29.5)/10.9 (1.5-20.5)/8.7 (4.3-16.9)/10.1 (4.2-29.8) ng/mg urinary creatinine) TGF-beta1 levels. A statistically significant correlation was observed between plasma TGF-beta1 and insulin dosage (U/kg/day) (r = 0.52, P = 0.037). CONCLUSIONS: The increased TGF-beta1 production observed herein was not modulated by glycaemic reduction and could be a response to immuno-inflammatory activation present at the onset of Type 1 diabetes. PMID- 15270784 TI - Sex-differences in prevalence of electrocardiographic left ventricular hypertrophy in Type 2 diabetes: the Casale Monferrato Study. AB - AIMS Although left ventricular hypertrophy (LVH) defined by either standard 12 lead ECG or echocardiography strongly predicts cardiovascular mortality, its prevalence in Type 2 diabetes is largely unknown. We have assessed prevalence of ECG-LVH and its relationship with clinical and metabolic variables in an Italian population-based cohort of subjects with Type 2 diabetes. METHODS The study-base was 965 (61.3%) subjects with Type 2 diabetes of the population-based cohort living in Casale Monferrato (Italy). LVH was defined by ECG Cornell voltage duration product. All measurements were centralized. RESULTS ECG-LVH was diagnosed in 165/965 subjects, giving a prevalence of 17.1% (95% CI 14.7-19.5). Large sex differences were found, with higher prevalence in women (23.5%, 19.9 27.0) than in men (8.4%, 5.6-11.0), even after adjustment for age, BMI and hypertension (OR 3.83, 95% CI 2.5-5.9). At the examination, subjects with ECG-LVH were older than those without it. Similar age- and sex-adjusted values of HbA(1c), plasma lipids, fibrinogen, uric acid and creatinine were found in the two subgroups. No differences in prevalence of hypertension, CHD, increased QT duration or dispersion, micro- and macro-albuminuria were found between subjects with ECG-LVH and those without it. In logistic regression analysis, variables independently associated with ECG-LVH, after age-adjustment, were sex and diastolic blood pressure. CONCLUSIONS: This population-based study shows: (i) a high prevalence of ECG-LVH in Type 2 diabetic subjects; (ii) 3-fold higher risk in women than in men, independently of age, BMI, and blood pressure; (iii) an independent association between ECG-LVH and diastolic blood-pressure. Screening for ECG-LVH in diabetic subjects is therefore recommended, particularly in diabetic women. PMID- 15270785 TI - Metformin, pre-eclampsia, and pregnancy outcomes in women with polycystic ovary syndrome. AB - AIMS: Was metformin during pregnancy in women with polycystic ovary syndrome (PCOS) associated with pre-eclampsia, and was it safe for mother and neonate? METHODS: In the current study, pre-eclampsia and other pregnancy outcomes were prospectively studied in 90 women with PCOS who conceived on metformin 1.5-2.55 g/day, and had > or = 1 live birth (97 pregnancies, 100 live births) compared with 252 healthy women (not known to have PCOS) with > or = 1 live birth, consecutively delivered in a community obstetrics practice. RESULTS: Women with PCOS were older than controls (33 +/- 5 vs. 29 +/- 6 years, P < 0.0001), more likely to be > 35 years old at conception (23 vs. 13%, P = 0.028), much heavier (93 +/- 23 vs. 72 +/- 18 kg, P < 0.0001, BMI 33.8 +/- 7.8 kg/m2 vs. 25.6 +/- 5.9, P < 0.0001), and more likely to be Caucasian (97 vs. 90%, P = 0.05), but there were similar numbers with preconception Type 2 diabetes mellitus [2/90 (2.2%) vs. 1/252 (0.4%), P = 0.17]. Pre-eclampsia in PCOS (5/97 pregnancies, 5.2%), did not differ (P = 0.5) from controls (9/252, 3.6%), nor did it differ (P = 1.0) in PCOS vs. control primigravidas [2/45 (4.4%) vs. 4/91 (4.4%)]. Development of gestational diabetes in PCOS did not differ from controls [9/95 pregnancies (9.5%) vs. 40/251 (15.9%), P = 0.12]. Of the 100 live births to 90 women with PCOS, there were no major birth defects. Mean +/- sd birth weight of the 80 live births > or = 37 weeks gestation in women with PCOS (3414 +/- 486 g) did not differ from controls' 206 live births > or = 37 weeks (3481 +/- 555 g), P = 0.34, nor did the percentage of > or = 37 week gestation neonates > or = 4000 g (12.5 vs. 17.5%, P = 0.3) or > or = 4500 g (1.3 vs. 2.9%, P = 0.7). CONCLUSIONS: Metformin is not associated with pre-eclampsia in pregnancy in women with PCOS, and appears to be safe for mother and fetus. PMID- 15270786 TI - Paraoxonase 1 Gln/Arg polymorphism is associated with the risk of microangiopathy in Type 2 diabetes mellitus. AB - AIMS: To investigate possible associations between diabetic microangiopathy and genetic polymorphisms in factors relevant to arterial thrombosis. METHODS: We conducted a case-control study on a total of 280 patients with Type 2 diabetes, comparing those without retinopathy or nephropathy (n = 92) and those with microangiopathies (n = 188), for the association of polymorphisms in four candidate genes, paraoxonase 1 (PON1), plasminogen activator inhibitor-1, fibrinogen, and platelet glycoprotein Ibalpha. RESULTS: There were no differences between the two study groups in gender distribution, age at diagnosis of diabetes (47.9 +/- 8.4 and 49.0 +/- 11.4 years, respectively), or duration of diabetes (14.9 +/- 4.5 and 14.5 +/- 8.4 years, respectively). Among the gene polymorphisms tested, the 192Gln/Arg polymorphism of PON1 was associated with the prevalence of retinopathy [odds ratio (OR) = 3.13, 95% confidence interval (CI) = 1.42-6.89, P = 0.0046, Gln/Gln vs. Gln/Arg and Arg/Arg]. This polymorphism was also associated with nephropathy (OR = 3.01, 95% CI = 1.30-6.98, P = 0.0103). There were no differences between the three PON1 genotypes (Gln/Gln, Gln/Arg, and Arg/Arg) with regard to the present disease status. Logistic regression analysis for the adjustment of other risk factors revealed that genotypes with PON1 192Arg were an independent predictor of retinopathy. No associations were found between microangiopathies and the other polymorphisms evaluated (plasminogen activator inhibitor-1, fibrinogen, and platelet glycoprotein Ibalpha). CONCLUSIONS: This study suggests that the presence of the 192Arg-allele in the PON1 gene is a genetic risk factor for microangiopathy in Type 2 diabetes mellitus. PMID- 15270787 TI - Mortality of South Asian patients with insulin-treated diabetes mellitus in the United Kingdom: a cohort study. AB - AIMS: To investigate mortality in South Asian patients with insulin-treated diabetes and compare it with mortality in non South Asian patients and in the general population. METHODS: A prospective cohort study was conducted of 828 South Asian and 27 962 non South Asian patients in the UK with insulin-treated diabetes diagnosed at ages under 50 years. The patients were followed for up to 28 years. Ethnicity was determined by analysis of names. Standardized mortality ratios (SMRs) were calculated, comparing mortality in the cohort with expectations from the mortality experience of the general population. RESULTS: SMRs were significantly raised in both groups of patients, particularly the South Asians, and especially in women and subjects with diabetes onset at a young age. The SMRs for South Asian patients diagnosed under age 30 years were 3.9 (95% CI 2.0-6.9) in men and 10.1 (5.6-16.6) in women, and in the corresponding non South Asians were 2.7 (2.6-2.9) and 4.0 (3.6-4.3), respectively. The SMR in women was highly significantly greater in South Asians than non South Asians. The mortality in the young-onset patients was due to several causes, while that in the patients diagnosed at ages 30-49 was largely due to cardiovascular disease, which accounted for 70% of deaths in South Asian males and 73% in females. CONCLUSIONS: South Asian patients with insulin-treated diabetes suffer an exceptionally high mortality. Clarification of the full reasons for this mortality are needed, as are measures to reduce levels of known cardiovascular disease risk factors in these patients. PMID- 15270788 TI - A 14-year prospective study of autonomic nerve function in Type 1 diabetic patients: association with nephropathy. AB - AIMS: Prospective studies of autonomic nerve function are rare. We have followed the progression of autonomic dysfunction in relation to nephropathy over 14 years in Type 1 diabetic patients. METHODS: Autonomic nerve function was assessed by heart-rate responses to deep breathing (E/I ratio) and tilting (acceleration and brake indices) and by the postural blood pressure reaction in 58 patients, 43 of whom were reassessed after 14 years. Nephropathy was evaluated by the degree of albuminuria (albuminuria > 20 micro g/min or > 0.03 g/24 h) and glomerular filtration rate ((51)Cr-EDTA plasma clearance). The acceleration index had deteriorated after 7 years (P = 0.0155), whereas the E/I ratio (P = 0.0070) and the diastolic postural blood pressure reaction (P = 0.0054) had deteriorated 14 years after the baseline examination (age-corrected values). All those with albuminuria at the third examination showed signs of autonomic neuropathy at baseline (10 of 10) compared with only nine of 22 without (P = 0.0016). Multiple regression analysis showed that the association between autonomic dysfunction and future albuminuria was due to the E/I ratio. In addition, individuals with an abnormal postural diastolic blood pressure fall (n = 7) at baseline showed a greater fall in glomerular filtration rate more than others 7-14 years later [29 (16.5) ml/min/1.72 m(2) vs. 11 (9) ml/min/1.72 m(2); P = 0.0074]. CONCLUSION: Autonomic nerve function had deteriorated after 14 years. Autonomic neuropathy and abnormal postural diastolic blood pressure falls at baseline were associated with future renal complications. PMID- 15270790 TI - Association of aldose reductase gene Z+2 polymorphism with reduced susceptibility to diabetic nephropathy in Caucasian Type 1 diabetic patients. AB - AIMS: The Z-2 allele of the (AC)n polymorphism in the aldose reductase gene (ALR2) confers increased risk of microvascular diabetic complications, whereas the Z+2 allele has been proposed to be a marker of protection. However data are conflicting. Therefore, we investigated whether this polymorphism is associated with diabetic nephropathy and retinopathy in Type 1 diabetes mellitus in a large case-control study and a family-based analysis. METHODS: A total of 431 Type 1 diabetic patients with diabetic nephropathy and 468 patients with longstanding Type 1 diabetes and persistent normoalbuminuria were genotyped for the case control study. In addition, 102 case trios and 98 control trios were genotyped for a family-based study. RESULTS: Thirteen different alleles were identified. In the case-control study, the Z+2 allele frequency was significantly higher in the normoalbuminuric diabetic than in patients with diabetic nephropathy (0.17 vs. 0.11, P = 0.008), suggesting a protective function of the Z+2 allele. No significant increase in the frequency of the putative risk allele Z-2 was found in patients with diabetic nephropathy vs. controls (0.39 vs. 0.36). No association with diabetic retinopathy was found. Although the results of the transmission of the Z-2 and Z+2 alleles in the independent family-based study were consistent with the association study, the number of informative families was limited and thus differences were not statistically significant. CONCLUSIONS: The Z+2 allele of the ALR2 promoter polymorphism is associated with a reduced susceptibility to diabetic nephropathy in Danish Type 1 diabetic patients, suggesting a minor role for the polyol pathway in the pathogenesis of diabetic kidney disease. No association of the ALR2 polymorphism with diabetic retinopathy was found. PMID- 15270789 TI - Sustained effects of pioglitazone vs. glibenclamide on insulin sensitivity, glycaemic control, and lipid profiles in patients with Type 2 diabetes. AB - AIMS: This study compared the effects of 52 weeks' treatment with pioglitazone, a thiazolidinedione that reduces insulin resistance, and glibenclamide, on insulin sensitivity, glycaemic control, and lipids in patients with Type 2 diabetes. METHODS: Patients with Type 2 diabetes were randomized to receive either pioglitazone (initially 30 mg QD, n = 91) or micronized glibenclamide (initially 1.75 mg QD, n = 109) as monotherapy. Doses were titrated (to 45 mg for pioglitazone and 10.5 mg for glibenclamide) to achieve glycaemic targets during the next 12 weeks: fasting blood glucose of < or = 7 mmol/l and 1-h postprandial blood glucose of < or = 10 mmol/l. Patients were maintained on the titrated dose for 40 weeks. RESULTS: Pioglitazone significantly increased insulin sensitivity compared with glibenclamide, as assessed by homeostasis model assessment (17.0% vs. -13.0%; P < 0.001), quantitative insulin sensitivity check index (0.011 vs. 0.007; P < 0.001) and fasting serum insulin (-1.3 pmol/l vs. 23.8 pmol/l; P = 0.007). The glibenclamide group had significantly lower HbA1c than the pioglitazone group after 12 weeks of therapy (7.8% vs. 8.3%, P = 0.015), but significantly higher HbA1c after 52 weeks of therapy (7.8% vs. 7.2%, P = 0.001). Pioglitazone significantly (vs. glibenclamide) increased mean HDL-C (P < 0.001), decreased mean triglycerides (P = 0.019), and decreased mean atherogenic index of plasma (AIP; P = 0.001) and mean total cholesterol/HDL-C (P = 0.004), without significantly elevating mean total cholesterol or mean LDL-C compared with glibenclamide. CONCLUSIONS These data suggest that the effects of pioglitazone are more sustained than those of glibenclamide for improving insulin sensitivity in patients with Type 2 diabetes, and that 52 weeks' treatment with pioglitazone has favourable effects on glycaemic control and lipoprotein profile. PMID- 15270791 TI - Targeted screening for undiagnosed diabetes reduces the number of diagnostic tests. Inter99(8). AB - AIMS: To determine the cost and performance of a Danish risk score, fasting plasma glucose (FPG), and HbA1c as single screening tests and in combination with targeted screening. SUBJECTS AND METHODS: In the Inter99 study, 12 934 inhabitants of Copenhagen County were invited to participate. All participants underwent anthropometric measurements, blood samples, and a 75-g standardized oral glucose tolerance test [N = 6784 (52.5%)]. RESULTS: Of the 6117 individuals included in the analysis, 252 (4.1%) had previously undiagnosed diabetes. As a stand-alone test, the FPG had the highest performance expressed by a significantly higher area under the receiver-operating curve [0.89; 95% confidence interval (CI) 0.86, 0.99] compared with the Danish risk score (0.78; 95% CI 0.76, 0.81) and HbA1c (0.76; 95% CI 0.72, 0.80). Targeted screening where the initial test was a risk score reduced the FPG measurements by 72% (100% vs. 27.8%). Using FPG in population-based screening, the cost per newly diagnosed diabetic individual was 583 euro compared with 270 euro if screened by questionnaire followed by FPG. The sensitivity and specificity were 78.6% and 87.7% for FPG, and 61.5% and 89.2% for the combination of the questionnaire and FPG, respectively. CONCLUSIONS: The performance of FPG was superior to a questionnaire and HbA1c used as single tests. However taking into account workload, the burden on the population and the cost per identified person with undiagnosed diabetes, targeted screening using a questionnaire followed by FPG appears to be the strategy of choice. PMID- 15270792 TI - Impact of adiposity and plasma adipocytokines on diabetic angiopathies in Japanese Type 2 diabetic subjects. AB - AIMS: Adipocytokines, products from adipose tissue, have biological activities on the vascular system, and may affect diabetic angiopathy. In this study, we assessed the relationship between adiposity and plasma adipocytokine levels, and investigated the clinical significance of adiposity and plasma adipocytokine levels on diabetic micro- and macroangiopathy in Type 2 diabetic subjects. METHODS: We studied 231 Japanese Type 2 diabetic subjects (135 men and 96 women, aged 60.4 +/- 12.3 years, body mass index 24.8 +/- 5.2 kg/m2). We measured adipocytokine [adiponectin, leptin, resistin, and tumour necrosis factor (TNF) alpha] levels, lipid profiles and urine albumin excretion. We also performed optic fundus examination and measured carotid intramedia thickness (IMT) using B mode ultrasonography, and the localization of fat with abdominal computed tomography. A group of 93 subjects with microalbuminuria or overt proteinuria was compared with the other 148 to assess the effect on nephropathy. A group of 191 eyes with simple retinopathy or more advanced changes was compared with 263 eyes without retinal changes to assess the effect on retinopathy. RESULTS: Plasma adiponectin level was negatively correlated with both visceral (r = -0.37, P < 0.01) and subcutaneous (r = -0.25, P < 0.01) fat areas. Resistin concentration was positively related with visceral fat area (r = 0.15, P < 0.05). Adiponectin concentration was positively correlated with age (r = 0.26, P < 0.01). TNF-alpha was correlated with IMT (r = 0.16, P < 0.05) after correction for age. Logistic regression analysis indicates a 4085 times greater chance of having retinopathy with a one unit increase in TNF-alpha (pmol/l) and a 30.64 times greater chance of having nephropathy with one unit increase in leptin (nmol/l). CONCLUSIONS: The present observations suggest that visceral and subcutaneous fat has an impact on microangiopathy as well as macroangiopathy, possibly through different adipocytokines. PMID- 15270794 TI - Initiation of glucose-lowering therapy in Type 2 diabetes mellitus patients in general practice. AB - AIM: The purpose of this study was to investigate which factors determine the initiation of glucose-lowering therapy in patients with Type 2 diabetes mellitus in general practice and their future glycaemic control. METHODS: All incident Type 2 diabetic patients in the general practices in a Dutch middle-sized town from 1994 to 2000 were identified. Factors associated with initiation of glucose lowering therapy were obtained from clinical files and examined by Cox's regression analyses. Using anova, the associations between clinical characteristics at diagnosis and future glycaemic control were investigated. RESULTS: In total, 603 newly diagnosed patients with Type 2 diabetes mellitus were included in the study. In the first month following diagnosis, 319 (53%) started with oral therapy. One, two and three years after diagnosis of diabetes, the cumulative incidences were 71% (95% CI 66-73%), 75% (71-79%) and 81% (77 84%), respectively. Age, gender, body weight, blood pressure, history of cardiovascular disease or total serum cholesterol values were not associated with time to start of drug therapy. An increased plasma glucose level at diagnosis was strongly related to faster initiation of drug therapy and worse future glycaemic control. Immediate initiation of glucose-lowering medication was not related to future glycaemic control. CONCLUSION: This study shows that the initial severity of diabetes, assessed by the degree of hyperglycaemia at time of diagnosis, is a major factor in determining the time to start of glucose-lowering drugs and the likelihood of achieving target levels of glycaemic control in the future, independent of glucose-lowering strategy. Therefore, patients with high glucose levels at diagnosis need close monitoring from the beginning of their disease. PMID- 15270793 TI - Effects of cardiac rehabilitation on exercise capacity in Type 2 diabetic patients with coronary artery disease. AB - AIM: To determine whether Type 2 diabetic patients with coronary disease can obtain, after cardiac rehabilitation, a similar benefit on exercise capacity to non-diabetic coronary individuals. RESEARCH DESIGN AND METHODS: Fifty-nine Type 2 diabetic patients and 36 age-matched non-diabetic patients were enrolled in a 2 month cardiac rehabilitation programme, after an acute coronary event. At the beginning and at the end of the cardiac rehabilitation programme, each subject underwent a cardiopulmonary exercise test to assess exercise capacity as measured by peak workload, duration of test, maximal heart rate, peak VO2 and anaerobic threshold. The two groups of patients were not different in age, sex ratio, type of coronary event or left ventricular ejection fraction. RESULTS: The baseline exercise capacity parameters were not different between diabetic and non-diabetic subjects. After cardiac rehabilitation, improvement of exercise capacity was significantly less in patients with diabetes compared with those without diabetes: peak workload (19% vs. 29%, P = 0.022), peak VO2 (13% vs. 30%, P = 0.002), anaerobic threshold (12% vs. 31%, P = 0.017). In the diabetic patients, a significant inverse relation between fasting blood glucose and change in peak VO2 was observed on both univariate (r = -0.40, P = 0.002) and multivariate (P = 0.001) analyses. CONCLUSIONS: The benefit of cardiac rehabilitation, after an acute ischaemic heart event, in exercise capacity is significantly lower in Type 2 diabetic patients. The response to cardiac rehabilitation in those with diabetes appears to be influenced by blood glucose levels. PMID- 15270795 TI - Baseline characteristics in the Collaborative AtoRvastatin Diabetes Study (CARDS) in patients with Type 2 diabetes. AB - AIM: To describe baseline characteristics of patients in the Collaborative AtoRvastatin Diabetes Study (CARDS), a randomized, placebo-controlled trial of lipid lowering with atorvastatin 10 mg daily for the primary prevention of major cardiovascular events in patients with Type 2 diabetes. METHODS: The main eligibility criteria were Type 2 diabetes, age 40-75 years, no previous history of coronary heart disease, stroke or other major cardiovascular events, a documented history of at least one of retinopathy, micro- or macroalbuminuria, hypertension or current smoking, LDL-cholesterol < or = 4.14 mmol/l and triglycerides < or = 6.78 mmol/l. RESULTS: Randomization of 2838 persons (909 women) into CARDS was completed in June 2001. At entry, mean age was 62 years, 12% were over 70 years old and median duration of diabetes was 6 years. Median fasting lipid levels were total cholesterol 5.4 mmol/l, LDL-cholesterol 3.1 mmol/l, HDL-cholesterol 1.4 mmol/l and triglyceride 1.7 mmol/l. There was a documented history of retinopathy in 30% of patients, micro/macroalbuminuria in 11% (additionally 17% had micro/macroalbuminuria based on two elevated pretreatment measurements of albumin-creatinine ratios), hypertension in 79% and 23% were current smokers. CONCLUSION: CARDS will contribute importantly to the evidence for the macrovascular and microvascular benefits of lipid lowering with atorvastatin in patients with Type 2 diabetes. The results are likely to have important implications for the management of patients. PMID- 15270796 TI - The metabolic syndrome and coronary heart disease in older women: findings from the British Women's Heart and Health Study. AB - AIMS: To compare two proposed definitions of the metabolic syndrome and to determine the clinical importance of the syndrome with respect to its association with coronary heart disease (CHD). METHODS: Cross-sectional study of 3770 women aged 60-79 years randomly selected from 23 British towns. RESULTS: The prevalence of the metabolic syndrome was high in this population and similar with both definitions: 28.2% (95% confidence interval 26.8, 29.7%) of the women had metabolic syndrome according to a modified version of the WHO definition, and 29.2% (27.7, 30.7%) had the ATP III-defined syndrome. There was reasonable agreement between the two definitions, with 79% of the participants being similarly classified by both definitions. The syndrome was associated with prevalent CHD, with the magnitude of the association with CHD being similar for both definitions. The odds ratio (95% confidence interval) for the age, smoking, physical activity, adult and childhood social class adjusted association of the WHO defined syndrome with prevalent CHD was 1.45 (1.19, 1.75) and for the ATP III defined syndrome was 1.53 (1.27, 1.85). Insulin resistance alone, hypertension alone and dyslipidaemia alone were all associated with CHD, with the magnitudes of these associations being similar to those for the WHO and ATP III-defined syndrome with CHD. CONCLUSIONS: The prevalence of the metabolic syndrome is high in older British women and is associated with CHD. There is reasonable agreement between a modified version of the WHO definition and the ATP III definition of the syndrome, and both are similarly associated with CHD. Single components of the syndrome are associated with CHD to a similar magnitude as the syndrome. PMID- 15270797 TI - Evaluating risk factors associated with severe hypoglycaemia in epidemiology studies-what method should we use? AB - AIMS: To determine the most appropriate regression models to use when assessing risk factors for severe hypoglycaemia and to investigate the impact of model misspecification and its clinical implications. METHODS: A total of 1229 children with Type 1 diabetes (mean age 11.7 years sd 4.1), of which 605 (49.2%) were males, were studied. Prospective assessment of severe hypoglycaemia (an event leading to loss of consciousness or seizure) was made over the 9-year period, 1992-2001. Patients were seen every 3 months and episodes of hypoglycaemia along with clinical data were recorded. Over 70% of children never experienced a severe hypoglycaemic event. Data were analysed using the Poisson regression, negative binomial, zero-inflated Poisson (ZIP) and zero-inflated negative binomial (ZINB) models. The over-dispersion and likelihood ratio statistics were calculated and the analytical methods compared. RESULTS: The Poisson regression model did not fit the data well. The negative binomial and the zero inflated Poisson and negative binomial models fitted the data better than Poisson. CONCLUSIONS: The commonly used Poisson regression models to analyse hypoglycaemia epidemiology may lead to biased parameter estimates and incorrect determination of risk factors for hypoglycaemia. We recommend the use of the negative binomial or zero inflated models to examine any risk factors associated with severe hypoglycaemia. Careful consideration must be given to the interpretation of hypoglycaemia surveys and their analysis. PMID- 15270798 TI - Clinical characteristics and outcomes of diabetic ketoacidosis in Pakistani adults with Type 2 diabetes mellitus. AB - AIMS: The aim of this study was to describe the clinical characteristics and outcomes of diabetic ketoacidosis (DKA) in Pakistani adult population with Type 2 diabetes mellitus. METHODS: We reviewed the medical records of all adult patients admitted with a diagnosis of DKA and Type 2 diabetes mellitus (DM) and followed their clinical course and outcome. Follow-up data were obtained by chart review or telephone contact where necessary. RESULTS: Fifty-seven patients fulfilled criteria for inclusion in the study. Their mean age was 48 +/- 7 years. The mean body mass index was 25.5 +/- 6.2 kg/m2. Forty-nine had a prior history of Type 2 DM but DKA was the initial presentation in 14%. Nine were on no treatment, 40 were using oral hypoglycaemic agents and eight were on insulin. A history of prior DKA was noted in eight patients. Infections were the most common precipitating factor (63%). There were 12 deaths. Follow-up after a period ranging between 12 and 43 months revealed that 30/45 patients remained on OHA without recurrence of DKA. CONCLUSION: This report highlights the need for the growing recognition of DKA occurring in adults with Type 2 DM in the South Asian population. Mortality rates are unacceptably high but the majority of survivors remain insulin independent. PMID- 15270799 TI - Amelanotic malignant melanoma disguised as a diabetic foot ulcer. AB - BACKGROUND/CASE REPORT A female patient with diet-controlled Type 2 diabetes mellitus, presented with disseminated malignancy. She had a 15-year history of a diabetic foot ulcer, which was subsequently found to be an amelanotic malignant melanoma. She had recently received immunosuppressive treatment for an episode of nephrotic syndrome secondary to focal segmental glomerulosclerosis. CONCLUSIONS: This case raises two important points. Firstly, whether non-healing diabetic foot ulcers should be biopsied, and secondly, whether the spread of the malignant melanoma was precipitated by immunosuppressive treatment. PMID- 15270800 TI - Serum amino acids in patients with mutations in the hepatocyte nuclear factor-1 alpha gene. AB - AIMS: Knockout mice lacking both copies of the hepatocyte nuclear factor 1 (HNF1) gene have altered serum levels of amino acids and generalized aminoaciduria. The aim of our study was to test whether alterations in serum amino acid levels were found in patients with mutations in the hepatocyte nuclear factor-1 alpha (HNF 1alpha) gene compared with controls. METHODS: Fasting serum from 20 patients with HNF-1alpha mutations and 20 age, sex and body mass index-matched controls was analysed for 16 amino acids. Means were compared between the two groups and Z scores calculated. RESULTS: There was no significant difference between patients with HNF-1alpha mutations and controls in serum levels of phenylalanine, arginine, citrulline or lysine as suggested by knockout mice models. Although serum levels of eight amino acids were different in the two groups, these were not significant after Bonferroni correction. CONCLUSIONS: The alterations in serum amino acid levels seen in mice models are not seen in patients with mutations in the HNF-1alpha gene. This suggests differences in mouse and man in the regulation of amino acid transport and has not provided us with a phenotypic marker to use before confirmatory genetic testing. PMID- 15270801 TI - Reduced forearm reactive hyperaemia in normoalbuminuric subjects with Type 1 diabetes and retinopathy. AB - AIM: To determine whether the forearm vasodilatory response to reactive hyperaemia (RH) is reduced in normoalbuminuric subjects with Type 1 diabetes mellitus and retinopathy compared with subjects with no retinopathy. METHODS: Forearm RH, an indicator of endothelial function, was measured, using strain gauge plethysmography, in 39 normoalbuminuric subjects (22 with retinopathy) with long-standing Type 1 diabetes mellitus. RESULTS: were evaluated in relation to conventional risk factors for atherosclerosis, and C-reactive protein (CRP), which we have recently determined to be an independent correlate of forearm RH. RESULTS: Forearm RH was decreased in subjects with retinopathy compared with those with no retinopathy (219 +/- 182 vs. 473 +/- 355, P < 0.01). Both retinopathy and CRP proved to be independent and negative predictors, and explain 27% of the variance, in forearm RH. CONCLUSION: Retinopathy in subjects with Type 1 diabetes mellitus may reflect a generalized process of endothelial dysfunction, even in the absence of microalbuminuria. PMID- 15270802 TI - Trends in mortality rates for death-certificate-coded diabetes mellitus in an English population 1979-99. AB - AIMS: Mortality statistics have customarily been coded and analysed using only one underlying cause of death. Rules for selecting the underlying cause, when more than one cause is certified on a death certificate, have changed twice in England over the past 20 years. We used data from death certificates for 1979-99 to compare mortality rates for diabetes mellitus certified anywhere on death certificates with those certified as the underlying cause. METHODS: Analysis of data from 18,917 death certificates that included diabetes mellitus in the former Oxford health region. RESULTS: Based on the underlying cause of death, mortality rates for diabetes varied substantially between the periods defined by rule changes. Based on mentions of diabetes anywhere on the death record, mortality rates were almost unchanged over time: they showed a non-significant rise of 0.1% per year (95% confidence interval -0.3, 0.6). Circulatory diseases were certified causes of death in 71% of all deaths in people with diabetes. Although mortality rates from circulatory diseases in the general population fell by 2.5% per year, rates for circulatory diseases in combination with diabetes did not fall. CONCLUSIONS: Two explanations are possible for the lack of change in mortality rates for diabetes based on all certified mentions between 1979 and 1999. Increasing prevalence and improved survival may have resulted in no net change; and/or there may have been no improvement in survival for people whose diabetes is associated with life-threatening pathology and in particular with circulatory diseases. PMID- 15270803 TI - Electrical stimulation therapy through stocking electrodes for painful diabetic neuropathy: a double blind, controlled crossover study. AB - AIMS: Peripheral neuropathy affects more than a third of diabetic patients, of whom a significant minority will have disabling symptoms. The aim of this study was to assess the efficacy of pulsed-dose electrical stimulation (through stocking electrodes) in the treatment of painful diabetic neuropathy. METHODS: Thirty patients with painful diabetic neuropathy were consecutively randomised to wear silver-plated nylon-dacron stocking electrodes for 8 h a night for 6 weeks (pulsed electric current of 50 micro amps delivered by a microcomputer). The control, identical stockings received an insignificant current (5 micro amps). Pre-treatment, weekly and end-of-treatment pain and sleep disturbance scores were recorded. RESULTS: Fourteen patients completed the study (the 16 non-completers withdrew during the first phase). Mean (+/- sd) age: 57.5 +/- 10.5 years; HbA(1c): 8.3 +/- 1.4%; median (IQR) duration of diabetes: 14.5 (7.6-19.3) years; duration of neuropathy: 4 (3-7) years. Active treatment and control produced similar reductions in pain scores [median (IQR): 40.1 (4.7-97.7)% vs. 49.2 (0.2 91.1)%, P = 0.70] and sleep disturbance scores [median (IQR): 31.1 (-4.6 to 85.4)% vs. 42.6 (-16.2 to 91.1)%, P = 0.70]. Non-completers (seven on active treatment, nine on control) withdrew for similar reasons (inconvenience, exacerbation of symptoms, dermatitis). CONCLUSIONS: There is no evidence from these results that this treatment is more effective than control in the treatment of painful diabetic neuropathy. PMID- 15270804 TI - Thiazolidinediones and congestive heart failure--exacerbation or new onset of left ventricular dysfunction? AB - BACKGROUND: Patients with diabetes mellitus have a high incidence of coronary heart disease and congestive heart failure (CHF). Thiazolidinediones (TZD) are a new class of pharmacological agents for the treatment of Type 2 diabetes mellitus, which have many beneficial cardiovascular effects. Peripheral oedema and weight gain have been reported in 4.8% of subjects on TZDs alone, with a higher incidence noted in those receiving combination insulin therapy (up to 15%), but there is limited data on the occurrence of CHF. METHODS AND RESULTS: In this paper, we report on six cases of TZD-induced fluid retention with symptoms and signs of peripheral oedema and/or CHF that occurred in subjects attending our diabetic clinic. The predominant finding in all cases was of diastolic dysfunction. All subjects were obese and hypertensive, with 5/6 having the additional risk factor of LVH, 5/6 subjects had microvascular complications, whilst 3/6 were also on insulin therapy. CONCLUSION: We suggest that obese, hypertensive diabetics may benefit from echocardiographic screening prior to commencement of TZDs, as these agents may exacerbate underlying undiagnosed left ventricular diastolic dysfunction. PMID- 15270805 TI - Nutritional factors and diabetic foot: a role for vitamin E? PMID- 15270806 TI - Genotype in human CD36 deficiency and diabetes mellitus. PMID- 15270807 TI - Post-prandial administration of insulin lispro with a high fat meal minimizes risk of hypoglycaemia in Type 1 diabetes. PMID- 15270808 TI - The statistical analysis of cardiovascular risks. PMID- 15270809 TI - Autoantibodies against GAD65 and IA-2 in recently diagnosed Type 1 diabetic children from Western India. PMID- 15270811 TI - Alcohol--the neglected risk factor in head and neck cancer. AB - Alcohol remains second only to cigarette smoking as a risk factor for head and neck cancer worldwide. The increase in incidence in head and neck cancer in a number of countries appears linked at least in part to contemporaneous rises in alcohol consumption. The relative increase in risk in women may also relate to increasing alcohol consumption. Women may be particularly sensitive to alcohol induced tumours in the oral/oropharyngeal sites. The risk is dose related, but with a non-linear increase for heavy drinkers (>100 g i.e. 12 units/day). The type of alcoholic beverage consumed seems less important. Potential mechanisms include local toxic cellular proliferation; carcinogenic action of metabolites e.g. acetaldehyde or impurities; induction of enzymes which activate procarcinogens; reduction of the protective retinoic acid; genetic polymorphism may play a part in certain geographic locations. Alcohol is also linked to stage at presentation, risk of second primary and the occurrence of comorbidity. Public awareness of the risks of alcohol remains disappointingly low. Those in identifiable high-risk groups should perhaps be targeted specifically for counselling. PMID- 15270812 TI - A systematic review of case-control studies of human papillomavirus infection in laryngeal squamous cell carcinoma. AB - A role for human papillomavirus (HPV) has been suggested in laryngeal squamous cell carcinoma (LSCC). In order to quantitate the available evidence, we reviewed studies examining the risk of laryngeal cancer-associated HPV. PubMed was searched for case-control studies conducted worldwide and published in any language since 1966. Relevant papers were hand-searched and cross-referenced. Six studies met the inclusion criteria. The studies are heterogeneous in the methods used to harvest tissue samples and techniques for detecting the virus within the tissue. HPV-16 positivity among cases ranged from 2.7% to 46.9% and 0-5.7% among controls. Two studies showed a significantly increased risk of LSCC if HPV-16 was present (OR 18.5, 95% CI 2.2-154.8, OR 2.6, 95% CI 1.1-6.0). An increased risk was also observed for glottic versus supraglottic cancer in one study (OR 9.69, 95% CI 1.47-64.04). The direction of effect is towards an increase in risk of LSCC in people with evidence of HPV-16 infection. There is marked heterogeneity in the methods used to detect the virus and frequency with which it is detected. An adequately powered study using a reliable detection technique is required to confirm and quantify this risk and to examine effect modification. PMID- 15270813 TI - Nutritional challenges in head and neck cancer. AB - In head and neck cancer patients malnutrition impacts on quality of life, complications of therapy and also prognosis, in part via altered immunity. Dysphagia assessment is extremely valuable but more work is needed to optimize the rehabilitation of the incompetent swallow in this particular patient group. Proper nutritional assessment is mandatory pre-/peri-/post-treatment. The range and palatability of nutritional supplements has greatly increased over the past few years. Many of the early problems of percutaneous gastrostomy feeding have been addressed but complication rates still remain high. As accelerated radiotherapy and chemoradiation techniques become more widely advocated, nutrition is likely to become increasingly important. The authorship includes two otolaryngologists, a nutritionist and a speech and language therapist with an interest in head and neck dysphagia, thereby aiming to provide a broad perspective of these issues. However, there appears to be a lack of prospective evaluation of many aspects of dysphagia/nutrition in head and neck cancer, which needs to be addressed. PMID- 15270814 TI - Myringoplasty using a subcutaneous soft tissue graft. AB - A retrospective study was performed on patients who underwent myringoplasty using an autologous subcutaneous soft tissue graft over a 5-year period. Details including age, site and size of perforation, grade of surgeon, surgical approach, postoperative dressings, overnight stay, complications and outcome were recorded and analysed. Fifty-two patients underwent myringoplasty using a subcutaneous soft tissue graft. Their ages ranged from 4 to 78 years (median = 36 years). The mean follow-up period was 19 months. Successful closure to give an intact tympanic membrane was obtained in 82.7% of patients. Thresholds improved on pure tone audiometry in 57.1% and deteriorated in only one patient. There was no case of dead ear as a result of surgery. Subcutaneous tissue graft has comparable outcomes with temporalis fascia graft with additional advantages of a smaller incision, minimum dissection and a lower risk of bleeding. PMID- 15270815 TI - The effect of otitis media with effusions on balance in children. AB - There is increasing interest in the effect, otitis media with effusions (OME) has on the balance in children. The aim of our investigation was to determine whether a universal effect on balance could be demonstrated in children with OME by using sway posturography. Assessment was made in 20 children with proven OME before and after the insertion of bilateral ventilation tubes. Sway posturography was performed on each occasion in each of the four recording conditions in the presence and absence of both reduced optic fixation and reduced proprioception. The pathlength traversed during the recording interval was measured and analysed by a split-unit anova. The results demonstrate an overall improvement in mean pathlength of 20% following treatment of the effusions (P < 0.001) (95% CI 14 25%). The effect of reduced optic fixation and of reduced proprioception were similar when increasing the mean pathlength by 22% (P < 0.001). These results demonstrate that OME has a universal effect on balance in all recording conditions. PMID- 15270816 TI - Emergence of ciprofloxacin-resistant pseudomonas in chronic suppurative otitis media. AB - The most frequently isolated organism in chronic suppurative otitis media (CSOM) is Pseudomonas aeruginosa. Ototopical ciprofloxacin has proven effectiveness against P. aeruginosa. The purpose of this study is to evaluate the patients with recurrent otorrhoea caused by CSOM that was unresponsive to topical ciprofloxacin. Eighty-eight patients (18-77 years of age) with otorrhoea due to CSOM were reviewed retrospectively. All of them were initially treated with ciprofloxacin eardrops but the otorrhoea failed to resolve. Bacteriological specimens were processed and identified with standard cultures. In vitro antimicrobial susceptibility of these bacterial isolates was assessed by an agar disc diffusion method. Isolates were tested against 16 antibiotics. Ciprofloxacin resistant P. aeruginosa were isolated in all cases. Imipenem was the most sensitive antibiotic agent with an overall susceptibility rate of 96.5%, followed by amikacin (55.6%), piperacillin/tazobactam (37.5%) and ceftazidime (31.8%). In our series, ciprofloxacin-resistant P. aeruginosa is increasing recently. Continuous surveillance is necessary to monitor antimicrobial resistance and to guide antibacterial therapy. PMID- 15270817 TI - Psycho-socio-economic outcomes in acoustic neuroma patients and their carers related to tumour size. AB - The objective was to explore psycho-socio-economic outcomes of a 2-year cohort of patients having surgery for an acoustic neuroma, and carers and their relationship to tumour size after surgery. The Wessex Patient Carer Questionnaire was designed in conjunction with Patients and Carers, to determine psycho-socio economic outcomes. The results were juxtaposed against clinical profiles. The House-Brackman (HB) scale was used to assess facial function at 6 and 12 months after operation. The cohort contained 102 patients. There were 87% effective responders. Half were aged below 54 years and 30% had school-aged children. The majority (93%) of patients were operated via the translabyrinthine approach. Patients with large tumours, i.e. greater than 3 cm (28%), had most post treatment physical problems, including hearing and balance difficulties, and 42% reported difficulty eating in public. Thirty-four per cent felt 'stressed' and 18%'depressed'. After 6 months, facial function was recorded as HB scale 5/6 in 21% of patients but by 1 year only 8% of patients were HB 5/6. Patients and carers were generally very satisfied with their in-patient neurosurgical care, but significantly dissatisfied with post-discharge care - particularly the shortcoming of the community services. The majority of families felt 'unsupported' and only 20% of patients had confidence in their General Practitioner's knowledge. Families faced severe socio-economic disruption and patients"time-off-work' was estimated to cost pound 954,000. Carers carried considerable post-discharge psychological burdens and costs to the public purse were calculated to be pound 52,000. PMID- 15270818 TI - Trends and outcomes in the Manchester adult cochlear implant series. AB - The adult cochlear implant programme in Manchester was established in 1988, initially using funding obtained from the HEAR (Help Ear and Allied Research: charity number: 519784) charity before government resources became available in the mid-1990s. Manchester was the first centre in the UK to implant multichannel devices on a regular basis. To date, over 250 adults have been implanted, including nine bilateral and eight deaf-blind patients. All the patients have a postlingual onset of severe-profound hearing loss; 73% (n = 175) of the implants performed used a Nucleus multichannel implant and 24% (n = 58) used a Medel multichannel implant. In addition, the team has implanted three Medel single channel devices, two Ineraid devices and one Clarion High Focus II device. This study is a retrospective analysis of the trends and outcomes in implant fitting during the first 14 years (1988-2002) of the programme. The paper describes the patient demographics and audiological complications for 240 implantations performed on 214 patients. Speech perception outcomes are reported for a subset of the patients. The average score for the Bench, Kowal, Bamford sentence test at the post-18-month stage of implant use is 66% and for Arthur Boothroyd words 53%. Trends in the series are analysed with respect to the change in criteria for adult implantation, the move towards bilateral implantation and the rate of uptake of cochlear implants by different ethnic groups. PMID- 15270819 TI - A computer program to calculate the size of tympanic membrane perforations. AB - Chronic suppurative otitis media is characterized by the presence of a persistent perforation of the tympanic membrane. Accurate estimation of the perforation size is helpful in clinical management and research. In this study, a computer program was developed to calculate the percentage of a perforation relative to the whole tympanic membrane including the pars tensa and pars flaccida. In order to demonstrate the variability of estimations of perforation size by different surgeons, we calculated the percentage of perforation for four tympanic membranes, and compared the results with those estimated by five experienced otologists and 10 senior residents. The results show that the estimations from both otologists and residents departed from the values calculated by the computer program quite substantially. Beside, the variance of the estimation is large. However, the variances of computer calculation are quite small, which means that the results obtain from different users are quite consistent. Therefore, we concluded that this program is necessary and useful to evaluate the size of the perforation as the differences in visual estimations can be very big and the variances can be large for different individuals, even by experienced otologists. PMID- 15270820 TI - Nasal physiological changes during pregnancy. AB - Rhinitis in pregnancy has been previously investigated with variable results. This study examines all the variables of the nasal airway simultaneously for the first time. Eighteen women were recruited in the first trimester of pregnancy and followed through to the postpartum period to monitor the changes that occurred. Measurements of the nasal airway included anterior rhinoscopy (AnR), peak inspiratory nasal flow, acoustic rhinometry, anterior rhinomanometry (ARM), and the saccharin test with rhinitis questionnaire scores providing a symptomatic measurement. All the tests showed a trend consistent with decreasing nasal patency when expressed as an average for the group as a whole, although only AnR, ARM, mucociliary clearance time and rhinitis questionnaire scores were statistically significant (P < or = 0.05). This confirms the effect of pregnancy on the nasal mucosa and coincides with the rise in the serum concentration of the female sex hormones with gestational age, returning to normal postpartum. Pharmacological antagonism of oestrogens may therefore relieve nasal congestion and is currently under further research. PMID- 15270821 TI - Investigation and endoscopic treatment for functional and anatomical obstruction of the nasolacrimal duct system. AB - The purpose of this study was to assess the results of endonasal DCR in patients with a functional obstruction of the nasolacrimal system. The study design was a prospective non-randomized interventional case series. A prospective series of 102 consecutive endoscopic DCRs were evaluated with preoperative dacrycytography (DCG) and lacrimal scintillography. Evaluation of the DCG and scintillography allowed the patients to be classified as either having an anatomical or functional obstruction of the nasolacrimal system. The powered endoscopic DCR was performed between January 1999 and December 2001. Patients were followed up for a minimum of 12 months after surgery. In the 70 powered endoscopic DCRs for anatomical obstruction, 68 (97%) have remained asymptomatic with a free flow of fluorescein from the conjunctiva to the nose. In the patients who were classified as having a functional obstruction, 27 of 32 were asymptomatic (84%) with a free flow of fluorescein into the nose. To conclude, powered endoscopic DCR produces excellent results in patients with anatomical obstruction and good results in patients with functional obstruction of the nasolacrimal system. PMID- 15270822 TI - Immediate effect of benzalkonium chloride in decongestant nasal spray on the human nasal mucosal temperature. AB - Benzalkonium chloride is a preservative commonly used in nasal decongestant sprays. It has been suggested that benzalkonium chloride may be harmful to the nasal mucosa. Decongestion with the vasoconstrictor xylometazoline containing benzalkonium chloride has been shown to cause a significant reduction of the nasal mucosal temperature. The purpose of the present study was to determine the short-term influence of xylometazoline nasal spray with and without benzalkonium chloride on the nasal mucosal temperature. Healthy volunteers (30) were included in the study. Fifteen volunteers received xylometazoline nasal spray (1.0 mg/mL) containing benzalkonium chloride (0.1 mg/mL) and 15 age-matched subjects, received xylometazoline nasal spray without benzalkonium chloride. Using a miniaturized thermocouple the septal mucosal temperature was continuously measured at defined intranasal detection sites before and after application of the nasal spray. The mucosal temperature values did not significantly differ between the group receiving xylometazoline containing benzalkonium chloride and the group receiving xylometazoline spray without benzalkonium chloride before and after decongestion (P > 0.05). In both study groups septal mucosal temperatures significantly decreased after decongestion (P < 0.05) because of a reduction of the nasal mucosal blood flow following vasoconstriction. This study indicates that benzalkonium chloride itself does not seem to influence nasal blood flow and nasal mucosal temperature in topical nasal decongestants. PMID- 15270823 TI - Consent for tonsillectomy. AB - Rulings in recent negligence cases reveal a shift towards what the 'reasonable patient' would expect in deciding the risks doctors must disclose to patients. This survey aimed to investigate whether the 'reasonable patient' and 'responsible body of medical opinion' agree about which risks should be discussed regarding tonsillectomy. Using questionnaires, surgeons were asked which of the 10 complications they routinely discussed and patients were asked how seriously they regarded these complications. The results were compared with the Test of Proportions. Most surgeons routinely mentioned otalgia, odynophagia, throat infection and re-operation. Most patients regarded potentially fatal bleeding, pneumonia and blood transfusion as very serious but only the minority of surgeons mentioned these (P < 0.001). When obtaining consent for tonsillectomy, surgeons do not routinely mention all the risks that the 'reasonable patient' would expect. The 'reasonable patient' would expect that re-operation, transfusion, pneumonia and fatal blood loss are discussed. PMID- 15270824 TI - Tonsillar size is an important indicator of recurrent acute tonsillitis. AB - This prospective study was designed to identify important clinical features in patients with recurrent acute tonsillitis. A total of 195 consecutive children aged from 1 to 16 years were examined and a history of recurrent acute tonsillitis recorded. Patients with obstructive sleep apnoea or recent acute tonsillitis were excluded. Tonsil size was measured on the Brodsky scale [Brodsky L. (1989) Paediatr Clin N Am 36, 1551], tonsil symmetry, cervical lymphadenopathy, and hyperaemia of the anterior pillars was recorded. Patients with a history of recurrent tonsillitis had larger tonsils than those without tonsillitis (P < 0.001). Tonsil asymmetry and cervical lymphadenopathy were more common in patients with recurrent tonsillitis (P < 0.001). Anterior pillar hyperaemia was also more frequent in recurrent tonsillitis (P < 0.01). In addition to the frequency and severity of tonsillitis it is suggested that the size and symmetry of the tonsils, plus cervical lymphadenopathy and anterior pillar hyperaemia should be taken into account when deciding which patients would benefit from tonsillectomy. PMID- 15270825 TI - Neoplastic invasion of laryngeal cartilage: the significance of cartilage sclerosis on computed tomography images. AB - Cartilage sclerosis has been cited as a sensitive and a specific sign of neoplastic cartilage invasion, on cross-sectional computed tomography (CT) images of the larynx. We retrospectively reviewed 36 consecutive patients, who underwent a total laryngectomy for squamous cell carcinoma of the larynx. Preoperative CT images were compared with formal histological sections of the larynx in order to assess cartilage invasion by tumour. Isolated asymmetrical cartilage sclerosis was found to have a sensitivity of 62% and a specificity of 42% for predicting neoplastic cartilage invasion when compared with histopathological sections of the tumour. In this study we found that cartilage sclerosis was not a useful early radiological sign of neoplastic cartilage invasion when taken in isolation. PMID- 15270826 TI - A UK multi-centre pilot study of speech and swallowing outcomes following head and neck cancer. AB - Speech and swallowing are important components of health-related quality of life following head and neck cancer treatment. The aim of this study was to demonstrate the value of prospective multi-centre evaluation by Speech and Language Therapists and to compare health-related quality of life with speech and swallowing impairments. The University of Washington Head and Neck questionnaire version 4 (UW-QOL) and Therapy Outcome Measures (TOM) were rated before and 6 months after cancer treatment in 95 patients from 12 centres. There was deterioration in TOM scores at 6 months. Pretreatment UW-QOL swallowing was ranked equal first, with speech fourth. At 6 months speech was first and swallowing second. There were positive correlations between UW-QOL swallowing and TOM dysphagia and between UW-QOL speech and TOM laryngectomy, voice, phonology and dysarthria disorders. Both outcome measures are suitable for routine practice. Adaptation of TOM scales for use with head and neck cancer patients may improve sensitivity, validity and therapist compliance. PMID- 15270827 TI - Evaluation of aural manifestations in temporo-mandibular joint dysfunction. AB - Thirty patients with temporo-mandibular joint dysfunction were selected to investigate the changes in otoacoustic emissions before and after conservative treatment of their temporo-mandibular joints. Pure tone audiometry, transient evoked otoacoustic emissions (TEOAE), distortion-product otoacoustic emissions (DPOAE) as well as a tinnitus questionnaire were administered to all patients before and after therapy. Therapy was conservative in the form of counselling, physiotherapy, anti-inflammatory agents, muscle relaxants, and occlusal splints. Results indicated insignificant changes in the TEOAEs, whereas there were significant increases in distortion product levels at most of the frequency bands. These results were paralleled to subjective improvement of tinnitus. PMID- 15270828 TI - Frontal ostium neo-osteogenesis and restenosis after modified endoscopic Lothrop procedure in an animal model. AB - All patients who undergo a modified endoscopic Lothrop procedure have postoperative narrowing of the enlarged frontal ostium. The aim of this study is to evaluate neo-osteogenesis and restenosis of the frontal ostium and its effect on mucociliary clearance. Fourteen sheep underwent an endoscopic modified Lothrop procedure. Pre- and postoperative nuclear medicine gamma scintigraphy of the frontal sinuses was performed. The sizes of the frontal ostia were measured and biopsies taken from the bone of the frontal ostium. Histological evidence of new bone formation was found in 56% of biopsies. The average preoperative mucociliary clearance half times (T1/2) at 15 and 30 min were 70 and 74 min, respectively, and postoperatively were 50 and 67 min. There was a non-significant trend towards poorer clearance in sinuses with neo-osteogenesis. The average size of the frontal ostium decreased by 28%. There was no relationship between the size of the ostium and neo-osteogenesis. Neo-osteogenesis was seen in 56% of biopsies with a 28% reduction in size of the frontal ostium after 224 days. Mucociliary clearance did not alter significantly. PMID- 15270837 TI - Autoimmune polyglandular syndrome Type 2: the tip of an iceberg? AB - Autoimmune polyglandular syndromes (APS) are conditions characterized by the association of two or more organ-specific disorders. Type 2 APS is defined by the occurrence of Addison's disease with thyroid autoimmune disease and/or Type 1 diabetes mellitus. Clinically overt disorders are considered only the tip of the autoimmune iceberg, since latent forms are much more frequent. Historical, clinical, genetic, and immunological aspects of Type 2 APS are reviewed. Furthermore, data on 146 personal cases of Type 2 APS are also reported. PMID- 15270838 TI - Interference with immune function by HTLV-1. PMID- 15270839 TI - The ratio of n-6 to n-3 fatty acids in maternal diet influences the induction of neonatal immunological tolerance to ovalbumin. AB - Prevalence of allergy is increasing in many countries and might be related to changed environmental factors, such as dietary fatty acids (FA). The present study investigates whether dietary ratio of n-6 to n-3 FA influences the induction of immunological tolerance to ovalbumin (OA) in neonatal rats. During late gestation and throughout lactation Sprague-Dawley rats were fed a diet containing 7% linseed oil (n-3 diet), sunflower oil (n-6 diet) or soybean oil (n 6/n-3 diet). At 10-16 days of age the rat offspring were subsequently exposed, or not, to OA via the milk. The offspring were weaned onto the same diets as the mothers and immunized with OA and the bystander antigen human serum albumin (HSA). In the offspring on the n-3 diet exposure to OA via the milk resulted in lower delayed type hypersensitivity reaction (DTH) and antibody responses against both OA and HSA, compared to those in the offspring not exposed to OA, indicating the induction of oral tolerance. In the offspring on the n-6 diet, the exposure to OA led to depressed specific immune responses against only OA, not HSA. In the offspring on the n-6/n-3 diet oral exposure to OA did not influence immune responses against OA, or HSA. The results indicate that the dietary ratio of n 6/n-3 FA is important for the induction of neonatal oral tolerance. Thus nonoptimal feeding may have effects on the development of immunological tolerance to dietary antigen ingested by the mother. The ratio of n-6/n-3 FA in the diet may be considered in the context of increased prevalence of allergy. PMID- 15270840 TI - Reduction of human anti-tetanus toxoid antibody in hu-PBL-SCID mice by immunodominant peptides of tetanus toxoid. AB - Immunotherapy of murine autoimmune and allergic diseases by administration of peptides corresponding to the dominant T cell epitope is a reality. However, problems remain in applying this therapy to reduce antibody responses in humans. To overcome these difficulties, a preclinical system was developed to test the effect of immunodominant peptides from a common antigen, tetanus toxoid (TT), on the long-term human anti-TT response. Individuals whose T cells proliferated against dominant TT peptides were identified. Peripheral blood leucocytes (PBL) from these donors were injected intraperitoneally (i.p.) into mice with severe combined immunodeficiency (SCID) that had been depleted of murine natural killer (NK) cells (hu-PBL-SCID mice). Peptides or PBS were injected i.p. before a further injection of PBL and immunization with TT. The concentration of human IgG and anti-TT in murine plasma was followed for 10 weeks. The total IgG was similar in both groups. By contrast, there was a statistically significant reduction in IgG anti-TT from eight weeks onwards. It is considered that the hu-PBL-SCID model system may provide a means by which the efficacy of peptide immunotherapy for reduction of pathological antibodies in humans can be examined. PMID- 15270841 TI - DNA vaccination encoding glutamic acid decarboxylase can enhance insulitis and diabetes in correlation with a specific Th2/3 CD4 T cell response in non-obese diabetic mice. AB - DNA vaccination encoding beta cell autoantigens has been shown very recently to prevent type I diabetes in non-obese diabetic (NOD) mice. However, DNA vaccination encoding microbial or reporter antigens is known to induce specific long-lasting CD4 Th1 and strong cytolytic CD8 T cell responses. As this immune phenotype is associated strongly with beta cell destruction leading to diabetes, we have chosen to study the effects of plasmids encoding glutamic acid decarboxylase (GAD), a crucial beta cell autoantigen, in female NOD mice that developed a 'moderate' diabetes incidence. In the present study, 3-week-old female NOD mice were vaccinated twice in tibialis muscles with plasmid-DNA encoding 65-kDa GAD or betagalactosidase. In GAD-DNA immunized mice, diabetes cumulative incidence (P < 3.10(-3)) and insulitis (P < 7.10(-3)) increased significantly. Simultaneously, DNA immunization induced GAD-specific CD4 T cells secreting interleukin (IL)-4 (P < 0.05) and transforming growth factor (TGF)-beta (P = 0.03). These cells were detected in spleen and in pancreatic lymph nodes. Furthermore, vaccination produced high amounts of Th2 cytokine-related IgG1 (P < 3.10(-3)) and TGF-beta-related IgG2b to GAD (P = 0.015). Surprisingly, diabetes onset was correlated positively with Th2-related GAD-specific IgG1 (P < 10(-4)) and TGF-beta-related IgG2b (P < 3.10(-3)). Moreover, pancreatic lesions resembled Th2-related allergic inflammation. These results indicate, for the first time, that GAD-DNA vaccination could increase insulitis and diabetes in NOD mice. In addition, our study suggests that Th2/3 cells may have potentiated beta cell injury. PMID- 15270842 TI - Beneficial effects of troglitazone on neutrophil dysfunction in multiple low-dose streptozotocin-induced diabetic mice. AB - Patients with poorly controlled diabetes are at high risk of acquiring bacterial infections. However, conflicting results have been reported on neutrophil function in diabetes. We periodically evaluated neutrophil dysfunction in multiple low-dose streptozotocin (STZ)-induced diabetic mice, and then evaluated the effects of troglitazone and other thiazolidinediones (TZDs) on the decline of neutrophil function. Zymosan was injected intraperitoneally and neutrophil infiltration and phagocytosis were evaluated. While phagocytosis of zymosan by peritoneal neutrophils was consistently reduced in diabetic mice, neutrophil infiltration was decreased on day 30, but increased on day 40 after STZ injection. The in vitro chemotactic and phagocytic activities of blood neutrophils in mice that did not receive zymosan were consistently reduced in diabetic mice. Phorbol myristate acetate (PMA)-stimulated superoxide production by zymosan-induced peritoneal neutrophils and the levels of zymosan-induced tumour necrosis factor (TNF)-alpha and interleukin (IL)-1beta in peritoneal exudate fluids were also reduced in the diabetic mice. Treatment of the diabetic mice with troglitazone beginning 2 weeks after STZ injection did not improve hyperglycaemia but did prevent the decline of zymosan-induced neutrophil infiltration on day 30, and additionally promoted the increased infiltration on day 40. Troglitazone also promoted the chemotactic activity of blood neutrophils isolated from normal mice in vitro. Rosiglitazone but not pioglitazone induced a similar effect. Neutrophil phagocytosis was not enhanced by troglitazone either in vivo or in vitro. Taken together, neutrophil function is impaired by STZ induced diabetes, but inflammatory infiltration does not always vary with the chemotactic disability or cytokine levels. Furthermore, troglitazone and rosiglitazone were suggested to improve at least neutrophil chemotactic activity in these animals. PMID- 15270843 TI - Anti-inflammatory effect of Viscum album agglutinin-I (VAA-I): induction of apoptosis in activated neutrophils and inhibition of lipopolysaccharide-induced neutrophilic inflammation in vivo. AB - Viscum album agglutinin-I (VAA-I) is a plant lectin which possesses antitumoral properties. This lectin is also known for its immunostimulatory effects when used at low concentrations (1-100 ng/ml). We have demonstrated recently that VAA-I is a potent inducer of human neutrophil apoptosis in vitro when used at higher concentrations. The role of VAA-I on activated neutrophils has not so far been investigated and its potential proinflammatory properties in vivo are poorly documented. Herein, we demonstrated that VAA-I (1000 ng/ml) induces apoptosis in lipopolysaccharide (LPS)-treated human neutrophils in vitro as well as in murine neutrophils isolated from lipopolysaccharide (LPS)-induced neutrophil influx. Using this model, we found that administration of VAA-I (100 or 1000 ng/ml) did not induce an inflammatory response. However, when used at 1 or 10 ng/ml, VAA-I was found to significantly induce a transitory inflammatory response, based on an increased leucocyte infiltration (>98% neutrophils). Also, we found that VAA-I inhibits LPS-induced neutrophil influx when administered simultaneously with LPS. In such conditions, some characteristic apoptotic neutrophils were observed in the pouch. Unlike LPS, which increased the production of some cytokines, VAA-I (1 or 10 ng/ml) did not increase the production of tumour necrosis factor (TNF) alpha, interleukin (IL)-1Ra, IL-1alpha, IL-beta, IL-8, IL-10 or IL-12 (p70) in human neutrophils. We conclude that VAA-I possesses the ability to induce apoptosis of preactivated neutrophils at a concentration that does not induce a proinflammatory response. Moreover, we conclude that VAA-I can inhibit a LPS induced proinflammatory response in vivo. These data may provide new clinical perspectives in future mistletoe therapy and favour its potential utilization based on anti-inflammatory activity that at first appears contradictory with its use as immunostimulant. PMID- 15270844 TI - T cell-derived tumour necrosis factor is essential, but not sufficient, for protection against Mycobacterium tuberculosis infection. AB - Tumour necrosis factor (TNF) is critical for sustained protective immunity against Mycobacterium tuberculosis infection. To investigate the relative contributions of macrophage- and T cell-derived TNF towards this immunity T cells from wild-type (WT) or TNF-/- mice were transferred into RAG-/- or TNF-/- mice which were then infected with M. tuberculosis. Infected RAG-/- mice and RAG-/- recipients of TNF deficient T cells developed overwhelming infection, with extensive pulmonary and hepatic necrosis and succumbed with a median of only 16 days infection. By contrast, RAG-/- recipients of WT T cells showed a significant increase in survival with a median of 32 days. Although initial bacterial growth was similar in all groups of RAG-/- mice, the transfer of WT, but not TNF-/-, T cells led to the formation of discrete foci of leucocytes and macrophages and delayed the development of necrotizing pathology. To determine requirements for macrophage-derived TNF, WT or TNF-/- T cells were transferred into TNF-/- mice at the time of M. tuberculosis infection. Transfer of WT T cells significantly prolonged survival and reduced the early tissue necrosis evident in the TNF-/- mice, however, these mice eventually succumbed indicating that T cell-derived TNF alone is insufficient to control the infection. Therefore, both T cell- and macrophage-derived TNF play distinct roles in orchestrating the protective inflammatory response and enhancing survival during M. tuberculosis infection. PMID- 15270845 TI - Mycoplasma infection induces a scleroderma-like centrosome autoantibody response in mice. AB - Development of autoantibodies to intracellular molecules is a universal feature of autoimmune diseases and parallels onset of chronic inflammatory pathology. Initiating antigens of disease-specific autoantibody responses are unknown. We previously showed that the major targets of autoantibodies in scleroderma are centrosomes, organelles involved in mitotic spindle organization. Here we show that centrosome autoantibodies are induced in mice by mycoplasma infection. The centrosome-specific antibody response involves class switching of preexisting IgM to IgG isotypes, suggesting a T cell-dependent mechanism. The antibody response spreads to include additional intracellular targets, with newly recruited autoantibody specificities arising as IgM isotypes. Antibiotic treatment of mice prevents autoantibody development. Centrosome autoantibodies may provide an aetiological link between infection and human autoimmunity and suggest novel therapeutic strategies in these disorders. PMID- 15270846 TI - Multiple antigenic peptides facilitate generation of anti-prion antibodies. AB - Recent reports have demonstrated the ability of anti-prion antibodies to inhibit PrPSc propagation. Due to the relatively poor immunogenic properties of both PrPC and PrPSc, the generation of anti-prion antibodies still causes a significant problem in the development of immunotherapeutic strategies. This study examines the potential of multiple antigenic peptides (MAPs) to raise an antibody response to prion derived sequences in mice. The MAP was constructed of a four spiked ring. Two spikes containing human or mouse derived prion amino acid sequences and two spikes containing the universally promiscuous tetanus toxoid sequence (aa 830 844) which was used to assist T-cell-dependent B-cell antibody production. Following vaccinations with the MAP or MAP plus adjuvant, sera were taken and antibody titres assessed. The MAP containing only the mouse sequence failed to elicit a significant antibody response. MAPs containing human prion sequences elicited antibody production to the corresponding prion sequence. Further analysis also demonstrated that these peptides were able to generate antibody responses that recognize conserved human and mouse sequences. These homologous sequences contain the heralded PrPSc specific sequence 'Tyr-Tyr-Arg' and therefore these MAPs may have some therapeutic potential. PMID- 15270847 TI - Immunomodulation by roquinimex decreases the expression of IL-23 (p19) mRNA in the brains of herpes simplex virus type 1 infected BALB/c mice. AB - Herpes simplex virus (HSV) is a common neurotropic virus which infects epithelial cells and subsequently the trigeminal ganglia (TG) and brain tissue. We studied how immunomodulation with roquinimex (Linomide) affects the course of corneal HSV infection in BALB/c mice. BALB/c mice have also been used in a model for HSV based vectors in treating an autoimmune disease of the central nervous system (CNS). We addressed the questions of how immunomodulation affects the local as well as the systemic immune response and whether roquinimex could facilitate the spread of HSV to the CNS. The cytokine response in the brain and TG was studied using a quantitative rapid real-time RT-PCR method. We were interested in whether immunomodulation affects the expression of the recently described Th1-cytokine IL 23p19 in the brain and TG. The expression of IL-23 mRNA was decreased in brains of roquinimex-treated BALB/c mice. Also the expression of IL-12p35 and IFN-gamma mRNAs decreased. No significant changes were seen in IL-4 and IL-10 mRNA expression. The cytokine response was also studied using supernatants of stimulated splenocytes by EIA. Roquinimex treatment suppressed the production of IFN-gamma and also the production of IL-10 in HSV-infected BALB/c mice. PMID- 15270848 TI - Prevention of experimental autoimmune encephalomyelitis in DA rats by grafting primary skin fibroblasts engineered to express transforming growth factor-beta1. AB - To determine whether primary fibroblasts producing latent transforming growth factor beta1 (TGF-beta1) are capable of down-regulating experimental autoimmune encephalomyelitis (EAE), a retroviral vector TGF-beta1-pBabe-neo (-5'UTR) was used for efficient gene transfer into primary skin fibroblasts of DA rats. After heat activation, conditioned medium from the transduced fibroblasts was found to inhibit significantly in vitro proliferation of lymphocytes from lymph nodes of DA rats with EAE. Intraperitoneal administration of TGF-beta1-transduced fibroblasts into DA rats during the priming phase of EAE resulted in a significant reduction in mortality and in the mean clinical and EAE scores versus the control immunized animals treated with non-transduced fibroblasts. PMID- 15270849 TI - Effect of prolactin on carcinoembryonic antigen-specific cytotoxic T lymphocyte response induced by dendritic cells. AB - The cytokine hormone prolactin (PRL) has been shown previously to modulate native cellular responses and maturation of antigen-presenting cells. Here we have addressed its effect on the antigen-specific response of cytotoxic T lymphocytes (CTL). CTL were generated from HLA-A2 lymphocytes after three rounds of stimulation with autologous dendritic cells loaded with HLA-A2-restricted carcinoembrionic antigen (CEA) Cap-1 (YLSGANLNL) peptide. Selected cultures were expanded on cytokine-supplemented feeder-layers, enriched for CD8+ lymphocytes and analysed for PRL-receptor (PRL-R) expression and PRL responsiveness. Resting CD8+ lymphocytes were negative for PRL-R, whereas antigen-activated CD8+ lymphocytes derived from long-term cultures were highly positive. Results of a 51Cr release assay showed CTL killing of CEA-loaded, but not unloaded, T2 cell line and the CEA-positive gastric carcinoma cell line KATO, but not of the CEA negative T leukaemia cell line Jurkat. Interferon (IFN)-gamma release, evaluated in an ELISPOT assay against CEA-loaded T2, was enhanced (P < 0.05) by concentrations of PRL (12-25 ng/ml) very close to the physiological levels (6-20 ng/ml), but was decreased (P < 0.05) by high concentrations (200 ng/ml). Pre incubation of the stimulators with the anti-MHC class I MoAb W6.32 induced a 40 60% decrease of the PRL-boosted IFN-gamma release, thus proving the MHC restriction of the lymphocyte response. Cytotoxicity against CEA-loaded T2 and KATO cell lines was also increased by 12-25 ng (P < 0.05) and decreased (P < 0.05) by 200 ng PRL. Pre-incubation of CTL with an antibody specific for the PRL R almost completely abrogated this effect. PMID- 15270850 TI - Disruption of MAP kinase activation and nuclear factor binding to the IL-12 p40 promoter in HIV-infected myeloid cells. AB - Progressive immunodeficiency in HIV infection is paralleled by a decrease in IL 12 production, a cytokine crucial for cellular immune function. Here we examine the molecular mechanisms by which HIV infection suppresses IL-12 p40 expression. HIV infection of THP-1 myeloid cells resulted in decreased LPS-induced nuclear factor binding to the NF-kappaB, AP-1, and Sp1 sites of the IL-12 p40 promoter. By site-directed mutagenesis we determined that each of these sites was necessary for transcriptional activation of the IL-12 p40 promoter. Binding of NF-kappaB p50, c-Rel, p65, Sp1, Sp3, c-Fos, and c-Jun proteins to their cognate nuclear factor binding sites was somewhat impaired by HV infection, although a role for other as yet unidentified factors cannot be dismissed. The cellular levels of these transcription factors were unaffected by HIV infection, with the exception of a decrease in expression of NF-kappaB p65, consistent with the observed decrease in its binding to the IL-12 p40 promoter following HIV infection. Analysis of regulation of upstream LPS-induced MAP kinases demonstrated impaired phosphorylation of JNK and p38 MAPK, and suppressed phosphorylation and degradation of IkappaBalpha following HIV infection. These results suggest that alterations in nuclear factor binding to numerous sites in the IL-12 p40 promoter, together may contribute to the suppression in IL-12 p40 transcription previously reported. These effects on nuclear factor binding may be a direct effect of HIV infection on the IL-12 p40 promoter, or may occur indirectly as a consequence of altered MAP kinase activation. PMID- 15270851 TI - Escherichia coli up-regulates proinflammatory cytokine expression in granulocyte/macrophage lineages of CD34 stem cells via p50 homodimeric NF-kappaB. AB - Umbilical cord blood has emerged as an alternative source of haematopoietic CD34+ cells for allogeneic stem cell transplantation. Although bacteraemia induced by Escherichia coli is considered one of the complications of transplantation, expression of proinflammatory cytokines is poorly understood. In this study, we report the altered expression of proinflammatory cytokines in CD34+ cells and their in vitro cultured cells following E. coli infection. CD34+ stem cells and their cultured cells up-regulated expression of proinflammatory cytokines such as interleukin (IL)-1alpha, IL-6, IL-8 and tumour necrosis factor (TNF)-alpha after infection with E. coli. Expression of the proinflammatory cytokines was generated mainly by the granulocyte-macrophage lineages. E. coli infection activated the signals of p50/p50 nuclear factor-kappaB (NF-kappaB) homodimers and IkappaB kinase. Furthermore, inhibition of NF-kappaB activation lowered the up-regulated expression of the proinflammatory cytokines. These results suggest that CD34+ cells and their cultured cells infected with E. coli induce the expression of proinflammatory cytokines via the NF-kappaB pathway. PMID- 15270852 TI - Environmental factors and not genotype influence the plasma level of interleukin 1 receptor antagonist in normal individuals. AB - Cytokine production may be regulated by both genotypic (single nucleotide or tandem repeat polymorphisms) and non-genotypic factors relating to the environment and inherent biology (i.e. gender). Interleukin (IL)-1 is one of the body's most highly proinflammatory cytokines and is implicated in the pathophysiology of numerous diseases, but also in the maintenance of homeostasis in a number of tissues. The cytokine IL-1 receptor antagonist (IL-1Ra) is the competitive inhibitor of the IL-1 agonists IL-1alpha and IL-1beta. In vivo IL-1Ra was measured in a cohort of 200 + blood donors and the effect of the IL-1 gene polymorphisms, environmental and biological factors assessed. In this study, we observed that possession of particular alleles of 5 IL-1 gene polymorphisms (IL1A 889, IL1Alpha VNTR, IL1B -511, IL1B +3953 and the IL1RN VNTR) did not correlate with higher plasma IL-1Ra levels. Environmental factors such as smoking and non steroidal anti-inflammatory drug ingestion were associated with higher in vivo IL 1Ra levels (P = 0.015 and 0.022, respectively), but biological factors such as gender, age and menstruation status did not have any impact upon in vivo IL-1Ra levels. Genotypic associations of IL-1 gene family polymorphisms with disease features may reflect characteristics of stressed rather than normal control circuits for cytokine production. PMID- 15270853 TI - Increase in Ksp37-positive peripheral blood lymphocytes in mild extrinsic asthma. AB - Killer-specific secretory protein of 37 kDa (Ksp37), identified as a Th1/Tc1 specific secretory protein is expressed preferentially in cytotoxic T lymphocytes (CTL) and natural killer (NK) cells and might be involved in essential processes of CTL-mediated immunity. Although extrinsic asthma is linked currently to a Th2 dominated pathogenesis, there is increasing evidence for Th1/Tc1-mediated processes in the aetiopathology of asthma. CTL from patients with asthma have been shown to express cytokines and effector molecules which were different from healthy controls. We hypothesized that Ksp37 could indicate the involvement of CTL in the pathogenesis of extrinsic asthma. We therefore investigated Ksp37 expression in PBMC from patients with mild extrinsic asthma (n = 7) and healthy controls (n = 7). Flow cytometric analysis was used to quantify Ksp37+ cells and to investigate cellular Ksp37 expression as relative mean fluorescence intensities (MFI). We found a significantly (P = 0.016) higher percentage of Ksp37+ cells within the total lymphocyte population obtained from patients with mild extrinsic asthma compared with healthy controls. Subdifferentiation revealed a significant difference limited exclusively to the CD8+ subset (P = 0.010). In addition, Ksp37 secretion from cultured peripheral blood mononuclear cells (PBMC) and MFI of Ksp37+ lymphocytes were increased in patients with asthma compared with healthy controls. We conclude that mild extrinsic asthma appears to be associated with an increased expression of the Tc1 related protein Ksp37. The functional role of Ksp37 in the pathogenesis of asthma remains to be elucidated. PMID- 15270854 TI - Nasal mucosa in natural colds: effects of allergic rhinitis and susceptibility to recurrent sinusitis. AB - The mechanisms of virus-induced airway hyperresponsiveness in asthma and allergy and the failure of host defence in patients suffering from secondary airway infections are still largely unknown. The aim of this study was to examine whether the presence of allergic rhinitis or susceptibility to recurrent sinusitis affects the structural and cellular changes in nasal mucosa during natural colds and convalescence. We compared the mucosal changes in biopsy samples during acute natural colds (days 2-4 of illness) and convalescence (3 weeks later) in patients with allergic rhinitis (n = 9), patients with susceptibility to sinusitis (n = 19) and healthy controls (n = 20). We saw similarly increased numbers of mucosal T and B lymphocytes and mast cells and increased vascular density during the acute colds compared to convalescence in all the three groups. The allergic subjects had elevated levels of eosinophils in the acute phase (P = 0.03), and the allergic and sinusitis-prone subjects had elevated levels of epithelial T cells (P = 0.04) and low levels of mast cells (P = 0.005) in convalescence compared to the control group. The sinusitis-prone subjects lacked intraepithelial cytotoxic cells in convalescence. In the allergic subjects, the reticular basement membrane was thicker in the acute phase compared to the convalescence (P = 0.05). These results suggest that various cells of the airways, including inflammatory and structural cells, are involved during viral respiratory infections in subjects with allergic rhinitis. The small numbers of mast cells and cytotoxic lymphocytes in the sinusitis-prone subjects may be related to their susceptibility to bacterial complications. PMID- 15270855 TI - Prospective immunological profiling in a case of immune dysregulation, polyendocrinopathy, enteropathy, X-linked syndrome (IPEX). AB - IPEX syndrome is a genetic autoimmune disease characterized by immune-mediated polyendocrinopathy, enteropathy, and X-linked inheritance. We describe a case of IPEX in which lymphocyte phenotypes were assessed at birth, before initiation of Cyclosporin A therapy, and at frequent intervals to 18 months of age. We performed flow cytometry for lymphocyte subtypes and for activation markers (HLA DR, CD25, and CD69 or CD71). The ratios of both T to B cells and CD4+ to CD8+ cells were elevated at birth, but CD4+ cells were not activated. HLA-DR+ and CD25+ activated T-cells increased in association with two episodes of clinical deterioration: colitis and the onset of type I diabetes mellitus. These results indicate that measures of activation, particularly HLA-DR+ and CD25+ frequency, correlate well with the development of early active disease and may presage clinical episodes. Continuous maintenance of immunosuppression, once started, appears critical for prevention of permanent tissue damage. PMID- 15270856 TI - beta-Defensin-3 and -4 in intestinal epithelial cells display increased mRNA expression in ulcerative colitis. AB - mRNA expression of two recently described human beta-defensins (hBD-3 and hBD-4) in epithelial cells of normal small and large intestine and the impact of chronic intestinal inflammation on their expression levels was investigated. Intestinal specimens from patients with ulcerative colitis (UC), Crohn's disease (CD) and controls with no history of inflammatory bowel disease were studied. hBD-3 and hBD-4 mRNAs were determined in freshly isolated epithelial cells by real-time quantitative reverse transcription-polymerase chain reaction (QRT-PCR) and by in situ hybridization. The effect of proinflammatory cytokines on hBD-3 and hBD-4 mRNA expression in colon carcinoma cells was also investigated. Purified epithelial cells of normal small and large intestine expressed both hBD-3 and hBD 4 mRNA, with higher expression levels of hBD-3 mRNA. In situ hybridization revealed higher levels of mRNA expression in the crypt- compared to the villus/luminal-compartment. Interferon (IFN)-gamma, but not tumour necrosis factor (TNF)-alpha or IL-1beta, augmented hBD-3 mRNA expression. None of these agents stimulated hBD-4 expression. Colonic epithelial cells from patients with UC displayed a significant increase in hBD-3 and hBD-4 mRNA compared to epithelial cells of controls. In contrast, small intestinal epithelial cells from CD patients did not show increased expression levels compared to the corresponding control cells. Moreover, Crohn's colitis did not show increased expression of hBD-4 mRNA, while the data are inconclusive for hBD-3 mRNA. We conclude that the chronic inflammatory reaction induced in the colon of UC patients enhances hBD-3 and hBD-4 mRNA expression in the epithelium, whereas in CD this is less evident. PMID- 15270857 TI - Anti-actin IgA antibodies in severe coeliac disease. AB - Anti-actin IgA antibodies have been found in sera of coeliacs. Our aim was to define the prevalence and clinical significance of anti-actin IgA in coeliacs before and after gluten withdrawal. One hundred and two biopsy-proven coeliacs, 95 disease controls and 50 blood donors were studied. Anti-actin IgA were evaluated by different methods: (a) antimicrofilament positivity on HEp-2 cells and on cultured fibroblasts by immunofluorescence; (b) anti-actin positivity by enzyme-linked immuosorbent assay (ELISA); and (c) presence of the tubular/glomerular pattern of anti-smooth muscle antibodies on rat kidney sections by immunofluorescence. Antimicrofilament IgA were present in 27% of coeliacs and in none of the controls. Antimicrofilament antibodies were found in 25 of 54 (46%) coeliacs with severe villous atrophy and in three of 48 (6%) with mild damage (P < 0.0001). In the 20 patients tested, antimicrofilaments IgA disappeared after gluten withdrawal in accordance with histological recovery. Our study shows a significant correlation between antimicrofilament IgA and the severity of intestinal damage in untreated coeliacs. The disappearance of antimicrofilament IgA after gluten withdrawal predicts the normalization of intestinal mucosa and could be considered a useful tool in the follow-up of severe coeliac disease. PMID- 15270858 TI - Role of peptide antigen for induction of inhibitory antibodies to Streptococcus mutans in human oral cavity. AB - The alanine-rich repeating region (A-region) in the surface protein antigen (PAc) of Streptococcus mutans has received much attention as an antigenic component for vaccines against dental caries. The PAc (residue 361-386) peptide in the A-region possesses a multiple binding motif (L- -V-K- -A) to various HLA-DR molecules and a B-cell core epitope (- Y- - -L- -Y- - - -) that recognizes the inhibiting antibody to S. mutans. In the present study, we investigated the immunogenicity of the PAc (361-386) peptide in humans and regulators of induction of the anti PAc (361-386) peptide IgA antibody (aPPA) in saliva. The PAc (361-386) peptide was confirmed as an ideal peptide antigen for induction of the inhibiting antibody to S. mutans in 151 healthy human subjects (36.6 +/- 12.6 years old) by quantitative analyses of oral bacteria and ELISA, as the aPPA titre in human saliva decreased significantly in an age-dependent manner. Homozygous DRB1*0405 and 1502, and heterozygous DRB1*0405/1502 showed a negative association with production of aPPA and tended to reduce the number of total streptococci in saliva. In contrast, the DRB1*1501 allele was significantly correlated with a high level of induction of the antibodies, and also tended to reduce lactobacilli and mutans streptococci. Further, peptide immunogenicity was confirmed in NOD SCID mice grafted with human peripheral blood mononuclear cells. Our results indicate that the interplay between regulators such as age, DRB1 genotype, cytokines, and peptide immunogenicity may provide a potential means for developing a vaccine useful for the prevention of dental caries as well as their diagnosis. PMID- 15270859 TI - Rapid simultaneous measurement of multiple cytokines using 100 microl sample volumes--association with neonatal sepsis. AB - Early diagnosis of neonatal infection has proved problematic due to the inadequacy of currently available laboratory tests. Neonatal sepsis is associated with an increase in plasma-derived cytokine levels, but an increase of a single cytokine cannot identify neonatal sepsis specifically and multiple cytokine levels are required. The time constraints and relatively large volume of plasma required to measure multiple cytokines from newborn infants by conventional enzyme-linked immunosorbent assay (ELISA) techniques is prohibitive. We therefore applied cytometric bead array (CBA) technology for simultaneous measurement of multiple cytokines from a group of 18 term neonates with infection confirmed by culture and a control group. 'Normal' ranges were established for each cytokine from 1-7-, 8-14- and 15-21-day-old newborns. There was no significant change in the levels of cytokines from infants in different control age groups, suggesting that basal cytokine levels are unchanged in the first 3 weeks of life. In the patient groups, however, there was a significant difference in several cytokines between the different age groups. Interleukin (IL)-6, IL-10 and IL-12 were increased significantly in the 1-7-day-old patient group compared to either the 8 14 and 15-21 age group, suggesting that infection in utero is associated with increased levels of these cytokines compared to infection acquired following birth. When individual patient cytokine levels were compared to normal control reference ranges, two patients failed to show significant elevation of any cytokine tested. All other patients showed elevated levels of between one and nine cytokines tested (mean of 4.6). There was no correlation between elevated cytokine levels and types of infective organism or patient age. In conclusion, neonatal sepsis is associated with the elevation of multiple plasma cytokines. The use of CBA kits is a rapid, easy, low sample volume and sensitive method to measure multiple plasma cytokines. PMID- 15270860 TI - CD56(+dim) and CD56(+bright) cell activation and apoptosis in hepatitis C virus infection. AB - CD3- CD56(+dim) natural killer (NK) cells, which are cytotoxic against virally infected cells, may be important in hepatitis C virus (HCV)-infected patients who are successfully treated with pegylated interferon (PEG-IFN)-alpha. We used flow cytometry to enumerate activated (CD69+) and apoptotic (annexin-V+) dim (CD3- CD56(+dim)) and bright (CD3- CD56(+bright)) NK cells obtained from HCV-infected patients before treatment (n=16) and healthy controls (n=15) in the absence and presence of pegylated interferon (PEG-IFN)-alpha-2b. A subset of HCV-infected patients, subsequently treated with PEG-IFN-alpha-2b in vivo, was determined to have a sustained virological response (SVR, n=6) or to not respond (NR) to treatment (n=5). In the absence of IFN, activated dim (CD3- CD56(+dim) CD69+) NK cells were significantly decreased (P=0.04) while activated apoptotic dim (CD3- CD56(+dim)CD69+ annexin-V+) NK cells tended to be increased (P=0.07) in SVR patients compared with NR patients. Activated bright (CD3-CD56(+bright)CD69+) and activated apoptotic bright (CD3- CD56(+bright)CD69+ annexin-V+) NK cells were significantly correlated (P=0.02 and P=0.01, respectively) with increasing hepatic inflammation. These findings suggest that in the absence of PEG-IFN, activated dim (CD3- CD56(+dim)CD69+) NK cell turnover may be enhanced in SVR compared with NR patients and that activated bright (CD3- CD56(+bright)CD69+) NK cells may play a role in liver inflammation. PMID- 15270861 TI - Isolation and functional analysis of circulating dendritic cells from hepatitis C virus (HCV) RNA-positive and HCV RNA-negative patients with chronic hepatitis C: role of antiviral therapy. AB - Hepatitis C virus (HCV) RNA has been localized in antigen-presenting dendritic cells (DCs) from patients with chronic hepatitis C (CHC). DCs from patients with CHC also exhibit impaired functional capacities. However, HCV RNA in DCs and functional impairment of DCs in CHC might be independent or interrelated events. Moreover, the impact of antiviral therapy on the functions of DCs in CHC is not well documented. In order to address these issues, we took advantage of antiviral therapy in these patients. Ten patients with CHC, expressing HCV RNA in circulating DCs, became negative for HCV RNA in circulating DCs after therapy with interferon-alpha and ribavirin for 4 weeks. The functions of DCs from HCV RNA+ patients (isolated before antiviral therapy) and HCV RNA- patients (isolated 4 weeks after antiviral therapy) were compared in allogenic mixed leucocyte reactions. In comparison to circulating DCs from normal control subjects, DCs from HCV RNA+ patients had a significantly decreased capacity to stimulate allogenic T lymphocytes (P < 0.01) and produce interleukin-12 (P < 0.05). However, the allostimulatory capacity of circulating DCs from HCV RNA- patients was several-fold higher compared to that of HCV RNA+ DCs from the same patient. DC from HCV RNA- patients also produced significantly higher levels of interleukin-12 compared to HCV RNA+ DCs from the same patient (P < 0.01). Taken together, this study is the first to provide experimental evidence regarding the impact of HCV RNA and antiviral therapy on the function of DCs in patients with CHC. PMID- 15270862 TI - HTLV-1 modifies the clinical and immunological response to schistosomiasis. AB - The immunological response in HTLV-1 infected individuals is characterized by a prominent Type-1 cytokine response with high production of IFN-gamma and TNF alpha. In contrast, helminthic infections and in particular chronic schistosomiasis are associated with a predominant production of IL-4, IL-5, IL-10 and IL-13. Liver fibrosis is the main pathological finding in schistosomiasis that occurs after many years of infection. This pathology is T cell dependent but the immune response mechanisms are not completely understood. The North-east region of Brazil is endemic for both HTLV-1 and schistosomiasis. In the present study the immune response, clinical severity, and therapeutic response to praziquantel of patients with schistosomiasis coinfected with HTLV-1 were compared with patients infected only with S. mansoni. Patients with HTLV-1 and S. mansoni had lower levels of IL-5 (P < 0.05) and higher levels of IFN-gamma (P < 0.05) in cultures stimulated with S. mansoni antigen and decreased S. mansoni antigen specific IgE levels when compared with patients with schistosomiasis without HTLV-1 coinfection. Liver fibrosis was mild in all HTLV-1 coinfected patients and efficacy of praziquantel was lower in patients dually infected than in patients infected only with S. mansoni. PMID- 15270863 TI - Osteoprotegerin (OPG) acts as an endogenous decoy receptor in tumour necrosis factor-related apoptosis-inducing ligand (TRAIL)-mediated apoptosis of fibroblast like synovial cells. AB - We examined the role of osteoprotegerin (OPG) on tumour necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis in rheumatoid fibroblast-like synovial cells (FLS). OPG protein concentrations in synovial fluid from patients with rheumatoid arthritis (RA) correlated with those of interleukin (IL)-1beta or IL-6. A similar correlation was present between IL-1beta and IL-6 concentrations. Rheumatoid FLS in vitro expressed both death domain-containing receptors [death receptor 4 (DR4) and DR5] and decoy receptors [decoy receptor 1 (DcR1) and DcR2]. DR4 expression on FLS was weak compared with the expression of DR5, DcR1 and DcR2. Recombinant TRAIL (rTRAIL) rapidly induced apoptosis of FLS. DR5 as well as DR4 were functional with regard to TRAIL-mediated apoptosis induction in FLS; however, DR5 appeared be more efficient than DR4. In addition to soluble DR5 (sDR5) and sDR4, OPG administration significantly inhibited TRAIL-induced apoptogenic activity. OPG was identified in the culture supernatants of FLS, and its concentration increased significantly by the addition of IL-1beta in a time dependent manner. Neither IL-6 nor tumour necrosis factor (TNF)-alpha increased the production of OPG from FLS. TRAIL-induced apoptogenic activity towards FLS was reduced when rTRAIL was added without exchanging the culture media, and this was particularly noticeable in the IL-1beta-stimulated FLS culture; however, the sensitivity of FLS to TRAIL-induced apoptosis itself was not changed by IL-1beta. Interestingly, neutralization of endogenous OPG by adding anti-OPG monoclonal antibody (MoAb) to FLS culture restored TRAIL-mediated apoptosis. Our data demonstrate that OPG is an endogenous decoy receptor for TRAIL-induced apoptosis of FLS. In addition, IL-1beta seems to promote the growth of rheumatoid synovial tissues through stimulation of OPG production, which interferes with TRAIL death signals in a competitive manner. PMID- 15270864 TI - The number of CD8+ T cells and NKT cells increases in the aqueous humor of patients with Behcet's uveitis. AB - To determine whether there are differences in the immunopathogenesis of different endogenous uveitis syndromes, the phenotypic characteristics of immune cells were analysed among patients with endogenous uveitis. The aetiology of the uveitis included idiopathic recurrent acute anterior uveitis (18 patients), idiopathic intermediate uveitis (13 patients), Behcet's uveitis (17 patients), Vogt-Koyanagi Harada syndrome (7 patients), and so on. Flow cytometric analysis was performed using immune cells of the aqueous humor and the peripheral blood during the active phase of intraocular inflammation, and monoclonal antibodies to CD3, CD4, CD8, CD14, CD19, CD56, TCR gammadelta, pan TCR alphabeta and Valpha24. CD8+ T cells were predominant in the aqueous humor of the patients with Behcet's uveitis, whereas CD4+ T cells were mainly found in the aqueous humor of patients other than those with Behcet's uveitis. The number of NKT (CD3+CD56+) cells was significantly higher both in the aqueous humor and the peripheral blood of the patients with Behcet's uveitis compared with the other groups (P < 0.05). CD8+CD56+ cells were the predominant subtype of the increased NKT cells in patients with Behcet's uveitis. In addition, intraocular infiltration of CD14+ cells significantly differed among the uveitis patients (P < 0.05). These results suggest that the immunopathogenesis of endogenous uveitis can vary between syndromes, and that CD8+CD56+ NKT cells may play an important role in the immunopathogenesis of Behcet's uveitis. PMID- 15270865 TI - Th1 and Th2 cytokine production is suppressed at the level of transcriptional regulation in Kawasaki disease. AB - To clarify the functional state of T cells in Kawasaki disease, we analysed mRNA expression levels of Th1/Th2 cytokines (IFN-gamma and IL-4) along with Th1/Th2 inducing transcription factors, T-bet and GATA-3, which play pivotal roles in the development of Th1 and Th2 cells, respectively. By real-time PCR, IFN-gamma mRNA levels in peripheral blood mononuclear cells (PBMNC) were significantly decreased in Kawasaki disease patients compared with those with measles, and tended to be lower than those in healthy controls. T-bet mRNA levels were significantly decreased in patients with Kawasaki disease compared with healthy controls. In addition, IL-4 and GATA-3 mRNA levels were significantly decreased in Kawasaki disease compared with healthy controls. Regulatory cytokine mRNA levels (TGF-beta and IL-10) were also decreased in Kawasaki disease. The mRNA levels of IFN-gamma showed a significant positive correlation with those of T-bet in Kawasaki disease. These results suggest that the suppressed function of Th1 and Th2, associated with the suppression of both T-bet and GATA-3 gene expression, may be one of the immunological characteristics of Kawasaki disease. PMID- 15270866 TI - Ethics and conflicts of interest in the BJD. PMID- 15270867 TI - Psoriasis: immunopathogenesis and evolving immunomodulators and systemic therapies; U.S. experiences. AB - BACKGROUND: Psoriasis is a chronic inflammatory skin disorder that is presently without a permanent cure. Up to 40% of patients with psoriasis also develop psoriatic arthritis. The mainstay armamentarium to treat psoriasis systemically includes methotrexate, cyclosporin and oral retinoids, all with significant potential for toxicity and the need for close laboratory supervision. The although the exact mechanism of psoriasis is still unclear, the involvement of T cell-mediated cytokine expression in the aetiology of psoriasis is becoming clearer. The goal of modern treatment is to target such immune responses that lead to the formation of psoriatic plaques and psoriatic arthritis using selective immunomodulating pharmacotherapy. The advantages of these biological agents are less toxic systemic side-effect profiles that will improve the quality of life in psoriatic patients. OBJECTIVES: This review article describes current and emerging selective immunotherapies and systemic therapies for the treatment of psoriasis, and will briefly discuss disease immunopathogenesis. METHODS: Literature review. RESULTS AND CONCLUSIONS: Given the role of the inflammatory immune responses in the pathogenesis of psoriasis, the goal of modern medicine and pharmacotherapy lies in the design and use of specific targets in cell mediated immune reactions and the modulation of the expression of various inflammatory cytokines. The clinical evidence of efficacy of some of these new biological immunomodulatory agents from several U.S.-based research studies and clinical experiences is convincing. PMID- 15270868 TI - The genetic epidemiology of alopecia areata in China. AB - BACKGROUND: Alopecia areata (AA) is hypothesized to be an organ-specific autoimmune disease with genetic predisposition and an environmental trigger. There are few clinical data in Asians. OBJECTIVES: To describe the genetic epidemiological features of AA patients in China and to determine the possible genetic model for AA. METHODS: Data for 1032 patients with AA were obtained by questionnaire in the Institute of Dermatology of Anhui Medical University in China from 2001 to 2003. Complex segregation analysis and heritability analysis were performed using Falconer's method, EPI INFO 6.0 and SAGE-REGTL programs. RESULTS: In total, 1032 AA patients (male/female ratio 1.1 : 1) were enrolled, representing 0.94% of the total number of cases seen in our outpatient clinic during that time. The mean +/- SD age of onset was 28.98 +/- 13.43 years. The difference between the mean age of onset in males and females was not significant. Most patients (82.6%) experienced their first episode of AA within the first four decades of life. A positive family history of AA was obtained in 87 patients (8.4%). The prevalence of AA in first-, second- and third-degree relatives of the proband with AA was 1.6%, 0.19% and 0.03%, respectively. These figures were higher than those in controls. A greater severity and longer duration of AA were seen in the early onset group than in the late-onset group. The early onset group also had more affected first- and second-degree relatives. The heritability of AA in first-, second- and third-degree relatives was 47.16%, 42.53% and 22.29%, respectively. Based on the REGTL results, the best model was a polygenic additive model for AA. CONCLUSIONS: The effect of genetic factors is strong in AA, but environmental factors such as infection and psychological stress may still play an important role. Our findings on the genetics of AA are consistent with a polygenic additive mode of inheritance. PMID- 15270869 TI - Psoriatic patients with arthropathy show significant expression of free HLA class I heavy chains on circulating monocytes: a potential role in the pathogenesis of psoriatic arthropathy. AB - BACKGROUND: Surface free heavy chains on monocytes were recently implicated in playing a role in the pathogenesis of several forms of arthritis. OBJECTIVES: To determine the expression of surface free heavy chains (recognized by monoclonal antibody HC10) on peripheral blood mononuclear cells of psoriatic patients with or without arthropathy. METHODS: Twenty-eight psoriatic patients from the dermatology outpatient clinic were included in this study. Blood samples were collected during outpatient visits and clinical characteristics of the patients were documented. Quantitative analyses of circulating mononuclear cells were performed using flow cytometry. RESULTS: Circulating monocytes showed higher expression of HC10 compared with circulating lymphocytes (P < 0.05). Psoriatic patients with arthropathy showed elevated expression of HC10 on peripheral blood monocytes compared with those without arthropathy (P < 0.05). Among the arthropathic group, those without the human leucocyte antigen (HLA)-B27 allele showed even higher expression of HC10 on circulating monocytes compared with those possessing HLA-B27 (P < 0.05). The polyarthropathic subgroup showed the highest HC10 expression, but the level of expression was not high enough to be of statistical significance compared with other arthropathic subgroups. No correlation was found between psoriatic skin involvement and the expression of HC10 on circulating monocytes. CONCLUSIONS: The presence of free heavy chains on circulating monocytes is closely associated with psoriatic arthropathy, while the expression of free heavy chains on circulating monocytes has no significant influence on psoriatic skin lesions. PMID- 15270870 TI - Nickel-responding T cells are CD4+ CLA+ CD45RO+ and express chemokine receptors CXCR3, CCR4 and CCR10. AB - BACKGROUND: Whereas T lymphocytes are widely accepted as effector cells determining the pathogenesis of allergic contact dermatitis, contradictory results have been found regarding the roles of different T-cell subsets. The use of various experimental models, involving long-term cultured T-cell lines or clones, may explain these contradictory results. OBJECTIVE: To investigate the involvement of distinct T-cell subsets in patients with nickel contact allergy. METHODS: Different T-cell subsets were directly isolated from peripheral blood mononuclear cells (PBMCs) of nickel-allergic patients, and their proliferative capacity, type-1 or type-2 cytokine secretion [measured by interferon (IFN)-gamma or interleukin (IL)-5 release] and phenotypical marker expression were analysed after stimulation with nickel. RESULTS: Only CD4+ CLA+ CD45RO+ and not CD8+ T cells proliferate and produce both type-1 (IFN-gamma) and type-2 (IL-5) cytokines in response to nickel. Moreover, cells expressing the marker CLA in combination with CD4, CD45RO or CD69 are increased after nickel-specific stimulation. Interestingly, in addition, CD45RA+ CLA+ cells showed an increased frequency after allergen-specific stimulation. Analysis of nickel-reactive T cells for expression of distinct chemokine receptors showed that both proliferative capacity and cytokine production are restricted to subsets expressing CXCR3, CCR4 but not CCR6. Fluorescence-activated cell sorting analysis of chemokine receptors expressed on nickel-stimulated T cells confirmed these results; a subset of T cells expressing CLA and CXCR3, CCR4 and, most importantly, CCR10 increased in response to allergen, while these CLA+ nickel-reactive T cells were all negative for CCR6. CONCLUSIONS: These findings demonstrate that freshly isolated nickel reactive T cells can be characterized as CD4+ CLA+ memory T cells which express the chemokine receptors CXCR3, CCR4 and CCR10, but not CCR6. PMID- 15270871 TI - Increased Fas ligand expression by T cells and tumour cells in the progression of actinic keratosis to squamous cell carcinoma. AB - BACKGROUND: In the counterattack model of tumorigenesis, it has been proposed that tumours develop resistance to attack from Fas ligand (FasL)-expressing cytotoxic T cells by downregulating Fas (immune escape), while at the same time upregulating FasL expression to induce apoptosis in Fas-expressing T cells (counterattack). OBJECTIVES: The aim of this study was to examine Fas and FasL expression on tumour cells and infiltrating T cells during the progression of actinic keratoses (AK), the benign precursor lesion, to squamous cell carcinoma (SCC). PATIENTS AND METHODS: Samples of AK (n = 20) and SCC (n = 20) were collected from immunocompetent patients attending dermatology clinics. Double label immunohistochemistry was performed on frozen sections using mouse monoclonal antibodies to Fas or FasL, simultaneously with a rabbit polyclonal antibody to either CD3 or cytokeratin, markers of T cells and keratinocytes, respectively. Cell densities and the optical density of tumour Fas expression were measured using image analysis. RESULTS: FasL-expressing T cells were observed in nine of 19 SCCs, compared with three of 20 AKs (P < 0.05). FasL expressing tumour cells were found in nine of 18 SCCs, compared with only one of 20 AK specimens (P < 0.005). There was no difference in the number of Fas expressing T cells infiltrating AK and SCC. Fas expression by keratinocytes, measured by optical density, was lower in SCC (range 0.1-40, median 17) compared with AK (range 4-62, median 25) (P < 0.05). CONCLUSIONS: These results suggest that the greater numbers of FasL-expressing T cells infiltrating into SCC compared with AK are targeting Fas-expressing tumour cells. As AK cells progress to SCC, they subvert this T-cell-mediated killing of tumour cells by downregulating their Fas expression (immune escape). Furthermore, tumour cells upregulate their expression of FasL, possibly as a counterattack measure to induce apoptosis in the increased number of tumour-infiltrating T cells. Thus changes in Fas/FasL-mediated interactions between T cells and tumour cells occur during the progression of AK into SCC. PMID- 15270872 TI - Stress-induced modulation of skin immune function: two types of antigen presenting cells in the epidermis are differentially regulated by chronic stress. AB - BACKGROUND: Inflammatory skin diseases are exacerbated by psychiatric stress. Previous studies have shown that the activity of epidermal antigen-presenting cells (APCs), Langerhans cells (LCs) and keratinocytes (KCs), is affected by stress. Hapten application causes migration of LCs to draining lymph nodes (DLNs). Recently, we found that hapten application also activates epidermal cells (ECs) to mature potent APCs, and that the main APCs in these populations are KCs. Thus, DLN cells and ECs following hapten application are available for estimating the APC function of LCs and KCs in stress studies. OBJECTIVES: To investigate the mechanism of exacerbation of skin inflammation by chronic stress by observing the effect of isolation stress transversally on the skin immune and neurohormonal systems. METHODS: Contact sensitivity (CS) was elicited in BALB/c mice. The APC function of LCs and ECs following hapten application was assessed by the CS inducing activity in the recipient mice. Levels of neurohormonal transmitters and proinflammatory cytokines were measured by enzyme-linked immunosorbent assay. Cell surface molecules were detected using flow cytometry. Expression of mRNA for cytokines, neurohormonal receptors and a differentiation marker by ECs was determined by reverse transcription-polymerase chain reaction. RESULTS: Acute stress (2-day isolation) suppressed induction of CS, while chronic stress (30-day isolation) markedly enhanced induction of CS. DLN cells from chronically stressed mice contained increased numbers of LCs and exhibited enhanced APC function for inducing CS. In contrast, the APC function of KCs from these mice was markedly suppressed. Serum corticosterone levels were enhanced in acute stress, while substance P (SP) levels were enhanced in chronic stress. Corticotrophin-releasing hormone receptor-1 mRNA expression in ECs was enhanced in acute stress, while SP receptor (i.e. neurokinin-1 receptor) mRNA expression in ECs was enhanced in chronic stress. Production and mRNA expression of the proinflammatory cytokines interleukin-1 alpha and tumour necrosis factor-alpha by ECs following hapten application was markedly suppressed in chronic stress. Expression by ECs of E cadherin, which adheres LCs and KCs homophilically, was suppressed in chronic stress. In addition, these cells exhibited impaired differentiation, i.e. suppressed spontaneous proliferation and enhanced mRNA expression for transglutaminase-3. CONCLUSIONS: Chronic isolation stress may enhance CS responses by upregulation of the APC activity of LCs and the SP system. However, dysregulation of KC function and differentiation by chronic stress suggests that KCs may not contribute to the enhancement of the CS response positively. These complex changes suggest that chronic isolation stress in mice may provide a possible model system for studying the mechanism of exacerbation of skin inflammation by stress. PMID- 15270873 TI - Hereditary 'white nails': a genetic and structural study. AB - BACKGROUND: Hereditary subtotal leuconychia is a rare nail disease. The gene(s) underlying this phenotype is (are) not known. Immunohistochemical and ultrastructural studies of nails are performed infrequently. OBJECTIVES: To perform genetic linkage analysis and to assess ultrastructure and soft/hard keratin expression in hereditary white nails. METHODS: We have analysed microscopically and ultrastructurally the white nails of a patient from a family in which the trait is inherited in an autosomal dominant manner as an isolated symptom. No skin lesions or hair abnormalities could be detected. Genetic linkage studies were performed on DNA samples obtained from several members of the affected family. A longitudinal surgical biopsy of the nail from a great toe was split in two parts. One part was fixed in formalin and processed for histopathology. Another part was further subdivided and embedded either in Epon, following fixation in 2% glutaraldehyde, or in Lowicryl K4M, after fixation in 3% paraformaldehyde. Dewaxed nail sections and Lowicryl ultrathin sections were also stained with various antikeratin antibodies. RESULTS: Genetic linkage studies of the family pointed to the disease gene mapping to the chromosomal 12q13 region. Genes mapping within this chromosomal region include the genes coding for type II (basic) cytokeratins and hard keratins. The nail matrix presented an abnormal hypergranulosis. The upper part of the nail plate, originating from the proximal nail matrix, had a nonhomogeneous lamellar appearance, with numerous intracellular 'lipidic' vacuoles and 'empty' spaces separating keratin filament bundles. These cells were progressively shed at the nail surface. The cell loss was compensated by hyperproliferation of the distal matrix and of the nail bed keratinocytes, with persistent marked parakeratosis and loose arrangement of keratin bundles. The distal matrix and the nail bed contributed equally to formation of the lower plate. This presented the characteristics of a tissue composed of soft keratins. Accordingly, there was virtually no labelling with the Hb1 antibody to a basic hard keratin in the white nail, whereas the labelling with AE3 antibody to all type II keratins and with KL1 recognizing suprabasal soft keratins was normal or even enhanced. CONCLUSIONS: Genetic linkage indicates that the gene defect underlying the leuconychia in the family studied resides on chromosome 12q13. As the type II keratins map within this chromosomal interval, it is possible that a mutation in one of these keratin genes may be a cause of the hereditary leuconychia. The white appearance of nails in this disease seems to be due to an abnormal keratinization of cells originating from the proximal nail matrix, leading to the presence of abundant intracellular vacuoles and to a lesser compactness of keratins. PMID- 15270874 TI - The detection of IgG and IgA autoantibodies to desmocollins 1-3 by enzyme-linked immunosorbent assays using baculovirus-expressed proteins, in atypical pemphigus but not in typical pemphigus. AB - BACKGROUND: We have shown previously that human desmocollin (Dsc) 1 is recognized by IgA autoantibodies of subcorneal pustular dermatosis (SPD) type IgA pemphigus. However, the presence of IgG anti-Dsc autoantibodies is still controversial, and antibodies to Dsc2 and Dsc3 have not been clearly identified. OBJECTIVES: To investigate this by producing recombinant proteins consisting of the entire extracellular domains of human Dsc1, 2 and 3 in baculovirus, and to use them to establish an enzyme-linked immunosorbent assay (ELISA). METHODS: By this ELISA, we examined in total 165 cases of various types of autoimmune bullous diseases, as well as 23 normal controls. RESULTS: None of 45 sera of classical pemphigus showed either IgG or IgA antibodies to any Dsc. In contrast, one atypical pemphigus serum showed both IgG and IgA antibodies to Dsc1, which were adsorbed by incubation with Dsc1 baculoprotein. Furthermore, this ELISA detected both IgA and IgG anti-Dsc3 antibodies in one atypical case, and IgA antibodies to both Dsc2 and Dsc3 in another. This reactivity was confirmed by positive IgA immunofluorescence with Dsc2 and Dsc3 expressed on COS-7 cells. These results show that both IgG and IgA autoantibodies against all of Dsc1-3 are present in the sera of particular cases of nonclassical pemphigus, except for IgG antibodies to Dsc2, but that they are not detected in classical pemphigus. Unexpectedly, although IgA antibodies of all of eight SPD type IgA pemphigus sera reacted with Dsc1 expressed on COS-7 cells, only one serum was positive in Dsc1 ELISA for IgA. CONCLUSIONS: This result indicates either that Dscs expressed by baculovirus may not adopt the correct conformation or that Dscs may need association with other molecules to express all the epitopes for autoantibodies. PMID- 15270875 TI - Evaluation of inflammatory infiltrate and fibrogenic cytokines in pseudopelade of Brocq suggests the involvement of T-helper 2 and 3 cytokines. AB - BACKGROUND: Pseudopelade of Brocq (PB) is an acquired progressive cicatricial alopecia which is characterized by some distinctive clinical features. It may represent either a distinct entity, i.e. an idiopathic primary scarring alopecia, or the end stage of various forms of scarring alopecia such as discoid lupus erythematosus (DLE) or lichen planopilaris (LPP). OBJECTIVES: The aim of the study was to evaluate a set of patients with a clinically defined PB, to ascertain whether their PB was idiopathic or secondary, and then to study the phenotype of the inflammatory infiltrate and the presence of any fibrogenic and antifibrogenic cytokines to identify idiopathic or secondary forms in more detail. METHODS: Twelve female patients with PB were studied by means of histology, direct immunofluorescence (DIF) and immunohistochemistry, by using monoclonal antibodies to cell markers (lymphocyte subtypes, Langerhans cells, macrophages, fibroblasts, mastocytes and activation markers) and fibrogenic and antifibrogenic cytokines. RESULTS: Using histology and DIF, we diagnosed two cases as DLE and three cases as LPP. Seven cases had nonspecific histology or DIF appearances and were classified as noncharacterized pseudopelade (NCPB). Two major phenotypic patterns of dermal infiltrate were identified by immunohistochemistry. These were: (i) a conspicuous infiltrate of CD3+ cells with a high CD4+/CD8+ ratio, variable numbers of macrophages, mast cells and fibroblasts always fewer than lymphocytes; (ii) an infiltrate of CD3+ cells with variable CD4+/CD8+ ratio and conspicuous amounts of macrophages, mast cells and fibroblasts, more numerous than infiltrating lymphocytes. The first pattern was typical of DLE and LPP, the second one was typical of NCPB. Fibrogenic cytokines were observed in all cases, but basic fibroblastic growth factor (bFGF) and transforming growth factor (TGF)-beta were more strongly expressed in NCPB. Interferon (IFN)-gamma was found in LPP. CONCLUSIONS: In our PB patients we identified five of 12 secondary PB and seven of 12 idiopathic PB by means of histology and DIF. The phenotypic pattern of infiltration allowed us to further differentiate secondary (richer in lymphocytes) from idiopathic PB (richer in resident cells). The pattern of cytokine expression showed the presence of fibrogenic molecules (interleukins 4 and 6, bFGF and TGF-beta) in all cases, suggesting the involvement of mechanisms mediated by T-helper 2 and 3 cytokines in PB. PMID- 15270876 TI - Temozolomide and interferon alpha 2b in metastatic melanoma stage IV. AB - BACKGROUND: A multicentre, centrally randomized, open-labelled study with temozolomide and interferon (IFN)-alpha 2b was carried out to study the therapeutic effect in patients with metastatic melanoma stage IV. OBJECTIVES: The response rate, efficacy, side-effects, reasons for discontinuation of therapy and survival rate of 47 patients treated with temozolomide in combination with two different dosing regimens of IFN-alpha 2b were documented. PATIENTS/METHODS: Twenty-nine male and 18 female patients (mean age 57.6 years, range 34-74) were centrally randomized to two different arms: 20 patients received a treatment schedule with temozolomide 150 mg m(-2) on days 1-5 orally every 28 days in combination with IFN-alpha 2b 10 MIU m(-2) every other day and 27 patients received temozolomide 150 mg m(-2) on days 1-5 every 28 days in combination with IFN-alpha 2b in a fixed dose of 10 MIU every other day. RESULTS: We observed an overall response rate of 27.6% comprising five complete remissions (10.6%: one patient group A, four patients group B), in two of these five patients at the last follow-up in the study (4.3%, both in group B); and eight partial remissions (17%: six patients in group A, two patients in group B), in three of these eight patients at the last follow-up in the study (6.4%, two patients in group A, one patient in group B). Three patients showed stable disease (6.4%: one patient in group A, two patients in group B). Mean survival was 14.5 months [95% confidence interval (CI) 10-19] with no significant differences between treatment groups. However, there was a significant correlation with response after three cycles (log rank test, P < 0.03). Within the 32 patients who completed at least three cycles of therapy, seven patients (three in group A and four in group B) with a partial or complete response showed a significantly better mean survival of 30.6 months (95% CI 19.1-42) compared with 25 patients who did not respond (13.7 months 95% CI 9.2-18.3). In total, patients with at least one complete remission showed the longest survival (37.1 months 95% CI 26.3-47.9), followed by patients with at least one partial response (17.4 95% CI 10.9-23.9). Major side-effects of the treatment were nausea, vomiting, headache, leucopenia, thrombopenia, elevation of liver function parameters and neurological symptoms. In five patients, the side-effects led to a discontinuation of treatment: neurological symptoms (two patients), sepsis (one patient), brain haemorrhage (one patient) and exanthema (one patient). There were no treatment-related deaths. CONCLUSIONS: The combination of temozolomide and IFN-alpha 2b can easily be administered and shows tolerable toxicity. When an objective response occurs after three cycles, it indicates a significant survival advantage. PMID- 15270877 TI - Muckle-Wells syndrome: clinical and histological skin findings compatible with cold air urticaria in a large kindred. AB - BACKGROUND: Muckle-Wells syndrome is a rare familial disease with autosomal dominant inheritance, characterized by cold sensitivity and polyarthralgias since childhood, with possible later development of nerve deafness and renal amyloidosis. The nature of the skin manifestations is, however, not well characterized. OBJECTIVES: To clarify the nature of cutaneous cold sensitivity in patients with Muckle-Wells syndrome by studying clinical aspects and histological features. METHODS: Eighteen members of a family with Muckle-Wells syndrome and the recently identified mutation of the CIAS1 gene at locus 260 of chromosome 1q44 were available for study. Examination included a thorough history, physical examination and a battery of laboratory tests. In two brothers, standard cold contact and cold air provocation tests were performed, as were biopsies from normal and lesional skin. RESULTS: All affected family members reported an increased sensitivity to cold, dampness or changes in temperature, and most had arthritis and conjunctivitis. Eight had developed hearing loss, four renal involvement, and amyloid deposits were found in three of five patients in whom rectal biopsies were performed. Laboratory tests showed leucocytosis and elevated C-reactive protein, but no serum cold agglutinins and cryoglobulins. Skin eruptions, with weals of 0.2-3 cm, lasted from 5 to 24 h and were associated with local itching or pain as well as fever, malaise and chills. On cold provocation of two patients, lesions could be reproduced by cold air, but not by contact with an ice cube or cold water. On histology, there was increased vasodilatation, marked infiltration with neutrophils and monocytes/macrophages, and increased expression of beta 2 integrins in lesional vs. normal skin. Numbers of mast cells as well as expression of interleukin-3 and tumour necrosis factor-alpha were unchanged. CONCLUSIONS: Cold-induced skin lesions in Muckle-Wells syndrome represent typical generalized cold air/wind inflammatory reactions, as also observed in familial cold urticaria. Microscopic features are similar to those observed in other types of urticaria. PMID- 15270878 TI - Increased advanced oxidation protein products in Behcet's disease: a new activity marker? AB - BACKGROUND: Behcet's disease (BD) is a chronic inflammatory disease with unknown pathogenesis. As various functions of neutrophils in peripheral blood, such as chemotaxis, phagocytosis and generation of reactive oxygen species (ROS) increase in BD, ROS-mediated oxidative stress related to neutrophil activation may have an important role in the pathogenesis of BD. OBJECTIVES: To investigate the importance of neutrophil activation as the main source of oxidative stress through protein oxidation in the pathogenesis of BD, and also to investigate whether one of the products of protein oxidation, advanced oxidation protein products (AOPP), may be used as an activity marker for BD. METHODS: Patients with BD (n = 49), at active and inactive stages, with or without evidence of uveitis, and healthy volunteers (n = 40) were entered into the study. A full blood count, peripheral blood smears, routine biochemical analyses, C-reactive protein (CRP) and erythrocyte sedimentation rate (ESR) measurements were performed in all patients preceding the study. Plasma myeloperoxidase (MPO) activity, representing neutrophil activation, and biomarkers of oxidative stress reflecting protein oxidation, such as levels of AOPP and thiol, were measured spectrophotometrically. Statistical comparisons were made using Mann-Whitney U tests, Student's t-tests, anova/post-anova tests and correlation analyses. RESULTS: In all patients, the results of full blood count, peripheral blood smears and routine biochemical analyses were in the normal range, but mean values of CRP and ESR were higher than laboratory reference values. Plasma MPO activity and AOPP levels were found to be higher and thiol values lower in the total patient group and individual subgroups than in controls. Patients with active BD had significantly higher MPO and AOPP levels and lower thiol levels than patients with inactive BD. There was no difference between uveitis-positive and uveitis negative subgroups in MPO and thiol levels, but AOPP levels were lower in the latter group. Patients with active BD +/- uveitis were shown to have increased MPO and AOPP but decreased thiol levels in comparison with the inactive BD, uveitis-negative subgroup. There were strong positive correlations between ESR and CRP, ESR and MPO, ESR and thiol, ESR and AOPP, CRP and MPO, CRP and AOPP, MPO and AOPP, and thiol and AOPP levels in patients with BD. CONCLUSIONS: Based on this first study, in which MPO-mediated AOPP formation has been demonstrated, it may be suggested that activated neutrophils may play an important role in the pathogenesis of BD and that chlorinated oxidants of neutrophil origin may lead to oxidative stress, notably protein oxidation. Therefore, AOPP may be a useful marker for monitoring the progress and the severity of the disease activity. PMID- 15270879 TI - U-serrated immunodeposition pattern differentiates type VII collagen targeting bullous diseases from other subepidermal bullous autoimmune diseases. AB - BACKGROUND: Epidermolysis bullosa acquisita (EBA) can be differentiated from other subepidermal bullous diseases by sophisticated techniques such as immunoelectron microscopy, salt-split skin antigen mapping, fluorescence overlay antigen mapping, immunoblot and enzyme-linked immunosorbent assay. OBJECTIVES: To determine whether the diagnosis can also be made by routine direct immunofluorescence microscopy. METHODS: We studied frozen skin biopsies from 157 patients with various subepidermal immunobullous diseases. RESULTS: We found three distinct 'linear' fluorescence patterns at the basement membrane zone: true linear, n-serrated and u-serrated. The true linear pattern, often seen in conjunction with either the n- or the u-serrated pattern, was found in any subepidermal immunobullous disease with nongranular depositions. In bullous pemphigoid, mucous membrane pemphigoid, antiepiligrin cicatricial pemphigoid, p200 pemphigoid and linear IgA disease the n-serrated pattern was found, corresponding with depositions located in hemidesmosomes, lamina lucida or lamina densa. However, in EBA and bullous systemic lupus erythematosus the u-serrated staining pattern was seen, corresponding with the ultralocalization of type VII collagen in the sublamina densa zone. The diagnosis of EBA with IgG or IgA autoantibodies directed against type VII collagen was confirmed by immunoelectron microscopy, salt-split skin antigen mapping, fluorescence overlay antigen mapping or immunoblotting. CONCLUSIONS: Using this pattern recognition by direct immunofluorescence microscopy we discovered several cases of EBA which would otherwise have been erroneously diagnosed as a form of pemphigoid or linear IgA disease. PMID- 15270880 TI - Immunophenotyping of inflammatory cells in lesional skin of the extrinsic and intrinsic types of atopic dermatitis. AB - BACKGROUND: There is a subgroup of atopic dermatitis (AD) patients with normal total and specific IgE levels and negative skin tests towards common allergens. This form of the disease has been referred to as the 'intrinsic' form of AD. Although previous studies have demonstrated differences in the cytokine profile between the extrinsic and intrinsic subtypes, the pathogenesis of both subtypes of AD remains unclear. OBJECTIVES: To compare the inflammatory micromilieu in both forms of AD. METHODS: Immunophenotyping of the inflammatory cells was performed in lesional and nonlesional skin from 18 patients with extrinsic and 17 with intrinsic AD. RESULTS: Immunohistochemical analysis revealed a high proportion of CD4+ T cells in the dermis, with a similar CD4/CD8 ratio in the two groups. The expression levels of other T-cell markers and epidermal Langerhans cells were increased in both forms of AD. Although the T-cell repertoires in the two subtypes were similar, dermal infiltration of eosinophils and eosinophil granular proteins was more prominent in the extrinsic type than in the intrinsic type. Eotaxin immunoreactivity was also significantly higher in the extrinsic subtype. CONCLUSIONS: The data suggest that although the overall inflammatory microenvironment in the two subtypes appears to be similar, differences in T-cell cytokine production might contribute to the differential tissue eosinophilia in these subtypes. PMID- 15270881 TI - Evaluation of a BP180-NC16a enzyme-linked immunosorbent assay in the initial diagnosis of bullous pemphigoid. AB - BACKGROUND: Bullous pemphigoid (BP) is the most common subepidermal immunobullous disease, characterized by circulating IgG autoantibodies targeting BP180 and BP230 hemidesmosomal proteins. Several immunological studies have demonstrated that the membrane proximal noncollagenous domain NC16a of BP180 is the immunodominant region targeted by BP autoantibodies. Recently, a commercial BP180 NC16a-specific enzyme-linked immunosorbent assay (ELISA) has become available for detecting pathogenic anti-BP180 autoantibodies in BP sera. However, it remains unclear whether the diagnostic potential of the ELISA is equivalent to that of the 'gold-standard' diagnostic technique of immunofluorescence (IF). OBJECTIVES: To examine the usefulness of a commercially available BP180-NC16a ELISA in the initial serodiagnosis of BP. METHODS: Sera from a large cohort of patients with BP (n = 102) and control subjects (age- and sex-matched normal volunteers, n = 60; pemphigus foliaceus, n = 18; pemphigus vulgaris, n = 16) were assayed by BP180-NC16a ELISA. All BP sera were obtained at presentation before initiation of systemic immunosuppressive therapy. The values of IgG antibody levels measured by ELISA were compared with those measured by indirect IF on salt-split skin. Results Receiver operating characteristic analysis was used to calculate the cut off value for the ELISA in the diagnosis of BP which maximizes both sensitivity and specificity, and to estimate the diagnostic accuracy of the ELISA as represented by the area under the curve (AUC = 0.965). A cut-off value of 9 was associated with a sensitivity of 89% (91 of 102 BP sera showed a positive result) and a specificity of 98%. Fifty-eight of 60 normal controls and all the pemphigus sera showed a negative result. There was a correlation between the mean ELISA values and indirect IF titres (Spearman rank correlation 0.286; P = 0.004). CONCLUSIONS: Our results suggest that the BP180-NC16a ELISA is a useful tool for the detection of pathogenic anti-BP180 IgG autoantibodies at the initial disease stage of BP. Because it is not only highly sensitive and specific, but is also easy to perform, is objective, and semiquantitative, the ELISA may provide valuable information for the accurate and reliable serodiagnosis of BP. PMID- 15270882 TI - Enhancement of topical 5-aminolaevulinic acid delivery by erbium:YAG laser and microdermabrasion: a comparison with iontophoresis and electroporation. AB - BACKGROUND: 5-aminolaevulinic acid (ALA) is used as a protoporphyrin IX-precursor for the photodynamic therapy of superficial skin cancer and cutaneous metastases of internal malignancies. However, the permeability of hydrophilic ALA across the skin is very low. OBJECTIVES AND METHODS: The objective of this study was to optimize and enhance the in vitro skin permeation of ALA by two resurfacing techniques: erbium:yttrium-aluminium-garnet (Erb:YAG) laser and microdermabrasion. Light microscopic changes in pig skin caused by these techniques were also compared. The electrically assisted methods, iontophoresis and electroporation, were also used to facilitate ALA permeation across laser- or microdermabrasion-treated skin. RESULTS: Among the modalities tested in this study the Erb:YAG laser showed the greatest enhancement of ALA permeation. The laser fluence was found to play an important role in controlling the drug flux, producing enhancement ratios from 4-fold to 246-fold relative to the control. The skin permeation of ALA across microdermabrasion-treated skin was approximately 5 15-fold higher than that across intact skin. Both the ablated effect of the stratum corneum (SC) and ALA flux were proportional to the treatment duration of microdermabrasion. The application of iontophoresis or electroporation alone also increased the ALA permeation by approximately 15-fold and 2-fold, respectively. The incorporation of iontophoresis or electroporation with the resurfacing techniques caused a profound synergistic effect on ALA permeation. CONCLUSIONS: This basic study has encouraged the further investigation of ALA permeation by laser or microdermabrasion. PMID- 15270883 TI - Mohs' micrographic surgery for treatment of basal cell carcinoma of the face- results of a retrospective study and review of the literature. AB - BACKGROUND: The incidence of skin cancer and especially basal cell carcinoma (BCC) has increased in the last decade and is still increasing. Many treatment modalities can be used to treat BCC; surgical excision is the most frequently used. Mohs' micrographic surgery (MMS) is an advanced excision technique which is often used to treat BCC in the U.S.A. In Europe it is practised less frequently. OBJECTIVE: The aim of this article was to evaluate the efficiency of MMS for the treatment of facial BCC. METHODS: In a retrospective study recurrence rates after the treatment of facial BCC by MMS were estimated by reviewing the records of all patients with BCCs (620 patients with 720 BCCs) treated by MMS in our department from April 1992 until December 1999. RESULTS: The 5-year recurrence rates estimated from this study were 3.2% for primary BCC and 6.7% for recurrent BCC. Prognostic factors for recurrence are: an aggressive histopathological subtype, more than four Mohs' stages, a large defect size and a recurrent BCC. CONCLUSION: Based on the fact that MMS provides the lowest recurrence rates, it is the treatment of first choice for primary facial BCCs with an aggressive histopathological subtype and for recurrent BCCs in the face. PMID- 15270884 TI - Topical treatment of basal cell carcinoma with tazarotene: a clinicopathological study on a large series of cases. AB - BACKGROUND: Basal cell carcinoma (BCC) is the most common cancer in humans. Medical treatment modalities offer cost reductions and clinical advantages in selected cases such as low-risk areas, surgically inaccessible sites, patients with multiple neoplasms, and older, infirm or anticoagulated subjects. Tazarotene has been proposed for the treatment of BCC; however, data on its efficacy are lacking. OBJECTIVES: To investigate the efficacy of tazarotene in a large series of BCCs, better to define the clinical advantages and the mechanisms of action in vivo. METHODS: Tazarotene 0.1% gel was applied daily for 24 weeks to 154 small superficial and nodular BBCs. Clinicopathological changes were followed during the therapy by dermoscopic and histological examination. Proliferation, retinoic acid receptors and apoptosis were investigated by immunohistochemistry and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick-end labelling on biopsies. RESULTS: At 24 weeks of therapy, 70.8% of the BCCs showed > 50% clinical and dermoscopic regression, and 30.5% healed without recurrences after 3 years of follow-up. At 12 weeks, biopsies showed that regression was associated with reduced proliferation and increased apoptosis of basaliomatous cells. Most unresponsive tumours displayed a keratotic differentiation. CONCLUSIONS: Tazarotene was effective in the majority of superficial and nodular undifferentiated BCCs treated, possibly by antiproliferative and proapoptotic actions in vivo. Keratotic BCCs were the major type among the unresponsive tumours, and were characterized by overexpression of p53 and cellular retinol binding protein-1 in comparison with undifferentiated tumours. Topical tazarotene represents an alternative medical choice for selected cases of BCC. PMID- 15270885 TI - Nuclear beta-catenin in basal cell carcinoma correlates with increased proliferation. AB - BACKGROUND: Virtually all BCCs have deregulation of the Hedgehog (Hh) signalling pathway and a proportion show nuclear beta-catenin accumulation. The latter is thought to be due to Hh pathway-directed Wnt expression but this has not been tested. An alternative cause of nuclear beta-catenin accumulation is gene mutation, which stabilizes the protein. Theoretically, reduced E-cadherin expression could also be important because it can sequester beta-catenin at the cell membrane. In turn, nuclear beta-catenin can increase expression of MYC and cyclin D1, thus potentially altering proliferation. OBJECTIVES: To assess whether nuclear beta-catenin occurs in BCC, and to look at potential causes and consequences. METHODS: Nuclear beta-catenin was assessed by immunohistochemistry, and its causes by analysis of E-cadherin expression, beta-catenin exon 3 mutation and WNT5A expression. Its consequences were assessed by analysing proliferation. RESULTS: We found nuclear beta-catenin in 20 of 86 paraffin-embedded sections of BCCs using immunohistochemistry. BCCs showed increased WNT5A relative to the surrounding skin. No mutations in exon 3 of the beta-catenin gene were found in 10 cases. There was no association between beta-catenin localization and E cadherin expression. Tumours with nuclear beta-catenin had significantly higher proliferation (P < 0.01). CONCLUSIONS: The absence of beta-catenin gene mutations indicate that the Hh pathway-directed Wnt signalling remains the most likely cause of nuclear beta-catenin accumulation in BCC. Additionally, the correlation with increased proliferation is the first evidence that nuclear beta-catenin may have a biological effect. However, a causal link between Hh pathway deregulation, Wnt ligand overexpression, nuclear beta-catenin accumulation and increased proliferation remains to be confirmed. PMID- 15270886 TI - Immunohistochemical staining of cutaneous tumours with G-81, a monoclonal antibody to dermcidin. AB - BACKGROUND: Recently, the novel antimicrobial peptide named dermcidin (DCD) was reported in human eccrine sweat glands. OBJECTIVES: We investigated the expression of DCD in a variety of cutaneous tumours in order to assess the usefulness of the monoclonal antibody (G-81), which recognizes a fragment of DCD. PATIENTS/METHODS: We studied the immunoreactivity of the G-81 antibody on 197 cutaneous tumours. RESULTS: A total of 13 of 26 cutaneous mixed tumours showed substantial immunoreactivity. In contrast all the following cases were completely unreactive: (i) epithelial tumours (seborrhoeic keratosis, squamous cell carcinoma, Bowen's disease, actinic keratosis, genital Paget's disease); (ii) follicular tumours (basal cell carcinoma, trichilemmoma, trichoepithelioma, trichoblastoma, keratoacanthoma, proliferating trichilemmal tumour, pilomatricoma); (iii) melanocytic tumours (malignant melanoma, naevus cell naevus, Spitz naevus, blue naevus); (iv) neural tumours (schwannoma, neurofibroma, Merkel cell neoplasm); (v) mesenchymal tumours (soft fibroma, dermatofibroma, dermatofibrosarcoma protuberans, vascular leiomyoma, leiomyosarcoma, lipoma, juvenile xanthogranuloma, angiomyoma); and (vi) other sweat gland tumours (poroid neoplasms, syringoma, cylindroma, clear cell hidradenoma, spiradenoma, syringoid eccrine carcinoma, mucinous carcinoma, apocrine cystadenoma, syringocystadenoma papilliferum, apocrine adenocarcinoma). Twenty-six cutaneous mixed tumours were considered from histopathological findings to be the apocrine type, but 13 of 26 mixed tumours contained some DCD immunopositive cells that possibly differentiate into eccrine secretory glands. CONCLUSIONS: We found the expression of DCD in tubular structures of 50% of cutaneous mixed tumours with apocrine differentiation. These results suggest that a number of cutaneous mixed tumours show both eccrine and apocrine differentiation in the same neoplasm. PMID- 15270887 TI - Risk and protective factors for sporadic basal cell carcinoma: results of a two centre case-control study in southern Germany. Clinical actinic elastosis may be a protective factor. AB - BACKGROUND: There are very few data regarding sun exposure behaviour of patients with basal cell carcinoma (BCC) in central Europe. OBJECTIVES: A case-control study of patients with sporadic BCC was conducted to assess the risk of occupational and leisure-time sun exposure behaviour, precursor lesions for skin cancer and phenotypic factors on the development of sporadic BCC in Ulm and Dresden, Germany. METHODS: A comparison was made of 213 patients with BCC (128 from Ulm, 85 from Dresden; 103 men and 110 women; median age at diagnosis 69 years) and 411 controls (237 from Ulm, 174 from Dresden; 197 men and 214 women; median age 58 years). Crude odds ratios (ORs) and corresponding 95% confidence intervals for all of 64 possible risk factors revealed strong associations in 33 items. Selection of important risk factors was performed in a multiple logistic regression. RESULTS: For sporadic BCC, an increased risk was shown for persons with actinic cheilitis (OR 7.1), actinic keratosis (OR 2.7) and solar lentigo (OR 2.5). The only phenotypic factor indicating risk of sporadic BCC was hair colour, with a higher risk for red/fair than brown/black hair (OR 4.3). There was an increased risk for persons with BCC in first-degree relatives (OR 5.1) and those with sunburn 20 years before sporadic BCC was diagnosed (OR 3.6). Additionally, occupational ultraviolet (UV) exposure appeared to be a risk factor (OR 2.4). In contrast, clinical actinic elastosis showed a protective effect (OR 0.1). CONCLUSIONS: In contrast to earlier reports, clinical actinic elastosis turned out to be the only protective factor for sporadic BCC. A special relationship between wrinkling and BCC risk could not be shown. For basic research, future work should be aimed at elucidating further the different forms of collagen repair processes after intermittent and/or chronic UV exposure. The data strongly support the recommendation that a change in recreational UV exposure habits in individuals, and sunburn avoidance in particular, are necessary not only because of the increased long-term risk of melanoma, but also because of the risk of other skin cancers such as sporadic BCC. PMID- 15270888 TI - The optimal time to determine the minimal phototoxic dose in skin photosensitized by topical 8 methoxypsoralen. AB - BACKGROUND: We recently investigated the characteristics of psoralen plus ultraviolet (UV) A erythema in skin photosensitized by topical 8-methoxypsoralen (8-MOP) in three independent studies. OBJECTIVES: In order to determine the optimal time to read the minimal phototoxic dose (MPD) after treatment with topical 8-MOP and irradiation with UVA, we assessed the overall data. METHODS: One forearm of each subject was immersed in 8-MOP solution for 15 min and test sites on the flexor surface of the forearm were immediately exposed to a UVA dose series. Erythema was assessed visually and objectively using a reflectance instrument at 24-h intervals for 7 days. RESULTS: Results were obtained from 44 subjects (predominantly Fitzpatrick skin phototype II). A broad erythemal plateau was evident beyond 72 h and the visual MPD was significantly lower at 96, 120 and 144 h than at 72 h (P < 0.01). Only 30% of subjects were at peak erythema at the conventional MPD assessment time of 72 h. The median time to reach maximal erythema was 96 h (range 48-144). Objectively, 85% of subjects were at peak erythema at or beyond 96 h. CONCLUSIONS: We recommend that (i) the optimal time to read the topical 8-MOP MPD is 4 days after UVA exposure as readings beyond this time may be difficult to interpret because of the development of pigmentation, and (ii) 40% of the topical 8-MOP MPD should be considered for the first treatment. PMID- 15270889 TI - Anger and acne: implications for quality of life, patient satisfaction and clinical care. AB - BACKGROUND: Acne is a common skin disorder with a significant psychological and social impact for some people. Little is known about how personality and emotional traits affect acne and its impact on quality of life and treatment. Trait anger (TA), which is related to heart disease and other morbidities, may also affect acne and patients' adjustment to it. OBJECTIVES: To evaluate the relationship between TA and acne severity, skin-related quality of life, satisfaction with treatment, and adherence to treatment. PARTICIPANTS AND METHODS: A sample of 479 individuals with acne completed a survey instrument to assess acne severity, skin care practices, skin-related quality of life, satisfaction with treatment, adherence, TA and demographic variables. Respondents who reported high TA were compared with individuals with low TA on outcome variables. Regression analyses adjusted for covariates and identified the significant predictors of quality of life, satisfaction and adherence. RESULTS: High TA was unrelated to acne severity (P = 0.2) or frequency of face washing (P = 0.9). Anger was significantly related to both global quality of life (P < 0.001) and skin-related quality of life (P = 0.002) as well as to satisfaction with treatment (P = 0.001) and adherence to treatment advice (P = 0.05) in bivariate analyses. Regression analyses revealed that high TA remained a significant predictor of global (P < 0.001) and skin-related quality of life (P = 0.003) and satisfaction with treatment (P = 0.04), but not adherence to treatment advice (P = 0.8) after controlling for covariates. CONCLUSIONS: Anger is associated with the quality of patients' lives and with their satisfaction with treatment. Care of acne patients should include attention to anger and other chronic emotional states, quality of life, as well as to clinical severity. Simple guidelines are suggested for how clinicians might approach this important aspect of care. PMID- 15270890 TI - Topical paricalcitol (19-nor-1 alpha,25-dihydroxyvitamin D2) is a novel, safe and effective treatment for plaque psoriasis: a pilot study. AB - BACKGROUND: There continues to be a need to develop new pharmacological approaches for treating psoriasis. Topical active vitamin D compounds have proven to be both safe and effective for treating psoriasis. Paricalcitol (19-nor-1 alpha,25-dihydroxyvitamin D(2)) is a novel vitamin D analogue which has been developed for the prevention of secondary hyperparathyroidism in patients with chronic renal failure. OBJECTIVES: To investigate the efficacy and safety of 12 weeks' therapy with a once-daily application of paricalcitol ointment (15 microg g(-1)) in comparison with placebo ointment. METHODS: This pilot double-blinded self-controlled study was initiated in 11 patients with moderate plaque psoriasis. To characterize the biological effects further and to evaluate the efficacy of topical paricalcitol treatment in psoriasis, we have analysed immunohistochemically the expression of one of the markers for epidermal differentiation (transglutaminase K) in paricalcitol-treated skin as compared with placebo treatment. RESULTS: Treatment with paricalcitol was superior to placebo treatment beginning at week 1. The global severity score for erythema, plaque elevation and scaling was improved significantly more by paricalcitol ointment than by placebo (P < 0.001). Similar results were obtained for assessments of scaling, erythema and plaque elevation. No symptoms of local skin irritation were noted. Laboratory parameters including serum calcium, phosphorus, parathyroid hormone and urinary calcium/creatinine ratio did not reveal any changes of clinical relevance during treatment. The immunoreactivity of transglutaminase K changed after 12 weeks of paricalcitol treatment almost completely to the pattern characteristic for nonlesional psoriatic skin. CONCLUSIONS: Once-daily application of paricalcitol ointment was safe and effective for the treatment of plaque psoriasis. PMID- 15270891 TI - A randomized controlled clinical trial of topical photodynamic therapy with methyl aminolaevulinate in the treatment of actinic keratoses in transplant recipients. AB - BACKGROUND: Transplant recipients have an increased propensity to develop multiple actinic keratoses, which demonstrate an increased transformation rate into invasive squamous cell carcinoma. OBJECTIVE: To evaluate the efficacy and tolerability of topical photodynamic therapy with the new highly tumour-selective photosensitizer methyl aminolaevulinate vs. placebo in the treatment of actinic keratoses in transplant recipients. METHODS: Seventeen transplant recipients with a total number of 129 mild to moderate actinic keratoses were enrolled in a prospective, randomized, double-blind, placebo-controlled study. Two lesional areas within a patient were randomized for two consecutive treatments of topical photodynamic therapy 1 week apart using either methyl aminolaevulinate or placebo cream. Sites were illuminated with 75 J cm(-2) of visible light delivered at 80 mW cm(-2) by a noncoherent light source. Complete resolution and reduction in the number or size of actinic keratoses within the lesional area relative to the initial findings were evaluated at weeks 4, 8 and 16 after treatment. RESULTS: The lesional areas treated with methyl aminolaevulinate were clinically cleared in 13 of 17 patients at 16 weeks. A partial response was recorded in a further three. No reduction in the size or number of actinic keratoses was observed in one area treated with methyl aminolaevulinate and in all placebo-treated areas. Adverse events, such as erythema, oedema and crust formation, were mild to moderate, and treatment was well tolerated by all patients. CONCLUSION: Photodynamic therapy using methyl aminolaevulinate is a safe and effective treatment for actinic keratoses in transplant recipients. It may also reduce the risk of transformation of actinic keratoses to invasive, potentially fatal, squamous cell carcinoma. PMID- 15270892 TI - The juvenile variant of papular-purpuric gloves and socks syndrome and its association with viral infections. AB - BACKGROUND: Papular-purpuric gloves and socks syndrome (PPGSS) occurs mostly in adults and has been shown to be related to several possible viral infections. However, childhood-onset PPGSS seems to be not so rare as previously thought in our clinical experience. OBJECTIVES: To survey the general characteristics of childhood-onset PPGSS and to determine the possible association between this juvenile variant of PPGSS and various viral infections. PATIENTS AND METHODS: Thirty-three children with erythematopurpuric papular eruptions on the hands and/or feet were enrolled. Detailed history-taking and physical examination were performed on all of them. Blood samples were obtained from 25 patients about 1-5 weeks after the appearance of cutaneous eruptions to check complete blood counts, differential white blood cell counts, and IgM and IgG antibodies to parvovirus B19, cytomegalovirus (CMV), viral capsid antigen of Epstein-Barr virus (EBV) and measles. RESULTS: The median age of these 33 patients was 23 months. The mean duration of the skin eruption was 4.8 weeks (SD 2.7, 95% CI 3.9-5.0). Lymphocytosis was present in 13 patients (52%) while mild eosinophilia occurred in only three patients (12%). Five patients (20%) were positive for IgM antibodies against CMV and seven (28%) were positive for IgM antibodies against EBV. Only one patient (4%) was detected to have IgM antibodies against parvovirus B19. CONCLUSIONS: Childhood-onset PPGSS shows somewhat different clinical features from the adult type. It may represent a nonspecific manifestation of several viral infections, including CMV, EBV and parvovirus B19 infections. PMID- 15270893 TI - The effect of haematopoietic stem cell transplant on papules with 'pebbly' appearance in Hunter's syndrome. AB - BACKGROUND: Hunter's syndrome is associated with several cutaneous findings. For instance, papules with 'pebbly' appearance are a specific marker for the disease. However, it remains uncertain whether they disappear after haematopoietic stem cell transplant (HSCT). OBJECTIVES: To investigate the papules with 'pebbly' appearance before and after HSCT in infants with Hunter's syndrome, and to clarify the effect of HSCT on papules. PATIENTS: We observed five Japanese boys with Hunter's syndrome who had received HSCT at 4-11 years of age. RESULTS: The post-HSCT physical examinations revealed that papules disappeared completely within 35 days after the transplant with progressive reduction of cutaneous tightness in all the patients. Histochemical findings showed that papules contained a large amount of hyaluronic acid in the extracellular materials of the dermis and sulphated acid mucopolysaccharides in dermal fibroblasts before HSCT. CONCLUSIONS: These results suggest that papules with a 'pebbly' appearance fade away through the digestion of a large amount of hyaluronic acid in cutaneous tissues by normal tissue histiocytes or enzymes of donor origin at an early stage after HSCT. PMID- 15270894 TI - Detection of human papillomavirus type 56 DNA, belonging to a mucous high-risk group, in hair follicles in the genital area of a woman no longer suffering from viral warts. AB - BACKGROUND: Human papillomaviruses (HPVs) parasitize human epithelium, but it is not clear where they reside when they do not cause apparent infection. Hair follicles are important candidates as reservoirs. OBJECTIVES: A patient reported previously by us as having perianal warts caused mainly by HPV 56, demonstrated hair follicles in her genital area which bulged a little from the surface and appeared somewhat enlarged. We therefore examined whether DNA of HPV 56, a member of the mucous high-risk group, might be detectable in these structures. METHODS: We obtained plucked hairs and performed an examination by polymerase chain reaction and subsequent reverse-phase dot blot hybridization (PCR-RDBH) and in situ hybridization (ISH). RESULTS: Strong positive signals were obtained not only with PCR-RDBH but also with ISH. CONCLUSIONS: Hair follicles in the genital area might serve as reservoirs for HPVs belonging to the mucous high-risk group. PMID- 15270895 TI - Syringotropic cutaneous T-cell lymphoma: an immunophenotypic and genotypic study of five cases. AB - There is uncertainty about the exact nosological relationship between mycosis fungoides, follicular mucinosis, syringolymphoid hyperplasia with alopecia (SLHA) and syringotropic cutaneous T-cell lymphoma (CTCL). We report the clinical, histological, immunophenotypic and genotypic characteristics of a series of five patients (three men and two women) with syringotropic CTCL. We also review the 15 cases of SLHA previously reported in the literature. We conclude that syringotropic CTCL is a distinct clinicopathological variant of mycosis fungoides which may present on its own with characteristic punctate erythema or more commonly in association with folliculotropic lesions. Syringotropic CTCL is characterized histologically by infiltration of sweat glands by atypical lymphocytes in association with syringolymphoid hyperplasia. Cases of SLHA represent a syringotropic form of CTCL in association with follicular involvement, and such cases need to be investigated using T-cell receptor gene analysis of both skin and blood. Only limited conclusions on prognosis can be derived from our preliminary data. However, a review of the literature suggests that the prognosis does not differ significantly from other types of mycosis fungoides of equivalent stage. PMID- 15270896 TI - Bilateral Bowen's disease. AB - Multiple Bowen's disease may be difficult to differentiate from bowenoid papulosis because of its clinicopathological resemblance to bowenoid papulosis. We experienced a case of bilaterally and symmetrically developed multiple bowenoid lesions in a 71-year-old man previously diagnosed as having chronic lymphocytic lymphoma (CLL). Based on histological findings and the results of human papillomavirus examinations, we finally diagnosed this case as bilateral Bowen's disease. We speculate that the underlying immunosuppressive state due to CLL may have been associated with onset of the disease. We report the unique clinical picture, the differential diagnosis and the aetiology. PMID- 15270897 TI - Infantile perianal pyramidal protrusion with hard stool history. PMID- 15270898 TI - Melanocytic naevus of the palm resembling callus. PMID- 15270899 TI - Naproxen-induced generalized bullous fixed drug eruption. PMID- 15270900 TI - Immunohistochemical analyses of p63 expression in normal human skin. PMID- 15270902 TI - Plasma cell leukaemia cutis preferentially localized to recent puncture sites. PMID- 15270901 TI - Imatinib mesylate inhibits the growth of metastatic lung lesions in a patient with dermatofibrosarcoma protuberans. PMID- 15270903 TI - Acneiform eruption induced by epidermal growth factor receptor inhibitors in patients with solid tumours. PMID- 15270904 TI - Exfoliative cheilitis successfully treated with topical tacrolimus. PMID- 15270905 TI - Acquired pseudoxanthoma elasticum of the elbow flexures. PMID- 15270906 TI - Streptococcal infection may make psoriasis worse but do antibiotics help? PMID- 15270907 TI - The syringe skin hook: the history and evolution of the improvised skin hook. PMID- 15270908 TI - Granuloma annulare restricted to Becker's naevus. PMID- 15270910 TI - Systemic steroids in the treatment of alopecia areata. PMID- 15270911 TI - Susceptibility to human herpesvirus-8 infection in a healthy population from Sardinia is not directly correlated with the expression of HLA-DR alleles. PMID- 15270912 TI - The histopathology of envenomation by Japanese viper bite. PMID- 15270913 TI - Primary cutaneous B-cell lymphoma mimicking pyoderma gangrenosum: first-line treatment with rituximab. PMID- 15270914 TI - Lichen planus pemphigoides and multiple keratoacanthomas associated with colon adenocarcinoma. PMID- 15270915 TI - Atypical varicella mimicking hand-foot-mouth disease in an adult patient with malignant lymphoma during chemotherapy. PMID- 15270920 TI - Teenage pregnancy: a problem or what? PMID- 15270921 TI - The impact of guidelines on mild hypertension in pregnancy: time series analysis. AB - OBJECTIVE: To determine the impact of a national strategy to promote implementation of a guideline on the management of mild, non-proteinuric hypertension in pregnancy. DESIGN: Simple, interrupted time series analysis. SETTING: Four maternity units in Scotland. POPULATION: Women delivering a live or stillborn baby. METHODS: Dissemination of the guideline under the auspices of a national clinical effectiveness programme, supported by a national launch meeting and feedback from a survey of obstetricians highlighting aspects of care that could be improved. MAIN OUTCOME MEASURES: Appropriateness of initial investigation and subsequent clinical management, and costs of guideline development and implementation activities. DATA COLLECTION: Twenty-four months pre-intervention and 12 months post-intervention data were abstracted from a random sample of case notes. RESULTS: Initial investigation was consistent with recommendations for 59.9% out of 1263 women and subsequent clinical management for 67.6% out of 1081 in whom a diagnosis could be made from available data. There were no significant changes in the appropriateness of initial investigation (10.6%; 95% confidence interval [CI] -0.1% to 19.3%; decreasing by 1.2% per month post-implementation, 95% CI -2.5% to 0.1%) or clinical management (-0.3%; 95% CI 8.7% to 11.2%). Guideline development and implementation cost an estimated pound 2784 per maternity unit in Scotland. CONCLUSIONS: Clinical care of mild hypertension in pregnancy remains highly inconsistent. The lack of the intervention effect may be related to the complexity of the guideline recommendations and the nature of the implementation strategy. PMID- 15270922 TI - A case-controlled study comparing clinical course and outcomes of pregnant and non-pregnant women with severe acute respiratory syndrome. AB - OBJECTIVE: To compare the clinical courses and outcomes of pregnant severe acute respiratory syndrome (SARS) patients and non-pregnant SARS patients. DESIGN: A case-control study. SETTING: Tertiary Hospital for Infectious Disease. Sample Ten pregnant and 40 non-pregnant female patients infected with SARS. METHODS: Clinical course and outcomes of pregnant SARS patients were compared with a group of non-pregnant SARS patient. Cases and controls were matched with respect to sex, age, timing of contracting SARS, health care workers status and underlying illness. MAIN OUTCOME MEASURES: The incidence of intensive care unit admission, intubation, medical complications and death rate. RESULTS: Pregnancy had no discernible impact on clinical symptoms and presentation delay. Four out of the 10 pregnant patients, nevertheless, required endotracheal intubation and six were admitted to the intensive care unit (ICU), as compared with 12.5% intubation rate (P= 0.065) and 17.5% ICU admission rate (P= 0.012) in the non-pregnant group. More pregnant SARS patients developed renal failure (P= 0.006) and disseminated intravascular coagulopathy (P= 0.006), as compared with non-pregnant SARS group. There were three deaths in the pregnant group, whereas there was no death in the non-pregnant control group (P= 0.006). CONCLUSION: Pregnant women with SARS experience a worse clinical course and poorer outcomes compared with non-pregnant women. PMID- 15270923 TI - Women and health care professionals' preferences for Down's Syndrome screening tests: a conjoint analysis study. AB - OBJECTIVE: To describe and compare women and health care professionals' preferences for Down's Syndrome screening tests with different test characteristics. DESIGN: Cross sectional questionnaire based conjoint analysis study. SETTING: London teaching hospital. SAMPLE: 291/383 women in their first or second trimester of pregnancy and 98/122 health care professionals (41 obstetricians, senior house officers and above and 57 qualified midwives) providing care at the same hospital. METHODS: Women completed a questionnaire while attending a clinic visit for a dating scan or a routine 20-week anomaly scan. Health care professionals completed a postal questionnaire. MAIN OUTCOME MEASURES: The relative values participants attach to Down's Syndrome screening test attributes: time of test, detection rate and risk of miscarriage of a baby unaffected by Down's Syndrome as a result of subsequent diagnostic tests. RESULTS: Pregnant women and health care professionals shared broadly similar relative values regarding the importance of safe tests, conducted early and with high detection rates. When asked to choose between different Down's Syndrome screening tests, health care professionals valued earlier tests more highly than did women. CONCLUSIONS: While pregnant women and health care professionals share similar relative values regarding optimal prenatal tests, health care professionals place a higher value on earlier tests. This may result in screening policies that overweight timing in the selection of a test to the relative neglect of tests associated with lower miscarriage rates and higher detection rates but conducted later in pregnancy. PMID- 15270924 TI - Thrombophilia and stillbirth: possible connection by intrauterine growth restriction. AB - OBJECTIVE: To define the association between thrombophilia and unexplained stillbirth. DESIGN: A case-control study. SETTING: Obstetric department in a university affiliated hospital (Ha'Emek Medical Center, Afula). POPULATION: A total of 53 women who delivered stillborns between March 1998 and June 2002 and 59 women with unremarkable obstetric history who delivered at the same period. METHODS: Presence of genetic and acquired markers of thrombophilia was investigated. MAIN OUTCOME MEASURE: Presence or absence of thrombophilia. RESULTS: Thrombophilia was found in 34% of the women who delivered stillborns and in 20% of the 59 women with normal pregnancies (non-significant). However, significantly higher prevalence of thrombophilia (73%) was found in women who delivered small for gestational age stillborns compared with women who delivered normal birthweight stillborns (73%vs 18.4%, P < 0.0001). CONCLUSIONS: There is no association between thrombophilia and stillbirth, overall. However, there is a clear association between thrombophilia and stillbirth of extremely growth restricted infants. PMID- 15270925 TI - Comparison of ultrasound and magnetic resonance imaging in 100 singleton pregnancies with suspected brain abnormalities. AB - OBJECTIVE: To compare the diagnostic accuracy of the current reference standard ultrasound with in utero magnetic resonance imaging, in a selected group of patients. DESIGN: Prospective study. SETTING: Five fetal maternal tertiary referral centres and an academic radiology unit. SAMPLE: One hundred cases of fetuses with central nervous system abnormalities where there has been diagnostic difficulties on ultrasound. In 48 cases the women were less than 24 weeks of gestation and in 52 cases later in pregnancy. METHODS: All women were imaged on a 1.5 T clinical system using a single shot fast spin echo technique. The results of antenatal ultrasound and in utero magnetic resonance were compared. MAIN OUTCOME MEASURES: The definitive diagnosis was made either at autopsy or by postmortem magnetic resonance imaging, in cases that went to termination of pregnancy, or a combination of postnatal imaging and clinical follow up in the others. RESULTS: In 52 of cases, ultrasound and magnetic resonance gave identical results and in a further 12, magnetic resonance provided extra information that was judged not to have had direct effects on management. In 35 of cases, magnetic resonance either changed the diagnosis (29) or gave extra information that could have altered management (6). In 11 of the 30 cases where magnetic resonance changed the diagnosis, the brain was described as normal on magnetic resonance. CONCLUSIONS: In utero magnetic resonance imaging is a powerful tool in investigating fetal brain abnormalities. Our results suggest that in selected cases of brain abnormalities, detected by ultrasound, antenatal magnetic resonance may provide additional, clinically useful information that may alter management. PMID- 15270926 TI - Premature death among teenage mothers. AB - OBJECTIVE: Some data suggest an association between teenage childbearing and premature death. Whether this possible increase in risk is associated with social circumstances before or after childbirth is not known. We studied premature death in relation to age at first birth, social background and social situation after first birth. DESIGN: Population-based cohort study. SETTING: Women born in Sweden registered in the 1985 Swedish Population Census. POPULATION: Swedish women born 1950-1964 who had their first infant before the age of 30 years (N= 460,434). METHODS: Information on the women's social background and social situation after first birth was obtained from Population Censuses. The women were followed up with regard to cause of death from December 1, 1990 to December 31, 1995. Mortality rate ratios and 95% confidence intervals (CI) were calculated. MAIN OUTCOME MEASURES: Mortality rates by cause of death. RESULTS: Independent of socio-economic background, teenage mothers faced an increased risk of premature death later in life compared with older mothers (rate ratio 1.6, 95% CI 1.4-1.9). The increased risk was most evident for deaths from cervical cancer, lung cancer, ischaemic heart disease, suicide, inflicted violence and alcohol-related diseases. Some, but not all, of these increases in risk were associated with the poorer social position of teenagers mothers. CONCLUSIONS: Teenage mothers, independent of socio-economic background, face an increased risk of premature death. Strategies to reduce teenage childbearing are likely to contribute to improved maternal and infant health. PMID- 15270927 TI - Home-based care after a shortened hospital stay versus hospital-based care postpartum: an economic evaluation. AB - OBJECTIVES: To compare the cost effectiveness of early postnatal discharge and home midwifery support with a traditional postnatal hospital stay. DESIGN: Cost minimisation analysis within a pragmatic randomised controlled trial. SETTING: The University Hospital of Geneva and its catchment area. POPULATION: Four hundred and fifty-nine deliveries of a single infant at term following an uncomplicated pregnancy. METHODS: Prospective economic evaluation alongside a randomised controlled trial in which women were allocated to either early postnatal discharge combined with home midwifery support (n= 228) or a traditional postnatal hospital stay (n= 231). MAIN OUTCOME MEASURES: Costs (Swiss francs, 2000 prices) to the health service, social services, patients, carers and society accrued between delivery and 28 days postpartum. RESULTS: Clinical and psychosocial outcomes were similar in the two trial arms. Early postnatal discharge combined with home midwifery support resulted in a significant reduction in postnatal hospital care costs (bootstrap mean difference 1524 francs, 95% confidence interval [CI] 675 to 2403) and a significant increase in community care costs (bootstrap mean difference 295 francs, 95% CI 245 to 343). There were no significant differences in average hospital readmission, hospital outpatient care, direct non-medical and indirect costs between the two trial groups. Overall, early postnatal discharge combined with home midwifery support resulted in a significant cost saving of 1221 francs per mother-infant dyad (bootstrap mean difference 1209 francs, 95% CI 202 to 2155). This finding remained relatively robust following variations in the values of key economic parameters performed as part of a comprehensive sensitivity analysis. CONCLUSIONS: A policy of early postnatal discharge combined with home midwifery support exhibits weak economic dominance over traditional postnatal care, that is, it significantly reduces costs without compromising the health and wellbeing of the mother and infant. PMID- 15270928 TI - Home-based versus hospital-based postnatal care: a randomised trial. AB - OBJECTIVE: To compare a shortened hospital stay with midwife visits at home to usual hospital care after delivery. DESIGN: Randomised controlled trial. SETTING: Maternity unit of a Swiss teaching hospital. POPULATION: Four hundred and fifty nine women with a single uncomplicated pregnancy at low risk of caesarean section. METHODS: Women were randomised to either home-based (n= 228) or hospital based postnatal care (n= 231). Home-based postnatal care consisted of early discharge from hospital (24 to 48 hours after delivery) and home visits by a midwife; women in the hospital-based care group were hospitalised for four to five days. MAIN OUTCOME MEASURES: Breastfeeding 28 days postpartum, women's views of their care and readmission to hospital. RESULTS: Women in the home-based care group had shorter hospital stays (65 vs 106 hours, P < 0.001) and more midwife visits (4.8 vs 1.7, P < 0.001) than women in the hospital-based care group. Prevalence of breastfeeding at 28 days was similar between the groups (90%vs 87%, P= 0.30), but women in the home-based care group reported fewer problems with breastfeeding and greater satisfaction with the help received. There were no differences in satisfaction with care, women's hospital readmissions, postnatal depression scores and health status scores. A higher percentage of neonates in the home-based care group were readmitted to hospital during the first six months (12%vs 4.8%, P= 0.004). CONCLUSIONS: In low risk pregnancies, early discharge from hospital and midwife visits at home after delivery is an acceptable alternative to a longer duration of care in hospital. Mothers' preferences and economic considerations should be taken into account when choosing a policy of postnatal care. PMID- 15270929 TI - Is home-based administration of prostaglandin safe and feasible for medical abortion? Results from a multisite study in Vietnam. AB - OBJECTIVES: To study the efficacy and acceptability of a simplified medical abortion regimen in Vietnam. DESIGN: Open-label study. SETTING: One peri-urban and three urban hospitals and four urban maternal-child health family planning clinics located in Northern, Central and Southern Vietnam. SAMPLE: A total of 1601 women seeking abortion services from January 2001 to December 2001. METHODS: Consenting women presenting for abortion services with gestations less than 56 days LMP who met the inclusion criteria were given 200 mg mifepristone and offered the choice of either home or clinic administration of 400 microg oral misoprostol two days later. MAIN OUTCOME MEASURE: Complete abortion rate of 89.2% (n= 1395), with 1.5% (n= 24) of the women lost to follow up. The majority of women (>90%) reported that their medical abortion experience was either 'very satisfactory' or 'satisfactory'. RESULTS: There was a strong preference for home administration of misoprostol, with more than four-fifths of the study population selecting to administer the prostaglandin at home. Location of misoprostol administration did not affect efficacy rate. Regardless of location selected, women expressed a high degree of satisfaction with the medical abortion experience. CONCLUSIONS: Medical abortion with the option of home administration of misoprostol is safe and feasible for introduction into the Vietnamese healthcare system. PMID- 15270930 TI - Emergency contraceptive pills in Sweden: evaluation of an information campaign. AB - OBJECTIVE: To evaluate a community-based intervention regarding emergency contraceptive pills, including a mass media campaign and information to women visiting family planning clinics. DESIGN: Quasi-experimental. SETTING: Two counties in Sweden. POPULATION: Eight hundred randomly selected women aged 16-30 years, 400 women in the intervention group and 400 in a comparison group. METHODS: Postal questionnaires before (2002) and after (2003) the intervention. MAIN OUTCOME MEASURES: Exposure to the intervention, knowledge, attitudes, practices and intention to use emergency contraceptive pills. RESULTS: Before the intervention, the response rate was 71% (n= 564); after the intervention, the corresponding figure was 83% (n= 467); overall response rate 58%. Two-thirds (64%) of the targeted women had noticed the information campaign. One out of six who had visited a family planning clinic during the intervention year recalled being given information about emergency contraceptive pills. Specific knowledge and attitudes improved over time in both groups, but there was no difference in change between the groups. The proportion of women who had used emergency contraceptive pills increased from 27% to 31% over time. Intention to use emergency contraceptive pills in case of need was reported by 74% of the women and remained stable over time, but logistic regression showed that information during the previous year contributed to willingness to use the method in the intervention group. CONCLUSIONS: Knowledge, attitudes and practices about emergency contraceptive pills increased in both groups. Emergency contraceptive pills is gradually becoming a more widely known, accepted and used contraceptive method in Sweden, a trend that may have limited the impact of the intervention. PMID- 15270931 TI - How long should urinary bladder catheterisation be continued after vaginal prolapse surgery? A randomised controlled trial comparing short term versus long term catheterisation after vaginal prolapse surgery. AB - OBJECTIVE: To determine whether prolonged urinary bladder catheterisation after vaginal prolapse surgery is advantageous. DESIGN: Randomised controlled trial. SETTING: A large training hospital in the Netherlands. POPULATION: Patients undergoing anterior colporrhaphy. METHODS: One hundred patients were included. Patients were randomised into two groups. In one group (n= 50), a transurethral catheter was in place for four days post-operatively and removed on the fifth post-operative day. In the other group (n= 50), catheterisation was not prolonged and the catheter was removed the morning after surgery. Residual volumes after removal of the catheter were measured by ultrasound scanning. Where residual volumes of >200 mL were found the patient was recatheterised for three more days. Urinary cultures were taken before removal of the catheter. Six patients were excluded: four in the standard prolonged catheterisation group and two in the not prolonged catheterisation group. MAIN OUTCOME MEASURES: Need for recatheterisation, urinary tract infection, mean duration of catheterisation and hospital stay. RESULTS: Residual volumes exceeding 200 mL and need for recatheterisation occurred in 9% in the standard prolonged catheterisation group versus 40% of patients in the not prolonged catheterisation group (OR 0.15, 95% CI 0.045-0.47). Positive urine cultures were found in 40% of cases in the standard prolonged catheterisation group compared with 4% in the not prolonged catheterisation group (OR 15, 95% CI 3.2-68.6). Mean duration of catheterisation was 5.3 days in the standard prolonged catheterisation group and 2.3 days in the not prolonged catheterisation group (P < 0.001). Mean duration of hospitalisation was 7 days in the standard prolonged catheterisation group and 5.7 days in the not prolonged group (P < 0.001). CONCLUSION: The disadvantages of prolonged catheterisation outweigh the advantages, therefore, removal of the catheter on the morning after surgery may be preferable and longer term catheterisation should only be undertaken where there are specific indications. PMID- 15270932 TI - Transvaginal repair of anterior and posterior compartment prolapse with Atrium polypropylene mesh. AB - OBJECTIVE: To determine the efficacy and safety of a new technique using Atrium polypropylene mesh (Atrium, Hudson, New Hampshire, USA) as an overlay graft for repair of large or recurrent anterior and posterior compartment prolapse. DESIGN: A retrospective review of women who had vaginal prolapse surgery with Atrium mesh reinforcement. SETTING: Tertiary referral urogynaecology unit in Australia. POPULATION: Forty-seven women where mesh was placed under the bladder base with lateral extensions onto the pelvic sidewall, 33 women where a Y-shaped mesh was placed from the sacrospinous ligaments to the perineal body and 17 women who had mesh placement in both compartments. METHODS: Women were assessed by site specific vaginal examination pre-operatively and post-operatively at six weeks, six months and two years. MAIN OUTCOME MEASURES: All complications. Rate of recurrent prolapse assessed by the Baden-Walker halfway classification system. RESULTS: Mean follow up was 29 months (range 6 to 52). Four of 64 women with anterior mesh placement (6%) developed a grade 2 asymptomatic cystocele. Five women (5%) required further surgery for recurrent prolapse at a non-mesh site. Erosion occurred in nine women (9%). Three healed after intravaginal oestrogen cream, five after excision of exposed mesh and vaginal closure and one woman also had surgical closure of a rectovaginal fistula. The risk of mesh erosion decreased over the study period. Urinary, coital and bowel symptoms were significantly improved following surgery. CONCLUSIONS: This technique shows promise in correcting pelvic organ prolapse. Vaginal mesh erosion is the most common complication and is related to surgical experience. PMID- 15270933 TI - The tension-free vaginal tape in older women. AB - OBJECTIVE: To evaluate peri-operative morbidity, continence outcome and patient satisfaction in older women (>/=65 years) compared with younger women undergoing tension-free vaginal tape. DESIGN: Case controlled study. SETTING: Tertiary Urogynaecology Unit. SAMPLE: Women undergoing tension-free vaginal tape for urodynamic stress incontinence between July 1999 and July 2002 were included. Those with detrusor overactivity, voiding difficulty at urodynamics or requiring concomitant prolapse surgery were excluded. METHODS: Older women were case matched to a younger cohort for BMI, parity, mode of anaesthesia and whether it was a primary or secondary continence procedure. MAIN OUTCOME MEASURES: Operative morbidity and continence outcome were assessed at six weeks. After a minimum six months follow up, patient satisfaction and continence outcome were assessed using the Genitourinary Treatment Satisfaction Score (GUTSS). RESULTS: The median hospital stay was one day and overall urinary tract infection rate was similar in both groups. Post-operative voiding difficulty rates were 3% in older versus 15% in younger women (P= 0.09). At six weeks, 65% of older versus 79% of younger women were dry (P= 0.2). At a median of 12 months, 15 (45%) of older versus 24 (73%) of younger women had no urinary symptoms (P= 0.05). Median GUTSS scores for satisfaction with continence outcome were lower for older 90% compared with 100% in younger women (P= 0.003). CONCLUSIONS: Tension-free vaginal tape is an effective continence intervention in older women but has a lower continence satisfaction rate compared with younger women. PMID- 15270934 TI - Comparison of HPV test versus conventional and automation-assisted Pap screening as potential screening tools for preventing cervical cancer. AB - OBJECTIVE: To evaluate new techniques in primary cervical cancer screening programmes. DESIGN: Cross sectional pilot study. SETTING: Department of Obstetrics and Gynaecology, Helsinki University Hospital. POPULATION: Consecutive 2032 human papillomavirus (HPV) DNA and Pap smear samples were taken. Histological diagnoses were obtained from 460 patients. METHODS: We compared the validity of the high risk (HR) HPV DNA detection test to automation-assisted and conventional Pap smear screening. MAIN OUTCOME MEASURES: Specificity and sensitivity of screening methods. RESULTS: Twenty-three percent of women were HPV positive. Forty-five of 46 had high grade lesions and cancers were HR HPV DNA positive, whereas 72/93 of low grade and more severe lesions were HR HPV DNA positive. When histologically verified high grade lesions were observed, the relative sensitivity of HR Hybrid Capture 2 (HR HC2) test was 98% compared with conventional Pap smear and Papnet tests, which performed 54%versus 58%, 83%versus 86% and 93%versus 98% relative sensitivity respectively, using cytological diagnoses HSIL (high grade squamous intraepithelial lesion), LSIL (low grade squamous intraepithelial lesion) or ASCUS (atypical squamous cells of undetermined significance) as the cutoff. The specificity of HC2 test (77-79%) was comparable with the ASCUS+ (ASCUS and more severe) cytology (68-79%), but lower when compared with LSIL+ (91-95%) or especially HSIL+ (97-99%) Pap smear results. CONCLUSION: Pap smear, as a screening test, is very different from HPV DNA detecting test HR HC 2. If cutoff LSIL or more severe lesions is used, primary Pap smear is clearly more specific than HR HC2, but markedly less sensitive. Due to high relative sensitivity of the HPV, only very few histologically confirmed high grade lesions would be detected among HPV negatives using simultaneous cytology. On the other hand, using HPV DNA test alone would lead to multifold amounts of referrals for colposcopy. A posterior Pap smear assessment among HPV positives might be helpful in increasing sensitivity and specificity of screening and defining those who need an immediate referral or treatment. We plan to incorporate primary HR HPV DNA test with posterior Pap smear reading of HPV positives into our ongoing randomised prospective multiarm trial evaluating new techniques in organised screening for cervical cancer in Finland. PMID- 15270935 TI - Preterm birth and maternal country of birth in a French district with a multiethnic population. AB - OBJECTIVES: This analysis explores the association between preterm birth and maternal country of birth in a French district with a multiethnic population. DESIGN: Prospective observational study. SETTING: District of Seine-Saint-Denis in France POPULATION: 48,746 singleton live births from a population-based birth register between October 1998 and December 2000. METHODS: We compare preterm birth rates by mother's country of birth controlling for demographic and obstetric factors as well as insurance coverage and timing of initiation of antenatal care. MAIN OUTCOME MEASURES: Overall preterm birth rates and preterm birth rates by timing of delivery (<33 weeks versus 33-36 weeks of gestation), mode of onset (spontaneous or indicated preterm birth) and the presence of hypertension in pregnancy. RESULTS: Women born in Northern Africa, Southern Europe and South/East Asia did not have higher preterm birth rates than women born in continental France. Rates were significantly higher for women born in the overseas French districts in the Caribbean and Indian Ocean and Sub-Saharan Africa. Excess risk was greatest for early preterm births, medically indicated births and preterm births associated with hypertension. CONCLUSIONS: Patterns of preterm birth with relation to timing, mode of onset and medical complications among of Afro-Caribbean origin should be confirmed in future research. PMID- 15270936 TI - Dramatic reduction in triplet and higher order births in England and Wales. AB - The proportion of multiple births has increased markedly since 1980 in England and Wales. A major contribution to this trend is thought to be the widespread introduction of assisted reproductive technologies. Despite a continuing (but slower) upward trend in twin maternities, analysis of recent data shows that the rate of triplet and higher order births in England and Wales has declined by one quarter since 1998. This probably reflects both voluntary and statutory regulation of treatment regimes. This downward trend will help alleviate the high burden of perinatal morbidity and mortality associated with multiple births. PMID- 15270937 TI - Definition of mild, moderate and severe incontinence on the 24-hour pad test. AB - The values for 'mild', 'moderate' and 'severe' urinary incontinence have not been determined for the 24-hour pad test. To define these values, a prospective observational study was performed on 110 women with the primary symptom of urinary incontinence. Consenting women performed two 1-hour pad tests one week apart, and seven 24-hour pad tests for seven consecutive days. The 1-hour pad test definitions for mild, moderate and severe were translated to centiles, and used to categorise the 24-hour test values. This revealed that the range for 'mild incontinence' was between 1.3 and 20 g, 'moderate incontinence' ranged from 21 to 74 g, and 'severe incontinence' was defined as 75 g or more in 24 hours. Severity of leakage was analysed in relation to urodynamic diagnosis, age, parity and pelvic floor muscle strength. Increasing severity was associated with increasing age and parity. Women with detrusor overactivity were most likely to have severe leakage. In conclusion, this study defines the three grades of severity for the 24-hour pad test, which may help to guide patients' choice between conservative and surgical treatment and is useful for stratified randomisation of controlled trial participants. PMID- 15270938 TI - Ketanserin in pre-eclamptic patients: transplacental transmission and disposition in neonates. AB - The aim of this prospective, observational study was to assess transplacental transmission of ketanserin, an antihypertensive drug used in pre-eclampsia, and to determine disposition and effects in the neonate after maternal ketanserin use. In 22 pregnant women with severe pre-eclampsia, admitted to the antenatal ward in the period 1999-2001, the ratio of drug levels in the umbilical cord to drug levels in maternal blood just before delivery was used as an indicator of placental transmission. Disposition of ketanserin was assessed using neonatal plasma concentrations of ketanserin in eight neonates after birth. A median placental transmission was found in the pre-eclamptic women of 0.95 (0.612-1.24) for ketanserin and for its metabolite, ketanserinol, of 0.60 (0.5-0.77). Pharmacologically relevant concentrations of ketanserin were found in the neonate after delivery. The elimination half-life of ketanserin in the neonate varied between 12.7 and 43.7 hours (median 19.3 hours) and of ketanserinol between 13.8 and 34.4 hours (median 18.7 hours). Despite the high placental transmission and disposition in the neonate, no apparent adverse effects in the neonates could be detected. In conclusion, a high placental transmission of ketanserin and its metabolite ketanserinol occurred after maternal treatment of pre-eclampsia with ketanserin and pharmacologically active concentrations of ketanserin are found in the neonate for a prolonged period after delivery. PMID- 15270939 TI - Comparison of the arrhythmogenic effects of tauro- and glycoconjugates of cholic acid in an in vitro study of rat cardiomyocytes. AB - Obstetric cholestasis is associated with intrauterine death. In obstetric cholestasis, primary bile acids are more commonly conjugated with taurine than glycine, while glycoconjugates predominate in normal pregnancy. Using an in vitro model of rat cardiomyocytes, we compared the effect of tauro- and glycoconjugated cholate on cardiomyocyte rhythm, contraction amplitude and network integrity. We demonstrated that taurocholate had a more marked effect on all of these parameters, and the effects of the glycoconjugates were fully reversible while those of tauroconjugates were not. The increased proportion of tauroconjugated bile acids in obstetric cholestasis may contribute to the aetiology of the intrauterine death associated with the condition. PMID- 15270940 TI - Women's experiences of student presence in consultations for problematic uterine bleeding. AB - Research suggests that although a high proportion of patients accept the presence of students in gynaecological consultations, issues of consent, privacy and comfort are important. This study considers women's views on the impact of student presence on communication in the consultation. Our research suggests that student presence may distort the flow of communication in the gynaecological consultation. There are implications for both patient satisfaction and clinician training. If students are introduced into the consultation, clinical tutors should take special care to maintain dedicated communication with the patient. PMID- 15270941 TI - National survey for intrapartum and postpartum bladder care: assessing the need for guidelines. AB - Variation in the practice of intrapartum and postpartum bladder care reported by 189 maternity units in England and Wales hospitals was evaluated by analysing the data obtained from a postal questionnaire completed by labour ward managers or heads of midwifery. The survey revealed that there was no consensus of opinion about the diagnostic criteria for postpartum urinary retention and therefore the optimum management for voiding dysfunction remains controversial. In spite of the increasing awareness of the risk management issues involved, the majority of the units were found to be non-compliant with the limited RCOG recommendations currently available. Although further research is needed to develop evidence based guidelines, all units should be timing and measuring the voided volume and ideally checking the first post-void residual volume to ensure that retention does not go unrecognised. PMID- 15270942 TI - A national survey of senior trainees surgical experience in hysterectomy and attitudes to the place of vaginal hysterectomy. AB - We set out to determine the current status of training in vaginal hysterectomy in the UK. In total, 255 year 4 or 5 'Calman' trainees were identified and sent an anonymous questionnaire assessing surgical experience, quality of training and attitudes towards vaginal hysterectomy. Our results demonstrate that senior trainees' experience in vaginal as opposed to abdominal hysterectomy is relatively poor. Despite this, trainees believed that the majority of hysterectomies should be done vaginally, and only a minority, laparoscopically. PMID- 15270943 TI - Arterial embolisation for persistent primary postpartum haemorrhage: before or after hysterectomy? AB - Arterial embolisation is a recognised treatment for postpartum haemorrhage (PPH). In this retrospective study, we evaluate its use in the management of persistent PPH. Records of all births during a 54 month period at a university hospital were analysed. Two sub-groups were identified. Group I (n= 5), underwent embolisation after hysterectomy and Group II (n= 4), had embolisation as a first-line theraphy without hysterectomy. Of 20,215 births, there were 636 cases of PPH (3.1%). Nine required embolisation to control bleeding (1.4%). Group I needed multiple surgical procedures, had a larger pre- and post-operative blood requirement (12 100, median 22 units, vs. 6-12, median 8.5 units), longer embolisation (33-93, median 54 minutes, vs 20-66, 47 minutes) with a larger radiation exposure (5194 9067, median 6301 dGy, vs. 269-3862, median 950 dGy), a longer intensive care stay (3-7, median four days vs. 0-1.5, median one day), and more complications, when compared with Group II. Three of four women from Group II resumed menstrual function. Embolisation prior to hysterectomy may be preferable to embolisation after hysterectomy for the control of PPH. PMID- 15270944 TI - Persistent maternal viremia after varicella infection during pregnancy as a possible cause of false positive prenatal diagnosis of fetal infection on amniotic fluid. PMID- 15270946 TI - Horner's syndrome postpartum. PMID- 15270950 TI - Evaluation of chitosan-alginate-hyaluronate complexes modified by an RGD containing protein as tissue-engineering scaffolds for cartilage regeneration. AB - In this study, a series of natural biodegradable materials in the form of chitosan (C)-alginate (A)-hyaluronate (H) complexes are evaluated as tissue engineering scaffolds. The weight ratio of C/A is 1 : 1 or 1 : 2. Sodium hyaluronate is mixed in 2%. The complexes can be cast into films or fabricated as scaffolds. Their surface can be further modified by an Arg-Gly-Asp (RGD) containing protein, a cellulose-binding domain-RGD (R). Cytocompatibility tests of the films are conducted using immortalized rat chondrocyte (IRC) as well as primary articular chondrocytes harvested from rabbits. The neocartilage formation in cell-seeded scaffolds is examined in vitro as well as in rabbits, where the scaffolds are implanted into the defect-containing joints. The results from cytocompatibility tests demonstrate that R enhances cell attachment and proliferation on C-A and C-A-H complex films. Complex C1A1HR (C : A = 1 : 1 with H and R) has better performance than the other formulation. Cells retain their spherical morphology on all C-A and C-A-H complexes. The in vitro evaluation of the seeded scaffolds indicates that the C1A1HR complex is the most appropriate for 3-D culture, manifested by the better cell growth as well as higher glycosaminoglycan and collagen contents. When the chondrocyte scaffolds are implanted into rabbit knee cartilage defects, partial repair is observed after 1 month in C1A1HR as well as in C1A1 (C : A = 1 : 1 without H and R) scaffolds. The defects are completely repaired in 6 months when C1A1HR constructs are implanted. It is concluded that C1A1HR is a potential tissue-engineering scaffold for cartilage regeneration. PMID- 15270951 TI - Power spectral analysis of the heart rate variability of goat fetuses during extrauterine incubation. AB - Our aim is to determine the relationship between heart rate and behavioral states of a fetal goat using power spectral analysis. Electrocardiograms, electrocortical activity, and fetal breathing movements are recorded from 7 goat fetuses during extrauterine incubation. The heart rate power spectrum is classified into very low, low, and high frequency bands, and behavioral states are classified into low-voltage electrocortical activity with fetal breathing movements (LVB), low-voltage electrocortical activity without fetal breathing movements (LVN), and high-voltage electrocortical activity (HVN). There is a significant difference in total power spectral density in the high frequency band between LVN and HVN, and LVN and LVB. The relationship between each fetal behavioral state is assessed by power spectral analysis. PMID- 15270952 TI - Development of a suction detection system for axial blood pumps. AB - Axial flow blood pumps for cardiac assistance have proven their clinical viability and benefit in recent years. However, the clinical systems to date have no direct mechanism to decrease pump speed when adequate supply is not available. This may lead to ventricular collapse or increase the probability of hemolysis and thrombotic risks. Based on various experiences with left ventricular assist device (LVAD) patients in various states of recovery, at implant, in the intensive care unit, in the standard ward, and during physical exercise, 11 different algorithms were developed for the automatic detection of ventricular suction. These detection algorithms analyze the flow pattern for the presence of distinct suction indicators. For selection and optimization of the algorithms, 1000 records from approximately 100 patients were collected. Each record contains 5 s of pump flow, current, and arterial pressure. Three experts classified these records in terms of suction probability and other abnormalities. The optimization was developed in Matlab, capable of solving a fifth-dimensional optimization problem with 256 different algorithm combinations. The optimization resulted in a set of 6 algorithms, each with specific thresholds. The system detects 100% of the known suction events with 0.28% of false-positive interpretations. If tuned to avoid any false-positive detection, 90.7% of the certain events would be detected. A strategy for the development of a robust suction detection system for axial blood pumps was found. This system will be integrated into an automatic pump speed control system to provide adequate perfusion for the LVAD recipient, without excessive unloading of the ventricle. PMID- 15270953 TI - New aspects on the role of blood pressure and arterial stiffness in mechanical assistance by intra-aortic balloon pump: in-vitro data and their application in clinical practice. AB - Despite the well-known beneficial effects of the intra-aortic balloon pump (IABP) generally, there are still some clinical conditions accompanied by IABP ineffectiveness. The aim of this study was the investigation of the independent effects of arterial stiffness and blood pressure on acute IABP effectiveness. For this purpose, a mock circulatory system and 20 patients with cardiogenic shock due to acute myocardial infarction, were employed. It was shown that IABP acute efficiency was determined primarily by arterial compliance (AC) rather than blood pressure alone. IABP induced low hemodynamic effects in patients with systolic blood pressure > 80 mm Hg but with increased AC, whereas IABP resulted in greater hemodynamic effectiveness in cases with systolic pressure < 70 mm Hg but lower AC. The present study provides evidence concerning the hemodynamic conditions, which might lead to optimization of IABP or to the prediction of its acute hemodynamic performance, based on both measurements of AC and blood pressure. PMID- 15270954 TI - Right versus left internal jugular vein catheterization for hemodialysis: complications and impact on ipsilateral access creation. AB - This study investigated whether the anatomical differences between right and left internal jugular vein catheterization (RJVC and LJVC) would lead to differences in the frequency of complications, in particular, central vein occlusion (CVO). A group of 479 jugular vein catheterizations, 403 RJVC and 77 LJVC done in 294 prevalent hemodialysis patients were analyzed. A right-sided carotid pseudoaneurysm was the only major puncture-related complication. A total of 78 RJVC and 17 LJVC were inserted more than once in the same position. Of the RJVC, 44 (10.9%) of 403 were removed because of infection compared with 16 (20.8%) of 77 LJVC (P < 0.02). The overall incidence of infections was 1.58 episodes of infection per 1000 catheter days, 1.57 for RJVC and 3.72 for LJVC, respectively. Catheter dwell times were not different. A group of 127 patients with former RJVC and 44 with LJVC had ipsilateral arteriovenous access (RJVC+ and LJVC+ group, respectively). Four diabetic LJVC+ developed severe arm swelling secondary to CVO leading ultimately to access ligation. The RJVC+ group had no access ligated. LJVC may cause CVO in diabetics. PMID- 15270955 TI - Composite device for attachment of polyvinyl alcohol-hydrogel to underlying bone. AB - Polyvinyl alcohol-hydrogels (PVA-H) have been specifically proposed as promising prosthetic biomaterials because of their excellent mechanical properties and biocompatibility. However, it is very difficult to obtain stable long-term fixation of implanted PVA-H and this problem is the key point to be solved in the further development of PVA-H in biomedical applications more widely in various fields. Orthopedic implants using PVA-H have been developed and in the process this problem was solved using a composite device. This device consists of PVA-H and a titanium fiber mesh (TFM) as a porous tissue-ingrowth material. In the present study, the mechanical properties of this composite device produced using a new improved production method (injection molding) were investigated and animal experiments for in vivo evaluation were performed. In vitro tests showed that the shear strength at the interface of the PVA-H and TFM, and the mechanical properties of PVA-H itself were promising. Moreover, the histological appearance of the composite device implant in vivo showed abundant bone ingrowth into the deep pore spaces of TFM, which indicated that this device attached firmly to the adjacent bone. These results indicated this composite device prepared by this new method can supply firm attachment of PVA-H onto underlying bone, and that it will be available for various biomedical applications using PVA-H. PMID- 15270956 TI - Injury in organs after cardiopulmonary bypass: a comparative experimental morphological study between a centrifugal and a new pulsatile pump. AB - The aim of this investigation was to assess organ injury provoked by a new pulsatile pump for cardiopulmonary bypass (CPB) with respect to a conventional centrifugal pump. Eight pigs in the pulsatile group (PG) and five in the centrifugal group (CG) underwent a partial CPB lasting 180 min. The animals were sacrificed 180 min after CPB was suspended, and a morphological study of fragments of ventricular wall, liver, lung, and kidney was performed. In CG, centrilobular hepatic necrosis was observed accompanied by sinusoidal dilatation and congestion, multiple focuses of myocardial ischemia, and minor to moderate pulmonary interstitial edema. In PG, diffuse centrilobular sinusoidal congestion in the liver, congestion and capillary dilatation of low intensity in the ventricular wall, and nonsignificant pulmonary interstitial septal edema was observed. In the kidney, both groups showed degenerative changes of the tubular cells and nonsignificant tubular dilatation. These results suggest a better peripheral circulation in the pulsatile group. PMID- 15270957 TI - Balloon-pump-induced pulsatility improves coronary and carotid flows in an experimental model of BioMedicus left ventricular assistance. AB - The aim of this study is to evaluate the benefit of the simultaneous use of a BioMedicus left ventricular assistance device (Medtronic, Minneapolis, MN, U.S.A.) and an intra-aortic balloon pump on regional blood flows, pressure, and pulsatility. Twelve pigs are studied. A BioMedicus pump was placed between the left atrium and the ascending aorta and an intra-aortic balloon pump was inserted through the left femoral artery. Blood flow and pressure were measured in the carotid, femoral, and coronary arteries and in the thoracic aorta below the intra aortic balloon in the basal experimental condition with a full-flow BioMedicus pump and with a full-flow BioMedicus pump + intra-aortic balloon. The BioMedicus pump eliminates pulsatility in all sites and significantly decreases coronary and carotid blood flow. The adjunction of an intra-aortic balloon restores pulsatility to values comparable to those recorded in basal conditions. Coronary and carotid flows even increase to values higher than in the basal conditions. The simultaneous use of an intra-aortic balloon combined with the BioMedicus pump provides a pulsatile flow and increases coronary and carotid blood flows in pigs. An intra-aortic balloon can easily be combined with a BioMedicus pump whenever possible and may improve myocardial recovery in patients with postcardiotomy ventricular failure. PMID- 15270958 TI - Molecular adsorbents recirculating system dialysis for liver insufficiency and sepsis following right ventricular assist device after cardiac surgery. AB - We report a case of right heart failure (RHF) and sepsis with liver insufficiency in a 70-year-old patient after coronary artery bypass graft surgery. Three hours after surgery the patient suddenly developed therapy refractory cardiac arrest caused by RHF. He had to have emergency surgery, under which the graft to the right coronary artery was revised and a right ventricular assist device was implanted. Heart function recovered and the assist device was explanted on day 1 after surgery. Thoracic closure was performed on day 5 after surgery. The patient went into septic shock on day 11. Liver dysfunction developed postoperatively and worsened the course of sepsis. Therefore, MARS (molecular adsorbents recirculating system) dialysis was performed once on day 20 after surgery. Liver function improved after MARS therapy and the patient recovered from sepsis. On day 46 the patient was transferred from the ICU of another hospital to one of the peripheral wards, to be finally discharged on day 67. PMID- 15270959 TI - Peritoneal T cell responses can be polarized toward Th1 or Th2 in children on chronic peritoneal dialysis. AB - Peritoneal T cell responses can be polarized toward Th1 or Th2 in children on chronic peritoneal dialysis. Previous studies on the peritoneal immune system described the presence of activated T lymphocytes in peritoneal effluents from subjects on chronic peritoneal dialysis (CPD). Since Th1/Th2 polarized response can influence the outcome of specific infectious diseases, we investigated if activated Th1/Th2 cells can be detected in peritoneal effluents during peritoneal dialysis, in order to better understand the role of T cells in the mechanisms of peritoneal defense. We have studied 8 children (4 males, 4 females, mean age 5.8 +/- 5.7 years, range 0.3-13.4) on CPD. Peritoneal cells have been isolated from peritoneal effluents by centrifugation. Immunofluorescent staining of intracellular cytokines for flow cytometric analysis was used to detect the percentage of T cells producing either IFN-gamma (Th1) or IL-4 (Th2). In the initial study 3 months after CPD initiation, high percentages of IFN-gamma positive peritoneal T cells (38% and 63%) were detected in two subjects; this finding is consistent with a Th1 polarization of peritoneal T cells. In another subject, high percentages of IL-4 positive T cells (31%) were detected, suggesting a Th2 polarization of peritoneal T cell response. Small amounts of either Th1 or Th2 T cells (2-4%) were also detected in the other subjects. At the 1 year follow-up, Th1 polarization persisted in one subject (18% IFN-gamma positive peritoneal T cells), in another a shift from Th1 to Th2 was observed, and in the other subject a down regulation of both T cell subsets occurred. The finding that a predominance of T cells producing either IFN-gamma or IL-4 was found in 3 out of 8 children strongly suggests that peritoneal T cell responses can be polarized toward Th1 or Th2. The decrease of Th1 and/or Th2 polarized T cells in the peritoneum of 4 out of 6 subjects (after 1 year) suggests that CPD can play an immunosuppressive role on T cell peritoneal responses. Further studies are needed in order to define whether different T helper activation patterns are associated with a higher risk of peritoneal infection or of peritoneal damage. PMID- 15270961 TI - Shift work, the anaesthetist and Santayana's warning. PMID- 15270962 TI - Plasma volume changes associated with two hydroxyethyl starch colloids following acute hypovolaemia in volunteers. AB - This randomised double blind prospective study compared the effective intravascular volume expansion and maintenance, with two types of starches following induced haemorrhagic hypovolaemia. Twenty healthy male volunteers aged between 18 and 65 year were bled 10% of their total blood volume in fully monitored conditions and under the supervision of a trained specialist doctor and research nurse. The lost blood volume was replaced using one of the starch solutions. Effective intravascular volume expansion was monitored hourly using the (51)Cr radio-labelled red blood cell dilution technique, we compared the effects of two hydroxyethyl starch colloid preparations, one a high molecular weight and the other a low molecular weight preparation, on the plasma volume changes over time. The large molecular weight starch (Hextend) provided a less well-sustained volume expansion effect than the smaller one (Voluven) PMID- 15270963 TI - Impact of spinal anaesthesia and obesity on maternal respiratory function during elective Caesarean section. AB - Spinal anaesthesia for Caesarean section has gained widespread acceptance. We assessed the impact of spinal anaesthesia and body mass index (BMI) on spirometric performance. In this prospective study, we consecutively assessed 71 consenting parturients receiving spinal anaesthesia with hyperbaric bupivacaine and fentanyl for elective Caesarean section. We performed spirometry during the antepartum visit (baseline), immediately after spinal anaesthesia, 10-20 min, 1 h, 2 h after the operation, and after mobilisation (3 h). Baseline values were within normal ranges. There was a significant decrease in all spirometric parameters after effective spinal anaesthesia that persisted throughout the study period. The decrease in respiratory function was significantly greater in obese (BMI > 30 kg x m(-2)) than in normal-weight parturients (BMI < 25 kg x m(-2)), e.g. median (IQR) vital capacity directly after spinal anaesthesia; -24 (-16 to 31)% vs. -11 (-6 to -16)%, p < 0.001 and recovery was significantly slower. We conclude that both spinal anaesthesia and obesity significantly impair respiratory function in parturients. PMID- 15270964 TI - Reversal of vecuronium with neostigmine in patients with diabetes mellitus. AB - Reversal of vecuronium-induced neuromuscular blockade with neostigmine was compared in two groups of 16 subjects: patients with Type 2 diabetes mellitus and normal controls. When the first twitch of the train-of-four had returned to 25% of the control value, neostigmine 40 microg x kg(-1) and atropine 20 microg x kg( 1) were given to reverse the neuromuscular blockade. The train-of-four ratio was lower at 3 min, 6 min, 9 min, 12 min and 15 min after reversal in the diabetic group than in the control group but the differences did not reach statistical significance. Fifteen minutes after reversal, the number of patients in whom recovery from neuromuscular blockade was judged insufficient to guarantee good respiratory function (train-of-four ratio < 0.74) did not differ between the groups. However, 15 min after reversal, the number of patients with a train-of four ratio < 0.9 was significantly higher in the Diabetic Group than in the Control Group (15 vs. 10, p = 0.033). PMID- 15270965 TI - Clearing the cervical spine after polytrauma: implementing unified management for unconscious victims in the intensive care unit. AB - Determining the best method for excluding cervical spine injury while a polytrauma victim is unconscious remains a controversial topic despite a number of published guidelines. A structured questionnaire demonstrated major differences between intensivists, neurosurgeons, orthopaedic surgeons and spinal surgeons with regard to the imaging modalities requested, the perception of their performance, the relative risks of missed injuries and the complications of immobilisation. Unconscious victims of polytrauma often come under the care of several subspecialties, with the direct consequence that management can be contradictory and lack standardisation. Advanced Trauma Life Support and Eastern Association for the Surgery of Trauma guidelines can reinforce and even contribute to non-standardised care. Having performed this clinician survey, we have now developed a multidisciplinary management protocol appropriate for Northern Ireland. PMID- 15270966 TI - The effect of critical care outreach on postoperative serious adverse events. AB - We proposed that critical care outreach would decrease the incidence of postoperative serious adverse events and so conducted a sequential cohort study with a surveillance-only phase (baseline) followed by an intervention phase. We studied high-risk patients in a large Australian hospital. A critical care qualified nurse reviewed patients for the first three days after return to the general wards. During the intervention phase the nurse intervened in patient care where appropriate. We examined the incidence of 11 categories of serious adverse events per 100 patients during the first three days on the general wards during the surveillance and intervention phases. The surveillance phase had 319 patients and the intervention phase 345 patients. In a subgroup analysis, there were four myocardial infarctions per 100 patients in the surveillance phase and seven per 100 patients during the intervention phase (95% confidence interval: 1-7 infarctions per 100 patients increase). For the other 10 serious adverse events there were 19 per 100 patients in the surveillance phase and 11 per 100 patients in the intervention phase (95% confidence interval: 4-11 serious adverse events per 100 patients decrease). Outreach may have led to greater detection of myocardial infarctions while reducing the incidence of other serious adverse events. PMID- 15270967 TI - Variation in the transcription of laboratory data in an intensive care unit. AB - Manual transcription of numerical data is prone to error. We quantified the transcription error rate for blood results recorded in a critical care setting by comparing the handwritten and printed laboratory results in 100 consecutive patients in the intensive care unit, Glasgow Royal Infirmary. Nine hundred and fifty-four sets of results with 4664 individual values were analysed. There was complete and accurate transcription in 67.6% (n = 645) of cases, a failure to transcribe in 23.6% (n = 225) and inaccurate transcription in 8.8% (n = 84). Transcription was significantly more accurate in the morning (p = 0.02). This study highlights that our current system of recording blood results is unreliable. These results strengthen the case for computerisation of the patient record in terms of data retrieval and transcription accuracy. PMID- 15270968 TI - Provision of long-term venous access procedures by UK anaesthetists: a postal survey. AB - Long-term venous access is widely used in hospital and in the community for cancer chemotherapy, total parenteral nutrition and long-term administration of antibiotics. There is a large variety of catheters, ports and other devices designed to facilitate these treatments. A postal survey of anaesthetic departments in England, Wales, Scotland and Northern Ireland was undertaken to assess the role of anaesthetists in this area of clinical practice. Two hundred and fifteen out of 276 (78%) anaesthetic departments responded. Forty-three percent of departments (92 out of 215) provided some form of long-term vascular access service. Twenty-two percent of departments which provided this service (20 out of 92) had anaesthetists with sessional allocation for such procedures. Such work represents a significant workload for anaesthetic departments which is likely to increase over time. PMID- 15270969 TI - An audit of peribulbar blockade using 15 mm, 25 mm and 37.5 mm needles, and sub Tenon's injection. AB - The efficacy of peribulbar anaesthesia performed with short, medium and long needles, with sub-Tenon's injection as a control, was audited. Two hundred patients undergoing cataract surgery underwent peribulbar injection using 25G needles of the following lengths: 15 mm, 25 mm or 37.5 mm. Sub-Tenon's injections were performed with a curved 25-mm sub-Tenon anaesthesia cannula. The injection technique, ocular akinesia and analgesia scoring system, and supplementary injection protocols were standardised. After initial injections of local anaesthetic via the sub-Tenon's cannula or with 37.5 mm, 25 mm and 15 mm needles, supplementation was required in one (2%), 13 (26%), 22 (44%) and 32 (64%) of patients, respectively; the total number of supplementary injections required were 1, 16, 35 and 47, respectively. It is concluded that the efficacy of peribulbar anaesthesia depends upon the proximity of the deposition of local anaesthetic solution either to the globe or orbital apex. These data justify the classification of peribulbar anaesthesia into: circum-ocular (sub-Tenon's, episcleral), peri-ocular (anterior, superficial); peri-conal (posterior, deep) and apical (ultra-deep) for teaching purposes. PMID- 15270970 TI - Effects of the European Working Time Directive on anaesthetic training in the United Kingdom. AB - Decreases in the hours worked by trainee anaesthetists are being brought about by both the New Deal for Trainees and the European Working Time Directive. Anticipated improvements in health and safety achieved by a decrease in hours will be at the expense of training time if the amount of night-time work remains constant. This audit examined the effects of a change from a partial to a full shift system on a cohort of trainee anaesthetists working in a large district general hospital in the South-west of England. Logbook and list analyses were performed for two 10-week periods: one before and one after the decrease in hours. An 18% decrease in the number of cases done and an 11% decrease in the number of weekly training lists were found for specialist registrars. A 22% decrease in the number of cases done and a 14% decrease in the number of weekly training lists were found for senior house officers. Furthermore, a decrease of one service list per specialist registrar per week was seen, which will have implications for consultant manpower requirements. PMID- 15270971 TI - Integrated approaches to academic anaesthesia - the Cambridge experience. AB - There is mounting concern about the pressures experienced by University Departments of Anaesthesia, which, if lost, could threaten undergraduate peri operative medicine teaching, development of critical appraisal skills among anaesthetists, and the future of coherent research programs. We have addressed these problems by establishing a foundation course in scientific methods and research techniques (the Cambridge SMART Course), complemented by competitive, fully funded, 12-month academic trainee attachments. Research conducted during academic attachments has been published and used to underpin substantive grant applications allowing work towards higher degrees. Following the attachment, a flexible scheme ensures safe reintroduction to clinical training. Research at consultant level is facilitated by encouraging applications for Clinician Scientist Fellowships, and by ensuring that the University Department champions, legitimises and validates the allocation of research time within the new consultant contract. We believe that these are important steps in safeguarding research and teaching in anaesthesia, critical care and peri-operative medicine. PMID- 15270972 TI - An American tale - professional conflicts in anaesthesia in the United States: implications for the United Kingdom. AB - Professional conflict between nurse anaesthetists and anaesthesiologists in the United States of America is well known in the UK but has not been explored and documented in detail. We present an account, based on critical analysis of published literature and other documentary evidence, of the historical, professional and financial factors which have led to this. In the USA, anaesthesia developed as a nursing specialty until physicians began to take on this work after the Second World War. Payment arrangements between the 1960s and the 1990s made anaesthesiology a lucrative career choice for medical graduates and this led both to considerable growth in the number of anaesthesiologists and to a strengthening of the resolve of nurse anaesthetists to retain their scope of work and preserve their professional status. Changes in payment regulations in the 1980s and 1990s threatened anaesthesiologists' income and led to re-appraisal of evidence over relative cost-effectiveness and safety of different provider models. More recently, the terms of engagement have shifted from disputes over evidence to political lobbying to promote the professional capabilities and status of each of the anaesthesia providers. Factors of relevance to possible changes in the provision of anaesthesia in the United Kingdom are highlighted. PMID- 15270973 TI - Ximelagatran, a new oral direct thrombin inhibitor, for the prevention of venous thromboembolic events in major elective orthopaedic surgery. Efficacy, safety and anaesthetic considerations. AB - The oral direct thrombin inhibitor ximelagatran shows great promise for prevention of venous thromboembolic events following major elective orthopaedic surgery. Its consistent and predictable pharmacokinetics and pharmacodynamics across a wide range of patient populations allow administration with fixed dosing and with no coagulation monitoring. In orthopaedic surgery clinical trials, ximelagatran was effective and well tolerated compared with standard therapy, with dose and timing relative to surgery important factors in determining its optimal profile. In European trials, an initial 3-mg postoperative dose of subcutaneous melagatran, the active form of ximelagatran, followed by oral ximelagatran 24 mg twice daily achieved similar efficacy and safety to enoxaparin. Although the risk of spinal haematoma following neuraxial anaesthesia is rare, it is increased by the concomitant use of anticoagulants. In orthopaedic surgery trials with ximelagatran to date, complications such as spinal haematoma have not been reported. The pharmacokinetic profile of ximelagatran suggests that concurrent use with neuraxial anaesthesia should require no further precautions than those currently necessary with low-molecular-weight heparin. PMID- 15270974 TI - Evaluation of Frova, single-use intubation introducer, in a manikin. Comparison with Eschmann multiple-use introducer and Portex single-use introducer. AB - In a randomised cross-over study, 48 anaesthetists attempted to place a Frova single-use introducer, an Eschmann multiple-use introducer and a Portex single use introducer in the trachea of a manikin set up to simulate a grade 3 laryngoscopic view. The anaesthetists were blinded to success (tracheal placement) or failure (oesophageal placement). Successful placement (proportion, 95% confidence interval) of either the Frova introducer (65%, 50-77%) or the Eschmann introducer (60%, 46-73%) was significantly more likely than with the Portex introducer (8%, 3-20%). There were no significant differences between the success rates for the Frova and the Eschmann introducers. A separate experiment revealed that the peak force exerted by the Frova and Portex introducers was two to three times greater than that which could be exerted by the Eschmann introducer, p < 0.0001, indicating that the single-use introducers are more likely to cause tissue trauma during placement. PMID- 15270975 TI - The air elimination capabilities of pressure infusion devices and fluid-warmers. AB - Pressurised infusion devices may have only limited capability to detect and remove air during pressurised infusions. In order to assess pressure infusion systems with regard to their actual air elimination capabilities four disposable pressure infusion systems and fluid warmers were investigated: The Level 1 (L-1), Ranger (RA), Gymar (GY), and the Warmflo (WF). Different volumes of air were injected proximal to the heat exchanger and the remaining amount of air that was delivered at the end of the tubing was measured during pressurised infusions. Elimination of the injected air (100-200 ml) was superior by the RA system when compared to L-1 (p < 0.01). The GY and WF systems failed to eliminate the injected air. In conclusion, air elimination was best performed by the RA system. In terms of the risk of air embolism during pressurised infusions, improvements in air elimination of the investigated devices are still necessary. PMID- 15270976 TI - Consent for publication of a case report. PMID- 15270977 TI - Prepreparation of succinylcholine. PMID- 15270978 TI - Methylene blue or patent blue? PMID- 15270979 TI - Failed gastric tube insertion in the LMA-ProSeal. PMID- 15270980 TI - Safety of the Pulsiocath for haemodynamic monitoring during magnetic resonance imaging. PMID- 15270981 TI - Blockage of a fluid administration set. PMID- 15270982 TI - Topiramate induced metabolic acidosis. PMID- 15270983 TI - Increased carbon dioxide during anaesthesia in patients with phaeochromocytoma. PMID- 15270984 TI - Proposal for a different ranking of anaesthesia journals. PMID- 15270985 TI - Critical incident due to a Guedel airway. PMID- 15270986 TI - A new use of rubber gloves for venous cannulation. PMID- 15270987 TI - Tracheal intubation through a laryngeal mask may kink the pilot tube. PMID- 15270988 TI - Death from chloroform? PMID- 15270989 TI - Euphemia Maclean, Agnes Sampson and pain relief during labour in 16th century Edinburgh. PMID- 15270990 TI - Breathing circuit failure due to surgical 'prep' solution. PMID- 15270991 TI - Another use for the LOCKIT epidural catheter clamp. PMID- 15270993 TI - Effects of racial diversity on complex thinking in college students. AB - An experiment varying the racial (Black, White) and opinion composition in small group discussions was conducted with college students (N = 357) at three universities to test for effects on the perceived novelty of group members' contributions to discussion and on participants' integrative complexity. Results showed that racial and opinion minorities were both perceived as contributing to novelty. Generally positive effects on integrative complexity were found when the groups had racial- and opinion-minority members and when members reported having racially diverse friends and classmates. The findings are discussed in the context of social psychological theories of minority influence and social policy implications for affirmative action. The research supports claims about the educational significance of race in higher education, as well as the complexity of the interaction of racial diversity with contextual and individual factors. PMID- 15270994 TI - Music lessons enhance IQ. AB - The idea that music makes you smarter has received considerable attention from scholars and the media. The present report is the first to test this hypothesis directly with random assignment of a large sample of children (N = 144) to two different types of music lessons (keyboard or voice) or to control groups that received drama lessons or no lessons. IQ was measured before and after the lessons. Compared with children in the control groups, children in the music groups exhibited greater increases in full-scale IQ. The effect was relatively small, but it generalized across IQ subtests, index scores, and a standardized measure of academic achievement. Unexpectedly, children in the drama group exhibited substantial pre- to post-test improvements in adaptive social behavior that were not evident in the music groups. PMID- 15270995 TI - The correlates and consequences of newspaper reports of research on sex differences. AB - Correlates and consequences of newspaper accounts of research on sex differences were examined. In Study 1, articles from high-circulation newspapers were coded for the degree to which biological factors were used to explain sex differences. Results showed that political conservatism and traditional attitudes toward gender roles coded from other newspaper sections predicted greater use of biological explanations than did political liberalism and less traditional attitudes toward gender roles. In Studies 2 and 3, participants read a fictional newspaper article reporting research on a gender difference that cited either biological or sociocultural factors as explaining the difference. Results showed that exposure to biological explanations significantly increased participants' endorsement of gender stereotypes. Moreover, exposure to social explanations significantly increased participants' belief in the mutability of human behavior. Together, these studies show that political ideology influences how the popular press reports research findings and that such reporting in turn affects readers' beliefs and attitudes. PMID- 15270996 TI - Using and being used by categories. The case of negative evaluations and daily well-being. AB - Three studies involving 257 undergraduates examined whether a simple choice reaction time task could predict daily experiences of affect. Individuals who were fast to make negative evaluations experienced more negative affect and more somatic symptoms and were less satisfied with their lives, compared with individuals who were slow to make negative evaluations. A fourth study, involving 89 undergraduates, indicated that performance on the task was relatively unaffected by transitory mood states. The results support the idea that categorization provides a useful perspective on personality functioning. PMID- 15270997 TI - Is there any "free" choice? Self and dissonance in two cultures. AB - Four experiments provided support for the hypothesis that upon making a choice, individuals justify their choice in order to eliminate doubts about culturally sanctioned aspects of the self, namely, competence and efficacy in North America and positive appraisal by other people in Japan. Japanese participants justified their choice (by increasing liking for chosen items and decreasing liking for rejected items) in the standard free-choice dissonance paradigm only when self relevant others were primed, either by questionnaires (Studies 1-3) or by incidental exposure to schematic faces (Study 4). In the absence of these social cues, Japanese participants showed no dissonance effect. In contrast, European Americans justified their choices regardless of the social-cue manipulations. Implications for cognitive dissonance theory are discussed. PMID- 15270998 TI - Decisions from experience and the effect of rare events in risky choice. AB - When people have access to information sources such as newspaper weather forecasts, drug-package inserts, and mutual-fund brochures, all of which provide convenient descriptions of risky prospects, they can make decisions from description. When people must decide whether to back up their computer's hard drive, cross a busy street, or go out on a date, however, they typically do not have any summary description of the possible outcomes or their likelihoods. For such decisions, people can call only on their own encounters with such prospects, making decisions from experience. Decisions from experience and decisions from description can lead to dramatically different choice behavior. In the case of decisions from description, people make choices as if they overweight the probability of rare events, as described by prospect theory. We found that in the case of decisions from experience, in contrast, people make choices as if they underweight the probability of rare events, and we explored the impact of two possible causes of this underweighting--reliance on relatively small samples of information and overweighting of recently sampled information. We conclude with a call for two different theories of risky choice. PMID- 15270999 TI - Thinking about low-probability events. An Exemplar-Cuing theory. AB - The way people respond to the chance that an unlikely event will occur depends on how the event is described. We propose that people attach more weight to unlikely events when they can easily generate or imagine examples in which the event has occurred or will occur than when they cannot. We tested this idea in two experiments with mock jurors using written murder scenarios. The results suggested that jurors attach more weight to the defendant's claim that an incriminating DNA match is merely coincidental when it is easy for them to imagine other individuals whose DNA would also match than when it is not easy for them to imagine such individuals. We manipulated the difficulty of imagining such examples by varying the description of the DNA-match statistic. Some of the variations that influenced the jurors were normatively irrelevant. PMID- 15271000 TI - Mechanisms of belief-desire reasoning. Inhibition and bias. AB - Biases in reasoning can provide insight into underlying processing mechanisms. We demonstrate a new bias in children's belief-desire reasoning. Children between 4 and 8 years of age were told a story in which a character was mistaken about which of three boxes contained some object. The character wanted to go to one of the boxes, but only if it did not contain the object. In this scenario, the character would be expected to avoid the box where she falsely believed the object to be, but might go to either of the remaining boxes. Though the character was equally likely to go to either box, children were biased to predict that the character would go to the box that contained the object. In a control task, the character had the same desire but did not have a false belief; in this case, children showed no bias, choosing the two correct answers equally often. The observed pattern of bias was predicted by a developmental model of belief-desire reasoning. Competent belief-desire reasoning depends on a process of selection by inhibition in which the best belief content emerges from a set of candidates. PMID- 15271001 TI - When development and learning decrease memory. Evidence against category-based induction in children. AB - Inductive inference is crucial for learning: If one learns that a cat has a particular biological property, one could expand this knowledge to other cats. We argue that young children perform induction on the basis of similarity of compared entities, whereas adults may induce on the basis of category information. If different processes underlie induction at different points in development, young children and adults would form different memory traces during induction, and would subsequently have different memory accuracy. Experiment 1 demonstrates that after performing an induction task, 5-year-olds exhibit more accurate memory than adults. Experiment 2 indicates that after 5-year-olds are trained to perform induction in an adultlike manner, their memory accuracy drops to the level of adults. These results, indicating that sometimes 5-year-olds exhibit better memory than adults, support the claim that, unlike adults, young children perform similarity-based rather than category-based induction. PMID- 15271002 TI - Scene consistency in object and background perception. AB - Does knowledge about which objects and settings tend to co-occur affect how people interpret an image? The effects of consistency on perception were investigated using manipulated photographs containing a foreground object that was either semantically consistent or inconsistent with its setting. In four experiments, participants reported the foreground object, the setting, or both after seeing each picture for 80 ms followed by a mask. In Experiment 1, objects were identified more accurately in a consistent than an inconsistent setting. In Experiment 2, backgrounds were identified more accurately when they contained a consistent rather than an inconsistent foreground object. In Experiment 3, objects were presented without backgrounds and backgrounds without objects; comparison with the other experiments indicated that objects were identified better in isolation than when presented with a background, but there was no difference in accuracy for backgrounds whether they appeared with a foreground object or not. Finally, in Experiment 4, consistency effects remained when both objects and backgrounds were reported. Semantic consistency information is available when a scene is glimpsed briefly and affects both object and background perception. Objects and their settings are processed interactively and not in isolation. PMID- 15271003 TI - Perception of three-dimensional shape from specular highlights, deformations of shading, and other types of visual information. AB - There have been numerous computational models developed in an effort to explain how the human visual system analyzes three-dimensional (3D) surface shape from patterns of image shading, but they all share some important limitations. Models that are applicable to individual static images cannot correctly interpret regions that contain specular highlights, and those that are applicable to moving images have difficulties when a surface moves relative to its sources of illumination. Here we describe a psychophysical experiment that measured the sensitivity of human observers to small differences of 3D shape over a wide variety of conditions. The results provide clear evidence that the presence of specular highlights or the motions of a surface relative to its light source do not pose an impediment to perception, but rather, provide powerful sources of information for the perceptual analysis of 3D shape. PMID- 15271004 TI - Yes, there is a preferential detection of negative stimuli. A response to Labiouse. PMID- 15271005 TI - Ethical issues with implantable defibrillators. PMID- 15271006 TI - Comments on ethical issues with implantable defibrillators by F. James Brennan. PMID- 15271007 TI - Comments on ethical issues with implantable defibrillators by F. James Brennan. PMID- 15271008 TI - Excessive increase in QT interval and dispersion of repolarization predict recurrent ventricular tachyarrhythmia after amiodarone. AB - Although chronic amiodarone has been proven to be effective to suppress ventricular tachycardia (VT) and ventricular fibrillation (VF), how we predict the recurrence of VT/VF after chronic amiodarone remains unknown. This study evaluated the predictive value of the QT interval, spatial, and transmural dispersions of repolarization (SDR and TDR) for further arrhythmic events after chronic amiodarone. Eighty-seven leads body surface ECGs were recorded before (pre) and one month after (post) chronic oral amiodarone in 50 patients with sustained monomorphic VT associated with organic heart disease. The Q-Tend (QTe), the Q-Tpeak (QTp), and the interval between Tpeak and Tend (Tp-e) as an index of TDR were measured automatically from 87-lead ECG, corrected Bazett's method (QTce, QTcp, Tcp-e), and averaged among all 87 leads. As an index of SDR, the maximum (max) minus minimum (min) QTce (max-min QTce) and standard deviation of QTce (SD-QTce) was obtained among 87 leads. All patients were prospectively followed (15 +/- 10 months) after starting amiodarone, and 20 patients had arrhythmic events. The univariate analysis revealed that post max QTce, post SD QTce, post max-min QTce, and post mean Tcp-e from 87-lead but not from 12-lead ECG were the significant predictors for further arrhythmic events. ROC analysis indicated the post max-min QTce > or = 106 ms as the best predictor of events (hazard ratio = 10.4, 95%, CI 2.7 to 40.5, P = 0.0008). Excessive QT prolongation associated with increased spatial and transmural dispersions of repolarization predict the recurrence of VT/VF after amiodarone treatment. PMID- 15271009 TI - Improving the acceptability of the atrial defibrillator: patient-activated cardioversion versus automatic night cardioversion with and without sedation (ADSAS 2). AB - Acceptability of the atrial defibrillator is partly limited by concerns about shock related anxiety and discomfort. Sedation and/or automatic cardioversion therapy during sleep may ease shock discomfort and improve patient acceptability. Three atrial cardioversion techniques were compared: patient-activated cardioversion with sedation, automatic night cardioversion with sedation, and automatic night cardioversion without sedation. Sedation was oral midazolam (15 mg). Fifteen patients aged 60 +/- 13 years were assigned each strategy randomly for three consecutive episodes of persistent atrial fibrillation requiring cardioversion. Patients completed questionnaires for multiple parameters immediately and again at 24 hours postcardioversion. Atrial cardioversion strategies with oral sedation (patient-activated and automatic) significantly reduced shock recall by 77% (P < 0.005), therapy dissatisfaction by 57%-71% (P < 0.03), shock discomfort by 61%-73% (P < 0.01), shock pain by 79%-83% (P < 0.001), and shock intensity by 73%-77% (P < 0.03), compared to automatic night cardioversion without sedation (P < 0.02). Atrial shock pain was short-lived and caused little disruption to the patients' daily routines. Automatic night cardioversion without sedation, resulted in sleep disturbances not seen with the other strategies (42% vs 0%, P < 0.001) as well as concerns about future pain or discomfort. Twelve patients (80%) chose patient-activated cardioversion with sedation as their preferred treatment, and three (20%) remainder chose automatic night cardioversion with sedation. Ninety percent of patients chose automatic night cardioversion without sedation as the least acceptable therapy. Sedation significantly increases atrial shock acceptability regardless of cardioversion method. Shocks without sedation are significantly less acceptable to patients using the atrial defibrillators. PMID- 15271010 TI - Situational syncope: response to head-up tilt testing and follow-up: comparison with vasovagal syncope. AB - Among sequential patients with neurally-mediated syncope, we studied the response to head-up tilt test (HUTT) in patients with situational syncope (SS) and their follow-up. Our findings were compared to those in patients with vasovagal syncope (VVS). The response to HUTT in patients with SS has not to date been fully investigated. Additionally, the prognosis of SS patients has not been systematically studied. We studied 162 consecutive patients with recurrent SS or VVS, all free of structural heart disease. Before study inclusion, they underwent an HUTT and were followed up for 12 months. Patients with SS were advised to avoid the trigger event. Patients with VVS were treated with propranolol or fluoxetine. For each patient we compared the number of syncopal spells during the last 12 months before study inclusion with that during follow-up. Among the 162 patients, 36 had SS and 126 had VVS. The response to HUTT and the number of syncopes before and during follow-up were similar in both groups. Among patients with SS, 10 (28%) had also experienced occasional episodes of VVS; however, they had a similar response to HUTT and prognosis to the remaining 26 SS patients without VVS attacks. Patients with SS have a similar response to HUTT and similarly benign clinical course to patients with VVS. The coexistence of occasional VVS episodes in patients with SS is not associated with a higher rate of positive HUTT or worse prognosis. PMID- 15271011 TI - High incidence of appropriate and inappropriate ICD therapies in children and adolescents with implantable cardioverter defibrillator. AB - Appropriate and inappropriate therapies of implantable cardioverter defibrillators have a major impact on morbidity and quality of life in ICD recipients, but have not been systematically studied in children and young adults during long-term follow-up. ICD implantation was performed in 20 patients at the mean age of 16 +/- 6 years, 11 of which had prior surgical repair of a congenital heart defect, 9 patients had other cardiac diseases. Implant indications were aborted sudden cardiac death in six patients, recurrent ventricular tachycardia in 9 patient, and syncope in 5 patients. Epicardial implantation was performed in 6 and transvenous implantation in 14 patients. Incidence, reasons and predictors (age, gender, repaired congenital heart disease, history of supraventricular tachycardia, and epicardial electrode system) of appropriate and inappropriate ICD therapies were analyzed during a mean follow-up period of 51 +/- 31 months range 18-132 months. There were a total 239 ICD therapies in 17 patients (85%) with a therapy rate of 2.8 per patient-years of follow-up. 127 (53%) ICD therapies in 15 (75%) patients were catagorized as appropriate and 112 (47%) therapies in 10 (50%) patients as inappropriate, with a rate of 1.5 appropriate and 1.3 inappropriate ICD therapies per patient-years of follow-up. Time to first appropriate therapy was 16 +/- 18 months. Appropriate therapies were caused by ventricular fibrillation in 29 and ventricular tachycardia in 98 episodes. Termination was successful by antitachycardia pacing in 4 (3%) and by shock therapy in 123 episodes (97%). Time to first inappropriate therapy was 16 +/- 17 months. Inappropriate therapies were caused by supraventricular tachycardia in 77 (69%), T wave oversensing in 19 (17%), and electrode defect in 16 episodes (14%). It caused shocks in 87 (78%) and only antitachycardia pacing in 25 episodes (22%). No clinical variable could be identified as predictor of either appropriate or inappropriate ICD therapies. There is a high rate of ICD therapies in young ICD recipients, the majority of which occur during early follow-up. The rate of inappropriate therapies is as high as 47% and is caused by supraventricular tachycardia and electrode complications in the majority of cases. Prospective trials are required to establish preventative strategies of ICD therapies in this young patient population. PMID- 15271012 TI - Ablation lesion size correlates with pacing threshold: a physiological basis for use of pacing to assess ablation lesions. AB - The virtual electrode model predicts that pacing stimulus strength should reflect proximity of the pacing electrode to excitable myocardium, allowing pacing threshold to assess radiofrequency (RF) ablation lesions and unexcitable scar. The purpose of this study is to correlate RF lesion size with pacing threshold and electrogram (EG) amplitude change at the ablation site. In four swine (32-58 kg, 20 ventricular RF lesions were created using a 4-mm tip electrode catheters under fluoroscopic and electroanatomic guidance. Unipolar pacing threshold and bipolar and unipolar EG amplitude were measured before and after ablation and compared with lesion size measured in the fixed, serially sectioned tissue. Lesion diameter ranged from 6.4 to 19 mm and volume ranged from 29 to 1920 mm3. Ablation increased the pacing threshold by 320%, from 0.9 +/- 0.3 to 3.6 +/- 2.6 mA, P < 0.001. The change in pacing threshold correlated with lesion volume R = 0.88, P < 0.001). Linear regression predicts that lesion volume (mm3) = 160 X rise in pacing threshold + 13. Ablation reduced peak to peak bipolar EG amplitude by 56%, from 2.5 +/- 2.0 mV to 1.1 +/- 0.6 mV (P = 0.005). Unipolar EG amplitude diminished by only 22% from 4.0 +/- 1.6 to 3.2 +/- 0.9 mV postablation (P = 0.005). The correlations of lesion volume with change in either bipolar R = 0.14, P = 0.6) or unipolar R = 0.18, P = 0.6) EG amplitude were poor. Pacing threshold correlates with RF ablation lesion size, consistent with the virtual electrode model. In normal myocardium, change in pacing threshold is likely to be a better marker of lesion size than electrogram amplitude. PMID- 15271013 TI - Is isolation of all four pulmonary veins necessary in patients with paroxysmal atrial fibrillation? AB - The upper pulmonary veins (PVs) are responsible for the majority of atrial fibrillation (AF) triggering foci, whereas the inferior PVs are more difficult to ablate and prone to postablation ostial stenosis. Most procedure failures can be attributed to incomplete isolation or recurrent PV left atrial reconnection rather than to identification of another focus. Furthermore, in certain patients AF triggers can be detected outside the PVs, and local denervation of the ganglionic plexus around the superior PV left atrial junctions following the ablation procedure may also play a role in eliminating AF. Based on these data, the authors propose that in AF patients the superior PVs should be ablated first, and in case of recurrence a second procedure should be performed for identification of PV left atrial reconnection or extrapulmonary foci and additional ablation of the inferior PVs. Such a staged approach might offer slightly lower success rates but with a significantly lower radiation exposure and procedural time and at a smaller risk of ablation induced PV stenosis. PMID- 15271014 TI - Surgical open-chest ventricular defibrillation: triphasic waveforms are superior to biphasic waveforms. AB - Triphasic shocks have been evaluated for endocardial defibrillation but not for open-chest epicardial defibrillation. The purpose of this study was to compare the efficacy and safety of biphasic versus triphasic shocks for epicardial defibrillation in a porcine model. Twenty-two adult swine (18-28 kg) were deeply anesthetized and intubated. After 30 seconds electrically induced VF, each pig received truncated exponential biphasic (7.2-ms positive pulse duration and 7.2 ms negative pulse duration, total waveform duration 14.4 ms) and triphasic (4.8/4.8/4.8 ms, total waveform duration 14.4 ms) epicardial shocks. Pigs in group 1 (n = 11) received epicardial biphasic and triphasic shocks from large hand held paddle electrodes (44.2 cm2); pigs in group 2 (n = 11) received shocks from small paddle electrodes (15.9 cm2). Shocks were given at five selected energy levels (3-30 J) in random sequence. Four shocks were delivered at each energy level to construct an energy versus percentage of success curve. In group 1 (large paddle electrodes), percentage of shock success was significantly higher for triphasic shocks at the energy levels of 3, 5, 10, and 20 J compared to biphasic shocks. In group 2 (small paddle electrodes), triphasic shocks yielded a significantly higher percentage of shock success than biphasic shocks at the energy levels of 5, 10, and 20 J). Shock induced ventricular tachycardia was similar for both waveforms; asystole was rare. For open-chest defibrillation, triphasic waveform shocks were superior to biphasic waveform shocks for VF termination at energy levels of 3-20 J and were as safe as biphasic shocks. PMID- 15271015 TI - Endocardial atrial pacing lead implantation and midterm follow-up in young patients with sinus node dysfunction after the fontan procedure. AB - The purpose of the study was to investigate the results of endocardial lead implantation, lead performance, and follow-up in young patients after the Fontan procedure. A retrospective study was conducted with patients who had endocardial atrial pacing for SND and intact AVN function after Fontan from two pediatric centers. Patient demographics, pacing, and sensing data of endocardial atrial leads were analyzed at the time of pacemaker implantation and follow-up visits. Fifteen patients (weight 42.6 +/- 35 kg) had transvenous endocardial atrial lead implantation at an average age of 11.4 +/- 6.5 years. Active-fixation leads were used in all patients and steroid elution was present in 12 (80%) patients. Adequate P wave sensing was obtained in patients with sinus rhythm (n = 10); the remaining four patients had junctional rhythm without measurable P waves. Lead failure was not observed in any patient during the follow-up period of 2.9 +/- 2.1 years. The energy threshold at implantation was 1.46 +/- 1.5 microJ, 1.54 +/- 0.75 microJ at 3 months, 0.62 +/- 0.45 microJ at 1 year, 0.72 +/- 0.65 microJ at 2 years, 0.75 +/- 0.55 microJ at 3 years, and 0.8 +/- 0.85 microJ at 5 years postimplant. The lead impedance was 648 +/- 298 omega at implantation, 714 +/- 163 omega at 3 months, 744 +/- 195 omega at 1 year, 734 +/- 198 omega at 2 years, 800 +/- 142 omega at 3 years and 830 +/- 200 omega 5 years postimplant. Anticoagulation therapy (aspirin n = 5, warfarin n = 8) was continued by 13 patients. Complications consisted of a pneumothorax at implantation and a transient ischemic attack in one patient 4 years after pacemaker implant. Endocardial atrial leads offer low energy thresholds and can be implanted relatively safely in Fontan patients. PMID- 15271016 TI - The world survey of cardiac pacing and cardioverter defibrillators: calendar year 2001. AB - A worldwide cardiac pacing and ICD survey was undertaken for calendar year 2001. Fifty countries, 22 from Europe, 16 from the Asia Pacific region, 3 from the Middle East and Africa, and 9 from the Americas contributed to the survey. The United States had by far the largest number of cardiac pacemaker implants, although Germany had the highest new implants per million population. Virtually all countries that participated in the 1997 survey showed significant increases in implant numbers over the 4 years. High degree atrioventricular block and sick sinus syndrome remain the major indications for implantation of a cardiac pacemaker with < 2% biventricular pacing in those countries that implanted such systems in 2001. There remains a high percentage of VVIR pacing in the developing countries with only a few countries using substantial numbers of single lead VDD and AAIR systems. There has been an increase in the use of DDDR systems in most countries since the 1997 survey. Pacing leads were predominantly transvenous, bipolar, and passive fixation. There was an increased use of active-fixation leads in the atrium. There was a significant rise in the use of ICDs with the largest usage occurring in the United States. A group of enthusiastic survey coordinators has now been established. Recruitment of new countries will hopefully continue to obtain a fully global experience of cardiac pacing and ICD usage. PMID- 15271017 TI - Cooled intramural needle catheter ablation creates deeper lesions than irrigated tip catheter ablation. AB - Endocardial radiofrequency ablation of the left ventricle does not create transmural lesions reliably even with active electrode cooling. The authors developed a prototype catheter with an internally cooled needle electrode that could be advanced an adjustable distance into the myocardium. Freshly excised hearts from eight male sheep were perfused and superfused using oxygenated ovine blood. Ablations were performed for 2 minutes using the prototype catheter and a conventional endocardial 5-mm irrigated tip ablation catheter at target temperatures of 80 degrees C and 50 degrees C, respectively. The prototype catheter needle was inserted 12 mm deep for all ablations. The maximal power and irrigation rate was 50 W, 20 mL/min for the irrigated tip catheter and 20 W, 10 mL/min for the intramural needle catheter. Intramural needle lesions were significantly deeper (13.5 +/- 2.3 vs 9.1 +/- 1.3 mm, P < 0.01) but less wide (8.7 +/- 1.5 vs 12.7 +/- 1.9 mm, P < 0.01) than irrigated tip lesions. Popping occurred during 12 (37%) of the 32 irrigated tip ablations. Popping did not occur during intramural needle ablation. The cooled intramural needle ablation catheter creates lesions that are significantly deeper than irrigated tip catheters with less tissue boiling. In contrast to irrigated tip ablation, electrode temperature monitoring can be used to determine if a lesion has been created during intramural needle ablation. The cooled intramural needle ablation lesions were of a clinically useful width, addressing one of the main recognized deficiencies of intramural needle ablation. PMID- 15271018 TI - Radiofrequency catheter ablation for arrhythmic storm in patients with an implantable cardioverter defibrillator. AB - The aim of the study was to analyze the usefulness of RFA in controlling arrhythmic storm due to hemodynamically stable VT in a cohort of patients with an ICD and to evaluate the incidence of arrhythmic storm among patients with an ICD. A group of 13 (3%) of 403 consecutive ICD recipients were submitted to RFA of VT during an arrhythmic storm. Two additional patients were referred from other institutions. Standard criteria were used for VT endocardial ablation. A transcatheter epicardial approach was required in three patients. A total of 18 procedures were performed in 15 patients. A mean of 13.2 +/- 9.7 pulses of RF were delivered. Clinical tachycardia was successfully ablated in 12 (80%) patients. One patient died in incessant VT, 1 patient underwent heart transplant, and 1 was treated with direct current ablation. During a mean follow-up of 12 +/- 17 months, only two patients suffered a single episode of VT. Arrhythmic storm requiring VT ablation was uncommon among patients with an ICD and occurred late after ICD implantation. The arrhythmic episode was successfully controlled in the majority of patients with endocardial or epicardial RFA. PMID- 15271019 TI - Comparison of the specificity of implantable dual chamber defibrillator detection algorithms. AB - The aim of the study was to compare the specificity of dual chamber ICDs detection algorithms for correct classification of supraventricular tachyarrhythmias derived from clinical studies according to their size to detect an impact of sample size on the specificity. Furthermore, the study sought to compare the specificities of detection algorithms calculated from clinical data with the specificity calculated from simulations of tachyarrhythmias. A survey was conducted of all available sources providing data regarding the specificity of five dual chamber ICDs. The specificity was correlated with the number of patients included, number of episodes, and number of supraventricular tachyarrhythmias recorded. The simulation was performed using tachyarrhythmias recorded in the electrophysiology laboratory. The range of the number of patients included into the studies was 78-1,029, the range of the total number of episodes recorded was 362-5,788, and the range of the number of supraventricular tachyarrhythmias used for calculation of the specificity for correct detection of these arrhythmias was 100 (Biotronik) to 1662 (Medtronic). The specificity for correct detection of supraventricular tachyarrhythmias was 90% (Biotronik), 89% (ELA Medical), 89% (Guidant), 68% (Medtronic), and 76% (St. Jude Medical). There was an inverse correlation (r = -0.9, P = 0.037) between the specificity for correct classification of supraventricular tachyarrhythmias and the number of patients. The specificity for correct detection of supraventricular tachyarrhythmias calculated from the simulation after correction for the clinical prevalence of the simulated tachyarrhythmias was 95% (Biotronik), 99% (ELA Medical), 94% (Guidant), 93% (Medtronic), and 92% (St. Jude Medical). In conclusion, the specificity of ICD detection algorithms calculated from clinical studies or registries may depend on the number of patients studied. Therefore, a direct comparison between different detection algorithms based on clinical data is difficult. In contrast, simulation of supraventricular tachyarrhythmias using a uniform database may be a better tool for direct comparison of the specificity of ICD detection algorithms. PMID- 15271020 TI - Accuracy of atrial tachyarrhythmia detection in implantable devices with arrhythmia therapies. AB - The clinical application of atrial tachyarrhythmia (AT) episode data stored by implantable devices is presently limited by the high proportion of inappropriate detection. We quantified the percentage of inappropriate AT detection in two implantable devices with AT diagnostics and therapies via meta-analysis of stored AT episodes from a number of clinical trials. The AT500 and GEM III AT, contain dual chamber logic to discriminate AT from ventricular tachycardia and far-field R wave (FFRW) oversensing using dual chamber bipolar electrograms. A subset of data from four clinical trials of 1,142 patients was considered. Manual analysis was performed on 21,553 stored episodes with atrial EGM and marker channel from 409 patients with stored episodes and the market-released device detection configuration. The percentage of episodes with inappropriate detection and termination was evaluated and compared between septal and nonseptal lead locations. The percentage of inappropriately detected episodes receiving ATP therapy was also determined. The percentage of episodes appropriately detected and the percentage of net episode duration (i.e., burden) recorded by the device were also determined from a separate analysis of 24-hour Holter recordings from a subset of 40 patients from one trial. Adjusted estimates of the percentage of appropriate [corrected] detection were 95.3% (93.5-96.7; 95% CI) for AT500 and 95.7% (84.3-98.9) for GEM III AT. Inappropriate detection was primarily due to FFRW oversensing or brief runs of premature atrial contractions (PACs). The device detected 100% of the sustained atrial arrhythmia episodes and 95.3% (range 76.1-99.9) of the net AT duration observed on the Holter recordings. AT detection was not influenced by atrial lead location. Appropriate detection of normal sinus rhythm at episode termination was 83.7% (80.7-86.3) for AT500 and 92.1% (84.5 96.2) for GEM III AT. Accurate detection and discrimination of FFRWs validates the reliability of AT diagnostic data and decreases the risk of inappropriate device therapy. PMID- 15271021 TI - Narrow QRS complex tachycardia: what is the mechanism? PMID- 15271023 TI - Unexpected loss of atrial tracking caused by interaction between temporary and permanent right ventricular leads during implantation of a biventricular pacemaker. AB - This report describes a patient who underwent implantation of an atriobiventricular pacemaker following AV junction ablation and insertion of a temporary right ventricular (RV) pacemaker. During implantation, intermittent loss of sinus P wave tracking occurred when the three permanent leads were connected to the generator. Analysis of marker annotation disclosed intermittent abnormal ventricular sensing that reinitiated postventricular atrial blanking and caused failure of P wave tracking. This phenomenon disappeared after removing the temporary RV lead, but not by turning off the temporary pacemaker. We assume that mechanical contact between the temporary and the permanent RV leads is the underlying mechanism. PMID- 15271024 TI - Complete loss of ICD programmability after magnetic resonance imaging. AB - The purpose of this case report is to describe the effects of an MRI performed on a patient without realizing that an ICD has been previously implanted. After a few seconds of imaging the adversity was recognized and the examination was stopped immediately. The patient was not pacemaker dependent and had neither physical complaints nor electrocardiographic changes in the surface ECG. A consecutively performed ICD assessment showed a backup mode with standard parameters for pacing (VVI 50 beats/min) and arrhythmia detection and treatment. The device could not be programmed by the external programmer. With the exception of printing out the parameters, all software functions were no longer feasible. A device examination by the manufacturer after ICD replacement showed that a major portion of the device memory was corrupt. Even ICDs of a newer generation are susceptible to magnetic interference, with the danger of complete loss of programmability. PMID- 15271025 TI - Catecholamine dependent accessory pathway automaticity. AB - Automaticity from extra nodal accessory pathways appears to be rare. We report the case of a man with the WPW syndrome who presented for repeat electrophysiological study and catheter ablation. After successful ablation of a para-Hisian accessory pathway, an isoproterenol challenge produced an accelerated wide complex rhythm that was dissociated from sinus rhythm and matched the previous pattern of maximal preexcitation. This automatic rhythm was transient and dependent on catecholamine administration. One month after successful ablation, an exercise treadmill test (ETT) did not demonstrate any pre-excitation or ectopy. PMID- 15271026 TI - Pacemaker lead infection secondary to Haemophilus parainfluenzae. AB - Cardiac device infections are a rare complication of pacing and defibrillator therapy. The number of implanted devices will likely continue to rise with increasing implantation of the cardioverter defibrillator and cardiac resynchronization devices. This report describes a case of an uncommon pathogen for device-associated endocarditis. PMID- 15271027 TI - Magnetic resonance imaging-based biventricular pacemaker upgrade. AB - This report describes a patient with drug refractory severe chronic ischemic heart failure, atrial fibrillation with bradycardia, and left bundle branch block who had a failed implantation of a biventricular pacemaker because of a high left ventricular pacing threshold. VVI pacemaker implantation had not improved the patient's condition. MRI-guided biventricular pacemaker upgrade had been performed with a left ventricular epicardial lead at the lateral region where a 4 mm thickening during systole had been proven. After 6 months of effective resynchronization, the patient's functional class improved to NYHA II without further need of hospitalization. PMID- 15271028 TI - Implantation of a biventricular pacing system in a patient with a persistent left superior vena cava. AB - The presence of a persistent left superior vena cava was encountered in a 64-year old man undergoing implantation of a biventricular pacing system. Leads with active fixation were positioned in the right atrium and right ventricle, through the persistent left superior vena cava and the proximal segment of the coronary sinus. For left ventricular pacing, a standard bipolar lead was positioned directly in the posterior branch of the coronary sinus without the use of special guiding catheters. PMID- 15271029 TI - Retrograde cycle length alternans during supraventricular tachycardia: an unusual tachycardia mechanism. AB - Cycle length alternans is occasionally seen during supraventricular tachycardia due to oscillations in the antegrade atrioventricular nodal (AVN) refractoriness. However, alternans due to retrograde variation in AVN conduction has not been reported. This report describes the case of a 36-year-old man with atypical AVN reentry tachycardia (AVNRT) whose episodes of tachycardia were characterized by continuous oscillations in retrograde AVN conduction. Ablation at one spot eliminated the tachycardia. Cycle length alternans due to oscillations in retrograde AVN conduction, although rare, can be seen during atypical AVNRT and should be considered. PMID- 15271030 TI - Brugada pattern electrocardiographic changes associated with profound electrolyte disturbance. AB - Only a few case reports of Brugada pattern ECG changes caused by electrolyte disturbance exist in the literature, all of which lack adequate electrophysiological exclusion (or inclusion) of an underlying Brugada syndrome. This report describes a case of an otherwise healthy 38-year-old man who presented to the hospital with diabetic ketoacidosis, profound electrolyte disturbance, and Brugada pattern ECG changes. Subsequent flecanide drug challenge and electrophysiological studies ruled out an underlying Brugada syndrome. PMID- 15271031 TI - Pulmonary vein isolation during atrial fibrillation using a circumferential cryoablation catheter. AB - Pulmonary vein (PV) isolation for the treatment of atrial fibrillation is limited by procedure related complications, such as PV stenosis and occlusions. We report about a PV isolation using a circumferential cryoablation catheter which applies the ablation energy simultaneously at the entire circumference by cooling down to a minimal temperature of -80 degrees C. PMID- 15271032 TI - Evolution of mapping and anatomic imaging of cardiac arrhythmias. PMID- 15271035 TI - Academic medicine in crisis. PMID- 15271036 TI - Learning by scar formation. PMID- 15271037 TI - The relationship between learning style and learning environment. PMID- 15271038 TI - Reflecting on learning portfolios. PMID- 15271039 TI - A wider perspective on assessment. PMID- 15271040 TI - Changing education, changing assessment, changing research? AB - BACKGROUND: In medical education, assessment of medical competence and performance, important changes have taken place in the last 5 decades. These changes have affected the basic concepts in all 3 domains. DEVELOPMENTS IN EDUCATION AND ASSESSMENT: In education constructivism has provided a completely new view on how students learn best. In assessment the change from trait orientated to competency- or role-orientated thinking has given rise to a whole range of new approaches. Certain methods of education, such as problem-based learning (PBL), and assessment, however, are often seen as almost synonymous with the underlying concepts, and one tends to forget that it is the concept that is important and that a particular method is but 1 way of using a concept. When doing this, one runs the risk of confusing means and ends, which may hamper or slow down new developments. LESSONS FOR RESEARCH: A similar problem seems to occur often in research of medical education. Here too, methods--or, rather, methodologies--are confused with research questions. This may lead to an overemphasis on research that fits well known methodologies (e.g. the randomised controlled trial) and neglect of what are sometimes even more important research questions because they do not fit well known methodologies. CONCLUSION: In this paper we advocate a return to the underlying concepts and a careful reflection of their use in various situations. PMID- 15271041 TI - Assessment of medical communication skills by computer: assessment method and student experiences. AB - BACKGROUND: A computer-assisted assessment (CAA) program for communication skills designated ACT was developed using the objective structured video examination (OSVE) format. This method features assessment of cognitive scripts underlying communication behaviour, a broad range of communication problems covered in 1 assessment, highly standardised assessment and rating procedures, and large group assessments without complex organisation. SETTING: The Academic Medical Centre (AMC) at the University of Amsterdam, the Netherlands. Aims To describe the development of the AMC Communication Test (ACT); to describe our experiences with the examination and rating procedures; to present test score descriptives, and to present the students' opinions of ACT. DESIGN: The ACT presents films on history taking, breaking bad news and shared decision making. Each film is accompanied by 3 types of short essay questions derived from our assessment model: "knows", "knows why/when" and "knows how". Evaluation questions about ACT were integrated into the assessment. Participants A total of 210 third year medical undergraduates were assessed. This study reports on the 110 (53%) students who completed all evaluation questions. RESULTS: Marking 210 examinations took about 17 days. The test score matched a normal distribution and showed a good level of discrimination of the students. About 75% passed the examination. Some support for the validity of our assessment model was found in the students' differential performance on the 3 types of questions. The ACT was well received. Student evaluations confirmed our efforts to develop realistic films that related well to the communication training programme. CONCLUSIONS: The ACT is a useful assessment method which complements interpersonal assessment methods for the evaluation of the medical communication skills of undergraduates. PMID- 15271042 TI - Factor analysis can be a useful standard setting tool in a high stakes OSCE assessment. AB - BACKGROUND: No method of standard setting for objective structured clinical examinations (OSCEs) is perfect. Using scores aggregated across stations risks allowing students who are incompetent in some core skills to pass an examination, which may not be acceptable for high stakes assessments. AIM: To assess the feasibility of using a factor analysis of station scores in a high stakes OSCE to derive measures of underlying competencies. METHODS: A 12-station OSCE was administered to all 192 students in the penultimate undergraduate year at the University of Aberdeen Medical School. Analysis of the correlation table of station scores was used to exclude stations performing unreliably. Factor analysis of the remaining station scores was carried out to characterise the underlying competencies being assessed. Factor scores were used to derive pass/fail cut-off scores for the examination. RESULTS: Four stations were identified as having unpredicted variations in station scores. Analysis of the content of these stations allowed the underlying problems with the station designs to be isolated. Factor analysis of the remaining 8 stations revealed 3 main underlying factors, accounting for 53% of the total variance in scores. These were labelled "examination skills", "communication skills" and "history taking skills". CONCLUSION: Factor analysis is a useful tool for characterising and quantifying the skills that are assessed in an OSCE. Standard setting procedures can be used to calculate cut-off scores for each underlying factor. PMID- 15271043 TI - Evaluating the impact of moving from discipline-based to integrated assessment. AB - BACKGROUND: The move from discipline-based to problem-based learning (PBL) at Adelaide University in 2000 offered exciting opportunities to integrate the teaching and learning of the basic and clinical sciences for medical undergraduates. However, several cohorts of students still needed to progress through the first 3 years of the more traditional curriculum. Paradoxically, their readiness to function in the integrated learning and assessment environment of the last 3 years was assessed in 7 separate discipline-based examinations at the end of third year. When considerable examination-related stress was noted in the 1997 cohort and students petitioned formally for a reduced examination load, it was considered to be time for assessment to lead the way in integrating the disciplines. AIM: After introducing third year integrated written assessments in 1998, we aimed to develop an integrated practical examination (IPE) linking theory to practice, and evaluate its impact on staff and students. METHODS: After extensive staff collaboration, a structured objective multistation IPE was developed and administered in 1999 and 2000. Its utility was evaluated using a model proposed earlier. RESULTS: Assessment validity was maximised by an extensive item review process. Reliability, as measured by Cronbach's alpha, was 0.79 and 0.80 in 1999 and 2000, respectively. An independent evaluation yielded qualitative data on the examination's educational impact, cost and acceptability. CONCLUSIONS: Investing time in changing from discipline-based to integrated assessment, integrating theory and practice, resulted in gains in assessment reliability, validity and educational impact on both staff and students. PMID- 15271044 TI - Training the trainers: do teaching courses develop teaching skills? AB - OBJECTIVE: This paper reports on consultants' self-assessed changes in their teaching and training practices over an 8-10-month period. It compares the changes between a group undergoing a 3-day teaching course (participants) and a sample group taken from the course waiting list (controls). METHOD: A questionnaire listing 18 teaching skills was given to the participants immediately prior to the course and 8-10 months later, and to the controls at the same time intervals. Respondents were asked to rate their ability, frequency of use of each skill, as well as their teaching confidence and effectiveness. Additionally, the second questionnaire asked respondents to identify changes they had made to their teaching. A total of 63% (54) of participants and 51% (23) of controls completed both questionnaires. Changes of 2 + on the rating scales were seen as genuine. The number of such changes was calculated for each individual and on each skill for the 2 groups. Data were analysed using a Mann-Whitney U test. RESULTS: The majority of course participants reported positive changes in teaching ability on a significantly greater number of skills than did the control group. As a group, changes in ability in 16 of the teaching skills were significantly greater for the participants than for the controls. Increased ability resulted in participants' increased frequency of use of only 4 of the teaching skills. The majority in the participant group reported changes to their teaching. Only a minority in the control group reported such changes. These changes were consistent with course topics and the teaching skills needed to meet General Medical Council recommendations for the education of new doctors. CONCLUSIONS: The teaching course is an effective vehicle for increasing consultants' teaching skills. PMID- 15271045 TI - Factors that influence doctors' participation in clinical research. AB - BACKGROUND: Although clinical investigators are regarded as an endangered species, no systematic investigation of the factors that influence doctor participation in clinical research has previously been performed. AIM: The objective of this study was to evaluate the influences of selected aspects of medical education, specialty selection and practice type upon current involvement in clinical research. METHODS: Data were obtained by a mail survey of 428 graduates from the 1985-95 classes of the Penn State College of Medicine. RESULTS: Among the 34% (n = 145) of doctors who were currently participating in clinical research, there was a higher rate of participation among those in medical and surgical specialties versus those in primary care or hospital-based specialties. Of those participating in clinical research, 46% (n = 65) had sought external funds for their research, and 82% (n = 51) of that group had been awarded funds. Those who had been awarded funds as the percentage of their time involved in clinical research increased were more likely to report that research carried out in medical school had positively influenced their current involvement in clinical research (P = 0.004). The gender distribution among both this latter funded group and the larger group of 145 who were participating in clinical research was 72% men and 28% women, whereas the gender distribution among all respondents was 60% men and 40% women. The 283 individuals who were not participating in clinical research cited financial, family, career plan and practice philosophy as reasons for not doing so. CONCLUSION: The authors conclude that participation in medical research as a medical student may be under recognised as a determinant of future involvement in clinical research and that the gender disparity of young doctors entering clinical research must be addressed. PMID- 15271046 TI - Improving the quality of outpatient clinic letters using the Sheffield Assessment Instrument for Letters (SAIL). AB - AIM: To improve the quality of outpatient letters used as communication between hospital and primary care doctors. METHODS: On 2 separate occasions, 15 unselected outpatient letters written by each of 7 hospital practitioners were rated by another hospital doctor and a general practitioner (GP) using the Sheffield Assessment Instrument for Letters (SAIL). Individualised feedback was provided to participants following the rating of the first set of letters. The audit cycle was completed 3 months later without forewarning by repeat assessment by the same hospital and GP assessors using the SAIL tool to see if there was any improvement in correspondence. SETTING: Single centre: general paediatric outpatient department in a large district general hospital. RESULTS: All 7 doctors available for reassessment completed the audit loop, each providing 15 outpatient letters per assessment. The mean of the quality scores, derived for each letter from the summation of a 20-point checklist and a global score, improved from 23.3 (95% CI 22.1-24.4) to 26.6 (95% CI 25.8-27.4) (P = 0.001). CONCLUSIONS: The SAIL provides a feasible and reliable method of assessing the quality and content of outpatient clinic letters. This study demonstrates that it can also provide feedback with a powerful educational impact. This approach holds real potential for appraisal and revalidation, providing an effective means for the quality improvement required by clinical governance. PMID- 15271047 TI - The effectiveness of problem-based learning compared to traditional teaching in undergraduate psychiatry. AB - OBJECTIVES: A change from traditional to problem-based learning (PBL) methods in a psychiatry attachment was evaluated by comparing the learning styles, attitudes to psychiatry and examination performance of 2 cohorts of students. It was hypothesised that the PBL curriculum would result in increased deep learning, decreased surface learning, more favourable attitudes to psychiatry and improved examination performance. It was predicted that students' examination success would be related to the use of deep and strategic learning and favourable attitudes. METHODS: Consecutive cohorts of Year 2 clinical students taught using a traditional psychiatry curriculum (n = 188) and a PBL curriculum (n = 191) were compared. Students completed the Study Process Questionnaire to assess their learning styles and the Attitudes to Psychiatry Scale at the beginning and end of the attachment. Students completed 2 end-of-attachment examinations, a multiple choice paper and a viva. RESULTS: The PBL curriculum resulted in significantly better examination performance than did the traditional teaching curriculum, both for multiple-choice questions and the viva. No differences in learning styles or attitudes to psychiatry were found between the curricula. Students were significantly more successful in the examinations if they had received the PBL curriculum, were female, and used strategic learning. CONCLUSIONS: Examination performance indicated that the PBL curriculum was more successful than the previous course, but that this improvement was not due to students using more effective learning styles or having more favourable attitudes towards psychiatry. It is possible that students learned more effectively during the teaching sessions in the PBL curriculum, but did not change their preferred learning styles. PMID- 15271048 TI - Teaching and evaluating first and second year medical students' practice of evidence-based medicine. AB - PURPOSE: To implement an evidence-based medicine (EBM) curriculum for Year 1 and 2 medical students, and to develop a method to evaluate their practice of EBM in discrete and relevant worksteps. METHODS: For the 100 students entering Year 1 of their medical education in 2000, we implemented a curriculum with 25-30 student contact hours of EBM instruction which used a variety of teaching formats and spanned the first and second years of their training. We developed an evaluation module that assessed the following 5 steps in the practice of EBM: generating well built questions; searching for evidence; critical appraisal; applying the evidence, and self-evaluation. We tested 2 different versions of the test module 3-months apart with the same cohort of second year students, and correlated their scores on the second module with examination components of a comprehensive assessment. We obtained feedback from the students regarding the EBM curriculum and evaluation method. RESULTS: Each test module took 2-4 hours to complete and 5 8 minutes to grade. There was moderate test-retest reliability for the total test scores (r = 0.35, P < 0.001). Step 1 scores correlated with the mock board examination scores (r = 0.23, P = 0.05). Step 2 scores correlated with the peer assessment factor "work habits" (r = 0.24, P = 0.02), and Step 3 scores correlated with clinical reasoning exercises (r = 0.31, P = 0.002). Step 4 scores lacked test-retest reliability and did not correlate with components of the comprehensive assessment. The majority of students felt there was too much focus on EBM during the first 2 years of the curriculum and they rated the EBM test module the lowest rated component of the comprehensive assessment. CONCLUSIONS: Although we have demonstrated preliminary reliability and validity of a new evaluation instrument that assess the domains of scientific knowledge, work habits and reasoning skills required in the practice of EBM, many of the correlations were weak, and we remain in the very early stages of determining if, when and how EBM instruction should occur in medical education. PMID- 15271049 TI - The impact of multifaceted educational structuring on learning effectiveness in a surgical clerkship. AB - INTRODUCTION: Various measures have been introduced to enhance learning experiences in clerkships, generally with limited success. This study evaluated the impact of a multifaceted approach on the effectiveness of learning in a surgical clerkship. In accordance with results obtained in continuing medical education, several interventions were introduced simultaneously. We compared students' evaluations of the traditional surgical clerkship with those of the restructured clerkship. METHODS: Two consecutive cohorts of students were asked to complete a questionnaire about the quality and quantity of their learning experiences. Cohort 1 (n = 28) undertook the traditional clerkship and cohort 2 (n = 72) the restructured clerkship. A Mann-Whitney test was used to compare outcomes between the 2 cohorts. RESULTS: There were few statistically significant differences between cohorts 1 and 2. Overall, quality indicators did not differ between the 2 cohorts. DISCUSSION: A short-term multifaceted intervention led to a slight increase in the performance of clinical skills and a slight decrease in time spent on activities of limited educational value. The intervention may have been too brief to produce substantial effects. Future interventions should also target teachers, including trainees, in order to assess their opinions and address their educational needs. PMID- 15271050 TI - Computer-aided learning in the real world of medical education: does the quality of interaction with the computer affect student learning? AB - BACKGROUND: Computerised learning clearly offers exciting potential for improving student learning, either as an aid to or as a replacement for traditional formats, or for the development of innovative approaches. However, rigorous evaluation of the utility of computer-aided learning (CAL) in enhancing student learning can be difficult. Many studies have compared CAL to more traditional learning formats, but there is little evidence to show which style of CAL leads to the best learning outcomes. AIM: This study aimed to test the hypothesis that a CAL tutorial, in which the learner actively interacts with the computer, will result in superior learning (ability to apply and retain knowledge) to that obtained in more passive CAL formats. METHODS: Third year medical undergraduates at Adelaide University, South Australia were randomly assigned to 4 groups. Following a pretest, only students in the "didactic", "problem-based" and "free text" groups had 2 weeks of free access to a neuroradiology CAL tutorial in their assigned format. Tutorial access was denied to all students 2 weeks before post testing. Learning was quantified by comparing the post- to pretest scores for each of the 4 groups. RESULTS: After active interaction with the computer material, students in the free text group demonstrated a statistically significant improvement in their ability to apply and retain knowledge compared to the control group, but no advantage compared to the didactic group. CONCLUSIONS: While users of an interactive CAL tutorial demonstrated significant learning gains compared to non-CAL users, these gains were not superior to those achieved from non-interactive CAL. When evaluating education interventions such as CAL packages, it is important to use a valid assessment tool to measure learning. PMID- 15271051 TI - Evaluation of a surgical simulator for learning clinical anatomy. AB - BACKGROUND: New techniques in imaging and surgery have made 3-dimensional anatomical knowledge an increasingly important goal of medical education. This study compared the efficacy of 2 supplemental, self-study methods for learning shoulder joint anatomy to determine which method provides for greater transfer of learning to the clinical setting. METHODS: Two groups of medical students studied shoulder joint anatomy using either a second-generation virtual reality surgical simulator or images from a textbook. They were then asked to identify anatomical structures of the shoulder joint as they appeared in a videotape of a live arthroscopic procedure. RESULTS: The mean identification scores, out of a possible score of 7, were 3.1 +/- 1.3 for the simulator group and 2.9 +/- 1.5 for the textbook group (P = 0.70). Student ratings of the 2 methods on a 5-point Likert scale were significantly different. The simulator group rated the simulator more highly as an effective learning tool than the textbook group rated the textbook (means of 3.2 +/- 0.7 and 2.6 +/- 0.5, respectively, P = 0.02). Furthermore, the simulator group indicated that they were more likely to use the simulator as a learning tool if it were available to them than the textbook group was willing to use the textbook (means of 4.0 +/- 1.2 and 3.0 +/- 0.9, respectively, P = 0.02). CONCLUSION: Our results show that this surgical simulator is at least as effective as textbook images for learning anatomy and could enhance student learning through increased motivation. These findings provide insight into simulator development and strategies for learning anatomy. Possible explanations and future research directions are discussed. PMID- 15271052 TI - Setting and maintaining professional role boundaries: an educational strategy. AB - AIM: To develop and evaluate a programme focused on assisting medical students in setting and maintaining social and sexual boundaries, within their training and in future medical practice. CONTEXT: In response to allegations of sexual misconduct by medical practitioners, a teaching programme was implemented with, and evaluated by, final year medical students who were undertaking 9 weeks of community health and general practice experience. OUTCOME: The consensus of the students was that professional role boundary issues were complex, their professional ethos had been challenged, and there was a need to incorporate teaching about setting and maintaining role boundaries throughout all facets of the medical curriculum. RESULTS: The pilot programme was successful in engaging students in the process of developing teaching to assist in setting and maintaining social and sexual boundaries. Recommendations to formalise the programme were approved. PMID- 15271053 TI - Teaching anatomy without cadavers. PMID- 15271054 TI - Teaching anatomy as a multimedia experience. PMID- 15271056 TI - First aid and cardiopulmonary resuscitation training for medical students. PMID- 15271057 TI - Neuroendocrinological and molecular aspects of insect reproduction. AB - This review summarizes recent advances and novel concepts in the area of insect reproductive neuroendocrinology. The role of 'classic' hormones, such as ecdysteroids and juvenoids, to control reproduction is well documented in a large variety of insect species. In adult gonads, ecdysteroids appear to induce a cascade of transcription factors, many of which also occur during the larval molting response. Recent molecular and functional data have created opportunities to study an additional level of regulation, that of neuropeptides, growth factors and their respective receptors. As a result, many homologs of factors playing a role in vertebrate reproductive physiology have been discovered in insects. This review highlights several neuropeptides controlling the biosynthesis and release of the 'classic' insect hormones, as well as various peptides and biogenic amines that regulate behavioural aspects of the reproduction process. In addition, hormone metabolizing enzymes and second messenger pathways are discussed with respect to their role in reproductive tissues. Finally, we speculate on future prospects for insect neuroendocrinological research as a consequence of the recent 'Genomics Revolution'. PMID- 15271058 TI - Glial limitans elasticity subjacent to the supraoptic nucleus. AB - Two previous studies from our laboratory have indicated that the ventral glial limitans subjacent to the hypothalamic supraoptic nucleus (SON-VGL) undergoes a reversible thinning upon chronic activation of the magnocellular neuroendocrine cells (MNCs) of the supraoptic nucleus (SON). Numerous other studies have shown that MNC somata hypertrophy with activation. One aim of the current study was to understand better how SON-VGL thinning occurs. A second aim was to quantify overall changes of the MNC somata region due to cellular hypertrophy to compare relative changes in dimensions. Here, we undertook a light microscopic stereological investigation of the SON and the subjacent SON-VGL of Nissl stained material under basal and activated conditions. Astrocyte numbers in the underlying SON-VGL remained stable across hydration state as did the overall volume of the SON-VGL and dendritic zone reference area. How these data are consistent with our earlier observations of SON-VGL thinning was resolved by the finding of a highly significant, 30% increase in the mediolateral dimension of the SON-VGL in dehydrated rats. These observations fit well with previous work from our laboratory that demonstrates a reorientation of SON-VGL astrocytes, from vertical to horizontal, which occurs in the activated SON-VGL. We found a significant, approximately 54%, increase in the overall volume of the MNC region of the SON. No significant rostrocaudal lengthening of the SON was detected, although a trend was evident. All the observed changes reversed with rehydration. These data indicate that elasticity of the SON-VGL acts to accommodate the volume expansion of the MNCs and enables the SON-VGL to continue as an interface between the underlying cerebrospinal fluid in the subarachnoid space and the expanded SON above. PMID- 15271059 TI - Chronic changes in peripheral growth hormone levels do not affect ghrelin stomach mRNA expression and serum ghrelin levels in three transgenic mouse models. AB - Ghrelin is an endogenous ligand for the growth hormone secretagogue (GHS) receptor. Ghrelin is involved in feeding behaviour and is a potent stimulator of GH release. Chronically increased GH concentrations are known to negatively regulate the pituitary GHS receptor. This study tested whether chronic changes in peripheral GH levels/action affect ghrelin mRNA expression and circulating concentrations of ghrelin. Stomach ghrelin mRNA expression and serum concentrations of ghrelin were measured in three groups of transgenic mice and the respective control animals: group 1, GH-receptor gene disrupted mice (GHR/KO); group 2, mice expressing bovine GH (bGH); and group 3, mice expressing GH-antagonist (GHA). Ghrelin mRNA expression in the stomach, pituitary and hypothalamus of young adult male rats were measured using reverse-transcription polymerase chain reaction. Ghrelin mRNA expression levels were approximately 3000 fold higher in rat stomach than in rat pituitary. Ghrelin mRNA expression in rat hypothalamus was below the detection limits of our assay. Stomach ghrelin mRNA expression, as well as serum concentrations of ghrelin, did not change significantly in any of the three mouse groups compared to the respective control group. These data support previous observations that the stomach is the main source of circulating ghrelin, and also indicate that stomach ghrelin mRNA expression and serum concentrations of ghrelin are not affected by chronic changes in peripheral GH/insulin-like growth factor-I levels/action. PMID- 15271060 TI - Glial aromatization decreases neural injury in the zebra finch (Taeniopygia guttata): influence on apoptosis. AB - Emerging evidence suggests a neuroprotective role for oestrogens following damage to the vertebrate brain. Aromatase (oestrogen synthase) is rapidly transcribed and translated in glial cells around areas of neural damage in several vertebrates. However, the potential neuroprotection afforded by locally up regulated glial aromatase immediately surrounding the injury remains to be tested. Towards this end, individual birds sustained penetrating mechanical injuries via a needle that contained either vehicle or the aromatase inhibitor fadrozole into contralateral hemispheres. Seventy-two hours later, the size of neural injury (as assessed by the extent of necrotic tissue) and the number of apoptotic cells around the injuries were evaluated. The size of injury in the hemisphere injected with fadrozole was significantly larger than the injury caused by vehicle injection. Furthermore, a greater number of apoptotic nuclei were found around the fadrozole-associated lesion relative to vehicle. Finally, constitutively expressed, neuronal aromatase close to the injury site did not differ between hemispheres. We conclude that local inhibition of glial aromatase immediately around the site of injury plays a neuroprotective role in the songbird brain and this protection involves apoptotic pathways. Local up regulation of glial aromatase may play a pivotal role in the limitation of secondary damage and/or the acceleration of restorative processes following injury to the vertebrate brain. PMID- 15271061 TI - Somatostatin-14 actions on dopamine- and pituitary adenylate cyclase-activating polypeptide-evoked Ca2+ signals and growth hormone secretion. AB - Using single-cell Ca(2+) imaging and a growth hormone (GH) radioimmunassay, we investigated somatostatin-14 (SS(14)) inhibition of cAMP-dependent, stimulated GH secretion from primary cultures of dispersed goldfish pituitary cells. The dopamine-D1 receptor agonist SKF-38393, and the hypothalamic neuropeptide pituitary adenylate cyclase-activating polypeptide (PACAP) both elevated intracellular Ca(2+) concentration ([Ca(2+)](i)) and stimulated GH release. When increases in [Ca(2+)](i) were prevented by intracellular loading of BAPTA, a Ca(2+) chelator, SKF-38393- and PACAP-stimulated GH release were inhibited, suggesting that these Ca(2+) signals are required for stimulated GH release. SS(14) inhibited SKF-38393- and PACAP-stimulated GH release, but did not prevent these Ca(2+) signals. Kinetic analysis revealed that SS(14) lowered the maximum amplitude of the SKF-38393- and PACAP-evoked Ca(2+) responses, but had no effect on other aspects of the Ca(2+) signal. We then examined the ability of SS(14) to act subsequent to dopamine-D1 or PACAP receptor activation using the adenylate cyclase activator forskolin, or the membrane permeant cAMP analogue 8Br-cAMP. Forskolin and 8Br-cAMP both increased [Ca(2+)](i) and GH secretion and, as expected, SS(14) inhibited the resultant GH release. Although SS(14) significantly increased the time to maximum amplitude of the forskolin-evoked Ca(2+) signals, it had no detectable effect on any of the kinetic parameters used to describe the Ca(2+) signals evoked by 8Br-cAMP. Taken together, these results establish that SS(14) has the ability to suppress Ca(2+)-dependent exocytosis by acting distal to elevations in [Ca(2+)](i). Furthermore, it appears likely that the cellular mechanisms underlying the observed effects of SS(14) on Ca(2+) signalling are upstream of cAMP and may be unrelated to those responsible for inhibiting GH release. PMID- 15271062 TI - Melanocortin peptides stimulate prolactin gene expression and prolactin accumulation in rat pituitary aggregate cell cultures. AB - Treatment for 40 h of reaggregate pituitary cell cultures from 14-day-old female rats with nanomolar concentrations of gamma3-melanocyte-stimulating hormone (MSH) increased prolactin mRNA but not growth hormone (GH) mRNA expression levels as measured by quantitative real-time reverse transcriptase-polymerase chain reaction (RT-PCR). During the 40 h incubation, gamma3-MSH stimulated prolactin accumulation in the culture medium. alpha-MSH, a potent agonist of the rat melanocortin-3 receptor (MC3R) and Ala(8)-gamma2-MSH, a very weak agonist of the MC3R, increased prolactin mRNA expression at a similar concentration range as gamma3-MSH. The effect of gamma3-MSH on prolactin mRNA expression was abolished when aggregates were cultured in the presence of thyroid or glucocorticoid hormones, but not of oestradiol. By contrast, oestradiol abolished the stimulatory effect of Ala(8)-gamma2-MSH on prolactin mRNA expression. In GH3 cells stably transfected with the enhanced green fluorescent protein (eGFP) gene under control of a 3-kb prolactin promoter fragment, a dose as low as 1 nMgamma3 MSH, added for 24 h, significantly increased eGFP fluorescence. Agouti-related protein (AgRP(83-132)), a known endogenous MC3R and MC4R antagonist, did not reduce the stimulation of prolactin mRNA expression by gamma3-MSH or Ala(8) gamma2-MSH. On its own, AgRP(83-132) significantly increased prolactin mRNA expression level and prolactin accumulation. Both gamma2-MSH and Ala(8)-gamma2 MSH increased [S(35)]GTPgammaS binding in membrane preparations of 14-day-old rat pituitaries and of GH3 cells. Whereas MC3R and MC5R mRNA were detectable by RT PCR in normal pituitary, these receptor mRNAs were undetectable in GH3 cells using various oligonucleotide primer sets. The present findings indicate that melanocortin peptides stimulate prolactin gene expression and production and that, at least in part, a receptor different from the classic MCR is involved. AgRP appears to have other actions than its known antagonistic activity on the MC3R and MC4R. PMID- 15271063 TI - Oestradiol restores cell proliferation in dentate gyrus and subventricular zone of streptozotocin-diabetic mice. AB - Type 1 diabetes mellitus correlates with several brain disturbances, including hypersensitivity to stress, cognitive impairment, increased risk of stroke and dementia. Within the central nervous system, the hippocampus is considered a special target for alterations associated with diabetes. Neurogenesis is a plastic event restricted to few adult brain areas: the subgranular zone of the dentate gyrus and the subventricular zone (SVZ). First, we studied the ability for neurogenesis in the dentate gyrus and SVZ of chronic diabetic mice induced by streptozotocin (STZ). Using bromodeoxyuridine (BrdU) labelling of cells in the S phase, we observed a strong reduction in cell proliferation rate in both brain regions of diabetic mice killed 20 days after STZ administration. Second, because oestrogens are active neuroprotective agents, we investigated whether 17beta oestradiol (200 micro g pellet implant in cholesterol during 10 days) restored brain cell proliferation in the diabetic mouse brain. Our results demonstrated a complete reversibility of dentate gyrus cell proliferation in oestrogen-treated diabetic mice. This plasticity change was not exclusive to the hippocampus because oestrogen treatment restored BrdU incorporation into newborn cells of the SVZ region of diabetic animals. Oestrogen treatment did not alter the hyperglycemic status of STZ-diabetic mice. Moreover, oestrogen did not modify BrdU incorporation in control animals. These data show that oestrogen treatment strongly stimulates brain neurogenesis of diabetic mice and open up new venues for understanding the potential neuroprotective role of steroid hormones in diabetic encephalopathy. PMID- 15271064 TI - Glutamic acid decarboxylase 67 (GAD67) gene expression in discrete regions of the rostral preoptic area change during the oestrous cycle and with age. AB - Gamma-aminobutyric acid (GABA), an inhibitory neurotransmitter, is important to the timing and amplitude of the gonadotrophin-releasing hormone (GnRH)-induced luteinizing hormone (LH) surge on pro-oestrus. Data suggest that GABA input in the preoptic area must decrease for a normal LH surge to occur in young rats. We have previously found that ageing alters the timing and amplitude of the LH surge. Therefore, this study focused on changes in GAD(67) gene expression, a reflection of GABA synthesis, in two regions of the rostral preoptic area, the organum vasculosum of the lamina terminalis (OVLT) and the anteroventral periventricular nucleus (AVPV) during the oestrous cycle and with age. We assessed the possibility that the expression of GAD(67) mRNA in these regions displays time-related and age-dependent changes on pro-oestrus. Our results demonstrate that, with age, overall expression of GAD(67) mRNA decreases in the area surrounding the OVLT and in the AVPV. Young rats display a diurnal rhythm in GAD(67) mRNA in both regions. GAD(67) mRNA expression is high during the early morning hours of pro-oestrus and then declines around the time of the GnRH induced LH surge. In addition, the diurnal rhythm disappears in the AVPV and is attenuated in the area surrounding the OVLT of middle-aged proestrous rats. These findings suggest that a loss of rhythmicity in GAD(67) gene expression and maintenance of inhibitory tone on proestrous afternoon may alter the timing and amplitude of the LH surge, as previously observed in middle-aged rats. PMID- 15271065 TI - Galanin-like peptide as a link between metabolism and reproduction. AB - Galanin-like peptide (GALP) is a hypothalamic neuropeptide that binds and activates galanin receptors in vitro. Following the discovery of GALP, researchers have attempted to properly place it in the context of galanin receptor physiology. Central injections of GALP have revealed some common actions with galanin, such as acutely increased food intake and suppression of the thyroid axis. Other actions are unique to GALP, such as long-term inhibition of food intake and stimulation of luteinizing hormone (LH) secretion in male rats. GALP and galanin also produce differential effects on expression of the immediate early gene product Fos in the brain. Determining which of these actions are dependent on galanin receptors (versus a putative GALP-specific receptor), as well as which actions represent the authentic physiology of endogenous GALP will require continued experimentation. GALP gene expression is positively regulated by several hormones involved in the control of energy balance and metabolism, namely leptin, insulin and thyroid hormone. Based on current evidence, GALP neurones may serve as a hypothalamic relay, transmitting information from the periphery to circuits within the brain involved in the physiological control of metabolism and reproduction. PMID- 15271066 TI - Systemic and mucosal immunity in rhesus macaques immunized with HIV-1 peptide and gp120 conjugated to Brucella abortus. AB - HIV vaccine testing in primates is an important method for determining the possibility of vaccine benefit in humans. Goals of HIV-1 vaccination include establishing neutralizing antibodies and a strong CD8(+) T-cell response. We tested a novel vaccine conjugate for its ability to elicit relevant immune responses to HIV proteins and peptides in rhesus macaques. A neutralizing epitope, V3 loop peptide from HIV-1 envelope, was coupled to heat-inactivated Brucella abortus (V3-HKBA). Rhesus macaques were immunized with this conjugate in the anterior thigh. After two immunizations V3-specific antibodies were found in the sera and at mucosal sites. Neutralizing activity of these antibodies was demonstrated by syncytia inhibition assays. Cellular immune recall responses were demonstrated by antigen-specific induction of interferon-gamma and Regulation on Activation Noraml T Cell Expressed and Secreted (RANTES) secretion in vitro. These results confirm and extend preliminary studies in mice that suggest HKBA is an effective carrier that promotes neutralizing antibody secretion at relevant mucosal sites, as well as cellular immune responses that are correlated with viral protection. PMID- 15271067 TI - Changes in hematology, biochemical values, and restraint ECG of rhesus monkeys (Macaca mulatta) following 6-month laboratory acclimation. AB - The aim of this study was to determine if 6-month acclimation would enable accurate evaluation of hematological, biochemical data, and ECG recorded under restraint for conscious rhesus monkeys of both sexes. Periodic evaluation of these parameters was made during the 6-month period of acclimation. The platelet count, alkaline phosphatase, glucose, and sodium levels significantly decreased, whereas creatinine increased, compared with pre-acclimation values. The heart rate was significantly reduced compared with pre-acclimation values. QT-RR relation followed the square root regression function, which means modification of Bazett's QTc formula can be applied even if the ECG is recorded under restraint. In conclusion, 6-month acclimation was effective for stabilizing the blood data and for allowing accurate evaluation of the ECG even under restraint. Current results show that an acclimation period at least 3 months may be necessary prior to using rhesus monkeys for chronic studies. PMID- 15271068 TI - Descriptive urological record of chimpanzees (Pan troglodytes) in the wild and limitations associated with using multi-reagent dipstick test strips. AB - Ten urine chemistry parameters were measured on 74 voided urine samples from 34 wild chimpanzees (Pan troglodytes). Multi-reagent urine dipstick tests were performed and results determined using colorimetric scales. Urine pH measured between 8 and 9 units in 91% of the chimpanzees. Test pads detected protein, erythrocytes, leukocyte esterase activity, and nitrites, ketones and bilirubin in 47, 32, 29, and <10% of the chimpanzees, respectively. No apparent association between positive test results for blood in adult females and reproductive status was found. Overall, 17 of the 34 chimpanzees had positive urine test results for protein, hemoglobin, erythrocytes, leukocytes, nitrites, ketones, and/or bilirubin. Dipstick urinalysis alone is an unreliable method for assessing health and physiological status of wild chimpanzees. However, if combined with other diagnostics it could prove to be a valuable health-monitoring tool. Limitations associated with this methodology need to be considered when interpreting urinary dipstick test results. PMID- 15271069 TI - White monkey syndrome in infant baboons (Papio species). AB - Over 23 months, zinc toxicosis was diagnosed in 35 baboons aged 5-12 months in one galvanized metal and concrete cage complex with conditions that led to excessive exposure to environmental zinc. Clinical signs included reduced pigmentation of hair, skin, and mucous membranes (whiteness), alopecia, dehydration, emaciation, cachexia, dermatitis, diarrhea and, in six cases, severe gangrenous dermatitis of extremities. The syndrome was characterized by pancytopenia, elevated zinc and low copper serum concentrations, low vitamin D and bone-specific alkaline phosphatase levels, and atypical myelomonocytic proliferation of bone marrow. This syndrome emphasizes the importance of proper husbandry and cage design and indicates the potential of infant baboons as a model to study the effects of excessive zinc on development. This is the first report describing the epidemiologic and clinical presentation of zinc toxicosis in infant baboons in captivity. PMID- 15271070 TI - Sebaceous gland adenoma in a rhesus monkey (Macaca mulatta). AB - A tumor mass was identified below the shoulder region of a 5-year-old male rhesus monkey (Macaca mulatta). The mass was excised and diagnosed as sebaceous gland adenoma based on the microscopic findings. Morphologically it appeared as an elevated, dome-shaped, circumscribed mass of 3.6 x 2.8 x 3.2 cm in dimension with tan speckled color. Histologically, the tumor was composed of mature, sebaceous cells (sebocytes), basal cells arranged in a mass of irregular shapes and sizes, with a characteristic appearance of poly or multilobular structure. Sebocytes were well differentiated with foamy cytoplasm in the center of the lobules and poorly or undifferentiated densely staining basal cells in the periphery of the lobules. Cellular changes in the adjacent lymph node included hyperplasia of plasma cells, macrophages and lymphoid elements with typical mitosis. PMID- 15271071 TI - Ecological diversification in a group of Indomalayan pitvipers (Trimeresurus): convergence in taxonomically important traits has implications for species identification. AB - We analyse molecular and phenotypic evolution in a group of taxonomically problematic Indomalayan pitvipers, the Trimeresurus sumatranus group. Mitochondrial DNA sequencing provides a well-resolved phylogeny, with each species representing a distinct lineage. Multivariate morphological analysis reveals a high level of phenotypic differentiation, which is congruent between the sexes but does not reflect phylogenetic history. An adaptive explanation for the observed pattern of differentiation is supported by independent contrasts analysis, which shows significant correlations between current ecology and the characters that most account for the variation between taxa, including those that are presently used to identify the species. Reduced precipitation and altitude, and increased temperature, are correlated with higher numbers of scales on the head, body and tail. It is hypothesized that scale number plays an important role in heat and water exchange by influencing the area of exposed of interstitial skin, and that colour pattern variation reflects selection pressures involving camouflage and thermoregulation. Ecological convergence in traits used for classification is found to have important implications for species identification where taxa are distributed over varying environments. PMID- 15271072 TI - Testing for microevolution in body size in three blue tit populations. AB - Quantifying the genetic variation and selection acting on phenotypes is a prerequisite for understanding microevolutionary processes. Surprisingly, long term comparisons across conspecific populations exposed to different environments are still lacking, hampering evolutionary studies of population differentiation in natural conditions. Here, we present analyses of additive genetic variation and selection using two body-size traits in three blue tit (Parus caeruleus) populations from distinct habitats. Chick tarsus length and body mass at fledging showed substantial levels of genetic variation in the three populations. Estimated heritabilities of body mass increased with habitat quality. The poorer habitats showed weak positive selection on tarsus length, and strong positive selection on body mass, but there was no significant selection on either trait in the good habitat. However, there was no evidence of any microevolutionary response to selection in any population during the study periods. Potential explanations for this absence of a response to selection are discussed, including the effects of spatial heterogeneity associated with gene flow between habitats. PMID- 15271073 TI - Sex-ratio distorter of Drosophila simulans reduces male productivity and sperm competition ability. AB - The aim of the present study was to determine whether the effects of sex-ratio segregation distorters on the fertility of male Drosophila simulans can explain the contrasting success of these X-linked meiotic drivers in different populations of the species. We compared the fertility of sex-ratio and wild-type males under different mating conditions. Both types were found to be equally fertile when mating was allowed, with two females per male, during the whole period of egg laying. By contrast sex-ratio males suffered a strong fertility disadvantage when they were offered multiple mates for a limited time, or in sperm competition conditions. In the latter case only, the toll on male fertility exceeded the segregation advantage of the distorters. These results indicate that sex-ratio distorters can either spread or disappear from populations, depending on the mating rate. Population density is therefore expected to play a major role in the evolution of sex-ratio distorters in this Drosophila species. PMID- 15271074 TI - Evolutionary genetics of dorsal wing colour in Colias butterflies. AB - The evolution of butterfly wing colouration is strongly affected by its multiple functions and by the correlated evolution of wing colour elements. Both factors may prevent local adaptation to ecological conditions. We investigated one aspect of wing colouration, the degree of dorsal wing melanization, in the butterfly Colias philodice eriphyle across an elevational gradient and its correlation with another aspect of wing colouration, ventral wing melanization. Dorsal wing melanization increased with elevation and these differences persisted in a common environment. Full-sibling analysis revealed high heritability for males but only intermediate heritability for females. The correlation between ventral and dorsal melanization showed significant elevational and sex-specific differences. In males the two traits were highly correlated, whereas in females the strength of the correlation decreased with increasing elevation. We conclude that uncoupling of ventral and dorsal melanization has evolved in females but not in males and discuss possible mechanisms underlying uncoupling. PMID- 15271075 TI - Interspecific aggression and character displacement in the damselfly Calopteryx splendens. AB - Problems in species recognition are thought to affect the evolution of secondary sexual characters mainly through avoidance of maladaptive hybridization. Another, but much less studied avenue for the evolution of sexual characters due to species recognition problems is through interspecific aggression. In the damselfly, Calopteryx splendens, males have pigmented wing spots as a sexual character. Large-spotted males resemble males of another species, Calopteryx virgo, causing potential problems in species recognition. In this study, we investigate whether there is character displacement in wing spot size and whether interspecific aggression could cause this pattern. We found first that wing spot size of C. splendens in populations decreased with increasing relative abundance of C. virgo. Secondly, C. virgo males were more aggressive towards large- than small-spotted C. splendens males. Thirdly, in interspecific contests C. virgo males had better territory holding ability than C. splendens males. These results suggest that interspecific aggression may have caused character displacement in wing spot size of C. splendens, because the intensity of aggression towards large spotted males is likely to increase with relative abundance of C. virgo males. Thus, interspecific aggression may be an evolutionarily significant force that is able to cause divergence in secondary sexual characters. PMID- 15271076 TI - The interaction between reproductive lifespan and protandry in seasonal breeders. AB - The timing and duration of reproductive activities are highly variable both at the individual and population level. Understanding how this variation evolved by natural selection is fundamental to understanding many important aspects of an organism's life history, ecology and behaviour. Here, we combine game theoretic principles governing reproductive timing and the evolutionary theory of senescence to study the interaction between protandry (the earlier arrival or emergence of males to breeding areas than females) and senescence in seasonal breeders. Our general model applies to males who are seeking to mate as frequently as possible over a relatively short period, and so is relevant to many organisms including annual insects and semelparous vertebrates. The model predicts that protandry and maximum reproductive lifespans should increase in environments characterized by high survival and by a low competitive cost of maintaining the somatic machinery necessary for survival. In relatively short seasons under these same conditions, seasonal declines in the reproductive lifespans of males of equivalent quality will be evolutionarily stable. However, over a broad range of potential values for daily survival and maintenance cost, reproductive lifespan is expected to be relatively short and constant throughout a large fraction of the season. We applied the model to sockeye (or kokanee) salmon Oncorhynchus nerka and show that pronounced seasonal declines in reproductive lifespan, a distinctive feature of semelparous Oncorhynchus spp., is likely part of a male mating strategy to maximize mating opportunities. PMID- 15271077 TI - Experimental reduction of codon bias in the Drosophila alcohol dehydrogenase gene results in decreased ethanol tolerance of adult flies. AB - The ethanol tolerance of adult transgenic flies of Drosophila containing between zero and ten unpreferred synonymous mutations that reduced codon bias in the alcohol dehydrogenase (Adh) gene was assayed. As the amino acid sequences of the ADH protein were identical in the four genotypes assayed, differences in ethanol tolerance were due to differences in the abundance of ADH protein, presumably driven by the effects of codon bias on translational efficiency. The ethanol tolerance of genotypes decreased with the number of unpreferred synonymous mutations, and a positive correlation between ADH protein abundance and ethanol tolerance was observed. This work confirms that the fitness effects of unpreferred synonymous mutations that reduce codon bias in a highly expressed gene are experimentally measurable in Drosophila melanogaster. PMID- 15271078 TI - Hybridization and regional sex ratios in Nemophila menziesii. AB - I tested whether a region of high female frequencies in the gynodioecious plant, Nemophila menziesii, may be due to hybridization between regionally distributed populations with different corolla colours. I crossed plants in the greenhouse from populations with different corolla colours and found that hybrid crosses yielded higher frequencies of females than within-colour crosses. In the field, I found that populations with high female frequencies had intermediate mean corolla colours and higher variance in corolla colour, two traits suggesting hybridization. Nemophila menziesii has nuclear-cytoplasmic sex inheritance, thus if populations with different corolla colours are fixed for different male sterile cytoplasms and matching nuclear restorer alleles, hybridization between populations with different corolla colour should yield high frequencies of females. Two populations that are all hermaphroditic in the field segregated females in hybrid crosses suggesting that field populations may contain sex ratio distorters but appear undistorted, a prediction of genomic conflict theory. PMID- 15271079 TI - Fine-scale genetic structure and gene dispersal in Centaurea corymbosa (Asteraceae) I. Pattern of pollen dispersal. AB - Pollen dispersal was characterized within a population of the narrowly endemic perennial herb, Centaurea corymbosa, using exclusion-based and likelihood-based paternity analyses carried out on microsatellite data. Data were used to fit a model of pollen dispersal and to estimate the rates of pollen flow and mutation/genotyping error, by developing a new method. Selfing was rare (1.6%). Pollen dispersed isotropically around each flowering plant following a leptokurtic distribution, with 50% of mating pairs separated by less than 11 m, but 22% by more than 40 m. Estimates of pollen flow lacked precision (0-25%), partially because mutations and/or genotyping errors (0.03-1%) could also explain the occurrence of offspring without a compatible candidate father. However, the pollen pool that fertilized these offspring was little differentiated from the adults of the population whereas strongly differentiated from the other populations, suggesting that pollen flow rate among populations was low. Our results suggest that pollen dispersal is too extended to allow differentiation by local adaptation within a population. However, among populations, gene flow might be low enough for such processes to occur. PMID- 15271080 TI - Variation in natural selection for growth and phlorotannins in the brown alga Fucus vesiculosus. AB - Directional selection for plant traits associated with resistance to herbivory tends to eliminate genetic variation in such traits. On the other hand, balancing selection arising from trade-offs between resistance and growth or spatially variable selection acts against the elimination of genetic variation. We explore both the amount of genetic variation and variability of natural selection for growth and concentration of phenolic secondary compounds, phlorotannins, in the brown alga Fucus vesiculosus. We measured variation in selection at two growing depths and two levels of nutrient availability in algae that had faced two kinds of past growing environments. Genetic variation was low for growth but high for phlorotannins. The form and strength of selection for both focal traits depended on the past growing environment of the algae. We found strong directional selection for growth rate in algae previously subjected to higher ultraviolet radiation, but not in algae previously subjected to higher nutrient availability. Stabilizing selection for growth occurred especially in the deep growing environment. Selection for phlorotannins was generally weak, but in some past environment-current-environment combinations we detected either directional selection against phlorotannins or stabilizing selection. Thus, phlorotannins are not selectively neutral but affect the fitness of F. vesiculosus. In particular, there may be a fitness cost of producing phlorotannins, but the realization of such a cost varies from one environment to another. Genetic correlations between selective environments were high for growth but nonexistent for phlorotannins, emphasizing the high phenotypic plasticity of phlorotannin production. The highly heterogeneous selection, including directional, stabilizing, and spatially variable selection as well as temporal change in selection due to responses to past environmental conditions, probably maintains a high amount of genetic variation in phlorotannins. Such variation provides the potential for rapid evolutionary response of phlorotannins under directional selection. PMID- 15271081 TI - Condition-dependent traits and the capture of genetic variance in male advertisement song. AB - The occurrence of additive genetic variance (VA) for male sexual traits remains a major problem in evolutionary biology. Directional selection normally imposed by female choice is expected to reduce VA greatly, yet recent surveys indicate that a substantial amount remains in many species. We addressed this problem, also known as the 'lek paradox', in Achroia grisella (Lepidoptera: Pyralidae), an acoustic moth in which males advertise to females with a pulsed ultrasonic song. Using a standard half-sib/full-sib breeding design, we generated F1 progeny from whom we determined VA and genetic covariance (COVA) among seven traits: three song characters, an overall index of song attractiveness, nightly singing period, adult lifespan, and body mass at adult eclosion. Because A. grisella neither feed nor drink as adults, the last trait, eclosion body mass, is considered a measure of 'condition'. We found significant levels of VA and narrow-sense heritabilities (h2) for all seven traits and significant genetic correlations (= COVAi,j / radical (VA i x VA j)) between most pairs of traits (i, j). Male attractiveness was positively correlated with body mass (condition), adult lifespan, and nightly singing period, which we interpret as an energy constraint preventing males in poor condition from singing attractively, from singing many hours per night, and from surviving an extended lifespan. The positive genetic correlation (r = 0.79) between condition and attractiveness, combined with significant levels of VA for both traits, indicates that much of the variation in male song can be explained by VA for condition. Finally, we discuss the morphological and physiological links between condition and song attractiveness, and the ultimate factors that may maintain VA for condition. PMID- 15271082 TI - Behavioural interactions, kin and disease susceptibility in the bumblebee Bombus terrestris. AB - Behavioural interactions are often analysed in terms of their costs and benefits to the actors [Hamilton, (1964) J. Theor. Biol.7 1-16; Gadagkar, (1993) Trends Ecol. Evol.8 232-234; Foster et al., (2001) Ann. Zool. Fenn.38 229-238]. Using the bumblebee Bombus terrestris, we wish to distinguish between two possible determinants of interaction behaviour between conspecifics, namely kin-directed behaviour that reflects genetic distance between individuals, or, alternatively, interactions guided by a functional distance between individuals, specifically, with respect to disease susceptibility. We find no relationship between contact rate of individuals and the genetic distance of their respective colonies. Interestingly, we do find a significant negative correlation between contact rate and the distance between the two colonies in susceptibility to a spectrum of parasite strains. This cannot be explained by either of the a priori alternatives so we propose two further testable hypotheses to explain our results. PMID- 15271083 TI - The cost of immunity in the yellow fever mosquito, Aedes aegypti depends on immune activation. AB - Although host immunity offers the obvious benefit of reducing parasite infection, it is often traded-off with other fitness components. We investigated whether the cost of an immune response in the yellow fever mosquito, Aedes aegypti, is modulated by the antigen that activates the melanization immune response. Thus, one of three different novel antigens were injected into the mosquito's thorax- either a glass bead, a negatively charged (C-25) Sephadex bead, or a neutral (G 25) Sephadex bead--and fecundity and bead melanization were observed. Glass beads are immunologically inert and were therefore used as an inoculation control. The fecundity of mosquitoes inoculated with these beads did not differ from the fecundity of mosquitoes that did not melanize negatively charged or neutral beads. The ability of A. aegypti to melanize negatively charged Sephadex beads was associated with reduced fecundity, showing a clear cost of immunity. In contrast, melanization of the neutral beads was quite strong but had no effect on fecundity. Thus, the cost of what appeared to be the same immune response- melanization of a bead--depended on the type of bead that stimulated the immune system. Such differences might help to explain variation of immune efficacy against different parasites in natural populations. PMID- 15271084 TI - Swift laboratory thermal evolution of wing shape (but not size) in Drosophila subobscura and its relationship with chromosomal inversion polymorphism. AB - Latitudinal clinal variation in wing size and shape has evolved in North American populations of Drosophila subobscura within about 20 years since colonization. While the size cline is consistent to that found in original European populations (and globally in other Drosophila species), different parts of the wing have evolved on the two continents. This clearly suggests that 'chance and necessity' are simultaneously playing their roles in the process of adaptation. We report here rapid and consistent thermal evolution of wing shape (but not size) that apparently is at odds with that suggestion. Three replicated populations of D. subobscura derived from an outbred stock at Puerto Montt (Chile) were kept at each of three temperatures (13, 18 and 22 degrees C) for 1 year and have diverged for 27 generations at most. We used the methods of geometric morphometrics to study wing shape variation in both females and males from the thermal stocks, and rates of genetic divergence for wing shape were found to be as fast or even faster than those previously estimated for wing size on a continental scale. These shape changes did not follow a neat linear trend with temperature, and are associated with localized shifts of particular landmarks with some differences between sexes. Wing shape variables were found to differ in response to male genetic constitution for polymorphic chromosomal inversions, which strongly suggests that changes in gene arrangement frequencies as a response to temperature underlie the correlated changes in wing shape because of gene inversion linkage disequilibria. In fact, we also suggest that the shape cline in North America likely predated the size cline and is consistent with the quite different evolutionary rates between inversion and size clines. These findings cast strong doubts on the supposed 'unpredictability' of the geographical cline for wing traits in D. subobscura North American colonizing populations. PMID- 15271085 TI - Extinction of the acoustic startle response in moths endemic to a bat-free habitat. AB - Most moths use ears solely to detect the echolocation calls of hunting, insectivorous bats and evoke evasive flight manoeuvres. This singularity of purpose predicts that this sensoribehavioural network will regress if the selective force that originally maintained it is removed. We tested this with noctuid moths from the islands of Tahiti and Moorea, sites where bats have never existed and where an earlier study demonstrated that the ears of endemic species resemble those of adventives although partially reduced in sensitivity. To determine if these moths still express the anti-bat defensive behaviour of acoustic startle response (ASR) we compared the nocturnal flight times of six endemic to six adventive species in the presence and absence of artificial bat echolocation sounds. Whereas all of the adventive species reduced their flight times when exposed to ultrasound, only one of the six endemic species did so. These differences were significant when tested using a phylogenetically based pairwise comparison and when comparing effect sizes. We conclude that the absence of bats in this habitat has caused the neural circuitry that normally controls the ASR behaviour in bat-exposed moths to become decoupled from the functionally vestigial ears of endemic Tahitian moths. PMID- 15271086 TI - Cheating is not always punished: killer female plants and pollination by deceit in the dwarf palm Chamaerops humilis. AB - Because the interests of mutualists are not perfectly aligned, conflicts between partners often arise, rendering mutualism unstable by allowing the evolution of cheating. The dwarf palm Chamaerops humilis is engaged in a nursery pollination mutualism with a specific weevil Derelomus chamaeropsis. In exchange for pollen dispersal, dwarf palms provide pollinators with food, shelter and egg-laying sites, but pollinators can develop only within male inflorescences. Here we show that weevils lay eggs in female inflorescences but processes associated with fruit development prevent larval development. The cost imposed by developing larvae probably differs between male and female plants, explaining why only females defend their inflorescences. Female palms thus cheat their pollinating weevil, and pollinators are expected to 'punish' (avoid) them. We found no evidence for such punishment: weevils visit female plants and the duration of visits to male and female inflorescences does not differ. Thus mutualists do not always co-operate and cheating may not be necessarily punished. PMID- 15271087 TI - Evolution of eusociality and the soldier caste in termites: a validation of the intrinsic benefit hypothesis. AB - In termites the evolution of reproductive altruism is not based on a particularly high relatedness between nestmates. For the evolution and maintenance of the ancestral sterile soldier caste, the benefits generated by the soldiers' presence must compensate the loss of the soldiers' reproductive potential. To study the impact of soldiers on colony's fitness, we manipulated the proportion of soldiers to nonsoldiers in colonies of the dry-wood termite Cryptotermes secundus.'Soldier less' colonies were obtained by removing soldiers and inhibiting their development with an extract of soldier heads. The colonies were set up for 1 year in experimental nests in the field. 'Soldier-less' colonies produced fewer soldiers. The reduction of soldiers neither affected colony survival nor helper growth, but fewer dispersing sexuals were produced in 'soldier-less' than in control colonies. This confirms what was only supposed so far, that in termites soldiers are maintained for their intrinsic benefit to cost ratio. PMID- 15271088 TI - 'Anti-bee' and 'pro-bird' changes during the evolution of hummingbird pollination in Penstemon flowers. AB - Floral phenotypes may be as much the result of selection for avoidance of some animal visitors as selection for improving the interaction with better pollinators. When specializing on hummingbird-pollination, Penstemon flowers may have evolved to improve the morphological fit between bird and flower, or to exclude less-efficient bees, or both. We hypothesized how such selection might work on four floral characters that affect the mechanics of pollen transfer: anther/stigma exsertion, presence of a lower corolla lip, width of the corolla tube, and angle of flower inclination. We surgically modified bee-pollinated P. strictus flowers changing one trait at a time to make them resemble hummingbird pollinated P. barbatus flowers, and measured pollen transfer by bumblebees and hummingbirds. Results suggest that, apart from 'pro-bird' adaptations, specific 'anti-bee' adaptations have been important in shaping hummingbird-flowers. Moreover, some trait changes may have been selected for only if changing in concert with other traits. PMID- 15271089 TI - Constrained sex allocation in a parasitoid due to variation in male quality. AB - The theory of constrained sex allocation posits that when a fraction of females in a haplodiploid population go unmated and thus produce only male offspring, mated females will evolve to lay a female-biased sex ratio. I examined evidence for constrained sex ratio evolution in the parasitic hymenopteran Uscana semifumipennis. Mated females in the laboratory produced more female-biased sex ratios than the sex ratio of adults hatching from field-collected eggs, consistent with constrained sex allocation theory. However, the male with whom a female mated affected her offspring sex ratio, even when sperm was successfully transferred, suggesting that constrained sex ratios can occur even in populations where all females succeed in mating. A positive relationship between sex ratio and fecundity indicates that females may become sperm-limited. Variation among males occurred even at low fecundity, however, suggesting that other factors may also be involved. Further, a quantitative genetic experiment found significant additive genetic variance in the population for the sex ratio of offspring produced by females. This has only rarely been demonstrated in a natural population of parasitoids, but is a necessary condition for sex ratio evolution. Finally, matings with larger males produced more female-biased offspring sex ratios, suggesting positive selection on male size. Because the great majority of parasitic hymenoptera are monandrous, the finding of natural variation among males in their capacity to fertilize offspring, even after mating successfully, suggests that females may often be constrained in the sex allocation by inadequate number or quality of sperm transferred. PMID- 15271090 TI - Relatedness affects competitive performance of a parasitic plant (Cuscuta europaea) in multiple infections. AB - Theoretical models predict that parasite relatedness affects the outcome of competition between parasites, and the evolution of parasite virulence. We examined whether parasite relatedness affects competition between parasitic plants (Cuscuta europaea) that share common host plants (Urtica dioica). We infected hosts with two parasitic plants that were either half-siblings or nonrelated. Relative size asymmetry between the competing parasites was significantly higher in the nonrelated infections compared to infections with siblings. This higher asymmetry was caused by the fact that the performance of some parasite genotypes decreased and that of others increased when grown in multiple infections with nonrelated parasites. This result agrees with the predictions of theories on the evolution of parasite virulence: to enhance parasite transmission, selection may favour reduced competition with genetically related parasites in hosts infected by several genotypes. However, in contrast to the most common predictions, nonrelated infections were not more virulent than the sibling infections. PMID- 15271091 TI - Delaying evolution of insect resistance to transgenic crops by decreasing dominance and heritability. AB - The refuge strategy is used widely for delaying evolution of insect resistance to transgenic crops that produce Bacillus thuringiensis (Bt) toxins. Farmers grow refuges of host plants that do not produce Bt toxins to promote survival of susceptible pests. Many modelling studies predict that refuges will delay resistance longest if alleles conferring resistance are rare, most resistant adults mate with susceptible adults, and Bt plants have sufficiently high toxin concentration to kill heterozygous progeny from such matings. In contrast, based on their model of the cotton pest Heliothis virescens, Vacher et al. (Journal of Evolutionary Biology, 16, 2003, 378) concluded that low rather than high toxin doses would delay resistance most effectively. We demonstrate here that their conclusion arises from invalid assumptions about larval concentration-mortality responses and dominance of resistance. Incorporation of bioassay data from H. virescens and another key cotton pest (Pectinophora gossypiella) into a population genetic model shows that toxin concentrations high enough to kill all or nearly all heterozygotes should delay resistance longer than lower concentrations. PMID- 15271093 TI - Phenotypic correlations among fitness and its components in a population of the housefly. AB - An individual's or a population's fitness is the result of a large number of interacting life history traits and the environment. Little information is available on the phenotypic correlations among fitness components and fitness itself, especially outside of Drosophila melanogaster. We also lack detailed information on trade-offs among life history traits. Here we present the relationship between adult progeny production and eight components of fitness, as well as some observed trade-offs between life history traits in the housefly (Musca domestica). We briefly discuss some of the ramifications of these relationships. PMID- 15271094 TI - Sources of stochasticity in models of sex allocation in spatially structured populations. AB - There are many ways to include stochastic effects in models of sex allocation evolution. These include variability in the number of mating partners and fecundity in a rich literature that goes back 20 years. The effects of variance in the fecundity and number of mating partners have typically been considered separately from the stochastic effects of mortality. However, I show that these processes produce mathematically equivalent models with subtly different biological details. These scenarios differ in the way that information becomes available to individuals because the parents often have information on mating partners while they are making sex allocation decisions, but must make these decisions before brood mortality takes place. This makes it possible to test which mechanism, stochastic mortality or variation in mating partners, is responsible for observed sex ratios. Alternatively, asymmetric variance between sexual functions can cause skewed sex allocation, even in the absence of local mate competition. This allows the evolution of either female- or male-biased sex ratios depending on which sexual function is more variable. PMID- 15271096 TI - Cardiac arrest: a role for vasopressin. PMID- 15271097 TI - A simple and rapid high-performance liquid chromatographic (HPLC) method for 5 fluorouracil (5-FU) assay in plasma and possible detection of patients with impaired dihydropyrimidine dehydrogenase (DPD) activity. AB - BACKGROUND: Dihydropyrimidine dehydrogenase (DPD) gene polymorphism may lead to severe toxicity with 5-fluorouracil (5-FU), a major anticancer drug extensively used in clinical oncology. Drug monitoring combined with early detection of patients at risk would enable timely dose adaptation so as to maintain drug concentrations within a therapeutic window. However, the best method to identify such patients remains to be determined. OBJECTIVE: The aim of this study was to develop a rapid and simple high-performance liquid chromatographic (HPLC) method for estimating uracil/dihydrouracil (U/UH2) ratio in plasma, as an index of DPD status, and for assaying 5-FU as part of drug level monitoring. METHOD: Assay of 5-FU, and U/UH2 detection were performed on a HPLC system equipped with UV detector. Analytes were separated at room temperature using a 5 microm particles, 25 cm RP-18 X-Terra column. The mobile-phase consisted of a KH(2)PO(4) salt solution (0.05 m) + 0.1% triethylamine (TEA) pumped at 0.4 mL/min. Detection of 5 FU and 5-bromouracil were performed at 254 nm; U and UH2 elution was monitored at 210 nm. RESULTS: The method was sensitive and specific for assaying 5-FU within the 5-500 ng/mL concentration range, which covers exposure levels currently met in clinical practice. The method was simple, and relatively cheap, and rapid, with an analytical run time of about 30 min. Data from a patient with 5-FU toxicity suggest that the method was capable of identifying DPD metabolic phenotype in cancer patients, based on measurement of plasma U/UH2 ratio. CONCLUSION: The method described should be suitable both for detecting patients at high risk of 5-FU toxicity, and for drug level monitoring during chemotherapy. PMID- 15271098 TI - Repeatability of measurements of the initial distribution volume of glucose in haemodynamically stable patients. AB - AIMS: The initial distribution volume of glucose (IDVG) has been proposed to provide a useful tool to estimate the central extracellular fluid volume. The purpose of this study was to determine the repetition interval of two consecutive measurements in haemodynamically stable patients without presence of recent changes in fluid status. METHODS: Twenty-nine patients admitted to the general intensive care unit of the University of Hirosaki Hospital were entered into this study. After achieving a haemodynamically stable state in each patient regardless of an infusion of vasoactive drugs, two glucose challenges at an interval of either 30 or 60 min, were carried out to calculate the IDVG. The IDVG was calculated using a one-compartment model after intravenous administration of glucose (5 g) followed by serial arterial blood sampling. RESULTS: Although plasma glucose levels immediately before the second glucose challenge in either group were increased compared with those of the first challenge (P < 0.001, respectively), the bias of the IDVG measurements was 0.08 +/- 0.32 L (SD) for the 30-min group and -0.19 +/- 0.28 L for the 60-min group. CONCLUSIONS: Our results indicate that IDVG determinations can be reliably repeated within a minimum interval of 30 min. PMID- 15271099 TI - Epoetin in haemodialysis patients: impact of change from subcutaneous to intravenous routes of administration. AB - BACKGROUND AND OBJECTIVE: Cases of erythroblastopenia, as an adverse effect to epoetin (EPREX), led to its use being restricted in Europe to the intravenous route (i.v.) since July 2002, in patients with chronic renal insufficiency. This work aimed at investigating the biological, pharmaceutical and economic impacts of this change in policy. METHODS: We retrospectively compared the characteristics of 99 haemodialysis patients treated with epoetin at the time of the recommendation (July 2002) and 5 months after the policy change (November 2002). RESULTS: In July 2002, 69 patients who were receiving EPREX subcutaneously (s.c./i.v. group) changed to the i.v. route of administration. Thirty other patients were already on i.v. epoetin (i.v. group). During the study period, the dose of epoetin increased significantly in the s.c./i.v. group but not in the i.v. group (46.83 +/- 10.20 UI/kg/week vs. 2.17 +/- 20.14 UI/kg/week respectively). This increased dosage was accounted for by a subgroup of 42 patients in the s.c./i.v. group while the others had dosage variations similar to those observed in the i.v. group. There were no significant clinical and biological changes associated with this change in route of administration. However, the change in policy led to the haemodialysis ward incurring an additional cost of 265,905 Euro (+32.7%) or an average annual extra cost of 1841 +/- 401 Euro per patient. CONCLUSION: Changing the route of administration of EPREX from the i.v. to the subcutaneous route required an increase in dosage and in substantial additional cost. PMID- 15271100 TI - The quality of essential antimicrobial and antimalarial drugs marketed in Rwanda and Tanzania: influence of tropical storage conditions on in vitro dissolution. AB - OBJECTIVE: The quality of 33 formulations of essential antimicrobial and antimalarial drugs (amoxicillin capsules, metronidazole tablets, sulfamethoxazole/trimethoprim tablets, quinine tablets and sulphadoxine/pyrimethamine tablets) marketed in Rwanda and Tanzania was assessed and the influence of tropical storage conditions on potency and in vitro dissolution investigated. METHODS: Drug content and in vitro dissolution were determined immediately after purchase and during 6-month storage under simulated tropical conditions (75% relative humidity, 40 degrees C) using the methods described in the USP 24 monographs on the drugs concerned. RESULTS AND DISCUSSION: At the time of purchase, the drug content of all the formulations was within the limits recommended by the USP 24, but after 6-month storage, the drug content of one sulfamethoxazole/trimethoprim and one quinine formulation were found to be substandard. Immediately after purchase, four formulations (three sulfamethoxazole/trimethoprim and one sulphadoxine/pyrimethamine combination) failed the USP 24 dissolution test. Except for three metronidazole and one quinine formulations, dissolution tests performed after 6 months of storage under simulated tropical conditions showed that drug release remained within the USP 24 recommended values. CONCLUSION: In both countries, essential drug formulations met pharmacopoeial potency requirements, but some had a poor in vitro drug release profiles. Some of the formulations tested were not stable upon storage under simulated tropical conditions. PMID- 15271101 TI - Developing the Medication Change Questionnaire. AB - OBJECTIVE: People who seek non-pharmaceutical interventions are often motivated by a desire to avoid or reduce orthodox medication. Effectiveness research in these areas needs to measure change in medication as an outcome. We set out to develop a data collection tool that is sensitive to changes in individual drug use over time. METHOD: A multi-disciplinary team designed, piloted, and revised the Medication Change Questionnaire (MCQ) on two occasions, and used qualitative interviews to understand the patient's perspective and ensure that the final product accurately reflected the medication that patients were taking. Thirty patients in one general practice completed the questionnaire on two occasions and a purposive sample of 14 were interviewed. The design sought to enable patients to record all their ingested medication accurately, both prescription and over the-counter drugs, over a period of 7 days. It was designed to be administered face-to face on the first occasion, and to be self-completed on subsequent occasions. RESULTS: In considering in detail what medication was taken each day, the interview data fully correlated with the MCQ data in only one of the five people who were interviewed after completing the first draft of the questionnaire, but in eight of the nine people who completed the second draft. Of these eight people all but one had made some change to their medication, either by stopping or starting a drug, varying the dose of a drug or always taking one or more drugs in varying doses. The interviews demonstrated the complex and individual ways that people took their medicines, and the disparity between what was prescribed and what was actually taken. The qualitative data were also useful for reflecting on the potential advantages and disadvantages of other data collection methods, such as single medication questions and pill counts. CONCLUSION: By involving patients at every level of research, we have developed a questionnaire that enables people to record their medication use accurately over a 7-day period, and to demonstrate changes in medication over time. Further work is required to assess its acceptability by different patient populations and its feasibility in terms of completion rates over longer periods of repeated use. We believe the MCQ to be an improvement over the variety of ad-hoc tools used in the past and it is freely available from the authors. PMID- 15271102 TI - High dose vancomycin for osteomyelitis: continuous vs. intermittent infusion. AB - OBJECTIVES: To compare the efficacy, ease of use and safety of intermittent vancomycin infusion (IVI) and continuous vancomycin infusion (CVI) in high-dose therapy of osteomyelitis. METHODS: Forty-four patients with an osteomyelitis requiring vancomycin for more than 4 weeks were prospectively included, 21 receiving IVI and 23, CVI. The target serum concentration of vancomycin was 20-25 mg/L. Pharmacokinetics, adverse effects, and clinical efficacy were recorded. RESULTS: The mean daily vancomycin dosing was the same in the two groups, but the serum vancomycin concentrations (trough or plateau) were lower in the IVI group than the CVI group (21.7 +/- 9.3 and 26.0 +/- 6.1 mg/L, respectively; P < 0.0001). The target concentrations were achieved quicker with CVI, and daily dosing was changed more frequently in the IVI group. After reaching the target, variability of vancomycin serum concentration (trough or plateau concentrations) was higher in the IVI group than in CVI group (standard deviation 7.9 mg/L vs. 5.6 mg/L, respectively; P = 0.001). CVI did not show clinical superiority, but adverse drug effects were more frequent in the IVI group as compared with the CVI group, 9 (42.9%) and 2 (8.7%), respectively (P = 0.03). Survival multiple regression using Cox's proportional hazard model showed that IVI (RR = 5.9, P = 0.03) and osteomyelitis of the foot (RR = 5.2, P = 0.01) were the only factors associated with adverse drug reactions leading to treatment termination. CONCLUSIONS: CVI is practical and effective, and may be a good alternative for patients requiring prolonged treatment with high vancomycin serum levels. PMID- 15271103 TI - A retrospective drug utilization evaluation of vancomycin usage in paediatric patients. AB - OBJECTIVE: To evaluate the appropriateness of use of vancomycin in paediatric patients at KK Women's and Children's Hospital, the major paediatric hospital in Singapore to identify potential problems in prescribing practices that may necessitate intervention to optimize vancomycin usage. METHODS: A retrospective drug utilization evaluation was performed for paediatric patients who received intravenous vancomycin from 1 June 1998 to 31 June 1999. The outcome measures were consistency of vancomycin indication with recommended guidelines, dosing regimens, microbiological data, monitoring of serum drug levels, renal function, clinical outcomes and adverse drug reactions (ADRs). RESULTS: A total of 96 cases was available for evaluation. Sixty-two (64.6%) courses of vancomycin were consistent with guidelines for indication of therapy. Eighty-six (89.6%) of the dosing regimen were consistent. All infusion times that were recorded (56.3%) were consistent with criteria. Of the patients treated with vancomycin for more than 1 day, peak and/or trough serum vancomycin levels were ordered for 70 cases. Of the 56 cases with paired levels ordered, 46 cases had at least one level that fell outside the therapeutic range. Nineteen (19.8%) cases of ADRs were documented. Fifty-eight (60.4%) cases received concurrent nephrotoxic drugs. However, a substantial portion of vancomycin courses were apparently not prescribed for appropriate indications, and there was poor recording of vancomycin administration information and sampling time. CONCLUSION: The majority of dosing regimens of vancomycin was consistent with guideline criteria. The most evident problem was the sub-optimal use of the monitoring of vancomycin serum levels. The information derived from this study may be used as a for further study and for the development of strategies for optimize vancomycin usage. PMID- 15271104 TI - Stabilization of a freeze-dried recombinant streptokinase formulation without serum albumin. AB - AIM: To evaluate the stability and content homogeneity of a new freeze-dried and albumin-free formulation of recombinant streptokinase (SKr) that has recently been approved by the Cuban National Center for the Quality Control of Medicaments. MATERIALS AND METHODS: The new formulation was stored at the intended recommended storage temperature of 4 degrees C, and under accelerated storage conditions (37 degrees C). The stability of the product was also examined after reconstitution and storage at room temperature (28 degrees C) for 24 h. Samples were periodically subjected to biological activity assays (S-2251 chromogenic-substrate method or in vitro clot-lysis assay), sodium dodecyl sulphate/polyacrylamide gel electrophoresis (SDS/PAGE), pyrogen and sterility testing, abnormal toxicity screening, organoleptic evaluation, and measurement of residual moisture and pH. RESULTS AND DISCUSSION: Accelerated storage (37 degrees C) data showed biochemical stability of SKr throughout the 6-month study with activity, remaining between 90 and 110% of its nominal value (0.75 x 10(6) IU/mL). SDS/PAGE-determined purity showed that SKr remained above 97 %. Furthermore, the formulation was non-pyrogenic, non-toxic, sterile and organoleptically acceptable. Real-time storage data confirmed the excellent biochemical long-term (30 months) stability of the new formulation of SKr. Comparison with other freeze-dried preparations showed that the new formulation was organoleptically better. The formulation was stable after reconstitution and storage at 28 degrees C for 24 h. The content homogeneity of this new formulation was also satisfactory. CONCLUSIONS: This study demonstrated the stability and the content homogeneity of this formulation, despite the absence of albumin as stabilizer. PMID- 15271105 TI - Comparing the fixed combination dorzolamide-timolol (Cosopt) to concomitant administration of 2% dorzolamide (Trusopt) and 0.5% timolol -- a randomized controlled trial and a replacement study. AB - PURPOSE: To compare the intraocular pressure (IOP) lowering effect of concomitant administration of 0.5% timolol and 2% dorzolamide and a fixed combination dorzolamide-timolol (Cosopt) To critically evaluate discrepancies between phase 3 clinical trials and prior replacement studies. DESIGN: A prospective, randomized, controlled clinical trial and a prospective, non-randomized comparative replacement trial. PARTICIPANTS/INTERVENTIONS: In a national multicentre trial, 131 patients were randomized to dorzolamide-timolol or a topical carbonic anhydrase inhibitor (CAI) and non-selective beta-blocker following a 1-month run in using the separate components. Peak (maximal drug effect) and trough (minimal drug effect) IOPs were measured at baseline and 1 month after treatment. The replacement therapy study enrolled 404 consecutive glaucoma patients using a non selective beta-blocker and dorzolamide and changed treatment to the fixed combination. Mean IOPs at the same time of day were compared before and 1 month after changeover. MAIN OUTCOME MEASURE: The main outcome measure was IOP, comparing baseline and on-therapy measurements at study conclusion between the two arms of the randomized trial and before and after switching therapy in the replacement trial. RESULTS: In the randomized trial, the mean baseline peak and trough IOPs were 18.4 and 21.0 mmHg in the group randomized to combination therapy and 17.6 and 19.8 mmHg in the dual drug group. After randomization and treatment for four weeks, the peak and trough IOPs were 17.6 and 19.5 mmHg in the combination group and 17.3 and 19.0 mmHg in the concomitant group. The percentage change in IOP was -3.2% at peak and -6.5% at trough for the combination and -0.3 and -3.2% for the concomitant group. These differences did not show statistical significance. In the replacement study, mean baseline IOP was 19.4 mmHg. Four weeks after initiation of treatment on the fixed combination, a significant additional IOP reduction of 1.7 mmHg (-8.8%) was observed (P < 0.0001). Overall, 81% of eyes exhibited equal or lower IOP on the fixed combination compared with concomitant therapy. CONCLUSION: The results of the randomized trial indicate that the fixed combination dorzolamide-timolol (Cosopt) was as effective as its components in controlling IOP, confirming results seen in phase 3 clinical trials. However, in the replacement study, utilization of the combination drug offered a statistically significant additional IOP reduction (P < 0.0001), which duplicates results from previous replacement studies. PMID- 15271106 TI - Determination of ciprofloxacin concentrations in human serum and urine by HPLC with ultraviolet and fluorescence detection. AB - OBJECTIVE: To develop a simple and sensitive high performance liquid chromatography method for the determination of ciprofloxacin concentrations in human serum and urine. METHOD: Serum proteins were removed by ultrafiltration through a filtering device after the addition of a displacing reagent. Urine samples were diluted with mobile phase prior to injection. Separation was achieved with a C18 reverse-phase column and using ultraviolet (UVD) and fluorescence detection (FD) for serum samples and UVD for urine samples. RESULTS: The quantitation limits of the assay were 20 ng/ml (FD) and 100 ng/ml (UVD) in serum and 1 microg/ml in urine. The assay was successfully applied to a pharmacokinetic study of ciprofloxacin in healthy volunteers. CONCLUSION: The method presented for ciprofloxacin assay in human serum and urine requires less sample clean up and is more sensitive than those reported in the literature. PMID- 15271107 TI - Determinants of prescribing patterns for type II diabetes at Primary Care Trust level. AB - OBJECTIVE: To estimate the effect of demography, deprivation and ethnicity on variations in prescribing of oral antidiabetics at the Primary Care Trust level. DESIGN: Cross-sectional survey. SETTING: All Primary Care Trusts in England. MAIN OUTCOME MEASURES: Percentage of variance explained. RESULTS: A model using measures of demography, ethnicity, deprivation and use of thiazolidinediones explained 59% of the variation in cost per registered patient although individual variables explained far less. CONCLUSIONS: This study shows the importance of using a multivariate approach in modelling the drivers of cost. The high use of thiazolidinediones is of concern in the light of NICE guidance. PMID- 15271108 TI - Effects of provider practice on functional independence in older adults. AB - OBJECTIVES: To examine provider determinants of new-onset disability in basic activities of daily living (ADLs) in community-dwelling elderly. DESIGN: Observational study. SETTING: King County, Washington. PARTICIPANTS: A random sample of 800 health maintenance organization (HMO) enrollees aged 65 and older participating in a prospective longitudinal cohort study of dementia and normal aging and their 56 primary care providers formed the study population. MEASUREMENTS: Incident ADL disability, defined as any new onset of difficulty performing any of the basic ADLs at follow-up assessments, was examined in relation to provider characteristics and practice style using logistic regression and adjusting for case-mix, patient and provider factors associated with ADL disability, and clustering by provider. RESULTS: Neither provider experience taking care of large numbers of elderly patients nor having a certificate of added qualifications in geriatrics was associated with patient ADL disability at 2 or 4 years of follow-up (adjusted odds ratio (AOR) for experience=1.29, 95% confidence interval (CI)=0.81-2.05; AOR for added qualifications=0.72, 95% CI=0.38-1.39; results at 4 years analogous). A practice style embodying traditional geriatric principles of care was not associated with a reduced likelihood of ADL disability over 4 years of follow-up (AOR for prescribing no high-risk medications=0.56, 95% CI=0.16-1.94; AOR for managing geriatric syndromes=0.94, 95% CI=0.40-2.19; AOR for a team care approach=1.35, 95% CI=0.66 2.75). CONCLUSION: Taking care of a large number of elderly patients, obtaining a certificate of added qualifications in geriatrics, and practicing with a traditional geriatric orientation do not appear to influence the development of ADL disability in elder, community dwelling HMO enrollees. PMID- 15271109 TI - A randomized, controlled trial of an intensive community nurse-supported discharge program in preventing hospital readmissions of older patients with chronic lung disease. AB - OBJECTIVES: To evaluate the effectiveness of an intensive community nurse (CN) supported discharge program in preventing hospital readmissions of older patients with chronic lung disease (CLD). DESIGN: Randomized, controlled trial. SETTING: Two acute hospitals in the same health region in Hong Kong. PARTICIPANTS: One hundred fifty-seven hospitalized patients aged 60 and older with a primary diagnosis of CLD and at least one hospital admission in the previous 6 months. INTERVENTION: CNs made home visits within 7 days of discharge, then weekly for 4 weeks and monthly until 6 months. CNs coordinated closely with a geriatric or respiratory specialist in hospital. Subjects had telephone access to CNs during normal working hours from Monday to Saturday. MEASUREMENTS: The primary outcome was the rate of unplanned readmission within 6 months. The secondary outcomes were the rate of unplanned readmission within 28 days, number of unplanned readmissions, hospital bed days, accident and emergency room attendance, functional and psychosocial status, and caregiver burden. RESULTS: One hundred forty hospitalized patients completed the trial. Intervention group subjects had a higher rate of unplanned readmission within 6 months than control group subjects (76% vs 62%, P=.080, chi2 test). There was no significant group difference in any of the secondary outcomes except that intervention group subjects did better on social handicap scores. CONCLUSION: There was no evidence that an intensive CN-supported discharge program can prevent hospital readmissions in older patients with CLD. PMID- 15271110 TI - Site of death in the hospital versus nursing home of Medicare skilled nursing facility residents admitted under Medicare's Part A Benefit. AB - OBJECTIVES: To determine factors that predict site of death (hospital vs nursing home (NH)), related costs, and geographic variation in site of death of NH residents admitted under the Medicare Part A Benefit. DESIGN: Retrospective cohort study. SETTING: NHs located in the United States (N=13,146). PARTICIPANTS: All persons admitted to skilled nursing facilities (SNFs) in 2001 who died in a SNF (n=101,307) or hospital (n=51,187). MEASUREMENTS: Patient, facility, and geographic characteristics associated with death in a hospital and receipt of Medicare payment. RESULTS: Absence of a do-not-resuscitate order, non-Caucasian ethnicity, greater functional independence, and higher cognitive status correlated with hospital as the site of death. Rural, hospital-based, and government-owned facilities had the lowest in-hospital death rates. Site of death varied widely from state to state. Of those who died in a hospital, 24.2% (12,410) died within 24 hours of transfer. The average daily combined stay Medicare payment for those who died in the hospital was $969, versus $300 for those who died in a NH. CONCLUSION: Patient and facility characteristics predict site of death of Medicare NH patients, but in-hospital death rather than NH death varies geographically and is associated with higher daily Medicare payment. PMID- 15271111 TI - The effect of a cognitive task on voluntary step execution in healthy elderly and young individuals. AB - OBJECTIVES: To investigate voluntary step behavior of healthy elderly individuals during single- and dual-task conditions and to compare it with those of young subjects. DESIGN: Laboratory-based study. SETTING: Tests of healthy elderly and young individuals from senior community centers and from the university population in Boston, Massachusetts. PARTICIPANTS: Sixty-six elderly and 12 young subjects. MEASUREMENTS: Forward, sideways, and backward rapid voluntary stepping performed as a reaction time task while standing on a force platform and (1) awaiting a cutaneous cue (single task) and (2) awaiting a cutaneous cue while performing an attention-demanding Stroop task (dual task). Step initiation phase, foot-off time, foot contact time, and preparatory and swing phases were extracted from center-of-pressure and ground reaction force data. RESULTS: Elderly subjects were significantly slower than young in all step parameters under both conditions. For dual compared with single task, the initiation phase increased 108% in the elderly group and 34% in the young. There was a short-term learning effect during the dual task in elderly subjects but not in the young. CONCLUSION: The disproportional increase in step initiation time during the dual task in the elderly group suggests that they lacked neural processing resources required for swift multitasking during a voluntary postural task. This may be a factor contributing to balance loss and the large number of falls in elderly persons. Training may improve this skill. Clinical tests of postural function should incorporate multitask conditions to capture a more complete assessment of an individual's ability. PMID- 15271112 TI - Prevalence and outcomes of low mobility in hospitalized older patients. AB - OBJECTIVES: To estimate the prevalence of different levels of mobility in a hospitalized older cohort, to measure the degree and rate of adverse outcomes associated with different mobility levels, and to examine the physician activity orders and documented reasons for bedrest in the lowest mobility group. DESIGN: A prospective cohort study. SETTING: An 800-bed university teaching hospital. PARTICIPANTS: Four hundred ninety-eight hospitalized medical patients, aged 70 and older. MEASUREMENTS: Using average mobility level, scored from 0 to 12, the low-mobility group was defined as having a score of 4 or less, intermediate as a score of higher than 4 to 8, and high as higher than 8. Outcomes were functional decline, new institutionalization, death, and death or new institutionalization. RESULTS: Low and intermediate levels of mobility were common, accounting for 80 (16%) and 157 (32%) study patients, respectively. Overall, any activity of daily living (ADL) decline occurred in 29%, new institutionalization in 13%, death in 7%, and death or new institutionalization in 22% of patients in this cohort. When compared with the high mobility group, the low and intermediate groups were associated with the adverse outcomes in a graded fashion, even after controlling for multiple confounders. The low-mobility group had an adjusted odds ratio (OR) of 5.6 (95% confidence interval (CI)=2.9-11.0) for ADL decline, 6.0 (95% CI=2.5 14.8) for new institutionalization, 34.3 (95% CI=6.3-185.9) for death, and 7.2 (95% CI=3.6-14.4) for death or new institutionalization. The intermediate group had adjusted ORs of 2.5 (95% CI=1.5-4.1), 2.9 (95% CI=1.4-6.0), 10.1 (95% CI=1.9 52.9), and 3.3 (95% CI=1.8-5.9) for ADL decline, new institutionalization, death, and death or new institutionalization, respectively. Bedrest was ordered at some point during hospitalization in 165 (33%) patients. For most patients, mobility was limited involuntarily (bedrest orders), and almost 60% of bedrest episodes in the lowest mobility group had no documented medical indication. CONCLUSION: Low mobility and bedrest are common in hospitalized older patients and are important predictors of adverse outcomes. This study demonstrated that the adverse outcomes associated with low mobility and bedrest may be viewed as iatrogenic events leading to complications, such as functional decline. PMID- 15271113 TI - The health status of elderly veteran enrollees in the Veterans Health Administration. AB - OBJECTIVES: To examine the health status of elderly veteran enrollees, stratified by age group, and compare with nonveteran populations. DESIGN: Cross-sectional study. SETTING: Outpatient. PARTICIPANTS: A total of 1,406,049 veteran enrollees were surveyed, and 887,775 returned the questionnaire (63.1%). Of these, 663,729 (74%) were aged 65 and older. MEASUREMENTS: Patient demographics, comorbid conditions, and health status, which was assessed using the Veterans 36-item short form (SF-36), a reliable and valid measure of health-related quality of life (HRQoL). RESULTS: Elderly veteran enrollees are a group with poor health status across all scales of the Veterans SF-36. Significant decline in HRQoL was found in patients grouped by increasing age (65-74, 75-84, and > or =85). Of the Veterans SF-36 scales, the role physical and role emotional scales and physical functioning presented the largest decrements by age group. The elderly veteran enrollees had poorer health status than older people enrolled in Medicare managed care, ranging from 0.5 to 1 standard deviations worse. CONCLUSION: Elderly veteran enrollees have substantial disease burden, as reflected by major impairments across multiple dimensions of HRQoL. These findings bear important implications for use of services, suggesting that the Veterans Health Administration will require considerable resources to provide care for its aging population. PMID- 15271114 TI - Ethnic differences in the prevalence and pattern of dementia-related behaviors. AB - OBJECTIVES: To determine the prevalence of dementia-related behaviors in a large, multiethnic sample of community-dwelling patients with moderate to severe dementia and to determine whether differences in patient or caregiver characteristics could explain any differences in prevalence of these behaviors between white and nonwhite patients. DESIGN: Cross-sectional study. SETTING: Community-based. PARTICIPANTS: A total of 5,776 Medicare patients (5,090 white, 469 black, 217 Latino; mean age 78.9) enrolled in the Medicare Alzheimer's Disease Demonstration and Evaluation study at eight sites across the United States between 1989 and 1991. MEASUREMENTS: Trained interviewers collected information on patient demographic characteristics, cognitive and functional status, and caregiver characteristics such as relationship to patient, functional status, depression, and burden. Ethnicity was obtained by self-report. Caregivers were asked if the patient typically demonstrated any of eight dementia-related behaviors. To determine the independent association between ethnicity and dementia-related behaviors, logistic regression models were developed for each of the behaviors, adjusting for patient and caregiver characteristics. RESULTS: Overall, 92% of patients had one or more dementia-related behaviors. Sixty-one percent of black and 57% of Latino patients were reported to have four or more behaviors, compared with 46% of white patients (P<.001). The prevalence of specific behaviors ranged from 24% for combativeness to 67% for wandering. After multivariate adjustment, black patients were significantly more likely than whites to be constantly talkative (odds ratio (OR)=1.41, 95% confidence interval (CI)=1.11-1.80), to have hallucinations (OR=1.89, 95% CI=1.49-2.40) and episodes of unreasonable anger (OR=1.70, 95% CI=1.34-2.15), to wander (OR=1.40, 95% CI=1.08-1.81), and to wake their caregiver (OR=1.33, 95% CI=1.04-1.69). Latinos had a significantly higher likelihood than whites of having hallucinations (OR=1.49, 95% CI=1.10-2.01), episodes of unreasonable anger (OR=1.59, 95% CI=1.18 2.16), combativeness (OR=1.59, 95% CI=1.17-2.17), and wandering (OR=1.59, 95% CI=1.21-2.26). For most behaviors, these adjusted ORs are similar in magnitude of effect and statistical significance to the unadjusted estimates. CONCLUSION: Black and Latino community-dwelling patients with moderate to severe dementia have a higher prevalence of dementia-related behaviors than whites. Therefore, as the aging minority population grows, it will be especially important to target caregiver education, in-home support, and resources to minority communities. PMID- 15271115 TI - Government expenditures at the end of life for short- and long-stay nursing home residents: differences by hospice enrollment status. AB - OBJECTIVES: To examine end-of-life government expenditures for short- and long stay Medicare- and Medicaid-eligible (dual-eligible) nursing home (NH) hospice and nonhospice residents. DESIGN: A retrospective cohort study. SETTING: Six hundred fifty-seven Florida NHs. PARTICIPANTS: Dual-eligible NH residents who died in Florida NHs between July and December 1999 (N=5,774). MEASUREMENTS: Nursing home stays of 90 days or less were considered short stays (n=1,739), and those over 90 days were long stays (n=4,035). Three diagnosis groups were studied: cancer without Alzheimer's disease or dementia, Alzheimer's disease or dementia, and other diagnoses. Eligibility and expenditure claims data for 1998 and 1999 were merged with vital statistics and NH resident assessment data to determine diagnoses, location of death, hospice enrollment, eligibility, and expenditures. RESULTS: Twenty percent of short-stay (n=350) and 26% of long-stay (n=958) NH decedents elected hospice; of these, 73% of short-stay and 58% of long stay NH residents had hospice stays of 30 days or less. Overall, mean government expenditures in the last month of life were significantly less for hospice than nonhospice residents (7,365 dollars; 95% confidence interval (CI)=7,144-7586 dollars vs 8,134 dollars; 95% CI=7,896-8,372 dollars), but 1-month expenditures were only significantly lower for hospice residents with short NH stays, not for those with long NH stays. CONCLUSION: Overall, hospice care in NHs does not appear to increase government expenditures. Because significantly lower expenditures are observed for short-stay NH hospice residents, policy restricting access to Medicare hospice for Medicare skilled nursing facility residents may represent a missed opportunity for savings. PMID- 15271116 TI - Black/White differences in pressure ulcer incidence in nursing home residents. AB - OBJECTIVES: To compare black and white nursing home residents with respect to the incidence of nursing home (NH)-acquired pressure ulcers (PUs) and to examine the role of resident characteristics and facility characteristics in explaining differences between the racial groups. DESIGN: Prospective cohort study conducted between 1992 and 1995. SETTING: Fifty-nine Maryland NHs. PARTICIPANTS: A total of 1,938 residents (301 black, 1,637 white) aged 65 and older newly admitted to participating NHs. MEASUREMENTS: The outcome variable was the first occurrence of a Stage 2, 3, or 4 PU as determined based on medical record review. The predictor variable was race (black, white). Eight resident characteristics (age, sex, number of activity of daily living dependencies, bedfast, PU on admission to facility, incontinence, dementia, and whether the resident was on Medicaid) and three facility characteristics (number of beds, for-profit ownership status, and urban/nonurban location) were considered as possible confounding variables. RESULTS: The incidence of PUs was 0.38 per person-year in the NH. The rate for blacks was significantly higher than for whites (0.56 vs 0.35 per person-year) (P<.001). In multivariate analysis, controlling for eight resident characteristics and three facility characteristics, race was significantly associated with PU incidence (hazard ratio comparing blacks with whites=1.31, 95% confidence interval=1.02-1.66). CONCLUSION: Blacks have a higher incidence of NH acquired PUs than whites; resident characteristics appear to mediate the higher risk. Future research should aim to identify modifiable factors that explain differences between racial groups in PU risk and to develop solutions to prevent the suffering and cost associated with PUs. PMID- 15271117 TI - Risk factors for deep vein thrombosis in inpatients aged 65 and older: a case control multicenter study. AB - OBJECTIVES: To identify independent risk factors of symptomatic deep vein thrombosis (DVT) in geriatric inpatients and to define high-risk patients likely to benefit from preventive treatment. DESIGN: Hospital-based case-control multicenter study with prospective data collection. SETTINGS: Geriatric university hospitals with long-, intermediate-, and short-term care facilities. PARTICIPANTS: All patients aged 65 and older in 19 geriatric departments were submitted to clinical surveillance over a 16-month period. MEASUREMENTS: Twenty three potential risk factors of phlebitis were screened for. Comparison using logistic regression of 310 consecutive patients with symptomatic DVT versus 310 randomly selected controls was performed. The risk for symptomatic DVT in geriatrics was then scored from the clinical risk factors identified using multivariate analysis. This score is defined by the sum of the odds ratio (OR) of each risk factor present. RESULTS: Six factors were identified as independently related to the development of DVT: restriction of mobility (from OR=1.73, limited mobility without immobilization, to OR=5.64, bedridden during <15 days), aged 75 and older (OR=1.5/10 years), history of DVT or pulmonary embolism (OR=3.38), acute heart failure (OR=2.52), chronic edema of the lower limbs (OR=2.51), and paresis or paralysis of a lower limb (OR=2.06). The defined score of 8 or higher corresponded to an 88.7% probability of having symptomatic DVT. CONCLUSION: Treatments to prevent symptomatic DVT in hospitalized elderly should be evaluated on patients with these factors. PMID- 15271118 TI - Providing nutrition supplements to institutionalized seniors with probable Alzheimer's disease is least beneficial to those with low body weight status. AB - OBJECTIVES: To examine whether providing a midmorning nutrition supplement increases habitual energy intake in seniors with probable Alzheimer's disease (AD) and to investigate the effects of body weight status and cognitive and behavioral function on the response to the intervention. DESIGN: Randomized, crossover, nonblinded clinical trial. SETTING: A fully accredited geriatric teaching facility affiliated with the University of Toronto's Medical School with a home for the aged. PARTICIPANTS: Thirty-four institutionalized seniors with probable AD who ate independently. INTERVENTION: Nutrition supplements were provided between breakfast and lunch for 21 consecutive days and compared with 21 consecutive days of habitual intake. MEASUREMENTS: Investigator-weighed food intake, body weight, cognitive function (Severe Impairment Battery and Global Deterioration Scale), behavioral disturbances (Neuropsychiatric Inventory-Nursing Home Version), and behavioral function (London Psychogeriatric Rating Scale). RESULTS: Relative to habitual intake, group mean analyses showed increased 24 hour energy, protein, and carbohydrate intake during the supplement phase, but five of 31 subjects who finished all study phases completely compensated for the energy provided by the supplement by reducing lunch intake, and 24-hour energy intake was enhanced in only 21 of 31 subjects. Compensation at lunch was more likely in subjects with lower body mass indices, increased aberrant motor behavior, poorer attention, and increased mental disorganization/confusion. CONCLUSION: Nutrition supplements were least likely to enhance habitual energy intake in subjects who would normally be targeted for nutrition intervention those with low body weight status. Those likely to benefit include those with higher body mass indices, less aberrant motor problems, less mental disorganization, and increased attention. PMID- 15271119 TI - Plasma hypertonicity: another marker of frailty? AB - OBJECTIVES: To determine whether plasma hypertonicity might be a marker of early frailty, this study tested the associations between plasma hypertonicity, incident disability, and mortality in nondisabled older adults. DESIGN: Longitudinal, observational study. SETTING: Community-based. PARTICIPANTS: Older adults (> or =70), who reported no disability and gave blood in the 1992 Duke Established Populations for Epidemiologic Studies of the Elderly survey (n=705), were re-interviewed in 1996 for functional status (n=561) and followed for all deaths up to January 1, 2000. MEASUREMENTS: Plasma tonicity was estimated from plasma glucose, sodium, and potassium measures and used to classify subjects as normo- (285-294 mOsm/L) or hypertonic (> or =300 mOsm/L). Disability was defined as any impairment on the Rosow-Breslau, activity of daily living (ADL), and instrumental activity of daily living (IADL) scales. The relative risk (RR) of any new disability and relative hazard of death associated with hypertonicity were estimated using logistic regression models and Cox proportional hazards models, respectively. All models were controlled for age, sex, race, weight status, current smoking, activity level, plasma blood urea nitrogen and creatinine, cognitive impairment, depression, and chronic disease status. To determine whether observed effects were attributable to plasma glucose alone, all models were repeated on a subsample of nondiabetic, normoglycemic subjects. RESULTS: Plasma hypertonicity (observed in 15% of subjects) was associated with increased risk of new Rosow-Breslau (RR=2.1, 95% confidence interval (CI)=1.2 3.6), IADL (RR=2.3, 95% CI=1.2-4.3), and ADL (RR=2.7 95% CI=1.3-5.6) disability by 1996 and mortality by 2000 (RR=1.4, 95% CI=1.0-1.9). Results were similar for the normoglycemic subgroup (ADL: RR=2.9, 95% CI=1.0-8.0; IADL: RR=2.5, 95% CI=1.0 6.3; Rosow-Breslau: RR=1.8, 95% CI=0.8-3.9; mortality: RR=1.5, 95% CI=0.9-2.3). CONCLUSION: Plasma hypertonicity may be a marker of early frailty. It was prevalent in this sample of nondisabled community-dwelling older adults and predicted incident disability and mortality. Further research to identify its determinants and consequences may help inform interventions against frailty. PMID- 15271120 TI - A comparative study of clinical features and outcomes in young and older adults with severe acute respiratory syndrome. AB - OBJECTIVES: To determine the clinical presentation, findings, and outcomes of older adults (> 60) with severe acute respiratory syndrome (SARS) and compare these with a control group of younger patients (< or or =60). DESIGN: Retrospective cohort study. SETTING: A community-based, acute hospital in Hong Kong. PARTICIPANTS: All adult inpatients with a clinical diagnosis of SARS. MEASUREMENTS: Clinical presentations, investigations, treatment, and 30- and 150 day mortality. RESULTS: There were 52 young and 25 older patients with a mean age +/- standard deviation of 39.5+/-11.7 and 72.1+/-7.2, respectively. Fever, chills, and diarrhea were more common in younger patients, whereas decrease in appetite and general condition occurred only in older patients. The prevalence of positive reverse-transcriptase polymerase chain reaction for SARS-associated coronavirus (SARS-CoV) in nasopharyngeal secretions and stool samples was similar in the two groups. The prevalence of positive serological tests for SARS-CoV was significantly lower in older patients (42% vs 92%, P<.001). This was largely due to incomplete testing in elderly patients. Older patients were more likely to develop secondary nosocomial infection, be admitted to an intensive care unit, and require mechanical ventilation. The cumulative 30- and 150-day mortality rates were 3.8% and 7.6%, respectively, in young patients with SARS and 56% and 60%, respectively, in older patients (P<.001). CONCLUSION: Older patients with SARS more often presented with nonspecific symptoms, and the prognosis was poor. Reverse-transcriptase polymerase chain reaction was useful in diagnosing SARS in older patients, but the role of serological tests in individual elderly is limited. PMID- 15271121 TI - An analysis of an older driver evaluation program. AB - OBJECTIVES: To identify elements of an older driver evaluation program that predict driving performance in older adults. SETTING: Outpatient medical clinic in an academic medical center. DESIGN: A retrospective analysis. PARTICIPANTS: Six hundred sixty-four older adults who were referred to an older driver evaluation program. MEASUREMENTS: A physician trained in geriatric medicine and a clinical geriatric nurse specialist oversaw an experienced driving evaluator and an occupational therapist who conducted assessments of older persons' functional status; reaction time; driving skills; and cognitive, hearing, and vision abilities. Self-report data along with a medical history submitted by patients' primary care physicians supplemented the clinical assessments. RESULTS: A multinomial logistic regression revealed that the Mini-Mental State Examination (MMSE), cues needed with the Trail Making Test, Part B, grip strength, and an interaction effect between the MMSE and reaction time constituted the most parsimonious model for predicting on-the-road performance. A receiver operating characteristic analysis indicated that this index had good sensitivity but low specificity. A binomial regression comparing imperfect and perfect drivers demonstrated the significance of the Traffic Sign and Visual Perception tests. CONCLUSION: Clinicians should employ a multilevel screening process that includes initial cognitive tests, such as the MMSE and the Trail Making Test, Part B, although more studies of driving evaluation programs in medical settings that include random samples of older drivers are needed. PMID- 15271122 TI - Hospice care in nursing homes: is site of care associated with visit volume? AB - OBJECTIVES: To determine factors associated with hospice visit volume and to examine whether visit volume differs by nursing home (NH) versus non-NH setting. DESIGN: Retrospective cohort study. SETTING: Twenty-one hospices across seven states under the ownership of one parent provider. PARTICIPANTS: Hospice patients from October 1998 through September 1999 in NH (n=9,460) and non-NH (n=15,484) settings. MEASUREMENTS: Data were from the provider's centralized information system. Average daily visit volume was the number of visits divided by the number of hospice routine home care days (days not in hospice inpatient or continuous home care). Multivariate logistic regression tested the association between site of care and an individual's probability of having average daily visits above the sample median. RESULTS: Average daily visits+/-standard deviation were 1.1+/-1.1 for NH and 1.2+/-1.3 for non-NH hospice patients. Site of care was not significantly associated with having an average daily visit volume above the sample median, but patients in NH settings had a lower probability of having a nurse average daily visit volume above the median (adjusted odds ratio (AOR)=0.59, 95% confidence interval (CI)=0.46-0.74) and a greater probability of having social worker (AOR=2.46, 95% CI=1.87-3.24), aide (AOR=1.97; 95% CI=1.11 3.48), and clergy (AOR=3.23, 95% CI=2.21-4.44) average daily visits above the median than those in non-NH settings. CONCLUSION: A different mix, not volume, of services appears to be used to address the physical, psychosocial, and spiritual needs of hospice patients/families who reside in NH settings than of those in non NH settings. PMID- 15271123 TI - Outcomes and characteristics of patients discharged alive from hospice. AB - OBJECTIVES: To describe outcomes and characteristics of patients discharged alive from hospice. DESIGN: Prospective cohort study using a telephone survey. SETTING: Hospices (n=18) participating in the Population-Based Palliative Care Research Network during the 1-year study period. PARTICIPANTS: English-speaking adults (n=164) who were discharged alive from participating hospices during the 1-year study period. MEASUREMENTS: Mortality within 6 months of hospice discharge. RESULTS: Thirty-five percent (n=48) of the 139 patients with known outcomes died within 6 months of hospice discharge, 15 of whom (31%) died without hospice readmission. There were no significant associations between sex (P=.77), length of hospice service (P=.99), diagnosis (P=.73), discharge disposition (P=.54), admission evidence of prognosis of less than 6 months to live (P=.22-.95), Karnofsky score at admission or change between admission and discharge (P=.39, P=.38, respectively), or duration of hospice care after stabilization (P=.83) and mortality within 6 months after hospice discharge. Age (P=.11), discharge Karnofsky score (P=.17), and reason for discharge being improved or stabilized condition (P=.13) trended toward statistical significance. The strongest predictor of mortality after hospice discharge was a report that the patient's condition had worsened (hazard ratio=10.2, 95% confidence interval 4.5-23.4). CONCLUSION: One-third of patients who were discharged from hospice died within 6 months of hospice discharge, indicating ongoing eligibility for hospice care even under the strictest interpretation of hospice eligibility criteria. Patients who are discharged from hospice care should be evaluated frequently, especially within the first weeks to months after discharge, for changes in status, unmet needs, and potential hospice readmission. PMID- 15271124 TI - Psychometric comparisons of the timed up and go, one-leg stand, functional reach, and Tinetti balance measures in community-dwelling older people. AB - OBJECTIVES: To compare the practicality, reliability, validity, and responsiveness of the timed up and go (TUG), one-leg stand (OLS), functional reach (FR), and Tinetti balance (TB) performance measures in people aged 65 and older. DESIGN: A prospective study. SETTING: Shin-Sher Township of Taichung County, west-central Taiwan. PARTICIPANTS: Twelve hundred community-dwelling older people. MEASUREMENTS: During an initial assessment at their residences, participants were interviewed for demographics, cognition, fall history, use of a walking aid, and activities of daily living (ADLs), in addition to completing the four balance tests. Falls were ascertained by telephone every 3 months for a 1 year follow-up; the four balance measures and ADLs were also reassessed at the end of the follow-up year. RESULTS: Of the four balance measures, the OLS had the lowest participation rate, and participation of people who were cognitively impaired had fallen in the previous year, used a walking aid, or suffered from an ADL disability was lower than for their counterparts. The time to complete the tests ranged from 58 seconds for OLS, to 160 seconds for the TB. All four balance measures exhibited excellent test-retest reliability and discriminant validity but poor responsiveness to fall status. The TB showed better discriminant, convergent, and predictive validities and responsiveness to ADL changes than the other three tests. CONCLUSION: According to psychometric properties, the most suitable performance measure for evaluating balance in community-dwelling older people was the TB, followed by the TUG. PMID- 15271125 TI - Risk factors for adverse drug events among older adults in the ambulatory setting. AB - OBJECTIVES: To gather information on patient-level factors associated with risk of adverse drug events (ADEs) that may allow focus of prevention efforts on patients at high risk. DESIGN: Nested case-control study. SETTING: Large multispecialty group practice in New England. PARTICIPANTS: All Medicare enrollees cared for by a multispecialty group practice during 1 year (N=30,397 person-years from July 1, 1999, through June 30, 2000). For each patient with an ADE, a control was randomly selected. MEASUREMENTS: Data were abstracted from medical records on age, sex, comorbidities, and medication use at the time of the event. RESULTS: ADEs were identified in 1,299 older adults. Independent risk factors included being female and aged 80 and older. There were dose-response associations with the Charlson Comorbidity Index and number of scheduled medications. Patients taking anticoagulants, antidepressants, antibiotics, cardiovascular drugs, diuretics, hormones, and corticosteroids were at increased risk. In the analysis of preventable ADEs, the dose-response relationship with comorbidity and number of medications remained. Patients taking nonopioid analgesics (predominantly nonsteroidal antiinflammatory drugs and acetaminophen), anticoagulants, diuretics, and anti-seizure medications were at increased risk. CONCLUSION: Prevention efforts to reduce ADEs should be targeted toward older adults with multiple medical conditions or taking multiple medications, nonopioid analgesics, anticoagulants, diuretics, and antiseizure medications. PMID- 15271126 TI - Age-related changes in treatment strategies for acute myocardial infarction: a population-based study. AB - OBJECTIVES: To compare across four age groups (<65, 65-74, 75-84, > or =85) the determinants of coronary reperfusion therapy (CRT) use in ST-segment elevation acute myocardial infarction (STE-AMI). DESIGN: Population-based, observational study. SETTING: Performed in the health district of Florence, Italy, where percutaneous coronary intervention (PCI) is the preferred CRT. PARTICIPANTS: Nine hundred thirty patients with STE-AMI prospectively enrolled in the Florence AMI registry. MEASUREMENTS: Use of CRT, clinical factors associated with CRT use. RESULTS: CRT use was reduced from 71% at younger than 65 to 31% at aged 85 and older (P<.001). After adjusting for chronic comorbidity, Killip class, admission hospital category, hospitalization delay, and AMI location, CRT use was 29% (P=.17) lower at age 75 to 84 and 63% (P<.001) lower at age 85 and older than at younger than 65. Within each age group, the probability of receiving CRT was three to five times greater in patients directly admitted to the hospital with PCI facilities. Acute cardiac failure and chronic comorbidity were associated with lower CRT use only in patients aged 65 and older. Patients aged less than 85 years who received reperfusive therapy had a significantly lower risk of death ( 44%, P=.045) at 1 year, whereas it was less evident and nonsignificant (-27%, P=.27) in patients aged 85 and older. CONCLUSION: Results confirm that, although they might substantially benefit from CRT during STE-AMI, older patients are excluded from CRT even when eligible. This further indicates that clinicians are not yet completely prepared to manage most efficiently frail elderly with AMI, a task requiring a specific interdisciplinary training program in geriatric cardiology. PMID- 15271127 TI - The tumor necrosis factor alpha -308G>A polymorphism is associated with dementia in the oldest old. AB - OBJECTIVES: To test the hypothesis that the tumor necrosis factor (TNF) -308 G>A promoter gene polymorphism is a risk factor in age-related dementia and longevity. DESIGN: A cross-sectional and a longitudinal study. SETTING: A population-based sample of Danish centenarians. PARTICIPANTS: One hundred-year old Danes (n=122) from "The Longitudinal Study of Danish Centenarians." Octogenarians (n=174) and healthy volunteers aged 18 to 30 (n=47) served as reference groups. METHODS: Whether the distribution of TNF -308 GG/GA/AA genotypes were different in centenarians than in younger age groups was investigated (Fischer exact test). Furthermore, whether the TNF -308 G>A polymorphism was associated with the prevalence of dementia (logistic regression analysis), the plasma level of TNF-alpha (analysis of variance), and mortality in the following 5 years (Cox regression analysis) within the cohort of centenarians was tested. RESULTS: The distribution of TNF -308 genotypes was not different across the three different age groups, but the GA genotype was associated with decreased prevalence of dementia in centenarians. The few centenarians with AA carrier status had higher mortality risk and tended to show higher plasma levels of TNF-alpha, but the significance was questionable due to a low number of subjects with this genotype. CONCLUSION: It is possible that the TNF -308 A allele is maintained during aging because subjects who are heterozygous for this polymorphism possess the optimal inflammatory response with regard to protection against age-related neurodegeneration. PMID- 15271128 TI - Normal values of balance tests in women aged 20-80. AB - OBJECTIVES: To determine normal values for four commonly used clinical functional balance tests from community-dwelling women aged 20 to 80 and to identify any significant decline due to aging. DESIGN: A cross-sectional study was undertaken to provide normative values for four clinical balance tests across 6 decade cohorts. SETTING: The Betty Byrne-Henderson Center for Women and Aging, Royal Womens' Hospital, Brisbane, Australia. PARTICIPANTS: Four hundred fifty-six community-dwelling, independently ambulant women with no obvious neurological or musculoskeletal-related disability, aged 20 to 80, were randomly recruited from a large metropolitan region. MEASUREMENTS: The clinical balance measures/tests were the Timed Up and Go test, step test, Functional Reach test, and lateral reach test. Multivariate analysis was used to test the effect for age, height, and activity level. RESULTS: Normal data were produced for each test across each decade cohort. Gradual decline in balance performance was confirmed, with significant effect for age demonstrated. CONCLUSION: New normative data across the adult age decades are available for these clinical tests. Use of clinical balance tests could complement other balance tests and be used to screen women aged 40 to 60 whose performance is outside the normal values for age and to decrease later falls risk. PMID- 15271129 TI - The identification of genetically related bacterial isolates using pulsed field gel electrophoresis on nursing home units: a clinical experience. AB - OBJECTIVES: To describe a laboratory-based technique to track nursing home infections. DESIGN: Retrospective data analysis. SETTING: A 721-bed skilled care facility with 14 nursing units. PARTICIPANTS: Residents in a nursing home, average age 76+/-10, 78% male. MEASUREMENTS: Bacterial isolates were listed for each nursing unit. Clusters of identical species and antibiotic susceptibility were identified followed by pulsed-field gel electrophoresis (PFGE). If the genetic analysis yielded related strains, the director of nursing performed a clinical investigation. PFGE is available through reference laboratories at a cost of approximately 75 dollars/isolate. RESULTS: Twenty-four clinical clusters of phenotypically identical bacteria (species, antibiotic susceptibility) were identified. Fourteen included genetically related isolates. CONCLUSION: Approximately half of the phenotypically identical clusters contained genetically related isolates. The identification of genetically related bacterial isolates on nursing units by PFGE provides staff with a specific circumstance to review secretion precautions. Genetic analysis may also demonstrate that apparent clusters are unrelated. PMID- 15271130 TI - Accuracy of nursing home medical record information about care-process delivery: implications for staff management and improvement. AB - Arguments have been made that the culture of nursing homes (NHs) must change to improve the quality of care, and two initiatives have been designed to accomplish this goal. One initiative is to provide resident outcome information (quality indicators) to NH management and consumers via public reporting systems. This initiative is based on the assumptions that resident outcomes are related to care processes implemented by NH staff, the NH industry will respond to market forces, and there are management systems in place within NHs to change the behavior of direct care staff if outcomes are poor. A separate staffing initiative argues that NH care will not improve until there are resources available to increase the number of direct care staff and improve staff training. This initiative also assumes that systems are in place to manage staff resources. Unfortunately, these initiatives may have limited efficacy because information useful for managing the behavior of direct care providers is unavailable within NHs. Medical record documentation about daily care-process implementation may be so erroneous that even the best-intentioned efforts to improve the care received by residents will not be successful. A culture of inaccurate documentation is largely created by a discrepancy between care expectations placed on NHs by regulatory guidelines and inadequate reimbursement to fulfill these expectations. Nursing home staff have little incentive to implement the technologies necessary to audit and assure data quality if accurate documentation reveals that care consistent with regulatory guidelines is not or cannot be provided. A survey process that largely focuses on chart documentation to assess quality provides further incentive for care-process documentation as opposed to care-process delivery. This article reviews methods to improve the accuracy of NH medical record documentation and to create data systems useful for staff training and management. PMID- 15271131 TI - Building academic geriatric capacity: an evaluation of the John A. Hartford Foundation Centers of Excellence initiative. AB - Almost 15 years ago, the John A. Hartford Foundation began its Centers of Excellence (CoE) program. In summer 2002, a quantitative and qualitative evaluation of the CoE program was conducted. The evaluation used previously collected quantitative data from surveys of program directors and graduates of fellowship programs, as well as interviews and surveys of currently funded CoEs. Since its inception, the CoE program has supported 163 geriatrics fellows, of whom 63% entered academic geriatrics. Almost half of these graduating fellows have gone to new academic institutions. CoEs have also supported 222 faculty, including some who were in disciplines other than geriatrics. The vast majority (82%) have remained in academics, and nearly two-thirds are currently in geriatrics. As the priorities and needs of the institutions and geriatrics programs changed, most centers shifted their CoE priorities. These changes predominantly took two forms: a refocus from one activity to another or an expansion of outreach or levels of support. Based upon this formal evaluation, the Hartford-supported CoE program has been successful in strengthening academic geriatrics, particularly in attracting, developing, and retaining geriatrics faculty. PMID- 15271133 TI - Confronting the geriatrician's nightmare. PMID- 15271134 TI - Accuracy (or lack thereof) of nursing home medical records: what do industry and professional organizations have to say? PMID- 15271135 TI - A cure for all ills. PMID- 15271136 TI - Thursdays with Caroline. PMID- 15271137 TI - Where do continuing care retirement community residents die? PMID- 15271138 TI - Skull fracture? Not! PMID- 15271139 TI - Increased arterial carboxyhemoglobin concentrations in elderly patients with silicosis. PMID- 15271140 TI - An unusual cause of diarrhea in an elderly woman: a case report. PMID- 15271141 TI - Blisters in a nursing home: bullous pemphigoid more often than we think? PMID- 15271142 TI - Acquired hemophilia in the elderly: a case report. PMID- 15271143 TI - Dysphagia and unexpected myasthenia gravis associated with primary biliary cirrhosis, ulcerative colitis and vitiligo. PMID- 15271144 TI - Diabetic amyotrophy and autoimmune thyroiditis: are they related? PMID- 15271145 TI - Cobalamin reduces homocysteine in older adults on folic acid-fortified diet: a pilot, double-blind, randomized, placebo-controlled trial. PMID- 15271146 TI - Benefits of pneumococcal vaccination for bedridden patients. PMID- 15271147 TI - Polymyalgia rheumatica complicating influenza vaccination. PMID- 15271148 TI - Mortality in donepezil-treated patients. PMID- 15271150 TI - The goal of culturally sensitive gerontological care. PMID- 15271152 TI - Analysis of the moral habitability of the nursing work environment. AB - BACKGROUND: Following health reform, nurses have experienced the tremendous stress of heavy workloads, long hours and difficult professional responsibilities. In recognition of these problems, a study was conducted that examined the impact of the working environment on the health of nurses. After conducting focus groups across Canada with nurses and others well acquainted with nursing issues, it became clear that the difficult work environments described had significant ethical implications. AIM: The aim of this paper is to report the findings of research that examined the moral habitability of the nursing working environment. METHODS: A secondary analysis was conducted using the theoretical work of Margaret Urban Walker. Moral practices and responsibilities from Walker's perspective cannot be extricated from other social roles, practices and divisions of labour. Moral-social orders, such as work environments in this research, must be made transparent to examine their moral habitability. Morally habitable environments are those in which differently situated people experience their responsibilities as intelligible and coherent. They also foster recognition, cooperation and shared benefits. FINDINGS: Four overarching categories were developed through the analysis of the data: (1) oppressive work environments; (2) incoherent moral understandings; (3) moral suffering and (4) moral influence and resistance. The findings clearly indicate that participants perceived the work environment to be morally uninhabitable. The social and spatial positioning of nurses left them vulnerable to being overburdened by and unsure of their responsibilities. Nevertheless, nurses found meaningful ways to resist and to influence the moral environment. CONCLUSIONS: We recommend that nurses develop strong moral identities, make visible the inseparability of their proximity to patients and moral accountability, and further identify what forms of collective action are most effective in improving the moral habitability of their work environments. PMID- 15271153 TI - Commentary: the discourse of moral suffering. PMID- 15271154 TI - Commentary: methodological approach and implications for practice. PMID- 15271155 TI - Implementing local pay systems in nursing and midwifery. AB - BACKGROUND: The paper is based on a case study, which was part of a Department of Health commissioned research study covering 10 National Health Service (NHS) trusts in England that had adopted a range of approaches to the employment terms and conditions of nurses, midwives, and other non-medical staff, as the precursor to evaluating Agenda for Change, the modernized pay system for the NHS. AIM: The aim of this paper is to discuss a case study of the effects of changing nurses' pay progression. METHODS: Fieldwork took place in 2000, and included interviews with managers, union representatives and other staff, and analysis of internal documents. FINDINGS: Findings discussed include the constraints on managers when devising a new pay system, the time and detailed work needed and the challenges of assessing their effects (particularly in relation to patient care). Although the latter are difficult to assess, staff involved in the scheme reported benefits from the closer focus on competences which resulted from the scheme. CONCLUSIONS: The effects of changes to pay schemes are difficult to assess, and the evaluation of the proposed national scheme (Agenda for Change) will be challenging. PMID- 15271156 TI - School nurses: policies, working practices, roles and value perceptions. AB - BACKGROUND: In the UK, school nursing has recently been at the forefront of policy change, with school nurses being considered pivotal to child-centred public health practice. There is very little literature on this topic and, in particular, little that is written from a practitioner perspective. AIM: This paper reports a survey that examines the work of school nurses compared with the expectations of their first line managers and policy-makers in government, in order to discover any potential practical or ideological areas of conflict. METHODS: We first applied a theoretical framework of sensitizing concepts to the historical, political, cultural and contextual background of school nursing. Following this, 46 school nurses in the West Midlands region of the UK were randomly selected and asked to complete a questionnaire about their personal characteristics, experience, training and working practices. The 38 nurses who completed this were then interviewed. Job descriptions for school nurses and governmental job expectations were obtained from various official sources and compared with the self-reported practices of school nurses. FINDINGS: All the nurses met the work criteria of their local employers, except in respect of health needs assessment activities, which two nurses had not yet attempted. All 38 also carried out a range of additional work activities, including providing sexual health services and parental support clinics. They also had a diverse range of skills and qualifications relevant to supporting the needs of their local communities. Qualitative data from interviews provided a useful insight into nurses' feelings of being valued by their clients and by local and national employers, and feelings of professional undervaluing by their peers. They felt positive about role changes in the last few years, and that they supported the child-centred public health role advocated by public policy. CONCLUSIONS: The practice of the school nurses in this study covered what employers and policy makers required, with the notable exception of health needs assessment, which nurses were uncomfortable and unconfident about. The theoretical framework used provides a useful starting point for examining how school nursing has developed into its current role. PMID- 15271157 TI - Nurses' attitudes towards lesbians and gay men. AB - BACKGROUND: During the last decade, official policy and Swedish legislation have strengthened the legal rights of homosexuals and demanded tolerance for this group. There is evidence in the literature that homosexual patients have experienced negative attitudes and poor quality care from nurses, and may be unwilling to disclose their sexuality because of fears of discriminatory treatment. AIMS: The aim of this paper is to report a study that investigated the attitudes of nurses towards lesbians and gay men and nurses beliefs about the causes of homosexuality. METHOD: The study had a descriptive, comparative design. The Attitudes Toward Homosexuality Scale was used, along with Causes of Homosexuality Questionnaire. The participants were Registered Nurses and Assistant Nurses from one infectious disease clinic in central Sweden (response rate 67%, n = 57), and students enrolled in a university nursing programme and in upper secondary assistant nurses' training (response rate 62%, n = 165). RESULTS: In general, participants expressed positive attitudes (62%). Nurses expressed the most positive attitudes, whereas the assistant nursing students expressed the least positive attitudes. A minority of the sample (30%) expressed neither positive nor negative attitudes. The most common belief about the cause of homosexuality was that it was congenital. Those who held this belief expressed more positive attitudes towards homosexuality than those who believed that homosexuality was acquired. Limitations of the study were that the sample was relatively small and not randomly selected. CONCLUSION: This study demonstrated that attitudes have improved towards homosexuals compared with earlier international studies, although more needs to be done to increase the positive attitudes among the nursing staff and students with neutral attitudes (neither positive nor negative attitudes) to enhance the wellbeing of homosexual persons. General education about homosexuality is a necessary beginning to make homosexual patients visible, which is an important aspect of practical nursing ethics. PMID- 15271158 TI - Preventing hypothermia during continuous veno-venous haemodiafiltration: a randomized controlled trial. AB - BACKGROUND: Continuous veno-venous haemodiafiltration is a common form of dialysis used in intensive care units. Unfortunately, patients often experience hypothermia as a side-effect of the therapy because of the necessity for extracorporeal blood flow. Intensive care nurses aim to prevent hypothermia developing. Intravenous fluid warmers are sometimes added to the dialysis circuit in an attempt to maintain patient temperature. However, the efficacy of this method has not been previously studied. AIM: This paper reports a study to investigate whether intravenous fluid warmers prevent hypothermia during continuous veno-venous haemodiafiltration. METHOD: A prospective randomized controlled trial was carried out in the intensive care unit of a metropolitan, tertiary-referral, teaching hospital. After Ethics Committee approval, 60 circuits in continuous veno-venous haemodiafiltration mode (200 mL/minute blood flow, 1 L/hour countercurrent dialysate, 3 L/hour pump-controlled ultrafiltration and prefilter fluid replacement of 1.7-2.0 L/hour) were studied. Circuits were randomized to have either an intravenous fluid warmer set at 38.5 degrees C on the dialysate and 1 L/hour of replacement fluid lines or no fluid warmer. Patient core temperature was recorded at baseline and then hourly. Hypothermia was defined as a core temperature <36.0 degrees C. RESULTS: Mean core temperature loss did not vary between circuits with or without a fluid warmer (0.92 degrees C vs. 1.11 degrees C, P = 0.339). Survival analysis found no difference in hypothermia incidence between groups (log rank = 0.47, d.f. = 1, P = 0.491). Lower baseline temperature (RR 0.142, 95% CI 0.044, 0.459, P = 0.001) and female gender (RR 0.185, 95% CI 0.060, 0.573, P = 0.003) were significant risks for hypothermia. CONCLUSIONS: Intravenous fluid warmers used as described do not prevent hypothermia during continuous veno-venous haemodiafiltration. Female patients and those with a low-normal baseline temperature are most likely to become hypothermic during this form of dialysis. Further research is needed to address effective ways of preventing hypothermia in critically ill patients receiving continuous renal replacement therapies. PMID- 15271159 TI - Health issues of aboriginal female adolescents in Taiwan. AB - BACKGROUND: The mortality rate and prevalence for some chronic diseases are higher for aborigines compared with the rest of the Taiwanese population. Health professionals in many countries have been concerned with the health of aborigines, but specific health policies for native adolescents have been neglected. AIMS: This paper reports a study comparing the general health status of aborigines and non-aborigine female adolescents, based on school enrollment health examinations. METHODS: A cross-sectional prevalence study with between group comparison was conducted. School enrollment health files were accessed between September 2000 and June 2001. A total of 320 selected female aboriginal students were compared with 237 non-aborigines. RESULTS: With the exception of eye problems, general health status was inferior for the aboriginal female adolescent students. They had lower mean red blood cell volumes, lower haemoglobin concentrations, more oral decay, a greater prevalence of hepatitis B virus carriers and were more likely to be overweight. CONCLUSIONS: It seems reasonable to suggest that health intervention strategies specific for indigenous adolescents should be provided. The design of these strategies should target lifestyle modification and provision of more accessible health information. Health professionals, and particularly nurses, are in a prime position to offer advice and support to this group. PMID- 15271160 TI - Interpretations of stillbirth. AB - BACKGROUND: Continuity is a major concept in the father-son domain of the Han Chinese value system in Taiwan. Aspects of continuity may include structure, interactions and other facets of family; however, providing descendants is the keystone of women's reality in these families. In a culture in which death is seen as a taboo subject and the unborn child has not been recognized as a real baby, losing a long-expected child at the end of pregnancy becomes a great challenge to women who have experienced stillbirth. AIM: The aim of this paper is to report a study exploring Taiwanese mothers' interpretations of stillbirth, and their unique sociocultural context. METHOD: An interpretive ethnographic approach was used. Over a two and a half-year period, 20 women who had experienced such losses after at least 20 weeks of pregnancy were interviewed to find out how they interpreted their babies' deaths. Interview data were analysed thematically. FINDINGS: The four major themes identified were: 'loss of control', 'broken dream', 'shattered self' and 'something wrong with me'. Interpretations of stillbirth among Taiwanese women indicate a strong sense of incompleteness and personal failure, triggering reactions in terms of not only maternal identity, but also female cultural roles. Many interviewees blamed themselves for the deaths of their unborn children, a viewpoint resulting in excessive guilt feelings. CONCLUSION: Culturally bound taboos against talking about death, participating in death-related events, and expressing grief in public affect the adaptation and grieving processes of Taiwanese women who have had a stillbirth. Nurses should, therefore, make an effort to listen to the perspectives of such patients in order to assist them with coming to terms with their loss. As part of their education, nurses require information on cultural beliefs so that they can provide appropriate care to grieving mothers. PMID- 15271161 TI - Anxiety and self-consciousness in patients with minor facial lacerations. AB - BACKGROUND: Although minor facial injuries are relatively common, their psychological impact is an area neglected in the literature. For physiologically major injuries (such as facial cancers, burns and fractures), the face has been suggested to be a psychologically significant area of the body and disfigurement has been found to have numerous potential social consequences for patients. AIMS: This paper reports the findings of an inquiry that explored the psychological impact of minor facial injuries and the influence of patient and scar characteristics in relation to self-consciousness and anxiety levels. METHOD: Data were collected in 2001 in an accident and emergency unit from patients with a visible laceration over 1.5 cm that was treatable in an outpatient setting. The Derriford Appearance Scale (with general and social self-consciousness subscales) and the State-Trait Anxiety Inventory were administered to 63 patients 1 week later; data on 50 patients were also available 6 months after the injury. RESULTS: Larger scar size, living alone and aetiology of injury were significantly related to self-consciousness and anxiety levels, although gender, age, socio-economic group, location of scar, satisfaction with appearance and number of scars were not. General self-consciousness improved at 6 months but social self-consciousness and anxiety remained the same. Patient factors were not related to changes in general self-consciousness over time. CONCLUSIONS: Minor facial scars can have significant psychological impact for some people. Awareness training for health professionals, social skills training for affected patients and a patient information leaflet are recommended. PMID- 15271162 TI - Priority setting in clinical nursing practice: literature review. AB - BACKGROUND: Time is a valuable resource. When nurses experience demands on their services which exceed their available time, then 'rationing' must occur. In clinical practice such rationing requires practitioners to set priorities for care. AIMS: The aim of this paper is establish what is currently known about priority setting in nursing, including how nurses set priorities and what factors influence this. METHOD: CINAHL, Medline, ASSIA, and PsychLit databases for the years 1982-2002 were searched, using the terms (clinical decision-making or problem-solving or planning) and (setting priorities or prioriti*). The publications found were used in a selective, descriptive review. FINDINGS: Priority setting is an important skill in nursing, and a skill deficit can have serious consequences for patients. Recent studies have suggested that it is a difficult skill for newly qualified nurses to acquire and may not be given sufficient attention in nurse education. Priority setting can be defined as the ordering of nursing problems using notions of urgency and/or importance, in order to establish a preferential order for nursing actions. A number of factors that may impact on priority setting have been identified in the literature. These include: the expertise of the nurse; the patient's condition; the availability of resources; ward organization; philosophies and models of care; the nurse-patient relationship; and the cognitive strategy used by the nurse to set priorities. However, very little empirical work has been conducted in this area. CONCLUSIONS: Further study of priority setting in a range of clinical practice settings is necessary. This could inform both practice and education, promote better use of limited resources and maximize patient outcomes. PMID- 15271163 TI - Emancipation in decision-making in women's health care. AB - BACKGROUND: Emancipation as a nursing concept is derived from a long-standing history of social oppression and is easily addressed by both critical social theory and feminist theory. It is the apparent concept to describe a phenomenon witnessed in nursing when caring for women in the decision-making process about health care issues. Emancipation has been recognized by expert clinical observation. AIM: The aim of this paper is to define the concept of emancipation for possible future application to nursing practice for the promotion of humanistic nursing care in women's health, specifically applied to the decision making process. METHOD: A literature search was carried out using the CINAHL database and the keywords nursing and emancipation, and covering the period 1985 2003. The Rodgers and Knafl (2000) method of concept analysis was then used to derive a conceptual meaning of emancipation that benefits patient care as well as professional nursing development. Emancipation is broken down into antecedents, attributes and consequences. Related concepts are also explored, compared and discussed to synthesize relevant characteristics. FINDINGS: This concept analysis identifies emancipation in decision-making as a nursing phenomenon by discussing the antecedent of oppression and exploring the identified attributes: (a) empowerment, (b) personal knowledge, (c) social norms, (d) reflection and (e) flexible environment. The consequence of emancipation is free choice. It is a futuristic concept with strong historical ties in need of exploration and development within the context of women's health care in relation to decision making. CONCLUSIONS: The concept model illustrates emancipated decision-making, with its five attributes in relation to oppression as a non-linear phenomenon. Areas for further study include the exploration of the contribution of each critical attribute and its relationship to emancipated decision-making, and the decision-making process in relation to patient satisfaction and how long the person continues to adhere to the decision. Also the professional nursing role in promotion of emancipated decision-making is virtually unexplored, but is an important concept in the paradigm of shared decision-making about health care alternatives. PMID- 15271164 TI - Attitudes towards hospitalized older adults. AB - BACKGROUND: A number of studies have explored nurses' attitudes toward older people. However many of those have produced inconclusive results. Moreover dated attitudinal measures have been used to answer a broad range of questions relating to older people. AIM: The aim of this paper is to report a study examining whether negative attitudes and beliefs toward older adults persist. METHODS: Focus group interviews were used identifying factors which may influence attitudes and beliefs both negatively and positively. Nine Registered Nurses from care of older adult areas, four from acute areas, six nurse teachers, and 17 nursing students participated in the study. Ten themes were identified from the findings. FINDINGS: The findings show that the student nurses had varying experiences in older adult settings. Some of which had the effect of turning them away from the specialty. However, nurses who worked with older adults were very positive about their work and the nursing opportunities they had to offer student nurses. They were rather critical of the content of the pre-registration curriculum, which they perceived to over-rely teaching the negative aspects of ageing, and there was also criticism of the currency of teachers' knowledge. Nurses who worked in acute settings also did not escape criticism, in that they were identified as attributing a lack of sense of humour to older adults. The limitations include the small sample size although it is congruent with qualitative research. CONCLUSIONS: All nursing staff need to be more aware of their influence on the attitudes of student nurses toward older people. Good practice includes the ability to demonstrate that older people in hospital settings are valued. Nurse teachers need to review the way they prepare students for this specialist work in order to avoid inadvertently conveying negative attitudes. PMID- 15271170 TI - The long and short of the risk of antibiotic treatment failure. PMID- 15271171 TI - Short course intravenous benzylpenicillin treatment of adults with meningococcal disease. AB - BACKGROUND: Short-course treatment of meningococcal disease (including meningitis) with 4-5 days of an i.v. beta-lactam is of proven efficacy. Since April 1998, all adult patients with meningococcal disease admitted to Auckland Hospital were prospectively treated with 3 days of i.v. benzylpenicillin. AIMS: To assess the clinical features, laboratory findings, disease complications and outcome of patients with meningococcal disease prospectively treated with 3 days of i.v. benzylpenicillin. METHODS: A retrospective chart review of all adult patients with meningococcal disease admitted to Auckland Hospital from April 1998 to December 2002 was conducted. RESULTS: Ninety patients with definite (n = 72) or -probable (n = 16) meningococcal disease were admitted during the study period. Two were excluded on the basis of treatment duration. The remaining 88 patients received a mean +/- standard deviation duration of treatment of 3.1 +/- 0.5 days (excluding those who died while receiving treatment). Six patients (7%) died, four of whom while on treatment. There were no relapses. CONCLUSION: Three days of i.v. benzylpenicillin for the treatment of adults with meningococcal disease is effective. PMID- 15271172 TI - Positron emission tomography scanning in the assessment of patients with lymphoma. AB - BACKGROUND: The detection of lymphoma by computed tomography (CT) scanning is known to be improved by positron emission tomography (PET) and/or gallium scanning, although the direct comparative accuracy of these imaging modalities remains a subject of ongoing review. AIMS: The aim of the present study was to compare PET scanning with conventional imaging (CT and/or gallium scanning) in patients with lymphoma. METHODS: A retrospective study of 38 patients (25 men; 13 women; median age 39.5 years; range 18.0-81.0 years) who had had PET scans (24 scans at initial staging and 46 scans at restaging, including suspected disease relapse) was carried out. Thirty-one concurrent gallium scans had been performed. Disease was validated with clinical follow up or biopsy. RESULTS: The sensitivities of PET and CT at initial staging were 96 and 71%, respectively. PET identified additional sites of disease compared with CT in 29% of patients. Of the 15 patients who had had all three imaging modalities, the sensitivities of PET, CT and gallium were 93, 67 and 87%, respectively. At treatment completion, the positive predictive values of PET, CT and gallium scans for relapse given a residual mass were 100, 33 and 0%, respectively (P = 0.006 for PET and CT comparison). The negative predictive values of PET, CT and gallium were 76, 0 and 70%, respectively (P-value not significant). In suspected disease relapse, PET results changed management in 50% of patients. CONCLUSION: Compared with CT and gallium scans, PET has superior accuracy in staging and restaging, and its greatest value lies in its positive predictive value for relapse in patients with residual masses. PMID- 15271173 TI - Analysis of clinical outcomes following in-hospital adult cardiac arrest. AB - AIMS: The outcome of in-hospital resuscitation following cardiac arrest depends on many factors related to the patient, the environment and the extent of resuscitation efforts. The aim of the present study was to determine predictors of successful resuscitation and survival to -hospital discharge following in hospital cardiac arrest and to assess functional outcomes of survivors (cerebral performance scores). METHODS: Medical records of adult patients sustaining in hospital cardiac arrest between June 2001 and January 2003 were reviewed. Successful resuscitation was defined as the return of spontaneous circulation at the completion of resuscitative efforts, irrespective of degree of inotropic/vasopressor support. Thirty demographic and clinical variables were analysed to determine predictors of successful resuscitation and in-hospital survival. RESULTS: In 105 patients with cardiac arrest, 46 patients (44%) were successfully resuscitated and 22 (21%) survived to hospital discharge. Predictors of successful resuscitation included a primary cardiac admission diagnosis, monitoring at the time of the arrest, a longer duration of resuscitation and the absence of the need for endotracheal intubation. Patients with ventricular tachycardia/fibrillation were more likely to survive to hospital discharge than those with asystolic or pulseless electrical activity (45 vs 12 vs 20%, P = 0.01). The sole independent predictor of survival to hospital discharge was the absence of the need for endotracheal intubation (odds ratio 0.14, 95% confidence interval 0.02-0.88, P < 0.01). The majority of survivors (73%) had normal cerebral performance scores. CONCLUSIONS: Identification of predictors of successful resuscitation following cardiac arrest is important for risk stratification. Ongoing appraisal of in-hospital cardiac arrests through a multicentre registry could improve clinical outcomes. PMID- 15271174 TI - Patient characteristics and quality of life among a sample of Australian chronic pain clinic attendees. AB - BACKGROUND: Multidisciplinary chronic pain management programs have proliferated widely in recent decades. The clinical characteristics of patients attending these clinics are becoming the subject of increased research. Recent European data suggests that patients attending these clinics report very low quality of life. AIMS: The present study profiles an Australian population in terms of demographics, clinical characteristics and quality of life, as measured by the Short Form 36 Quality of Life Questionnaire (SF-36). METHODS: Data were collected prospectively from consecutive patients presenting to a multidisciplinary chronic pain clinic at a major Sydney metropolitan teaching-hospital. Cross-sectional analysis of demographic and clinical characteristics and quality of life were then undertaken. RESULTS: Descriptive analysis of demographics and clinical characteristics suggest a patient population group reporting significant pain severity and reduced quality of life. The comparison of SF-36 domain scores between clinic patients and Australian norm values indicates a greatly reduced score on all SF-36 domains for clinic patients. Pain clinic patients reported the most profound effect upon quality of life in the role physical, physical function and social function domains of the SF-36. Stepwise multiple regression indicated impaired coping ability and depressive disability as the most significant correlates of low quality of life. CONCLUSION: Patients who attend chronic pain clinics are likely to report low quality of life with an inability to cope. These findings suggest that future intervention research should explore the impacts of behavioural and self-management interventions. Psychological distress and ability to cope could be used as indices of improvement. PMID- 15271175 TI - Aciclovir or ganciclovir universal prophylaxis of cytomegalovirus infection in liver transplantation: an economic analysis. AB - BACKGROUND: Cytomegalovirus (CMV) following orthotopic liver transplantation can result in significant morbidity and mortality. Prophylaxis with oral aciclovir (ACV) or ganciclovir (GCV) for all transplant recipients (universal prophylaxis) may be beneficial, but which agent is more cost-effective is unknown. METHODS: A single centre, retrospective study of all patients who had OLT at the Western Australian Liver Transplantation Service was performed. Patients received ACV from 1992 to 1998, and GCV from 1999 to 2001. A comparative cost-effectiveness analysis for the two groups was performed based on the mean total cost of the number of cases of CMV infection and disease as the clinical end-point. RESULTS: The ACV group comprised of 55 patients and there were 24 in the GCV group. The incidence of CMV disease was 7% and 4% for the ACV and GCV groups, respectively (P > 0.05). For CMV infection it was 16% and 8%, respectively (P > 0.05). GCV prevented more cases of CMV infection and disease than ACV but at an incremental cost of dollars A20,000 (dollars US10,172) per case prevented. Overall, ACV was more cost-effective than GCV by dollars A2200 (dollars US1119) per person. The cost benefit of ACV was derived principally through a reduced pharmaceutical cost. Both agents were well tolerated without development of antiviral resistance. CONCLUSIONS: Universal prophylaxis of CMV infection-following liver transplantation with aciclovir is more cost-effective than with ganciclovir. PMID- 15271176 TI - Repetitive strain injuries: has the Australian epidemic burnt out? AB - In the 1980s Australia experienced an epidemic of medically certified claims for non-specific arm symptoms described as repetitive strain injury. Although a number of factors were mooted as causal of the epidemic, no single factor emerged as a compelling putative candidate. The present paper discusses the results of research which was published only after the epidemic had waned. It provides possible insights into the rise and fall of repetitive strain injury. PMID- 15271177 TI - Obstructive sleep apnoea and cardiovascular disease. AB - Obstructive sleep apnoea (OSA) leads to both acute and chronic physiological effects on the cardiovascular system. There is now a large amount of evidence showing that OSA is independently associated with a wide spectrum of clinical cardiovascular disease (CVD). Evidence for a causative effect of OSA is strongest for hypertension, but is weaker for other cardiovascular disorders. Large prospective trials are ongoing and when results become available the link between OSA and CVD is likely to be strengthened. Treatment of OSA with continuous positive airway pressure has been shown to improve blood pressure, particularly in those with hypertension, and also left ventricular ejection fraction in those with congestive heart failure. Given the high prevalence of OSA in the community and its effects on the cardiovascular system, symptoms of this disorder should be sought in patients being investigated or treated for CVD. PMID- 15271178 TI - Scorpion stings in Australia: five definite stings and a review. AB - Despite scorpions being locally abundant in many parts of Australia, scorpion sting is a poorly defined clinical condition in Australia. Many health-care workers are unaware of the effects of their stings and scorpions are often feared based on their international reputation. Five scorpion stings that occurred in different parts of Australia where the scorpion was caught at the time of the sting and identified by a professional arachnologist are reported in the present paper. The spectrum of clinical effects of scorpion stings in Australia and the potential for significant effects are discussed. These cases and recent prospective case series demonstrate that in Australia scorpion stings cause only minor effects. The main effect is localized pain lasting for several hours, associated less commonly with systemic effects, local numbness and paraesthesia. Most stings are from smaller scorpions from the family Buthidae and often occur indoors at night. The stings from Australian buthid scorpions cause more severe effects than from the larger species in the families Urodacidae (genus Urodacus) and Liochelidae (genus Liocheles). PMID- 15271179 TI - Staged autologous peripheral blood progenitor cell transplantation for Ewing sarcoma and rhabdomyosarcoma. PMID- 15271181 TI - Displaced colon: Chilaiditi's sign. PMID- 15271180 TI - Ethics and public health. AB - This paper examines the differences between clinical and public health ethics and provides several examples of contemporary public health challenges that pose ethical questions. The relations among ethics, rights and obligations are explored, because public health philosophers propose these as different ways for public health to align its activities to what we value. Resources for education in relation to the ethics of public health are identified. PMID- 15271182 TI - Hepatotoxicity possibly due to paracetamol with carbamazepine. PMID- 15271183 TI - Management of spontaneous pneumothorax. PMID- 15271184 TI - Ramipril: clinical and economic benefits. PMID- 15271186 TI - Genetic testing: a round table conversation. PMID- 15271187 TI - Genetic testing: a round table discussion. PMID- 15271188 TI - Genetic testing: a round table conversation. PMID- 15271189 TI - Genetic testing: a round table conversation. PMID- 15271191 TI - Changing face of colonoscopy: a comparison between audits of colonoscopy at Wellington Hospital from 1987 to 2002. PMID- 15271193 TI - Ciclopirox: a broad-spectrum antifungal with antibacterial and anti-inflammatory properties. PMID- 15271194 TI - Efficacy of different concentrations of ciclopirox shampoo for the treatment of seborrheic dermatitis of the scalp: results of a randomized, double-blind, vehicle-controlled trial. AB - BACKGROUND: Seborrheic dermatitis is a common inflammatory skin disorder affecting 1-3% of the population. It is thought to be linked to dandruff via a common etiology, yeasts of the genus Malassezia. Ciclopirox is a broad-spectrum, hydroxypyridone-derived, synthetic antifungal agent with anti-inflammatory properties. METHODS: A total of 203 patients were enrolled in this vehicle controlled, double-blind, randomized study designed to compare vehicle with three different concentrations of ciclopirox shampoo: 0.1%, 0.3% and 1%, with each applied twice a week. The main efficacy parameters were based on 6-point ordinal scales describing the disease's signs and symptoms (scaling, inflammation and itching), global status of disease, and global change in disease. RESULTS: A tendency towards improvement of the sum score from baseline was found in all ciclopirox treatment groups. The most pronounced improvement was found in the ciclopirox 1% group, which changed from a baseline sum score of 8.3 to 4.4 at the end of the 4-week study period (P-value vs. vehicle 0.0372). In addition, the therapeutic index showed increasing efficacy with the use of increased concentrations of ciclopirox. CONCLUSIONS: The study supports the use of 1% ciclopirox shampoo in the treatment of seborrheic dermatitis of the scalp. Furthermore, ciclopirox shampoo at each concentration was found to be safe and well tolerated. PMID- 15271195 TI - Rationale of frequency of use of ciclopirox 1% shampoo in the treatment of seborrheic dermatitis: results of a double-blind, placebo-controlled study comparing the efficacy of once, twice, and three times weekly usage. AB - BACKGROUND: Seborrheic dermatitis is a common inflammatory skin disorder. Although the precise etiology of seborrheic dermatitis is uncertain, yeasts of the genus Malassezia have been implicated. Ciclopirox is a broad-spectrum, hydroxypyridone-derived, synthetic antifungal agent with anti-inflammatory properties. METHODS: A total of 183 patients were enrolled in this randomized, parallel-group, double-blind, vehicle-controlled trial designed to compare three different application frequencies of ciclopirox 1% shampoo: once, twice, and three times weekly. The main efficacy parameters were based on 6-point ordinal scales describing the disease's signs and symptoms (scaling, inflammation and itching), global status of seborrheic dermatitis, and global change of seborrheic dermatitis. RESULTS: Each application frequency of ciclopirox 1% shampoo was found to significantly improve the signs and symptoms of seborrheic dermatitis of the scalp after 4 weeks of treatment. Furthermore, the therapeutic index calculated from the global change in seborrheic dermatitis score increased with increasing application frequency; therapeutic index scores increased from vehicle (1.25) to ciclopirox 1% once (3.30), twice (3.50), and three times (3.56) weekly. No serious adverse events were recorded during the course of the study. CONCLUSIONS: Ciclopirox 1% shampoo, applied once, twice or three times weekly, is a safe and acceptable preparation that improves seborrheic dermatitis of the scalp, suggesting that this is an alternative treatment for this indication. PMID- 15271196 TI - Safety and efficacy of ciclopirox 1% shampoo for the treatment of seborrheic dermatitis of the scalp in the US population: results of a double-blind, vehicle controlled trial. AB - BACKGROUND: Seborrheic dermatitis is a common inflammatory skin disorder. Yeasts of the genus Malassezia have been implicated in the etiology of seborrheic dermatitis, although this connection remains controversial. Ciclopirox is a synthetic, hydroxypyridone-derived, broad-spectrum antifungal agent with anti inflammatory properties. METHODS: A total of 499 US patients with seborrheic dermatitis of the scalp were randomized to apply either ciclopirox shampoo 1% or vehicle twice weekly for 4 weeks. The main efficacy parameters were based on 6 point ordinal scales describing the disease's signs and symptoms (scaling, erythema and itching) and a 6-point scale providing a global evaluation of the status of seborrheic dermatitis. RESULTS: Ciclopirox was significantly better than vehicle in effectively treating seborrheic dermatitis. 'Effective treatment' (score of 0 or 1 for disease status, scaling and erythema) was achieved in 26.0% of ciclopirox-treated patients compared with 12.9% of vehicle-treated patients (P = 0.0001; OR: 2.383, 95% CI: 1.494-3.799). The majority of subjects experienced adverse events that were mild in intensity, with skin and appendage reactions the most commonly reported, at similar frequency in both groups. CONCLUSIONS: Ciclopirox shampoo 1% is effective and safe in the treatment of seborrheic dermatitis of the scalp. PMID- 15271197 TI - Testicular dysgenesis syndrome: new epidemiological evidence. PMID- 15271198 TI - The A-type cyclins and the meiotic cell cycle in mammalian male germ cells. AB - There are two mammalian A-type cyclins, cyclin Al and A2. While cyclin A1 is limited to male germ cells, cyclin A2 is widely expressed. Cyclin A2 promotes both Gl/S and G2/M transitions in somatic cells and cyclin A2-deficient mice are early embryonic lethal. We have shown that cyclin Al is essential for passage of spermatocytes into meiosis I (MI) by generating mice null for the cyclin A1 gene Ccna1. Both Ccna1(-/-) males and females were healthy but the males were sterile because of a cell cycle arrest before MI. This arrest was associated with desynapsis abnormalities, low M-phase promoting factor activity, and apoptosis. We have now determined that human cyclin A1 is expressed in similar stages of spermatogenesis and are exploring its role in human male infertility and whether it may be a novel target for new approaches for male contraception. PMID- 15271199 TI - Meiotic segregation of translocations during male gametogenesis. AB - Balanced reciprocal and Robertsonian translocations are the most common structural chromosomal abnormalities in humans. Generally, they are without consequence for the carrier, but for various degrees of oligoasthenoteratozoospermia in men. As these carriers can produce a significant percentage of gametes with an unbalanced combination of the parental rearrangement, there is a more or less significant risk, according to cases, of chromosomal imbalances for their offspring. Therefore, techniques were developed to study the meiotic segregation of these translocations in males. Direct investigation of human sperm chromosomes became possible by karyotyping spermatozoa after penetration of zona-free hamster oocytes and, more recently, using fluorescent in situ hybridization (FISH). This paper reviews the results obtained using these techniques in Robertsonian and reciprocal translocations. The studies on spermatozoa from translocation carriers help the comprehension of the mechanisms of the meiotic segregation. They should be integrated in the genetic exploration of the infertile men, in order to give them a personalized risk assessment of unbalanced spermatozoa, specially as a correlation was found recently between the percentage of abnormal spermatozoa and that of abnormal embryos. PMID- 15271200 TI - Risk factors for hypospadias in Norwegian boys - association with testicular dysgenesis syndrome? AB - It has been proposed that hypospadias, cryptorchidism and testicular cancer, as well as decreasing sperm quality are symptoms of an underlying entity called testicular dysgenesis syndrome (TDS). We wanted to study the risk factors for hypospadias and compare them with those of the other conditions belonging to TDS. A large case-control study was undertaken on data on all live-born boys registered in the Medical Birth Registry of Norway during the period 1967-1998 (n = 961 396; hypospadias cases = 2382). Logistic regression analysis was used to study the association between potential risk factors and hypospadias, estimated by odds ratio (OR). The risk factors for hypospadias were divided into four categories: (i) maternal characteristics, e.g. low parity [p(trend) < 0.001], hypertension (OR = 1.49) and bleeding (OR = 1.39) during index pregnancy, and (pre)eclampsia (OR = 1.84); (ii) complications during delivery, e.g. retained placenta (OR = 1.67) or Caesarean section (OR = 1.36); (iii) characteristics of the newborn, e.g. low birth weight [p(trend) < 0.001], small for gestational age (OR = 2.16), and presence of congenital malformations other than hypospadias (OR = 2.72), e.g. inguinal hernia (OR = 5.65); (iv) prevalence among relatives of hypospadias cases, e.g. brother with hypospadias (OR = 20.81). The novel finding of retained placenta as a risk factor indicates that early malfunction of placenta could be a causative factor for hypospadias. When comparing with previously published risk factors for hypospadias, cryptorchidism and testicular cancer, we found that the following risk factors were common to all three conditions: low parity, low birth weight, low gestational age, inguinal hernia, bleeding during pregnancy and Caesarean section. In conclusion, our results support the notion that the conditions of TDS share risk factors. PMID- 15271201 TI - A novel instrument-independent no-scalpel vasectomy - a comparative study against the standard instrument-dependent no-scalpel vasectomy. AB - An instrument-independent no-scalpel vasectomy (IINSV) technique is reported. This technique does not use the standard specific instruments, but comparatively retains the advantages of minimally invasive instrument-dependent no-scalpel vasectomy (IDNSV). Between July 1999 and June 2002, 449 men were prospectively randomized to be vasectomized at two hospitals in Taipei. Of those who accepted, 215 underwent IDNSV at one hospital and the remaining 234 underwent IINSV at the other. The intra-operative conditions of each group were recorded. The postoperative pain and life conditions were self-reported through a questionnaire that had been carefully designed prior to the operations, in which the pain level was assessed using a 10-cm visual analogue scale under varying situations. Men vasectomized using the IINSV method experienced less operation time and postoperative complications (haematomas, infections and granulomas) (p < 0.05). There were no significant differences between the two groups with respect to incision length, postoperative pain, pain at coitus, time of return to work, time of resuming intercourse, self-reported satisfaction in sexual life, postoperative psychological status, postoperative body weight change and vasectomy failure as evidenced by sperm analysis (p > 0.05 for all items). Thus, the IINSV technique can offer an alternative option for vasectomists whenever the specific instruments of standard no-scalpel vasectomy are unavailable. The IINSV technique shortens the operation time and reduces the incidence of operative complications when compared with the IDNSV technique, while still retaining the advantages of minimally invasive vasectomy. PMID- 15271202 TI - The role of reactive oxygen species and apoptosis in the pathogenesis of varicocele in a rat model and efficiency of vitamin E treatment. AB - We investigated role of reactive oxygen species (ROS) and apoptosis in the pathogenesis of infertility in experimental model of varicocele. The protective effect of vitamin E was also examined. Three groups of rats were constructed as the first group had sham operation, experimental varicoceles were established by partial ligation of the left renal vein in later two groups. Third group had received vitamin E. Production of ROS was determined by chemiluminescence assay (CL). The in situ end labelling technique was utilized to investigate apoptosis. Tissue vitamin E levels were measured by high performance liquid chromatography. The differences between luminol enhanced CL levels of groups were not statistically significant. However, the difference between CL levels of lucigenin probe in left testicles of sham and varicocele groups were statistically significant ( p = 0.0007). Similarly, the results of the third group receiving vitamin E significantly differed from the varicocele group ( p = 0.0025). The difference of apoptotic index was also statistically significant between sham and varicocele groups ( p = 0.0038). Although the values of apoptotic index detected in the vitamin E group were lower compared with the varicocele group, the difference was not significant. This study proposes that ROS production and apoptosis in the testicles were induced with experimental varicocele. Vitamin E had a protective role. An increased rate of apoptosis with experimental varicocele suggests a molecular alteration, which may involve ROS overproduction as the triggering mechanism. Consequently, this study indicates an association between varicocele and infertility at molecular level through stimulation of ROS and apoptosis. PMID- 15271203 TI - Disrupted expression of intermediate filaments in the testis of rhesus monkey after experimental cryptorchidism. AB - Cytoskeletons in Sertoli cell play an important role in process of spermatogenesis. The expression and distribution of the intermediate filaments, vimentin, keratin and desmin, were studied in the Sertoli cells of the cryptorchid testis of rhesus monkey. Vimentin was localized in the perinuclear region of Sertoli cells of the normal testis. An intense increase in vimentin immunoreactivity was observed with appearance of disorganized staining in the Sertoli cells of the cryptorchid testes. Cytokeratin 18, a marker of immature Sertoli cells, re-expressed in the cells of the adult cryptorchid testes. Desmin was also observed in the Sertoli cells in addition to the peritubular myoid cells on 30 days after the cryptorchid operation. These data suggest that Sertoli cells in primate can be affected by the heat stress. The altered changes in intermediate filaments could be possible to induce the Sertoli cell functional changes that would partially contribute to the germ cell apoptosis leading to azoospermia or oligozoospermia. PMID- 15271204 TI - EAA/EMQN best practice guidelines for molecular diagnosis of y-chromosomal microdeletions. State of the art 2004. AB - Microdeletions of the Y chromosome are the second most frequent genetic cause of spermatogenetic failure in infertile men after the Klinefelter syndrome. The molecular diagnosis of Y-chromosomal microdeletions is routinely performed in the workup of male infertility in men with azoospermia or severe oligozoospermia. Since 1999, the European Academy of Andrology (EAA) and the European Molecular Genetics Quality Network (EMQN) support the improvement of the quality of the diagnostic assays by publication of the laboratory guidelines for molecular diagnosis of Y-chromosomal microdeletions and by offering external quality assessment trials. The present revision of the 1999 laboratory guidelines summarizes the results of a 'Best Practice Meeting' held in Florence (Italy) in October 2003. The basic protocol for microdeletion screening suggested in the 1999 guidelines proved to be very accurate, sensitive and robust. In the light of the recent advance in the knowledge of the Y chromosome sequence and of the mechanism of microdeletion it was agreed that the basic 1999 protocol, based on two multiplex polymerase chain reactions each covering the three AZF regions, is still fully valid and appropriate for accurate diagnosis. PMID- 15271205 TI - Transcription of a lepidopteran cytochrome P450 promoter is modulated by multiple elements in its 5' UTR and repressed by 20-hydroxyecdysone. AB - The biochemical response to the phytochemical xanthotoxin encountered in the diet of black swallowtail larvae is the induction of P450s capable of detoxifying this and other toxic furanocoumarins. As the xenobiotic response element to xanthotoxin (XRE-xan) is necessary but not sufficient for transcription of the CYP6B1v3 gene in Sf9 cells, sequences upstream of it, such as a putative EcRE, and downstream of it, such as a putative C/EBP binding site and Inr, have been tested for their roles in regulation. Mutation of the putative EcRE has indicated that it affects basal transcription of this promoter but not repression by 20 hydroxyecdysone. Mutation of the more proximal promoter sequence, including the C/EBP and Inr, have indicated that many core promoter elements between the TATA box and translation start site modulate basal and xanthotoxin-inducible expression of this composite promoter. PMID- 15271206 TI - Storage and secretion of the peritrophic matrix protein Ag-Aper1 and trypsin in the midgut of Anopheles gambiae. AB - The gene Ag-Aper1 encodes a peritrophic matrix (PM) protein from the mosquito Anopheles gambiae. Ag-Aper1 gene expression and protein localization in the mosquito midgut were studied during the course of a blood meal. Ag-Aper1 mRNA abundance does not change appreciably during the course of blood ingestion and digestion. Prior to a blood meal, the protein is stored in secretory vesicles of midgut epithelial cells. Moreover, Ag-Aper1 colocalizes to the same secretory vesicles as trypsin, indicating that these proteins use a common secretory pathway. Blood feeding triggers the secretion of vesicle contents into the midgut lumen, after which Ag-Aper1 is incorporated into the PM. Newly synthesized Ag Aper1 protein was again detected within the midgut epithelial cells at 60 h after blood ingestion. PMID- 15271207 TI - Distribution and prevalence of Wolbachia in Japanese populations of Lepidoptera. AB - Wolbachia are cytoplasmically inherited bacteria that are reported to infect at least 18-30% of all insect species. Our survey of Lepidoptera indicated that 44.9% of forty-nine species and 77.8% of nine families tested positive for Wolbachia using PCR with wsp primers. Nineteen species had not been described previously as infected. In particular, although Pieris rapae, which is a common species in Japan, is infected by Wolbachia, the prevalence was very low (3.4%) and there were some localities where Wolbachia could not be detected. The probability of detection of Wolbachia depends on the number of screened individuals of P. rapae. The results indicate that the actual number of species that are positive for Wolbachia may be higher than previously reported. PMID- 15271208 TI - Molecular discrimination of Wolbachia in the Culex pipiens complex: evidence for variable bacteriophage hyperparasitism. AB - The medically important members of the Culex pipiens species complex provide an enigma for systematists, evolutionary biologists, and vector biologists. The species complex is composed of forms that differ in their ecology, behaviour, physiology and vector competence. Cytoplasmic incompatibility (CI) caused by endosymbiotic Wolbachia bacteria is thought to play an important role in restricting gene flow and the evolution of the Culex complex. Here we describe the first molecular marker useful for discriminating between Wolbachia infections in Culex. A putative bacteriophage locus (orf7) varies between Culex forms in copy number and sequence. We provide evidence that the orf7 loci are strictly associated with Wolbachia and are maternally inherited. PMID- 15271209 TI - Variation in an intron sequence of the voltage-gated sodium channel gene correlates with genetic differentiation between Anopheles gambiae s.s. molecular forms. AB - We present the results of a geographical survey of genetic variation in Anopheles gambiae M and S molecular forms from ten African countries at Intron I of the voltage-gated sodium channel gene. We found two major haplotypes separated by a single mutational step, which cosegregate almost completely with the rDNA sites that identify M and S, consistent with previous estimates of strong reductions of gene flow between the two forms. We also report ten additional haplotypes stemming from the two major haplotypes, mostly present in single localities. The low levels of genetic variation found in this intronic region are discussed in light of a possible selective sweep. These findings offer additional elements to the ongoing debate on the amount of genetic differentiation and isolation between the two molecular forms and on their taxonomic status. PMID- 15271210 TI - Genetic mapping of genes conferring permethrin resistance in the malaria vector, Anopheles gambiae. AB - Resistance to permethrin in an East African population of the major malaria vector, Anopheles gambiae is multifactorial. A mutated sodium channel allele and enhanced insecticide metabolism contribute to the resistance phenotype. We used microsatellite markers to scan the genome for quantitative trait loci (QTL) associated with permethrin resistance. Two major and one minor QTL were identified. The first QTL, rtp1, colocalizes with the sodium channel gene on chromosome 2L thus further supporting the importance of mutations in this gene in conferring permethrin resistance. The second two loci are located on the third chromosome and one of these, rtp2, flanks a large cluster of cytochrome P450 genes. Further detailed mapping of these regions will help elucidate the molecular mechanisms of metabolic resistance to insecticides. PMID- 15271211 TI - Identification and molecular characterization of two immune-responsive chitinase like proteins from Anopheles gambiae. AB - Two haemolymph proteins that are processed rapidly and specifically in response to exposure to bacteria have been identified from Anopheles gambiae. Both proteins, Anopheles gambiae bacteria-responsive 1 (AgBR1) and AgBR2, are similar to chitinases but belong to a family of proteins that have lost chitinolytic activity. AgBR1 and AgBR2 are converted to smaller forms in vivo or in vitro on exposure to bacteria, and AgBR2 also can be processed on exposure to peptidoglycan alone. AgBR1 and AgBR2 do not bind to bacteria or chitin beads. The AgBR1 and AgBR2 genes are expressed in all developmental stages. In adults, AgBR1 expression is restricted to the fat body, whereas AgBR2 is expressed in many tissues. PMID- 15271212 TI - Baculovirus and dsRNA induce Hemolin, but no antibacterial activity, in Antheraea pernyi. AB - Hemolin is one of the haemolymph proteins most strongly induced upon bacterial infection in Lepidoptera. When we applied RNA interference (RNAi) to suppress Hemolin expression in the Chinese oak silk moth Antheraea pernyi, we discovered that Hemolin is induced by double-stranded RNA (dsRNA) per se. As dsRNA is recognized as a virus pattern molecule, we then investigated the effect of a baculovirus (ApNPV) infection. We found that Hemolin is induced and expressed with similar kinetics as upon dsRNA injection. Notably, no Attacin gene expression or antibacterial activity was recorded. When baculovirus and high amounts of dsRNA were coinjected, the viral symptoms appeared earlier with Hemolin dsRNA than with GFP dsRNA. This indicates that silencing of hemolin affected the progress of the viral infection. PMID- 15271213 TI - Molecular cloning and expression patterns of the molt-regulating transcription factor HHR3 from Helicoverpa armigera. AB - Molt-regulating transcription factors, hormone receptor 3 (HR3), play important roles in regulating expression of tissue-specific genes involved in insect molting and metamorphosis. A 1668 bp cDNA encoding a molt-regulating transcription factor (HHR3) was cloned from Helicoverpa armigera, which encodes a protein made up of 556 amino acids. This 62 kDa protein was found to have an isoelectric point (pI) of 6.52. There was no signal peptide or N-glycosylation site found in this cDNA. A DNA-binding region signature of nuclear hormone receptor was found from amino acids 107-133. A possible outside to inside transmembrane helice was found from amino acids 72-90. Northern blots of the larvae revealed five bands of HHR3 named as band 0, 1, 2, 3 and 4 with molecular masses determined as 2.1, 2.6, 3.6, 4.5 and 5.5 kb, respectively. The expression patterns of HHR3 in vivo were variable with developmental stages and tissues. Results showed that band 1-4 of HHR3 was only briefly expressed during molting, which suggested these bands are involved in the regulation of molting cascade, whereas band 0 was expressed in both molting and feeding larvae. Band 1 and 2 of HHR3 could be induced from epidermis of newly molted 6th instar larvae by non steroidal ecdysone agonist, RH-2485. PMID- 15271214 TI - Population structure of the beetle pests Phyllodecta vulgatissima and P. vitellinae on UK willow plantations. AB - Phyllodecta (= Phratora) vulgatissima and P. vitellinae (Coleoptera: Chrysomelidae) are important pests of willows and poplars. Their differences in host species preference may provide a non-chemical control strategy for pest control. However, little is known about population structure with respect to hosts, regions or seasons. Using five microsatellites, 850 P. vulgatissima and 1100 P. vitellinae individuals, comprising 17 and 22 UK samples, respectively, were genotyped. High diversity was observed at all loci. Migrant numbers exchanged per generation (Nm) were high (2.1-12.6 for P. vulgatissima and 0.9 12.2 for P. vitellinae), suggesting high genetic exchange between samples. Estimates of population differentiation (FST) and analyses of the data using Bayesian methods (Partition and Structure) showed little evidence of subdivision in relation to geography, sampling time or host. PMID- 15271215 TI - Stress and transcriptional regulation of tick ferritin HC. AB - We previously identified a partial Dermacentor variabilis cDNA encoding ferritin HC (HC) subunit homolog (DVFER) that was differentially upregulated in Rickettsia montanensis infected ticks (Mulenga et al., 2003a). We have used rapid amplification of cDNA ends to clone full-length DVFER cDNA and its apparent ortholog from the wood tick, D. andersoni (DAFER), both of which show high sequence similarity to vertebrate than insect ferritin. Both DVFER and DAFER contain the stem-loop structure of a putative iron responsive element in the 5' untranslated region (nucleotide positions, 16-42) and the feroxidase centre loop typical for vertebrate ferritin HC subunits. Quantitative Western and Northern blotting analyses of protein and RNA from unfed and partially fed whole tick as well as dissected tick tissues demonstrated that DVFER is constitutively and ubiquitously expressed. Based on densitometric analysis of detected protein and mRNA bands, DVFER is predominantly expressed in the midgut, and to a lesser extent in the salivary glands, ovary and fatbody. Sham treatment (mechanical injury) and Escherichia coli challenge of D. variabilis ticks stimulated statistically significant (approximately 1.5- and approximately 3.0-fold, respectively) increases in DVFER mRNA abundance over time point matched naive control ticks. These data suggest that DVFER mRNA is nonspecifically up regulated in response to mechanical injury or bacterial infection induced stress. PMID- 15271216 TI - Use of a PCR-based approach for sequencing whole mitochondrial genomes of insects: two examples (cockroach and dragonfly) based on the method developed for decapod crustaceans. AB - Recent development of a PCR-based approach for sequencing vertebrate mitochondrial genomes has attracted much attention as being more rapid and economical than traditional methods using cloned mtDNA and primer walking. Such a method has not been available for insect mitochondrial genomes, despite widespread use of them for the molecular phylogenetic, biogeographical and population genetic markers. A recently developed PCR-based approach for sequencing whole mitochondrial genomes of decapod crustaceans, which included the design of many versatile PCR primers for the latter, was applied with the same primers sets to mitochondrial genomes of two insects, smoky-brown cockroach Periplaneta fuliginosa (Serville, 1839) and skimmer dragonfly Orthetrum triangulare melania (Selys, 1883). Almost the entire region of the two mitochondrial genomes was successfully sequenced. Features of the two mitochondrial genomes are described and the usefulness of this PCR-based approach for sequencing insect mitochondrial genomes demonstrated. PMID- 15271217 TI - Identification of mariner elements from house flies (Musca domestica) and German cockroaches (Blattella germanica). AB - Full-length mariner elements were isolated and sequenced from house flies (Musca domestica) and German cockroaches (Blattella germanica). The amino acid sequence of the house fly mariner element (accession number: AF373028) showed 99.5% identity with Mos1 and peach elements, whereas the German cockroach mariner element (accession number: AF355143) showed 98.8% and 99.8% identity, respectively. Sequence analysis revealed that the mariner elements in house flies and German cockroaches differed from the active Mos1 mariner element by seven and 15 nucleotides, respectively. Four essential nucleotide substitutions at positions 64, 154, 305, and 1203, which have been proposed to contribute to the loss of activity of the inactive elements, were detected in the German cockroach mariner element. In contrast, although the mariner element in house flies contained substitutions at positions 64, 154, and 305, it retained T at position 1203, identical to active mariner elements. Mariner is present in approximately eight copies in the German cockroach genome. PMID- 15271218 TI - Low fasting low high-density lipoprotein and postprandial lipemia. AB - BACKGROUND: Low levels of high density lipoprotein (HDL) cholesterol and disturbed postprandial lipemia are associated with coronary heart disease. In the present study, we evaluated the variation of triglyceride (TG) postprandially in respect to serum HDL cholesterol levels. RESULTS: Fifty two Greek men were divided into 2 main groups: a) the low HDL group (HDL < 40 mg/dl), and b) the control group. Both groups were further matched according to fasting TG (matched low HDL, and matched-control groups). The fasting TG concentrations were higher in the low HDL group compared to controls (p = 0.002). The low HDL group had significantly higher TG at 4, 6 and 8 h postprandially compared to the controls (p = 0.006, p = 0.002, and p < 0.001, respectively). The matched-low HDL group revealed higher TG only at 8 h postprandially (p = 0.017) compared to the matched control group. ROC analysis showed that fasting TG > or = 121 mg/dl have 100% sensitivity and 81% specificity for an abnormal TG response (auc = 0.962, p < 0.001). CONCLUSIONS: The delayed TG clearance postprandially seems to result in low HDL cholesterol even in subjects with low fasting TG. The fasting TG > 121 mg/dl are predictable for abnormal response to fatty meal. PMID- 15271219 TI - An active electrode for biopotential recording from small localized bio-sources. AB - BACKGROUND: Laser bio-stimulation is a well-established procedure in Medical Acupuncture. Nevertheless there is still a confusion as to whether it works or the effect is just placebo. Although a plethora of scientific papers published, showing positive clinical results, there is still a lack of objective scientific proofs about the bio-stimulation effect of lasers used in Acupuncture. The objective of this work was to design and build a body surface electrode and an amplifier for biopotential recording from acupuncture points, considered here as small localized bio-sources (SLB). The design is aimed for studying SLB potentials provoked by laser stimulus, in search for objective proofs of the bio stimulation effect of lasers used in Medical Acupuncture. METHODS: The active electrode presented features a new adjustable anchoring system and fractionation of the biopotential amplifier between the electrode and the cabinet's location. The new adjustable electrode anchoring system is designed to reduce the electrode skin contact impedance, its variation and motion artifacts. That is achieved by increasing the electrode-skin tension and decreasing its relative movement. Additionally the sensing element provides local constant skin stretching thus eliminating the contribution of the skin potential artifact. The electrode is attached to the skin by a double-sided adhesive pad, where the sensing element is a stainless steel, 4 mm in diameter. The fractionation of the biopotential amplifier is done by incorporating the amplifier's front-end op-amps at the electrodes, thus avoiding the use of extra buffers. The biopotential amplifier features two selectable modes of operation: semi-AC-mode with a -3 dB bandwidth of 0.32-1000 Hz and AC-mode with a bandwidth of 0.16-1000 Hz. RESULTS: The average measured DC electrode-skin contact impedance of the proposed electrode was 450 kOmega, with electrode tension of 0.3 kg/cm2 on an unprepared skin of the inner forearm. The peak-to-peak noise voltage measured at the amplifier output, with input terminals connected to common, was 10 mVp-p, or 2 microVp-p referred to the input. The common-mode rejection ratio of the amplifier was 96 dB at 50 Hz, measured with imbalanced electrodes' impedances. The prototype was also tested practically and sample records were obtained after a low intensity SLB laser stimulation. All measurements showed almost a complete absence of 50 Hz interference, although no electrolyte gel or skin preparation was applied. CONCLUSION: The results showed that the new active electrode presented significantly reduced the electrode-skin impedance, its variation and motion artifact influences. This allowed SLB signals with relatively high quality to be recorded without skin preparation. The design offers low noise and major reduction in parts, size and power consumption. The active electrode specifications were found to be better or at least comparable to those of other existing designs. PMID- 15271220 TI - World health system performance revisited: the impact of varying the relative importance of health system goals. AB - BACKGROUND: In 2002, the World Health Organization published a health system performance ranking for 191 member countries. The ranking was based on five indicators, with fixed weights common to all countries. METHODS: We investigate the feasibility and desirability of using mathematical programming techniques that allow weights to vary across countries to reflect their varying circumstances and objectives. RESULTS: By global distributional measures, scores and ranks are found to be not very sensitive to changes in weights, although differences can be large for individual countries. CONCLUSIONS: Building the flexibility of variable weights into calculation of the performance index is a useful way to respond to the debates and criticisms appearing since publication of the ranking. PMID- 15271221 TI - A quantitative analysis of qualitative studies in clinical journals for the 2000 publishing year. AB - BACKGROUND: Quantitative studies are becoming more recognized as important to understanding health care with all of its richness and complexities. The purpose of this descriptive survey was to provide a quantitative evaluation of the qualitative studies published in 170 core clinical journals for 2000. METHODS: All identified studies that used qualitative methods were reviewed to ascertain which clinical journals publish qualitative studies and to extract research methods, content (persons and health care issues studied), and whether mixed methods (quantitative and qualitative methods) were used. RESULTS: 60 330 articles were reviewed. 355 reports of original qualitative studies and 12 systematic review articles were identified in 48 journals. Most of the journals were in the discipline of nursing. Only 4 of the most highly cited health care journals, based on ISI Science Citation Index (SCI) Impact Factors, published qualitative studies. 37 of the 355 original reports used both qualitative and quantitative (mixed) methods. Patients and non-health care settings were the most common groups of people studied. Diseases and conditions were cancer, mental health, pregnancy and childbirth, and cerebrovascular disease with many other diseases and conditions represented. Phenomenology and grounded theory were commonly used; substantial ethnography was also present. No substantial differences were noted for content or methods when articles published in all disciplines were compared with articles published in nursing titles or when studies with mixed methods were compared with studies that included only qualitative methods. CONCLUSIONS: The clinical literature includes many qualitative studies although they are often published in nursing journals or journals with low SCI Impact Factor journals. Many qualitative studies incorporate both qualitative and quantitative methods. PMID- 15271222 TI - Family-based clusters of cognitive test performance in familial schizophrenia. AB - BACKGROUND: Cognitive traits derived from neuropsychological test data are considered to be potential endophenotypes of schizophrenia. Previously, these traits have been found to form a valid basis for clustering samples of schizophrenia patients into homogeneous subgroups. We set out to identify such clusters, but apart from previous studies, we included both schizophrenia patients and family members into the cluster analysis. The aim of the study was to detect family clusters with similar cognitive test performance. METHODS: Test scores from 54 randomly selected families comprising at least two siblings with schizophrenia spectrum disorders, and at least two unaffected family members were included in a complete-linkage cluster analysis with interactive data visualization. RESULTS: A well-performing, an impaired, and an intermediate family cluster emerged from the analysis. While the neuropsychological test scores differed significantly between the clusters, only minor differences were observed in the clinical variables. CONCLUSIONS: The visually aided clustering algorithm was successful in identifying family clusters comprising both schizophrenia patients and their relatives. The present classification method may serve as a basis for selecting phenotypically more homogeneous groups of families in subsequent genetic analyses. PMID- 15271223 TI - Caveolin-2 associates with intracellular chlamydial inclusions independently of caveolin-1. AB - BACKGROUND: Lipid raft domains form in plasma membranes of eukaryotic cells by the tight packing of glycosphingolipids and cholesterol. Caveolae are invaginated structures that form in lipid raft domains when the protein caveolin-1 is expressed. The Chlamydiaceae are obligate intracellular bacterial pathogens that replicate entirely within inclusions that develop from the phagocytic vacuoles in which they enter. We recently found that host cell caveolin-1 is associated with the intracellular vacuoles and inclusions of some chlamydial strains and species, and that entry of those strains depends on intact lipid raft domains. Caveolin-2 is another member of the caveolin family of proteins that is present in caveolae, but of unknown function. METHODS: We utilized a caveolin-1 negative/caveolin-2 positive FRT cell line and laser confocal immunofluorescence techniques to visualize the colocalization of caveolin-2 with the chlamydial inclusions. RESULTS: We show here that in infected HeLa cells, caveolin-2, as well as caveolin-1, colocalizes with inclusions of C. pneumoniae (Cp), C. caviae (GPIC), and C. trachomatis serovars E, F and K. In addition, caveolin-2 also associates with C. trachomatis serovars A, B and C, although caveolin-1 did not colocalize with these organisms. Moreover, caveolin-2 appears to be specifically, or indirectly, associated with the pathogens at the inclusion membranes. Using caveolin-1 deficient FRT cells, we show that although caveolin-2 normally is not transported out of the Golgi in the absence of caveolin-1, it nevertheless colocalizes with chlamydial inclusions in these cells. However, our results also show that caveolin-2 did not colocalize with UV-irradiated Chlamydia in FRT cells, suggesting that in these caveolin-1 negative cells, pathogen viability and very likely pathogen gene expression are necessary for the acquisition of caveolin-2 from the Golgi. CONCLUSION: Caveolin-2 associates with the chlamydial inclusion independently of caveolin-1. The function of caveolin-2, either in the uninfected cell or in the chlamydial developmental cycle, remains to be elucidated. Nevertheless, this second caveolin protein can now be added to the small number of host proteins that are associated with the inclusions of this obligate intracellular pathogen. PMID- 15271224 TI - Integrated web service for improving alignment quality based on segments comparison. AB - BACKGROUND: Defining blocks forming the global protein structure on the basis of local structural regularity is a very fruitful idea, extensively used in description, and prediction of structure from only sequence information. Over many years the secondary structure elements were used as available building blocks with great success. Specially prepared sets of possible structural motifs can be used to describe similarity between very distant, non-homologous proteins. The reason for utilizing the structural information in the description of proteins is straightforward. Structural comparison is able to detect approximately twice as many distant relationships as sequence comparison at the same error rate. RESULTS: Here we provide a new fragment library for Local Structure Segment (LSS) prediction called FRAGlib which is integrated with a previously described segment alignment algorithm SEA. A joined FRAGlib/SEA server provides easy access to both algorithms, allowing a one stop alignment service using a novel approach to protein sequence alignment based on a network matching approach. The FRAGlib used as secondary structure prediction achieves only 73% accuracy in Q3 measure, but when combined with the SEA alignment, it achieves a significant improvement in pairwise sequence alignment quality, as compared to previous SEA implementation and other public alignment algorithms. The FRAGlib algorithm takes approximately 2 min. to search over FRAGlib database for a typical query protein with 500 residues. The SEA service align two typical proteins within circa approximately 5 min. All supplementary materials (detailed results of all the benchmarks, the list of test proteins and the whole fragments library) are available for download on-line at http://ffas.ljcrf.edu/darman/results/. CONCLUSIONS: The joined FRAGlib/SEA server will be a valuable tool both for molecular biologists working on protein sequence analysis and for bioinformaticians developing computational methods of structure prediction and alignment of proteins. PMID- 15271225 TI - Guidelines for preventing health-care-associated pneumonia, 2003: buyer beware! PMID- 15271226 TI - Expiratory chest compression for atelectasis: No harm, no foul-oops! PMID- 15271228 TI - Effects of expiratory rib cage compression combined with endotracheal suctioning on gas exchange in mechanically ventilated rabbits with induced atelectasis. AB - INTRODUCTION: In Japan, expiratory rib cage compression (a chest physiotherapy technique) is frequently used with mechanically ventilated patients. It has not been determined whether rib cage compression combined with endotracheal suctioning improves oxygenation, ventilation, and mucus clearance. We evaluated the effects of rib cage compression with and without endotracheal suctioning on P(aO(2)), P(aCO(2)), dynamic compliance of the respiratory system (C(RS)), and mucus clearance in rabbits with induced atelectasis. METHODS: Anesthetized adult rabbits had an 18-gauge catheter placed into the airway, together with a tracheal tube via tracheostoma, and were mechanically ventilated. To create atelectasis, artificial mucus was infused into the airway via the catheter. Each rabbit was randomly assigned to one of 4 groups (= 7 in each): (1) control, (2) received endotracheal suctioning alone, (3) received rib cage compression alone, and (4) received both rib cage compression and endotracheal suctioning. After these interventions, for 30 min, each animal was placed supine without intervention for 120 min. RESULTS: In the groups that received rib cage compression, oxygenation, ventilation, and C. PMID- 15271229 TI - An evaluation of 2 new devices for nasal high-flow gas therapy. AB - BACKGROUND: The traditional nasal cannula with bubble humidifier is limited to a maximum flow of 6 L/min to minimize the risk of complications. We conducted a bench study of 2 new Food and Drug Administration-approved nasal cannula/humidifier products designed to deliver at flows> 6 L/min. METHODS: Using a digital psychrometer we measured the relative humidity and temperature of delivered gas from each device, at 5 L/min increments over the specified functional high-flow range. RESULTS: The Salter Labs unit achieved 72.5-78.7% relative humidity (5-15 L/min range) at ambient temperature (21-23 degrees C). The Vapotherm device achieved 99.9% relative humidity at a temperature setting of 37 degrees C (5-40 L/min). CONCLUSIONS: Both devices meet minimum humidification standards and offer practical new treatment options. The patient-selection criteria are primarily the severity of the patient's condition and cost. PMID- 15271230 TI - An assessment of the appropriateness of respiratory care delivered at a 450-bed acute care Veterans affairs hospital. AB - INTRODUCTION: Respiratory care is expensive and time-intensive, inappropriate care wastes resources, and failure to provide necessary and appropriate respiratory care may adversely affect patient outcomes. OBJECTIVE: To determine the appropriateness of basic respiratory care delivered at a 450-bed Veterans Affairs hospital during a 3-month interval. METHODS: We determined (1) the percentage of delivered respiratory care that was not indicated (based on standardized clinical practice guidelines), (2) the percentage of respiratory care that was indicated but not ordered (based on standardized clinical practice guidelines), and (3) the labor cost and potential savings of protocol-based respiratory care at our hospital. We selected 5 assessment days, occurring at 2 week intervals. All patients who received basic respiratory care underwent a complete respiratory care assessment, including medical records review, patient interview, physical assessment, and measurement of blood oxygen saturation (via pulse oximetry) and inspiratory capacity. Intensive care patients were excluded from the study. The assessment instrument provided a standardized format based on American Association for Respiratory Care clinical practice guidelines. RESULTS: We assessed 75 patients. A mean of 24.8% of the delivered respiratory therapies reviewed were not indicated. The percentages of ordered but not indicated therapies were: oxygen 17.7%; all categories of aerosolized medications (bronchodilators, mucolytics, anti-inflammatory agents) 32.4%; chest physiotherapy 37.5%; lung expansion therapy 7.7%. A mean of 11.8% of the patients assessed were not receiving respiratory care that was indicated. The percentages of indicated but not ordered therapies were: oxygen 5.3%; bronchodilator 5.3%; lung expansion therapy 36%. CONCLUSION: A mean of 24.8% of the basic respiratory care procedures delivered were not indicated and 11.8% of patients were not receiving care that was indicated. Inappropriate utilization of respiratory care services may increase costs and adversely affect morbidity, mortality, and duration of stay. We believe that implementation of respiratory care assessment protocols based on nationally accepted clinical practice guidelines can reduce unnecessary care, optimize care delivered, and may reduce costs and improve outcomes. PMID- 15271231 TI - Respiratory therapists' attitudes about participative decision making: relationship between managerial decision-making style and job satisfaction. AB - BACKGROUND: Studies of non-health-care work environments indicate that non managerial employee job satisfaction is higher in companies that use participative (as opposed to autocratic) decision making. It has not been determined whether managerial decision-making style influences job satisfaction among respiratory therapists (RTs) and which managerial decision-making style RTs prefer. METHODS: We surveyed Nebraska RTs' attitudes regarding their job satisfaction, their perceptions of their managers' decision-making styles (autocratic, consultative, and/or delegative), and which decision-making style they would prefer their managers to use. We sought to determine whether there is a significant correlation between RTs' perceptions of their managers' decision making styles and the RTs' job satisfaction. The study population was 792 licensed and practicing non-managerial RTs in Nebraska, from which we randomly selected 565 RTs to survey. The self-administered, descriptive survey used 2 Likert scales (one for decision-making style and one for job satisfaction) and inquired about 57 items. The survey was mailed on October 1, 1999. On October 28, 1999, we sent a second mailing to RTs who had not responded. RESULTS: We received 271 responses (response rate 47.9%). The respondents were generally satisfied with their jobs (mean +/- SD Minnesota Satisfaction Questionnaire score 73.46 +/- 11.63). The sub-scale scores ranged from 20 ("very dissatisfied") to 100 ("very satisfied"). The respondents did not want autocratic managerial decision making (mean +/- SD autocratic sub-scale score 4.29 +/- 0.60). Autocratic decision making was associated with lower job satisfaction (r = 0.49), whereas consultative and delegative decision making were associated with higher job satisfaction (r = -0.31 and -0.48, respectively). RTs who worked in departments that had < 25 RT employees reported higher job satisfaction than did RTs in larger departments (p = 0.029). CONCLUSIONS: Our survey data indicate that (1) RTs prefer delegative and consultative managerial decision making, (2) job satisfaction was highest in departments that had < 25 RTs in the department and in which the manager practiced participative decision making. These findings offer guidance for organizing optimal work environments for RTs. PMID- 15271232 TI - Methemoglobinemia: sudden dyspnea and oxyhemoglobin desaturation after esophagoduodenoscopy. PMID- 15271233 TI - Consciousness and neurosurgery. AB - The neuronal basis of consciousness is the greatest challenge to the scientific worldview. Much relevant empirical work is carried out on the minimal neuronal mechanisms underlying any one specific conscious percept. Two broad approaches are popular among brain scientists: electrophysiological recordings from individual neurons in the cortex of behaving monkeys or behavior combined with functional brain imaging in humans. However, many aspects of consciousness are problematic or remain off-limits to the former approach, while the latter one lacks sufficient spatial and temporal resolution to monitor individual neurons that are key to perception, thought, memory, and action. It is here that neurosurgeons, probing the living human brain on a daily basis, can play a decisive role. This article explores the contributions of neurosurgeons to this quest and outlines some of the results that have already been achieved. PMID- 15271234 TI - Genetic and cellular therapies for cerebral infarction. AB - Neurosurgeons, working as surgical scientists, can have a prominent role in developing and implementing genetic and cellular therapies for cerebral ischemia. The rapid emergence of both genetic and cellular therapies for neural regeneration warrants a careful analysis before implementation of human studies to understand the pitfalls and promises of this strategy. In this article, we review the topic of genetic and cellular therapy for stroke to provide a foundation for practicing neurosurgeons and clinical scientists who may become involved in this type of work. In Part 1, we review preclinical approaches with gene transfer, such as 1) improved energy delivery, 2) reduction of intracellular calcium availability, 3) abrogation of effects of reactive oxygen species, 4) reduction of proinflammatory cytokine signaling, 5) inhibition of apoptosis mediators, and 6) restorative gene therapy, that are paving the way to develop new strategies to treat cerebral infarction. In Part 2, we discuss the results of studies that address the possibility of using cellular therapies for stroke in animal models and in human trials by reviewing 1) the basics of stem cell biology, 2) exogenous and 3) and endogenous cell sources for therapy, and 4) clinical considerations in cell therapy applications. These emerging technologies based on the advancements made in recent years in the fields of genetics, therapeutic cloning, neuroscience, stem cell biology, and gene therapy provide significant potential for new therapies for stroke. PMID- 15271235 TI - Bleeding after radiosurgery for cerebral arteriovenous malformations. AB - OBJECTIVE: Obliteration is progressive after radiosurgery (RS) for cerebral arteriovenous malformation (AVM), and until it is complete, there is still a risk of hemorrhage. The aim of our study was to evaluate the severity of hemorrhage after RS, the actuarial risk of hemorrhage, and the parameters associated with hemorrhage. METHODS: Of 756 patients treated by linear accelerator RS for AVM, 51 (6.5%) had one or more hemorrhages after the RS. We studied the clinical, anatomic, and dosimetric parameters and obliteration rate before hemorrhage and then calculated the actuarial risk per patient and per hemorrhage before and after RS. Correlations between parameters and risk were studied by univariate and multivariate analysis using Kaplan-Meier hemorrhage-free survival curves and the Cox model. RESULTS: Apart from one exclusively ventricular hemorrhage, which caused the death of the patient, only parenchymal hemorrhages were associated with morbidity and neurological deficits (64.5% of all cases of hemorrhage had neurological deficits, 45% had a permanent deficit). The overall mortality rate per hemorrhage was 7.14%. The overall morbidity rate was 47.6%, 26.2% with a permanent deficit. In all but one patient, the AVM was not cured before hemorrhage; thus, the mean obliteration rate before hemorrhage was 24%. The actuarial hemorrhage rates were 3.08% per year per patient and 3.31% per year per hemorrhage. The actuarial rate per patient increased from 1.66% the 1st year to 3.87% in the 5th year after RS but was not statistically different from the rate before radiosurgery. The parameters found to be correlated with hemorrhage risk after RS using multivariate analysis were intranidal or paranidal aneurysms, complete coverage, and minimum dose. CONCLUSION: The risk of hemorrhage after RS would seem to be the sum of hemorrhage risk factors of the AVM and factors predicting a poor level of obliteration. These factors can be predicted in some cases but rarely avoided. PMID- 15271236 TI - Comparison of endovascular and surface cooling during unruptured cerebral aneurysm repair. AB - OBJECTIVE: To compare endovascular versus surface methods for the induction and reversal of hypothermia during neurosurgery in a multicenter, prospective, randomized study. METHODS: Patients undergoing elective open craniotomy for repair of an unruptured cerebral aneurysm (n = 153) were randomly assigned (2:1) to undergo whole-body hypothermia to 33 degrees C, either with an endovascular cooling device placed in the inferior vena cava via the femoral vein (n = 92) or with a surface convective air blanket (n = 61). Active rewarming was accomplished using the same devices. RESULTS: Cooling rates in endovascular and surface blanket groups averaged 4.77 and 0.87 degrees C/h, respectively (P < 0.001). When the first temporary arterial or aneurysm clip was placed, 99% of endovascular patients and 20% of surface blanket patients had reached the target of 33 degrees C (P < 0.001). Obese patients were cooled efficiently with the endovascular approach (3.56 degrees C/h). Rewarming rates averaged 1.88 degrees C/h for endovascular patients and 0.69 degrees C/h for surface blanket patients (P < 0.001). By the end of surgery, 89 and 53% of these patients, respectively, had rewarmed to at least 35 degrees C (P < 0.001). On leaving the operating room, 14% of endovascular patients and 28% of surface blanket patients were still intubated (P = 0.035). The overall safety of the two procedures was comparable. No clinically significant catheter-related thrombotic, bleeding, or infectious complications were reported in the endovascular group. CONCLUSION: Endovascular cooling provided superior induction, maintenance, and reversal of hypothermia compared with the surface blanket, without an increase in complications. Endovascular cooling may have clinical benefit for patients undergoing cerebrovascular surgery, as well as patients with acute stroke, head injury, or acute myocardial infarction. PMID- 15271237 TI - Giant vertebrobasilar aneurysms: endovascular treatment and long-term follow-up. AB - OBJECTIVE: To report long-term imaging follow-up and clinical outcome of 13 patients with a giant vertebrobasilar aneurysm treated by parent artery occlusion (PAO). METHODS: From 1994 to 2000, 13 consecutive patients with a giant vertebrobasilar aneurysm were treated by PAO. Symptoms were related to mass effect in nine patients and to a subarachnoid hemorrhage in four. Endovascular treatment consisted of aneurysm trapping in nine patients and occlusion of one or both vertebral arteries in four. We assessed the clinical outcome and imaging findings in all patients during a 28-month period. RESULTS: Endovascular treatment resulted in clinical improvements in eight patients, worsening of symptoms in four, and death in one. One woman with a ruptured vertebral aneurysm died from a rebleeding after PAO without trapping. One man developed a brainstem infarction after lower basilar artery occlusion and incurred hemiparesis. In three patients, symptoms of mass effect increased after the procedure. Long-term follow-up revealed good or excellent clinical outcome in all patients and a sharp decrease in size of the thrombosed aneurysm in nine patients. One basilar aneurysm recanalized despite selective coiling and subsequent bilateral vertebral artery occlusion; one vertebral aneurysm and one basilar aneurysm did not decrease in size despite complete occlusion. CONCLUSION: Giant vertebrobasilar aneurysms are rare and challenging lesions for both neurosurgeons and neurointerventionalists. Their treatment by endovascular PAO remains safe and effective. Early clinical worsening may be observed, but long-term follow-up shows good or excellent results in most patients. This treatment can be carried out with minimal morbidity and mortality using clinical and angiographic monitoring. PMID- 15271238 TI - Predicting stroke risk in pediatric moyamoya disease with xenon-enhanced computed tomography. AB - OBJECTIVE: To determine whether estimates of regional cerebral blood flow (rCBF) using xenon computed tomography (XeCT) in children with moyamoya disease can predict stroke risk before and after treatment. METHOD: Seven patients with moyamoya disease underwent 22 serial Xe computed tomographic scans. Estimates of rCBF were obtained at three computed tomographic levels by use of a 5-minute inhalation of 28% Xe. Acetazolamide challenge was performed in eight scans. For comparison of abnormal vessel distribution and areas of infarction, 17 intra arterial digital subtraction angiograms, 47 computed tomographic scans, and 15 magnetic resonance imaging scans were available. Follow-up exceeded 36 months in all patients. Mean follow-up for the interventional group was 65.2 months (n = 5; range, 37-109 mo) and 38 months for the nonoperative patients (n = 2; 36 and 40 mo). RESULTS: Of six Xe computed tomographic scans obtained at diagnosis, four revealed regions of oligemia, augmented vertebrobasilar flow, and regions of carotid steal after acetazolamide. In the delay between diagnosis and treatment, three patients had strokes in ischemic areas identified by XeCT. Of the 10 posttreatment scans obtained from 4 patients, 2 revealed improved tissue perfusion with angiography confirming successful encephaloduroangiomyosynangiosis. In 2 others, XeCT performed 6 months posttreatment revealed improved perfusion without angiographic change, and angiography at 1 year revealed failed encephaloduroangiomyosynangiosis and new native collaterals. None of the patients with improved rCBF had new strokes. Eleven of 14 Xe computed tomographic scans were obtained within 30 days of angiography. Comparison of these studies demonstrates that regions of oligemia were confined to areas associated with vessel stenosis and little neovascularity or collateral pathways. CONCLUSION: XeCT, particularly with acetazolamide challenge, objectively quantifies rCBF. Our preliminary data suggest that it may permit assessment of stroke risk in children with moyamoya disease and may predict surgical outcome earlier than angiography. PMID- 15271239 TI - Trigeminal neuralgia caused by venous compression. AB - OBJECTIVE: The purpose of this study is to clarify whether venous compression on the trigeminal nerve really causes trigeminal neuralgia or not, and to identify which veins are the offending veins. METHODS: We used microvascular decompression in operations on 121 patients with typical trigeminal neuralgia. We analyzed the intraoperative findings and surgical results in these 121 cases. RESULTS: In 7 of the 121 cases, only the vein was identified as a compressive factor on the trigeminal nerve. In 6 of these 7 cases, single venous compression was found, whereas the remaining case had two offending veins. The transverse pontine vein was most frequently found as the offending vein near Meckel's cave. All patients showed complete relief of trigeminal pain after decompression of the veins, but four of them developed facial numbness after surgery, which tended to be slight and did not require any treatment. CONCLUSION: Our surgical experiences showed that venous compression could cause trigeminal neuralgia by itself and that the transverse pontine vein should be carefully observed because it is most frequently the offending vein. PMID- 15271240 TI - Evidence for a clinically distinct new subtype of grade II astrocytomas in patients with long-term epilepsy. AB - OBJECTIVE: The authors tested the hypothesis that among Grade II astrocytomas with a particularly long seizure history, a subgroup is hidden with a different prognosis and possibly histological characteristics. To do so, clinical and histological characteristics of two groups of World Health Organization Grade II astrocytoma patients were analyzed: the long-term epilepsy-associated tumor (LEAT) astrocytoma group, with a mean duration of 12.5 years of seizures (n = 19), and the ordinary astrocytomas (n = 87), with a mean length of seizure history of 1.5 years (Non-LEAT). METHODS: All astrocytomas operated on between 1988 and 1999 were collected and followed up for 2 to 13 years (median, 7.0 yr). The 19 LEAT astrocytomas belonged to a group of 207 long-term epilepsy-associated tumors from the epilepsy surgery program. The 87 Non-LEAT cases were 60 so-called ordinary or diffuse World Health Organization Grade II astrocytomas and 27 oligoastrocytomas without long-term epilepsy operated on during the same time period. All tumor cases have been reviewed and partly reclassified as a result of the use of modern immunohistochemical techniques. Statistical analyses for possible discriminating factors included chi(2) test, Fisher's exact test, Kaplan Meier curves, and multifactorial analysis. RESULTS: Histological subtyping revealed a possible new isomorphic astrocytoma subtype in seven patients. By use of Kaplan-Meier curves, this isomorphic subtype had 50% fewer recurrences at 7.5 years and an estimated long-term survival of 80%. LEAT astrocytomas differed from ordinary Non-LEAT astrocytomas in overall length of history, younger age at first seizure, and higher percentage of 10-year survivors (80%). Temporal location did not influence outcome, and the presence of epilepsy per se was also not a prognostic factor. CONCLUSION: Differences between astrocytomas with a very long seizure history and those with a very short seizure history do exist. Significant factors for prognosis were age at surgery and presence of postoperative tumor residue but not the presence of epilepsy per se. A new subtype of astrocytomas, provisionally called isomorphic LEA astrocytoma, has putatively been identified with significantly better survival and lower recurrence rate. The negative prognostic factor of a gemistocytic differentiation pattern in diffuse astrocytomas was confirmed. PMID- 15271241 TI - Diagnosis and management of pineocytomas. AB - OBJECTIVE: Pineocytomas are associated with the most favorable prognosis of all pineal tumors. However, a subset of pineocytomas may have a predilection for recurrence and therefore behave aggressively. PATIENTS AND METHODS: Records of nine patients (five men, four women; mean age, 44 yr; range, 24-63 yr) with histologically diagnosed pineocytomas consecutively treated between 1990 and 2003 were reviewed retrospectively to identify factors predictive of aggressiveness. Eight patients presented with hydrocephalus and four with tectal compression. Three patients underwent gross total resection, and six underwent subtotal resection or biopsy. RESULTS: Three local recurrences necessitated reoperation. One recurrence involved the obex of the fourth ventricle. The mean time to recurrence was 3.5 years (range, 1-7 yr). There was no correlation between histological features and tumor recurrence. Patients undergoing radiosurgery showed stable or attenuated local disease (mean follow-up, 19.3 mo; range, 6-36 mo). Mean radiographic follow-up was 34 months (range, 6 mo to 10 yr). Mean clinical follow-up was 36 months (range, 1 mo to 10 yr). CONCLUSION: A subset of pineocytomas demonstrates the potential for symptomatic recurrence. We advocate an attempt at gross total tumor resection for all symptomatic patients with tectal plate compression, reserving radiosurgery for small, subtotally resected, or recurrent lesions. Patients must be followed closely for recurrence. Radiosurgery seems to be beneficial for local tumor control. Further investigation is needed to identify histological markers for pineocytomas that behave aggressively. PMID- 15271242 TI - Volumetric assessment of glioma removal by intraoperative high-field magnetic resonance imaging. AB - OBJECTIVE: To investigate the contribution of high-field intraoperative magnetic resonance imaging (iMRI) for further reduction of tumor volume in glioma surgery. METHODS: From April 2002 to June 2003, 182 neurosurgical procedures were performed with a 1.5-T magnetic resonance system. Among patients who underwent these procedures, 47 patients with gliomas (14 with World Health Organization Grade I or II glioma, and 33 with World Health Organization Grade III or IV glioma) who underwent craniotomy were investigated retrospectively. Completeness of tumor resection and volumetric analysis were assessed with intraoperative imaging data. RESULTS: Surgical procedures were influenced by iMRI in 36.2% of operations, and surgery was continued to remove residual tumor. Additional further resection significantly reduced the percentage of final tumor volume compared with first iMRI scan (6.9% +/- 10.3% versus 21.4% +/- 13.8%; P < 0.001). Percentages of final tumor volume also were significantly reduced in both low grade (10.3% +/- 11.5% versus 25.8% +/- 16.3%; P < 0.05) and high-grade gliomas (5.4% +/- 9.9% versus 19.5% +/- 13.0%; P < 0.001). Complete resection was achieved finally in 36.2% of all patients (low-grade, 57.1%; high-grade, 27.3%). Among the 17 patients in whom complete tumor resection was achieved, 7 complete resections (41.2%) were attributable to further tumor removal after iMRI. We did not encounter unexpected events attributable to high-field iMRI, and standard neurosurgical equipment could be used safely. CONCLUSION: Despite extended resections, introduction of high-field iMRI in conjunction with functional navigation did not translate into an increased risk of postoperative deficits. The use of high-field iMRI increased radicality in glioma surgery without additional morbidity. PMID- 15271243 TI - Endoscopic aqueductoplasty and interventriculostomy for the treatment of isolated fourth ventricle in children. AB - OBJECTIVE: There are different approaches for the treatment of isolated fourth ventricle in children, including a suboccipital ventriculoperitoneal shunt, suboccipital craniotomy with microsurgical fenestration, and endoscopic fenestration. We discuss the indications, surgical methods, and outcome of 18 patients who underwent endoscopic treatment for isolated fourth ventricle. METHODS: We retrospectively reviewed the medical histories of 18 patients with an isolated fourth ventricle. Surgical procedures included endoscopic aqueductoplasty, endoscopic aqueductoplasty with a stent, endoscopic interventriculostomy (lateral ventricle or third ventricle to fourth ventricle), and endoscopic interventriculostomy with a stent. Operations were performed between July 1997 and June 2002. The mean age of the patients at the time of surgery was 3 years. The mean follow-up was 29 months. All patients had a supratentorial ventriculoperitoneal shunt. RESULTS: Clinical symptoms (impairment of consciousness, tetraparesis, and ataxia) improved in all patients. Reduction of the size of the fourth ventricle was observed in all patients. Seven patients required reoperation because of restenosis (39% revision rate). Restenosis occurred between 2 weeks and 7 months after surgery (average, 3 mo). Four patients underwent reoperation with stent placement, and three patients underwent reaqueductoplasty. We had the following complications: one infection, one asymptomatic subdural hygroma, one transient oculomotor paresis, and one permanent oculomotor paresis (4 [22%] of 18 patients). CONCLUSION: The significant failure rate of fourth ventricle shunts has led to the development of alternative treatment methods. Endoscopic aqueductoplasty or interventriculostomy presents an effective, minimally invasive, and safe procedure for the treatment of isolated fourth ventricle in pediatric patients. Compared with suboccipital craniotomy and microsurgical fenestration, endoscopic aqueductoplasty is less invasive, and compared with fourth ventricle shunts, it is more reliable and effective. PMID- 15271244 TI - Posterior cranial fossa volume in patients with rickets: insights into the increased occurrence of Chiari I malformation in metabolic bone disease. AB - OBJECTIVE: Some have proposed that the calvarial thickening seen in patients with rickets results in an increased rate of Chiari I malformation (CIM) in these patients. The present study measures the posterior fossa volume in children with rickets to verify previous case reports indicting a small posterior fossa as the cause for an increased rate of CIM in children with rickets. METHODS: Patients were chosen by use of a computer database to search for individuals diagnosed with rickets. Nineteen patients were identified with this diagnosis. Seven patients were found from this cohort to have imaging of the head. Axial computed tomographic and magnetic resonance images were analyzed by use of the Cavalieri method to define posterior fossa volumes. These data were then compared with those from age-matched control subjects. RESULTS: Mean volumes of the posterior fossa were significantly reduced in all patients compared with age-matched control subjects (P < 0.0001). CONCLUSION: We have found that the volume of the posterior fossa is significantly smaller in children with rickets versus age matched control subjects. Furthermore, 29% of our study group had an associated CIM. We may hope that these data will aid in the further understanding of the pathophysiology of CIM in cases of metabolic bone disease. PMID- 15271245 TI - The unique characteristics of "upper" lumbar disc herniations. AB - OBJECTIVE: To compare the characteristics, presentation, and surgical outcome of patients with microdiscectomies at L1-L2 and L2-L3 with those we treated at L3 L4. We further sought to compare these results with those reported in the literature for discectomies at the L4-L5 and L5-S1 levels. METHODS: We reviewed the clinical data collected from 69 patients who had 72 L1-L2, L2-L3, and L3-L4 microdiscectomies performed from 1989 to 1999 at the New York University Medical Center. Patients who had surgery at L1-L2 or L2-L3 were grouped and compared with those treated at the L3-L4 level. Patients' charts were retrospectively reviewed at a mean of 12.9 months after surgery for presenting signs and symptoms, patient characteristics, and surgical outcome. Long-term follow-up via telephone interview was obtained at an average of 81.3 months after surgery. RESULTS: In the L1-L2 + L2-L3 group, 58% of the patients had previous lumbar disc surgery, compared with only 10% of those in the L3-L4 group, and 20% in the L1-L2 + L2-L3 group required a fusion during the procedure compared with only 10% in the L3-L4 group. These differences are both statistically significant. The short-term chart review demonstrates that only 58% and 53% of patients in the L1-L2 + L2-L3 group were improved with regard to radicular and back pain, respectively, whereas those in the L3-L4 group reported 94 and 87% rates of improvement in the same categories, both highly statistically significant findings. The long-term follow up confirmed a highly statistically significantly worse outcome in the L1-L2 + L2 L3 group, with only 33% of patients reporting an improvement in their economic or functional status, compared with an 88% rate of improvement in the L3-L4 group. The outcome of our patients with L3-L4 herniations was similar to that reported in the literature for herniations at L4-L5 and L5-S1. CONCLUSION: Herniated discs at the L1-L2 or L2-L3 level are different entities from those at lower levels of the lumbar spine. The surgical outcome in terms of postoperative back and radicular pain is worse for herniated discs at L1-L2 and L2-L3 compared with those treated at L3-L4. Our patients with L1-L2 or L2-L3 surgically treated herniated discs were more likely to have had previous lumbar surgery and required a fusion more often than their counterparts with L3-L4 herniated discs. PMID- 15271246 TI - Correlation between withdrawal symptoms and medication pump residual volume in patients with implantable SynchroMed pumps. AB - OBJECTIVE: To investigate whether patients with implantable SynchroMed pumps (Medtronic, Inc., Minneapolis, MN) develop symptoms of drug withdrawal at residual medication volumes that exceed 2 ml (the alarm residual volume recommended by the manufacturer). METHODS: The data sheets of 88 patients with implantable SynchroMed pumps were retrospectively reviewed. The following parameters were analyzed: development of symptoms of drug withdrawal; drugs used in the pump; disease state; drug residual volume in the pump; intake of orally administered medications; time of development of withdrawal symptoms; drug flow volume through the pump; daily intrathecally administered drug dose; and drug concentration in the pump. RESULTS: Of 88 patients, 21 (24%) consistently developed symptoms of drug withdrawal 1 to 7 days before the drug residual volume reached a mean of 2.7 ml (range, 2.1-3.8 ml; median, 2.6 ml). Symptoms first developed 1 to 18 months after surgery. In all patients, symptoms of drug withdrawal subsided after pump refill and did not recur after the alarm volume was increased to 4 ml. Symptom development did not correlate with intake of orally administered medication, drug flow volume through the pump, intrathecally administered drug dose, drug concentration in the pump, drugs used in the pump, or disease state. CONCLUSION: Some patients develop symptoms of drug withdrawal at residual volumes that exceed 2 ml. We could not identify factors that predict this occurrence. Withdrawal symptoms did not recur when the alarm volume was increased to 4 ml. PMID- 15271247 TI - Superficial temporal artery-to-middle cerebral artery bypass grafting for cerebral revascularization. AB - Superficial temporal artery-to-middle cerebral artery bypass procedures are an important tool in the armamentarium of cerebrovascular surgeons for the treatment of carotid occlusion and revascularization for complex aneurysms and brain tumors. This article enumerates the essential steps in performing superficial temporal artery-to-middle cerebral artery bypass procedures. The nuances of this technique reflect the extensive experience of the senior authors. PMID- 15271248 TI - The accessory atlantoaxial ligament. AB - OBJECTIVE: The stability of the joints connecting the cranium to the upper cervical spine is of vital importance. The ligaments of this region, for the most part, have been thoroughly investigated, with the exception of the accessory atlantoaxial ligament. METHODS: Ten cadaveric specimens were examined to observe the anatomy of this ligament. RESULTS: This ligament was found in all specimens, and in each, it not only connected the atlas to the axis but also continued cephalically to the occipital bone. The approximate dimensions of this structure were 3 cm x 5 mm. Functionally, this ligament became maximally taut with a rotation of the head of 5 to 8 degrees. Laxity was observed with cervical extension, and maximal tautness was seen at 5 to 10 degrees of cervical flexion. CONCLUSION: The accessory atlantoaxial ligament seems to participate in craniocervical stability and perhaps should be renamed the accessory alar ligament or accessory atlantoaxialoccipital ligament; both of these terms better denote its anatomic characteristics. Perhaps in the future, better magnetic resonance imaging techniques and machines will be able to identify this structure so as to appreciate its integrity after upper cervical spine trauma. PMID- 15271249 TI - Functional improvement with low-dose dopaminergic grafts in hemiparkinsonian rats. AB - OBJECTIVE: The beneficial functional effects of neural transplantation in Parkinson's disease are often directly attributed to the number of surviving dopaminergic cells within a graft. However, recent clinical trials of fetal neural transplantation suggest that a high number of dopaminergic cells may induce serious side effects. In this study, we explored the ability of low-dose dopaminergic grafts to produce functional benefits in the 6-hydroxydopamine rodent model of Parkinson's disease over a long period of observation. METHODS: Twelve rats received either 50,000 or 400,000 fetal ventral mesencephalic cells implanted into the striatum. Rotational behavior was assessed after the lesion and at 3, 6, 9, and 12 weeks after transplantation. Twelve weeks after transplantation, animals were perfused, and microtome sections were stained for tyrosine hydroxylase, glial fibrillary acidic protein, heat-shock protein 27, and vimentin. RESULTS: The low-dose group had a three-fold increase in tyrosine hydroxylase-positive cell survival rate compared with the high-dose group rate. The low-dose group also had a mean cell diameter significantly higher than the high-dose group. There was no significant difference between groups in fiber density; however, a higher percentage of longer fibers was encountered in the low dose group. The low-dose group had a lower degree of trauma in the striatum, as assessed by optical density scores from glial fibrillary acidic protein, heat shock protein 27, and vimentin staining. There was significant improvement in rotational behavior in the high-dose group at 3 weeks after transplantation, whereas the rotational behavior normalized in the low-dose group at 6 weeks after grafting. There was no significant difference in rotational behavior scores between groups at 6 weeks after grafting. CONCLUSION: This study demonstrates that over time, a low-dose dopaminergic graft has the capability of eliciting the same functional effect as a high-dose graft. Furthermore, low-dose grafts may increase graft survival, fiber outgrowth, and dopamine production and decrease trauma to the brain. PMID- 15271250 TI - Tumor necrosis factor alpha expression and protein levels after fluid percussion injury in rats: the effect of injury severity and brain temperature. AB - OBJECTIVE: Tumor necrosis factor alpha (TNFalpha) is elevated in some models of traumatic brain injury (TBI). However, it is unclear how TNFalpha messenger ribonucleic acid (mRNA) expression and protein levels are affected by injury severity and posttraumatic temperature modification. This study determined the regional and temporal profile of TNFalpha levels after moderate and severe TBI and assessed the effects of posttraumatic hypothermia or hyperthermia on this proinflammatory cytokine. METHODS: Adult male Sprague-Dawley rats were subjected to sham procedures (no injury), moderate fluid-percussion TBI (1.8-2.2 atm), or severe fluid-percussion TBI (2.4-2.6 atm). After 1 to 72 hours of survival, animals were killed, and brain samples, cerebrospinal fluid, and serum were harvested for enzyme-linked immunosorbent assay quantification of TNFalpha levels. In a subsequent study, a 3-hour period of posttraumatic hypothermia (33 degrees C) or hyperthermia (39.5 degrees C) was applied, followed by immediate killing and cytokine assay. Another group was subjected to moderate TBI (1.8-2.2 atm), followed by killing at 15 minutes or at 1, 3, or 24 hours for TNFalpha reverse transcriptase-polymerase chain reaction analysis. RESULTS: A significant increase in TNFalpha mRNA and protein levels in cellular lysates of injured cortex and ipsilateral hippocampus was noted by 1 hour after TBI; it was sustained to 3 hours, followed by a rapid decline. Increased injury severity was associated with increased protein levels at remote injury sites and in the injured cerebral cortex at 72 hours. Posttraumatic hypothermia significantly reduced TNFalpha mRNA expression in the hippocampus compared with that in normothermic rats. In contrast, no temperature effects on TNFalpha protein levels were documented. CONCLUSION: Rapid and marked increase in TNFalpha mRNA expression and protein levels follows moderate and severe TBI. Injury severity and posttraumatic temperature play a modest but significant role on TNFalpha expression and protein levels. These findings suggest that the effects of posttraumatic temperature on histopathological and behavioral outcome primarily may involve secondary mediators that do not operate directly through their effect on TNFalpha. PMID- 15271251 TI - PTK787/ZK222584, an inhibitor of vascular endothelial growth factor receptor tyrosine kinases, decreases glioma growth and vascularization. AB - OBJECTIVE: The aim of this study was to test the efficacy of PTK787/ZK222584, an inhibitor of vascular endothelial growth factor (VEGF) receptor tyrosine kinases, on VEGF-dependent glioma vascularization and growth. METHODS: C6 rat glioma cells were transfected with VEGF(164) in a sense (V(+)) or antisense (V(-)) direction. Spheroids generated from V(+) or V(-) cells were implanted orthotopically into 60 rat brains. Expression of VEGF and fetal liver kinase-1 (VEGF receptor 2) was assessed immunohistochemically. Animals with V(+) gliomas received orally administered PTK787/ZK222584 on postoperative Day (POD) 1 to 12 or POD 7 to 12. Untreated animals served as negative controls, and animals with V(-) gliomas served as positive controls. Growth and vascularization were evaluated by magnetic resonance imaging and immunohistochemistry. RESULTS: Flk-1 expression was positive within tumor vessels in V(+) gliomas, whereas all C6 clones were negative for fetal liver kinase-1 in vitro. Early (POD 1-12) and delayed (POD 7 12) application of PTK787/ZK222584 in V(+) glioma-bearing animals resulted in a significant reduction of tumor size (71% and 36%, P < 0.05) as measured by magnetic resonance imaging volumetry. Early treated V(+) gliomas reached similar volumes compared with V(-) gliomas. Vessel density was significantly reduced (42.3% and 25.7%, P < 0.05), and areas of intratumoral necrosis were enlarged (by 1.7-fold after early treatment). Additionally, proliferation was decreased by 89% and 72% (P < 0.05). There was no growth-inhibiting effect of PTK787/ZK222584 on V(-) cells observed. CONCLUSION: PTK787/ZK222584 significantly halted VEGF mediated glioma growth by inhibition of neovascularization and proliferation, providing a promising new tool in malignant glioma therapy. PMID- 15271255 TI - Intrinsic regenerative ability of mature CNS neurons. AB - A prevailing view in neuroscience is that the mature CNS has relatively little capacity to respond adaptively to injury. Recent data indicating a high degree of structural plasticity in the adult brain provides an impetus to reexamine how central neurons react to trauma. An analysis of both in vivo and in vitro experimental studies demonstrates that certain brain neurons may have an intrinsic ability to respond to structural injury by an attempt at regenerative sprouting. Indeed, aberrant sprouting following neuronal injury may be the cause of epilepsy following brain trauma and may underlie the neuronal changes stimulated by plaque formation in Alzheimer's disease. An understanding of the stereotypical reaction to injury of different CNS neurons, as well as the role of nonneuronal cells, may provide new avenues for therapeutic intervention for a range of neurodegenerative diseases and "acquired" forms of CNS injury. PMID- 15271256 TI - Electrical signaling in central orexin/hypocretin circuits: tuning arousal and appetite to fit the environment. AB - In mammals, alertness and foraging are linked to the light-dark cycle and body energy levels. This link is crucially dependent on the novel peptide transmitters orexins/hypocretins. The firing of orexin neurons encodes the overall internal (body energy levels) and external (time of day) environment. In turn, orexins modulate arousal and appetite by innervating and electrically exciting wakeand appetite-promoting neurons. Electrical signaling in orexin circuits thereby couples arousal to the environment and synchronizes foraging with states of high alertness. PMID- 15271257 TI - Inhibitory interneurons in the olfactory bulb: from development to function. AB - Identifying and defining the characteristic features of the inhibitory neurons in the nervous system has become essential for achieving a cellular understanding of complex brain activities. For this, the olfactory bulb is ideally suited because it is readily accessible, it is a laminated structure where local interneurons can be easily distinguished from projecting neurons, and, more important, GABAergic interneurons are continuously replaced. How the newly generated neurons integrate into a preexisting neural network and how basic network functions are maintained when a large percentage of neurons are subjected to continuous renewal are important questions that have recently received new insights. Here, it is seen that the production of bulbar interneurons is specifically adapted to experience-dependent regulation of adult neural networks. In particular, the authors report the degree of sensitivity of the bulbar neurogenesis to the activity level of sensory inputs and, in turn, how the adult neurogenesis adjusts the neural network functioning to optimize information processing. By maintaining a constitutive neurogenesis sensitive to environmental cues, this neuronal recruitment leads to improving sensory abilities. This review brings together recently described properties and emerging principles of interneuron functions that may convey, into bulbar neuronal networks, a degree of circuit adaptation unmatched by synaptic plasticity alone. PMID- 15271258 TI - Eph receptors, ephrins, and synaptic function. AB - Compelling new findings have revealed that receptor tyrosine kinases of the Eph family, along with their ephrin ligands, play an essential role in regulating the properties of developing mature excitatory synapses in the central nervous system. The cell surface localization of both the Eph receptors and the ephrins enables these proteins to signal bidirectionally at sites of cell-to-cell contact, such as synapses. Eph receptors and ephrins have indeed been implicated in multiple aspects of synaptic function, including clustering and modulating N methyl-D-aspartate receptors, modifying the geometry of postsynaptic terminals, and influencing long-term synaptic plasticity and memory. In this review, we discuss how Eph receptors and ephrins are integrated into the molecular machinery that supports synaptic function. PMID- 15271259 TI - Calcium dynamics and circadian rhythms in suprachiasmatic nucleus neurons. AB - The hypothalamic suprachiasmatic nucleus (SCN) has a pivotal role in the mammalian circadian clock. SCN neurons generate circadian rhythms in action potential firing frequencies and neurotransmitter release, and the core oscillation is thought to be driven by "clock gene" transcription-translation feedback loops. Cytosolic Ca2+ mobilization followed by stimulation of various receptors has been shown to reset the gene transcription cycles in SCN neurons, whereas contribution of steady-state cytosolic Ca2+ levels to the rhythm generation is unclear. Recently, circadian rhythms in cytosolic Ca2+ levels have been demonstrated in cultured SCN neurons. The circadian Ca2+ rhythms are driven by the release of Ca2+ from ryanodine-sensitive internal stores and resistant to the blockade of action potentials. These results raise the possibility that gene translation/transcription loops may interact with autonomous Ca2+ oscillations in the production of circadian rhythms in SCN neurons. PMID- 15271261 TI - Pathway-specific maturation, visual deprivation, and development of retinal pathway. AB - One of the fundamental features of the visual system is the segregation of neural circuits that process increments and decrements of luminance into ON and OFF pathways. In mature retina, the dendrites of retinal ganglion cells (RGCs) in the inner plexiform layer (IPL) of retina are separated into ON or OFF sublamina specific stratification. At an early developmental stage, however, the dendrites of most RGCs are ramified throughout the IPL. The maturation of RGC ON/OFF dendritic stratification requires neural activities mediated by afferent inputs from bipolar and amacrine cells. The synchronized spontaneous burst activities in early postnatal developing retina regulate RGC dendritic filopodial movements and the maintenance or elimination of dendritic processes. After eye opening, visual experience further remodels and consolidates the retinal neural circuit into mature forms. Several neurotransmitter systems, including glutamatergic, acetylcholinergic, GABAergic, and glycinergic systems, might act together to modulate the RGC dendritic refinement. In addition, both the bipolar cells and cholinergic amacrine cells may provide laminar cues for the maturation of RGC dendritic stratification. PMID- 15271262 TI - Regulation of arm and leg movement during human locomotion. AB - Walking can be a very automated process, and it is likely that central pattern generators (CPGs) play a role in the coordination of the limbs. Recent evidence suggests that both the arms and legs are regulated by CPGs and that sensory feedback also regulates the CPG activity and assists in mediating interlimb coordination. Although the strength of coupling between the legs is stronger than that between the arms, arm and leg movements are similarly regulated by CPG activity and sensory feedback (e.g., reflex control) during locomotion. PMID- 15271260 TI - Signaling cascades regulating NMDA receptor sensitivity to ethanol. AB - One of the major targets for ethanol (alcohol) in the brain is the N-methyl-D aspartate (NMDA) receptor, a glutamate-gated ion channel. Intriguingly, the effects of ethanol on the NMDA receptor are not homogeneous throughout the brain. This review focuses on recent studies revealing molecular mechanisms that mediate the actions of ethanol on the NMDA receptor in different brain regions via changes in NMDA receptor phosphorylation and compartmentalization. Specifically, the role of the scaffolding protein RACK1 and the regulatory protein DARPP-32 in mediating the distinct effects of ethanol is presented. PMID- 15271263 TI - Neuroanatomical markers for dyslexia: a review of dyslexia structural imaging studies. AB - A neuroanatomical description of dyslexia has been elusive, due in part to the complex cognitive nature of dyslexia. People with dyslexia have varying degrees of impairment in reading skills that engage oral and written language (reading) neural networks. Although findings for the inferior parietal lobule, inferior frontal gyrus, and cerebellum have been relatively consistent across studies, these studies also demonstrate that anatomical patterns of results vary according to the reading skills that characterize dyslexia. The number and likelihood of atypical anatomical findings in oral and/or written language systems appears to be related to the pattern of impairments in measures of phonology, orthography, and fluency. A comprehensive neurobiological understanding of dyslexia will depend on studies of dyslexic individuals with homogeneous perceptual, cognitive, and genetic backgrounds. PMID- 15271264 TI - Mapping changes in the human cortex throughout the span of life. AB - In this review, the authors summarize the literature on brain morphological changes that occur throughout the human life span from childhood into old age. They examine changes observed postmortem and in vivo where various brain MRI analytic methods have been applied. They evaluate brain changes observed with volumetric image analytic methods and voxel-based morphometric methods that may be used to better localize where changes occur. The primary focus of the review is on recent studies using state-of-the-art cortical pattern-matching techniques to assess age-related changes in cortical asymmetries, gray matter distribution, and brain growth across various age spans. The authors attempt to integrate findings from the in vivo studies with results from postmortem studies and analyze the complicated question of when brain maturation stops and brain aging begins. Analyzing the regional patterns of change initiated at various ages may help elucidate relationships between changing brain morphology and changing cognitive functions that occur throughout life. Long-range longitudinal studies, correlations between imaging and postmortem data, and more advanced image acquisition and analysis technologies will be needed to fully interpret brain morphological changes observed in vivo in relation to development and aging. PMID- 15271265 TI - Residency in the United States, subjective well-being, and depression in an older Mexican-origin sample. AB - OBJECTIVE: To compare the mental health and well-being of Mexican immigrants with native-born Mexican Americans living in the Lower Rio Grande Valley of Texas. METHODS: A randomly stratified sample of 353 Hispanics aged 45 and older were interviewed. The immigrant group (n = 148) was compared with native-born Mexican Americans (n = 205). RESULTS: The findings showed that the native-born group was not significantly different from the immigrant group on measures of depression, health status, life satisfaction, or self-esteem. The immigrant group was found to report significantly more stress than the Mexican American group. Income, age, gender, and acculturation were significant predictors of well-being, whereas immigration status and years of residency were not. DISCUSSION: The well-being of Mexican immigrants in the United States is confounded by such variables as income, age, gender, and acculturation, along with various other contextual factors that characterize their life experiences in the United States. PMID- 15271266 TI - Religious involvement and 6-year course of depressive symptoms in older Dutch citizens: results from the Longitudinal Aging Study Amsterdam. AB - OBJECTIVES: Expanding on cross-sectional studies, associations are examined between religious involvement and the 6-year course of depressive symptoms in older adults. METHODS: Subjects are 1,840 community-dwelling older adults (aged 55 to 85) participating in three measurement cycles of the Longitudinal Aging Study, Amsterdam. Assessments include aspects of religious involvement, depressive symptoms, physical health, self-perceptions, social integration, urbanization, and alcohol use. RESULTS: Church attendance is negatively associated with the course of depressive symptoms, also after adjustment for explanatory variables. Among respondents with functional limitations, lower depression scores are found for those who attend church on a regular basis. For respondents who are bereaved or nonmarried, however, slightly higher depression scores are found for those with high levels of orthodox beliefs. DISCUSSION: There is a consistent negative association over time between church attendance and depressive symptoms in older Dutch citizens. Both stress-buffering as well as depression-evoking effects of religious involvement are found. PMID- 15271267 TI - The life course of activity limitations: exploring indicators of functional limitations over time. AB - OBJECTIVE: To strengthen the foundations for the use of survey-based measures of functional limitations and to explore associations between limitations in a variety of activities across the adult life course. METHOD: Five panels of data from the young and mature women's cohorts of the National Longitudinal Surveys are used to (a) examine patterns of limitations in activities as women age, (b) compare how limitations develop over the life course, (c) explore how limitations in one activity are associated with limitations in others, and (d) investigate whether limitations develop incrementally or occur in clusters. RESULTS: We find that scales of functional limitations are not dependent on the age of the respondent, activity limitations emerge in clusters, and relationships between items do not consistently fall into upper and lower body groups. DISCUSSION: Scales of functional limitations are equally applicable to younger and older women, but further research is needed to compare substantive results using different methods of scale construction. PMID- 15271268 TI - The effects of social networks on disability in older Australians. AB - OBJECTIVE: To investigate the effects of total social networks and specific social networks with children, relatives, friends, and confidants on disability in mobility and Nagi functional tasks. METHODS: Six waves of data from the Australian Longitudinal Study of Ageing were used. Data came from 1,477 participants aged 70 years or older. The effects of total social networks and those with children, relatives, friends, and confidants on transitions in disability status were analyzed using binary and multinomial logistic regression. RESULTS: After controlling for a range of health, environmental, and personal factors, social networks with relatives were protective against developing mobility disability (OR = 0.89; 95% CI = 0.79 to 1.00) and Nagi disability (OR = 0.85; 95% CI = 0.74 to 0.96). Other social subnetworks did not have a consistent effect on the development of disability. DISCUSSION: The effects of social relationships extend beyond disability in activities of daily living. Networks with relatives protect against disability in mobility and Nagi tasks. PMID- 15271269 TI - Associations between anticipated support, physical functioning, and education level among a nationally representative sample of older adults. AB - OBJECTIVE: The purpose of this study is to determine the extent to which levels of anticipated support reported by older adults are associated with functional disability. In addition, level of education is tested as a modifier of this relationship. METHOD: Survey data were collected from a nationwide sample of 1,103 adults aged 60 to 95. Ordinary least squares regression analysis was used to test a range of model specifications involving anticipated support and functional disability. RESULTS: Levels of anticipated support are inversely associated with functional disability. Risk for functional disability is confined mostly to those reporting lower than average levels of anticipated support. This association is particularly strong with respect to instrumental support and among those older adults with low levels of education. DISCUSSION: These findings expand our understanding of the benefits of social relationships by suggesting that the perception of access to social support may enhance physical functioning in older age. PMID- 15271270 TI - A short-term intervention to enhance cognitive and affective functioning in older adults. AB - OBJECTIVE: This study was designed to investigate the benefits of a short-term intervention for older adults that targeted cognitive functioning and quality of life issues important for independent living. METHOD: One hundred twenty-four community-dwelling participants (aged 60 to 86) took part in one of three study conditions: theater arts (primary intervention), visual arts (non-content specific comparison group), and no-treatment controls. RESULTS: After 4 weeks of instruction, those given theater training made significantly greater gains than did no-treatment controls on both cognitive and psychological well-being measures. A comparison of theater and visual arts training showed fewer benefits in fewer areas for visual arts. DISCUSSION: The authors suggest reasons why various aspects of theater training appear to enhance healthy aging. PMID- 15271272 TI - A single mathematical model predicts physicians' recommendations and postmenopausal women's decisions to participate in a clinical trial to prevent breast cancer or coronary heart disease. AB - Few eligible postmenopausal women participate in clinical trial research to prevent breast cancer or coronary heart disease, making it impossible to adequately assess the efficacy of tested interventions for this vulnerable group. To elucidate the causal factors and decision model underlying participation behavior, 180 white, African American, and Hispanic postmenopausal women judged their likelihood of participation in a breast cancer or coronary heart disease prevention clinical trial in scenarios with varied cost/remuneration, perceived risk, doctor's recommendation, and expected toxicity. In addition, 293 white, African American, and Hispanic male and female physicians judged the strength of their participation recommendation in scenarios with varied cost/remuneration, expected toxicity, patient's age, and the source of the information about the clinical trial. An additive and constant-weight-averaging model were rejected. The same configural-weight-range model accounted for judgments in both breast cancer and coronary heart disease scenarios, with different parameter values for each group. According to this model, white and Hispanic women under 70 years of age are most likely to participate, even under somewhat adverse conditions; costs and high toxicity levels act as severe barriers to physicians' positive recommendations and women's participation. Perceived risk was the most important factor for women, yet only 8% and 15% reported ever having received risk information from their doctor for breast cancer and coronary heart disease, respectively. For these two diseases, respectively, 75% and 48% of women rated their risk of the disease as low and 76% and 88% reported they had never heard of a randomized clinical trial or of a prevention clinical trial being conducted. These results have implications for education, information dissemination, and prevention clinical-trial planners. PMID- 15271273 TI - Refining estimates of major depression incidence and episode duration in Canada using a Monte Carlo Markov model. AB - BACKGROUND: Serial period prevalence estimates for recurrent diseases such as major depression are available more frequently than fully detailed longitudinal data, but it is difficult to estimate incidence and episode duration from such data. Incidence and episode duration are critical decision modeling parameters for recurrent diseases. OBJECTIVES: To reduce bias that would otherwise occur in national incidence and duration-of-episode estimates for major depressive episodes deriving from studies using serial period prevalence data and to illustrate a methodological approach for the estimation of incidence from such studies. METHODS: Monte Carlo simulation was applied to a Markov process describing incidence and recovery from major depressive episodes. RESULTS: The annual incidence and episode duration were found to be 3.1% and 17.1 weeks, respectively. These estimates are expected to be less subject to bias than those generated without modeling. CONCLUSIONS: These results highlight the usefulness of Markov models for analysis of longitudinal data. The methods described here may be useful for decision modeling and may be generalizable to other chronic diseases. PMID- 15271274 TI - Decision aids for benign prostatic hyperplasia: applicability across race and education. AB - BACKGROUND/METHOD: Decision aids have not been widely tested in diverse audiences. The authors conducted interviews in a 2 x 2 race by education design with participants who were 50 years old (n = 188). The decision aid was a benign prostatic hyperplasia videotape. RESULTS: There was an increase in knowledge equal in all groups, with baseline knowledge higher in whites. The decision stage increased in all groups and was equivalent in the marginal-illiterate subgroup (n = 0.15). CONCLUSION: Contrary to expectations, results show no difference by race or college education in knowledge gain or increase in reported readiness to decide. The video appeared to produce change across race and education. The end decision stage was high, especially in less educated men. Results suggest that decision aids may be effective without tailoring, as suggested previously to enhance health communication in diverse audiences. Research should test findings in representative samples and in clinical encounters and identify types of knowledge absorbed from decision aids and whether the shift to decision reflects data/knowledge or shared decision-making message. PMID- 15271275 TI - Positive association between current health and health values for hypothetical disease states. AB - BACKGROUND: Valuations of hypothetical health state scenarios can be affected by participant characteristics. METHODS: The authors interviewed 108 veterans using the visual analogue scale (VAS), standard gamble (SG), time tradeoff (TTO), and willingness to pay (WTP) to measure health values for 1) current health and 2) 3 hypothetical health states portrayed in written scenarios describing cervical spondylotic myelopathy (CSM), a degenerative spine condition. They used bivariate rank order and multivariate regression analyses to assess the relationship between CSM values, participants' characteristics, and participants' current health values. RESULTS: Participants were predominantly male (89.8%) and Caucasian (75.9%), with a median age of 58.3 years and a median annual income of $15,000. Median values for current health were VAS, 0.75; SG, 0.80; TTO, 0.80; and WTP, $25,000. In the multivariate analysis, higher CSM values were associated with better current health as measured with the SG, TTO, and WTP (for all, P < 0.001); there was no association with VAS values (P = 0.157). CONCLUSIONS: Health values for CSM are positively associated with the current health of the study population. PMID- 15271276 TI - Women's preferences for doctor's involvement in decisions about mammography screening. AB - OBJECTIVES: To assess women's preferences for doctor's involvement in mammography screening decisions. METHODS: Mail survey of 50- to 69-year-old women residing in Geneva, Switzerland (N = 2216). RESULTS: Women considered that the decision to undergo mammography screening should be made by the doctor alone (5.6%), doctor primarily (42.6%), shared equally between woman and doctor (45.0%), woman primarily (4.2%), and woman alone (2.4%). These subgroups differed considerably. Compared to women in the shared equally group, doctor alone respondents were more likely to be older, to be born outside Switzerland, and to wish to know as late as possible about having cancer. In contrast, woman alone respondents were more likely to report no previous mammogram, to expect bad news from mammograms, and to feel nervous about screening. CONCLUSIONS: Most women wished to see their doctor involved in the decision to undergo a screening mammogram. Nevertheless, notable minorities held other opinions. PMID- 15271277 TI - Classification algorithms for hip fracture prediction based on recursive partitioning methods. AB - This article presents 2 modifications to the classification and regression tree. The authors improved the robustness of a split in the test sample approach and developed a cost-saving classification algorithm by selecting noninferior to the optimum splits from variables with lower cost or being used in parent splits. The new algorithm was illustrated by 43 predictive variables for 5-year hip fracture previously documented in the Study of Osteoporotic Fractures. The authors generated the robust optimum classification rule without consideration of classification variable costs and then generated an alternative cost-saving rule with equivalent diagnostic utility. A 6-fold cross-validation study proved that the cost-saving alternative classification is statistically noninferior to the optimal one. Their modified classification and regression tree algorithm can be useful in clinical applications. A dual X-ray absorptiometry hip scan and information from clinical examinations can identify subjects with elevated 5-year hip fracture risk without loss of efficiency to more costly and complicated algorithms. PMID- 15271278 TI - Exploration of a bayesian updating methodology to monitor the safety of interventional cardiovascular procedures. AB - Appropriate methods for monitoring of the safety of medical devices introduced into clinical practice have been elusive to develop and implement. A novel approach is the application of Bayesian updating, which incorporates existing knowledge regarding event rates into the estimation of risk. This framework has been shown in other domains to be data efficient and to address some of the limitations of conventional statistical methods. In this article, the authors propose a methodologic framework for developing initial prior probability distributions in risk-stratified patient groups and a mechanism for incorporating accumulating procedure safety experience. In addition, they use this methodology to retrospectively analyze the clinical outcomes of 309 patients undergoing an infrequent interventional cardiology procedure, rotational atherectomy. These exploratory analyses demonstrate the feasibility of Bayesian updating applied to medical device safety evaluation and indicate that the methodology is capable of generating stable estimates of risk in a variety of patient risk groups. PMID- 15271279 TI - Presidential reflections on the 25th anniversary of the society for medical decision making. PMID- 15271280 TI - Law and ethics. PMID- 15271281 TI - Two-layered quasi-3D finite element model of the oesophagus. AB - Analysis of oesophageal mechanoreceptor-dependent responses requires knowledge about the distribution of stresses and strains in the layers of the organ. A two layered and a one-layered quasi-3D finite element model of the rat oesophagus were used for simulation. An exponential pseudo-strain energy density function was used as the constitutive equation in each model. Stress and strain distributions at the distension pressures 0.25 and 1.0 kPa were studied. The stress and strain distributions depended on the wall geometry. In the one-layered model, the stress ranged from -0.24 to 0.38 kPa at a pressure of 0.25 kPa and from -0.67 to 2.57 kPa at a pressure of 1.0 kPa. The stress in the two-layered model at the pressure of 0.25 and 1.0 kPa varied from -0.52 to 0.64 kPa and from 1.38 to 3.84 kPa. In the two-layered model, the stress was discontinuous at the interface between the muscle layer and the mucosa-submucosa layer. The maximum stress jump was 1.67 kPa at the pressure of 1.0 kPa. The present study provides a numerical simulation tool for characterising the mechanical behaviour of a multi layered, complex geometry organ. PMID- 15271282 TI - Numerical study of nonlinear pulsatile flow in S-shaped curved arteries. AB - The nonlinear pulsatile blood flow in S-shaped curved arteries was studied with finite element method. Numerical simulations for flows in two models of S-shaped curved arteries with different diameters and under the same boundary conditions were performed. The temporal and spatial distributions of hemodynamic variables during the cardiac cycle such as velocity field, secondary flow, pressure, and wall shear stresses in the arteries were analyzed. Results of numerical simulations showed that the secondary flow in the larger S-shaped curved artery is more complex than that in the smaller one; stronger eddy flow occurred in the inner bends of curved arteries; pressure and wall shear stresses changed violently in the curved arteries, especially in the larger model. These hemodynamic variables in curved arteries will cause important effects on the function of arterial endothelium in the region. For instance, they may lead to the proliferation of smooth muscle cells and the thickening of the intima, and cardiovascular diseases such as atherosclerosis may develop in such regions. Due to having the special blood flow characteristics in the S-shaped arteries, it is worthwhile to study flow in this kind of curved artery. The comprehensive theoretical foundation showed in the present study can be extended to approach problems of nonlinear pulsatile flow in curved arteries with more complex geometrical shape. PMID- 15271283 TI - A low computational complexity algorithm for ECG signal compression. AB - In this work, a filter bank-based algorithm for electrocardiogram (ECG) signals compression is proposed. The new coder consists of three different stages. In the first one--the subband decomposition stage--we compare the performance of a nearly perfect reconstruction (N-PR) cosine-modulated filter bank with the wavelet packet (WP) technique. Both schemes use the same coding algorithm, thus permitting an effective comparison. The target of the comparison is the quality of the reconstructed signal, which must remain within predetermined accuracy limits. We employ the most widely used quality criterion for the compressed ECG: the percentage root-mean-square difference (PRD). It is complemented by means of the maximum amplitude error (MAX). The tests have been done for the 12 principal cardiac leads, and the amount of compression is evaluated by means of the mean number of bits per sample (MBPS) and the compression ratio (CR). The implementation cost for both the filter bank and the WP technique has also been studied. The results show that the N-PR cosine-modulated filter bank method outperforms the WP technique in both quality and efficiency. PMID- 15271284 TI - The effect of duty cycle and frequency on muscle torque production using kilohertz frequency range alternating current. AB - We investigated the frequency and duty cycle dependence of maximal electrically induced torque (MEIT) of the wrist extensors. Fifty hertz burst modulated sinusoidal alternating current (AC) in the frequency range 0.5-20 kHz was used, with duty cycles ranging from a minimum (one cycle) to maximum (continuous AC). MEITs were similar at low frequencies but decreased markedly above 2.5 kHz. MEITs also decreased markedly above a 20% duty cycle. Subjective reports of discomfort were fewest at 4 kHz and at duty cycles in the range 20-25%. Our conclusion is that for maximum torque production, a frequency of 1 kHz and a duty cycle of 20% are indicated. When comfort is a major consideration, a frequency of 2.5 kHz provides an acceptable trade-off between MEIT and comfort. The findings also suggest that low duty cycle, burst modulated AC stimulation may be more effective than stimulation using conventional low-frequency pulsed current. PMID- 15271285 TI - Factors affecting reaction times to short anterior postural disturbances. AB - One source of falls in the elderly may be an inability to sufficiently adjust to transient postural perturbations or slips. Identifying useful predictors of fall potential, as well as factors that affect the ability of an individual to detect a movement of the standing support surface may provide insight into postural stability and methods to increase stability in elders. The effects of acceleration, displacement, neurological status, and age on movement detection reaction times were studied in 25 individuals--1 young adults, seven neurologically intact elderly adults, and six elders with (diabetic) peripheral neuropathy. Acceleration detection thresholds for anterior perturbations of 1, 4, and 16 mm of the support surface was previously determined for each subject via a two-alternative forced choice (2AFC) protocol, with longer (16 mm) moves yielding lower 2AFC thresholds (12-39 mm/s(2)) that varied by group. Using the acceleration threshold value determined, and 125% of that threshold (suprathreshold), reaction times to the start of the platform movement were determined for all three displacements. Reaction times to an additional superthreshold movement (4 mm at 100 mm/s(2)) were also measured. Lower acceleration values over longer moves required longer reaction times for motion detection. Reaction times were also influenced by peak energy imparted to the subject through the move. The higher prevalence of falls in the elderly and elderly diabetic may be due to slowing reaction times compounded by larger amounts of imparted energy needed for detection of a slipping event. PMID- 15271286 TI - Changes in nuclear composition following cyclic compression of the intervertebral disc in an in vivo rat-tail model. AB - While in vitro studies have shown that mechanical loading can result in changes in the composition of intervertebral disc matrix, the effects of cyclic loading in vivo have not been considered. The objective of this study was to assess the effect of static and cyclic compression of different frequencies on the nuclear composition of the intervertebral disc. Thirty-six Sprague-Dawley rats were randomly divided into a control group (no pin insertion, no loading), a sham group (pins inserted in sixth and seventh caudal vertebrae, no loading), a static loading group (compression applied via pins) and cyclic loading groups (loading at 0.5, 1.5 or 2.5 Hz). Loading was applied for 1 h each day from the third to 17th day following pin insertion, and the caudal 5-6, 6-7 and 7-8 discs harvested to quantify proteoglycan content, collagen content and chondrocyte density in the nucleus pulposus. Static compression resulted in a significant reduction in total proteoglycan content as compared with the adjacent control disc, but this effect was not seen in any of the cyclic loading groups. However, comparison with the sham group appears to indicate an overall decrease in total proteoglycan content at the targeted and adjacent levels following cyclic loading. The 0.5 Hz loading group showed a significantly greater total proteoglycan content than all other compression groups, and also showed a lower total collagen content than the sham group. Results suggest that frequency dependent changes in composition occur in response to cyclic loading, but are not limited to the directly loaded disc alone. Further studies are required to verify this, but the choice of control appears to need careful consideration in all studies of this nature. PMID- 15271287 TI - Development of a cost-effective loading machine for biomechanical evaluation of mouse transgenic models. AB - Recent advances in molecular biology have enabled the widespread use of transgenic mouse models. Whether these transgenics present with a skeletal phenotype is often of interest, particularly to orthopedic researchers. Unfortunately, the expense of commercial mechanical testing systems often impedes their routine use in biology laboratories. In this article, the development of a small-scale, relatively inexpensive loading machine, that is ideal for the biomechanical bend testing of mouse long bones, is detailed and the system in transgenic mouse tibiae testing is utilized. PMID- 15271288 TI - The comparison of electromyographic pattern classifications with active and passive electrodes. AB - Electromyographic (EMG) signals are usually acquired using surface electrodes, and they commonly serve as the control sources of myoelectric prosthetic limbs. The use of passive electrodes and amplifiers with adjustable gain is very popular in laboratories for the development of new control strategies. However, active electrodes without conductive jelly are used in most clinical applications of myoelectric hand control. There remains an important question: Are there any differences between using active and passive electrodes in EMG pattern classifications? Autoregressive and cepstral coefficients were used to evaluate recognition rates via both types of electrodes. The results showed that the estimated recognition rates in the passive electrodes were comparable to those in the active ones (averaged recognition rate, 88.5 vs. 85.84% in the autoregressive coefficients, and 84.84 vs. 83.5%, in the cepstral coefficients, respectively). Aside from the lack of significant statistical differences between them, the results imply that the differences between the recognition rates via these electrodes could be negligible. This would be helpful for the myoelectric control of assistive devices. PMID- 15271289 TI - Joint kinematics and spatial-temporal parameters of gait measured by an ultrasound-based system. AB - Since measuring and recording techniques were developed, gait analysis has been frequently used in almost all fields of human locomotion such as rehabilitation medicine, orthopaedics, sports science, and other related fields. The measuring range of usual ultrasound-based devices is limited because the ultrasound sources must be always in visual contact with the microphones (markers). Our technique for the expansion of the measuring range is presented. Our approach is based on a mechanical axiom, which states that the position and orientation of a segment of the human body is determined by an array of three points per segment. The position of an invisible anatomical point of the segment could be determined by its position in relation to the fundamental points, being in visible contact on the body segment. Before measurement, the position of investigated anatomical points in relation to the fundamental points has to be given by an ultrasound based pointer. The position of fundamental points of each segment of the human body has to be measured during motion by the ultrasound-based device. A computer code calculates the position of anatomical points from the above data on-line. This approach provides an opportunity to investigate a discretional number and posture (lateral, medial, posterior and anterior) of anatomical points. Our model consists of 19 anatomical and anthropometrical points. Based on the spatial coordinates of the anatomical points investigated, the spatial-temporal parameters of gait and anatomical joint angles are estimated. No significant statistical difference was observed between the values presented and those found in literature. Several clinical applications can be proposed such as monitoring of rehabilitation progress after orthopaedic surgery and gait analysis in neurological diseases. PMID- 15271291 TI - Cell surface receptors in lysophospholipid signaling. AB - The lysophospholipids, lysophosphatidic acid (LPA) and sphingosine 1-phosphate (S1P), regulate various signaling pathways within cells by binding to multiple G protein-coupled receptors. Receptor-mediated LPA and S1P signaling induces diverse cellular responses including proliferation, adhesion, migration, morphogenesis, differentiation and survival. This review will focus on major components of lysophospholipid signaling: metabolism, identification and expression of LPA and S1P receptors, general signaling pathways and specific signaling mechanisms in mouse embryonic fibroblasts. Finally, in vivo effects of LP receptor gene deletion in mice will be discussed. PMID- 15271292 TI - Sphingosine 1-phosphate and lysophosphatidic acid receptors: agonist and antagonist binding and progress toward development of receptor-specific ligands. AB - Sphingosine 1-phosphate and lysophosphatidic acid are two phospholipid growth factors whose importance in physiology and pathophysiology is becoming more and more apparent. Structure-activity relationships for agonism and antagonism at the thirteen known cell-surface and one intracellular receptor are described. Particular emphasis is placed on ligands having different selectivity than the parent molecules. Structural insights regarding agonist and antagonist recognition by the receptors from both computational modeling studies and crystallography are also discussed. PMID- 15271293 TI - Mechanisms of lysophosphatidic acid production. AB - Lysophosphatidic acid is one of the most attractive phospholipid mediator with multiple biological functions and is implicated in various human diseases. In the past ten years much has been learned about the physiological roles of LPA through series of studies on LPA actions and its receptors. However, the molecular mechanisms of LPA have been poorly understood. LPA is produced in various conditions both in cells and in biological fluids, where multiple synthetic reactions occur. At least two pathways are postulated. In serum and plasma, LPA is mainly converted from lysophospholipids. By contrast, in platelets and some cancer cells, LPA is converted from phosphatidic acid. In each pathway, at least two phospholipase activities are required: phospholipase A1 (PLA1)/PLA2 plus lysophospholipase D (lysoPLD) activities are involved in the first pathway and phospholipase D (PLD) plus PLA1/PLA2 activities are involved in the second pathway. Now multiple phospholipases are identified that account for PLA1, PLA2, PLD, and lysoPLD activities. In the absence of specific inhibitors and genetically modified animals and individuals, the contribution of each phospholipase to LPA production can not be easily determined. However, apparently certain extracellular phospholipases such as secretory PLA2 (sPLA2-IIA), membrane associated PA-selective PLA1 (mPA-PLA1), lecithin-cholesterol acyltransferase (LCAT), and lysoPLD are involved in LPA production. PMID- 15271294 TI - Lysophosphatidic acid and sphingosine 1-phosphate biology: the role of lipid phosphate phosphatases. AB - The biological actions of the lysolipid agonists sphingosine 1-phosphate and lysophosphatidic acid, in addition to other bioactive lipid phosphates such as phosphatidic acid and ceramide 1-phosphate, can be influenced by a family of lipid phosphate phosphatases (LPP), including LPP1, LPP2, LPP3, the Drosophila homologues Wunen (Wun) and Wunen2 (Wun2) and sphingosine 1-phosphate phosphatases 1 and 2 (SPP1, SPP2). This review describes the characteristic of these enzymes and their potential physiological roles in regulating intracellular and extracellular actions and amounts of these lipids in addition to the involvement of these phosphatases in development. PMID- 15271295 TI - Biology of LPA in health and disease. AB - The functions of lysophosphatidic acid (LPA) can be broadly divided into two classes: (1) physiological and (2) pathological roles. The role of LPA in embryonic development can be seen as early as oocyte formation. It continues in postnatal homeostasis, through its ability to impart a level of protection from both stress and local injury, by regulating cellular proliferation, apoptosis, and the reorganization of cytoskeletal fibers. LPA may function as a double-edged sword. While it helps maintain homeostasis against stress and insult, it may also augment the development and spread of pathological processes, including cancers. PMID- 15271296 TI - Physiological and pathological actions of sphingosine 1-phosphate. AB - Sphingosine 1-phosphate (S1P), a product of sphingomyelin (SM) metabolism, occurs widely in nature. Although, originally described as an intracellular second messenger, its role as an extracellular lipid mediator in higher organisms has recently been shown with the discovery of the G protein-coupled receptors (GRCR) for S1P. In mammals, S1P receptors are widely expressed and are thought to regulate important physiological actions, such as immune cell trafficking, vascular development, vascular tone control, cardiac function, and vascular permeability, among others. In addition, S1P may participate in various pathological conditions. For example, S1P has been implicated as an important mediator in autoimmunity, transplant rejection, cancer, angiogenesis, vascular permeability, female infertility, and myocardial infarction. It is important to emphasize that these findings represent an early understanding of the physiological and pathological roles of S1P. The ubiquity of the mediator and its receptors, as well as the evolutionary conservation of S1P metabolism and action, argues that it is a potent and ubiquitous physiological factor in many contexts, and warrant a fuller understanding of its actions at the molecular, cellular and organismal levels. PMID- 15271297 TI - Modulation of adaptive immune responses by sphingosine-1-phosphate. AB - Sphingosine-1-phosphate (S1P) has long been recognized as a mediator of a variety of cell functions. A growing body of evidence has accumulated demonstrating its role in cell migration and as a mediator of growth factor-induced events. In recent years, it has become apparent that S1P also mediates many cytokine and chemokine functions. Cells of the immune system function and migrate in response to a complex network of cytokines and chemokines, and the outcome is determined by the interplay of the effects of these molecules on the target cell. S1P may be a bona fide component of these networks and influence the responses of cells to these immune modulators. PMID- 15271298 TI - Death and taxis: what non-mammalian models tell us about sphingosine-1-phosphate. AB - Sphingosine-1-phosphate (S1P) is a signaling molecule that regulates critical events including mammalian cell proliferation, survival, migration and cell-cell interactions. Most of these signals are triggered by engagement of sphingosine-1 phosphate receptors of the Edg family. However, accumulating evidence derived from investigation of non-mammalian models that lack Edg receptors suggests that sphingosine-1-phosphate-like molecules can act through alternative mechanisms and thereby contribute to morphogenesis, development, reproduction and survival. This review provides an overview of sphingosine-1-phosphate metabolism, the isolation of genes in this pathway employing yeast genetics, the evidence for its influence on non-mammalian development, and the pertinence of these findings to human disease. PMID- 15271300 TI - Gastrula organiser and embryonic patterning in the mouse. AB - Embryonic patterning of the mouse during gastrulation and early organogenesis engenders the specification of anterior versus posterior structures and body laterality by the interaction of signalling and modulating activities. A group of cells in the mouse gastrula, characterised by the expression of a repertoire of "organiser" genes, acts as a source and the conduit for allocation of the axial mesoderm, floor plate and definitive endoderm. The organiser and its derivatives provide the antagonistic activity that modulates WNT and TGFbeta signalling. Recent findings show that the organiser activity is augmented by morphogenetic activity of the extraembryonic and embryonic endoderm, suggesting embryonic patterning is not solely the function of the organiser. PMID- 15271301 TI - A gene network establishing polarity in the early mouse embryo. AB - In mammalian embryos, molecular cross-talk with extraembryonic tissues is essential to elaborate the primary body axes. Here, we review a series of reciprocal interactions that occur shortly after implantation in the uterus, and discuss how they are integrated in a complex signaling network to establish antero-posterior and dorso-ventral polarity. At the heart of this signaling network is the TGFbeta-related protein Nodal which acts on extraembryonic tissues to induce positive and negative feedback regulators at opposite poles of the egg cylinder. This likely results in an activity gradient which is instrumental to pattern the embryo proper. PMID- 15271302 TI - First cell fate decisions and spatial patterning in the early mouse embryo. AB - A growing body of evidence indicates that although the early mouse embryo retains flexibility in responding to perturbations, its patterning is initiated at the earliest developmental stages. There are a few spatial cues that are able to influence the pattern of cleavage divisions: one of these lies in the vicinity of the previous meiotic division, the second is associated with the sperm entry and, related to this, the third is the cell shape. Furthermore, the first cleavage separates the zygote into two cells that tend to follow distinguishable fates: one contributes mainly to the embryonic part of the blastocyst, and the other to the abembryonic. The cumulative effect of the early asymmetries generated through cleavage might lead to asymmetric interactions between the first lineages of cells. This could influence development of patterning after implantation. These early polarity cues serve to bias patterning and not as definitive determinants. PMID- 15271303 TI - Lineage development and polar asymmetries in the peri-implantation mouse blastocyst. AB - The early events of mouse embryogenesis lead to the formation of three distinct cell lineages by the blastocyst: the pluripotent epiblast and the two extraembryonic lineages, the trophoblast and primitive endoderm. Segregation of the lineages depends on the relative levels of expression of key transcription factors, whose localized expression must be controlled by the earlier events of compaction and polarization of the morula. Soon after lineage specification, the two extraembryonic lineages show evidence of early polarities that may relate to the polarity of the postimplantation embryo at gastrulation. The exact relationship between lineage segregation, preimplantation polarities and the postimplantation axes remain to be determined but are now open to molecular and cellular investigation. PMID- 15271304 TI - Lineage allocation and cell polarity during mouse embryogenesis. AB - The first developmental lineage allocation during the generation of the mouse blastocyst is to outer trophoblast or to inner pluriblast (inner cell mass; ICM) cells. This allocation seems to be initiated at the 8-cell stage, when blastomeres polarise. Polarisation is followed by differentiative divisions at the subsequent two cleavage divisions to generate polar outer and non-polar inner 16- and 32-cells. The key events in polarisation are regulated post translationally through a cell contact-mediated pathway, which imposes a heritable determinant-like organisation on the blastomere cortex. Two proteins in particular, E-cadherin and ezrin, are intimately involved in the generation and stabilisation of developmentally significant information. Transcriptional differences between lineages appear to follow and may coincide with the lineage commitment of cells. PMID- 15271305 TI - The developmental origins of mammalian oocyte polarity. AB - Drosophila has been an excellent model system to study the cell and molecular determinants of oocyte axis specification, a problem which is little known in mammalian species. Recent evidence supports the notion that mammalian oocytes utilize axis-orienting properties during the course of oogenesis. Among these, axis specification in relation to the oocyte cortex, germinal vesicle (GV) position, anchoring of GV and spindle, and patterning of follicle cell/oocyte attachments are proposed as conserved features of oogenesis in mammals that may be important to the survival and development of the preimplantation embryo. PMID- 15271306 TI - Polarity in the rabbit embryo. AB - The main aim of the gastrulation process is commonly regarded to be the generation of the definitive germ layers known as mesoderm, endoderm and ectoderm. Here we discuss how the topography of gene expression, cellular migration and proliferative activity in the preliminary germ layers (hypoblast and epiblast) of the rabbit embryo reveal the sequence of events that establishes the three major body axes. We present a testable model in which a combination of cellular movement in the hypoblast with a morphogen gradient created by the (extraembryonic) trophoblast creates morphological polarity in the embryo and, hence, the co-ordinates for germ layer formation. PMID- 15271307 TI - Diversity of germ layer and axis formation among mammals. AB - The class mammalia is composed of approximately 4800 extant species. This class is divided into three subclasses, the prototheria (monotremes), metatheria (marsupials), and eutheria. Surprisingly, there is relatively little knowledge about germ layer and axis formation in mammalian species. Most knowledge about these embryonic processes has been obtained from one species, the mouse, Mus musculus. Here we discuss major variations in germ layer and axis formation among mammals. We suggest that more studies of embryonic development in diverse mammalian species are required for an understanding of germ layer and axis formation to provide insights into human biology and disease. PMID- 15271308 TI - Impact on rectal dose from the use of a prostate immobilization and rectal localization device for patients receiving dose escalated 3D conformal radiation therapy. AB - PURPOSE: When >25% of the rectum is irradiated to > or = 70 Gy, the risk of developing Grade 2 or higher rectal complications is significantly increased. This study evaluates the impact on dose to the rectum from the use of an intrarectal (IR) balloon device, previously shown to immobilize the prostate gland and localize the rectum, in patients receiving dose escalated 3-dimentional (3D) conformal radiation therapy. MATERIALS AND METHODS: From July 2001 through February 2003, 28 consecutive patients with prostate cancer underwent computerized tomography-based simulation with and without the IR balloon in place. Treatment planning was performed for three clinical paradigms in which the IR balloon was not used at all (0 Gy), used during the cone-down for 15 treatments (28.35 Gy), or used for the entire course of 40 treatments (75.6 Gy). The three plans were compared for differences in the percent of rectum receiving >70 Gy. RESULTS: Dose volume histogram (DVH) analysis revealed that the median(range) of percent rectal volume exceeding 70 Gy was 25% (12.7-41.5%), 7.5% (0.9-19.5%), and 3.6% (0-8.7%) for patients in whom the IR balloon was used for 0, 15, and 40 treatments, respectively. The percent of rectum exceeding 70 Gy was significantly different for all treatment plan comparisons (P < 0.0001). CONCLUSIONS: Grade 2 or higher rectal toxicity may be minimized during dose escalated 3D conformal radiation therapy through the use of an IR balloon during the 3-week cone down portion of an 8-week treatment course. PMID- 15271309 TI - Time trends in pathologic features of radical prostatectomy--impact of family history. AB - We investigated whether the clinical or pathological features of patients with a family history of prostate cancer treated by radical prostatectomy differ from patients without a family history. A retrospective analysis of patients treated by radical prostatectomy between 1989 through 2000 was performed. The clinical and pathologic features of patients with a family history (defined as at least one first-degree relative with prostate cancer, N = 103) were compared with those with no family history (N = 456). In addition, the patients were stratified into two groups, those treated from 1989 through 1992 and those treated after 1992. In the entire cohort from 1989 through 2000, patients with a family history had a greater proportion of well-differentiated tumors than the NFH group (26.2% vs. 17.8%; P = 0.05). From 1989 to 1992 there was no statistical difference between patients with a family history (FH) and those without a family history (NFH) with respect to age, prostate specific antigen (PSA), PSA density, clinical or pathologic stage, Gleason grade, or total tumor volume. However, after 1992 the FH group tended to be younger than the NFH group (61.1 vs. 63.4; P = 0.02) and have a lower PSA (6.8 vs. 7.9; P = 0.01) at the time of diagnosis. We believe these differences are predominantly driven by more aggressive screening in patients with a family history of prostate cancer rather than any true genetic differences. PMID- 15271310 TI - Prospective metastatic risk assignment in clinical stage I nonseminomatous germ cell testis cancer: a single institution pilot study. AB - OBJECTIVE: To gain initial experience with a histopathologic model to assign metastatic risk in patients with clinical stage I nonseminomatous germ cell testis cancer (CSI NSGCTC). MATERIALS AND METHODS: Histopathologic factors were recorded prospectively, and metastatic risk assigned according to the proposed model. In the model tested, percentage of embryonal carcinoma (%EMB) > or = 80% and/or vascular invasion (+VI) denoted high (> 50%) occult disease risk, while %EMB < 80% plus absence of VI denoted low (< or = 10%) risk. Risk stratification was correlated with outcome and assessed statistically. RESULTS: There were 54 patients with CSI testis cancer evaluated during the study period. Patients with pure seminoma (n = 30), Sertoli cell tumor (n = 1), and Leydig cell tumor (n = 1) were excluded from analysis. Twenty-two patients had CSI NSGCTC and comprise the pilot study cohort. The median follow-up duration from the time of study entry is 31 months (range, 20-61 months). Utilizing the model tested, a statistically significant higher likelihood of occult disease in the high risk cohort compared to the low risk cohort was observed (67% vs. 0%; Fisher's exact test, P = 0.005). CONCLUSIONS: The results of the present pilot study are encouraging, particularly in the potential of identifying a cohort at low metastatic risk. In the appropriate setting, such a patient might be considered for surveillance alone following orchiectomy. High risk assignment was associated with a positive predictive value (PPV) of 67%. This level of risk is superior to single factor PPV, and if confirmed, could influence clinical decision making. Further experience with this model in an expanded setting is required to establish its reproducibility and predictive value. PMID- 15271311 TI - Benefit of radical cystectomy in the elderly patient with significant co morbidities. AB - Although recent series have demonstrated that radical cystectomy can be safely performed in elderly patients, few if any, have examined the long-term success of this procedure. We sought to determine the long-term benefit and survival outcomes after radical cystectomy in the elderly, high operative risk patient. We reviewed the records of all patients undergoing radical cystectomy between July 1994 and January 2000. Of these 382 patients, we identified 38 patients with transitional cell carcinoma who met our predetermined selection criteria of elderly, high peri-operative risk patients [age > or = 75 years and American Society of Anesthesiologists (ASA) classification > or = 3]. We analyzed patient characteristics, presenting symptoms, pathology, outcomes, and survival. Median age was 79 years (75-87 years). All but a single patient underwent surgery for symptomatic disease. No patient died in the early perioperative period. At a mean follow-up of 22 months (3-90 months), 11/38 (29%) patients are alive. Of the patients with < or = pT2B pathology, 9/27 (33%) are alive and are disease-free. There are 2/11 patients (18%) with > or = pT3 pathology still alive with 1 of those patients (pT4a) alive with disease 34 months after his radical cystectomy. Kaplan-Meier survival curves demonstrate that patients with organ confined disease (< or = pT2B) had a significantly longer mean overall survival than patients with nonorgan confined disease (> or = pT3): 31 months vs. 18 months, P = 0.046. Cause of death was known in 17 patients, with the majority (14/17) because of bladder cancer. However, there were no local recurrences, and palliative goals were achieved in all patients. Our results validate radical cystectomy as a safe and effective treatment choice in the elderly patient with significant co-morbidities. These patients, most of whom are symptomatic, can achieve palliation of their symptoms, local control, and long term survival, especially if their bladder cancer is organ confined. Reluctance to offer timely, aggressive local therapy may compromise ultimate survival, even amongst high operative risk, elderly patients. PMID- 15271312 TI - A contemporary evaluation of cytoreductive nephrectomy with tumor thrombus: Morbidity and long-term survival. AB - BACKGROUND: Metastatic renal cell carcinoma (RCC) is an aggressive entity that frequently invades the venous system. We evaluated the morbidity and survival of patients with tumor thrombus who undergo cytoreductive nephrectomy. MATERIALS AND METHODS: We identified 56 patients from our institution's database who had a primary renal tumor in place and documented metastases at the time of surgery. We reviewed demographic and pathologic characteristics from these patients as well as complications and overall survival. RESULTS: Median age was 58 (37-77). There were 33 patients (59%) who had tumor thrombus with 21 (64%) involving the renal vein, 10 (30%) involving the infradiaphragmatic inferior vena cava (IVC), and 2 (6%) involving the supradiaphragmatic IVC. Median tumor size for thrombus patients was 12 cm (5-29). There were 8 (14.2%) who had complications, including 1 death. Thrombus patients were significantly more likely to have a complication (P = 0.008). Median survival for all patients was 10.7 months (0.3-61). There was no significant difference in overall survival between patients with and without thrombus (P = 0.76). CONCLUSIONS: Patients who undergo cytoreductive nephrectomy with a tumor thrombus have a higher rate of complications as compared to patients undergoing cytoreductive nephrectomy without tumor thrombus. The long-term survival, however, was not statistically different and thus aggressive surgery for select metastatic RCC patients is warranted. PMID- 15271313 TI - Cytotoxicity of cisplatin in bladder cancer is significantly enhanced by application of bcl-2 antisense oligonucleotides. AB - BACKGROUND: The aim of our study was to examine the effects of the combined application of cisplatin and bcl-2 antisense oligonucleotide on human bladder cancer cell lines to determine the possible synergistic effects in cytotoxicity and to estimate its potential value for subsequent in vivo trials. MATERIALS AND METHODS: Human bladder cancer cell lines (UM-UC 3, RT 112, T24/83 and HT 1197) were treated with bcl-2 antisense oligonucleotide, cisplatin, or a combination of both and incubated for 48 h under standard conditions. Cell survival was determined using a Neubauer haemocytometer or standard MTT assay. BCL-2 expression was verified using western blotting. RESULTS: The combined treatment resulted in significant lower cell survival rates compared to individual treatment. Additionally, there was a decrease in cell survival rate with an increase in cisplatin concentration in combined treatment that was not observed in cisplatin mono treatment. CONCLUSIONS: For the combined treatment with oligonucleotides and cisplatin a synergistic effect can be strongly suggested. Therefore, further investigations and in vivo trials have to be done to determine the possible benefits for clinical applications. PMID- 15271314 TI - Complete radiological and metabolic response of metastatic renal cell carcinoma to SU5416 (semaxanib) in a patient with probable von Hippel-Lindau syndrome. AB - We report a case of a patient with probable von Hippel-Lindau (VHL) syndrome and metastatic renal cell cancer (RCC) who had a complete radiological and metabolic response to SU5416 (semaxanib). The patient was enrolled on a clinical study examining the efficacy of SU5416 in patients with metastatic cancer. Treatment with SU5416 was given at a dose of 145 mg/m2 intravenously twice-weekly for 11 doses. The patient achieved an early metabolic response on an F-18 fluorodeoxyglucose (FDG) Positron Emission Tomographic (PET) scan within 2 weeks of therapy. Subsequent computerized tomography (CT) and PET scans (9 and 12 months after treatment, respectively) confirmed ongoing complete radiological and metabolic response. He remains tumor-free 18 months after treatment. This is the first documented report of metastatic RCC in the setting of presumed VHL syndrome responding to treatment with SU5416. While vascular endothelial growth factor (VEGF) inhibitors have been shown to produce a modest response in sporadic metastatic RCC, further studies utilizing VEGF inhibitors in patients with VHL syndrome and RCC warrants exploration. PMID- 15271315 TI - Importance of lymph node dissection in urologic cancers. PMID- 15271316 TI - The role of lymphadenectomy in prostate cancer. PMID- 15271318 TI - The evolving role of pelvic lymphadenectomy in the treatment of bladder cancer. AB - Regional lymphadenectomy is integral to the surgical management of high-grade invasive bladder cancer. A growing body of evidence suggests that a lymph node dissection may provide not only improved prognostic information, but also a clinically significant therapeutic benefit for both lymph node positive and negative patients undergoing radical cystectomy. While the inclusion of lymph node resection in conjunction with radical cystectomy for patients with clinically negative nodes is well accepted, the extent of the nodal dissection remains highly contentious. Similarly, the benefit of node dissection for patients with advanced disease and gross adenopathy or for those with superficial disease (Ta, T1 or TIS) remains a topic of heated debate. This review describes the historical evolution of lymphadenectomy in the surgical treatment of bladder cancer and provides a comprehensive review of the current literature addressing the role of lymph node dissection in the treatment of bladder cancer. PMID- 15271320 TI - The role of lymphadenectomy in the surgical management of renal cell carcinoma. AB - After decades of evaluation, the role of lymphadenectomy in the management of renal cell carcinoma remains a controversy. Contemporary series suggest that the true incidence of isolated lymph node metastases in clinically localized disease is small, and the location of such metastases is unpredictable. While several institutional series have suggested a therapeutic benefit for extended lymphadenectomy, there remains a lack of randomized data to support its routine use. Despite this, there remains a role for lymphadenectomy in individuals with high risk of lymph node metastasis or known lymphadenopathy in whom few other options exist for aggressive, potentially curative therapy. PMID- 15271322 TI - The role of retroperitoneal lymph node dissection in the management of testicular cancer. AB - Despite continued refinement in terms of technique and the integration of retroperitoneal lymph node dissection (RPLND) in the management of patients with testicular cancer, RPLND remains an essential component in the ultimate cure of these patients. The failure to eradicate all disease in the retroperitoneum exposes patients to the risk of late relapse events with potentially lethal consequences. For patients with low-stage nonseminomatous germ cell tumor (NSGCT), primary RPLND is an important staging tool to define subsequent treatment requirements, simplify the follow-up of patients by obviating the need for routine abdominal imaging, and limit the exposure of patients to the long term toxicity of chemotherapy. RPLND alone is curative in up to 90% of patients with low-volume retroperitoneal disease. In the post-chemotherapy setting, the inability to reliably exclude the presence of teratoma or viable germ cell cancer in the retroperitoneum mandates that post-chemotherapy RPLND be performed for all NSGCT patients with residual masses. With improvements in surgical technique and perioperative care, RPLND is associated with minimal short- and long-term morbidity in the hands of experienced surgeons at dedicated centers. This article reviews the role of RPLND in the management of patients with NSGCT at all stages and its role in advanced seminoma. PMID- 15271324 TI - The role of lymphadenectomy in penile cancer. AB - In patients with squamous carcinoma of the penis, the presence and extent of metastases involving the inguinal nodes are the most important factors predictive of survival. Favorable prognostic indicators of cure in surgically treated patients in whom metastases develop include: (1) minimal nodal disease, (2) unilateral involvement, (3) no evidence of extranodal extension of cancer, and (4) absence of pelvic nodal metastases. Prophylactic lymphadenectomy in select patients at high risk for metastasis seems reasonable in lieu of prospective randomized trials because novel procedures have significantly decreased the morbidity of surgical staging. Patients with poor prognostic indicators either before or after surgery should be considered for multimodal therapy. PMID- 15271326 TI - Critique of laparoscopic lymphadenectomy in genitourinary oncology. AB - Regional lymphadenectomy is prognostic and selectively therapeutic in urologic oncology. The role of lymphadenectomy continues to be defined with the evolving multimodal management of genitourinary malignancies. Laparoscopy is playing a greater role in the management of genitourinary malignancies and thus, it is germane to critique the role of laparoscopic lymphadenectomy in the management of these tumors. Review of the literature suggests that laparoscopic pelvic lymphadenectomy is feasible with nodal yields commensurate to those in open published series. Although laparoscopic retroperitoneal lymph node dissection for nonseminomatous germ cell tumor is feasible, the technique and efficacy of this procedure require further investigation. PMID- 15271328 TI - When is a negative lymph node really negative? Molecular tools for the detection of lymph node metastasis from urological cancer. PMID- 15271342 TI - Polymerix corporation: polymer drugs for device and injection. PMID- 15271343 TI - Daptomycin structure and mechanism of action revealed. AB - Daptomycin kills otherwise antibiotic-resistant gram-positive pathogens and is the first lipopeptide antibiotic to reach the clinic. Elucidation of its 3D structure and mechanism of action, reported in this issue of Chemistry & Biology, will facilitate the design and engineering of new, potentially life-saving antibiotics. PMID- 15271344 TI - Mapping key elements of a protein motif. AB - In this issue of Chemistry & Biology, Weiss and colleagues use phage display to map residues in the engrailed homeodomain that influence DNA recognition. Their shotgun scanning data provides surprising new insights into the importance of regions outside the recognition helix and N-terminal arm for DNA binding. PMID- 15271345 TI - Cellular addresses; step one in creating a glycocode. AB - In this issue of Chemistry & Biology, a library screening approach reveals at least four types of enzymes that attach galactosamine to build cell surface mucin type glycoproteins. A better molecular understanding of how these information carrying oligosaccharides are created sets the stage for designing more selective inhibitors and potential therapeutics. PMID- 15271346 TI - Novel mechanism for priming aromatic polyketide synthases. AB - In this issue of Chemistry & Biology, a novel priming mechanism is proposed for aromatic polyketide biosynthesis, with an iterative type I polyketide synthase generating a starter unit primed for a type II polyketide synthase. This novel priming system participates in hedamycin biosynthesis, a DNA alkylating agent. PMID- 15271347 TI - Ribozyme diagnostics comes of age. AB - Biosensing ribozymes could soon be used to diagnose viral infection. The Kossen group from Sirna Therapeutics have developed a sensitive, high-throughput means of screening for hepatitis C virus, using their target activated half-ribozyme technology, as reported in the June issue of Chemistry & Biology. PMID- 15271348 TI - A human single-chain antibody specific for integrin alpha3beta1 capable of cell internalization and delivery of antitumor agents. AB - Selective antitumor chemotherapy can be achieved by using antibody-drug conjugates that recognize surface proteins upregulated in cancer cells. One such receptor is integrin alpha3beta1, which is overexpressed on malignant melanoma, prostate carcinoma, and glioma cells. We previously identified a human single chain Fv antibody (scFv), denoted Pan10, specific for integrin alpha3beta1 that is internalized by human pancreatic cancer cells. Herein, we describe the chemical introduction of reactive thiol groups onto Pan10, the specific conjugation of the modified scFv to maleimide-derivatized analogs of the potent cytotoxic agent duocarmycin SA, and the properties of the resultant conjugates. Our findings provide evidence that Pan10-drug conjugates maintain the internalizing capacity of the parent scFv and are cytotoxic at nanomolar concentrations. Our Pan10-drug conjugates may be promising candidates for targeted chemotherapy of malignant diseases associated with overexpression of integrin alpha3beta1. PMID- 15271349 TI - Chemical genetic identification of the histamine H1 receptor as a stimulator of insulin-induced adipogenesis. AB - A large collection of bioactive compounds with diverse biological effects can be used as probes to elucidate new biological mechanisms that influence a particular cellular process. Here we analyze the effects of 880 well-known small-molecule bioactives or drugs on the insulin-induced adipogenesis of 3T3-L1 fibroblasts, a cell-culture model of fat cell differentiation. Our screen identified 86 compounds as modulators of the adipogenic differentiation of 3T3-L1 cells. Examination of their chemical and pharmacological information revealed that antihistamine drugs with distinct chemical scaffolds inhibit differentiation. Histamine H1 receptor is expressed in 3T3-L1 cells, and its knockdown by small interfering RNA impaired the insulin-induced adipogenic differentiation. Histamine receptors and histamine-like biogenic amines may play a role in inducing adipogenesis in response to insulin. PMID- 15271350 TI - Characterization and investigation of substrate specificity of the sugar aminotransferase WecE from E. coli K12. AB - WecE gene, encoding a sugar aminotransferase (SAT), has been cloned from E. coli K12 and expressed in E. coli BL21 (DE3). The enzyme was purified and characterized. WecE used TDP-4-keto-6-deoxy-D-glucose (TDP-D-Glc4O) and L glutamate as a good amino acceptor and donor, respectively, leading to the production of TDP-4-amino-4,6-dideoxy-D-galactose (TDP-Fuc4N), which was identified by NMR studies. WecE also showed a similar activity for TDP-4-keto 6 deoxy-D-mannose (TDP-D-Man4O), but no activity for GDP-4-keto-6-deoxy-D-mannose (GDP-D-Man4O), suggesting that the nucleotide moiety would become a key determinant to the substrate specificity of amine acceptor for the activity of the SAT. Multiple alignments showed that SATs have four highly conserved motifs located around the active site and could be divided into three subgroups (VIalpha, VIbeta, and VIgamma) that might be closely related with their substrate specificities. PMID- 15271351 TI - The biosynthetic gene cluster for a monocyclic beta-lactam antibiotic, nocardicin A. AB - The monocyclic beta-lactam antibiotic nocardicin A is related structurally and biologically to the bicyclic beta-lactams comprised of penicillins/cephalosporins, clavams, and carbapenems. Biosynthetic gene clusters are known for each of the latter, but not for monocyclic beta-lactams. A previously cloned gene encoding an enzyme specific to the biosynthetic pathway was used to isolate the nocardicin A cluster from Nocardia uniformis. Sequence analysis revealed the presence of 14 open reading frames involved in antibiotic production, resistance, and export. Among these are a two-protein nonribosomal peptide synthetase system, p-hydroxyphenylglycine biosynthetic genes, an S adenosylmethionine-dependent 3-amino-3-carboxypropyl transferase (Nat), and a cytochrome P450. Gene disruption mutants of Nat, as well as an activation domain of the NRPS system, led to loss of nocardicin A formation. Several enzymes involved in antibiotic biosynthesis were heterologously overproduced, and biochemical characterization confirmed their proposed activities. PMID- 15271352 TI - A distance ruler for RNA using EPR and site-directed spin labeling. AB - As a basic model study for measuring distances in RNA molecules using continuous wave (CW) EPR spectroscopy, site-directed spin-labeled 10-mer RNA duplexes and HIV-1 TAR RNA motifs with various interspin distances were examined. The spin labels were attached to the 2'-NH2 positions of appropriately placed uridines in the duplexes, and interspin distances were measured from both molecular dynamics simulations (MD) and Fourier deconvolution methods (FD). The 10-mer duplexes have interspin distances ranging from 10 A to 30 A based on MD; however, dipolar line broadening of the CW EPR spectrum is only observed for the RNAs for predicted interspin distances of 10-21 A and not for distances over 25 A. The conformational changes in TAR (transactivating responsive region) RNA in the presence and in the absence of different divalent metal ions were monitored by measuring distances between two nucleotides in the bulge region. The predicted interspin distances obtained from the FD method and those from MD calculations match well for both the model RNA duplexes and the structural changes predicted for TAR RNA. These results demonstrate that distance measurement using EPR spectroscopy is a potentially powerful method to help predict the structures of RNA molecules. PMID- 15271353 TI - Structural transitions as determinants of the action of the calcium-dependent antibiotic daptomycin. AB - Daptomycin is a cyclic anionic lipopeptide antibiotic recently approved for the treatment of complicated skin infections (Cubicin). Its function is dependent on calcium (as Ca2+). Circular dichroism spectroscopy indicated that daptomycin experienced two structural transitions: a transition upon interaction of daptomycin with Ca2+, and a further transition upon interaction with Ca2+ and the bacterial acidic phospholipid, phosphatidyl glycerol. The Ca2+-dependent insertion of daptomycin into model membranes promoted mild and more pronounced perturbations as assessed by the increase of lipid flip-flop and membrane leakage, respectively. The NMR structure of daptomycin indicated that Ca2+ induced a conformational change in daptomycin that increased its amphipathicity. These results are consistent with the hypothesis that the association of Ca2+ with daptomycin permits it to interact with bacterial membranes with effects that are similar to those of the cationic antimicrobial peptides. PMID- 15271354 TI - The hedamycin locus implicates a novel aromatic PKS priming mechanism. AB - The biosynthetic gene cluster for the pluramycin-type antitumor antibiotic hedamycin has been cloned from Streptomyces griseoruber. Sequence analysis of the 45.6 kb region revealed a variety of unique features such as a fabH homolog (KSIII), an acyltransferase (AT) gene, a set of type I polyketide synthase (PKS) genes, and two putative C-glycosyltransferase genes. As the first report of the cloning of the biosynthetic gene cluster for the pluramycin antibiotics, this work suggests that the biosynthesis of pluramycins utilize an iterative type I PKS system for the generation of a novel starter unit that subsequently primes the type II PKS system. It also implicates the involvement of a second catalytic ketosynthase (KSIII) to regulate this unusual priming step. Gene disruption is used to confirm the importance of both type I and II PKS genes for the biosynthesis of hedamycin. PMID- 15271355 TI - Characterization and genetic manipulation of peptide synthetases in Pseudomonas aeruginosa PAO1 in order to generate novel pyoverdines. AB - PvdD, a nonribosomal peptide synthetase (NRPS) of Pseudomonas aeruginosa PAO1, incorporates two L-threonines into the siderophore pyoverdine. A pvdD mutant did not synthesize pyoverdine and lacked a high Mr iron-regulated cytoplasmic protein (IRCP). Analysis of other IRCPs and the P. aeruginosa genome enabled the remaining pyoverdine NRPSs to be identified. The pvdD mutation could be complemented in trans, enabling design of plasmid-based systems for the generation of novel pyoverdines. Introduction of a truncated pvdD gene resulted in attenuated forms of pyoverdine, and introduction of L-threonine-incorporating NRPSs from other organisms restored pyoverdine production to mutant cells. This is the first successful rational in vivo modification of NRPS modules outside of Bacillus subtilis. The systems employed did not allow incorporation of other residues into pyoverdine, indicating that there are multiple elements contributing toward substrate specificity in NRPSs. PMID- 15271356 TI - Directed evolution of epoxide hydrolase from A. radiobacter toward higher enantioselectivity by error-prone PCR and DNA shuffling. AB - The enantioselectivity of epoxide hydrolase from Agrobacterium radiobacter (EchA) was improved using error-prone PCR and DNA shuffling. An agar plate assay was used to screen the mutant libraries for activity. Screening for improved enantioselectivity was subsequently done by spectrophotometric progress curve analysis of the conversion of para-nitrophenyl glycidyl ether (pNPGE). Kinetic resolutions showed that eight mutants were obtained with up to 13-fold improved enantioselectivity toward pNPGE and at least three other epoxides. The large enhancements in enantioselectivity toward epichlorohydrin and 1,2-epoxyhexane indicated that pNPGE acts as an epoxyalkane mimic. Active site mutations were found in all shuffled mutants, which can be explained by an interaction of the affected amino acid with the epoxide oxygen or the hydrophobic moiety of the substrate. Several mutations in the shuffled mutants had additive effects. PMID- 15271357 TI - Quantum chemical calculations and mutational analysis suggest heat shock protein 90 catalyzes trans-cis isomerization of geldanamycin. AB - The affinity of geldanamycin (GA) for binding to heat shock protein 90 (HSP90) is 50- to 100-fold weaker than is the affinity of the structurally distinct natural product radicicol. X-ray crystallography shows that although radicicol maintains its free conformation when bound to HSP90, the conformation of GA is dramatically altered from an extended conformation with a trans amide bond to a kinked shape in which the amide group in the ansa ring has the cis configuration. We have performed ab initio quantum chemical calculations to demonstrate that the trans cis isomeriztion of GA in solution is both kinetically and thermodynamically unfavorable. Thus, we propose that HSP90 catalyzes the isomerization of GA. We identify Ser113, a conserved residue outside the ATP binding pocket, as essential for the isomerization of GA. In support of this model, we show that radicicol binds equally well to both wild-type HSP90 and the Ser113 mutant, whereas the binding of GA to the Ser113 mutant is decreased significantly from its binding to wild-type HSP90. Based on this finding, a mechanism of keto-enol tautomerization of GA catalyzed by HSP90 is proposed. The added requirement of isomerization prior to tight binding may explain the enhanced binding affinity of GA for HSP90 in a cell extract versus in a purified form. PMID- 15271358 TI - A general approach to detect protein expression in vivo using fluorescent puromycin conjugates. AB - Understanding the expression of known and unknown gene products represents one of the key challenges in the post-genomic world. Here, we have developed a new class of reagents to examine protein expression in vivo that does not require transfection, radiolabeling, or the prior choice of a candidate gene. To do this, we constructed a series of puromycin conjugates bearing various fluorescent and biotin moieties. These compounds are readily incorporated into expressed protein products in cell lysates in vitro and efficiently cross cell membranes to function in protein synthesis in vivo as indicated by flow cytometry, selective enrichment studies, and Western analysis. Overall, this work demonstrates that fluorescent-puromycin conjugates offer a general means to examine protein expression in vivo. PMID- 15271359 TI - Deconvoluting the functions of polypeptide N-alpha acetylgalactosaminyltransferase family members by glycopeptide substrate profiling. AB - The polypeptide N-alpha-acetylgalactosaminyltransferases (ppGalNAcTs) play a key role in mucin-type O-linked glycan biosynthesis by installing the initial GalNAc residue on the protein scaffold. The preferred substrates and functions of the >20 isoforms in mammals are not well understood. However, current data suggest that glycosylated mucin domains are created by the successive, often hierarchical, action of several specific ppGalNAcTs. Herein we analyzed the glycopeptide substrate preferences of several ppGalNAcT family members using a library screening approach. A 56-member glycopeptide library designed to reflect a diversity of glycan clustering was assayed for substrate activity with ppGalNAcT isoforms using an azido-ELISA. The data suggest that the ppGalNAcTs can be classified into at least four types, which working together, are able to produce densely glycosylated mucin glycoproteins. PMID- 15271360 TI - Dissecting the Engrailed homeodomain-DNA interaction by phage-displayed shotgun scanning. AB - Phage-displayed alanine shotgun scanning was used to dissect contributions by engrailed homedomain (En-HD) residues 17 through 46, which indirectly influence recognition of DNA. The relative contributions of such indirect contacts, quantified by shotgun scanning, highlight previously unexplored En-HD residues. Two motifs dominate En-HD function in this region. First, two surface-exposed aromatic residues (F20 and Y25) bracket the hydrophobic core. Second, two sets of turn-forming residues are highlighted, including carboxamide-requiring residues E22/N23 and a leucine/isoleucine splint. The En-HD hydrophobic core exhibits a surprising degree of malleability, as demonstrated by homolog shotgun scanning. Most selectants from in vitro shotgun scanning mirror the consensus human homeodomain sequence. Thus, natural evolution and in vitro selection use similar selection criteria: affinity, specificity, and stability. However, homolog shotgun scanning identified mutations capable of improving the affinity and specificity of En-HD. PMID- 15271361 TI - Renal imaging: what the urologist wants to know. AB - Preoperative imaging in renal surgery is of utmost importance in contemporary surgical practice. From a diagnostic standpoint, imaging discovers many renal tumors incidentally before they become symptomatic. These tumors often are amenable to partial renal resection or minimally invasive surgical approaches. In general, surgical interventions for renal abnormalities have evolved to a less invasive endourologic or laparoscopic approach. Selection of the appropriate surgical intervention for renal tumors, collecting system tumors, and hydronephrosis depends heavily on the anatomy of the renal pathology. Thus, renal imaging is crucial in clinical decision-making. This article reviews the contribution of imaging to the surgical management of renal tumors, upper tract urothelial tumors, and ureteropelvic junction obstruction. PMID- 15271362 TI - MR imaging of cystic renal masses. AB - MR imaging has proven to be an important imaging modality in the evaluation of cystic renal masses. Because it is becoming more widely used, it is necessary to be able to characterize cystic renal masses accurately using MR imaging alone. We review the indications for the use of MR imaging and discuss the findings present in a variety of cystic renal masses. The role of MR imaging in the characterization of cystic lesions of the kidney is summarized and compared with the CT findings in the Bosniak renal cyst classification. It is expected that the increasing experience with the use of MR imaging in cystic renal masses will add to our ability to diagnose and manage these cases successfully. PMID- 15271363 TI - MR evaluation of solid renal masses. AB - After diagnosis of a suspicious renal mass on ultrasound or CT, renal MR imaging typically is ordered to characterize the mass further, stage the mass, or resolve discordant ultrasound and CT results. MR imaging may also be ordered in cases in which ultrasound is poor or in instances in which contrast-enhanced CT may be ill advised. PMID- 15271364 TI - Image-guided ablation of renal cell carcinoma. AB - An increasing number of small asymptomatic renal cell carcinomas (RCCs) are being detected by cross-sectional imaging. Because of the nonaggressive biologic behavior of many of these tumors, there is increasing interest in minimally invasive treatment modalities,particularly for the elderly, infirm, and patients with comorbid conditions. Radiofrequency(RF) ablation, cryoablation, microwave ablation, and laser ablation have all shown promise for the treatment of RCC, with high local control and low complication rates for RF ablation and cryoablation. However, the clinical trial data remain early, and survival data are not yet available for a definitive comparison with conventional surgical techniques for removal of RCC. PMID- 15271365 TI - Cystic renal disease. AB - Renal cysts are relatively common radiographic and clinical abnormalities. Cystic renal disease is a heterogeneous entity comprised of heritable, developmental, and acquired disorders. During the last decade, considerable progress has been made in reaching a consensus for standard terminologies and classifications of cystic renal disease among radiologists, pathologists, nephrologists, and urologists. This article discusses more common MR imaging-related types of cystic renal disease. PMID- 15271366 TI - Functional renal MR imaging. AB - MR imaging is the only noninvasive test that may provide a complete picture of renal status with minimal risk to the patient, simultaneously improving diagnosis and lowering costs. This article reviews several MR renography techniques, including approaches for quantifying renal perfusion and glomerular filtration rate. Also discussed are clinical applications for the diagnosis and follow-up of renovascular disease, hydronephrosis,and renal transplant dysfunction. The article concludes with an overview of technical problems and challenges facing MR renography. PMID- 15271367 TI - Renal MR angiography. AB - The high accuracy of renal MR angiography makes it well suited for diagnosing renal vascular disease. A comprehensive examination includes three-dimensional gadolinium MR angiography to assess lumenal anatomy and functional techniques to assess the hemodynamic significance of any stenosis identified. Postprocessing is critical to provide reformations, maximum intensity projections, and optional volume-rendered images to display arteries in an angiographic format for optimal demonstration of any vascular lesions. It is important to review source images to avoid missing pathologic findings. As MR imaging continues to develop, the renal MR angiography examination will likely expand to include extensive functional information about creatinine clearance, flow, and response to pharmacologic agents as well as spectroscopy, diffusion, perfusion, phase contrast, and other techniques. PMID- 15271368 TI - Kidney transplantation: evaluation of donors and recipients. AB - MR imaging provides a comprehensive method for noninvasive evaluation of renal donor anatomy. Although multidetector helical CT can provide similar information, MR imaging has the advantage of avoiding exposure to ionizing radiation and potentially nephrotoxic contrast material. These are important considerations in screening a generally healthy donor population. MR imaging also can provide complete evaluation of the kidney after transplantation, where avoidance of potentially nephrotoxic agents and preservation of maximal renal function are critical. PMID- 15271369 TI - MR imaging of the adrenal glands. AB - MR imaging is used commonly for imaging the adrenal glands. Its high-contrast resolution and multiplanar imaging capability enables the detection and characterization of many adrenal masses. The advent of chemical-shift imaging revolutionized the role of MR imaging in characterizing adrenal masses. In this article, the authors discuss the range of MR appearances of common and uncommon adrenal masses, focusing on the nonfunctioning incidentally discovered mass and its characterization methods. MR imaging is continuously improving. The increasing use of higher strength magnets and the introduction of newer coils, sequences, and techniques will help detect and characterize very small adrenal masses, quantify their fat content, and provide exquisite morphologic images of the gland and its vascular supply. PMID- 15271370 TI - MR imaging of the bladder. AB - MR imaging is a useful modality for evaluating diseases of the bladder. MR imaging can detect and stage bladder cancer by determining the presence and depth of muscle invasion. Direct multiplanar imaging and superb soft-tissue contrast make MR imaging an ideal modality for evaluating less common neoplastic diseases of the bladder, such as urachal carcinoma, and tumors that develop within bladder diverticula. Dynamic breath-held fast T2-weighted imaging can evaluate for cystocele and other components of pelvic floor relaxation. PMID- 15271371 TI - MR imaging and MR spectroscopic imaging of prostate cancer. AB - The primary indication for prostate MR and MR spectroscopic imaging is evaluating men with newly diagnosed prostate cancer who are deciding whether to undergo surgery or radiotherapy. Other applications of MR and MR spectroscopic imaging in prostate cancer are not defined fully. Areas of research include volumetric localization of prostate cancer, in vivo MR spectroscopic imaging at high field strength, in vitro MR spectroscopic imaging at very high field strength, novel spectroscopic markers of malignancy,and interventional MR guidance of biopsy and therapy. MR spectroscopic imaging remains a relatively novel technique, and successful implementation is demanding. Nonetheless, only MR and MR spectroscopic imaging allow structural and metabolic evaluation of prostate cancer location, aggressiveness, and stage, and MR imaging provides clinically and therapeutically relevant information on prostatic and periprostatic anatomy. PMID- 15271372 TI - MR imaging staging of pelvic lymph nodes. AB - Accurate pelvic nodal staging is important in the workup of many pelvic tumors for assessing prognosis and directing therapy. Advances in diagnostic imaging have played an integral role in the staging of these tumors. Cross-sectional imaging, including MR imaging,however, uses size criteria and morphology to infer malignancy within a node,which is neither sensitive nor specific. This article reviews the normal pelvic nodal anatomy and techniques of conventional MR imaging for optimal nodal evaluation as well as introducing the recent technique of ultra small supraparamagnetic iron oxide(USPIO)-enhanced MR lymphangiography, which uses nodal function rather than structural criteria in assessing for metastatic nodes. PMID- 15271374 TI - Human interleukin-5 expression is synergistically regulated by histone acetyltransferase CBP/p300 and transcription factors C/EBP, NF-AT and AP-1. AB - Interleukin-5 (IL-5) plays a central role in the growth and differentiation of eosinophils and contributes to several disease states including asthma. There has been considerable interest in the identification of the transcriptional mechanisms controlling the synthesis of this cytokine. The regulation of IL-5 message is primarily at the level of transcription and is likely to be controlled, to a large extent, by regulatory elements in the promoter region that can influence the transcriptional activity of the gene. In this study, we performed a series of transient transfection experiments with IL-5 promoter reporter gene construct and expression plasmid for E1A, an inhibitor of CBP/p300, or for the CBP/p300-binding defective E1A Delta 2-36. The results showed that E1A repressed IL-5 promoter activity, while E1A Delta 2-36 had no effect on it. This suggested that CBP/p300 was involved in regulation of IL-5 gene expression. Transcriptional coactivator CBP/p300 and transcription factors C/EBP, NF-AT, and c-Fos synergistically activated IL-5 promoter. Furthermore, we found that ectopic expression of p300 increased endogenous IL-5 mRNA expression. Thus, the histone acetyltransferase (HAT) activity of CBP/p300 was required to activate IL-5 expression. This report provided evidence, for the first time, that CBP/p300 was involved in IL-5 gene expression and the HAT activity was important in regulation of IL-5 expression. PMID- 15271375 TI - Effects of vitamin C on intracytoplasmic cytokine production in human whole blood monocytes and lymphocytes. AB - BACKGROUND: Vitamin C (ascorbic acid) is an essential water-soluble nutrient which primarily exerts its effect on immune homeostasis as physiological antioxidant. However, conflicting data exist regarding the effect of vitamin C on the expression of pro-inflammatory cytokines. METHODS: It was the aim of this study to investigate the impact of vitamin C on intracytoplasmic production of pro-inflammatory cytokines in monocytes and lymphocytes by flow cytometry after human whole blood assay. RESULTS: Vitamin C dose dependently inhibited the LPS induced number of monocytes producing IL-6 (e.g., 41.0% reduction, p < 0.001, 20 mM vitamin C) and TNF-alpha (e.g., 26.0% reduction, p < 0.005, 20 mM vitamin C). Simultaneously, the number of lymphocytes producing IL-2 after PMA/ionomycin stimulation was dose dependently reduced (e.g., 24.2% inhibition, p < 0.005, 20 mM vitamin C). Notably, the number of IL-1 and IL-8 producing monocytes as well as TNF-alpha and IFN-gamma producing lymphocytes were not significantly affected by 20 mM vitamin C. CONCLUSIONS: These data suggest that vitamin C selectively influences intracytoplasmic cytokine production and therefore, further studies are needed to elucidate the underlying mechanisms of immunomodulation, i.e. regulation of NF kappa B activation which is mandatory for the induction of pro inflammatory cytokines. PMID- 15271376 TI - Orally administered IL-6 induces elevated intestinal GM-CSF gene expression and splenic CFU-GM. AB - Orally administered interleukin (IL)-6 has been shown to be of benefit in eliminating Campylobacter infection and in preventing sepsis following hemorrhage. In related experiments, it was seen that proliferating cells were found in the spleens of untreated mice given IL-6 by oral gavage. Injection of the DNA label, BrdU, showed that significant proliferation began at 4 h and peaked at 24 h in the splenic red pulp of animals given oral IL-6. Mice given saline showed no increase in splenic BrdU uptake. Histological analysis suggested a hematopoietic lineage for these cells. Clonogenic assays performed on spleen cells taken from mice given oral IL-6 revealed that increased granulocyte macrophage colony forming units (GM-CFU) were present at 24 h post-IL-6 administration. No increase in GM colonies occurred in mice fed IL-3, granulocyte colony stimulating factor (G-CSF) or granulocyte-macrophage (GM)-CSF. RT-PCR analysis of intestinal mRNA from treated mice revealed that GM-CSF mRNA was elevated at 4 h after oral IL-6 administration, but not in mice fed other cytokines. It is suggested that oral administration of IL-6 induces both proliferation and a brief elevation of GM-CFU in the hematopoietic spleens of mice. This increase appears to be the result of increased GM-CSF mRNA production in the intestines of mice fed IL-6. PMID- 15271377 TI - Continuous interferon-gamma or tumor necrosis factor-alpha exposure of enterocytes attenuates cell death responses. AB - Short-term stimulation (i.e. <2 days) with tumor necrosis factor-alpha (TNF alpha) or interferon-gamma (IFN-gamma) cause growth arrest and sensitize epithelial cells to CD95 (Fas/Apo-1)-mediated cell death. The effect of long-term cytokine exposure on viability, proliferation, and apoptosis response of colonic epithelial cells is unknown and addressed in this study. In the present study HT29 and DLD-1 colonic cells were stimulated with either TNF-alpha or IFN-gamma at varying concentrations for 2-9 days. Viability and proliferation was assessed. CD95-mediated cell death response was determined. IFN-gamma caused decreased viability at high concentrations (1 nM), whereas lower concentrations (10-100 pM) only caused a transient growth arrest. TNF-alpha (100 pM) did not affect cell growth. Cells stimulated for 8 days with IFN-gamma (10 pM) or TNF-alpha (100 pM) had higher proliferation rates than controls or cells stimulated for 2 days (p < 0.05). Whereas the spontaneous cell death increased slightly during continuous cytokine exposure the CD95L response decreased (P < 0.01). Colonic cells continuously exposed to IFN-gamma or TNF-alpha had cell turnover characteristics that resemble findings in patients with UC. Increased proliferation and decreased cell death response may act as a counter regulatory mechanism that limits the damaging effects of cytokines. PMID- 15271378 TI - Comparison of results using the gel microdrop cytokine secretion assay with ELISPOT and intracellular cytokine staining assay. AB - The gel microdrop (GMD) secretion assay involves encapsulating single cells in a biotinylated agarose matrix, addition of a streptavidin bridge, diffusion of a biotinylated capture antibody, and detection of secreted molecules using a fluorescently labeled reporter antibody. Using flow cytometry, encapsulated cells can be analyzed or recovered based on cell type and secretory profile. Using murine Th2 cell line D10.G4.1 as a model, we recently demonstrated the feasibility of using the GMD cytokine secretion assay combined with flow cytometry to detect IL-4-producing cells after stimulation with the mitogen, Con A. In addition, subpopulations of encapsulated cells secreting IL-4 were simultaneously characterized by immunophenotyping. We found good correlation between results using the GMD cytokine secretion assay and results with the standard ELISPOT and intracellular cytokine (ICC) assays. The GMD cytokine secretion assay permits simultaneous detection of secreted cytokine and determination of cell surface phenotype on viable, single cells. Moreover, using fluorescence activated cell sorting (FACS), secreting cells of interest can be sorted, recovered, and cultured for further studies. PMID- 15271379 TI - Quantifying Aotus monkey cytokines by real-time quantitative RT-PCR. AB - Aotus spp. monkeys are considered the ideal model for studying the progress of malarial infection and the immune response it elicits. We describe the use of a recently developed technique, real-time quantitative RT-PCR, to quantify several Aotus monkey cytokine mRNAs involved in Th1/Th2 responses (IL-4, IL-10, TNF-beta and IFN-gamma). Specific primers were designed for each cytokine and standard curves were constructed using serial dilutions of pDNA containing each target sequence. Results were normalized to GAPDH housekeeping gene expression levels. Standard curves showed high correlation coefficients and were linear over a wide range of copy numbers. Quantification of Aotus samples showed little intra- and inter-experiment variation, thus, the technique has proven to be highly reproducible and sensitive allowing us to detect as little as 25 copies/microl of target DNA. This technique will allow studying Th1 and Th2 cytokine patterns elicited in response to infection for prospectively evaluating the efficacy of malarial vaccines. PMID- 15271380 TI - SNP genotyping with FRET probes. Optimizing the resolution of heterozygotes. AB - Analysis of single nucleotide polymorphisms by PCR with fluorescence resonance energy transfer (FRET) probes often can produce a result where the melting peak corresponding to perfectly matched sequence (A allele) has a smaller area than the peak corresponding to the allele with a mismatch (B allele). This imbalance can make it difficult to distinguish heterozygous individuals from BB homozygotes. These results suggested that the higher strength in the binding of the perfect match probe to the A allele could cause the selective amplification of the B allele, possibly by interfering with the elongation of the PCR product. In order to optimize the detection of heterozygotes in allelic discrimination assays with FRET probes, we tested several modifications aimed at minimizing the apparent interference of the probes with the amplification process. We observed, in agreement with our hypothesis, that lowering the probe concentration or adding the probes after the amplification step more accurately resolved heterozygotes. PMID- 15271381 TI - Specific identification of Habronema microstoma and Habronema muscae (Spirurida, Habronematidae) by PCR using markers in ribosomal DNA. AB - Gastric or cutaneous habronemosis caused by Habronema microstoma Creplin, 1849 and Habronema muscae Carter, 1865 is a parasitic disease of equids transmitted by muscid flies. There is a paucity of information on the epidemiology of this disease, which is mainly due to limitations with diagnosis in the live animal and with the identification of the parasites in the intermediate hosts. To overcome such limitations, a molecular approach, based on the use of genetic markers in the second internal transcribed spacer (ITS-2) of ribosomal DNA, was established for the two species of Habronema. Characterisation of the ITS-2 revealed sequence lengths and G+C contents of 296 bp and 29.5% for H. microstoma, and of 334 bp and 35.9% for H. muscae, respectively. Exploiting the sequence difference (approximately 40%) between the two species of nematode, primers were designed and tested by the polymerase chain reaction (PCR) for their specificity using a panel of control DNA samples from common equid endoparasites, and from host tissues, faeces or muscid flies. Effective amplification from each of the two species of Habronema was achieved from as little as 10 pg of genomic DNA. Hence, this molecular approach allows the specific identification and differentiation of the DNA from H. microstoma and H. muscae, and could thus provide a molecular tool for the specific detection of Habronema DNA (irrespective of developmental stage) from faeces, skin and muscid fly samples. The establishment of this tool has important implications for the specific diagnosis of clinical cases of gastric and cutaneous habronemosis in equids, and for studying the ecology and epidemiology of the two species of Habronema. PMID- 15271382 TI - DNA microarray analysis of predominant human intestinal bacteria in fecal samples. AB - A microarray method was developed for the detection of 40 bacterial species reported in the literature to be predominant in the human gastrointestinal tract. The 40 species include seven species each of Bacteroides and Clostridium, six species of Ruminococcus, five species of Bifidobacterium, four species of Eubacterium, two species each of Fusobacterium, Lactobacillus and Enterococcus, and single species each of Collinsella, Eggerthella, Escherichia, Faecalibacterium and Finegoldia. Three 40-mer oligos specific for each bacterial species were designed based on comparison of the 16S rDNA sequences available in the GenBank database, and were used to make the DNA-array on epoxy slides. Using two universal primers, the 16S rRNA gene from bacteria present in fecal samples were amplified and labeled with Cyanine5-dCTP by PCR, and then hybridized to the DNA-array. After resolving some difficulties caused by sequence conflicts in GenBank and inaccurate reference strains, all 40 bacterial reference species gave positive results. The microarray method was used to screen fecal samples obtained from 11 healthy human volunteers for the presence of these intestinal bacteria. The results indicated that 25-37 of the 40 species could be detected in each fecal sample and that 33 of the species were found in a majority of the samples. PMID- 15271383 TI - A comparison of 14 antibodies for the biochemical detection of the cystic fibrosis transmembrane conductance regulator protein. AB - Interest in the biochemical detection of the cystic fibrosis (CF) transmembrane conductance regulator (CFTR) protein followed soon after cloning of the gene and prediction of the protein structure. Ever since, antibodies (Abs) have been produced and used to detect CFTR in both heterologously and endogenously expressing cells and tissues. Although designed to be sensitive and specific, these Abs produce, in most cases, unsatisfactory results when used for the biochemical detection of CFTR either by Western blot or by immunoprecipitation. The lack of Abs that can reliably detect the CFTR protein is a major constraint to studies of CF. We compared 14 different Abs for their ability to detect CFTR in both stably transfected and endogenously expressing cell lines. PMID- 15271384 TI - PCR-DIG ELISA with biotinylated primers is unsuitable for use in whole blood samples from patients with brucellosis. AB - In an attempt to avoid some of the inconveniences associated with conventional PCR, such as electrophoresis in ethidium bromide, we developed and analyzed the yield of a digoxigenin-based enzyme-linked immunosorbent PCR assay (PCR-DIG ELISA) for the detection of specific Brucella target DNA. During the DNA amplification process in healthy subjects and controls (Brucella abortus B-19) non-specific amplification of fragments was formed between genomic DNA and specific biotin-labeled primers. The labeled non-specific fragments bound to streptavidin-coated wells, saturating the solid phase streptavidin by biotin streptavidin interaction. The formation of these non-specific PCR products was demonstrated by reduction in absorbance with hemin, a Taq polymerase inhibitor, and identified by use of a silver stained method which improves the sensitivity of nucleic acid visualization. PMID- 15271385 TI - Unexpected detection of DNA by nucleic acid sequence-based amplification technique. AB - Nucleic acid sequence-based amplification (NASBA) is a technique that has been previously shown to selectively mediate the detection of RNA in microbial cells. In a series of tests, nucleic acids were extracted from Salmonella enterica serotype Typhimurium and Mycobacterium avium subsp. paratuberculosis, and subjected to four enzymatic treatments prior to NASBA. These enzymatic treatments were DNase, RNase, S1 nuclease, and RNase/S1 nuclease. The results obtained were different for the two bacteria. With S. enterica serotype Typhimurium, RNase and RNase/S1 nuclease abolished the NASBA signal, as expected. But with M. avium subsp. paratuberculosis RNase, S1 nuclease, and RNase/S1 nuclease had no effect on the NASBA signal, whereas DNase treatment abolished it. This indicates that in the latter bacterium, NASBA can detect DNA, and demonstrates the necessity of verifying the nucleic acid origin of a NASBA signal if detection of RNA is objective. PMID- 15271386 TI - Identification of ciprofloxacin-resistant Campylobacter jejuni and analysis of the gyrA gene by the LightCycler mutation assay. AB - A real-time PCR assay was developed to identify ciprofloxacin-resistant Campylobacter jejuni. Ciprofloxacin resistance in C. jejuni has been associated with a C-->T nucleotide point mutation occurring at the 86 codon of the gyrA gene. Other nucleotide substitutions have been identified in proximity to or at the same codon in the gyrA gene, but their role in ciprofloxacin resistance is still unknown. The LightCycler assay is based on the fluorescence resonance energy transfer technology using melting peak analysis of two fluorescent probes hybridized on PCR amplicons. This assay was used to detect the 86-codon mutation conferring ciprofloxacin resistance, as well as other nucleotides substitutions occurring within the same site in the gyrA gene. This gyrA mutation assay allows a rapid and reproducible screening method of ciprofloxacin resistant strains and was applied to C. jejuni strains isolated in Italy in 2000. PMID- 15271387 TI - Amplification of Echoviruses genomic regions by different RT-PCR protocols--a comparative study. AB - In the present report, the results of a comparative study in the detection of all Echoviruses reference strains as well as of 38 clinical isolates are presented. Using RT-PCR with already published primer pairs (UG(52)-UC(53), 292-222, 012-011 and EUG2a, 2b, 2c-EUC2) from the 5'UTR, the VP1 region as well as a long genomic fragment including the VP1 3' end, the entire coding sequence of 2A, 2B, and the 5' moiety of the 2C-coding region amplification was effective with all reference and clinical Echovirus isolates with primer pair UG(52)-UC(53) while with 292-222 and 012-011 were amplified 27/28 reference Echovirus strains and all clinical isolates. As far as EUG2a,2b,2c-EUC2 is concerned, the RT-PCR gave a positive result for 26/28 reference Echovirus strains and 34/38 clinical isolates. The sequence analysis of a large part of the 5'UTR has revealed that there is no correlation between 5'UTR identity and the currently recognized human enterovirus species. It has been suggested that part of VP1 coding sequence would correlate well with serotype since a number of important neutralization epitopes, as well as receptor recognition sequences, lie within the VP1 coding sequence. Therefore, UG(52)-UC(53) and 292-222 primer pairs seem to be the most appropriate for Echovirus detection and, moreover, UG(52)-UC(53) is useful for the classification of enteroviruses into genetic clusters (sub-groups) while 292-222 for the identification of enteroviruses by amplicon sequencing. PMID- 15271388 TI - Cloning and expression of Clostridium difficile toxin A gene (tcdA) by PCR amplification and use of an expression vector. AB - Toxigenic Clostridium difficile isolates harbor a 19 kb pathogenicity locus that encodes the genes for toxins A and B. Toxins A and B are among the largest known bacterial toxins expressing potent cytotoxicity and enterotoxicity, and thus the major virulence factors in C. difficile associated diarrhea. Cloning and sequencing of toxin genes is of interest for studies of molecular pathogenesis. We report the amplification and cloning of the complete toxin A gene into an Escherichia coli expression vector. Ten clones analyzed contained the complete toxin A gene. Four of these clones showed cytotoxic activity in cell culture, and were positive for toxin A as determined by ELISA. Toxin A expression was confirmed by Western immunoblot analysis. The presence of cytotoxic activity in cell culture suggests that toxin A activity is independent of other genes in the pathogenicity locus. PMID- 15271389 TI - A real-time multiplexed PCR assay for rapid detection and differentiation of Campylobacter jejuni and Campylobacter coli. AB - Campylobacter species are the leading agents of bacterial gastroenteritis worldwide. C. jejuni and C. coli together are responsible for more than 95% of all cases of Campylobacter-induced diarrheal disease in the United States. Detection of campylobacteria in clinical samples by conventional culture is problematic and slow due to their complex taxonomy, fastidious growth requirements, and biochemical inertness. The current study describes a rapid, sensitive, and specific real-time polymerase chain reaction (PCR) assay capable of detecting and differentiating C. jejuni (hippuricase gene, hipO) and C. coli (serine hydroxymethyltransferase gene, glyA) in a single reaction, directly from clinical isolates and human feces. The analytical specificity of the assay was demonstrated with a diverse range of Campylobacter species, related organisms, and other diarrhea-inducing bacterial pathogens. The analytical sensitivity of the multiplexed, PCR assay was 10 genome copies for both C. jejuni and C. coli. Following a rapid DNA extraction method (QIAGEN QIAamp DNA stool Mini Kit), the multiplexed PCR identified C. jejuni or C. coli in 100% of fecal samples containing 10(3) colony-forming units (CFU) per gram of feces. This assay represents the first real-time PCR method capable of detecting and differentiating C. jejuni and C. coli in a single reaction. PMID- 15271390 TI - Development of a multiplex PCR assay for the identification of pathogenic genes of Escherichia coli in milk and milk products. AB - A multiplex PCR for the simultaneous detection of some pathogenic genes of enteropathogenic, enterotoxigenic and verocytotoxin-producing Escherichia coli was developed. In this study primers found in literature as well as primers to the purpose designed were used. In this way, it was possible to generate specific fragments of 96, 170, 229, 285, 348, 414 and 510 bp for Hlya, St, EaeA, Lt, Vt1, UidA and Vt2 genes, respectively. When applied to bacterial strains experimentally inoculated in milk and milk products, the proposed PCR showed a detection limit of 5 x 10(4)CFU/ml for Hyla, St, Eaea, Vt1 primers, while for Lt and Vt2 primers the limit resulted of 10(6)CFU/ml. PMID- 15271391 TI - Animal models of chronic myelogenous leukemia. AB - Of the current mouse chronic myelogenous leukemia (CML) models,the murine bone marrow (BM) transduction and transplantation model most efficiently mimics many of the central features of human CML. In this model, lethally irradiated mice are reconstituted with primary murine BM cells transduced with a P210BCR/ABL retrovirus. All recipient mice develop a fatal peripheral blood and BM granulocytosis and splenomegaly, a disease termed the murine CML-like myeloproliferative disorder. This model has been used to establish the causative role of Bcr/Abl in CML, identify those signaling pathways and regions of Bcr/Abl critical for leukemogenesis, and explore the limitations of targeted CML therapy. Future refinements in this CML mouse model will make it a more effective tool for studying imatinib-resistant CML, reproducing chronic- and blastic-phase human CML, and performing CML progenitor studies. PMID- 15271392 TI - Biology of chronic myelogenous leukemia--signaling pathways of initiation and transformation. AB - Chronic myeloid leukemia (CML) is caused by the Bcr-Abl oncoprotein,the product of the t(9;22) chromosomal translocation that generates the Philadelphia chromosome. Different disease phenotypes are associated with each of the three Bcr-Abl isoforms: p190Bcr-Abl, p210Bcr-Abl, and p230Bcr-Abl all of which have a constitutively activated tyrosine kinase. Mechanisms associated with malignant transformation include altered cellular adhesion, activation of mitogenic signaling pathways, inhibition of apoptosis, and proteasomal degradation of physiologically important cellular proteins.CML is subject to an inexorable progression from an "indolent" chronic phase to a terminal blast crisis. Disease progression is presumed to be associated with the phenomenon of genomic instability. PMID- 15271393 TI - Natural history and staging of chronic myelogenous leukemia. AB - The natural history of chronic myelogenous leukemia (CML) has changed in recent years, partly due to earlier diagnosis but mostly as a consequence of the availability of effective therapies that have the potential to eradicate the Philadelphia chromosome-positive clone. Highly effective therapy with imatinib has changed the prognostic significance of clinical features traditionally associated with poor outcome. Achieving a complete cytogenetic response and a major molecular response early during the course of therapy with imatinib may be the most important factor in determining longterm outcome. Therefore, treatment modalities that increase the probability of achieving this goal should be pursued. This article describes the natural history of CML and its prognostic factors,with emphasis on changes due to the emergence of imatinib. PMID- 15271394 TI - Interferon therapy in chronic myelogenous leukemia. AB - Interferon (IFN)-alpha, the molecule used in the treatment of chronic myelogenous leukemia and initially prepared from human leucocytes,is now produced essentially by recombinant techniques. Polyethylene glycol (PEG) modifications of proteins could be more effective than the regular molecules; thus, pegylated IFNs more recently have been tested in chronic myelogenous leukemia. PEG modification of proteins reduces sensitivity to proteolysis. Moreover,administration of pegylated IFNs results in less antigenicity and immunogenicity, and prolongation of their plasma half-life has been assessed by pharmacokinetic studies. It is assumed, therefore, that this compound could be more effective and better tolerated. Given the results recently obtained with imatinib, however, whether IFN-alpha will still have a therapeutic role is questionable. PMID- 15271395 TI - Imatinib therapy in chronic myeloid leukemia. AB - This article briefly recaps the key clinical trials that involve imatinib and focuses on the ongoing results and implications of these studies for future research in imatinib-based therapy in chronic myeloid leukemia (CML). Imatinib by no means has replaced allografting in the treatment of CML, although clearly, there has been a radical shift in thinking in considering which patients should undergo transplant. The route to cure with minimal toxicity may involve creative use of drug and transplant technologies. PMID- 15271396 TI - Investigational strategies in chronic myelogenous leukemia. AB - Imatinib is the cornerstone of therapy in chronic myelogenous leukemia (CML) and a model for the development of novel agents directed at specific targets. The results of imatinib therapy continue to improve with approaches such as higher doses of imatinib and, possibly, with combinations of imatinib and interferon alpha with or without cytarabine. There are multiple targets with agents directed to them that may prove to be synergistic with imatinib. These approaches are attractive, particularly when dealing with imatinib resistant CML, to prevent resistance and improve the probability of cure. The continued understanding of the biology of CML and mechanisms of resistance to imatinib and the ability to develop target-specific therapies should lead to the increased probability of cure for most patients who have CML. PMID- 15271397 TI - Clinical resistance to imatinib: mechanisms and implications. AB - Although responses to imatinib in chronic phase chronic myelogenous leukemia have been durable in most cases, most patients in advanced disease develop resistance and relapse after a short duration of therapy. The mechanisms of drug resistance are diverse, but in most cases, mutations are found at the time of resistance that change amino acids within the kinase domain of BCR-ABL. Cytogenetic and molecular analyses at the time of resistance are suggested to guide therapy. PMID- 15271398 TI - Monitoring of minimal residual disease in chronic myeloid leukemia. AB - Detection and monitoring of minimal residual disease has become one of the most prevalent topics in chronic myeloid leukemia(CML) therapy. The goal of early detection of residual disease is to allow timely therapeutic intervention before overt relapse of therapy resistant disease occurs. The most powerful tool to serve this purpose is polymerase chain reaction (PCR). Major improvements in assay techniques have advanced PCR from a purely qualitative test with considerable variability of test results to a real-time quantitative assay with far more reproducible results than were possible before. At the same time, treatment of CML has changed dramatically since the introduction of imatinib. Integration of therapy and molecular assays such as PCR, in addition to a profound understanding of the pathophysiology of CML, has assumed even more importance. Quantitative PCR testing has become the standard monitoring strategy for patients undergoing stem cell transplantation. Although correlations have been established between positive test results and probability of relapse, no absolute guidelines for monitoring exist, especially for patients treated with imatinib. PMID- 15271399 TI - Clonal evolution in chronic myelogenous leukemia. AB - Clonal evolution (CE) may be a marker of disease progression in chronic myelogenous leukemia (CML) and is thought to reflect the genetic instability of the highly proliferative CML progenitors. The frequency of CE increases with advancing stage, rising from 30%in accelerated phase and up to 80% in blast crisis. Given its association with disease progression, CE is considered a feature that defines accelerated-phase CML; however, not all studies have demonstrated a uniformly poor outcome for patients with CE. Chromosomal abnormalities in Ph chromosome negative metaphases increasingly have been recognized in patients treated with imatinib. The true incidence of this phenomenon is not clear but appears to occur in 2% to 17% of imatinib-treated patients. Regardless of the precise mechanism and the long-term clinical implications, the findings described in this article underscore the importance of routine cytogenetic analysis for patients treated with imatinib. The continued study of these phenomena will help us to understand better the pathogenesis of CML and improve the long-term treatment of patients with CML. PMID- 15271400 TI - Allogeneic hematopoietic stem cell transplantation for chronic myeloid leukemia. AB - There never has been a more difficult time to advise patients newly diagnosed with chronic myeloid leukemia (CML). Until recently,the options comprised noncurative but relatively nontoxic chemotherapy or potentially curative allogeneic stem cell transplantation,with its attendant morbidity and mortality. There now is the additional option of the tyrosine kinase inhibitor imatinib mesylate that has been in clinical practice for almost 5 years. Although data are emerging regarding the efficacy of imatinib, solid evidence of any prolongation in survival will be delayed for several years. Future management of CML will continue to depend on a combination of approaches that use allografting and targeted drug therapy. The next decade undoubtedly will witness the introduction of a number of agents capable of inhibiting signal transduction pathways,perhaps controlling CML in cases of primary or acquired imatinib resistance. In the absence of long-term outcome data for imatinib,it remains reasonable to propose that young patients with newly diagnosed CML who have HLA-identical or well matched unrelated donors should undergo allogeneic transplantation using a standard or nonmyeloablative conditioning regimen. Similarly,patients who fail to respond to imatinib should be rescued with a transplant strategy. The role of molecular monitoring in these two strategies cannot be underestimated. PMID- 15271401 TI - Nonmyeloablative stem cell transplantation for chronic myeloid leukemia. AB - Conventional myeloablative hematopoietic stem cell transplantation carries risks of morbidity and mortality from regimen-related toxicities that have restricted its use to relatively young patients in good medical condition. In nonmyeloablative allogeneic hematopoietic stem cell transplantation, enhanced immunosuppression rather than myeloablative cytotoxic conditioning has allowed the engraftment of allogeneic stem cells, with lower early transplant related mortality and morbidity. This approach shifts tumor eradication to the graft versus-leukemia effect mediated by donor T lymphocytes. The development of nonmyeloablative transplantation has allowed the application of a potentially curative procedure to elderly or medically infirm patients who would not be able to tolerate high-dose conditioning regimens. PMID- 15271402 TI - Autologous hematopoietic cell transplantation for chronic myelogenous leukemia. AB - Experimental and clinical evidence for persistence of polyclonal Philadelphia chromosome negative (Ph-) progenitors in chronic myelogenous leukemia (CML) patients has provided the rationale for autologous transplantation. Clinical trials of autologous transplantation suggest that this procedure can induce cytogenetic remissions in a subset of patients and may be associated with longer than-expected patient survival. Most autologous transplant recipients, however, continue to have evidence of persistent leukemia. Recent reports indicating that it is possible to collect sufficient numbers of Ph- peripheral blood stem cells for autologous transplantation from most patients in complete cytogenetic remission on imatinib treatment have rekindled interest in autologous transplantation in CML. Additional approaches to eliminate residual disease in autografts and to sustain cytogenetic response after transplantation, however, will be required to achieve long-term restoration of Ph- hematopoiesis. Several promising methods to improve purging of the autograft and for more effective elimination of residual leukemia are being explored. PMID- 15271403 TI - Immunity to chronic myelogenous leukemia. AB - Given the high rate of cytogenetic responses to imatinib mesylate in chronic myelogenous leukemia (CML), logical future treatment strategies will include combinations of tyrosine kinase inhibitors and immunotherapies such as vaccines. Increased understanding of highly specific immune responses will lead to novel and improved immunotherapy strategies for CML patients. Such advances can be expected to revolutionize the field much in the same way that imatinib mesylate and other targeted small molecules have revolutionized our conception of traditional chemotherapy. This article begins with a brief discussion of why CML may represent a model disease for immunotherapy-based strategies. Laboratory evidence of the immunoresponsiveness of CML is discussed and used to highlight the principles for understanding tumor immunity. Finally,the authors discuss how advances in the understanding of the molecular and cellular nature of immunity are being translated into new therapeutic strategies for the treatment of CML. PMID- 15271404 TI - Accelerated and blastic phases of chronic myelogenous leukemia. AB - Chronic myelogenous leukemia (CML) may have a biphasic or triphasic course, whereby patients who were initially diagnosed in the chronic phase (CP) develop more aggressive disease, frequently pass through an intermediate or accelerated phase (AP), and finally evolve into an acute leukemia like blastic phase (BP). A slowing in the rate of development of AP or BP has accompanied successive improvements in therapy for patients who have CP CML. Variable diagnostic criteria for AP and BP are used in the literature, rendering comparisons difficult. The management of patients in AP or BP consistently has been less effective than the management of those inCP for all modalities of therapy. This article reviews the current diagnostic criteria, therapeutic strategies, outcomes, and investigational therapies for AP and BP CML. PMID- 15271406 TI - Executive dysfunction and visuospatial ability among depressed elders in a community setting. AB - Visuospatial ability is frequently compromised among elderly depressed patients, but it is unclear whether the impairment is a consequence of a visuospatial memory deficit or of an executive dysfunction that impacts visuospatial ability. The Boston Qualitative Scoring System is a method of scoring the Rey-Osterrieth Complex Figure (ROCF) that assesses the process used to draw the figure, the executive aspect of the task, as well as the accuracy and location of the completed elements. The hypotheses that executive scores as measured by the BQSS would separate diagnostic groups and that executive function would mediate the relationship between depression and nonverbal recall were tested using a between groups design with elderly depressed volunteers (N = 31) and healthy controls (N = 31). Participants were screened for other Axis I disorders with the Structured Clinical Interview for DSM-IV Diagnosis, diagnosed for major depression per DSM IV criteria, and administered the ROCF. The copy and recall drawings were scored using BQSS criteria, and scores were grouped into executive and drawing scores from both copy and recall phases. Executive scores during the copy phase and drawing scores from the recall phase separated the diagnostic groups [F(1,59), = 4.14, P = .05] and [F(1,59) = 6.88, P = .01], respectively. Follow-up ANCOVAS showed that copy Planning, the score that quantified the process by which the figure was drawn, separated the diagnostic groups. Planning also mediated the association between depression and the percent of the figure recalled after the short delay (Z = 1.84, P < .05). The significance of the depression-to-recall pathway was eliminated when Planning was controlled for, but Planning remained related to percent recalled [B = -6.90, P < .007]. A dimension of executive dysfunction, represented here by Planning, may be one underlying source of the observed decline in nonverbal recall among elderly depressed patients. This result is consistent with the theory that dysfunction of the prefrontal cortex is a critical feature of late-life depression. PMID- 15271407 TI - The construct of problem solving in higher level neuropsychological assessment and rehabilitation. AB - Three inter-related studies examine the construct of problem solving as it relates to the assessment of deficits in higher level outpatients with traumatic brain injury (TBI). Sixty-one persons with TBI and 58 uninjured participants completed measures of problem solving and conceptually related constructs, which included neuropsychological tests, self-report inventories, and roleplayed scenarios. In Study I, TBI and control groups performed with no significant differences on measures of memory, reasoning, and executive function, but medium to large between-group differences were found on timed attention tasks. The largest between-group differences were found on psychosocial and problem-solving self-report inventories. In Study II, significant-other (SO) ratings of patient functioning were consistent with patient self-report, and for both self-report and SO ratings of patient problem solving, there was a theoretically meaningful pattern of correlations with timed attention tasks. In Study III, a combination of self-report inventories that accurately distinguished between participants with and without TBI, even when cognitive tests scores were in the normal range, was determined. The findings reflect intrinsic differences in measurement approaches to the construct of problem solving and suggest the importance of using a multidimensional approach to assessment. PMID- 15271408 TI - Validation of the Meyers Short Battery on mild TBI patients. AB - This manuscript reports the results of two studies focusing on patients with mild Traumatic Brain Injury (TBI). The first assesses the validity of the Meyers Short Battery (MSB) of neuropsychological tests. The second study reports on the reliability of the MSB. The groups consisted of normal controls, depressed, chronic pain patients, and patients with mild TBI. Validity was assessed using a discriminant function analysis comparing the non-TBI participants with the TBI participants, which showed a 96.1% correct classification rate. When patients were assessed at least 6 months post-injury and re-assessment 12-14 months later, an overall reliability of r = .86 was obtained. This indicates that the MSB has adequate psychometric properties for clinical use. The results are consistent with previous published research indicating that the MSB is sensitive not only to the presence of mild TBI but also to the degree of cognitive impairment based on loss of consciousness. PMID- 15271409 TI - Relationship between plasticity, mild cognitive impairment and cognitive decline. AB - A topic of great interest in gerontology research is the prediction of cognitive deterioration which marks the transition from mild cognitive impairment (MCI) to dementia. In this area the term plasticity is a construct of prime importance. Previous studies have demonstrated the existence of plasticity in healthy older persons, and it is thought that this is what discriminates between healthy individuals and those at risk for dementia. The aim of the present study is to demonstrate that plasticity exists in persons with MCI, and that a lack of plasticity may be one of the risk factors related to cognitive decline. An adapted version of the Auditory Verbal Learning Test-the AVLT of Learning Potential-was used to assess plasticity. Participants in the research were 203 older persons whose cognitive status had previously been determined using a cognitive screening test. The results show that plasticity exists in persons with MCI and that its presence is associated with less marked cognitive decline. PMID- 15271410 TI - The Tower of London and neuropsychological assessment of ADHD in adults. AB - Executive function refers to a variety of behaviors and abilities related to planning and strategy use, as well as the maintenance of attention and behavior in the pursuit of some goal; these behaviors are generally deficient in individuals with Attention Deficit Hyperactivity Disorder (ADHD). The Tower of London (TOL) is one task used in the assessment of executive function. For adults with ADHD, there is minimal research on the extent to which they demonstrate impaired performance on tower tasks. With a sample of 102 individuals between the ages of 16 and 33 years, the extent to which performance on the TOL-Drexel Edition (TOL(DX)) was related to performance on other measures of executive function and diagnostic grouping was investigated. Results indicated that TOL(DX) variables are not correlated significantly with age or Global Assessment of Functioning (GAF). Of the TOL(DX) variables, only Rule Violations correlated with multiple other executive function variables. Rule Violations correlated minimally, but significantly, with cognitive ability, perceptual skills, Matrix Reasoning, Processing Speed, and immediate memory. As might be expected, Processing Speed also significantly correlated with Total Time and Time Violations. Notably, scores on the TOL(DX) did not correlate significantly with behavioral self-report; no between-group (ADHD, Clinical Control, No Diagnosis) differences emerged for any of the TOL(DX) variables. Further, with this sample, mean scores across the TOL(DX) variables were well within the average range. Taken together, these results suggest that while the TOL(DX) measures aspects of ability not tapped by other measures, and may therefore provide additional information on individual functioning, results should not be interpreted as indicative of the presence or absence of a disorder. PMID- 15271411 TI - Predicting competency in automated machine use in an acquired brain injury population using neuropsychological measures. AB - The purpose of the current study was to explore whether performance on standardised neuropsychological measures could predict functional ability with automated machines and services among people with an acquired brain injury (ABI). Participants were 45 individuals who met the criteria for mild, moderate or severe ABI and 15 control participants matched on demographic variables including age- and education. Each participant was required to complete a battery of neuropsychological tests, as well as performing three automated service delivery tasks: a transport automated ticketing machine, an automated teller machine (ATM) and an automated telephone service. The results showed consistently high relationship between the neuropsychological measures, both as single predictors and in combination, and level of competency with the automated machines. Automated machines are part of a relatively new phenomena in service delivery and offer an ecologically valid functional measure of performance that represents a true indication of functional disability. PMID- 15271412 TI - Detecting symptom- and test-coached simulators with the test of memory malingering. AB - The ability of the Test of Memory Malingering (TOMM; Tombaugh, 1996) to detect feigned-memory impairment was explored. The TOMM was administered to three groups: (a) a control group instructed to perform optimally, (b) a symptom coached group instructed to feign memory problems after being educated about traumatic brain injury symptomatology, and (c) a test-coached group instructed to feign memory problems after being educated about test-taking strategies to avoid detection. The recommended cutoff scores (Tombaugh, 1996) on Trial 2 and the Retention Trial produced overall classification accuracy rates of 96%, with high levels of sensitivity and specificity. Although the symptom-coached group performed more poorly on the TOMM relative to the test-coached group, the test was equally sensitive in detecting suboptimal effort across the different coaching paradigms. PMID- 15271413 TI - RET oncogene mutations in 75 cases of familial medullary thyroid carcinoma in Japan. AB - The familial form of medullary thyroid carcinoma (MTC) is caused by mutations of the RET protooncogene. We registered 60 multiple endocrine neoplasia (MEN) 2A patients, 12 familial non-MEN medullary carcinoma (FMTC) patients, and three MEN2B patients with a confirmed RET germline mutation. All 60 MEN2A patients had RET mutations in a cysteine-rich domain. Seven of the FMTC patients had a mutation in cysteine-rich domain, and the other five had a mutation in codon 768, which encodes a tyrosine-kinase domain. Two of the MEN2B patients had a mutation in codon 918, and one patient had a double mutation, one in codon 804 and the other in codon 806, both of which are all encoded tyrosine-kinase domain. The genotype-phenotype correlations of our data will allow individualized recommendations for the optimal timing of prophylactic surgery. PMID- 15271414 TI - Clinical manifestations of familial medullary thyroid carcinoma. AB - We conducted a large-scale nation-wide questionnaire survey to ascertain the status of familial medullary thyroid carcinoma (MTC) in Japan in 2002. Out of a total of 271 MTC cases (male to female ratio 1:1.4), multiple endocrine neoplasia (MEN) 2A accounted for 83 cases (30.6%), familial MTC (FMTC) for 14 cases (5.1%), MEN for 11 cases (4.1%), and sporadic MTC for 163 cases (60.1%). Mean age at the time of diagnosis was 35.6 in MEN2A, 34.6 in FMTC, 30.5 in MEN2B, and 47.6 in sporadic MTC. Forty-five percent of MEN2A patients had pheochromocytoma and 11% of MEN2A patients had parathyroid disorders when MTC was diagnosed. Finally, the RET oncogene test yielded the largest number of initial findings that led to diagnosis of familial MTC. PMID- 15271415 TI - Large needle aspiration biopsy results of palpable thyroid nodules diagnosed by fine-needle aspiration as a microfollicular nodule with atypical cells or suspected cancer. AB - Among 1875 patients with palpable thyroid nodules examined with FNA cytology and LNAB histology, 132 with a preoperative FNA diagnosis of microfollicular nodule with atypical cells (n = 50) or suspected cancer (n = 82) were operated on. The 50 nodules showed the following preoperative LNAB finding: inadequate (8), benign (15), microfollicular (20), microfollicular with atypical cells (5), suspected cancer (2). The postoperative cancer incidence in the nodules with the benign LNAB diagnosis was 0% while it was 10%, 60% (P = 0.008), 100% (P = 0.007) in the other three LNAB diagnostic categories. The 82 nodules showed the following preoperative LNAB finding: inadequate (21), benign (21), microfollicular (15), microfollicular with atypical cells (15), suspected cancer (10). The postoperative incidence of cancer in the 21 (14%) and 10 (80%) nodules diagnosed by LNAB as benign nodule or suspected cancer, respectively, was significantly different (P = 0.0007). These data suggest that LNAB can be used for the preoperative selection of the palpable thyroid nodules diagnosed by FNA as a microfollicular nodule with atypical cells or suspected cancer. PMID- 15271416 TI - From the bench to the bedside. Galectin-3 immunodetection for improving the preoperative diagnosis of the follicular thyroid nodules. AB - The authors discuss the principal aspects concerning the preoperative characterization of thyroid nodules, in particular those with follicular histology, and illustrate the potential clinical impact of a new diagnostic test method, named "galectin-3 thyrotest", which is based on the immunodetection of galectin-3 molecule in cytological specimens derived from thyroid nodular lesions. This diagnostic test method, which consistently improves the accuracy of conventional cytology, has been recently validated in a large international multicenter study and is going to impact hardly the clinical management of patients bearing thyroid nodular diseases. The rationale of this new diagnostic approach, the possibility to improve its performance in selected cases by using large needle aspiration biopsy (LNAB) together with technical and operative details are presented and discussed. PMID- 15271417 TI - Radiological and surgical management of thyroid neoplasms. AB - Recent advances in the radiological diagnosis in thyroid neoplasms have been achieved by high-resolution ultrasonography and color-Doppler, and the ultrasound guided fine-needle aspiration biopsy and ultrasound-guided percutaneous ethanol injection therapy have been developed on the basis of these modalities. Ultrasonography and ultrasound-guided fine-needle aspiration biopsy have made minimally invasive thyroid surgery possible. The surgical procedures are classified into three main categories according to the approach, and each approach has its own advantages and disadvantages. Surgeons have to select the most suitable approach from one of these categories of approaches for each patient with a thyroid neoplasm. PMID- 15271418 TI - Nuclear factor kappa B--molecular biomedicine: the next generation. AB - Nuclear factor kappa B (NFkappaB) as a transcription factor plays an important integrating role in the intracellular regulation of immune response, inflammation and cell cycle regulation. Nouvelle insights into the structure and regulation of activation of NFkappaB have brought a detailed picture of the function of this transcription factor. In this review the findings of interactions of NFkappaB with its inhibitors, tumour necrosis factor alpha and glucocorticoids are presented. The results from the latest in vivo studies show the capability of specific NFkappaB inhibitors in the clinical use. This article summarizes the most important facts regarding NFkappaB participation in the pathogenesis of diseases and its potential as a target of pharmacological agents. PMID- 15271419 TI - Tissue distribution of the low molecular weight heparin, tinzaparin, following administration to rats by the oral route. AB - Heparins are antithrombotic drugs given by intravenous and subcutaneous routes. However, we have observed that heparins have antithrombotic activity in a rat model when administered orally despite low plasma levels, with low molecular weight heparins (LMWHs) being effective at lower single doses than unfractionated heparins (UFH). Since LMWHs may have other pharmaceutical uses and little is known regarding the pharmacokinetics of oral LMWHs, our objectives were to determine the distribution of the LMWH tinzaparin (Logiparin) following oral dosing. To study distribution at different doses, 0.025-15 mg/kg tinzaparin was given by stomach tube to rats. Gut and non-gut tissues were sampled 4 h later. In a time course study, plasma and tissue samples were collected at eight time points within 24 h after oral administration (60 mg/kg, 4 rats/time interval). Accumulated urine and faeces were collected over 4 and 24 h using metabolic cages. Gut tissue and washes, faeces, urine and non-gut tissue were extracted and analysed for heparin by agarose gel electrophoresis with toluidine blue staining. Activated partial thromboplastin time (APTT) and anti-Xa activity, by Heptest and chromogenic assay, estimated plasma tinzaparin concentrations. Stomach and lung tinzaparin concentrations demonstrated a dose-effect. Peak concentrations in tissue and washes of stomach, duodenum, jejunum, ileum and colon were at 6-30, 15 30, 30 min, 2 and 4 h, respectively. Amounts found at peak times in combined tissue and washes accounted for 46% and 0.5% in stomach (15 min) and colon (4 h), respectively. Tinzaparin was recovered from liver, lung, endothelial samples, and urine at 24 h, but not in faeces. Non-significant increases were seen in APTT and the Heptest, however, anti-Xa activity was significantly greater than control at all times examined, peaking at 2 h. No bleeding was observed. Results are consistent with oral absorption of tinzaparin with wide tissue distribution, likely on endothelium with little in plasma, as previously observed for UFH. Oral administration of LMWHs should be further studied. PMID- 15271420 TI - Down-regulation of lipoxin A4 receptor by thromboxane A2 signaling in RAW246.7 cells in vitro and bleomycin-induced lung fibrosis in vivo. AB - Lipoxins (LXs) are members of eicosanoid family that can be endogenously produced during cell-to-cell interactions such as platelet-leukocyte interactions. Anti inflammatory function of lipoxin A4 (LXA4) as "braking signals" is mediated by the receptor. On the other hand, thromboxane A2 (TXA2) produced by catalysis of cyclooxygenase and thromboxane synthetase is released during platelet aggregation as a vasoconstrictor and a pro-inflammatory factor. To investigate interaction of TXA2 receptor (TP) and LXA4 receptor, effects of a TP agonist and a thromboxane synthetase inhibitor on expression of LXA4 receptor were examined in vitro and in vivo. A TP agonist, U46619 showed a down-regulation of LXA4 receptor induced by interleukin-1beta (IL-1beta) in RAW246.7 cells. In bleomycin-induced lung fibrosis in mice, administration of a thromboxane synthetase inhibitor DP-1904 increased LXA4 receptor mRNA and decreased type I collagen mRNA. In vitro experiments indicate that LXA4 significantly prevented enhanced proliferation of NIH3T3 fibroblasts and the collagen expression by transforming growth factor-beta (TGF-beta). These results suggest that TXA2-TP signaling could cause negative regulation of lipoxin A4 receptor under the transcriptional level during inflammatory process mediated by IL-1beta and TGF-beta induce the expression of LXA4 receptor. Furthermore, the down-regulation of LXA4 receptor by TXA2 implies a possibility that a cellular signaling by TXA2 may have a novel and potential function as a pro-inflammatory factor to inhibit anti-inflammatory effect of LXA4. Concomitantly, selective blockade of TXA2-TP signaling could be suggested to lead to anti-inflammation through active role of LXA4. PMID- 15271421 TI - In vitro antileishmanial activity of acetogenins from Annonaceae. AB - Twelve acetogenins from Annonaceae were evaluated in vitro for their antileishmanial activities in order to search for new lead-compounds having antileishmanial properties. The compounds were comparatively evaluated by the 50% inhibitory concentrations (IC50) determination on promastigote forms of wild-type and four drug-resistant lines of Leishmania donovani. In addition, after testing the toxicity on mouse peritoneal macrophages, the compounds were evaluated on amastigote infected macrophages and a therapeutic index was calculated. The IC50 of the acetogenins against promastigote forms of L. donovani was in a range 4.7 47.3 microM. The most active compound was Rolliniastatin 1 (IC50 at 4.7 microM). On the intramacrophage amastigote in vitro model, only seven compounds exhibited measurable antileishmanial activity with IC50 values in a range 2.5-29.7 microM. Rollinistatin 1 was the most interesting compound with IC50 of 2.5 microM and it appears as the most promising one on the basis of therapeutic index (18.08). Isoannonacin, which is active against intramacrophagic amastigotes (IC50 of 6.2 microM) with a therapeutic index of 2.05, exhibited a strong action on drug resistant strains (IC50 from 5.1 to 9.8 microM). Acetogenins are a new chemical series with interesting in vitro antileishmanial activity and further studies will be focused on the understanding of this selectivity in regard to the membrane and mitochondrial action using specific probes. PMID- 15271422 TI - Linear furanocoumarin protects rat myocardium against lipidperoxidation and membrane damage during experimental myocardial injury. AB - The antioxidant activity and the membrane effects of linear furanocoumarin marmesinin isolated from Aegle marmelose was evaluated during experimental myocardial injury. Isoproterenol (150 mg kg(-1) intraperitonially twice at an interval of 24 h) caused increase in the levels of serum marker enzymes via creatinekinase (CK), creatinekinase-MB (CK-MB) isoenzyme, lactatedehydrogenase (LDH) and lactatedehydrogenase isoenzyme (LDH1). It also produced electrocardiographic changes such as increased heart rate, reduced R amplitude and ST elevation. Marmesinin at a dose of 200 mg kg(-1), when administered orally, demonstrated a decrease in serum enzyme levels and restored the electrocardiographic changes towards normalcy. Myocardial injury was accompanied by the disintegration of lipidperoxides and the impairment of natural scavengers. Marmesinin oral treatment for 2 days before and during isoproterenol administration decreased the effect of lipidperoxidation. It was also shown to have a membrane stabilizing action by inhibiting the release of beta glucuronidase from the subcellular fractions. Thus, linear furanocoumarin marmesinin could have the protective effect against the damage caused by experimental myocardial injury. PMID- 15271423 TI - Ornithine decarboxylase, polyamines and CD11b expression in HL-60 cells during differentiation induced by retinoic acid. AB - Polyamines (PA) and retinoic acid affect mammalian cell growth, differentiation and apoptosis. Retinoic acid induces granulocytic differentiation of mieloid cell lines and, during this process, is responsible for the expression of CD11b, a surface antigen. In this study we investigate the effects of retinoic acid on HL 60 cells, monitoring ornithine decarboxylase (ODC) activity (enzyme rate of PA), putrescine (PUT), spermidine (SPD), spermine (SPM) levels, CD11b myeloid surface marker differentiation, cell cycle, and apoptosis. ODC activity and PUT levels are correlated with mieloid cell differentiation induced by retinoic acid treatment. Only the ODC/PUT ratio is connected with retinoic acid treated HL-60 cells. Treated cultures show a decrease of proliferation and a cell block in the G0/G1 phase, with consequent diminished S phase. The G0/G1 and S phases are significantly related to ODC activity and to PUT and SPD behavior, whereas in differentiating condition only the decrease of PUT is related to the S phase. CD11b expression, stimulated by retinoic acid treatment, is associated with the SPM trend. Total PA behavior agrees with apoptotic cell increase after 96 h of stimulation. Our data show that retinoic acid treatment modifies ODC activity and the turnover of PA. PUT, SPD and SPM, therefore, have a different role, and may be involved in the differentiative/apoptotic program of retinoic acid treated HL 60 cells. PMID- 15271424 TI - Equine ocular anatomy and ophthalmic examination. AB - This article is intended to provide the practitioner with a succinct but complete source regarding equine orbital and ocular anatomy,instrumentation available for ophthalmic examination, a methodical examination technique, sedation and regional nerve blocks, and diagnostic procedures involving the eye. Such knowledge of orbital and ocular anatomy is essential to allow recognition of normal,normal variations, or an abnormality of the equine eye and orbit. PMID- 15271425 TI - The precorneal tear film in horses: its importance and disorders. AB - The precorneal tear film (PTF) is of critical importance in the physiologic and pathologic findings of the cornea. Clinicians should recall that quantitative and qualitative disorders of the PTF can be a cause of corneal diseases as well as a clinical sign of ocular surface diseases. It is also important to consider that some systemic and topical treatments may affect the PTF volume and composition. Not all interactions are known at this time. There is a continued need for basic research into PTF components in healthy and diseased equine eyes, because much remains unknown. Until additional basic information about the biochemical composition and pH of the PTF as well as the interactions between equine corneal pathogens and specific PTF deficits becomes available, it will not be possible to define the cause and effect relations precisely between the various ocular surface diseases and deficiencies,excesses, and imbalances of PTF components. It is quite possible that a number of equine corneal diseases may be manifestations of qualitative PTF disorders. PMID- 15271426 TI - Corneal epithelial disease. AB - This article reviews conditions that primarily affect the corneal epithelium as distinct from corneal stromal diseases. Corneal ulceration is discussed elsewhere in this issue. The other corneal conditions include a variety of more subtle epithelial disease,which might colloquially be termed epitheliopathies, as well as uninfected indolent superficial ulcerations, corneal neoplasia, and eosinophilic keratitis. The fungal plaque is an unusual and somewhat chronic form of epithelial infection. Although less common than corneal ulcers, this collection of lesions may present uniquely challenging obstacles to diagnosis and identification of a cure. Newer therapies and surgical strategies are discussed. PMID- 15271427 TI - Inflammatory stromal keratopathies: medical management of stromal keratomalacia, stromal abscesses, eosinophilic keratitis, and band keratopathy in the horse. AB - This article discusses the diagnosis and medical treatment of stromal keratomalacia or "melting ulcers," stromal abscesses, eosinophilic keratitis (EK), and calcific band keratopathy. These are common and important inflammatory keratopathies of the equine corneal stroma. Keratomalacia and stromal abscesses are associated with infection, leukocytic invasion of the stroma, and loss of tissue and tear film proteinase homeostasis. Eosinophils infiltrate the stroma in response to unknown stimuli in EK. Calcium is deposited in the stroma and epithelium secondary to chronic equine recurrent uveitis in calcific band keratopathy. They are all associated with varying degrees of iridocyclitis. PMID- 15271428 TI - Equine corneal surgery and transplantation. AB - Corneal disease is common in equine ophthalmology and requires vigilant monitoring and appropriate therapy to optimize the outcome. Many equine corneal diseases, particularly those that progress rapidly, may benefit from surgical intervention. These include descemetoceles, deep corneal lacerations and ulcers, corneal perforation/iris prolapse, ulcerative keratitis, corneal stromal abscesses, and corneoscleral neoplasia. Indications for corneal transplantation include optical, tectonic, therapeutic, and cosmetic purposes. Corneal transplantation is most often implemented in equine patients for tectonic and therapeutic reasons when a cornea is compromised by corneal stromal abscess, iris prolapse, or neoplasia. This article provides an outline of when to consider surgical intervention for corneal disease, the procedures available and expected outcomes, and how appropriate early surgical intervention can dramatically improve the end result. PMID- 15271429 TI - Equine glaucoma. AB - Glaucoma is a diverse group of vision-impairing disorders that have as a common bond an elevation of intraocular pressure(IOP) to a level incompatible with the health of the eye. Glaucoma can be congenital, primary, or secondary. Congenital equine glaucoma is associated with developmental abnormalities of the iridocorneal angle or, in many cases, with the more severe anterior segment dysgenesis. PMID- 15271430 TI - The lens and cataracts. AB - It is conservatively estimated that some form of lens opacity is present in 5% to 7% of horses with otherwise clinically normal eyes.These opacities can range from small epicapsular remnants of the fetal vasculature to dense and extensive cataract. A cataract is defined technically as any opacity or alteration in the optical homogeneity of the lens involving one or more of the following: anterior epithelium, capsule, cortex, or nucleus. In the horse, cataracts rarely involve the entire lens structure (ie, complete cataracts) and are more usually localized to one anatomic landmark or sector of the lens. Complete cataracts are invariably associated with overt and significant visual disability. Focal or incomplete cataracts alone seldom cause any apparent visual dysfunction in affected horses,however. PMID- 15271431 TI - Equine recurrent uveitis: new methods of management. AB - Equine recurrent uveitis (ERU) is one of the most common causes of blindness in horses. Until recently, treatment of this condition consisted only of symptomatic therapy, typically with steroidal and nonsteroidal medications. A better understanding of the disease process(es) has permitted new medical and surgical therapies that have recently been described. This article highlights clinical features of ERU, the causes of ERU, and new management and treatment options for horses with ERU. PMID- 15271432 TI - Ocular conditions of neonatal foals. AB - A discussion of ocular conditions of foals with an emphasis on congenital and inherited disorders is presented. An understanding of the normal postnatal development of the eye and adnexae is important. Recognition of inherited abnormalities is essential when giving advice on breeding suitability, and prompt attention or referral of deteriorating ocular conditions in foals ensures the best outcome for future use. Congenital conditions may be recognized for the first time in older animals during their first thorough eye examination. PMID- 15271433 TI - Ophthalmic imaging. AB - The availability of advanced imaging modalities in veterinary medicine has greatly widened the diagnostic imaging capabilities possible. Ultrasonography provides a rapid noninvasive modality that provides detailed examination and resolution of the intraocular structures and soft tissues surrounding the orbit of opaque eyes. Ultrasonography is cost-effective and widely available to practitioners,referral centers, and academic institutions. In many areas,mobile specialist ultrasonographers are available to supplement the equipment and skills of the practitioner. The added strengths of CT and MRI lie in their cross sectional capability and better image quality. Unfortunately, the cost of CT and MRI currently limits their availability to referral centers and academic institutions. Primarily because of financial considerations, CT is currently more widely available for evaluation of equine disorders than MRI. A thorough evaluation of the multiple images and an understanding of normal anatomy and abnormal tissue patterns are indicated to maximize the use of each modality. Unlike ultrasonography, which can be performed in awake horses, the costs and contraindications of general anesthesia in some critical patients should also be considered when using CT and MRI. Finally, imaging artifacts are frequently encountered with each of these modalities. Thus, a thorough understanding of the various types of artifacts that occur is important so as to avoid interpretation pitfalls. PMID- 15271434 TI - The prepurchase examination. AB - The prepurchase examination is performed to aid a prospective purchaser in determining the suitability of a specific patient for an anticipated use and, perhaps, an intended rider. The ophthalmic portion of that examination can be performed satisfactorily with minimal equipment by following a systematic protocol that includes examination of all segments of the globe and an evaluation of the patient's vision. Findings of the examination should be discussed with and made in writing to the buyer so that he or she can make an informed decision about the suitability of the purchase. PMID- 15271435 TI - Cosmetic globe surgery in the horse. AB - Effects of traumatic injury or inflammation on the equine eye can be catastrophic. These ocular conditions can frequently result in blindness or chronic pain. In addition to blindness and pain, permanent unsightly cosmetic defects can occur. This article addresses options available for improved cosmetic outcome in horses with ocular scars or requiring enucleation. Many of these options have been described in detail previously. New information has been added to the discussion of each option where pertinent. PMID- 15271436 TI - Standing sedation and pain management for ophthalmic patients. AB - Several ocular procedures, including examination, removal of corneal foreign bodies, nictitans surgery, eyelid repair, and tumor excision,can be successfully performed in the appropriately restrained and sedated standing horse. Sedation is best achieved with xylazine,with or without the addition of acepromazine. Additional analgesia can be provided with appropriate local anesthetic blocks. Surgical conditions are greatly improved by using an auriculopalpebral and supraorbital block and topical anesthetics. More elaborate standing sedation involving continuous rate infusions of lidocaine or detomidine combined with butorphanol may facilitate more involved surgery with appropriate support staff and equipment in animals that are at high risk for general anesthesia or when the latter is not an option. Short-term or long-term analgesia is most commonly provided with nonsteroidal anti-inflammatory drugs, but several newer techniques, including lidocaine and butorphanol infusions, may be effective. Topical treatment with opioids to provide analgesia and opioid antagonists to enhance corneal healing is an exciting new development that may revolutionize our approach to corneal ulcer therapy in the future if current research findings are supportive. PMID- 15271438 TI - Proceedings of the 15th International Congress on Animal Reproduction. Porto Seguro, Brazil, 8-12 August 2004. PMID- 15271439 TI - Production of bioproducts through the use of transgenic animal models. AB - Transgenic livestock that produce recombinant proteins in their milk can provide an economic and safe system for production of valuable proteins, such as pharmaceutical proteins for treatment or prevention of human disease or biomaterials for medical use. This method of production is frequently referred to as biopharming. The promise of biopharming, that is the actual commercial production of pharmaceuticals and other bioproducts, is nearing fulfillment. Improvements in molecular and reproductive techniques and strong economic incentives have continued to drive the implementation of transgenic technology to domestic animals. Nuclear transfer using transgenic donor cells is rapidly becoming the predominant technique used in the production of transgenic livestock, replacing the direct injection of DNA into the zygotic pronuclei. Production of transgenic founder animals by nuclear transfer in combination with traditional reproductive technologies can result in the propagation of transgenic herds of sufficient size to meet market demands for commercially important proteins. While some of the companies that have established transgenic programs have run into setbacks owing to a combination of economic, scientific and regulatory difficulties, other companies are continuing to make significant advances. While further improvements are needed to increase efficiencies of production, economically viable production of recombinant proteins using livestock species is not only possible but should be a commercial reality in the very near future. PMID- 15271440 TI - Genome activation and developmental block in bovine embryos. AB - The ultimate goal of in vitro embryo culture systems is to perfectly mimic the condition of oocyte maturation, fertilization and embryo development. These systems are far more complex than standard in vitro cell culture because of the various environments through which the gametes and embryos pass during in vivo development. Improvement of the medium and other culture conditions has allowed for full development of a percentage of the fertilized oocytes but the great majority of bovine zygotes stop developing within a few cell cycles after initiating cleavage. This developmental block arises in the bovine embryo at the eight-cell-stage and is likely correlated with the cytoplasmic quality of the oocyte. Oocytes harbor all mRNAs and proteins needed to reach the fourth or fifth cell cycle, however, embryos that fail to transcribe their own genome fail to further develop. In this article, we review some of the advances in developmental block knowledge and describe a possible role of active embryo transcription that drives incompetent embryos to block and death. PMID- 15271441 TI - International training programs in reproductive sciences for conservation of Latin American felids. AB - Survival of the ten non-domestic felid species endemic to Latin America is imperiled by habitat loss, poaching and poor captive management. Over the past 10 years, conservation of these felids has been the primary focus of a reproductive research and training program conducted in Brazil, Mexico, and the USA. The objectives of this program were to: (1) provide intensive training in reproductive sciences to Latin American scientists, (2) conduct collaborative studies investigating basic and applied reproduction in endangered felids, and (3) establish a highly-trained scientific cohort to conduct independent conservation-based research. Four formal training courses, consisting of didactic lectures and hands-on instruction in research techniques, including semen collection, sperm cryopreservation and laparoscopic artificial insemination (AI), were taught in Brazil and Mexico between 1995 and 1998. Several of these scientists received further training in conducting fecal hormone analysis in the USA, and a number of research studies, many in collaboration with American scientists, were initiated in Latin American felids. Research findings have characterized basal reproductive traits in several felid species, including ocelots, margay, tigrinas and jaguars, and established that Latin American felids exhibit only minimal seasonal variation in most reproductive traits. Other studies have explored the impact of acute and chronic stressors on adrenocortical activity and demonstrated the importance of environmental enrichment in captivity, especially in small felids. Additional research has examined ovarian and immunological responsiveness of Latin American felids to exogenous gonadotropins and assessed the impact of nutrition on sperm production and oocyte quality. Applied reproductive studies have investigated sperm cryopreservation in both captive and wild felid populations and demonstrated the production of viable offspring in ocelots and tigrinas following laparoscopic AI. Ongoing studies are investigating the potential of in vitro fertilization (IVF), embryo cryopreservation and embryo transfer for genetic management of ocelots and tigrinas. To date, over 75 Brazilian ocelot and 50 tigrina IVF embryos have been cryopreserved and two pregnancies have been established in ocelots following transfer of frozen-thawed embryos. Findings from these studies are helping to improve husbandry, population management, and breeding of Latin American felids in captivity. Continued advances in assisted reproduction eventually may provide an alternative route for exchanging genetic material among Latin American felid populations. Most importantly, this collaborative program has been essential for building scientific capacity, within Brazil and Mexico, in establishing a core group of highly-trained reproductive biologists that will continue applying their new knowledge and skills to the conservation of Latin American felids. PMID- 15271442 TI - Placentation in species of phylogenetic importance: the Afrotheria. AB - Afrotheria, one of four mammalian superorders, comprises elephants, sea cows, hyraxes, aardvark, elephant shrews, tenrecs and golden moles. Their placentas either form an equatorial band or are discoid in shape. The interhemal region, separating fetal and maternal blood, is endotheliochorial in elephants, aardvark and possibly the sea cows, but hemochorial in the remaining orders. There is a secondary epitheliochorial placenta in elephant shrews while a similar structure in tenrecs erodes maternal tissues. Specialized hemophagous regions are a striking characteristic of some of these placentas yet absent in hyraxes, elephant shrews, and golden moles. It is possible that the common ancestor of the Afrotheria had an endotheliochorial placenta. Establishment of a hemochorial condition, as seen in rock hyraxes, elephant shrews, tenrecs, and golden moles, would be a more recent development. The elephant, manatee, and aardvark all have circumferential placentas. Thus the formation of a discoid placenta with a more or less extensive secondary placenta in elephant shrews and tenrecs would also be a derived state. PMID- 15271443 TI - Reproductive problems directly attributable to long-term captivity--asymmetric reproductive aging. AB - Problems attributable to long-term captivity have been identified and are responsible for the difficulties in establishing successful reproduction in captive populations of wildlife, specifically, elephants and rhinoceroses. Historically, non-reproductive periods of 10-15 years in nulliparous female rhinoceroses and elephants have not been considered problematic. New evidence suggests that prolonged exposure to endogenous sex steroids and that long stretches of non-reproductive periods induce asymmetric reproductive aging in captive animals. The consequences are reduced fertility, shortened reproductive life-span and, eventually, irreversible acyclicity. Although age-related reproductive lesions have also been documented in male rhinoceroses, they continue to maintain a longer reproductive life-span than females. Since human and domestic animal models have already indicated that early pregnancy provides natural protective mechanism against asymmetric reproductive aging processes and premature senescence, it is imperative that appropriate counter measures such as assisted reproductive technologies (ART) be utilized to ensure early pregnancy in captive animals for their preservation and to ensure increased genetic diversity of the captive populations. PMID- 15271444 TI - DNA methylation in the preimplantation embryo: the differing stories of the mouse and sheep. AB - In mammals, active demethylation of cytosine methylation in the sperm genome prior to forming a functional zygotic nucleus is thought to be a function of the oocyte cytoplasm important for subsequent normal development. Furthermore, a stepwise passive loss of DNA methylation in the embryonic nucleus has been observed as DNA replicates between two-cell and morula stages, with somatic cell levels of methylation being re-established by, or after the blastocyst stage when differentiated lineages are formed. The ability of oocyte cytoplasm to also reprogram the genome of a somatic cell by nuclear transfer (SCNT) has raised the possibility of directing reprogramming of a somatic nucleus ex ovo by mimicking the epigenetic events normally induced by maternal factors from the oocyte. Whilst examining DNA methylation changes in normal sheep fertilization, we were surprised to observe no demethylation of the sheep male pronucleus at any point in the first cell cycle. Furthermore, using quantitative image analysis, we observed limited demethylation of the sheep embryonic genome only between the two and eight-cell stages and no evidence of remethylation by the blastocyst stage. We suggest that the dramatic differences in DNA methylation between the sheep and other mammalian species examined call in to question the requirement and role of DNA methylation in early mammalian embryonic development. PMID- 15271445 TI - Integration of sperm sexing technology into the ART toolbox. AB - Sex-sorting of mammalian spermatozoa has applications for genetic improvement of farm animals, in humans for the control of sex-linked disease, and in wildlife as a captive management strategy and for the re-population of endangered species. Considerable research has been undertaken worldwide on the Beltsville sperm sexing technology, the only effective method for pre-selection of sex of offspring. The combination of this method with assisted reproductive technologies has resulted in the birth of offspring in a wide range of animals, including cattle, the only livestock species in which sperm sexing is used commercially. Major improvements in the efficiency of sorting, in particular the development of high speed sorting (15 million X and Y spermatozoa per hour) have led to the production of offspring using conventional and low dose AI and the successful cryopreservation of sorted spermatozoa in cattle, sheep, horses and elk. A major limitation remains the short viable lifespan of sorted spermatozoa in the female genital tract, in most species necessitating sperm deposition deep in the uterus, and close to the expected time of ovulation, for acceptable fertility after in vivo insemination. Special deep uterine insemination technology has been employed to produce offspring in pigs and horses using low sperm doses. Considerable attention has been paid to reduction of the damage and capacitation-like changes to spermatozoa that result from flow cytometric sorting and from freezing and thawing. However, high-purity sorting of liquid-stored or frozen-thawed spermatozoa for immediate use, or re-cryopreservation for later use, does not reduce its fertilizing capacity in vitro, allowing its combination with in vitro fertilization or juvenile in vitro embryo transfer to produce blastocysts, and offspring in sheep and cattle after embryo transfer. Further research into sorting and preservation methods that incorporate strategies to prevent destabilization of sperm membranes may improve the fertilizing lifespan of flow cytometrically sorted spermatozoa. With continued improvement in sorting instrumentation and biological handling, sorting efficiency should reach a point where commercially acceptable pregnancy rates may be achieved in a number of species after conventional or deep uterine insemination. PMID- 15271446 TI - Interpretation of reprogramming to predict the success of somatic cell cloning. AB - In the context of mammalian somatic cell cloning, the term reprogramming refers to the processes that enable a somatic cell nucleus to adopt the role of a zygotic nucleus. Gene re-expression is one measure of reprogramming if correlated with subsequent developmental potential. This paper describes several experiments utilizing pre-implantation gene expression to evaluate reprogramming and clone viability. We have established a direct correlation between Oct4 expression in mouse clones at the blastocyst stage and their potential to maintain pluripotent embryonic cells essential for post-implantation development. Furthermore, the quality of gene expression in clones dramatically improves when genetically identical clones are combined in clone-clone aggregate chimeras. Clone--clone aggregates exhibit a higher developmental potential than single clones both in vitro and in vivo. This could be mediated by complementation between blastomeres from epigenetically different clones within the aggregate rather than by the increase in cell number resulting from aggregation. We also discuss the use of tetraploid embryos as a model to evaluate reprogramming using gene expression and demonstrate that somatic cell nuclei can be reprogrammed by blastomeres to re express embryonic specific genes but not to contribute to post-implantation development. PMID- 15271447 TI - Follicular development: the role of the follicular microenvironment in selection of the dominant follicle. AB - The importance of endocrine signals in the regulation of follicular development has long been recognized. However, the follicular microenvironment also plays a critical role in determining follicular fate. This review summarizes our studies on the role of the intrafollicular IGF system in selection of the dominant follicle (DF) in cattle. During the bovine estrous cycle, the largest antral follicles develop in two or three successive waves of follicular recruitment and selection of a DF. High concentrations of estradiol in the follicular fluid are the hallmark of dominant and preovulatory follicles and are associated with lower concentrations of low molecular weight (MW) insulin-like growth factor binding proteins (IGFBP-2, -4, and -5), which can prevent binding of IGF to its receptor. Our studies have shown that dominant and preovulatory follicles also have much higher levels of an IGFBP-4/-5 protease activity, which is the bovine equivalent of the human IGFBP-4 protease, pregnancy-associated plasma protein-A (PAPP-A). Studies of follicles isolated just after the emergence of the DF showed that PAPP A is present in the follicular fluid of the DF as soon as it can be detected as morphologically dominant. To examine whether higher levels of PAPP-A in one follicle of the cohort (the future DF) precedes morphological dominance, the four largest follicles were isolated from pairs of bovine ovaries obtained before one follicle of the cohort was significantly larger the others, around the time that one follicle was first detected as morphologically dominant and after dominance was well established. Analysis of the temporal sequence of changes in estradiol, low MW IGFBPs, free IGF, and PAPP-A in the follicular fluid suggested that an increase in PAPP-A is the earliest biochemical difference yet detected in the future DF and that follicular selection is the result of a progressive series of changes beginning with the acquisition of PAPP-A, which leads to a decrease in IGFBP-4 and -5 and an increase in free IGF, which synergizes with FSH to increase estradiol production. Co-dominant follicles, induced by injection of small doses of recombinant bovine (rb) FSH, both had levels of PAPP-A similar to the single DF of control heifers, supporting the hypothesized role of FSH in the induction of PAPP-A in the DF. Taken together, these results suggest a critical role for FSH-induced PAPP-A, and thus for free IGF, in the selection of the DF. In contrast, other experiments provided evidence for a deleterious effect of IGF on the initiation of bovine follicular growth and the survival of primordial and primary follicles in vitro. These results underscore the importance of the follicular microenvironment in determining follicular fate and indicate that its effects can be stage-specific. PMID- 15271448 TI - Vascular control of ovarian function: ovulation, corpus luteum formation and regression. AB - Hemodynamic changes are involved in the cyclic remodeling of ovarian structures. A transrectal color Doppler ultrasonography was used to assess the blood flow and changes in the vasculature that take place in the follicle wall and within the corpus luteum (CL) during specific physiological events such as ovulation, CL development, and CL regression in cows. To investigate the local release of vasoactive peptides, steroid hormones, and prostaglandins (PGs) in the ovarian microenvironment, the capillary membranes (0.2mm diameter and 5-10mm length) of a microdialysis system (MDS) were implanted into the follicle wall and the CL in vitro. Furthermore, in vivo experiments were conducted with the same MDS membranes surgically implanted in follicle wall or on CL along with ovarian venous and jugular catheters to collect simultaneous, real-time information on the ovarian and systemic changes in the secretion of factors regulating vascular function. Based on the results obtained from the series of in vitro and in vivo experiments, we propose that a functional "cross-talk" occurs between the vascular components (endothelial cells) and steroidogenic cells to control follicular and luteal functions in the bovine ovary. PMID- 15271449 TI - Evidence for the role of oviduct secretions in sperm function, fertilization and embryo development. AB - The oviduct is a dynamic organ which facilitates gamete function, fertilization and embryo development. Secretions of the oviduct, recovered by tissue culture or cannulation techniques have been used to define the composition of the oviduct milieu, as well as functions associated with stage of the reproductive cycle or region of the oviduct. Several oviduct proteins have been shown to associate with the gametes and embryos. Ongoing studies are directed at identifying oviduct proteins and determining their function. Oviduct-specific glycoproteins (OSG) have been purified from the oviduct and shown in vitro to have positive affects on sperm capacitation, sperm-ovum binding, ovum penetration and embryo development. Osteopontin, another oviduct secretion, also has been shown to stimulate fertilization and embryo development. The picture emerging is that some components of the oviduct milieu have overlapping functions to collectively provide a failsafe system to ensure fertility in vivo so that success is not dependent on a single component. PMID- 15271450 TI - Role of leptin in the regulation of gonadotropin secretion in farm animals. AB - The recently discovered protein, leptin, which is secreted by fat cells, has been implicated in regulation of feed intake or energy balance and the neuroendocrine axis in rodents, humans and large domestic animals. Leptin was first identified as the gene product found to be deficient in the obese (ob/ob) mouse. Administration of leptin to ob/ob mice restored reproduction as well as reducing feed intake and causing weight loss. The leptin receptor (LR) which has been cloned and is a member of the class 1 cytokine family of receptors, is found in the brain and pituitary of all species studied to date. Neuropeptide Y has been proposed as the primary mediator of leptin action in the hypothalamus to regulate luteinizing hormone (LH) and growth hormone (GH) secretion. In vitro studies using both hypothalamic explants and pituitary cell culture provided evidence that supports a direct action of leptin at the level of brain and pituitary gland in the pig, but only the pituitary in cattle. Central administration of leptin increased LH secretion in the fasted cow and ewe, but not in control fed animals, indicating that metabolic state is an important factor in modulating the hypothalamic-pituitary response to leptin. Changing serum leptin concentrations and leptin mRNA expression were associated with onset of puberty in heifers and gilts. Thus, leptin appears to be an important link between metabolic status and the neuroendocrine axis. PMID- 15271451 TI - Effects of maternal nutrition on fetal and neonatal reproductive development and function. AB - Maternal undernutrition and, under certain circumstances overnutrition, before or during pregnancy or during early postnatal life can alter reproductive function of the offspring. Effects can be exerted at many stages of development, from prior to conception until after birth and may be expressed at the time of the nutritional insult or later. Since patterns of development differ between species, it is probably more appropriate to consider effects in relation to a stage of development rather than relative to the time of birth. Effects exerted at one stage of development may be expressed later, even if the nutritional influence is no longer present. The signals by which maternal nutrition affects the offspring must be related to maternal nutritional state and must have the capacity to reach the embryo, to be 'read' by it and to modify expression of selected genes. It is suggested that single nutrients and/or metabolites are unlikely to have direct impacts on the pattern of development of the reproductive system and it is postulated that multiple endocrine and metabolic signals are involved. Whilst it has been shown that many components of the hypothalamic pituitary-gonadal system are modified by early life nutritional influences, understanding of the mechanisms through which these effects are exerted remains limited. PMID- 15271452 TI - Effects of ghrelin on food intake and neuroendocrine function in sheep. AB - Ghrelin, a novel acylated peptide, is the endogenous ligand for growth hormone secretagogue (GHS) receptor. Ghrelin is produced mainly in the oxyntic glands of the stomach, but also produced in the intestines, kidneys, hypothalamus and pituitary gland. Circulating ghrelin levels have been shown to rise before a meal and fall afterwards, suggesting that anticipation of a meal may stimulate secretion. In some species, ghrelin administration has been shown to stimulate growth hormone (GH) secretion, and to cause weight gain by increasing food intake and reducing metabolic utilization of fat. Furthermore, intracerebroventricular and intravascular administration of ghrelin increases gastric acid output in a dose-dependent manner. Thus, ghrelin may play an important role in controlling feeding behavior and energy homeostasis. We have investigated the role of ghrelin in the control of feeding and neuroendocrine function in ruminants using sheep as an experimental model. This mini review describes mechanisms regulating ghrelin secretion at feeding time, and also focuses on the neuroendocrine functions of ghrelin. PMID- 15271453 TI - Canine brucellosis. AB - This review discusses the prevalence, etiology, pathogenesis, clinical findings, diagnostic methods, therapy, management and public health considerations of Brucella canis infection in dogs. Canine brucellosis is a contagious infection produced by a gram-negative coccobacilus called Brucella canis. The main sources of infection are vaginal fluids of infected females and urine in males. Routes of entry are venereal, oronasal, conjunctivae mucosa and placenta. The most significant symptoms are late abortions in bitches, epididymitis in males and infertility in both sexes, as well as generalized lymphadenitis, discospondylitis and uveitis. Diagnosis is complex because serology can give false positive results and chronic cases can give negative results, needing to be complemented with bacteriological studies. No antibiotic treatment is 100% effective and the infection often recurs in animals apparently treated successfully. Infected animals must be removed from the kennels and no longer used for breeding. Preferably, males should be castrated and females spayed. Human contagion is not frequent, although it has been reported, and is easily treated. PMID- 15271454 TI - Canine fresh and cryopreserved semen evaluation. AB - There is little doubt that objective assessment of multiple parameters related to the functional and morphological characteristics of spermatozoa, increase the predictability of the fertilizing potential of a semen sample. Conventional microscopic methods for sperm evaluation in combination with the more objective computer-assisted sperm motility and morphology analyzers and flow cytometry, have allowed investigators to obtain precise information about the morphofunctional status of spermatozoa, which already has resulted in a better understanding of sperm biology and of some of the mechanisms involved in sperm cryoinjury. In addition, assays based on in vitro fertilization provide valuable information about the functional ability of spermatozoa when interacting with the oocyte. Although objective methods for dog sperm evaluation are available for many researchers, considerable effort has still to be invested in order to standardize physiological sperm parameters for the results to be interpretable under unified criteria. In the present paper some of the classical and new methods currently used for dog sperm evaluation are reviewed, though not exhaustively, covering some aspects of the sperm cell that are useful in estimating its functional ability. PMID- 15271455 TI - Using genetic technologies for promoting canine health and temperament. AB - The entire canine genome is scheduled to be sequenced by researchers at the Whitehead Institute/MIH Center for Genome Research by the end of 2004. Thus, new genetic technologies are likely to be developed that can soon predict certain aspects of health and temperament in dogs. The C. familiaris is similar in size to that of humans and other mammals, with an estimated 2.8 billion base pairs. Although, the boxer was chosen as the first breed to sequence, it will have application for all dog breeds. Once the entire genome is sequenced, genetic markers for specific diseases and temperaments may be developed, which can guide breeders to make informed decisions concerning breeding management. Such a technology may be useful for guide and service dog organizations that have breeding colonies. It is important that the human-animal bond be preserved for as long as possible, both for pet owners and also for those disabled individuals who depend upon a dog for independence and mobility. Because genetic diseases may not manifest in carriers, and some genetic diseases do not manifest until after a dog is older and has already produced other animals with the same defect, genetic markers to identify some of the over 400 genetic diseases could be very useful in promoting canine health. PMID- 15271456 TI - Natural methods for increasing reproductive efficiency in small ruminants. AB - This paper describes three strategies to improve the reproductive performance of small ruminants in ways that lead to "clean, green and ethical" animal production. The first is aimed at control of the timing of reproductive events for which we turn to the socio-sexual inputs of the "male effect" to induce synchronised ovulation in females that would otherwise be anovulatory. The second strategy, "focussed feeding", is based on our knowledge of the responses to nutrition and aims to develop short programs of nutritional supplements that are precisely timed and specifically designed for individual events in the reproductive process, such as gamete production, embryo survival, fetal programming and colostrum production. The third strategy aims to maximise offspring survival by a combination of management, nutrition and genetic selection for behavior (temperament). All of these approaches involve non pharmacological manipulation of the endogenous control systems of the animals and complement the detailed information from ultrasound that is now becoming available. The use of such clean, green and ethical tools in the management of our animals can be cost-effective, increase productivity and, at the same time, greatly improve the image of meat and milk industries in society and the marketplace. PMID- 15271457 TI - Fecundity genes in sheep. AB - Since 1980 there has been increasing interest in the identification and utilisation of major genes for prolificacy in sheep. Mutations that increase ovulation rate have been discovered in the BMPR-1B, BMP15 and GDF9 genes, and others are known to exist from the expressed inheritance patterns although the mutations have not yet been located. In the case of BMP15, four different mutations have been discovered but each produces the same phenotype. The modes of inheritance of the different prolificacy genes include autosomal dominant genes with additive effects on ovulation rate (BMPR-1B; Lacaune), autosomal over dominant genes with infertility in homozygous females (GDF9), X-linked over dominant genes with infertility in homozygous females (BMP15), and X-linked maternally imprinted genes (FecX2). The size of the effect of one copy of a mutation on ovulation rate ranges from an extra 0.4 ovulations per oestrus for the FecX2 mutation to an extra 1.5 ovulations per oestrus for the BMPR-1B mutation. DNA tests enable some of these mutations to be used in genetic improvement programmes based on marker assisted selection. PMID- 15271458 TI - Advanced assisted reproduction technologies (ART) in goats. AB - Assisted reproduction technologies (ART) are reviewed with special emphasis on goat genetic improvement programs. Estrous synchronization and artificial insemination are the most commonly used ART worldwide because of their simplicity and excellent cost/benefit, especially when proven sires are used. Multiple ovulation and embryo transfer (MOET) has not become widely used due to its unpredictability. In vitro embryo production using oocytes collected by laparoscopy from valuable donors has the potential to improve the results obtained from MOET and expand its applications (for example, using prepubertal donors). However, the costs and inefficiencies of the system might restrict its use to special situations. Finally, transgenesis and cloning are expected to have a significant impact on the future genetic improvement of livestock. However, because of low efficiencies and high costs, their present use is restricted to applications with high returns such as the production of recombinant proteins of pharmaceutical and biomedical interest. PMID- 15271459 TI - Major histocompatibility antigen expression on the bovine placenta: its relationship to abnormal pregnancies and retained placenta. AB - In viviparous animals, regulation of expression of major histocompatibility complex (MHC) class I antigens by the trophoblast cells, which constitute the outermost layer of the placenta, seems to be critical for maternal immunological acceptance of an allogeneic fetus. Cattle are unusual in this regard, since the bovine trophoblast cells, in specific regions of the uterine/placental interface, normally express MHC class I antigens during the third trimester of gestation. This expression appears to be biologically relevant as MHC class I compatibility between a cow and her fetus has been associated with an increased incidence of placental retention. We have found significant differences in lymphocyte populations, cytokine production, and trophoblast cell apoptosis in the placentomes of MHC-compatible and -incompatible pregnancies at parturition. This suggests that maternal immunological recognition of fetal MHC class I proteins triggers an immune/inflammatory response that contributes to placental separation at parturition in cattle. Early in pregnancy, a complete shutdown of MHC class I expression by trophoblast cells appears to be critical for normal placental development and fetal survival. In bovine somatic cell nuclear transfer (SCNT) pregnancies, there is an extremely high rate of fetal loss between days 30 and 90 of pregnancy. We have shown that in bovine SCNT pregnancies, between days 34 and 63 of gestation, there is both abnormal expression of MHC class I antigens by trophoblast cells and an abnormal accumulation of lymphocytes within the uterine stroma. Consequently, it is likely that activation of the maternal mucosal immune system, within the uterus at the same time when placentomes are being established, interferes with the process of placentome development and leads to immune-mediated abortion. Our data suggest that bovine MHC-compatible pregnancies provide a unique model for studying regulation of the uterine immune system, as well as immune-mediated placental rejection. PMID- 15271460 TI - Steroidal regulation of uterine immune defenses. AB - Progesterone suppresses uterine immune defenses and predisposes postpartum animals to nonspecific uterine infections. Progesterone can also suppress uterine eicosanoid synthesis. This effect of progesterone seems to be an important factor in the onset of uterine infections because eicosanoids can enhance uterine immune defenses. In fact, exogenous prostaglandin F(2alpha) (PGF(2alpha)), an eicosanoid that stimulates uterine PGF(2alpha) production, enhances uterine immune defenses and promotes the ability of ewes and sows to resolve uterine infections, even when progesterone is maintained at luteal phase concentrations. Prostaglandin F(2alpha) is also a proinflammatory molecule that stimulates the production of proinflammatory cytokines and may enhance uterine production of leukotriene B(4) (LTB(4)), which stimulates various neutrophil functions. Neutrophils seem to mount the initial response to bacteria that enter the uterus, and proinflammatory cytokines and LTB(4) enhance phagocytic activity of neutrophils. Even though there are clear associations among PGF(2alpha), LTB(4), proinflammatory cytokines, phagocytosis, and the ability of the uterus to resist or resolve infections, the mechanisms of action of exogenous PGF(2alpha) in mitigating the immunosuppressive effects of progesterone have not yet been defined. However, defining the PGF(2alpha) mechanisms should yield important new information that can be used to develop novel prevention and treatment strategies that do not rely on antibiotic and antimicrobial compounds for managing uterine infections. PMID- 15271461 TI - Postpartum uterine health in cattle. AB - Uterine health is often compromised in cattle because postpartum contamination of the uterine lumen by bacteria is ubiquitous, and pathogenic bacteria frequently persist causing clinical disease. The subfertility associated with uterine infection involves perturbation of the hypothalamus, pituitary and ovary, in addition to the direct effects on the uterus, and appears to persist even after clinical resolution of the disease. Absorption of bacterial components from the uterus can prevent the follicular phase LH surge and ovulation. In addition, the first postpartum dominant follicle has a slower growth rate and secretes less estradiol at the end of the growth phase. There are also localised ovarian effects of high uterine bacterial growth density, because fewer first dominant follicles are selected in the ovary ipsilateral than contralateral to the previously gravid uterine horn. Thus, it is important to diagnose and treat uterine disease promptly and effectively. Examination of the contents of the vagina for the presence of pus is the most useful method for diagnosis of endometritis. The character and odor of the vaginal mucus can be scored and this endometritis score is correlated with the growth density of pathogenic bacteria in the uterus, and is prognostic for the likely success of treatment. The challenge for the future is to design prevention and control programs to reduce the incidence of disease, and understand how the immune and endocrine systems are integrated. PMID- 15271462 TI - Bicarbonate and its role in mammalian sperm function. AB - This paper deals with the effects of bicarbonate induced signaling pathways on plasma membrane lipid organization and downstream protein signaling, and their role in sperm-egg interactions. It also provides an overview of results that indicate that bicarbonate responses are not related to cell death or apoptosis. The information presented shows that only those sperm cells that have functionally completed maturation in the epididymis are sensitive to bicarbonate whereas immature sperm fail to respond to this physiological challenge. Therefore, it is important to selectively analyze the responsive sperm subpopulations when studying sperm capacitation. Moreover, bicarbonate induced signaling responses differ within the diverse sperm structures (e.g. the tail versus the head). Consequently, dissecting sperm structures and signaling areas from each other deserves more attention in sperm capacitation research. The information discussed was obtained from a variety of mammalian species but the basic bicarbonate-mediated sperm responses are similar in most Eutherian species despite some species to species variations (most notably in kinetics rather than the sequence of events). The objective of the paper is to provide a comparative experimental overview of bicarbonate mediated sperm capacitation in the hope that this information will lead to a better understanding of the complex biochemical nature of the involvement of bicarbonate in mammalian sperm capacitation. PMID- 15271463 TI - Post-testicular sperm environment and fertility. AB - When mammalian spermatozoa exit the testis, they show a highly specialized morphology; however, they are not yet able to carry out their task: to fertilize an oocyte. This property, that includes the acquisition of motility and the ability to recognize and to fuse with the oocyte investments, is gained only after a transit through the epididymis during which the spermatozoa from the testis travel to the vas deferens. The exact molecular mechanisms that turn these cells into fertile gametes still remain mysterious, but surface-modifying events occurring in response to the external media are key steps in this process. Our laboratory has established cartographies of secreted (secretomes) and present proteins (proteomes) in the epididymal fluid of different mammals and have shown the regionalized variations in these fluid proteins along the epididymis. We have found that the main secreted proteins are common in different species and that enzymatic activities, capable of controlling the sperm surface changes, are present in the fluid. Our studies also indicate that the epididymal fluid is more complex than previously thought; it contains both soluble and particulate compartments such as exosome-like vesicles (epididymosomes) and certainly specific glycolipid-protein micelles. Understanding how these different compartments interplay to modify sperm components during their transit will be a necessary step if one wants to control and to ameliorate sperm quality and to obtain valuable fertility markers helpful to establish a male fertility based genetic selection. PMID- 15271464 TI - Male fertility markers, myth or reality. AB - For the past many years my laboratory has been interested in the function of the epididymis in sperm maturation which allows the male gamete to acquire its fertilizing ability. Our work has focused on proteins secreted by the epididymis and the mechanisms by which these proteins are added to the sperm surface. We have focused on sperm proteins involved in the binding to the egg's extra cellular coat-the zona pellucida. For this purpose, we have used different animal models as well as humans. We have proposed that some epididymal sperm proteins can be used as marker of male fertility in humans and animals. PMID- 15271465 TI - Physiological and cellular adaptations of zebu cattle to thermal stress. AB - During their separate evolution from Bos taurus, zebu cattle (Bos indicus) have acquired genes that confer thermotolerance at the physiological and cellular levels. Cattle from zebu breeds are better able to regulate body temperature in response to heat stress than are cattle from a variety of B. taurus breeds of European origin. Moreover, exposure to elevated temperature has less deleterious effects on cells from zebu cattle than on cells from European breeds. Superior ability for regulation of body temperature during heat stress is the result of lower metabolic rates as well as increased capacity for heat loss. As compared to European breeds, tissue resistance to heat flow from the body core to the skin is lower for zebu cattle while sweat glands are larger. Properties of the hair coat in zebu cattle enhance conductive and convective heat loss and reduce absorption of solar radiation. At the cellular level, genetic adaptations to resist deleterious effects of elevated temperature result in preimplantation embryos from zebu being less likely to be inhibited in development by elevated temperature than are embryos from European breeds. The zebu genotype has been utilized in crossbreeding systems to develop cattle for beef and dairy production systems in hot climates but success has been limited by other unfavorable genetic characteristics of these cattle. An alternative scheme is to incorporate specific thermotolerance genes from zebu cattle into European breeds while avoiding undesirable genes. Once specific genes responsible for thermotolerance in zebu have been identified or mapped, breeding strategies such as marker-assisted selection and transgenics can be applied to further the exploitation of the zebu genotype for cattle production systems. PMID- 15271466 TI - Puberty in South American Bos indicus (Zebu) cattle. AB - Puberty in Zebu heifers follows a pattern characterized by a decrease in the steroid feedback mechanism and an increase in LH concentration, which result in the first ovulation followed by a short estrous cycle and the onset of normal cycles thereafter. These events are similar to those observed in Bos taurus cattle but occur at a later age. The late onset of puberty is both genetic and environmental in origin and is reflected by the age at first calving that can be at 40 months of age or older in these animals. Age at puberty in Zebu heifers has been shown to have a high heritability. Consequently, selecting precocious heifers may be an effective means of reducing age at puberty in these animals and this approach is being adopted in commercial practice. Genetic selection is not the sole solution to the problem because environmental improvements are necessary, particularly in terms of improved nutrition. South American Zebu cattle are usually subject to sub-optimum nutritional and management conditions and, hence, exhibit late onset of puberty. Hybrids of Zebu and Bos taurus cattle exhibit heterosis in respect of the age of puberty with earlier onset than expected in crossbred animals. Recently, purebred South American Zebu cattle have been shown to have Bos taurus genes, indicating that there have been previous attempts to improve their productivity using this approach. It was concluded that the age at first calving in South American Zebu cattle can be reduced by exposing well-fed, yearling heifers to bulls and selecting, over several generations, those animals that become pregnant at an early age. PMID- 15271467 TI - Pre-pubertal and postpartum anestrus in tropical Zebu cattle. AB - Bos indicus breeds, commonly known as Zebu cattle, have spread from their center of origin in Western Asia into large areas of Asia (including the Asia-Pacific basin), Africa, South and Central America (including the Caribbean islands). The original Zebu genotype, however, has been modified by planned and unplanned cross breeding programs involving many native and Bos taurus breeds in their new habitats. Though accurate estimates are not available, more than half of the world's cattle population includes a proportion of B. indicus germ plasma. B. indicus native breeds have developed by natural selection over centuries for their ability to survive in rough, harsh tropical environments. Most of these non described breeds still exhibit high fertility, in terms of calving rates, and disease resistance but they grow very slowly and take well over 3 years to reach puberty and produce only a few liters of milk over a short lactation period. Selection has been carried out in some areas and distinct Zebu breeds have been developed that have moderately high growth rate and milk production. However, they are slow breeders and have extended pre-pubertal and postpartum anestrous periods, compared to their temperate counterparts exposed to similar environment and management. The reproductive biology of B. indicus is similar to that of B. taurus. Most of the proven management, nutritional, hormonal and biotechnological interventions developed through experimentation with B. taurus breeds are equally applicable to B. indicus and their crosses. Zebu breeds predominate in most tropical countries where the majority of the human population lives. If meat and milk production are to be increased in the tropics, Zebu cow productivity, in terms of number of calves produced per lifetime or per unit area of land, must be increased and the time from birth to slaughter must be reduced. This goal could be achieved either by selection within local Zebu populations or through planned cross-breeding with B. taurus breeds. Because the productive and reproductive potentials of Zebu cattle are relatively low, worthwhile gains could only be achieved by selection over many generations. This would require substantial investment in labor, feed and drugs that may not be economic since the return from such investment is relatively low. However, many studies have shown that cross-breeding with B. taurus, which combines additive, dominance and epistatic effects of the two genotypes, ensures high productive and reproductive performance. Therefore, planned cross-breeding with suitable B. taurus breeds, although demanding additional investment in labor, feed and drugs, will still be economic because the return far exceeds the costs. PMID- 15271468 TI - Endocrine aspects in pathogenesis of mastitis in postpartum dairy cows. AB - In well-managed dairy herds some environmental pathogens including Gram-negative (GN) strains (E. coli and others) have been recognized recently as the predominant causative microbes of mastitis in the peri-parturient period. In early weeks of lactation hyperketonaemia may predispose the high-producing cows for GN mastitis. In GN mastitis cytokines, eicosanoids and oxygen radicals are released, which are responsible for the local and systemic symptoms. Experimental administration of endotoxin induces a complex endocrine cascade. Similar changes in plasma levels of cortisol, insulin, insulin-like growth factor-I and thyroid hormones are seen also in severe cases of GN mastitis. However, leptin is not responsible for the anorexia associated with severe mastitis in ruminants. Mastitis can postpone the resumption of ovarian cyclic activity in dairy cows when its outbreak occurs between days 15 and 28 after calving (at the expected time of first ovulation). In cyclic cows severe cases of GN mastitis can induce premature luteolysis or prolong the follicular phase. PMID- 15271469 TI - Immune cell infiltration of normal and impaired sow endometrium. AB - The paper reviews the physiological infiltration of immune cells, leukocytes, in the sow endometrium during different stages of the normal oestrous cycle, after mating and during early pregnancy. The mechanisms for development of endometritis in relation to oestrous cycle stages are also described. PMID- 15271470 TI - Fetal death: comparative aspects in large domestic animals. AB - Although the majority of pregnancy failures occur during the embryonic period, reports indicate that approximately 5% of detected pregnancies are lost during the fetal period, underlining the fact that fetal death is a substantial cause of economic loss. However, examination for fetal development or death during pregnancy is not performed routinely in domestic animals, and reference curves for normal fetal growth are, therefore, scarce. In this paper, the numerous possible causes of fetal death are reviewed briefly, with emphasis on the role of placental problems in fetal death and impaired fetal viability. In this respect, the role of placental insufficiency as a cause of pregnancy loss in twin pregnancies in monotocous species is well known, whereas the abnormal placental development leading to retarded fetal growth during pregnancies in recipients of in vitro produced (IVP) or nuclear transfer (NT) embryos has been less extensively documented. Fetal viability or death can be evaluated using hormonal, chemical and ultrasonographic parameters. For example, the viability of the feto placental unit can be examined by measuring maternal plasma concentrations of oestrone sulphate or the placental proteins, including pregnancy-associated glycoprotein (PAG) and pregnancy-specific protein B-60 (PSPB-60). Low concentrations of any of these three indicate either no pregnancy, or if pregnancy was confirmed earlier, fetal death and abnormally high or low levels can indicate fetal abnormality. Ultrasound can be used to examine the fetal heart rate (FHR), the incidence of fetal movements (FM), the appearance of fetal fluids and the development of the fetus and placenta. However, although abnormal FHRs have been correlated to subsequent fetal death, it is important to remember that there is a large physiological variation in FHR at the end of gestation, due to different behavioural states and differences in FM patterns. Although monitoring fetal viability and death using hormonal and ultrasonographic evaluations is possible during pregnancy in domestic animals, there is considerable physiological variations in the 'normal' values. Therefore, suitable combinations of tests need to be identified and more accurate reference values generated before such approaches can be considered reliable for monitoring the status of individual fetuses. PMID- 15271471 TI - Oocyte-somatic cell interactions during follicle development in mammals. AB - Our current perspectives on the relationship between the oocyte and its surrounding somatic cells are changing as we gain a greater understanding of factors regulating folliculogenesis. It is now widely accepted that the oocyte plays a very active role in promoting follicle growth and directing granulosa cell differentiation. The oocyte achieves this, in part, by secreting soluble paracrine growth factors that act on its neighboring granulosa cells, which in turn regulate oocyte development. In preantral follicles, the oocyte directs granulosa cells to regulate oocyte growth, and oocytes may also directly drive follicle growth. In antral follicles, the oocyte governs the behaviour of cells in its immediate vicinity, thereby actively regulating its own microenvironment. As such, the oocyte establishes and maintains the distinct cumulus lineage of granulosa cells. This oocyte-cumulus cell interaction, in general, prevents luteinization of cumulus cells by promoting growth, regulating steroidogenesis and inhibin synthesis, and suppressing luteinizing hormone receptor expression. Conversely, mural granulosa cells in antral follicles, which have no direct physical contact with the oocyte and, presumably, experience a more diffuse concentration of oocyte-secreted factors, proceed to a different phenotype. In the ovulating follicle, oocyte-secreted factors also play vital roles in enabling cumulus cell expansion and regulating extracellular matrix stability, thus facilitating ovulation. The identities of these oocyte-secreted growth factors regulating such key ovarian functions remain unknown, although growth differentiation factor-9 (GDF-9), GDF-9B and/or bone morphogenetic protein-6 (BMP 6) are likely candidate molecules, probably forming complex local interactions with other related members of the transforming growth factor-beta (TGF-beta) superfamily. Elucidating the nature of oocyte-somatic cell interactions at the various stages of follicle development will have important implications for our understanding of factors regulating folliculogenesis, ovulation rate and fecundity. PMID- 15271472 TI - Physiology of GDF9 and BMP15 signalling molecules. AB - Two related oocyte-derived members of the transforming growth factor-beta (TGF beta) superfamily, namely growth differentiation factor 9 (GDF9) and bone morphogenetic protein 15 (BMP15, also known as GDF9B), have recently been shown to be essential for ovarian follicular growth. In addition, both proteins have been shown to regulate ovulation rate in sheep, and although it is evident that these growth factors interact both with one another and with other intra- and extra-ovarian factors, the precise mechanisms by which they influence follicular growth and ovulation rate have not been thoroughly elucidated. PMID- 15271473 TI - Endocrine and paracrine control of follicular development and ovulation rate in farm species. AB - Productivity in farm species is controlled by many factors, including ovulation rate. In cattle, single ovulations occur most frequently and in sheep (and goats) the number of ova released can range from one to many depending upon the breed, whilst the pig is polyovular. The processes of recruitment and selection determine the number of ovulatory follicles in all these species with FSH and subsequently LH playing major roles. GnRH-agonist models in which endogenous gonadotrophin secretion is suppressed and exogenous LH and/or FSH are administered at specific concentrations in defined patterns, are useful in all three species for elucidating the precise roles of specific hormones in stimulating follicular development. Differences in the hypothalamic-pituitary ovarian feedback response lead to the differences in the number of ovulatory follicles, as does the pool of antral follicles from which the ovulatory ones are selected. Precocious development of follicles is also associated with more ovulations, as is the case with the Booroola due to the single gene acting through bone morphogenetic proteins (BMPs). It is well established that ovulation rate can also be influenced by exogenous hormone administration and by environmental factors such as nutrition. It has become apparent that these nuritional effects are mediated by a direct action at the level of the ovary, involving insulin, insulin-like growth factors (IGF) I and II and their binding proteins among other factors. These factors can also affect the quality of the oocyte and consequently embryo development and survival. Recently, the regulation of follicular angiogenesis has been shown to be important for the development of ovulatory follicles, particularly vascular endothelial growth factor (VEGF) which is produced primarily by the granulosa cells within the ovary and can be stimulated by gonadotrophins. Administration of VEGF has been shown to stimulate pre-antral follicular growth and increase the number of pre-ovulatory follicles. In summary both extra- and intra-ovarian factors are involved in the control of ovulation rate. Manipulation of the angiogenic process may also provide new opportunities for regulating the quality and number of follicles that ovulate. PMID- 15271474 TI - The use of hormonal treatments to improve reproductive performance of anestrous beef cattle in tropical climates. AB - Most of the world's bovine herd is found in tropical regions. Bos indicus predominates, due to their adaptation to the climate and management conditions. Anestrous is the main factor that negatively affects reproductive performance of animals bred in these regions of the globe. Several factors affect postpartum anestrous, including suckling and maternal-offspring bond, and pre- and postpartum nutritional status. The short duration of estrus and the tendency to show estrus during the night, greatly affect the efficiency of artificial insemination (AI) programs in B. indicus cattle managed in tropical areas. Several restricted suckling or weaning procedures (temporary or permanent), and hormonal treatments have been used to induce ovulation and cyclicity in postpartum cows. Most hormonal treatments are based on progesterone/progestogen (P4) releasing devices associated with estradiol benzoate (EB), or a combination of GnRH/PGF(2alpha)/GnRH (Ovsynch). Treatments with GnRH/PGF(2alpha)/GnRH has presented inconsistent results, probably due to the variable number of cows in anestrous. Treatments using P4 devices and EB have resulted in apparently more consistent results than Ovsynch programs in B. indicus cattle; however, pregnancy rates are low in herds presenting high anestrous rates and moderate to low body condition. The addition of an eCG treatment at the time of device removal, which increased plasma progesterone concentrations and pregnancy rates in anestrous postpartum suckled B. indicus cows, may be useful to improve reproductive performance of beef cattle in tropical climates. PMID- 15271475 TI - Hormonal induction of estrous cycles in anestrous Bos taurus beef cows. AB - A significant proportion of postpartum beef cows are anestrus at the onset of the breeding season. Much progress has been made in understanding anestrus and the changes that lead to spontaneous resumption of reproductive function. Likewise, knowledge regarding the impact of hormonal interventions on the endocrine and ovarian changes normally associated with spontaneous resumption of estrous cycles continue to accumulate. A wide range of hormonal treatment programs designed to induce estrous cycles in anestrous cows to coincide with the start of the breeding season have been developed. Programs structured to provide for increased progesterone, estradiol and LH concentrations at the appropriate times during the period leading to the first ovulation, and an induced preovulatory gonadotropin surge when the dominant ovarian follicle is of appropriate maturity have been demonstrated to induce estrous cycles of normal duration and acceptable fertility in a majority of anestrous, Bos taurus beef cows. PMID- 15271476 TI - The use of hormonal treatments to improve the reproductive performance of lactating dairy cows in feedlot or pasture-based management systems. AB - Hormonal interventions have been used to increase the probability of estrous detection and insemination, and to increase pregnancy rates of dairy cattle under a variety of management systems. The present review addresses the basic principles of hormonal intervention and presents typical examples that illustrate the methodology. The hormones used to control the estrous cycle mimic the reproductive hormones found within the normal cow. Most estrous synchronization systems employ a method for controlling follicular wave development, promoting ovulation in anestrous cows, regressing the corpus luteum in cyclic cows, and synchronizing estrus and (or) ovulation at the end of treatment. A wide range of reproductive systems are in place on dairy farms. In most herds, a non intervention period is practiced where postpartum cows are observed estrus estrus. Cows not observed in estrus are then treated. A number of studies in pasture-based and confinement systems have demonstrated net benefits of whole herd synchronization. Despite the advantages of whole-herd reproductive programs, their uptake has been inconsistent globally. The benefits of a timed artificial insemination (AI) system increase under conditions of poor estrous detection rate and poor conception rate. The unpopular nature of timed AI programs in pasture fed cows relates to high rates of estrous detection and conception for pasture based dairying. Regardless of production system, some cows must be re-inseminated because they are not pregnant after first insemination. The presence of "phantom cows" (non-pregnant cows that do not return to estrus) creates a serious reproductive challenge for both pasture-based and confinement-style operations. Early pregnancy diagnosis and second insemination timed AI may reduce the effects of phantom cows on dairy herds. Fundamental research into anestrous, the hormonal control of the estrous cycle, and early pregnancy detection should elucidate new methods that can be used to strengthen reproductive programs on dairy farms. PMID- 15271477 TI - The effect of embryonic death rates in cattle on the efficacy of estrus synchronization programs. AB - Reproductive failure in inseminated cattle results from poor fertilization and embryo survival. Recent studies utilizing dairy and beef cattle indicate that fertilization rates are higher for nulliparous dairy and beef heifers and nonlactating beef cows than lactating beef and dairy cows and nonlactating dairy cows. Several factors affect fertilization rates, but the greatest impact was observed for high producing cows under heat stress, when fertilization was only 55%. Once fertilization has occurred, the fate of a successful pregnancy is then determined by the survival of the embryo and fetus. Losses of pregnancy are characterized by early embryonic death, which occurs prior to the period of corpus luteum (CL) maintenance in the cow at days 15-17 of the cycle, and late embryonic death, which occurs from CL maintenance to the end of the differentiation stage, at approximately 42 days of gestation. After 50 days of gestation, pregnancy losses are less frequent and characterize fetal death. Most pregnancy losses occur prior to the period of maintenance of the CL, but in high producing lactating dairy cattle, substantial losses continue to occur up to 42 56 days after insemination. Several factors affect pregnancy losses in cattle, such as compromised oocytes, which result in poorly developed embryos incapable of cross-talking with the endometrial epithelial cells, to inadequate uterine environment and infectious agents resulting in death of the embryo from undernourishment. Recently, studies have indicated that anovulation/anestrous, the metabolic status of the animal, some dietary ingredients, as well as occurrence of diseases, predispose the cow to experience embryonic and fetal death. Although some insemination protocols might impact embryo survival, when timed AI has been implemented properly, it has not influenced embryonic or fetal death in cattle. Improvements in reproductive programs in the future will have to focus on enhancing fertilization rates and minimizing embryonic losses to optimize conception rates in dairy and beef cattle. PMID- 15271478 TI - Conceptus signals for establishment and maintenance of pregnancy. AB - Establishment and maintenance of pregnancy results from signaling by the conceptus (embryo/fetus and associated extraembryonic membranes) and requires progesterone produced by the corpus luteum. In most mammals, hormones produced by the trophoblast maintain progesterone production by acting directly or indirectly to maintain the corpus luteum. In domestic animals (ruminants and pigs), hormones from the trophoblast are antiluteolytic in that they act on the endometrium to prevent uterine release of luteolytic prostaglandin F2alpha. In cyclic and pregnant sheep, progesterone negatively autoregulates progesterone receptor gene expression in the endometrial luminal and superficial glandular epithelium. In cyclic sheep, loss of the progesterone receptor is closely followed by increases in epithelial estrogen receptors and then oxytocin receptors, allowing oxytocin to induce uterine release of luteolytic prostaglandin F2alpha pulses. In pregnant sheep, the conceptus trophoblast produces interferon tau that acts on the endometrium to inhibit transcription of the estrogen receptor alpha gene directly and the oxytocin receptor gene indirectly to abrogate development of the endometrial luteolytic mechanism. Subsequently, sequential, overlapping actions of progesterone, interferon tau, placental lactogen, and growth hormone comprise a hormonal servomechanism that regulates endometrial gland morphogenesis and terminal differentiated function to maintain pregnancy in sheep. In pigs, the conceptus trophoblast produces estrogen that alters the direction of prostaglandin F2alpha secretion from an endocrine to exocrine direction, thereby sequestering luteolytic prostaglandin F2alpha within the uterine lumen. Conceptus estrogen also increases expression of fibroblast growth factor 7 in the endometrial lumenal epithelium that, in turn, stimulates proliferation and differentiated functions of the trophectoderm, which expresses the fibroblast growth factor 7 receptor. Strategic manipulation of these physiological mechanisms may improve uterine capacity, conceptus survival, and reproductive health. PMID- 15271479 TI - Placental hormones and fetal-placental development. AB - Production of growth promoting substances by the placenta is regulated differently from the way production of similar compounds is regulated by maternal organs in various cases. Gene duplication is one of the mechanisms that facilitated the evolution of placental specific endocrine activity. Cattle, sheep and goats, although evolutionarily related, differ significantly from each other in the way their placental growth hormone (GH) and prolactin (PRL)-like hormones have evolved. Cattle carry one copy of the GH gene and there is no evidence yet for expression of that single GH gene copy in the placenta. On the other hand, the ovine GH gene has been duplicated and both oGH copies are expressed in the placenta during early stages of gestation. Prolactin gene duplication in ruminants resulted in the formation of specific placental-expressed prolactin related genes including the placental lactogen (PL) gene. In homologous state, ovine PL manifests PRL activity, but antagonizes GH activity. Ovine PL activity which can be mediated by PRL receptors or by hetero-dimerization of GH and PRL receptors, provide a novel regulatory mechanism for somatogenic activity dependent on the coexistence of both GH and PRL receptors in the same cells. Another mechanism for specific placental endocrine activity is silencing of the alleles through genetic imprinting. Disruption of genetic imprinting of placental genes has been proposed as one of the explanations for the loss of cloned fetuses generated by somatic cell nuclear transfer. PMID- 15271480 TI - Mechanisms responsible for parturition; the use of experimental models. AB - In this review, our knowledge, gleaned from a range of species, of what determines gestation length, how fetal maturation and birth are synchronized and how the uterotonic mechanisms are activated at birth are discussed. Accumulated data indicate that fetal glucocorticoids are involved in, but do not necessarily play a causative role in, the initiation of parturition in eutherian mammals generally. Present observations are consistent with a complex, positive regulatory interaction between estrogens, prostaglandins and oxytocin and are consistent with a role for prostaglandins as the final, common effector in myometrial activation. We are, however, left with the possibility that the initial mechanism for the timing of birth is encoded in the fetal genome and is closely linked to, and activated when, certain prerequisite developmental events have occurred in the fetus. Our understanding of these events in the sheep have led to its extensive use as an experimental model for the study of human clinical correlates of fetal maturation and development and the control of the initiation of parturition. PMID- 15271481 TI - Molecular regulation of blastocyst formation. AB - Preimplantation development encompasses the interval from insemination until embryo implantation and thus includes the 'freeliving' period of oviduct and uterine development. Formation of the blastocyst is required for implantation and establishment of pregnancy, and is a principal determinant of embryo quality prior to embryo transfer. Development through this period is regulated by the expression of specific gene families that encode for cell polarity, cell junctional, cytoskeletal, ion transporter, and water channel gene products that direct the acquisition of cell polarity and differentiation of the outer cells of the early embryo. This results in the formation of the trophectoderm, which is the first epithelium of development. This review considers the roles of each of these gene families in trophectoderm differentiation and blastocyst formation. The principal hypothesis under investigation is that blastocyst formation is regulated by a Na/K-ATPase-generated trans-trophectoderm ion gradient that promotes the accumulation of water across the epithelium. This, combined with the formation of the tight junction seal controlling paracellular movement of water between adjacent trophectoderm cells, results in the formation of a fluid-filled blastocyst cavity and the expansion of the blastocyst. Results from recent experiments, however, have cast some doubt on the role of Na/K-ATPase in mediating these events and have defined water channels or Aquaporins (AQPs) as physiological mediators of fluid movement across the trophectoderm. In addition, studies have now implicated mitogen-activated protein kinase (MAPK) signaling as an important mediator of development to the blastocyst stage. Such studies define the physiology of blastocyst formation and serve to support the application of assisted reproductive technologies (ART) to both human and animal species. PMID- 15271482 TI - Gene expression patterns in in vitro-produced and somatic nuclear transfer derived preimplantation bovine embryos: relationship to the large offspring syndrome? AB - A considerable proportion of the offspring born from somatic nuclear transfer (sNT)-derived and in vitro-produced (IVP) embryos, particularly in ruminants and mice, is affected by multiple abnormalities of which a high birth weight is the predominant feature; a phenomenon that has been called "large offspring syndrome (LOS)". The underlying mechanisms are largely unknown at present, but changes in epigenetic modifications occurring during preimplantation development resulting in perturbed embryonic and fetal gene expression patterns are thought to be involved in the syndrome. This review summarizes results from studies comparing mRNA expression patterns from IVP and sNT-derived embryos to those of their in vivo counterparts, which are regarded as the "gold standard". Numerous aberrations have been observed ranging from suppression of expression to de novo overexpression or more frequently to a significant up- or down-regulation of a specific gene. These observations emphasize the need for further studies during preimplantation embryo development to gain insight in the molecular, preferentially epigenetic, mechanisms regulating embryonic and fetal development. Understanding these mechanisms will help to improve biotechnologies applied to early embryos in all species including humans. PMID- 15271483 TI - Regulation of ribosomal RNA gene expression in porcine oocytes. AB - In vitro production (IVP) of porcine embryos including in vitro maturation (IVM) of oocytes followed by in vitro fertilization (IVF) and in vitro culture (IVC) of the resultant embryos may result in live offspring, but it is still associated with great inefficiencies probably due to incomplete cytoplasmic maturation of the oocytes in vitro. Therefore, fundamental knowledge on the regulation of transcription during the oocyte growth phase when the messengers and protein synthetic machinery necessary for oocyte developmental competence are formed, is of great importance. In mammals, synthesis of RNA, up to 60-70% of which is ribosomal (rRNA), increases during oocyte growth and reaches a peak at the beginning of follicular antrum formation. In oocytes at the end of the growth phase, acquisition of full meiotic competence coincides with a markedly decreased rRNA transcriptional activity in the gametes. Our recent studies on the porcine oocyte growth phase have revealed a deeper molecular and biological insight into the complex regulation of rRNA transcription at different stages of follicular development. The data indicate that the so-called pocket protein, p130, is involved in the down-regulation of rRNA transcription at the end of the oocyte growth phase through an inhibition of the action of upstream binding factor (UBF). The latter protein is necessary for the function of RNA polymerase I (RNA Pol I), which is the actual enzyme driving rRNA gene transcription. Moreover, rRNA transcription also appears to be down-regulated by a decrease in the expression of mRNA encoding PAF53, an RNA Pol I-associated factor also required for the polymerase to exert its action. At the ultrastructural level, these molecular changes are paralleled by marginalization of the fibrillar centres of the oocyte nucleolus followed by compaction of the nucleolus into an inactive sphere of fibrils. PMID- 15271484 TI - Oocyte transfer and gamete intrafallopian transfer in the mare. AB - Methods for the collection and transfer of equine oocytes have been developed, and uses of these techniques have resulted in new clinical and research possibilities. Because oocyte transfer avoids reproductive problems associated with the oviduct, uterus, and cervix, pregnancies can be produced from many mares that cannot carry a pregnancy or produce embryos. Oocytes for clinical transfers are usually collected from preovulatory follicles and cultured for a short interval or transferred directly into a recipient's oviduct. For oocyte transfer, the recipient is inseminated within the uterus. A large number (1 x 10(9) to 2 x 10(9)) of motile sperms are preferred for inseminations. In contrast, sperm and oocyte are transferred into the oviduct during gamete intrafallopian transfer (GIFT). Therefore, a lower number (1 x 10(5) to 2 x 10(5)) of sperm can be used. Potentially, GIFT could be used in situations where sperm numbers are limited. Use of oocyte transfer and GIFT in clinical and research settings will aid us in understanding the interactions between oocyte, sperm, and oviduct in the equine. PMID- 15271485 TI - Low dose insemination in the mare: an update. AB - The generally recommended minimum number of spermatozoa required for conventional artificial insemination in the mare is in excess of 200 x 10(6) progressively motile spermatozoa. Recent developments in different insemination techniques such as deep uterine, hysteroscopic and oviductal insemination, which have been designed to use significantly fewer spermatozoa, are reviewed in this paper. A number of studies have demonstrated that ultrasound guided deep uterine insemination of 5 x 10(6) fresh spermatozoa can produce satisfactory pregnancy rates. The use of hysteroscopic insemination enables the insemination dose to be successfully reduced to 1 x 10(6) fresh or 3 x 10(6) frozen-thawed spermatozoa. Further reduction of the insemination dose is possible and satisfactory pregnancy rates can also be achieved by surgical oviductal insemination of mares with as few as 2 x 10(5) fresh spermatozoa. The refinement of these insemination techniques will allow us to maximise the use of frozen-thawed semen, use new technology such as sex-preselection of spermatozoa and to conserve and utilise epididymal spermatozoa at the time of castration or death of a stallion. PMID- 15271486 TI - Suppressing reproductive activity in horses using GnRH vaccines, antagonists or agonists. AB - There are a number of situations in which it is desirable to suppress part or all of the reproductive endocrine system in a horse, notably the competing animal whose tractability during training, or performance during competition, is compromised by the expression of sexual or aggressive behavior. The current therapeutic approaches to reproductive endocrine suppression include gonadectomy and progestagen administration, where the former carries surgical risks and entails irreversible loss of breeding potential, and effective progestagen therapy requires frequent administration for extended periods and is banned in some competing animals as potentially anabolic. In this context, preventing the action of gonadotrophin releasing hormone (GnRH) by blocking its pituitary receptors is an attractive alternative for reversibly rendering mares anestrus or depressing testosterone secretion or spermatogenesis in stallions. This paper reviews the data on effects, efficacy, reversibility, and side effects of GnRH vaccines, antagonists, and agonists for suppressing reproductive activity in horses, within the context of their potential place in the pharmacological armoury of the veterinary clinician. PMID- 15271488 TI - Presynaptic induction and expression of NMDA-dependent LTP. AB - It is almost axiomatic in neuroscience that postsynaptically induced NMDA dependent long-term potentiation (LTP) is a cellular substrate of memory. As a result, memory impairments caused by experimental reductions of NMDA currents are often interpreted as evidence that this form of plasticity is involved in memory formation. However, a recent Nature study indicates that not all NMDA-dependent LTP is induced postsynaptically. PMID- 15271489 TI - Parsing brain activity with fMRI and mixed designs: what kind of a state is neuroimaging in? AB - Neuroimaging is often pilloried for providing little more than pretty pictures that simply show where activity occurs in the brain. Strong critics (notably Uttal) have even argued that neuroimaging is nothing more than a modern day version of phrenology: destined to fail, and fundamentally uninformative. Here, I make the opposite case, arguing that neuroimaging is in a vibrant and healthy state of development. As recent investigations of memory illustrate, when used well, neuroimaging goes beyond asking 'where' activity is occurring, to ask questions concerned more with 'what' functional role the activity reflects. PMID- 15271490 TI - Do spines and dendrites distribute dye evenly? PMID- 15271491 TI - Milestones of neuronal development in the adult hippocampus. AB - Adult hippocampal neurogenesis originates from precursor cells in the adult dentate gyrus and results in new granule cell neurons. We propose a model of the development that takes place between these two fixed points and identify several developmental milestones. From a presumably bipotent radial-glia-like stem cell (type-1 cell) with astrocytic properties, development progresses over at least two stages of amplifying lineage-determined progenitor cells (type-2 and type-3 cells) to early postmitotic and to mature neurons. The selection process, during which new neurons are recruited into function, and other regulatory influences differentially affect the different stages of development. PMID- 15271492 TI - Computational models of the basal ganglia: from robots to membranes. AB - With the rapid accumulation of neuroscientific data comes a pressing need to develop models that can explain the computational processes performed by the basal ganglia. Relevant biological information spans a range of structural levels, from the activity of neuronal membranes to the role of the basal ganglia in overt behavioural control. This viewpoint presents a framework for understanding the aims, limitations and methods for testing of computational models across all structural levels. We identify distinct modelling strategies that can deliver important and complementary insights into the nature of problems the basal ganglia have evolved to solve, and describe methods that are used to solve them. PMID- 15271493 TI - Corticostriatal plasticity: life after the depression. AB - Experience-dependent changes in corticostriatal transmission efficacy are likely to support the role of the striatum in reinforcement-based motor learning. Whereas long-term depression at glutamatergic corticostriatal synapses has long been regarded as the normal form of striatal plasticity, recent work provides evidence that use-dependent potentiation can naturally occur at these connections through an increase in both synaptic efficacy and postsynaptic intrinsic excitability. By decreasing the weight of cortical inputs required to fire striatal output neurons, short-term and long-term potentiation at corticostriatal connections can jointly participate in the formation of sensorimotor links by which specific context-dependent patterns of cortical activity can engage selected motor programs. PMID- 15271494 TI - Putting a spin on the dorsal-ventral divide of the striatum. AB - Since its conception three decades ago, the idea that the striatum consists of a dorsal sensorimotor part and a ventral portion processing limbic information has sparked a quest for functional correlates and anatomical characteristics of the striatal divisions. But this classic dorsal-ventral distinction might not offer the best view of striatal function. Anatomy and neurophysiology show that the two striatal areas have the same basic structure and that sharp boundaries are absent. Behaviorally, a distinction between dorsolateral and ventromedial seems most valid, in accordance with a mediolateral functional zonation imposed on the striatum by its excitatory cortical, thalamic and amygdaloid inputs. Therefore, this review presents a synthesis between the dorsal-ventral distinction and the more mediolateral-oriented functional striatal gradient. PMID- 15271495 TI - Correlating function and gene expression of individual basal ganglia neurons. AB - Functional studies at the level of individual neurons have greatly contributed to our current understanding of basal ganglia function and dysfunction. However, identification of the expressed genes responsible for these distinct neuronal phenotypes is less advanced. Qualitative and quantitative single-cell gene expression profiling, combined with electrophysiological analysis, allows phenotype-genotype correlations to be made for individual neurons. In this review, progress on gene-expression profiling of individual, functionally characterized basal ganglia neurons is discussed, focusing on ion channels and receptors. In addition, methodological issues are discussed and emerging novel techniques are introduced that will enable a genome-wide comparison of function and gene expression for individual neurons. PMID- 15271496 TI - Control of neuronal phenotype: what targets tell the cell bodies. AB - Assembly of neuronal circuits is controlled by the sequential acquisition of neuronal subpopulation-specific identities at progressive developmental steps. Whereas neuronal features involved in initial phases of differentiation are already established at cell-cycle exit, recent findings, based mainly on work in the peripheral nervous system, suggest that the timely integration of signals encountered en route to targets and from the target region itself is essential to control late steps in connectivity. As neurons project towards their targets they require target-derived signals to establish mature axonal projections and acquire neuronal traits such as the expression of distinct combinations of neurotransmitters. Recent evidence presented in this review shows that this principle, of a signaling interplay between target-derived signals and neuronal cell bodies, is often mediated through transcriptional events and is evolutionarily conserved. PMID- 15271497 TI - Energetic basis of brain activity: implications for neuroimaging. AB - The complex activities of the brain need not distract us from the certainty that it uses energy and performs work very efficiently. The human brain, which claims approximately 2% of our body mass, is responsible for approximately 20% of our body oxygen consumption. In vivo magnetic resonance spectroscopy (MRS) follows the metabolic pathways of energy production (as glucose oxidation) and work (as monitored by the cycling of glutamate and GABA neurotransmitters). In the resting awake state, approximately 80% of energy used by the brain supports events associated with neuronal firing and cycling of GABA and glutamate neurotransmitters. Small differences in neuronal activity between stimulation and control conditions can be measured and localized using functional magnetic resonance imaging (fMRI). MRS and fMRI experiments show that the majority of cerebral activity, which is often disregarded in imaging experiments, is ongoing even when the brain appears to be doing nothing. PMID- 15271498 TI - Viewing and doing: similar cortical mechanisms for perceptual and motor learning. AB - Historically, different groups of researchers have investigated the mechanisms of perceptual learning and motor learning. For sensory cortex, neurophysiological and psychophysical findings have linked changes in perception with altered neuronal tuning properties. However, less information has been forthcoming from motor cortex. This review compares recent findings on perceptual and motor learning, and suggests that similar mechanisms govern both. These mechanisms involve changes in both the center of neuronal tuning functions and their width or slope. The former reflects the values of the sensory or motor parameters that a neuron encodes, and the latter adjusts the encoding sensitivity. These similarities suggest that specific unifying principles for neural coding and computation exist across sensory and motor domains. PMID- 15271499 TI - Plasticity of interneuronal species diversity and parameter variance in neurological diseases. AB - Interneuronal diversity reflects the division of labor between numerous highly specialized interneuronal species, each performing a set of specific functions in neuronal networks. The rich diversity of interneurons found in the normal healthy brain is often significantly altered in neurological and psychiatric diseases. In genetic and developmental disorders, the diversity of interneuronal networks is compromised because of disturbances in the generation, specification or migration of specific interneuronal subtypes. Following insults related to trauma and seizures, the relative abundance of interneuronal subtypes might change, and entire interneuronal species can be lost from the network. In addition to the complete or partial loss of interneuronal subgroups, heterogeneity can also be altered in more subtle ways, as a result of changes in cell-to-cell variance of a particular parameter within specific interneuronal populations. Computational and experimental studies show that alterations in cellular and synaptic GABAergic heterogeneity can significantly modulate both firing rates and network coherence, indicating that plasticity of interneuronal diversity is likely to be an important mechanistic component of malfunctioning cortical networks in many pathological states. PMID- 15271500 TI - Interactions between metal ions and carbohydrates: the coordination behavior of neutral erythritol to zinc and europium nitrate. AB - The single crystals of coordinated complexes of neutral erythritol (C4H10O4) with zinc nitrate and europium nitrate were synthesized and studied using FT-IR and single crystal X-ray diffraction analysis. In the structure of Zn(NO3)2.C4H10O4, ZnEN (E denotes erythritol, N represents nitrate), Zn2+ is coordinated to four hydroxyl groups from two erythritol molecules and two oxygen atoms from two nitrates. Two Zn2+ are connected by one erythritol molecule to form Zn(C4H10O4)(NO3)2 chain, and layers formed by above chain pile to produce 3D structures. In the structure of Eu(NO3)3.C4H10O4.C2H5OH, EuEN, Eu3+ is 10 coordinated by six oxygen atoms from three nitrate ions, three hydroxyl groups from one erythritol molecule and one hydroxyl group from ethanol. In the above erythritol complexes, two hydroxyl groups of erythritol coordinate to one metal ion and the other two to another metal ion or erythritol acts as three-hydroxyl groups donor. The OH groups of erythritol act as ligand to coordinate to metal ions on one hand, one the other hand, OH groups form hydrogen bonds network to build three-dimensional structures. PMID- 15271501 TI - Studies on activities, cell uptake and DNA binding of four trans planaramineplatinum(II) complexes of the form: trans-PtL(NH3)Cl2, where L=2 hydroxypyridine, imidazole, 3-hydroxypyridine and imidazo(1,2-alpha)pyridine. AB - Four trans-planaramineplatinum(II) complexes code named YH9, YH10, YH11 and YH12 each of the form trans-PtL(NH(3))Cl(2), where L=2-hydroxypyridine and 3 hydroxypyridine, imidazole, and imidazo(1,2-alpha)pyridine for YH9, YH10, YH11 and YH12, respectively, have been synthesized and the activity of the compounds against human cancer cell lines, cell uptake, DNA-binding and nature of interaction with pBR322 plasmid DNA have been studied. The compound having imidazo(1,2-alpha)pyridine ligand as one the carrier ligands in the trans configuration is found to be significantly more active than cis-platin against ovarian A2780(cisR) cancer cell line corresponding with higher Pt-DNA binding. All other compounds have resistance factors less than that for cis-platin in the A2780 and A2780(cisR) cell lines. A greater prevention of BamH1 digestion with increasing concentration of the compounds indicates that as the compounds bind with nucleobases in DNA, the DNA conformation is changed sufficiently so as to prevent BamH1 digestion at the specific GG site. Gel electrophoresis results also indicate that as the compounds bind to DNA, unwinding of supercoiled form I DNA takes place to change it from the negatively supercoiled form I through relaxed circular form I to the positively supercoiled form I. PMID- 15271502 TI - Copper(II) Schiff base coordination compounds of dien with heterocyclic aldehydes and 2-amino-5-methyl-thiazole: synthesis, characterization, antiproliferative and antibacterial studies. Crystal structure of CudienOOCl2. AB - A new series of coordination compounds of the starting materials [Cu(dienX(2)Y(2))] and their adducts [Cu(dienXXY(2))(2a-5mt)] (where dien=diethylenetriamine, dienXX=Schiff bases of diethylenetriamine with 2 furaldehyde or 2-thiophene-carboxaldehyde, X=O, S, Y=Cl, Br, NO(3) and 2a-5mt=2 amino-5-methylthiazole) were synthesized by stepwise reactions and their structures were established by C, H, N, Cu analysis, spectroscopic, magnetic and molar conductivity measurements. The isolated compounds are monomers, paramagnetic and electrolytic compounds of the type 1:1. In all cases, the pentadentate Schiff base (dienXX) is bonded in a tridentate fashion through the 3 N atoms. In the CudienXXY(2) compounds the coordination sphere is completed by two Cl or Br or NO(3) groups in a square pyramidal arrangement. The proposed structure for this type of compound was further supported by X-ray diffraction analysis of the compound [Cu(dienOO)Cl(2)]. Its basal plane consists of three Cu N contacts [2.017(2), 2.025(2) and 2.012(2) A] from dienOO, and the Cl(1) atom, while the Cl(2) atom possesses the apical position, the relevant distances being 2.2732(7) A for Cu-Cl(1) and 2.6051(7) A Cu-Cl(2). In the CudienX(2)Y(2).2a-5mt adducts the coordination sphere of copper is further completed by the nitrogen ring atom of the 2a-5mt, forming an octahedral configuration. The study of the biological activity of the compounds synthesized against a panel of different normal and cancer cell lines (MRC5, HeLa, MCF7, HT-29, OAW42, T47D) and bacteria (E. coli, B. cereus, B. subtilis) showed that the adducts of the type [Cu(dienXXY(2))(2a-5mt)] exhibit increased activity both in cancer cells and in bacteria, compared to the starting material of type [Cu(dienXXY(2))]. PMID- 15271503 TI - Interactions between metal ions and carbohydrates: the coordination behavior of neutral erythritol to transition metal ions. AB - The single crystals of coordinated complexes of neutral erythritol (C4H10O4) with various transition metal ions were synthesized and studied using FT-IR and single crystal X-ray diffraction analysis. Two CuCl2-erythritol complexes (denoted as CuE(I) and CuE(II)) were obtained. In CuE(I), Cu2+ coordinates with two chloride ions and four OH groups from two erythritol molecules. Two copper centers are linked by one erythritol molecule to form a zigzag chain. For CuE(II), each Cu2+ coordinates with two OH groups from an erythritol molecule and two chloride ions. The crystal of CuE(II) contains complexed and free erythritol, the dimers of [Cu2Cl4(C4H10O4)] further form a [Cu2Cl4(C4H10O4)]infinity chain via secondary Cu...Cl bonds, both the dimer unit of [Cu2Cl4.(C4H10O4)] and non-coordinated C4H10O4 unit exist side by side in the crystal. MnCl2-erythritol complex whose structure is similar to CuE(I) is also acquired. The OH groups of erythritol act as ligand to coordinate to metal ions on one hand, one the other hand, OH groups form hydrogen bonds network that link chain and layer together to build three dimensional structures. PMID- 15271504 TI - Conformational analysis of octa- and tetrabromo tetraphenylporphyrins and their Ni(II) and Tb(III) complexes. AB - Molecular mechanics (MM) calculations were used to analyze the puckering of metalloporphyrins as a function of metal ion size and the position of substituents on the porphyrin periphery, on a three series of octa- and tetrabromo tetraphenylporphyrins: without metal, and with Ni(II), and Tb(III) as representative small and large metal ions, respectively. Molecular energy optimization calculations were carried out using the Consistent Force Field (CFF) program, with the parameters developed previously and new parameters for bromine atom. Normal-coordinate structural decomposition (NSD) analysis was performed on the equilibrium structures obtained by MM calculations. The conformers are also stereochemically characterized, compared with available X-ray structures and with the conformers obtained in our previous MM study using chloro instead of bromo beta-pyrrole substituents. PMID- 15271505 TI - Chromium-containing biomimetic cation triaqua-mu3-oxo-mu hexapropionatotrichromium(III) inhibits colorectal tumor formation in rats. AB - The cation [Cr(3)O(propionate)(6)(H(2)O)(3)](+) has recently been found by oral or intravenous administration to increase insulin sensitivity and improve serum lipids in healthy and type 2 diabetic model rats. Serum insulin concentrations and, hence, insulin sensitivity are in part responsible for the relationship between diet and the incidence of colorectal cancer. This strongly suggested that the synthetic cation, which in vitro is able to stimulate insulin receptor tyrosine kinase activity in a fashion similar to the oligopeptide chromodulin, could influence the incidence of colorectal cancer. Consequently, the effects of the cation on the inhibition of colon tumorigenesis induced by 1,2 dimethylhydrazine in male Sprague Dawley rats were examined. Gavage administration of aqueous solutions of the complex (1000 microg Cr/kg body mass daily for 6 months) resulted in significantly decreased colon tumor incidence (P<0.003). PMID- 15271506 TI - N-benzoyl-N'-alkylthioureas and their complexes with Ni(II), Co(III) and Pt(II) - crystal structure of 3-benzoyl-1-butyl-1-methyl-thiourea: activity against fungi and yeast. AB - The thiourea derivatives of N-butylmethylamine (3-benzoyl-1-butyl-1-methyl thiourea) (1), N-ethylisopropylamine (3-benzoyl-1-ethyl-1-isopropyl-thiourea) (2) and the corresponding complexes of 1 and 2 with Ni(II), Co(III) and Pt(II) have been synthesized. The compounds obtained were characterized by elemental analysis, spectroscopic methods (FT-IR, UV-Vis and NMR) and mass spectrometry. Compound 1, crystallized in the triclinic space group. The antifungal activities of compounds 1 and 2 and their corresponding complexes against the fungus Penicillium digitatum and against the yeast Saccharomyces cerevisiae were investigated. In general, fungal growth inhibition was higher with compound 1 and its complexes than with compound 2, except for the Co(III) complex of 2. PMID- 15271507 TI - Crystal structure and catecholase-like activity of a mononuclear complex [Cu(EDTB)]2+ (EDTB=N,N,N',N'-tetrakis(2'-benzimidazolyl methyl)-1,2 ethanediamine). AB - The crystal structure and catecholase-like activity of a mononuclear complex, Cu(EDTB)(NO3)2.C2H5OH (here EDTB stands N,N,N',N'-tetrakis(2'-benzimidazolyl methyl)-1,2-ethanediamine) has been studied in comparison with a binuclear complex Cu2(EDTB)(NO3)4.3H2O. The results show that the reactive rate constants increase with increases of reaction temperature and pH value of intermediate. Electrospray ionization mass spectrum (ESI-MS) shows that tautomerism isomers of catechol with the title complex exist in reaction solution, and catechol is oxidized to quinone, then it is further oxidized resulting in muconic acid and its derivatives via an intradiol mechanism, just like that catalyzed by a mononuclear non-heme iron-containing dioxygenase. PMID- 15271508 TI - Coordination mode of adenosine 5'-diphosphate in ternary systems containing Cu(II), Cd(II) or Hg(II) ions and polyamines. AB - The interactions of adenosine 5'-diphosphate (ADP) with some polyamines (PA) (1,3 diaminopropane (tn), 1,4-diaminobutane (Put), 1,7-diamino-4-azaheptane (3,3-tri) and 1,8-diamino-4-azaoctane (Spd)) both in presence and in the absence of metal ions (Cu(II), Cd(II) and Hg(II)) have been studied. In the metal-free systems the formation of adducts (ADP)Hx(PA) has been observed, in which the main reaction centres are the endocyclic nitrogen atoms of the purine ring, the phosphate group of the nucleotide and the protonated nitrogen atoms of the polyamine. The effectiveness of the phosphate group in formation of adducts has been found to decrease in the series Put > Spd > Spm and to be lower than in the reactions with shorter homologues of biogenic amines. In the ternary systems with metal ions the formation of molecular complexes (ML L' type) has been evidenced in which the protonated polyamine interacts with the nitrogen atoms N(1) or N(7) of the purine ring of the nucleotide. In the ternary systems Cu(II)/ADP/polyamine the coordination dichotomy observed in the binary system Cu(II)/ADP disappears. In the systems with Hg(II) ions the pH range of the dichotomy is extended, while for the systems Cd(II)/ADP/polyamine no changes of the range relative to the binary system Cd(II)/ADP have been noted. PMID- 15271509 TI - Inhibition of alcohol dehydrogenase by bismuth. AB - Bismuth compounds have been widely used for the treatment of ulcers and Helicobacter pylori infection, and enzyme inhibition was thought to be crucial for bismuth anti-microbial activity. We have investigated the interaction of colloidal bismuth subcitrate (CBS) with alcohol dehydrogenase and our results demonstrate that bismuth can effectively inhibit the enzyme. Kinetic analysis revealed that CBS acted as a non-competitive inhibitor of yeast alcohol dehydrogenase. Both UV-vis and fluorescence data show that interaction of CBS with the enzyme exhibits biphasic processes. Bismuth can replace only half of Zn(II) from the enzyme (i.e., about one Zn(II) per monomer). Surprisingly, binding of CBS also induces the enzyme dissociation from its native form, tetramer into dimers. The inhibition of Bi(III) on the enzyme is probably due to its direct interference with the zinc sites. This study is likely to provide an insight into the mechanism of action of bismuth drugs. PMID- 15271510 TI - Effect of cadmium on ferredoxin:NADP+ oxidoreductase activity. AB - Ferredoxin:NADP(+) oxidoreductase (FNR) was treated with cadmium and after that its diaphorase reaction in the presence of dibromothymoquinone (DBMIB) or ferricyanide (FeCy, K(3)Fe(CN)(6)) was examined. CdSO(4) (5 mM) caused 50% inhibition after half hour incubation. At least two components were distinguishable in the time-course inhibition, suggesting that more than one amino acid residues were engaged in reaction with the metal ion. The Lineweaver Burk plots indicate that Cd(2+) is an uncompetitive inhibitor for DBMIB reduction but exerts non-competitive inhibition for the NADPH oxidation. The FeCy reduction did not follow Michaelis-Menten kinetics. Zn(2+) diminished inhibitory effect of Cd(2+) on the DBMIB reduction but enhanced inhibition of the FeCy reduction. Incubation with additional chelator (beta-mercaptoethanol, or histidine) abolished inhibitory effect of Cd(2+) on the FeCy reduction but not on the DBMIB reduction. The mode of Cd(2+) action on the diaphorase activity of FNR in the presence of DBMIB or FeCy is briefly discussed with the special reference to the implication of two distinct sites at the FNR molecule, which might be involved in the reduction of various non-physiological substrates. PMID- 15271511 TI - Nucleophilic reaction by carbonic anhydrase model zinc compound: characterization of intermediates for CO2 hydration and phosphoester hydrolysis. AB - The partially hydrophilic and hydrophobic tripodal ligands, tris(hydroxy-2 benzimidazolylmethyl)amine L1h and tris(2-benzimidazolyl)amine L1 were used for the preparation of biomimetic complex of carbonic anhydrase. The CO(2) hydration using [L1hZn(OH)]ClO(4).1.5H(2)O provided the zinc-bound and free HCO(3)(-)s, which were formed by nucleophilic attack of Zn-OH toward CO(2) in dimethyl sulfoxide (DMSO). The phenolic OH in L1h can recognize water molecules through hydrogen bonds to facilitate the collection of the water molecules around a biomimetic zinc compound; the molecular structure of [L1hZn(OH)](+) was revealed. The packing diagram has demonstrated the all the water molecules are hydrogen bonded to each phenolic OH. The nucleophilic attack of zinc-bound OH(-) to substrate is used to catalyze the CO(2) hydration and phosphoester hydrolysis. The carbonic anhydrase model compound [L1Zn(OH(2))](2+) was applied for the hydrolysis of phosphoesters, parathion and bis(p-nitrophenyl)phosphate (BNPP(-)). The low reactivity of [L1Zn(OH)](+) for parathion hydrolysis is attributed to the stability of the intermediate [L1Zn(OP(S)(OEt)(2))](+). Since the structures of the intermediates [L1Zn(OH(2))](BNPP)(2) (1) and [L1Zn(OP(S)(OEt)(2))]ClO(4) (2) formed on the way of hydrolysis are too stable to realize the catalytic cycle and are not active for hydrolysis, carbonic anhydrase model compound [L1Zn(OH(2))](2+) was not suitable for phosphoester hydrolysis; the zinc model compound surrounded by three benzimidazolyl groups is used to have the steric hindrance for bulky substrate, such as parathion and BNPP(-). PMID- 15271512 TI - Synthesis, X-ray crystal structure, anti-fungal and anti-cancer activity of [Ag2(NH3)2(salH)2] (salH2=salicylic acid). AB - [Ag(2)(NH(3))(2)(salH)(2)] (salH(2)=salicylic acid) was synthesised from salicylic acid and Ag(2)O in concentrated aqueous NH(3) and the dimeric Ag(I) complex was characterised using X-ray crystallography. The complex is centrosymmetric with each metal coordinated to a salicylate carboxylate oxygen and to an ammonia nitrogen atom in an almost linear fashion. The two [Ag(NH(3))(salH)] units in the complex are linked by an Ag-Ag bond. Whilst metal free salH(2) did not prevent the growth of the fungal pathogen Candida albicans [Ag(2)(NH(3))(2)(salH)(2)], [Ag(2)(salH)(2)] and some simple Ag(I) salts greatly inhibited cell reproduction. SalH(2), [Ag(2)(NH(3))(2)(salH)(2)] [Ag(2)(salH)(2)] and AgClO(4) produced a dose-dependent cytotoxic response against the three human derived cancer cell lines, Cal-27, Hep-G2 and A-498, with the Ag(I)-containing reagents being the most effective. PMID- 15271513 TI - Mechanistic information on the reaction of cis- and trans-[RuCl2(DMSO)4] with d(T2GGT2) derived from MALDI-TOF and HPLC studies. AB - Reactions of trans and cis isomers of the Ru(II) complex [RuCl(2)(DMSO)(4)] with single-stranded hexanucleotide d(T(2)GGT(2)) were studied in aqueous solutions in the absence and presence of excess chloride by high performance liquid chromatography (HPLC) and matrix-assisted laser desorption/ionisation time of flight mass spectrometry (MALDI-TOF MS). Despite the different reactive species formed from the two isomers in aqueous solution, similar reaction products are obtained in their interaction with d(T(2)GGT(2)). Both [RuCl(2)(DMSO)(4)] isomers bind to the oligonucleotide in the bidentate mode to form thermodynamically stable bis-guanosine adducts, Ru(G-N7)(2). Significant differences were observed in the reaction rates, however the reaction with trans- [RuCl(2)(DMSO)(4)] is ca. 5-10 times faster in comparison to that observed for the cis analogue. This difference is interpreted in terms of different rate-limiting steps for the trans and cis complexes, respectively. It is suggested that the rate of the reaction with the trans isomer is controlled by dissociation of a Cl(-) ligand from the initially formed trans,cis,cis-[RuCl(2)(DMSO)(2)(H(2)O)(2)]. In the contrast, release of a dimethyl sulfoxide molecule from the reactive species cis,fac [RuCl(2)(DMSO)(3)(H(2)O)] is likely to be rate limiting for the cis analogue. Significant influence of electrostatic interactions on the reaction rate was observed for the trans isomer. Mechanistic interpretation of the observed reactivity trends based on data obtained from UV-Vis spectroscopy, HPLC and MALDI TOF MS studies is presented and discussed within the paper. PMID- 15271514 TI - Cytotoxicity of some platinum(IV) complexes with ethylenediamine-N,N'-di-3 propionato ligand. AB - The cytotoxicities of two platinum(IV) complexes of formula [PtX2(eddp)].nH2O (eddp=ethylenediamine-N,N'-di-3-propionate, X=chloro [I] or bromo [II], n=1 or 1.24) are reported. The complexes have been obtained by direct reaction of potassium hexahaloplatinate(IV) with H2eddp.2HCl followed by addition of a base (LiOH). The crystal and molecular structure has confirmed that the complex with bromo ligands, similarly to the complex with chloro ligands previously reported, has trans configuration of the halogens. In both the chloro and bromo complexes there appear to be intramolecular N-H...X interactions which account for a narrowing of the corresponding X-Pt-N angles below 90degrees. The trans isomer (configuration index OC-6-13, two nitrogens and two oxygens of eddp bound in the equatorial plane) is the only one obtained in the reaction of hexahaloplatinate(IV) with the eddp ligand while a similar reaction performed with ethylenediamine-N,N'-diacetate (edda) affords exclusively the symmetrical cis-isomer (configuration index OC-6-33, equatorial nitrogen and axial oxygen atoms of edda). The longer chain of the propionato groups (as compared to the acetato ones) is responsible for such a change in preferred configuration. NMR data have revealed a very large diastereotopic splitting of the propionato methylene protons to the nitrogens (0.88 ppm). The trans disposition of the halogen ligands in the compounds with eddp leads to deactivation of platinum(IV) complexes in comparison to those with edda having cis disposition of the leaving chlorides (human ovarian cancer cell line A2780, IC50 [muM] of 92.6 +/- 12 and 30.3 +/- 7.5 for [I] and [II], respectively). PMID- 15271515 TI - Antiviral properties and cytotoxic activity of platinum(II) complexes with 1,10 phenanthrolines and acyclovir or penciclovir. AB - Platinum compounds containing an aromatic diimine (1,10-phenanthroline or 2,9 dimethyl-1,10-phenanthroline; phen and Me(2)phen, respectively) and antiviral guanosine-type ligands (acyclovir or penciclovir; acy and pen, respectively) have been synthesised. These compounds maintain the antiviral activity against Herpes Symplex Virus (HSV) and have greater efficacy than free acyclovir or penciclovir against Cytomegalovirus (CMV); in both cases the species with Me(2)phen are more active. The same complexes are effective against tumor cell proliferation which also results to be dependent upon the nature of the diimine ligand: all compounds containing Me(2)phen being more active than those containing phen. Although in vivo some complexes significantly reduce tumor cell proliferation, nevertheless, they do not appear to significantly affect the life time expectancy of the treated mice. The greater cytotoxicity of compounds with Me(2)phen may result from a higher reactivity towards cellular components, such as glutathione, which could cause release of the diimine, known to be highly cytotoxic. PMID- 15271516 TI - Coordination ability of pentapeptides with two dehydro-amino acid residues inserted into their sequences. AB - The study on the binding ability of tested ligands have shown that insertion of two dehydro-amino acid residues into peptide sequences makes them more effective in metal ion binding than ligands with one dehydro-amino acid residue. The ligand with two Z(Delta)Phe residue form more stable complexes than his analogues with one Z(Delta)Phe residue. Interesting is this that position of Z(Delta)Phe residue in peptide chain have impact on Cu(II)-complexes formation. PMID- 15271517 TI - Homoleptic ruthenium (II) complexes containing asymmetric tridentate 2 (benzimidazole-2-yl)-1,10-phenanthroline likes ligands: syntheses, characterization and DNA binding. AB - Two novel homoleptic ruthenium (II) complexes containing asymmetric tridentate ligands, 2-(benzimidazole-2-yl)-1,10-phenanthroline (PHBI) and 2-(naphtho[2,3 d]imidazole-2-yl)-1,10-phenanthroline (PHNI) have been synthesized and characterized by elemental analysis, mass spectra, (1)H NMR, and electronic spectroscopy. The DNA-binding properties of the complexes have been investigated by spectroscopic methods and viscosity measurements. The results indicate that the two complexes interact with DNA via electrostatic interaction, and the mechanisms of DNA binding with the complexes have also been discussed. PMID- 15271518 TI - DNA-binding and photoactivated enantiospecific cleavage of chiral polypyridyl ruthenium(II) complexes. AB - A series of enantiomeric polypyridyl ruthenium(II) complexes Delta- and Lambda [Ru(bpy)2CNOIP](PF6)2 (Delta-1 and Lambda-1; BPY=2,2'-bipyridine, CNOIP=2-(2 chloro-5-nitrophenyl)imidazo[4,5-f][1,10]phenanthroline), Delta- and Lambda [Ru(bpy)2HPIP](PF6)2 (Delta-2 and Lambda-2; HPIP=2-(2-hydroxyphenyl)imidazo[4,5 f][1,10]phenanthroline), Delta- and Lambda-[Ru(bpy)2DPPZ](PF6)2 (Delta-3 and Lambda-3; DPPZ=dipyrido[3,2:a-2',3':c]-phenazine), Delta- and Lambda [Ru(bpy)2TAPTP](PF6)2 (Delta-4 and Lambda-4; TAPTP=4,5,9,18 tetraazaphenanthreno[9,10-b] triphenylene) have been synthesized. Binding of these chiral complexes to calf thymus DNA has been studied by spectroscopic methods, viscosity, and equilibrium dialysis. The experimental results indicated that all the enantiomers of these complexes bound to DNA through an intercalative mode, but the binding affinity of each chiral complex to DNA was different due to the different shape and planarity of the intercalative ligand. After binding to DNA, the luminescence property of complex 1 was distinctly different from complexes 2 to 4. Upon irradiation at 302 nm, complexes 2-4 were found to promote the cleavage of plasmid pBR 322 DNA from supercoiled form I to nicked form II, and obvious enantioselectively was observed on DNA cleavage for the enantiomers of complexes 2 and 4. The mechanisms for DNA cleavage by these enantiomeric complexes were also proposed. PMID- 15271519 TI - Zinc and cadmium specifically interfere with RNA-binding activity of human iron regulatory protein 1. AB - The cellular pro-oxidative stress induced by high zinc concentrations or cadmium is most likely mediated by disruption of redox (mainly thiol) homeostasis or by mishandling of redox-active transition metals. The impact of zinc and cadmium on the main regulators of iron homeostasis in metazoans, the iron regulatory proteins (IRP) 1 and 2, has been probed with the human recombinant proteins. Using purified proteins or extracts of yeast producing human IRP, zinc and cadmium were shown to interfere with the IRE-binding activity of IRP1, but not with that of IRP2 or the aconitase activity of IRP1. The IRP1 active site cysteines in positions 437, 503 and 506 were not directly involved in the effects of zinc and cadmium. The loss of RNA-binding activity is due to the reversible and specific aggregation of the IRP1 apoprotein with zinc and cadmium, since precipitation did not occur with other divalent metals such as manganese, cobalt or magnesium. The reported data suggest a new mechanism for the biological toxicity of cadmium and high zinc concentrations by interference with iron metabolism. PMID- 15271520 TI - 1H NMR relaxometric characterization of bovine lactoferrin. AB - Lactoferrin (Lf) is a mammalian iron binding protein present in external secretions and in polymorphonuclear leukocytes. Its role in host defense mechanisms related to the non-immune defense system has been definitively established. Lf has two identical iron-binding sites, far from each other (44.3 A) and magnetically non-interacting. Fe(III) ions are six-coordinated, with four donor atoms provided by protein sidechains (two Tyr, one His, one Asp) and two oxygen atoms from a bridged HCO(3)(-). This set of ligands provides an ideal coordination scheme for stable and reversible iron binding. Nuclear magnetic relaxation dispersion (NMRD) profiles of Lf are consistent with a closest distance for a single water hydrogen atom of 3.1 A. By looking at the X-ray structure of Lf (PDB ID code: 1BLF) we can locate two water oxygens at 3.95 and 4.27 A from each Fe(III), respectively. Temperature dependence data suggest that an important contribution to the overall paramagnetic contribution to the solvent water relaxation rate arises from one or more second sphere water molecules in slow exchange with the bulk. A decreasing value of the exchange rate is obtained, ranging from 1.2 to 0.7 micros in the observed temperature range (25-65 degrees C), with an activation enthalpy of 7.3+/-0.8 kJ mol(-1). The low exchange rate obtained from NMRD data can be explained by the observation that both water molecules are bound to several polar groups of the protein backbone and side chains. By increasing the pH from 6.5 to 12 two distinct titrations are observed, consistent with sequential removal of both water molecules. PMID- 15271521 TI - A peptide model of the copper-binding region of lysyl oxidase. AB - The copper-binding site of lysyl oxidase remains extremely poorly characterized and although models have been suggested for copper(II) coordination by three histidine ligands, as has been found for other copper-containing amine oxidases, there has been no experimental confirmation of these suggestions. In this work, two synthetic peptides with 24 and 34-amino acid residues, respectively, were chosen from the highly conserved histidine-rich sequence previously suggested as the copper-binding region of lysyl oxidase. These peptides each bind one equivalent of Cu(II), at the same site in the two peptides. Spectroscopic (NMR, electron paramagnetic resonance (EPR), CD, visible absorption and fluorescence) techniques were employed to investigate the nature of the resulting complexes. The results indicate that at neutral pH three histidine ring nitrogen atoms and one carboxylate oxygen atom coordinate as the in-plane ligands of the copper, which is in an approximately tetragonally-distorted octahedral geometry. Modeling of the copper-peptides using the consistent force field (CFF91) produces a minimum energy configuration with three histidines and one water molecule as the copper ligands. CD, EPR and fluorescence results are reported for lysyl oxidase and compared with results for the peptides. PMID- 15271522 TI - Syntheses, crystal structures, and oxidative DNA cleavage of some Cu(II) complexes of 5-amino-3-pyridin-2-yl-1,2,4-triazole. AB - Three new monomeric Cu(II) complexes of 5-amino-3-pyridin-2-yl-1,2,4-triazole (Hapt), [Cu(Hapt)(H(2)O)(2)(SO(4))] (1), [Cu(Hapt)(2)(H(2)O)(NO(3))](NO(3)) (2), and [Cu(Hapt)(2)(NCS-N)](NCS).H(2)O (3), have been prepared and characterized by single crystal X-ray diffraction. One distorted [CuN(2)O(2)+O(')] square pyramidal (1), one distorted [CuN(3)O+N(')+O(')] octahedral (2), and one distorted [CuN(4)+N(')] intermediate between square-pyramidal and trigonal bipyramidal (3) coordination configuration were found and are suggested to be due to the chelating nature of the ligand, which interacts with Cu(II) through the N4(triazole) and N(pyridine) atoms. Spectral properties of these chelates are in accordance with the X-ray structural data. With ascorbate and H(2)O(2) activation, compound 2 exhibits higher nuclease activity than compound 1. The influence on the DNA cleavage process of different scavengers of reactive oxygen species: dimethyl sulfoxide (DMSO), tert-butyl alcohol, sodium azide, 2,2,6,6 tetramethyl-4-piperidone and superoxide dismutase enzyme (SOD), and of the minor groove binder distamycin, is also studied. PMID- 15271523 TI - Synthesis, structural, physico-chemical and biological properties of new palladium(II) complexes with 2,6-dimethyl-4-nitropyridine. AB - This paper describes the synthesis and properties of two new palladium(II) complexes with 2,6-dimethyl-4-nitro-pyridine (dmnp): mononuclear [Pd(dmnp)2Cl2] and dinuclear [Pd2(dmnp)2Cl4]. Complexes were characterized on the basis of chemical and chromatographic analyses, MS and conductometric measurements, as well as by IR and NMR (1H and 13C) spectral studies. The crystal structures of ligand and mononuclear complex, trans-dichlorobis(2,6-dimethyl-4-nitro pyridine)palladium(II), were determined by three-dimensional X-ray methods. The crystals of both compounds are monoclinic, space groups P21/c with a=19.075(4), b=5.419(1), c=15.045(3) A and beta=108.15(3)degrees for (dmnp), and a=7.544(2), b=14.509(3), c=8.032(2) A and beta=90.32(3)degrees for [Pd(dmnp)2Cl2]. In the (dmnp) there are two crystallographically independent molecules in the unit cell. The nitro groups and methyl C atoms are coplanar with the ring plane. The hydrogen bond of the type C-H...O links the molecules into pairs around center of symmetry. These dimers are held together by contacts of the van der Waals type. In the crystal structure of [Pd(dmnp)2Cl2] the Pd atom lies on an inversion center and is four-coordinated by two pyridine N atoms and by two Cl atoms in trans positions. The coordination geometry is square-planar, with Pd-N and Pd-Cl distances of 2.033(2) and 2.311(1) A, respectively. The two pyridine rings are mutually parallel, but they are twisted from the PdN2Cl2 coordination plane by about 88.5degrees. The preliminary assessments of anti-tumor properties of both complexes and ligand were evaluated as in vitro anti-proliferative activity in four human cancer cell lines: SW707 (adenocarcinoma of the rectum), T47D (breast cancer), HCV (bladder cancer) and A549 (non-small cell lung carcinoma). The [Pd(dmnp)2Cl2] exhibits strong cytotoxic activity against all cell lines whereas the free ligand and dinuclear [Pd2(dmnp)2Cl4] are only moderate active. PMID- 15271524 TI - The kinetics and mechanisms of reactions of iron(III) with caffeic acid, chlorogenic acid, sinapic acid, ferulic acid and naringin. AB - The kinetics and mechanisms of the reactions of iron(III) with the hydroxy cinnamic acid based ligands caffeic, chlorogenic, sinapic and ferulic acids and the flavonoid naringin have been investigated in aqueous solution. The mechanisms for caffeic and chlorogenic acid are generally consistent with the formation of a 1:1 complex that subsequently decays through an electron transfer reaction. On reaction with iron(III), ferulic and sinapic acids undergo an electron transfer without the prior formation of any complex. There was no evidence of electron transfer occurring in the complex formed when iron(III) is reacted with naringin. Rate constants for k1 (formation) and k(-1) (dissociation) have been evaluated for the complex formation reactions of [Fe(H2O)6(OH)]2+ with caffeic acid, chlorogenic acid and naringin. Analysis of the kinetic data yielded stability constants, equilibrium constants for protonation of the iron(III) chlorogenic acid complex initially formed, together with the rate constants for complex decomposition through intramolecular electron transfers and in the case of caffeic acid and chlorogenic acid, rate constants for the iron(III) assisted decomposition of the initial complex formed. Some of the suggested mechanisms and calculated rate constants are validated by calculations carried out using global analysis of time dependent spectra. PMID- 15271525 TI - Non-chelating inhibition of the H101N variant of human liver arginase by EDTA. AB - Recombinant wild-type human liver arginase (EC 3.5.3.1) expressed in Escherichia coli was markedly resistant to inhibition by ethylene diamine tetraacetic acid (EDTA). In contrast, half and fully activated species of the H101N variant were totally inactive in the presence of approximately 1 mM EDTA. Dilution of inhibited species in metal-free buffer lead to a time dependent recovery of activity, even when measured in the absence of added Mn2+. The inhibition was mixed type, with predominance of a competitive component (Kii=0.31 mM; Kis=0.022 mM). The structurally related N,N,N',N'-tetramethylethylenediamine was not inhibitory, indicating the importance of the carboxyl groups in EDTA inhibition. We conclude that EDTA inhibition of H101N arginase is not due to interaction with a weakly bound Mn2+ or chelation of essential metal ions. PMID- 15271526 TI - Shame and scandal: Clinical and Canon Law perspectives on the crisis in the priesthood. PMID- 15271527 TI - Characteristics of adolescent females in juvenile detention. PMID- 15271528 TI - Child homicide and Infanticide in New Zealand. PMID- 15271529 TI - A structured clinical model for violence risk intervention. PMID- 15271530 TI - Duty, burden, and ambivalence: families of forensic psychiatric patients in Hong Kong. PMID- 15271531 TI - The relationship between defendant's social attributes, psychiatric assessment and sentencing--a case study in Switzerland. PMID- 15271532 TI - Length of incarceration: was there parity for mentally ill offenders? PMID- 15271533 TI - Immunologic factors in the pathogenesis of osteonecrosis. AB - Osteonecrosis is a disease in which death of cellular elements of bone occurs as a result of diminished arterial blood supply. The pathogenetic mechanisms of osteonecrosis remain unresolved. Extravascular pressure and subsequent tamponade of the arterial vessels or an intravascular thrombosis have been suggested. Immunologic factors may also play an important role. In autoimmune disorders, small vessel vasculitis or other disease-associated features, as well as antiphospholipid antibodies, have been involved in the development of osteonecrosis. PMID- 15271534 TI - Coagulation abnormalities in patients with hip osteonecrosis. AB - The relatively high frequency of coagulation abnormalities in patients with hip osteonecrosis might represent increased risk factors for the development of bone necrosis by predisposing these patients to thromboembolic phenomena. The recognition of this association may increase as more patients with osteonecrotic lesions are tested for haemostatic abnormalities. Early diagnosis of hypercoagulability in the group of patients at risk may allow pharmacologic intervention that may prevent this devastating process from developing. PMID- 15271535 TI - Classification systems for osteonecrosis: an overview. AB - Currently a number of classification systems for osteonecrosis are in use. The use of different systems often leads to confusion and makes it difficult to compare the results of different methods of treatment. Because the management of osteonecrosis is determined in large part by the stage of the disease, it is important to use an effective and reliable method of staging and classification. This article provides an overview of the systems most commonly used so the reader can better understand and compare the outcome of studies that report their results using different methods of classification. The essential features of the ideal system are outlined to enable the reader to decide which of the available classification systems best meets these goals. PMID- 15271536 TI - Vascular anatomy and microcirculation of skeletal zones vulnerable to osteonecrosis: vascularization of the femoral head. AB - Bone, regardless of its type or location, is a highly vascular structure with unique features in its internal blood flow. Changes that occur in blood flow through bone have important implications in disease, and several attempts have been made to correlate vascular patterns with the clinical incidence of osteonecrosis. Examination of the arterial anatomy of bones that undergo osteonecrosis in other regions of the body has allowed identification of types of vascular interruptions that place particular bones at risk. Although the role of an impaired blood supply of the femoral head in the pathogenesis of osteonecrosis has not been clarified, several studies have found abnormal blood supply in patients with osteonecrosis. PMID- 15271537 TI - Imaging of early stages of osteonecrosis of the knee. AB - Osteonecrosis of the knee can present as a spontaneous and primary or a secondary clinical entity. The natural history of osteonecrosis follows a course of several sequential stages, and the later stages of both entities seem to be irreversible. Early diagnosis of osteonecrosis is crucial: the earlier the stage of the lesion at the time of diagnosis, the better the prognosis.Clinically, early diagnosis and treatment of osteonecrosis might prevent unnecessary surgery in cases with a concomitant degenerative meniscal tear. Early-stage osteonecrosis should be ruled out before surgery, because arthroscopy has lately been associated with osteonecrosis. Not every imaging method is equally suitable for detecting pathognomonic changes in each stage of osteonecrosis. Early-stage osteonecrosis is difficult to diagnose,because various differential diagnoses must be kept in mind. Moreover, there is a diagnostic window between the onset of symptoms and the appearance of pathognomonic changes on plain radiographs and MRI. PMID- 15271538 TI - Analysis of failures after vascularized fibular grafting in femoral head necrosis. AB - Evaluation of graft-host bone interactions after failed vascularized fibular grafting of femoral head necrosis may elucidate the reasons for failure of the procedure. According to the authors' study, the vascularized fibula implanted into the femoral head before collapse has the potential for restructuring the major segment of the affected head and delaying joint degeneration for many years if circumferential graft-host union is established. Asymmetric bone healing and non-union between the graft and the necrotic subchondral bone in the weight bearing area lead to failure, progression of symptoms, and subsequent early hip replacement. PMID- 15271539 TI - Pathologic conditions mimicking osteonecrosis. AB - MRI has become increasingly helpful in establishing an early diagnosis of avascular necrosis(AVN). AVN often demonstrates a classic pattern on MRI; findings earlier in the course of the disease are less specific. Many pitfalls can complicate interpretation, and a number of pathologic conditions can share features of early AVN on MRI and plain radiographs. These entities should be distinguished from AVN, because treatment and prognosis may differ significantly. PMID- 15271540 TI - Painful bone marrow edema of the knee: differential diagnosis and therapeutic concepts. AB - Bone marrow edema (BME) is a common finding when patients with knee pain are evaluated by MRI. The typical MRI signal patterns for BME are nonspecific, however. This article categorizes painful BME of the knee joint into three distinct etiologic groups and briefly describes therapeutic approaches for each of the 12 different types of BME. PMID- 15271541 TI - Non-union of femoral neck fractures with osteonecrosis of the femoral head: treatment with combined free vascularized fibular grafting and subtrochanteric valgus osteotomy. AB - Femoral neck fractures, frequently complicated by non-union and femoral head osteonecrosis,present a difficult clinical situation, especially when young patients are concerned. Existing treatment options are valgus osteotomy to address the biomechanical factors or bone grafting to address the biologic factor. The authors describe the operative technique and results of combined subtrochanteric valgus osteotomy and free vascularized fibular grafting in management of five young patients with both non-union and avascular necrosis. PMID- 15271542 TI - Can rotation osteotomy remain effective for more than ten years? AB - Trochanteric rotation osteotomies displace the necrotic zone of the femoral head outside the major acetabular weight-bearing zone and rotate the head anteriorly or posteriorly. Nineteen consecutive patients were selected for rotation osteotomy based on age,absence of progressive disease, and preoperative imaging studies predicting that rotation osteotomy would move the entire necrotic zone away from the acetabular roof. Factors associated with failure were head flattening and necrosis deeper than one third of the femoral head diameter. Among patient subsets with identical disease stages, outcomes seemed better after posterior rotation than after anterior rotation. Rotation osteotomies,fixed by a nail plate, can be recommended in a few selected patients with shallow necrosis involving less than one third of the femoral head diameter and without osteoarthritis or head flattening. Under these conditions, good outcomes may be achieved for 10 years or longer. PMID- 15271543 TI - Results of free vascularized fibular grafting for femoral head osteonecrosis in patients with systemic lupus erythematosus. AB - Osteonecrosis of the femoral head is a known complication of systemic lupus erythematosus. Patients often require surgical intervention at an early age. Options include total hip arthroplasty, core decompression, and free vascularized fibular grafting. Because these patients are usually young, any procedure that avoids total hip arthroplasty, especially if it does not make joint replacement more difficult, would be most desirable. Free vascularized fibular grafting is an effective method of treating osteonecrosis of the femoral head. Inpatients with lupus erythematosus, results of free vascularized fibular grafting at a minimum 2 year follow-up are similar to those in patients without a diagnosis of lupus. PMID- 15271544 TI - Effectiveness of total hip arthroplasty in the management of hip osteonecrosis. AB - Total hip replacement initially showed universally bad results when performed in hips with advanced stages of osteonecrosis. Newer techniques and implants remarkably improved these results. Today cementless or hybrid total hip arthroplasty for osteonecrosis is proven to be safe and effective and to have survivorship similar to cases with osteoarthritis. Newer,more durable bearing surfaces will further improve the longevity of this procedure. PMID- 15271545 TI - Spontaneous osteonecrosis of the knee: tibial plateaus. AB - Spontaneous osteonecrosis of the medial tibial plateau is less recognized than osteonecrosis of the medial femoral condyle, but it presents in a similar manner. These patients have a sudden onset of pain on the medial side of the knee associated with a spectrum of MRI changes in the tibial subchondral bone. The small lesions can resolve with only minimal residual scar remaining in the subchondral zone. If the lesion is large, it can collapse or show MRI changes of osteonecrosis. Recognition of this problem may help avoid unnecessary intra articular surgical intervention. PMID- 15271546 TI - Diagnosis and management of the osteonecrotic triad of the knee. AB - Generalized osteonecrosis of the knee may include, in addition to osteonecrosis of the medial femoral condyle that occurs most frequently, osteonecrosis of the patella or the tibial plateau. Such involvement is known as the osteonecrotic triad of the knee. Although the clinical picture of idiopathic osteonecrosis of the medial femoral condyle seems similar to several other disorders, certain distinct features, including its typical location,clinical symptoms, and late onset of cartilaginous erosion, facilitate differential diagnosis. Despite the progress made in the diagnosis and treatment of idiopathic osteonecrosis of the medial femoral condyle, the prognosis remains severe. More than 80% of the patients deteriorate to the extent that surgical reconstruction is necessary, whereas only about 20%of the patients demonstrate spontaneous resolution or no additional deterioration of the osteonecrotic lesion. PMID- 15271547 TI - Avascular necrosis of the talus. AB - Avascular necrosis (AVN) of the talus has always been a surgical challenge because the talus is hidden by its anatomic location and has a precarious blood supply. Most cases (75%) of talar AVN are traumatically induced in association with talar body and talar neck fractures.AVN of the talus can be a significant problem because collapse of the talar dome leads to degenerative changes and pain and disability of the ankle and subtalar joints. Although there are many published treatments for posttraumatic AVN of the talus, critical outcome studies are still lacking. PMID- 15271548 TI - Pathogenesis and treatment of osteonecrosis of the shoulder. AB - Osteonecrosis of the humeral head is an uncommon condition that occurs when bone vascularity is impaired. The causes underlying osteonecrosis of the humeral head seem to be multifactorial and can be traumatic or nontraumatic. Although clinical symptoms and causative factors affect the treatment of each patient, staging is the most objective criterion in deciding on the most appropriate treatment. Conservative treatment can be effective in stages I and II. Surgical intervention is advocated when symptoms are persistent and signs of collapse are evident. The surgical options include arthroscopic debridement, core decompression, vascularized bone grafting, and shoulder arthroplasty. PMID- 15271549 TI - Vascularity and osteonecrosis of the wrist. AB - This article outlines the vascular anatomy of the carpus, describing the extraosseous and intraosseus vascular systems and emphasizing the carpal bones at risk for osteonecrosis. Separate discussions of etiology, diagnosis, and treatment of osteonecrosis of the commonly involved carpal bones are included. PMID- 15271550 TI - Osteonecrosis of the human skeleton. PMID- 15271551 TI - Thermodynamic analysis of lysozyme adsorbed to silica. AB - The structural stability of hen egg white lysozyme in solution and adsorbed to small colloidal silica particles at various surface concentrations was investigated using hydrogen-deuterium (H/D) exchange in combination with mass spectrometry (HDX-MS) and differential scanning calorimetry (DSC). The combination of HDX-MS and DSC allows a full thermodynamic analysis of the lysozyme structure as both the enthalpy and the Gibbs free energy can be derived from the various measurements. Moreover, both HDX-MS and DSC provide information on the relative structural heterogeneity of lysozyme in the adsorbed state compared to that in solution. Results demonstrated that at high surface coverage, the structural stability of lysozyme was only marginally affected by adsorption to silica particles whereas the unfolding enthalpy decreased by more than 10%, meaning that the entropy of lysozyme increased with a similar value upon adsorption. Furthermore, the structural heterogeneity increased considerably. At lower surface concentrations, the structural heterogeneity increased further whereas the enthalpy of unfolding decreased. Further analyses of the HDX-MS experiments clearly indicated that folding/unfolding of lysozyme occurs through a two-domain process. These two domains had a similar amount of structural elements and a difference in stabilization energy of 8 kJ/mol, regardless if lysozyme was in solution or adsorbed to silica. PMID- 15271552 TI - Effect of sucrose on the properties of caffeine adsorption layers at the air/solution interface. AB - Sweet and bitter tastes are known to be mediated by G-protein-coupled receptors. The relationship between the chemical structure of gustable molecules and their molecular organization in saliva (aqueous solution) near the surface of the tongue provides a useful tool for elucidating the mechanism of chemoreception. The interactions between stimulus and membrane receptors occur in an anisotropic system. They might be influenced by the molecular packing of gustable molecules within an aqueous solvent (saliva) close to the receptor protein. To investigate the molecular organization of a sweet molecule (sucrose), a bitter molecule (caffeine), and their mixture in an aqueous phase near a "wall", a hydrophobic phase, we modeled this using an air/liquid interface as an anisotropic system. The experimental (tensiometry and ellipsometry) data unambiguously show that caffeine molecules form an adsorption layer, whereas sucrose induces a desorption layer at the air/water interface. The adsorption of caffeine molecules at the air/water interface gradually increases with the volume concentration and is delayed when sucrose is added to the solution. Spectroscopic ellipsometry data show that caffeine in the adsorption layer has optical properties practically identical to those of the molecule in solution. The results are interpreted in terms of molecular association of caffeine with itself at the interface with and without sucrose in the subphase, using the theory of ideal gases. PMID- 15271553 TI - Prediction of the adsorption equilibrium of mixtures composed of supercritical gases. AB - A new model was proposed to predict the adsorption equilibrium of mixtures composed of supercritical gases. The adsorbed phase was visualized as a two dimensional nonideal compressed gas. Pore size distribution was used to describe the energetic heterogeneity of the surface, and the two-dimensional virial equation was used as the local adsorption isotherm. The new model obtained is thermodynamically rigorous because it reduces to Henry's law as pressure approaches zero. The prediction performance of the new model was verified and compared with other models using the experimental data of a ternary mixture of CH4/N2/H2 and two binary mixtures of CH4/C2H4 and CH4/N2. Better performance was shown for all systems tested. PMID- 15271554 TI - Determination of standard isotherms of the sorption of some vapors with cellulose. AB - Standard isotherms of the sorption of water, methanol, and benzene vapors on cellulose using a cellulose standard are determined. The standard, namely, mesoporous cellulose with specific surface of up to 350 m2/g, is obtained by the method of exchanging water in swollen cellulose with organic solvents. A comparison of the experimental sorption isotherm with the standard isotherm makes it possible to determine the specific surface of celluloses accessible a the given sorbate and, in combination with the Brunauer-Emmett-Teller adsorption equation, to characterize their surface properties. The identity of the sorption properties of the initial and dewatered (porous) celluloses relative to active vapors is shown, which evidences the assumed mechanism of swelling as the sorbent's division into morphological structures with the formation of new surface. A comparative analysis of the sorption properties of cellulose and silica, whose nature of active sorption centers is similar (weak acid hydroxyl groups), has been made. The affinity of the standard isotherms and close values of the cross-sectional area of different sorbates on both sorbents testify the similarity in their sorption behavior. Thus, the processes of sorption with rigid and swelling sorbents can be regarded in a unified context. Therefore, the adsorption models developed for rigid sorbents can be applied to cellulose sorbents to analyze their sorption properties. PMID- 15271555 TI - Self-association of water-soluble fluorinated diblock copolymers in solutions. AB - The self-association of the fluorinated diblock copolymer, poly(methacrylic acid) block-poly(perfluorooctylethyl methacrylate) (PMAA-b-PFMA), in water has been investigated by light scattering, potentiometry, atomic force microscopy, and transmission electron microscopy. The size of the polymer micelles increases, as the degree of dissociation of the PMAA blocks increases. Since the charged PMAA block takes the stretched structure, PMAA-b-PFMA can easily form large micelles due to the low steric hindrance of PMAA blocks. Addition of NaCl shielded electrostatic repulsion in the PMAA chain and induced the formation of smaller micelles than water without NaCl did because of the bulky structure of the PMAA chain in the shell of the micelles. The micelle of PMAA-b-PFMA in ethanol is larger than that of poly(t-butyl methacrylate)-block-poly(perfluorooctylethyl methacrylate) (PtBMA-b-PFMA) in ethanol as a result of the higher steric hindrance of the PtBMA block. The dimensions of the core and shell of the micelles were estimated. The micelle of PMAA-b-PFMA in water possesses a rather thick shell and a large volume per molecule, consistent with the extended PMAA chain. On the other hand, the shell of the micelle in an ethanol solution of PtBMA-b-PFMA is thin but has a large surface area. Facts are consistent with the shrunk structure of the PtBMA block in poor solvent. PMID- 15271556 TI - Adsorption of poly(N-isopropylacrylamide) on glass substrata. AB - The adsorption of poly(N-isopropylacrylamide) (PNIPAAM), a well known thermosensitive polymer, on glass was investigated by X-ray photoelectron spectroscopy (XPS) and atomic force microscopy (AFM). The polymer was dissolved in water at low (0.02 g/L) and high (2 g/L) concentration and the tested temperatures were below (25 degrees C) and above (50 degrees C) the lower critical solubility temperature (LCST). Whatever the conditions, a smooth layer of adsorbed molecules spread along the surface was observed. The thickness was about twice higher for high concentration compared to low concentration. The cohesion in the adsorbed layer, as revealed by scraping tests performed by AFM, was higher above the LCST than below the LCST. On top of this adsorbed layer, single-chain coils, globules, or aggregates were present, depending on concentration and temperature. The observation of these additional adsorbed entities was poorly reproducible, presumably due to the lack of shear control upon rinsing. These results emphasize the importance of the characterization of surface morphology to interpret amounts of adsorbed polymers. PMID- 15271557 TI - Direct force measurement of the stability of poly(ethylene glycol) polyethylenimine graft films. AB - The stability and passivity of poly(ethylene glycol)-polyethylenimine (PEG-PEI) graft films are important for their use as antifouling coatings in a variety of biotechnology applications. We have used AFM colloidal-probe force measurements combined with optical reflectometry to characterize the surface properties and stability of PEI and dense PEG-PEI graft films on silica. Initial contact between bare silica probes and PEI-modified surfaces yields force curves that exhibit a long-range electrostatic repulsion and short-range attraction between the surfaces, indicating spontaneous desorption of PEI in the aqueous medium. Further transfer of PEI molecules to the probe occurs with subsequent application of forces between FR = 300 and 500 microN/m. The presence of PEG reduces the adhesive properties of the PEI surface and prevents transfer of PEI molecules to the probe with continuous contact, though an initial desorption of PEI still occurs. Glutaraldehyde crosslinking of the graft films prevents both the initial desorption and subsequent transfer of the PEI, resulting in sustained attractive interaction forces of electrostatic origin between the negatively charged probe and the positively charged copolymer graft films. PMID- 15271558 TI - Comparison of the shape parameters of DNA-cationic lipid complexes and model polyelectrolyte-lipid complexes. AB - In this study, we used a rheological method to study the shape of DNA-cationic lipid complexes and model polyelectrolyte-lipid complexes. We introduced two kinds of anionic polyelectrolytes, sodium polygalacturonate (PGU) and sodium dextran sulfate (DSS), of varying size, as models for DNA. The prepared complexes were incubated under laminar flow conditions. The results show the same quantitative relation between the shape parameter of lipoplexes and the length of anionic polyelectrolytes, including DNA. The rheological behavior of PGU and DSS were similar to that of DNA. PMID- 15271559 TI - Selective separation of arsenopyrite from pyrite by biomodulation in the presence of Acidithiobacillus ferrooxidans. AB - Effective methods for selective separation using flotation or flocculation of arsenopyrite from pyrite by biomodulation using Acidithiobacillus ferrooxidans are presented here. Adhesion of the bacterium to the surface of arsenopyrite was very slow compared to that to pyrite, resulting in a difference in surface modification of the minerals subsequent to interaction with cells. The cells were able to effectively depress pyrite flotation in presence of collectors like potassium isopropyl xanthate and potassium amyl xanthate. On the other hand the flotability of arsenopyrite after conditioning with the cells was not significantly affected. The activation of pyrite by copper sulfate was reduced when the minerals were conditioned together, resulting in better selectivity. Selective separation could also be achieved by flocculation of biomodulated samples. PMID- 15271560 TI - Preparation of thin films comprising palladium nanoparticles by a solid-liquid interface reaction technique. AB - A new approach to the formation of palladium nanoparticulate films with diameter between 6 and 50 nm by the solid-liquid interface reaction technique (SLIRT) has been presented. A solid film of palladium nitrate was formed by the modified spin coating method. This film is subsequently immersed in a reducing solution to initiate a reaction at the interface and ultimately transforms it to a palladium metal film. The kinetics of palladium reduction has been studied by UV-visible spectroscopy. The characterization of the palladium film has been performed by various physicochemical techniques such as XRD, ED, XPS, SEM, EDX, TEM, and UV visible spectroscopy. The texture and morphology of the materials has been investigated by atomic force microscopy (AFM). At a constant palladium nitrate concentration, the average diameter of palladium nanoparticles decreases with an increase of hydrazine concentration. The effect of concentration of hydrazine on the particle size has been discussed. The palladium film formation mechanism has been proposed for the SLIRT. PMID- 15271561 TI - Preparation of Y2O3:Eu3+ nanoparticles in reverse micellar systems and their photoluminescence properties. AB - Y2O3:Eu3+ phosphor nanoparticles (4-8 nm in size) with spherical morphology and narrow size distribution were obtained by calcination of composite Y-Eu hydroxide nanoparticles, which were prepared in sodium bis(2-ethylhexyl)sulfosuccinate (AOT)/isooctane or polyethylene glycol mono-4-nonylphenyl ether (NP 5)/cyclohexane reverse micellar systems. This was achieved by the incorporation of the Y-Eu hydroxide nanoparticles into polyurea (PUA) via in situ polymerization of hexamethylene diisocyanate (HDI) in the reverse micellar solution and subsequent calcination of the resulting PUA materials. The emission intensity of the Y2O3:Eu3+ nanoparticles, prepared in the AOT/isooctane system, was significantly lower than that of the micrometer-size particles prepared in a homogeneous aqueous solution, since the calcined nanoparticles contained Na2SO4 impurity derived from the remaining AOT surfactant. The nanoparticles prepared in the NP-5/cyclohexane system, in contrast, showed higher emission intensity compared to the nanoparticles prepared in the AOT/isooctane system and longer luminescence lifetime compared to the micrometer-size particles prepared in the homogeneous aqueous solution. The photoluminescence intensity of Y2O3:Eu3+, prepared via the proposed process was found to decrease with decreasing the particle size. PMID- 15271562 TI - Surface characterization of plasma sprayed oxide materials: estimation of surface acidity using mass titration. AB - The aim of this study was to characterize the influence of plasma spraying on the point of zero charge (PZC) of Al2O3-, Cr2O3- and TiO2-based materials. PZC is one of the most important parameter, which describes the acidity of oxide material in aqueous environments. PZC values of several plasma sprayed oxides were determined using mass titration method. Studies were performed for initial spray powders and plasma sprayed coating materials. In addition, mass titration experiments were performed for water-washed and nonwashed samples. It was found that mass titration is a suitable method to estimate the surface acidity of relatively coarse sample powders. It was found for most of the studied materials that the limiting pH values (assumed to be close PZC) were close to those reported in literature for the PZC values of traditionally manufactured oxide materials. On the other hand, mass titration curves of some oxide samples showed unexpected deviation in curve shapes and limiting pH. These deviations were probably due to selective dissolution of sample contaminations or sample material. PMID- 15271563 TI - Physical properties of organic particulate UV absorbers used in sunscreens II. UV attenuating efficiency as function of particle size. AB - In this study the UV-attenuating properties of microparticles consisting of a benzotriazole derivative were investigated, which are used as absorbers for UV radiation in cosmetic sunscreens. The particles were micronized in presence of a dispersing agent by means of a ball milling process. According to the energy input different particle sizes were produced in the range of 0.16 to 4 microm. In order to study even smaller particles, the sample with particle size 0.16 microm was fractionated further by centrifugation. Particle sizes were measured using fiberoptic quasi-elastic light scattering (FOQELS) and laser diffractometry. The UV-attenuating properties of the dispersions with different particle sizes were assessed using UV spectroscopy. With decreasing particle size the efficiency of the UV extinction of the dispersion increases up to a particle size of 80 nm. For particles smaller than 80 nm the UV extinction decreases again indicating an optimum at 80 nm. From reflection spectroscopic measurements it was found that scattering makes about 10%, and absorption 90%, of the UV-attenuating effect of the particles, which are obtained at the end of the milling process. PMID- 15271564 TI - Effects of vacuum pyrolysis conditions on the characteristics of activated carbons derived from pistachio-nut shells. AB - Preparation of effective adsorbents from pistachio-nut shells was carried out. Optimization of the vacuum pyrolysis parameters prior to activation was carried out to study the effects of vacuum pyrolysis temperature, hold time, and heating rate on the properties of chars and activated carbons, while CO2 activation conditions were fixed at a temperature of 900 degrees C, an activation time of 30 min, a heating rate of 10 degrees C/min, a CO2 flow rate of 100 cm3/min, and a nitrogen flow rate of 150 cm3/min. The optimum vacuum pyrolysis conditions for preparing activated carbons with high surface area and pore volume were identified. The microstructure and microcrystallinity of the activated carbons prepared were examined by scanning electron microscopy and powder X-ray diffraction techniques respectively while the Fourier transform infrared spectra determined any changes in the surface functional groups produced during different preparation stages. Experimental results show that it is feasible to prepare activated carbons with high BET surface area from pistachio-nut shells. PMID- 15271565 TI - Some questions on the mechanism of changing of silica sorbents structural characteristics under hydrothermal treatment. AB - Obtaining wide porous silicas by means of hydrothermal treatment (HT) of mesoporous silicas is the most reliable method from the point of view of the reproducibility of obtained data. Silica gels with relatively narrow pore distributions by size could be obtained during the first hours of HT. Increase of the duration and rise of the temperature of HT leads to uncontrolled change of the silica gel structure. Existing theories explaining the mechanism of the change of porous characteristics during hydrothermal treatment do not completely describe the reasons for one or another change. A new approach explaining the mechanism of the change in porous characteristics of the silica during HT, based on the assumption that the water in the pores is in "motion state," has been suggested in the present work. Theories did not consider water state in the pores during HT. An attempt to explain the changes of the porous structure of the silica gels during HT is undertaken in the present manuscript. PMID- 15271566 TI - Dynamic surface tension measurements with submillisecond resolution using a capillary-jet instability technique. AB - An oscillating capillary jet method is implemented to measure surface tension of aqueous nonionic surfactant solutions as a function of surface age from the jet orifice. The experimental technique captures the evolution of jet swells and necks continuously along the jet propagation axis and is used in conjunction with an existing linear, axisymmetric, constant-property model to determine surface tension of liquids. The method is first validated using deionized water and isopropyl alcohol (constant surface tension test fluids) and a procedure is described to identify the optimum wavelength from the breakup point, which produces the smallest error in surface tension measurements. Dynamic surface tension data of concentrated aqueous Tergitol NP-8 surfactant solutions is then presented. The measurements are performed over a spatial length of approximately 1.5 wavelengths, a span corresponding to 0.6-4.2 ms time window from the jet orifice. Submillisecond surface age measurements are made possible by decreasing the jet diameter. Increased surfactant concentrations make the liquid jet more stable and allow measurements at higher surface ages. The correlation of Hua and Rosen fits well the dynamic surface tension data, which includes submillisecond surface ages. Finally, the time required for surface tension to reach equilibrium levels is estimated using a simple adsorption kinetics theory of surfactant molecules on the liquid/air interface. PMID- 15271567 TI - Electrospray emission from nonwetting flat dielectric surfaces. AB - Electrosprays are devices in which nanometer sized droplets and/or solvated ions are electrically extracted from a liquid surface and accelerated to high velocities. They are usually constructed from conductive capillaries with one of the ends tapered down to a sharp tip with the purpose of enhancing the local electric field that produces the instability that develops into the Taylor cone structure from which emission occurs. In an alternative configuration, the conductive needles are replaced by small holes through a dielectric, nonwetting flat block. An electrostatic model shows that if the emitter material has low dielectric constant then the local electric field near the emission site is enhanced in a very similar way as with sharp metallic needles. Furthermore, the combination of the nonwetting property of the material and the sharp corner formed in the hole-surface interface effectively anchors the Taylor cone to the edge of the hole, thus simplifying the process of manufacturing. The possible microfabrication of this configuration makes it especially attractive for producing arrays of large numbers of individual emitters. Such arrays may find use as space propulsion thrusters and in the analytical industry to improve the characteristics of mass-spectrometric analyses. PMID- 15271568 TI - Effects of poly(ethylene oxide)-poly(propylene oxide)-poly(ethylene oxide) triblock copolymers on structure and stability of liposomal dioleoylphosphatidylethanolamine. AB - The effects caused by poly(ethylene oxide)-poly(propylene oxide)-poly(ethylene oxide) (PEO-PPO-PEO; Pluronic) copolymers on the structure and stability of dioleoylphosphatidylethanolamine (DOPE) liposomes were studied by means of turbidity, leakage, and cryo-transmission electron microscopy investigations. The results show that by inclusion of Pluronics in the DOPE dispersion it is possible to stabilize the lamellar Lalpha phase and to produce liposomes that are stable and nonleaky at low pH (pH 5). The stabilizing capacity was observed to depend critically on the molecular composition of the Pluronics. Block copolymers with comparably long PPO and PEO segment lengths, such as F127 and F108, most effectively protected DOPE liposomes prepared at high pH from aggregation and subsequent structural rearrangements induced by acidification. A sufficiently long PPO block was found to be the most decisive parameter in order to obtain adequate coverage of the liposome surface at low Pluronic concentrations. Upon increasing the copolymer concentration, however, Pluronics with comparably short PPO and PEO segment lengths, such as F87 and P85, could also be used to stabilize the DOPE liposomes. Essentially the same trends were observed when the Pluronics were added to preformed DOPE liposomes instead of being included in the preparation mixture. In this case the least effective copolymers failed, however, to completely prevent the DOPE liposomes from releasing encapsulated hydrophilic markers. PMID- 15271569 TI - Static and dynamic light-scattering studies on micellar solutions of alkyldimethylbenzylammonium chlorides. AB - Static (SLS) and dynamic (DLS) light-scattering techniques were applied to the study of the aggregation of dodecyl- (C12DBACl), tetradecyl- (C14DBACl), and hexadecyldimethylbenzylammonium (C16DBACl) chlorides in water and in 0.01 and 0.05 m NaCl aqueous solutions at 25 degrees C. Results of SLS measurements yielded critical micelle concentration (cmc) values for aqueous and NaCl solutions. The aggregation numbers of the micelles for the homologous surfactants are low but increase with chain length and ionic strength of the solution. Various patterns of changes of the diffusion coefficient, D, as a function of chain length, molality, and with ionic strength were found for the studied surfactants. Transformations in the structure of micelles of C14DBACl in 0.01 m NaCl occur at a concentration of surfactant of about 0.01 m. Such transformations, presumably due to rodlike structure, are the more extensive the higher the concentration of NaCl. The concentration of C16DBACl in 0.05 m NaCl covers the range where already repulsive interactions between micelles occur, as judged by the strongly negative slope of the D versus molality plot. To provide additional information on the suggested transformations, complementary viscosity measurements for C14DBACl in 0.01 m of NaCl have been performed. PMID- 15271570 TI - Effect of structure of PEO-PPO-PEO copolymers on reverse micelle formation induced by compressed CO2. AB - The micellization of PEO-PPO-PEO block copolymers in p-xylene has been studied in the presence of CO2. With the application of CO2, some copolymers with suitable molecular weights and EO ratios can form reverse micelles with critical micellization pressure up to 5.8 MPa. For the copolymers with the same length of PO block, higher EO ratios facilitate reverse micelle formation. For the copolymers with the same composition, higher molecular weight is favorable to form reverse micelles. With the suitable composition and molecular weight, the critical micelle pressure (CMP) of copolymers decreases with the increase in the lengths of PEO and PPO blocks due to the hydrophilic and folding effects, respectively. Both the EO ratios and the molecular weights are important for the formation of reverse micelle. The reverse micelle solution can solubilize water with W0 (molar ratio of water to EO segment) up to 3.3. PMID- 15271571 TI - Foam drainage on the microscale I. Modeling flow through single Plateau borders. AB - The drainage of liquid through a foam involves flow in channels, also called Plateau borders, which generally are long and slender. We model this flow by assuming the flow is unidirectional, the shear is transverse to the flow direction, and the liquid/gas interfaces are mobile and characterized by a Newtonian surface viscosity, which does not depend on the shear rate. Numerical finite difference simulations are performed, and analytical approximations for the velocity fields inside the channels and the films that separate the bubbles are given. We compare the liquid flow rates through interior channels, exterior channels (i.e., channels contacting container walls) and films. We find that when the number of exterior channels is comparable to the number of interior channels, i.e., narrow container geometries, the exterior channels can significantly affect the dynamics of the drainage process. Even for highly mobile interfaces, the films do not significantly contribute to the drainage process, unless the amount of liquid in the films is within a factor of ten of the amount of liquid in the channels. PMID- 15271572 TI - Foam drainage on the microscale II. Imaging flow through single Plateau borders. AB - The liquid in foam forms an interconnected network, which is composed of Plateau borders, nodes, and films. One of the dominant pathways for foam drainage is flow through Plateau borders, and we use confocal microscopy to obtain experimental results for the flow fields inside individual Plateau borders. For three types of surfactants detailed comparisons are made with a model based upon the influence of surface viscosity at free boundaries between the gas in the bubbles and the liquid in the Plateau borders. The model describes the flows well, and we find good agreement between the surface viscosity predicted by this model and representative values found in the literature. We also give a qualitative description of the flow in the nodes. PMID- 15271573 TI - Physicochemical properties of anionic triple-chain surfactants in alkaline solutions. AB - Physicochemical properties of two types of anionic triple-chain surfactants of disodium 2,10-didecyl-6-dodecyl-3,6,9-triazaundecanedioate (3C12dtda) with three hydrocarbon chains and two carboxylate headgroups, and trisodium tris(N-2 ethylamino-N'-2-dodecylcarboxylato)azane (3C12tata) with three hydrocarbon chains and three carboxylate headgroups were characterized by several techniques such as static surface tension, fluorescence spectroscopy, and dynamic and static light scattering in alkaline solutions at pH 13. 3C12dtda shows a much lower critical micelle concentration (cmc) and higher efficiency in lowering the surface tension than 3C12tata. 3C12dtda also provides a very small occupied area per molecule as compared to 3C12tata and packs closely at the interface. The fluorescent intensity ratio of the first to third band in the emission spectra of pyrene for two triple-chain surfactants starts to decrease gradually from around the cmc obtained by the surface tension method, and decreases sharply from the concentration of 10-18 fold cmc. It is also found that the polarity of the triple chain surfactant micelle is very much lower than that of the single-chain surfactant micelle. The results of dynamic and static light-scattering measurements show the difference behavior in the aggregation process for two triple-chain surfactants; that is, 3C12dtda forms large aggregates, and 3C12tata forms two aggregates of normal micelle size and a larger one in solution. This indicates that the aggregation states of triple-chain surfactants differ by the number of carboxylate groups, and the large aggregates are derived from the strong attractive interaction between multiple hydrocarbon chains. PMID- 15271574 TI - Surfactants treatment of crude oil contaminated soils. AB - This study reports experimental measurements investigating the ability of a biological (rhamnolipid) and a synthetic (sodium dodecyl sulfate, SDS) surfactant to remove the North Sea Ekofisk crude oil from various soils with different particle size fractions under varying washing conditions. The washing parameters and ranges tested were as follows: temperature (5 to 50 degrees C), time (5 to 20 min), shaking speed (80 to 200 strokes/min), volume (5 to 20 cm3), and surfactant concentration (0.004 to 5 mass%). The contaminated soils were prepared in the laboratory by mixing crude oil and soils using a rotating cylindrical mixer. Two contamination cases were considered: (1) weathered contamination was simulated by keeping freshly contaminated soils in a fan assisted oven at 50 degrees C for 14 days, mimicking the weathering effect in a natural hot environment, and (2) nonweathered contamination which was not subjected to the oven treatment. The surfactants were found to have considerable potential in removing crude oil from different contaminated soils and the results were comparable with those reported in literature for petroleum hydrocarbons. The removal of crude oil with either rhamnolipid or SDS was within the repeatability range of +/-6%. The most influential parameters on oil removal were surfactant concentration and washing temperature. The soil cation exchange capacity and pH also influenced the removal of crude oil from the individual soils. However, due to the binding of crude oil to soil during weathering, low crude oil removal was achieved with the weathered contaminated soil samples. PMID- 15271575 TI - Experimental investigation of capillary pressure influence on breaking of emulsions stabilized by solid particles. AB - Experimental investigations of the stability of emulsions stabilized by modified silica particles were carried out under high capillary pressure in the emulsion films. Emulsions were placed on a ceramic porous plate or in a centrifugal field. The capillary pressure was created by decreasing the pressure in the continuous phase of the emulsion. The dependence of the emulsion's lifetime on the capillary pressure magnitude and on the contact-angle value is discussed. PMID- 15271576 TI - Influence of ionic specificity on the microstructure and the strength of gelled colloidal silica suspensions. AB - The dynamic rheological behavior of gelled Ludox suspensions in the presence of the structure-breaker ammonium counterion and the structure-maker sodium counterion has been investigated. Depending on the nature of the electrolyte and its concentration, the results highlight the crucial effect of the network microstructure on the rheological results. Within the same microstructure, the strength of gelled networks has been found to be greater in the presence of the poorly hydrated ammonium counterions at any Ludox volume fraction. The obtained results were discussed with respect to recent contradictory trends reported in the literature concerning the behavior of silica suspensions. PMID- 15271577 TI - Nonlinear electrokinetics and "superfast" electrophoresis. AB - Nonlinear and nonequilibrium electrophoresis of spherical particles of radius a is shown to be possible when the solid surface allows field or current penetration. At low particle Peclet numbers, transient capacitative charging occurs until the surface polarization completely screens the external field. For a DC applied field [see text], the resulting electrokinetic velocity reaches Dukhin's maximum value of [formula: see text], where [see text] and mu are the liquid permittivity and viscosity. At high Peclet numbers, electroosmotic convection of the electroneutral bulk stops the transient charging before complete field-line exclusion. For an ion-selective and conducting spherical granule, the polarization is then determined by the steady-state Ohmic current driven by the penetrated external field. The high-Peclet electrokinetic velocity is lower, diffusivity-dependent and scales as [see text]. PMID- 15271578 TI - Use of a chiral surfactant for enantioselective reduction of a ketone. AB - The influence of a chiral surfactant and a polymer-supported chiral additive on reduction of ketones using sodium borohydride will be described. Initial preparations involved methylation of (S)-leucinol to give (2S)-N , N-dimethyl-2 amino-4-methyl-1-pentanol (1) (67%). The chiral surfactant (2) was synthesized by reacting (1) with bromohexadecane (71%). The functionalized styrene for the polymer-supported chiral additive (5) was synthesized by reacting (1) with 4 vinylbenzyl chloride. Polymerization was carried out with 10% of the functionalized monomer (4), 5% cross-linking agent divinylbenzene, and 85% styrene with AIBN as the initiator. The activity of the chiral surfactant and polymeric additive were examined by using them as additives in a standard reduction of 2-pentanone with sodium borohydride to yield (R)- and (S)-2-pentanol (3) (20%). The resulting alcohol was analyzed by polarimetry (ee 9.5%) and also esterified with (2S)-methylbutyric acid prior to characterization by NMR. 13C NMR indicated an enantiomeric excess of 5.2% when the chiral surfactant was used, and 7% when the polymeric additive was used. PMID- 15271579 TI - An ellipsometry-based Alzheimer plaque mimic: Effect of beta-amyloid, lipoprotein identity and apolipoprotein E isoform. AB - Ca2+-induced deposition of low-density lipoprotein (LDL), intermediate-density lipoprotein (IDL), and high-density lipoprotein (HDL) at proteoheparan sulfate modified surfaces was investigated as a function of beta-amyloid (Abeta) presence and apolipoprotein E isoform. Presence of beta-amyloid resulted in an increased deposition, as did the E4/E4 isoform compared to the corresponding E3/E3 isoform. The results are compatible with findings reported in literature on plaque formation in Alzheimer's disease, and suggest that, although simplistic, the present model system may have some potential in biosensor studies of Alzheimer plaque formation. PMID- 15271580 TI - Is impaired neurogenesis relevant to the affective symptoms of depression? PMID- 15271581 TI - Depression: a case of neuronal life and death? AB - Preclinical and clinical studies have demonstrated that stress or depression can lead to atrophy and cell loss in limbic brain structures that are critically involved in depression, including the hippocampus. Studies in experimental animals demonstrate that decreased birth of new neurons in adult hippocampus could contribute to this atrophy. In contrast, antidepressant treatment increases neurogenesis in the hippocampus of adult animals and blocks the effects of stress. Moreover, blockade of hippocampal neurogenesis blocks the actions of antidepressants in behavioral models of depression, demonstrating a direct link between behavior and new cell birth. This perspective reviews the literature in support of the hypothesis that altered birth of new neurons in the adult brain contributes to the etiology and treatment of depression and considers research strategies to test this hypothesis. PMID- 15271582 TI - Neurogenesis and depression: etiology or epiphenomenon? AB - The concept that decreased neurogenesis might be the cause of depression is supported by the effects of stress on neurogenesis and the demonstration that neurogenesis seems to be necessary for antidepressant action. Data from the animal models tested to date show that decreasing the rate of neurogenesis does not lead to depressive behavior. Furthermore, evidence shows that an effective treatment for depression, transcranial magnetic stimulation, does not alter rates of neurogenesis. On the basis of these findings, it is suggested that neurogenesis might play a subtle role in depression but that it is not the primary factor in the final common pathway leading to depression. PMID- 15271583 TI - Effects of cyclic adenosine monophosphate response element binding protein overexpression in the basolateral amygdala on behavioral models of depression and anxiety. AB - BACKGROUND: Chronic antidepressant administration increases the cyclic adenosine monophosphate response element binding protein (CREB) in the amygdala, a critical neural substrate involved in the physiologic responses to stress, fear, and anxiety. METHODS: To determine the role of CREB in the amygdala in animal models of depression and anxiety, a viral gene transfer approach was used to selectively express CREB in this region of the rat brain. RESULTS: In the learned helplessness model of depression, induction of CREB in the basolateral amygdala after training decreased the number of escape failures, an antidepressant response. However, expression of CREB before training increased escape failures, and increased immobility in the forced swim test, depressive effects. Expression of CREB in the basolateral amygdala also increased behavioral measures of anxiety in both the open field test and the elevated plus maze, and enhanced cued fear conditioning. CONCLUSIONS: Taken together, these data demonstrate that CREB expression in the basolateral amygdala influences behavior in models of depression, anxiety, and fear. Moreover, in the basolateral amygdala, the temporal expression of CREB in relation to learned helplessness training, determines the qualitative outcome in this animal model of depression. PMID- 15271584 TI - Neonatal quinpirole treatment impairs Morris water task performance in early postweanling rats: relationship to increases in corticosterone and decreases in neurotrophic factors. AB - BACKGROUND: Past studies from this laboratory have shown that quinpirole administration from postnatal day (P) 1-21 produces persistent supersensitization of the dopamine D2 receptor that persists throughout the animal's lifetime. METHODS: In Experiment 1, both male and female rats were treated with quinpirole or saline from P1-21 and tested on the place and match-to-place versions of the Morris water task (MWT) from P22-28. In Experiment 2, both male and female rats were administered either acute or chronic injections of quinpirole (1 mg/kg) or saline beginning on P1 until analysis for corticosterone (CORT) on P7, 14, or 21. RESULTS: Neonatal quinpirole treatment produced deficits on both versions of the MWT compared with saline control. One day after behavioral testing, brain tissue was harvested, and the hippocampus was analyzed for nerve growth factor (NGF) and brain-derived nerve growth factor (BDNF); NGF was found to be significantly decreased by neonatal quinpirole treatment. Acute or chronic quinpirole treatment on P14 produced a larger increase in CORT than controls and produced larger increases in CORT than control rats on P21. CONCLUSIONS: These results demonstrate that neonatal quinpirole treatment produces cognitive deficits that could be related to decreases in hippocampal NGF and increases in CORT, resulting in abnormalities in hippocampal development. PMID- 15271585 TI - Is the G72/G30 locus associated with schizophrenia? single nucleotide polymorphisms, haplotypes, and gene expression analysis. AB - BACKGROUND: The genes G72/G30 were recently implicated in schizophrenia in both Canadian and Russian populations. We hypothesized that 1) polymorphic changes in this gene region might be associated with schizophrenia in the Ashkenazi Jewish population and that 2) changes in G72/G30 gene expression might be expected in schizophrenic patients compared with control subjects. METHODS: Eleven single nucleotide polymorphisms (SNPs) encompassing the G72/G30 genes were typed in the genomic deoxyribonucleic acid (DNA) from 60 schizophrenic patients and 130 matched control subjects of Ashkenazi ethnic origin. Case-control comparisons were based on linkage disequilibrium (LD) and haplotype frequency estimations. Gene expression analysis of G72 and G30 was performed on 88 postmortem dorsolateral prefrontal cortex samples. RESULTS: Linkage disequilibrium analysis revealed two main SNP blocks. Haplotype analysis on block II, containing three SNPs external to the genes, demonstrated an association with schizophrenia. Gene expression analysis exhibited correlations between expression levels of the G72 and G30 genes, as well as a tendency toward overexpression of the G72 gene in schizophrenic brain samples of 44 schizophrenic patients compared with 44 control subjects. CONCLUSIONS: It is likely that the G72/G30 region is involved in susceptibility to schizophrenia in the Ashkenazi population. The elevation in expression of the G72 gene coincides with the glutamatergic theory of schizophrenia. PMID- 15271586 TI - Positive association between synapsin II and schizophrenia. AB - BACKGROUND: Synapsin II encodes a neuron-specific phosphoprotein that selectively binds to small synaptic vesicles in the presynaptic nerve terminal. The expressions of messenger ribonucleic acid and protein of synapsin II have been reported to be significantly reduced in the brains of schizophrenia patients. The synapsin II gene is located on 3p25, a region that has been implicated to be associated with schizophrenia by genetic linkage. All these findings suggest synapsin II as a candidate gene for schizophrenia. METHODS: In this work, we studied four markers (two single nucleotide polymorphisms (SNPs): rs308963 and rs795009; and two insertion/deletion polymorphisms: rs2307981 and rs2308169) covering 144.2 kilobase pairs (kb) with an average interval of 38 kb in synapsin II in a sample of 654 schizophrenic patients and 628 normal control subjects to explore the mechanism underlying schizophrenia. RESULTS: We found significant differences in allele frequency distribution of SNP rs795009 (p =.000018, odds ratio 1.405, 95% confidence interval 1.202-1.641) between patients and control subjects. The T allele was significantly higher in patients than in control subjects. Moreover, the overall frequency of haplotype showed significant differences between patients and control subjects (p <.000001). CONCLUSIONS: This study suggests a positive association between synapsin II and schizophrenia, implying that synapsin II is involved in the etiology of schizophrenia. PMID- 15271587 TI - Abnormal cortical folding in high-risk individuals: a predictor of the development of schizophrenia? AB - BACKGROUND: A number of studies have found localized differences in the appearance and extent of cortical folding between the brains of schizophrenic patients and healthy control subjects. This study aimed to determine whether, within individuals at genetic high risk for schizophrenia, there are pre-existing differences in gyral folding between those who subsequently develop the disease and those who remain unaffected. METHODS: Assessment was conducted on baseline magnetic resonance imaging scans of 30 young adults grouped into 14 who remained unaffected and 16 who subsequently developed schizophrenia. The gyrification index (GI), the ratio of the inner and outer cortical surface contours, was measured bilaterally on every second 1.88-mm image slice in four specifically defined lobar regions. Independent t tests and volume and genetic liability correlations were conducted for each region, followed by a post hoc examination. RESULTS: Right prefrontal lobe GI values were significantly increased in individuals who subsequently developed schizophrenia. Post hoc examination suggested that the areas of greatest increase lay anteriorly and laterally in Brodmann areas 9 and 10. Correlations with volume and analysis of covariance suggested some overlap between GI and volume measures. CONCLUSIONS: Differences in frontal lobe GI might reflect disturbed or abnormal connectivity predictive of subsequent schizophrenia. PMID- 15271588 TI - Functional lateralization of the sensorimotor cortex in patients with schizophrenia: effects of treatment with olanzapine. AB - BACKGROUND: Earlier cross-sectional studies with functional magnetic resonance imaging (fMRI) in treated patients with schizophrenia have reported abnormalities of cortical motor processing, including reduced lateralization of primary sensory motor cortex. The objective of the present longitudinal study was to evaluate whether such cortical abnormalities represent state or trait phenomena of the disorder. METHODS: Seventeen acutely ill, previously untreated patients were studied after 4 weeks and after 8 weeks of olanzapine therapy. Seventeen matched healthy subjects served as control subjects. All subjects underwent two fMRI scans 4 weeks apart during a visually paced motor task using a simple periodic block design. Functional magnetic resonance imaging data were analyzed in Statistical Parametric Mapping (SPM99). Region of interest analyses were used to determine a laterality quotient (an index of lateralization) of motor cortical regions. RESULTS: The fMRI data indicated that patients had reduced activation of the primary sensory motor cortex at 4 weeks but not at 8 weeks; however, the laterality quotient in the primary sensory motor cortex was reduced in patients at both time points. CONCLUSIONS: These results suggest that some cortical abnormalities during motor processing represent state phenomena, whereas reduced functional lateralization of the primary sensory motor cortex represents an enduring trait of schizophrenia. PMID- 15271589 TI - Twenty-four-hour cortisol secretion patterns in prepubertal children with anxiety or depressive disorders. AB - BACKGROUND: Previous studies found few abnormalities in hypothalamic-pituitary adrenal (HPA) axis function in prepubertal children with anxiety or depressive disorders. In this study, we combined data from two independent, consecutive studies to achieve a larger sample size. Our goal was to identify potential alterations in the circadian pattern of cortisol secretion in anxious or depressed children. METHODS: A total of 124 prepubertal subjects from two independent samples (76 with major depressive disorder, 31 with anxiety disorders, and 17 healthy control subjects) were studied. Blood samples collected for cortisol at hourly intervals over a 24-hour period were examined. Analyses were performed aligning cortisol samples by clock-time. Additional analyses aligning samples by sleep-onset time were performed with a subsample of subjects. RESULTS: In the combined sample, significant findings emerged that were previously undetected. Anxious children exhibited significantly lower nighttime cortisol levels and an initially sluggish rise in cortisol during the nighttime when compared with depressed and healthy control children. In contrast, depressed children did not show a clear-cut pattern of differences compared with healthy control children. CONCLUSIONS: Anxious children seem to exhibit an altered pattern of nighttime cortisol secretion, with an initially sluggish or delayed nocturnal rise before reaching similar peak levels of cortisol near the time of awakening. These findings suggest subtle alterations in HPA axis function in prepubertal children with anxiety disorders. PMID- 15271590 TI - Saliva cortisol in posttraumatic stress disorder: a community epidemiologic study. AB - BACKGROUND: Stress activates the hypothalamic-pituitary-adrenal (HPA) axis, so it was expected that posttraumatic stress disorder (PTSD) would be associated with activation of this axis; however, studies have found both increased and decreased cortisol in PTSD. To address this question, we collected saliva cortisol at home in a subsample of a longitudinal epidemiologic sample. METHODS: Six hundred eighty-four persons randomly selected from the total sample of 913 were requested to collect saliva samples upon awakening and in the early evening. Of these, 538 responded with samples, 516 of whom met inclusion criteria. These were 68 exposed to trauma with lifetime PTSD, 265 exposed to trauma with no PTSD, and 183 never exposed to trauma. RESULTS: In a comparison of these three groups, lifetime PTSD revealed elevated evening saliva cortisol compared with exposed/no PTSD. When lifetime comorbidity with major depressive disorder (MDD) was included in the analysis, only persons with comorbid PTSD and MDD showed this evening elevation in cortisol. Persons with PTSD alone (never MDD) showed normal saliva cortisol levels, as did subjects with lifetime MDD alone. CONCLUSIONS: Neither exposure to trauma nor PTSD alone is associated with alterations in saliva cortisol; however, elevated cortisol is found in PTSD comorbid with lifetime MDD. PMID- 15271591 TI - From Freud to Gehrig to Rapaport to DiMaggio. AB - Lacking the size, talent, and speed to play shortstop for the New York Yankees, I chose instead to become a psychologist/psychoanalyst. Yet, baseball and psychoanalysis share much in common. Each is played in the inner diamonds of one's mind, values the past and what is passing, and requires a strong work ethic. Both are timeless and involve an almost boring leisureliness between occasional moments of crisis. Then too, each calls for an attitude of consistency and patience and a responsibility to do one's best. From my parents and brother to my wife and son and his family, from my teachers and supervisors to my colleagues, and from Menninger to the Society for Personality Assessment (SPA), I feel deeply grateful for the partners and teammates I have and have had. PMID- 15271592 TI - Psychometric deviance measured by MMPI in adoptees at high risk for schizophrenia and their adoptive controls. AB - Psychometric deviance in personality traits as assessed by the Minnesota Multiphasic Personality Inventory (MMPI; Dahlstrom, Welsh, & Dahlstrom, 1982) was compared between adopted-away, high-risk (HR) offspring of schizophrenic biologic mothers and low-risk (LR) controls. A subsample of the Finnish Adoptive Family Study (Tienari et al., 2000) included 60 HR adoptees and 76 LR control adoptees who were tested by the MMPI before the onset of any psychiatric disorder at the mean age of 24 years. The HR group was found to be distinguishable based on deviant scores on the scales HOS and HYP, indicating emotional unresponsiveness, restricted affectivity, and decreased energy. These may also be considered possible premorbid and prodromal signs of future schizophrenia among the HR adoptees. PMID- 15271593 TI - The Depressive Personality Disorder Inventory and its relationship to quality of life, hopefulness, and optimism. AB - The construct validity of the Depressive Personality Disorder Inventory (DPDI; Huprich, Margrett, Barthelemy, & Fine, 1996) was examined through its relationship to the constructs of hope, optimism, and quality of life (QOL). Three hundred thirty-two undergraduate students were administered the DPDI and measures of the aforementioned constructs. As predicted, the DPDI negatively correlated with all measures. Individuals classified with a depressive personality disorder had significantly higher scores on measures of hope, optimism, and QOL compared to a control group. Stepwise regression analysis indicated that optimism, QOL, and one component of hope significantly predicted DPDI scores, although more variance was accounted for in women than men. These findings are explained in light of Carver and Scheier's (2000) explanation of optimism and its relationship to hope. In sum, it appears that the construct validity of the DPDI is supported within an undergraduate sample. PMID- 15271594 TI - Psychological entitlement: interpersonal consequences and validation of a self report measure. AB - Nine studies were conducted with the goal of developing a self-report measure of psychological entitlement and assessing its interpersonal consequences. The Psychological Entitlement Scale (PES) was found to be reliable and valid (Study 1, 2), not associated with social desirability (Study 2), stable across time (Study 3), and correlated negatively with two of the Big Five factors: agreeableness and emotional stability (Study 4). The validity of the PES was confirmed in studies that assessed willingness to take candy designated for children (Study 5) and reported deservingness of pay in a hypothetical employment setting (Study 6). Finally, the PES was linked to important interpersonal consequences including competitive choices in a commons dilemma (Study 7), selfish approaches to romantic relationships (Study 8), and aggression following ego threat (Study 9). Psychological entitlement has a pervasive and largely unconstructive impact on social behavior. PMID- 15271595 TI - Association of Rorschach and MMPI psychosis indicators and schizophrenia spectrum diagnoses in a Russian clinical sample. AB - In this study, I investigated the relationships among psychological test variables and schizophrenia spectrum diagnoses in a Russian sample of 180 psychiatric patients. Schizophrenia is understood somewhat differently in Russia than in the West. Analyses compared Rorschach (SCZI, PTI; Exner, 2001) and MMPI (Berezin, Mitroshinkov, & Sokolova, 1994) psychosis indicators (Sc, Sc3, Sc6, and BIZ) and 3 diagnostic systems: (a) Russian traditional, (b) the Russian-modified International Classification of Diseases (9th ed. [ICD-9]; Ministerstvo Zdravokhraneniya SSSR, 1982), and (c) the nonmodified ICD-10 (World Health Organization, 1992; comparable to the Diagnostic and Statistical Manual of Mental Disorders [4th ed.], American Psychiatric Association, 1994). Results showed modest support for the SCZI and PTI but not the MMPI indicators. While the field awaits further evidence, psychologists should proceed with caution when using the Rorschach and MMPI to assess for psychosis among Russians. PMID- 15271596 TI - The Millon Adolescent Clinical Inventory: is it valid and reliable for Mexican American youth? AB - In this study, we examined the internal consistency reliability, construct related validity, and mean base rate scores of the Millon Adolescent Clinical Inventory (MACI; Millon, Millon, & Davis, 1993) Personality and Clinical scales in a sample (N = 131) of juvenile offending, educationally at-risk, and substance dependent Mexican American youth. Overall, the MACI scales demonstrated good reliability estimates. Using methods derived from contrast analysis (Westen & Rosenthal, 2003), the scales performed as would be expected based on predictions derived from Millon's (1969, 1981) theory. In addition, the scales differentiated in an expected fashion among the 3 groups. The results of this study provide preliminary support for the use of the MACI among Mexican Americans. PMID- 15271597 TI - NEO-PI-R neuroticism scores in substance-dependent outpatients: internal consistency and self-partner agreement. AB - We evaluated internal consistency reliabilities and self-partner agreement on Revised NEO Personality Inventory (Costa & McCrae, 1992) Neuroticism (N) domain and facet scores (anxiety, angry hostility, depression, self-consciousness, impulsiveness, vulnerability) in a sample of 48 substance-dependent outpatients. Low internal consistency was seen for self-rated impulsiveness (alpha =.36). Agreement between self and partner reports was low for impulsiveness (.22) and vulnerability (.24) and was modest for the remaining facets of N (.31 to.34) and the N domain score (.31). PMID- 15271598 TI - Review of computer-based test interpretation software for the MMPI-2. PMID- 15271601 TI - Introduction to the special section on protective factors in the relation between community violence exposure and adjustment in youth. PMID- 15271602 TI - Exposure to community violence and violence perpetration: the protective effects of family functioning. AB - Although research has found that urban youth are exposed to excessive levels of community violence, few studies have focused on the factors that alter the risk of exposure to violence or the processes through which youth who are exposed to community violence do better or worse. This study investigates the risk of exposure to community violence and its relation to violence perpetration among a sample of 263 African American and Latino male youth living in inner-city neighborhoods. The study also examines the role that family functioning plays in moderating the risk. The study finds that youth from struggling families--those that consistently used poor parenting practices and had low levels of emotional cohesion--were more likely to be exposed to community violence. It also finds a relation between exposure to violence and later violence perpetration. However, youth exposed to high levels of community violence but living in families that functioned well across multiple dimensions of parenting and family relationship characteristics perpetrated less violence than similarly exposed youth from less well-functioning families. PMID- 15271603 TI - Social support factors as moderators of community violence exposure among inner city African American young adolescents. AB - Using both surveys and the experience sampling method (ESM), community violence exposure, social support factors, and depressive and anxiety symptoms were assessed longitudinally among inner-city African American adolescents. Moderator models were tested to determine protective factors for youth exposed to community violence. Several social support factors emerged as protective-stabilizing forces for witnesses of violence both cross-sectionally and longitudinally, including maternal closeness, time spent with family, social support, and daily support (ESM). Contrary to hypotheses, several social support factors demonstrated a promotive-reactive effect such that, in conditions of high victimization, they failed to protect youth from developing symptoms. Effects did not differ by outcome or sex, though sex differences in findings emerged. Protective stabilizing effects occurred more for witnessing violence, whereas promotive reactive patterns occurred more for victimization. Results affirm social support factors as protective from the adverse effects of violence exposure, but they also suggest that some factors typically conceived as contributing to resilience might at times fail to protect youth in conditions of extreme risk. PMID- 15271604 TI - Urban adolescents' exposure to community violence: the role of support, school safety, and social constraints in a school-based sample of boys and girls. AB - This study examined recent exposure to violence in the community and in other settings, protective factors, and current psychological functioning among 349 young adolescents from 9 urban middle schools. The majority (76%) of adolescents reported witnessing or being victimized by at least 1 violent event in the prior 6 months. Nearly half of adolescents who had talked about their experience of a violent event reported feeling constrained from sharing their thoughts or feelings because of others' reactions. After controlling for daily hassles, more exposure to violence was associated with more self-reported posttraumatic stress disorder (PTSD) and depressive symptoms. Exposure to violence was not a significant predictor of teachers' ratings of adaptive functioning or internalizing symptoms. Support from specific individuals, perceived school safety, and lower constraints for discussing violence showed protective effects in the relation between exposure to violence and specific dimensions of psychological functioning. The implications of this research for school-based interventions are discussed. PMID- 15271605 TI - Violence exposure and adjustment in inner-city youth: child and caregiver emotion regulation skill, caregiver-child relationship quality, and neighborhood cohesion as protective factor. AB - This short-term, longitudinal interview study used an ecological framework to explore protective factors within the child, the caregiver, the caregiver-child relationship, and the community that might moderate relations between community violence exposure and subsequent internalizing and externalizing adjustment problems and the different patterns of protection they might confer. Participants included 101 pairs of African American female caregivers and one of their children (56% male, M = 11.15 yrs, SD = 1.28) living in high-violence areas of a mid-sized southeastern city. Child emotion regulation skill, felt acceptance from caregiver, observed quality of caregiver-child interaction, and caregiver regulation of emotion each were protective, but the pattern of protection differed across level of the child's ecology and form of adjustment. Implications for prevention are discussed. PMID- 15271606 TI - Relation between witnessing violence and drug use initiation among rural adolescents: parental monitoring and family support as protective factors. AB - This study examined the relation between witnessing violence and drug use initiation among 6th graders attending middle schools in 5 rural counties and investigated the extent to which family support and parental monitoring moderated this relation. Data were obtained from 1,282 adolescents at 2 time points during the 6th grade. Witnessing violence predicted subsequent initiation of cigarette, beer and wine, liquor, and advanced alcohol use. Adolescents who reported high levels of family support and parental monitoring were less likely to initiate use across all drug categories except beer and wine. High levels of parental monitoring and family support were effective in buffering the relation between witnessing violence and initiation of cigarette and advanced alcohol use at low levels of witnessing violence. With increasing levels of witnessing violence, however, the protective effects of monitoring and support were substantially diminished. These findings have important implications for research and intervention efforts. PMID- 15271607 TI - Children's exposure to community violence: implications for understanding risk and resilience. AB - The 5 articles included in this special section are reviewed in this article. The studies encompassed were all focused on pre- or early adolescents, and samples were generally from inner-city areas, with 1 involving rural youth. Considered collectively, the results point to 3 major conclusions: Many children in America are regularly exposed to violence in communities; such exposure carries risk for psychopathology; and parents and other adults can provide valuable support but are limited in how much they can offset the effects of ongoing violence exposure. Intervention implications are, foremost, that community violence itself must be reduced and, second, that positive relationships with significant adults should be fostered to the degree possible among children living in high-risk, violence prone communities. PMID- 15271608 TI - Story comprehension and the impact of studying on recall in children with attention deficit hyperactivity disorder. AB - This study examined the impact of studying on story comprehension and recall among children with attention deficit hyperactivity disorder (ADHD). Participants in the study were 36 children with ADHD and 43 nonreferred comparison children ages 7 to 11. The participants listened to 1 of 2 folktales and recalled the story both before and after studying a written version of the story for up to 10 min. The stories had been divided into individual events, and each event was coded for the number of causal connections it had to other story events. Each event was presented on a separate page of the study booklet so that time spent on each event could be recorded. All of the transcribed recalls were coded for which story events the participant correctly recalled. For both groups, recall increased as the number of causal connections increased, but the effect of the number of causal connections on recall was stronger for comparison children than for children with ADHD. The results revealed no group differences in studying behavior. However, when recall before studying was included as a predictor of recall after studying, studying was found to be more effective for higher IQ comparison children than for higher IQ children with ADHD, especially at the highest levels of causal connections. The results offer important leads for the development of academic interventions that are specific to the story comprehension deficits of children with ADHD. PMID- 15271609 TI - Predictors of outcome for children with behavior problems served in public mental health. AB - This study investigated whether pervasiveness of problems across settings predicted successful reduction of impairment in school, home, and interactions with others after controlling for other variables that may be stronger predictors. The data of 4,434 youths between the ages of 7 and 17 years in public mental health services in Michigan were examined employing logistic regressions, with 4 sets of predictors, as follows: demographic characteristics, risk factors, therapist's perception of impairment in the youth's caregiving environment, and pervasiveness of the youth's problems. The results indicated that pervasiveness of problems was the strongest predictor of poor outcomes for each domain. In addition, impaired caregiving environment, previous hospitalization for psychiatric or substance use problems, and placement out-of-home were also negatively associated with successful outcome. The implications of the findings for practice and research are discussed. PMID- 15271610 TI - Relation of adolescent mothers' history of antisocial behavior to child conduct problems and social competence. AB - We examined the extent to which maternal antisocial behavior (ASB) is directly related to child conduct problems and social competence and assessed the potential mediating role of negative parenting. The sample included 93 adolescent mothers and their children (44 boys, 49 girls). Mothers retrospectively reported about their ASB since the child's birth, through Grade 2. Negative parenting was coded during a parent-child interaction task (PCIT) at Grade 2. Teachers assessed child outcomes at Grade 3. Maternal ASB during the child's life was directly related to parenting and both child outcomes. In the overall sample, negative parenting partially mediated the relation between maternal ASB and child conduct problems. However, the pattern of relations differed by sex. For boys, maternal ASB was directly related to conduct problems, independent of parenting. For girls, maternal ASB was strongly related to parenting but not conduct problems. Negative parenting did not mediate the relation between maternal ASB and child social competence. Implications for intervention and future research are discussed. PMID- 15271611 TI - Comparing the validity of clinician-generated diagnosis of conduct disorder to the diagnostic interview schedule for children. AB - Clinician diagnoses of conduct disorder (CD) were compared to the diagnoses of CD generated by a structured interview against an observed criterion. Participants were 534 youth from a large residential program in the Midwest for delinquent youth. Rates of in-program CD behaviors were gathered from staff observations of the youth over a 9-month time period. Youth diagnosed with CD by the Diagnostic Interview Schedule for Children (DISC) displayed significantly more CD behaviors in the first 6 months of treatment compared to both youth without an externalizing disorder and youth diagnosed with CD by a clinician. Youth diagnosed with CD by a clinician had rates of CD identical to youth without an externalizing disorder. Clinicians may have weighted contextual information more heavily, as this group was significantly more likely to have an arrest record. Results support the use of structured interviews and provide evidence that typical clinician diagnoses may lack adequate validity. PMID- 15271612 TI - Multi-informant assessment of temperament in children with externalizing behavior problems. AB - We examined the criterion validity of parent and self-report versions of the Junior Temperament and Character Inventory (JTCI) in children with high levels of externalizing problems. The sample included 412 children (206 participants and 206 siblings) participating in a family study of attention and aggressive behavior problems. Criterion validity analyses included (a) correlations between temperament scales and emotional and behavioral scales and (b) correlations between temperament and intelligence and achievement scales. Temperament scales displayed strong convergent and discriminant validity. Across informants and samples, JTCI scales assessing novelty seeking and harm avoidance discriminated between internalizing and externalizing problems. Reward dependence, persistence, cooperativeness, and self-directedness displayed similar patterns of negative relations to emotional and behavioral scales and positive relations to intelligence, achievement, and competence. PMID- 15271613 TI - The utility of measures of child and adolescent anxiety: a meta-analytic review of the Revised Children's Manifest Anxiety Scale, the State-Trait Anxiety Inventory for Children, and the Child Behavior Checklist. AB - We evaluated the ability of the Revised Children's Manifest Anxiety Scale (RCMAS), the State-Trait Anxiety Inventory for Children (STAIC), and the Child Behavior Checklist (CBCL) to (a) discriminate between youth with an anxiety disorder and youth without a disorder, (b) discriminate between youth with an anxiety disorder and youth with either externalizing disorders or affective disorders, and (c) measure treatment change. In addition, variables, including age and sex, were explored as possible moderators of instrument utility. A meta analysis of 43 articles was conducted. A large effect size was found when the instruments were used to compare youth with an anxiety disorder to youth without a disorder. When comparing anxious youth to psychiatric control groups, the picture was mixed; the instruments were found to be useful when discriminating between youth with an anxiety disorder and youth with an externalizing disorder, but not between youth with an anxiety disorder and children and adolescents with an affective disorder. The RCMAS, STAIC, and CBCL were found to be moderately sensitive to treatment gains. PMID- 15271614 TI - The Dating Anxiety Scale for Adolescents: scale development and associations with adolescent functioning. AB - Given the importance of romantic and dating relationships during adolescence, the purpose of the study was to develop and evaluate the psychometric properties of the Dating Anxiety Scale for Adolescents (DAS-A). Participants were 757 high school students (56% girls, ages 15 to 18 years). Adolescents completed the DAS A, the Social Anxiety Scale for Adolescents (SAS-A), a Dating Questionnaire, and the Revised Beck Depression Inventory-II (BDI-II). Factor analysis of the DAS-A yielded a 3-factor solution with acceptable internal consistencies: fear of negative evaluation in dating situations (FNE-Dating); social distress when interacting with real or potential dating partners (SD-Date); and social distress when in a group of mixed-sex peers (SD-Group). Confirmatory factor analysis confirmed the 3-factor solution. Results indicated that younger adolescents reported more dating anxiety than older adolescents, and boys reported more SD Group than girls. Dating anxiety was associated with peer-related anxiety and depressive symptoms and was a significant predictor of adolescents' current and usual dating status, even when controlling for adolescents' peer-related social anxiety. The findings provide support for the reliability and validity of the DAS A. Clinical and research implications are discussed. PMID- 15271615 TI - Relations between parent- and teacher-reported behavioral inhibition and behavioral observations of this temperamental trait. AB - This study examined the relation between the Behavioral Inhibition Scale (BIS), a measure that was specifically developed for assessing behavioral inhibition, and behavioral observations of this trait. Children ages 6 to 10 years participated in a series of experimental tasks assessing behavioral features of the inhibited temperament. The parents and teachers of these children completed the BIS. Results showed that the BIS (in particular the parent version) was significantly related to the observational index of behavioral inhibition. An additional aim of the study was to examine the relation between behavioral inhibition as indexed by the BIS and observations, on the one hand, and measures of anxiety symptoms, fear, and behavioral symptoms, on the other hand. As expected, significant correlations between behavioral inhibition indexes and symptoms of anxiety and withdrawal were observed. The BIS is found to be a brief and easy-to-administer instrument, which seems to provide a meaningful first impression of children's level of behavioral inhibition, and this is particularly true when using the parent version of the BIS. PMID- 15271616 TI - Parenting and children's externalizing problems in substance-abusing families. AB - This study tested associations in path models among positive and negative parenting and children's rule-breaking behavior, aggressive and oppositional behavior, and attention problems for families with a drug-dependent parent. A structural model tested relations between parenting and children's externalizing problems for 251 families with 399 children between the ages of 6 and 18, controlling for nonindependence of ratings at the family level. The model also tested potential moderators, including child age, gender, and ethnicity (White vs. other), and caregiver gender (families with a female substance-abusing caregiver vs. families with a male substance-abusing caregiver). Results indicated that caregiver ratings of monitoring predicted rule-breaking behavior and use of inconsistent discipline predicted ratings of all 3 externalizing syndromes, after controlling parenting and externalizing problems for the effects of the moderators and after controlling significant relations among types of parenting and types of externalizing problems. PMID- 15271617 TI - Therapist verbal behavior early in treatment: relation to successful completion of parent-child interaction therapy. AB - We examined the role of specific therapist verbal behaviors in predicting successful completion of Parent-Child Interaction Therapy (PCIT) in 22 families, including 11 families that successfully completed treatment and 11 that discontinued treatment prematurely. The children were 3 to 6 years old and diagnosed with oppositional defiant disorder (ODD). Chamberlain et al.'s (1986) Therapy Process Code (TPC) was used to measure therapist verbalizations during therapist-parent interactions during the initial clinical interview and the second treatment session. Results indicated that therapists' use of the categories Question, Facilitate, and Support during these sessions accurately predicted treatment dropout versus completion for 73% of families. Findings suggest that the early therapist-parent relationship in PCIT may be critical to successful treatment completion. PMID- 15271618 TI - The cognitive content of thought-listed worry episodes in clinic-referred anxious and nonreferred children. AB - We investigated the cognitive content of worry in 8- to 13-year-old clinic referred anxious (n = 38) and nonreferred (n = 51) children. The children were interviewed individually. They thought-listed their latest worry episodes, rated the uncontrollability of the episodes, and reported on the strategies they used to terminate worry. Content analyses showed that children's worry episodes contained predominantly thoughts reflecting negative outcome anticipation, but other types of thought content also were present. These included problem-solving, ruminating, and self-blaming thoughts. Compared to clinic-referred children, nonreferred children reported more problem solving and less ruminating. In the nonreferred group, increasing age was associated with more problem solving and less ruminating. No such age-related associations were found in the clinic referred group. The 2 groups did not differ in the types of worry-termination strategies they reported, but clinic-referred children were more likely to keep worrying until the perceived threat was removed. The results suggest that the problem-solving function of worry is still emerging during late childhood and that developmental delays in problem solving may be associated with excessive and uncontrollable worrying. PMID- 15271619 TI - Pairwise multiple comparison test procedures: an update for clinical child and adolescent psychologists. AB - Locating pairwise differences among treatment groups is a common practice of applied researchers. Articles published in this journal have addressed the issue of statistical inference within the context of an analysis of variance (ANOVA) framework, describing procedures for comparing means, among other issues. In particular, 1 article (Jaccard & Guilamo-Ramos, 2002b) presented some new methods of performing contrasts of means whereas another presented a framework for obtaining robust tests within this same context (Jaccard & Guilamo-Ramos, 2002a). The purpose of this article is to add to these contributions by presenting some newer methods for conducting pairwise comparisons of means, that is by extending the contributions of the first article and applying the framework of the second article to pairwise multiple comparisons. The newer methods are intended to provide additional sensitivity to detect treatment group differences and provide tests that are robust to the effects of variance heterogeneity, nonnormality, or both. PMID- 15271620 TI - Lesbian cruising: an examination of the concept and methods. AB - The concept of cruising has typically focused on gay male activity and, as such, has been narrowly focused and limited. This view has reinforced negative stereotypical images of gay men and has devalued women's more subtle styles and longer-term relationships. This paper argues that lesbians do cruise but also problematizes the definition of cruising in lesbian culture. Utilizing observations and in-depth interviews, I have broadened the definition of cruising to include a range of behavior as evidenced in both the lesbian and gay community. I have developed a typology of cruising, highlighting seven styles based on method, whether the individual approaches or not, intent, and investment. PMID- 15271621 TI - Summer holiday camps for gay and lesbian young adults: an evaluation of their impact on social support and mental well-being. AB - This study evaluates the effects of participating in a summer holiday camp for Flemish (Belgium) gay and lesbian young adults (N 197). Analysis showed to what extent participation affects the constitution and quality of friendships, the availability of confidant and appraisal support and reported levels of hopelessness, self-esteem, and depression. The study is based on panel data with a six-months interval. At follow-up, respondents reported a higher proportion of gays and lesbians in their friendship network, more satisfaction with these friendships, higher levels of confidant and appraisal support, less hopelessness, higher self-esteem, and lower levels of depression. PMID- 15271622 TI - Willingness to speak out about gay laws reform: some cause for optimism. AB - We examined the extent to which people's private attitudes to gay law reform are influenced by the attitudes of others. Ninety-six university students were told that they were either in a minority or in a majority relative to their university group on their attitudes to gay law reform. Contrary to a number of assumptions made in the social psychological literature, participants who supported gay law reform were more prepared to act in line with their attitudes than were those who opposed gay law reform. Furthermore, anti-gay law reform participants appeared to reassess their attitudes when they were told they were in a minority; in contrast, pro-gay law reform participants were unaffected by the group norm. This suggests that anti-gay law reform attitudes are softer and more easily influenced than are pro-gay law reform attitudes. The implications of these results for activists are discussed. PMID- 15271623 TI - Denial of equal civil rights for lesbian and gay men in The Netherlands,1980 1993. AB - In six national samples (a total of 11,863 respondents) of the Dutch population, aged 16 and over, the denial of equal rights (in housing, inheriting, and adoption) for lesbians and gay men decreased from 1980 and 1985, and remained stable between 1985 and 1993. The denial of equal rights for lesbians and gay men was subscribed to more strongly by social categories that have been exposed to traditional socializing agents and socializing circumstances in which traditional norms prevailed:members of denominations, people who frequently attend church, and older cohorts, especially the ones born before 1948, as well as by those who have presumably not dissociated themselves from these traditional norms, i.e., the lower educated. PMID- 15271624 TI - Sexual minority identity formation in an adult population. AB - This study reflects an initial empirical attempt to assess the validity of a widely accepted model of sexual identity formation proposed by Cass (1979). Using a cross-sectional sample of 143 sexual minority participants, factor analytic and multidimensional scaling results suggest that sexual minorities view the identity formation process as occurring in two phases, rather than multiple discrete linear stages. These two phases can be best characterized by individuals having either an "unintegrated" or "fully integrated" sense of sexual orientation into one's self-identity. Future research is suggested utilizing a longitudinal design in order to conduct a more rigorous test of this model. PMID- 15271625 TI - The identity transformation of biological parents in lesbian/gay stepfamilies. AB - This paper investigates the identity transformation of lesbian and gay biological parents in homosexual stepfamilies. I explore previous models typically used to describe gay and lesbian identity formation, arguing that these models provide little understanding of the experiences of those who have been previously married and have children. Lesbians and gays who leave heterosexual unions and form homosexual stepfamily units undergo a series of transformations. The results show that the transition is relatively positive and less internally stigmatizing and stressful than that experienced by younger, childless lesbians and gays. PMID- 15271626 TI - Changes in psychosocial well-being during stages of gay identity development. AB - The current study evaluated the stage theory of Homosexual Identity Formation (HIF) developed by Cass (1979), in terms of the relationship between stage of gay identity development and psychosocial well-being. Four hundred twenty-five males (12 to 64 years, M = 29.2) reporting sexual attraction to other men provided demographic information and completed psychosocial measures: the Happiness Sadness Scale (McGreal & Joseph, 1993), the Satisfaction with Life Scale (Diener, Emmons, Larsen & Griffin, 1985), the UCLA Loneliness Scale (Russell, Peplau & Ferguson, 1978), the Index of Self-Esteem (Hudson, 1982), and the Gay Identity Questionnaire (Brady & Busse, 1994). Correlation analysis and ANCOVAs controlling for age and nationality demonstrated that the 6 sequential stages of HIF were associated with a U-shaped function for the psychosocial variables. Well-being was high during the initial Confusion and Comparison stages of HIF, was reduced during the middle Tolerance and Acceptance stages, and was again high in the later Pride and Synthesis stages. Each of the psychosocial variables was significantly different according to stage of development (p <.001). Qualitative analysis of subjects' comments also revealed support for the U-shaped pattern. PMID- 15271627 TI - The relative importance of ethnicity and religion in predicting attitudes towards gays and lesbians. AB - Using data from a sample of college students from several campuses throughout the United States, this research examines whether ethnic differences (African Americans versus European Americans) in attitudes toward gays and lesbians are a function of religious attendance. Multiple regressions were run separately for attitudes towards lesbians and attitudes toward gay men. When predicting attitudes toward lesbians, ethnic differences were present in the absence of religious attitudes; however, when religious attitudes were entered into the model, ethnic differences disappeared. In predicting attitudes toward gay men, ethnic differences were never present, while religious attitudes were always statistically significant. We conclude that differences in attitudes toward homosexuals in general, and gay men specifically, are not necessarily a function of ethnicity but possibly of religious attendance and the effect of the "Black church." We end with a discussion of the link between ethnicity, religion, HIV/AIDS, and heterosexism in the African American community. PMID- 15271628 TI - Characteristics of gay, lesbian, and bisexual Asians, Asian Americans, and immigrants from Asia to the USA. AB - An exploratory study of self-identified lesbian, gay, and bisexual Asians and Asian Americans surveyed respondents (60 women and 254 men) using a questionnaire in four languages from nineteen different sources in Korea, Japan, China, and the US. Respondents were compared in terms of country of residence, whether they immigrated to the US, having a same-sex lover, living with the lover, being open about their sexual orientation to the family, and age. Respondents in the US were generally more open about their sexual orientation. Openness to the family was related to other variables suggesting an affirmative lesbian, gay, or bisexual identity. PMID- 15271630 TI - Internet use by physicians and its impact on medical practice-an exploratory study. AB - Internet use by physicians has played a vital role in medical practices for many years. A number of related studies have emerged to examine the impact of Internet use on medical practice. However, there is yet to be a comprehensive study on the impact of Internet use by physicians on their medical practice. This study examines a preliminary step to explore the major implications of physicians' Internet use on the traditional areas, such as health education and learning, physician-patient relationship, and medical marketing. Barriers to Internet use are also investigated. Implication of use of the Internet in the medical practice and limitations of this study are discussed as well. PMID- 15271631 TI - Service quality assessment of workers compensation health care delivery programs in New York using SERVQUAL. AB - Preferred provider organizations (PPOs) provide healthcare services to an expanding proportion of the U.S. population. This paper presents a programmatic assessment of service quality in the workers' compensation environment using two different models: the PPO program model and the fee-for-service (FFS) payor model. The methodology used here will augment currently available research in workers' compensation, which has been lacking in measuring service quality determinants and assessing programmatic success/failure of managed care type programs. Results indicated that the SERVQUAL tool provided a reliable and valid clinical quality assessment tool that ascertained that PPO marketers should focus on promoting physician outreach (to show empathy) and accessibility (to show reliability) for injured workers. PMID- 15271632 TI - The identification of factors linked to the potential acceptance of transgenic biopharmaceuticals: an exploratory study. AB - In this exploratory study, Rogers' diffusion of innovation theory was used to identify which factors are likely to contribute to the potential acceptance of transgenic biopharmaceuticals (TG-Bs). These products are not yet available to the general public. A scale was designed to assess three of five core attributes related to the potential adoption rate of innovations (Rogers 1995), as well as to measure potential acceptance characteristics for biotechnology products. These attributes were relative advantage, compatibility with existing values, and complexity. In addition, two other characteristics were included: knowledge (Gartrell and Gartrell 1979) and perceived risks (Bauer 1960). The survey was completed by 74 consumers (78% response rate) using convenience sampling. The research findings show that Rogers' three core attributes are supported, but that knowledge andperceived risks were excluded from the model. The model for transgenic biopharmaceuticals consists of: 1. Consumer-related benefits (positively correlated to potential adoption). 2. New types of animals (negatively correlated to potential 3. Perceived complexity (negatively correlated to potential adoption). All the scaled items developed for this study were highly significant, which indicates that they can be used successfully by other researchers working in this field. As TG-Bs are a discontinuous innovation, biotechnology companies may need to present the benefits of these products, as well as the ease of their use prior to their launch, in order to increase their potential acceptance by consumers. PMID- 15271633 TI - Developing a profile of consumer intention to seek our health information beyond a doctor. AB - The health care consumer of the new millennium is becoming increasingly involved in his/her health choices. The explosion in the number of media outlets offering health-specific information has further propelled the growth in active health orientation. The goal of this research is to understand the profile of the actively oriented health care consumer. It constructs a psychographic plot of additional health information seeking by investigating variables such as health consciousness, environmental consciousness, and consumerism. Based on the study results, strategic recommendations are made for health information targeting and delivery. PMID- 15271634 TI - An investigation of factors that influence the consumption of dietary supplements. AB - This study finds that consumers take dietary supplements to improve their physical health and gain peace of mind. Several factors influence the consumption of dietary supplements, but the advice of a physician underlies most consumer behavior. Few dietary supplement users over-consume these types of products, but those who do could be partaking in potentially dangerous consumption practices. More importantly, those who consume multivitamins, concentrates, and herbs/flowers/ roots express different consumer behaviors from those who only take multivitamins and concentrates. Conclusions are drawn in light of the media's impact on the behaviors of dietary supplement consumers. PMID- 15271637 TI - General practitioners' knowledge, confidence and attitudes in the diagnosis and management of dementia. AB - OBJECTIVE: To measure general practitioners' knowledge of, confidence with and attitudes to the diagnosis and management of dementia in primary care. SETTING: 20 general practices of varying size and prior research experience in Central Scotland, and 16 similarly varied practices in north London. PARTICIPANTS: 127 general practitioners who had volunteered to join a randomised controlled trial of educational interventions about dementia diagnosis and management. METHODS: Self-completion questionnaires covering knowledge, confidence and attitudes were retrieved from practitioners prior to the educational interventions. RESULTS: General practitioners' knowledge of dementia diagnosis and management is good, but poor awareness of its epidemiology leads to an over-estimate of caseload. Knowledge of local diagnostic and support services is less good, and one third of general practitioners expressed limited confidence in their diagnostic skills, whilst two-thirds lacked confidence in management of behaviour and other problems in dementia. The main difficulties identified by general practitioners were talking with patients about the diagnosis, responding to behaviour problems and coordinating support services. General practitioners perceived lack of time and lack of social services support as the major obstacles to good quality care more often than they identified their own unfamiliarity with current management or with local resources. Attitudes to the disclosure of the diagnosis, and to the potential for improving the quality of life of patients and carers varied, but a third of general practitioners believed that dementia care is within a specialist's domain, not that of general practice. More experienced and male general practitioners were more pessimistic about dementia care, as were general practitioners with lower knowledge about dementia. Those reporting greater difficulty with dementia diagnosis and management and those with lower knowledge scores were also less likely to express attitudes endorsing open communication with patient and carer. CONCLUSION: Educational support for general practitioners should concentrate on epidemiological knowledge, disclosure of the diagnosis and management of behaviour problems in dementia. The availability and profile of support services, particularly social care, need to be enhanced, if earlier diagnosis is to be pursued as a policy objective in primary care. PMID- 15271638 TI - Hydrochlorothiazide induced hepato-cholestatic liver injury. AB - Hydrochlorothiazide and other thiazide-like diuretics are considered as a first line drug for initial therapy in uncomplicated arterial hypertension [1]. There are several reports [2-6] of thiazide-induced cholecystitis, but here we report a case of serious hepatotoxicity associated with hydrochlorothiazide treatment. PMID- 15271639 TI - Effects of dementia on perceived daily pain in home-dwelling elderly people: a population-based study. AB - BACKGROUND: Pain is a significant problem in the elderly, but the impact of dementia on perceived pain has not been studied in population-based study settings. OBJECTIVES: To analyse the prevalence of daily pain and analgesic use among home-dwelling older people with and without dementia. DESIGN: A cross sectional population-based survey. SETTING: Population of Kuopio city, Finland. SUBJECTS: 523 home-dwelling subjects aged 75 years and older. METHODS: Structured clinical examination and interview. RESULTS: Prevalence rates for any pain, any daily pain, pain every day interfering with routine activities, and daily pain at rest were significantly lower in those subjects with dementia (43%, 23%, 19% and 4%, respectively) compared to those subjects without dementia (69%, 40%, 36% and 13%, respectively). The subjects with dementia were less likely to use analgesics (33%) than the non-demented (47%). CONCLUSION: Dementia was related to a lower prevalence of reported pain and analgesic use among home-dwelling elderly people. PMID- 15271640 TI - Lesson of the week: perils of pessaries. AB - Vaginal pessaries are widely considered to be a safe alternative to surgery in older women. We report a case of near fatal septicaemia in a 75-year-old woman associated with a shelf pessary, the presence of which was identified during an exploratory laparotomy. This case highlights the importance of the gynaecological history and examination when assessing older women with septicaemia of unknown source. PMID- 15271641 TI - Cutaneous angiosarcoma presenting as an unusual facial bruise. AB - Angiosarcoma is a rare vascular tumour accounting for 2% of soft tissue sarcomas, which together represent less than 1% of all cancers. Although well described in specialist literature, it is unusual in general medical practice. We describe a case in which the initial appearance as a bruise gave rise to diagnostic uncertainty and delay. PMID- 15271642 TI - Left ventricular systolic dysfunction and atrial fibrillation in older people in the community--a need for screening? AB - BACKGROUND: Heart failure and stroke are major causes of morbidity and mortality in older people. Angiotensin converting enzyme inhibitors improve symptoms and survival in left ventricular systolic dysfunction. Anticoagulants are effective in stroke prevention in atrial fibrillation with aspirin being a less effective alternative. OBJECTIVES: To determine the prevalence of left ventricular systolic dysfunction, health services utilisation and prescribing of diuretics and angiotensin converting enzyme inhibitors in left ventricular systolic dysfunction, and the prevalence of atrial fibrillation and anti platelet/thrombotic therapy in atrial fibrillation in older people in the community. METHODS: 500 subjects were drawn by two-stage random sampling from 5,002 subjects aged 70 years and over living at home. Subjects were screened for atrial fibrillation and left ventricular systolic dysfunction using electrocardiography and echocardiography. RESULTS: The population prevalence amongst older people of left ventricular systolic dysfunction was 9.8% and of atrial fibrillation 7.8%. More than two-thirds of those with left ventricular systolic dysfunction were not on angiotensin converting enzyme inhibitors. Of those in atrial fibrillation, 35% were taking aspirin, 24% were taking warfarin and 41% were on neither aspirin nor warfarin. Nearly 90% of older people in the community have had contact with their general practitioner over the past year, and over half of those with left ventricular systolic dysfunction have had contact with hospital-based services over the past 2 years. CONCLUSIONS: Left ventricular systolic dysfunction is under-treated in older people in the community. Despite the high level of contact with hospital and community-based services, the majority of those with systolic left ventricular dysfunction are not on angiotensin converting enzyme inhibitors and a significant proportion of those in atrial fibrillation are not on any treatment for stroke prevention. PMID- 15271643 TI - Guidelines for reporting statistics in journals published by the American Physiological Society. PMID- 15271644 TI - Pancreatic beta-cell growth and survival in the onset of type 2 diabetes: a role for protein kinase B in the Akt? AB - The control of pancreatic beta-cell growth and survival in the adult plays a pivotal role in the pathogenesis of type 2 diabetes. In certain insulin-resistant states, such as obesity, the increased insulin-secretory demand can often be compensated for by an increase in beta-cell mass, so that the onset of type 2 diabetes is avoided. This is why approximately two-thirds of obese individuals do not progress to type 2 diabetes. However, the remaining one-third of obese subjects that do acquire type 2 diabetes do so because they have inadequate compensatory beta-cell mass and function. As such, type 2 diabetes is a disease of insulin insufficiency. Indeed, it is now realized that, in the vast majority of type 2 diabetes cases, there is a decreased beta-cell mass caused by a marked increase in beta-cell apoptosis that outweighs rates of beta-cell mitogenesis and neogenesis. Thus a means of promoting beta-cell survival has potential therapeutic implications for treating type 2 diabetes. However, understanding the control of beta-cell growth and survival at the molecular level is a relatively new subject area of research and still in its infancy. Notwithstanding, recent advances have implicated signal transduction via insulin receptor substrate-2 (IRS-2) and downstream via protein kinase B (PKB, also known as Akt) as critical to the control of beta-cell survival. In this review, we highlight the mechanism of IRS-2, PKB, and anti-apoptotic PKB substrate control of beta-cell growth and survival, and we discuss whether these may be targeted therapeutically to delay the onset of type 2 diabetes. PMID- 15271645 TI - Role of incretin hormones in the regulation of insulin secretion in diabetic and nondiabetic humans. AB - The available evidence suggests that about two-thirds of the insulin response to an oral glucose load is due to the potentiating effect of gut-derived incretin hormones. The strongest candidates for the incretin effect are glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide 1 (GLP-1). In patients with type 2 diabetes, however, the incretin effect is lost or greatly impaired. It is hypothesized that this loss explains an important part of the impaired insulin secretion in patients. Further analysis of the incretin effects in patients has revealed that the secretion of GIP is near normal, whereas the secretion of GLP-1 is decreased. On the other hand, the insulintropic effect of GLP-1 is preserved, whereas the effect of GIP is greatly reduced, mainly because of a complete loss of the normal GIP-induced potentiation of second-phase insulin secretion. These two features, therefore, explain the incretin defect of type 2 diabetes. Strong support for the hypothesis that the defect plays an important role in the insulin deficiency of patients is provided by the finding that administration of excess GLP-1 to patients may completely restore the glucose induced insulin secretion as well as the beta-cells' sensitivity to glucose. Because of this, analogs of GLP-1 or GLP-1 receptor activations are currently being developed for diabetes treatment, so far with very promising results. PMID- 15271646 TI - Components of a platelet-activating factor-signaling loop are assembled in the ovine endometrium late in the estrous cycle. AB - Pulsatile release of uterine prostaglandin F(2alpha) (PGF(2alpha)) induces luteolysis in ruminants. Exogenous PAF is well known to cause PGF(2alpha) release from the ovine uterus. This study examines whether the components of a PAF signaling loop exist in sheep at the time luteolysis is normally initiated. Day 14 of the cycle was the first day the uterus responded to an infusion of PAF, inducing a significant short-term increase in circulating levels of the PGF(2alpha) metabolite. There was a significant increase of PAF concentration (P < 0.001) in the endometrium and PAF release by tissue explants (P < 0.001) from day 10 to day 16 of the cycle. Endometrial tissue PAF receptor mRNA expression was induced (P < 0.01) by estradiol and progesterone treatment of animals, and transcripts were present between days 10 and 16 of the estrous cycle. Western analysis of endometrial tissue showed marked upregulation of PAF receptor protein expression from day 14 of the cycle, and immunolocalization studies showed that the receptor expression was predominantly around the endometrial glands. PAF:acetylhydrolase was primarily located within the lumen of the endometrial glands. The study shows that a PAF-signaling loop was assembled within the ovine endometrium at the time that PGF(2alpha) pulsatility was first observed. PMID- 15271647 TI - Hormone-sensitive lipase-independent adipocyte lipolysis during beta-adrenergic stimulation, fasting, and dietary fat loading. AB - In white adipose tissue, lipolysis can occur by hormone-sensitive lipase (HSL) dependent or HSL-independent pathways. To study HSL-independent lipolysis, we placed HSL-deficient mice in conditions of increased fatty acid flux: beta adrenergic stimulation, fasting, and dietary fat loading. Intraperitoneal administration of the beta(3)-adrenergic agonist CL-316243 caused a greater increase in nonesterified fatty acid level in controls (0.33 +/- 0.05 mmol/l) than in HSL(-/-) mice (0.12 +/- 0.01 mmol/l, P < 0.01). Similarly, in isolated adipocytes, lipolytic response to CL-316243 was greatly reduced in HSL(-/-) mice compared with controls. Fasting for 0.01) parvocellular divisions. In addition, we analyzed spinally projecting neurons of the RVLM and found a significantly greater percentage were Fos positive in water-deprived rats than in control rats (26 +/- 3 vs. 3 +/- 1%, respectively; P < 0.001). Collectively, the present findings indicate that water deprivation evokes a distinct cellular response in sympathetic-regulatory neurons of the PVN and RVLM. PMID- 15271658 TI - LDL receptor-related protein LRP6 regulates proliferation and survival through the Wnt cascade in vascular smooth muscle cells. AB - Initial studies have established expression of low-density lipoprotein (LDL) receptor-related protein 6 (LRP6) in vascular smooth muscle cells (VSMCs). We hypothesized that LRP6 is a critical mediator governing the regulation of the canonical Wnt/beta-catenin/T cell factor 4 (Tcf-4) cascade in the vasculature. This hypothesis was based on our previous work demonstrating a role for the beta catenin/Tcf-4 pathway in vascular remodeling as well as work in other cell systems establishing a role for LRP family members in the Wnt cascade. In line with our hypothesis, LRP6 upregulation significantly increased Wnt-1-induced Tcf activation. Moreover, a dominant interfering LRP6 mutant lacking the carboxyl intracellular domain (LRP6DeltaC) abolished Tcf activity. LRP6-induced stimulation of Tcf was blocked in VSMCs harboring constitutive expression of a dominant negative Tcf-4 transgene lacking the beta-catenin binding domain, suggesting that LRP6-induced activation of Tcf was mediated through a beta catenin-dependent signal. Expression of the dominant interfering LRP6DeltaC transgene was sufficient to abolish the Wnt-induced survival as well as cyclin D1 activity and cell cycle progression. In conclusion, these findings provide the first evidence of a role for an LDL receptor-related protein in the regulation of VSMC proliferation and survival through the evolutionary conserved Wnt signaling cascade. PMID- 15271659 TI - Combined NO and PG inhibition augments alpha-adrenergic vasoconstriction in contracting human skeletal muscle. AB - Sympathetic alpha-adrenergic vasoconstrictor responses are blunted in the vascular beds of contracting muscle (functional sympatholysis). We tested the hypothesis that combined inhibition of nitric oxide (NO) and prostaglandins (PGs) restores sympathetic vasoconstriction in contracting human muscle. We measured forearm blood flow via Doppler ultrasound and calculated the reduction in forearm vascular conductance in response to alpha-adrenergic receptor stimulation during rhythmic handgrip exercise (6.4 kg) and during a control nonexercise vasodilator condition (using intra-arterial adenosine) before and after combined local inhibition of NO synthase (NOS; via N(G)-nitro-L-arginine methyl ester) and cyclooxygenase (via ketorolac) in healthy men. Before combined inhibition of NO and PGs, the forearm vasoconstrictor responses to intra-arterial tyramine (which evoked endogenous noradrenaline release), phenylephrine (a selective alpha1 agonist), and clonidine (an alpha2-agonist) were significantly blunted during exercise compared with adenosine treatment. After combined inhibition of NO and PGs, the vasoconstrictor responses to all alpha-adrenergic receptor stimuli were augmented by approximately 10% in contracting muscle (P <0.05), whereas the responses to phenylephrine and clonidine were also augmented by approximately 10% during passive vasodilation in resting muscle (P <0.05). In six additional subjects, PG inhibition alone did not alter the vasoconstrictor responses in resting or contracting muscles. Thus in light of our previous findings, it appears that inhibition of either NO or PGs alone does not affect functional sympatholysis in healthy humans. However, the results from the present study indicate that combined inhibition of NO and PGs augments alpha-adrenergic vasoconstriction in contracting muscle but does not completely restore the vasoconstrictor responses compared with those observed during passive vasodilation in resting muscle. PMID- 15271660 TI - cAMP protects endothelial barrier functions by preventing Rac-1 inhibition. AB - cAMP enhances endothelial barrier properties and is protective against various inflammatory mediators both in vivo and in vitro. However, the mechanisms whereby cAMP stabilizes the endothelial barrier are largely unknown. Recently we demonstrated that the Rho family GTPase Rac-1 is required for maintenance of endothelial barrier functions in vivo and in vitro. Therefore, in the present study we investigated the effect of forskolin (5 microM)- and rolipram (10 microM)-induced cAMP increase on reduction of barrier functions in response to Rac-1 inhibition by Clostridium sordellii lethal toxin (LT). Forskolin and rolipram treatment blocked LT (200 ng/ml)-induced hydraulic conductivity (Lp) increase in mesenteric microvessels in vivo. Likewise, LT-induced intercellular gap formation in monolayers of cultured microvascular myocardial endothelial (MyEnd) cells and LT-induced loss of adhesion of vascular endothelial cadherin coated microbeads were abolished. Inhibition of PKA by myristoylated inhibitor peptide (14-22) of PKA (100 microM) reduced the protective effect of cAMP on LT induced Lp increase in vivo and gap formation in vitro, indicating that the effect of cAMP on Rac-1 inhibition was PKA dependent. Glucosylation assays demonstrated that cAMP prevents inhibitory Rac-1 glucosylation by LT, indicating that one way that cAMP enhances endothelial barrier functions may be by regulating Rac-1 signaling. Our study suggests that cAMP may provide its well established protective effects at least in part by regulation of Rho proteins. PMID- 15271661 TI - Increase of PTP levels in vascular injury and in cultured aortic smooth muscle cells treated with specific growth factors. AB - Migration and proliferation of vascular smooth muscle cells are key events in injury-induced neointima formation. Several growth factors and ANG II are thought to be involved in neointima formation. A recent report indicated that vascular injury is associated with increased mRNA levels of protein tyrosine phosphatase (PTP)-1B (PTP-1B). In the present study, we tested the following hypotheses: 1) rat carotid artery injury induces the expression of PTP-1B, Src homology-2 domain phosphatase (SHP-2), and PTP-proline, glutamate, serine, and threonine sequence (PEST) protein; and 2) polypeptide growth factors as well as ANG II increase the levels of tyrosine phosphatases in cultured rat aortic smooth muscle cells. We found that vascular injury induced by balloon catheter increases the protein levels of aforementioned phosphatases and that these effects occur in a PTP specific, as well as temporally and regionally specific, manner. Moreover, treatment of cultured primary rat aortic smooth muscle cells with PDGF or bFGF, but not with IGF1, EGF, or ANG II, increases PTP-1B, SHP-2, and PTP-PEST protein levels. These results suggest that increased PDGF and bFGF levels, occurring after vascular injury, may induce expression of several PTPs. PMID- 15271662 TI - Adenosine A1/A2a receptor agonist AMP-579 induces acute and delayed preconditioning against in vivo myocardial stunning. AB - The purpose of this study was to determine whether the adenosine A1/A2a receptor agonist AMP-579 induces acute and delayed preconditioning against in vivo myocardial stunning. Regional stunning was produced by 15 min of coronary artery occlusion and 3 h of reperfusion (RP) in anesthetized open-chest pigs. In acute protection studies, animals were pretreated with saline, low-dose AMP-579 (15 microg/kg iv bolus 10 min before ischemia), or high-dose AMP-579 (50 microg/kg iv at 14 microg/kg bolus + 1.2 microg.kg(-1).min(-1) for 30 min before coronary occlusion). The delayed preconditioning effects of AMP-579 were evaluated 24 h after administration of saline vehicle or high-dose AMP-579 (50 microg/kg iv). Load-insensitive contractility was assessed by measuring regional preload recruitable stroke work (PRSW) and PRSW area. Acute preconditioning with AMP-579 dose dependently improved regional PRSW: 129 +/- 5 and 100 +/- 2% in high- and low-dose AMP-579 groups, respectively, and 78 +/- 5% in the control group at 3 h of RP. Administration of the adenosine A1 receptor antagonist 8-cyclopentyl-1,3 dipropylxanthine (0.7 mg/kg) blocked the acute protective effect of high-dose AMP 579, indicating that these effects are mediated through A1 receptor activation. Delayed preconditioning with AMP-579 significantly increased recovery of PRSW area: 64 +/- 5 vs. 33 +/- 5% in control at 3 h of RP. In isolated perfused rat heart studies, kinetics of the onset and washout of AMP-579 A1 and A2a receptor mediated effects were distinct compared with those of other adenosine receptor agonists. The unique nature of the adenosine agonist AMP-579 may play a role in its ability to induce delayed preconditioning against in vivo myocardial stunning. PMID- 15271663 TI - Biphasic modulation of vascular nitric oxide catabolism by oxygen. AB - Endothelium-derived nitric oxide (NO) plays an important role in the regulation of vascular tone. Lack of NO bioavailability can result in cardiovascular disease. NO bioavailability is determined by its rates of generation and catabolism; however, it is not known how the NO catabolism rate is regulated in the vascular wall under normoxic, hypoxic, and anaerobic conditions. To investigate NO catabolism under different oxygen concentrations, studies of NO and O2 consumption by the isolated rat aorta were performed using electrochemical sensors. Under normoxic conditions, the rate of NO consumption in solution was enhanced in the presence of the rat aorta. Under hypoxic conditions, NO consumption decreased in parallel with the O2 concentration. Like the inhibition of mitochondrial respiration by NO, the inhibitory effects of NO on aortic O2 consumption increased as O2 concentration decreased. Under anaerobic conditions, however, a paradoxical reacceleration of NO consumption occurred. This increased anaerobic NO consumption was inhibited by the cytochrome c oxidase inhibitor NaCN but not by the free iron chelator deferoxamine, the flavoprotein inhibitor diphenylene iodonium (10 microM), or superoxide dismutase (200 U/ml). The effect of O2 on the NO consumption could be reproduced by purified cytochrome c oxidase (CcO), implying that CcO is involved in aortic NO catabolism. This reduced NO catabolism at low O2 tensions supports the maintenance of effective NO levels in the vascular wall, reducing the resistance of blood vessels. The increased anaerobic NO catabolism may be important for removing excess NO accumulation in ischemic tissues. PMID- 15271664 TI - FKBP12.6 overexpression decreases Ca2+ spark amplitude but enhances [Ca2+]i transient in rat cardiac myocytes. AB - Ryanodine receptors/Ca2+-release channels (RyR2) from the sarcoplasmic reticulum (SR) provide the Ca2+ required for contraction at each cardiac twitch. RyR2 are regulated by a variety of proteins, including the immunophilin FK506 binding protein (FKBP12.6). FKBP12.6 seems to be important for coupled gating of RyR2 and its deficit and alteration may be involved in heart failure. The role of FKBP12.6 on Ca2+ release has not been analyzed directly, but rather it was inferred from the effects of immunophilins, such us FK506 and rapamycin, which, among other effects, dissociates FKBP12.6 from the RyR2. Here, we investigated directly the effects of FKBP12.6 on local (Ca2+ sparks) and global [intracellular Ca2+ concentration ([Ca2+]i) transients] Ca2+ release in single rat cardiac myocytes. The FKBP12.6 gene was transfected in single myocytes using the adenovirus technique with a reporter gene strategy based on green fluorescent protein (GFP) to check out the success of transfections. Control myocytes were transfected with only GFP (Ad-GFP). Rhod-2 was used as the Ca2+ indicator, and cells were viewed with a confocal microscope. We found that overexpression of FKBP12.6 decreases the occurrence, amplitude, duration, and width of spontaneous Ca2+ sparks. FK506 had diametrically opposed effects. However, overexpression of FKBP12.6 increased the [Ca2+]i transient amplitude and accelerated its decay in field-stimulated cells. The associated cell shortening was increased. SR Ca2+ load, estimated by rapid caffeine application, was increased. In conclusion, FKBP12.6 overexpression decreases spontaneous Ca2+ sparks but increases [Ca2+]i transients, in relation with enhanced SR Ca2+ load, therefore improving excitation-contraction coupling. PMID- 15271665 TI - Time course of changes in collateral blood flow and isolated vessel size and gene expression after femoral artery occlusion in rats. AB - The objectives of this study were to assess the time course of enlargement and gene expression of a collateral vessel that enlarges following occlusion of the femoral artery and to relate these responses to the increases in collateral dependent blood flow to the calf muscles in vivo. We employed exercise training to stimulate collateral vessel development. Rats were exercise trained or kept sedentary for various times of up to 25 days postbilateral occlusion (n=approximately 9/time point). Collateral blood flow to the calf muscles, determined with microspheres, increased modestly over the first few days to approximately 40 ml.min(-1).100 g(-1) in sedentary animals; the increase continued over time to approximately 80 ml.min(-1).100 g(-1) in the trained animals. Diameters of the isolated collateral vessels increased progressively over time, whereas an increased vessel compliance observed at low pressures was similar across time. These responses were greater in the trained animals. The time course of upregulation of vascular endothelial growth factor and placental growth factor, and particularly endothelial nitric oxide synthase and fms-like tyrosine kinase 1, mRNAs in the isolated collateral vessel implicates these factors as integral to the arteriogenic process. Collateral vessel enlargement and increased compliance at low pressures contribute to the enlarged circuit available for collateral blood flow. However, modulation of the functioning collateral vessel diameter, by smooth muscle tone, must occur to account for the observed increases in collateral blood flow measured in vivo. PMID- 15271666 TI - Heterogeneity of action potential durations in isolated mouse left and right atria recorded using voltage-sensitive dye mapping. AB - An imaging system for di-4-ANEPPS (4-[beta-[2-(di-n-butylamino)-6 naphthylvinyl]pyridinium]) voltage-sensitive dye recordings has been adapted for recording from an in vitro mouse heart preparation that consists of both atria in isolation. This approach has been used to study inter- and intra-atrial activation and conduction and to monitor action potential durations (APDs) in the left and right atrium. The findings from this study confirm some of our previous findings in isolated mouse atrial myocytes and demonstrate that many electrophysiological properties of mouse atria closely resemble those of larger mammals. Specifically, we made the following observations: 1) Activation in mouse atria originates in the sinoatrial node and spreads into the right atrium and, after a delay, into the left atrium. 2) APD in the left atrium is shorter than in the right atrium. 3) Sites in the posterior walls have longer APDs than sites in the atrial appendages. 4) Superfusion of this preparation with 4-aminopyridine and tetraethylammonium resulted in increases in APD, consistent with their inhibitory effects on the K+ currents known to be expressed in mouse atria. 5) The muscarinic agonist carbachol shortened APD in all areas of the preparation, except the left atrial appendage, in which carbachol had no statistically significant effect on APD. These results validate a new approach for monitoring activation, conduction, and repolarization in mouse atria and demonstrate that the physiological and pharmacological properties of mouse atria are sufficiently similar to those of larger animals to warrant further studies using this preparation. PMID- 15271667 TI - Decreased cardiac mitochondrial NADP+-isocitrate dehydrogenase activity and expression: a marker of oxidative stress in hypertrophy development. AB - Mitochondrial dysfunction subsequent to increased oxidative stress and alterations in energy metabolism is considered to play a role in the development of cardiac hypertrophy and its progression to failure, although the sequence of events remains to be elucidated. This study aimed at characterizing the impact of hypertrophy development on the activity and expression of mitochondrial NADP+ isocitrate dehydrogenase (mNADP+-ICDH), a metabolic enzyme that controls redox and energy status. We expanded on our previous finding of its inactivation through posttranslational modification by the lipid peroxidation product 4 hydroxynonenal (HNE) in 7-wk-old spontaneously hypertensive rat (SHR) hearts before hypertrophy development (Benderdour et al. J Biol Chem 278: 45154-45159, 2003). In this study, we used 7-, 15-, and 30-wk-old SHR and Sprague-Dawley (SD) rats with abdominal aortic coarctation. Compared with age-matched control Wistar Kyoto (WKY) rats, SHR hearts showed a significant 25% decrease of mNADP+-ICDH activity, which preceded in time 1) the decline in its protein and mRNA expression levels (between 10% and 35%) and 2) the increase in hypertrophy markers. The chronic and persistent loss of mNADP+-ICDH activity in SHR was associated with enhanced tissue accumulation of HNE-mNADP+-ICDH and total HNE protein adducts at all ages and contrasted with the profile of changes in the activity of other mitochondrial enzymes involved in antioxidant or energy metabolism. Two-way ANOVA of the data also revealed a significant effect of age on most parameters measured in SHR and WKY hearts. The mNADP+-ICDH activity, protein, and mRNA expression were reduced between 25% and 35% in coarctated SD rats and were normalized by treatment of SHR or coarctated SD rats with renin angiotensin system inhibitors, which prevented or attenuated hypertrophy. Altogether, our data show that cardiac mNADP+-ICDH activity and expression are differentially and sequentially affected in hypertrophy development and, to a lesser extent, with aging. Decreased cardiac mNADP+-ICDH activity, which is attributed at least in part to HNE adduct formation, appears to be a relevant early and persistent marker of mitochondrial oxidative stress-related alterations in hypertrophy development. Potentially, this could also contribute to the aetiology of cardiomyopathy. PMID- 15271668 TI - cAMP modulates cGMP-mediated cerebral arteriolar relaxation in vivo. AB - No studies have specifically addressed whether cAMP can influence nitric oxide (NO)/cGMP-induced cerebral vasodilation. In this study, we examined whether cAMP can enhance or reduce NO-induced cerebral vasodilation in vivo via interfering with cGMP efflux or through potentiating phosphodiesterase 5 (PDE5)-mediated cGMP breakdown, respectively, in cerebral vascular smooth muscle cells (CVSMCs). To that end, we evaluated, in male rats, the effects of knockdown [via antisense oligodeoxynucleotide (ODN) applications] of the cGMP efflux protein multidrug resistance protein 5 (MRP5) and PDE5 inhibition on pial arteriolar NO donor [S nitroso-N-acetyl penicillamine (SNAP)]-induced dilations in the absence and presence of cAMP elevations via forskolin. Pial arteriolar diameter changes were measured using well-established protocols in anesthetized rats. In control (missense ODN treated) rats, forskolin elicited a leftward shift in the SNAP dose response curves (approximately 50% reduction in SNAP EC50). However, in MRP5 knockdown rats, cAMP increases were associated with a substantial reduction in SNAP-induced vasodilations (reflected as a significant 35-50% lower maximal response). In the presence of the PDE5 inhibitor MY-5445, the repression of the NO donor response accompanying forskolin was prevented. These findings suggest that cAMP has opposing effects on NO-stimulated cGMP increases. On the one hand, cAMP limits CVSMC cGMP loss by restricting cGMP efflux. On the other, cAMP appears to enhance PDE5-mediated cGMP breakdown. However, because increased endogenous cAMP seems to potentiate NO/cGMP-induced arteriolar relaxation when MRP5 expression is normal, the effect of cAMP to reduce cGMP efflux appears to predominate over cAMP stimulation of cGMP hydrolysis. PMID- 15271669 TI - Dynamical effects of diffusive cell coupling on cardiac excitation and propagation: a simulation study. AB - Cell coupling is considered to be important for cardiac action potential propagation and arrhythmogenesis. We carried out computer simulations to investigate the effects of stimulation strength and cell-to-cell coupling on action potential duration (APD) restitution, APD alternans, and stability of reentry in models of isolated cell, one-dimensional cable, and two-dimensional tissue. Phase I formulation of the Luo and Rudy action potential model was used. We found that stronger stimulation resulted in a shallower APD restitution curve and onset of APD alternans at a faster pacing rate. Reducing diffusive coupling between cells prolonged APD. Weaker diffusive currents along the direction of propagation steepened APD restitution and caused APD alternans to occur at a slower pacing rate in tissue. Diffusive current due to curvature changed APD but had little effect on APD restitution slope and onset of instability. Heterogeneous cell coupling caused APD inhomogeneities in space. Reduction in coupling strength either uniformly or randomly had little effect on the rotation period and stability of a reentry, but random cell decoupling slowed the rotation period and, thus, stabilized the reentry, preventing it from breaking up into multiple waves. Therefore, in addition to its effects on action potential conduction velocity, diffusive cell coupling also affects APD in a rate-dependent manner, causes electrophysiological heterogeneities, and thus modulates the dynamics of cardiac excitation. These effects are brought about by the modulation of ionic current activation and inactivation. PMID- 15271670 TI - Insulin resistance does not diminish eNOS expression, phosphorylation, or binding to HSP-90. AB - Previously, using an animal model of syndrome X, the obese Zucker rat (OZR), we documented impaired endothelium-dependent vasodilation. The aim of this study was to determine whether reduced expression or altered posttranslational regulation of endothelial nitric oxide synthase (eNOS) underlies the vascular dysfunction in OZR rats. There was no significant difference in the relative abundance of eNOS in hearts, aortas, or skeletal muscle between lean Zucker rats (LZR) and OZR regardless of age. There was no difference in eNOS mRNA levels, as determined by real-time PCR, between LZR and OZR. The inability of insulin resistance to modulate eNOS expression was also documented in two additional in vivo models, the ob/ob mouse and the fructose-fed rat, and in vitro via adenoviral expression of protein tyrosine phosphatase 1B in endothelial cells. We next investigated whether changes in the acute posttranslational regulation of eNOS occurs with insulin resistance. Phosphorylation of eNOS at S632 (human S633) and T494 was not different between LZR and OZR; however, phosphorylation of S1176 was significantly enhanced in OZR. Phosphorylation of S1176 was not different in the ob/ob mouse or in fructose-fed rats. The association of heat shock protein 90 with eNOS, a key regulatory step controlling nitric oxide and aberrant O2- production, was not different between OZR and LZR. Taken together, these results suggest that changes in eNOS expression or posttranslation regulation do not underlie the vascular dysfunction seen with insulin resistance and that other mechanisms, such as altered localization, reduced availability of cofactors, substrates, and the elevated production of O2-, may be responsible. PMID- 15271671 TI - Protein kinase C-alpha-induced hypertrophy of neonatal rat ventricular myocytes. AB - Protein kinase C (PKC) isoenzymes play a critical role in cardiomyocyte hypertrophy. At least three different phorbol ester-sensitive PKC isoenzymes are expressed in neonatal rat ventricular myocytes (NRVMs): PKC-alpha, -delta, and epsilon. Using replication-defective adenoviruses (AdVs) that express wild-type (WT) and dominant-negative (DN) PKC-alpha together with phorbol myristate acetate (PMA), which is a hypertrophic agonist and activator of all three PKC isoenzymes, we studied the role of PKC-alpha in signaling-specific aspects of the hypertrophic phenotype. PMA induced nuclear translocation of endogenous and AdV WT PKC-alpha in NRVMs. WT PKC-alpha overexpression increased protein synthesis and the protein-to-DNA (P/D) ratio but did not affect cell surface area (CSA) or cell shape compared with uninfected or control AdV beta-galactosidase (AdV betagal)-infected cells. PMA-treated uninfected cells displayed increased protein synthesis, P/D ratio, and CSA and elongated morphology. PMA did not further enhance protein synthesis or P/D ratio in AdV-WT PKC-alpha-infected cells. To assess the requirement of PKC-alpha for these PMA-induced changes, AdV-DN PKC alpha or AdV betagal-infected NRVMs were stimulated with PMA. Without PMA, AdV-DN PKC-alpha had no effects on protein synthesis, P/D ratio, CSA, or shape vs. AdV betagal-infected NRVMs. PMA increased protein synthesis, P/D ratio, and CSA in AdV betagal-infected cells, but these parameters were significantly reduced in PMA-stimulated AdV-DN PKC-alpha-infected NRVMs. Overexpression of DN PKC-alpha enhanced PMA-induced cell elongation. Neither WT PKC-alpha nor DN PKC-alpha affected atrial natriuretic factor gene expression. Insulin-like growth factor-1 also induced nuclear translocation of endogenous PKC-alpha. PMA but not WT PKC alpha overexpression induced ERK1/2 activation. However, AdV-DN PKC-alpha partially blocked PMA-induced ERK activation. Thus PKC-alpha is necessary for certain aspects of PMA-induced NRVM hypertrophy. PMID- 15271672 TI - Transgenic upregulation of IK1 in the mouse heart leads to multiple abnormalities of cardiac excitability. AB - To assess the functional significance of upregulation of the cardiac current (IK1), we have produced and characterized the first transgenic (TG) mouse model of IK1 upregulation. To increase IK1 density, a pore-forming subunit of the Kir2.1 (green fluorescent protein-tagged) channel was expressed in the heart under control of the alpha-myosin heavy chain promoter. Two lines of TG animals were established with a high level of TG expression in all major parts of the heart: line 1 mice were characterized by 14% heart hypertrophy and a normal life span; line 2 mice displayed an increased mortality rate, and in mice < or =1 mo old, heart weight-to-body weight ratio was increased by >100%. In adult ventricular myocytes expressing the Kir2.1-GFP subunit, IK1 conductance at the reversal potential was increased approximately 9- and approximately 10-fold in lines 1 and 2, respectively. Expression of the Kir2.1 transgene in line 2 ventricular myocytes was heterogeneous when assayed by single-cell analysis of GFP fluorescence. Surface ECG recordings in line 2 mice revealed numerous abnormalities of excitability, including slowed heart rate, premature ventricular contractions, atrioventricular block, and atrial fibrillation. Line 1 mice displayed a less severe phenotype. In both TG lines, action potential duration at 90% repolarization and monophasic action potential at 75-90% repolarization were significantly reduced, leading to neuronlike action potentials, and the slow phase of the T wave was abolished, leading to a short Q-T interval. This study provides a new TG model of IK1 upregulation, confirms the significant role of IK1 in cardiac excitability, and is consistent with adverse effects of IK1 upregulation on cardiac electrical activity. PMID- 15271674 TI - Emerging role of relaxin in renal and cardiovascular function. AB - Although traditionally associated with reproductive processes, relaxin is emerging as an important player in renal and cardiovascular function. Much of our recently acquired understanding of relaxin in this new context has arisen from studies of maternal renal and cardiovascular adaptations to pregnancy in rats where the hormone is turning out to be an important mediator. First, we highlight the influence of relaxin on renal hemodynamics and glomerular filtration rate, as well as on other peripheral circulations. Second, we discuss the effect of relaxin on both the steady and pulsatile systemic arterial load, as well as on the heart, in particular, coronary blood flow. Third, we consider the impact of the hormone on cultured endothelial and vascular smooth muscle cells. Fourth, we address the interaction of relaxin with renal and cardiac disease, as well as its role in angiogenesis. Finally, in Perspectives, we point out several key research questions in need of investigation that relate to a potential autocrine/paracrine role of relaxin in renal and cardiovascular tissues. Furthermore, on the basis of its potent vasodilatory and matrix-degrading attributes, we speculate about the therapeutic potential of relaxin in renal and cardiovascular diseases. PMID- 15271675 TI - Cardiac neuronal hierarchy in health and disease. AB - The cardiac neuronal hierarchy can be represented as a redundant control system made up of spatially distributed cell stations comprising afferent, efferent, and interconnecting neurons. Its peripheral and central neurons are in constant communication with one another such that, for the most part, it behaves as a stochastic control system. Neurons distributed throughout this hierarchy interconnect via specific linkages such that each neuronal cell station is involved in temporally dependent cardio-cardiac reflexes that control overlapping, spatially organized cardiac regions. Its function depends primarily, but not exclusively, on inputs arising from afferent neurons transducing the cardiovascular milieu to directly or indirectly (via interconnecting neurons) modify cardiac motor neurons coordinating regional cardiac behavior. As the function of the whole is greater than that of its individual parts, stable cardiac control occurs most of the time in the absence of direct cause and effect. During altered cardiac status, its redundancy normally represents a stabilizing feature. However, in the presence of regional myocardial ischemia, components within the intrinsic cardiac nervous system undergo pathological change. That, along with any consequent remodeling of the cardiac neuronal hierarchy, alters its spatially and temporally organized reflexes such that populations of neurons, acting in isolation, may destabilize efferent neuronal control of regional cardiac electrical and/or mechanical events. PMID- 15271676 TI - Preoptic thermoregulatory mechanisms in detail. PMID- 15271677 TI - Anorexia: the toll for lipopolysaccharide recognition. PMID- 15271678 TI - Control analysis, mitochondrial bioenergetics and programmed cell death: the Krogh principle in practice. PMID- 15271679 TI - Role for CD14, TLR2, and TLR4 in bacterial product-induced anorexia. AB - The cell surface component CD14 and the toll-like receptors 2 and 4 (TLR2 and TLR4) are important in mediating the immune responses to bacterial products in mammals. Using mice genetically deficient in CD14, TLR2, or TLR4, we studied the role of these molecules in the anorectic effects of LPS and muramyl dipeptide (MDP). CD14 or TLR2 knockout (KO) and TLR4-deficient (TLR4-DEF) mice as well as corresponding wild-type (WT) colittermates were injected intraperitoneally at dark onset with LPS (2 microg/mouse), MDP (10 mg/kg), interleukin-1 beta (IL-1 beta, 150 ng/mouse), or vehicle, and food intake was recorded. LPS and MDP reduced food intake in WT mice of all genotypes tested. The anorectic effect of LPS was attenuated (P < 0.04) in CD14-KO and TLR4-DEF mice but not in TLR2-KO (P > 0.05). The anorectic effect of MDP was blunted in CD14-KO and TLR2-KO (P < 0.02) mice but not in TLR4-DEF mice. IL-1 beta reduced food intake similarly in all genotypes tested. These results indicate that CD14 is involved in mediating the anorectic effects of both LPS and MDP. Furthermore, TLR4 and TLR2 are specifically involved in mediating the anorectic effects of LPS and MDP, respectively. The results are consistent with the hypothesis that TLR4 functions as the true LPS receptor and that TLR2 is involved in recognition of gram positive bacterial products. PMID- 15271680 TI - Acute and chronic effects of FR-149175, a beta 3-adrenergic receptor agonist, on energy expenditure in Zucker fatty rats. AB - Clinical therapies for both obesity and obese non-insulin-dependent diabetes mellitus require maintenance of reduced body weight after the initial successful reduction resulting from calorie control, exercise, or medication. Although beta(3)-adrenergic receptor (beta(3)-AR) agonists have been shown to stimulate whole body energy expenditure and lipid mobilization, whether stimulatory effects on oxygen consumption and lipolysis are influenced by chronic exposure to agonists has not been fully characterized. We therefore examined the acute and chronic effects of FR-149175, a selective beta(3)-AR agonist, on whole body oxygen consumption in genetically obese Zucker fatty rats. Chronic treatment with FR-149175 caused a decrease in both body weight gain and white fat pad weight at doses that induced lipolysis in acute treatment (1 and 3.2 mg/kg p.o.). Single administration of FR-149175 (0.1, 1, and 3.2 mg/kg p.o.) dose dependently increased whole body oxygen consumption. Repetitive administration did not cause attenuation of the thermogenic response at lower doses (0.1 and 1 mg/kg 2 times daily), whereas the highest dose (3.2 mg/kg 2 times daily) induced a progressive increase in oxygen consumption. PCR analyses of retroperitoneal white adipose tissue indicated little or no change in beta(3)-AR mRNA levels. Uncoupling protein 1 gene expression increased at 1 mg/kg, and drastic upregulation was detected at 3.2 mg/kg. FR-149175 also increased HSL mRNA levels in a dose-related manner, whereas there was no effect on genes involved in beta-oxidation. These results support that the thermogenic effect of beta(3)-AR agonists is not attenuated by chronic exposure to agonists. PMID- 15271681 TI - Differential cardiac parasympathetic innervation--what is the functional significance? PMID- 15271682 TI - Inhibition of salt appetite in rats by central oxytocin. PMID- 15271684 TI - New insights into the pathophysiology of the dysnatremias: a quantitative analysis. AB - Recent theoretical considerations have played an important role in advancing our understanding of the physiological mechanisms responsible for perturbing the plasma water sodium concentration ([Na(+)](pw)) in health and disease. Central to these considerations is the original empirical relationship between the [Na(+)](pw) and total exchangeable sodium (Na(e)), total exchangeable potassium (K(e)), and total body water (TBW) initially discovered by Edelman and colleagues (Edelman IS, Leibman J, O'Meara MP, and Birkenfeld LW. J Clin Invest 37: 1236 1256, 1958). The non-zero values of the slope and y-intercept in the Edelman equation are a consequence of the effects of the osmotic coefficient of Na(+) salts at physiological concentrations and Gibbs-Donnan and osmotic equilibrium. Moreover, in addition to Na(e), K(e), and TBW, the physiological components of the y-intercept in this equation play a role in modulating the [Na(+)](pw) and in the generation of the dysnatremias. In this review, the pathophysiological mechanisms underlying the generation and treatment of the dysnatremias are analyzed theoretically and quantitatively. Importantly, the non-zero values of both the slope and y-intercept in the Edelman equation result in several theoretical predictions that can be tested experimentally and have been mathematically incorporated into recently derived equations used to analyze both the generation and the optimal treatment of the dysnatremias. In addition, we review current concepts regarding 1) the role of Gibbs-Donnan and osmotic equilibrium in the determination of the [Na(+)](pw); 2) the modulating effect of osmotically inactive exchangeable Na(+) and K(+) on the [Na(+)](pw); 3) the effect of glucose on the [Na(+)](pw) as reflected by changes in Na(e), K(e), and TBW as well as changes in several components of the y-intercept resulting from the hyperglycemia; and 4) the complex role of K(+) in modulating the [Na(+)](pw). PMID- 15271685 TI - Cold ischemic injury of transplanted kidneys: new insights from experimental studies. AB - Kidney transplantation is the preferred and definitive treatment for end-stage renal disease (ESRD), and kidneys from deceased donors are a major source for it. These kidneys are routinely cold stored to prolong viability, which, however, when prolonged can cause injury, resulting in reduced graft function and survival. Recent experimental studies have identified the release of iron and free radicals, activation of calpain, and formation of F(2)-isoprostanes as important components of cold ischemic injury, as are the swelling of mitochondria and activation of mitochondrial apoptotic pathways. Moreover, studies have also suggested that fortifying the storage solution with deferoxamine or preconditioning the donor kidneys with hemeoxygenase-1 may prove viable clinical strategies to limit cold ischemic injury. This review will summarize these and other new experimental data that have implications for reducing cold ischemic transplant injury, a step necessary to improve deceased-donor allograft survival. PMID- 15271686 TI - Homocysteine clearance and methylation flux rates in health and end-stage renal disease: association with S-adenosylhomocysteine. AB - Hyperhomocysteinemia is a risk factor for cardiovascular disease and occurs frequently in end-stage renal disease (ESRD), but its pathogenesis is poorly understood. We aimed to evaluate one-carbon flux rates of methionine and homocysteine (Hcy) in ESRD patients and healthy controls. Transmethylation (TM), remethylation (RM), and transsulfuration (TS), as well as Hcy clearance by TS (i.e., TS/plasma total Hcy concentration) and by RM (i.e., RM/plasma total Hcy concentration) were evaluated in relation to body composition, vitamins, and S adenosylhomocysteine (AdoHcy) and S-adenosylmethionine (AdoMet) levels. After a fixed protein diet for 3 days, primed-continuous infusion of [(2)H(3)-methyl-1 (13)C]methionine was performed in the postabsorptive state in 12 hemodialysis patients and 16 healthy volunteers. Hcy clearance by TS (-80%, P < 0.001) and by RM (-77%, P < 0.001) in ESRD patients was decreased compared with healthy controls. The absolute flux rates of TM (-27%, P < 0.01) and RM (-28%, P = 0.02) were lower in the ESRD patients. After adjustment for age, TS was not significantly reduced. Whole blood AdoHcy was significantly elevated in ESRD and was a significant determinant of TM (standardized beta = -1.24, P = 0.01) and RM (standardized beta = -1.43, P = 0.03). In conclusion, patients with ESRD have impaired Hcy clearance by TS and RM. Elevated whole blood AdoHcy levels are associated with impaired RM and TM flux rates in these patients, and AdoHcy may be a key regulatory compound in one-carbon flux. PMID- 15271687 TI - Tetanization-induced pelvic-to-pudendal reflex plasticity in anesthetized rats. AB - Reflex plasticity between pelvic afferent and pudendal efferent nerve fibers was examined in anesthetized rats. Brief high-frequency electric stimulation (300 pulses at 100 Hz) of the pelvic nerve afferent fiber produced a long-lasting potentiation of the pelvic-to-pudendal reflex (PPR). This tetanization-induced potentiation was abolished by a selective N-methyl-D-aspartate (NMDA) receptor antagonist and attenuated by a non-NMDA excitatory amino acid receptor antagonist. However, the GABA(A)-receptor antagonist had no effect on this potentiation. Both intrathecal glutamate (0.1 mM, 2-5 microl it) and NMDA (0.1 mM, 2-5 microl it) induced a potentiation of PPR similar to that of tetanization. Agonist-induced potentiation was shorter than tetanization-induced potentiation. The duration of the contraction wave of intraurethral pressure, elicited by PPR, was elongated by tetanization-induced potentiation, whereas the peak pressure was not affected. All these results demonstrate that brief high-frequency stimulation of the pelvic nerve afferent fiber can induce a distinct and long-lasting modulation in PPR activity and this change may be involved in nociceptive C afferent-induced obstructive urinary dysfunctions. PMID- 15271688 TI - High- and low-affinity transport of L-leucine and L-DOPA by the hetero amino acid exchangers LAT1 and LAT2 in LLC-PK1 renal cells. AB - The present study examined the functional characteristics of the inward and outward L-[14C]DOPA and L-[14C]leucine transporters in LLC-PK1 cells. Uptake was initiated by the addition of Hanks' medium with a given concentration of L [14C]DOPA or L-[14C]leucine. Saturation experiments were performed in cells incubated for 6 min with 0.25 microM concentration of the substrates in the absence and the presence of increasing concentrations of the nonlabeled substrates. Fractional outflow of intracellular L-[14C]DOPA or L-[14C]leucine was evaluated in cells loaded with 2.5 microM L-[14C]DOPA or 1 microM L-[14C]leucine for 6 min and then the corresponding efflux was monitored over 24 min. The high affinity (Km = 5.1 microM) uptake of L-[14C]leucine and the low-affinity (Km = 120.0 microM) uptake of L-[14C]DOPA were largely promoted through a Na+ independent transporter. The uptake of the substrates was insensitive to N (methylamino)-isobutyric acid but competitively inhibited by 2 aminobicyclo(2,2,1)-heptane-2-carboxylic acid (BCH). L- And D-neutral amino acids, but not acidic and basic amino acids, markedly inhibited L-[14C]DOPA and L [14C]leucine accumulation. The uptake of L-[14C]leucine was a pH-insensitive process, whereas that of L-[14C]DOPA was sensitive to pH. The efflux of L [14C]DOPA and L-[14C]leucine was markedly increased (P < 0.05) by L-cysteine, L leucine, BCH, and L-DOPA but not by L-arginine. RT-PCR detected LAT1 and LAT2 transcripts in LLC-PK1 cells. It is concluded that LLC-PK1 cells express both LAT1 and LAT2 transcripts and transport L-[14C]leucine through the Na+ independent pH-insensitive and high-affinity LAT1 transporter, whereas L [14C]DOPA is mainly transported through the Na+-independent pH-insensitive and low-affinity LAT2 transporter and a minor component through a Na+-dependent transporter. PMID- 15271689 TI - Alpha1-antitrypsin as a risk for infant and adult respiratory outcomes in a national birth cohort. AB - Reduced alpha1-antitrypsin (AAT) encoded by the gene SERPINA1 is a potential risk for pulmonary disease. We investigated SERPINA1 polymorphism as a risk for infant and adult pulmonary morbidity, and adult respiratory function and its change between 43 and 53 yr. We used data on a British national representative sample (n = 5,362) studied since birth in 1946 to age 53 yr (when n = 3,035), when DNA was first obtained. SERPINA1 Z and, to a lesser extent, S carriers had an increased risk of infant lower respiratory infection compared with those who were neither S nor Z carriers (Z carriers: odds ratio = 2.32, 95% confidence interval = 1.37 3.92; S but not Z carriers odds ratio = 1.58, 95% confidence interval = 1.10 2.28) after adjustment for environmental, socioeconomic, and developmental factors, and breast-feeding. There was no difference in the adult outcomes at 53 yr according to genotype, nor was there any association of genotype with change in forced expiratory volume at 1 s between 43 and 53 yr. Lower alpha1 antitrypsin, as indicated by carrier status for the Z and S alleles, was a risk for infant lower respiratory infection, but not for adult respiratory outcomes. PMID- 15271690 TI - Effects of testosterone and resistance training in men with chronic obstructive pulmonary disease. AB - Dysfunction of the muscles of ambulation contributes to exercise intolerance in chronic obstructive pulmonary disease (COPD). Men with COPD have high prevalence of low testosterone levels, which may contribute to muscle weakness. We determined effects of testosterone supplementation (100 mg of testosterone enanthate injected weekly) with or without resistance training (45 minutes three times weekly) on body composition and muscle function in 47 men with COPD (mean FEV(1) = 40% predicted) and low testosterone levels (mean = 320 ng/dl). Subjects were randomized to 10 weeks of placebo injections + no training, testosterone injections + no training, placebo injections + resistance training, or testosterone injections + resistance training. Testosterone injections yielded a mean increase of 271 ng/dl in the nadir serum testosterone concentration (to the middle of the normal range for young men). The lean body mass (by dual-energy X ray absorptiometry) increase averaged 2.3 kg with testosterone alone and 3.3 kg with combined testosterone and resistance training (p < 0.001). Increase in one repetition maximum leg press strength averaged 17.2% with testosterone alone, 17.4% with resistance training alone, and 26.8% with testosterone + resistance training (p < 0.001). Interventions were well tolerated with no abnormalities in safety measures. Further studies are required to determine long-term benefits of adding testosterone supplementation and resistance training to rehabilitative programs for carefully screened men with COPD and low testosterone levels. PMID- 15271691 TI - Critical care use during the course of serious illness. AB - Despite its expense and importance, it is unknown how common critical care use is. We describe longitudinal patterns of critical care use among a nationally representative cohort of elderly patients monitored from the onset of common serious illnesses. A retrospective population-based cohort study of elderly patients in fee-for-service Medicare is used, with 1,108,060 Medicare beneficiaries at least 68 years of age and newly diagnosed with serious illnesses: 1 of 9 malignancies, stroke, congestive heart failure, hip fracture, or myocardial infarction. Medicare inpatient hospital claims from diagnosis until death (65.1%) or fixed-right censoring (more than 4 years) were reviewed. Distinct hospitalizations involving critical care use (intensive care unit or critical care unit) were counted and associated reimbursements were assessed; repeated use was defined as five or more such hospitalizations. Of the cohort, 54.9% used critical care at some time after diagnosis. Older patients were much less likely to ever use critical care (odds ratio, 0.31; comparing patients more than 90 years old with those 68-70 years old), even after adjustment. A total of 31,348 patients (2.8%) were repeated users of critical care; they accounted for 3.6 billion dollars in hospital charges and 1.4 billion dollars in Medicare reimbursement. We conclude that critical care use is common in serious chronic illness and is not associated solely with preterminal hospitalizations. Use is uneven, and a minority of patients who repeatedly use critical care account for disproportionate costs. PMID- 15271692 TI - Influence of calibration on densitometric studies of emphysema progression using computed tomography. AB - The fundamental importance of calibration for any measuring device is indisputable, but computed tomography (CT) calibration in longitudinal lung densitometry studies is largely unexplored. Although the validity of CT as a measure of emphysema has been confirmed in cross-sectional studies, there are limited data on long-term reproducibility, and this is critically important for validating its use as an outcome measure in therapeutic trials. A general understanding of the strengths and pitfalls of CT densitometry is critical for physicians reviewing the published literature using this methodology. In our study of 57 patients with alpha-1 antitrypsin deficiency (phenotype PiZ), progression of voxel index determined from three successive annual scans acquired with a fully calibrated scanner was intimately associated with changes in CT air densitometry, sampled from patient images. Images were therefore reanalyzed, using a correction technique validated in phantom studies that adjusted for changes in measured air density, and the reliability of the voxel index as a measure of emphysema progression was improved. Comparison of adjusted voxel index thresholds indicated the optimum threshold was -950 Hounsfield units. Internal air calibration is therefore critical in longitudinal and multicenter lung densitometry studies of emphysema and incorporation of a correction factor is essential for quantitative image analysis. PMID- 15271693 TI - An effective strategy for diagnosing occupational asthma: use of induced sputum. AB - Monitoring airway inflammation by means of induced sputum cell counts seems to improve the management of asthma. We sought to assess whether such monitoring at the end of periods at and away from work combined with the monitoring of PEF could improve the diagnosis of occupational asthma. We enrolled subjects suspected of having occupational asthma. Serial monitoring of PEF was performed during 2 weeks at and away from work. At the end of each period, induced sputum was collected. Specific inhalation challenge was subsequently performed. PEF graphs were interpreted visually by five independent observers. Forty-nine subjects, including 23 with positive specific inhalation challenge, completed the study. The addition of sputum cell counts to the monitoring of PEF increased the specificity of this test, respectively, by 18 (range [r] 13.7-25.5) or 26.8% (r 24.8-30.4) depending if an increase of sputum eosinophils greater than 1 or 2% when at work was considered as significant. The sensitivity increased by 8.2% (r 4.1-13.4) or decreased by 12.3% (r 3.1-24.1) depending on the cutoff value in sputum eosinophils chosen (greater than 1 or 2%, respectively). The addition of sputum cell counts to PEF monitoring is useful to improve the diagnosis of occupational asthma. PMID- 15271694 TI - Pulmonary surfactant, lung function, and endobronchial inflammation in cystic fibrosis. AB - Cystic fibrosis (CF) lung disease is primarily a disease of the small airways. We hypothesized that even in patients with normal lung function, a reduced surfactant function would be present and favor small airway obstruction. Bronchoalveolar lavages from 76 patients with CF (5-31 years, median 11) with well-conserved lung function (FEV1 94% predicted, range 78-121) and from 10 healthy control subjects were investigated. The deviation of the biophysical surfactant performance from normal, assessed in a bubble surfactometer, was small; however, the ability of the surfactant to maintain the patency of a narrow airway (% open) was significantly reduced. Surfactant protein (SP)-C level was increased, SP-B and SP-D were unchanged, whereas SP-A was decreased. Among the patients with CF, neutrophilic inflammation was modestly related to a poorer surfactant activity, but not to lung function. SP-D was reduced in proportion to the degree of inflammation and in the presence of bacteria. These findings in a large cohort of patients with CF with normal lung function show that the endobronchial airway inflammation is linked to early perturbations of the biophysical properties and immunologic components of pulmonary surfactant and opens fields for novel therapeutic interventions. PMID- 15271695 TI - Influence of ambient and ventilator output temperatures on performance of heated wire humidifiers. AB - Although heated humidifiers are considered the most efficient humidification devices for mechanical ventilation, endotracheal tube occlusion caused by dry secretions has been reported with heated-wire humidifiers. We tested the hypothesis that inlet chamber temperature, influenced by ambient air and ventilator output temperatures, may affect humidifier performance, as assessed by hygrometry. Hygrometry was measured with three different humidifiers under several conditions, varying ambient air temperatures (high, 28-30 degrees C; and normal, 22-24 degrees C), ventilators with different gas temperatures, and two VE levels. Clinical measurements were performed to confirm bench measurements. Humidifier performance was strongly correlated with inlet chamber temperature in both the bench (p < 0.0001, r2 = 0.93) and the clinical study. With unfavorable conditions, absolute humidity of inspired gas was much lower than recommended (approximately 20 mg H2O/L). Performance was improved by specific settings or new compensatory algorithms. Hygrometry could be evaluated from condensation on the wall chamber only when ambient air temperature was normal but not with high air temperature. An increase in inlet chamber temperature induced by high ambient temperature markedly reduces the performance of heated-wire humidifiers, leading to a risk of endotracheal tube occlusion. Such systems should be avoided in these conditions unless automatic compensation algorithms are used. PMID- 15271696 TI - Elevated plasma ghrelin level in underweight patients with chronic obstructive pulmonary disease. AB - Ghrelin, a novel growth hormone-releasing peptide, has been shown to cause a positive energy balance by reducing fat use and stimulating food intake. This study investigated whether plasma ghrelin is associated with clinical parameters in patients with chronic obstructive pulmonary disease. Plasma ghrelin was measured in 50 patients and 13 control subjects, together with anabolic and catabolic factors. Patients were divided into two groups based on body mass index: underweight patients (n = 26) or normal weight patients (n = 24). Plasma ghrelin was significantly higher in underweight patients than in normal weight patients and healthy control subjects. Circulating tumor necrosis factor-alpha, interleukin-6, and norepinephrine were significantly higher in underweight patients than in normal weight patients. Plasma ghrelin correlated negatively with body mass index and correlated positively with catabolic factors such as tumor necrosis factor-alpha and norepinephrine. In addition, plasma ghrelin correlated positively with percent predicted residual volume and residual volume to-total lung capacity ratio. In conclusion, plasma ghrelin was elevated in underweight patients with chronic obstructive pulmonary disease, and the level was associated with a cachectic state and abnormality of pulmonary function. PMID- 15271697 TI - The new anesthesia diet plan: keeping perioperative carbs in check. PMID- 15271698 TI - Maintenance of normoglycemia during cardiac surgery. AB - We used the hyperinsulinemic normoglycemic clamp technique, i.e., infusion of insulin at a constant rate combined with dextrose titrated to clamp blood glucose at a specific level, to preserve normoglycemia during elective cardiac surgery. Ten nondiabetic and seven diabetic patients entered the clamp protocols. Perioperative glucose control was also assessed in 19 nondiabetic and 11 diabetic patients (control group) receiving a conventional insulin infusion sliding scale. In patients of the clamp group, a priming bolus of insulin (2 U) was started before the induction of anesthesia followed by infusions of insulin at 5 mU. kg( 1). min(-1) and of variable amounts of dextrose. Arterial blood glucose was measured every 5 min in the clamp group and every 20 min in the control group. Control of normoglycemia was defined as > or =95% of the glucose levels within 4.0-6.0 mmol/L. Glucose concentration was recorded before surgery, 15 min before cardiopulmonary bypass (CPB), during early and late CPB, and at sternal closure. Patients of the control group became progressively hyperglycemic during surgery (late CPB; nondiabetics, 9.0 +/- 3.2 mmol/L; diabetics, 10.1 +/- 3.6 mmol/L), whereas normoglycemia was achieved in the study group (late CPB; nondiabetics, 5.5 +/- 0.7 mmol/L; diabetics, 4.9 +/- 0.6 mmol/L; P < 0.05 versus control group). In conclusion, it seems that normal blood glucose concentration during open heart surgery can be reliably maintained in nondiabetic and diabetic patients by using the hyperinsulinemic normoglycemic clamp technique. PMID- 15271699 TI - The effect of diabetes on the interrelationship between jugular venous oxygen saturation responsiveness to phenylephrine infusion and cerebrovascular carbon dioxide reactivity. AB - In this study, we examined whether cerebrovascular carbon dioxide (CO(2)) reactivity was related to the response of jugular venous oxygen saturation (SjvO(2)) to phenylephrine infusion in diabetic patients during cardiopulmonary bypass. Forty diabetic patients scheduled for coronary artery bypass graft surgery were studied, and 40 age-matched nondiabetic cardiopulmonary bypass patients served as controls. Cerebrovascular CO(2) reactivity was measured continuously using transcranial Doppler. Mean arterial blood pressure (MAP) was increased by repeated phenylephrine infusion until reaching 100% of baseline values. There was a significant difference in absolute CO(2) reactivity between the diabetic and control groups (controls, 2.8 +/- 0.7 cm. s(-1). mm Hg(-1); diabetics, 2.2 +/- 1.1 cm. s(-1). mm Hg(-1); P = 0.02). Among the diabetics, absolute CO(2) reactivity in insulin-dependent patients was less than that in noninsulin-dependent patients (diet therapy group, 3.2 +/- 0.7; glibenclamide group, 2.6 +/- 0.7; insulin-dependent group, 1.0 +/- 0.7; P < 0.01). There was a correlation between absolute CO(2) reactivity and the mean slope of SjvO(2) versus MAP for increasing MAP (r = 0.54; P < 0.0001). In conclusion, we found that the interrelationship between SjvO(2) responsiveness to phenylephrine infusion and cerebrovascular CO(2) reactivity, as well as impaired cerebrovascular autoregulation, were associated with previous hyperglycemia. PMID- 15271700 TI - The effects of load on systolic mitral annular velocity by tissue Doppler imaging. AB - Tissue Doppler Imaging (TDI) provides information on systolic function through its systolic mitral annulus velocity wave (Sm), reflecting the peak velocity of shortening of the myocardial fibers oriented in the longitudinal direction. In this study, we evaluated the effect of load changes on Sm. Forty-two cardiac surgical patients with left ventricular ejection fraction >60% were consecutively evaluated. In 24 patients, load was changed with an IV bolus of phenylephrine (50 100 microg) or nitroglycerine (300-500 microg); in 18 patients, preload was changed with a rapid infusion of 500 mL of a gelatin solution. The sample volume of TDI was placed at the lateral side of the mitral annulus in the mid-esophageal 4-chamber view. Changing loading conditions with phenylephrine or nitroglycerine had no effect on Sm; the increase of preload in 18 patients resulted in a statistically significant increase of Sm (baseline, 8.4 +/- 2.6 cm/s; after increase of preload, 9.6 +/- 2.5 cm/s; P = 0.001). We conclude that Sm is dependent on changes in preload obtained by volume loading and cannot be recommended as an index of ventricular contractile performance in critically ill patients where significant changes in ventricular filling occur. PMID- 15271701 TI - Midazolam: an effective antiemetic after cardiac surgery--a clinical trial. AB - Cardiac surgery has been associated with a significant incidence of postoperative nausea and vomiting (PONV). To assess the antiemetic property of midazolam, we undertook this double-blinded, randomized trial in 200 patients undergoing cardiac surgery involving cardiopulmonary bypass, and we compared its efficacy with that of ondansetron in preventing PONV. Assessments on the occurrence of PONV were made at regular intervals for the first 24 h after tracheal extubation, along with sedation and pain scoring. We report a 6% incidence of nausea and no incidence of vomiting in the midazolam group, compared with a 21% incidence of PONV in the ondansetron group (P < 0.001). All 21 patients (18 women and 3 men) in the ondansetron group and none of the 6 patients (all women) in the midazolam group required a rescue antiemetic drug (P < 0.001). The sedation scores and postoperative pain scores were comparable in both groups. We conclude that midazolam, instituted as a continuous infusion in a dose of 0.02 mg. kg(-1). h( 1), is a more effective antiemetic than ondansetron in a dose of 0.1 mg/kg IV every 6 h for the prevention of PONV after cardiac surgery. PMID- 15271702 TI - Regional cerebral oxygen saturation is a sensitive marker of cerebral hypoperfusion during orthotopic liver transplantation. AB - Neurological complications contribute significantly to morbidity and mortality of patients after orthotopic liver transplantation (OLT). One possible cause of postoperative neurological complications is cerebral ischemia during the surgical procedure. In this study, we investigated the relationship between intraoperative changes in regional cerebral oxygen saturation (rSo(2)) and postoperative values of neuron-specific enolase (NSE) and S-100, which are specific variables that indicate cerebral disturbances due to hypoxia/ischemia. The rSo(2) was monitored continuously by near-infrared spectroscopy in 16 patients undergoing OLT. In addition, NSE and S-100 were determined in arterial blood before surgery and 24 h after reperfusion of the donor liver. Interestingly, clamping of the recipient's liver led to a significant decline in rSo(2) in eight patients, whereas the others tolerated clamping without major changes in rSo(2). The decrease in rSo(2) after clamping correlated significantly with postoperative increases in NSE (r(2) = 0.57) and S-100 (r(2) = 0.52). However, there were no significant differences between patients with and without rSo(2) decline concerning hemodynamic variables. There were no significant correlations between DeltarSo(2) and cardiac output (r(2) = 0.20), NSE and cardiac output (r(2) = 0.37), or S-100 and cardiac output (r(2) = 0.24). Monitoring of rSo(2) may be a useful noninvasive tool to estimate disturbances in rSo(2) during OLT. PMID- 15271703 TI - Postoperative death in a patient with unrecognized arrhythmogenic right ventricular dysplasia syndrome. AB - Arrhythmogenic right ventricular dysplasia is an inherited disease causing fatty replacement of heart tissue. This disease often presents as T-wave inversion in the anterior leads of the electrocardiogram (ECG) with life-threatening ventricular arrhythmias. In older patients, progressive right and left ventricular failure can develop. This is a case report of postoperative death occurring in a 59-yr-old woman with undiagnosed arrhythmogenic right ventricular dysplasia after hepatic cystectomy. The patient had T-wave inversion in the inferior ECG leads and no history of arrhythmias. During general anesthesia, cardiovascular collapse occurred in the absence of arrhythmias that was unresponsive to resuscitation. PMID- 15271704 TI - Diagnosis and treatment of intracardiac thrombosis during orthotopic liver transplantation. AB - Intracardiac thrombus formation during orthotopic liver transplantation can be a catastrophic event leading to death. Most often this devastating complication occurs after reperfusion and may be related to massive blood transfusion, marginal liver grafts, tendencies towards hypercoagulability, or the potential role of antifibrinolytics. We report a case of an intracardiac thrombus occurring during the hepatectomy stage (stage I) of orthotopic liver transplantation. Transesophageal echocardiography was used to quickly diagnose the thrombus, allowing rapid pharmacological intervention and later guide surgical evacuation of the intracardiac thrombus via the inferior vena cava. PMID- 15271705 TI - The role of transesophageal echocardiography in rapid diagnosis and treatment of migratory tumor embolus. AB - Transesophageal echocardiography (TEE) is sometimes used in renal cell carcinoma excision for evaluating the extension of tumor in the inferior vena cava (IVC), characterizing the tumor anatomy, monitoring the tumor during surgical mobilization, and assessing cardiac function. Although the risk for embolization is small, when embolization does occur, its consequences can be catastrophic. In this case report, we describe the crucial role of TEE in diagnosing an intraoperative migratory embolus from the IVC to the pulmonary artery and also provide both single-frame photographs and Internet-accessible videos of the event. Our case illustrates the key role that TEE played in the intraoperative management of a patient with renal cell carcinoma undergoing surgical excision of tumor. TEE aided in accurately defining the cephalad extent of the thrombus, provided continuous monitoring of the thrombus during surgical manipulation, and allowed immediate identification of its embolization and proper notification of the surgeons. This case illustrates the crucial role TEE played in the management of a migratory tumor embolus and argues for its routine use during excision of renal cell carcinomas invading the IVC. PMID- 15271706 TI - Does halothane really preserve cardiac baroreflex better than sevoflurane? A noninvasive study of spontaneous baroreflex in children anesthetized with sevoflurane versus halothane. AB - Heart rate profiles during the induction of anesthesia differ markedly between the administration of sevoflurane and halothane. Previous investigations have shown that halothane preserves cardiac parasympathetic activity more than sevoflurane. Because vagal drive to the sinus node is the main effector of arterial baroreflex control of heart rate, halothane may preserve cardiac baroreflex better than sevoflurane. To investigate cardiac baroreflex in anesthetized children, we used two noninvasive methods providing different approaches to the arterial blood pressure (BP) and R-R interval (RRI) relationship: the sequence methods investigating beat-to-beat changes in BP and RRI (time domain) and the cross-spectral analysis investigating relationships between oscillations of BP and RRI (frequency domain). Children were randomly assigned to mask induction with sevoflurane in 100% oxygen, sevoflurane in 50% nitrous oxide/50% oxygen, or halothane in 50% nitrous oxide/50% oxygen. After tracheal intubation, the inspired fraction of volatile anesthetic was reduced to 1 minimum alveolar anesthetic concentration (MAC). The spontaneous baroreflex (SBR) sensitivity was calculated with the sequence method at baseline, during induction, and after intubation. The cardiac baroreflex was also estimated with cross-spectral analysis at baseline and at 1 MAC (stationary conditions). In the three groups, the induction of anesthesia was associated with a marked decrease of SBR sensitivity, which occurred earlier with sevoflurane than with halothane. Five minutes after intubation (1 MAC), the sequence method showed a similar decrease of the SBR sensitivity in the three groups. Similarly, the cross spectral analysis between systolic blood pressure and RRI showed a decrease of the gain calculated in the low-frequency band, but the gain in the respiratory band was higher with halothane compared with sevoflurane. In children, the induction of anesthesia with halothane and sevoflurane is associated with a marked decrease of cardiac baroreflex activity. The persistence of respiratory RRI fluctuations under halothane might reflect reflex respiratory arrhythmia rather than efficient parasympathetic baroreflex activity. PMID- 15271707 TI - Hepatitis after sevoflurane exposure in an infant suffering from primary hyperoxaluria type 1. AB - An 11-mo-old child with primary hyperoxaluria was scheduled for a nephroureteromia procedure. Anesthesia was induced and maintained with sevoflurane. Two days after the operation, a hepatomegaly was diagnosed, and a considerable increase in liver enzymes was observed. These pathologic findings disappeared without treatment within 7 days. In a subsequent operation 2 wk later, general anesthesia was performed (sevoflurane was avoided). After the second operation, no pathologic findings could be detected. Nothing in this patient's disease or the conduct of the anesthesia suggested a cause for the injury other than an idiosyncratic response to sevoflurane. PMID- 15271708 TI - The use of caudal morphine for pediatric liver transplantation. AB - A 3-yr-old female with cryptogenic cirrhosis presented for a liver transplant. After the induction and intubation, we performed a supplemental caudal block with a 22-gauge B-bevel needle in the usual sterile fashion, and 0.6 mg of Duramorph was injected without complications. Initially, the 14.9-kg child received a total of 110 microg of fentanyl in the first 2 h of the 6-h operation and was maintained on air-oxygen-isoflurane. The child was easily tracheally extubated and remained hemodynamically stable. In the pediatric intensive care unit, she was weaned off oxygen, out of bed, and required minimal pain control in the first 18 h. PMID- 15271709 TI - The analgesic effects of gabapentin in monitored anesthesia care for ear-nose throat surgery. AB - We investigated the efficacy and safety of gabapentin in rhinoplasty or endoscopic sinus surgery patients. Patients received either oral placebo or gabapentin 1200 mg 1 h before surgery. After standard premedication, 25 patients in each group received propofol, fentanyl, and local anesthesia at the operative site. Sedation was maintained by a continuous infusion of propofol adjusted according to the Ramsay scale. Sedation and pain scores were obtained at 5, 15, 30, 45, and 60 min during surgery and 30 min and 2, 4, 6, 8, 12, 16, 20, and 24 h after the procedure. Diclofenac 75 mg IM was administered as a rescue analgesic. Postoperative pain scores and intraoperative pain scores at 45 and 60 min were significantly lower in the gabapentin group. Fentanyl (122 +/- 40 microg versus 148 +/- 42 microg; P < 0.05) and diclofenac (33 +/- 53 mg versus 111 +/- 92 mg; P < 0.001) consumption was smaller and the time to first analgesic request (18 +/- 9 h versus 9 +/- 7 h; P < 0.001) was longer in the gabapentin group. A more frequent incidence of dizziness was found in the gabapentin (versus placebo) group (24% versus 4%, respectively). We conclude that gabapentin provided a significant analgesic benefit for intraoperative and postoperative pain relief in patients undergoing ambulatory rhinoplasty or endoscopic sinus surgery; however, dizziness may be a handicap for ambulatory use. PMID- 15271710 TI - Capnography accurately detects apnea during monitored anesthesia care. AB - Apnea and airway obstruction are common during monitored anesthesia care (MAC). Because their early detection is essential, we sought to measure the efficacy of capnography as an indicator of apnea during MAC at a variety of oxygen flow rates compared with thoracic impedance. Anesthesia care providers using standard American Society of Anesthesiologists monitors were blinded to capnography and thoracic impedance monitoring. Ten (26%) of the 39 patients studied developed 20 s of apnea; none was detected by the anesthesia provider, but all were detected by capnography and impedance monitoring. There was no difference in detection rates between the two methods. Higher oxygen flow rates decreased the amplitude of the capnograph but did not interfere with apnea detection. This pilot study revealed that apnea of at least 20 s in duration may occur in every fourth patient undergoing MAC. Although these episodes were undetected by the anesthesia provider, they were reliably detected by both capnography and respiratory plethysmography. Monitoring of nasal end-tidal CO(2) is an important way to improve safety in patients undergoing MAC. PMID- 15271711 TI - Unsuspected temporomandibular joint pathology leading to a difficult endotracheal intubation. AB - A 40-yr-old woman with an unremarkable medical history and no prior surgeries presented for ambulatory surgery. Physical examination revealed normal jaw opening. On induction of general anesthesia, her jaw was found to be locked in a nearly closed position. We discuss anesthetic considerations and the pathology of temporomandibular joint anterior disk dislocation without reduction. A simple maneuver to reduce the dislocation is described. PMID- 15271712 TI - Neuromuscular pharmacodynamics of rocuronium in patients with major burns. AB - Rocuronium, which has a short onset time and is free of hyperkalemic effects, could be considered for rapid-sequence induction of anesthesia in patients with burns. In this study, we assessed the neuromuscular pharmacodynamics of rocuronium in patients with major burns. Adults aged 18-59 yr who had a major burn injury (n = 56) and a control group of 44 nonburned patients were included. Rocuronium was used at 3 times (0.9 mg/kg) or 4 times (1.2 mg/kg) the 95% effective dose. Anesthesia consisted of propofol and fentanyl with nitrous oxide and oxygen. Neuromuscular block was monitored with an acceleromyograph by using train-of-four stimulation. The onset time to 95% neuromuscular block was prolonged in burned compared with nonburned patients (115 +/- 58 s versus 68 +/- 16 s for 0.9 mg/kg; 86 +/- 20 s versus 57 +/- 11 s for 1.2 mg/kg). Dose escalation shortened the onset time, prolonged the duration of action, and improved intubating conditions in burned patients. All recovery profiles were significantly shorter in burned versus nonburned groups with both doses. Resistance to the neuromuscular effects of rocuronium was partially overcome by increasing the dose. A dose up to 1.2 mg/kg provides good tracheal intubating conditions after major burns. PMID- 15271713 TI - Propofol inhibits human platelet aggregation induced by proinflammatory lipid mediators. AB - Lysophosphatidic acid (LPA), platelet-activating factor (PAF), and thromboxane A(2) are proinflammatory lipid mediators that activate surface receptors on platelets, producing increased intracellular calcium, which is necessary for aggregation. We investigated propofol's effect on platelet aggregation and intracellular calcium mobilization caused by these three agonists. Platelets from human volunteers were incubated in buffers containing LPA (1 microM), U46619 (thromboxane A(2) analog; 1 microM), or PAF (10 nM). Propofol emulsion or 2,6 diisopropylphenol (propofol without fat emulsion) dissolved in ethanol was added to achieve concentrations of propofol used clinically: 5 or 10 microg/mL. After 2 min, aggregation or intracellular calcium concentrations were measured with optical techniques. Propofol emulsion and propofol in ethanol produced similar inhibition of platelet aggregation induced by LPA, PAF, and U46619 in a dose dependent fashion. LPA, PAF, and U46619 each caused significant increases in intracellular calcium that were not modified by propofol. Because propofol does not significantly alter intracellular calcium increases caused by receptor activation, inhibition appears to act distal to platelet receptors, inositol phosphate 3, and phospholipase C. Because the three lipid mediators play a key role in inflammation, their inhibition by propofol might be clinically important. PMID- 15271714 TI - Thiopental and propofol affect different regions of the brain at similar pharmacologic effects. AB - Propofol has a greater amnesic effect than thiopental. In this study we tested whether different brain regions were affected by propofol and thiopental at similar drug effects. Changes in regional cerebral blood flow (rCBF) were identified by using SPM99 analysis of images obtained with positron emission tomography with (15)O water. Ten right-handed male volunteers (age, 35 +/- 10 yr; weight, 74.1 +/- 7.5 kg; mean +/- sd) were randomized to receive thiopental (n = 4) or propofol (n = 6) to target sedative and hypnotic concentrations with bispectral index (BIS) monitoring. Four positron emission tomography images were obtained during various tasks at baseline and with sedative and hypnotic effects. Two participants receiving propofol were unresponsive at sedative concentrations and were not included in the final analyses. Median serum concentrations were 1.2 and 2.7 microg/mL for sedative and hypnotic propofol effects, respectively. Similarly, thiopental concentrations were 4.8 and 10.6 microg/mL. BIS decreased similarly in both groups. The pattern of rCBF change was markedly different for propofol and thiopental. Propofol decreased rCBF in the anterior (right-sided during sedation) brain regions, whereas thiopental decreased rCBF primarily in the cerebellar and posterior brain regions. At similar levels of drug effect, propofol and thiopental affect different regions of the brain. These differences may help to identify the loci of action for the nonsedative effects of propofol, such as amnesia. PMID- 15271715 TI - Halothane and propofol modulation of gamma-aminobutyric acidA receptor single channel currents. AB - Halothane and propofol enhance the activity of the gamma-aminobutyric acid (GABA) system, which is one of the most important systems in the mechanism of anesthesia. To determine whether halothane and propofol enhance GABAergic responses by the same mechanism, we performed single-channel patch-clamp experiments with rat cortical neurons in primary culture. Each of the open-time and closed-time distributions of GABA(A) receptor single channels was expressed by a sum of fast and slow time constants. Neither halothane nor propofol changed the single-channel conductance. Halothane increased the probability of the channel being open via a prolongation of the slow phase of open time, whereas propofol increased the channel open probability via a shortening of the slow phase of closed time. Thus, although both halothane and propofol augmented the channel open probability, thereby causing an increase in charge transfer during inhibitory transmitter action, they acted by different mechanisms. PMID- 15271716 TI - Modulation of Xenopus laevis Ca-activated Cl currents by protein kinase C and protein phosphatases: implications for studies of anesthetic mechanisms. AB - Ca-activated Cl currents (I(Cl(Ca))) are used frequently as reporters in functional studies of anesthetic effects on G protein-coupled receptors using Xenopus laevis oocytes. However, because anesthetics affect protein kinase C (PKC), they could indirectly affect I(Cl(Ca)) if this current is regulated by phosphorylation. We therefore studied the effect of modulation of either PKC or protein phosphatases PP1alpha and PP2A on I(Cl(Ca)) stimulated either by lysophosphatidate (LPA) signaling or by microinjection of Ca. X. laevis oocytes were studied under voltage clamp. Rat PP1alpha and PP2A were overexpressed in oocytes. PP, inositoltrisphosphate (IP(3)), the PP inhibitor okadaic acid (OA), the PKC inhibitor chelerythrine, or CaCl(2) were directly injected into the oocyte. Responses to agonists (LPA 10(-6) M, IP(3) 10(-4) M, CaCl(2) 0.5 M) were measured at a holding potential of -70 mV in the presence or absence of the PP inhibitors cantharidin or OA. PP1 alpha and PP2A inhibited I(Cl(Ca)) from 7.6 +/- 0.9 microC to 2.5 +/- 0.9 microC and 3.2 +/- 1.4 microC, respectively. PP inhibition enhanced I(Cl(Ca)) in control oocytes and reversed the inhibitory effect in oocytes expressing PP1 alpha or PP2A. PKC inhibition by chelerythrine enhanced both LPA- and CaCl(2)-induced I(Cl(Ca)). Our data indicate that the Xenopus I(Cl(Ca)) is modulated by phosphorylation. This may complicate design and interpretation of studies of G protein-coupled receptors using this model. PMID- 15271717 TI - Sevoflurane promotes endothelium-dependent smooth muscle relaxation in isolated human omental arteries and veins. AB - Anesthesia with sevoflurane is accompanied by vasodilatation. This could be due to the effects of sevoflurane on endothelium-dependent relaxation. We measured muscle tension of isolated human omental arteries and veins in response to substance P or glyceryl trinitrate in the presence of sevoflurane (0%, 1%, 2%, or 4%). Vascular levels of guanosine 3', 5'-cyclic monophosphate were measured with enzyme-linked immunosorbent assay. Substance P induced an endothelium- and concentration-dependent relaxation in omental vessels that was not affected by sevoflurane. In the presence of L-N(G)-nitroarginine methyl ester (nitric oxide synthase inhibitor), KCl (prevention of hyperpolarization), or both, sevoflurane at 4% enhanced the relaxation in the arteries (P < 0.05). In the vein segments, the relaxation was enhanced by sevoflurane at 4% in the presence of KCl and 2% and 4% in the presence of both L-N(G)-nitroarginine methyl ester and KCl (P < 0.05). The glyceryl trinitrate-induced endothelium-independent relaxation was enhanced by sevoflurane at 4% in both artery and vein segments (P < 0.05). Substance P increased the levels of guanosine 3', 5'-cyclic monophosphate similarly in the presence and absence of sevoflurane. These results show that sevoflurane, in contrast to its effect in animal models, promotes endothelium dependent relaxation in human omental arteries and veins via an enhancement of the smooth muscle response to relaxing second messengers. PMID- 15271718 TI - The delay of gastric emptying induced by remifentanil is not influenced by posture. AB - Posture has an effect on gastric emptying. In this study, we investigated whether posture influences the delay in gastric emptying induced by opioid analgesics. Ten healthy male subjects underwent 4 gastric emptying studies with the acetaminophen method. On two occasions the subjects were given a continuous infusion of remifentanil (0.2 microg. kg(-1). min(-1)) while lying either on the right lateral side in a 20 degrees head-up position or on the left lateral side in a 20 degrees head-down position. On two other occasions no infusion was given, and the subjects were studied lying in the two positions. When remifentanil was given, there were no significant differences between the two postures in maximal acetaminophen concentration (right side, 34 micromol. L(-1); versus left side, 16 micromol. L(-1)), time taken to reach the maximal concentration (94 versus 109 min), or area under the serum acetaminophen concentration time curve from 0 to 60 min (962 versus 197 min. micromol. L(-1)). In the control situation, there were differences between the postures in maximal acetaminophen concentration (138 versus 94 micromol. L(-1); P < 0.0001) and area under the serum acetaminophen concentration time curves from 0 to 60 min (5092 versus 3793 min. micromol. L( 1); P < 0.0001), but there was no significant difference in time taken to reach the maximal concentration (25 versus 47 min). Compared with the control situation, remifentanil delayed gastric emptying in both postures. We conclude that remifentanil delays gastric emptying and that this delay is not influenced by posture. PMID- 15271719 TI - Volatile anesthetics and succinylcholine in cardiac ryanodine receptor defects. AB - Familial polymorphic (catecholaminergic) ventricular tachycardia is an arrhythmogenic cardiac disorder caused by mutations of the myocardial isoform of the ryanodine receptor gene (RyR2). Mutations of the corresponding gene in the skeletal muscle (RyR1) predispose its carriers to malignant hyperthermia upon use of volatile anesthetics or succinylcholine, which further deteriorate the inherited intracellular calcium release disorder. We report a series of patients with cardiac RyR defects who underwent general anesthesia without complications. Succinylcholine and volatile anesthetics did not have a clinically significant effect on RyR2 defects. PMID- 15271720 TI - Anesthetic management of patients with severe peripheral ischemia due to calciphylaxis. AB - Calciphylaxis is a small-vessel disease associated with renal failure. Here, we report the management of a 43-yr-old man with calciphylaxis who received left lower leg amputation with prostaglandin E(1) (PGE(1)) under monitoring by laser Doppler blood flowmetry in the left second and third fingers. Anesthesia was induced with midazolam, fentanyl, and vecuronium and was maintained with oxygen, nitrous oxide, and sevoflurane. The peripheral blood flow varied and decreased gradually; therefore, we added PGE(1) 20 ng. kg(-1). min(-1), which increased blood flow of the tissues. Three weeks after the operation, we again anesthetized the patient. We maintained the blood flow with PGE(1) throughout anesthesia. Monitoring by laser Doppler blood flowmetry and PGE(1) 20 ng. kg(-1). min(-1) could be useful for patients with impaired peripheral circulation, as in calciphylaxis. PMID- 15271721 TI - Aspirin withdrawal and acute lower limb ischemia. AB - Aspirin is used mainly to prevent arterial events in patients with arteriopathy. Myocardial infarction and cerebrovascular events have been described after recent aspirin withdrawal. Experimental data suggest rebound platelet activity after aspirin discontinuation. Among a retrospective cohort of 181 patients admitted for acute lower limb ischemia for 4 yr, we studied 11 patients who had recently stopped taking aspirin. Aspirin was administered for vascular event prevention. The median duration of aspirin treatment without vascular events was 12 mo (range, 6-60 mo). The median time between aspirin withdrawal and lower limb ischemia was 23 days (range, 7-60 days). Four of the 11 patients stopped aspirin before a surgical procedure, without any substitution. In five patients, a recent diagnosis of neoplasia was observed. This study should alert clinicians to the risk of discontinuing chronic aspirin therapy in patients with severe peripheral vascular disease. PMID- 15271722 TI - Increased carbon monoxide concentration in exhaled air after surgery and anesthesia. AB - Heme oxygenase-1 (HO-1) is induced by oxidative stress and is thought to confer protection against oxidative tissue injuries. HO-1 catalyzes the conversion of the heme moiety of hemeproteins, such as hemoglobin, myoglobin, and cytochrome P450, to biliverdin, liberating carbon monoxide (CO) in the process. CO reacts with hemoglobin to form carboxyhemoglobin. In this study, to examine the effect of anesthesia and/or surgery on endogenous CO production, we measured the amount of exhaled CO and the arterial carboxyhemoglobin concentration of patients who underwent surgery under general or spinal anesthesia. Both CO and carboxyhemoglobin concentrations were significantly larger on the day after surgery than during the preoperative period (P < 0.05) and in the recovery room (P < 0.05), regardless of anesthesia. However, neither index differed between general and spinal anesthesia. These results suggest that oxidative stress caused by anesthesia and/or surgery may induce HO-1, which catalyzes heme to produce CO, leading to increased exhaled CO concentration. PMID- 15271723 TI - Changes in thrombelastograph variables associated with aging. AB - Aging is associated with hypercoagulability. To assess thrombelastography (TEG) variables associated with aging, 132 adult patients of various ages undergoing orthopedic surgery for fracture repair had venous blood samples withdrawn for testing of recalcified TEG before the induction of anesthesia. Age was weakly correlated with all TEG variables: r time (R) (r = -0.45, P < 0.001; R = 19.5 - 0.09 x age), k time (K) (r = -0.49, P < 0.001; K = 6.5 - 0.04 x age), maximum amplitude (MA) (r = 0.25, P < 0.01; MA = 53.3 + 0.07 x age), and alpha (r = 0.52, P < 0.001; alpha = 52.8 + 0.2 x age). The correlation was stronger for men than for women. Only R was significantly correlated with age when the women were separately analyzed. Part of the correlation may be attributable to a concurrent decrease in hemoglobin with aging, but age remained an independent predictor of R, K, and alpha on forward stepwise linear multiple regression analysis. Aging was weakly associated with changes in TEG variables, which should be allowed for when interpreting TEG measurements in the elderly. PMID- 15271724 TI - Towards a mechanisms-based approach to pain medicine. PMID- 15271725 TI - Differential analgesic sensitivity of two distinct neuropathic pain models. AB - Progressive tactile hypersensitivity (PTH) manifesting after sciatic nerve crush and spared nerve injury (SNI) are two distinct rodent experimental models of neuropathic pain. PTH develops months after recovery from the nerve crush in response to repeated intermittent low-threshold mechanical stimulation of the reinnervated sciatic nerve skin territory and represents a model of stimulus induced pain. SNI is characterized by an early and sustained increase in stimulus evoked pain sensitivity in the intact skin territory of the spared sural nerve after sectioning of the two other terminal branches of the sciatic nerve. We examined the effects of morphine (0.5-10 mg/kg), gabapentin (30-200 mg/kg), MK801 (0.01-0.02 mg/kg), amitriptyline (10-25 mg/kg), and carbamazepine (5-7.5 mg/kg) in both models. Morphine, gabapentin, and carbamazepine both reversed and prevented stimulus-induced PTH, whereas MK801 and amitriptyline reduced but did not prevent stimulus-induced PTH. In contrast, the stimulus-evoked behavioral hypersensitivity in the SNI model was poorly modified by these drugs. Independent neuropathic pain models show differential sensitivity to analgesic drug treatment. We suggest that this is due to the different mechanisms responsible for the neuropathic pain-related behavior. Multiple models are required, therefore, to study the mechanisms that contribute to neuropathic pain and to predict analgesic efficacy for different components of the neuropathic pain syndrome. PMID- 15271726 TI - The effects of class Ic antiarrhythmics on tetrodotoxin-resistant Na+ currents in rat sensory neurons. AB - IV or oral administration of antiarrhythmics has been reported to be effective for relieving neuropathic pain. Recent reports have indicated that tetrodotoxin resistant (TTX-R) Na(+) channels play important roles in the nerve conduction of nociceptive sensation. In the present study, we investigated the effects of flecainide, pilsicainide (class Ic antiarrhythmics), and lidocaine (a class Ib drug) on TTX-R Na(+) currents in rat dorsal root ganglion neurons using the whole cell patch-clamp method. Flecainide, pilsicainide, and lidocaine reversibly blocked the peak amplitude of TTX-R Na(+) currents in a concentration-dependent manner with half-maximum inhibitory concentration values of 8.5 +/- 6.6 microM (n = 7), 78 +/- 6.9 microM (n = 7), and 73 +/- 6.8 microM (n = 7), respectively. Each drug shifted the inactivation curve for the TTX-R Na(+) currents in the hyperpolarizing direction and caused a use-dependent block. We also studied an interaction between these antiarrhythmics on TTX-R Na(+) channels. Additional application of flecainide or pilsicainide to lidocaine resulted in an additive increase of tonic and use-dependent block. These results suggest that the inhibition of TTX-R Na(+) currents of dorsal root ganglion neurons by such antiarrhythmics is attributable, at least partly, to their antinociceptive effects. PMID- 15271727 TI - Intradiscal thermal annuloplasty for the treatment of lumbar discogenic pain in patients with multilevel degenerative disc disease. AB - Symptomatic degenerative disc disease (DDD) may lead to significant deterioration of quality of life and increased disability. Intradiscal thermal annuloplasty (IDTA) is a minimally invasive treatment for painful DDD. We hypothesized that there may be an improvement in pain scores and the pain disability index (PDI) of patients who have multilevel DDD after IDTA. Patients 24-66 yr old, male and female with multilevel DDD (MDDD) and matched 1 or 2 level DDD (1,2-DDD) patients were enrolled in the study. Visual analog pain scale (VAS) score and PDI were observed for 12 mo. The 1,2-DDD patient group had a 2.5 +/- 2.4 VAS score at 12 mo after annuloplasty compared to 7.7 +/- 2 before the procedure. The MDDD VAS score was 4.9 +/- 2.9 at 12 mo compared to 7.4 +/- 1.8 before the procedure. Similar improvements in PDI were found. The pain relief and PDI were significantly better in patients with 1,2-DDD than in the MDDD group (P = 0.0037 and P = 0.041, respectively). We concluded that IDTA is an effective treatment of discogenic pain and that the number of discs affected by degeneration is an important determinant of the procedure outcome. PMID- 15271728 TI - An analysis of the relationship between activity and pain in chronic and acute low back pain. AB - We studied the temporal relationship between pain and activity in patients with acute or chronic low back pain. We studied 15 patients with acute low back pain and 15 patients with chronic low back pain over 3 wk. The activity levels were collected automatically using a wrist accelerometer and were sampled every minute. The pain levels were recorded at least every 90 min using a pocket-sized electronic diary. The time series from each patient were then analyzed using the cross-correlation function at various time offsets. We found that during the first 7 days of acute low back pain, there was a significant (P < 0.01) degree of cross-correlation between activity and pain. On average, pain followed activity by approximately 30 min. As these patients improved and reported less pain, the relationship between activity and pain disappeared. There was no such relationship at any point among the patients with chronic low back pain. PMID- 15271729 TI - Ketamine as adjuvant analgesic to opioids: a quantitative and qualitative systematic review. AB - Animal studies on ketamine and opioid tolerance have shown promising results. Clinical trials have been contradictory. We performed a systematic review of randomized, double-blind clinical trials of ketamine added to opioid analgesia. Thirty-seven trials with 51 treatment arms and 2385 patients were included. Studies were divided into 5 subgroups: IV ketamine as single dose (n = 11), continuous infusion (n = 11), patient-controlled analgesia (PCA) (n = 6), epidural ketamine with opioids (n = 8), and studies in children (n = 4). Outcome measures included pain scores, time to first request for analgesia, supplemental analgesics, and adverse events. Efficacy was estimated by statistical significance (P < 0.05) of outcome measures as reported in studies and also by calculation of weighted mean difference for pain scores during the first 24 h after surgery. As compared to morphine alone, IV PCA with ketamine and morphine did not improve analgesia. Intravenous infusion of ketamine decreased IV and epidural opioid requirements in 6 of 11 studies. A single bolus dose of ketamine decreased opioid requirements in 7 of 11 studies. Five of 8 trials with epidural ketamine showed beneficial effects. Adverse effects were not increased with small dose ketamine. We conclude that small dose ketamine is a safe and useful adjuvant to standard practice opioid-analgesia. PMID- 15271730 TI - Lumbar segmental nerve blocks with local anesthetics, pain relief, and motor function: a prospective double-blind study between lidocaine and ropivacaine. AB - Selective segmental nerve blocks with local anesthetics are applied for diagnostic purposes in patients with chronic back pain to determine the segmental level of the pain. We performed this study to establish myotomal motor effects after L4 spinal nerve blocks by lidocaine and ropivacaine and to evaluate the relationship with pain. Therefore, 20 patients, of which 19 finished the complete protocol, with chronic lumbosacral radicular pain without neurological deficits underwent segmental nerve blocks at L4 with both lidocaine and ropivacaine. Pain intensity scores (verbal numeric rating scale; VNRS) and the maximum voluntary muscle force (MVMF; using a dynamometer expressed in newtons) of the tibialis anterior and quadriceps femoris muscles were measured on the painful side and on the control side. The median VNRS decrease was 4.0 (P < 0.00001; Wilcoxon's signed rank test), without significant differences between ropivacaine and lidocaine (Mann-Whitney U-test). A difference in effect on MVMF was found for affected versus control side (P = 0.016; Tukey test). Multiple regression revealed a significant negative correlation for change in VNRS score versus change in median MVMF (Spearman R = -0.48: P = 0.00001). This study demonstrates that in patients with unilateral chronic low back pain radiating to the leg, pain reduction induced by local anesthetic segmental nerve (L4) block is associated with increased quadriceps femoris and tibialis anterior MVMF, without differences for lidocaine and ropivacaine. PMID- 15271731 TI - The effect of epidural clonidine on perioperative cytokine response, postoperative pain, and bowel function in patients undergoing colorectal surgery. AB - The postoperative period is associated with an increased production of cytokines, which augment pain sensitivity. We investigated the hypothesis that epidural clonidine premedication and postoperative patient-controlled epidural analgesia (PCEA) including clonidine would decrease the release of proinflammatory (interleukin (IL)-6, IL-1beta, IL-8, and tumor necrosis factor (TNF)-alpha) and antiinflammatory (IL-1 receptor antagonist (RA)) cytokines in patients who underwent elective colorectal surgery and that they would provide better postoperative analgesia. Forty patients were randomly assigned to 1 of 2 groups of 20 each: the control group received normal saline 10 mL, whereas the clonidine group received epidural clonidine 150 microg diluted with 9 mL of normal saline 30 min before surgery. Venous blood samples for cytokine levels were obtained before induction, at the end of surgery, and after surgery at 12 and 24 h. After surgery, the clonidine group patients received PCEA with morphine (0.1 mg/mL) and clonidine (1.5 microg/mL) in 0.2% ropivacaine 100 mL, whereas control group patients received only PCEA morphine and ropivacaine. Patients in the clonidine group exhibited longer PCEA trigger times, lower pain scores at rest and while coughing, less morphine consumption, and a faster return of bowel function throughout the 72-h postoperative observation period, compared with patients in the control group. For patients in the clonidine group, production of IL-1RA, IL 6, and IL-8 was significantly less increased at the end of the surgical procedure and at 12 and 24 h after surgery. However, the concentrations of IL-1beta and TNF alpha were not significantly increased. PMID- 15271732 TI - Persistent pain as a disease entity: implications for clinical management. AB - Pain has often been regarded merely as a symptom that serves as a passive warning signal of an underlying disease process. Using this model, the goal of treatment has been to identify and address the pathology causing pain in the expectation that this would lead to its resolution. However, there is accumulating evidence to indicate that persistent pain cannot be regarded as a passive symptom. Continuing nociceptive inputs result in a multitude of consequences that impact on the individual, ranging from changes in receptor function to mood dysfunction, inappropriate cognitions, and social disruption. These changes that occur as a consequence of continuing nociceptive inputs argue for the consideration of persistent pain as a disease entity in its own right. As with any disease, the extent of these changes is largely determined by the internal and external environments in which they occur. Thus genetic, psychological and social factors may all contribute to the perception and expression of persistent pain. Optimal outcomes in the management of persistent pain may be achieved not simply by attempting to remove the cause of the pain, but by addressing both the consequences and contributors that together comprise the disease of persistent pain. PMID- 15271733 TI - Preliminary report on the use of high-fidelity simulation in the training of study coordinators conducting a clinical research protocol. AB - Training of health care research personnel is a critical component of quality assurance in clinical trials. Interactivity (such as simulation) is desirable compared with traditional methods of teaching. We hypothesized that the addition of an interactive simulation exercise to standard training methods would increase the confidence of study coordinators. A simulation exercise was developed to replicate a complex clinical trial. Eighteen study coordinators completed pre- and postexercise confidence questionnaires. Questions were targeted at key trial components using a 0-10 scale (not confident to confident) and were categorized using Bloom's Taxonomy. The primary analysis compared overall mean pre- and postexercise responses. Secondary analyses assessed affective, psychomotor, and cognitive confidence. Significance was at P < 0.05. A significant increase in overall confidence (8.64 versus 5.77; P < 0.0001) was reproduced in the subcategory analyses (affective, 8.24 versus 4.89; P < 0.0001; cognitive, 8.75 versus 6.42; P = 0.0003; psychomotor, 8.63 versus 5.26; P < 0.0001). A high level of internal consistency and reliability in question responses within domains was observed, validating the questionnaire tool. In this preliminary report, we confirmed that addition of a simulation exercise to the training of study coordinators resulted in increased confidence. Simulation exercises should be considered when training study coordinators for clinical research trials. PMID- 15271734 TI - Increased cerebral tissue oxygen tension after extensive hemodilution with a hemoglobin-based oxygen carrier. AB - Transfusion of anemic patients with hemoglobin-based oxygen carriers (HBOCs) may improve cerebral oxygen delivery. Conversely, cerebral vasoconstriction, associated with HBOC transfusion, could limit optimal cerebral tissue oxygenation. We hypothesized that hemodilution with a HBOC would maintain cerebral tissue oxygenation, despite the occurrence of cerebral vasoconstriction. Isoflurane-anesthetized rats (100% oxygen) underwent direct measurement of mean arterial blood pressure (MAP), caudate tissue oxygen tension (P(Br)o(2)), and regional cortical cerebral blood flow (rCBF) before and after 50% of the estimated blood volume (30 mL/kg) was exchanged with either an HBOC (hemoglobin raffimer; Hemolink) or pentastarch (n = 6). Hemodilution with hemoglobin raffimer caused a transient increase in P(Br)o(2) from 24.9 +/- 13.3 mm Hg to 32.2 +/- 19.1 mm Hg (P < 0.05), a sustained increase in MAP, and no change in rCBF. Arterial blood oxygen content was maintained despite an increase in methemoglobin and reduced oxygen saturation. Hemodilution with pentastarch caused a transient increase in MAP, no change in P(Br)o(2), and a sustained increase in rCBF (P < 0.05), whereas the hemoglobin concentration and oxygen content were significantly reduced. Hemodilution with hemoglobin raffimer augmented P(Br)o(2) and prevented the increase in rCBF observed after similar hemodilution with pentastarch. These data suggest that transfusion with hemoglobin raffimer may help to maintain cerebral oxygenation during severe anemia. PMID- 15271735 TI - The early systemic and gastrointestinal oxygenation effects of hemorrhagic shock resuscitation with hypertonic saline and hypertonic saline 6% dextran-70: a comparative study in dogs. AB - The smaller volemic state from hypertonic (7.5%) saline (HS) solution administration in hemorrhagic shock can determine lesser systemic oxygen delivery and tissue oxygenation than conventional plasma expanders. In a model of hemorrhagic shock in dogs, we studied the systemic and gastrointestinal oxygenation effects of HS and hyperoncotic (6%) dextran-70 in combination with HS (HSD) solutions in comparison with lactated Ringer's (LR) and (6%) hydroxyethyl starch (HES) solutions. Forty-eight mongrel dogs were anesthetized, mechanically ventilated, and subjected to splenectomy. A gastric air tonometer was placed in the stomach for intramucosal gastric CO(2) (Pgco(2)) determination and for the calculation of intramucosal pH (pHi): The dogs were hemorrhaged (42% of blood volume) to hold mean arterial blood pressure at 40-50 mm Hg over 30 min and were then resuscitated with LR (n = 12) in a 3:1 relation to removed blood volume; HS (n = 12), 6 mL/kg; HSD (n = 12), 6 mL/kg; and HES (mean molecular weight, 200 kDa; degree of substitution, 0.5) (n = 12) in a 1:1 relation to the removed blood volume. Hemodynamic, systemic, and gastric oxygenation variables were measured at baseline, after 30 min of hemorrhage, and 5, 60, and 120 min after intravascular fluid resuscitation. After fluid resuscitation, HS showed significantly lower arterial pH and mixed venous Po(2) and higher systemic oxygen uptake index and systemic oxygenation extraction than LR and HES (P < 0.05), whereas HSD showed significantly lower arterial pH than LR and HES (P < 0.05). Only HS and HSD did not return arterial pH and pHi to control levels (P < 0.05). In conclusion, all solutions improved systemic and gastrointestinal oxygenation after hemorrhagic shock in dogs. However, the HS solution showed the worst response in comparison to LR and HES solutions in relation to systemic oxygenation, whereas HSD showed intermediate values. HS and HSD solutions did not return regional oxygenation to control values. PMID- 15271736 TI - Prevention of hemodilution-induced inhibition of hypoxic pulmonary vasoconstriction by N-acetylcysteine in dogs. AB - We investigated the possible contributions of reactive oxygen species and of viscosity changes to hemodilution-induced inhibition of hypoxic pulmonary vasoconstriction (HPV) in dogs. Fourteen isoflurane-anesthetized dogs were randomly assigned to receive N-acetylcysteine (NAC) 200 mg/kg IV (n = 7) or placebo (n = 7). Mean pulmonary artery pressure (Ppa) was measured with cardiac output maintained constant by a manipulation of venous return in hyperoxia (fraction of inspired oxygen, 0.4) and in hypoxia (fraction of inspired oxygen, 0.1) at baseline and after stepwise reductions in hematocrit from 40% to 20%. Measured Ppa was compared with predicted Ppa by using a viscoelastic model. HPV was expressed as hypoxic Ppa minus hyperoxic Ppa. Hemodilution was associated with a decrease in HPV from 7 +/- 1 mm Hg to 3 +/- 1 mm Hg (P < 0.01), and this was completely prevented by NAC (HPV was unchanged, from 8 +/- 1 to 8 +/- 1 mm Hg; not significant). Hemodilution in the model decreased HPV from 8 +/- 1 mm Hg to 6 +/- 1 mm Hg (P < 0.05). We conclude that hemodilution-induced inhibition of HPV is in part explained by viscosity changes and can be prevented by the administration of NAC, which is possibly explained by the scavenging of reactive oxygen species. PMID- 15271737 TI - Patient-controlled analgesia with fentanyl for burn dressing changes. AB - In this randomized, double-blinded study in 60 ASA I or II adults with >20% body surface area thermal burns, we investigated the feasibility of patient-controlled analgesia (PCA) with fentanyl for pain management during dressing changes and determined the optimal PCA-fentanyl demand dose. An initial loading dose of IV fentanyl 1 microg/kg was administered. Patients received on-demand analgesia with fentanyl (10, 20, 30, and 40 microg) whenever their visual analog scale (VAS) score was >2. Mean VAS scores in the 10 and 20 microg groups (7.73 +/- 1.33 and 7.20 +/- 1.21, respectively) were significantly higher than those in the 30 and 40 microg groups (4.47 +/- 0.83 and 3.90 +/- 0.63, respectively) (all P = 0.000). Demand/delivery ratios were significantly larger in the 10 and 20 microg groups (3.03 +/- 1.06 and 2.54 +/- 0.49, respectively) than those in the 30 and 40 microg groups (1.36 +/- 0.34 and 1.37 +/- 0.36, respectively) (all P = 0.000). VAS scores and demand/delivery ratios were comparable in the 30 and 40 microg groups (P = 0.260 and P = 0.977, respectively), which suggests comparable analgesic efficacy. There was no hemodynamic instability or respiratory depression. The optimal demand dose of PCA-fentanyl was 30 microg (5-min lockout interval) after an initial loading dose of IV fentanyl 1 microg/kg. PMID- 15271738 TI - The protective effect of protein kinase C and adenosine triphosphate-sensitive potassium channel agonists against inflammation in rat endothelium and vascular smooth muscle in vitro and in vivo. AB - Volatile anesthetic pretreatment protects the vasculature from inflammation induced injury via mechanisms involving the activation of adenosine triphosphate sensitive potassium (K(ATP)) channels and/or protein kinase C (PKC). Therefore, we hypothesized that K(ATP) and PKC agonists may mimic the protective effects of volatile anesthetics in vitro and in vivo. In vitro, rat vascular smooth muscle cells (VSM) and aortic endothelial cells (AEC) were used to evaluate whether pretreatment with a K(ATP) agonist, cromakalim (CRK), or a PKC agonist, phorbol 12-myristate 13-acetate (PMA), decreases lipopolysaccharide (LPS)-induced cell injury. Cell survival was determined by trypan blue staining after 6 h. In vivo, rats received systemic LPS or saline with or without pretreatment with PMA or CRK. Mean arterial blood pressure, the response to endothelium-dependent (acetylcholine; ACH) and -independent (sodium nitroprusside) vasodilators, and arterial blood gases were determined after 6 h. Cell survival in VSM and AEC control cultures was more than 90%, which was not altered in the presence of PMA or CRK, whereas LPS significantly decreased cell survival. PMA (0.1-10 microM) significantly attenuated the LPS-induced decrease in cell survival by 28%-37% in VSM and 39%-53% in AEC, and CRK (1 mM) increased cell survival by 24% in VSM and 22% in AEC. In vivo, PMA and CRK pretreatment had no significant effect on measured variables in control rats. LPS decreased mean arterial blood pressure and vasodilation to ACH and sodium nitroprusside and caused hypoglycemia. PMA, but not CRK, increased ACH-dependent vasodilation (46%) at 6 h, but neither agonist altered the other detrimental effects of LPS. In conclusion, PKC and K(ATP) agonists appear to protect AEC and VSM cells against inflammation in vitro, but the systemic administration of PKC and K(ATP) agonists appeared to exert minimal or no protection in our in vivo model. PMID- 15271739 TI - The use of ultrasound for axillary artery catheterization through pectoral muscles: a new anterior approach. AB - A palpable axillary artery pulse is a prerequisite for introducing an arterial line. The close proximity of four nerves to the artery increases the chance of nerve injury, especially in anesthetized patients. The highly colonized entry site results in frequent infection. Approaching the axillary artery through the pectoral muscles by using real-time imaging should improve success, decrease infection, and prevent nerve and vessel injuries because these structures and the needle can be visualized directly. I describe three patients who had successful axillary lines placed through the pectoral muscles by using real-time sonography. The ability to see the artery, surrounding nerves, and vein and to observe the needle going through the tissues should increase safety and success, although a large study is needed to prove these hypotheses. PMID- 15271740 TI - Evidence-based management of critically ill patients: analysis and implementation. AB - A number of important clinical trials focusing on critically ill patients have been completed in the last few years. These trials have been among the first critical care clinical trials to demonstrate mortality reduction in the critically ill. As in any adaptation of evidence-based medicine, it is essential to closely examine the trials and to determine whether the demonstrated benefits can be translated to the individual patient. In addition to the primary outcome, usually survival benefit, it is also important to examine cost-effectiveness. All of the trials examined in this review were able to demonstrate mortality reduction. Most focused on patients with severe sepsis, because this population has been associated with both frequent mortality and increased hospital costs. Some of the interventions, such as small tidal volume mechanical ventilation in patients with acute lung injury or the administration of low-dose corticosteroids for patients with septic shock, are cost-effective and relatively simple to implement. Others, such as use of activated protein C in patients with severe sepsis or "tight" glycemic control in patients with hyperglycemia, require either significant pharmaceutical expenditure or, possibly, additional health care personnel. Nevertheless, the trials discussed represent significant advances in the field of critical care medicine and should at least be considered for implementation in all intensive care units. PMID- 15271741 TI - The effect of propofol sedation on the intracranial pressure of patients with an intracranial space-occupying lesion. AB - The fear of producing CO(2) retention and a secondary increase of intracranial pressure (ICP) sometimes precludes the use of sedation for the spontaneously breathing patient in the presence of an intracranial space-occupying lesion. In this study we assessed the effect of moderately deep propofol sedation on the ICP of patients undergoing stereotactic brain tumor biopsy under regional anesthesia. Thirty patients were randomized into 2 groups to receive propofol titrated to a level of 2 on the Observer's Assessment of Alertness/Sedation Scale or no sedation. ICP was measured via the biopsy needle. Preoperative data were similar in both groups. During surgery, patients receiving propofol had a higher arterial Pco(2) (48 +/- 8 mm Hg versus 41 +/- 3 mm Hg; P = 0.005) (95% confidence interval, 43-53 mm Hg and 39-43 mm Hg, respectively), resulting in a lower arterial pH (P = 0.002) than patients in the no-sedation group. The median ICP (95% confidence interval) for both groups was similar-13 mm Hg (8.2-16.2 mm Hg) and 15 mm Hg (8.3-21.7 mm Hg)-for the propofol and no-sedation groups, respectively (P = 0.66). Cerebral perfusion pressure was lower in the propofol group (76 +/- 18 mm Hg versus 89 +/- 18 mm Hg; P = 0.003). Moderately deep propofol sedation does not result in a higher ICP than no sedation in patients undergoing stereotactic brain tumor biopsy. Further studies are needed to assess the effect on ICP of other sedative medications. PMID- 15271742 TI - Thoracic epidural anesthesia: asleep at the wheal? PMID- 15271743 TI - Paraplegia after delayed detection of inadvertent spinal cord injury during thoracic epidural catheterization in an anesthetized elderly patient. AB - We report a case of permanent paraplegia in an 81-yr-old patient who had thoracic epidural catheterization performed under general anesthesia for abdominal surgery. The epidural needle was introduced at the T9-10 interspace, and 3 passes were made to locate the epidural space with the loss-of-resistance-to-air technique. During the postoperative epidural pump infusion, the patient was unaware of the progressive motor and sensory impairment. Sensory loss below T11 and paraplegia with no movement of either lower extremity were identified 8 h after surgery. Magnetic resonance imaging demonstrated an intramedullary split like lesion extending from T4 to T12 and an intramedullary air bubble at T9. Spinal cord injury caused by an intracord catheterization with subsequent local anesthetic injection was diagnosed. Little improvement was noted after large-dose IV methylprednisolone for initial treatment and subsequent rehabilitation for 6 mo. The possible causes of the delayed detection of the neurologic deficits and the timing of performing epidural anesthesia are discussed. PMID- 15271744 TI - Nerve stimulator-assisted evoked motor response predicts the latency and success of a single-injection sciatic block. AB - Variable onset latency of single-injection sciatic nerve block (SNB) may result from drug deposition insufficiently close to all components of the nerve. We hypothesized that this variability is caused by the needle tip position relative to neural components, which is objectified by the type of evoked motor response (EMR) elicited before local anesthetic injection. One-hundred ASA I-II patients undergoing reconstructive ankle surgery received infragluteal-parabiceps SNB using 0.4 mL/kg (maximum 35 mL) of levobupivacaine 0.625%. The end-point for injection was the first elicited EMR: inversion (I), plantar flexion (PF), dorsiflexion (DF), or eversion (E) at 0.2-0.4 mA. The frequencies of the EMRs were: I 40%, PF 43%, E 14%, and DF 3%. SNB was considered complete if both tibial and common peroneal nerves were blocked and failed if either analgesia to pinprick was not observed at 30 min or anesthesia at 60 min. Patients with an EMR of I demonstrated shorter mean times (+/-95% confidence interval [CI]) to complete the block with 8.5 (95% CI, 6.2-10.8) min compared to 27.0 (95% CI, 20.6 33.4) min after PF (P < 0.001) and 30.4 (95% CI, 24.9-35.8) min after E (P < 0.001). No rescue blocks were required in group I compared with 24% (P = 0.001) and 71% (P < 0.001) of patients in groups PF and E, respectively. We conclude that EMR type during nerve stimulator-assisted single-injection SNB predicts latency and success of complete SNB because the observed EMR is related to the positioning of the needle tip relative to the tibial and common peroneal nerves. PMID- 15271745 TI - Pain relief after arthroscopic shoulder surgery: a comparison of intraarticular analgesia, suprascapular nerve block, and interscalene brachial plexus block. AB - In this prospective, randomized, blinded study, we assessed the analgesic efficacy of interscalene brachial plexus block (ISB), suprascapular nerve block (SSB), and intraarticular local anesthetic (IA) after arthroscopic acromioplasty. One-hundred-twenty patients were divided into 4 groups of 30. In Group SSB, the block was performed with 10 mL of 0.25% bupivacaine. In Group IA, 20 mL of 0.25% bupivacaine was administered intraarticularly at the end of surgery. In Group ISB, the block was performed with 20 mL of 0.25% bupivacaine. A control group was included for comparison. General anesthesia was administered to all patients. Patients were observed during the first 24 h. Pain scores, supplemental analgesia, satisfaction scores, and side effects were recorded at 4 and 24 h. No significant difference was observed between the IA and control groups. When compared with these groups, Groups SSB and ISB had significantly lower pain scores. At 4-h follow-up, better pain relief on movement was noted in Group ISB than in Group SSB. When compared with controls, a significant reduction in morphine consumption and a better satisfaction score were noted only in Group ISB. We conclude that ISB is the most efficient analgesic technique after arthroscopic acromioplasty. SSN block would be a clinically appropriate alternative. PMID- 15271746 TI - Seizures after a Bier block with clonidine and lidocaine. AB - A 47-yr-old man with history of complex regional pain syndrome type 1 underwent an IV Bier block with a mixture of lidocaine and clonidine. The tourniquet was deflated after 60 min, and approximately 10 min later he presented with complex partial seizures. The possible mechanisms for this are discussed, and the effects of clonidine, lidocaine, and the mixture of both are reviewed, as are four additional published cases reporting seizures after the administration of clonidine. PMID- 15271747 TI - Intraoperative recombinant activated factor VII for emergent epidural hematoma evacuation. AB - We report a case of a chronically anticoagulated 59-yr-old woman who underwent an L4 to L5 epidural block to relieve her low back pain and subsequently developed a T7 to L5 epidural hematoma with cauda equina and conus compression. Fresh frozen plasma and vitamin K were given before surgery, whereas recombinant activated factor VII was administered during surgery to reverse the coagulopathy and to enable the emergent laminectomy and hematoma evacuation. Recombinant activated factor VII administration proved to be a useful adjunct in the emergent surgical management of a thoracolumbar epidural hematoma. PMID- 15271748 TI - A simple glucose insulin regimen for perioperative blood glucose control: the Vellore regimen. AB - In this study, we sought a simple, easily implemented method of intraoperative control of blood glucose in diabetic patients in a large multispecialty teaching hospital. The Vellore regimen, which offers the advantages of a combined glucose insulin and variable rate infusion was evaluated. For every 1 to 50-mg/dL increase in blood glucose concentration more than 100 mg/dL, 1 U of insulin was added to the injection port of a 100-mL measured volume set containing 5% dextrose in water. Hourly monitoring of blood glucose was performed. The blood glucose control was compared with the different existing techniques followed in the hospital in 204 randomized patients: 98 in the study and 106 in the control group. The study group had a mean +/- sd blood glucose value of 156 +/- 36 mg/dL, and the control group's value was 189 +/- 63 mg/dL (P = 0.003). The percentage of patients who were poorly controlled (outside 100 to 200-mg/dL range) decreased from 51% to 28% (no patient less than 60 mg/dL) with this regimen as compared with the control group in which it increased from 49% to 72% (10 patients less than 60 mg/dL) (P = 0.0013). We conclude that the Vellore regimen is simple, effective, and safe for intraoperative blood glucose control. PMID- 15271749 TI - Prediction of difficult tracheal intubation in thyroid surgery. AB - The incidence of difficult endotracheal intubation (DEI) for patients undergoing thyroidectomy has rarely been studied, and evaluation of factors linked to DEI is limited to a few studies. We undertook this prospective study to investigate the incidence of DEI in the presence of goiter (an enlargement of the thyroid gland) and to evaluate factors linked to DEI. We studied 320 consecutive patients scheduled for thyroidectomy. DEI was evaluated by an intubation difficulty scale. The trachea was intubated by an unassisted anesthesiologist, and the intubation difficulty scale was calculated. A univariate analysis was performed to identify potential factors predicting DEI, followed by a multivariate analysis. DEI was reported in 17 patients. The rate of easy tracheal intubation was 36.9%; the rate for patients who had minor difficulty of intubation was 57.8%. Sex (male), body mass index, Mallampati class, thyromental distance, neck mobility, Cormack grade, cancerous goiter, and tracheal deviation or compression were identified in the univariate analysis as potential DEI risk factors. With multivariate analysis, two criteria were recognized as independent for DEI (Cormack Grade III or IV and cancerous goiter). We conclude that the large goiter is not associated with a more frequent DEI. However, the presence of a cancerous goiter is a major factor for predicting DEI. PMID- 15271750 TI - Emergency tracheal intubation: complications associated with repeated laryngoscopic attempts. AB - Repeated conventional tracheal intubation attempts may contribute to patient morbidity. Critically-ill patients (n = 2833) suffering from cardiovascular, pulmonary, metabolic, neurologic, or trauma-related deterioration were entered into an emergency intubation quality improvement database. This practice analysis was evaluated for airway and hemodynamic-related complications based on a set of defined variables that were correlated to the number of attempts required to successfully intubate the trachea outside the operating room. There was a significant increase in the rate of airway-related complications as the number of laryngoscopic attempts increased (2 attempts): hypoxemia (11.8% versus 70%), regurgitation of gastric contents (1.9% versus 22%), aspiration of gastric contents (0.8% versus 13%) bradycardia (1.6% versus 21%), and cardiac arrest (0.7% versus 11%; P < 0.001). Although predictable, this analysis provides data that confirm the number of laryngoscopic attempts is associated with the incidence of airway and hemodynamic adverse events. These data support the recommendation of the ASA Task Force on the Management of the Difficult Airway to limit laryngoscopic attempts to three in lieu of the considerable patient injury that may occur. PMID- 15271751 TI - Potassium permanganate reduces protein contamination of reusable laryngeal mask airways. AB - We tested the hypothesis that supplementary cleaning with potassium permanganate 2 mg/L eliminates protein deposits from reusable laryngeal mask airways (LMAs). Sixty previously used classic LMAs were hand-washed, machine-washed, dried, autoclaved, and then randomly allocated into two groups for supplementary cleaning. In Group A, the cuff was immersed in potassium permanganate 2 mg/L at 20 degrees C for 20 min. In Group B (control), the cuff was immersed in sterile water at 20 degrees C for 20 min. After supplementary cleaning, the LMAs were immersed in a protein staining solution and rinsed, and a high-resolution digital image was taken of the dorsal surface. The severity of staining was scored by an observer blinded to the type of supplementary cleaning. The severity of protein contamination was reduced after supplementary cleaning in potassium permanganate (P < 0.00001). Protein contamination was detected on 20% of LMAs after supplementary cleaning in potassium permanganate, compared with all LMAs in the control group. We conclude that supplementary cleaning with potassium permanganate 2 mg/L does not eliminate protein deposits from all LMAs, but it does reduce the number of devices contaminated from 100% to 20%. PMID- 15271752 TI - Cerebrospinal fluid rhinorrhea after thermometer insertion through the nose. AB - Cerebrospinal fluid (CSF) rhinorrhea is a rare clinical condition. Most leaks either are caused by traumatic head injury or are a complication of surgical procedures on the base of the skull. CSF rhinorrhea from nasal tube placement has been reported previously. We report a case of nasal thermometer placement during anesthesia complicated by a CSF leakage. We reemphasize that any material- including thermometers, nasogastric tubes, and endotracheal tubes--should be directed posteriorly after introduction into the external naris. PMID- 15271753 TI - Is hydroxyethyl starch safe in brain injury? PMID- 15271754 TI - A well-fertilized bulb should blossom. PMID- 15271755 TI - Falsely low pulse oximetry values in patients receiving docetaxel (Taxotere). PMID- 15271756 TI - Risperidol and asystole. PMID- 15271757 TI - The quest for new devices to improve postoperative pain control. PMID- 15271758 TI - An uncommon reason for damage to the intubating laryngeal mask airway (ILMA) endotracheal tube cuff-inflation system. PMID- 15271759 TI - Peripartum management of a suspected spinal hematoma after epidural puncture. PMID- 15271760 TI - Failure to advance the guidewire when the Seldinger technique is used for central venous cannulation: safe and reliable recovery. PMID- 15271761 TI - The natural half life of a large stock of reusable laryngeal mask airways at a teaching hospital. PMID- 15271762 TI - What is wrong with this picture of pain management? PMID- 15271763 TI - Inadvertent femoral nerve impalement and intraneural injection visualized by ultrasound. PMID- 15271764 TI - Percutaneous dilatational cricothyroidotomy: airway control via CobraPLA. PMID- 15271765 TI - Lifestyle changes in U.S. academic anesthesia: quo vadis? PMID- 15271766 TI - Latex allergy: oh, what a surprise! Another reason why all anesthesia equipment should be latex-free. PMID- 15271767 TI - A nasal bridle for securing nasotracheal tubes. PMID- 15271768 TI - Bilateral brachial plexus block versus segmental epidural anesthesia. PMID- 15271769 TI - The use of tourniquets in patients with sickle cell disease. PMID- 15271770 TI - When a DMARD fails, should patients switch to sulfasalazine or add sulfasalazine to continuing leflunomide? AB - OBJECTIVE: To evaluate the efficacy and safety of adding sulfasalazine to leflunomide treatment compared with switching to sulfasalazine alone in patients with RA with an inadequate response to leflunomide monotherapy. METHODS: Patients with active RA ((DAS28) >3.2) who were enrolled in the first open label phase of the RELIEF study received leflunomide for 24 weeks. Inadequate responders then entered the double blind phase and received a further 24 weeks' treatment with leflunomide (20 mg once daily) plus sulfasalazine (final dose 2 g once daily), or placebo plus sulfasalazine (dose as above). The primary efficacy variable was the DAS28 response rate, and secondary efficacy outcomes were ACR 20%, 50%, and 70% response rates. Adverse events, including standard laboratory tests, were recorded. RESULTS: 106 inadequate responders entered the double blind phase; 56 received leflunomide plus sulfasalazine, and 50 placebo plus sulfasalazine. In the intention to treat population, more patients receiving leflunomide plus sulfasalazine (25/56 (45%)) achieved a DAS28 response than those receiving placebo plus sulfasalazine (17/50 (34%)) (p = 0.179). In week 24 completers, more patients receiving leflunomide plus sulfasalazine (17/56 (30%)) were DAS28 responders than those receiving placebo plus sulfasalazine (10/50 (20%)) (p = 0.081). Comparable numbers in each group were ACR 20% responders; the ACR 50% response rate was significantly higher in the leflunomide plus sulfasalazine group (8.9%) than in the placebo plus sulfasalazine group (0%) (p = 0.038). The safety profiles of both groups were comparable. CONCLUSION: Patient numbers are small and firm conclusions cannot be reached, but a non-significant benefit is indicated for combining leflunomide with sulfasalazine compared with switching to sulfasalazine alone in patients inadequately responding to leflunomide. PMID- 15271771 TI - Dactylitis in psoriatic arthritis: a marker for disease severity? AB - AIM: To describe dactylitis in a large cohort of patients with psoriatic arthritis followed prospectively in a specialist clinic, and identify whether it is associated with a worse prognosis. METHODS: Between 1979 and 1999, 537 patients were registered in the psoriatic arthritis clinic and entered onto a longitudinal database. Patients were followed prospectively at six to 12 month intervals according to a standard protocol, and all information was entered onto a database. The database was searched for patients with dactylitis. Descriptive statistics were used to describe the population and chi(2) tests to relate dactylitis to radiographic changes. RESULTS: Dactylitis was documented in 260 patients (48%); 69% of the episodes were recorded at presentation to the clinic. Dactylitis affected feet only in 65% of cases, hands only in 24%, and both hands and feet in 12%. Recurrent dactylitis occurred in 44% of the patients. Increased radiological progression was noted in digits showing dactylitis compared with those without dactylitis (50% v 38%, respectively; p<0.0001). CONCLUSIONS: Dactylitis is common among patients with psoriatic arthritis. It most often affects the feet, in an asymmetrical distribution. It is associated with a greater degree of radiological damage than occurs in digits not affected by dactylitis. PMID- 15271772 TI - Lower limb arterial incompressibility and obstruction in rheumatoid arthritis. AB - BACKGROUND: Despite increased cardiovascular morbidity and mortality in rheumatoid arthritis, the peripheral arteries remain understudied. OBJECTIVE: To examine the lower limb arteries in age and sex matched, non-smoking subjects with and without rheumatoid arthritis. METHODS: The ankle-brachial index (ABI) was measured at the posterior tibial and dorsal pedal arteries. Arteries were classified as obstructed with ABI < or =0.9, normal with ABI >0.9 but < or =1.3, and incompressible with ABI >1.3. Multinomial logistic regression was used to estimate differences in ABI between patients and controls, adjusting for cardiovascular risk factors, rheumatoid arthritis manifestations, inflammation markers, and glucocorticoid dose. RESULTS: 234 patients with rheumatoid arthritis and 102 controls were studied. Among the rheumatoid patients, 66 of 931 arteries (7%) were incompressible and 30 (3%) were obstructed. Among the controls, three of 408 arteries (0.7%) were incompressible (p = 0.002) and four (1%) were obstructed (p = 0.06). At the person level, one or more abnormal arteries occurred among 45 rheumatoid patients (19%), v five controls (5%, p = 0.001). The greater frequency of arterial incompressibility and obstruction in rheumatoid arthritis was independent of age, sex, and cardiovascular risk factors. Adjustment for inflammation markers, joint damage, rheumatoid factor, and glucocorticoid use reduced rheumatoid arthritis v control differences. Most arterial impairments occurred in rheumatoid patients with 20 or more deformed joints. This subgroup had more incompressible (15%, p< or =0.001) and obstructed arteries (6%, p = 0.005) than the controls, independent of covariates. CONCLUSIONS: Peripheral arterial incompressibility and obstruction are increased in rheumatoid arthritis. Their propensity for patients with advanced joint damage suggests shared pathogenic mechanisms. PMID- 15271773 TI - Large differences in cost of illness and wellbeing between patients with fibromyalgia, chronic low back pain, or ankylosing spondylitis. AB - OBJECTIVE: To compare the cost of illness of three musculoskeletal conditions in relation to general wellbeing. METHODS: Patients with fibromyalgia, chronic low back pain (CLBP), and ankylosing spondylitis who were referred to a specialist and participated in three randomised trials completed a cost diary for the duration of the study, comprising direct medical and non-medical resource utilisation and inability to perform paid and unpaid work. Patients rated perceived wellbeing (0-100) at baseline. Univariate differences in costs between the groups were estimated by bootstrapping. Regression analyses assessed which variables, in addition to the condition, contributed to costs and wellbeing. RESULTS: 70 patients with fibromyalgia, 110 with chronic low back pain, and 111 with ankylosing spondylitis provided data for the cost analyses. Average annual disease related total societal costs per patient were 7813 euro for fibromyalgia, 8533 euro for CLBP, and 3205 euro for ankylosing spondylitis. Total costs were higher for fibromyalgia and CLBP than for ankylosing spondylitis, mainly because of cost of formal and informal care, aids and adaptations, and work days lost. Wellbeing was lower in fibromyalgia (mean, 48) and low back pain (mean, 42) than in ankylosing spondylitis (mean, 67). No variables other than diagnostic group contributed to differences in costs or wellbeing. CONCLUSIONS: In patients under the care of a specialist, there were marked differences in costs and wellbeing between those with fibromyalgia or CLBP and those with ankylosing spondylitis. In particular, direct non-medical costs and productivity costs were higher in fibromyalgia and CLBP. PMID- 15271774 TI - Molecular systematics of Boraginaceae tribe Boragineae based on ITS1 and trnL sequences, with special reference to Anchusa s.l. AB - BACKGROUND AND AIMS: Boragineae is one of the main tribes of Boraginaceae, but delimitation and intergeneric classification of this group are unclear and have not yet been studied using DNA sequences. In particular, phylogenetic relationships in Anchusa s.l. still need to be elucidated in order to assess its taxonomic boundaries with respect to the controversial segregate genera Hormuzakia, Gastrocotyle, Phyllocara and Cynoglottis. METHODS: Phylogenetic relationships among 51 taxa of tribe Boragineae were investigated by comparative sequencing of the trnL(UAA) intron of the plastid genome and of the ITS1 region of the nuclear ribosomal DNA. Exemplar taxa from 16 genera of Boragineae and all subgenera of Anchusa s.l. were included, along with two selected outgroups from tribes Lithospermeae and Cynoglosseae. KEY RESULTS: Phylogenies generated by maximum parsimony and combined ITS1-trnL sequences support the monophyly of the tribe and a split into two clades, Pentaglottis and the remainder of Boragineae. The latter contains two large monophyletic groups. The first consists of three moderately to well-supported branches, Borago-Symphytum, Pulmonaria-Nonea and Brunnera. In the Pulmonaria-Nonea subclade, the rare endemic Paraskevia cesatiana is sister to Pulmonaria, and Nonea appears to be paraphyletic with respect to Elizaldia. The second main group corresponds to the well-supported clade of Anchusa s.l., with the megaphyllic, polyploid herb Trachystemon orientalis as sister taxon, although with low support. Anchusa s.l. is highly paraphyletic to its segregate genera and falls into four subclades: (1) Phyllocara, Hormuzakia, Anchusa subgenus Buglossum and A. subgenus Buglossoides; (2) Gastrocotyle; (3) A. subgenus Buglossellum and Cynoglottis; and (4) A. subgenus Anchusa, Lycopsis and Anchusella. All species of Anchusa subg. Anchusa, including the South African A. capensis, are included in a single unresolved clade. Anchusa subgenus Limbata is also included here despite marked divergence in floral morphology. The low nucleotide variation of ITS1 suggests a recent partly adaptive radiation within this group. CONCLUSIONS: Molecular data show that nine of the usually accepted genera of the Boragineae consisting of two or more species are monophyletic: Anchusella, Borago, Brunnera, Cynoglottis, Gastrocotyle, Hormuzakia, Nonea, Pulmonaria and Symphytum. In addition, the tribe includes the four monotypic genera Paraskevia, Pentaglottis, Phyllocara and Trachystemon. The morphologically well-characterized segregate genera in Anchusa s.l. are all confirmed by DNA sequences and should be definitively accepted. Most of the traditionally recognized subgenera of Anchusa are also supported as monophyletic groups by both nuclear and plastid sequence data. In order to bring taxonomy in line with phylogeny, the institution of new, independent generic entities for subgenera Buglossum, Buglossellum and Buglossoides and a narrower but more natural concept of Anchusa are advocated. PMID- 15271775 TI - Influence of initial organic N reserves and residual leaf area on growth, N uptake, N partitioning and N storage in alfalfa (Medicago sativa) during post cutting regrowth. AB - BACKGROUND AND AIMS: The influence of initial residual leaf area and initial N reserves on N uptake, final N distribution, and yield in alfalfa regrowing after cutting, were studied. METHODS: The effects of two levels of initial residual leaf area (plants cut to 15 cm, with (L+) or without (L-) their leaves) and two initial levels of N status [high N (HN) or low N (LN)] on growth, N uptake and N partitioning, allocation and storage after 29 d of post-cutting regrowth were analysed. KEY RESULTS: During most of the regrowth period (8-29 d after the initial harvest), HN and L+ plants had higher net N uptake rates than LN and L- plants, respectively, resulting in a greater final mineral N uptake for these treatments. However, the final partitioning of exogenous N to the regrowing shoots was the same for all treatments (67 % of total exogenous N on average). Final shoot growth, total plant N content, and N allocation to the different taproot N pools were significantly lower in plants with reduced initial leaf area and initial N reserve status. CONCLUSIONS: Although both initial residual leaf area and initial N reserves influenced alfalfa regrowth, the residual leaf area had a greater effect on final forage production and N composition in the taproot, whereas the N uptake rate and final total N content in plant were more affected by the initial N reserve status than by the residual leaf area. Moreover, N storage as proteins (especially as vegetative storage proteins, rather than nitrate or amino acids) in the taproot allowed nitrate uptake to occur at significant rates. This suggests that protein storage is not only a means of sequestering N in a tissue for further mobilization, utilization for growth or tissue maintenance, but may also indirectly influence both N acquisition and reduction capacities. PMID- 15271776 TI - EC_oligos: automated and whole-genome primer design for exons within one or between two genomes. AB - SUMMARY: EC_oligos designs oligonucleotides (oligos) from exons of annotated genomic sequence information. It can automatically and rapidly select oligos that are conserved between two sets of sequence data, and can pair up oligos for use as PCR primers. It can do this on a whole-genome scale and according to user defined criteria. AVAILABILITY: The source code, executable program and user manual are available at ftp://ftp.ebi.ac.uk/pub/software/dos/EC_oligos/. PMID- 15271777 TI - SelSim: a program to simulate population genetic data with natural selection and recombination. AB - SelSim is a program for Monte Carlo simulation of DNA polymorphism data for a recombining region within which a single bi-allelic site has experienced natural selection. SelSim allows simulation from either a fully stochastic model of, or deterministic approximations to, natural selection within a coalescent framework. A number of different mutation models are available for simulating surrounding neutral variation. The package enables a detailed exploration of the effects of different models and strengths of selection on patterns of diversity. This provides a tool for the statistical analysis of both empirical data and methods designed to detect natural selection. AVAILABILITY: http://www.stats.ox.ac.uk/mathgen/software.html. SUPPLEMENTARY INFORMATION: http://www.stats.ox.ac.uk/mathgen/software.html. PMID- 15271778 TI - Microarray Data Analysis Toolbox (MDAT): for normalization, adjustment and analysis of gene expression data. AB - SUMMARY: We introduce a novel Matlab toolbox for microarray data analysis. This toolbox uses normalization based upon a normally distributed background and differential gene expression based on five statistical measures. The objects in this toolbox are open source and can be implemented to suit your application. AVAILABILITY: MDAT v1.0 is a Matlab toolbox and requires Matlab to run. MDAT is freely available at http://microarray.omrf.org/publications/2004/knowlton/MDAT.zip. PMID- 15271779 TI - Data exploration tools for the Gene Ontology database. AB - MOTIVATION: To improve the ability of biologists (both researchers and students) to ask biologically interesting questions of the Gene Ontology (GO) database and to explore the ontologies by seeing large portions of the ontology graphs in context, along with details of individual terms in the ontologies. RESULTS: GoGet and GoView are two new tools built as part of an extensible web application system based on Java 2 Enterprise Edition technology. GoGet has a user interface that enables users to ask biologically interesting questions, such as (1) What are the DNA binding proteins involved in DNA repair, but not in DNA replication? and (2) Of the terms containing the word triphosphatase, which have associated gene products from mouse, but not fruit fly? The results of such queries can be viewed in a collapsed tabular format that eases the burden of getting through large tables of data. GoView enables users to explore the large directed acyclic graph structure of the ontologies in the GO database. The two tools are coordinated, so that results from queries in GoGet can be visualized in GoView in the ontology in which they appear, and explorations started from GoView can request details of gene product associations to appear in a result table in GoGet. AVAILABILITY: Free access to the GoGet query tool and free download of the GoView ontology viewer are provided to all users at http://db.math.macalester.edu/goproject. In addition, source code for the GoView tool is also available from this site, along with a user manual for both tools. PMID- 15271780 TI - A probabilistic measure for alignment-free sequence comparison. AB - MOTIVATION: Alignment-free sequence comparison methods are still in the early stages of development compared to those of alignment-based sequence analysis. In this paper, we introduce a probabilistic measure of similarity between two biological sequences without alignment. The method is based on the concept of comparing the similarity/dissimilarity between two constructed Markov models. RESULTS: The method was tested against six DNA sequences, which are the thrA, thrB and thrC genes of the threonine operons from Escherichia coli K-12 and from Shigella flexneri; and one random sequence having the same base composition as thrA from E.coli. These results were compared with those obtained from CLUSTAL W algorithm (alignment-based) and the chaos game representation (alignment-free). The method was further tested against a more complex set of 40 DNA sequences and compared with other existing sequence similarity measures (alignment-free). AVAILABILITY: All datasets and computer codes written in MATLAB are available upon request from the first author. PMID- 15271781 TI - ORFcurator: molecular curation of genes and gene clusters in prokaryotic organisms. AB - The ability to detect clusters of functionally related genes in multiple microbial genomes has enormous potential for enhancing studies on gene function and microbial evolution. The staggering amount of new genome sequence data presents a largely untapped resource for gene cluster discovery. To date, gene cluster analysis has not been fully automated, and one must rely on manual, tedious and time-consuming manipulation of sequences. To facilitate accurate and rapid identification of conserved gene clusters, we developed a database-driven web application, called ORFcurator. We used ORFcurator to find clusters containing any genes similar to those of the 14-gene Widespread Colonization Island of Actinobacillus actinomycetemcomitans. From 126 genomes, ORFcurator identified all 73 clusters previously determined by manual searching. AVAILABILITY: ORFcurator and all associated scripts are freely available as supplementary information. SUPPLEMENTARY INFORMATION: http://www.genomecurator.org/ORFcurator/ PMID- 15271782 TI - GeneNetwork: an interactive tool for reconstruction of genetic networks using microarray data. AB - Inferring genetic network architecture from time series data generated from high throughput experimental technologies, such as cDNA microarray, can help us to understand the system behavior of living organisms. We have developed an interactive tool, GeneNetwork, which provides four reverse engineering models and three data interpolation approaches to infer relationships between genes. GeneNetwork enables a user to readily reconstruct genetic networks based on microarray data without having intimate knowledge of the mathematical models. A simple graphical user interface enables rapid, intuitive mapping and analysis of the reconstructed network allowing biologists to explore gene relationships at the system level. AVAILABILITY: Download from http://genenetwork.sbl.bc.sinica.edu.tw/. SUPPLEMENTARY INFORMATION: Supplement documentation of algorithms for the four approaches is downloadable at the above location. PMID- 15271783 TI - ViTO: tool for refinement of protein sequence-structure alignments. AB - ViTO is a graphical application, including an editor, of multiple sequence alignment and a three-dimensional (3D) structure viewer. It is possible to manipulate alignments containing hundreds of sequences and to display a dozen structures. ViTO can handle so-called 'multiparts' alignments to allow the visualization of complex structures (multi-chain proteins and/or small molecules and DNA) and the editing of the corresponding alignment. The 3D viewer and the alignment editor are connected together allowing rapid refinement of sequence structure alignment by taking advantage of the immediate visualization of resulting insertions/deletions and strict conservations in their structural context. More generally, it allows the mapping of informations about the sequence conservation extracted from the alignment onto the 3D structures in a dynamic way. ViTO is also connected to two comparative modelling programs, SCWRL and MODELLER. These features make ViTO a powerful tool to characterize protein families and to optimize the alignments for comparative modelling. AVAILABILITY: http://bioserv.cbs.cnrs.fr/VITO/DOC/. SUPPLEMENTARY INFORMATION: http://bioserv.cbs.cnrs.fr/VITO/DOC/index.html. PMID- 15271784 TI - Systematic analysis of snake neurotoxins' functional classification using a data warehousing approach. AB - MOTIVATION: Sequence annotations, functional and structural data on snake venom neurotoxins (svNTXs) are scattered across multiple databases and literature sources. Sequence annotations and structural data are available in the public molecular databases, while functional data are almost exclusively available in the published articles. There is a need for a specialized svNTXs database that contains NTX entries, which are organized, well annotated and classified in a systematic manner. RESULTS: We have systematically analyzed svNTXs and classified them using structure-function groups based on their structural, functional and phylogenetic properties. Using conserved motifs in each phylogenetic group, we built an intelligent module for the prediction of structural and functional properties of unknown NTXs. We also developed an annotation tool to aid the functional prediction of newly identified NTXs as an additional resource for the venom research community. AVAILABILITY: We created a searchable online database of NTX proteins sequences (http://research.i2r.a star.edu.sg/Templar/DB/snake_neurotoxin). This database can also be found under Swiss-Prot Toxin Annotation Project website (http://www.expasy.org/sprot/). PMID- 15271785 TI - PCOAT: positional correlation analysis using multiple methods. AB - PCOAT (Positional COrrelation Analysis Tool) is a program to perform positional correlation analysis for protein multiple sequence alignment in order to identify structurally or functionally important interactions between positions in a protein family. We implement different statistical methods to detect highly correlated position pairs, amino acid pairs, individual positions and networks of correlated positions, and utilize multiple sequence weighting and sampling methods to eliminate background correlations caused by phylogeny and stochastic events. Our program runs relatively fast and is suitable for analyzing alignments containing large number of sequences. AVAILABILITY: ftp://iole.swmed.edu/pub/PCOAT/. SUPPLEMENTARY INFORMATION: The PCOAT ftp site contains a detailed description of the program, and the results of PCOAT analysis on C2H2 alignment and ACT domain alignment. PMID- 15271786 TI - Effect of exposure to cigarette smoke on carotid artery intimal thickening: the role of inducible NO synthase. AB - OBJECTIVE: We investigated the role of inducible NO synthase (iNOS) in intimal thickening with exposure to cigarette smoke (CS). METHODS AND RESULTS: Intimal thickening in wild-type (WT) and iNOS-deficient (iNOS-/-) mice subjected to CS exposure was induced by placement of a cuff around the carotid artery. CS exposure in WT mice was associated with increased arterial iNOS expression, superoxide production, activator protein-1 (AP-1) activation, and serum NO. Intimal thickening 21 days after cuff placement was significantly greater in mice exposed to CS compared with air (0.023+/-0.013 mm(2) versus 0.009+/-0.008 mm(2); P<0.05). iNOS inhibitor mercaptoethylguanidine-treated WT mice exposed to CS had reduced iNOS activity and intimal thickening (0.006+/-0.005 mm(2); P<0.05). Intimal thickening was significantly less in iNOS-/- mice compared with WT mice (0.006+/-0.005 mm(2); P<0.01) and was not augmented with CS (0.002+/-0.002 mm(2)). The aryl hydrocarbon receptor (AhR) was detected in arteries in vivo and in smooth muscle cells (SMCs) in vitro. CS condensate treatment of SMCs increased AhR binding to the core xenobiotic-responsive element of the iNOS promoter and increased iNOS expression. CONCLUSIONS: Increased arterial expression of iNOS, mediated at least in part by AhR signaling, may be an important mechanism by which CS increases carotid intimal thickening. CS exposure in mice was associated with increased arterial iNOS expression, superoxide production, AP-1 activation, serum NO expression, and intimal thickening. Inhibition or deletion of iNOS abrogated the effects of CS. PMID- 15271787 TI - Angiotensin II-induced insulin resistance and protein tyrosine phosphatases. AB - Although the importance of protein tyrosine phosphorylation by tyrosine kinases in mitogenic signaling is well-accepted, recent studies also suggest that tyrosine dephosphorylation by protein tyrosine phosphatases (PTPases) play an equally important role. For example, both angiotensin II (Ang II) and insulin are known to mediate protein tyrosine phosphorylation and dephosphorylation events. These apparently paradoxical effects of Ang II and insulin suggest that both convergent and divergent intracellular signaling cascades are stimulated downstream of their respective receptors, producing diverse cellular responses. In this review, we discuss the hypothesis that the protein tyrosine phosphatase (PTPase), PTP-1B, plays a central role in Ang II-induced insulin resistance by inhibiting activation of the insulin receptor. We hypothesize that Ang II-induced PTP-1B activation leads to dephosphorylation of the insulin receptor and that this signaling pathway underlies the maladaptive responses observed in diabetic vascular and renal tissue during type II diabetes. PMID- 15271788 TI - Proinflammatory cytokines regulate LOX-1 expression in vascular smooth muscle cells. AB - OBJECTIVE: Atherogenesis represents a type of chronic inflammation and involves elements of the immune response, eg, the expression of proinflammatory cytokines. In advanced atherosclerotic lesions, lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) is expressed in endothelial cells, macrophages, and smooth muscle cells (SMCs). In vitro, the expression of LOX-1 is induced by inflammatory cytokines like TNF-alpha and transforming growth factor (TGF)-beta. Therefore, LOX-1 is thought to be upregulated locally in response to cytokines in vivo. METHODS AND RESULTS: We determined by reverse-transcription polymerase chain reaction (PCR) and Western blot analysis whether the mediators of the acute phase response in inflammation, IL-1alpha, IL-1beta, and TNF-alpha, regulate LOX-1 expression in cultured SMC, and whether this regulation is influenced by peroxisome proliferator-activated receptor gamma (PPARgamma). We studied by immunohistochemistry whether these cytokines are spatially correlated with LOX-1 expression in advanced atherosclerotic lesions. We found upregulation of LOX-1 expression in SMC in a dose- and time-dependent manner after incubation with IL 1alpha, IL-1beta, and TNF-alpha. Simultaneous incubation with these cytokines at saturated concentrations had an additive effect on LOX-1 expression. The PPARgamma activator, 15d-PGJ(2), however, inhibited IL-1beta-induced upregulation of LOX-1. In the intima of atherosclerotic lesions regions of IL-1alpha, IL 1beta, and TNF-alpha expression corresponded to regions of LOX-1 expression. CONCLUSIONS: We suppose that upregulated LOX-1 expression in SMC of advanced atherosclerotic lesions is a response to these proinflammatory cytokines. Moreover, the proinflammatory effects of these cytokines can be decreased by the antiinflammatory effect of PPARgamma. PMID- 15271789 TI - Leukotriene B4 strongly increases monocyte chemoattractant protein-1 in human monocytes. AB - OBJECTIVE: Leukotriene B4 (LTB4), a product of the 5-lipoxygenase (5-LO) pathway of arachidonic acid metabolism, has been implicated in atherosclerosis. However, the molecular mechanisms for the atherogenic effect of LTB4 are not well understood. This study is to determine candidate mechanisms. METHOD AND RESULTS: Primary human monocytes were treated with LTB4 and the supernatant was analyzed for cytokine/chemokine production by an immuno-protein array. This analysis revealed a strong increase of the monocyte chemoattractant protein-1 (MCP-1), a proinflammatory cytokine. Follow-up analyses with MCP-1 enzyme-linked immunosorbent assay (for quantitation of MCP-1 protein) and real-time polymerase chain reaction (PCR) (for MCP-1 mRNA) demonstrated that LTB4 strongly induced expression of MCP-1 protein and mRNA in a time-dependent and dose-dependent fashion. This induction was effectively abolished by CP-105,696, an antagonist for the LTB4 receptor BLT1. Selective inhibitors of ERK1/2 or JNK MAPK effectively blocked the LTB4-induced MCP-1 production. Furthermore, LTB4 increased NF-[kappa]B DNA binding activity, which was blocked by CP-105,696. CONCLUSIONS: LTB4 strongly induces MCP-1 production in primary human monocytes. This induction is mediated through the BLT1 pathway increasing MCP-1 transcription. Activation of ERK1/2 or JNK MAPK is essential for this induction. The NF-[kappa]B activation may be involved in LTB4-increased MCP-1 expression. The LTB4-induced MCP-1 in human monocytes may play a critical role in the atherogenicity of LTB4. PMID- 15271790 TI - C-reactive protein genotypes affect baseline, but not exercise training-induced changes, in C-reactive protein levels. AB - OBJECTIVE: The goal of this study is to determine whether C-reactive protein (CRP) gene variants affect baseline and training-induced changes in plasma CRP levels. METHODS AND RESULTS: Sixty-three sedentary men and women aged 50 to 75 years old underwent baseline testing (Vomax, body composition, CRP levels). They repeated these tests after 24 weeks of exercise training while on a low-fat diet. The CRP +219G/A variant significantly associated with CRP levels before and after training after accounting for the effects of demographic and biological variables. CRP -732A/G genotype was significantly related on a univariate basis to CRP levels after training. The CRP +29T/A variant did not affect CRP levels before or after training. In regression analyses, the +219 and -732 variants each had significant effects on CRP levels before and after training. Subjects homozygous for the common A/G -732/+219 haplotype exhibited the highest CRP levels, and having the rare allele at either site was associated with significantly lower CRP levels. CRP levels decreased significantly with training (-0.38+/-0.18 mg/L; P=0.03). However, none of the CRP variants was associated with the training-induced CRP changes. CONCLUSIONS: CRP +219G/A and -732A/G genotypes and haplotypes and exercise training appear to modulate CRP levels. However, training-induced CRP reductions appear to be independent of genotype at these loci. PMID- 15271791 TI - The transcriptionally active form of AML1 is required for hematopoietic rescue of the AML1-deficient embryonic para-aortic splanchnopleural (P-Sp) region. AB - Acute myelogenous leukemia 1 (AML1; runt-related transcription factor 1 [Runx1]) is a member of Runx transcription factors and is essential for definitive hematopoiesis. Although AML1 possesses several subdomains of defined biochemical functions, the physiologic relevance of each subdomain to hematopoietic development has been poorly understood. Recently, the consequence of carboxy terminal truncation in AML1 was analyzed by the hematopoietic rescue assay of AML1-deficient mouse embryonic stem cells using the gene knock-in approach. Nonetheless, a role for specific internal domains, as well as for mutations found in a human disease, of AML1 remains to be elucidated. In this study, we established an experimental system to efficiently evaluate the hematopoietic potential of AML1 using a coculture system of the murine embryonic para-aortic splanchnopleural (P-Sp) region with a stromal cell line, OP9. In this system, the hematopoietic defect of AML1-deficient P-Sp can be rescued by expressing AML1 with retroviral infection. By analysis of AML1 mutants, we demonstrated that the hematopoietic potential of AML1 was closely related to its transcriptional activity. Furthermore, we showed that other Runx transcription factors, Runx2/AML3 or Runx3/AML2, could rescue the hematopoietic defect of AML1-deficient P-Sp. Thus, this experimental system will become a valuable tool to analyze the physiologic function and domain contribution of Runx proteins in hematopoiesis. PMID- 15271792 TI - KSHV viral cyclin binds to p27KIP1 in primary effusion lymphomas. AB - Primary effusion lymphomas (PELs) represent a unique non-Hodgkin lymphoma that is consistently infected by Kaposi sarcoma herpesvirus (KSHV). PEL cells express high levels of the cell cycle inhibitor p27(KIP1) and yet proliferate actively. KSHV genome encodes a viral cyclin homolog, v-cyclin, which has previously been implicated in down-regulation of p27(KIP1) levels. To address how PEL cells can tolerate high p27(KIP1) levels, we investigated functional interactions between v cyclin and p27(KIP1) using PEL-derived cell lines as a model system. Here we demonstrate that v-cyclin and p27(KIP1) stably associate in PEL cells in vivo suggesting an attractive model by which p27(KIP1) is inactivated in the actively proliferating PEL cells. Moreover, we show that v-cyclin and cyclin-dependent kinase 6 (CDK6) form an active kinase without p27(KIP1) and that CDK6 is the in vivo catalytic subunit of v-cyclin in PEL cells. These findings suggest that KSHV may promote oncogenesis in PEL by expressing v-cyclin, which both overrides negative cell cycle controls present in the PEL precursor cells and induces a strong proliferative signal via CDK6 kinase activity. PMID- 15271793 TI - Gene expression profiling of normal and malignant CD34-derived megakaryocytic cells. AB - Gene expression profiles of bone marrow (BM) CD34-derived megakaryocytic cells (MKs) were compared in patients with essential thrombocythemia (ET) and healthy subjects using oligonucleotide microarray analysis to identify differentially expressed genes and disease-specific transcripts. We found that proapoptotic genes such as BAX, BNIP3, and BNIP3L were down-regulated in ET MKs together with genes that are components of the mitochondrial permeability transition pore complex, a system with a pivotal role in apoptosis. Conversely, antiapoptotic genes such as IGF1-R and CFLAR were up-regulated in the malignant cells, as was the SDF1 gene, which favors cell survival. On the basis of the array results, we characterized apoptosis of normal and ET MKs by time-course evaluation of annexin V and sub-G1 peak DNA stainings of immature and mature MKs after culture in serum free medium with an optimal thrombopoietin concentration, and annexin-V-positive MKs only, with decreasing thrombopoietin concentrations. ET MKs were more resistant to apoptosis than their normal counterparts. We conclude that imbalance between proliferation and apoptosis seems to be an important step in malignant ET megakaryocytopoiesis. PMID- 15271794 TI - Human immunodeficiency virus-driven expansion of CD4+CD25+ regulatory T cells, which suppress HIV-specific CD4 T-cell responses in HIV-infected patients. AB - The present study demonstrates that CD4(+)CD25(+) T cells, expanded in peripheral blood of HIV-infected patients receiving highly active antiretroviral therapy (HAART), exhibit phenotypic, molecular, and functional characteristics of regulatory T cells. The majority of peripheral CD4(+)CD25(+) T cells from HIV infected patients expressed a memory phenotype. They were found to constitutively express transcription factor forkhead box P3 (Foxp3) messengers. CD4(+)CD25(+) T cells weakly proliferated to immobilized anti-CD3 monoclonal antibody (mAb) and addition of soluble anti-CD28 mAb significantly increased proliferation. In contrast to CD4(+)CD25(-) T cells, CD4(+)CD25(+) T cells from HIV-infected patients did not proliferate in response to recall antigens and to p24 protein. The proliferative capacity of CD4 T cells to tuberculin, cytomegalovirus (CMV), and p24 significantly increased following depletion of CD4(+)CD25(+) T cells. Furthermore, addition of increasing numbers of CD4(+)CD25(+) T cells resulted in a dose-dependent inhibition of CD4(+)CD25(-) T-cell proliferation to tuberculin and p24. CD4(+)CD25(+) T cells responded specifically to p24 antigen stimulation by expressing transforming growth factor beta (TGF-beta) and interleukin 10 (IL 10), thus indicating the presence of p24-specific CD4(+) T cells among the CD4(+)CD25(+) T-cell subset. Suppressive activity was not dependent on the secretion of TGF-beta or IL-10. Taken together, our results suggest that persistence of HIV antigens might trigger the expansion of CD4(+)CD25(+) regulatory T cells, which might induce a tolerance to HIV in vivo. PMID- 15271795 TI - Megakaryocyte proliferation and ploidy regulated by the cytoplasmic tail of glycoprotein Ibalpha. AB - We have investigated the ability of glycoprotein (GP) Ibalpha, a megakaryocytic gene product, to sequester the signal transduction protein 14-3-3xi and to influence megakaryocytopoiesis. Using a Gp1ba(-/-) mouse colony, we compared the rescued phenotypes produced by a wild-type human GP Ibalpha allele or a similar allele containing a 6-residue cytoplasmic tail truncation that abrogates binding to 14-3-3xi. The observed phenotypes illustrate an involvement for GP Ibalpha in thrombopoietin-mediated events of megakaryocyte proliferation, polyploidization, and the expression of apoptotic markers in maturing megakaryocytes. We developed a hypothesis for the involvement of a GP Ibalpha/14-3-3xi/PI-3 kinase complex in regulating thrombopoietin-mediated responses. An observed increase in thrombopoietin-mediated Akt phosphorylation in the truncated variant supported the hypothesis and led to the development of a model in which the GP Ibalpha cytoplasmic tail sequestered signaling proteins during megakaryocytopoiesis and, as such, became a critical regulator in the temporal sequence of events that led to normal megakaryocyte maturation. PMID- 15271796 TI - PDGF-D induces macrophage recruitment, increased interstitial pressure, and blood vessel maturation during angiogenesis. AB - Platelet-derived growth factor-D (PDGF-D) is a recently characterized member of the PDGF family with unknown in vivo functions. We investigated the effects of PDGF-D in transgenic mice by expressing it in basal epidermal cells and then analyzed skin histology, interstitial fluid pressure, and wound healing. When compared with control mice, PDGF-D transgenic mice displayed increased numbers of macrophages and elevated interstitial fluid pressure in the dermis. Wound healing in the transgenic mice was characterized by increased cell density and enhanced recruitment of macrophages. Macrophage recruitment was also the characteristic response when PDGF-D was expressed in skeletal muscle or ear by an adeno associated virus vector. Combined expression of PDGF-D with vascular endothelial growth factor-E (VEGF-E) led to increased pericyte/smooth muscle cell coating of the VEGF-E-induced vessels and inhibition of the vascular leakiness that accompanies VEGF-E-induced angiogenesis. These results show that full-length PDGF D is activated in tissues and is capable of increasing interstitial fluid pressure and macrophage recruitment and the maturation of blood vessels in angiogenic processes. PMID- 15271797 TI - NADPH oxidase mediates vascular endothelial cadherin phosphorylation and endothelial dysfunction. AB - Vascular endothelial activation is an early step during leukocyte/endothelial adhesion and transendothelial leukocyte migration in inflammatory states. Leukocyte transmigration occurs through intercellular gaps between endothelial cells. Vascular endothelial cadherin (VE-cadherin) is a predominant component of endothelial adherens junctions that regulates intercellular gap formation. We found that tumor necrosis factor (TNF) caused tyrosine phosphorylation of VE cadherin, separation of lateral cell-cell junctions, and intercellular gap formation in human umbilical vein endothelial cell (HUVEC) monolayers. These events appear to be regulated by intracellular oxidant production through endothelial NAD(P)H (nicotinamide adenine dinucleotide phosphate) oxidase because antioxidants and expression of a transdominant inhibitor of the NADPH oxidase, p67(V204A), effectively blocked the effects of TNF on all 3 parameters of junctional integrity. Antioxidants and p67(V204A) also decreased TNF-induced JNK activation. Dominant-negative JNK abrogated VE-cadherin phosphorylation and junctional separation, suggesting a downstream role for JNK. Finally, adenoviral delivery of the kinase dead PAK1(K298A) decreased TNF-induced JNK activation, VE cadherin phosphorylation, and lateral junctional separation, consistent with the proposed involvement of PAK1 upstream of the NADPH oxidase. Thus, PAK-1 acts in concert with oxidase during TNF-induced oxidant production and loss of endothelial cell junctional integrity. PMID- 15271798 TI - The contribution of endothelial cell P-selectin to the microvascular flow of mouse sickle erythrocytes in vivo. AB - Microvascular occlusion in sickle cell disease can be initiated by adhesion of sickle red blood cells (RBCs) to the endothelium. Our objective in this study was to verify the relevance in vivo of our discovery that sickle RBCs adhere abnormally to endothelial P-selectin in vitro. We used computer-assisted intravital microscopy to characterize RBC flow velocity (V(RBC)) in mice. We found faster V(RBC) of sickle RBCs in P-selectin knock-out and control mice than in sickle cell mice, which have increased endothelial cell P-selectin expression. Agonist peptide for murine protease-activated receptor-1 (PAR-1), which selectively activates mouse endothelial cells but not platelets, was used to assess the effects of endothelial cell P-selectin on microvascular flow. Suffusion of venules with this agonist stopped flow promptly in normal and sickle mice but not in P-selectin knock-out mice or in control mice pretreated with anti P-selectin monoclonal antibody or unfractionated heparin (UFH). Agonist-induced slowing of flow was reversed rapidly by suffusion with UFH, provided flow had not already stopped. We conclude that endothelial cell P-selectin contributes to the microcirculatory abnormalities in sickle cell disease and that blocking P selectin may be useful for preventing painful vasoocclusion in sickle cell disease. PMID- 15271799 TI - G6PD is indispensable for erythropoiesis after the embryonic-adult hemoglobin switch. AB - Glucose 6-phosphate dehydrogenase (G6PD) (EC 1.1.1.42) is an essential enzyme for the rapid production of NADPH, as required on exposure to oxidative stress. Mouse embryonic stem (ES) cells can produce all embryonic and fetal/adult cell types. By studying the in vitro differentiation of embryoid bodies produced from G6pdDelta ES cells that are totally unable to produce G6PD protein, we found that these cells are able to differentiate into mesodermal cells, cardiomyocytes, hepatocytes, and primitive erythroid cells. However, we show here that, after the hemoglobin switch has taken place, definitive erythrocytes die by apoptosis. This apoptotic death is delayed by reducing agents and by a caspase inhibitor, but it is prevented only by the restoration of G6PD activity. Thus, G6PD proves indispensable for definitive erythropoiesis. PMID- 15271800 TI - Interleukin-2-induced survival of natural killer (NK) cells involving phosphatidylinositol-3 kinase-dependent reduction of ceramide through acid sphingomyelinase, sphingomyelin synthase, and glucosylceramide synthase. AB - Interleukin 2 (IL-2) rescued human natural killer (NK) KHYG-1 cells from apoptosis along with a reduction of ceramide. Conversely, an increase of ceramide inhibited IL-2-rescued survival. IL-2 deprivation-induced activation of acid sphingomyelinase (SMase) and inhibition of glucosylceramide synthase (GCS) and sphingomyelin synthase (SMS) were normalized by IL-2 supplementation. A phosphatidyl inositol-3 (PI-3) kinase inhibitor, LY294002, inhibited IL-2-rescued survival, but a mitogen-activated protein kinase inhibitor, PD98059, and an inhibitor of Janus tyrosine kinase/signal transducer and activator of transcription pathway, AG490, did not. LY294002 inhibited IL-2-induced reduction of ceramide through activation of acid SMase and inhibition of GCS and SMS, suggesting the positive involvement of PI-3 kinase in ceramide reduction through enzymatic regulation. Indeed, a constitutively active PI-3 kinase enhanced growth rate and ceramide reduction through inhibition of acid SMase and activation of GCS and SMS. Further, LY294002 inhibited IL-2-induced changes of transcriptional level as well as mRNA and protein levels in acid SMase and GCS but did not affect the stability of the mRNAs. These results suggest that PI-3 kinase-dependent reduction of ceramide through regulation of acid SMase, GCS, and SMS plays a role in IL-2-rescued survival of NK cells. PMID- 15271801 TI - Identification of a novel natural regulatory CD8 T-cell subset and analysis of its mechanism of regulation. AB - The immune system contains natural regulatory T cells that control the magnitude of the immune response during physiologic and pathologic conditions. Although this suppressive function was historically attributed to CD8 T cells, most recent reports have focused on natural regulatory CD4 T cells. In the present study, we describe a new subset of natural CD8 regulatory T cells in normal healthy animals. This subset expresses low levels of CD45RC at its surface (CD45RC(low)); produces mainly interleukin-4 (IL-4), IL-10, and IL-13 cytokines upon in vitro stimulation; expresses Foxp3 and cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4); and is not cytotoxic against allogeneic targets. This subset suppresses the proliferation and differentiation of autologous CD4 T cells into type-1 cytokines producing T cells after stimulation with allogeneic accessory cells. We also provide evidence that this regulatory subset mediates its suppression by cell-to-cell contact and not through secretion of suppressive cytokines. Finally, the regulatory activity of CD8 CD45RC(low) cells is also demonstrated in vivo in a rat model of CD4-dependent graft-versus-host disease. Collectively, these data demonstrate for the first time that freshly isolated rat CD8 CD45RC(low) T cells contain T cells with regulatory properties, a result that enlarges the general picture of T-cell-mediated regulation. PMID- 15271802 TI - Visualization of lymphatic vessels through NF-kappaB activity. AB - The molecular biology of lymphatics is only rudimentary owing to the long standing absence of specific markers, and scanty is the information regarding bladder lymphatic vessels. By using mice with a reporter gene for nuclear factor kappaB (NF-kappaB) activity (kappaB-lacZ) in combination with immunohistochemical staining with a specific lymphatic marker (LYVE-1), we show, for the first time, that NF-kappaB is constitutively active in lymphatic endothelium in the urinary bladder, uterus, intestine, heart, and airways. Tie2-lacZ mice confirmed that the structures observed in kappaB-lacZ mice were not blood vessels. In addition, acute instillation of lipopolysaccharide (LPS) or tumor necrosis factor alpha (TNF-alpha) into the kappaB-lacZ mouse bladder revealed the capacity of this transgenic in reporting inducible NF-kappaB activity. Our findings demonstrate an overriding constitutive NF-kappaB activity in the lymphatic system. They also provide a working model for detecting lymphatic vessels and evoke testable hypotheses regarding the role of lymphatic vessels in health and disease. PMID- 15271803 TI - Treating depression in later life. PMID- 15271804 TI - Which drugs should be available over the counter? PMID- 15271805 TI - Social networks and collateral health effects. PMID- 15271806 TI - Condoms and prevention of HIV. PMID- 15271807 TI - UK Health Commission downgrades four foundation trusts. PMID- 15271808 TI - Blair pledges 150m pounds sterling to support and protect AIDS orphans. PMID- 15271810 TI - Shipman inquiry recommends tighter rules on controlled drugs. PMID- 15271811 TI - FDA's counsel accused of being too close to drug industry. PMID- 15271812 TI - Drug companies frame rules to work with NHS. PMID- 15271815 TI - US government to open national bank of "approved" embryonic stem cells. PMID- 15271816 TI - South African doctors charged with involvement in organ trade. PMID- 15271817 TI - Doctor shortage forces leading African children's hospital to refuse emergencies. PMID- 15271818 TI - UN considers sanctions against Sudanese government for obstructing aid. PMID- 15271819 TI - Doctors' group publishes archive of doctors registered in Nazi era. PMID- 15271820 TI - Draft guidance on clinical trials recognises needs of non-commercial research. PMID- 15271821 TI - Inquiry into death of boy from induced illness recommends tightening of child protection rules. PMID- 15271823 TI - Commission invites discussion on the future of genetics in reproduction. PMID- 15271824 TI - Canada accused of failing women patients. PMID- 15271825 TI - MSF reports encouraging results with AIDS treatment. PMID- 15271826 TI - Bush's AIDS plan criticised for emphasising abstinence and forbidding condoms. PMID- 15271828 TI - US doctors and patients are split on approval of generic thyroid hormone. PMID- 15271829 TI - Bush accused of pressuring countries to stop producing generic drugs. PMID- 15271830 TI - Acquisition of Helicobacter pylori infection after outbreaks of gastroenteritis: prospective cohort survey in institutionalised young people. PMID- 15271831 TI - Anterior uveitis is associated with travoprost. PMID- 15271832 TI - Ruling a diagnosis in or out with "SpPIn" and "SnNOut": a note of caution. PMID- 15271833 TI - Sex, sun, sea, and STIs: sexually transmitted infections acquired on holiday. PMID- 15271834 TI - Acute visual loss and pituitary apoplexy after surgery. PMID- 15271835 TI - Intensive care management and control of infection. PMID- 15271836 TI - National Institute for Clinical Excellence and its value judgments. PMID- 15271837 TI - Challenges for the National Institute for Clinical Excellence. PMID- 15271838 TI - Bone scanning in lung cancer: evidence is not sufficient to justify routine bone scanning. PMID- 15271839 TI - Firm foundation for senior house officers: a firm foundation for general practice. PMID- 15271840 TI - Firm foundation for senior house officers: quality assurance programme needs to be in place. PMID- 15271841 TI - Firm foundation for senior house officers: junior doctors should become progressively more enabled. PMID- 15271842 TI - Firm foundation for senior house officers: five years is long enough. PMID- 15271843 TI - Bone scanning in lung cancer: pretest probability is of value. PMID- 15271844 TI - Low back pain: sacroiliac joint pain may be myth. PMID- 15271845 TI - Monitoring procalcitonin is of value in acute pancreatitis. PMID- 15271846 TI - Responsibility for ancillary care in clinical trials: research patrimony and unintended coercion are hazards. PMID- 15271847 TI - Responsibility for ancillary care in clinical trials: finding out what participants think may be way forward. PMID- 15271848 TI - A time to live or a time to die? Are we losing our humanity? PMID- 15271849 TI - A time to live or a time to die?: Let those who have ears hear. PMID- 15271851 TI - Pharmacologic treatment of erectile dysfunction. PMID- 15271852 TI - Screening for abdominal aortic aneurysm. PMID- 15271853 TI - STK15 polymorphism and breast cancer risk in a population-based study. AB - STK15 is considered a potential cancer susceptibility gene owing to its functions in normal cell mitosis. Two common coding region polymorphisms in the gene (F31I and V57I) may affect ubiquitin-dependent degradation and thus the half-life of the encoded protein. There are limited data on the relevance of these polymorphisms to population cancer rates. To examine whether functional variation in STK15 may affect breast cancer risk, we genotyped a large series of incident breast cancer cases (n = 941) and age-matched population controls (n = 830) for the F31I and V57I polymorphisms. Individually, neither the F31I polymorphism [odds ratio (OR) 1.54; 95% confidence interval (CI) 0.96-2.47, comparing 31I with 31F homozygotes] nor the V57I polymorphism (OR 0.92; 95% CI 0.50-1.71, comparing 57I with 57V homozygotes) was significantly associated with breast cancer risk. A relatively common genotype, combining the two polymorphisms (31I-57V/31I-57V, 3% of controls) was related to a significant 2-fold increase in the risk of post menopausal breast cancer (OR 1.96; 95% CI 1.01-3.79). No interaction was detected between STK15 variants and estrogenic risk factors, although the power of these analyses was limited. These results suggest that STK15 may represent a low penetrance type breast cancer susceptibility gene. PMID- 15271854 TI - Curcumin-induced GADD153 gene up-regulation in human colon cancer cells. AB - Ingestion of plant products containing the phenolic phytochemical, curcumin, has been linked to lower incidences of colon cancer, suggesting that curcumin has cancer chemopreventive effects. Supporting this suggestion at the cellular level, apoptosis occurs in human colon cancer cells exposed to curcumin. However, the mechanism is unclear, prompting this investigation to further clarify the molecular effects of curcumin. HCT-116 colonocytes were incubated with 0-20 microM curcumin for 0-48 h. In concentration-dependent and time-dependent manners, curcumin induced DNA damage, resulting later in the appearance of cellular features characteristic of apoptosis. To identify a potential pro apoptotic gene that could be responsive to the DNA damage in curcumin-treated cells, growth arrest and DNA damage-inducible gene 153 (GADD153) was considered. Curcumin increased GADD153 mRNA (and also protein) expression, which was prevented by actinomycin D and also by a broad protein kinase C inhibitor, but not by selective MAPK inhibitors. These findings suggest that curcumin-induced up regulation of GADD153 mRNA expression was at the level of transcription, but apparently without depending on upstream MAPK. In determining the involvement of reactive oxygen species in mediating the effect of curcumin on GADD153, the antioxidants pyrrolidine dithiocarbamate and N-acetylcysteine (NAC), but neither alpha-tocopherol nor catalase, also blunted or prevented up-regulation of GADD153 mRNA expression caused by curcumin. Most noteworthy, when NAC was tested, it inhibited the DNA damage and apoptosis caused by curcumin. Because expression of GADD153 protein was detected before the appearance of apoptotic features, this observation raises the possibility that GADD153 protein might be important for curcumin-induced apoptosis. PMID- 15271855 TI - Altered vegetable intake affects pivotal carcinogenesis pathways in colon mucosa from adenoma patients and controls. AB - The evidence from epidemiological and experimental studies that vegetables reduce the risk of colorectal cancer is convincing. However, the involved genes and genetic pathways are not clear. The aim of this study was to identify genes that are modulated in vivo in colorectal mucosa by vegetables, and to investigate whether colon adenoma patients respond differently compared with healthy controls. Twenty female adenoma patients and eight healthy controls were randomly split into two groups of ten and four persons, respectively, receiving either a 50% decreased (=75 g/day) or doubled (=300 g/day) intake of vegetables for 2 weeks. In order to assess the effects on gene expression at the target level, colorectal biopsies were collected before and after the intervention. Total RNA was isolated from the biopsies to measure gene expression of 597 genes relevant for responses to xenobiotics by microarray technology, followed by confidence analyses to identify differentially expressed genes. Mainly genes related to cell cycle control and genes for oxidoreductase activities were over-represented in the list of modulated genes. Twenty genes were modulated, which are known to be related to (colon)carcinogenesis. Seven genes were similarly modulated in patients and controls, for example fos proto-oncogene and ornithine decarboxylase. Thirteen genes were modulated differently in patients compared with controls, including cyclooxygenase-2 and human mdm2-A in patients and cytochrome P45027B1, -2C19, -2D6, -2C9 and -3A4 in controls. Almost all the effects on modulating the expression of genes by altering vegetable intake can be mechanistically linked to cellular processes that explain either prevention of colorectal cancer risk by high vegetable intake or increased colorectal cancer risk by low vegetable intake. Furthermore, it seems that vegetables in patients affect genes involved in the late stage of colorectal cancer, whereas in controls genes involved in the initiation phase are modulated. PMID- 15271856 TI - Functional Phe31Ile polymorphism in Aurora A and risk of breast carcinoma. AB - Aurora-A/BTAK/STK15, involved in regulating centrosomes and chromosome segregation, is overexpressed in human breast carcinoma and other cancers. The Phe31-->Ile polymorphism in Aurora A alters the kinase function, with the Ile31 variant being preferentially amplified and associated with degree of aneuploidy in human tumors. We have previously shown that the Phe31Ile polymorphism is associated with the occurrence and advanced disease status of esophageal cancer. This case-control study examined the contribution of this polymorphism to susceptibility to development and progression of breast cancer. Aurora A genotypes were determined in 520 patients with breast carcinoma, 191 patients with benign breast diseases (BBD) and 520 controls. It was found that the Aurora A Ile/Ile genotype was significantly associated with increased risk of breast carcinoma occurrence [odds ratio (OR) 1.66; 95% confidence interval (95% CI) 1.29 2.12] compared with the Phe/Phe or Phe/Ile genotype. The increased risk for BBD and breast carcinoma related to the Ile/Ile genotype was more pronounced in younger subjects. Moreover, we found that patients carrying the Ile/Ile genotype tended to have ER-carcinomas (OR 2.56; 95% CI 1.24-5.26). No significant association was observed between the polymorphism and metastasis and disease stage of the cancer. These findings suggest that the Phe31Ile polymorphism in Aurora A may be a genetic modifier for developing breast carcinoma. PMID- 15271857 TI - Vascular endothelial growth factor activation of sterol regulatory element binding protein: a potential role in angiogenesis. AB - By stimulating the migration and proliferation of endothelial cells (ECs), vascular endothelial growth factor (VEGF) is a potent angiogenic factor. However, the molecular mechanism involved in the VEGF-induced angiogenesis remains elusive. We hypothesized that sterol regulatory element binding proteins (SREBPs), transcription factors governing cellular lipid homeostasis, play an important role in regulating angiogenesis in response to VEGF. VEGF activated SREBP1 and SREBP2 in ECs, as demonstrated by the increased SREBPs, their cleavage products, and the upregulation of the targeted genes. VEGF-induced SREBP activation depended on SREBP cleavage-activating protein (SCAP), because knocking down SCAP by RNA interference (RNAi) inhibited SREBP activation in response to VEGF. SREBP activation was also blocked by 25-hydroxycholesterol (25-HC). To verify the functional implication of SREBPs in VEGF-induced angiogenesis, we tested the role of SREBPs in EC migration and proliferation. SCAP RNAi or 25-HC inhibited VEGF-induced pseudopodia extension and migration of ECs. Both treatments inhibited VEGF-induced EC proliferation, with cell growth arrested at the G(0)/G(1) phase and a concomitant decrease of the S phase. Blocking the PI3K Akt pathway inhibited the VEGF-activated SREBPs, demonstrating that PI3K-Akt regulates SREBPs. Consistent with our in vitro data, SREBP1 was detected in newly developed microvasculatures in a rabbit skin partial-thickness wound-healing model. SREBP inhibition also markedly suppressed VEGF-induced angiogenesis in chick embryos. In summary, this study identifies SREBPs as the key molecules in regulating angiogenesis in response to VEGF. PMID- 15271858 TI - Requirement for Rac1-dependent NADPH oxidase in the cardiovascular and dipsogenic actions of angiotensin II in the brain. AB - We have shown that intracellular superoxide (O(2)(*-)) production in CNS neurons plays a key role in the pressor, bradycardic, and dipsogenic actions of Ang II in the brain. In this study, we tested the hypothesis that a Rac1-dependent NADPH oxidase is a key source of O(2)(*-) in Ang II-sensitive neurons and is involved in these central Ang II-dependent effects. We performed both in vitro and in vivo studies using adenoviral (Ad)-mediated expression of dominant-negative Rac1 (AdN17Rac1) to inhibit Ang II-stimulated Rac1 activation, an obligatory step in NADPH oxidase activation. Ang II induced a time-dependent increase in Rac1 activation and O(2)(*-) production in Neuro-2A cells, and this was abolished by pretreatment with AdN17Rac1 or the NADPH oxidase inhibitors apocynin or diphenylene iodonium. AdN17Rac1 also inhibited Ang II-induced increases in NADPH oxidase activity in primary neurons cultured from central cardiovascular control regions. In contrast, overexpression of wild-type Rac1 (AdwtRac1) caused more robust NADPH oxidase-dependent O(2)(*-) production to Ang II. To extend the in vitro studies, the pressor, bradycardic, and drinking responses to intracerebroventricularly (ICV) injected Ang II were measured in mice that had undergone gene transfer of AdN17Rac1 or AdwtRac1 to the brain. AdN17Rac1 abolished the increase in blood pressure, decrease in heart rate, and drinking response induced by ICV injection of Ang II, whereas AdwtRac1 enhanced these physiological effects. The exaggerated physiological responses in AdwtRac1 treated mice were abolished by O(2)(*-) scavenging. These results, for the first time, identify a Rac1-dependent NADPH oxidase as the source of central Ang II induced O(2)(*-) production, and implicate this oxidase in cardiovascular diseases associated with dysregulation of brain Ang II signaling, including hypertension. PMID- 15271859 TI - Kappa free light chains in cerebrospinal fluid as markers of intrathecal immunoglobulin synthesis. AB - BACKGROUND: Intrathecal immunoglobulin synthesis is observed in several inflammatory disorders of the central nervous system, but its detection by current laboratory tests is either tedious or relatively insensitive. We assessed the diagnostic accuracy of an assay for kappa free light chains (kappaFLC) in cerebrospinal fluid (CSF) and serum, and compared it with traditional tests for intrathecal immunoglobulin synthesis. METHODS: kappaFLCs were measured by nephelometry in CSF/serum pairs from 112 patients. Samples were excluded if blood contamination of CSF as a result of traumatic lumbar puncture (n = 12) or monoclonal bands in both CSF and serum (n = 5) were present. The remaining sample pairs were grouped according to the presence (n = 71) or absence (n = 24) of oligoclonal bands. Data were analyzed as kappaFLC concentrations in CSF, as kappaFLC CSF/serum ratios, and by use of the quotient diagram described previously for immunoglobulins. RESULTS: Both kappaFLC concentrations in CSF and the kappaFLC CSF/serum ratio identified patients with oligoclonal bands with high specificity and sensitivity. The areas under the ROC curves were 0.991 (95% confidence interval, 0.944-0.998) and 0.978 (0.924-0.996), respectively. Exclusion of patients with impaired blood-CSF barrier function further improved diagnostic accuracy. To account for patients with impaired blood-CSF barrier function, data were also analyzed in a quotient diagram. Only two patients without detectable oligoclonal bands would have been misclassified by this approach. CONCLUSIONS: Our data indicate that the nephelometric assay for kappaFLCs in CSF reliably detects intrathecal immunoglobulin synthesis. This automated and quantitative method could simplify the diagnostic procedure for CSF analysis in the routine laboratory. PMID- 15271860 TI - Validated gas chromatographic-negative ion chemical ionization mass spectrometric method for delta(9)-tetrahydrocannabinol in sweat patches. AB - BACKGROUND: A sensitive gas chromatography-negative ion chemical ionization mass spectrometry (GC/MS-NICI) method was developed and validated for the measurement of Delta(9)-tetrahydrocannabinol (THC) in human sweat patches. METHODS: THC-d(0) and THC-d(3) were added to worn blank sweat patches (PharmChek; PharmChem Incorporated) and extracted with 3 mL of methanol-0.2 mol/L sodium acetate buffer (pH 5.0, 3:1 by volume) on a reciprocating shaker at ambient temperature for 30 min. Extracted solution (2 mL) was diluted with 8 mL of 0.1 mol/L sodium acetate buffer (pH 4.5) and extracted by use of solid-phase extraction columns (CleanScreen; United Chemical Technologies). Dried extracts were derivatized with trifluoroacetic acid and analyzed with an Agilent 6890 gas chromatograph interfaced with an Agilent 5973 mass selective detector operated in NICI-selected ion-monitoring mode. RESULTS: The lower limits of detection and quantification for THC in human sweat were 0.2 and 0.4 ng/patch, respectively. The calibration curve was linear from 0.4 to 10 ng/patch (R(2) >0.995). Overall recovery of THC from blank worn patches to which 0.6, 4.0, and 8.0 ng of THC had been added was 44-46%. Assay imprecision, expressed as CV, was <10% for 0.6, 4.0, and 8.0 ng/patch quality-control samples. Twenty-one potential interfering compounds (50 ng/patch) added to low quality-control samples (0.6 ng/patch) did not influence THC quantification. CONCLUSIONS: This GC/MS-NICI assay for THC in human sweat provides adequate sensitivity and performance characteristics for analyzing THC in sweat patches and meets the requirements of the proposed Substance Abuse and Mental Health Administration's guidelines for sweat testing. PMID- 15271862 TI - Staying connected without connexin43: can you hear me now? PMID- 15271863 TI - And what about the endothelium? On the predominance of cerebral superoxide formation for angiotensin II-induced systemic hypertension. PMID- 15271864 TI - Myocardial protection at a crossroads: the need for translation into clinical therapy. AB - Over the past 30 years, hundreds of experimental interventions (both pharmacologic and nonpharmacologic) have been reported to protect the ischemic myocardium in experimental animals; however, with the exception of early reperfusion, none has been translated into clinical practice. The National Heart, Lung, and Blood Institute convened a working group to discuss the reasons for the failure to translate potential therapies for protecting the heart from ischemia and reperfusion and to recommend new approaches to accomplish this goal. The Working Group concluded that cardioprotection in the setting of acute myocardial infarction, cardiac surgery, and cardiac arrest is at a crossroads. Present basic research approaches to identify cardioprotective therapies are inefficient and counterproductive. For 3 decades, significant resources have been invested in single-center studies that have often yielded inconclusive results. A new paradigm is needed to obviate many of the difficulties associated with translation of basic science findings. The Working Group urged a new focus on translational research that emphasizes efficacy and clinically relevant outcomes, and recommended the establishment of a system for rigorous preclinical testing of promising cardioprotective agents with clinical trial-like approaches (ie, blinded, randomized, multicenter, and adequately powered studies using standardized methods). A national preclinical research consortium would enable rational translation of important basic science findings into clinical use. The Working Group recommended that the National Institutes of Health proactively intervene to remedy current problems that impede translation of cardioprotective therapies. Their specific recommendations include the establishment of a preclinical consortium and the performance of 2 clinical studies that are likely to demonstrate effectiveness (phase III clinical trials of adenosine in acute myocardial infarction and cardiac surgery). PMID- 15271865 TI - Is the failing heart energy starved? On using chemical energy to support cardiac function. AB - The requirement of chemical energy in the form of ATP to support systolic and diastolic work of the heart is absolute. Because of its central role in cardiac metabolism and performance, the subject of this review on energetics in the failing heart is ATP. We briefly review the basics of myocardial ATP metabolism and describe how this changes in the failing heart. We present an analysis of what is now known about the causes and consequences of these energetic changes and conclude by commenting on unsolved problems and opportunities for future basic and clinical research. PMID- 15271866 TI - Red blood cell-mediated hypoxic vasodilatation: a balanced physiological viewpoint. PMID- 15271867 TI - Reliability of comparative genomic hybridization to detect chromosome abnormalities in first polar bodies and metaphase II oocytes. AB - BACKGROUND: Preimplantation Genetic Diagnosis (PGD) using FISH to analyze up to nine chromosomes to discard chromosomally abnormal embryos has resulted in an increase of pregnancy rates in certain groups of patients. However, the number of chromosomes that can be analyzed is a clear limitation. We evaluate the reliability of using comparative genomic hybridization (CGH) to detect the whole set of chromosomes, as an alternative to PGD using FISH. METHODS AND RESULTS: We have analysed by CGH both, first polar bodies (1PBs) and metaphase II (MII) oocytes from 30 oocytes donated by 24 women. The aneuploidy rate was 48%. Considering two maternal age groups, a higher number of chromosome abnormalities were detected in the older group of oocytes (23% versus 75%, P < 0.02). About 33% of the 1PB-MII oocyte doublets diagnosed as aneuploid by CGH would have been misdiagnosed as normal if FISH with nine chromosome probes had been used. CONCLUSION: We demonstrate the reliability of 1PB analysis by CGH, to detect almost any chromosome abnormality in oocytes as well as unbalanced segregations of maternal translocations in a time frame compatible with regular in vitro fertilization (IVF). The selection of euploid oocytes could help to increase implantation and pregnancy rates of patients undergoing IVF treatment. PMID- 15271868 TI - Half-dose depot triptorelin in pituitary suppression for multiple ovarian stimulation in assisted reproduction technology: a randomized study. AB - BACKGROUND: Pituitary suppression by depot GnRH agonist may be excessive for ovarian stimulation in assisted reproduction technology. This study compares the efficacy of standard and half-dose depot triptorelin in a long protocol. METHODS: A total of 180 patients were randomized into two groups using sealed envelopes. Pituitary desensitization was obtained in group 1 (90 patients) with half-dose (1.87 mg) triptorelin depot in the mid-luteal phase of their menstrual cycle, and in group 2 (90 patients) with full-dose (3.75 mg) triptorelin. RESULTS: There was no premature LH surge, with LH levels being lower in the full-dose group (1.04+/ 0.05 versus 0.7+/-0.06 IU/l on the day of hCG). The number of FSH ampoules used was lower in group 1 (42+/-2 versus 59+/-3). The numbers of mature oocytes (10.1+/-0.54 versus 7.4+/-0.55), of fertilized oocytes (8.24+/-0.35 versus 6.34+/ 0.37) and of embryos (7.8+/-0.36 versus 5.9+/-0.37) were significantly higher in group 1. No significant differences were found in pregnancy (38.8 versus 25.3%), implantation (22.6 versus 13.8%) or abortion (6.1 versus 5.0%) rates. Cumulative pregnancy (fresh plus frozen embryo transfers: 56.8 versus 35.4%) rate was significantly higher in group 1. CONCLUSION: A half-dose of depot triptorelin can be successfully used in ovarian stimulation for IVF and produce a higher number of good quality embryos with a good chance of implantation. PMID- 15271869 TI - Toluidine blue cytometry test for sperm DNA conformation: comparison with the flow cytometric sperm chromatin structure and TUNEL assays. AB - BACKGROUND: Sperm DNA integrity (SDI) is an important factor in the prognosis of male fertility. Here we compare the toluidine blue (TB) image cytometry test, recently proposed by us for SDI assessment, with two other tests-the sperm chromatin structure assay (SCSA) and the terminal nick-end labelling (TUNEL) assay. METHODS: Sperm samples from 35 men were evaluated for standard sperm parameters and subjected to the TB test and SCSA. Eighteen of the 35 samples were also subjected to the TUNEL assay. RESULTS: The proportion of sperm cells with abnormal DNA integrity assayed by the TB test correlated strongly with the proportion of abnormal cells detected by the SCSA and TUNEL assay (rho=-0.84 and rho=0.80, P<0.001, respectively). Furthermore, the fractions of abnormal cells by the TB test corresponded closely to the sum of two SCSA parameters, the DNA fragmentation index (DFI) and the fraction of highly DNA-stainable cells (HDS) (medians 33.0 versus 32.0%, P=0.6). CONCLUSIONS: Abnormal cells in a TB test correspond to the sum of DFI and HDS fractions in the SCSA. TB-positive cells may represent sperm with fragmented DNA and/or abnormal chromatin structure. Because the TB test is an easy and inexpensive method, its potential use as a routine test for sperm DNA integrity, complementary to standard semen parameters, should be investigated further. PMID- 15271870 TI - The FLUSH trial--flushing with lipiodol for unexplained (and endometriosis related) subfertility by hysterosalpingography: a randomized trial. AB - BACKGROUND: To assess the effectiveness of flushing with the oil-soluble contrast medium lipiodol in women with unexplained infertility. METHODS: An open randomized controlled trial design in a single centre secondary and tertiary level infertility service setting. A total of 158 women with unexplained infertility were stratified into two populations: 96 women without confirmed endometriosis and 62 women with endometriosis who had normal Fallopian tubes and ovaries. Randomization was computer-generated, with allocation concealment by opaque sequentially numbered envelopes. Lipiodol flushing was tested versus no intervention. The main outcome measures were clinical pregnancy (assessed at 6 months following randomization) and live birth. RESULTS: Lipiodol flushing resulted in a significant increase in pregnancy [48.0 versus 10.8%, relative risk (RR) 4.44, 95% confidence interval (CI) 1.61-12.21] and live birth (40.0 versus 10.8%, RR 3.70, 95% CI 1.30-10.50) rates versus no intervention for women with endometriosis, although there was no significant difference in pregnancy (33.3 versus 20.8%, RR 1.60, 95% CI 0.81-3.16) or live birth (27.1 versus 14.6%, RR 1.86, 95% CI 0.81-4.25) rates for women with unexplained infertility without confirmed endometriosis. CONCLUSIONS: Lipiodol flushing is an effective treatment for couples with unexplained infertility (based on meta-analysis data), but is particularly effective for women with endometriosis who have normal Fallopian tubes and ovaries. PMID- 15271871 TI - Mifepristone does not induce cervical softening in non-pregnant women. AB - BACKGROUND: Many techniques have been developed to soften the cervix to reduce complications following surgical dilatation. Progesterone inhibits myometrial contractility and its secretion during pregnancy ensures cervical competence. We used the progesterone antagonist mifepristone as a cervical ripening agent and evaluated its effect prior to office hysteroscopy. METHODS: Fifty-eight healthy non-pregnant women aged 18-50 were studied in a randomized double-blind study. They received mifepristone (200 mg) or placebo 30 h prior to hysteroscopy. A Hegar test was performed prior to drug administration and again before hysteroscopy. A visual analogue pain scale was used to assess pain. RESULTS: Medical history, physical examination and blood tests were similar in both groups, except for serum progesterone which was higher in the study group. Hegar measurement prior to drug ingestion was similar in both groups and after a mean time of 30.3 h increased in both groups. Neither the DeltaHegar measurement nor the pain scale was different in the two groups. There was also no effect of the high progesterone levels. CONCLUSIONS: Unlike its dramatic effect in the pregnant uterus, mifepristone administered 30 h prior to hysteroscopy was not effective in ripening the cervix of non-pregnant women. PMID- 15271872 TI - Breast cancer risk associated with being treated for infertility: results from the French E3N cohort study. AB - BACKGROUND: The use of fertility drugs (FDs) is steadily increasing in Western countries and concern has been raised as to the possible impact of fertility treatments on breast cancer risk. METHODS: We analysed this association in the French E3N study. In this prospective cohort, data on treatment against infertility, duration and time of administration were collected at entry through self-administered questionnaires. Cox regression analysis was used to estimate adjusted relative risks (RRs). RESULTS: Among the 92 555 women from the study population, 6602 women were treated for infertility. During the 10 year follow-up period, 2571 cases of primary invasive breast cancer were diagnosed (183 in treated women). Our study showed no overall significant association between breast cancer risk and treatment for infertility (RR = 0.95, confidence interval 0.82-1.11), after surgery or FDs, and whatever the type, the duration of use and the age at first use of FDs. However, infertility treatment was associated with an increased risk, of borderline significance, of breast cancer among women with a family history of breast cancer. This last result had limited statistical power. CONCLUSIONS: Our study provides evidence that treatment for infertility does not influence breast cancer risk overall. An interaction with a familial history of breast cancer is possible but should be investigated further. PMID- 15271873 TI - A possible mechanism for the action of adrenomedullin in brain to stimulate stress hormone secretion. AB - Adrenomedullin (AM) has been reported to have actions at each level of the hypothalamo-pituitary-adrenal (HPA) axis, suggesting that the peptide plays a role in the organization of the neuroendocrine responses to stress. We examined the mechanism by which AM regulates the central nervous system branch of the HPA axis as well as the possible role of AM in the modulation of the releases of two other hormones, prolactin and GH, whose secretions also are altered by stress. Intracerebroventricular administration of AM led to elevated plasma corticosterone levels in unrestrained, conscious male rats. This effect was abrogated by pretreatment with a CRH antagonist, suggesting that AM activates the HPA axis by causing the release of CRH into hypophyseal portal vessels. In addition, AM given intracerebroventricularly stimulated the release of prolactin but did not alter the secretion of GH. We propose that AM produced in the brain may be an important neuromodulator of the hormonal stress response. PMID- 15271874 TI - Regulation of the follicle-stimulating hormone beta gene by the LHX3 LIM homeodomain transcription factor. AB - FSH is a critical hormone regulator of gonadal function that is secreted from the pituitary gonadotrope cell. Human patients and animal models with mutations in the LHX3 LIM-homeodomain transcription factor gene exhibit complex endocrine diseases, including reproductive disorders with loss of FSH. We demonstrate that in both heterologous and pituitary gonadotrope cells, specific LHX3 isoforms activate the FSH beta-subunit promoter, but not the proximal LHbeta promoter. The related LHX4 mammalian transcription factor can also induce FSHbeta promoter transcription, but the homologous Drosophila protein LIM3 cannot. The actions of LHX3 are specifically blocked by a dominant negative LHX3 protein containing a Kruppel-associated box domain. Six LHX3-binding sites were characterized within the FSHbeta promoter, including three within a proximal region that also mediates gene regulation by other transcription factors and activin. Mutations of the proximal binding sites demonstrate their importance for LHX3 induction of the FSHbeta promoter and basal promoter activity in gonadotrope cells. Using quantitative methods, we show that the responses of the FSHbeta promoter to activin do not require induction of the LHX3 gene. By comparative genomics using the human FSHbeta promoter, we demonstrate structural and functional conservation of promoter induction by LHX3. We conclude that the LHX3 LIM homeodomain transcription factor is involved in activation of the FSH beta-subunit gene in the pituitary gonadotrope cell. PMID- 15271875 TI - Involvement of brainstem catecholaminergic inputs to the hypothalamic paraventricular nucleus in estrogen receptor alpha expression in this nucleus during different stress conditions in female rats. AB - In the present study, we determined the involvement of brainstem catecholaminergic inputs to the paraventricular nucleus (PVN) on estrogen receptor alpha (ERalpha) expression in this nucleus during conditions of 48-h fasting, 2-deoxy-d-glucose (2DG)-induced acute glucoprivation and 1-h immobilization, in ovariectomized rats. Our approach was to examine the effect of lesioning catecholaminergic inputs to the PVN using DSAP [saporin-conjugated anti DBH (dopamine-beta-hydroxylase)]. Bilateral injection of DSAP into the PVN, 2 wk before stress, prevented fasting-, glucoprivation-, and immobilization-induced increase in ERalpha-immunopositive cells in the PVN. The DBH-immunoreactive (ir) terminals in the PVN were severely depleted by DSAP injection in all experimental groups. Among the brainstem noradreneregic cell groups examined, DBH-ir cell bodies were significantly reduced in the A2 region of all experimental groups treated with DSAP compared with the saporin- and vehicle-injected controls. PVN DSAP injection caused a small, but not significant, decrease in A1 DBH-ir cell bodies in fasted and immobilized rats, and a significant, but slight, reduction in A1 DBH-ir cell bodies of iv 2DG- injected rats compared with PVN vehicle injected or PVN saporin-injected controls. The A6 DBH-ir cell bodies in all experimental groups treated with DSAP, saporin, or vehicle did not show any significant difference. These results suggest that the brainstem catecholaminergic inputs to the PVN, especially from the A2 cell group, may play a major role in mediating the induction of ERalpha expression in the PVN by metabolic stressors such as fasting, acute glucoprivation, and less specific stressors, such as immobilization, in female rats. PMID- 15271876 TI - Evidence that cocaine- and amphetamine-regulated transcript is a novel intraovarian regulator of follicular atresia. AB - We recently obtained evidence that cocaine- and amphetamine-regulated transcript (CART), a potent anorectic neuropeptide, is expressed in the bovine ovary. The objectives of this study were to characterize bovine ovarian CART and determine its localization, regulation, and regulatory role during follicular development. CART mRNA was detected in stroma of adult ovaries and in large follicles, but was undetectable in several peripheral tissues, fetal ovaries, and corpora lutea. Within the ovary, CART mRNA and peptide were localized to the granulosal layer of some, but not all, antral follicles, with low, but detectable, expression in oocytes and cumulus cells. CART mRNA was undetectable in granulosal cells of dominant ovulatory follicles collected before and after the preovulatory gonadotropin surge, but was detected in the granulosal layer of adjacent subordinate follicles. In addition, amounts of CART mRNA and follicular fluid concentrations of CART peptide were greater in subordinate follicles vs. dominant follicles of the first follicular wave. Furthermore, CART treatment inhibited basal estradiol production, but not progesterone production, by granulosal cells in a dose-dependent fashion, and the effect was dependent on stage of cell differentiation. We conclude that granulosal cell CART expression is temporally regulated and potentially associated with follicle health status, and CART can inhibit granulosal cell estradiol production. Thus, CART may be a novel local regulator of follicular atresia in the bovine ovary. PMID- 15271877 TI - Progesterone receptor and the cell cycle modulate apoptosis in granulosa cells. AB - Our previous studies showed that exposure of bovine preovulatory follicles to the LH surge-induced resistance of granulosa cells, but not theca cells, to apoptosis. Here, the temporal development of resistance to apoptosis and potential roles of progesterone receptor (PR) and alterations in the cell cycle in mediating this effect were examined. Injection of cows with GnRH induced an LH surge within 2 h. Granulosa cells isolated 0, 6, and 10 h after GnRH were sensitive to Fas ligand-induced apoptosis, but cells isolated at 14 h were resistant. PR was first detectable in granulosa cells at 10 and 14 h and was not detectable in theca. Treatment of granulosa cells isolated 14 h after GnRH with the PR antagonist, RU486, induced susceptibility to apoptosis, an effect mediated by PR and not glucocorticoid receptor. After GnRH treatment, granulosa cells, but not theca cells, exited the cell cycle, expression of cyclin D2 was reduced, and p27(Kip1) was elevated. Treatment of granulosa cells isolated from small antral follicles with the G1 phase blocker, mimosine, reduced Fas ligand-induced killing, suggesting that nonproliferating cells are resistant to apoptosis. Treatment of granulosa cells isolated 14 h after GnRH with RU486 induced reentry of some cells into the cell cycle and reversed resistance to apoptosis, suggesting that cycling cells became susceptible to apoptosis. Treatment with mimosine prevented the ability of RU486 to promote susceptibility to apoptosis. In summary, the LH surge induces expression of PR by granulosa cells and withdrawal from the cell cycle, and these events promote resistance to apoptosis. PMID- 15271878 TI - Intestinal lipoprotein production is stimulated by an acute elevation of plasma free fatty acids in the fasting state: studies in insulin-resistant and insulin sensitized Syrian golden hamsters. AB - It is not known whether intestinal lipoprotein production is stimulated by an acute elevation of plasma free fatty acids (FFA). We examined the effect of an intralipid and heparin infusion on the intestinal lipoprotein production rate (PR) in insulin-sensitive [chow-fed (CHOW)], insulin-resistant [60% fructose (FRUC) or 60% fat-fed (FAT)], and insulin-sensitized [FRUC or FAT plus rosiglitazone (RSG)-treated] Syrian Golden hamsters. After 5 wk of treatment, overnight-fasted hamsters underwent in vivo Triton WR-1339 studies for measurement of apolipoprotein B48 (apoB48) PR in large (Svedberg unit, >400) and small (Svedberg unit, 100-400) lipoprotein fractions, with an antecedent 90-min infusion of 20% intralipid and heparin (IH) to raise plasma FFA levels approximately 5- to 8-fold vs. those in the saline control study. IH markedly increased apoB48 PR in CHOW by 3- to 5-fold, which was confirmed ex vivo in pulse chase experiments in primary cultured hamster enterocytes. Oleate, but not glycerol, infusion was associated with a similar elevation of apoB48 PR as IH. In FRUC and FAT, basal (saline control) apoB48 PR was approximately 4-fold greater than that in CHOW; there was no additional stimulation with IH in vivo and only minimal additional stimulation ex vivo. RSG partially normalized basal apoB48 PR in FAT and FRUC, and PR was markedly stimulated with IH. We conclude that intestinal lipoprotein production is markedly stimulated by an acute elevation of plasma FFAs in insulin-sensitive hamsters, in which basal production is low, but minimally in insulin-resistant hamsters, in which basal production is already elevated. With RSG treatment, basal PR is partially normalized, and they become more susceptible to the acute FFA stimulatory effect. PMID- 15271879 TI - Metabolic consequences of hypoxia from birth and dexamethasone treatment in the neonatal rat: comprehensive hepatic lipid and fatty acid profiling. AB - Neonatal hypoxia is a common condition resulting from pulmonary and/or cardiac dysfunction. Dexamethasone therapy is a common treatment for many causes of neonatal distress, including hypoxia. The present study examined the effects of dexamethasone treatment on both normoxic and hypoxic neonatal rats. We performed comprehensive hepatic fatty acid/lipid profiling and evaluated changes in pertinent plasma hormones and lipids and a functional hepatic correlate, i.e. hepatic lipase activity. Rats were exposed to hypoxia from birth to 7 d of age. A 4-d tapering dose regimen of dexamethasone was administered on: postnatal day (PD)3 (0.5 mg/kg), PD4 (0.25 mg/kg), PD5 (0.125 mg/kg), and PD6 (0.05 mg/kg). The most significant finding was that dexamethasone attenuated nearly all hypoxia induced changes in hepatic lipid profiles. Hypoxia increased the concentration of hepatic triacylglyceride and free fatty acids and, more specifically, increased a number of fatty acid metabolites within these lipid classes. Administration of dexamethasone blocked these increases. Hypoxia alone increased the plasma concentration of cholesterol and triacylglyceride, had no effect on plasma glucose, and only tended to increase plasma insulin. Dexamethasone administration to hypoxic pups resulted in an additional increase in plasma lipid concentrations, an increase in insulin, and a decrease in plasma glucose. Hypoxia and dexamethasone treatment each decreased total hepatic lipase activity. Normoxic pups treated with dexamethasone displayed increased plasma lipids and insulin. The effects of dexamethasone on hepatic function in the hypoxic neonate are dramatic and have significant implications in the assessment and treatment of metabolic dysfunction in the newborn. PMID- 15271880 TI - Grape seed-derived procyanidins have an antihyperglycemic effect in streptozotocin-induced diabetic rats and insulinomimetic activity in insulin sensitive cell lines. AB - Flavonoids are functional constituents of many fruits and vegetables. Some flavonoids have antidiabetic properties because they improve altered glucose and oxidative metabolisms of diabetic states. Procyanidins are flavonoids with an oligomeric structure, and it has been shown that they can improve the pathological oxidative state of a diabetic situation. To evaluate their effects on glucose metabolism, we administered an extract of grape seed procyanidins (PE) orally to streptozotocin-induced diabetic rats. This had an antihyperglycemic effect, which was significantly increased if PE administration was accompanied by a low insulin dose. The antihyperglycemic effect of PE may be partially due to the insulinomimetic activity of procyanidins on insulin-sensitive cell lines. PE stimulated glucose uptake in L6E9 myotubes and 3T3-L1 adipocytes in a dose dependent manner. Like insulin action, the effect of PE on glucose uptake was sensitive to wortmannin, an inhibitor of phosphoinositol 3-kinase and to SB203580, an inhibitor of p38 MAPK. PE action also stimulated glucose transporter 4 translocation to the plasma membrane. In summary, procyanidins have insulin like effects in insulin-sensitive cells that could help to explain their antihyperglycemic effect in vivo. These effects must be added to their antioxidant activity to explain why they can improve diabetic situations. PMID- 15271881 TI - Region-specific leptin resistance within the hypothalamus of diet-induced obese mice. AB - Leptin resistance in diet-induced obese (DIO) mice is characterized by elevated serum leptin and a decreased response to exogenous leptin and is caused by unknown defects in the central nervous system. Leptin normally acts on several brain nuclei, but a detailed description of leptin resistance within individual brain regions has not been reported. We first mapped leptin-responsive cells in brains from DIO mice using phospho-signal transducer and activator of transcription (P-STAT3) immunohistochemistry. After 16 wk of high-fat-diet feeding, leptin-activated P-STAT3 staining within the arcuate nucleus (ARC) was dramatically decreased. In contrast, other hypothalamic and extrahypothalamic nuclei remained leptin sensitive. Reduced leptin-induced P-STAT3 in the ARC could also be detected after 4 wk and as early as 6 d of a high-fat diet. To examine potential mechanisms for leptin-resistant STAT3 activation in the ARC of DIO mice, we measured mRNA levels of candidate signaling molecules in the leptin receptor-STAT3 pathway. We found that the level of suppressor of cytokine signaling 3 (SOCS-3), an inhibitor of leptin signaling, is specifically increased in the ARC of DIO mice. The study suggests that the ARC is selectively leptin resistant in DIO mice and that this may be caused by elevated suppressor of cytokine signaling 3 in this hypothalamic nucleus. Defects in leptin action in the ARC may play a role in the pathogenesis of leptin-resistant obesity. PMID- 15271882 TI - Evidence that basal activity, but not transactivation, of the epidermal growth factor receptor tyrosine kinase is required for insulin-like growth factor I induced activation of extracellular signal-regulated kinase in oral carcinoma cells. AB - IGF-I receptor (IGF-IR) is involved in numerous biological functions via its major downstream signaling molecules, extracellular signal-regulated kinase (ERK) and phosphatidylinositol 3'-kinase/Akt. The IGF-I-induced activation of ERK, but not that of Akt, is reportedly mediated by the transactivation of the epidermal growth factor receptor (EGFR) tyrosine kinase (TK). The mechanism for the EGFR-TK dependent activation, however, still remains largely unknown. We found that an oral carcinoma cell line overexpressing EGFR, Ca9-22, exhibited IGF-I-induced activation of both Akt and ERK, but that only the latter was significantly decreased by a specific inhibitor of EGFR-TK, tyrphostin AG1478. In this report we provide evidence for the existence in this cell line of a novel mechanism by which IGF-I induces ERK activation in a manner that is dependent on the basal level of EGFR-TK activity, but is independent of receptor transactivation. In addition, we show that c-Raf kinase is likely to be a key regulator of this mechanism. The elucidation of such a unique mechanism involving cross-talk between EGFR and heterologous receptors may shed additional light on the clinical use of EGFR-TK inhibitors in antitumor therapies. PMID- 15271883 TI - Cocaine- and amphetamine-regulated transcript activates the hypothalamic pituitary-adrenal axis through a corticotropin-releasing factor receptor dependent mechanism. AB - Cocaine- and amphetamine-regulated transcript (CART) is a highly expressed hypothalamic transcript that is concentrated in areas associated with the stress response. There is evidence for a role of CART in the regulation of the hypothalamic-pituitary-adrenal (HPA) axis. However, it is not clear whether CART regulates activity of the HPA axis by directly stimulating ACTH release from pituitary corticotropes or through interaction with hypothalamic factors. To address this issue, the effects of central and peripheral administration of CART on the HPA axis were compared. Central administration of CART(55-102) (1 microg) significantly increased circulating levels of ACTH (481 +/- 122 vs. 93 +/- 14 pg/ml; CART vs. vehicle) and corticosterone (460 +/- 29 vs. 179 +/- 62 ng/ml; CART vs. vehicle). In contrast, iv injection of CART(55-102) (0.09-9.0 nmol/kg) did not significantly affect circulating levels of ACTH or corticosterone. The corticotropin-releasing factor (CRF) receptor antagonist Astressin B was used to determine whether CART(55-102) elicits ACTH secretion via a CRF receptor dependent mechanism. Injection of Astressin B (50 microg/kg, iv) inhibited CART(55-102)-induced ACTH and corticosterone responses. The effects of CART(55 102) on CRF and arginine vasopressin (AVP) expression were also examined in static hypothalamic explants. RT-PCR analysis revealed a significant up regulation of CRF and AVP mRNA levels after CART(55-102) (10 nm and 1 microm) treatment. Last, the effects of CART(55-102) on CRF- and AVP-mediated ACTH release was investigated in dispersed rat anterior pituitary cells. Incubation of CART(55-102) (10-100 nm) did not significantly affect ACTH release from anterior pituitary cells. Findings from the present study suggest that CART regulates activity of the HPA axis through a CRF-dependent central mechanism and not by means of direct interaction with pituitary corticotropes. PMID- 15271884 TI - Adenovirus-mediated transfer of thyroid transcription factor-1 induces radioiodide organification and retention in thyroid cancer cells. AB - Loss of thyroid-specific gene expression and functions accompanied by loss of thyroid transcription factors render them unresponsive to radioiodide therapy in poorly differentiated and anaplastic thyroid cancer. In anticipation of reactivation of thyroid functions, we investigated the effect of thyroid transcription factor-1 (TTF-1) gene transfer on thyroid cancer cells. Reexpression of thyroperoxidase (TPO) and thyroglobulin (Tg) mRNA and protein was detected in poorly differentiated human thyroid cancer cells that were infected with an adenovirus vector containing TTF-1 (AdTTF-1). Although TTF-1 gene transfer faintly induced iodide uptake, the induction of sodium/iodide symporter (NIS) mRNA was not observed in AdTTF-1-infected cells. To analyze the effect of TTF-1 on iodide metabolism, we transfected an NIS expression vector into BHP18 21v cells and cloned a cell line (N-BHP18-21v) that stably expressed NIS. The treatment of N-BHP18-21v cells with AdTTF-1 significantly increased the amount of protein-bound radioiodide and prolonged the iodide efflux. AdTTF-1 injections significantly induced iodide retention and organification in tumors formed from N BHP18-21v cells in nude mice. These results indicate that AdTTF-1 specifically induces iodide organification and retards iodide efflux in thyroid cancer cells in vitro and in vivo. PMID- 15271885 TI - In vivo and in vitro inhibition of cyp19 gene expression by prostaglandin F2alpha in murine luteal cells: implication of GATA-4. AB - A major function of the corpus luteum (CL) is to secrete progesterone. In rats, this gland also produces significant amounts of 17beta-estradiol. Progesterone and 17beta-estradiol are important regulators of rat luteal cell function. Estrogen biosynthesis is catalyzed by P450aromatase (P450arom), which is encoded by the cyp19 gene. In the rat CL, P450arom is expressed throughout pregnancy until the day before parturition, when it rapidly decreases. The mechanisms that control P450arom expression in luteal cells, particularly, the one or more factors that cause its rapid fall before parturition, are not known. Inasmuch as prostaglandin (PG) F(2alpha) plays a key role in the regulation of luteal function at the end of pregnancy, the purpose of this investigation was to determine whether PGF(2alpha) affect the expression of P450arom in the CL before parturition. PGF(2alpha) decreased luteal P450arom mRNA and protein levels in vivo and in vitro. A decrease in P450arom mRNA was also observed in mice CL just before parturition, but this change did not take place in PGF(2alpha) receptor knockout mice. The time course of the decrease in P450arom mRNA by PGF(2alpha) reflected the P450arom mRNA half-life determined by actinomycin D. Moreover, nuclear run-on assay showed that PGF(2alpha) attenuates P450arom gene transcription. Gel shift assays revealed that GATA-4 binds to the P450aromatase promoter, and that such binding is increased by PGF(2alpha). It is concluded that PGF(2alpha) decreases luteal P450arom mRNA levels at the end of pregnancy in rodents by inhibiting cyp19 expression. PMID- 15271886 TI - Interleukin (IL)-6, but not IL-1, induction in the brain downstream of cyclooxygenase-2 is essential for the induction of febrile response against peripheral IL-1alpha. AB - IL-1 is an endogenous pyrogen produced upon inflammation or infection. Previously, we showed that, upon injection with turpentine, IL-1 is induced in the brain in association with the development of fever. The role of endogenous IL 1 in the brain and the signaling cascade to activate thermosensitive neurons, however, remain to be elucidated. In this report, febrile response was analyzed after peripheral injection of IL-1alpha. We found that a normal febrile response was induced even in IL-1alpha/beta-deficient mice, indicating that production of IL-1 in the brain is not necessarily required for the response. In contrast, IL-6 deficient mice did not exhibit a febrile response. Cyclooxygenase (Cox)-2 expression in the brain was strongly induced 1.5 h after injection of IL-1alpha, whereas IL-6 expression was observed 3 h after the injection. Cox-2 expression in the brain was not influenced by IL-6 deficiency, whereas indomethacin, an inhibitor of cyclooxygenases, completely inhibited induction of IL-6. These observations suggest a mechanism of IL-1-induced febrile response in which IL-1 in the blood activates Cox-2, with the resulting prostaglandin E(2) inducing IL-6 in the brain, leading to the development of fever. PMID- 15271887 TI - Modulation and utilization of host cell phosphoinositides by Salmonella spp. PMID- 15271888 TI - Expression of members of the 28-kilodalton major outer membrane protein family of Ehrlichia chaffeensis during persistent infection. AB - The 28-kDa immunodominant outer membrane proteins (P28 OMPs) of Ehrlichia chaffeensis are encoded by a multigene family. As an indirect measure of the in vivo expression of the members of the p28 multigene family of E. chaffeensis, sera from two beagle dogs experimentally infected with E. chaffeensis were evaluated for the presence of specific antibodies to P28 OMPs by enzyme-linked immunosorbent assay. Antigenic peptides unique to each of the P28s were identified within the first hypervariable region of each P28 OMP. Serological responses to peptides derived from all P28 OMPs were detected from day 30 postinoculation to day 468 and from day 46 until day 159 in the two beagles. Although antibody titers to the peptides fluctuated, the peak response to all of the peptides appeared simultaneously in each dog. The antibody responses to another outer membrane protein of E. chaffeensis (GP120) showed similar temporal and quantitative changes. These data suggest that the P28 OMPs are expressed concurrently during persistent Ehrlichia infection. PMID- 15271889 TI - Alpha 1-antitrypsin binds to and interferes with functionality of EspB from atypical and typical enteropathogenic Escherichia coli strains. AB - Enteropathogenic Escherichia coli (EPEC), including diffusely adhering atypical E. coli, strains use a type III secretion system to deliver effector proteins into the membrane and cytoplasm of infected cells. The E. coli secreted proteins A, B, and D (EspA, EspB, and EspD) are required for the formation of the characteristic attaching and effacing (A/E) lesions. EspB and EspD are thought to form a translocation pore in the host cell membrane through which effector proteins are injected into the host cytosol. Besides its function in pore formation, EspB has been found in the cytosol and implicated to function as an effector protein. We screened for putative host cell proteins interacting with EspB of atypical EPEC strains and identified alpha(1)-antitrypsin (AAT) as a binding partner for EspB. AAT binds to EspB in pull-down and overlay experiments and also to EspD in overlay experiments. In agreement with the role of EspB and EspD in pore formation, EPEC-mediated hemolysis of red blood cells is strongly reduced by AAT in a concentration-dependent manner, indicating that AAT interferes with type III secretion by inhibiting the formation of the translocation pore. This is further supported by a decreased actin polymerization after infection of HeLa or CaCo-2 cells with EPEC in the presence of physiologically relevant concentrations of AAT. In this study, we identify AAT as a new binding partner for EspB and EspD, suggesting a previously unappreciated role for AAT in host cell defense against EPEC infections and potentially also against other bacterial pathogens. PMID- 15271890 TI - Cleavage of human transferrin by Porphyromonas gingivalis gingipains promotes growth and formation of hydroxyl radicals. AB - Porphyromonas gingivalis, a gram-negative anaerobic bacterium associated with active lesions of chronic periodontitis, produces several proteinases which are presumably involved in host colonization, perturbation of the immune system, and tissue destruction. The aims of this study were to investigate the degradation of human transferrin by gingipain cysteine proteinases of P. gingivalis and to demonstrate the production of toxic hydroxyl radicals (HO*) catalyzed by the iron containing transferrin fragments generated or by release of iron itself. Analysis by polyacrylamide gel electrophoresis and Western immunoblotting showed that preparations of Arg- and Lys-gingipains of P. gingivalis cleave transferrin (iron free and iron-saturated forms) into fragments of various sizes. Interestingly, gingival crevicular fluid samples from diseased periodontal sites but not samples from healthy periodontal sites contained fragments of transferrin. By using (55)Fe-transferrin, it was found that degradation by P. gingivalis gingipains resulted in the production of free iron, as well as iron bound to lower-molecular mass fragments. Subsequent to the degradation of transferrin, bacterial cells assimilated intracellularly the radiolabeled iron. Growth of P. gingivalis ATCC 33277, but not growth of an Arg-gingipain- and Lys-gingipain-deficient mutant, was possible in a chemically defined medium containing 30% iron-saturated transferrin as the only source of iron and peptides, suggesting that gingipains play a critical role in the acquisition of essential growth nutrients. Finally, the transferrin degradation products generated by Arg-gingipains A and B were capable of catalyzing the formation of HO*, as determined by a hypoxanthine/xanthine oxidase system and spin trapping-electron paramagnetic resonance spectrometry. Our study indicates that P. gingivalis gingipains degrade human transferrin, providing sources of iron and peptides. The iron-containing transferrin fragments or the release of iron itself may contribute to tissue destruction by catalyzing the formation of toxic HO*. PMID- 15271891 TI - Chlamydia trachomatis-specific human CD8+ T cells show two patterns of antigen recognition. AB - Chlamydia trachomatis is an intracellular gram-negative bacteria which causes several clinically important diseases. T-cell-mediated immunity and the production of gamma interferon (IFN-gamma) are known to be essential for the clearance of the bacteria in vivo. Here we have investigated CD8(+)-T-cell responses to C. trachomatis in patients with previous episodes of chlamydia infection. To isolate C. trachomatis-specific CD8(+)-T-cell lines, dendritic cells (DC) were infected with C. trachomatis and cocultured with purified CD8(+) T cells to generate C. trachomatis-specific CD8(+)-T-cell lines which were then cloned. Two patterns of recognition of C. trachomatis-infected cells by CD8(+)-T cell clones were identified. In the first, C. trachomatis antigens were recognized in association with classical class I HLA antigens, and responses were inhibited by class I HLA-specific monoclonal antibodies. The second set of clones was unrestricted by classical HLA class I, and further studies showed that CD1 molecules were also not the restriction element for those clones. Both types of clones produced IFN-gamma in response to C. trachomatis and were able to lyse C. trachomatis-infected target cells. However, unrestricted clones recognized C. trachomatis-infected cells at much earlier time points postinfection than HLA restricted clones. Coculture of C. trachomatis-infected DC with the C. trachomatis-specific clones induced DC activation and a rapid enhancement of interleukin-12 (IL-12) production. Early production of IL-12 during C. trachomatis infection, facilitated by unrestricted CD8(+)-T-cell clones, may be important in ensuring a subsequent Th1 T-cell-mediated response by classical major histocompatibility complex-restricted CD4(+) and CD8(+) T cells. PMID- 15271892 TI - Intranasal coadministration of the Cry1Ac protoxin with amoebal lysates increases protection against Naegleria fowleri meningoencephalitis. AB - Cry1Ac protoxin has potent mucosal and systemic adjuvant effects on antibody responses to proteins or polysaccharides. In this work, we examined whether Cry1Ac increased protective immunity against fatal Naegleria fowleri infection in mice, which resembles human primary amoebic meningoencephalitis. Higher immunoglobulin G (IgG) than IgA anti-N. fowleri responses were elicited in the serum and tracheopulmonary fluids of mice immunized by the intranasal or intraperitoneal route with N. fowleri lysates either alone or with Cry1Ac or cholera toxin. Superior protection against a lethal challenge with 5 x 10(4) live N. fowleri trophozoites was achieved for immunization by the intranasal route. Intranasal immunization of N. fowleri lysates coadministered with Cry1Ac increased survival to 100%; interestingly, immunization with Cry1Ac alone conferred similar protection to that achieved with amoebal lysates alone (60%). When mice intranasally immunized with Cry1Ac plus lysates were challenged with amoebae, both IgG and IgA mucosal responses were rapidly increased, but only the increased IgG response persisted until day 60 in surviving mice. The brief rise in the level of specific mucosal IgA does not exclude the role that this isotype may play in the early defense against this parasite, since higher IgA responses were detected in nasal fluids of mice intranasally immunized with lysates plus either Cry1Ac or cholera toxin, which, indeed, were the treatments that provided the major protection levels. In contrast, serum antibody responses do not seem to be related to the protection level achieved. Both acquired and innate immune systems seem to play a role in host defense against N. fowleri infection, but further studies are required to elucidate the mechanisms involved in protective effects conferred by Cry1Ac, which may be a valuable tool to improve mucosal vaccines. PMID- 15271893 TI - Long-term multiepitopic cytotoxic-T-lymphocyte responses induced in chimpanzees by combinations of Plasmodium falciparum liver-stage peptides and lipopeptides. AB - Preclinical immunogenicity studies of 12 malaria peptides, selected from four Plasmodium falciparum antigens (Ags), namely, LSA1, LSA3, SALSA, and STARP, that are expressed at the pre-erythrocytic (sporozoite and liver) stages of the human parasite were carried out in chimpanzees. To strengthen their immunogenicity, six of these synthetic peptides were modified by the C-terminal addition of a single palmitoyl chain (lipopeptides) and delivered without adjuvant, whereas the remaining six unmodified peptides were emulsified and delivered by using Montanide ISA51 adjuvant. We have previously reported that these peptides and lipopeptides induce high B-cell and CD4(+)-T-helper responses in chimpanzees. In this report, we show their ability to induce multiepitopic and long-lasting antigen-specific CD8(+) cytotoxic-T-lymphocyte (CTL) responses. The magnitude, consistency, and memory of CTL responses generated by LSA3 peptides point to the strong immunogenicity of this liver-stage Ag. These findings support the screening strategy used to select the four P. falciparum pre-erythrocytic Ags and emphasize their valuable immunogenic properties. The successful immunization of nonhuman primates with combinations of corresponding peptides in a mineral oil emulsion (ISA51) and lipopeptides in saline provide a vaccine formulation that can be tested in humans. PMID- 15271894 TI - Mycobacterium tuberculosis diverts alpha interferon-induced monocyte differentiation from dendritic cells into immunoprivileged macrophage-like host cells. AB - Dendritic cells (DCs) are critical for initiating a pathogen-specific T-cell response. During chronic infections the pool of tissue DCs must be renewed by recruitment of both circulating DC progenitors and in loco differentiating monocytes. However, the interaction of monocytes with pathogens could affect their differentiation. Mycobacterium tuberculosis has been shown to variably interfere with the generation and function of antigen-presenting cells (APCs). In this study we found that when alpha interferon (IFN-alpha) is used as an inductor of monocyte differentiation, M. tuberculosis inhibits the generation of DCs, forcing the generation of immunoprivileged macrophage-like cells instead. Cells derived from M. tuberculosis-infected monocyte-derived macrophages (M. tuberculosis-infected MoMphi) retained CD14 without acquiring CD1 molecules and partially expressed B7.2 but did not up-regulate B7.1 and major histocompatibility complex (MHC) class I and II molecules. They synthesized tumor necrosis factor alpha and interleukin-10 (IL-10) but not IL-12. They also showed a reduced ability to induce proliferation and functional polarization of allogeneic T lymphocytes. Thus, in the presence of IFN-alpha, M. tuberculosis may hamper the renewal of potent APCs, such as DCs, generating a safe habitat for intracellular growth. M. tuberculosis-infected MoMphi, in fact, showed reduced expression of both signal 1 (CD1, MHC classes I and II) and signal 2 (B7.1 and B7.2), which are essential for mycobacterium-specific T-lymphocyte priming and/or activation. These data further suggest that M. tuberculosis has the ability to specifically interfere with monocyte differentiation. This ability may represent an effective M. tuberculosis strategy for eluding immune surveillance and persisting in the host. PMID- 15271895 TI - Potential involvement of gelatinases and their inhibitors in Mannheimia haemolytica pneumonia in cattle. AB - Mannheimia haemolytica infection of the lower respiratory tract of cattle results in a bronchofibrinous pneumonia characterized by massive cellular influx and lung tissue remodeling and scarring. Since altered levels of gelatinases and their inhibitors have been detected in a variety of inflammatory conditions and are associated with tissue remodeling, we examined the presence of gelatinases in lesional and nonlesional lung tissue obtained from calves experimentally infected with M. haemolytica. Lesional tissue had elevated levels of progelatinase A and B and active gelatinase A and B when compared with nonlesional tissue obtained from the same lung lobe. In vitro, M. haemolytica products stimulated production of gelatinase B, but not its activation, by bovine monocytes. Alveolar macrophages showed constitutive production of gelatinase B but no change in response to M. haemolytica products. Bovine neutrophils exposed to M. haemolytica products also released gelatinase B, and there was a significant increase in the activated form of this enzyme. These effects were virtually identical when recombinant O sialoglycoprotease was used to stimulate these cells. M. haemolytica products also enhanced the expression by bovine monocytes and alveolar macrophages of the tissue inhibitor of metalloproteinase 1. Our results provide evidence that matrix metalloproteinases are activated in lung lesions from cattle with shipping fever and that M. haemolytica virulence products induce production, release, and especially activation of gelatinase B by bovine inflammatory cells in vitro. PMID- 15271896 TI - Sequence and binding activity of the autolysin-adhesin Ami from epidemic Listeria monocytogenes 4b. AB - Ami is an autolytic amidase from Listeria monocytogenes that is targeted to the bacterial surface via its C-terminal cell wall anchoring (CWA) domain. We recently showed that the CWA domain from Ami of L. monocytogenes EGD (serovar 1/2a) (Ami 1/2a) mediated bacterial binding to mammalian cells. Here we studied the sequence and binding properties of Ami from CHUT 82337 (serovar 4b) (Ami 4b). The Ami 4b polypeptide is predicted to be 770 amino acids long (compared with the 917 amino acids of Ami 1/2a from EGD). Ami 1/2a and Ami 4b are almost identical in the N-terminal enzymatic domain (approximately 98% amino acid identity), but the sequence is poorly conserved in the C-terminal CWA domain, with only approximately 54% amino acid identity and eight GW modules in Ami 1/2a compared with six GW modules in Ami 4b. The purified Ami 4b CWA domain efficiently bound serovar 4b bacterial cells and only poorly bound serovar 1/2a bacterial cells. The Ami 4b CWA domain was also significantly less able to bind Hep-G2 human hepatocytic cells than the Ami 1/2a CWA domain. We sequenced the ami regions encoding CWA domains of reference strains belonging to the 12 L. monocytogenes serovars. The phylogenic tree constructed from the sequences yielded a binary division into group I (serovars 1/2a, 1/2b, 1/2c, 3a, 3b, 3c, and 7) and group II (serovars 4a, 4b, 4c, 4d, and 4e). This is the first direct evidence of divergence between serovars 1/2a and 4b in a gene involved in the adhesion of L. monocytogenes to mammalian cells, as well as the first demonstration of allelic polymorphism correlated with the somatic antigen in this species. PMID- 15271897 TI - Analysis of serum cytokines in patients with severe acute respiratory syndrome. AB - Severe acute respiratory syndrome (SARS) is an acute infectious disease of the respiratory system. Although a novel coronavirus has been identified as the causative agent of SARS, the pathogenic mechanisms of SARS are not understood. In this study, sera were collected from healthy donors, patients with SARS, patients with severe SARS, and patients with SARS in convalescence. The serum concentrations of interleukin-1 (IL-1), IL-4, IL-6, IL-8, IL-10, tumor growth factor beta (TGF-beta), tumor necrosis factor alpha (TNF-alpha), and gamma interferon (IFN-gamma) were measured by enzyme-linked immunosorbent assays (ELISA). The concentrations of IL-1 and TNF-alpha were not significantly different in different groups. The IL-6 concentration was increased in SARS patients and was significantly elevated in severe SARS patients, but the IL-6 concentrations were similar in convalescent patients and control subjects, suggesting that there was a positive relationship between the serum IL-6 concentration and SARS severity. The concentrations of IL-8 and TGF-beta were decreased in SARS patients and significantly reduced in severe SARS patients, but they were comparable in convalescent SARS patients and control subjects, suggesting that there was a negative relationship between the IL-8 and TGF-beta concentrations and SARS severity. The concentrations of IFN-gamma, IL-4, and IL 10 showed significant changes only in convalescent SARS patients. The IFN-gamma and IL-4 levels were decreased, while the levels of IL-10 were increased, and the differences between convalescent SARS patients and other patient groups were statistically significant. These results suggest that there are different immunoregulatory events during and after SARS and may contribute to our understanding of the pathogenesis of this syndrome. PMID- 15271898 TI - Induction of maturation and cytokine release of human dendritic cells by Helicobacter pylori. AB - Helicobacter pylori causes a persistent infection in the human stomach, which can result in chronic gastritis and peptic ulcer disease. Despite an intensive proinflammatory response, the immune system is not able to clear the organism. However, the immune escape mechanisms of this common bacterium are not well understood. We investigated the interaction between H. pylori and human dendritic cells. Dendritic cells (DCs) are potent antigen-presenting cells and important mediators between the innate and acquired immune system. Stimulation of DCs with different concentrations of H. pylori for 8, 24, 48, and 72 h resulted in dose dependent interleukin-6 (IL-6), IL-8, IL-10 and IL-12 production. Lipopolysaccharide (LPS) from Escherichia coli, a known DC maturation agent, was used as a positive control. The cytokine release after stimulation with LPS was comparable to that induced by H. pylori except for IL-12. After LPS stimulation IL-12 was only moderately released compared to the large amounts of IL-12 induced by H. pylori. We further investigated the potential of H. pylori to induce maturation of DCs. Fluorescence-activated cell sorting analysis of cell surface expression of maturation marker molecules such as CD80, CD83, CD86, and HLA-DR revealed equal upregulation after stimulation with H. pylori or LPS. We found no significant differences between H. pylori seropositive and seronegative donors of DCs with regard to cytokine release and upregulation of surface molecules. These data clearly demonstrate that H. pylori induces a strong activation and maturation of human immature DCs. PMID- 15271899 TI - Effects of the Enterococcus faecalis hypR gene encoding a new transcriptional regulator on oxidative stress response and intracellular survival within macrophages. AB - In order to identify regulators of the oxidative stress response in Enterococcus faecalis, an important human pathogen, several genes annotated as coding for transcriptional regulators were inactivated by insertional mutagenesis. One mutant, affected in the ef2958 locus (designated hypR [hydrogen peroxide regulator]), appeared to be highly sensitive to oxidative challenge caused by hydrogen peroxide. Moreover, testing of the hypR mutant by using an in vivo-in vitro macrophage infection model resulted in a highly significant reduction in survival compared to the survival of parent strain JH2-2. Northern blot analyses were carried out with probes specific for genes encoding known antioxidant enzymes, and they showed that the ahpCF (alkyl hydroperoxide reductase) transcript was expressed less in mutant cells. Mobility shift protein-DNA binding assays revealed that HypR regulated directly the expression of hypR itself and the ahpCF operon. Our combined results showed that HypR appeared to be directly involved in the expression of ahpCF genes under oxidative stress conditions and suggested that this regulator could contribute to the virulence of E. faecalis. PMID- 15271900 TI - Gamma interferon production, but not perforin-mediated cytolytic activity, of T cells is required for prevention of toxoplasmic encephalitis in BALB/c mice genetically resistant to the disease. AB - We previously showed the requirement of both T cells and gamma interferon (IFN gamma)-producing non-T cells for the genetic resistance of BALB/c mice to the development of toxoplasmic encephalitis (TE). In order to define the role of IFN gamma production and the perforin-mediated cytotoxicity of T cells in this resistance, we obtained immune T cells from spleens of infected IFN-gamma knockout (IFN-gamma(-/-)), perforin knockout (PO), and wild-type BALB/c mice and transferred them into infected and sulfadiazine-treated athymic nude mice, which lack T cells but have IFN-gamma-producing non-T cells. Control nude mice that had not received any T cells developed severe TE and died after discontinuation of sulfadiazine treatment due to the reactivation of infection. Animals that had received immune T cells from either wild-type or PO mice did not develop TE and survived. In contrast, nude mice that had received immune T cells from IFN-gamma( /-) mice developed severe TE and died as early as control nude mice. T cells obtained from the spleens of animals that had received either PO or wild-type T cells produced large amounts of IFN-gamma after stimulation with Toxoplasma gondii antigens in vitro. In addition, the amounts of IFN-gamma mRNA expressed in the brains of PO T-cell recipients did not differ from those in wild-type T-cell recipients. Furthermore, PO mice did not develop TE after infection, and their IFN-gamma production was equivalent to or higher than that of wild-type animals. These results indicate that IFN-gamma production, but not perforin-mediated cytotoxic activity, by T cells is required for the prevention of TE in genetically resistant BALB/c mice. PMID- 15271901 TI - Mouse susceptibility to anthrax lethal toxin is influenced by genetic factors in addition to those controlling macrophage sensitivity. AB - Bacillus anthracis lethal toxin (LT) produces symptoms of anthrax in mice and induces rapid lysis of macrophages (M phi) derived from certain inbred strains. We used nine inbred strains and two inducible nitric oxide synthase (iNOS) knockout C57BL/6J strains polymorphic for the LT M phi sensitivity Kif1C locus to analyze the role of M phi sensitivity (to lysis) in LT-mediated cytokine responses and lethality. LT-mediated induction of cytokines KC, MCP-1/JE, MIP-2, eotaxin, and interleukin-1 beta occurred only in mice having LT-sensitive M phi. However, while iNOS knockout C57BL/6J mice having LT-sensitive M phi were much more susceptible to LT than the knockout mice with LT-resistant M phi, a comparison of susceptibilities to LT in the larger set of inbred mouse strains showed a lack of correlation between M phi sensitivity and animal susceptibility to toxin. For example, C3H/HeJ mice, harboring LT-sensitive M phi and having the associated LT-mediated cytokine response, were more resistant than mice with LT resistant M phi and no cytokine burst. Toll-like receptor 4 (Tlr4)-deficient, lipopolysaccharide-nonresponsive mice were not more resistant to LT. We also found that CAST/Ei mice are uniquely sensitive to LT and may provide an economical bioassay for toxin-directed therapeutics. The data indicate that while the cytokine response to LT in mice requires M phi lysis and while M phi sensitivity in the C57BL/6J background is sufficient for BALB/cJ-like mortality of that strain, the contribution of M phi sensitivity and cytokine response to animal susceptibility to LT differs among other inbred strains. Thus, LT-mediated lethality in mice is influenced by genetic factors in addition to those controlling M phi lysis and cytokine response and is independent of Tlr4 function. PMID- 15271902 TI - The major subunit of the toxin-coregulated pilus TcpA induces mucosal and systemic immunoglobulin A immune responses in patients with cholera caused by Vibrio cholerae O1 and O139. AB - Diarrhea caused by Vibrio cholerae is known to give long-lasting protection against subsequent life-threatening illness. The serum vibriocidal antibody response has been well studied and has been shown to correlate with protection. However, this systemic antibody response may be a surrogate marker for mucosal immune responses to key colonization factors of this organism, such as the toxin coregulated pilus (TCP) and other factors. Information regarding immune responses to TCP, particularly mucosal immune responses, is lacking, particularly for patients infected with the El Tor biotype of V. cholerae O1 or V. cholerae O139 since highly purified TcpA from these strains has not been available previously for use in immune assays. We studied the immune responses to El Tor TcpA in cholera patients in Bangladesh. Patients had substantial and significant increases in TcpA-specific antibody-secreting cells in the circulation on day 7 after the onset of illness, as well as similar mucosal responses as determined by an alternate technique, the assay for antibody in lymphocyte supernatant. Significant increases in antibodies to TcpA were also seen in sera and feces of patients on days 7 and 21 after the onset of infection. Overall, 93% of the patients showed a TcpA-specific response in at least one of the specimens compared with the results obtained on day 2 and with healthy controls. These results demonstrate that TcpA is immunogenic following natural V. cholerae infection and suggest that immune responses to this antigen should be evaluated for potential protection against subsequent life-threatening illness. PMID- 15271903 TI - CD4+ Th1 cells induced by dendritic cell-based immunotherapy in mice chronically infected with Leishmania amazonensis do not promote healing. AB - The susceptibility of mice to Leishmania amazonensis infection is thought to result from an inability to develop a Th1 response. Our data show that the low levels of gamma interferon (IFN-gamma) produced by the draining lymph node (DLN) cells of chronically infected mice could be enhanced in vitro and in vivo with L. amazonensis antigen-pulsed bone marrow-derived dendritic cells (BM-DC) and the Th1-promoting cytokine interleukin-12 (IL-12). Given intralesionally to chronically infected mice, this treatment induced the upregulation of mRNA levels for IFN-gamma, the transcription factor T-box expressed in T cells, and IL-12 receptor beta 2 in CD4(+) T cells from the DLN and an increase in parasite specific immunoglobulin G2a in the serum. However, this Th1 response was not associated with healing, and the antigen-specific enhancement of IFN-gamma production remained impaired in the DLN. However, addition of IL-12 to the in vitro recall response was able to recover this defect, suggesting that antigen presenting cell-derived IL-12 production may be limited in infected mice. This was supported by the fact that L. amazonensis amastigotes limited the production of IL-12p40 from BM-DC in vitro. Altogether, our data indicate that the immune response of mice chronically infected with L. amazonensis can be enhanced towards a Th1 phenotype but that the presence of Th1 CD4(+) T cells does not promote healing. This suggests that the phenotype of the CD4(+) T cells may not always be indicative of protection to L. amazonensis infection. Furthermore, our data support growing evidence that antigen-presenting cell function, such as IL-12 production, may limit the immune response in L. amazonensis-infected mice. PMID- 15271904 TI - Production of the subdomains of the Plasmodium falciparum apical membrane antigen 1 ectodomain and analysis of the immune response. AB - The apical membrane antigen 1 of Plasmodium falciparum is one of the leading candidate antigens being developed as a vaccine to prevent malaria. This merozoite transmembrane protein has an ectodomain that can be divided into three subdomains (D I, D II, and D III). We have previously expressed a major portion of this ectodomain and have shown that it can induce antibodies that prevent merozoite invasion into red blood cells in an in vitro growth and invasion assay. To analyze the antibody responses directed against the individual subdomains, we constructed six different genes that express each of the domains separately (D I, D II, or D III) or in combination with another domain (D I+II, D II+III, or D I+III). These proteins were purified and used to immunize rabbits to raise construct-specific antibodies. We demonstrated that D I+II induced a significant amount of the growth-inhibitory antibodies active in the growth and invasion assay. PMID- 15271905 TI - Quantitative gene expression profiling implicates genes for susceptibility and resistance to alveolar bone loss. AB - Periodontal disease is one of the most prevalent chronic inflammatory diseases. There is a genetic component to susceptibility and resistance to this disease. Using a mouse model, we investigated the progression of alveolar bone loss by gene expression profiling of susceptible and resistant mouse strains (BALB/cByJ and A/J, respectively). We employed a novel and sensitive quantitative real-time PCR method to compare basal RNA transcription of a 48-gene set in the gingiva and the spleen and the subsequent changes in gene expression due to Porphyromonas gingivalis oral infection. Basal expression of interleukin-1 beta (Il1b) and tumor necrosis factor alpha (Tnf) mRNA was higher in the gingiva of the susceptible BALB/cByJ mice than in the gingiva of resistant A/J mice. Gingival Il1b gene expression increased further and Stat6 gene expression was turned on after P. gingivalis infection in BALB/cByJ mice but not in A/J mice. The basal expression of interleukin-15 (Il15) in the gingiva and the basal expression of p selectin (Selp) in the spleen were higher in the resistant A/J mice than in the susceptible BALB/cByJ mice. In the resistant A/J mice the expression of no genes detectably changed in the gingiva after infection. These results suggest a molecular phenotype in which discrete sets of differentially expressed genes are associated with genetically determined susceptibility (Il1b, Tnf, and Stat6) or resistance (Il15 and Selp) to alveolar bone loss, providing insight into the genetic etiology of this complex disease. PMID- 15271906 TI - Helicobacter pylori induces apoptosis of human monocytes but not monocyte-derived dendritic cells: role of the cag pathogenicity island. AB - Monocytes are circulating precursors of the dendritic cell subset, professional antigen-presenting cells with a unique ability to initiate the innate and adaptive immune response. In this study, we have investigated the effects of wild type Helicobacter pylori strains and their isogenic mutants with mutations in known bacterial virulence factors on monocytes and monocyte-derived dendritic cells. We show that H. pylori strains induce apoptosis of human monocytes by a mechanism that is dependent on the expression of a functional cag pathogenicity island. This effect requires an intact injection organelle for direct contact between monocytes and the bacteria but also requires a still-unidentified effector that is different from VacA or CagA. The exposure of in vitro-generated monocyte-derived dendritic cells to H. pylori stimulates the release of inflammatory cytokines by a similar mechanism. Of note is that dendritic cells are resistant to H. pylori-induced apoptosis. These phenomena may play a critical role in the evasion of the immune response by H. pylori, contributing to the persistence of the infection. PMID- 15271907 TI - Characterization of an I-E-restricted, gp63-specific, CD4-T-cell clone from Leishmania major-resistant C3H mice that secretes type 2 cytokines and exacerbates infection with L. major. AB - A T-cell clone (designated KLmB-3) was derived from resistant C3H mice 2 weeks after infection with Leishmania major. KLmB-3 was a CD4-T-cell clone that utilized the V beta 8.1 T-cell receptor. When adoptively transferred to naive C3H mice, KLmB-3 unexpectedly exacerbated infection with L. major (it increased the cutaneous lesion size and the parasite burden within the lesion). The ability of KLmB-3 to exacerbate disease correlated with its ability to produce the type 2 associated cytokines interleukin-4 (IL-4), IL-5, IL-10, and transforming growth factor beta. Interestingly, KLmB-3 was specific for an epitope in the amino terminal end of the L. major surface gp63 zinc metalloproteinase (leishmanolysin) that has been shown to be capable of inducing a protective immune response. Moreover, KLmB-3 was activated when this epitope was presented in the context of H-2 I-E rather than H-2 I-A. PMID- 15271908 TI - Immunostimulatory CpG oligodeoxynucleotide confers protection in a murine model of infection with Burkholderia pseudomallei. AB - Although CpG oligodeoxynucleotides (CpG ODNs) are known to enhance resistance against infection in a number of animal models, little is known about the CpG induced protection against acute fatal sepsis such as that associated with the highly virulent bacterium Burkholderia pseudomallei. We previously demonstrated in an in vitro study that immunostimulatory CpG ODN 1826 enhances phagocytosis of B. pseudomallei and induces nitric oxide synthase and nitric oxide production by mouse macrophages. In the present study, CpG ODN 1826 given intramuscularly to BALB/c mice 2 to 10 days prior to B. pseudomallei challenge conferred better than 90% protection. CpG ODN 1826 given 2 days before the bacterial challenge rapidly enhanced the innate immunity of these animals, judging from the elevated serum levels of interleukin-12 (IL-12)p70 and gamma interferon (IFN-gamma) over the baseline values. No bacteremia was detected on day 2 in 85 to 90% of the CpG treated animals, whereas more than 80% of the untreated animals exhibited heavy bacterial loads. Although marked elevation of IFN-gamma was found consistently in the infected animals 2 days after the bacterial challenge, it was ameliorated by the CpG ODN 1826 pretreatment (P = 0.0002). Taken together, the kinetics of bacteremia and cytokine profiles presented are compatible with the possibility that protection by CpG ODN 1826 against acute fatal septicemic melioidosis in this animal model is associated with a reduction of bacterial load and interference with the potential detrimental effect of the robust production of proinflammatory cytokines associated with B. pseudomallei multiplication. PMID- 15271909 TI - A specific genomic location within the icm/dot pathogenesis region of different Legionella species encodes functionally similar but nonhomologous virulence proteins. AB - Legionella pneumophila, the major causative agent of Legionnaires' disease, is a facultative intracellular pathogen that grows within human macrophages and amoebae. Intracellular growth involves the formation of a replicative phagosome that requires the Icm/Dot type IV secretion system. Part of the icm/dot region in L. pneumophila contains the icmTSRQPO genes. The proteins encoded by the icmR and icmQ genes were shown to exhibit a chaperone-substrate relationship. Analysis of this region from other pathogenic Legionella species, i.e., L. micdadei and L. longbeachae, indicated that the overall organization of this region is highly conserved, but it was found to contain a favorable site for gene variation. In the place where the icmR gene was expected to be located, other open reading frames that are nonhomologous to each other or to any entry in the GenBank database were found (migAB in L. micdadei and ligB in L. longbeachae). Examination of these unique genes revealed an outstanding phenomenon; by use of interspecies complementation, the icmR, migB, and ligB gene products were found to be functionally similar. In addition, the function of these proteins was usually dependent on the presence of the corresponding IcmQ proteins. Moreover, each of these proteins (IcmR, LigB, and MigB) was found to interact with the corresponding IcmQ proteins, and the genes encoding these proteins were found to be regulated by CpxR. This study reveals new evidence of gene variation occurring in the same genomic location within the icm/dot locus in various Legionella species. The genes found at this site were shown to be similarly regulated and to encode species-specific, nonhomologous, but functionally similar proteins. PMID- 15271910 TI - Cytotoxicity of hydrogen peroxide produced by Enterococcus faecium. AB - Although the opportunistic bacterial pathogen Enterococcus faecium is a leading source of nosocomial infections, it appears to lack many of the overt virulence factors produced by other bacterial pathogens, and the underlying mechanism of pathogenesis is not clear. Using E. faecium-mediated killing of the nematode worm Caenorhabditis elegans as an indicator of toxicity, we determined that E. faecium produces hydrogen peroxide at levels that cause cellular damage. We identified E. faecium transposon insertion mutants with altered C. elegans killing activity, and these mutants were altered in hydrogen peroxide production. Mutation of an NADH oxidase-encoding gene eliminated nearly all NADH oxidase activity and reduced hydrogen peroxide production. Mutation of an NADH peroxidase-encoding gene resulted in the enhanced accumulation of hydrogen peroxide. E. faecium is able to produce hydrogen peroxide by using glycerol-3-phosphate oxidase, and addition of glycerol to the culture medium enhanced the killing of C. elegans. Conversely, addition of glucose, which leads to the down-regulation of glycerol metabolism, prevented both C. elegans killing and hydrogen peroxide production. Lastly, detoxification of hydrogen peroxide either by exogenously added catalase or by a C. elegans transgenic strain overproducing catalase prevented E. faecium mediated killing. These results suggest that hydrogen peroxide produced by E. faecium has cytotoxic effects and highlight the utility of C. elegans pathogenicity models for identifying bacterial virulence factors. PMID- 15271911 TI - Intranasal vaccination against cutaneous leishmaniasis with a particulated leishmanial antigen or DNA encoding LACK. AB - We have previously demonstrated that oral delivery of a disease-promoting particulated antigen of Leishmania amazonensis (LaAg) partially protects mice against cutaneous leishmaniasis. In the present work, we sought to optimize a mucosal vaccine by using the intranasal route for delivery of different antigen preparations, including (i) LaAg, (ii) soluble recombinant p36/LACK leishmanial antigen (LACK), and (iii) plasmid DNA encoding LACK (LACK DNA). BALB/c mice that received two intranasal doses of 10 microg of LaAg and were challenged 1 week postvaccination with L. amazonensis developed delayed but effective control of lesion growth. A diminished parasite burden was accompanied by enhancement of both gamma interferon (IFN-gamma) and interleukin-10 levels in the lesion draining lymph nodes. The vaccine efficacy improved with time. At 4 months postvaccination, when a strong parasite-specific TH1-type response was present in vivo, the infection was controlled for at least 5 months after challenge. In contrast to the particulated LaAg, soluble LACK (10 microg/dose) had no effect. Interestingly, LACK DNA (30 microg/dose), but not empty DNA, promoted rapid and durable protective immunity. Parasite growth was effectively controlled, and at 5 months after challenge LACK-reactive cells in both the mucosal and lesion draining lymph nodes produced high levels of IFN-gamma. These results demonstrate for the first time the feasibility of using the intranasal route for long-lived memory vaccination against cutaneous leishmaniasis with adjuvant-free crude antigens or DNA. PMID- 15271912 TI - Expression of the LspA1 and LspA2 proteins by Haemophilus ducreyi is required for virulence in human volunteers. AB - Haemophilus ducreyi colocalizes with polymorphonuclear leukocytes and macrophages and evades phagocytosis during experimental infection of human volunteers. H. ducreyi contains two genes, lspA1 and lspA2, which encode predicted proteins of 456 and 543 kDa, respectively. Compared to its wild-type parent, an lspA1 lspA2 double mutant does not inhibit phagocytosis by macrophage and myelocytic cell lines in vitro and is attenuated in an experimental rabbit model of chancroid. To test whether expression of LspA1 and LspA2 was necessary for virulence in humans, six volunteers were experimentally infected. Each volunteer was inoculated with three doses (ranging from 85 to 112 CFU) of the parent (35000HP) in one arm and three doses (ranging from 60 to 822 CFU) of the mutant (35000HP Omega 12) in the other arm. The papule formation rates were 88% (95% confidence interval [95% CI], 76.8 to 99.9%) at 18 parent sites and 72% (95% CI, 44.4 to 99.9%) at 18 mutant sites (P = 0.19). However, papules were significantly smaller at mutant sites (mean size, 24.8 mm(2)) than at parent sites (mean size, 39.1 mm(2)) 24 h after inoculation (P = 0.0002). The pustule formation rates were 44% (95% CI, 5.8 to 77.6%) at parent sites and 0% (95% CI, 0 to 39.4%) at mutant sites (P = 0.009). With the caveat that biosafety regulations preclude testing of a complemented mutant in human subjects, these results indicate that expression of LspA1 and LspA2 facilitates the ability of H. ducreyi to initiate disease and to progress to pustule formation in humans. PMID- 15271913 TI - Protection against pneumococcal pneumonia in mice by monoclonal antibodies to pneumolysin. AB - Pneumolysin (PLY) is an important virulence factor of Streptococcus pneumoniae. We examined the ability of three murine monoclonal antibodies (MAbs) to PLY (PLY 4, PLY-5, and PLY-7) to affect the course of pneumococcal pneumonia in mice. The intravenous administration of antibodies PLY-4 and PLY-7 protected the mice from the lethal effect of the purified toxin. Mice treated with PLY-4 before intranasal inoculation of S. pneumoniae type 2 survived longer (median survival time, 100 h) than did untreated animals (median survival time, 60 h) (P < 0.0001). The median survival time for mice treated with a combination of PLY-4 and PLY-7 was 130 h, significantly longer than that for mice given isotype matched indifferent MAbs (P = 0.0288) or nontreated mice (P = 0.0002). The median survival time for mice treated with a combination of three MAbs was significantly longer (>480 h) than that for mice treated with PLY-5 (48 h; P < 0.0001), PLY-7 (78 h; P = 0.0007), or PLY-4 (100 h; P = 0.0443) alone. Similarly, the survival rate for mice treated with three MAbs (10 of 20 mice) was significantly higher than the survival rate obtained with PLY-5 (1 of 20; P = 0.0033), PLY-4 (2 of 20; P = 0.0138), or PLY-7 (3 of 20; P = 0.0407) alone. These results suggest that anti-PLY MAbs act with a synergistic effect. Furthermore, MAb administration was associated with a significant decrease in bacterial lung colonization and lower frequencies of bacteremia and tissue injury with respect to the results for the control groups. PMID- 15271914 TI - Icm/dot-independent entry of Legionella pneumophila into amoeba and macrophage hosts. AB - Legionella pneumophila, the causative agent of Legionnaires' disease, expresses a type IVB secretion apparatus that translocates bacterial proteins into amoeba and macrophage hosts. When stationary-phase cultures are used to infect hosts, the type IVB apparatus encoded by the icm/dot genes is required for entry, delay of phagosome-lysosome fusion, and intracellular multiplication within host cells. Null mutants with mutations in icm/dot genes are defective in these phenotypes. Here a new model is described in which hosts are infected with stationary-phase cultures that have been incubated overnight in pH 6.5 buffer. This model is called Ers treatment because it enhances the resistance to acid, hydrogen peroxide, and antibiotic stress beyond that of stationary-phase cultures. Following Ers treatment entry into amoeba and macrophage hosts does not require dotA, which is essential for Legionella virulence phenotypes when hosts are infected with stationary-phase cultures, dotB, icmF, icmV, or icmX. Defective host entry is also suppressed for null mutants with mutations in the KatA and KatB catalase-peroxidase enzymes, which are required for proper intracellular growth in amoeba and macrophage hosts. Ers treatment-induced suppression of defective entry is not associated with increased bacterial adhesion to host cells or with morphological changes in the bacterial envelope but is dependent on protein expression during Ers treatment. By using proteomic analysis, Ers treatment was shown to induce a protein predicted to contain eight tetratricopeptide repeats, a motif previously implicated in enhanced entry of L. pneumophila. Characterization of Ers treatment-dependent changes in expression is proposed as an avenue for identifying icm/dot-independent factors that function in the entry of Legionella into amoeba and macrophage hosts. PMID- 15271915 TI - A STAT4-dependent Th1 response is required for resistance to the helminth parasite Taenia crassiceps. AB - To determine the role of STAT4-dependent Th1 responses in the regulation of immunity to the helminth parasite Taenia crassiceps, we monitored infections with this parasite in resistant mice lacking the STAT4 gene. While T. crassiceps infected STAT4(+/+) mice rapidly resolved the infection, STAT4(-/-) mice were highly susceptible to infection and displayed large parasite loads. Moreover, the inability of STAT4(-/-) mice to control the infection was associated with the induction of an antigen-specific Th2-type response characterized by significantly higher levels of Th2-associated immunoglobulin G1 (IgG1) and total IgE as well as interleukin-4 (IL-4), IL-10, and IL-13 than those in STAT4(+/+) mice, who produced significantly more gamma interferon. Furthermore, early after infection, macrophages from STAT4(-/-) mice produced lower levels of the pro-inflammatory cytokines IL-12, tumor necrosis factor alpha, IL-1 beta, and nitric oxide (NO) than those from STAT4(+/+) mice, suggesting a pivotal role for macrophages in mediating protection against cysticercosis. These findings demonstrate a critical role for the STAT4 signaling pathway in the development of a Th1-type immune response that is essential for mediating protection against the larval stage of T. crassiceps infection. PMID- 15271916 TI - Toll-like receptor 2 represses nonpilus adhesin-induced signaling in acute infections with the Pseudomonas aeruginosa pilA mutant. AB - Expression of pili and associated proteins is an important means of host invasion by bacterial pathogens. Recent evidence has suggested that the binding of Pseudomonas aeruginosa through nonpilus adhesins may also be important in respiratory diseases, since adhesins bind mucins. Using wild-type C57BL/6 and TLR2KO mice, we compared the induction levels of the host response to P. aeruginosa that either expressed pili or lacked pilus expression due to a mutation in the structural gene pilA. In C57BL/6 mice, deletion of pili led to a decreased immune response, evidenced by a lower secretion of cytokines and a lack of neutrophil chemotaxis. By contrast, the P. aeruginosa pilA mutant induced a hyperresponsive phenotype in TLR2KO mice. TLR2KO mice showed an increased number of neutrophils in lavage fluid compared to the levels seen when either mouse strain was exposed to wild-type P. aeruginosa. Further analysis indicated that the increased neutrophil influx was associated with an increased expression of calgranulins, possibly through an induction of Toll-like receptor 4 (TLR4) expression. The hyperresponsive phenotype of TLR2KO mice exposed to the P. aeruginosa pilA mutant was associated with TLR4 induction and indicated that nonpilus adhesin-induced signaling was repressed by TLR2 function and, if not blocked by the host, could induce airway hyperresponsiveness. PMID- 15271917 TI - Uropathogenic Escherichia coli triggers oxygen-dependent apoptosis in human neutrophils through the cooperative effect of type 1 fimbriae and lipopolysaccharide. AB - Type 1 fimbriae are the most commonly expressed virulence factor on uropathogenic Escherichia coli. In addition to promoting avid bacterial adherence to the uroepithelium and enabling colonization, type 1 fimbriae recruit neutrophils to the urinary tract as an early inflammatory response. Using clinical isolates of type 1 fimbriated E. coli and an isogenic type 1 fimbria-negative mutant (CN1016) lacking the FimH adhesin, we investigated if these strains could modulate apoptosis in human neutrophils. We found that E. coli expressing type 1 fimbriae interacted with neutrophils in a mannose- and lipopolysaccharide (LPS)-dependent manner, leading to apoptosis which was triggered by the intracellular generation of reactive oxygen species. This induced neutrophil apoptosis was abolished by blocking FimH-mediated attachment, by inhibiting NADPH oxidase activation, or by neutralizing LPS. In contrast, CN1016, which did not adhere to or activate the respiratory burst of neutrophils, delayed the spontaneous apoptosis in an LPS dependent manner. This delayed apoptosis could be mimicked by adding purified LPS and was also observed by using fimbriated bacteria in the presence of d-mannose. These results suggest that LPS is required for E. coli to exert both pro- and antiapoptotic effects on neutrophils and that the difference in LPS presentation (i.e., with or without fimbriae) determines the outcome. The present study showed that there is a fine-tuned balance between type 1 fimbria-induced and LPS mediated delay of apoptosis in human neutrophils, in which altered fimbrial expression on uropathogenic E. coli determines the neutrophil survival and the subsequent inflammation during urinary tract infections. PMID- 15271918 TI - phgABC, a three-gene operon required for growth of Streptococcus pneumoniae in hyperosmotic medium and in vivo. AB - To cause disease, bacterial pathogens need to be able to adapt to the physiological conditions found within the host, including an osmolality of approximately 290 mosmol kg(-1). While investigating Streptococcus pneumoniae genes contained within pneumococcal pathogenicity island 1, we identified a three gene operon of unknown function termed phgABC. PhgC has a domain with similarity to diacylglycerol kinases of eukaryotes and is the first described member of a family of related proteins found in many gram-positive bacteria. phgA and phgC mutant strains were constructed by insertional duplication mutagenesis and found to have impaired growth under conditions of high osmotic and oxidative stress. The compatible solutes proline and glycine betaine improved growth of the wild type and the phgA mutant strains in hyperosmolar medium, and when analyzed by electron microscopy, the cellular morphology of the phgA mutant strain was unaffected by osmotic stress. The phgA and phgC mutant strains were reduced in virulence in models of both systemic and pulmonary infection. As the virulence of the phgA mutant strain was not restored in gp91phox(-/-) mice and the phgA and phgC mutant strains had reduced growth in both blood and serum, the reduced virulence of these strains is unlikely to be due to increased sensitivity to the respiratory burst of phagocytes but is, instead, due to impaired growth at physiological osmolality. PMID- 15271919 TI - The plague virulence protein YopM targets the innate immune response by causing a global depletion of NK cells. AB - Yersinia pestis, the etiologic agent of plague, delivers six Yersinia outer proteins (Yops) into host cells upon direct bacterial contact. One of these, YopM, is necessary for virulence in a mouse model of septicemic plague, but its pathogenic function is unknown. We report here the immune processes affected by YopM during infection. To test whether the innate or adaptive immune system is targeted by YopM, C57BL/6 (B6) and B6 SCID mice were infected with either the conditionally virulent Y. pestis KIM5 or a yopM deletion mutant and evaluated for bacterial growth in spleen and liver. Both B6 and SCID mice succumbed to infection with Y. pestis KIM5, whereas both mouse strains survived infection by the YopM(-) mutant. These data showed that YopM counteracts innate defenses present in SCID mice. The YopM(-) strain grew more slowly than the parent Y. pestis during the first 4 days of infection in both mouse strains, indicating an early pathogenic role for YopM. In B6 mice, populations of cells of the immune system were not differentially affected by the two Y. pestis strains, with one major exception: the parent Y. pestis KIM5 but not the YopM(-) mutant caused a significant global decrease in NK cell numbers (blood, spleen, and liver), beginning early in infection. NK cells and macrophages isolated early (day 2) from livers and spleens of mice infected with either Y. pestis strain contained comparable levels of cytokine mRNA: interleukin (IL)-1 beta, IL-12, IL-15, IL-18, and tumor necrosis factor alpha in macrophages and gamma interferon in NK cells. However, by day 4 postinfection, cells from mice infected with the parent Y. pestis expressed lower levels of these messages, while those from mice infected with the mutant retained strong expression. Significantly, mRNA for the IL-15 receptor alpha chain was not expressed in NK cells from Y. pestis KIM5-infected mice as early as day 2 postinfection. These findings suggest that YopM interferes with innate immunity by causing depletion of NK cells, possibly by affecting the expression of IL-15 receptor alpha and IL-15. PMID- 15271920 TI - Inbred strains derived from feral mice reveal new pathogenic mechanisms of experimental leishmaniasis due to Leishmania major. AB - Two inbred mouse strains, derived from feral founders, are susceptible to experimental leishmaniasis due to Leishmania major and support a disease of a severity intermediate between those observed in strains C57BL/6 and BALB/c. Mice of the MAI strain develop a severe, nonhealing, but nonfatal disease with no resistance to a secondary parasite challenge. The immunological responses showed a TH2 dominance characterized by an early peak of interleukin-4 (IL-4) and IL-13. However, neutralization of IL-4, which leads to a resistance phenotype in BALB/c mice, has no effect on disease progression in MAI mice. Mice of strain PWK develop a protracted but self-healing disease, characterized by a mixed TH1-plus TH2 pattern of immune responses in which IL-10 plays an aggravating role, and acquire resistance to a secondary challenge. These features are close to those observed in human cutaneous leishmaniasis due to L. major and make PWK mice a suitable model for the human disease. PMID- 15271921 TI - Gamma interferon production by bovine gamma delta T cells following stimulation with mycobacterial mycolylarabinogalactan peptidoglycan. AB - A large percentage of lymphocytes in the blood of cattle express the gamma delta T-cell receptor, but specific functions for these cells have not yet been clearly defined. There is evidence, however, that human, murine, and bovine gamma delta T cells have a role in the immune response to mycobacteria. This study investigated the ability of bovine gamma delta T cells to expand and produce gamma interferon (IFN-gamma) in response to stimulation with mycobacterial products. Bovine gamma delta T cells, isolated from the peripheral blood of healthy cattle, expanded following in vitro stimulation with live mycobacteria, mycobacterial crude cell wall extract, and Mycobacterium bovis culture filtrate proteins. In addition, purified gamma delta T cells, cocultured with purified monocytes and interleukin 2, consistently produced significant amounts of IFN-gamma in response to mycobacterial cell wall. The IFN-gamma-inducing component of the cell wall was further identified as a proteolytically resistant, non-sodium dodecyl sulfate soluble component of the mycolylarabinogalactan peptidoglycan. PMID- 15271922 TI - Multiple functions of the leucine-rich repeat protein LrrA of Treponema denticola. AB - The gene lrrA, encoding a leucine-rich repeat protein, LrrA, that contains eight consensus tandem repeats of 23 amino acid residues, has been identified in Treponema denticola ATCC 35405. A leucine-rich repeat is a generally useful protein-binding motif, and proteins containing this repeat are typically involved in protein-protein interactions. Southern blot analysis demonstrated that T. denticola ATCC 35405 expresses the lrrA gene, but the gene was not identified in T. denticola ATCC 33520. In order to analyze the functions of LrrA in T. denticola, an lrrA-inactivated mutant of strain ATCC 35405 and an lrrA gene expression transformant of strain ATCC 33520 were constructed. Characterization of the mutant and transformant demonstrated that LrrA is associated with the extracytoplasmic fraction of T. denticola and expresses multifunctional properties. It was demonstrated that the attachment of strain ATCC 35405 to HEp-2 cell cultures and coaggregation with Tannerella forsythensis were attenuated by the lrrA mutation. In addition, an in vitro binding assay demonstrated specific binding of LrrA to a portion of the Tannerella forsythensis leucine-rich repeat protein, BspA, which is mediated by the N-terminal region of LrrA. It was also observed that the lrrA mutation caused a reduction of swarming in T. denticola ATCC 35405 and consequently attenuated tissue penetration. These results suggest that the leucine-rich repeat protein LrrA plays a role in the attachment and penetration of human epithelial cells and coaggregation with Tannerella forsythensis. These properties may play important roles in the virulence of T. denticola. PMID- 15271923 TI - Pneumocystis carinii cell wall biosynthesis kinase gene CBK1 is an environmentally responsive gene that complements cell wall defects of cbk deficient yeast. AB - Pneumocystis species remain an important cause of life-threatening pneumonia in immunocompromised hosts, including those with AIDS. Responses of the organism to environmental cues both within the lung and elsewhere have been poorly defined. Herein, we report the identification of a cell wall biosynthesis kinase gene (CBK1) homologue in Pneumocystis carinii, isolated by differential display PCR, that is expressed optimally at physiological pH (7 to 8) as opposed to more acidic environments. Expression of Pneumocystis CBK1 was also induced by contact with lung epithelial cells and extracellular matrix. Translation of this gene revealed extensive homology to other fungal CBK1 kinases. Pneumocystis CBK1 expression was equal in the cyst and trophic life forms of the organisms. We further demonstrate that Pneumocystis CBK1 expressed in cbk1 Delta Saccharomyces cerevisiae cells restored defective cell wall separation during proliferation. Consistent with this, Pneumocystis CBK1 expression also stimulated transcription of the CTS1 chitinase in cbk1 Delta mutant yeast cells, an event necessary for cell wall separation. In addition, Pneumocystis CBK1 cDNA supported normal mating projection formation in response to alpha-factor in the cbk1 Delta yeast cells. Site-directed mutations of serine-303 and threonine-494, potential regulatory phosphorylation sites in Pneumocystis CBK1, abolished mating projection formation, indicating a role for these amino acid residues in CBK1 activity. These findings indicate that Pneumocystis CBK1 is an environmentally responsive gene that may function in signaling pathways necessary for cell growth and mating. PMID- 15271924 TI - Mucosally delivered Salmonella live vector vaccines elicit potent immune responses against a foreign antigen in neonatal mice born to naive and immune mothers. AB - The development of effective vaccines for neonates and very young infants has been impaired by their weak, short-lived, and Th-2 biased responses and by maternal antibodies that interfere with vaccine take. We investigated the ability of Salmonella enterica serovars Typhi and Typhimurium to mucosally deliver tetanus toxin fragment C (Frag C) as a model antigen in neonatal mice. We hypothesize that Salmonella, by stimulating innate immunity (contributing to adjuvant effects) and inducing Th-1 cytokines, can enhance neonatal dendritic cell maturation and T-cell activation and thereby prime humoral and cell-mediated immunity. We demonstrate for the first time that intranasal immunization of newborn mice with 10(9) CFU of S. enterica serovar Typhi CVD 908-htrA and S. enterica serovar Typhimurium SL3261 carrying plasmid pTETlpp on days 7 and 22 after birth elicits high titers of Frag C antibodies, previously found to protect against tetanus toxin challenge and similar to those observed in adult mice. Salmonella live vectors colonized and persisted primarily in nasal tissue. Mice vaccinated as neonates induced Frag C-specific mucosal and systemic immunoglobulin A (IgA)- and IgG-secreting cells, T-cell proliferative responses, and gamma interferon secretion. A mixed Th1- and Th2-type response to Frag C was established 1 week after the boost and was maintained thereafter. S. enterica serovar Typhi carrying pTETlpp induced Frag C-specific antibodies and cell mediated immunity in the presence of high levels of maternal antibodies. This is the first report that demonstrates the effectiveness of Salmonella live vector vaccines in early life. PMID- 15271925 TI - Attenuation of the bacterial load in blood by pretreatment with granulocyte colony-stimulating factor protects rats from fatal outcome and brain damage during Streptococcus pneumoniae meningitis. AB - A model of pneumococcal meningitis in young adult rats receiving antibiotics once the infection was established was developed. The intent was to mimic clinical and histopathological features of pneumococcal meningitis in humans. The primary aim of the present study was to evaluate whether medical boosting of the peripheral neutrophil count affected the outcome of the meningitis. The risk of terminal illness over the first 7 days after infection was significantly reduced for rats who had elevated peripheral white blood cell counts after receiving granulocyte colony-stimulating factor (G-CSF) prior to the infection compared to that for untreated rats (P = 0.039 by the log rank test). The improved outcome was associated with reduced signs of cerebral cortical damage (P = 0.008). Furthermore, the beneficial effects of G-CSF were associated with reduced bacterial loads in the cerebrospinal fluid (median, 1.1 x 10(5) versus 2.9 x 10(5) CFU/ml; P = 0.023) and in blood (median, 2.9 x 10(2) versus 6.3 x 10(2) CFU/ml; P = 0.024), as well as attenuated pleocytosis (median, 800 x 10(6) versus 1,231 x 10(6) cells/liter; P = 0.025), 24 h after the infection. Conversely, initiation of G-CSF therapy 28 h postinfection did not alter the clinical or histological outcome relative to that for non-G-CSF-treated rats. The magnitude of bacteremia and pretreatment with G-CSF were found to be prognostic factors for both outcome and brain damage. In summary, elevated neutrophil levels prior to the development of meningitis result in reduced risks of death and brain damage. This beneficial effect is most likely achieved through improved control of the systemic disease. PMID- 15271926 TI - Role of the two-component regulator CpxAR in the virulence of Salmonella enterica serotype Typhimurium. AB - The CpxAR (Cpx) two-component regulator controls the expression of genes in response to a variety of environmental cues. The Cpx regulator has been implicated in the virulence of several gram-negative pathogens, although a role for Cpx in vivo has not been demonstrated directly. Here we investigate whether positive or negative control of gene expression by Cpx is important for the pathogenesis of Salmonella enterica serotype Typhimurium. The Cpx signal pathway in serotype Typhimurium was disrupted by insertional inactivation of the cpxA and cpxR genes. We also constitutively activated the Cpx pathway by making an internal in-frame deletion in cpxA (a cpxA* mutation). Activation of the Cpx pathway inhibited induction of the envelope stress response pathway controlled by the alternative sigma factor sigma(E) (encoded by rpoE). Conversely, the Cpx pathway was highly up-regulated (>40-fold) in a serotype Typhimurium rpoE mutant. The cpxA* mutation, but not the cpxA or the cpxR mutation, significantly reduced the capacity of serotype Typhimurium to adhere to and invade eucaryotic cells, although intracellular replication was not affected. The cpxA and cpxA* mutations significantly impaired the ability of serotype Typhimurium to grow in vivo in mice. To our knowledge, this is the first demonstration that the Cpx system is important for a bacterial pathogen in vivo. PMID- 15271927 TI - Role of p38 mitogen-activated protein kinase in middle ear mucosa hyperplasia during bacterial otitis media. AB - Hyperplasia of the middle ear mucosa contributes to the sequelae of acute otitis media. Understanding the signal transduction pathways that mediate hyperplasia could lead to the development of new therapeutic interventions for this disease and its sequelae. Endotoxin derived from bacteria involved in middle ear infection can contribute to the hyperplastic response. The p38 mitogen-activated protein kinase (MAPK) is known to be activated by endotoxin as well as cytokines and other inflammatory mediators that have been documented in otitis media. We assessed the activation of p38 in the middle ear mucosa of an in vivo rat bacterial otitis media model. Strong activity of p38 was observed 1 to 6 h after bacterial inoculation. Activity continued at a lower level for at least 7 days. The effects of p38 activation were assessed using an in vitro model of rat middle ear mucosal hyperplasia in which mucosal growth is stimulated by nontypeable Haemophilus influenzae during acute otitis media. Hyperplastic mucosal explants treated with the p38 alpha and p38 beta inhibitor SB203580 demonstrated significant inhibition of otitis media-stimulated mucosal growth. The results of this study suggest that intracellular signaling via p38 MAPK influences the hyperplastic response of the middle ear mucosa during bacterial otitis media. PMID- 15271928 TI - Chronic Helicobacter pylori infection with Sydney strain 1 and a newly identified mouse-adapted strain (Sydney strain 2000) in C57BL/6 and BALB/c mice. AB - The mouse model of Helicobacter pylori-induced disease using Sydney strain 1 (SS1) has been used extensively in Helicobacter research. Herein we describe the isolation and characterization of a new mouse-colonizing strain for use in comparative studies. One strain capable of persistent mouse colonization was isolated from a total of 110 clinical isolates and is named here SS2000 (Sydney strain 2000). Genome typing revealed a number of differences between SS1 and SS2000 as well as between them and the respective original clinical isolates. In particular, SS2000 lacked the entire cag pathogenicity island, while SS1 contained all 27 genes of the island. C57BL/6 and BALB/c mice were infected with SS1 or SS2000 or were treated with broth medium (controls). After 6 months host specific effects were evident, including lower colonization levels in the BALB/c animals. Few pathological differences were observed between SS1- and SS2000 infected animals. However, by 15 months postinfection, SS1-infected C57BL/6 mice had developed more severe gastritis than the SS2000-infected animals. In contrast SS2000-infected BALB/c mice showed increased accumulation of mucosa-associated lymphoid tissue compared to those infected with SS1. This improved comparative model of H. pylori-induced disease allowed dissection of both host and strain effects and thus will prove useful in further studies. PMID- 15271929 TI - Characterization of the Streptococcus mutans P1 epitope recognized by immunomodulatory monoclonal antibody 6-11A. AB - Monoclonal antibody (MAb) 6-11A directed against Streptococcus mutans surface adhesin P1 was shown previously to influence the mucosal immunogenicity of this organism in BALB/c mice. The specificity of anti-P1 serum immunoglobulin G (IgG) and secretory IgA antibodies and the subclass distribution of anti-P1 serum IgG antibodies were altered, and the ability of elicited serum antibodies to inhibit S. mutans adherence in vitro was in certain cases increased. MAb 6-11A is known to recognize an epitope dependent on the presence of the proline-rich region of the protein, although it does not bind directly to the isolated P-region domain. In this report, we show that MAb 6-11A recognizes a complex discontinuous epitope that requires the simultaneous presence of the alanine-rich repeat domain (A region) and the P-region. Formation of the core epitope requires the interaction of these segments of P1. Residues amino terminal to the A-region also contributed to recognition by MAb 6-11A but were not essential for binding. Characterization of the MAb 6-11A epitope will enable insight into potential mechanisms of immunomodulation and broaden our understanding of the tertiary structure of P1. PMID- 15271930 TI - In vitro models of tissue penetration and destruction by Porphyromonas gingivalis. AB - Porphyromonas gingivalis is a gram-negative anaerobic bacterium that is considered the key etiologic agent of chronic periodontitis. Arg- and Lys gingipain cysteine proteinases produced by P. gingivalis are key virulence factors and are believed to be essential for significant tissue component degradation, leading to host tissue invasion by periodontopathogens. Two in vitro models were used to determine the extent to which P. gingivalis can reach connective tissue. The tissue penetration potential of P. gingivalis was first investigated by using an engineered human oral mucosa model composed of normal human epithelial cells and fibroblasts. Internalized bacteria were assessed by transmission electron microscopy. Bacteria were observed within multilayered gingival epithelial cells and in the space between the stratified epithelium and the lamina propria. A gingipain-null mutant strain of P. gingivalis was found to be less potent in penetrating tissue than the wild-type strain. Proinflammatory responses to P. gingivalis infection were evaluated. P. gingivalis increased the secretion of interleukin-1 beta, interleukin-6, interleukin-8, and tumor necrosis factor alpha. In the second part of the study, the contribution of P. gingivalis gingipains to tissue penetration was investigated by using a reconstituted basement membrane model (Matrigel). The penetration of (14)C-labeled P. gingivalis cells through Matrigel was significantly reduced when leupeptin, a specific inhibitor of Arg-gingipain activity, was added or when a gingipain-null mutant was used. The results obtained with these two relevant models support the capacities of P. gingivalis to infiltrate periodontal tissue and to modulate the proinflammatory response and suggest a critical role of gingipains in tissue destruction. PMID- 15271931 TI - Contribution of the alanine-rich region of Streptococcus mutans P1 to antigenicity, surface expression, and interaction with the proline-rich repeat domain. AB - Streptococcus mutans is considered to be the major etiologic agent of human dental caries. Attachment of S. mutans to the tooth surface is required for the development of caries and is mediated, in part, by the 185-kDa surface protein variously known as antigen I/II, PAc, and P1. Such proteins are expressed by nearly all species of oral streptococci. Characteristics of P1 include an alanine rich repeat region and a centrally located proline-rich repeat region. The proline-rich region of P1 has been shown to be important for the translational stability and translocation of P1 through the bacterial membrane. We show here that (i) several anti-P1 monoclonal antibodies require the simultaneous presence of the alanine-rich and proline-rich regions for binding, (ii) the proline-rich region of P1 interacts with the alanine-rich region, (iii) like the proline-rich region, the alanine-rich region is required for the stability and translocation of P1, (iv) both the proline-rich and alanine-rich regions are required for secretion of P1 in Escherichia coli, and (v) in E. coli, P1 is secreted in the absence of SecB. PMID- 15271932 TI - Novel phage display-based subtractive screening to identify vaccine candidates of Brugia malayi. AB - This study describes a novel phage display method based on an iterative subtraction strategy to identify candidate vaccine antigens of Brugia malayi. A cDNA library of the infective larval stage of B. malayi expressed on the surface of T7 phage was sequentially screened with sera samples from human subjects showing different manifestations of the disease. Antigens that selectively and specifically bind to immune sera were then enriched using a multi-step panning procedure. This strategy identified five antigens, four of which were previously reported (ALT-2, TPX-2, VAH and COX-2) and the other one was a novel cuticular collagen (Col-4). Sera from immune individuals specifically recognized all the five antigens. However, ALT-2 appeared to be the most predominantly recognized antigen by the immune sera. Therefore, it was decided to evaluate the vaccine potential of recombinant ALT-2 (rALT-2) in a mouse and jird model. The results presented show that immunization with rALT-2 conferred over 73% protection against a challenge infection in the jird model and over 64% protection in the mouse model. The present study suggests that phage display-based cDNA screening may be a powerful tool to identify candidate vaccine antigens of infectious agents. PMID- 15271933 TI - Adherence to, invasion by, and cytokine production in response to serotype VIII group B Streptococci. AB - The adherence to and invasion of the human epithelial cell line A549 by group B streptococcus (GBS) serotype VIII strains were compared with those of serotype III strains by a conventional method and the dynamic in vitro attachment and invasion system. Twenty GBS strains, including nine vaginal isolates and one invasive isolate each of serotypes III and VIII, were used in the conventional attachment and invasion assay. Adherence to and invasion of A549 cells by serotype VIII GBS strains were significantly greater (P < 0.0001) than those by serotype III strains for both the invasive strain and vaginal isolates. Cytokine production by A549 cells following stimulation with GBS serotypes III and VIII or their purified capsular polysaccharides (CPS) was measured. Serotype III strains stimulated significantly greater tumor necrosis factor alpha (TNF-alpha) (P < 0.0001) and interleukin-10 (IL-10) (P < 0.05) production than did serotype VIII strains. IL-8 production in response to serotype VIII was significantly higher (P < 0.001) than that in response to serotype III. TNF-alpha, IL-8, and IL-10 production was greater in A549 cells infected with GBS than in the untreated control cells. TNF-alpha production was significantly greater (P < 0.005) after stimulation with purified GBS serotype III CPS than after stimulation with serotype VIII CPS, a result similar to that after stimulation with whole GBS. IL 12 production by A549 cells was observed only in response to infection with GBS serotype III, resulting in the possibility of a greater TH1 response in serotype III GBS. These results suggest differences in immune responses to infection with GBS serotypes III and VIII. PMID- 15271934 TI - Reinfection with Anaplasma phagocytophilum in BALB/c mice and cross-protection between two sympatric isolates. AB - Infection with Anaplasma phagocytophilum in white-footed mice results in partial protection against reinfection with the same agent. However, humans and domestic animals may be sequentially exposed to different isolates of the agent circulating in the same or adjacent foci. We investigated whether immune response to a tick-borne infection with A. phagocytophilum provides protection against homologous and heterologous challenges. BALB/c mice were infected with one of the two sympatric isolates of A. phagocytophilum via tick bite and challenged 16 weeks later by Ixodes scapularis nymphs infected with either the same or the alternative isolate. As controls, groups of infected mice were challenged by uninfected ticks to confirm an absence of reactivation of the original infection or groups of naive mice were fed upon by ticks from cohorts used for an infectious challenge. Xenodiagnostic I. scapularis larvae were fed upon each mouse at 14 and 21 days postchallenge (PCH) and tested for the presence of A. phagocytophilum as freshly molted nymphs. Blood samples for quantitative PCR were collected at 7, 14, 21, and 70 days PCH. Serum samples were collected weekly to monitor development of immune response. The proportion of infected animals, levels of bacteremia, and the prevalence of infection in xenodiagnostic ticks were higher in groups of control mice exposed to A. phagocytophilum for the first time than in mice reinfected with either homologous or heterologous isolates. The presence of antibodies against A. phagocytophilum did not protect mice from a challenge with either homologous or heterologous isolates, however the ensuing reinfection was significantly milder and of a shorter duration than the first infection with either isolate. PMID- 15271935 TI - Disruption of the Aspergillus fumigatus gene encoding nucleolar protein CgrA impairs thermotolerant growth and reduces virulence. AB - Aspergillus fumigatus CgrA is the ortholog of a yeast nucleolar protein that functions in ribosome synthesis. To determine how CgrA contributes to the virulence of A. fumigatus, a Delta cgrA mutant was constructed by targeted gene disruption, and the mutant was reconstituted to wild type by homologous introduction of a functional cgrA gene. The Delta cgrA mutant had the same growth rate as the wild type at room temperature. However, when the cultures were incubated at 37 degrees C, a condition that increased the growth rate of the wild type and reconstituted strains approximately threefold, the Delta cgrA mutant was unable to increase its growth rate. The absence of cgrA function caused a delay in both the onset and rate of germination at 37 degrees C but had little effect on germination at room temperature. The Delta cgrA mutant was significantly less virulent than the wild-type or reconstituted strain in immunosuppressed mice and was associated with smaller fungal colonies in lung tissue. However, this difference was less pronounced in a Drosophila infection model at 25 degrees C, which correlated with the comparable growth rates of the two strains at this temperature. To determine the intracellular localization of CgrA, the protein was tagged at the C terminus with green fluorescent protein, and costaining with propidium iodide revealed a predominantly nucleolar localization of the fusion protein in living hyphae. Together, these findings establish the intracellular localization of CgrA in A. fumigatus and demonstrate that cgrA is required for thermotolerant growth and wild-type virulence of the organism. PMID- 15271936 TI - The V antigen of Pseudomonas aeruginosa is required for assembly of the functional PopB/PopD translocation pore in host cell membranes. AB - Pseudomonas aeruginosa efficiently intoxicates eukaryotic cells through the activity of the type III secretion-translocation system (TTSS). Gene deletions within the translocation operon pcrGVH-popBD abolish pore-forming activity of P. aeruginosa strains with macrophages and TTSS-dependent hemolysis. Here we investigated the requirements for PcrV, PopB, and PopD in pore formation by analyzing specific mutants using red blood cells (RBCs) and fibroblasts expressing green fluorescent protein fused to actin. Simultaneous secretion of three proteins, PopB, PopD, and PcrV, was required to achieve wild-type hemolysis and effector translocation. Deletion of pcrV in a cytotoxic strain did not affect secretion of PopB and PopD but abolished hemolytic activity and translocation of effectors into fibroblasts. Notably, the PcrV-deficient mutant was not capable of inserting PopD into host cell membranes, whereas PopB and PopD, but not PcrV, were readily found within membranes of wild-type-infected RBCs. Immunoprecipitation experiments performed by using a liposome model of pore assembly revealed a direct interaction between PopD and PopB but not between PopD and PcrV. Consequently, PcrV is necessary for the functional assembly of the PopB/D translocon complex but does not interact directly with pore-forming Pop proteins. PMID- 15271937 TI - Interaction of Chlamydia trachomatis serovar L2 with the host autophagic pathway. AB - Chlamydiae are obligate intracellular pathogens that replicate within a membrane bound compartment (the inclusion) and are associated with important human diseases, such as trachoma, pneumonia, and atherosclerosis. We have examined the interaction of the host autophagic pathway with Chlamydia trachomatis serovar L2 by using the specific autophagosomal stain monodansylcadaverine, antibodies to autophagosome-associated markers, and traditionally used autophagic inhibitors, particularly 3-methyladenine and amino acids. Chlamydial inclusions did not sequester monodansylcadaverine, suggesting absence of fusion with autophagosomes. Interestingly, exposure of cultures infected for 19 h to 3-methyladenine or single amino acids until the end of infection (44 h) caused various degrees of abnormalities in the inclusion maturation and in the progeny infectivity. Incubation of host cells with chemicals throughout the entire period of infection modulated the growth of Chlamydia even more dramatically. Remarkably, autophagosomal markers MAP-LC3 and calreticulin were redistributed to the inclusion of Chlamydia, a process that appears to be sensitive to 3-methyladenine and some amino acids. The present data indicate the lack of autophagosomal fusion with the inclusion because it was devoid of monodansylcadaverine and no distinct rim of autophagosomal protein-specific staining around the inclusion could be observed. However, high sensitivity of Chlamydia to conditions that could inhibit host autophagic pathway and the close association of MAP-LC3 and calreticulin with the inclusion membrane still suggest a potential role of host autophagy in the pathogenesis of Chlamydia. PMID- 15271938 TI - Experimental infections of neonatal mice with cysts of Giardia lamblia clone GS/M 83-H7 are associated with an antigenic reset of the parasite. AB - Transmission of the protozoan parasite Giardia lamblia from one to another host individuum occurs through peroral ingestion of cysts which, following excystation in the small intestine, release two trophozoites each. Many studies have focused on the major surface antigen, VSP (for variant surface protein), which is responsible for the antigenic variability of the parasite. By using trophozoites of G. lamblia clone GS/M-83-H7 (expressing VSP H7) and the neonatal mouse model for experimental infections, we quantitatively assessed the process of antigenic variation of the parasite on the transcriptional level. In the present study, variant-specific regions identified on different GS/M-83-H7 vsp sequences served as targets for quantitative reverse transcription-PCR to monitor alterations in vsp mRNA levels during infection. Respective results demonstrated that antigenic switching of both the duodenal trophozoite and the cecal cyst populations was associated with a massive reduction in vsp H7 mRNA levels but not with a simultaneous increase in transcripts of any of the subvariant vsp genes analyzed. Most importantly, we also explored giardial variant-type formation and vsp mRNA levels after infection of mice with cysts. This infection mode led to an antigenic reset of the parasite in that a VSP H7-negative inoculum "converted" into a population of intestinal trophozoites that essentially consisted of the original VSP H7 type. This antigenic reset appears to be associated with excystation rather than with a selective process which favors expansion of a residual population of VSP H7 types within the antigenically diversified cyst inoculum. Based on these findings, the VSP H7 type has to be regarded as a predominant variant of G. lamblia clone GS/M-83-H7 which (re-)emerges during early-stage infection and may contribute to an optimal establishment of the parasite within the intestine of the experimental murine host. PMID- 15271939 TI - Anaplasma phagocytophilum utilizes multiple host evasion mechanisms to thwart NADPH oxidase-mediated killing during neutrophil infection. AB - Anaplasma phagocytophilum, the etiologic agent of human anaplasmosis, is a bacterial pathogen that specifically colonizes neutrophils. Neutrophils utilize the NADPH oxidase complex to generate superoxide (O(2)(-)) and initiate oxidative killing of microorganisms. A. phagocytophilum's unique tropism for neutrophils, however, indicates that it subverts and/or avoids oxidative killing. We therefore examined the effects of A. phagocytophilum infection on neutrophil NADPH oxidase assembly and reactive oxygen species (ROS) production. Following neutrophil binding, Anaplasma invasion requires at least 240 min. During its prolonged association with the neutrophil plasma membrane, A. phagocytophilum stimulates NADPH oxidase assembly, as indicated by increased cytochrome b(558) mobilization to the membrane, as well as colocalization of Rac and p22(phox). This initial stimulation taxes the host neutrophil's finite oxidase reserves, as demonstrated by time- and bacterial-dose-dependent decreases in secondary activation by N formyl-methionyl-leucyl-phenylalanine (FMLP) or phorbol myristate acetate (PMA). This stimulation is modest, however, and does not diminish oxidase stores to nearly the extent that Escherichia coli, serum-opsonized zymosan, FMLP, or PMA do. Despite the apparent activation of NADPH oxidase, no change in ROS-dependent chemiluminescence is observed upon the addition of A. phagocytophilum to neutrophils, indicating that the bacterium may scavenge exogenous O(2)(-). Indeed, A. phagocytophilum rapidly detoxifies O(2)(-) in a cell-free system. Once internalized, the bacterium resides within a protective vacuole that excludes p22(phox) and gp91(phox). Thus, A. phagocytophilum employs at least two strategies to protect itself from neutrophil NADPH oxidase-mediated killing. PMID- 15271940 TI - YAPI, a new Yersinia pseudotuberculosis pathogenicity island. AB - Pathogenicity islands (PAIs) are chromosomal clusters of pathogen-specific virulence genes often found at tRNA loci. In the Yersinia pseudotuberculosis 32777 chromosome, we characterized a 98-kb segment that has all of the characteristic features of a PAI, including insertion in a (phenylalanine) tRNA gene, the presence of a bacteriophage-like integrase-encoding gene, and direct repeats at the integration sites. The G+C content of the segment ranges from 31 to 60%, reflecting a genetic mosaic: this is consistent with the notion that the sequences were horizontally acquired. The PAI, termed YAPI (for Yersinia adhesion pathogenicity island), carries 95 open reading frames and includes (i) the previously described pil operon, encoding a type IV pilus that contributes to pathogenicity (F. Collyn et al., Infect. Immun. 70:6196-6205, 2002); (ii) a block of genes potentially involved in general metabolism; (iii) a gene cluster for a restriction-modification system; and (iv) a large number of mobile genetic elements. Furthermore, the PAI can excise itself from the chromosome at low frequency and in a precise manner, and deletion does not result in a significant decrease of bacterial virulence compared to inactivation of the fimbrial gene cluster alone. The prevalence and size of the PAI vary from one Y. pseudotuberculosis strain to another, and it can be found integrated into either of the two phe tRNA loci present on the species' chromosome. YAPI was not detected in the genome of the genetically closely related species Y. pestis, whereas a homologous PAI is harbored by the Y. enterocolitica chromosome. PMID- 15271941 TI - Identification and characterization of a Neospora caninum microneme-associated protein (NcMIC4) that exhibits unique lactose-binding properties. AB - Microneme proteins have been shown to play an important role in the early phase of host cell adhesion, by mediating the contact between the parasite and host cell surface receptors. In this study we have identified and characterized a lectin-like protein of Neospora caninum tachyzoites which was purified by alpha lactose-agarose affinity chromatography. Upon separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, this lactose-binding protein migrated at 70 and 55 kDa under reducing and nonreducing conditions, respectively. Immunofluorescence and immunogold electron microscopy with affinity-purified antibodies showed that the protein was associated with the tachyzoite micronemes. Mass spectrometry analyses and expressed sequence tag database mining revealed that this protein is a member of the Neospora microneme protein family; the protein was named NcMIC4 (N. caninum microneme protein 4). Upon two-dimensional gel electrophoresis, NcMIC4 separated into seven distinct isoforms. Incubation of extracellular parasites at 37 degrees C resulted in the secretion of NcMIC4 into the medium as a soluble protein, and the secreted protein exhibited a slightly reduced M(r) but retained its lactose-binding properties. Immunofluorescence was used to investigate the temporal and spatial distribution of NcMIC4 in tachyzoites entering their host cells and showed that reexpression of NcMIC4 took place 30 min after entry into the host cell. Incubation of secreted fractions and purified NcMIC4 with Vero cells demonstrated binding of NcMIC4 to Vero cells as well as binding to chondroitin sulfate A glycosaminoglycans. PMID- 15271942 TI - Murine model of pulmonary anthrax: kinetics of dissemination, histopathology, and mouse strain susceptibility. AB - Bioweapons are most often designed for delivery to the lung, although this route is not the usual portal of entry for many of the pathogens in the natural environment. Vaccines and therapeutics that are efficacious for natural routes of infection may not be effective against the pulmonary route. Pulmonary models are needed to investigate the importance of specific bacterial genes in virulence, to identify components of the host immune system that are important in providing innate and acquired protection, and for testing diagnostic and therapeutic strategies. This report describes the characteristics of host and Bacillus anthracis interactions in a murine pulmonary-infection model. The infective dose varied depending on the route and method of inoculation. The germination process in the lung began within 1 h of inoculation into the lung, although growth within the lung was limited. B. anthracis was found in the lung-associated lymph nodes approximately 5 h after infection. Minimal pneumonitis was associated with the lung infection, but significant systemic pathology was noted after dissemination. Infected mice typically succumbed to infection approximately 3 to 4 days after inoculation. The 50% lethal doses differed among inbred strains of mice, but within a given mouse strain, neither the age nor the sex of the mice influenced susceptibility to B. anthracis. PMID- 15271943 TI - Protective and nonprotective human immunoglobulin M monoclonal antibodies to Cryptococcus neoformans glucuronoxylomannan manifest different specificities and gene use profiles. AB - The features of protective murine antibodies to the Cryptococcus neoformans capsular polysaccharide glucuronoxylomannan (GXM) have been rigorously investigated; however, the characteristics of protective human antibodies to GXM have not been defined. We produced monoclonal antibodies (MAbs) from XenoMouse mice (transgenic mice that express human immunoglobulin M [IgM], IgG2, and kappa) which were immunized with a C. neoformans serotype D strain 24067 GXM-diphtheria toxoid conjugate. This study reports the specificity and efficacy of three human IgM MAbs, G14, G15, and G19, generated from these mice. Each MAb was specific for GXM, but G14 and G19 had different specificity based on their binding to serotype A strain H99 and SB4 GXMs, to which G15 did not bind. Nucleic acid sequence analysis revealed that G15 uses V(H)3-64 in the germ line configuration. G14 and G19 use V(H)6-1, which has somatic mutations. All of the MAbs use V kappa DPK22/A27. Studies of MAb efficacy in BALB/c mice showed that administration of 0.1 mg, but not 1 or 0.01 mg, of G15 prolonged survival against lethal C. neoformans strain 24067 challenge, whereas G14 and G19 were not protective at any dose. This panel of MAbs illustrates that serotype D GXM has epitopes that elicit human antibodies that can be either protective or nonprotective. Our findings suggest that V(H) gene use may influence GXM specificity and efficacy, and they provide insights into the possible contribution that V(H) gene use may have in resistance and susceptibility to cryptococcosis. PMID- 15271944 TI - Molecular interactions of surface protein peptides of Streptococcus gordonii with human salivary components. AB - Oral streptococci play a large role in dental biofilm formation, and several types interact as early colonizers with the enamel salivary pellicle to form the primary biofilm, as well as to incorporate other bacteria on tooth surfaces. Interactions of surface molecules of individual streptococci with the salivary pellicle on the tooth surface have an influence on the etiological properties of an oral biofilm. To elucidate the molecular interactions of streptococci with salivary components, binding between surface protein (SspB and PAg) peptides of Streptococcus gordonii and Streptococcus sobrinus were investigated by utilizing BIAcore biosensor technology. The analogous peptide [change of T at position 400 to K in SspB(390-402), resulting in the SspB(390-T400K-402) peptide] from S. gordonii showed the greatest response for binding to salivary components and inhibited the binding of Streptococcus sanguis by more than 50% in a competitive inhibition assay in a comparison with other SspB and PAg peptides. This peptide also bound to the high-molecular-weight protein complex of salivary components and the agglutinin (gp340/DMBT1) peptide (scavenger receptor cysteine-rich domain peptide 2 [SRCRP 2]). In addition, the SspB(390-T400K-402) peptide was visualized by two surface positive charges in connection with the positively charged residues, in which lysine was a key residue for binding. Therefore, the region containing lysine may have binding activity in S. gordonii and S. sanguis, and the SRCRP 2 region may function as a receptor for the binding. These findings may provide useful information regarding the molecular mechanism of early biofilm formation by streptococci on tooth surfaces. PMID- 15271945 TI - Interaction of Dr adhesin with collagen type IV is a critical step in Escherichia coli renal persistence. AB - The pathogenic mechanism of recurrent or chronic urinary tract infection is poorly understood. Escherichia coli cells bearing Dr fimbriae display unique tropism to the basement membrane (BM)-renal interstitium that enables the bacteria to cause chronic pyelonephritis in experimental mice. The renal receptors for Dr-fimbriated E. coli are type IV collagen and decay-accelerating factor (DAF). We hypothesized that type IV collagen receptor-mediated BM interstitial tropism is essential for E. coli to cause chronic pyelonephritis. To test the role of the type IV collagen tropism of Dr-fimbriated E. coli in renal persistence, we constructed an isogenic mutant in the DraE adhesin subunit that was unable to bind type IV collagen but retained binding to DAF and examined its virulence in the mouse model. The collagen-binding mutant DrI113T was eliminated from the mouse renal tissues in 6 to 8 weeks, while the parent strain caused persistent renal infection that lasted at least 14 weeks (P < or = 0.02). Transcomplementation with the intact Dr operon restored collagen-binding activity, BM-interstitial tropism, and the ability to cause persistent renal infection. We conclude that type IV collagen binding mediated by DraE adhesin is a critical step for the development of persistent renal infection in a murine model of E. coli pyelonephritis. PMID- 15271946 TI - Proapoptotic effect of proteolytic activation of matrix metalloproteinases by Streptococcus pyogenes thiol proteinase (Streptococcus pyrogenic exotoxin B). AB - Streptococcus pyogenes thiol proteinase, also known as streptococcal pyrogenic exotoxin B (SpeB), has been suggested to be a major virulence factor in S. pyogenes infection. SpeB was reported to induce apoptosis of host cells, but its mechanism of action is not yet fully understood. In this study, we examined the involvement of matrix metalloproteinases (MMPs) in SpeB-induced apoptosis. We first developed a large-scale preparation of recombinant SpeB and precursors of human MMP-9 and -2 (proMMPs) by using Escherichia coli Rosetta (DE3)pLysS and baculovirus-insect cell expression systems, respectively. Treatment with SpeB induced effective proteolytic activation of both proMMP-9 and -2. When RAW264 murine macrophages were incubated with SpeB-activated proMMP-9, the level of tumor necrosis factor alpha (TNF-alpha) in conditioned medium (CM), assessed by an enzyme immunoassay, was elevated. This increase was completely inhibited by addition of the MMP inhibitor SI-27 to the cell culture. The CM also produced marked induction of apoptosis of U937 human monocytic cells. Similarly, soluble Fas ligand (sFasL) was detected in CM of cultures of SW480 cells expressing FasL after treatment with SpeB-activated proMMPs; this CM also induced apoptosis in U937 cells. SpeB had a direct effect as well and caused the release of TNF-alpha and sFasL from the cells. SpeB-dependent production of MMP-9 and -2 and proapoptotic molecules (TNF-alpha and sFasL) was evident in a murine model of severe invasive S. pyogenes infection. These results suggest that SpeB or SpeB activated MMPs contribute to tissue damage and streptococcal invasion in the host via extracellular release of TNF-alpha and sFasL. PMID- 15271947 TI - Aeromonas hydrophila beta-hemolysin induces active chloride secretion in colon epithelial cells (HT-29/B6). AB - The diarrheal mechanisms in Aeromonas enteritis are not completely understood. In this study we investigated the effect of aeromonads and of their secretory products on ion secretion and barrier function of monolayers of human intestinal cells (HT-29/B6). Ion secretion was determined as a short-circuit current (I(SC)) of HT-29/B6 monolayers mounted in Ussing-type chambers. Transepithelial resistance (R(t)) served as a measure of permeability. A diarrheal strain of Aeromonas hydrophila (strain Sb) added to the mucosal side of HT-29/B6 monolayers induced a significant I(SC) (39 +/- 3 microA/cm(2)) and decreased the R(t) to approximately 10% of the initial value. A qualitatively identical response was obtained with sterile supernatant of strain Sb, and Aeromonas supernatant also induced a significant I(SC) in totally stripped human colon. Tracer flux and ion replacement studies revealed the I(SC) to be mainly accounted for by electrogenic Cl(-) secretion. Supernatant applied serosally completely abolished basal I(SC). The supernatant-induced I(SC) was inhibited by the protein kinase C inhibitor chelerythrine, whereas a protein kinase A inhibitor (H8) and a Ca(2+) chelator (BAPTA-AM) had no effect. Physicochemical properties indicated that the supernatant's active compound was an aerolysin-related Aeromonas beta-hemolysin. Accordingly, identical I(SC) and R(t) responses were obtained with Escherichia coli lysates harboring the cloned beta-hemolysin gene from strain SB or the aerA gene encoding for aerolysin. Sequence comparison revealed a 64% homology between aerolysin and the beta-hemolysin cloned from Aeromonas sp. strain Sb. In conclusion, beta-hemolysin secreted by pathogenic aeromonads induces active Cl(-) secretion in the intestinal epithelium, possibly by channel insertion into the apical membrane and by activation of protein kinase C. PMID- 15271948 TI - Human diffusely adhering Escherichia coli expressing Afa/Dr adhesins that use human CD55 (decay-accelerating factor) as a receptor does not bind the rodent and pig analogues of CD55. AB - Afa/Dr diffusely adhering Escherichia coli (DAEC) bacteria that are responsible for recurrent urinary tract and gastrointestinal infections recognized as a receptor the glycosylphosphatidylinositol (GPI)-anchored protein decay accelerating factor (DAF; CD55) at the brush border of cultured human intestinal cells. Results show that Afa/Dr DAEC C1845 bacteria were poorly associated with the mucosa of the gastrointestinal tract of infected mice. We conducted experiments with Chinese hamster ovary (CHO) cells stably transfected with mouse (GPI or transmembrane forms), pig, or human CD55 or mouse Crry cDNAs or transfected with empty vector pDR2EF1 alpha. Recombinant E. coli AAEC185 bacteria expressing Dr or F1845 adhesins bound strongly to CHO cells expressing human CD55 but not to the CHO cells expressing mouse (transmembrane and GPI anchored), rat, or pig CD55 or mouse Crry. Positive clustering of CD55 around Dr-positive bacteria was observed in human CD55-expressing CHO cells but not around the rarely adhering Dr-positive bacteria randomly distributed at the cell surface of CHO cells expressing mouse, rat, or pig CD55. PMID- 15271949 TI - The luxS gene is not required for Borrelia burgdorferi tick colonization, transmission to a mammalian host, or induction of disease. AB - luxS mutants of Borrelia burgdorferi strain 297 naturally colonized their arthropod (Ixodes scapularis) vector, were maintained in ticks throughout the molting process (larvae to nymphs), were tick transmitted to uninfected mice, and elicited histopathology in mice indistinguishable from that induced by wild-type B. burgdorferi. PMID- 15271950 TI - Elevated nitric oxide production in children with malarial anemia: hemozoin induced nitric oxide synthase type 2 transcripts and nitric oxide in blood mononuclear cells. AB - Experiments outlined here investigate the role of nitric oxide (NO) in the pathogenesis of Plasmodium falciparum-induced malarial anemia (MA). The results show that ex vivo and in vitro NO synthase (NOS) activity in peripheral blood mononuclear cells (PBMCs) is significantly elevated in children with MA and inversely associated with hemoglobin levels. Additional experiments using PBMCs from non-malaria-exposed donors demonstrate that physiologic amounts of P. falciparum-derived hemozoin augment NOS type 2 (NOS2) transcripts and NO production. Results of these experiments illustrate that elevated NO production in children with MA is associated with decreased hemoglobin concentrations and that hemozoin can induce NOS2-derived NO formation in cultured blood mononuclear cells. PMID- 15271951 TI - Characterization of adenylate cyclase-hemolysin gene duplication in a Bordetella pertussis isolate. AB - We describe a clinical isolate of Bordetella pertussis, the agent responsible for whooping cough, composed of at least two clones harboring one or two copies of the cya locus encoding one of the major toxins, adenylate cyclase-hemolysin. No difference was observed between the two clones in murine and cellular models, probably due to the high instability of the cya locus duplication. PMID- 15271952 TI - Genetic susceptibility of mice to Candida albicans vaginitis correlates with host estrogen sensitivity. AB - We compared susceptibility to Candida vaginitis in derived murine substrains differing in sensitivity to estrogen (CD-1 and CD10, resistant; CD3 and C57BL/6 responsive), and in F1 crosses. The order of decreasing resistance was CD-1 > or = CD10 > or = CD10 x CD3F1 > CD10 x B6F1 > CD3 > C57BL/6 and correlated with estrogen responsiveness in endocrine disruptor assays. Resistance to Candida vaginitis appears additive in CD10 x B6F1 animals and dominant in CD10 x CD3F1 animals. PMID- 15271953 TI - Mycoplasma penetrans is capable of activating V gamma 9/V delta 2 T cells while other human pathogenic mycoplasmas fail to do so. AB - While most mycoplasma species appear to have evolutionarily lost the ability to synthesize isoprenoid precursors, Mycoplasma penetrans has retained the nonmevalonate pathway that proceeds via the immunogenic intermediate (E)-4 hydroxy-3-methyl-but-2-enyl pyrophosphate (HMB-PP). Consequently, this pathogen is capable of stimulating human V gamma 9/V delta 2 T cells. PMID- 15271954 TI - N19 polyepitope as a carrier for enhanced immunogenicity and protective efficacy of meningococcal conjugate vaccines. AB - N19, a string of human universal CD4 T-cell epitopes from various pathogen derived antigens, was shown to exert a stronger carrier effect than CRM197 for the induction of anti-group C Neisseria meningitidis capsular polysaccharide (MenC), after immunization of mice with various dosages of N19-MenC or CRM-MenC conjugate vaccines. After two immunizations, the N19-based construct induced anti MenC antibody and protective bactericidal antibody titers higher than those induced by three doses of the CRM-MenC conjugate and required lower amounts of conjugate. N19-based conjugates are superior to CRM-based conjugates to induce protective immune responses to MenC conjugates. PMID- 15271955 TI - Biofilm formation in vitro and virulence in vivo of mutants of Klebsiella pneumoniae. AB - One of the early stages of Klebsiella pneumoniae airway infections may involve biofilm formation. Bacterial biofilm formation is frequently investigated using in vitro techniques that facilitate identification and analysis of individual genes. We investigated the correlation between K. pneumoniae biofilm formation in vitro and ability to cause infection in vivo following construction of a bank of mini-Tn5 mutants. PMID- 15271956 TI - Inhibition of nitric oxide synthase exacerbates group B streptococcus sepsis and arthritis in mice. AB - The role of nitric oxide in group B Streptococcus (GBS) infection was evaluated by inhibiting its production with aminoguanidine (AG). AG-treated mice displayed higher mortality rates and more frequent and severe arthritis than controls. Worsening of arthritis correlated with a higher number of GBS cells in the joints and local interleukin-1 beta production. PMID- 15271957 TI - Inactivation of the ciaH Gene in Streptococcus mutans diminishes mutacin production and competence development, alters sucrose-dependent biofilm formation, and reduces stress tolerance. AB - Many clinical isolates of Streptococcus mutans produce peptide antibiotics called mutacins. Mutacin production may play an important role in the ecology of S. mutans in dental plaque. In this study, inactivation of a histidine kinase gene, ciaH, abolished mutacin production. Surprisingly, the same mutation also diminished competence development, stress tolerance, and sucrose-dependent biofilm formation. PMID- 15271958 TI - Chlamydia pneumoniae decreases smooth muscle cell proliferation through induction of prostaglandin E2 synthesis. AB - Chlamydia pneumoniae may modulate the proliferation of smooth muscle cells (SMC) in atherosclerotic plaques. Conditioned medium from C. pneumoniae-infected SMC decreased the proliferation of uninfected SMC. Treatment of infected cells with the cyclooxygenase-2 inhibitor NS-398 [N-[2-(cyclohexyloxy)-4-nitrophenyl] methanesulfonamide] suppressed the up-regulation of prostaglandin E(2) (PGE(2)) and abolished the antimitogenic effect of conditioned medium, suggesting that C. pneumoniae can decrease SMC proliferation via stimulation of PGE(2) synthesis. PMID- 15271959 TI - Role of flagellum and motility in pathogenesis of Vibrio vulnificus. AB - To assess the role of the flagellum which was detected by immunoscreening of surface proteins of Vibrio vulnificus, an flgE-deleted mutant was constructed and tested for its pathogenicity. The ability of this nonmotile mutant to adhere to INT-407 cells and its role in biofilm were decreased, as was its lethality to mice. PMID- 15271960 TI - Intact purine biosynthesis pathways are required for wild-type virulence of Brucella abortus 2308 in the BALB/c mouse model. AB - Brucella abortus 2308 derivatives with mini-Tn5 insertions in purE, purL, and purD display significant attenuation in the BALB/c mouse model, while isogenic mutants with mini-Tn5 insertions in pheA, trpB, and dagA display little or no attenuation in cultured murine macrophages or mice. These experimental findings confirm the importance of the purine biosynthesis pathways for the survival and replication of the brucellae in host macrophages. In contrast to previous reports, however, these results indicate that exogenous tryptophan and phenylalanine are available for use by the brucellae in the phagosomal compartment. PMID- 15271961 TI - Involvement of the chemokine RANTES (CCL5) in resistance to experimental infection with Leishmania major. AB - The expression and putative role of chemokines during infection with Leishmania major in mice were investigated. CCL5 expression correlates with resistance, and blockade of CCL5 rendered mice more susceptible to infection. CCL5 is part of the cascade of events leading to efficient parasite control in L. major infection. PMID- 15271962 TI - DNA-Salmonella enterica serovar Typhimurium primer-booster vaccination biases towards T helper 1 responses and enhances protection against Leishmania major infection in mice. AB - Successful resolution of infections by intracellular pathogens requires gamma interferon (IFN-gamma). DNA vaccines promote T helper 1 (Th1) responses by triggering interleukin-12 (IL-12) release by dendritic cells (DC) through Toll like receptor 9 (TLR9). In humans TLR9 is restricted to plasmacytoid DC. Here we show that DNA-Salmonella enterica serovar Typhimurium primer-booster vaccination, which provides alternative ligands to bind TLR4 on myeloid DC, strongly biases towards Th1 responses compared to vaccination with DNA alone. This results in higher immunoglobulin G2a (IgG2a) responses compared to IgG1 responses, higher IFN-gamma responses compared to IL-10 CD4(+)-T-cell responses, and enhanced protection against Leishmania major infection in susceptible BALB/c mice. PMID- 15271963 TI - Roles of the Maltese cross form of Babesia microti in the development of parasitemia in B. microti infection. PMID- 15271965 TI - Dentin regeneration by dental pulp stem cell therapy with recombinant human bone morphogenetic protein 2. AB - Regenerative medicine is based on stem cells, signals, and scaffolds. Dental pulp tissue has the potential to regenerate dentin in response to noxious stimuli, such as caries. The progenitor/stem cells are responsible for this regeneration. Thus, stem cell therapy has considerable promise in dentin regeneration. Culture of porcine pulp cells, as a three-dimensional pellet, promoted odontoblast differentiation compared with monolayers. The expression of dentin sialophosphoprotein (Dspp) and enamelysin/matrix metalloproteinase 20 (MMP20) mRNA confirmed the differentiation of pulp cells into odontoblasts and was stimulated by the morphogenetic signal, bone morphogenetic protein 2 (BMP2). Based on the in vitro experiments, an in vivo evaluation of pulp progenitor/stem cells in the dog was performed. The autogenous transplantation of the BMP2 treated pellet culture onto the amputated pulp stimulated reparative dentin formation. In conclusion, BMP2 can direct pulp progenitor/stem cell differentiation into odontoblasts and result in dentin formation. PMID- 15271966 TI - Mechanical strain delivers anti-apoptotic and proliferative signals to gingival fibroblasts. AB - Physical forces play a critical role in the survival and proliferation of many cell types, including fibroblasts. Gingival fibroblasts are exposed to mechanical stress during mastication, orthodontic tooth movement, and wound healing following periodontal surgery. The aim of this study was to examine the effect of mechanical strain on human gingival fibroblasts (hGF). Cells were subjected to short-term (up to 60 min) and long-term (up to 48 hrs) 20% average elongation at 0.1 Hz. We monitored survival signaling by evaluating the phosphorylation status and localization of Forkhead box (FoxO) family members, which are mediators of apoptosis. We also examined strain-induced proliferation by measuring the level of proliferating cell nuclear antigen (PCNA). We observed that cyclic strain caused the phosphorylation and retention in the cytoplasm of FoxO family members. Moreover, mechanical strain resulted in increased ERK kinase phosphorylation and PCNA expression. In conclusion, cyclic strain delivers anti-apoptotic and proliferative stimuli to hGF. PMID- 15271967 TI - Cementoblast gene expression is regulated by Porphyromonas gingivalis lipopolysaccharide partially via toll-like receptor-4/MD-2. AB - Lipopolysaccharides are potent inflammatory mediators considered to contribute to destruction of periodontal tissues. Here, we hypothesized that Porphyromonas gingivalis lipopolysaccharide (P-LPS) treatment would regulate gene expression in murine cementoblasts through Toll-like receptor 4. Real-time (RT)-PCR and Northern blot analysis indicated that P-LPS decreased expression of transcripts for osteocalcin (OCN) and receptor activator of nuclear factor kappaB ligand (RANKL). In contrast, a dose-dependent up-regulation in mRNA levels for osteopontin (OPN) and osteoprotegerin (OPG) was observed. Similarly, ELISA demonstrated decreased RANKL and increased OPG levels. A monoclonal antibody specific for mouse TLR-4/MD-2 partially neutralized the P-LPS effect on cementoblasts. These results indicate that exposure of cementoblasts to P-LPS can alter cell function by regulating markers of osteoclastic activity (e.g., RANKL/OPG), thereby potentially affecting the inflammation-associated resorption of mineralized tissues. PMID- 15271968 TI - Amelogenesis imperfecta in a new animal model--a mutation in chromosome 5 (human 4q21). AB - Candidate genes for amelogenesis imperfecta (AI) and dentinogenesis imperfecta (DI) are located on 4q21 in humans. We tested our hypothesis that mutations in the portion of mouse chromosome 5 corresponding to human chromosome 4q21 would cause enamel and dentin abnormalities. Male C3H mice were injected with ethylnitrosourea (ENU). Within a dominant ENU mutagenesis screen, a mouse mutant was isolated with an abnormal tooth enamel (ATE) phenotype. The structure and ultrastructure of teeth were studied. The mutation was located on mouse chromosome 5 in an interval of 9 cM between markers D5Mit18 and D5Mit10. Homozygotic mutants showed total enamel aplasia with exposed dentinal tubules, while heterozygotic mutants showed a significant reduction in enamel width. Dentin of mutant mice showed a reduced content of mature collagen cross-links. We were able to demonstrate that a mutation on chromosome 5 corresponding to human chromosome 4q21 can cause amelogenesis imperfecta and changes in dentin composition. PMID- 15271969 TI - Induction of calcification in MC3T3-E1 cells by inorganic polyphosphate. AB - Relatively large amounts of inorganic polyphosphate [poly(P)] (400 microM) have been found in normal osteoblasts. The effect of poly(P) with an average chain length of 65 phosphate residues on cell calcification was therefore investigated with the use of MC3T3-E1 cells. Expression of both osteopontin and osteocalcin was induced by poly(P) (0.1 approximately 1 mM), and cells treated with poly(P) were strongly stained by alizarin red. In addition, the level of alkaline phosphatase activity induced in poly(P)-treated cells was two-fold higher than that in either orthophosphate-treated or control cells but not higher than that in cells treated with beta-glycerophosphate and ascorbic acid. In contrast, however, polyphosphatase activities were activated by poly(P) treatment to levels up to six-fold greater than that in controls. MC3T3-E1 cells may utilize poly(P) as a phosphate source for calcification rather than phosphate sources that are mainly produced by ALPase. Poly(P)-dependent induction of polyphosphatase activities may therefore promote calcification in MC3T3-E1 cells. PMID- 15271970 TI - Oxidative stress-induced DNA damage in the synovial cells of the temporomandibular joint in the rat. AB - Synovial hyperplasia is a feature of degenerative temporomandibular joint (TMJ) disease. However, the mechanism by which hyperplasia progresses in the TMJ is unknown. Based on the hypothesis that the oxidative stress generated by mechanical loading causes degenerative changes in the TMJ synovium, we investigated the generation of the highly reactive species, peroxynitrite, and the occurrence of DNA damage in the synovium. After condylar hypermobility of rat TMJs, a marker of peroxynitrite, nitrotyrosine, was localized to the nuclei and cytoplasm of the synovial lining cells and fibroblasts in synovitis-induced TMJ. DNA single-strand breaks were found in the nuclei of the synovial cells only after enzyme treatment, whereas DNA double-strand breaks were not detected. These findings indicate that condylar hypermovement induces the proliferation of synovial cells, and suggest that oxidative stress leads to the progression of synovial hyperplasia via DNA damage of the synovial cells in TMJs after mechanical loading. PMID- 15271971 TI - Denervation resulting in dento-alveolar ankylosis associated with decreased Malassez epithelium. AB - Inferior alveolar nerve denervation causes appreciable decreases in the distribution of epithelial rests of Malassez. To explore roles of the Malassez epithelium, we attempted to evaluate possible changes in dento-alveolar tissues surrounding this epithelium by experimental denervation. We found that denervation led to dento-alveolar ankylosis with a decrease in the width of the periodontal spaces. Interestingly, with regeneration of the Malassez epithelium 10 weeks after the denervation, the periodontal space width showed a correspondingly significant increase. These findings suggest that the Malassez epithelium may be involved in the maintenance of periodontal space and that sensory innervation might be indirectly associated with it. In addition, it is of interest that denervation activated root resorption of the coronal root surface and that the consequently resorbed lacunae were repaired by cellular cementum. It is suggested that Malassez epithelium may negatively regulate root resorption and induce acellular cementum formation. PMID- 15271972 TI - An eight-year follow-up to a randomized clinical trial of participant satisfaction with three types of mandibular implant-retained overdentures. AB - Studies have shown that mandibular implant overdentures significantly increase satisfaction and quality of life of edentulous elders. Improved chewing ability appears to have a positive impact on nutritional state. Therefore, it is important to determine the best design of this prosthesis over the long term. In this randomized controlled trial, three groups of edentulous participants with atrophic mandibles wore 3 types of implant overdentures. During an eight-year follow-up, only seven of the 110 participants had dropped out of this study. Almost all participants were still satisfied with their overdentures. Participant satisfaction concerning retention and stability of the mandibular overdenture had decreased significantly in the two-implant ball attachment group, whereas the opinion of participants in the single- and triple-bar groups was still at the same level. The long-term results suggest that a mandibular overdenture retained by 2 implants with a single bar may be the best treatment strategy for edentulous people with atrophic ridges. PMID- 15271973 TI - Estimating rates of new root caries in older adults. AB - Although older adults are keeping their teeth longer, no national data are available on new caries in this age group. To characterize the extent of caries among older adults, we systematically reviewed studies on root caries incidence, increment, attack rate, and annual total (root + coronal) caries increment. We used a random-effects model to estimate annual summary measures and their 95% confidence intervals (CI) and tested for heterogeneity. For the 9 studies reporting root caries incidence, the summary measure equaled 23.7% (CI = 17.1 30.2%). For the 9 studies reporting root caries increment, the summary measure was 0.47 surfaces (CI = 0.34-0.61). For the 7 studies reporting total caries increment, the summary measure equaled 1.31 surfaces (95% CI = 1.01-1.61 surfaces). Because of heterogeneity, summary measures should be interpreted with caution. This research suggests, however, that older adults experience high rates of new caries and could benefit from caries-prevention programs. PMID- 15271974 TI - Prevalence of human herpesvirus-8 salivary shedding in HIV increases with CD4 count. AB - Human herpesvirus-8 (HHV-8) is the etiologic agent of Kaposi's sarcoma (KS), which occurs in epidemic form in human immunodeficiency virus(HIV)-infected individuals. Saliva is the only mucosal fluid in which infectious HHV-8 has been identified, although factors associated with HHV-8 salivary shedding remain unclear. Our study performed PCR analysis for HHV-8 DNA in saliva (and other body fluids) in 66 HIV- and HHV-8-co-infected women without KS so that we could examine predictors for HHV-8 DNA detection. CD4 count was the most significant predictor of HHV-8 salivary shedding, with increased prevalence of HHV-8 salivary DNA at higher CD4 counts. The odds of salivary HHV8 shedding at CD4 counts > = 350 cells/microL was 63 times the odds of shedding at CD4 < 350 (95%CI, 1.3 3078), with an increase in effect size when the analysis was restricted to those with a CD4 nadir > 200. Analysis of these data suggests an increased potential for HHV-8 transmission early in HIV infection, with implications for HHV-8 prevention. PMID- 15271975 TI - Water-dependent interfacial transition zone in resin-modified glass-ionomer cement/dentin interfaces. AB - The function of the interfacial transition zone (absorption layer) in resin modified glass-ionomer cements bonded to deep dentin remains obscure. This study tested the hypotheses that the absorption layer is formed only in the presence of water derived from hydrated dentin and allows for better bonding of resin modified glass-ionomer cements to dentin. Ten percent polyacrylic acid conditioned, hydrated, and dehydrated deep dentin specimens were bonded with 2 resin-modified glass-ionomer cements and sealed with resins to prevent environmental water gain or loss. A non-particulate absorption layer was identified over hydrated dentin only, and was clearly discernible from the hybrid layer when bonded interfaces were examined with transmission electron microscopy. This layer was relatively more resistant to dehydration stresses, and remained intact over the dentin surface after tensile testing. The absorption layer mediates better bonding of resin-modified glass-ionomer cements to deep dentin, and functions as a stress-relieving layer to reduce stresses induced by desiccation and shrinkage. PMID- 15271976 TI - Laboratory stresses and tractional forces on the TMJ disc surface. AB - The etiology of degenerative disease of the TMJ may involve fatigue produced by surface tractional forces and compressive stresses. This study tested the time dependent effects of compressive loading and stress-field translation on TMJ disc surface tractional forces and stresses. In laboratory experiments with 50 porcine discs, an acrylic indenter imposed 10 N static loads for 10 and 60 sec, followed by translation of the loaded indenter along the mediolateral axis of the disc. Maximum tractional forces were found to occur following 60 sec of static loading (p < 0.001), and increased with translation velocity (R(2) = 0.73); whereas maximum compressive stresses occurred after 10 sec of static loading (p < 0.001). Overall, the results were consistent with current mechanical theories of the time dependent effects of compressive loading of cartilage. PMID- 15271977 TI - A distal region in the interferon-gamma gene is a site of epigenetic remodeling and transcriptional regulation by interleukin-2. AB - Interferon-gamma (IFN-gamma) is a multifunctional cytokine that defines the development of Th1 cells and is critical for host defense against intracellular pathogens. IL-2 is another key immunoregulatory cytokine that is involved in T helper differentiation and is known to induce IFN-gamma expression in natural killer (NK) and T cells. Despite concerted efforts to identify the one or more transcriptional control mechanisms by which IL-2 induces IFN-gamma mRNA expression, no such genomic regulatory regions have been described. We have identified a DNase I hypersensitivity site approximately 3.5-4.0 kb upstream of the transcriptional start site. Using chromatin immunoprecipitation assays we found constitutive histone H3 acetylation in this distal region in primary human NK cells, which is enhanced by IL-2 treatment. This distal region is also preferentially acetylated on histones H3 and H4 in primary Th1 cells as compared with Th2 cells. Within this distal region we found a Stat5-like motif, and in vitro DNA binding assays as well as in vivo chromosomal immunoprecipitation assays showed IL-2-induced binding of both Stat5a and Stat5b to this distal element in the IFNG gene. We examined the function of this Stat5-binding motif by transfecting human peripheral blood mononuclear cells with -3.6 kb of IFNG luciferase constructs and found that phorbol 12-myristate 13-acetate/ionomycin induced transcription was augmented by IL-2 treatment. The effect of IL-2 was lost when the Stat5 motif was disrupted. These data led us to conclude that this distal region serves as both a target of chromatin remodeling in the IFNG locus as well as an IL-2-induced transcriptional enhancer that binds Stat5 proteins. PMID- 15271978 TI - Multiple pools of phosphatidylinositol 4-phosphate detected using the pleckstrin homology domain of Osh2p. AB - Phosphatidylinositol (PtdIns) phosphate (PtdInsP) lipids are used as intracellular signposts for the recruitment and activation of peripheral membrane proteins. Whereas the distribution of most PtdInsPs is restricted to a single organelle, PtdIns(4)P is unique in that it exists in several discrete pools, and so proteins that bind PtdIns(4)P must use extra receptors to achieve a restricted localization. Here we compare the two highly related pleckstrin homology (PH) domains from Osh1p and Osh2p, yeast homologues of oxysterol-binding protein (OSBP), that target membranes using PtdIns(4)P, and in vitro bind both PtdIns(4)P and PtdIns(4,5)P2. We show that Golgi targeting is specified by an additional site on PH(Osh1), which lies on a face of the domain not previously known to interact with receptors. In contrast, PH(Osh2) does not have a demonstrable second site, and targets multiple pools of PtdInsPs, each dependent on a different PtdIns 4-kinase. This lack of a second site in PH(Osh2) allows it to be used as an unbiased reporter for altered distribution of 4-phosphorylated PtdIns. For example, in cells with excess PtdIns(4)P caused by inactivation of the phosphatase Sac1p, PH(Osh2) indicates that PtdIns(4)P accumulates on the plasma membrane, whereas other Golgi-targeted PH domains fail to detect this change. PMID- 15271979 TI - Alteration of nucleic acid structure and stability modulates the efficiency of minus-strand transfer mediated by the HIV-1 nucleocapsid protein. AB - During human immunodeficiency virus type 1 minus-strand transfer, the nucleocapsid protein (NC) facilitates annealing of the complementary repeat regions at the 3'-ends of acceptor RNA and minus-strand strong-stop DNA ((-) SSDNA). In addition, NC destabilizes the highly structured complementary trans activation response element (TAR) stem-loop (TAR DNA) at the 3'-end of (-) SSDNA and inhibits TAR-induced self-priming, a dead-end reaction that competes with minus-strand transfer. To investigate the relationship between nucleic acid secondary structure and NC function, a series of truncated (-) SSDNA and acceptor RNA constructs were used to assay minus-strand transfer and self-priming in vitro. The results were correlated with extensive enzymatic probing and mFold analysis. As the length of (-) SSDNA was decreased, self-priming increased and was highest when the DNA contained little more than TAR DNA, even if NC and acceptor were both present; in contrast, truncations within TAR DNA led to a striking reduction or elimination of self-priming. However, destabilization of TAR DNA was not sufficient for successful strand transfer: the stability of acceptor RNA was also crucial, and little or no strand transfer occurred if the RNA was highly stable. Significantly, NC may not be required for in vitro strand transfer if (-) SSDNA and acceptor RNA are small, relatively unstructured molecules with low thermodynamic stabilities. Collectively, these findings demonstrate that for efficient NC-mediated minus-strand transfer, a delicate thermodynamic balance between the RNA and DNA reactants must be maintained. PMID- 15271980 TI - Autocrine and exogenous transforming growth factor beta control cell cycle inhibition through pathways with different sensitivity. AB - Human colon carcinoma cells HCT116 that lack transforming growth factor beta (TGF beta) type II receptor (RII) demonstrated restoration of autocrine TGF-beta activity upon reexpression of RII without restoring inhibitory responses to exogenous TGF-beta treatment. RII transfectants (designated RII Cl 37) had a longer lag phase relative to NEO-transfected control cells (designated NEO pool) before entering exponential growth in tissue culture. The prolonged growth arrest of RII Cl 37 cells was associated with markedly reduced cyclin-dependent kinase (CDK)2 activity. Our results demonstrate that p21 induction by autocrine TGF-beta is responsible for reduced CDK2 activity, which at least partially contributes to prolonged growth arrest and reduced cell proliferation in RII Cl 37 cells. In contrast to RII transfectants, HCT116 cells transfected with chromosome 3 (designated HCT116Ch3), which bears the RII gene, restored the response to exogenous TGF-beta as well as autocrine TGF-beta activity. Autocrine TGF-beta activity in HCT116Ch3 cells induced p21 expression as seen in RII Cl 37 cells; however, in addition to autocrine activity, HCT116Ch3 cells responded to exogenous TGF-beta as decreased CDK4 expression and reduced pRb phosphorylation mediated a TGF-beta inhibitory response in these cells. These results indicate that autocrine TGF-beta regulates the cell cycle through a pathway different from exogenous TGF-beta in the sense that p21 is a more sensitive effector of the TGF beta signaling pathway, which can be induced and saturated by autocrine TGF-beta, whereas CDK4 inhibition is a less sensitive effector, which can only be activated by high levels of exogenous TGF-beta PMID- 15271981 TI - Myofibroblastic differentiation leads to hyaluronan accumulation through reduced hyaluronan turnover. AB - During the initiation and progression of fibrosis there is extensive differentiation of cells to a myofibroblastic phenotype. Because the synthesis of hyaluronan (HA) was recently linked to oncogenic epithelial-mesenchymal transformation, the present study investigated whether increased HA synthesis was also associated with myofibroblastic differentiation. HA synthesis and size were measured by incorporation of [(3)H]glucosamine, ion exchange, and size exclusion chromatography. Hyaluronan synthase (HAS) or hyaluronidase (HYAL) mRNA levels were assessed by reverse transcription-PCR. HYAL was detected by immunoblotting and the degradation of [(3)H]HA. Between 2- and 3-fold more HA appeared in the conditioned medium and became associated with the cells upon myofibroblastic differentiation. Inhibition of HAS and examination of HAS mRNA expression demonstrated that this was not the result of increased synthesis of HA or the induction of HAS 2. After differentiation, however, myofibroblasts metabolized exogenously supplied [(3)H]HA at a slower rate than fibroblasts and expressed lower levels of both HYAL 1 and HYAL 2 mRNA. Immunoblotting revealed more HYAL 1 and 2 in the myofibroblast conditioned medium. After acidification, however, there was no difference in HA degradation. This suggests that much of the released HYAL is inactive and that the observed differences in HA degradation are caused by cell-associated rather than secreted activity. This was confirmed by immunohistochemical staining for HYAL 1 and HYAL 2. This finding indicates the potential importance of the HYAL enzymes in controlling fibrotic progression and contrasts HA synthesis as a mediator of oncogenic transformation with that of HA degradation controlling fibrogenic differentiation. PMID- 15271982 TI - Nucling recruits Apaf-1/pro-caspase-9 complex for the induction of stress-induced apoptosis. AB - Nucling is a novel protein isolated from murine embryonal carcinoma cells with an up-regulated expression during cardiac muscle differentiation. We show here that Nucling was up-regulated by proapoptotic stimuli and important for the induction of apoptosis after cytotoxic stress. We further demonstrated that overexpressed Nucling was able to induce apoptosis. In Nucling-deficient cells, the expression levels of Apaf-1 and cytochrome c, which are the major components of an apoptosis promoting complex named apoptosome, were both down-regulated under cellular stress. A deficiency of Nucling also conferred resistance to apoptotic stress on the cell. After UV irradiation, Nucling was shown to reside in an Apaf-1/pro caspase-9 complex, suggesting that Nucling might be a key molecule for the formation and maintenance of this complex. Nucling induced translocation of Apaf 1 to the nucleus, thereby distributing the Nucling/Apaf-1/pro-caspase-9 complex to the nuclear fraction. These findings suggest that Nucling recruits and transports the apoptosome complex during stress-induced apoptosis. PMID- 15271983 TI - Depletion of intracellular ascorbate by the carcinogenic metals nickel and cobalt results in the induction of hypoxic stress. AB - Exposure of cells to carcinogenic compounds of nickel(II) and cobalt(II) causes activation of the HIF-1 transcription factor and up-regulates a battery of hypoxia-inducible genes. However, the mechanism of HIF-1 activation by these metals is not known. It was shown recently that hydroxylation of prolines in the HIFalpha subunit of HIF-1 is required for its binding with the von Hippel-Lindau tumor suppressor protein and the subsequent proteasomal destruction. Here we show that responsible prolyl hydroxylases are targets for both nickel(II) and cobalt(II) because degradation of a reporter protein containing the oxygen dependent degradation domain (Pro-402/564) of HIFalpha was abolished in a von Hippel-Lindau-dependent manner in cells exposed to nickel(II) or cobalt(II). The enzymatic activity of prolyl hydroxylases depends on iron as the activating metal, 2-oxoglutarate as a co-substrate, and ascorbic acid as a cofactor. Hydroxylase activity can be impaired by the depletion of any of these factors. We found that exposure of cells to nickel(II) or cobalt(II) did not affect the level of intracellular iron. Instead, nickel(II) or cobalt(II) exposure greatly depleted intracellular ascorbate. Co-exposure of cells to metals and ascorbate resulted in the increase of intracellular ascorbate and reversed both metal induced stabilization of HIF-1alpha and HIF-1-dependent gene transcription. Because ascorbate is essential for maintaining iron in prolyl hydroxylases in the active iron(II) state, we suggest that the observed depletion of ascorbate by nickel(II) or cobalt(II) favors iron oxidation and thus inactivation of the enzyme. PMID- 15271984 TI - Phosphorylation of the platelet-derived growth factor receptor-beta by G protein coupled receptor kinase-2 reduces receptor signaling and interaction with the Na(+)/H(+) exchanger regulatory factor. AB - G protein-coupled receptor kinase-2 (GRK2) can phosphorylate and desensitize the platelet-derived growth factor receptor-beta (PDGFRbeta) in heterologous cellular systems. To determine whether GRK2 regulates the PDGFRbeta in physiologic systems, we examined PDGFRbeta signaling in mouse embryonic fibroblasts from GRK2 null and cognate wild type mice. To discern a mechanism by which GRK2-mediated phosphorylation can desensitize the PDGFRbeta, but not the epidermal growth factor receptor (EGFR), we investigated effects of GRK2-mediated phosphorylation on the association of the PDGFRbeta with the Na(+)/H(+) exchanger regulatory factor (NHERF), a protein shown to potentiate dimerization of the PDGFRbeta, but not the EGFR. Physiologic expression of GRK2 diminished (a) phosphoinositide hydrolysis elicited through the PDGFRbeta but not heterotrimeric G proteins; (b) Akt activation evoked by the PDGFRbeta but not the EGFR; and (c) PDGF-induced tyrosyl phosphorylation of the PDGFRbeta itself. PDGFRbeta desensitization by physiologically expressed GRK2 correlated with a 2.5-fold increase in PDGF promoted PDGFRbeta seryl phosphorylation. In 293 cells, GRK2 overexpression reduced PDGFRbeta/NHERF association by 60%. This effect was reproduced by S1104D mutation of the PDGFRbeta, which also diminished PDGFRbeta activation and signaling (like the S1104A mutation) to an extent equivalent to that achieved by GRK2-mediated PDGFRbeta phosphorylation. GRK2 overexpression desensitized only the wild type but not the S1104A PDGFRbeta. We conclude that GRK2-mediated PDGFRbeta seryl phosphorylation plays an important role in desensitizing the PDGFRbeta in physiologic systems. Furthermore, this desensitization appears to involve GRK2-mediated phosphorylation of PDGFRbeta Ser(1104), with consequent dissociation of the PDGFRbeta from NHERF. PMID- 15271985 TI - Particulate adenylate cyclase plays a key role in human sperm olfactory receptor mediated chemotaxis. AB - Human sperm chemotaxis is a critical component of the fertilization process, but the molecular basis for this behavior remains unclear. Recent evidence shows that chemotactic responses depend on activation of the sperm olfactory receptor, hOR17 4. Certain floral scents, including bourgeonal, activate hOR17-4, trigger pronounced Ca(2+) fluxes, and evoke chemotaxis. Here, we provide evidence that hOR17-4 activation is coupled to a cAMP-mediated signaling cascade. Multidimensional protein identification technology was used to identify potential components of a G-protein-coupled cAMP transduction pathway in human sperm. These products included various membrane-associated adenylate cyclase (mAC) isoforms and the G(olf)-subunit. Using immunocytochemistry, specific mAC isoforms were localized to particular cell regions. Whereas mAC III occurred in the sperm head and midpiece, mAC VIII was distributed predominantly in the flagellum. In contrast, G(olf) was found mostly in the flagellum and midpiece. The observed spatial distribution patterns largely correspond to the spatiotemporal character of hOR17-4-induced Ca(2+) changes. Behavioral and Ca(2+) signaling responses of human sperm to bourgeonal were bioassayed in the presence, or absence, of the adenylate cyclase antagonist SQ22536. This specific agent inhibits particulate AC, but not soluble AC, activation. Upon incubation with SQ22536, cells ceased to exhibit Ca(2+) signaling, chemotaxis, and hyperactivation (faster swim speed and flagellar beat rate) in response to bourgeonal. Particulate AC is therefore required for induction of hOR17-4-mediated human sperm behavior and represents a promising target for future design of contraceptive drugs. PMID- 15271986 TI - Mutational analysis of ThiH, a member of the radical S-adenosylmethionine (AdoMet) protein superfamily. AB - Thiamine pyrophosphate (TPP) is an essential cofactor for all forms of life. In Salmonella enterica, the thiH gene product is required for the synthesis of the 4 methyl-5-beta hydroxyethyl-thiazole monophosphate moiety of TPP. ThiH is a member of the radical S-adenosylmethionine (AdoMet) superfamily of proteins that is characterized by the presence of oxygen labile [Fe-S] clusters. Lack of an in vitro activity assay for ThiH has hampered the analysis of this interesting enzyme. We circumvented this problem by using an in vivo activity assay for ThiH. Random and directed mutagenesis of the thiH gene was performed. Analysis of auxotrophic thiH mutants defined two classes, those that required thiazole to make TPP (null mutants) and those with thiamine auxotrophy that was corrected by either L-tyrosine or thiazole (ThiH* mutants). Increased levels of AdoMet also corrected the thiamine requirement of members of the latter class. Residues required for in vivo function were identified and are discussed in the context of structures available for AdoMet enzymes. PMID- 15271987 TI - Aberrant regulation of survivin by the RB/E2F family of proteins. AB - Survivin is a putative oncogene that is aberrantly expressed in cancer cells. It has been hypothesized to play a central role in cancer progression and resistance to therapy in diverse tumor types. Although some of the transcriptional processes regulating its expression have been established, the diversity of genes that may be controlling the levels of its expression in both normal cells as well as in cancer cells has not been fully explored. The most common genetically mutated pathways in human malignancies are the p53 tumor suppressor pathway and the RB/E2F pathway. Both of these pathways, when intact, provide essential checkpoints in the maintenance of normal cell growth and protect the cell from DNA damage. Using non-transformed embryonic fibroblasts, we provide evidence of a molecular link between the regulation of survivin transcription and the RB/E2F family of proteins. We demonstrate that both pRB and p130 can interact with the survivin promoter and can repress survivin transcription. We also show that the E2F activators (E2F1, E2F2, and E2F3) can bind to the survivin promoter and induce survivin transcription. Genetically modified cells that harbor deletions in various members of the RB/E2F family confirm our data from the wild-type cells. Our findings implicate several members of the RB/E2F pathway in an intricate mechanism of survivin gene regulation that, when genetically altered during the process of tumorigenesis, may function within cancer cells to aberrantly alter survivin levels and enhance tumor progression. PMID- 15271988 TI - The twisted abdomen phenotype of Drosophila POMT1 and POMT2 mutants coincides with their heterophilic protein O-mannosyltransferase activity. AB - Walker-Warburg syndrome, caused by mutations in protein O-mannosyltransferase-1 (POMT1), is an autosomal recessive disorder characterized by severe brain malformation, muscular dystrophy, and structural eye abnormalities. As humans have a second POMT, POMT2, we cloned each Drosophila ortholog of the human POMT genes and carried out RNA interference (RNAi) knock-down to investigate the function of these proteins in vivo. Drosophila POMT2 (dPOMT2) RNAi mutant flies showed a "twisted abdomen phenotype," in which the abdomen is twisted 30-60 degrees , similar to the dPOMT1 mutant. Moreover, dPOMT2 interacted genetically with dPOMT1, suggesting that the dPOMTs function in collaboration with each other in vivo. We expressed dPOMTs in Sf21 cells and measured POMT activity. dPOMT2 transferred a mannose to the dystroglycan protein only when it was coexpressed with dPOMT1. Likewise, dPOMT1 showed POMT activity only when coexpressed with dPOMT2, and neither dPOMT showed any activity by itself. Each dPOMT RNAi fly totally reduced POMT activity, despite the specific reduction in the level of each dPOMT mRNA. The expression pattern of dPOMT2 mRNA was found to be similar to that of dPOMT1 mRNA using whole mount in situ hybridization. These results demonstrate that the two dPOMTs function as a protein O-mannosyltransferase in association with each other, in vitro and in vivo, to generate and maintain normal muscle development. PMID- 15271989 TI - Loss of cell wall mannosylphosphate in Candida albicans does not influence macrophage recognition. AB - The outer layer of the cell wall of the human pathogenic fungus Candida albicans is enriched with heavily mannosylated glycoproteins that are the immediate point of contact between the fungus and cells of the host, including phagocytes. Previous work had identified components of the acid-labile fraction of N-linked mannan, comprising beta-1,2-linked mannose residues attached via a phosphodiester bond, as potential ligands for macrophage receptors and modulators of macrophage function. We therefore isolated and disrupted the CaMNN4 gene, which is required for mannosyl phosphate transfer and hence the attachment of beta-1,2 mannose oligosaccharides to the acid-labile N-mannan side chains. With the mannosylphosphate eliminated, the mnn4Delta null mutant was unable to bind the charged cationic dye Alcian Blue and was devoid of acid-labile beta-1,2-linked oligomannosaccharides. The mnn4Delta mutant was unaffected in cell growth and morphogenesis in vitro and in virulence in a murine model of systemic C. albicans infection. The null mutant was also not affected in its interaction with macrophages. Mannosylphosphate is therefore not required for macrophage interactions or for virulence of C. albicans. PMID- 15271990 TI - Rad53 kinase activation-independent replication checkpoint function of the N terminal forkhead-associated (FHA1) domain. AB - Saccharomyces cerevisiae Rad53 has crucial functions in many aspects of the cellular response to DNA damage and replication blocks. To coordinate these diverse roles, Rad53 has two forkhead-associated (FHA) phosphothreonine-binding domains in addition to a kinase domain. Here, we show that the conserved N terminal FHA1 domain is essential for the function of Rad53 to prevent the firing of late replication origins in response to replication blocks. However, the FHA1 domain is not required for Rad53 activation during S phase, and as a consequence of defective downstream signaling, Rad53 containing an inactive FHA1 domain is hyperphosphorylated in response to replication blocks. The FHA1 mutation dramatically hypersensitizes strains with defects in the cell cycle-wide checkpoint pathways (rad9Delta and rad17Delta) to DNA damage, but it is largely epistatic with defects in the replication checkpoint (mrc1Delta). Altogether, our data indicate that the FHA1 domain links activated Rad53 to downstream effectors in the replication checkpoint. The results reveal an important mechanistic difference to the homologous Schizosaccharomyces pombe FHA domain that is required for Mrc1-dependent activation of the corresponding Cds1 kinase. Surprisingly, despite the severely impaired replication checkpoint and also G(2)/M checkpoint functions, the FHA1 mutation by itself leads to only moderate viability defects in response to DNA damage, highlighting the importance of functionally redundant pathways. PMID- 15271991 TI - Obligatory role of Src kinase in the signaling mechanism for TRPC3 cation channels. AB - Members of the canonical transient receptor potential (TRPC) subfamily of cation channels are candidates for capacitative and non-capacitative Ca2+ entry channels. When ectopically expressed in cell lines, TRPC3 can be activated by phospholipase C-mediated generation of diacylglycerol or by addition of synthetic diacylglycerols, independently of Ca2+ store depletion. Apart from this mode of regulation, little is known about other receptor-dependent signaling events that modulate TRPC3 activity. In the present study the role of tyrosine kinases in receptor- and diacylglycerol-dependent activation of TRPC3 was investigated. In HEK293 cells stably expressing TRPC3, pharmacological inhibition of tyrosine kinases, and specifically of Src kinases, abolished activation of TRPC3 by muscarinic receptor stimulation and by diacylglycerol. Channel regulation was lost following expression of a dominant-negative mutant of Src, or when TRPC3 was expressed in an Src-deficient cell line. In both instances, wild-type Src restored TRPC3 regulation. We conclude that Src plays an obligatory role in the mechanism for receptor and diacylglycerol activation of TRPC3. PMID- 15271992 TI - Perturbing the linker regions of the alpha-subunit of transducin: a new class of constitutively active GTP-binding proteins. AB - The GDP-GTP exchange activity of the retinal G protein, transducin, is markedly accelerated by the photoreceptor rhodopsin in the first step of visual transduction. The x-ray structures for the alpha subunits of transducin (alpha(T)) and other G proteins suggest that the nucleotide-binding (Ras-like) domain and a large helical domain form a "clam shell" that buries the GDP molecule. Thus, receptor-promoted G protein activation may involve "opening the clam shell" to facilitate GDP dissociation. In this study, we have examined whether perturbing the linker regions connecting the Ras-like and helical domains of Galpha subunits gives rise to a more readily exchangeable state. The sole glycine residues in linkers 1 and 2 were individually changed to proline residues within an alpha(T)/alpha(i1) chimera (designated alpha(T)(*)). Both alpha(T)(*) linker mutants showed significant increases in their basal rates of GDP-GTP exchange when compared either to retinal alpha(T) or recombinant alpha(T)(*). The alpha(T)(*) linker mutants were responsive to aluminum fluoride, which binds to alpha-GDP complexes and induces changes in Switch 2. Although both linker mutants were further activated by light-activated rhodopsin together with the betagamma complex, their activation was not influenced by betagamma alone, arguing against the idea that the betagamma complex helps to pry apart the helical and Ras-like domains of Galpha subunits. Once activated, the alpha(T)(*) linker mutants were able to stimulate the cyclic GMP phosphodiesterase. Overall, these findings highlight a new class of activated Galpha mutants that constitutively exchange GDP for GTP and should prove valuable in studying different G protein-signaling systems. PMID- 15271993 TI - The chicken serotonin transporter discriminates between serotonin-selective reuptake inhibitors. A species-scanning mutagenesis study. AB - The serotonin transporter (SERT) belongs to a family of sodium chloride-dependent transporters responsible for uptake of amino acids and biogenic amines from extracellular spaces. SERT represents the main pharmacological target in the treatment of several clinical conditions, including depression and anxiety. Serotonin-selective reuptake inhibitors and tricyclic antidepressants are the most predominantly prescribed drugs in the treatment of depression. In addition to antidepressants also psychostimulants, like cocaine and amphetamines, are important SERT antagonists. In the present study, we report the cloning and characterization of chicken SERT. Although the uptake kinetic was very similar to human SERT, the pharmacological profiles differed considerably for the two species. We find that chicken SERT is capable of discriminating between different serotonin-selective reuptake inhibitors; thus, the potency of S-citalopram and paroxetine is reduced more than 40-fold. A cross-species chimera strategy was undertaken and followed by species-scanning mutagenesis. Differences in pharmacological profiles were tracked to amino acid residues 169, 172, and 586 in human SERT. Structure-activity studies on structurally related compounds indicated that species divergences in drug sensitivity between human and chicken SERT were arising from differences in coordination or recognition of an important aminomethyl pharmacophoric substructure, which is shared by all high affinity antidepressants. Consequently, we suggest that Ala(169) and Ile(172) of human SERT are important residues in sensing the N-methylation state of SERT antagonists. PMID- 15271994 TI - Molecular cloning and functional analysis of zebrafish neutral ceramidase. AB - Almost all observations on the functions of neutral ceramidase have been carried out at cellular levels but not at an individual level. Here, we report the molecular cloning of zebrafish neutral ceramidase (znCD) and its functional analysis during embryogenesis. We isolated a cDNA clone encoding znCD by 5' and 3' rapid amplification of cDNA ends-PCR. It possessed an open reading frame of 2,229 base pairs encoding 743 amino acids. A possible signal/anchor sequence near the N terminus and four potential O-glycosylation and eight potential N glycosylation sites were found in the putative sequence. The enzyme activity at neutral pH increased markedly after transformation of Chinese hamster CHOP and zebrafish BRF41 cells with the cDNA. The overexpressed enzyme was found to be distributed in endoplasmic reticulum/Golgi compartments as well as the plasma membranes. The antisense morpholino oligonucleotide (AMO), which was designed based on the sequence of znCD mRNA, successfully blocked the translation of znCD in a wheat germ in vitro translation system. The knockdown of znCD with AMO led to an increase in the number of zebrafish embryos with severe morphological and cellular abnormalities such as abnormal morphogenesis in the head and tail, pericardiac edema, defect of blood cell circulation, and an increase of apoptotic cells, especially in the head and neural tube regions, at 36 h post fertilization. The ceramide level in AMO-injected embryos increased significantly compared with that in control embryos. Simultaneous injection of both AMO and synthetic znCD mRNA into one-cell-stage embryos rescued znCD activity and blood cell circulation. These results indicate that znCD is essential for the metabolism of ceramide and the early development of zebrafish. PMID- 15271995 TI - Reverse two-hybrid screening identifies residues of JNK required for interaction with the kinase interaction motif of JNK-interacting protein-1. AB - The development of specific inhibitors for the c-Jun N-terminal kinase (JNK) family of mitogen-activated protein kinases (MAPKs) has been a recent research focus because of the association of JNK with cell death in conditions such as stroke and neurodegeneration. We have demonstrated previously the presence of critical inhibitory residues within an 11-mer peptide (TI-JIP) based on the sequence of JNK-interacting protein-1 (JIP-1). However, the corresponding region of JNK bound by this JIP-1-based peptide was unknown. To identify this region, we used a novel reverse two-hybrid approach with TI-JIP as bait. We screened a library of JNK1 mutants that had been generated by random PCR mutagenesis and found three mutants of JNK1 that failed to interact with TI-JIP. The mutations in JNK1 were L131R, R309W, and Y320H. Of these mutated residues, Leu-131 and Tyr-320 were located on a common face of the JNK protein close to other residues implicated previously in the interactions of MAPKs with substrates, phosphatases, and scaffolds. To test whether these JNK1 mutants were thus affected in their regulation, we evaluated their activation in mammalian cells in response to hyperosmolarity or cotransfection with a constitutively active upstream kinase or their direct phosphorylation by either MAPK kinase (MKK)4 or MKK7. In each situation, all three JNK mutants were not activated or phosphorylated to the same level as wild-type JNK. Therefore, the results of our unbiased reverse two-hybrid screening approach have identified residues of JNK responsible for binding JIP-1 based peptides as well as MKK4 or MKK7. PMID- 15271996 TI - Proteomic identification of Bcl2-associated athanogene 2 as a novel MAPK activated protein kinase 2 substrate. AB - The p38 MAPK cascade is activated by various stresses or cytokines. Downstream of p38 MAPKs, there are diversification and extensive branching of signaling pathways. Fluorescent two-dimensional difference gel electrophoresis of phosphoprotein-enriched samples from HeLa cells in which p38 MAPK activity was either suppressed or activated enabled us to detect approximately 90 candidate spots for factors involved in p38-dependent pathways. Among these candidates, here we identified four proteins including Bcl-2-associated athanogene 2 (BAG2) by peptide mass fingerprintings. BAG family proteins are highly conserved throughout eukaryotes and regulate Hsc/Hsp70-mediated molecular chaperone activities and apoptosis. The results of two-dimensional immunoblots suggested that the phosphorylation of BAG2 was specifically controlled in a p38 MAPK dependent manner. Furthermore, BAG2 was directly phosphorylated at serine 20 in vitro by MAPK-activated protein kinase 2 (MAPKAP kinase 2), which is known as a primary substrate of p38 MAPK and mediates several p38 MAPK-dependent processes. We confirmed that MAPKAP kinase 2 is also required for phosphorylation of BAG2 in vivo. Thus, p38 MAPK-MAPKAP kinase 2-BAG2 phosphorylation cascade may be a novel signaling pathway for response to extracellular stresses. PMID- 15271997 TI - The S box of major histocompatibility complex class II promoters is a key determinant for recruitment of the transcriptional co-activator CIITA. AB - Tightly regulated expression of major histocompatibility complex (MHC) class II genes is critical for the immune system. A conserved regulatory module consisting of four cis-acting elements, the W, X, X2 and Y boxes, controls transcription of MHC class II genes. The X, X2, and Y boxes are bound, respectively, by RFX, CREB, and NF-Y to form a MHC class II-specific enhanceosome complex. The latter constitutes a landing pad for recruitment of the transcriptional co-activator CIITA. In contrast to the well defined roles of the X, X2, and Y boxes, the role of the W region has remained controversial. In vitro binding studies have suggested that it might contain a second RFX-binding site. We demonstrate here by means of promoter pull-down assays that the most conserved subsequence within the W region, called the S box, is a critical determinant for tethering of CIITA to the enhanceosome complex. Binding of CIITA to the enhanceosome requires both integrity of the S box and a remarkably stringent spacing between the S and X boxes. Even a 1-2-base pair change in the native S-X distance is detrimental for CIITA recruitment and promoter function. In contrast to current models, binding of RFX to a putative duplicated binding site in the W box is thus not required for either CIITA recruitment or promoter activity. This paves the way for the identification of novel factors mediating the contribution of the S box to the activation of MHC class II promoters. PMID- 15271998 TI - Probing the dynamics of a mobile loop above the active site of L1, a metallo-beta lactamase from Stenotrophomonas maltophilia, via site-directed mutagenesis and stopped-flow fluorescence spectroscopy. AB - A structural feature shared by the metallo-beta-lactamases is a flexible loop of amino acids that extends over their active sites and that has been proposed to move during the catalytic cycle of the enzymes, clamping down on substrate. To probe the movement of this loop (residues 152-164), a site-directed mutant of metallo-beta-lactamase L1 was engineered that contained a Trp residue on the loop to serve as a fluorescent probe. It was necessary first, however, to evaluate the contribution of each native Trp residue to the fluorescence changes observed during the catalytic cycle of wild-type L1. Five site-directed mutants of L1 (W39F, W53F, W204F, W206F, and W269F) were prepared and characterized using metal analyses, CD spectroscopy, steady-state kinetics, stopped-flow fluorescence, and fluorescence titrations. All mutants retained the wild-type tertiary structure and bound Zn(II) at levels comparable with wild type and exhibited only slight (<10-fold) decreases in k(cat) values as compared with wild-type L1 for all substrates tested. Fluorescence studies revealed a single mutant, W39F, to be void of the fluorescence changes observed with wild-type L1 during substrate binding and catalysis. Using W39F as a template, a Trp residue was added to the flexile loop over the active site of L1, to generate the double mutant, W39F/D160W. This double mutant retained all the structural and kinetic characteristics of wild-type L1. Stopped-flow fluorescence and rapid-scanning UV visible studies revealed the motion of the loop (k(obs) = 27 +/- 2 s(-1)) to be similar to the formation rate of a reaction intermediate (k(obs) = 25 +/- 2 s( 1)). PMID- 15271999 TI - Interleukin-13 stimulates the transcription of the human alpha2(I) collagen gene in human dermal fibroblasts. AB - Interleukin (IL)-13 is a novel lymphokine produced by activated Type 2 helper cells. In this study, we examined the target genes of IL-13 by the cDNA microarray analysis in human dermal fibroblasts. We focused on the human alpha2(I) collagen gene, which was one of the IL-13-induced genes by the microarray analysis. IL-13 induced type I collagen protein as well as mRNA in a dose-dependent manner. Actinomycin D, an RNA synthesis inhibitor, significantly blocked the IL-13-mediated up-regulation of alpha2(I) collagen mRNA expression, whereas cycloheximide, a protein synthesis inhibitor, did not block this up regulation. In addition, IL-13 treatment induced the promoter activity of alpha2(I) collagen by nuclear run-on transcription assay and chloramphenicol acetyltransferase assay. IL-13-mediated transcriptional activation of alpha2(I) collagen gene or type I collagen protein up-regulation was inhibited by the treatment of fibroblasts with a selective phosphoinositide 3-kinase (PI3K) inhibitor, LY294002, or STAT6 antisense oligonucleotide, but not by PD98059, a specific inhibitor of MEK/ERK, or SB202190 or SB203580, specific inhibitors of p38 MAPK; IL-13 induced the phosphorylation of PI3K p85 regulatory subunit and STAT6. These results suggest that IL-13 may play a role in the regulation of extracellular matrix and indicate the possible therapeutic value of the blockade of IL-13 signaling pathways via PI3K and STAT6 in fibrosis. PMID- 15272000 TI - Identification of the tRNA-binding protein Arc1p as a novel target of in vivo biotinylation in Saccharomyces cerevisiae. AB - Biotin is an essential cofactor of cell metabolism serving as a protein-bound coenzyme in ATP-dependent carboxylation, in transcarboxylation, and certain decarboxylation reactions. The involvement of biotinylated proteins in other cellular functions has been suggested occasionally, but available data on this are limited. In the present study, a Saccharomyces cerevisiae protein was identified that reacts with streptavidin on Western blots and is not identical to one of the known biotinylated yeast proteins. After affinity purification on monomeric avidin, the biotinylated protein was identified as Arc1p. Using 14C labeled biotin, the cofactor was shown to be incorporated into Arc1p by covalent and alkali-stable linkage. Similar to the known carboxylases, Arc1p biotinylation is mediated by the yeast biotin:protein ligase, Bpl1p. Mutational studies revealed that biotinylation occurs at lysine 86 within the N-terminal domain of Arc1p. In contrast to the known carboxylases, however, in vitro biotinylation of Arc1p is incomplete and increases with BPL1 overexpression. In accordance to this fact, Arc1p lacks the canonical consensus sequence of known biotin binding domains, and the bacterial biotin:protein ligase, BirA, is unable to use Arc1p as a substrate. Arc1p was shown previously to organize the association of MetRS and GluRS tRNA synthetases with their cognate tRNAs thereby increasing the substrate affinity and catalytic efficiency of these enzymes. Remarkably, not only biotinylated but also the biotin-free Arc1p obtained by replacement of lysine 86 with arginine were capable of restoring Arc1p function in both arc1Delta and arc1Deltalos1Delta mutants, indicating that biotinylation of Arc1p is not essential for activity. PMID- 15272001 TI - Lipid utilization, gluconeogenesis, and seedling growth in Arabidopsis mutants lacking the glyoxylate cycle enzyme malate synthase. AB - The aim of this research was to test the role of the glyoxylate cycle enzyme malate synthase (MLS) in lipid utilization, gluconeogenesis, and seedling growth in Arabidopsis. We hypothesized that in the absence of MLS, succinate produced by isocitrate lyase (ICL) could still feed into the tricarboxylic acid cycle, whereas glyoxylate could be converted to sugars using enzymes of the photorespiratory pathway. To test this hypothesis we isolated knock-out mls mutants and studied their growth and metabolism in comparison to wild type and icl mutant seedlings. In contrast to icl seedlings, which grow slowly and are unable to convert lipid into sugars (Eastmond, P. J., Germain, V., Lange, P. R., Bryce, J. H., Smith, S. M. & Graham, I. A. (2000) Proc. Natl. Acad. Sci. U. S. A. 97, 5669-5674), mls seedlings grow faster, use their lipid more rapidly, and are better able to establish as plantlets. Transcriptome and metabolome analyses show that icl seedlings exhibit many features characteristic of carbohydrate starvation, whereas mls seedlings differ relatively little from wild type. In the light mls seedlings generate more sugars than icl seedlings, and when fed with [14C]acetate, 14C-labeling of sugars is three times greater than in icl seedlings and more than half that in wild type seedlings. The mls seedlings also accumulate more glycine and serine than icl or wild type seedlings, consistent with a diversion of glyoxylate into these intermediates of the photorespiratory pathway. We conclude that, in contrast to bacteria and fungi in which MLS is essential for gluconeogenesis from acetate or fatty acids, MLS is partially dispensable for lipid utilization and gluconeogenesis in Arabidopsis seedlings. PMID- 15272002 TI - Diphtheria toxin-induced autophagic cardiomyocyte death plays a pathogenic role in mouse model of heart failure. AB - It is still not clear whether loss of cardiomyocytes through programmed cell death causes heart failure. To clarify the role of cell death in heart failure, we generated transgenic mice (TG) that express human diphtheria toxin receptor in the hearts. A mosaic expression pattern of the transgene was observed, and the transgene-expressing cardiomyocytes (17.3% of the total cardiomyocytes) were diffusely scattered throughout the ventricles. Intramuscular injection of diphtheria toxin induced complete elimination of the transgene-expressing cardiomyocytes within 7 days, and approximately 80% of TG showed pathophysiological features characteristic of heart failure and were dead within 14 days. Degenerated cardiomyocytes of the TG heart showed characteristic features indicative of autophagic cell death such as up-regulated lysosomal markers and abundant autophagosomes containing cytosolic organelles like cardiomyocytes of human dilated cardiomyopathy. The heart failure-inducible TG are a useful model for dilated cardiomyopathy, and provided evidence indicating that myocardial cell loss through autophagic cell death plays of a causal role in the pathogenesis heart failure. PMID- 15272003 TI - Serine and threonine phosphorylation of the low density lipoprotein receptor related protein by protein kinase Calpha regulates endocytosis and association with adaptor molecules. AB - The low density lipoprotein receptor-related protein (LRP) is a large receptor that participates in endocytosis, signaling pathways, and phagocytosis of necrotic cells. Mechanisms that direct LRP to function in these distinct pathways likely involve its association with distinct cytoplasmic adaptor proteins. We tested the hypothesis that the association of various adaptor proteins with the LRP cytoplasmic domain is modulated by its phosphorylation state. Phosphoamino acid analysis of metabolically labeled LRP revealed that this receptor is phosphorylated at serine, threonine, and tyrosine residues within its cytoplasmic domain, whereas inhibitor studies identified protein kinase Calpha (PKCalpha) as a kinase capable of phosphorylating LRP. Mutational analysis identified critical threonine and serine residues within the LRP cytoplasmic domain that are necessary for phosphorylation mediated by PKCalpha. Mutating these threonine and serine residues to alanines generated a receptor that was not phosphorylated and that was internalized more rapidly than wild-type LRP, revealing that phosphorylation reduces the association of LRP with adaptor molecules of the endocytic machinery. In contrast, serine and threonine phosphorylation was necessary for the interaction of LRP with Shc, an adaptor protein that participates in signaling events. Furthermore, serine and threonine phosphorylation increased the interaction of LRP with other adaptor proteins such as Dab-1 and CED-6/GULP. These results indicate that phosphorylation of LRP by PKCalpha modulates the endocytic and signaling function of LRP by modifying its association with adaptor proteins. PMID- 15272004 TI - Overexpression of beta2-adrenergic receptors cAMP-dependent protein kinase phosphorylates and modulates slow delayed rectifier potassium channels expressed in murine heart: evidence for receptor/channel co-localization. AB - The cardiac slow delayed rectifier potassium channel (IKs), comprised of (KCNQ1) and beta (KCNE1) subunits, is regulated by sympathetic nervous stimulation, with activation of beta-adrenergic receptors PKA phosphorylating IKs channels. We examined the effects of 2-adrenergic receptors (beta2-AR) on IKs in cardiac ventricular myocytes from transgenic mice expressing fusion proteins of IKs subunits and hbeta2-ARs. KCNQ1 and beta2-ARs were localized to the same subcellular regions, sharing intimate localization within nanometers of each other. In IKs/B2-AR myocytes, IKs density was increased, and activation shifted in the hyperpolarizing direction; IKs was not further modulated by exposure to isoproterenol, and KCNQ1 was found to be PKA-phosphorylated. Conversely, beta2-AR overexpression did not affect L-type calcium channel current (ICaL) under basal conditions with ICaL remaining responsive to cAMP. These data indicate intimate association of KCNQ1 and beta2-ARs and that beta2-AR signaling can modulate the function of IKs channels under conditions of increased beta2-AR expression, even in the absence of exogenous beta-AR agonist. PMID- 15272005 TI - Direct binding of visual arrestin to microtubules determines the differential subcellular localization of its splice variants in rod photoreceptors. AB - Proper function of visual arrestin is indispensable for rapid signal shut-off in rod photoreceptors. Dramatic light-dependent changes in its subcellular localization are believed to play an important role in light adaptation of photoreceptor cells. Here we show that visual arrestin binds microtubules. The truncated splice variant of visual arrestin, p44, demonstrates dramatically higher affinity for microtubules than the full-length protein (p48). Enhanced microtubule binding of p44 underlies its earlier reported preferential localization to detergent-resistant membranes, where it is anchored via membrane associated microtubules in a rhodopsin-independent fashion. Experiments with purified proteins demonstrate that arrestin interaction with microtubules is direct and does not require any additional protein partners. Most importantly, arrestin interactions with microtubules and light-activated phosphorylated rhodopsin are mutually exclusive, suggesting that microtubule interaction may play a role in keeping p44 arrestin away from rhodopsin in dark-adapted photoreceptors. PMID- 15272006 TI - A novel role for nuclear factor of activated T cells in receptor tyrosine kinase and G protein-coupled receptor agonist-induced vascular smooth muscle cell motility. AB - In addition to their role in cytokine gene regulation in T cells, nuclear factors of activated T cells (NFATs) have been shown to be involved in cardiac development and hypertrophy. We have reported previously that NFATs play an important role in the regulation of vascular smooth muscle cell (VSMC) proliferation by receptor tyrosine kinase (RTK) and G protein-coupled receptor (GPCR) agonists, platelet-derived growth factor-BB (PDGF-BB) and thrombin, respectively. To understand the role of NFATs in vascular disease and development, we have now studied the role of these transcriptional factors in VSMC motility. PDGF-BB and thrombin induced VSMC motility in a dose-dependent manner. Blockade of NFAT activation resulted in substantial reduction in PDGF-BB- and thrombin-induced VSMC motility. PDGF-BB and thrombin also induced interleukin 6 (IL-6) expression in NFAT-dependent manner. Furthermore, IL-6 dose-dependently caused VSMC motility. A neutralizing anti-rat IL-6 antibody inhibited VSMC motility induced by IL-6, PDGF-BB, and thrombin. In addition, exogenous addition of IL-6 rescued both PDGF-BB- and thrombin-induced VSMC motility from inhibition by the blockade of NFAT activation. Together, these results for the first time demonstrate that NFATs mediate both RTK and GPCR agonist-induced VSMC motility via induction of expression of IL-6. PMID- 15272007 TI - Tyrosine-phosphorylated and nonphosphorylated sodium channel beta1 subunits are differentially localized in cardiac myocytes. AB - Voltage-gated sodium channel alpha and beta subunits expressed in mammalian heart are differentially localized to t-tubules and intercalated disks. Sodium channel beta subunits are multifunctional molecules that participate in channel modulation and cell adhesion. Reversible, receptor-mediated changes in beta1 tyrosine phosphorylation modulate its ability to recruit and associate with ankyrin. The purpose of the present study was to test our hypothesis that tyrosine-phosphorylated beta1 (pYbeta1) and nonphosphorylated beta1 subunits may be differentially localized in heart and thus interact with different cytoskeletal and signaling proteins. We developed an antibody that specifically recognizes pYbeta1 and investigated the differential subcellular localization of beta1 and pYbeta1 in mouse ventricular myocytes. We found that pYbeta1 colocalized with connexin-43, N-cadherin, and Nav1.5 at intercalated disks but was not detected at the t-tubules. Anti-pYbeta1 immunoprecipitates N-cadherin from heart membranes and from cells transfected with beta1 and N-cadherin in the absence of other sodium channel subunits. pYbeta1 does not associate with ankyrinB in heart membranes. N-cadherin and connexin-43 associate with Nav1.5 in heart membranes as assessed by co-immunoprecipitation assays. We propose that sodium channel complexes at intercalated disks of ventricular myocytes are composed of Nav1.5 and pYbeta1 and that these complexes are in close association with both N-cadherin and connexin-43. beta1 phosphorylation appears to regulate its localization to differential subcellular domains. PMID- 15272008 TI - Monitoring conformational rearrangements in the substrate-binding site of a membrane transport protein by mass spectrometry. AB - Combined biochemical, biophysical, and crystallographic studies on the lactose permease of Escherichia coli suggest that Arg-144 (helix V) forms a salt bridge with Glu-126 (helix IV), which is broken during substrate binding, thereby permitting the guanidino group to form a bidentate H-bond with the C-4 and C-3 O atoms of the galactopyranosyl moiety and an H-bond with Glu-269 (helix VIII). To examine the relative interaction of Arg-144 with these two potential salt bridge partners (Glu-126 and Glu-269) in the absence of substrate, the covalent modification of the guanidino group was monitored with the Arg-specific reagent butane-2,3-dione using electrospray ionization mass spectrometry. In a functional background, the reactivity of Arg-144 with butane-2,3-dione is low ( approximately 25%) and is reduced by a factor of approximately 2 by preincubation with ligand. Interestingly, although replacement of Glu-126 with Ala results in a 3-fold increase in the reactivity of Arg-144, replacement of Glu-269 with Ala elicits virtually no effect. Taken together, these results suggest that in the absence of substrate the interaction between Arg-144 and Glu-126 is much stronger than the interaction with Glu-269, supporting the contention that sugar recognition leads to rearrangement of charge-paired residues essential for sugar binding. PMID- 15272009 TI - Plasmodium induces swelling-activated ClC-2 anion channels in the host erythrocyte. AB - Intraerythrocytic growth of the human malaria parasite Plasmodium falciparum depends on delivery of nutrients. Moreover, infection challenges cell volume constancy of the host erythrocyte requiring enhanced activity of cell volume regulatory mechanisms. Patch clamp recording demonstrated inwardly and outwardly rectifying anion channels in infected but not in control erythrocytes. The molecular identity of those channels remained elusive. We show here for one channel type that voltage dependence, cell volume sensitivity, and activation by oxidation are identical to ClC-2. Moreover, Western blots and FACS analysis showed protein and functional ClC-2 expression in human erythrocytes and erythrocytes from wild type (Clcn2(+/+)) but not from Clcn2(-/-) mice. Finally, patch clamp recording revealed activation of volume-sensitive inwardly rectifying channels in Plasmodium berghei-infected Clcn2(+/+) but not Clcn2(-/-) erythrocytes. Erythrocytes from infected mice of both genotypes differed in cell volume and inhibition of ClC-2 by ZnCl(2) (1 mm) induced an increase of cell volume only in parasitized Clcn2(+/+) erythrocytes. Lack of ClC-2 did not inhibit P. berghei development in vivo nor substantially affect the mortality of infected mice. In conclusion, activation of host ClC-2 channels participates in the altered permeability of Plasmodium-infected erythrocytes but is not required for intraerythrocytic parasite survival. PMID- 15272010 TI - The DeltaF508 mutation disrupts packing of the transmembrane segments of the cystic fibrosis transmembrane conductance regulator. AB - The most common mutation in cystic fibrosis (deletion of Phe-508 in the first nucleotide binding domain (DeltaF508)) in the cystic fibrosis transmembrane conductance regulator (CFTR) causes retention of the mutant protein in the endoplasmic reticulum. We previously showed that the DeltaF508 mutation causes the CFTR protein to be retained in the endoplasmic reticulum in an inactive and structurally altered state. Proper packing of the transmembrane (TM) segments is critical for function because the TM segments form the chloride channel. Here we tested whether the DeltaF508 mutation altered packing of the TM segments by disulfide cross-linking analysis between TM6 and TM12 in wild-type and DeltaF508 CFTRs. These TM segments were selected because TM6 appears to line the chloride channel, and cross-linking between these TM segments has been observed in the CFTR sister protein, the multidrug resistance P-glycoprotein. We first mapped potential contact points in wild-type CFTR by cysteine mutagenesis and thiol cross-linking analysis. Disulfide cross-linking was detected in CFTR mutants M348C(TM6)/T1142C(TM12), T351C(TM6)/T1142C(TM12), and W356C(TM6)/W1145C(TM12) in a wild-type background. The disulfide cross-linking occurs intramolecularly and was reducible by dithiothreitol. Introduction of the DeltaF508 mutation into these cysteine mutants, however, abolished cross-linking. The results suggest that the DeltaF508 mutation alters interactions between the TM domains. Therefore, a potential target to correct folding defects in the DeltaF508 mutant of CFTR is to identify compounds that promote correct folding of the TM domains. PMID- 15272011 TI - Fluorescent biosensor for quantitative real-time measurements of inositol 1,4,5 trisphosphate in single living cells. AB - The second messenger inositol 1,4,5-trisphosphate (IP(3)) plays a central role in the generation of a variety of spatiotemporally complex intracellular Ca(2+) signals involved in the regulation of many essential physiological processes. Here we describe the development of "LIBRA", a novel ratiometric fluorescent IP(3) biosensor that allows for the quantitative monitoring of intracellular IP(3) concentrations in single living cells in real time. LIBRA consists of the IP(3)-binding domain of the rat type 3 IP(3) receptor fused between the fluorescence resonance energy transfer pair cyan fluorescent protein and yellow fluorescent protein and preceded by a membrane-targeting signal. We show that the LIBRA fluorescent signal is highly selective for IP(3) and unaffected by concentrations of Ca(2+) and ATP in the physiological range. In addition, LIBRA can be calibrated in situ. We demonstrate the utility of LIBRA by monitoring the temporal relationship between the responses intracellular IP(3) and Ca(2+) concentrations in SH-SY5Y cells following acetylcholine stimulation. PMID- 15272012 TI - Sustained entry of Ca2+ is required to activate Ca2+-calmodulin-dependent phosphodiesterase 1A. AB - Regulation of adenylyl cyclases (ACs) by Ca2+ requires capacitative Ca2+ entry (CCE) (Cooper, D. M. F. (2003) Biochem. J. 375, 517-529), but whether Ca2+ sensitive phosphodiesterases (PDEs) are similarly discriminating has never been addressed. In the present study, a variety of conditions were devised to manipulate [Ca2+]i so that we could ask whether PDE1 selectively responds to different modes of elevating [Ca2+]i, viz. Ca2+ released from intracellular stores and various modes of Ca2+ entry. In 1321N1 human astrocytoma cells, the endogenous PDE1 (identified as PDE1A by reverse transcriptase-PCR) was largely insensitive to Ca2+ released from carbachol-sensitive stores but was robustly stimulated by a similar rise in [Ca2+]i due to carbachol-induced Ca2+ influx. Gd3+, which effectively blocked thapsigargin-induced CCE and its effect on PDE1A, also inhibited the activation of PDE1A by carbachol-induced Ca2+ entry. However, non-selective ionomycin-mediated Ca2+ entry also activated PDE1A, so that, unlike Ca2+-sensitive ACs, PDE1A cannot discriminate between the different sources of Ca2+ entry. Fractionation of the cells revealed that the Ca2+-calmodulin stimulated PDE activity was not present at the plasma membrane but was associated with the cytosol and the organellar compartments of the cell. Therefore, the apparent disparity between PDE1A and ACs is likely to be the consequence of their differential subcellular localization. Nevertheless, in a physiological context, where artificial modes of elevating [Ca2+]i are not available, as with ACs, a dependence on CCE would be evident, and it would be the duration of this influx of Ca2+ that would determine how long PDE1A was activated. PMID- 15272013 TI - SHEP1 function in cell migration is impaired by a single amino acid mutation that disrupts association with the scaffolding protein cas but not with Ras GTPases. AB - SHEP1 is a signaling protein that contains a guanine nucleotide exchange factor like domain, which binds Ras family GTPases and also forms a stable complex with the scaffolding protein Crk-associated substrate (Cas). SHEP1 and Cas have several common functions, such as increasing c-Jun N-terminal kinase activity, promoting T cell activation, and regulating the actin cytoskeleton. However, it is unclear whether a physical association between SHEP1 and Cas is required for these activities. We reported previously that SHEP1 is tyrosine-phosphorylated downstream of the EphB2 receptor; in this study, we further demonstrate that activated EphB2 inhibits SHEP1 association with Cas. To investigate whether phosphorylation negatively regulates the SHEP1-Cas complex, we have identified by mass spectrometry several SHEP1 tyrosine phosphorylation sites downstream of EphB2; of particular interest among them is tyrosine 635 in the Cas association/exchange factor domain. Mutation of this tyrosine to glutamic acid, but not to phenylalanine, disrupts Cas binding to SHEP1 without inhibiting Ras GTPase binding. The glutamic acid mutation also makes SHEP1 unable to promote Cas Crk association, membrane ruffling, and cell migration toward epidermal growth factor (EGF), implying that these activities of SHEP1 depend upon a physical interaction with Cas. Association with Cas also seems to be necessary for EGF induced SHEP1 tyrosine phosphorylation, which is mediated by a Src family kinase. It is noteworthy that EGF stimulation does not cause dissociation of SHEP1 from Cas. These data show that SHEP1 regulates membrane ruffling and cell migration and that binding to Cas is probably critical for these functions. Furthermore, the SHEP1-Cas complex may have different roles downstream of EphB2 and the EGF receptor. PMID- 15272014 TI - Augmentation of tissue transglutaminase expression and activation by epidermal growth factor inhibit doxorubicin-induced apoptosis in human breast cancer cells. AB - Tissue transglutaminase (TGase) exhibits both a GTP binding/hydrolytic capability and an enzymatic transamidation activity. Increases in TGase expression and activation often occur in response to stimuli that promote cellular differentiation and apoptosis, yet the signaling mechanisms used by these stimuli to regulate TGase expression and activation and the role of TGase in these cellular processes are not well understood. Retinoic acid (RA) consistently induces TGase expression and activation, and it was shown recently that RA induced TGase expression was inhibited in NIH3T3 mouse fibroblasts co-stimulated with epidermal growth factor (EGF). Here we investigate whether EGF also antagonized RA-induced TGase expression in breast cancer cells. We found that EGF stimulation affected TGase expression and activation very differently in these cancer cells. Not only did EGF fail to block RA-induced TGase expression, but also EGF alone was sufficient to potently up-regulate TGase expression and activation in SKBR3 cells, as well as MDAMB468 and BT-20 cells. Inhibiting phosphoinositide 3-kinase activity severely diminished the ability of EGF and RA to increase TGase protein levels, whereas a constitutively active form of phosphoinositide 3-kinase potentiated the induction of TGase expression by EGF in SKBR3 cells. Because EGF is an established antiapoptotic factor, we examined whether the protection afforded by EGF was dependent on its ability to up regulate TGase activity in SKBR3 and BT-20 cells. Exposure of cells to a TGase inhibitor or expression of a dominant-negative form of TGase potently inhibited EGF-mediated protection from doxorubicin-induced apoptosis. Moreover, expression of exogenous TGase in SKBR3 cells mimicked the survival advantage of EGF, suggesting that TGase activation is necessary and sufficient for the antiapoptotic properties of EGF. These findings indicate for the first time that EGF can induce TGase expression and activation in human breast cancer cells and that this contributes to their oncogenic potential by promoting chemoresistance. PMID- 15272015 TI - A spontaneous point mutation produces monoamine oxidase A/B knock-out mice with greatly elevated monoamines and anxiety-like behavior. AB - A spontaneous monoamine oxidase A (MAO A) mutation (A863T) in exon 8 introduced a premature stop codon, which produced MAO A/B double knock-out (KO) mice in a MAO B KO mouse colony. This mutation caused a nonsense-mediated mRNA decay and resulted in the absence of MAO A transcript, protein, and catalytic activity and abrogates a DraI restriction site. The MAO A/B KO mice showed reduced body weight compared with wild type mice. Brain levels of serotonin, norepinephrine, dopamine, and phenylethylamine increased, and serotonin metabolite 5 hydroxyindoleacetic acid levels decreased, to a much greater degree than in either MAO A or B single KO mice. Observed chase/escape and anxiety-like behavior in the MAO A/B KO mice, different from MAO A or B single KO mice, suggest that varying monoamine levels result in both a unique biochemical and behavioral phenotype. These mice will be useful models for studying the molecular basis of disorders associated with abnormal monoamine neurotransmitters. PMID- 15272016 TI - In vitro modification of human centromere protein CENP-C fragments by small ubiquitin-like modifier (SUMO) protein: definitive identification of the modification sites by tandem mass spectrometry analysis of the isopeptides. AB - Protein sumoylation by small ubiquitin-like modifier (SUMO) proteins is an important post-translational regulatory modification. A role in the control of chromosome dynamics was first suggested when SUMO was identified as high-copy suppressor of the centromere protein CENP-C mutants. CENP-C itself contains a consensus sumoylation sequence motif that partially overlaps with its DNA binding and centromere localization domain. To ascertain whether CENP-C can be sumoylated, tandem mass spectrometry (MS) based strategy was developed for high sensitivity identification and sequencing of sumoylated isopeptides present among in-gel-digested tryptic peptides of SDS-PAGE fractionated target proteins. Without a predisposition to searching for the expected isopeptides based on calculated molecular mass and relying instead on the characteristic MS/MS fragmentation pattern to identify sumolylation, we demonstrate that several other lysine residues located not within the perfect consensus sumoylation motif psiKXE/D, where psi represents a large hydrophobic amino acid, and X represents any amino acid, can be sumolylated with a reconstituted in vitro system containing only the SUMO proteins, E1-activating enzyme and E2-conjugating enzyme (Ubc9). In all cases, target sites that can be sumoylated by SUMO-2 were shown to be equally susceptible to SUMO-1 attachments which include specific sites on SUMO 2 itself, Ubc9, and the recombinant CENP-C fragments. Two non-consensus sites on one of the CENP-C fragments were found to be sumoylated in addition to the predicted site on the other fragment. The developed methodologies should facilitate future studies in delineating the dynamics and substrate specificities of SUMO-1/2/3 modifications and the respective roles of E3 ligases in the process. PMID- 15272017 TI - Osmotic diuretics induce adenosine A1 receptor expression and protect renal proximal tubular epithelial cells against cisplatin-mediated apoptosis. AB - Osmotic diuretics are used successfully to alleviate acute tubular necrosis (ATN) produced by chemotherapeutic agents and aminoglycoside antibiotics. The beneficial action of these agents likely involves rapid elimination of the nephrotoxic agents from the kidney by promoting diuresis. Adenosine A1 receptor (A1AR) subtype present on renal proximal tubular epithelial and cortical collecting duct cells mediates the antidiuretic and cytoprotective actions of adenosine. These receptors are induced by activation of nuclear factor (NF) kappaB, a transcription factor reported to mediate hyperosmotic stress-induced cytoprotection in renal medullary cells. In this study, we tested the hypothesis that induction of the A1AR in renal proximal tubular cells by NF-kappaB contributes to the cytoprotection afforded by osmotic diuretics. Exposure of porcine renal proximal tubular epithelial (LLC-PK1) cells to mannitol or NaCl produced a significant increase in A1AR. This increase was preceded by adenosine release and NF-kappaB activation. Expression of an IkappaB-alpha mutant, which acts as a superrepressor of NF-kappaB, abrogated the increase in A1AR. Cells exposed to mannitol demonstrated increased reactive oxygen species (ROS) generation, which was attenuated by inhibiting xanthine oxidase with allopurinol. Allopurinol attenuated both the increase in A1AR expression and NF-kappaB activation produced by osmotic diuretics, indicating a role of adenosine metabolites in these processes. Treatment of LLC-PK1 cells with cisplatin (8 microm) resulted in apoptosis, which was attenuated by mannitol but exacerbated by selective A1AR blockade. Administration of mannitol to mice increases A1AR expression and activation of NF-kappaB in renal cortical sections. Taken together, these data provide novel mechanisms of nephroprotection by osmotic diuretics, involving both activation and induction of the A1AR, the latter mediated through activation of a xanthine oxidase pathway leading to ROS generation and promoting activation of NF-kappaB. PMID- 15272018 TI - Gi2 signaling enhances proliferation of neural progenitor cells in the developing brain. AB - Our previous study showed that the pertussis toxin-sensitive G protein, Gi2, is selectively localized in the ventricular zone of embryonic brains, where the neuroepithelial cells undergo active proliferation. In order to clarify the role of Gi2 in this site, we first administered pertussis toxin by an exo-utero manipulation method into the lateral ventricle of mouse brain at embryonic day 14.5. Examination at embryonic day 18.5 revealed that pertussis toxin-injected embryos had brains with thinner cerebral cortices, made up of fewer constituent cells. Bromodeoxyuridine labeling revealed fewer numbers of bromodeoxyuridine positive cells in the cerebral cortices of pertussis toxin-injected embryos, suggesting impaired proliferation of neuroepithelial cells. Next we cultured neural progenitor cells from rat embryonic brains and evaluated the mitogenic effects of agonists for several Gi-coupled receptors that are known to be expressed in the ventricular zone. Among agonists tested, endothelin most effectively stimulated the incorporation of [3H]thymidine in the presence of fibronectin, via the endothelin-B receptor. This was associated with phosphorylation of extracellular signal-regulated kinase, and pertussis toxin partially inhibited both endothelin-stimulated DNA synthesis and phosphorylation of extracellular signal-regulated kinase. Injection of endothelin-3 into the ventricle of embryonic brains increased numbers of bromodeoxyuridine-positive cells in the cerebral cortex, whereas injection of an endothelin-B receptor antagonist decreased them. These findings indicate that Gi2 mediates signaling from receptors such as the endothelin-B receptor to maintain mitogenic activity in the neural progenitor cells of developing brain. PMID- 15272019 TI - Two different contact sites are recruited by cardiotrophin-like cytokine (CLC) to generate the CLC/CLF and CLC/sCNTFRalpha composite cytokines. AB - The cytokines of the interleukin-6 family are multifunctional proteins that regulate cell growth, differentiation, and other cell functions in a variety of biological systems including the immune, inflammatory, hematopoietic, and nervous systems. One member of this family, ciliary neurotrophic factor (CNTF), displays biological functions more restricted to the neuromuscular axis. We have recently identified two additional ligands for the CNTF receptor complex. Both are composite cytokines formed by cardiotrophin-like cytokine (CLC) associated to either the soluble type I cytokine receptor CLF or the soluble form of CNTF receptor alpha (CNTFRalpha). The present study was aimed at analyzing the interactions between the cytokine CLC and its different receptor chains. For this purpose, we modeled CLC/receptor interactions to define the residues potentially involved in the contact sites. We then performed site-directed mutagenesis on these residues and analyzed the biological interactions between mutants and receptor chains. Importantly, we found that CLC interacts with the soluble forms of CNTFRalpha and CLF via sites 1 and 3, respectively. For site 1, the most crucial residues involved in the interaction are Trp67, Arg170, and Asp174, which interact with CNTFRalpha. Surprisingly, the residues that are important for the interaction of CLC with CLF are part of the conserved FXXK motif of site 3 known to be the interaction site of LIFRbeta. Obtained results show that the Phe151 and Lys154 residues are effectively involved in the interaction of CLC with LIFRbeta. This study establishes the molecular details of the interaction of CLC with CLF, CNTFRalpha, and LIFRbeta and helps to define the precise role of each protein in this functional receptor complex. PMID- 15272020 TI - Skeletal muscle FOXO1 (FKHR) transgenic mice have less skeletal muscle mass, down regulated Type I (slow twitch/red muscle) fiber genes, and impaired glycemic control. AB - FOXO1, a member of the FOXO forkhead type transcription factors, is markedly up regulated in skeletal muscle in energy-deprived states such as fasting and severe diabetes, but its functions in skeletal muscle have remained poorly understood. In this study, we created transgenic mice specifically overexpressing FOXO1 in skeletal muscle. These mice weighed less than the wild-type control mice, had a reduced skeletal muscle mass, and the muscle was paler in color. Microarray analysis revealed that the expression of many genes related to the structural proteins of type I muscles (slow twitch, red muscle) was decreased. Histological analyses showed a marked decrease in size of both type I and type II fibers and a significant decrease in the number of type I fibers in the skeletal muscle of FOXO1 mice. Enhanced gene expression of a lysosomal proteinase, cathepsin L, which is known to be up-regulated during skeletal muscle atrophy, suggested increased protein degradation in the skeletal muscle of FOXO1 mice. Running wheel activity (spontaneous locomotive activity) was significantly reduced in FOXO1 mice compared with control mice. Moreover, the FOXO1 mice showed impaired glycemic control after oral glucose and intraperitoneal insulin administration. These results suggest that FOXO1 negatively regulates skeletal muscle mass and type I fiber gene expression and leads to impaired skeletal muscle function. Activation of FOXO1 may be involved in the pathogenesis of sarcopenia, the age related decline in muscle mass in humans, which leads to obesity and diabetes. PMID- 15272021 TI - Identification of placenta growth factor determinants for binding and activation of Flt-1 receptor. AB - Placenta growth factor (PlGF) belongs to the vascular endothelial growth factor (VEGF) family and represents a key regulator of angiogenic events in pathological conditions. PlGF exerts its biological function through the binding and activation of the seven immunoglobulin-like domain receptor Flt-1, also known as VEGFR-1. Here, we report the first detailed mutagenesis studies that provide a basis for understanding molecular recognition between PlGF-1 and Flt-1, highlighting some of the residues that are critical for receptor recognition. Mutagenesis analysis, performed on the basis of a structural model of interaction between PlGF and the minimal binding domain of Flt-1, has led to the identification of several PlGF-1 residues involved in Flt-1 recognition. The two negatively charged residues, Asp-72 and Glu-73, located in the beta3-beta4 loop, are critical for Flt-1 binding. Other mutations, which bring about a significant decrease in PlGF binding activity, are Gln-27, located in the N-terminal alpha helix, and Pro-98 and Tyr-100 on the beta6 strand. The mutation of one of the two glycosylated residues of PlGF, Asn-84, generates a PlGF variant with reduced binding activity. This indicates that, unlike in VEGF, glycosylation plays an important role in Flt-1 binding. The double mutation of residues Asp-72 and Glu 73 generates a PlGF variant unable to bind and activate the receptor molecules on the cell surface. This variant failed to induce in vitro capillary-like tube formation of primary endothelial cells or neo-angiogenesis in an in vivo chorioallantoic membrane assay. PMID- 15272022 TI - A ternary complex of transcription factors, Nished and NFATc4, and co-activator p300 bound to an intronic sequence, intronic regulatory element, is pivotal for the up-regulation of myosin light chain-2v gene in cardiac hypertrophy. AB - Transcriptional up-regulation of the myosin light chain-2 (MLC-2v) gene is an established marker for hypertrophic response in cardiomyocytes. Despite the documentation on the role of several cis-elements in the MLC-2v gene and their cognate proteins in transcription, the mechanism that dictates the preferential increase in MLC-2v gene expression during myocardial hypertrophy has not been delineated. Here we describe the properties of a cardiac specific intronic activator element (IRE) that shares sequence homology with the repressor element, the cardiac specific sequence, in the chicken MLC-2v gene. The transcription factor, Nished, that recognizes both IRE and the cardiac specific sequence potentiates the transcription of the MLC-2v gene via interaction with another transcription factor, nuclear factor of activated T cells, and the co-activator p300 at the IRE site. Angiotensin II (Ang II), a potent agonist of hypertrophy, causes induction of the MLC-2v gene transcription, which correlates well with the enhanced binding of Nished-nuclear factor of the activated T cells-p300 complex to IRE in the gel mobility shift assay. Losartan, an antagonist of Ang II receptor (AT1), abolishes the agonist-dependent stimulation of IRE/protein interaction and the consequent increase in MLC-2v gene transcription. These results together have thus established a transcriptional role of IRE as a direct target sequence of Ang II-mediated signaling that appears to be pivotal in the mechanism underlying the up-regulation of the MLC-2v gene during cardiac hypertrophy. PMID- 15272023 TI - Necdin interacts with the Msx2 homeodomain protein via MAGE-D1 to promote myogenic differentiation of C2C12 cells. AB - Necdin is a potent growth suppressor that is expressed predominantly in postmitotic cells such as neurons and skeletal muscle cells. Necdin shows a significant homology to MAGE (melanoma antigen) family proteins, all of which contain a large homology domain. MAGE-D1 (NRAGE, Dlxin-1) interacts with the Dlx/Msx family homeodomain proteins via an interspersed hexapeptide repeat domain distinct from the homology domain. Here we report that necdin associates with the Msx homeodomain proteins via MAGE-D1 to modulate their function. In vitro binding and co-immunoprecipitation analyses revealed that MAGE-D1 directly interacted with necdin via the homology domain and Msx1 (or Msx2) via the repeat domain. A ternary complex of necdin, MAGE-D1, and Msx2 was formed in vitro, and an endogenous complex containing these three proteins was detected in differentiating embryonal carcinoma cells. Co-expression of necdin and MAGE-D1 released Msx-dependent transcriptional repression. C2C12 myoblast cells that were stably transfected with Msx2 cDNA showed a marked reduction in myogenic differentiation, and co-expression of necdin and MAGE-D1 canceled the Msx2 dependent repression. These results suggest that necdin and MAGE-D1 cooperate to modulate the function of Dlx/Msx homeodomain proteins in cellular differentiation. PMID- 15272024 TI - A cascade of 24 histatins (histatin 3 fragments) in human saliva. Suggestions for a pre-secretory sequential cleavage pathway. AB - The systematic search by tandem mass spectrometry of human saliva from four different subjects, of 136 possible fragments originated from histatin 3, allowed the detection of 24 different peptides. They include, with the exception of histatin 4, all the known histatin 3 fragments, namely histatins 5-12 and the peptides corresponding to 15-24, 26-32, 29-32 residues, and 13 new fragments corresponding to 1-11, 1-12, 1-13, 5-13, 6-11, 6-13, 7-11, 7-12, 7-13, 14-24, 14 25, 15-25, and 28-32 residues of histatin 3. On the contrary, none of 119 possible fragments of histatin 1, including histatin 2, was detected. The results suggest that the genesis of histatin 3-related peptides, being under the principal action of trypsin-like activities, is probably not a random process but rather follows a sequential fragmentation pathway. Lack of detection of C terminal fragments, with the exception of 26-32, 28-32, and 29-32 fragments, suggested that arginine 25 should be the first cleavage site, generating histatin 6 and 26-32 fragments. The genesis of 28-32 and 29-32 fragments and histatin 5 should implicate a subsequent exo-protease action. Similarly, lack of detection of fragments having Lys-5 and Arg-6 at the N terminus and Arg-25 at the C terminus strongly suggested that sequences KRKF (11-14 residues) and AKR (4-6 residues) should be the second and the third cleavage sites, respectively. Lys-17 and Arg-22 are not cleaved at all. PMID- 15272025 TI - Insulin receptor substrate-1/SHP-2 interaction, a phenotype-dependent switching machinery of insulin-like growth factor-I signaling in vascular smooth muscle cells. AB - Insulin-like growth factor-I (IGF-I) plays a role in mutually exclusive processes such as proliferation and differentiation in a variety of cell types. IGF-I is a potent mitogen and motogen for dedifferentiated vascular smooth muscle cells (VSMCs) in vivo and in vitro. However, in differentiated VSMCs, IGF-I is only required for maintaining the differentiated phenotype. Here we investigated the VSMC phenotype-dependent signaling and biological processes triggered by IGF-I. In differentiated VSMCs, IGF-I activated a protein-tyrosine phosphatase, SHP-2, recruited by insulin receptor substrate-1 (IRS-1). The activated SHP-2 then dephosphorylated IRS-1 Tyr(P)-895, resulting in blockade of the pathways from IRS 1/Grb2/Sos to the ERK and p38 MAPK. Conversely, such negative regulation was silent in dedifferentiated VSMCs, where IGF-I activated both MAPKs via IRS 1/Grb2/Sos interaction-linked Ras activation, leading to proliferation and migration. Thus, our present results demonstrate that the IRS-1/SHP-2 interaction acts as a switch controlling VSMC phenotype-dependent IGF-I-induced signaling pathways and biological processes, and this mechanism is likely to be applicable to other cells. PMID- 15272026 TI - Protein phosphatase 2B dephosphorylates twitchin, initiating the catch state of invertebrate smooth muscle. AB - "Catch" is the state where some invertebrate muscles sustain high tension for long periods at low ATP hydrolysis rates. Physiological studies using muscle fibers have not yet fully provided the details of the initiation process of the catch state. The process was extensively studied by using an in vitro reconstitution assay with several phosphatase inhibitors. Actin filaments bound to thick filaments pretreated with the soluble protein fraction of muscle homogenate and Ca2+ (catch treatment) in the presence of MgATP at a low free Ca2+ concentration (the catch state). Catch treatment with > 50 microm okadaic acid, > 1 microm microcystin LR, 1 microm cyclosporin A, 1 microm FK506, or 0.2 mm calcineurin autoinhibitory peptide fragment produced almost no binding of the actin filaments, indicating protein phosphatase 2B (PP2B) was involved. Use of bovine calcineurin (PP2B) and its activator calmodulin instead of the soluble protein fraction initiated the catch state, indicating that only PP2B and calmodulin in the soluble protein fraction are essential for the initiation process. The initiation was reproduced with purified actin, myosin, twitchin, PP2B, and calmodulin. 32P autoradiography showed that only twitchin was dephosphorylated during the catch treatment with either the soluble protein fraction or bovine calcineurin and calmodulin. These results indicate that PP2B directly dephosphorylates twitchin and initiates the catch state and that no other component is required for the initiation process of the catch state. PMID- 15272027 TI - Functional dissection and molecular characterization of calcium-sensitive actin capping and actin-depolymerizing sites in villin. AB - All proteins of the villin superfamily, which includes the actin-capping and severing proteins such as gelsolin, scinderin, and severin, are calcium-regulated actin-modifying proteins. Like some of these proteins, villin has morphologically distinct effects on actin assembly depending on the free calcium concentrations. At physiological calcium (Ca2+) villin nucleates and bundles actin, whereas at higher concentrations it caps (>50 microm) and severs (>200 microM) actin filaments. Although Ca(2+)-binding sites have been described in villin, the functional characterization of these sites has not been done previously. In the present study we functionally dissect the calcium-dependent actin-capping and depolymerizing sites in villin. Our analysis reveals that villin binds Ca2+ with a Kd of 80.5 microM, a stoichiometry of 5.97, and a Hill's coefficient of 1.2. Using the NMR structure of villin 14T and the gelsolin-actin/Ca2+ crystal structure, six putative sites that result in Ca(2+)-induced conformational changes were identified in human villin and confirmed by mutational analysis. Molecular dynamics studies support the mutational analysis and provide a model for structural difference in the A93G mutant that prevents the calcium-induced conformational changes in the S1 domain of villin. Furthermore, we determined that villin expresses at least two types of Ca(2+)-sensitive sites that determine separate functional properties; site 1 (Glu-25, Asp-44, and Glu-74) regulates actin-capping, whereas sites 1 and 2 (Asp-86, Ala-93, and Asp-61), together with the intra-domain calcium-sensitive sites in villin, regulate actin depolymerization by villin. This is the first study that employs sequential mutagenesis to biochemically and functionally characterize the calcium-sensitive sites in villin. Such mutational analysis and functional characterization of the actin-capping and -depolymerizing sites are unknown for other proteins of the villin family. PMID- 15272028 TI - Dynamics of rat entorhinal cortex layer II and III cells: characteristics of membrane potential resonance at rest predict oscillation properties near threshold. AB - Neurones generate intrinsic subthreshold membrane potential oscillations (MPOs) under various physiological and behavioural conditions. These oscillations influence neural responses and coding properties on many levels. On the single cell level, MPOs modulate the temporal precision of action potentials; they also have a pronounced impact on large-scale cortical activity. Recent studies have described a close association between the MPOs of a given neurone and its electrical resonance properties. Using intracellular sharp microelectrode recordings we examine both dynamical characteristics in layers II and III of the entorhinal cortex (EC). Our data from EC layer II stellate cells show strong membrane potential resonances and oscillations, both in the range of 5-15 Hz. At the resonance maximum, the membrane impedance can be more than twice as large as the input resistance. In EC layer III cells, MPOs could not be elicited, and frequency-resolved impedances decay monotonically with increasing frequency or has only a small peak followed by a subsequent decay. To quantify and compare the resonance and oscillation properties, we use a simple mathematical model that includes stochastic components to capture channel noise. Based on this model we demonstrate that electrical resonance is closely related though not equivalent to the occurrence of sag-potentials and MPOs. MPO frequencies can be predicted from the membrane impedance curve for stellate cells. The model also explains the broad-band nature of the observed MPOs. This underscores the importance of intrinsic noise sources for subthreshold phenomena and rules out a deterministic description of MPOs. In addition, our results show that the two identified cell classes in the superficial EC layers, which are known to target different areas in the hippocampus, also have different preferred frequency ranges and dynamic characteristics. Intrinsic cell properties may thus play a major role for the frequency-dependent information flow in the hippocampal formation. PMID- 15272029 TI - Ca(2+) and K(+) (BK) channels in chick hair cells are clustered and colocalized with apical-basal and tonotopic gradients. AB - Electrical resonance is a mechanism used by birds and many vertebrates to discriminate between frequencies of sound, and occurs when the intrinsic oscillation in the membrane potential of a specific hair cell corresponds to a specific stimulus sound frequency. This intrinsic oscillation results from an interplay between an inward Ca(2+) current and the resultant activation of a hyperpolarizing Ca(2+)-activated K(+) current. These channels are predicted to lie in close proximity owing to the fast oscillation in membrane potential. The interplay of these channels is widespread in the nervous system, where they perform numerous roles including the control of synaptic release, burst frequency and circadian rhythm generation. Here, we used confocal microscopy to show that these two ion channels are clustered and colocalized in the chick hair cell membrane. The majority of Ca(2+) channels were colocalized while the proportion of colocalized BK channels was markedly less. In addition, we report both an apical-basal gradient of these clusters in individual hair cells, as well as a gradient in the number of clusters between hair cells along the tonotopic axis. These results give physical confirmation of previous predictions. Since the proportion of colocalized channels was a constant function of Ca(2+) channels, and not of BK channels, these results suggest that their colocalization is determined by the former. The molecular mechanisms underpinning their clustering and colocalization are likely to be common to other neuronal cells. PMID- 15272030 TI - Role of ATP-conductive anion channel in ATP release from neonatal rat cardiomyocytes in ischaemic or hypoxic conditions. AB - It is known that the level of ATP in the interstitial spaces within the heart during ischaemia or hypoxia is elevated due to its release from a number of cell types, including cardiomyocytes. However, the mechanism by which ATP is released from these myocytes is not known. In this study, we examined a possible involvement of the ATP-conductive maxi-anion channel in ATP release from neonatal rat cardiomyocytes in primary culture upon ischaemic, hypoxic or hypotonic stimulation. Using a luciferin-luciferase assay, it was found that ATP was released into the bulk solution when the cells were subjected to chemical ischaemia, hypoxia or hypotonic stress. The swelling-induced ATP release was inhibited by the carboxylate- and stilbene-derivative anion channel blockers, arachidonic acid and Gd3+, but not by glibenclamide. The local concentration of ATP released near the cell surface of a single cardiomyocyte, measured by a biosensor technique, was found to exceed the micromolar level. Patch-clamp studies showed that ischaemia, hypoxia or hypotonic stimulation induced the activation of single-channel events with a large unitary conductance (approximately 390 pS). The channel was selective to anions and showed significant permeability to ATP4- (PATP/PCl approximately 0.1) and MgATP2- (PATP/PCl approximately 0.16). The channel activity exhibited pharmacological properties essentially identical to those of ATP release. These results indicate that neonatal rat cardiomyocytes respond to ischaemia, hypoxia or hypotonic stimulation with ATP release via maxi-anion channels. PMID- 15272032 TI - Variations in excitability of single human motor axons, related to stochastic properties of nodal sodium channels. AB - Threshold variability of single human motor axons was studied by delivering 0.1 ms constant current stimuli of randomly varied intensity over the ulnar nerve at the elbow, and recording all-or-none potentials from flexor carpi ulnaris. In nine normal subjects, a single unit was tested with 8-11,000 stimuli at intervals of 0.5 s. After allowing for slow changes in excitability, the probability of excitation was in all cases well fitted by a cumulative Gaussian function. The relative spread (RS) of thresholds (S.D./mean) averaged 1.65 +/- 0.26% (mean +/- S.D., n = 9). When threshold was tested 20 ms after the start of a polarizing current, RS increased on depolarization and decreased on hyperpolarization. The product RS x mean threshold also increased on depolarization, especially when threshold was reduced by more than 50%. When the mean was reduced by 90%, RS increased above 50% and the axon sometimes fired spontaneously. Threshold variability was simulated by a computer model of a single node of Ranvier, in which the variability arose because of the stochastic behaviour of nodal sodium channels. The observed values of RS, and potential dependence of RS, were well modelled by a node with 60,000 sodium channels, of which about 1% were modelled as persistent sodium channels. Threshold variations in the model at resting potential were not primarily due to fluctuations in the state of the node before the stimulus was delivered, but rather to the variable activation of channels by the stimulus pulse. On depolarization, however, current through (mainly persistent) sodium channels caused appreciable fluctuations in membrane potential, which increased RS and the probability of spontaneous firing. PMID- 15272031 TI - Non-selective cationic channels of smooth muscle and the mammalian homologues of Drosophila TRP. AB - Throughout the body there are smooth muscle cells controlling a myriad of tubes and reservoirs. The cells show enormous diversity and complexity compounded by a plasticity that is critical in physiology and disease. Over the past quarter of a century we have seen that smooth muscle cells contain--as part of a gamut of ion handling mechanisms--a family of cationic channels with significant permeability to calcium, potassium and sodium. Several of these channels are sensors of calcium store depletion, G-protein-coupled receptor activation, membrane stretch, intracellular Ca2+, pH, phospholipid signals and other factors. Progress in understanding the channels has, however, been hampered by a paucity of specific pharmacological agents and difficulty in identifying the underlying genes. In this review we summarize current knowledge of these smooth muscle cationic channels and evaluate the hypothesis that the underlying genes are homologues of Drosophila TRP (transient receptor potential). Direct evidence exists for roles of TRPC1, TRPC4/5, TRPC6, TRPV2, TRPP1 and TRPP2, and more are likely to be added soon. Some of these TRP proteins respond to a multiplicity of activation signals- promiscuity of gating that could enable a variety of context-dependent functions. We would seem to be witnessing the first phase of the molecular delineation of these cationic channels, something that should prove a leap forward for strategies aimed at developing new selective pharmacological agents and understanding the activation mechanisms and functions of these channels in physiological systems. PMID- 15272033 TI - Functional and molecular evidence of MaxiK channel beta1 subunit decrease with coronary artery ageing in the rat. AB - Large-conductance, voltage- and Ca2+ -activated K+ channels (MaxiK, BK) are key regulators of vascular tone. Vascular MaxiK are formed by the pore-forming alpha subunit and the modulatory beta1 subunit, which imprints unique kinetics, Ca2+/voltage sensitivities and pharmacology to the channel. As age progresses, alpha subunit functional expression and protein levels diminish in coronary myocytes. However, whether ageing modifies beta1 subunit expression or the mechanism of alpha subunit reduction is unknown. Thus, we examined functional and pharmacological characteristics of MaxiK, as well as alpha and beta1 transcript levels in coronary myocytes from young and old F344 rats. The mechanism of age dependent alpha subunit protein reduction involves its transcript downregulation. A corresponding loss of beta1 transcripts was also detected in old myocytes, suggesting a proportional age-dependent decrease of beta1 to alpha subunit protein. Indeed, MaxiK channel properties, defined by coassembly of beta1 and alpha subunits, were equivalent in young versus old, for example in terms of (i) activation kinetics, (ii) sensitivity to Ca2+ levels > 1 microm (iii) dehydrosoyasaponin-I-induced activation, and (iv) iberiotoxin blockade. Consistent with less MaxiK expression/function in older myocytes, the ability of iberiotoxin to contract coronary rings was reduced approximately 50% with ageing confirming our previous findings. 5-Hydroxytryptamine (5-HT) contractile efficacy was reduced by iberiotoxin pretreatment in young > old coronary arteries (explained by larger iberiotoxin-induced contraction and decreased dynamic range for 5-HT contraction in young versus old) with no apparent differences in nitroglycerine-induced relaxation. We propose that the age-related MaxiK reduction involves a parallel decrease of alpha and beta1 functional expression via a transcript downregulatory mechanism; a major impact on basal and possibly stimulated coronary contraction may contribute to altered coronary flow regulation and coronary morbidity in the elderly. PMID- 15272034 TI - Regulation of tissue oxygen levels in the mammalian lens. AB - Opacification of the lens nucleus is a major cause of blindness and is thought to result from oxidation of key cellular components. Thus, long-term preservation of lens clarity may depend on the maintenance of hypoxia in the lens nucleus. We mapped the distribution of dissolved oxygen within isolated bovine lenses and also measured the rate of oxygen consumption (QO2) by lenses, or parts thereof. To assess the contribution of mitochondrial metabolism to the lens oxygen budget, we tested the effect of mitochondrial inhibitors on (QO2) and partial pressure of oxygen (PO2). The distribution of mitochondria was mapped in living lenses by 2 photon microscopy. We found that a steep gradient of PO2 was maintained within the tissue, leading to PO2 < 2 mmHg in the core. Mitochondrial respiration accounted for approximately 90% of the oxygen consumed by the lens; however, PO2 gradients extended beyond the boundaries of the mitochondria-containing cell layer, indicating the presence of non-mitochondrial oxygen consumers. Time constants for oxygen consumption in various regions of the lens and an effective oxygen diffusion coefficient were calculated from a diffusion-consumption model. Typical values were 3 x 10(-5) cm(2) s(-1) for the effective diffusion coefficient and a 5 min time constant for oxygen consumption. Surprisingly, the calculated time constants did not differ between differentiating fibres (DF) that contained mitochondria and mature fibres (MF) that did not. Based on these parameters, DF cells were responsible for approximately 88% of lens oxygen consumption. A modest reduction in tissue temperature resulted in a marked decrease in (QO2) and the subsequent flooding of the lens core with oxygen. This phenomenon may be of clinical relevance because cold, oxygen-rich solutions are often infused into the eye during intraocular surgery. Such procedures are associated with a strikingly high incidence of postsurgical nuclear cataract. PMID- 15272035 TI - Role of adenosine transport in gestational diabetes-induced L-arginine transport and nitric oxide synthesis in human umbilical vein endothelium. AB - Gestational diabetes is associated with increased L-arginine transport and nitric oxide (NO) synthesis, and reduced adenosine transport in human umbilical vein endothelial cells (HUVEC). Adenosine increases endothelial L-arginine/NO pathway via A(2) purinoceptors in HUVEC from normal pregnancies. It is unknown whether the effect of gestational diabetes is associated with activation of these purinoceptors or altered expression of human cationic amino acid transporter 1 (hCAT-1) or human equilibrative nucleoside transporter 1 (hENT1), or endothelial NO synthase (eNOS) in HUVEC. Cells were isolated from normal or gestational diabetic pregnancies and cultured up to passage 2. Gestational diabetes increased hCAT-1 mRNA expression (2.4-fold) and activity, eNOS mRNA (2.3-fold), protein level (2.1-fold), and phosphorylation (3.8-fold), but reduced hENT1 mRNA expression (32%) and activity. Gestational diabetes increased extracellular adenosine (2.7 microM), and intracellular L-arginine (1.9 mM) and L-citrulline (0.7 mM) levels compared with normal cells (0.05 microM, 0.89 mM, 0.35 mM, respectively). Incubation of HUVEC from normal pregnancies with 1 microM nitrobenzylthioinosine (NBMPR) mimicked the effect of gestational diabetes, but NBMPR was ineffective in diabetic cells. Gestational diabetes and NBMPR effects involved eNOS, PKC and p42/44(mapk) activation, and were blocked by the A(2a) purinoceptor antagonist ZM-241385. Thus, gestational diabetes increases the L arginine/NO pathway involving activation of mitogen-activated protein (MAP) kinases, protein kinase C (PKC) and NO cell signalling cascades following activation of A(2a) purinoceptors by extracellular adenosine. A functional relationship is proposed between adenosine transport and modulation of L-arginine transport and NO synthesis in HUVEC, which could be determinant in regulating vascular reactivity in diabetes mellitus. PMID- 15272036 TI - Principal neuron spiking: neither necessary nor sufficient for cerebral blood flow in rat cerebellum. AB - Neuronal activity, cerebral blood flow, and metabolic responses are all strongly coupled, although the mechanisms behind the coupling remain unclear. One of the key questions is whether or not increases in spiking activity in the stimulated neurons are sufficient to drive the activity-dependent rises in cerebral blood flow (CBF) that form the basis of the signals used in functional neuroimaging such as the blood oxygen level-dependent (BOLD) signal. To this end the present study examined the effect of enhanced spike activity per se on CBF in rat cerebellar cortex under conditions of disinhibition, achieved by blocking GABA(A) receptors using either bicuculline or picrotoxin. Purkinje cell spiking activity and local field potentials were recorded by glass microelectrodes, and laser Doppler flowmetry was used to monitor CBF. Disinhibition increased Purkinje cell spiking rate to 200-300% of control without incurring any increase in basal CBF. This demonstrates that increased spike activity per se is not sufficient to affect basal CBF. The neurovascular coupling between excitatory synaptic activity and CBF responses evoked by inferior olive (climbing fibre) stimulation was preserved during disinhibition. Thus, the unchanged basal CBF in the presence of the dramatic rise in Purkinje cell spiking rate was not explained by impaired synaptic activity-CBF coupling. On the basis of our previous and the present studies, we conclude that increased spiking activity of principal neurons is neither sufficient nor necessary to elicit CBF responses and in turn BOLD signals, and that activation-dependent vascular signals reflect excitatory synaptic activity. PMID- 15272037 TI - Effects of cortical stimulation on auditory-responsive thalamic neurones in anaesthetized guinea pigs. AB - In the present study, we investigated neuronal responses to acoustic stimuli and cortical stimulation in the medial geniculate body (MGB) through in vivo intracellular recordings in anaesthetized guinea pigs. Of the 54 neurones examined with acoustic stimuli, 36 showed excitatory postsynaptic potential (EPSP) responses and 19 showed inhibitory postsynaptic potential (IPSP) responses to acoustic stimuli. Of the 36 EPSP neurones examined with corticofugal modulation, 29 received corticofugal depolarization, 3 corticofugal inhibition, and 4 showed no effect. Of the 19 IPSP neurones, 17 received corticofugal inhibition and 2 were not affected. The mean amplitude of the EPSPs evoked by acoustic stimuli was similar to that evoked by the electrical cortical stimulation (9.19 +/- 5.55 mV versus 9.22 +/- 5.16 mV). There was a significant correlation between the parameters of the EPSPs evoked by an acoustic stimulus and those evoked by cortical stimulation. The mean amplitude of the IPSP evoked by electrical cortical stimulation was significantly greater than that evoked by acoustic stimuli (11.6 +/- 3.8 mV versus 9.1 +/- 3.7 ms, P < 0.05). Seven auditory EPSP and 7 IPSP neurones were examined with corticofugal modulation and labelled with Neurobiotin. Of the 7 EPSP neurones, 5 showed excitatory responses to cortical stimulation and 2 demonstrated no effects. Four of the 5 neurones that received corticofugal depolarization were located in the lemniscal MGB and 1 in the non-lemniscal MGB; of the remaining 2, 1 was located in the lemniscal and the other in the non-lemniscal MGB. Of the 7 IPSP neurones, 1 received an excitatory corticofugal input followed by an inhibitory input and 4 received only an inhibitory corticofugal input, while the remainder demonstrated no corticofugal effects. All 7 neurones were located in the non-lemniscal MGB. The result that both ascending and descending inputs caused similarly shaped EPSPs reflects a neuronal endogenous characteristic irrespective of the physical locations of the synapses. The IPSP responses to both acoustic stimuli and electrical cortical stimulation are likely to be caused by feedback from the thalamic reticular nucleus. PMID- 15272038 TI - In vivo intracellular responses of the medial geniculate neurones to acoustic stimuli in anaesthetized guinea pigs. AB - In the present study, we investigated the auditory response features of the medial geniculate neurones, using in vivo intracellular recordings in anaesthetized guinea pigs. Of the 76 neurones examined, 9 showed 'off' or 'on off' responses to an acoustic stimulus and thus were defined as 'off' or 'on-off' neurones. Among the remaining 67 neurones, 42 showed an excitatory postsynaptic potential (EPSP) to acoustic stimuli and 25 showed either a pure inhibitory postsynaptic potential (IPSP, 7 neurones), or an IPSP preceded by an EPSP (EPSP IPSP type, 18 neurones). The EPSP responses exhibited a mean latency of 15.7 +/- 6.1 ms, which was significantly shorter than that of the IPSP responses (21.3 +/- 8.6 ms, P < 0.01). The IPSP responses also showed a significantly greater duration than the EPSP responses (208.5 +/- 128.2 ms versus 122.4 +/- 84.8 ms, P < 0.05), while there were no significant differences between the amplitudes of IPSP and EPSP (8.3 +/- 3.2 mV versus 8.7 +/- 5.3 mV). Of the 11 neurones that showed EPSP responses to acoustic stimuli and were histologically labelled, 7 were located in the lemniscal medial geniculate body (MGB) and 4 in the non lemniscal MGB. Another 6 labelled neurones that showed IPSP responses to acoustic stimuli were located in the non-lemniscal MGB. With a membrane potential of above -72 mV, the neurones showed greater EPSP or IPSP to an acoustic stimulus when their membrane potential was depolarized. However, upon hyperpolarization to below -74 mV, the neurones shifted to low-threshold calcium spikes (LTS)/LTS bursts. In response to auditory stimuli of different durations, 'off' neurones that responded to the offset of the acoustic stimulus and were located in the non lemniscal MGB showed different response latencies or deviations of latencies in addition to exhibiting different numbers of spikes, suggesting that the timing of the spikes could be another component utilized by thalamic neurones to encode information on the stimulus. Given that some non-lemniscal neurones are multisensory and project to the entire auditory cortex, the selective corticofugal inhibition in the non-lemniscal MGB would enable the ascending pathway to prepare the auditory cortex to receive subsequent auditory information, avoiding the interference of other sensory inputs. PMID- 15272039 TI - Magnesium-inhibited, TRPM6/7-like channel in cardiac myocytes: permeation of divalent cations and pH-mediated regulation. AB - Cardiac tissue expresses several TRP proteins as well as a Mg2+ -inhibited, non selective cation current (IMIC) that bears many characteristics of TRP channel currents. We used the whole-cell voltage clamp technique in pig and rat ventricular myocytes to characterize the permeation, blockage properties and regulation of the cardiac IMIC channels in order to compare them with TRP channels, in particular with Mg2+ -sensitive TRPM6 and TRPM7. We show that removing extracellular divalent cations unmasks large inward and outward monovalent currents, which can be inhibited by intracellular Mg2+. Inward currents are suppressed upon replacing extracellular Na+ by NMDG+. Divalent cations block monovalent IMIC and, at 10-20 mm, carry measurable currents. Their efficacy sequence in decreasing outward IMIC (Ni2+ = Mg2+ > Ca2+ > Ba2+) and in inducing inward IMIC (Ni2+ >> Mg2+ = Ca2+ approximately Ba2+), and their permeabilities calculated from reversal potentials are similar to those of TRPM6 and TRPM7 channels. The trivalent cations Gd3+ and Dy3+ also block IMIC in a voltage-dependent manner (delta = 0.4-0.5). In addition they inhibit the inward current carried by divalent cations. IMIC is regulated by pH. Decreasing or increasing extracellular pH decreased and increased IMIC, respectively (pH0.5 = 6.9, nH = 0.98). Qualitatively similar results were obtained on IMIC in rat basophilic leukaemia cells. These effects in cardiac myocytes were absent in the presence of high intracellular buffering by 40 mm Hepes. Our results suggest that IMIC in cardiac cells is due to TRPM channels, most probably to TRPM6 or TRPM7 channels or to their heteromultimeres. PMID- 15272040 TI - An estimate of the resting membrane resistance of frog olfactory receptor neurones. AB - The ability of a frog olfactory receptor neurone (ORN) to respond to odorous molecules depends on its resting membrane properties, including membrane resistance and potential. Quantification of these properties is difficult because of a shunt conductance at the membrane-pipette seal that is in parallel with the true membrane conductance. In physiological salines, the sum of these two conductances averaged 235 pS. We used ionic substitution and channel blockers to reduce the membrane conductance as much as possible. This yielded a lower limit for the membrane conductance of 158 pS. The upper limit of resting membrane resistance, then, is 6 GOmega. The membrane is permeable to K+ and, to a lesser extent, other cations. No resting Cl- conductance was detectable. Correcting measured zero-current potentials for distortion by the shunt suggests that the resting membrane potential is no more negative than -75 mV. The present results help to explain why frog ORNs are excitable at rest. PMID- 15272041 TI - Cannabinoids suppress synaptic input to neurones of the rat dorsal motor nucleus of the vagus nerve. AB - Cannabinoids bind central type 1 receptors (CB1R) and modify autonomic functions, including feeding and anti-emetic behaviours, when administered peripherally or into the dorsal vagal complex. Western blots and immunohistochemistry indicated the expression of CB1R in the rat dorsal vagal complex, and tissue polymerase chain reaction confirmed that CB1R message was made within the region. To identify a cellular substrate for the central autonomic effects of cannabinoids, whole-cell patch-clamp recordings were made in brainstem slices to determine the effects of CB1R activation on synaptic transmission to neurones of the dorsal motor nucleus of the vagus (DMV). A subset of these neurones was identified as gastric related after being labelled retrogradely from the stomach. The CB1R agonists WIN55,212-2 and anandamide decreased the frequency of spontaneous excitatory or inhibitory postsynaptic currents in a concentration-related fashion, an effect that persisted in the presence of tetrodotoxin. Paired pulse ratios of electrically evoked postsynaptic currents were also increased by WIN55,212-2. The effects of WIN55,212-2 were sensitive to the selective CB1R antagonist AM251. Cannabinoid agonist effects on synaptic input originating from neurones in the nucleus tractus solitarius (NTS) were determined by evoking activity in the NTS with local glutamate application. Excitatory and inhibitory synaptic inputs arising from the NTS were attenuated by WIN55,212-2. Our results indicate that cannabinoids inhibit transfer of synaptic information to the DMV, including that arising from the NTS, in part by acting at receptors located on presynaptic terminals contacting DMV neurones. Inhibition of synaptic input to DMV neurones is likely to contribute to the suppression of visceral motor responses by cannabinoids. PMID- 15272042 TI - Persistent decrease in synaptic efficacy induced by nicotine at Schaffer collateral-CA1 synapses in the immature rat hippocampus. AB - Neuronal nicotinic acetylcholine receptors (nAChRs) are widely distributed within the brain where they contribute to the regulation of higher cognitive functions. The loss of the cholinergic function in Alzheimer's disease patients, along with the well-known memory enhancing effect of nicotine, emphasizes the role of cholinergic signalling in memory functions. The hippocampus, a key structure in learning and memory, is endowed with nAChRs localized at pre- and postsynaptic levels. In previous work on the immature hippocampus we have shown that, at low probability (P) synapses, activation of alpha7 nAChRs by nicotine or by endogenously released acetylcholine persistently enhanced glutamate release and converted 'presynaptically silent' synapses into functional ones. Here we show that in the same preparation, at high P synapses, nicotine induces long-term depression of AMPA- and NMDA-mediated synaptic currents. This effect was mediated by presynaptic alpha7- and beta2-containing receptors and was associated with an increase in the paired pulse ratio and in the coefficient of variation. High P synapses could be converted into low P and vice versa by changing the extracellular Ca2+/Mg2+ ratio. In these conditions nicotine was able to persistently potentiate or depress synaptic responses depending on the initial P values. A bi-directional control of synaptic plasticity by nicotine would considerably enhance the computational properties of the network during a critical period of postnatal development thus contributing to sculpt the neuronal circuit. PMID- 15272043 TI - Two types of non-selective cation channel opened by muscarinic stimulation with carbachol in bovine ciliary muscle cells. AB - In the ciliary muscle, the tonic contraction requires a sustained influx of Ca2+ through the cell membrane. However, little has hitherto been known about the route(s) of Ca2+ influx in this tissue that lacks voltage-gated Ca2+ channels. To identify ion channels as the Ca2+ entry pathway we studied the effects of carbachol (CCh) on freshly isolated bovine ciliary muscle cells by whole-cell voltage clamp. Experiments were carried out using pipettes filled with K+ -free solution containing 100 mm caesium aspartate, 5 mm BAPTA and 180 microm GTP (pH 7.0; the intracellular free Ca2+ concentration, [Ca2+]i = 70 nm). CCh evoked an inward current showing polarity reversal at a holding potential near 0 mV. Analysis of the current noise distinguished two types of non-selective cation channel (NSCCL and NSCCS) with widely different unitary conductances (35 pS and 100 fS). The ratios of the permeabilities to Li+, Na+, Cs+, Mg2+, Ca2+, Sr2+ and Ba2+, estimated by cation replacement procedures, were 0.9: 1.0: 1.5: 0.2: 0.3: 0.4: 0.5 for NSCCL, and 1.0: 1.0: 1.8: 2.5: 2.6: 3.2: 5.0 for NSCCS. NSCCS, but not NSCCL, was strongly inhibited by elevation of [Ca2+]i. Both NSCCL and NSCCS were dose-dependently inhibited by 1-100 microm SKF96365, La3+ and Gd3+, which also inhibited the tonic component of the contraction produced in muscle bundles by CCh without markedly affecting the initial phasic component. NSCCL and/or NSCCS may serve as a major Ca2+ entry pathway required for sustained contraction of the bovine ciliary muscle. RT-PCR experiments in the bovine ciliary muscle (whole tissue) detected mRNAs of several transient receptor potential (TRP) channel homologues (TRPC1, TRPC3, TRPC4 and TRPC6), which are now regarded as possible molecular candidates for receptor-operated cation channels. PMID- 15272044 TI - Neurotransmitter modulation of extracellular H+ fluxes from isolated retinal horizontal cells of the skate. AB - Self-referencing H(+)-selective microelectrodes were used to measure extracellular H(+) fluxes from horizontal cells isolated from the skate retina. A standing H(+) flux was detected from quiescent cells, indicating a higher concentration of free hydrogen ions near the extracellular surface of the cell as compared to the surrounding solution. The standing H(+) flux was reduced by removal of extracellular sodium or application of 5-(N-ethyl-N-isopropyl) amiloride (EIPA), suggesting activity of a Na(+)-H(+) exchanger. Glutamate decreased H(+) flux, lowering the concentration of free hydrogen ions around the cell. AMPA/kainate receptor agonists mimicked the response, and the AMPA/kainate receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) eliminated the effects of glutamate and kainate. Metabotropic glutamate agonists were without effect. Glutamate-induced alterations in H(+) flux required extracellular calcium, and were abolished when cells were bathed in an alkaline Ringer solution. Increasing intracellular calcium by photolysis of the caged calcium compound NP-EGTA also altered extracellular H(+) flux. Immunocytochemical localization of the plasmalemma Ca(2+)-H(+)-ATPase (PMCA pump) revealed intense labelling within the outer plexiform layer and on isolated horizontal cells. Our results suggest that glutamate modulation of H(+) flux arises from calcium entry into cells with subsequent activation of the plasmalemma Ca(2+)-H(+)-ATPase. These neurotransmitter-induced changes in extracellular pH have the potential to play a modulatory role in synaptic processing in the outer retina. However, our findings argue against the hypothesis that hydrogen ions released by horizontal cells normally act as the inhibitory feedback neurotransmitter onto photoreceptor synaptic terminals to create the surround portion of the centre-surround receptive fields of retinal neurones. PMID- 15272045 TI - State-dependent trapping of flecainide in the cardiac sodium channel. AB - Flecainide is a Class I antiarrhythmic drug and a potent inhibitor of the cardiac (Nav1.5) sodium channel. Although the flecainide inhibition of Nav1.5 is typically enhanced by depolarization, the contributions of the open and inactivated states to flecainide binding and inhibition remain controversial. We further investigated the state-dependent binding of flecainide by examining its inhibition of rapidly inactivating and non-inactivating mutants of Nav1.5 expressed in Xenopus oocytes. Applying flecainide while briefly depolarizing from a relatively negative holding potential resulted in a low-affinity inhibition of the channel (IC(50) = 345 microM). Increasing the frequency of stimulation potentiated the flecainide inhibition (IC(50) = 7.4 microM), which progressively increased over the range of voltages where Nav1.5 channels activated. This contrasts with sustained depolarizations that effectively stabilize the channels in inactivated states, which failed to promote significant flecainide inhibition. The voltage sensitivity and strong dependence of the flecainide inhibition on repetitive depolarization suggests that flecainide binding is facilitated by channel opening and that the drug does not directly bind to closed or inactivated channels. The binding of flecainide to open channels was further investigated in a non-inactivating mutant of Nav1.5. Flecainide produced a time-dependent decay in the current of the non-inactivating mutant that displayed kinetics consistent with a simple pore blocking mechanism (K(D) = 11 microM). At hyperpolarized voltages, flecainide slowed the recovery of both the rapidly inactivating (tau = 81 +/- 3 s) and non-inactivating (tau = 42 +/- 3 s) channels. Mutation of a conserved isoleucine of the D4S6 segment (I1756C) creates an alternative pathway that permits the rapid diffusion of anaesthetics out of the Nav1.5 channel. The I1756C mutation accelerated the recovery of both the rapidly inactivating (tau = 12.6 +/- 0.4 s) and non-inactivating (tau = 7.4 +/- 0.1 s) channels, suggesting that flecainide is trapped and not tightly bound within the pore when the channels are closed or inactivated. The data indicate that flecainide rapidly gains access to its binding site when the channel is open and inhibits Na(+) current by a pore blocking mechanism. Closing of either the activation or the inactivation gate traps flecainide within the pore resulting in the slow recovery of the drug-modified channels at hyperpolarized voltages. PMID- 15272046 TI - Ghrelin stimulates neurogenesis in the dorsal motor nucleus of the vagus. AB - Ghrelin, a gastric peptide hormone, has been reported to regulate growth hormone secretion and energy homeostasis. Here we show that ghrelin promotes neural proliferation in vivo and in vitro in the rat dorsal motor nucleus of the vagus (DMNV). Ghrelin receptor mRNA and immunoreactivity were detected in tissues from DMNV. Systemic administration of ghrelin (130 nmol kg(-1)) significantly increased 5-bromo-2'-deoxyuridine (BrdU) incorporation in the DMNV in adult rats with cervical vagotomy (BrdU positive cells; from 27 +/- 4 to 69 +/- 14 n = 5, P < 0.05). In vitro, exposure of cultured DMNV neurones to ghrelin significantly increased the percentage of BrdU incorporation into cells in both dose-dependent (10(-9) -10(-6)m), and time-dependent (6 h to 48 h) manners. Ghrelin significantly increased voltage-activated calcium currents in isolated single DMNV neurones from a mean maximal change of 141 +/- 26 pA to 227 +/- 37 pA. Upon removal of ghrelin, calcium currents slowly returned to baseline. Blocking L-type calcium channels by diltiazem (10 microm) significantly attenuated ghrelin mediated increments in BrdU incorporation (n = 5, P < 0.05). Ghrelin acts directly on DMNV neurones to stimulate neurogenesis. PMID- 15272047 TI - Functional properties of K+ currents in adult mouse ventricular myocytes. AB - Although the K+ currents expressed in hearts of adult mice have been studied extensively, detailed information concerning their relative sizes and biophysical properties in ventricle and atrium is lacking. Here we describe and validate pharmacological and biophysical methods that can be used to isolate the three main time- and voltage-dependent outward K+ currents which modulate action potential repolarization. A Ca2+ -independent transient outward K+ current, Ito, can be separated from total outward current using an 'inactivating prepulse'. The rapidly activating, slowly inactivating delayed rectifier K+ current, IKur, can be isolated using submillimolar concentrations of 4-aminopyridine (4-AP). The remaining K+ current, Iss, can be obtained by combining these two procedures: (i) inactivating Ito and (ii) eliminating IKur by application of low concentration of 4-AP. Iss activates relatively slowly and shows very little inactivation, even during depolarizations lasting several seconds. Our findings also show that the rate of reactivation of Ito is more than 20-fold faster than that of IKur. These results demonstrate that the outward K+ currents in mouse ventricles can be separated based on their distinct time and voltage dependence, and different sensitivities to 4-AP. Data obtained at both 22 and 32 degrees C demonstrate that although the duration of the inactivating prepulse has to be adapted for the recording temperature, this approach for separation of K+ current components is also valid at more physiological temperatures. To demonstrate that these methods also allow separation of these K+ currents in other cell types, we have applied this same approach to myocytes from mouse atria. Molecular approaches have been used to compare the expression levels of different K+ channels in mouse atrium and ventricle. These findings provide new insights into the functional roles of IKur, Ito and Iss during action potential repolarization. PMID- 15272048 TI - Nicotine increases initial blood flow responses to local heating of human non glabrous skin. AB - Nicotine affects the regulation of skin blood flow (SkBF), but the mechanisms involved are not well understood. We tested the hypothesis that acute exposure to nicotine inhibits both the initial neurally mediated component and the later sustained component of SkBF responses to local heating of non-glabrous skin in humans. SkBF (measured by laser-Doppler) responses to local heating of forearm skin from 32 to 42 degrees C were measured in 11 chronic smokers. Heating occurred at one site over 15 min (RAMP) and over 90 s (STEP) at another site, and was maintained for an additional 30 min. STEP heating was also applied to a site pretreated with bretylium via iontophoresis to inhibit noradrenergic neurotransmission. Responses were measured before and after acute administration of nicotine via cigarettes or nasal spray in two experimental sessions. Nicotine decreased resting skin blood flow (P < 0.05); this response was inhibited by bretylium. During RAMP, nicotine increased the initial SkBF at 42 degrees C (by approximately 12%, P < 0.05). For STEP, nicotine increased the initial peak response (by approximately 25%, P < 0.05), and decreased the sustained plateau value (by approximately 10%, P < 0.05). In skin pretreated with bretylium, the increase caused by nicotine in the initial peak value persisted, but the plateau value was not different from pre-nicotine. These data suggest that in abstinent cigarette smokers, nicotine augments initial responses to both gradual and rapid non-painful heating of non-glabrous skin by sensitizing the sensory nerves that mediate the axon reflex associated with rapid vasodilatation. In contrast, nicotine decreases SkBF responses to prolonged heating by activating noradrenergic nerves. PMID- 15272049 TI - Demonstration of a second rapidly conducting cortico-diaphragmatic pathway in humans. AB - Functional imaging studies in normal humans have shown that the supplementary motor area (SMA) and the primary motor cortex (PMC) are coactivated during various breathing tasks. It is not known whether a direct pathway from the SMA to the diaphragm exists, and if so what properties it has. Using transcranial magnetic stimulation (TMS) a site at the vertex, representing the diaphragm primary motor cortex, has been identified. TMS mapping revealed a second area 3 cm anterior to the vertex overlying the SMA, which had a rapidly conducting pathway to the diaphragm (mean latency 16.7 +/- 2.4 ms). In comparison to the vertex, the anterior position was characterized by a higher diaphragm motor threshold, a greater proportional increase in motor-evoked potential (MEP) amplitude with voluntary facilitation and a shorter silent period. Stimulus response curves did not differ significantly between the vertex and anterior positions. Using paired TMS, we also compared intracortical inhibition/facilitation (ICI/ICF) curves. In comparison to the vertex, the MEP elicited from the anterior position was not inhibited at short interstimulus intervals (1-5 ms) and was more facilitated at long interstimulus intervals (9-20 ms). The patterns of response were identical for the costal and crural diaphragms. We conclude that the two coil positions represent discrete areas that are likely to be the PMC and SMA, with the latter wielding a more excitatory effect on the diaphragm. PMID- 15272050 TI - Atelocollagen-mediated synthetic small interfering RNA delivery for effective gene silencing in vitro and in vivo. AB - Silencing gene expression by siRNAs is rapidly becoming a powerful tool for the genetic analysis of mammalian cells. However, the rapid degradation of siRNA and the limited duration of its action call for an efficient delivery technology. Accordingly, we describe here that Atelocollagen complexed with siRNA is resistant to nucleases and is efficiently transduced into cells, thereby allowing long-term gene silencing. Site-specific in vivo administration of an anti luciferase siRNA/Atelocollagen complex reduced luciferase expression in a xenografted tumor. Furthermore, Atelocollagen-mediated transfer of siRNA in vivo showed efficient inhibition of tumor growth in an orthotopic xenograft model of a human non-seminomatous germ cell tumor. Thus, for clinical applications of siRNA, an Atelocollagen-based non-viral delivery method could be a reliable approach to achieve maximal function of siRNA in vivo. PMID- 15272051 TI - Transcriptional response to corticotropin-releasing factor in AtT-20 cells. AB - Corticotropin-releasing factor (CRF) plays a central role in the regulation of the hypothalamic-pituitary-adrenal axis, mediating endocrine and behavioral responses to various stressors. Two high-affinity receptors for CRF have been described. Although many of the intracellular signaling pathways activated by CRF have been studied extensively, our knowledge of transcriptional responses downstream of the CRF receptor 1 (CRFR1) is still limited. To elucidate gene networks regulated by CRF and CRFR1, we applied microarray technology to explore transcriptional response to CRF stimulation. Therefore, mouse pituitary-derived AtT-20 cells were exposed continuously to CRF either in the presence or absence of the specific CRFR1 antagonist R121919. Transcriptional responses to different treatments were studied in a time course ranging from 0.5 to 24 h. Microarray data were analyzed using classic microarray data analysis tools such as correspondence factor analysis, cluster analysis, and fold-change filtering. Furthermore, spectral map analysis was applied, a recently introduced unsupervised multivariate analysis method. A broad and transient transcriptional response to CRF was identified that could be blocked by the antagonist. This way, several known CRF-induced target genes and novel CRF responsive genes were identified. These include transcription factors such as cAMP-responsive element modulator (7x increased), secreted peptides such as cholecystokinin (1.5x), and proteins involved in modulating intracellular signaling, such as regulator of G protein signaling 2 (11x). Up-regulation of many of these genes can be explained as negative feedback, attenuating CRF-activated pathways. In addition, spectral map analysis proved to be a promising new tool for microarray data analysis. PMID- 15272052 TI - RNA interference suggests a primary role for monoacylglycerol lipase in the degradation of the endocannabinoid 2-arachidonoylglycerol. AB - The endogenous cannabinoid 2-arachidonoylglycerol (2-AG) is produced by neurons and other cells in a stimulus-dependent manner and undergoes rapid biological inactivation through transport into cells and catalytic hydrolysis. The enzymatic pathways responsible for 2-AG degradation are only partially understood. We have shown previously that overexpression of monoacylglycerol lipase (MGL), a cytosolic serine hydrolase that cleaves 1- and 2-monoacylglycerols to fatty acid and glycerol, reduces stimulus-dependent 2-AG accumulation in primary cultures of rat brain neurons. We report here that RNA interference-mediated silencing of MGL expression greatly enhances 2-AG accumulation in HeLa cells. After stimulation with the calcium ionophore ionomycin, 2-AG levels in MGL-silenced cells were comparable with those found in cells in which 2-AG degradation had been blocked using methyl arachidonyl fluorophosphonate, a nonselective inhibitor of 2-AG hydrolysis. The results indicate that MGL plays an important role in the degradation of endogenous 2-AG in intact HeLa cells. Furthermore, immunodepletion experiments show that MGL accounts for at least 50% of the total 2-AG-hydrolyzing activity in soluble fractions of rat brain, suggesting that this enzyme also contributes to 2-AG deactivation in the central nervous system. PMID- 15272053 TI - Meclizine is an agonist ligand for mouse constitutive androstane receptor (CAR) and an inverse agonist for human CAR. AB - The constitutive androstane receptor (CAR, NR1I3) is a key regulator of xenobiotic and endobiotic metabolism. The ligand-binding domains of murine (m) and human (h) CAR are divergent relative to other nuclear hormone receptors, resulting in species-specific differences in xenobiotic responses. Here we identify the widely used antiemetic meclizine (Antivert; Bonine) as both an agonist ligand for mCAR and an inverse agonist for hCAR. Meclizine increases mCAR transactivation in a dose-dependent manner. Like the mCAR agonist 1,4-bis[2-(3,5 dichloropyridyloxy)]benzene, meclizine stimulates binding of steroid receptor coactivator 1 to the murine receptor in vitro. Meclizine administration to mice increases expression of CAR target genes in a CAR-dependent manner. In contrast, meclizine suppresses hCAR transactivation and inhibits the phenobarbital-induced expression of the CAR target genes, cytochrome p450 monooxygenase (CYP)2B10, CYP3A11, and CYP1A2, in primary hepatocytes derived from mice expressing hCAR, but not mCAR. The inhibitory effect of meclizine also suppresses acetaminophen induced liver toxicity in humanized CAR mice. These results demonstrate that a single compound can induce opposite xenobiotic responses via orthologous receptors in rodents and humans. PMID- 15272054 TI - The vitamin D receptor is present in caveolae-enriched plasma membranes and binds 1 alpha,25(OH)2-vitamin D3 in vivo and in vitro. AB - The steroid hormone 1 alpha,25(OH)(2)-vitamin D(3) (1,25D) regulates gene transcription through a nuclear receptor [vitamin D receptor (VDR)] and initiation of rapid cellular responses through a putative plasma membrane associated receptor (VDR(mem)). This study characterized the VDR(mem) present in a caveolae-enriched membrane fraction (CMF), a site of accumulation of signal transduction agents. Saturable and specific [(3)H]-1,25D binding in vitro was found in CMF of chick, rat, and mouse intestine; mouse lung and kidney; and human NB4 leukemia and rat ROS 17/2.8 osteoblast-like cells; in all cases the 1,25D K(D) binding dissociation constant = 1-3 nM. Our data collectively support the classical VDR being the VDR(mem) in caveolae: 1) VDR antibody immunoreactivity was detected in CMF of all tissues tested; 2) competitive binding of [(3)H]-1,25D by eight analogs of 1,25D was significantly correlated between nuclei and CMF (r(2) = 0.95) but not between vitamin D binding protein (has a different ligand binding specificity) and CMF; 3) confocal immunofluorescence microscopy of ROS 17/2.8 cells showed VDR in close association with the caveolae marker protein, caveolin-1, in the plasma membrane region; 4) in vivo 1,25D pretreatment reduced in vitro [(3)H]-1,25D binding by 30% in chick and rat intestinal CMF demonstrating in vivo occupancy of the CMF receptor by 1,25D; and 5) comparison of [(3)H]-1,25D binding in VDR KO and WT mouse kidney tissue showed 85% reduction in VDR KO CMF and 95% reduction in VDR KO nuclear fraction. This study supports the presence of VDR as the 1,25D-binding protein associated with plasma membrane caveolae. PMID- 15272055 TI - Ectodomain shedding-dependent transactivation of epidermal growth factor receptors in response to insulin-like growth factor type I. AB - Diverse extracellular stimuli activate the ERK1/2 MAPK cascade by transactivating epidermal growth factor (EGF) receptors. Here, we have examined the role of EGF receptors in IGF-I-stimulated ERK1/2 activation in several cultured cell lines. In human embryonic kidney 293 cells, IGF-I triggered proteolysis of heparin binding (HB)-EGF, increased tyrosine autophosphorylation of EGF receptors, stimulated EGF receptor inhibitor (AG1478)-sensitive ERK1/2 phosphorylation, and promoted EGF receptor endocytosis. In a mixed culture system that employed IGF-I receptor null murine embryo fibroblasts (MEFs) (R(-) cells) to detect paracrine signals produced by MEFs expressing the human IGF-I receptor (R(+) cells), stimulation of R(+) cells provoked rapid activation of green fluorescent protein tagged ERK2 in cocultured R(-) cells. The R(-) cell response was abolished by either the broad-spectrum matrix metalloprotease inhibitor batimastat or by AG1478, indicating that it resulted from the proteolytic generation of an EGF receptor ligand from adjacent R(+) cells. These data suggest that the paracrine production of EGF receptor ligands leading to EGF receptor transactivation is a general property of IGF-I receptor signaling. In contrast, the contribution of transactivated EGF receptors to IGF-I-stimulated downstream events, such as ERK1/2 activation, varies in a cell type-dependent manner. PMID- 15272056 TI - Acanthamoeba induces cell-cycle arrest in host cells. AB - Acanthamoeba can cause fatal granulomatous amoebic encephalitis (GAE) and eye keratitis. However, the pathogenesis and pathophysiology of these emerging diseases remain unclear. In this study, the effects of Acanthamoeba on the host cell cycle using human brain microvascular endothelial cells (HBMEC) and human corneal epithelial cells (HCEC) were determined. Two isolates of Acanthamoeba belonging to the T1 genotype (GAE isolate) and T4 genotype (keratitis isolate) were used, which showed severe cytotoxicity on HBMEC and HCEC, respectively. No tissue specificity was observed in their ability to exhibit binding to the host cells. To determine the effects of Acanthamoeba on the host cell cycle, a cell cycle-specific gene array was used. This screened for 96 genes specific for host cell-cycle regulation. It was observed that Acanthamoeba inhibited expression of genes encoding cyclins F and G1 and cyclin-dependent kinase 6, which are proteins important for cell-cycle progression. Moreover, upregulation was observed of the expression of genes such as GADD45A and p130 Rb, associated with cell-cycle arrest, indicating cell-cycle inhibition. Next, the effect of Acanthamoeba on retinoblastoma protein (pRb) phosphorylation was determined. pRb is a potent inhibitor of G1-to-S cell-cycle progression; however, its function is inhibited upon phosphorylation, allowing progression into S phase. Western blotting revealed that Acanthamoeba abolished pRb phosphorylation leading to cell-cycle arrest at the G1-to-S transition. Taken together, these studies demonstrated for the first time that Acanthamoeba inhibits the host cell cycle at the transcriptional level, as well as by modulating pRb phosphorylation using host cell-signalling mechanisms. A complete understanding of Acanthamoeba-host cell interactions may help in developing novel strategies to treat Acanthamoeba infections. PMID- 15272057 TI - Amphotericin B enhances the synthesis and release of the immunosuppressive agent gliotoxin from the pulmonary pathogen Aspergillus fumigatus. AB - Exposure of the pulmonary pathogen Aspergillus fumigatus to amphotericin B alters membrane permeability as indicated by the escape of amino acids and protein from the mycelium. Amphotericin B exposure for periods of 2-4 h also leads to increased release of the immunosuppressive agent gliotoxin into the surrounding culture medium. Examination of the intracellular gliotoxin concentration following exposure to amphotericin B indicated elevated levels within the hyphae as well as in the culture medium--an effect which was also evident upon exposure of A. fumigatus to DMSO. These results indicate that in parallel with the ability of amphotericin B to act as a fungistatic agent it can also induce the synthesis of gliotoxin and facilitate its release by increasing the permeability of the fungal cell membrane. Increased synthesis of gliotoxin may result from the commencement of secondary metabolism in the presence of amphotericin B. The ability of amphotericin B to enhance the synthesis and release of gliotoxin may exacerbate the effects of the toxin and facilitate fungal invasion of pulmonary tissue. PMID- 15272058 TI - Legionella-induced acute lung injury in the setting of hyperoxia: protective role of tumour necrosis factor-alpha. AB - Among the main characteristics of Legionella pneumophila pneumonia are acute lung injury and severe hypoxemia. Although high oxygen supplementation is a valuable supportive therapy in these patients, oxygen itself is known to be a risk factor for acute lung injury. The effects of hyperoxia on lung injury of mice with Legionella pneumonia were examined. Hyperoxia treatment reduced survival of the infected mice in an oxygen concentration- and exposure time-dependent manner. The enhanced lethality was associated with an increase in total lung weight and apoptosis markers, but not with bacterial burden in the lungs. Hyperoxia decreased the levels of the antioxidant glutathione (GSH) in infected lungs. Exogenous tumour necrosis factor-alpha (TNF-alpha) improved the survival of infected mice kept under hyperoxia. TNF-alpha effects were associated with restoration of total lung weight and histone DNA and GSH levels on day 2, whereas the lung bacterial burden did not differ significantly. Moreover, upregulation of GSH by TNF-alpha was observed in the lungs of mice without infection. These results demonstrate that hyperoxia exacerbates L. pneumophila pneumonia. The data suggest that TNF-alpha may be a potential therapeutic candidate for these individuals, not only through modulating host antibacterial systems, but also by mediating induction of the antioxidant GSH. PMID- 15272059 TI - Lipopolysaccharides of Bacteroides fragilis, Chlamydia trachomatis and Pseudomonas aeruginosa signal via toll-like receptor 2. AB - Recognition of bacterial lipopolysaccharide (LPS) is critical in the host defence against Gram-negative infection. While enterobacterial LPS signals via Toll-like receptor 4 (TLR4), it has recently been reported that the LPS of Leptospira interrogans, Legionella pneumophila, Rhizobium species Sin-1 and at least one strain of Porphyromonas gingivalis are capable of signalling via TLR2. Using a TLR transfection assay and measurement of an NF-kappaB-sensitive promoter region, the results show that the LPS of Bacteroides fragilis NCTC-9343, Chlamydia trachomatis LGV-1 and Pseudomonas aeruginosa PAC-611 also signal via TLR2 and it is pointed out that all TLR2-signalling LPS discovered to date demonstrate relatively weak endotoxicity in some models and structural features distinct from those LPS shown to signal via TLR4. PMID- 15272060 TI - Influence of normal microbiota on some aspects of the immune response during experimental infection with Trypanosoma cruzi in mice. AB - To study the influence of normal associated microbiota on systemic immunological responses during experimental Chagas' disease, germ-free and conventional NIH Swiss mice were infected with Y strain of Trypanosoma cruzi. Although no statistical differences in mortality and parasitaemia were found, conventional mice showed IFN-gamma, TNF-alpha and NO production (P < 0.05) by spleen cell cultures and higher blood levels of immunoglobulins of the IgG2a isotype (P < 0.05) when compared to their germ-free counterparts. Moreover, higher levels of IgG1 were also found in conventional animals. On the other hand, no differences in IL10 production were found between germ-free and conventional mice after infection (P < 0.05). Interestingly, spleen cell cultures from non-infected germ free mice spontaneously produced higher levels of IL10 than cultures from conventional mice. Moreover, cultures from non-infected germ-free mice responded to T. cruzi antigens with IFN-gamma production, contrary to cultures from conventional animals. In conclusion, the presence of the normal microbiota skews the immune response towards production of inflammatory cytokines during experimental infection with T. cruzi in mice. However, the increase in production of cytokines that is linked to resistance to this parasite did not alter the outcome of infection significantly, probably due to high virulence of the Y strain. PMID- 15272061 TI - Comparison of culture and PCR for detection of Bordetella pertussis and Bordetella parapertussis under routine laboratory conditions. AB - A PCR assay for the detection of Bordetella pertussis and Bordetella parapertussis was compared with the conventional culture method under routine laboratory conditions. Detection of B. pertussis was based on the amplification of a section of the IS481 insertion sequence and confirmation of positive results was based on a sequence of the pertussis toxin promoter region. Detection of B. parapertussis was based on the amplification of a section of the IS1001 insertion sequence. An internal control was included. Data were available for the period 28 November 2000 to 9 July 2003. In this period, 3096 patients were examined for infection with B. pertussis and B. parapertussis by culture and PCR on the same day. B. pertussis was found in 496 (16 %) patients; 208 (42 %) were diagnosed by PCR alone whereas 17 (3 %) were diagnosed by culture alone. B. parapertussis was found in 64 (2 %) patients. The sensitivity of the PCR was 97 % and of culture 58 %. The specificity of PCR was 93 % when regarding culture as 100 % sensitive. There was a significant relationship between laboratory method and age, as the superiority of PCR was most marked in the age group 0.5-3 years. The PCR assay proved highly sensitive for the diagnosis of pertussis. The specificity estimate of the PCR assay suffers from the influence of a gold-standard method with a low sensitivity. The PCR assay is considered highly specific due to the amplification of two different sequences in two separate assays. PMID- 15272062 TI - Underdiagnosis of urinary tract infection caused by Methylobacterium species with current standard processing of urine culture and its clinical implications. AB - Methylobacterium species are environmental opportunistic bacteria, and urinary tract infection (UTI) caused by these pathogens has not yet been documented. Four cases of UTI with Methylobacterium bacteraemia in immunocompetent female patients are reported. Their urine cultures, processed according to standard procedures (i.e. incubation at 35 degrees C in ambient air for 24 h before incubation at room temperature for a further 24 h), were either negative or positive for Escherichia coli. Specially designed experiments indicated that colonies of Methylobacterium species were visualized on blood agar only after incubation at 35 degrees C for at least 40 h, and growth was completely suppressed when concurrently incubated with much smaller inocula of E. coli. The isolates were variably susceptible to cephalosporins, but 100 % susceptible to aminoglycosides. This study suggests an underdiagnosis of UTI caused by Methylobacterium species when the standard procedure of processing urine cultures is used, and implies that administration of aminoglycosides is important when treatment of UTIs with cephalosporin fails. PMID- 15272063 TI - Antimicrobial-resistance and enterotoxin-encoding genes among staphylococci isolated from expressed human breast milk. AB - Resistance traits and the presence of enterotoxin-encoding genes were investigated in staphylococcus isolates obtained from expressed human breast milk. A total of 54 staphylococcal isolates identified as Staphylococcus epidermidis (53.6 %), Staphylococcus warneri (20.4 %), Staphylococcus haemolyticus (13 %) and Staphylococcus aureus (13 %) were investigated. By using a disc-diffusion method, higher rates of resistance, including intermediate resistance, were observed for penicillin (87 %) and erythromycin (59.3 %). All strains were susceptible to clindamycin and vancomycin. Minimal inhibitory concentration (MIC) was determined by a macrodilution method for four clinically relevant antimicrobial drugs. High rates of resistance or intermediate resistance were observed for erythromycin, gentamicin and oxacillin. Additionally, three isolates showed reduced susceptibility to vancomycin (MIC, 8 microg ml(-1)). Genetic determinants of resistance were detected by using PCR and the results showed good correlation with the macrodilution tests. Moreover, in four staphylococcus isolates, the presence of enterotoxin-encoding genes (seg, seh and sea) was identified. The results demonstrated that expressed human breast milk can be a reservoir of multiresistant staphylococci that may also harbour important virulent determinants. PMID- 15272064 TI - Anaerobic incubation conditions enhance pyrazinamide activity against Mycobacterium tuberculosis. AB - Pyrazinamide (PZA) is an unconventional front line tuberculosis drug characterized by high in vivo sterilizing activity, but poor in vitro activity. This disparity in PZA activity may reflect differences between the in vivo tissue environment and in vitro culture conditions. This study examined the effect of anaerobic conditions, which exist in granulomatous lesions in vivo, on PZA activity in vitro. Low oxygen enhanced the activity of PZA against Mycobacterium tuberculosis, with anaerobic conditions resulting in greater enhancement than microaerobic conditions. ATPase and respiratory chain enzyme inhibitors enhanced PZA activity under normal atmospheric conditions, but not under anaerobic conditions. Furthermore, the inhibitors did not enhance isoniazid or rifampicin activity. Nitrate as an alternative electron acceptor antagonized PZA activity under anaerobic conditions. These findings provide further support for a proposed mechanism of action of PZA in which the active form of PZA (pyrazinoic acid) depletes the membrane energy reserve. They also provide another explanation for the higher sterilizing activity of PZA within in vivo lesions with low oxygen than under in vitro drug susceptibility testing conditions with ambient oxygen. PMID- 15272065 TI - Ultrastructural examination of two cases of stromal microsporidial keratitis. AB - Two cases with chronic stromal keratitis are described in immunocompetent hosts where the diagnosis was originally thought to be herpetic or adenoviral disease. Light microscopy and ultrastructural examination of corneal tissue by electron microscopy were performed following penetrating keratoplasty (case 1) and corneal biopsy (case 2). Specimens from both cases were analysed for viral identification by PCR. Two different species of Microsporidia were identified. Case 1 represents the fourth reported case of corneal stromal Vittaforma corneae where the spores measured 3.3 x 1.4 microm, arranged in characteristic linear groups of about four to eight. Each spore contained a diplokaryotic nucleus and a single row of ten polar tube coils. By contrast, case 2 is the first reported case of stromal keratitis caused by Trachipleistophora hominis. In this case, spores measured 4 x 2.4 microm, located typically within packets. In this species, the polar tube was arranged as a single row of about 10-13 profiles. Viral DNA could not be amplified by PCR. In conclusion, microsporidial stromal keratitis should be considered in culture-negative cases refractory to medical therapy. As microbiological culture techniques are unsuccessful, diagnosis may only be established following histopathological and ultrastructural examination of corneal tissue. PMID- 15272066 TI - A fast, practical and reproducible procedure for the standardization of the cell density of an Aspergillus suspension. AB - The progressive increase of invasive disease and reports of resistance among Aspergillus species emphasizes the need for reproducible antifungal susceptibility testing. Inoculum standardization is a crucial step in such procedures. The objective of this study was to develop a fast and precise method of evaluating the cell density of an Aspergillus spore suspension, as an alternative to spectrometric readings or cell-counting with a haemocytometer. Densimat (bioMerieux) is a portable photometer that shows a good correlation with spectrometric readings and can advantageously replace the cumbersome, time consuming method of cell-counting. Thus, Densimat brings significant improvement to the reproducibility and feasibility of standardization of a fungal inoculum. PMID- 15272067 TI - Is there any relationship between Chlamydophila pneumoniae infection and juvenile idiopathic arthritis? AB - The role of Chlamydophila pneumoniae in the development and exacerbation of juvenile idiopathic arthritis (JIA) was investigated. Blood samples were taken from 60 JIA patients during an active disease period and for 4 weeks after. Synovial fluid samples were obtained from 20 of the 60 patients. In addition, 22 patients with familial Mediterranean fever (FMF) during the active period and 35 healthy children were included in the study as control groups. Synovial fluid samples were also obtained from three children with FMF. IgG, IgM and IgA levels against C. pneumoniae in serum samples were studied by immunofluorescence and IgG antibody and PCR studies were performed for C. pneumoniae DNA in synovial fluid samples. Twenty-nine (48.3 %) patients with JIA, 18 (81.8 %) patients with FMF and 22 (62.8 %) healthy children were found to be pre-infected with C. pneumoniae. Pre-infection with C. pneumoniae among FMF patients was found to be significantly higher than among those with JIA. We did not find a significant difference between JIA patients and healthy children. Chronic C. pneumoniae infection was observed only in six JIA patients, one FMF patient and two healthy children. Synovial fluid antibodies were found at higher than 1/512-fold dilution in one JIA patient and four times higher than normal serum in three JIA patients. C. pneumoniae DNA was not detected in any synovial fluid sample from FMF or JIA patients by PCR. In conclusion, C. pneumoniae infection does not have a triggering or a progressive effect on the clinical situation in JIA aetiopathogenesis, as a result of a multifactorial aetiology. New, extensive and serial studies (especially PCR studies of synovial tissue) are needed in order to confirm the indirect results. PMID- 15272068 TI - Molecular epidemiology of human P[8],G9 rotaviruses in Hungary between 1998 and 2001. AB - Increasing numbers of studies have documented the widespread distribution of human G9 rotaviruses and demonstrated that these strains may represent a fifth epidemiologically important G serotype. Serotype G9 strains were identified in Hungary for the first time in the 1997-1998 rotavirus season. Contrary to numerous surveys that reported several unexpected P-G combinations among recent G9 isolates (e.g. genotypes P[4], P[6] and P[19]), all Hungarian strains characterized to date possess the globally most common P-type, P[8], which was found among the first G9 isolates that were identified during the 1980s in the USA (WI61) and Japan (F45). To study the genetic variability within Hungarian G9 strains, RNA profile analysis and nucleotide sequencing were performed on a subset of samples that were collected between 1998 and 2001. These strains could be classified into four major RNA profiles, of which two were characteristic for epidemiologically major and two for epidemiologically minor G9 strains. Phylogenetic analysis demonstrated substantial sequence differences between the VP7 gene of Hungarian G9 strains and early strains that were isolated in the USA (WI61), Japan (F45) and India (116E) and a few recently identified isolates, e.g. from China (97'SZ37) and the USA (OM67) (< 90 % nucleotide sequence similarity). In contrast, the VP7 genes of Hungarian G9 strains were related very closely to the vast majority of G9 strains that were isolated in a variety of countries over the last several years (> 96 % nucleotide sequence similarity). With respect to the VP4 gene, Hungarian G9 rotaviruses fell into two of the major genetic lineages of genotype P[8], one corresponding to the epidemic strains (lineage II; P-like) and the other for two unique strains (lineage I; Wa-like), suggesting independent introduction of distinct P[8],G9 strains into Hungary or genetic reassortment between locally circulating P[8] strains and descendants of G9 isolates that were imported into the country at an earlier time. The unexpected heterogeneity found for G9 VP7 genes from several countries suggests that genetic variation among these strains has not yet been fully explored. PMID- 15272069 TI - Detection of Aspergillus DNA by a nested PCR assay is superior to blood culture in an experimental murine model of invasive aspergillosis. AB - For diagnosing invasive aspergillosis (IA), an increasing clinical problem in immunocompromised patients, molecular tools are gaining in importance. Detection of Aspergillus DNA in blood samples was investigated by a nested PCR assay in a murine model of experimentally induced IA. Ex vivo, the detection threshold of the PCR assay was determined in blood and organ homogenates of mice. After intravenous injection of Aspergillus fumigatus conidia on different days, growth of colonies was determined in cultures of blood and organs from immunocompetent and immunosuppressed mice and Aspergillus DNA was detected from blood samples by a nested PCR assay. The detection threshold of the PCR assay was as low as 1 c.f.u. ml(-1). The assay proved to be more sensitive than cultures of blood, with sensitivity rates between 17.6 and 87.5 % depending on the fungal burden. In conclusion, the nested PCR assay is superior to cultural methods in detecting Aspergillus spp. in murine blood samples. PMID- 15272070 TI - Chemotactic response of Helicobacter pylori to human plasma and bile. AB - To clarify further the role of chemotaxis in Helicobacter pylori colonization, the in vitro bacterium response to human plasma and bile (secretions containing chemoeffector compounds that are present in the gastric mucus layer) was examined. Human plasma, after dilution to 1 % (v/v) with buffer, was found to be a chemoattractant for the motile bacillus. Human gall-bladder bile, after dilution to 2 % (v/v) with buffer, was found to be a chemorepellent, but did not cause the motility of the bacillus to be diminished after prolonged exposure. The basis of the chemoattractant effect of plasma was explored by examining how urea and 12 amino acids found in plasma affected the taxis of H. pylori. Urea and the amino acids histidine, glutamine, glycine and arginine were the strongest chemoattractants. Other amino acids were chemoattractants, with the exceptions of aspartic and glutamic acids, which were chemorepellents. The basis of the chemorepellent effect of bile was explored by examining how the six most abundant conjugated bile acids in human bile affected the taxis of H. pylori. All the bile acids were chemorepellents, with the greatest effects being demonstrated by taurocholic and taurodeoxycholic acids. The implications of these findings for H. pylori colonization of gastric epithelium are discussed. PMID- 15272071 TI - Genetic diversity of the dnaJ gene in the Mycobacterium avium complex. AB - The Mycobacterium avium complex (MAC) is associated with various diseases in humans as a zoonosis. The dnaJ gene was partially sequenced in Schaefer's 28 reference strains of MAC, 14 human MAC isolates and 22 veterinary isolates. From substitutions affecting 21-32 nucleotides, all strains could be classified into 14 groups. Most nucleotide substitutions did not alter amino acid sequences. Approximately 8 % genetic diversity was seen in these strains, which divided into two clusters: cluster I (0.8 % genetic diversity), comprising the reference strain serotypes 1-6, 8-11 and 21 and all isolates; and cluster II (7 % genetic diversity), comprising the remaining reference strains. Analysis of the dnaJ gene in MAC may be useful in epidemiological studies. PMID- 15272072 TI - Isolated high-level ciprofloxacin resistance in Salmonella enterica subsp. enterica serotype Paratyphi A. PMID- 15272073 TI - Genetic basis for the evolution of vertebrate mineralized tissue. AB - Mineralized tissue is vital to many characteristic adaptive phenotypes in vertebrates. Three primary tissues, enamel (enameloid), dentin, and bone, are found in the body armor of ancient agnathans and mammalian teeth, suggesting that these two organs are homologous. Mammalian enamel forms on enamel-specific proteins such as amelogenin, whereas dentin and bone form on collagen and many acidic proteins, such as SPP1, coordinately regulate their mineralization. We previously reported that genes for three major enamel matrix proteins, five proteins necessary for dentin and bone formation, and milk caseins and salivary proteins arose from a single ancestor by tandem gene duplications and form the secretory calcium-binding phosphoprotein (SCPP) family. Gene structure and protein characteristics show that SCPP genes arose from the 5' region of ancestral sparcl1 (SPARC-like 1). Phylogenetic analysis on SPARC and SPARCL1 suggests that the SCPP genes arose after the divergence of cartilaginous fish and bony fish, implying that early vertebrate mineralization did not use SCPPs and that SPARC may be critical for initial mineralization. Consistent with this inference, we identified SPP1 in a teleost genome but failed to find any genes orthologous to mammalian enamel proteins. Based on these observations, we suggest a scenario for the evolution of vertebrate tissue mineralization, in which body armor initially formed on dermal collagen, which acted as a reinforcement of dermis. We also suggest that mammalian enamel is distinct from fish enameloid. Their similar nature as a hard structural overlay on exoskeleton and teeth is because of convergent evolution. PMID- 15272074 TI - Centromeric DNA sequences in the pathogenic yeast Candida albicans are all different and unique. AB - In an approach to clone and characterize centromeric DNA sequences of Candida albicans by chromatin immunoprecipitation, we have used antibodies directed against an evolutionarily conserved histone H3-like protein, CaCse4p (CENP-A homolog). Sequence analysis of clones obtained by this procedure reveals that only eight relatively small regions (approximately 3 kb each) of the Can. albicans genome are selectively enriched. These CaCse4-bound sequences are located within 4- to 18-kb regions lacking ORFs and occur once in each of the eight chromosomes of Can. albicans. Binding of another evolutionarily conserved kinetochore protein, CaMif2p (CENP-C homolog), colocalizes with CaCse4p. Deletion of the CaCse4p-binding region of chromosome 7 results in a high rate of loss of the altered chromosome, confirming that CaCse4p, a centromeric histone in the CENP-A family, indeed identifies the functional centromeric DNA of Can. albicans. The CaCse4p-rich regions not only lack conserved DNA motifs of point (<400 bp) centromeres and repeated elements of regional (>40 kb) centromeres, but also each chromosome of Can. albicans contains a different and unique CaCse4p-rich centromeric DNA sequence, a centromeric property previously unobserved in other organisms. PMID- 15272075 TI - Critical role for granulocyte colony-stimulating factor in inflammatory arthritis. AB - Granulocyte colony-stimulating factor (G-CSF) is a well known regulator of granulopoiesis, but the role of endogenous G-CSF in inflammatory joint disease has not been explored. We studied the response of G-CSF-deficient mice in experimental models of joint inflammation. We show that G-CSF deficiency protects mice from acute and chronic arthritis. Reduced severity was associated with blunted mobilization of granulocytic cells from the bone marrow and less cellular infiltrate and cellular activation in inflamed joints. We also demonstrate that G CSF blockade in established collagen-induced arthritis in WT mice markedly reduces disease manifestations and is as effective as tumor necrosis factor blockade. Our results reveal a critical role for G-CSF in driving joint inflammation and highlight G-CSF as a potential therapeutic target in inflammatory joint diseases, such as rheumatoid arthritis. PMID- 15272076 TI - Nitrate assimilation in plant shoots depends on photorespiration. AB - Photorespiration, a process that diminishes net photosynthesis by approximately 25% in most plants, has been viewed as the unfavorable consequence of plants having evolved when the atmosphere contained much higher levels of carbon dioxide than it does today. Here we used two independent methods to show that exposure of Arabidopsis and wheat shoots to conditions that inhibited photorespiration also strongly inhibited nitrate assimilation. Thus, nitrate assimilation in both dicotyledonous and monocotyledonous species depends on photorespiration. This previously undescribed role for photorespiration (i) explains several responses of plants to rising carbon dioxide concentrations, including the inability of many plants to sustain rapid growth under elevated levels of carbon dioxide; and (ii) raises concerns about genetic manipulations to diminish photorespiration in crops. PMID- 15272077 TI - Nuclear scaffold/matrix attached region modules linked to a transcription unit are sufficient for replication and maintenance of a mammalian episome. AB - The activation of mammalian origins of replication depends so far on ill understood epigenetic events, such as binding of transcription factors, chromatin structure, and nuclear localization. Understanding these mechanisms is not only a scientific challenge but also represents a prerequisite for the rational design of nonviral episomal vectors for mammalian cells. In this paper, we demonstrate that a tetramer of a 155-bp minimal nuclear scaffold/matrix attached region DNA module linked to an upstream transcription unit is sufficient for replication and mitotic stability of a mammalian episome in the absence of selection. Fluorescence in situ hybridization analyses, crosslinking with cis diammineplatinum(II)-dichloride and chromatin immunoprecipitation demonstrate that this vector associates with the nuclear matrix or scaffold in vivo by means of specific interaction of the nuclear scaffold/matrix attached region with the nuclear matrix protein SAF-A. Results presented in this paper define the minimal requirements of an episomal vector for mammalian cells on the molecular level. PMID- 15272078 TI - Simultaneous absence of dopamine D1 and D2 receptor-mediated signaling is lethal in mice. AB - Dopamine (DA) controls a wide variety of physiological functions in the central nervous system as well as in the neuroendocrine and gastrointestinal systems. DA signaling is mediated by five cloned receptors named D1-D5. Knockout mouse models for the five receptors have been generated, and, albeit impaired for some important DA-mediated functions, they are viable and can reproduce. D1 and D2 receptors are the most abundant and widely expressed DA receptors. Cooperative/synergistic effects mediated by these receptors have been suggested, in particular, in the control of motor behaviors. To analyze the extent of such interrelationship, we have generated double D1/D2 receptor mutants. Interestingly, in contrast to single knockouts, we found that concurrent ablation of the D1 and D2 receptors is lethal during the second or third week after birth. This dramatic phenotype is likely to be related to altered feeding behavior and dysfunction of the gastrointestinal system, especially because major anatomical changes were not identified in the brain. Similarly, in the absence of functional D1, heterozygous D2 mutants (D1r(-/-);D2r(+/-)) showed severe growth retardation and did not survive their postweaning period. The analysis of motor behavior in D1r/D2r compound mutants showed that loss of D2-mediated functions reduces motor abilities, whereas the effect of D1r ablation on locomotion strongly depends on the experimental paradigms used. These studies highlight the interrelationship between D1 and D2 receptor-mediated control of motor activity, food intake, and gastrointestinal functions, which has been elusive in the single-gene ablation studies. PMID- 15272079 TI - CSTX-13, a highly synergistically acting two-chain neurotoxic enhancer in the venom of the spider Cupiennius salei (Ctenidae). AB - The survival of the spider Cupiennius salei depends on its hunting success, which largely relies on its immediately paralyzing multicomponent venom. Here, we report on the isolation and characterization of CSTX-13, a neurotoxic enhancer in the spider venom. De novo elucidation of the disulfide bridge pattern of CSTX-13 and the neurotoxin CSTX-1 by tandem MS revealed an identical arrangement. However, in contrast to CSTX-1, CSTX-13 is a two-chain peptide with two interchain and two intrachain disulfide bridges. Furthermore, the insecticidal activity of CSTX-13 is synergistically increased in the presence of K+ ions as well as of the cytolytic peptide cupiennin 1a. We demonstrated that the weakly neurotoxic CSTX-13 enhances the paralytic activity of the neurotoxin CSTX-1 by 65% when it is administered with the latter at its entirely nontoxic physiological concentration, which is 440 times below its LD50 concentration. PMID- 15272080 TI - Chromatin landscape dynamics of the Il4-Il13 locus during T helper 1 and 2 development. AB - Il4 and Il13 encode the canonical T helper 2 (TH2) cytokines responsible both for promoting immune responses against extracellular pathogens and, when misregulated, causing allergic and autoimmune disease. The expression potential of these genes undergoes developmentally programmed repression and enhancement during commitment of naive CD4+ T cells to the mature T helper 1 (TH1) and TH2 fates, respectively. Thus, like the globin locus, the TH2 cytokine locus provides a highly tractable system to study a developmental fate choice leading to alternative transcriptional states of either silence or permissivity. We used quantitative chromatin immunoprecipitation and RT-PCR to correlate changes in the transcriptional states of Il4 and Il13 with markers of permissive chromatin across the Il4-Il13 locus in naive CD4+ T cells undergoing TH1 and TH2 differentiation. We provide evidence that DNaseI hypersensitive site V in the Il4 3' enhancer is the likely target for signals maintaining Il4 and Il13 transcriptional permissivity in naive cells. We also demonstrate rapid acquisition of differences in H3 acetylation between TH1- and TH2-primed cells, indicating a developmentally early role for cytokine signaling in the process of TH cell fate determination. Finally, we show that transcriptional repression correlates with the disappearance of permissive H3 modifications from everywhere in the Il4-Il13 locus except hypersensitive site IV, suggesting a critical role for this element in the maintenance of transcriptional repression. Our findings are consistent with a progressive regulatory element activation/deactivation model of TH1/TH2 development. PMID- 15272081 TI - 5' Long serial analysis of gene expression (LongSAGE) and 3' LongSAGE for transcriptome characterization and genome annotation. AB - Complete genome annotation relies on precise identification of transcription units bounded by a transcription initiation site (TIS) and a polyadenylation site (PAS). To facilitate this process, we developed a set of two complementary methods, 5' Long serial analysis of gene expression (LS) and 3'LS. These analyses are based on the original SAGE and LS methods coupled with full-length cDNA cloning, and enable the high-throughput extraction of the first and the last 20 bp of each transcript. We demonstrate that the mapping of 5'LS and 3'LS tags to the genome allows the localization of TIS and PAS. By using 537 tag pairs mapping to the region of known genes, we confirmed that >90% of the tag pairs appropriately assigned to the first and last exons. Moreover, by using tag sequences as primers for RT-PCRs, we were able to recover putative full-length transcripts in 81% of the attempts. This large-scale generation of transcript terminal tags is at least 20-40 times more efficient than full-length cDNA cloning and sequencing in the identification of complete transcription units. The apparent precision and deep coverage makes 5'LS and 3'LS an advanced approach for genome annotation through whole-transcriptome characterization. PMID- 15272082 TI - Herpes simplex virus type-1 induces IFN-alpha production via Toll-like receptor 9 dependent and -independent pathways. AB - Type I IFN production in response to the DNA virus herpes simplex virus type-1 (HSV-1) is essential in controlling viral replication. We investigated whether plasmacytoid dendritic cells (pDC) were the major tissue source of IFN-alpha, and whether the production of IFN-alpha in response to HSV-1 depended on Toll-like receptor 9 (TLR9). Total spleen cells or bone marrow (BM) cells, or fractions thereof, including highly purified pDC, from WT, TLR9, and MyD88 knockout mice were stimulated with known ligands for TLR9 or active HSV-1. pDC freshly isolated from both spleen and BM were the major source of IFN-alpha in response to oligodeoxynucleotides containing CpG motifs, but in response to HSV-1 the majority of IFN-alpha was produced by other cell types. Moreover, IFN-alpha production by non-pDC was independent of TLR9. The tissue source determined whether pDC responded to HSV-1 in a strictly TLR9-dependent fashion. Freshly isolated BM pDC or pDC derived from culture of BM precursors with FMS-like tyrosine kinase-3 ligand, produced IFN-alpha in the absence of functional TLR9, whereas spleen pDC did not. Heat treatment of HSV-1 abolished maturation and IFN alpha production from all TLR9-deficient DC but not WT DC. Thus pDC and non-pDC produce IFN-alpha in response to HSV-1 via both TLR9-independent and -dependent pathways. PMID- 15272083 TI - Neutron crystallographic study on rubredoxin from Pyrococcus furiosus by BIX-3, a single-crystal diffractometer for biomacromolecules. AB - The structure of a partially deuterated rubredoxin from the hyperthermophilic archaeon Pyrococcus furiosus, an organism that grows optimally at 100 degrees C, was determined by using the neutron single-crystal diffractometer dedicated for biological macromolecules (BIX-3) at the JRR-3M reactor of the Japan Atomic Energy Research Institute. Data were collected at room temperature up to a resolution of 1.5 A, and the completeness factor of the data set was 81.9%. The model contains 306 H and 50 D atoms. A total of 37 hydration water molecules were identified, with 15 having all three atoms fully located and the remaining D2O molecules partially defined. The model has been refined to final agreement factors of R = 18.6% and Rfree = 21.7%. Several orientations of the O-D bonds of side chains, whose assignments from x-ray data were previously ambiguous, were clearly visible in the neutron structure. Although most backbone N-H bonds had undergone some degree of H/D exchange throughout the rubredoxin molecule, 5 H atom positions still had distinctly negative (H) peaks. The neutron Fourier maps clearly showed the details of an extensive set of H bonds involving the ND3+ terminus that may contribute to the unusual thermostability of this molecule. PMID- 15272084 TI - Detection of 91 potential conserved plant microRNAs in Arabidopsis thaliana and Oryza sativa identifies important target genes. AB - MicroRNAs (miRNAs) are an extensive class of tiny RNA molecules that regulate the expression of target genes by means of complementary base pair interactions. Although the first miRNAs were discovered in Caenorhabditis elegans, >300 miRNAs were recently documented in animals and plants, both by cloning methods and computational predictions. We present a genome-wide computational approach to detect miRNA genes in the Arabidopsis thaliana genome. Our method is based on the conservation of short sequences between the genomes of Arabidopsis and rice (Oryza sativa) and on properties of the secondary structure of the miRNA precursor. The method was fine-tuned to take into account plant-specific properties, such as the variable length of the miRNA precursor sequences. In total, 91 potential miRNA genes were identified, of which 58 had at least one nearly perfect match with an Arabidopsis mRNA, constituting the potential targets of those miRNAs. In addition to already known transcription factors involved in plant development, the targets also comprised genes involved in several other cellular processes, such as sulfur assimilation and ubiquitin-dependent protein degradation. These findings considerably broaden the scope of miRNA functions in plants. PMID- 15272085 TI - siRNA-mediated gene silencing: a global genome view. AB - The task of specific gene knockdown in vitro has been facilitated through the use of short interfering RNA (siRNA), which is now widely used for studying gene function, as well as for identifying and validating new drug targets. We explored the possibility of using siRNA for dissecting cellular pathways by siRNA-mediated gene silencing followed by gene expression profiling and systematic pathway analysis. We used siRNA to eliminate the Rb1 gene in human cells and determined the effects of Rb1 knockdown on the cell by using microarray-based gene expression profiling coupled with quantitative pathway analysis using the GenMapp and MappFinder software. Retinoblastoma protein is one of the key cell cycle regulators, which exerts its function through its interactions with E2F transcription factors. Rb1 knockdown affected G1/S and G2/M transitions of the cell cycle, DNA replication and repair, mitosis, and apoptosis, indicating that siRNA-mediated transient elimination of Rb1 mimics the control of cell cycle through Rb1 dissociation from E2F. Additionally, we observed significant effects on the processes of DNA damage response and epigenetic regulation of gene expression. Analysis of transcription factor binding sites was utilized to distinguish between putative direct targets and genes induced through other mechanisms. Our approach, which combines the use of siRNA-mediated gene silencing, mediated microarray screening and quantitative pathway analysis, can be used in functional genomics to elucidate the role of the target gene in intracellular pathways. The approach also holds significant promise for compound selection in drug discovery. PMID- 15272086 TI - A YY1-binding site is required for accurate human LINE-1 transcription initiation. AB - The initial step in Long Interspersed Element-1 (LINE-1) retrotransposition requires transcription from an internal promoter located within its 5' untranslated region (5'-UTR). Previous studies have identified a YY1 (Yin Yang 1) binding site as an important sequence in LINE-1 transcription. Here, we demonstrate that mutations in the YY1-binding site have only minor effects on transcription activation of the full-length 5'-UTR and LINE-1 mobility in a single round cultured cell retrotransposition assay. Instead, these mutations disrupt proper initiation of transcription from the +1 site of the 5'-UTR. Thus, we propose that the YY1-binding site functions as a component of the LINE-1 core promoter to direct accurate transcription initiation. Indeed, this sequence may explain the evolutionary success of LINE-1 by enabling full-length retrotransposed copies to undergo autonomous retrotransposition in subsequent generations. PMID- 15272087 TI - TFIIB-facilitated recruitment of preinitiation complexes by a TAF-independent mechanism. AB - Gene activators contain activation domains that are thought to recruit limiting components of the transcription machinery to a core promoter. VP16, a viral gene activator, has served as a model for studying the mechanistic aspects of transcriptional activation from yeast to human. The VP16 activation domain can be divided into two modules--an N-terminal subdomain (VPN) and a C-terminal subdomain (VPC). This study demonstrates that VPC stimulates core promoters that are either independent or dependent on TAFs (TATA-box Binding Protein-Associated Factors). In contrast, VPN only activates the TAF-independent core promoter and this activity increases in a synergistic fashion when VPN is dimerized (VPN2). Compared to one copy of VPN (VPN1), VPN2 also displays a highly cooperative increase in binding hTFIIB. The increased TFIIB binding correlates with VPN2's increased ability to recruit a complex containing TFIID, TFIIA and TFIIB. However, VPN1 and VPN2 do not increase the assembly of a complex containing only TFIID and TFIIA. The VPN subdomain also facilitates assembly of a complex containing TBP:TFIIA:TFIIB, which lacks TAFs, and provides a mechanism that could function at TAF-independent promoters. Taken together, these results suggest the interaction between VPN and TFIIB potentially initiate a network of contacts allowing the activator to indirectly tether TFIID or TBP to DNA. PMID- 15272088 TI - New invMED1 element cis-activates human multidrug-related MDR1 and MVP genes, involving the LRP130 protein. AB - The MDR1 gene is a key component of the cytotoxic defense network and its overexpression results in the multidrug resistance (MDR) phenotype. However, the molecular mechanisms that regulate the MDR1 gene and coordinate multiple MDR related genes expression are poorly understood. In a previous study, we identified a new 12 bp cis-activating region in the 5'-flanking region of the human MDR1 gene, which we called inverted MED1. In the present study, we characterized the precise binding element, which we named invMED1, and revealed the presence of the LRP130 protein as the nuclear factor. Its binding intensity increases with the endogenous MDR1 geneexpression and with the MDR level of CEM leukemia cells. Interestingly, the LRP130 level did not vary with the chemoresistance level. We observed the involvement of LRP130 in the transcriptional activity of the MDR1 gene promoter, and moreover, in that of the MDR-related, invMED1-containing, MVP gene promoter. We used siRNAs and transcriptional decoys in two unrelated human cancer cell lines to show the role of the invMED1/LRP130 couple in both MDR1 and MVP endogenous genes activities. We showed that invMED1 was localized in the -105/-100 and -148/-143 regions of the MDR1 and MVP gene promoters, respectively. In addition, since the invMED1 sequence is primarily located in the -160/-100 bp region of mammalian MDR-related genes, our results present the invMED1/LRP130 couple as a potential central regulator of the transcription of these genes. PMID- 15272089 TI - Cardiovascular reactivity to work stress predicts subsequent onset of hypertension: the Air Traffic Controller Health Change Study. AB - OBJECTIVE: The hypothesis that increased blood pressure reactivity to stress is an early risk marker of hypertension was tested in a 1994 follow-up of the 1974 to 1978 Air Traffic Controller Health Change Study sample. METHODS: Assessments in 1974 to 1978 included physical examinations and recordings (every 20 minutes for 5 hours) of both workload (planes within controller airspace) and blood pressure reactivity. Individual differences in reactivity were used to predict 1994 self-report of ever having been told by a physician to take antihypertensive medication, assessed in a telephone survey of 218 respondents who were normotensive or stage 1 hypertensive in 1974 to 1978. RESULTS: Each SD increase in baseline systolic reactivity was associated with a 1.7 (p <.019) increase in the relative-odds of 1994 hypertension, after controlling for age, body mass index, and clinic systolic and diastolic blood pressure at clinical examination, with effects comparable for baseline normotensives and stage 1 hypertensives. CONCLUSION: A 20-year follow-up of originally normotensive and stage I hypertensive workers suggests that increased systolic blood pressure reactivity to work stress is associated with long-term risk of hypertension. PMID- 15272090 TI - Depression and late mortality after myocardial infarction in the Enhancing Recovery in Coronary Heart Disease (ENRICHD) study. AB - OBJECTIVE: The Enhancing Recovery in Coronary Heart Disease study was a multicenter clinical trial in which patients with depression and/or low perceived social support after an acute myocardial infarction were randomly assigned to an intervention consisting of cognitive behavior therapy and, in some cases, sertraline, or to usual care. There was no difference in survival between the groups. A possible reason why the intervention failed to affect survival is that too many patients with mild, transient depression were enrolled. Another is that some patients died too soon to complete the intervention. This analysis evaluates whether there was a difference in late (ie, > or =6 months after the myocardial infarction) mortality among initially depressed patients who had a Beck Depression Inventory score > or =10 and a past history of major depression, and who completed the 6-month post-treatment assessment. It also examines the relationship between change in depression and late mortality. METHODS: Out of the 1,165 (47%) of the Enhancing Recovery in Coronary Heart Disease study participants who met our criteria, 57 died in the first 6 months, and 858 (409 usual care, 449 intervention) completed the 6-month assessment. Cox regression was used to analyze survival. RESULTS: The intervention did not affect late mortality. However, intervention patients whose depression did not improve were at higher risk for late mortality than were patients who responded to treatment. CONCLUSIONS: Patients whose depression is refractory to cognitive behavior therapy and sertraline, two standard treatments for depression, are at high risk for late mortality after myocardial infarction. PMID- 15272091 TI - Psychosocial treatment within sex by ethnicity subgroups in the Enhancing Recovery in Coronary Heart Disease clinical trial. AB - OBJECTIVE: Intervening in depression and/or low perceived social support within 28 days after myocardial infarction (MI) in the Enhancing Recovery in Coronary Heart Disease (ENRICHD) clinical trial did not increase event-free survival. The purpose of the present investigation was to conduct post hoc analyses on sex and ethnic minority subgroups to assess whether any treatment subgroup is at reduced or increased risk of greater morbidity/mortality. METHODS: The 2481 patients with MI (973 white men, 424 minority men, 674 white women, 410 minority women) who had major or minor depression and/or low perceived social support were randomly allocated to usual medical care or cognitive behavior therapy. Total mortality or recurrent nonfatal MI (ENRICHD primary endpoint) and cardiac mortality or recurrent nonfatal MI (secondary endpoint) were analyzed as composite endpoints by group for time to first event using Cox proportional hazards regression. RESULTS: There was a trend in the direction of treatment efficacy for white men for the primary endpoint (hazard ratio [HR], 0.80; 95% confidence interval, 0.61 1.05; p =.10) and a significant (p <.006, Bonferroni corrected) effect for the secondary endpoint (HR, 0.63; 95% CI, 0.46-0.87; p =.004). In contrast, the HRs for each of the other three subgroups were nonsignificant. The magnitude of differences in treatment effects between white men and the other subgroups remained significant for the secondary endpoint (p =.04) after adjustment for age, education, living alone, antidepressant use, comorbidity score, cardiac catheterization, ejection fraction, history of hypertension, and major depression. CONCLUSIONS: White men, but not other subgroups, may have benefited from the ENRICHD intervention, suggesting that future studies need to attend to issues of treatment design and delivery that may have prevented benefit among sex and ethnic subgroups other than white men. PMID- 15272092 TI - Association of fear of terror with low-grade inflammation among apparently healthy employed adults. AB - OBJECTIVE: Based on evidence that psychological stress may induce a chronic inflammatory process, we hypothesized that the stress caused by chronic fear of terror may be associated with low-grade inflammation. This hypothesis was examined in employed men and women with the presence of low-grade inflammation measured by high sensitivity C-reactive protein (CRP). METHODS: Apparently healthy employed adults (N = 1153) undergoing periodic health check-ups in a tertiary hospital in Israel completed a questionnaire. Fear of terror (scored 1 5) was assessed by three items measuring the extent to which respondents have deep concern for personal safety, elevated tension in crowded places, and fear of terror strikes causing harm to one's self or one's family members. The main outcome measure was the presence or absence of an elevated CRP level (>3.0 mg/L). RESULTS: Women scored significantly higher on fear of terror compared with men (M = 2.16 vs. M = 1.68, respectively; p <.0001). Most of the study participants who scored high (4 or 5) on fear of terror, reported having experienced this feeling for 1 year or more. In women only, there was a positive association between fear of terror and risk of elevated CRP level (adjusted OR = 1.7, 95% CI 1.2-2.4) in a multivariate model adjusting for generalized anxiety, depressive symptoms, and potentially confounding demographic and biomedical variables. CONCLUSIONS: Chronic fear of terror in women, but not in men, is associated with elevated CRP levels, which suggests the presence of low-grade inflammation and a potential risk of cardiovascular disease. PMID- 15272093 TI - Exaggerated platelet and hemodynamic reactivity to mental stress in men with coronary artery disease. AB - OBJECTIVE: This study compared the effects of acute mental stress on cardiovascular and subjective responses and platelet activation in male patients with established coronary artery disease (CAD) and age-matched controls. METHODS: We assessed 17 male CAD patients aged 44 to 59 years and 22 healthy male controls. Blood pressure, heart rate, and hemodynamics were assessed before, during, and up to 2 hours after administration of color/word and mirror tracing tasks. Blood was sampled at baseline, after tasks, and at 30 and 75 minutes after stress, and platelet activation was assessed by measuring platelet-leukocyte aggregates (PLAs) using flow cytometry. RESULTS: CAD patients showed significantly greater systolic blood pressure stress responses than controls (mean increases of 43.9 and 28.3 mm Hg, adjusted for income, body mass index, waist/hip ratio, and medication), together with larger increases in heart rate (14.1 and 4.7 bpm) and cardiac index. Total peripheral resistance increased during the poststress recovery period in CAD patients but not in controls. PLAs increased with stress in both groups, but remained elevated at 75 minutes in CAD patients, returning to baseline in controls. Heart rate and cardiac index responses were correlated with increases in subjective stress and with depression ratings, whereas PLA responses were associated with ratings of task difficulty. CONCLUSION: Acute mental stress stimulated heightened cardiovascular responses in CAD patients, coupled with more prolonged platelet activation. These factors may contribute to plaque rupture and thrombogenesis, and partly mediate stress induced triggering of acute coronary syndromes. PMID- 15272094 TI - Elevated Plasma C-reactive protein in chronically distressed subjects who carry the A allele of the TNF-alpha -308 G/A polymorphism. AB - OBJECTIVE: Sustained psychological stress may result in a state operationalized as "vital exhaustion." Exhaustion predicted coronary artery disease (CAD) events whereby increased inflammatory activity might mediate this link. Moreover, there is an emerging importance of gene-environmental interactions in CAD. We investigated the effect of exhaustion severity on plasma levels of C-reactive protein (CRP) and whether exhaustion might regulate CRP levels via the -308G/A polymorphism of the tumor necrosis factor (TNF)-alpha gene. METHODS: We assessed exhaustion in 275 industrial employees (mean age +/- SD, 41 +/- 9 years, 88% men) using the Maastricht Questionnaire. Subjects were stratified as per exhaustion severity: none (N = 80), moderate (N = 128), and severe (N = 67). The TNF-alpha polymorphism was determined by real-time polymerase chain reaction, and plasma CRP levels were measured by a high-sensitivity immunoassay. RESULTS: There was a significant interaction between exhaustion and the TNF-alpha polymorphism, explaining 4.5% in the variance of plasma CRP values (F(5,271) = 2.47, p =.033); the result held after controlling for classic cardiovascular risk factors. Adjusted mean CRP levels across exhaustion strata in GA (N = 70) and AA (N = 3) carriers combined were 0.91 mg/l (none), 1.78 mg/l (moderate), and 2.61 mg/l (severe) as compared with 1.24 mg/l, 1.61 mg/l, and 1.36 mg/l for the GG wild type (N = 202). CONCLUSION: The findings suggest that the A allele of the TNF alpha -308 G/A polymorphism may mediate inflammation with exhaustion in a dose response relationship, while with the GG wild-type exhaustion severity seems unrelated to CRP levels. The finding provides a rationale for gene-environmental interactions by which psychosocial factors may promote atherosclerosis and CAD. PMID- 15272095 TI - Hostility is related to blunted beta-adrenergic receptor responsiveness among middle-aged women. AB - OBJECTIVE: Based on previous findings in men, the hypothesis that hostility would be associated with blunted responsiveness of cardiovascular beta-adrenergic receptors was tested in a study sample of middle-aged women. The roles of the sympathetic nervous system and of social support in this putative relationship were also evaluated. METHODS: Subjects were 80 healthy women (n = 23 African American; n = 57 white), aged 47 to 55 years. Hostility was assessed using the Cook-Medley Hostility Scale and social support was assessed with the Brief Social Support Questionnaire. Intravenous isoproterenol challenge was used to evaluate cardiac and vascular beta-adrenergic receptor responsiveness. Twenty-four-hour urinary catecholamine excretion was used to index sympathetic nervous system activity. RESULTS: Hostility was related to blunted cardiac (R = 0.33, p <.01) and vascular (R = 0.23, p <.05) beta-adrenergic receptor responsiveness in simple correlation analysis and in hierarchical regression analyses controlling for race, menopausal status, weight, and resting heart rate. Low social support was also related to blunted beta-adrenergic receptor responsiveness (R = 0.3, p <.01). Twenty-four-hour norepinephrine excretion was related both to hostility (R = 0.32, p <.01) and to cardiac (R = 0.25, p <.05) and vascular (R = 0.24, p <.05) beta-adrenergic receptor responsiveness. CONCLUSIONS: These observations replicate and extend previous findings in men by demonstrating that higher levels of hostility and low levels of social support are associated with blunted beta adrenergic receptor responsiveness in middle-aged women. They also suggest that heightened sympathetic nervous system activity associated with hostility may contribute to beta-adrenergic receptor blunting. Because blunted beta-adrenergic receptor sensitivity is a characteristic feature of a broad range of cardiovascular diseases, these findings may reflect an early preclinical manifestation of pathophysiology accompanying hostility and low social support. PMID- 15272096 TI - Predictors of depression three months after cardiac hospitalization. AB - OBJECTIVE: Depression occurs comorbidly in patients hospitalized for a range of cardiac conditions and procedures. This study examines the fluctuations in depressive symptomatology from index hospitalization to 3 months after hospitalization and determines predictors of depression 3 months after hospital admission for a cardiac condition or procedure. METHODS: Baseline clinical and demographic variables collected from a prospective study of the natural history of depression in 833 hospitalized cardiac patients were entered into a multinomial regression analysis. RESULTS: Similar proportions of participants were found to have no, mild, or moderate to severe depression at baseline and at 3 months, although 35.8% of participants had moved from one depression level to another during that period. Baseline characteristics predicting depression at 3 months after hospitalization were: a mild or moderate to severe level of depressive symptoms at hospitalization; younger age; smoking; self-reported previous diagnosis of a cardiac condition; and self-reported history of depression, anxiety, or stress. CONCLUSIONS: The five clinically accessible variables identified as predictors in this study may assist physicians in identification of cardiac patients who are at risk of persistent depression and who may require active intervention. Given that depression in cardiac patients is related to increased mortality and morbidity and that it is currently poorly diagnosed, these findings may have implications for preventing adverse outcomes. PMID- 15272097 TI - Posttraumatic stress, nonadherence, and adverse outcome in survivors of a myocardial infarction. AB - OBJECTIVE: Posttraumatic stress disorder (PTSD) symptoms have been reported in patients with coronary vascular disease, after the trauma of a myocardial infarction (MI). The effect of these symptoms on post-MI disease control has not been elucidated. We conducted a study that sought to determine whether PTSD symptoms post-MI are associated with increased likelihood of cardiovascular readmission and with nonadherence to treatment recommendations. METHODS: Patients were recruited during a visit in a cardiology clinic 6 months post-MI and were followed for 1 year. Adherence to aspirin was measured by platelet thromboxane production (an indication of aspirin's effect). Medical outcome was measured as rate of admission due to cardiovascular causes during the follow-up period. Self report measures of PTSD (Impact of Event Scale), Depression, and Global Distress (SCL-90-R) were administered at enrollment. RESULTS: Seventy-three patients were studied. Above-threshold PTSD symptom scores at enrollment, but not depression or global distress scores, were significant predictors of nonadherence to aspirin and of an increased likelihood of cardiovascular readmission over the course of the following year. CONCLUSIONS: PTSD symptoms predicted poor disease control in this cohort of MI survivors. The data suggest that screening MI survivors for symptoms of PTSD may be beneficial if this high-risk population is to be targeted for interventions. PMID- 15272098 TI - Role of spousal anxiety and depression in patients' psychosocial recovery after a cardiac event. AB - OBJECTIVE: The purposes of this study were to a) compare emotional responses and perception of control of patients and their spouses to myocardial infarction or coronary revascularization; and b) examine the relationship between spouses' emotional distress and patients' emotional distress and psychosocial adjustment to the cardiac event. METHODS: A total of 417 patient-spouse pairs were recruited after the patient was hospitalized for either acute myocardial infarction or coronary revascularization. We compared emotional responses of patients and spouses. The relationship between spouse anxiety and depression, and patient psychosocial distress was then determined. RESULTS: Spouses had higher levels of anxiety (p <.001) and depression (p <.001) than did patients, but there were no differences in level of hostility. Patients also expressed higher levels of perceived control than did spouses (p <.001). Spouse anxiety, depression, and perceived control remained correlated with patient psychosocial adjustment to illness, even when patient anxiety and depression were kept constant. Patients' psychosocial adjustment to illness was worse when spouses were more anxious or depressed than patients, and it was best when patients were more anxious or depressed than spouses, whereas psychosocial adjustment to illness was intermediate to these 2 extremes when patient and spouse anxiety and depression levels were similar (p =.001). CONCLUSION: Spouses often experience greater anxiety and depression and less perceived control than patients themselves. Attention to the psychological distress experienced by spouses of patients who have suffered a cardiac event may improve outcomes in patients. PMID- 15272099 TI - Depression and bone mineral density in young adults: results from NHANES III. AB - OBJECTIVE: The purpose of this cross-sectional population-based study was to assess the association of major depressive episode (MDE) and dysthymia with bone mineral density (BMD) in young adults. METHODS: Data are from a nationally representative sample of 5,171 people aged 20 to 39 years from the Third National Health and Nutrition Examination Survey. Total proximal femoral BMD was measured using dual energy x-ray absorptiometry. MDE and dysthymia were measured using the Diagnostic Interview Schedule. RESULTS: MDE was associated with lower BMD in multivariate models in men (mean BMD = 1.038 vs. 1.068 g/cm(2); odds ratio (OR) per 1 SD decline in BMD = 1.65, 95% confidence interval (CI) = 1.08-2.52; p = 0.02) but not in women (mean BMD = 0.982 vs. 0.979 g/cm(2); OR = 0.96, 95% CI = 0.71-1.30; p =.79). The same divergence by gender was seen for dysthymia. CONCLUSION: The relationship between BMD and MDE or dysthymia in young adults varies by gender. PMID- 15272100 TI - Treatment of somatoform disorders with St. John's wort: a randomized, double blind and placebo-controlled trial. AB - OBJECTIVE: To investigate efficacy and safety of St. John's wort (SJW) LI 160 in somatoform disorders. METHODS: In a prospective, randomized, placebo-controlled, and double-blind parallel group study, 184 outpatients with somatization disorder (ICD-10 F45.0), undifferentiated somatoform disorder (F45.1), and somatoform autonomic dysfunction (F45.3), but not major depression, received either 300 mg of SJW extract LI 160 twice daily or matching placebo for 6 weeks. Six outcome measures were evaluated as a combined measure by means of the Wei Lachin test: Somatoform Disorders Screening Instrument--7 days (SOMS-7), somatic subscore of the HAMA, somatic subscore of the SCL-90-R, subscores "improvement" and "efficacy" of the CGI, and the global judgment of efficacy by the patient. RESULTS: In the intention to treat population (N=173), for each of the six primary efficacy measures as well as for the combined test, statistically significant medium to large-sized superiority of SJW treatment over placebo was demonstrated (p <.0001). Of the SJW patients, 45.4% were classified as responders compared with 20.9% with placebo (p =.0006). Tolerability of SJW treatment was equivalent to placebo. CONCLUSIONS: Administration of 600 mg of SJW extract LI 160 daily is effective and safe in the treatment of somatoform disorders, thereby confirming results from a previous study. PMID- 15272101 TI - Total serum cholesterol and suicidality in anorexia nervosa. AB - OBJECTIVE: No published study has evaluated the relationship between serum cholesterol and suicidality in anorexia nervosa (AN). AIMS: To assess psychiatric and nutritional correlates of serum cholesterol in a sample of AN patients. METHODS: Serum cholesterol and nutritional status were evaluated in a sample of 74 AN patients, before starting any type of refeeding. All subjects underwent a structured clinical interview and completed the Hopkins Symptom Checklist. RESULTS: Subjects who reported previous suicide attempts, impulsive self injurious behavior, or current suicidal ideation showed significantly lower cholesterol levels than subjects without suicidality. Cholesterol levels were negatively correlated with the severity of depressive symptoms in all the patients with the exception of those with recurrent binge eating. A multivariate analysis showed that the relationships between cholesterol levels and suicidal behavior and ideation do not seem to be affected by the nutritional and metabolic factors considered in the study. CONCLUSIONS: Notwithstanding the influence of important metabolic factors affecting cholesterolemia in AN, our research tends to confirm previous studies that have found an association between low cholesterol levels and suicidality. PMID- 15272102 TI - Childhood socioeconomic status and host resistance to infectious illness in adulthood. AB - OBJECTIVE: Low childhood socioeconomic status (SES) is a risk factor for adult morbidity and mortality primarily attributable to cardiovascular disease. Here, we examine whether childhood SES is associated with adult host resistance to infectious illness, and whether the effect is limited to a critical period of low SES exposure, can be undone by changes in childhood SES, and is explained by adult SES. METHODS: Three hundred thirty-four healthy volunteers reported their own and their parents' level of education and the ages during their childhood when their parents owned their homes. Volunteers' current home ownership was recorded from real estate records. Subsequently, they were given nasal drops containing 1 of 2 rhinoviruses and were monitored in quarantine for infection and signs/symptoms of a common cold. RESULTS: For both viruses, susceptibility to colds decreased with the number of childhood years during which their parents owned their home (odds ratios by tertiles adjusted for demographics, body mass, season, and prechallenge viral-specific immunity were 3.7 for fewest years, 2.6 and 1). This decreased risk was attributable to both lower risk of infection and lower risk of illness in infected subjects. Moreover, those whose parents did not own their home during their early life but did during adolescence were at the same increased risk as those whose parents never owned their home. These associations were independent of parent education level, adult education and home ownership, and personality characteristics. CONCLUSIONS: A marker of low income and wealth during early childhood is associated with decreased resistance to upper respiratory infections in adulthood. Higher risk is not ameliorated by higher SES during adolescence and is independent of adult SES. PMID- 15272103 TI - Self-rated health is related to levels of circulating cytokines. AB - OBJECTIVE: Self-rated health is a powerful and independent predictor of long-term health, but its biological basis is unknown. Because factors associated with poor self-rated health (eg, pain, daily discomforts, and low energy and fitness) resemble symptoms of a generalized cytokine-induced sickness response, we examined the relationship between circulating cytokines and self-rated health. METHODS: In 265 consecutive primary health care patients (174 women and 91 men), we examined self-rated and physician-rated health, circulating levels of interleukin (IL)-1beta, IL-1 receptor antagonist (IL-1ra), IL-6, and tumor necrosis factor (TNF)-alpha as determined from plasma samples using high sensitivity enzyme-linked immunoassay. RESULTS: Self-rated health correlated with levels of IL-1beta (r = 0.27; p <.001), IL-1ra (r = 0.19; p <.05) and TNF-alpha (r = 0.46; p <.001) in women but not in men. Thus, poorer subjective health was associated with higher levels of inflammatory cytokines. Even when controlling for age, education, physical health, and diagnoses in multiple regression analyses, self-rated health was an independent and more robust predictor of cytokine levels than physician-rated health. CONCLUSIONS: The present findings suggest that an individual's health perception may be coupled to circulating cytokines. Because epidemiological research established that self-rated health predicts morbidity and mortality, the biological correlates and mechanisms of self-rated health need to be understood. PMID- 15272104 TI - Neurokinin-1 receptor mediates stress-exacerbated allergic airway inflammation and airway hyperresponsiveness in mice. AB - BACKGROUND: A wealth of clinical observation has suggested that stress and asthma morbidity are associated. We have previously established a mouse model of stress exacerbated allergic airway inflammation, which reflects major clinical findings. OBJECTIVE: The aim of the current study was to investigate the role of the neurokinin- (NK-)1 receptor in the mediation of stress effects in allergic airway inflammation. METHODS: BALB/c mice were systemically sensitized with ovalbumin (OVA) on assay days 1, 14, and 21 and repeatedly challenged with OVA aerosol on days 26 and 27. Sound stress was applied to the animals for 24 hours, starting with the first airway challenge. Additionally, one group of stressed and one group of nonstressed mice received the highly specific NK-1 receptor antagonist RP 67580. Bronchoalveolar lavage fluid was obtained, and cell numbers and differentiation were determined. Airway hyperreactivity was measured in vitro by electrical field stimulation of tracheal smooth-muscle elements. RESULTS: Application of stress in sensitized and challenged animals resulted in a significant increase in leukocyte number in the bronchoalveolar lavage fluid. Furthermore, stressed animals showed enhanced airway reactivity. The increase of inflammatory cells and airway reactivity was blocked by treatment of animals with the NK-1 receptor antagonist. CONCLUSION: These data indicate that the NK-1 receptor plays an important role in mediating stress effects in allergen-induced airway inflammation. PMID- 15272105 TI - Cynical hostility, socioeconomic position, health behaviors, and symptom load: a cross-sectional analysis in a Danish population-based study. AB - OBJECTIVE: To analyze the cross-sectional association between cynical hostility and high symptom load in a Danish population-based study. Furthermore, the aim was to investigate to what extent health risk behaviors mediated this association. METHODS: Data were based on a postal questionnaire in a Danish random sample of 3426 men and 3699 women aged 40 or 50 years. Cynical hostility was measured by the 8-item Cynical Distrust Scale. High symptom load was assessed by physiological and mental symptoms experienced within the last 4 weeks. Confounders were age and socioeconomic position, while potential mediators were alcohol consumption, smoking, physical activity, and BMI. RESULTS: Higher cynical hostility was associated with self-reported symptom load. Health behaviors did not seem to mediate this effect. Socioeconomic position was a strong confounder for the effect on both health and health behaviors. After adjustment the effects of hostility on health remained with odds ratios of 2.1 (1.7-2.6) for women and 2.3 (1.8-2.8) for men. CONCLUSION: After adjustment for socioeconomic position, cynical hostility has an effect on self-reported high symptom load, and this effect is not mediated by health behaviors. PMID- 15272106 TI - Changes in tolerance to rectal distension correlate with changes in psychological state in patients with severe irritable bowel syndrome. AB - OBJECTIVE: Reduced tolerance to rectal distension has been regarded as a biological marker for irritable bowel syndrome (IBS), but longitudinal studies are few. This study determined whether change in tolerance to rectal distension after psychological treatments was associated with: 1) change in abdominal pain; 2) change in psychological symptoms; 3) a reported history of sexual abuse. METHODS: Participants completed a visual analogue scale of abdominal pain, SCL-90 and Hamilton rating scale of depression; discomfort threshold to rectal distension was determined using a double random staircase protocol. These were measured at entry to a trial of psychotherapy or paroxetine (selective serotonin reuptake inhibitor antidepressant) and 3 months later (N = 52). Analysis of change scores were adjusted for treatment group and baseline values. RESULTS: Increased tolerance to distension after treatment was associated with reduction in depression (r = -0.37, p =.008) but not abdominal pain. Patients who reported prior sexual abuse showed greater increase in tolerance than the remainder (changes in volume threshold: -24.7 ml [SEM = 12.1] vs. 3.6 ml [SEM = 6.2], adjusted p =.045; changes in pressure threshold: -4.7 [SEM = 1.7] mm Hg vs. 0.96 [SEM=0.9], adjusted p =.005). Multiple regression indicated that reduction in depression score and a reported history of sexual abuse were independently associated with improved tolerance to distension. CONCLUSIONS: In patients with severe IBS, increased tolerance to rectal distension after psychological treatment is significantly associated with improved depression and reported sexual abuse. These results suggest that in some patients with severe IBS psychological rather than biological processes are primarily responsible for reduced tolerance to rectal distension. PMID- 15272107 TI - Elevated resting blood pressure and dampened emotional response. AB - OBJECTIVE: Increased blood pressure is associated with decreased reports of aversiveness for both physical pain and psychosocial stressors. Based on these findings, higher blood pressure could be associated with altered emotional responses to a broader range of stimuli. There are at least 3 ways this could happen: a) less dire response to negative stimuli with no change in response to positive stimuli; b) more positive responses to both negative and positive stimuli; or c) dampened emotional responses to both positive and negative stimuli. METHODS: Sixty-five normotensive volunteers had their resting blood pressure measured, then rated their emotional responses to a series of positive and negative photographs. RESULTS: Resting systolic blood pressure was significantly and negatively correlated with subjective emotional ratings of both positive (r = -.26) and negative (r = -.35) photographs. CONCLUSION: Results were consistent with emotion dampening for elevated resting blood pressure and may reflect homeostatic integration of neurocirculatory control and affect regulation. PMID- 15272108 TI - Responses to controlled diesel vapor exposure among chemically sensitive Gulf War veterans. AB - OBJECTIVE: A significant proportion of Gulf War veterans (GWVs) report chemical sensitivity, fatigue, and unexplained symptoms resulting in ongoing disability. GWVs frequently recall an association between diesel and petrochemical fume exposure and symptoms during service. The purpose of the present study among GWVs was to evaluate the immediate health effects of acute exposure to chemicals (diesel vapors with acetaldehyde) with and without stress. METHODS: In a single, controlled exposure to 5 parts per million (ppm) diesel vapors, symptoms, odor ratings, neurobehavioral performance, and psychophysiologic responses of 12 ill GWVs (GWV-I) were compared with 19 age- and gender-matched healthy GWVs (GWV-H). RESULTS: Relative to baseline and to GWV-H, GWV-I reported significantly increased symptoms such as disorientation and dizziness and displayed significantly reduced end-tidal CO(2) just after the onset of exposure. As exposure increased over time, GWV-I relative to GWV-H reported significantly increased symptoms of respiratory discomfort and general malaise. GWV-I were also physiologically hyporeactive in response to behavioral tasks administered during but not before exposure. CONCLUSIONS: Current symptoms among GWV-I may be exacerbated by ongoing environmental chemical exposures reminiscent of the Gulf War. Both psychologic and physiologic mechanisms contribute to current symptomatic responses of GWV-I. PMID- 15272109 TI - Sensory and affective pain discrimination after inhalation of essential oils. AB - OBJECTIVE: The purpose of this investigation was to examine the effects of olfactory absorption of two commonly used therapeutic essential oils on sensory and affective responses to experimentally induced pain. METHODS: A sex-balanced (13 men and 13 women) randomized crossover design was used to obtain pre- and posttreatment change scores for quantitative sensory ratings of contact heat, pressure, and ischemic pain across separate inhalation treatment conditions using essential oil of lavender, essential oil of rosemary, and distilled water (control). Subjective reports of treatment-related changes in pain intensity and pain unpleasantness were obtained for each condition using a visual analog scale. We interpret our findings with respect to the separate dimensions of sensory and affective processing of pain. RESULTS: Analyses revealed the absence of changes in quantitative pain sensitivity ratings between conditions. However, retrospectively, subjects' global impression of treatment outcome indicated that both pain intensity and pain unpleasantness were reduced after treatment with lavender and marginally reduced after treatment with rosemary, compared with the control condition. CONCLUSION: These findings suggest that aromatherapy may not elicit a direct analgesic effect but instead may alter affective appraisal of the experience and consequent retrospective evaluation of treatment-related pain. PMID- 15272110 TI - Attenuation of laboratory-induced stress in humans after acute administration of Melissa officinalis (Lemon Balm). AB - OBJECTIVE: Melissa officinalis (lemon balm) is contemporaneously used as a mild sedative and/or calming agent. Although recent research has demonstrated modulation of mood in keeping with these roles, no studies to date have directly investigated the effects of this herbal medication on laboratory-induced psychological stress. METHODS: In this double-blind, placebo-controlled, randomized, balanced crossover experiment, 18 healthy volunteers received two separate single doses of a standardized M. officinalis extract (300 mg, 600 mg) and a placebo, on separate days separated by a 7-day washout period. Modulation of mood was assessed during predose and 1-hour postdose completions of a 20 minute version of the Defined Intensity Stressor Simulation (DISS) battery. Cognitive performance on the four concurrent tasks of the battery was also assessed. RESULTS: The results showed that the 600-mg dose of Melissa ameliorated the negative mood effects of the DISS, with significantly increased self-ratings of calmness and reduced self-ratings of alertness. In addition, a significant increase in the speed of mathematical processing, with no reduction in accuracy, was observed after ingestion of the 300-mg dose. CONCLUSION: These results suggest that the potential for M. officinalis to mitigate the effects of stress deserves further investigation. PMID- 15272111 TI - Influences of distress and alcohol consumption on the development of a delayed type hypersensitivity skin test response. AB - OBJECTIVE: This paper reports the effects of naturally occurring levels of distress on the development of delayed-type hypersensitivity (DTH) skin test responses. These in vivo measures provide a biologically relevant assessment of cellular immune competence. METHODS: Subjects (N = 166) were immunized (baseline) with the novel antigen, keyhole limpet hemocyanin (KLH), and DTH skin test responses against KLH were assessed 3 weeks later (follow-up). The CMI Multitest (Merieux, France), which evaluates DTH responses to a panel of seven antigens, was also administered at follow-up. Emotional distress was assessed at baseline and follow-up by the Profile of Mood States. RESULTS: Distress levels at baseline were associated with a reduced likelihood of developing DTH responses against KLH at follow-up (r = -0.22, p =.01). There was no relationship between distress at follow-up and cutaneous DTH in response to KLH (r = 0.09, p =.24) or in the Multitest (r = -0.03, p =.70). In addition, higher levels of alcohol consumption at baseline (r = -0.19, p =.02) and at follow-up (r = -0.20, p =.01) were associated with a decreased likelihood of developing cutaneous DTH against KLH. CONCLUSIONS: Everyday levels of distress predicted the capacity of the cellular arm of the immune system to exhibit recall responses to an antigen, when the experimental paradigm allowed the assessment of distress at the time of antigen sensitization. Moderate alcohol consumption independently affected cutaneous DTH. PMID- 15272112 TI - Association between poorer quality of life and psychiatric morbidity in patients with different dermatological conditions. AB - OBJECTIVE: To determine the relationship between skin-related quality of life and psychiatric morbidity in patients with different skin conditions. METHODS: We recruited all adults attending the outpatient clinics of the Dermatological Institute IDI-IRCCS, Rome, Italy, during 14 predetermined days. Eligible patients, who gave their informed consent, completed the Skindex-29 and the 12 item General Health Questionnaire (GHQ-12). We used a stringent cut-off threshold (> or =5 on the GHQ-12) for identification of psychiatric morbidity. Skindex-29 scale scores were computed separately for GHQ noncases and GHQ cases. RESULTS: A total of 2,136 patients were included in the analysis. For all skin conditions, GHQ cases had substantially poorer score in all 3 domains of quality of life, Symptoms, Emotions, and Functioning. Most differences remained significant after adjusting for clinical severity, age, sex, and education in multiple regression models. These differences were not as marked in the Symptoms scale for some conditions known to be nearly asymptomatic (eg, alopecia, vitiligo, nevi), suggesting that, although patients with psychiatric morbidity might be more burdened by their symptoms, nevertheless they do not perceive nonexistent symptoms. CONCLUSION: In most skin conditions we considered, psychiatric morbidity was strongly associated with poorer quality of life. Although the cross sectional nature of our study does not allow identification of the direction of this association, care for the psychological condition of patients might have an impact on their quality of life. PMID- 15272113 TI - Academic examinations and immunity: academic stress or examination stress? PMID- 15272114 TI - Wising up to diabetes. PMID- 15272115 TI - Aging cartilage and osteoarthritis--what's the link? AB - Cartilage aging can contribute to the development of osteoarthritis (OA), the most common cause of chronic pain and disability in older adults. Articular cartilage is a unique tissue from the perspective of aging in that the cells (chondrocytes) and the majority of the extracellular matrix proteins experience little turnover, resulting in a tissue that must withstand years of use and can also accumulate years of aging-associated changes. Accumulation of advanced glycation end products (AGEs) occurs in cartilage, and the potential role of AGEs in the development of OA is being investigated. An age-associated reduction in growth factor signaling and an increase in oxidative stress may also play an important role in the age-OA connection. Further elucidation of mechanisms that affect chondrocyte function with aging should lead to novel interventions designed to slow the aging process in cartilage with the goal of preventing age associated OA. PMID- 15272116 TI - "siRNAs and miRNAs": a meeting report on RNA silencing. PMID- 15272117 TI - RNA editing of a miRNA precursor. AB - Micro RNAs comprise a large family of small, functional RNAs with important roles in the regulation of protein coding genes in animals and plants. Here we show that human and mouse miRNA22 precursor molecules are subject to posttranscriptional modification by A-to-I RNA editing in vivo. The observed editing events are predicted to have significant implications for the biogenesis and function of miRNA22 and might point toward a more general role for RNA editing in the regulation of miRNA gene expression. PMID- 15272118 TI - Using an RNA secondary structure partition function to determine confidence in base pairs predicted by free energy minimization. AB - A partition function calculation for RNA secondary structure is presented that uses a current set of nearest neighbor parameters for conformational free energy at 37 degrees C, including coaxial stacking. For a diverse database of RNA sequences, base pairs in the predicted minimum free energy structure that are predicted by the partition function to have high base pairing probability have a significantly higher positive predictive value for known base pairs. For example, the average positive predictive value, 65.8%, is increased to 91.0% when only base pairs with probability of 0.99 or above are considered. The quality of base pair predictions can also be increased by the addition of experimentally determined constraints, including enzymatic cleavage, flavin mono-nucleotide cleavage, and chemical modification. Predicted secondary structures can be color annotated to demonstrate pairs with high probability that are therefore well determined as compared to base pairs with lower probability of pairing. PMID- 15272119 TI - Detection of a novel sense-antisense RNA-hybrid structure by RACE experiments on endogenous troponin I antisense RNA. AB - Conformational changes in the troponin/tropomyosin complex significantly alter the mechanical properties of cardiac muscle. Phosphorylation of cardiac troponin I, part of the troponin/tropomyosin complex, reduces calcium affinity, which leads to increased relaxation of cardiac muscle. Because cardiac troponin I plays a central role in tuning the heart to different work demands, detailed knowledge of troponin I protein regulation is required. Our group previously detected naturally occurring antisense RNA for troponin I in human and rat hearts, and here, attempt to unravel the structure of rat cardiac troponin I antisense RNA. We performed rapid amplification of cDNA ends (RACE) experiments and discovered antisense sequences identical to a copy of the sense mRNA, which led us to conclude that the antisense RNA must be transcribed from troponin I mRNA in the cytoplasm. Moreover, we isolated RNA structures comprising sense and antisense sequences in one continuous molecule. As we found no homolog structures described in the literature, we called this "hybrid RNA." Because a duplex formation was demonstrated previously we concluded that hybrid RNA is a consequence of a tight interaction between sense and antisense troponin I RNA in vivo, which we discuss in the article. PMID- 15272120 TI - Both U2 snRNA and U12 snRNA are required for accurate splicing of exon 5 of the rat calcitonin/CGRP gene. AB - Two classes of spliceosome are present in eukaryotic cells. Most introns in nuclear pre-mRNAs are removed by a spliceosome that requires U1, U2, U4, U5, and U6 small nuclear ribonucleoprotein particles (snRNPs). A minor class of introns are removed by a spliceosome containing U11, U12, U5, U4atac, and U6 atac snRNPs. We describe experiments that demonstrate that splicing of exon 5 of the rat calcitonin/CGRP gene requires both U2 snRNA and U12 snRNA. In vitro, splicing to calcitonin/ CGRP exon 5 RNA was dependent on U2 snRNA, as preincubation of nuclear extract with an oligonucleotide complementary to U2 snRNA abolished exon 5 splicing. Addition of an oligonucleotide complementary to U12 snRNA increased splicing at a cryptic splice site in exon 5 from <5% to 50% of total spliced RNA. Point mutations in a candidate U12 branch sequence in calcitonin/CGRP intron 4, predicted to decrease U12-pre-mRNA base-pairing, also significantly increased cryptic splicing in vitro. Calcitonin/CGRP genes containing base changes disrupting the U12 branch sequence expressed significantly decreased CGRP mRNA levels when expressed in cultured cells. Coexpression of U12 snRNAs containing base changes predicted to restore U12-pre-mRNA base pairing increased CGRP mRNA synthesis to the level of the wild-type gene. These observations indicate that accurate, efficient splicing of calcitonin/CGRP exon 5 is dependent upon both U2 and U12 snRNAs. PMID- 15272121 TI - New tertiary constraints between the RNA components of active yeast spliceosomes: a photo-crosslinking study. AB - Elucidation of the three-dimensional (3D) structures of the two sequential active sites in spliceosomes is essential for understanding the mechanism of premessenger RNA splicing. The mechanism is predicted to be catalyzed by the small nuclear RNA (snRNA) components of spliceosomes. To obtain new tertiary constraints between the RNA components, we produced and mapped crosslinks between U6 snRNA and the proximal RNAs of active yeast spliceosomes ("yeast" in this report is Saccharomyces cerevisiae). Thus, specific sites in U6, when substituted with a photoreactive 4-thiouridine or 5-iodouridine, produced spliceosome dependent crosslinks to U2 snRNA, or in one case, to the pre-mRNA substrate. One set of U2-U6 crosslinks formed before the Prp2p-dependent step of spliceosome assembly, whereas another set formed during or after this step but before the first chemical step of splicing. This latter set of crosslinks formed across U2 U6 helix I. Importantly, this set provides new tertiary constraints for developing 3D models of fully assembled yeast spliceosomes, which are poised for the first chemical step of splicing. PMID- 15272122 TI - In vivo selection reveals combinatorial controls that define a critical exon in the spinal muscular atrophy genes. AB - Humans have two near identical copies of the survival of motor neuron (SMN) gene, SMN1 and SMN2. In spinal muscular atrophy (SMA), SMN2 is not able to compensate for the loss of SMN1 due to an inhibitory mutation at position 6 (C6U mutation in transcript) of exon 7. We have recently shown that C6U creates an extended inhibitory context (Exinct) that causes skipping of exon 7 in SMN2. Previous studies have shown that an exonic splicing enhancer associated with Tra2 (Tra2 ESE) is required for exon 7 inclusion in both SMN1 and SMN2. Here we describe the method of in vivo selection that determined the position-specific role of wild type nucleotides within the entire exon 7. Our results confirmed the existence of Exinct and revealed the presence of an additional inhibitory tract (3'-Cluster) near the 3'-end of exon 7. We also demonstrate that a single nucleotide substitution at the last position of exon 7 improves the 5' splice site (ss) such that the presence of inhibitory elements (Exinct as well as the 3'-Cluster) and the absence of Tra2-ESE no longer determined exon 7 usage. Our results suggest that the evolutionary conserved weak 5' ss may serve as a mechanism to regulate exon 7 splicing under different physiological contexts. This is the first report in which a functional selection method has been applied to analyze the entire exon. This method offers unparallel advantage for determining the relative strength of splice sites, as well as for identifying the novel exonic cis elements. PMID- 15272123 TI - Distinct ensemble codes in hippocampal areas CA3 and CA1. AB - The hippocampus has differentiated into an extensively connected recurrent stage (CA3) followed by a feed-forward stage (CA1). We examined the function of this structural differentiation by determining how cell ensembles in rat CA3 and CA1 generate representations of rooms with common spatial elements. In CA3, distinct subsets of pyramidal cells were activated in each room, regardless of the similarity of the testing enclosure. In CA1, the activated populations overlapped, and the overlap increased in similar enclosures. After exposure to a novel room, ensemble activity developed slower in CA3 than CA1, suggesting that the representations emerged independently. PMID- 15272124 TI - Pairing gap and in-gap excitations in trapped fermionic superfluids. AB - We consider trapped atomic Fermi gases with Feshbach-resonance enhanced interactions in pseudogap and superfluid temperatures. We calculate the spectrum of radio-frequency (or laser) excitations for transitions that transfer atoms out of the superfluid state. The spectrum displays the pairing gap and also the contribution of unpaired atoms, that is, in-gap excitations. The results support the conclusion that a superfluid, in which pairing is a manybody effect, was observed in recent experiments on radio-frequency spectroscopy of the pairing gap. PMID- 15272125 TI - Observation of the pairing gap in a strongly interacting Fermi gas. AB - We studied fermionic pairing in an ultracold two-component gas of 6Li atoms by observing an energy gap in the radio-frequency excitation spectra. With control of the two-body interactions through a Feshbach resonance, we demonstrated the dependence of the pairing gap on coupling strength, temperature, and Fermi energy. The appearance of an energy gap with moderate evaporative cooling suggests that our full evaporation brought the strongly interacting system deep into a superfluid state. PMID- 15272126 TI - Comparison of preperfusion and postperfusion magnetic resonance angiography in acute stroke. AB - BACKGROUND AND PURPOSE: The multimodal magnetic resonance imaging study in acute stroke includes perfusion-weighted imaging (PWI) after administration of contrast and magnetic resonance angiography (MRA). However, MRA may overestimate the degree of vessel obstruction caused by limitations to detect low flow states. Our aim was to determine the usefulness of a new fast imaging protocol combining classical MRA, PWI, and postperfusion MRA to improve the diagnostic management in acute ischemic stroke. METHODS: We studied 31 patients with a middle cerebral artery (MCA) infarction within the first 12 hours from the onset of symptoms. All patients had an MCA stenosis or occlusion. The study protocol included a preperfusion MRA and a postperfusion MRA. Modified thrombolysis in myocardial infarction (TIMI) classification was used to assess the patency of vessels. RESULTS: In 17 patients (group A, 55%), preperfusion MRA and postperfusion MRA accorded in the estimation of vascular status, whereas in 14 patients (group B, 45%) postperfusion MRA showed a better vascular flow than preperfusion MRA. The improvement in the depiction of flow was from a complete occlusion (TIMI I) to a partial occlusion (TIMI II) in 9 patients and from TIMI II to normal patency (TIMI III) in 5 patients. Thirty-six percent of the patients with suspected internal carotid artery occlusion in the preperfusion MRA showed flow in the intracranial internal carotid artery in the postperfusion MRA. CONCLUSIONS: Postperfusion contrast-enhanced MRA can demonstrate arterial segments with low flow and avoid overestimation of vascular obstruction. PMID- 15272127 TI - Systemic inflammatory response depends on initial stroke severity but is attenuated by successful thrombolysis. AB - BACKGROUND AND PURPOSE: To determine whether body temperature, c-reactive protein (CRP), and white blood cell (WBC) count within the first days after stroke onset correlate with infarct size and stroke severity, and to examine whether successful thrombolysis reduces poststroke inflammation. METHODS: Out of 1500 consecutive acute ischemic stroke patients, 346 cases (43 patients with thrombolysis) were selected according to the following criteria: admission to hospital < or =24 hours after event, absence of prestroke and poststroke infectious disease, no intracerebral hemorrhage or brain stem stroke, and data availability. Body temperature, WBC within 3 days, and CRP within 5 days of event were determined daily. Lesion volume was measured by planimetry on computed tomography or MRI scans. Successful thrombolysis was defined as improvement on the National Institutes of Health Stroke Scale of > or =4 points within 24 hours. RESULTS: Increase of inflammatory parameters correlated significantly with lesion volume and stroke severity. This was shown for body temperature on days 2 and 3 (P<0.001), CRP on days 1 to 5 (P<0.05), and WBC on days 1 to 3 (P<0.01). Patients with successful thrombolysis had reduced body temperature on day 3, WBC on days 2 and 3, and CRP on days 3 to 5 (P<0.05). CONCLUSIONS: Patients with a larger stroke volume and more severe stroke deficits have higher body temperature, CRP, and WBC count in the acute phase after stroke. Successful thrombolysis is related to a significantly attenuated inflammatory response. PMID- 15272128 TI - Day of the week and ischemic stroke: is it Monday high or Sunday low? AB - BACKGROUND AND PURPOSE: The study aim was to examine the incidence of ischemic stroke (IS) by day of the week and its relationship with age, sex, and socioeconomic status (SES). METHODS: A total of 12,801 IS events in men and women aged 25 to 99 years was recorded in a population-based stroke register (FINMONICA), which was functioning in Finland from 1982 to 1992. We analyzed the weekly variation in IS incidence by pooling the data and stratifying by sex and age. Taxable income and level of education were used as indicators of SES. RESULTS: We observed a significant weekly variation in IS occurrence, but the analysis by age group demonstrated a difference by weekday only in the age group 60 to 74, both in men and women (P<0.001 and P=0.02, respectively). The increase in the number of IS events from Sunday to Monday was pronounced in men (29.2% increase from Sunday to Monday). When stratifying by age, Monday excess in IS incidence was associated with lower SES among persons >59 years of age. No Monday excess was observed in persons with high SES. CONCLUSIONS: Because the incidence of IS is much higher in persons with low SES than in those with high SES, the Monday excess in persons with low SES is of substantial public health interest. This finding may suggest reasons for the higher IS incidence in persons with low socioeconomic positions and open up some possibilities for prevention. PMID- 15272129 TI - Individual patient data meta-analysis of randomized controlled trials of community occupational therapy for stroke patients. AB - BACKGROUND AND PURPOSE: Trials of occupational therapy for stroke patients living in the community have varied in their findings. It is unclear why these discrepancies have occurred. METHODS: Trials were identified from searches of the Cochrane Library and other sources. The primary outcome measure was the Nottingham Extended Activities of Daily Living (NEADL) score at the end of intervention. Secondary outcome measures included the Barthel Index or the Rivermead ADL (Personal ADL), General Health Questionnaire (GHQ), Nottingham Leisure Questionnaire (NLQ), and death. Data were analyzed using linear or logistic regression with a random effect for trial and adjustment for age, gender, baseline dependency, and method of follow-up. Subgroup analyses compared any occupational therapy intervention with control. RESULTS: We included 8 single blind randomized controlled trials incorporating 1143 patients. Occupational therapy was associated with higher NEADL scores at the end of intervention (weighted mean difference [WMD], 1.30 points, 95% confidence intervals [CI], 0.47 to 2.13) and higher leisure scores at the end of intervention (WMD, 1.51 points; 95% CI, 0.24 to 2.79). Occupational therapy emphasizing activities of daily living (ADL) was associated with improved end of intervention NEADL (WMD, 1.61 points; 95% CI, 0.72 to 2.49) and personal activities of daily living (odds ratio [OR], 0.65; 95% CI, 0.46 to 0.91), but not NLQ. Leisure-based occupational therapy improved end of intervention NLQ (WMD, 1.96 points; 95% CI, 0.27 to 3.66) but not NEADL or PADL. CONCLUSIONS: Community occupational therapy significantly improved personal and extended activities of daily living and leisure activity in patients with stroke. Better outcomes were found with targeted interventions. PMID- 15272130 TI - Late measures of brain injury after neonatal hypoxia-ischemia in mice. AB - BACKGROUND AND PURPOSE: This work was undertaken to determine to what degree long term neurofunctional outcome of neonatal hypoxic-ischemic (HI) brain injury in mice correlates with anatomical extent of cerebral damage assessed by magnetic resonance imaging (MRI) and histopathology. METHODS: On postnatal day 7, mice were subjected to HI. At 7 to 9 weeks after HI neurofunctional outcome was assessed by water-maze, rota-rod, and open-field test performance, followed by cerebral MRI and histopathology evaluation. RESULTS: At 10 weeks after HI, MRI revealed ipsilateral brain atrophy alone or with porencephalic cyst formation and contralateral ventriculomegaly. Adult HI-affected mice, especially those that developed a porencephalic cyst, demonstrated significant neurofunctional deficit compared with age-matched naive mice. HI-affected mice with ipsilateral cerebral atrophy but without porencephaly demonstrated no or an intermediate level of neurofunctional deficit. Neurobehavioral assessment of mice subjected to HI insult revealed a strong correlation between degree of brain injury and functional neurohandicap. CONCLUSIONS: This is the first study to demonstrate that long-term neurofunctional outcome in mice after a neonatal HI correlates tightly with anatomical pattern/extent of cerebral damage, defined by MRI and histopathology. PMID- 15272131 TI - Statin therapy after acute ischemic stroke in the heart protection study: is the role in recurrent stroke prevention now defined? PMID- 15272132 TI - Are there time-dependent differences in diffusion and perfusion within the first 6 hours after stroke onset? AB - BACKGROUND AND PURPOSE: Stroke heterogeneity in computed tomography-based studies has been attributed as main cause for missing efficacy of intravenous tissue plasminogen activator (tPA) therapy within 3 to 6 hours. We investigated early time-dependent differences in acute stroke pathophysiology by multiparametric magnetic resonance imaging (MRI). METHODS: Stroke MRI of 112 acute ischemic stroke patients within <6 hours were dichotomized into a <3-hour group (n=52) and a 3- to 6-hour group (n=60). Infarct volume was determined on days 5 to 8. Lesion volumes were determined for apparent diffusion coefficient (ADC_man) and the subregion with ADC values <550x10(-9) mm/s2 (ADC <550), and for the time-to-peak (TTP) delay of 2 to 4 seconds, 4 to 6 seconds, 6 to 8 seconds, and >8 seconds. A subsample analysis was performed for occlusions of the middle carotid artery (MCA) trunk (n=36) and MCA branches (n=30), and for all patients treated by intravenous tPA (n=70). RESULTS: ADC and TTP lesion volumes were not different within <3 hours compared with volumes at 3 to 6 hours. In patients receiving intravenous tPA (n=70), there were no significant differences in ADC_man, TTP >2 seconds, and infarct volume (days 5 to 8) between the 2 groups. There was a greater proportion of ADC <550/ADC_man, which was most pronounced in patients with MCA trunk occlusions after 3 to 6 hours and a larger mismatch in the <3-hour group compared with that of the 3- to 6-hour group. In MCA branch occlusions, there was a less severe TTP delay after 3 to 6 hours. However, all differences missed the significance level (P=0.05) after correction for multiple testing. CONCLUSIONS: We observed no significant time-dependent differences within 6 hours after stroke onset in degree and volume of diffusion and perfusion impairment. An exclusion from intravenous tPA solely based on a rigid 3-hour time window seems unjustified in MRI-confirmed ischemic stroke. PMID- 15272133 TI - Declining mortality from subarachnoid hemorrhage: changes in incidence and case fatality from 1985 through 2000. AB - BACKGROUND AND PURPOSE: Northern Sweden has one of the highest incidence rates of subarachnoid hemorrhage (SAH) among the populations covered by the WHO MONICA (Monitoring of Trends and Determinants in Cardiovascular Disease) Project, approximately twice as high as in the other populations in Europe. In this study, trends in incidence, 28-day case fatality (CF), and mortality in SAH were followed over a 16-year period. METHODS: Since 1985, all SAHs in northern Sweden among patients 25 to 74 years old have been validated using strict MONICA criteria. From 1985 through 2000, 392 men and 592 women had SAH. During 3 years, 1997 to 1999, SAH among those aged 75 and older were also included. RESULTS: The total incidence among those 25 years and older was 13.3 per 100 000 in men and 24.4 per 100 000 in women. During the 16 years of observation, age standardized incidence in the group aged 25 to 74 years decreased significantly in men (P for trend <0.0001) but remained essentially unchanged in women (P for trend=0.64). The 28-day CF for all years was 36.5% in men, with no significant trend (P=0.7). In women, average CF was 35%, with a significant decline (P=0.003). The annual mortality decreased significantly in both sexes (by 3.87 [95% CI+/-2.75 percentage points] in men and 3.97 [95% CI+/-2.29] in women). CONCLUSIONS: The decline in SAH mortality has different explanations in men (declining incidence) and in women (declining CF). PMID- 15272134 TI - Cardiac autonomic derangement and arrhythmias in right-sided stroke with insular involvement. AB - BACKGROUND AND PURPOSE: The insula of the right cerebral hemisphere may have a major role in cardiac autonomic control. This study was aimed at assessing the effects of acute right insular ischemic damage on heart rate variability (HRV) and arrhythmias. METHODS: Holter monitoring for 24 hours was performed in 103 consecutive patients with first-ever acute ischemic stroke. Time and frequency domain measures of HRV and arrhythmias were considered in all cases. RESULTS: Forty-nine patients (47.5%) had a right-sided infarction, whereas 54 (52.5%) had a left-sided infarction. Insular involvement was present in 33 patients with right-sided stroke (67.3%) and in 36 patients with left-sided stroke (66.6%). When compared with all other stroke patients, subjects with right-sided insular damage showed significantly lower values of the standard deviation of all normal to-normal (SDNN) R wave to R wave (RR) intervals and of the root mean square of differences (rMSSD) of adjacent normal-to-normal RR intervals, and higher low frequency/high-frequency ratio values (P<0.05). Right insular stroke was also associated with more complex arrhythmias than any other localization (P<0.05). Moreover, in the whole population of stroke patients, lower values of SDNN were associated with the presence of more frequent and complex arrhythmias. CONCLUSIONS: These findings further support the notion that the right insula is implicated in the autonomic control of cardiac activity and that acute right insular damage may lead to a derangement of cardiac function with potential prognostic implications. PMID- 15272136 TI - Metabolic rate constants for hydroquinone in F344 rat and human liver isolated hepatocytes: application to a PBPK model. AB - Hydroquinone (HQ) is an important industrial chemical that also occurs naturally in foods and in the leaves and bark of a number of plant species. Exposure of laboratory animals to HQ may result in species-, sex-, and strain-specific nephrotoxicity. The sensitivity of male F344 versus female F344 and Sprague Dawley rats or B6C3F1 mice appears to be related to differences in the rates of formation of key nephrotoxic metabolites. Metabolic rate constants for the conversion of HQ through several metabolic steps to the mono-glutathione conjugate and subsequent detoxification via mercapturic acid formation were measured in suspension cultures of hepatocytes isolated from male F-344 rats and humans. A mathematic kinetic model was used to analyze each metabolic step by simultaneously fitting the disappearance of each substrate and the appearance of subsequent metabolites. An iterative, nested approach was used whereby downstream metabolites were considered first, and the model was constrained by the requirement that rate constants determined during analysis of individual steps must also satisfy the complete, integrated metabolism scheme, including competitive pathways. The results from this study indicated that the overall capacity for metabolism of HQ and its mono-glutathione conjugate is greater in hepatocytes from humans than in those from rats, suggesting a greater capacity for detoxification of the glutathione conjugates in humans. Metabolic rate constants were applied to an existing physiologically based pharmacokinetic model, which was used to predict total glutathione metabolites produced in the liver. The results showed that body burdens of these metabolites will be much higher in rats than in humans. PMID- 15272135 TI - A critical role for MAP kinases in the control of Ah receptor complex activity. AB - The heterodimeric complex of aromatic hydrocarbon receptor (AHR) and Ah receptor nuclear translocator (ARNT) plays a pivotal role in controlling the expression of drug metabolism genes, such as the cytochromes p450 (Cyp) 1a1 and 1b1, believed to be responsible for most toxic effects of the environmental contaminant 2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD). In this study, we show that activation of Jun N-terminal kinase (JNK) and extracellular signal-regulated kinase (ERK) modulates ARNT transcription activity and potentiates the transcriptional activity of AHR/ARNT complexes. Inhibition of ERK by chemical compounds and ablation of JNK caused significant decreases in CYP1A1 induction by TCDD. Compared to wild type, JNK2 ablation significantly reduced TCDD-stimulated CYP1A1 expression in mouse thymus and testis, but not in liver. In contrast, CYP1B1 expression was unaffected in all three tissues of the knockout mice. These data suggest that JNK and ERK modulate ARNT activity and AHR/ARNT-dependent gene expression, contributing to the gene-specific and tissue-specific toxicity of environmental contaminants. PMID- 15272137 TI - Evaluation of the developmental and reproductive toxicity of methoxychlor using an anuran (Xenopus tropicalis) chronic exposure model. AB - The chronic toxicity of methoxychlor to the South African clawed frog, Xenopus (Silurana) tropicalis, was evaluated using a life cycle approach. The chronic exposure period ranged from mid-cell blastula stage [NF (Nieuwkoop and Faber, 1994) stage 8] to 90 days of exposure, during which time the organisms generally completed metamorphosis and emerged as juvenile frogs. Methoxychlor concentrations ranged from 1 to 100 micrograms/l. Methoxychlor concentrations >10 micrograms/l caused delayed development. Organisms exposed to 10 micrograms/l methoxychlor for 30 days showed enlarged thyroid glands with follicular hyperplasia. No increase in mortality or external malformation was observed at any of the test concentrations during early embryo-larval development (NF stage 8 to NF stage 46; ca. 2 days exposure). A concentration-dependent increase in external malformations and internal abnormalities of the liver and gonads were noted after 90 days of exposure, however. Skewing of the sex ratio toward the female gender decreased ovary weight and number of oocytes, and increased oocyte immaturity and necrosis were noted at methoxychlor concentrations of 100 micrograms/l. Reductions in testis weight and sperm cell count were also detected at 100 micrograms/l methoxychlor. Results from these studies suggested that methoxychlor was capable of altering the rate of larval development, but did not adversely affect early embryo-larval development (2 days of exposure) as manifested in external malformations. Internal malformations, increases in the ratio of phenotypic females, were induced by chronic methoxychlor exposure. In addition, reproductive endpoints, most notably in the female specimens, were adversely affected by methoxychlor exposure. These studies add to the standardization and validation of a useful amphibian test methods capable of evaluating both reproductive and developmental effects of potential endocrine disrupting chemicals over a life cycle exposure. PMID- 15272138 TI - Blood pressure in acute stroke and its prognostic value. PMID- 15272139 TI - Preventing cancer, cardiovascular disease, and diabetes: a common agenda for the American Cancer Society, the American Diabetes Association, and the American Heart Association. AB - Collectively, cardiovascular disease (including stroke), cancer, and diabetes account for approximately two thirds of all deaths in the United States and about 700 billion dollars in direct and indirect economic costs each year. Current approaches to health promotion and prevention of cardiovascular disease, cancer, and diabetes do not approach the potential of the existing state of knowledge. A concerted effort to increase application of public health and clinical interventions of known efficacy to reduce prevalence of tobacco use, poor diet, and insufficient physical activity-the major risk factors for these diseases-and to increase utilization of screening tests for their early detection could substantially reduce the human and economic cost of these diseases. In this article, the ACS, ADA, and AHA review strategies for the prevention and early detection of cancer, cardiovascular disease, and diabetes, as the beginning of a new collaboration among the three organizations. The goal of this joint venture is to stimulate substantial improvements in primary prevention and early detection through collaboration between key organizations, greater public awareness about healthy lifestyles, legislative action that results in more funding for and access to primary prevention programs and research, and reconsideration of the concept of the periodic medical checkup as an effective platform for prevention, early detection, and treatment. PMID- 15272140 TI - The predictive value of cortical activation by passive movement for motor recovery in stroke patients. AB - PURPOSE: Contralateral primary sensori-motor cortex (SM1) activation by passive movement was investigated by functional MRI (fMRI) at the early stage of stroke, to determine whether SM1 activation can be used to predict the degree of motor recovery of the hemiplegic hand. METHODS: We studied 17 stroke patients who showed complete paralysis of a hemiplegic hand at onset. The motor function of the hemiplegic hand was assessed on 4 separate occasions (at onset, at fMRI evaluation (performed < 4 weeks after onset), and 3 and 6 months after onset). Significant motor recovery was defined as recovery of the affected hand to the extent of it being able to prehend an object against gravity at least at 6 months after onset. RESULTS: The patients having an activated contralateral SM1 showed better motor recovery than those who did not. Only a fourth of the patients with an activated contralateral SM1 experienced a significant motor recovery, whereas none of the patients with an inactivated SM1 showed an improvement 6 months after onset, however, the incidence of significant motor recovery was not significantly difference between the two groups. CONCLUSION: It appears that contralateral SM1 activation by passive movement in the early stage of stroke has a low predictive value for the motor recovery of the hemiplegic hand, because the activation of the contralateral SM1 by passive movement appears to be mediated by somatosensory input to the cortex from the thalamus rather than from the motor pathway. PMID- 15272141 TI - Bridging short nerve defects by direct repair under tension, nerve grafts or longitudinal sutures. AB - PURPOSE: To compare the longitudinal suture model for bridging nerve defects with direct approximation under tension or with autologuos nerve grafting. METHODS: Seven mm nerve defects in the rat sciatic nerve were repaired by either of these three methods. Evaluation was performed at twelve weeks by morphometry of the tibial nerve distal to the repair site and by weight of the gastrocnemius muscle, an indicator of target reinnervation. RESULTS: The number of nerve fibers and myelin areas in the tibial nerve were similar for all repair methods as were the weight of the gastrocnemius muscle. CONCLUSIONS: Longitudinal sutures can be used to bridge short nerve defects and could be an alternative to nerve grafting. PMID- 15272142 TI - Inhibition of vascular endothelial growth factor receptor (VEGFR) signaling by BSF476921 attenuates regional cerebral edema following traumatic brain injury in rats. AB - PURPOSE: In the present study we assessed the ability of BSF476921, an inhibitor of vascular endothelial growth factor receptor (VEGFR) kinase signal transduction, to reduce edema formation and neurologic motor dysfunction following lateral fluid percussion (FP) brain injury in rats. METHODS: Anesthetized adult male rats were subjected to either lateral FP brain injury of moderate severity (n = 37) or sham injury (n = 22, surgery without brain injury). Animals were randomized to receive i.p. injections of either BSF476921 (30 mg/kg bw; injured n = 15, sham n = 11) or sterile water (injured n = 14, sham n = 11) at 1, 11 and 22 hours post-injury. After assessment of motor function using a standard 28-point neuroscore, animals were sacrificed 24 hours following trauma and their brains evaluated for regional water content using the wet-weight/dry weight technique. RESULTS: Although brain-injured animals showed a significant motor deficit compared to uninjured animals, no differences were detected between BSF476921- and vehicle-treated animals at the acute 24 hour post-injury time point. However, BSF476921 significantly attenuated regional edema formation in brain-injured animals in the ipsilateral hippocampus (p < 0.05) and in the cortex adjacent to the injury (p < 0.05) when compared to vehicle treatment. CONCLUSIONS: To our knowledge, this is the first report of a small molecule VEGFR kinase inhibitor reducing cerebral edema in a widely accepted model of brain injury. PMID- 15272143 TI - Connection between p53 gene Bam HI RFLP polymorphism with the volume of brain infarction in patients with carotid atherothrombotic ischemic stroke. AB - PURPOSE: The aim of our investigation was to study the connection between p53 gene Bam HI RFLP polymorphism and the brain infarction volume in patients with atherothrombotic ischemic stroke that could highlight certain genetic aspects of the individual sensibility of brain tissue to acute ischemia. MATERIALS AND METHODS: Diallelic Bam HI RFLP polymorphism in 5' flanking region p53 gene was studied in 96 patients with carotid atherothrombotic stroke from Moscow population. Magnetic resonance imaging was conducted on day 7 after the stroke onset. The manual morphometry and "Osiris" morphometric hardware (by the Hospital of the University of Geneva) were used for assessment of the infarction volume. RESULTS: The predominance of small-size infarctions (< 40 cm3) was revealed in patients with (-/-) Bam HI RFLP p53 genotype versus patients with (-/+) (X2 = 19.7; P < 0.001) and (+/+) (X2 = 12.288; P < 0.001) genotypes. According to the Bayesian's statistics, in patients with (-/-) p53 Bam HI genotype the development of a small-size infarction in atherothrombotic ischemic stroke can be prognosticated with probability more than 65%. CONCLUSIONS: A significant association between p53 gene Bam HI RFLP polymorphism and the infarction volume was found in patients with carotid atherothrombotic stroke from Moscow population. These results additionally confirm that apoptosis plays an important role in the formation of ischemic brain lesion and that drugs with anti-apoptotic properties may prove beneficial in stroke patients. PMID- 15272144 TI - Insights into oxidative stress and potential novel therapeutic targets for Alzheimer disease. AB - PURPOSE: Since the description of his original patient Auguste D. with cognitive disability, Alois Alzheimer persisted in his clinical investigations to understand this complex disease. Although more than a century later the underlying cellular dysfunctions that can initiate and determine the course of this neurodegenerative disease remain evasive, significant strides continue to elucidate the complex nature of Alzheimer dementia and define potential effective strategies for its prevention and treatment. METHODS AND RESULTS: In this article, we examine the cellular mechanisms that define Alzheimer disease. They are diverse in nature and involve pathways of oxidative stress that extend well beyond the pathological hallmarks of beta-amyloid aggregates and neurofibrillary tangles. Oxidative stress precipitates both nuclear DNA degradation and membrane phosphatidylserine exposure in neuronal and vascular cells to promote loss of cellular integrity, microglial phagocytosis, and thrombotic destruction. Critical in the ability to foster cell survival during oxidative stress is the modulation of the metabotropic glutamate system, cell cycle regulation in post-mitotic neurons, and control of GSK-3beta activity and presenilin integrity. These cellular pathways ultimately converge upon more central cellular mechanisms that involve maintenance of mitochondrial membrane permeability through Bcl-2 family members, trophic factors, and mitochondrial energy reserves. CONCLUSIONS: By targeting critical elements that determine neuronal and vascular survival during Alzheimer disease, successful development of clinical applications can emerge for the treatment of neurodegenerative disorders. PMID- 15272145 TI - Erythropoietin as a novel neuroprotectant. AB - PURPOSE: To provide an overview of the current knowledge on neuroprotective properties of Erythropoietin (Epo), mechanisms by which Epo produces neuroprotection, and signaling pathways regulated by Epo in the nervous system. METHODS: The Medline database was searched for articles on the neuroprotective properties of Epo. Experimental and clinical studies were systematically reviewed. RESULTS: In addition to promoting the survival, proliferation, and differentiation of immature erythroid cells, Epo and the Epo receptor (EpoR) have recently been shown to exist and function in the nervous system. The Epo/EpoR system plays a critical role in neurodevelopment and neuroprotection. Epo ameliorates or prevents neuronal injury by neuroprotective, anti-apoptotic, anti inflammatory, anti-oxidant, angiogenic, neurogenic and neurotrophic effects in cell culture and animal models of neurological diseases. The clinical effectiveness of recombinant human Epo in ischemic stroke in human patients has also been reported recently. CONCLUSION: Recent studies suggest that Epo is a potential novel neurotherapeutic agent and further clinical studies are warranted. PMID- 15272146 TI - Role of brain-derived neurotrophic factor and NF-kappaB in neuronal plasticity and survival: From genes to phenotype. AB - PURPOSE: Brain-derived neurotrophic factor (BDNF) is a member of the family of neurotrophins and promotes diverse effects in neurons including development, maintenance of function, synaptic plasticity, and survival in different animal models. We present advances in our understanding of the genomics of the BDNF gene (bdnf) and its regulation by calcium-activated transcription factors, including cAMP response element binding protein (CREB) and more recently, nuclear factor kappaB (NF-kappaB) and discuss these findings in the context of neuronal plasticity and survival. METHODS: We used amplified bdnf complementary DNAs (cDNAs) and genomic DNA templates for direct sequencing and sequence variant discovery, information mining of public databases, and conventional molecular and cellular biology approaches to screen bdnf for novel regulatory elements, alternatively spliced exons, and functional sequence variants. RESULTS: We discovered a candidate NF-kappaB site in promoter 3 of bdnf and showned that activation of N-methyl-D-aspartate (NMDA) inotropic glutamate receptors increased bdnf expression through an NF-kappaB-dependent pathway and extended the finding to show that NF-kappaB was required for NMDA neuroprotection in vitro. In addition, sequence analysis of bdnf cDNAs from different brain regions predicted at least three pre-pro-BDNF protein isoforms, two of which were previously unknown. Each isoform differs at the amino terminus and may have functional importance. CONCLUSIONS: Given the central role that BDNF plays in the developing and adult nervous system, understanding how BDNF is regulated and how it functions will enhance our knowledge of its diverse effects, which may lead to more effective treatments for neurodegenerative disorders and reveal the role of BDNF in complex phenotypes related to behavior. PMID- 15272147 TI - APP, NGF & the 'Sunday-driver' in a Trolley on the Road. AB - Alzheimer's disease (AD) is one of the most common and challenging neurodegenerative diseases in humans and is characterized by: progressive impairment in cognitive function, degeneration of cholinergic neurons of the basal forebrain (CBF), neurofibrillary tangles and amyloid beta-peptide (Abeta) depositions. The amyloid precursor protein (APP) is a transmembrane protein of which abnormal processing produces Abeta that is associated with the pathogenesis of AD. Neurotrophic factors have attracted much attention for their potential as a remedy for neurological disorders. In this regard, nerve growth factor (NGF) has generated a great interest as a potential target for the treatment of AD. This interest is based on the observation that CBF neurons, which provide the major source of cholinergic innervation to the cerebral cortex and hippocampus, undergo selective and severe degeneration in advanced AD and that the survival of CBF neurons depends upon NGF and its receptors, namely, trkA and p75NTR. This review focuses on recent findings about APP, NGF and their potential signaling connections to the protein encoded by the 'Sunday-driver' (SYD) gene. PMID- 15272148 TI - Hemorheological disturbances in patients with chronic cerebrovascular diseases. AB - Hemorheological disturbances may occur in more than 40% of patients with ischemic cerebrovascular diseases. In this study the changes of rheological factors- hematocrit, plasma fibrinogen concentration, whole blood and plasma viscosity, red blood cell aggregation and deformability were investigated in 297 patients (173 males, 124 females, mean age 60 +/- 11 years) with transient ischemic attack or chronic phase (> 3 months after onset) ischemic stroke, and in 73 healthy volunteers (35 males, 38 females, mean age 38 +/- 7 years). Hematocrit, plasma and whole blood viscosity were significantly (p < 0.0001) elevated in cerebrovascular patients compared to controls. Plasma fibrinogen concentration (p < 0.001), red blood cell aggregation (p < 0.05) and deformability (p < 0.01) were also impaired in stroke patients. Hemorheological disturbances were dominant in stroke patients with diabetes, hyperlipidemia and smoking habits. Hematocrit, plasma viscosity and red blood cell aggregation showed a significant (p < 0.025 0.001) correlation with the severity of carotid artery stenosis. We could not find any characteristic distribution of rheological parameters among the three subtypes of brain ischemia. Our results show that all of the measured rheological parameters are significantly impaired in chronic ischemic cerebrovascular disorders, especially in diabetic, smoking and alcoholic patients. They correlate with the severity of the carotid artery stenosis, but there is no association with the type of ischemic stroke. PMID- 15272149 TI - Post-occlusive reactive hyperemia in patients with peripheral vascular disease. AB - We measured tissue oxygen saturation (StO2) changes during a post-occlusive reactive hyperemia (PORH) test on lower extremities of peripheral vascular disease (PVD) patients and healthy controls. The StO2 changes were measured by the near-infrared tissue oximeter prototyped by ViOptix Inc. We found that PVD indicators may include the post-occlusive StO2 reperfusion rate, the time duration of the reperfusion phase in a PORH test, and the StO2 dispersion rate in reactive hyperemia. We suggest critical values of these indicators for PVD assessments. PMID- 15272150 TI - Analyzing shear stress-elongation index curves: comparison of two approaches to simplify data presentation. AB - Ektacytometry is frequently used to study red blood cell (RBC) deformability. This method is capable of measuring RBC deformation under a wide range of shear stresses, and the results are usually presented as shear stress-elongation index curves. These curves are very useful for detailed analyses of RBC mechanical behavior under various shearing conditions, yet may not be appropriate for clinical and experimental studies where a global parameter of deformability is satisfactory. That is, presenting data at a selected shear stress may not always be appropriate, since the selected stress level may not accurately reflect variations of the whole curve. We have thus compared two approaches to calculate parameters that represent the entire shear stress-elongation index curve; data were obtained using a commercial laser diffraction ektacytometer (LORCA) and compared in terms of their power to detect a difference between groups. Usage of these parameters (i.e., shear stress at half maximal deformation and maximal deformation) appears to offer a simplified approach to data presentation and interpretation in clinical and experimental hemorheological studies. PMID- 15272151 TI - In vivo imaging of the rat cerebral microvessels with optical coherence tomography. AB - A technique called optical coherence tomography (OCT) was applied to in vivo observation of microcirculation in the rat cerebral cortex. The OCT system used in this study provided cross-sectional images of the cerebral cortical tissue up to about 1 mm depth with longitudinal resolution up to 8 microm. It could visualize cross-sectional structure of the dura, arachnoid membrane, cortical tissue, and pial microvessels through the cranial window. Pial microvessels with diameter larger than several 10 microm could be detected to observe their cross sectional shape, while the microvessels within the cortical tissue with smaller diameter were not discernible. The OCT observation revealed that the pial microvessels showed different spatial configurations depending on the cerebral preparations with intact dura and without dura. Stimulus responses of the somatosensory cortices were also different among the preparation methods; Delayed swelling of the cortical surface appeared in the somatosensory cortex following the electrical stimulation of the hind paw in the case of dura removal, which was restricted to a thin surface layer with less than several 10 microm. It is considered to reflect the reactive hyperemia accompanying the neuronal activation. Doppler frequency shift due to the blood flow was detected in pial arterioles. This phenomenon is promising to provide the velocity profile within microvessels and may be applicable to the functional imaging of the brain. PMID- 15272152 TI - Polymorphonuclear leukocyte membrane fluidity and cytosolic Ca(2+) content in young adults with acute myocardial infarction. Evaluation at the initial stage and after 12 months. AB - Our aim was to examine two aspects of polymorphonuclear leukocyte (PMN) rheology (membrane fluidity and cytosolic Ca2+ content), at baseline and after in vitro activation, in a group of young adults with acute myocardial infarction (AMI) at the initial stage and after 12 months. We enrolled 21 AMI subjects aged < or = 45 years (mean age 41.1 +/- 3.5 years) and evaluated PMN membrane fluidity, labelling intact PMN cells with the fluorescent probe 1,4-(trimethylamino)-phenyl 4-phenylhexatriene and the PMN cytosolic Ca2+ content marking PMN cells with the fluorescent probe Fura 2-AM, at baseline and after in vitro activation with 4 phorbol 12-myristate 13-acetate (PMA) and N-formyl-methionyl-leucyl-phenylalanine (fMLP). During the initial stage PMN membrane fluidity and cytosolic Ca2+ content did not distinguish AMI patients from control subjects; after 12 months, when compared with the initial stage, PMN cytosolic Ca2+ content was significantly increased. In vitro PMN activation with PMA and fMLP caused no variation of the two PMN parameters in control subjects, while in AMI patients membrane fluidity decreased and cytosolic Ca2+ content increased; the same behaviour pattern was observed after 12 months. The constant functional alteration of PMN cells in young AMI patients highlights the role of activated leukocytes as a component of the inflammatory reaction that follows ischemia. PMID- 15272153 TI - Hemorheological changes in women with severe preeclampsia. AB - The objective of this study was to determine the differences of hemorheological parameters in association to haematological tests in patients with severe preeclampsia (blood pressure (diastolic) > 100 mm Hg, blood pressure (systolic) > 180 mm Hg and proteinuria > 3 g/24 h). Blood samples of 45 primigravidas by hospital admission were studied. The control group were 45 pregnant women--age and weight matched--with normal blood pressure and without obstetric complications. We measured red cell aggregation (stasis, low shear), red cell elongation with the ectacytometer, blood cell indices (Hct, Hbg, MCV, MCHC, reticulocytes, white cells, platelets), fibrinogen haptoglobin and factor VIIIR:Ag, cholesterol and triclycerides. In comparison between patients with severe preeclampsia and normal pregnant women we found statistically elevated values of hematocrit, hemoglobin, red cell aggregation (stasis, low shear rate), MVC and factor VIIIR:Ag. Non-significant changes were observed in values of plasma viscosity, white cells, platelets, haptoglobin, MCHC, reticulocytes, triglycerides and cholesterol. The red cell deformability measured as cell elongation was statistically reduced by high shear stress application in patients with severe preeclampsia. Our results suggest that hemorheological parameters play an important role in severe preeclampsia, especially at microcirculatory regions with high shear stress such as intervillous space of placenta. PMID- 15272154 TI - Preventive mechanism of genistein on coronary endothelial dysfunction in ovariectomized rats: an isolated arrested heart model. AB - The effects of genistein on coronary endothelial dysfunction in bilateral ovariectomized rats were examined. Female Wistar rats were subjected to a bilateral ovariectomy (OVX rat). The animals were divided into three groups: sham treated with vehicle (DMSO 100 microl/day, Sham-DMSO), OVX treated with vehicle (DMSO 100 microl/day, OVX-DMSO), and OVX treated with genistein (0.25 mg/kgBW/day, OVX-genistein). Mean arterial pressure (MAP), heart rate (HR), body weight (BW), uterine weight and plasma E2 were monitored at 4- and 10-week after the treatment. We investigated the endothelium-dependent and -independent vasorelaxation by using acetylcholine (Ach 10(-5) M) and sodium nitroprusside (SNP 10(-7) M), respectively. The experimental results indicated that the uterine weights of all OVX rats were significantly decreased as compared to the sham groups. HR and MAP of both OVX-DMSO and OVX-genistein on 4 and 10 weeks were no significantly increased as compared to the Sham groups. The present coronary vasodilatation responses demonstrated only the significant decrement of endothelium-dependent, not for endothelium-independent, in OVX rats. The treatment of genistein could significantly attenuate this abnormality (% changes of vessel diameter obtained after Ach 10(-6) M: Sham-DMSO(10-wk) = 10.96 +/- 1.2%, OVX-DMSO(10-wk) = 3.2 +/- 0.77%, OVX-genistein(10-wk) = 11.45 +/- 1.85%), (% changes of vessels diameter obtained after SNP 10(-7)M: Sham-DMSO(10-wk) = 16.05 +/- 2.82%, OVX-DMSO(10-wk) = 12.73 +/- 2.72%, OVX-genistein(10-wk) = 16.4 +/- 4.71%) (p < 0.05). However, the lipid profiles monitored from all groups of 4 and 10 weeks did not demonstrate any significant changes. Therefore, it implied that endothelial dysfunction was not primarily cause by the lipid profiles changing in ovariectomized rats. Moreover, such effects of estrogen lacking on coronary endothelial-dependent vasodilatation could be attenuated by genistein supplementation. The present findings suggest that genistein might be used as an therapeutic agent for preventing the menopausal vascular complications. PMID- 15272155 TI - Chronic venous diseases: roles of various pathophysiological factors. AB - Disturbances in haemodynamic, biochemical and enzymatic factors have been observed in chronic venous diseases (CVD). These changes lead to the development of varices, telangiectasies and skin disorders. They affect vessels, blood, skin tissues and cells. It is now possible to describe their time course and interdependance of these changes. Orthostatism pressure on vein wall may lead to fluid leakage and oedema, these resulting in vein enlargement. These processes may be further influenced by genetic or acquired risk factors. Skin microvessels suffer more from hypoxia than from hypertension. Indeed, hypoxia affects not only endothelial cells, but also red and white blood cells and modifies particularly, but not exclusively, TGF-beta1 production. This substance is, an important modulator of zinc dependent-metallo-proteinases and their tissue inhibitor of metallo-proteinases (TIMP) in the skin. Imbalance in this enzymatic system seems to lead either to sclerosis or ulcer. Of course, other biochemical events (also in this review) play a role in vessel wall and skin deterioration in CVD. The aim of the present review is to assess the role of pathophysiological factors in CVD and the influence of different therapies, including the venotropic agent calcium dobesilate, on some of these haemodynamic or biochemical aspects. PMID- 15272156 TI - The crystallographic structure of the aldose reductase-IDD552 complex shows direct proton donation from tyrosine 48. AB - The X-ray crystal structure of human aldose reductase (ALR2) in complex with the inhibitor IDD552 was determined using crystals obtained from two crystallization conditions with different pH values (pH 5 and 8). In both structures the charged carboxylic head of the inhibitor binds to the active site, making hydrogen-bond interactions with His110 and Tyr48 and electrostatic interactions with NADP+. There is an important difference between the two structures: the observation of a double conformation of the carboxylic acid moiety of the inhibitor at pH 8, with one water molecule interacting with the main configuration. This is the first time that a water molecule has been observed deep inside the ALR2 active site. Furthermore, in the configuration with the lower occupancy factor the difference electron-density map shows a clear peak (2.5sigma) for the H atom in the hydrogen bond between the inhibitor's carboxylic acid and the Tyr48 side-chain O atom. The position of this peak implies that this H atom is shared between both O atoms, indicating possible direct proton transfer from this residue to the inhibitor. This fact agrees with the model of the catalytic mechanism, in which the proton is donated by the Tyr48 hydroxyl to the substrate. These observations are useful both in drug design and in understanding the ALR2 mechanism. PMID- 15272157 TI - PRODRG: a tool for high-throughput crystallography of protein-ligand complexes. AB - The small-molecule topology generator PRODRG is described, which takes input from existing coordinates or various two-dimensional formats and automatically generates coordinates and molecular topologies suitable for X-ray refinement of protein-ligand complexes. Test results are described for automatic generation of topologies followed by energy minimization for a subset of compounds from the Cambridge Structural Database, which shows that, within the limits of the empirical GROMOS87 force field used, structures with good geometries are generated. X-ray refinement in X-PLOR/CNS, REFMAC and SHELX using PRODRG generated topologies produces results comparable to refinement with topologies from the standard libraries. However, tests with distorted starting coordinates show that PRODRG topologies perform better, both in terms of ligand geometry and of crystallographic R factors. PMID- 15272158 TI - A neutron crystallographic analysis of a rubredoxin mutant at 1.6 A resolution. AB - A neutron diffraction study has been carried out at 1.6 A resolution on a mutant rubredoxin from Pyrococcus furiosus using the BIX-3 single-crystal diffractometer at the JRR-3 reactor of the Japan Atomic Energy Research Institute. In order to study the unusual thermostability of rubredoxin from P. furiosus (an organism that grows optimally at 373 K), the hydrogen-bonding patterns were compared between the wild-type protein and a 'triple-mutant' variant. In this mutant protein, three residues were changed (Trp3-->Tyr3, Ile23-->Val23, Leu32-->Ile32) so that they are identical to those in a mesophilic rubredoxin from Clostridium pasteurianum. In the present study, some minor changes were found between the wild-type and mutant proteins in the hydrogen-bonding patterns of the Trp3/Tyr3 region. In this investigation, the H/D-exchange ratios in the protein were also studied. Because the target protein was soaked in D2O during the crystallization procedure, most of the N-H and O-H bonds have become deuterated, while essentially all of the C-H bonds have not. In particular, the H/D-exchange pattern of the N-H amide bonds of the protein backbone is of interest because it may contain some indirect information about the mechanism of unfolding of this small protein. The results are in broad agreement with those from solution NMR studies, which suggest that the backbone amide bonds near the four Cys residues of the FeS4 redox center are most resistant to H/D exchange. Finally, the detailed geometries of the water molecules of hydration around the rubredoxin molecule are also reported. The 1.6 A resolution of the present neutron structure determination has revealed a more detailed picture than previously available of some portions of the water structure, including ordered and disordered O-D bonds. Crystallographic details: space group P2(1)2(1)2(1) (orthorhombic), unit-cell parameters a = 34.48, b = 35.70, c = 43.16 A; final agreement factors R = 0.196 and Rfree = 0.230 for 19,384 observed and 6548 unique neutron reflections collected at room temperature; crystal size 4 mm3; a total of 423 non-H atoms, 290 H atoms and 88 D atoms were located in this study. PMID- 15272159 TI - The three-dimensional structure of catalase from Enterococcus faecalis. AB - Enterococcus faecalis haem catalase was crystallized using lithium sulfate at neutral pH. The crystals belong to space group R3, with unit-cell parameters a = b = 236.9, c = 198.1 A. The three-dimensional structure was determined by molecular replacement using a subunit of the Proteus mirabilis catalase structure. It was refined against 2.3 A synchrotron data to a free R factor of 21.8%. Like other catalases, the E. faecalis catalase is a homotetramer with a fold and structure similar to those of its structurally closest relative P. mirabilis. The solvent structure in the active site is identical in the four subunits but differs from that found in other catalases. The structural consequences of the Ramachandran outlier Ser196 are discussed. PMID- 15272160 TI - Structures of the hyperthermophilic chromosomal protein Sac7d in complex with DNA decamers. AB - The protein Sac7d belongs to a class of small chromosomal proteins from the hyperthermophilic archaeon Sulfolobus acidocaldarius. Two new crystal forms of Sac7d in complex with the DNA decamers CCTATATAGG and CCTACGTAGG were obtained and their structures were determined by molecular replacement. The refined models yielded R/Rfree values of 0.221/0.257 and 0.248/0.290 at 1.9 and 2.2 A resolution, respectively. The protein structures are similar to the previously determined structure of Sac7d-GCGATCGC (PDB code 1azp), but the DNA molecules are more bent overall, by 14-20 degrees. The relative positions of the Sac7d protein and the bound DNA also differ by rotations of 6-10 degrees and translations of 1.0-2.4 A. In addition to the water molecules in the central cavity, three additional conserved water molecules are found that mediate the protein-DNA interactions. The decamer DNA fragments form virtual double helices in the crystal, with a unit length of eight base pairs. The molecular packing of the new crystal forms differs from that of 1azp. The terminal nucleotides are opened up and form triple base pairs with other DNA molecules. Through lattice contacts, the Sac7d molecule also makes additional interactions with DNA, whereas only limited protein-protein interactions are seen. PMID- 15272161 TI - The role of substrate-binding groups in the mechanism of aspartate-beta semialdehyde dehydrogenase. AB - The reversible dephosphorylation of beta-aspartyl phosphate to L-aspartate-beta semialdehyde (ASA) in the aspartate biosynthetic pathway is catalyzed by aspartate-beta-semialdehyde dehydrogenase (ASADH). The product of this reaction is a key intermediate in the biosynthesis of diaminopimelic acid, an integral component of bacterial cell walls and a metabolic precursor of lysine and also a precursor in the biosynthesis of threonine, isoleucine and methionine. The structures of selected Haemophilus influenzae ASADH mutants were determined in order to evaluate the residues that are proposed to interact with the substrates ASA or phosphate. The substrate Km values are not altered by replacement of either an active-site arginine (Arg270) with a lysine or a putative phosphate binding group (Lys246) with an arginine. However, the interaction of phosphate with the enzyme is adversely affected by replacement of Arg103 with lysine and is significantly altered when a neutral leucine is substituted at this position. A conservative Glu243 to aspartate mutant does not alter either ASA or phosphate binding, but instead results in an eightfold increase in the Km for the coenzyme NADP. Each of the mutations is shown to cause specific subtle active-site structural alterations and each of these changes results in decreases in catalytic efficiency ranging from significant (approximately 3% native activity) to substantial (<0.1% native activity). PMID- 15272162 TI - Structure, crystal packing and molecular dynamics of the calponin-homology domain of Schizosaccharomyces pombe Rng2. AB - Schizosaccharomyces pombe Rng2 is an IQGAP protein that is essential for the assembly of an actomyosin ring during cytokinesis. Rng2 contains an amino terminal calponin-homology (CH) domain, 11 IQ repeats and a RasGAP-homology domain. CH domains are known mainly for their ability to bind F-actin, although they have other ligands in vivo and there are only few examples of actin-binding single CH domains. The structures of several CH domains have already been reported, but this is only the third report of an actin-binding protein that contains a single CH domain (the structures of calponin and EB1 have been reported previously). The 2.21 A resolution crystal structure of the amino terminal 190 residues of Rng2 from Br- and Hg-derivatives includes 40 residues (150-190) carboxyl-terminal to the CH domain that resemble neither the extended conformation seen in utrophin, nor the compact conformation seen in fimbrin, although residues 154-160 form an unstructured coil which adopts a substructure similar to dystrophin residues 240-246 in the carboxyl-terminal portion of the CH2 domain. This region wraps around the stretch of residues that would be equivalent to the proposed actin-binding site ABS1 and ABS2 from dystrophin. This distinctive feature is absent from previously published CH-domain structures. Another feature revealed by comparing the two derivatives is the presence of two loop conformations between Tyr92 and Arg99. PMID- 15272163 TI - Two orthorhombic crystal structures of a galactose-specific lectin from Artocarpus hirsuta in complex with methyl-alpha-D-galactose. AB - Based on their carbohydrate specificity, the jacalin family of lectins can be divided into two groups: galactose-specific and mannose-specific. The former are cytoplasmic proteins, whereas the latter are localized in the storage vacuoles of cells. It has been proposed that the post-translational modification in some of the lectins that splits their polypeptide chains into two may be crucial for galactose specificity. The mannose-specific members of the family are single chain proteins that lack the above modification. Although the galactose-specific and the mannose-specific jacalin-type lectins differ in their sequences, they share a common fold: the beta-prism I fold, which is characteristic of Moraceae plant lectins. Here, two crystal structures of a jacalin-related lectin from Artocarpus hirsuta, which is specific for galactose, in complex with methyl-alpha D-galactose are reported. The lectin crystallized in two orthorhombic forms and one hexagonal form under similar conditions. The crystals had an unusually high solvent content. The structure was solved using the molecular-replacement method using the jacalin structure as a search model. The two orthorhombic forms were refined using data to 2.5 and 3.0 A resolution, respectively. The structures of the A. hirsuta lectin and jacalin are identical. In orthorhombic form I the crystal packing provides three different micro-environments for sugar binding in the same crystal. The observed difference in the specificity for oligosaccharides between the A. hirsuta lectin and jacalin could only be explained based on differences in the molecular associations in the packing and variation of the C terminal length of the beta-chain. The observed insecticidal activity of A. hirsuta lectin may arise from its similar fold to domain II of the unrelated delta-endotoxin from Bacillus thuringiensis. PMID- 15272164 TI - Spectacular improvement of X-ray diffraction through fast desiccation of protein crystals. AB - Succeeding in getting a protein to crystallize is not always the final hurdle in the determination of its three-dimensional structure. A relatively frequent and particularly vexing situation is the production of macroscopically well formed crystals that exhibit no suitable diffraction pattern. In this paper, three independent cases (i.e. proteins and crystallization conditions) are reported of spectacular diffraction-pattern improvement through a simple crystal-handling procedure that was discovered serendipitously. The procedure basically consists of removing a non-diffracting frozen crystal from the X-ray beam, plunging it into a soaking solution made of the original crystallization solution supplemented with a traditional cryoprotectant and then letting it dry in the evaporating sitting drop for some time (15 min to several hours). The treated crystals are then remounted and exhibit a huge improvement in their diffraction intensity and resolution. In all three cases presented here, the crystal quality shifted from unusable to perfectly suitable for structure determination. In addition to being a 'last resort' procedure for experimentalists struggling with non-diffracting crystals, this puzzling effect constitutes one more challenging problem for theoretical protein crystallographers. PMID- 15272165 TI - Calf spleen purine-nucleoside phosphorylase: crystal structure of the binary complex with a potent multisubstrate analogue inhibitor. AB - Purine-nucleoside phosphorylase (PNP) deficiency in humans leads to inhibition of the T-cell response. Potent membrane-permeable inhibitors of this enzyme are therefore considered to be potential immunosuppressive agents. The binary complex of the trimeric calf spleen phosphorylase, which is highly homologous to human PNP, with the potent ground-state analogue inhibitor 9-(5,5-difluoro-5 phosphonopentyl)guanine (DFPP-G) was crystallized in the cubic space group P2(1)3, with unit-cell parameter a = 93.183 A and one monomer per asymmetric unit. High-resolution X-ray diffraction data were collected using synchrotron radiation (EMBL Outstation, DESY, Hamburg, station X13). The crystal structure was refined to a resolution of 2.2 A and R and Rfree values of 19.1 and 24.2%, respectively. The crystal structure confirms that DFPP-G acts as a multisubstrate analogue inhibitor as it binds to both nucleoside- and phosphate-binding sites. The structure also provides the answers to some questions regarding the substrate specificity and molecular mechanism of trimeric PNPs. The wide access to the active-site pocket that was observed in the reported structure as a result of the flexibility or disorder of two loops (residues 60-65 and 251-266) strongly supports the random binding of substrates. The putative hydrogen bonds identified in the base-binding site indicate that N1-H and not O6 of the purine base defines the specificity of trimeric PNPs. This is confirmed by the fact that the contact of guanine O6 with Asn243 Odelta1 is not a direct contact but is mediated by a water molecule. Participation of Arg84 in the binding of the phosphonate group experimentally verifies the previous suggestion [Blackburn & Kent (1986), J. Chem. Soc. Perkin Trans. I, pp. 913-917; Halazy et al. (1991), J. Am. Chem. Soc. 113, 315-317] that fluorination of alkylphosphonates yields compounds with properties that suitably resemble those of phosphate esters and in turn leads to optimized interactions of such analogues with the phosphate-binding site residues. DFPP-G shows a Ki(app) in the nanomolar range towards calf and human PNPs. To date, no high-resolution X-ray structures of these enzymes with such potent ground-state analogue inhibitors have been available in the Protein Data Bank. The present structure may thus be used in the rational structure-based design of new PNP inhibitors with potential medical applications. PMID- 15272166 TI - The production and purification of the human T-cell receptors, the CD3epsilongamma and CD3epsilondelta heterodimers: complex formation and crystallization with OKT3, a therapeutic monoclonal antibody. AB - Human CD3 is an essential multisubunit complex that plays a fundamental role in T cell signalling, T-cell development and surface expression of the alphabeta T cell receptor. The CD3 complex comprises the CD3epsilongamma and CD3epsilondelta heterodimers and the CD3zetazeta homodimer. Here, the expression of the human CD3epsilongamma and CD3epsilondelta heterodimers, both of which were expressed as single-chain polypeptides, is reported. Following refolding, functional heterodimers were immunoaffinity purified from improperly folded heterodimers using OKT3, a therapeutic monoclonal antibody specific for the CD3epsilon chain. Subsequently, the Fab fragment of OKT3 was used to complex individually with the CD3epsilongamma and CD3epsilondelta heterodimers. Crystals of scCD3epsilongamma FabOKT3 were grown using 15%(w/v) PEG 3350, 200 mM potassium fluoride, 100 mM Tris-HCl pH 8.0. Crystals of scCD3epsilondelta-FabOKT3 were grown using 20%(w/v) PEG 3350, 200 mM potassium formate, 100 mM Tris-HCl pH 8.0, 2%(v/v) MPD. Crystals of both complexes diffract to beyond 3 A resolution. scCD3epsilongamma-FabOKT3 crystals belonged to space group P2(1), with unit-cell parameters a = 67.70, b = 55.77, c = 96.05 A, beta = 100.85 degrees and one complex per asymmetric unit. scCD3epsilondelta-FabOKT3 crystals belong to space group P2(1), with unit-cell parameters a = 101.67, b = 50.36, c = 138.7 A, beta = 108.84 degrees, suggesting two complexes per asymmetric unit. PMID- 15272167 TI - Expression, crystallization and crystallographic analysis of DegS, a stress sensor of the bacterial periplasm. AB - Regulated proteolysis is a key event in transmembrane signalling between intercellular compartments. In Escherichia coli, a protein DegS has been identified as being a periplasmic stress sensor for unfolded or misfolded outer membrane proteins (OMPs). Activation of DegS initiates a proteolytic cascade which results in the transcription of periplasmic genes under sigmaE control, most importantly chaperones and proteases. DegS has been cloned and expressed as full-length protein and in an N-terminally truncated form. Both proteins were tested for crystallization and two forms of well diffracting crystals of the truncated form were obtained. Crystals of form I diffract to 3.5 A and belong to space group P2(1)3, while crystals of form II diffract to 2.2 A and belong to space group I23. Crystals of form II were soaked with a consensus peptide representing the C-termini of outer membrane proteins and data to 2.4 A resolution were collected. Molecular-replacement trials using a homologous protease domain indicate the presence of two molecules in the asymmetric unit of crystal form I. The correctness of the molecular-replacement solution was verified by identifying radiation-damage-induced structural changes. PMID- 15272168 TI - Facile crystallization of Escherichia coli ketol-acid reductoisomerase. AB - Ketol-acid reductoisomerase (EC 1.1.1.86) catalyses the second reaction in the biosynthesis of branched-chain amino acids. The reaction involves an Mg2+ dependent alkyl migration followed by an NADPH-dependent reduction of the 2-keto group. Here, the crystallization of the Escherichia coli enzyme is reported. A form with a C-terminal hexahistidine tag could be crystallized under 18 different conditions in the absence of NADPH or Mg2+ and a further six crystallization conditions were identified with one or both ligands. With the hexahistidine tag on the N-terminus, 20 crystallization conditions were found, some of which required the presence of NADPH, NADP+, Mg2+ or a combination of ligands. Finally, the selenomethionine-substituted enzyme with the N-terminal tag crystallized under 15 conditions. Thus, the enzyme is remarkably easy to crystallize. Most of the crystals diffract poorly but several data sets were collected at better than 3.2 A resolution; attempts to phase them are currently in progress. PMID- 15272169 TI - Expression, purification, crystallization and preliminary X-ray diffraction data of methylmalonate-semialdehyde dehydrogenase from Bacillus subtilis. AB - Methylmalonate-semialdehyde dehydrogenase from Bacillus subtilis was cloned and overexpressed in Escherichia coli. Suitable crystals for X-ray diffraction experiments were obtained by the hanging-drop vapour-diffusion method using ammonium sulfate as precipitant. The crystals belong to space group P2(1)2(1)2(1), with unit-cell parameters a = 195.2, b = 192.5, c = 83.5 A, and contain one tetramer per asymmetric unit. X-ray diffraction data were collected to 2.5 A resolution using a synchrotron-radiation source. The crystal structure was solved by the molecular-replacement method. PMID- 15272170 TI - Crystallization and X-ray diffraction of a halogenating enzyme, tryptophan 7 halogenase, from Pseudomonas fluorescens. AB - Chlorination of natural products is often required for their biological activity; notable examples include vancomycin, the last-ditch antibiotic. It is now known that many chlorinated natural products are made not by haloperoxidases, but by FADH2-dependent halogenases. The mechanism of the flavin-containing enzymes is obscure and there are no structural data. Here, crystals of PrnA (tryptophan 7 halogenase), an enzyme that regioselectively chlorinates tryptophan, cocrystallized with tryptophan and FAD are reported. The crystals belong to the tetragonal space group P4(3)2(1)2 or P4(1)2(1)2, with unit-cell parameters a = b = 67.8, c = 276.9 A. A data set to 1.8 A with 93% completeness and an Rmerge of 7.1% has been collected from a single flash-cooled crystal. A method for incorporating selenomethionine in a Pseudomonas fluorescens expression system also is reported. PMID- 15272171 TI - Crystallization and preliminary crystallographic studies of the mitochondrial F1 ATPase from the yeast Saccharomyces cerevisiae. AB - A genetically modified (His6-tagged) form of the mitochondrial F1-ATPase (MW = 370 kDa) has been purified from the yeast Saccharomyces cerevisiae and crystallized in the presence of polyethelene glycol (PEG) 6000 as a precipitant, 1 mM NiCl2, 1 mM Mg AMP-PNP and 50 microM Mg ADP. X-ray diffraction data were obtained on three separate occasions using synchrotron radiation, with a progression in the quality of the diffraction data, which improved from 3.3 to 3.0 to 2.8 A. On the second occasion, the diffraction was improved by a crystal annealing procedure. The crystals belong to the monoclinic space group P2(1), with unit-cell parameters a = 110.6, b = 294.2, c = 190.4 A, beta = 101.6 degrees. The asymmetric unit contains three molecules of yeast F1, with a corresponding volume per protein weight (VM) of 2.8 A3 Da(-1) and a solvent content of 55%. PMID- 15272172 TI - Crystallization and preliminary X-ray analysis of carboxypeptidase 1 from Thermus thermophilus. AB - Carboxypeptidase 1 from the thermophilic eubacterium Thermus thermophilus (TthCP1, 58 kDa), a member of the M32 family of metallocarboxypeptidases, was crystallized by the sitting-drop vapour-diffusion method using PEG 8000 as the precipitant. The crystals diffracted X-rays to beyond 2.6 A resolution using a synchrotron-radiation source. The crystals belonged to the orthorhombic space group C222(1), with unit-cell parameters a = 171.0, b = 231.6, c = 124.9 A. The crystal contains three molecules in an asymmetric unit (VM = 2.11 A3 Da(-1)) and has a solvent content of 61.5%. PMID- 15272173 TI - Crystallization and preliminary X-ray crystallographic study of UDP-glucose pyrophosphorylase (UGPase) from Helicobacter pylori. AB - UDP-glucose pyrophosphorylase (UGPase) catalyzes the synthesis of UDP-glucose, an essential metabolite in all living organisms. An X-ray crystallographic study of UGPase from Helicobacter pylori has been performed in order to elucidate its role in the regulation of this important metabolic pathway. UGPase was crystallized from 0.1 M sodium acetate trihydrate pH 4.6, 2.0 M ammonium sulfate and 0.1 M guanidine-HCl. According to diffraction data collected at a resolution of 2.9 A using a synchrotron-radiation source, the crystal belongs to the orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 91.47, b = 98.61, c = 245.70 A, alpha = beta = gamma = 90.0 degrees. PMID- 15272174 TI - Purification, crystallization and preliminary crystallographic analysis of the glycine-cleavage system component T-protein from Pyrococcus horikoshii OT3. AB - The glycine-cleavage system component T-protein is a folate-dependent enzyme that catalyzes the formation of ammonia and 5,10-CH2-tetrahydrofolate from the aminomethyl intermediate bound to the lipoate cofactor of H-protein. T-protein from Pyrococcus horikoshii OT3 has been cloned, overexpressed in Escherichia coli, purified and crystallized by the microbatch method using PEG 4000 as a precipitant at 296 K. X-ray diffraction data have been collected to 1.50 A resolution at 100 K using synchrotron radiation. The crystals belong to the orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 78.980, b = 95.708, c = 118.331 A. Assuming one homodimer per asymmetric unit gives a VM value of 2.4 A3 Da(-1) and a solvent content of 49.0%. PMID- 15272175 TI - Crystallization and initial X-ray analysis of phenoxazinone synthase from Streptomyces antibioticus. AB - Phenoxazinone synthase, an oligomeric multicopper oxidase produced by Streptomyces antibioticus, is responsible for the six-electron oxidative coupling of two molecules of 4-methyl 3-hydroxyanthraniloyl pentapeptide to form the phenoxazinone chromophore of the antineoplastic agent actinomycin D. Spectroscopic studies have shown that the enzyme contains one type I (blue) and three to four type II copper centers. However, the exact arrangement of the copper centers in this multicopper oxidase is unknown. As a first step towards determining the three-dimensional structure of the enzyme, phenoxazinone synthase has been crystallized. The hexameric form of phenoxazinone synthase was purified from 72 h cultures of S. lividans containing the plasmid pIJ702. Purified hexamers were concentrated to 75 mg ml(-1) and used to grow two forms of crystals. Data collected from the two crystal forms were processed in two separate space groups. Crystals of both forms were grown at 288 K using the sitting-drop vapour-diffusion method. Native data sets extending to resolutions of 3.35 and 2.30 A have been collected and processed in space groups R32 and P1, respectively. PMID- 15272176 TI - Crystallization and preliminary analysis of active nitroalkane oxidase in three crystal forms. AB - Nitroalkane oxidase (NAO), a flavoprotein cloned and purified from Fusarium oxysporum, catalyzes the oxidation of neutral nitroalkanes to the corresponding aldehydes or ketones, with the production of H2O2 and nitrite. In this paper, the crystallization and preliminary X-ray data analysis of three crystal forms of active nitroalkane oxidase are described. The first crystal form belongs to a trigonal space group (either P3(1)21 or P3(2)21, with unit-cell parameters a = b = 103.8, c = 487.0 A) and diffracts to at least 1.6 A resolution. Several data sets were collected using 2theta and kappa geometry in order to obtain a complete data set to 2.07 A resolution. Solvent-content and Matthews coefficient analysis suggests that crystal form 1 contains two homotetramers per asymmetric unit. Crystal form 2 (P2(1)2(1)2(1); a = 147.3, b = 153.5, c = 169.5 A) and crystal form 3 (P3(1) or P3(2); a = b = 108.9, c = 342.5 A) are obtained from slightly different conditions and also contain two homotetramers per asymmetric unit, but have different solvent contents. A three-wavelength MAD data set was collected from selenomethionine-enriched NAO (SeMet-NAO) in crystal form 3 and will be used for phasing. PMID- 15272177 TI - Crystallization and preliminary crystallographic analysis of a thermostable family 52 beta-D-xylosidase from Geobacillus stearothermophilus T-6. AB - Beta-D-xylosidases (EC 3.2.1.37) are hemicellulases that hydrolyze short xylooligosaccharides into single xylose units. In this study, the first crystallization and preliminary X-ray analysis of a family 52 glycoside hydrolase, the beta-D-xylosidase (XynB2) from Geobacillus stearothermophilus T-6, is described. XynB2 is a dimeric protein consisting of two identical subunits of 705 amino acids with a calculated molecular weight of 79 894 Da. XynB2 was crystallized by the hanging-drop vapour-diffusion method and the crystals were found to belong to space group P1, with unit-cell parameters a = 80.6, b = 97.5, c = 107.2 A, alpha = 107.4, beta = 98.2, gamma = 106.6 degrees. The native crystals diffracted X-rays to a resolution of 2.0 A. PMID- 15272178 TI - Crystallization and preliminary X-ray crystallographic studies on a Kunitz-type potato serine protease inhibitor. AB - Interest in protease inhibitors has been renewed because of their potent activity in preventing carcinogenesis in a wide variety of in vivo and in vitro model systems. Potato tubers contain a wide range of such protease inhibitors. In cv. Elkana potato tubers, protease inhibitors represent about 50% of the total amount of soluble protein. Potato serine protease inhibitor (PSPI), one of the isoforms of the most abundant group of protease inhibitors, is a dimeric double-headed Kunitz-type inhibitor. No high-resolution structural information on this type of inhibitor has so far been obtained, as all currently known structures are of the monomeric single-headed or monomeric double-headed types. Crystals were grown in 0.1 M HEPES pH 7.5, 10% PEG 8000 and 8% ethylene glycol complemented with 9 mM 1 s-octyl-beta-D-thioglucoside or 0.1 M glycine. Data were collected from a single crystal under cryoconditions to 1.8 A resolution. The protein crystallized in space group P2(1), with unit-cell parameters a = 54.82, b = 93.92, c = 55.44 A, beta = 100.7 degrees; the scaling Rsym is 0.044 for 45,456 unique reflections. PMID- 15272179 TI - Crystallization and preliminary X-ray analysis of the outer membrane pyoverdine receptor FpvA from Pseudomonas aeruginosa. AB - FpvA, the pyoverdine outer-membrane receptor from Pseudomonas aeruginosa, is involved in iron uptake when bacteria grow under iron limitation. Crystals of the in vivo pyoverdine-loaded FpvA were obtained under several crystallization conditions using different detergents. A native data set was collected at 3.6 A resolution and a three-wavelength MAD data set was collected at 3.6 A resolution using crystals of selenomethionine-substituted protein. The crystals grew under similar conditions and both belong to space group C2, but have different unit cell parameters. PMID- 15272180 TI - Preparation and preliminary X-ray analysis of the catalytic module of beta-1,3 xylanase from the marine bacterium Vibrio sp. AX-4. AB - Beta-1,3-xylanase (1,3-beta-D-xylan xylanohydrolase; EC 3.2.1.32) is an enzyme capable of hydrolyzing beta-1,3-xylan. The newly cloned beta-1,3-xylanase from the marine bacterium Vibrio sp. AX-4 (XYL4) exhibited a modular structure consisting of three modules: an N-terminal catalytic module belonging to glycoside hydrolase family 26 and two C-terminal xylan-binding modules belonging to carbohydrate-binding module family 31. Despite substantial crystallization screening, crystallization of the recombinant XYL4 was not accomplished. However, the deletion mutant of XYL4, composed of a catalytic module without a xylan binding module, was crystallized. The crystal belonged to space group P2(1)2(1)2(1), with unit-cell parameters a = 51.6, b = 75.8, c = 82.0 A. X-ray diffraction data were collected to 1.44 A resolution. PMID- 15272181 TI - Complex assembly, crystallization and preliminary X-ray crystallographic studies of MHC H-2Kd complexed with an HBV-core nonapeptide. AB - In order to establish a system for structural studies of the murine class I major histocompatibility antigen complex (MHC) H-2Kd, a bacterial expression system and in vitro refolding preparation of the complex of H-2Kd with human beta2m and the immunodominant peptide SYVNTNMGL from hepatitis B virus (HBV) core-protein residues 87-95 was employed. The complex (45 kDa) was crystallized; the crystals belong to space group P222(1), with unit-cell parameters a = 89.082, b = 110.398, c = 47.015 A, alpha = beta = gamma = 90 degrees. The crystals contain one complex per asymmetric unit and diffract X-rays to at least 2.06 A resolution. The structure has been solved by molecular replacement and is the first crystal structure of a peptide-H-2Kd complex. PMID- 15272183 TI - Crystallization and preliminary X-ray diffraction analysis of phosphoglucose/phosphomannose isomerase from Pyrobaculum aerophilum. AB - Phosphoglucose isomerase from the crenarchaeon Pyrobaculum aerophilum (PaPGI/PMI) shows virtually no sequence similarity to its counterparts from bacterial and eukaryotic sources and belongs to a unique group within the PGI superfamily. Whereas conventional PGIs show strict substrate specificity for glucose 6 phosphate and fructose 6-phosphate, PaPGI/PMI can also catalyse the isomerization of mannose 6-phosphate. In order to establish its relatedness within the PGI family and to elucidate the structural basis for its broader specificity, this enzyme was crystallized. The crystals belong to space group P2(1) and a complete data set extending to 1.6 A resolution has been collected. PMID- 15272182 TI - Crystallization and preliminary crystallographic analysis of endonuclease VIII in its uncomplexed form. AB - The Escherichia coli DNA repair enzyme endonuclease VIII (EndoVIII or Nei) excises oxidized pyrimidines from damaged DNA substrates. It overlaps in substrate specificity with endonuclease III and may serve as a back-up for this enzyme in E. coli. The three-dimensional structure of Nei covalently complexed with DNA has been recently determined, revealing the critical amino-acid residues required for DNA binding and catalytic activity. Based on this information, several site-specific mutants of the enzyme have been tested for activity against various substrates. Although the crystal structure of the DNA-bound enzyme has been fully determined, the important structure of the free enzyme has not previously been analyzed. In this report, the crystallization and preliminary crystallographic characterization of DNA-free Nei are described. Four different crystal habits are reported for wild-type Nei and two of its catalytic mutants. Despite being crystallized under different conditions, all habits belong to the same crystal form, with the same space group (I222) and a similar crystallographic unit cell (average parameters a = 57.7, b = 80.2, c = 169.7 A). Two of these crystal habits, I and IV, appear to be suitable for full crystallographic analysis. Crystal habit I was obtained by vapour diffusion using PEG 8000, glycerol and calcium acetate. Crystal habit IV was obtained by a similar method using PEG 400 and magnesium chloride. Both crystals are mechanically strong and stable in the X-ray beam once frozen under cold nitrogen gas. A full diffraction data set has recently been collected from a wild-type Nei crystal of habit I (2.6 A resolution, 85.2% completeness, Rmerge = 9.8%). Additional diffraction data were collected from an Nei-R252A crystal of habit IV (2.05 A resolution, 99.9% completeness, Rmerge = 6.0%) and an Nei-E2A crystal of habit IV (2.25 A resolution, 91.7% completeness, Rmerge = 6.2%). These diffraction data were collected at 95-100 K using a synchrotron X-ray source and a CCD area detector. All three data sets are currently being used to obtain crystallographic phasing via molecular-replacement techniques. PMID- 15272184 TI - Crystallization and preliminary X-ray diffraction study of the catalytic subunit of archaeal H+ -transporting ATP synthase from Pyrococcus horikoshii OT3. AB - H+ -transporting ATP synthase (H+ -ATPase) is a multi-subunit complex which acts to produce ATP molecules. The catalytic subunit A of the archaeal-type H+ -ATPase from Pyrococcus horikoshii OT3 was cloned, expressed in Escherichia coli, purified and crystallized by the hanging-drop vapour-diffusion method with MPD as a precipitant. X-ray intensity data were collected to 2.55 A resolution at beamline BL41XU of SPring-8. The crystals belong to the tetragonal space group P4(1)2(1)2 or P4(3)2(1)2, with unit-cell parameters a = b = 128.0, c = 104.7 A, and contain one molecule per asymmetric unit. PMID- 15272185 TI - Crystallization and preliminary X-ray diffraction studies of the hyperthermophilic archaeal sulredoxin having the unique Rieske [2Fe-2S] cluster environment. AB - The hyperthermophilic archaeal sulredoxin from Sulfolobus tokodaii is a water soluble high-potential Rieske [2Fe-2S] protein with unique pH-dependent redox properties compared with its mesophilic homologues in cytochrome bc1/b6f complexes. The oxidized recombinant sulredoxin has been crystallized by the hanging-drop vapour-diffusion method using 30%(v/v) polyethylene glycol 400, 0.1 M cadmium chloride and 0.1 M sodium acetate pH 4.6. The crystals diffracted to beyond 2.0 A resolution and belong to the cubic space group F4(1)32, with unit cell parameter a = 163.00 +/- 0.05 A. The asymmetric unit contains one sulredoxin molecule. Three-wavelength MAD data were collected. PMID- 15272186 TI - Crystallization and preliminary X-ray study of alkaline mannanase from an alkaliphilic Bacillus isolate. AB - An alkaline mannanase (EC 3.2.1.78) from the alkaliphilic Bacillus sp. strain JAMB-602 was cloned and sequenced. The deduced amino-acid sequence of the enzyme suggested that the enzyme consists of a catalytic and unknown additional domains. The recombinant enzyme expressed by B. subtilis was crystallized using the hanging-drop vapour-diffusion method at 277 K. X-ray diffraction data were collected to 1.65 A. The crystal belongs to space group P2(1)2(1)2(1), with unit cell parameters a = 70.7, b = 79.5, c = 80.4 A. The asymmetric unit contains one protein molecule, with a corresponding VM of 2.26 A3 Da(-1) and a solvent content of 45.6%. Molecular replacement for initial phasing was carried out using the three-dimensional structure of a mannanase from Thermomonospora fusca as a search model, which corresponds to the catalytic domain of the alkaline mannanase. It gave sufficient phases to build the unknown domain. PMID- 15272187 TI - Crystallization and preliminary X-ray diffraction analysis of the lectin from Canavalia gladiata seeds. AB - The seed lectin from Canavalia gladiata was purified and crystallized. Orthorhombic crystals belonging to space group C222(1) grew within three weeks at 293 K using the hanging-drop vapour-diffusion method. Using synchrotron X-ray radiation, a complete structural data set was collected at 2.3 A resolution. The preliminary crystal structure of the lectin, determined by molecular replacement, had a correlation coefficient of 0.569 and an R factor of 0.412. PMID- 15272188 TI - Crystallization of the major cytosolic glutathione S-transferase from Onchocerca volvulus. AB - Glutathione S-transferases (GSTs) are a family of detoxification enzymes that catalyse the conjugation of glutathione to xenobiotic and endogenous electrophilic compounds, thus facilitating their elimination from cells. The recombinant Onchocerca volvulus GST2 has been expressed in Escherichia coli, purified and crystallized by the hanging-drop vapour-diffusion technique. Two different crystal forms were grown under identical conditions. They belong to space groups P2(1)2(1)2 and P2(1), respectively. The unit-cell parameters obtained are a = 112.6, b = 84.3, c = 45.1 A for the P2(1)2(1)2 crystal form and a = 51.6, b = 82.3, c = 56.7 A, beta = 95.89 degrees for the P2(1) form. Complete data sets to 2.6 and 1.5 A, respectively, have been collected at 100 K with synchrotron radiation. PMID- 15272189 TI - Crystallization and preliminary crystallographic analysis of monodehydroascorbate radical reductase from cucumber. AB - Monodehydroascorbate (MDA) radical reductase (EC 1.6.5.4) is an FAD enzyme that catalyzes the univalent reduction of MDA radical to ascorbate using NAD(P)H as an electron donor. The recombinant MDA reductase from cucumber was crystallized using polyethylene glycol 6000 as a precipitant. The crystals belong to space group P2(1), with unit-cell parameters a = 60.8, b = 138.6, c = 61.7 A, beta = 114.5 degrees, and contained two molecules per asymmetric unit. The Matthews coefficient (VM) and the solvent content are 2.46 A3 Da(-1) and 50.0%, respectively. Diffraction data were collected to a resolution of 2.4 A at 100 K using Cu Kalpha radiation with a multi-wire area detector and gave a data set with an overall Rsym of 10.0% and a completeness of 92.5%. PMID- 15272190 TI - Purification, crystallization and preliminary X-ray diffraction study of a recombinant cytokinin oxidase from Zea mays. AB - Cytokinins are hormones that are involved in plant growth and development. They are irreversibly degraded by cytokinin oxidases/dehydrogenases, flavoenzymes which contain a covalently bound flavine adenine dinucleotide (FAD) cofactor. Cytokinin oxidase from Zea mays (ZmCKO1) was overexpressed in the yeast Yarrowia lipolytica, purified (molecular weight 69 kDa) and crystallized using the hanging drop method. Crystals belong to the monoclinic space group C2, with unit-cell parameters a = 250.6, b = 50.6, c = 51.5 A, beta = 94.1 degrees. A complete data set has been collected at 100 K to 1.95 A resolution on an X-ray synchrotron source. PMID- 15272191 TI - C3 exoenzyme from Clostridium botulinum: structure of a tetragonal crystal form and a reassessment of NAD-induced flexure. AB - C3 exoenzyme from Clostridium botulinum (C3bot1) ADP-ribosylates and thereby inactivates Rho A, B and C GTPases in mammalian cells. The structure of a tetragonal crystal form has been determined by molecular replacement and refined to 1.89 A resolution. It is very similar to the apo structures determined previously from two different monoclinic crystal forms. An objective reassessment of available apo and nucleotide-bound C3bot1 structures indicates that, contrary to a previous report, the protein possesses a rigid core formed largely of beta strands and that the general flexure that accompanies NAD binding is concentrated in two peripheral lobes. Tetragonal crystals disintegrate in the presence of NAD, most likely because of disruption of essential crystal contacts. PMID- 15272192 TI - Heavy-atom derivatives in lipidic cubic phases: results on hen egg-white lysozyme tetragonal derivative crystals with Gd-HPDO3A complex. AB - Gd-HPDO3A, a neutral gadolinium complex, is a good candidate for obtaining heavy atom-derivative crystals by the lipidic cubic phase crystallization method known to be effective for membrane proteins. Gadolinium-derivative crystals of hen egg white lysozyme were obtained by co-crystallizing the protein with 100 mM Gd HPDO3A in a monoolein cubic phase. Diffraction data were collected to a resolution of 1.7 A using Cu Kalpha radiation from a rotating-anode generator. Two binding sites of the gadolinium complex were located from the strong gadolinium anomalous signal. The Gd-atom positions and their refined occupancies were found to be identical to those found in derivative crystals of hen egg-white lysozyme obtained by co-crystallizing the protein with 100 mM Gd-HPDO3A using the hanging-drop technique. Moreover, the refined structures are isomorphous. The lipidic cubic phase is not disturbed by the high concentration of Gd-HPDO3A. This experiment demonstrates that a gadolinium complex, Gd-HPDO3A, can be used to obtain derivative crystals by the lipidic cubic phase crystallization method. Further studies with membrane proteins that are known to crystallize in lipidic cubic phases will be undertaken with Gd-HPDO3A and other Gd complexes to test whether derivative crystals with high Gd-site occupancies can be obtained. PMID- 15272193 TI - Effect of silver-carrying photocatalyst "Hikari-Gintech" on mycobacterial growth in vitro. AB - The antimycobacterial activity of "Hikari-Gintech" powder, which has photocatalytic activity, was examined in vitro. Both powder dissolved in liquid and Hikari-Gintech-coated cloths showed strong antimycobacterial activity against Mycobacterium tuberculosis H37Rv, M. bovis BCG Pasteur, multi-drug-resistant M. tuberculosis (a clinical isolate) and M. avium. Hikari-Gintech powder appeared to affect mycobacterial cell wall metabolism rather than mycobacterial DNA because no damage to mycobacterial DNA was detected after spraying with Hikari-Gintech solution. PMID- 15272194 TI - Microbiota composition of the intestinal mucosa: association with fecal microbiota? AB - The fecal and mucosal microbiota of infants with rectal bleeding and the fecal microbiota of healthy age-matched controls were investigated by fluorescent in situ hybridization. Bifidobacteria were the main genus in both the feces and mucosa. The other genera tested, Bacteroides, Clostridium, Escherichia coli and lactobacilli/enterococci, represented only minor constituents. No differences in fecal microbiota were observed between patients and controls. In the patients, however, four times greater numbers of bifidobacteria were observed in the feces when compared to the mucosa. Notwithstanding this difference, a strong positive correlation prevailed for bifidobacteria in feces and mucosal samples. The genera assessed accounted for 16% of total bacterial counts on mucosal samples and for 47% of total bacterial counts in feces. This indicates that the unidentified part of the microbiota, especially on the mucosa, deserves more attention. PMID- 15272196 TI - Antibody reactive to a hepatitis C virus (HCV)-derived peptide capable of inducing HLA-A2 restricted cytotoxic T lymphocytes is detectable in a majority of HCV-infected individuals without HLA-A2 restriction. AB - Hepatitis C virus (HCV) is a single-strand RNA virus. Approximately 170 million people around the world are persistently infected and are at risk of liver cirrhosis or cancer. There is an urgent need to develop both therapeutic and diagnostic modalities of HCV. One approach to achieve these goals would be to determine highly immunodominant HCV peptides which are recognized by both cellular and humoral immunities. This study reports one such peptide, HCV-core protein at positions 35-44, having HLA-A2 binding motifs. IgG specific to this CTL-epitope peptide is consistently detectable in a majority of the patients with HCV infection regardless of the different HLA types, different disease conditions, and different HCV-genotypes tested. The sequence LPRR at positions 37 40 is considered to be the fine epitope recognized by the IgG. These results may provide new insights for the development of both therapeutic and diagnostic modalities of HCV at lower costs. PMID- 15272197 TI - Development and evaluation of a rapid flow-through immuno filtration test using recombinant filarial antigen for diagnosis of brugian and bancroftian filariasis. AB - There is an imperative need to develop a rapid antibody test that can be used for diagnosis of clinical cases in travelers and expatriates, primary surveillance in areas of unknown endemicity, detection of early infection in childhood and for monitoring chemotherapeutic programs. A rapid-format, simple and qualitative flow through immuno filtration test has been developed for the identification of total IgG antibodies to recombinant filarial antigen WbSXP-1. This test system employs colloidal gold-protein A reagent as the antibody capture reagent. The sensitivity and specificity of the test was evaluated in a total of 1,230 serum samples. The sensitivity of the test was found to be 90.8% with brugian (n = 70) and 91.4% with bancroftian (n = 140) microfilaraemic subjects. The test showed minimum reactivity (4/10) with Loa loa microfilaria (MF) positive sera and no reactivity (0/20) with Onchocerca MF positive sera. This rapid diagnosis is found to be non reactive with individuals having other parasitic diseases including schistosomiasis (n = 10), soil-transmitted helminthiases (n = 34) and protozoan infections (n = 33) indicating the potential of this test as a prospective method of diagnosis for both brugian and bancroftian lymphatic filariasis. Stability kinetics was studied at different temperatures and different time periods. The rapid flow-through immuno filtration test is advantageous since it can be stored at room temperature, is user friendly and is particularly applicable in the field as an initial screening method, for epidemiological monitoring of filarial infections in bancroftian and brugian endemic regions of the world. PMID- 15272198 TI - Isolation and phylogenetic analysis of arcobacter spp. in ground chicken meat and environmental water in Japan and Thailand. AB - Prevalence of Arcobacter spp. in chicken meat samples and environmental water samples in Japan and Thailand was investigated. Arcobacter was isolated from 48% of chicken meat samples (20/41) and 23% of river water samples (4/17) from Japan, and 100% of chicken meat samples (10/10) and 100% of canal water samples (7/7) from Thailand. A. butzleri was among the species isolated from all positive samples. About 10% genetic diversity was seen in the rpoB-rpoC in Arcobacters, and phylogenetic trees were divided into two clusters. In both countries, the results suggested that chicken and environmental water were highly contaminated with a genetically diverse population of Arcobacter. PMID- 15272199 TI - Ultrastructure of a Japanese Rickettsial strain genetically identified as Rickettsia helvetica which was originally found in Europe. AB - A rickettsial strain IO-1 has been isolated from a tick, Ixodes ovatus, in Japan and genetically identified as Rickettsia helvetica, a member of the spotted fever group rickettsiae. Ultrastructural observations were made on the microorganism. The ultrastructure of R. helvetica IO-1 appeared to be generally the same as that previously shown for other rickettsiae of the spotted fever and typhus groups. The rickettsiae were primarily found free in the cytoplasm of L929 cultured cells. Occasionally, the rickettsiae may also invade the host cell nucleus; however, the frequency of the nuclear localization was very low. PMID- 15272200 TI - Autolysis of Porphyromonas gingivalis is accompanied by an increase in several periodontal pathogenic factors in the supernatant. AB - Porphyromonas gingivalis autolyzes in the culture media. To examine in more detail the molecular components of the autolysate, two-dimensional gel electrophoresis was performed. Many protein spots varied both in number and volume. One of these spots included Arg-gingipain (Rgp) as determined by N terminal amino acid sequencing. Corresponding to the increase in spot volume, Rgp activity also increased during autolysis. The results of this study suggested that Rgp and other proteins in the P. gingivalis autolysate may be involved with the prolongation of periodontal disease, even after the death of P. gingivalis cells. PMID- 15272201 TI - Distribution of virulence-associated genes in Vibrio mimicus isolates from clinical and environmental origins. AB - Distribution of virulence-associated genes in Vibrio mimicus was studied including the toxin genes ctxA, tdh, st and vmh and the genes necessary for regulation of toxin production, toxR, toxS, toxT, tcpA and tcpP. Approximately half of clinical V. mimicus isolates possessed one or more genes encoding V. cholerae enterotoxic factors such as ctxA, tdh and st. All of the clinical and environmental isolates possessed vmh encoding V. mimicus hemolysin (VMH). The ctxA encoding cholera toxin was detected in only 2 strains, 5% of the clinical isolates. Furthermore, there were very few strains possessing tcpP and toxT needed for the expression of ctxA. These results may suggest that VMH is a more important pathogenic factor than well recognized toxins such as cholera toxin (CT) in V. mimicus infection. PMID- 15272195 TI - New World relapsing fever Borrelia found in Ornithodoros porcinus ticks in central Tanzania. AB - Ticks were collected from 8 houses in Mvumi Mission village, near Dodoma, Tanzania. All ticks were examined for Borrelia infestation by flagellin gene based nested polymerase chain reaction. All houses were highly infested with ticks, and all ticks collected were of the Ornithodoros porcinus species. Fifty one out of 120 ticks were infected with spirochetes, and a flagellin gene sequence comparison showed that most of the spirochetes belonged to Borrelia duttonii, which is the causative agent of tick-borne relapsing fever in East Africa. The rest of the spirochetes were quite different from B. duttonii and instead resembled the New World tick-borne relapsing fever borreliae. Phylogenetic analysis using 16S ribosomal RNA gene sequences also supported the interpretation that the spirochete was a Borrelia species distinct from previously described members of the genus. PMID- 15272202 TI - Epidemiological characterization of Salmonella Typhimurium DT104 prevalent among food-producing animals in the Japanese veterinary antimicrobial resistance monitoring program (1999-2001). AB - In the course of nationwide investigation on epidemiological characteristics in Salmonella Typhimurium isolates from food-producing animals in Japan between 1999 and 2001, fifty-seven isolates of S. Typhimurium DT104 and 104B obtained from cattle and swine at farm level in Japan between 1999 and 2001 were classified with pulsotype and antimicrobial resistance type. Most of the isolates were resistant to five or more antimicrobials and were genotyped into four groups. The present nationwide investigation shows that at least 11 types of S. Typhimurium related to DT104 are prevalent among food-producing animals across the country. PMID- 15272203 TI - Detection of the impairment of CD80 expression on circulating monocytes in HIV infected Thai children. AB - The mechanism of progressive anergic response in HIV-infected children has yet to be adequately described. One possibility is inappropriate delivery of an essential second signal for T-cell activation due to the inappropriate presentation of co-stimulatory molecules. To determine whether the ligand for the secondary signal is impaired in pediatric AIDS, we compared the level of CD80 expression by circulating monocytes in HIV-infected and-noninfected children (15 mild/asymptomatic, 13 symptomatic and 12 HIV seronegative children). By two-color flow cytometry analysis, there was no statistically significant difference in the percentage of monocytes expressing CD80 among the groups (i.e., 63.2 +/- 15.8, 60.9 +/- 12.7, 61.04 +/- 10.9 for uninfected children, mild-asymptomatic children and symptomatic children, respectively). However, both infected groups showed statistically significant lower levels of CD80 expression, with mean fluorescent intensities of 40.9 +/- 15.9 and 38.8 +/- 10.7 compared to 57.05 +/- 16.3 for the uninfected control group. Our data demonstrated a correlation between HIV infection and impairment of CD80 by circulating monocytes. Whether the impairment on CD80 expression contributes to destruction of the immunological network in HIV infected children requires further investigation. PMID- 15272204 TI - On the mechanism of levosimendan-induced dopamine release in the striatum of freely moving rats. AB - The Ca(2+) sensitizer levosimendan (LEV) improves myocardial contractility by enhancing the sensitivity of the contractile apparatus to Ca(2+). In addition, LEV promotes Ca(2+) entry through L-type channels in human cardiac myocytes. In this study, which was performed using microdialysis, infusion of LEV at 0.25 microM for 160 min increased dopamine (DA) concentrations (up to fivefold baseline) in dialysates from the striatum of freely moving rats. Ca(2+) omission from the perfusion fluid abolished baseline DA release and greatly decreased LEV induced DA release. Reintroduction of Ca(2+) in the perfusion fluid restored LEV induced DA release. Chelation of intracellular Ca(2+) by co-infusing 1,2-bis (o amino-phenoxy)ethane-N,N,N',N'-tetraacetic acid tetra (acetoxymethyl) ester (BAPTA-AM, 0.2 mM) did not affect basal DA release and scarcely affected LEV induced increases in dialysate DA. In addition, co-infusion of the L-type (Ca(v) 1.1-1.3) voltage-sensitive Ca(2+)-channel inhibitor nifedipine failed to inhibit LEV-induced increases in dialysate DA, which, in contrast, was inhibited by co infusion of the N-type (Ca(v) 2.2) voltage-sensitive Ca(2+)-channel inhibitor omega-conotoxin GVIA. We conclude that LEV promotes striatal extracellular Ca(2+) entry through N-type Ca(2+) channels with a consequent increase in DA release. PMID- 15272205 TI - Doxazosin effects on cholinergic and adrenergic responses in rat isolated detrusor smooth muscle preparations from obstructed bladder. AB - We investigated the effect of doxazosin on cholinergic and adrenergic agonists responses in detrusor smooth muscle preparations from sham-operated and 2-week partially obstructed rat bladders. Male Wistar albino rats, 200-250 g, were randomly allocated to 4 experimental groups consisting of 12 animals each: sham operated bladder, sham-operated bladder treated with doxazosin, partially obstructed bladder, and partially obstructed bladder treated with doxazosin. Partial outlet obstruction of the rat was surgically induced. The response to carbachol (10(-7)-10(-4) M), isoproterenol (10(-6)-10(-3) M), and 80 mM KCl were recorded. Carbachol caused concentration-dependent contractile responses in the detrusor smooth muscles from sham-operated and partially obstructed bladder. Isoproterenol produced concentration-dependent relaxation responses in the detrusor strips from all groups. Dose-response curves for carbachol and isoproterenol showed a shift to the left in rat detrusor smooth muscles from partially obstructed bladder when compared with the results obtained in detrusor muscles from sham-operated bladder. These responses were reversed to normal by doxazosin treatment in rat detrusor smooth muscles from partially obstructed bladder. KCl produced contractile responses in rat detrusor smooth muscles from all groups. The contractile responses to KCl were not significantly changed in all groups. We have shown that carbachol and isoproterenol responses were shifted to the left in rat detrusor smooth muscles from partially obstructed bladder and these responses were reversed by doxazosin treatment. PMID- 15272206 TI - Validation of a [3H]astemizole binding assay in HEK293 cells expressing HERG K+ channels. AB - A radioligand binding assay for the HERG (human ether-a-go-go-related gene) K(+) channel was developed to identify compounds which may have inhibitory activity and potential cardiotoxicity. Pharmacological characterization of the [(3)H]astemizole binding assay for HERG K(+) channels was performed using HERG expressing HEK293 cells. The assay conditions employed yielded 90% specific binding using 10 microg/well of membrane protein with 1.5 nM of [(3)H]astemizole at 25 degrees C. The K(d) and B(max) values were 5.91 +/- 0.81 nM and 6.36 +/- 0.26 pmol/mg, respectively. The intraassay and interassay variations were 11.4% and 14.9%, respectively. Binding affinities for 32 reference compounds (including dofetilide, cisapride, and terfenadine) with diverse structures demonstrated a similar potency rank order for HERG inhibition to that reported in the literature. Moreover, the [(3)H]astemizole binding data demonstrated a rank order of affinity that was highly correlated to that of inhibitory potency in the electrophysiological studies for HERG in HEK293 (r(SP) = 0.91, P<0.05). In conclusion, the [(3)H]astemizole binding assay is rapid and capable of detecting HERG inhibitors. PMID- 15272207 TI - Phosphodiesterase type IV inhibitors prevent ischemia-reperfusion-induced gastric injury in rats. AB - The effects of selective inhibitors of phosphodiesterase type IV (PDE4) on ischemia-reperfusion-induced gastric injuries were investigated in rats. Gastric ischemia was induced by applying a small clamp to the celiac artery, and reoxygenation was performed by removal of the clamp. Ischemia-reperfusion produced gastric hemorrhagic injuries and increased the content of the proinflammatory cytokine tumor necrosis factor-alpha (TNF-alpha) and myeloperoxidase (MPO) activity in gastric mucosa. Rolipram (0.03-0.3 mg/kg, s.c.) and Ro-20-1724 (0.3-3 mg/kg, s.c.) prevented the development of gastric injury in a dose-dependent manner, and it also inhibited the increase in mucosal TNF-alpha content and MPO activity induced by ischemia-reperfusion. The anti-ulcer drug irsogladine (1-10 mg/kg, p.o.), which is known to possess a PDE4 inhibitory action, also inhibited the gastric injury produced by ischemia-reperfusion, as well as the increase in TNF-alpha levels and MPO activity. It is concluded that the ability of PDE4 inhibitors to inhibit cytokine TNF-alpha synthesis and the infiltration of polymorphonuclear leukocytes underlies their gastroprotective effects in ischemia-reperfusion-induced gastric injury. Our experiments suggest that drugs that inhibit PDE4 isoenzyme, such as the anti-ulcer drug irsogladine, may be a useful adjunct therapy for the treatment of the gastric damage that follows ischemia-reperfusion. PMID- 15272208 TI - Failure of repeated electroconvulsive shock treatment on 5-HT4-receptor-mediated depolarization due to protein kinase A system in young rat hippocampal CA1 neurons. AB - We previously demonstrated that repeated electroconvulsive shock (ECS) treatment enhanced serotonin (5-HT)(1A)- and 5-HT(3)-receptor-mediated responses in hippocampal CA1 pyramidal neurons. The electrophysiological studies were performed to elucidate the effects of ECS treatment on depolarization, which was an additional response induced by 5-HT, and the second messenger system involved in this depolarization of hippocampal CA1 neurons. Both application of 5-HT (100 microM) induced depolarization of the membrane potential in the presence of 5 HT(1A)-receptor antagonists. This depolarization was mimicked by 5-HT(4)-receptor agonists, RS 67506 (1-30 microM) and RS 67333 (0.1-30 microM), in a concentration dependent manner. 5-HT- and RS 67333-induced depolarization was attenuated by concomitant application of RS 39604, a 5-HT(4)-receptor antagonist. H-89, a protein kinase A (PKA) inhibitor, inhibited 5-HT-, RS 67506-, and RS 67333 induced depolarizations, while forskolin (10 microM), an activator of adenylate cyclase, induced depolarization. Furthermore, RS 67333-induced depolarization was not significantly different between hippocampal slices prepared from rats administered ECS once a day for 14 days and those from sham-treated rats. These findings suggest that 5-HT(4)-receptor-mediated depolarization is caused via the cAMP-PKA system. In addition, repeated ECS-treatment did not modify 5-HT(4) receptor functions in contrast to 5-HT(1A)- and 5-HT(3)-receptor functions. PMID- 15272209 TI - Effects of norepinephrine and cardiotrophin-1 on phospholipase D activity and incorporation of myristic acid into phosphatidylcholine in rat heart. AB - The present study is part of a project on phospholipase D (PLD) in cardiac hypertrophy and analyzed effects on PLD activity of two growth stimuli, norepinephrine (NE) and cardiotrophin-1 (CT-1), in incubated rat heart. Phosphatidylcholine (PC) was labeled by (3)H-myristic acid. PLD produced (3)H phosphatidylethanol ((3)H-PEth) from (3)H-PC in the presence of ethanol and maintained a basal formation of (3)H-PEth. Short-term and long-term exposure to NE for 2 or 13 h, respectively, enhanced the formation of (3)H-PEth, which was blocked by prazosin. Long-term pretreatment with NE or CT-1 increased the incorporation of (3)H-myristic acid into PC, which was blocked by atenolol. When the (3)H-PEth formation was expressed as a fraction of (3)H-PC, PLD activity seemingly was unchanged (NE) or markedly reduced (CT-1); the true effects, namely, stimulation by NE and nonresponsiveness towards CT-1, were unraveled by atenolol (NE) or when PLD activity was expressed as (3)H-PEth per ng protein. In conclusion, alpha-adrenoceptor activation increased PLD activity. Long-term treatment with NE (via beta-receptors) or CT-1 enhanced the (3)H-myristic acid incorporation into a PC compartment, that was not available for the alpha receptor-mediated PLD activation. These results were discussed in regard to cellular mechanisms of cardiac hypertrophy and to the transphosphatidylation assay of PLD. PMID- 15272210 TI - Effects of mannitol on ischemia-induced degeneration in rat hippocampus. AB - Although mannitol has been used as an osmotherapeutic drug on brain injury, the clinical efficiency of the drug are still controversial. In the present study, we examined the effects of mannitol on the edema in a hippocampal slice due to brief ischemia. To evaluate the effects, we employed an image analysis system that consists of an infrared-differential interference contrast (IR-DIC) microscope, an infrared CCD camera, and a computer with custom-made software. By this system, severity of the edema can be quantified as the coefficient of variation (CV) of digitalized slice images. The dose-dependent improvement on the deteriorated hippocampal slices could be obtained by administration of mannitol (10, 50, and 100 mM) after 10-min ischemia. However, field excitatory postsynaptic potentials (fEPSP) in CA1 stratum radiatum, which disappeared during 10-min ischemia, were never recovered by mannitol after more than 20-min treatment. fEPSP were blocked by the effective dose of mannitol for morphological recovery, but the effects found to be reversible. Although we failed to find positive rescuing effects of mannitol on the synaptic activities after ischemia, the protective effects of the drug on ischemic edema may rescue the secondary damages around the infarct area. PMID- 15272211 TI - Expression of iNOS mRNA and inhibitory effect of NO on uterine contractile activity in rats are determined by local rather than systemic factors of pregnancy. AB - Our purpose was to investigate whether the local or systemic factors of pregnancy are associated with inducible nitric oxide synthase (iNOS) mRNA expression and to determine the inhibitory effects of pharmacological agents that increase cGMP levels in rat myometrium. iNOS mRNA expression was determined in uterine tissues from nonpregnant rats and on day 17 of gestation in the pregnant and non-pregnant uterine horns by RT-PCR. In addition, uterine rings from the pregnant and non pregnant uterine horns were placed in Krebs-Henseleit solution for isometric recordings of spontaneous contractions. Concentration-inhibition relationships to diethylamine/nitric oxide complex, 8-bromo-cGMP, and the selective phosphodiesterase V inhibitor were obtained. Compared to nonpregnant rats, expression of iNOS mRNA in myometrium increased during pregnancy, which was maximal on day 17, followed by a decrease on day 21 of gestation. Expression of iNOS mRNA at day 17 of gestation was greater in pregnant uterine horns than in nonpregnant ones. Maximal inhibition of phosphodiesterase V and increasing cGMP induced similar inhibition of spontaneous contractions in nonpregnant and pregnant uterine horns, while NO induced less inhibition in the former. The results suggest that the local pregnancy factor is needed for signal transduction from NO to soluble guanylate cyclase at a time when maximal expression of iNOS mRNA is evident. PMID- 15272212 TI - Antiepileptic effects of single and repeated oral administrations of S-312-d, a novel calcium channel antagonist, on tonic convulsions in spontaneously epileptic rats. AB - We investigated the effects of single and repeated administrations of S-312-d (methyl-4,7-dihydro-3-isobutyl-6-methyl-4-(3-nitrophenyl)-thieno-[2,3-b]pyridine 5-carboxylate), a newly synthesized L-type Ca(2+)-channel blocker, on tonic convulsions and absence-like seizures in the spontaneously epileptic rat (SER: zi/zi, tm/tm), a genetically based animal model of human epilepsy. Single oral administrations of S-312-d dose-dependently inhibited tonic convulsions and the effects lasted for more than 2 h, although they did not attenuate the absence like seizures. We also examined the effects of repeated administrations of S-312 d at 1 mg/kg once a day for 4 days on SER. A significant decrease in the number and total duration of tonic convulsions was observed 45 and 75 min after the first administration of the drug, respectively. The effects lasted for 24 h without changes in the background EEG or blood pressure. This inhibitory effect on the tonic convulsions was gradually strengthened by subsequent daily administrations of S-312-d and lasted for 3 days after the cessation of drug treatment. In contrast, the repeated treatment with S-312-d did not influence absence-like seizures of SER. These results suggest that S-312-d is a candidate drug that has antiepileptic effects against the convulsive seizures in human epilepsy. PMID- 15272213 TI - Sevoflurane inhibition of the slowly activating delayed rectifier K+ current in guinea pig ventricular cells. AB - Single ventricular cells were enzymatically isolated from guinea pig hearts and the effects of sevoflurane on the delayed rectifier K(+) current were investigated by the patch clamp method. The rapidly (I(Kr)) and slowly activating delayed rectifier K(+) current (I(Ks)) were isolated using chromanol 293B, a selective blocker for I(Ks) or E4031 (N-[4-[[1-[2-(6-methyl-2-pyridinyl)ethyl]-4 piperidinyl]carbonyl]phenyl]methanesulfonamide dihydrochloride), a blocker for I(Kr). Sevoflurane and halothane decreased I(Ks) in a concentration-dependent manner with an IC(50) value of 0.38 mM for sevoflurane and 1.05 mM for halothane. I(Ks) inhibition was characterized by suppression of maximum conductance with little effect on activation kinetics. Inhibition occurred immediately after anesthetic application and recovered upon wash-out. In contrast to the marked inhibition of I(Ks), I(Kr) was hardly affected by sevoflurane. Under the current clamp, sevoflurane prolonged the action potential duration in a reversible manner and this effect was more marked when I(Kr) was inhibited by E4031. The results suggest that sevoflurane inhibits I(Ks), and not I(Kr), in a concentration dependent manner at clinically relevant concentrations. The resulting prolongation of ventricular repolarization may partly account for the clinical observation of excessive QT prolongation by these anesthetics. PMID- 15272214 TI - Potential anxiolytic and antidepressant-like activities of SNC80, a selective delta-opioid agonist, in behavioral models in rodents. AB - In the present study, we investigated the antidepressant- and anxiolytic-like effects of (+)-4-[(aR)-a-((2S,5R)-4-allyl-2,5-dimethyl-1-piperazinyl)-3 methoxybenzyl]-N,N-diethylbenzamide (SNC80), a non-peptidic selective delta opioid receptor agonist, in various animal models in rodents. SNC80 significantly reduced the duration of immobility in the forced swimming test. Furthermore, in the elevated plus-maze test, SNC80 dose-dependently and significantly increased the time spent in the open arms of the plus-maze. These effects were completely antagonized by a selective delta-opioid-receptor antagonist, naltrindole. In the conditioned fear stress test, which examines psychological stress-induced motor suppression, desipramine did not produce any significant effect on the conditioned suppression of locomotor activity. However, SNC80 completely attenuated the conditioned suppression of locomotor activity in the conditioned fear stress test. In conclusion, our results suggest that delta-opioid receptors may play an important role in the regulation of emotional responses. Furthermore, it is possible that delta-opioid-receptor agonists might be novel and potent antidepressants that also have anxiolytic-like effects. PMID- 15272215 TI - A pegylated liposomal platform: pharmacokinetics, pharmacodynamics, and toxicity in mice using doxorubicin as a model drug. AB - Aims were to observe pharmacokinetics, pharmacodynamics, and toxicity for constructing a Sino-pegylated liposomal platform. Human hepatocarcinoma cells (Bel7402) and murine hepatocarcinoma cells (H(22)) were used for the cytotoxicity assay and the in vivo solid xenograft tumor model in mice, respectively. Pharmacokinetic results in mice showed that the pegylated liposomal doxorubicin markedly prolonged the blood circulation of doxorubicin. Elimination half-time (T(1/2,gamma)) of pegylated, regular liposomal doxorubicin and free doxorubicin were 46.09 +/- 14.44, 26.04 +/- 3.34, and 23.72 +/- 5.13 h, respectively. The area under the concentration-time curves (AUC(0- infinity )) (h. microg/g) of the pegylated and regular liposomal doxorubicin were 6.8- and 2.6-fold higher than that of free doxorubicin, respectively. Cytotoxicity and antitumor activity in vivo indicated that activity of the pegylated liposomal doxorubicin was higher than that of the regular or the free one, respectively. After two weeks of tail intravenous injection of the pegylated liposomal doxorubicin at a single dose of 10 mg/kg, no significant damage was observed in gastric, intestinal mucosa, and heart muscle, but pronounced damages were found in the control group after dosing free doxorubicin. The results demonstrate that the pegylated liposomes improve the efficacy of toxics and reduce the toxicity, therefore providing favorable evidence for building a pegylated liposomal platform. PMID- 15272216 TI - Captopril enhances cardiac vagal but not sympathetic neurotransmission in pithed rats. AB - The effect of captopril on neurally evoked bradycardia and tachycardia was investigated in pithed rats. Captopril enhanced the vagal nerve stimulation evoked bradycardia. Angiotensin I reduced the vagal bradycardia, which was reversed by subsequent administration of captopril. Bradykinin did not affect the neurally evoked bradycardia. Captopril and angiotensin I affected neither the exogenous acetylcholine-evoked bradycardia nor the sympathetic nerve stimulation evoked tachycardia. These results suggest that the interruption of angiotensin II formation by captopril causes less presynaptic inhibition of acetylcholine release via angiotensin II receptors without affecting cardiac sympathetic neurotransmission. PMID- 15272217 TI - Combined effects of benidipine and diltiazem in a rat model of experimental angina. AB - We examined the combined effects of the calcium channel blockers 1,4 dihydropyridine (benidipine) and benzothiazepine (diltiazem) on vasopressin induced myocardial ischemia in anesthetized rats, an experimental model of angina. Benidipine (3, 10 microg/kg, i.v.) and diltiazem (300, 1000 microg/kg, i.v.) caused dose-related inhibition of vasopressin-induced S-wave depression, an index of myocardial ischemia. Co-administration of low doses of benidipine (3 microg/kg) and diltiazem (300 microg/kg) almost completely inhibited the S-wave depression, where the efficacy was similar to that obtained with the use of high doses of benidipine (10 microg/kg) or diltiazem (1000 microg/kg). These results suggest that the administration strategy employed may be useful in the treatment of angina pectoris. PMID- 15272218 TI - Effect of cilnidipine on L- and T-type calcium currents in guinea pig ventricle and action potential in rabbit sinoatrial node. AB - Cilnidipine, a dihydropyridine Ca(2+) channel antagonist, is known to have inhibitory effects on both L- and N-type Ca(2+) currents. In the present study, we examined the effect of cilnidipine on myocardial L- and T-type Ca(2+) currents and sinoatrial node action potential configuration. In voltage clamped guinea pig ventricular myocytes, cilnidipine concentration-dependently decreased L- and T type Ca(2+) currents. In rabbit sinoatrial node tissue, cilnidipine increased cycle length through reduction of phase 4 depolarization slope. In conclusion, cilnidipine has inhibitory effects on T-type Ca(2+) current, which may contribute to its negative chronotropic potency. PMID- 15272219 TI - Incidence of dementia in a rural community in Spain: the Girona cohort study. AB - BACKGROUND: Information on dementia incidence in Spanish populations is still scarce, and there is a dearth of prospective studies. OBJECTIVE: To estimate the incidence rates of dementia, Alzheimer's disease (AD) and vascular dementia (VaD) in a population cohort aged 75 and over in a rural area in Spain. METHODS: A prospective population cohort study over a 5-year period in 8 rural villages in the province of Girona. The baseline study in 1990 identified 200 prevalent cases of dementia. The dementia-free cohort included 1,260 persons aged 75 and over. This was the sample used for the incidence study. We rescreened and selectively reexamined this group in 1995 using a two-phase procedure consisting of a screening interview at home using the MMSE. Diagnoses of dementia, AD and VaD were established using the Cambridge Examination for Mental Disorders of the Elderly for surviving participants. For deceased participants, we used the Retrospective Collateral Dementia Interview to establish a diagnosis of dementia and AD according to DSM-III-R diagnostic criteria. RESULTS: Information was obtained for 91% of the subjects at risk; 122 incident cases of dementia were identified. Incidence rates per 1,000 person-years at risk were 23.2 (95% CI = 19.1-27.3) for dementia, 10.8 (95% CI = 7.8-13.7) for AD and 9.5 (95% CI = 6.7 12.1) for VaD. All dementia subtypes showed an age-dependent pattern. Females had a relative risk of 1.8 (95% CI = 1.0-3.4) to develop AD. The inclusion of deceased cases with manifestations of dementia increased the rate of dementia incidence in 7.1 cases/1,000 person-years at risk. CONCLUSION: Incidence rates were similar to those reported by other cohort studies. All dementia subtypes increased with age, but incidence rates did not increase exponentially in the oldest old. Females were at increased risk for AD. The inclusion of information about dementia symptoms from relatives of deceased participants was useful in order to avoid underestimation of the dementia incidence rates. Underestimation of the incidence rates was more important in those aged 75-84 years. PMID- 15272220 TI - Cerebral haemodynamics in the elderly: the rotterdam study. AB - Transcranial Doppler ultrasonography allows the non-invasive assessment of cerebral haemodynamics. Data on the frequency distribution of these parameters in the oldest old are scarce, which makes a distinction between physiological and pathophysiological ageing difficult. We studied the relation between cerebral haemodynamic parameters and age and sex in 1,720 participants of a population based study by means of transcranial Doppler ultrasonography. The end-diastolic, peak systolic and mean cerebral blood flow velocity, and cerebrovascular CO2 reactivity declined significantly with increasing age up to 90 years (per year 0.6 cm/s; -0.3 cm/s; -0.5 cm/s, and -0.6%/kPa, respectively). The pulsatility index increased with age (per year 0.01). End-diastolic, peak systolic and mean cerebral blood flow velocities were lower in men compared to women (age-adjusted difference 1.6 cm/s; 4.1 cm/s, and 2.5 cm/s, respectively). Cerebrovascular CO2 reactivity was higher in men compared to women. Adjusting for carotid atherosclerosis did not change the observed sex differences. These findings provide insight into physiologic changes of haemodynamics during ageing and may serve as a starting point for investigations on determinants of pathophysiologic changes in cerebral haemodynamics in the elderly. PMID- 15272221 TI - Misclassification is likely in the assessment of mild cognitive impairment. AB - We estimated the relative frequency of isolated memory impairment versus isolated and comorbid impairment in executive control function (ECF). One hundred and ninety-three noninstitutionalized residents of a single Comprehensive Care Retirement Community (mean age 79.2 years) were investigated. The subjects were tested with multiple measures of memory and ECF. Test scores were standardized to minimize scaling effects. 'Impairment' was defined as performance < or =1.5 standard deviations below the mean for the entire sample (i.e., a z score < or = 1.5). Disability was estimated as the sum of self-reported activities of daily living and instrumental activities of daily living. The cognitive test performance was significantly associated with functional impairment, independently of age. ECF and memory measures were significantly intercorrelated. Both were significantly and independently associated with disability ratings. 6 10% of the subjects had memory impairment; 25-35% of the memory-impaired subjects had comorbid ECF impairments. An additional 4-7% of the subjects had isolated ECF impairment. A significant fraction of the cases otherwise meeting the criteria for 'mild cognitive impairment' may have comorbid ECF impairment. This raises the issue of whether they might be more properly classified as 'demented'. In addition, isolated ECF impairment may affect almost as many persons as isolated memory impairment. Isolated ECF impairment is not consistent with the natural history of preclinical Alzheimer's disease, suggests other conditions, and can be disabling, independently of age and/or memory loss. PMID- 15272222 TI - Incidence of amyotrophic lateral sclerosis and of the parkinsonism-dementia complex of Guam, 1950-1989. AB - Studies representing the accumulated information from the first 30 years of research effort on Guam (1950-1979) have demonstrated a varying degree of decline in the incidence of amyotrophic lateral sclerosis (ALS) and the parkinsonism dementia complex (PDC) of Guam. Analysis with more complete information for the period 1980-1989 provides more valid estimates of the later patterns in the incidence of ALS and PDC and affords a more extensive assessment of trends over a 40-year period. The annual age-adjusted incidence of ALS was 7/100,000 and the annual age-adjusted incidence of PDC was 22/100,000 in 1989. The incidence was much higher for the period 1980-1989 than suggested in previous reports. These findings provide compelling evidence that this spectrum of neurodegenerative diseases continues to have a significant impact on the health of the Chamorro people of Guam. PMID- 15272223 TI - Epidemiology of multiple system atrophy: a prevalence and pilot risk factor study in Aquitaine, France. AB - We investigated the prevalence of multiple system atrophy (MSA) in Gironde, France, through a network of 120 public and private specialists and assessed the relationship between some environmental factors and MSA in a case-control study involving 50 MSA patients, 50 Parkinson's disease (PD) patients and 50 healthy controls. The occupational exposure to pesticides was evaluated through a job exposure matrix. On prevalence day (November 1, 1998), the crude prevalence of MSA in Gironde was 1.94/100,000 inhabitants. We found no significant relationship between occupational exposure to pesticides and MSA. PD patients were significantly less frequently ever-smokers than controls and the same tendency was observed for MSA patients. We also described the clinical features that heralded the disease among this nonselected population. PMID- 15272224 TI - Does rivastigmine improve cognitive functions or disability in patients with Alzheimer's disease? PMID- 15272225 TI - A new perspective on cardiotoxicity of 5-fluorouracil. A novel research tool 'cardiac ultrasonic integrated backscatter analysis' indicates transient, subclinical myocardial dysfunction due to high-dose leucovorin and infusional 5 fluorouracil regimen. AB - BACKGROUND: The pathophysiology of 5-fluorouracil (5-FU) cardiotoxicity is still controversial. The objective of this study was to assess the influence of high dose leucovorin and infusional 5-FU regimen (HDLV5FU) on cardiac tissues. METHODS: We monitored 28 patients (median age 68 years) under HDLV5FU chemotherapy with complete blood counts, cardiac enzymes, C-reactive protein, coagulation tests, Holter electrocardiogram, and conventional echocardiography. Cardiac ultrasonic tissue characterization with integrated backscatter (IBS) analysis was performed in the 16 last enrolled patients. RESULTS: The magnitude of both anterior and posterior cardiac IBS values significantly decreased at the 48th hour of treatment compared to both 0th hour and day 15 (p < 0.003). Cardiac IBS values on the 15th day were not different from the 0th hour. Clinical cardiotoxicity was not observed and other monitored parameters did not change significantly in any patient (p > 0.5 for all). CONCLUSION: Cardiac IBS analysis suggests that 5-FU might cause reversible subclinical myocardial dysfunction. PMID- 15272226 TI - Hypoxia-inducible factor 1alpha- mediated resistance to phenolic anticancer. AB - BACKGROUND: Phenolic compounds EGCG [(-)-epigallocatechin-3-gallate], resveratrol (3,4',5-trihydroxy-trans-stilbene) and capsaicin (trans-8-methyl-N-vanillyl-6 nonenamide) are worth investigating for clinical application in cancer prevention and chemotherapy. Hypoxia-induced drug resistance is a major obstacle in the development of effective cancer chemotherapy. Therefore, we examined whether drug resistance to these phenolic compounds is acquired by hypoxia. METHODS: Hep3B hepatoma, Caki-1 renal carcinoma, SK-N-MC neuroblastoma, and HEK293 cell lines were cultured under normoxic or hypoxic conditions. Drug sensitivities to the phenolic compounds and expression of hypoxia-inducible factor-1alpha (HIF-1alpha) and the multidrug resistance genes were examined in these cell lines. RESULTS: Drug resistance was acquired 24 h after hypoxia and subsided 8 h after reoxygenation. Protein synthesis inhibitors abolished this drug resistance. A transfection study demonstrated that HIF-1alpha enhanced this hypoxia-induced resistance and that its dominant-negative isoform suppressed resistance acquisition. However, MDR1 and MRP1, which provide multidrug resistance to conventional anticancer agents, were not induced by hypoxia. CONCLUSIONS: These results suggest that HIF-1alpha-dependent gene expression participates in the cellular process of the hypoxia-induced resistance to phenolic compounds. PMID- 15272227 TI - Antibacterial resistance of community-acquired respiratory tract pathogens recovered from patients in Germany and activity of the Ketolide Telithromycin: results from the PROTEKT surveillance study (1999-2000). AB - BACKGROUND: The Prospective Resistant Organism Tracking and Epidemiology for the Ketolide Telithromycin (PROTEKT) longitudinal global surveillance study examines the antibacterial susceptibility of community-acquired respiratory pathogens. METHODS AND RESULTS: Data from isolates collected in Germany in 1999-2000 in the PROTEKT study show that 8.3% of pneumococcal isolates (n = 325) had reduced susceptibility to penicillin and 2.2% were fully resistant. Erythromycin resistance was 15.7% overall and particularly high in Leipzig (31.6%). All penicillin- and erythromycin-resistant strains were inhibited by telithromycin (MIC < or =0.5 mg/l) and linezolid (MIC < or =2 mg/l). Beta-lactamase was produced by 3.2% of Haemophilus influenzae (9/284) and 89.5% of Moraxella catarrhalis strains (111/124). All Streptococcus pyogenes isolates (n = 87) were susceptible to penicillin, although 9.2% were resistant to macrolides. CONCLUSIONS: Penicillin resistance in Germany remains low; however, the prevalence of antimicrobial resistance among common respiratory pathogens is rising, particularly against macrolides. Continued surveillance is necessary to guide optimal empirical therapy, and new antimicrobials, like telithromycin, need to be developed with improved potency against target pathogens and low propensity for the development of resistance. PMID- 15272228 TI - Synergism of ciprofloxacin and trimethoprim against Salmonella enterica serovar typhi isolates showing reduced susceptibility to ciprofloxacin. AB - The activity of the combination of ciprofloxacin and trimethoprim against 16 Salmonella enterica serovar typhi isolates from blood cultures were tested by agar dilution checkerboard technique. When used in combination, minimum inhibitory concentrations (MICs) of ciprofloxacin and trimethoprim ranged from 0.5 to 1.25 and from 10 to 125 microg/ml, respectively, and fractional inhibitory concentrations (FICs) from 0.025 to 0.125 and from 2.5 to 10 microg/ml, respectively. The FIC index was 0.140-0.483, indicating a marked synergy between ciprofloxacin and trimethoprim against trimethoprim-resistant S. enterica serovar typhi isolates (100%) with high MICs for ciprofloxacin. PMID- 15272229 TI - Combined effect of atovaquone and pyrrolidine dithiocarbamate in the treatment of acute murine toxoplasmosis. PMID- 15272230 TI - Calcium cycling proteins in heart failure, cardiomyopathy and arrhythmias. AB - A growing body of evidence, including studies using genetically engineered mouse models, has shown that Ca2+ cycling and Ca2+-dependent signaling pathways play a pivotal role in cardiac hypertrophy and heart failure. In addition, recent studies identified that mutations of the genes encoding sarcoplasmic reticulum (SR) proteins cause human cardiomyopathies and lethal ventricular arrhythmias. The regulation of Ca2+ homeostasis via the SR proteins may have potential therapeutic value for heart diseases such as cardiomyopathy, heart failure and arrhythmias. PMID- 15272231 TI - Human beta-defensin 2 is induced by interleukin-1beta in the corneal epithelial cells. AB - Mammalian epithelia produce the various antimicrobial peptides against the bacterial or viral infection, thereby acting as the active immune modulators in the innate immunity. In this study, we examined the effects of the various proinflammatory cytokines or LPS on cell viability and antimicrobial beta defensin gene expressions in human corneal epithelial cells. Results showed that the cytokines or LPS did not exert severe cytotoxic effects on the cells, and that beta-defensin 1 was constitutively expressed, while beta-defensin 2 was specifically induced by IL-1beta, supporting the idea that these cytokines or LPS involve the defense mechanism in the cornea. Furthermore, the reporter and gel shift assay to define the induction mechanism of beta-defensin 2 by IL-1beta demonstrated that the most proximal NF-kappaB site on the promoter region of beta defensin 2 was not critical for the process. Data obtained from the normal or patients with the varying ocular diseases showed that our in vitro results were relevant in the clinical settings. Our results clearly demonstrated that beta defensin 1 and 2 are important antimicrobial peptides in the corneal tissues, and that the mechanistic induction process of beta-defensin 2 by IL-1beta is not solely dependent on proximal NF-kappaB site activation, thus suggesting that the long distal portion of the promoter is needed for the full responsiveness toward IL-1beta. PMID- 15272232 TI - Beta ig-h3 promotes renal proximal tubular epithelial cell adhesion, migration and proliferation through the interaction with alpha3beta1 integrin. AB - Betaig-h3 (betaig-h3) is a secretory protein composed of fasciclin I-like repeats containing sequences that allows binding of integrins and glycosaminoglycans in vivo. Expression of betaig-h3 is responsive to TGF-Beta and the protein is found to be associated with extracellular matrix (ECM) molecules, implicating betaig-h3 as an ECM adhesive protein of developmental processes. We previously observed predominant expression of betaig-h3 expression in the basement membrane of proximal tubules of kidney. In this study, the physiological relevance of such localized expression of betaig-h3 was examined in the renal proximal tubular epithelial cells (RPTEC). RPTEC constitutively expressed betaig-h3 and the expression was dramatically induced by exogenous TGF-Beta1 treatment. betaig-h3 and its second and fourth FAS1 domain were able to mediate RPTEC adhesion, spreading and migration. Two known alpha3beta1 integrin-interaction motifs including aspartatic acid and isoleucine residues, NKDIL and EPDIM in betaig-h3 were responsible to mediate RPTEC adhesion, spreading, and migration. By using specific antibodies against integrins, we confirmed that alpha3beta1 integrin mediates the adhesion and migration of RPTECs on betaig-h3. In addition, it also enhanced proliferation of RPTECs through NKDIL and EPDIM. These results indicate that betaig-h3 mediates adhesion, spreading, migration and proliferation of RPTECs through the interaction with alpha3beta1 integrin and is intimately involved in the maintenance and the regeneration of renal proximal tubular epithelium. PMID- 15272233 TI - Involvement of thromboxane A2 and tyrosine kinase in the synergistic interaction of platelet activating factor and calcium ionophore A23187 in human platelet aggregation. AB - The present study was carried out to examine the mechanisms of the synergistic interaction of PAF and A23187 mediated platelet aggregation. We found that platelet aggregation mediated by subthreshold concentrations of PAF (5 nM) and A23187 (1 mM) was inhibited by PAF receptor blocker (WEB 2086, IC50 = 0.65 mM) and calcium channel blockers, diltiazem (IC50 = 13 mM) and verapamil (IC50 = 18 mM). Pretreatment of platelets with PAF and A23187 induced rise in intracellular calcium and this effect was also blocked by verapamil. While examining the role of the down stream signaling pathways, we found that platelet aggregation induced by the co-addition of PAF and A23187 was also inhibited by low concentrations of phospholipase C (PLC) inhibitor (U73122; IC50 = 10 mM), a cyclooxygenase inhibitor (indomethacin; IC50 = 0.2 mM) and inhibitor of TLCK, herbimycin A with IC50 value of 5 mM. The effect was also inhibited by a specific TXA2 receptor antagonist, SQ 29548 with very low IC50 value of 0.05 mM. However, the inhibitors of MAP kinase, PD98059 and protein kinase C, chelerythrine had no effect on PAF and A23187-induced platelet aggregation. These data suggest that the synergism between PAF and A23187 in platelet aggregation involves activation of thromboxane and tyrosine kinase pathways. PMID- 15272234 TI - Interleukin-1beta stimulates matrix metalloproteinase-2 expression via a prostaglandin E2-dependent mechanism in human chondrocytes. AB - IL-1beta is known promote cyclooxygenase-2 (COX- 2) and matrix metalloproteinase 2 (MMP-2) expression. This study focuses on the characterization of the signaling cascade associated with IL-1beta-induced matrix metalloproteinase-2 (MMP-2) regulation in human chondrocytes. The decrease in collagen levels in the conditioned media was prevented by a broad spectrum MMP inhibitor, suggesting that IL-1beta promotes the proteolytic process leading to MMP-2 activation. IL 1beta-related MMP-2 expression was found to be dependent on prostaglandin E2 (PGE2) production. In addition, the induction of COX-2 and MMP-2 was inhibited by the pretreatment of chondrocytes with a SB203580 or Ro 31-8220, indicating the involvement of protein kinase C (PKC) or p38 mitogen-activated protein kinase (MAPK). However, there is no cross-talk between PKC and p38 MAPK in the IL-1beta induced MMP-2 activation. Taken together, these results demonstrated that IL 1beta induces MMP-2 expression through the PGE2-dependent mechanism in human chondrocytes. PMID- 15272235 TI - Association of DNA-dependent protein kinase with hypoxia inducible factor-1 and its implication in resistance to anticancer drugs in hypoxic tumor cells. AB - Tumor hypoxia contributes to the progression of a malignant phenotype and resistance to ionizing radiation and anticancer drug therapy. Many of these effects in hypoxic tumor cells are mediated by expression of specific set of genes whose relation to therapy resistance is poorly understood. In this study, we revealed that DNA-dependent protein kinase (DNA-PK), which plays a crucial role in DNA double strand break repair, would be involved in regulation of hypoxia inducible factor-1 (HIF-1). HIF-1beta-deficient cells showed constitutively reduced expression and DNA-binding activity of Ku, the regulatory subunit of DNA-PK. Under hypoxic condition, the expression and activity of DNA- PK were markedly induced with a concurrent increase in HIF-1alpha expression. Our result also demonstrated that DNA-PK could directly interact with HIF-1, and especially DNA-PKcs, the catalytic subunit of DNA-PK, could be involved in phosphorylation of HIF-1alpha, suggesting the possibility that the enhanced expression of DNA- PK under hypoxic condition might attribute to modulate HIF 1alpha stabilization. Thus, the correlated regulation of DNA-PK with HIF-1 could contribute to therapy resistance in hypoxic tumor cells, and it provides new evidence for developing therapeutic strategies enhancing the efficacy of cancer therapy in hypoxic tumor cells. PMID- 15272236 TI - Growth impairment of primary chondrocyte cells by serum of rats with chronic renal failure. AB - Insulin-like growth factor (IGF)/IGF binding protein (IGFBP) abnormalities may be important in the pathogenesis of growth failure in chronic renal failure (CRF). We induced experimental CRF by 5/6 nephrectomy in Sprague Dawley rats (100 g) and observed for 2 weeks comparing with sham-operated pair-fed control rats (Sham- C). CRF rats gained 30% less height than Sham- C rats (P < 0.01). Serum IGFBP profiles by Western ligand blot revealed that IGFBP4 was elevated two fold in CRF rats (P < 0.01 vs. Sham-C). However, IGFBP4 mRNA levels in liver or skeletal muscle were not different in two groups. To determine if the increase of serum IGFBP4 in CRF retarded the growth of cartilage, epiphyseal chondrocytes were isolated from CRF or control rats and cultured in the presence of control or CRF rat sera. Incubation with 10% CRF serum reduced proliferations of normal chondrocytes and L6 rat skeletal muscle cells. In contrast, 10% CRF serum did not inhibit the growth of CRF chondrocytes. Rat sera from two groups were separated into two different fractions, high (>10 kDa, containing IGFBPs) and low (<10 kDa, containing free IGF) molecular weight fractions using a gel filtration column. Both fractions obtained from CRF sera decreased the growth of control chondrocytes up to 40% compared with those from control sera. We suggest that the pathogenesis of growth failure in CRF may be involved in the increase of circulating IGFBP4 as well as the unidentified small molecular weight uremic serum factors which block the growth of chondrocytes in growth plate. PMID- 15272237 TI - Gene expression in uremic left ventricular hypertrophy: effects of hypertension and anemia. AB - Hypertension and anemia may be causes of left ventricular hypertrophy (LVH) in uremia but the molecular mechanism is not known. Uremia was induced in male Spraugue Dawley rats by 5/6 nephrectomy. The following groups of rats were studied for 6 weeks; uremic rats (U) fed ad. lib., control rats (C) pair-fed with U, U rats given hydralazine (100 mg/kg/day) (UH), U rats given erythropoietin (48 U/kg/week, i.p.) (UE). Both diastolic and mean arterial pressures are higher (P < 0.01) in U and UE compared with C whereas both pressures in UH were normalized. Hemoglobin in U was lower than in C, and was normalized in UE. U, UH and UE had higher heart weight/body weight ratios (HW/BW) as well as left ventricular weight/body weight ratios (LV/BW) compared with C (P < 0.01). Compared with U, UH has lower HW/BW and LV/BW (P < 0.05) and UE has normal HW/BW but lower LV/BW than U (P < 0.05). To see if the gene expression in uremic LVH is similar to that described in pressure overload LVH in which mRNA levels of angiotensin converting enzyme (ACE), transforming growth factor-beta1 (TGF-beta1), atrial natriuretic factors (ANF) and skeletal a- actin were increased, we measured these mRNA levels by Northern analysis. TGF-beta1, ACE and alpha-actin mRNA levels were not changed in all 4 groups. ANF mRNA in U and UE was increased 3 fold over C, and normalized in UH. Treatment of anemia with erythropoietin improved uremic LVH but did not change ANF mRNA; whereas treatment of hypertension with hydralazine normalized ANF mRNA but did not completely correct uremic LVH. Thus, gene expression in uremic LVH is distinct from that in pressure-overload LVH, suggesting that other unidentified factor(s) might be involved in uremic LVH. PMID- 15272238 TI - Paradoxical effects of elastase inhibitor guamerin on the tissue repair of two different wound models: sealed cutaneous and exposed tongue wounds. AB - Innate elastase inhibitors are known to be putatively involved in the regulation of tissue inflammation by inhibiting polymorphonuclear leukocyte (PMN) derived proteinases. The aim of this study was to evaluate affects of leukocyte elastase suppression and PMN infiltration on wound healing in mouse by administering the recombinant elastase inhibitor guamerin (rEIG) in two different wound models; 1) impaired pin-punctured dorsal mucosa of anterior tongue wound, 60 mice, treated with saline containing rEIG that were fed ad libitum and 2) stable linear excisional cutaneous wound, 40 mice, covered with fibrin sealant containing rEIG. The progress of healing was analyzed by histological methods. The tongue wounds treated with rEIG became edematous around the pin-punctured tongue wound, and influx of inflammatory cells and PMN into the underlying stromal tissue were seen rapidly after wounding and peaked between 2-4 days. Whereas the control mice showed almost no wheal formation in the pin-punctured wound, a far lesser levels of PMN infiltration, and almost complete wound closure in 4 days. In the other model, the liner excisional cutaneous wound treated with fibrin sealant containing rEIG showed early wound constriction, lesser degree of inflammatory cells influx, and complete reepithelialization in 4-5 days, whereas the wound of control mice with the fibrin sealant alone showed contrary delayed reepithelialization, greater degree of inflammatory cell infiltration, and consequencial formation of greater granulation tissue at wound site. Taken together, these data suggest paradoxical effects of rEIG on the wound healing where in the wound exposed to infiltrating milieu of microorganisms in the oral cavity, the rEIG aggravates the wound healing by interfering with other innate defensive factors and extended greater flux of PMNs to inflamed wound site, while in the wound enclosed by fibrin, the rEIG accelerated wound healing by inhibiting the inflammation-generated proteases and the acute inflammatory reaction. PMID- 15272239 TI - Molecular technology and the recombinant TSH have changed diagnostics of thyroid carcinoma with positive I-131 whole body scan but low serum thyroglobulin. AB - The early detection of recurrent differentiated thyroid carcinoma (DTC) cells in the post surgery DTC patients relies on the sensitivity of measuring both the level of thyroglobulin (Tg) and 131-Iodine distribution by Whole Body Scan (WBS). Undetectable level of Tg associated with negative WBS or elevated levels of Tg associated with positive WBS ("concordant") is ordinarily indicative of either absence or presence of disease. At times, elevated level of Tg with negative WBS or low levels of Tg with positive WBS ("discordant") could also occur. In the present study, we retrospectively reviewed series of 573 patients with DTC followed in the Diagnostic Imaging and Radiotherapy of the University "Federico II" of Naples between 1993 and 1997. We focused on 9 out of 573 patients (1.56%) who had a discordant pattern with low level of Tg/positive WBS in the post surgical follow-up. Four patients were metastatic at presentation while 5 patients with metastasis during follow-up still remained in persistently low levels of Tg (<5 ng/mL). This result does point to some flaw in the evaluation of "discordant" cases. Reviewing data previously described series by resetting cut off values of Tg <1 ng/ml as undetectable changed the apparent "discordant" subgroup of patients into "concordant". Recent introduction of recombinant human TSH (rhTSH) to enhance the expression level of Tg brought significant increase in the sensitivity of diagnostic evaluation of thyroid cancer patients. The role of burdensome WBS in the follow up evaluation of DTC patients is significantly reduced over time especially in low-risk patients while the relevance of Tg assay is steadily increased. Sensitive Tg assays, significantly improved our ability to assess disease status in follow-up of DTC. Given the possibility of late disease relapses, the need for long-term follow-up, and reduced delay in treatment of persistent disease, there is still need for greater sensitive diagnostic tools for DTC. PMID- 15272240 TI - Molecular variations in Th1-specific cell surface gene Tim-3. AB - The family of T-cell immunoglobulin domain and mucin domain (TIM) proteins is identified to be expressed on T cells. A member of Tim family, Tim-3 (T cell immunoglobulin mucin 3) is selectively expressed on the surface of differentiated Th1 cells. Tim-3 might have an important role in the induction of autoimmune diseases by regulating macrophage activation and interacts with Tim-3 ligand to regulate Th1 responses. To determine the variation sites in the coding and promoter region of human Tim-3 gene, we performed variation scanning by direct sequencing using the genomic DNA isolated from the patients with asthma or allergic rhinitis and healthy controls without asthma and allergic rhinitis. We identified four single nucleotide polymorphisms (SNPs) including one novel SNPs ( 1541C>T) and two variation sites (-1292_-1289delTAAA and -1282_-1278dupTAAAA) in the coding and promoter region of human Tim-3 gene in both the patients and healthy groups. PMID- 15272241 TI - Real-Time PCR analysis of af4 and dek genes expression in acute promyelocytic leukemia t (15;17) patients. AB - Among several newly identified oncogenes, dek and af4 are attractive targets for researchers interested with leukemia. In this study quantitative Real-Time RT-PCR technique was used to define alterations in expression of dek and af4 genes associated with acute promyelocytic leukaemia (APL) t (15; 17). RNA samples obtained from bone marrow aspirates of fourteen APL patients, cDNA portions were labelled with Syber Green 1 dye and LightCycler analysis have been performed. Expression changes in patients were found not significant in comparison to healthy donors for af4 (P = 0.192) and dek (P = 0.0895). We suggest that af4 gene may have a role in leukomogenesis restricted to lymphoblastic lineage; also further studies must carry on with a larger series of patients in order to understand the relationship between the dek gene and APL. Our study was the first attempt for analysing dek and af4 genes in APL t (15; 17) patients by quantitative Real-Time RT-PCR. This rapid and sensitive method could be used to screen these genes in different types of leukaemia. PMID- 15272242 TI - Markers of beta cell function in type 1 diabetes mellitus. AB - Type 1 diabetes mellitus is a multifactorial autoimmune disease characterized by destruction of insulin producing pancreatic beta cells that results in insulin deficiency and fasting hyperglycemia. It is now well known that the clinical onset of the disease represents the end stage of an immunological process that occurs over a course of months to years. During this period the presence of autoantibodies against different islet antigens can be detected by the use of standardized assays. The rate of beta cell loss is quite variable among different individuals and at onset ketoacidosis represents still a life threatening complication of the disease. The Diabetes Control and Complication Trial (DCCT) has clearly shown that the preservation of beta cell function in type 1 diabetic subjects results in a better metabolic control and significantly reduces the risk of microvascular complications. Consequently, markers of beta cell function represent important tools to make an early diagnosis and to evaluate the impact of new therapies on the natural history of the disease. The present review will focus on clinical markers currently available (intravenous glucose tolerance test, i.v.GTT, oral glucose tolerance test, OGTT, basal and stimulated C-peptide) to assess the beta cell function in type 1 diabetes. PMID- 15272243 TI - Rediscovery of insulin pump treatment of childhood type 1 diabetes. AB - Currently, goals for the treatment of children and adolescents with type 1 diabetes mellitus are to achieve near normal glycemia; minimize the risks of severe hypoglycemia and excessive weight gain; optimize psychosocial functioning and quality of life (for children and their families); and prevent or delay long term microvascular complications. Continuous subcutaneous insulin infusion (CSII), or insulin pump therapy, provides a treatment option that can assist in the attainment of all of these goals in all ages of children. Insulin pump therapy provides the opportunity for greater flexibility in meal timing and content due to the convenience of its bolus delivery of insulin. Insulin pump therapy can potentially reduce the risk of exercise-related and nocturnal hypoglycemia, through the use of programmable variable basal infusion rates. In pediatric patients, usage of CSII has been demonstrated to reduce both glycosylated hemoglobin levels and frequency of severe hypoglycemia, without sacrifices in safety, quality of life, or weight gain, particularly in conjunction with the use of new insulin analogs and improvements in pump technology. Clinical studies of safety and efficacy of CSII in children and the use of continuous glucose monitoring to optimize insulin pump therapy are reviewed. PMID- 15272244 TI - Gene therapy for type 1 diabetes. New approaches. AB - Type 1 or insulin-dependent diabetes mellitus is a metabolic disease that results from the destruction of pancreatic beta-cells. Most patients receive subcutaneous insulin injections to reduce blood glucose levels. However, strict glucose control by multiple insulin injections is associated with an increased risk of hypoglycemia and weight gain, while a less strict glucose control is insufficient to prevent chronic complications such as nephropathy, neuropathy and retinopathy. Gene therapy holds a tremendous therapeutic potential to improve glycemic control by restoring endogenous insulin production. This review focuses on recent advances directed at producing insulin in an ectopic tissue as well as inducing pancreatic beta-cell neogenesis. The strategies include constitutive and promoter regulated insulin expression in the liver; increasing hepatic glucose oxidation; insulin production from intestinal cells; and islet cell neogenesis in liver and pancreas. PMID- 15272245 TI - Prophylaxis of autoimmune diabetes by antigen based immune modulation: are we there yet? AB - Autoimmune type 1 diabetes (early onset/juvenile and slow onset T1D) is a disease mediated by autoreactive T cells accompanied by autoantibodies against islet associated antigens. The genetic predisposition, the association with certain HLA alleles (DR4 and DQ8) as well as the possibility to identify high-risk subjects by autoantibody measurement, fueled hope in regards to potential prophylaxis by vaccination. Numerous preclinical studies along with accumulating evidence for the role of certain autoantigens and putative dominant epitopes created the premises of clinical trials addressing this objective. However, the disappointing outcomes of many of such trials made it clear that significant challenges to clinical application of this concept must be addressed: we do not know at this point, how to choose the most appropriate set of autoantigens, how to optimally use them as a vaccine, whether to define and enroll only a subset of recipients, how to monitor their T cell responses to the vaccine and lastly, which the end points of clinical T1D trials from an immunologic or metabolic perspective should be? Herein, we will approach some of the opportunities and obstacles associated with this intriguing strategy to fight autoimmune diabetes based on promising novel insight gained form studies with animal models. PMID- 15272246 TI - Prioritizing diabetes nutrition recommendations based on evidence. AB - Recommendations for carbohydrate, protein, dietary fat, micronutrients, and alcohol are classified according the level of available evidence based on the American Diabetes Association evidence grading system. The grading of recommendations can be used to prioritize nutrition care as those graded A are the most robust and can be emphasized first. Strong evidence suggests that the total amount of carbohydrate in meals (or snacks) is more important than the source or type. All persons with diabetes can benefit from basic information concerning carbohydrate foods, portion sizes, and amounts to select for meals. Patients on intensive (physiological) insulin therapy or insulin pumps can adjust their bolus insulin according to the amount of carbohydrate they plan to ingest. Therefore, the first priority is to identify a food/meal plan that can be used to integrate an insulin regimen into the person's lifestyle. Nutrition therapy for type 2 diabetes progresses from prevention of obesity or weight gain to improving insulin resistance to contributing to improved metabolic control. Research supports nutrition therapy as an effective therapy in reaching treatment goals for glycemia, lipids, and blood pressure. Monitoring of outcomes is essential to assess the outcomes of lifestyle interventions and/or to determine if changes in medication(s) are necessary. PMID- 15272247 TI - Role of TNF-alpha producing T-cells in bone loss induced by estrogen deficiency. AB - Many study in literature have suggested a possible role of T cells and tumor necrosis factor-alpha (TNF-alpha) in the pathogenesis of bone loss that occurs in pathological conditions, such as systemic inflammatory diseases; the molecular bases through which this phenomenon occurs and the relevance of this mechanism also in estrogen deficiency induced bone loss remain unclear. In our study we observed that TNF-alpha knock-out mice (TNF-/-), as well as transgenic mice without thymus (and therefore without mature T cell), do not lose bone after ovariectomy like observed for mice of normal genetic background (wild type, WT). Moreover, after transfer into athymic mice of T cell isolated from WT ovariectomized animals (and so stimulated by estrogen deficiency to proliferate and to produce TNF-alpha), ovariectomy recovers its ability to induce bone loss; whereas there is no change in bone density after injection into athymic mice of T cell purified from TNF-/- animals which, even if mature, are unable to produce TNF-alpha. Therefore the presence of TNF-alpha producing T-cell is essential for estrogen deficiency to influence bone metabolism. In the following study of the research group of Prof. Pacifici it has been shown that the increased activation of TNF-alpha producing T-cell in the ovariectomized mice is due to increased INF gamma levels, resulting from ovariectomy-induced enhanced secretion of IL-12 and IL-18 by macrophages. INF-gamma promotes expression in immunocompetent cells of class II transactivator (CIITA), that, up-regulating expression of the major system of histocompatibility of class II, makes the macrophages more active in antigen presentation to T-cells, which in turn start producing TNF. For the first time an immune mechanism is involved in the pathogenesis of post-menopausal osteoporosis; nevertheless the applicability of these conclusions also in humans remains still to be proved. PMID- 15272248 TI - [Lipoprotein-associated phospholipase A2: importance and perspectives]. AB - Type VII phospholipase A2 associated to low density lipoproteins (LDL), also known as platelet-activating factor acetylhydrolase, has been recently indicated as a new non traditional and independent risk factor of coronary disease. After the classification of phospholipase A2 family enzymes, a review is made of the recent physiologic and biochemical knowledges on A2 type VII phospholipase LDL lipoproteins-associated and the role developed in lipoproteins metabolism and atherogenesis. Finally, future therapeutic implications and perspectives depending on these knowledges are pointed out especially by using molecules inhibiting the activity of the enzyme in atherosclerosis therapy. The evaluation of circulating activity of the enzyme may be useful in the prevention and recognition of acute coronary syndromes. PMID- 15272249 TI - [The dependence medical index (DMI): validation and comparison with the activity daily living and the instrumental activity daily living]. AB - AIM: The aim of this study was the validation of the dependence medical index (DMI), a disability medical assessment tool in the elderly. METHODS: Study sample included 1054 subjects aged 65 and over, consecutively admitted to the University Department of Geriatric Medicine of Turin, Italy. A total of 356 of these subjects was classified as dependent to activity of daily living (ADL) and instrumental activity of daily living (IADL) scales. Some conditions causing medical dependence were detected, such as strength and/or motility impairment, incontinence, pressure sores, disturbances in speech and communication, decline in sight and/or hearing, terminal illness (death expected within 6 months), need for multiple and complex therapies, episodic disorientation, dizziness with tendency to fall, use of the wheel-chair. The relationship between dependence and the DMI was studied by discriminant analysis. A scale was created using the discriminant scores of each 15 medical indications for disability. RESULTS: The discriminant model of DMI was validated by cross-validation statistical method: its application permitted to classify correctly 73.1% of the sample. The DMI permitted to classify the dependent subjects in variable percentages: from 67% (DMI score > or =1) to 90% (DMI score > or =7). The best ratio between specificity and sensibility was for score 4 to DMI. CONCLUSION: The conclusion is drawn that DMI can be used to detect and evaluate the disability for medical reasons in elderly people. PMID- 15272250 TI - Plasmodium falciparum parasitized red blood cells modulate the production of endothelin-1 by human endothelial cells. AB - AIM: Plasmodium falciparum (P. falciparum) malaria is the most important parasitic infection of humans, responsible for about 2,000,000 deaths every year. Cytoadherence of P. falciparum parasitized erythrocytes (pRBC) to vascular endothelium contributes to the pathogenesis of severe malaria causing microcirculatory obstruction and subsequent tissue hypoxia. Several cytokines and vasoactive mediators are involved in this process. The aim of this paper was to investigate the production of endothelin-1 (ET-1), a potent vasoconstrictor agent, by endothelial cells from large vessels (human umbilical vein endothelial cells, HUVEC) or the microvasculature (human microvascular endothelial cells, HMEC-1), co-cultured with different strains of P. falciparum pRBC under normoxic or hypoxic conditions. METHODS: HMEC-1, immortalized by SV 40 large Tontigen, were maintained in MCDB 131 medium supplement ed with 10% fetal calf serum, 10 ng/ml of epidermal growth factor, 1 microg/ml of hydrocortisone, 2 mM glutamine, 100 U/ml of penicillin, 100 microg/ml of streptomycin and 20 mM Hepes buffer. The levels of ET-1 in the supernatants were measured by immunoenzymatic assay. RESULTS: The results indicated that IL1-beta and hypoxia were able to induce ET-1 production by both HUVEC and HMEC-1. However, the co-incubation of HUVEC or HMEC 1 with pRBC induced a dose-dependent decrease of both constitutive and IL1- or hypoxia-induced ET-1 production. The inhibition was independent from the parasite strain used and from the origin of endothelial cells. CONCLUSION: These results show that pRBC by modulating both constitutive and stimulated ET-1 release from endothelial cells can induce modifications of the vascular tone in different anatomical districts. This could be of relevance in the pathogenesis of severe malaria. PMID- 15272251 TI - "Falling leaves": a survey of the history of apoptosis. AB - Cell death has long been defined using morphological criteria. A first important concept, "necrosis", was early identified by Areteo from Cappadocia and by Galen. The term apoptosis was introduced by Kerr in 1972 to indicate a particular form of death in which cells commit suicide by chopping themselves into membrane bounded apoptotic bodies. Apoptosis is distinguished from necrosis, or accidental cell death, which is characterized by nuclear autolysis and cell disintegration. The aim of this study was an evaluation of the concepts of apoptosis and necrosis, starting from the first definition of cell death by Rudolph Virchow in 1859. In recent years substantial progress has been made in the understanding of apoptotic and necrotic cell death. In particular, cell death researchers have evolved a paradigm change, from one in which apoptosis and necrosis were considered distinct forms of cell demise, to one in which the 2 cell deaths share common features, as an integral part of a same cell death process. Since pure apoptosis and necrosis are only extremes in a continuum spectrum of aponecrotic response, a mixture of features associated with both apoptosis and necrosis represents the more typical tissue and cell response to damaging stimuli. PMID- 15272252 TI - Efficacy of tizanidine hydrochloride in the treatment of myofascial face pain. AB - AIM: The aim of the present investigation was to evaluate the usefulness of tizanidine hydrochloride in the treatment of myofascial pain of the masticatory muscles. METHODS: This work is a preliminary report of clinical experience with the use of tizanidine hydrochloride at the Section of Prosthetic Dentistry, Department of Neuroscience, University of Pisa, Pisa, Italy. During the period from January 2000 to March 2003, 145 patients were given Research Diagnostic Criteria for Temporomandibular Disorders (RDC/TMD) axis I group I diagnosis of myofascial pain in the absence of any painful temporomandibular joint condition. Seventy-eight subjects agreed to suspend any kind of treatment they had undergone and to start assuming tizanidine hydrochloride (SIRDALUD) 4 mg/die per os (2 2- mg tablets a day, 1 in the morning and 1 after dinner) for 2 weeks. All underwent a clinical assessment according to RDC/TMD guidelines at baseline time and were re-evaluated at the end of the treatment period. RESULTS: At the end of the treatment period all patients had improved; 42/78 patients (53.8%) showed absence of clinical symptoms; 18/78 (23.1%) showed a good improvement, still presenting a low number of painful sites, but not satisfying RDC/TMD parameters for diagnosis of myofascial pain; 18/78 (23.1%) showed only a slight improvement, still presenting a high number of painful sites and satisfying RDC/TMD parameters for diagnosis of myofascial pain. CONCLUSION: Given the absence of papers on the use of tizanidine hydrochloride in the treatment of myofascial pain of the masticatory muscles, the present investigation could provide some preliminary data about its possible efficacy. Randomized and controlled clinical trials are needed to confirm these results. PMID- 15272253 TI - Irritable bowel syndrome, functional dyspepsia, and functional abdominal pain syndrome. AB - Recurrent or chronic abdominal pain is a description and not a diagnosis. The clinician should consider both disease and functional pain. In the absence of obvious disease, adolescents fulfilling symptom-based criteria for functional gastrointestinal disorders can be treated for their problems without initially performing extensive diagnostic studies. Most of these patients will have symptoms resembling IBS, functional dyspepsia, or functional abdominal pain syndrome. It is imperative that the clinician takes a biopsychosocial approach in dealing with these patients. Although the clinician still evaluates for biologic disease, he or she maintains an appreciation that psychosocial events may have a profound impact on physiology and symptom production. PMID- 15272254 TI - Gastroesophageal reflux in adolescents. AB - Gastroesophageal reflux disease (GERD) occurs in adolescents but its frequency and severity is less than in adults. Typical symptoms of heartburn and regurgitation generally do not require a diagnostic evaluation unless they are associated with alarm signs including odynophagia, dysphagia, upper gastrointestinal bleeding, weight loss, atypical chest pain, or respiratory disease. Empiric treatment with proton pump inhibitors (PPIs) provides relief in most patients. Patients with persistent symptoms requiring PPI therapy should undergo endoscopy. Those with chronic GERD require medical or surgical therapy, whereas those with nonerosive reflux disease often benefit from changes in lifestyle or intermittent, on-demand medical therapy with a therapeutic aim of symptom relief. Surgical therapy is rarely required but may have a role in adolescents with respiratory complications of gastroesophageal reflux or neurologic handicap. PMID- 15272255 TI - Adolescent constipation: evaluation and management. AB - It is important for physicians who deal with adolescents to become familiar with the multiple facets of the symptom of constipation. Constipation is not a life threatening problem; however, affected patients may suffer a great deal of angst and have their lives disrupted, especially when it is left untreated. A thorough history and physical examination will uncover the majority of predisposing problems that may contribute to constipation. Most adolescents are adequately treated by appropriate behavioral and lifestyle changes and pharmacologic interventions. Failure of treatment despite good compliance requires reconsideration of the differential diagnosis, further evaluation, and possible referral to the subspecialist. PMID- 15272256 TI - Helicobacter pylori in children and adolescents. AB - There is now considerable evidence that suggests that the H. pylori organism isa human pathogen. The strong association between H. pylori and gastroduodenal disease is well documented. A number of hypotheses have been suggested for the pathogenic mechanisms of H. pylori-induced gastroduodenal disease, including the presence of bacterial virulence factors, the production of inflammatory mediators, disregulation of acid secretion, and the host immune response. At the present time, treatment with a combination of a proton pump inhibitor and antimicrobial agents continues to be recommended for the treatment of H. pylori associated peptic ulcer disease. PMID- 15272257 TI - Inflammatory bowel disease. AB - Crohn's disease (CD) and ulcerative colitis (UC), collectively known as inflammatory bowel disease (IBD) are idiopathic, life-long, destructive chronic inflammatory conditions of the gastrointestinal tract that typically manifest during late childhood and adolescence. These chronic relapsing diseases may have devastating effects on patients. New medical progress in IBD includes genetics, gut ecology and microflora, immune mechanisms, and targeted biologic therapies. This article reviews the current understanding of the etiopathogenesis of IBD, the emerging epidemiologic data in pediatric IBD, clinical presentations, diagnostic evaluation, distinctions between adult and pediatric-onset disease, and a comprehensive review of both conventional and new therapies, highlighting age-specific issues such as growth, sexual delay, and psychological and behavioral health. PMID- 15272258 TI - Celiac disease. AB - Childhood celiac disease--gluten-sensitive enteropathy--is defined in its most salient form by malabsorption and disturbed growth in association with a specific histologic lesion of the small intestine. Celiac disease occurs in response to grain consumption in susceptible individuals. Although subtotal villus atrophy with crypt hyperplasia has, in the past, been considered essential for the diagnosis, a spectrum of histologic lesions are now appreciated. The advances in the field of diagnosis (specifically, the recent development of serologic diagnostic tests) have been instrumental in correlating the histopathology of patients with subtle clinical features of the disease. Through observations, an expansion of the clinical findings associated with the condition has evolved that has helped to unfold the pathophysiology. As a result, celiac disease is now recognized as an autoimmune enteropathy of the small intestine with a broad spectrum of clinical manifestations. PMID- 15272259 TI - Allergic bowel disease. AB - The allergic bowel diseases of the adolescent have been traditionally lumped under the diagnosis of allergic or eosinophilic gastroenteritis. Over the past 20 years, clinical criteria have been established to distinguish three distinct clinical syndromes: eosinophilic esophagitis, eosinophilic gastroenteritis, and eosinophilic ascites. Each has a characteristic infiltration of eosinophils in, respectively,the esophagus only, the stomach, small bowel, and/or large bowel, or the serosal surface of the bowel. These conditions are distinguished by clinical presentation, diagnostic features, and treatment alternatives. Teenagers with these conditions may present to their pediatrician, family physician, allergist, or gastroenterologist, so each physician must appreciate the extent of appropriate diagnostic investigations and the relative value of dietary or anti inflammatory therapy. PMID- 15272260 TI - Gastrointestinal polyps and polyp syndromes in adolescents. AB - Although gastrointestinal polyps are more common in the first decade of life than during adolescence, underlying genetic polyposis syndromes are more likely in adolescents. In the past decade, the discovery of gene defects associated with polyposis syndromes has improved classification of these disorders, assisted in the stratification of cancer risk, and permitted more precise diagnosis. Genetic testing is now clinically available for the gene defects that occur in familial adenomatous polyposis coli, Peutz-Jeghers syndrome, Cowden syndrome, and juvenile polyposis syndrome. This review outlines clinical features, genetics, and management strategies for the major polyposis syndromes that affect adolescents. PMID- 15272261 TI - Autoimmune hepatitis. AB - Autoimmune hepatitis affects all ages with a peak incidence in preadolescent girls. The pathogenesis of autoimmune hepatitis has not been defined. Susceptibility, clinical manifestations, and treatment outcomes are affected by environmental factors, individual immunoregulatory responses, genetic factors, age and gender. An international panel has developed diagnostic criteria for autoimmune hepatitis, and a scoring system to assess the strength of the diagnosis has also been proposed. This article discusses the diagnosis and treatment of the three types of autoimmune hepatitis proposed based on characteristic autoantibody profiles, as well as de novo autoimmune hepatitis, a new type of autoimmune hepatitis that has been recently described in liver transplant recipients without previous autoimmune disease. PMID- 15272262 TI - Chronic viral hepatitis. AB - Although less common in childhood, hepatitis B virus (HBV) and hepatitis C virus (HCV) remain the most common causes of chronic hepatitis in the United States and worldwide. Children with chronic HBV or HCV are often asymptomatic, with normal or mildly elevated serum transaminases. Although chronic HBV and HCV are indolent diseases in childhood, they cause significant morbidity and mortality later in life. Because the dreaded complications of chronic HBV and HCV--cirrhosis with liver failure and hepatocellular carcinoma--can be seen in childhood, routine follow-up with a pediatric gastroenterologist or hepatologist is recommended. The most important role of the primary care physician and pediatric gastroenterologist or hepatologist is prevention of chronic viral hepatitis through education and screening programs. PMID- 15272263 TI - Nonalcoholic fatty liver disease. AB - Nonalcoholic fatty liver disease (NAFLD) is a spectrum of disorders that encompasses simple hepatic steatosis and the more serious nonalcoholic steatohepatitis (NASH) that can progress to cirrhosis. Although the prevalence of NAFLD in childhood is not clear, it is apparently more common than originally thought. The major association with NAFLD is obesity, and as the prevalence of obesity in childhood and adolescence increases, fatty liver is recognized with greater frequency. Although the factors associated with progression of liver disease have not been determined fully, the pathogenesis of NASH is a "two hit" process that includes disturbed lipid homeostasis, resistance to the effects of insulin and subsequent hyperinsulinemia, and local toxic effects of triglyceride on hepatocytes. Treatment options are currently limited. PMID- 15272264 TI - Wilson's disease and hemochromatosis. AB - Wilson's disease (WD) and hereditary hemochromatosis (HH) are two inherited disorders with potentially devastating and life-threatening complications. Their eminent treatability makes diagnosis in adolescence or young adulthood critical. WD is the result of abnormal copper homeostasis, causing copper overload and end organ damage. Chelation therapy can be highly efficacious in preventing manifestations of WD. HH is caused by inappropriate absorption of dietary iron, typically as the result of a specific mutation, C282Y, in the HFE gene. End-organ disease from iron accumulation is protean and includes progressive damage of the liver, pancreas, skin, heart, and pituitary. It is important to permit therapeutic phlebotomy to commence before the onset of complications. PMID- 15272265 TI - Exploring the mind field. PMID- 15272266 TI - Molecular pathways to neurodegeneration. AB - The molecular bases underlying the pathogenesis of neurodegenerative diseases are gradually being disclosed. One problem that investigators face is distinguishing primary from secondary events. Rare, inherited mutations causing familial forms of these disorders have provided important insights into the molecular networks implicated in disease pathogenesis. Increasing evidence indicates that accumulation of aberrant or misfolded proteins, protofibril formation, ubiquitin proteasome system dysfunction, excitotoxic insult, oxidative and nitrosative stress, mitochondrial injury, synaptic failure, altered metal homeostasis and failure of axonal and dendritic transport represent unifying events in many slowly progressive neurodegenerative disorders. PMID- 15272267 TI - Protein aggregation and neurodegenerative disease. AB - Neurodegenerative diseases such as Alzheimer's disease (AD), Parkinson's disease (PD), Huntington's disease (HD), amyotrophic lateral sclerosis (ALS) and prion diseases are increasingly being realized to have common cellular and molecular mechanisms including protein aggregation and inclusion body formation. The aggregates usually consist of fibers containing misfolded protein with a beta sheet conformation, termed amyloid. There is partial but not perfect overlap among the cells in which abnormal proteins are deposited and the cells that degenerate. The most likely explanation is that inclusions and other visible protein aggregates represent an end stage of a molecular cascade of several steps, and that earlier steps in the cascade may be more directly tied to pathogenesis than the inclusions themselves. For several diseases, genetic variants assist in explaining the pathogenesis of the more common sporadic forms and developing mouse and other models. There is now increased understanding of the pathways involved in protein aggregation, and some recent clues have emerged as to the molecular mechanisms of cellular toxicity. These are leading to approaches toward rational therapeutics. PMID- 15272268 TI - Potential role of presenilin-regulated signaling pathways in sporadic neurodegeneration. AB - Neurodegenerative diseases can be genetic or sporadic in origin. Genetic analysis has changed the study of the pathogenesis of these disorders by showing the causative functions of rare mutations. Yet, in the most common age-associated neurodegenerative diseases such as Alzheimer's and Parkinson's diseases, the causes of neurodegeneration remain to be clarified. The observations that presenilin modulates proteolysis and turnover of several signaling molecules have led to speculations that pathways that are important in development may contribute to neurodegeneration. In this article, the possibility that these presenilin-regulated molecules may contribute to neurodegeneration is reviewed. PMID- 15272269 TI - Stem cell therapy for human neurodegenerative disorders-how to make it work. AB - Recent progress shows that neurons suitable for transplantation can be generated from stem cells in culture, and that the adult brain produces new neurons from its own stem cells in response to injury. These findings raise hope for the development of stem cell therapies in human neurodegenerative disorders. Before clinical trials are initiated, we need to know much more about how to control stem cell proliferation and differentiation into specific phenotypes, induce their integration into existing neural and synaptic circuits, and optimize functional recovery in animal models closely resembling the human disease. PMID- 15272270 TI - Genetic clues to the pathogenesis of Parkinson's disease. AB - Recent years have seen an explosion in the rate of discovery of genetic defects linked to Parkinson's disease. These breakthroughs have not provided a direct explanation for the disease process. Nevertheless, they have helped transform Parkinson's disease research by providing tangible clues to the neurobiology of the disorder. PMID- 15272271 TI - The controversial protein-only hypothesis of prion propagation. AB - Prion diseases are some of the most intriguing infectious disorders affecting the brains of humans and animals. The prevalent hypothesis proposes that the infectious agent is a misfolded protein that propagates in the absence of nucleic acid by transmission of its altered folding to the normal host version of the protein. This article details the evidence for and against the prion hypothesis, including results of recent studies in yeast, in which a prion phenomenon has also been identified. The evidence in favor of the prion model is very strong, but final proof-consisting of the generation of infectious prions in vitro-is still missing. PMID- 15272275 TI - Microsatellite instability in gastric MALT lymphoma. AB - The role of microsatellite instability and defects in DNA mismatch repair mechanism in the pathogenesis of gastric lymphoma of mucosa-associated lymphoid tissue (MALT) type is still controversial, as both negative and positive findings have been reported. This may be explained mainly by arbitrary selection of the tested loci, the use of various techniques of microsatellite instability analysis and by different definitions of replication error positive phenotype. The aim of our study was to evaluate the instability at selected microsatellite markers using the GeneScan Analysis Software. DNA from paraffin-embedded tissue blocks of 13 previously untreated patients with localized gastric MALT lymphoma was extracted. Five microsatellite markers, which are located in hMSH2, hMLH1, P16, APC and MLL loci, were selected from the genetic database. We found genetic instability in tumors of 9/13 patients with gastric MALT lymphoma (69%). Seven of them had replication-error-positive phenotype (54%). Microsatellite instability was found in 39% of the samples in the MLL locus, 39% in the APC, 46% in the P16, 23% in the hMLH1 and none in the hMSH2. This study demonstrates that microsatellite instability has more prominent role in pathogenesis of gastric MALT lymphoma than reported to date. We suggest that microsatellite instability should be analyzed with markers adjacent to chromosomal loci that are involved in lymphomas. Our findings support the 'Real Common Target genes' theory of high rate of microsatellite instability in specific genes, which are associated with related tumors. PMID- 15272276 TI - Chronic idiopathic myelofibrosis: independent prognostic importance of bone marrow microvascular density evaluated by CD105 (endoglin) immunostaining. AB - Microvascular density (MVD) is substantially increased in bone marrow biopsies of patients with chronic idiopathic myelofibrosis (CIMF). CD105, a useful molecule for assessing MVD in various malignancies, is preferentially expressed by recently formed microvessels. Increased serum-soluble CD105 in patients with chronic myeloproliferative disorders, including CIMF, was documented. CD105 MVD has not so far been investigated in CIMF: to this end, the results in 55 patients with CIMF and 21 controls were compared with the conventional CD34 immunostaining as well as traditional histological and clinical disease features. The MVD mean values estimated by both CD105 and CD34 were significantly higher in CIMF patients than in controls (P<0.00001). In addition, the proportion of CD105 positive megakaryocytes was significantly higher in CIMF than in controls (P<0.0001). A degree of reticulin fibrosis >2 correlated with increased CD105 MVD (P=0.05). A multivariate analysis confirmed that CD105-positive MVD was an independent adverse prognosticator. This study demonstrates that while MVD, as assessed by both CD34 and CD105 immunostaining, is significantly increased in CIMF, only CD105-determined MVD correlates with the degree of fibrosis and is prognostically relevant. These findings provide a rationale for the investigational use of anti-CD105-targeted drugs in CIMF. PMID- 15272277 TI - Progesterone receptor by immunohistochemistry and clinical outcome in breast cancer: a validation study. AB - Progesterone receptor is a surrogate marker of estrogen receptor activity in breast cancer and its utility in helping predict clinical outcome has been established using biochemical assays. However, most laboratories worldwide have switched to immunohistochemistry to assess progesterone receptor, but unfortunately no validated immunohistochemical assay exists for progesterone receptor. The purpose of this study was to develop and validate an immunohistochemical assay for progesterone receptor in breast cancer. The assay was based on monoclonal antibody 1294 (DakoCytomation) and slides were scored microscopically using the 'Allred score' on a scale of 0-8. The assay was compared to ligand-binding assay in 1235 breast cancers, and a subset (n=362) that received only hormonal therapy was used to define a cutoff for progesterone receptor-positive. Clinical utility was validated in an independent set of samples (n=423) from a clinical trial randomizing premenopausal breast cancer patients to tamoxifen+oophorectomy vs observation following surgery. A cutoff of >2 (corresponding to >1% positive cells) dichotomized patients with significantly better or worse clinical outcome (P=0.0014). Progesterone receptor by immunohistochemistry provided significantly better results than progesterone receptor by ligand-binding assay in predicting clinical outcome. In the clinical trial, a positive result in univariate analyses was associated with significantly improved disease-free and overall survival both in untreated (hazard ratios/P=0.656/0.060 and 0.479/0.005, respectively) and hormonally treated patients (hazard ratios/P=0.529/0.017 and 0.451/0.007, respectively). Positive progesterone receptor remained significant for improved disease-free and overall survival (hazard ratios/P=0.666/0.038 and 0.549/0.007, respectively) in multivariate analyses including the standard variables of tumor size, nodal status, treatment, histological grade, and HER-2/neu status. Estrogen and progesterone receptors are codependent variables and progesterone receptor was a weaker predictor of response to endocrine therapy than estrogen receptor when both were included in multivariate analysis. This is the first comprehensive study assessing the clinical usefulness of progesterone receptor by immunohistochemistry in archival tissue in breast cancer. Progesterone receptor assessed by immunohistochemistry provides useful information about clinical outcome and it is better than progesterone receptor measured by ligand-binding assay. PMID- 15272278 TI - Expression of cancer-testis antigens in endometrial carcinomas using a tissue microarray. AB - Cancer-testis (CT) antigens are expressed in a variety of malignant tumors, but in normal adult tissue, they are only expressed in testicular germ cells. Owing to this tumor-associated expression pattern, these antigens are of major interest as potential targets for immunotherapy and possibly for diagnostic purposes. This study was performed to analyze the expression of four CT antigens, NY-ESO-1, MAGE A3, MAGE-A4, and CT7/MAGE-C1, in endometrial carcinoma using immunohistochemistry, and to correlate expression with histologic subtypes, grade, and expression of WT1 and p53. Formalin-fixed paraffin-embedded tissues of 130 endometrial carcinomas of the following types and grades were analyzed using a tissue microarray: 85 endometrioid carcinomas (FIGO grade 1, 39; grade 2, 11; and grade 3, 35), 18 papillary serous carcinomas, 12 clear cell carcinomas, 13 malignant mixed mullerian tumors, one mucinous adenocarcinoma, and one undifferentiated carcinoma. The following anti-CT monoclonal antibodies/antigens were studied by immunohistochemistry: monoclonal antibody ES121/NY-ESO-1, monoclonal antibody M3H67/MAGE-A3, monoclonal antibody 57B/MAGE-A4, and monoclonal antibody CT7-33/CT7. The CT expression data were compared to WT1 and p53 protein expression as analyzed in a previous study. Positive staining with anti-CT monoclonal antibodies was graded as follows: focal, <5% positive cells; 1+, 5-25% cells; 2+, 26-50% cells; 3+, 51-75%; and 4+, >75% cells. The 3+ and 4+ staining patterns were considered homogeneous patterns of potential clinical significance and were scored positive for statistical analysis. In low-grade tumors, the most immunoreactivity was seen with mAb M3H67 but little labeling was observed with the other monoclonal antibodies. In high-grade tumors, monoclonal antibodies M3H67 (25%), 57B (23%), and CT7-33 (20%) showed the highest reactivity, while ES121 showed the lowest immunoreactivity (6%). The staining pattern was mostly heterogeneous. Statistical significance was found solely for the correlation of monoclonal antibody 57B staining and p53 expression. No correlation was found for any anti-CT monoclonal antibody staining and clinical stage or for anti-CT staining and WT1 expression. CT antigens CT7, MAGE-A3 and MAGE-A4, but not NY-ESO-1, are expressed in high-grade endometrial carcinomas, and expression of MAGE-A4 is correlated with the presence of overexpressed p53. PMID- 15272279 TI - Galectin-3, fibronectin-1, CITED-1, HBME1 and cytokeratin-19 immunohistochemistry is useful for the differential diagnosis of thyroid tumors. AB - The diagnosis of thyroid tumors is critical for clinical management; however, tumors with follicular architecture often present problems. We evaluated the diagnostic use of the protein expression of four genes that were found to be upregulated in papillary thyroid carcinoma compared to normal thyroid (LGALS3, FN1, CITED1 and KRT19), and of the mesothelial cell surface protein recognized by monoclonal antibody HBME1 in thyroid tumors. Tissues from 85 carcinomas (67 papillary, six follicular, eight Hurthle cell and four anaplastic) and 21 adenomas were evaluated by immunohistochemistry for the expression of these gene protein products, for example, galectin-3 (GAL3), fibronectin-1 (FN1), CITED1, cytokeratin-19 (CK19) and HBME1. Non-neoplastic thyroids (29 adenomatous and 14 thyrotoxic hyperplasia, and 59 normal) were also studied. The expression of all five proteins was significantly associated with malignancy, and highly specific (> or = 90%) for carcinoma compared to adenoma. GAL3, FN1 and/or HBME1 expression was seen in 100% of carcinomas (85/85) and in 24% of adenomas (5/21). Coexpression of multiple proteins was seen in 95% of carcinomas and only 5% of adenomas (P<0.0001). Coexpression of FN1 and GAL3 (FN1+ GAL3+, 70/85) or FN1 and HBME1 (FN1+ HBME1+, 53/85) was restricted to carcinomas, while their concurrent absence (FN1- GAL3- or FN1- HBME1-, 18/21 adenoma) was highly specific (96%) for benign lesions. Among non-neoplastic thyroids, adenomatous hyperplasia frequently expressed GAL3 (n=16), CK19 (n=9) and CITED1 (n=7), but the expression was predominantly focal in contrast to the diffuse expression in carcinomas. An immunohistochemical panel consisting of GAL3, FN1 and HBME1 may be useful in the diagnosis of follicular cell-derived thyroid tumors. PMID- 15272280 TI - Practice guidelines for the renal biopsy. AB - Biopsy of the kidney should never be undertaken without careful consideration of the risks vs benefits. Given the importance of a correct diagnosis in the treatment and prognosis of renal disease, the pathological evaluation should use all available modalities. Native kidney biopsies require examination by light microscopy, immunohistochemistry and electron microscopy. The processing of the renal biopsy is complex and requires the support of a fully equipped anatomic pathology laboratory. Technical expertise is required to process the small fragments of tissue and to produce sections of highest quality. The correct diagnosis requires a well-trained renal pathologist with thorough knowledge of not only renal pathology but also renal medicine in order to correlate intricate tissue-derived information with detailed clinical data. In view of the importance and consequences of the pathologic diagnosis, the Renal Pathology Society appointed an Ad Hoc Committee on Practice Guidelines, to define the essential ingredients necessary to provide quality renal pathology diagnoses. This document incorporates the consensus opinions of the committee and the RPS membership at large. PMID- 15272281 TI - Does HPV play a role in the etiopathogenesis of ameloblastoma? An immunohistochemical, in situ hybridization and polymerase chain reaction study of 18 cases using laser capture microdissection. AB - Ameloblastomas are epithelial tumors of odontogenic origin, biologically characterized by local recurrence. Among different etiologic factors, HPV infection has been recently postulated to be somehow involved in ameloblastoma etiopathogenesis. To address this issue, we studied 18 ameloblastomas by means of immunohistochemistry, in situ hybridization (conventional and amplified), polymerase chain reaction and nested-polymerase chain reaction analyses using laser capture microdissection in order to detect the occurrence of HPV in this setting. No evidence of HPV infection was detected by morphological examination, immunohistochemistry, in situ hybridization and conventional polymerase chain reaction, while nested-polymerase chain reaction showed a weak positive band in two cases. However, the subsequent restriction enzyme analysis carried out from the nested-polymerase chain reaction amplification products of these two samples excluded the presence of HPV subtypes 16, 18, 31, 33, 35, 52, and 58. The search for HPV 6 and 11 in the same specimens was also negative. In conclusion, our data do not support an etiopathogenetic evidence for HPV in ameloblastoma. PMID- 15272282 TI - Human papillomavirus testing as a cytology gold standard: comparing Surinam with the Netherlands. AB - Polymerase chain reaction to detect high-risk human papillomavirus has been suggested as a gold standard for cytology. The Netherlands and Surinam were prospectively compared in regard to the proportions of Negative, Atypical Squamous Cells of Undetermined Significance, and Squamous Intraepithelial Lesion smears that had detectable high-risk human papillomavirus. For the Netherlands, 14 600 negative, 270 Atypical Squamous Cells of Undetermined Significance and 120 Squamous Intraepithelial Lesion smears were evaluated by polymerase chain reaction. For Surinam, 150 negative, 50 Atypical Squamous Cells of Undetermined Significance, and 150 Squamous Intraepithelial Lesion smears were evaluated by polymerase chain reaction. In all, 4% of Dutch and 80% of Surinamese negative smears had detectable high-risk human papillomavirus (chi2=1313, P<0.00001). In total, 41.9% of Dutch and 84% of Surinamese Atypical Squamous Cells of Undetermined Significance smears had detectable high-risk human papillomavirus (chi2=28, P<0.00001). Totally, 67.5% of Dutch and 94% of Surinamese SIL smears had detectable high-risk human papillomavirus (chi2=30, P<0.00001). The Negative: Atypical Squamous Cells of Undetermined Significance odds ratio was 0.058 for the Netherlands and 0.762 for Surinam (chi2homog=31, P<0.00001). The Negative: Squamous Intraepithelial Lesion odds ratio was 0.020 for the Netherlands and 0.255 for Surinam (chi2homog=31, P<0.00001). The Atypical Squamous Cells of Undetermined Significance: Squamous Intraepithelial Lesion odds ratio was 0.347 for the Netherlands and 0.335 for Surinam (chi2homog=0.005, P>0.75). Human papillomavirus DNA testing may not be a suitable gold standard in general because its use would make specificity and sensitivity prevalence-dependent. A new statistic, the percent of Negative pap smears with detectable high-risk human papillomavirus, is posited, which may be important if human papillomavirus DNA testing is used clinically. PMID- 15272283 TI - p185HER2 overexpression and HER2 oncogene amplification in recurrent vulvar Paget's disease. AB - We evaluated p186Her2 overexpression and HER2 oncogene amplification in recurrent vulvar Paget's disease. We identified six patients with recurrent vulvar Paget's disease in our archives. The number of recurrences ranged from 1 to 11 over a time period of 1-168 months. Recurrences were evaluated immunohistochemically for p185Her2 overexpression with the HercepTest and for HER2 oncogene amplification with fluorescence in situ hybridization. p185Her2 overexpression was scored as 3 in two patients, as 2 in two patients, and as 1 in two patients. All patients with scores 2 and 3 showed HER2 oncogene amplification. Overexpression of p185Her2 and HER2 oncogene amplification appears to be common in recurrent vulvar Paget's disease. PMID- 15272284 TI - COX-2 expression correlates with VEGF-C and lymph node metastases in patients with head and neck squamous cell carcinoma. AB - Recent observations suggest an implication of the cyclooxygenase-2 (COX-2) in tumor lymphangiogenesis through an upregulation of vascular endothelial growth factor-C expression. It is unknown whether this mechanism also acts in squamous cell carcinoma of the head and neck region. We performed a retrospective study of 70 patients with head and neck squamous cell carcinoma in order to investigate whether COX-2 immunohistochemical expression correlates with vascular endothelial growth factor-C expression. We also examined the association of the expression of these molecules with clinicopathologic parameters (especially lymph node status) and outcome for these patients. We performed immunostaining on formalin-fixed, paraffin-embedded tissue sections by the routine streptavidin-biotin peroxidase labeled procedure. Increased cyclooxygenase-2 expression was observed in 30 of the 68 tumor samples (44%), while high vascular endothelial growth factor-C expression occurred in 26 of the 68 tumor samples (38%). High expression of the two proteins was correlated with the presence of lymph node metastasis at the time of diagnosis, and the observed association was even stronger when there was overexpression for both the antibodies (P<0.001). High expression of vascular endothelial growth factor-C, but not of COX-2 was correlated with increased mortality in patients with oral-larynx squamous cell carcinoma. When multivariate Cox regression model was applied, the presence of lymph node metastasis at the time of diagnosis, combined with overexpression of both the antibodies, was the only independent prognostic factor for mortality of these patients. Our results suggest that a lymphangiogenic pathway, in which COX-2 overexpression stimulates vascular endothelial growth factor-C upregulation, probably exists in head and neck squamous cell carcinoma. Also, the predictive ability for mortality of regional lymph node metastasis can be improved with the combined evaluation of the immunohistochemical expression of these two proteins. PMID- 15272285 TI - Comparative genomic hybridization analysis of thymic neuroendocrine tumors. AB - Thymic neuroendocrine (carcinoid) tumors are a rare neoplasm of the anterior mediastinum. The tumors frequently exhibit a wide spectrum of histology and appear to follow a more aggressive behavior than their nonthymic counterparts. Given the differing clinicopathologic manifestations, thymic neuroendocrine tumors may also possess different cytogenetic abnormalities from those that occur in foregut carcinoid tumors. In this study, we employed comparative genomic hybridization to detect genomic instability in 10 sporadic thymic neuroendocrine tumors and one multiple endocrine neoplasia type 1 (MEN1)-associated case. Gross chromosomal imbalances were found in nine cases, including gains of chromosomal material on regions X, 8, 18 and 20p and losses on 3, 6, 9q, 13q and 11q. We did not observe deletion at locus 11q13 where the MEN1 gene is located. These findings were essentially dissimilar to those reported in sporadic and MEN1 associated foregut carcinoid tumors. Consequently, we consider that a distinctive cytogenetic mechanism is at work in the development of thymic neuroendocrine tumors, which is different from that of foregut carcinoid tumors. PMID- 15272286 TI - Pulmonary pathology of severe acute respiratory syndrome in Toronto. AB - The severe acute respiratory syndrome (SARS) pandemic in Toronto resulted in a large number of autopsies on its victims. We describe the pulmonary pathology of patients who died in the 2003 Toronto outbreak. Autopsy material from the lungs of 20 patients who died between March and July 2003 were characterized by histology, molecular biology, and immunohistochemistry for cytokeratins, thyroid transcription factor-1, CD68, Epstein-Barr virus, cytomegalovirus, and human herpes simplex viruses. Matched controls were obtained from patients who died of other causes over the same interval. The mean duration of illness was 27 days (range 5-108 days). Post-mortem lung tissues from 19 of 20 patients with probable SARS were positive for SARS-associated coronavirus by RT-PCR. Histologically, all patients showed varying degrees of exudative and proliferative phase acute lung injury, evidenced in conventional and immunohistochemical stains by edema, inflammatory infiltrate, pneumocyte hyperplasia, fibrinous exudates, and organization. Eight of 20 patients showed predominantly a diffuse alveolar damage pattern of acute lung injury, six showed predominantly an acute fibrinous and organizing pneumonia pattern, and the remainder showed an admixture of the two patterns. Squamous metaplasia and scattered multinucleate giant cells were present in most cases. Vascular fibrin thrombi were a common finding and were often associated with pulmonary infarcts. Special stains demonstrated vascular endothelial damage of both small- and mid-sized pulmonary vessels. Two cases were complicated by invasive fungal disease consistent with Aspergillosis, and another by coinfection with cytomegalovirus. Our findings indicate that the lungs of patients who die of SARS are almost always positive for the SARS-associated coronavirus by RT-PCR, and may show features of both diffuse alveolar damage and acute fibrinous and organizing pneumonia patterns of acute injury. Cases of SARS may be complicated by coexistent infections and therapy-related lung injury. PMID- 15272287 TI - Botulinum toxin treatment for acute traumatic complete sixth nerve palsy. AB - AIMS: To investigate the benefits of botulinum toxin (BTX) injection for acute unilateral complete sixth nerve palsy caused by trauma. METHODS: We retrospectively reviewed patients treated for acute unilateral complete sixth nerve palsy caused by head injury during a 10-year period (between March 1993 and February 2003) in our hospital. The BTX treatment group was defined as patients who received BTX injection within 3 months of injury. Patients who presented within 3 months of trauma, and had no previous BTX injection or surgery were enrolled as the conservative treatment group. Comparison of the patient demographics, palsy characteristics, angle of deviations, and recovery rates were made between the two groups. RESULTS: In all, 33 patients were enrolled by our inclusion criteria. Of these, 19 patients were treated conservatively, and 14 patients were treated with BTX. A total of 79% of our patients presented with abduction deficit of grade -5. The results showed that there was no significant difference in the outcome for the two groups based on age, gender, time to presentation, severity, and initial angle of deviation. The BTX group had a higher recovery rate than the conservative treatment group (64.3 vs 26.3%, P=0.028). Among 26 patients with grade -5 abduction deficit, the recovery rate was higher in the BTX-treated patients than in the conservatively treated patients, which had no statistical significance (50 vs 18.8%, P=0.09). CONCLUSION: BTX facilitates recovery of acute traumatic complete sixth nerve palsy in severely injured patients. PMID- 15272288 TI - A review of trabeculectomy in East Asian people--the influence of race. AB - Glaucoma is the leading cause of irreversible blindness worldwide. East Asians account for approximately half of all glaucoma sufferers. It is likely that trabeculectomy will be needed for many of these people as the intraocular pressure is to be maintained at a satisfactorily low level. The eyes of East Asian people differ in some aspects from those of other races. This review describes the natural history of the eye after trabeculectomy in East Asians. PMID- 15272289 TI - Neurofibromatosis type 1 presenting with Horner's syndrome. PMID- 15272290 TI - Microscopic polyangitis presenting with sub-acute reversible optic neuropathy. PMID- 15272291 TI - Is measurement of blood pressure worthwhile in the diabetic eye clinic? AB - PURPOSE: The UK Prospective Diabetic Study has confirmed the importance of blood pressure (BP) as a major risk factor for diabetic retinopathy (DR). We wanted to investigate whether measuring the BP in the diabetic eye clinic could identify new hypertensive patients and monitor control in existing ones. PATIENTS AND METHODS: We compared BP in patients attending the diabetic eye clinic with home blood pressure measurement (HBPM) and ambulatory BP measurement (ABPM). In all, 106 patients attending a diabetic eye clinic were selected at random from clinic attendees. BP measurement (on an Omron 705 CP) was performed in the eye clinic and also compared to HBPM three times per day with an Omron 705 CP machine, and was compared to diabetic clinic measurements. In addition, 11 randomly chosen patients had 24 h ABPM to validate the above techniques. RESULTS: In all, 106 patients (70 male and 36 female) were recruited for the study, of which 71 were known to be hypertensive on antihypertensive medication. Of the total, 75 patients (70.8%) had BP > 140/85 in the eye clinic, of which 51 (68%) were known to be hypertensive on treatment and this was confirmed in 46 (90%) on HBPM. A total of, 24 patients (22.6%) were newly diagnosed as hypertensive in the eye clinic, which was confirmed by HBPM in 22 patients (92%). The mean BP of the measurements performed in the eye clinic was significantly higher than that carried out in the diabetic clinic (P < 0.01). Tropicamide 1% and phenylephrine 2.5% eye drop instillation had no effect on BP. In 11 randomly chosen patients, 24 h ABPM validated both diabetic eye clinic and home BP measurements. CONCLUSION: Attendance at the diabetic eye clinic is an important chance to detect both new patients with systemic hypertension and those with inadequate BP control. Ophthalmologists should be encouraged to measure BP in their diabetic patients attending diabetic eye clinics, as it is an important risk factor for DR. On the basis of our findings, good BP control is a goal yet to be achieved in diabetic patients with retinopathy. PMID- 15272292 TI - Severe bilateral posterior ischemic optic neuropathy as a complication of spinal surgery. PMID- 15272294 TI - Statistical analysis of agreement in measurement comparison studies. PMID- 15272295 TI - Ocular motility findings in chronic progressive external ophthalmoplegia. AB - AIMS: To characterise the ocular motility features of chronic progressive external opthalmoplegia by quantitative and semiquantitative means. To assess the prevalence of diplopia and the binocular adaptations to nonaligned visual axes. METHOD: We studied 25 patients with chronic progressive external ophthalmoplegia. In each case muscle biopsies were consistent with mitochondrial myopathy. All patients underwent cover test in the primary position, assessment of binocular status, and measurement of uniocular fields of fixation using the Goldmann perimeter. RESULTS: A total of 23 (92%) patients had an exo-deviation, with six (26%) of those having an associated vertical deviation: 12 patients were binocular. Of the 13 patients with a manifest deviation seven had diplopia and six had suppression. Of all paired extra-ocular muscles (EOM), 68% had symmetry of movement within 5 degrees of each other. CONCLUSION: Almost all patients had an exo-deviation. Diplopia was more common than expected. The majority of patients had symmetry of EOM limitation. PMID- 15272296 TI - Diode laser trans-scleral cyclophotocoagulation in the management of glaucoma in patients with long-term intravitreal silicone oil. AB - AIMS: To analyse the results of contact trans-scleral cyclodiode photocoagulation (TSCPC) in reducing intraocular pressures (IOP) refractory to medical treatment in a group of patients with intravitreal silicone oil. METHODS: The medical records of 18 patients who received TSCPC were evaluated retrospectively. Success was evaluated primarily in terms of IOP control. Success was defined as a final IOP of less than 22 mmHg. RESULTS: In all, 18 eyes of 18 patients were followed up for an average of 21.8 months (range 6-36) (SD=8.6). The mean pre-treatment IOP was 39.6 mmHg (range 25-56 mmHg) (SD=9.3). This reduced to 20.0 mmHg (range 0 48, SD=13.5) after TSCPC, producing a mean reduction of 49% from mean pretreatment IOP levels. The overall success rate was 44% (eight eyes). Chronic hypotony occurred in two patients. IOP remained above that required to meet the IOP reduction criteria in eight patients. CONCLUSION: Our results raise doubts about the efficacy of TSCPC in the management of glaucoma in eyes retaining silicone oil. This may relate to the long duration of silicone oil in our patients. Further studies are required to identify risk factors for treatment failure. PMID- 15272297 TI - Residual debris as a potential cause of postphacoemulsification endophthalmitis. PMID- 15272298 TI - HMGB1 is an endogenous immune adjuvant released by necrotic cells. AB - Immune responses against pathogens require that microbial components promote the activation of antigen-presenting cells (APCs). Autoimmune diseases and graft rejections occur in the absence of pathogens; in these conditions, endogenous molecules, the so-called 'innate adjuvants', activate APCs. Necrotic cells contain and release innate adjuvants; necrotic cells also release high-mobility group B1 protein (HMGB1), an abundant and conserved constituent of vertebrate nuclei. Here, we show that necrotic HMGB1(-/-) cells have a reduced ability to activate APCs, and HMGB1 blockade reduces the activation induced by necrotic wild type cell supernatants. In vivo, HMGB1 enhances the primary antibody responses to soluble antigens and transforms poorly immunogenic apoptotic lymphoma cells into efficient vaccines. PMID- 15272299 TI - Helicase Motif III in SecA is essential for coupling preprotein binding to translocation ATPase. AB - The SecA ATPase is a protein translocase motor and a superfamily 2 (SF2) RNA helicase. The ATPase catalytic core ('DEAD motor') contains the seven conserved SF2 motifs. Here, we demonstrate that Motif III is essential for SecA-mediated protein translocation and viability. SecA Motif III mutants can bind ligands (nucleotide, the SecYEG translocase 'channel', signal and mature preprotein domains), can catalyse basal and SecYEG-stimulated ATP hydrolysis and can be activated for catalysis. However, Motif III mutation specifically blocks the preprotein-stimulated 'translocation ATPase' at a step of the reaction pathway that lies downstream of ligand binding. A functional Motif III is required for optimal ligand-driven conformational changes and kinetic parameters that underlie optimal preprotein-modulated nucleotide cycling at the SecA DEAD motor. We propose that helicase Motif III couples preprotein binding to the SecA translocation ATPase and that catalytic activation of SF2 enzymes through Motif III-mediated action is essential for both polypeptide and nucleic-acid substrates. PMID- 15272300 TI - What's new about RNAi? Meeting on siRNAs and miRNAs. PMID- 15272301 TI - Dynamic movement of actin-like proteins within bacterial cells. AB - Actin proteins are present in pro- and eukaryotes, and have been shown to perform motor-like functions in eukaryotic cells in a variety of processes. Bacterial actin homologues are essential for cell viability and have been implicated in the formation of rod cell shape, as well as in segregation of plasmids and whole chromosomes. We have generated functional green fluorescent protein fusions of all three Bacillus subtilis actin-like proteins (MreB, Mbl and MreBH), and show that all three proteins form helical filaments underneath the cell membrane, the pattern of which is distinct for each protein. Time-lapse microscopy showed that the filaments are highly dynamic structures. A number of separate filaments of MreB and Mbl continuously move through the cell along helical tracks underneath the cell membrane. The speed of extension of the growing end of filaments is within the range of known actin polymerization (0.1 microm/s), generating a potential poleward or centreward pushing velocity at 0.24 microm/min for MreB or Mbl, respectively. During nutritional downshift and a block in topoisomerase IV activity, the filaments rapidly disintegrated, showing that movement occurs only in growing cells. Contrary to Mbl and MreBH filaments, MreB filaments were generally absent in cells lacking DNA, providing a further distinction between the three orthologues. PMID- 15272302 TI - MAPKKKalpha is a positive regulator of cell death associated with both plant immunity and disease. AB - Many plant pathogens cause disease symptoms that manifest over days as regions of localized cell death. Localized cell death (the hypersensitive response; HR) also occurs in disease-resistant plants, but this response appears within hours of attempted infection and may restrict further pathogen growth. We identified a MAP kinase kinase kinase gene (MAPKKKalpha) that is required for the HR and resistance against Pseudomonas syringae. Significantly, we found that MAPKKKalpha also regulates cell death in susceptible leaves undergoing P. syringae infection. Overexpression of MAPKKKalpha in leaves activated MAPKs and caused pathogen independent cell death. By overexpressing MAPKKKalpha in leaves and suppressing expression of various MAPKK and MAPK genes by virus-induced gene silencing, we identified two distinct MAPK cascades that act downstream of MAPKKKalpha. These results demonstrate that signal transduction pathways associated with both plant immunity and disease susceptibility share a common molecular switch. PMID- 15272303 TI - Activation of a vinculin-binding site in the talin rod involves rearrangement of a five-helix bundle. AB - The interaction between the cytoskeletal proteins talin and vinculin plays a key role in integrin-mediated cell adhesion and migration. We have determined the crystal structures of two domains from the talin rod spanning residues 482-789. Talin 482-655, which contains a vinculin-binding site (VBS), folds into a five helix bundle whereas talin 656-789 is a four-helix bundle. We show that the VBS is composed of a hydrophobic surface spanning five turns of helix 4. All the key side chains from the VBS are buried and contribute to the hydrophobic core of the talin 482-655 fold. We demonstrate that the talin 482-655 five-helix bundle represents an inactive conformation, and mutations that disrupt the hydrophobic core or deletion of helix 5 are required to induce an active conformation in which the VBS is exposed. We also report the crystal structure of the N-terminal vinculin head domain in complex with an activated form of talin. Activation of the VBS in talin and the recruitment of vinculin may support the maturation of small integrin/talin complexes into more stable adhesions. PMID- 15272304 TI - A hydrocarbon ruler measures palmitate in the enzymatic acylation of endotoxin. AB - The ability of enzymes to distinguish between fatty acyl groups can involve molecular measuring devices termed hydrocarbon rulers, but the molecular basis for acyl-chain recognition in any membrane-bound enzyme remains to be defined. PagP is an outer membrane acyltransferase that helps pathogenic bacteria to evade the host immune response by transferring a palmitate chain from a phospholipid to lipid A (endotoxin). PagP can distinguish lipid acyl chains that differ by a single methylene unit, indicating that the enzyme possesses a remarkably precise hydrocarbon ruler. We present the 1.9 A crystal structure of PagP, an eight stranded beta-barrel with an unexpected interior hydrophobic pocket that is occupied by a single detergent molecule. The buried detergent is oriented normal to the presumed plane of the membrane, whereas the PagP beta-barrel axis is tilted by approximately 25 degrees. Acyl group specificity is modulated by mutation of Gly88 lining the bottom of the hydrophobic pocket, thus confirming the hydrocarbon ruler mechanism for palmitate recognition. A striking structural similarity between PagP and the lipocalins suggests an evolutionary link between these proteins. PMID- 15272305 TI - Crystal structure of glycogen synthase: homologous enzymes catalyze glycogen synthesis and degradation. AB - Glycogen and starch are the major readily accessible energy storage compounds in nearly all living organisms. Glycogen is a very large branched glucose homopolymer containing about 90% alpha-1,4-glucosidic linkages and 10% alpha-1,6 linkages. Its synthesis and degradation constitute central pathways in the metabolism of living cells regulating a global carbon/energy buffer compartment. Glycogen biosynthesis involves the action of several enzymes among which glycogen synthase catalyzes the synthesis of the alpha-1,4-glucose backbone. We now report the first crystal structure of glycogen synthase in the presence and absence of adenosine diphosphate. The overall fold and the active site architecture of the protein are remarkably similar to those of glycogen phosphorylase, indicating a common catalytic mechanism and comparable substrate-binding properties. In contrast to glycogen phosphorylase, glycogen synthase has a much wider catalytic cleft, which is predicted to undergo an important interdomain 'closure' movement during the catalytic cycle. The structures also provide useful hints to shed light on the allosteric regulation mechanisms of yeast/mammalian glycogen synthases. PMID- 15272306 TI - CD95 ligand induces motility and invasiveness of apoptosis-resistant tumor cells. AB - The apoptosis-inducing death receptor CD95 (APO-1/Fas) controls the homeostasis of many tissues. Despite its apoptotic potential, most human tumors are refractory to the cytotoxic effects of CD95 ligand. We now show that CD95 stimulation of multiple apoptosis-resistant tumor cells by CD95 ligand induces increased motility and invasiveness, a response much less efficiently triggered by TNFalpha or TRAIL. Three signaling pathways resulting in activation of NF kappaB, Erk1/2 and caspase-8 were found to be important to this novel activity of CD95. Gene chip analyses of a CD95-stimulated tumor cell line identified a number of potential survival genes and genes that are known to regulate increased motility and invasiveness of tumor cells to be induced. Among these genes, urokinase plasminogen activator was found to be required for the CD95 ligand induced motility and invasiveness. Our data suggest that CD95L, which is found elevated in many human cancer patients, has tumorigenic activities on human cancer cells. This could become highly relevant during chemotherapy, which can cause upregulation of CD95 ligand by both tumor and nontumor cells. PMID- 15272307 TI - Structure of the bifunctional and Golgi-associated formiminotransferase cyclodeaminase octamer. AB - Mammalian formiminotransferase cyclodeaminase (FTCD), a 0.5 million Dalton homo octameric enzyme, plays important roles in coupling histidine catabolism with folate metabolism and integrating the Golgi complex with the vimentin intermediate filament cytoskeleton. It is also linked to two human diseases, autoimmune hepatitis and glutamate formiminotransferase deficiency. Determination of the FTCD structure by X-ray crystallography and electron cryomicroscopy revealed that the eight subunits, each composed of distinct FT and CD domains, are arranged like a square doughnut. A key finding indicates that coupling of three subunits governs the octamer-dependent sequential enzyme activities, including channeling of intermediate and conformational change. The structure further shed light on the molecular nature of two strong antigenic determinants of FTCD recognized by autoantibodies from patients with autoimmune hepatitis and on the binding of thin vimentin filaments to the FTCD octamer. PMID- 15272308 TI - Chk1, but not Chk2, inhibits Cdc25 phosphatases by a novel common mechanism. AB - Cdc25 phosphatases activate cyclin-dependent kinases (Cdks) and thereby promote cell cycle progression. In vertebrates, Chk1 and Chk2 phosphorylate Cdc25A at multiple N-terminal sites and target it for rapid degradation in response to genotoxic stress. Here we show that Chk1, but not Chk2, phosphorylates Xenopus Cdc25A at a novel C-terminal site (Thr504) and inhibits it from C-terminally interacting with various Cdk-cyclin complexes, including Cdk1-cyclin A, Cdk1 cyclin B, and Cdk2-cyclin E. Strikingly, this inhibition, rather than degradation itself, of Cdc25A is essential for the Chk1-induced cell cycle arrest and the DNA replication checkpoint in early embryos. 14-3-3 proteins bind to Chk1 phosphorylated Thr504, but this binding is not required for the inhibitory effect of Thr504 phosphorylation. A C-terminal site presumably equivalent to Thr504 exists in all known Cdc25 family members from yeast to humans, and its phosphorylation by Chk1 (but not Chk2) can also inhibit all examined Cdc25 family members from C-terminally interacting with their Cdk-cyclin substrates. Thus, Chk1 but not Chk2 seems to inhibit virtually all Cdc25 phosphatases by a novel common mechanism. PMID- 15272310 TI - Crystal structure of the bacterial nucleoside transporter Tsx. AB - Tsx is a nucleoside-specific outer membrane (OM) transporter of Gram-negative bacteria. We present crystal structures of Escherichia coli Tsx in the absence and presence of nucleosides. These structures provide a mechanism for nucleoside transport across the bacterial OM. Tsx forms a monomeric, 12-stranded beta-barrel with a long and narrow channel spanning the outer membrane. The channel, which is shaped like a keyhole, contains several distinct nucleoside-binding sites, two of which are well defined. The base moiety of the nucleoside is located in the narrow part of the keyhole, while the sugar occupies the wider opening. Pairs of aromatic residues and flanking ionizable residues are involved in nucleoside binding. Nucleoside transport presumably occurs by diffusion from one binding site to the next. PMID- 15272309 TI - Xenopus paraxial protocadherin has signaling functions and is involved in tissue separation. AB - Protocadherins have homophilic adhesion properties and mediate selective cell cell adhesion and cell sorting. Knockdown of paraxial protocadherin (PAPC) function in the Xenopus embryo impairs tissue separation, a process that regulates separation of cells of ectodermal and mesodermal origin during gastrulation. We show that PAPC can modulate the activity of the Rho GTPase and c jun N-terminal kinase, two regulators of the cytoskeletal architecture and effectors of the planar cell polarity pathway. This novel signaling function of PAPC is essential for the regulation of tissue separation. In addition, PAPC can interact with the Xenopus Frizzled 7 receptor, and both proteins contribute to the development of separation behavior by activating Rho and protein kinase Calpha. PMID- 15272312 TI - Morphine induces terminal micro-opioid receptor desensitization by sustained phosphorylation of serine-375. AB - Morphine is a poor inducer of micro-opioid receptor (MOR) internalization, but a potent inducer of cellular tolerance. Here we show that, in contrast to full agonists such as [D-Ala(2)-MePhe(4)-Gly-ol]enkephalin (DAMGO), morphine stimulated a selective phosphorylation of the carboxy-terminal residue 375 (Ser(375)). Ser(375) phosphorylation was sufficient and required for morphine induced desensitization of MOR. In the presence of full agonists, morphine revealed partial agonistic properties and potently inhibited MOR phosphorylation and internalization. Upon removal of the drug, DAMGO-desensitized receptors were rapidly dephosphorylated. In contrast, morphine-desensitized receptors remained at the plasma membrane in a Ser(375)-phosphorylated state for prolonged periods. Thus, morphine promotes terminal MOR desensitization by inducing a persistent modification of Ser(375). PMID- 15272313 TI - Enhanced paclitaxel cytotoxicity and prolonged animal survival rate by a nonviral mediated systemic delivery of E1A gene in orthotopic xenograft human breast cancer. AB - Paclitaxel (Taxol) is a promising frontline chemotherapeutic agent for the treatment of human breast and ovarian cancers. The adenoviral type 5 E1A gene has been tested in multiple clinical trials for its anticancer activity. E1A has also been shown to sensitize paclitaxel-induced killing in E1A-expressing cells. Here, we show that E1A can sensitize paclitaxel-induced apoptosis in breast cancer cells in a gene therapy setting by an orthotopic mammary tumor model. We first showed that expression of E1A enhanced in vitro paclitaxel cytotoxicity, as compared to the control cells. We then compared the therapeutic efficacy of paclitaxel between orthotopic tumor models established with vector-transfected MDA-MB-231 (231-Vect) versus 231-E1A stable cells, using tumor weight and apoptotic index (TUNEL assay) as the parameters. We found paclitaxel was more effective in shrinking tumors and inducing apoptosis in tumor models established with stable 231-E1A cells than the control 231-Vect cells. We also tested whether E1A could directly enhance paclitaxel-induced killing in nude mice, by using a nonviral, surface-protected cationic liposome to deliver E1A gene via the mouse tail vein. We compared the therapeutic effects of E1A gene therapy with or without Taxol chemotherapy in the established orthotopic tumor model of animals inoculated with MDA-MB-231 cells, and found that a combination of systemic E1A gene therapy and paclitaxel chemotherapy significantly enhanced the therapeutic efficacy and dramatically repressed tumor growth (P < .01). In addition, survival rates were significantly higher in animals treated with combination therapy than in the therapeutic control groups (both P < .0001). Thus, the E1A gene therapy indeed enhances the sensitivity of tumor cells to chemotherapy in a gene therapy setting and, the current study provides preclinical data to support combination therapy between E1A gene and chemotherapy for future clinical trials. PMID- 15272311 TI - Involvement of BNIP1 in apoptosis and endoplasmic reticulum membrane fusion. AB - BNIP1, a member of the BH3-only protein family, was first discovered as one of the proteins that are capable of interacting with the antiapoptotic adenovirus E1B 19-kDa protein. Here we disclose a totally unexpected finding that BNIP1 is a component of the complex comprising syntaxin 18, an endoplasmic reticulum (ER) located soluble N-ethylmaleimide-sensitive factor (NSF) attachment protein (SNAP) receptor (SNARE). Functional analysis revealed that BNIP1 participates in the formation of the ER network structure, but not in membrane trafficking between the ER and Golgi. Notably, a highly conserved leucine residue in the BH3 domain of BNIP1 plays an important role not only in the induction of apoptosis but also in the binding of alpha-SNAP, an adaptor that serves as a link between the chaperone ATPase NSF and SNAREs. This predicts that alpha-SNAP may suppress apoptosis by competing with antiapoptotic proteins for the BH3 domain of BNIP1. Indeed, overexpression of alpha-SNAP markedly delayed staurosporine-induced apoptosis. Our results shed light on possible crosstalk between apparently independent cellular events, apoptosis and ER membrane fusion. PMID- 15272314 TI - Therapy of human pancreatic carcinoma based on suppression of HMGA1 protein synthesis in preclinical models. AB - Pancreatic carcinoma is one of the most aggressive tumors, and, being refractory to conventional therapies, is an excellent target for new therapeutic approaches. Based on our previous finding of high HMGA1 expression in pancreatic cancer cells compared to normal pancreatic tissue, we evaluated whether suppression of HMGA1 protein expression could be a treatment option for patients affected by pancreatic cancer. Here we report that HMGA1 proteins are overexpressed in pancreatic carcinoma cell lines, and their downregulation through an adenovirus carrying the HMGA1 gene in an antisense orientation (Ad Yas-GFP) results in the death of three human pancreatic carcinoma cell lines (PANC1, Hs766T and PSN1). Pretreatment of PANC1 and PSN1 cells with Ad Yas-GFP suppressed and reduced, respectively, their ability to form xenograft tumors in nude mice. To further verify the role of HMGA1 in pancreatic tumorigenesis, we used a HMGA1 antisense phosphorothioate oligodeoxynucleotide (ODN); its addition induced a decrease in HMGA1 protein levels and a significant reduction of the proliferation rate of PANC1-, Hs766T- and PSN1-treated cells. Therefore, suppression of HMGA1 protein synthesis by an HMGA1 antisense approach seems to be a feasible treatment strategy in pancreatic carcinomas. PMID- 15272315 TI - Regulation of Hsp27 expression and cell survival by the POU transcription factor Brn3a. PMID- 15272316 TI - A novel method for the combined flow cytometric analysis of cell cycle and cytochrome c release. PMID- 15272317 TI - Deficiency of Bax and Bak protects photoreceptors from light damage in vivo. AB - Photoreceptors of bax(-/-)bak(-/-) but neither bax(-/-) mice nor bak(-/-) mice are protected from developmental apoptosis, suggesting that bax(-/-)bak(-/-) photoreceptors may also be protected from pathologic apoptosis. To test this possibility, we exposed bax(-/-)bak(-/-) and bax(-/-) mice to bright light, which normally induces photoreceptor death. Photoreceptors in bax(-/-)bak(-/-) mice were protected from death compared to bax(-/-) mice as indicated by a reduction in the number of TUNEL-positive photoreceptor nuclei 24 h following light damage and almost complete preservation of photoreceptors 7 days following light damage. These results provide the first in vivo evidence that combined deficiency of Bax and Bak can rescue cells from a pathologic stimulus more effectively than Bax deficiency and suggest that combined deficiency of Bax and Bak may also protect cells from other insults. PMID- 15272318 TI - Genome-wide RNAi identifies p53-dependent and -independent regulators of germ cell apoptosis in C. elegans. AB - We used genome-wide RNA interference (RNAi) to identify genes that affect apoptosis in the C. elegans germ line. RNAi-mediated knockdown of 21 genes caused a moderate to strong increase in germ cell death. Genetic epistasis studies with these RNAi candidates showed that a large subset (16/21) requires p53 to activate germ cell apoptosis. Apoptosis following knockdown of the genes in the p53 dependent class also depended on a functional DNA damage response pathway, suggesting that these genes might function in DNA repair or to maintain genome integrity. As apoptotic pathways are conserved, orthologues of the worm germline apoptosis genes presented here could be involved in the maintenance of genomic stability, p53 activation, and fertility in mammals. PMID- 15272319 TI - Two's company, three's a crowd. PMID- 15272320 TI - Learning outcomes. PMID- 15272321 TI - The best position. PMID- 15272322 TI - Unusual conditions. PMID- 15272323 TI - A clear message? PMID- 15272324 TI - Drugs for vegetarians. PMID- 15272325 TI - No fly zone. PMID- 15272326 TI - Incidental finding. PMID- 15272327 TI - Hemorrhagic tendencies. PMID- 15272328 TI - Clinical techniques. PMID- 15272329 TI - Non-payment. PMID- 15272330 TI - Editorial policy. PMID- 15272337 TI - Research in primary dental care Part 6: Data analysis. AB - The analysis of data from a research project seeks to answer the research question which the investigators set at the outset of the study. This article provides information on data analysis for both quantitative and qualitative data, always referring the reader back to their initial research question. Advice is also given on software which can be used for the analysis of data. A guide is provided to the choice of appropriate statistics for studies involving quantitative data. PMID- 15272338 TI - Presentation of a case of variant CJD in general dental practice. AB - This case report describes the initial presentation of variant CJD to a general dental practitioner. The case highlights the importance of prompt referral of patients presenting with a history of atypical facial symptoms. PMID- 15272339 TI - Severe Acute Respiratory Syndrome (SARS) and the GDP. Part I : Epidemiology, virology, pathology and general health issues. AB - The health profession faces a new challenge with the emergence of a novel viral disease Severe Acute Respiratory Syndrome (SARS), a form of atypical pneumonia caused by a coronavirus termed SARS-CoV. This highly infectious disease has spread through 32 countries, infecting more than 8,400 patients with over 790 deaths in just over 6 months. Over one quarter of those infected were unsuspecting healthcare workers. The major transmission mode of SARS-coronavirus appears to be through droplet spread with other minor subsidiary modes of transmission such as close contact and fomites although air borne transmission has not been ruled out. There is as yet no definitive treatment protocol. Although the peak period of the outbreak is likely to have passed and the risk of SARS in the UK is therefore assessed to be low, the World Health Organisation has asked all countries to remain vigilant lest SARS re-emerges. Recent laboratory acquired cases of SARS reported from Taiwan and Beijing, China are a testimony to this risk. Until reliable diagnostic tests, vaccine and medications are available, control of SARS outbreaks depends on close surveillance, early identification of index cases, quick isolation of carriers and effective infection control and public health measures. PMID- 15272346 TI - Quality improvement of referrals to a department of restorative dentistry following the use of a referral proforma by referring dental practitioners. AB - AIM: To assess the quality improvement of new patient referrals to a restorative department comparing a standard referral proforma and a normal referral letter. DESIGN: A prospective analysis of a consecutive sample of all referral letters and replied proforma until a total of 100 had been achieved. METHOD: The study covered the period from November 2000 to June 2001. Once the letters and corresponding proforma were matched, they were compared for data capture and hence quality. RESULTS: There was an increase in 29.3% of information provided. Specific categories of data showed high increases such as patient's telephone number, relevant medical history, treatment already given, recorded signs and symptoms, urgency of the referral and whether treatment or advice was requested. CONCLUSIONS: In this study, the quality of restorative referral increased with the use of a referral proforma. PMID- 15272348 TI - Comparing lecture and e-learning as pedagogies for new and experienced professionals in dentistry. AB - OBJECTIVE: To evaluate the relative effectiveness of e-learning versus lecture learning in VDPs and trainers. DESIGN: Experimental comparison of two groups' learning retention. SETTING: VDPs and trainers from two regions were assessed by independent researchers. METHOD: One region's VDPs and trainers received e learning; another's received a traditional one hour lecture. Retention and understanding were tested and compared. Personal preference was assessed in group interviews. RESULTS: Significantly greater retention for the trainees occurred from lecturing rather than e-learning, and for the trainers e-learning was significantly more successful than lecturing. CONCLUSIONS: Small numbers in this study preclude wide generalisation. However, the results point to the benefits of face-to-face interaction for inexperienced staff, and the benefits of the speed and manageability of e-learning for busy, more experienced staff. The need for a discussion facility to be incorporated into ICT innovations to CPD (via, for example, online 'chatrooms') is also highlighted, with the potential of greatly enhancing e-learning efficacy. PMID- 15272347 TI - 'It's difficult being a dentist': stress and health in the general dental practitioner. AB - OBJECTIVE: The aim of the study was to investigate overall stress, work-stress and health in general dental practitioners (GDPs). DESIGN, SETTING AND SUBJECTS: A nationwide anonymous cross-sectional survey was undertaken using stratified random sampling of 2,441 GDPs in the UK.Main outcome measures Measures included perceived stress, Work Stress Inventory for Dentists, job dissatisfaction, measures of health symptoms and health behaviour, dental and demographic information. RESULTS: The main findings were that perceived stress was significantly correlated with measures of dental stress. Work-related factors: fragility of dentist-patient relationship, time and scheduling pressures, staff and technical problems, job dissatisfaction, percentage NHS, and number of hours worked per week together explained nearly a half of GDPs overall stress in their life (linear multiple regression, adjusted r(2) = 0.48, F (2, 2404) = 509.68, P < 0.0001). Health behaviours such as alcohol use was associated with work stress (r = 0.18, P < 0.001) and over a third of GDPs were overweight or obese. Sixty per cent of GDPs reported being nervy, tense or depressed, 58.3% reported headache, 60% reported difficulty in sleeping and 48.2% reported feeling tired for no apparent reason. These were all related to work stress (one way analysis of variance, F (1,2211) = 241.53 P < 0.0001, F (1,2214) = 86.17 P < 0.0001; F (1, 2215) = 125.55 P < 0.0001; F (1,2211) = 209.67 P < 0.0001 respectively). Levels of minor psychiatric symptoms were high, with 32.0% of cases identified. The amount of backache was also high (reported by 68.3% of GDPs). CONCLUSION: A high percentage of NHS dentistry was associated with high levels of overall stress in GDPs' lives, indicating that the nature of NHS dentistry should be carefully investigated to try to improve GDPs working conditions. A comparatively large number of dentists reported high levels of psychological stress symptoms, such as being nervy, tense and depressed, showing minor psychiatric symptoms, with alcohol use being related to stress. Other factors reported which were not related to stress but may be related to the actual practice of dentistry were that a third of dentists were overweight or obese and over 60% reported backache. Overall, these findings indicate the stressful nature of dentistry and difficulties in working conditions. The next step should be to develop interventions to help dentists to reduce stress in the dental surgery. PMID- 15272364 TI - Dentist on TV. AB - Why is it that dentistry just doesn't seem to cut the mustard when the programme planners are looking to fill next year's schedules? PMID- 15272365 TI - Visualizing the Needle in the Haystack: In Situ Hybridization With Fluorescent Dendrimers. AB - In situ hybridization with 3DNA trade mark dendrimers is a novel tool for detecting low levels of mRNA in tissue sections and whole embryos. Fluorescently labeled dendrimers were used to identify cells that express mRNA for the skeletal muscle transcription factor MyoD in the early chick embryo. A small population of MyoD mRNA positive cells was found in the epiblast prior to the initiation of gastrulation, two days earlier than previously detected using enzymatic or radiolabeled probes for mRNA. When isolated from the epiblast and placed in culture, the MyoD mRNA positive cells were able to differentiate into skeletal muscle cells. These results demonstrate that DNA dendrimers are sensitive and precise tools for identifying low levels of mRNA in single cells and tissues. PMID- 15272366 TI - Tin-free radical chemistry using the persistent radical effect: alkoxyamine isomerization, addition reactions and polymerizations. AB - In this tutorial review applications of alkoxyamines as C-radical precursors for the conduction of tin-free radical reactions are presented. These processes are controlled by the Persistent Radical Effect. A brief introduction on the Persistent Radical Effect is provided. In addition, the use of microwave irradiation to conduct thermal radical reactions is discussed. Finally, the use of alkoxyamines as initiators/mediators for the controlled/living radical polymerization is highlighted. PMID- 15272367 TI - Selected strategies for the synthesis of triphenylenes. AB - This tutorial review surveys the most useful strategies for the synthesis of triphenylenes. It is aimed at organic chemists in general and, in particular, synthetic chemists interested in the development of advanced materials for organic electronic devices. The synthetic strategies considered are classified according to the structures of their key intermediates. Selected examples illustrate the variety of target structures and the method(s) of choice for their synthesis. PMID- 15272368 TI - The NiCl2-Li-arene(cat.) combination: a versatile reducing mixture. AB - The NiCl2.2H2O-Li-arene(cat.) combination described in this tutorial review has shown to be a useful and versatile mixture able to reduce a broad range of functionalities bearing carbon-carbon multiple bonds, as well as carbon heteroatom and heteroatom-heteroatom single and multiple bonds. The analogous deuterated combination, NiCl2.2D2O-Li-arene(cat.), allows the easy incorporation of deuterium in the reaction products. Alternatively, the anhydrous NiCl2-Li arene (or polymer-supported arene)(cat.) system generates a highly reactive metallic nickel, which in the presence of molecular hydrogen at atmospheric pressure is able to catalyze the hydrogenation of almost the same type of functionalities mentioned above. PMID- 15272369 TI - Highly ordered structures of peptides by using molecular scaffolds. AB - Protein secondary structures such as alpha-helices, beta-sheets, and beta-turns are important in inducing the three-dimensional structure and biological activity of proteins. Designing secondary structure mimics composed of short peptides has attracted much attention not only to gain fundamental insight into the factors affecting protein folding but also to develop pharmacologically useful compounds, artificial receptors, asymmetric catalysts, and new materials. In this tutorial review, we focus on molecular scaffolds employed to induce beta-sheet-like structure in attached peptide chains, thereby creating highly ordered molecular structures, and discuss the versatility of these molecular scaffolds to regulate the attached peptide strands in the appropriate dimensions. PMID- 15272370 TI - Cooperative multi-catalyst systems for one-pot organic transformations. AB - One-pot co-catalyst systems are covered in this tutorial review. It is divided into three sections according to the reaction types: i) one catalyst performs a desired reaction as the second catalyst restores the first catalytic species back into its original state for the next catalytic cycles, ii) two catalysts carry out sequential organic transformations, in which the first step is carried out by one catalyst to afford certain intermediates being to be subjected to the second catalyst for the next step, and iii) cooperative catalytic actions on both substrates by suitable catalysts proceed in a substrate-selective manner followed by the subsequent coupling of the two activated adducts providing the desired products. PMID- 15272371 TI - The syntheses and catalytic applications of unsymmetrical ferrocene ligands. AB - Interest in the chemistry of ferrocene remains intense, largely due to applications within catalysis. New synthetic routes to unsymmetrical ferrocene ligands have provided another facet to this area, as substituents can be designed to be electronically- and/or sterically-distinct in order to affect the environment around the catalytically-active metal centre. This critical review provides a concise summary of the synthetic routes that have been applied to the synthesis of unsymmetrical ferrocene ligands, along with a systematic survey of the applications of these ligands in homogeneous catalysis. The aim is to help the reader select a suitable ferrocenediyl ligand for a particular synthetic application, and in the synthesis of ligands that require particular structural and/or electronic features. PMID- 15272372 TI - The choice between epidermal and neural fate: a matter of calcium. AB - The development of the vertebrate embryo, which includes the induction and the patterning of the three germ layers, requires signaling among cells. In vertebrates, cells of the embryonic ectoderm have a choice during gastrulation between to fates; they give rise to epidermal progenitors on the ventral side and neural progenitors on the dorsal side. It was first shown by Spemann and Mangold in 1924 that the dorsal mesoderm (also called the Spemann-Mangold Organizer) has a potent biological capacity to induce nervous system from the adjacent ectoderm. A similar observation was reported in amniotes, indicating that neural induction by the Organizer is a conserved process controlling the initial steps in vertebrates neurogenesis. Molecular studies of the last decade have led to the identification of signaling molecules which participate in these embryonic decisions. Epidermis induction occurs through a signaling cascade involving Bone Morphogenetic Proteins (BMP 2, 4, 7) and receptor-regulated Smad proteins which translocate into the nucleus to form active transcriptional complexes. These molecules mediate BMPs effect to activate epidermal-specific gene expression and to repress neural-specific gene transcription. Neural fate is revealed by factors secreted by the dorsal mesoderm (Noggin, Chordin, Follistatin, ...) which act by blocking BMP signaling. Consequently, epidermal fate is an induced fate while neural fate is interpreted as a default state of the ectoderm. This review describes the signaling pathways which act to determine the fate of the ectoderm and discuss an alternative model in which neural fate is not a default state. This new model integrates the activation of a calcium-dependent signaling pathway due to an influx of calcium through L-type calcium channels. In this model, calcium plays a central regulatory role. While calcium is required for neural determination, epidermal determination occurs when calcium-dependent signaling pathways are inactive. PMID- 15272373 TI - Skin field formation: morphogenetic events. AB - This chapter is mostly a review of the pioneering work of the Philippe Sengel school in Grenoble carried out in the late sixties and the seventies. The questions raised concerning the morphogenesis of feather tracts were approached by means of microsurgery on chick embryos. P. Sengel and his wife M. Kieny had the feeling that proteins synthesized by the neural tube were required for the formation of feather fields. It was my pleasure to carry on the story from the beginning. Although some clarifications concerning this morphogenesis have been contributed by my group and by a few other laboratories interested in this subject, the most important contributions to recent research have been the elucidation of the nature of the required messages, which will be explored further in other papers in this Issue. PMID- 15272374 TI - Molecular mechanisms controlling dorsal dermis generation from the somitic dermomyotome. AB - The initiation of the development of skin appendages (hair/feathers/scales) requires a signal from the competent dense dermis to the epidermis (Dhouailly, 1977). It is therefore essential to understand how to make a competent dermis. In recent years, a few studies have focused on the development of the dorsal dermis from the somitic dermomyotome. Our first aim in this review is to attempt to reconcile the available data on the origin of the dorsal dermis and summarize the present knowledge on the molecular mechanisms implicated in dermal lineage induction. Secondly, we open the discussion on the formation of a loose pre dermal mesenchyme and more importantly of a dense dermis capable of participating in appendage development. To go further we draw a comparison between the chick and mouse systems to gain a new insight into how to initiate appendage morphogenesis and regulate the extent of hair/feather fields. PMID- 15272375 TI - Ventral vs. dorsal chick dermal progenitor specification. AB - The dorsal and the ventral trunk integuments of the chick differ in their dermal cell lineage (originating from the somatic and somatopleural mesoderm respectively) and in the distribution of their feather fields. The dorsal macropattern has a large spinal pteryla surrounded by semi-apteria, whereas the ventral skin has a true medial apterium surrounded by the ventral pterylae. Comparison of the results of heterotopic transplantations of distal somatopleure in place of somatic mesoderm (Mauger 1972) or in place of proximal somatopleure (our data), leads to two conclusions. These are that the fate of the midventral apterium is not committed at day 2 of incubation and that the signals from the environment which specify the ventral and dorsal featherforming dermal progenitors are different. Effectively, Shh, but not Wnt -1 signalling can induce the formation of feather forming dermis from the embryonic somatopleure. Shh is not able, however, to trigger the formation of a feather forming dermis from the extra embryonic somatopleure. This brief report constitutes the first attempt, by comparing old and new preliminary results, to understand whether dermal progenitors at different sites are specified by different signalling pathways. PMID- 15272376 TI - The different steps of skin formation in vertebrates. AB - Skin morphogenesis occurs following a continuous series of cell-cell interactions which can be subdivided into three main stages: 1- the formation of a dense dermis and its overlying epidermis in the future appendage fields (macropattern); 2- the organization of these primary homogeneous fields into heterogeneous ones by the appearance of cutaneous appendage primordia (micropattern) and 3- cutaneous appendage organogenesis itself. In this review, we will first show, by synthesizing novel and previously published data from our laboratory, how heterogenetic and heterospecific dermal/epidermal recombinations have allowed us to distinguish between the respective roles of the dermis and the epidermis. We will then summarize what is known from the work of many different research groups about the molecular signaling which mediates these interactions in order to introduce the following articles of this Special Issue and to highlight what remains to done. PMID- 15272378 TI - How and when the regional competence of chick epidermis is established: feathers vs. scutate and reticulate scales, a problem en route to a solution. AB - Most of the chick body is covered with feathers, while the tarsometatarsus and the dorsal face of the digits form oblong overlapping scales (scuta) and the plantar face rounded nonoverlapping scales (reticula). Feathers and scuta are made of beta-keratins, while the epidermis of reticula and inter-appendage or apteria (nude regions) express a-keratins. These regional characteristics are determined in skin precursors and require an epidermal FGF-like signal to be expressed. Both the initiation of appendages, their outline and pattern depend on signals from the dermis, while their asymmetry and outgrowth depend on epidermal competence. For example, the plantar dermis of the central foot pad induces reticula in a plantar or feathers in an apteric epidermis, in a hexagonal pattern starting from the medial point. By manipulating Shh levels in the epidermis, the regional appendage type can be changed from scuta or reticula to feather, whereas the inhibition of Wnt7a, together with a downregulation of Shh gives rise to reticula and in extreme cases, apteria. During morphogenesis of plantar skin, the epidermal expression of En-1, acting as a repressor both of Wnt7a and Shh, is linked to the formation of reticula. Finally, in birds, the complex formation of feathers, which can be easily triggered, even in the extra-embryonic somatopleure, may result from a basic genetic program, whereas the simple formation of scales appears secondarily derived, as requiring a partial (scuta) or total (reticula) inhibition of epidermal outgrowth and beta-keratin gene expression, an inhibition lost for the scuta in the case of feathered feet breeds. PMID- 15272377 TI - Integument pattern formation involves genetic and epigenetic controls: feather arrays simulated by digital hormone models. AB - Pattern formation is a fundamental morphogenetic process. Models based on genetic and epigenetic control have been proposed but remain controversial. Here we use feather morphogenesis for further evaluation. Adhesion molecules and/or signaling molecules were first expressed homogenously in feather tracts (restrictive mode, appear earlier) or directly in bud or inter-bud regions ( de novo mode, appear later). They either activate or inhibit bud formation, but paradoxically colocalize in the bud. Using feather bud reconstitution, we showed that completely dissociated cells can reform periodic patterns without reference to previous positional codes. The patterning process has the characteristics of being self-organizing, dynamic and plastic. The final pattern is an equilibrium state reached by competition, and the number and size of buds can be altered based on cell number and activator/inhibitor ratio, respectively. We developed a Digital Hormone Model which consists of (1) competent cells without identity that move randomly in a space, (2) extracellular signaling hormones which diffuse by a reaction-diffusion mechanism and activate or inhibit cell adhesion, and (3) cells which respond with topological stochastic actions manifested as changes in cell adhesion. Based on probability, the results are cell clusters arranged in dots or stripes. Thus genetic control provides combinational molecular information which defines the properties of the cells but not the final pattern. Epigenetic control governs interactions among cells and their environment based on physical-chemical rules (such as those described in the Digital Hormone Model). Complex integument patterning is the sum of these two components of control and that is why integument patterns are usually similar but non-identical. These principles may be shared by other pattern formation processes such as barb ridge formation, fingerprints, pigmentation patterning, etc. The Digital Hormone Model can also be applied to swarming robot navigation, reaching intelligent automata and representing a self-re-configurable type of control rather than a follow-the instruction type of control. PMID- 15272379 TI - Drm/Gremlin, a BMP antagonist, defines the interbud region during feather development. AB - The pattern of feather buds in a tract is thought to result from the relative ratios between activator and inhibitor signals through a lateral inhibition process. We analyse the role of Drm/Gremlin, a BMPs antagonist expressed during feather pattern formation, in the dermal precursor, the dense dermis, the interbud dermis and in the posterior dermal condensation. We have altered the activity of Drm in embryonic chick skin using retroviral vectors expressing drm/ gremlin and bmps. We show that expression of endogenous drm is under the control of a feedback loop induced by the BMP pathway, and that overexpression of drm results in fusion between adjacent feather buds. We propose that endogenous BMP proteins induce drm expression in the interbud dermis. In turn, the Drm/Gremlin protein limits the inhibitory effect of BMPs, allowing the adjacent row of feathers to form. Thus, the balance between BMPs and its antagonist Drm would regulate the size and spacing of the buds. PMID- 15272380 TI - Spacing patterns on tongue surface-gustatory papilla. AB - The dorsal surface of the mammalian tongue is covered with four kinds of papillae, fungiform, circumvallate, foliate and filiform papillae. With the exception of the filiform papillae, these types of papillae contain taste buds and are known as the gustatory papillae. The gustatory papillae are distributed over the tongue surface in a distinct spatial pattern. The circumvallate and foliate papillae are positioned in the central and lateral regions respectively and the fungiform papillae are distributed on the anterior part of the tongue in a stereotyped array. The patterned distribution and developmental processes of the fungiform papillae indicate some similarity between the fungiform papillae and the other epithelial appendages, including the teeth, feathers and hair. This is because 1) prior to the morphological changes, the signaling molecules are expressed in the fungiform papillae forming area with a stereotyped pattern; 2) the morphogenesis of the fungiform papillae showed specific structures in early development, such as epithelial thickening and mesenchymal condensation and 3) the fungiform papillae develop through reciprocal interactions between the epithelium and mesenchymal tissue. These results led us to examine whether or not the early organogenesis of the fungiform papillae is a good model system for understanding both the spacing pattern and the epithelial-mesenchymal interaction during embryogenesis. PMID- 15272381 TI - Hair follicle differentiation and regulation. AB - Ten years ago, Hardy (1992) wrote a timely review on the major features of hair follicle development and hair growth which she referred to as a secret life. Many of these secrets are now being revealed. The information discussed in this brief review comprises the structure of the hair and hair follicle, the continuing characterisation of the genes for keratin and keratin associated proteins, the determination of the location of their expression in the different cell layers of the hair follicle, molecular signals which control keratin gene expression and post-translational events in the terminal stages of hair formation. PMID- 15272382 TI - Inherited disorders of the skin in human and mouse: from development to differentiation. AB - The last ten years has revealed some of the key players in the development and differentiation of the hair follicle and the epidermis in general. In this review, we discuss how our current understanding of these processes has been made possible by the elucidation of the molecular basis of human inherited diseases and mouse mutants which display defects in the hair and epidermis. For examples, the study of ectodermal dysplasias and the basal cell carcinoma predisposition disease Gorlin syndrome have allowed the determination of signalling hierarchies critical in the formation of the hair follicle. Epidermolytic diseases and hyperkeratoses have focussed attention on the importance of the programs of keratin expression, while ichthyoses provide insight in the final stage of epidermal development, cornification. Finally, the increasing range of diseases and mouse models exhibiting alopecias are revealing the critical pathways in control of the hair follicle cycle. PMID- 15272384 TI - Sharpening the focus; the business of epithelial cell biology. An interview with Chris Potten by David Pearton. AB - Dr Chris Potten is a singularly influential figure in the field of epithelial biology. His contributions have been seminal and include the introduction of the epidermal proliferative unit and of the concept of epidermal stem cells. With around 400 scientific papers and reviews to his credit as well as two books, he has certainly made his mark. His contributions have been recognised by the award of the Curie medal and recently the Weiss medal for radiation biology. Dr. Potten graciously agreed to be interviewed for this Special Issue of The International Journal of Developmental Biology. This interview was conducted via e-mail during June -August 2003. PMID- 15272383 TI - The biology of feather follicles. AB - The feather is a complex epidermal organ with hierarchical branches and represents a multi-layered topological transformation of keratinocyte sheets. Feathers are made in feather follicles. The basics of feather morphogenesis were previously described (Lucas and Stettenheim, 1972). Here we review new molecular and cellular data. After feather buds form (Jiang et al., this issue), they invaginate into the dermis to form feather follicles. Above the dermal papilla is the proliferating epidermal collar. Distal to it is the ramogenic zone where the epidermal cylinder starts to differentiate into barb ridges or rachidial ridge. These neoptile feathers tend to be downy and radially symmetrical. They are replaced by teleoptile feathers which tend to be bilateral symmetrical and more diverse in shapes. We have recently developed a "transgenic feather" protocol that allows molecular analyses: BMPs enhance the size of the rachis, Noggin increases branching, while anti- SHH causes webbed branches. Different feather types formed during evolution (Wu et al., this issue). Pigment patterns along the body axis or intra-feather add more colorful distinctions. These patterns help facilitate the analysis of melanocyte behavior. Feather follicles have to be connected with muscles and nerve fibers, so they can be integrated into the physiology of the whole organism. Feathers, similarly to hairs, have the extraordinary ability to go through molting cycles and regenerate. Some work has been done and feather follicles might serve as a model for stem cell research. Feather phenotypes can be modulated by sex hormones and can help elucidate mechanisms of sex hormone-dependent growth control. Thus, the developmental biology of feather follicles provides a multi-dimension research paradigm that links molecular activities and cellular behaviors to functional morphology at the organismal level. PMID- 15272385 TI - Transdifferentiation of corneal epithelium: evidence for a linkage between the segregation of epidermal stem cells and the induction of hair follicles during embryogenesis. AB - Corneal epithelium transdifferentiation into a hair-bearing epidermis provides a particularly useful system for studying the possibility that transient amplifying (TA) cells are able to activate different genetic programs in response to a change in their fibroblast environment, as well as to follow the different steps of rebuilding an epidermis from induced stem cells. Corneal stem and TA cells are found in different locations - stem cells at the periphery, in the limbus, and TA cells more central. Moreover, the TA cells already express the differentiating corneal-type keratin pair K3/K12, whereas the limbal keratinocytes express the basal keratin pair K5/K14. In contrast, suprabasal epidermal keratinocytes express keratin pair K1-2/K10, and basal keratinocytes the keratin pair K5/K14. The results of tissue recombination experiments show that adult central corneal cells are able to respond to specific information originating from embryonic dermis. First, the cells located at the base of the corneal epithelium show a decrease in expression of K12 keratin, followed by an increase in K5 expression; they then proliferate and form hair follicles. The first K10 expressing cells appear at the junction of the new hair follicles and the covering corneal epithelium. Their expansion finally gives rise to epidermal strata, which displace the corneal suprabasal keratinocytes. Corneal TA cells can thus be reprogrammed to form epidermal cells, first by reverting to a basal epithelial type, then to hair pegs and probably concomitantly to hair stem cells. This confirms the role of the hair as the main reservoir of epidermal stem cells and raises the question of the nature of the dermal messages which are both involved in hair induction and stem cell specification. PMID- 15272386 TI - Derivation of keratinocyte progenitor cells and skin formation from embryonic stem cells. AB - Despite numerous elegant transgenic mice experiments, the absence of an appropriate in vitro model system has hampered the study of the early events responsible for epidermal and dermal commitments. Embryonic stem (ES) cells are derived from the pluripotent cells of the early mouse embryo. They can be expanded infinitely in vitro while maintaining their potential to spontaneously differentiate into any cell type of the three germ layers, including epidermal cells. We recently reported that ES cells have the potential to recapitulate the reciprocal instructive ectodermal-mesodermal commitments, which are characteristic of embryonic skin formation. Derivation of epidermal cells from murine ES cells has been successfully established by exposing the cells to precisely controlled instructive influences normally found in the body, including extracellular matrix and the morphogen BMP-4. These differentiated ES cells are able to form, in culture, a multilayered epidermis coupled with an underlying dermal compartment similar to native skin. This bioengineered skin provides a powerful tool for studying the molecular mechanisms controlling skin development and epidermal stem cell properties. PMID- 15272387 TI - Genetic control of epidermis differentiation in Drosophila. AB - In arthropods, the animal body is isolated from the external environment by a protective exoskeleton called the cuticle. The cuticle of young larvae has certainly been the most scrutinized structure in Drosophila and genetic studies of the pattern of cuticular extensions has provided the main source of our comprehension of the control of embryonic development. However, the complex structure of the cuticle remains poorly understood and analysis of the underlying epidermis has started only recently. Here I review different aspects of epidermis differentiation with the aim of presenting an integrated view of the organisation of the Drosophila integument. Although profound differences in epidermis organisation are observed across species, accumulated results suggest that epidermis formation and differentiation might share an unsuspected number of homologies between Drosophila and vertebrates. PMID- 15272388 TI - Skin development in bony fish with particular emphasis on collagen deposition in the dermis of the zebrafish (Danio rerio). AB - The first part of this article is a review of the current status of knowledge of the fish skin, with particular attention to its development. In the second part we present original results obtained in zebrafish (Danio rerio), with particular emphasis on the deposition and organisation of the dermal collagenous stroma. Using a series of zebrafish specimens aged between 15 hours postfertilization (hpf) and 4.5 years old, we have combined Transmission Electron Microscopy (TEM) observations and in situ hybridisation using type I collagen a2 chain (Col1a2) probe. Collagen fibrils, with a diameter of 22 nm, appear first in an acellular subepidermal space at 24 hpf, are first all oriented in the same direction, and form the primary dermal stroma. Subsequently, three events occur. (1) From 5-7 days pf (dpf) onwards the collagen fibrils self-organise into several lamellae arranged in a plywood-like structure, starting in the upper layers and progressing throughout the entire thickness of the dermis. (2) At 20-26 dpf, fibroblasts of unknown origin progressively invade the acellular collagenous stroma, some of them accumulating below the epidermis. (3) Concomitant with the invasion of fibroblasts, the collagen fibrils increase progressively in diameter to reach 160 nm towards the end of the fish life. In situ hybridisation experiments reveal that, between 24 and 48 hpf, the collagen matrix is produced by the epidermis only. From 72 hpf to 20-26 dpf, both the basal epidermal cells and the dermal cells bordering the deep region of the dermis are involved in the production of collagen. When the fibroblasts invade the plywood-like structure, the epidermal cells progressively cease to synthesise collagen, which from this point is produced only by the fibroblasts. This suggests that the fibroblasts secrete a still unidentified signalling molecule that downregulates collagen production by the epidermis. PMID- 15272389 TI - Scale development in fish: a review, with description of sonic hedgehog (shh) expression in the zebrafish (Danio rerio). AB - In the first part of this paper we review current knowledge regarding fish scales, focusing on elasmoid scales, the only type found in two model species, the zebrafish and the medaka. After reviewing the structure of scales and their evolutionary origin, we describe the formation of the squamation pattern. The regularity of this process suggests a pre-patterning of the skin before scale initiation. We then summarise the dynamics of scale development on the basis of morphological observations. In the absence of molecular data, these observations support the existence of genetic cascades involved in the control of scale development. In the second part of this paper, we illustrate the potential that scale development offers as a model to study organogenesis mediated by epithelial mesenchymal interactions. Using the zebrafish (Danio rerio), we have combined alizarin red staining, light and transmission electron microscopy and in situ hybridisation using an anti-sense RNA probe for the sonic hedgehog (shh) gene. Scales develop late in ontogeny (30 days post-fertilisation) and close to the epidermal cover. Only cells of the basal epidermal layer express shh. Transcripts are first detected after the scale papillae have formed. Thus, shh is not involved in the mechanisms controlling squamation patterning and scale initiation. As the scales enlarge, shh expression is progressively restricted to a subset of basal epidermal cells located in the region that overlies their posterior field. This pattern of expression suggests that shh may be involved in the control of scale morphogenesis and differentiation in relationship with the formation of the epidermal fold in the posterior region. PMID- 15272391 TI - Evidence-based prevention: increasing the efficiency of HIV intervention trials. PMID- 15272390 TI - Evo-Devo of amniote integuments and appendages. AB - Integuments form the boundary between an organism and the environment. The evolution of novel developmental mechanisms in integuments and appendages allows animals to live in diverse ecological environments. Here we focus on amniotes. The major achievement for reptile skin is an adaptation to the land with the formation of a successful barrier. The stratum corneum enables this barrier to prevent water loss from the skin and allowed amphibian / reptile ancestors to go onto the land. Overlapping scales and production of beta-keratins provide strong protection. Epidermal invagination led to the formation of avian feather and mammalian hair follicles in the dermis. Both adopted a proximal - distal growth mode which maintains endothermy. Feathers form hierarchical branches which produce the vane that makes flight possible. Recent discoveries of feathered dinosaurs in China inspire new thinking on the origin of feathers. In the laboratory, epithelial - mesenchymal recombinations and molecular mis-expressions were carried out to test the plasticity of epithelial organ formation. We review the work on the transformation of scales into feathers, conversion between barbs and rachis and the production of "chicken teeth". In mammals, tilting the balance of the BMP pathway in K14 noggin transgenic mice alters the number, size and phenotypes of different ectodermal organs, making investigators rethink the distinction between morpho-regulation and pathological changes. Models on the evolution of feathers and hairs from reptile integuments are discussed. A hypothetical Evo-Devo space where diverse integument appendages can be placed according to complex phenotypes and novel developmental mechanisms is presented. PMID- 15272392 TI - Some design issues in trials of microbicides for the prevention of HIV infection. AB - Trials for the prevention of human immunodeficiency virus (HIV) infection that evaluate microbicides provide significant design challenges. Three of these design issues deserve more careful consideration. The first issue relates to the benefits of using both blinded and unblinded control groups when the placebo regimen may not be inert and when the effectiveness of an intervention heavily depends on behavioral, as well as biological, factors. The second issue relates to the strength of evidence required for regulatory approval for the marketing of drugs and biologics when only a single pivotal phase 3 clinical trial has provided such evidence. The third issue relates to the appropriate next step after the completion of phase 1 trials, as well as the specific merits of conducting phase 2b screening trials that assess the effects on the same clinical efficacy end point that will be the primary end point in a phase 3 trial. The issues considered in microbicide trials for the prevention of HIV infection are also of importance in many other clinical scenarios. PMID- 15272393 TI - Nucleoside analogue use before and during highly active antiretroviral therapy and virus load rebound. AB - Patients who use nucleoside reverse-transcriptase inhibitors (NRTIs) before highly active antiretroviral therapy (HAART) have an increased rate of virus rebound. Study of rebound according to specific NRTIs used might inform which NRTIs retain activity once others have failed. We focused on 2280 patients who had received zidovudine and either didanosine or lamivudine before starting HAART, started HAART that included zidovudine with didanosine or lamivudine or stavudine with didanosine or lamivudine, and had virus loads <500 copies/mL within 24 weeks. In a Cox model, the relative hazard (RH) of virus rebound for having switched from zidovudine to stavudine (vs. retaining zidovudine) was 0.94 (95% confidence interval [CI], 0.77-1.15), which suggests little or no benefit in terms of reduced rebound rate. Having switched from didanosine to lamivudine, or vice versa, was associated with a reduced rebound rate (RH, 0.59 [95% CI, 0.48 0.73]), which suggests that these drugs retain appreciable activity after use of the other and of zidovudine. PMID- 15272394 TI - Changes in mitochondrial DNA in peripheral blood mononuclear cells from HIV infected patients with lipoatrophy randomized to receive abacavir. AB - It has been suggested that lipoatrophy associated with exposure to nucleoside analogues is caused by depletion of mitochondrial DNA (mtDNA). The aim of the present study was to determine whether switching treatment from a thymidine analogue to abacavir was associated with an increase in the mtDNA copy number in peripheral blood mononuclear cells (PBMCs). Of 111 patients with lipoatrophy who were randomized to have treatment switched to abacavir or to continue treatment with thymidine analogues, 94 patients had PBMCs obtained at baseline and at weeks 4, 12, and 24, for quantification of the mtDNA copy number. During the 24-week study, there was no significant change in mtDNA copy numbers in PBMCs in either treatment group, despite improvement in peripheral lipoatrophy among patients whose treatment was switched to abacavir. PMID- 15272395 TI - Frequent reactivation of herpes simplex virus among HIV-1-infected patients treated with highly active antiretroviral therapy. AB - The effect of highly active antiretroviral therapy (HAART) on control of herpes simplex virus (HSV) in human immunodeficiency virus (HIV) type 1-infected subjects is not known. Among 28 HAART-treated and 49 untreated subjects with HIV 1 and HSV-2 infections, mucosal HSV shedding (median, 18% and 29% of days positive for HSV DNA, respectively; P=.08) and HSV DNA level (median, 56,250 and 50,000 copies/mL, respectively; P=.20) were similar. Treated subjects reported significantly fewer days with HSV lesions, compared with untreated subjects (2.8% vs. 11.3% of days, respectively; P=.001). Thus, mucosal HSV shedding and HSV-2 reactivation were still frequent among treated subjects, even though HAART was associated with fewer days with HSV lesions. PMID- 15272396 TI - Long-term survivors in Nairobi: complete HIV-1 RNA sequences and immunogenetic associations. AB - To investigate African long-term survivors (LTSs) infected with non-subtype B human immunodeficiency virus type 1 (HIV-1), we obtained full-length HIV-1 RNA sequences and immunogenetic profiles from 6 untreated women enrolled in the Pumwani Sex Worker Cohort in Nairobi, Kenya. There were no discernible sequence changes likely to cause attenuation. CCR2-V64I, an immunogenetic polymorphism linked to LTSs, was detected in 4 women, all of whom carried the HLA B58 allele. Further investigation of 99 HIV-1-infected Nairobi women found an association between CCR2-V64I and HLA B58 (P=.0048). Studying the interaction among immunogenetics, immune responses, and viral sequences from all HIV-1 subtypes may increase our understanding of slow HIV-1 disease progression. PMID- 15272397 TI - Safety and immunogenicity of an HIV subtype B and E prime-boost vaccine combination in HIV-negative Thai adults. AB - ALVAC-HIV (vCP1521) and AIDSVAX B/E were evaluated in a phase 1/2 trial of human immunodeficiency virus (HIV)-negative Thai adults. Of 133 volunteers enrolled, 122 completed the trial. There were no serious vaccine-related adverse events, nor were there intercurrent HIV infections. Lymphoproliferative responses to glycoprotein 120 E were induced in 63% of the volunteers, and HIV-specific CD8 cytotoxic T lymphocyte responses were induced in 24%. Antibody responses increased in frequency and magnitude in association with the dose level of AIDSVAX B/E. Binding and neutralizing antibodies to the MN strain were induced in 100% and 98%, respectively, of the volunteers receiving 600 microg of AIDSVAX B/E, and such antibodies to E strains were induced in 96% and 71%, respectively, of these volunteers. This vaccine combination was well tolerated and was immunogenic, meeting milestones for advancement to phase 3 evaluation. PMID- 15272398 TI - A case-control study to investigate serological correlates of clinical failure of 23-valent pneumococcal polysaccharide vaccine in HIV-1-infected Ugandan adults. AB - We have investigated the association between the concentration of anti polysaccharide pneumococcal capsule-specific (anti-PS) immunoglobulin G and the killing activity, in serum, in invasive pneumococcal disease (IPD) events and response to 23-valent polysaccharide vaccine in human immunodeficiency virus (HIV)-infected Ugandans. Case patients with IPD had lower concentrations of anti PS IgG before and after vaccination and before the IPD event (P<.01 for 5 [i.e., 4, 9V, 14, 18C, and 19F] of 6 serotypes assessed). After vaccination, case patients were less likely than were control subjects to develop detectable serum killing activity against the 2 serotypes tested--for 19F, this activity was detected in 16% of case patients versus 37% of control subjects (P=.08); for 23F, it was detected in 11% of case patients versus 40% of control subjects (P=.02). Thus, absolute concentration of anti-PS IgG and an attenuated response to polysaccharide are associated with risk of IPD in HIV-infected adults. PMID- 15272399 TI - Loss of viral control in early HIV-1 infection is temporally associated with sequential escape from CD8+ T cell responses and decrease in HIV-1-specific CD4+ and CD8+ T cell frequencies. AB - The outcome of human immunodeficiency virus type 1 (HIV-1) infection is related to the set-point plasma virus load (pVL) that emerges after primary HIV-1 infection (PHI). This set-point pVL generally remains stable but eventually increases with progression to disease. However, the events leading to loss of viremic control are poorly understood. Here, we describe an individual who presented with symptomatic PHI and subsequently progressed rapidly, after an initial period of 1 year during which viral replication was well controlled. Escalation of viral replication in this atypical case was preceded by the emergence of escape variants in many epitopes targeted by dominant CD8+ T cell responses and a marked decrease in HIV-1-specific CD4+ and CD8+ T cell frequencies. There were no changes in viral tropism, replication kinetics, or neutralizing antibody titers. These findings demonstrate the temporal relationship between viral escape from CD8+ T cell activity, decrease in HIV-1 specific T cell frequencies, and loss of control of viral replication. PMID- 15272400 TI - Correlation between HIV-Specific CD8 cell production of interferon- gamma and plasma levels of HIV RNA in perinatally infected pediatric populations. AB - BACKGROUND: CD8 cell responses to human immunodeficiency virus (HIV) have been correlated with virus control in adults, and this study outcome has been controversial. Attempts to establish the same correlation in small numbers of children have also been made, with similar controversy resulting. METHODS: A total of 110 perinatally infected children were studied. Nine of the children (mean age, 1.9 years vs. 11.8 years for the remaining 101 children) received treatment with antiretrovirals within the first 3 months of life. CD4 cell and HIV RNA levels were measured. Production of interferon- gamma after exposure to recombinant vaccinia vectors was measured by enzyme-linked immunospot (ELISPOT) assay. RESULTS: Responses to Pol and Gag antigens exceeded those to Nef and Env antigens, with responses significantly approximated by a quadratic function for which peak responses occurred at plasma HIV RNA levels of 103-104 HIV RNA copies/mL. Children who are treated early in life with highly active antiretroviral therapy have fewer total responses of ELISPOT-forming cells to HIV antigens than do children who are treated later in life. PMID- 15272401 TI - Progress toward characterization of the group A Streptococcus metagenome: complete genome sequence of a macrolide-resistant serotype M6 strain. AB - We describe the genome sequence of a macrolide-resistant strain (MGAS10394) of serotype M6 group A Streptococcus (GAS). The genome is 1,900,156 bp in length, and 8 prophage-like elements or remnants compose 12.4% of the chromosome. A 8.3 kb prophage remnant encodes the SpeA4 variant of streptococcal pyrogenic exotoxin A. The genome of strain MGAS10394 contains a chimeric genetic element composed of prophage genes and a transposon encoding the mefA gene conferring macrolide resistance. This chimeric element also has a gene encoding a novel surface exposed protein (designated "R6 protein"), with an LPKTG cell-anchor motif located at the carboxyterminus. Surface expression of this protein was confirmed by flow cytometry. Humans with GAS pharyngitis caused by serotype M6 strains had antibody against the R6 protein present in convalescent, but not acute, serum samples. Our studies add to the theme that GAS prophage-encoded extracellular proteins contribute to host-pathogen interactions in a strain-specific fashion. PMID- 15272402 TI - Evolution of erythromycin-resistant Streptococcus pneumoniae from Asian countries that contains erm(B) and mef(A) genes. AB - To investigate the genetic characteristics of erythromycin-resistant Streptococcus pneumoniae in Asian countries, 110 pneumococcal isolates were analyzed by multilocus sequence typing (MLST). MLST analysis showed that 2 clonal complexes--CC236 (Taiwan19F-14 clone) and CC81 (Spain23F-1 clone)--are the major lineages of erythromycin-resistant S. pneumoniae in the Asian region. Pneumococcal isolates containing both the erm(B) and the mef(A) genes are thought to have originated from the Taiwan19F-14 clone containing the mef(A) gene, after introduction of the erm(B) gene. Further evolution of this variant clone has generated resistant strains with different sequence types. Dissemination of these variant clones of the Taiwan19F-14 could be the main reason for the high frequency of pneumococcal isolates containing both erm(B) and mef(A) in some Asian countries. Data suggest that the high prevalence of erythromycin-resistant S. pneumoniae in the Asian region is partly due to the clonal spread of a few multidrug-resistant clones. PMID- 15272403 TI - Activity of Staphylococcus epidermidis phenol-soluble modulin peptides expressed in Staphylococcus carnosus. AB - Staphylococcus epidermidis releases a group of peptides termed phenol-soluble modulin (PSM) that stimulate macrophages. The structure of 3 peptides (PSM alpha, PSM beta, and PSM gamma ) have been described. We report a fourth peptide (PSM delta ), which is a 23mer with the structure fMSIVSTIIEVVKTIVDIVKKFKK. The gene for each of the 4 peptides was introduced singly into Staphylococcus carnosus, and the PSM-like activity of culture medium and bacterial extract were significantly greater than those of the parent strain. PSM peptides from each of the S. carnosus-expressing strains were purified and analyzed by liquid chromatography-mass spectrometry. The products, which appeared to form aggregates, were active in the activation of human immunodeficiency virus type 1 long-terminal repeat and the production of tumor necrosis factor- alpha by the macrophage cell line THP-1. These findings suggest that PSM peptides are responsible, in part, for the modulin-like activity of staphylococci and may contribute to the development of severe staphylococcal sepsis. PMID- 15272404 TI - Increased Interleukin-4 production by CD8 and gammadelta T cells in health-care workers is associated with the subsequent development of active tuberculosis. AB - We evaluated immune responses to Mycobacterium tuberculosis in 10 health-care workers (HCWs) and 10 non-HCWs and correlated their immune status with the development of active tuberculosis (TB). Twenty individuals were randomly recruited, tested, and monitored longitudinally for TB presentation. Peripheral blood mononuclear cells (PBMCs) from donors were stimulated with M. tuberculosis and tested for cell proliferation and the production of interferon (IFN)- gamma, interleukin (IL)-5, and IL-4, by use of enzyme-linked immunosorbent or flow cytometric assays. HCWs had higher levels of cell proliferation (24,258 cpm) and IFN- gamma (6373 pg/mL) to M. tuberculosis than did non-HCWs (cell proliferation, 11,462 cpm; IFN- gamma, 3228 pg/mL). Six of 10 HCWs showed increased median percentages of CD8+IL-4+ (4.7%) and gammadelta +IL-4+ (2.3%) T cells and progressed to active TB. HCWs who remained healthy showed increased median percentages of CD8+IFN- gamma+ (25.0%) and gammadelta +IFN- gamma+ (8.0%) and lower percentages of CD8+IL-4+ (0.05%) and gammadelta +IL-4+ (0.03%) T cells. PMID- 15272405 TI - Immunization with the C-Domain of alpha -Toxin prevents lethal infection, localizes tissue injury, and promotes host response to challenge with Clostridium perfringens. AB - Clostridium perfringens gas gangrene is characterized by rapid tissue destruction, impaired host response, and, often, death. Phospholipase C (alpha toxin) is the virulence factor most responsible for these pathologies. The present study investigated the efficacy of active immunization with the C terminal domain of alpha -toxin (Cpa247-370) in a murine model of gas gangrene. Primary end points of the study were survival, progression of infection, and tissue perfusion. Secondary end points, which were based on findings of histologic evaluation of tissues, included the extent of tissue destruction and microvascular thrombosis, as well as the magnitude of the tissue inflammatory response. Survival among C-domain-immunized animals was significantly greater than that among sham-immunized control animals. Furthermore, immunization with the C-domain localized the infection and prevented ischemia of the feet. Histopathologic findings demonstrated limited muscle necrosis, reduced microvascular thrombosis, and enhanced granulocytic influx in C-domain-immunized mice. We conclude that immunization with the C-domain of phospholipase C is a viable strategy for the prevention of morbidity and mortality associated with C. perfringens gas gangrene. PMID- 15272406 TI - Induction of protective immunity against lethal anthrax challenge with a patch. AB - BACKGROUND: Transcutaneous immunization (TCI) is a needle-free technique that delivers antigens and adjuvants to potent epidermal immune cells. To address critical unmet needs in biodefense against anthrax, we have designed a novel vaccine delivery system using a dry adhesive patch that simplifies administration and improves tolerability of a subunit anthrax vaccine. METHODS: Mice and rabbits were vaccinated with recombinant protective antigen of Bacillus anthracis and the heat-labile toxin of Escherichia coli. Serologic changes, levels of toxin neutralizing antibodies (TNAs), and pulmonary and nodal responses were monitored in the mice. A lethal aerosolized B. anthracis challenge model was used in A/J mice, to demonstrate efficacy. RESULTS: The level of systemic immunity and protection induced by TCI was comparable to that induced by intramuscular vaccination, and peak immunity could be achieved with only 2 doses. The addition of adjuvant in the patch induced superior TNA levels, compared with injected vaccination. CONCLUSIONS: Anthrax vaccine patches stimulated robust and functional immune responses that protected against lethal challenge. Demonstration of responses in the lung suggests that a mechanism exists for protection against challenge with aerosolized anthrax spores. A formulated, pressure-sensitive, dry adhesive patch, which is stable and can be manufactured in large scale, elicited comparable immunoglobulin G and TNA responses, suggesting that an anthrax vaccine patch is feasible and should advance into clinical evaluation. PMID- 15272407 TI - Transmission of Yersinia pestis from an infectious biofilm in the flea vector. AB - Transmission of plague by fleas depends on infection of the proventricular valve in the insect's foregut by a dense aggregate of Yersinia pestis. Proventricular infection requires the Y. pestis hemin storage (hms) genes; here, we show that the hms genes are also required to produce an extracellular matrix and a biofilm in vitro, supporting the hypothesis that a transmissible infection in the flea depends on the development of a biofilm on the hydrophobic, acellular surface of spines that line the interior of the proventriculus. The development of biofilm and proventricular infection did not depend on the 3 Y. pestis quorum-sensing systems. The extracellular matrix enveloping the Y. pestis biofilm in the flea appeared to incorporate components from the flea's blood meal, and bacteria released from the biofilm were more resistant to human polymorphonuclear leukocytes than were in vitro-grown Y. pestis. Enabling arthropod-borne transmission represents a novel function of a bacterial biofilm. PMID- 15272408 TI - Rashes occurring after immunization with a mixture of viruses in the Oka vaccine are derived from single clones of virus. AB - Vaccination against chickenpox causes a varicella-like rash in up to 5% of healthy children and 50% of children with leukemia. The vaccine may establish latency and reactivate to cause herpes zoster, albeit more rarely than wild-type virus. All vaccine preparations are composed of a mixture of varicella-zoster virus strains that show genotypic variation at several loci. We have shown, by DNA sequencing of 40 polymorphic loci, that viruses sampled from vesicles in varicella-like and herpes zoster rashes are single clones. This finding suggests that, between the time of inoculation of the vaccine and development of rash, selection of single strains occurs. The results have general implications for the pathogenesis of varicella-zoster virus. PMID- 15272409 TI - Disease-specific B Cell epitopes for serum antibodies from patients with severe acute respiratory syndrome (SARS) and serologic detection of SARS antibodies by epitope-based peptide antigens. AB - Severe acute respiratory syndrome (SARS) has emerged as a highly contagious, sometimes fatal disease. To find disease-specific B cell epitopes, phage displayed random peptide libraries were panned on serum immunoglobulin (Ig) G antibodies from patients with SARS. Forty-nine immunopositive phage clones that bound specifically to serum from patients with SARS were selected. These phageborne peptides had 4 consensus motifs, of which 2 corresponded to amino acid sequences reported for SARS-associated coronavirus (SARS-CoV). Synthetic peptide binding and competitive-inhibition assays further confirmed that patients with SARS generated antibodies against SARS-CoV. Immunopositive phage clones and epitope-based peptide antigens demonstrated clinical diagnostic potential by reacting with serum from patients with SARS. Antibody-response kinetics were evaluated in 4 patients with SARS, and production of IgM, IgG, and IgA were documented as part of the immune response. In conclusion, B cell epitopes of SARS corresponded to novel coronavirus. Our epitope-based serologic test may be useful in laboratory detection of the virus and in further study of the pathogenesis of SARS. PMID- 15272410 TI - Size and charge characteristics of the protein leak in dengue shock syndrome. AB - BACKGROUND: he mechanism underlying the transient vascular leak syndrome of dengue hemorrhagic fever (DHF) is unknown. We aimed to determine whether molecular size and charge selectivity, which help restrict plasma proteins within the intravascular space, are altered in patients with DHF and whether a disturbance of the anionic glycosaminoglycan (GAG) layer on the luminal endothelial surface contributes to disease pathogenesis. METHODS: We measured serial plasma levels and fractional clearances of proteins with different size and charge characteristics in 48 children with dengue shock syndrome (DSS) and urinary excretion profiles of heparan sulfate, chondroitin-4-sulfate, and chondroitin-6-sulfate in affected children and healthy control subjects. RESULTS: Compared with convalescent values, acute plasma concentrations of all proteins were reduced, with increased fractional clearances. Smaller proteins were more affected than larger molecules. Albumin, which is normally protected from leakage by its strong negative charge, demonstrated a clearance pattern similar to that of transferrin, a neutral molecule of similar size. Urinary heparan sulfate excretion was significantly increased in children with DSS. CONCLUSIONS: The endothelial size-dependent sieving mechanism for plasma proteins is at least partially retained, whereas selective restriction based on negative charge is impaired. The increased heparan sulfate excretion suggests a role for GAGs in the pathogenesis of the vascular leak. PMID- 15272411 TI - Iron regulates hepatitis C virus translation via stimulation of expression of translation initiation factor 3. AB - BACKGROUND: Although the response to treatment with interferon- alpha in individuals with chronic hepatitis C virus (HCV) infection is negatively associated with increased liver iron stores, the underlying mechanisms at work have remained elusive to date. The translation initiation factor 3 (eIF3) is essential for HCV translation, and thus the effects that iron perturbations have on eIF3 expression and HCV translation were studied here. METHODS: eIF3 expression was analyzed by TaqMan polymerase chain reaction, Northern and Western blot analysis of HepG2 cells, and liver biopsies. Functional effects of iron on HCV mRNA translation were estimated by use of transient transfection experiments with bicistronic vectors. RESULTS: Iron treatment of HepG2 cells increased eIF3 mRNA and protein expression, whereas iron chelation reduced it. Accordingly, iron dependent stimulation of eIF3 specifically induced the expression of reporter genes under the control of regulatory HCV mRNA stem-loop structures. Moreover, a positive association between liver iron levels, eIF3 expression, and HCV expression was found when liver-biopsy samples from HCV-infected patients were analyzed. CONCLUSION: Iron promotes the translation of HCV by stimulating the expression of eIF3, which may be one reason for the negative association between liver iron overload and HCV infection. Modulation of the affinity of eIF3 to bind to HCV mRNA may be a promising target for the treatment of chronic HCV infection. PMID- 15272412 TI - Viral and bacterial pathogens at the maternal-fetal interface. AB - We studied the incidence of pathogenic bacteria and concurrent infections with human cytomegalovirus (CMV) and herpes simplex virus (HSV) type 1 and 2 in biopsy samples from the placenta and decidua of women with healthy pregnancies. By polymerase chain reaction analysis, we found that 38% of placental samples were positive for selected bacteria and viruses. CMV, HSV-1, and HSV-2 were detected in isolation or with bacteria in first- and second-trimester samples. Certain bacteria were detected more often during the second trimester than during the first--Ureaplasma urealyticum, Mycoplasma hominis, and Gardnerella/Bifidobacterium species. In paired samples from first-trimester tissues, the detection rate for viruses, compared with most bacteria, was higher in the decidua than in the adjacent placenta. In contrast, bacteria were more frequently detected in placenta. Analyses of immunoglobulin G isolated from the placenta support the hypothesis that immune responses suppress CMV reactivation in the presence of pathogenic bacteria at the maternal-fetal interface. PMID- 15272413 TI - Increases in human T helper 2 cytokine responses to Schistosoma mansoni worm and worm-tegument antigens are induced by treatment with praziquantel. AB - Levels of Schistosoma mansoni-induced interleukin (IL)-4 and IL-5 and posttreatment levels of immunoglobulin E recognizing the parasite's tegument (Teg) correlate with human resistance to subsequent reinfection after treatment. We measured changes in whole-blood cytokine production in response to soluble egg antigen (SEA), soluble worm antigen (SWA), or Teg after treatment with praziquantel (PZQ) in a cohort of 187 individuals living near Lake Albert, Uganda. Levels of SWA-induced IL-4, IL-5, IL-10, and IL-13 increased after treatment with PZQ, and the greatest relative increases were seen in the responses to Teg. Mean levels of Teg-specific IL-5 and IL-10 increased ~10-15 fold, and mean levels of IL-13 increased ~5-fold. Correlations between the changes in cytokines suggested that their production was positively coregulated by tegumentally derived antigens. Levels of SEA-, SWA-, and Teg-induced interferon- gamma were not significantly changed by treatment, and, with the exception of IL-10, which increased slightly, responses to SEA also remained largely unchanged. The changes in cytokines were not strongly influenced by age or intensity of infection and were not accompanied by corresponding increases in the numbers of circulating eosinophils or lymphocytes. PMID- 15272414 TI - Inhibition of heme aggregation by chloroquine reduces Schistosoma mansoni infection. AB - Adult Schistosoma mansoni digest large amounts of host hemoglobin and release potentially toxic heme inside their guts. We have previously demonstrated that free heme in S. mansoni is detoxified through aggregation, forming hemozoin (Hz). Possible mechanisms of heme aggregation and the effects of chloroquine (CLQ) on formation of Hz and on the viability of this parasite have now been investigated. Different fractions isolated from S. mansoni, such as crude whole-worm homogenates, total lipid extracts, and Hz itself promoted heme aggregation in vitro in a CLQ-sensitive manner. Treatment of S. mansoni-infected mice with CLQ led to remarkable decreases in total protein, Hz content, and viability of the worms, as well as in parasitemia and deposition of eggs in mouse livers. These results indicate that inhibition of formation of Hz in S. mansoni, by CLQ, led to an important decrease in the overall severity of experimental murine schistosomiasis. Taken together, the results presented here suggest that formation of Hz is a major mechanism of heme detoxification and a potential target for chemotherapy in S. mansoni. PMID- 15272415 TI - Molecular diagnosis of resistance to antimalarial drugs during epidemics and in war zones. AB - Plasmodium falciparum mutations pfcrt K76T and the dhfr/dhps "quintuple mutant" are molecular markers of resistance to chloroquine and sulfadoxine-pyrimethamine, respectively. During an epidemic of P. falciparum malaria in an area of political unrest in northern Mali, where standard efficacy studies have been impossible, we measured the prevalence of these markers in a cross-sectional survey. In 80% of cases of infection, pfcrt K76T was detected, but none of the cases carried the dhfr/dhps quintuple mutant. On the basis of these results, chloroquine was replaced by sulfadoxine-pyrimethamine in control efforts. This example illustrates how molecular markers for drug resistance can provide timely data that inform malaria-control policy during epidemics and other emergency situations. PMID- 15272416 TI - Detection of antigen in bronchial epithelium and macrophages in acute Kawasaki disease by use of synthetic antibody. AB - BACKGROUND: Kawasaki disease (KD) is the most common acquired cardiac disease in children in developed nations. The etiology is unknown, but a ubiquitous infectious agent appears to be likely. Immunoglobulin A plasma cells infiltrate inflamed tissues in acute KD, producing oligoclonal, antigen-driven antibodies. METHODS: To identify antigens important in the pathogenesis of KD, oligoclonal KD antibodies were prepared in vitro and tested by immunohistochemistry experiments on tissues from patients with acute KD and from control subjects and were also tested for reactivity with human inflammatory proteins. RESULTS: By use of synthetic antibody A, specific binding to a cytoplasmic antigen in proximal bronchial epithelium was observed in 10 of 13 patients with acute KD but in 0 of 9 control subjects (P=.001). A subset of macrophages was positive in at least 1 inflamed tissue from all 17 patients with acute KD. Antigen was detected in 9 of 12 acute KD coronary artery aneurysms but in 0 of 10 control coronary arteries (P<.001). The antigen is not immunoglobulin or any of 40 common inflammatory proteins. CONCLUSIONS: We report the first demonstration of a KD-associated antigen in the tissues targeted by the disease. Our findings are consistent with the theory that KD is caused by a previously unidentified respiratory infectious agent with tropism for vascular tissue. PMID- 15272417 TI - A major lung cancer susceptibility locus maps to chromosome 6q23-25. AB - Lung cancer is a major cause of death in the United States and other countries. The risk of lung cancer is greatly increased by cigarette smoking and by certain occupational exposures, but familial factors also clearly play a major role. To identify susceptibility genes for familial lung cancer, we conducted a genomewide linkage analysis of 52 extended pedigrees ascertained through probands with lung cancer who had several first-degree relatives with the same disease. Multipoint linkage analysis, under a simple autosomal dominant model, of all 52 families with three or more individuals affected by lung, throat, or laryngeal cancer, yielded a maximum heterogeneity LOD score (HLOD) of 2.79 at 155 cM on chromosome 6q (marker D6S2436). A subset of 38 pedigrees with four or more affected individuals yielded a multipoint HLOD of 3.47 at 155 cM. Analysis of a further subset of 23 multigenerational pedigrees with five or more affected individuals yielded a multipoint HLOD score of 4.26 at the same position. The 14 families with only three affected relatives yielded negative LOD scores in this region. A predivided samples test for heterogeneity comparing the LOD scores from the 23 multigenerational families with those from the remaining families was significant (P=.007). The 1-HLOD multipoint support interval from the multigenerational families extends from C6S1848 at 146 cM to 164 cM near D6S1035, overlapping a genomic region that is deleted in sporadic lung cancers as well as numerous other cancer types. Parametric linkage and variance-components analysis that incorporated effects of age and personal smoking also supported linkage in this region, but with somewhat diminished support. These results localize a major susceptibility locus influencing lung cancer risk to 6q23-25. PMID- 15272418 TI - Replication study supports evidence for linkage to 9p24 in obsessive-compulsive disorder. AB - Obsessive-compulsive disorder (OCD) is a severe psychiatric illness that is characterized by intrusive and senseless thoughts and impulses (obsessions) and by repetitive behaviors (compulsions). Family, twin, and segregation studies support the presence of both genetic and environmental susceptibility factors, and the only published genome scan for OCD identified a candidate region on 9p24 at marker D9S288 that met criteria for suggestive significance (Hanna et al. 2002). In an attempt to replicate this finding, we genotyped 50 pedigrees with OCD, using microsatellite markers spanning the 9p24 candidate region, and analyzed the data, using parametric and nonparametric linkage analyses under both a narrow phenotype model (DSM-IV OCD definite; 41 affected sib pairs) and a broad phenotype model (DSM-IV OCD definite and probable; 50 affected sib pairs). Similar to what was described by Hanna et al. (2002), our strongest findings came with the dominant parameters and the narrow phenotype model: the parametric signal peaked at marker D9S1792 with an HLOD of 2.26 ( alpha =0.59), and the nonparametric linkage signal (NPL) peaked at marker D9S1813 with an NPL of 2.52 (P=.006). These findings are striking in that D9S1813 and D9S1792 lie within 0.5 cM (<350 kb) of the original 9p24 linkage signal at D9S288; furthermore, pedigree based association analyses also implicated the 9p24 candidate region by identifying two markers (D9S288 and GATA62F03) with modest evidence (P=.046 and .02, respectively) for association. PMID- 15272419 TI - The future of association studies: gene-based analysis and replication. AB - Historically, association tests were limited to single variants, so that the allele was considered the basic unit for association testing. As marker density increases and indirect approaches are used to assess association through linkage disequilibrium, association is now frequently considered at the haplotypic level. We suggest that there are difficulties in replicating association findings at the single-nucleotide-polymorphism (SNP) or the haplotype level, and we propose a shift toward a gene-based approach in which all common variation within a candidate gene is considered jointly. Inconsistencies arising from population differences are more readily resolved by use of a gene-based approach rather than either a SNP-based or a haplotype-based approach. A gene-based approach captures all of the potential risk-conferring variations; thus, negative findings are subject only to the issue of power. In addition, chance findings due to multiple testing can be readily accounted for by use of a genewide-significance level. Meta-analysis procedures can be formalized for gene-based methods through the combination of P values. It is only a matter of time before all variation within genes is mapped, at which point the gene-based approach will become the natural end point for association analysis and will inform our search for functional variants relevant to disease etiology. PMID- 15272422 TI - Neonatal anesthesia. AB - The physiology of the preterm and term neonate is characterized by a high metabolic rate, limited pulmonary, cardiac and thermoregulatory reserve, and decreased renal function. Multisystem immaturity creates important developmental differences in drug handling and response when compared to the older child or adult. Neonatal anesthetic management requires an understanding of the pharmacophysiologic limitations of the neonate as well as the pathophysiology of coexisting surgical disease. This review addresses the pertinent aspects of neonatal physiology and pharmacology, general considerations in the anesthetic care of surgical neonates, and concludes with a brief review of the anesthetic management of neonates with necrotizing enterocolitis, diaphragmatic hernia, and tracheoesophageal fistula. PMID- 15272420 TI - A genomewide scan for early-onset coronary artery disease in 438 families: the GENECARD Study. AB - A family history of coronary artery disease (CAD), especially when the disease occurs at a young age, is a potent risk factor for CAD. DNA collection in families in which two or more siblings are affected at an early age allows identification of genetic factors for CAD by linkage analysis. We performed a genomewide scan in 1,168 individuals from 438 families, including 493 affected sibling pairs with documented onset of CAD before 51 years of age in men and before 56 years of age in women. We prospectively defined three phenotypic subsets of families: (1) acute coronary syndrome in two or more siblings; (2) absence of type 2 diabetes in all affected siblings; and (3) atherogenic dyslipidemia in any one sibling. Genotypes were analyzed for 395 microsatellite markers. Regions were defined as providing evidence for linkage if they provided parametric two-point LOD scores >1.5, together with nonparametric multipoint LOD scores >1.0. Regions on chromosomes 3q13 (multipoint LOD = 3.3; empirical P value <.001) and 5q31 (multipoint LOD = 1.4; empirical P value <.081) met these criteria in the entire data set, and regions on chromosomes 1q25, 3q13, 7p14, and 19p13 met these criteria in one or more of the subsets. Two regions, 3q13 and 1q25, met the criteria for genomewide significance. We have identified a region on chromosome 3q13 that is linked to early-onset CAD, as well as additional regions of interest that will require further analysis. These data provide initial areas of the human genome where further investigation may reveal susceptibility genes for early-onset CAD. PMID- 15272423 TI - The pediatric airway. AB - Good airway management technique is an essential skill for physicians in most specialties. This article begins with a review of basic airway anatomy and the physiology of the uninstrumented airway. This subject is of particular importance given the increasing use of procedural sedation and the increased recognition of sleep-disordered breathing in infants and children. A discussion of the various artificial airways and their advantages and disadvantages follows. The difficult airway is an important contributor to both patient morbidity and mortality. It is important to have a planned management approach available for the anticipated and, more importantly, the unanticipated difficult airway. The recommendations of the American Society of Anesthesiologists Taskforce on the Management of the Difficult Airway have good application for this important problem. The fetus with the prenatal diagnosis of a lesion that predicts a difficult airway presents a particular challenge. The utilization of an ex-utero intrapartum treatment method is presented as an important approach for the delivery and airway management of these infants. This section closes with a discussion of the prehospital airway management of the pediatric patient. PMID- 15272424 TI - Sedation and analgesia for procedures outside the operating room. AB - The volume of literature concerning sedation and analgesia for procedures outside the operating room has increased greatly over the past several years. Information relating to sedation risks and complications, the development of sedation guidelines, and now specific sedation techniques has appeared. The following section will trace the development of the most recent sedation guidelines and identify the key elements for inclusion into institutional sedation guidelines. The risks and complications associated with sedation will be addressed and an approach to providing sedation and analgesia for procedures outside the operating room will be presented. The important characteristics of several common drugs used for sedation and analgesia will also be discussed. PMID- 15272425 TI - Perioperative implications of common respiratory problems. AB - Pediatric surgical patients can present with a number of challenging common respiratory problems. This article reviews potential perioperative implications and anesthetic management of asthma, upper respiratory tract infections, bronchopulmonary dysplasia, and the effects of passive environmental smoke on children presenting for surgery. PMID- 15272426 TI - Anesthesia for minimally invasive surgery. AB - Minimally invasive surgery has become the surgical approach of choice for a number of operative procedures in adults, but remains a challenge in children, especially the youngest, due to the physiologic changes that occur during the procedure as well as limitations posed by equipment. These surgical techniques can present unique problems to the pediatric anesthesiologist in designing and implementing the anesthetic management plan. In this article, we will outline the physiologic impact, anesthetic management, and potential challenges and complications associated with minimally invasive surgery in children. PMID- 15272427 TI - Single-ventricle physiology: perioperative implications. AB - Neonates with functional single ventricles have pulmonary and systemic circulations that are supplied in parallel, creating significant cyanosis and ventricular volume overload. The goal of palliative surgery, excluding transplantation, is to convert single-ventricle circulation from a parallel to a series arrangement. This will ultimately require a complete cavopulmonary anastomosis (Fontan-type procedure) in which vena caval blood is rerouted directly into the pulmonary circulation. Various factors require that this palliation occur in stages. Stage I surgery, which is often a Norwood procedure, is done in the neonatal period and stabilizes, but does not resolve, parallel circulation. The tenuous balance between pulmonary and systemic perfusion during this stage makes noncardiac surgery hazardous, and it should be restricted to urgent or emergent indications. Stage II surgery, or partial cavopulmonary anastomosis, relieves both parallel circulation and volume overload, but not cyanosis. Relatively stable hemodynamics during this stage create favorable conditions for elective surgery. Patients who have undergone stage III surgery, the Fontan-type repair, vary in age from toddlers to adults, and in physical status from well-compensated to significantly debilitated. Fontan patients require thorough preoperative assessment when elective surgery is contemplated. Optimal communication between surgeons, anesthesiologists, and cardiologists is essential when caring for the patient with single-ventricle physiology. PMID- 15272428 TI - Postoperative neuraxial pain relief in the pediatric patient. AB - Pain serves as a useful warning function of potential tissue damage. The systemic response to pain is characterized by activation of the sympathetic nervous system. The ensuing neuroendocrine response results in a myriad of adverse effects on the various organ systems, which can adversely affect surgical outcomes. When effective neuraxial anesthesia and analgesic techniques are employed, the surgical patient may benefit from decreases in morbidity, mortality, and prevention of subsequent development of chronic pain. The use of intrathecal and epidural techniques in the pediatric population is well established and allows these patients to benefit from superior pain relief and improved surgical outcomes. PMID- 15272429 TI - Anesthetic considerations for pediatric outpatient surgery. AB - At least 60% of all pediatric surgical procedures are performed as outpatients. Successful outpatient practice requires that both the patients and the procedures are appropriate for outpatient management. Appropriate scheduling ensures that the overall function of the outpatient unit is not compromised. While not all scheduled patients need be perfectly healthy, care must be taken to provide adequate resources to those patients who are not, without diverting resources away from other patients. Anesthesia must be managed with attention to the unique physiologic and psychological challenges of children. Anesthetic management of postoperative nausea and vomiting and provision of adequate postoperative analgesia, including regional techniques, are important elements of the care plan. PMID- 15272430 TI - Prokaryote phylogeny without sequence alignment: from avoidance signature to composition distance. AB - This is a review of a new and essentially simple method of inferring phylogenetic relationships from complete genome data without using sequence alignment. The method is based on counting the appearance frequency of oligopeptides of a fixed length (up to K = 6) in the collection of protein sequences of a species. It is a method without fine adjustment and choice of genes. Applied to prokaryotic genomes it has led to results comparable with the bacteriologists' systematics as reflected in the latest 2002 outline of the Bergey's Manual of Systematic Bacteriology. The method has also been used to compare chloroplast genomes and to the phylogeny of Coronaviruses including human SARS-CoV. A key point in our approach is subtraction of a random background from the original counts by using a Markov model of order K-2 in order to highlight the shaping role of natural selection. The implications of the subtraction procedure is specially analyzed and further development of the new approach is indicated. PMID- 15272431 TI - Computing highly specific and noise-tolerant oligomers efficiently. AB - The sequencing of the genomes of a variety of species and the growing databases containing expressed sequence tags (ESTs) and complementary DNAs (cDNAs) facilitate the design of highly specific oligomers for use as genomic markers, PCR primers, or DNA oligo microarrays. The first step in evaluating the specificity of short oligomers of about 20 units in length is to determine the frequencies at which the oligomers occur. However, for oligomers longer than about fifty units this is not efficient, as they usually have a frequency of only 1. A more suitable procedure is to consider the mismatch tolerance of an oligomer, that is, the minimum number of mismatches that allows a given oligomer to match a substring other than the target sequence anywhere in the genome or the EST database. However, calculating the exact value of mismatch tolerance is computationally costly and impractical. Therefore, we studied the problem of checking whether an oligomer meets the constraint that its mismatch tolerance is no less than a given threshold. Here, we present an efficient dynamic programming algorithm solution that utilizes suffix and height arrays. We demonstrated the effectiveness of this algorithm by efficiently computing a dense list of numerous oligo-markers applicable to the human genome. Experimental results show that the algorithm runs faster than well-known Abrahamson's algorithm by orders of magnitude and is able to enumerate 65% approximately 76% of qualified oligomers. PMID- 15272432 TI - cWINNOWER algorithm for finding fuzzy dna motifs. AB - The cWINNOWER algorithm detects fuzzy motifs in DNA sequences rich in protein binding signals. A signal is defined as any short nucleotide pattern having up to d mutations differing from a motif of length l. The algorithm finds such motifs if a clique consisting of a sufficiently large number of mutated copies of the motif (i.e., the signals) is present in the DNA sequence. The cWINNOWER algorithm substantially improves the sensitivity of the winnower method of Pevzner and Sze by imposing a consensus constraint, enabling it to detect much weaker signals. We studied the minimum detectable clique size qc as a function of sequence length N for random sequences. We found that qc increases linearly with N for a fast version of the algorithm based on counting three-member sub-cliques. Imposing consensus constraints reduces qc by a factor of three in this case, which makes the algorithm dramatically more sensitive. Our most sensitive algorithm, which counts four-member sub-cliques, needs a minimum of only 13 signals to detect motifs in a sequence of length N = 12,000 for (l, d) = (15, 4). PMID- 15272433 TI - Statistical and visual morph movie analysis of crystallographic mutant selection bias in protein mutation resource data. AB - Structural studies of the effects of non-silent mutations on protein conformational change are an important key in deciphering the language that relates protein amino acid primary structure to tertiary structure. Elsewhere, we presented the Protein Mutant Resource (PMR) database, a set of online tools that systematically identified groups of related mutant structures in the Protein DataBank (PDB), accurately inferred mutant classifications in the Gene Ontology using an innovative, statistically rigorous data-mining algorithm with more general applicability, and illustrated the relationship of these mutant structures via an intuitive user interface. Here, we perform a comprehensive statistical analysis of the effect of PMR mutations on protein tertiary structure. We find that, although the PMR does contain spectacular examples of conformational change, in general there is a counter-intuitive inverse relationship between conformational change (measured as C-alpha displacement or RMS of the core structure) and the number of mutations in a structure. That is, point mutations by structural biologists present in the PDB contrast naturally evolved mutations. We compare the frequency of mutations in the PMR/PDB datasets against the accepted PAM250 natural amino acid mutation frequency to confirm these observations. We generated morph movies from PMR structure pairs using technology previously developed for the Macromolecular Motions Database (http://molmovdb.org), allowing bioinformaticians, geneticists, protein engineers, and rational drug designers to analyze visually the mechanisms of protein conformational change and distinguish between conformational change due to motions (e.g., ligand binding) and mutations. The PMR morph movies and statistics can be freely viewed from the PMR website, http://pmr.sdsc.edu. PMID- 15272435 TI - Index-based similarity search for protein structure databases. AB - We propose new methods for finding similarities in protein structure databases. These methods extract feature vectors on triplets of SSEs (Secondary Structure Elements) of proteins. The feature vectors are then indexed using a multidimensional index structure. Our first technique considers the problem of finding proteins similar to a given query protein in a protein dataset. It quickly finds promising proteins using the index structure. These proteins are then aligned to the query protein using a popular pairwise alignment tool such as VAST. We also develop a novel statistical model to estimate the goodness of a match using the SSEs. Our second technique considers the problem of joining two protein datasets to find an all-to-all similarity. Experimental results show that our techniques improve the pruning time of VAST 3 to 3.5 times, while keeping the sensitivity similar. Our technique can also be incorporated with DALI and CE to improve their running times by a factor of 2 and 2.7 respectively. The software is available online at http://bioserver.cs.ucsb.edu/. PMID- 15272434 TI - Combining microarrays and biological knowledge for estimating gene networks via bayesian networks. AB - We propose a statistical method for estimating a gene network based on Bayesian networks from microarray gene expression data together with biological knowledge including protein-protein interactions, protein-DNA interactions, binding site information, existing literature and so on. Microarray data do not contain enough information for constructing gene networks accurately in many cases. Our method adds biological knowledge to the estimation method of gene networks under a Bayesian statistical framework, and also controls the trade-off between microarray information and biological knowledge automatically. We conduct Monte Carlo simulations to show the effectiveness of the proposed method. We analyze Saccharomyces cerevisiae gene expression data as an application. PMID- 15272436 TI - Logos: a modular bayesian model for de novo motif detection. AB - The complexity of the global organization and internal structure of motifs in higher eukaryotic organisms raises significant challenges for motif detection techniques. To achieve successful de novo motif detection, it is necessary to model the complex dependencies within and among motifs and to incorporate biological prior knowledge. In this paper, we present LOGOS, an integrated LOcal and GlObal motif Sequence model for biopolymer sequences, which provides a principled framework for developing, modularizing, extending and computing expressive motif models for complex biopolymer sequence analysis. LOGOS consists of two interacting submodels: HMDM, a local alignment model capturing biological prior knowledge and positional dependency within the motif local structure; and HMM, a global motif distribution model modeling frequencies and dependencies of motif occurrences. Model parameters can be fit using training motifs within an empirical Bayesian framework. A variational EM algorithm is developed for de novo motif detection. LOGOS improves over existing models that ignore biological priors and dependencies in motif structures and motif occurrences, and demonstrates superior performance on both semi-realistic test data and cis regulatory sequences from yeast and Drosophila genomes with regard to sensitivity, specificity, flexibility and extensibility. PMID- 15272438 TI - Optimal, efficient reconstruction of phylogenetic networks with constrained recombination. AB - A phylogenetic network is a generalization of a phylogenetic tree, allowing structural properties that are not tree-like. In a seminal paper, Wang et al.(1) studied the problem of constructing a phylogenetic network, allowing recombination between sequences, with the constraint that the resulting cycles must be disjoint. We call such a phylogenetic network a "galled-tree". They gave a polynomial-time algorithm that was intended to determine whether or not a set of sequences could be generated on galled-tree. Unfortunately, the algorithm by Wang et al.(1) is incomplete and does not constitute a necessary test for the existence of a galled-tree for the data. In this paper, we completely solve the problem. Moreover, we prove that if there is a galled-tree, then the one produced by our algorithm minimizes the number of recombinations over all phylogenetic networks for the data, even allowing multiple-crossover recombinations. We also prove that when there is a galled-tree for the data, the galled-tree minimizing the number of recombinations is "essentially unique". We also note two additional results: first, any set of sequences that can be derived on a galled tree can be derived on a true tree (without recombination cycles), where at most one back mutation per site is allowed; second, the site compatibility problem (which is NP hard in general) can be solved in polynomial time for any set of sequences that can be derived on a galled tree. Perhaps more important than the specific results about galled-trees, we introduce an approach that can be used to study recombination in general phylogenetic networks. This paper greatly extends the conference version that appears in an earlier work.(8) PowerPoint slides of the conference talk can be found at our website.(7). PMID- 15272437 TI - An empirical study of the universal chemical key algorithm for assigning unique keys to chemical compounds. AB - In this paper, we introduce an algorithm that assigns an essentially unique key called the Universal Chemical Key (UCK) to molecular structures. The molecular structures are represented as labeled graphs whose nodes abstract atoms and whose edges abstract bonds. The algorithm was tested on 236,917 compounds obtained from the National Cancer Institute (NCI) database of chemical compounds. On this database, the UCK algorithm assigned unique keys for chemicals with distinct molecular structures. PMID- 15272439 TI - Protein structure alignment and fast similarity search using local shape signatures. AB - We present a new method for conducting protein structure similarity searches, which improves on the efficiency of some existing techniques. Our method is grounded in the theory of differential geometry on 3D space curve matching. We generate shape signatures for proteins that are invariant, localized, robust, compact, and biologically meaningful. The invariancy of the shape signatures allows us to improve similarity searching efficiency by adopting a hierarchical coarse-to-fine strategy. We index the shape signatures using an efficient hashing based technique. With the help of this technique we screen out unlikely candidates and perform detailed pairwise alignments only for a small number of candidates that survive the screening process. Contrary to other hashing based techniques, our technique employs domain specific information (not just geometric information) in constructing the hash key, and hence, is more tuned to the domain of biology. Furthermore, the invariancy, localization, and compactness of the shape signatures allow us to utilize a well-known local sequence alignment algorithm for aligning two protein structures. One measure of the efficacy of the proposed technique is that we were able to perform structure alignment queries 36 times faster (on the average) than a well-known method while keeping the quality of the query results at an approximately similar level. PMID- 15272446 TI - Ethanol-inducible gene expression: first step towards a new green revolution? AB - The introduction of dwarf varieties of cereals was fundamental to the green revolution, but in the post-genomic era, the manipulation of plant morphology could be more sophisticated. A recent publication by Tahar Ait-ali et al. describes the use of the ethanol-inducible transgene expression system to re examine plant architecture, and the genes that determine it. Their findings have implications for the manipulation of plant height and yield, and demonstrate the efficacy of regulated transgene expression for functional genomics. PMID- 15272447 TI - Research shows 'poppers' use in connected with infection among MSM. Little is done to address problem. AB - Researchers and experts on the use of inhaled nitrates among men who have sex with men (MSM) say clinicians, public health officials, and AIDS groups are ignoring the impact of poppers, a popular party inhalant, on HIV risk behaviors and even seroconversion. PMID- 15272448 TI - AIDS is a major killer of African-American women. Many women are in denial. AB - African-American women's share of AIDS cases has been growing steadily in the past decade, and now represents one-third of all new AIDS cases reported among African-Americans, CDC data show. The challenge for public health officials is developing prevention programs that do not treat African-American women as one monolithic group, according to Victoria Cargill, MD, of the Office of AIDS Research. PMID- 15272449 TI - Some innovative ideas for preventing HIV. Educators meet on their turf. AB - Clinicians, AIDS service organizations, and public health officials who are looking for new strategies for reaching African-American women through HIV prevention programs might find useful ideas in how two very different programs handle HIV education. PMID- 15272450 TI - Data reveal high sexual risk among Asian MSMs. More intervention work needed. AB - Investigators have found a disturbing trend of increased levels of sexual risk behavior among a small, little-studied group: Asian/Pacific Islander MSM. PMID- 15272451 TI - HIV and older Americans: forgotten in AIDS focus? Denial runs rampant in public perception of AIDS. AB - Sometimes clinicians and health officials who work with HIV-infected people who are older than 50 may feel like John the Baptist crying out in the wilderness: They talk and talk, and no one seems to be listening. But in Florida, their share of the epidemic runs to 14% of the population. PMID- 15272452 TI - FDA approves Hep C drugs. PMID- 15272453 TI - FDA notifications. Oral fluid-based rapid HIV test approved. PMID- 15272454 TI - Updated antiretroviral guidelines available. PMID- 15272455 TI - The disappearance of thymine dimers from DNA: an error-correcting mechanism. 1963. PMID- 15272456 TI - Release of ultraviolet light-induced thymine dimers from DNA in E. coli K-12. 1964. PMID- 15272457 TI - U.S. researcher starts treatment fund in Uganda: interview with David Bangsberg, M.D., M.P.H. AB - $400 a year will save a life in Uganda. A U.S. scientist who works there has created a fund that has guaranteed five years of treatment for ten people so far. PMID- 15272458 TI - President Bush on AIDS: more questions than answers. AB - The president spoke on AIDS June 23 in Philadelphia. His comments looked great in headlines, but details raised major questions. PMID- 15272459 TI - Abstinence, abstinence-only, faith-based, and the psychology of stigma. AB - If abstinence is 100% effective in preventing sexual transmission, why does abstinence-only not work well? And what is the personal psychology of the stigma that prevents individuals, communities, and nations from protecting themselves against the epidemic? We offer some fairly obvious analysis that has been largely overlooked in the public discussion. PMID- 15272460 TI - Medical marijuana: important vote coming, you can help. AB - Congress will vote soon on an amendment to stop Attorney General Ashcroft's crusade against medical marijuana, in states where laws recognize medical use. This amendment received 152 votes in Congress last year. PMID- 15272461 TI - ADVAX, new DNA vaccine in human trial; HIV-negative volunteers needed in New York City or Rochester, NY areas. AB - HIV-negative volunteers are needed for an important vaccine trial. PMID- 15272462 TI - [Growth, mortality and exploitation rate of Priacanthus arenatus (Perciformes: Priacanthidae), in the trawl fisheries of northeast Venezuela]. AB - We analyzed growth, mortality and exploitation rate of Priacanhus arenatus, captured by the shrimp trawling fishery (1989-1996), in northeastern Venezuela. The growth coefficient (K) and the asymptotic length (L8) were estimated by length-frequency data using the Battacharya method and other routines of the FISAT program. Total mortality (Z) and exploitation (E) rates were obtained by length-converted catch curve analysis, based on length-frequency data, and the Berverton and Holt's yield per recruit model, respectively. The mean growth parameters L and K were estimated as 474.7 mm and 0.69 year(-1), respectively. Mean total mortality was 4.03 and the exploitation rate range was 0.70-0.80. Results indicated that the population is overexploited. PMID- 15272463 TI - Comparison of the efficacy of a subunit and a live streptomycin-dependent porcine pleuropneumonia vaccine. AB - OBJECTIVE: To evaluate the efficacy of two new-generation porcine pleuropneumonia vaccines when challenged with Australian isolates of Actinobacillus pleuropneumoniae of serovars 1 and 15. DESIGN: The Porcilis APP vaccine and an experimental streptomycin-dependent strain of A pleuropneumoniae were evaluated in a standardised pen trial. Each vaccine/challenge group consisted of 10 pigs. RESULTS: With the serovar 1 challenge, the Porcilis APP vaccine and the live vaccine, compared with the control group, gave significant protection in terms of clinical signs, lung lesions, re-isolation scores and average daily gain (ADG) postchallenge. Only the Porcilis APP vaccine provided significant protection against mortality. In the serovar 15 challenged pigs, the only significant difference detected was that the Porcilis APP vaccinated pigs had a better postchallenge ADG than the controls. None of the Porcilis APP vaccinated pigs showed signs of depression postvaccination and none were euthanased after challenge with either serovar 1 or 15. The pigs vaccinated with the live vaccine showed obvious depression after each vaccination and a total of 3 pigs were euthanased after challenge (one with serovar 1 and two with serovar 15). CONCLUSIONS: Both of the vaccines provided significant protection against a severe challenge with serovar 1 A pleuropneumoniae. Neither vaccine was effective against a serovar 15 A pleuropneumoniae challenge. There was evidence that the Porcilis APP vaccine did provide some protection against the serovar 15 challenge because the ADG, after challenge of pigs given this vaccine, was greater than the control pigs. PMID- 15272464 TI - [Malaria in Italy in 1902. Epidemiological and preventive research]. PMID- 15272465 TI - Sarcocystis neurona (Protozoa: Apicomplexa): description of oocysts, sporocysts, sporozoites, excystation, and early development. AB - Equine protozoal myeloencephalitis is a major cause of neurological disease in horses from the Americas. Horses are considered accidental intermediate hosts. The structure of sporocysts of the causative agent, Sarcocystis neurona, has never been described. Sporocysts of S. neurona were obtained from the intestines of a laboratory-raised opossum fed skeletal muscles from a raccoon that had been fed sporocysts. Sporocysts were 11.3 by 8.2 microm and contained 4 sporozoites. The appearance of the sporocyst residuum was variable. The residuum of some sporocysts was composed of many dispersed granules, whereas some had granules mixed with larger globules. Excystation was by collapse of the sporocyst along plates. The sporocysts wall was composed of 3 layers: a thin electron-dense outer layer, a thin electron-lucent middle layer, and a thick electron-dense inner layer. The sporocyst wall was thickened at the junctions of the plates. Sporozoites were weakly motile and contained a centrally or posteriorly located nucleus. No retractile or crystalloid body was present, but lipidlike globules about 1 microm in diameter were usually present in the conoidal end of sporozoites. Sporozoites contained 2-4 electron-dense rhoptries and other organelles typical of coccidian zoites. Sporozoites entered host cells in culture and underwent schizogony within 3 days. PMID- 15272466 TI - [Fibroepithelial polyp of the ureter. Report of one case]. AB - OBJECTIVES: To report the rare case of a patient with a ureteral polyp. METHODS: We describe the case of a 55-year-old female patient receiving care at the Celia Sanchez Manduley University Hospital in Manzanillo, Cuba, who was fortuitously diagnosed of a fibroepithelial polyp of the right ureter during the work up and treatment of an ovarian tumor. RESULTS: This case is the first of its kind in this hospital after 22 years, which confirms the rarity of ureteral tumors, specifically those of benign etiology. The absence of symptoms, specifically hematuria and pain, does not correspond to the reviewed articles. The chosen treatment was exeresis of the polyp at its base and frozen biopsy, followed by re establishment of the urinary passage, as various authors recommend. Currently the endoscopical approach is recommended for its multiple advantages. CONCLUSIONS: We conclude that this disease is very rare, may have a symptomatic course and the treatment of choice is surgery with very good results. PMID- 15272467 TI - [The advantages of axillary artery cannulation in surgical treatment of type A acute aortic dissection]. AB - There are more alternative cannulation techniques during surgery of type A aortic dissection. The most frequently used femoro-atrial cannulation method provides limited possibility for brain protection during surgery. This theory is confirmed by relatively high frequency of major brain complications in patients operated on while using this cannulation technique. During the last years cannulationis used more often, as it may provide more protection for the brain than other methods. In 2003 seven patients underwent aortic reconstruction because of type A acute aortic dissection using axillary cannulation. All patients except one were discharged after uneventful recovery. There were no postoperative neurological complications following surgery. We lost one patient due to distal progression of the dissection. He was the only patient with clinical evidence of transient postoperative brain damage. We are strongly convinced that the spectacular improvement in our results for the surgery of type A acute aortic dissection is due to the axillary cannulation, the anterograde flow and the isolated cerebral perfusion. We recommend the axillary cannulation technique as the first choice in type A acute aortic dissection. PMID- 15272468 TI - Researches on the etiology of sleeping sickness. 1903. PMID- 15272469 TI - Glycine encephalopathy (nonketotic hyperglycinaemia) : review and update. AB - This article summarizes data and issues covered in the workshop on Glycine Encephalopathy using headings that cover important topics in our present knowledge of this disease. PMID- 15272470 TI - Congenital disorders of glycosylation (CDG): update and new developments. AB - After a brief overview on CDG, this workshop concentrated on the experience with (mostly) known CDG in a European country (the Czech Republic) and on the Australasian experience, on recent developments regarding congenital muscular dystrophies due to O-mannoslyglycan assembly defects, and on new presentations of CDG. It was concluded that we are still at the beginning of 'explosive' research on CDG and that we need to apply new and known technologies to the diagnosis, understanding of pathophysiology, and treatment of CDG. PMID- 15272471 TI - Two genomes are better than one: widespread paleopolyploidy in plants and evolutionary effects. PMID- 15272472 TI - HIV treatment to be tested in patients. PMID- 15272473 TI - FDA issues new rules on donor tissue eligibility. PMID- 15272474 TI - The GC-MS detection and characterization of reticuline as a marker of opium use. AB - Reticuline (a precursor of opium alkaloids) was detected and characterised as its trimethylsilyl ethers, acetyl esters and methyl ethers by GC-EIMS and GC-CIMS in opium and the urine of opium users after hydrolysis by acid or beta-glucuronidase as coextractive of morphine. Because this compound cannot be detected in heroin and poppy seeds, it is suggested as a differentiating marker between opium and heroin use, opium and poppy seeds use, or opium and "pharmaceutical" codeine use in cases when opiate use has been confirmed by detection of morphine and codeine in the urine. As well as being a constituent of opium, reticuline in the urine of opium users may also result from the metabolic demethylation of the three other benzyltetrahydroisoquinoline opium alkaloids: codamine, laudanosine and laudanine. PMID- 15272475 TI - Biological underpinnings of treatment resistance in schizophrenia: an hypothesis. PMID- 15272476 TI - Prisoner or patient? The official debate on the criminal lunatic in nineteenth century Ireland. AB - Nineteenth-century Ireland was colonized and strictly controlled from Britain. In this highly regulated society, reports of the Inspectorate of Lunacy in Ireland were used to express an official medical view on cirminal lunacy. This view was based on experiences gained in the Central Criminal Lunatic Asylum for Ireland, opened at Dundrum in 1820. This paper will examine some of the ideas expressed in these reports, including views on the treatment of criminal lunatics, on their potential for dangerous behaviour, and on emigration as a form of after-care. PMID- 15272477 TI - A single-LTR HIV-1 vector optimized for functional genomics applications. AB - The development of high-throughput methods of converting simple expression cassettes into lentiviral vectors and expediting the process of retrieving vector genomes that carry candidate genes from host DNA will facilitate the use of lentiviral vectors as an efficient means of screening novel gene function. To optimize lentiviral vectors for functional genomic applications we have developed a shuttle HIV-1 vector containing a single LTR. Incorporation of a LoxP site and the Sbfl restriction enzyme site into the vector LTR allowed for the rescue of integrated vector genomes into individual bacterial clones. Vector DNA isolated from bacteria was used for a second round of functional screening. Furthermore, we identified a continuous DNA sequence containing all the cis elements required for vector production. Incorporating the isolated sequence into expression cassettes resulted in the generation of HIV-1 vectors in a single cloning step, which imparts a simplified procedure for converting cDNA expression cassettes into single-LTR lentiviral vectors. PMID- 15272478 TI - Hypnosis in Sweden during the twentieth century - the life and work of John Bjorkhem. AB - In this paper we present a study of hypnosis in Swedish medicine during the twentieth century. We focus on the life and work of the hypnotist and Swedish physician John Bjorkhem. He was a controversial person and his therapeutic ideas were not accepted in the medical establishment. The biography of Bjorkhem and the history of hypnosis in Swedish medicine during the twentieth century are intimately related to each other. We describe an outsider in medicine, both with respect to person and topic. In the case of Bjorkhem the situation was especially complicated because of his multidisciplinary activities, with three different faculties involved. Finally we deal with the condition of a pioneer, and his struggle for acceptance and understanding. PMID- 15272479 TI - Transduction of cell lines and primary cells by FIV-packaged HIV vectors. AB - Human immunodeficiency virus type 1 (HIV-1), simian immunodeficiency virus, and feline immunodeficiency virus (FIV) are capable of packaging viral RNA derived from heterologous as well as homologous lentiviruses, a phenomenon referred to as "cross packaging." To remove the possibility of seroconversion to HIV proteins, and to avoid potential problems arising due to targeting of vector or packaging construct by antiviral genes, we investigated the feasibility of using an FIV based packaging system to deliver human immunodeficiency virus type 2 (HIV-2) based vectors bearing anti-HIV-1 RNA expression cassettes to target cells. In the absence of FIV rev, FIV was packaged by HIV-2 at only 3% the efficiency of FIV packaging by FIV, but this was increased to 39% of homologous controls by supplying FIV rev in trans. HIV-2 vectors were packaged by FIV at levels equal to or exceeding the homologous HIV-2 packaging system in the absence of HIV-1 tat and rev, and levels increased approximately four- to fivefold with the addition of tat and rev in trans. HIV-2 vectors bearing a polyribozyme cassette targeting multiple regions of HIV RNA were efficiently packaged by FIV and transferred to target cells. Upon challenge with cell-free HIV-1 (m.o.i. = 0.1) a significant reduction in replication was observed. These findings demonstrate that packaging HIV vectors with FIV is a viable alternative, which avoids use of HIV structural proteins. PMID- 15272480 TI - A new vector, based on the PolII promoter of the U1 snRNA gene, for the expression of siRNAs in mammalian cells. AB - Several vectors for the induction of RNA interference in mammalian cells have been described,based mainly on polIII-dependent promoters. They transcribe short hairpin RNAs (shRNA) that,after being processed into short interfering RNAs (siRNAs), mediate the degradation of the target mRNA. Here, we describe the construction of a new siRNA-expressing vector (psiUx) based on the strong and ubiquitous polII-dependent promoter of the human U1 small nuclear RNA (snRNA)gene. In psiUx, the only constraint for the shRNA sequence is a purine at position +1, since specific 3'-end formation is achieved by a box element located downstream of the transcribed region. Several constructs were designed against the lamin A/C target. Depending on the structure of the shRNA transcribed, a preferential or exclusive accumulation of the antisense strand is obtained, thus avoiding possible nonspecific targeting by the sense strand. In all cases tested, very effective siRNAs were produced, thus providing a proof-of-principle that a snRNA-type polII promoter can be used for the expression of siRNAs. We show that psiUx ensures high levels of expression and efficient knock down of the target gene also in stable cell lines. PMID- 15272481 TI - Requirements for the group health insurance market; non-federal governmental plans exempt from HIPAA Title I requirements. Final rule. AB - This rule finalizes existing exemption election requirements that apply to self funded non-Federal governmental plans. In it, we clarify the conditions under which plan sponsors may exempt these plans from most of the requirements of title XXVII of the PHS Act, and provide guidance on the procedures, limitations, and documentation associated with exemption elections. Finally, we revise the requirements to reinforce beneficiary protections for exemption elections. PMID- 15272483 TI - [Review of: Healy, David. The Creation of psychopharmacology. Cambridge, MA: Harvard University Press, 2002]. PMID- 15272482 TI - Manic-depressive illness in children: an early twentieth-century view by Theodor Ziehen (1862-1950). Introduction. AB - The German psychiatrist and philosopher Theodor Ziehen (1862-1950), little known in psychiatry today, wrote on of the first systematic treatises on child psychiatry in the early years of the twentieth century. This report provides the first English translation of Ziehen's chapters on major mood disorders of children and adolescents, with detailed biographical notes and commentary on his contributions to the foundation of child psychiatry and to characterizing manic despressive illness in this age group. PMID- 15272484 TI - [Developments in civil and disciplinary law. A view of the revision of the Netherlands Society for Dentistry (NMT) administration of justice]. AB - According to some people, the renewed Dutch civil disciplinary law is not fulfilling the expectations. Some procedures seem to be reductant for accused dentists. Besides the civil disciplinary law, also the professional disciplinary law of Dutch Dental Association is questionable. The Association is planning to change the current professional disciplinary law. However, the proposed amendments may have the consequence that a Dutch dentist more frequently will be faced with a civil disciplinary law procedure or normal civil law procedure. PMID- 15272485 TI - Karl Jaspers' general psychopathology: the history of the English translation. PMID- 15272486 TI - Mammogram underuse concerns researchers. PMID- 15272487 TI - Hospital-based falls are surprisingly common in younger patients. PMID- 15272488 TI - Standards for high-risk surgeries would save lives, study says. PMID- 15272489 TI - Study reveals wide variations in guideline-based heart failure care. PMID- 15272491 TI - G-protein-coupled receptors: the devil is the detail. PMID- 15272490 TI - Evolutionary genetics: Ambiguous role of CCR5 in Y. pestis infection. AB - Mecsas and colleagues suggest that a deficiency in the chemokine receptor CCR5 in humans is unlikely to confer protection against plague, based on their study of Yersinia pestis infection in Ccr5-deficient mice. They were testing the hypothesis that a mutation in the CCR5 gene, frequently found in Caucasians, may have been selected for in the past because it provided protection against (bubonic) plague; the mutation, called CCR5Delta32, is characterized by a 32-base pair deletion. We have also tested this hypothesis by using Y. pestis infection in mice and, in addition, we have done phagocytosis experiments with macrophages from wild-type and Ccr5-deficient mice. Although, like Mecsas et al., we did not see any difference in the survival of the two groups of mice, we did find that there was a significantly reduced uptake of Y. pestis by Ccr5-deficient macrophages in vitro. Our results indicate that the role of Ccr5 in Y. pestis infection may therefore be more complex than previously thought. PMID- 15272492 TI - How academia can help drug discovery. PMID- 15272493 TI - Merck's statin first to receive over-the-counter status. PMID- 15272494 TI - Lessons learnt from Genasense's failure. PMID- 15272495 TI - Establishing novelty and relevant prior art. PMID- 15272496 TI - An audience with... Robert S. Langer [interviewed by Kenneth J. Germeshausen]. PMID- 15272497 TI - Targeted lung cancer therapies. PMID- 15272498 TI - Cetuximab. PMID- 15272499 TI - The state of GPCR research in 2004. PMID- 15272500 TI - The genomics evolution. PMID- 15272501 TI - The paradox of the insulin/IGF-1 signaling pathway in longevity. AB - Ageing may be controlled by a genetic-hormonal system that may have originated from a very early common ancestor. One of the pathways that has been implicated in ageing is the insulin/insulin-like growth factor (IGF-1) signaling, which is involved in many functions that are necessary for metabolism, growth, and fertility in animal models like flies, nematodes, and mammalians. While disruption of the insulin/IGF-1 receptor in nematodes and flies increases lifespan significantly, mammals with genetic or acquired defects in insulin signaling pathway are at risk for age-related diseases and increased mortality. This contradiction can be explained by the acquisition of more complicated metabolic pathways in mammalians over evolution. Mammals have insulin/IGF-1 receptors in many organs, but their functions are opposite if they are located in the central nervous system or in the periphery; whereas lower species have insulin/IGF-1 receptors signaling mainly through the nervous system. Furthermore, mammalians have different and very specific receptors for insulin and IGF-1, with distinct pathways and diverse functions. Striking evidence suggests that decreased IGF-1 levels and signaling during early development, but not the insulin signaling may modulate longevity in many species. Thus, paradoxical outcomes follow the decrease of insulin and/or IGF-1 signal pathway in invertebrates and in mammals, prolonging life in the former and shortening it in the latter. In this review we focus on the downstream cascade of events in the insulin and IGF-1 signaling to identify specific pathways that are relevant to human longevity. PMID- 15272502 TI - Long-term neurological complications of traumatic brain injuries. PMID- 15272503 TI - The new amnesty. PMID- 15272504 TI - DNA damage and aging. AB - The hypothesis discussed here is that a major component of aging in metazoans is oxidative damage to nuclear DNA. Such a viewpoint would be consistent with the fact that all of the thus far identified premature aging syndromes in mammals involve mutations in nuclear proteins. Several of these nuclear proteins are enzymes that are related to DNA metabolism or DNA repair. Among the single- and double-stranded DNA damage repair pathways present in eukaryotes, only one pathway often fails to restore the full information content of the genome and typically would result in a deletion of a few base pairs. This pathway is called nonhomologous DNA end joining (NHEJ) and it is a major pathway for the repair of double-strand DNA breaks. Repetitive DNA content may determine the extent to which any organism can use this pathway, and therefore, may dictate a key factor in the balance between oxidation and organismal lifespan. PMID- 15272505 TI - Many uninsured children qualify for Medi-Cal or Health Families. AB - Nearly 1.5 million California children ages 0-17 did not have health insurance coverage for all or part of the year in 2002. However, almost two-thirds of these uninsured children were eligible for one of the state's two public health insurance programs--Medi-Cal or Healthy Families. Uninsured children who were eligible but not enrolled in these programs were spread throughout the state, with wide variations between local areas. This policy brief presents data on children ages 0-17 in California who did not have health insurance coverage for some or all of the year and who were eligible for the Medi-Cal or Healthy Families programs. These data highlight the geographic variations in children's uninsured eligibility rates for state Assembly, Senate and Congressional districts, as well as for counties and the Los Angeles Service Planning Areas (SPAs). Uninsured-eligibility rates at local levels were estimated by applying a small-area methodology to multiple data sources, including the 2001 California Health Interview Survey (CHIS 2001), 2000-2002 Current Population Surveys, and the 2000 Census. PMID- 15272506 TI - [Hip prosthesis implantation--an interdisciplinary clinical pathway]. PMID- 15272507 TI - [Guideline of Cochlear implantation and development of otology]. PMID- 15272508 TI - Charting a future course for the MCH Journal. PMID- 15272509 TI - Alginate impressions for fixed prosthodontics. A 20 year follow up study. AB - The aim of this study was to estimate whether the survival ratios after 20 years of fixed prosthodontics made of alginate impressions was higher, equivalent or lower, compared to the survival ratios, shown in studies, where different impression materials were used. 151 females and 104 males were screened regarding the condition and age of the restorations at the annual check-up in one of the author's surgeries. Average ages were 55 and 54 years respectively, when the fixed prosthodontics were seated. A total of 1.271 units were produced during the twenty years, 911 abutment teeth and 360 pontics. The type of prosthetic work was divided into three groups: 1) larger fixed prosthodontics 6-14 units (469), 2) smaller fixed prosthodontics 2-5 units (541) and 3) single crowns (261). The results show that alginate impressions can produce fixed prosthodontics with survival ratios similar to those presented in other studies, after 5 years (99%), 1o years (93-96%) and 15 years (74-96%). After 20 years the survival ratio was 61 63%. In conclusion, fixed prosthodontics made according to the syringe-tray alginate impression method may have the same success rates after 20 years compared to that of fixed prosthodontics presented in previous longitudinal clinical studies where other impression materials had been used. In this study, caries and root fractures were the main reasons for removing abutment teeth and pontics. PMID- 15272510 TI - Dental treatment of the primary dentition in 7-12 year-old Swedish children in relation to caries experience at 6 years of age. AB - In most Swedish counties, epidemiological data on the permanent dentition are collected on patients between 7 and 19 years of age. However, for the primary dentition, epidemiological data are only available for the 3-6-year-old age groups. As far as we know, no studies have investigated the relation between caries prevalence in the early primary dentition and caries prevalence or treatment performed in the late primary dentition. Thus, the purpose of the present study was to investigate the relationship between dental treatment in the primary dentition of Swedish children 7-12 years of age and caries experience in the primary dentition at 6 years of age. This retrospective study is based on record data from a randomly selected sample comprising 10% of all children born in 1987 in Jonkoping County (n=433). Of these children, 381 had been treated regularly at the Public Dental Service clinics between 7 and 12 years of age and were included in the study. It was found that children with previous caries experience at 6 years of age received significantly more treatment during the studied period compared to children who were caries free at the same age. The children with caries experience required a mean of 3.5 times more treatments compared to caries-free children. The present study underlines the importance of early detection and prevention of caries in the primary dentition if optimal dental health is to be expected in the late primary dentition. PMID- 15272511 TI - The microbial outcome observed with polymerase chain reaction in subjects with recurrent periodontal disease following local treatment with 25% metronidazole gel. AB - The aim of this study was to evaluate the microbial outcome in patients with recurrent periodontal disease following treatment with 25% metronidazole gel using the polymerase chain reaction (PCR). Twenty subjects in a maintenance care program but with recurrent periodontal disease participated. Three months after scaling and root planing a total of 40 sites, 2 in each patient, with pocket probing depth of > or = 5 mm were selected. One site randomly selected was treated with 25% metronidazole gel (test) and the other site with a placebo gel (control). A bacterial sample was collected on paperpoint from each test and control site at baseline and 12 weeks after treatment. The following pathogens were analysed and detected with PCR:Actinobacillus actinomycetemcomitans (A.a.), Porphyromonas gingivalis (P.g.) and Prevotella nigrescens (P.n.). At baseline, A.a., P.g. and P.n. were detected in 30, 60 and 70% of all test sites and in 32, 58 and 21% of all control sites. There was a statistically significant difference between the test and control sites for P.n. at baseline. The major difference after treatment with 25% metronidazole gel was the increase of positive control sites for P.g. and P.n. However, there were no statistically significant differences in the occurrence rate of A.a., P.g. and P.n. at test and control sites after treatment. This study has shown that 25% metronidazole gel treatment did not seem to influence the microbial outcome, when PCR was used to analyse the presence/absence of A.a., P.g. and P.n. in this group of subjects with recurrent periodontal disease. PMID- 15272512 TI - Image quality of digital and film radiographs in applications sent to the Dental Insurance Office in Sweden for treatment approval. AB - In July 2002, a new dental insurance program was introduced in Sweden. For all patients over 65 years, prior approval for all prosthetic work would need to be obtained from the Dental Insurance Office. From October to December 2002, 540 cases were randomly selected for evaluation from the 14,624 applications that had been sent from throughout Sweden to the Dental Insurance Office in Lund. Our aims were to appraise the quality of the radiographic examinations and to compare the quality of the digital with the film (X-ray film) radiographs. The radiographic examinations were evaluated as a whole in relation to the proposed treatment and in detail using specific criteria such as density, contrast, unsharpness, angulation, and receptor position error. The quality variables were evaluated as acceptable or unacceptable. A total of 4,687 intra-oral and 206 panoramic radiographs were evaluated. Thirteen per cent of the intra-oral radiographs and 9% of the panoramic radiographs were taken with a digital technique. Most of the digital radiographs--7o% of the intra-oral and 61% of the panoramic radiographs- were submitted on microdisk. Twenty-eight per cent of the intra-oral digital radiographs, however, were submitted on paper. The radiographic quality in 15o cases (28%) were found to be unacceptable for assessment of the proposed treatment. The most common error--both in digital and X-ray film radiographs--was in receptor position. Significantly more errors were found in the intraoral digital radiographs compared to the radiographs taken with X-ray film. Most of the errors in the digital radiographs were detected in the paper copies. In conclusion, it is possible to improve the radiographic quality in applications for treatment approval, and the dentists had more difficulties with the digital technique than with X-ray film. PMID- 15272513 TI - The efficacy of ropivacaine as a dental local anaesthetic. AB - The objectives was to investigate the efficacy of ropivacaine in dentistry. This open-labelled, parallel-group study included 41 subjects randomised to 1 ml or 2 ml ropivacaine 7.5 mg/ml and to maxillary infiltration or nerve block of the inferior alveolar nerve. Pinprick pain, electrical pulpal testing, and numbness were used as efficacy measures. Following onset, the duration was measured until baseline status was re-established or after a maximum of 6 h. A high frequency of anaesthesia was obtained. The pulpal anaesthesia mean onset time was 2.1 and 1.6 min after end of infiltration and 2.9 min and 4.5 min following end of nerve block injection for the 1 and 2 ml ropivacaine respectively. Pulpal anaesthesia mean duration was 0.4 and 1.3 h after infiltration and 3.7 and 4.3 h for nerve block respectively. The mean lip numbness duration ranged from 3.7 to 5.1 h for the upper lip and from 7.5 to 8.4 h of the lower lip. In conclusion 1 and 2 ml ropivacaine 7.5 mg/ml provided a high frequency of anaesthesia with short onset time and long duration following nerve block. Ropivacaine may be suitable for time-consuming oral procedures and/or when prolonged postoperative analgesia is desirable. PMID- 15272514 TI - Factors affecting the duration of orthodontic treatment in children. A retrospective study. AB - Factors that affect the duration of orthodontic treatment in children were studied from treatment records, dental and cephalic radiographs, and dental plaster casts of a random sample of 93 orthodontic patients who were treated during the period 1983-1994 and were aged 7 to 13 years at the start of treatment. The duration of treatment and 15 variables describing the patients' age at the start of treatment, the gender distribution of the patients, the progress of treatment, the occlusal status, skeletal deviations, malocclusion classes (Angle), and patients' age at the start of treatment, were recorded. The data were analysed by means of analysis of variance, chi-square test, and multiple regression analysis. The mean treatment time was 2.9 years. The regression analysis revealed that the variations in sex variables, malocclusion class, patients' age at the start of treatment, type of appliances used, number of appliances used, number of missed appointments, and main additional diagnosis explained about 41% of variation in treatment time. Treatment of Class I and Class II/1 patients with combinations of fixed and removable appliances, early start, high number of appliances used, and high number of missed appointments prolonged the duration of treatment. Treatment of Class I patients with frontal crossbite was relatively short. PMID- 15272515 TI - Age dependence of compliance with orthodontic treatment in children with large overjet. An interview study. AB - Large overjet has been associated with an increased risk of trauma of the permanent maxillary incisors, especially before 10 years of age. The prevalence of an overjet of more than 6 mm in 10-year-old Swedish children is about 15%. To prevent trauma by reducing the overjet, this treatment be set in early in life. However, compliance with orthodontic treatment is a significant and well known problem and may be associated with the age of the patient. This study concerns children treated with an open activator with built in headgear (HG activator). The aim of treatment was to reduce the trauma risk in these patients as early as possible, and an objective of the study was to gain better knowledge of young children's (6-13 years of age) motivation for and response to correction of large overjet. In-depth interviews focusing on motivation were held with 18 children. Interviews were transcribed verbatim and analysed according to Grounded Theory, a qualitative method. The results indicate the importance of parental involvement for younger children's compliance. Older children seem to have a higher degree of internal motivation for treatment and less need for parental support for compliance with treatment. If treatment compliance can not be ensured through parents' wholehearted involvement and control it seems to be better to delay treatment until the child is older. PMID- 15272516 TI - [Arrhythmias of primary hypertrophic cardiomyopathy]. AB - Primary hypertrophic cardiomyopathy is a genetic disease causing sarcomere dysfunction. The structural and functional myocardial changes combine to produce cardiac arrhythmias related to reentry phenomena and to triggered automatic activity. The commonest arrhythmias are atrial fibrillation and ventricular arrhythmias; junctional tachycardias via the bundle of Kent are rare. Atrial fibrillation and the Wolff-Parkinson-White syndrome are more commonly associated with certain genetic mutations. Their treatment is mainly based on medication with amiodarone or on radiofrequency ablation in cases of junctional tachycardia. Ventricular arrhythmias are mainly isolated ventricular extrasystoles and non sustained ventricular tachycardia. The prognostic significance of the latter has been subject of debate for many years but recent studies report a poor prognosis with non-sustained ventricular tachycardia especially in the young patients. Sustained ventricular tachycardia and ventricular fibrillation, though life threatening complications of hypertrophic cardiomyopathy, are rarely documented and justify implantation of an automatic defibrillator as the risk of recurrence is high. The main objective of the cardiologist in cases of primary hypertrophic cardiomyopathy is to identify the patient at high risk of sudden death. This requires analysis of several parameters: clinical, anatomical, haemodynamic, rhythmic, functional and genetic. The presence of at least two risk factors for sudden death justifies preventive measures. The implantation of an automatic defibrillator is the most reliable form of treatment. PMID- 15272517 TI - [Concealed phenomena in electrocardiography]. AB - Concealed phenomena in electrocardiography are events which, having no direct effect on the electrocardiogramme, nevertheless change the following sequences giving rise to very unusual appearances. They are diagnosed by deduction. First of all, there is concealed anterograde or retrograde conduction in the AV node, the consequences of which are either a block or the suppression of an escape rhythm or even facilitation of the transmission of activation. Concealed conduction is also possible in the Bundle of His and its branches, explaining the frequency of dependent blocks and aberrant conduction. There is also concealed conduction in accessory pathways. The second form is concealed rhythms: hisian extrasystoles giving rise to pseudo A-V block, and concealed extrasystoles and parasystoles. With temporary pacing of tachycardia, it has finally been possible to describe "visible" concealed phenomena, not on the surface but on the endocavitary electrogramme. PMID- 15272518 TI - [Bidirectional ventricular tachycardias]. AB - Bidirectional tachycardias are rare arrhythmias. Nevertheless in the sixties and seventies these arrhythmias prompted much work relating to their mechanism. Discussions about the supposed supra-ventricular origin of certain bidirectional tachycardias essentially rested on presumptive arguments based on electrocardiographic analysis. All the electrophysiological investigations which could be performed in tachycardia showed a ventricular origin. The current hypotheses concerning the electrophysiological mechanism favour non-unifocal mechanisms as well as a very diverse aetiology: an automatic focus, or the triggered activities being associated with alternating conduction, or re-entry between the left hemibranches. Although the classic context is of excess digitalis with advanced cardiopathy, readily in atrial fibrillation with a poor prognosis as a corollary, the most recent description of catecholergic ventricular tachycardias with the very characteristic appearance of bidirectional tachycardias justifies updating the understanding of these unusual tachycardias. PMID- 15272519 TI - [Anatomy of the atria for rhythmologists]. AB - The anatomy of the atria is always in the mind of interventional rhythmologists. There is a mental superposition of the anatomical structures and the references obtained by different incidences of fluoroscopy and the endocavitary electrocardiogram. But understanding the anatomy also requires a certain knowledge of dissection to determine, for example, the orientation of bundles of muscle fibres and anatomical sections. The sino-atrial node is situated at a distance from the endocardium. It is long and protected by its own artery which makes it difficult to reach. The atrio-ventricular node has multiple posterior expansions which correspond to the sites where radiofrequency ablation is effective. The cavo-tricuspid isthmus is the target zone for the treatment of atrial flutter but radiofrequency ablation which must be long may be applied at three different levels: inferolateral, median (the most common site) or inferoseptal. Finally, atrial fibrillation has incited many studies of the muscular extensions of the left atrium to the pulmonary veins, the morphological variations of these veins and the organisation of the muscle fibres of the left atrial wall. They have inspired new concepts of atrial fibrillation. PMID- 15272520 TI - [Arrhythmogenic effects of non cardiovascular drugs]. AB - Among the unwanted effects of drugs, arrhythmogenic effects are particularly fearsome. Although rare they are serious, and responsible for syncope or sudden death, often linked with torsades de pointe on established long QT. For non cardiovascular drugs, detection is difficult because patients do not undergo systematic cardiological surveillance. Nevertheless understanding the risk, identification of predisposing factors, and consideration of the contra indications are the rules of prescription, which are even more indispensable when the pathology being treated is benign. In effect, the implicated drugs are mainly anti-histamines, antibiotics, neuroleptics and antidepressants. The pharmaceutical companies, regulatory agencies, and pharmacological surveillance services must recognise the greatest possible risk of a drug, thanks to pre clinical data, experimental electrophysiology both in vivo (measurement of the QT interval) and in vitro (action potential duration) or even in the elementary channel (essentially analysis of the iKr current). Correlated with clinical data (QT changes, pharmacokinetic interactions), a risk/benefit profile can therefore be established, which is even more demanding when the pathology is benign or when alternative drugs are available. PMID- 15272521 TI - [Complications of permanent cardiac pacing]. AB - The implantation of a pacemaker is an everyday medical procedure. New indications are under evaluation. However, it should be recalled that this is a surgical intervention with implantation of a prosthesis with possible complications. This should, therefore, be a considered decision. There are early complications which occur in the first 6 weeks after implantation. Their incidence is underestimated (up to 7%) as is their seriousness. There are late complications. Some are responsible for pacemaker dysfunction, the risk of which is proportional to the dependence of the patient on permanent cardiac pacing. The migration of a pacing catheter or the fracture of an Accufix catheter expose the patient to much greater risk. Venous complications are overlooked as they are usually asymptomatic. The superior vena cava syndrome is, however, a serious complication of cardiac pacing. Two recent studies (MOST and DAVID) underline the deleterious haemodynamic effects of unnecessary right ventricular pacing. This right ventricular pacing may have a pro-arrhythmic effect on the ventricles and be responsible for sudden death. It may also cause atrial arrhythmia even if atrio ventricular synchronisation is preserved. Infectious complications are also under reported, partially because of the difficulty of diagnosis. They may be life threatening and require extraction of the implanted material. In conclusion, it is wrong to think that even if a patient does not benefit from his implanted device this cannot have deleterious consequences. Pacemakers should be adjusted especially to avoid inappropriate right ventricular stimulation. PMID- 15272522 TI - [Atypical forms of atrioventricular nodal reentrant tachycardia]. AB - The concept of functional dissociation of atrioventricular nodal conduction on an underlying structure with different nodal inputs is well established. In the common form of AVNRT, the circuit comprises the slow pathway for antegrade conduction and the fast pathway for retrograde conduction. However, the tachycardia circuit may be slightly or totally different as it is not based on discrete anatomical pathways but on functional pathways due to non-uniform anisotropy of the perinodal atrial tissue involved in the circuit. First, there are variations of the common form. The fast retrograde pathway may be posterior or left instead of being anteriorly-located. The so-called uncommon forms of AVNRT consist in slow-slow and fast-slow forms. The slow-slow form uses two different slow pathways as antegrade and retrograde limb of the circuit. These pathways may be posterior or left. The fast-slow form appears to be complex and heterogeneous. The retrograde slow pathway may be located posteriorly, anteriorly, in-between or sometimes left-sided. Whatever the circuit, targeting the antegrade or retrograde slow pathway remains the aim of ablation and the only therapeutic issue. However, a 1%-AV block rate has to be kept in mind. PMID- 15272523 TI - [Sudden death and semi-automatic defibrillation]. AB - While cardiac arrest in hospital poses few immediate management problems, this is not the case outside hospital. For this reason semi-automatic defibrillators are easy to handle devices designed to deliver an early electric shock in the context of usage by non-specialist people following minimum training. These devices have shown a clear improvement in survival compared to the exclusive use of a manual defibrillator by highly trained emergency services, especially in confined areas such as casinos or aircraft, or where a significant number of potential patients are concentrated, such as airports. It is now important to be able to improve public access to defibrillation by various means currently being studied, and probably by relaxing the rules which allow the use of these devices. PMID- 15272524 TI - Assessing the prognosis of gastrointestinal stromal tumors: a growing role for molecular testing. PMID- 15272525 TI - The diagnosis of bronchioloalveolar carcinoma by cytologic means. PMID- 15272526 TI - Lowering the detection limits of HIV-1 viral load using real-time immuno-PCR for HIV-1 p24 antigen. AB - Presently, the assay that attains maximal sensitivity and dynamic range of HIV-1 viral copy number (50 copies per milliliter) is nucleic acid amplification of HIV RNA in plasma. Enzyme-linked immunosorbent assay (ELISA) methods for quantification of HIV-1 p24 antigen have been relatively insensitive. In this report, we show data that indicate real-time immuno-polymerase chain reaction (IPCR), a combination of the ELISA and PCR techniques, is more sensitive for HIV 1 p24 antigen detection than other currently reported methods. When derived from an IPCR standard curve, a dose response was observed from patient samples with known viral loads diluted within a 3-log range (1.68-6,514 viral RNA copies per milliliter). IPCR detected 42% (22/52) of patient samples that had fewer than 50 viral RNA copies per milliliter by reverse transcriptase-PCR. IPCR shows the potential to become the most analytically sensitive test available for determination of HIV-1 viral load by the detection of HIV-1 p24 antigen. PMID- 15272527 TI - Point prevalence of Cryptosporidium, Cyclospora, and Isospora infections in patients being evaluated for diarrhea. AB - From March to September 2001, 315 specimens from "nonrepeat" patients that were submitted for ova and parasite examination were stained using the Kinyoun modified acid-fast stain to detect the intestinal coccidians. Four patients (1.3%) were infected with coccidians, 2 with Cryptosporidium parvum and 2 with Cyclospora cayetanensis. No infections with Isospora belli were detected. In comparison, 15 patients (4.8%) had infections with one or more intestinal parasites detected by routine trichrome staining: 5 had Giardia lamblia; 2, Dientamoeba fragilis; 3, Strongyloides stercoralis; 1, Iodamoeba butschlii; 3, Endolimax nana; 6, Blastocystis hominis; and 1, Entamoeba coli. Four patients were multiply infected. Coccidians made up 29% of the clinically significant parasitic infections. The coccidians were missed in all 4 cases because no special staining was ordered. Clinicians need to be reminded that additional tests should be ordered to fully evaluate patients with chronic diarrhea in which no diagnosis is found by routine testing. PMID- 15272528 TI - Role of p16/INK4a in gastrointestinal stromal tumor progression. AB - Because the p16 locus is involved consistently in chromosomal losses found in malignant gastrointestinal stromal tumors (GISTs), we studied p16 in a series of 21 GISTs with complete follow-up using immunohistochemical analysis, semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR) and methylation-specific PCR (MSP). A fraction of cells of more than 20% with low or absent p16 immunostaining was detected in 12 GISTs, including all showing malignancy. RT-PCR revealed decreased p16 transcription in all except 2 p16 protein-deficient GISTs. By MSP, 7 cases showed p16 promoter methylation (all hypoexpressing p16; 6 malignant). A fraction of p16-deficient cells of more than 20% was associated with clinical malignancy (P = .003; log-rank test). The percentage of cells underexpressing p16, size, cellularity, mitotic count, and coagulative necrosis were associated with malignancy by Cox proportional hazards univariate analysis; only the former factor was selected by multivariate analysis (P = .039). Thus, p16 down-regulation, partly due to p16 promoter methylation, is implied in GIST progression. Furthermore, p16 immunohistochemical assessment seems a promising method for GIST prognostication. PMID- 15272529 TI - Cytopathologic diagnosis of bronchioloalveolar carcinoma: does it correlate with the 1999 World Health Organization definition? AB - We identified 29 bronchial washing, bronchoalveolar lavage, sputum, and fine needle aspiration specimens with corresponding surgical pathology specimens with features of bronchioloalveolar carcinoma (BAC). Surgical pathology correlates were reclassified according to the 1999 World Health Organization classification into pure BAC, mixed adenocarcinoma-BAC (AD-BAC), and papillary adenocarcinoma (PAP-AD). Twelve cases of invasive pulmonary adenocarcinoma (INV-AD) without a bronchioloalveolar component were reviewed for comparison. The cytology slides were evaluated for 12 features of BAC. No statistically significant feature permitted separation of BAC from AD-BAC or from PAP-AD. However, comparison of BAC with INV-AD identified 9 statistically significant cytologic features: clean background, absence of 3-dimensional clusters, neoplastic cells in flat sheets, orderly arrangement of cells with round uniform nuclei, predominance of mucinous cells, absence of nuclear overlap, absence of irregular nuclear membranes, fine granular chromatin, and nuclear grooves that were features of BAC cases. Although cytologic evaluation cannot prospectively diagnose BAC, the bronchioloalveolar pattern may be recognized and suggests in situ proliferation that is present in BAC, AD-BAC, or PAP-AD. The bronchioloalveolar pattern must be correlated with clinical, radiographic, and histologic parameters to determine whether the tumor is localized, multifocal, or diffuse and whether there is parenchymal invasion. PMID- 15272530 TI - The use of cell line standards to reduce HER-2/neu assay variation in multiple European cancer centers and the potential of automated image analysis to provide for more accurate cut points for predicting clinical response to trastuzumab. AB - Immunohistochemical analysis, the most efficient way of assaying for HER-2/neu overexpression in patients with invasive breast cancer, is subject to variation in sensitivity and evaluation when used in multiple laboratories. Cell lines with differing but constant levels of HER-2/neu expression have been advocated as standard material against which assay sensitivity can be gauged. Automated image analysis could provide more precise linear measurements of HER-2/neu expression than the subjective and categorical scoring system originally designed for the HER-2/neu clinical trials. Multiple European laboratories (range, 92-126) stained 7 cell line standards on 6 successive occasions using a variety of immunohistochemical assays for HER-2/neu. During the 2-year study period, a trend toward a standard sensitivity level was observed, with significant improvement in numbers of laboratories achieving appropriate results. Image analysis gave reproducible and significantly different linear values for a total of 621 HER 2/neu results on cell lines previously categorized manually as 3+, 2+, 1+, or 0. The use of cell line standards and image analysis have the potential to assist in standardizing immunohistochemical results for predictive markers and provide more accurate and quantifiable cut points for predicting clinical response to therapy, respectively. PMID- 15272531 TI - Differential expression of MUC1, MUC2, and MUC5AC in carcinomas of various sites: an immunohistochemical study. AB - We studied immunohistochemical expression of MUC1, MUC2, and MUC5AC in 194 carcinomas of different primary sites to determine whether differential expression patterns could be used to distinguish different carcinomas. MUC1 was expressed by most (except adrenocortical and hepatocellular carcinomas). MUC2 was expressed infrequently (positive immunoreactivity primarily in tumors of gastrointestinal origin). MUC5AC was expressed by most pancreatic ductal and endocervical adenocarcinomas and a variable number of tumors of the gastrointestinal tract. A MUC1+/MUC2-/MUC5AC- immunophenotype was observed in most breast, lung, kidney, bladder, endometrial, and ovarian carcinomas; MUC1+/MUC2-/MUC5AC+ was characteristic of pancreatic ductal adenocarcinomas and cholangiocarcinomas. Adrenocortical and hepatocellular carcinomas were negative for all mucins. Carcinomas of gastrointestinal origin exhibited variable expression of each mucin examined and no consistent immunoreactivity pattern. Many carcinomas can exhibit distinct MUC1, MUC2, and MUC5AC expression patterns, which might be valuable diagnostically in specific settings (eg, distinguishing cholangiocarcinoma from hepatocellular carcinoma or renal from adrenocortical carcinoma). However the overlapping and heterogeneous patterns of MUC1, MUC2, and MUC5AC expression observed in many tumors, particularly those of gastrointestinal origin, preclude use of these markers in the routine immunohistochemical assessment of carcinomas of an unknown primary site. PMID- 15272532 TI - Primary melanoma of the skin and cutaneous melanomatous metastases: comparative histologic features and immunophenotypes. AB - Through careful clinicopathologic correlation, we identified 37 metastatic melanomas in the skin, all of which had intraepidermal components. These were compared with 43 microscopically similar primary melanomas with a predetermined panel of immunostains in general use in surgical pathology, including bcl-2 protein, mutant p53 protein, Ki-67 (MIB-1), proliferating cell nuclear antigen (PCNA), alpha-isoform actin, and CD117 (c-kit protein). There was no significant difference in bcl-2 or alpha-isoform actin staining patterns of primary vs secondary cutaneous melanomas. The expression of Ki-67 generally was higher in metastatic melanomas than in primary lesions, and the same was true of mutant p53 protein labeling; however, some overlap was observed. CD117 staining was retained in 65% of metastatic melanomas (24/37) when they originated from ocular primary tumors; nevertheless, that marker was lost in virtually all of the other metastatic melanocytic neoplasms, whereas primary melanomas demonstrated consistent reactivity for c-kit protein. Although they are not definitive, these trends in immunoreactivity could facilitate the process of distinguishing the multiple primary melanoma syndrome from melanomatous metastases to the skin. That undertaking is best approached with circumspection, because clinicopathologic discriminators for this diagnostic separation are still imperfect. PMID- 15272533 TI - E-cadherin expression in patients with esophageal squamous cell carcinoma: promoter hypermethylation, Snail overexpression, and clinicopathologic implications. AB - Hypermethylation in the E-cadherin promoter region and expression of the transcription factor Snail were analyzed in 41 cases of esophageal squamous cell carcinoma (ESCC) and paired normal squamous epithelium by methylation-specific polymerase chain reaction (PCR) and reverse transcription-polymerase chain reaction (RT-PCR) to clarify the mechanism regulating E-cadherin deletion; 93 cases of ESCC were analyzed immunohistochemically to determine the clinicopathologic impact of E-cadherin deletion. Hypermethylation of the E cadherin promoter and Snail overexpression were detected in 25 cases (61%) by methylation-specific PCR and 34 cases (83%) by RT-PCR, respectively. Reduced E cadherin expression, observed immunohistochemically in 55 cases (59%), correlated with hypermethylation (P = .0011) but not Snail overexpression (P = .685). Hypermethylation and Snail overexpression correlated significantly with E cadherin deletion (P = .0018). Snail overexpression was unrelated to clinicopathologic factors. Reduced E-cadherin expression correlated with tumor invasion (P = .019) and vascular invasion (P = .052) but not other factors. E cadherin deletion had prognostic impact in univariate (P = .023) and multivariate (P = .034) analyses. E-cadherin deletion was regulated by hypermethylation and Snail expression. Examination of reduced E-cadherin expression is important for assessing biologic behavior, including clinical outcome, in patients with ESCC. PMID- 15272534 TI - Immunoreactivity of hepatocyte paraffin 1 monoclonal antibody in cutaneous metastatic tumors. AB - Hepatocyte paraffin 1 (Hep Par 1), a monoclonal antibody recognizing an antigen thought to be specific for hepatocyte mitochondria, is considered the most specific and sensitive marker of normal and neoplastic hepatocytes and has been used in diagnosing hepatocellular carcinomas. Recent data suggest that the specificity of Hep Par 1 for hepatic neoplasms is not absolute; nonhepatic neoplasms might express this marker. We assessed the value of Hep Par 1 in the diagnosis of cutaneous metastases of visceral tumors by immunostaining 65 biopsy or excision specimens with Hep Par 1. Hepatocarcinomas (primary and metastatic to the skin) showed specific (coarse granular intracytoplasmic) immunoreactivity. A similar reactivity pattern was found in 5 of 10 metastases of bronchial adenocarcinoma. Nonspecific (weak, nongranular) cytoplasmic immunoreactivity was observed in 5 cases of nonhepatic skin metastases. Hep Par 1 seems to be a sensitive marker of hepatocellular carcinomas metastatic to the skin, but its specificity is not absolute because it might be expressed in metastases of nonhepatic tumors, namely bronchial adenocarcinomas. Hep Par 1 should be used with caution in the investigation of cutaneous metastases from an unknown primary site, preferably in conjunction with other markers of neoplastic hepatocytes. PMID- 15272535 TI - Human papillomavirus typing with hybrid capture II on archived liquid-based cytologic specimens: is HPV typing always reproducible? AB - Reproducibility of human papillomavirus (HPV) typing on archived ThinPrep (Cytyc, Boxborough, MA) specimens was evaluated repeating Hybrid Capture II (HCII) (Digene, Gaithersburg, MD) testing after 25 to 40 months (mean, 31.3 months; group 1), 6 to 11 months (mean, 8.4 months; group 2), and 0 to 5 months (mean, 3.5 months; group 3). Another ThinPrep slide was prepared to evaluate cellularity and reproducibility of the cytologic diagnosis. The mean residual relative light units (RLU) calculated for each group showed a strong decrease of RLU values at the second typing (group 1, 21.69%; group 2, 26.47%; and group 3, 32.25% of original values). No residual HPV DNA was shown in group 1 in 8 (13%) of 60 cases or in groups 2 and 3 in 2 (3%) of 60 cases each. These cases were associated mostly with poor cellularity and reproducibility of the initial cytologic diagnosis in the final cytologic examination. Intergroup statistical analysis of mean relative percentages for cases with satisfactory residual cellularity revealed a significant difference only between groups 1 and 3 (P < .05). Although mostly reproducible, HPV typing results by HCII on archived specimens are influenced by material consumption. In addition, results might be affected by some DNA degradation after long-term sample storage. PMID- 15272536 TI - Patterns of colonic involvement at initial presentation in ulcerative colitis: a retrospective study of 46 newly diagnosed cases. AB - Studies have shown that rectal sparing and patchiness develop in treated and longstanding ulcerative colitis (UC), making the distinction from Crohn colitis increasingly difficult after treatment is initiated. However, no histologic studies of the incidence of rectal sparing in adults at UC onset have been performed. Colectomy specimens from 46 patients with classic UC histologic features and no Crohn disease features were identified. Biopsy specimens obtained before medical therapy were retrieved and examined blindly by 2 pathologists, along with appropriate control samples. Slides were scored for chronicity (crypt branching, subcryptal plasma cells, lamina propria plasma cells) and activity (cryptitis, crypt abscesses, epithelial injury). In 28 cases, only rectal biopsy specimens were taken; for 16, rectal and at least 1 proximal biopsy specimen were taken. All cases showed rectal involvement; none had rectal sparing at initial biopsy. Of 16 cases with rectal and more proximal biopsy specimens, 5 (31%) showed relative rectal sparing (lower scores in rectum than in more proximal sites). In 16 cases with rectal and more proximal biopsy specimens, chronicity and activity scores were higher in the rectum than in more proximal sites (P = .01; chronicity and activity). The mean overall chronicity score decreased in a linear manner from rectum to cecum. The rectum is involved and shows evidence of chronicity and activity at disease onset in UC, using colectomy as the gold standard for diagnosis. Because rectal sparing at UC onset has been reported, a prospective study using uniform biopsy protocols is needed to establish the true incidence of rectal sparing at presentation. PMID- 15272537 TI - Human achaete-scute homologue (hASH1) mRNA level as a diagnostic marker to distinguish esthesioneuroblastoma from poorly differentiated tumors arising in the sinonasal tract. AB - Distinction of high-grade esthesioneuroblastomas from other poorly differentiated tumors arising in the nasal cavity is an important diagnostic challenge because it determines patient management and prognosis. The human achaete-scute homologue (hASH1) gene is critical in olfactory neuronal differentiation and is expressed in immature olfactory cells; therefore, it could have potential use as a diagnostic marker The aim of the present study was to determine the value of hASH1 messenger RNA (mRNA) levels in differentiating esthesioneuroblastoma from other poorly differentiated tumors. A real-time polymerase chain reaction assay was developed, permitting the comparative determination of hASH1 mRNA levels in triplicate in a double-blind pilot study including 24 frozen cases of esthesioneuroblastoma and poorly differentiated tumors. All 4 positive cases were esthesioneuroblastomas, and all 19 poorly differentiated tumors were negative. In addition, there was an inverse association between the grade of esthesioneuroblastomas and hASH1 mRNA levels. The hASH1 mRNA level might represent a useful tool for distinguishing esthesioneuroblastoma from poorly differentiated tumors of the sinonasal region. PMID- 15272538 TI - Papillary lung carcinoma with prominent "morular" component. AB - Three cases of primary pulmonary papillary carcinomas with a prominent "morular" component involved 2 women and 1 man (age range, 25-68 years). The patients had symptoms related to the pulmonary mass, including chest pain, cough, and dyspnea. Radiographic evaluation of the thorax revealed the presence of a pulmonary mass. Surgical biopsies were obtained and reported as non-small cell carcinoma. All patients underwent lobectomy. Two tumors were located in the right upper lobe and 1 in the left upper lobe. The tumors were soft, white to tan, without evidence of necrosis or hemorrhage, and 2.5 to 3.5 cm in greatest diameter. The tumors were characterized predominantly by papillary architecture containing numerous "morules" composed of spindle cells without nuclear atypia or mitotic activity. Some morules were floating freely within papillary spaces; others seemed to detach from the papillary structures. Immunohistochemical studies of 2 tumors showed positivity for thyroid transcription factor-1, keratin, and carcinoembryonic antigen and negativity for thyroglobulin. The morules showed positive thyroid transcription factor-1 staining, weak keratin staining, and negative staining for smooth muscle actin, desmin, and HMB-45. These cases highlight an unusual phenomenon, that of primary papillary carcinomas of the lung with a prominent morular component. PMID- 15272539 TI - Detection and quantitation of HER-2 gene amplification and protein expression in breast carcinoma. AB - We compared fluorescence in situ hybridization (FISH), immunohistochemical analysis, immunocytochemical analysis, and relative quantification assays using polymerase chain reaction (PCR) as methods for estimating HER-2 gene amplification and protein overexpression infine-needle aspirate (FNA) specimens and paraffin-embedded tissue samples from 49 cases of breast cancer: FISH can be performed successfully on FNA smears. Immunohistochemical and immunocytochemical staining intensity of 3+ corresponds to a FISH ratio of more than 2.5. Immunohistochemical and immunocytochemical staining of 2+ and 1+ are not necessarily associated with gene amplification. Increased DNA PCR ratios might be seen without amplification, reflecting polysomy. HER-2 messenger RNA relative quantitation scores correlate well with HER-2 gene amplification. Owing to the ease with which it can be performed and interpreted, we conclude that FISH is the test of choice for HER-2 estimation and, when possible, should be performed on whole nuclei, which are readily available in FNA smears or imprint cytology. FISH may be used primarily or to confirm immunohistochemical, immunocytochemical, and PCR results. PMID- 15272540 TI - An unusual case of leukemic mantle cell lymphoma with a blastoid component showing loss of CD5 and aberrant expression of CD10. AB - Characteristically, mantle cell lymphoma (MCL) expresses surface immunoglobulin (sIg), CD19, CD20, and CD5 and lacks CD10 and CD23. Rare CD5-MCL variants have been described. This report describes a case of leukemic MCL with morphologically and immunophenotypically distinct classic MCL and blastoid-variant MCL (BV-MCL) components. The classic MCL had typical morphologic features and immunophenotype (kappa sIg light chain-restricted and CD5+; CD10- and CD23-). The BV-MCL had larger nuclei and open chromatin; these cells also were kappa sIg light chain restricted; however, they were CD10+ and CD5-. Fluorescence in situ hybridization studies demonstrated cyclin D1-immunoglobulin heavy chain gene fusion in both components; the bone marrow biopsy cellularity was replaced by CD10+ and cyclin D1+ and CD5-BV-MCL. This case illustrates the phenotypic heterogeneity of MCL and underscores the need for histopathologic correlation and, in some instances, ancillary genetic studies to accurately classify B-cell lymphomas. PMID- 15272541 TI - Identification of unsuspected PNH-type cells in flow cytometric immunophenotypic analysis of peripheral blood and bone marrow. AB - In this report, the flow cytometric expression patterns for CD14 on monocytic cells and CD16 on granulocytic cells in peripheral blood or bone marrow specimens are illustratedfor 15 patients proven to have a paroxysmal nocturnal hemoglobinuria (PNH) phenotype by flow cytometric analysis for CD55 and CD59. The varied clinical manifestations of PNH and its rarity may make it difficult to recognize clinically. As a result, blood or bone marrow samples may be submitted for flow cytometric analysis to exclude bone marrow neoplasia or dysplasia in patients with cytopenias rather than to exclude PNH. This was true in 5 of 15 study cases. Unlike CD55 and CD59, CD14 and/or CD16 are assessed routinely in the flow cytometric analysis of blood and bone marrow samples. Recognition of abnormal patterns of CD14 and CD16 expression might permit the identification of clinically unsuspected PNH by routine flow cytometric analysis. PMID- 15272542 TI - Flow cytometric detection of nonneoplastic antigenic polymorphisms of donor origin after allogeneic marrow transplant: a report of two cases. AB - Multidimensional flow cytometry found an abnormal lack of CD16 expression detected on neutrophils after hematopoietic stem cell transplantation (HSCT) in bone marrow specimens from 2 patients. Chimerism studies were used to determine that the CD16- myeloid cells were donor in origin, consistent with a normal polymorphic variant, most likely a previously described Fc gamma receptor IIIB gene deletion. Identical phenotypes were recognized in peripheral blood samples obtained subsequently from the respective donors. These donor-specific phenotypic abnormalities were transferred to patients who underwent successful HSCT. The distinction of these phenotypic abnormalities from neoplastic changes such as secondary myelodysplasias or relapse of original disease is important in posttransplantation monitoring of hematopoietic diseases. PMID- 15272543 TI - Enigmatic Kikuchi-Fujimoto disease: a comprehensive review. AB - To determine the clinicopathologic significance of Kikuchi-Fujimoto disease (KFD) and review the literature on this condition, we conducted a MEDLINE search of English-language articles published between 1972 and December 2003. KFD has a worldwide distribution, and Asiatic people have a higher prevalence. Its pathogenesis remains controversial. Patients are young and seek care because of acute tender, cervical lymphadenopathy and low-grade fever. Histologic findings include paracortical areas of coagulative necrosis with abundant karyorrhectic debris. Karyorrhectic foci consist of various types of histiocytes, plasmacytoid monocytes, immunoblasts, and small and large lymphocytes. There is an abundance of T cells with predominance of CD8+ over CD4+ T cells. Differential diagnosis includes lymphoma, lymphadenitis associated with systemic lupus erythematosus, and even adenocarcinoma. KFD is an uncommon, self-limited, and perhaps underdiagnosed process with an excellent prognosis. Accurate clinicopathologic recognition is crucial, particularly because KFD can be mistaken for malignant lymphoma. PMID- 15272544 TI - A molecular mechanism of formalin fixation and antigen retrieval. PMID- 15272545 TI - Fetal-maternal hemorrhage quantitation by flow cytometry. PMID- 15272546 TI - CT diagnosis of postoperative abdominal complications. PMID- 15272547 TI - Postoperative anatomic and pathologic findings at CT following colonic resection. AB - CT is frequently used for postoperative evaluation in patients who have undergone colonic resection. This pictorial article reviews and demonstrates the CT findings of normal postoperative anatomic changes, as well as different postoperative complications following various colonic operative techniques. PMID- 15272548 TI - Radiographic manifestations of normal postoperative anatomy and gastrointestinal complications of bariatric surgery, with emphasis on CT imaging findings. AB - Recently, there has been a tremendous increase in the frequency of utilization of surgery to control morbid obesity that is very common and increasing in incidence in Western industrialized nations. Imaging plays an important role in the evaluation and management of patients before and after bariatric surgery. In this article, we discuss the imaging findings relating to bariatric procedures, focusing on the role of computed tomography (CT) in the evaluation of normal postoperative anatomy and gastrointestinal complications. PMID- 15272549 TI - Pancreatoduodenectomy: imaging and image-guided interventional treatment. AB - Though the mortality of pancreatoduodenectomy (Whipple surgery) is under 4%, the morbidity continues to be high. The interventional radiologist plays an important role in the management of postoperative complications, such as abdominal abscess, bilomas, liver abscess, biliary obstruction, pseudocyst, and hemorrhage. Identification of the normal postoperative anatomy is crucial to correctly interpreting CT scans for short-term complications and long-term tumor recurrence. PMID- 15272550 TI - Postoperative radiology of endovascular abdominal aortic aneurysm repair. AB - This article addresses the imaging appearances following endovascular abdominal aortic aneurysm repair (EVAR). EVAR is gaining popularity and hence there is increasing likelihood that radiologists who are unfamiliar with the procedure will report imaging investigations on these patients. We describe the technique, failure modes, complications, and postoperative imaging features of this procedure. PMID- 15272551 TI - Ureteral injuries: CT diagnosis. AB - Ureteral injury is a rare, yet very serious, complication of various abdominal, pelvic, and even spinal procedures. It is often clinically unsuspected as symptoms are nonspecific and the patient may present weeks and even months after the injury. Therefore the diagnosis of ureteral injury is often delayed, leading to more serious morbidity. A ureteral injury may be first diagnosed on CT in a patient evaluated after surgery. A high index of suspicion is essential and a CT study should then include a delayed scan in order to establish the diagnosis of ureteral injury resulting in a urinoma. This may obviate the need for additional invasive imaging studies or unnecessary exploration. PMID- 15272552 TI - Postoperative pneumoperitoneum: prevalence, duration, and possible significance. AB - Free intraperitoneal air after abdominal surgery is a confounding finding with uncertain significance. A diagnostic dilemma often arises as to its origin: does it merely represent residual postoperative pneumoperitoneum (PP), which will need no intervention, or does it indicate a complication such as an anastomotic leak or a perforation of the gastrointestinal tract. Residual PP is usually well tolerated, as it will be absorbed over time and requires no therapy. On the other hand, air escaping through a gastrointestinal tract perforation or leak usually represents an intra-abdominal catastrophe requiring urgent intervention. This intriguing subject has been dealt with quite extensively based on plain film radiography findings in the past 50 years, and has lately also been studied on CT. This review discusses factors influencing the prevalence of PP and its range of duration. PMID- 15272553 TI - Retained surgical materials in the postoperative abdomen and pelvis. AB - The imaging appearances of surgical materials in the postoperative abdomen and pelvis can be confusing and difficult to interpret. With the increasing complexity of surgical procedures and more frequent use of postoperative imaging, the radiologist needs to be familiar with the imaging characteristics of a variety of intentionally and unintentionally placed surgical materials and devices. In addition, they must be differentiated from postoperative complications such as hematoma or abscess. PMID- 15272554 TI - [SOS-inducible DNA polymerases and adaptive mutagenesis]. AB - Stability of genomes of living organisms is maintained by various mechanisms that ensure high fidelity of DNA replication. However, cells can reversibly enhance the level of replication errors in response to external factors. As mutable states are potentially involved in carcinogenesis, aging, and resistance for pathogenic agents, the existence of these states is of great importance for human health. A well-known system of inducible mutation is SOS response, whose key component is replication of damaged DNA regions. Inducible mutation implies a contribution of SOS response to the adaptation of a bacterial population to adverse environments. There is ample evidence indicating the primary role of SOS response genes in the phenomenon of adaptive mutation. The involvement of the SOS system in adaptive mutagenesis is discussed. PMID- 15272555 TI - [Isolation and primary identification of methylotrophic yeast Hansenula polymorpha mutants for peroxisome biogenesis]. AB - After exposure of cells of the methylotrophic yeast Hansenula polymorpha HF246 leu1-1 to N-nitro-N-nitrosoguanidine, a collection of 227 mutants unable to grow on methanol at elevated temperature (45 degrees C) was obtained. Ninety four ts mutants (35% of the total number of mutants), which were unable to grow on methanol only at 45 degrees C but could grow at optimal temperature (37 degrees C), were isolated. Complementation analysis of mutants using 12 deletion mutants for genes of peroxisome biogenesis (PEX) (available in this yeast species by the beginning of our work) allowed to assign 51 mutants (including 16 ts) to the separate group of mutants unable to complement deletion mutants with defects in eight PEX genes. These mutants were classified into three groups: group 1 contained 10 pex10 mutants (4 ts mutants among them); group 2 included 19 mutants that failed to complement other pex testers: 1 pex1; 2 pex4 (1 ts); 6 pex5 (5 ts); 3 pex8; 6 (3ts)- pex19; group 3 contained 22 "multiple" mutants. In mutants of group 3, hybrids with several testers do not grow on methanol. All mutants (51) carried recessive mutations, except for mutant 108, in which the mutation was dominant only at 30 degrees C, which suggests that it is ts-dominant. Recombination analysis of mutants belonging to group 2 revealed that only five mutants (two pex5 and three pex8) carried mutations for the corresponding PEX genes. The remaining 14 mutants yielded methanol-utilizing segregants in an arbitrarily chosen sample of hybrids with the pex tester, which indicates mutation location in other genes. In 19 mutants, random analysis of ascospores from hybrids obtained upon crossing mutants of group 3 with a strain lacking peroxisomal disorders (ade11) revealed a single mutation causing the appearance of a multiple phenotype. A more detailed study of two mutants from this group allowed the localization of this mutation in the only PEX gene (PEX or PEX2). The revealed disorder of complementation interactions between nonallelic genes is under debate. PMID- 15272556 TI - [Characterization of missense mutations in the SUP45 gene of Saccharomyces cerevisiae encoding translation termination factor eRF1]. AB - Collection of missense mutations in the SUP45 gene of Saccharomyces cerevisiae encoding translation termination factor eRF1 has been obtained by different approaches. It has been shown that most of isolated mutations cause amino acid substitutions in the N-terminal part of eRF1 and do not decrease the eRF1 amount. Most of mutations studied do not abolish eRF1-eRF3 interaction. The role of the N terminal part of eRF1 in stop codon recognition is discussed. PMID- 15272557 TI - [Bipolar manifestation of cataleptic reactions in rats]. AB - Selection of GC rats for the predisposition to cataleptic freezing has increased not only the frequency, intensity, and duration of freezing, but also the proportion of irritable or "nervous" rats with enhanced anxiety, defensive behavior with vocalization, jerky running, and jumpiness. An increased amplitude of the startle reflex is a correlate of this "nervousness." The results of the comparison of some behavioral characters in the nervous and freezing GC rats, as well as in F1 and F2 offspring from homogeneous crosses between nervous and freezing GC rats suggest that cataleptic freezing and nervousness are two poles of the same bipolar catatonic reaction. They have a common mechanism, with the alternative or preferential expression of one particular form of the reaction is determined by the external and internal environments or the set of modifier genes in the given individual. PMID- 15272558 TI - [Cytogenetic maps of lampbrush chromosomes of newts of the genus Pleurodeles: an algorithm of lampbrush chromosome identification in Pleurodeles waltl by immunocytochemical marker-loop staining with polyclonal anti-Ro52 antisera]. AB - Our work was aimed at developing a simple and effective method of identification of most or all chromosomes of Pleurodeles newts. To this end, we used DAPI staining of the chromomeres of newt lampbrush chromosomes and immunochemical reactions between the ribonucleoprotein (RNP) marker loops and polyclonal antibodies against human zinc-finger protein Ro52 (52-kDa Ro/SS-A). A method has been developed to obtain newt lampbrush chromosome preparations. Cytological maps of P. waltl chromosomes (Spanish population/subspecies) showing distributions of chromomeres and marker loops along the chromosome length were constructed. PMID- 15272559 TI - [Analysis of the expression system of gene radius incompletes in Drosophila melanogaster]. AB - The genetic expression system of gene radius incompletes was studied by substituting regions of chromosome 3 from line th st sr ca, which has drastically reduced ri expression, to regions of chromosome 3 from a "selection" line ri s having very high expression of this gene. Virtually all chromosome 3 was shown to carry polygenes controlling phenotypic expression of oligogenic mutation ri. Chromosome regions making large (as well as small and even negative) contributions to sizes of the distal and proximal fragments of the latitudinal wing vein were found. These results suggest that the genetic system of expression does not correspond to the generally accepted postulates of K. Mather on equal, small, and additive contributions of polygenes. PMID- 15272560 TI - [Construction of immune lines with complex resistance to leaf rust and powdery mildew in common spring wheat cultivar Saratovskaya 29]. AB - Immune lines resistant both to leaf rust and to powdery mildew were constructed on the basis of common wheat cultivar Saratovskaya 29. Synthetic wheat Triticum timopheevii/Aegilops squarrosa (AAGGDD, 2n = 42) of Savov (Bulgaria) was used as a source of resistance genes. Using cytological analysis of BC2, we selected resistant plants (21") free from meiosis 1 (M1) defects. With these plants and continuous selection, BC8-BC9 immune lines were obtained. The lines were shown to carry new resistance genes differing from the known ones, and were proposed as donors of immunity to the diseases. PMID- 15272561 TI - [Expression of fertility during morphogenesis in self-pollinated backcrossed progenies of barley-wheat amphiploids]. AB - The fertility characteristics expressed during morphogenesis in first-generation self-pollinated backcrossed progenies (BC1) obtained from amphiploid barley-wheat hybrids [Hordeum geniculatum All. (2n = 28) x Triticum aestivum L. (2n = 42)] (2n = 70) backcrossed with common wheat were studied. It was found that, in the case of self-pollination of BC1 plants, karyotype stabilization leads to the formation of alloplasmic euploid (2n = 42), telocentric substitution (2n = 40 + 2t), and telocentric addition (2n = 42 + 2t), (2n = 42 + 2t) plant forms, which may serve as the sources of the respective alloplasmic lines of common wheat. That the expression of fertility characters in BC1F8 plants was shown to depend on growth conditions. The main mechanism of hybrid incompatibility of BC1F1-BC1F8 plants was expressed as grass-clump dwarfism. PMID- 15272562 TI - [Phylogenetic relationships and intraspecific variation of D-genome Aegilops L. as revealed by RAPD analysis]. AB - RAPD analysis was carried out to study the genetic variation and phylogenetic relationships of polyploid Aegilops species, which contain the D genome as a component of the alloploid genome, and diploid Aegilops tauschii, which is a putative donor of the D genome for common wheat. In total, 74 accessions of six D genome Aegilops species were examined. The highest intraspecific variation (0.03 0.21) was observed for Ae. tauschii. Intraspecific distances between accessions ranged 0.007-0.067 in Ae. cylindrica, 0.017-0.047 in Ae. vavilovii, and 0.00 0.053 in Ae. juvenalis. Likewise, Ae. ventricosa and Ae. crassa showed low intraspecific polymorphism. The among-accession difference in alloploid Ae. ventricosa (genome DvNv) was similar to that of one parental species, Ae. uniaristata (N), and substantially lower than in the other parent, Ae. tauschii (D). The among-accession difference in Ae. cylindrica (CcDc) was considerably lower than in either parent, Ae. tauschii (D) or Ae. caudata (C). With the exception of Ae. cylindrica, all D-genome species--Ae. tauschii (D), Ae. ventricosa (DvNv), Ae. crassa (XcrDcrl and XcrDcrlDcr2), Ae. juvenalis (XjDjUj), and Ae. vavilovii (XvaDvaSva)--formed a single polymorphic cluster, which was distinct from clusters of other species. The only exception, Ae. cylindrica, did not group with the other D-genome species, but clustered with Ae. caudata (C), a donor of the C genome. The cluster of these two species was clearly distinct from the cluster of the other D-genome species and close to a cluster of Ae. umbellulata (genome U) and Ae. ovata (genome UgMg). Thus, RAPD analysis for the first time was used to estimate and to compare the interpopulation polymorphism and to establish the phylogenetic relationships of all diploid and alloploid D genome Aegilops species. PMID- 15272563 TI - [Effective size of subpopulations in early-run sockeye salmon Oncorhynchus nerka from Azabach'e Lake (Kamchatka): the effect of relative reproductive success of different-year cohorts]. AB - The effect of variation in reproductive success of cohorts of different year of birth (within generation) on the effective subpopulation (breeding group) size in early-run sockeye salmon Oncorhynchus nerka from Azabach'e Lake (Kamchatka). The annual variation in census size and overlapping of year classes reduced the ratio of the effective subpopulation size to the census size by 7 to 88% in different subpopulations. The total effect of the variance of reproductive success in individual years and the variance of reproductive success of different cohorts reduced the effective size/census size ratio by 68-96%. PMID- 15272564 TI - [Inheritance of hypodontia in Kerry Blue Terrier dogs]. AB - Integral segregation analysis, which earlier proved efficient in studying complex hereditary diseases in humans and have been introduced in animal genetics for several years, was used to analyze the inheritance of hypodontia by premolars in Kerry Blue Terrier dogs. Dental formulas have been determined in 598 out of 911 animals united into a single large, complex pedigree. The results of integrated segregation analysis indicated heterogeneous genetic control of different forms of hypodontia. The geneses of different premolars in dogs have been demonstrated to be described by different models of inheritance: the absence of the second premolars can be described by the recessive major-gene model, whereas the agenesis of the fourth premolars have a more complex genetic mechanism and cannot be described by the model of a simple major-gene control. PMID- 15272565 TI - [Integrated population genetic and medical genetic study of two raions of the Tver oblast]. AB - An integrated medical genetic an population genetic study has been performed in two raions (administrative districts) of the Tver oblast (region) of Russia: the Udomlya raion located in the zone affected by the Kalininskaya Nuclear Power Plant and the Ostashkov raion, which served as a control district. No significant differences has been found with respect to the genetic parameters studied. The values of these parameters in the populations of the town of Udomlya, the town of Ostashkov, the Udomlya raion, and the Ostashkov raion, respectively, are the following: random inbreeding, 0.00006, 0.00011, 0.000167, and 0.000366; endogamy index, 0.05, 0.43, 0.30, and 0.42; local inbreeding, 0.0003, 0.00045, 0.0009, and 0.0011; the degree of isolation by distance, 0.0003, 0.00045, 0.0009, and 0.0005; sigma, 2098, 1338, 1473, and 1189; the load of autosomal dominant (AD) diseases, 0.71, 0.92, 0.92, and 1.37; the load of autosomal recessive (AR) diseases, 0.68, 0.69, 0.67, and 0.82; and the load of X-linked diseases, 0.18, 0.64, 0.83, and 0.27. PMID- 15272566 TI - [Genetic demographic description of the Ust-Aldan rural population of Sakha Republic (Yakutia): ethnic, sex, and age compositions, vital statistics, and surname structure]. AB - Information on the sex, age, and ethnic compositions; reproductive parameters; intensity of natural selection (Crow's indices); and surname diversity of three rural populations (the Byadi, Dyupsya, and Cheriktey villages) of the Ust-Aldan ulus (district) of Sakha Republic (Yakutia) has been analyzed. The rural Yakut population of the Ust-Aldan ulus is demographically young (the mean age 25-31 years) and characterized by low outbreeding, unfavorable sex ratio in both prereproductive and reproductive ages, and high fertility (3.58-5.45 children surviving until the reproductive age per woman that has completed the reproductive period), although the actual reproductively active period is shorter than half its physiological duration. In the structure of total selection, the differential-fertility component is considerably greater than the differential mortality component (Itot = 0.625, Im = 0.093, and If = 0.487). In the villages studied, some surnames are accumulated (45-65% of the population have five most frequent surnames), which determines the low surname diversity (alpha = 11.62 25.19) and high random isonymy (Ir = 0.0391-0.0823). PMID- 15272567 TI - [Genetic demographic description of the Ust-Aldan rural population of Sakha Republic (Yakutia): migrations and marriage structure]. AB - Migrations, dynamics of the gametic structure of rural populations, and marriage structure with respect to birthplaces and inbreeding estimated from isonymy have been studied in the Ust-Aldan ulus (administrative district) of Sakha Republic (Yakutia). The villages studied (Byadi, Dyupsya, and Cheriktey) are characterized by intense migration; however, the migration radius is small (most migrations occur within the district). The rural populations studied differ in the intensities and directions of gamete flows and their dynamics. There is no substantial gamete flow into the Ust-Aldan population from outside Sakha Republic. About 50% of marriages contracted in this population are homolocal (between residents of the same district); the endogamy is low (15%). In most cases of heterolocal marriages (contracted between residents of different districts), one of the spouses is a local resident. The inbreeding estimated from isonymy is FTT = 0.002930 in Yakuts; it is mainly accounted for by the nonrandom component (FIS = 0.002232 and FST = 0.000700). PMID- 15272568 TI - [STR polymorphism in populations of indigenous Daghestan ethnic groups]. AB - Genomic diversity of 21 STR loci has been studied in six ethnic populations of Daghestan (the Caucasus), namely, Avars, Dargins, Kubachians, Lezgins, and Nogais, and the results have been compared with these data for European, African, and East Asian ethnic groups. Daghestan is unique in its ethnic diversity, which is the greatest in the Caucasus: 26 out of approximately 50 autochthonous ethnic groups of the Caucasus live there. The genetic origin of this wide ethnic diversity of Daghestan and the Caucasus as a whole is still obscure. The genetic heterogeneity of Daghestan populations has been found to be lower than that of most other populations in the world. This is explained by a prolonged isolation and gene drift in their demographic history. Generalized genetic distances between ethnic groups calculated for the whole set of loci studied allow differentiating Asian populations from African ones, with European populations occupying intermediate positions. All Daghestan ethnic populations form a distinct common group together with some European populations (Finnish, Polish, and French). Nogais are genetically close to Southeast Asian populations. The genetic closeness and the apparently equal genetic diversity of Daghestan and European populations suggest that the ethnic differentiation of the ancestral populations of Daghestan and European ethnic groups occurred in the earliest populations of modern humans. PMID- 15272569 TI - [Population study of frequency of methylenetetrahydrofolate reductase C677T gene polymorphism in Yakutia]. AB - The enzyme methylenetetrahydrofolate reductase (MTHFR) catalyzes synthesis of 5' methylenehydrofolate, which is the methyl donor for the conversion of homocysteine to methionine. According to the numerous literature data, polymorphic variant of the MTHFR-encoding gene, C677T, is associated with hyperhomocysteinemia, vascular pathologies, neural tube defects, dementia, perinatal mortality, mental disorders, long-term neurodegenerative disorders, lens displacement, arachnodactyly, and venous thromboses. The present study was focused on the analysis of the C677T polymorphism (missence mutation leading to the replacement of cytosine by thymine at position 677) of the MTHFR gene in three indigenous populations of the Republic of Sakha (Yakutia), living in the settlements of Cheriktei, Byadi, and Dyupsya. Comparison of the genotype and allele frequencies revealed no substantial differences between the three Yakut populations, as well as between Yakuts and other Mongoloid ethnic groups. PMID- 15272570 TI - [Analysis of the fixation of discrete genetic structures by means of an integral model of evolution in a Mendelian one-locus population of diploid organisms]. AB - An integral model of the evolution of a Mendelian one-locus population of diploid organisms with continual allele diversity developing under density-limiting conditions or without density limitation has been proposed and analyzed. The model was used to study the mechanism of the appearance of discrete genetic structures, i.e., the fixation of a limited number of alleles. Local resistance of the resultant genetic distributions to homogeneous equiprobable mutations has been demonstrated. PMID- 15272571 TI - [Study of the mechanism for regulating ribR gene activity in Bacillus subtilis]. AB - Analysis of the phenotypic manifestation of inactivation of several genes from the ytlI-ytnM operon containing the ribR gene and results of Northern hybridization showed that the ribR gene does not have the self promoter and is transcribed from the main promoter of the ytlI-ytnM operon. Two sites of single nucleotide substitutions leading to derepresson of ribR gene were identified between the putative main promoter and transcription start of the ytlI-ytnM operon regulatory region. PMID- 15272572 TI - Making the business case for quality: what do consumers and employers want--and can they get it? AB - As winter moved into spring this year, the debate ratcheted up in the halls of Congress, in hospital corridors, in payer meetings, and in executive boardrooms on the question of what is quality healthcare--and how can it best be monitored and delivered? And, can it be viewed as the panacea to escalating healthcare costs? The answers, it seems, depend on to whom you are talking. Some say more measures are needed. Some say the measures available are confusing. And some say reengineering of the system is what's needed. PMID- 15272573 TI - AHRQ, IOM weigh in on developing a health-literate America. AB - Low health literacy is "a very underappreciated problem," Institute of Medicine (IOM) president Harvey Fineberg, MD, said at a Washington, DC, media briefing that introduced reports on health literacy from the IOM and the Agency for Healthcare Research and Quality. PMID- 15272574 TI - Registries are evolving to meet needs of chronically ill patients. AB - Registries for chronic disease management have been around for years. But as disease registries are adopted more widely, their definitions and functions are evolving--accompanied by a better understanding of how to manage a population of patients with chronic diseases--according to a new report. PMID- 15272575 TI - [Proteasome inhibitors]. AB - Bortezomib, one of the proteasome inhibitors, has been approved in the United States for multiple myeloma as a second-line chemotherapy. In Japan, bortezomib has been used for for phase I clinical trials for multiple myeloma. As the action mechanism, it has been proposed that bortezomib inhibits NF-kappaB via IkappaB alpha. It also demonstrated positive results for non-Hodgkin's lymphoma and non small cell lung cancer, but not for colorectal cancer or chronic lymphocytic leukemia. The mechanism of drug resistance has been well analized. Bortezomib is very effective but still induces adverse effects including hypotension especially when there is an overdose. Medical oncologists or hematology/oncologists must exert due caution. PMID- 15272576 TI - [Biomarker in gynecologic malignancies]. AB - We selected ovarian cancer as being the most representative of the gynecologic malignancies since it has the largest number of tumor markers now in clinical use. First, variety of serum tumor markers were developed and regularly used to detect the existence of ovarian cancer and its stage. These markers of monoclonal antibodies could detect three different classes of cell surface antigen. CA 125, CA 130, CA 602 are antibodies raised against the core protein of the proteoglycan molecule, whereas CA 19-9, CA 50, KMO-1 and CA 72-4, STN, CA 546 are antibodies against a different portion of the glycosaminoglycan chain from the core protein. These markers, when combined with another group of tumor markers for discriminating the variety of pathological types of ovarian cancer, might have the potential to provide a better predictive value. Second, biomarkers for the detection of early-stage ovarian cancer are urgently needed and developed also in gynecologic tumors. These include a ovarian cancer-specific proteomic patterns generated by mass spectroscopy and single nucleotide polymorphism (SNP) digital analysis combined with assessment of allelic imbalance. Third, for monitoring the effect of treatment with cytostatic drugs, various types of biomarkers could be used as surrogate markers for the treatment depending on the mechanism of the effects of the drug used. For treatment with Bryostatin, a strong protein kinase C (PKC) stimulator, PKC activity promises to be an effective marker. For the trials with bevacizumab, anti-VEGF antibody, VEGF and its associated bFGF, CD-31, and thrombospondin-1 (TSP-1), are leading candidates for monitoring markers. PMID- 15272577 TI - [Significance of current bio- and molecular-markers in urological cancers]. AB - In order to reconfirm their diagnostic and predictive significance, bio- and molecular-markers of urological cancers (prostate, urothelial, kidney, and testicular cancer) are reviewed. In prostate cancer, although prostatic specific antigen (PSA) is a gold standard marker for its screening and monitoring, recent studies have revealed that PSA-related markers have additional information for patients with a gray zone PSA score. Prostate-specific membrane antigen (PSMA) may provide new applications for detection of high-grade cancer or microscopic circulating prostate cancer cells in the blood. In urothelial cancers, several urinary markers are available and may be helpful in the diagnosis of lower-grade urothelial cancers, which have a low sensitivity of urinary cytology. In testicular cancer, lactate dehydrogenase (LDH), alpha-fetoprotein (AFP), and human chorionic gonadotropin (hCG) are essential markers not only to determine the tumor stage but also the prognosis of the patient. In renal cell carcinoma (RCC), unfortunately, no definitive biomarker is available for its diagnosis and monitoring. Thus, we must find new specific bio- and molecular-markers that reflect the biological activity of RCC. PMID- 15272578 TI - [Biomarkers for neoplasmas in digestive organs]. AB - This review is concerned with the usefulness and the problem of biomarkers for cancer of digestive organs. Carcinoembryonic antigen (CEA) is a most popular and useful tumor marker for cancer of digestive organs. Squamous cell carcinoma (SCC) antigen and CYFRA have been reported as a useful tumor marker for esophageal cancer. CEA and CA 19-9 are a good prognostic factor in gastric cancer patients. The post-operative increase of serum CEA can be a predictive marker for the patients of colorectal cancer. Development of a radioimmunoassay for highly sensitive detection of tumor markers, they are considered to be useful for monitoring after treatment. But are not useful for the early diagnosis. The diagnosis of hepatocellular carcinoma (HCC) is based mainly on serological markers, such as alpha-fetoprotein and PIVKA-II. The two are useful complementary markers of HCC because they do not correlate with each other. But the problem of the false-positive rate for the patients with chronic hepatitis or liver cirrhosis is still remained. A typical marker of pancreatic and bile duct cancer is carbohydrate antigen, but the sensitivity of these markers is only 50%. Recent molecular biological analysis may be used as effective biomarkers in the diagnosis, prognosis, therapy, and risk assessment of digestive cancer. PMID- 15272579 TI - [Biomarkers in breast cancer]. AB - Biomarkers are measured in the management of breast cancer patients for the following purposes. (1) Early detection of breast cancer: blood tumor markers such as CA 15-3 are useless for this detection because of a low sensitivity. Proteomics profiling has recently been investigated using blood or nipple aspirate fluid for the detection. Measurement of CEA and HER 2 in abnormal nipple discharge has been approved for diagnosis of breast cancer in Japan. (2) Monitoring of breast cancer patients: serum tumor markers are routinely measured for early detection of recurrent diseases, evaluation of therapeutic response and monitoring outcome of patients by a majority of breast cancer experts in Japan. Study results investigated by the Study Group of the Japanese Breast Cancer Society in 2001 are presented with regard to the questionnaire survey on the present status of tumor marker measurement and the clinical study on usefulness of tumor markers for the evaluation for therapeutic response. (3) Prognostic factors: new biomarkers have been investigated to select patients at high risk for distant metastases, which could not be selected by classic prognostic factors. Three prognostic factors (UPA/PAI-1, cyclin E, gene profiling), which were discussed at the 8th St. Gallen International Consensus Meeting last year, are mainly discussed. (4) Predictive factors for therapeutic response: hormone receptors (HR) have been used as reliable predictive factors for response to endocrine therapy. Other biomarkers have been investigated to select patients with tumors HR-positive but unresponsive to endocrine therapy. Current status, clinical significance, problems and future directions on predictive factors for response to cytotoxic chemotherapy are also discussed. PMID- 15272580 TI - [Biochemical markers in bone metastasis]. AB - Certain types of cancers have a strong propensity to metastasize to bone, which requires combination of multiple factors responsible for the different steps of metastasis. Bone metabolic markers are now widely used in clinical practice and give useful information on the ongoing bone metabolism, reflecting the activity of bone-resorbing osteoclasts and bone-forming osteoblasts. Bone markers have a potential as diagnostic tools for bone metastasis, and are useful in monitoring the response to anticancer as well as antiresorptive therapies. Since bone metabolic markers alone are insufficient for the diagnosis and assessment of bone metastasis, it is important to combine bone markers with tumor-related markers and imaging studies such as scintigraphy and MRI. More recently, soluble receptor activator of nuclear factor-kappaB ligand (RANKL) and osteoprotegerin (OPG) have been implicated as markers for osteoclastogenic activity. Serum levels of these factors and/or their ratios may provide additional information on the severity of bone disease and the prognosis. PMID- 15272581 TI - [Molecular-target drug]. AB - Cancer metastasis involves the complicated steps of tumor growth, angiogenesis, invasion and adhesion. At present new drugs targeting particular molecule (s) responsible for such cancer progression and metastasis have been developed in clinics. Major endpoints for cancer treatment should be prolongation of survival and maintenance of QOL. However, clinical development of such molecular-target based drugs is associated with difficulties in evaluating the efficacy in phase I/II studies prior to entering phase III study, because many of the targeted drugs seem cytostatic rather than cytocidal to tumors. New approaches incorporating technologies of genomics and proteomics may provide an expanding repertoire of molecular targeted therapeutics for clinical evaluation. In this review, the significance and problems of biomarkers available for clinical evaluation of molecular targeted drugs are discussed. PMID- 15272582 TI - [Effect of Cepharanthin to prevent radiation induced xerostomia in head and neck cancer]. AB - We retrospectively examined the effect of Cepharanthin to prevent radiation xerostomia in 37 cases of head and neck cancer. In the Cepharanthin group, the degree of xerostomia was milder than in the non-Cepharanthin group in spite of higher normal tissue complication probability (NTCP) and mean dose (MD) of parotid glands. In the non-Cepharanthin group, the degree of xerostomia was significantly correlated with NTCP and MD of parotid glands. MD of parotid glands and use of Cepharanthin were significantly related to more severe xerostomia by multivariate analysis with logistic regression. Cepharanthin may prevent radiation xerostomia after radiotherapy for head and neck cancer. PMID- 15272583 TI - [Examination of second-line therapies following administration of low-dose TS 1+CDDP for highly-advanced gastric cancer]. AB - Sixteen patients with highly-advanced gastric cancer were administered low-dose TS-1 and CDDP as a first-line treatment, followed by either paclitaxel or CPT 11/CDDP as a second-line treatment. The results of the 2 second-line treatments are reported herein. Overall response rate for the first-line treatment was 55.6%. For the second-line treatments, responses were noted in both the paclitaxel group and the CPT-11/CDDP group. Overall MST was 16.3 months and 1 year survival was 60%. The paclitaxel group, however, showed significantly better prognoses than the CPT-11/CDDP group. Adverse reactions to the first-line treatment were grade 3 leukopenia in 1 patient, with no other reactions over grade 2 observed. No adverse reaction greater than grade 2 was noted during administration of the second-line treatments. These results appear to present ample data that a first-line treatment of low-dose TS-1/CDDP followed by a second line treatment of paclitaxel at 1/week in the outpatient setting yields improved prognoses and minimal adverse reactions. PMID- 15272584 TI - [Choice of chemotherapeutic drugs for colorectal cancers by DPD and OPRT activities in cancer tissues]. AB - 5-FU is the most widely used anticancer drug for digestive system cancers, and it has been recently reported that the effect of 5-FU varies with the amount of enzymes that are involved in the drug metabolism in the cancer tissue. In this report, we measured DPD and OPRT activities in the normal mucosa and colorectal cancer tissues and compared them with clinicopathological factors to examine the choice of chemotherapy for colorectal cancers. Forty-six patients with colorectal carcinoma (28 colon and 18 rectal cancers), which were resected in our department from 1999 to March 2003, were examined. There was no significant difference in the DPD activities between the normal and cancer tissues, whereas OPRT showed significantly higher activity in the cancer tissues. Although we did not find a relation between the DPD activity and clinicopathological factors, the OPRT activity showed a significant difference between gender and classification of tumor, and mucinous adenocarcinoma showed significantly lower OPRT activity than differentiated adenocarcinoma. The DPD activity of mucinous adenocarcinoma is related to catabolism of 5-FU is equal to that of adenocarcinoma, however, the OPRT activity is related to a main pathway of 5-FU phosphorylation is significantly lower. Thus, mucinous adenocarcinoma of the colon was more resistant to 5-FU than the differentiated adenocarcinoma. Therefore, for chemotherapy of mucinous adenocarcinoma, anticancer agents other than 5-FU should be selected. PMID- 15272585 TI - Cisplatin and gemcitabine in patients with metastatic cervical cancer. AB - PURPOSE: To determine the therapeutic efficacy of cisplatin plus gemcitabine in the treatment of patients with metastatic cervical cancer. METHODS AND MATERIALS: A total of 51 patients were enrolled in this study. The median age was 46 years (34-67). Thirty-five patients were previously treated to the pelvis by radical radiotherapy, one patient was treated by surgery and one by surgery and postoperative radiotherapy. There were 14 patients with stage IVB cervical cancer who were previously untreated. The sites of metastases were 10 in the lungs, 5 in the supraclavicular nodes, 9 in the paraaortic nodes, 4 in the liver, 3 in the inguinal nodes, 5 in both supraclavicular nodes and lungs, 9 in both supraclavicular and paraaortic nodes, 1 in both supraclavicular and inguinal nodes, 1 in both lung and inguinal nodes, 2 in both lung and paraaortic nodes and 2 in both liver and lungs. Fine needle biopsies were done in all metastatic sites except for multiple lung and multiple liver metastases. Cisplatin was administered as an i.v. infusion on day 1 (70 mg/m2). Gemcitabine was administered as an i.v. infusion for over 30 minutes on day 1 and 8 (1,250 mg/m2) in a 21 day cycle. RESULTS: Forty patients were available for evaluation, with 3/40 (7.5%) achieving a complete response, 27/40 (67.5%) achieving a partial response (OR=30/40, 75%), 5/40 (12.5%) having a stable disease, and 5/40 (12.5%) a progressive one. Eleven patients were not assessable because of patient refusal for further treatment and loss of follow up. Myelosuppression was the major toxicity. Grade 3 or 4 anemia and granulocytopenia occurred at a frequency of 25.5% and 29.4%, respectively. Grade 3-4 thrombocytopenia was found in 3.8%. There were 3 (5.8%) patients who developed grade 4 neutropenia and fever. No other major side effects were found other than alopecia and usual gastrointestinal toxicities such as anorexia, nausea and vomiting. There were no treatment related deaths. The median time to progression was 8.3 months and the median survival was 9.6 months. With a median follow up of 7.7 months (range, 0.3 to 22.1), 30% of the patients were alive at 12 months. CONCLUSION: In this study of patients with metastatic cervical cancer, the combination of cisplatin and gemcitabine treatment induced a high response rate. PMID- 15272586 TI - [Effect of Cepharanthin in preventing radiation induced normal tissue damage in prostate cancer]. AB - A total of 97 patients with prostate cancer who underwent radiotherapy were retrospectively reviewed to analyze the protective efficacy of Cepharanthin for acute or late toxicity to the bladder/urethra and rectum. Cepharanthin administrations were divided into 3 groups: intravenous, oral, and non administration. Acute urinary toxicity was significantly milder for the intravenous group than for the oral and non-administration groups. The protective efficacy of Cepharanthin was not approved for acute rectal toxicity, but late rectal toxicity was significantly milder for the intravenous group than for the oral group. Intravenous Cepharanthin administration may prevent acute or late toxicity by radiotherapy for prostate cancer. PMID- 15272587 TI - [A case of brain metastasis of small cell lung cancer improved with nogitecan hydrochloride (topotecan) after prophylactic irradiation]. AB - A 55-year old woman was treated with chemoradiotherapy for limited type small cell lung cancer. She was then treated with prophylactic cranial irradiation. However, local recurrence and metastatic brain tumors were detected after a year. Three courses of chemotherapy with nogitecan (topotecan) ameliorated the size and numbers of the tumors pronouncedly. Nogitecan (topotecan) is suggested to be an effective therapeutic agent for brain metastasis of small cell lung cancer after prophylactic cranial irradiation. PMID- 15272588 TI - [A case of advanced cervical and upper thoracic esophageal carcinoma completely responding to chemoradiotherapy with TS-1 and cisplatin]. AB - A 52-year-old man was hospitalized for evaluation of dysphagia. Esophagography depicted an irregular narrowing extending 7 cm from the cervical esophagus to the upper thoracic esophagus. Esophagoscopy with biopsy showed cervical esophageal cancer narrowing the lumen. Surgery was contraindicated because of a previous cardiac infarction. The patient selected concurrent chemoradiotherapy with TS-1 and cisplatin. The first course included 30 Gy of radiotherapy given over 3 weeks, together with daily oral administration of TS-1 (120 mg/day) for 2 weeks, and a 24-h infusion of cisplatin (70 mg/m2) on day 8. After a second course of chemoradiotherapy, 4 courses of chemotherapy with TS-1 and cisplatin were administered at 4-week intervals. After additional chemotherapy, esophagoscopy and cervical CT showed that the primary lesion had disappeared. Two years after initial hospitalization, no recurrence has been detected. PMID- 15272589 TI - [A case of advanced gastric cancer with a tumor embolus in the portal vein successfully treated with TS-1 and CDDP]. AB - A 71-year-old man was admitted to our hospital in February 2002 with a diagnosis of advanced gastric cancer with a tumor embolus in the portal vein. TS-1 (120 mg/day) was administered orally daily for 21 days, and CDDP (90 mg/day) was infused intravenously on day 8. After 1 course of this regimen, medication was discontinued in accordance with the patient's request. The patient was readmitted with a history of tarry stools in July 2003. Despite no cancer treatment for almost 1.5 years, the primary lesion and the metastatic lymph nodes had decreased significantly in size and the tumor embolus in the portal vein had disappeared completely on the CT scan. He was therefore treated with TS-1 alone (120 mg/day) under a 4-weeks-on and 2-weeks-off regimen. After 1 course of TS-1 administered alone, the primary lesion showed a further significant decrease in size as viewed by GI endoscopy, and biopsies did not reveal any evidence of malignancy. PMID- 15272590 TI - [A case of effective treatment of oral TS-1 and intravenous MMC administration for multiple metastases of gastric cancer]. AB - A 49-year old man underwent distal gastrectomy (D3) for circumferential type 3 cancer at the gastric antrum and cholecystectomy in September 2002. During the surgery, multiple metastases were observed predominantly in the left lobe of the liver, and lateral segmentectomy was performed as non-curative (curability-C) resection leaving the small metastases in the right lobe of the liver. Based on the results of chemo-sensitivity tests (5-FU 15.0%, CDDP 34.0%, MMC 35.3%, TXT 0.0%), we started to administer TS-1 (100 mg/day for 4 weeks followed by a 2-week rest interval) and MMC (10 mg/body on day 1). Due to leukocytopenia, the regimen was changed to TS-1 (100 mg/day for 4 weeks followed by a 2-week rest interval) and MMC (4 mg/body every other week [day 1, 14]) from the second course. Levels of tumor markers dropped and liver metastatic lesions remarkably decreased in size by CT after the third course. In conclusion, a combination of TS-1/MMC may be regarded as one option for postoperative adjuvant chemotherapy for outpatients. PMID- 15272591 TI - [A case of TS-1 resistant recurrent gastric cancer responding to TS-1 +CPT-11 combination therapy]. AB - A 71-year-old man underwent distal partial gastrectomy for gastric cancer. Four years after surgery, the tumor marker was elevated. Examinations by computed tomography (CT) revealed para-aortic lymphnode swelling and hydronephrosis. The patient treated oral administration of TS-1 (120 mg/day). After 3 courses of treatment of TS-1, progressive disease was observed. TS-1+CPT-11 (TS-1 120 mg/day day 1-14, CPT-11 100 mg/day day 1, 15) combination therapy was then chosen as second-line chemotherapy. After 5 courses of combination therapy, the tumor marker was decreased and para-aortic lymphnodes could not be detected by CT. Only grade 2 leukopenia was observed as an adverse event during the therapy. TS-1+CPT 11 combination therapy could be useful as the second-line chemotherapy for cases of TS-1 resistant recurrent gastric cancer. PMID- 15272592 TI - [A case of inoperable scirrhous gastric cancer that responded remarkably to a combination of TS-1+paclitaxel and showed complete loss of ascites]. AB - Complete loss of malignant ascites by combination chemotherapy of TS-1+paclitaxel was experienced. The case was a 56-year-old woman who was diagnosed with inoperable scirrhous gastric cancer with malignant ascites. The administered regimen of chemotherapy was TS-1 100 mg/day (80 mg/m2) for 2 weeks. Paclitaxel 60 mg/day (50 mg/m2) on day 1 and 8 of TS-1 intake, followed by 2-weeks rest. Partial response was confirmed by gastrography and gastrofiberscope after 3 courses were performed. Furthermore, computed tomography (CT) showed complete loss of malignant ascites. Adverse reactions were grade 3 leukopenia and grade 2 nausea, vomiting and diarrhea. This result indicates the possibility of combination chemotherapy of TS-1+paclitaxel becoming an effective option in treating inoperable scirrhous gastric cancer. PMID- 15272593 TI - [A case of advanced colon cancer responding to l-LV+5-FU as neoadjuvant chemotherapy]. AB - The patient was a 45-year-old woman who had ascending colon cancer with multiple paramesenteric and paraaortic lymph node metastases. Combined l-LV 400 mg/body (250 mg/m2) +5-FU 950 mg/body (600 mg/m2) therapy was carried out as neoadjuvant chemotherapy. After 2 cycles of this therapy, the lymph node metastases were not detectable on computed tomography. Thus, right hemicolectomy was performed (D3, CurB). It is suggested that l-LV+5-FU therapy may be useful for advanced colon cancer as neoadjuvant chemotherapy. PMID- 15272594 TI - [Successful treatment of 5-FU intraarterial infusion on the third postoperative day for a patient with multiple liver metastases from colon cancer and high risk of liver failure]. AB - A 68-year-old man with multiple liver metastases from stage 4 advanced descending colon cancer who underwent partial resection of the colon and simultaneous catheter insertion into the gastroduodenal artery for arterial infusion chemotherapy. On postoperative day 3, the multiple liver metastases had enlarged so rapidly that there was high risk of liver failure. Intraarterial infusion of 5 FU 600 mg/m2 (1,000 mg/body) for 6 hours weekly and intravenous administration of methylpredonisolone 125 mg were started for emergency chemotherapy on the third postoperative day. Only 1 course was sufficient for the patient be rid of oncologic emergencies and liver failure. After 3 courses, liver metastases showed significant reduction. PMID- 15272595 TI - [A case of advanced descending colon cancer with peritoneal dissemination responding to weekly high-dose l-leucovorin/5-fluorouracil combination therapy]. AB - We report a case in which l-leucovorin/5-fluorouracil (l-LV/5-FU) combination therapy was remarkably effective for non-resectable advanced descending colon cancer with carcinomatous peritonitis. A 65-year-old man complained of severe abdominal distension, abdominal pain and pulmonary failure, and was diagnosed as having ileus due to descending colon cancer. The patient underwent urgent open laparotomy to release the ileus condition on March 5, 2002. During the laparotomy, we established a diagnosis of nonresectable descending colon cancer accompanied by severe peritoneal dissemination and therefore performed only double-barreled transverse colostomy. The postoperative course was very good. Pathological examination of omental dissemination demonstrated moderately differentiated adenocarcinoma and cytology of ascites was class II. The levels of serum CEA and CA19-9 were within the normal ranges. l-LV/5-FU therapy was initiated postoperatively. Seven cycles of this chemotherapy regimen was performed with no apparent side effect during the treatment. There was no postoperative accumulation of carcinomatous ascites. The patient gained 15 kg compared with body weight admission. On abdominal computed tomography (CT), the primary lesion of the colon decreased 98% and there was no ascites found. To date, there has not been any sign of recurrence during the 19 months of follow-up after this therapy, and we are currently discussing closure of the transverse colostomy. This therapy may be useful for patients with advanced colon cancer accompanied by peritoneal dissemination. PMID- 15272596 TI - [Tamoxifen-induced severe hypertriglyceridemia--report of 3 cases]. AB - Tamoxifen, an anti-estrogen, has been used for a long time as an adjuvant therapy in cases of estrogen receptor positive breast cancer. Tamoxifen also demonstrates some weak estrogenic activity. A small increase in serum triglycerides is commonly found after tamoxifen administration. Herein we report 3 cases of sever hypertriglyceridemia due to tamoxifen. Case 1 recovered with tamoxifen withdrawal. Tamoxifen was replaced with toremifene in case 2. The level of triglyceride decreased significantly after the change of agent. Tamoxifen was discontinued and anastrozole administration was started in the third patient. Her triglyceride levels improved. Tamoxifen-induced severe hypertriglyceridemia seen in these patients was an effect of its estrogen action. Anastrozole has been used to treat postmenopausal metastatic breast cancer, and several clinical trials in the adjuvant setting are ongoing. Anastrozole does not affect lipid metabolism. Therefore, anstrozole might be safe for patiens with abnormal triglyceride profiles during tamoxifen treatment. We recommended that a periodic serum triglyceride check is needed for patients treated with tamoxifen. PMID- 15272597 TI - [Successful treatment of multiple pulmonary metastases of adenoid cystic carcinoma of the external auditory meatus with TS-1]. AB - The patient, a 67-year old man, was referred to us with refractory otorrhea of the left ear. The diagnosis was adenoid cystic carcinoma of the external auditory meatus, so the primary focus was excised. During the postoperative observation period, multiple metastases to the lung were noted, for which TS-1 (120 mg/day, given for 28 consecutive days, each course followed by a discontinuation of 14 days) was administered. After the completion of 2 courses, reductions in the metastatic pulmonary lesions and the amount of pleural effusion were noted, while adverse effects were limited to mild anorexia. Thus, it was possible to continue medication for 15 months on an ambulatory basis. It was indicated that TS-1 may be effective in the treatment of multiple pulmonary metastases of adenoid cystic carcinoma. PMID- 15272598 TI - [Five cases of de novo acute myeloid leukemia with trilineage myelodysplasia (AML/TMDS) achieved CR with the continuous drip infusion of low-dose etoposide and low-dose cytosine arabinoside combined with mitoxantrone (MEtA)]. AB - From 1998 to 2001, 5 consecutive cases of AML/TMDS entered our hospital and achieved complete remission (CR) with continuous drip infusion of low-dose etoposide and low-dose Ara-C combined with mitoxantrone (MEtA regimen). The ages of the 5 patients (4 males and 1 female) ranged 32 to 50 years-old, respectively. WBCs were 1,560-45,150/microl, blasts were 12-62%. Bone marrow aspirates revealed trilineage myelodysplasia with various number of blasts. These patients had an acute onset and no preceding hematologic disorders. They were diagnosed M2/TMDS or M4/TMDS. Continuous drip infusion of etoposide (50 mg/body/day) and Ara-C (30 mg/body/day) were given for 11-14 days and a bolus injection of mitoxantrone (8 mg/m2) was added for 2-3 days. Patient 5 was given additional MIT (6.7 mg/m2 on day 6). All cases achieved CR in 21-24 days after the end of the therapy. Toxicities were nausea, vomiting, stomatitis, alopecia and fever due to infection. All were well tolerable, however. Two patients are alive more than 4 years without relapse. MEtA regimen is available for AML/TMDS. PMID- 15272599 TI - [How can sentinel navigation surgery abbreviate mediastinal lymph node dissection for lung cancer?]. AB - Sentinel nodes (SNs) were examined for 101 patients who had peripheral type non small cell lung cancer less than 5 cm and had undergone systemic mediastinal lymph node dissection. The surgical procedure was lobectomy in 91, pneumonectomy in 3, and segmentectomy with lymph node dissection in 7. In the CT room, the site for RI injection was marked on the skin, and the angle and depth of the needle required to reach the peritumoral region was determined. The RI was then injected in the RI room. The radioactivity in the lymph nodes was counted before dissection (in vivo counting), and after dissection that (ex vivo counting). SNs were defined as any node for which the count was > or = 10 times than the background count. SN identification was finally based on ex vivo data. Of the 101 patients, SNs could be identified in 80 patients (80%). Patients whose SNs could not be identified had a significantly lower FEV1/FVC than those with identifiable SNs (p=0.025). Twenty six patients (33%) had SN in the mediastinum, the distribution of which depended on the lobe, ie the #4 lymph node station in the right upper lobe, the #4 in the right middle lobe, the #4 and 7 in the right lower lobe, the #5 in the left upper lobe, and the #7 in the left lower lobe. One false negative SN was detected in 25 patients with N 1 or N 2 disease (4%). In vivo and ex vivo counting showed 73% concurrence for the identification of SNs in mediastinal lymph node stations, of which rate was significantly higher than 40% in hilar lymph node stations (p<0.001). CONCLUSION: The SNs were identifiable in 80% of lung cancer patients, with 4% false negative by using a Tc-99 m tin colloid. SNs were difficult to identify in patients with a low level of FEV1/FVC, such as those with chronic obstructive pulmonary disease. The in vivo identification of mediastinal SNs was reliable as much as the ex vivo. Therefore, the in vivo identification of SNs in the mediastinum could be useful approach to guide mediastinal lymph node sampling or dissection. PMID- 15272600 TI - [Clinical usefulness of circulating tumor markers]. AB - There are many molecular tumor markers for diagnosing and monitoring cancer patients. Especially, quantitative assay for serum levels of tumor markers; such as AFP, CEA, PSA, hCG, CA 19-9 and CA 125, are frequently used in daily practice because of their relative specificities and usefulness to the common cancers. Though not suitable for early diagnosis, but they are used in monitoring patients with advanced caner, especially after treatments. Two of them, AFP and PSA, are also used in the screening and monitoring of high-risk groups, namely patients with chronic viral hepatitis and old male, who are the high risk for hepatoma and proste cancer respectively. Problems in using serum markers are; relatively low specificity and low sensitivity to cancer, confusing naming for similar markers that recognize almost the same molecule of cancer. Users must understand that CA 19-9, CA 50, KM-O 1 and SPAN-1 are in the same sialylated Lewis A group, and CA 125, CA 130 and CA 602; in the mucin antigen group, and STN, CA 54/61 and CA 72 4; in the sialyl Tn antigen group. Combination of two or more markers may inform us the biological characteristics of the cancer. For example, a germ-cell tumors may produce hCG and placental marker. That is of the choriocarcinoma type. Those with hCG and fetal antigens are the ordinal type of germ cell tumors, and those with AFP, CEA and cytokeratin are teratoma, and those with LDH and ALP only but negative for hCG and AFP must be seminoma. For the bronchial and alveolar carcinomas, CEA, SCC, NSE and cytokeratin 19 fragments are useful. Combination may be difficult for beginners but once understood, it will be an art in clinical oncology. PMID- 15272601 TI - Natural occurrence of trichothecenes on lodged and water-damaged domestic rice in Japan. AB - In 1998, a typhoon struck before rice harvesting in Japan, and the unpolished rice was found to be stained brown. Samples were collected and analyzed for the presence of trichothecenes using GC/MS. The mycotoxins deoxynivalenol (DON), fusarenon-X (Fus.-X) and nivalenol (NIV) were detected and confirmed with GC/MS. The quantity of trichothecenes was determined using GC-ECD. To our knowledge, this is the first report of the presence of the trichothecene Fus.-X in rice. PMID- 15272602 TI - [Studies on analysis of terpene resin in imported chewing gums]. AB - We have developed an analytical method for terpene resins in chewing gum. The fraction including terpene resins was prepared by means of hexane extraction and two silica gel column chromatography treatments (hexane and ethyl acetate) from chewing gum. The terpene resin fraction was analyzed with LC/MS and IR. The terpene resins are mixtures of polymeric pinene and/or limonene, which have a monomer molecular weight of 136. The MS spectrum of the terpene resin peak on the LC/MS total ion chromatogram showed protonated molecular ion (M + H)+ peaks at intervals of m/z 136, characteristic of a complex mixture of polyterpenes. IR spectroscopy is a suitable technique to identify the terpene resin type, ie., pinene or limonene. When the method was applied to imported chewing gum sold in Japan, terpene resins were clearly detected. PMID- 15272603 TI - [The toxification of juvenile cultured kusafugu Takifugu niphobles by oral administration of crystalline tetrodotoxin]. AB - Non-toxic cultured juvenile kusafugu were fed with diet containing crystalline tetrodotoxin (TTX) for 30 days and then fed with non-toxic diet for 170 days. During this period, 5 fish were sampled and the toxicity of each tissue was determined periodically. The amount of total accumulated toxin in the fish was 90 microg, representing 50% of the administered TTX (180 microg/fish) at the 60th day. It decreased to 54 microg (30%) at the 80th day and then remained unchanged up to the 200th day. The amount of toxin in the liver amounted to 40 microg (45% of total accumulated toxin) at the 30th day and gradually decreased to 5 microg (10%) at the 200th day. The toxin amount in the skin reached the highest level with 30 microg (30%) at the 50th day and then remained unchanged during the experimental period. The testes had almost no toxicity. Although the ovaries were immature, the toxin amounts increased as the weight of the tissues increased. With administration of crystalline TTX, all kusafugu used in the experiment became toxic and retained the toxin at the level of 30% of the administered toxin for about 5 months thereafter, while being fed with non-toxic diet. PMID- 15272604 TI - Mechanism of the decrease of tetrodotoxin activity in modified seawater medium. AB - This study was designed to clarify the mechanism of the decrease of tetrodotoxin (TTX) toxicity during storage in a modified seawater medium (MSWM). When TTX was added to sterilized MSWM, the toxicity of TTX in the medium markedly decreased within 1 day, as determined by a mouse bioassay. HPLC (high-performance liquid chromatography) analysis showed that the peak of TTX was reduced and new unidentified peaks were observed. Omission of the P-1 metal solution from MSWM suppressed the decrease in TTX toxicity and the disappearance of TTX. Further studies indicated that boric acid in the P-1 metal solution triggers this toxicity decrease, indicating that TTX is chemically, not microbiologically, converted to unknown compounds in MSWM. PMID- 15272605 TI - [Simultaneous determination of stevioside, rebaudioside A and glycyrrhizic acid in foods by HPLC]. AB - A method for the simultaneous determination of stevioside (Stev), rebaudioside A (RebA) and glycyrrhizic acid (GA) in foods was developed. These sweeteners were extracted from foods, except for dried fishes and shellfishes, by dialysis against Tris-HCl buffer (pH 9.0). Dried fishes and shellfishes were extracted with Tris-HCl buffer--methanol (2:8). The extracts were cleaned up with an Oasis MAX cartridge. The cartridge was washed with 0.05 mol/L sodium acetate (pH 4.0)- methanol (19:1), and the three sweeteners were eluted with 0.1 mol/L phosphoric acid--acetonitrile (1:1). Stev, RebA and GA in the eluate were chromatographed on a Develosil RPAQUEOUS-AR-5 (4.6 mm i.d. x 250 mm) column with 0.02 mol/L phosphoric acid-acetonitrile--methanol (90:55:5) as a mobile phase and monitored at 210 nm for Stev and RebA, and at 254 nm for GA. The recoveries of Stev, RebA and GA from 8 kinds of foods spiked at the level of 0.1 g/kg were 81.7-101%, 81.5 100% and 78.6-95.0%, respectively. The determination limits were 0.01 g/kg in samples. PMID- 15272606 TI - [Investigation of pesticide residues in foods distributed in Kitakyushu City]. AB - We investigated 160 kinds of pesticide residues in 715 samples of 116 kinds of foods distributed in Kitakyushu city. Sixty kinds of pesticides were detected in 55 kinds of foods (204 samples) in the range of 0.002-22 mg/kg. Five kinds of pesticides in 7 samples violated the residue standards and the indication of "unused". The detection ratios of unregulated pesticide in domestic and imported foods were 27.8 and 33.0%, respectively. Iprodione, dicofol, diethofencarb, procymidone and chlorfenapyr (for domestic food) and total bromine, benomyl, chlorpyrifos, dicofol, fenvalerate, cypermethrin and dimethoate (for imported food) showed relatively high detection ratios. Chinese cabbage, garland chrysanthemum, tomatoes and green teas (domestic) and broccoli, bananas, grapefruit, lemons, oranges, frozen edamame and frozen kidney beans (imported) showed high relative pesticide detection ratios. Residual pesticides were detected with relatively high frequency in imported fruits, imported frozen foods and imported processed foods. PMID- 15272607 TI - Application of ion-trap LC/MS/MS for determination of acrylamide in processed foods. AB - Ion-trap LC/MS/MS was evaluated for use in the determination of acrylamide (AA) in processed foods. Multiple reaction monitoring (MRM) analysis of a series of AA standard solutions containing deuterium-labeled acrylamide (AA-d3) as an internal standard was performed. A linear relationship between the concentration of AA and the ratio of peak area (AA/AA-d3) in the extracted ion chromatogram (m/z 55, 58 derived from m/z 72, 75, respectively) was obtained over a wide range of 2-20,000 ng/mL. The quantification limit of AA was 2 ng/mL. In analyses of 37 commercial foods, AA was detected in a potato snack at the maximum value of 3,570 ng/g and found in 23 foods prepared or cooked at high temperature. The samples were analyzed in triplicate and the relative standard deviations (RSD) were less than 15% in many processed foods. PMID- 15272608 TI - [Inhibitory effect of zinc on beta-hexosaminidase release from RBL-2H3 cells by synthetic chemicals]. AB - The effects of foods and chemicals related to food hygiene on degranulation were evaluated using a method for assaying the enzyme activity of beta-hexosaminidase as an index of chemical mediator release from RBL-2H3 cells in vitro. Using a previously developed assay system, we had found a large number of inhibitors and promoters of degranulation of RBL-2H3 cells. In the present study, we examined the inhibitory effect of zinc chloride on the degranulation (beta-hexosaminidase release) from RBL-2H3 cells with or without antigen in the presence of the degranulation-promotive chemicals, namely, 4 food additives, 7 pesticides and 2 veterinary drugs. These promotive chemicals were classified into two types on the basis of inhibitory profile by zinc chloride: 1) those which showed marked degranulation-inhibitory action when the cells were stimulated with antigen, such as butylhydroxyanisole, dibutylhydroxytoluene, EPN, cis- and trans-permethrin, prothiofos, pyridaben, terbufos, 2) those which showed marked degranulation inhibitory action whether the cells were stimulated with antigen or not, such as butyl p-hydroxybenzoate, o-phenylphenol, bitertanol, salinomycin. In conclusion, zinc had a dramatic inhibitory effect on enhanced degranulation induced by synthetic chemicals in vitro. PMID- 15272609 TI - [Association between progress in muscular dystrophy research and my study]. PMID- 15272610 TI - [Development of visual function: a neurophysiological point of view]. AB - Recent findings on the development of visual function in children are summarized. First, visual evoked potentials (VEPs) in normal school children, following two types of visual stimuli (pattern reversal and light emitting diode stimulation) by transient and steady-state stimulation, are presented. Reproducible VEPs with the 4 stimulation conditions can be obtained. Transient and steady-state methods provide complementary results. Second, mechanisms for photosensitive epilepsy (PSE) are discussed. We recorded flicker VEPs to different color combinations and quantified the effects of changes in color and luminance combinations. Two amplitude peaks (9 and 18 Hz) were observed for all kinds of isoluminant color combination stimuli against temporal frequency. In addition, this characteristic was modulated by luminance. Normal children showed much higher sensitivity to contrast changes and color combination compared with normal young adults, which may be responsible for PSE in childhood. Third, cognitive function for facial expression of normal children and adults is mentioned. For Chernoff's faces showing anger and sadness produced by computer, children showed higher scores compared with adults, suggesting higher sensitivity for facial expression. Knowledge of developmental changes in children allows us to understand the maturational and degenerative changes of the central nervous system. PMID- 15272611 TI - [A questionnaire study on psychological problems in home medical care for children with chronic illness]. AB - We performed a questionnaire study on psychological problems in home medical care, including mechanical ventilation, oxygen therapy, continuous ambulatory peritoneal dialysis and parenteral nutrition, for children with chronic illnesses in Osaka prefecture, Japan. One hundred two pediatricians (46%) answered the questionnaire. The majority of the doctors regarded psychological support for the children and their families as an important issue. In some cases, home medical care was interrupted because of psychological problems such as psychological burden, anxiety and stress of the children and/or their families. And some other cases, home medical care was impossible because of parental refusal to treatment and/or their child. The cases with mechanical ventilation and oxygen therapy mainly accounted for these cases. Since the number of cases undergoing home medical care is estimated to increase in the future, medical staff should be more aware of psychological support for the families as well as their children. PMID- 15272612 TI - [Efficacy of secretin for the treatment of autism]. AB - We administered secretin intravenously to 14 patients with autism (9 to 14 years, 10 males; 4 females), and evaluated its clinical effect. We also measured cerebrospinal fluid (CSF) levels of homovanillic acid (HVA) and 5-hydroxy-indole 3-acetic acid (5HIAA) before and after 4 weeks treatment, and compared them with the grade of improvement of the clinical symptoms assessed by the scores of Autism Diagnostic Interview-Revised (ADI-R). After injection of secretin, the ADI R score increased in 8 patients, but declined in 3. Improvement was observed in functions such as sociability (interpersonal relationships), communication and speech improved, whereas in the others. symptoms such as hyperkinesias and stereotyped behavior became worse. The CSF levels of HVA was significantly increased in all of the patients showing an improvement in the ADI-R score. SHIAA levels also tended to increase, although this increase was not significant. These findings suggest that secretin promotes the metabolism of serotonin and dopamine in the central nervous system, which may contribute to improvement in clinical symptoms of autism. PMID- 15272613 TI - [Visual perception of Kanji characters and complicated figures. Part 3. Visual P300 event-related potentials in patients with attention deficit/hyperactivity disorders]. AB - In order to evaluate visual perception, the P300 event-related potentials (ERPs) for visual oddball tasks were recorded in 11 patients with attention deficit/hyperactivity disorders (AD/HD), 12 with mental retardation (MR) and 14 age-matched healthy controls. With the aim of revealing trial-to-trial variabilities which are neglected by investigating averaged ERPs, single sweep P300s (ss-P300s) were assessed in addition to averaged P300. There were no significant differences of averaged P300 latency and amplitude between controls and AD/HD patients. AD/HD patients showed an increased variability in the amplitude of ss-P300s, while MR patient showed an increased variability in latency. These findings suggest that in AD/HD patients general attention is impaired to a larger extent than selective attention and visual perception. PMID- 15272614 TI - [Etiology and bronchofiberscopic treatment of pulmonary atelectasis in patients with severe motor and intellectual disabilities]. AB - Atelectasis is a common respiratory complication in patients with severe motor and intellectual disabilities. We encountered 3 patients with irreversible atelectasis due to delayed therapy, emphasizing the necessity of performing flexible bronchofiberscopy. A total of 21 patients with atelectasis were studied to assess the etiology and efficacy of bronchofiberscopy. The underlying condition was bronchitis/pneumonia in 19 cases, tracheal hemorrhage in 1, and lung cancer in 1. Most of the patients had predisposing factors, such as a bedridden status in 90% and a weak or absent cough reflex in 81%. It was statistically suggested that atelectasis is likely to occur on the side contralateral to thoracic scoliosis. Among the 18 patients who underwent bronchofiberscopy within 2 weeks after the diagnosis of atelectasis, 16 (89%) showed recanalization and resultant improvement of respiratory failure. Bronchofiberscopy is useful for treating atelectasis in patients with severe motor and intellectual disabilities. PMID- 15272615 TI - [Abnormal findings of dichotic listening test in patients with childhood adrenoleukodystrophy]. AB - Ten Japanese boys with childhood-onset adrenoleukoqdystrophy (ALD) were evaluated with dichotic listening test (DLT). Six cases showed abnormal findings especially of laterality index (L.I.) calculated from the score of each ear. Some of them showed no abnormal findings with other auditory examinations containing auditory brainstem responses (ABR). One patient showed abnormal L.I. of DLT at an early stage. The abnormality of laterality index was similar to the so-called "strong left-ear suppression" in patients who underwent callosotomy. Although all of these six patients had a high signal lesion at the splenium of the corpus callosum in a T3-weighted MRI sequence, it was difficult to evaluate the width of demyelinated area. DLT could detect the early damage of connecting fibers mediating inter-hemispheric transfer of auditory information, and might be a useful method for evaluating the cerebral function of auditory processing in patients with childhood ALD. PMID- 15272616 TI - [Survey of vaccination and viral infections for children with severe myoclonic epilepsy in infancy]. AB - A study group for establishment of a proposed immunization program for neurologically high risk children (Chief, Kihei Maekawa) sponsored by the Ministry of Health, Labour and Welfare of Japan is preparing a proposal for patients with epilepsy. Severe myoclonic eplepsy in infancy (SMEI) is an intractable epilepsy which often presents with status epilepticus and triggered by hyperthermia and viral infections. In this study we investigated the history of vaccination in children with SMEI to compare the risk of vaccination with that of natural contraction of infection. Fifty-eight patients with SMEI, aged from 2 to 25 years, were enrolled in this study. A total of 359 vaccines were given to these subjects. The vaccination rates were high for BCG (71%) and polio (1st; 71%, 2nd; 53%), and then fell gradually after the onset of SMEI (DPT-1st; 48%. DPT-2nd; 45%, DPT-3rd; 38%, DPT-4th; 24%, mumps; 21%, varicella; 19%, rubella 31%). However, the measles vaccine was given at a relatively high rate (55%) before the age of three. When patients suffered from measles, rubella, mumps or influenza, they had a high risk of severe neurological complications, such as convulsive status, disturbance of consciousness and encephalopathy. These complications were documented in 63% of all episodes of naturally contracted infections. This rate was significantly higher (p < 0.0001) than that associated with vaccination (7.2%). However, hyperthermia and convulsion developed more frequently (p = 0.012) after measles vaccine was given, as compared to other vaccines. Thus, administration of these vaccines to patients with SMEI in conjunction with other preventive measures against seizure induced by hyperthermia, may meet the needs of their parents. PMID- 15272617 TI - [Case of methylmalonic acidemia presenting clinically Leigh encephalopathy]. AB - A 1-year-old boy with methylmalonic acidemia had symmetrical lesions of the bilateral basal ganglia, which suggested Leigh encephalopathy. The findings on brain magnetic resonance imaging (MRI) and his physical condition greatly improved by the intravenous administration of vitamin B1. We hypothesized that in this case, clinical Leigh encephalopathy was caused by a impairment of the activity of pyruvate carboxylase induced by the accumulation of methylmalonyl CoA and an impairment of energy production due to a lack of vitamin B1, especially impairment of the activity of pyruvate dehydrogenase complex during an acute worsening of methylmalonic acidemia. Thus, in the treatment of methylmalonic acidemia, attention should be paid to vitamin B1 deficiency. During an acute worsening, vitamin B1 should be administered by intravenous drip injection. PMID- 15272618 TI - [A case of infant with paroxysmal downgaze]. AB - We reported a girl showing paroxysmal downgaze as the only manifestation of neurological abnormality. The movement was first noted at the age of 75 postnatal days, increased in frequency gradually to 10 weeks, and decreasing gradually, and disappeared at 35 weeks. The eye movements comprised first rapid downward and floating and rapid upward to primary position. The phenomenon was only provoked by looking at her own finger in front of her at a distance of 5 cm, and looked like a overshoot of vertical saccade. The suppression of the saccade neurons in the superior colliculus by inhibitory systems from the frontal association cortex through the direct pathway of the basal ganglia was also immature in the baby. PMID- 15272619 TI - [Magnetic resonance findings in a case of Gradenigo syndrome: widespread inflammation involving the paranasal sinuses and middle ear]. AB - Gradenigo syndrome is a rare condition consisting of otitis media, trigeminal neuralgia and abducens palsy. We report here a 6-year-old girl with this syndrome. Cranial magnetic resonance imaging (MRI) demonstrated inflammatory lesions in the left petrous apex and cavernous sinus, as well as stenosis of the left carotid artery. She was conservatively treated with antibiotics, which successfully improved her clinical and MRI findings. Surgical treatment including mastoidectomy was avoided. This case illustrates the usefulness of MRI in the diagnosis and management of Gradenigo syndrome. PMID- 15272620 TI - [Reevaluation of serum isocitrate dehydrogenase concentration as liver function marker in patients with Duchenne muscular dystrophy]. PMID- 15272621 TI - [Duchenne muscular dystrophy born prematurely showed normal CK value until 35 weeks of gestation age]. PMID- 15272622 TI - Covering all bases: early thoughts for population bases for the 2011 Census. AB - This article discusses early thinking on the options for the population bases to be used in the 2011 Census. It describes some of the work being conducted to enable a decision about bases to be reached, including historic and international reviews of censuses. It also discusses the implications that our rapidly changing society has for census planning. PMID- 15272623 TI - Small area population estimates project: data quality of administrative datasets. AB - The Office for National Statistics (ONS) has set up a project to investigate the feasibility of producing postcensal small area population estimates on a nationally consistent basis for England and Wales. Research has taken place to identify datasets that could potentially be used within a method to produce small area population estimates. Following an evaluation of a number of different administrative datasets, the most suitable have been short-listed for further consideration. This article presents the findings of the evaluation, based on 2001 data, and summarises the characteristics of these short-listed data sources. This article does not cover the methods that are being evaluated as part of the feasibility assessment. PMID- 15272624 TI - Immigration, emigration and the ageing of the overseas-born population in the United Kingdom. AB - This article uses data from the 1971 and 2001 Censuses, the 1999-2003 Labour Force Survey and the 1977 to 2002 International Passenger Survey to investigate the migration processes contributing to the age structure and ageing of the UK's overseas-born population. Overall almost half of recent decades' immigrants to the UK emigrate again within five years of arrival, but with large variation by overseas country of birth. Between half and two thirds of the immigrants born in the continental European Union, North America and Oceania emigrate again within five years, while 15 per cent of those born in the Indian subcontinent do so. Significant cumulative emigration more than five years after arrival is seen among earlier immigrants from the Indian subcontinent, the Caribbean Commonwealth and Europe. Large country-of-origin variations in the ratio of pension-age population to working-age population primarily reflect the country composition of immigration streams 30 or more years before. PMID- 15272625 TI - [Corpus callosum, interhemispheric interaction and brain right hemisphere function]. AB - A complex clinico-neuropsychological study, using A.R. Luria's method, has been conducted before and after surgery in 36 patients with arteriovenous malformations of the corpus callosum. Symptoms of local lesions in different compartments of the corpus callosum are described. The symptoms of the partial corpus callosum lesion are shown to be modally specific, but only to a certain extent. Isolated appearance of dyscopia or dysgraphia or symptoms is possible. A combination of lesions in the medial brain areas (cingulated gyrus, frontal lobes) with corpus callosum malformation significantly augmented their dysfunction. In case of impairment in frontal parts of the corpus callosum, symptoms of frontal lobes dysfunction were observed. The corpus callosum malformations resulted in right hemisphere dysfunction in the sphere of emotions, perception and spatial activity. As it was shown earlier, right hemisphere integrates impulses from both sides of the space, being first involved in activity, realizing its initial stages. The author suggests that such a synthetic right hemisphere activity, in the presence of closed relations to "conscious" left hemisphere, is necessary for formation of the whole picture of the separate objects, as well as of certain types of activity. In this view, right hemisphere may be relatively considered as a dominant one but not vice versa. PMID- 15272626 TI - [The disorders of blood circulation and consciousness in non-psychotic and delirious alcohol withdrawal syndrome]. AB - One hundred and fifty-seven men with alcohol withdrawal syndrome, including 127 with delirium of different severity, have been studied. During delirium, sequestration of erythrocytes in the vessels and exudation of the fluid into the cellular space developed. There were marked capillary blood flow disturbances with erythrocyte aggregation, metabolic acidosis, compensated with respiratory alkalosis, total lactate dehydrogenase activity elevation, Na+ and K+ decrease in blood plasma and K+ reduction in erythrocytes in a severe form of alcohol delirium. The authors suggest that the complex of disorders observed is similar to shock process, and hemodynamic disturbances may be involved in the development of consciousness disorders. PMID- 15272627 TI - [Acute stress reactions in children and adolescents in emergency situations]. AB - Psychological and psychiatric examination, along with medico-psychological assistance, have been conducted for 48 children and adolescents, aged 9-18 years, who have been saved during the fire. An examination was carried out on day 2-7 after the disaster. A diagnosis of acute stress reaction (ASR) was made using DSM IV-TR criteria with definition of 4 groups of symptoms: recurrent trauma surviving, avoidant symptoms, elevated excitability and dissociated symptoms. Besides the clinical examination, the questionnaires adapted for the study and drawing techniques have been used. Symptoms of recurrent trauma surviving occurred most often (79.2%). Avoidant symptoms were detected in 75% of the cases, elevated excitability--in 64.4%, dissociated symptoms--in 62.5%. Symptoms of all the groups were found in 43.8% of the cases that suggested the presence of pronounced distress. No statistically significant correlation between symptom severity and subject's age and sex was revealed. The presence of siblings and friends among those been lost, affected the severity of ASR expression, in particular, avoidant symptoms and dissociation, recurrent trauma surviving symptoms being less influenced. Symptoms of excitability in ARS were shown to be more specific for the boys. PMID- 15272628 TI - [Efficacy of cytoflavin in spondylogenic radiculomyeloischemia]. AB - The efficacy of cytoflavin in the treatment of 60 patients (39 women, 21 men, age 32-64 year) with spondylogenic cervical and lumbosacral radiculomyeloischemia due to degenerative dystrophic spinal lesions was studied in a randomized double blind placebo controlled study according to GCP rules. During 10 days 40 patients received intravenous cytoflavin dropper injections once daily; 20 patients (control group) received 5% glucose solution as a placebo. Considerable improvement was observed in 70% patients with radiculomyeloischemia of cervical localization and in 65%--with radiculomyeloischemia of lumbosacral segments. In the control group, positive dynamics of neurologic symptoms was 25-30% lower. Cytoflavin significantly reduced cognitive disturbances, improved reparative processes both in the central and peripheral neurons, and may be recommended in ischemic neuronal spinal lesions. PMID- 15272629 TI - [The peculiarities of hypnotic states in patients with borderline mental disorders]. AB - Two hundreds and eighteen patients with borderline mental disorders have been examined. After the first session of hypnosis suggesting therapy, 5 clinical variants of hypnotic states (physiological, asthenic, phobic, hysterical and mixed) were distinguished. Additionally, 40 patients have been interviewed for obtaining a formalized, according to adjectives used by the patients, description of experiences during the hypnotic session. Using cluster analysis, 5 groups were formed characteristics of which strongly correlated with earlier clinical description and therapeutical efficacy. The results allow better understanding of hypnotic states specificity as well as more precise prognosis of a therapeutic effect. PMID- 15272630 TI - [Computer tests for research of the oculomotor reactions in patients with complains of dizziness]. AB - New methodological principles for a study of vestibular function, intersensory interactions, and eye tracking function are proposed by parameters of spontaneous oculomotor responses and those induced by visual and vestibular stimuli using the series of computer stimulating programs, which provided polymodal, separate and combined stimulation of the visual and vestibular inputs for the OCULOSTIM unit. The method suggested allows to objectivity the subjective complaints of dizziness and equilibrium disturbances, to differentiate diagnosis of cerebral organic process and psychogenic disease of central nervous system (CNS), and to specify physiological systems involved in their pathogenesis as well as to diagnose subclinical cerebral vestibulopathy. Qualitative and quantitative characteristics of eye tracking movements with and without a visual background (retinal optokinetic stimulation) are the most informative in the differential diagnosis of organic and functional psychogenic CNS disturbances. PMID- 15272631 TI - [Cytochemical activity of mitochondrial enzymes in Parkinson's disease]. AB - Using cytochemical computerized morphometric method, activity of the key enzymes of energetic metabolism (succinate dehydrogenase, alpha-glycerophosphate dehydrogenase, malate dehydrogenase, glutamate dehydrogenase and lactate dehydrogenase) was studied in blood lymphocytes of 75 patients with Parkinson's disease and 15 healthy controls. The signs of systemic mitochondrial insufficiency, which correlated with the disease duration and severity, were found in all the patients, including those with juvenile parkinsonism. These data may provide a basis for introducing cytochemical monitoring as well as for administration of modern "mitochondrial" drugs (yantavit, coenzyme Q10, L carnitine, etc). PMID- 15272632 TI - [Serotonin transporter gene polymorphism and factors influencing mental and physical health in aging]. AB - Genetic predisposition is thought to exert a certain influence on the indices related to longevity and quality of life. Many of the indices, namely cognitive functioning, stress resistance, metabolism control, may be related to serotonin activity. To study polymorphic serotonin transporter gene variants and their association with features relevant for survival and longevity prognosis, a sample of elderly Russians from Moscow community recruited in the project "Stress related mechanisms in Russia", comprising 196 subjects, mean age 76.2+/-5.3 years, 155 men, 41 women, has been genotyped. Allele and genotype frequencies have been estimated in 3 groups, aged 60-69, 70-79 and 80-87 years, respectively. A trend (chi2=4.1; p=0.12) to the prevalence of individuals with SS genotype (21.8%), as compared to expected level (14.6%), was found in the group of octogenarians (n=55, mean age 82.8+/-1.9 years). An association analysis between genotype and physiological traits revealed a genotype contribution to past smoking on tendency level (p=0.069), waist to hip ratio (WHR) (p=0.012) and plasma insulin concentration (p=0.02), with a higher frequency of SS genotype among non-smokers and subjects with lower WHR and insulin concentration. Genotype effect on the traits was stronger, being considered in interaction with the age above 80 years. Genotype was not associated with cognitive functioning (MMSE), but proved to be a significant predictor of MMSE performance (p=0.03) in octogenarians. The results obtained are in line with current concepts of serotonin role in smoking, obesity and cognitive functioning. PMID- 15272633 TI - [Epidemiology of headache in children and adolescent]. PMID- 15272634 TI - [Methodological aspects of psychiatric care for patients of general hospitals and outpatient clinics]. PMID- 15272635 TI - [Movalis in the treatment of acute dorsopathy]. PMID- 15272636 TI - [Signopam treatment of insomnia in patients with somatic pathology]. PMID- 15272637 TI - [Salt regimens modulation of neuroprotective effects of Magne B6 on the model of global cerebral stroke]. PMID- 15272638 TI - [Patent foramen ovale: a role in pathogenesis of some neurological diseases and methods of their treatment]. PMID- 15272639 TI - [Primary headache in children]. PMID- 15272640 TI - [Some problem of art-therapy in psychiatry]. PMID- 15272641 TI - Pustular impetigo with good response to clarithromycin. AB - Impetigo is a contagious superficial pyogenic infection of the skin caused by Staphylococcus aureus and/or by group A Streptococcus. Two main clinical forms are recognized: bullous impetigo and non-bullous impetigo. We present an unusual case of pustular impetigo in a 35-year-old man. The pustules were localized symmetrically in the groin and the patient was successfully treated with clarithromycin. In bullous impetigo, exfoliative toxins produced by Staphylococcus aureus are accepted as the basis for the bulla formation just below the stratum granulosum. Although clarithromycin is considered to be a second-choice therapy for bullous impetigo, it was highly effective in our case. PMID- 15272642 TI - Increased fibrin specificity and reduced paradoxical thrombin activation of the combined thrombolytic regimen with reteplase and abciximab versus standard reteplase thrombolysis. AB - In patients with acute myocardial infarction treated with thrombolytics, platelet activation as well as alterations of the hemostatic and fibrinolytic systems have been described favoring early infarct-related artery reocclusion. We investigated the effects of a newer thrombolytic regimen with half-dose double-bolus reteplase (2 x 5 IU, 20 patients) combined with abciximab versus full-dose reteplase (2 x 10 IU, 18 patients) on the fibrinolytic and the hemostatic system in patients with acute ST-segment elevation (in the electrocardiogram) myocardial infarction. The thrombolytic regimen with half-dose reteplase in combination with abciximab caused in vivo a lower systemic plasminemia and a lower paradoxical activation of the contact phase of the coagulation system (measured as activated factor XII); a lower paradoxical thrombin activation/generation; and a lesser extent of fibrinogen breakdown compared with the reteplase regimen. These results could be, at least in part, a possible explanation for the observed significantly lower rates of reinfarction until 7 days after enrollment and of recurrent ischemia in the combination group in the Global Use of Strategies to Open Occluded Coronary Arteries V (GUSTO V) trial. PMID- 15272643 TI - Effect of angiotensin-converting enzyme inhibitor on collagenolytic enzyme activity in patients with acute myocardial infarction. AB - Matrix metalloproteinases and their tissue inhibitors are key enzymes degrading myocardial collagen in acute myocardial infarction (AMI). The aim of the present study was to determine whether angiotensin-converting enzyme inhibitors (ACEI) influence collagenase-1 (MMP-1) and their tissue inhibitor (TIMP-1) activity in AMI patients. Plasma levels of MMP-1, TIMP-1 and MMP-1/TIMP-1 complex were measured in 24 patients (aged 58.4 +/- 13.9 years) with AMI. Thirteen patients received perindopril 4 mg/day (group A) and 11 did not (group B). Plasma samples collected on admission and at 0, 3, 6, 9, 12, 18, 24, 36 and 48 hours and on days 3, 4, 5, 7, 15 and 30 thereafter were analyzed by relevant ELISA kits. Ejection fraction (EF) was assessed by ventriculography and end-diastolic diameter (EDD) echo-study on days 6 and 30. Values of collagenolytic enzymes of group A compared with those in group B were on average lower by 34%, 18.3% and 40%, respectively. The difference in values between groups at 0 h, 3 h and 9 h was significant (p < 0.048). ANOVA repeated measurement analysis showed significance within subjects for MMP-1 alone (p < 0.043) and for MMP-1 and ACEI (p < 0.046), while for TIMP-1 and MMP-1/TIMP-1 complex significance was only p < 0.0009. Regarding EDD changes, patients in group A showed minimal or no changes (51.23 +/- 1.8 mm to 51.6 +/- 2.13 mm), their EF was 38.8% and infarct size was medium to large. In contrast, group B showed a trend to increase EDD (41 +/- 0.78 mm to 42.33 +/- 0.59 mm), their EF was 50.5% and infarct size was small to medium. In conclusion, early initiation of ACEI treatment reduces collagenolytic activity. This effect may be considered an alternative mechanism for beneficial effects on postinfarction remodeling. PMID- 15272644 TI - Antalgic effect and clinical tolerability of hyaluronic acid in patients with degenerative diseases of knee cartilage: an outpatient treatment survey. AB - A total of 40 outpatients (28 men and 12 women) aged between 18 and 82 years with primary or secondary symptomatic knee osteoarthritis (OA) were selected for this retrospective study. The patients were treated weekly with an intra-articular injection of hyaluronic acid of biofermentative origin. A total of five injections were given, with a follow-up visit at week 7. The aims of this study were to analyze the antalgic effect and tolerability of the procedure, evaluated by overall tolerance, Lequesne's Algo-Functional Index (AFI), pain level evolution and analgesic consumption. No systemic adverse effects related with the device were reported. Global tolerability was judged as excellent/good by almost all the patients and the investigator; 16 patients reported a mild burning sensation at the injection site, which was more frequent during the first injection and resolved within a few minutes. The mean value of the AFI decreased from 7.9 at the initial visit to 3.2 at the final visit, parallel to a decrease into the relative scores of the pain scale. We can thus conclude that intra articular injection of hyaluronic acid of biofermentative origin appears to be a safe and effective therapy for gonarthritic pain. PMID- 15272645 TI - Effects of chronic administration of D-003, a mixture of sugar cane wax high molecular acids, in beagle dogs. AB - D-003 is a mixture of high molecular weight aliphatic primary acids purified from sugar cane wax (Saccharum officinarum, L) with cholesterol-lowering and antiplatelet effects. Previous studies, including a 6-month study conducted in rats, have shown no D-003-related toxicity. The present study was undertaken to investigate the effects of D-003 orally administered for 9 months in beagle dogs. The animals were randomly distributed in three groups: a control group receiving the vehicle only and two groups orally administered D-003 (200 and 400 mg/kg). Body weight gain, food consumption and clinical signs were controlled throughout the study. The effects of D-003 on collagen-induced platelet aggregation, bleeding time (BT) and coagulation parameters (prothrombin time and kaolin activated thromboplastin time) were also investigated. Most blood biochemistry and hematological parameters were assessed at baseline and after 6 and 9 months of treatment, while total cholesterol (TC), triglycerides, platelet aggregation, BT and coagulation parameters were determined at baseline and after 9 months of treatment. At study completion, the animals were sacrificed. D-003 at a dose of 200 and 400 mg/kg significantly reduced TC (p < 0.05), significantly inhibited platelet aggregation and increased BT compared with levels in controls. Data analyses of body weight gain, food consumption, clinical observations, the remaining blood biochemistry and hematology indicators (including coagulation parameters, organ weight ratios and histopathological findings) showed no trends with D-003 doses or significant differences between control animals and treated groups. In conclusion, D-003 administered for 9 months to beagle dogs induced the expected effects with no evidence of drug-related toxicity. PMID- 15272646 TI - Sweet schizophrenia. PMID- 15272647 TI - Medicare eventually may cover dental costs--but what about the children? AB - Government programs, particularly Medicare, provide extremely limited funds for dental services. As a result, the government absorbs fewer dental costs than the costs for other health services. Despite the need for support of dental services for children, it may become politically expedient to support the expansion of Medicare to include dental care for the increasing numbers of dentate baby boomers. The economic case for Medicare dentistry is presented as a harbinger for continued underfunding by the federal government for dental care for children. PMID- 15272648 TI - Where's the best place to be born? AB - A series of large city rankings developed from federal agency reports were reviewed to increase appreciation of the factors that impact the general health of newborns. Efforts to provide the "right start" for the beginning of life may decrease the incidence of children with developmental disabilities. PMID- 15272649 TI - Oral trauma in an urban emergency department. AB - The purpose of this investigation was to determine the type of traumatic orofacial injuries and the referral pattern seen in children up to 15 years of age who came to the pediatric emergency department at an urban hospital during an 18-month period. The majority of injuries treated, 87% in preschool children and 71% in children ages 6 to 15 years old, were lacerations and/or abrasions. The primary mechanism of injury was falls, accounting for 78% of traumatic orofacial injuries in preschoolers and 47% in children ages 6 to 15 years. Children ages 6 to 15 years were more likely to be injured in sports-related activities and more likely to injure the dentition than preschool children. When referrals for follow up care were documented, the majority was to the department of dentistry. PMID- 15272650 TI - Parent satisfaction with emergency dental treatment at a children's hospital. AB - PURPOSE: This study evaluated parental satisfaction with emergency dental treatment. METHODS: One hundred twenty-two parents of children requiring emergency extraction of 1 or more primary teeth completed a survey designed to test the effect of provider, treatment, and demographic variables on parental satisfaction. RESULTS: Most parents (>80%) indicated satisfaction with the treatment provided. Parents were most satisfied with treatment during clinic hours, treatment provided by an attending pediatric dentist, and treatment provided by male dentists. Satisfaction was correlated with the clarity of the provider explanation. Explanations by male dentists were perceived most positively. Parents of children receiving molar extraction(s) were more satisfied than parents of children with incisor extraction(s). Satisfaction did not correlate with ethnicity of the parent or patient, parent education level, funding sources, or use of an immobilization device. Parents preferred sedation for behavior management of the emergency patient. CONCLUSIONS: To address the expectations and concerns of parents, dental professionals need to be attentive to the quality of dentist-parent communication and parental expectations during emergency services. PMID- 15272651 TI - Systemic and local teething disturbances: prevalence in a clinic for infants. AB - The aim of this study was to present data as reported by parents on primary tooth eruption and the occurrence of local and systemic manifestations in children ages 0 to 3 years seen at the baby clinic of the Aracatuba Dental School, UNESP, Sao Paulo, Brazil. Data from 1,813 records were analyzed, yielding 1,165 records suitable for review. Some type of local and/or systemic manifestation during primary tooth eruption was reported for 95% (1,129) children studied. The predominant manifestation was gingival irritation (85%), while the least frequent symptom (26%) was a runny nose. PMID- 15272652 TI - Prevalence of periodontal disease in the primary dentition of children with cerebral palsy. AB - PURPOSE: The aim of this study was to evaluate periodontal disease prevalence in the primary dentition of children with cerebral palsy (CP). METHODS: The experimental group consisted of 100 children with CP and the control group, 100 healthy children. The indices used in the oral examination were: debris index, calculus index, gingival index, Simplified Oral Hygiene Index (OHI-S), and gingival hyperplasia index. RESULTS: It was observed that the mean values for the debris index, OHI-S, and gingival index were higher in the children with CP than in the control group. In the CP group, the percentage of children who had their teeth brushed by their parents or another person was greater than in the control group. CONCLUSIONS: From this study, it was concluded that children with CP had greater prevalence of periodontal disease in the primary dentition than children in the control group. PMID- 15272653 TI - Salivary gland disease in pediatric HIV patients: an update. AB - Oral manifestations are one of the earliest clinical indicators of HIV infection and progression in children. Prompt recognition of these signs and symptoms by dental providers can help in the diagnosis and intervention of delaying the progression of HIV disease to AIDS. Salivary gland disease is a common manifestation of HIV infection in pediatric patients, presenting either as gland enlargement and/or xerostomia. The parotid glands by far are most frequently affected, though the other major glands are commonly involved. Diseases of the salivary glands and the corresponding quantitative changes in saliva affect the homeostasis of the oral cavity and account for significant morbidity during the progression of HIV disease. This paper summarizes the research on HIV-related salivary gland disease and outlines treatment and management considerations. PMID- 15272654 TI - Comparison of submandibular/sublingual salivary flow rates in children and adolescents. AB - PURPOSE: This study compared age related changes in submandibular/sublingual salivary flow rates in children and adolescents. METHODS: Twenty-nine children between the ages of 5 and 14 years were evaluated. A group of adults ages 18 to 39 years were also evaluated. No subject was taking medication which could have affected salivary flow rate. Salivary flow was stimulated using a single application of 3% citric acid during a 5-minute test period. A group of 12 children ages 5 to 9 years was compared to a group of 16 adolescents ages 10 to 14 years. RESULTS: Analysis using a Wilcoxon Rank Sum Test with a .05 level used for significance showed that the salivary flow rates of the 2 groups were not significantly different. CONCLUSIONS: No significant difference was found between the stimulated submandibular salivary flow rates of children ages 5 to 9 years and 10 to 14 years. PMID- 15272656 TI - Comparison of rotary and manual instrumentation techniques on cleaning capacity and instrumentation time in deciduous molars. AB - PURPOSE: The aim of this study was to evaluate, in vitro, the cleaning capacity and time needed for instrumentation of root canals of deciduous molars by manual and rotary instrumentation. METHODS: Thirty-three deciduous molar root canals were injected with India ink and divided into 3 groups: group I--the root canal instrumented manually with K files; group II--the root canal instrumented with rotary Profile .04 instruments; group III--control group, (ie, root canals not instrumented). Instrumentation time was recorded. The teeth were cleared and the removal of India ink was measured in the cervical, middle, and apical thirds. RESULTS: There was no significant difference for cleaning capacity between manual and rotary techniques in the 3 root thirds (P>.05), but both techniques were different from the control group (P<.001). Significantly less time was needed for instrumentation with the rotary technique (3.46 minutes) than with the manual technique (9.06 minutes). CONCLUSIONS: Although no differences were found for cleaning capacity, the reduction of instrumentation time by the rotary technique was a relevant clinical factor for endodontic treatment. PMID- 15272655 TI - Penetration and microleakage of dental sealants in artificial fissures. AB - This study investigated sealant penetration and dye microleakage of a resin composite system, a compomer system, and a resin-modified glass-ionomer cement in artificially grooved fissures in human molars. Ionosit Seal penetrated 99% of the artificial crevices, whereas Dyract Seal penetrated 97%. The penetration of Helioseal F at 90% was statistically different (P<.0001) from the other 2 materials. Microleakage dye penetration occurred in 22% of the Dyract Seal samples, while it occurred in 5% of Healioseal F and 7% of Ionosit Seal samples. The viscosity and flow properties of the 3 sealants allowed the materials to penetrate the artificial grooves, but they did not seem to affect their sealing capacity. PMID- 15272657 TI - Positional changes of the upper canine and posterior teeth, hard palate, and sinus floor from primary to permanent dentition. AB - This cross-sectional study investigated normal positional changes of the upper permanent canine and posterior teeth, hard palate, and sinus floor in normal Taiwanese children from the deciduous to early permanent dentition. In total, 261 panoramic radiographs were used. During the observation period, almost all structures changed their positions toward the distal and occlusal direction. The vertical positional changes of crowns of all teeth were greater than those of the root apices through all developmental stages. There were small positional changes in the mesial surfaces of the crowns of the upper buccal teeth until their roots had formed. There were continuous positional changes in the crowns and root apices of the permanent molars during tooth development and eruption. There were no significant changes in tooth inclination for any of the buccal teeth, whereas there were conspicuous changes in tooth inclination for the permanent molars. The floor of the maxillary sinus remarkably changed its position in an occlusal direction during the active eruption period of the first molar and buccal teeth. PMID- 15272658 TI - Evaluation of the bonding mechanism of an adhesive material to primary teeth. AB - PURPOSE: The aim of this study was to evaluate with scanning electron microscopy (SEM) the bonding mechanism of the one-bottle adhesive Prime&Bond NT (Dentsply, PBNT) to enamel and dentin of deciduous teeth, following different methods of substrate treatment. METHODS: Eighteen extracted posterior deciduous teeth were randomly divided into 6 groups (N=3). The experimental groups differed for substrate and method of substrate conditioning prior to bonding with PBNT. Group 1: 36% phosphoric acid (PA)/PBNT on dentin; group 2: PA/PBNT on enamel; group 3: non-rinsing conditioner (NRC) (Dentsply)/PBNT on dentin; group 4: NRC/PBNT on enamel; group 5: PBNT on dentin without any previous conditioning; group 6: PBNT on enamel without any previous conditioning. On all the specimens, following the application of the adhesive solution, Dyract AP was layered on top and light cured. The bonded specimens were processed for SEM observations. RESULTS: When used in combination with 36% phosphoric acid, PBNT was able to form a hybrid layer with resin tags on both enamel and dentin. Following conditioning with NRC, a thinner hybrid layer with shorter resin tags was developed on dentin; on enamel an etching pattern was still detectable. When only PBNT was applied without any previous conditioning, on dentin neither hybrid layer nor resin tags were visible; no sign of micromechanical bonding could be seen on the untreated enamel. CONCLUSIONS: The bonding mechanism of the one-bottle adhesive Prime&Bond NT on enamel and dentin of deciduous teeth is effective only following substrate conditioning with 36% phosphoric acid or NRC. PMID- 15272659 TI - Effectiveness of chlorhexidine-thymol varnish in preventing caries lesions in primary molars. AB - PURPOSE: The objective of this study was to evaluate the effect of chlorhexidine thymol varnish on the prevention of caries lesions in primary molars among schoolchildren ages 6 to 7 in relation to their previous experience with caries. METHODS: Two groups of schoolchildren of lower-middle socioeconomic level were followed up in a clinical trial: one group of 86 children, treated with a chlorhexidine-thymol varnish (Cervitec) and another group of 95 children who served as controls. The varnish was reapplied every 3 months, and the caries lesion increments were compared at 24 months. RESULTS: There was no statistically significant difference between these 2 groups in the increment in decayed and filled primary molars. The children in the varnish group with no decayed or filled primary teeth at baseline showed a significantly lower (P<.05) incidence of caries lesions in primary molars (at 24 months) compared with the control group. CONCLUSIONS: Chlorhexidine-thymol varnish can be said to reduce caries lesions in the primary molars of schoolchildren ages 6 to 7 with no previous caries lesion experience. PMID- 15272660 TI - Glass-fiber reinforced composite in management of avulsed central incisor: a case report. AB - Reimplantation failure of avulsed anterior tooth in an adolescent patient requires removal of the failed tooth and consideration of restorative options. These options may include a removable partial denture, conventional 3-unit fixed partial denture, implant, or a resin-bonded appliance with a metal substructure (Maryland bridge). The glass-fiber reinforced composite material (everSTICK, StickTech Ltd, Turku, Finland) offers a restorative solution that is conservative and esthetic when compared to other restorations. Advantages include reduction of cost compared to conventional bridges, saving of time, elimination of second visit, ease of application, absence of metal allergy, ease of cleaning, and naturalness of feel. Its limitations include occlusal factors, and the presence of unsuitable abutment teeth. Another traditional contraindication is the presence of diastemas, which may limit the potential esthetic gains. This case of an 11-year-old girl, addresses the indications, preparation guidelines, and restorative procedures for an everSTICK bridge. PMID- 15272661 TI - Dental fusion and dens evaginatus in the permanent dentition: literature review and clinical case report with conservative treatment. AB - This clinical case report describes concomitant developmental disorders in the permanent dentition. The concurrence of bilateral dens evaginatus and dental fusion is a rare dental anomaly. Dental fusion is characterized by the union of 2 adjacent teeth at the crown level, which produces the formation of an enlarged clinical crown. Dens evaginatus is an enamel elevation similar to a cusp, generally located in the main groove of molars and premolars. The etiology of both anomalies is uncertain. The purpose of this paper was to review the related literature and present a clinical case where both anomalies are present. PMID- 15272662 TI - Odontomas in the primary dentition: literature review and case report. AB - Odontomas are hamartomatous developmental malformations of dental tissues. They are very rarely diagnosed in the primary dentition. The purpose of this paper was to review the literature in relation to odontomas in the primary dentition, and describe the dental management of a child, age 4 years and 8 months, who had a complex odontoma. PMID- 15272663 TI - Unerupted mandibular second primary molar with an unusual histopathological finding: a case report. AB - This is the case of a healthy 6-year old female with a clinically absent right mandibular second primary molar with no history of that tooth ever being present. Radiographic examination revealed a well-circumscribed pericoronal radiolucency surrounding the mandibular right primary second molar. The mandibular right second premolar was displaced mesially. Treatment consisted of enucleation of the lesion with removal of both the unerupted primary second molar and second premolar. The histopathology of the excised lesion revealed a hyperplastic dental follicle with a focal proliferation of odontogenic epithelium and duct-like structures, probably representing an incipient adenomatoid odontogenic tumor. PMID- 15272664 TI - Multidisciplinary treatment of "twinned" permanent teeth: two case reports. AB - Twinned teeth usually appear in the anterior region of the dental arch. Besides orthodontic malocclusions, such as protrusion, crowding, or diastema, they also cause esthetic problems. Different treatment methods can be used according to the requirements of the situation. This article reports multidisciplinary treatment of 2 patients whose maxillary central teeth were twinned with supernumerary incisors. The twinned tooth was reshaped following endodontic treatment in 1 of the patients and orthodontic treatment was performed with edgewise mechanics. In the second case, however, it was determined that the twinned tooth had 2 separate roots. There was also another supernumerary lateral incisor in the dental arch that increased the severity of the crowding. The twinned tooth was hemisected and the other supernumerary tooth was extracted. The patient wore a removable appliance for initial tooth movements and treatment was finished with fixed appliances. PMID- 15272665 TI - Pediatric cervicofacial actinomycosis: a case report. AB - Cervicofacial actinomycosis affects many soft tissue and bony structures in the head and neck, and has both granulomatous and suppurative features. Pathogenesis of actinomycosis is still unclear, but trauma provides a portal of entry for the infection. It usually presents as a diffuse swelling with multiple sinus tracts containing macroscopic colonies of the organism known as "sulphur granules." Cervicofacial actinomycosis in children is rare. This article reports a case of actinomycosis in a 10-year-old-boy overlying the left ramus of the mandible. PMID- 15272666 TI - Dental management of an adolescent with trisomy 13 syndrome: a case report. AB - This first fully documented dental case report reviews the medical aspects of trisomy 13, summarizes the dental literature on this topic, and describes the dental findings and treatment of a 16-year-old female long-term survivor. PMID- 15272667 TI - Stretching the limits of "rights talk": securing health care entitlements for children. AB - Health care rights by their very nature have to be considered not only in a traditional legal context structured around the ideas of human autonomy but in a new analytical framework based on the notion of human interdependence. "A healthy life depends upon [social] interdependence: the quality of air, water, and sanitation which the [state] maintains for the public good; the quality of one's caring relationships, which are highly correlated to health; [as well as] the quality of health care and support furnished officially by medical institutions and provided informally by family, friends, [and the community]." PMID- 15272668 TI - Effects of gel composition on the radiation induced density change in PAG polymer gel dosimeters: a model and experimental investigations. AB - Due to a density change that occurs in irradiated polyacrylamide gel (PAG), x-ray computed tomography (CT) has emerged as a feasible method of performing polymer gel dosimetry. However, applicability of the technique is currently limited by low sensitivity of the density change to dose. This work investigates the effect of PAG composition on the radiation induced density change and provides direction for future work in improving the sensitivity of CT polymer gel dosimetry. A model is developed that describes the PAG density change (delta(rho)gel) as a function of both polymer yield (%P) and an intrinsic density change, per unit polymer yield, that occurs on conversion of monomer to polymer (delta(rho)polymer). %P is a function of the fraction of monomer consumed and the weight fraction of monomer in the unirradiated gel (%T). Applying the model to experimental CT and Raman spectroscopic data, two important fundamental properties of the response of PAG density to dose (delta(rho)gel dose response) are discovered. The first property is that delta(rho)polymer)depends on PAG %C (cross-linking fraction of total monomer) such that low and high %C PAGs exhibit a higher deltarho(polymer)than do more intermediate %C PAGs. This relationship is opposite to the relationship of polymer yield to %C and is explained by the effect of %C on the type of polymer formed. The second property is that the delta(rho)gel dose response is linearly dependent on %T. From the model, the inference is that, at least for %T < or = 2%, monomer consumption and delta(rho)polymer depend solely on %C. In terms of optimizing CT polymer gel dosimetry for high sensitivity, these results indicate that delta(rho)polymer can be expected to vary with each polymer gel system and thus should be considered when choosing a polymer gel for CT gel dosimetry. However, delta(rho)polymerand %P cannot be maximized simultaneously and maximizing %P, by choosing gels with intermediate %C and high %T, is found to have the greatest impact on increasing the sensitivity of PAG density to dose. As such, future research into new gel formulations for high sensitivity CT polymer gel dosimetry should focus on gels that exhibit an intrinsic density change and maximizing polymer yield in these systems. PMID- 15272669 TI - Evidence for using Monte Carlo calculated wall attenuation and scatter correction factors for three styles of graphite-walled ion chamber. AB - The basic equation for establishing a 60Co air-kerma standard based on a cavity ionization chamber includes a wall correction term that corrects for the attenuation and scatter of photons in the chamber wall. For over a decade, the validity of the wall correction terms determined by extrapolation methods (K(w)K(cep)) has been strongly challenged by Monte Carlo (MC) calculation methods (K(wall)). Using the linear extrapolation method with experimental data, K(w)K(cep) was determined in this study for three different styles of primary standard-grade graphite ionization chamber: cylindrical, spherical and plane parallel. For measurements taken with the same 60Co source, the air-kerma rates for these three chambers, determined using extrapolated K(w)K(cep) values, differed by up to 2%. The MC code 'EGSnrc' was used to calculate the values of K(wall) for these three chambers. Use of the calculated K(wall) values gave air kerma rates that agreed within 0.3%. The accuracy of this code was affirmed by its reliability in modelling the complex structure of the response curve obtained by rotation of the non-rotationally symmetric plane-parallel chamber. These results demonstrate that the linear extrapolation technique leads to errors in the determination of air-kerma. PMID- 15272670 TI - Inverse treatment planning using volume-based objective functions. AB - The results of optimization of inverse treatment plans depend on a choice of the objective function. Even when the optimal solution for a given cost function can be obtained, a better solution may exist for a given clinical scenario and it could be obtained with a revised objective function. In the approach presented in this work mixed integer programming was used to introduce a new volume-based objective function, which allowed for minimization of the number of under- or overdosed voxels in selected structures. By selecting and prioritizing components of this function the user could drive the computations towards the desired solution. This optimization approach was tested using cases of patients treated for prostate and oropharyngeal cancer. Initial solutions were obtained based on minimization/maximization of the dose to critical structures and targets. Subsequently, the volume-based objective functions were used to locate solutions, which satisfied better clinical objectives particular to each of the cases. For prostate cases, these additional solutions offered further improvements in sparing of the rectum or the bladder. For oropharyngeal cases, families of solutions were obtained satisfying an intensity modulated radiation therapy protocol for this disease site, while offering significant improvement in the sparing of selected critical structures, e.g., parotid glands. An additional advantage of the present approach was in providing a convenient mechanism to test the feasibility of the dose-volume histogram constraints. PMID- 15272671 TI - Artificial neural network modelling of megavoltage photon dose distributions. AB - An artificial neural network (NN) has been used to model the two-dimensional dose distributions from a Varian 2100C linac. The network was trained using depth dose data for 6 and 10 MV x-rays, collected during the linac commissioning phase. During training, the number of iterations and hidden nodes was adjusted manually until acceptable agreement between measured and predicted data was obtained. In order to validate the network a subset of the data was set aside and not used for training. This enabled the performance of the network to be investigated in terms of generalization and accuracy, together with its ability to interpolate between different field sizes and positions in the beam. Finally, the network was used to generate data points over a 2D grid so that isodose distributions could be visualized. Good agreement was found between measured data and that produced by the trained neural network. PMID- 15272672 TI - Optimization and performance evaluation of the microPET II scanner for in vivo small-animal imaging. AB - MicroPET II is a newly developed PET (positron emission tomography) scanner designed for high-resolution imaging of small animals. It consists of 17,640 LSO crystals each measuring 0.975 x 0.975 x 12.5 mm3, which are arranged in 42 contiguous rings, with 420 crystals per ring. The scanner has an axial field of view (FOV) of 4.9 cm and a transaxial FOV of 8.5 cm. The purpose of this study was to carefully evaluate the performance of the system and to optimize settings for in vivo mouse and rat imaging studies. The volumetric image resolution was found to depend strongly on the reconstruction algorithm employed and averaged 1.1 mm (1.4 microl) across the central 3 cm of the transaxial FOV when using a statistical reconstruction algorithm with accurate system modelling. The sensitivity, scatter fraction and noise-equivalent count (NEC) rate for mouse- and rat-sized phantoms were measured for different energy and timing windows. Mouse imaging was optimized with a wide open energy window (150-750 keV) and a 10 ns timing window, leading to a sensitivity of 3.3% at the centre of the FOV and a peak NEC rate of 235,000 cps for a total activity of 80 MBq (2.2 mCi) in the phantom. Rat imaging, due to the higher scatter fraction, and the activity that lies outside of the field of view, achieved a maximum NEC rate of 24,600 cps for a total activity of 80 MBq (2.2 mCi) in the phantom, with an energy window of 250 750 keV and a 6 ns timing window. The sensitivity at the centre of the FOV for these settings is 2.1%. This work demonstrates that different scanner settings are necessary to optimize the NEC count rate for different-sized animals and different injected doses. Finally, phantom and in vivo animal studies are presented to demonstrate the capabilities of microPET II for small-animal imaging studies. PMID- 15272673 TI - Feasibility of a novel design of high resolution parallax-free Compton enhanced PET scanner dedicated to brain research. AB - A novel concept for a positron emission tomography (PET) camera module is proposed, which provides full 3D reconstruction with high resolution over the total detector volume, free of parallax errors. The key components are a matrix of long scintillator crystals and hybrid photon detectors (HPDs) with matched segmentation and integrated readout electronics. The HPDs read out the two ends of the scintillator package. Both excellent spatial (x, y, z) and energy resolution are obtained. The concept allows enhancing the detection efficiency by reconstructing a significant fraction of events which underwent Compton scattering in the crystals. The proof of concept will first be demonstrated with yttrium orthoaluminate perovskite (YAP):Ce crystals, but the final design will rely on other scintillators more adequate for PET applications (e.g. LSO:Ce or LaBr3:Ce). A promising application of the proposed camera module, which is currently under development, is a high resolution 3D brain PET camera with an axial field-of-view of approximately 15 cm dedicated to brain research. The design philosophy and performance predictions based on analytical calculations and Monte Carlo simulations are presented. Image correction and reconstruction tools required to operate this transmissionless device in a research environment are also discussed. Better or similar performance parameters were obtained compared to other known designs at lower fabrication cost. The axial geometrical concept also seems to be promising for applications such as positron emission mammography. PMID- 15272674 TI - Quantitative evaluation of dual-energy digital mammography for calcification imaging. AB - Dual-energy digital mammography (DEDM), where separate low- and high-energy images are acquired and synthesized to cancel the tissue structures, may improve the ability to detect and visualize microcalcifications. Under ideal imaging conditions, when the mammography image data are free of scatter and other biases, DEDM could be used to determine the thicknesses of the imaged calcifications. We present quantitative evaluation of a DEDM technique for calcification imaging. The phantoms used in the evaluation were constructed by placing aluminium strips of known thicknesses (to simulate calcifications) across breast-tissue-equivalent materials of different glandular-tissue compositions. The images were acquired under narrow-beam geometry and high exposures to suppress the detrimental effects of scatter and random noise. The measured aluminium thicknesses were found to be approximately linear with the true aluminium thicknesses and independent of the underlying glandular-tissue composition. However, the dual-energy images underestimated the true aluminium thickness due to the presence of scatter from adjacent regions. Regions in the DEDM image that contained no aluminium yielded very low aluminium thicknesses (<0.07 mm). The aluminium contrast-to-noise ratio in the dual-energy images increased with the aluminium thickness and decreased with the glandular-tissue composition. The changes to the aluminium contrast-to noise ratio and the contrast of the tissue structures between the low-energy and DEDM images are also presented. PMID- 15272675 TI - Optimization of a fully 3D single scatter simulation algorithm for 3D PET. AB - We describe a new implementation of a single scatter simulation (SSS) algorithm for the prediction and correction of scatter in 3D PET. In this implementation, out of field of view (FoV) scatter and activity, side shields and oblique tilts are explicitly modelled. Comparison of SSS predictions with Monte Carlo simulations and experimental data from uniform, line and cold-bar phantoms showed that the code is accurate for uniform as well as asymmetric objects and can model different energy resolution crystals and low level discriminator (LLD) settings. Absolute quantitation studies show that for most applications, the code provides a better scatter estimate than the tail-fitting scatter correction method currently in use at our institution. Several parameters such as the density of scatter points, the number of scatter distribution sampling points and the axial extent of the FoV were optimized to minimize execution time, with particular emphasis on patient studies. Development and optimization were carried out in the case of GSO-based scanners, which enjoy relatively good energy resolution. SSS estimates for scanners with lower energy resolution may result in different agreement, especially because of a higher fraction of multiple scatter events. The algorithm was applied to a brain phantom as well as to clinical whole-body studies. It proved robust in the case of large patients, where the scatter fraction increases. The execution time, inclusive of interpolation, is typically under 5 min for a whole-body study (axial FoV: 81 cm) of a 100 kg patient. PMID- 15272676 TI - LiF:W as a scintillator for dosimetry in diagnostic radiology. AB - The use of ionization chambers in diagnostic radiology is not feasible in measurement situations requiring small and robust dose sensors. The composition of LiF:W developed as a scintillator for the measurement of thermal neutrons seems profitable for an application in dosimetry of low-energy photons. Properties of a small LiF:W scintillator were determined with DV- and DH-standard radiation qualities. For a tube voltage range of 40 to 150 kV (corresponding to a HVL of 1.56 to 13.69 mm Al) a maximum variation in sensitivity of +/-13% was determined for a scintillator thickness of 3.7 mm. The scintillator signal was linear in a range from 6.6 mGy min(-1) to at least 13.7 Gy min(-1). Higher dose rates could not be obtained in the measurement setup. The temperature dependence of the luminescence response was found to decrease from +3.7% (at +2.5 degrees C) to -7.0% (at +45 degrees C) with respect to the luminescence response at 20 degrees C. LiF:W appears to be an interesting choice as a dosimetric scintillator in diagnostic radiology making immediate measurements of dose and dose rate with high spatial resolution feasible. PMID- 15272677 TI - A geometric model for evaluating the effects of inter-fraction rectal motion during prostate radiotherapy. AB - A geometric model is presented which allows calculation of the dosimetric consequences of rectal motion in prostate radiotherapy. Variations in the position of the rectum are measured by repeat CT scanning during the courses of treatment of five patients. Dose distributions are calculated by applying the same conformal treatment plan to each imaged fraction and rectal dose-surface histograms produced. The 2D model allows isotropic expansion and contraction in the plane of each CT slice. By summing the dose to specific volume elements tracked by the model, composite dose distributions are produced that explicitly include measured inter-fraction motion for each patient. These are then used to estimate effective dose-surface histograms (DSHs) for the entire treatment. Results are presented showing the magnitudes of the measured target and rectal motion and showing the effects of this motion on the integral dose to the rectum. The possibility of using such information to calculate normal tissue complication probabilities (NTCP) is demonstrated and discussed. PMID- 15272678 TI - An accurate calibration method of the multileaf collimator valid for conformal and intensity modulated radiation treatments. AB - Because for IMRT treatments the required accuracy on leaf positioning is high, conventional calibration methods may not be appropriate. The aim of this study was to develop the tools for an accurate MLC calibration valid for conventional and IMRT treatments and to investigate the stability of the MLC. A strip test consisting of nine adjacent segments 2 cm wide, separated by 1 mm and exposed on Kodak X-Omat V films at Dmax depth, was used for detecting leaf-positioning errors. Dose profiles along the leaf-axis were taken for each leaf-pair. We measured the dose variation on each abutment to quantify the relative positioning error (RPE) and the absolute position of the abutment to quantify the absolute positioning error (APE). The accuracy of determining the APE and RPE was 0.15 and 0.04 mm, respectively. Using the RPE and the APE the MLC calibration parameters were calculated in order to obtain a flat profile on the abutment at the correct position. A conventionally calibrated Elekta MLC was re-calibrated using the strip test. The stability of the MLC and leaf-positioning reproducibility was investigated exposing films with 25 adjacent segments 1 cm wide during three months and measuring the standard deviation of the RPE values. A maximum shift over the three months of 0.27 mm was observed and the standard deviation of the RPE values was 0.11 mm. PMID- 15272679 TI - A six-bank multi-leaf system for high precision shaping of large fields. AB - In this study, we present the design for an alternative MLC system that allows high precision shaping of large fields. The MLC system consists of three layers of two opposing leaf banks. The layers are rotated 60 degrees relative to each other. The leaves in each bank have a standard width of 1 cm projected at the isocentre. Because of the symmetry of the collimator set-up it is expected that collimator rotation will not be required, thus simplifying the construction considerably. A 3D ray tracing computer program was developed in order to simulate the fluence profile for a given collimator and used to optimize the design and investigate its performance. The simulations show that a six-bank collimator will afford field shaping of fields of about 40 cm diameter with a precision comparable to that of existing mini MLCs with a leaf width of 4 mm. PMID- 15272680 TI - Modelling of electron contamination in clinical photon beams for Monte Carlo dose calculation. AB - The purpose of this work is to model electron contamination in clinical photon beams and to commission the source model using measured data for Monte Carlo treatment planning. In this work, a planar source is used to represent the contaminant electrons at a plane above the upper jaws. The source size depends on the dimensions of the field size at the isocentre. The energy spectra of the contaminant electrons are predetermined using Monte Carlo simulations for photon beams from different clinical accelerators. A 'random creep' method is employed to derive the weight of the electron contamination source by matching Monte Carlo calculated monoenergetic photon and electron percent depth-dose (PDD) curves with measured PDD curves. We have integrated this electron contamination source into a previously developed multiple source model and validated the model for photon beams from Siemens PRIMUS accelerators. The EGS4 based Monte Carlo user code BEAM and MCSIM were used for linac head sinulation and dose calculation. The Monte Carlo calculated dose distributions were compared with measured data. Our results showed good agreement (less than 2% or 2 mm) for 6, 10 and 18 MV photon beams. PMID- 15272681 TI - The effect of patient inhomogeneities in oesophageal 192Ir HDR brachytherapy: a Monte Carlo and analytical dosimetry study. AB - The effect of patient inhomogeneities surrounding the oesophagus on the dosimetry planning of an upper thoracic oesophageal 192Ir HDR brachytherapy treatment is studied. The MCNPX Monte Carlo code is used for dosimetry in a patient-equivalent phantom geometry and results are compared in terms of isodose contours as well as dose volume histograms with corresponding calculations by a contemporary treatment planning system software featuring a full TG-43 dose calculation algorithm (PLATO BPS version 14.2.4). It is found that the presence of patient inhomogeneities does not alter the delivery of the planned dose distribution to the planning treatment volume. Regarding the organs at risk, the common practice of current treatment planning systems (TPSs) to consider the patient geometry as a homogeneous water medium leads to a dose overestimation of up to 13% to the spinal cord and an underestimation of up to 15% to the sternum bone. These findings which correspond to the dose region of about 5-10% of the prescribed dose could only be of significance when brachytherapy is used as a boost to external beam therapy. Additionally, an analytical dosimetry model, which is efficient in calculating dose in mathematical phantoms containing inhomogeneity shells of materials of radiobiological interest, is utilized for dosimetry in the patient-equivalent inhomogeneous phantom geometry. Analytical calculations in this work are in good agreement with corresponding Monte Carlo results within the bone inhomogeneities of spinal cord and sternum bone but, like treatment planning system calculations, the model fails to predict the dose distribution in the proximal lung surface as well as within the lungs just as the TPS does, due to its inherent limitation in treating lateral scatter and backscatter radiation. PMID- 15272682 TI - A simplified model of the source channel of the Leksell GammaKnife tested with PENELOPE. AB - Monte Carlo simulations using the code PENELOPE have been performed to test a simplified model of the source channel geometry of the Leksell GammaKnife. The characteristics of the radiation passing through the treatment helmets are analysed in detail. We have found that only primary particles emitted from the source with polar angles smaller than 3 degrees with respect to the beam axis are relevant for the dosimetry of the Gamma Knife. The photon trajectories reaching the output helmet collimators at (x, v, z = 236 mm) show strong correlations between rho = (x2 + y2)(1/2) and their polar angle theta, on one side, and between tan(-1)(y/x) and their azimuthal angle phi, on the other. This enables us to propose a simplified model which treats the full source channel as a mathematical collimator. This simplified model produces doses in good agreement with those found for the full geometry. In the region of maximal dose, the relative differences between both calculations are within 3%, for the 18 and 14 mm helmets, and 10%, for the 8 and 4 mm ones. Besides, the simplified model permits a strong reduction (larger than a factor 15) in the computational time. PMID- 15272683 TI - MR-guided stereotactic neurosurgery--comparison of fiducial-based and anatomical landmark transformation approaches. AB - For application in magnetic resonance (MR) guided stereotactic neurosurgery, two methods for transformation of MR-image coordinates in stereotactic, frame-based coordinates exist: the direct stereotactic fiducial-based transformation method and the indirect anatomical landmark method. In contrast to direct stereotactic MR transformation, indirect transformation is based on anatomical landmark coregistration of stereotactic computerized tomography and non-stereotactic MR images. In a patient study, both transformation methods have been investigated with visual inspection and mutual information analysis. Comparison was done for our standard imaging protocol, including t2-weighted spin-echo as well as contrast enhanced t1-weighted gradient-echo imaging. For t2-weighted spin-echo imaging, both methods showed almost similar and satisfying performance with a small, but significant advantage for fiducial-based transformation. In contrast, for t1-weighted gradient-echo imaging with more geometric distortions due to field inhomogenities and gradient nonlinearity than t2-weighted spin-echo imaging, mainly caused by a reduced bandwidth per pixel, anatomical landmark transformation delivered markedly better results. Here, fiducial-based transformation yielded results which are intolerable for stereotactic neurosurgery. Mean Euclidian distances between both transformation methods were 0.96 mm for t2-weighted spin-echo and 1.67 mm for t1-weighted gradient-echo imaging. Maximum deviations were 1.72 mm and 3.06 mm, respectively. PMID- 15272684 TI - Image reconstruction on PI-lines by use of filtered backprojection in helical cone-beam CT. AB - Recently, we have derived a general formula for image reconstruction from helical cone-beam projections. Based upon this formula, we have also developed an exact algorithm for image reconstruction on PI-line segments from minimum data within the Tam-Danielsson window. This previous algorithm can be referred to as a backprojection-filtration algorithm because it reconstructs an image by first backprojection of the data derivatives and then filtration of the backprojections on PI-line segments. In this work, we propose an alternative algorithm, which reconstructs an image by first filtering the modified data along the cone-beam projections of the PI-lines onto the detector plane and then backprojecting the filtered data onto PI-line segments. Therefore, we refer to this alternative algorithm as the filtered-backprojection algorithm. A preliminary computer simulation study was performed for validating and demonstrating this new algorithm. Furthermore, we derive a practically useful expression to accurately compute the derivative of the data function for image reconstruction. The proposed filtered-backprojection algorithm can reconstruct the image within any selected ROI inside the helix and thus can handle naturally the long object problem and the super-short scan problem. It can also be generalized to reconstruct images from data acquired with other scanning configurations such as the helical scan with a varying pitch. PMID- 15272685 TI - Half-scan and single-plane intensity diffraction tomography for phase objects. AB - A reconstruction theory for intensity diffraction tomography (I-DT) has been proposed that permits reconstruction of a weakly scattering object without explicit knowledge of phase information. In this work, we examine the application of I-DT, using either planar- or spherical-wave incident wavefields, for imaging three-dimensional (3D) phase objects. We develop and investigate two algorithms for reconstructing phase objects that utilize only half of the measurements that would be needed to reconstruct a complex-valued object function. Each reconstruction algorithm reconstructs the phase object by use of different sets of intensity measurements. Although the developed reconstruction algorithms are equivalent mathematically, we demonstrate that their numerical and noise propagation properties differ considerably. We implement numerically the reconstruction algorithms and present reconstructed images to demonstrate their use and to corroborate our theoretical assertions. PMID- 15272686 TI - Theoretical evaluation of accuracy in position and size of brain activity obtained by near-infrared topography. AB - Near-infrared (NIR) topography can obtain a topographical distribution of the activated region in the brain cortex. Near-infrared light is strongly scattered in the head, and the volume of tissue sampled by a source-detector pair on the head surface is broadly distributed in the brain. This scattering effect results in poor resolution and contrast in the topographic image of the brain activity. In this study, a one-dimensional distribution of absorption change in a head model is calculated by mapping and reconstruction methods to evaluate the effect of the image reconstruction algorithm and the interval of measurement points for topographic imaging on the accuracy of the topographic image. The light propagation in the head model is predicted by Monte Carlo simulation to obtain the spatial sensitivity profile for a source-detector pair. The measurement points are one-dimensionally arranged on the surface of the model, and the distance between adjacent measurement points is varied from 4 mm to 28 mm. Small intervals of the measurement points improve the topographic image calculated by both the mapping and reconstruction methods. In the conventional mapping method, the limit of the spatial resolution depends upon the interval of the measurement points and spatial sensitivity profile for source-detector pairs. The reconstruction method has advantages over the mapping method which improve the results of one-dimensional analysis when the interval of measurement points is less than 12 mm. The effect of overlapping of spatial sensitivity profiles indicates that the reconstruction method may be effective to improve the spatial resolution of a two-dimensional reconstruction of topographic image obtained with larger interval of measurement points. Near-infrared topography with the reconstruction method potentially obtains an accurate distribution of absorption change in the brain even if the size of absorption change is less than 10 mm. PMID- 15272687 TI - Diffuse optical tomography with physiological and spatial a priori constraints. AB - Diffuse optical tomography is a typical inverse problem plagued by ill-condition. To overcome this drawback, regularization or constraining techniques are incorporated in the inverse formulation. In this work, we investigate the enhancement in recovering functional parameters by using physiological and spatial a priori constraints. More accurate recovery of the two main functional parameters that are the blood volume and the relative saturation is demonstrated through simulations by using our method compared to actual techniques. PMID- 15272688 TI - Mapping human skeletal muscle perforator vessels using a quantum well infrared photodetector (QWIP) might explain the variability of NIRS and LDF measurements. AB - Near-infrared spectroscopy (NIRS) and laser Doppler flowmetry (LDF) have become the techniques of choice allowing the non-invasive study of local human skeletal muscle metabolism and blood perfusion on a small tissue volume (a few cm3). However, it has been shown that both NIRS and LDF measurements may show a large spatial variability depending on the position of the optodes over the investigated muscle. This variability may be due to local morphologic and/or metabolic characteristics of the muscle and makes the data interpretation and comparison difficult. In the present work, we use a third method to investigate this problem which permits fast, non-invasive mapping of the intramuscular vessel distribution in the human vastus latelralis muscle. This method uses an advanced, passive, infrared imaging sensor called a QWIP (quantum well infrared photodetector). We demonstrate, using a recovery-enhanced infrared imaging technique, that there is a significant presence of perforator vessels in the region of interest of approximately 30 x 18 cm (the number of vessels being: 14, 9, 8, 33, 17 and 18 for each subject, respectively). The presence of these vessels makes the skeletal muscle highly inhomogeneous, and may explain the observed NIRS and LDF spatial variability. We conclude that accurate comparison of the metabolic activity of two different muscle regions is not possible without reliable maps of vascular 'singularities' such as the perforator vessels, and that the QWIP-based imaging system is one method to obtain this information. PMID- 15272689 TI - Influence of intra-fractional breathing movement in step-and-shoot IMRT. AB - Efforts have been made to extend the application of intensity-modulated radiotherapy to a variety of organs. One of the unanswered questions is whether breathing-induced organ motion may lead to a relevant over- or underdosage, e.g., in treatment plans for the irradiation of lung cancer. Theoretical considerations have been made concerning the different kinds of IMRT but there is still a lack of experimental data. We examined 18 points in a fraction of a clinical treatment plan of a NSCLC delivered in static IMRT with a new phantom and nine ionization chambers. Measurements were performed at a speed of 12 and 16 breathing cycles per minute. The dose differences between static points and moving target points ranged between -2.4% and +5.5% (mean: +0.2%, median: -0.1%) when moving with 12 cycles min(-1) and between -3.6% and +5.0% (mean: -0.4%, median: -0.6%) when moving with 16 cycles min(-1). All differences of measurements with and without movements were below 5%, with one exception. In conclusion, our results underline that at least in static IMRT breathing effects (concerning target dose coverage) due to interplay effects between collimator leaf movement and target movement are of secondary importance and will not reduce the clinical value of IMRT in the step-and-shoot technique for irradiation of thoracic targets. PMID- 15272690 TI - America's best hospitals. PMID- 15272691 TI - Behind the rankings. PMID- 15272692 TI - Hidden specialties. PMID- 15272693 TI - For the sickest kids. PMID- 15272694 TI - Soul of a surgeon. PMID- 15272695 TI - You're never too old. Surgery on patients of 80, 90, and up? It's gaining acceptance. PMID- 15272696 TI - Special report--Best hospitals. PMID- 15272697 TI - Special report--Pioneers of medicine. Walter Willett. The truth on foods and fats. PMID- 15272698 TI - Special report--Pioneers of medicine. Ronald Levy. Cancer's natural enemy. PMID- 15272699 TI - Special report--Pioneers of medicine. Robert Jennings. Attacking the heart attack. PMID- 15272700 TI - Special report--Pioneers of medicine. Phill Wilson. AIDS: darkening in America. PMID- 15272701 TI - Nitrous oxide emission and reduction in a laboratory-incubated paddy soil response to pretreatment of water regime. AB - A laboratory incubation experiment was conducted to investigate nitrous oxide (N2O) emission and reduction in a paddy soil (Stagnic Anthrosol) response to the pretreatment of water regime. The paddy soil was maintained under either air dried (sample D) or submerged (sample F) conditions for 110 d before the soil was adjusted into soil moisture of 20% , 40% , 60% , 80% and 100% water holding capacity (WH(-2)) respectively, and then incubated with or without 10% (v/v) acetylene for 138 h at 25 degrees C. At lower soil water content (< or = 60% WHC), N2O emission from the sample F was 2.29 times higher than that from the sample D (P < 0.01). While, N2O emission from the sample F was only 29 and 14 percent of that from the sample D at the soil moisture of 80% and 100% WHC, respectively (P < 0.01). The maximal N2O emissions observed at soil moisture of 80% WHC were about 24 and 186 times higher than the minima obtained at the soil moisture of 20% WHC for the sample F and D, respectively. But at the soil moisture of 80% and 100% WHC, N2O emission from the sample F with acetylene (F + ACE) was comparable to that of the sample D with acetylene (D + ACE). The results showed that the F sample produced N2O ability in denitrification was similar to the sample D, however, the sample F was in the better reduction of N2O to N, than the sample D even after the soil moisture was adjusted into the same level of 80% or 100% WHC. Therefore, the pretreatment of water regime influenced the strength and product composition of denitrification and N2O emission from the paddy soil. PMID- 15272702 TI - Effect of nitrogen and phosphorus on the water quality in the Three Gorges Reservoir Area during and after its construction. AB - A survey concerning the concentration of the nutrients in the Three Gorges Reservoir Area was carried out. This paper presents the parameters(NO3- -N, NO2- N, Kjeldahl-N, non-ionic ammonia, P-PO4 and TP) determined at 16 sampling sites from 1997 to 1999. The dominant soluble nitrogen form was NO3- -N followed by Kjeldahl-N, NO1- -N and non-ionic ammonia. Mean values of NO3- -N, NO2- -N, Kjeldahl-N and non-ionic ammonia ranged from 0.50 to 2.37 mg/L, 0.022 to 0.084 mg/L, 0.33 to 0.99 mg/L and 0.007 to 0.092 mg/L respectively. Mean values of P PO4 at most sampling sites were higher than 0.1 mg/L for subject to eutrophication. The major factors influencing the concentrations of N and P might be agricultural runoff, municipal and industrial effluents. In addition, 6 kinds of soil were sampled at the area where would inundated after the dam completed. Two approaches were adopted to simulate the N and P release from the inundated soils. The results showed that the soils would release nitrogen and phosphorus to the overlying water when the soils were inundated. The characteristics of soil affected the equilibrium concentrations of N and P between the soil and the overlying water. PMID- 15272703 TI - Degradation of nitrobenzene in wastewater by gamma-ray irradiation. AB - The degradation of nitrobenzene (NB) by gamma-ray irradiation was studied. The influences of dose rate and initial NB concentration were investigated in details. At a dose rate of 55 Gy/min, the degradation kinetics was pseudo-first order at NB concentrations from 0.2 mmol/L to 4.0 mmol/L. At an initial NB concentration of 0.8 mmol/L, the degradation of NB at various dose rates also followed pseudo-first-order kinetics. Dissolved oxygen was found to have a positive effect on NB degradation. The degradation products were identified as nitrophenol, nitrosobenzene, and hydroquinone, and so on. Based on the product analysis, possible degradation pathways of nitrobenzene were proposed. PMID- 15272704 TI - An innovative integrated oxidation ditch with vertical circle (IODVC) for wastewater treatment. AB - The oxidation ditch process is economic and efficient for wastewater treatment, but its application is limited in case where land is costly due to its large land area required. An innovative integrated oxidation ditch with vertical circle (IODVC) system was developed to treat domestic and industrial wastewater aiming to save land area. The new system consists of a single-channel divided into two ditches(the top one and the bottom one by a plate), a brush, and an innovative integral clarifier. Different from the horizontal circle of the conventional oxidation ditch, the flow of IODVC system recycles from the top zone to the bottom zone in the vertical circle as the brush is running, and then the IODVC saved land area required by about 50% compared with a conventional oxidation ditch with an intrachannel clarifier. The innovative integral clarifier is effective for separation of liquid and solids, and is preferably positioned at the opposite end of the brush in the ditch. It does not affect the hydrodynamic characteristics of the mixed liquor in the ditch, and the sludge can automatically return to the down ditch without any pump. In this study, experiments of domestic and dye wastewater treatment were carried out in bench scale and in full scale, respectively. Results clearly showed that the IODVC efficiently removed pollutants in the wastewaters, i.e., the average of COD removals for domestic and dye wastewater treatment were 95% and 90%, respectively, and that the IODVC process may provide a cost effective way for full scale dye wastewater treatment. PMID- 15272705 TI - Heavy metals pollution in poultry and livestock feeds and manures under intensive farming in Jiangsu Province, China. AB - The heavy metals pollution in poultry and livestock feeds and manures under intensive farming in Jiangsu Province was investigated. 97 feed and manure samples were sampled from 31 farming plants in 10 major cities of Jiangsu. 14 metals, including Zn, Cu, Pb, Cd, Cr, Ni, Mo, Mn, Ba, Co, Sr, Ti, As and Hg, were analyzed after samples acid digestion. The results showed that the most feed samples contained high concentration of metals exceeding National Hygienical Standard for Feeds. Meanwhile, it was found that Cu, Zn, Pb, Cd and Cr concentrations in animal manures were also high, for example, Cu concentration in a manure sample reached to as much as 1726.3 mg/kg. Heavy metals loading quantities in soil per year were then calculated when metals contaminated organic fertilizers were applied, and its effects on soil environmental quality were further evaluated. PMID- 15272706 TI - Photocatalytic degradation of dye effluent by titanium dioxide pillar pellets in aqueous solution. AB - Photocatalytic oxidation (PCO) process is an effective way to deal with organic pollutants in wastewater which could be difficult to be degraded by conventional biological treatment methods. Normally the TiO2 powder in nanometre size range was directly used as photocatalyst for dye degradation in wastewater. However the titanium dioxide powder was arduous to be recovered from the solution after treatment. In this application, a new form of TiO2 (i.e. pillar pellets ranging from 2.5 to 5.3 mm long and with a diameter of 3.7 mm) was used and investigated for photocatalytic degradation of textile dye effluent. A test system was built with a flat plate reactor (FPR) and UV light source (blacklight and solar simulator as light source respectively) for investigating the effectiveness of the new form of TiO2. It was found that the photocatalytic process under this configuration could efficiently remove colours from textile dyeing effluent. Comparing with the TiO2 powder, the pellet was very easy to recovered from the treated solution and can be reused in multiple times without the significant change on the photocatalytic property. The results also showed that to achieve the same photocatalytic performance, the FPR area by pellets was about 91% smaller than required by TiO2 powder. At least TiO2 pellet could be used as an alternative form of photocatalyst in applications for textile effluent treatment process, also other wastewater treatment processes. PMID- 15272707 TI - Alternating shortcut nitrification-denitrification for nitrogen removal from soybean wastewater by SBR with real-time control. AB - A novel treating technology for nitrogen removal from soybean wastewater was studied. The process for nitrogen removal was achieved by alternating aeration and mixing, combined with real-time control strategies. Results showed that the COD and total nitrogen removal rates are more than 90% and 92% at COD and total nitrogen loads of 1.0 - 1.2 kg COD/(kgMLSS x d) and 0.20 - 0.27 kg TN/(kgMLSS x d), respectively. In addition, it could improve sludge settling property. SVI value is less than 70 g/ml during the whole cycles. The method not only may be adapted to treat soybean wastewater with high nitrogen, but also may be applied to treat other high nitrogen wastewater. PMID- 15272708 TI - Biological removal of methanol from process condensate for the purpose of reclamation. AB - The biological removal of methanol from condensate of ammonia manufacturing processes for the purpose of reclamation using contact type reactor was studied. Methanol of 60 mg/L was removed completely under an HRT of 1.12 h. Optimal inorganic nutrient dose was determined on evaluating methanol removal performance and dehydrogenase activities (DHA) under different nutrition doses. The optimal inorganic nutrient dose only gave an increase of conductivity of ca. 10 micros/cm2 in the effluent on treating synthetic condensate containing methanol of 30 mg/L. The results demonstrated that biological removal of methanol was effective for the purpose of recovering the methanol-bearing condensate. PMID- 15272709 TI - Conversion regular patterns of acetic acid, propionic acid and butyric acid in UASB reactor. AB - On the basis of continuous tests and batch tests, conversion regular patterns of acetate, propionate and butyrate in activated sludge at different heights of the UASB reactor were conducted. Results indicated that the conversion capacity of the microbe is decided by the substrate characteristic when sole VFA is used as the only substrate. But when mixed substrates are used, the conversion regulations would have changed accordingly. Relationships of different substrates vary according to their locations. In the whole reactor, propionate's conversion is restrained by acetate and butyrate of high concentration. On the top and at the bottom of the reactor, conversion of acetate, but butyrate, is restrained by propionate. And in the midst, acetate's conversion is accelerated by propionate while that of butyrate is restrained. It is proved, based on the analysis of specific conversion rate, that the space distribution of the microbe is the main factor that affects substrates' conversion. The ethanol-type fermentation of the acidogenic-phase is the optimal acid-type fermentation for the two-phase anaerobic process. PMID- 15272710 TI - Photochemical surface modification of poly(arylsulfone) ultrafiltration membrane and covalent immobilization of enzyme. AB - The sensitivity of poly(arylsulfone) (PSf) for UV irradiation in different solvents (water and ethanol) was investigated. It is confirmed that acrylic acid (AA) and acrylamide (AAm) are grafted only onto the surface of the membrane instead of the interior by FTIR and scanning electron microscope (SEM). The membrane performance (deltaJ/J0 and contact angle theta) after photografting was studied. In the range of conditions used, the grafting yield increases with irradiation time and monomer concentration growing. After photografting and N-3 dimethylaminopropyl-N'-ethycarbodiimide hydrochloride (EDC) activation, PSf membrane was immobilized with hydrogen peroxide oxidoreductase, and showed a higher activity than the control membrane. PMID- 15272711 TI - Stabilization effects of surplus soft clay with cement and GBF slag. AB - Utilization of industrial waste and surplus construction soft clay as construction material was recommended, and many attempts at geotechnical waste utilization were undertaken. This study aimed at the application of cement and a kind of industrial wastes, i.e. granulated blast furnace slag, on stabilization of surplus soft clay. The results showed that the cement and slag can successfully stabilize Ariake clays even though this high organic clay fails to be stabilized by lime and cement. Addition of slag in cement for stabilization induces higher strength than cement alone for longer curing time. The application of the cement with slag is more suitable than cement alone for stabilization because of economical consideration. PMID- 15272712 TI - Agroecosystem functional assessment and its difficulties. AB - Agroecosystem functional assessment indicators provide a necessary bridge between decision-makers and scientists. The development of acceptable indicators, however, remains a difficult task because the current knowledge and understanding of ecosystems is not sufficient to allow an objective assessment of all ecosystem functions. These difficulties were summarized from three perspectives. First, there are difficulties in individual function assessment. Of the four functions associated with agroecosystems-energy flow, materials cycling, information flow and value flow-data on material cycling and information flow remain difficult to obtain and the indicators relatively immature. Secondly, there are difficulties of integration. During the assessment process, the integration of the agroecosystem functions remains the biggest obstacle. Until now, there has been no practical or effective methodology established to resolve the problem. At present, the makeshift approach has been to weight the various indicators and then add them together. Thirdly, there is the problem of obscure concepts and concept confusion. When assessments of agroecosystems are conducted, concepts such as structure, function, benefit, and resource utilization are used extensively. To date, no logical relationship (either real or implied) has been developed between any of these concepts. Are they causes and results such that the relationship between them is linear, or are they independent from one another such that the relationship is parallel? Thus far, the essence of this question is yet to be explored. PMID- 15272713 TI - Kinetics study of aqueous sorption of phenanthrene to humic acids and sediments. AB - The sorption behavior was determined for a model polycyclic aromatic hydrocarbon (PAH), i.e., phenanthrene(PHN), from water to three humic acids (HAs) and three sediments in different reacting time. The chemical compositions of HA samples were measured using cross polarization magic angle spinning carbon-13 (CPMAS 13C NMR along with elemental analysis. The dissolved humic substances dissociating from solid HAs and sediments were characterized by 'H NMR. The experiments indicated that the sorption modes and mechanisms of natural sorbents for PHN varied significantly between short (< 7 d) and long contact time and the reaction time should be taken into consideration in studying the overall sorption process. The sorption capacity (K'f) and exponent (n) might be relative to the properties of dissolved humic materials in initial stage but the solid aromatic organic matter after long time reaction. According to the experiments performed in this investigation and the previous researches, a conceptive sorption model was established. PMID- 15272714 TI - Influences of excessive Cu on photosynthesis and growth in ectomycorrhizal Pinus sylvestris seedlings. AB - Growth and photosynthesis responses were measured for Scots pine (Pinus sylvestris L. cv.) inoculated with ectomycorrhizal fungi (Suillus bovinus) under 6.5 and 25 mg/L Cu treatments to evaluate ectomycorrhizal seedlings' tolerance to heavy metal stress. Results showed that excessive Cu can significantly impair the growth and photosynthesis of pine seedlings, but such impairment is much smaller to the ectomycorrhizal seedlings. Under 25 mg/L Cu treatment, the dry weight of ectomycorrhizal seedlings is 25% lower than the control in contrary to 53% of the non-mycorrhizal seedlings, and the fresh weight of ectomycorrhizal roots was significantly higher than those of non-mycorrhizal roots, about 25% and 42% higher at 6.5 and 25 mg/L Cu treatments respectively. Furthermore, ectomycorrhizal fungi induced remarkable difference in the growth rate and pigment content of seedlings under excessive Cu stress. At 25 mg/L Cu, the contents of total chlorophyll, chlorophyll-a and chlorophyll-b were 30% higher in ectomycorrhizal plants than those in non-mycorrhizal plants. O2 evolution and electron transport of PSI and PSII were restrained by elevated Cu stress. However, no significant improvement was observed in reducing the physiological restraining in ectomycorrhizal seedlings over the non-mycorrhizal ones. PMID- 15272715 TI - Interaction of PACls with sulfate. AB - This article discusses the influential factors on Al13 separation considering the interaction of sulfate with various polyaluminum chloride (PACl). The experimental results showed that the basicity (B = [OH/[Al]), the concentration of PACl and Al/SO4 ratio exhibited significant roles in the PACl-sulfate reaction. It indicated that different species in various PACl underwent different reaction pathway with sulfate. The Alc, colloidal species, formed precipitation quickly with sulfate, while Alb, oligomers and polymers, underwent slow crystallization. And Ala, monomers, reacted with sulfate to form soluble complexes. The kinetic difference of reaction made it possible to realize the separation of Alb and further purification. The decrease of Ala resulted in the limit of ferron method was also mentioned. PMID- 15272716 TI - Holographic quantitative structure-activity relationship for prediction acute toxicity of benzene derivatives to the guppy (Poecilia reticulata). AB - Holographic quantitative structure-activity relationship (HQSAR) is an emerging QSAR technique with the combined application of molecular hologram, which encoded the frequency of occurrence of various molecular fragment types, and the subsequent partial least squares (PLS) regression analysis. In this paper, the acute toxicity data to the guppy (Poecilia reticulata) for a series of 56 substituted benzenes, phenols, aromatic amines and nitro-aromatics were subjected and this resulted in a model with a high predictive ability. The influence of fragment size and fragment distinction parameters on the quality of HQSAR model was investigated. The robustness and predictive ability of the model were also validated by leave-one-out (LOO) cross-validation procedure and external testing data set. PMID- 15272717 TI - Dissimilatory reduction of FeIII (EDTA) with microorganisms in the system of nitric oxide removal from the flue gas by metal chelate absorption. AB - In the system of nitric oxide removal from the flue gas by metal chelate absorption, it is an obstacle that ferrous absorbents are easily oxidized by oxygen in the flue gas to ferric counterparts, which are not capable of binding NO. By adding iron metal or electrochemical method, FeIII(EDTA) can be reduced to FeII(EDTA). However, there are various drawbacks associated with these techniques. The dissimilatory reduction of FeIII(EDTA) with microorganisms in the system of nitric oxide removal by metal chelate absorption was investigated. Ammonium salt instead of nitrate was used as the nitrogen source, as nitrates inhibited the reduction of FeIII due to the competition between the two electron acceptors. Supplemental glucose and lactate stimulated the formation of FeII more than ethanol as the carbon sources. The microorganisms cultured at 50 degrees C were not very sensitive to the other experimental temperature, the reduction percentage of FeIII varied little with the temperature range of 30-50 degrees C. Concentrated Na2CO3 solution was added to adjust the solution pH to an optimal pH range of 6-7. The overall results revealed that the dissimilatory ferric reducing microorganisms present in the mix-culture are probably neutrophilic, moderately thermophilic FeIII reducers. PMID- 15272718 TI - Sonolysis and mineralization of pentachlorophenol by means of varying parameters. AB - Degradation effect of organic pollutant on pentachlorophenol (PCP) is researched by ultrasound. PCP is treated by low frequency (16 kHz) and high frequency (800 +/- 1 kHz), and bi-frequency. The results of investigation on the ultrasonic destruction of PCP showed that the rate of PCP degradation at the same conditions is the highest at bi-frequency, and the lowest at 16 kHz. In the presence of Fenton type reagent the degradation rate of PCP is the highest at bi-frequency (20.93 times) as compared to the stirring system. This ratio is 4.91 and 1.06 at 800 kHz and 16 kHz, respectively. The studies showed the bi-frequency is an effective method for pollutants degradation, but it is need make further study. Taking 800 kHz for example, under the same conditions, the smaller pH of the solution, the higher is the reaction rate. A similar situation applied to TOC, but the TOC removal lags behind degradation of PCP. This indicated the PCP is not completely mineralized. The ultrasound is somewhat enhanced for degradation of PCP and mineralization with only addition of CuSO4, but the combination of ultrasound and Fenton type reagent is effective method for PCP degradation and mineralization. The rate of PCP degradation and TOC removal appeared to follow pseudo-first-order reaction kinetic law. PMID- 15272719 TI - Effects of land-use pattern change on rainfall-runoff and runoff-sediment relations: a case study in Zichang watershed of the Loess Plateau of China. AB - The purpose of this article is to identify the effect of land-use pattern on rainfall-runoff and runoff-sediment relations in Zichang Watershed of the Loess Plateau. From 1986 to 1997, many farmlands changed into grassland or woodland, especially the farmland in steep slope positions or far away from the river. The change of land-use pattern altered the rainfall-runoff and runoff-sediment relationships, and led to higher slope of trend curves (STCs) of annual rainfall runoff mass curve and runoff-sediment mass curve in 1990s than that in 1980s. It is implied that more soil and water loss yielded in 1990s. In order to reduce soil loss, more attentions should be paid to land-use pattern and some grass or other herbaceous filter strips should be built along rivers. PMID- 15272720 TI - Flocculation performance of a novel synthesized flocculant with low ecological risk. AB - Combined flocculants with low ecological risk are urgently required in water supply and wastewater treatment in China. A novel flocculant was thus developed under the condition of low ecological risk (noted as CAS). The experiments to examine wastewater treatment performance of the new product showed that there was favourable performance in the flocculation process in contrast to commercial flocculants in treating kaolin suspensions, municipal effluent and domestic wastewater. Flocculation performance included the turbidity removal rate, sediment character and a decrease in COD (chemical oxygen demand). The sediment time of flocculation is short and the removal rate of turbidity treated by CAS is high compared with PAC (polyaluminum-chloride), PAM (polyacrylamide) and the combined addition of PAC and PAM. The optimal concentration required to affect flocculation processes is dependent on kaolin concentration and the character of the wastewater within the range examined. It also showed that CAS is effective to treat wastewater with high turbidity. PMID- 15272721 TI - Arsenic toxicity in mice and its possible amelioration. AB - Oral administration of arsenic trioxide (3 and 6 mg/kg body weight/d) for 30 d caused, as compared with vehicle control, dose-dependent significant reductions in body weight, absolute weight, protein, glycogen, as well as, total, dehydro and reduced ascorbic acid contents both in the liver and kidney of arsenic treated mice. Succinic dehydrogenase (SDH) and phosphorylase only in the liver activities were significantly reduced in a dose-dependent manner. Acid phosphatase activity was significantly decreased in the liver of low dose arsenic treated animals; however, significant rise in its activity was observed in high dose group. As compared with vehicle control, treatment also caused significant dose-dependent reductions in SDH, alkaline phosphatase and acid phosphatase activities in the kidney of mice. Vitamin E cotreatment as well as, 30 d withdrawal of arsenic trioxide treatment with or without vitamin E caused significant amelioration in arsenic-induced toxicity in mice. Administration of vitamin E during withdrawal of treatment also caused significant amelioration as compared from only withdrawal of the treatment. It is concluded that vitamin E ameliorates arsenic-induced toxicities in the liver and kidney of mice. PMID- 15272722 TI - Effects of traffic pollution on the genetic structure of Poa annua L. populations. AB - The genetic composition of Poa annua L. populations with a series of traffic pollution was studied by starch electrophoresis. Five enzyme systems were stained. The results showed that: (1) Traffic pollution can dramatically change genotypic frequencies at some loci of P. annua populations. Significant deviations from Hardy-Weinberg equilibrium were observed on loci Fe-1 and Me due to the excess of heterozygotes in some populations. (2) The effective number of alleles per locus and the observed and expected heterozygosity were higher in the pollution series than in the clear control site (Botanic Park population), but the increase was not related with the pollution extent. (3) Most genetic variation was found within populations, and only 6.21% was among populations of the polluted series. Slightly higher differentiation (FST = 7.98%) was observed when the control population was included. (4) The calculated gene flow (Nm) is 2.8841 per generation. The mean of genetic identity is 0.9864 and the genetic distance average to 0.0138. PMID- 15272723 TI - Effect of inoculation with arbuscular mycorrhizal fungi on the degradation of DEHP in soil. AB - The effect of inoculation with arbuscular mycorrhizal (AM) fungi (Acaulospora lavis) on the degradation of di(2-ethylhexyl) phthalate (DEHP) in soil was studies. Cowpea plants (Pigna sinensis) were used as host plants and grown in a specially designed rhizobox. The experimental results indicated that, both in sterile and non-sterile soil, mycorrhizal colonization rates were much higher in the mycorrhizal plants than in the non-mycorrhizal plants. Addition of 4 mg/kg DEHP slightly affected mycorrhizal colonization, but the addition of 100 mg/kg DEHP significantly decreased mycorrhizal colonization. DEHP degradation in the mycorrhizosphere (Ms) and hyphosphere (Hs), especially in the Hs, increased after inoculation with Acaulospora lavis. It is concluded that mycorrhizal hyphae play an important role in the plant uptake, degradation and translocation of DEHP. The mechanism might be attributed to increased numbers of bacteria and actinomycetes and activity of dehydrogenase, urease and acid phosphatase in the Ms and Hs by mycorrhizal fungi. PMID- 15272724 TI - Removal of NOx from flue gas with radical oxidation combined with chemical scrubber. AB - In this paper, removal of NOx (namely DeNOx) from flue gas by radical injection combined with NaOH solution (26% by weight of NaOH in water) scrubbing was investigated. The experimental results showed that the steady streamer corona occurs through adjusting the flow rate of the oxygen fed into the nozzles electrode. The vapor in the oxygen has influence on the V-I characteristics of corona discharge. Both HNO2 and HNO3 come into being in the plasma reactor and the DeNOx efficiency in the plasma reactor is more than 60%. The overall DeNOx efficiency of the whole system reaches 81.7% when the NaOH solution scrubbing is collaborated. PMID- 15272725 TI - Environmental impact of aquaculture-sedimentation and nutrient loadings from shrimp culture of the southeast coastal region of the Bay of Bengal. AB - Nutrient loadings were measured for surface seawater and bottom sediments of semi intensive and improved extensive shrimp culture pond, adjacent estuary, and fallow land in the south-east coastal region of Bangladesh during August, 2000 January, 2001 to evaluate the impact of shrimp culture. The mean levels of nutrients found in the pond surface water were 108.780 mg/L for CaCO3, 0.526 mg/L for NH4+ -N, 3.075 wt% for organic carbon, 7.00 mg/L for PO4-P, 5.57 mg/L for NO3 N, and 7.33 mg/L for chlorophyll-a. The maximum mean value of H2S (0.232 mg/L) was found in estuarine water. Nutrients loading were found to be decreased with distance from the shrimp farm discharge unit in estuarine water. The mean level of organic matter, total nitrogen, and organic carbon were found in higher concentrations in sediments of cultured pond compared to bottom soil of adjacent fallow land at the same elevation. Extractable Ca values were found in higher concentration (550.33 ppt) in adjacent fallow land, as the shrimps for molting in shrimp ponds use extractable Ca. The relation between seawater H2S value and sediment pH (r = - 0.94); sediment organic carbon and sediment pH values (r = 0.76), sediment total nitrogen and sediment pH (r = -0.74) were found to be highly negatively correlated. Whereas the relation between seawater H2S value and sediment total nitrogen (r = 0.92), water NH4+ -N and sediment pH (r = 0.66) were found to be positively correlated. The results revealed that load of nutrients at eutrophic level in estuarine water, and decrease of soil pH; leading to acid sulphate soil formation indicates a negative impact of shrimp culture. PMID- 15272726 TI - Binary competitive adsorption of naphthalene compounds onto an aminated hypercrosslinked macroporous polymeric adsorbent. AB - An aminated hypercrosslinked macroporous polymeric adsorbent was synthesized and characterized. Adsorption isotherms for 1-amino-2-naphthol-4-sulfonic acid (1,2,4 acid) and 2-naphthol obtained from various binary adsorption environments can be well fitted by Freundlich equation, which indicated a favorable adsorption process in the studied range. Adsorption for 1,2,4-acid was an endothermic process in comparison with that for 2-naphthol of an exothermic process. 2 naphthol molecules put a little influence on the adsorption capacity for 1,2,4 acid. However, the adsorption to 1,2,4-acid depressed that to 2-naphthol in a large extent for the stronger electrostatic interaction between 1,2,4-acid and adsorbent. The predominant mechanism can be contributed to the competition for adsorption sites. And the simultaneous environment was confirmed to be helpful to the selective adsorption towards 1,2,4-acid based on the larger selectivity index. PMID- 15272727 TI - Performance of a subsurface-flow constructed wetland in southern China. AB - The operational performance of a full-scale subsurface-flow constructed wetland, which treated the mixed industrial and domestic wastewater with BOD5/COD mean ratio of 0.33 at Shatian, Shenzhen City was studied. The constructed wetland system consists of screens, sump, pumping station, and primary settling basin, facultative pond, first stage wetland and secondary stage wetland. The designed treatment capacity is 5000 m3/d, and the actual influent flow is in the range of < 2000 to > 10000 m3/d. Under normal operational conditions, the final effluent quality well met the National Integrated Wastewater Discharge Standard (GB 8978 1996), with the following parameters(mean values): COD 33.90 mg/L, BOD, 7.65 mg/L, TSS 7.92 mg/L, TN 9.11 mg/L and TP 0.56 mg/L. Seven species of plants were selected to grow in the wetland: Reed, Sweetcane flower Silvergrass, Great Bulrush, Powdery Thalia and Canna of three colours. The growing season is a whole year-round. The seasonal discrepancy could be observed and the plants growing in the wetland are vulnerable to lower temperature in winter. The recycling of the effluent in the first stage of the wetland system is an effective measure to improve the performance of the wetland system. The insufficient DO value in the wetland system not only had significant effect on pollutants removal in the wetland, but also was unfavourable to plant growth. The recycling of effluent to the inlet of wetland system and artificial pond to increase DO value of influent to the wetland is key to operate the subsurface constructed wetland steadily and effectively. PMID- 15272728 TI - A novel method to synthesize polyaluminum chloride with a membrane reactor. AB - Al13 or Alb is usually regarded as the most efficient species of polyaluminum chloride (PAC), the performance flocculant for water treatment. This paper was intended to report a new method to synthesize PAC with high content Alb, by using the membrane reactor. NaOH solutions were managed to permeate slowly through the micropores of ultrafiltration membrane into AICl3 solutions under the suitable transmembrane pressure(TMP). Meanwhile NaOH drops size was limited to nano-scale, resulting in dramatical reduction of the characteristic diffusion time and great increment of contact interface between the strong base and Al ions in solution to favor the formation of Al(OH)4-, the precursor of Al13, so few precipitates and much Alb are produced. When the initial concentration of AlCl3/NaOH is 0.40/2.0 (mol/L), MWCO = 10000, TMP = 0.0085 MPa, T = 305 K and B (molar ratio of OH /Al3+) = 2.25, the quantity of Alb attains about 80%. The results of 27Al-NMR determination showed that the Al13 content is equal to Alb content. And our PAC product has shown better flocculation effects than the commercial product. PMID- 15272729 TI - Cometabolic microbial degradation of trichloroethylene in the presence of toluene. AB - Trichloroethylene (TCE), a common groundwater pollutant, was cometabolized by microorganisms in the presence of toluene as a growth substrate. The effect of concentrations of toluene and TCE and temperature on biodegradation was discussed. Acclimated microorganisms degraded TCE after a lag period of 5 to 22 h depending on toluene concentrations. Approximately 60%, 90% and 64% of TCE were degraded at toluene to TCE concentration ratios of 23:1, 115:1 and 230:1, respectively. At a TCE concentration of 1.46 microg/ml, 80% of TCE and 98.4% of toluene were removed. But less degradation of TCE and toluene was observed when TCE concentration was above 48.8 microg/ml. The lag time of TCE decreased and the TCE biodegradation rates increased with the increase of temperature. PMID- 15272730 TI - Landscape eco-environmental research on littoral zone in China. AB - Littoral zone is a special land/landscape type. As an important kind of land resource in support, the use of littoral zone is vital to eastern coastal areas in China. And the research on littoral zone relates to the key theory of landscape ecology. Based on the theory of landscape ecology, the littoral zone was divided into four types: mud flat, sand beach, bench, and biological flat. The distribution of each type in China is pointed out. As a typical open system, littoral zone has six landscape ecological characteristics: (1) high sensitivity to disturbance; (2) distinct edge effect; (3) spatial aggregation of natural resources; (4) frequently spatial oscillation; (5) obviously spatial heterogeneity; and (6) noticeably spatial differentiation. Some proposals are also put forward on the land use and development of littoral zone for environmental protection and environmental management. PMID- 15272731 TI - Determination of volatile organic compounds in river water by solid phase extraction and gas chromatography. AB - A simple, rapid, and reproducible method is described employing solid-phase extraction (SPE) using dichloromethane followed by gas chromatography (GC) with flame ionization detection (FID) for determination of volatile organic compound (VOC) from the Buriganga River water of Bangladesh. The method was applied to detect the benzene, toluene, ethylbenzene, xylene and cumene (BTEXC) in the sample collected from the surface or 15 cm depth of water. Two-hundred ml of n hexane-pretreated and filtered water samples were applied directly to a C18 SPE column. BTEXC were extracted with dichloromethane and average concentrations were obtained as 0.104 to 0.372 microg/ml. The highest concentration of benzene was found as 0.372 microg/ml with a relative standard deviation (RSD) of 6.2%, and cumene was not detected. Factors influencing SPE e.g., adsorbent types, sample load volume, eluting solvent, headspace and temperatures, were investigated. A cartridge containing a C18 adsorbent and using dichloromethane gave better performance for extraction of BTEXC from water. Average recoveries exceeding 90% could be achieved for cumene at 4 degrees C with a 2.7% RSD. PMID- 15272732 TI - Effect of atmospheric precipitation on the dissolved loads of the Dongjiang River, China. AB - The atmospheric precipitation plays an important role in influencing the river chemistry of the Dongjiang River. The atmospheric contribution to river water is estimated by reference to Cl concentration called Clref. The Clref of 41.97 micromol/L represents the highest chloride concentration of the rainwater inputs to river water, thus sea salts are responsible for total Cl concentration of the Dongjiang River. According to the principal compositions of precipitation and river water, two approaches-sea salt correction and precipitation correction were proposed in order to correct the contribution proportions of atmospheric precipitation on the solutes and to calculate chemical weathering rate. The results reflected that the atmospheric contribution ratios fluctuate from approximately 5% to approximately 20% of TDS (total dissolved solids) in the Dongjiang River. As compared with the other world watersheds, the lower dissolved ion contents and high runoff may result in the obvious influence of precipitation on river chemistry in the Dongjiang basin. The major elemental chemistry is mainly controlled by silicate weathering, with the anion HCO3- and cation Ca2+ and Na+ dominating the major compositions in this basin. The estimated chemical weathering rate of 15.78-23.48 t/(km2 x a) is only 40%-60% of a global average in the Dongjiang basin. Certainly, the estimated results are still under correction gradually because the effect of human activities on the precipitation chemistry has never been quantified in detail. PMID- 15272733 TI - Research on buildings impacting on aerosol diffusing in urban area using remote sensing. AB - Employing remote sensing method to interpret the building volumetric ratio and aerosol status in Guangzhou, China. The relation between them and identified characteristics of their spatial distribution was analyzed. Results showed that building density and aerosol status are strongly correlated. It is indicated that the resistance of building to aerosol diffusing is one of factors influencing air pollution in urban area. On the basis of calculated results, building voluminous ratio of 5.6 is taken as the threshold impacting on aerosol diffusing, so the building voluminous ratio of Guangzhou should be limited to less than 5.6 in order to alleviate air pollution. PMID- 15272734 TI - Effect of linear alkyl benzene sulfonates (LAS) on the fate of phenanthrene in a model ecosystem (water-lava-plant-air). AB - Advanced closed chamber system was used to study the fate of phenanthrene (3 rings PAHs) in the presence of linear alkylbenzene sulphonates (LAS). The results showed mineralization and metabolism of phenanthrene are fast in the "culture solution-lava-plant-air" model ecological system. The distribution proportions of applied 14C-activity in this simulative ecological system were 41%-45%, 14% to 10% and 1% in plant, lava and culture solution respectively, and 18% to 29%, 11% to 8% recovered in the forms of VOCs and CO2. Main parts of the applied 14C activity exist in two forms, one is polar metabolites (25%) which mainly distribute in the root (23%), the other is unextractable part (23%) which have been constructed into plant root (8.98%), shoot (0.53%) or bonded to lava (13.2%). The main metabolites of phenanthrene were polar compounds (25% of applied 14C-activity), and small portion of 14C-activity was identified as non polar metabolites (6% of applied 14C-activity) and apparent phenanthrene (1.91% of applied 14C-activity). Phenanthrene and its metabolites can be taken up through plant roots and translocated to plant shoots. The presence of LAS significantly increased the the concentration of 14C-activity in the plant and production of VOCs, at the same time it decreased the phenanthrene level in the plant and the production of CO2 at the concentration of 200 mg/L. PMID- 15272735 TI - Two stages kinetics of municipal solid waste inoculation composting processes. AB - In order to understand the key mechanisms of the composting processes, the municipal solid waste (MSW) composting processes were divided into two stages, and the characteristics of typical experimental scenarios from the viewpoint of microbial kinetics was analyzed. Through experimentation with advanced composting reactor under controlled composting conditions, several equations were worked out to simulate the degradation rate of the substrate. The equations showed that the degradation rate was controlled by concentration of microbes in the first stage. The degradation rates of substrates of inoculation Run A, B, C and Control composting systems were 13.61 g/(kg x h), 13.08 g/(kg x h), 15.671 g/(kg x h), and 10.5 g/(kg x h), respectively. The value of Run C is around 1.5 times higher than that of Control system. The decomposition rate of the second stage is controlled by concentration of substrate. Although the organic matter degradation rates were similar to all Runs, inoculation could reduce the values of the half velocity coefficient Km and could be more efficient to make the composting stable. Particularly, for Run C, the degradation rate is high in the first stage, and Km is low in the second stage. The results indicated that the inoculation was efficient for the composting processes. PMID- 15272736 TI - Degradation of phenol in an upflow anaerobic sludge blanket (UASB) reactor at ambient temperature. AB - A synthetic wastewater containing phenol as sole substrate was treated in a 2.8 L upflow anaerobic sludge blanket (UASB) reactor at ambient temperature. The operation conditions and phenol removal efficiency were discussed, microbial population in the UASB sludge was identified based on DNA cloning, and pathway of anaerobic phenol degradation was proposed. Phenol in wastewater was degraded in an UASB reactor at loading rate up to 18 gCOD/(L x d), with a 1:1 recycle ratio, at 26 +/- 1 degrees C, pH 7.0-7.5. An UASB reactor was able to remove 99% of phenol up to 1226 mg/L in wastewater with 24 h of hydraulic retention time (HRT). For HRT below 24 h, phenol degradation efficiency decreased with HRT, from 95.4% at 16 h to 93.8% at 12 h. It further deteriorated to 88.5% when HRT reached 8 h. When the concentration of influent phenol of the reactor was 1260 mg/L (corresponding COD 3000 mg/L), with the HRT decreasing (from 40 h to 4 h, corresponding COD loading increasing), the biomass yields tended to increase from 0.265 to 3.08 g/(L x d). While at 12 h of HRT, the biomass yield was lower. When HRT was 12 h, the methane yield was 0.308 L/(gCOD removed), which was the highest. Throughout the study, phenol was the sole organic substrate. The effluent contained only residual phenol without any detectable intermediates, such as benzoate, 4-hydrobenzoate or volatile fatty acids (VFAs). Based on DNA cloning analysis, the sludge was composed of five groups of microorganisms. Desulfotomaculum and Clostridium were likely responsible for the conversion of phenol to benzoate, which was further degraded by Syntrophus to acetate and H2/CO2. Methanogens lastly converted acetate and H2/CO2 to methane. The role of epsilon-Proteobacteria was, however, unsure. PMID- 15272737 TI - Cysticercosis: IgG-ELISA evaluations of peak1 antigen and <30 kDa antigen of delipidized extract of Taenia solium metacestodes. AB - The antigenicity of ether-delipidized Taenia solium metacestode extract (DLPAg) was investigated by IgG-ELISA. The antigen showed higher antigenicity than that of non-delipidized antigen (NDLPAg). Then the DLPAg was subjected to Sephacryl S 200 gel chromatography and a partially purified antigen (DLPP1Ag) was identified as the promised antigen by IgG-ELISA using 25 sera from cysticercosis cases, 177 cases of 24 heterologous infections, and healthy controls. Sensitivity was 52% and specificity was 91.8% at the cut-off value (X + 7SD), 0.399. Cross-reactivity occurred with 17 cases of eight diseases: cystic echinococcosis (7/11), taeniasis (1/16), gnathostomiasis (2/8), strongyloidiasis (1/12), angiostrongyliasis (1/12), paragonimiasis heterotremus (2/15), opisthorchiasis (1/9) and fascioliasis (2/7). When DLPP1Ag was fractionated through Ultra free centrifugal tube (retained 30 kDa) and Amicon (PM10), MWCOP1Ag (<30-10> kDa) was obtained; the antigen showed better results than DLPP1Ag with 88% sensitivity and 95.6% specificity at the cut-off value (X + 4SD), 0.264. Nine cases of six diseases cross-reacted with this antigen: cystic echinococcosis (2/11), gnathostomiasis (2/8), trichinellosis (2/12), toxocariasis (1/5), schistosomiasis (1/6), and fascioliasis (1/7). MWCOP1Ag gave higher sensitivity than that of DLPP1Ag but some cross-reactivity occurred. PMID- 15272738 TI - A comparative study on mouse MHC class I sequences detected in Schistosoma japonicum recovered from BALB/c (H-2d) and C57BL/6 (H-2b) mice. AB - The mouse major histocompatibility complex (MHC) class I sequence was detected in all the 8-week-old Schistosoma japonicum recovered from BALB/c (H-2d) and C57BL/6 (H-2b) mice by in situ polymerase chain reaction (in situ PCR). The signals of the mouse class I MHC sequence were observed in the nuclei of the mesenchymal and reproductive cells of 8-week-old S. japonicum. Furthermore, the class I MHC sequence was detected in each DNA extracted from S. japonicum cercariae maintained in BALB/c and C57BL/6 mice by nested PCR. To prove both horizontal and vertical transmission of this sequence in schistosomes, we have used cercariae obtained from parasites maintained in BALB/c mice to infect C57BL/6 and BALB/c mice, and vice versa. The MHC sequences from adult worms were compared to the cercarial MHC and host MHC sequences. Nucleotide sequence comparisons between adult worm DNA, host (H-2d and H-2b mice) DNA and cercarial DNA used for the infection suggested that the sequence of mouse class I MHC was incorporated into schistosome adults and inherited throughout their life-cycle. PMID- 15272739 TI - Parasites detected from diarrheal stool samples collected in Nepal. AB - Intestinal parasites were investigated in 396 diarrheal stool samples collected from individuals aged 1 to 68 years (males: 239 and females: 157) in Nepal. Samples were collected at different medical centers located in Kathmandu and from two public schools in a village setting in Kathmandu Valley and outside, during October 1999 to January 2001. The stool samples were mixed with 2% dichromate solution and transported to Japan for investigations. Parasites were detected by employing the formal-ether sedimentation technique. Of a total of 396 fecal samples investigated, 193 (49%) were positive for some kind of parasite. Altogether, 15 species of parasites were detected. Giardia intestinalis topped the list of protozoa, whereas Trichuris trichiura was the most frequently detected among helminth parasites. Of the 193 positive samples, 109 (56%) had single parasite infections, whereas 84 (43%) had multiple infections with a maximum of five species. Of the total positive, 45 (23%) had both protozoa and helminths whereas 37 (19%) had only protozoa. Females (52%) and children (15 years and under) (52%) had a marginally higher prevalence compared with males (46%) and adults (45%), respectively (p > 0.05). Samples collected from two public schools in a village setting inside Kathmandu Valley and outside had a significantly higher positive rate compared with those observed in individuals visiting different medical centers in the city and suburban areas in Kathmandu (p < 0.05). PMID- 15272740 TI - Review on human toxoplasmosis in Malaysia: the past, present and prospective future. AB - We reviewed various studies regarding human toxoplasmosis in Malaysia. They showed a varying prevalence of specific Toxoplasma antibodies among the Malaysian population. The Malays have shown the highest seroprevalence of toxoplasmosis, by most studies, when compared to other races. Demographic profiles have shown that Toxoplasma seropositivity is higher in males than females, lower in people with higher incomes, higher in the unemployed and tends to increase with age. In general, the route of transmission, such as contact with a cat, consumption of undercooked meat and blood transfusion were shown to have no significant association with Toxoplasma seropositivity (p > 0.05). The immune status (CD4 cell count < 200 cell/mm3) was strongly associated with toxoplasmic encephalitis (p < 0.05). PMID- 15272741 TI - Association of intestinal helminths with decreased liver size and sCD23 concentration during falciparum malaria. AB - To determine if intestinal helminths and the CD23/nitric oxide pathway had an influence on liver size, we conducted a cross-sectional study on 438 patients with confirmed P. falciparum malaria admitted at the Hospital for Tropical Diseases in Bangkok. For all patients the liver size was measured as number of centimeters below the rib cage, a stool examination was conducted, and CD23 and reactive nitrogen intermediates were measured. The median liver size was smaller in helminth-infected patients than in helminth-free patients (chi2 for trend = 9.1, p = 0.003). Liver size significantly increased with the concentration of sCD23 (p < 0.0001). The median sCD23 concentration (OD) was significantly lower in helminth-infected patients than in helminth-free patients, respectively 0.33 (quartiles 0.24-0.57) and 0.45 (quartiles 0.27-0.59), (p = 0.01). There was a negative correlation between sCD23 concentrations and RNI (Spearman's rho = 0.40, p < 0.0001). All the above results remained significant after controlling for potential confounders. These results are compatible with a CD23/NO-mediated decrease in liver size in helminth-infected patients. PMID- 15272742 TI - Evaluation of the KAT-Quick Malaria Rapid Test for rapid diagnosis of falciparum malaria in Thailand. AB - In recent years, several rapid diagnostic tests for falciparum malaria have been developed. KAT test results were compared with microscopy on 90 consecutive patients hospitalized at the Hospital for Tropical Diseases, Bangkok, Thailand. Fifty-one patients had P. falciparum infections while 49 had malaria due to other plasmodium species. For a geometric mean +/-SD (Min;Max;range) parasitemia of 11,481 +/- 5.0 (88;713,838;713,750), the sensitivity of the KAT test was 96% (95% CI = 86-99.5), the specificity was 92% (95% CI = 80-99), the accuracy was 94% and the reliability was 85%. These findings suggest that the KAT test is of potential interest in the diagnosis of falciparum malaria in Thailand. PMID- 15272743 TI - A mixture model application in disease mapping of malaria. AB - Disease mapping, a method for displaying the geographical distribution of disease occurrence, has received attention for more than 2 decades. Because traditional approaches to disease mapping have some deficiencies and disadvantages in presenting the geographical distribution of disease, the mixture model--as an alternative approach--overcomes some of these deficiencies and provides a clearer picture of the spatial risk structure. The purpose of this study was twofold: (1) to investigate the geographical distribution of malaria in Thailand during 1995, 1996, and 1997 by applying the mixture model to disease mapping, and (2) to investigate the dynamic nature of malaria in Thailand during the 3-year time frame by applying the space-time mixture model. Non-parametric maximum likelihood estimation was employed to estimate the parameters of both the mixture model and the space-time mixture model. Applying Bayes' theorem, the 76 provinces of Thailand were classified into component risk levels by the rate of malaria for each province. Malaria intensively occurred in 4 provinces on the Thai-Myanmar border and in 2 provinces on the Thai-Cambodian border. Of the 76 provinces studied, 10 showed an increasing trend over the 3-year period. A comparison of the map based on the mixture model with the map based on the traditional percentiles method indicates that the non-parametric mixture model removes random variability from the map and provides a clearer picture of the spatial risk structure. The advantage of the mixture model approach to disease mapping is the graphical visual presentation of the prevalence of disease. The space-time mixture model more adequately investigates the dynamic nature of disease than does the mixture model. PMID- 15272744 TI - Malaria infection among the migrant population along the Thai-Myanmar border area. AB - A population based case-control study was performed to determine factors associated with malaria infection among the migrant population, foreign nationals aged 15 years or over. Data were obtained from 217 malaria and 217 non-malaria patients attending the Vector-Borne Disease Control Units 6-9 (Thong Pha Phum and Sangkhla Buri districts) in Kanchanaburi Province and at the Vector-Borne Disease Control Units 1,9 (Mae Fa Luang and Mae Sai districts) in Chiang Rai Province, between June and December 2002. All study subjects were interviewed by trained interviewers using a structured interview form. The statistical analysis was carried out by the chi-square test and multivariate logistic regression: a p value of less than 0.05 was considered to be statistically significant. The results showed that the study subjects were predominantly Thai-Yai and Myanmar. Plasmodium falciparum was the major type of the malaria (60.8%). Logistic regression analysis, controlling for possible confounding factors, revealed that residence located in the forest increased the risk of malaria infection by a factor of 6.29 (OR = 6.29, 95% CI = 1.56-25.42); outdoor stay < 7 and > 7 days prior to the blood examination also increased the risk by a factor of 4.34 and 4.13 respectively (OR = 4.34, 95% CI = 1.05-17.99; OR = 4.13, 95% CI = 1.29 13.13). PMID- 15272745 TI - Knowledge, attitudes and practices with regard to malaria control in an endemic rural area of Myanmar. AB - Malaria is a global health problem, in particular, a major health problem within Southeast Asia. This study aimed to investigate malaria control within a rural area of Myanmar, where traditionally non-western medicine is the preferred treatment. Whilst malaria was perceived by the local people to be a major health problem, knowledge about the mode of transmission and correct treatment for malaria was relatively low. Consequently, the practices of the local people to control malaria were often ill-informed or based on cultural and traditional beliefs. PMID- 15272746 TI - Impact of a filariasis control program on intestinal helminthic infections; a pilot study in Narathiwat Province, Thailand. AB - This study was conducted in 9 villages located in endemic areas for brugian filariasis in Narathiwat Province, Thailand. Parasitological and anthropometric examinations were cross-sectionally performed to assess the prevalence of intestinal parasitic infections of 539 villagers. Paired stool samples were collected before and after mass treatment for the filariasis control program in 150 participants in order to study the impact of the filariasis control program on intestinal helminthiasis. The results found that 50.3% of the villagers were infected with one or more types of intestinal parasites. Double and triple infections were found in 10.9% and 1.6% of infected individuals respectively. The prevalence of intestinal parasitic infections peaked in the 1-10 year old age group, which are pre-school and young school-age children. A significant reduction of intestinal helminthic infections in the post-treatment stool sample was observed in the 150 participants who were examined six months after mass treatment. Integrating an intestinal helminthic control program alongside the existing filariasis control program would be an appropriate and cost-effective strategy in the control of intestinal helminths. However, reinfection of parasites was observed. PMID- 15272747 TI - Space spraying of bacterial and chemical insecticides against Anopheles balabacensis Baisas for the control of malaria in Sabah, East Malaysia. AB - A pilot study was undertaken to determine the effectiveness of space application of insecticides for the control of malaria in Ranau, a district in Sabah. A village each was treated monthly: with chemical adulticide--alpha cypermethrin (Fendona SC(R)/10SC(R)) at 2 g a.i./10,000 m2 in Pahu; with biological larvicides -Bacillus thuringiensis israelensis (Vectobac 12AS(R)) at 500 ml/10,000 m2 or B. sphaericus (Vectolex WG(R)) at 500 g/10,000 m2 in Pinawantai; and with a mixture of chemical adulticide and biological larvicide in Togop Laut. All sprayings were conducted using a portable mist blower. During the study period all villages, including Tarawas the untreated village, received the conventional malaria control measures. Entomological and epidemiological surveillance was used to measure the effectiveness of the space application. The entomological surveillance indicated that the An. balabacensis population was significantly reduced by alpha cypermethrin in Pahu and Togop Laut and B. sphaericus in Pinawantai; but was not reduced by B.t.i. in Pinawantai. There was a significant reduction in the number of malaria cases and in the slide positivity rate in the treated villages during the study period. The pilot study does indicate that space application of larvicides/adulticides or a mixture of both is able to reduce the malaria vector population and the malaria transmission. A larger scale study needs to be undertaken in a malarious village/province to determine whether space application of insecticides together with other malaria control measures will be able to eradicate malaria. PMID- 15272748 TI - Effect of ten chlorophytes on larval survival, development and adult body size of the mosquito Aedes aegypti. AB - The effect of ten microalgal chlorophytes isolated from mosquito breeding containers on the survival, larval development and adult body size of the mosquito Aedes aegypti was investigated. All larvae fed with six of the microalgal isolates died after 7 days. These isolates were found to be resistant to digestion by mosquito larvae. Delayed pupation and body size reduction of the mosquitos fed with Chlorococcum UMACC 218 and Scenedesmus UMACC 220 were observed. In contrast, larvae fed with Ankistrodesmus convolutus UMACC 101 and Chlorococcum UMACC 213 were bigger in size than those fed with normal insectory feed. The present study showed that microalgal chlorophytes have the potential to be used as larvicidal agents for mosquitos. PMID- 15272749 TI - Correlation studies on Widal agglutination reaction and diagnosis of typhoid fever. AB - A total of 80 patients at the University of Uyo Teaching Hospital (UUTH) suspected of having enteric infections were screened for the presence of Salmonella species using blood, urine and stool samples along with Widal agglutination tests. Although 39 patients tested positive for the Widal agglutination test with titers ranging from 1:80 to 1:320, no Salmonella organism was encountered in some cultures. Statistical analysis revealed significant differences (chi2) at the 5% probability level between the Widal test and the cultures of the clinical samples. The results suggest that serological investigations alone may not be a reliable index for the diagnosis of Salmonella infections. PMID- 15272750 TI - Research note: Molecular subtyping of Salmonella enterica serovar Tshiongwe recently isolated in Malaysia during 2001-2002. AB - Pulsed field gel electrophoresis (PFGE) and antimicrobial susceptibility analysis were undertaken on twenty-three strains of Salmonella enterica serovar Tshiongwe, an unusual serovar, which recently emerged in Malaysia. Antimicrobial susceptibility analysis showed that all the strains were sensitive to ampicilin, chloramphenicol, cotrimoxazole, and kanamycin. Twenty (87%) and 8 (3.5%) strains had resistance to tetracycline and streptomycin respectively. PFGE analysis subtyped 23 strains into 10 profiles (Dice coefficient of similarity, F = 0.7 1.0). The predominant profile, X1 was found in both clinical and environmental isolates and was widely distributed in different parts of Malaysia during the study period. In addition, isolates recovered from food, a hand-towel, apron and the surface of a table-top in one particular location had unique, indistinguishable profiles (X4/4a) and identical antibiograms. Similarly, isolates from cooked meat and a chopping board had PFGE profiles similar to some human isolates. These probably indicated cross-contamination and poor hygiene in food practices, hence contributing to Salmonellosis. Factors causing the emergence of this rare Salmonella serovar being responsible for food poisoning episodes during the study period remained unclear. The study reiterated the usefulness and versatility of PFGE in the molecular subtyping of this rare Salmonella serovar in Malaysia. PMID- 15272751 TI - Perceptions of Shigella and of Shigella vaccine among rural Chinese: compatibility with Western models of behavioral change. AB - Shigella remain a major source of morbidity and mortality in developing countries, including China. In response, national and international researchers are actively working to develop vaccines that will be effective against dysentery and diarrhea caused by shigella dysentariae. With the growing recognition of the problems associated with sustained vaccine acceptance and usage, researchers and policy makers recognize the importance of conducting theory-based qualitative research to inform vaccine development program efforts. Accordingly we undertook this qualitative study involving 81 residents of one of China's rural communities with high rates of dysentery. The semi-structured interviews suggest that a Western model of behavioral change offered a useful research construct. Consistent with the model is the community's strong perception of 'response efficacy' of vaccines, particularly in comparison with water and sanitation and disease treatment. Residents were eager to vaccinate their children despite variable perception of disease severity, while they were less consistent in their interest in vaccinating adults; this enthusiasm for vaccinating children was attributed to China's 'one child per couple' policy. Intervention implications are discussed. PMID- 15272752 TI - Case report: Brucellosis: a re-emerging disease in Thailand. AB - Brucellosis is a zoonotic disease prevalent in many countries, but it has been reported only once in Thailand, 36 years ago. We describe here two consecutive cases of brucellosis in Bangkok, Thailand. Both cases presented with prolonged fever and weight loss. Blood cultures taken from 2 patients yielded Brucella melitensis. The slide agglutination test of blood samples were also positive, with a titer of 1:64 for antibodies to Brucella. The first patient responded to a combination of doxycycline, gentamicin, and ciprofloxacin; the other responded to doxycycline and rifampicin. Brucellosis is a potential public health threat, therefore, preventive measures should be actively implemented. This clinical syndrome should be included in the differential diagnosis of patients presenting with prolonged fever, particularly those with contact to animals which could serve as reservoirs. PMID- 15272753 TI - A new member of the trombiculid mite family Neotrombicula nagayoi (Acari: Trombiculidae) induces human dermatitis. AB - We present the first definitive evidence that the mite Neotrombicula nagayoi bites humans under natural conditions in Japan. Initially, bites resulted in mild pruritus without pain. However, skin reactions increased gradually year by year with severe pruritus with pain being reported by the victim after being bitten repeatedly. Six species of trombiculid mites comprising three genera were isolated from soil samples collected from August to October in both 2001 and 2002 at a study site where a man was bitten by N. nagayoi. The dominant species was L. intermedium (72.4%) followed by L. pallidum (8.3%) and N. nagayoi (8.1%). N. nagayoi was found only in August and September. We did not detect the pathogen Orientia tsutsugamushi in any of the unfed larvae, including N. nagayoi, collected from the soil samples. PMID- 15272754 TI - Aspergillosis of the central nervous system: a catastrophic opportunistic infection. AB - The clinical features and outcome of the treatment of aspergillosis of the central nervous system (CNS) in Thai patients are presented. The patients who were diagnosed as having CNS aspergillosis by tissue biopsy or culture from January 1, 1991 to December 31, 2000 were retrospectively reviewed. The study variables including age, sex, underlying disease, symptoms and signs, neuro imaging studies, pathological findings and outcome of treatment, are described. There were seven cases of aspergillosis of the central nervous system. Four patients were male. The median age was 65 years (range 36-78 years). The most common underlying disease was diabetes mellitus (4/7; 57.1%). Two patients (28.6%) had no underlying disease. The most common primary site of infection was the paranasal sinuses (6/7; 85.7%). The most common clinical presentation was headache (6/7; 85.7%). Common neurological signs included multiple cranial nerve palsies (5/7; 71.4%) and alteration of consciousness (3/7; 42.9%). The median duration of the symptoms prior to admission was 60 days (range 8-180 days). All patients were treated with intravenous antifungal agents at high doses. Extensive surgery was performed in 6 patients. The mortality rate was very high (6/7; 85.7%). The median time from diagnosis and treatment to death was 53 days (22-720 days). Aspergillosis of the CNS should be considered in those with clinical features of headache, multiple cranial nerve palsies and alteration of consciousness accompanied by sinusitis, especially in elderly and diabetic patients. It remains a catastrophic opportunistic infection in spite of the current intensive and aggressive treatment. PMID- 15272755 TI - Gender difference in treatment seeking behaviors of tuberculosis cases in rural communities of Bangladesh. AB - This descriptive cross-sectional study was conducted to investigate gender differences in the epidemiological factors associated with the treatment seeking behaviors of TB cases in the rural communities of Bangladesh. The study reveals that there is significant gender difference in treatment seeking behaviors of rural TB cases and the majority of them (52%) have taken prior treatment from various traditional healers, 70% of them are females who attended health centers (UZHCs) as the other choice (adjusted OR: 4.2, 95% CI: 2.0-8.4). It was found that the mean patient delay was 63 days (range 14-210 days) where half of the females delayed more than 60 days while they were spreading their disease. The study findings reveal gender differences in treatment seeking behaviors associated with socio-cultural barriers, particularly among females in their access to TB care. Fifty-five percent of cases wanted the diagnosis of TB remain confidential to avoid being labeled as TB patients, where 82.7% were female, 85.6% of female TB patients had problems in their relationships with their spouse (61%) and family members (58%) after being diagnosed with TB. The results of the TB service factors found that 39% of females were not satisfied with their provider's behaviors, which was significantly associated with treatment seeking behavior (adjusted OR: 2.6, 95% CI: 1.0-6.6). The study findings strongly suggest that there was a significant gender difference in treatment seeking behavior in rural Bangladesh. Based on the study findings, we recommend developing an appropriate gender strategy for developing a TB control program, comprised of operational, socio-cultural and community awareness interventions aimed at treating undiscovered reservoirs of female TB cases in rural Bangladesh. PMID- 15272756 TI - A subset of clinical status of pulmonary tuberculosis in Southern Taiwan. AB - The aims of this study were to present the clinical status of pulmonary tuberculosis in Southern Taiwan and to analyze the reasons for failure of antituberculosis treatment in order to achieve a higher rate of success after treatment. Two hundred and senventeen adult patients, aged 15 to 90 years old who presented to the Chest Division, Kaohsiung Medical University Hospital from 1999 to 2002 with a diagnosis of Pulmonary Tuberculosis, were retrospectively studied. We compared the rate of recurrence of pulmonary tuberculosis by dividing the cases into 2 groups: those who completed treatment and those who did not. We also determined the age distributions for when initial diagnosis of pulmonary tuberculosis was made among these 217 cases. In 90 culture proven cases, antituberculosis drug susceptibility was tested to determine the rate of drug resistance. We also assessed the reasons for failure of treatment. Age distribution analysis showed that initial infection began at a young age, was widely spread, and occurred regardless of age. There were 116 cases that completed antituberculosis treatment and 101 cases that did not. Of the 116 cases, only 16 relapsed, whereas 79 of the 101 cases relapsed. In cases where completely treated patients relapsed, the period before recurrence was indefinite. Most of the cases of incompletely treated patients relapsed earlier. In the 90 culture proven cases in which antituberculosis drug susceptibility was tested, 39 patients showed resistance to at least one drug, 9 patients were resistant to only one drug, 9 patients were resistant to two drugs and 21 patients were resistant to more than 3 drugs. The common reasons for failure of treatments were: 1) poor patient compliance to medication: 50 cases, 2) multiple drug resistance: 30 cases, 3) delayed treatment: 19 cases. Some cases included a combination of the above. PMID- 15272757 TI - Characteristics of HIV-infected tuberculosis patients in Kota Bharu Hospital, Kelantan from 1998 to 2001. AB - To characterize the demographic profiles, clinical features, radiological patterns and outcomes of treatment of HIV-infected TB patients, a descriptive study was carried out on 149 HIV-infected TB cases diagnosed from 1998 through 2001 at Kota Bharu Hospital, Kelantan, Malaysia. The majority of the patients were males (94.6%), single (45.0%), ethnic Malay (94.0%) with a mean age of 34 years (standard deviation 7.8, range 18-76). The most common HIV transmission category was through injecting drug use (73.8%) and being the inmates or former inhabitants of drug rehabilitation centers and prisons were the commonest high risk groups. One hundred and seventeen patients were diagnosed as having pulmonary TB, while about 20% were extra-pulmonary in type with 9 cases of milliary TB. The majority (45%) presented with cough symptoms while only 51% had a positive sputum smear. Fifty-five percent were found to have pulmonary lesions on chest x-ray, such as localized, milliary or diffuse pulmonary infiltrates, or opacities. Eight (5.4%) had pleural lesions while another 8 cases had hilar or mediastinal lymph node lesions. Overall, fifty-eight (38.9%) patients had died by the completion of data collection. The median weeks or survival from the time of starting TB treatment was 13.5 (range 1-56) and the majority of them (74%) died without completing the 6-month regime of treatment. PMID- 15272758 TI - Evaluation of CD4 counts and percentages in the HIV infected Indian population. AB - This study was designed to evaluate the absolute CD4+ counts and percentages in HIV subtype C infected patients at a tertiary care hospital in northern India. The CD4+ counts of 377 HIV seropositive subjects were estimated by a FACS Calibur (BD) flow cytometer. Dual color immunophenotyping was performed on each sample, which was acquired and analysed using CellQUEST software. Discordance between CD4+ counts and percentages were found more in the early stage ie Group A (37.2%) when compared with Group B (31.6%) and Group C (28.8%), with the counts remaining in the normal range but percentages being severely depressed. PMID- 15272759 TI - Hepatitis C virus infection among Thai blood donors: antibody prevalence, risk factors and development of risk screening form. AB - Hepatitis C virus (HCV) infection is an important blood-borne infection in many countries, including Thailand. For epidemiological surveillance and controlling the infection, 2167 blood donors were screened for antibody to HCV by an enzyme immunoassay method and interviewed by using a structured questionnaire which consisted of personal health history and some risk behaviors. The prevalence and risk factors were assessed and the risk screening form was developed. The results revealed that the prevalence of anti-HCV was 2.90%. Male blood donors had relatively higher anti-HCV positive rate than females (3.21% vs 1.77%). The significant risk factors from univariate analysis were: (a) gender as male, OR = 1.94 (p = 0.042), (b) education to the primary level, OR = 4.15 (p < 0.001), (c) occupation as laborer or agriculture workers, OR = 2.87 (p < 0.001), police and military, OR = 1.82 (p = 0.046), (d) residence in a rural area, OR = 3.09 (p < 0.001), (e) a history of receiving blood or blood products, OR = 5.21 (p < 0.001), (f) a history of tattooing, OR = 1.70 (p = 0.043), (g) a history of IDU (Infecting Drug Use), OR = 41.43 (p < 0.001), (h) a history of STDs (sexually transmitted diseases) in the last year, OR = 3.87 (p = 0.021), and (i) a history of sexual service, OR = 4.24 (p = 0.017). After multivariate analysis, four variables related to HCV infection among the studied samples included education to the primary level, OR = 3.34 (p = 0.0036), occupation as a laborer or agriculture worker, OR = 2.14 (p = 0.0092), a history of receiving blood or blood products, OR = 4.13 (p = 0.0029), and a history of IDU, OR = 3.82 (p < 0.0001). The risk screening form was developed using risk scores. The validity was calculated by the Receiving Operating Curve. The sensitivity of this form was approximately 55.3% and the specificity was 85.7% when a cut-off score at risk > or =7 was used. If the cut-off score was > or =6, the screening form showed 77.1% of specificity and 61.3% sensitivity. This risk screening form should be applied not only for blood donation but also for pre-marital health screening. PMID- 15272760 TI - Infection risk to travelers going to dengue fever endemic regions. AB - The risk of dengue virus infection to travelers visiting dengue fever endemic regions was studied through the use of mathematical modeling. A Susceptible Infected-Recovered (SIR) model is used to describe the transmission of dengue fever (DF) in an endemic region into which tourists enter. The dynamics of a new class of human, the traveler, is incorporated into the systems of first order differential equations in the SIR describing the dynamics of the transmission in the host region. Using standard dynamic analysis methods, the numbers of travelers who become infected with the dengue virus are calculated as a function of the length of time the tourist stays in the region. PMID- 15272761 TI - Case report: Dengue hemorrhagic fever with encephalopathy in an adult. AB - Encephalopathy in dengue hemorrhagic fever is a very rare condition and usually occurrs in the febrile stage. We report a 29-year-old woman, who presented with acute fever, thrombocytopenia and positive IgM antibodies for dengue virus. On the fourth hospital day, the fever subsided and she developed a confusional stage. CT scan and MRI of the brain were within normal limits. Electroencephalography (EEG) revealed generalized theta waves. Cerebrospinal fluid was normal. She was treated with supportive treatment. Five days later, she was fully recovered without any neurological deficits. This is a first case of encephalopathy in dengue hemorrhagic fever that developed after the fever subsided. PMID- 15272762 TI - Effect of bird-to-bird transmission of the West Nile virus on the dynamics of the transmission of this disease. AB - Two recent publications report that direct bird-to-bird transmission of West Nile virus is possible. The effect of a bird-to-bird transmission on the transmission dynamics of this virus is studied through mathematical modeling. The model still treats the bird-to-mosquito-to-bird as the main transmission route. The results of numerical calculations show that there are changes in the dynamics of the transmission of West Nile fever in humans when the non-mosquito transmission route becomes more important. PMID- 15272763 TI - Case report: A Thai patient with Leber's hereditary optic neuropathy linked to mitochondrial DNA 14484 mutation. AB - A young Thai male presented with bilateral visual loss and disc pallor. The 14484 mutation responsible for Leber's hereditary optic neuropathy (LHON) was identified on blood mitochondrial analysis. His visual loss was more severe than the visual loss described in Caucasian and Japanese patients and showed no improvement. He had no other identifiable mutations related to LHON nor any associated neurological disorder. This is the first case report of LHON with the 14484 mutation in a Thai patient. PMID- 15272764 TI - Small vocal cord polyps: completely resolved with conservative treatment. AB - Vocal fold polyps usually occur on the anterior or middle part of the membranous vocal fold and are the commonest laryngeal pathology requiring surgical removal. We report on six cases of small vocal polyps (4 cases angiomatous polyps and 2 gelatinous) that completely resolved using conservative treatment. Not every case of polyps requires surgical removal. PMID- 15272765 TI - Treatment of eosinophilic meningitis with a combination of albendazole and corticosteroid. AB - To study the efficacy of the combination of albendazole and prednisolone for the treatment of eosinophilic meningitis, we conducted a pilot study among Thai patients with eosinophilic meningitis. Patients were given a 2-week course of prednisolone, 60 mg/day and albendazole, 15 mg/kg/ day. The primary observation parameter was the number of patients who still had headache after the 2-week course of treatment. Twenty-six patients were enrolled in the study. There were 3 (11.5%) patients who still had headache after the 2-week course of treatment and the median length of time until complete disappearance of headache was 4 days. Serious side effects were not detected. Treatment for 2 weeks with the combination regimen of albendazole and prednisolone is safe and effective for the treatment of eosinophilic meningitis. PMID- 15272766 TI - Case report: Diazepam in severe tetanus treatment. AB - The causes of death in tetanus are muscle spasms and spasm of the larynx, which are caused by blocking the release of inhibitory neurotransmitters in the spinal synapses, causing the uncontrolled spread of impulses. Diazepam controls the spasms by blocking the polysynaptic reflexes, working peripherally, without depressing the cortical center and has no cardiovascular or endocrine effects. High dose diazepam had been used and proved to be a good muscle relaxant. Diazepam seems to work better with tetanus than pancuronium bromide, but both drugs need mechanical ventilation. In cases where the dose exceeds 240 mg per day in a child, a ventilator should be on hand, and if the dose required is more than 480 mg per day, other drugs should be considered. In three cases of severe tetanus presented here, the first two were managed by diazepam and pancuronium bromide and the last case by high dose diazepam only. In the first case, the dose of diazepam was up to 480 mg/day. By using high dose diazepam in severe tetanus, management of the clinical manifestations of autonomic nerve involvement and the weaning process become easier. Most complications of severe tetanus became more manageable. PMID- 15272767 TI - Antimicrobial use in children under five years with diarrhea in a central region province, Thailand. AB - This cross-sectional study aimed to estimate the prevalence of appropriate antimicrobial prescribing for treating childhood diarrhea within the public hospital system in a central region province, Thailand. Reported are findings of a prospective clinical audit of 424 cases treated by 38 physicians. Appropriate use of antimicrobials was defined as prescribing antimicrobials for managing an invasive bacterial-type, bloody diarrhea or not prescribing antimicrobials for managing a watery-type or non-bloody diarrhea. Among 424 cases with diarrhea, 12.5% were invasive bacterial-type. Of the 66 diarrheal episodes in which stool samples were cultured, 7 stool specimens were positive, two with Shigella sonnei, two with Vibrio cholerae Ogawa and three with E. coli. Based on the presence of mucus and blood in stools, 27.4% of 424 cases received appropriate antimicrobial drugs. Cotrimoxazole was the most commonly prescribed drug (51%), followed by colistin sulfate (15.3%), norfloxacin (11%), and nalidixic acid (0.5%). The average number of antimicrobials per case of inpatients was higher than outpatients (1.15 vs 0.84, p < 0.001). There was a trend toward prescribing norfloxacin in childhood diarrhea. The Ministry of Public Health should continue providing effective interventions aimed at improving physicians' knowledge of diarrhea treatment. Similar efforts should be directed toward improving caretakers' knowledge about home care for childhood diarrhea and encouraging widespread correct use of oral rehydration therapy (ORT) in the community. Hopefully, such activities will help reduce the inappropriate use of antimicrobial agents in treating diarrheal disease. PMID- 15272768 TI - Primary verification: is the TRISS appropriate for Thailand? AB - The Trauma and Injury Severity Score (TRISS) is a well-accepted model used to evaluate the quality of trauma care in the US. This research aims to study whether TRISS can be applied to evaluate trauma care and classify outcomes of road traffic injury patients in Thailand. A retrospective study was used to review the Thailand's Injury Surveillance System database from the 1st January to the 31st of December 1996. The study subjects were severe road traffic injury patients with blunt injuries. The TRISS model was applied to compute the survival probability for each patient. The chi-square goodness-of-fit was used to compare the survival probability distribution between the American Major Trauma Outcome of Study (MTOS) and the road traffic injuries in Thailand. The accuracy, sensitivity and specificity of the survival prediction by TRISS were evaluated. The distribution of survival probability between American trauma patients and Thai road traffic injury patients was significantly different (p-value < 0.00001). The TRISS model had high accuracy and sensitivity, but low specificity, in predicting the survival of Thai road traffic injuries. The MTOS and Thai road traffic injuries had different distributions for various factors such as the Revised Trauma Score (RTS), Injury Severity Score (ISS), and ages which effect injury survival. Due to these factors the distribution of survival probability between MTOS and Thai road traffic injuries was also significantly different. By applying TRISS, the survival prediction of Thai road traffic injuries resulted in a high number of false positives. PMID- 15272769 TI - Research report: Frequencies of mica gene polymorphism: a comparison between Indonesians on Bacan Island and suburban Japanese. AB - MHC class I chain related gene A (MICA) is located near the HLA-B gene on the short arm of human chromosome 6. In the transmembrane (TM) of region of MICA, there is a trinucleotide repeat (GCT/AGC) microsatellite polymorphism in exon 5. Five alleles with 4, 5, 6 and 9 repetitions or 5 repetitions with 1 additional nucleotide insertion (GGCT) are identified and they were named A4, A5, A5.1, A6, and A9 respectively. We report the allele frequencies of 127 Indonesians on Bacan Island and 250 Japanese in the Kanto area. From the genotyping result, the frequency among Indonesians was as follows: A4 15.4%, A5 26.0%, A5.1 16.5%, A6 5.5%, and A9 36.6%. The frequency among Japanese was as follows: A4 20.6%, A5 28.1%, A5.1 10.8%, A6 27.2%, and A9 13.2%. Allele 9 is significantly increased and allele 6 significantly decreased in Indonesians compared with Japanese subjects. The results suggested that MICA microsatellite polymorphism are quite different in each race. Among Indonesians, the frequency of MICA-A9 allele, which was reported to be negatively associated with Behcet's disease, was significantly higher, whereas the MICA-A6 allele frequency, which was reported to be positively associated with Behcet's disease, was significantly lower among Japanese. PMID- 15272770 TI - Incidence of enteric bacteria and Staphylococcus aureus in day care centers in Akwa Ibom State, Nigeria. AB - The incidence of enteric bacteria and Staphylococcus aureus in four day care centers in Akwa Ibom State was studied using culture techniques. The percentage frequencies of the isolates from 124 samples were Staphylococcus aureus (33.9), Escherichia coli (19.0), Klebsiella sp (14.4), Citrobacter sp (12.5) and Proteus mirabilis (7.4). The sources of contamination were floors, chairs, skin, bed linen, door handles, fans, children's tables, walls, windows, ceiling, headmistress's table and chairs, drinking water and wash water. Cultures from Aunty Chimmy's Day Care and Nursery School, Eket and Ideal Day Care and Nursery School, Eket yielded more organisms than those from Trinity International Nursery School, Ikot Ekpene and Adiaha Obong Day Care Center in Uyo. The results revealed the insanitary conditions in these day care centers. The enforcement of an effective public health enlightenment program is advocated in order to attract sufficient attention of the proprietors of these establishments to the role of fomites as reservoirs of pathogenic microorganisms. PMID- 15272771 TI - A study of job strain and dissatisfaction among lecturers in the School of Medical Sciences Universiti Sains Malaysia. AB - Job stress has now become one of the most significant health and safety issues in the workplace and one of the least understood areas of organizational cost. A cross-sectional study to assess job strain and dissatisfaction in lecturers of the School of Medical Sciences, Universiti Sains Malaysia (USM) was undertaken between August 2001 and May 2002. The original English version of the Job Content Questionnaire (JCQ) version 1.7 (revised 1997) by Robert Karasek was self administered to 73 (response rate 58.4%) lecturers in School of Medical Sciences USM. The prevalence of job strain (defined by low decision latitude and high psychological demands) in USM was 23.3%. The risk factors of job strain in the lecturers were psychological stressors (adjusted OR 1.2, 95% CI 1.0, 1.4), created skill (adjusted OR 0.4, 95% CI 0.2, 0.8) and working in clinical-based departments (adjusted OR 18.7, 95% CI 1.6, 22.7). The prevalence of job dissatisfaction was 42.6%. Associated factors of job dissatisfaction in USM lecturers were decision authority (p < 0.001) and psychological job demand (p < 0.001). We conclude that psychological stressors and created skill were non protective and protective, respectively, against job strain in USM lecturers. Clinical-based lecturers experienced higher job strain compared to non-clinical based lecturers. Psychological job demand was strongly associated with job dissatisfaction, and decision authority was protective against job dissatisfaction. PMID- 15272772 TI - Cigarette smoking and its relation to pulmonary tuberculosis in adults. AB - The purpose of this hospital-based case-control study is to determine the effect of passive and active smoking on pulmonary TB in adults. The study subjects were 100 new pulmonary TB cases diagnosed at TB Division, and age-sex matched 100 non TB cases from patients admitted to Taksin Hospital and healthy subjects who came for annual physical check-up at either the outpatient clinic of the TB division or Taksin Hospital, during May 2001 to October 2001. All subjects had blood tests and only persons who were HIV-negative, DM-negative and free of other lung diseases were included. Data were collected by direct interview using questionnaires. Multivariate analysis of cigarette smoking related to pulmonary TB in adults was performed. The factors related to pulmonary TB in adults were current active smoking regardless of passive smoking exposure. There was a significant association between early age at initiation of smoking and TB. Active (current + ex-active) smokers who started smoking at age 15-20 years had a higher risk of pulmonary TB compared to others (OR = 3.18, 95% CI = 1.15-8.77); as well as the long duration of smoking: persons who had smoked >10 years had a higher risk of pulmonary TB (OR = 2.96, 95% CI = 1.06-8.22). There was a relationship between pulmonary TB and the amount of smoking exposure. Those who smoked >10 cigarettes/day (OR = 3.98, 95% CI = 1.26-12.60) or >3 days/week (OR = 2.68, 95% CI = 1.01-7.09) had higher risk of pulmonary TB compared to non-smokers. Passive smokers who were exposed to tobacco smoke >3 times/week outside the home had a higher risk of pulmonary TB than those with exposure < or =3 times/week (OR = 3.13, 95% CI = 1.07-9.17). It was also found that the effects of passive smoking in the office and/or neighborhood were strong. Persons with such exposures had a higher risk of pulmonary TB than no exposure or exposure < or =3 times/week from either or both places (OR = 4.62, 95% CI = 1.68-14.98). Therefore, an effective anti-smoking campaign is expected to have a positive repercussion on TB incidence. Smoking cessation must be considered and promoted by all levels of health care providers. PMID- 15272773 TI - Methamphetamine abuse during pregnancy and its health impact on neonates born at Siriraj Hospital, Bangkok, Thailand. AB - To ascertain the impact of intrauterine methamphetamine exposure on the overall health of newborn infants at Siriraj Hospital, Bangkok, Thailand, birth records of somatic growth parameters and neonatal withdrawal symptoms of 47 infants born to methamphetamine-abusing women during January 2001 to December 2001 were compared to 49 newborns whose mothers did not use methamphetamines during pregnancy. The data on somatic growth was analyzed using linear regression and multiple linear regression. The association between methamphetamine use and withdrawal symptoms was analyzed using the chi-square. Home visitation and maternal interview records were reviewed in order to assess for child-rearing attitude, and psychosocial parameters. Infants of methamphetamine-abusing mothers were found to have a significantly smaller gestational age-adjusted head circumference (regression coefficient = -1.458, p < 0.001) and birth weight (regression coefficient = -217.9, p < or = 0.001) measurements. Methamphetamine exposure was also associated with symptoms of agitation (5/47), vomiting (11/47) and tachypnea (12/47) when compared to the non-exposed group (p < 0r =0.001). Maternal interviews were conducted in 23 cases and showed that: 96% of the cases had inadequate prenatal care (<5 visits), 48% had at least one parent involved in prostitution, 39% of the mothers were unwilling to take their children home, and government or non-government support were provided in only 30% of the cases. In utero methamphetamine exposure has been shown to adversely effect somatic growth of newborns and cause a variety of withdrawal-like symptoms. These infants are also psychosocially disadvantaged and are at greater risk for abuse and neglect. PMID- 15272774 TI - Motorcycle helmet use and related risk behaviors among adolescents and young adults in Northern Thailand. AB - The objective of this study was to assess the prevalence of not wearing a helmet (unprotected) while riding a motorcycle and associated risk behaviors among adolescents and young adults in Northern Thailand. Participants were 1725 students, aged 15-21 years, from 3 vocational schools in Chiang Rai Province; 51.8% were male. Participants completed a classroom-based computer-assisted self interview (ACASI). Of men 72.7% and of women 64.4% reported unprotected motorcycle riding 3 times or more in the past week. Logistic regression analysis showed the variables independently associated with unprotected riding to be history of ever riding after having had 3 or more alcoholic drinks (odds ratio (OR) = 2.21, 95% confidence interval (CI) = 1.76-2.21), attending technical school (OR = 2.09, 95% Cl = 1.55-2.83), living with the family (OR = 1.38, 95% CI = 1.10-1.73), and having ever had a traffic accident (OR = 1.20, 95% CI = 1.12 1.29). Being of hill tribe ethnicity (vs Thai lowlander) was associated with protected riding (OR = 0.42, 95% CI = 0.20-0.90). Adolescents and young adults in Chiang Rai are at high risk for riding a motorcycle without a helmet buckled on the head. Public education in combination with enforcement of compulsory helmet use while riding a motorcycle is recommended. PMID- 15272775 TI - [The relationship between growth habit of Lonicera japonica and quality of Flos Lonicerae]. AB - The relation of growing and flowering rhythm of Lonicera japonica to the quality of Flos Lonicerae was studied. The result showed that the yield and quality of Flos Lonicerae were different in different plant age and collecting time,the best collecting time was in the completely white flower bud stage, the first and the second batch flower buds had the highest yield and the best quality. PMID- 15272776 TI - [Effect of boron, manganese and rare soil trace fertilizer on Angelica sinensis]. AB - The experiment of spraying boron, manganese and rare soil trace fertilizer was carried out on Angelica sinensis in leave flourish stage and root expand stage respectively. The result showed that trace fertilizer supplement, especially manganese sulfate and rare soil,could significantly enhance the yield over 60% and improve the proportion of high quality product. PMID- 15272777 TI - [Studies on random amplified polymorphic DNA fingerprinting of Cortex Magnoliae Officinalis]. AB - OBJECTIVE: To identify Chinese traditional medicine (CTM) "Hou-pu" (Cortex Magnoliae Officinalis), its counterfeits and substitues. METHODS: Total genomic DNA samples of ten plant species were amplified by randomly amplified polymorphic DNA (RAPD). RESULTS: Ten samples were able to be distinguished through their amplified DNA banding patterns on the agarose gels after electrophoresis. CONCLUSION: RAPD is able to identify "Hou-pu", its counterfeits and substitutes quickly and truly, which is also quite valuable for correctly introducing plant. PMID- 15272778 TI - [Chemical constituents from Typhonium flagelliforme]. AB - OBJECTIVE: To study the major chemical constituents from the root tuber of Typhyonium flagelliforme. METHOD: Compounds were separated and purified with silica gel column and preparative HPLC, and their structures were elucidated on the basis of spectroscopic evidence (UV, IR, MS, NMR and 2D NMR). RESULT: Four compounds were identified as 1-O-beta-glucopyranosyl-2-[(2-hydroxyloctadecanoyl) amido]-4,8-octadecadiene-1,3-diol (1), coniferin (2), beta-sitosterol (3) and beta-daucosterol (4). CONCLUSION: A cerebroside with significant antihepatotoxic activity and a phenylpropanoid glycoside were isolated from Typhyonium flagelliforme for the first time. PMID- 15272779 TI - [Determination of phenylethanoid glycosides from Cistanche deserticola in spring and autumn with LC-MS]. AB - OBJECTIVE: To have a contrast study on phenylethanoid glycosides from Cistanche deserticola Y. C. Ma collected in different seasons. METHODS: LC/MS method has been applied for the analysis of four kinds of phenylethanoid glycosides compunds (echinacoside, acteoside, cistanoside A and 2'-acetylacteoside) from Cistanche deserticola Y. C. Ma in spring and autumn. RESULTS: According to the special MS spectra and HPLC chromatogram, this four kinds of phenylethanoid glycosides compounds were detected in each Cistanche deserticola Y. C. Ma, but the content is considerable different except the acteoside. CONCLUSION: The content of phenylethanoid glycosides from Cistanche deserticola Y. C. Ma in different seasons has a difference from each other, the quality of Cistanche deserticola Y. C. Ma is also different. PMID- 15272780 TI - [Supercritical CO2 extraction of fatty oils from bee pollen and its GC-MS analysis]. AB - The extraction of fatty oils from bee pollen with supercritical CO2 was studied. The effects of extraction pressure, extraction temperature and the grinding size of the bee pollen on the yields were discussed. The optimal condition of this method was: extraction pressure 30MPa,extraction temperature 55 degrees C, separator I pressure 14MPa, separator I temperature 45 degrees C, separator II pressure 6MPa, separator II temperature 40 degrees C, extraction period 2 hours. Compared with the traditional solvent extraction, this method had some virtues, such as shortening the extraction period greatly, escaping oxidation of unsaturated fatty acid, and the products from which having better quality. With a GC-MS analysis, the main composition of the fatty oil from the bee pollen was: oleic acid, palmitic acid, linoic acid, pentacosane, octacosane and so on. The contents of alpha-linolenic acid in the fatty oils from separator I and separator II were 42% and 8.8%, which differed greatly. It proved that this method succeeded in extraction, separating and enriching alpha-linolenic acid. PMID- 15272781 TI - [Effects of flavone from leaves of Diospyros kaki on adventitial fibroblasts proliferation by advanced oxidation protein products in vitro]. AB - OBJECTIVE: To observe the effects of flavone from leaves of Diospyros kaki on adventitial fibroblasts proliferation by advanced oxidation protein products (AOPP) in vitro. METHODS: NIH-3T3 cells were cultured in vitro and treated with AOPP and flavone from leaves of Diospyros kaki, respectively, and observed in comparison with the control group. The ratio of cell proliferation was determined by non-radioactive MTS/PES assay. RESULTS: The ratio of cell proliferation was 1.789 +/- 0.299 in the control group, and 2.064 +/- 0.141, 2.149 +/- 0.218, 2.108 +/- 0.165, 2.124 +/- 0.131 and 2.087 +/- 0.125 in AOPP groups corresponding to AOPP concentrations of 100, 50, 10, 1 and 0.1 microg/ml, respectively. It showed that AOPP significantly induced the fibroblasts proliferation when the concentration was above 100 ng/ml (P < 0.05). The ratio of cell proliferation was 1.714 +/- 0.179 in flavone from leaves of Diospyros kaki group corresponding to concentration of 50 microg/ml. It also showed that flavone from leaves of Diospyros kaki alone had no effect on fibroblasts proliferation (P > 0.05). With AOPP stimulation, flavone from leaves of Diospyros kaki significantly inhibited fibroblasts proliferation (P < 0.05). CONCLUSIONS: Flavone from leaves of Diospyros kaki can significantly inhibit the adventitial fibroblasts proliferation stimulated by AOPP in vitro. PMID- 15272782 TI - [Antioxidative activities of two metabolites of cultured marine fungus, Halorosellinia oceanicum 323 in vitro]. AB - OBJECTIVE: To investigate the antioxidative effects of 323-A and 323-B, two isomers extracted from the metabolites of cultured marine fungus, Halorosellinia oceanicum 323 in vitro. METHODS: NADH-PMS-NBT system was used to produce superoxide free radical (O2*-), EDTANa2-Fe(II)-H2O2 system to generate hydroxyl free radical (*OH), H2O2 to stimulate oxidative hemolysis of erythrocytes of rats, Cys-Fe2+ to induce malondialdehyde (MDA) production in homogenates, and ferrous-ascorbic acid system to increase the turbidity of mitochondria suspension in the liver of rats. And the antioxidative activities of 323-A and 323-B were studied. RESULTS: 323-A and 323-B not only scavenge O2*- and *OH produced by the experimental systems directly, but also inhibit H2O2 stimulated oxidative hemolysis of erythrocytes of rats, depress MDA production in homogenates induced by Cys-Fe2+ system, and reduce the turbidity of mitochondria suspension in the liver of rats increased by ferrous-ascorbic acid system in vitro. CONCLUSION: 323 A and 323-B showed comprehensive cleaning actions on free radicals and protective effects on the functions of tissues and cells against oxidative lesion. The results suggested that the marine microorganic metabolites might be a novel and profound source of antioxidative reagents. PMID- 15272783 TI - [Immunosuppressive activity of exopolysaccharide from Paecilomyces gunnii]. AB - OBJECTIVE: To investigate bioactivities of Paecilomyces gunnii exopolysaccharide (PGEP). METHODS: The effects of PGEP on mice spleen lymphocytes proliferation, mice peritoneal macrophage (PMphi) phagocytosis and cytotoxin T lymphocytes (CTL) activity were studied by MTT method, neutral red colorimetry, respectively. RESULTS: In vitro, mice spleen lymphocytes proliferation, mice peritoneal macrophage (PMphi) phagocytosis of neutral red and CTL activity were significant suppressed by PGEP, especially in 10 microg/ml. CONCLUSION: PGEP had the immunosuppressive activities. PMID- 15272784 TI - [Influence of Shenshuai Capsule on renal tissue apoptosis of rats with chronic renal failure]. AB - OBJECTIVE: To explore the influence of Shenshuai Capsule on renal tissue apoptosis of Wistar rats with chronic renal failure. METHOD: Wistar rats were fed with adenine to establish chronic renal failure model, and then they were divided into three groups separately fed with Shenshuai capsule (high or low dosage) and Niaoduqing. RIA, immunohistochemical method and electron microscopy were used. RESULTS: The parameters of renal function between the treatment group (high dosage) and the Niaoduqing group were significantly different respectively. Shenshuai capsule could inhibit tubular epicyte apoptosis and Fas genetic expression, and improve Bcl-2 genetic expression. CONCLUSION: Shenshuai capsule could inhibit epicyte apoptosis, and adjust Fas genetic expression. Bcl-2 genetic expression indicated that this may be one of the mechanism to postpone the renal failure by Shunshuai capsule. PMID- 15272785 TI - [Effect of Composite Yinchen Tablet on lymphocyte proliferation in mice]. AB - OBJECTIVE: To observe the effect of Composite Yinchen Tablet (CYT) on lymphocytes proliferation in mice. METHODS: By the method of 3H-TdR admixing, the effect of CYT on lymphocytes proliferation in mice were detected in vivo and in vitro. RESULTS: CYT in high, medium and low dose could significantly increase the cpm level of 3H-TdR. CYT could improve the lymphocytes function of mice. CONCLUSION: CYT could enhance cellular immune function. PMID- 15272786 TI - [Pathological observations of Fructus Schisandrae polysaccharide on anti-tumor effects in S180-bearing mice]. AB - OBJECTIVE: To study the effects of Fructus Schisandrae Polysaccharide (FSP) on anti-tumor in S180-bearing mice. METHODS: The transplantable carcinosarcoma model was established in S180-bearing mice by subcutaneous implantation. Tumor size and histopathological changes were observed under electron microscopy by HE staining after 8 days treatment. RESULTS: Morphological changes showed that slight cataplasia and apoptosis were induced by high concentration FSP, as well as the numbers of inflammatory cells in tumor were increased and tumor cells in infiltration areas around tumor cataplasised partly. These suggested that high concentration FSP can inhibit tumor growth lightly. In addition, high concentration FSP combined with cyclophosphamide can promote the effect of anti tumor. CONCLUSIONS: FSP can inhibit tumor growth, this effect may be correlated with apoptosis and the activation of immunocytes, but not depend on killing tumor cells directly. In addition, high concentration FSP combined with cyclophosphamide can promote the capacity on anti-tumor in S180-bearing mice. PMID- 15272787 TI - [Studies on antitussive, antiasthmatic and expectorant action of chenodeoxycholine acid]. AB - OBJECTIVE: In this paper, antitussive, antiasthmatic and expectorant action of chenodeoxycholine acid (CDCA) were studied. METHOD: The method of strong aqua ammoniae induced cough and sulfur dioxide induced cough, the method of tracheal volume and tracheal spiral line, and phenolsulfonphthalein excretion test. RESULTS: CDCA was able to prolong cough latent period and decrease cough rate significantly in mice resulted from strong aqua ammoniae and sulfur doxide, and inhibit contraction of whole-body or extracorpored tracheal smooth muscle in guinea pig resulted from histamine phosphate. Besides, CDCA can increase phenolsulfonphthalein excretion quantity of trachea in mice, singnificantly. CONCLUSION: CDCA has antitussive, antiasthmatic, expectorant actions. PMID- 15272788 TI - [Study on the extraction process for free isofraxidin and hydrolytic condition of combined isofraxidin in Ciwujia with orthogonal design]. AB - OBJECTIVE: To optimize the extraction and hydrolytic condition of isofraxidin in Ciwujia. METHODS: The study was carried out through orthogoral experimental design. The content of isofraxidin was determined as marker to evaluate the effect on the extraction and hydrolytic condition of different process. RESULTS: The optimum extraction condition was 12 times methanol and 3 times for sixty minutes every time. And the optimum hydrolytic condition was that hydrolysis for two hours, ten times amount of 15% sulfuric acid at 100 degrees C. PMID- 15272789 TI - A PVDF transducer for low-frequency acceleration measurements. AB - A unique acceleration transducer, using piezoelectric PVDF, has been developed for low-frequency vibration monitoring. The paper develops the theoretical model for this low-cost, robust sensor. The theoretical model is validated using experimental results from laboratory tests. The sensor was also installed in an underground potash mine alongside a commercial geophone for a three-month in-mine test producing results that show a close correspondence between the two transducers. PMID- 15272790 TI - Robust and efficient vision system for group of cooperating mobile robots with application to soccer robots. AB - In this paper a global vision scheme for estimation of positions and orientations of mobile robots is presented. It is applied to robot soccer application which is a fast dynamic game and therefore needs an efficient and robust vision system implemented. General applicability of the vision system can be found in other robot applications such as mobile transport robots in production, warehouses, attendant robots, fast vision tracking of targets of interest and entertainment robotics. Basic operation of the vision system is divided into two steps. In the first, the incoming image is scanned and pixels are classified into a finite number of classes. At the same time, a segmentation algorithm is used to find corresponding regions belonging to one of the classes. In the second step, all the regions are examined. Selection of the ones that are a part of the observed object is made by means of simple logic procedures. The novelty is focused on optimization of the processing time needed to finish the estimation of possible object positions. Better results of the vision system are achieved by implementing camera calibration and shading correction algorithm. The former corrects camera lens distortion, while the latter increases robustness to irregular illumination conditions. PMID- 15272791 TI - Embedded estimator in predictive feedback control. AB - In this work, a new approach to design predictive feedback control for SISO systems is presented. The proposed formulation relies on the development of a single step predictor based on an autoregressive moving average with external input (ARMAX) model. Although no explicit observer is actually involved in the implementation, this predictor implicitly includes one since the input-output model subsumes an observer. Exploiting this idea the resulting ARMAX model is extended to include extra outputs to improve the quality of the prediction for systems with large time delay and nonmeasurable disturbances. The resulting predictor is used to develop a predictive feedback controller. This new formulation of predictive feedback control includes feedback and feedforward actions. Simulations of two linear systems illustrate the applicability of the control algorithm. PMID- 15272792 TI - Flexible-structure control: a strategy for releasing input constraints. AB - In this paper output unreachability under input saturation phenomenon is studied: under a large disturbance or setpoint change, the process output may never reach the set point even when the manipulated variable has driven to saturation. The process output can be brought back to the set point only by activating an auxiliary manipulated variable. A new control structure for designing and implementing a control system capable of solving this problem is proposed by transferring the control from one variable to another and taking into account the different dynamics involved in the system. The control structure, called flexible structure control due to its ability to adapt the control structure to the operating conditons, is a generalization of the split-range control. It can be summarized as two controllers connected through a piecewise linear function. This function decides, based on the value of one manipulated variable, when and how the control structure changes. Its parameters control the interaction between both manipulated variables and leave the capability for handling the balance between control quality and other goals to the operator. PMID- 15272793 TI - Robust covariance control for discrete system by Takagi-Sugeno fuzzy controllers. AB - Variances of the system states or outputs often play vital roles in the problem for performance requirements of many stochastic control systems. For linear stochastic systems, the covariance control technique has been applied to deal with the variance constrained design problem. This paper extends this technique to a class of discrete-time nonlinear perturbed stochastic systems, which are modeled by the Takagi-Sugeno (TS) fuzzy systems. By fuzzy IF-THEN rules, which represent local linear input-output relations, the nonlinear systems can be described by TS fuzzy models. According to the parallel distributed compensation (PDC) concept, the discrete-time nonlinear perturbed stochastic systems can be driven by the linear feedback gains. The purpose of this paper is to provide a method to design an output feedback fuzzy controller, which is based on the upper bound state covariance control technique and PDC concept, for the discrete-time perturbed stochastic systems using TS fuzzy models. PMID- 15272794 TI - Covariance control with observed-state feedback gains for continuous nonlinear systems using T-S fuzzy models. AB - The design problem of state variance constrained control for stochastic systems has received rather extensive attention in recent years. This paper solves the state variance constrained controller design problem by using the covariance control theory, with observed-state feedback gains for continuous Takagi-Sugeno (TS) fuzzy models. By incorporating the technique of state estimation into the practical covariance control theory, a variance constrained control methodology is developed for the continuous TS fuzzy models. Finally, a numerical example is shown to demonstrate the efficiency and applicability of the proposed approach. PMID- 15272795 TI - Precision motion control with a high gain disturbance compensator for linear motors. AB - In this paper, we address the problem relating to the precision control of permanent magnet linear motors to track repeated motion trajectories. A high gain disturbance compensator is developed to improve the control performance degraded due to the presence of significant disturbances. An inverse gain of the overall system model is used to set up a disturbance observer. The observed disturbance is then used to generate a "knocker" signal, to be augmented to the control signal, which can provide the additional energy necessary to overcome the effects of the disturbances. A learning scheme is used to adjust the knocker signal iteratively over the repeated cycles. Simulation and experimental results are furnished to demonstrate the effectiveness of the proposed control scheme. PMID- 15272796 TI - A new self-tuned PID-type fuzzy controller as a combination of two-term controllers. AB - The present paper is a venture into the domain of proportional-integral derivative (PID) -type adaptive fuzzy logic controllers (FLC's) and proposes a new algorithm which is realized by a self-tuned PI-type FLC (in velocity form) in parallel with a self-tuned PD-type FLC (in position form). Each of these PI/PD controllers implements a supervisory static FLC for adaptive online modification of the output scaling factor (SF) of a static PI/PD FLC. The proposed scheme is developed with a view to having a PID-type FLC with an architecture, simple enough for practical implementation, which at the same time has substantially satisfactory performance for a wide class of processes. Simulation studies on a range of processes reveal that the proposed controller has better performance compared to many of its existing counterparts. PMID- 15272797 TI - Electronic cam motion generation with special reference to constrained velocity, acceleration, and jerk. AB - Electronic cam motion involves velocity tracking control of the master motor and trajectory generation of the slave motor. Special concerns such as the limits of the velocity, acceleration, and jerk are beyond the considerations in the conventional electronic cam motion control. This study proposes the curve-fitting of a Lagrange polynomial to the cam profile, based on trajectory optimization by cubic B-spline interpolation. The proposed algorithms may yield a higher tracking precision than the conventional master-slaves control method does, providing an optimization problem is concerned. The optimization problem contains three dynamic constraints including velocity, acceleration, and jerk of the motor system. PMID- 15272798 TI - Manipulated variable based PI tuning and detection of poor settings: an industrial experience. AB - A new method, based on the physical interpretation of the manipulated variable, is developed for tuning of proportional-integral-derivative (PID) controllers and detection of poor settings. This variable is used to set specifications and tune controllers for set-point changes. Here, these specifications are to have a closed-loop time response that is similar to the open-loop time response and a no static error to nonzero mean disturbances acting at the process input. The method is applicable to self-regulating first- and second-order models (overdamped) having stable or unstable zeros as well as delays to integrating first-order processes. For second-order processes, an approximation of equal time constants is used in order to facilitate the identification with step changes and tuning of PI controllers. Results show that the tuning is near optimal even when the time constants are very different. The method has been taught to people with different backgrounds: process engineers, maintenance technicians, production operators, as well as managers. Results have shown better understanding of the possibilities and limitations of process control and improved communication between the different groups. PMID- 15272799 TI - Model-based fault diagnosis in continuous dynamic systems. AB - Traditional fault detection and isolation methods are based on quantitative models which are sometimes difficult and costly to obtain. In this paper, qualitative bond graph (QBG) reasoning is adopted as the modeling scheme to generate a set of qualitative equations. The QBG method provides a unified approach for modeling engineering systems, in particular, mechatronic systems. An input-output qualitative equation derived from QBG formalism performs continuous system monitoring. Fault diagnosis is activated when a discrepancy is observed between measured abnormal behavior and predicted system behavior. Genetic algorithms (GA's) are then used to search for possible faulty components among a system of qualitative equations. In order to demonstrate the performance of the proposed algorithm, we have tested it on a laboratory scale servo-tank liquid process rig. Results of the proposed model-based fault detection and diagnosis algorithm for the process rig are presented and discussed. PMID- 15272800 TI - Development of a novel SCADA system for laboratory testing. AB - This document summarizes the supervisory control and data acquisition (SCADA) system that allows communication with, and controlling the output of, various I/O devices in the renewable energy systems and components test facility RESLab. This SCADA system differs from traditional SCADA systems in that it supports a continuously changing operating environment depending on the test to be performed. The SCADA System is based on the concept of having one Master I/O Server and multiple client computer systems. This paper describes the main features and advantages of this dynamic SCADA system, the connections of various field devices to the master I/O server, the device servers, and numerous software features used in the system. The system is based on the graphical programming language "LabVIEW" and its "Datalogging and Supervisory Control" (DSC) module. The DSC module supports a real-time database called the "tag engine," which performs the I/O operations with all field devices attached to the master I/O server and communications with the other tag engines running on the client computers connected via a local area network. Generic and detailed communication block diagrams illustrating the hierarchical structure of this SCADA system are presented. The flow diagram outlining a complete test performed using this system in one of its standard configurations is described. PMID- 15272801 TI - Stigma, community, ethnography: Joan Ablon's contribution to the anthropology of impairment-disability. AB - Joan Ablon has helped establish the anthropology of impairment-disability and significantly contributed to the role of anthropology in disability studies. In this article, we review the development of and situate Ablon's ethnographic research in the anthropology of impairment-disability. We then address various methodological issues in her work including her ethnographic approach, her grounding in action anthropology and her support for the development of the academic study of disability in anthropology and the careers of disabled anthropologists. The next section of the article examines Ablon's use of the notion of stigma, her understanding of community, and her engagement with disability rights. As examples of themes important to disability studies, we present her discussion of the implications of the ideal of the body beautiful, and gender differences in negotiating intimacy for people with physical differences. We close with a discussion of the future of an anthropology of impairment-disability. PMID- 15272802 TI - Middle Eastern masculinities in the age of new reproductive technologies: male infertility and stigma in Egypt and Lebanon. AB - Worldwide, male infertility contributes to more than half of all cases of childlessness; yet, it is a reproductive health problem that is poorly studied and understood. This article examines the problem of male infertility in two Middle Eastern locales, Cairo, Egypt, and Beirut, Lebanon, where men may be at increased risk of male infertility because of environmental and behavioral factors. It is argued that male infertility may be particularly problematic for Middle Eastern men in their pronatalist societies; there, both virility and fertility are typically tied to manhood. Thus, male infertility is a potentially emasculating condition, surrounded by secrecy and stigma. Furthermore, the new reproductive technology called intracytoplasmic sperm injection (ICSI), designed specifically to overcome male infertility, may paradoxically create additional layers of stigma and secrecy, due to the complex moral and marital dilemmas associated with Islamic restrictions on third-party donation of gametes. PMID- 15272803 TI - Sexuality, color, and stigma among Northeast Brazilian women. AB - Despite its international image as a sexually free-spirited country, local attitudes toward morality of sexual behavior remain complex throughout Brazil, especially in rural areas and the conservative Northeast region. In addition, notwithstanding its official ideology of nonracism, African ancestry as judged through personal appearance (color) constitutes a significant social and economic disadvantage. Using Goffman's idea of "spoiled identity" as a starting point, I show how locals use sexual behavior as a multivocal symbol of moral status in women, and how spoiled sexual reputation interacts with other stigmatized statuses, especially color. I also consider how the acquisition of sexually stigmatized status jeopardizes women's well-being and that of their children. PMID- 15272804 TI - Multiple chemical sensitivities: stigma and social experiences. AB - Multiple Chemical Sensitivity (MCS), an intolerance to everyday chemical and biological substances in amounts that do not bother other people, is a medically contested condition. In addition to symptoms and the ongoing difficulties of living with this condition, this hidden and stigmatized disability strongly impacts social relationships and daily life. Based on an ethnographic study, this article introduces the context of MCS in terms of cultural themes, the media, and the economic power of industries that manufacture the products that make people with MCS sick. Participants' experiences with family members and friends, in work and school settings, and with physicians exemplify the difficulties of living with MCS. I dedicate this article to Joan Ablon, my professor and mentor, whose work has always inspired my thinking and research topics. PMID- 15272805 TI - Childhood asthma on the northern Mexico border. AB - Children with asthma living on the northern Mexico border suffer not only from the physical aspects of this condition, but also from the lack of a clear biomedical definition and treatment plan for the illness. An ethnographic study involving participant observation and focused interviews in Tijuana, Mexico, sought to understand the intersection of diagnostic uncertainties surrounding childhood asthma on the part of parents, particularly mothers, living in acute poverty. Environmental factors such as dust and insects in impoverished homes probably acted as asthma triggers among many of the children in the study. Furthermore, management of children's asthma took place not only in biomedical clinics, but also in homes, traditional medical settings, and pharmacies, where mothers often sought remedies for their children's asthma attacks on an emergency basis. In all treatment settings, including biomedical ones, they often faced significant barriers to effective care, including the misuse of antibiotics. Thus, the role of pharmaceutical sales clerks, as well as pediatric asthma specialists, is explored in this article. PMID- 15272806 TI - Free markets and dead mothers: the social ecology of maternal mortality in post socialist Mongolia. AB - Beginning in 1990, Mongolia, a former client state of what was then the Soviet Union, undertook liberal economic reforms. These came as a great shock to Mongolia and Mongolians, and resulted in food shortages, reports of famine, widespread unemployment, and a collapse of public health and health care. Although economic conditions have stabilized in recent years, unemployment and poverty are still at disturbingly high levels. One important consequence of the transition has been the transformation of the rural, primarily pastoral, economy. With de-collectivization, herding households have been thrown into a highly insecure subsistence mode of production, and, as a consequence, have become vulnerable to local fluctuations in rainfall and availability and quality of forage, and many now lack access to traded staples and essential commodities. Household food insecurity, malnutrition, and migration of impoverished households to provincial centers and the capital of Ulaanbaatar are one result. Reductions to investments in the health sector have also eroded the quality of services in rural areas, and restricted access to those services still functioning. Evidence suggests that women are particularly vulnerable to these political-ecological changes, and that this vulnerability is manifested in increasing rates of poor reproductive health and maternal mortality. Drawing on case-study ethnographic and epidemiological data, this article explores the links between neoliberal economic reform and maternal mortality in Mongolia. PMID- 15272807 TI - Deadly inequality in the health care "safety net": uninsured ethnic minorities' struggle to live with life-threatening illnesses. AB - Forty-three million Americans are uninsured. This article explores the difficulties people experience in seeking health care through the health care "safety net," which provides most of the health care that uninsured people receive, and critiques the gaps, inconsistencies, and failures of such care. In research with 176 African Americans and Latinos who had no health insurance, it was found that they delay seeking care because of cost, do without medications, have negative views of safety net health care, and experience discrimination. As a consequence of dissatisfaction with safety net care, avoidance of the health care system was commonplace. It is concluded that safety net health care facilitates the development of unhealthy practices, such as delays in seeking care. The inadequacy of safety net health care is thus injurious to people's health. PMID- 15272808 TI - Population ageing in the Balkan countries. AB - INTRODUCTION: As we enter the third millennium, we are witnessing an unprecedented rapid expansion of the population of older persons in both the developed and developing world. The total worldwide aged population (aged 60 and older) is projected to rise from 605 million in 2000 to 1.2 billion in 2025 and to nearly 2 billion in 2050. AIM: To study the process of population ageing in Bulgaria and Balkan countries and to reveal its main characteristics, similarities and differences as compared to other European countries. MATERIAL AND METHODS: Primary information was obtained from the European Database "Health for all", the European Public Health Information Network for Eastern Europe and the US Census Bureau International Database. Total, male and female old and youth dependency ratios and ratio of people over 65 to 0-14 years were calculated and trends were followed for 30 years. Countries were ranked and compared with Europe and East European countries. RESULTS: From 1970 to 1999 the old dependency ratio increased dramatically in Bulgaria (14.2-23.6%), Greece (17.2-24.7%), Romania and Slovenia. The ratio of old to young people surpassed 100% in Greece (107.3%, for female 122.6%) and in Bulgaria (99.7% and 116.8%, respectively). Albanian and Turkish populations differ substantially from the other Balkan countries. DISCUSSION: Population ageing creates great socio-economic and health problems- slower economic growth, lack of young labour force, higher tax charges on working age population, etc. The process in Balkan countries is aggravated by the rise of inflation, unemployment and other socio-economic factors. CONCLUSION: All Balkan countries face multiple challenges posed by population ageing. Most of them experience the same demographic tendencies as for market economy countries even though their economic development is far worse. This is especially true for Bulgaria. PMID- 15272809 TI - A study of the effect of age on depressivity in Bulgarian urban population. AB - AIM: Depression is the most prevalent mental disorder in the population. The aim of the present study is to investigate the effect of age on depressivity. MATERIAL AND METHODS: The study is community-based survey and includes 12,600 persons over 15 years of age. The design is multi-stage randomized cluster double sampling. The short self-evaluating screening test for depression of Roglev (STD 1-R) was administered. Gathered data were analyzed with descriptive statistics, Student's t-test, ANOVA with Tukey's Honestly Significant Difference Test and chi2-test with Bonferroni correction for multiple comparisons, using SPSS 10.0. RESULTS AND CONCLUSIONS: The prevalence of depressivity increases steadily with age. The lowest rate of depressivity is found in the 15-45 age group, while the highest rate--in the senior age group. PMID- 15272810 TI - The holistic approach to rehabilitation of patients after total hip joint replacement. AB - AIM: The objective of the present study was to present the principles of a program used jointly by the Clinic of Orthopaedics and Traumatology and the Clinic of Physical Medicine and Early Rehabilitation at St. George University Hospital, Plovdiv for rehabilitation of patients with total hip replacement surgery. MATERIALS AND METHODS: We reviewed 486 cases of endoprostheses implanted by two surgical teams in the Clinic of Orthopaedics and Traumatology over a period of 6 years. We lay the stress on the fact that early mobilization of patients (3 to 5 days after operation) results in better functional results. We particularly emphasize the necessity of using a comprehensive approach that can have an effect on the psychoemotional status and the motivation of patients. RESULTS: The treatment results we evaluated by taking into account not only all functional parameters of restoring the range of hip joint motion but also the psychoemotional aspects of the recovery of patients after surgery. We took into consideration the range of motion of both the operated and the unoperated hip joints, the muscle strength, the walking distance, the type of gait, and the subjective evaluation of the patients (satisfied, not very satisfied, unsatisfied of operation). Particular attention was paid to the psychoemotional status of patients stimulating the positive feedback and their willingness to put in serious efforts toward improvement of their quality of life. CONCLUSION: The new approach to rehabilitation of patients with total hip replacement focuses not only on the collaboration between orthopedists, traumatologists and physical therapeutists, but also on the integral effect both on the physical state of patients and on their psychoemotional status with the aim of improving their quality of life. PMID- 15272811 TI - Surgical treatment of bronchobiliary fistulas due to complicated echinococcosis of the liver: case report and literature review. AB - Echinococcosis still remains a widespread disease associated with the development of a number of complications. The present study presents three patients, successfully treated surgically of bronchobiliary fistulas--in two of the patients the fistulas were due to complicated echinococcosis of the liver, and in the third patient the fistula was a result of a hepatic abscess which developed in a residual cavity after echinococcectomy. The patients were discharged from hospital in good health. The authors consider thoracophrenotomy to be the method of choice in the surgical treatment of patients with bronchobiliary fistulas; it allows performance of echinococcectomy and elimination of the bronchobiliary fistula. The results are discussed in relation to relevant data in literature. PMID- 15272812 TI - Anti-lactoferrin antibodies in patients with connective tissue diseases. AB - AIM: To study the prevalence of antineutrophil cytoplasmic antibodies against lactoferrin in patients with connective tissue diseases and to characterize the antibody response to lactoferrin in these patients. METHODS: Sera from 60 patients with systemic lupus erythematosus, 98 with rheumatoid arthritis, 11 with systemic sclerosis, and 6 with mixed connective tissue disease were studied. The presence of anti-lactoferrin antibodies was detected by enzyme-linked immunosorbent assay. The IgG subclass reactivity of antibodies to lactoferrin was analyzed by biotin-extravidin amplified enzyme-linked immunosorbent assay and monoclonal antibodies to human IgG subclasses. RESULTS: Anti-lactoferrin antibodies were found in 15 patients (5 patients with rheumatoid arthritis and 10 with systemic lupus erythematosus). IgG1 was the predominant subclass for antibodies to lactoferrin. CONCLUSION: Patients with connective tissue diseases are known to develop multiple auto-antibodies; anti-lactoferrin antibodies mainly of IgG1 isotype can also be found in patients with rheumatoid arthritis and more often in patients with systemic lupus erythematosus. PMID- 15272813 TI - Formation of the basal lamina in human embryonal adipose cells- immunohistochemical and ultrastructural evidence. AB - AIM: The aim of the present study was to investigate the formation of the basal lamina in adipose cells during human embryonal development using transmission electronmicroscopy and immunohistochemistry (reactions for collagen type IV and laminin). MATERIAL AND METHODS: Material (subcutaneous tissue from the gluteal region) was taken from human embryos aged 6 to 12 weeks of gestation. Part of the material was prepared for immunohistochemical investigation for collagen type IV and laminin by the peroxidase-antiperoxidase (PAP) method with universal PAP mouse kit (Dako Corporation, Santa Barbara, CA, USA) and primary antibodies- monoclonal mouse anti-human collagen type IV in dilution 1:100 and monoclonal mouse anti-human laminin in dilution 1:100 (Dako Corporation, Santa Barbara, CA, USA). Part of the same material was frozen at -18 degreesC and on fresh 5 microm thick cryostat sections Sudan III-hematoxylin staining was performed for lipids' demonstration after Daddi (1986). Another part of the same material was prepared for electron microscopic examination with TEM Philips CM 12. RESULTS: The results showed that from the 6th week of gestation onward human preadipose cells accumulated lipid droplets in their cytoplasm and expressed positive immunoreactivity for collagen type IV and laminin. First signs of basal lamina could be seen at places outside the plasmalema. Well-developed rough endoplasmatic reticulum was observed which could be associated with the production of collagen and laminin. A time and place related expression was seen in the immunohistochemical and the electronmicroscopical results. CONCLUSION: Formation of the basal lamina of the human adipocytes begins with the onset of adipogenesis and differentiation during the early human embryogenesis (6th week of gestation). The role of collagen type IV and laminin molecules is suggested in this process. PMID- 15272814 TI - Early clinical results from the use of 5% potassium nitrate in polycarboxylate cement for biological treatment of reversible pulpitis. AB - The aim of the present study was to obtain immediate and early clinical results from the use of 5% potassium nitrate (KNO3) in polycarboxylate cement for biological treatment of reversible pulpitis. MATERIAL AND METHODS: The method was applied in 140 teeth, mostly premolars and molars, with a clinical diagnosis of reversible pulpitis and indications for biological treatment of the inflamed dental pulp by means of indirect pulp capping. All teeth had clinically healthy periodontium and no non-carious lesions of the hard dental tissues. The study sample consisted of 127 patients aged between 16 and 40 years. The control group included 92 teeth in 78 patients, which were treated by a calcium hydroxide liner. The follow-up examinations were conducted 24 hours after treatment and on day 3 for the immediate results, and on days 7, 14, 30, and 90 for the early clinical results assessing the functional condition of the teeth, presence of thermal stimulus-induced pain and the measurements from the electric pulp vitality tests. The results were analysed with the analysis of variance and graphic analysis. RESULTS: The immediate and early clinical results of the biological treatment of reversible pulpitis with 5% KNO3 in polycarboxylate cement show that pain as a symptom of the initial inflammatory process in the dental pulp is rapidly and effectively relieved, leaving the treated teeth in good functional condition. The comparison of the measurements from the electric pulp tests of all teeth reveals a statistically significant difference (u = 8.51; P < 0.001) in favour of the group treated with potassium nitrate and polycarboxylate cement. CONCLUSIONS: 1. The immediate and early clinical results of our study suggest that treating reversible pulpitis with 5% KNO3 in polycarboxylate cement has an a very good analgesic action 2. The normal electrical excitability of the treated teeth is restored faster than that in the teeth treated by calcium hydroxide-based materials (P < 0.001). PMID- 15272815 TI - Effectiveness and ergonomic features of different light sources for polymerization of composite resin materials. AB - INTRODUCTION: Besides the halogen curing lights, already accepted as standard equipment, diode and plasma-arc units for polymerization of filling materials have also been commercially available in Bulgaria in recent years. The present study was prompted by the lack of detailed scientific information about these novel light curing devices. AIM: 1. To evaluate the quality of polymerization achieved by different light curing units by measuring the quantity of residual monomer in the test specimens. 2. To classify the units according to their convenience features and polymerization abilities. MATERIALS AND METHODS: Five diode units from three different manufacturers, one plasma-arc unit and one halogen unit were used. The test specimens, made from the hybrid photopolymerizing material Illumine (Ortho Plus, France), Type II, radiopaque, color C1, were 2 mm high and 4 mm in diameter. They were cured for 40 seconds with the respective unit after which they were ground and 25 mg from each sample were immersed in 8 ml of 95% ethanol for extraction of the residual monomer. The quantity of the residual monomer was determined by the method of the standard straight line. Statistical analysis was performed with the analysis of variance and F-test. The comparisons in each group were made by the Tukey-Kramer test, the level of significance for the null hypothesis being P = 0.05. The complex evaluation of the units was made with the help of a scale, developed by us. RESULTS: The material polymerized by the halogen light demonstrated the smallest quantities of residual monomer of all the specimens. There was no statistically significant difference between the quantities of the residual monomer in the specimens polymerized by the plasma arc unit (PAC), OL2 and Elipar Free Light exp. Such a difference, however, existed when comparing the PAC unit and the 3M ESPE unit, working in a continuous irradiation mode, the former having more residual monomer. Polymerization by Elipar Free Light produced the smallest quantities of residual monomer of all other diode lights. The only exception was the comparison between OL2 and Elipar Free Light exp., where there was no difference. The diode light of 3M ESPE performed best of all units according to the criteria we used for evaluation of the light curing devices. CONCLUSIONS: 1. The intensity of radiation of the light source and the duration of polymerization are of crucial importance for the complete conversion of the monomer into polymer. 2. Diode technology has not yet been able to reach any halogen equipment with respect to quality of polymerization, despite some of its unquestionable advantages. 3. When choosing a light curing unit, the quality of polymerization is more important than the price and the offered additional features. PMID- 15272816 TI - Diagnostic difficulties in a case of isolated sarcoidosis of the nose and sinuses. AB - Sarcoidosis still remains a diagnosis of exclusion. It is unusual for the disease to be localised in the nose and sinuses and to manifest its symptoms in this site; the diagnosis in such cases is rather a difficult task. There is no symptom pathognomonic of the disease. A major role in the diagnosis of sarcoidosis is played by histologic evidence on the basis of which additional tests can be used. We present a case of primary sarcoidosis of the nose and sinuses which involved the orbit and had non-specific symptoms in the sinonasal region. The diagnosis of sarcoidosis was made ultimately only after decalcification of sample material taken from the ethmoidal labyrinth. Interspersed among the bone trabeculae there were the typical epithelioid cellular granulomas composed of epithelioid cells with round nuclei and prominent nucleoli, Langhans'-type giant cells, and a tender rim of chronic inflammatory infiltrate in the periphery. After initial beneficial response to the administered cortisone therapy (40 mg prednisolone daily for 6 months and then a daily maintenance dose of 10 mg) the control examination showed that the peri- and retrobulbar infiltrate persisted and the eye symptoms recurred. We therefore increased the dose and proceeded to a pulse therapy (120 mg of urbason daily and a daily maintenance dose of 30 mg). At present the patient is still receiving this therapy which has reduced as a result the local manifestations but Cushing's syndrome has developed as a side effect. PMID- 15272817 TI - Quantitative assessment of pterygomasseteric hypertrophy. AB - The aim of this study was to quantify the pterygomasseteric hypertrophy in a clinical case with significant facial asymmetry and temporomandibular joint dysfunction. Magnetic resonance imaging (MRI) technique was used to measure the surface areas of affected and contralateral pterygomasseteric muscle groups on six coronal scans. The results suggest that MRI is a very effective method for quantification of muscular hypertrophy and better explanation of associated clinical findings, therefore motivating the need to preserve or restore the bilateral masticatory function. PMID- 15272818 TI - [The study of mosquito fauna (Diptera, Culicidae) in Volgograd city in light of the outbreak of West Nile fever in Volgograd region, 1999]. AB - Mosquito sampling was carried out in Volgograd city and its vicinity in August 2001 and 2002. In total 16,000 individuals belonging to 6 genera and 12 species were collected. Nine species were anthropophilic. Culex modestus and Aedes vexans dominated in all outdoor samples collected in Volgograd city. In addition to these species, Coquillettidia richiardii and Ae. caspius were abundant in the vicinity of Volgograd. Autogenous Cx. popiens dominated among six species sampled indoors in Volgograd city. The possible role of different mosquito species in West Nile virus circulation in Volgograd city is discussed. PMID- 15272819 TI - [Morphofunctional changes in the midgut of tick nymphs of the genus Ixodes (Acarina: Ixodidae) during and after feeding]. AB - The midgut epithelium of feeding nymph is represented by the digestive cells of larval phase. Digestion of the main part of feed is performed by the one generation of digestive cells of nymphal phase after detachment, during moult. This period precedes the apolysis. The generation of secretory cells is absent on the nymphal phase. Secretory vacuoles are formed in the digestive cells of larval phase. All functioning cells form a peritrophic matrix on their apical surface. The replacement of the digestive cells of larval phase by the digestive cells of nymphal phase proceeds gradually, during the first 5-10 days after detachment. The beginning of the accumulation of digestive inclusions in the young digestive cells of nymphal phase takes place in the 10-15 days after detachment. PMID- 15272820 TI - [Morphology of Urospora chiridotae (Sporozoa: Gregarinomorpha: Eugregarinida) from sea cucumber Chiridota laevis (Echinodermata: Holothuroidea: Apoda)]. AB - Several morphological forms (morphotypes) of Urospora chiridotae gamontes are found in White Sea holothuroid Chiridota laevis. All these morphotypes are differed by localization in the body of host, form and cytological features. The gregarines are situated in several host biotopes, such as blood vessels, intestine and mesenteries. In the blood vessels elongate skittle-like cells supplied with long thin cytopillia are observed. On the external surface of the intestine spherical gregarines are found. These parasites commonly covered with one layer of coelomic epithelium's cells. In some holothuria intratissue spherical cells of parasites located in intestinal epithelium are presented. Both of these types of parasites lack cytopillia, and folds or ridges on its surface. On different mesenteries, connections between intestine and body wall, and also on intestine elongate ounce-shaped cells and gamontocysts are observed. These cells are situated on the apices of finger-like processes of the intestine and mesenteries surface. Ounce-shaped gregarines have cytopillia shorter than in skittle-like gregarines. The differences between morphotypes of Urospora chiridotae are probably caused by different environmental conditions. In the narrow rift of blood vessel elongate cells are developed. The cytopillia may serve for making more or less wide space around gregarines, which is necessary for food uptake. Spherical cells surrounded by host's cells and have the form typical for tissue parasites. In the wide coelomic cavity where convection of liquid proceeds better than in blood vessel, ounce-shaped gregarines with short cytopillia are developed. We found only typical for Urospora chiridotae ovoid oocysts with dissimilar ends, anterior collar and spine-like posterior end. Thus, the all above-mentioned morphotypes undoubtedly belong to the same species. The relationships between defense host cells and the different morphotypes of trophozoites are variable. PMID- 15272821 TI - [Microsporidiosis in the wax moth Galleria mellonella (Lepidoptera: Pyralidae) caused by Vairimorpha ephestiae (Microsporidia: Burenellidae)]. AB - An experimental microsporidiosis of the wax moth caterpillars from laboratory population had been caused by oral infecting of early stages larvae and by intracavity injections of the spores of the microsporidian species Vairimorpha ephestiae. Peculiarities of microsporidiosis proceeding, manifestations of host defence reactions, and also an effect of the temperature of caterpillars cultivation and conditions of spores keeping on liability of the insects to the infection were studied. The effect of the microsporidia on the host organism was the early death or the delay of larvae development, but in several cases external manifestations of the effect of the parasite on the host were absent. The development of the parasites from the moment of infecting to the appearing of the mature spores congestions in the host organism proceeded 6 days. Microsporidia invaded insect fat body and caused its hypertrophy and disappearance of lipid granules. In the intestine and salivary glands microsporidia were not observed in the period from 6 to 16 day of the development. On the final stage of microsporidiosis the all contents of fatty tissue cells were replaced by spores of microsporidia. Under microscope only diplocaryotic spores of the Nozema type had been found in infected and died specimens, but not octospores. The spores threw out polar tubes under the change of pH in incubating solution from neutral to alkaline. The effects of microsporidiosis on the wax moth haemolymph were the increased rate of prohaemocytes, appearing of multinuclear free-circulating cells at 6 day after infection, and suppression of the reaction of haemolymph melanization with the mass sporogenesis of the parasite. The characteristic symptom of the wax moth microsporidiosis had been revealed, accumulation of black points and small spots of irregular form under cuticle ("reaction of attretization"). Increase of the temperature of insect cultivation up to 32 degrees C during 3 days after infection contributed to the full deliverance of the insects from the infection in first and second generations. It can be considered as a method of treatment of wax moth laboratory colonies from microsporidiosis. Oral infection of III and IV stage caterpillars by the spores being kept during 3-6 months under 4 degrees C in form of water suspension caused the death of 63.0-61.5 and 91% of caterpillars being cultivated under 25 and 21 degrees C respectively. Under the temperature of cultivation equal 30 degrees C the mortality did not differ from the control sample (8-10%). The spores extracted from dried bodies of caterpillars lost their vitality. It was demonstrated by the test on infectious ability in vivo and by acridine orange staining. This host-parasite system appears to be perspective in investigations of resistance mechanisms in insects and immunosuppressive features of entomopathogen microsporidia. PMID- 15272822 TI - [Parasite fauna of amur sleeper Perccottus glehni (Eleotridae) from Primor'e region]. AB - Ecological analysis of parasite fauna associated with Perccottus glehni from water basins of the Primorye has been carried out. The 31 species of parasites have been found: protozoan--10, monogeneans--3, trematodes--6, cestodes--2, proboscis worms--3, nematodes--2, crustaceans--3, hirudinis--1, and molluscs--1 species. PMID- 15272823 TI - [On the validity of the genus Ichthyobronema Gnedina et Savina, 1930 (Nematoda, Spirurida: Quimperiidae)]. AB - The author maintains Moravec's (1994) point of view about attachment of valid taxon status to the genus Ichthyobronema Gnedina et Savina, 1930 with type and single species I. hamulatum (Moulton, 1931) Moravec, 1994. PMID- 15272824 TI - [Experimental study of the infecting ability of the flea Coptopsylla lamellifer rostrata in the Kyzylkum natural focus of plague]. AB - Experimental infecting of the fleas Coptopsylla lamellifer rostrata by plague was carried out. The fleas were infected by feeding through biomembrane (skin of a white mouse) with a mixture of defibrinated blood of guinea-pig and plague microbes. Under concentration 2-3 milliard of the microbes in 1 millilitre, from 60 to 100% of the fleas were infected. Forming of the block of proventriculus was observed in 9-13 day. Mean percent of blocking for all experiments was less than 1%. Maximal rate of blocking (2.8%) was observed in the experiment under everyday 3-hour feeding. After planting of the fleas with blocks on great gerbil (2 fleas on one gerbil) 20% of the gerbils were infected. The conclusion had been made that C. lamellifer rostrata has only insignificant effectiveness as a vector of plague agent in Kyzylkum. Probably it is caused by some specific features of this subspecies. PMID- 15272825 TI - [The first finding of Corynosoma shackletoni Zdzitowiecki, 1978 (Acanthocephala: Polymorphidae) in marine mammals]. AB - The acanthocephal Corynosoma shackletoni Zdzitowiecki, 1978 is recorded for the first time as the parasite of sea leopard Hydrurga leptonix (Blainville, 1820) from the Pacific sector of Antarctica. A description and drawings of examined specimens are given. PMID- 15272826 TI - Structural interpretation of the topological index. 2. The molecular connectivity index, the Kappa index, and the atom-type E-State index. AB - The structural interpretation is extended to the topological indices describing cyclic structures. Three representatives of the topological index, such as the molecular connectivity index, the Kappa index, and the atom-type E-State index, are interpreted by mining out, through projection pursuit combining with a number theory method generating uniformly distributed directions on unit sphere, the structural features hidden in the spaces spanned by the three series of indices individually. Some interesting results, which can hardly be found by individual index, are obtained from the multidimensional spaces by several topological indices. The results support quantitatively the former studies on the topological indices, and some new insights are obtained during the analysis. The combinations of several molecular connectivity indices describe mainly three general categories of molecular structure information, which include degree of branching, size, and degree of cyclicity. The cyclicity can also be coded by the combination of chi cluster and path/cluster indices. The Kappa shape indices encode, in combination, significant information on size, the degree of cyclicity, and the degree of centralization/separation in branching. The size, branch number, and cyclicity information has also been mined out to interpret atom-type E-State indices. The structural feature such as the number of quaternary atoms is searched out to be an important factor. The results indicate that the collinearity might be a serious problem in the applications of the topological indices. PMID- 15272827 TI - Periodic systems of molecules as elements of Shchukarev's "supermatrix", i.e. the chemical element periodic system. AB - Generations of Soviet scientists contributed invaluable insights into molecular classification. Unfortunately, this research is little appreciated in much of the world. Among these workers S. A. Shchukarev was of great importance. His and his followers' legacy includes a host of graphical displays showing enthalpies of formation of gaseous molecules from free atoms DeltaH(a) and standard enthalpies of formation of substances plotted on the atomic number of the central elements, on their oxidation states, their internuclear separations, and other variables for a wide range of molecules. These graphs serve as databases, from which data can be extracted, to moderate precision, visually. We discuss graphs for one very limited set, or "pleiade" (gas-phase oxides of nitrogen), and for three much broader sets, or subsystems (gas-phase fluorides of all main subgroup atoms and oxides of transition-metal atoms in gas-phase and in STP conditions). When dissolved in water, molecules lose their identities but periodicity is echoed in the acids and aquocations that are formed. We show, as an example in tabular form, that redox potentials of high-oxygen acids containing S, Se, and Te change concomitantly with DeltaH(a ) and DeltaH(f) of their hexafluorides. We present graphical evidence that three properties for cations of groups 1-3 (in the short version of the periodic chart) behave similarly and share the periodicity of the elements. One of the properties is related to the ionization potential, which is shown in a tabular example to vary concomitantly with energy of hydration. It was the ultimate goal of S. A. Shchukarev that the transformation of any one graphical database into any other, having different molecules under different conditions, would be made mathematically. PMID- 15272828 TI - The most stable class of benzenoid hydrocarbons-new topological correlations of strain-free total resonant sextet benzenoids. AB - While briefly reviewing how the concepts of strictly pericondensed, strain-free, Clar's aromatic sextet, and symmetry are interconnected in the topological correspondence between strictly pericondensed and total resonant sextet (TRS) benzenoid hydrocarbons, new structural correlations in isomer numbers, symmetry distributions, and empty rings between various strain-free TRS benzenoids made up of only fused hexagonal rings are presented. These correlations are made evident by the Formula Periodic Table for Total Resonant Sextet Benzenoid Hydrocarbons [Table PAH6(sextet)] and application of the leapfrog operation. A new perimeter topology relationship for fused strain-free TRS benzenoids is derived. This work represents a contribution toward understanding formula/structure relationships of benzenoid hydrocarbons. PMID- 15272829 TI - Topomers: a validated protocol for their self-consistent generation. AB - The hypothesis underlying topomer development is that describing molecular structures consistently may be at least as productive as describing them more realistically but incompletely. A general protocol is detailed for deterministically generating self-consistent shapes of molecular fragments from their topologies. These and other extensions to the topomer methodology are validated by repetition of earlier benchmark studies. PMID- 15272830 TI - Maximum spectrum of continuous wavelet transform and its application in resolving an overlapped signal. AB - To estimate the number of peaks and to find the individual peak positions in an overlapped signal, a new method called maximum spectrum of continuous wavelet transform (MSCWT) was developed by extracting the maximum coefficients of continuous wavelet transform (CWT). The peak position in MSCWT was the same as that in its original signal. In this process, CWT was performed not on a single dilation but on an appreciation dilation range. To obtain such a range, a new criterion was introduced to choose a center dilation, which was used to form the dilation range. If Cdilation denoted the center dilation, the proper dilation range was [Cdilation -6 +/- 2, Cdilation +1 +/- 1]. The Mexican Hat function was an analytical wavelet. Utilizing the information of the peak number and the position detected by MSCWT, a fitting route was performed to recover the original signal. One simulated and four true overlapped signals, including high performance liquid chromatography (HPLC), ultraviolet-visible (UV) spectrum, and differential pulse voltammetric (DPV), were processed, and the results indicated that MSCWT could detect an overlapped peak number and position, and the curve fitting based on information of MSCWT had a higher accuracy. The proposed method was an efficient one in resolving different types of overlapped signals. PMID- 15272831 TI - A protein folding degree measure and its dependence on crystal packing, protein size, secondary structure, and domain structural class. AB - Comparing two or more protein structures with respect to their degree of folding is common practice in structural biology despite the fact that there is no scale for a folding degree. Here we introduce a formal definition of a folding degree, capable of quantitative characterization. This enables ordering among protein chains based on their degree of folding. The folding degree of a data set of 152 representative nonhomologous proteins is then studied. We demonstrate that the variation in the folding degree seen for this data set is not due to crystallization artifacts or experimental conditions, such as resolution, refinement protocol, pH, or temperature. A good linear relationship is observed between the folding degree and the percentages of secondary structures in the protein. The folding degree is able to account for the small changes produced in the structure due to crystal packing and temperature. Automating the classification of proteins into their respective structural domain classes, namely mainly-alpha, mainly-beta, and alpha-beta, is also possible. PMID- 15272832 TI - A FAST pattern matching algorithm. AB - The advent of digital computers has made the routine use of pattern-matching possible in various applications. This has also stimulated the development of many algorithms. In this paper, we propose a new algorithm that offers improved performance compared to those reported in the literature so far. The new algorithm has been evolved after analyzing the well-known algorithms such as Boyer-Moore, Quick-search, Raita, and Horspool. The overall performance of the proposed algorithm has been improved using the shift provided by the Quick-search bad-character and by defining a fixed order of comparison. These result in the reduction of the character comparison effort at each attempt. The best- and the worst- case time complexities are also presented in this paper. Most importantly, the proposed method has been compared with the other widely used algorithms. It is interesting to note that the new algorithm works consistently better for any alphabet size. PMID- 15272833 TI - Comparative study of QSAR/QSPR correlations using support vector machines, radial basis function neural networks, and multiple linear regression. AB - Support vector machines (SVMs) were used to develop QSAR models that correlate molecular structures to their toxicity and bioactivities. The performance and predictive ability of SVM are investigated and compared with other methods such as multiple linear regression and radial basis function neural network methods. In the present study, two different data sets were evaluated. The first one involves an application of SVM to the development of a QSAR model for the prediction of toxicities of 153 phenols, and the second investigation deals with the QSAR model between the structures and the activities of a set of 85 cyclooxygenase 2 (COX-2) inhibitors. For each application, the molecular structures were described using either the physicochemical parameters or molecular descriptors. In both studied cases, the predictive ability of the SVM model is comparable or superior to those obtained by MLR and RBFNN. The results indicate that SVM can be used as an alternative powerful modeling tool for QSAR studies. PMID- 15272834 TI - Support vector machines-based quantitative structure-property relationship for the prediction of heat capacity. AB - The support vector machine (SVM), as a novel type of learning machine, for the first time, was used to develop a Quantitative Structure-Property Relationship (QSPR) model of the heat capacity of a diverse set of 182 compounds based on the molecular descriptors calculated from the structure alone. Multiple linear regression (MLR) and radial basis function networks (RBFNNs) were also utilized to construct quantitative linear and nonlinear models to compare with the results obtained by SVM. The root-mean-square (rms) errors in heat capacity predictions for the whole data set given by MLR, RBFNNs, and SVM were 4.648, 4.337, and 2.931 heat capacity units, respectively. The prediction results are in good agreement with the experimental value of heat capacity; also, the results reveal the superiority of the SVM over MLR and RBFNNs models. PMID- 15272835 TI - Similarity search profiles as a diagnostic tool for the analysis of virtual screening calculations. AB - An analysis method termed similarity search profiling has been developed to evaluate fingerprint-based virtual screening calculations. The analysis is based on systematic similarity search calculations using multiple template compounds over the entire value range of a similarity coefficient. In graphical representations, numbers of correctly identified hits and other detected database compounds are separately monitored. The resulting profiles make it possible to determine whether a virtual screening trial can in principle succeed for a given compound class, search tool, similarity metric, and selection criterion. As a test case, we have analyzed virtual screening calculations using a recently designed fingerprint on 23 different biological activity classes in a compound source database containing approximately 1.3 million molecules. Based on our predefined selection criteria, we found that virtual screening analysis was successful for 19 of 23 compound classes. Profile analysis also makes it possible to determine compound class-specific similarity threshold values for similarity searching. PMID- 15272836 TI - Zigzags, railroads, and knots in fullerenes. AB - Two connections between fullerene structures and alternating knots are established. Knots may appear in two ways: from zigzags, i.e., circuits (possibly self-intersecting) of edges running alternately left and right at successive vertices, and from railroads, i.e., circuits (possibly self-intersecting) of edge sharing hexagonal faces, such that the shared edges occur in opposite pairs. A z knot fullerene has only a single zigzag, doubly covering all edges: in the range investigated (n /= 38, all chiral, belonging to groups C(1), C(2), C(3), D(3), or D(5). An r-knot fullerene has a railroad corresponding to the projection of a nontrivial knot: examples are found for C(52) (trefoil), C(54) (figure-of-eight or Flemish knot), and, with isolated pentagons, at C(96), C(104), C(108), C(112), C(114). Statistics on the occurrence of z-knots and of z-vectors of various kinds, z uniform, z-transitive, and z-balanced, are presented for trivalent polyhedra, general fullerenes, and isolated-pentagon fullerenes, along with examples with self-intersecting railroads and r-knots. In a subset of z-knot fullerenes, so called minimal knots, the unique zigzag defines a specific Kekule structure in which double bonds lie on lines of longitude and single bonds on lines of latitude of the approximate sphere defined by the polyhedron vertices. PMID- 15272837 TI - On the stability of CoMFA models. AB - Abrupt, smooth, and box methods for the calculation of electrostatic and steric field values in the comparative molecular field analysis (CoMFA) 3D QSAR technique are assessed on three diverse data sets of medicinal chemistry interest. While the standard CoMFA settings are robust to small changes in the position of the lattice, superior results may sometimes be obtained by use of only one field. However, if only the electrostatic field is used, then sometimes large differences between models are apparent. This appears to be due to a lack of column dropping, and these difficulties can be remedied by use of the box method. PMID- 15272838 TI - Identification of symmetries in molecules and complexes. AB - An algorithm is presented that quickly detects local and global symmetries of single molecules and complexes. Based upon the Morgan Naming Algorithm, the algorithm involves traversing the molecule from a starting atom and building up a molecule name based upon the names of the atoms encountered along the traversal. Additional molecule names are generated from other starting atoms, and name-name matches are identified as corresponding to symmetry operations. A number of enhancements relative to prior methods yield increased efficiency and extended functionality. In particular, the present method detects not only global symmetries but also local symmetries associated with bond rotations as well as symmetries that are only apparent when alternate resonance forms are considered. Importantly, the present method works not only for single molecules but also for multimolecular complexes. As a consequence, it is well, and perhaps uniquely, suited to applications in supramolecular and host-guest chemistry. Applications include filtering out redundant conformations during conformational searching and free energy calculations; accelerating ligand-receptor docking calculations by reducing the sampling ranges of rotatable bonds linked to locally symmetric groups, such as phenyls; and automating the calculation of symmetry numbers for thermochemical applications. PMID- 15272839 TI - Addition patterns in carbon allotropes: independence numbers and d-codes in the Klein and related graphs. AB - The problem of predicting stoichiometries and patterns of chemical addition to a carbon framework, subject solely to the restriction that each addend excludes neighboring sites up to some distance d, is equivalent to determination of d codes of a graph, and for d = 2 to determination of maximum independent sets. Sizes, symmetries, and numbers of d-codes are found for the all-heptagon Klein graph (prototype for "plumber's nightmare" carbon) and for three related graphs. The independence number of the Klein graph is 23, which increases to 24 for a related, but sterically relaxed, all-heptagon network with the same number of vertices and modified adjacencies. Expansion of the Klein graph and its relaxed analogue by insertion of hexagonal faces to form leapfrog graphs also allows all heptagons to achieve their maximum of 3 addends. Consideration of the pi system that is the complement of the addition pattern imposes a closed-shell requirement on the adjacency spectrum, which typically reduces the size of acceptable independent sets. The closed-shell independence numbers of the Klein graph and its relaxed analogue are 18 and 20, respectively. PMID- 15272840 TI - Accurate classification of homodimeric vs other homooligomeric proteins using a new measure of information discrepancy. AB - It has been shown that protein primary sequence encodes quaternary structure information. In this present work, function of degree of disagreement (FDOD), a new measure of information discrepancy, is applied to discriminating between homodimers and other homooligomeric proteins from the primary structure. This new approach is based on subsequence distributions of the primary sequences, so the effect of residue order on protein structure is taken into account. When the length of subsequence is 4, the overall accuracy of the 10-fold cross-validation test attains to 82.5%, which is much better than that of the previous method on the same data set. Our tests demonstrate that the residue order along protein sequences plays an important role in the prediction of homooligomers. In addition, our results suggest that FDOD measure is a simple and powerful tool for the prediction of protein multimeric states. PMID- 15272841 TI - Use of computer-assisted methods for the modeling of the retention time of a variety of volatile organic compounds: a PCA-MLR-ANN approach. AB - A hybrid method consisting of principal component analysis (PCA), multiple linear regressions (MLR), and artificial neural network (ANN) was developed to predict the retention time of 149 C(3)-C(12) volatile organic compounds for a DB-1 stationary phase. PCA and MLR methods were used as feature-selection tools, and a neural network was employed for predicting the retention times. The regression method was also used as a calibration model for calculating the retention time of VOCs and investigating their linear characteristics. The descriptors of the total information index of atomic composition, IAC, Wiener number, W, solvation connectivity index, X1sol, and number of substituted aromatic C(sp(2)), nCaR, appeared in the MLR model and were used as inputs for the ANN generation. Appearance of these parameters shows the importance of the dispersion interactions in the mechanism of retention. Comparison of the MLR and 5-2-1 ANN models indicates the superiority of the ANN over that of the MLR model. The values of 0.913 and 0.738 were obtained for the standard error of prediction set of MLR and ANN models, respectively. PMID- 15272842 TI - Detailed mechanism generation. 1. Generalized reactive properties as reaction class substructures. AB - This paper, the first in a series on the automatic generation of detailed hydrocarbon mechanisms for hydrocarbons, describes how generalized reaction types, specifically the 25 types outlined by Curran et al. for heptane and isooctane combustion, can be translated into general reaction classes. Each type description from Curran et al. ranges from specific, such as differentiating among 1 degrees, 2 degrees, and 3 degrees centers, to nonspecific such as specifying intermediate specific as "products". In the latter case, additional interpretation of the reaction type must be extrapolated from other sources. The types are translated to the substructure "patterns" representing reaction classes. The success of fully translating all the rate and structural information described in these reaction types as they are defined in the literature demonstrates the power of the approach used here. A comparison of the generated reactions using these 25 types with that of a hand-produced heptane mechanism showed that, aside from well-defined exceptions, the set of reactions were almost identical. Often just the elimination of selected species (and sometimes reactions) was enough to make them identical. A notable exception was the use of lumped species in the hand-produced mechanism. The purpose of this paper is to show that an established set of reaction type descriptions can be translated to reaction classes suitable for automatic mechanism generation. A further goal of this paper is to show that these classes can be used to generate a detailed mechanism that mimics a hand produced one. PMID- 15272843 TI - Detailed mechanism generation. 2. Aldehydes, ketones, and olefins. AB - Generalized reaction classes for the consumption and decomposition of aldehydes, ketones, and olefins are described. These classes are important for generating not only reactions for the consumption of the branching agents of low-temperature hydrocarbon combustion but also reactions of the oxidation of alkenes and the decomposition of cyclic ethers. These reaction classes have been extrapolated from specific reactions of existing validated mechanisms. The reaction patterns making up the class were derived by identifying the reactive center of the specific reactions and the important surrounding functional groups. The rates used currently are definitely "first guesses" based on these specific reactions. The reaction classes in this paper supplement the reaction classes derived from accepted reaction types in the previous paper in this series. The purpose of this paper is to outline a complete (with very few exceptions) set of reaction classes which describe the C(5) and C(6) products of the low-temperature and cyclic ether path in the heptane and isooctane mechanisms. PMID- 15272844 TI - Sequential superparamagnetic clustering for unbiased classification of high dimensional chemical data. AB - For the clustering of chemical structures that are described by the Similog, ISIS count, and ISIS binary fingerprints, we propose a sequential superparamagnetic clustering approach. To appropriately handle nonbinary feature keys, we introduce an extension of the binary Tanimoto similarity measure. In our applications, data sets composed of structures from seven chemically distinct compound classes are evaluated and correctly clustered. The comparison, with results from leading methods, indicates the superiority of our sequential superparamagnetic clustering approach. PMID- 15272845 TI - "In silico" design of new uranyl extractants based on phosphoryl-containing podands: QSPR studies, generation and screening of virtual combinatorial library, and experimental tests. AB - This paper is devoted to computer-aided design of new extractants of the uranyl cation involving three main steps: (i) a QSPR study, (ii) generation and screening of a virtual combinatorial library, and (iii) synthesis of several predicted compounds and their experimental extraction studies. First, we performed a QSPR modeling of the distribution coefficient (logD) of uranyl extracted by phosphoryl-containing podands from water to 1,2-dichloroethane. Two different approaches were used: one based on classical structural and physicochemical descriptors (implemented in the CODESSA PRO program) and another one based on fragment descriptors (implemented in the TRAIL program). Three statistically significant models obtained with TRAIL involve as descriptors either sequences of atoms and bonds or atoms with their close environment (augmented atoms). The best models of CODESSA PRO include its own molecular descriptors as well as fragment descriptors obtained with TRAIL. At the second step, a virtual combinatorial library of 2024 podands has been generated with the CombiLib program, followed by the assessment of logD values using developed QSPR models. At the third step, eight of these hypothetical compounds were synthesized and tested experimentally. Comparison with experiment shows that developed QSPR models successfully predict logD values for 7 of 8 compounds from that "blind test" set. PMID- 15272846 TI - Representation of the molecular topology of cyclical structures by means of cycle graphs. 2. Application to clustering of chemical databases. AB - The great size of chemical databases and the high computational cost required in the atom-atom comparison of molecular structures for the calculation of the similarity between two chemical compounds necessitate the proposal of new clustering models with the aim of reducing the time of recovery of a set of molecules from a database that satisfies a range of similarities with regard to a given molecule pattern. In this paper we make use of the information corresponding to the cycles existing in the structure of molecules as an approach for the classification of chemical databases. The clustering method here proposed is based on the representation of the topological structure of molecules stored in chemical databases through its corresponding cycle graph. This method presents a more appropriate behavior for others described in the bibliography in which the information corresponding to the cyclicity of the molecules is also used. PMID- 15272847 TI - Use of electron density critical points as chemical function-based reduced representations of pharmacological ligands. AB - In this paper, we propose a reduced representation of molecules of pharmacological interest based on their chemical functions. The proposed representations of the molecules are obtained by a topological analysis of their electron density maps at medium resolution, leading to graphs of critical points. The distribution of the different types of critical points are compared at various levels of resolution for a training set of 22 molecules in order to define the optimal resolution level leading to the best representation of the various chemical functions. The reduced representations can in the future be used for molecular similarity research and pharmacophore proposals. PMID- 15272848 TI - Data mining and machine learning techniques for the identification of mutagenicity inducing substructures and structure activity relationships of noncongeneric compounds. AB - This paper explores the utility of data mining and machine learning algorithms for the induction of mutagenicity structure-activity relationships (SARs) from noncongeneric data sets. We compare (i) a newly developed algorithm (MOLFEA) for the generation of descriptors (molecular fragments) for noncongeneric compounds with traditional SAR approaches (molecular properties) and (ii) different machine learning algorithms for the induction of SARs from these descriptors. In addition we investigate the optimal parameter settings for these programs and give an exemplary interpretation of the derived models. The predictive accuracies of models using MOLFEA derived descriptors is approximately 10-15%age points higher than those using molecular properties alone. Using both types of descriptors together does not improve the derived models. From the applied machine learning techniques the rule learner PART and support vector machines gave the best results, although the differences between the learning algorithms are only marginal. We were able to achieve predictive accuracies up to 78% for 10-fold cross-validation. The resulting models are relatively easy to interpret and usable for predictive as well as for explanatory purposes. PMID- 15272849 TI - Creating artificial binding pocket boundaries to improve the efficiency of flexible ligand docking. AB - Traditionally, algorithms for binding site characterization or identification focus on the problem of identifying atoms within a macromolecule that might be responsible for ligand binding. In this manuscript, we focus on the binding pocket problem from a different perspective as a challenge of calculating an artificial binding pocket boundary that is sufficient to isolate binding pocket volume. The approach involves the calculation of a macromolecule encapsulating surface (MES) that separates binding pocket volume from outside space. We show that the MES can be used to increase the efficiency of flexible docking as implemented in AutoDock 3.0. The most significant improvement in docking efficiency is seen when the entire protein is searched and results show additional support for the use of AutoDock, in and of itself, as a feasible tool for binding-site identification for cases in which a protein ligand is known. PMID- 15272850 TI - GRID formalism for the comparative molecular surface analysis: application to the CoMFA benchmark steroids, azo dyes, and HEPT derivatives. AB - Shape analysis is a powerful tool in chemistry and drug design, and molecular surface defines shape in the molecular scale. In the current publication we presented a novel formalism for the comparative molecular surface analysis (s CoMSA). The method enables both quantitative modeling of 3D-QSAR and finding possible pharmacophoric sites. The method provides very predictive models for the CBG activity of the benchmark steroid series, tinctorial properties of the heterocyclic azo dyes and anti-HIV activity of the HEPT series. PMID- 15272851 TI - Group vector space method for estimating enthalpy of vaporization of organic compounds at the normal boiling point. AB - The specific position of a group in the molecule has been considered, and a group vector space method for estimating enthalpy of vaporization at the normal boiling point of organic compounds has been developed. Expression for enthalpy of vaporization Delta(vap)H(T(b)) has been established and numerical values of relative group parameters obtained. The average percent deviation of estimation of Delta(vap)H(T(b)) is 1.16, which show that the present method demonstrates significant improvement in applicability to predict the enthalpy of vaporization at the normal boiling point, compared the conventional group methods. PMID- 15272852 TI - Development of QSAR models to predict and interpret the biological activity of artemisinin analogues. AB - This work presents the development of Quantitative Structure-Activity Relationship (QSAR) models to predict the biological activity of 179 artemisinin analogues. The structures of the molecules are represented by chemical descriptors that encode topological, geometric, and electronic structure features. Both linear (multiple linear regression) and nonlinear (computational neural network) models are developed to link the structures to their reported biological activity. The best linear model was subjected to a PLS analysis to provide model interpretability. While the best linear model does not perform as well as the nonlinear model in terms of predictive ability, the application of PLS analysis allows for a sound physical interpretation of the structure-activity trend captured by the model. On the other hand, the best nonlinear model is superior in terms of pure predictive ability, having a training error of 0.47 log RA units (R2 = 0.96) and a prediction error of 0.76 log RA units (R2 = 0.88). PMID- 15272853 TI - Efficient 3D database screening for novel HIV-1 IN inhibitors. AB - We describe the use of pharmacophore modeling as an efficient tool in the discovery of novel HIV-1 integrase (IN) inhibitors. A three-dimensional hypothetical model for the binding of diketo acid analogues to the enzyme was built by means of the Catalyst program. Using these models as a query for virtual screening, we found several compounds that contain the specified 3D patterns of chemical functions. Biological testing shows that our strategy was successful in searching for new structural leads as HIV-1 IN inhibitors. PMID- 15272854 TI - Chemical proteomic tool for ligand mapping of CYP antitargets: an NMR-compatible 3D QSAR descriptor in the Heme-Based Coordinate System. AB - Chemical proteomic strategies strive to probe and understand protein-ligand interactions across gene families. One gene family of particular interest in drug and xenobiotic metabolism are the cytochromes P450 (CYPs), the topic of this article. Although numerous tools exist to probe affinity of CYP-ligand interactions, fewer exist for the rapid experimental characterization of the structural nature of these interactions. As a complement to recent advances in X ray crystallography, NMR methods are being developed that allow for fairly high throughput characterization of protein-ligand interactions. One especially promising NMR approach involves the use of paramagnetic induced relaxation effects to measure distances of ligand atoms from the heme iron in CYP enzymes. Distances obtained from these T(1) relaxation measurements can be used as a direct source of 1-dimensional structural information or to restrain a ligand docking to generate a 3-dimensional data set. To facilitate such studies, we introduce the concept of the Heme-Based Coordinate System and present how it can be used in combination with NMR T(1) relaxation data to derive 3D QSAR descriptors directly or in combination with in silico docking. These descriptors should have application in defining the binding preferences of CYP binding sites using 3D QSAR models. They are especially well-suited for the biasing of fragment assembly and combinatorial chemistry drug design strategies, to avoid fragment or reagent combinations with enhanced affinity for CYP antitargets. PMID- 15272855 TI - Steric trigger as a mechanism for CB1 cannabinoid receptor activation. AB - To determine the moiety that behaves as the steric trigger to activate the CB(1) cannabinoid receptor, conformational properties of the nonclassical cannabinoid CP55244, one of the most potent CB(1) receptor agonists, were characterized by conformational analysis, rotational barrier calculations, and molecular dynamics (MD) simulations. It was shown from the present MD simulations that the torsion angles phi1 and phi4 of the C3 side chain showed the most dramatic change when compared with the ground-state receptor-bound conformation, indicating that rotation around these torsion angles is responsible for releasing the ligand strain energy. Multiple stages would be involved in the ligand conformational change. As a molecular mechanism for the ligand-induced CB(1) receptor conformational change, we propose that the C3 side chain serves as the steric trigger, while the ACD-ring moiety of CP55244 serves as the plug. Steric clash with helices within the binding pocket would induce microconformational adaptation within the protein. This mechanism would suggest that rotational flexibility in a ligand may be as important a determinant of agonist activity as the pharmacophoric elements that can be identified. PMID- 15272856 TI - Global and local computational models for aqueous solubility prediction of drug like molecules. AB - The aim of this study was to develop in silico protocols for the prediction of aqueous drug solubility. For this purpose, high quality solubility data of 85 drug-like compounds covering the total drug-like space as identified with the ChemGPS methodology were used. Two-dimensional molecular descriptors describing electron distribution, lipophilicity, flexibility, and size were calculated by Molconn-Z and Selma. Global minimum energy conformers were obtained by Monte Carlo simulations in MacroModel and three-dimensional descriptors of molecular surface area properties were calculated by Marea. PLS models were obtained by use of training and test sets. Both a global drug solubility model (R(2) = 0.80, RMSE(te) = 0.83) and subset specific models (after dividing the 85 compounds into acids, bases, ampholytes, and nonproteolytes) were generated. Furthermore, the final models were successful in predicting the solubility values of external test sets taken from the literature. The results showed that homologous series and subsets can be predicted with high accuracy from easily comprehensible models, whereas consensus modeling might be needed to predict the aqueous drug solubility of datasets with large structural diversity. PMID- 15272857 TI - Models of steroid binding based on the minimum deviation of structurally assigned 13C NMR spectra analysis (MiDSASA). AB - This paper develops a quantitative k-nearest neighbors modeling technique. The technique is used to demonstrate that a compound's biological binding activity to a receptor can be calculated from the minimum of the square root of the sum of squared deviations (SSSD) of a structurally assigned chemical shift on a template between the unknown compound to be predicted and a set of known compounds with known activities. When building models of biological activity, nonlinear relationships are built into the input training data. If a model is developed by selecting only compounds with minimum structurally assigned chemical shift deviations from the unknown compound, some of the nonlinear relationships can be removed. The smaller the total chemical shift deviation between a compound with known activity and another compound with unknown activity, the more likely it will have similar biological, chemical, and physical properties. This means that a model can be produced without rigorous statistics or neural networks. This technique is similar to structure-activity relationship (SAR) modeling, but instead of relying on substructure fragments to produce a model, this new model is based on minimum chemical shift differences on those substructure fragments. We refer to this method as minimum deviation of structurally assigned spectra analysis (MiDSASA) modeling. Modeling by the minimum deviation concept can be applied to other chemoinformatic data analyses such as metabolite concentrations in metabolic pathways for metabolomics research. A MiDSASA template model for 30 steroids binding the corticosterone binding globulin based on the activity factors of the two nearest compounds had a correlation of 0.88. A MiDSASA template model for 50 steroids binding the aromatse enzyme based on the average activity of the four nearest compounds had a correlation of 0.71. PMID- 15272858 TI - Prediction of P-glycoprotein substrates by a support vector machine approach. AB - P-glycoproteins (P-gp) actively transport a wide variety of chemicals out of cells and function as drug efflux pumps that mediate multidrug resistance and limit the efficacy of many drugs. Methods for facilitating early elimination of potential P-gp substrates are useful for facilitating new drug discovery. A computational ensemble pharmacophore model has recently been used for the prediction of P-gp substrates with a promising accuracy of 63%. It is desirable to extend the prediction range beyond compounds covered by the known pharmacophore models. For such a purpose, a machine learning method, support vector machine (SVM), was explored for the prediction of P-gp substrates. A set of 201 chemical compounds, including 116 substrates and 85 nonsubstrates of P-gp, was used to train and test a SVM classification system. This SVM system gave a prediction accuracy of at least 81.2% for P-gp substrates based on two different evaluation methods, which is substantially improved against that obtained from the multiple-pharmacophore model. The prediction accuracy for nonsubstrates of P gp is 79.2% using 5-fold cross-validation. These accuracies are slightly better than those obtained from other statistical classification methods, including k nearest neighbor (k-NN), probabilistic neural networks (PNN), and C4.5 decision tree, that use the same sets of data and molecular descriptors. Our study indicates the potential of SVM in facilitating the prediction of P-gp substrates. PMID- 15272859 TI - Prediction of chemical carcinogenicity from molecular structure. AB - Carcinogens represent a serious threat to human health. In vivo determination of carcinogenicity is time-consuming and expensive, thus in silico models to predict chemical carcinogenicity are highly desirable for virtual screening of compound libraries of both pharmaceutically and other commercially interesting molecules. In the present study, a PLS-DA (partial least squares discriminant analysis) model was developed to predict carcinogenicities in each of four rodent models: male mouse (MM), female mouse (FM), male rat (MR), and female rat (FR). The data set that was used contained over 520 compounds from both the NTP and the FDA databases. All the models were built from the same molecular descriptor system, which is based on atom typing [Sun, H. J. Chem. Inf. Comput. Sci. 2004, 44, 748 757], enabling the comparison of atomic contributions to carcinogenicity with respect to species and gender. Using four components, the models were able to achieve excellent fitting and prediction, with r(2) = 0.987 and q(2) = 0.944 for MM, r(2) = 0.985 and q(2) = 0.950 for FM, r(2) = 0.989 and q(2) = 0.962 for MR, and r(2) = 0.990 and q(2) = 0.965 for FR. The models were further validated by response permutation testing and external validation, and the results indicated that the models were both statistically significant and predictive. Variable influence on projection (VIP) analysis identified the key atom types and fragments that contributed to carcinogenicities and response differences across species and gender. PMID- 15272861 TI - Nitric oxide signalling functions in plant-pathogen interactions. AB - Nitric oxide (NO) is a highly reactive molecule that rapidly diffuses and permeates cell membranes. During the last few years NO has been detected in several plant species, and the increasing number of reports on its function in plants have implicated NO as a key molecular signal that participates in the regulation of several physiological processes; in particular, it has a significant role in plant resistance to pathogens by triggering resistance associated cell death and by contributing to the local and systemic induction of defence genes. NO stimulates signal transduction pathways through protein kinases, cytosolic Ca2+ mobilization and protein modification (i.e. nitrosylation and nitration). In this review we will examine the synthesis of NO, its effects, functions and signalling giving rise to the hypersensitive response and systemic acquired resistance during plant-pathogen interactions. PMID- 15272862 TI - Conserved features of type III secretion. AB - Type III secretion systems (TTSSs) are essential mediators of the interaction of many Gram-negative bacteria with human, animal or plant hosts. Extensive sequence and functional similarities exist between components of TTSS from bacteria as diverse as animal and plant pathogens. Recent crystal structure determinations of TTSS proteins reveal extensive structural homologies and novel structural motifs and provide a basis on which protein interaction networks start to be drawn within the TTSSs, that are consistent with and help rationalize genetic and biochemical data. Such studies, along with electron microscopy, also established common architectural design and function among the TTSSs of plant and mammalian pathogens, as well as between the TTSS injectisome and the flagellum. Recent comparative genomic analysis, bioinformatic genome mining and genome-wide functional screening have revealed an unsuspected number of newly discovered effectors, especially in plant pathogens and uncovered a wider distribution of TTSS in pathogenic, symbiotic and commensal bacteria. Functional proteomics and analysis further reveals common themes in TTSS effector functions across phylogenetic host and pathogen boundaries. Based on advances in TTSS biology, new diagnostics, crop protection and drug development applications, as well as new cell biology research tools are beginning to emerge. PMID- 15272863 TI - Expression and distribution of beta1 integrins in in vitro-induced M cells: implications for Yersinia adhesion to Peyer's patch epithelium. AB - Beta1 integrins are anchored on the basal membrane of enterocytes, but little is known about their localization in M cells, which are the main entry route into the intestinal mucosa for many bacterial pathogens. In particular, it has been suggested that adhesion of enteropathogenic Yersinia to M cells is mediated by interaction of the bacterial protein invasin and apical beta1 integrins. Using a novel in vitro model of M cells, we demonstrate an augmented apical and basolateral targeting of beta1 integrins in M cells associated with increased total alpha chain synthesis. The alpha3 and alpha6 subunits were targeted to the basal pole, but alpha2 subunit was targeted at both poles. No other alpha subunit was found associated with apical beta1 integrins on M cells. Interestingly, Y. enterocolitica still adhered to the apical surface of M cells, despite the fact that alpha2beta1 is not a receptor for invasin. We therefore studied the adhesive properties of invasin-mutant Y. enterocolitica and invasin-expressing Escherichia coli on the apical surface of M cells. We show that it is not invasin, but the product of an as yet unidentified bacterial chromosomal gene, that is involved in the adhesion of Y. enterocolitica to the apical membrane of M cells. PMID- 15272864 TI - Host cell actin polymerization is required for cellular retention of Trypanosoma cruzi and early association with endosomal/lysosomal compartments. AB - One of the hallmarks of Trypanosoma cruzi invasion of non-professional phagocytes is facilitation of the process by host cell actin depolymerization. Host cell entry by invasive T. cruzi trypomastigotes is accomplished by exploiting a cellular wound repair process involving Ca(2+)-regulated lysosome exocytosis (i.e. lysosome-dependent) or by engaging a recently recognized lysosome independent pathway. It was originally postulated that cortical actin microfilaments present a barrier to lysosome-plasma membrane fusion and that transient actin depolymerization enhances T. cruzi entry by increasing access to the plasma membrane for lysosome fusion. Here we demonstrate that cytochalasin D treatment of host cells inhibits early lysosome association with invading T. cruzi trypomastigotes by uncoupling the cell penetration step from lysosome recruitment and/or fusion. These findings provide the first indication that lysosome-dependent T. cruzi entry is initiated by plasma membrane invagination similar to that observed for lysosome-independent entry. Furthermore, prolonged disruption of host cell actin microfilaments results in significant loss of internalized parasites from infected host cells. Thus, the ability of internalized trypomastigotes to remain cell-associated and to fuse with host cell lysosomes is critically dependent upon host cell actin reassembly, revealing an unanticipated role for cellular actin remodelling in the T. cruzi invasion process. PMID- 15272865 TI - Induction of the CD23/nitric oxide pathway in endothelial cells downregulates ICAM-1 expression and decreases cytoadherence of Plasmodium falciparum-infected erythrocytes. AB - Cytoadherence of parasitized red blood cells (PRBCs) to postcapillary venules and cytokine production are clearly involved in the pathogenesis of cerebral malaria. Nitric oxide and TNF-alpha have been proposed as major effector molecules both in protective and physiopathological processes during malaria infections. Nitric oxide production has been shown to be induced by engagement of CD23 antigen. This study aimed to investigate the potential role of the CD23/nitric oxide pathway in the control of the cytoadherence of PRBCs on human endothelial cells. We demonstrate that normal human lung endothelial cells (HLECs) are able to express the low affinity receptor for IgE (Fc in RII/CD23), following cell incubation with interleukin 4 or PRBCs. Ligation of the CD23 antigen by a specific anti-CD23 monoclonal antibody at the cell surface of HLECs was found to induce iNOS mRNA and protein expression, NO release and P. falciparum killing. In addition, the specific CD23-engagement on these cells also induced a significant decrease in ICAM-1 expression, an adhesion molecule implicated in PRBCs cytoadherence. These data not only described for the first time the expression of a CD23 antigen at the cell surface of endothelial cells but also suggest a possible new regulatory mechanisms via the CD23/NO pathway during malaria infection. PMID- 15272866 TI - Simultaneous analysis of host and pathogen interactions during an in vivo infection reveals local induction of host acute phase response proteins, a novel bacterial stress response, and evidence of a host-imposed metal ion limited environment. AB - A fundamental goal in the study of infections is to understand the dynamic interplay between host and pathogen; however, direct in vivo interrogation of this disease process via transcriptional profiling has been lacking. Here we describe the development and application of novel bacterial RNA amplification technology to simultaneously identify key elements of both host and pathogen responses in a murine infection model. On the bacterial side, we found induction of an unusual pattern of stress response genes, a response to host-induced metal ion limitation, and a failure to achieve stationary phase in vivo. On the mammalian side, we observed the surprising induction of several genes encoding acute phase response proteins including hepcidin, haptoglobin, complement C3 and metallothionein 1 at the site of infection, as well as other mediators of innate immunity. Thus, our results reveal host-pathogen cross-talk not predicted by previous in vitro analyses and provide the framework to eavesdrop on a broad array of host-pathogen interactions in vivo. As described here, the comprehensive examination of host-pathogen interactions during an infection is critical to the discovery of novel approaches for intervention not predicted by current models. PMID- 15272867 TI - Non-encapsulated strains reveal novel insights in invasion and survival of Streptococcus suis in epithelial cells. AB - Streptococcus suis is a porcine and human pathogen causing invasive diseases, such as meningitis or septicaemia. Host cell interactions of S. suis have been studied mainly with serotype 2 strains, but multiple capsular serotypes as well as non-typeable strains exist with diverse virulence features. At present, S. suis is considered an extracellular pathogen. However, whether or not it can also invade host cells is a matter of controversial discussions. We have assessed adherence and invasion of S. suis for HEp-2 epithelial cells by comparing 10 serotype 2 strains and four non-typeable (NT) strains. Only the NT strains and a non-encapsulated serotype 2 mutant strain, but none of the serotype 2 strains, adhered strongly and were invasive. Invasion seemed to be affected by environmental signals, as suggested from comparison of strains grown in different media. Further phenotypic and genotypic characterization revealed a high diversity among the different strains. Electron microscopic analysis of invasion of selected invasive NT strains indicated different uptake mechanisms. One strain induced large invaginations comparable to those seen in 'caveolae' mediated uptake, whereas invasion of the other strains was accompanied by formation of filipodia-like membrane protrusions. Invasion of all strains, however, was similarly susceptible to hypertonic sucrose, which inhibits receptor-mediated endocytosis. Irrespective of the uptake pathway, streptococci resided in acidified phago-lysosome like vacuoles. All strains, except one, survived intracellularly as well as extracellular acidic conditions. Survival seemed to be associated with the AdiS protein, an environmentally regulated arginine deiminase of S. suis. Concluding, invasion and survival of NT strains of S. suis in epithelial cells revealed novel evidence that S. suis exhibits a broad variety of virulence-associated features depending on genetic variation and regulation. PMID- 15272868 TI - An early imbalance of interleukin 12 influences the adjuvant effect of mannoproteins of Cryptococcus neoformans. AB - Mannoprotein from Cryptococcus neoformans induces protective response against a lethal challenge with this fungus or with Candida albicans. This phenomenon is largely related to early production of interleukin 12 (IL-12) and induction of T helper 1 response. Our study assesses whether the early absence of this critical cytokine could account for the incomplete activation of cellular response and whether the immune system compensates this imbalance. The results show that the neutralization of early IL-12 enhanced IL-18 production but decreased IFN-gamma secretion and IL-12R expression by splenic CD4 T cells. In contrast, IL-18R was not augmented despite an increase in IL-18 production. The co-stimulatory pathway was partially dysregulated because splenic macrophages showed unmodified B7-2, and a decrease of B7-1 expression. This dysregulation led to incomplete proliferative response of T cells in response to Cryptococcus neoformans and to increased fungal load in the brain 21 days post infection. The inability to dispose early IL-12, forced the immune system to compensate the imbalance and produced a series of long-lasting dysregulations involving the co-stimulatory pathway and T cell activation. PMID- 15272869 TI - Metabolic engineering of ketocarotenoid formation in higher plants. AB - Although higher plants synthesize carotenoids, they do not possess the ability to form ketocarotenoids. In order to generate higher plants capable of synthesizing combinations of ketolated and hydroxylated carotenoids the genes responsible for the carotene 4,4' oxygenase and 3,3' hydroxylase have been transformed into tomato and tobacco. The gene products were produced as a polyprotein. Subsequent cleavage of the polyprotein, targeting of the two enzymes to the plastid and enzyme activities have been shown for both gene products. Metabolite profiling has shown the formation of ketolated carotenoids from beta-carotene and its hydroxylated intermediates in tobacco and tomato leaf. In the nectary tissues of tobacco flowers a quantitative increase (10-fold) as well as compositional changes were evident, including the presence of astaxanthin, canthaxanthin and 4 ketozeaxanthin. Interestingly, in this tissue the newly formed carotenoids resided predominantly as esters. These data are discussed in terms of metabolic engineering of carotenoids and their sequestration in higher plant tissues. PMID- 15272870 TI - Global changes in transcription orchestrate metabolic differentiation during symbiotic nitrogen fixation in Lotus japonicus. AB - Research on legume nodule metabolism has contributed greatly to our knowledge of primary carbon and nitrogen metabolism in plants in general, and in symbiotic nitrogen fixation in particular. However, most previous studies focused on one or a few genes/enzymes involved in selected metabolic pathways in many different legume species. We utilized the tools of transcriptomics and metabolomics to obtain an unprecedented overview of the metabolic differentiation that results from nodule development in the model legume, Lotus japonicus. Using an array of more than 5000 nodule cDNA clones, representing 2500 different genes, we identified approximately 860 genes that were more highly expressed in nodules than in roots. One-third of these are involved in metabolism and transport, and over 100 encode proteins that are likely to be involved in signalling, or regulation of gene expression at the transcriptional or post-transcriptional level. Several metabolic pathways appeared to be co-ordinately upregulated in nodules, including glycolysis, CO(2) fixation, amino acid biosynthesis, and purine, haem, and redox metabolism. Insight into the physiological conditions that prevail within nodules was obtained from specific sets of induced genes. In addition to the expected signs of hypoxia, numerous indications were obtained that nodule cells also experience P-limitation and osmotic stress. Several potential regulators of these stress responses were identified. Metabolite profiling by gas chromatography coupled to mass spectrometry revealed a distinct metabolic phenotype for nodules that reflected the global changes in metabolism inferred from transcriptome analysis. PMID- 15272871 TI - Functional interactions between a glutamine synthetase promoter and MYB proteins. AB - In Scots pine (Pinus sylvestris), ammonium assimilation is catalysed by glutamine synthetase (GS) [EC 6.3.1.2], which is encoded by two genes, PsGS1a and PsGS1b. PsGS1b is expressed in the vascular tissue throughout the plant body, where it is believed to play a role in recycling ammonium released by various facets of metabolism. The mechanisms that may underpin the transcriptional regulation of PsGS1b were explored. The PsGS1b promoter contains a region that is enriched in previously characterized cis-acting elements, known as AC elements. Pine nuclear proteins bound these AC element-rich regions in a tissue-specific manner. As previous experiments had shown that R2R3-MYB transcription factors could interact with AC elements, the capacity of the AC elements in the PsGS1b promoter to interact with MYB proteins was examined. Two MYB proteins from loblolly pine (Pinus taeda), PtMYB1 and PtMYB4, bound to the PsGS1b promoter were able to activate transcription from this promoter in yeast, arabidopsis and pine cells. Immunolocalization experiments revealed that the two MYB proteins were most abundant in cells previously shown to accumulate PsGS1b transcripts. Immunoprecipitation analysis and supershift electrophoretic mobility shift assays implicated these same two proteins in the formation of complexes between pine nuclear extracts and the PsGS1b promoter. Given that these MYB proteins were previously shown to have the capacity to activate gene expression related to lignin biosynthesis, we hypothesize that they may function to co-regulate lignification, a process that places significant demands on nitrogen recycling, and GS, the major enzyme involved in the nitrogen recycling pathway. PMID- 15272872 TI - Isolation and identification of phosphatidic acid targets from plants. AB - Phosphatidic acid (PA) is emerging as an important lipid signalling molecule. In plants, it is implicated in various stress-signalling pathways and is formed in response to wounding, osmotic stress, cold stress, pathogen elicitors, Nod factors, ethylene and abscisic acid. How PA exerts its effects is still unknown, mainly because of the lack of characterized PA targets. In an approach to isolate such targets we have used PA-affinity chromatography. Several PA-binding proteins were present in the soluble fraction of tomato and Arabidopsis cells. Using mass spectrometric analysis, several of these proteins, including Hsp90, 14-3-3 proteins, an SnRK2 serine/threonine protein kinase and the PP2A regulatory subunit RCN1 could be identified. As an example, the binding of one major PA binding protein, phosphoenolpyruvate carboxylase (PEPC), was characterized further. Competition experiments with different phospholipids confirmed specificity for PA. Hypo-osmotic treatment of the cells increased the amount of PEPC that bound the PA beads without increasing the absolute amount of PEPC. This suggests that PEPC's affinity for PA had increased. The work shows that PA affinity chromatography/mass spectrometry is an effective way to isolate and identify PA-binding proteins from plants. PMID- 15272873 TI - Transcriptional profiling by cDNA-AFLP and microarray analysis reveals novel insights into the early response to ethylene in Arabidopsis. AB - A comprehensive transcriptome analysis by means of cDNA-amplified fragment length polymorphism (AFLP) and cDNA-microarray technology was performed in order to gain further understanding of the molecular mechanisms of immediate transcriptional response to ethylene. Col-0 plants were treated with exogenous ethylene and sampled at six different time-points ranging from 10 min until 6 h. In order to isolate truly ethylene-responsive genes, both the ethylene-insensitive mutant ein2-1 and the constitutive mutant (ctr1-1) were analysed in parallel by cDNA AFLP while ein2-1 was included for the microarray experiment. Out of the cDNA transcript profiling covering about 5% of the Arabidopsis transcriptome, 46 ethylene-responsive genes were isolated, falling in different classes of expression pattern and including a number of novel genes. Out of the 6008 genes present on the chip, 214 genes were significantly (alpha = 0.001) differentially expressed between Col-0 and ein2-1 over time. Cluster analysis and functional grouping of co-regulated genes allowed to determine the major ethylene-regulated classes of genes. In particular, a large number of genes involved in cell rescue, disease and defence mechanisms were identified as early ethylene-regulated genes. Furthermore, the data provide insight into the role of protein degradation in ethylene signalling and ethylene-regulated transcription and protein fate. Novel interactions between ethylene response and responses to several other signals have been identified by this study. Of particular interest is the overlap between ethylene response and responses to abscisic acid, sugar and auxin. In conclusion, the data provide unique insight into early regulatory steps of ethylene response. PMID- 15272874 TI - Metabolism of avenanthramide phytoalexins in oats. AB - Oat leaves produce phytoalexins, avenanthramides, in response to infection by pathogens or treatment with elicitors. The metabolism of avenanthramides was investigated using low molecular weight, partially deacetylated chitin as an elicitor. When oat leaf segments are floated on the elicitor solution, avenanthramides accumulate in the solution. The transfer of elicited oat leaves to solutions containing stable-isotope-labeled avenanthramides resulted in a rapid decrease in the labeled avenanthramides, suggesting the metabolism of avenanthramides. The rate of decrease was enhanced by elicitor treatment, and was dependent on the species of avenanthramides, with avenanthramide B decreasing most rapidly. The rates of biosynthesis and metabolism of avenanthramides A and B were measured using a model of isotope-labeling dynamics. Avenanthramide B was found to be more actively biosynthesized and metabolized than avenanthramide A. Radiolabeled avenanthramide B was incorporated into the cell wall fraction and 99% of incorporated activity was released by alkaline treatment. Gel filtration indicated that high-molecular-weight compounds derived from avenanthramide B were released by alkaline treatment. The decrease in stable-isotope-labeled avenanthramides was suppressed by catalase, salicylhydroxamic acid, and sodium ascorbate, suggesting the involvement of peroxidase in the metabolism. Consistent with this, peroxidase activity that accepts avenanthramide B as a substrate was induced in apoplastic fractions by elicitor treatment. The appearance of multiple basic isoperoxidases was observed by activity staining with 3-amino-9 ethylcarbazole coupled with isoelectric focusing of proteins from elicitor treated leaves. These findings suggest that accumulated avenanthramides are further metabolized in apoplasts in oat leaves by inducible isoperoxidases. PMID- 15272875 TI - The S haplotype-specific F-box protein gene, SFB, is defective in self-compatible haplotypes of Prunus avium and P. mume. AB - Many Prunus species, including sweet cherry and Japanese apricot, of the Rosaceae, display an S-RNase-based gametophytic self-incompatibility (GSI). The specificity of this outcrossing mechanism is determined by a minimum of two genes that are located in a multigene complex, termed the S locus, which controls the pistil and pollen specificities. SFB, a gene located in the S locus region, encodes an F-box protein that has appropriate S haplotype-specific variation to be the pollen determinant in the self-incompatibility reaction. This study characterizes SFBs of two self-compatible (SC) haplotypes, S(4') and S(f), of Prunus. S(4') of sweet cherry is a pollen-part mutant (PPM) that was produced by X-ray irradiation, while S(f) of Japanese apricot is a naturally occurring SC haplotype that is considered to be a PPM. DNA sequence analysis revealed defects in both SFB(4') and SFB(f). A 4 bp deletion upstream from the HVa coding region of SFB(4') causes a frame-shift that produces transcripts of a defective SFB lacking the two hypervariable regions, HVa and HVb. Similarly, the presence of a 6.8 kbp insertion in the middle of the SFB(f) coding region leads to transcripts for a defective SFB lacking the C-terminal half that contains HVa and HVb. As all reported SFBs of functional S haplotypes encode intact SFB, the fact that the partial loss-of-function mutations in SFB are present in SC mutant haplotypes of Prunus provides additional evidence that SFB is the pollen S gene in GSI in Prunus. PMID- 15272876 TI - From pollen tubes to infection threads: recruitment of Medicago floral pectic genes for symbiosis. AB - While the biology of nitrogen-fixing root nodules has been extensively studied, little is known about the evolutionary events that predisposed legume plants to form symbiosis with rhizobia. We have studied the presence and the expression of two pectic gene families in Medicago, polygalacturonases (PGs) and pectin methyl esterases (PMEs) during the early steps of the Sinorhizobium meliloti-Medicago interaction and compared them with related pollen-specific genes. First, we have compared the expression of MsPG3, a PG gene specifically expressed during the symbiotic interaction, with the expression of MsPG11, a highly homologous pollen specific gene, using promoter-gus fusions in transgenic M. truncatula and tobacco plants. These results demonstrated that the symbiotic promoter functions as a pollen-specific promoter in the non-legume host. Second, we have identified the presence of a gene family of at least eight differentially expressed PMEs in Medicago. One subfamily is represented by one symbiotic gene (MtPER) and two pollen-expressed genes (MtPEF1 and MtPEF2) that are clustered in the M. truncatula genome. The promoter-gus studies presented in this work and the homology between plant PGs, together with the analysis of the PME locus structure and MtPER expression studies, suggest that the symbiotic MsPG3 and MtPER could have as ancestors pollen-expressed genes involved in polar tip growth processes during pollen tube elongation. Moreover, they could have been recruited after gene duplication in the symbiotic interaction to facilitate polar tip growth during infection thread formation. PMID- 15272877 TI - Antisense downregulation of the barley limit dextrinase inhibitor modulates starch granule size distribution, starch composition and amylopectin structure. AB - The barley protein limit dextrinase inhibitor (LDI), structurally related to the alpha-amylase/trypsin inhibitor family, is an inhibitor of the starch debranching enzyme limit dextrinase (LD). In order to investigate the function of LDI, and the consequences for starch metabolism of reduced LDI activity, transgenic barley plants designed to downregulate LDI by antisense were generated. Homozygous antisense lines with reduced LDI protein level and activity were analysed and found to have enhanced free LD activity in both developing and germinating grains. In addition the antisense lines showed unpredicted pleiotropic effects on numerous enzyme activities, for example, alpha- and beta-amylases and starch synthases. Analysis of the starch showed much reduced numbers of the small B-type starch granules, as well as reduced amylose relative to amylopectin levels and reduced total starch. The chain length distribution of the amylopectin was modified with less of the longer chains (>25 units) and enhanced number of medium chains (10-15 units). These results suggest an important role for LDI and LD during starch synthesis as well as during starch breakdown. PMID- 15272878 TI - Molecular events in senescing Arabidopsis leaves. AB - Senescence is the final stage of leaf development. Although it means the loss of vitality of leaf tissue, leaf senescence is tightly controlled by the development to increase the fitness of the whole plant. The molecular mechanisms regulating the induction and progression of leaf senescence are complex. We used a cDNA microarray, containing 11 500 Arabidopsis DNA elements, and the whole-genome Arabidopsis ATH1 Genome Array to examine global gene expression in dark-induced leaf senescence. By monitoring the gene expression patterns at carefully chosen time points, with three biological replicates each time, we identified thousands of up- or down-regulated genes involved in dark-induced senescence. These genes were clustered and categorized according to their expression patterns and responsiveness to dark treatment. Genes with different expression kinetics were classified according to different biological processes. Genes showing significant alteration of expression patterns in all available biochemical pathways were plotted to envision the molecular events occurring in the processes examined. With the expression data, we postulated an innovative biochemical pathway involving pyruvate orthophosphate dikinase in generating asparagine for nitrogen remobilization in dark-treated leaves. We also surveyed the alteration in expression of Arabidopsis transcription factor genes and established an apparent association of GRAS, bZIP, WRKY, NAC, and C2H2 transcription factor families with leaf senescence. PMID- 15272879 TI - Phosphate transport in Arabidopsis: Pht1;1 and Pht1;4 play a major role in phosphate acquisition from both low- and high-phosphate environments. AB - Of the mineral nutrients essential for plant growth, phosphorus plays the widest diversity of roles and a lack of phosphorus has profound effects on cellular metabolism. At least eight members of the Arabidopsis Pht1 phosphate (Pi) transporter family are expressed in roots and Pht1;1 and Pht1;4 show the highest transcript levels. The spatial and temporal expression patterns of these two genes show extensive overlap. To elucidate the in planta roles of Pht1;1 and Pht1;4, we identified loss-of-function mutants and also created a double mutant, lacking both Pht1;1 and Pht1;4. Consistent with their spatial expression patterns, membrane location and designation as high-affinity Pi transporters, Pht1;1 and Pht1;4 contribute to Pi transport in roots during growth under low-Pi conditions. In addition, during growth under high-Pi conditions, the double mutant shows a 75% reduction in Pi uptake capacity relative to wildtype. Thus, Pht1;1 and Pht1;4 play significant roles in Pi acquisition from both low- and high-Pi environments. PMID- 15272880 TI - Antisense phenotypes reveal a role for SHY, a pollen-specific leucine-rich repeat protein, in pollen tube growth. AB - SHY, a pollen-specific gene identified in a screen for genes upregulated at pollen germination, encodes a leucine-rich repeat (LRR) protein that is predicted to be secreted. To test if SHY plays an important role during pollen germination, we generated transgenic plants expressing an antisense (AS) copy of the SHY cDNA in pollen. Primary transformants exhibited poor seed set, but homozygous lines could be identified. In these lines, nearly all pollen tubes failed to reach the ovules; tube growth was arrested at the apex of the ovary and the pollen tubes exhibited abnormal callose deposits throughout the tube and in the tips. We show that a SHY::eGFP fusion protein is targeted to the cell wall. The structure of the SHY protein is nearly identical to other extracellular matrix glycoproteins that are composed of LRRs, such as the polygalacturonase inhibitor proteins (PGIP) of plants. PGIPs may function as defense proteins by inhibiting fungal endo-polygalacturonases, but enzyme assays with extracts of AS-SHY pollen do not support such an inhibitor role for SHY. The tomato ortholog of SHY interacts with a tomato receptor kinase (LePRK2) in yeast two-hybrid and pull-down assays; this, and the AS-SHY phenotypes, suggest instead that SHY might function in a signal transduction pathway mediating pollen tube growth. PMID- 15272881 TI - Stromule formation is dependent upon plastid size, plastid differentiation status and the density of plastids within the cell. AB - Stromules are motile extensions of the plastid envelope membrane, whose roles are not fully understood. They are present on all plastid types but are more common and extensive on non-green plastids that are sparsely distributed within the cell. During tomato fruit ripening, chloroplasts in the mesocarp tissue differentiate into chromoplasts and undergo major shifts in morphology. In order to understand what factors regulate stromule formation, we analysed stromule biogenesis in tobacco hypocotyls and in two distinct plastid populations in tomato mesocarp. We show that increases in stromule length and frequency are correlated with chromoplast differentiation, but only in one plastid population where the plastids are larger and less numerous. We used tobacco hypocotyls to confirm that stromule length increases as plastids become further apart, suggesting that stromules optimize the plastid-cytoplasm contact area. Furthermore, we demonstrate that ectopic chloroplast components decrease stromule formation on tomato fruit chromoplasts, whereas preventing chloroplast development leads to increased numbers of stromules. Inhibition of fruit ripening has a dramatic impact on plastid and stromule morphology, underlining that plastid differentiation status, and not cell type, is a significant factor in determining the extent of plastid stromules. By modifying the plastid surface area, we propose that stromules enhance the specific metabolic activities of plastids. PMID- 15272882 TI - Kinetics of labelling of organic and amino acids in potato tubers by gas chromatography-mass spectrometry following incubation in (13)C labelled isotopes. AB - Metabolic pathways of primary metabolism of discs isolated from potato tubers were evaluated by the use of a gas chromatography-mass spectrometry (GC-MS) method generated specifically for this purpose. After testing several possible methods including chemical ionization, it was decided for reasons of sensitivity, reproducibility and speed to use electron impact ionization-based GC-MS analysis. The specific labelling and label accumulation of over 30 metabolites including a broad number of sugars, organic and amino acids was analysed following the incubation of tuber discs in [U-(13)C]glucose. The reproducibility of this method was similar to that found for other GC-MS-based analyses and comparison of flux estimates from this method with those obtained from parallel, yet less comprehensive, radiolabel experiments revealed close agreement. Therefore, the novel method allows quantitatively evaluation of a broad range of metabolic pathways without the need for laborious (and potentially inaccurate), chemical fractionation procedures commonly used in the estimation of fluxes following incubation in radiolabelled substrates. As a first experiment the GC-MS method has been applied to compare the metabolism of wild type and well-characterized transgenic potato tubers exhibiting an enhanced sucrose mobilization. The fact that this method is able to rapidly yield further comprehensive information into primary metabolism illustrates its power as a further phenotyping tool for the analysis of plant metabolism. PMID- 15272883 TI - A delay they can ill afford: delays in obtaining Attendance Allowance for older, terminally ill cancer patients, and the role of health and social care professionals in reducing them. AB - Despite a growing interest in holistic care for the terminally ill, financial needs are often not addressed. This is reflected in the fact that some people with a terminal illness are not accessing disability benefits, despite eligibility. The present paper is based on a study investigating delays experienced by cancer patients in obtaining Attendance Allowance (AA) by special rules, and missed opportunities for professionals to assist with claims. The study took place in a hospice where patients were referred to social work professionals for assistance in claiming AA. In each case, the patient had been eligible for some time before the referral. Over a 5-month period, all 22 patients who were referred completed a questionnaire. Data were collected to show their personal characteristics, how they came to be referred for assistance and their level of knowledge of AA. The length of time that people had already been eligible and the time taken to claim were recorded to show the amount of lost benefit. The health and social care professionals whom these patients had seen since becoming eligible were also recorded. A wide range of people experienced delays in accessing AA. Their total lost income ranged from pound 110.60 to pound 1106.00. The median was pound 387.10 and four people died before being awarded AA. Only four patients were fully aware of their eligibility. Every person had seen between one and four professionals since becoming eligible for the benefit, without the meeting resulting in a claim. Increased income aids the management of illness, and information and assistance to claim disability benefits need to be made available in a consistent manner at the earliest opportunity. Health and social care professionals are in a position to provide this. However, changes to the claims process, proposed by the present author, could ensure that AA is received automatically, without delay and without extensive paperwork. PMID- 15272884 TI - An exploration of nutrition and eating disabilities in relation to quality of life at 6 months post-stroke. AB - Quality of life (QoL) is increasingly recognised as an important healthcare outcome, especially for those living with enduring disability. Stroke is a major source of long-term disablement and many aspects of life after stroke have been explored. Little attention has been paid to nutritional issues despite the cultural and hedonistic importance of food and eating, and the deleterious effects of malnutrition. The present study employed an epidemiological survey to investigate the contribution of dietary and nutritional factors in relation to QoL after stroke. The participants were 206 survivors of a cohort of acute stroke patients consecutively admitted to a National Health Service trust hospital in South London, UK, between March 1998 and April 1999. They were interviewed in their homes at 6 months post-stroke. Cognitively or communication-impaired patients were precluded from interview except where a live-in carer participated as a proxy (n = 10). The participation rate for those who were eligible and could be contacted was 206 out of 218 (94%). Participants were assessed using standardised, validated tools for functional abilities in activities of daily living and eating, cognition and mood state, social support and economic indices, nutritional status, dietary intake, and QoL. Overall group scores demonstrated relatively minor degrees of physical disablement; exclusion of those with limited cognition and communication precluded assessment of a small subgroup with greater disablement at hospital discharge. Nonetheless, the overall assessment results were not dissimilar to other reported groups. Indices of poor nutritional status and substantial dietary inadequacy were revealed, linked with reduced appetite and depression. Multiple regression analyses revealed the dominant impact of mood state in relation to QoL scores; additional significant effects were identified for social support, eating-related disabilities and age. The effects of mood and social support are well-recognised, whilst nutrition-related effects have previously received little attention. Intervention in these areas might achieve improvements in survivors' perceived QoL. PMID- 15272885 TI - Involving people with learning disabilities in research: issues and possibilities. AB - Advances in the social position of people with learning disabilities have led to a situation where research and evaluation studies are increasingly required to include the views and opinions of people with learning disabilities. One key outcome of this shift is that some of the major funding bodies now insist on the inclusion of people with learning disabilities as a condition of research funding. This has produced new possibilities and new challenges for researchers, and it has real consequences for people working in health and social care. The present paper sets out to explore some of the developments and challenges in research with people with learning disabilities. The author provides a selective overview of developments with the aim of demonstrating the richness, ingenuity and potential of research involving people with learning disabilities. The paper is divided into three broad sections that focus on: (1) the ethics and philosophy of participatory research; (2) the methodologies employed at particular points in the research process that are designed to ensure the involvement of participants in research; and (3) building capacity in participatory research as a precondition to the further development of this approach. An investment in capacity would enable this approach to move into the mainstream of research activity involving people with learning disabilities. PMID- 15272886 TI - Conceptualising successful partnerships. AB - Partnership working has become a central feature of British social welfare policy since 1997. Although this development is applicable to all areas of public welfare, nowhere is it more evident than in the planning and provision of care that overlaps health and social services. The literature survey described in the present paper focused on research examining the impact of partnership working in these areas to assess the evidence concerning its effects and to investigate how partnership 'success' is conceptualised. The literature conceptualised the success of partnerships in two main ways: (1) process issues, such as how well the partners work together in addressing joint aims and the long-term sustainability of the partnership; and (2) outcome issues, including changes in service delivery, and subsequent effects on the health or well-being of service users. The authors found that research into partnerships has centred heavily on process issues, while much less emphasis has been given to outcome success. If social welfare policy is to be more concerned with improving service delivery and user outcomes than with the internal mechanics of administrative structures and decision-making, this is a knowledge gap that urgently needs to be filled. PMID- 15272887 TI - Is client-centred care planning for chronic disease sustainable? Experience from rural South Australia. AB - This qualitative evaluation of a chronic disease self-management project in rural South Australia considers the sustainability of client-centred care planning under current organisational and funding arrangements. The study involved consultation with a range of five stakeholder types over two stages (40 in the beginning stage and 39 in the middle stage) about their satisfaction with the care planning and self-management approach used in the project. All stakeholder types valued the client-centred approach because they perceived that clients were better able to accept and deal with the long-term management of their condition. However, this required that care planning should deal with a wider range of issues than just medical management, and so it took longer, which raised its sustainability in general practice under the current funding through the national health insurance programme (Medicare). The study concludes that sustainability may be addressed through further research into the role of and funding for peer led self-management groups and the employment of care planners in organisational settings that are conducive to a client-centred approach. PMID- 15272888 TI - A family support service for homeless children and parents: users' perspectives and characteristics. AB - The objective of the present study was to establish the psychosocial characteristics and perspectives of 49 consecutive homeless families who received input from a new designated family support worker (FSW) post at a large statutory hostel for homeless parents and children. The FSW provided: assessment of social, educational and health needs; support and parent training; and liaison with and referral to specialist services. Measures included quantitative questionnaires (i.e. the Hospital Anxiety and Depression Scale, the Parenting Daily Hassles Scale, the Eyberg Child Behaviour Inventory, and the Health of the Nation Outcome Scales for Children and Adolescents), and a qualitative (semistructured) interview on service experiences and satisfaction. The psychosocial measures indicated high rates of parenting difficulties, mental health and related needs among children and their parents. Parenting difficulties were associated with child behaviour problems. Parents expressed satisfaction with the service whilst they were residents at the hostel, but they were often not clear about the objectives of agencies and interventions. Family support interventions have a key role in service provision for homeless and other vulnerable families by providing direct parenting interventions and ensuring that specialist agencies are appropriately involved. Family support worker involvement needs to continue when families are re-housed in the community. PMID- 15272889 TI - Primary care mental health workers: role expectations, conflict and ambiguity. AB - A number of professionals are involved in mental health in primary care. The NHS Plan proposed the introduction of a new professional, the primary care mental health worker (PCMHW), to improve care in this setting. The present study was conducted to examine pilot PCMHW-type roles currently in existence, to explore staff expectations concerning the new PCMHW role and to consider the issues relating to roles in primary care mental health that are raised by this new worker. The study used a case study design, and involved qualitative interviews with 46 managers and clinicians from primary care and specialist mental health services, including pilot PCMHW-type roles. The key findings were as follows: The pilot PCMHW-type roles were almost exclusively related to client work, whereas respondents had far wider role expectations of the new PCMHWs, relating to perceived gaps in current service provision. This highlights the potential for role conflict. Secondly, there was disagreement and ambiguity among some respondents as to the nature of the new PCMHW's role in client work, and its relationship with the work undertaken by other mental health professionals such as counsellors, psychologists and nurses. Given that multiple professionals are involved in mental health care in primary care, issues relating to roles are likely to be crucial in the effective implementation of the new PCMHWs. PMID- 15272890 TI - Collaborative learning for collaborative working? Initial findings from a longitudinal study of health and social care students. AB - This paper presents the initial findings from a longitudinal quantitative study of two cohorts of students who entered the 10 pre-qualifying programmes of the Faculty of Health and Social Care, University of the West of England (UWE), Bristol, UK. The overall aim of the study is to explore students' attitudes to collaborative learning and collaborative working, both before and after qualification. On entry to the faculty, 852 students from all 10 programmes completed the UWE Entry Level Interprofessional Questionnaire, which gathered baseline data concerning their self-assessment of communication and teamwork skills, and their attitudes towards interprofessional learning and interprofessional interaction. Comparative analysis of these data was undertaken in terms of demographic variables such as age (i.e. older or younger than 21 years), experience of higher education, prior work experience and choice of professional programme. The results indicate that most students rated their communication and teamwork skills positively, and were favourably inclined towards interprofessional learning, but held negative opinions about interprofessional interaction. Some student groups differed in their responses to some sections of the questionnaire. Mature students, and those with experience of higher education or of working in health or social care settings, displayed relatively negative opinions about interprofessional interaction; social work and occupational therapy students were particularly negative in their responses, even after adjustment for confounding demographic variables. The paper concludes by considering the implications of the findings for interprofessional educational initiatives and for professional practice. PMID- 15272891 TI - 'How can they tell?' A qualitative study of the views of younger people about their dementia and dementia care services. AB - There is growing interest in eliciting the views of younger people with dementia (i.e. those under 65 years of age) within health and social care research. The often erroneous view that these individuals are not capable of expressing their views and experiences has now been seriously challenged. The present paper draws on the findings from 14 qualitative in-depth interviews with younger people with dementia conducted in the South-west of England, and considers some of the issues involved in interviewing people with dementia. Purposive and snowballing techniques were used to recruit participants. Data were transcribed and subjected to comparative textual analysis to index, code and analyse the data for emergent themes. Four major themes emerged: (1) the general experience of having dementia; (2) dementia diagnosis; (3) the importance of age; and (4) risk and danger issues. The results indicate that the majority of participants were articulate and insightful about their experiences and needs. The present paper concludes by arguing that the challenge for health and social care professionals is to engage with and consult such individuals about their experiences and what they want from dementia care services, and the authors consider some of the issues involved in interviewing people with dementia. PMID- 15272893 TI - Joubert syndrome: review and report of seven new cases. AB - Joubert syndrome (JS) is an autosomal-recessive disorder, characterized by hypotonia, ataxia, global developmental delay and molar tooth sign on magnetic resonance imaging. A variety of other abnormalities have been described in children with JS, including abnormal breathing, abnormal eye movements, a characteristic facial appearance, delayed language, hypersensitivity to noise, autism, ocular and oculomotor abnormalities, meningoencephaloceles, microcephaly, low-set ears, polydactyly, retinal dysplasia, kidney abnormalities (renal cysts), soft tissue tumor of the tongue, liver disease and duodenal atresia. Even within siblings the phenotype may vary, making it difficult to establish the exact clinical diagnostic boundaries of JS. We review the clinical characteristics of seven cases that fulfill the criteria of JS. PMID- 15272894 TI - Mortality from multiple sclerosis in Austria 1970-2001: dynamics, trends, and prospects. AB - A divergence in earlier multiple sclerosis (MS) mortality rates observed within Europe, prompted us to determine the MS mortality rate in Austria and several European countries. Our aim was to examine the temporal and geographical variations within Austria and to determine future MS mortality rates based on a projection model. MS mortality data set, differentiated by age groups, sex, and region at death for the period 1970-2001 were obtained. Prognostic MS mortality trends for the period 2002-2020 were estimated using the simultaneous multiple cause-delay (SIMCAD) method. Our findings indicate a decline (47%) in the MS mortality rate from 1.41 (1970-79) to 0.96 (1980-89) and 0.70 (1990-2001) per 100 000 in Austria during the 32-year period observed. Conversely, the scenarios of our projection for the period 2002-2020, reveal an increasing MS mortality rate. The median age at death because of MS increased with 0.7 years for men and 2.9 years for women during the observed period (1970-2001). Austria, like many other European countries, has experienced a decreasing MS mortality rate over the last three decades. An increased MS mortality rate is however expected over the next decades in Austria. This increase will be most prominent in the elderly population cohorts because of demographic shifts. PMID- 15272895 TI - A novel mutation (Thr116Ile) in the presenilin 1 gene in a patient with early onset Alzheimer's disease. AB - We report a novel presenilin 1 (PSN1) mutation (Thr116Ile) in a woman with early onset Alzheimer's disease (AD). This mutation was not found in 100 healthy controls, indicating that this is not a common polymorphism. The patient presented with forgetfulness at age 45, followed over the next 3 years by a worsening of the memory loss and frequent episodes of confusion and spatial disorientation. Neuroimaging studies were consistent with AD. The analysis of the family's pedigree showed that the proband was apparently the only member affected. Because the early death of several close relatives (i.e. the mother and the grandmother) and the demonstration that the father is not a mutation carrier, it is suggested that either a de novo mutation or a censor effect might have occurred. Our finding supports the indication that PSN1 mutations should be searched for in early-onset AD, particularly when a censor effect precludes a precise genetic analysis. PMID- 15272896 TI - CSF isoelectrofocusing in a large cohort of MS and other neurological diseases. AB - The present study was performed in order to confirm the diagnostic value of isoelectrofocusing (IEF) in a large multiple sclerosis (MS) cohort and to evaluate the various neurological diseases probably to present a similar IEF profile. The cerebrospinal fluid (CSF) of 1292 patients with neurological diseases was studied by IEF. After a follow-up of 2-36 months, we only included patients with a definite MS or confirmed diagnosis of other neurological diseases (OND). MS was diagnosed in 407 patients and OND in 593 patients. For patients in whom three or more oligoclonal bands (OCB) were detected, IEF results showed a sensitivity of 85% and a specificity of 92% for the diagnosis of MS. The positive and negative predictive values were 86.5 and 90%, respectively. Inflammatory and infectious disorders of the central nervous system represented the main affections associated with OCB, including human immunodeficiency virus encephalitis, Lyme disease and less frequently Sjogren syndrome. Furthermore, when OCB were observed, 10 or more bands were more frequently found in MS than in OND (P < 0.0001). IEF of the CSF is a reliable method for the diagnosis of MS. The absolute number of bands may help to discriminate between MS and OND. PMID- 15272897 TI - The screening for X-linked Emery-Dreifuss muscular dystrophy amongst young patients with idiopathic heart conduction system disease treated by a pacemaker implant. AB - The X-linked Emery-Dreifuss muscular dystrophy (X-EDMD) is a hereditary muscle disorder associated with cardiac involvement. Sinus node dysfunction and atrioventricular conduction defects, typical of X-EDMD, occur in both males and females and may result in sudden cardiac death unless treated by permanent pacing. The objective of the study was to determine the frequency and relevance of X-EDMD in heart conduction system disease in young individuals treated with a pacemaker implant. The medical history of 3450 paced individuals in the region of South Moravia, Czech republic, was reviewed. Thirty-five patients, 20 males and 15 females, with idiopathic heart conduction disease of onset before age 40 were identified and screened for X-EDMD. Within these 35 individuals, only one male was found to carry a mutation in X-EDMD gene. We conclude that the clinical relevance of X-EDMD in heart conduction system disease is very low. It should, however, be included into the diagnostic work-up of young male individuals with idiopathic cardiac conduction disturbances. PMID- 15272898 TI - Excessive daytime somnolence in Japanese patients with Parkinson's disease. AB - To investigate the prevalence and severity of excessive daytime somnolence (EDS) in Japanese patients with Parkinson's disease (PD) and to examine the main cause of EDS. Fifty-three Japanese patients with PD (PDs: 32 females and 21 males) and 17 controls (10 females and seven males) were evaluated using the Epworth Sleepiness Scale (ESS). The severity of the disease was evaluated by Unified Parkinson's disease Rating Scale (UPDRS), and information about quality and quantity of medications was collected. The correlations amongst EDS and age, severity of PD, duration of illness and medications were analyzed. The mean ESS score was significantly higher in advanced PDs than in controls, and correlated with the UPDRS score (r(s) = 0.743, P < 0.0001). Age, duration of illness and the dose of levodopa weakly correlated with ESS score. The intake of dopamine agonists did not affect the severity of EDS. The mean ESS score in PDs was lower than that reported in PD in European and American studies. EDS in Japanese patients with PD was milder compared with Caucasian patients, which might be due to the lower doses of the medications used in Japan. The results suggest that EDS in PD is mainly because of neuropathological changes of the disease itself. PMID- 15272899 TI - Prednisolone therapy in Duchenne muscular dystrophy prolongs ambulation and prevents scoliosis. AB - Steroids may have a beneficial effect on the course of Duchenne muscular dystrophy (DMD). However, results vary in different studies. This study consisted of 66 DMD boys who were in the therapy group and 22 DMD boys in the control group. The mean ages were 6.8 +/- 2.1 years (range 2.5-12.5) and 7.0 +/- 1.3 years (range 5.0-9.0), respectively. We assessed muscle strength, 10-m walking, ankle contracture, and loss of independent walking ability age and onset of scoliosis. Treatment regimen was oral prednisolone 0.75 mg/kg on alternate days, plus vitamin D 600-1200 units/day and a calcium-enriched diet. After a follow-up period of 2.75 +/- 1.1 years (range 1.5-5) and when compared with controls, there was a statistically significant change in muscle strength between the two groups after 12 months (P < 0.05). Although 10-m walking time decreased in therapy group (P < 0.05), there was not significance between the groups in the end. Boys in the control group developed significantly less ankle contractures (P < 0.05). None of the therapy group had scoliosis during the follow-up period (mean age 10.8 +/- 1.2 years), whereas seven boys of the control group had scoliosis at a mean age of 11.7 +/- 2 years. Loss of walking ability age was statistically different between groups (P < 0.05). Our results indicate that, alternate-day prednisolone regimen may prolong ambulation and scoliosis can be delayed or prevented. PMID- 15272900 TI - Therapeutic outcome in neuropathic pain: relationship to evidence of nervous system lesion. AB - Treatment outcome in patients with neuropathic pain (NP) is often variable and disappointing. We tested the hypothesis that patients with clear evidence of nervous system lesion respond better to pharmacological treatment with documented effect on NP than patients with poor or no evidence of nervous system lesion. Furthermore, we examined whether specific symptoms or signs were associated with treatment outcome. A total of 214 patients with suspected non-cancer NP were divided into four groups with graded evidence of nervous system lesion based on medical history, bedside sensory examination, quantitative sensory tests, electrophysiology, and neuroimaging. Patients were treated with imipramine guided by plasma-drug concentrations. Gabapentin 2400 mg/day was given in case of treatment failure or if imipramine treatment was not possible. Two hundred patients completed the study. Global pain relief was similar in the four groups. There was no association between evidence of nervous system lesion and treatment outcome. Classical NP signs: abnormal temporal summation, cold and brush allodynia, and abnormal sensibility to temperature were also unrelated to outcome. Treatment outcome was similar in peripheral and central definite NP. Neither definite evidence of nervous system lesion nor abnormal sensory phenomena seems to predict for good outcome of therapy with imipramine or gabapentin in patients with suspected neuropathic pain. PMID- 15272901 TI - A negative personal and family history for venous thrombotic events is not sufficient to exclude thrombophilia in patients with cerebral venous thrombosis. AB - A hereditary thrombophilia is found in 20-30% of patients with cerebral venous thrombosis (CVT). These patients might have an increased rate of a positive personal or family history of venous thrombotic events. We investigated the diagnostic value of a structured personal and family history for venous thrombotic events in 56 consecutive cases of CVT. Fourteen of 56 patients (25%) had a hereditary thrombophilia, mostly factor V Leiden. Patients with both CVT and hereditary thrombophilia had more frequently a positive family and personal history than patients affected by CVT only but the difference was not strong enough to differ from the 42 CVT patients without thrombophilia (43% vs. 31%; P = 0.52 and 14% vs. 10 %; P = 0.63). We conclude that a negative personal and family history of venous thrombotic events is not sufficient to exclude thrombophilia and patients with CVT should be tested for inherited thrombophilia regardless of the patient's past personal and family history for venous thrombotic events. PMID- 15272902 TI - Asymptomatic myasthenia gravis influences pregnancy and birth. AB - Women with myasthenia gravis (MG) have an increased risk of complications and adverse pregnancy outcome. This study has examined if this is true also for asymptomatic MG. Using data from the Medical Birth Registry of Norway, births of women prior to receiving an MG diagnosis or in complete clinical MG remission were compared with all non-MG births in Norway in the same period (1967-2000). Forty-nine births occurred in 37 women, 11 of them in clinical remission, and six thymectomized. The perinatal mortality was increased (P = 0.02) and induction of birth (P = 0.007) occurred more frequently. Protracted labor occurred more frequently in the target group (P = 0.03). One of the three children that died had Potter's syndrome. Both mothers with children who died were in complete clinical MG remission. One had previously given and one subsequently gave birth to a child with neonatal MG. The results indicate that complications in birth and pregnancy are not only related to clinical MG disease severity but to the underlying immunological dysfunction. PMID- 15272903 TI - The Cotard syndrome. Report of two patients: with a review of the extended spectrum of 'delire des negations'. AB - The Cotard syndrome is characterized by the delusion where an individual insists that he has died or part of his body has decayed. Although described classically in schizophrenia and bipolar disorder, physical disorders including migraine, tumour and trauma have also been associated with the syndrome. Two new cases are described here, the one associated with arteriovenous malformations and the other with probable multiple sclerosis. The delusion has been embarrassing to each patient. Study of such cases may have wider implications for the understanding of the psychotic interpretation of body image, for example that occurring in anorexia nervosa. PMID- 15272904 TI - Flail arm syndrome: a clinical variant of amyotrophic lateral sclerosis. AB - We describe a case of a 65-year old patient diagnosed with amyotrophic lateral sclerosis. The clinical findings, with symmetric, predominantly proximal wasting and weakness of both arms (especially of the infra-, supraspinatus and deltoideus) leading to severe functional disability and contrasting with preserved independent ambulation and sparing of bulbar muscles, were consistent with the proposed criteria of the so-called flail arm syndrome. Based on our case we characterize the clinical features of flail arm syndrome and review the literature. PMID- 15272905 TI - Postmenopausal HRT is not independent risk factor for dural sinus thrombosis. AB - While a dural sinus thrombosis (DST), is a well-known consequence of the use of oral contraceptives, the role of hormone replacement therapy (HRT) in DST was not previously evaluated. We report two postmenopausal women, presenting with DST under HRT. Antiphospholipid antibodies in one case and borderline protein S deficiency in another were diagnosed. Only five cases of DST under HRT were previously reported and in two of them additional prothrombotic risk factors were found. According to these and previous cases, HRT is not an independent risk factor for DST. PMID- 15272906 TI - Greater occipital neuralgia and arthrosis of C1-2 lateral joint. PMID- 15272910 TI - Uses for recombinant human TSH in patients with thyroid cancer and nodular goiter. AB - Recombinant human TSH (rhTSH) has revolutionized the care of patients with differentiated thyroid cancer. Since its approval for clinical use in 2001 in Europe (1998 in the USA), rhTSH has greatly enhanced the surveillance of these patients by allowing the avoidance of hypothyroidism for TSH stimulation. Previously, a hypothyroid state was required for TSH stimulated diagnostic whole body radio-iodine scans (DxWBS) and thyroglobulin (Tg) levels. Patients generally prefer rhTSH as a mechanism for TSH stimulation because symptoms of hypothyroidism can be completely avoided. Currently, rhTSH is only approved for diagnostic monitoring of differentiated thyroid cancer patients. There are many other potential uses for rhTSH, including facilitation of treatment of patients with thyroid cancer and nodular goiter. The diagnostic and therapeutic role of rhTSH in patients with differentiated thyroid cancer and nodular goiter will be discussed in this review. PMID- 15272911 TI - Aldosterone synthase gene variation and adrenocortical response to sodium status, angiotensin II and ACTH in normal male subjects. AB - OBJECTIVE: Aldosterone synthase, a key enzyme in the terminal steps of aldosterone synthesis, is encoded by the CYP11B2 gene. A polymorphism in the 5' coding region of this gene (-344 C/T) is associated with hypertension, particularly with elevation of the aldosterone to renin ratio. A second polymorphism (a conversion in intron 2 to resemble that of the neighbouring 11beta-hydroxylase (CYP11B1) gene) is found in close linkage dysequilibrium with the variant at -344 C/T. The mechanism by which these variants predispose to cardiovascular disease and the precise intermediate phenotype associated with them remains speculative. DESIGN: We performed a focused physiological study in normal volunteers stratified by CYP11B2 genotype. PATIENTS: Twenty-three subjects homozygous for the T allele and 21 homozygous for the C allele of the -344 C/T polymorphism of CYP11B2 were studied. MEASUREMENTS: Basal and angiotensin II stimulated plasma and 24-h urinary steroid excretion during low (60 mmol/day) and high (160 mmol/day) sodium intake and plasma steroids after ACTH stimulation were measured. RESULTS: No influence of polymorphic variation on basal or stimulated plasma cortisol or aldosterone or other plasma steroid concentrations during either dietary phase was seen. However, excretion of tetrahydro-11-deoxycortisol (the urinary metabolite of 11-deoxycortisol), which is the precursor of cortisol) was increased in TT subjects during sodium restriction, consistent with impairment of zona fasciculata 11beta-hydroxylation. CONCLUSIONS: We conclude that this polymorphism has no major influence on normal zona glomerulosa function but is associated with a change in 11beta-hydroxylation in the zona fasciculata. The mechanism remains uncertain, but alteration of 11-deoxycortisol levels without change in cortisol suggests altered efficiency of 11beta-hydroxylation. In the long term, this may lead to a minor but chronic increase in ACTH drive to the gland, which may have consequences for steroid synthesis and predispose to the risk of cardiovascular disease. PMID- 15272912 TI - Proton magnetic resonance spectroscopy and single photon emission computed tomography study of the brain in asymptomatic young hyperlipidaemic Asian Indians in North India show early abnormalities. AB - OBJECTIVE: To evaluate brain metabolism and cerebral blood flow in young patients with hyperlipidaemia. PATIENTS AND METHODS: Proton magnetic resonance spectroscopy ((1)H MRS) and single photon emission computed tomography (SPECT) of the brain was carried out in 19 asymptomatic young patients with hyperlipidaemia (mean age 32.6 +/- 6.0 years, range 22-45 years) and 21 age-matched healthy controls divided into the following three groups; (i) hyperlipidaemics on pharmacological treatment (group 1, n = 13), (ii) hyperlipidaemics not on pharmacological treatment (group 2, n = 6) and (iii) control group of healthy subjects (group 3, n = 21). RESULTS: No statistical difference was observed in the brain metabolite ratios between controls and hyperlipidaemic patients (both treatment naive and treated) in the (1)H NMR study. Two hyperlipidaemic patients showed a lactate peak and one had a lipid peak. The SPECT study was abnormal in seven hyperlipidaemic patients. In the pooled data, 50% subjects with high serum triglyceride (TG) levels as opposed to 14% subjects with normal serum TG levels showed cerebral hypoperfusion. The choline/creatine (Cho/Cr) ratio of the occipital region of the brain showed correlation with the excess percentage of body fat (%BF) and low levels of high density lipoprotein cholesterol (HDL-C) compared to those with normal %BF and normal HDL-C levels, respectively, in pooled data of all subjects. The N-acetyl aspartate (NAA)/Cho ratio also showed correlation with hypercholesterolaemia. Serum TG levels were positively correlated with the NAA/Cr ratio (r = 0.62, P < 0.05) and the Cho/Cr ratio (r = 0.63, P < 0.05) in the parieto-temporal area in group 1 patients. CONCLUSION: The study revealed no difference in the brain metabolite ratios between controls and hyperlipidaemic patients, while some hyperlipidaemic patients showed abnormalities of cerebral blood flow. Brain metabolite ratios were also influenced by certain parameters of body composition and lipids. As abnormal body composition, hypertriglyceridaemia and low levels of HDL-C are prevalent in Asian Indians, such data are important and indicate a need for further study. PMID- 15272913 TI - Genetics of endometriosis: a role for the progesterone receptor gene polymorphism PROGINS? AB - OBJECTIVE: Endometriosis is a steroid-dependent disease with a particular genetic background, but the locations of possible genomic aberrations are still poorly clarified. We have investigated the potential association between endometriosis and the PROGINS 306 base pair insertion polymorphism in intron G of the progesterone receptor (PR) gene, which has been reported previously to segregate with this disease. DESIGN: In a case-control study, we examined the PROGINS polymorphism of the progesterone receptor gene in 131 Italian women affected by endometriosis diagnosed according to published criteria for the definition of the definite disease. Control subjects were represented by 127 Italian women without laparoscopic evidence of the disease. MEASUREMENTS: Peripheral blood samples, DNA extraction and polymerase chain reaction (PCR) were used to genotype women for the presence of the PROGINS polymorphism. RESULTS: We found a statistically significant difference in the distribution of PROGINS genotypes between patients with and without endometriosis. The frequency of the PROGINS allele T2 was 17.2% and 11%, respectively, in affected women and in controls [odds ratio (OR) = 1.7, 95% confidence interval (CI) 1.0-2.8]. This association was stronger in patients with more severe forms of endometriosis, such as an infiltrating disease or a disease characterized by severe pelvic adhesions (OR 2.4, 95% CI 1.2-4.8; and OR 2.7, 95% CI 1.4-5.3, respectively). Combination of the results from an earlier study and the current data indicates that carrying the allele variant T2 is associated with a twofold increase in the risk of developing endometriosis (OR 2.0, 95% CI 1.3-2.9). CONCLUSIONS: Our results further support the idea that the PROGINS polymorphism of the progesterone receptor may be associated with an increased risk of endometriosis. PMID- 15272914 TI - A polymorphic CA repeat in the IGF-I gene is associated with gender-specific differences in body height, but has no effect on the secular trend in body height. AB - OBJECTIVE: A polymorphism near the promoter region of the IGF-I gene has been associated with serum IGF-I levels, age-related decline of serum IGF-I levels, body height, birth weight and intima media thickness in hypertensive subjects. DESIGN AND METHODS: We investigated the association between the length of the IGF I alleles of this promoter polymorphism and IGF-I levels and body height. Furthermore, we investigated the potential influence of this polymorphism on final height in relationship to the secular trend of individuals born between 1917 and 1945. All subjects were participants of the Rotterdam Study. RESULTS: We observed, in analyses including only homozygous carriers, the highest IGF-I levels in homozygous carriers of the 192-bp allele (18.7 nmol/l +/- 0.6) and homozygous carriers of the 194-bp allele (17.7 nmol/l +/- 1.4). IGF-I levels were significantly lower in individuals with homozygous longer alleles [> 194-bp (12.0 nmol/l +/- 1.2; P < 0.001)] and homozygous shorter alleles [< 192-bp (15.6 nmol/l +/- 1.4; P < 0.05)] compared to homozygous carriers of the 192-bp and the 194-bp allele. In males and females separately, an optimum for serum IGF-I was also observed in homozygous carriers of the 192-bp and 194-bp allele. Only in males, homozygous carriers of the 192-bp allele were significantly taller than homozygous carriers of the shorter alleles (174.9 cm +/- 0.2 vs. 171.5 cm +/- 1.4; P = 0.01). When all subjects genotyped for the IGF-I promoter polymorphism were included in the analysis, a clear optimum for IGF-I levels and body height was observed in carriers of the 192-bp and/or 194-bp allele in the total population. Between 1917 and 1945, a secular trend in body height was observed in our Dutch population. Mean final body height was significantly higher in carriers of the most frequent alleles (192-bp and/or the 194-bp), than carriers of the remaining shorter and longer genotypes (P-trend < 0.01). CONCLUSIONS: In conclusion, we observed an optimum in IGF-I levels and final body height for the 192-bp and 194-bp allele of the IGF-I gene. A gender-specific effect of the IGF-I alleles on body height was observed. The secular trend in body height observed in our elderly Dutch population was similar for the different genotypes; carriers of the 192-bp and/or the 194-bp allele remained significantly taller throughout time. PMID- 15272915 TI - Lack of independent relationship between plasma adiponectin, leptin levels and bone density in nondiabetic female adolescents. AB - OBJECTIVES: Adiponectin has been implicated in the pathophysiology of metabolic syndrome and coronary artery disease in humans. Whether adiponectin is related to bone mineralization remains unclear in adults as well as in adolescents. In this study, we aimed to determine the relationship between plasma adiponectin, leptin concentrations and bone density, including total-body bone mineral density (BMD) and bone mineral content (BMC) in adolescence. PATIENTS AND MEASUREMENTS: We studied 105 nondiabetic female adolescents [mean age 15.4 +/- 1.9 years, and mean body mass index (BMI), 23.1 +/- 4.0 kg/m(2)]. A venous blood sample was taken after 12 h of fasting to measure fasting plasma adiponectin and leptin levels. BMD and BMC of the whole body were measured by dual-energy X-ray absorptiometry. RESULTS: In simple correlation analysis, plasma adiponectin and leptin levels correlated significantly with total-body BMD (r =-0.523 and r = 0.443, P < 0.001, respectively) and BMC (r =-0.471 and r = 0.396, P < 0.001, respectively). However, plasma adiponectin and leptin were related to both BMD and BMC in opposite directions. In multivariate linear regression analyses, only BMI or fat mass (FM) and Tanner stage, but not plasma adiponectin and leptin, were significantly related to BMD and BMC following adjustment for other variables. CONCLUSIONS: The results suggest that plasma adiponectin and leptin concentrations are not related to the total-body BMD and BMC independent of the chronological age, BMI or FM, and Tanner stage in nondiabetic female adolescents, although they were highly correlated in simple correlation analyses. The biologic roles of adiponectin in bone still need further clarification. PMID- 15272916 TI - Cabergoline addition to depot somatostatin analogues in resistant acromegalic patients: efficacy and lack of predictive value of prolactin status. AB - BACKGROUND: Somatostatin analogues (SA) are currently the mainstay in the medical treatment of acromegaly. However, even high doses of depot SA for prolonged periods do not achieve GH-IGF-I normalization in some patients. Even though some data were reported about the addition of cabergoline, a long-acting dopamine agonist (DA), to SA in resistant patients, definite data are still lacking. DESIGN: Prospective open trial. PATIENTS: In 19 acromegalic patients with active disease (34-82 years old, seven males, 12 females) resistant to chronic (9-12 months) depot SA (octreotide-LAR, 30 mg/28 days in 13 patients, lanreotide, 60 mg/28 days intramuscularly in six patients) cabergoline was added (combined treatment). In these patients, SA treatment had partially relieved GH and IGF-I hypersecretion but no patient had achieved 'safe' GH and normal IGF-I-values. Eight patients had PRL levels greater than 15 micro g/l (range 16-60 micro g/l; 1 micro g = 21.2 mIU). Immunohistochemistry (IHC) was positive for PRL in four out of eight operated patients. RESULTS: The addition of cabergoline, using the minimal effective and the maximal tolerated dose (range 1-3.5 mg/week), decreased GH from 6.6 +/- 0.9 to 4.6 +/- 0.6 micro g/l (P = 0.018), and IGF-I from 552 +/- 44 to 428 +/- 54 micro g/l (P = 0.019) after 6 months (median, range 3-18 Months). Combined treatment decreased GH to < 2.5 micro g/l in four patients (21%) and normalized IGF-I for age in eight patients (42%). It obtained a decline of both GH and IGF-I (-49 +/- 7%, and -47 +/- 5%, respectively) in nine patients (47%), and a partial improvement in six (32%) patients (GH decreased by 43 +/- 8% in four, and IGF-I by 35-38% in two patients). No change was observed in two patients, and worsening in two other patients. Results were not dependent on PRL status (serum levels or IHC). Combined treatment was well tolerated. CONCLUSIONS: The addition of cabergoline to depot SA-resistant acromegalic patients is effective, not dependent on PRL values and normalizes IGF-I levels in 42% of patients. The association of long-acting DA and SA deserves a more relevant role in the therapeutical algorithm of acromegaly. PMID- 15272917 TI - Polymorphisms in oestrogen and progesterone receptor genes: possible influence on prolactin levels in women. AB - OBJECTIVE: Oestrogen and progesterone are known to influence the release of human prolactin. The present study was undertaken in order to investigate the possible influence of polymorphisms of the genes encoding the oestrogen receptor (ER)alpha, ERbeta and the progesterone receptor (PGR), on prolactin levels in premenopausal women. DESIGN AND MEASUREMENTS: Serum levels of prolactin were measured in the follicular phase of the menstrual cycle. Subjects were genotyped with respect to a TA repeat polymorphism of the ERalpha gene, a CA repeat polymorphism of the ERbeta gene, and two polymorphisms of the PGR gene: one insertion polymorphism (PROGINS) and one single nucleotide polymorphism (G331A). SUBJECTS: A population-based cohort of 270 42-year-old women. RESULTS: The CA repeat polymorphism of the ERbeta gene and the G331A polymorphism of the PGR gene appeared to be associated with prolactin levels. In contrast, we found no evidence for an influence of the PROGINS polymorphism of the PGR gene or the TA repeat polymorphism of the ERalpha gene on the levels of this hormone. CONCLUSIONS: These data suggest that genetic variants of both the ERbeta and the PGR may influence prolactin release. PMID- 15272918 TI - Systematic dose-extension of octreotide LAR: the importance of individual tailoring of treatment in patients with acromegaly. AB - OBJECTIVE: The depot long-acting somatostatin analogue octreotide LAR (LAR) provides effective and well-tolerated treatment for acromegaly. Despite a 4 weekly recommended injection frequency, prolonged duration of GH suppression has been observed in some patients following treatment with long-acting somatostatin analogues. The aim of our study was to perform a prospective systematic study to determine whether extending the interval between doses of LAR allows maintenance of 'safe' GH in selected patients with acromegaly. PATIENTS AND METHODS: Twenty two patients (15 men, seven women), mean age 58.9 years (35-81 years) with active acromegaly (mGH > 5 mU/l), requiring treatment were selected to receive treatment with LAR. Eleven patients had received previous treatment with both transsphenoidal surgery and radiotherapy, while six had received surgery alone. All patients were commenced on treatment with 20 mg LAR intramuscularly (i.m.) every 4 weeks. Mean GH (mGH) was measured after three consecutive injections immediately prior to the fourth injection. The dose frequency was systematically reduced after every four injections if mGH < 5 mU/l. Once mGH > 5 mU/l, the dose frequency was increased and mGH reassessed. RESULTS: The dosing interval was successfully increased to greater than 4 weeks in 20/22 patients (90.9%). Six of 22 (27.3%) were receiving injections every 8 weeks and 3/22 (13.6%) every 12 weeks. GH and IGF-I were lower on treatment compared with baseline (P < 0.01). There was no difference in individual mGH and IGF-I between the values on 4 weekly dosing and those at final dose frequency. There was no relationship between final dose frequency and either mean GH or IGF-I prior to LAR, patient age, or previous treatment. The percentage suppression following 100 micro g octreotide subcutaneously did not predict subsequent dose frequency of LAR. The drug cost if patients had continued at 4-weekly intervals would be UK pound 187 850, compared with UK pound 101 065 for the individually titrated dose frequency (P < 0.01). This represents a final cost of 53.8% of the 4-weekly injection price. CONCLUSION: Individual tailoring of LAR administration maintains control of acromegaly, with reduced injection frequency and improved cost-effectiveness. PMID- 15272919 TI - Subclinical hypothyroidism is associated with a low-grade inflammation, increased triglyceride levels and predicts cardiovascular disease in males below 50 years. AB - OBJECTIVE: Mild thyroid failure is associated with an increased risk for development of atherosclerosis, but whether subclinical hypothyroidism is related to risk for cardiovascular disease is controversial. The purpose of the present study was to examine a possible association between subclinical hypothyroidism and cardiovascular disease. DESIGN: Cross-sectional study of a general population. PATIENTS: Twelve hundred and twelve subjects, men and women, between 20 and 69 years old without thyroid disease not treated with drugs interfering with thyroid function or analysis of TSH were included. MEASUREMENTS: Clinical signs of cardiovascular disease based on a questionnaire and medical records and laboratory analysis of lipids, atherothrombotic risk markers, C-reactive protein and TSH. RESULTS: The main findings were a high incidence of subclinical hypothyroidism (19.7%) in a general population. Subclinical hypothyroidism was associated with higher concentrations of triglycerides and C-reactive protein. Below 50 years of age cardiovascular disease was more frequent in males with subclinical hypothyroidism compared to euthyroid males. Subclinical hypothyroidism was a predictor of cardiovascular disease in males below 50 years with an odds ratio of 3.4 (95% confidence interval 1.6-6.8) for developing cardiovascular disease compared to euthyroid age-matched males. CONCLUSION: Our study demonstrates that patients with subclinical hypothyroidism have increased levels of triglycerides and signs of low-grade inflammation (raised C-reactive protein levels) and that subclinical hypothyroidism might be a risk factor for development of cardiovascular disease in younger males. PMID- 15272920 TI - BRAF mutations in an Italian cohort of thyroid cancers. AB - OBJECTIVE: Recently, a somatic point mutation of the BRAF gene (V599E) has been identified as the most common genetic event in papillary thyroid carcinoma (PTC) with a variable frequency (about 25-70%) in different series from USA, Japan, Portugal and Ukraine. DESIGN: In the present study, the genetic analysis of BRAF in an Italian cohort of 65 thyroid tumours with corresponding normal tissues and 21 thyroid benign disorders is reported. METHODS: For BRAF analysis, the somatic DNA was PCR amplified by means of specific intronic primers and PCR products were directly sequenced. Statistical analyses were obtained by means of Fisher's exact test. RESULTS: All mutations detected involved a T > A transversion at 1796 (V599E) and were heterozygous. Overall, BRAF(V599E) mutation was found in 18/56 (32.1%) PTCs. According to the histological type of the tumour, the mutation was present in 38.3% of cases of conventional PTC (18/47), in 0/6 follicular variant of PTC, in 0/3 oncocytic variant of PTC. No BRAF mutations were detected either in five follicular carcinomas, or in four poorly differentiated or undifferentiated cancers or in benign thyroid disorders. No statistically significant correlation of BRAF mutation with patient age and gender, with multicentricity of the tumour, with the lymphocytic infiltration of the tissue, with the stage and with the recurrence rate, was found. BRAF(V599E) tended to be associated, although not significantly, with a greater volume and extension of the tumour and with lymph-nodal metastases at surgery. CONCLUSIONS: In conclusion, the present study on the first Italian series of thyroid cancers shows a frequency of 38.3% of BRAF(V599E) in the classical variant of PTC, confirming the key role of this mutation in promoting tumourigenesis. PMID- 15272921 TI - The relationship between circulating osteoprotegerin levels and bone mineral metabolism in healthy women. AB - OBJECTIVE: Osteoprotegerin (OPG) is a recently identified cytokine that acts as a decoy receptor for the RANK ligand. Moreover, OPG has been shown to be an important inhibitor of osteoclastogenesis in animal models. However, the relationship between circulating OPG levels and female bone status in human populations is unclear. In this study we undertook to investigate the relationship between circulating OPG levels and bone mineral metabolism in healthy women. PATIENTS AND MEASUREMENTS: Our subjects were 287 women aged 37-73 years (mean age 51.5 years). The serum concentrations of OPG were determined by enzyme-linked immunosorbent assay (ELISA). The biochemical markers of bone turnover and FSH were measured using standard methods. Bone mineral densities at the lumbar spine and femoral neck were measured by dual-energy X-ray absorptiometry. RESULTS: Postmenopausal women had a significantly higher mean value of serum OPG than premenopausal women (1358.5 +/- 32.5 pg/ml vs. 1228.8 +/- 33.3 pg/ml, P < 0.01). Serum OPG levels were positively correlated with age (r = 0.169, P < 0.01), as were urine deoxypyridinoline levels (r = 0.133, P < 0.05) and serum FSH levels (r = 0.187, P < 0.01) in a bivariate correlation analyses. In a multiple regression analysis, only urine calcium excretion was identified as a significant predictor for serum OPG levels. CONCLUSIONS: Circulating OPG levels were found to be associated with urine calcium excretion and menopause in healthy women. Our observations suggest that circulating OPG levels reflect an antiresorptive activity in bone, and they are related to endogenous oestrogen levels. PMID- 15272922 TI - Marked GH secretion after ghrelin alone or combined with GH-releasing hormone (GHRH) in obese patients. AB - OBJECTIVES: Ghrelin is a 28-amino-acid peptide, predominantly produced by the stomach. It displays a strong GH-releasing activity mediated by the hypothalamus pituitary GH secretagogue (GHS)-receptor (GHS-R). There are different studies that suggest the importance of ghrelin in feeding and weight homeostasis. In obesity there is a markedly decreased GH secretion. For both children and adults, the greater the body mass index (BMI), the lower the GH response to provocative stimuli, including the response to GHRH. However, the response to the natural GH secretaogogue ghrelin is unclear at the present time. The aim of the present study was to evaluate the GH response to ghrelin alone or combined with GHRH in a group of obese patients, in order to further understand the deranged GH secretory mechanisms in obesity and to clarify the mechanism of action of ghrelin. PATIENTS AND MEASUREMENTS: Six obese female patients (31 +/- 3.4 years) with a BMI of 36.1 +/- 7.7 kg/m(2) were studied. As a control group, six normal nonobese female subjects of similar age and sex were studied. Four tests were performed: placebo, GHRH [1 micro g/kg, no more than 100 micro g, intravenous (i.v.)], ghrelin (1 micro g/kg, no more than 100 micro g, i.v.) and GHRH (1 micro g/kg, no more than 100 micro g, i.v.) plus ghrelin (1 micro g/kg, no more than 100 micro g, i.v.). Blood samples were taken at appropriate intervals for determination of GH. Statistical analyses were performed by Wilcoxon and by Mann-Whitney tests. RESULTS: After GHRH, the median peak GH secretion in obese patients was 2.4 micro g/l (range 0.9-8.9 micro g/l). Ghrelin-induced GH secretion showed in obese patients a median peak of 24.4 micro g/l (range 7.4-85.0 micro g/l), significantly greater than the response after GHRH (P < 0.05). After the combined administration of GHRH plus ghrelin in obese patients the median peak GH secretion was 39.9 micro g/l (range 19.2-120.0 micro g/l), significantly greater than the response after GHRH (P < 0.05) or ghrelin (P < 0.05). GHRH-induced GH secretion in normal control subjects showed a median peak of 25.0 micro g/l (range 16.5-33.4 micro g/l). Ghrelin-induced GH secretion in normal showed a median peak of 68.5 micro g/l (range 22.5-119.5 micro g/l), significantly greater than the response after GHRH (P < 0.05). After the combined administration of GHRH plus ghrelin, in normal subjects the median peak GH secretion was 117.8 micro g/l (range 77.5-280.1 micro g/l), significantly greater than the response after GHRH or ghrelin alone (P < 0.05). When we compare the response of normal and obese patients, after GHRH alone, it was markedly decreased in obese people when compared with normal patients (P < 0.05) with a median GH peak of 25.0 micro g/l (range 16.5-33.4 micro g/l) and 2.4 micro g/l (range 0.9-8.9 micro g/l) for normal and obese patients, respectively. When we compare the response of normal and obese patients, after ghrelin alone or GHRH plus ghrelin, it was only blunted in obese subjects when compared with normal subjects with a median GH peak of 68.5 micro g/l (range 22.5-119.5 micro g/l) and 24.4 micro g/l (range 7.4-85 micro g/l) for normal and obese subjects, respectively, after ghrelin alone (P < 0.05) and a median GH peak of 117.8 micro g/l (range 77.5-280.1 micro g/l) and 39.9 micro g/l (range 19.2-120.0 micro g/l) for normal and obese patients, respectively, after GHRH plus ghrelin (P < 0.05). CONCLUSIONS: This study has demonstrated a massive GH response to ghrelin alone or combined with GHRH in obese patients, suggesting that altered ghrelin secretion could play a major role in the blunted GH secretion present in obese patients. PMID- 15272923 TI - Effects of clomiphene and raloxifene on gonadotrophin secretion in postmenopausal women: evidence of nongenomic action. AB - OBJECTIVE: To evaluate the effect of raloxifene (R) and clomiphene (Cl) on FSH and LH secretion in postmenopausal women. DESIGN: Postmenopausal volunteer women participated in two experimental (Exp) procedures. In Group 1, the women received R (180 mg/day orally) for 30 days plus oestradiol (E2) through skin patches (100 microg/24 h) from days 21 to 30 (R-Exp). After a month's break the same women received Cl (150 mg/day orally) for 30 days plus E2 as above (Cl-Exp). In Group 2, the women received E2 for 30 days plus R from days 21 to 30 (R-Exp) and after a month's break they received E2 for 30 days plus Cl from days 21 to 30 (Cl-Exp). Daily doses were as in Group 1. A GnRH test (100 microg intravenously) was performed in all women on days 0, 10, 20 and 30 of each experiment. PATIENTS: Sixteen healthy postmenopausal women were divided into two groups (eight women in each group). MEASUREMENTS: The area under the curve (AUC) of DeltaFSH and DeltaLH response to GnRH (net increase above the basal value) was calculated. RESULTS: In Group 1, basal levels of FSH and LH did not change significantly during the R Exp, while they decreased significantly in the Cl-Exp (P < 0.001). The addition of E2 did not have any effect. The AUC of LH response to GnRH increased significantly in the R-Exp (P < 0.05) and that of FSH in the Cl-Exp (P < 0.05). In Group 2, basal levels of FSH and LH declined significantly during treatment with E2 in both the R-Exp (P < 0.01) and the Cl-Exp (P < 0.001). However, the addition of Cl (for 10 days) interrupted this decrease, while the addition of R stimulated FSH levels significantly (P < 0.05). E2 suppressed significantly the AUC of LH in both experiments (P < 0.05). The addition of Cl did not affect the AUC in response to GnRH, while the addition of R increased the AUC of both LH and FSH (P < 0.05). CONCLUSIONS: These results demonstrate for the first time that in contrast to Cl, R does not exert oestrogenic effects on basal gonadotrophin secretion. Although the antioestrogenic action of these drugs was evident only after pretreatment with E2, both R and Cl stimulated GnRH-induced gonadotrophin secretion in oestrogen-deprived women. It is hypothesized that these two compounds sensitize the pituitary to GnRH through mechanisms not involving the oestrogen receptor complex (nongenomic). PMID- 15272924 TI - Association of a thyroglobulin gene polymorphism with Hashimoto's thyroiditis in the Japanese population. AB - OBJECTIVE: The aetiology of the autoimmune thyroid diseases (AITDs), Graves' disease (GD) and Hashimoto's thyroiditis is largely unknown. However, genetic susceptibility is believed to play a major role. Two whole genome scans from Japan and from the USA identified a locus on chromosome 8q24 which showed evidence for linkage with AITD and HT. Recent studies have demonstrated an association between a Tg polymorphisms and AITD, suggesting that Tg is the susceptibility gene on 8q24. PATIENTS: We studied 308 Japanese AITD patients (194 GD and 114 HT patients) and 417 Japanese control subjects in association studies. DESIGN: Case-control association studies were performed using D8S284 and D8S272, microsatellite markers located in the 8q24 region, Tgms1 and Tgms2, microsatellite markers in introns 10 and 27, respectively, of Tg, and a SNP in exon 33 of Tg. RESULTS: No differences in allele frequencies were observed between AITD patients and controls for D8S284, D8S272 and Tgms1. Similarly, for Tgms2 and the exon 33 SNP no significant differences in allele frequency distribution were observed for all AITD patients. However, when analysing the HT patients alone we found a significant association between the 330 bp/352 bp genotype of Tgms2 and HT (HT = 16.7%, controls = 7.1%; corrected P-value = 0.01, OR = 2.6). CONCLUSION: Our results confirm the previous reports of an association between the Tg gene and AITD and suggest that Tg is an AITD susceptibility gene. PMID- 15272925 TI - A single nucleotide polymorphism in the CD40 gene on chromosome 20q (GD-2) provides no evidence for susceptibility to Graves' disease in UK Caucasians. AB - OBJECTIVE: A genome-wide screen in Graves' disease (GD) has shown linkage to chromosome 20q, designated GD-2. The gene encoding CD40, which stimulates lymphocyte proliferation and differentiation, maps to this region, and a single nucleotide polymorphism (SNP) at position -1 of the Kozak sequence within the gene has been reported to be associated with GD. The aim of this study was to determine whether this SNP of the CD40 gene confers susceptibility to GD in UK Caucasians. DESIGN: A large case-control cohort consisting of 800 patients with GD, and 785 control subjects with no history of autoimmune disease, was used to genotype this SNP by polymerase chain reaction restriction fragment length polymorphism. RESULTS: Despite adequate power (> 99%) to detect an effect, if present (odds ratio of 1.5), no significant difference in allele or genotype frequency of the CD40 SNP was observed between patients with GD and control subjects (P = 0.087 and P = 0.145, respectively). CONCLUSIONS: These data suggest that this polymorphism of the CD40 gene is not associated with GD in the UK and is therefore not contributing to disease susceptibility in the chromosomal region designated GD-2. PMID- 15272926 TI - Abnormal regulation of thirst and vasopressin secretion following surgery for craniopharyngioma. AB - OBJECTIVE: In this study we aimed to establish the frequency of postoperative diabetes insipidus and the incidence and characteristics of abnormalities of thirst in a cohort of patients with craniopharyngioma, in whom neurosurgery had been performed. DESIGN: Diabetes insipidus was determined by either standard criteria for diagnosis in the immediate postoperative period, or by water deprivation test, in all craniopharyngioma and pituitary tumour patients who underwent surgery in Beaumont Hospital between the years 1986 and 1998. Osmoregulated thirst and vasopressin release were studied during a 2-h infusion of hypertonic (5%) saline followed by a 30-min period of free access to water. PATIENTS: Data on the incidence of postoperative diabetes insipidus was collected in 26 patients with craniopharyngioma and 154 patients with pituitary adenomata. We recruited 16 healthy control patients, 16 patients with cranial diabetes insipidus following pituitary tumour surgery and 16 patients with cranial diabetes insipidus following craniopharyngioma resection for the hypertonic saline infusion study. RESULTS: Twenty-five patients out of 26 (96%) patients developed diabetes insipidus after surgery for craniopharyngioma, a much higher incidence than after surgery for suprasellar (26/88, 30%, P < 0.001) or intrasellar pituitary tumours (9/66, 14%, P < 0.001). Hypertonic saline infusion identified abnormal thirst responses in five of the 16 craniopharygioma patients studied; all of the pituitary tumour patients had a normal thirst response. Three of the craniopharyngioma patients had adipsic diabetes insipidus whilst two had polydipsic diabetes insipidus. CONCLUSION: This study demonstrates following surgery for craniopharyngioma there is a high incidence of cranial diabetes insipidus and a significant incidence of abnormal thirst responses to osmotic stimuli. PMID- 15272927 TI - The expression of PAX8-PPARgamma rearrangements is not specific to follicular thyroid carcinoma. PMID- 15272928 TI - Endothelin axis expression in medullary thyroid carcinoma: a potential therapeutic target. PMID- 15272929 TI - Manifestation of thyroid autoimmunity in patients successfully treated for hypercortisolism. PMID- 15272930 TI - Accounting for regional background and population size in the detection of spatial clusters and outliers using geostatistical filtering and spatial neutral models: the case of lung cancer in Long Island, New York. AB - BACKGROUND: Complete Spatial Randomness (CSR) is the null hypothesis employed by many statistical tests for spatial pattern, such as local cluster or boundary analysis. CSR is however not a relevant null hypothesis for highly complex and organized systems such as those encountered in the environmental and health sciences in which underlying spatial pattern is present. This paper presents a geostatistical approach to filter the noise caused by spatially varying population size and to generate spatially correlated neutral models that account for regional background obtained by geostatistical smoothing of observed mortality rates. These neutral models were used in conjunction with the local Moran statistics to identify spatial clusters and outliers in the geographical distribution of male and female lung cancer in Nassau, Queens, and Suffolk counties, New York, USA. RESULTS: We developed a typology of neutral models that progressively relaxes the assumptions of null hypotheses, allowing for the presence of spatial autocorrelation, non-uniform risk, and incorporation of spatially heterogeneous population sizes. Incorporation of spatial autocorrelation led to fewer significant ZIP codes than found in previous studies, confirming earlier claims that CSR can lead to over-identification of the number of significant spatial clusters or outliers. Accounting for population size through geostatistical filtering increased the size of clusters while removing most of the spatial outliers. Integration of regional background into the neutral models yielded substantially different spatial clusters and outliers, leading to the identification of ZIP codes where SMR values significantly depart from their regional background. CONCLUSION: The approach presented in this paper enables researchers to assess geographic relationships using appropriate null hypotheses that account for the background variation extant in real-world systems. In particular, this new methodology allows one to identify geographic pattern above and beyond background variation. The implementation of this approach in spatial statistical software will facilitate the detection of spatial disparities in mortality rates, establishing the rationale for targeted cancer control interventions, including consideration of health services needs, and resource allocation for screening and diagnostic testing. It will allow researchers to systematically evaluate how sensitive their results are to assumptions implicit under alternative null hypotheses. PMID- 15272931 TI - Multi-component based cross correlation beat detection in electrocardiogram analysis. AB - BACKGROUND: The first stage in computerised processing of the electrocardiogram is beat detection. This involves identifying all cardiac cycles and locating the position of the beginning and end of each of the identifiable waveform components. The accuracy at which beat detection is performed has significant impact on the overall classification performance, hence efforts are still being made to improve this process. METHODS: A new beat detection approach is proposed based on the fundamentals of cross correlation and compared with two benchmarking approaches of non-syntactic and cross correlation beat detection. The new approach can be considered to be a multi-component based variant of traditional cross correlation where each of the individual inter-wave components are sought in isolation as opposed to being sought in one complete process. Each of three techniques were compared based on their performance in detecting the P wave, QRS complex and T wave in addition to onset and offset markers for 3000 cardiac cycles. RESULTS: Results indicated that the approach of multi-component based cross correlation exceeded the performance of the two benchmarking techniques by firstly correctly detecting more cardiac cycles and secondly provided the most accurate marker insertion in 7 out of the 8 categories tested. CONCLUSION: The main benefit of the multi-component based cross correlation algorithm is seen to be firstly its ability to successfully detect cardiac cycles and secondly the accurate insertion of the beat markers based on pre-defined values as opposed to performing individual gradient searches for wave onsets and offsets following fiducial point location. PMID- 15272932 TI - Ciprofibrate therapy in patients with hypertriglyceridemia and low high density lipoprotein (HDL)-cholesterol: greater reduction of non-HDL cholesterol in subjects with excess body weight (The CIPROAMLAT study). AB - BACKGROUND: Hypertriglyceridemia in combination with low HDL cholesterol levels is a risk factor for cardiovascular disease. Our objective was to evaluate the efficacy of ciprofibrate for the treatment of this form of dyslipidemia and to identify factors associated with better treatment response. METHODS: Multicenter, international, open-label study. Four hundred and thirty seven patients were included. The plasma lipid levels at inclusion were fasting triglyceride concentrations between 1.6-3.9 mM/l and HDL cholesterol < or = 1.05 mM/l for women and < or = 0.9 mM/l for men. The LDL cholesterol was below 4.2 mM/l. All patients received ciprofibrate 100 mg/d. Efficacy and safety parameters were assessed at baseline and at the end of the treatment. The primary efficacy parameter of the study was percentage change in triglycerides from baseline. RESULTS: After 4 months, plasma triglyceride concentrations were decreased by 44% (p < 0.001). HDL cholesterol concentrations were increased by 10% (p < 0.001). Non-HDL cholesterol was decreased by 19%. A greater HDL cholesterol response was observed in lean patients (body mass index < 25 kg/m2) compared to the rest of the population (8.2 vs 19.7%, p < 0.001). In contrast, cases with excess body weight had a larger decrease in non-HDL cholesterol levels (-20.8 vs -10.8%, p < 0.001). There were no significant complications resulting from treatment with ciprofibrate. CONCLUSIONS: Ciprofibrate is efficacious for the correction of hypertriglyceridemia / low HDL cholesterol. A greater decrease in non-HDL cholesterol was found among cases with excess body weight. The mechanism of action of ciprofibrate may be influenced by the pathophysiology of the disorder being treated. PMID- 15272933 TI - Stimulation of MMP-11 (stromelysin-3) expression in mouse fibroblasts by cytokines, collagen and co-culture with human breast cancer cell lines. AB - BACKGROUND: Matrix metalloproteinases (MMPs) are central to degradation of the extracellular matrix and basement membrane during both normal and carcinogenic tissue remodeling. MT1-MMP (MMP-14) and stromelysin-3 (MMP-11) are two members of the MMP family of proteolytic enzymes that have been specifically implicated in breast cancer progression. Expressed in stromal fibroblasts adjacent to epithelial tumour cells, the mechanism of MT1-MMP and MMP-11 induction remains unknown. METHODS: To investigate possible mechanisms of induction, we examined the effects of a number of plausible regulatory agents and treatments that may physiologically influence MMP expression during tumour progression. Thus NIH3T3 and primary mouse embryonic fibroblasts (MEFs) were: a) treated with the cytokines IL-1beta, IL-2, IL-6, IL-8 and TGF-beta for 3, 6, 12, 24, and 48 hours; b) grown on collagens I, IV and V; c) treated with fibronectin, con-A and matrigel; and d) co-cultured with a range of HBC (human breast cancer) cell lines of varied invasive and metastatic potential. RESULTS: Competitive quantitative RT PCR indicated that MMP-11 expression was stimulated to a level greater than 100%, by 48 hour treatments of IL-1beta, IL-2, TGF-beta, fibronectin and collagen V. No other substantial changes in expression of MMP-11 or MT1-MMP in either tested fibroblast culture, under any treatment conditions, were observed. CONCLUSION: We have demonstrated significant MMP-11 stimulation in mouse fibroblasts using cytokines, matrix constituents and HBC cell lines, and also some inhibition of MT1-MMP. Our data suggest that the regulation of these genes in the complex stromal-epithelial interactions that occur in human breast carcinoma, is influenced by several mechanisms. PMID- 15272934 TI - The six-minute walk test in community dwelling elderly: influence of health status. AB - BACKGROUND: The 6 minutes walk test (6MWT) is a useful assessment instrument for the exercise capacity of elderly persons. The impact of the health status on the 6MWT-distance in elderly, however, remains unclear, reducing its value in clinical settings. The objective of this study was to investigate to what extent the 6MWT-distance in community dwelling elderly is determined by health conditions. METHODS: One hundred and fifty-six community dwelling elderly people (53 male, 103 female) were assessed for health status and performed the 6MWT. After clinical evaluation, electrocardiography and laboratory examination participants were categorized into a stratified six-level classification system according to their health status, going from A (completely healthy) to D (signs of active disease at the moment of examination). RESULTS: The mean 6MWT-distance was 603 m (SD = 178). The 6MWT-distance decreased significantly with increasing age (ANOVA p = 0.0001) and with worsening health status (ANCOVA, corrected for age p < 0.001).A multiple linear regression model with health status, age and gender as independent variables explained 31% of the 6MWT-distance variability. Anthropometrical measures (stature, weight and BMI) did not significantly improve the prediction model. A significant relationship between 6MWT-distance and stature was only present in category A (completely healthy). CONCLUSIONS: Significant differences in 6MWT-distance are observed according to health status in community-dwelling elderly persons. The proposed health categorizing system for elderly people is able to distinguish persons with lower physical exercise capacity and can be useful when advising physical trainers for seniors. PMID- 15272935 TI - Accuracy of cDNA microarray methods to detect small gene expression changes induced by neuregulin on breast epithelial cells. AB - BACKGROUND: cDNA microarrays are a powerful means to screen for biologically relevant gene expression changes, but are often limited by their ability to detect small changes accurately due to "noise" from random and systematic errors. While experimental designs and statistical analysis methods have been proposed to reduce these errors, few studies have tested their accuracy and ability to identify small, but biologically important, changes. Here, we have compared two cDNA microarray experimental design methods with northern blot confirmation to reveal changes in gene expression that could contribute to the early antiproliferative effects of neuregulin on MCF10AT human breast epithelial cells. RESULTS: We performed parallel experiments on identical samples using a dye-swap design with ANOVA and an experimental design that excludes systematic biases by "correcting" experimental/control hybridization ratios with control/control hybridizations on a spot-by-spot basis. We refer to this approach as the "control correction method" (CCM). Using replicate arrays, we identified a decrease in proliferation genes and an increase in differentiation genes. Using an arbitrary cut-off of 1.7-fold and p values <0.05, we identified a total of 32 differentially expressed genes, 9 with the dye-swap method, 18 with the CCM, and 5 genes with both methods. 23 of these 32 genes were subsequently verified by northern blotting. Most of these were <2-fold changes. While the dye-swap method (using either ANOVA or Bayesian analysis) detected a smaller number of genes (14 16) compared to the CCM (46), it was more accurate (89-92% vs. 75%). Compared to the northern blot results, for most genes, the microarray results underestimated the fold change, implicating the importance of detecting these small changes. CONCLUSIONS: We validated two experimental design paradigms for cDNA microarray experiments capable of detecting small (<2-fold) changes in gene expression with excellent fidelity that revealed potentially important genes associated with the anti-proliferative effects of neuregulin on MCF10AT breast epithelial cells. PMID- 15272937 TI - What is the role of theory in health behavior change interventions? PMID- 15272936 TI - Graph-based iterative Group Analysis enhances microarray interpretation. AB - BACKGROUND: One of the most time-consuming tasks after performing a gene expression experiment is the biological interpretation of the results by identifying physiologically important associations between the differentially expressed genes. A large part of the relevant functional evidence can be represented in the form of graphs, e.g. metabolic and signaling pathways, protein interaction maps, shared GeneOntology annotations, or literature co-citation relations. Such graphs are easily constructed from available genome annotation data. The problem of biological interpretation can then be described as identifying the subgraphs showing the most significant patterns of gene expression. We applied a graph-based extension of our iterative Group Analysis (iGA) approach to obtain a statistically rigorous identification of the subgraphs of interest in any evidence graph. RESULTS: We validated the Graph-based iterative Group Analysis (GiGA) by applying it to the classic yeast diauxic shift experiment of DeRisi et al., using GeneOntology and metabolic network information. GiGA reliably identified and summarized all the biological processes discussed in the original publication. Visualization of the detected subgraphs allowed the convenient exploration of the results. The method also identified several processes that were not presented in the original paper but are of obvious relevance to the yeast starvation response. CONCLUSIONS: GiGA provides a fast and flexible delimitation of the most interesting areas in a microarray experiment, and leads to a considerable speed-up and improvement of the interpretation process. PMID- 15272938 TI - How can Health Behavior Theory be made more useful for intervention research? AB - BACKGROUND: The present paper expresses the author's views about the practical utility of Health Behavior Theory for health behavior intervention research. The views are skeptical and perhaps even a bit exaggerated. They are, however, also based on 20-plus years of in-the-trenches research focused on improving health behavior practice through research. DISCUSSION: The author's research has been theoretically driven and has involved measurement of varying variables considered to be important theoretical mediators and moderators of health behavior. Regretfully, much of this work has found these variables wanting in basic scientific merit. Health Behavior Theory as we have known it over the last 25 years or so has been dominated by conceptualizations of behavior change processes that highlight cognitive decision-making. Although much of health behavior practice targets what people do rather than what they think, the logic of focusing on thoughts is that what people think about is the key to what they will do in the future, and that interventions that can measure and harness those processes will succeed to a greater extent than those that do not. Unfortunately, in the author's experience, the premise of cognitive theories has fallen short empirically in a number of ways. The cognitive schemata favored by most health behavior theories are difficult to measure, they do not predict behavioral outcomes very well, there is little evidence that they cause behavior, and they are hard to change directly. SUMMARY: It is suggested that health behavior researchers reconsider their use of these theories in favor of models whose variables are more accessible to observation and experimental manipulation and that most importantly have strong empirical support. PMID- 15272939 TI - Proposed methods for reviewing the outcomes of health research: the impact of funding by the UK's 'Arthritis Research Campaign' AB - BACKGROUND: External and internal factors are increasingly encouraging research funding bodies to demonstrate the outcomes of their research. Traditional methods of assessing research are still important, but can be merged into broader multi dimensional categorisations of research benefits. The onus has hitherto been on public sector funding bodies, but in the UK the role of medical charities in funding research is particularly important and the Arthritis Research Campaign, the leading medical charity in its field in the UK, commissioned a study to identify the outcomes from research that it funds. This article describes the methods to be used. METHODS: A case study approach will enable narratives to be told, illuminating how research funded in the early 1990s was (or was not) translated into practice. Each study will be organised using a common structure, which, with careful selection of cases, should enable cross-case analysis to illustrate the strengths of different modes and categories of research. Three main interdependent methods will be used: documentary and literature review; semi structured interviews; and bibliometric analysis. The evaluative framework for organising the studies was previously used for assessing the benefits from health services research. Here, it has been specifically amended for a medical charity that funds a wide range of research and is concerned to develop the careers of researchers. It was further refined in three pilot studies. The framework has two main elements. First, a multi-dimensional categorisation of benefits going from the knowledge produced in peer reviewed journal articles through to the health and potential economic gain. The second element is a logic model, which, with various stages, should provide a way of organising the studies. The stock of knowledge is important: much research, especially basic, will feed into it and influence further research rather than directly lead to health gains. The cross case analysis will look for factors associated with outcomes. CONCLUSIONS: The pilots confirmed the applicability of the methods for a full study which should assist the Arthritis Research Campaign to demonstrate the outcomes from its funding, and provide it with evidence to inform its own policies. PMID- 15272940 TI - A unique dedifferentiated tumor of the retroperitoneum. AB - BACKGROUND: Dedifferentiated liposarcomas represent heterogeneous tumors with lipomatous and nonlipomatous elements starkly juxtaposed. It is thought that the high grade nonlipomatous elements of the tumor portend a worse prognosis. CASE PRESENTATION: A 19.8 kg heterogeneous retroperitoneal tumor was successfully and completely resected. Because of its extent, no additional treatment modalities were practicable. The tumor soon recurred. The recurrent tumor differed from the primary tumor in that it was more homogeneous, consisting mainly of nonlipogenic, calcific tissue. CONCLUSIONS: Dedifferentiated liposarcomas are known to have a very high recurrence rate. The biological behavior of dedifferentiated liposarcomas is likely dictated by the most aggressive element of these heterogeneous tumors. PMID- 15272941 TI - Quality of life and emotional distress in advanced prostate cancer survivors undergoing chemotherapy. AB - Prostate cancer continues to occur in over 230,000 men each year. Although the majority of these will be diagnosed in the early stages, there remains a proportion who will either be diagnosed in late stage disease or develop progressive disease. In patients with advanced disease, research has recently focused on using chemotherapy for symptom management and palliation. Given that the focus is not on cure, the effect of chemotherapy on quality of life is of utmost importance. The present article will 1) summarize the current chemotherapeutic studies that have included a quality of life component, with a particular focus on pain and fatigue, 2) discuss the issue of distress in advanced prostate cancer patients treated with chemotherapy, and 3) suggest future research directions. From the studies that have investigated quality of life, it appears that several chemotherapeutic agents reduce pain and fatigue, although the development of fatigue is often the dose-limiting factor of some agents. The assessment of overall quality of life has occurred in several studies, however, an examination into the impact of chemotherapy on functional status and interpersonal relationships has not been studied. Finally, in contrast to the numerous studies in early stage prostate cancer patients, the presence and effect of distress in chemotherapy-treated prostate patients has not been examined. As such, increased attention is needed to quality of life during phase I-III chemotherapy trials. PMID- 15272943 TI - Attributional biases in the service of stereotype maintenance: a schema maintenance through compensation analysis. AB - Six experiments were conducted to test assumptions of a schema-maintenance through compensation analysis. The results of these experiments indicated that perceivers can compensate for the inconsistent action of one individual (the target) by altering their attribution concerning the action of a fellow group member. When the target performed an inconsistent behavior, perceivers compensated by making especially extreme stereotypically consistent attributions concerning a fellow group member's subsequent action. In addition, in Experiment 5, perceivers compensated via a fellow group member for a target's inconsistent action while maintaining their general view of group members. Experiments also provided tests of the capability and motivation assumptions of the schema maintenance through compensation analyses. PMID- 15272942 TI - Using a Geographical Information System to investigate the relationship between reported cryptosporidiosis and water supply. AB - BACKGROUND: This paper reports on a study investigating the epidemiology of sporadic cryptosporidiosis in the North West of England and Wales using a Geographical Information System (GIS) to map location of residence of cases. Some 747 reports of cases were made to CDSC North West of which 649 reports were suitable for analysis. Cases were plotted on the maps of water supply zone and water quality area boundaries, provided by the two main water utilities. RESULTS: It was notable that there were major spatial variations in attack rate across the North West and Wales. The most dramatic example was the large difference between the Greater Manchester conurbation with many reports and Liverpool with none. Given the distribution of previously detected waterborne outbreaks in the region it was initially thought that drinking water source may be an explanation. However, an analysis of the distribution of cases in the Greater Manchester area showed no correlation with any of five water supplies that serve the conurbation. CONCLUSIONS: Our study has shown a dramatic variation in the incidence of laboratory confirmed cryptosporidiosis within two regions of the United Kingdom. Further analysis has not been able to prove drinking water as a likely explanation of this variation which so far remains unexplained. PMID- 15272944 TI - Younger achievement age predicts shorter life for governors: testing the precocity-longevity hypothesis with artifact controls. AB - McCann's precocity-longevity hypothesis suggests that the prerequisites, concomitants, and consequences of early peaks in career achievement may foster the conditions for premature death. In the present test of the precocity longevity hypothesis, it was predicted that state governors elected at younger ages live shorter lives. Two competing explanatory frameworks, the life expectancy artifact and the selection bias artifact, also were tested. In a sample of 1,672 male governors, the precocity-longevity prediction was supported, and it was demonstrated with correlation, regression, and subsample construction strategies that the life expectancy and selection bias artifacts were not sufficient to account or the significant positive correlation between election age and death age. The positive correlation also was maintained when year of birth, years of service, span of service, and state of election were statistically controlled. PMID- 15272945 TI - Choosing social situations: the relation between automatically activated racial attitudes and anticipated comfort interacting with african americans. AB - This research explored how White students' automatically activated racial attitudes and motivations to control prejudiced reactions (MCPR) related to their self-reported comfort and willingness to interact with Black partners in a variety of situations. Participants completed the MCPR scale and a priming task that provides an unobtrusive measure of automatically activated racial attitudes. In a later session, participants rated their comfort and willingness to enter eight situations varying in their degree of intimacy and scriptedness and then rerated each situation while imagining different interaction partners, including a Black individual. When the situations were scripted or participants were low on the restraint to avoid dispute factor of the MCPR scale, participants expressed willingness to interact with Black partners. When the situation was unscripted and participants were characterized by higher restraint scores, anticipated comfort varied as a function of automatically activated racial attitudes and the concern with acting prejudiced factor of the MCPR scale. PMID- 15272946 TI - Respect for group members: intragroup determinants of collective identification and group-serving behavior. AB - This experiment examined how (disrespectful vs. respectful) treatment and (negative vs. positive) performance evaluation, both received from the same fellow group members, affects collective identification and willingness to engage in group-serving behavior. It was predicted and found that respectful as opposed to disrespectful intragroup treatment increased collective identification and willingness to engage in group-serving behavior in the immediate group situation, irrespective of whether intragroup evaluation was positive or negative. There was also evidence of a mediating role of collective identification. Regression analyses based on the measurement of perceived intragroup treatment and perceived intragroup evaluation as continuous variables corroborated these findings but also pointed to limits of the positive effects of respectful intragroup treatment. Finally, the interrelation of treatment and evaluation by fellow group members as two possible components of intragroup respect are discussed as well as the political dimension of research on intragroup respect. PMID- 15272947 TI - A case of collective responsibility: who else was to blame for the Columbine high school shootings? AB - Two studies examined perceptions of collective responsibility for the April 20, 1999, shootings at Columbine High School in Littleton, Colorado. Collective responsibility refers to the perception that others, besides the wrongdoers themselves, are responsible for the event. In Study 1, the authors assessed perceptions of the shooters' parents and their peer group (the Trenchcoat Mafia), whereas Study 2 tested perceptions of collective responsibility across a range of groups. In both studies, perceptions of a target group's entitativity predicted judgments of collective responsibility. This relationship was mediated by two situational construals that justify applying collective responsibility: responsibility by commission (encouraging or facilitating the event) and responsibility by omission (failing to prevent the event). Study 2 also determined that perceptions of authority predicted judgments of collective responsibility for the Columbine shootings and was mediated by inferences of omission. Future directions in collective responsibility research are discussed. PMID- 15272948 TI - Mediator of moderators: temporal stability of intention and the intention behavior relation. AB - Intention certainty, past behavior, self-schema, anticipated regret, and attitudinal versus normative control all have been found to moderate intention behavior relations. It is argued that moderation occurs because these variables produce "strong" intentions. Stability of intention over time is a key index of intention strength. Consequently, it was hypothesized that temporal stability of intention would mediate moderation by these other moderators. Participants (N = 185) completed questionnaire measures of theory of planned behavior constructs and moderator variables at two time points and subsequently reported their exercise behavior. Findings showed that all of the moderators, including temporal stability, were associated with significant improvements in consistency between intention and behavior. Temporal stability also mediated the effects of the other moderators, supporting the study hypothesis. PMID- 15272949 TI - Relations between high and low power groups: the importance of legitimacy. AB - Using a social identity perspective, two experiments examined the effects of power and the legitimacy of power differentials on intergroup bias. In Experiment 1, 125 math-science students were led to believe that they had high or low representation in a university decision-making body relative to social-science students and that this power position was either legitimate or illegitimate. Power did not have an independent effect on bias; rather, members of both high and low power groups showed more bias when the power hierarchy was illegitimate than when it was legitimate. This effect was replicated in Experiment 2 (N = 105). In addition, Experiment 2 showed that groups located within an unfair power hierarchy expected the superordinate power body to be more discriminatory than did those who had legitimately high or low power. The results are discussed in terms of their implications for group relations. PMID- 15272950 TI - Framing social judgment: self-ingroup comparison and perceived discrimination. AB - Members of disadvantaged groups often report that they are less vulnerable to discrimination than is the average member of their group. In two experiments, we examined how the framing of self-ingroup comparisons moderates this phenomenon. In Experiment 1, participants estimated the relative likelihood that either they, compared to the average member of their ingroup, would experience discrimination or that the average member of their ingroup, compared to themselves, would experience discrimination. In Experiment 2, a direct manipulation of self-ingroup similarity was added to this framing manipulation. Across the experiments, the results demonstrated that conditions highlighting differences between the individual and the group led participants to perceive themselves as much less vulnerable to discrimination than their group, relative to conditions highlighting similarities between the individual and the group. PMID- 15272951 TI - The excluded player in coalition formation. AB - Coalition research generally assumes that people strive to maximize their share of the coalition payoff and that they exclude others from joining a coalition if these others are not needed to obtain the coalition payoff. In two experiments, the authors show that this view is too narrow and that willingness to include such others is dependent on the extent to which people feel that exclusion affects the payoff of the excluded player. This finding was moderated by social value orientations. Proselfs were not affected by the consequences for the excluded players. Prosocials were less willing to exclude others the more harmful were the consequences of exclusion. Results are related to research on social exclusion, the do-no-harm principle, and social value orientations. PMID- 15272952 TI - The first year: influences on the satisfaction, involvement, and persistence of new community volunteers. AB - This investigation tests an elaborated form of Omoto and Snyder's volunteer process model, which explains how the helping behavior of volunteers is influenced by antecedent factors and by subjective experiences while volunteering. Two-hundred-thirty-eight community volunteers from nine different organizations were recruited at the time of initial orientation and completed measures of personality and motivation. They were contacted at four times during their first year of volunteering and queried regarding their emotional reactions (sympathy, distress), satisfaction, and degree to which their motivations for volunteering were being fulfilled. Consistent with the elaborated model, feelings of sympathy, distress, and motive fulfillment were substantially predicted by antecedent factors, and satisfaction with the volunteer work was substantially predicted by these subjective experiences. Also consistent with the elaborated model, volunteer involvement (hours per week volunteered) was predicted by satisfaction, although volunteer persistence over time was not. Implications of these findings are discussed. PMID- 15272953 TI - "Isn't it fun to get the respect that we're going to deserve?" Narcissism, social rejection, and aggression. AB - Across four studies, narcissists were more angry and aggressive after experiencing a social rejection than were nonnarcissists. In Study 1, narcissism was positively correlated with feelings of anger and negatively correlated with more internalized negative emotions in a self-reported, past episode of social rejection. Study 2 replicated this effect for a concurrent lab manipulation of social rejection. In Study 3, narcissists aggressed more against someone who rejected them (i.e., direct aggression). In Study 4, narcissists were also more aggressive toward an innocent third party after experiencing social rejection (i.e., displaced aggression). Narcissists were not more aggressive after social acceptance. Self-esteem plays little role in predicting aggression in response to rejection. These results suggest that the combination of narcissism and social rejection is a powerful predictor of aggressive behavior. PMID- 15272954 TI - In the privacy of their own homes: using the internet to assess racial bias. AB - Recent studies suggest that research participants show reduced distortion of their taboo attitudes and behaviors when they take part in Internet-based procedures from outside the laboratory. We explored whether such procedures would reduce distortion in the assessment of racial bias. In Study 1, White participants who completed the study in the laboratory evaluated Black targets more favorably than White targets. This unexpected "outgroup-favoring" pattern occurred in both pencil-and-paper and Internet versions of the study, showing that modality did not produce it; but when participants worked outside the laboratory via the Internet, this pattern disappeared. Study 2 replicated the above findings and further indicated that the reduced distortion in Internet based studies was due to the removal of the experimenter rather than removing the participants from the laboratory environment. The implications of these findings for the study of controlled processes of prejudice and the nature of Internet based social communication are discussed. PMID- 15272955 TI - The effects of categorically based expectations on minority influence: the importance of congruence. AB - The role of congruence and incongruence in diverse decision-making groups is examined by manipulating opinion agreement within and between members of different social categories. Congruence occurs when ingroup members agree with one another and outgroup members disagree, whereas incongruence occurs when an ingroup member disagrees with a majority composed of ingroup and outgroup members. The results of two studies, one using a scenario methodology and the second using simulated work teams with two ingroup members and one outgroup member, show that regardless of the task-relevance of salient differences, individuals respond most favorably when categorical and opinion differences are congruent. Study 1 examined individuals' emotional reactions and group efficacy. Study 2 examined group performance, the minority influence process, and efforts to maintain congruence. The findings suggest that outgroup minority opinion holders may be more influential in diverse group decision-making settings than ingroup minority opinion holders. PMID- 15272956 TI - Inductive reasoning and judgment interference: experiments on Simpson's paradox. AB - In a series of experiments on inductive reasoning, participants assessed the relationship between gender, success, and a covariate in a situation akin to Simpson's paradox: Although women were less successful then men according to overall statistics, they actually fared better then men at either of two universities. Understanding trivariate relationships of this kind requires cognitive routines similar to analysis of covariance. Across the first five experiments, however, participants generalized the disadvantage of women at the aggregate level to judgments referring to the different levels of the covariate, even when motivation was high and appropriate mental models were activated. The remaining three experiments demonstrated that Simpson's paradox could be mastered when the salience of the covariate was increased and when the salience of gender was decreased by the inclusion of temporal cues that disambiguate the causal status of the covariate. PMID- 15272957 TI - The weight of obesity in evaluating others: a mere proximity effect. AB - Previous research demonstrates that we tend to derogate individuals who are perceived to be in a social relationship with stigmatized persons. Two experiments examined whether this phenomenon also occurs for individuals seen in the presence of an obese person and whether a social relationship is necessary for stigmatization to spread. The results from both experiments revealed that a male job applicant was rated more negatively when seen with an overweight compared to a normal weight female and that just being in the mere proximity of an overweight woman was enough to trigger stigmatization toward the male applicant. Experiment 2 examined possible moderating effects of the proximity finding. Applicants seated next to heavy (vs. average weight) individuals were denigrated consistently regardless of the perceived depth of the relationship, the participant's anti-fat attitudes or gender, and whether or not positive information was presented concerning the woman. The profound nature of the obesity stigma and implications for impression formation processes are discussed. PMID- 15272958 TI - Immunizing the self: self-concept stabilization through reality-adaptive self definitions. AB - Processes of self-concept immunization are introduced as a way of reconciling self-concept protection against threatening information with the necessity of acknowledging own failure or losses. Self-immunization works by adaptively changing the subjective operationalization of personal traits, such that skills that individuals believe themselves to be good at are conceived as highly diagnostic, whereas skills that persons do not believe they possess are considered less diagnostic. Three studies are presented to investigate this stabilizing process. Correlational as well as experimental and longitudinal data support the assumption that self-immunization stabilizes central and abstract aspects of the self-concept without ignoring negative information on concrete skills. PMID- 15272959 TI - Remembering everyday experience through the prism of self-esteem. AB - Two studies examined whether global self-esteem was associated with bias in memory for autobiographical experience. For 7 days, participants described specific events and made ratings of their experience (i.e., state self-esteem, positive and negative emotion, and perceived valence of the event) in response to each event. Later, participants were presented with their event descriptions and were asked to recall their experience ratings from memory. As hypothesized, higher global self-esteem predicted positive shifts in memory for experience, whereas lower global self-esteem predicted negative shifts in memory for experience. Patterns of bias were strongest for remembered state self-esteem, moderate for positive emotion, and minimal for event valence. Self-esteem did not predict bias for negative emotion. Mood at the time of recall (measured in Study 2) generally did not account for the patterns. These findings strengthen the view that self-esteem is a rich source of knowledge about the self that can influence memory for some kinds of autobiographical experience. PMID- 15272960 TI - The vulnerability of values to attack: inoculation of values and value-relevant attitudes. AB - Based on the values-as-truisms hypothesis and inoculation theory, two experiments tested whether providing cognitive defenses for the value of equality induces resistance against a message attacking this value. Experiment 1 found that participants who generated cognitive support in an active-supportive or an active refutational defense were less persuaded by a subsequent message attacking equality than were participants who engaged in no prior defense. Experiment 2 examined the effects of an active-refutational defense and a passive-refutational defense, which simply asked participants to read reasons supporting or opposing equality. Results indicated additive effects of the active and passive defenses, such that participants were most resistant to the anti-equality message when they were given both defenses. Mediational analysis across both experiments revealed that the defenses increased counterargumentation of the anti-equality message, which led to increased post-attack importance of equality and predicted more favorable equality-relevant attitudes and values. PMID- 15272961 TI - Watching your troubles away: television viewing as a stimulus for subjective self awareness. AB - Three studies explored the role of television viewing in eliciting subjective self-awareness and positive self-feelings. Study 1 assessed the effects of self awareness manipulations via exposure to a neutral television program on actual ideal discrepancies. Those who watched television showed significantly smaller self-discrepancies than those who did not, independent of mood. Study 2 demonstrated the ecological validity of this finding by replicating it with people watching television in their own homes. Study 3 investigated whether manipulations of self-feelings affected television watching. Results indicated that those who received failure feedback watched television longer than those in a control condition who likewise watched television longer than those who received success feedback. Television appears to be an effective stimulus to direct the focus away from oneself and to render people less aware of how they are falling short of their standards. PMID- 15272962 TI - The effects of mindset on behavior: self-regulation in deliberative and implemental frames of mind. AB - The effects of deliberative and implemental mindsets--cognitive and motivational states associated with predecisional and postdecisional frames of mind, respectively--were examined in the context of the self-regulation of behavior. Participants who had been induced to deliberate the merits of participating in a specified task formulated more pessimistic expectations about this task than did participants who had been induced to imagine implementing a plan to complete the task. Moreover, participants in the deliberation condition underperformed relative to the participants in the implemental condition, demonstrating that deliberative and implemental thinking can influence behavior as well as cognition. PMID- 15272963 TI - Can't quite commit: rumination and uncertainty. AB - Why do some individuals persist in self-destructive rumination? Two studies investigated the relation between a ruminative response style and the reluctance to initiate instrumental behavior. In Study 1, ruminators were compared to nonruminators regarding their evaluation of a self-generated plan to revise their university housing system and, in Study 2, concerning their plan to redesign the undergraduate curriculum. In both studies, on relevant composite measures, ruminators expressed less satisfaction and confidence with regard to their plans than did nonruminators. They were also less likely to commit to the plans they generated. The findings suggest that in addition to its documented detrimental effects on thinking and problem solving, self-focused rumination may inhibit instrumental behavior by increasing uncertainty, resulting in further rumination and behavioral paralysis. PMID- 15272964 TI - Perceptions of trait typicality in gender stereotypes: examining the role of attribution and categorization processes. AB - Gender stereotypes are understood as the ascription of different personality traits to men and women. Data from American and Italian samples showed that consistent with the attribution hypothesis, the estimated prevalence of a trait in a target group predicted perceptions of trait typicality well. In contrast, there was no support for the categorization hypothesis, according to which perceived differences in trait prevalence between groups should independently predict trait typicality. Nevertheless, participants overestimated gender differences in personality as predicted by the principle of intercategory accentuation. The implications of these findings for the rationality and accuracy of gender stereotyping are discussed. PMID- 15272965 TI - White guilt and racial compensation: the benefits and limits of self-focus. AB - In two studies, the authors investigated guilt as a response to group-based advantage. Consistent with its conceptualization as a self-focused emotion, White guilt was based in self-focused beliefs in racial inequality. Thus, guilt was associated with belief in White privilege (Study 1) and resulted from seeing European Americans as perpetrators of racial discrimination (Study 2). Just as personal guilt is associated with efforts at restitution, White guilt was predictive of support for affirmative action programs aimed at compensating African Americans. White guilt was not, however, predictive of support for noncompensatory efforts at promoting equality, such as affirmative action programs that increase opportunities (Study 2). In contrast, the other-focused emotion of group-based sympathy was a more general predictor of support for different affirmative action policies. Our findings demonstrate the benefits and limits of group-based guilt as a basis of support for social equality and highlight the value of understanding the specific emotions elicited in intergroup contexts. PMID- 15272966 TI - Predicting the paths of peripherals: the interaction of identification and future possibilities. AB - Two studies investigated how both degree of identification and the individual's position within the group influence aspects of group loyalty. The authors considered ingroup position in terms of both the individual's current position within a group and expectations concerning the likelihood that one's position might change in the future. Peripheral group members learned that their acceptance by other group members would improve in the future or that they could expect rejection by other group members. Various indices of group loyalty (ingroup homogeneity, motivation to work for the group, and evaluation of a motivated group member) showed that when group members anticipated future rejection, the lower the identification the less loyal they were. In contrast, those who expected future acceptance were more loyal (more motivated to work for the group) the lower their identification. Current group behavior depends on both intragroup future expectations and level of identification. PMID- 15272967 TI - Synergistic person x situation interaction in distributive justice behavior. AB - A person x situation interaction is synergistic when a personality trait amplifies the effect of a situational factor. The present study tested how individuals' justice attitudes and situational factors jointly affect the allocation of financial burdens. Six insurance cases were described to 80 participants. Economic status of client (high, low) and responsibility of client for damage of the insured (high, low) were manipulated between subjects. Participants suggested a percentage of the total costs that they considered a fair contribution by the client. In accordance with the synergistic model, justice attitude (person factor) and responsibility for damage (situation factor) interacted and explained 5% of the variance of the dependent variable. With increasing negativity of attitude toward equality, the effect of responsibility was larger. Several cognitive mechanisms, such as motivated perception, selective attention, and the availability of attitude congruent situation schemas, that may account for synergistic interactions in justice behavior and in other domains were discussed. PMID- 15272968 TI - Using questionnaires in qualitative interviews. PMID- 15272969 TI - Weapons of mass seduction. PMID- 15272970 TI - Soft governance and attitudes to clinical quality in English general practice. AB - OBJECTIVES: English primary care organisations (primary care groups and trusts - PCGs, PCTs) were, and are, responsible for the quality of general practice but lack hierarchical structures and, frequently, contractual relationships through which to influence it. The theory of soft governance describes how managers can influence professional practice by other means. This study examines the hypothesis that PCG/Ts have used 'soft' clinical governance. METHODS: Survey in 2000/01 of general practitioners' (GPs') attitudes, opinions and self-reported activity in six PCGs and six PCTs using a semi-structured mailed questionnaire. To assess how representative respondents were of English GPs generally, four questions from a national sample survey of English GPs were included and the results compared. RESULTS: Responses were obtained from 437 (52%) GPs. They most often mentioned the technical aspects of clinical governance. Managerial, policy and resourcing implications were next most frequently mentioned, usually in unfavourable terms. Most GPs reported that their clinical practice had changed because of clinical governance activities, although nearly 40% also reported little difference in the quality of care provided. The National Service Framework for coronary heart disease influenced practice independently of PCG/T activities. CONCLUSION: English primary care organisations are exercising soft governance (although not by that name) over some but not all aspects of GPs' clinical practice. However, this soft governance is complex, not easy to sustain and appears hard to extend beyond essentially clinical domains. PMID- 15272971 TI - 'Questerviews': using questionnaires in qualitative interviews as a method of integrating qualitative and quantitative health services research. AB - OBJECTIVES: Multi-method approaches are increasingly advocated in health services research (HSR). This paper examines the use of standardised self-completion questionnaires and questions, during in-depth interviews, a technique termed 'questerviews'. METHODS: 'Questerview' techniques were used in four empirical studies of health perceptions conducted by the authors. The studies included both standardised self-completion questions or questionnaires and in-depth interviews. Respondents were tape-recorded while they completed the standardised questionnaires and were encouraged to discuss their definitions of terms and responses to items in-depth. In all studies, 'questerviews' were fully transcribed and data analysis involved the scrutinising of transcripts to identify emergent themes. RESULTS: Responses to the standardised items led to rich sources of qualitative data. They proved to be useful triggers as respondents discussed their understanding and definitions of terms, often explaining their responses with stories from their own experiences. The items triggered detailed exploration of the complex factors that comprise health, illness and healthcare seeking, and gave considerable insight into the ways in which people respond to standardised questions. Apparently simple questions and response categories conceal considerable complexity. CONCLUSIONS: Inclusion of standardised survey questions in qualitative interviews can provide an easy and fruitful method to explore research issues and provide triggers to difficult or contested topics. Well designed and validated questionnaires produce data of immense value to HSR, and this value could be further enhanced by their use within a qualitative interview. We suggest that the technique of 'questerviews' is a tangible and pragmatic way of doing this. PMID- 15272972 TI - Evidence-based priority-setting: what do the decision-makers think? AB - OBJECTIVES: Resource scarcity dictates the need for health organisations to set priorities. Although such activity should be based, at least in part, on evidence, there are limited examples in the literature of decision-makers reflecting on their use of evidence in priority-setting. METHODS: A participatory action-research project was conducted in a single health authority in Alberta. It included in-depth interviews and focus groups with senior decision-makers both before and after development and implementation of a macro-level priority-setting framework (programme budgeting and marginal analysis, PBMA). Data were thematically coded and information on the use of evidence in priority-setting is reported. RESULTS: Barriers to the use of evidence in priority-setting identified by decision-makers included crisis-orientated management, time constraints and a lack of skills. Decision-makers suggested using a mix of 'soft' and 'hard' forms of evidence in priority-setting. Following PBMA implementation, decision-makers wanted better information on capacity to benefit, but preferred to do this pragmatically from multiple sources of information rather than using a single metric. CONCLUSION: In examining the perspectives of decision-makers in using evidence to support priority-setting, valuable information was derived which should provide insight for such processes in other jurisdictions. The main finding of a desire for pragmatic assessment of benefit is informative for those involved in both decision-making and research. PMID- 15272973 TI - Influencing prescribing in English primary care: the views of primary care organisations. AB - OBJECTIVE: The rapid rise of prescribing expenditure is a concern in many industrialised countries and methods to manage medicines are widely employed. The purpose of this study was to identify the approaches to improve primary care prescribing by primary care organisations (PCOs) in the National Health Service (NHS) in England. METHODS: A questionnaire (Management of Medicines, MANMED) was mailed to prescribing advisers and prescribing leads in 332 PCOs. RESULTS: A response rate of 66% (220/332) was achieved. Most PCOs report the improvement of the quality of prescribing as their top priority, followed by budget adherence at both practice and PCO levels. Prescribing advisers typically offer several forms of support: practice visits, prescribing reviews, indicators of prescribing, prescribing newsletters, hands-on support, seminars and local formularies. PCOs are pursuing a wide range of prescribing initiatives, covering, on average, seven different therapeutic areas. National targets are the main driver for prescribing initiatives but other key influences include inappropriate prescribing and clinical governance. Although cost considerations are important, improving the quality of prescribing is perceived as the overriding principle on which PCO prescribing strategy is based. Multifaceted prescribing support is widespread and national targets are the largest single factor influencing choice of therapeutic area for prescribing initiatives. CONCLUSIONS: Diversity in approaches presents the opportunity to improve the evidence base for medicines management. Not only could such research inform PCOs in their central aim of improving the quality of prescribing within the NHS, but it may also offer insights of relevance to other countries if the influence of process and context upon the effectiveness of medicines management is systematically explored. PMID- 15272974 TI - In the absence of evidence, who chooses? A qualitative study of patients' needs after treatment for colorectal cancer. AB - OBJECTIVES: To identify the range of patient pathways following surgery for colorectal cancer and explore patients' needs and preferences for follow-up. METHODS: A survey of hospitals within the UK Colorectal Cancer Services Collaborative and qualitative thematic analysis of 39 in-depth narrative interviews with colorectal cancer patients. Participants volunteered or were contacted through hospital consultants, support groups and general practitioners (GPs). Most of the interviews were collected in respondents' homes, throughout the UK. RESULTS: Thirty-five (70%) hospitals supplied details of their follow-up regime. There was a wide variation: only three hospitals specifically stated that patients were given a choice about the type of follow-up. The patients' interviews highlighted their need for a responsive GP and realistic information about recovery, resources and diet. Choice is particularly important because patients differ in their views of the benefits of hospital follow-up. CONCLUSIONS: The absence of evidence about what constitutes ideal clinical follow up for colorectal cancer is reflected in current hospital practice. In such circumstances, the preferences of individual patients are particularly important. Not all patients want repeated specialist investigations but those without stomas, and therefore no access to a stoma nurse, need another source of advice about recovery and long-term practical help. Follow-up care organised by GPs may be acceptable to many patients. We suggest a list of topics for GPs to discuss with their patients about follow-up. The needs described by patients are not extensive and could often be met by existing resources. PMID- 15272975 TI - Are data on the uptake of disability benefits a useful addition to census data in describing population health care needs? AB - OBJECTIVE: To determine if administrative data relating to the uptake of disability benefits held by government departments in Northern Ireland are fair and unbiased indicators of relative need for health care and therefore a useful addition to census data. METHOD: This is an ecological study of the 566 electoral wards in Northern Ireland. The variation in uptake of a health-related benefit (Disability Living Allowance) was regressed against three other indicators of health (limiting long-term illness, mortality and the numbers of patients admitted to hospital). All the indicators had been indirectly standardised by age and sex. The unstandardised residuals from this model were regressed against the social and geographical factors, namely area deprivation score, religious denominational composition and urban/rural dwelling to see if they influenced benefit uptake above and beyond any association with ill health. RESULTS: The health factors alone explained 77.2% of the variation in benefit uptake, with limiting long-term illness being the major determinant; however, even after controlling for differences in health status, benefit uptake was shown to be higher in more urban areas and in areas that had a greater proportion of Catholics. Area deprivation was not associated with benefit uptake once health differences had been controlled for. CONCLUSION: Administrative data on disability benefits can undoubtedly provide useful additional information for describing the levels of relative disadvantage or ill health of areas. However, because they also reflect variations in uptake, which appears to be confounded by social and geographical factors, we would urge caution when they are used to identify priority areas or to allocate resources. PMID- 15272976 TI - Issues arising from the use of qualitative methods in health economics. AB - As health economists begin to embrace qualitative methodology they inevitably face a number of issues. This paper explores these, distinguishing between those associated with the conflict between quantitative and qualitative methodologies (that has already been faced in a number of other research areas) and those associated with the potential for challenges to the discipline of mainstream economics. The former include both the acceptability of the methods (because of issues such as generalisability and reflexivity) and the acceptability of presentation. The latter appear to be essentially concerned with identity within economics. The paper concludes by noting the positive aspects of conducting qualitative research in health economics: the interesting and motivating nature of the research and, particularly, the possibilities for increasing the relevance associated with economic theory in the context of health and health services. PMID- 15272977 TI - Interdisciplinarity in health services research: dreams and nightmares, maladies and remedies. AB - Interdisciplinarity has become popular in health services research. Advocates suggest that interdisciplinary approaches may produce more accessible, applicable, exciting and realistic knowledge than traditional disciplinary approaches. To date, there has been surprisingly little analysis of the institutional and intellectual demands of interdisciplinarity as a methodology or practice. This paper (1) identifies some basic intellectual and institutional features of interdisciplinary research, (2) describes typical interdisciplinary 'dreams' and corresponding 'nightmares' that researchers might encounter in practice, (3) highlights maladies of interdisciplinary research careers and suggests practical remedies, and (4) discusses implications for health research policy. Individual researchers can avoid pitfalls of interdisciplinarity through strategies that include selective collaboration, cross-training, sustained relationships, good humour, participation in peer review, declaring the place of one's work, and balancing dissemination of research between peer and other audiences. Interdisciplinary activities span institutional boundaries and make novel demands on academic resources and allegiances. Research organizations can improve their hospitality to interdisciplinary work by encouraging straightforward communication, recognising interdisciplinary productivity, making allowances for the higher time and energy costs of interdisciplinary liaisons, and providing the necessary institutional support and stability to cultivate projects to fruition. Alongside the creation of large new interdisciplinary networks and organizations, we should invest in the highly valuable contributions of small and enduring interdisciplinary teams, modest interdisciplinary stretches and evolving interdisciplinary creatures. PMID- 15272978 TI - The condition of the working class in England. PMID- 15272979 TI - What will the new genetic information do for us? AB - The usefulness of new genetic information is frequently assumed, but depends crucially on asking the right research questions. Successful applications of the new knowledge will require effective integration with the ideas, concerns and expectations not only of patients and their families, but also health professionals in primary care. The answers to some of the most exciting questions, concerning new taxonomies of health and disease, are to be found not in teaching hospitals or academic centres, but in the population at large. New types of partnership are required between colleagues in biomedicine, epidemiology, social science and primary care. The greatest challenge, however, is to our social imagination. Can we develop the sustained relationships with families and communities, based on goodwill, mutuality and trust, on which the whole enterprise depends? PMID- 15272980 TI - Ethics of health care and research. PMID- 15272981 TI - An ounce of prevention buys a pound of cure. PMID- 15272982 TI - [Genotoxicity from exposure to cigarettes in young smokers in Colombia]. AB - OBJECTIVE: To evaluate the frequency of chromosome aberrations (CAs) in the peripheral blood lymphocytes of young cigarette smokers in the city of Popayan, Colombia. METHODS: In this cytogenetic case-control study there were 32 young cigarette smokers and 32 nonsmokers. All of them were between 19 and 29 years old and none used psychoactive drugs, suffered from chronic or infectious diseases, or had been exposed to chemotherapy or radiation therapy or to chemical agents in their work. A survey was used to obtain demographic information, occupational information (type of employment, type of and length of exposure to chemical agents), lifestyle information (consumption of alcoholic beverages and psychoactive drugs), and information on smoking (current or former smoker, number of cigarettes smoked daily, length of time smoking, and type of cigarettes smoked). The cases were matched with the controls by age (+/- 5 years) and sex. The microscopic study of the CAs using lymphocyte cultures was carried out under the light microscope with 100X magnification. For each study participant, 100 complete metaphase cells (2n = 46 chromosomes) were analyzed, counting the structural CAs (chromatid breaks and chromosome breaks) and numerical CAs (change in the number of chromosomes). The frequency of CAs was adjusted for alcohol consumption, using a univariate linear model. RESULTS: The frequency of total CAs was significantly greater in the young cigarettes smokers (6.02 +/- 0.52) than in the nonsmokers (3.04 +/- 0.50), and the greatest number of CAs (7.77 +/- 0.88) was found in those who had a pack-year value of more than 3.0. In addition, there was a dose-effect relationship, shown by the increase in the frequency of CAs with an increase in the pack-years of consumption (coefficient of determination = 0.2257). CONCLUSION: We confirmed the association between cigarette consumption and CAs in young people who smoked relatively little. These results should be taken into account in order to formulate national smoking prevention policies and to evaluate their outcomes, from both the social, economic, and environmental standpoint and the standpoint of the health of future generations. PMID- 15272983 TI - The use of the female condom by women in Brazil participating in HIV prevention education sessions. AB - OBJECTIVES: To study HIV-positive women and women at risk of becoming infected with HIV who attended HIV prevention education group sessions at a university hospital in Brazil and to compare the use of the female condom and the male condom by these two groups of women. METHODS: The study subjects were 165 women participating in HIV prevention education group sessions at the Medical School Hospital of Ribeirao Preto of the University of Sao Paulo, in the city of Ribeirao Preto, Sao Paulo, Brazil. Women could be enrolled in the study from August 2000 to June 2001, and the follow-up observation time period was from August 2000 to July 2001. Male condoms and female condoms were freely distributed to all the participants at the end of each educational session and also at the end of each follow-up visit that the participants made. Each woman took part in an initial interview and was asked to return monthly. At each follow-up visit an additional short interview was carried out in order to investigate use of the male condom and of the female condom. Variables that were examined for the study included age, education, ethnic group, marital or relationship status, number of children, the women's use of male condoms and female condoms, commercial sex (whether the women had ever had sex in exchange for money, gifts, or favors), and previous knowledge of the female condom. RESULTS: The 165 women studied fell into the following three categories: 132 of them (80.0%) were HIV-positive, 26 of them (15.8%) had a sexually transmitted disease (STD) other than HIV and did not have an HIV-positive partner, and 7 of them (4.2%) had an HIV-positive partner but did not have HIV or any other STD. The women ranged in age from 15 to 64 years, with a mean of 30.3 years. Of the women in the study, 69.7% of them were married or were cohabitating, and 90.9% of them had a sexual partner. Just over two-thirds of the women had seven years of formal schooling or less. Out of 163 women, a total of 31 of them (19.0%) had never used the male condom with a partner, and 49 of the 163 (30.1%) had not used a male condom at the time of the last sexual intercourse. Out of the 165 women, 74 of them (44.8%) returned for at least one follow-up visit. Of these 74 women, 58 of them (78.3%) reported using the female condom between the initial interview and the first follow-up visit. The majority of the 74 women who returned for a visit liked using the female condom, and the women reported that their partners also generally accepted the female condom. In comparison to women at risk of HIV, HIV-positive women were more likely to have used the male condom with a partner before the initial interview. Women who continued returning over a longer follow-up period were more likely to have used the female condom at the time of the last sexual intercourse. No association was found between female condom use at the time of last sexual intercourse and the woman's HIV infection status. CONCLUSIONS: In comparison to the women at risk of HIV, the HIV-positive women in our study were more likely to use male condoms with their partners, to return for follow-up visits, and to have a longer follow up period. The acceptance of the female condom among the HIV-positive women in this study, as reported at their first follow-up visit, appears to be similar to the acceptance of the female condom among women in general in Brazil. PMID- 15272984 TI - [Recommendation of the Brazilian Society of Pediatrics for antibiotic therapy in children and adolescents with community-acquired pneumonia]. AB - OBJECTIVE: To standardize the use of antibiotics to treat community-acquired pneumonia in children and adolescents in Brazil. METHODS: The following data sources were utilized: the Medline and LILACS (Literatura Latino-Americana e do Caribe em Ciencias da Saude) bibliographic databases; World Health Organization and Pan American Health Organization documents published between 1980 and 2002; Internet materials in Portuguese, Spanish, or English located using the search terms "pneumonia, child, adolescent, etiology, treatment"; and interviews with renowned experts in the field. RESULTS: Hospitalization is always required for children younger than 2 months of age. That age group should be treated with ampicillin and aminoglycosides or third-generation cephalosporins. Children older than 2 months must be hospitalized if there is severe pneumonia. Tachypnea should be used as the criterion to distinguish between acute respiratory infection and pneumonia. Pneumonia is considered to be very severe when there are seizures, sleepiness, stridor at rest, severe malnutrition, no ingestion of fluids, or signs of respiratory failure such as central cyanosis. Children who are 2 months of age or older may receive outpatient treatment with amoxicillin or penicillin G procaine. In the case of inpatient treatment, crystalline penicillin or ampicillin may be used for severe cases, and oxacillin and chloramphenicol or ceftriaxone for very severe cases. A macrolide, preferably erythromycin, should be employed when the etiologic agent is suspected to be Chlamydia trachomatis, C. pneumoniae, Mycoplasma pneumoniae, or Bordetella pertussis. CONCLUSIONS: The diagnosis of pneumonia and the need for hospitalization can be based on clinical assessment. The main antibiotics to be used are amoxicillin, penicillin, erythromycin, ampicillin, oxacillin, chloramphenicol, ceftriaxone, and aminoglycosides, depending on the age of the patient and the severity of the disease. PMID- 15272985 TI - An ecological analysis of childhood-onset type 1 diabetes incidence and prevalence in Latin America. AB - OBJECTIVE: To explore, for Latin America, by means of an ecological correlation analysis, the possible relationships between both the incidence and prevalence of childhood type 1 diabetes and selected hypothesized etiological factors, including race/ethnicity, geographical latitude, breastfeeding rates, per capita milk supply and coffee consumption, and wealth-related indicators such as infant mortality rate, life expectancy at birth, and national human development index. METHODS: Recently published data on incidence and prevalence of type 1 diabetes in children < or = 14 years of age in Latin American countries were utilized. Risk indicators were selected based on existing genetic and environmental hypotheses. Indicator data were obtained from publicly available resources. Spearman correlation coefficients were used to measure the association between both incidence and prevalence of type 1 diabetes and the selected indicators. RESULTS: A strong negative correlation was found between the proportion of Amerindians in a country's population and both incidence (r = -0.75; P = 0.008) and estimated prevalence (r = -0.78; P < 0.0001) of childhood type 1 diabetes. The per capita supply of milk showed a strong positive correlation with both incidence (r = 0.70; P = 0.025) and prevalence (r = 0.55; P = 0.018). Wealth related indicators correlated with prevalence but not with incidence. CONCLUSIONS: Our results suggest that the presence of the Amerindian component of the population in Latin America provides protection against childhood-onset type 1 diabetes. Our results also confirm the association previously reported in other countries and regions of the world of type 1 diabetes and milk consumption. Further studies are needed to develop and test potential genetic and environmental hypotheses that could help to better understand the interplay between genetic susceptibility and environment in type 1 diabetes across different ethnic groups. PMID- 15272986 TI - [Prevalence of tobacco use among pediatric residents in Argentina]. AB - OBJECTIVES: To determine the prevalence of smoking among pediatric residents in Argentina, to evaluate the risk factors associated with that habit, and to analyze the attitudes of these professionals with regard to tobacco use by their patients. METHODS: A cross-sectional study was conducted using anonymous self administered questionnaire surveys. The surveys were used in May 2002 with pediatric residents in eight hospitals in five provinces of Argentina: Buenos Aires, Cordoba, La Plata, Mendoza, and Neuquen. The study variables were: sex, age, the year of residence, the number of times per week on night duty, if the resident's mother or father smoked, the age at which began to smoke, the place and the hospital activities in which most often smoked, if the immediate supervisor was a smoker, if had increased tobacco use after becoming a resident, attitude toward tobacco use by patients and their parents, and knowledge concerning the risks of smoking. We calculated the frequencies of the studied variables and the odds ratios (ORs) and their 95% confidence intervals (CIs). Multiple logistic regression was used in a model with all the possible predictive variables. The level of significance was P < 0.05. RESULTS: A total of 349 responses were obtained (98.8% of the residents present at the time of the survey). The prevalence of smokers among the surveyed pediatric residents was 22.2%. Among the smokers, 38.9% of them said they smoked more since becoming a resident, and 63.9% of them said that night duty was the hospital activity in which they most often smoked. After adjusting for the other variables, the remaining risk factors for smoking were having a mother who smoked (OR: 2.7; 95% CI: 1.57 to 4.84) and living alone (OR: 3.15; 95% CI: 1.58 to 6.26). Out of all the residents (both smokers and nonsmokers), only 26.5% of them said that they explained to their patients the risks of tobacco use, and only 23.2% of them suggested quitting smoking or not beginning; there were no statistically significant differences between the smoker residents and the nonsmoker residents. CONCLUSIONS: The prevalence of smoking among the pediatric residents is high and is close to the level found in other Argentine physicians. The factors associated with smoking were having a mother who smoked and living alone. The residents should take a more active role with patients or relatives of patients who smoke. Activities need to be put into place that improve the level of training on this subject in medical school and during residency. PMID- 15272987 TI - The PACARDO research project: youthful drug involvement in Central America and the Dominican Republic. AB - OBJECTIVE: To estimate the occurrence and school-level clustering of drug involvement among school-attending adolescent youths in each of seven countries in Latin America, drawing upon evidence from the PACARDO research project, a multinational collaborative epidemiological research study. METHODS: During 1999 2000, anonymous self-administered questionnaires on drug involvement and related behaviors were administered to a cross-sectional, nationally representative sample that included a total of 12,797 students in the following seven countries: Costa Rica (n = 1,702), the Dominican Republic (n = 2,023), El Salvador (n = 1,628), Guatemala (n = 2,530), Honduras (n = 1,752), Nicaragua (n = , 419), and Panama (n = 1,743). (The PACARDO name concatenates PA for Panama, CA for Centroamerica, and RDO for Republica Dominicana). Estimates for exposure opportunity and actual use of alcohol, tobacco, inhalants, marijuana, cocaine (crack/coca paste), amphetamines and methamphetamines, tranquilizers, ecstasy, and heroin were assessed via responses about questions on age of first chance to try each drug, and first use. Logistic regression models accounting for the complex survey design were used to estimate the associations of interest. RESULTS: Cumulative occurrence estimates for alcohol, tobacco, inhalants, marijuana, and illegal drug use for the overall sample were, respectively: 52%, 29%, 5%, 4%, and 5%. In comparison to females, males were more likely to use alcohol, tobacco, inhalants, marijuana, and illegal drugs; the odds ratio estimates were 1.3, 2.1, 1.6, 4.1, and 3.2, respectively. School-level clustering was noted in all countries for alcohol and tobacco use; it was also noted in Costa Rica, El Salvador, Guatemala, and Panama for illegal drug use. CONCLUSIONS: This report sheds new light on adolescent drug experiences in Panama, the five Spanish-heritage countries of Central America, and the Dominican Republic, and presents the first estimates of school-level clustering of youthful drug involvement in these seven countries. Placed in relation to school survey findings from North America and Europe, these estimates indicate lower levels of drug involvement in these seven countries of the Americas. For example, in the United States of America 70% of surveyed youths had tried alcohol and 59% had smoked tobacco. By comparison, in these seven countries, only 51% have tried alcohol and only 29% have smoked tobacco. Future research will help to clarify explanations for the observed variations across different countries of the world. In the meantime, strengthening of school-based and other prevention efforts in the seven-country PACARDO area may help these countries slow the spread of youthful drug involvement, reduce school-level clustering, and avoid the periodic epidemics of illegal drug use that have been experienced in North America. PMID- 15272988 TI - Crime in Trinidad and Tobago: the effect of alcohol use and unemployment. AB - OBJECTIVE: The purpose of this study was to determine whether unemployment and alcohol consumption were associated with different types of crime in Trinidad and Tobago. METHODS: This study made use of secondary data from the Central Statistical Office of Trinidad and Tobago for the period 1990-1997. Pearson product moment correlations and stepwise multiple regression analysis were used to identify significant predictors of crime. RESULTS: Unemployment accounted for 69.2% of the variance for serious crimes. Beer available for home consumption explained 64% of the variance for minor offenses, and both unemployment and beer available for home consumption accounted for 92.2% of the variance for minor crimes. CONCLUSIONS: This study provides information that is potentially useful for developing public policies for unemployment and for the sale of beer for home consumption, both of which are associated with crime in Trinidad and Tobago. Reductions in beer available for home consumption-a major public health concern would significantly reduce the occurrence of minor offenses in this country. Further research is needed on the relationship between unemployment and crime. PMID- 15272989 TI - [Health, equity, and the Millennium Development Goals]. AB - In September 2000 representatives of 189 countries met for the Millennium Summit, which the United Nations convened in New York City, and adopted the declaration that provided the basis for formulating the Millennium Development Goals (MDGs). The eight goals are part of a long series of initiatives that governments, the United Nations system, and international financial institutions have undertaken to reduce world poverty. Three of the eight goals deal with health, so the health sector will be responsible for implementing, monitoring, and evaluating measures proposed to meet targets that have been formulated: to reduce by two-thirds the mortality rate in children under 5 years of age between 1990 and 2015; to reduce by three-quarters the maternal mortality rate between 1990 and 2015; and to halt and begin to reverse the spread of HIV/AIDS by the year 2015, as well as to halt and begin to reverse the incidence of malaria, tuberculosis, and other major diseases. The health sector must also work with other parties to achieve targets connected with two other of the goals: to improve access to affordable essential drugs, and to reduce the proportion of persons who do not have safe drinking water. Adopting a strategy focused on the most vulnerable groups-ones concentrated in locations and populations with the greatest social exclusion would make possible the largest total reduction in deaths among children, thus reaching the proposed target as well as producing greater equity. In the Region of the Americas the principal challenges in meeting the MDGs are: improving and harmonizing health information systems; designing health programs related to the MDGs that bring together the set of services and interventions that have the greatest impact, according to the special characteristics of the populations who are intended to be the beneficiaries; strengthening the political will to support the MDGs; and guaranteeing funding for the measures undertaken to attain the MDGs. PMID- 15272990 TI - Perceptions of a fluid consensus: uniqueness bias, false consensus, false polarization, and pluralistic ignorance in a water conservation crisis. AB - A 5-day field study (N = 415) during and right after a shower ban demonstrated multifaceted social projection and the tendency to draw personality inferences from simple behavior in a time of drastic consensus change. Bathers thought showering was more prevalent than did non-bathers (false consensus) and respondents consistently underestimated the prevalence of the desirable and common behavior--be it not showering during the shower ban or showering after the ban (uniqueness bias). Participants thought that bathers and non-bathers during the ban differed greatly in their general concern for the community, but self reports demonstrated that this gap was illusory (false polarization). Finally, bathers thought other bathers cared less than they did, whereas non-bathers thought other non-bathers cared more than they did (pluralistic ignorance). The study captures the many biases at work in social perception in a time of social change. PMID- 15272991 TI - Two functions of verbal intergroup discrimination: identity and instrumental motives as a result of group identification and threat. AB - In two studies, the authors examined the circumstances under which discrimination has an identity confirmation function or an instrumental function (instigating collective action). In Study 1, participants (N = 601) described a situation in which they had discriminated and then completed measures of functionality, group identification, and group threat. Both functions were predicted by group identification, whereas the instrumental function (but not identity confirmation) operated under group threat. In Study 2, "die-hard" soccer fans (N = 1,546) suggested soccer chants in reaction to either a group-reinforcing (own team scores) or group-threatening (other team scores) situation and rated the perceived functionality of the song. Although both of these conditions evoked discriminating songs, as predicted, these served a more identity-confirming function in the reinforcing situation but a more instrumental function (pepping up the team) in the threat situation. Results are discussed in terms of a contextual-functional model of intergroup discrimination. PMID- 15272992 TI - Solitude experiences: varieties, settings, and individual differences. AB - Solitude may be positive or negative, depending on situational and personal factors. From prior research, nine types of solitude were identified. Based on data from a questionnaire study of undergraduate participants, factor analysis suggests that these nine types can be reduced to three dimensions, two positive and one negative. These are, respectively, Inner-Directed Solitude (characterized by self-discovery and inner peace), Outer-Directed Solitude (characterized by intimacy and spirituality), and Loneliness. Personality and value correlates, as well as situational correlates, of the various types of solitude also were explored. PMID- 15272993 TI - Discrimination and the positive-negative asymmetry effect: ideological and normative processes. AB - Research using the minimal group paradigm demonstrates that categorization and ingroup identification can foster intergroup discrimination. However, the positive-negative asymmetry effect shows that less discrimination occurs when negative rather than positive outcomes are distributed. The normative hypothesis explains this asymmetry by the stronger inappropriateness of discrimination in negative than in positive outcome distributions. Results obtained in this minimal group paradigm study (N = 257) did not replicate the asymmetry effect: discrimination occurred in both positive and negative outcome distributions, even if norms against discrimination were stronger in negative than in positive outcome distributions. The absence of the asymmetry effect is explained by the effect of the discrimination-justifying ideology. PMID- 15272994 TI - Variation in black anti-white bias and target distancing cues: factors that influence perceptions of "ambiguously racist" behavior. AB - Experiment 1 indicated that when the White supervisor's negative treatment of a Black subordinate was unconstrained, participant race had no impact on attributions. Conversely, when the treatment was constrained, Black participants reported greater racist attributions than did White participants. Experiment 2 indicated that when the supervisor reported no response or a minimal negative response (i.e., indicating that he did not support his actions) after his negative treatment of the Black subordinate, Black participants reported greater racist attributions than did White participants. Conversely, when the supervisor's negative treatment was followed by a more extreme negative response, participant race had no impact on attributions. Experiment 3 indicated that Black participants were less likely than White participants to perceive a minimal negative response as reflecting a White supervisor's lack of support for his negative actions. Conversely, participant race had no impact on attributions of a Black supervisor's negative actions. PMID- 15272995 TI - The invalidity of disclaimers about the effects of social feedback on self esteem. AB - Despite the fact that several theories suggest that people's self-esteem is affected by social approval and disapproval, many individuals steadfastly maintain that how other people regard them has no effect on how they feel about themselves. To examine the validity of these beliefs, two experiments compared the effects of social approval and disapproval on participants who had indicated either that their self-esteem is affected by how other people evaluate them or that their self-esteem is unaffected by interpersonal evaluation. Results of both studies converged to show that approval and disapproval clearly affected the self esteem of even those individuals who denied that social evaluations affected their feelings about themselves. PMID- 15272996 TI - Fear of the dark: interactive effects of beliefs about danger and ambient darkness on ethnic stereotypes. AB - Two studies examined effects of ambient darkness and chronic beliefs about danger on activation of stereotypes about Blacks. Chronic beliefs were measured by a Belief in a Dangerous World (BDW) questionnaire. In Study, 1, participants in either a dimly lit or dark room saw photos of Black men and rated the extent to which specific traits described the cultural stereotype of Blacks. In Study 2, participants in either a well-lit or dark room completed reaction-time tasks assessing implicit associations between Blacks and evaluative attributes. Separate measures assessed stereotypes connoting danger versus those that are merely derogatory. Results revealed BDW x Darkness interactions on activation of danger-relevant stereotypes: BDW positively predicted activation in dark but not in light conditions. It appears that chronic beliefs about danger can facilitate activation of functionally relevant stereotypes, but this effect occurs primarily under circumstances (such as darkness) that heuristically suggest vulnerability to harm. Conceptual implications are discussed. PMID- 15272997 TI - Fear appeals motivate acceptance of action recommendations: evidence for a positive bias in the processing of persuasive messages. AB - Three experiments are reported that tested the hypothesis that the use of fear appeals in health persuasion may lead to positively biased systematic processing of a subsequent action recommendation aimed at reducing the health threat and, consequently, to more persuasion, regardless of the quality of the arguments in the recommendation. The levels of participants' vulnerability to as well as the severity of a health risk were varied independently, followed by a manipulation of the quality of the arguments in the subsequent action recommendation. The dependent variables included measures of persuasion (attitude, intention, and action), negative affect, and cognitive responses. The results show that participants who felt vulnerable to the health threat were more persuaded, experienced more negative emotions, and had more favorable cognitive responses. Both negative emotions concerning one's vulnerability and positive thoughts concerning the recommendation mediated the effects of vulnerability on persuasion. PMID- 15272998 TI - Lying words: predicting deception from linguistic styles. AB - Telling lies often requires creating a story about an experience or attitude that does not exist. As a result, false stories may be qualitatively different from true stories. The current project investigated the features of linguistic style that distinguish between true and false stories. In an analysis of five independent samples, a computer-based text analysis program correctly classified liars and truth-tellers at a rate of 67% when the topic was constant and a rate of 61% overall. Compared to truth-tellers, liars showed lower cognitive complexity, used fewer self-references and other-references, and used more negative emotion words. PMID- 15272999 TI - Tactical differences in coping with rejection sensitivity: the role of prevention pride. AB - Prevention pride reflects a person's subjective history of success in preventing negative outcomes, leading to a strategic avoidance of errors of commission (e.g., explicit mistakes) in new situations. Two studies examined the impact of prevention pride on the strategies that highly rejection sensitive (HRS) people use to cope with the anxiety of anticipated rejection and the negative feelings elicited by perceived rejection. It was hypothesized that prevention pride orientation would lead HRS people toward covert and passive rather than overt and active forms of negative coping. Results indicated that HRS individuals who were also high in prevention pride reported increased use of self-silencing, presumably to prevent rejection. When rejection was perceived, however, they expressed hostility passively, by reducing positive behavior (e.g., withdrawing love and support) while inhibiting direct, active acts of hostility (e.g., yelling). PMID- 15273000 TI - Jealousy and the meaning (or nonmeaning) of violence. AB - Previous research has indicated that jealousy is one of the major triggers of domestic violence. Three studies here examined North Americans' ambivalent feelings about jealousy and jealousy-related aggression. In Study 1, it was shown that participants believed both that jealousy can be a sign of insecurity and a sign of love. In Study 2, it was shown that this equating of jealousy with love can lead to the tacit acceptance of jealousy-related violence. In Study 3, it was shown that a relative acceptance of jealousy-related aggression extends to cases of emotional and sexual abuse by husbands against their wives. In both Studies 2 and 3, men who hit or abused their wives over a jealousy-related matter were judged to romantically love their wives as much as those who did not engage in abuse. Violence in the context of a non-jealousy-related argument was seen quite negatively, but it lost a great deal of its negativity in the jealousy case. PMID- 15273001 TI - The ingroup as pars pro toto: projection from the ingroup onto the inclusive category as a precursor to social discrimination. AB - In an approach to intergroup discrimination and tolerance, it is assumed that the outgroup's difference from the ingroup is evaluated with reference to the prototype of the higher-order category that includes both groups. Two correlational studies yielded evidence that (a) group members tend to perceive their ingroup as relatively prototypical for the inclusive category (projection), (b) members highly identified with both ingroup and inclusive category (dual identity) tend to project most, and (c) relative prototypicality is related to negative attitudes toward the outgroup. The latter relation was further specified in Study 3, manipulating the valence of the inclusive category. Projection was related to more negative attitudes toward the outgroup when the inclusive category was primed positively but to more positive attitudes when it was primed negatively. The meaning of dual identities for intergroup relations is discussed. PMID- 15273002 TI - Contemporary racism in australia: the experiences of Aborigines. AB - In recent decades, social psychologists have suggested that contemporary racism is more subtle in nature than it had been in previous times. However, such theorizing has been from the perspective of the perpetrators. The present study follows a small number of other studies that have focused on the perspective of the victims of racism. It investigated the experiences of racism reported by 34 Aboriginal Australians during semi-structured, open-ended interviews. The data suggest that racism is experienced commonly and frequently by the participants and that much of it is overt or old-fashioned rather than subtle and modern. It is argued that if the data are reflective of what happens in intergroup encounters, social scientists may have embraced the theories of modern racism too readily. This may have contributed to the maintenance of social institutions that impact negatively on the minority populations in the community. PMID- 15273003 TI - Dominant group members in intergroup interaction: safety or vulnerability in numbers? AB - An experiment examined how low- and high-prejudice dominant group members' (LPs' and HPs') reactions to intergroup contact were affected by whether they were accompanied by fellow ingroup members who exhibited prejudice-relevant behavior. Participants answered questions alone or in a group and then estimated how they were viewed by an observer who was an ingroup or an outgroup member. HPs believed that they were viewed more negatively by an outgroup member in the individual than the group condition. LPs showed the opposite effect, which led them to evaluate the outgroup member more negatively in the group condition. All participants in the group condition expected an outgroup member to exaggerate their similarity to the other ingroup members present, and LPs evaluated the other ingroup members more negatively when the observer was an outgroup member. The results suggest that intergroup attitudes guide the types of intergroup contact situations that are experienced most positively. PMID- 15273004 TI - Getting to know you: the relational self-construal, relational cognition, and well-being. AB - Individuals with a highly relational self-construal define the self in terms of their close relationships with others. Consequently, they seek to nurture and develop new relationships. These studies examine individual differences in the self-construal in the context of a new roommate relationship, with a focus on cognitive aspects of relationship development. Study 1 revealed that persons with a highly relational self-construal were better able than others to predict a new roommate's values and beliefs. Study 2 showed that highly relational individuals tended to think optimistically about a new roommate's feelings about the relationship. The relational self-construal was more strongly related to these measures of relationship cognition in distant relationships than in very close relationships. Participants' self-construals and their perceptions of the closeness of the roommate relationship interacted in predicting well-being, revealing an unexpected negative relation between closeness and well-being for participants with a low relational self-construal. PMID- 15273005 TI - Mood as information in making attributions to discrimination. AB - Previous research demonstrates that people use their mood as information when making a variety of judgments. The present research examines the extent to which people use their current mood as information when making attributions to discrimination. Women were given a positive or negative mood induction and either provided with an external attribution for their current mood state or not. They then reported on discrimination occurring to themselves and other women. When an external attribution for induced mood was not provided, women in positive moods were less likely to report discrimination across three measures than were women in negative moods. When an external attribution was provided, mood had no effect. Implications for understanding the effects of context and individual differences in the perception and reporting of experiences with discrimination are discussed. PMID- 15273006 TI - Confronting prejudice (literally): reactions to confrontations of racial and gender bias. AB - Participants in two studies reported how they would feel, think, and behave after being confronted about either gender-biased or equivalent racial-biased responses. In Study 2, whether the confrontation was from a target group member (Black or female) or nontarget (White or male) group member was manipulated. Regardless of confronter status, allegations of racial bias elicited more guilt and apologetic-corrective responses and greater concern over having offended the confronter than similar confrontations of gender bias, which elicited more amusement. Target confrontations elicited less guilt but greater discomfort than nontarget confrontations and were associated with feelings of irritation and antagonism among more prejudiced participants. In addition, participants perceived a target's confrontation as more of an overreaction than the same confrontation from a nontarget. The implications of these findings for prejudice reduction efforts are discussed. PMID- 15273007 TI - Conflict over emotional expression: implications for interpersonal communication. AB - The current study examined the role of conflict over emotional expression for subjective and interpersonal functioning. The Ambivalence Over the Expression of Emotion Questionnaire (AEQ) was administered to female students who were videotaped while engaging in a conflict-resolution and feedback task with their boyfriends. External ratings showed ambivalent women to be less positive in their verbal statements and to be more constricted in their nonverbal expressions. When mood and other personality constructs were controlled for, ambivalence entailed greater overt submissiveness. Ambivalent women also displayed lower congruence between their verbal and nonverbal communication, irrespective of depression and other personality variables. These data suggest that conflict over emotional expression entails less congruent communication, less positivity in close relationships, and a subordinate stance for the ambivalent individual. PMID- 15273008 TI - Assessing anti-White attitudes and predicting perceived racism: the Johnson-Lecci scale. AB - Five studies are herein reported to describe the development and preliminary validation of the Johnson-Lecci Scale (JLS), a multicomponent self-report measure of anti-White attitudes held among Blacks. Items were generated from the everyday experiences of Black respondents using an act-frequency approach, and the scale configuration was derived using factor analysis. The factor structure was shown to be robust because it was cross-validated in an independent sample. The resulting JLS factors (subscales) were ingroup-directed stigmatization and discriminatory expectations, outgroup-directed negative beliefs, negative views toward ingroup-outgroup relations, and negative verbal expression toward the outgroup. These subscales were shown to predict the interpretations of ambiguously racist scenarios (i.e., perceived racism) and converged with peer evaluations of the target's anti-White attitudes. The subscales also demonstrate both convergent and discriminant validity with other self-report assessments of bias relating to age, gender, education, socioeconomic status, and race. PMID- 15273009 TI - Self-guide framing and persuasion: responsibly increasing message processing to ideal levels. AB - The current research examines the effect that framing persuasive messages in terms of self-guides (ideal vs. ought) has on the attitudes and cognitive responses of individuals with chronic ideal versus ought self-guides. The strength of participants' ideal and ought self-guides and the magnitude of participants' ideal and ought self-discrepancies were measured using a computerized reaction time program. One week later, participants read a persuasive message about a fictional breakfast product, framed in terms of either ideals or oughts. Matching framing to stronger self-guide led to enhanced message processing activity, especially among individuals who were low in need for cognition. Individuals who read messages framed to match their stronger self guides paid more attention to argument quality, as reflected in their attitudes and cognitive responses. Messages with self-guide framing that matched individuals' stronger self-discrepancies did not have this effect on processing. PMID- 15273010 TI - Sex differences in judgments of physical attractiveness: a social relations analysis. AB - Men and women rated the physical attractiveness of other men and women who were sitting nearby and were rated by them in return. They also provided meta perceptions of how they thought those others rated them. Attractiveness ratings were partly a function of both the target being rated and the perceiver providing the ratings regardless of the sex of the perceiver or target, but the highest levels of consensus occurred when men judged the attractiveness of women and the highest levels of idiosyncrasy occurred when men rated other men. Meta perceptions were also idiosyncratic; some believed that they were consistently considered attractive, whereas others thought they were seen as unattractive. People were aware of what others thought of them and, in particular, women's meta perceptions were highly related to men's judgments of them. People agree about others' attractiveness, and those who are attractive to others know they are pretty or handsome. PMID- 15273011 TI - Procedural justice and social regulation across group boundaries: does subgroup identity undermine relationship-based governance? AB - The relational model of authority suggests that people are inclined to accept the decisions of ethnic outgroup authorities when they identify with a superordinate category they share with the authority, and when the authority satisfies their relational justice concerns. Using responses from a random sample of African Americans, Latinos, and Whites about their cross-ethnic interactions with legal authorities, the findings indicated that those who are highly identified with the superordinate category of America indicate greater reliance on relational concerns and less on instrumental concerns when evaluating the authority's decision. In contrast, identification with one's ethnic subgroup did not moderate the linkage between relational concerns and acceptance. Across all ethnic groups, there were positive rather than negative correlations between measures of American and ethnic identity. Together, these findings indicate that subgroup identity does not undermine the relational basis of social regulation and that relationship-based governance is compatible with multiculturalism. PMID- 15273012 TI - Devaluation versus enhancement of attractive alternatives: a critical test using the calibration paradigm. AB - The calibration paradigm was used to test the competing hypotheses that (a) commitment motivates unduly negative evaluations of attractive alternatives (devaluation) versus (b) low commitment motivates exaggerated positive evaluations of attractive alternatives (enhancement). Single participants and dating participants low and high in relationship commitment were presented with an attractive, available person of the opposite sex and asked to judge the person's romantic appeal from their own perspective or from the perspective of their friends. Contrary to predictions based on the enhancement hypothesis, single and low-commitment participants did not provide higher ratings from their own perspective. In support of devaluation and calibration hypotheses, committed participants did provide lower ratings from their own perspective. Singles did not rate the target less attractive in a third condition in which the target was unavailable. However, dating participants, regardless of commitment level, rated the unavailable alternative negatively, consistent with social comparison processes and interdependence theory. PMID- 15273013 TI - Believing is seeing: the effects of racial labels and implicit beliefs on face perception. AB - Two studies tested whether racial category labels and lay beliefs about human traits have a combined effect on people's perception of, and memory for, racially ambiguous faces. Participants saw a morphed target face accompanied by a racial label (Black or White). Later, they were asked to identify the face from a set of two new morphed faces, one more Black and the other more White than the target. As predicted, entity theorists, who believe traits are immutable, perceived and remembered the target face as consistent with the racial label, whereas incremental theorists, who believe traits are malleable, perceived and remembered the face as inconsistent with the racial label. In Study 2, participants also drew the target face more consistently (entity theorists) or less consistently (incremental theorists) with the racial label. Results of both studies confirm that social variables can affect how physical features are seen and remembered. PMID- 15273014 TI - Stereotype threat in the classroom: dejection mediates the disrupting threat effect on women's math performance. AB - Research on stereotype threat, which is defined as the risk of confirming a negative stereotypic expectation about one's group, has demonstrated that the applicability of negative stereotypes disrupts the performance of stigmatized social groups. While it has been shown that a reduction of stereotype threat leads to improved performance by members of stigmatized groups, there is a lack of clear-cut findings about the mediating processes. The aim of the present study is to provide a better understanding of the mechanisms that stereotype threat causes in women working on mathematical problems. In addition, the study set out to test stereotype threat theory in a natural environment: high school classrooms. The experiment involved the manipulation of the gender fairness of a math test. The results indicate that the stereotype threat effect exists in this everyday setting. Moreover, it appears that dejection emotions mediate the effect of threat manipulation. PMID- 15273015 TI - Confusions of self with close others. AB - This article explores the cognitive underpinnings of interpersonal closeness in the theoretical context of "including other in the self" and, specifically, the notion of overlap between cognitive representations of self and close others. In each of three studies, participants first rated different traits for self, close others (e.g., romantic partner, best friend), and less close others (e.g., media personalities), followed by a surprise source recognition task (who was each trait rated for?). As predicted, in each study, there were more source confusions between traits rated for self and close others (e.g., a trait rated for self recalled as having been rated for the close other) than between self (or close others) and non-close others. Furthermore, several results suggest that the greater confusions between self and close others are due specifically to interpersonal closeness and not to greater familiarity or similarity with close others PMID- 15273016 TI - Effects of visual exposure to the opposite sex: cognitive aspects of mate attraction in human males. AB - This research is an investigation into the cognitive aspects of mate attraction in human males. Two experiments demonstrate that visual exposure to women (in person or within photographs) can prime large changes in the attitudes, mood states, and personality trait descriptions of male participants. These changes, furthermore, are such that participants show greater conformity to female mate preferences as described in the extant literature: In particular, men exposed to potential mates reported higher valuations of material wealth, greater momentary feelings of ambition, higher valuations of other indicators of social status, and personality trait descriptions indicative of high surgency/extraversion. All such effects occurred without participants' awareness that their responses had been affected by the experimental manipulations. These findings suggest a model of mate attraction mechanisms in which input cues from potential mates can prime those psychological representations that facilitate the behavioral expression of courtship tactics. PMID- 15273017 TI - Autocracy bias in informal groups under need for closure. AB - Two experiments investigated the tendency of groups with members under high (vs. low) need for cognitive closure to develop an autocratic leadership structure in which some members dominate the discussion, constitute the "hubs" of communication, and influence the group more than other members. The first experiment found that high (vs. low) need for closure groups, as assessed via dispositional measure of the need for closure, manifested greater asymmetry of conversational floor control, such that members with autocratic interactional style were more conversationally dominant and influential than less autocratic members. The second experiment manipulated the need for closure via time pressure and utilized a social network analysis. Consistent with expectation, groups under time pressure (vs. no pressure) showed a greater asymmetry of participation, of centrality, and of prestige among the group members, such that the more focal members were perceived to exert the greater influence over the groups' decisions. PMID- 15273018 TI - Effects of objective feedback and "single other" or "average other" social comparison feedback on performance judgments and helping behavior. AB - In two studies, participants received positive or negative feedback about their performance on a verbal task and then provided hints to another person on a subsequent, different task. It was expected that participants would give more helpful hints after positive than after negative feedback but that this would be more apparent when the feedback was based on performance comparisons with the "average participant" than on comparisons with another person or an objective standard. This effect was expected to be mediated by judgments of one's performance on the first task. These predictions were supported. Participants seemed aware of the effect of feedback on their hint choices, and their hint choices did not alter their affect levels. Also, participants receiving comparative (single or aggregated target) feedback exhibited changes in self ascribed importance of the performance domain. Implications for social comparison theory and self-evaluation maintenance theory are discussed. PMID- 15273019 TI - Introduction to the special issue on low frequency noise. PMID- 15273020 TI - Vibroacoustic disease. AB - Vibroacoustic disease (VAD) is a whole-body, systemic pathology, characterized by the abnormal proliferation of extra-cellular matrices, and caused by excessive exposure to low frequency noise (LFN). VAD has been observed in LFN-exposed professionals, such as, aircraft technicians, commercial and military pilots and cabin crewmembers, ship machinists, restaurant workers, and disk-jockeys. VAD has also been observed in several populations exposed to environmental LFN. This report summarizes what is known to date on VAD, LFN-induced pathology, and related issues. In 1987, the first autopsy of a deceased VAD patient was performed. The extent of LFN induced damage was overwhelming, and the information obtained is, still today, guiding many of the associated and ongoing research projects. In 1992, LFN-exposed animal models began to be studied in order to gain a deeper knowledge of how tissues respond to this acoustic stressor. In both human and animal models, LFN exposure causes thickening of cardiovascular structures. Indeed, pericardial thickening with no inflammatory process, and in the absence of diastolic dysfunction, is the hallmark of VAD. Depressions, increased irritability and aggressiveness, a tendency for isolation, and decreased cognitive skills are all part of the clinical picture of VAD. LFN is a demonstrated genotoxic agent, inducing an increased frequency of sister chromatid exchanges in both human and animal models. The occurrence of malignancies among LFN-exposed humans, and of metaplastic and displastic appearances in LFN-exposed animals, clearly corroborates the mutagenic outcome of LFN exposure. The inadequacy of currently established legislation regarding noise assessments is a powerful hindrance to scientific advancement. VAD can never be fully recognized as an occupational and environmental pathology unless the agent of disease--LFN- is acknowledged and properly evaluated. The worldwide suffering of LFN-exposed individuals is staggering and it is unethical to maintain this status quo. PMID- 15273021 TI - Low frequency noise and stress: bronchitis and cortisol in children exposed chronically to traffic noise and exhaust fumes. AB - A correlation of respiratory diseases to traffic related air pollution and noise was observed in an interview study. Since in that study the exposure was subjectively assessed, in the present field study nitrogen dioxide as indicator for vehicle exhausts and the mean night-time noise level were measured outside the children's windows in representative locations. Based on these measurements each child was placed in one of the following categories: low, medium or high traffic immission (ambient emissions). The physician contacts due to bronchitis of 68 children were assessed retrospectively from the files of the participating paediatricians. Saliva samples were collected from all children and the cortisol concentration was estimated. Children under high noise exposure (L(night, 8h) = 54-70dB(A)) had in comparison to all other children significantly increased morning saliva cortisol concentrations, indicating an activation of the hypothalamus-pituitary-adrenal (HPA) axis. Analysing a subgroup of children without high noise exposure showed, that children with frequent physician contacts due to bronchitis did not have increased morning saliva cortisol. However, multiple regression analysis with stepwise exclusion of variables showed that bronchitis was correlated more closely to morning salvia cortisol than to traffic immissions. On the other hand, the rate of physician contacts due to bronchitis increased in a dose dependent manner and significantly with increasing traffic immissions. From these results it can be concluded that high exposure to traffic noise, especially at nighttime, activates the HPA axis and this leads in the long term to an aggravation of bronchitis in children. This seems to be more important than the effect of exhaust fumes on bronchitis symptoms. The results of the present study should be subjected to further investigation using specially designed studies. PMID- 15273022 TI - Disturbing effects of low frequency sound immissions and vibrations in residential buildings. AB - Noise immissions with predominant low frequency sound components may exert considerably disturbing effects in dwellings. This applies in particular to sounds which are excitated by transmission of structure-borne noise, and to low frequency sounds emitted by ventilators. Exposed persons usually declare such immissions as being "intolerable" even at very low A-weighted sound levels. If mechanical vibrations in the frequency range below 20 Hz (ground-borne vibrations) affect dwelling rooms, the annoying effects are perceived only by a small portion of exposed individuals as a physical effect. For the most part the immissions are observed as vibratory effects on the building and on objects inside the dwelling. The disturbing effects of vibration frequencies above 20 Hz (structure-borne sound) are determined by the airborne sound field generated inside a particular room and its given surface and extension. PMID- 15273024 TI - Low frequency noise and annoyance. AB - Low frequency noise, the frequency range from about 10 Hz to 200 Hz, has been recognised as a special environmental noise problem, particularly to sensitive people in their homes. Conventional methods of assessing annoyance, typically based on A-weighted equivalent level, are inadequate for low frequency noise and lead to incorrect decisions by regulatory authorities. There have been a large number of laboratory measurements of annoyance by low frequency noise, each with different spectra and levels, making comparisons difficult, but the main conclusions are that annoyance of low frequencies increases rapidly with level. Additionally the A-weighted level underestimates the effects of low frequency noises. There is a possibility of learned aversion to low frequency noise, leading to annoyance and stress which may receive unsympathetic treatment from regulatory authorities. In particular, problems of the Hum often remain unresolved. An approximate estimate is that about 2.5% of the population may have a low frequency threshold which is at least 12 dB more sensitive than the average threshold, corresponding to nearly 1,000,000 persons in the 50-59 year old age group in the EU-15 countries. This is the group which generates many complaints. Low frequency noise specific criteria have been introduced in some countries, but do not deal adequately with fluctuations. Validation of the criteria has been for a limited range of noises and subjects. PMID- 15273023 TI - Hearing at low and infrasonic frequencies. AB - The human perception of sound at frequencies below 200 Hz is reviewed. Knowledge about our perception of this frequency range is important, since much of the sound we are exposed to in our everyday environment contains significant energy in this range. Sound at 20-200 Hz is called low-frequency sound, while for sound below 20 Hz the term infrasound is used. The hearing becomes gradually less sensitive for decreasing frequency, but despite the general understanding that infrasound is inaudible, humans can perceive infrasound, if the level is sufficiently high. The ear is the primary organ for sensing infrasound, but at levels somewhat above the hearing threshold it is possible to feel vibrations in various parts of the body. The threshold of hearing is standardized for frequencies down to 20 Hz, but there is a reasonably good agreement between investigations below this frequency. It is not only the sensitivity but also the perceived character of a sound that changes with decreasing frequency. Pure tones become gradually less continuous, the tonal sensation ceases around 20 Hz, and below 10 Hz it is possible to perceive the single cycles of the sound. A sensation of pressure at the eardrums also occurs. The dynamic range of the auditory system decreases with decreasing frequency. This compression can be seen in the equal-loudness-level contours, and it implies that a slight increase in level can change the perceived loudness from barely audible to loud. Combined with the natural spread in thresholds, it may have the effect that a sound, which is inaudible to some people, may be loud to others. Some investigations give evidence of persons with an extraordinary sensitivity in the low and infrasonic frequency range, but further research is needed in order to confirm and explain this phenomenon. PMID- 15273025 TI - Effects of low frequency noise up to 100 Hz. AB - This review concentrates on the effects of low frequency noise (LFN) up to 100 Hz on selected physiological parameters, subjective complaints and performance. The results of laboratory experiments and field studies are discussed in relation to the thresholds of hearing, of vibrotactile sensation and of aural pain. The effects of LFN may be mediated trough different ways. Temporary or permanent hearing threshold shifts seem to be due to acoustic stimuli above the individual hearing threshold. However, non-aural physiological and psychological effects may be caused by levels of low frequency noise below the individual hearing threshold. The dynamic range between the thresholds of hearing and of aural pain diminishes with decreasing frequency. This should be taken into account by the setting of limits concerning the health risks. Sufficient safety margins are recommended. The use of a frequency weighting with an attenuation of the low frequencies (e.g. G-weighting) does not seem to be appropriate for the evaluation of the health risks caused by LFN up to 100 Hz. It may be proposed to measure third octave band spectra or narrow band spectra. A comparison with the known human responses caused by the measured levels and frequencies could help to evaluate the health risks. Some proposals for further investigations were given: (1) experimental methods to discover the ways mediating the effects of low frequency noise, (2) consideration of the individual hearing threshold or hearing threshold shift and of the vibrotactile threshold in the low frequency range to be able to judge the effects, (3) consideration of combined body vibration caused by airborne low frequency noise or by other sources, (4) modelling to analyse the transmission of the acoustic energy from the input into the body to the structures containing sensors, (5) consideration of probable risk groups like children or pregnant women. PMID- 15273026 TI - Effects of low frequency noise on sleep. AB - Low frequency noise (20-200 Hz) is emitted by numerous sources in the society. As low frequencies propagate with little attenuation through walls and windows, many people may be exposed to low frequency noise in their dwellings. Sleep disturbance, especially with regard to time to fall asleep and tiredness in the morning, are commonly reported in case studies on low frequency noise. However, the number of studies where sleep disturbance is investigated in relation to the low frequencies in the noise is limited. Based on findings from available epidemiological and experimental studies, the review gives indications that sleep disturbance due to low frequency noise warrants further concern. PMID- 15273027 TI - Videoconferencing in child and adolescent telepsychiatry: a systematic review of the literature. AB - A systematic review of child and adolescent telepsychiatry was conducted. It was based on a search of the electronic databases MEDLINE and PsycINFO covering the period 1966 to June 2003. Studies were selected for review if they concerned videoconferencing for patient care or consultation, evaluated a clinical service or education, or assessed satisfaction with videoconferences. Twenty-seven articles were identified that fulfilled the selection criteria. These comprised two reports of randomized controlled experiments, 10 of descriptive questionnaire studies or observational surveys, seven case studies and eight other reports. Only three of the studies presented some calculations of cost-effectiveness. When classified by 'Quality of Evidence' criteria, only two studies were in category I (the highest), one was in II-2 and the rest fell into category III (the lowest). Most studies of child and adolescent telepsychiatry examined satisfaction with videoconferencing or described programmes or care regimens. Videoconferencing seemed to improve the accessibility of services and served an educational function. Some papers also mentioned savings in time, costs and travel. Problems with non-verbal communication and the audiovisual quality of the videoconference were mentioned as drawbacks. Telepsychiatry therefore seems to offer several benefits, at least in sparsely populated regions. Well designed and properly controlled trials are required to evaluate the clinical value of this promising method in child psychiatry, where there is a constantly increasing need for services. PMID- 15273028 TI - Diagnostic accuracy of and patient satisfaction with telemedicine for the follow up of paediatric burns patients. AB - Videoconferencing has become a routine technique for the post-acute burns care of children in Queensland. We compared the agreement between clinical assessments conducted via videoconference and assessments conducted in the conventional, face to-face manner (FTF). A total of 35 children with a previous burn injury were studied. Twenty-five children received three consecutive assessments: first FTF by a consultant in the outpatient department, then by a second consultant who reviewed the patient via videoconference, and then by the second consultant in person. The second consultant also reviewed another 10 children twice. At each review, the following variables were measured: scar colour, scar thickening, contractures, range of motion, the patient's level of general activity, any breakdown of the graft site, and adequacy of the consultation. Agreement between the two consultants when seeing patients FTF was moderately high, with an overall concordance of 85%. When videoconferencing was used, the level of agreement was almost the same, at 84%. If one consultant reviewed patients FTF first and then via videoconference, the overall concordance was 98%; if the process was reversed, the overall concordance was 97%. This study confirms that the quality of information collected during a videoconference appointment is comparable to that collected during a traditional, FTF appointment for a follow-up burns consultation. PMID- 15273029 TI - Telerehabilitation - a new model for community-based stroke rehabilitation. AB - Community resources for stroke clients are underdeveloped in Hong Kong and stroke survivors often face difficulties in community reintegration. We have examined the feasibility of using videoconferencing for community-based stroke rehabilitation. The sample comprised 21 stroke patients living at home. All the subjects participated in an eight-week intervention programme at a community centre for seniors. The intervention, which comprised educational talks, exercise and psychosocial support, was conducted by a physiotherapist via a videoconference link. The Berg Balance Scale (BBS), State Self-Esteem Scale (SSES), Medical Outcomes Study 36-item Short Form (SF-36) and a stroke knowledge test were administered at the start and end of the programme. In addition, at the start of the study the Geriatric Depression Scale 15-item Short Form, the Elderly Mobility Scale and the Lawton Instrumental Activities of Daily Living Scale were used to assess subjects' baseline status, and a focus group was also held at the end of the programme to gather qualitative findings. Nineteen subjects completed the eight-week intervention. The baseline functional status was high, although 52% had symptoms of depression. After the intervention, there were significant improvements in BBS, SSES and knowledge test scores and scores on all subscales of the SF-36. All the subjects accepted the use of videoconferencing for delivery of the intervention. The pilot study demonstrated the feasibility, efficacy and high level of acceptance of telerehabilitation for community-dwelling stroke clients. PMID- 15273030 TI - Comparison of an auto-stereoscopic display and polarized stereoscopic projection for macroscopic pathology. AB - We investigated stereoscopic imaging for gross examination in telepathology. A conventional macroscopic station was equipped with two cameras mounted 6.5 cm apart and images were produced of 30 different routine pathology specimens. Still images were displayed on a three-dimensional auto-stereoscopic display with a lenticular plate (which did not require the viewer to wear special glasses) and as a three-dimensional projection that required the viewer to wear glasses with polarized lenses. Nine observers (pathologists, laboratory technicians and engineers) viewed the three-dimensional images first on the auto-stereoscopic display and then with polarized projection. The observers scored the images for spatial reproduction, surface structure, proportions, colour and sharpness (10 indices in total, each rated on a five-point Likert scale of 1-5, with lower scores indicating better quality). Results were compared with those from five observers who had previously viewed the corresponding two-dimensional images on a conventional (two-dimensional) display. The mean scores across each of the 10 indices were 2.9 (two-dimensional display), 2.1 (auto-stereoscopic display) and 1.6 (polarized projection). All observers stated that the polarized projection had superior image quality with regard to resolution, colour and surface structures. The results obtained in the present study with still images have encouraged us to integrate stereoscopy into a dynamic telepathology system. PMID- 15273031 TI - Development and evaluation of a teleradiology and videoconferencing system. AB - We developed a teleradiology system linking a general hospital on Sado Island to tertiary care hospitals in Niigata City. The island is 40 km from Niigata City on the mainland and has only one diagnostic radiologist (for 72,000 islanders). Fibre optic cables between Sado Island and Niigata City were used for transmission. The introduction of the teleradiology system facilitated diagnostic and therapeutic consultation with specialists in Niigata City. The performance of the system was evaluated (on a scale of 0-6, with higher scores indicating better performance) by five diagnostic radiologists, who rated 32 features of the system twice, once in April 2002 and once in September 2003. The performance ratings improved from 1.38 to 2.86. While many of the initial problems with the software had been resolved, many still remained. PMID- 15273032 TI - A study of a rural telemedicine system in the Amazon region of Peru. AB - Voice and data communication facilities (email via VHF radio) were installed in 39 previously isolated health facilities in the province of Alto Amazonas in Peru. A baseline study was carried out in January 2001 and a follow-up evaluation in May 2002, after nine months of operation. We measured the reliability of the technology and the effect the system had on staff access to medical training and information. We also measured the indirect effects on the general population of access to better health-care. The experimental data were collected from 35 of the 39 sites in face-to-face questionnaire interviews. Before installation of the system, the mean consultation rate was 3 per month per facility (95% CI 1.5 to 4.5). At the end of the study, the mean consultation rate was 23 per month per facility (95% CI 14.7 to 31.5). There were 205 emergency transfers from the 39 health facilities. The system was employed in all these cases to alert the referral centre. The mean time required for evacuation was reduced from 8.6 h to 5.2 h. Health-care personnel reported that in 58 of the emergency cases (28%) the use of the system saved the life of the patient. The study shows that the use of communication technologies appropriate to local needs solves many problems in rural primary care, and that voice and email communication via VHF radio are feasible and useful for rural telemedicine. PMID- 15273033 TI - Health professionals' responses to the introduction of a home telehealth service. AB - An ethnographic (participant observation) study was undertaken of the socio technical processes involved in the implementation, within a randomized controlled trial, of a home telehealth nursing service for patients with chronic obstructive pulmonary disease (COPD). Ethnographic field notes were taken about technology-related tasks and the interplay between the research team and the 12 nurses who were to use the telehealth equipment. Views of the technology were linked to views of professional self-image and status. The technology was sometimes seen as unhelpful in establishing effective relationships with patients. Considerable work by all participants, over a period of months, was required to develop the technology in ways that minimized the risk to the stability of the specialist service and existing nurse-patient relationships. Our work highlights the complex problems that health professionals encounter when they try to integrate new technologies into routine service delivery. The concerns arising from the interplay of new technology with existing professional practices and relationships go beyond simple issues of training. PMID- 15273034 TI - A comparison of face-to-face and videoconference administration of the Hamilton Depression Rating Scale. AB - To determine whether the mode of administration affected the psychometric properties of the Hamilton Depression Rating Scale (HAMD), 21 subjects with an affective disorder were administered two HAMD interviews on the same day, once via videoconference (at 384 kbit/s) and once face-to-face, by two independent interviewers. The interviewers were blind to the results of the other interview. The mean HAMD scores were almost identical (18.4 for videoconferencing and 18.2 for face to face). The intra-class correlation between the two sets of scores was 0.88. When another group of 21 subjects had the HAMD administered in two face-to face interviews on the same day, the inter-rater reliability was not significantly different from that for the videoconference cohort. Most patients (91%) thought videoconferencing was a useful way to receive a psychological evaluation when other means were unavailable or limited. The study shows the psychometric equivalence of remote and face-to-face HAMD administration. PMID- 15273035 TI - Development of a telenursing system for patients with chronic conditions. AB - We have evaluated the feasibility of a telenursing system for patients with a chronic condition who are in receipt of home care services. The Internet-based system allows patients (equipped with a laptop computer), nurses and physicians to access information from a central database through a wireless network (128 kbit/s). Email and video-mail messages as well as vital signs data can be sent daily by the patient to a server at a regional health-care centre, and can be accessed by a nurse or physician, who can then decide on appropriate care. The system was tested by a male patient with type 2 diabetes mellitus, to see whether it would enhance his own management of his condition. During a 73-day baseline introductory period, no specific educational material was provided on the system Website about the management of diabetes (during this time the technical operation of the system was tested). During a second, 71-day period, educational material was provided. The telenursing system helped the patient to manage his condition, as shown by significant improvements in his levels of blood glucose and glycosylated haemoglobin (HbA(1c)) and in his blood pressure. Our findings suggest that the system is feasible. PMID- 15273036 TI - Motivation and tele-ophthalmology. PMID- 15273037 TI - Investigating e-health policy - tools for the trade. PMID- 15273038 TI - Nine tips for patients. Few of us like being patients, but there are ways to take charge of the situation and make the best of it. PMID- 15273039 TI - All-in-one pills for heart disease. A polypill may make sense, but drug companies are coming up with their own combinations. PMID- 15273040 TI - Fainting. Simple maneuvers could keep you from losing consciousness if you're feeling faint. PMID- 15273041 TI - The end of glasses? First came LASIK. Now there's a procedure for baby boomers who don't want to wear reading glasses. But does it work? PMID- 15273042 TI - What is a healthy bowel movement? The characteristics of feces can offer clues to health problems, digestive and otherwise. PMID- 15273043 TI - Almonds, oh joy! But peanuts better? They're good for us, but which nut is the best? PMID- 15273044 TI - By the way, doctor. I'm 70 and recently heard about a new blood test for something called lipoprotein(a) that might help determine if I'll get heart disease. Should I get this blood test? PMID- 15273045 TI - By the way, doctor. I read in the newspaper that angioplasty and stents don't do any good because narrowed arteries aren't the cause of most heart attacks. Is that true? PMID- 15273046 TI - Timing, target critical for cholesterol-lowering statins. Whether you are just starting a statin or have been taking one for years, make sure you're getting a "therapeutic dose" and hit your target. PMID- 15273047 TI - Benefits from moderate drinking extended? A drink a day may be okay for some people with high blood pressure. PMID- 15273048 TI - Exercise your right to health. Don't let inconsistent guidelines on how much or what kind of exercise is best for your heart keep you seated in frustration. PMID- 15273049 TI - Split thinking on low cholesterol and heart failure. Even though low cholesterol may signal trouble for people with heart failure, taking a cholesterol-lowering statin may help. PMID- 15273050 TI - Even cholesterol has its seasons. PMID- 15273051 TI - Just say no to heart screening tests. PMID- 15273052 TI - Vitamin, mineral intake low or high--not just right. PMID- 15273053 TI - Ask the doctor. After having had vague leg pain for a few months, I finally went to the doctor. She said I have venous insufficiency. What does that mean and what can I do about it? PMID- 15273054 TI - Ask the doctor. When checking my blood pressure at home, I always put the cuff on my right arm. Just for fun, one day I put it on my left arm and got a different reading. Did I make a mistake, or can someone have different blood pressure readings depending on which arm is used? PMID- 15273055 TI - Hormone therapy's unanswered questions. PMID- 15273056 TI - Artificial sweeteners: okay in moderation. PMID- 15273057 TI - Hemorrhoids and what to do about them. A few simple strategies can help ease the pain and trouble associated with these bothersome bulging blood vessels. PMID- 15273058 TI - Shoring up your immunity. Your immune system is always on patrol. Does it need any help from you? PMID- 15273060 TI - Statins for type 2 diabetes. PMID- 15273059 TI - Calcium may cut kidney stone risk in younger women. PMID- 15273061 TI - By the way, doctor. I feel tired, and my joints ache. I've been told I have a vitamin B12 deficiency. What is this--and what causes it? PMID- 15273063 TI - Insect bites and stings. PMID- 15273062 TI - By the way, doctor. I'm only 52 years old and I've been diagnosed with degenerative arthritis in my lower back. What exactly does this mean? I take ibuprofen daily, and it helps some. What else can I do? PMID- 15273064 TI - Scanning the scans: comparing CT, MRI, and PET scans. PMID- 15273065 TI - Fish oil: staging a comeback. PMID- 15273066 TI - On call. I developed severe ulcerative colitis in my late teens, and I had my entire colon and rectum removed at age 24. It was certainly worthwhile, since I adjusted easily and I've been healthy ever since. But now I'm 52, and I'm starting to think about prostate cancer. My doctor obviously can't do a rectal exam, and he doesn't want to do a PSA test because he says he won't be able to send me for a prostate biopsy if the test is high. Do I have any options? PMID- 15273067 TI - The addicted brain. Neuroscientific research reveals new ways of understanding drug and alcohol dependence. PMID- 15273068 TI - Hypochondria. PMID- 15273069 TI - Emotion in economic decisions. PMID- 15273070 TI - Do women make better leaders? PMID- 15273071 TI - Questions & answers. What is a repetition compulsion, and how can it be changed? PMID- 15273072 TI - Effects of creatine supplementation on renal function. AB - Creatine is a popular supplement used by athletes in an effort to increase muscle performance. The purpose of this review was to assess the literature evaluating the effects of creatine supplementation on renal function. A PubMed search was conducted to identify relevant articles using the keywords, creatine, supplementation, supplements, renal dysfunction, ergogenic aid and renal function. Twelve pertinent articles and case reports were identified. According to the existing literature, creatine supplementation appears safe when used by healthy adults at the recommended loading (20 gm/day for five days) and maintenance doses (100-kb) plasmids (designated types A, B, and C) and one 10.1-kb plasmid (type D) that carry the blaCMY-2 gene. In the present study, the distribution of these four known blaCMY-2-carrying plasmids among 35 ceftriaxone-resistant Salmonella isolates obtained from 1998 to 2001 was examined. Twenty-three of these isolates were Salmonella enterica serotype Newport, 10 were Salmonella enterica serotype Typhimurium, 1 was Salmonella enterica serotype Agona, and 1 was Salmonella enterica serotype Reading. All 23 serotype Newport isolates carried a type C plasmid, and 5, 4, and 1 serovar Typhimurium isolate carried type B, A, and C plasmids, respectively. Both the serotype Agona and serotype Reading isolates carried type A plasmids. None of the isolates carried a type D plasmid. Hybridization data suggested that plasmid types A and C were highly related replicons. DNA sequencing revealed that the region surrounding blaCMY-2 was highly conserved in all three plasmid types analyzed (types B, C, and D) and was related to a region surrounding blaCMY-5 from the Klebsiella oxytoca plasmid pTKH11. These findings are consistent with a model in which blaCMY-2 has been disseminated primarily through plasmid transfer, and not by mobilization of the gene itself, to multiple Salmonella chromosomal backbones. PMID- 15273089 TI - Prediction of mechanisms of action of antibacterial compounds by gene expression profiling. AB - We have generated a database of expression profiles carrying the transcriptional responses of the model organism Bacillus subtilis following treatment with 37 well-characterized antibacterial compounds of different classes. The database was used to build a predictor for the assignment of the mechanisms of action (MoAs) of antibacterial compounds by the use of support vector machines. This predictor was able to correctly classify the MoA class for most compounds tested. Furthermore, we provide evidence that the in vivo MoA of hexachlorophene does not match the MoA predicted from in vitro data, a situation frequently faced in drug discovery. A database of this kind may facilitate the prioritization of novel antibacterial entities in drug discovery programs. Potential applications and limitations are discussed. PMID- 15273091 TI - Cellular pharmacokinetics and pharmacodynamics of the glycopeptide antibiotic oritavancin (LY333328) in a model of J774 mouse macrophages. AB - The intracellular pharmacokinetics and pharmacodynamics of oritavancin (LY333328) were studied in cultured cells. Oritavancin was avidly accumulated by J774 and THP-1 macrophages and rat fibroblasts and to a lesser extent by LLC-PK1 and Caco 2 cells. In J774 macrophages, the level of accumulation reached a plateau (at 370 fold the extracellular concentration) within 24 h and was partly defeated by a rise in serum protein levels. Efflux was incomplete (with a plateau at two-thirds of the original level at 6 h). In short-term kinetic studies, oritavancin uptake was linear for up to 4 h (as was the case for horseradish peroxidase and small latex beads, used as markers of the fluid phase and adsorptive endocytosis, respectively), which was in contrast to azithromycin and chloroquine uptake (which accumulate in cells by diffusion and segregation). The rates of clearance of oritavancin and latex beads were comparable (150 and 120 microl x mg of protein(-1) x h(-1), respectively) and were approximately 200 times higher than that of horseradish peroxidase. Oritavancin accumulation was partially reduced by monensin but was unaffected by acidic pH (these conditions abolished chloroquine accumulation). Cell-associated oritavancin was found in lysosomal fractions after homogenization of J774 macrophages and fractionation by isopycnic centrifugation. Oritavancin was bactericidal against intracellular Staphylococcus aureus (phagolysosomal infection) but was unable to control the intracellular growth of Listeria monocytogenes (cytosolic infection), even though its cellular concentration largely exceeded the MIC (0.02 mg/liter) and minimal bactericidal concentration (2 mg/liter). We conclude that oritavancin enters cells by adsorptive endocytosis (favored by its lipophilic side chain and/or the presence of three protonatable amines), which drives it to lysosomes, where it exerts antibiotic activity. PMID- 15273092 TI - Effects of an antibiotic cycling program on antibiotic prescribing practices in an intensive care unit. AB - Various interventions have been proposed to combat the increase of antibiotic resistance and influence antibiotic prescribing practices. A prospective cohort study in a medical intensive care unit was conducted to determine the effect of an antibiotic cycling program on patterns of antibiotic use and to determine patient factors associated with cycling adherence. Four major classes of antibiotics for empirical therapy of suspected gram-negative bacterial infections were rotated at 3- and 4-month intervals. During the study, 1,003 patients received antibiotic therapy with at least one of the study drugs; of the 792 receiving cycle antibiotics during the cycling period, 598 (75.5%) received an on cycle drug. Compared to the baseline, cycling recommendations increased the use of the target cycle agent: the use of cephalosporins increased during cycle 1 (56 to 64% of total antibiotic days, P < 0.001), fluoroquinolone use increased in cycle 2 (24 to 55%, P < 0.001), carbapenem use increased during cycle 3 (14 to 38%, P < 0.001), and use of extended-spectrum penicillins increased in cycle 4 (5 to 36%, P < 0.001). Overall, 48% of total cycle antibiotic days were compliant with the cycling protocol. On average, 8.8 days per patient were spent receiving on-cycle drugs (range, 1 to 109). Cycle periods that specified carbapenem and fluoroquinolone use had the highest number of off-cycle days (62 and 64%). Predictors of on-cycle antibiotic use were increased severity of illness, as measured by an acute physiology and chronic health evaluation II score, and greater length of intensive care unit stay. In conclusion, the successful implementation of this cycling protocol increased antibiotic heterogeneity over time in the study unit. PMID- 15273093 TI - Pilot randomized double-blind trial of treatment of Mycobacterium ulcerans disease (Buruli ulcer) with topical nitrogen oxides. AB - Mycobacterium ulcerans disease (Buruli ulcer) is a serious ulcerating skin disease which is common in many tropical countries. Standard treatment, by extensive excision and skin grafting, is not available in rural communities where the disease is common. We evaluated the efficacy and safety of treatment with topical nitrogen oxides. Thirty-seven patients with a clinical diagnosis of Buruli ulcer caused by M. ulcerans disease were randomly assigned to one of two groups. In one group, two creams containing sodium nitrite (6%, wt/wt) or citric acid monohydrate (9%, wt/wt) were applied daily for 6 weeks, while the other group received a placebo. In the second 6 weeks, both groups received the nitrogen oxide-generating combination of creams. Treatment was continued for another 4 weeks for patients whose ulcers were not healed after 12 weeks. The ulcer surface area was monitored by weekly tracings made by assessors blinded to the treatment. In the first 6 weeks, patients on sodium nitrite and citric acid monohydrate (group I, active treatment) showed a rapid decrease in ulcer size from 28.6 +/- 5.6 cm2 (mean +/- standard error) to 12.6 +/- 3.2 cm2, a decrease significantly greater than that in group II (from 15.3 +/- 3.1 to 11.7 +/- 3.7 cm2; P = 0.03). Five ulcers in the placebo group enlarged during this period, compared with one in the active group. In the second 6 weeks (both groups on active treatment), the rates of healing were similar for the two groups and there was a significant reduction in ulcer size in group II (previously on placebo) compared to the first 6 weeks. Yellow pigmentation of the skin, which disappeared 3 days after treatment was stopped, was the only side effect to date. We conclude that creams releasing nitrogen oxides increase the healing rate of ulcers caused by M. ulcerans infection with minimal adverse events. This is the first controlled trial of any form of therapy which demonstrates efficacy in treating this disease. PMID- 15273094 TI - Synergy of daptomycin with oxacillin and other beta-lactams against methicillin resistant Staphylococcus aureus. AB - We previously observed marked synergy between daptomycin and both rifampin and ampicillin against vancomycin-resistant enterococci (VRE). Because the synergy between daptomycin and ampicillin was observed for 100% of VRE strains with high level ampicillin resistance (ampicillin MIC of > or =128 microg/ml), we looked for synergy between daptomycin and other beta-lactams against 18 strains of methicillin-resistant Staphylococcus aureus (MRSA) by employing a time-kill method using Mueller-Hinton broth supplemented to 50 mg of Ca2+/liter. All strains were resistant to oxacillin (16 of 18 strains were resistant at drug concentrations of > or =256 microg/ml), and all strains were susceptible to daptomycin (the MIC at which 90% of the tested isolates were inhibited was 1 microg/ml). Daptomycin was tested at concentrations of 2, 1, 0.5, 0.25, 0.125, and 0.0625 microg/ml alone or in combination with oxacillin at a fixed concentration of 32 microg/ml. Synergy was found for all 18 strains with daptomycin at one-half the MIC in combination with 32 microg of oxacillin/ml, and synergy was found for 11 of 18 strains (61%) with daptomycin at one-fourth the MIC or less in combination with oxacillin. At 24 h, the daptomycin-oxacillin combination with daptomycin at one-half the MIC showed bactericidal activity against all 18 strains, and the combination with one-fourth the daptomycin MIC showed bactericidal activity against 9 of 18 strains. We also used a novel screening method to look for synergy between daptomycin and other beta-lactams. In this approach, daptomycin was incorporated into Ca(2+)-supplemented Mueller Hinton agar at subinhibitory concentrations, and synergy was screened by comparing test antibiotic Kirby-Bauer disks on agar with and without daptomycin. By this method, daptomycin with ampicillin-sulbactam, ticarcillin-clavulanate, or piperacillin-tazobactam showed synergy comparable to or greater than daptomycin with oxacillin. For seven of the eight strains tested, time-kill studies confirmed synergy between daptomycin and ampicillin-sulbactam with ampicillin in the range of 2 to 8 microg/ml. The combination of daptomycin and beta-lactams may be useful for the treatment of MRSA infection, but further studies are needed to elucidate the mechanisms and to determine the in vivo efficacy of the combination. PMID- 15273095 TI - Inhibition of hepatitis C virus replication by arsenic trioxide. AB - Hepatitis C virus (HCV) is a serious global problem, and present therapeutics are inadequate to cure HCV infection. In the present study, various antiviral assays show that As2O3 at submicromolar concentrations is capable of inhibiting HCV replication. The 50% effective concentration (EC50) of As2O3 required to inhibit HCV replication was 0.35 microM when it was determined by a reporter-based HCV replication assay, and the EC50 was below 0.2 microM when it was determined by quantitative reverse transcription-PCR analysis. As2O3 did not cause cellular toxicity at this concentration, as revealed by an MTS [3-(4,5-dimethylthiozol-2 yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt] assay. A combination of As2O3 and alpha interferon exerted synergistic effects against HCV, as revealed by a multiple linear logistic model and isobologram analysis. Furthermore, in an alternative HCV antiviral system that may recapitulate additional steps involved in HCV infection and replication, As2O3 at 0.3 microM totally abolished the HCV signal, whereas alpha interferon at a high dose (5,000 IU/ml) only partially suppressed the HCV signal. The study highlights the indications for use of a novel class of anti-HCV agent. Further elucidation of the exact antiviral mechanism of As2O3 may lead to the development of agents with potent activities against HCV or related viruses. PMID- 15273096 TI - Effect of amoxicillin use on oral microbiota in young children. AB - Dental plaque samples from 40 children were screened for the presence of bacteria resistant to amoxicillin. Fifteen children had used amoxicillin and 25 had not used any antibiotic in the 3 months prior to sample collection. All (100%) of the children harbored amoxicillin-resistant oral bacteria. The median percentage of the total cultivable oral microbiota resistant to amoxicillin was 2.4% (range, 0.1 to 14.3%) in children without amoxicillin use and 10.9% (range, 0.8 to 97.3%) in children with amoxicillin use, with the latter value being significantly higher (P < 0.01). A total of 224 amoxicillin-resistant bacteria were isolated and comprised three main genera: Haemophilus spp., Streptococcus spp., and Veillonella spp. The biodiversity of the amoxicillin-resistant microbiota was similar among the isolates from children with and without previous antibiotic use. The amoxicillin MIC at which 90% of the isolates were inhibited for isolates from children who had used amoxicillin in the previous 3 months was higher (64 mg liter(-1)) than that obtained for the isolates from subjects who had not used antibiotics (16 mg liter(-1)). The majority of the amoxicillin-resistant isolates (65%) were also resistant to at least one of the three antibiotics tested (penicillin, erythromycin, and tetracycline), with resistance to penicillin (51% of isolates) being the most frequently encountered. However, significantly more (P < 0.05) of the amoxicillin-resistant isolates from subjects with previous amoxicillin use were also resistant to erythromycin. This study has demonstrated that a diverse collection of amoxicillin-resistant bacteria is present in the oral cavity and that the number, proportions, MICs, and resistance to erythromycin can significantly increase with amoxicillin use. PMID- 15273097 TI - Antibiotic-inducible promoter regulated by the cell envelope stress-sensing two component system LiaRS of Bacillus subtilis. AB - Soil bacteria are among the most prodigious producers of antibiotics. The Bacillus subtilis LiaRS (formerly YvqCE) two-component system is one of several antibiotic-sensing systems that coordinate the genetic response to cell wall active antibiotics. Upon the addition of vancomycin or bacitracin, LiaRS autoregulates the liaIHGFSR operon. We have characterized the promoter of the lia operon and defined the cis-acting sequences necessary for antibiotic-inducible gene expression. A survey for compounds that act as inducers of the lia promoter revealed that it responds strongly to a subset of cell wall-active antibiotics that interfere with the lipid II cycle in the cytoplasmic membrane (bacitracin, nisin, ramoplanin, and vancomycin). Chemicals that perturb the cytoplasmic membrane, such as organic solvents, are also weak inducers. Thus, the reporter derived from P(liaI) (the liaI promoter) provides a tool for the detection and classification of antimicrobial compounds. PMID- 15273098 TI - Impact of cethromycin (ABT-773) therapy on microbiological, histologic, immunologic, and respiratory indices in a murine model of Mycoplasma pneumoniae lower respiratory infection. AB - Mycoplasma pneumoniae is a major etiologic agent of acute lower respiratory infections. We evaluated the antimicrobial and immunologic effects of cethromycin (ABT-773), a ketolide antibiotic, for the treatment of M. pneumoniae pneumonia in a mouse model. Eight-week-old BALB/c mice were inoculated intranasally once with 10(6) CFU of M. pneumoniae on day 0. Treatment was started 24 h after inoculation. Groups of mice were treated subcutaneously with cethromycin at 25 mg/kg of body weight or with placebo daily until sacrifice. Five to ten mice per group were evaluated at days 1, 4, 7, and 10 after inoculation. Outcome variables included bronchoalveolar lavage (BAL) for M. pneumoniae quantitative culture and cytokine and chemokine concentration determinations by enzyme-linked immunosorbent assay (tumor necrosis factor alpha [TNF-alpha], gamma interferon [IFN-gamma], interleukin-1beta [IL-1beta], IL-2, IL-4, IL-12, granulocyte macrophage colony-stimulating factor, IL-8, monocyte chemoattractant protein 1 [MCP-1], and macrophage inflammatory protein 1alpha [MIP-1alpha]), histopathologic score of the lungs (HPS), and pulmonary function tests (PFT) using whole-body, unrestrained plethysmography at the baseline and post methacholine exposure as indicators of airway obstruction (AO) and airway hyperresponsiveness (AHR), respectively. The cethromycin-treated mice had a greater reduction in M. pneumoniae culture titers than placebo-treated mice, reaching statistical significance on days 7 and 10 (P < 0.05). HPS was significantly reduced in cethromycin-treated mice compared with placebo-treated mice on days 4, 7, and 10 (P < 0.05). Cytokine concentrations in BAL samples were reduced in mice that received cethromycin, and the differences were statistically significant for 7 of the 10 cytokines measured (TNF-alpha, IFN-gamma, IL-1beta, IL-8, IL-12, MCP-1, and MIP-1alpha) on day 4 (P < 0.05). PFT values were improved in the cethromycin-treated mice, with AO and AHR significantly reduced on day 4 (P < 0.05). In this mouse model, treatment with cethromycin significantly reduced M. pneumoniae culture titers in BAL samples, cytokine and chemokine concentrations in BAL samples, histologic inflammation in the lungs, and disease severity as defined by AO and AHR. PMID- 15273099 TI - Molecular characterization of a cephamycin-hydrolyzing and inhibitor-resistant class A beta-lactamase, GES-4, possessing a single G170S substitution in the omega-loop. AB - The nosocomial spread of six genetically related Klebsiella pneumoniae strains producing GES-type beta-lactamases was found in a neonatal intensive care unit, and we previously reported that one of the six strains, strain KG525, produced a new beta-lactamase, GES-3. In the present study, the molecular mechanism of cephamycin resistance observed in strain KG502, one of the six strains described above, was investigated. This strain was found to produce a variant of GES-3, namely, GES-4, which was responsible for resistance to both cephamycins (cefoxitin MIC, >128 microg/ml) and beta-lactamase inhibitors (50% inhibitory concentration of clavulanic acid, 15.2 +/- 1.7 microM). The GES-4 enzyme had a single G170S substitution in the Omega-loop region compared with the GES-3 sequence. This single amino acid substitution was closely involved with the augmented hydrolysis of cephamycins and carbapenems and the decreased affinities of beta-lactamase inhibitors to GES-4. A cloning experiment and sequencing analysis revealed that strain KG502 possesses duplicate bla(GES-4) genes mediated by two distinct class 1 integrons with similar gene cassette configurations. Moreover, the genetic environments of the bla(GES-4) genes found in strain KG502 were almost identical to that of bla(GES-3) in strain KG525. From these findings, these two phenotypically different strains were suggested to belong to a clonal lineage. The bla(GES-4) gene found in strain KG502 might well emerge from a point mutation in the bla(GES-3) gene harbored by its ancestor strains, such as strain KG525, under heavy antibiotic stress in order to acquire extended properties of resistance to cephamycins and carbapenems. PMID- 15273100 TI - Estimation of serum-free 50-percent inhibitory concentrations for human immunodeficiency virus protease inhibitors lopinavir and ritonavir. AB - Using measured free fraction and 50% inhibitory concentration (IC50) values for the human immunodeficiency virus protease inhibitors lopinavir (LPV) and ritonavir (RTV) in tissue culture media with various protein concentrations ranging from 5 to 50%, we estimated serum-free IC50 values for each drug. The range of serum-free IC50 values (0.64 to 0.77 ng/ml for LPV and 3.0 to 5.0 ng/ml for RTV) did not exhibit a trend with increasing protein concentrations, despite a 10-fold difference in the free fraction value (0.006 to 0.063) for LPV and a 5 fold difference in the free fraction value (0.013 to 0.057) for RTV. The mean serum-free IC50 by the MTT-MT4 assay (0.69 ng/ml for LPV and 4.0 ng/ml for RTV) may be the most accurate parameter for the estimation of the inhibitory quotient (IQ), a relative measure of in vivo potency defined as the ratio of the minimal free drug concentration in plasma (C(trough,free)) for a specific patient population and the serum-free IC50. Using this approach, we calculated the average IQs for protease inhibitor-naive patients for LPV and RTV to be 67 and 5.6, respectively. PMID- 15273101 TI - Therapeutic efficacy of "nubiotics" against burn wound infection by Pseudomonas aeruginosa. AB - "Nubiotics" are a novel class of proprietary protonated nucleic acid-based drugs shown to have potent in vitro antibacterial activities against a number of gram positive and gram-negative bacteria. These nubiotics are evaluated here for their in vivo therapeutic efficacy for the treatment of burn wound infection caused by Pseudomonas aeruginosa. To achieve this, a burn wound infection model was established in mice by using a highly pathogenic burn wound clinical isolate of P. aeruginosa. Lethal doses of the bacteria were determined for two routes of infection (subcutaneous and topical), representing systemic and local forms of infection, respectively. Using this infection model, treatment with nubiotics by various routes of drug administration was evaluated and optimized. A total of 12 nubiotics and their analogues were tested and of these, Nu-2, -3, -4, and -5 were found to be extremely efficacious in the postexposure treatment of burn wound infection (60 to 100% survival rates versus 0% for untreated control [P < 0.05]). These nubiotics were effective when given either systemically by intravenous and/or subcutaneous administration or given locally to the affected site in the skin by topical application. Treatment by these two routes resulted in almost 100% survival rates and complete eradication of the bacteria from infection sites in the livers, spleens, and blood. These nubiotics were found to be as effective as intravenously administered ciprofloxacin, a potent and broad-spectrum fluoroquinolone. These results suggest that nubiotics may be a promising and effective approach for the treatment of burn wound infection caused by P. aeruginosa. PMID- 15273102 TI - Rapid real-time PCR genotyping of mutations associated with sulfadoxine pyrimethamine resistance in Plasmodium falciparum. AB - The resistance of Plasmodium falciparum to sulfadoxine-pyrimethamine (SP) is an emerging public health threat. Resistance to these drugs is associated with point mutations in the genes encoding dihydropteroate synthase (DHPS) and dihydrofolate reductase (DHFR). We describe here an assay using real-time PCR and sequence specific probes that detects these mutations. Using DNA from plasmids, cultured strains, and clinical samples, real-time PCR could distinguish four DHPS polymorphisms (codons 437, 540, 581, and 613) and three DHFR polymorphisms (codons 51, 59, and 108). This assay is rapid and sensitive, with a detection limit of 10 copies in most cases. This assay is amenable to large-scale studies of drug resistance. PMID- 15273103 TI - Ribosomal P0 protein domain involved in selectivity of antifungal sordarin derivatives. AB - The ribosomal stalk protein P0 is involved in the susceptibility to the antifungal sordarin derivatives, as reported for a number of Saccharomyces cerevisiae resistant mutants. Mammals and some lower eukaryotes are naturally resistant to these compounds. It is shown here that expression in S. cerevisiae of the ribosomal protein P0 from Homo sapiens and from other sordarin-resistant organisms results in a decrease in the sensitivity of the cells to an agent of this class. To further characterize the P0 region responsible for inducing sordarin resistance, a series of protein chimeras containing complementary regions of the human and yeast P0 proteins were constructed and expressed in yeast. The chimeras complement the absence of the native yeast P0 except in chimeras containing the human P0 carboxyl-terminal domain. Resistance to sordarins was found to be associated with the presence of an HsP0 amino acid sequence comprising P118 to F138, which unexpectedly led to higher resistance than the presence of the complete human P0. A comparison of the corresponding region in P0 from yeast and sordarin-insensitive organisms, followed by site directed mutagenesis, indicates that residues in positions 119, 124, and 126 have an important role in determining resistance to sordarins. Moreover, since sordarins block the eukaryotic elongation factor 2 (EF2) function, the P0 region affecting sordarin susceptibility must correspond to EF2-interacting domains of the ribosomal stalk protein, which affects the drug-binding site in the elongation factor. PMID- 15273104 TI - Novel azasterols as potential agents for treatment of leishmaniasis and trypanosomiasis. AB - This paper describes the design and evaluation of novel azasterols as potential compounds for the treatment of leishmaniasis and other diseases caused by trypanosomatid parasites. Azasterols are a known class of (S)-adenosyl-L methionine: Delta24-sterol methyltransferase(24-SMT) inhibitors in fungi, plants, and some parasitic protozoa. The compounds prepared showed activity at micromolar and nanomolar concentrations when tested against Leishmania spp. and Trypanosoma spp. The enzymatic and sterol composition studies indicated that the most active compounds acted by inhibiting 24-SMT. The role of the free hydroxyl group at position 3 of the sterol nucleus was also probed. When an acetate was attached to the 3beta-OH, the compounds did not inhibit the enzyme but had an effect on parasite growth and the levels of sterols in the parasite, suggesting that the acetate group was removed in the organism. Thus, an acetate group on the 3beta-OH may have application as a prodrug. However, there may be an additional mode(s) of action for these acetate derivatives. These compounds were shown to have ultrastructural effects on Leishmania amazonensis promastigote membranes, including the plasma membrane, the mitochondrial membrane, and the endoplasmic reticulum. The compounds were also found to be active against the bloodstream form (trypomastigotes) of Trypanosoma brucei rhodesiense, a causative agent of African trypanosomiasis. PMID- 15273105 TI - Isoniazid pharmacokinetics-pharmacodynamics in an aerosol infection model of tuberculosis. AB - Limited data exist on the pharmacokinetic-pharmacodynamic (PK-PD) parameters of the bactericidal activities of the available antimycobacterial drugs. We report on the PK-PD relationships for isoniazid. Isoniazid exhibited concentration (C) dependent killing of Mycobacterium tuberculosis H37Rv in vitro, with a maximum reduction of 4 log10 CFU/ml. In these studies, 50% of the maximum effect was achieved at a C/MIC ratio of 0.5, and the maximum effect did not increase with exposure times of up to 21 days. Conversely, isoniazid produced less than a 0.5 log10 CFU/ml reduction in two different intracellular infection models (J774A.1 murine macrophages and whole human blood). In a murine model of aerosol infection, isoniazid therapy for 6 days produced a reduction of 1.4 log10 CFU/lung. Dose fractionation studies demonstrated that the 24-h area under the concentration-time curve/MIC (r2 = 0.83) correlated best with the bactericidal efficacy, followed by the maximum concentration of drug in serum/MIC (r2 = 0.73). PMID- 15273106 TI - Modulation of fibronectin adhesins and other virulence factors in a teicoplanin resistant derivative of methicillin-resistant Staphylococcus aureus. AB - The impact of glycopeptide resistance on the molecular regulation of Staphylococcus aureus virulence and attachment to host tissues is poorly documented. We compared stable teicoplanin-resistant methicillin-resistant S. aureus (MRSA) strain 14-4 with its teicoplanin-susceptible MRSA parent, strain MRGR3, which exhibits a high degree of virulence in a rat model of chronic foreign body MRSA infection. The levels of fibronectin-mediated adhesion and surface display of fibronectin-binding proteins were higher in teicoplanin resistant strain 14-4 than in its teicoplanin-susceptible parent or a teicoplanin susceptible revertant (strain 14-4rev) that spontaneously emerged during tissue cage infection. Quantitative reverse transcription-PCR (qRT-PCR) showed four- and twofold higher steady-state levels of fnbA and fnbB transcripts, respectively, in strain 14-4 than in its teicoplanin-susceptible counterparts. Analysis of global regulatory activities by qRT-PCR revealed a strong reduction in the steady-state levels of RNAIII and RNAII in the teicoplanin-resistant strain compared to in its teicoplanin-susceptible counterparts. In contrast, sarA mRNA levels were more than fivefold higher in strain 14-4 than in MRGR3 and 14-4rev. Furthermore, the alternative transcription factor sigma B had a higher level of functional activity in the teicoplanin-resistant strain than in its teicoplanin-susceptible counterparts, as evidenced by significant increases in both the sigma B-dependent asp23 mRNA levels and the sarA P3 promoter-derived transcript levels, as assayed by qRT-PCR and Northern blotting, respectively. These data provide further evidence that the emergence of glycopeptide resistance is linked by still poorly understood molecular pathways with significant pleiotropic changes in the expression and regulation of some major virulence genes. These molecular and phenotypic changes may have a profound impact on the bacterial adhesion and colonization properties of such multiresistant organisms. PMID- 15273107 TI - Disposition of artesunate and dihydroartemisinin after administration of artesunate suppositories in children from Papua New Guinea with uncomplicated malaria. AB - A detailed pharmacokinetic analysis was performed with 47 children from Papua New Guinea with uncomplicated falciparum or vivax malaria treated with artesunate (ARTS) suppositories (Rectocaps) given in two doses of approximately 13 mg/kg of body weight 12 h apart. Following an intensive sampling protocol, samples were assayed for ARTS and its primary active metabolite, dihydroartemisinin (DHA), by liquid chromatography-mass spectrometry. A population pharmacokinetic model was developed to describe the data. Following administration of the first dose, the mean maximal concentrations of ARTS and DHA were 1,085 nmol/liter at 0.9 h and 2,525 nmol/liter at 2.3 h, respectively. The absorption half-life for ARTS was 2.3 h, and the conversion half-life (ARTS to DHA) was 0.27 h, while the elimination half-life of DHA was 0.71 h. The mean common volumes of distribution for ARTS and DHA relative to bioavailability were 42.8 and 2.04 liters/kg, respectively, and the mean clearance values relative to bioavailability were 6 and 2.2 liters/h/kg for ARTS and DHA, respectively. Substantial interpatient variability was observed, and the bioavailability of the second dose relative to that of the first was estimated to be 0.72. The covariates age, sex, and alpha thalassemia genotype were not influential in the pharmacokinetic model development; but the inclusion of weight as a covariate significantly improved the performance of the model. An ARTS suppositories dose of 10 of 20 mg/kg is appropriate for use in children with uncomplicated malaria. PMID- 15273108 TI - Relationship between triclosan and susceptibilities of bacteria isolated from hands in the community. AB - The possible association between triclosan and bacterial susceptibility to antibiotic was examined among staphylococci and several species of gram-negative bacteria (GNB) isolated from the hands of individuals in a community setting. Hand cultures from individuals randomized to using either antibacterial cleaning and hygiene products (including a hand soap containing 0.2% triclosan) or nonantibacterial cleaning and hygiene products for a 1-year period were taken at baseline and at the end of the year. Although there was no statistically significant association between triclosan MICs and susceptibility to antibiotic, there was an increasing trend in the association the odds ratios (ORs) for all species were compared at baseline (OR = 0.65, 95% confidence interval [95%CI] = 0.33 to 1.27) versus at the end of the year (OR = 1.08, 95%CI = 0.62 to 1.97) and for GNB alone at baseline(OR = 0.66, 95%CI = 0.29 to 1.51) versus the end of year (OR = 2.69, 95%CI = 0.78 to 9.23) regardless of the hand-washing product used. Moreover, triclosan MICs were higher in some of the species compared to earlier reports on household, clinical, and industrial isolates, and some of these isolates had triclosan MICs in the range of concentrations used in consumer products. The absence of a statistically significant association between elevated triclosan MICs and reduced antibiotic susceptibility may indicate that such a correlation does not exist or that it is relatively small among the isolates that were studied. Still, a relationship may emerge after longer-term or higher-dose exposure of bacteria to triclosan in the community setting. PMID- 15273109 TI - Role of positional hydrophobicity in the leishmanicidal activity of magainin 2. AB - The emergence of membrane-active antimicrobial peptides as new alternatives against pathogens with multiantibiotic resistance requires the design of better analogues. Among the different physicochemical parameters involved in the optimization of linear antimicrobial peptides, positional hydrophobicity has recently been incorporated. This takes into consideration the concept of the topological distribution of hydrophobic residues throughout the sequence rather than the classical concept of hydrophobicity as a global parameter of the peptide, calculated as the summation of the individual hydrophobicities of the residues. In order to assess the contribution of this parameter to the leishmanicidal mechanisms of magainin 2 analogues, the activities of two of these analogues, MG-H1 (GIKKFLHIIWKFIKAFVGEIMNS) and MG-H2 (IIKKFLHSIWKFGKAFVGEIMNI), which have similar charges, amino acid compositions, and hydrophobicities but different positional hydrophobicities, against Leishmania donovani promastigotes were assayed (T. Tachi, R. F. Epand, R. M. Epand, and K. Matsuzaki, Biochemistry 41:10723-10731, 2002). The activities were compared with that of the parental peptide, F5W-magainin 2 (GIGKWLHSAKKFGKAFVGEIMNS). The three peptides were active at micromolar concentrations, in the order MG-H2 > MG-H1 > F5W-magainin 2. These activities differ from their hemolytic and bactericidal activities. The results demonstrate that positional hydrophobicity, which reflects the presence of short stretches of sequences rich in hydrophobic amino acids, plays an important role in the activities of leishmanicidal peptides. PMID- 15273110 TI - Pharmacodynamic evaluation of the neutralization of endotoxin by PMX622 in mice. AB - Polymyxin B (PMB) binds to and neutralizes endotoxin, but its systemic clinical utility is limited by neuro- and nephrotoxicity. PMX622 is a covalent conjugate of PMB and Dextran-70 designed to retain the ability of PMB to neutralize endotoxin and to retain the favorable colloidal, pharmacokinetic, and metabolic properties of Dextran-70. PMX622 has demonstrated efficacy in a number of animal models and effectively neutralized endotoxin in phase I clinical trials. Here, we systematically evaluated the pharmacodynamic properties of PMX622 in a murine model of endotoxin-induced lethality in galactosamine-sensitized mice. PMX622 completely and dose dependently inhibited lethality in this model. A stoichiometric relationship was found between the endotoxin challenge dose and the dose of PMX622 needed for protection. PMX622 neutralized endotoxin from four different genera of gram-negative bacteria but not Neisseria meningitidis. PMX622 was significantly less toxic than PMB in the mouse, suggesting that PMX622 has a better margin of safety than PMB. The timing of PMX622 administration relative to endotoxin was crucial. PMX622 was active for several hours prior to the endotoxin challenge; however, PMX622 did not protect mice if administered >/=15 min after endotoxin challenge. This suggests that PMX622 would best be clinically used prophylactically rather than therapeutically. These studies will be crucial in designing and interpreting human clinical trials assessing PMX622 efficacy. PMID- 15273111 TI - Nucleotide and amino acid polymorphisms at drug resistance sites in non-B-subtype variants of human immunodeficiency virus type 1. AB - We have compared nucleotide substitutions and polymorphisms at codons known to confer drug resistance in subtype B strains of human immunodeficiency virus type 1 (HIV-1) with similar substitutions in viruses of other subtypes. Genotypic analysis was performed on viruses from untreated individuals. Nucleotide and amino acid diversity at resistance sites was compared with a consensus subtype B reference virus. Among patients with non-subtype B infections, polymorphisms relative to subtype B were observed at codon 10 in protease (PR). These included silent substitutions (CTC-->CTT, CTA, TTA) and an amino acid mutation, L10I. Subtype A viruses possessed a V179I substitution in reverse transcriptase (RT). Subtype G viruses were identified by silent substitutions at codon 181 in RT (TAT ->TAC). Similarly, subtype A/G viruses were identified by a substitution at position 67 in RT (GAC-->GAT). Subtype C was distinguished by silent substitutions at codons 106 (GTA-->GTG) and 219 (AAA-->AAG) in RT and codon 48 (GGG-->GGA) in PR. Variations relative to subtype B were seen at RT position 215 (ACC-->ACT) for subtypes A and A/E. These substitutions and polymorphisms reflect different patterns of codon usage among viruses of different subtypes. However, the existence of different subtypes may only rarely affect patterns of drug resistance-associated mutations. PMID- 15273112 TI - Comparison of the susceptibilities of Burkholderia pseudomallei to meropenem and ceftazidime by conventional and intracellular methods. AB - The effect of the two antibiotics ceftazidime and meropenem on a collection of 46 Burkholderia pseudomallei isolates representing clinical and environmental sources across northern Australia was investigated by using a series of in vitro test methods. The susceptibility testing methods used included Kirby-Bauer disk diffusion, Etest MIC, broth microdilution MIC, and a modification of the microdilution method in which Acanthamoeba cells were added to simulate the effect of a professional phagocytic cell on test outcome. In a semiquantitative validation coculture series, the majority of bacteria were intracellular up to a multiplicity of infection of 10 bacteria to one ameba. The optical density and bacterial count (log10 CFU/ml) correlated across the range tested (r2 = 0.77; P < 0.0001). Susceptibility test results were compared against clinical outcomes. The MICs of ceftazidime were consistently higher than those of meropenem by all three methods. The MICs of both agents were significantly higher when Acanthamoeba trophozoites were added to the broth microdilution method. Conventional and intracellular MIC results were consistent for clinical isolates from the Western Australian outbreak cluster despite the wide variety of clinical outcomes. Further development of the intracellular MIC method is expected to help assess the efficacy of antimicrobial agents on this bacterial species in an intracellular setting. PMID- 15273113 TI - Nitric oxide generated from isoniazid activation by KatG: source of nitric oxide and activity against Mycobacterium tuberculosis. AB - Isonicotinic acid hydrazide (INH) is a frontline antituberculosis agent. Once taken up by Mycobacterium tuberculosis, INH requires activation by the catalase peroxidase KatG, converting INH from its prodrug form into a range of bactericidal reactive species. Here we used 15N-labeled INH together with electron paramagnetic resonance spin trapping techniques to demonstrate that nitric oxide (NO*) is generated from oxidation at the hydrazide nitrogens during the activation of INH by M. tuberculosis KatG. We also observed that a specific scavenger of NO* provided protection against the antimycobacterial activity of INH in bacterial culture. No significant increases in mycobacterial protein nitration were detected, suggesting that NOdot; and not peroxynitrite, a nitrating metabolite of NO*, is involved in antimycobacterial action. In conclusion, INH-derived NO* has biological activity, which directly contributes to the antimycobacterial action of INH. PMID- 15273115 TI - Emergence of Streptococcus pneumoniae with very-high-level resistance to penicillin. AB - Penicillin resistance threatens the treatment of pneumococcal infections. We used sentinel hospital surveillance (1978 to 2001) and population-based surveillance (1995 to 2001) in seven states in the Active Bacterial Core surveillance of the Emerging Infections Program Network to document the emergence in the United States of invasive pneumococcal isolates with very-high-level penicillin resistance (MIC > or = 8 microg/ml). Very-high-level penicillin resistance was first detected in 1995 in multiple pneumococcal serotypes in three regions of the United States. The prevalence increased from 0.56% (14 of 2,507) of isolates in 1995 to 0.87% in 2001 (P = 0.03), with peaks in 1996 and 2000 associated with epidemics in Georgia and Maryland. For a majority of the strains the MICs of amoxicillin (91%), cefuroxime (100%), and cefotaxime (68%), were > or =8 microg/ml and all were resistant to at least one other drug class. Pneumonia (50%) and bacteremia (36%) were the most common clinical presentations. Factors associated with very highly resistant infections included residence in Tennessee, age of <5 or > or =65 years, and resistance to at least three drug classes. Hospitalization and case fatality rates were not higher than those of other pneumococcal infection patients; length of hospital stay was longer, controlling for age. Among the strains from 2000 and 2001, 39% were related to Tennessee(23F) 4 and 35% were related to England(14-)9. After the introduction of the pneumococcal conjugate vaccine, the incidence of highly penicillin resistant infections decreased by 50% among children <5 years of age. The emergence, clonality, and association of very-high-level penicillin resistance with multiple drug resistance requires further monitoring and highlights the need for novel agents active against the pneumococcus. PMID- 15273114 TI - Possible mechanism of miltefosine-mediated death of Leishmania donovani. AB - Miltefosine causes leishmanial death, but the possible mechanism(s) of action is not known. The mode of action of miltefosine was investigated in vitro in Leishmania donovani promastigotes as well as in extra- and intracellular amastigotes. Here, we demonstrate that miltefosine induces apoptosis-like death in L. donovani based on observed phenomena such as nuclear DNA condensation, DNA fragmentation with accompanying ladder formation, and in situ labeling of DNA fragments by the terminal deoxyribonucleotidyltransferase-mediated dUTP-biotin nick end labeling method. Understanding of miltefosine-mediated death will facilitate the design of new therapeutic strategies against Leishmania parasites. PMID- 15273116 TI - Activities of different fluoroquinolones against Bacillus anthracis mutants selected in vitro and harboring topoisomerase mutations. AB - Three sets of mutants of Bacillus anthracis resistant to fluoroquinolones were selected on ciprofloxacin and moxifloxacin in a stepwise manner from a nalidixic acid-resistant but fluoroquinolone-susceptible plasmidless strain harboring a Ser85Leu GyrA mutation. A high level of resistance to fluoroquinolones could be obtained in four or five selection steps. In each case, ParC was the secondary target. However, in addition to the GyrA mutation, expression of high-level resistance required (i) in the first set of mutants, active drug efflux associated with a mutation in the QRDR of ParC; (ii) in the second set, two mutations in the QRDR of ParC associated with a mutation in GyrB; and (iii) in the third set, two QRDR mutations, one in ParC and one in GyrA. Interestingly, several selection steps occurred without obvious mutations in the QRDR of any topoisomerase, thereby implying the existence of other resistance mechanisms. Among the fluoroquinolones tested, garenoxacin showed the best activity. PMID- 15273117 TI - Impact of specific pbp5 mutations on expression of beta-lactam resistance in Enterococcus faecium. AB - We tested the impact of individual PBP 5 mutations on expression of ampicillin resistance in Enterococcus faecium using a shuttle plasmid designed to facilitate expression of cloned pbp5 in ampicillin-susceptible E. faecium D344SRF. Substitutions that had been implicated in contributing to the resistance of clinical strains conferred only modest levels of resistance when they were present as single point mutations. The levels of resistance were amplified when some mutations were present in combination. In particular, a methionine-to alanine change at position 485 (in close proximity to the active site) combined with the insertion of a serine at position 466 (located in a loop that forms the outer edge of the active site) was associated with the highest levels of resistance to all beta-lactams. Affinity for penicillin generally correlated with beta-lactam MICs for the mutants, but these associations were not strictly proportional. PMID- 15273118 TI - Inhibitors of Leishmania mexicana CRK3 cyclin-dependent kinase: chemical library screen and antileishmanial activity. AB - The CRK3 cyclin-dependent kinase of Leishmania has been shown by genetic manipulation of the parasite to be essential for proliferation. We present data which demonstrate that chemical inhibition of CRK3 impairs the parasite's viability within macrophages, thus further validating CRK3 as a potential drug target. A microtiter plate-based histone H1 kinase assay was developed to screen CRK3 against a chemical library enriched for protein kinase inhibitors. Twenty seven potent CRK3 inhibitors were discovered and screened against Leishmania donovani amastigotes in vitro. Sixteen of the CRK3 inhibitors displayed antileishmanial activity, with a 50% effective dose (ED50) of less than 10 microM. These compounds fell into four chemical classes: the 2,6,9-trisubstituted purines, including the C-2-alkynylated purines; the indirubins; the paullones; and derivatives of the nonspecific kinase inhibitor staurosporine. The paullones and staurosporine derivatives were toxic to macrophages. The 2,6,9-trisubstituted purines inhibited CRK3 in vitro, with 50% inhibitory concentrations ranging from high nanomolar to low micromolar concentrations. The most potent inhibitors of CRK3 (compounds 98/516 and 97/344) belonged to the indirubin class; the 50% inhibitory concentrations for these inhibitors were 16 and 47 nM, respectively, and the ED50s for these inhibitors were 5.8 and 7.6 microM, respectively. In culture, the indirubins caused growth arrest, a change in DNA content, and aberrant cell types, all consistent with the intracellular inhibition of a cyclin dependent kinase and disruption of cell cycle control. Thus, use of chemical inhibitors supports genetic studies to confirm CRK3 as a validated drug target in Leishmania and provides pharmacophores for further drug development. PMID- 15273119 TI - Pharmacodynamics of telavancin (TD-6424), a novel bactericidal agent, against gram-positive bacteria. AB - Telavancin (TD-6424) is a novel lipoglycopeptide that produces rapid and concentration-dependent killing of clinically relevant gram-positive organisms in vitro. The present studies evaluated the in vivo pharmacodynamics of telavancin in the mouse neutropenic thigh (MNT) and mouse subcutaneous infection (MSI) animal models. Pharmacokinetic-pharmacodynamic studies in the MNT model demonstrated that the 24-h area under the concentration-time curve (AUC)/MIC ratio was the best predictor of efficacy. Telavancin produced dose-dependent reduction of thigh titers of several organisms, including methicillin-susceptible Staphylococcus aureus (MSSA) and methicillin-resistant Staphylococcus aureus (MRSA), penicillin-susceptible and -resistant strains of Streptococcus pneumoniae, and vancomycin-resistant Enterococcus faecalis. The 50% effective dose (ED50) estimates for telavancin ranged from 0.5 to 6.6 mg/kg of body weight (administered intravenously), and titers were reduced by up to 3 log10 CFU/g from pretreatment values. Against MRSA ATCC 33591, telavancin was 4- and 30-fold more potent (on an ED50 basis) than vancomycin and linezolid, respectively. Against MSSA ATCC 13709, telavancin was 16- and 40-fold more potent than vancomycin and nafcillin, respectively. Telavancin, vancomycin, and linezolid were all efficacious and more potent against MRSA ATCC 33591 in the MSI model compared to the MNT model. This deviation in potency was, however, disproportionately greater for vancomycin and linezolid than for telavancin, suggesting that activity of telavancin is less affected by the immune status. The findings of these studies collectively suggest that once-daily dosing of telavancin may provide an effective approach for the treatment of clinically relevant infections with gram positive organisms. PMID- 15273120 TI - Susceptibility to thrombin-induced platelet microbicidal protein is associated with increased fluconazole efficacy against experimental endocarditis due to Candida albicans. AB - Platelet microbicidal proteins (PMPs) are believed to be integral to host defense against endovascular infection. We previously demonstrated that susceptibility to thrombin-induced PMP 1 (tPMP-1) in vitro negatively influences Candida albicans virulence in the rabbit model of infective endocarditis (IE). This study evaluated the relationship between in vitro tPMP-1 susceptibility (tPMP-1s) or resistance (tPMP-1r) and efficacy of fluconazole (FLU) therapy of IE due to C. albicans. Candida IE was established in rabbits with either tPMP-1s or tPMP-1r strains. Treatment groups received FLU (100 mg/kg/day) intraperitoneally for 7 or 14 days; control animals received no therapy. At these time points, cardiac vegetations, kidneys, and spleens were quantitatively cultured to assess fungal burden. At both 7 and 14 days and in all target tissues, the extent of candidal clearance by FLU was greater in animals infected with the tPMP-1s strain than in those infected with the tPMP-1r strain. These differences were statistically significant in the spleen and kidney. Corroborating these in vivo data, FLU (a candidastatic agent), in combination with tPMP-1, exerted an enhanced fungicidal effect in vitro against tPMP-1s and tPMP-1r C. albicans, with the extent of this effect greatest against the tPMP-1s strain. Collectively, these results support the concept that tPMP-1 susceptibility contributes to the net efficacy of FLU against C. albicans IE in vivo, particularly in tissues in which platelets and tPMP-1 likely play significant roles in host defense. PMID- 15273121 TI - Regulation of the expression of cell wall stress stimulon member gene msrA1 in methicillin-susceptible or -resistant Staphylococcus aureus. AB - Genome-wide transcriptional profiling studies of the response of Staphylococcus aureus to cell wall-active antibiotics have led to the discovery of a cell wall stress stimulon of genes induced by these agents. msrA1, encoding methionine sulfoxide reductase, is a highly induced member gene of this stimulon. In the present study we show that msrA1 induction by oxacillin is common to all methicillin-susceptible strains studied but did not occur in two homogeneous and two heterogeneous methicillin-resistant strains. However, msrA1 was induced by vancomycin and/or D-cycloserine in methicillin-resistant strains. Lysozyme and lysostaphin treatment did not induce msrA1 expression. Oxacillin-induced msrA1 expression was enhanced by ca. 30% in a SigB+ derivative (SH1000) of the SigB defective RN450 (NCTC 8325-4) strain. msrA1 expression was not affected in mutants in the global regulatory systems agr and sar. Glycerol monolaurate, an inhibitor of signal transduction, inhibited the oxacillin-induced transcription of msrA1 and other cell wall stress stimulon member genes, vraS and dnaK. These observations suggest that the cell wall stress stimulon is induced by inhibition of the process of peptidoglycan biosynthesis, and the inhibitory effects of glycerol monolaurate indicate that gene expression is dependent on a signal transduction pathway. PMID- 15273122 TI - Comparison of gene expression profiles of Candida albicans azole-resistant clinical isolates and laboratory strains exposed to drugs inducing multidrug transporters. AB - Azole resistance in Candida albicans can be due to upregulation of multidrug transporters belonging to ABC (ATP-binding cassette) transporters (CDR1 and CDR2) or major facilitators (CaMDR1). Upregulation of these genes can also be achieved by exposure to fluphenazine, resulting in specific upregulation of CDR1 and CDR2 and by exposure to benomyl, resulting in specific CaMDR1 upregulation. In this study, these two different states of gene upregulation were used to determine coregulated genes that often share similar functions or similar regulatory regions. The transcript profiles of a laboratory strain exposed to these drugs were therefore determined and compared with those of two matched pairs of azole susceptible and -resistant strains expressing CDR1 and CDR2 (CDR strains) or CaMDR1 (MDR isolates). The results obtained revealed that, among 42 commonly regulated genes (8.6% of all regulated genes) between fluphenazine-exposed cells and CDR isolates, the most upregulated were CDR1 and CDR2 as expected, but also IFU5, RTA3 (which encodes putative membrane proteins), HSP12 (which encodes heat shock protein), and IPF4065 (which is potentially involved in stress response). Interestingly, all but HSP12 and IPF4065 contain a putative cis-acting drug responsive element in their promoters. Among the 57 genes (11.5% of all regulated genes) commonly regulated between benomyl-exposed cells and MDR isolates, the most upregulated were CaMDR1 as expected but also genes with oxido-reductive functions such as IFD genes, IPF5987, GRP2 (all belonging to the aldo-keto reductase family), IPF7817 [NAD(P)H oxido-reductase], and IPF17186. Taken together, these results show that in vitro drug-induced gene expression only partially mimics expression profiles observed in azole-resistant clinical strains. Upregulated genes in both drug-exposed conditions and clinical strains are drug resistance genes but also genes that could be activated under cell damage conditions. PMID- 15273123 TI - Novel polymorphisms in mec genes and a new mec complex type in methicillin resistant Staphylococcus aureus isolates obtained in rural Wisconsin. AB - We determined allelic polymorphisms in the mec complexes of 524 methicillin resistant Staphylococcus aureus isolates by partial or complete sequencing of three mec genes, mecA, mecI, and mecR1. The isolates had been collected over a 10 year period from patients living in rural Wisconsin, where the use of antibiotics was expected to be lower than in the bigger cities. Of the 18 mutation types identified, 16 had not been described previously. The five most common mutations were a mutation 7 nucleotides (nt) upstream from the start site (G-->T) in the mecA promoter (34.7%), an E246G change encoded by mecA (2.2%), a cytosine insertion at codon 257 in mecA (13.2%), an N121K change encoded by mecI (7.8%), and a major mecI-mecR1 deletion designated as a class B1 mec complex deletion type (25.4%). There was a high degree of conservation of the amino acid sequence of MecR1. Strains with the same mutations had variable resistance to oxacillin, and the median MIC for isolates that harbored the 7-nt-upstream mutation was lower than that for strains harboring other mutations. Our data suggest that the mecA promoter mutation plays a more important role in determining methicillin resistance than mutations in mecI and mecA do. Eighty-five percent of the tested isolates (n = 148) with the mecA promoter mutation and the class B1 mec complex deletion belonged to the same major clonal group, identified as MCG-2, and harbored the type IV staphylococcal cassette chromosome mec DNA. There was also a wide range of oxacillin MICs for strains with wild-type mecA, mecI, and mecR1 sequences, suggesting that the genetic backgrounds of clinical strains are significant in determining susceptibility to methicillin. PMID- 15273124 TI - Doripenem versus Pseudomonas aeruginosa in vitro: activity against characterized isolates, mutants, and transconjugants and resistance selection potential. AB - Doripenem is a broad-spectrum parenteral carbapenem under clinical development in Japan and North America. Its activities against (i) Pseudomonas aeruginosa isolates with graded levels of intrinsic efflux-type resistance, (ii) mutants with various combinations of AmpC and OprD expression, (iii) PU21 transconjugants with class A and D beta-lactamases, and (iv) P. aeruginosa isolates with metallo beta-lactamases were tested by the agar dilution method of the National Committee for Clinical Laboratory Standards. Selection of resistant P. aeruginosa mutants was investigated in single- and multistep procedures. Doripenem MICs for isolates without acquired resistance mostly were 0.12 to 0.5 microg/ml, whereas meropenem MICs were 0.25 to 0.5 microg/ml and imipenem MICs were 1 to 2 microg/ml. The MICs of doripenem, meropenem, ertapenem, and noncarbapenems for isolates with increased efflux-type resistance were elevated, whereas the MICs of imipenem were less affected. The MICs of doripenem were increased by the loss of OprD but not by derepression of AmpC; nevertheless, and as with other carbapenems, the impermeability-determined resistance caused by the loss of OprD corequired AmpC activity and was lost in OprD- mutants also lacking AmpC. The TEM, PSE, PER, and OXA enzymes did not significantly protect P. aeruginosa PU21 against the activity of doripenem, whereas MICs of > or =16 microg/ml were seen for clinical isolates with VIM and IMP metallo-beta-lactamases. Resistant mutants seemed to be harder to select with doripenem than with other carbapenems (or noncarbapenems), and the fold increases in the MICs were smaller for the resistant mutants. Single-step doripenem mutants were mostly resistant only to carbapenems and had lost OprD; multistep mutants had broader resistance, implying the presence of additional mechanisms, putatively including up-regulated efflux. Most mutants selected with aminoglycosides and quinolones had little or no cross-resistance to carbapenems, including doripenem. PMID- 15273125 TI - 1,2-dithiole-3-ones as potent inhibitors of the bacterial 3-ketoacyl acyl carrier protein synthase III (FabH). AB - The enzyme FabH catalyzes the initial step of fatty acid biosynthesis via a type II dissociated fatty acid synthase. The pivotal role of this essential enzyme, combined with its unique structural features and ubiquitous occurrence in bacteria, has made it an attractive new target for the development of antibacterial and antiparasitic compounds. We have searched the National Cancer Institute database for compounds bearing structural similarities to thiolactomycin, a natural product which exhibits a weak activity against FabH. This search has yielded several substituted 1,2-dithiole-3-ones that are potent inhibitors of FabH from both Escherichia coli (ecFabH) and Staphylococcus aureus (saFabH). The most potent inhibitor was 4,5-dichloro-1,2-dithiole-3-one, which had 50% inhibitory concentration (IC50) values of 2 microM (ecFabH) and 0.16 microM (saFabH). The corresponding 3-thione analog exhibited comparable activities. Analogs in which the 4-chloro substituent was replaced with a phenyl group were also potent inhibitors, albeit somewhat less effectively (IC50 values of 5.7 and 0.98 microM for ecFabH and saFabH, respectively). All of the 5 chlorinated inhibitors were most effective when they were preincubated with FabH in the absence of substrates. The resulting enzyme-inhibitor complex did not readily regain activity after excess inhibitor was removed, suggesting that a slow dissociation occurs. In stark contrast, a series of inhibitors in which the 5-chloro substituent was replaced with the isosteric and isoelectronic trifluoromethyl group were poorer inhibitors (IC50 values typically ranging from 25 to >100 microM for both ecFabH and saFabH), did not require a preincubation period for maximal activity, and generated an enzyme-inhibitor complex which readily dissociated. Possible modes of binding of 5-chloro-1,2-dithiole-3-ones and 5-chloro-1,2-dithiole-3-thiones with FabH which account for the role of the 5 chloro substituent were considered. PMID- 15273126 TI - Emergence in Italy of a Neisseria meningitidis clone with decreased susceptibility to penicillin. AB - A rise in invasive diseases due to Neisseria meningitidis C:2b:P1.5 with decreased penicillin susceptibility occurred in Italy during the last 2 years. Real-time PCR identified the Peni phenotype, and the penA sequence revealed the mosaicism of the gene. Molecular analyses assigned the isolates to a single emergent clone of the hypervirulent A4 cluster. PMID- 15273127 TI - In vitro activities of ravuconazole and four other antifungal agents against fluconazole-resistant or -susceptible clinical yeast isolates. AB - The activities of ravuconazole and four other antifungal agents were tested against a collection of 1,796 clinical yeast isolates, including fluconazole susceptible and -resistant strains. Ravuconazole was active against the majority of fluconazole-resistant isolates; but for 102 of 562 (18%) resistant isolates, mainly Candida tropicalis, Candida glabrata, and Cryptococcus neoformans, ravuconazole MICs were > or =1 microg/ml. PMID- 15273128 TI - Omiganan pentahydrochloride (MBI 226), a topical 12-amino-acid cationic peptide: spectrum of antimicrobial activity and measurements of bactericidal activity. AB - The activity of omiganan pentahydrochloride (formerly MBI 226; a synthetic cationic peptide) was assessed against 1,437 recent clinical bacterial isolates and 214 recent clinical yeast isolates. The omiganan was highly active, and minimal bactericidal concentrations or minimal fungicidal concentrations were either equal to or two- to fourfold higher than MICs. Kill curve experiments showed a clear pattern of bactericidal activity. PMID- 15273129 TI - Inhibitory activities of colicins against Escherichia coli strains responsible for postweaning diarrhea and edema disease in swine. AB - The efficacies of colicins E1 and N against Escherichia coli strains responsible for postweaning diarrhea and edema disease, two of the most prevalent disease problems for pigs in the United States, were determined in vitro. These proteins may provide an environmentally sound means for the prevention of these infections in swine. PMID- 15273130 TI - Distribution of human immunodeficiency virus type 1 protease and reverse transcriptase mutation patterns in 4,183 persons undergoing genotypic resistance testing. AB - In a sample of 6,156 sequences from 4,183 persons, the top 30 patterns of protease inhibitor, nucleoside reverse transcriptase (RT) inhibitor, and nonnucleoside RT inhibitor mutations accounted for 55, 46, and 66%, respectively, of sequences with drug resistance mutations. Characterization of the phenotypic and clinical significance of these common patterns may lead to improved treatment recommendations for a large proportion of patients for whom antiretroviral therapy is failing. PMID- 15273131 TI - In vitro activity and potency of an intravenously injected antimicrobial peptide and its DL amino acid analog in mice infected with bacteria. AB - We report that intravenous injection (3 mg/kg of body weight twice daily) of a diastereomer (containing 33% D amino acids) of an antimicrobial peptide, K6L9 (LKLLKKLLKKLLKLL-NH2), but not the all-L-amino-acid parental peptide, cures neutropenic mice infected with gentamicin-sensitive Pseudomonas aeruginosa and gentamicin-resistant Acinetobacter baumannii bacteria. Various biophysical experiments suggest a membranolytic-like effect. PMID- 15273132 TI - In vitro killing of Mycobacterium ulcerans by acidified nitrite. AB - Mycobacterium ulcerans, which causes Buruli ulcer, was exposed to acidified nitrite or to acid alone for 10 or 20 min. Killing was rapid, and viable counts were reduced below detectable limits within 10 min of exposure to 40 mM acidified nitrite. M. ulcerans is highly susceptible to acidified nitrite in vitro. PMID- 15273133 TI - Lack of activity of orally administered clofazimine against intracellular Mycobacterium tuberculosis in whole-blood culture. AB - The activity of oral clofazimine against intracellular Mycobacterium tuberculosis was compared to that of ofloxacin in healthy volunteers by the use of whole-blood cultures. Clofazimine was inactive whether it was tested alone or combined with other drugs that are used to treat multidrug-resistant tuberculosis, despite a total dose of 2 g. Kanamycin was the most active drug tested. PMID- 15273134 TI - Activities of doripenem (S-4661) against drug-resistant clinical pathogens. AB - Doripenem (formerly S-4661), a new 1-beta-methyl carbapenem, was challenged with a worldwide collection of 394 drug-refractory isolates. For endemic extended spectrum beta-lactamase- and stably derepressed AmpC-producing enteric bacilli, the doripenem MICs at which 90% of the isolates were inhibited (MIC90s) were 0.03 to 0.5 microg/ml, generally lower than those of comparator carbapenems. A greater proportion of strains among carbapenem-resistant nonfermentative gram-negative bacilli were inhibited by doripenem at < or =4 microg/ml, and doripenem was the most active carbapenem (MIC90, 1 to 4 microg/ml) against penicillin-resistant streptococci. PMID- 15273135 TI - Lack of evidence that DNA in antibiotic preparations is a source of antibiotic resistance genes in bacteria from animal or human sources. AB - Although DNA encoding antibiotic resistance has been discovered in antibiotic preparations, its significance for the development of antibiotic resistance in bacteria is unknown. No phylogenetic evidence was obtained for recent horizontal transfer of antibiotic resistance genes from antibiotic-producing organisms to bacteria from human or animal sources. PMID- 15273136 TI - Effect of pH on the in vitro activities of amphotericin B, itraconazole, and flucytosine against Aspergillus isolates. AB - The in vitro susceptibilities of 21 Aspergillus isolates were tested against three antifungal agents in RPMI 1640 and yeast nitrogen base at pH 5.0 and 7.0 by a broth microdilution format of the NCCLS method. The MICs of amphotericin B and itraconazole were higher, while those of flucytosine were lower, at pH 5.0 than at pH 7.0. The poor correlation between in vitro results and clinical outcome could be due to a difference in pH between the in vitro susceptibility test and at the site of infection. PMID- 15273137 TI - Pharmacokinetics of intravenous itraconazole in stable hemodialysis patients. AB - The pharmacokinetics of intravenous itraconazole (ITC) was studied in dialysis patients. Dialysis had no effect on the half-life and clearance of ITC or OH-ITC. However, dialysis allowed the clearance of hydroxypropyl-beta-cyclodextrin (HP beta-CD). The area under the concentration-time curve from time zero to infinity (AUC(0- infinity)) for HP-beta-CD administered before dialysis was lower than the AUC(0- infinity) when it was administered after dialysis (P < 0.01). Administration of ITC intravenously just prior to hemodialysis appears to produce adequate systemic exposures of ITC and OH-ITC while allowing dialysis clearance of HP-beta-CD. Studies of multiple administrations are warranted. PMID- 15273138 TI - Effect of granulocyte colony-stimulating factor combination therapy on efficacy of posaconazole (SCH56592) in an inhalation model of murine pulmonary aspergillosis. AB - Using an inhalation model of pulmonary aspergillosis, we observed modest differences in the survival rates of mice treated with granulocyte colony stimulating factor (G-CSF) and posaconazole (POS) and those treated with POS alone. This finding is in contrast to a previous report that suggested that G-CSF had a significant antagonistic effect on the antifungal activity of POS. PMID- 15273139 TI - High prevalence of extended-spectrum beta-lactamase-producing strains among blood isolates of Enterobacter spp. collected in a tertiary hospital during an 8-year period and their antimicrobial susceptibility patterns. AB - Of 72 blood isolates of Enterobacter spp. collected over an 8-year period, 50% (36 of 72) were derepressed or partially derepressed AmpC mutants. The extended spectrum beta-lactamase (ESBL) production rate was 43% (31 of 72 isolates), and 67.3% (31 of 46) of extended-spectrum cephalosporin-resistant strains produced ESBLs. Thus, a confirmatory test for ESBL production is necessary for extended spectrum cephalosporin-resistant Enterobacter spp. PMID- 15273140 TI - Temporin A soaking in combination with intraperitoneal linezolid prevents vascular graft infection in a subcutaneous rat pouch model of infection with Staphylococcus epidermidis with intermediate resistance to glycopeptides. AB - The efficacy of linezolid and temporin A in the prevention of prosthetic graft infection due to methicillin-resistant Staphylococcus epidermidis with intermediate resistance to glycopeptides was investigated in a subcutaneous rat pouch model. Linezolid and temporin A, alone or combined, greatly reduced the bacterial numbers compared to the effect with control drugs. PMID- 15273141 TI - In vitro activities of the newer quinolones garenoxacin, gatifloxacin, and gemifloxacin against human mycoplasmas. AB - The activities of garenoxacin, gatifloxacin, and gemifloxacin were compared with those of four fluoroquinolones against human mycoplasmas and ureaplasmas, including fluoroquinolone-resistant genetically characterized strains. Garenoxacin exhibited the highest activity, followed by gemifloxacin, moxifloxacin, and gatifloxacin. The minimal bactericidal activities of these three compounds were lower than those of the four fluoroquinolones. PMID- 15273142 TI - In vitro activities of telithromycin, linezolid, and quinupristin-dalfopristin against Streptococcus pneumoniae with macrolide resistance due to ribosomal mutations. AB - To date, 86 of 7,746 macrolide-resistant Streptococcus pneumoniae isolates from 1999 to 2002 PROTEKT (Prospective Resistant Organism Tracking and Epidemiology for the Ketolide Telithromycin) surveillance studies were negative for methylase and efflux mechanisms. Mutations in 23S rRNA or the genes encoding riboprotein L4 or L22 were found in 77 of 86 isolates. Six isolates were resistant to quinupristin-dalfopristin and two were resistant to linezolid, while telithromycin demonstrated good activities against all isolates. PMID- 15273143 TI - CMY-13, a novel inducible cephalosporinase encoded by an Escherichia coli plasmid. AB - An IncN plasmid (p541) from Escherichia coli carried a Citrobacter freundii derived sequence of 4,252 bp which included an ampC-ampR region and was bound by two directly repeated IS26 elements. ampC encoded a novel cephalosporinase (CMY 13) with activity similar to that of CMY-2. AmpR was likely functional as indicated in induction experiments. PMID- 15273144 TI - Rv2686c-Rv2687c-Rv2688c, an ABC fluoroquinolone efflux pump in Mycobacterium tuberculosis. AB - The Mycobacterium tuberculosis Rv2686c-Rv2687c-Rv2688c operon, encoding an ABC transporter, conferred resistance to ciprofloxacin and, to a lesser extent, norfloxacin, moxifloxacin, and sparfloxacin to Mycobacterium smegmatis. The resistance level decreased in the presence of the efflux pump inhibitors reserpine, carbonyl cyanide m-chlorophenylhydrazone, and verapamil. Energy dependent efflux of ciprofloxacin from M. smegmatis cells containing the Rv2686c Rv2687c-Rv2688c operon was observed. PMID- 15273145 TI - Extended-spectrum-cephalosporin resistance in Salmonella enterica isolates of animal origin. AB - A total of 112 out of 5,709 Salmonella enterica isolates from domestic animal species exhibited decreased susceptibilities to ceftiofur and ceftriaxone, and each possessed the blaCMY gene. Ten Salmonella serotypes were significantly more likely to include resistant isolates. Isolates from turkeys, horses, cats, and dogs were significantly more likely to include resistant isolates. PMID- 15273146 TI - A lipid-peptide microbicide inactivates herpes simplex virus. AB - A microbicide combining the lipid-ether 1-0-octyl-sn-glycerol (OG; 3 mM) and peptide D2A21 (9 microM) reduced herpes simplex virus type 1 (HSV-1) and HSV-2 titers by at least 1,000-fold, more than the sum of the inactivations produced by OG and D2A21 alone. OG plus D2A21 reduced HSV-1 and HSV-2 titers by > or =1,000 fold in < or =10 and < or =20 min, respectively, whereas OG and D2A21 used alone produced almost no virus inactivation during these times. PMID- 15273147 TI - Remarkable increase in central Japan in 2001-2002 of Neisseria gonorrhoeae isolates with decreased susceptibility to penicillin, tetracycline, oral cephalosporins, and fluoroquinolones. AB - Four hundred sixty-two clinical isolates of Neisseria gonorrhoeae recovered from 1999 through 2002 in central Japan were examined for MICs of antimicrobial agents. The majority was sensitive to ceftriaxone and spectinomycin, but a remarkable increase in isolates with decreased susceptibility to penicillin, tetracycline, oral cephalosporins, and fluoroquinolones was observed from 2001 through 2002. PMID- 15273148 TI - Antianaerobic activity of a novel fluoroquinolone, WCK 771, compared to those of nine other agents. AB - Agar dilution MIC methodology was used to compare the activity of WCK 771 with those of ciprofloxacin, levofloxacin, moxifloxacin, gatifloxacin, piperacillin, piperacillin-tazobactam, imipenem, clindamycin, and metronidazole against 350 anaerobes. Overall, the MICs (in micrograms per milliliter) at which 50 and 90%, respectively, of the isolates tested were inhibited were as follows: WCK 771, 0.5 and 2.0; ciprofloxacin, 2.0 and 32.0; levofloxacin, 1.0 and 8.0; gatifloxacin, 0.5 and 4.0; moxifloxacin, 0.5 and 4.0; piperacillin, 2.0 and 64.0; piperacillin tazobactam, < or =0.125 and 8.0; imipenem, 0.125 and 1.0; clindamycin, 0.125 and 16.0; and metronidazole, 1.0 and >16.0. PMID- 15273149 TI - High prevalence of the ermB gene among erythromycin-resistant streptococcus pneumoniae isolates in Germany during the winter of 2000-2001 and in vitro activity of telithromycin. AB - Of 595 isolates of Streptococcus pneumoniae from outpatients with respiratory tract infections, collected from 17 microbiology laboratories, 14.1% were resistant to erythromycin. Eighty-three erythromycin-resistant isolates were genetically analyzed, 83.1% of which harbored the ermB gene. Only four isolates (4.8%) harbored the mefA gene. Telithromycin exhibited potent activity against all isolates. PMID- 15273150 TI - Novel antimalarial compounds isolated in a survey of self-medicative behavior of wild chimpanzees in Uganda. AB - Following a veterinary and behavioral survey of chimpanzees from a natural population in Uganda, leaf samples of Trichilia rubescens were collected because of the unusual method of ingestion observed. The methanolic crude extract of T. rubescens leaves exhibited significant antimalarial activity in vitro. Bioassay directed fractionation provided two new limonoids, trichirubines A and B. A greater understanding of the role of secondary compounds in the primate diet may be helpful in recovering naturally occurring compounds of medicinal significance for human medicine. PMID- 15273151 TI - Susceptibility of Chlamydia trachomatis to excipients commonly used in topical microbicide formulations. AB - Commonly used "inactive" pharmaceutical excipients were tested in a previously developed minimum cidal concentration assay to assess their ability to kill Chlamydia trachomatis topically. Sixteen excipients were evaluated in these studies under various conditions. A range of activities was found among the excipients that could be tested in our assay system. PMID- 15273152 TI - Role of beta-lactamases and porins in resistance to ertapenem and other beta lactams in Klebsiella pneumoniae. AB - High-level resistance to ertapenem was produced by beta-lactamases of groups 1, 2f, and 3 in a strain of Klebsiella pneumoniae deficient in Omp35 and Omp36. From a wild-type strain producing ACT-1 beta-lactamase, ertapenem-resistant mutants for which the ertapenem MICs were up to 128 microg/ml and expression of outer membrane proteins was diminished could be selected. PMID- 15273153 TI - nimE gene in a metronidazole-susceptible Veillonella sp. strain. PMID- 15273154 TI - Spectrum of pncA mutations in multidrug-resistant Mycobacterium tuberculosis isolates obtained in Latvia. PMID- 15273155 TI - Challenges in designing animal studies to detect antagonism of polyene activity. PMID- 15273159 TI - the JigCell model builder and run manager. AB - SUMMARY: We describe the JigCell Model Builder (JCMB), a tool for creating biochemical reaction network models. JCMB is designed for ease of use and its interface uses the standard spreadsheet metaphor. The JigCell Run Manager (JCRM) is a tool for organizing the large collections of simulation runs typically required by reaction network modeling activities. AVAILABILITY: JCMB and JCRM are part of the JigCell suite available at http://jigcell.biol.vt.edu. PMID- 15273160 TI - Identification of potential candidates for varicella vaccination by history: questionnaire and seroprevalence study. PMID- 15273162 TI - Blinded by the light. PMID- 15273163 TI - Extracorporeal shock wave therapy for plantar fasciitis: randomised controlled multicentre trial. PMID- 15273164 TI - Anterior cruciate ligament rupture: is osteoarthritis inevitable? PMID- 15273165 TI - Effect of arthrographic shoulder joint distension with saline and corticosteroid for adhesive capsulitis. PMID- 15273166 TI - Postactivation potentiation: role in performance. PMID- 15273168 TI - A survey of flexibility training protocols and hamstring strains in professional football clubs in England. AB - OBJECTIVES: To investigate the relation between current flexibility training protocols, including stretching, and hamstring strain rates (HSRs) in English professional football clubs. METHOD: Questionnaire based data on flexibility training methods and HSRs were collected from 30 English professional football clubs in the four divisions during the 1998/99 season. Data were coded and analysed using cross tabulation, correlation, and multiple regression. RESULTS: Flexibility training protocols were characterised by wide variability, with static stretching the most popular stretching technique used. Hamstring strains represented 11% of all injuries and one third of all muscle strains. About 14% of hamstring strains were reinjuries. HSRs were highest in the Premiership (13.3 (9.4)/1000 hours) with the lowest rates in Division 2 (7.8 (2.9)/1000 hours); values are mean (SD). Most (97%) hamstring strains were grade I and II, two thirds of which occurred late during training/matches. Forwards were injured most often. Use of the standard stretching protocol (SSP) was the only factor significantly related to HSR (r = -0.45, p = 0.031) in the correlation analysis, suggesting that the more SSP is used, the lower the HSR. About 80% of HSR variability was accounted for by stretching holding time (SHT), SSP, and stretching technique (STE) in the multiple regression equation: HSR = 37.79 - (0.33SHT + [corrected] 10.05SSP + 2.24STE) +/- 2.34. SHT (negatively correlated with HSR) was the single highest predictor, and accounted for 30% of HSR variability, and an additional 40% in combination with SSP. CONCLUSIONS: Flexibility training protocols in the professional clubs were variable and appeared to depend on staffing expertise. Hamstring stretching was the most important training factor associated with HSR. The use of SSP, STE, and SHT are probably involved in a complex synergism which may reduce hamstring strains. Modification of current training patterns, especially stretching protocols, may reduce HSRs in professional footballers. PMID- 15273169 TI - A pilot study of the eccentric decline squat in the management of painful chronic patellar tendinopathy. AB - OBJECTIVES: This non-randomised pilot study investigated the effect of eccentric quadriceps training on 17 patients (22 tendons) with painful chronic patellar tendinopathy. METHODS: Two different eccentric exercise regimens were used by subjects with a long duration of pain with activity (more than six months). (a) Nine consecutive patients (10 tendons; eight men, one woman; mean age 22 years) performed eccentric exercise with the ankle joint in a standard (foot flat) position. (b) Eight patients (12 tendons; five men, three women; mean age 28 years) performed eccentric training standing on a 25 degrees decline board, designed to increase load on the knee extensor mechanism. The eccentric training was performed twice daily, with three sets of 15 repetitions, for 12 weeks. Primary outcome measures were (a) 100 mm visual analogue scale (VAS), where the subject recorded the amount of pain during activity, and (b) return to previous activity. Follow up was at 12 weeks, with a further limited follow up at 15 months. RESULTS: Good clinical results were obtained in the group who trained on the decline board, with six patients (nine tendons) returning to sport and showing a significantly reduced amount of pain over the 12 week period. Mean VAS scores fell from 74.2 to 28.5 (p = 0.004). At 15 months, four patients (five tendons) reported satisfactory results (mean VAS 26.2). In the standard squat group the results were poor, with only one athlete returning to previous activity. Mean VAS scores in this group were 79.0 at baseline and 72.3 at 12 weeks (p = 0.144). CONCLUSION: In a small group of patients with patellar tendinopathy, eccentric squats on a decline board produced encouraging results in terms of pain reduction and return to function in the short term. Eccentric exercise using standard single leg squats in a similar sized group appeared to be a less effective form of rehabilitation in reducing pain and returning subjects to previous levels of activity. PMID- 15273170 TI - Do team gymnasts compete in spite of symptoms from an injury? AB - BACKGROUND: Gymnasts practise many hours a week, and symptoms from injuries do not seem to stop them from continuing with practice. They may even compete with symptoms from injuries, which could increase the risk of reinjury, or of the occurrence of a more severe injury. OBJECTIVES: To investigate whether team gymnasts compete at high level in spite of symptoms from an injury. METHODS: 188 male and female competitors participating in the Swedish Cup for juniors and seniors answered a questionnaire about symptoms from injuries on the day of the competition. RESULTS: More than half the gymnasts (58%) competed despite having symptoms from an injury on the day of the competition. More seniors than juniors competed in spite of symptoms from an injury (p = 0.006). Two of three team gymnasts (65%) reported symptoms from the lower extremities and around one in five (22%) reported back symptoms. Fifty five per cent of the gymnasts reported recurrence of an injury at the same site (reinjury). CONCLUSIONS: There was a high prevalence of symptoms from injuries on the day of competition. This did not stop the team gymnasts from competing. PMID- 15273171 TI - Off seasonal and pre-seasonal assessment of circulating energy sources during prolonged running at the anaerobic threshold in competitive triathletes. AB - OBJECTIVES: To compare changes in circulating energy sources during prolonged exercise in off season (OS) and pre-season (PS) training of triathletes. METHODS: Nine athletes of the Swiss national triathlon team (three female, mean (SD) age 28.7 (4.9) years, height 169.8 (6.0) cm, weight 57.0 (6.2) kg, VO(2)max 66.5 (5.3) ml/min/kg; six male, mean (SD) age 24.0 (4.1) years, height 181.4 (6.9) cm, weight 73.5 (6.0) kg, VO(2)max 75.9 (4.9) ml/min/kg) were tested twice (2.5 months apart) during a 25 km aerobic capacity test run at the end of the OS and just before the season. The average training load during the OS was 9.9 h/week, and this increased to 14.4 h/week in the PS. With heart rates as reference, exercise intensity during the aerobic capacity test was 97.0 (4.9)% of the anaerobic threshold and 91.2 (4.5)% of VO(2)max. Blood samples were collected before, during, and after the aerobic capacity test. Samples were collected every 5 km during three minute rest intervals. RESULTS: Blood was analysed for triglyceride (TG), free fatty acids, cholesterol, high density lipoprotein cholesterol, glucose, insulin, lactate, and changes in plasma volume. A two factor (season by distance) repeated measures analysis of variance revealed an increase in capacity for prolonged exercise in the PS by a decrease in running intensity during the aerobic capacity test (% of speed at 2.0 mmol/l lactate threshold, p = 0.008), an increase in running speed at the anaerobic threshold (p = 0.003) and at 4.0 and 2.0 mmol/l (p<0.001) of the lactate threshold. A significant season by distance interaction was found for TG (p<0.001). TG concentrations peaked at 5 km and decreased logarithmically throughout the OS (1.48 (0.34) to 0.86 (0.20) mmol/l) and PS (1.90 (0.31) to 0.73 (0.18) mmol/l) tests. From the OS to the PS, there was an increase in the difference in TG at 5 15 km with a concomitant increase at 2.0 mmol/l of the lactate threshold. The peak TG concentrations at 5 km followed by a logarithmic decrease suggest that TG may also provide circulating energy. A greater logarithmic decrease in TG occurred in the PS than in the OS, indicating a higher rate of use. There was an increase in the difference in TG at 5-15 km similar to the increase in the speed at 2.0 mmol/l of the lactate threshold between the two seasons. Glucose, insulin, lactate, and free fatty acids were similar in the two seasons. CONCLUSION: Free fatty acid and TG concentrations were much higher than expected, and the two training seasons showed significantly different patterns of TG concentration during prolonged running. These responses may be related to aerobic capacity of prolonged exercise. PMID- 15273172 TI - Master's performance in the New York City Marathon 1983-1999. AB - BACKGROUND: Physical activity in older people is believed to slow down the natural aging process through its effects on disuse atrophy. OBJECTIVES: To show that elite master (age above 50) male and female athletes, as a group, have improved their running times over the last two decades at a greater rate than their younger counterparts. METHODS: Running time, age, and sex of all 415,000 runners in the New York City Marathon from 1983 to 1999 were examined using linear regression analysis. RESULTS: The number of master participants increased at a greater rate than their younger counterparts (p<0.05). Running times for the top 50 male and female finishers over the past two decades showed significantly greater improvement in the master groups than in the younger age groups (p<0.001). CONCLUSIONS: Participation in the New York City Marathon is increasing at a higher rate in the master groups than in other age groups. Male and female masters continued to improve running times at a greater rate than the younger athletes, whose performance levels have plateaued. This is the largest study to compare master athletic performance with younger counterparts and men with women. PMID- 15273173 TI - Shoulder pathoanatomy in marathon kayakers. AB - OBJECTIVES: To determine the prevalence of soft and hard tissue abnormalities and their interrelations in the shoulders of marathon kayakers and to examine the pathoanatomical factors that predispose these athletes to injury. METHODS: Fifty two long distance kayakers completed a questionnaire. Their shoulders were examined for range of motion, pain, and stability using a standard set of 10 clinical tests. The shoulder was subsequently scanned by magnetic resonance imaging (MRI) in three planes and evaluated for evidence of injury or other abnormality. The relation of clinical symptoms and MRI findings was investigated with respect to kayaker's age, number of years kayaking, and number of marathon races completed. RESULTS: Thirty subjects were asymptomatic at the time of scanning, and twenty two showed symptoms of pain and/or instability. MRI showed acromioclavicular hypertrophy, acromial or clavicular spur, supraspinatus tendinitis, and partial tear of the supraspinatus as the most common abnormalities. Kayaker's age, number of years kayaking, and number of races completed did not relate significantly to symptoms or to the presence of an abnormality on MRI scan. Of all the pathoanatomical findings that are reported to predispose to rotator cuff injury, only acromial and clavicular spurs were found to correlate highly with supraspinatus muscle pathology. CONCLUSIONS: Rotator cuff injuries make up a large portion of the injuries seen in marathon kayakers, about twice the number reported for sprint kayakers. These injuries are the result of secondary impingement factors associated with overuse, possibly specific to kayakers, and not the result of bony restrictions around the shoulder joint. Acromioclavicular hypertrophy is a common finding in marathon kayakers, but is possibly the result of portaging or a previous injury. PMID- 15273174 TI - Mutations in the hereditary haemochromatosis gene HFE in professional endurance athletes. AB - BACKGROUND: Hereditary haemochromatosis, a disease that affects iron metabolism, progresses with a greater or lesser tendency to induce iron overload, possibly leading to severe organ dysfunction. Most elite endurance athletes take iron supplements during their active sporting life, which could aggravate this condition. OBJECTIVE: To determine the prevalence and discuss potential clinical implications of mutations of HFE (the gene responsible for hereditary haemochromatosis) in endurance athletes. METHODS: Basal concentrations of iron, ferritin, and transferrin and transferrin saturation were determined in the period before competition in 65 highly trained athletes. Possible mutations in the HFE gene were evaluated in each subject by extracting genomic DNA from peripheral blood. The restriction enzymes SnaBI and BclI were used to detect the mutations 845G-->A (C282Y) and 187C-->G (H63D). RESULTS: Our findings indicate a high prevalence of HFE gene mutations in this population (49.2%) compared with sedentary controls (33.5%). No association was detected in the athletes between mutations and blood iron markers. CONCLUSIONS: The findings support the need to assess regularly iron stores in elite endurance athletes. PMID- 15273176 TI - Evaluation of physiological standard pressures of the forearm flexor muscles during sport specific ergometry in sport climbers. AB - BACKGROUND: Chronic exertional compartment syndromes (CECS) are well known in sports medicine. Most commonly affected is the tibialis anterior muscle compartment in runners and walkers. Only a few cases of CECS of the forearm flexor muscles have been reported. OBJECTIVES: To determine pressure levels inside the deep flexor compartment of the forearms during a sport specific stress test. METHOD: Ten healthy, high level climbers were enrolled in a prospective study. All underwent climbing specific ergometry, using a rotating climbing wall (step test, total climbing time 9-15 minutes). Pressure was measured using a slit catheter placed in the deep flexor compartment of the forearm. Pressure, blood lactate, and heart rate were recorded every three minutes and during recovery. RESULTS: In all the subjects, physical exhaustion of the forearms defined the end point of the climbing ergometry. Blood lactate increased with physical stress, reaching a mean of 3.48 mmol/l. Compartment pressure was related to physical stress, exceeding 30 mm Hg in only three subjects. A critical pressure of more than 40 mm Hg was never observed. After the test, the pressure decreased to normal levels within three minutes in seven subjects. The three with higher pressure levels (>30 mm Hg) required a longer time to recover. CONCLUSIONS: For further clinical and therapeutic consequences, an algorithm was derived. Basic pressure below 15 mm Hg and stress pressure below 30 mm Hg as well as pressures during the 15 minute recovery period below 15 mm Hg are physiological. Pressures of 15-30 mm Hg during recovery suggest high risk of CECS, and pressures above 30 mm Hg confirm CECS. PMID- 15273177 TI - Community football players' attitudes towards protective equipment--a pre-season measure. AB - BACKGROUND: The Australian football injury prevention project (AFIPP) was a randomised controlled trial examining the effects of protective equipment on injury rates in Australian Football. OBJECTIVE: To present the results of the AFIPP baseline survey of community football players' attitudes towards protective equipment. METHODS: Teams of players were recruited from the largest community football league in Victoria, Australia, during the 2001 playing season; 301 players were enrolled in the study and all were surveyed before the season began about their attitudes towards protective headgear and mouthguards. RESULTS: Almost three quarters of the players (73.6%) reported wearing mouthguards during the previous playing season (year 2000) compared with only 2.1% wearing headgear. The most common reasons for not wearing headgear and mouthguards (in non-users) were: "I don't like wearing it" (headgear: 44.8%; mouthguards: 30.6%), and "It is too uncomfortable" (headgear: 40.7%; mouthguards: 45.8%). CONCLUSIONS: The higher mouthguard usage reflects the favourable attitudes towards mouthguards by Australian football players generally. Similarly, the low headgear usage reflects the low acceptance of this form of protection in this sport. Further research should be directed towards establishing the reasons why players seem to believe that headgear plays a role in injury prevention yet few wear it. PMID- 15273178 TI - A double blind, randomised, parallel group study on the efficacy and safety of treating acute lateral ankle sprain with oral hydrolytic enzymes. AB - OBJECTIVE: To compare the effectiveness and safety of the triple combination Phlogenzym (rutoside, bromelain, and trypsin) with double combinations, the single substances, and placebo. DESIGN: Multinational, multicentre, double blind, randomised, parallel group design with eight groups structured according to a factorial design. SETTING: Orthopaedic surgery and emergency departments in 27 European hospitals. PARTICIPANTS: A total of 721 patients aged 16-53 years presenting with acute unilateral sprain of the lateral ankle joint. PRIMARY EFFICACY CRITERIA: (a) Pain on walking one or two steps, as defined by the patient on a visual analogue scale. (b) The range of motion, as measured by the investigator and expressed as a sum of flexion and extension. (c) The volume of the injured ankle measured with a volometer. RESULTS: At the primary end point at seven days, the greatest reduction in pain was in the bromelain/trypsin group (73.7%). The Phlogenzym group showed a median reduction of 60.3%, and the placebo group showed a median reduction of 73.3%. The largest increase in range of motion (median) was in the placebo group (60% change from baseline). The Phlogenzym group showed a median increase of 42.9%. The biggest decrease in swelling was in the trypsin group (3.9% change from baseline). The Phlogenzym group showed a 2.30% change from baseline and the placebo group a -2.90% change. In the subgroup analysis of patients who did not use a Caligamed brace, Phlogenzym was superior to placebo for the summarising directional test of the primary efficacy criteria (MW = 0.621; LB-CI 0.496; p = 0.029; one sided Wei-Lachin procedure). The vast majority of doctors and patients rated the tolerability of all treatments tested as very good or at least good. CONCLUSIONS: Phlogenzym was not found to be superior to the three two-drug combinations, the three single substances, or placebo for treatment of patients with acute unilateral sprain of the lateral ankle joint. The small subgroup of patients treated without the support of a Caligamed brace showed evidence of superiority of Phlogenzym over placebo. Further research is warranted to study this effect of Phlogenzym in patients treated without ankle support. PMID- 15273179 TI - Effect of incorporating low intensity exercise into the recovery period after a rugby match. AB - BACKGROUND: The psychological and physiological condition of athletes affect both their performance in competitions and their health. Rugby is an intense sport which appears to impose psychological and physiological stress on players. However, there have been few studies of the most appropriate resting techniques to deliver effective recovery from a match. OBJECTIVES: To compare the difference in recovery after a match using resting techniques with or without exercise. METHODS: Fifteen Japanese college rugby football players were studied. Seven performed only normal daily activities and eight performed additional low intensity exercise during the post-match rest period. Players were examined just before and immediately after the match and one and two days after the match. Blood biochemistry and two neutrophil functions, phagocytic activity and oxidative burst, were measured to assess physiological condition, and the profile of mood states (POMS) scores were examined to evaluate psychological condition. RESULTS: Immediately after the match, muscle damage, decreases in neutrophil functions, and mental fatigue were observed in both groups. Muscle damage and neutrophil functions recovered with time almost equally in the two groups, but the POMS scores were significantly decreased only in subjects in the low intensity exercise group. CONCLUSIONS: Rugby matches impose both physiological and psychological stress on players. The addition of low intensity exercise to the rest period did not adversely affect physiological recovery and had a significantly beneficial effect on psychological recovery by enhancing relaxation. PMID- 15273181 TI - Risk factors associated with exertional medial tibial pain: a 12 month prospective clinical study. AB - OBJECTIVE: To investigate in a military setting the potential role of intrinsic biomechanical and anthropometric risk factors for, and the incidence of, exertional medial tibial pain (EMTP). METHODS: A prospective clinical outcome study in a cohort of 122 men and 36 women at the Australian Defence Force Academy. Each cadet underwent measurements of seven intrinsic variables: hip range of motion, leg length discrepancy, lean calf girth, maximum ankle dorsiflexion range, foot type, rear foot alignment, and tibial alignment. Test retest reliability was undertaken on each variable. A physician recorded any cadet presenting with diagnostic criteria of EMTP. Records were analysed at 12 months for EMTP presentation and for military fitness test results. RESULTS: 23 cadets (12 men, 11 women) met the criteria for EMTP after 12 months, with a cross gender (F/M) odds ratio of 3.1. In men, both internal and external range of hip motion was greater in those with EMTP: left internal (12 degrees, p = 0.000), right internal (8 degrees, p = 0.014), left external (8 degrees, p = 0.042), right external (9 degrees, p = 0.026). Lean calf girth was lower by 4.2% for the right leg (p = 0.040) but by only 2.9% for the left leg (p = 0.141). No intrinsic risk factor was associated with EMTP in women. EMTP was the major cause for non completion of the run component of the ADFA fitness test in both men and women. CONCLUSIONS: Greater internal and external hip range of motion and lower lean calf girth were associated with EMTP in male military cadets. Women had high rates of injury, although no intrinsic factor was identified. Reasons for this sex difference need to be identified. PMID- 15273182 TI - Clinical examination of athletes with groin pain: an intraobserver and interobserver reliability study. AB - BACKGROUND: Groin pain is a diagnostic and therapeutic challenge to sports medicine. The literature provides no consensus on definitions of or diagnostic criteria for groin pain in athletes. To compare the results of research and treatments, the methods used to diagnose and evaluate the degree of groin pain must be clearly defined and reproducible. OBJECTIVES: To describe clinical examination techniques for groin pain in athletes and evaluate the intraobserver and interobserver reliability of these. METHODS: Eighteen athletes, nine with sports related groin pain and nine without groin pain, were examined by two doctors and two physiotherapists. The examiners were trained in the examination techniques before the study. The examiners were blinded to the symptoms and identity of the subjects. The subjects were examined twice by each examiner in random order. The examinations included evaluation of adductor muscle related pain and strength, iliopsoas muscle related pain, strength, and flexibility, abdominal muscle related pain, and strength and pain at the symphysis joint. Kappa statistics and percentage of agreement were used to evaluate the data. RESULTS: Overall, the kappa values and percentage of agreement were in accordance and showed good reliability of the examinations. The kappa values for the intraobserver agreement were above 0.60 in 11 of 14 tests, and those for the interobserver agreement of the pain tests were above 0.60 in eight of 10 tests. The only test without acceptable interobserver reliability was the strength test for iliopsoas muscle. CONCLUSION: All but one of the tests investigated were reproducible and subject only to limited intraobserver and interobserver variation. PMID- 15273183 TI - Left ventricular systolic function and diastolic filling after intermittent high intensity team sports. AB - BACKGROUND: Prolonged steady state exercise can lead to a decrease in left ventricular (LV) function as well as promote the release of cardiac troponin T (cTnT). There is limited information on the effect of intermittent high intensity exercise of moderate duration. OBJECTIVES: To determine the effect of intermittent high intensity exercise of moderate duration on LV function. METHODS: Nineteen male rugby and football players (mean (SD) age 21 (2) years) volunteered. Assessments, before, immediately after, and 24 hours after competitive games, included body mass, heart rate (HR), and systolic blood pressure (sBP) as well as echocardiography to assess stroke volume (SV), ejection fraction (EF), systolic blood pressure/end systolic volume ratio (sBP/ESV), and global diastolic filling (E:A) as well as to indirectly quantify preload (LV internal dimension at end diastole (LVIDd)). Serum cTnT was analysed using a 3rd generation assay. Changes in LV function were analysed by repeated measures analysis of variance. cTnT data are presented descriptively. RESULTS: SV (91 (26) v 91 (36) v 90 (35) ml before, after, and 24 hours after the game respectively), EF (71 (8) v 70 (9) v 71 (7)%), and sBP/ESV (4.2 (1.8) v 3.8 (1.9) v 4.1 (1.6) mm Hg/ml) were not significantly altered (p>0.05). Interestingly, whereas LVIDd was maintained after the game (50 (5) v 50 (6) mm), sBP was transiently but significantly reduced (131 (3) v 122 (3) mm Hg; p<0.05). E:A was moderately (p<0.05) reduced after the game (2.0 (0.4) v 1.5 (0.4)) but returned to baseline within 24 hours. No blood sample contained detectable levels of cTnT. CONCLUSIONS: In this cohort, LV systolic function was not significantly altered after intermittent activity. A transient depression in global diastolic filling was partially attributable to a raised HR and could not be explained by myocyte disruption as represented by cTnT release. PMID- 15273185 TI - Does generalised ligamentous laxity increase seasonal incidence of injuries in male first division club rugby players? AB - OBJECTIVES: To investigate if ligamentous laxity increases seasonal incidence of injury in male first division club rugby players, and to determine if strength protects against injury in hypermobile and tight players. METHODS: Fifty one male first division club rugby players were examined for ligamentous laxity using the Beighton-Horan assessment and graded with an overall laxity score ranging from 0 (tight) to 9 (hyperlax). Each participant was classified into a group determined by their laxity score: tight (0-3), hypermobile (4-6), or excessively hypermobile (7-9). The incidence of joint injuries was recorded prospectively throughout the rugby season and correlated with laxity score. Differences between the groups were analysed. RESULTS: The overall prevalence of generalised joint hypermobility was 24% (12/51). The incidence of injuries was significantly higher in hypermobile (116.7 per 1000 hours) than tight (43.6 per 1000 hours) players (p = 0.034). There were no significant differences in peak strength between the hypermobile and tight groups. CONCLUSIONS: The laxity of the players may explain the differences in injury rates between these groups. Peak strength does not protect the hypermobile joint against injury. It appears that hypermobility may cause an increase in the injury rate of male first division club rugby players. PMID- 15273187 TI - Higher tibial quantitative ultrasound in young female swimmers. AB - BACKGROUND: It has been found that swimming, a non-impact sport, generally has no effect on bone mineral density. OBJECTIVES: To examine bone properties, as measured by quantitative ultrasound, among female swimmers in comparison with control girls and women. METHODS: Subjects included 61 swimmers and 71 controls aged 8.5 to 26.5 years. None of the swimmers was at the elite level and none had included resistance training in her schedule. Bone speed of sound (SOS) was measured bilaterally at the distal radius and the mid-tibia. RESULTS: No differences were observed between swimmers and controls in body mass (mean (SD): 49.7 (12.3) v 50.7 (12.4) kg, respectively), although swimmers were taller (159 (12) v 155 (12) cm) and had lower body fat (18.3 (4.2)% v 22.3 (5.4)%). No difference was found in time since menarche (5.2 (4.0) and 4.5 (2.9) years in swimmers and controls, respectively; 21 swimmers and 25 control were premenarcheal). Radial speed of sound (SOS) increased with age but did not differ between swimmers and controls (non-dominant: 3904 (172) and 3889 (165) m/s for swimmers and controls, respectively). Tibial SOS also increased with age and was significantly higher in swimmers than in controls (non-dominant: 3774 (155) v 3712 (171) m/s). No differences were found between dominant and non-dominant sides. CONCLUSIONS: Swimming appears to be associated with higher bone SOS in the lower but not in the upper extremities. Further studies are needed to assess whether this difference reflects higher habitual activity among the swimmers or swimming specific mechanisms. PMID- 15273188 TI - The Football Association medical research programme: an audit of injuries in academy youth football. AB - OBJECTIVES: To undertake a prospective epidemiological study of the injuries sustained in English youth academy football over two competitive seasons. METHODS: Player injuries were annotated by medical staff at 38 English football club youth academies. A specific injury audit questionnaire was used together with a weekly return form that documented each club's current injury status. RESULTS: A total of 3805 injuries were reported over two complete seasons (June to May) with an average injury rate of 0.40 per player per season. The mean (SD) number of days absent for each injury was 21.9 (33.63), with an average of 2.31 (3.66) games missed per injury. The total amount of time absent through injury equated to about 6% of the player's development time. Players in the higher age groups (17-19 years) were more likely to receive an injury than those in the younger age groups (9-16 years). Injury incidence varied throughout the season, with training injuries peaking in January (p<0.05) and competition injuries peaking in October (p<0.05). Competition injuries accounted for 50.4% of the total, with 36% of these occurring in the last third of each half. Strains (31%) and sprains (20%) were the main injury types, predominantly affecting the lower limb, with a similar proportion of injuries affecting the thigh (19%), ankle (19%), and knee (18%). Growth related conditions, including Sever's disease and Osgood-Schlatter's disease, accounted for 5% of total injuries, peaking in the under 13 age group for Osgood-Schlatter's disease and the under 11 age group for Sever's disease. The rate of re-injury of exactly the same anatomical structure was 3%. CONCLUSIONS: Footballers are at high risk of injury and there is a need to investigate ways of reducing this risk. Injury incidence at academy level is approximately half that of the professional game. Academy players probably have much less exposure to injury than their full time counterparts. Areas that warrant further attention include the link between musculoskeletal development and the onset of youth related conditions such as Sever's disease and Osgood Schlatter's disease, the significant number of non-contact injuries that occur in academy football, and the increased rates of injury during preseason training and after the mid season break. This study has highlighted the nature and severity of injuries that occur at academy level, and the third part of the audit process now needs to be undertaken: the implementation of strategies to reduce the number of injuries encountered at this level. PMID- 15273189 TI - Validation of fan beam dual energy x ray absorptiometry for body composition assessment in adults aged 18-45 years. AB - BACKGROUND: Pencil beam dual energy x ray absorptiometry (DXA) has been shown to provide valid estimates of body fat (%BF), but DXA fan beam technology has not been adequately tested to determine its validity. OBJECTIVE: To compare %BF estimated from fan beam DXA with %BF determined using two and three compartment (2C, 3C) models. METHODS: Men (n = 25) and women (n = 31), aged 18-41 years, participated in the study. Body density, from hydrostatic weighing, was used in the 2C estimate of %BF; DXA was used to determine bone mineral content (BMC) for the 3C estimate of %BF calculated using body density and BMC (3C(BMC)). DXA was also used to determine %BF. Analysis of variance was used to test for significant differences in %BF between sexes and among methods. RESULTS: Women were significantly shorter, weighed less, had less fat free mass, and a higher %BF than men. No significant differences were found among methods (2C, 3C(BMC), DXA) for determination of %BF in either sex. Although not significant, Bland-Altman plots showed that DXA gave higher values for %BF than the 2C and 3C(BMC) methods. CONCLUSION: DXA determination of %BF was not different from that of the 2C and 3C(BMC) models in this group of young adults. However, to validate fan beam DXA fully as a method for body composition assessment in a wide range of individuals and populations, comparisons are needed that use a 4C model with a measure of total body water and BMC. PMID- 15273190 TI - A one season prospective cohort study of volleyball injuries. AB - OBJECTIVE: To estimate the overall incidence of acute and overuse volleyball injuries, and to describe factors associated with ankle sprains. METHODS: 486 players from the second and third Dutch national volleyball divisions participated in the study and were followed prospectively during a whole season. Three measurements were made during the season (baseline, follow up 1, and follow up 2), where all players completed a questionnaire on demographic variables (only at baseline), sports participation, use of preventive measures, and previous injuries. Volleyball exposure during training and matches was recorded for each individual player by the coach on a weekly exposure form. In case of injury the coach provided the injured player with an injury registration form, which had to be completed within one week after the onset of injury. RESULTS: 100 injuries were reported, resulting in an overall injury incidence of 2.6 injuries/1000 hours. The incidence of acute injuries was 2.0/1000 hours. Ankle sprains (n = 41) accounted for most of the acute injuries, and 31 (75%) of all players with an ankle sprain reported a previous ankle sprain. Twenty five overuse injuries were reported. The overall incidence of overuse injuries was 0.6/1000 hours; the back and the shoulder were the most common sites. CONCLUSIONS: Ankle sprain is the most common injury in volleyball, accounting for 41% of all volleyball related injuries. Previous injury seems to be an important risk factor for an ankle sprain. Injury prevention programmes should focus on ankle sprains and concentrate on players with previous ankle sprains. PMID- 15273191 TI - Objective evaluation of small bowel and colonic transit time using pH telemetry in athletes with gastrointestinal symptoms. AB - BACKGROUND: Gastrointestinal (GI) disturbances are often reported by long distance runners and are more common in women, particularly after prolonged high intensity exercise. OBJECTIVES: To determine whether these symptoms could be associated with alterations in GI motility. METHODS: Small bowel and colonic transit were measured using pH telemetry in a group of 11 female athletes (age 22 to 53 years), six of whom experienced lower GI symptoms during exercise. Subjects participated in two experimental sessions: a control measurement, where small bowel transit was estimated during a rest period (R) of six hours; and an exercise session (E), where small bowel transit was measured during a one hour period of high intensity exercise (cross country running) at >70% VO(2)max. Colonic transit was estimated indirectly from determinations of whole gut transit time by radio-opaque marker. RESULTS: Small bowel transit time was 3.5 to 10.6 h (R) and 3.0 to 8.7 h (E) in asymptomatic athletes, versus 4.0 to 6.6 h (R) and 4.6 to 7.3 h (E) in symptomatic athletes (NS). Colonic transit time was 35.0 to 62.5 h (R) and 30.5 to 70.9 h (E) in asymptomatic athletes versus 20.4 to 42.9 h (R) and 21.5 to 67.2 h (E) in symptomatic athletes (NS). CONCLUSIONS: Small bowel and colonic transit times were similar in the two groups in the rest and exercise sessions. The diarrhoea seen in this study did not result from accelerated colonic transit. Other mechanisms must be sought. PMID- 15273192 TI - Serum electrolyte concentrations and hydration status are not associated with exercise associated muscle cramping (EAMC) in distance runners. AB - OBJECTIVES: To determine whether acute exercise associated muscle cramping (EAMC) in distance runners is related to changes in serum electrolyte concentrations and hydration status. METHODS: A cohort of 72 runners participating in an ultra distance road race was followed up for the development of EAMC. All subjects were weighed before and immediately after the race. Blood samples were taken before the race, immediately after the race, and 60 minutes after the race. Blood samples were analysed for glucose, protein, sodium, potassium, calcium, and magnesium concentrations, as well as serum osmolality, haemoglobin, and packed cell volume. Runners who suffered from acute EAMC during the race formed the cramp group (cramp, n = 21), while runners with no history of EAMC during the race formed the control group (control, n = 22). RESULTS: There were no significant differences between the two groups for pre-race or post-race body weight, per cent change in body weight, blood volume, plasma volume, or red cell volume. The immediate post-race serum sodium concentration was significantly lower (p = 0.004) in the cramp group (mean (SD), 139.8 (3.1) mmol/l) than in the control group (142.3 (2.1) mmol/l). The immediate post-race serum magnesium concentration was significantly higher (p = 0.03) in the cramp group (0.73 (0.06) mmol/l) than in the control group (0.67 (0.08) mmol/l). CONCLUSIONS: There are no clinically significant alterations in serum electrolyte concentrations and there is no alteration in hydration status in runners with EAMC participating in an ultra-distance race. PMID- 15273193 TI - A congested football calendar and the wellbeing of players: correlation between match exposure of European footballers before the World Cup 2002 and their injuries and performances during that World Cup. AB - OBJECTIVES: To investigate the correlation between exposure of footballers in European clubs to match play in the months before the World Cup 2002 and their injuries and performances during that World Cup. METHODS: The team doctors at 11 of the best football clubs in Europe prospectively recorded players' exposure and injuries during the 2001-2002 season (July 2001-May 2002). Sixty five players participated in the World Cup in Korea/Japan (June 2002). During the World Cup, the clubs reported injuries sustained by these players, and their performance was evaluated by three international experts. RESULTS: The number of team matches during the season varied between 40 and 76 for the different countries involved. The individual player had a mean of 36 matches during the season. Top players played more matches, especially during the final period of the season. Players who participated in the World Cup played more matches during the season than those who did not (46 v 33 matches). World Cup players did not show any increased risk of injury during the season. About 29% incurred injuries during the World Cup, and 32% performed below their normal standard. The players who underperformed had played more matches during the 10 weeks before the World Cup than those who performed better than expected (12.5 v 9, p<0.05). Twenty three (60%) of the 38 players who had played more than one match a week before the World Cup incurred injuries or underperformed during the World Cup. CONCLUSIONS: There is considerable variation in the number of matches played per season in European professional football leagues. Top level players are obliged to play many matches especially during the final period of the season. PMID- 15273194 TI - The wear and tear of 26.2: dermatological injuries reported on marathon day. AB - Whether it is to take on the challenge, to get in shape and lose weight, to relieve stress, or to enjoy the outdoors, people have increasingly turned to the marathon as their sporting event of choice. Although there are many health benefits, beginners should be aware that injuries are quite common in marathon runners. Among these are the wear and tear injuries to the skin. This is a review of the most commonly reported dermatological injuries on marathon day. PMID- 15273195 TI - Lumbar spine region pathology and hamstring and calf injuries in athletes: is there a connection? AB - This paper discusses the theory that subtle lumbosacral canal impingement of the L5 nerve root may be a relatively common occurrence in older footballers and may in fact be a common underlying basis for the age related predisposition towards hamstring and calf strains. PMID- 15273197 TI - Electronic muscular stimulators: a novel unsuspected cause of rhabdomyolysis. PMID- 15273198 TI - From catastrophe to complexity: a novel model of integrative central neural regulation of effort and fatigue during exercise in humans. AB - It is a popular belief that exercise performance is limited by metabolic changes in the exercising muscles, so called peripheral fatigue. Exercise terminates when there is a catastrophic failure of homoeostasis in the exercising muscles. A revolutionary theory is presented that proposes that exercise performance is regulated by the central nervous system specifically to ensure that catastrophic physiological failure does not occur during normal exercise in humans. PMID- 15273199 TI - Desbaric air embolism during diving: an unusual complication of Osler-Weber-Rendu disease. AB - Cerebral manifestations of Osler-Weber-Rendu disease (OWRD, hereditary haemorrhagic telangiectasia) including telangiectases, venous malformations, and arteriovenous malformations, are usually under-recognised. The highest complication rate is observed in high flow cerebral arteriovenous malformations, which may present with headache, epilepsy, ischaemia, or haemorrhage. Cerebral air embolism during self-contained underwater breathing apparatus (scuba) diving as the first manifestation of pulmonary arteriovenous malformation (PAVM) in OWRD patients has never been reported before. Here we report a 31 year old male who presented desbaric air embolism as the first manifestation of PAVM. As far as we know, this is the first such case published in English medical literature. PMID- 15273200 TI - Video illustration of staple gun use to rapidly repair on-field head laceration. AB - A professional rugby league player sustained a left eyebrow laceration during a match which immediately started to bleed. Within seconds he was taken to the sideline and had the laceration closed with six staples by the team doctor. Bleeding was arrested and he returned to play, taking the ball within 80 s of suffering the initial laceration, and within 40 s of the stapling procedure. This sequence was captured on video and appeared on television. The staples were removed after the match and the wound sutured. Repair of the wound was uneventful. The staple gun allows bleeding lacerations to be closed within seconds and for players to safely and quickly return to play, whilst minimising the risk of blood-borne infection transmission. PMID- 15273201 TI - Subdural haematoma associated with an arachnoid cyst after repetitive minor heading injury in ball games. AB - We report the case of a chronic subdural haematoma caused by repetitive heading of a football which led to the diagnosis of a middle fossa arachnoid cyst. The association between arachnoid cysts and subdural haematoma is discussed as are safety implications in sporting injuries. PMID- 15273202 TI - Upper airway obstruction masquerading as exercise induced bronchospasm in an elite road cyclist. AB - This case concerns an elite road cyclist who complained of occasional dyspnoea and inspiratory difficulty during intense exercise. Clinical examination was normal and the final diagnosis was vocal cord dysfunction, a paradoxical closure of the vocal cords during inspiration which is highly associated with inspiratory stridor at high rates of ventilation. Awareness by the sports physician of this not uncommon condition is important to avoid misdiagnosis. PMID- 15273203 TI - Cardiovascular stress on an elite basketball referee during national competition. AB - This case report examined the cardiovascular stress imposed on an experienced elite basketball referee during national competition. The average heart rate was similar for all matches, approximated 73% of maximum heart rate, and was experienced for most (>63%) of the match. Similar relative exercise intensity was demonstrated regardless of match play (men's v women's) and officiating type (two v three-referee). Further study is needed to document the physiological characteristics of elite basketball referees for greater performance. PMID- 15273204 TI - An unusual presentation of immersion foot. AB - We report a case of "green foot" in a child with a plaster cast applied for a fractured metatarsal who subsequently re-presented with circulatory compromise. The foot was green and smelly and profuse Pseudomonas aeruginosa was cultured. The infection cleared with simple exposure to air. Perhaps this diagnosis should be considered in patients presenting with circulatory compromise in a cast as severe infection can result in amputation. PMID- 15273205 TI - Traumatic humeral articular cartilage shear (THACS) lesion in a professional rugby player: a case report. AB - A 20 year old male professional rugby player was seen at the clinic for evaluation of shoulder pain after rugby play. Magnetic resonance imaging showed extensive subchondral bone bruising of the humeral head with defect of the articular cartilage. Arthroscopy showed that the inferior half of the humeral head had extensive articular cartilage loss with nearly 70% of the inferior head having lost its cartilage. Sports medicine doctors should be aware that the shoulder joint in young competitive athletes playing contact sports may be exposed to greater risk of this kind of injury. PMID- 15273206 TI - Avulsion fracture of peroneus longus at the first metatarsal insertion: a case report. AB - Reports of isolated avulsion fracture at the planter lateral base of the first metatarsal without injury of the tarsometatarsal joint are very rare. A 24 year old man sustained an avulsion fracture at the plantar lateral base of the first metatarsal. Normal alignment of metatarsal bones and tarsometatarsal joint was maintained. In this paper, we describe internal fixation of the displaced fragment using x ray and minimally invasive surgery with good results. PMID- 15273207 TI - Little league shoulder syndrome in an adolescent cricket player. AB - The first case of little league shoulder syndrome in a cricket player is reported. The condition has been reported in baseball pitchers and is characterised by a proximal humeral epiphyseolysis. PMID- 15273208 TI - The reliability of team-based primary data collectors for the collection of exposure and protective equipment use data in community sport. AB - OBJECTIVE: Reliable data allows for the generalisation of study findings to the wider population. The aim of this study was to assess the reliability of using community team based primary data collectors for the collection of exposure and protective equipment use data. METHODS: Nine clubs (23 teams) from a metropolitan Australian Football league in Victoria each provided one primary data collector to monitor exposure and protective equipment use over a regular playing season. Four random audits of this data collection for each team were conducted throughout the regular playing season. The audits were compared with data collected by the club data collectors and the level of agreement assessed. RESULTS: Although exposure data agreement was higher during competition and protective equipment use agreement higher during training, there was no significant difference in data collected by the primary data collectors and the random audits. CONCLUSIONS: The use of trained data collectors associated with Australian Football teams provides reliable information about player exposure and protective equipment use in community intervention studies. PMID- 15273209 TI - The dipsomania of great distance: water intoxication in an Ironman triathlete. AB - Of 371 athletes (62% of all finishers) whose weights were measured before and after the 226 km South African Ironman Triathlon, the athlete who gained the most weight (3.6 kg) during the race was the only competitor to develop symptomatic hyponatraemia. During recovery, he excreted an excess of 4.6 litres of urine. This case report again confirms that symptomatic hyponatraemia is caused by considerable fluid overload independent of appreciable NaCl losses. Hence prevention of the condition requires that athletes be warned not to drink excessively large volumes of fluid (dipsomania) during very prolonged exercise. This case report also shows that there is a delayed diuresis in this condition and that it is not caused by renal failure. PMID- 15273210 TI - Hip fracture-dislocation in football: a report of two cases and review of the literature. AB - Soccer is the world's most popular sport, with over 200 million participants world wide. Fractures account for only 4-9% of acute injuries, and hip fracture dislocation is extremely uncommon. The potentially serious long term sequelae require that team physicians have an awareness of this injury. Two cases of traumatic hip fracture-dislocation are here reported in recreational soccer players sustained by low energy mechanisms. Prompt reduction and fixation are important to produce a stable and congruent joint. PMID- 15273211 TI - Effect of submaximal contraction intensity in contract-relax proprioceptive neuromuscular facilitation stretching. AB - OBJECTIVE: To determine if submaximal contractions used in contract-relax proprioceptive neuromuscular facilitation (CRPNF) stretching of the hamstrings yield comparable gains in hamstring flexibility to maximal voluntary isometric contractions (MVICs). METHOD: Randomised controlled trial. A convenience sample of 72 male subjects aged 18-27 was used. Subjects qualified by demonstrating tight hamstrings, defined as the inability to reach 70 degrees of hip flexion during a straight leg raise. Sixty subjects were randomly assigned to one of three treatment groups: 1, 20% of MVIC; 2, 60% of MVIC; 3, 100% MVIC. Twelve subjects were randomly assigned to a control group (no stretching). Subjects in groups 1-3 performed three separate six second CRPNF stretches at the respective intensity with a 10 second rest between contractions, once a day for five days. Goniometric measurements of hamstring flexibility using a lying passive knee extension test were made before and after the stretching period to determine flexibility changes. RESULTS: Paired t tests showed a significant change in flexibility for all treatment groups. A comparison of least squares means showed that there was no difference in flexibility gains between the treatment groups, but all treatment groups had significantly greater flexibility than the control group. CONCLUSION: CRPNF stretching using submaximal contractions is just as beneficial at improving hamstring flexibility as maximal contractions, and may reduce the risk of injury associated with PNF stretching. PMID- 15273212 TI - Impediments to implementing evidence-based mental health in developing countries. PMID- 15273213 TI - Functional recovery is limited in people with bipolar disorder. PMID- 15273214 TI - DemTect effective in screening for mild cognitive impairment and mild dementia. PMID- 15273215 TI - The Seasonal Health Questionnaire is more effective at detecting seasonal affective disorder than the Seasonal Pattern Adjustment Questionnaire. PMID- 15273216 TI - Review: lithium reduces relapse rates in people with bipolar disorder. PMID- 15273217 TI - Review: psychological therapies can improve psychological symptoms in children who have been sexually abused. PMID- 15273218 TI - Cognitive-behavioural therapy modestly reduces post-traumatic stress symptoms resulting from physical injury. PMID- 15273220 TI - Memantine plus donepezil improves physical and mental health in people with Alzheimer's disease. PMID- 15273219 TI - Cognitive therapy is more effective than fluoxetine in people with generalised social phobia. PMID- 15273221 TI - Donepezil is more effective than galantamine for mild to moderate Alzheimer's disease. PMID- 15273222 TI - Review: interactive, multisession, and targeted programmes most effective in preventing eating disorders. PMID- 15273223 TI - Review: current therapies for men committing acts of domestic violence are of limited benefit. PMID- 15273224 TI - Review: disease management programmes improve detection and care of people with depression. PMID- 15273225 TI - Review: reminiscence and life review are effective therapies for depression in the elderly. PMID- 15273226 TI - Sertraline improves depression scores in the elderly in the short term, regardless of medical comorbidity status. PMID- 15273227 TI - Risperidone improves severe tardive dyskinesia in people with schizophrenia. PMID- 15273228 TI - An early detection programme reduces the duration of untreated first episode psychosis. PMID- 15273229 TI - Multiple family group treatment reduces distress for caregivers of people with schizophrenia. PMID- 15273230 TI - Checklist improves communication between doctors and patients and results in changes in care. PMID- 15273231 TI - History of alcohol abuse reduces response to antipsychotics in people with first episode psychosis. PMID- 15273232 TI - Traumatic brain injury increases the risk of psychiatric illness. PMID- 15273233 TI - ICD-10 predicts risk of relapse and suicide in people diagnosed with a single depressive episode. PMID- 15273234 TI - Depression increases in women during early to late menopause but decreases after menopause. PMID- 15273235 TI - Review: rapid cycling bipolar disorder associated with female gender and bipolar type II subgroup. PMID- 15273236 TI - Review: in older people, decline of cognitive function is more likely than improvement, but rate of change is very variable. PMID- 15273237 TI - Quantitative assessment of hemoglobin-induced endothelial barrier dysfunction. AB - Hemoglobin (Hb)-based O2 carriers (HBOC) are undergoing extensive development as potential "blood substitutes." A major problem associated with these molecules is an increase in microvascular permeability and peripheral vascular resistance. In this paper, we utilized bovine lung microvascular endothelial cell monolayers and simultaneously measured Hb-induced changes in transendothelial electrical resistance, diffusive albumin permeability, and diffusive Hb permeability (PDH) for three forms of Hb: natural tetrameric human Hb-A and two polymerized recombinant HBOCs containing alpha-human and beta-bovine chains designated Hb Polytaur (molecular mass: 500 kDa) and Hb-(Polytaur)n (molecular mass: approximately 1,000,000 Da), respectively. Hb-Polytaur and Hb-(Polytaur)n are being evaluated for clinical use as HBOCs. All three Hb molecules induce a rapid decline of transendothelial electrical resistance to 30% of baseline. Diffusive albumin permeabiltiy increases, on average, approximately ninefold (2.78 x 10(-7) vs. 2.47 x 10(-6) cm/s) in response to Hb exposure. All three Hb molecules induce an increase in their own permeability, a process that we have called Hb-induced Hb permeability. The magnitude of change of PDH is also related to Hb size. When PDH is corrected for the diffusive coefficient for each Hb species, no evidence of restricted diffusion is found. Immunofluorescent images demonstrate Hb-induced actin stress fiber formation and large intercellular gaps. These data provide the first quantitative assessment of the effect of polymerized HBOC on their own diffusion rates over time. We discuss the importance of these findings in terms of Hb extravasation rates, molecular sieving, and clinical consequences of HBOC use. PMID- 15273238 TI - Respiratory response to passive limb movement is suppressed by a cognitive task. AB - Feedback from muscles stimulates ventilation at the onset of passive movement. We hypothesized that central neural activity via a cognitive task source would interact with afferent feedback, and we tested this hypothesis by examining the fast changes in ventilation at the transition from rest to passive leg movement, under two conditions: 1) no task and 2) solving a computer-based puzzle. Resting breathing was greater in condition 2 than in condition 1, evidenced by an increase in mean +/- SE breathing frequency (18.2 +/- 1.1 vs. 15.0 +/- 1.2 breaths/min, P = 0.004) and ventilation (10.93 +/- 1.16 vs. 9.11 +/- 1.17 l/min, P < 0.001). In condition 1, the onset of passive movement produced a fast increase in mean +/- SE breathing frequency (change of 2.9 +/- 0.4 breaths/min, P < 0.001), tidal volume (change of 233 +/- 95 ml, P < 0.001), and ventilation (change of 6.00 +/- 1.76 l/min, P < 0.001). However, in condition 2, the onset of passive movement only produced a fast increase in mean +/- SE breathing frequency (change of 1.3 +/- 0.4 breaths/min, P = 0.045), significantly smaller than in condition 1 (P = 0.007). These findings provide evidence for an interaction between central neural cognitive activity and the afferent feedback mechanism, and we conclude that the performance of a cognitive task suppresses the respiratory response to passive movement. PMID- 15273239 TI - Passive triceps surae stretch inhibits vasoconstriction in the nonexercised limb during posthandgrip muscle ischemia. AB - We investigated whether selective muscle mechanoreceptor activation in the lower limb opposes arm muscle metaboreceptor activation-mediated limb vasoconstriction. Seven subjects completed two trials: one control trial and one stretch trial. Both trials included 2 min of handgrip and 2 min of posthandgrip exercise muscle ischemia (PEMI). In the stretch trial, a 2-min sustained triceps surae stretch, by brief passive dorsiflexion of the right foot, was performed simultaneously during PEMI. Mean arterial pressure, heart rate, and forearm blood flow (FBF) in the nonexercised arm and forearm vascular conductance (FVC) in the nonexercised arm were measured. During PEMI in the control trial, mean arterial pressure was significantly greater and FBF and FVC were significantly lower than baseline values (P < 0.05 for each). In contrast, FBF and FVC during PEMI in the stretch trial exhibited different responses than in the control trial. FBF and FVC were significantly greater in the stretch trial than in the control trial (FBF, 5.5 +/ 0.4 vs. 3.8 +/- 0.4 ml x 100 ml(-1) x min(-1); FVC, 0.048 +/- 0.004 vs. 0.033 +/ 0.003 unit, respectively; P < 0.05). These results indicate that passive triceps surae stretch can inhibit vasoconstriction in the nonexercised forearm mediated via muscle metaboreceptor activation in the exercised arm. PMID- 15273240 TI - Estimation of paracellular conductance of primary rat alveolar epithelial cell monolayers. AB - Freshly isolated rat type II pneumocytes, when grown on permeable tissue culture treated polycarbonate filters, form confluent alveolar epithelial cell monolayers (RAECM). Cells in RAECM undergo transdifferentiation, exhibiting over time morphological and phenotypic characteristics of type I pneumocytes in vivo. We recently reported that transforming growth factor-beta(1) (TGF-beta(1)) decreases overall monolayer resistance (R(te)) and stimulates short-circuit current in a dose-dependent manner. In this study, we investigated the effects of TGF-beta(1) (50 pM) or 10% newborn bovine serum (NBS) on modulation of paracellular passive ion conductance and its contribution to total passive ion conductance across RAECM. On days 5-7 in culture, tight-junctional resistance (R(tj), kOmegacm(2)) of RAECM, cultured in minimally defined serum-free medium (MDSF) with or without TGF-beta(1) or NBS, was estimated from the relationship between observed transmonolayer voltage and resistance after addition of gramicidin D to apical potassium isethionate Ringer solution under open-circuit conditions. NaCl Ringer solution bathed the basolateral side throughout the experimental period. Results showed that transmonolayer conductance (1/R(te)) and tight-junctional conductance (1/R(tj)) are 0.59 and 0.14 mS/cm(2) for control monolayers in MDSF, 1.59 and 0.38 mS/cm(2) for monolayers exposed to TGF-beta(1), and 0.38 and 0.18 mS/cm(2) for monolayers grown in the presence of NBS. The contributions to total transepithelial ion conductance by the paracellular pathway are estimated to be 23, 23, and 47% for control, TGF-beta(1)-exposed, and newborn bovine serum (NBS) treated RAECM, respectively. PMID- 15273241 TI - Chronic fluoxetine microdialysis into the medullary raphe nuclei of the rat, but not systemic administration, increases the ventilatory response to CO2. AB - In conscious rats, focal CO2 stimulation of the medullary raphe increases ventilation, whereas interference with serotonergic function here decreases the ventilatory response to systemic hypercapnia. We sought to determine whether repeated administration of a selective serotonin reuptake inhibitor in this region would increase the ventilatory response to hypercapnia in unanesthetized rats. In rats instrumented with electroencephalogram-electromyogram electrodes, 250 or 500 microM fluoxetine or artificial cerebrospinal fluid (aCSF) was microdialyzed into the medullary raphe for 30 min daily over 15 days. To compare focal and systemic treatment, two additional groups of rats received 10 mg x kg( 1) x day(-1) fluoxetine or vehicle systemically. Ventilation was measured in normocapnia and in 7% CO2 before treatment (day 0), acutely (days 1 or 3), on day 7, and on day 15. There was no change in normocapnic ventilation in any treatment group. Rats that received 250 microM fluoxetine microdialysis showed a significant 13% increase in ventilation in wakefulness during hypercapnia on day 7, due to an increase in tidal volume. In rats microdialyzed with 500 microM fluoxetine, there were 16 and 32% increases in minute ventilation during hypercapnia in wakefulness and sleep on day 7, and 20 and 28% increases on day 15, respectively, again due to increased tidal volume. There was no change in the ventilatory response to CO2 in rats microdialyzed with aCSF or in systemically treated rats. Chronic fluoxetine treatment in the medullary raphe increases the ventilatory response to hypercapnia in an unanesthetized rat model, an effect that may be due to facilitation of chemosensitive serotonergic neurons. PMID- 15273242 TI - Cellular distribution of GPR14 and the positive inotropic role of urotensin II in the myocardium in adult rat. AB - Urotensin II is a cyclic neuropeptide recently shown to play a role via its receptor GPR14 in regulating vascular tone in the mammalian cardiovascular system. The existence of GPR14 in rat heart has been validated by ligand binding assay and RT-PCR. In the present study, we investigated the cellular distribution of GPR14 protein in rat heart by using immunohistochemistry and confocal microscopic immunofluorescence double staining with antipeptide polyclonal antibodies against GPR14 and cell type markers for myocytes and endothelial cells. The direct effect of urotensin II on left ventricular contractility was further evaluated in isolated left ventricular papillary muscles of the rat. In paraffin-embedded heart sections, positive immunohistochemical staining was observed in the left ventricle but not in the right ventricle and atria. Immunofluorescence double staining revealed the cardiac myocyte as the only cell type expressing GPR14 protein in frozen heart sections as well as in isolated cardiac myocytes. There was no visible signal for GPR14 in intramyocardial coronary arteries and capillaries. The existence of GPR14 protein in rat heart was further validated by immunoprecipitation and Western blot analysis. In isolated rat left ventricular papillary muscle preparations, urotensin II induced an increase in active contractile force. GPR14 mRNA was also detected in rat heart by RT-PCR. These data provide the first direct evidence for the cellular localization of GPR14 receptor protein and a positive inotropic effect of urotensin II in normal rat heart. PMID- 15273243 TI - Ventilatory responses to carbon dioxide at low and high levels of oxygen are elevated after episodic hypoxia in men compared with women. AB - We hypothesized that the acute ventilatory response to carbon dioxide in the presence of low and high levels of oxygen would increase to a greater extent in men compared with women after exposure to episodic hypoxia. Eleven healthy men and women of similar race, age, and body mass index completed a series of rebreathing trials before and after exposure to eight 4-min episodes of hypoxia. During the rebreathing trials, subjects initially hyperventilated to reduce the end-tidal partial pressure of carbon dioxide (PetCO2) below 25 Torr. Subjects then rebreathed from a bag containing a normocapnic (42 Torr), low (50 Torr), or high oxygen gas mixture (150 Torr). During the trials, PetCO2 increased while the selected level of oxygen was maintained. The point at which minute ventilation began to rise in a linear fashion as PetCO2 increased was considered to be the carbon dioxide set point. The ventilatory response below and above this point was determined. The results showed that the ventilatory response to carbon dioxide above the set point was increased in men compared with women before exposure to episodic hypoxia, independent of the oxygen level that was maintained during the rebreathing trials (50 Torr: men, 5.19 +/- 0.82 vs. women, 4.70 +/- 0.77 l x min( 1) x Torr(-1); 150 Torr: men, 4.33 +/- 1.15 vs. women, 3.21 +/- 0.58 l x min(-1) x Torr(-1)). Moreover, relative to baseline measures, the ventilatory response to carbon dioxide in the presence of low and high oxygen levels increased to a greater extent in men compared with women after exposure to episodic hypoxia (50 Torr: men, 9.52 +/- 1.40 vs. women, 5.97 +/- 0.71 l x min(-1) x Torr(-1); 150 Torr: men, 5.73 +/- 0.81 vs. women, 3.83 +/- 0.56 l x min(-1) x Torr(-1)). Thus we conclude that enhancement of the acute ventilatory response to carbon dioxide after episodic hypoxia is sex dependent. PMID- 15273244 TI - Effects of midodrine on exercise-induced hypotension and blood pressure recovery in autonomic failure. AB - We tested the hypothesis that the oral alpha1-adrenergic agonist, midodrine, would limit the fall in arterial pressure observed during exercise in patients with pure autonomic failure (PAF). Fourteen subjects with PAF underwent a stand test, incremental supine cycling exercise (25, 50, and 75 W), and ischemic calf exercise, before (control) and 1 h after ingesting 10 mg midodrine. Heart rate (ECG), beat-to-beat blood pressure (MAP, arterial catheter), cardiac output (Q, open-circuit acetylene breathing), forearm blood flow (FBF, Doppler ultrasound), and calf blood flow (CBF, venous occlusion plethysmography) were measured. The fall in MAP after standing for 2 min was similar ( approximately 60 mmHg; P = 0.62). Supine MAP immediately before cycling was greater after midodrine (124 +/- 6 vs 117 +/- 6 mmHg; P < 0.03), but cycling caused a workload-dependent hypotension (P < 0.001), whereas increases in Q were modest but similar. Midodrine increased MAP and total peripheral resistance (TPR) during exercise (P < 0.04), but the exercise-induced fall in MAP and TPR were similar during control and midodrine (P = 0.27 and 0.14). FBF during cycling was not significantly reduced by midodrine (P > 0.2). By contrast, recovery of MAP after cycling was faster (P < 0.04) after midodrine ( approximately 25 mmHg higher after 5 min). Ischemic calf exercise evoked similar peak CBF in both trials, but midodrine reduced the hyperemic response over 5 min of recovery (P < 0.02). We conclude midodrine improves blood pressure and TPR during exercise and dramatically improves the recovery of MAP after exercise. PMID- 15273245 TI - Disruption of a salt bridge dramatically accelerates subunit exchange in duck delta2 crystallin. AB - Intragenic complementation is a unique property of oligomeric enzymes with which to study subunit-subunit interactions. Complementation occurs when different subunits, each possessing distinct mutations that render the individual homomutant proteins inactive, interact to form a heteromutant protein with partial recovery of activity. In this paper, complementation events between human argininosuccinate lyase (ASL) and its homolog, duck delta2 crystallin, were characterized. Different active site mutants in delta2 crystallin complement by the regeneration of native-like active sites as reported previously for ASL. The complementarity of the ASL and delta2 crystallin subunit interfaces was illustrated by the in vivo formation of active hybrid tetramers from inactive ASL and inactive delta2 crystallin mutants. Subunits of both ASL and delta2 crystallin do not dissociate and reassociate in vitro at room temperature, even after 6 days of incubation, indicating that the multimerization interface is very strong. However, disruption of a salt bridge network in the tetrameric interface of delta2 crystallin caused a drastic acceleration of subunit dissociation. Double mutants combining these interface mutants with active site mutants of delta2 crystallin were able to dissociate and reassociate to form active tetramers in vitro within hours. These results suggest that exchange of subunits may occur without unfolding of the monomer. Intragenic complementation in these interface mutants occurs by reintroducing the native salt bridge interaction upon hetero-oligomerization. Our studies demonstrate the value of intragenic complementation as a tool for investigating subunit-subunit interactions in oligomeric proteins. PMID- 15273246 TI - Regulation of histone acetylation during memory formation in the hippocampus. AB - Formation of long term memory begins with the activation of many disparate signaling pathways that ultimately impinge on the cellular mechanisms regulating gene expression. We investigated whether mechanisms regulating chromatin structure were activated during the early stages of long term memory formation in the hippocampus. Specifically, we investigated hippocampal histone acetylation during the initial stages of consolidation of long term association memories in a contextual fear conditioning paradigm. Acetylation of histone H3 in area CA1 of the hippocampus was regulated in contextual fear conditioning, an effect dependent on activation of N-methyl-D-aspartic acid (NMDA) receptors and ERK, and blocked using a behavioral latent inhibition paradigm. Activation of NMDA receptors in area CA1 in vitro increased acetylation of histone H3, and this effect was blocked by inhibition of ERK signaling. Moreover, activation of ERK in area CA1 in vitro through either the protein kinase C or protein kinase A pathways, biochemical events known to be involved in long term memory formation, also increased histone H3 acetylation. Furthermore, we observed that elevating levels of histone acetylation through the use of the histone deacetylase inhibitors trichostatin A or sodium butyrate enhanced induction of long term potentiation at Schaffer-collateral synapses in area CA1 of the hippocampus, a candidate mechanism contributing to long term memory formation in vivo. In concert with our findings in vitro, injection of animals with sodium butyrate prior to contextual fear conditioning enhanced formation of long term memory. These results indicate that histone-associated heterochromatin undergoes changes in structure during the formation of long term memory. Mimicking memory associated changes in heterochromatin enhances a cellular process thought to underlie long term memory formation, hippocampal long term potentiation, and memory formation itself. PMID- 15273247 TI - A mechanism of sulfite neurotoxicity: direct inhibition of glutamate dehydrogenase. AB - Exposure of Neuro-2a and PC12 cells to micromolar concentrations of sulfite caused an increase in reactive oxygen species and a decrease in ATP. Likewise, the biosynthesis of ATP in intact rat brain mitochondria from the oxidation of glutamate was inhibited by micromolar sulfite. Glutamate-driven respiration increased the mitochondrial membrane potential (MMP), and this was abolished by sulfite but the MMP generated by oxidation of malate and succinate was not affected. The increased rate of production of NADH from exogenous NAD+ and glutamate added to rat brain mitochondrial extracts was inhibited by sulfite, and mitochondria preincubated with sulfite failed to reduce NAD+. Glutamate dehydrogenase (GDH) in rat brain mitochondrial extract was inhibited dose dependently by sulfite as was the activity of a purified enzyme. An increase in the Km (glutamate) and a decrease in Vmax resulting in an attenuation in Vmax/Km (glutamate) at 100 microm sulfite suggest a mixed type of inhibition. However, uncompetitive inhibition was noted with decreases in both Km (NAD+) and Vmax, whereas Vmax/Km (NAD+) remained relatively constant. We propose that GDH is one target of action of sulfite, leading to a decrease in alpha-ketoglutarate and a diminished flux through the tricarboxylic acid cycle accompanied by a decrease in NADH through the mitochondrial electron transport chain, a decreased MMP, and a decrease in ATP synthesis. Because glutamate is a major metabolite in the brain, inhibition of GDH by sulfite could contribute to the severe phenotype of sulfite oxidase deficiency in human infants. PMID- 15273248 TI - Regulation of the fusion pore conductance during exocytosis by cyclin-dependent kinase 5. AB - Cyclin-dependent kinase 5 (Cdk5) is a serine/threonine kinase involved in synaptogenesis and brain development, and its enzymatic activity is essential for slow forms of synaptic vesicle endocytosis. Recent work also has implicated Cdk5 in exocytosis and synaptic plasticity. Pharmacological inhibition of Cdk5 modifies secretion in neuroendocrine cells, synaptosomes, and brain slices; however, the specific mechanisms involved remain unclear. Here we demonstrate that dominant-negative inhibition of Cdk5 increases quantal size and broadens the kinetics of individual exocytotic events measured by amperometry in adrenal chromaffin cells. Conversely, Cdk5 overexpression narrows the kinetics of fusion, consistent with an increase in the extent of kiss-and-run exocytosis. Cdk5 inhibition also increases the total charge and current of catecholamine released during the amperometric foot, representing a modification of the conductance of the initial fusion pore connecting the granule and plasma membrane. We suggest that these effects are not attributable to an alteration in catecholamine content of secretory granules and therefore represent an effect on the fusion mechanism itself. Finally, mutational silencing of the Cdk5 phosphorylation site in Munc18, an essential protein of the late stages of vesicle fusion, has identical effects on amperometric spikes as dominant-negative Cdk5 but does not affect the amperometric feet. Cells expressing Munc18 T574A have increased quantal size and broader kinetics of fusion. These results suggest that Cdk5 could, in part, control the kinetics of exocytosis through phosphorylation of Munc18, but Cdk5 also must have Munc18-independent effects that modify fusion pore conductance, which may underlie a role of Cdk5 in synaptic plasticity. PMID- 15273249 TI - Promyelocytic leukemia is a direct inhibitor of SAPK2/p38 mitogen-activated protein kinase. AB - The promyelocytic leukemia gene (PML) encodes a growth/tumor suppressor protein that is essential for the induction of apoptosis in response to various apoptotic signals. The mechanism by which PML plays a role in the regulation of cell death is still unknown. In the current study, we demonstrate that PML negatively regulated the SAPK2/p38 signaling pathway by sequestering p38 from its upstream kinases, MKK3, MKK4, and MKK6, whereas PML did not affect the SAPK1/c-Jun NH(2) terminal kinase pathway. PML associated with p38 both in vitro and in vivo and the carboxyl terminus of PML mediated the interaction. In contrast to other studies of PML and PML-nuclear bodies (NB), our study shows that the formation of PML-NBs was not required for PML to suppress p38 activity because PML was still able to bind and inhibit p38 activity under the conditions in which PML-NBs were disrupted. In addition, we show that the promotion of Fas-induced cell death by PML correlated with the extent of p38 inhibition by PML, suggesting that PML might regulate apoptosis through manipulating SAPK2/p38 pathways. Our findings define a novel function of PML as a negative regulator of p38 kinase and provide further understanding on the mechanism of how PML induces multiple pathways of apoptosis. PMID- 15273250 TI - Identification of a carboxyl-terminal motif essential for the targeting of Na+ HCO-3 cotransporter NBC1 to the basolateral membrane. AB - The Na+-HCO3- cotransporter NBC1 is located exclusively on the basolateral membrane and mediates vectorial transport of bicarbonate in a number of epithelia, including kidney and pancreas. To identify the motifs that direct the targeting of kidney NBC1 to basolateral membrane, wild type and various carboxyl terminally truncated kidney NBC1 mutants were generated, fused translationally in frame to GFP, and transiently expressed in kidney epithelial cells. GFP was linked to the NH2 terminus of NBC1, and labeling was examined by confocal microscopy. Full-length (1035 aa) and mutants with the deletion of 3 or 20 amino acids from the COOH-terminal end of NBC1 (lengths 1032 and 1015 aa, respectively) showed strong and exclusive targeting on the basolateral membrane. However, the deletion of 26 amino acid residues from the COOH-terminal end (length 1010 aa) resulted in retargeting of NBC1 to the apical membrane. Expression studies in oocytes demonstrated that the NBC1 mutant with the deletion of 26 amino acid residues from the COOH-terminal end is functional. Additionally, the deletion of the last 23 amino acids or mutation in the conserved residue Phe at position 1013 on the COOH-terminal end demonstrated retargeting to the apical membrane. We propose that a carboxyl-terminal motif with the sequence QQPFLS, which spans amino acid residues 1010-1015, and specifically the amino acid residue Phe (position 1013) are essential for the exclusive targeting of NBC1 to the basolateral membrane. PMID- 15273251 TI - Site-specific acetylation of the fetal globin activator NF-E4 prevents its ubiquitination and regulates its interaction with the histone deacetylase, HDAC1. AB - Acetylation provides one mechanism by which the functional diversity of individual transcription factors can be expanded. This is valuable in the setting of complex multigene loci that are regulated by a limited number of proteins, such as the human beta-globin locus. We have studied the role of acetylation in the regulation of the transcription factor NF-E4, a component of a protein complex that facilitates the preferential expression of the human gamma-globin genes in fetal erythroid cells. We have shown that NF-E4 interacts directly with, and serves as a substrate for, the acetyltransferase co-activator PCAF. Acetylation of NF-E4 is restricted to a single residue (Lys(43)) in the amino terminal domain of the protein and results in two important functional consequences. Acetylation of NF-E4 prolongs the protein half-life by preventing ubiquitin-mediated degradation. This stabilization is PCAF-dependent, since enforced expression in fetal/erythroid cells of a mutant form of PCAF lacking the histone acetyltransferase domain (PCAFDeltaHAT) decreases NF-E4 stability. Acetylation of Lys(43) also reduces the interaction between NF-E4 and HDAC1, potentially maximizing the activating ability of the factor at the gamma promoter. These results provide further demonstration that co-activators, such as PCAF, can influence individual transcription factor properties at multiple levels to alter their effects on gene expression. PMID- 15273252 TI - Crystal structure of a ternary complex of DnrK, a methyltransferase in daunorubicin biosynthesis, with bound products. AB - One of the final steps in the biosynthesis of the widely used anti-tumor drug daunorubicin in Streptomyces peucetius is the methylation of the 4-hydroxyl group of the tetracyclic ring system. This reaction is catalyzed by the S-adenosyl-L methionine-dependent carminomycin 4-O-methyltransferase DnrK. The crystal structure of the ternary complex of this enzyme with the bound products S adenosyl-L-homocysteine and 4-methoxy-epsilon-rhodomycin T has been determined to a 2.35-angstroms resolution. DnrK is a homodimer, and the subunit displays the typical fold of small molecule O-methyltransferases. The structure provides insights into the recognition of the anthracycline substrate and also suggests conformational changes as part of the catalytic cycle of the enzyme. The position and orientation of the bound ligands are consistent with an SN2 mechanism of methyl transfer. Mutagenesis experiments on a putative catalytic base confirm that DnrK most likely acts as an entropic enzyme in that rate enhancement is mainly due to orientational and proximity effects. This contrasts the mechanism of DnrK with that of other O-methyltransferases where acid/base catalysis has been demonstrated to be an essential contribution to rate enhancement. PMID- 15273253 TI - Human adipocyte fatty acid-binding protein (aP2) gene promoter-driven reporter assay discriminates nonlipogenic peroxisome proliferator-activated receptor gamma ligands. AB - Peroxisome proliferator-activated receptors (PPARs) regulate storage and catabolism of fats and carbohydrates. PPARgamma activity increases insulin sensitivity and adipocyte differentiation at the expense of adipogenesis and weight gain. The goal of this study was to 1) clone the promoter of the human adipocyte fatty acid binding protein (aP2) gene, namely fatty acid-binding protein-4, 2) characterize its pharmacological regulation, and 3) determine its putative predictability for adipogenesis. Among the selected PPAR agonists, rosiglitazone and pioglitazone displayed the highest maximal efficacy (E(max)) on reporter-gene assays in COS-7 cells cotransfected by either a galactosidase 4 response element-based or a human aP2 promoter-based Luc reporter vector, along with either chimeric or full-length human PPAR expression plasmids. The non subtype-selective 2-(4-[2-(3-[2,4-difluorophenyl]-1-heptylureido)ethyl]phenoxy)-2 methyl-butyric acid (GW-2331) and the compounds [4-[3-(4-acetyl-3-hydroxy-2 propylphenoxy)-propoxyl]phenoxy]-acetic acid (L-165041), (4-((2S,5S)-5-(2 (bis(phenylmethyl)amino)-2-oxoethyl)-2-heptyl-4-oxo-3-thiazolidinyl)butyl) benzoic acid (GW-0072), and indomethacin behaved as partial agonists relative to pioglitazone in full-length human aP2-PPARgamma2. Beyond their partial PPARgamma agonist properties, these compounds elicited a lower maximal up-regulation of mouse aP2 mRNA in 3T3-L1 adipocytes as compared with pioglitazone; these properties paralleled a time-dependent increase in neutral lipids. By contrast, the selective PPARalpha agonist 2,2-dichloro-12-(4-chlorophenyl)dodecanoic acid (BM-17.0744) neither stimulated the human aP2-PPARalpha promoter reporter-gene assay, thus demonstrating a specific interaction between PPARgamma and the aP2 promoter, nor affected lipogenesis in 3T3-L1 cells. Altogether, these data characterized a functional promoter of the human aP2 gene; its in vitro pharmacological regulation in PPARgamma-mediated reporter-gene assay may represent an interesting complement or an alternative to time-consuming procedures aiming at discriminating PPAR ligands with low lipogenic properties. PMID- 15273254 TI - Relationship between antiapoptotic molecules and metastatic potency and the involvement of DNA-dependent protein kinase in the chemosensitization of metastatic human cancer cells by epidermal growth factor receptor blockade. AB - The failure to treat metastatic cancer with multidrug resistance is a major problem for successful cancer therapy, and the molecular basis for the association of metastatic phenotype with resistance to therapy is still unclear. In this study, we revealed that various metastatic cancer cells showed consistently higher levels of antiapoptotic proteins, including Bcl-2, nuclear factor-kappaB, MDM2, DNA-dependent protein kinase (DNA-PK), and epidermal growth factor receptor (EGFR), and lower levels of proapoptotic proteins, including Bax and p53 than low metastatic parental cells. This was followed by chemo- and radioresistance in metastatic cancer cells compared with their parental cells. EGFR and DNA-PK activity, which are known to be associated with chemo- and radioresistance, were demonstrated to be mutually regulated by each other. Treatment with PKI166, an EGFR inhibitor, suppressed etoposide-induced activation of DNA-PK in A375SM metastatic melanoma cells. In addition, PKI166 enhanced markedly the chemosensitivities of metastatic cancer cell sublines to various anticancer drugs in comparison with those of low metastatic cancer cells. These results suggest that the activities of DNA-PK and EGFR, which is positively correlated with each other, may contribute to metastatic phenotype as well as therapy resistance, and the EGFR inhibitor enhances the effect of anticancer drugs against therapy-resistant metastatic cancer cells via suppression of stress responses, including activation of DNA-PK. PMID- 15273256 TI - Bush administration health policy. PMID- 15273255 TI - Evaluation of second generation amiloride analogs as therapy for cystic fibrosis lung disease. AB - Epithelial sodium channel (ENaC) blockers have been proposed as a therapy to restore mucus clearance (MC) in cystic fibrosis (CF) airways. The therapeutic effects of the first generation ENaC blocker, amiloride, in CF patients, however, were minimal. Because the failure of amiloride reflected both its low potency and short duration of action on airway surfaces, we investigated whether the increased potency of benzamil and phenamil would produce more favorable pharmacodynamic properties. In vitro potency, maximal efficacy, rate of recovery from maximal block of ENaC, and rate of drug absorption were compared for amiloride, benzamil, and phenamil in cultured human and ovine bronchial epithelial cells. In both human and ovine bronchial epithelia, the rank order of potency was benzamil > phenamil >> amiloride, the maximal efficacy was benzamil = phenamil = amiloride, the recovery to baseline sodium transport was phenamil < benzamil << amiloride, and the rate of drug absorption was phenamil > benzamil >> amiloride. Based on greater potency, benzamil was compared with amiloride in in vivo pharmacodynamic studies in sheep, including tracheal mucus velocity (TMV) and MC. Benzamil enhanced MC and TMV, but acute potency or duration of effect did not exceed that of amiloride. In conclusion, our data support the hypothesis that ENaC blocker aerosol therapy increases MC. However, rapid absorption of benzamil from the mucosal surface offset its greater potency, making it equieffective with amiloride in vivo. More potent, less absorbable, third generation ENaC blockers will be required for an effective aerosol CF pharmacotherapy. PMID- 15273257 TI - Take Care New York: a focused health policy. PMID- 15273258 TI - The obesity epidemic. PMID- 15273259 TI - Measuring progress in meeting healthy people goals for low birth weight and infant mortality among the 100 largest cities and their suburbs. AB - We examined the progress of the nation's 100 largest cities and their surrounding suburban areas toward achieving Healthy People 2000/2010 goals for two measures of infant health: low birth weight (LBW) and infant mortality (IM). Using data from the National Center for Health Statistics, we compared 1990 and 2000 urban and suburban LBW and IM rates to target rates for Healthy People 2000 and 2010 objectives. Although the 2000 LBW weight rate for the 100 largest cities was higher than the average for the suburbs (8.9% vs. 7.1%), the increase in LBW rates for the suburbs was nearly four times that of the cities (15.7% vs. 4.1%). Suburban and urban white infants led the increases in LBW rates; urban and suburban black infants showed a slight decrease or no change in LBW rates. Neither cities nor suburbs, on average, met the 2000 target rate of 5%. It appears unlikely that most of the 100 largest cities and suburbs will meet the Healthy People 2010 goal, which remains at 5%, without reductions in preterm births, nationally on the rise. The IM rate declined across most cities and suburbs between 1990 and 2000. However, the 100 largest cities on average did not meet the 2000 IM rate target of 7 infant deaths per 1000 live births; their suburbs did (8.5 vs. 6.4, respectively). The cities and suburbs that did not meet the 2000 target may be especially challenged to meet the 2010 goal for IM unless rates of preterm births are reduced. With the continuing black-white disparities in LBW and IM rates and the overall differences in the racial composition of the largest cities and suburbs, strategies for meeting Healthy People goals will likely need to be targeted to the specific populations they serve. PMID- 15273260 TI - Unintended pregnancy among the urban poor. AB - This article seeks to determine the proportion of pregnancies that are unintended among poor women in New York City, compare the New York City rate to national data, and examine factors associated with unintended pregnancy in this population. Pregnancy testing data collected between June 1, 1998, and June 1, 2001, from field sites operated by the Office of Family Health, New York City Department of Health and Mental Hygiene were analyzed. Pregnancy planning (intended vs. unintended) was examined by age group, race/ethnicity, marital status, frequency of contraceptive use, number of previous pregnancies, drug and alcohol use, and smoking. Odds ratios were calculated to determine if pregnancies were more likely to be unintended among women with certain characteristics. Logistic regression was used to examine independent risk factors for unintended pregnancy. Of the 20,518 women who had a pregnancy test during the study period, 9,406 (45.8%) were pregnant. Of the pregnancies, 82% were unintended. Marital status was the strongest predictor of unintended pregnancy, increasing the risk 2.5-fold for unmarried women. Adolescents and those who drank alcohol were also at increased risk of unintended pregnancy. The extremely high percentage of pregnancies that were unintended among the study population suggests that national unintended pregnancy rates are not representative of what occurs among low-income women in an urban setting. Unintended pregnancy interventions should be tailored for the urban poor and target unmarried, young women. PMID- 15273261 TI - Weighing social and economic determinants related to inequalities in mortality. AB - It is well known that there are social inequalities in health. Following the ecological approach, unemployment has been one of the most used indicators to study social inequalities. The aim of the present study was to investigate the relationships between indicators of extreme poverty and social unrest, along with unemployment, and mortality in Barcelona, during the years 1989 to 1993. A cross sectional ecological study was carried out using Primary Health Care Areas (PHCAs) as the unit of analysis. The study population consisted of residents in Barcelona City. The indicators studied as dependent variables were the age standardized mortality rates of the following causes of death: total mortality; lung cancer; bronchitis, emphysema, and asthma; cirrhosis; cerebrovascular disease; ischemic heart disease; breast cancer; traffic accidents; acquired immunodeficiency syndrome (AIDS); and drug overdose. Independent variables were male unemployment rate of the primary health care areas and indicators of extreme poverty and social conflict. A descriptive analysis, a bivariate analysis using Spearman correlation coefficients, and a multivariate analysis fitting Poisson regression models were carried out. For the main results, one group of causes of death was associated only with unemployment: bronchitis, emphysema and asthma, cerebrovascular disease, and ischemic heart disease (both men and women); lung cancer (only among men); total mortality and cirrhosis (only among women). Among men, another group of causes of death was associated with extreme poverty and/or social unrest, as well as unemployment: total mortality, cirrhosis, and drug overdose. AIDS in men was only associated with extreme poverty and social unrest. We concluded that we see different types of relationships between deprivation and mortality. Unemployment has been related to mortality because of pathologies with socially accepted risk factors (tobacco and alcohol). Causes of death with risk factors not socially accepted (illegal drug use) have been related to indicators of marginality as well as unemployment. PMID- 15273262 TI - Measuring urbanization pattern and extent for malaria research: a review of remote sensing approaches. AB - Within the next 30 years, the proportion of urban dwellers will rise from under half to two thirds of the world's population. Such a shift will entail massive public health consequences, and most of this urban transition will occur in low income regions of the world. Urban populations face very different health risks compared to those in rural areas, particularly in terms of malaria. To target effective and relevant public health interventions, the need for clear, consistent definitions of what determines urban areas and urban communities is paramount. Decision makers are increasingly seeing remote sensing as a cost effective solution to monitoring urbanization at a range of spatial scales. This review focuses on the progress made within the field of remote sensing on mapping, monitoring, and modeling urban environments and examines existing challenges, drawbacks, and future prospects. We conclude by exploring some of the particular relevance of these issues to malaria and note that they are of more general relevance to all those interested in urban public health. PMID- 15273263 TI - Estimating numbers of injecting drug users in metropolitan areas for structural analyses of community vulnerability and for assessing relative degrees of service provision for injecting drug users. AB - This article estimates the population prevalence of current injection drug users (IDUs) in 96 large US metropolitan areas to facilitate structural analyses of its predictors and sequelae and assesses the extent to which drug abuse treatment and human immunodeficiency virus (HIV) counseling and testing are made available to drug injectors in each metropolitan area. We estimated the total number of current IDUs in the United States and then allocated the large metropolitan area total among large metropolitan areas using four different multiplier methods. Mean values were used as best estimates, and their validity and limitations were assessed. Prevalence of drug injectors per 10,000 population varied from 19 to 173 (median 60; interquartile range 42-87). Proportions of drug injectors in treatment varied from 1.0% to 39.3% (median 8.6%); and the ratio of HIV counseling and testing events to the estimated number of IDUs varied from 0.013 to 0.285 (median 0.082). Despite limitations in the accuracy of these estimates, they can be used for structural analyses of the correlates and predictors of the population density of drug injectors in metropolitan areas and for assessing the extent of service delivery to drug injectors. Although service provision levels varied considerably, few if any metropolitan areas seemed to be providing adequate levels of services. PMID- 15273264 TI - An HIV prevalence-based model for estimating urban risk populations of injection drug users and men who have sex with men. AB - Issues of cost and complexity have limited the study of the population sizes of men who have sex with men (MSM) and injection drug users (IDUs), two groups at clearly increased risk for human immunodeficiency virus (HIV) and other acute and chronic diseases. We developed a prototypical, easily applied estimation model for these populations and applied it to Miami, Florida. This model combined HIV prevalence estimates, HIV seroprevalence rates, and census data to make plausible estimates of the number and proportion of MSM and IDUs under a number of assumptions. Sensitivity analyses were conducted to test the robustness of the model. The model suggests that approximately 9.5% (plausible range 7.7%-11.3%) of Miami males aged 18 years or older are MSM (point estimate, N = 76,500), and 1.4% (plausible range 0.9%-1.9%) of the total population aged 18 years or older are IDUs (point estimate, N = 23,700). Males may be about 2.5 times more likely than females to be IDUs. The estimates were reasonably robust to biases. The model was used to develop MSM and IDU population estimates in selected urban areas across Florida and should be replicable in other medium-to-large urban areas. Such estimates could be useful for behavioral surveillance and resource allocation, including enhanced targeting of community-based interventions for primary and secondary HIV prevention. PMID- 15273265 TI - Prevalence of problematic cocaine consumption in a city of southern Europe, using capture-recapture with a single list. AB - This study aims to determine the magnitude and characteristics of problematic cocaine consumption in the city of Barcelona, Spain. Capture-recapture with a single source was used to estimate prevalence. Log-linear regression models with interaction terms were fitted to the total sample and to subgroups according to other drugs consumed. Emergency room indicator data were obtained from the Barcelona Drug Information System. Drug-related emergencies of Barcelona residents for 1999 were analyzed. During 1999, a total of 4,035 drug-related emergencies were seen in Barcelona hospitals. Of these, 1,656 (41%) involved cocaine consumption; 41% of these patients had consumed cocaine with an opiate; 29% used cocaine with other substances; and 30% used cocaine alone. It was estimated that there was a total of 25,988 problematic cocaine users (95% confidence interval 11,782-58,064), yielding a rate of 31.27 per 1,000 inhabitants aged 15 to 54 years (95% confidence interval 14.2-69.9). The number of cocaine-related emergencies was high enough to allow capture-recapture to be applied, thus obtaining an estimate of the prevalence of problematic cocaine consumption, and high enough to characterize users according to different profiles. The use of capture-recapture with a single source can be interesting for problems related to the urban context. PMID- 15273267 TI - Risk factors for hepatitis C infection and perception of antibody status among male prison inmates in the Hepatitis C Incidence and Transmission in Prisons Study cohort, Australia. AB - The objective of this study was to compare the prevalence of risk factors for hepatitis C virus (HCV) infection among male prison inmates enrolling into a prospective cohort in Australia. We tested 121 inmates who were previously untested or were previously known to be anti-HCV antibody negative for anti-HCV antibodies by enzyme-linked immunosorbent assay. HCV-positive inmates were classified as cases (n = 25) and HCV-negative inmates as controls (n = 96). The study found that cases were less educated than controls and confirmed that prior imprisonment, drug injection, and a longer duration of injecting were risk factors for HCV infection. More than half of those who tested HCV positive perceived that they did not have HCV infection, and 44% were unsure of their HCV status. Those inmates who were incorrect about their HCV status tended to be less educated and were more likely to have been previously imprisoned than those who were correct about their HCV status. Inmates who were unsure of their HCV status were less likely to have been tested for HCV than those who had a clear perception of their HCV status, even if incorrect. Three (12%) inmates who tested positive denied injecting drug use, but reported other risk factors. Prisons are likely to remain an important site for the diagnosis of HCV infection and targeted interventions aimed at risk reduction among inmates with low education levels and a previous imprisonment history. PMID- 15273266 TI - Injection drug use and the hepatitis C virus: considerations for a targeted treatment approach--the case study of Canada. AB - Infection with the hepatitis C virus (HCV) is a major public health burden in Canada and globally. The literature shows that injection drug use is currently the primary transmission route for HCV, and that a majority of injection drug users (IDUs) are currently infected with HCV in Canada. This article first reviews the burden of HCV within IDU populations and the transmission risks and the treatment implications specific to IDUs. Traditionally, IDUs have been excluded from HCV treatment unless abstaining from illicit drug use. However, recent research suggests that categorical exclusion is not medically necessary. A series of key questions about the feasibility of offering HCV treatment to IDUs in the specific Canadian context are considered, including concerns related to the motivation of treatment for IDUs, treatment delivery, treatment side effects, HCV reinfection, and the social environment. The article concludes that treatment of HCV-infected illicit drug users is both feasible and may be necessary to reduce transmission and adverse outcomes in this high-risk population. PMID- 15273268 TI - Cost-effectiveness of universal screening for chlamydia and gonorrhea in US jails. AB - Universal screening for the sexually transmitted diseases (STDs) of chlamydia and gonorrhea on intake in jails has been proposed as the most effective strategy to decrease morbidity in inmates and to reduce transmission risk in communities after release. Most inmates come from a population that is at elevated risk for STDs and has limited access to health care. However, limited resources and competing priorities force decision makers to consider the cost of screening programs in comparison to other needs. The costs and cost-effectiveness of universal screening in correctional settings have not been documented. We estimated the incremental cost-effectiveness of universal urine-based screening for chlamydia and gonorrhea among inmates on intake in US jails compared to the commonly used practice of presumptive treatment of symptomatic inmates without laboratory testing. Decision analysis models were developed to estimate the cost effectiveness of screening alternatives and were applied to hypothetical cohorts of male and female inmates. For women, universal screening for chlamydia only was cost-saving to the health care system, averting more health care costs than were incurred in screening and treatment. However, for men universal chlamydia screening cost $4,856 more per case treated than presumptive treatment. Universal screening for both chlamydia and gonorrhea infection cost the health care system $3,690 more per case of pelvic inflammatory disease averted for women and $650 more per case of infection treated for men compared to universal screening for chlamydia only. Jails with a high prevalence of chlamydia and gonorrhea represent an operationally feasible and cost-effective setting to universally test and treat women at high risk for STDs and with limited access to care elsewhere. PMID- 15273269 TI - Identification of gaps in the diagnosis and treatment of childhood asthma using a community-based participatory research approach. AB - The goal of this investigation was to use a community-based participatory research approach to develop, pilot test, and administer an asthma screening questionnaire to identify children with asthma and asthma symptoms in a community setting. This study was conducted as the recruitment effort for Community Action Against Asthma, a randomized trial of a household intervention to reduce exposure to environmental triggers of asthma and was not designed as a classic prevalence study. An asthma screening questionnaire was mailed and/or hand delivered to parents of 9,627 children, aged 5 to 11 years, in two geographic areas of Detroit, Michigan, with predominantly African American and Hispanic populations. Additional questionnaires were distributed via community networking. Measurements included parent report of their child's frequency of respiratory symptoms, presence of physician diagnosis of asthma, and frequency of doctor-prescribed asthma medication usage. Among the 3,067 completed questionnaires, 1,570 (51.2% of returned surveys, 16.3% of eligible population) were consistent with asthma of any severity and 398 (12.9% of returned surveys, 4.1% of eligible population) met criteria for moderate-to-severe asthma. Among those meeting criteria for moderate to-severe asthma, over 30% had not been diagnosed by a physician, over one half were not taking daily asthma medication, and one quarter had not taken any physician-prescribed asthma medication in the past year. Screening surveys conducted within the context of a community-based participatory research partnership can identify large numbers of children with undiagnosed and/or undertreated moderate-to-severe asthma. These children are likely to benefit from interventions to reduce morbidity and improve quality of life. PMID- 15273270 TI - Workplace health and safety concerns in service organizations in the inner city. AB - There is little known about occupational health and safety concerns or programs in workplaces in the inner city. This work was part of a needs assessment for development of occupational health and safety programs for workplaces in the inner city. Its key objective was to identify inner-city worker concerns regarding specific hazards. The work involved two phases. The first sampled workers in an inner-city hospital and church, and the second involved both paid and volunteer workers in inner-city community outreach programs. The key concerns raised by inner-city workers were infectious disease and personal safety and violence. Occupational health and safety programs need to address infectious disease and personal safety issues in this environment. Further research is needed regarding workplace health and safety in inner-city workplaces, both regarding hazards particular to the inner city and occupational health programs for the workers, both paid and volunteer, who work there. PMID- 15273271 TI - Availability of antibiotics without prescription in New York City. AB - Misuse of antibiotics in the community has been associated with emergence of increasingly antibiotic-resistant bacterial strains. Although antibiotics in the United States are to be prescribed by a health care provider, the extent to which they are obtained by other means is not known. The purpose of this article is to describe a survey of the availability of nonprescription antibiotics in neighborhood independent businesses in several Manhattan, New York, neighborhoods. A survey was conducted of 101 stores in three neighborhoods--one primarily Hispanic; one primarily black, non-Hispanic; and one primarily white, non-Hispanic. Antibiotics were available in all stores in the Hispanic neighborhood (n = 34), but in none of the others (P < .001). If efforts to rationalize the use of antibiotics are to be successful, the beliefs and cultural norms of subpopulations must be considered, and interventions must be culturally relevant. PMID- 15273272 TI - Attempted suicide among injecting and noninjecting cocaine users in Sydney, Australia. AB - A sample of 183 current cocaine users, 120 primary injecting cocaine users (ICUs), and 63 primary noninjecting cocaine users (NICUs) were administered a structured interview to ascertain attempted suicide histories, methods used, and factors associated with suicide attempts. All respondents were volunteers and current cocaine users recruited through a wide range of sources. The mean age of participants was 30.1 years, and 65% were male. The ICUs were older (32.3 vs. 26.7 years, respectively), more likely to be male (72% vs. 54%, respectively), to be unemployed (84% vs. 23%, respectively) and to have a prison history (53% vs. 1%, respectively) compared to NICUs. Of the sample, 31% had attempted suicide, 18% had done so on more than one occasion, and 8% had made an attempt in the preceding 12 months. Overall, 28% of the sample had been treated by a medical practitioner after an attempt. ICUs (38%) were significantly more likely than NICUs (10%) to have attempted suicide and to have done so on more than one occasion (23% vs. 3%, respectively). The most common method used among both groups was self-poisoning (ICUs 28%, NICUs 8%), primarily by drug overdose. Violent methods had been used by 22% of ICUs and 3% of NICUs. Multivariate analyses revealed that injecting, female gender, and more extensive polydrug use were independent predictors of a suicide attempt. The prevalence of suicide in this study indicates that it represents a major clinical issue among ICUs and to a lesser extent among noninjectors of the drug. Those treating cocaine users for drug dependence need to be aware of the salience of suicide as a problem, among injectors in particular. PMID- 15273273 TI - The Junior Fellows Program: motivating urban youth toward careers in health, science, and medicine. AB - Lack of diversity in the health, science, and medical professions has been documented as a contributor to health disparities in the United States, and early intervention is essential for the recruitment of underrepresented students into the health professions. The Junior Fellows Program, a partnership between the New York Academy of Medicine, New York City public schools, and regional academic medical centers, is designed to stimulate students' interest in health, science, medicine, and research. From seminars designed to advance Junior Fellows' skills in identifying concrete strategies for improving health and preventing illness, to understanding the research process and the nature of scientific inquiry, the program engages Junior Fellows in project-based learning, works to enhance their critical thinking skills, and helps them to foster positive interactions with practicing physicians and health professionals. Surveys of program graduates indicate the program has been influential in creating a high level of motivation to pursue careers related to health, science, and medicine. The program continues to work on enhancing educational opportunities for urban public school students and promoting career awareness for the health professions, with a special emphasis on improving the proportion of minorities and women who enter these fields. PMID- 15273274 TI - Caught in the act: visualization of an intermediate in the DNA base-flipping pathway induced by HhaI methyltransferase. AB - Rotation of a DNA or RNA nucleotide out of the double helix and into a protein pocket ('base flipping') is a mechanistic feature common to some DNA/RNA-binding proteins. Here, we report the structure of HhaI methyltransferase in complex with DNA containing a south-constrained abasic carbocyclic sugar at the target site in the presence of the methyl donor byproduct AdoHcy. Unexpectedly, the locked south pseudosugar appears to be trapped in the middle of the flipping pathway via the DNA major groove, held in place primarily through Van der Waals contacts with a set of invariant amino acids. Molecular dynamics simulations indicate that the structural stabilization observed with the south-constrained pseudosugar will not occur with a north-constrained pseudosugar, which explains its lowered binding affinity. Moreover, comparison of structural transitions of the sugar and phosphodiester backbone observed during computational studies of base flipping in the M.HhaI-DNA-AdoHcy ternary complex indicate that the south-constrained pseudosugar induces a conformation on the phosphodiester backbone that corresponds to that of a discrete intermediate of the base-flipping pathway. As previous crystal structures of M.HhaI ternary complex with DNA displayed the flipped sugar moiety in the antipodal north conformation, we suggest that conversion of the sugar pucker from south to north beyond the middle of the pathway is an essential part of the mechanism through which flipping must proceed to reach its final destination. We also discuss the possibility of the south constrained pseudosugar mimicking a transition state in the phosphodiester and sugar moieties that occurs during DNA base flipping in the presence of M.HhaI. PMID- 15273275 TI - Sequence-specific artificial ribonucleases. I. Bis-imidazole-containing oligonucleotide conjugates prepared using precursor-based strategy. AB - Antisense oligonucleotide conjugates, bearing constructs with two imidazole residues, were synthesized using a precursor-based technique employing post synthetic histamine functionalization of oligonucleotides bearing methoxyoxalamido precursors at the 5'-termini. The conjugates were assessed in terms of their cleavage activities using both biochemical assays and conformational analysis by molecular modelling. The oligonucleotide part of the conjugates was complementary to the T-arm of yeast tRNA(Phe) (44-60 nt) and was expected to deliver imidazole groups near the fragile sequence C61-ACA-G65 of the tRNA. The conjugates showed ribonuclease activity at neutral pH and physiological temperature resulting in complete cleavage of the target RNA, mainly at the C63 A64 phosphodiester bond. For some constructs, cleavage was completed within 1-2 h under optimal conditions. Molecular modelling was used to determine the preferred orientation(s) of the cleaving group(s) in the complexes of the conjugates with RNA target. Cleaving constructs bearing two imidazole residues were found to be conformationally highly flexible, adopting no preferred specific conformation. No interactions other than complementary base pairing between the conjugates and the target were found to be the factors stabilizing the 'active' cleaving conformation(s). PMID- 15273276 TI - Changing the recognition specificity of a DNA-methyltransferase by in vitro evolution. AB - The gene coding for the SinI DNA-methyltransferase, a modification enzyme able to recognize and methylate the internal cytosine of the GG(A)/(T)CC sequence, was subjected to in vitro mutagenesis, DNA-shuffling and a strong selection for relaxed GGNCC recognition specificity. As a result of this in vitro evolution experiment, a mutant gene with the required phenotype was selected. The mutant SinI methyltransferase carried five amino acid substitutions. None of these was found in the 'variable region' that were thought to be responsible for sequence specificity. Three were located near the N-terminal end, preceding the first conserved structural motif of the enzyme; two were found between conserved motifs VI and VII. A clone engineered to carry out only the latter two replacements (L214S and Y229H) displays relaxed recognition specificity similar to that of the parental mutant, whereas the clone carrying only the N-terminal replacements showed a much weaker change in recognition specificity. The enzyme with two internal mutations was purified and characterized. Its catalytic activity (kcat/Km) was approximately 5-fold lower towards GG(A)/(T)CC and 20-fold higher towards GG(G)/(C)CC than that of the wild-type enzyme. PMID- 15273277 TI - Identification of important chemical groups of the hut mRNA for HutP interactions that regulate the hut operon in Bacillus subtilis. AB - HutP is an RNA binding protein that regulates the expression of the histidine utilization (hut) operon in Bacillus species by binding to cis-acting regulatory sequences on hut mRNA. We recently solved the HutP crystal structure, which revealed a novel fold where three dimers are arranged in a 3-fold axis to form the hexamer. We also identified a minimal RNA binding element sufficient for HutP binding: three UAG trinucleotide motifs, each separated by 4 nt, located just upstream of the terminator. In the present study we have identified important RNA chemical groups essential for HutP interactions, by combining an in vitro selection strategy and analyses by site-specific base substitutions. These analyses suggest that each HutP molecule recognizes one UAG motif, where the first base (U) can be substituted with other bases, while the second and third bases (A and G) are required for the interactions. Further analyses of the chemical groups of the A and G bases in the UAG motif by modified base analogs suggested the importance of the exocyclic NH2 group in these bases. Also, in this motif, only the 2'-OH group of A is important for HutP recognition. Considering the important chemical groups identified here, as well as the electrostatic potential analysis of HutP, we propose that Glu137 is one of the important residues for the HutP-RNA interactions. PMID- 15273278 TI - The adenovirus priming protein pTP contributes to the kinetics of initiation of DNA replication. AB - Adenovirus (Ad) precursor terminal protein (pTP) in a complex with Ad DNA polymerase (pol) serves as a primer for Ad DNA replication. During initiation, pol covalently couples the first dCTP with Ser-580 of pTP. By using an in vitro reconstituted replication system comprised of purified proteins, we demonstrate that the conserved Asp-578 and Asp-582 residues of pTP, located close to Ser-580, are important for the initiation activity of the pTP/pol complex. In particular, the negative charge of Asp-578 is essential for this process. The introduced pTP mutations do not alter the binding capacity to DNA or polymerase, suggesting that the priming mechanism is affected. The Asp-578 or Asp-582 mutations increase the Km for dCTP incorporation, and higher dCTP concentrations or Mn2+ replacing Mg2+ partially relieve the initiation defect. Moreover, the kcat/Km values are reduced as a consequence of the pTP mutations. These observations demonstrate that pTP influences the catalytic activity of pol in initiation. Since both Asp residues are situated close to the pol active site during initiation, they may contribute to correct positioning of the OH group in Ser-580. Our results indicate that specific amino acids of the protein primer influence the ability of Ad5 DNA polymerase to initiate DNA replication. PMID- 15273279 TI - Detection of protein-DNA interaction with a DNA probe: distinction between single strand and double-strand DNA-protein interaction. AB - A simple, direct method for the detection of DNA-protein interaction was developed with electrochemical methods. Single-stranded DNA (ss-DNA) probes were prepared through the chemical bonding of an oligonucleotide to a polymer film bearing carboxylic acid groups, and double-stranded DNA (ds-DNA) probes were prepared through hybridization of the complementary sequence DNA on the ss-DNA probe. Impedance spectroscopy and differential pulse voltammetry (DPV) distinguished the interaction between the DNA probes with mouse Purbeta (mPurbeta), an ss-DNA binding protein, and with Escherichia coli MutH, a ds-DNA binding protein. Impedance spectra obtained before and after the interaction of DNA probes with these proteins clearly showed the sequence-specific ss-DNA preference of mPurbeta and the sequence-specific ds-DNA preference of MutH. The concentration dependence of proteins on the response of the DNA probes was also investigated, and the detection limits of MutH and mPurbeta were 25 and 3 microg/ml, respectively. To confirm the impedance results, the variation of the current oxidation peak of adenine of the DNA probe was monitored with DPV. The formation constants of the complexes formed between the probe DNA and the proteins were estimated based on the DPV results. PMID- 15273280 TI - Endometrial TIMP-4 mRNA is high at midcycle and in hyperplasia, but down regulated in malignant tumours. Coordinated expression with MMP-26. AB - We have previously reported that endometrial expression of matrix metalloproteinase (MMP)-26 mRNA comes to a maximum in the early secretory phase. Since tissue inhibitor of metalloproteinase (TIMP)-4 is a potent inhibitor of MMP 26, the objective of this study was to identify the pattern of TIMP-4 mRNA expression in the normal endometrial cycle. We also evaluated hyperplastic, pre malignant (atypical hyperplasia) and malignant endometrial tissue. Endometrial TIMP-4 mRNA was localized in tissue sections using in situ hybridization, and quantified in tissue extracts using real-time PCR. Estrogen receptor alpha (ERalpha) was assayed in the same set of samples using immunohistochemistry. In situ hybridization demonstrated TIMP-4 mRNA in the stroma of both normal and pathological tissues. TIMP-4 mRNA increased in the proliferative phase to a maximum in the early secretory phase, and then decreased in the late part of the cycle. Expression was comparable in normal and hyperplastic (including atypical) endometrial samples, whereas lower levels were detected in malignant tumours. Since this general pattern of expression suggests estrogen dependence, we evaluated ERalpha in our samples. Tissue sections from the normal proliferative phase, hyperplasia and pre-malignant atypical hyperplasia tissue stained strongly for ERalpha, whereas weak staining was seen in the secretory phase and in malignant tumours. Thus, low level of ERalpha was accompanied by down-regulated TIMP-4 mRNA, supporting the hypothesis that ERalpha contributes to regulation of the TIMP-4 gene. In addition, we identified a putative estrogen response element (ERE) in the promoter region of the TIMP-4 gene at position -930 to -916. Similarities in the cyclic patterns of TIMP-4 mRNA and MMP-26 mRNA, together with the fact that TIMP-4 is a potent inhibitor of MMP-26, suggest a functional relationship, and furthermore a role in human implantation. PMID- 15273281 TI - Analysis of the codon 72 polymorphism of the TP53 gene in patients with endometriosis. AB - Endometriosis is a benign gynaecologic disease that is associated with a certain risk for malignant degeneration. The disease has a genetic background, but the locations of possible genomic aberrations are still poorly clarified. In this context, the proline form of TP53 codon 72 polymorphism has been recently associated with the risk of developing endometriosis. In this case-control study, we aimed to investigate further the potential association between endometriosis and this polymorphism in order to evaluate whether this genetic variant may influence the susceptibility to the disease. Genomic DNA was obtained from a consecutive series of 303 Italian Caucasian women of reproductive age who underwent laparoscopy for benign gynaecological pathologies. Endometriosis was defined according to the criteria of Holt and Weiss [Holt V and Weiss NS (2000) Recommendations for the design of epidemiologic studies of endometriosis. Epidemiol 11,654-659] for the definite disease. Subjects of similar age without laparoscopic evidence of the disease served as control group. Molecular analysis of TP53 codon 72 polymorphism was performed by PCR amplification. Endometriosis was documented in 151 women. We found no statistically significant difference in the distribution of TP53 codon 72 polymorphism genotypes between patients with and without endometriosis. The respective proportions of arginine homozygotes, heterozygotes and proline homozygotes were 55.6, 39.7 and 4.6% in the group with endometriosis and 59.9, 30.9 and 9.2% in the control group. Moreover, no statistically significant association was demonstrated between TP53 codon 72 polymorphism and the various clinical manifestations of the disease, although a non-significant tendency towards an increased frequency of the proline allele was observed in association with specific manifestations of the disease reflecting a more severe form. Our results suggest that the TP53 codon 72 polymorphism does not confer genetic susceptibility to endometriosis in the Italian population. However, a possible susceptibility role of this polymorphism in endometriosis development towards very severe forms cannot be ruled out. PMID- 15273282 TI - Sphingosine 1-phosphate activates Weibel-Palade body exocytosis. AB - Sphingosine 1-phosphate (S1P) not only regulates angiogenesis, vascular permeability and vascular tone, but it also promotes vascular inflammation. However, the molecular basis for the proinflammatory effects of S1P is not understood. We now show that S1P activates endothelial cell exocytosis of Weibel Palade bodies, releasing vasoactive substances capable of causing vascular thrombosis and inflammation. S1P triggers endothelial exocytosis in part through phospholipase C-gamma signal transduction. However, S1P also modulates endothelial cell exocytosis by activating endothelial nitric oxide synthase production of nitric oxide, which inhibits exocytosis. Thus S1P plays a dual role in regulating endothelial exocytosis, triggering pathways that both promote and inhibit endothelial exocytosis. Regulation of endothelial exocytosis may explain part of the proinflammatory effects of S1P. PMID- 15273283 TI - Mapping of sudden infant death with dysgenesis of the testes syndrome (SIDDT) by a SNP genome scan and identification of TSPYL loss of function. AB - We have identified a lethal phenotype characterized by sudden infant death (from cardiac and respiratory arrest) with dysgenesis of the testes in males [Online Mendelian Inheritance in Man (OMIM) accession no. 608800]. Twenty-one affected individuals with this autosomal recessive syndrome were ascertained in nine separate sibships among the Old Order Amish. High-density single-nucleotide polymorphism (SNP) genotyping arrays containing 11,555 single-nucleotide polymorphisms evenly distributed across the human genome were used to map the disease locus. A genome-wide autozygosity scan localized the disease gene to a 3.6-Mb interval on chromosome 6q22.1-q22.31. This interval contained 27 genes, including two testis-specific Y-like genes (TSPYL and TSPYL4) of unknown function. Sequence analysis of the TSPYL gene in affected individuals identified a homozygous frameshift mutation (457_458insG) at codon 153, resulting in truncation of translation at codon 169. Truncation leads to loss of a peptide domain with strong homology to the nucleosome assembly protein family. GFP-fusion expression constructs were constructed and illustrated loss of nuclear localization of truncated TSPYL, suggesting loss of a nuclear localization patch in addition to loss of the nucleosome assembly domain. These results shed light on the pathogenesis of a disorder of sexual differentiation and brainstem mediated sudden death, as well as give insight into a mechanism of transcriptional regulation. PMID- 15273284 TI - The total synthesis of (-)-crispatene. AB - The total synthesis of the molluscan polypropionate (-)-crispatene is described. The synthesis features a palladium-catalyzed cross-coupling to establish a sensitive conjugated tetraene and its Lewis acid-catalyzed cycloisomerization to yield the bicyclo[3.1.0]hexene core of the natural product. The absolute configuration of (-)-crispatene and related molecules is established. PMID- 15273285 TI - Molecular-timetable methods for detection of body time and rhythm disorders from single-time-point genome-wide expression profiles. AB - Detection of individual body time (BT) via a single-time-point assay has been a longstanding unfulfilled dream in medicine, because BT information can be exploited to maximize potency and minimize toxicity during drug administration and thus will enable highly optimized medication. To achieve this dream, we created a "molecular timetable" composed of >100 "time-indicating genes," whose gene expression levels can represent internal BT. Here we describe a robust method called the "molecular-timetable method" for BT detection from a single time-point expression profile. The power of this method is demonstrated by the sensitive and accurate detection of BT and the sensitive diagnosis of rhythm disorders. These results demonstrate the feasibility of BT detection based on single-time-point sampling, suggest the potential for expression-based diagnosis of rhythm disorders, and may translate functional genomics into chronotherapy and personalized medicine. PMID- 15273286 TI - Red cell membrane and plasma linoleic acid nitration products: synthesis, clinical identification, and quantitation. AB - Nitric oxide (*NO) and its reactive metabolites mediate the oxidation, nitration, and nitrosation of DNA bases, amino acids, and lipids. Here, we report the structural characterization and quantitation of two allylic nitro derivatives of linoleic acid (LNO(2)), present as both free and esterified species in human red cell membranes and plasma lipids. The LNO(2) isomers 10-nitro-9-cis, 12-cis octadecadienoic acid and 12-nitro-9-cis, 12-cis-octadecadienoic acid were synthesized and compared with red cell and plasma LNO(2) species based on chromatographic elution and mass spectral properties. Collision-induced dissociation fragmentation patterns from synthetic LNO(2) isomers were identical to those of the two most prevalent LNO(2) positional isomers found in red cells and plasma. By using [(13)C]LNO(2) as an internal standard, red cell free and esterified LNO(2) content was 50 +/- 17 and 249 +/- 104 nM, respectively. The free and esterified LNO(2) content of plasma was 79 +/- 35 and 550 +/- 275 nM, respectively. Nitrated fatty acids, thus, represent the single largest pool of bioactive oxides of nitrogen in the vasculature, with a net LNO(2) concentration of 477 +/- 128 nM, excluding buffy coat cells. These observations affirm that basal oxidative and nitrating conditions occur in healthy humans to an extent that is sufficient to induce abundant membrane and lipoprotein-fatty acid nitration. Given that LNO(2) is capable of mediating cGMP and non-cGMP-dependent signaling reactions, fatty acid nitration products are species representing the convergence of ()NO and oxygenated lipid cell-signaling pathways. PMID- 15273287 TI - Identification of candidate cancer-causing genes in mouse brain tumors by retroviral tagging. AB - Murine retroviruses may cause malignant tumors in mice by insertional mutagenesis of host genes. The use of retroviral tagging as a means of identifying cancer causing genes has, however, almost entirely been restricted to hematopoietic tumors. The aim of this study was to develop a system allowing for the retroviral tagging of candidate genes in malignant brain tumors. Mouse gliomas were induced by a recombinant Moloney murine leukemia virus encoding platelet-derived growth factor (PDGF) B-chain. The underlying idea was that tumors evolve through a combination of PDGF-mediated autocrine growth stimulation and insertional mutagenesis of genes that cooperate with PDGF in gliomagenesis. Common insertion sites (loci that were tagged in more than one tumor) were identified by cloning and sequencing retroviral flanking segments, followed by blast searches of mouse genome databases. A number of candidate brain tumor loci (Btls) were identified. Several of these Btls correspond to known tumor-causing genes; these findings strongly support the underlying idea of our experimental approach. Other Btls harbor genes with a hitherto unproven role in transformation or oncogenesis. Our findings indicate that retroviral tagging with a growth factor-encoding virus may be a powerful means of identifying candidate tumor-causing genes in nonhematopoietic tumors. PMID- 15273288 TI - Semaphorin 3B (SEMA3B) induces apoptosis in lung and breast cancer, whereas VEGF165 antagonizes this effect. AB - Semaphorin 3B (SEMA3B) is a secreted member of the semaphorin family, important in axonal guidance. We and others have shown that SEMA3B can act as a tumor suppressor by inducing apoptosis either by reexpression in tumor cells or applied as a soluble ligand. The common method of inactivation of SEMA3B is by allele loss and tumor-acquired promoter methylation. We studied the mechanism of SEMA3B induced tumor cell apoptosis and found that vascular endothelial growth factor (VEGF)165 significantly decreased the proapoptotic and antimitotic effect of transfected or secreted SEMA3B on lung and breast cancer cells. VEGF165 binds to neuropilin, receptors for SEMA3B, and we found that SEMA3B competed for binding of 125I-VEGF165 to lung and breast cancer cells. We also found that small interfering RNA knockdown of tumor-produced VEGF-A or the use of an anti-VEGF neutralizing antibody (Ab) significantly inhibited tumor cell growth in vitro. By contrast, VEGF121, a VEGF variant that lacks binding to neuropilin (NP)-1 or NP-2 receptors, was not expressed in tumor cells and had no effect on SEMA3B growth suppressing activities. In conclusion, we hypothesize that VEGF165, produced by tumor cells, acts as an autocrine survival factor and that SEMA3B mediates its tumor-suppressing effects, at least in part, by blocking this VEGF autocrine activity. PMID- 15273289 TI - Mutations in herpes simplex virus glycoprotein D that prevent cell entry via nectins and alter cell tropism. AB - Glycoprotein D (gD) determines which cells can be infected by herpes simplex virus (HSV) by binding to one of the several cell surface receptors that can mediate HSV entry or cell fusion. These receptors include the herpesvirus entry mediator (HVEM), nectin-1, nectin-2, and sites in heparan sulfate generated by specific 3-O-sulfotransferases. The objective of the present study was to identify residues in gD that are critical for physical and functional interactions with nectin-1 and nectin-2. We found that double or triple amino acid substitutions at positions 215, 222, and 223 in gD caused marked reduction in gD binding to nectin-1 and a corresponding inability to function in cell fusion or entry of HSV via nectin-1 or nectin-2. These substitutions either enhanced or did not significantly inhibit functional interactions with HVEM and modified heparan sulfate. These and other results demonstrate that different domains of gD, with some overlap, are critical for functional interactions with each class of entry receptor. Viral entry assays, using gD mutants described here and previously, revealed that nectins are the principal entry receptors for selected human cell lines of neuronal and epithelial origin, whereas HVEM or nectins could be used to mediate entry into a T lymphocyte line. Because T cells and fibroblasts can be infected via HVEM, HSV strains carrying gD mutations that prevent entry via nectins may establish transient infections in humans, but perhaps not latent infections of neurons, and are therefore candidates for development of safe live virus vaccines and vaccine vectors. PMID- 15273290 TI - Heterodimerization of type II phytochromes in Arabidopsis. AB - Coimmunoprecipitation of members of the phytochrome red/farred photoreceptor family from plant extracts has been used to analyze their heteromeric binding interactions. Phytochrome (phy)B or phyD apoproteins with six myc epitopes fused to their N termini are biologically active when expressed in Arabidopsis. Immunoprecipitation of either of these tagged proteins from seedling extracts coprecipitates additional type II phytochromes: six myc (myc6)-phyB coprecipitates phyC-phyE; and myc6-phyD coprecipitates phyB and phyE. No interaction of the epitope-tagged proteins with type I phyA was detected. Gel filtration chromatography shows that all five of the Arabidopsis phytochromes are present in seedlings as dimers, and that the heteromeric type II phytochrome complexes migrate at molecular masses characteristic of heterodimers. Similar levels of heterodimer formation are observed in extracts of dark-grown seedlings, where the phytochromes are cytosolic, and light-grown seedlings, where they are predominantly nuclear. These findings indicate that Arabidopsis, which until now has been thought to contain five homodimeric forms of phytochrome, in fact contains multiple species of both homodimeric and heterodimeric phytochromes. The conservation of the phytochrome family throughout angiosperms suggests that heterodimeric red/far-red receptors may be present in many flowering plants. PMID- 15273291 TI - Watching the photosynthetic apparatus in native membranes. AB - Over the last 9 years, the structures of the various components of the bacterial photosynthetic apparatus or their homologues have been determined by x-ray crystallography to at least 4.8-A resolution. Despite this wealth of structural information on the individual proteins, there remains an urgent need to examine the architecture of the photosynthetic apparatus in intact photosynthetic membranes. Information on the arrangement of the different complexes in a native system will help us to understand the processes that ensure the remarkably high quantum efficiency of the system. In this work we report images obtained with an atomic force microscope of native photosynthetic membranes from the bacterium Rhodospirillum photometricum. Several proteins can be seen and identified at molecular resolution, allowing the analysis and modeling of the lateral organization of multiple components of the photosynthetic apparatus within a native membrane. Analysis of the distribution of the complexes shows that their arrangement is far from random, with significant clustering both of antenna complexes and core complexes. The functional significance of the observed distribution is discussed. PMID- 15273292 TI - Staphylococcal toxin induced preferential and prolonged in vivo deletion of innate-like B lymphocytes. AB - Contributing to host defenses from the adaptive immune system, splenic marginal zone (MZ) B cells, with their preactivated state and special topographical location, serve essential roles as primary defenders from blood-borne microbes. From studies designed to define the immunologic impact of protein A of Staphylococcus aureus (SpA), a virulence factor with targeted B cell antigen receptor-binding properties, we found that within minutes of in vivo exposure, SpA became surface associated with B lymphocytes and induced trafficking. Within several hours, MZ were completely effaced of affected B cells. This was rapidly followed by massive B cell apoptosis, with accelerated preferential deletion of targeted MZ B cells and impaired responsiveness to T independent immunogens. Subsequently, the temporal recovery of MZ B cells was significantly delayed compared to peripheral follicular B cells (B-2 cells). These studies elucidate the cellular program induced by a natural toxin that is shown to be highly efficient at depleting innate-like B cells important for defense from systemic infection. PMID- 15273293 TI - Arabidopsis thaliana GLN2-encoded glutamine synthetase is dual targeted to leaf mitochondria and chloroplasts. AB - In higher plants, photorespiratory Gly oxidation in leaf mitochondria yields ammonium in large amounts. Mitochondrial ammonium must somehow be recovered as glutamate in chloroplasts. As the first step in that recovery, we report glutamine synthetase (GS) activity in highly purified Arabidopsis thaliana mitochondria isolated from light-adapted leaf tissue. Leaf mitochondrial GS activity is further induced in response to either physiological CO(2) limitation or transient darkness. Historically, whether mitochondria are fully competent for oxidative phosphorylation in actively photorespiring leaves has remained uncertain. Here, we report that light-adapted, intact, leaf mitochondria supplied with Gly as sole energy source are fully competent for oxidative phosphorylation. Purified intact mitochondria efficiently use Gly oxidation (as sole energy, NH(3), and CO(2) source) to drive conversion of l-Orn to l-citrulline, an ATP dependent process. An A. thaliana genome-wide search for nuclear gene(s) encoding mitochondrial GS activity yielded a single candidate, GLN2. Stably transgenic A. thaliana ecotype Columbia plants expressing a p35S::GLN2::green fluorescent protein (GFP) chimeric reporter were constructed. When observed by laser scanning confocal microscopy, leaf mesophyll and epidermal tissue of transgenic plants showed punctate GFP fluorescence that colocalized with mitochondria. In immunoblot experiments, a 41-kD chimeric GLN2::GFP protein was present in both leaf mitochondria and chloroplasts of these stably transgenic plants. Therefore, the GLN2 gene product, heretofore labeled plastidic GS-2, functions in both leaf mitochondria and chloroplasts to faciliate ammonium recovery during photorespiration. PMID- 15273294 TI - Functional analysis of a 450-amino acid N-terminal fragment of phytochrome B in Arabidopsis. AB - Phytochrome, a major photoreceptor in plants, consists of two domains: the N terminal photosensory domain and the C-terminal domain. Recently, the 651-amino acid photosensory domain of phytochrome B (phyB) has been shown to act as a functional photoreceptor in the nucleus. The phytochrome (PHY) domain, which is located at the C-terminal end of the photosensory domain, is required for the spectral integrity of phytochrome; however, little is known about the signal transduction activity of this domain. Here, we have established transgenic Arabidopsis thaliana lines expressing an N-terminal 450-amino acid fragment of phyB (N450) lacking the PHY domain on a phyB-deficient background. Analysis of these plants revealed that N450 can act as an active photoreceptor when attached to a short nuclear localization signal and beta-glucuronidase. In vitro spectral analysis of reconstituted chromopeptides further indicated that the stability of the N450 Pfr form, an active form of phytochrome, is markedly reduced in comparison with the Pfr form of full-length phyB. Consistent with this, plants expressing N450 failed to respond to intermittent light applied at long intervals, indicating that N450 Pfr is short-lived in vivo. Taken together, our findings show that the PHY domain is dispensable for phyB signal transduction but is required for stabilizing the Pfr form of phyB. PMID- 15273296 TI - The Ustilago maydis a2 mating-type locus genes lga2 and rga2 compromise pathogenicity in the absence of the mitochondrial p32 family protein Mrb1. AB - The Ustilago maydis mrb1 gene specifies a mitochondrial matrix protein with significant similarity to mitochondrial p32 family proteins known from human and many other eukaryotic species. Compatible mrb1 mutant strains were able to mate and form dikaryotic hyphae; however, proliferation within infected tissue and the ability to induce tumor development of infected maize (Zea mays) plants were drastically impaired. Surprisingly, manifestation of the mrb1 mutant phenotype selectively depended on the a2 mating type locus. The a2 locus contains, in addition to pheromone signaling components, the genes lga2 and rga2 of unknown function. Deletion of lga2 in an a2Deltamrb1 strain fully restored pathogenicity, whereas pathogenicity was partially regained in an a2Deltamrb1Deltarga2 strain, implicating a concerted action between Lga2 and Rga2 in compromising pathogenicity in Deltamrb1 strains. Lga2 and Rga2 localized to mitochondria and Mrb1 interacted with Rga2 in the yeast two-hybrid system. Conditional expression of lga2 in haploid cells reduced vegetative growth, conferred mitochondrial fragmentation and mitochondrial DNA degradation, and interfered with respiratory activity. The consequences of lga2 overexpression depended on the expression strength and were greatly exacerbated in Deltamrb1 mutants. We propose that Lga2 interferes with mitochondrial fusion and that Mrb1 controls this activity, emphasizing a critical link between mitochondrial morphology and pathogenicity. PMID- 15273295 TI - Global transcription profiling reveals multiple sugar signal transduction mechanisms in Arabidopsis. AB - Complex and interconnected signaling networks allow organisms to control cell division, growth, differentiation, or programmed cell death in response to metabolic and environmental cues. In plants, it is known that sugar and nitrogen are critical nutrient signals; however, our understanding of the molecular mechanisms underlying nutrient signal transduction is very limited. To begin unraveling complex sugar signaling networks in plants, DNA microarray analysis was used to determine the effects of glucose and inorganic nitrogen source on gene expression on a global scale in Arabidopsis thaliana. In whole seedling tissue, glucose is a more potent signal in regulating transcription than inorganic nitrogen. In fact, other than genes associated with nitrate assimilation, glucose had a greater effect in regulating nitrogen metabolic genes than nitrogen itself. Glucose also regulated a broader range of genes, including genes associated with carbohydrate metabolism, signal transduction, and metabolite transport. In addition, a large number of stress responsive genes were also induced by glucose, indicating a role of sugar in environmental responses. Cluster analysis revealed significant interaction between glucose and nitrogen in regulating gene expression because glucose can modulate the effects of nitrogen and vise versa. Intriguingly, cycloheximide treatment appeared to disrupt glucose induction more than glucose repression, suggesting that de novo protein synthesis is an intermediary event required before most glucose induction can occur. Cross talk between sugar and ethylene signaling may take place on the transcriptional level because several ethylene biosynthetic and signal transduction genes are repressed by glucose, and the repression is largely unaffected by cycloheximide. Collectively, our global expression data strongly support the idea that glucose and inorganic nitrogen act as both metabolites and signaling molecules. PMID- 15273297 TI - Functional specialization amongst the Arabidopsis Toc159 family of chloroplast protein import receptors. AB - The initial stages of preprotein import into chloroplasts are mediated by the receptor GTPase Toc159. In Arabidopsis thaliana, Toc159 is encoded by a small gene family: atTOC159, atTOC132, atTOC120, and atTOC90. Phylogenetic analysis suggested that at least two distinct Toc159 subtypes, characterized by atToc159 and atToc132/atToc120, exist in plants. atTOC159 was strongly expressed in young, photosynthetic tissues, whereas atTOC132 and atTOC120 were expressed at a uniformly low level and so were relatively prominent in nonphotosynthetic tissues. Based on the albino phenotype of its knockout mutant, atToc159 was previously proposed to be a receptor with specificity for photosynthetic preproteins. To elucidate the roles of the other isoforms, we characterized Arabidopsis knockout mutants for each one. None of the single mutants had strong visible phenotypes, but toc132 toc120 double homozygotes appeared similar to toc159, indicating redundancy between atToc132 and atToc120. Transgenic complementation studies confirmed this redundancy but revealed little functional overlap between atToc132/atToc120 and atToc159 or atToc90. Unlike toc159, toc132 toc120 caused structural abnormalities in root plastids. Furthermore, when proteomics and transcriptomics were used to compare toc132 with ppi1 (a receptor mutant that is specifically defective in the expression, import, and accumulation of photosynthetic proteins), major differences were observed, suggesting that atToc132 (and atToc120) has specificity for nonphotosynthetic proteins. When both atToc159 and the major isoform of the other subtype, atToc132, were absent, an embryo-lethal phenotype resulted, demonstrating the essential role of Toc159 in the import mechanism. PMID- 15273298 TI - The Arabidopsis microtubule-associated protein AtMAP65-1: molecular analysis of its microtubule bundling activity. AB - The 65-kD microtubule-associated protein (MAP65) family is a family of plant microtubule-bundling proteins. Functional analysis is complicated by the heterogeneity within this family: there are nine MAP65 genes in Arabidopsis thaliana, AtMAP65-1 to AtMAP65-9. To begin the functional dissection of the Arabidopsis MAP65 proteins, we have concentrated on a single isoform, AtMAP65-1, and examined its effect on the dynamics of mammalian microtubules. We show that recombinant AtMAP65-1 does not promote polymerization and does not stabilize microtubules against cold-induced microtubule depolymerization. However, we show that it does induce microtubule bundling in vitro and that this protein forms 25 nm cross-bridges between microtubules. We further demonstrate that the microtubule binding region resides in the C-terminal half of the protein and that Ala409 and Ala420 are essential for the interaction with microtubules. Ala420 is a conserved amino acid in the AtMAP65 family and is mutated to Val in the cytokinesis-defective mutant pleiade-4 of the AtMAP65-3/PLEIADE gene. We show that AtMAP65-1 can form dimers and that a region in the N terminus is responsible for this activity. Neither the microtubule binding region nor the dimerization region alone could induce microtubule bundling, strongly suggesting that dimerization is necessary to produce the microtubule cross-bridges. In vivo, AtMAP65-1 is ubiquitously expressed both during the cell cycle and in all plant organs and tissues with the exception of anthers and petals. Moreover, using an antiserum raised to AtMAP65-1, we show that AtMAP65-1 binds microtubules at specific stages of the cell cycle. PMID- 15273299 TI - Determination of enzyme mechanisms by molecular dynamics: studies on quinoproteins, methanol dehydrogenase, and soluble glucose dehydrogenase. AB - Molecular dynamics (MD) simulations have been carried out to study the enzymatic mechanisms of quinoproteins, methanol dehydrogenase (MDH), and soluble glucose dehydrogenase (sGDH). The mechanisms of reduction of the orthoquinone cofactor (PQQ) of MDH and sGDH involve concerted base-catalyzed proton abstraction from the hydroxyl moiety of methanol or from the 1-hydroxyl of glucose, and hydride equivalent transfer from the substrate to the quinone carbonyl carbon C5 of PQQ. The products of methanol and glucose oxidation are formaldehyde and glucolactone, respectively. The immediate product of PQQ reduction, PQQH- [-HC5(O-)-C4(=O)-] and PQQH [-HC5(OH)-C4(=O)-] converts to the hydroquinone PQQH2 [-C5(OH)=C4(OH)-]. The main focus is on MD structures of MDH * PQQ * methanol, MDH * PQQH-, MDH * PQQH, sGDH * PQQ * glucose, sGDH * PQQH- (glucolactone, and sGDH * PQQH. The reaction PQQ-->PQQH- occurs with Glu 171-CO2- and His 144-Im as the base species in MDH and sGDH, respectively. The general-base-catalyzed hydroxyl proton abstraction from substrate concerted with hydride transfer to the C5 of PQQ is assisted by hydrogen-bonding to the C5=O by Wat1 and Arg 324 in MDH and by Wat89 and Arg 228 in sGDH. Asp 297-COOH would act as a proton donor for the reaction PQQH(-)-->PQQH, if formed by transfer of the proton from Glu 171-COOH to Asp 297 CO2- in MDH. For PQQH-->PQQH2, migration of H5 to the C4 oxygen may be assisted by a weak base like water (either by crystal water Wat97 or bulk solvent, hydrogen-bonded to Glu 171-CO2- in MDH and by Wat89 in sGDH). PMID- 15273300 TI - Caspase activation, inhibition, and reactivation: a mechanistic view. AB - Caspases, a unique family of cysteine proteases, execute programmed cell death (apoptosis). Caspases exist as inactive zymogens in cells and undergo a cascade of catalytic activation at the onset of apoptosis. The activated caspases are subject to inhibition by the inhibitor-of-apoptosis (IAP) family of proteins. This inhibition can be effectively removed by diverse proteins that share an IAP binding tetrapeptide motif. Recent structural and biochemical studies have revealed the underlying molecular mechanisms for these processes in mammals and in Drosophila. This paper reviews these latest advances. PMID- 15273301 TI - Orientation and helical conformation of a tissue-specific hunter-killer peptide in micelles. AB - Hunter-killer peptides are chimeric synthetic peptides that selectively target specific cell types for an apoptotic death. These peptides, which are models for potential therapeutics, contain a homing sequence for receptor-mediated interactions and a pro-apoptotic sequence. Homing domains have been designed to target angiogenic tumor cells, prostate cells, arthritic tissue and, most recently, adipose tissue. After a receptor-mediated internalization, the apoptotic sequence, which contains D-enantiomer amino acids, initiates apoptosis through mitochondrial membrane disruption. We have begun structure and functional studies on a peptide (HKP1) that specifically targets angiogenic tumor cells for apoptosis. As a model for mitochondrial membrane disruption, we have examined peptide-induced leakage of a calcein fluorophore from large unilamellar vesicles. These experiments demonstrate more potent leakage activity by HKP1 than the peptide lacking the homing domain. Circular dichroism and 2D homonuclear NMR experiments demonstrate that this tumor-specific HKP adopts a left-handed amphipathic helix in association with dodecylphosphorylcholine micelles in a parallel orientation to the lipid-water interface with the homing domain remaining exposed to solvent. The amphipathic helix of the apoptotic domain orients with nonpolar leucine and alanine residues inserting most deeply into the lipid environment. PMID- 15273302 TI - Insights into the DNA repair process by the formamidopyrimidine-DNA glycosylase investigated by molecular dynamics. AB - Formamidopyrimidine-DNA glycosylase (Fpg) identifies and removes 8-oxoguanine from DNA. All of the X-ray structures of Fpg complexed to an abasic site containing DNA exhibit a common disordered region present in the C-terminal domain of the enzyme. However, this region is believed to be involved in the damaged base binding site when the initial protein/DNA complex is formed. The dynamic behavior of the disordered polypeptide (named Loop) in relation to the supposed scenario for the DNA repair mechanism was investigated by molecular dynamics on different models, derived from the X-ray structure of Lactococcus lactis Fpg bound to an abasic site analog-containing DNA and of Bacillus stearothermophilus Fpg bound to 8-oxoG. This study shows that the presence of the damaged base influences the dynamics of the whole enzyme and that the Loop location is dependent on the presence and on the conformation of the 8-oxoG in its binding site. In addition, from our results, the conformation of the 8-oxoG seems to be favored in syn in the L. lactis models, in agreement with the available X-ray structure from B. stearothermophilus Fpg and with a possible catalytic role of the flexibility of the Loop region. PMID- 15273303 TI - NMR structure of CXCR3 binding chemokine CXCL11 (ITAC). AB - CXCL11 (ITAC) is one of three chemokines known to bind the receptor CXCR3, the two others being CXCL9 (Mig) and CXCL10 (IP-10). CXCL11 differs from the other CXCR3 ligands in both the strength and the particularities of its receptor interactions: It has a higher affinity, is a stronger agonist, and behaves differently when critical N-terminal residues are deleted. The structure of CXCL11 was determined using solution NMR to allow comparison with that of CXCL10 and help elucidate the source of the differences. CXCL11 takes on the canonical chemokine fold but exhibits greater conformational flexibility than has been observed for related chemokines under the same sample conditions. Unlike related chemokines such as IP-10 and IL-8, ITAC does not appear to form dimers at millimolar concentrations. The origin for this behavior can be found in the solution structure, which indicates a beta-bulge in beta-strand 1 that distorts the dimerization interface used by other CXC chemokines. PMID- 15273304 TI - Experimentally biased model structure of the Hsc70/auxilin complex: substrate transfer and interdomain structural change. AB - A model structure of the Hsc70/auxilin complex has been constructed to gain insight into interprotein substrate transfer and ATP hydrolysis induced conformational changes in the multidomain Hsc70 structure. The Hsc70/auxilin system, which is a member of the Hsp70/Hsp40 chaperone system family, uncoats clathrin-coated vesicles in an ATP hydrolysis-driven process. Incorporating previous results from NMR and mutant binding studies, the auxilin J-domain was docked into the Hsc70 ATPase domain lower cleft using rigid backbone/flexible side chain molecular dynamics, and the Hsc70 substrate binding domain was docked by a similar procedure. For comparison, J-domain and substrate binding domain docking sites were obtained by the rigid-body docking programs DOT and ZDOCK, filtered and ranked by the program ClusPro, and relaxed using the same rigid backbone/flexible side chain dynamics. The substrate binding domain sites were assessed in terms of conserved surface complementarity and feasibility in the context of substrate transfer, both for auxilin and another Hsp40 protein, Hsc20. This assessment favors placement of the substrate binding domain near D152 on the ATPase domain surface adjacent to the J-domain invariant HPD segment, with the Hsc70 interdomain linker in the lower cleft. Examining Hsc70 interdomain energetics, we propose that long-range electrostatic interactions, perhaps due to a difference in the pKa values of bound ATP and ADP, could play a major role in the structural change induced by ATP hydrolysis. Interdomain electrostatic interactions also appear to play a role in stimulation of ATPase activity due to J-domain binding and substrate binding by Hsc70. PMID- 15273305 TI - A classification of disulfide patterns and its relationship to protein structure and function. AB - We report a detailed classification of disulfide patterns to further understand the role of disulfides in protein structure and function. The classification is applied to a unique searchable database of disulfide patterns derived from the SwissProt and Pfam databases. The disulfide database contains seven times the number of publicly available disulfide annotations. Each disulfide pattern in the database captures the topology and cysteine spacing of a protein domain. We have clustered the domains by their disulfide patterns and visualized the results using a novel representation termed the "classification wheel." The classification is applied to 40,620 protein domains with 2-10 disulfides. The effectiveness of the classification is evaluated by determining the extent to which proteins of similar structure and function are grouped together through comparison with the SCOP and Pfam databases, respectively. In general, proteins with similar disulfide patterns have similar structure and function, even in cases of low sequence similarity, and we illustrate this with specific examples. Using a measure of disulfide topology complexity, we find that there is a predominance of less complex topologies. We also explored the importance of loss or addition of disulfides to protein structure and function by linking classification wheels through disulfide subpattern comparisons. This classification, when coupled with our disulfide database, will serve as a useful resource for searching and comparing disulfide patterns, and understanding their role in protein structure, folding, and stability. Proteins in the disulfide clusters that do not contain structural information are prime candidates for structural genomics initiatives, because they may correspond to novel structures. PMID- 15273306 TI - Evolutionary constraints associated with functional specificity of the CMGC protein kinases MAPK, CDK, GSK, SRPK, DYRK, and CK2alpha. AB - Amino acid residues associated with functional specificity of cyclin-dependent kinases (CDKs), mitogen-activated protein kinases (MAPKs), glycogen synthase kinases (GSKs), and CDK-like kinases (CLKs), which are collectively termed the CMGC group, were identified by categorizing and quantifying the selective constraints acting upon these proteins during evolution. Many constraints specific to CMGC kinases correspond to residues between the N-terminal end of the activation segment and a CMGC-conserved insert segment associated with coprotein binding. The strongest such constraint is imposed on a "CMGC-arginine" near the substrate phosphorylation site with a side chain that plays a role both in substrate recognition and in kinase activation. Two nearby buried waters, which are also present in non-CMGC kinases, typically position the main chain of this arginine relative to the catalytic loop. These and other CMGC-specific features suggest a structural linkage between coprotein binding, substrate recognition, and kinase activation. Constraints specific to individual subfamilies point to mechanisms for CMGC kinase specialization. Within casein kinase 2alpha (CK2alpha), for example, the binding of one of the buried waters appears prohibited by the side chain of a leucine that is highly conserved within CK2alpha and that, along with substitution of lysine for the CMGC-arginine, may contribute to the broad substrate specificity of CK2alpha by relaxing characteristically conserved, precise interactions near the active site. This leucine is replaced by a conserved isoleucine or valine in other CMGC kinases, thereby illustrating the potential functional significance of subtle amino acid substitutions. Analysis of other CMGC kinases similarly suggests candidate family specific residues for experimental follow-up. PMID- 15273307 TI - Solution structure of the RWD domain of the mouse GCN2 protein. AB - GCN2 is the alpha-subunit of the only translation initiation factor (eIF2alpha) kinase that appears in all eukaryotes. Its function requires an interaction with GCN1 via the domain at its N-terminus, which is termed the RWD domain after three major RWD-containing proteins: RING finger-containing proteins, WD-repeat containing proteins, and yeast DEAD (DEXD)-like helicases. In this study, we determined the solution structure of the mouse GCN2 RWD domain using NMR spectroscopy. The structure forms an alpha + beta sandwich fold consisting of two layers: a four-stranded antiparallel beta-sheet, and three side-by-side alpha helices, with an alphabetabetabetabetaalphaalpha topology. A characteristic YPXXXP motif, which always occurs in RWD domains, forms a stable loop including three consecutive beta-turns that overlap with each other by two residues (triple beta-turn). As putative binding sites with GCN1, a structure-based alignment allowed the identification of several surface residues in alpha-helix 3 that are characteristic of the GCN2 RWD domains. Despite the apparent absence of sequence similarity, the RWD structure significantly resembles that of ubiquitin conjugating enzymes (E2s), with most of the structural differences in the region connecting beta-strand 4 and alpha-helix 3. The structural architecture, including the triple beta-turn, is fundamentally common among various RWD domains and E2s, but most of the surface residues on the structure vary. Thus, it appears that the RWD domain is a novel structural domain for protein-binding that plays specific roles in individual RWD-containing proteins. PMID- 15273308 TI - Biophysical and kinetic analysis of wild-type and site-directed mutants of the isolated and native dehydroquinate synthase domain of the AROM protein. AB - Dehydroquinate synthase (DHQS) is the N-terminal domain of the pentafunctional AROM protein that catalyses steps 2 to 7 in the shikimate pathway in microbial eukaryotes. DHQS converts 3-deoxy-D-arabino-heptulosonate-7-phosphate (DAHP) to dehydroquinate in a reaction that includes alcohol oxidation, phosphate beta elimination, carbonyl reduction, ring opening, and intramolecular aldol condensation. Kinetic analysis of the isolated DHQS domains with the AROM protein showed that for the substrate DAHP the difference in Km is less than a factor of 3, that the turnover numbers differed by 24%, and that the Km for NAD+ differs by a factor of 3. Isothermal titration calorimetry revealed that a second (inhibitory) site for divalent metal binding has an approximately 4000-fold increase in KD compared to the catalytic binding site. Inhibitor studies have suggested the enzyme could act as a simple oxidoreductase with several of the reactions occurring spontaneously, whereas structural studies have implied that DHQS participates in all steps of the reaction. Analysis of site-directed mutants experimentally test and support this latter hypothesis. Differential scanning calorimetry, circular dichroism spectroscopy, and molecular exclusion chromatography demonstrate that the mutant DHQS retain their secondary and quaternary structures and their ligand binding capacity. R130K has a 135-fold reduction in specific activity with DAHP and a greater than 1100-fold decrease in the kcat/Km ratio, whereas R130A is inactive. PMID- 15273309 TI - Detecting hidden sequence propensity for amyloid fibril formation. AB - The preponderance of evidence implicates protein misfolding in many unrelated human diseases. In all cases, normal correctly folded proteins transform from their proper native structure into an abnormal beta-rich structure known as amyloid fibril. Here we introduce a computational algorithm to detect nonnative (hidden) sequence propensity for amyloid fibril formation. Analyzing sequence structure relationships in terms of tertiary contact (TC), we find that the hidden beta-strand propensity of a query local sequence can be quantitatively estimated from the secondary structure preferences of template sequences of known secondary structure found in regions of high TC. The present method correctly pinpoints the minimal peptide fragment shown experimentally as the likely local mediator of amyloid fibril formation in beta-amyloid peptide, islet amyloid polypeptide (hIAPP), alpha-synuclein, and human acetylcholinesterase (AChE). It also found previously unrecognized beta-strand propensities in the prototypical helical protein myoglobin that has been reported as amyloidogenic. Analysis of 2358 nonhomologous protein domains provides compelling evidence that most proteins contain sequences with significant hidden beta-strand propensity. The present method may find utility in many medically relevant applications, such as the engineering of protein sequences and the discovery of therapeutic agents that specifically target these sequences for the prevention and treatment of amyloid diseases. PMID- 15273310 TI - The comparative study on the solution structures of the oxidized bovine microsomal cytochrome b5 and mutant V45H. AB - A comparative study on the solution structures of bovine microsomal cytochrome b5 (Tb5) and the mutant V45H has been achieved by 1D and 2D 1H-NMR spectroscopy to clarify the differences in the solution conformations between these two proteins. The results reveal that the global folding of the V45H mutant in solution is unchanged, but the subtle changes exist in the orientation of the axial ligand His39, and heme vinyl groups. The side chain of His45 in V45H mutant extends to the outer edge of the heme pocket leaving a cavity at the site originally occupied by the inner methyl group of Val45 residue. In addition, the imidazole ring of axial ligand His39 rotates counterclockwise by approximately 3 degrees around the His-Fe-His axis, and the 4-heme vinyl group turns to the space vacated by the removed side chain due to the mutation. Furthermore, the helix III of the heme pocket undergoes outward displacement, while the linkage between helix II and III is shifted leftward. These observations are not only consistent with the pattern of the pseudocontact shifts of the heme protons, but also well account for the lower stability of V45H mutant against heat and urea. PMID- 15273311 TI - Proteome-wide functional classification and identification of prokaryotic transmembrane proteins by transmembrane topology similarity comparison. AB - We propose a new method for classifying and identifying transmembrane (TM) protein functions in proteome-scale by applying a single-linkage clustering method based on TM topology similarity, which is calculated simply from comparing the lengths of loop regions. In this study, we focused on 87 prokaryotic TM proteomes consisting of 31 proteobacteria, 22 gram-positive bacteria, 19 other bacteria, and 15 archaea. Prior to performing the clustering, we first categorized individual TM protein sequences as "known," "putative" (similar to "known" sequences), or "unknown" by using the homology search and the sequence similarity comparison against SWISS-PROT to assess the current status of the functional annotation of the TM proteomes based on sequence similarity only. More than three-quarters, that is, 75.7% of the TM protein sequences are functionally "unknown," with only 3.8% and 20.5% of them being classified as "known" and "putative," respectively. Using our clustering approach based on TM topology similarity, we succeeded in increasing the rate of TM protein sequences functionally classified and identified from 24.3% to 60.9%. Obtained clusters correspond well to functional superfamilies or families, and the functional classification and identification are successfully achieved by this approach. For example, in an obtained cluster of TM proteins with six TM segments, 109 sequences out of 119 sequences annotated as "ATP-binding cassette transporter" are properly included and 122 "unknown" sequences are also contained. PMID- 15273312 TI - The C-terminal domain of dimeric serine hydroxymethyltransferase plays a key role in stabilization of the quaternary structure and cooperative unfolding of protein: domain swapping studies with enzymes having high sequence identity. AB - The serine hydroxymethyltransferase from Bacillus subtilis (bsSHMT) and B. stearothermophilus (bstSHMT) are both homodimers and share approximately 77% sequence identity; however, they show very different thermal stabilities and unfolding pathways. For investigating the role of N- and C-terminal domains in stability and unfolding of dimeric SHMTs, we have swapped the structural domains between bs- and bstSHMT and generated the two novel chimeric proteins bsbstc and bstbsc, respectively. The chimeras had secondary structure, tyrosine, and pyridoxal-5'-phosphate microenvironment similar to that of the wild-type proteins. The chimeras showed enzymatic activity slightly higher than that of the wild-type proteins. Interestingly, the guanidium chloride (GdmCl)-induced unfolding showed that unlike the wild-type bsSHMT, which undergoes dissociation of native dimer into monomers at low guanidium chloride (GdmCl) concentration, resulting in a non-cooperative unfolding of enzyme, its chimera bsbstc, having the C-terminal domain of bstSHMT was resistant to low GdmCl concentration and showed a GdmCl-induced cooperative unfolding from native dimer to unfolded monomer. In contrast, the wild-type dimeric bstSHMT was resistant to low GdmCl concentration and showed a GdmCl-induced cooperative unfolding, whereas its chimera bstbsc, having the C- terminal domain of bsSHMT, showed dissociation of native dimer into monomer at low GdmCl concentration and a GdmCl-induced non cooperative unfolding. These results clearly demonstrate that the C-terminal domain of dimeric SHMT plays a vital role in stabilization of the oligomeric structure of the native enzyme hence modulating its unfolding pathway. PMID- 15273313 TI - Kinetic traps in the folding/unfolding of procaspase-1 CARD domain. AB - We have examined the folding and unfolding of the caspase recruitment domain of procaspase-1 (CP1-CARD), a member of the alpha-helical Greek key protein family. The equilibrium folding/unfolding of CP1-CARD is described by a two-state mechanism, and the results show CP1-CARD is marginally stable with a DeltaG(H2O) of 1.1 +/- 0.2 kcal/mole and an m-value of 0.65 +/- 0.06 kcal/mole/M (10 mM Tris HCl at pH 8.0, 1 mM DTT, 25 degrees C). Consistent with the equilibrium folding data, CP1-CARD is a monomer in solution when examined by size exclusion chromatography. Single-mixing stopped-flow refolding and unfolding studies show that CP1-CARD folds and unfolds rapidly, with no detectable slow phases, and the reactions appear to reach equilibrium within 10 msec. However, double jump kinetic experiments demonstrate the presence of an unfolded-like intermediate during unfolding. The intermediate converts to the fully unfolded conformation with a half-time of 10 sec. Interrupted refolding studies demonstrate the presence of one or more nativelike intermediates during refolding, which convert to the native conformation with a half-time of about 60 sec. Overall, the data show that both unfolding and refolding processes are slow, and the pathways contain kinetically trapped species. PMID- 15273314 TI - Global analysis of three-state protein unfolding data. AB - A new method for analyzing three-state protein unfolding equilibria is described that overcomes the difficulties created by direct effects of denaturants on circular dichroism (CD) and fluorescence spectra of the intermediate state. The procedure begins with a singular value analysis of the data matrix to determine the number of contributing species and perturbations. This result is used to choose a fitting model and remove all spectra from the fitting equation. Because the fitting model is a product of a matrix function which is nonlinear in the thermodynamic parameters and a matrix that is linear in the parameters that specify component spectra, the problem is solved with a variable projection algorithm. Advantages of this procedure are perturbation spectra do not have to be estimated before fitting, arbitrary assumptions about magnitudes of parameters that describe the intermediate state are not required, and multiple experiments involving different spectroscopic techniques can be simultaneously analyzed. Two tests of this method were performed: First, simulated three-state data were analyzed, and the original and recovered thermodynamic parameters agreed within one standard error, whereas recovered and original component spectra agreed within 0.5%. Second, guanidine-induced unfolding titrations of the human retinoid X-receptor ligand-binding domain were analyzed according to a three-state model. The standard unfolding free energy changes in the absence of guanidine and the guanidine concentrations at zero free-energy change for both transitions were determined from a joint analysis of fluorescence and CD spectra. Realistic spectra of the three protein states were also obtained. PMID- 15273315 TI - Probing folding and fluorescence quenching in human gammaD crystallin Greek key domains using triple tryptophan mutant proteins. AB - Human gammaD crystallin (HgammaD-Crys), a major component of the human eye lens, is a 173-residue, primarily beta-sheet protein, associated with juvenile and mature-onset cataracts. HgammaD-Crys has four tryptophans, with two in each of the homologous Greek key domains, which are conserved throughout the gamma crystallin family. HgammaD-Crys exhibits native-state fluorescence quenching, despite the absence of ligands or cofactors. The tryptophan absorption and fluorescence quenching may influence the lens response to ultraviolet light or the protection of the retina from ambient ultraviolet damage. To provide fluorescence reporters for each quadrant of the protein, triple mutants, each containing three tryptophan-to-phenylalanine substitutions and one native tryptophan, have been constructed and expressed. Trp 42-only and Trp 130-only exhibited fluorescence quenching between the native and denatured states typical of globular proteins, whereas Trp 68-only and Trp 156-only retained the anomalous quenching pattern of wild-type HgammaD-Crys. The three-dimensional structure of HgammaD-Crys shows Tyr/Tyr/His aromatic cages surrounding Trp 68 and Trp 156 that may be the source of the native-state quenching. During equilibrium refolding/unfolding at 37 degrees C, the tryptophan fluorescence signals indicated that domain I (W42-only and W68-only) unfolded at lower concentrations of GdnHCl than domain II (W130-only and W156-only). Kinetic analysis of both the unfolding and refolding of the triple-mutant tryptophan proteins identified an intermediate along the HgammaD-Crys folding pathway with domain I unfolded and domain II intact. This species is a candidate for the partially folded intermediate in the in vitro aggregation pathway of HgammaD-Crys. PMID- 15273316 TI - Double-stranded DNA bacteriophage prohead protease is homologous to herpesvirus protease. AB - Double-stranded DNA bacteriophages and herpesviruses assemble their heads in a similar fashion; a pre-formed precursor called a prohead or procapsid undergoes a conformational transition to give rise to a mature head or capsid. A virus encoded prohead or procapsid protease is often required in this maturation process. Through computational analysis, we infer homology between bacteriophage prohead proteases (MEROPS families U9 and U35) and herpesvirus protease (MEROPS family S21), and unify them into a procapsid protease superfamily. We also extend this superfamily to include an uncharacterized cluster of orthologs (COG3566) and many other phage or bacteria-encoded hypothetical proteins. On the basis of this homology and the herpesvirus protease structure and catalytic mechanism, we predict that bacteriophage prohead proteases adopt the herpesvirus protease fold and exploit a conserved Ser and His residue pair in catalysis. Our study provides further support for the proposed evolutionary link between dsDNA bacteriophages and herpesviruses. PMID- 15273317 TI - Asymmetric amino acid compositions of transmembrane beta-strands. AB - In contrast to water-soluble proteins, membrane proteins reside in a heterogeneous environment, and their surfaces must interact with both polar and apolar membrane regions. As a consequence, the composition of membrane proteins' residues varies substantially between the membrane core and the interfacial regions. The amino acid compositions of helical membrane proteins are also known to be different on the cytoplasmic and extracellular sides of the membrane. Here we report that in the 16 transmembrane beta-barrel structures, the amino acid compositions of lipid-facing residues are different near the N and C termini of the individual strands. Polar amino acids are more prevalent near the C termini than near the N termini, and hydrophobic amino acids show the opposite trend. We suggest that this difference arises because it is easier for polar atoms to escape from the apolar regions of the bilayer at the C terminus of a beta-strand. This new characteristic of beta-barrel membrane proteins enhances our understanding of how a sequence encodes a membrane protein structure and should prove useful in identifying and predicting the structures of trans-membrane beta barrels. PMID- 15273320 TI - Specificity of the STAR/GSG domain protein Qk1: implications for the regulation of myelination. AB - Inadequate formation and maintenance of myelin is the basis for several neurodegenerative disorders, including leukodystrophy and multiple sclerosis. In mice, oligodendrocyte differentiation and subsequent formation of myelin requires the Quaking gene. Mutation of this gene leads to embryonic lethality or to a trembling phenotype characteristic of dysmyelination. Quaking encodes Qk1, a member of the highly conserved STAR/GSG family of RNA-binding proteins that function as master developmental regulators in higher eukaryotes. Qk1 has been implicated in the regulation of alternative splicing, stability, and translation control of mRNAs that code for myelin structural components in glial cells. We have used quantitative gel mobility shift and fluorescence polarization assays to define the nucleotide sequence specificity of the Qk1 STAR/GSG domain, and to probe the interaction between Qk1 and the 3'-untranslated region (UTR) of myelin basic protein (MBP) mRNA. The results show that Qk1 recognizes a hexanucleotide consensus element that is similar although not identical to the specificity determinant recognized by the Caenorhabditis elegans STAR/GSG protein GLD-1. Several consensus sites are present in the 3'-UTR of MBP mRNA. The highest affinity site is located within the RNA localization region, suggesting a possible role for Qk1 in restricting MBP mRNA to the myelin compartment. PMID- 15273321 TI - High-affinity binding site for a group II intron-encoded reverse transcriptase/maturase within a stem-loop structure in the intron RNA. AB - Mobile group II introns encode proteins that have reverse transcriptase and maturase activities and bind specifically to the intron RNA to promote both RNA splicing and intron mobility. Previous studies with the Lactococcus lactis Ll.LtrB intron showed that the intron-encoded protein (LtrA) has a high-affinity binding site in intron subdomain DIVa, an idiosyncratic structure containing the translation initiation region of the LtrA open reading frame, and that this binding site consists of a small stem-loop emanating from a purine-rich internal loop. The binding of LtrA to DIVa is important for translational regulation, RNA splicing, and intron mobility. Here, we show by in vitro selection that part of the purine-rich internal loop can be closed by base pairing, enabling the LtrA binding site to be represented as an extended stem-loop structure with a bulged A (A556) required for tight binding of LtrA. The deletion or pairing of A556 has relatively little effect on maturase-promoted RNA splicing, but significantly inhibits intron mobility. The wild-type DIVa structure has a second bulged A (A553), which is selected against in tightly binding variants. As expected from the selection, the deletion or pairing of A553 results in tighter binding of LtrA, but surprisingly, also inhibits intron mobility. These findings suggest that the binding of LtrA to DIVa is delicately balanced, so that either too weak or too tight binding can be deleterious. The nature of the maturase/DIVa interaction and its role in translational regulation are reminiscent of the coat protein/RNA hairpin interactions of single-stranded RNA phages. PMID- 15273322 TI - Functional analysis of mRNA scavenger decapping enzymes. AB - Eukaryotic cells primarily utilize exoribonucleases and decapping enzymes to degrade their mRNA. Two major decapping enzymes have been identified. The hDcp2 protein catalyzes hydrolysis of the 5' cap linked to an RNA moiety, whereas the scavenger decapping enzyme, DcpS, functions on a cap structure lacking the RNA moiety. DcpS is a member of the histidine triad (HIT) family of hydrolases and catalyzes the cleavage of m7GpppN. HIT proteins are homodimeric and contain two conserved 100-amino-acid HIT fold domains with independent active sites that are each sufficient to bind and hydrolyze cognate substrates. We carried out a functional characterization of the DcpS enzyme and demonstrate that unlike previously described HIT proteins, DcpS is a modular protein that requires both the core HIT fold at the carboxyl-terminus and sequences at the amino-terminus of the protein for cap binding and hydrolysis. Interestingly, DcpS can efficiently compete for and hydrolyze the cap structure even in the presence of excess eIF4E, implying that DcpS could function to alleviate the accumulation of complexes between eIF4E and cap structure that would otherwise accumulate following mRNA decay. Using immunofluorescence microscopy, we demonstrate that DcpS is predominantly a nuclear protein, with low levels of detected protein in the cytoplasm. Furthermore, analysis of the endogenous hDcp2 protein reveals that in addition to the cytoplasmic foci, it is also present in the nucleus. These data reveal that both decapping enzymes are contained in the nuclear compartment, indicating that they may fulfill a greater function in the nucleus than previously appreciated. PMID- 15273323 TI - Detection of genome-scale ordered RNA structure (GORS) in genomes of positive stranded RNA viruses: Implications for virus evolution and host persistence. AB - Discrete RNA secondary and higher-order structures, typically local in extent, play a fundamental role in RNA virus replication. Using new bioinformatics analysis methods, we have identified genome-scale ordered RNA structure (GORS) in many genera and families of positive-strand animal and plant RNA viruses. There was remarkably variability between genera that possess this characteristic; for example, hepaciviruses in the family Flaviviridae show evidence for extensive internal base-pairing throughout their coding sequences that was absent in both the related pestivirus and flavivirus genera. Similar genus-associated variability was observed in the Picornaviridae, the Caliciviridae, and many plant virus families. The similarity in replication strategies between genera in each of these families rules out a role for GORS in a fundamentally conserved aspect of this aspect of the virus life cycle. However, in the Picornaviridae, Flaviviridae, and Caliciviridae, the existence of GORS correlated strongly with the ability of each genus to persist in their natural hosts. This raises the intriguing possibility of a role for GORS in the modulation of innate intracellular defense mechanisms (and secondarily, the acquired immune system) triggered by double-stranded RNA, analogous in function to the expression of structured RNA transcripts by large DNA viruses. Irrespective of function, the observed evolutionary conservation of GORS in many viruses imposes a considerable constraint on genome plasticity and the consequent narrowing of sequence space in which neutral drift can occur. These findings potentially reconcile the rapid evolution of RNA viruses over short periods with the documented examples of extreme conservatism evident from their intimate coevolution with their hosts. PMID- 15273324 TI - Ligand-induced changes in 2-aminopurine fluorescence as a probe for small molecule binding to HIV-1 TAR RNA. AB - Replication of human immunodeficiency virus type 1 (HIV-1) is regulated in part through an interaction between the virally encoded trans-activator protein Tat and the trans-activator responsive region (TAR) of the viral RNA genome. Because TAR is highly conserved and its interaction with Tat is required for efficient viral replication, it has received much attention as an antiviral drug target. Here, we report a 2-aminopurine (2-AP) fluorescence-based assay for evaluating potential TAR inhibitors. Through selective incorporation of 2-AP within the bulge (C23 or U24) of a truncated form of the TAR sequence (delta TAR-ap23 and delta TAR-ap24), binding of argininamide, a 24-residue arginine-rich peptide derived from Tat, and Neomycin has been characterized using steady-state fluorescence. Binding of argininamide to the 2-AP deltaTAR constructs results in a four- to 11-fold increase in fluorescence intensity, thus providing a sensitive reporter of that interaction (KD approximately 1 mM). Similarly, binding of the Tat peptide results in an initial 14-fold increase in fluorescence (KD approximately 25 nM), but is then followed by a slight decrease that is attributed to an additional, lower-affinity association(s). Using the deltaTAR ap23 and TAR-ap24 constructs, two classes of Neomycin binding sites are detected; the first molecule of antibiotic binds as a noncompetitive inhibitor of Tat/argininamide (KD approximately 200 nM), whereas the second, more weakly bound molecule(s) becomes associated in a presumably nonspecific manner (KD approximately 4 microM). Taken together, the results demonstrate that the 2-AP fluorescence-detected binding assays provide accurate and general methods for quantitatively assessing TAR interactions. PMID- 15273325 TI - Chemical synthesis and binding activity of the trypanosomatid cap-4 structure. AB - Leishmania and other trypanosomatids are early eukaryotes that possess unusual molecular features, including polycistronic transcription and trans-splicing of pre-mRNAs. The spliced leader RNA (SL RNA) is joined to the 5' end of all mRNAs, thus donating a 5' cap that is characterized by complex modifications. In addition to the conserved m7GTP, linked via a 5'-5'-triphosphate bound to the first nucleoside of the mRNA, the trypanosomatid 5' cap includes 2'-O methylations on the first four ribose moieties and unique base methylations on the first adenine and the fourth uracil, resulting in the cap-4 structure, m7Gpppm3(6,6,2')Apm2'Apm2' Cpm2(3,2')U, as reported elsewhere previously. A library of analogs that mimic the cap structure to different degrees has been synthesized. Their differential affinities to the cap binding proteins make them attractive compounds for studying the molecular basis of cap recognition, and in turn, they may have potential therapeutic significance. The interactions between cap analogs and eIF4E, a cap-binding protein that plays a key role in initiation of translation, can be monitored by measuring intrinsic fluorescence quenching of the tryptophan residues. In the present communication we describe the multistep synthesis of the trypanosomatid cap-4 structure. The 5' terminal mRNA tetranucleotide fragment (pm3(6,6,2')Apm2'Apm2'Cpm2(3,2')U) was synthesized by the phosphoramidite solid phase method. After deprotection and purification, the 5'-phosphorylated tetranucleotide was chemically coupled with m7GDP to yield the cap-4 structure. Biological activity of this newly synthesized compound was confirmed in binding studies with eIF4E from Leishmania that we recently cloned (LeishIF4E-1), using the fluorescence time-synchronized titration method. PMID- 15273326 TI - Optimization of apolipoprotein B mRNA editing by APOBEC1 apoenzyme and the role of its auxiliary factor, ACF. AB - Expression and purification to homogeneity of the apolipoprotein B mRNA editing subunit, APOBEC1, has allowed the demonstration that this apoenzyme has considerable residual enzymatic activity on a minimal apoB mRNA substrate, even in the absence of any auxiliary factors. Assay of this activity as a function of various experimental conditions has led to substantial optimization of assay conditions through the use of incomplete factorial and response surface experiments. Surprisingly, the apoenzyme is thermostable, and has a temperature optimum near 45 degrees C. We have used these optimized conditions, to assess steady-state kinetic parameters for APOBEC1 mRNA editing activity with and without the auxiliary factor, ACF. An important effect of the auxiliary factor is to broaden the temperature range of APOBEC1 activity, lowering the optimal temperature and enabling it to function optimally at lower temperatures. A model consistent with this observation is that at lower temperatures ACF promotes a conformational transition in the RNA substrate that occurs spontaneously at higher temperature. Notably, the substantial RNA editing activity of APOBEC1 alone may be responsible for the "hyperediting" observed upon overexpression of APOBEC1 in transgenic mice. PMID- 15273327 TI - Solution structure of an RNA stem-loop derived from the 3' conserved region of eel LINE UnaL2. AB - The eel long interspersed element (LINE) UnaL2 and its partner short interspersed element (SINE) share a conserved 3' tail containing a stem-loop that is critical for their retrotransposition. Presumably, the first step of retrotransposition is the recognition of their 3' tails by UnaL2-encoded reverse transcriptase. The solution structure of a 17-nucleotide RNA derived from the 3' tail of UnaL2 was determined by NMR. The GGAUA loop forms a specific structure in which the uridine is exposed to solvent with the third and fifth adenosines stacked. A sharp turn in the phosphodiester backbone occurs between the second guanosine and third adenosine. When the uridine is mutated (but not deleted), all mutants form the loop structure, indicating that the loop structure requires an exposed fourth residue. The retrotransposition assay in HeLa cells revealed that retrotransposition requires the second guanosine, although any nucleoside functions at the fourth position, suggesting that UnaL2 reverse transcriptase specifically recognizes the 5' side of the GGANA loop. PMID- 15273328 TI - The yeast Apq12 protein affects nucleocytoplasmic mRNA transport. AB - An important step in mRNA biogenesis is the export of mRNA from the nucleus to the cytoplasm. In this work, we provide evidence that the previously uncharacterized gene APQ12 functions in nucleocytoplasmic mRNA transport in Saccharomyces cerevisiae. First, apq12delta strains manifest 3' hyperadenylated mRNA similar to other previously characterized RNA export mutants. Second, bulk poly(A)+ RNA is retained in the nucleus in apq12delta cells. Third, an Apq12p-GFP chimeric protein is localized to the nuclear periphery. Fourth, mRNA in apq12delta cells is stabilized, consistent with a defect in the rate of nuclear export. Interestingly, apq12delta mutants are severely compromised for growth and display atypical cell morphology. Because this aberrant cell morphology is not seen with other viable export mutants, Apq12p must have either an additional cellular function, or preferentially impinge on the export of mRNAs regulating cell growth. Together, these findings support a role for APQ12 in nucleocytoplasmic transport of mRNA. PMID- 15273329 TI - Severe acute respiratory syndrome: prognostic implications of chest radiographic findings in 52 patients. AB - PURPOSE: To retrospectively assess prognostic implications of radiographic findings in severe acute respiratory syndrome (SARS). MATERIALS AND METHODS: Radiographic findings were reviewed by two radiologists for 52 patients with SARS. On each radiograph, each lung was separated into upper, middle, and lower zones. A four-point scale was used to score extent of SARS-related lesions in each zone; points from all zones were added for a cumulative score. Patient sex, age, comorbidities, duration of developing lesions, lesion score for each radiograph, need for mechanical ventilation, and percentage of lung affected were compared between patients who died (n = 20) and survivors (n = 32). Continuous and categorical variables were analyzed with Mann-Whitney test and Fisher exact or chi(2) test, respectively. RESULTS: Survival and mortality groups showed no significant differences with respect to patient sex, duration of SARS-related lesions, development of lesion shifting, and acute respiratory distress syndrome. Patients who died were significantly older (mean +/- standard deviation, 56.9 years +/- 17.2 vs 40.4 years +/- 16.6; P =.002) and had higher frequency of comorbid lung illnesses (nine of 20 vs two of 32, P =.001), maximal lesion extent score of 7 or higher (20 of 20 vs five of 32, P <.001), involvement of four or more lung zones (17 of 20 vs four of 32, P <.001), bilateral lung involvement (19 of 20 vs 14 of 32, P <.001), need for mechanical ventilation (18 of 20 vs two of 32, P <.001), and higher percentage of affected areas (41.5% +/- 8.6 vs 16.4% +/- 10.0, P <.001) than those of survivors. CONCLUSION: On chest radiographs, maximal SARS-related lesion extent score of 7 or higher is a strong predictor of mortality, especially in patients with comorbid lung illnesses and involvement of four or more lung zones. PMID- 15273330 TI - Patient exposure and associated radiation risks from fluoroscopically guided vertebroplasty or kyphoplasty. AB - PURPOSE: To derive normalized data for the estimation of effective, gonadal, and peak skin doses to patients undergoing vertebroplasty or kyphoplasty and to investigate the potential for cancer induction, genetic effects, and radiation induced skin injury after such procedures. MATERIALS AND METHODS: Dose values normalized over dose-area product were determined for all radiosensitive organs and tissues by using a humanoid phantom and thermoluminescence dosimetry separately for anteroposterior and lateral projections. Measurements were obtained for treatments of the fifth, eighth, and 11th thoracic vertebrae and the first, third, and fifth lumbar vertebrae. Total fluoroscopy time and resultant dose-area product from each fluoroscopic exposure were monitored in 11 consecutive patients (seven women and four men) undergoing kyphoplasty. The age range of these patients was 41-78 years, and the mean age was 58 years. RESULTS: Mean total fluoroscopy time for kyphoplasty was 10.1 minutes +/- 2.2 (standard deviation). Mean effective dose to patients from kyphoplasty was 8.5-12.7 mSv, and mean gonadal dose was 0.04-16.4 mGy, depending on the level of the treated vertebra. Skin injuries after kyphoplasty are improbable if source-to-skin distance is 35 cm or more; however, such injuries may occur if the total fluoroscopy time per projection is extended and/or the source-to-skin distance is less than 35 cm during the procedure. CONCLUSION: Patient radiation exposure and associated risks from vertebroplasty or kyphoplasty may be considerable. Data obtained in the current study may be used to establish patient effective dose, gonadal dose, and entrance skin exposure, as well as associated risks, from these fluoroscopically guided surgical treatments of spinal disorders. PMID- 15273331 TI - Rectal cancer: local staging and assessment of lymph node involvement with endoluminal US, CT, and MR imaging--a meta-analysis. AB - PURPOSE: To perform a meta-analysis to compare endoluminal ultrasonography (US), computed tomography (CT), and magnetic resonance (MR) imaging in rectal cancer staging. MATERIALS AND METHODS: Relevant articles published between 1985 and 2002 were included if more than 20 patients were studied, histopathologic findings were the reference standard, and data were presented for 2 x 2 tables; articles were excluded if data were reported elsewhere in more detail. Two reviewers independently extracted data on study characteristics and results. Bivariate random-effects approach was used to obtain summary estimates of sensitivity and specificity for invasion of muscularis propria, perirectal tissue, and adjacent organs and for lymph node involvement. Summary receiver operating characteristic (ROC) curves were fitted for perirectal tissue invasion and lymph node involvement. RESULTS: Ninety articles fulfilled all inclusion criteria. For muscularis propria invasion, US and MR imaging had similar sensitivities; specificity of US (86% [95% confidence interval [CI]: 80, 90]) was significantly higher than that of MR imaging (69% [95% CI: 52, 82]) (P =.02). For perirectal tissue invasion, sensitivity of US (90% [95% CI: 88, 92]) was significantly higher than that of CT (79% [95% CI: 74, 84]) (P <.001) and MR imaging (82% [95% CI: 74, 87]) (P =.003); specificities were comparable. For adjacent organ invasion and lymph node involvement, estimates for US, CT, and MR imaging were comparable. Summary ROC curve for US of perirectal tissue invasion showed better diagnostic accuracy than that of CT and MR imaging. Summary ROC curves for lymph node involvement showed no differences in accuracy. CONCLUSION: For local invasion, endoluminal US was most accurate and can be helpful in screening patients for available therapeutic strategies. PMID- 15273332 TI - Stereotactic 11-gauge vacuum-assisted breast biopsy: influence of number of specimens on diagnostic accuracy. AB - PURPOSE: To determine whether number of specimens obtained at stereotactic 11 gauge vacuum-assisted breast biopsy with the patient prone influences diagnostic accuracy and to determine whether this number varies depending on mammographic appearance of lesions as masses or microcalcifications. MATERIALS AND METHODS: Biopsy was prospectively performed in 100 patients (median age, 55 years; range, 31-81 years) with 100 lesions that were mammographically evident as masses (n = 50) and microcalcifications (n = 50) with standardized protocol to acquire 20 specimens per lesion in three 360 degrees probe rotations at one skin entry site. Specimens were histologically evaluated sequentially, and findings were compared with results of surgical excision or of mammographic follow-up for at least 24 months. Differences in diagnostic yield after each probe rotation and differences in diagnostic yield between masses and microcalcifications were determined with chi(2) test. RESULTS: Up to 12 specimens harvested within two 360 degrees probe rotations were necessary to yield correct diagnosis in 96% of patients with masses and 92% of patients with microcalcifications. Diagnostic yield was not improved with more than 12 specimens for masses or microcalcifications. In two (4%) of 47 patients with lesions that were eventually diagnosed as cancer, results at stereotactic biopsy indicated they were benign. Underestimation of diagnosis of lesions as atypical ductal hyperplasia and ductal carcinoma in situ occurred in two (50%) of four and two (17%) of 12 lesions, respectively. With 20 specimens harvested during three probe rotations, there was no statistically significant difference in diagnostic yield between patients with masses and those with microcalcifications (P =.68). CONCLUSION: At 11-gauge vacuum-assisted biopsy, highest diagnostic yield was achieved with 12 specimens per lesion, independent of mammographic appearance of the lesion. Even with standardized retrieval of 20 specimens per lesion, underestimation of disease still occurs. PMID- 15273333 TI - Estimated radiation risks potentially associated with full-body CT screening. AB - PURPOSE: To estimate the radiation-related cancer mortality risks associated with single or repeated full-body computed tomographic (CT) examinations by using standard radiation risk estimation methods. MATERIALS AND METHODS: The estimated dose to the lung or stomach from a single full-body CT examination is 14-21 mGy, which corresponds to a dose region for which there is direct evidence of increased cancer mortality in atomic bomb survivors. Total doses for repeated examinations are correspondingly higher. The authors used estimated cancer risks in a U.S. population derived from atomic bomb-associated cancer mortality data, together with calculated organ doses from a full-body CT examination, to estimate the radiation risks associated with single and multiple full-body CT examinations. RESULTS: A single full-body CT examination in a 45-year-old adult would result in an estimated lifetime attributable cancer mortality risk of around 0.08%, with the 95% credibility limits being a factor of 3.2 in either direction. A 45-year-old adult who plans to undergo annual full-body CT examinations up to age 75 (30 examinations) would accrue an overall estimated lifetime attributable risk of cancer mortality of about 1.9%, with the 95% credibility limits being a factor of 2 in either direction. CONCLUSION: The authors provide estimates of lifetime cancer mortality risks from both single and annual full-body CT examinations. These risk estimates are needed to assess the utility of full-body CT examinations from both an individual and a public health perspective. PMID- 15273334 TI - Detection of recurrence in patients with rectal cancer: PET/CT after abdominoperineal or anterior resection. AB - PURPOSE: To assess diagnostic accuracy of combined positron emission tomography (PET) and computed tomography (CT) in detection of pelvic recurrence in patients with rectal cancer who underwent abdominoperineal or anterior resection. MATERIALS AND METHODS: Sixty-two patients were enrolled; 37 were men, and 25 were women. Seventeen patients underwent abdominoperineal resection and 45 underwent anterior resection with an anastomosis in the pelvic region before referral for PET/CT. Pelvic sites of fluorine 18 ((18)F) fluorodeoxyglucose (FDG) uptake were rated separately on PET and PET/CT images as benign or malignant on the basis of shape, location, and intensity of (18)F FDG uptake (1-2 = benign and/or physiologic, 3 = equivocal, 4-5 = malignant). Two readers interpreted images in consensus. Altered pelvic anatomy and presence of presacral abnormalities were assessed with CT. Pelvic recurrence was confirmed with histologic analysis or clinical and imaging follow-up. Sensitivity, specificity, positive and negative predictive values, and accuracy of PET and PET/CT in the detection of pelvic recurrence were compared with lesion- and patient-based analyses by using the chi(2) test. Clinical relevance of PET/CT assessment was determined. RESULTS: Of 81 pelvic sites with increased (18)F FDG uptake, 44 were malignant. Sensitivity, specificity, positive predictive value, negative predictive value, and accuracy for differentiating malignant from benign (18)F FDG uptake in the pelvis were 98%, 96%, 90%, 97%, and 93% for PET/CT and 82%, 65%, 73%, 75%, and 74% for PET, respectively. The most common cause for false-positive interpretation of PET findings was physiologic (18)F FDG uptake in displaced pelvic organs. Presacral CT abnormalities were present in 30 (48%) of 62 patients, and seven (23%) abnormalities were malignant. PET/CT was used to distinguish benign and malignant presacral abnormalities with a sensitivity, specificity, positive predictive value, and negative predictive value of 100%, 96%, 88%, and 100%, respectively. PET/CT findings were clinically relevant in 29 (47%) of 62 patients. CONCLUSION: PET/CT is an accurate technique in the detection of pelvic recurrence after surgical removal of rectal cancer. PMID- 15273335 TI - Non-Hodgkin lymphoma and Hodgkin disease: coregistered FDG PET and CT at staging and restaging--do we need contrast-enhanced CT? AB - PURPOSE: To retrospectively compare diagnostic value of coregistered fluorine 18 fluorodeoxyglucose positron emission tomographic (PET) and computed tomographic (CT) scans obtained with low-dose nonenhanced CT (PET/CT) with those routinely obtained with contrast material-enhanced CT for staging and restaging of disease in patients with Hodgkin disease or high-grade non-Hodgkin lymphoma. MATERIALS AND METHODS: Sixty patients (mean age, 39.6 years +/- 17.1 [standard deviation]) with Hodgkin disease (n = 42) or high-grade non-Hodgkin lymphoma (n = 18) were included in this retrospective study. All patients underwent PET/CT and contrast enhanced CT within a maximum of 24 days (mean, 9.1 days +/- 7.0) of each other for staging (n = 19) or first follow-up examination (n = 41). Findings were extracted from original written reports (PET/CT, contrast-enhanced CT) and compared with findings of reference standard, which included biopsy or follow-up with clinical, laboratory, or other imaging findings. For statistical analysis, sensitivity and specificity were calculated with findings of the reference standard. Agreement of both methods was determined with Cohen kappa and McNemar tests on a per-patient basis. RESULTS: For evaluation of lymph node involvement, sensitivity of PET/CT and contrast-enhanced CT was 94% and 88%, and specificity was 100% and 86%, respectively. For evaluation of organ involvement, sensitivity of PET/CT and contrast-enhanced CT was 88% and 50%, and specificity was 100% and 90%, respectively. Agreement of both methods was excellent (kappa = 0.84) for assignment of lymph node involvement but only fair (kappa = 0.50) for extranodal disease. A difference with P <.05 (McNemar test) was considered significant in regard to exclusion of disease with PET/CT, compared with contrast-enhanced CT. CONCLUSION: PET/CT performed with nonenhanced CT is more sensitive and specific than is contrast-enhanced CT for evaluation of lymph node and organ involvement, especially regarding exclusion of disease, in patients with Hodgkin disease and high-grade non-Hodgkin lymphoma. PMID- 15273336 TI - Intraoperative US in patients undergoing surgery for liver neoplasms: comparison with MR imaging. AB - PURPOSE: To retrospectively compare intraoperative ultrasonography (US) and preoperative magnetic resonance (MR) imaging with contrast material enhancement for the depiction of liver lesions in patients undergoing hepatic resection. MATERIALS AND METHODS: A radiologist (D.V.S.) and a surgeon (K.K.T.) retrospectively identified 79 patients (36 female and 43 male patients; age range, 10-78 years; mean age, 57 years) who had undergone surgical resection for primary liver tumor or metastasis and had also undergone preoperative contrast enhanced MR imaging within 6 weeks before surgery. MR imaging was performed with a 1.5-T system. Dedicated intraoperative US of the liver was performed or supervised by a gastrointestinal radiologist using a 7.5-MHz linear-array transducer, after adequate hepatic mobilization by the surgeon. Histopathologic evaluation of the 159 resected hepatic lesions served as the reference standard. The lesion distribution included colon cancer metastasis (n = 122), hepatocellular carcinoma (n = 23), cholangiocarcinoma (n = 6), cavernous hemangioma (n = 4), focal nodular hyperplasia (n = 2), hamartoma (n = 1), and metastatic embryonal sarcoma (n = 1). RESULTS: Of 159 lesions, 138 (86.7%) were identified at both MR imaging and intraoperative US. Twelve additional lesions (7.5%) in 10 patients were detected only at intraoperative US (eight metastases, one hepatocellular carcinoma, one cholangiocarcinoma, one hemangioma, and one biliary hamartoma). Both modalities failed to depict nine lesions (5.6%) (four metastases, four hepatocellular carcinomas, and one cholangiocarcinoma). The sensitivities of MR imaging and intraoperative US for liver lesion depiction were 86.7% and 94.3%, respectively. Surgical management was altered on the basis of the intraoperative US findings in only three of 10 patients (4%). CONCLUSION: Contrast-enhanced MR imaging is as sensitive as intraoperative US in depicting liver lesions before hepatic resection. PMID- 15273337 TI - Prostate cancer: gadolinium-enhanced MR imaging at 3 weeks compared with needle biopsy at 6 months after cryoablation. AB - PURPOSE: To determine if nonenhancing tissue on gadolinium-enhanced magnetic resonance (MR) images obtained 3 weeks after cryoablation of the prostate helps reliably and accurately predict nonviable cryoablated tissue at 6-month biopsy. MATERIALS AND METHODS: Fifty-four consecutive patients with prostate cancer who underwent cryoablation were followed up prospectively. Fifty-one underwent gadolinium-enhanced MR imaging at 3 weeks (three had gadolinium allergy); 49, biopsy at 6 months (three refused and two had other primary malignancies); and all, prostate-specific antigen (PSA) tests at 6 weeks, 3 months, and every 3 months thereafter. MR images were evaluated and scored according to the degree of signal void and were correlated with the 6-month biopsy reports and, to a lesser degree, PSA levels. The biopsy reports were examined for the presence or absence of cancerous tissue, viable tissue, and nonviable tissue. A one-way analysis of variance was used for statistical and regression analyses. RESULTS: The correlation of MR imaging scores with PSA levels and MR imaging scores with biopsy findings resulted in P values of.337 and.780, respectively. A slight statistically significant trend existed for the relation of biopsy results with PSA levels, with a P value of.041, which was expected. CONCLUSION: Findings of postoperative gadolinium-enhanced MR imaging are not predictive of 6-month biopsy results or follow-up PSA levels. PMID- 15273338 TI - Effect of monitor luminance and ambient light on observer performance in soft copy reading of digital chest radiographs. AB - PURPOSE: To examine the combined effects of monitor luminance and ambient light on observer performance for detecting abnormalities in a soft-copy interpretation of digital chest radiographs. MATERIALS AND METHODS: A total of 254 digital chest radiographs were displayed on a high-resolution cathode ray tube monitor at three luminance levels (25, 50, and 100 foot-lamberts) under three ambient light levels (0, 50, and 460 lux). Six chest radiologists reviewed each image in nine modes of combined luminance and ambient light. The observers were allowed to adjust the window width and level of the soft-copy images. The abnormalities included nodule, pneumothorax, and interstitial disease. Observer performance was analyzed in terms of the receiver operating characteristics. The observers reported their subjective level of visual fatigue with each viewing mode. A statistical test was conducted for each of the abnormalities and for fatigue score by using repeated measures two-way analysis of variance with an interaction. RESULTS: The detection of nodules was the only reading that was affected by the ambient light with a statistically significant difference (P <.05). Otherwise, observer performance for detecting a nodule, pneumothorax, and interstitial disease was not significantly different in the nine-mode comparison. There was no evidence that the luminance of the monitors was related to the ambient light for any of the abnormalities. The fatigue score showed a statistically significant difference due to both the luminance and ambient light. CONCLUSION: When adequate window width and level are applied to soft-copy images, the primary diagnosis with chest radiographs on the monitor is unlikely to be affected under low ambient light and a monitor luminance of 25 foot-lamberts or more. PMID- 15273339 TI - Percutaneous vertebroplasty for malignant compression fractures with epidural involvement. AB - PURPOSE: To evaluate safety and effectiveness of performance of percutaneous vertebroplasty (PVP) in patients with malignant compression fractures and involvement of the epidural space. MATERIALS AND METHODS: PVP was performed in 50 patients with metastatic disease or multiple myeloma between June 1998 and April 2002. Twenty-five women (mean age, 62.3 years; range, 38-85 years) and 25 men (mean age, 63.1 years; range, 37-92 years) were included. Cases were retrospectively reviewed. Patients who had undergone cross-sectional imaging were classified into three groups. First group had no epidural involvement; second group, mild epidural involvement without contact with spinal cord or nerve roots; third group, moderate involvement and contact with spinal cord or nerve roots. Procedural safety was evaluated with review of all post-PVP complications and their treatment. Effectiveness was evaluated with follow-up phone calls for assessment of change in pain level and activity after PVP. Follow-up calls were performed at 1 day; 2 weeks; 1, 3, and 6 months; and 1 and 2 years. Differences between groups were assessed with singly ordered Kruskal-Wallis test. RESULTS: Fourteen patients were classified in the first group, 18 in the second, and 18 in the third. There were no significant differences in pain or mobility outcomes among groups. At the last follow-up call, 41 (82%) of 50 patients reported improvement in pre-PVP pain. Six (12%) reported no change, and three (6%) reported increased pain. After PVP in 26 (52%) patients, there was a period of increased mobility; in 19 (38%), no improvement in mobility occurred; and in five (10%), decreased mobility was reported. Complications included acute increased pain or new areas of pain in seven (14%) patients. None of these required surgery; four were treated with nerve root block; two, with central epidural injection; and one, with overnight intravenous steroids. CONCLUSION: PVP can be performed safely and effectively with conscious sedation in patients with malignant compression fractures and epidural involvement. PMID- 15273340 TI - Blood flow in coronary artery bypass vein grafts: volume versus velocity at cardiovascular MR imaging. AB - Forty-nine patients with previous bypass surgery underwent coronary angiography and cardiovascular magnetic resonance (MR) imaging of single-vein bypass grafts. Volume flow and velocity analyses were performed and compared on MR velocity maps. Bland-Altman analysis showed close agreement between the two types of analysis. Comparison of areas under the receiver operating characteristic curve revealed no significant differences between the analyses for detection of stenoses of 70% or greater. Diagnostic accuracy for volume flow and velocity parameters was 92% and 93%, respectively. Velocity analysis appears to be the preferred method, because it is less time-consuming and has a similar diagnostic accuracy to volume flow analysis. PMID- 15273341 TI - Low-voltage digital selenium radiography: detection of simulated interstitial lung disease, nodules, and catheters--a phantom study. AB - PURPOSE: To compare three tube voltages in digital selenium radiography for the detection of simulated interstitial lung disease, nodules, and catheters. MATERIALS AND METHODS: Simulated catheters, nodules, and ground-glass, linear, miliary, and reticular patterns were superimposed over an anthropomorphic chest phantom. Digital selenium radiography was performed with different tube voltages (70, 90, and 150 kVp). Hard-copy images were generated. Detection performance of five radiologists was compared by using receiver operating characteristic (ROC) analysis involving 54,000 observations. RESULTS: The detection of ground-glass, linear, miliary, and reticular patterns over lucent lung and of nodules equal to, smaller than, and larger than 10 mm increased when 70 kVp and/or 90 kVp was used. However, only the reticular pattern was significantly better detected at lower peak voltage (P <.05). Simulated catheters and nodules over the mediastinum showed smaller areas under the ROC curve at lower peak voltage. These results were not statistically significant (P >.05). CONCLUSION: The diagnostic performance of digital selenium radiography at lower peak voltage is at least as good as that at higher peak voltage for interstitial lung disease over lucent lung. Performance is equivalent for nodules and catheters over obscured chest regions at lower peak voltages compared with that at 150 kVp. Our results implicate that the use of high-voltage technique in digital selenium radiography should be reassessed. PMID- 15273344 TI - Recent advances in dosimetry using the optically stimulated luminescence of Al2O3:C. AB - This paper presents an overview of some very recent developments in optically stimulated luminescence (OSL) dosimetry using aluminium oxide (Al(2)O(3):C), with special emphasis given to the work of the research group at Oklahoma State University. Some of the advances are: (i) the development of a real-time optical fibre system for in vivo dosimetry applied to radiotherapy; (ii) the development of a fibre dosimetry system for remote detection of radiological contaminants in soil; (iii) the characterisation of Al(2)O(3):C in heavy charged particle fields and the study of ionisation density dependence of the OSL from Al(2)O(3):C; and (iv) fast and separate assessment of beta and gamma components of the natural dose rate in natural sediments. These achievements highlight the versatility of the OSL technique associated with the high-sensitivity of Al(2)O(3):C for the development of new dosimetry applications. PMID- 15273345 TI - Standards for standard-makers? A testing time. AB - The International Commission on Radiological Protection (ICRP) is now reviewing its recommendations with a view to publishing their revision in 2005. The last set of recommendations issued by the ICRP has caused some concern to neutron dosimetrists. This paper attempts to explain these concerns. Technological developments make it likely that exposure to high-linear energy transfer (LET) radiations will increase in the future. It is in the area of the dosimetry of high-LET radiations, particularly neutrons, where some experts feel that ICRP recommendations have been unclear. This paper discusses the process of setting protection limits in toxicology and its application to radiation protection. The development of radiation protection quantities and models is described, and the problems found with effective dose described. Suggestions for improvements are made that would enable effective dose to serve in two modes--both as a limiting quantity and also as the measurable (operational) quantity required by dosimetrists. PMID- 15273346 TI - Spectra of scattered photons in large absorbers and their importance for the values of radiation weighting factor wR. AB - In its review of the present values of radiation weighting factor w(R) and of possible revisions of this factor, the German Radiation Protection Commission has recommended to maintain the approach of ICRP 60 to base the selection of the w(R) value for a given radiation (e.g. fission neutrons) on observed values of the relative biological effectiveness (RBE) of this radiation 'regardless of whether the reference radiation is X rays or gamma rays'. The physical background of the German recommendation is the buildup of a strong field of energy-degraded Compton scattered photons in the human body if exposed to an external field of high energy photons, so that the total radiation field inside the body is a mixture comprising low and high photon energies. Therefore, it is appropriate that the selection of the w(R) value of the given radiation is guided by RBE values averaged over X rays and gamma rays as the reference radiations. In support of this rationale, the present paper provides a sample of Monte Carlo calculated scattered photon spectra in large absorbers exposed to high-energy photons. Depth dependent fractional dose contributions of the scattered photons are tabulated for incident energies from 1 to 10 MeV, and estimates of the influence of their degraded energies on the biological effectiveness of the incoming radiation are presented. Accordingly, we point out that it is appropriate to use, for the purposes of 'risk projection', RBE values averaged over X and gamma reference radiations. PMID- 15273347 TI - Why it is advisable to keep wR = 1 and Q = 1 for photons and electrons. AB - It has been well known for a long time that the biological effectiveness of photons and electrons depends on the mean linear energy transfer (LET) of the radiation considered, e.g. (60)Co gamma rays are less effective than soft or hard X rays. Nevertheless, the protection and operational dose quantities applied in radiological protection include weighting factors, w(R) or Q, respectively, which were set to 1 for all low-LET radiations. Lack of precise information, simplicity and general practical considerations are the main arguments for this convention. However, a more detailed discussion on the practical aspects supporting this procedure is missing. The paper discusses in more detail some of these aspects regarding internal and external exposure situations, which may support the idea of continuing to use w(R) = 1 for photons and electrons and, correspondingly, using Q(L) = 1 for L < 10 keV microm(-1). PMID- 15273348 TI - The impact of ICRP publication 92 on the conversion coefficients in use for cosmic ray dosimetry. AB - ICRP Publication 92 presents a proposal to achieve coherence between radiation weighting factors and quality factors. In particular, the radiation weighting factors for incident protons and neutrons have been revised and new values have been proposed. On the basis of the proposed values, sets of conversion coefficients fluence-to-effective dose for protons and neutrons have been derived for the irradiation geometries of interest for cosmic ray dosimetry. PMID- 15273349 TI - Adaptation of the present concept of dosimetric radiation protection quantities for external radiation to radiation protection practice. AB - The present concept of dosimetric radiation protection quantities for external radiation is reviewed. For everyday application of the concept some adaptations are recommended. The check of the compliance with dose limits should be performed either by the comparison with values of the respective operational quantities directly or by the calculation of the protection quantity by means of the operational quantity, the appertaining conversion coefficient and additional information of the radiation field. Only four operational quantities are regarded to be sufficient for most applications in radiation protection practice. The term equivalent should be used in the connection dose equivalent only. Proposals are made for names of frequently used operational quantities which are denoted up to now by symbols only. PMID- 15273350 TI - Calibration measurements and standards for radiation protection dosimetry. AB - The safe use of ionising radiation for applications in medicine, electric power production and industrial processes requires accurate measurements that are traceable to national standards. Radiological calibration laboratories provide measurements that may be used to determine the calibration coefficients for personal dosemeters and survey meters. The wide range of ionising radiation applications results in the need for a wide range of reference radiation types and intensities to be available in the calibration laboratory. The methods used and the problems encountered while developing reference radiations are discussed. PMID- 15273351 TI - The skin in radiological protection--recent advances and residual unresolved issues. AB - Exposure of the skin is important in radiological protection because, as the most superficial organ of the body, it can often receive the highest absorbed dose from an external exposure. It also has the highest radiation-induced cancer incidence risk factor of any organ (although mortality is very low). The ICRP and NCRP have, particularly over the past 15 y, been able to set dose limits for the exposure of the skin on the basis of an extensive body of radiobiological, clinical and epidemiological data. Some of the main advances in skin dose limitation in radiological protection and some of the remaining unresolved issues are reviewed. PMID- 15273352 TI - The effects of ionisation density on the thermoluminescence response (efficiency) of LiF:Mg,Ti and LiF:Mg,Cu,P. AB - In this paper, the various models dealing with the effects of ionisation density on the thermoluminescence (TL) response (efficiency) of TL LiF dosemeters are discussed. These include (i) the Unified Interaction Model (UNIM), which models photon/electron linear/supralinear dose response; (ii) the Extended Track Interaction Model (ETIM), which models heavy charged particle (HCP) TL fluence response; (iii) Modified Track Structure Theory (MTST), which models relative HCP TL efficiencies; and (iv) Microdosimetric Target Theory (MTT), which models both relative HCP efficiencies and photon energy response. PMID- 15273353 TI - Radiation protection aspects of the cosmic radiation exposure of aircraft crew. AB - Aircraft crew and frequent flyers are exposed to elevated levels of cosmic radiation of galactic and solar origin and secondary radiation produced in the atmosphere, the aircraft structure and its contents. Following recommendations of the International Commission on Radiological Protection in Publication 60, the European Union introduced a revised Basic Safety Standards Directive, which included exposure to natural sources of ionising radiation, including cosmic radiation, as occupational exposure. The revised Directive has been incorporated into laws and regulations in the European Union Member States. Where the assessment of the occupational exposure of aircraft crew is necessary, the preferred approach to monitoring is by the recording of staff flying times and calculated route doses. Route doses are to be validated by measurements. This paper gives the general background, and considers the radiation protection aspects of the cosmic radiation exposure of aircraft crew, with the focus on the situation in Europe. PMID- 15273354 TI - The relative transit time of high-rigidity and low-rigidity cosmic rays through the solar system. AB - A study of neutron monitor count rates with vertical cut-off rigidities from 0.6 to 13 GV indicates that high-energy and low-energy galactic cosmic rays arrive at earth orbit with small time differences compared to a month. This is in contradiction to the theory of the deceleration potential which states that cosmic-ray particles with rigidities greater than 36 V arrive at earth orbit about three months before cosmic-ray particles with lesses rigidines. PMID- 15273355 TI - Detectors/Dosemeters of galactic and solar cosmic rays. AB - Different passive multidetector stacks have been developed at the Italian National Agency for Environmental Protection (ANPA-stack), which makes it possible to measure directly ionising radiations, low-energy and high-energy neutrons, and high-energy charged (HZE) particles. The stack consists of several types of passive devices, namely recoil-track and fission-track detectors, bubble detectors, thermoluminescence dosemeters and an electronic personal dosemeter. Most of these detectors have been used on earth for the assessment of the occupational exposure, or in outer space for cosmic ray physics and/or for the assessment of the dose received by astronauts. A great deal of efforts and new developments have been required to make these detectors useful for in-flight measurements. As outcome of these extensive efforts, different new detectors have been developed, which exploit some of the most successful principles of radiation detection, such as the use of avalanche processes to facilitate the registration of nuclear tracks and the use of coincidence-counting to increase the signal-to noise ratio. On the basis of these new detectors, different systems (generally referred to as ANPA-stack) have been obtained, which have been successfully applied for a variety of different measurements of cosmic ray radiation fields and doses. PMID- 15273356 TI - Track-etched detectors for the dosimetry of the radiation of cosmic origin. AB - Cosmic rays contribute to the exposure on the Earth's surface as well as in its surroundings. At the surface and/or at aviation altitudes, there are mostly secondary particles created through the cosmic rays interaction in the atmosphere, which contribute to this type of exposure. Onboard a spacecraft, the exposure comes mostly from primary cosmic rays. Track-etched detectors (TED) are able to characterise both these types of exposure. The contribution of neutrons, of cosmic origin, on the Earth's surface was studied at altitudes from few hundreds to 3000 m using TED in a moderator sphere. The results obtained are compared with other data on this type of natural radiation background. The results of studies performed onboard aircraft and/or spacecraft are presented afterwards. We used TED-based neutron dosemeter, as well as a spectrometer of linear energy transfer based on a chemically etched TED. The results of studies performed onboard aircraft, as well as spacecraft, are presented and discussed, including an attempt to estimate a neutron component onboard the spacecraft. It was found that they correlate with the results of other independent investigations. PMID- 15273357 TI - The development of protection standards for intakes of radionuclides (1955-2005). AB - The last 50 y has seen a dramatic increase in understanding of the behaviour of radionuclides in the body as well as of their effects. This has resulted in substantial improvements to the way that protection standards are set. This paper aims to chart the development of radiological protection as it applies to intakes of radionuclides by those who are occupationally exposed and by members of the public. It is not concerned with medical uses of radionuclides. PMID- 15273358 TI - A review of contributions of human tissue studies to biokinetics, bioeffects and dosimetry of plutonium in man. AB - This paper briefly reviews the contributions made by human tissue studies to improved understanding of the biokinetics, dosimetry and potential bioeffects of plutonium in man. It includes consideration of tissue donations from both environmental and occupational populations, along with a brief history of human experience with plutonium and consideration of the bioethical aspects of post mortem human tissue sampling. PMID- 15273359 TI - Radiation protection dosimetry for diagnostic radiology patients. AB - The radiation protection of patients undergoing medical X-ray examinations is governed by the principles of justification and optimisation. Radiation dosimetry is required to inform medical practitioners of the levels of exposure and hence the risks from the diagnostic procedures that they have to justify and to assist the operators of X-ray imaging equipment to determine whether their procedures are optimised. This paper describes the main dosimetric methods that have been developed to meet these requirements. Suitable radiation risk projection models are used to predict the risks to patients in the UK from computed tomography examinations, as a function of age at exposure and sex, and show that the lifetime risk of fatal cancer can reach 1 in 1000 for children. The concept of 'diagnostic reference levels' as an aid to the optimisation of medical exposures is described, and progress in implementing them in the UK is reported. PMID- 15273360 TI - The history of radon from a Swedish perspective. AB - Beginning in the 16th century, what was later found to be radon was thought to be causing sickness among miners. During the first decades of the 20th century, exposure to radon was seen as being healthy. During the 1940s, 1950s and 1960s researchers thought that the gamma radiation in residences could produce genetic damage. It was not until approximately 1970 that a quantitative risk estimate for lung cancer could be calculated for miners, and not until the 1990s that a risk estimate could be established based on epidemiological studies on radon in dwellings and lung cancer. PMID- 15273361 TI - Power reactor health physics in the 1960s. AB - This note is a personal re-collection of some of the problems and experiences of the Health Physics Department staff at Dungeness Power Station during the period 1963-1971. This was a period of construction, commissioning, operating and learning. It was an interesting period for power reactor health physicists as appropriate instrumentation; information and experience were all in fairly short supply. PMID- 15273363 TI - The old file-drawer problem. PMID- 15273364 TI - Scientific publishing. U.K. lawmakers urge prompt access to published papers. PMID- 15273366 TI - Physics. Energy curve confirms paired-up Fermi condensate. PMID- 15273365 TI - Scientific publishing. Congress puts similar heat on NIH. PMID- 15273367 TI - Glycobiology. Synthetic vaccine is a sweet victory for Cuban science. PMID- 15273368 TI - Stem cell research. Advocates keep pot boiling as Bush plans new centers. PMID- 15273369 TI - Research management. Security, safety probes shut down Los Alamos national lab. PMID- 15273370 TI - Scientific meetings. NIH scientists in a spin over foreign travel. PMID- 15273371 TI - Space astronomy. U.S. academy panel urges NASA to upgrade Hubble telescope. PMID- 15273372 TI - Theoretical physics. Physics enters the twilight zone. PMID- 15273373 TI - Space science. NASA reining in PI-led planetary missions. PMID- 15273374 TI - Psychopharmacology. Volatile chemistry: children and antidepressants. PMID- 15273375 TI - HIV/AIDS. International AIDS meeting finds global commitment lacking. PMID- 15273376 TI - The health benefits of eating salmon. PMID- 15273377 TI - Risk-benefit analysis of eating farmed salmon. PMID- 15273378 TI - Cancer risk and salmon intake. PMID- 15273379 TI - Contaminant levels in farmed salmon. PMID- 15273380 TI - Keeping drugs at the proper temperature. PMID- 15273381 TI - Medicine. Magic bullet gone astray: medications and the Internet. PMID- 15273382 TI - Neuroscience. Epileptic neurons go wireless. PMID- 15273383 TI - Physics. Charging atoms, one by one. PMID- 15273384 TI - Paleoecology. The rise and fall of forests. PMID- 15273385 TI - Applied physics. Designing optimal micromixers. PMID- 15273386 TI - Public information: from nosy neighbors to cultural evolution. AB - Psychologists, economists, and advertising moguls have long known that human decision-making is strongly influenced by the behavior of others. A rapidly accumulating body of evidence suggests that the same is true in animals. Individuals can use information arising from cues inadvertently produced by the behavior of other individuals with similar requirements. Many of these cues provide public information about the quality of alternatives. The use of public information is taxonomically widespread and can enhance fitness. Public information can lead to cultural evolution, which we suggest may then affect biological evolution. PMID- 15273387 TI - Comet or asteroid shower in the late Eocene? AB - The passage of a comet shower approximately 35 million years ago is generally advocated to explain the coincidence during Earth's late Eocene of an unusually high flux of interplanetary dust particles and the formation of the two largest craters in the Cenozoic, Popigai and the Chesapeake Bay. However, new platinum group element analyses indicate that Popigai was formed by the impact of an L chondrite meteorite. Such an asteroidal projectile is difficult to reconcile with a cometary origin. Perhaps instead the higher delivery rate of extraterrestrial matter, dust, and large objects was caused by a major collision in the asteroid belt. PMID- 15273388 TI - Controlling the charge state of individual gold adatoms. AB - The nature and control of individual metal atoms on insulators are of great importance in emerging atomic-scale technologies. Individual gold atoms on an ultrathin insulating sodium chloride film supported by a copper surface exhibit two different charge states, which are stabilized by the large ionic polarizability of the film. The charge state and associated physical and chemical properties such as diffusion can be controlled by adding or removing a single electron to or from the adatom with a scanning tunneling microscope tip. The simple physical mechanism behind the charge bistability in this case suggests that this is a common phenomenon for adsorbates on polar insulating films. PMID- 15273389 TI - Rapid late Pleistocene incision of Atlantic passive-margin river gorges. AB - The direct and secondary effects of rapidly changing climate caused large rivers draining the Atlantic passive margin to incise quickly into bedrock beginning about 35,000 years ago. Measured in samples from bedrock fluvial terraces, 10 beryllium shows that both the Susquehanna and Potomac Rivers incised 10- to 20 meter-deep gorges along steep, convex lower reaches during the last glacial cycle. This short-lived pulse of unusually rapid down-cutting ended by 13,000 to 14,000 years ago. The timing and rate of downcutting are similar on the glaciated Susquehanna and unglaciated Potomac Rivers, indicating that regional changes, not simply glacial meltwater, initiated incision. PMID- 15273390 TI - GRACE measurements of mass variability in the Earth system. AB - Monthly gravity field estimates made by the twin Gravity Recovery and Climate Experiment (GRACE) satellites have a geoid height accuracy of 2 to 3 millimeters at a spatial resolution as small as 400 kilometers. The annual cycle in the geoid variations, up to 10 millimeters in some regions, peaked predominantly in the spring and fall seasons. Geoid variations observed over South America that can be largely attributed to surface water and groundwater changes show a clear separation between the large Amazon watershed and the smaller watersheds to the north. Such observations will help hydrologists to connect processes at traditional length scales (tens of kilometers or less) to those at regional and global scales. PMID- 15273391 TI - Large perturbations of the carbon cycle during recovery from the end-permian extinction. AB - High-resolution carbon isotope measurements of multiple stratigraphic sections in south China demonstrate that the pronounced carbon isotopic excursion at the Permian-Triassic boundary was not an isolated event but the first in a series of large fluctuations that continued throughout the Early Triassic before ending abruptly early in the Middle Triassic. The unusual behavior of the carbon cycle coincides with the delayed recovery from end-Permian extinction recorded by fossils, suggesting a direct relationship between Earth system function and biological rediversification in the aftermath of Earth's most devastating mass extinction. PMID- 15273392 TI - Shifts in deep-sea community structure linked to climate and food supply. AB - A major change in the community structure of the dominant epibenthic megafauna was observed at 4100 meters depth in the northeast Pacific and was synchronous to a major El Nino/La Nina event that occurred between 1997 and 1999. Photographic abundance estimates of epibenthic megafauna from 1989 to 2002 show that two taxa decreased in abundance after 1998 by 2 to 3 orders of magnitude, whereas several other species increased in abundance by 1 to 2 orders of magnitude. These faunal changes are correlated to climate fluctuations dominated by El Nino/La Nina. Megafauna even in remote marine areas appear to be affected by contemporary climatic fluctuations. Such faunal changes highlight the importance of an adequate temporal perspective in describing biodiversity, ecology, and anthropogenic impacts in deep-sea communities. PMID- 15273393 TI - Phosphorylation by cyclin B-Cdk underlies release of mitotic exit activator Cdc14 from the nucleolus. AB - Budding yeast protein phosphatase Cdc14 is sequestered in the nucleolus in an inactive state during interphase by the anchor protein Net1. Upon entry into anaphase, the Cdc14 early anaphase release (FEAR) network initiates dispersal of active Cdc14 throughout the cell. We report that the FEARnetwork promotes phosphorylation of Net1 by cyclin-dependent kinase (Cdk) complexed with cyclin B1 or cyclin B2. These phosphorylations appear to be required for FEAR and sustain the proper timing of late mitotic events. Thus, a regulatory circuit exists to ensure that the arbiter of the mitotic state, Cdk, sets in motion events that culminate in exit from mitosis. PMID- 15273394 TI - Functional adaptation of BabA, the H. pylori ABO blood group antigen binding adhesin. AB - Adherence by Helicobacter pylori increases the risk of gastric disease. Here, we report that more than 95% of strains that bind fucosylated blood group antigen bind A, B, and O antigens (generalists), whereas 60% of adherent South American Amerindian strains bind blood group O antigens best (specialists). This specialization coincides with the unique predominance of blood group O in these Amerindians. Strains differed about 1500-fold in binding affinities, and diversifying selection was evident in babA sequences. We propose that cycles of selection for increased and decreased bacterial adherence contribute to babA diversity and that these cycles have led to gradual replacement of generalist binding by specialist binding in blood group O-dominant human populations. PMID- 15273395 TI - A synthetic conjugate polysaccharide vaccine against Haemophilus influenzae type b. AB - Glycoconjugate vaccines provide effective prophylaxis against bacterial infections. To date, however, no commercial vaccine has been available in which the key carbohydrate antigens are produced synthetically. We describe the large scale synthesis, pharmaceutical development, and clinical evaluation of a conjugate vaccine composed of a synthetic capsular polysaccharide antigen of Haemophilus influenzae type b (Hib). The vaccine was evaluated in clinical trials in Cuba and showed long-term protective antibody titers that compared favorably to licensed products prepared with the Hib polysaccharide extracted from bacteria. This demonstrates that access to synthetic complex carbohydrate-based vaccines is feasible and provides a basis for further development of similar approaches for other human pathogens. PMID- 15273396 TI - Large-scale copy number polymorphism in the human genome. AB - The extent to which large duplications and deletions contribute to human genetic variation and diversity is unknown. Here, we show that large-scale copy number polymorphisms (CNPs) (about 100 kilobases and greater) contribute substantially to genomic variation between normal humans. Representational oligonucleotide microarray analysis of 20 individuals revealed a total of 221 copy number differences representing 76 unique CNPs. On average, individuals differed by 11 CNPs, and the average length of a CNP interval was 465 kilobases. We observed copy number variation of 70 different genes within CNP intervals, including genes involved in neurological function, regulation of cell growth, regulation of metabolism, and several genes known to be associated with disease. PMID- 15273397 TI - Acquired dendritic channelopathy in temporal lobe epilepsy. AB - Inherited channelopathies are at the origin of many neurological disorders. Here we report a form of channelopathy that is acquired in experimental temporal lobe epilepsy (TLE), the most common form of epilepsy in adults. The excitability of CA1 pyramidal neuron dendrites was increased in TLE because of decreased availability of A-type potassium ion channels due to transcriptional (loss of channels) and posttranslational (increased channel phosphorylation by extracellular signal-regulated kinase) mechanisms. Kinase inhibition partly reversed dendritic excitability to control levels. Such acquired channelopathy is likely to amplify neuronal activity and may contribute to the initiation and/or propagation of seizures in TLE. PMID- 15273399 TI - Methods of epigenetic analysis. AB - Epigenetics encompasses heritable changes in DNA or its associated proteins except mutations in gene sequence. Many investigators in the field of epigenetics focus on histone modifications and DNA methylation, two molecular mechanisms that are often linked and interdependent. A variety of methods are applied to the study of epigenetic processes, and the past decade has witnessed an exponential increase in novel approaches to elucidate the molecular mysteries of epigenetic inheritance. This chapter summarizes some of the most contemporary methods used to study epigenetics presented throughout the book. PMID- 15273400 TI - Chromatin immunoprecipitation assays. AB - Chromatin immunoprecipitation (ChIP) is a powerful tool to study protein-DNA interaction and is widely used in many fields to study proteins associated with chromatin, such as histone and its isoforms and transcription factors, across a defined DNA domain. Here, we show the step-by-step methods currently used in our lab to immunoprecipitate the formaldehyde crosslinked chromatin and further analyze the immuprecipitated DNA by semiquantitative PCR. PMID- 15273401 TI - Native chromatin immunoprecipitation. AB - Chromatin immunoprecipitation (ChIP) is a technique widely used for determining the genomic location of modified histones and other chromatin-associated factors. Here we describe the methodology we have used in our laboratory for the immunoprecipitation of chromatin isolated from cells in the absence of crosslinking. Chromatin released from nuclei by micrococcal nuclease digestion is centrifuged through sucrose gradients to allow selection of mono- or dinucleosomes. This allows a protein or modification at a particular gene or locus to be mapped at higher resolution than in a crosslinked ChIP experiment. Two methods for the immunoprecipitation of chromatin are described: a large-scale fractionation by which it is possible to visualize the proteins of the immunoprecipitate by polyacrylamide gel electrophoresis, PAGE and a small-scale method that is more appropriate when the quantity of chromatin is limited. The sequence content of DNA extracted from the immunoprecipitated chromatin is analyzed by hybridization of Southern or slot blots, or by quantitative polymerase chain reaction. Enrichment of particular sequences in the immunoprecipitated fraction reveals the presence and extent of the modification at this location. PMID- 15273402 TI - Q-PCR in combination with ChIP assays to detect changes in chromatin acetylation. AB - Quantitative polymerase chain reaction (Q-PCR) allows for the accurate and reproducible determination of the amount of target DNA in a sample through the measurement of PCR product accumulation in "real time." This method determines starting target DNA quantity over a large assay dynamic range and requires no post-PCR sample manipulation. When used in combination with the method of chromatin immunoprecipitation (ChIP), the amount of protein binding to a specific region of DNA can be accurately and rapidly determined. A method for quantifying the presence of acetylated histones H3 and H4 on different regions of a target locus using Q-PCR after ChIP is described. PMID- 15273403 TI - Restriction endonuclease accessibility as a determinant of altered chromatin structure. AB - Active genes in eukaryotic genomes are typically found in open, nuclease sensitive regions of chromatin. This chapter presents an overview of the techniques used to assay restriction endonuclease cleavage of chromosomal DNA as an approach to assess general accessibility at a genomic region of interest. We describe protocols to (1) prepare nuclei templates, (2) treat chromosomal DNA with a restriction enzyme(s), and (3) visualize and quantify chromosomal cleavage(s), with an emphasis on ligation-mediated (LM) PCR techniques. PMID- 15273404 TI - Measuring changes in chromatin using micrococcal nuclease. AB - This chapter documents a simple protocol to identify the nucleosome positioning of any given genes. The procedure includes partitioning 200-bp DNA fragments constituting the nucleosomal core region by micrococcal nuclease digestion and semiquantitative polymerase chain reaction amplification using multiple sets of primers covering arbitrary regions of approximately 150 bp in the gene. If the nuclease-digested 200-bp DNA is efficiently amplified, the region is inside the core. If the amplification is poor, the region spans the linker region. By a combination of this method with direct methylation mapping, the core region of a maize gene, ZmMI1, was shown to be less methylated than the linker region. The potential usefulness of the technique is discussed. PMID- 15273405 TI - DNaseI hypersensitivity analysis of chromatin structure. AB - Transcriptionally inactive DNA is packaged into condensed chromatin such that it is unavailable to the transcription initiation complex. Activation of the silenced genes during processes such as differentiation first requires that the chromatin structure be remodeled into a transcriptionally permissive configuration, with the DNA "exposed" and accessible to transcription factors. The change in chromatin structure associated with transcriptional competence can be detected as increased sensitivity of the exposed DNA to digestion with DNaseI. This increased susceptibility is referred to as DnaseI-hypersensitivity. DNaseI hypersensitive sites are often located in the recognition sites for transcription factors, including promoters and enhancers. This chapter describes the protocols necessary to perform and analyze DNaseI hypersensitivity assays, a technique becoming increasingly important given the rapid advances in our understanding of the chromatin remodeling processes. PMID- 15273406 TI - Inhibition of histone deacetylases. AB - Reversible histone acetylation, governed dynamically by histone acetyltransferases (HATs) and histone deacetylases (HDACs), plays a pivotal role in regulation of gene expression through remodeling chromatin structure. Manipulation of the equilibrium between acetylation and deacetylation of histones by specific HDAC inhibitors is thus a useful tool to study functional role(s) for histone hyper-/hypoacetylation in controlling gene transcription and many other cellular activities. By using the trans-activating effect of trichostatin A (TSA), a widely used HDAC inhibitor, on the telomerase reverse transcriptase (hTERT) gene as an example, we summarize various aspects of HDAC inhibitors and provide a general strategy for their in vitro application in studies of gene regulation. PMID- 15273407 TI - Site-specific analysis of histone methylation and acetylation. AB - Covalent modifications on the nucleosomal histones are essential in chromatin regulation and gene expression. Patterns of histone modifications may be somatically maintained and can thereby maintain locus-specific repression/activity in defined lineages or throughout development. During recent years, histone acetylation and methylation have emerged as key players in the repression or activation of genes and chromosomal domains. Histone methylation and acetylation patterns (and other histone modifications) can be analyzed by chromatin immunoprecipitation (ChIP). This chapter describes how ChIP can be performed on native chromatin prepared from cells and tissues, in order to analyze histone methylation and acetylation at specific sites in the genome. We also present different PCR-based assays that can be applied to analyze loci of interest in immunoprecipitated chromatin fractions. PMID- 15273408 TI - Analysis of mammalian telomere position effect. AB - Methods relating to the positioning of a transgene next to a newly formed telomere in human (HeLa) cells and the subsequent analysis of the resulting clones are described. These include vector design, analysis of integration sites by Southern blotting, pharmacological relief of silencing, and enhancement of silencing by telomere elongation. Several potential pitfalls of applying these techniques to other cell lines are discussed. In addition, detailed instructions are provided for several more general methods related to human telomeres including terminal restriction fragment analysis and purification of telomeres from digested genomic DNA. This chapter summarizes the techniques currently in use that relate to human telomere position effect. PMID- 15273409 TI - Activity assays for poly-ADP ribose polymerase. AB - Poly(ADP-ribose) polymerase (PARP-1) is a nuclear enzyme that has traditionally been thought to require discontinuous or "damaged" DNA (dcDNA) as a coenzyme, a preconception that has limited research mainly to its role in cell pathology, i.e., DNA repair and apoptosis. Recent evidence has shown that this enzyme is broadly involved in normal cell physiological functions including chromatin modeling and gene regulation when DNA strand breaks are absent. We have recently shown that double-stranded DNA (dsDNA) serves as a more efficient coenzyme for PARP-1 than dcDNA, providing a mechanistic basis for PARP-1 function in normal cell physiology. Here we provide a detailed outline of methods for analyzing PARP 1 enzymatic activity using dsDNA as a coenzyme compared with broken or damaged DNA. Two procedures are described, one for analysis of auto-, and the other for trans-ADP-ribosylation. These assays provide a means of investigating the physiological role(s) of PARP-1 in normal cells. PMID- 15273410 TI - Multigenerational selection and detection of altered histone acetylation and methylation patterns: toward a quantitative epigenetics in Drosophila. AB - Quantitative epigenetics (QE) is a new area of research that combines some of the techniques developed for global quantitative trait loci (QTL) mapping analyses with epigenetic analyses. Quantitative traits such as height vary, not in a discrete or discontinuous fashion, but continuously, usually in a normal distribution. QTL analyses assume that allelic DNA sequence variation in a population is partly responsible for the trait variation, and the aim is to deduce the locations of the contributing genes. QE analyses assume that epigenetic variation in a population is partly responsible for the trait variation, and the aim is to associate inheritance of the trait with segregation of informative epigenetic polymorphisms, or epialleles. QTL and QE analyses are thus complementary, but the latter has several advantages. QTL mapping is limited in resolution because of meiotic recombination and population size, placing quantitative traits on genomic regions that are each typically several megabase pairs long, and requires DNA sequence variation. In contrast, QE analysis can make use of powerful emerging mapping techniques that allow the positioning of epialleles defined by chromatin variation to individual genes or chromosomal regions, even in the absence of DNA sequence variation. In this chapter, we present a case study for QE analysis-epigenetic mapping of enhancers of the KrIf 1 ectopic eye bristle phenotype in an isogenic strain of Drosophila melanogaster. PMID- 15273411 TI - Profiling DNA methylation by bisulfite genomic sequencing: problems and solutions. AB - The surge of interest in DNA methylation during the last two decades has triggered an urgent need for an effective method to detect the methylation status of the cytosines in the genome. Bisulfite genomic sequencing is the most attractive choice so far for many laboratories. Various protocols have been established, but difficulties are often encountered, particularly by individuals who have limited experience in this field. This analysis presents a simple protocol that has consistently worked well in our laboratory. Discussions of potential technical problems and corresponding solutions are also included to facilitate the reproducibility of this protocol. PMID- 15273412 TI - Methylation-sensitive single-strand conformation analysis: a rapid method to screen for and analyze DNA methylation. AB - The last few years have seen a growing interest in the study of DNA methylation because of its now acknowledged implication in cancer. The use of bisulfite to convert unmethylated cytosine to uracil, even as methylated cytosine remains unchanged, constitutes the basis for differentiating between methylated and unmethylated specific CpG sites in CpG islands. This technique therefore is critical to the success of this approach. Different parameters have to be considered in order to achieve a total conversion of cytosines to uracils. Several bisulfite-based methods are available for analyzing DNA methylation status. Methylation-sensitive single-strand conformation analysis (MS-SSCA) yields specific and semiquantitative data. The method is based on bisulfite treatment of DNA followed by polymerase chain reaction using primers without a CpG site to avoid selective amplification of either methylated or unmethylated DNA, and finally by single-strand conformation analysis (SSCA). The method allows one to establish clonal variations in the DNA methylation status for clones representing as little as 5-10% of the total cell population. MS-SSCA has, furthermore, a broad application field since it is the appropriate method for the analysis of frozen, fixed, and even microdissected tissues. PMID- 15273413 TI - SIRPH analysis: SNuPE with IP-RP-HPLC for quantitative measurements of DNA methylation at specific CpG sites. AB - This chapter describes a detailed protocol using single-nucleotide primer extension (SNuPE) for quantitative analysis of DNA methylation on specific CpG sites. The first step DNA sample to be studied is treated with sodium bisulfite, which converts selectively unmethylated cytosines to uracil, while methylated cytosines remain unconverted. Subsequently, a SNuPE reaction is performed, with an oligo just flanking a CpG site, using a purified polymerase chain reaction product derived from bisulfite-treated DNA as a template. The oligo is extended by either ddCTP or ddTTP depending on whether the site is methylated or unmethylated, respectively. The reaction is quantitative and linear, and two to three sites can be studied simultaneously in a multiple reaction. The SNuPE product, without further purification, is separated by ion-pair reverse-phase (IP RP) high-performance liquid chromatography (using an alkylated nonporous polysterene-divinylbenzene cartridge) that allows an easy, semiautomated method for separation of the extended and unextended products and an accurate quantification of the extended products. The ratio of the ddCTP to the ddTTP gives the fraction of the methylated cytosines at that specific CpG site. PMID- 15273414 TI - Real-time PCR-based assay for quantitative determination of methylation status. AB - The best studied epigenetic modification in mammals is the methylation of cytosine. During the development and progression of malignant neoplasia, a global hypomethylation is often accompanied by a locus-specific increase in methylation. Also, during normal development specific alterations in DNA methylation patterns take place. In recent years it has become clear that in many situations only quantitative changes in methylation levels occur and that the pure qualitative detection of cytosine methylation misses important biological and pathophysiological information. Therefore, several protocols were developed for the quantitative detection of cytosine methylation. Here, we describe a real-time polymerase chain reaction-based assay for the sensitive and precise quantification of methylated and unmethylated alleles after bisulfite treatment of genomic DNA. In addition to providing quantitative methylation data, this methodology is suitable for high-throughput analysis. PMID- 15273415 TI - Denaturing gradient gel electrophoresis to detect methylation changes in DNA. AB - Denaturing gradient gel electrophoresis (DGGE) is a technique that fractionates DNA molecules on the basis of their melting behavior and thereby permits the separation of DNA fragments with local variations in base composition. The separation of DNA fragments by DGGE is determined by the nucleotide sequence, rather than size. This approach is effective when part of the molecule is relatively dense in G+C pairs. This separation is possible because of the pronounced drop in electrophoretic mobility in a polyacrylamide gel that occurs when a region of a DNA molecule melts, thereby forming a structure that is partly helical and partly random chain. The electrophoretic mobility of these partly melted DNA fragments is much lower than that of fully helical or fully dissociated molecules. The low residual mobility of the fragment restricts migration into more strongly denaturing regions of the gradient gel and results in focusing of the band. This property can be applied to detect the difference in melting temperature between methylated and nonmethylated DNA fragments after chemical treatment, or to enrich genomic regions in which aberrant methylation occurs. In this chapter, the application of DGGE to the analysis of genomic DNA methylation is reviewed. PMID- 15273416 TI - Photocrosslinking oligonucleotide hybridization assay for concurrent gene dosage and CpG methylation analysis. AB - The phenotypic effects of aberrant gene expression are indistinguishable, regardless of whether the underlying mutation is one of gene copy number (deletion or duplication) or modification of differentially methylated CpG sites occurring in critical regulatory sequences in gene promoters. The XLnt photocrosslinking oligonucleotide technology provides for the hybridization dependent covalent attachment of probe to target, allowing survival of the probe/target complex under otherwise denaturing conditions. Posthybridization wash stringency can be substantially higher than under standard techniques, leading to a marked increase in signal-to-noise ratios. In addition, the reduction in nonspecific background provides linearity to XLnt-based oligonucleotide assays comparable to that otherwise achieved only with very long probes. The technology is thus ideally suited for combining the high-throughput capacity of oligonucleotide hybridization platforms with accurate measurement of relative gene dosage. By integrating the XLnt system with an assay design separating probe/target immobilization and signal elaboration functions, relative gene dosage assessment can be applied to the quantitation of fractional resistance to methylation-sensitive restriction enzyme (MSRE) digestion. The method described below provides for the development of photocrosslinking oligonucleotide assays for relative gene dosage and fractional locus CpG methylation in a microtiter plate-based format. PMID- 15273417 TI - Methylation-specific oligonucleotide microarray. AB - The methylation-specific oligonucleotide (MSO) microarray is a high-throughput approach capable of detecting DNA methylation in genes across several CpG sites. Based on the bisulfite modification of DNA that converts unmethylated cytosines to uracil but leaves the 5'methylcytosine intact, the method utilizes short oligonucleotides corresponding to the methylated and unmethylated alleles as probes affixed on solid support and products amplified from bisulfite-treated DNA as targets for hybridization. MSO is suitable for examining a panel of genes across multiple clinical samples. This approach can generate a robust dataset for discovering profiles of gene methylation in cancer with aberrant DNA methylation in the neoplastic genome and widespread hypermethylation in tumor suppressor genes. MSO and other oligonucleotide-based arrays have been applied successfully for analyses of single genes and have been useful in delineating and predicting tumor subgroups using clustering methods. Here we focus on design criteria important to the interrogation of multiple CpG sites across several genes. PMID- 15273418 TI - Methylation-specific PCR in situ hybridization. AB - Methylation-specific polymerase chain reaction (PCR) in situ hybridization, using paraffin-embedded, formalin-fixed tissues or formalin-fixed cell preparations, allows one to determine which specific cells have silencing of a given gene owing to hypermethylation of the promoter. Standard in situ hybridization, after conversion of nonmethylated bases, would not have sufficient sensitivity to detect the one or two copies of the promoter region of interest. The framework of the methodology is the same as solution-phase methylation-specific PCR. However, when working with intact tissue or cell preparations, adequate formalin fixation and protease digestion are essential for satisfactory results. Furthermore, since one often uses an oligoprobe, probe size (at least 40 bp), labeling method (3' end tailing), probe concentration, and posthybridization stringency conditions can all have an impact on the final results. PMID- 15273419 TI - Relative quantitation of DNA methyltransferase mRNA by real-time RT-PCR assay. AB - DNA methylation is one mechanism of epigenetic gene regulation and influences gene expression by recruiting methylcytosine-binding proteins and/or inducing changes in chromatin structure. In mammals, DNA methylation is mediated by at least four DNA methyltransferase (Dnmt) enzymes, including Dnmt1, Dnmt2, Dnmt3a, and Dnmt3b. To understand fully how DNA methylation is involved in gene regulation, knowledge of Dnmt mRNA transcript levels is required, both as a surrogate measure of Dnmt protein levels and also to facilitate an understanding of the regulation of expression of the corresponding genes. Measurement of transcript levels has traditionally been achieved by Northern blot analysis and more recently either by the ribonuclease protection assay or by reverse transcription polymerase chain reaction (RT-PCR), followed by agarose gel electrophoresis. In the past few years, a form of PCR has been developed that measures the accumulation of PCR product in real time. In conjunction with RT, real-time RT-PCR has become a widely accepted tool for measuring mRNA transcript levels and is now probably the method of choice. This technique is both sensitive and specific and allows for the rapid assessment of Dnmt mRNA transcript levels as well transcripts for other genes that may be involved in DNA methylation. PMID- 15273420 TI - DMB (DNMT-magnetic beads) assay: measuring DNA methyltransferase activity in vitro. AB - DNA methylation is an epigenetic modification of DNA that leads to heritable alterations in transcriptional regulation and conformational changes in chromatin structure of higher eukaryotes. Mammalian DNA methyltransferases, which are the enzymes responsible for DNA methylation, have attracted the attention of both basic and clinical researchers because they appear to participate in embryogenesis and carcinogenesis via chromatin modification. DNA methyltransferase catalyzes the transfer of a methyl group into DNA strands. Since traditional assays for DNA methyltransferase activity in vitro have insufficient reproducibility, there is a need in the art for more sensitive and quantitative methods for measuring enzymatic activity. We report a novel assay system, in which the activity of a DNA methyltransferase is measured as the incorporation of tritium into biotinylated DNA oligonucleotides. The DNA is immobilized onto magnetic beads with streptavidin covalently attached to the bead surface. The radioactive DNA can easily be separated from the unreacted radioactive substrate using a magnet. The radioactivity is counted by the liquid scintillation system. This DMB assay is simple and easy, has very low background, and, most importantly, is highly reproducible for the precise enzymatic analysis of any DNA methyltransferase in vitro. PMID- 15273421 TI - Impaired neuroendocrine response to stress following a short-term fat-enriched diet. AB - Unbalanced diets and stressful situations disrupt energy homeostasis and are implicated in the development of severe pathologies. The present study investigated the effects of a 7-day diet, enriched in corn oil (20%) and proportionally lower in protein and carbohydrate, on the major regulators of energy expenditure and stress response of adult male Wistar rats exposed to acute swimming stress at the end of the dietary treatment. Food intake and body weight gain were lower in diet-fed as compared with normal-chow-fed controls. The circulating leptin levels were elevated in both nonstressed and stressed diet-fed rats, while the glucose levels were significantly increased only in the diet-fed group subjected to stress. The plasma insulin levels were not affected by the diet, but were significantly reduced in acutely stressed rats. Acute swimming increased corticosterone levels both in chow-fed and diet-fed rats. No significant effect of diet was observed on corticosterone levels. Northern blot analysis showed increased glucocorticoid receptor mRNA levels in the hypothalamus of normally fed rats subjected to stress. This increase was not observed in the diet-fed stressed group, which on the contrary showed reduced glucocorticoid receptor mRNA levels following stress. The data presented indicate that even a moderately unbalanced, fat-enriched diet can within a short time disrupt the metabolic neuroendocrine balance and the stress response, rendering the organism more vulnerable to potential stressful insults. PMID- 15273422 TI - Acute effects of cyclooxygenase-2 inhibition on renal function in heterozygous ren-2-transgenic rats on normal or low sodium intake. AB - BACKGROUND/AIMS: Since there are no data available so far on the role of renal cyclooxygenase-2 (COX-2) in hypertensive Ren-2-transgenic rats (TGR), in the present study we evaluated renal cortical COX-2 protein expression and prostaglandin E2 (PGE2) concentrations as well as renal functional responses to acute COX-2 inhibition in male heterozygous TGR and in normotensive Hannover Sprague-Dawley (HanSD) rats fed either a normal-sodium (NS) or a low-sodium (LS) diet. METHODS: In rats fed either the NS or the LS diet for 12 days and prepared for clearance experiments with left ureteral catheterization, the renal functional responses of the left kidney were evaluated after intrarenal COX-2 inhibition with DuP-697 or NS-398. In renal cortical tissue, COX-2 protein expression was assessed by immunoblotting, and the concentration of PGE2 as a marker of COX-2 activity was determined by enzyme immunoassay. Mean arterial pressure in the right femoral artery was monitored by means of a pressure transducer. RESULTS: In heterozygous TGR, to our surprise, the LS diet normalized the mean arterial pressure. Despite significantly higher renocortical expression of COX-2 and PGE2 concentrations as well as urinary PGE2 excretion in TGR as compared with HanSD rats kept on the NS diet, selective intrarenal COX-2 inhibition did not influence renal function either in TGR or in HanSD rats. The LS diet increased renocortical COX-2 expression and PGE2 concentrations as well as urinary PGE2 excretion significantly stronger in TGR than in HanSD rats. Regardless of these increases, the intrarenal COX-2 inhibition caused comparable decreases in glomerular filtration rate, in absolute and fractional sodium excretion, as well as in urinary PGE2 excretion in TGR and HanSD rats kept on the LS diet. CONCLUSIONS: The present data show that a LS diet normalizes the mean arterial pressure in heterozygous male TGR. This first study on the role of renal COX-2 in TGR also demonstrates that COX-2-derived vasodilatory prostanoids do not act as renal compensatory vasodilator and natriuretic substances in this model of hypertension. PMID- 15273423 TI - Safety and efficacy of single bolus anticoagulation with enoxaparin for chronic hemodialysis. Results of an open-label post-certification study. AB - BACKGROUND: Low-molecular-weight heparin (LMWH) is supposed to be advantageous compared to unfractionated heparin for chronic hemodialysis (HD) with respect to lipid and bone metabolism, polymorphonuclear cell stimulation, induction of antibody-mediated thrombocytopenia, and aldosterone suppression. Due to longer biological half-life, LMWH offers the possibility of single bolus administration. METHODS: To assess safety and efficacy of single bolus anticoagulation with enoxaparin for chronic HD, 781 stable HD patients from 79 German dialysis centers (mean age 62 years; 31% ESRD due to diabetes mellitus) were monitored by clinical and laboratory parameters for 32 weeks. Additionally, in a single dialysis center, 22 chronic HD patients were investigated by molecular markers of coagulation during chronic HD under conditions of single bolus or continuous anticoagulation regimens. Anti-Xa activity and the thrombin- antithrombin-III complex (TAT) were determined before the enoxaparin bolus, after 15 min, 2 h, and at the end of HD in venous and arterial blood lines. RESULTS: Chronic HD was performed in 24,117 HD treatments with enoxaparin at a median dose of 70.1 IU/kg (5,000 IU median total dose) for single bolus anticoagulation. In 83.0% of HD treatments, enoxaparin was given as single bolus. In 98.3% of patients no adverse event was reported. No drug-related severe adverse event occurred. Significant clotting problems were observed in only 0.3% of HD treatments with single bolus anticoagulation. As assessed in 257 HD treatments, essentially identical anti-Xa levels were detected at the end of HD with single bolus (50 IU/kg) or continuous (mean total dose 43 IU/kg) anticoagulation regimens. Bolus anticoagulation resulted in higher TAT generation at the end of HD. However, this was not associated with increased macroscopic clot formation. CONCLUSION: Single bolus anticoagulation with enoxaparin was safe and effective for chronic HD. For a duration of 4 h HD, a median dose of 70 IU/kg can be recommended for regular use, which is in accordance with the manufacturer's instructions for use of enoxaparin recommending a range of 50-100 IU/kg. PMID- 15273424 TI - Combination therapy with angiotensin-converting enzyme inhibitor and oral adsorbent of uremic toxins can delay the appearance of glomerular sclerosis and interstitial fibrosis in established renal failure. AB - BACKGROUND/AIM: Angiotensin II plays a central role in the progression of chronic renal failure (CRF), and administration of angiotensin-converting enzyme inhibitor (ACEI) in rats delays the progression of CRF. However, ACEI has little effect on CRF progression in rats with established CRF. We therefore examined whether combination therapy with ACEI and oral adsorbent for uremic toxins in the gastrointestinal tract has the desired effect. METHODS: Rats subjected to subtotal nephrectomy were given enalapril at 20 mg/kg (n = 10, group E), AST-120 at 5 g (n = 10, group A), enalapril and AST-120 together at the same doses (n = 10, group EA), or no treatment (n = 10, group C) 8 weeks after the operation. The substances were administered in 100 g rat chow. All animals were pair-fed, and all were killed after 8 weeks of pair-feeding. RESULTS: Body weight did not differ between groups during the study. Blood pressure at week 8 was significantly lower in groups E and EA than in groups C and A (p < 0.05). Urinary protein excretion level and renal plasma flow rate at week 8 were significantly less in groups E and EA than in group C (p < 0.05, p < 0.01). The glomerular filtration rate at week 8 was significantly higher in group EA than in group C (p < 0.05). The glomerular sclerosis index and interstitial fibrosis area at week 8 were significantly less in group EA than in group C (p < 0.01). CONCLUSION: ACEI and AST-120 in combination can delay progression of established CRF in rats by inhibiting the appearance of glomerular sclerosis and interstitial fibrosis. PMID- 15273425 TI - Identifying chemokines as therapeutic targets in renal disease: lessons from antagonist studies and knockout mice. AB - Chemokines, in concert with cytokines and adhesion molecules, play multiple roles in local and systemic immune responses. In the kidney, the temporal and spatial expression of chemokines correlates with local renal damage and accumulation of chemokine receptor-bearing leukocytes. Chemokines play important roles in leukocyte trafficking and blocking chemokines can effectively reduce renal leukocyte recruitment and subsequent renal damage. However, recent data indicate that blocking chemokine or chemokine receptor activity in renal disease may also exacerbate renal inflammation under certain conditions. An increasing amount of data indicates additional roles of chemokines in the regulation of innate and adaptive immune responses, which may adversively affect the outcome of interventional studies. This review summarizes available in vivo studies on the blockade of chemokines and chemokine receptors in kidney diseases, with a special focus on the therapeutic potential of anti-chemokine strategies, including potential side effects, in renal disease. PMID- 15273426 TI - Does a nephron deficit in rats predispose to salt-sensitive hypertension? AB - AIM: This study tested the hypothesis that a nephron deficit predisposes rats to salt-sensitive hypertension in adulthood. METHODS: Female Wistar-Kyoto rats were fed a low (9%) or a normal (20%) protein diet during pregnancy and lactation. Male, birth-weight-matched offspring were paired. One rat from each pair was perfusion fixed at 4 weeks of age and the other rat at 40 weeks of age. Kidneys were removed and nephron number and total renal filtration surface area (FSA) determined using unbiased stereological techniques. The rats that were allowed to grow to adulthood had tail-cuff systolic blood pressure and body weight determined twice weekly. Between 30 and 40 weeks of age, a normal or a high-salt diet was fed to the rats. RESULTS: The offspring of rats fed the low-protein diet were significantly smaller at birth, and at 4 weeks of age they had a significant reduction in kidney volume, nephron number, and total renal FSA when compared to controls. Tail-cuff systolic blood pressure in the offspring from 4 to 29 weeks of age did not significantly differ between the two groups. Administration of a high-salt diet from 30 to 40 weeks of age led to a significant increase in blood pressure in both dietary treatment groups; however, it was not exacerbated in the rats exposed to the low-protein diet in utero. CONCLUSIONS: Maternal protein restriction in rats did not lead to salt-sensitive hypertension. Nephron endowment and FSA did not correlate with blood pressure in adulthood. PMID- 15273427 TI - Treatment of pubertal gynecomastia with the specific aromatase inhibitor anastrozole. AB - Gynecomastia can be detected in up to 70% of boys during puberty and in about one third of adult males. An imbalance of estrogen to androgen tissue levels is believed to be the major reason for the development of gynecomastia; as a result most medical treatments so far have tried to lower the estrogen level. Five boys with pubertal gynecomastia and breast tenderness were treated for 6 months with the selective aromatase inhibitor anastrozole. Initial plasma levels of estradiol (E2), testosterone (T), androstenedione, dehydroepiandrosterone sulfate (DHEA-S) and gonadotropins were normal. DHEA-S showed a significant rise during treatment. T and androstenedione showed no significant change during treatment. E2 decreased with therapy, although to no statistically significant extent. The E2/T ratio decreased significantly during the treatment. Breast size decreased in 4 out of 5 patients, and in 1 of these 4 boys glandular breast tissue disappeared completely. The longer the duration of gynecomastia before anastrozole administration, the smaller was the reduction of breast size. Breast tenderness was resolved in all boys within 4 weeks. No adverse effects were recorded. Since the aim of medical treatment is the total disappearance of breast tissue, anastrozole, as previous aromatase inhibitors, is of limited effect. However, anastrozole seems to be of benefit for the treatment of tenderness in gynecomastia and for patients in whom surgery is particularly risky. However, as spontaneous disappearance of pubertal gynecomastia is common, further double blinded, placebo-controlled trials are necessary before a definite conclusion can be drawn about the effectiveness and the side effects of this therapy. PMID- 15273428 TI - Blink reflex abnormalities in chronic alcoholics. AB - PURPOSE: The aim of the present study was to evaluate the efficacy of blink reflex as a method for obtaining early diagnosis of cranial nerve involvement in alcoholic patients. MATERIALS AND METHODS: The study was conducted on 30 male alcoholics with a mean age of 43 years. They had histories of alcohol abuse for at least 6 years (mean: 25). At the time of recording, they had undergone detoxification treatment for a mean of 27 days. RESULTS: R1 (early response), R2Y (second ipsilateral response), and R2C (second contralateral response) latencies in alcoholics were prolonged relative to controls and the differences were statistically significant (p < 0.02, p < 0.001, p < 0.001, respectively). According to the defined criteria, 40% of the patients had abnormal responses, and the most common abnormality was the unilateral prolongation of R1 (13%). CONCLUSION: Finding abnormal blink reflex responses in alcoholic patients has suggested that blink reflex testing is a useful method for the evaluation of subclinical cranial nerve involvement in alcoholic patients. Blink reflex testing may be useful in detecting early changes and in the follow-up of alcoholic disorder. PMID- 15273429 TI - Beta-interferon treatment reduces human herpesvirus-6 viral load in multiple sclerosis relapses but not in remission. AB - To determine whether the DNA prevalence of human herpesvirus-6 (HHV-6), the viral load and the prevalence of both HHV-6 variants in relapsing-remitting multiple sclerosis (RRMS) patients in exacerbation are altered by beta-interferon (IFN beta) treatment, in comparison with RRMS patients in remission, we analyzed HHV-6 (A and B) genomes in 189 serum samples by quantitative real-time polymerase chain reaction: 105 of the RRMS patients were receiving IFN-beta treatment (48 in exacerbation) and 84 were untreated (36 in relapse). The results were as follows. (1) Prevalence decrease because of IFN-beta treatment was not significant: 25% of RRMS patients in relapse vs. 15.9% in remission (p = 0.45). (2) Viral load was twice as much in untreated patients in relapse than in treated ones. (3) We only found variant A. Since IFN-beta treatment is able to significantly reduce HHV-6 viral load in RRMS patients in relapse, but not in remission, we suggest a role for HHV-6 in the pathogenesis of multiple sclerosis exacerbations and an antiviral role for IFN-beta treatment in RRMS. PMID- 15273430 TI - Cognitive, but not mood dysfunction develops in multiple sclerosis during 7 years of follow-up. AB - Long-term development of psychological deficits in disability-free early multiple sclerosis (MS) was evaluated in 27 female patients over a period of 7 years and compared with healthy controls. Physical and cognitive parameters deteriorated significantly but not depression scores. In particular, the self-assessed somatic complaints remained non-similar between patients and controls. This indicates that although depression is clinically relevant and frequent in MS, in contrast to cognition it is not related to physical disease progression. PMID- 15273431 TI - DNA haplotype analysis of CAG repeat in Taiwanese Huntington's disease patients. AB - We studied the expanded CAG repeat and adjacent CCG repeat in 53 Huntington's disease (HD) patients and 172 unrelated normal subjects matched to the patients for ethnic origin. The range of the CAG repeat varied from 38 to 109 in the HD patients and from 10 to 29 in the control group. A significant negative correlation was found between the age at onset and the CAG expansion, with no significant influence of the adjacent CCG repeat on the age at onset by multiple regression analysis. Allelic association using CCG repeat and 2 flanking dinucleotide repeat markers within 150 kb of the HD gene revealed linkage disequilibrium for 2 of 3 markers. Haplotype analysis of 24 HD families using these markers identified 3 major haplotypes underlying 87.5% of HD chromosomes. The data suggested frequent haplotypes in the Taiwanese population on which one or more mutational events leading to the disease occurred. PMID- 15273432 TI - Cerebral venous infarction: the pathophysiological concept. AB - Cerebral venous occlusion represents an often underdiagnosed cause for acute or slowly progressive neurological deterioration. The underlying pathophysiological basis is not well understood, but is different from those of arterial occlusion reflecting therefore different anatomical and physiological features of the cerebral venous system. Extensive collateral circulation within the cerebral venous system allows for a significant degree of compensation in the early stages of venous occlusion. Elevated cerebral venous pressure due to cerebral venous occlusion can result in a spectrum of phenomena including a dilated venous and capillary bed, development of interstitial edema, increased cerebrospinal fluid production, decreased cerebrospinal fluid absorption and rupture of venous structures (hematoma). All of these pathophysiological changes may explain the clinical observation that cerebral venous occlusion, if promptly diagnosed and adequately managed, contains reversible alterations and need not always lead to venous infarction. The present review outlines this different pathophysiological behavior of venous compared to arterial occlusion in the cerebral vasculature; special reference is given to the effect of these changes on the therapeutic impact. PMID- 15273433 TI - Mutations in the lysyl oxidase gene not associated with intracranial aneurysm in central European families. AB - BACKGROUND: Lysyl oxidase is a promising candidate gene for a mutation search in intracranial aneurysm families because (a) it controls the processing, cross linking and maturation of collagen and elastin fibers in the blood vessel wall, (b) its expression levels and activity are altered in different animal models of aneurysm pathogenesis, and (c) it is encoded within the chromosome 5q22-31 region of suggestive linkage to intracranial aneurysms. METHODS: We have performed genomic sequencing of all 7 exons including the intron-exon splice sites and of the putative promoter region for lysyl oxidase in 25 patients from intracranial aneurysm multiplex families resident in Central Europe. RESULTS: We observed 4 genetic variants including 2 novel polymorphisms, 1 in the noncoding sequence of exon 7 and the other upstream from the lysyl oxidase promoter. None of these single nucleotide polymorphisms showed an allelic association or cosegregation with intracranial aneurysm in the families. CONCLUSIONS: Genetic variants in the lysyl oxidase gene do not appear to be a major factor in the etiology of intracranial aneurysms in Central Europe. PMID- 15273434 TI - Serum cardiac troponin I in acute stroke is related to serum cortisol and TNF alpha. AB - BACKGROUND: Serum cardiac troponin I (cTnI) is a specific marker of myocardial injury related to in-patient fatality and cardiac injury in acute stroke. We investigated whether cTnI in acute stroke is related to serum cortisol, acute inflammatory response, and insular damage. We also investigated whether cTnI predicted outcome at 3 months. PATIENTS AND METHODS: The study was based on 155 patients with CT-confirmed acute cerebral infarction and study inclusion within 24 h (50% within 12 h) of stroke onset. Blood samples were obtained on inclusion. Stroke severity was assessed by the Scandinavian Stroke Scale (SSS) and outcome was assessed by the modified Rankin Scale (mRS), death or dependency was defined as mRS > or =3 three months after stroke. RESULTS: 35% of all patients and 63% of patients who died within 3 months were troponin positive. Tumor necrosis factor alpha (TNF-alpha) and cortisol were independently related to detection of cTnI: TNF-alpha(+100 pg/ml) OR 1.5 (CI 95% 1.1-2.2), cortisol(+100 nmol/l) OR 1.1 (CI 95% 1.01-1.2). SSS and age were also included in this model and did not reach significance. cTnI positivity was, together with age, stroke severity and prestroke mRS, but not s-cortisol, an independent explanatory variable of outcome at 3 months (death or dependency) with OR 4.1 (CI 95% 1.1-14.5). cTnI did not relate to insular involvement. CONCLUSION: In this study, cortisol and TNF-alpha were independently related to cTnI, which was predictive of 3-month prognosis. PMID- 15273435 TI - Early carotid surgery in acute stroke: a multicentre randomised pilot study. AB - BACKGROUND: Standard practice has been to delay carotid endarterectomy (CEA) for 2 months following acute stroke to avoid a perceived risk of cerebral haemorrhage. We investigated whether early CEA reduces early recurrent stroke and improves outcome in partial anterior circulation infarction (PACI). METHODS: Patients with PACI and a Barthel score of >18 before stroke underwent carotid duplex and CT imaging within 7 days of stroke. Forty consenting patients fit for surgery with greater than 70% ipsilateral carotid stenosis were randomised, 19 to 'early' (within 24 h) and 21 to 'delayed' surgery (at 8 weeks). Modified Rankin and Barthel scores were recorded at 1 week, 2 months, 6 and 12 months. RESULTS: Rankin scores improved more rapidly following 'early' surgery to a score of 1 (0 4) at 2 and 6 months compared with 2.5 and 2 (1-4), respectively, for delayed surgery (p < 0.05). Barthel scores were also significantly improved following 'early' CEA at 7 days but both groups reached a median score of 20 by 2 months. Four 'delayed' and 3 'early' patients suffered extension or recurrence of neurological deficits with 1 death in each group. CONCLUSIONS: Early CEA within 7 days of ischaemic stroke improved functional outcome with earlier hospital discharge. A large multicentre study is needed to exclude the possibility that 'early' CEA increases the risk of cerebral haemorrhage or death. PMID- 15273436 TI - Salivary cortisol, a biological marker of stress, is positively associated with 24-hour systolic blood pressure in patients with acute ischaemic stroke. AB - BACKGROUND/AIM: The cause of elevated blood pressure (BP) in acute stroke is unknown. Stress is often suggested as a main contributing factor. We aimed to investigate the relationship between BP and stress in patients with acute stroke. METHODS: 58 patients with clinical symptoms of stroke were recruited prospectively after exclusion of haemorrhage by CT scan within 14 h and 15 min (mean) after symptom onset (range 2 h and 45 min-23 h and 40 min). The mean age of the patients was 66 years (range 39-86 years), and the mean National Institute of Health Stroke Scale score was 7 (range 1-26). BP and pulse rate were recorded by non-invasive automatic monitoring hourly for 24 h. Stress was evaluated by testing the level of salivary cortisol. Four samples of saliva were obtained at inclusion, on the evening of the inclusion day (20.00-22.00 h), on the morning of the next day (7.00-9.00 h) and on the afternoon of the inclusion day/next day (15.00-17.00 h) within 24 h after inclusion in the study. Logarithmic transformation was done for cortisol levels. RESULTS: The 24-hour mean cortisol level (geometric mean 13.6 nmol/l) was related to 24-hour mean systolic BP [SBP; r = 0.36, p = 0.01, multivariate p = 0.02], mean night-time (22.00-6.00 h) SBP (r = 0.43, p = 0.001, multivariate p < 0.005) and mean night-time diastolic BP (r = 0.31, p = 0.02, multivariate p = 0.02). Cortisol levels at inclusion (r = 0.31, p = 0.02, multivariate p = 0.05 for 24-hour SBP) and in the evening were also statistically significantly related to the above BP variables. The morning cortisol (r = 0.28, p = 0.04, multivariate p = 0.04) was related to night-time SBP. CONCLUSIONS: Salivary cortisol was positively correlated with 24-hour SBP and night-time BP, suggesting that stress is a contributing factor for high BP in acute stroke. PMID- 15273437 TI - C-reactive protein and white blood cell count increases in the first 24 hours after acute stroke. AB - Levels of C-reactive protein (CRP) and white blood cell count (WBC) in acute stroke may reflect the stroke lesion itself or pre-existing factors such as infections, smoking or atherosclerosis. The aim of this study was to investigate the relation between CRP and WBC levels and time from onset of stroke, stroke severity and outcome. PATIENTS AND METHODS: The analyses were based on 719 patients in whom WBC test material was obtained within 9 h of stroke onset and CRP test material within 24 h of stroke onset. Stroke severity was assessed by the Scandinavian Stroke Scale Score on admission and outcome by death 7 days, 3 months and 1 year after symptom onset as well as modified Rankin Scale 3 months after stroke onset. RESULTS: CRP and WBC levels correlated significantly with time from symptom onset as well as with stroke severity and outcome. Levels of CRP and WBC were higher in later determinations in severe stroke. In multivariate logistic regression analysis, CRP(+10 mg/l) was independently related to 1-year mortality (OR 1.1, 95% CI 1.02-1.2). CONCLUSION: Levels of WBC and CRP increase within the first 24 h in patients with severe stroke. CRP but not WBC is related to long-term mortality possibly by reflecting the vascular risk profile. PMID- 15273438 TI - Reliability of a semi-automated technique of cerebral infarct volume measurement with CT. AB - BACKGROUND: A reliable method of infarct volume measurement is needed if infarct volume is to be used as an outcome measure in clinical stroke trials. We investigated the reproducibility of a semi-automated method of computed tomography (CT) infarct volume measurement amongst three stroke research fellows with no formal neuroradiology training and two consultant neuroradiologists. METHODS: CT brain scans for volumetric analysis were performed at 5 to 7 days in 34 patients with acute ischaemic stroke, of which 28 scans showed visible recent infarction. Five observers independently traced the infarct boundary on digitised images with a cursor. Volumetric analysis incorporated pixel thresholding with preset Hounsfield thresholds. One of the observers repeated the analyses on 21 of the scans in order to assess intraobserver variation. RESULTS: Median infarct volume was 35.7 cm3 (range 0.2-318 cm3). The closest limits of observed agreement (mean +/- 1.96 SD) between pairs of observers were between a research fellow and neuroradiologist (-29 to 21 cm3). The widest limits of agreement were between a different research fellow and the same neuroradiologist (-39.1 to 41.4 cm3). The limits of agreement between infarct volumes measured on two separate occasions by one of the research fellows were -7 to 8 cm3. CONCLUSIONS: Intraobserver reliability of CT infarct volume measurements performed by a stroke research fellow was superior to interobserver reliability between any pair of observers. The wide limits of agreement between different observers using manual tracing may not be acceptable in multicentre trials of acute ischaemic stroke treatment, but volume measurement by a single observer appears to be more reliable. PMID- 15273439 TI - First-year results of the third stroke registry in Tartu, Estonia. AB - BACKGROUND: The main goal of the Third Stroke Registry in Tartu was to determine the incidence and 28-day case-fatality rates for first-ever stroke in an Estonian population. METHODS: The data collection started on 01.12.2001. All patients with first-ever stroke living in Tartu were registered. RESULTS: During the first year, 234 first-ever stroke cases were registered. The incidence rate of first ever stroke age-standardised to the European population was 195/100,000, 214 (95% CI, 185-243) for men and 181 (95% CI, 155-208) per 100,000 for women. Sixty-eight patients (29%) died within 28 days of stroke onset. CONCLUSIONS: The incidence and 28-day case-fatality rate of stroke in Estonia are high compared to other countries. It might be related to higher risk factor prevalence, stress and socioeconomic status. PMID- 15273440 TI - Magnetic resonance imaging: significance of early ischemic changes on computed tomography. AB - The significance of early ischemic changes (EICs) on CT remains controversial. MRI may provide relevant information in patients with EICs. METHODS: EICs were assessed in patients with acute ischemic stroke. MRI was promptly performed at presentation after CT and repeated on day 1. The relationship between EICs and MRI parameters was assessed with one-way ANOVA for analysis of continuous variables and by the chi2 test for the analysis of variables with a binary outcome. RESULTS: Fourty-eight patients underwent CT and MR imaging before treatment with recombinant tissue plasminogen activator (age: 63 +/- 14 years). EICs were graded as absent in 28 patients, <33% in 15 patients, and >33% of the middle cerebral artery (MCA) territory in 5 patients. NIHSS score was higher in patients with EICs that covered more than one third of the MCA territory (19 +/- 3) compared to those without EICs (12 +/- 5; p = 0.04). Patients who had major EICs had a larger acute lesion volume in diffusion-weighted imaging (DWI; 140 +/- 78 cm3) compared to those without EICs (33 +/- 51 cm3, p < 0.0001). Regional cerebral blood flow, regional cerebral blood volume, time to peak and mean transit time values were not significantly different in the study groups. CONCLUSION: EICs reflect mainly a larger DWI lesion. PMID- 15273441 TI - Importance of anatomical asymmetries of transverse sinuses: an MR venographic study. AB - Time-of-flight (TOF) magnetic resonance venography (MRV) is often used to examine the intracranial dural sinuses, particularly in the evaluation of dural sinus thrombosis. The goal of the study was to evaluate the use of TOF MRV in assessing the normal anatomy of dural sinuses and their variations as sources of potential pitfalls in the diagnosis of venous sinus thrombosis. Cerebral TOF MRV obtained in 105 persons with normal MR studies were reviewed to determine the presence, aplasia and hypoplasia of the transverse sinuses. Twenty-one (20%) aplasias of the left sinus, 41 (39%) hypoplasia of the left sinus, 33 (31%) symmetric, 6 (6%) hypoplasia of the right sinus, and 4 (4%) aplasias of the right sinus cases were determined in the asymmetry in sizes of transverse sinuses. These results suggested that transverse sinus flow gaps or aplasias can be observed in approximately 24% of normal population on MR imaging. The rate of these gaps in normal subjects must be kept in mind because it can be a source of misdiagnosis in cases of suspected dural sinus thrombosis. PMID- 15273442 TI - The evaluation of Wallerian degeneration in chronic paediatric middle cerebral artery infarction using diffusion tensor MR imaging. AB - BACKGROUND: The long-term neuromotor outcome in paediatric strokes ranges from normal to varying degrees of hemiplegia. We evaluated the indices of diffusion tensor magnetic resonance imaging (DTI), fractional anisotropy and mean diffusivity to determine if these indices can identify and quantify the presence of Wallerian degeneration in paediatric patients with chronic middle cerebral artery infarction, and to determine if these quantitative parameters correlate with the neuromotor outcome. METHODS: Eleven children (mean age 8.1 years) with evidence of unilateral middle cerebral artery stroke on magnetic resonance imaging and 10 control subjects (mean age 8.7 years) were studied. Neuromotor outcome was based on functions of the affected hand: mild (n = 3), moderate (n = 6), and severe (n = 2) hemiparesis. Fractional anisotropy and mean diffusivity of the ipsilateral corticospinal tract were compared with matched contralateral regions using the Mann-Whitney U test. Spearman's test was performed to study the relationship between neuromotor outcome and the following: ipsilateral-to contralateral ratio of fractional anisotropy, mean diffusivity and cerebral peduncle area, and the largest infarction size. RESULTS: For control subjects, there were no significant differences in fractional anisotropy and mean diffusivity of the corticospinal tract between the right and left side. For patients, fractional anisotropy decreased by 18% and mean diffusivity increased by 8% in the ipsilateral compared to the contralateral corticospinal tract. Neuromotor outcome correlated with the ipsilateral-to-contralateral ratio of fractional anisotropy (r = -0.638, p = 0.035) but not with the mean diffusivity ratio, cerebral peduncle area ratio and largest infarction size. CONCLUSION: DTI can be used to detect and quantify Wallerian degeneration in chronic paediatric middle cerebral artery infarction. Our preliminary data show that loss of anisotropy in the corticospinal tract correlates with neuromotor outcome. PMID- 15273443 TI - Acute effects of nonlethal in utero hypoxia on fetal guinea pig heart and lack of persistent cardiac or cerebral effects in the neonate. AB - This study assesses the vulnerability of fetal guinea pig heart to metabolic changes during acute nonlethal in utero hypoxia. Guinea pigs (50-55 days gestation) were exposed to 7% O2 for 2 h and room air for 4 h. Fetal hearts were harvested before hypoxia, at the end of hypoxia, and 4 h after hypoxia, and analyzed for: apoptosis (TUNEL), histology, lipid peroxidation and ATP. A group of posthypoxic dams was taken to gestation. Within 48 h postpartum, the function of neonatal hearts was tested and cerebral histology examined. Fetal heart ATP was decreased by 27% at the end of hypoxia and by 32% 4 h after hypoxia. The lipid peroxides, 4-hydroxynonenal and malondialdehyde, were decreased by 37 and 46%, respectively, by 4 h after hypoxia. The apoptotic index increased from 2% in prehypoxic hearts to 8.4% by 4 h after hypoxia. Fetal heart morphology was unremarkable. Postpartum neonatal cardiac function was not affected and cerebral histology was unremarkable. These results support the conclusion that nonlethal in utero hypoxia has acute effects on the fetal heart but no persistent cardiac or cerebral effects in the postpartum neonate. PMID- 15273444 TI - Benign B-cell precursors (hematogones) are the predominant lymphoid population in the bone marrow of preterm infants. AB - Bone marrow (BM) findings in 3rd-trimester stillborns and full-term living neonates have been previously described. However, there is no information regarding BM composition in living preterm infants. Specifically, it is unknown whether the BM lymphocytosis seen in full-term infants at 1-4 weeks of age also occurs in preterm infants. Furthermore, the lineage of these cells has never been investigated. We used a panel of immunohistochemical stains to characterize the BM composition in 11 neonates (8 living and 3 deceased). Unlike in the other age groups, immature B cells (hematogones) were the most common lymphoid population, accounting for 10-60% (mean 34%) of all cells. In two additional cases (both living patients), flow cytometry revealed a level of 3.8% of immature B cells in a <1-week-old neonate and 25.7% in a 19-week-old infant. Immature B cells were not identified in 6 peripheral blood samples from preterm neonates. These findings are pertinent for the interpretation of BM and peripheral blood samples in this age group as survival improves and diagnostic samples become more common. PMID- 15273446 TI - Efficacy of pneumococcal immunization in patients with renal disease--what is the data? AB - BACKGROUND/AIMS: There is an increased incidence of invasive pneumococcal disease in patients with renal allografts, chronic renal insufficiency (CRI), or nephrotic syndrome (NS). Routine pneumococcal immunization (PI) has been recommended for these patients, but the efficacy of PI in this population is not well established. METHODS: A review was done of studies that reported the immunologic response, efficacy, or safety of PI in patients with renal allografts, CRI, or NS. RESULTS: On review of 26 published studies of PI in this population, all studies demonstrated a serologic response by the majority of patients to at least some pneumococcal serotypes. Use of steroids did not alter this response. In the studies with a greater than 6-month follow-up, declining antibody titers were consistently reported, and this decline was usually more rapid than in healthy controls. However, because the studies of the efficacy of PI in this population involve small numbers of patients and are not controlled, the significance of this decline in titers is not known. The incidence of serious adverse reactions to PI is very low. CONCLUSION: Pending more data, patients with renal transplants, CRI, or NS should continue to be offered PI. PMID- 15273447 TI - Determinants of cardiac electrophysiological properties in mice. AB - INTRODUCTION: The transgenic mouse is a popular model for human inherited cardiac disease. Electrophysiology (EP) studies have recently been performed in transgenic mice to characterize the electrical phenotype of the heart. However, little is known regarding the impact of experimental conditions or model selection on the outcome of EP studies in mice. METHODS AND RESULTS: We investigated the effects of experimental conditions on mouse cardiac EP by (1) comparing the findings of transesophageal pacing with those of invasive intracardiac pacing, (2) elucidating the effects of commonly used anesthetic agents, and (3) determining the impact of changes in body temperature. We also investigated the effects of model selection by (1) studying the dependence on mouse strain, and (2) exploring the effects of age. We found that EP parameters derived by both transesophageal and intracardiac pacing/recordings methods were similar. On the other hand, the anesthetic mixture of ketamine, xylazine, and acepromazine had profound effects on cardiac EP compared to sodium pentobarbital or isoflurane. Meanwhile, compared to normal body temperature (97-99 F), low body temperature (92-94 F) prolonged most cardiac EP parameters, while high body temperature (102-104 F) had little effect. Heart rate was a sensitive indicator of changes in body temperature. Significant differences were observed in specialized conduction system properties among the mouse strains studied (FVB, C57, and DBA). Furthermore, atrial electrical remodeling was evidently associated with age, while ventricular electrical properties were virtually unaltered. In comparison with corresponding invasive EP parameters, we found that the QT interval was not a reliable EP index in the mouse. CONCLUSIONS: Cardiac EP variability may result from differences in experimental techniques including anesthesia and body temperature and from differences in mouse selection including strain and age. The influence of these factors should be considered when characterizing the electrical phenotype of transgenic mice in cardiovascular research. PMID- 15273448 TI - Complementary role of echocardiography and cardiac magnetic resonance in the non invasive evaluation of suspected arrhythmogenic right ventricular cardiomyopathy. PMID- 15273449 TI - Myocarditis as a cause of alternating left bundle branch block. PMID- 15273450 TI - Maximum P-wave duration and P-wave dispersion predict recurrence of paroxysmal atrial fibrillation in patients with Wolff-Parkinson-White syndrome after successful radiofrequency catheter ablation. AB - BACKGROUND: Identification of patients at risk for PAF recurrence after AP ablation is important because of the necessity for additional therapies. OBJECTIVES: To determine whether Maximum P-wave duration (Pmax) and P-wave dispersion (Pd ) detected on surface ECG after successful accessory pathway (AP) ablation can predict the recurrence of paroxysmal atrial fibrillation (PAF). METHODS: Seventy-eight patients with Wolff-Parkinson-White (WPW) syndrome who had at least one documented PAF episode and underwent catheter ablation were enrolled. Pmax, minimum P-wave duration (Pmin) and Pd were determined on a surface ECG recorded on a high resolution computer screen on day 2 after ablation of the AP. RESULTS: There was no significant difference in terms of basic clinical data and electrophysiological findings between patients with (Group-1, n = 19) and without (Group-2, n = 59) recurrence of PAF during follow-up of 21 +/- 10 months. Pmax and Pd were significantly higher in Group-1 than Group-2 (120 +/- 15 vs. 96 +/- 10 ms and 47 +/- 12 vs. 25 +/- 7 ms, respectively; p < 0.001 for both). Pmin didn't differ significantly. A Pmax value of > or = 103 ms separated Group-1 from Group-2 with a sensitivity of 84.2%, specificity of 72.9%, positive predictive value of 50%, and negative predictive value of 93.5%. A Pd value of > or = 32.5 ms separated Group-1 from Group-2 with a sensitivity of 89.5%, specificity of 84.7%, positive predictive value of 65.4%, and negative predictive value of 96.2%. Pmax (p < 0.010) and Pd (p < 0.001) were found to be significant univariate predictors of PAF, whereas only Pd remained significant in multivariate analysis (p = 0.037). CONCLUSION: Pd > or = 32.5 ms and Pmax > or = 103.0 ms predict the recurrence of PAF after ablation with acceptable positive and negative predictive values. Pd > or = 32.5 ms is an independent predictor of recurrence of PAF after catheter ablation in patients with WPW syndrome. PMID- 15273451 TI - Accessory pathway with two conduction times. AB - A 48-year old man was referred for frequent paroxysmal narrow QRS tachycardias. Either a rapid orthodromic tachycardia (220 b/min) using a fast-conducting left lateral concealed atrioventricular (AV) accessory pathway (AP) for its retrograde conduction or a slow orthodromic tachycardia (125 b/min) using the same concealed AP which was slow-conducting, were induced. One application of radiofrequency energy at the earliest site of retrograde conduction suppressed both forms of tachycardias. Reciprocating tachycardias presenting with different retrograde conduction times were related to a single reentrant circuit. PMID- 15273452 TI - Masked complete atrioventricular block in a patient with ventricular preexcitation. AB - A 45-year-old male with a preexcited QRS consistent with WPW syndrome was hospitalized for syncope. ECG monitoring revealed episodes of advanced atrioventricular block. An electrophysiologic study demonstrated right anteroseptal preexcitation and revealed an intermittent block in the accessory pathway and AV complete block causing long periods of spontaneous asystole. A DDD pacemaker was implanted without ablation of the accessory pathway. PMID- 15273453 TI - Dynamic substrate mapping and ablation of ventricular tachycardias in right ventricular dysplasia. AB - BACKGROUND: Ablation of ventricular tachycardias in arrhythmogenic right ventricular dysplasia (ARVD-VTs) still remains a clinical challenge. We reported the value of abnormal electrophysiological substrate mapping for guiding ablation of ARVD-VTs using a non-contact mapping system. METHODS AND RESULTS: Dynamic substrate mapping was performed in three male ARVD patients during sinus rhythm. The sites of earliest activation, exit point and activation sequence were mapped for each induced VT. Three different patterns of substrates were determined in 3 patients and located in right ventricular outflow tract, anterior right ventricular wall, and anterolateral right ventricular wall, respectively. Five different clinical VTs (mean CL, 348 +/- 65 ms) were induced. Of 5 VTs, three originated from or near the boundary of substrate, and two had a remote origin. One VT conducted through the substrate. Linear ablations were created between the sites of the earliest ventricular activation and the VT exit point, or across the critical isthmus. The five clinical VTs were successfully ablated with a median of 17 radiofrequency applications. One patient was treated with amiodarone for a VT not clinically observed. There were no VT recurrences during 8.6 months of follow-up. CONCLUSIONS: Defining the abnormal anatomical VT substrates is useful for understanding the mechanisms of ARVD-VTs and determining an ablation strategy. Linear ablation across a critical isthmus or between the early activation and the exit point can effectively cure these arrhythmias. PMID- 15273454 TI - Doppler evaluation of the descending aorta in patients with hypertrophic cardiomyopathy: potential for assessing the functional significance of outflow tract gradients and for optimizing pacemaker function. AB - We evaluated whether analysis of aortic flow could be useful for determining the functional significance of left ventricular outflow gradients and for optimizing pacing therapy in patients with hypertrophic cardiomyopathy (HOCM). METHODS: Doppler echocardiography was performed in 32 patients with HOCM. Eleven patients with pacemakers (PPM) also underwent treadmill and quality-of-life (QOL) testing in a randomized crossover trial (1 month of backup pacing (AAI at 30 beats per minute), 1 month with an atrioventricular interval (AVI) of 30 ms (DDD30), and 1 month with an "optimized" AVI (DDDop) that maximized the descending aortic Doppler velocity time integral. RESULTS: Patients with HOCM displayed a notch in the aortic Doppler flow profile. The location of the notch in systole corresponded with the development of the peak left ventricular outflow gradient. Aortic flow after the notch was variable ranging from 6-48% of the total flow. In patients with pacemakers, improved response to pacing was noted in those patients that developed the notch early in systole and had subsequent attenuation of aortic flow. Optimizing the AVI was associated with improved exercise tolerance (AAI: 4.6 +/- 2.3 min., DDD30: 5.5 +/- 2.2 min., and DDDop: 7.7 +/- 2.5 min.; p < 0.05) and improved QOL. CONCLUSIONS: Patients with HOCM have a notch in their aortic Doppler flow profile. The location of the notch correlates with the development of the peak left ventricular outflow gradient and flow after the notch is variable. Patients with an early notch and attenuated flow after the notch appear to have the greatest response to pacing therapy. PMID- 15273455 TI - The use of the Doppler pulmonary artery velocity time integral to optimize placement of a ventricular pacing lead in a patient with Ebstein's anomaly. AB - Ebstein's anomaly is a rare congenital heart defect. Patients with severe symptomatic tricuspid regurgitation requiring surgical correction often have conduction system disease. We present a case of a 14 year-old girl with Ebstein's malformation and bioprosthetic tricuspid valve who required permanent pacing for symptomatic bradycardia. The placement of the right ventricular pacing lead was facilitated by the use of the Doppler pulmonary artery velocity time integral as a surrogate for stroke volume. This case demonstrates the importance of site specific pacing and the utility of Doppler echocardiography to optimize lead placement and cardiac performance in patients with Ebstein's anomaly and advanced conduction system disease. PMID- 15273456 TI - Implantable cardioverter defibrillator therapy in patients with ischemic or non ischemic cardiomyopathy and nonsustained ventricular tachycardia. AB - BACKGROUND: Mortality benefit from implantable cardioverter defibrillator (ICD) therapy in ischemic cardiomyopathy (ICM) with non-sustained ventricular tachycardia (NS-VT) and inducible VT is well defined. Although NS-VT may suggest an increased risk of sudden cardiac death (SCD) in non-ischemic cardiomyopathy (NICM), the role of ICD therapy is unclear. This retrospective study compares follow-up data in these two groups after ICD implantation. METHODS: 153 consecutive patients with ICD implantation for NS-VT were analyzed. ICM patients received an ICD if they had inducible VT at electrophysiology study (EPS). NICM patients did not routinely undergo EPS before ICD implantation. RESULTS: There were 48 patients (33 males) in NICM group and 105 patients (89 males) in the ICM group. Baseline characteristics including mean ejection fraction (EF), distribution in various New York Heart Association (NYHA) classes, and the mean duration of follow up in the two groups were similar. 50% of the patients in the NICM group and 36% in the ICM group received appropriate therapies (p = 0.106). The mean number of appropriate therapies in the two groups were similar (23.3 +/- 56.7 and 22.5 +/- 59.5 respectively, p = NS). The percentage of patients with inappropriate therapies in the two groups were 27% and 23% respectively (p = NS). Patients in the NICM group received appropriate ICD discharges at a greater rate (p = 0.02). CONCLUSION: Patients undergoing ICD implantation for NICM and NS-VT receive appropriate ICD therapy at a greater rate than those implanted for ICM, NS-VT, and a positive EPS. Although these data do not prove survival benefit in NICM, they suggest a beneficial effect. PMID- 15273457 TI - T wave oversensing in implantable cardioverter defibrillators. AB - The implantable cardioverter defibrillator is the treatment of choice for patients with ventricular tachycardia, especially in the setting of structural heart disease, but inappropriate therapy continues to be a problem. In this report we describe a short case series of patients who presented with T wave oversensing. We propose a classification in 2 categories, T wave oversensing occuring in the setting of persistently low R wave amplitude, and T wave oversensing occuring in the setting of transiently low R wave amplitude. PMID- 15273458 TI - Inappropriate single chamber ICD discharges due to supraventricular tachycardia with high degree atrioventricular block. AB - Supraventricular tachycardia with rapid ventricular response is well recognized as the more frequent cause of single chamber ICD inappropriate therapies. We report here a 18-year-old-woman with surgically corrected transposition of the great arteries who received repetitive inappropriate discharges from an ICD implanted for ventricular tachycardia. Rapid atrial activity during episodes of supraventricular tachycardia with high degree atrioventricular block was oversensed as ventricular fibrillation by a single chamber ICD causing repetitive painful discharges. Pharmacological treatment of the supraventricular tachycardia solved the problem. PMID- 15273459 TI - Obituary: a giant of modern clinical electrophysiology: Philippe Coumel (1935 2004). PMID- 15273460 TI - Management of severe burn injuries in neonates. AB - Although rare, burns suffered by neonates can be fatal. Many complex difficulties are faced during the management of burns in neonates because of the neonate's complex physiological and pathological changes. We compiled a retrospective review from the treatment of four burned neonates (including a premature neonate). All four neonates suffered bath-related burns in the hospital as a result of careless nursing when being bathed. The total body surface area burned ranged from 1 to 60% in these patients, and all survived the burn injury. All the patients were treated in the Burn Intensive Care Unit with close co-operation of burn surgeons and neonatologists. Based on our experience as well as a review of literature, management recommendations are proposed as the following: 1) prompt and aggressive fluid resuscitation, 2) early administration of oxygen and keeping the patient warm, 3) application of specific biological dressing and recombinant human growth hormone if necessary, 4) establishment of a multidisciplinary team, and 5) removal of necrosis tissue early and aggressively. Furthermore, a very important issue is also discussed, which is about the prevention of newborn burns in the neonate unit in developing countries. PMID- 15273461 TI - Hot beverage scalds in Australian children. AB - Our objective was to compile data on the mechanism and severity of injuries associated with hot beverage burns in children. We identified 152 children over a 3-year period who attended a tertiary level burns center, representing 18% of all children treated. Their median age was 17.5 months and median body surface area burned was 4% (range, 0.25% to 32%). Significantly, 52% of children required admission, 18% received a split skin graft, and 26% required long-term scar management. In 70% of all cases, the mechanism of injury was the child pulling the hot beverage over himself or herself. In 80% of incidents, a primary care giver witnessed the injury. These findings indicate that scalding from hot beverages carries significant morbidity and is an important pediatric public health issue. It is clear that further research towards effective education programs for primary caregivers is warranted. PMID- 15273462 TI - Personality characteristics and perceived health problems after burn injury. AB - The relationship between personality traits and the perceived outcome of burn injury 1 to 18 years (mean, 9.2 years) after severe burn injury was evaluated in 166 individuals treated at the Uppsala Burn Unit. The perceived outcome was measured with the Burn Specific Health Scale-Brief (BSHS-B) and was related to personality traits evaluated by means of the Swedish universities Scales of Personality. After controlling for age at inquiry, time since injury, burn area, and sex, a stepwise logistic regression analysis revealed an association between the Swedish universities Scales of Personality domain Neuroticism and Bad outcome in all BSHS-B domains, both psychosocial and physical, and Insufficient outcome in the domains Work, Body image, Affect, and BSHS-B total score. The neurotic traits Somatic trait anxiety, Psychic trait anxiety, Stress susceptibility, Embitterment, and Mistrust each or in different combinations explained the observed relationships. The data suggest that personality is related to health status because it is perceived a long time after severe burn injury and that its effect is not confined only to psychological but also to physical aspects of life. PMID- 15273463 TI - Effect of transcutaneous electrical nerve stimulation for the management of burn pruritus: a pilot study. PMID- 15273464 TI - Sentinel lymph node biopsy identifies occult nodal metastases in patients with Marjolin's ulcer. AB - Since Marjolin's description, the management of burn scar carcinoma has remained controversial. A multitude of options and recommendations exist for the management of both primary lesions and regional nodal metastasis. This work reviews six cases of Marjolin's ulcer staged using sentinel lymph node biopsy. All primary lesions were confirmed to be squamous cell carcinoma and occurred a median of 29.5 years after burn. No patient had clinically detectable lymphadenopathy. In all cases, preoperative lymphoscintigraphy successfully identified a single draining regional nodal basin. Subsequent intraoperative lymphatic mapping/sentinel lymph node (SLN) biopsy was successful in five of six cases (83%). A successful intraoperative lymphatic mapping/SLN biopsy was defined as the identification of blue (uptake of isosulfan blue dye) or "hot" (uptake of radiolabeled sulfur colloid as measured with a handheld gamma counter) node(s) and subsequent excision. Four of five SLN biopsies identified previously occult nodal metastasis. SLN biopsy represents a minimally invasive and accurate staging procedure for Marjolin's ulcer. PMID- 15273465 TI - Intravenous immunoglobulin does not improve outcome in toxic epidermal necrolysis. AB - Intravenous Immunoglobulin (IVIG) has been proposed as a beneficial therapy for toxic epidermal necrolysis (TEN). However, this has been based on a limited amount of Class 5 evidence. To compare outcomes in TEN patients treated in our burn unit since 1999, when we began to use IVIG (IG group), with TEN patients treated between 1995 and 1999 who did not receive IVIG (control group). Retrospective cohort review of the records of all TEN patients admitted between April 5, 1995 and December 4, 2002. There were 16 patients in the IG group (age 53 +/- 21 years, with initial rash involving 65 +/- 29% TBSA) and 16 patients in the control group (age 52 +/- 20 years, with initial rash involving 65 +/- 27% TBSA). The IG group received 0.7 +/- 0.2 g/kg/day of IVIG for 4 +/- 1 days. There were no significant differences between the groups with respect to the length of stay, duration of mechanical ventilation, severity of systemic inflammatory response syndrome and multiple organ dysfunction syndrome, or the incidence of sepsis. Significant progression of the wound occurred in 13% of the IG patients and in 27% of control patients, whereas no wound progression was observed in 47% of the IG patients and in 18% of the control patients (P =.299). The time to healing did not differ between IG and control groups (11.2 +/- 3.6 vs 11.4 +/- 2.6 days, respectively). There was no significant difference in the mortality rate between the IG group (25%) and the control group (38%). There were no complications from IVIG aside from one case of hyponatremia from the hypotonic IVIG solution. Although there may have been a trend towards less severe wound progression in patients who received IVIG, this was not associated with any substantial improvement in outcome in our TEN patients. A prospective randomized study with a larger sample size is needed to confirm our findings. PMID- 15273466 TI - Evaluation of a pediatric scald burn clinical pathway. AB - Scald burns make up more than 75% of the pediatric burns patients who are admitted to our burns unit. A pediatric scald burn pathway was implemented at our center in November 1999, the aim of which was to improve consistency in the management of the acute phase of injury. This study assessed the effectiveness of the first 18 months of this scald burn pathway. Aspects of the management of pediatric scald burn patients were reviewed. This included all the patients admitted over a 6-month period 1 year before the implementation of the pathway and the first 6 months and between 12 and 18 months after the implementation of the pathway. Data collected included patient demographics, total body surface area burned, and key management guidelines, including nasogastric feeding, intravenous cannulation, analgesia prescription, multidisciplinary referrals, and family education Thirty-seven patients were enrolled in the first 6 months of the pathway's use. Between 12 and 18 months, a further 38 patients were enrolled. The patients within the three groups were similar in age and burn size. A comparison among the groups with respect to compliance with the treatment guidelines is presented graphically. In conclusion, the management of pediatric scald burns in the acute phase is more consistent since the implementation of the clinical pathway. PMID- 15273467 TI - Favorable short- and long-term outcomes of prolonged translaryngeal intubation in critically ill children. AB - In those children who require protracted mechanical ventilation, we use long-term intubation in order to avoid the consequences of tracheostomy in young children. A retrospective 9-year review was performed to document the efficacy and safety of this practice. A retrospective review of children admitted from January 1, 1991, to December 31, 1998, who required mechanical ventilatory support for at least 7 consecutive days was performed. Data are presented as mean +/- standard deviation. There were 98 children, ventilated for a total of 1967 days, who satisfied review criteria. They had an average age of 6.1 +/- 5.3 years (range, 3 months to 17 years) a total body surface area burn of 53 +/- 25% (range, 0-100%), and 71 of 98 (72%) had suffered an inhalation injury. They were ventilated for 19.7 +/- 16.8 days (range, 7-92 days) and were hospitalized for 67.8 +/- 48.9 days (range, 9-211 days). Ninety-three percent (91 of 98) of the patients were maintained on morphine infusions at a mean hourly rate of 0.35 +/- 0.33 mg/kg/hr (range, 0.01-4.38) and 78% (76 of 98) on midazolam infusions at a mean hourly rate of 0.14 +/- 0.17 mg/kg/hr (range, 0.01-1.82). Neuromuscular blocking agents were administered in 39% (38 of 98) of patients during all or part of 355 (18%) of the 1967 ventilator days. Patients were ventilated with an oral endotracheal (ET) tube in 82% of ventilator days and nasal ET tube in 18% of ventilator days. Two patients (2%) required tracheostomies for long-term management, and five patients (5.1%) died during the study period unrelated to airway issues. There were five unplanned extubation events, for an incidence rate of 2.54 per 1000 ventilator days. All patients were reintubated successfully. Thirteen ET tubes needed to be changed for issues such as leaking cuffs. Patients were followed up for a mean of 2.99 +/- 2.24 years (range, 1 month to 8 years). Possible sequelae related to prolonged intubation were noted in follow-up visits in 8 patients, including sinusitis (one; resolved without treatment), subglottic stenosis (one; required reconstructive surgery), persistent cough (three; all resolved spontaneously), occipital breakdown because of ET ties (one; healed after 1 month), soft voice (two; resolved spontaneously), and decreased pharyngeal sensation (one; resolved without treatment). Translaryngeal intubation is a safe and effective method to provide long-term ventilatory support in children. PMID- 15273468 TI - A review of keratinocyte delivery to the wound bed. AB - Over the last 20 years, confluent sheets of cultured epithelial autograft have been used for patients with major burns. Problems with the lack of "take" and long-term durability, as well as the time delay to produce such grafts, have led to the development of delivery systems to transfer keratinocytes to the wound bed. This review article describes the problems of using cultured epithelial autograft and the advantages of using preconfluent keratinocytes. Despite the numerous delivery systems that have been reported, most studies are limited to animal wound bed models. There are a few small clinical studies that have demonstrated enhanced healing using mainly subjective methods. There is a need for controlled, randomized clinical trials to prove the efficacy of keratinocyte delivery systems. Proposals for the use of this technology are made. PMID- 15273469 TI - Glycosaminoglycan hydrogels as supplemental wound dressings for donor sites. AB - Chemically crosslinked glycosaminoglycan (GAG) hydrogel films were evaluated as biointeractive dressings in a porcine model for donor-site autograft wounds. Multiple 5 x 5 x 0.03 cm wounds were created on the dorsum of pigs. Half of the wounds were treated with a GAG film plus an occlusive dressing (Tegaderm), whereas the other half were treated with Tegaderm alone. At 3, 5, or 7 days after surgery, the partially healed wounds were excised and evaluated histologically for three animals at each time point. By day 3, epithelial cells had proliferated and migrated from wound edges and from epithelial islands associated with residual hair follicles to begin to cover the wound bed. A statistically significant increase in coverage was observed for GAG + Tegaderm-dressed wounds than for those with Tegaderm alone at day 3 and day 5 post-surgery. By day 7, all treatment groups were completely healed. Thus, GAG hydrogels accelerated wound healing by enhancing re-epithelialization. PMID- 15273470 TI - Effects of exogenous growth hormone on resting pulmonary function in children with thermal injury. AB - Burned children living beyond the acute phase of injury often have extensive physical functional limitations, such as impaired spirometry pulmonary function (PF). In patients with both lung disease and nutritional compromise, such as cystic fibrosis, studies suggest that growth hormone (GH) therapy improves PF. However, whether GH will improve PF in burned children is presently unknown. We therefore evaluated whether GH administration of 0.05 mg/kg/day for 1 year would improve PF in burned children. Thirty children, aged 7 to 18, with a 40% or more total body surface area burned were randomized into two groups and studied. One group received GH (n = 17) and the other received saline (n = 13). No differences were noted at hospital discharge between groups in age, % total body surface area, height, and weight. At 12 months after burn, both groups had similar height and weight. Baseline PF were below normal in both groups, but no statistical differences were noted between groups. At 1 year, there was a significant increase in PF in both groups; however, this increase in PF was similar in both groups. We conclude that the response in PF in burned children from the administration of GH prescribed for up to 1 year is limited. PMID- 15273471 TI - Treating sleep problems in patients with burn injuries: practical considerations. AB - Sleep disorders are a frequent but under-addressed complication of burn injuries. Burn injuries can potentially disrupt sleep for a variety of reasons, including the physiological effects of trauma as well as ramifications of treatment (ie, intensive care unit environment, pain, itching, medications). The literature on sleep disorders and burn injuries is reviewed, and suggestions for treatment are provided. Treatment is divided into two major types: nonpharmacologic and pharmacologic. Nonpharmacologic treatment, also referred to as behavioral techniques, may include any one or combination of the following: sleep hygiene, stimulus control, sleep restriction, relaxation techniques, cognitive, and light therapy. Pharmacologic therapies may include hypnotics (benzodiazepine, nonbenzodiazepine, or benzodiazepine receptor agonists), antidepressants, over the-counter preparations, hormone replacement therapy, herbs, and melatonin. PMID- 15273472 TI - Serum levels of prolactin, growth hormone, and cortisol in burn patients: correlations with severity of burn, serum cytokine levels, and fatality. AB - In this study, we measured serum prolactin (PRL), cortisol, growth hormone, interleukin (IL)-1beta, IL-6, IL-8, IL-10, IL-12, and tumor necrosis factor-alpha in patients admitted with small-to-moderate burn injuries. Serum samples were obtained at the time of admission from 49 adult male burn patients with ages ranging from 18 to 91 years and TBSA ranging from 0.001 to 60%. The levels of serum PRL, IL-8, IL-6, and IL-1beta correlated positively with the TBSA, whereas only serum IL-8 levels correlated positively with fatality. Each of these factors were increased at least 2-fold at the higher burn severity. Not surprisingly, there was a large degree of variability in the hormone and cytokine levels in this patient population, which presumably reflects individual levels of stress, as well as other physiological variables. We also studied relationships between serum hormone levels and serum cytokine levels in this context. Linear regression analysis revealed a significant positive correlation between the serum PRL level and the levels of IL-10, IL-6, and IL-8. These results indicate that PRL responds to burn injury at early time points and that a subset of cytokines are involved in the early response to burn injury. PMID- 15273473 TI - Burns from illegal drug manufacture: case series and management. AB - This case series presents our experience with burns sustained while manufacturing illegal drugs. All adult burn admissions in an 18-month period were retrospectively reviewed. All patients suspected of sustaining burns from illegal drug manufacture were contacted. Information regarding the burn mechanism was sought. Nine of the 64 adult burn admissions were caused by explosions during the manufacture of cannabis oil. Young males with hand and face burns were heavily represented. First-aid treatment was often ignored in favor of hiding incriminating evidence. Only two patients gave honest admission histories. Illegal drug manufacture is becoming more common as synthetic drugs become more consumer desirable. Burns sustained may be thermal and/or chemical. Dishonest patient histories negatively influence burn management. A high level of suspicion is required for diagnosing and treating burns from illegal drug manufacture. Public education is unlikely to be effective as the financial rewards outweigh the perceived risks. PMID- 15273474 TI - Postburn subtalar dislocation in a major foot contracture. AB - An unusual postburn contracture, which consists of calcaneal bone dislocation and severe dorsiflexion contracture of foot and toes, is presented. To our knowledge, subtalar dislocation as the result of postburn contracture has not been previously reported in the literature. PMID- 15273475 TI - Burn Education Awareness Recognition and Support (BEARS): a community-based juvenile firesetters assessment and treatment program. AB - In response to the continued staggering statistics of fires set by juveniles and the devastating personal and property costs that are associated with these fires, the Burn and Shock Trauma Institute of Loyola University Medical Center, in collaboration with the State Fire Marshal's Office; the Illinois Fire Safety Alliance; and representatives from the firefighting community, law enforcement, emergency medicine and mental health, came together to create the Burn Education Awareness Recognition and Support Program. Through financial grant support from the International Association of Firefighters, the Illinois Fire Safety Alliance, and other private donations, the Burn Education Awareness Recognition and Support Program is able to provide a free resource to anyone who is concerned about a child playing with fire. Specially trained firefighters assess each child using the tool developed by the Federal Emergency Management Agency. In 2002, we assessed 42 children; 29 of those children were referred through the courts. So far, none of the children treated in our program have returned to fire-setting behaviors. PMID- 15273476 TI - Quantification and risk analysis of occupational burns: Oregon workers' compensation claims, 1990 to 1997. AB - This study examined all accepted Oregon workers' compensation claims for occupational burn injuries during the period of 1990 to 1997 (N = 3,158). The Current Population Survey was used to derive employee population baselines for establishing rate estimates. It was estimated that the average occupational burn claim rate was 2.89 per 10,000 workers (95% confidence interval [CI] 2.76, 3.02). The majority of claimants (71.7%) were males, the largest proportion (32.6%) was aged 25 years or less, and almost half (48.7%) had less than 1 year of job tenure. The most frequent burn type cited was heat/scald burns (78.9%) followed by chemical burns (19.3%). Costs averaged over 1.6 million dollars annually. The average indemnity period was 16 days. Higher relative risks were found for evening workers (2.97, 95% CI 2.96, 2.98) and night workers (2.13, 95% CI 2.12, 2.13) compared with dayshift workers. Kitchen workers had the highest burn rate of all occupations, with 62.5 claims per 10,000 workers. PMID- 15273477 TI - Posttraumatic stress disorder and the intensive care unit patient: implications for staff and advanced practice critical care nurses. AB - Posttraumatic Stress Disorder (PTSD) is a rather common psychiatric diagnosis, and potentially is a very debilitating disorder. In PTSD, patients exhibit specific debilitating symptoms in response to exposure to an extreme stressor. Conditions in the intensive care unit (ICU) can exacerbate previously diagnosed newly developed PTSD, and in some cases cause PTSD. This diagnosis potentially puts both the patient and nursing staff at increased risk for harm, and is associated with increased utilization of medical services. Critical care staff and APNs can take actions to screen for at-risk patients, emplace safety protocols, and advocate for affected patients within the healthcare team. PMID- 15273479 TI - Ludwig's angina. AB - Ludwig's angina is a serious, often fatal infectious disease process that requires prompt intervention of life-saving therapies. The critical care nurses caring for a patient with Ludwig's angina must be able to recognize subtle changes in the patient's status and intervene quickly to prevent death by airway edema or profound sepsis. PMID- 15273480 TI - A review of rhabdomyolysis. AB - Rhabdomyolysis can be a life-threatening disease if not treated immediately. Once the process occurs, several complications both short-term and long-term can develop. The purpose of this article is to educate all types of nurses (particularly critical care nurses because of the life-threatening complications) about the disease process and treatment of rhabdomyolysis. PMID- 15273481 TI - Spirituality as a coping mechanism. AB - Spirituality as a coping mechanism can be observed to be a powerful resource in the provision of comfort, peace, and resolution for patients confronted with critical illness. While the exact machinery of spirituality in adaptation and adjustment to illness is enigmatic, the complementary benefits are clearly illustrated in the analysis of recounted personal experiences. Analysis of interactions with patients living the experience of coping with critical illness provides nurses with a means of reflection and transformational learning which improves and preserves the spiritual heritage of nursing care. PMID- 15273483 TI - Turning the tide on medical errors in intensive care units: a human factors approach. AB - Errors occur in all nursing settings. The current healthcare climate tends to focus on individuals as the cause of errors rather than addressing issues that may be inherently wrong with the healthcare system that predisposes the individual to make errors. Human factors engineering (HFE), which is focused on removing human factors as much as possible from errors, has the potential to greatly impact medical errors in intensive care units. Applied in other high-risk industries, HFE has been critical in understanding and preventing errors at a systems level. Knowledge concerning the role systems play in errors and improvements to medical systems using HFE is intended to empower nurses to be advocates for systems change, resulting in a safer work environment and a safer healthcare delivery system. PMID- 15273485 TI - A personal reflection: the intimacy of dying: an act of presence. AB - The focus of critical care education is usually on the development of knowledge and skills needed to manage care for the critically ill patient. In today's intensive care units, what is also needed is the education and development of the skills that enable nurses the ability to enter into a relationship with the patients and families that touch the spirit, and provide another dimension to healing. PMID- 15273487 TI - Clinical research and the development of new drugs: issues for nurses. AB - Critical care nurses are frequently exposed to clinical trials for either new medications, equipment, or procedures. This article discusses the FDA regulatory requirements as well as the guidelines for conducting human research involving investigational new drugs. The role of the critical care nurse caring for patients enrolled in clinical trials is also discussed. PMID- 15273491 TI - New strategies for cervical cancer screening in adolescents. AB - PURPOSE OF REVIEW: This paper reviews the epidemiology and natural history of human papillomavirus (HPV) infection in young women, the development of new technologies for cervical cytology screening, proper cervical cytology sampling technique, the new Bethesda system for reporting cervical cytology findings, and several recent professional society guidelines for cervical cytology screening and management of cytologic abnormalities in adolescents. RECENT FINDINGS: Natural history studies of HPV infection in healthy young women show that infection is quite prevalent, but is generally transient. New and sensitive technologies such as HPV DNA testing and liquid-based cytology are more likely to detect cytologic abnormalities in young women who are at low risk for actual invasive cervical disease. This sensitivity potentially places adolescents at risk for increased anxiety, testing, and intervention. The multi-center ASCUS LSIL Triage Study has shown that HPV DNA testing can be used safely to minimize intervention in many cases. SUMMARY: HPV infection is common in young women, but rarely progresses to invasive cervical disease. Providers need to inform themselves about new professional society guidelines that suggest delaying initiation of cervical cancer screening to within 3 years of onset of sexual activity. Given the idiosyncrasies of this population, the authors counsel using clinical discretion when applying these guidelines to individual teenagers. In light of the extremely low likelihood of invasive disease in this age group, providers must separate the provision of contraceptive services and sexually transmitted disease screening from requirements for cervical cancer screening. PMID- 15273492 TI - Fainting freshmen and sinking sophomores: cardiovascular issues of the adolescent. AB - PURPOSE OF REVIEW: Syncope is a common symptom in adolescents. The vast majority of cases are the result of benign neurocardiogenic syncope, without associated risk of sudden death. This paper reviews the mainstays of diagnosis and treatment for syncopal episodes, differentiation of syncope from life-threatening arrhythmia and aborted sudden cardiac death, and the patient populations at highest risk for cardiac symptoms and cardiac disease. RECENT FINDINGS: A detailed history (including past medical history and family history that focus on cardiac disease) combined with dynamic physical examination and electrocardiogram identifies the vast majority of adolescents with significant heart disease. Further diagnostic modalities have limited utility. Reassurance and supportive measures remain the treatment of choice, although drug therapy can sometimes be helpful, even if data are limited. Divergent approaches to the screening of the young competitive athlete exist. Particular attention is required in adolescents and young adults with exercise-associated syncope, eating disorders, chronic fatigue syndrome, or history of congenital heart disease. Their symptoms may be either more serious or challenging to manage. SUMMARY: Syncope in the adolescent patient is very common; true cardiac disease is not. The traditional diagnostic screen of history and physical combined with an electrocardiogram will identify the overwhelming majority of patients with significant disease. Patients with abnormalities on this initial office evaluation, history of cardiac disease, or complicating medical illness may benefit from referral to a cardiologist. Even within this patient subset, many will prove to have benign disease. PMID- 15273493 TI - Cholesterol (and cardiovascular risk) in adolescence. AB - PURPOSE OF REVIEW: Atherosclerosis is a significant cause of adult morbidity and mortality. New evidence confirms that it begins in childhood and accelerates at adolescence. Many new studies have solidified the understanding of its risk factors and changed the approach to their clinical management. RECENT FINDINGS: The important descriptions of known risk factors and their clear relation to the pathophysiologic process of atherosclerosis are reviewed here. Some emerging risk factors are discussed. Furthermore, the many new studies on treatment options are reviewed. SUMMARY: These new findings make it imperative that adolescents be screened for traditional risk factors and that they be treated with therapeutic lifestyle change. With the weight of new evidence on the safety and efficacy of statins in this age group, clinicians can feel more confident in their use when indicated. Research into emerging risk factors like obesity, metabolic syndrome, diabetes. and homocysteine will be important in the future management. PMID- 15273494 TI - Controversies in male adolescent health: varicocele, circumcision, and testicular self-examination. AB - PURPOSE OF REVIEW: The authors review three common clinical controversies encountered by primary care providers of adolescent males: management of varicoceles, the role of circumcision in the acquisition and transmission of sexually transmitted infections, and the value of teaching testicular self examination. RECENT FINDINGS: Recent findings in adolescent varicoceles have advanced knowledge regarding the cause of varicoceles, the mechanism by which they may lead to infertility, new screening methods, and optimal surgical management. Accumulating evidence shows circumcision to be protective against acquisition and transmission of sexually transmitted infections, and preliminary work also indicates the potential for protection against the spread of AIDS in Africa. Testicular self-examination remains an unproven screening modality that is suboptimally performed by at-risk patients. SUMMARY: This review updates the provider on these topics and clarifies issues involved in these controversies PMID- 15273495 TI - Adolescent smoking cessation. AB - PURPOSE OF REVIEW: Approximately 25% of high school students report current cigarette use, 85% of adolescents think about quitting, and around 80% of current smokers made a quit attempt in the past year. This review analyzes recent additions to the adolescent smoking cessation literature from June 1, 2003 to May 1, 2003. RECENT FINDINGS: Adolescent attitudes toward smoking cessation are largely affected by their smoking history. Youth cessation interventions largely focus on behavioral interventions, and research concerning these interventions has yielded mixed results. Little data exist about the effectiveness of nicotine replacement therapy in adolescents, but there is growing evidence that youth use this pharmacotherapy. Recent research has explored the use of nicotine replacement therapy as an adjunct for enhanced smoking reduction in adults, and future research may focus on this tactic for youth as well. Internet cessation adjuncts and telephone quit lines also serve as future frontiers for adolescent smoking cessation research. SUMMARY: Information concerning adolescent smoking behaviors, effective interventions, and smoking cessation therapy continue to grow and provide data that improve our understanding of adolescent smoking cessation. Although we cannot directly extrapolate the adult findings to this population, adult cessation research continues to inform future adolescent cessation efforts. PMID- 15273496 TI - Update on hemangiomas of infancy. AB - PURPOSE OF REVIEW: Recent clinical and basic science research has led to advances in the understanding of hemangiomas of infancy. RECENT FINDINGS: New developments include (1) the establishment of a relation between hemangiomas of infancy and placental tissue, (2) the discovery of unique immunohistochemical markers for hemangiomas of infancy, (3) the importance of morphology and location in determining potential risk for underlying complications, and (4) the discovery of becaplermin 1% gel as an effective therapy for refractory ulceration. SUMMARY: The morphology and location of a hemangioma of infancy are critically important factors in determining potential risk for complications. Ongoing research is bringing closer an understanding of the cause of hemangioma, which will provide opportunities for the development of interventional, and ultimately preventative, therapies. PMID- 15273497 TI - Sunscreens and insect repellents. PMID- 15273498 TI - Acne update: 2004. AB - PURPOSE OF REVIEW: Acne vulgaris is a common skin disorder among children and young adults that carries enormous financial and psychosocial impact. Contemporary therapies attempt to address factors underlying acne as a disorder of the pilosebaceous unit. These longstanding paradigms regarding pathogenesis and treatment continue to evolve in light of recent work on this ubiquitous disease. RECENT FINDINGS: This review focuses on new literature that has emerged regarding the biology of the folliculosebaceous unit, the identification of particular mediators responsible for inflammatory acne, the use of topical and systemic retinoids in acne therapy, and approaches to address the emergence of antibiotic-resistant Propionibacterium acnes strains. In addition, the use of several novel therapeutic avenues is discussed, including combination therapies, lipoxygenase inhibitors, and lasers. SUMMARY: As the understanding of the factors that initiate and exacerbate acne vulgaris continues to increase, so does the diversity of therapeutic options. Rational use of available treatment options based on the type and severity of acne lesions is a key component of successful acne therapy and allows the physician who treats adolescents with acne to provide optimum care. PMID- 15273499 TI - Lumps and bumps in children-when to worry: recent trends in recognition and pathology of hemangiomas of infancy and Spitz nevi. AB - PURPOSE OF REVIEW: Hemangiomas are common, and most lesions are benign. Careful attention needs to be paid to these lesions; however, because their growth can be unpredictable, they can have important complications and can serve as markers of underlying malformations. The consequences of hemangiomas are extensive and can be challenging to recognize. RECENT FINDINGS: Recent knowledge of the pathogenesis and diagnosis of these lesions has helped raise awareness of their classification and clinical significance. Spitz nevi can also be a challenging lesion to diagnose and treat, although for different reasons. These are lesions composed of melanocytes with clinical and histopathological findings that we easily misdiagnosed and can have significant implications for patient care. SUMMARY: Both of these important lesions in infants and children are reviewed in further detail with particular attention to recent trends. PMID- 15273500 TI - Atopic dermatitis update. AB - PURPOSE OF REVIEW: Atopic dermatitis, one of the most common chronic illnesses of childhood, is encountered routinely by all providers of health care to children. RECENT FINDINGS: In recent years there has been a dramatic rise in the prevalence of atopic dermatitis and therefore a rapid increase in the number of studies investigating various aspects of the disease. Consequently, hundreds of publications are released each year, and it is difficult to stay up to date on the latest advances. SUMMARY: This review will examine and summarize recent literature on the diagnosis, epidemiology, pathogenesis, treatment, and complications of atopic dermatitis. PMID- 15273501 TI - Human papillomavirus infections in children. AB - PURPOSE OF REVIEW: The human papillomavirus is a ubiquitous 55-nm DNA virus that causes a variety of clinical disease states in children, commonly referred to as warts. The natural history of warts is spontaneous regression through the development of a complex blend of cell-mediated and humoral immunity. Although spontaneous immunity can develop, as many as one third of children will have persistent human papillomavirus infection beyond 2 years. Therapeutic modalities are manifold, primarily because no therapy is universally effective. The purpose of this review is to update the reader with the latest information on the human papillomavirus and its therapeutics in children. RECENT FINDINGS: Recently, encouraging research has been conducted in human papillomavirus, including destructive and immunologic therapies. Vaccines tailored to genital human papillomavirus strains are just coming into clinical use. SUMMARY: Manipulation of the immune system through medications or vaccination will likely help contain human papillomavirus in the future and prevent secondary human papillomavirus oncogenesis of the skin and cervix. PMID- 15273502 TI - What's new in pediatric dermatology: update for the pediatrician. AB - PURPOSE OF REVIEW: Common pediatric skin conditions such as infantile atopic dermatitis, vitiligo, hemangiomas of infancy, warts, and molluscum contagiosum do not always respond to standard therapy. In some settings pediatricians will use "off-label" medications if the benefit-to-risk ratio is favorable. This article reviews important literature from the past year related to "off-label" immune based treatment of skin disease, using the topical immunomodulators tacrolimus, pimecrolimus, and imiquimod, as well as intravenous Ig. RECENT FINDINGS: The topical immunomodulators tacrolimus and pimecrolimus have been embraced by pediatricians as long awaited alternatives for treating atopic dermatitis in children 2 years of age and older. Their unique appeal as nonsteroidal topical agents with good safety profiles has led to their frequent use for unapproved indications. A number of recent publications detail their use in infantile atopic dermatitis in children as young as 3 months of age, as well as use in other conditions such as vitiligo. Imiquimod, another topical immunomodulator, approved for genital wart treatment in adults, has also been examined for "off-label" pediatric use in nongenital warts, molluscum contagiosum, hemangiomas of infancy, and basal cell carcinoma. Finally, "off-label" use of intravenous Ig has been evaluated for the life-threatening dermatoses Stevens-Johnson syndrome and toxic epidermal necrolysis. SUMMARY: In the absence of larger controlled trials, pediatricians must consider the cumulative weight of smaller studies with their personal experience when assessing any role for "off label" therapy. The recent literature reviewed herein will facilitate such assessments of the non-steroid topical immune modifiers tacrolimus, pimecrolimus, and imiquimod as well as intravenous immunoglobulin. PMID- 15273504 TI - Human disorders of ubiquitination and proteasomal degradation. AB - PURPOSE OF REVIEW: The goal of this review is to provide an overview of rapidly evolving information on a new group of genetic inborn errors affecting ubiquitination and proteasomal degradation of proteins and to suggest a classification scheme for these disorders. The relevant genes encode ubiquitin, ubiquitin enzymes (E1 and many E2s and E3s), deubiquitinating enzymes, proteasomal subunits, and substrates undergoing ubiquitination. RECENT FINDINGS: Since the initial recognition that Angelman syndrome is caused by maternal deficiency of the E6-AP ubiquitin E3 ligase (gene symbol UBE3A), several. other disorders of E3 ligases have been identified, including autosomal recessive juvenile Parkinson disease, the APECED form of autoimmune polyendocrinopathy syndrome, von Hippel-Lindau syndrome, and congenital polycythemia. Disorders that disturb ubiquitin regulatory signaling include at least two subtypes of Fanconi anemia, the BRCA1 and BRCA2 forms of breast and ovarian cancer susceptibility, incontinentia pigmenti, and cylindromatosis. Many disorders affect ubiquitin pathways secondarily. SUMMARY: The authors propose both a genetic and a functional classification for disorders of ubiquitination and proteasomal degradation, as follows. Genetic classes include mutations in (1) the UBB ubiquitin gene; (2) enzymes of ubiquitination including E1, E2, E3, and related proteins; (3) deubiquitinases; (4) proteasomal subunits; and (5) substrates of ubiquitination. Functional classes include defects in (1) proteolytic degradation, (2) ubiquitin signaling, and (3) subcellular localization of substrates. Additional functional classes are likely to be defined, and individual disorders may involve multiple functional defects. PMID- 15273505 TI - Recent developments and new applications of tandem mass spectrometry in newborn screening. AB - PURPOSE OF REVIEW: To summarize recent developments in the field of newborn screening related to the use of tandem mass spectrometry as an analytic platform. RECENT FINDINGS: Novel inborn errors of metabolism with informative amino acid and/or acylcarnitine profiles have been characterized, increasing the complexity of the differential diagnosis of abnormal results. In addition, methods have been developed for the analysis in dried blood spots of steroids and lysosomal enzymes. Previously unrecognized genotype/phenotype correlations have been found among cohorts of patients whose conditions were diagnosed by screening rather than clinically. Several government entities and professional organizations have issued position statements on newborn screening, and worldwide outcome studies continue to underscore the clinical and financial benefits of expanded newborn screening. SUMMARY: Although it is done inconsistently, newborn screening in the United States is undergoing a rapid expansion driven by the introduction of tandem mass spectrometry in at least 34 state programs. This technology is also used to detect disease markers beyond acylcarnitines and amino acids, as both primary and second-tier tests. In addition to analytic improvements, there is a trend toward the development of joint programs not limited to contiguous geographic areas, often based upon public-private partnerships. This review will summarize several new developments in the field that have occurred since early 2003 and will mention others likely to occur in the near future. PMID- 15273506 TI - Congenital disorders of glycosylation: a booming chapter of pediatrics. AB - PURPOSE OF REVIEW: The detection and identification of new congenital disorders of glycosylation continues at a rapid pace. Sine June 2003, four new congenital disorders of glycosylation have been reported, making a total of 20 diseases (on average nearly 1 disease per year since the first report in 1980; 12 of these congenital disorders of glycosylation were identified in the past 6 years). RECENT FINDINGS: Three of these newly discovered CDG are caused by defects in early steps of dolichol-linked oligosaccharide biosynthesis. Affected patients have a neurologic or a multisystem disease. The fourth new CDG is a completely new CDG type caused by a defect in an endoplasmic reticulum-Golgi shuttle protein carrying multiple glycosyltransferases and nucleotide-sugar transporters. SUMMARY: Disorders of nearly all organs and systems have been reported and continue to be reported in congenital disorders of glycosylation. Therefore, it is strongly recommended that congenital disorders of glycosylation be considered in any child with an unexplained clinical syndrome. PMID- 15273507 TI - Systemic effects of inhaled corticosteroids in children. AB - PURPOSE OF REVIEW: Inhaled corticosteroids (ICS) are first-line therapy for persistent asthma in children. Major safety concerns about long-term ICS therapy include suppression of adrenal function, growth, and bone development. Proper interpretation of ICS safety studies requires knowledge of differences between various ICS drug/delivery device systems. RECENT FINDINGS: Dosage, type of inhaler device used, patient technique, and characteristics of the individual drug influence systemic effects of ICS. Reports of adrenal insufficiency occur but are rare and are confined to children receiving high doses of ICS. Dose related inhibition of growth is detectable as ICS dosage increases, but appears temporary, more pronounced in childhood, and is not associated with reduction in final height. Moderate-dose ICS therapy is not associated with significant changes in measurements of bone density, but more studies of high doses and of therapy in adolescents are needed. SUMMARY: Recent studies confirm that benefits of ICS, properly prescribed and used, clearly outweigh not only their potential adverse effects but also the risks associated with poorly controlled asthma. PMID- 15273508 TI - Celiac disease as a cause of growth retardation in childhood. AB - PURPOSE OF REVIEW: Celiac disease is a syndrome characterized by damage of the small intestinal mucosa caused by the gluten fraction of wheat proteins and similar alcohol-soluble proteins (prolamines) of barley and rye in genetically susceptible subjects. The presence of gluten in these subjects leads to a self- perpetuating mucosal damage, and the elimination of gluten results in full mucosal recovery. The clinical manifestations of celiac disease are protean in nature and vary markedly with the age of the patient, the duration and extent of disease, and the presence of extraintestinal pathologic changes. In addition to the classic gastrointestinal form, a variety of other clinical manifestations of the disease have been described, including atypical and asymptomatic forms. Although the typical form of celiac disease, characterized by failure to thrive, is still the most frequent presentation in the pediatric age group, severe growth delay is less commonly seen in developed countries. RECENT FINDINGS: Recent epidemiologic studies suggest that celiac disease-associated growth retardation is becoming a tangible health problem in developing countries, where the problem has been historically overlooked. Given the protean nature of the clinical presentation of celiac disease, the diagnosis is extremely challenging and relies on a sensitive and specific algorithm that allows the identification of different manifestations of the disease. Serologic tests developed in the past decade provide a noninvasive tool for screening individuals at risk for the disease as well as the general population. SUMMARY: The current gold standard for the diagnosis of celiac disease remains histologic confirmation of the intestinal damage in serologically positive individuals. The keystone treatment of celiac disease patients is a lifelong elimination diet in which food products containing gluten are avoided. PMID- 15273509 TI - Immunizations, neonatal jaundice, and animal-induced injuries. AB - PURPOSE OF REVIEW: Published studies during the past year about three topics important to the pediatric clinician-- immunizations, neonatal jaundice, and animal-induced injuries-are concisely reviewed. RECENT FINDINGS: Recent updates regarding vaccines including the questionable link with autism, implementation of universal influenza vaccination for young children, the efficacy of pneumococcal vaccine against invasive disease, and new information on pertussis, varicella, hepatitis A, hepatitis B, measles, and rotavirus vaccination are discussed. No association between measles/mumps/rubella vaccine or thimerosal-containing pertussis vaccine and autism is evident. Universal influenza vaccination for children 6 to 23 months of age will be recommended for the 2004-2005 flu season, and this implementation should reduce significant school absenteeism as well as complications seen last year including encephalopathy, seizures, respiratory failure, and pneumonia. Pneumococcal vaccine significantly reduces rates of invasive pneumococcal vaccine in healthy and HIV-infected children, although it does not appear to greatly affect otitis media rates. A reduction in post-vaccine febrile seizures appears to be present since the introduction of acellular pertussis vaccine. Multiple outbreaks in varicella have been reported since the introduction of the varicella vaccine, and a booster vaccination may be necessary in the future. Methods for detecting and preventing severe neonatal hyperbilirubinemia are reviewed, as well as anticipated recommendations from the American Academy of Pediatrics for the detection and management of hyperbilirubinemia. High bilirubin levels in preterm infants may result in hearing dysfunction and developmental impairment. The American Academy of Pediatrics has recommended a higher level of monitoring for newborn jaundice and treatment of hyperbilirubinemia in an effort to prevent kernicterus and sequelae from elevated bilirubin levels, including post-discharge follow-up appointment by day 3 to 5 of age. Dog bites in children with resultant post-traumatic stress disorder, rabies, and salmonellosis from pet reptiles in the home are also addressed. Clinicians need to be aware of the risk for rabies bites, need to recognize that dog bites in children appear to cause post-traumatic stress disorder in more than half of cases, and need to know how to educate patients on how to prevent salmonellosis from pet reptiles and amphibians. SUMMARY: Progress has been made in immunizations, especially immunization for influenza, pneumonia, and pertussis. It is recommended that monitoring for neonatal hyperbilirubinemia be more thorough to prevent the consequences of this condition. Rabies, post traumatic stress disorder from dog bites, and salmonellosis associated with pet reptiles constitute an important area for patient education. PMID- 15273512 TI - Understanding nurse work. PMID- 15273510 TI - Bibliography current world literature. Adolescent medicine. PMID- 15273514 TI - Quality in your hands: using PDA technology for data collection. PMID- 15273515 TI - Model Rules and Regulations for CNS Title Protection and Scope of Practice. PMID- 15273516 TI - Taking the high road: what should you do when an adverse event occurs? Part II. PMID- 15273517 TI - The role of neuroleptics in managing morphine-induced terminal delirium: implications for the clinical nurse specialist. PMID- 15273518 TI - Appraisal of tools to enhance evidence-based clinical practice. PMID- 15273520 TI - A genetics course for advanced clinical nursing practice. AB - PURPOSE: To describe a genetics course for advanced clinical nursing practice. BACKGROUND: The Human Genome Project is yielding new discoveries in genetics. The sheer volume of new information threatens to overwhelm healthcare providers as they must find ways to interpret and use these discoveries. DESCRIPTION OF PROJECT: The course was developed in recognition of the fact that graduate nursing students in advanced practice programs need to understand and apply genetic information in the care of their patients. The course builds on core competencies identified by the National Coalition for Health Professional Education in Genetics and the International Society of Nurses in Genetics. OUTCOMES: Students demonstrate that they can access, evaluate, and synthesize current information on a genetic condition and develop a comprehensive plan of care for patients with the selected condition. INTERPRETATION/CONCLUSION: Students learn basic and applied genetic concepts and skills. They are able to utilize resources appropriately to identify patient care needs and develop strategies for meeting those needs. IMPLICATIONS FOR NURSING PRACTICE: The need to remain current in understanding emerging technologies and genetic information will continue to grow as scientific discoveries are made that affect greater numbers of the public. Nurses in advanced practice are in a position to support patients and their families as they deal with the significance of these discoveries for their current and future health. PMID- 15273521 TI - Importance of physical activity, nutrition, and social support for optimal aging. AB - PURPOSE/OBJECTIVES: The purpose of this article was to provide the clinical nurse specialist (CNS), practicing in settings across the healthcare delivery continuum, with information about physical activity, nutrition, and social support that is essential for optimal aging. BACKGROUND/RATIONALE: Lifestyle choices older adults make concerning physical activity, diet, and social support greatly impact how well they age, their quality of life, and their well-being. DESCRIPTION OF THE PROJECT: This article focuses on 3 key factors essential to successful aging: physical activity, nutrition, and social support. It provides information and assessment and intervention strategy tools that can be used by CNSs to promote optimal aging for older clients. IMPLICATIONS FOR NURSING PRACTICE: This article provides the CNS with resources (literature, assessment tools, and tools for intervention strategies) to promote optimal aging and wellness. CNSs can assist their clients in various settings to age more slowly, delay and prevent certain age-related diseases, and promote more healthful, productive longevity by guiding health-promotion and wellness activities for their older clients. PMID- 15273522 TI - Clinical nurse specialists' knowledge specific to Medicare structures and processes. AB - PURPOSE: The purpose of this study was to describe clinical nurse specialists' (CNSs) knowledge of Medicare structures and processes using a multiple-choice examination. DESIGN: This descriptive study used the survey method with a convenient sample. SETTING: Potential subjects received the research study packet at the address designated by them for membership mailings sent from the National Association of Clinical Nurse Specialists (NACNS). SAMPLE: The population of interest was practicing CNSs. The accessible population was NACNS members drawn from the 2002 mailing list organized by zip code. Every third mailing label was skipped. The remaining list members (N = 950) received a study packet. There was a 14.42% response rate with 137 respondents. Eighty-two percent held a master's degree in nursing. METHODS: A 46 item multiple-choice examination was developed based upon a content map established after a review of the available Medicare related literature and consultation with Medicare advanced practice nurse providers. Content validity was established. Survey packets with the instrument, Scantron answer key, informational letter, and demographic profile were sent to potential subjects. Returned responses were keyed for correct answers. Relationships between demographic variables, raw score, and self-reported Medicare expertise were analyzed. Qualitative data elicited on the demographic form were thematically analyzed. FINDINGS: The examination had a.67 coefficient. Correlation of examination raw score to self-reported Medicare content expertise was 0.982 (P =.000). One third of the examination questions were incorrectly answered by 30% or more of respondents. Percentage correct scores ranged from 60.9 to 95.7 (M = 62.7%; SD = 10.46). Respondents offered 40 comments. Eleven comments related to a self-recognized Medicare knowledge deficit. Four responses highlighted that graduate studies had not provided them with the knowledge necessary for understanding reimbursement issues. Ten respondents expressed an interest in locating some type of educational resource addressing the Medicare system, reimbursement processes, and opportunities. CONCLUSIONS: Findings suggest that NACNS members have Medicare program knowledge deficits. Respondents recognize their deficiencies and are interested in obtaining more information about Medicare but are uncertain as to available resources or networking opportunities that might support them in this process. Graduate nursing programs may need to evaluate the amount of information provided within curriculums specific to insurance reimbursement and Medicare structures and processes. Study limitations include the low response rate and indeterminate sample representativeness. IMPLICATIONS FOR PRACTICE: Educators should examine CNS curriculums to ensure the adequacy of Medicare program information. Continuing education programs and conference workshops addressing Medicare information should be offered to CNSs interested in increasing their expertise or exploring provider opportunities. There may be entrepreneurial opportunities available to CNSs interested in developing an expertise in Medicare programs. Future research studies should quantify potential benefits to Medicare beneficiaries when CNSs participate as providers. PMID- 15273523 TI - Anne Shirah Dykes, MSN. PMID- 15273524 TI - Risk compensation: a "side effect" of sport injury prevention? PMID- 15273525 TI - Acromio-humeral distance variation measured by ultrasonography and its association with the outcome of rehabilitation for shoulder impingement syndrome. AB - OBJECTIVE: First, to validate an ultrasonographic measure of the acromio-humeral distance (AHD); second, to compare the AHD variation during active abduction in patients with shoulder impingement syndrome (SIS) and healthy subjects; and third, to evaluate the relationship between functional status and AHD variations before and after rehabilitation in SIS subjects. DESIGN: This study has 3 components: (1) a reliability study, (2) a case-control study, and (3) a preliminary pretreatment/posttreatment clinical trial. SETTING: Primary care hospital setting. PARTICIPANTS: Seven SIS patients and 13 healthy subjects. INTERVENTIONS: For the clinical trial, the SIS subjects participated in 12 sessions of a rehabilitation program over 4 weeks. MAIN OUTCOME MEASURES: First, intraclass correlation coefficient for interobserver reliability; second, AHD measured at 0 degrees, 45 degrees, and 60 degrees of active abduction; and third, Western Ontario Rotator Cuff Index. RESULTS: Intraclass correlation coefficient for interobserver reliability ranged from 0.86 to 0.92 for the 3 shoulder positions. A significant reduction of the AHD was found within groups between rest and active abduction (P < 0.05). Comparison of AHD between groups was not statistically different (P = 0.06; beta < 0.80). In pre-post rehabilitation analysis, improvement of the Western Ontario Rotator Cuff Index score was positively correlated to the reduction of the AHD narrowing as the arm was abducted (r = 0.86; P = 0.01). CONCLUSIONS: The ultrasound measure of AHD is reliable and sensitive. Although a distinct pattern of AHD variation in SIS patients could not be confirmed, a strong positive relationship was found between the reduction of AHD narrowing and functional improvement following rehabilitation. Ultrasound measurement of AHD might help identify SIS patients who will benefit from rehabilitation. PMID- 15273526 TI - Effect of bracing on patellofemoral joint stress while ascending and descending stairs. AB - OBJECTIVE: To test the hypothesis that individuals who respond favorably to bracing will exhibit decreased patellofemoral joint stress during stair ambulation. DESIGN: A repeated-measures, cross-sectional study. BACKGROUND: Ascending and descending stairs is one of the most painful activities of daily living for persons with patellofemoral pain (PFP). Although patellar bracing has been shown to reduce symptoms during such tasks, the underlying mechanism has not been identified. METHODS: Fifteen subjects with a diagnosis of PFP completed 2 phases of data collection: (1) magnetic resonance imaging to determine patellofemoral joint contact area, and (2) gait analysis during stair ascent and descent. Data were obtained under braced and non-braced conditions. Variables obtained from both data collection sessions were used as input variables into a biomechanical model to quantify patellofemoral joint stress. RESULTS: Although subjects reported an average decrease in pain of 56%, bracing did not reduce peak stress during stair ascent and descent. This finding can be explained by the fact that despite improvements in contact area, bracing resulted in greater knee extensor muscle moments and joint reaction forces. CONCLUSIONS: Our results do not support the hypothesis that individuals with PFP would demonstrate reduced patellofemoral stress during stair ambulation following the application of a patellar brace. CLINICAL RELEVANCE: Although bracing did not decrease patellofemoral joint stress during stair ascent and descent, the decrease in pain, increase in quadriceps utilization, and tolerance of joint reaction forces would appear to be beneficial consequences of bracing. PMID- 15273527 TI - Effect of training on postural control in figure skaters: a randomized controlled trial of neuromuscular versus basic off-ice training programs. AB - OBJECTIVES: To compare the effect of a neuromuscular training program and a basic exercise program on postural control in figure skaters. DESIGN: Two groups; parallel design; prospective, randomized controlled trial. SETTING: Postural control laboratory, arenas, September 2001 to December 2002. PARTICIPANTS: Forty four young, healthy figure skaters (18 years +/- 3 years). INTERVENTIONS: Participants were randomly assigned to receive a neuromuscular training program (n = 22) or a basic exercise training program (n = 22). Both programs were completed 3 times per week for 4 weeks, and each session was supervised. MAIN OUTCOME MEASUREMENTS: Participants completed baseline and postintervention measures of postural control on a force plate. Postural control was quantified as the center of pressure (CoP) path length during tests of single-limb standing balance that mimicked figure skating skills and challenged the postural control system to varying degrees. The primary outcome measure was the CoP path length observed during a landing jump test completed with eyes closed. RESULTS: The post intervention CoP path lengths during the more challenging tests were significantly (P < 0.05) lower (indicating better postural control) for the neuromuscular trained group than for the basic exercise-trained group. For the landing jump test completed with eyes closed, the percent improvement in the neuromuscular trained group was significantly greater (mean = 21.0 +/- 22.0%) than the basic exercise trained group (mean = -4.9 +/- 24.9%; P < 0.05). The magnitude of improvement in the neuromuscular-trained group ranged from approximately 1% to 21%, depending on the specific postural control test used. CONCLUSIONS: The results suggest that off-ice neuromuscular training can significantly improve postural control in figure skaters, whereas basic exercise training does not. PMID- 15273528 TI - Efficacy of celecoxib, a COX-2-specific inhibitor, and naproxen in the management of acute ankle sprain: results of a double-blind, randomized controlled trial. AB - OBJECTIVE: To assess the efficacy and safety of celecoxib and naproxen in the treatment of acute ankle sprain. DESIGN: Double-blind, parallel-group, randomized trial. SETTING: Multicenter outpatient. PATIENTS: Adult patients (n = 397) with acute first-degree or second-degree ankle sprain. INTERVENTIONS: Patients randomized to celecoxib 200 mg b.i.d. (n = 198) or naproxen 500 mg b.i.d. (n = 198) for 7 days. MAIN OUTCOME MEASURES: Primary measures of efficacy were Patient's Assessment of Ankle Pain Visual Analogue Scale (VAS) and Patient's Global Assessment of Ankle Injury. Secondary efficacy measures included Physician's Global Assessment of Ankle Injury, Patient's Return to Normal Function/Activity, and Patients' and Physicians' Satisfaction Assessments. Adverse events (AEs) were reported by investigators during the study. RESULTS: For the primary endpoints at day 4, the mean pain VAS scores were 31.9 mm +/- 1.96 for celecoxib and 29.0 mm +/- 1.91 for naproxen, and the responder rate for Patient's Global Assessment of Ankle Injury was 71% in the celecoxib group and 72% in the naproxen group, differences that were not statistically significant. In addition, noninferiority analysis demonstrated treatment differences that were within prespecified minimal clinical important differences. Gastrointestinal AEs were the most common AE, accounting for 14% in the celecoxib group and 21% in the naproxen group. The incidence of dyspepsia was 3% for celecoxib compared with 12% for naproxen (P = 0.032). CONCLUSIONS: Celecoxib is as effective as naproxen in treating acute first-degree or second-degree ankle sprains but causes significantly less dyspepsia. PMID- 15273529 TI - Prostaglandin E2 affects proliferation and collagen synthesis by human patellar tendon fibroblasts. AB - OBJECTIVE: To determine the effect of prostaglandin E2 on proliferation and collagen synthesis by human patellar tendon fibroblasts. DESIGN AND SETTING: Controlled laboratory study. METHODS: Human patellar tendon fibroblasts were treated with different concentrations (1, 10, 100 ng/mL) of prostaglandin E2 in cultures. Fibroblasts without prostaglandin E2 treatment were used as the control group. The fibroblast proliferation and collagen synthesis were measured using 3 (4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay and Sircol collagen assay, respectively. MAIN OUTCOME MEASURED: Changes in proliferation and collagen production of human patellar tendon fibroblasts. RESULTS: : At 1 ng/mL of prostaglandin E2, there was no significant effect on fibroblast proliferation compared with the control group. At concentrations of 10 ng/mL and 100 ng/mL prostaglandin E2, however, fibroblast proliferation significantly decreased, by 7.3% (P = 0.002) and 10.8% (P < 0.0001), respectively, compared with the control group. At 1 ng/mL of prostaglandin E2, collagen production of the tendon fibroblasts was unaffected. However, at both 10 ng/mL and 100 ng/mL prostaglandin E2, collagen production was significantly decreased, by 45.2% (P < 0.0001) and 45.7% (P < 0.0001), respectively, compared with the control group. The levels of collagen production between these 2 dosages did not differ significantly. CONCLUSIONS: Prostaglandin E2 affects the proliferation of and collagen production by human patellar tendon fibroblasts in a dosage-dependent manner. CLINICAL RELEVANCE: Based on these in vitro findings, we speculate that production of prostaglandin E2 in tendons might play some role in the acellularity and matrix disorganization seen in exercise-induced tendinopathy. PMID- 15273530 TI - Anterior compartment syndrome following an Achilles tendon repair: an unusual complication. PMID- 15273531 TI - Lesser trochanteric bursitis: a rare cause of anterior hip pain. PMID- 15273532 TI - Isolated tear of the pectoralis minor. PMID- 15273534 TI - Fluid replacement during marathon running. PMID- 15273536 TI - Evaluating the dislocated shoulder joint with MRI and arthroscopy. PMID- 15273537 TI - Intra-articular lidocaine or intravenous sedation for reduction of shoulder dislocations? PMID- 15273538 TI - Predicting the effects of sports-related concussion in young athletes. PMID- 15273539 TI - Physical activity and changes in mobility in older persons. PMID- 15273540 TI - Long-term prognosis after operation for adhesive small bowel obstruction. AB - AIM OF STUDY: The objective of this study was to determine the pattern of recurrence after one or more episodes of adhesive small bowel obstruction (ASBO) during a follow-up period of up to 40 years. Furthermore, we wanted to analyze possible factors with an influence on the recurrence rate and to study the magnitude of "everyday" abdominal pain among these patients. PATIENTS AND METHODS: Hospital records of 500 patients operated on for adhesive obstruction at Haukeland University Hospital from 1961 to 1995 were studied. The patients were followed until death, loss to follow-up, or end of study (February 2002), with a median follow-up of 10 years and a maximum follow-up time of 40 years. A questionnaire was sent to all living patients to obtain information on recurrences and abdominal complaints. RESULTS: The cumulative recurrence rate for patients operated once for ASBO was 18% after 10 years and 29% at 30 years. For patients admitted several times for ASBO, the relative risk of recurrent ASBO increased with increasing number of prior ASBO episodes. The cumulative recurrence rate reached 81% for patients with 4 or more ASBO admissions. Other factors influencing the recurrence rate were the method of treatment of the last previous ASBO episode (conservative versus surgical) and the number of abdominal operations prior to the initial ASBO operation. Compared to results from the general populations, more ASBO patients suffer from abdominal pain at home. Women and patients having matted adhesions have significantly more complaints about abdominal pain than men and patients with band adhesions. CONCLUSION: The risk of recurrence increased with increasing number of ASBO episodes. Most recurrent ASBO episodes occur within 5 years after the previous one, but a considerable risk is still present 10 to 20 years after an ASBO episode. Surgical treatment decreased the risk of future admissions for ASBO, but the risk of new surgically treated ASBO episodes was the same regardless of the method of treatment. People treated for ASBO seem to be more prone to experiencing abdominal pain than the normal population, especially those having matted adhesions. PMID- 15273541 TI - Intra-abdominal adhesion prevention: are we getting any closer? PMID- 15273542 TI - Classification of surgical complications: a new proposal with evaluation in a cohort of 6336 patients and results of a survey. AB - OBJECTIVE: Although quality assessment is gaining increasing attention, there is still no consensus on how to define and grade postoperative complications. This shortcoming hampers comparison of outcome data among different centers and therapies and over time. PATIENTS AND METHODS: A classification of complications published by one of the authors in 1992 was critically re-evaluated and modified to increase its accuracy and its acceptability in the surgical community. Modifications mainly focused on the manner of reporting life-threatening and permanently disabling complications. The new grading system still mostly relies on the therapy used to treat the complication. The classification was tested in a cohort of 6336 patients who underwent elective general surgery at our institution. The reproducibility and personal judgment of the classification were evaluated through an international survey with 2 questionnaires sent to 10 surgical centers worldwide. RESULTS: The new ranking system significantly correlated with complexity of surgery (P < 0.0001) as well as with the length of the hospital stay (P < 0.0001). A total of 144 surgeons from 10 different centers around the world and at different levels of training returned the survey. Ninety percent of the case presentations were correctly graded. The classification was considered to be simple (92% of the respondents), reproducible (91%), logical (92%), useful (90%), and comprehensive (89%). The answers of both questionnaires were not dependent on the origin of the reply and the level of training of the surgeons. CONCLUSIONS: The new complication classification appears reliable and may represent a compelling tool for quality assessment in surgery in all parts of the world. PMID- 15273543 TI - Measuring morbidity. PMID- 15273544 TI - Clinical application of bioartificial liver support systems. AB - OBJECTIVE: To review the present status of bioartificial liver (BAL) devices and their obtained clinical results. BACKGROUND: Acute liver failure (ALF) is a disease with a high mortality. Standard therapy at present is liver transplantation. Liver transplantation is hampered by the increasing shortage of organ donors, resulting in high incidence of patients with ALF dying on the transplantation waiting list. Among a variety of liver assist therapies, BAL therapy is marked as the most promising solution to bridge ALF patients to liver transplantation or to liver regeneration, because several BAL systems showed significant survival improvement in animal ALF studies. Until today, clinical application of 11 different BAL systems has been reported. METHODS: A literature review was performed using MEDLINE and additional library searches. Only BAL systems that have been used in a clinical trial were included in this review. RESULTS: Eleven BAL systems found clinical application. Three systems were studied in a controlled trial, showing no significant survival benefits, in part due to the insufficient number of patients included. The other systems were studied in a phase I trial or during treatment of a single patient and all showed to be safe. Most BAL therapies resulted in improvement of clinical and biochemical parameters. CONCLUSIONS: Bioartificial liver therapy for bridging patients with ALF to liver transplantation or liver regeneration is promising. Its clinical value awaits further improvement of BAL devices, replacement of hepatocytes of animal origin by human hepatocytes, and assessment in controlled clinical trials. PMID- 15273545 TI - Open repair of pectus excavatum with minimal cartilage resection. AB - OBJECTIVE: To summarize the clinical experience with a new open repair for pectus excavatum (PE), with minimal cartilage resection. SUMMARY BACKGROUND DATA: A wide variety of modified techniques of the Ravitch repair for PE have been used over the past 5 decades, with the complications and results being inconsistent. Extensive subperiosteal costal cartilage resection and perichondrial sheath detachment from the sternum may not be necessary for optimal repair. METHODS: During a 12-month period, 75 consecutive patients with symptomatic PE underwent open repair using a new less invasive technique. After exposing the deformed costal cartilages, a short chip was resected medially adjacent to the sternum and laterally at the level where the chest had a near normal contour, allowing the cartilage to be elevated to the desired level with minimal force. A transverse anterior sternal osteotomy was used on most patients. A substernal support strut was used for 66 patients; the strut was placed anterior to the sternum in 9 patients under age 12 and over age 40 years. The strut was routinely removed within 6 months. RESULTS: With a mean follow-up of 8.2 months, all but 1 patient regarded the results as very good or excellent. Mean operating time was 174 minutes; mean hospitalization was 2.7 days. There were no major complications or deaths. CONCLUSIONS: The open repair using minimal cartilage resection is effective for all variations of PE in patients of all ages, uses short operating time, provides a stable early postoperative chest wall, causes only mild postoperative pain, and produces good physiologic and cosmetic results. PMID- 15273546 TI - The early effect of the Roux-en-Y gastric bypass on hormones involved in body weight regulation and glucose metabolism. AB - OBJECTIVE: To evaluate the early effect of Roux-en-Y (RYGB) gastric bypass on hormones involved in body weight regulation and glucose metabolism. SIGNIFICANT BACKGROUND DATA: The RYGB is an effective bariatric procedure for which the mechanism of action has not been elucidated yet. Reports of hormonal changes after RYGB suggest a possible endocrine effect of the operation; however, it is unknown whether these changes are the cause or rather the effect of surgically induced weight loss. We speculated that if the mechanism of action of the RYGB involves an endocrine effect, then hormonal changes should occur early after surgery, prior to substantial body weight changes. METHODS: Ten patients with a mean preoperative body mass index (BMI) of 46.2 kg/m (40-53 kg/m) underwent laparoscopic RYGB. Six patients had type 2 diabetes treated by oral hypoglycemic agents. Preoperatively and 3 weeks following surgery, all patients were tested for fasting glucose, insulin, glucagon, insulin-like growth factor 1 (IGF-1), leptin, gastric inhibitory polypeptide (GIP), glucagon-like peptide-1 (GLP-1), cholecystokinin (CCK), adrenocorticotropic hormone (ACTH), corticosterone, and neuropeptide Y (NPY). RESULTS: Changes in mean BMI were rather minimal (43.2 kg/m; P = not significant), but there was a significant decrease in blood glucose (P = 0.005), insulin (P = 0.02), IGF-1 (P < 0.05), leptin (P = 0.001), and an increase in ACTH levels (P = 0.01). The other hormones were not significantly changed by surgery. All the 6 diabetic patients had normal glucose and insulin levels and did not require medications after surgery. The RYGB reduced GIP levels in diabetic patients (P < 0.01), whereas no changes in GIP levels were found in nondiabetics. CONCLUSIONS: Roux-en-Y gastric bypass determines considerable hormonal changes before significant BMI changes take place. These results support the hypothesis of an endocrine effect as the possible mechanism of action of RYGB. PMID- 15273547 TI - Effects of bariatric surgery in older patients. AB - OBJECTIVE: Evaluate the safety and efficacy of bariatric surgery in older patients. BACKGROUND: Because of an increased morbidity in older patients who may not be as active as younger individuals, there remain concerns that they may not tolerate the operation well or lose adequate amounts of weight. METHODS: The database of patients who had undergone bariatric surgery since 1980 and National Death Index were queried for patients <60 and >/= 60 years of age. GBP was the procedure of choice after 1985. Data evaluated at 1 and 5 years included weight lost, % weight lost (%WL), % excess weight loss (%EWL), % ideal body weight (%IBW), mortality, complications, and obesity comorbidity. RESULTS: Eighty patients underwent bariatric surgery: age 63 +/- 3 years, 78% women, 68 white, 132 +/- 22 kg, BMI 49 +/- 7 kg/m, 217 +/- 32%IBW. Preoperative comorbidity, was greater (P < 0.001) in patients >/= 60 years. There were no operative deaths but 11 late deaths. COMPLICATIONS: 4 major wound infections, 2 anastomotic leaks, 10 symptomatic marginal ulcers, 5 stomal stenoses, 3 bowel obstructions, 26 incisional hernias (nonlaparoscopic), and 1 pulmonary embolism. At 1 year after surgery (94% follow-up), patients lost 38 +/- 11 kg, 57%EWL, 30%WL, BMI 34.5 +/- 7 kg/m, %IBW 153 +/- 31. Comorbidities decreased (P < 0.001); however, %WL and %EWL and improvement in hypertension and orthopedic problems, although significant, were greater in younger patients. At 5 years after surgery (58% follow-up), they had lost 31 +/- 18 kg, 50%EWL, 26%WL, BMI 35 +/- 8 kg/m, and %IBW 156 +/- 36. CONCLUSIONS: Bariatric surgery was effective for older patients with a low morbidity and mortality. Older patients had more pre- and post operative comorbidities and lost less weight than younger patients. However the weight loss and improvement in comorbidities in older patients were clinically significant. PMID- 15273548 TI - Surgical manipulation of the intestine results in quantitative and qualitative alterations in luminal Escherichia coli. AB - OBJECTIVES: To look at the qualitative and quantitative changes in the luminal bacterial flora in response to surgical manipulation of the small intestine. SUMMARY BACKGROUND DATA: The barrier function of the intestine is compromised in pathologic conditions, such as shock, trauma, or surgical stress. Our earlier work has shown that surgical manipulation results in oxidative stress in the intestinal mucosa leading to permeability alterations. METHODS: Studies were done on rats, which were randomly divided into four groups (n = 8): group I, control, group II, III, IV different time periods, such as 8, 12, and 24 hours after surgical manipulation, which was simulated by opening the abdominal wall and handling the intestine. The cecal wall and cecal luminal contents were harvested under sterile conditions and processed for quantitation for aerobes and anaerobes. Adherence assays using Hep-2 cells were carried out on Escherichia coli isolated under different experimental conditions. In addition, control E. coli were exposed to superoxide or hydrogen peroxide, followed by subculture and adherence studies. RESULTS: Surgical manipulation of the intestine resulted in qualitative and quantitative alterations in the aerobic bacteria. There was an increase in the number and relative proportion of E. coli in the cecal flora, and there was also an increase in adherence of E. coli to cecal mucosa, which was confirmed by in vitro bacterial adherence studies with HEp-2 cells. These changes were maximum at 12 hours following surgical manipulation and by 24 hours, this came back to control pattern. Control E. coli after in vitro exposure to oxidants also showed increased adherence. CONCLUSION: These studies suggest that oxidative stress in the mucosa following surgical manipulation results in alterations in the luminal bacteria leading to increased bacterial adherence onto mucosal epithelium, which may contribute to postsurgical complications. PMID- 15273549 TI - Anastomotic leakage is predictive of diminished survival after potentially curative resection for colorectal cancer. AB - OBJECTIVE: The aim of this study was to determine whether anastomotic leakage has an independent association with overall survival and cancer-specific survival. SUMMARY BACKGROUND DATA: There are many known prognostic indicators following surgery for colorectal cancer (CRC). However, the impact of anastomotic leakage has not been adequately assessed. METHODS: Consecutive patients undergoing resection between 1971 and 1999 were recorded prospectively in the Concord Hospital CRC database. Total anastomotic leakage was defined as any leak, whether local, general, or radiologically diagnosed. Patients were followed until death or to December 31, 2002. The association between anastomotic leakage and both overall survival and cancer-specific survival was examined by proportional hazards regression with adjustment for other patient and tumor characteristics influencing survival. Confidence intervals (CI) were set at the 95% level. RESULTS: From an initial 2980 patients, 1722 remained after exclusions. The total leak rate was 5.1% (CI 4.1-6.2%). In patients with a leak, the 5-year overall survival rate was 44.3% (CI 33.5-54.6%) compared to 64.0% (CI 61.5-66.3%) in those without leak. In proportional hazards regression-after adjustment for age, gender, urgent resection, site, size, stage, grade, venous invasion, apical node metastasis and serosal surface involvement-anastomotic leakage had an independent negative association with overall survival (hazard ratio [HR] 1.6, CI 1.2-2.0) and cancer-specific survival (HR 1.8, CI 1.2-2.6). CONCLUSION: Apart from its immediate clinical consequences, anastomotic leakage also has an independent negative association with survival. PMID- 15273550 TI - Anterior resection for rectal cancer with mesorectal excision: a prospective evaluation of 622 patients. AB - OBJECTIVE: This study aims to review the operative results and oncological outcomes of anterior resection for rectal and rectosigmoid cancer. Comparison was made between patients with total mesorectal excision (TME) for mid and distal cancer and partial mesorectal excision (PME) for proximal cancer, when a 4- to 5 cm mesorectal margin could be achieved. Risk factors for local recurrence and survival were also analyzed. SUMMARY BACKGROUND DATA: Anterior resection has become the preferred treatment option rectal cancer. TME with sharp dissection has been shown to be associated with a low local recurrence rate. Controversies still exist as to the need for TME in more proximal tumor. METHODS: Resection of primary rectal and rectosigmoid cancer was performed in 786 patients from August 1993 to July 2002. Of these, 622 patients (395 men and 227 women; median age, 67 years) underwent anterior resection. The technique of perimesorectal dissection was used. Patients with mid and distal rectal cancer were treated with TME while PME was performed for those with more proximal tumors. Prospective data on the postoperative results and oncological outcomes were reviewed. Risk factors for anastomotic leakage, local recurrence, and survival of the patients were analyzed with univariate and multivariate analysis. RESULTS: The median level of the tumor was 8 cm from the anal verge (range, 2.5-20 cm) and curative resection was performed in 563 patients (90.5%). TME was performed in 396 patients (63.7%). Significantly longer median operating time, more blood loss, and a longer hospital stay were found in patients with TME. The overall operative mortality and morbidity rates were 1.8% and 32.6%, respectively, and there were no significant differences between those of TME and PME. Anastomotic leak occurred in 8.1% and 1.3% of patients with TME and PME, respectively (P < 0.001). Independent factors for a higher anastomotic leakage rate were TME, the male gender, the absence of stoma, and the increased blood loss. The 5-year actuarial local recurrence rate was 9.7%. The advanced stage of the disease and the performance of coloanal anastomosis were independent factors for increased local recurrence. The 5-year cancer-specific survival was 74.5%. The independent factors for poor survival were the advanced stage of the disease and the presence of lymphovascular and perineural invasion. CONCLUSIONS: Anterior resection with mesorectal excision is a safe option and can be performed in the majority of patients with rectal cancer. The local recurrence rate was 9.7% and the cancer specific survival was 74.5%. When the tumor requires a TME, this procedure is more complex and has a higher leakage rate than in those higher tumors where PME provides adequate mesorectal clearance. By performing TME in patients with mid and distal rectal cancer, the local control and survival of these patients are similar to those of patients with proximal cancers where adequate clearance can be achieved by PME. PMID- 15273551 TI - Innate immunity genes influence the severity of acute appendicitis. AB - OBJECTIVE: Using acute appendicitis as a model, we tested the hypothesis that polymorphisms in genes involved in host defense can be associated with the severity of local infection-inflammation in humans. SUMMARY BACKGROUND DATA: Innate immunity is the body's front-line system for antimicrobial host defense. Local inflammation is a major innate immune mechanism for containing and destroying microbes, but it may also contribute to tissue injury. METHODS: We studied 134 patients with acute appendicitis treated at an urban hospital. We looked for associations between the severity of appendicitis (uncomplicated vs. perforated or gangrenous), plasma and peritoneal cytokine concentrations, and single nucleotide polymorphisms in genes involved in recognizing bacterial molecules [CD14 (-159 C-->T); TLR4 (896 A-->G)] and in mounting an inflammatory response [IL-6 (-174 G-->C), TNF-alpha (-308 G-->A), IL-1beta (-31 C -->T)]. RESULTS: Ninety-one patients (68%) had uncomplicated appendicitis and 43 (32%) had complicated disease. The SNPs in the CD14, TLR4, IL-1beta, and TNF-alpha genes were not associated with the severity of appendicitis. A strong association was found between C-allele carriage at -174 in the IL-6 gene and decreased risk of complicated disease (adjusted odds ratio = 0.24, 95% CI = 0.07-0.76). Lower plasma and peritoneal fluid IL-6 concentrations in the IL-6 -174 C-carriers than in the GG homozygotes suggest that this polymorphism contributes to decreased IL 6 production in vivo. CONCLUSIONS: Polymorphism in the IL-6 gene was associated with the severity of appendicitis, even after adjustment for duration of symptoms. The risk for developing appendiceal perforation or gangrene may be determined, in part, by variation in the IL-6 gene. PMID- 15273552 TI - Incomplete cytoreduction in 174 patients with peritoneal carcinomatosis from appendiceal malignancy. AB - OBJECTIVE: The aim of this study was to analyze the survival of patients with peritoneal dissemination of appendiceal malignancy having incomplete cytoreductive surgery. SUMMARY BACKGROUND DATA: Cytoreductive surgery plus perioperative intraperitoneal chemotherapy has emerged as a new and potentially curative treatment option for patients with peritoneal dissemination of appendiceal mucinous tumors. The goal of surgery is to remove all visible disease. Nevertheless, in some patients, complete cytoreduction is not possible. METHODS: Over a 30-year period, 645 patients with epithelial peritoneal surface malignancy of appendiceal origin were treated with cytoreductive surgery and intraperitoneal chemotherapy by a single surgeon. One hundred seventy-four (27.1%) of these patients had an incomplete cytoreduction. A critical statistical analysis of the impact of selected clinical features on survival was performed from a prospective database. RESULTS: Mortality and morbidity rates were 0% and 33.3%, respectively. Median survival of these 174 patients was 20.5 months and their 1-year, 3-year, and 5-year survival rates were 71%, 34%, and 15%, respectively. By multivariate analysis, the presence of signet ring cells and lymph node involvement were independent prognostic indicators of poor survival (P = 0.047 and P < 0.001, respectively). Patients who underwent more than 1 cytoreduction or repeat intraperitoneal chemohyperthermia showed significant improvement in survival (P = 0.018 and P < 0.001, respectively) CONCLUSION: Incomplete cytoreduction plus perioperative intraperitoneal chemotherapy of peritoneal dissemination from appendiceal malignancy results in limited long-term survival. Patients with signet ring histology or lymph node involvement have an especially poor outcome. Repeat cytoreduction and intraperitoneal chemohyperthermia may improve outcome. PMID- 15273553 TI - Outcome of splenectomy for thrombocytopenia associated with systemic lupus erythematosus. AB - OBJECTIVE: To determine the efficacy of splenectomy for treating thrombocytopenia associated with systemic lupus erythematosus (SLE). SUMMARY BACKGROUND DATA: The role of splenectomy has been controversial in this patient population. METHODS: Between 1975 and 2001, 25 consecutive adults with SLE underwent splenectomy specifically for thrombocytopenia. Surgical indications, operative mortality and morbidity, and hematological outcomes were followed in both the short-term (first 30 days) and the long-term (last recorded platelet count, last contact, or death). Response to splenectomy was rated as: complete (CR: platelets >/=150 x 10/L for at least 4 weeks), partial (PR: platelets 50-149 x 10/L for at least 4 weeks), or none (NR: platelets < 50 x 10/L at all times). Relapse occurred if platelets fell below 50 x 10/L after CR or PR. RESULTS: Indications for splenectomy included: thrombocytopenia refractory to (64%), dependent on (20%), or patient intolerance of (16%) medical treatments. Perioperative mortality was 0% and morbidity was 24%. After a median of 9.5 years, 9 patients (36%) had died, with only 1 death being secondary to bleeding. Early partial or complete response rate to splenectomy was 88%. After a median follow-up of 6.6 years, 16 (64%) patients had sustained complete or partial response without relapse. Eight (32%) of these patients required adjunctive medical therapy, whereas the other 8 (32%) did not. The remaining 9 (36%) patients relapsed, but 5 (20%) of the 9 patients were subsequently salvaged to at least partial response with further treatments. The overall PR or CR to splenectomy combined with medical therapy was 84%. CONCLUSION: Splenectomy should be considered safe and efficacious for thrombocytopenia associated with SLE. PMID- 15273554 TI - Prognostic nomogram for patients undergoing resection for adenocarcinoma of the pancreas. AB - BACKGROUND: Predictive nomograms are becoming increasingly used to define and predict outcome. They can be developed at presentation or following treatment and include variables not conventionally used in standard staging systems. METHODS: We use a predictive nomogram based on prospectively collected data from 555 pancreatic resections for adenocarcinoma at a single institution. At last follow up, 481 (87%) had died, defining a mature and comprehensive database. We used a 1 , 2-, and 3-year follow-up, as the number of patients alive beyond 3 years is sufficiently limited to provide insufficient events. RESULTS: Based on a Cox model, we then developed a nomogram that predicts the probability that a patient will survive pancreatic cancer for 1, 2, and 3 years from the time of the initial resection, assuming that there is not death from an alternate cause. Calibration between observed and corrected is good, and variables not conventionally associated with standard staging systems improved the predictivity of the model. CONCLUSIONS: This nomogram can serve as a basis for investigating other potentially predictive variables that are proposed of prognostic importance for patients undergoing resection for adenocarcinoma of the pancreas. PMID- 15273555 TI - Salvage surgery following downstaging of unresectable hepatocellular carcinoma. AB - OBJECTIVE: We reported here a series of 49 patients with unresectable hepatocellular carcinoma (HCC) who underwent nonsurgical treatment to downstage the disease followed by salvage surgery, their long-term outcome, and pattern of recurrence. SUMMARY BACKGROUND DATA: Most HCC patients present with unresectable disease and are treated with chemotherapy or intra-arterial therapy with a palliative intent. Occasionally, there are good responses to treatment so that salvage surgery becomes feasible afterward. However, long-term outcomes of these patients are seldom reported. METHODS: Patients with unresectable hepatocellular carcinoma, from September 1993 to June 2002, who received salvage surgery after downstaging by systemic chemotherapy, intra-arterial yttrium-90 microspheres, or sequential treatment were included in this study. Systemic chemotherapy consisted of combination doxorubicin, cisplatin, interferon-alpha and 5-fluorouracil (5 FU), or single-agent doxorubicin. The choice of treatment was according to stage of disease and contemporary clinical trial protocol. Survival, recurrence pattern, and surgical outcome were studied. RESULTS: There were 49 patients in this study with 40 males and 9 females, age ranged from 12 to 69 years. Forty patients (81.6%) were hepatitis B positive. Thirty-two patients had combination chemotherapy alone (65.3%), 8 patients had single agent chemotherapy alone (16.3%), 4 patients received intra-arterial yttrium-90 microspheres alone (8.2%), and 5 patients received sequential therapy (10.2%). Twenty-eight (57.1%) patients received major hepatic resection. Thirteen patients (26.5%) had complete necrosis of the tumor after treatment. Twenty-one patients (42.9%) had recurrence after surgery, and 14 of them were intrahepatic recurrence. The median survival was 85.9 months. The 1-year, 3-year, and 5-year survival rates were 98%, 64%, and 57%, respectively. CONCLUSIONS: Salvage surgery after successful downstaging can provide long-term control of disease in a small proportion of patients with unresectable hepatocellular carcinoma. PMID- 15273556 TI - Prognostic value of lymphangiogenesis and lymphovascular invasion in invasive breast cancer. AB - OBJECTIVE: The aim of this study was to investigate the prognostic relevance of lymphangiogenesis and lymphovascular invasion in a large cohort of breast cancer patients. INTRODUCTION: Invasion of tumor cells into blood and lymphatic vessels is one of the critical steps for metastasis. The presence or absence of lymph node metastasis is one of the main decision criteria for further therapy. One shortcoming of previous morphologic studies was the lack of specific markers that could exact discriminate between blood and lymphatic vessels. The aim of this study was to evaluate the prognostic relevance of lymphangiogenesis and lymphovascular invasion in breast cancer patients. METHODS: We investigated 374 tissue specimens of patients suffering from invasive breast cancer by immunostaining for the lymphatic endothelial specific marker podoplanin. Lymphangiogenesis, quantified by evaluating the lymphatic microvessels density (LMVD), and lymphovascular invasion (LVI) were correlated with various clinical parameters and prognostic relevance. RESULTS: LMVD correlated significantly with LVI (P = 0.001). LVI was associated significantly with a higher risk for developing lymph-node metastasis (P = 0.004). Calculating the prognostic relevance, LVI presented as an independent prognostic parameter for disease free as well as overall survival (P = 0.001, and P = 0.001, respectively). CONCLUSION: Our data provide evidence that the biologic system of lymphangiogenesis constitutes a potential new target for development of anti-breast cancer therapeutic concepts. Our results further suggest that young, premenopausal patients with low differentiated breast tumors and high LMVD and LVI would, in particular, benefit from lymphangiogenesis-associated therapeutic strategies. PMID- 15273557 TI - Paradoxical effect of IL-18 therapy on the severe and mild Escherichia coli infections in burn-injured mice. AB - OBJECTIVE: To investigate the effects of IL-18 therapy on severe and mild bacterial infection after burn injury. SUMMARY BACKGROUND DATA: IL-18 therapy restores IFN-gamma production in immunosuppressive mice following burn injury and up-regulate host response to LPS and experimental bacterial peritonitis. On the other hand, the overproduction of IFN-gamma could induce an exaggerated inflammation. Therefore, in this study, we focus on the beneficial and deleterious effects of IL-18-induced IFN-gamma and investigate the behavior of IL 18 in infections. METHODS: Burn injury was induced in C57BL/6 mice and then they were i.p. injected with IL-18 (0.2 microg) on alternate days. After 1 week, severe and mild infections were made in mice by an Escherichia coli challenge (5 x 10 CFU and 1 x 10 CFU i.v., respectively). RESULTS: IL-18 therapy decreased the mortality of burn-injured mice followed by a severe infection, whereas it unexpectedly increased the mortality of burned mice with a mild infection. The IL 18 therapy increased the number of liver mononuclear cells (MNCs), especially NK cells, and greatly up-regulated the impaired IFN-gamma production from the liver and spleen MNCs in mice with severe infection. Both the serum IFN-gamma concentrations recovered while the bacterial count in the liver decreased. In contrast, the serum IFN-gamma concentrations of the burned mice with mild infection did not decrease in comparison to the unburned mice, whereas IL-18 therapy greatly up-regulated the serum IFN-gamma levels in burned mice. However, IL-18 therapy significantly elevated the serum ALT and creatinine levels, thus suggesting that the mortality was induced by an exaggerated form of shock/multiorgan failure. These beneficial and deleterious effects of IL-18 therapy in mice with severe and mild infections, respectively, were all inhibited by anti-IFN-gamma Ab pretreatment. CONCLUSION: IL-18 therapy can be a potent therapeutic tool against severe bacterial infection in immunocompromised hosts, but careful attention should also be paid to its adverse effects. PMID- 15273558 TI - Adrenomedullin and adrenomedullin binding protein-1 attenuate vascular endothelial cell apoptosis in sepsis. AB - OBJECTIVE: To determine whether vascular endothelial cell apoptosis occurs in the late stage of sepsis and, if so, whether administration of a potent vasodilatory peptide adrenomedullin and its newly reported specific binding protein (AM/AMBP 1) prevents sepsis-induced endothelial cell apoptosis. SUMMARY BACKGROUND DATA: Polymicrobial sepsis is characterized by an early, hyperdynamic phase followed by a late, hypodynamic phase. Our recent studies have shown that administration of AM/AMBP-1 delays or even prevents the transition from the hyperdynamic phase to the hypodynamic phase of sepsis, attenuates tissue injury, and decreases sepsis induced mortality. However, the mechanisms responsible for the beneficial effects of AM/AMBP-1 in sepsis remain unknown. METHODS: Polymicrobial sepsis was induced by cecal ligation and puncture in adult male rats. Human AMBP-1 (40 microg/kg body weight) was infused intravenously at the beginning of sepsis for 20 minutes and synthetic AM (12 microg/kg body weight) was continuously administered for the entire study period using an Alzert micro-osmotic pump, beginning 3 hours prior to the induction of sepsis. The thoracic aorta and pulmonary tissues were harvested at 20 hours after cecal ligation and puncture (ie, the late stage of sepsis). Apoptosis was determined using TUNEL assay, M30 Cytodeath immunostaining, and electromicroscopy. In addition, anti-apoptotic Bcl-2 and pro apoptotic Bax gene expression and protein levels were assessed by RT-PCR and Western blot analysis, respectively. RESULTS: Vascular endothelial cells underwent apoptosis formation at 20 hours after cecal ligation and puncture as determined by three different methods. Moreover, partial detached endothelial cell in the aorta was observed. Bcl-2 mRNA and protein levels decreased significantly at 20 hours after the onset of sepsis while Bax was not altered. Administration of AM/AMBP-1 early after sepsis, however, significantly reduced the number of apoptotic endothelial cells. This was associated with significantly increased Bcl-2 protein levels and decreased Bax gene expression in the aortic and pulmonary tissues. CONCLUSION: The above results suggest that vascular endothelial cell apoptosis occurs in late sepsis and the anti-apoptotic effects of AM/AMBP-1 appear to be in part responsible for their beneficial effects observed under such conditions. PMID- 15273559 TI - Albumin protects against gut-induced lung injury in vitro and in vivo. AB - OBJECTIVE: Since albumin has the ability to detoxify, we assessed whether low dose albumin could protect against trauma/hemorrhagic shock (T/HS)-induced endothelial cell, lung, gut, and red blood cell (RBC) injury in vivo and endothelial cell injury in vitro. SUMMARY BACKGROUND DATA: T/HS cause ischemic insult to the gut, resulting in the release of biologically active factors into the mesenteric lymph, which then cause injury to multiple distant organs. METHODS: In vitro experiments tested the ability of albumin to reduce the cytotoxicity of mesenteric lymph from male rats subjected to T/HS (laparotomy + MAP 30 mm Hg for 90 minutes) for human umbilical vein endothelial cell (HUVEC). In subsequent in vivo experiments, the ability of albumin given as part of the resuscitation regimen to protect against T/HS-induced injury was tested by comparing the magnitude of injury in T/HS rats receiving human albumin (shed blood + 0.12, 0.24, or 0.36 g/kg) or lactated Ringer's solution (shed blood + 2 x volume of shed blood as LR) with that observed in rats subjected to trauma/sham shock. Rats were killed after a 3-hour recovery period and had lung permeability evaluated by bronchoalveolar lavage and myeloperoxidase assays, intestinal microvillous injury by histology, and RBC deformability using ektacytometry. RESULTS: Both bovine and human albumin prevented T/HS lymph-induced HUVEC cytotoxicity in vitro, even when added 30 minutes after the lymph (viability 15 +/- 4% to 88 +/- 3%, P < 0.01). In vivo RBC deformability was better preserved by blood plus albumin than blood plus lactated Ringer's solution (P < 0.01). Likewise, albumin administration reduced T/HS-induced lung permeability and neutrophil sequestration in a dose-dependent fashion, with 0.36 g/kg of albumin effecting total lung protection (P < 0.01). In contrast, albumin treatment did not prevent T/HS-induced gut injury. CONCLUSIONS: Low-dose albumin protects against gut lymph-induced lung, HUVEC, and RBC injury by neutralizing T/HS lymph toxicity. PMID- 15273560 TI - Insulin treatment improves hepatic morphology and function through modulation of hepatic signals after severe trauma. AB - OBJECTIVE: The purpose of the present study was to determine the effect of insulin therapy on hepatic function, structure, and hepatic mRNA and protein cytokine expression during the hypermetabolic cascade post burn. SUMMARY BACKGROUND DATA: Liver function and morphology are crucial for survival of patients suffering from trauma, operations, or infections. Insulin decreased mortality and prevented the incidence of multiorgan failure in critically ill patients. METHODS: Rats received a thermal injury and were randomly divided into the insulin or control group. Our outcome measures encompassed the effect of insulin on hepatic proteins, hepatic pro- and anti-inflammatory cytokines mRNA and proteins, hepatocyte proliferation, including Bcl-2 and hepatocyte apoptosis, with caspases-3 and caspases-9. RESULTS: Insulin significantly improved hepatic protein synthesis by increasing albumin and decreasing c-reactive protein and fat (P < 0.05). Insulin decreased the hepatic inflammatory response signal cascade by decreasing hepatic pro-inflammatory cytokines mRNA and proteins IL-1beta and tumor necrosis factor at pretranslational levels. Insulin increased hepatic cytokine mRNA and protein expression of IL-2 and IL-10 at a pretranslational level when compared with controls (P < 0.05). Insulin increased hepatocyte proliferation along with Bcl-2 concentration, while decreasing hepatocyte apoptosis along with decreased caspases-3 and -9 concentration, thus improving liver morphology (P < 0.05). CONCLUSIONS: Our data provide insight that insulin attenuates the inflammatory response by decreasing the pro-inflammatory and increasing the anti-inflammatory cascade, thus restoring hepatic homeostasis, which has been shown to be critical for organ function and survival of critically ill patients. PMID- 15273561 TI - Live donor liver transplantation without blood products: strategies developed for Jehovah's Witnesses offer broad application. AB - OBJECTIVE: Developing strategies for transfusion-free live donor liver transplantation in Jehovah's Witness patients. SUMMARY BACKGROUND DATA: Liver transplantation is the standard of care for patients with end-stage liver disease. A disproportionate increase in transplant candidates and an allocation policy restructuring, favoring patients with advanced disease, have led to longer waiting time and increased medical acuity for transplant recipients. Consequently, Jehovah's Witness patients, who refuse blood product transfusion, are usually excluded from liver transplantation. We combined blood augmentation and conservation practices with live donor liver transplantation (LDLT) to accomplish successful LDLT in Jehovah's Witness patients without blood products. Our algorithm provides broad possibilities for blood conservation for all surgical patients. METHODS: From September 1998 until June 2001, 38 LDLTs were performed at Keck USC School of Medicine: 8 in Jehovah's Witness patients (transfusion-free group) and 30 in non-Jehovah's Witness patients (transfusion eligible group). All transfusion-free patients underwent preoperative blood augmentation with erythropoietin, intraoperative cell salvage, and acute normovolemic hemodilution. These techniques were used in only 7%, 80%, and 10%, respectively, in transfusion-eligible patients. Perioperative clinical data and outcomes were retrospectively reviewed. Data from both groups were statistically analyzed. RESULTS: Preoperative liver disease severity was similar in both groups; however, transfusion-free patients had significantly higher hematocrit levels following erythropoietin augmentation. Operative time, blood loss, and postoperative hematocrits were similar in both groups. No blood products were used in transfusion-free patients while 80% of transfusion-eligible patients received a median of 4.5+/- 3.5 units of packed red cell. ICU and total hospital stay were similar in both groups. The survival rate was 100% in transfusion-free patients and 90% in transfusion-eligible patients. CONCLUSIONS: Timely LDLT can be done successfully without blood product transfusion in selected patients. Preoperative preparation, intraoperative cell salvage, and acute normovolemic hemodilution are essential. These techniques may be widely applied to all patients for several surgical procedures. Chronic blood product shortages, as well as the known and unknown risk of blood products, should serve as the driving force for development of transfusion-free technology. PMID- 15273562 TI - Laparoscopic live donor nephrectomy: trends in donor and recipient morbidity following 381 consecutive cases. AB - OBJECTIVE: To review a single-institution 6-year experience with laparoscopic live donor nephrectomy detailing the technical modifications, clinical results, as well as the trends in donor and recipient morbidity. SUMMARY BACKGROUND DATA: Since 1995, laparoscopic donor nephrectomy has had a significant impact on the field of renal transplantation, resulting in decreased donor morbidity, without jeopardizing procurement of a high-quality renal allograft. This technique has become the preferred method of allograft procurement for many transplantation centers worldwide but still remains technically challenging with a steep learning curve. METHODS: Records from 381 consecutive laparoscopic donor nephrectomies were reviewed with evaluation of both donor and recipient outcomes. Trends in donor and recipient complications were assessed over time by comparing the outcomes between four equally divided groups. RESULTS: All 381 kidneys were procured and transplanted successfully with only 8 (2.1%) open conversions. Mean operative time was 252.9 +/- 55.7 minutes, estimated blood loss 344.2 +/- 690.3 mL, warm ischemia time 4.9 +/- 3.4 minutes, and donor length of stay was 3.3 +/- 4.5 days. There was a significant decline in total donor complications, allograft loss, and rate of vascular thrombosis with experience. The rate of ureteral complications declined significantly when comparing our early (Group A) versus later (Groups B-D) experience. CONCLUSION: Laparoscopic donor nephrectomy has remained a safe, less invasive, and effective technique for renal allograft procurement. Over our 6-year experience and with specific refinements in surgical technique, we have observed a decline in both donor and recipient morbidity following laparoscopic live donor nephrectomy. PMID- 15273563 TI - Resuscitation with 100% oxygen causes intestinal glutathione oxidation and reoxygenation injury in asphyxiated newborn piglets. AB - OBJECTIVE: To compare mesenteric blood flow, oxidative stress, and mucosal injury in piglet small intestine during hypoxemia and reoxygenation with 21%, 50%, or 100% oxygen. SUMMARY BACKGROUND DATA: Necrotizing enterocolitis is a disease whose pathogenesis likely involves hypoxia-reoxygenation and the generation of oxygen-free radicals, which are known to cause intestinal injury. Resuscitation of asphyxiated newborns with 100% oxygen has been shown to increase oxidative stress, as measured by the glutathione redox ratio, and thus may predispose to free radical-mediated tissue injury. METHODS: Newborn piglets subjected to severe hypoxemia for 2 hours were resuscitated with 21%, 50%, or 100% oxygen while superior mesenteric artery (SMA) flow and hemodynamic parameters were continuously measured. Small intestinal tissue samples were analyzed for histologic injury and levels of oxidized and reduced glutathione. RESULTS: SMA blood flow decreased to 34% and mesenteric oxygen delivery decreased to 9% in hypoxemic piglets compared with sham-operated controls. With reoxygenation, SMA blood flow increased to 177%, 157%, and 145% of baseline values in piglets resuscitated with 21%, 50%, and 100% oxygen, respectively. Mesenteric oxygen delivery increased to more than 150% of baseline values in piglets resuscitated with 50% or 100% oxygen, and this correlated significantly with the degree of oxidative stress, as measured by the oxidized-to-reduced glutathione ratio. Two of eight piglets resuscitated with 100% oxygen developed gross and microscopic evidence of pneumatosis intestinalis and severe mucosal injury, while all other piglets were grossly normal. CONCLUSIONS: Resuscitation of hypoxemic newborn piglets with 100% oxygen is associated with an increase in oxygen delivery and oxidative stress, and may be associated with the development of small intestinal hypoxia-reoxygenation injury. Resuscitation of asphyxiated newborns with lower oxygen concentrations may help to decrease the risk of necrotizing enterocolitis. PMID- 15273564 TI - The educational impact of bench model fidelity on the acquisition of technical skill: the use of clinically relevant outcome measures. AB - OBJECTIVE: To evaluate the impact of bench model fidelity on the acquisition of technical skill using clinically relevant outcome measures. METHODS: Fifty junior surgery residents participated in a 1-day microsurgical training course. Participants were randomized to 1 of 3 groups: 1) high-fidelity model training (live rat vas deferens; n = 21); 2) low-fidelity model training (silicone tubing; n = 19); or 3) didactic training alone (n = 10). Following training, all participants were assessed on the high- and low-fidelity bench models. Immediate outcome measures included procedure times, blinded, expert assessment of videotaped performance using checklists and global rating scales, anastomotic patency, suture placement precision, and final product ratings. Delayed outcome measures (obtained from the live rat vas deferens 30 days following training) included anastomotic patency, presence of a sperm granuloma, and the presence of sperm on microscopy. RESULTS: Following training, checklist (P < 0.001) and global rating scores (P < 0.001) on the bench model simulators were higher among subjects who received hands-on training, irrespective of model fidelity. Immediate anastomotic patency rates of the rat vas deferens were higher with increasing model fidelity training (P = 0.048). Delayed anastomotic patency rates were higher among subjects who received bench model training, irrespective of model fidelity (P = 0.02). Rates of sperm presence on microscopy were higher among subjects who received high-fidelity model training compared with subjects who received didactic training (P = 0.039) but did not differ among subjects in the high- and low-fidelity groups. CONCLUSIONS: Surgical skills training on low fidelity bench models appears to be as effective as high-fidelity model training for the acquisition of technical skill among novice surgeons. PMID- 15273565 TI - Are actual standard fluid regimens in major surgery safe? PMID- 15273566 TI - Prognostic factors following curative resection for pancreatic adenocarcinoma. PMID- 15273568 TI - Effects of intravenous fluid restriction on postoperative complications: comparison of two perioperative fluid regimens: a randomized assessor-blinded multicenter trial. PMID- 15273570 TI - Effect of duodenal-jejunal exclusion in a non-obese animal model of type 2 diabetes: a new perspective for an old disease. PMID- 15273572 TI - Laparoscopic adjustable silicone gastric banding versus vertical banded gastroplasty in morbidly obese patients. PMID- 15273573 TI - Laparoscopic adjustable silicone gastric banding versus vertical banded gastroplasty in morbidly obese patients. PMID- 15273576 TI - Predictors of repeat testing and HIV seroconversion in a sexually transmitted disease clinic population. AB - OBJECTIVE: This study assessed the extent of and characteristics associated with repeat HIV testing in persons presenting to a sexually transmitted disease (STD) clinic. METHODS: The study population included all 101 newly diagnosed HIV infected subjects and 411 matched HIV-uninfected subjects identified over a 5 year period in a publicly funded STD clinic in the southeastern United States. RESULTS: Of the 508 subjects (99%) with available records, 160 (32%) had tested previously. Age, race, return for posttest counseling, and the client's stated reason for coming to the clinic were associated with repeat testing. Among the 160 subjects who had tested previously, self-identifying as a man who has sex with men or having a history of incarceration was strongly associated with HIV seroconversion (adjusted odds ratio [OR], 51.82; 95% confidence interval [CI], 9.10-295.13; adjusted OR, 83.98, 95% CI, 17.26-408.69, respectively). Presenting for STD-related reasons (STD symptoms or requesting an STD check) had a negative association with HIV seroconversion (adjusted OR, 0.07; 95% CI, 0.01-0.90) compared with presenting for the sole purpose of requesting an HIV test. CONCLUSIONS: Repeat HIV testing is common among patients receiving services at an STD clinic. The role of repeat testing in HIV prevention efforts is complex and poorly understood. Results from this study could be used to identify and target those testing previously at highest risk for contracting HIV for risk-reduction interventions. PMID- 15273575 TI - Quality of sexually transmitted infection clinical management and contact tracing outcomes in a remote area of high sexually transmitted infection endemicity. AB - OBJECTIVES: The objectives of this study were to assess health professionals' adherence to Western Australian sexually transmitted infection (STI) management guidelines and to document the outcomes of contact tracing in a remote Indigenous setting. METHODS: This article comprises 2 parts: a retrospective clinical audit of quality of clinical STI management and outcomes of contact tracing and an analysis of completeness of relevant laboratory investigations. RESULTS: Documented clinical STI management of index cases varied from 94% receiving treatment in accordance with the Guidelines, whereas only 48% underwent a clinical examination. Sexual contacts who underwent STI consultation had concordant (30%) and discordant (17%) STI(s). The proportion of patients with STI(s) in whom all appropriate laboratory investigations had been requested increased from 25% in 1998 to 9% to 49% in 2001-2002. CONCLUSIONS: This study demonstrates that quality of clinical STI management comparable to that observed overseas is possible despite the challenges of healthcare delivery in a remote setting. PMID- 15273577 TI - Receptivity for probiotic products among premenopausal female students in an African university. AB - OBJECTIVE: The objective of this study was to examine the receptivity for probiotic products among premenopausal female students in an African university. GOAL: The goal of this study was to determine the local knowledge in Nigeria of probiotics and the willingness of young women to use them should they be introduced. STUDY: Closed-ended questionnaires were administered to a sample of 280 participants and these addressed age, marital status, perceived risk of HIV infection for the next 3 years, and history of urogenital infections. The participants were also asked whether they would welcome a probiotic product in oral/vaginal form and in milk-based food products, willingness to purchase and use, how often they would use these products, preference of form, price, and where they would like to buy the products. The second questionnaire was open ended. It asked the participants to freely list any concerns or worries they had in relation to probiotic products. RESULTS: Of the 280 participants, 55.3% indicated that they believed they were at risk of acquiring HIV within the next 3 years, illustrating the enormity of the problem in Africa and the feelings among women that they cannot easily control sexual relationships and have partners use condoms. Eighty-two percent of the subjects stated they would welcome probiotic products in capsular form for vaginal instillation or to be taken orally to improve vaginal health. Over one third (36%) of women indicated they would be willing to use the probiotic products as part of their daily self-care. One hundred nine (39.6%) respondents were willing to purchase the probiotic products at a reasonable price of 0.08 US dollars per dose, whereas 71.5% were willing to pay up to 0.38 US dollars. Some subjects (25%) raised some concern over safety of probiotics. CONCLUSIONS: The findings revealed that female university students are receptive to probiotic products in Nigeria and indicate strongly a need to consider women's concerns about urogenital health. Furthermore, the study identified a need for appropriate educational materials about probiotics, including benefits and safety information, in an African country suffering severely from the HIV epidemic. PMID- 15273578 TI - Assay for establishing whether microbicide applicators have been exposed to the vagina. AB - OBJECTIVES: To develop an accurate, rapid, and inexpensive method for verifying vaginal applicator use. GOAL: To develop a method for assessing compliance in microbicide clinical trials. STUDY DESIGN: Single use Microlax applicators containing a placebo formulation either were or were not exposed to the vagina. Three assays were developed to determine whether the applicators had been used vaginally. RESULTS: Blinded examiners were able to discern 63% of the time whether or not applicator tips had been exposed to the vagina. Optical density (to measure lactobacilli), increased in media exposed to used applicators but not in media exposed to unused applicators. When tips of applicators were stained with trypan blue, used applicators could be distinguished easily from unused applicators. CONCLUSION: Staining of applicator is accurate, simple, rapid, and inexpensive. This method could be be used in clinical settings in the developing world. Dying applicator tips could prove useful in excluding non-compliant subjects, analyzing data, or developing social intervention strategies to improve compliance. PMID- 15273579 TI - Is HIV/sexually transmitted disease prevention counseling effective among vulnerable populations?: a subset analysis of data collected for a randomized, controlled trial evaluating counseling efficacy (Project RESPECT). AB - OBJECTIVE: The objective of this study was to evaluate counseling efficacy among high-risk groups. STUDY: We conducted a subset analysis of data collected from July 1993 through September 1996 during a randomized, controlled trial (Project RESPECT). Participants (n = 4328) from 5 public U.S. sexually transmitted disease (STD) clinics were assigned to enhanced counseling, brief counseling, or educational messages. For 9 subgroups (sex, age, city, education, prior HIV test, STD at enrollment, race/ethnicity, injection drug use, exchanging sex for money/drugs), we compared STD outcomes for those assigned either type of counseling with STD outcomes for those assigned educational messages. RESULTS: After 12 months, all subgroups assigned counseling (brief or enhanced) had fewer STDs than those assigned educational messages. STD incidence was similar for most subgroups assigned enhanced or brief counseling. All subgroups had an appreciable number of STDs prevented per 100 persons counseled, especially adolescents (9.4 per 100) and persons with STD at enrollment (8.4 per 100). CONCLUSIONS: HIV/STD prevention counseling (brief or enhanced counseling) resulted in fewer STDs than educational messages for all subgroups of STD clinic clients, including high-risk groups such as adolescents and persons with STDs at enrollment. PMID- 15273580 TI - An opportunity for prevention: prevalence, incidence, and sexual risk for HIV among young Asian and Pacific Islander men who have sex with men, San Francisco. AB - OBJECTIVES: We investigated HIV prevalence and incidence and related risk factors among young Asian and Pacific Islander (API) men who have sex with men (MSM). DESIGN: We conducted a cross-sectional survey. METHODS: A venue-based sample of 496 young API MSM in San Francisco completed face-to-face questionnaires and received HIV counseling and testing. RESULTS: HIV prevalence was 2.6% and annualized HIV incidence was 1.8% per year. In multivariate analysis, being American-born, having 51+ lifetime sex partners, and having attended a "circuit party" (multiday large MSM gatherings) were associated with HIV infection. Forty seven percent of the sample reported unprotected anal intercourse in the past 6 months. CONCLUSIONS: The current levels of HIV prevalence, HIV incidence, and sexual risk behavior suggest an emerging HIV epidemic among young API MSM in San Francisco. PMID- 15273581 TI - Sexual behavior is more risky in rural than in urban areas among young women in Nyanza province, Kenya. AB - BACKGROUND: HIV/sexually transmitted disease interventions in sub-Saharan Africa have largely been focused on urban areas, where sexual behavior is supposed to be more risky than in rural areas. GOAL: The goal of this study was to measure sexual risk behavior among young adults in Nyanza province in Kenya and to compare rural and urban areas. STUDY: In a cross-sectional study, 584 household members aged 15 to 29 years in Kisumu town and the rural districts Siaya and Bondo were selected by multistage random sampling and were administered a face-to face questionnaire. RESULTS: For women, sexual behavior was more risky in rural than in urban areas, also after adjusting for sociodemographic differences. Rural women reported less frequently being a virgin at marriage, a higher number of lifetime partners, and less consistent condom use with nonspousal partners. For men, sexual risk behavior was equally high in urban and rural areas. CONCLUSIONS: The potential for further HIV spread in rural Nyanza is large. HIV/sexually transmitted disease interventions should be expanded from urban to rural areas in Nyanza. PMID- 15273582 TI - Personal digital assistants used to document compliance of bacterial vaginosis treatment. AB - OBJECTIVE: The objective of this study was to estimate patient compliance with oral and vaginal metronidazole treatment of bacterial vaginosis using personal digital assistants (PDAs) and paper diaries. GOAL: The goal of this study was to assess a novel compliance documentation approach. STUDY: After each dose of intravaginal or oral metronidazole, 71 subjects recorded the time on a paper diary and answered questions on a PDA. All PDA entries were unknowingly time-date stamped. Subjects returned for 2- and 6-week posttreatment examinations. Compliance was calculated using a repeated-measures multivariate analysis of variance (ANOVA). RESULTS: Mean patient compliance rates within the oral metronidazole group were greater with the paper diary compared with the PDA (68.3% and 50.0%, respectively, P = 0.001). The observed rate of compliance agreement for PDA versus paper diary was 69.0% (kappa = 0.4). The majority of noncompliant subjects reported they were compliant with the PDA and paper diary. CONCLUSIONS: PDAs could more accurately document true compliance rates and could be reasonable instruments to assess compliance in intravaginal antimicrobial drug or contraceptive trials. PMID- 15273583 TI - Unprotected anal intercourse associated with recreational drug use among young men who have sex with men depends on partner type and intercourse role. AB - OBJECTIVE: The objective of this study was to measure associations of unprotected anal intercourse (UAI) and substance use by sexual partner (regular vs. casual) and role [insertive (I) vs. receptive (R)]. GOAL: The goal of this study was to identify determinants of the association of specific drugs and UAI. STUDY: We conducted a prospective study of young men who have sex with men (MSM), 1997 2002. Odds ratios (ORs) for association of substance use and UAI during the previous year were adjusted for age and calendar year. RESULTS: UAI was significantly associated with sexual situation-specific use of marijuana (OR, 1.43), crystal methamphetamine (OR, 1.75), ecstasy (OR, 1.88), and ketamine (OR, 2.17); global use associations were similar. Situation-specific associations with alcohol (OR, 1.93) and gamma-hydroxybutyrate (GHB; OR, 1.98) were not seen with global measures. GHB and ketamine were specifically associated with IUAI with regular partners, and methamphetamine with RUAI with casual partners. CONCLUSION: Type of drug use measure, partner, and role are important determinants of the association of specific substances and UAI. PMID- 15273584 TI - Comparison of first void urine and urogenital swab specimens for detection of Mycoplasma genitalium and Chlamydia trachomatis by polymerase chain reaction in patients attending a sexually transmitted disease clinic. AB - OBJECTIVE: The objective of this study was to compare urogenital swab specimens and first void urine (FVU) specimens from male and female patients at a sexually transmitted disease clinic for the detection of Mycoplasma genitalium and Chlamydia trachomatis infections using in-house, inhibitor-controlled polymerase chain reaction (PCR). STUDY DESIGN: Urethral swabs and FVU were collected from 1856 men and 753 women who also had a cervical swab collected. A positive diagnosis of infection was made if any 1 of the specimens tested positive and were confirmed in a second PCR assay targeting independent genes. RESULTS: M. genitalium DNA and C. trachomatis DNA were detected in 126 (6.8%) and 246 (13.3%) of the male sample sets and in 51 (6.8%) and 73 (9.7%) of the female specimen sets, respectively. Using our in-house PCR and sample preparation methods, FVU was found to be the most sensitive diagnostic specimen for both pathogens, but for optimal sensitivity, it should be supplemented with a cervical specimen in women. In a small subset of female FVUs, storage at -20 degrees C led to false negative M. genitalium PCR results in 27% of specimens found positive when a sample preparation was performed before freezing. The age-specific prevalence of M. genitalium in men was almost constant between 18 and 45 years of age in contrast to C. trachomatis infections, which were more common in younger men. CONCLUSION: Urine appeared to be a better diagnostic specimen than the urethral swab for M. genitalium and C. trachomatis detection by PCR in this cohort of sexually transmitted disease clinic attendees but should be supplemented with a cervical specimen in women. PMID- 15273585 TI - Development and use of a type-specific antibody avidity test based on herpes simplex virus type 2 glycoprotein G. AB - OBJECTIVES: It is difficult to discriminate between lesions resulting from recently acquired versus established genital herpes simplex virus type 2 (HSV-2) infection. Methods not based on history or serum IgM status are needed. GOAL: Our goal was to use type-specific gG-2 antibody avidity determinations based on HerpeSelect HSV-2 enzyme-linked immunosorbent assay (ELISA) to identify new infections. STUDY: Sera (N = 168) from 71 patients with first-episode genital herpes and 45 sera from 21 patients with recurrent episodes were tested. RESULTS: Median avidity increased from 30.2 in sera drawn 6 weeks after infection (P <0.001). Patients with recurrent episodes and established HSV 2 infections (median, 6.1 years' duration) had higher avidity antibodies (median, 92.7; range, 55.1-100) than patients after first episodes (median, 33.7; range, 6.4-73.9; P <0.001). CONCLUSION: Avidity testing based on HerpeSelect ELISA could be a cost-effective method to identify patients with new HSV-2 infections. PMID- 15273586 TI - The sixth right of medication administration: right response. PMID- 15273587 TI - Commissioning ceremony: "I want to belong...I pledge...I accept". PMID- 15273588 TI - Creative collaborative clinical quickies. PMID- 15273589 TI - Dear Florence: tips and strategies for faculty. PMID- 15273590 TI - Clinical preparation for beginning nursing students: an experiential learning activity. PMID- 15273591 TI - A clinical strategy to help students with leadership/management NCLEX questions. PMID- 15273592 TI - Lights, camera, action: using feature films to stimulate emancipatory learning in the RN to BSN student. AB - Nurse educators are continually challenged to develop and implement effective activities to stimulate reflective learning in the RN to BSN student. The authors discuss the successful use of the feature film My Life as a reflective learning activity for a family health systems course.While feature films have been used constructively to teach family systems and social development, there is scant literature on the use of feature film as a teaching strategy within the discipline of nursing. The authors present evidence of how a film promoted stimulating and powerful transformative learning. PMID- 15273593 TI - A qualitative tool for critical thinking skill development. AB - Quantitative measures were not meeting the needs of faculty for evaluating critical thinking among Baccalaureate of Science Nursing students. The authors discuss the critical thinking self-reflection tool that was developed for teaching critical thinking and qualitatively evaluating changes over time with established interrater reliability and content validity. The tool is consistent with the concepts of critical thinking in the curriculum and was implemented systematically in all clinical nursing courses. PMID- 15273594 TI - Writing Across the Curriculum: using Healthy People 2010 and the DHHS Secretary's Award for Innovations in Health Promotion and Disease Prevention. AB - The authors detail a Writing Across the Curriculum (WAC) exercise that combines the Department of Health and Human Services' Secretary's Award for Innovations in Health Promotion and Disease Prevention (Secretary's Award) and the DHHS document Healthy People 2010. The authors discuss the writing competition as a way to encourage innovative problem solving and provide curricular instructions for using Healthy People 2010 and the Secretary's Award as a WAC exercise. PMID- 15273595 TI - Assessing student critical thinking through online discussions. AB - Critical thinking is integral to nursing practice and education. The introduction of Internet-based and Internet-assisted instruction provides an opportunity to explore critical thinking with nursing students in a new format. The author reports on a hybrid or blended Internet-assisted course structure that incorporates online discussion and a new method for assessing student critical thinking. PMID- 15273596 TI - Enhancing diagnostic reasoning skills in nurse practitioner students: a teaching tool. AB - The Advanced Health Assessment course provides an opportunity to focus on critical thinking and diagnostic reasoning skills as students transition to the nurse practitioner role. The authors describe a structured teaching tool designed for faculty to use in facilitating diagnostic reasoning and clinical judgment. This tool gives faculty a format to use in the classroom or online for helping students decide which data to collect and which data are important in developing hypothesis driven differential diagnoses. PMID- 15273597 TI - Munchausen syndrome and Munchausen syndrome by proxy in a student nurse. AB - Most faculty are not prepared for the possibility of encountering Munchausen syndrome (MS) in nursing students and Munchausen syndrome by proxy (MSP) in nursing students' children. When confronted with MS or MSP, their first reaction is often hostility coupled with a sense of betrayal. Given that individuals with this condition are attracted to helping professions, the authors describe both conditions in a case in which a nursing student presented with MS and the student's daughter was a victim of MSP. The focus is on protection of any children and the public, psychiatric treatment for the offender, and assistance for faculty. PMID- 15273598 TI - Learning nursing research through faculty-mentored projects. AB - Baccalaureate nursing educators have incorporated nursing research content into the curriculum for many years. Our university also proposes that students have a faculty-mentored learning experience at least once during their undergraduate education. Student research, guided by a faculty mentor, provides an excellent opportunity for students to learn about and participate in nursing research. The authors describe a unique experience of nursing students working as research assistants in a faculty research project using a human patient simulator. PMID- 15273599 TI - Health literacy: a concern for case managers. PMID- 15273600 TI - Triumph and tragedy: using physicians to impact length of stay, part I. PMID- 15273601 TI - Enhancing the role of case managers with specialty populations: development and evaluation of a palliative care education program. AB - Palliative home care is an important component of the care system for patients at the end of life and case management is considered an essential element of the Canadian home care system. Case managers play a critical role in allocating resources, thus influencing the costs and the viability of palliative home care. Case management education programs focused on care coordination with specialty palliative care populations are nonexistent. An education program targeted at improving the knowledge and skills of case managers in allocating resources to palliative care populations was developed and pilot-tested in a metropolitan Canadian city home care program. Core curriculum was based on an initial learning needs assessment and used case-based problem solving to enhance case-management skills. An improvement in knowledge was noted on posttests and case managers described increased comfort and confidence in their role as case managers to this patient population. Home care organizations caring for palliative care populations must ensure case managers are prepared for case management roles with specialty populations if the home is to be rendered an appropriate and viable care setting for patients at the end of life. PMID- 15273603 TI - The relationship between diabetes and depression: improving the effectiveness of case management interventions. AB - Diabetes places an immense burden on U.S. healthcare delivery systems. The Centers for Disease Control and Prevention estimates the economic impact of diabetes at nearly 100 billion dollars a year. Diabetes affects 17 million people in the United States and ranks as the sixth leading cause of death. Diabetes is among the most psychologically and behaviorally demanding chronic medical illnesses and the presence of diabetes doubles the odds of comorbid depression. In addition, recent research demonstrates that depressive symptoms serve as a predictor of type 2 diabetes. Case managers should maintain a high index of suspicion for the possibility of depression in chronically ill clients, as the severity of depressive symptoms is associated with poorer treatment adherence and higher healthcare costs. Three valid and reliable tools available to support depression screening in the case management setting are presented. PMID- 15273604 TI - An interdisciplinary, evidence-based process of clinical pathway implementation increases pathway usage. AB - Clinical pathways have been implemented in many healthcare settings as a link between evidence and practice. Most published research concludes that when clinical pathways are implemented and used by health professionals, there is a positive impact on health outcomes. However, some research also suggests that utilization of clinical pathways by health professionals is low and that implementation strategies for linking evidence with clinical practice often prove to be weak or ineffective. This paper describes a before and after study to determine whether an interdisciplinary, genuinely collaborative, and evidence based process of clinical pathway implementation resulted in increased documented use of an acute myocardial infarction (AMI) clinical pathway by health professionals in a regional Australian hospital. Underpinning the design and implementation process was the belief that true team involvement would lead to ownership, acceptance, and, ultimately, to increased usage of the pathway. Documented clinical pathway usage was measured in two ways: (1) the presence of the AMI clinical pathway in the medical records of patients diagnosed with an AMI and (2) the proportion of the AMI clinical pathway completed when it was present in the medical record. A total of 195 medical records of those diagnosed with an AMI were audited before (n = 124) and after (n = 71) the implementation process. The interdisciplinary, truly collaborative, and evidence-based implementation process resulted in a statistically significant increase in documented usage of the AMI pathway (22.6% vs. 57.7%; p <.000). Results indicate that involvement of key users in the design and implementation of a clinical pathway significantly increases staff utilization of the document. PMID- 15273605 TI - Development of a training module on therapeutic boundaries for mental health clinicians and case managers. AB - This article describes the development of a training model to teach mental health clinicians and case managers about therapeutic boundaries. Awareness of boundary transgressions is vital for establishing and maintaining the moral and ethical integrity of mental health treatment. Our structured teaching model was presented to staff from many different treatment settings throughout our mental health organization. A major portion of each session was utilized for exploration of common boundary dilemmas in the clinical setting as they relate to current policies and ethics codes. Efforts were made to mention the variety of boundary transgressions that may occur in various settings. Participants reported a broadened perspective in their work with clients, having learned about types of boundary issues that they had not previously recognized. Maintaining an understanding of boundary issues among staff requires ongoing educational efforts in training. The health organization's policies on boundaries must be clear and specific to guide the staff. Although training was given in a mental health setting, these lessons can be extended beyond the mental health arena. Safe therapeutic boundaries must be practiced in all settings. PMID- 15273606 TI - Back to basics: "the four Rs". PMID- 15273607 TI - Congenital heart disease: an overview of the condition and treatment options. PMID- 15273608 TI - Negligence, a new twist to an old issue. PMID- 15273610 TI - Computer-aided diagnosis and the evaluation of lung disease. PMID- 15273611 TI - Flat panel detector-based volumetric CT: prototype evaluation with volumetry of small artificial nodules in a pulmonary phantom. AB - PURPOSE: To evaluate amorphous silicone-based flat panel detector volumetric CT (VCT) in volumetric assessment of small nodules in a pulmonary phantom, and to perform comparative experiments with 4-row multislice CT (MSCT). MATERIALS AND METHODS: Seventy synthetic nodules (volume range (VR): 0.99-185.77 mm; estimated diameter range (ED): 1.4-7.8 mm) were scanned in spherical shape and after iso volumetric deformation with VCT and MSCT using 0.63 mm (MSCT I) and 1.25 mm (MSCT II) collimations. Measured volumes and percent measurement errors (PME) were compared between the 3 CT modes before and after nodule deformation. For each measurement pair before and after deformation, the post-deformation relative volumetric inaccuracy (RIA) was determined. Volume, PME, and RIA differences were tested using Wilcoxon and Friedman methods. RESULTS: The volumes of the smallest nodules (VR = 0.99-2.83 mm, ED = 1.4-1.9 mm) were computable only from VCT scans. In VCT, measured volumes and PMEs before and after deformation differed significantly less compared with MSCT (VCT: P = 0.06 and 0.56, respectively; MSCT I: P = 0.0012 and 0.006, respectively; and MSCT II: P < 0.0001 for measured volumes and PMEs). In VCT PMEs of 5.51-32.21 mm nodules (ED = 2.4-4.1 mm) before and after deformation were significantly below MSCT (VCT averages = 1.43-1.91% and 1.98-3.48%, for spherical and deformed nodules, respectively; MSCT I averages = 9.97-26.1% and 12.16-38.10%, respectively; MSCT II averages = 17.79-46.18 and 18.14-54.66%, respectively, P < 0.0001) and RIAs in VCT were significantly below MSCT (VCT: 0.50-2.62%, MSCT I: 3.35-15.97%, and MSCT II: 4.29-18.46%; P = 0.0001 0.0039). CONCLUSION: VCT volumetry is highly accurate in volumetry of smallest nodules with estimated diameters of 1.4-4.1 mm. PMID- 15273612 TI - Trends in thoracic radiology over a decade at a large academic medical center. AB - OBJECTIVE: To investigate thoracic radiology usage over and above the secular trends associated with hospital-wide changes in the number of patients over a decade. MATERIALS AND METHODS: We retrospectively reviewed administrative data from our 905-bed tertiary-care hospital between January 1, 1992, to December 31, 2001. Three points of entry to the radiology department were identified: inpatient (IP), outpatient (OP), and the emergency room (ER). The total numbers of patients, imaging studies, chest radiographs, chest CTs, CTs for pulmonary embolism, pulmonary angiograms, ventilation/perfusion scintigrams (V/Qs), lung biopsies, cardiac and chest MRIs, and FDG-PET scans for lung nodules and masses were collected. The significance of trends using linear regression analysis was evaluated. RESULTS: IP and OP numbers have significantly increased over a decade (P = 0.04 and P = 0.01 respectively); ER patient numbers have not. There has been an increase in the ratio of chest radiographs per patient arising from the ER area (P = 0.0002). All 3 areas demonstrated an increase in the ratio of chest CTs per patient: IP (P = 0.0002), OP (P = <0.0001), and ER (P = <0.0001). IP and ER areas demonstrated an increase in the ratio of CTs for pulmonary embolism per patient (P = 0.006, P = 0.04 respectively). There was a decrease in the ratios of pulmonary angiograms and V/Qs per IP (P = 0.02 & P = 0.0003 respectively). Cardiac MRIs per patient demonstrated an increase (IP P = 0.01, OP P = 0.02). FDG PET for lung nodules and masses per patient demonstrated an increase in IP (P = 0.03) and OP (P = 0.003) areas. The total number of chest imaging studies divided by the total number of imaging studies demonstrated an increase in IP and ER areas (P = 0.02 and P = 0.02 respectively). CONCLUSION: There has been an increase in thoracic radiology usage above secular trends, particularly in the regions of chest CT and FDG-PET. CT is replacing more traditional techniques to diagnose pulmonary embolism for inpatients. PMID- 15273613 TI - HRCT findings of proximal interruption of the right pulmonary artery. AB - The purpose of this study is to present the characteristic HRCT findings of the lung parenchyma in patients with proximal interruption of the right main pulmonary artery. HRCT findings of proximal interruption of the right pulmonary artery demonstrated reticular opacities, septal thickening, subpleural consolidation, cystic lung changes, and pleural thickening in all 5 patients; bronchial dilation and bronchial wall thickening in 4 patients; and subpleural ground glass opacity (GGO) in 3 patients. The changes may be caused by absent pulmonary artery perfusion and development of systemic vessel collateralization. PMID- 15273614 TI - Ultrafast contrast-enhanced MR pulmonary perfusion: clinical applications and imaging findings. PMID- 15273615 TI - High resolution CT anatomy of the pulmonary fissures. AB - RATIONALE AND OBJECTIVES: Pulmonary interlobar fissures are important landmarks for proper identification of normal pulmonary anatomy and evaluation of disease. The purpose of this study was to define the radiologic anatomy of the pulmonary fissures using high resolution computed tomography (HRCT) in a large population. METHODS: HRCT of the lungs from aortic arch to diaphragm was performed in 622 patients, with a slice thickness of 1 mm and slice interval of 10 mm. Major, minor, and accessory fissures were studied for their orientation and completeness. RESULTS: Both major fissures were mostly facing laterally in their upper parts (100% and 89% right and left, respectively). The left major fissure faced medially (69%) while the right major fissure faced lateral (60%) in their lower parts. The right major fissure was more often incomplete (48% as compared with 43% on the left, P < 0.05). Minor fissures were convex superiorly with the apex in the anterolateral part of the base of the upper lobe, and were incomplete in 63% of cases. Azygos, inferior accessory, superior accessory, and left minor fissures were also seen in 1.2%, 8.6%, 4.6%, and 6.1% of the cases, respectively. CONCLUSION: The pulmonary fissures are highly variable and the right major fissure differs considerably from the left. The fissures are often incomplete. PMID- 15273616 TI - Thymic masses of the middle mediastinum: report of 2 cases and review of the literature. AB - Although thymic lesions are relatively common causes of anterior mediastinal masses, they can rarely arise in other mediastinal compartments, as it is well recognized that thymic tissue can lie in ectopic intrathoracic locations. A thymic mass within the middle mediastinum has rarely been reported, with only a single case of a thymic cyst described and no reports of a middle mediastinal thymoma. We report 2 thymic masses (1 thymoma and 1 thymic cyst) found to arise in the middle mediastinum. PMID- 15273617 TI - Granulomatous Pneumocystis carinii pneumonia in a non-AIDS patient: an atypical presentation. AB - We present the computed tomographic findings of pulmonary involvement by granulomatous Pneumocystis carinii pneumonia in a 73-year-old woman recently tapered from a high-dose long-term systemic corticosteroid therapy for Factor VII deficiency. PMID- 15273618 TI - Lymphocytic interstitial pneumonitis in a patient with systemic lupus erythematosus: radiographic and high-resolution CT findings. AB - Reports associating lymphocytic interstitial pneumonitis (LIP) and systemic lupus erythematosus (SLE) are exceedingly rare. This case demonstrates high-resolution CT (HRCT) findings of LIP including multiple thin-walled cysts throughout the lungs, which are known to occur in LIP, but have not been described in the radiologic literature in association with SLE. PMID- 15273619 TI - Fibrosing mediastinitis with peripheral airway dilatation and central pulmonary artery occlusion. AB - We present a case of fibrosing mediastinitis causing peripheral airway abnormalities on CT consisting of bronchial dilatation and wall thickening due to longstanding central pulmonary artery obstruction. The peripheral airway dilatation is similar in appearance to that which has been described in patients with chronic pulmonary embolism. PMID- 15273620 TI - Disseminated toxoplasmosis after bone marrow transplantation: high-resolution CT appearance. AB - A 16-year-old female patient, who had undergone bone marrow transplantation 35 days earlier, presented with dry cough, dyspnea, and fever for 4 days. Chest radiography showed poorly-defined bilateral opacities. High-resolution CT revealed bilateral ground glass opacities with superimposed septal thickening and intralobular linear opacities. Laboratory results were nonspecific and empiric treatment with multiple drugs was initiated. The patient had no response to therapy and died 12 days after the admission. At autopsy the patient had disseminated toxoplasmosis with involvement of the central nervous system, myocardium, bone marrow, and lungs. PMID- 15273621 TI - Notes from the 2003 annual meeting of the Korean Society of Thoracic Radiology. PMID- 15273624 TI - State of the society: Jaclyn Tropp MSN, CRNI, CFNP INS President 2003-2004. PMID- 15273626 TI - Chief Executive Officer's Report: Mary Alexander, BS, CRNI. PMID- 15273627 TI - Presidential Address: Mary Walsh, BS, CRNI INS President 2004-2005. PMID- 15273628 TI - Test your knowledge: preparing to take the CRNI exam. AB - The CRNI Certification Examination consists of 200 multiple-choice questions covering the nine core content areas of infusion nursing: Technology and Clinical Applications, Fluid and Electrolyte Balance, Pharmacology, Infection Control, Pediatrics, Transfusion Therapy, Antineoplastic Therapy, Parenteral Nutrition, and Quality Assurance. The review questions provided below are modeled on the CRNI exam and are intended to help exam candidates test their knowledge of infusion therapy practice. This special section is a regular addition to the Journal of Infusion Nursing, with each edition focusing on a single content area. PMID- 15273629 TI - Consideration of barrier protection and latex protein allergy in the evaluation of medical gloves. AB - The issues surrounding medical gloves have become increasingly complicated over the past 15 years. The AIDS epidemic, allergies, needlesticks, new glove materials, and, most recently, an increased risk of biohazards all factor into the choice of medical gloves. This article examines the research available on two of the issues affecting medical glove choice: barrier protection and latex protein allergy. PMID- 15273630 TI - A nursing process model: quantifying infusion therapy resource consumption. AB - Comprehensive measurement of the effort associated with a procedure, treatment, or project involves a myriad of planned observations and analyses. When accurately synthesized, these measurements show the amount of resources or the level of effort required for the intervention. In an era of unprecedented healthcare cost scrutiny, it is imperative for clinicians to understand and apply these components and methods to program development, budget management, staffing, and cost-to-outcome analysis. Through a series of on-site observations and interviews at in-office infusion centers providing a nonchemotherapeutic biologic therapy, specifically infliximab (Remicade), the investigators were able to isolate and assign value to the multiple factors that contribute to the cost of this procedure. The investigators also were able to establish a process model for clinicians who are inevitably involved in the "business" of healthcare. PMID- 15273631 TI - The impact of safety product use on catheter-related infections. AB - This article provides an overview of the issues that affect the use and proliferation of safety infusion products. In particular, the associated risks and benefits of needleless infusion systems are discussed. Recent legislation and regulations address healthcare worker exposure to bloodborne pathogens and mandate the use of these devices to mitigate the risk of healthcare worker exposure to bloodborne pathogens. This article examines whether safety devices increase the risk for catheter-related infections among patients, and evaluates the implications for clinical practice and compliance. Clinician education and standards of care also are discussed as methods to ensure the optimal safety of both healthcare workers and the patients for whom they are responsible. PMID- 15273632 TI - Safety and cost-effectiveness of paclitaxel administered as a 1-hour infusion versus a 3-hour infusion for various malignancies. AB - This study challenges the current practice of administering paclitaxel for a variety of malignancies over 3 hours and documents the safety and cost effectiveness of 1-hour administration in the outpatient setting. The authors investigated opportunities to save nursing time and costs in a cancer clinic without compromising patient safety. These savings are referred to as "opportunity-cost savings" that enable the clinic to schedule more patients during the time normally required to administer a 3-hour paclitaxel dose. Over a 2-year period, the authors were able to document significant time savings with no increase in adverse drug reactions. PMID- 15273633 TI - Comparison of hemodynamic and biochemical effects of furosemide by continuous infusion and intermittent bolus in critically ill patients. AB - Positive fluid balance in critically ill patients is a common problem in the intensive care unit (ICU) often associated with a poor outcome. In addition, clinically important changes in hemodynamic variables have been found to occur after diuretic therapy. This study was conducted to evaluate the safety and relative effectiveness of two diuretic protocols in the ICU. Twenty-two patients in the medical ICU with pulmonary edema or fluid overload and PaO2/FIO2 pressure less than 300, were randomized to diuretic therapy by either continuous infusion or intermittent bolus. Hemodynamic and biochemical measurements were recorded. Protocol-guided diuretic management can be readily and safely implemented in the ICU. Although both continuous and bolus diuretic regimens appear to be equally effective in achieving negative fluid balance, the clinician may consider a continuous infusion of furosemide in the hemodynamically and electrolytically unstable patient to ensure more controlled diuresis with less hemodynamic and electrolyte alteration. From a nursing perspective, a continuous infusion of furosemide is a more efficient means of drug delivery. PMID- 15273634 TI - Epidemiology, diagnosis, and treatment of patients with metastatic cancer and high-grade gliomas of the central nervous system. AB - Cancer is the hyperactive state of cell growth in which the multiplication and division of cells occur abnormally. Malignant cancer to the brain frequently begins and ends with the loss of self or quality of life. Cancer of the central nervous system can be in the form of a primary or secondary brain tumor commonly known as metastatic cancer. Primary brain tumors can be benign or malignant on the basis of the cell type or location within the brain. Metastatic cancer has a primary source of origin, from which it has traveled to the brain by direct extension (tumors arising from the skull or vertebral column), or most commonly by hematogenous spread (through the blood supply, lymphatic system, or cerebral spinal fluid). As the cancer grows, the individual can experience headache, seizures, or focal neurologic deficits, all impinging on quality of life. This article addresses malignant central nervous system cancer including metastatic cancer and malignant gliomas (anaplastic astrocytoma, grade III, and glioblastoma multiforme, grade IV). Epidemiology, diagnostic workup, treatment, and outcome also are reviewed. PMID- 15273636 TI - Current challenges and concepts in the preparation of root canal systems: a review. AB - Nickel-titanium rotary instruments are important adjuncts in endodontic therapy. This review attempts to identify factors that influence shaping outcomes with these files, such as preoperative root-canal anatomy and instrument tip design. Other, less significant factors include operator experience, rotational speed, and specific instrument sequence. Implications of various working length definitions and desired apical widths are correlated with clinical results. Despite the existence of one ever-present risk factor, dental anatomy, shaping outcomes with nickel-titanium rotary instruments are mostly predictable. Current evidence indicates that wider apical preparations are feasible. Nickel-titanium rotary instruments require a preclinical training period to minimize separation risks and should be used to case-related working lengths and apical widths. However, and despite superior in vitro results, randomized, clinical trials are required to evaluate outcomes when using nickel-titanium instruments. PMID- 15273638 TI - Microbiological evaluation of one- and two-visit endodontic treatment of teeth with apical periodontitis: a randomized, clinical trial. AB - The antimicrobial efficacy of endodontic procedures performed in one-visit (including a 10-min intraappointment dressing with 5% iodine-potassium-iodide) was compared with a two-visit procedure (including an interappointment dressing with calcium-hydroxide paste). Teeth with apical periodontitis (n = 96) were randomly assigned to either group. Root canal sampling and culturing were performed before and immediately after instrumentation, and after medication. Initial sampling demonstrated the presence of microorganisms in 98% of the teeth. Postinstrumentation sampling showed reduction of cultivable microbiota. Antibacterial dressing further reduced the number of teeth with surviving microbes. In the postmedication samples, residual microorganisms were recovered in 29% of the one-visit teeth and in 36% of the two-visit treated teeth. No statistically significant differences between the groups were discerned. It was concluded that from a microbiological point of view, treatment of teeth with apical periodontitis performed in two appointments was not more effective than the investigated one-visit procedure. PMID- 15273637 TI - Anesthetic efficacy of articaine for inferior alveolar nerve blocks in patients with irreversible pulpitis. AB - The purpose of this prospective, randomized, double-blind study was to compare the anesthetic efficacy of 4% articaine with 1:100,000 epinephrine to 2% lidocaine with 1:100,000 epinephrine for inferior alveolar nerve blocks in patients experiencing irreversible pulpitis in mandibular posterior teeth. Seventy-two emergency patients diagnosed with irreversible pulpitis of a mandibular posterior tooth randomly received, in a double-blind manner, 2.2 ml of 4% articaine with 1:100,000 epinephrine or 2.2 ml of 2% lidocaine with 1:100,000 epinephrine using a conventional inferior alveolar nerve block. Endodontic access was begun 15 min after solution deposition, and all patients were required to have profound lip numbness. Success was defined as none or mild pain (Visual Analogue Scale recordings) on endodontic access or initial instrumentation. The success rate for the inferior alveolar nerve block using articaine was 24% and for the lidocaine solution success was 23%. There was no significant difference (p = 0.89) between the articaine and lidocaine solutions. Neither solution resulted in an acceptable rate of anesthetic success in patients with irreversible pulpitis. PMID- 15273639 TI - Root canal treatment in a population-based adult sample: status of teeth after endodontic treatment. AB - The purpose of this study was to quantify treatment outcomes of endodontically treated teeth, in a representative, population-based adult sample. The "Florida Dental Care Study" was a prospective, longitudinal, cohort study of 873 subjects who had at least one tooth at baseline and who were 45 yr or older. An in-person interview and clinical examination were conducted at baseline, 24 months, and 48 months after baseline, with 6-monthly telephone interviews between those times. Dental record information was abstracted afterward. Seventy-five teeth had conventional root-canal therapy (RCT) performed and met the inclusion criteria of 1-yr of follow-up or extraction. The mean duration of follow-up after RCT was 24.8 months. The mean (SD) attachment loss (AL) on teeth receiving RCT was only 3.3 (1.4) mm. Permanent restorations were placed in 79% of teeth a mean of 4.4 months after the RCT. However, 12% of teeth did not have any restorative treatment after RCT. After RCT had been completed, 81% of teeth were retained and 19% were ultimately extracted. Taken as a whole, this community-based study of a representative sample of adults suggests a higher failure rate than reported from studies in highly controlled environments or insured populations. It also suggests that the length of time to initial restoration of RCT-treated teeth is less than optimal. RCT was almost never performed on teeth with advanced periodontal attachment loss. PMID- 15273640 TI - Coronal microleakage of three temporary restorative materials: an in vitro study. AB - The sealing properties of three temporary restorative materials, Cavit, IRM, and a polycarboxylate-based cement, Ultratemp Firm, were investigated in vitro. Standardized access cavities were prepared in 45, intact, extracted, human molars. The teeth were randomly assigned to three groups and the access openings filled with one of three temporary filling materials. In five teeth (negative control), no restorative material was placed but the preparations were coated entirely with sticky wax. The five teeth of the positive control group had no restorative material and no sticky wax applied. After thermocycling for 500 cycles (5-55 degrees C), the experimental teeth were dipped in molten sticky wax to the CEJ. The coronal enamel was subsequently coated with two layers of nail varnish, leaving an area of 1 mm around the filling material uncovered. The samples were then immersed in 2% methylene blue dye solution for leakage assessment. The teeth were sectioned and the greatest depth of dye penetration was recorded. Positive control sections exhibited complete dye penetration, whereas negative controls had none. There was no statistically significant difference in marginal leakage between Cavit, IRM, and Ultratemp Firm (p > 0.05). All materials leaked at the interface material-dentin, whereas some IRM specimens absorbed the dye into the bulk of the material. PMID- 15273641 TI - Q-switched versus free-running Er:YAG laser efficacy on the root canal walls of human teeth: a SEM study. AB - Twenty-one teeth with one root canal were prepared by the step-back technique, divided into three groups, and split longitudinally. Group A served as a control. In group B, 20 to 150 pulses of 100 micros, 30 to 70 mJ per pulse at 1 to 4 Hz from a free-running Er:YAG laser were applied to the root-canal dentin. In group C, the Q-switched Er:YAG laser, with the same energy parameters and a 190-ns pulse duration was used. Scanning electron microscopy examination revealed that control specimens had debris and smear layer obscuring the dentinal tubules at all levels in the canals without crack formation. Both groups of laser-treated dentin were clean with opened dentinal tubules except around the lased area in which there was an intact smear layer. Cracks were observed in both laser groups with higher frequency in group C. In group B, craters with different depth levels at the root canal walls were produced and the energy apparently was distributed equally, because craters were well-shaped. In contrast, the ablation efficiency in group C was questionable with the parameters used in this study. Consequently, suitable parameters of the free-running Er:YAG laser must be found before its careful use as an adjunct in endodontic therapy. PMID- 15273642 TI - Effect of solvents on bonding to root canal dentin. AB - The long-term success of resin cementation of post/cores is likely increased with improvement in resin-root canal dentin bonding. The adverse effect of some irrigation constituents (NaOCl, H2O2) or medications (eugenol) on the bond strengths of resins to dentin have been reported. The purpose of this in vitro study was to evaluate the effect of two gutta-percha solvents (chloroform versus halothane) on microtensile bond strength to root canal dentin. Thirty, extracted, human, single-rooted teeth were instrumented to a #70 file and randomly divided into 3 groups of 10 each. The root canals were treated with water, chloroform, or halothane for 60 s. All root canals were obturated using C&B Metabond. After 24 h of storage in distilled water, serial 1-mm-thick cross-sections were cut and trimmed. Microtensile bond strength to apical, middle, and coronal root canal dentin were measured using an Instron machine. Using pooled data, the results indicated that water-treated roots had significantly higher resin-dentin bond strengths compared with chloroform or halothane treatment groups (control: 23.9 MPa; chloroform: 18.3 MPa; halothane: 17 MPa; p < 0.05). Gutta-percha solvents have an adverse effect on bond strengths of adhesive cements to root canal dentin. PMID- 15273643 TI - Lack of genotoxicity of formocresol, paramonochlorophenol, and calcium hydroxide on mammalian cells by comet assay. AB - Formocresol, paramonochlorophenol, and calcium hydroxide are widely used in dentistry because of their antibacterial activities in root canal disinfection. However, the results of genotoxicity studies using these materials are inconsistent in literature. The goal of this study was to examine the genotoxic potential of formocresol, paramonochlorophenol, and calcium hydroxide using mouse lymphoma cells and human fibroblasts cells in vitro by the comet assay. Data were assessed by Kruskal-Wallis nonparametric test. The results showed that all compounds tested did not cause DNA damage for the tail moment or tail intensity parameters. These findings suggest that formocresol, paramonochlorophenol, and calcium hydroxide do not promote DNA damage in mammalian cells and that the comet assay is a suitable tool to investigate genotoxicity. PMID- 15273644 TI - Nonsurgical endodontic treatment of dens invaginatus with large periradicular lesion: a case report. AB - The endodontic treatment of teeth with severe Type 3 dens invaginatus, characterized by an infolding of enamel and dentin, extending deep into the pulp cavity near the root apex, may be complicated and challenging. Because of the bizarre root canal anatomy and widely open apex, a combination of nonsurgical and surgical endodontic treatment or extraction is the most common choice of therapy. This article describes a nonsurgical endodontic treatment of a tooth with severe Type 3 dens invaginatus and an associated large periradicular lesion. After complete removal of the invaginated central mass of hard tissue and long-term calcium hydroxide treatment, nonsurgical endodontic treatment was performed. Complete healing of the periradicular lesion was observed at 25-month and 74 month follow-up examinations. PMID- 15273645 TI - Taurodontism in six molars: a case report. AB - This case report describes taurodontism in six molars of one patient. There were two mandibular second molars and four maxillary molars. All of them were hypertaurodont. PMID- 15273647 TI - Beating psychosis to the punch: the treatment options debate. PMID- 15273648 TI - Premorbid cognitive and behavioral functioning in military recruits experiencing the first episode of psychosis. AB - OBJECTIVE: To characterize the premorbid cognitive and behavioral abilities in apparently healthy adolescents who at a later time will be diagnosed with schizophreniform disorder or schizophrenia. BACKGROUND: Clarifying the pathological relationship between subtle intellectual and behavioral abnormalities and disease could provide markers for the early prediction of future psychosis. METHOD: Premorbid data on young male patients admitted to the Department of Psychiatry of the Central Military Hospital in Bucharest, Romania, between 1996 and 2002 and diagnosed with a first episode of psychosis or schizophreniform disorder were collected. The premorbid data consisted in the test scores of intellectual functioning and personality traits were collected by the Romanian Draft Board in order to assesses their aptitude to serve in the military. Premorbid cognitive and behavioral scores of male patients (cases=157) were compared with the scores of healthy male individuals (non-cases=169) matched for age, education, and geographic area of residence. The tests were administered when subjects were 18 years of age (initial screening) and the entire assessment was completed and concluded when subjects reached 20 years of age (actual conscription). RESULTS: As a group, apparently healthy males later admitted for a first episode of psychosis or schizophreniform disorder, obtained lower (worse) scores on the Raven Progressive Matrices test and on relevant personality traits in comparison to controls. CONCLUSION: The results add to the accumulating body of evidence suggesting that aspects of schizophrenia manifest years before the illness is formally diagnosed. Despite these results, more studies are needed to improve the diagnosing specificity and predictive value of the premorbid cognitive and behavioral manifestations, before they can be used as markers in models of primary or secondary prevention. PMID- 15273649 TI - Prodromal schizophrenia: the dilemma of prediction and early intervention. PMID- 15273650 TI - The use of pharmacotherapy in the prodrome of schizophrenia. AB - In treating schizophrenia there has been a shift in focus, with more attention being paid to early intervention based on the notion that effective treatment at this point can improve outcome. Most of this work has centred on pharmacotherapeutic interventions during the first psychotic break. More recently, attention has turned to the potential value of intervening even earlier, that is during the so-called "prodrome" that has been identified as predating the first psychotic break by as much as 4-5 years. We now have a limited number of published reports addressing this topic and these are reviewed here. PMID- 15273651 TI - The diagnosis and assessment of individuals prodromal for schizophrenic psychosis. AB - Accurate identification of individuals in the earliest symptomatic stages of psychosis offers new hope for developing more effective treatment strategies. Recently, research clinics have been set up to identify and possibly treat individuals who are seen as being at ultra-high risk of developing a psychotic disorder. This article reviews and describes measures that are currently being used and further developed in order to determine both the diagnostic criteria for the prodromal state and the severity of these putatively prodromal symptoms. PMID- 15273652 TI - Identifying vulnerability markers in prodromal patients: a step in the right direction for schizophrenia prevention. AB - Research has shown that many of the long-term deficits that are observable in schizophrenia populations are present prior to the emergence of psychotic symptoms. Recent research suggests schizophrenia has a "prodromal" period, whereby significant changes from premorbid functioning can be observed. Accurate classification of this period could have far-reaching implications for schizophrenia prevention. This article aims to provide an indepth evaluation of the perceived benefits of vulnerability marker research in this unique phase. It is hoped that identification of such markers may improve the predictive potency of prodromal criteria, and perhaps pave the way for future screening and primary prevention strategies. PMID- 15273653 TI - Severity of obsessive-compulsive symptoms is related to self-directedness character trait in obsessive-compulsive disorder. AB - OBJECTIVE: The present study examined the psychobiological Temperament and Character model of personality on obsessive-compulsive disorder (OCD) patients, as well as the relation of temperament and/or character dimensions on the severity of obsessive-compulsive symptoms. METHODS: Fifty-four subjects diagnosed with OCD, were assessed with the Temperament and Character Inventory, the Yale Brown Obsessive-Compulsive scale and the Hamilton Rating Scales for depression and anxiety. RESULTS: Compared with controls, OCD subjects displayed increased harm avoidance and lower self-directedness and cooperativeness. Low self directedness and high Hamilton depression scores were associated with increased severity of obsessive-compulsive symptoms. CONCLUSION: The Temperament and Character profile of OCD patients characterized in the present study was in agreement with previous reports using the same personality model and can be linked to some of their behavioral features. Furthermore, our data provides support of the influence that some personality traits may have on the severity of OCD symptoms. PMID- 15273654 TI - Local injection of receptor tyrosine kinase inhibitor MAE 87 reduces retinal neovascularization in mice. AB - PURPOSE: Retinal neovascularization occurs under the influence of angiogenic factors like vascular endothelial growth factor (VEGF). VEGF signaling is enhanced by insulin-like growth factor-1 (IGF-1). In vitro, the oxoindolinone MAE 87 inhibits angiogenic signal transduction by blocking tyrosine kinase receptors including VEGF receptor 2 (VEGFR-2), IGF-1R, fibroblast GF-1R and epidermal GFR. We investigated the effect of MAE 87 in vivo using the mouse model for oxygen induced retinopathy. METHODS: From postnatal day seven (P7) on, C57BL/6J mice were kept in a 75% oxygen environment for five days. On postnatal day 12 (P12) they received an intravitreal injection of MAE 87 in one eye and control substance in the fellow eye. The animals were sacrificed by intracardial perfusion with fluorescein-dextran solution on P17. Retinal whole mounts were prepared and ischemic retinopathy was evaluated in 26 animals using a standardized retinopathy score. RESULTS: After a single intravitreal injection of MAE 87 there were significantly less angioproliferative changes (blood vessel tufts, extra-retinal neovascularization, and blood vessel tortuosity) than in the fellow eye (p=0.007). The median retinopathy score (maximal 13) for the MAE 87 treated eyes was 6 (25th percentile: 5; 75th percentile: 7) and 8 for the control eyes (25th percentile: 5; 75th percentile: 10). CONCLUSIONS: The tyrosine kinase inhibitor MAE 87 may be a promising substance for local treatment of retinal neovascularization. Due to its ability to inhibit not only the VEGF but also the IGF-1 cascade, MAE 87 may prove especially valuable for the treatment of diabetic retinopathy. PMID- 15273655 TI - Fibroblast growth factor 2: roles of regulation of lens cell proliferation and epithelial-mesenchymal transition in response to injury. AB - PURPOSE: To examine the role of fibroblast growth factor 2 (FGF2) in regulating lens cell proliferation and epithelial-mesenchymal transition (EMT) in response to injury. METHODS: The amount of FGF2 protein was determined in healing, injured rat lenses by enzyme immunoassay. The effects of FGF2 and transforming growth factor beta2 (TGFbeta2) on cell proliferation of alphaTN4 cells (a mouse lens epithelial cell line) were determined. FGF2-knockout mice were used to further examine the role of endogenous FGF2 on injury-induced epithelial cell proliferation and EMT. The anterior lens capsule was injured by a hypodermic needle under both general and topical anesthesia in one eye of 34 fgf2+/+ mice and 42 fgf2-/- mice. At days 2, 5, and 10 post-injury the mice were sacrificed following a 2 h labeling period with bromo-deoxyuridine (BrdU). The number of BrdU-positive cells in each specimen was determined. RESULTS: A capsular break caused a 10 fold increase of FGF2 protein accumulated in rat lens 14 days after injury. Addition of 3.43 ng/ml FGF2 enhanced proliferation of alphaTN4 cells. This occurred in the presence or absence of exogenous TGFbeta2, that has an inhibitory effect on alphaTN4 cell proliferation. Significantly fewer BrdU labeled cells were found in fgf2-/- mice than in fgf2+/+ mice during healing post injury. However, lacking FGF2 did not alter the expression patterns of alpha smooth muscle actin and collagen type I, markers of EMT in lens cells. CONCLUSIONS: Endogenous FGF2 is required for increased cell proliferation but not essential for EMT during the lens response to injury. PMID- 15273656 TI - Null type of glutathione S-transferase M1 polymorphism is associated with early onset pterygium. AB - PURPOSE: To investigate the association of glutathione S-transferase M1 (GSTM1) null genotype with pterygium. METHODS: One hundred and twenty seven pterygium patients and 102 volunteers without pterygium were enrolled in this study. Polymerase chain reaction based analysis was used to resolve the GSTM1 null and positive genotypes. RESULTS: There was no significant difference between total pterygium and the control group. When stratified by the age, there was a significantly higher frequency of the GSTM1 null genotype in younger patients. The difference was not significant between patients and controls older than 60 years old. CONCLUSIONS: These results indicate GSTM1 null genotype is associated with early onset pterygium, but not associated with late onset pterygium. PMID- 15273657 TI - Atrial natriuretic peptide in the vitreous humor and epiretinal membranes of patients with proliferative diabetic retinopathy. AB - PURPOSE: Atrial natriuretic peptide (ANP) has been recently described as an endogenous inhibitor of the synthesis and angiogenic action of vascular endothelial growth factor (VEGF). Given VEGF's key role in promoting neovascularization in proliferative diabetic retinopathy (PDR), this study was designed to evaluate the possibility that ANP could be involved in the neovascular and fibrotic complications of PDR. METHODS: We determined ANP by radioimmunoassay in plasma and vitreous humor samples collected from diabetic patients with and without PDR and from non-diabetic subjects. ANP was also immunohistochemically localized in the epiretinal membranes of patients with PDR. RESULTS: Vitreous ANP concentrations were significantly higher in patients with active PDR compared to patients with quiescent PDR, diabetes without PDR or controls <0.05. Significant differences were also observed between vitreous ANP levels in diabetic patients without PDR and control subjects. There was no significant correlation between serum and vitreous ANP levels in any of the patient groups. ANP was detected in the fibrovascular epiretinal tissue of patients with PDR. CONCLUSIONS: Diabetic patients with active neovascularization have significantly higher levels of ANP in the vitreous humor than those without active PDR. Diabetic patients without PDR were also found to have significantly higher vitreous ANP levels than non-diabetic patients. Since plasma and vitreous ANP concentrations were found to be unrelated, we suggest intraocular ANP synthesis and/or an increase in the release of ANP into the vitreous, as opposed to diffusion from the blood, as the main factors contributing to the high vitreous ANP levels observed in diabetic patients. In the fibrovascular epiretinal tissue of these patients, ANP was found to be localized in vascular, glial, fibroblast-like and retinal pigment epithelium cells. Our findings suggest a role for ANP in PDR. PMID- 15273658 TI - Autofluorescence of transplantable hepatoma A22 (MH-A22): prospects of tumor tissue optical biopsy. AB - AIM: Autofluorescence of experimental tumor (hepatoma A22 (MH-A22)) was employed to discriminate the optical differences between necrotic and non-necrotic tumor, hemorrhagic tumor and healthy tissue. METHODS: The experiment was performed ex vivo using the transplantable tumor from the right haunch of hybrid mice (C57Bl/CBA). Blue LED light (lambda em=405 nm) was applied for autofluorescence excitation and fibre optics based spectrofluorimeter was used for spectra detection. RESULTS: We observed that necrotic tumor tissue is characterized by the absence of endogenous porphyrins fluorescence, and registered spectra do not possess differences in the red spectral region (600-710 nm) in comparison with normalized autofluorescence spectra of muscle. Moreover, only certain segments of non-necrotic tumor bear the fluorescence of endogenous porphyrins. CONCLUSIONS: Based on the experimental results, we suggest that the absence of long-waved fluorescence differences between necrotic tumor tissue and healthy tissue, e.g. muscle can impede the demarcation between healthy and tumor tissue. The uneven distribution of endogenous porphyrins in non-necrotic tumor tissue as well as the absence of endogenous porphyrins fluorescence in the small experimental tumors complicates the localization of cancerous tissue based on the autofluorescence registration. PMID- 15273659 TI - Caspases and cancer: mechanisms of inactivation and new treatment modalities. AB - Elimination of superfluous or mutated somatic cells is provided by various mechanisms including apoptosis. Deregulation of apoptotic signaling pathways may contribute to oncogenesis. Aspartate specific cysteine proteases, termed caspases are the key effector molecules in apoptosis. The aim of this review is to summarize the various defects in caspase-dependent cell death machinery identified in the neoplastic cells. These include not only mutations, but also alterations of gene methylation, and altered mRNA stability. Among the molecules that we discuss are elements of the extrinsic death pathway like CD95 (APO 1/Fas), FADD, FLIPs, FLICE, other apical caspases, components of the intrinsic apoptotic pathway like Apaf-1, caspase-9, and modulators of apoptotic pathways like IAPs, Smac/DIABLO, OMI/HtrA2, and other apoptosis regulating proteins. We also discuss recent data on cancer-specific agents that target effector mechanisms of apoptosis. Particular emphasis is given to the prospects for combining cell suicide-activating approaches with classical cancer therapies. PMID- 15273660 TI - Endocrine disrupting pesticides: a leading cause of cancer among rural people in Pakistan. AB - Evidence on the relationship between cancer and occupational exposure to pesticides and endocrine disrupting chemicals is reviewed. In animal studies it has been proved that majority of endocrine disrupting pesticides are carcinogenic. In humans, pesticides have been classified as carcinogens by the International Agency for Research on Cancer. Farmers may therefore be at higher risk for acute and chronic health effects associated with pesticides. Human data, however, are limited by the small number of studies that evaluate individual endocrine disrupting pesticide. Cancer of the breast, ovary, prostate, testis, and thyroid are hormone-dependent, which fostered research on the potential risk associated with occupational and environmental exposure to the so-called endocrine-disrupting pesticides. Professional as well as public exposure to pesticides raises cancer risk. Interaction with adjuvant and with other toxicants increases the actual risk. On the other hand, organochlorine pesticides and triazine herbicides require further investigation for a possible etiologic role in some hormone-dependent cancers. PMID- 15273661 TI - Integrin receptors in primary lung cancer. AB - In the present review recent data regarding the role of integrins--an important category of adhesion molecules mediating interactions among cells and components of the extracellular matrix--in lung cancer development is discussed. Investigations have shown that down-regulation of alpha3 integrin subunit may contribute to enhanced tumorigenicity of c-myc-overexpressing small cell lung carcinoma, while the loss of an integrin expression is correlated with recurrence in node-negative lung carcinoma. Increased expression of alpha1beta1 and alpha2beta1 integrins have been shown to be positively correlated with increased metastatic ability in squamous cell carcinoma. alpha3beta1 integrin is a most critical integrin for pulmonary development and epithelium integrity and its reduced expression in small cell lung cancer is probably related to the increased aggressiveness of this type. Pulmonary cancer cells generally express fewer integrin receptors than the normal epithelium. Additionally, since the ability of malignant cells to interact with extracellular matrix components is thought to be important, integrin dependent migration of lung cancer cells is a crucial process. PMID- 15273662 TI - Comparative study of human breast carcinoma MCF-7 cells differing in their resistance to doxorubicin: effect of ionizing radiation on apoptosis and TGF-beta production. AB - AIM: The aim of the study was to investigate the survival and growth of human breast carcinoma MCF-7 cells with different sensitivity to doxorubicin and production of transforming growth factor beta-(TGF-beta) in dependence on the dose- and duration of X-ray in order to check if the cross-resistance to doxorubicin and radiation effects exists. METHODS: Determination of cell number and valiability using trypan blue (0.1% (w/v)) exclusion method, Western blot analysis of p53 protein expression, biological testing of TGF-beta activity, lectinocytochemical analysis for apoptosis quantitative estimation in unirradiated and irradiated cells of both sublines of MCF-7 cells--sensitive (MCF 7(wt)) and resistant (MCF-7(DOX/R)) to doxorubicin. RESULTS: It was found that doxorubicin-resistant breast cancer cells were also more refractory to X radiation-dependent growth inhibition. There were revealed different effects of distinct doses of X-ray on p53 protein expression by cells of both sublines. The level of production of TGF-beta was compared in non-irradiated MCF-7 cells and in these cells exposed to X-radiation. It was shown that X-radiation increased TGF beta activity in the conditioned medium of the irradiated cells of both doxorubicin-sensitive and -resistant lines. CONCLUSIONS: The results of our study suggest that the biological effects of X-radiation on human breast cancer MCF-7 cells can be at least partly mediated by TGF-beta. Taking into account that TGF beta is a potent natural immunosupressor, one may consider that an increased activity of this cytokine can intensify negative effects of X-radiation. PMID- 15273663 TI - Influence of temperature on the efficiency of photodestruction of Ehrlich ascites carcinoma cells sensitized by hematoporphyrin derivative. AB - AIM: To elucidate the mechanism of the potentiating influence of heating associated with photoirradiation on the antitumor efficiency of photodynamic therapy (PDT) with hematoporphyrin derivative (HPD). METHODS: The study was carried out on Ehrlich ascites carcinoma (EAC) cells, which were loaded with HPD in a serum-free medium and then irradiated with red light (lambda max=630 nm) at various temperatures. Cytotoxicity was estimated by the trypan blue exclusion assay. RESULTS: Our data support the view that in PDT the hyperthermia (around 44 degrees C) produced by irradiation can enhance synergistically the HPD photoinduced tumor eradication; it was found that raising the irradiation temperature from 30 to 44 degrees C caused a substantial (approximately 1.5 fold) increase in the rate of HPD-photosensitized inactivation of EAC cells, while hyperthermia (44 degrees C) itself showed little toxic effects towards the cells. CONCLUSION: Studies indicated that the potentiating effect of heating on the antitumor efficiency of HPD-PDT could be largely explained by the stimulation of reactive oxygen species formation such as H2O2, superoxide and hydroxyl radicals. It was also found that photosensitization of EAC cells by HPD caused a strong fall in the activity of catalase (CAT) and glutathione (GSH) peroxidase, and that heating sensitized the H2O2-detoxifying enzymes to HPD-photoinduced inactivation. Under HPD-PDT, these events could result in loss of protection against accumulating H2O2; we revealed that cell-bound CAT and the GSH redox cycle play an important role in the protection of EAC cells against HPD-PDT. Moreover, our findings suggest that during PDT with HPD, an increase in the temperature of tumors could enhance the efficiency of this therapy via the stimulation of a chlorin-type photoproduct formation. PMID- 15273664 TI - Selectivity of effects of redox-active cobalt(III) complexes on tumor tissue. AB - AIM: To estimate the selectivity of action of cobalt complexes on tumor tissue. MATERIALS AND METHODS: Cobalt(III) complexes containing both the tetradentate Schiff-base ligand derived from acetylacetone and ethylenediamine, and compounds of the vitamin PP series or their synthetic analogs, viz. nicotinamide, isonicotinamide or nicotinic acid, as extra (axial) ligands, were tested in vivo on transplanted mice tumors, namely Lewis lung carcinoma (3LL), melanoma B16, and mammary adenocarcinoma Ca755. concentrations of malondialdehyde in tissue extracts were measured by standard biochemical methods. The rate of DNA unwinding was used to detect DNA damage in tumor cells. Level of tumor hypoxia as well as bioenergetic status were estimated using 31P NMR spectroscopy in perchloric acid extracts of tissue. RESULTS: A significant and selective increase of malondialdehyde in tumor tissue reflecting activation of lipid peroxidation was found after administration of the complexes. The bioenergetic status in tumor was also selectively affected by the complexes: minimization of signals of high energy phosphates was observed two hours after injection of the complexes. An increase of the number of DNA single-strand breaks was registered in tumor tissue, supporting the suggestion that the complexes may directly affect DNA. A correlation between the above tumor effects and the structure of axial ligands was demonstrated. CONCLUSION: Cobalt(III) complexes affect tumor tissue with a very high level of selectivity; in particular they activate lipid peroxidation, induce DNA single-strand breaks, suppress the bioenergetic status, and enhance hypoxia. It is supposed that the selective action of these complexes on tumor tissue is due to peculiarities of tumor microphysiology, in particular significant tumor hypoxia. PMID- 15273665 TI - Antisera to polycyclic aromatic hydrocarbons and their application on the detection of chemical carcinogens in blood sera of oncological patients. AB - AIM: To investigate the cross-reactivity of the hyperimmune antisera of animals (rabbits) and the sera of oncological patients to polycyclic aromatic hydrocarbons (PAHs) of a similar chemical structure. METHODS: Reactions of antibodies with haptens have been estimated by non-competitive and competitive ELISA with synthesized protein conjugates of PAHs as a coating antigene. RESULTS: All the model rabbit antisera have been stated to react with anthracene, chrysene, pyrene, benzo(a)pyrene and benz(a)anthracene irrespective of the hapten used to immunize animals. The cross-reactivity of serum antibodies to all five PAHs has been found in blood samples of oncological patients. CONCLUSION: It is recommended to use the conjugates of anthracene, chrysene and pyrene for detecting human antibodies to more dangerous environmental carcinogens- benzo(a)pyrene and benz(a)anthracene. PMID- 15273666 TI - Activity of irinotecan, cisplatin and dacarbazine (CPD) combination in previously treated patients with advanced colorectal carcinoma. AB - AIM: Irinotecan is an active drug after fluorouracil (FU) failure in patients with colorectal cancer (CRC). Also a modest activity of cisplatin and dacarbazine combination in FU resistant patients have been reported. We aimed to assess the efficacy of irinotecan, cisplatin and dacarbazine combination in previously treated patients with measurable advanced CRC. METHODS: Treatment schedule was irinotecan 150 mg/m2, iv, d1; cisplatin 20 mg/m2 and dacarbazine 200 mg/m2 iv, d1 d3; every 21 days. 48 patients with a median age of 51 were entered the study. RESULTS: Objective response rate was 33.3%. The overall disease stabilization rate was 75.6%. The median survival was 14 months, and the median progression free survival was 7 months. Main toxicities were grade 2-3 vomiting (39.2%) and grade 3-4 neutropenia (17.4%). CONCLUSION: CPD combination seems to be very active, with acceptable safety profile, in patients with advanced CRC resistant to FUFA. PMID- 15273667 TI - Sea-buckthorn juice protects mice against genotoxic action of cisplatin. AB - AIM: To study the influence of sea-buckthorn (SB) juice on the micronucleus (MN) frequency in bone marrow cells and sperm abnormality induced by cisplatin (CP, cis-dichlordiammineplatinum-II). MATERIALS AND METHODS: The experiments were performed with male Swiss albino mice. SB juice (0.3 ml) prepared ex tempore was given to mice by gavage during 5 or 10 days. 3 h after the last gavage, mice received CP at doses either 1.2 mg/kg or 2.4 mg/kg i.p. MN frequency was studied in bone marrow polychromatic erythrocytes 24 h after the injection of the drug. The abnormality of sperm heads was studied by microscopy. RESULTS: The SB juice decreased significantly the number of MN in bone marrow cells induced by CP at dose of 1.2 mg/kg by 36.5% and 47.9% (when it was given 5 and 10 days, consequently), and by 19.0% (p > 0.05) at dose of 2.4 mg/kg. The SB juice decreased significantly also the damaging effect of CP on sperm head at low dose and not significantly at the higher one. Antigenotoxic effect of juice was 45.0% and 16.6%, respectively. CONCLUSION: SB juice decreased significantly the genotoxic effect of CP at dose of 1.2 mg/kg on somatic (bone marrow) and germ (sperm) cells of mice. At higher dose of the drug the effects was not statistically significant. PMID- 15273668 TI - CD4+ T cells tumor specific response exists in L615 leukemia mice: adoptive transfer in combination with cyclophosphamide. AB - AIM: L615 leukemia cell line is a transplantable acute lymphocytic leukemia model with the CD4 positive phenotype. In this study, we explored whether tumor response specific T cells can be separated from the live leukemia mice or not. METHODS: The mutant HGPRT- L615 cell line was first established. The splenocytes from HGPRT- L615 leukemia mice were cultured and expanded in mixed tumor lymphocytes culture manner. The expanded T cells were sorted with FACScan. Then their killing capacity, IFN-gamma release as well as antitumor capacity in adoptive transfer experiments were analyzed. RESULTS: The expanded response T cells are mostly CD4 positive. The CD4 positive T cells showed high release of IFN-gamma upon stimulation but lacked significant cytotoxicity. In immunochemotherapy model, these CD4 positive T cells can cure most leukemia mice. CONCLUSIONS: We demonstrated the feasibility of separation of tumor response specific CD4+ T cells from CD4+ L615 leukemia mice. These CD4+ T cells can cure leukemia mice upon adoptive transfer in combination with cyclophosphamide pretreatment. PMID- 15273669 TI - Analysis of malignancy-associated DNA changes in interphase nuclei of buccal epithelium in persons with breast diseases. AB - AIM: To study from the point of view of statistical and geometrical theory of pattern recognition, the peculiarities of the distribution of optical density of DNA in the interphase nuclei of mammary buccal epithelium upon pathology. METHODS: Cytogenetic investigation of buccal smears and computer-based image analysis were used. RESULTS: It is shown that in malignant neoplasms of the mammary glands compared with its values in benign, the optical density of DNA in the nuclei of buccal epithelium increase in a range from 0.15 to 0.30 conventional units. The sensitivity of that criterium varied from 80.5% to 96.8%, and specificity was 92.3%. CONCLUSION: The method proposed may be recommended as an additional one for improvement of the diagnosis of mammary pathologies. PMID- 15273670 TI - Stage of differentiation, proliferative index of tumor cells and cytotoxic activity of peripheral blood lymphocytes in colorectal cancer patients. AB - AIM: To evaluate prognostic value of activity of cytotoxic lymphocytes of peripheral blood (CTL) of patients with colorectal cancer (CRC). METHODS: 2-year survival of 30 patients with colorectal cancer stage II (T2-3N0-1M0G1-3) was investigated in relation with clinical data, cytotoxic activity of peripheral blood lymphocytes (by MTT-test) and immunohistochemical peculiarities of tumor cells using anti-Ki-67-MABs. RESULTS: It was revealed that some factors had prognostic significance, namely regional lymph nodes involvement (r = -0.69), proliferative activity of tumor cells (Ki-67-positivity) (r = -0.67), restoration of CTL function after operation (r = 0.46), adjuvant chemotherapy (r = 0.41) and differentiation stage of tumor (r = 0.40). Negative correlation between CTL activity before operation and tumor proliferative activity also exists (r = 0.35). CONCLUSION: the data point to the involvement of natural killer cells in the control of tumor growth. PMID- 15273671 TI - Getting started in clinical research. PMID- 15273672 TI - Wristwatch-distal radial fracture as a marker for osteoporosis investigation: a controlled trial of patient education and a physician alerting system. AB - This article reports a controlled trial to investigate the effectiveness of patient education and a physician alerting system in altering secondary osteoporosis prevention after a low-trauma (fragility) wrist fracture and to record the current rate of osteoporosis investigation following such fractures. Fifty-one women and men aged 50 years or older with a low-trauma wrist fracture were identified (41 women and 10 men; mean age [95% CI], 71.51 [67.31-74.81]). The intervention group received a four-part intervention aimed at both the patient and the family physician in addition to the usual care for the fracture. The control group only received usual care for the fracture. Data were collected for both groups at six weeks and six months. Results indicate that 92% of the intervention subjects were investigated for osteoporosis, compared with the usual care group, in which only 23% were investigated. Early osteoporosis intervention has the potential to limit disease impact, and hand therapists can play a key role in early identification of osteoporosis. PMID- 15273673 TI - Tendinopathies in the upper extremity: a paradigm shift. AB - Epicondylitis and de Quervain's tenosynovitis are two common diagnoses seen by hand therapists in clinical practice. Traditionally, these conditions have been viewed as being due to an inflammatory process and treated as such. New research shows that tendons exhibit areas of degeneration and a distinct lack of inflammatory cells. Tendinosis is degeneration of the collagen tissue due to aging, microtrauma, or vascular compromise. This article reviews key literature related to tendinopathies in the lower extremity and comments on the dearth of similar articles for the elbow and forearm. Hand therapists are encouraged to embrace this new terminology and to engage in research in this difficult area to improve treatment regimens and outcome measures. PMID- 15273674 TI - A comparison of high-profile and low-profile dynamic mobilization splint designs. AB - Despite claims that the high-profile dynamic mobilization splint design requires less frequent adjustments than the low-profile design, the authors are not aware of biomechanical evidence supporting such claims. The purpose of this study was to reexamine this claim and quantitatively analyze each design as well as the differences between designs with respect to the actual deviation from a 90 degrees angle of applied force for 60 degrees, 30 degrees, 20 degrees, and 10 degrees gains in proximal interphalangeal joint (PIP) extension. Additionally, for 10 degrees, 20 degrees, and 30 degrees gains in PIP extension, the authors determined the corrective and shear forces as a function of the deviation from a 90 degrees angle of applied force for each design, as well as the difference between the designs. Results show that in all instances examined, the actual difference between the designs is quite small. Implications of such findings are discussed along with newly identified relationships of potential utility to the hand therapist. PMID- 15273675 TI - Provocative test for carpal tunnel syndrome. AB - A new clinical provocative test for diagnosing carpal tunnel syndrome (CTS) is described. The examiner uses his or her fingers to simultaneously exert dorsal pressure on the first metacarpal and pisotriquetral complex and volar pressure on the lunate. The test result is considered positive if symptoms of CTS are reproduced. Thirty-four patients (41 hands) with carpal CTS and 21 control subjects (41 hands) were evaluated. The sensitivity of the suggested provocative test was 98%, the specificity was 98%, the positive predictive value was 98%, and the negative predictive value was 98%. These data support the use of the provocative test for CTS. PMID- 15273676 TI - Does training improve writer's cramp? An evaluation of a behavioral treatment approach using kinematic analysis. AB - Patients with writer's cramp (WC) show uncontrollable muscle co-contractions of agonists and antagonists and unusual postures of the upper limb during writing; their handwriting is inefficient and exhausting. Currently the treatment of choice is to inject botulinum toxin in selective hand muscles. However, this treatment has two drawbacks: it is short-lasting and may be associated with adverse side effects. An alternative behavioral treatment, namely, the handwriting training developed by Mai and coworkers, was carried out and evaluated in 50 patients with WC. A digitizing tablet was used to record the handwriting movements before and after training, and then again after a follow-up period. The results indicate the efficacy of the handwriting training. Speed and smoothness of the handwriting increased during the training with effects observed after an extended follow-up period. Handwriting training leads to significant improvements, but does not bring handwriting performance to normal levels. This training should be considered as an alternative or supplement to the traditional treatment with botulinum toxin or other programs based on neuroplasticity. PMID- 15273677 TI - Individual finger strength: are the ulnar digits "powerful"? AB - This study determined the test-retest reliability of a grip device that measures the contribution of individual fingers to grip strength and described the pattern of contribution in subjects without hand pathology. Subjects repeated a set of three maximal grip efforts on two occasions separated by two to seven days. Intraclass correlation reliability coefficients were high (>0.75) for eight out of ten strength measures. The percentage contributions of the index, middle, ring, and small fingers to grip were approximately 25%, 35%, 25%, and 14%, respectively. Grip and finger strengths were highly correlated. Anthropometric measures of body size or finger length were moderately correlated with strength measures. These data suggest that there is a predictable pattern by which individual fingers contribute to overall grip strength, which is partially related to body size. The ulnar side of the hand contributes to the smaller proportion of overall grip (approximately 60% radial, 40% ulnar). The clinical utility of finger strength measures should be explored. PMID- 15273678 TI - Effect of pressure garment work gloves on hand function in patients with hand burns: a pilot study. AB - This study aims to compare the effects of pressure garment work gloves (PGWGs) and standard-pressure garment gloves (SPGGs) on functional hand use in individuals with hand burns. A quasi-experimental, repeated-measure design was used. Each glove was worn for one week prior to testing. Grip strength, hand function, functional sensation, and functional task scores were assessed. Participants were asked to report which glove they preferred and why. Two individuals with three burned hands underwent this study. Grip and pinch strengths and functional sensation were decreased overall with the PGWG. However, better performance was noted with functional tasks involving gross and static fine-motor movements, and simple, dynamic fine-motor movements. These findings were found to be significant using a Cochran's test for related observations. Moreover, there was unanimous participant preference for the PGWG. In conclusion, use of PGWGs facilitated select functional performance, warranting further investigation. PMID- 15273679 TI - An ulnar boost splint for midcarpal instability. PMID- 15273680 TI - Modification to a pediatric thumb splint. PMID- 15273685 TI - Caenorhabditis elegans ABL-1 antagonizes p53-mediated germline apoptosis after ionizing irradiation. AB - c-Abl, a conserved nonreceptor tyrosine kinase, integrates genotoxic stress responses, acting as a transducer of both pro- and antiapoptotic effector pathways. Nuclear c-Abl seems to interact with the p53 homolog p73 to elicit apoptosis. Although several observations suggest that cytoplasmic localization of c-Abl is required for antiapoptotic function, the signals that mediate its antiapoptotic effect are largely unknown. Here we show that worms carrying an abl 1 deletion allele, abl-1(ok171), are specifically hypersensitive to radiation induced apoptosis in the Caenorhabditis elegans germ line. Our findings delineate an apoptotic pathway antagonized by ABL-1, which requires sequentially the cell cycle checkpoint genes clk-2, hus-1 and mrt-2; the C. elegans p53 homolog, cep-1; and the genes encoding the components of the conserved apoptotic machinery, ced 3, ced-9 and egl-1. ABL-1 does not antagonize germline apoptosis induced by the DNA-alkylating agent ethylnitrosourea. Furthermore, worms treated with the c-Abl inhibitor STI-571 (Gleevec; used in human cancer therapy), two newly synthesized STI-571 variants or PD166326 had a phenotype similar to that generated by abl 1(ok171). These studies indicate that ABL-1 distinguishes proapoptotic signals triggered by two different DNA-damaging agents and suggest that C. elegans might provide tissue models for development of anticancer drugs. PMID- 15273686 TI - The imprinted signaling protein XL alpha s is required for postnatal adaptation to feeding. AB - Genomic imprinting, by which maternal and paternal alleles of some genes have different levels of activity, has profound effects on growth and development of the mammalian fetus. The action of imprinted genes after birth, in particular while the infant is dependent on maternal provision of nutrients, is far less well understood. We disrupted a paternally expressed transcript at the Gnas locus, Gnasxl, which encodes the unusual Gs alpha isoform XL alpha s. Mice with mutations in Gnasxl have poor postnatal growth and survival and a spectrum of phenotypic effects that indicate that XL alpha s controls a number of key postnatal physiological adaptations, including suckling, blood glucose and energy homeostasis. Increased cAMP levels in brown adipose tissue of Gnasxl mutants and phenotypic comparison with Gnas mutants suggest that XL alpha s can antagonize Gs alpha-dependent signaling pathways. The opposing effects of maternally and paternally expressed products of the Gnas locus provide tangible molecular support for the parental-conflict hypothesis of imprinting. PMID- 15273687 TI - A cis-acting control region is required exclusively for the tissue-specific imprinting of Gnas. AB - Genomic imprinting brings about allele-specific silencing according to parental origin. Silencing is controlled by cis-acting regulatory regions that are differentially marked during gametogenesis and can act over hundreds of kilobases to silence many genes. Two candidate imprinting control regions (ICRs) have been identified at the compact imprinted Gnas cluster on distal mouse chromosome 2, one at exon 1A upstream of Gnas itself and one covering the promoters for Gnasxl and the antisense Nespas (ref. 8). This imprinted cluster is complex, containing biallelic, maternally and paternally expressed transcripts that share exons. Gnas itself is mainly biallelically expressed but is weakly paternally repressed in specific tissues. Here we show that a paternally derived targeted deletion of the germline differentially methylated region at exon 1A abolishes tissue-specific imprinting of Gnas. This rescues the abnormal phenotype of mice with a maternally derived Gnas mutation. Imprinting of alternative transcripts, Nesp, Gnasxl and Nespas (ref. 13), in the cluster is unaffected. The results establish that the differentially methylated region at exon 1A contains an imprinting control element that specifically regulates Gnas and comprises a characterized ICR for a gene that is only weakly imprinted in a minority of tissues. There must be a second ICR regulating the alternative transcripts. PMID- 15273688 TI - Antagonism between DNA hypermethylation and enhancer-blocking activity at the H19 DMD is uncovered by CpG mutations. AB - Imprinted expression at the H19-Igf2 locus depends on a differentially methylated domain (DMD) that acts both as a maternal-specific, methylation-sensitive insulator and as a paternal-specific site of hypermethylation. Four repeats in the DMD bind CCCTC-binding factor (CTCF) on the maternal allele and have been proposed to attract methylation on the paternal allele. We introduced point mutations into the DMD to deplete the repeats of CpGs while retaining CTCF binding and enhancer-blocking activity. Maternal inheritance of the mutations left H19 expression and Igf2 imprinting intact, consistent with the idea that the DMD acts as an insulator. Conversely, paternal inheritance of these mutations disrupted maintenance of DMD methylation, resulting in biallelic H19 expression. Furthermore, an insulator was established on the paternally inherited mutated allele in vivo, reducing Igf2 expression and resulting in a 40% reduction in size of newborn offspring. Thus, the nine CpG mutations in the DMD showed that the two parental-specific roles of the H19 DMD, methylation maintenance and insulator assembly, are antagonistic. PMID- 15273689 TI - Interaction between differentially methylated regions partitions the imprinted genes Igf2 and H19 into parent-specific chromatin loops. AB - Imprinted genes are expressed from only one of the parental alleles and are marked epigenetically by DNA methylation and histone modifications. The paternally expressed gene insulin-like growth-factor 2 (Igf2) is separated by approximately 100 kb from the maternally expressed noncoding gene H19 on mouse distal chromosome 7. Differentially methylated regions in Igf2 and H19 contain chromatin boundaries, silencers and activators and regulate the reciprocal expression of the two genes in a methylation-sensitive manner by allowing them exclusive access to a shared set of enhancers. Various chromatin models have been proposed that separate Igf2 and H19 into active and silent domains. Here we used a GAL4 knock-in approach as well as the chromosome conformation capture technique to show that the differentially methylated regions in the imprinted genes Igf2 and H19 interact in mice. These interactions are epigenetically regulated and partition maternal and paternal chromatin into distinct loops. This generates a simple epigenetic switch for Igf2 through which it moves between an active and a silent chromatin domain. PMID- 15273690 TI - Continuing role for mouse Numb and Numbl in maintaining progenitor cells during cortical neurogenesis. AB - Neural progenitor cells in the developing neocortex change over time to produce different neurons, a phenomenon that is also observed in other regions of the nervous system. Mouse Numb (also known as m-numb) and Numbl (also known as numblike or Nbl) are redundant but essential in maintaining virtually all progenitor cells during early neurogenesis. They do this by allowing cells to choose progenitor over neuronal fates. To determine whether their roles change as neurogenesis progresses, we conditionally ablated both genes in the embryonic dorsal forebrain after initial waves of neurogenesis. Here we report that these proteins continue to be required for progenitor-cell maintenance, contrary to recently reported findings. As occurs during early neurogenesis, the loss of Numb and Numbl causes premature progenitor-cell depletion and, consequently, a highly specific malformation of the neocortex and hippocampus. Because progenitor cells can proliferate without Numb and Numbl before neurogenesis, we propose that Numb mediated asymmetric cell divisions, which diversify many cell fates in Drosophila melanogaster, represent a general mechanism in mammals for stem cells to balance self-renewal and differentiation. PMID- 15273691 TI - Early life experience alters response of adult neurogenesis to stress. AB - Maternal deprivation produces persistent abnormalities in behavioral and neuroendocrine functions associated with the hippocampus, a brain region that shows considerable structural change in response to experience throughout life. Here we show that adverse experience early in life affects the regulation of adult neurogenesis in the hippocampus. More specifically, a decrease in cell proliferation and immature neuron production are observed in the dentate gyrus of adult rats that are maternally separated as pups. Although maternally separated rats show normal basal levels of corticosterone, the suppression of cell proliferation in these rats can be reversed by lowering corticosterone below the control value. In addition, normal stress-induced suppression of cell proliferation and neurogenesis, despite normal activation of the hypothalamic pituitary adrenal (HPA) axis, is not observed in maternally separated rats. Our results suggest that early adverse experience inhibits structural plasticity via hypersensitivity to glucocorticoids and diminishes the ability of the hippocampus to respond to stress in adulthood. PMID- 15273692 TI - Multiplexing using synchrony in the zebrafish olfactory bulb. AB - In the olfactory bulb (OB) of zebrafish and other species, odors evoke fast oscillatory population activity and specific firing rate patterns across mitral cells (MCs). This activity evolves over a few hundred milliseconds from the onset of the odor stimulus. Action potentials of odor-specific MC subsets phase-lock to the oscillation, defining small and distributed ensembles within the MC population output. We found that oscillatory field potentials in the zebrafish OB propagate across the OB in waves. Phase-locked MC action potentials, however, were synchronized without a time lag. Firing rate patterns across MCs analyzed with low temporal resolution were informative about odor identity. When the sensitivity for phase-locked spiking was increased, activity patterns became progressively more informative about odor category. Hence, information about complementary stimulus features is conveyed simultaneously by the same population of neurons and can be retrieved selectively by biologically plausible mechanisms, indicating that seemingly alternative coding strategies operating on different time scales may coexist. PMID- 15273693 TI - Caged phosphopeptides reveal a temporal role for 14-3-3 in G1 arrest and S-phase checkpoint function. AB - Using classical genetics to study modular phosphopeptide-binding domains within a family of proteins that are functionally redundant is difficult when other members of the domain family compensate for the product of the knocked-out gene. Here we describe a chemical genetics approach that overcomes this limitation by using UV-activatable caged phosphopeptides. By incorporating a caged phosphoserine residue within a consensus motif, these reagents simultaneously and synchronously inactivate all phosphoserine/phosphothreonine-binding domain family members in a rapid and temporally regulated manner. We applied this approach to study the global function of 14-3-3 proteins in cell cycle control. Activation of the caged phosphopeptides by UV irradiation displaced endogenous proteins from 14 3-3-binding, causing premature cell cycle entry, release of G1 cells from interphase arrest and loss of the S-phase checkpoint after DNA damage, accompanied by high levels of cell death. This class of reagents will greatly facilitate molecular dissection of kinase-dependent signaling pathways when applied to other phosphopeptide-binding domains including SH2, Polo-box and tandem BRCT domains. PMID- 15273694 TI - Replication protein A interacts with AID to promote deamination of somatic hypermutation targets. AB - Activation-induced cytidine deaminase (AID) is a single-stranded (ss) DNA deaminase required for somatic hypermutation (SHM) and class switch recombination of immunoglobulin genes. Class switch recombination involves transcription through switch regions, which generates ssDNA within R loops. However, although transcription through immunoglobulin variable region exons is required for SHM, it does not generate stable ssDNA, which leaves the mechanism of AID targeting unresolved. Here we characterize the mechanism of AID targeting to in-vitro transcribed substrates harbouring SHM motifs. We show that the targeting activity of AID is due to replication protein A (RPA), a ssDNA-binding protein involved in replication, recombination and repair. The 32-kDa subunit of RPA interacts specifically with AID from activated B cells in a manner that seems to be dependent on post-translational AID modification. Thus, our study implicates RPA as a novel factor involved in immunoglobulin diversification. We propose that B cell-specific AID-RPA complexes preferentially bind to ssDNA of small transcription bubbles at SHM 'hotspots', leading to AID-mediated deamination and RPA-mediated recruitment of DNA repair proteins. PMID- 15273699 TI - Smad3 is required for dedifferentiation of retinal pigment epithelium following retinal detachment in mice. AB - Retinal pigment epithelial (RPE) cells dedifferentiate and undergo epithelial mesenchymal transition (EMT) following retinal detachment, playing a central role in formation of fibrous tissue on the detached retina and vitreous retraction (proliferative vitreoretinopathy (PVR)). We have developed a mouse model of subretinal fibrosis with implications for PVR in which retinal detachment is induced without direct damage to the RPE cells. Transforming growth factor-beta (TGF-beta) has long been implicated both in EMT of RPEs and the development of PVR. Using mice null for Smad3, a key signaling intermediate downstream of TGF beta and activin receptors, we show that Smad3 is essential for EMT of RPE cells induced by retinal detachment. De novo accumulation of fibrous tissue derived from multilayered RPE cells was seen following experimental retinal detachment in eyes of wild type, but not Smad3-null mice. Expression of alpha-smooth muscle actin, a hallmark of EMT in this cell type, and extracellular matrix components, lumican and collagen VI, were also not observed in eyes of Smad3-null mice. Our data show that induction of PDGF-BB by Smad3-dependent TGF-beta signaling is likely an important secondary proliferative component of the disease process. The results suggest that blocking the Smad3 pathway might be beneficial in prevention/treatment of PVR. PMID- 15273700 TI - Identification of differentially expressed genes in a renal cell carcinoma tumor model after endostatin-treatment. AB - Endostatin is a cleavage product of collagen XVIII that has shown to inhibit tumor-angiogenesis in experimental tumor models. At present, the exact molecular mechanism of action of endostatin is not completely elucidated. In this study, we wanted to identify specific target genes of endostatin. For this purpose, the human renal cell carcinoma RC-9 was subcutaneously implanted in nude mice and treated with endostatin. Tumor growth was inhibited by endostatin after 4 days of treatment. Using immunohistochemistry and the hypoxia marker pimonidazole, we demonstrate disintegration of blood vessels and hypoxia and anoxia as a result of the treatment. Hereafter, we applied the polymerase chain reaction (PCR)-based subtractive suppression hybridization (SSH) method, together with the mirror orientation selection (MOS) technique to identify specifically induced and suppressed genes after endostatin-treatment. We found eight genes to be specifically induced and 11 to be suppressed by the endostatin-treatment. Among other genes, core binding factor a-1/osteoblast-specific factor-2 (cbfa1/osf2) was found to be specifically suppressed by endostatin. Unexpectedly, cbfa1/osf2 was found to be specifically expressed in granulocytes in the tumor, not only in the experimental RC-9 tumor model, but in sections of human breast cancer as well. Since an effect of antiangiogenic therapy on granulocytes has been reported before, this might lead to new insights in the role of granulocytes in antiangiogenic therapy in general. In conclusion, the SSH-PCR implemented with the MOS-technique is a powerful tool to identify differentially expressed genes. Using these techniques, we have identified several target genes of endostatin, of which cbfa1/osf2 was found to be specifically expressed in granulocytes in the tumor. PMID- 15273701 TI - Autosomal differences between males and females in hybrid zones: a first report from Barbus barbus and Barbus meridionalis (Cyprinidae). AB - Narrow hybrid zones are generally subjected to the action of two forces: dispersal, which tends to homogenise the hybridising taxa, and selection against hybrids, which, in contrast, produces steep clines of introgression for diagnostic markers. Although differences between sexes in dispersal abilities or in susceptibility to hybrid counterselection are common in hybrid zones, autosomal genetic differences between males and females have never been reported to our knowledge. Barbus barbus and Barbus meridionalis (Cyprinidae) form a hybrid zone along the Lergue river. By carrying out a genetic analysis of males and females in six samples from two central stations of the hybrid zone using codominant markers (six allozymes and four microsatellite loci), we revealed significant multilocus and monolocus differences between the sexes. This could reflect a genetic difference among sexes within a same cohort, caused either by a survival (or fertility) differential among sexes or by a sex-specific pattern of dispersal. Alternatively, this may be due to genetic differentiation between cohorts, since male and female barbel exhibit different maturation, growth and survival patterns leading to different age distributions among sexes, and particularly among reproducers. PMID- 15273704 TI - Risk assessment and outcome of chronic graft-versus-host disease after allogeneic peripheral blood progenitor cell transplantation in pediatric patients. AB - We retrospectively evaluated the incidence, risk factors for chronic graft-versus host disease (cGvHD) and outcome in 80 pediatric patients (36 male) (median age 13 years) who underwent allogeneic peripheral blood progenitor cell transplantation. Patients were grafted from an HLA-identical sibling after myeloablative conditioning (total body irradiation (TBI) based 52; non-TBI 28). GvHD prophylaxis used were: cyclosporin A (CsA)+ short methotrexate (MTX) in 52 and CsA+/-prednisone in 28. The median number of CD34+ cells infused were 5.8 x 10(6)/kg (range: 1.4-32.8). The median follow-up was 24 months (range: 3-94). In all, 28 patients had cGvHD (confidence interval (CI): 54.2+/-10%). Factors that were significant on univariate analysis were diagnosis (P=0.03) and GvHD prophylaxis administered (P=0.04). On multivariate analysis, only GvHD prophylaxis used was associated with a significant risk of cGvHD (hazard ratio (HR): 3.94; 95% CI: 1.41-10.91, P=0.009). The CI of cGvHD for patients receiving CsA+MTX was 40.9+/-12 vs 76.5+/-18% for patients who did not (P=0.03). The probability of relapse was 36+/-6% for all patients (12.5+/-8% for patients with cGvHD vs 47.9+/-8% without cGvHD). The probability of disease-free survival was better for patients with cGvHD (69.9+/-10 vs 37.9+/-7%; HR: 3.59, 95% CI: 1.47 5.56; P=0.001). Our data suggest that the GvHD prophylaxis used is the most relevant predictor of cGvHD. Patients with cGvHD had a lower risk of relapse and a better survival. PMID- 15273705 TI - Tacrolimus as monotherapy or combined with minidose methotrexate for graft-versus host disease prophylaxis after allogeneic peripheral blood stem cell transplantation: long-term outcomes. AB - We report long-term outcomes for allogeneic peripheral blood stem cell (PBSC) recipients, evaluating cumulative incidence (CI) of acute and chronic graft versus-host disease (GVHD), after use of tacrolimus with or without minidose methotrexate for GVHD prophylaxis. From April 1996 to April 1998, 97 adults underwent allogeneic PBSC transplantation. The first 49 received tacrolimus monotherapy as GVHD prophylaxis and the subsequent 48 received combination therapy. The median follow-up for survivors was 67 months. Compared to combination therapy, tacrolimus monotherapy resulted in enhanced neutrophil engraftment, shortened hospitalization, comparable rates of GVHD, and equivalent 100-day survival. The CI of grades II-IV acute GVHD was 35% with tacrolimus and 23% with the combination (P=0.19). The CI of III-IV GVHD was 22% for tacrolimus and 19% for the combination. CI of chronic GVHD was similar between the two groups (P=0.5). Patients with good-risk disease had superior overall survival (OS) when compared to those with poor-risk features (P<0.001). Within the good risk disease category, patients receiving methotrexate had a trend towards improved OS (P=0.07). Multivariate analysis indicated good-risk disease status and methotrexate were independently associated with improved OS, progression-free survival (PFS), and relapse. In addition, patients developing chronic GVHD had a significantly reduced risk of relapse and improved PFS. PMID- 15273706 TI - A randomised study comparing peripheral blood progenitor mobilisation using intermediate-dose cyclophosphamide plus lenograstim with lenograstim alone. AB - We conducted a prospective randomised study to compare the efficiency of out patient progenitor cell mobilisation using either intermediate-dose cyclophosphamide (2 g/m(2)) and lenograstim at 5 micrograms/kg (Cyclo-G-CSF group, n=39) or lenograstim alone at 10 micrograms/kg (G-CSF group, n=40). The end points were to compare the impact of the two regimens on mobilisation efficiency, morbidity, time spent in hospital, the number of apheresis procedures required and engraftment kinetics. Successful mobilisation was achieved in 28/40 (70%) in the G-CSF group vs 22/39 (56.4%) for Cyclo-G-CSF (P=0.21). The median number of CD34+ cells mobilised was 2.3 x 10(6)/kg and 2.2 x 10(6)/kg for G-CSF and cyclo-G-CSF arms following a median of two apheresis procedures. Nausea and vomiting and total time spent in the hospital during mobilisation were significantly greater after Cyclo-G-CSF (P<0.05). Rapid neutrophil and platelet engraftment was achieved in all transplanted patients in both groups. In conclusion, G-CSF at 10 micrograms/kg was as efficient at mobilising progenitor cells as a combination of cyclophosphamide and G-CSF with reduced hospitalisation and side effects and prompt engraftment. When aggressive in-patient cytoreductive regimens are not required to both control disease and generate progenitor cells, the use of G-CSF alone appears preferable to combination with intermediate-dose cyclophosphamide. PMID- 15273707 TI - Reduced-intensity conditioning prior to allogeneic transplantation of hematopoietic stem cells: the need for T cells early after transplantation to induce a graft-versus-lymphoma effect. AB - In patients with poor-risk relapse of aggressive lymphoma, reduced-intensity conditioning followed by allogeneic PBSCT may have its limitations because of rapid regrowth of the tumor. We tried to address this problem by intermediate intensity conditioning followed by allogeneic SCT. A total of 21 patients received fludarabine, busulfan and cyclophosphamide prior to allogeneic SCT. In the first 10 patients, GVHD prophylaxis by CD34+ selection of the grafts was employed (group I). The next 11 patients received nonmanipulated grafts and mycophenolat mofetil plus cyclosporinA (group II). In group I, no GVHD was observed. In contrast, patients in group II had a significant risk of acute GVHD (aGVHD) (six patients with grade II-IV acute GVHD). However, in group I, all surviving patients progressed within 9 months. In contrast, eight of nine surviving patients of group II remain in remission after a median observation time of 10.5 months (range 4-22 months). Survival differed significantly between the groups (P=0.004). Multivariate analysis identified intensive GVHD prophylaxis as important risk factor for survival. These results support the existence of a clinically relevant GVL effect in aggressive lymphoma. T-cell depletion (or CD34 selection) of grafts is not recommended in patients with poor-risk aggressive NHL. PMID- 15273708 TI - Sirolimus for GVHD prophylaxis in allogeneic stem cell transplantation. AB - Sirolimus is a novel macrolide immunosuppressant widely used in solid organ transplantation. We have conducted three clinical trials using this compound as prophylaxis against GVHD after allogeneic stem cell transplantation. Our studies have demonstrated excellent GVHD control even when mismatched and unrelated donors were used. The morbidity and mortality associated with transplantation were reduced due to the omission or reduction in methotrexate dose. Furthermore, CMV reactivation and fungal infection rates were low. However, we have noted that sirolimus may be associated with increased rates of thrombotic microangiopathy after transplantation. Sirolimus has other uses, such as the treatment of established acute and chronic GVHD, and may be useful for treatment of post transplant lymphoproliferative disorder and perhaps as an antineoplastic agent against a wide variety of hematologic and solid neoplasms. PMID- 15273709 TI - HER2 overexpression as a prognostic factor in metastatic breast cancer patients treated with high-dose chemotherapy and autologous stem cell support. AB - We retrospectively evaluated the predictive and prognostic role of HER2 expression in 44 metastatic breast cancer (MBC) patients treated with high-dose consolidation chemotherapy (HDCT) and autologous stem cell support after induction chemotherapy (IC) with six courses of epirubicin+paclitaxel (22 patients) or gemcitabine+epirubicin+paclitaxel (22 patients). HER2 expression was evaluated by an immunohistochemical method (Herceptest, Dako). A total of 13 patients (29.5%) showed a HER2 overexpression (score 3+). After IC, nine patients were in complete response (CR), 30 in partial response (PR), and five in stable disease (SD); after HDCT, 20 (45.5%) obtained a CR, and 23 were in PR, for a conversion rate of 48.5%. Conversion rate for HER2-positive patients was 87.5 vs 37% for HER2-negative patients (P=0.018). The median progression-free (PFS) and overall survivals (OS) were 17.6 (95% CI 13.2-22.0) and 44 (95% CI 25.9-62.3) months, respectively. Patients with HER2 overexpression experienced a significantly (P=0.0042) shorter median PFS (15.3 months, 95% CI 11.1-19.5) compared to HER2-negative patients (21.3 months, 95% CI 14.3-28.4). The median OS was 27.6 months (95% CI 4.5-50.7) in HER2-positive patients and 50.3 months (95% CI 38.7-62.0) in HER2-negative patients (P=0.345). These results indicate that HER2 overexpression predicts a worse outcome for patients with MBC treated with HDCT, despite the high CR rate obtained in this subset of patients. PMID- 15273710 TI - Donor leukocyte infusions in relapsed Hodgkin's lymphoma following allogeneic stem cell transplantation: CD3+ cell dose, GVHD and disease response. AB - Nine patients with advanced Hodgkin's lymphoma (HL) who had undergone allogeneic stem cell transplantation (allo-SCT) received donor leukocyte infusions (DLIs) for treatment of persistent or progressive disease (PD). A total of 15 DLIs were performed, with four patients receiving more than one DLI. In four patients, prior salvage chemotherapy was administered. The median CD3+ cell dose administered was 77.5 x 10(6)/kg (range 5-285). GVHD developed in all but one patient. The response rate was 4/9 (44%). Three of these four responders developed GVHD and 3/4 had received chemotherapy. No correlation was observed between CD3+ cell dose infused and disease response. At the latest follow-up, three patients are alive and six have expired (PD n=3, nonrelapse mortality n=3). The median response duration was 7 months (range 4-9), with one response currently ongoing. These data suggest that DLIs for immunotherapy of recurrent HL have significant activity, although they frequently leads to GVHD. The small sample size does not allow any conclusion as to whether chemotherapy administration increases the chance of response. The CD3 cell dose infused does not seem to correlate with disease response. PMID- 15273711 TI - Evidence for a graft-versus-mast-cell effect after allogeneic bone marrow transplantation. AB - Mast cell leukemia (MCL) is a rare form of aggressive mastocytosis with a reported median survival below 6 months. Casuistic reports suggest the effectiveness of allogeneic bone marrow transplantation (BMT) for MCL. However, these reports lack clear evidence for a graft-versus-mast-cell (GvMC) effect. We prospectively investigated the GvMC at different time points after allogeneic BMT and donor-lymphocyte infusions (DLI). Samples were gathered from a patient with MCL treated with allogeneic BMT from an unrelated HLA identical donor. Parameters for detection of a GvMC effect included flow cytometrical analysis of mast cell (MC) populations in peripheral blood and BM, BM smear and histology, chimerism analysis of flow cytometrically sorted BM CD117+/CD34- MC and testing for anti mast cell reactivity of donor lymphocytes by interferon (IFN)-gamma ELISPOT. DLIs reduced MC from 5 to 0.5%. MC chimerism analysis demonstrated a complete recipient genotype after BMT, suggesting that the persistent mastocytosis was part of residual neoplastic disease. At 3.7 years after BMT, there is some evidence for relapse. In summary, BMT and DLIs attenuated the mastocytosis from an aggressive to an indolent form and may have improved the patients' prognosis. The in vitro data of our study indicate for the first time the existence of a GvMC effect. PMID- 15273712 TI - Granulocyte transfusions in the G-CSF era. Where do we stand? AB - The efficacy of granulocytes transfusions (GTX) in either the prevention or treatment of neutropenic sepsis has been a controversial issue. Early studies employing steroid mobilised GTX showed variable, dose-dependent results and significant pulmonary toxicity was reported. With the introduction of the recombinant myeloid growth factor, granulocyte-colony stimulating factor (G-CSF), the quantity of granulocytes that could be harvested was substantially increased leading to renewed interest in the clinical application of GTX. The administration of G-CSF to normal donors leads to significantly higher pre harvest neutrophil counts and consequently larger granulocyte harvests. Infusion of G-CSF stimulated GTX results in measurable increases in the recipients' neutrophil count and may reduce the duration and severity of neutropenia. However, the efficacy of these GTX in treating or preventing established neutropenic sepsis remains to be established in prospective controlled clinical trials. PMID- 15273713 TI - High sensitivity of megakaryocytic progenitor cells contained in placental/umbilical cord blood to the stresses during cryopreservation. AB - In placental/umbilical cord blood (PCB) banking and PCB transplantation (PCBT), long-term cryopreservation of hematopoietic stem and progenitor cells is a unique requirement as compared to that for bone marrow transplantation and cytokine mobilized peripheral blood transplantation. A long period of severe thrombocytopenia is a problem in many patients after PCBT. The object of this study was to define whether megakaryocytic progenitor cells (CFU-Meg), which produce platelets, are more sensitive to cryopreservation than the other myeloid progenitor cells in PCB. The leukocyte concentrates (LCs) obtained from clinical PCB banks were cryopreserved, and progenitor cell recoveries were determined by differential count of colony-forming cells (CFCs). The LCs were exposed to stresses which cells face during freezing, thawing, and washing out cryoprotectants. Most of the myeloid progenitor cells contained in the LCs showed good survival when cryopreserved at slow cooling rates, although cellular injury was observed at higher cooling rates and higher osmolalities. In contrast, the recovery rate of CFU-Meg was significantly lower than other progenitor cells, indicating a higher sensitivity to the various stresses they were exposed to during cryopreservation. Thrombocytopenia observed in patients receiving PCBT may be explained, at least in part, by the disappearance of CFU-Meg during cryopreservation. PMID- 15273714 TI - High-dose CEB vs BEAM with autologous stem cell transplant in lymphoma. AB - Between January 1996 and July 2002, 72 patients with non-Hodgkin's lymphoma or Hodgkin's disease underwent high-dose chemotherapy with autologous stem cell transplant conditioned with either cyclophosphamide, etoposide, carmustine (CEB) or carmustine, etoposide, cytarabine, melphalan (BEAM) at a single institution. In all, 52 patients received CEB and 20 patients received the BEAM regimen. Patient characteristics that were significantly different between the two groups are tumor grade and extranodal involvement (P = 0.0196, 0.0341, respectively). Regimen-related toxicities examined yielded only diarrhea occurring at a higher rate in the BEAM group (81 vs 51%, P = 0.0026), although cases were milder (92 vs 57%). Patients treated with CEB developed mucositis at a slightly higher rate (79%) than patients treated with BEAM (75%), but this difference did not reach statistical significance. However, the mucositis that occurred within the BEAM group was predominately mild (67%) in contrast to the predominance of moderate to severe cases in the CEB group (74%). In addition, patients treated with CEB required growth factor support for a longer time than patients treated with BEAM (P = 0.0399). Response rates were high in both groups, with trends favoring the BEAM group. Overall survival was higher after treatment with BEAM than with CEB (84 vs 60%). PMID- 15273715 TI - Alternative mutations of BRAF, RET and NTRK1 are associated with similar but distinct gene expression patterns in papillary thyroid cancer. AB - Papillary thyroid carcinoma (PTC) is associated with RET and NTRK1 rearrangements and BRAF mutations. A series of 60 PTCs collected in a single center from Italian patients were histologically re-examined and subclassified as well differentiated or tall cell variant. The sample collection was analysed for the presence of all the reported PTC-associated genetic alterations through DNA or cDNA amplification, followed by automated sequencing. The analysis of exons 11 and 15 of BRAF gene revealed the T1796A (V599E) mutation in 32% of cases, and this alteration is significantly associated with PTC tall cell variant. Oncogenic rearrangements of RET and NTRK1 receptors were found in 33 and 5% of cases, respectively. No Ras mutations were detected. Overall, genetic alterations were detected in two-thirds of samples, and in no single case more than one mutational event was found simultaneously. Gene expression profiling of a subset of 31 tumors performed using cDNA microarray chips showed no strong differences in global gene expression among the different cases. However, a supervised analysis of the obtained data identified a subset of genes differentially expressed in tumors carrying BRAF mutation or RTK rearrangement. PMID- 15273716 TI - CAS promotes invasiveness of Src-transformed cells. AB - CAS ('Crk-associated substrate') is an Src substrate found at sites of integrin mediated cell adhesion and linked to cell motility and survival. In this study, the involvement of CAS in oncogenic transformation was evaluated through analysis of mouse embryo fibroblast populations expressing an activated Src mutant, either in the presence or absence of CAS expression. CAS was not found to be a critical determinant of either Src-mediated morphologic transformation or anchorage independent growth. However, CAS had a profound effect on other aspects of oncogenic Src function. CAS expression led to a substantial increase in the phosphotyrosine content of FAK and paxillin, supporting a role for CAS as a positive regulator of Src activity at integrin adhesion sites. Importantly, CAS expression resulted in a striking enhancement of the capacity of Src-transformed cells to invade through Matrigel. The increased invasiveness was associated with increased activation of matrix metalloproteinase MMP-2 and formation of large actin-rich podosomal aggregates appearing as ring and belt structures. Thus, elevated CAS-associated tyrosine phosphorylation signaling events occurring at sites of integrin-mediated cell adhesion can have a major role in the development of an invasive cell phenotype. PMID- 15273717 TI - Regulation of caspase-6 and FLIP by the AMPK family member ARK5. AB - Colorectal cancer cells are unique in that they escape Fas-mediated cell death in the presence of Fas ligand, and we recently reported that AMP-activated protein kinase-related kinase 5 (ARK5) suppresses cell death signaling mediated by cell death receptor in Akt-dependent manner. In the current study, therefore, we examined whether ARK5 is involved in the escape from Fas-mediated cell death of colorectal cancer cells. Among 10 cell lines, ARK5 mRNA expression was observed in LoVo, SW480, and SW1116 cell lines. Interestingly, SW480 and SW1116 cell lines, but not LoVo cell line, showed expressions of both Fas ligand (FasL) and Fas mRNAs. SW620 cell line also showed FasL mRNA; however, Fas and ARK5 mRNAs were not detected. Furthermore, well-coincided expression among ARK5, FasL, and Fas mRNAs was observed in tumor tissues from patients with colorectal cancer, suggesting the suppression of FasL/Fas system-induced cell death by ARK5 in colorectal cancer cell lines. Intensive cell death, which was dependent on the FasL/Fas system was encountered when ARK5 antisense RNA (ARK5/AS) was introduced into SW480 cells. FLIP was expressed in only ARK5 mRNA-expressing cell lines, and ARK5/AS induced FLIP cleavage in a caspase-6-dependent manner. Amino-acid sequence analysis of caspase-6 revealed two putative sites of phosphorylation by ARK5 at Ser80 and Ser257. Although active caspase-6 overexpression induced cell death in SW480 and DLD-1 cell lines, SW480 cells, but not DLD-1 cells, exhibited strong resistance to procaspase-6 overexpression. Moreover, mutant caspase-6, in which the Ser257 was substituted by Ala (caspase-6/SA), induced cell death and FLIP degradation, even in SW480 cells. Active ARK5 was found to phosphorylate wild-type caspase-6 in vitro, but not caspase-6/SA, and the prevented activation of caspase-6 was promoted due to its phosphorylation by active ARK5 in vitro. On the basis of the results of this study, we propose that ARK5 negatively regulates procaspase-6 by phosphorylation at Ser257, leading to resistance to the FasL/Fas system. PMID- 15273718 TI - Enhanced retinoid-induced apoptosis of MDA-MB-231 breast cancer cells by PKC inhibitors involves activation of ERK. AB - Retinoids are vitamin A derivatives, which cause growth inhibition, differentiation and/or apoptosis in various cell types, including some breast cancer cells. In general, estrogen receptor (ER)-positive cells are retinoic acid (RA) sensitive, whereas ER-negative cells are resistant. In this report, we show that ER-negative MDA-MB-231 cells are strongly growth inhibited by retinoids in combination with a PKC inhibitor. While neither RA nor GF109203X (GF) has a significant growth inhibitory effect in these cells, RA+GF potently suppress proliferation. We found that RA+GF induce apoptosis, as shown by an increase in fragmented DNA, Annexin-V-positive cells and caspase-3 activation. Apoptosis was also induced by GF in combination with two synthetic retinoids. Expression of phosphorylated as well as total PKC was decreased by GF and this was potentiated by RA. In addition, treatment with GF caused a strong and sustained activation of ERK1/2 and p38-MAPK, as well as a weaker activation of JNK. Importantly, inhibition of ERK but not p38 or JNK suppressed apoptosis induced by RA+GF, indicating that activation of ERK is specifically required. In support of this novel finding, the ability of other PKC inhibitors to cause apoptosis in combination with RA correlates with ability to cause sustained activation of ERK. PMID- 15273719 TI - Isoforms of the APC tumor suppressor and their ability to inhibit cell growth and tumorigenicity. AB - Mutation of the APC tumor suppressor gene is one of the earliest events in the development of most colorectal tumors. The APC gene encodes multiple protein isoforms through a complicated pattern of expression and alternative splicing. The role that each isoform plays in cellular physiology is unknown, although the presence of some of these isoforms in postmitotic cells suggests a role in controlling cell growth or promoting differentiation. Three APC isoforms that differ in their amino-terminal domains were evaluated by gene transfer experiments using a colon cancer cell line that lacks functional APC. All three isoforms alter cellular morphology and affect cell growth by elongating the G1 phase of the cell cycle. The conventional APC and brain-specific APC isoforms suppress the tumorigenic phenotype of cultured cells, while the 0.3 APC isoform does not. In support of these experiments, BrdU incorporation as a marker for S phase entry occurs at a higher level in transiently transfected cells with 0.3 APC when compared to cells transfected with the other isoforms. All three APC isoforms colocalize with microtubules and dramatically reduce beta-catenin activity to the same extent in transiently transfected cancer cells, suggesting that the different effects of each isoform on tumorigenesis may be nontranscriptional in origin. PMID- 15273720 TI - Cbl-b interacts with ubiquitinated proteins; differential functions of the UBA domains of c-Cbl and Cbl-b. AB - Cbl proteins are ubiquitin protein ligases, which ubiquitinate activated tyrosine kinases and target them for degradation. Both c-Cbl and Cbl-b have an ubiquitin associated (UBA) domain at their C-terminal end. We observed that high molecular weight ubiquitinated proteins constitutively coimmunoprecipitated with transfected and endogenous Cbl-b, but not c-Cbl. The binding site for these ubiquitinated proteins was mapped to the UBA domain of Cbl-b (UBAb). GST-fusion proteins containing the UBAb interacted with ubiquitinated proteins and polyubiquitin chains in vitro, whereas those containing the UBA domain of c-Cbl (UBAc) did not. The UBAb had a much greater affinity for polyubiquitin chains than for monoubiquitin. Analysis of the UBAb and UBAc demonstrate that the affinity for ubiquitin is determined by multiple amino-acid differences between the two domains. Overexpression of the UBAb, but not overexpression of the UBAc, inhibited a variety of ubiquitin-mediated processes such as degradation of ubiquitinated proteins (i.e. EGFR, Mdm-2, and Siah-1). This in vivo result is consistent with the differences in ubiquitin binding observed in vitro between the UBAb and UBAc. This difference in ubiquitin-binding may reflect distinct regulatory functions of c-Cbl and Cbl-b. PMID- 15273721 TI - Identification of novel AP-1 target genes in fibroblasts regulated during cutaneous wound healing. AB - Mesenchymal-epithelial interactions are increasingly considered to be of vital importance for epithelial homeostasis and regeneration. In skin, the transcription factor AP-1 was shown to be critically involved in the communication between keratinocytes and dermal fibroblasts. After skin injury, the release of IL-1 from keratinocytes induces the activity of the AP-1 subunits c-Jun and JunB in fibroblasts leading to a global change in gene expression. To identify AP-1 target genes in fibroblasts, which are involved in the process of cutaneous repair, we performed gene expression profiling of wild-type, c-jun- and junB-deficient fibroblasts in response to IL-1, mimicking the initial phase of wound healing. Using a 15K cDNA collection, over 1000 genes were found to be Jun dependent and additional 300 clones showed IL-1 responsiveness. Combinatorial evaluation allowed for the dissection of the specific contribution of either AP-1 subunit to gene regulation. Besides previously identified genes that are involved in cutaneous repair, we have identified novel genes regulated during wound healing in vivo and showed their expression by fibroblasts on wound sections. The identification of novel Jun target genes should provide a basis for understanding the molecular mechanisms underlying mesenchymal-epithelial interactions and the critical contribution of AP-1 to tissue homeostasis and repair. PMID- 15273722 TI - Mitochondrial proliferation during apoptosis induced by anticancer agents: effects of doxorubicin and mitoxantrone on cancer and cardiac cells. AB - Doxorubicin is one of the most largely prescribed antitumor drug for the treatment of breast, liver and colon cancers as well as leukemia, but the cardiotoxicity of this anthracycline derivative limits its clinical use. Although doxorubicin is toxic to both cancer and cardiac cells, there are evidences suggesting that the mechanism of cell death is different for the two cell types. To investigate further this issue, we have compared the proapoptotic effects of doxorubicin and the functionally related anthracenedione compound mitoxantrone, which is also used in the clinic for the treatment of cancer. After evaluating the toxicity of the two drugs to mammary adenocarcinoma MTLn3 cells and H9C2 cardiomyocytes, we dissected the drug-induced apoptotic machinery by measuring the effects on the cell cycle progression, DNA condensation and fragmentation, production of endogenous peroxides and caspase activation. Both doxorubicin and mitoxantrone are potent inducers of apoptosis in H9C2 cardiomyocytes and MTLn3 breast cancer cells, but there are significant differences between the two cell types in terms of kinetics and order of the events. In particular, flow cytometry measurements of drug-induced changes in mitochondrial transmembrane potential and mitochondrial mass with different fluorescent probes suggested that the two drugs induced a progressive increase in mitochondrial mass in the cancer cells but not in the cardiac cells. The hypothesis was validated by means of electron microscopy, which revealed a significant increase in the number of mitochondria in drug-treated MTLn3 but not in H9C2 cells. The mitochondrial proliferation precedes the nuclear apoptosis in doxorubicin-treated MTLn3 cells. The changes in the architecture and number of mitochondria are linked to the drug-induced perturbation of the cell cycle progression and apoptosis. The proliferation of mitochondria could explain the higher toxicity of doxorubicin to cancer cells compared to cardiac cells and this suggests novel therapeutic opportunities to better control the cardiotoxicity of anthracyclines. PMID- 15273723 TI - Deregulation of the carbohydrate (chondroitin 4) sulfotransferase 11 (CHST11) gene in a B-cell chronic lymphocytic leukemia with a t(12;14)(q23;q32). AB - The t(12;14)(q23;q32) breakpoints in a case of B-cell chronic lymphocytic leukemia (B-CLL) were mapped by fluorescence in situ hybridization (FISH) and Southern blot analysis and cloned using an IGH switch-gamma probe. The translocation affected a productively rearranged IGH allele and the carbohydrate (chondroitin 4) sulfotransferase 11 (CHST11) locus at 12q23, with a reciprocal break in intron 2 of the CHST11 gene. CHST11 belongs to the HNK1 family of Golgi associated sulfotransferases, a group of glycosaminoglycan-modifying enzymes, and is expressed mainly in the hematopoietic lineage. Northern Blot analysis of tumor RNA using CHST11-specific probes showed expression of two CHST11 forms of abnormal size. 5'- and 3'-Rapid Amplification of cDNA Ends (RACE) revealed IGH/CHST11 as well as CHST11/IGH fusion RNAs expressed from the der(14) and der(12) chromosomes. Both fusion species contained open reading frames making possible the translation of two truncated forms of CHST11 protein. The biological consequence of t(12;14)(q23;q32) in this case presumably is a disturbance of the cellular distribution of CHST11 leading to deregulation of a chondroitin-sulfate dependent pathway specific to the hematopoietic lineage. PMID- 15273724 TI - Role of protein-protein interactions in the antiapoptotic function of EWS-Fli-1. AB - In the majority of Ewing's family tumors, chromosomal translocation t(11;22) leads to aberrant fusion of RNA-binding protein EWS with DNA-binding ETS transcriptional factor Fli-1. EWS-Fli-1 has altered the transcriptional activity and modulating its downstream target genes through this transcriptional activity is thought to be responsible for this tumor. We have previously shown that both EWS-Fli-1 and Fli-1 have antiapoptotic activity against several apoptotic inducers. Here, we show that the transcriptional activity of EWS-Fli-1 and Fli-1 is not essential for its antiapoptotic activity. We also demonstrate that EWS-Fli 1 and Fli-1 interact with CBP through its amino-terminal region and inhibit the CBP-dependent transcriptional activity of RXR. This activity appears to be independent of DNA-binding activity of EWS-Fli-1. Introduction of the dominant negative form of CBP into Ewing's sarcoma cells sensitizes these cells against genotoxic or retinoic-acid induced apoptosis. These results suggest that the ability of EWS-Fli-1/Fli-1 to target transcriptional cofactor(s) and modulate apoptotic pathways may be responsible for its antiapoptotic and tumorigenic activities. PMID- 15273725 TI - Ha-Ras(G12V) induces senescence in primary and immortalized human esophageal keratinocytes with p53 dysfunction. AB - Oncogenic Ras induces premature senescence in primary cells. Such an oncogene induced senescence involves activation of tumor suppressor genes that provide a checkpoint mechanism against malignant transformation. In mouse, the ARF-p53 pathway mediates Ha-Ras(G12V)-induced senescence, and p19(ARF-/-) and p53(-/-) cells undergo transformation upon Ras activation. In addition, mouse cells, unlike human cells, express constitutively active telomerase and have long telomeres. However, it is unclear how Ras activation affects human cells of epithelial origin with p53 mutation and/or telomerase activation. In order to address this question, Ha-Ras(G12V) was expressed ectopically in primary as well as hTERT-immortalized human esophageal keratinocytes stably expressing dominant negative p53 mutants. In human esophageal keratinocytes, we found that Ha Ras(G12V) induced senescence regardless of p53 status and telomerase activation. Ras activation resulted in changes of cellular morphology, activation of senescence-associated beta-galactosidase, and suppression of cell proliferation, all coupled with reduction in the hyperphosphorylated form of the retinoblastoma protein (pRb). Furthermore, Ha-Ras(G12V) upregulated p16(INK4a) and downregulated cyclin-dependent kinase Cdk4 in human esophageal keratinocytes. Thus, Ras mediated senescence may involve distinct mechanisms between human and mouse cells. Inactivation of the pRb pathway may be necessary for Ras to overcome senescence and transform human esophageal epithelial cells. PMID- 15273726 TI - Tumor suppressor ARF inhibits HER-2/neu-mediated oncogenic growth. AB - HER2/neu, a receptor tyrosine kinase oncogene, promotes mitogenic growth and antiapoptotic activity in cancer cells. Strong expression of HER2/neu in cancers has been associated with poor prognosis. Alternative reading frame protein (ARF), a tumor suppressor protein encoded by a gene located in the Ink4a/ARF gene locus, is frequently inactivated in human cancers. Little is known about the tumor suppressor role of ARF in HER2/neu-overexpressing cancers. Here, we applied the ARF gene as a tumor-suppressive agent for HER2/neu-overexpressing cells under the control of a tetracycline (tet)-regulated gene expression system. We found that ARF antagonized protein kinase B (PKB)/Akt-mediated p27Kip1 phosphorylation and increased p27 stability in HER2/neu-overexpressing cells. ARF expression also led to decreased levels of Cul1 and Skp2, two proteins involved in p27 degradation. We also found that ARF caused apoptosis in HER2/neu-overexpressing cells, and sensitized cells to apoptosis induced by the chemotherapeutic agents taxol and 2 methoxyestradiol. Most significantly for clinical application, we found that ARF inhibited HER2/neu-mediated cell growth, transformation, and tumorigenesis. These findings indicate that modulation of ARF activity may be a useful therapeutic intervention in HER2-overexpressing cancers. PMID- 15273727 TI - Adenoviral-mediated mda-7 expression suppresses DNA repair capacity and radiosensitizes non-small-cell lung cancer cells. AB - The melanoma differentiation-associated gene-7 (mda-7) was identified by virtue of its enhanced expression in human melanoma cells induced into terminal differentiation. Enforced expression of mda-7 in human cancer cell lines of diverse origins results in the suppression of growth and induction of apoptosis. We have shown that adenoviral-mediated mda-7 (Ad-mda7) radiosensitizes non-small cell lung cancer (NSCLC) cells by enhancing the apoptotic pathway. To identify the mechanism of this radiosensitization, we examined the level of proteins involved in the nonhomologous end-joining (NHEJ) pathway of DNA double-strand break (DSB) repair. Western blot analysis indicated that the expression of NHEJ pathway components Ku70, XRCC4, and DNA ligase IV was downregulated in NSCLC cells--A549 with Ad-mda7 treatment. No such change was observed in normal human CCD16 fibroblasts previously shown not to be radiosensitized by Ad-mda7. The biological significance of these changes of expression of proteins critical for repair of radiation-induced DSBs was confirmed via the analysis of DSB rejoining kinetics using pulsed field gel electrophoresis and assessment of host cell reactivation capacity following Ad-mda7 treatment. Based on these results, we hypothesize that Ad-mda7 sensitizes NSCLC cells to ionizing radiation by suppressing the activity of NHEJ, a pathway essential for repair of radiation induced DSBs. PMID- 15273728 TI - Role of pescadillo in the transformation and immortalization of mammalian cells. AB - The murine and human homologs of the zebrafish pescadillo protein (Pes1 and PES1, respectively) play important roles in ribosome biogenesis and DNA replication. We investigated the effect of Pes1 on the growth of mouse embryo (3T3-like) fibroblasts and conditionally immortalized human fibroblasts expressing the SV40 T antigen (AR5 cells). Increased expression of Pes1 causes transformation of mouse and human fibroblasts in culture (colony formation in soft agar). Although Pes1 can replace the SV40 T antigen in inducing colony formation in soft agar, it cannot substitute the T antigen in the immortalization of human fibroblasts, indicating that it distinguishes between the two functions. As the biological effects of Pes1 are similar to those of the insulin receptor substrate-1 (IRS-1), we investigated the interactions of Pes1 with IRS-1 itself and with the SV40 T antigen. The Pes1 protein (which localizes to the nuclei and nucleoli of cells) interacts with both IRS-1 and the SV40 T antigen, and markedly decreases the interaction of T antigen with p53. Taken together, these results suggest mechanisms for the ability of Pes1 to transform cells, and its failure to immortalize them. PMID- 15273729 TI - Direct FGF receptor 1 activation through an anti-idiotypic strategy mimicks the biological activity of FGF-2 and inhibits the progression of the bladder carcinoma derived from NBT-II cells. AB - The hypothesis that tumor growth is angiogenesis-dependent has been documented by a considerable body of direct and indirect experimental data. Since the discovery of the vascular endothelial growth factor (VEGF), most attention has been focused on the VEGF system. Although fibroblast growth factors 1 and 2 (FGF-1 and FGF-2) can exert a strong angiogenic activity when they are supplied as a single pharmacological agent, their role in pathological angiogenesis in preclinical models remains controversial. To decipher the contribution of FGF receptors in various models of angiogenesis, we took advantage of the anti-idiotypic strategy to obtain circulating agonists specific for FGFR-1 and FGFR-2 (AIdF-1 and AIdF 2). They mimicked FGF-1 and FGF-2 for receptor binding, signal transduction, proliferation of endothelial cells and differentiation of the bladder carcinoma cell NBT-II which expresses FGFR-2b but not FGFR-1. The constitutive expression of FGFR-1 allowed binding of FGF-2 and AIdF-2 and inhibition of the proliferation of NBT-II cells. AIdF-1 and AIdF-2 induced angiogenesis in the corneal pocket assay. Although FGFR-1 dimerization achieved by AIdF-2 injection led to highly differentiated and smaller NBT-II tumors, no sign of reduction of tumor angiogenesis was observed, thus suggesting that endothelial cells are resistant to FGF. PMID- 15273730 TI - 5-aza-2'-deoxycytidine upregulates caspase-9 expression cooperating with p53 induced apoptosis in human lung cancer cells. AB - Treating lung cancer cell lines using low-dose 5-aza-2'-deoxycytidine (DAC) caused an accumulation of procaspase-9 through mRNA upregulation, but the cells did not undergo apoptosis. However, when cells were treated with DAC and infected with a low dose of a recombinant wild-type p53 adenovirus vector (Ad-p53), a synergistic growth inhibitory effect was observed. Combination treatment induced Apaf-1 and procaspase-9 expression in which cytochrome c releases by Ad-p53 triggered the mitochondrial pathway of apoptosis. Selective blockage of caspase-9 activities by Z-LEHD-FMK completely attenuated DAC-induced enhancement of apoptosis mediated by Ad-p53 infection, and ectopic overexpression of procaspase 9 sensitized cells to Ad-p53-induced apoptosis in p53-null cells. In addition, DAC sensitized lung cancer cells to cisplatin and paclitaxel. Induction of the mitochondrial pathway of apoptosis using a slightly toxic dose of DAC may therefore be a strategy for treating lung cancer, and DAC treatment may have clinical implications when combined with chemotherapy or apoptosis-inducing gene therapy. PMID- 15273731 TI - Induction of centrosome amplification and chromosome instability in p53-null cells by transient exposure to subtoxic levels of S-phase-targeting anticancer drugs. AB - Chromosome instability (CIN) is one of the most important phenotypes in tumor progression, introducing multiple mutations required for acquisition of further malignant characteristics. Abnormal amplification of centrosomes, which is frequently observed in human cancer, has been shown to contribute to CIN by increasing the frequency of mitotic defects. Here, we show that transient exposure to subtoxic concentrations of commonly used anticancer drugs that target DNA synthesis induces centrosome amplification in cells lacking p53 tumor suppressor protein, by allowing continuous centrosome duplication in the absence of DNA synthesis. When these cells are released from cell cycle arrest by removal of drugs, cells suffer extensive destabilization of chromosomes. Considering that p53 is the most frequently mutated gene in human cancer and that CIN is known to be associated with acquisition of malignant phenotypes, our observations may explain why recurrent tumors, after chemotherapy, often exhibit more malignant characteristics than the original tumors. The tumor cells that are exposed to subtoxic levels of DNA synthesis-targeting drugs will be arrested and undergo centrosome amplification. Upon cessation of chemotherapy, these cells will re enter cell cycling, and experience extensive CIN due to the presence of amplified centrosomes. This in turn promotes generation of tumor cells equipped with further malignant characteristics. PMID- 15273732 TI - Genetic alterations of the MYH gene in gastric cancer. AB - Increased oxygen free radicals produced in gastric mucosa by H. pylori induce DNA damage and lead to dyspalsia and gastric cancers. However, only a small percentage of individuals that carry H. pylori develop gastric cancer, indicating that other factors are involved. We have screened a set of 95 sporadic gastric cancers for mutations and allele loss of the DNA glycosylase MYH gene, which excises adenine misincorporated opposite unrepaired 8-oxoG. Two of 95 cancers had bi-allelic mutations of the MYH gene with somatic missense mutation of one allele and loss of the remaining allele. The mutations were missense mutations, P391S and Q400R, in exon 13 encoding NUDIX hydrolase domain of the gene. The patients were H. pylori-positive and the tumors were of advanced intestinal-type gastric cancer with lymph node metastasis. In addition, 4 (17.4%) of 23 informative cases showed allele loss at MYH locus. Therefore, our data suggest that somatic mutations of base excision repair gene MYH contribute to the development of a sub set of sporadic gastric cancers. PMID- 15273733 TI - PTEN can inhibit in vitro organotypic and in vivo orthotopic invasion of human bladder cancer cells even in the absence of its lipid phosphatase activity. AB - Recent studies have found a higher frequency of the PTEN tumor-suppressor gene alterations in invasive bladder carcinoma than in superficial disease, suggesting that PTEN is important in this process. A role of PTEN in bladder cancer invasion is further suggested by the fact that PTEN is a regulator of cell motility, a necessary component of tumor invasion. However, it is unknown whether PTEN is mechanistically involved in 'in vivo' tumor invasion or merely an epiphenomenon and, if the former is true, whether this process is dependent on its protein or lipid phosphatase activities. To address these issues, we stably transfected several commonly used human bladder cancer cell lines with known invasive phenotypes with either wild-type PTEN constructs or those deficient in the lipid phosphatase (G129E) or both protein and lipid phosphatase (G129R) activities. Here we show that chemotaxis was inhibited by both the wild-type and G129E mutant of PTEN but not by G129R-transfected cells. Using a novel organotypic in vitro invasion assay, we evaluated the impact of wild-type and mutant PTEN transgene expression on the invasive ability of T24T, a human bladder cancer cell line with a functionally impaired PTEN. Results indicate that the G129E mutant blocks invasion as efficiently as wild-type PTEN transfection. In contrast to the wild type gene, this mutant has no effect on cell clonogenicity in agar. To further establish the role of PTEN in tumor invasion, we evaluated vector- and PTEN transfected T24T cells in an orthotopic in vivo assay that faithfully reproduces human disease. Microscopic examination of murine bladders at the completion of this experiment parallels the results obtained with the organotypic assay. Our results are the first demonstration: (1) that the inhibitory effects of PTEN on cell motility translate into suppression of in vivo invasion; (2) that PTEN can inhibit tumor invasion even in the absence of its lipid phosphatase activity; (3) how organotypic in vitro approaches can be used as surrogates of in vivo invasion allowing rapid dissection of molecular processes leading to this phenotype while reducing the number of animals used in research. PMID- 15273734 TI - Sensitization for anticancer drug-induced apoptosis by the chemopreventive agent resveratrol. AB - Current attempts to improve the survival of cancer patients largely depend on strategies to target tumor cell resistance. Naturally occurring dietary compounds such as resveratrol have gained considerable attention as cancer chemopreventive agents. Here, we report that resveratrol acts as potent sensitizer for anticancer drug-induced apoptosis by inducing cell cycle arrest, which in turn resulted in survivin depletion. Concomitant analysis of cell cycle and apoptosis revealed that pretreatment with resveratrol resulted in cell cycle arrest in S phase and apoptosis induction preferentially out of S phase upon subsequent drug treatment. Likewise, cell cycle arrest in S phase by cell cycle inhibitors enhanced drug induced apoptosis. Resveratrol-mediated cell cycle arrest sensitized for apoptosis by downregulating survivin expression through transcriptional and post transcriptional mechanisms. Similarly, downregulation of survivin expression using survivin antisense oligonucleotides sensitized for drug-induced apoptosis. Importantly, downregulation of survivin and enhanced drug-induced apoptosis by resveratrol occurred in various human tumor cell lines irrespective of p53 status. Thus, this combined sensitizer (resveratrol)/inducer (cytotoxic drugs) concept may be a novel strategy to enhance the efficacy of anticancer therapy in a variety of human cancers. PMID- 15273735 TI - Involvement of cadherins 7 and 20 in mouse embryogenesis and melanocyte transformation. AB - We have determined the expression profiles of cdh7, and the related cdh20 during development. Both transcripts are found in the adult brain, but only cadherin-20 mRNA was detected during embryogenesis. In mouse embryos, cadherin-20 is synthesized by the forebrain, anterior neural ridge, developing visual system, primitive external granular layer of the cerebellum and a subset of neural crest cells likely to develop into melanoblasts. We found that the other embryonic tissues in which cadherin-20 was synthesized depended on genetic background. Melanoma cell lines contained transcripts for cadherin-7 but not for cadherin-20. The majority of the malignant melanoma cell lines produced N-cadherin (N-Cad) and/or cadherin-7 whereas melanocyte cell lines did not. The converse was observed for E-cadherin (E-Cad). Our data suggest that during development cadherin-20 is a key player in compartmentalization of the neural tube and establishment of neural circuitry. Finally, during oncogenesis, cadherin-7, N-cad and E-cad could be used as an efficient marker set for melanoma. PMID- 15273736 TI - Promoter hypermethylation downregulates RUNX3 gene expression in colorectal cancer cell lines. AB - It was recently reported that RUNX3 gene expression is significantly downregulated in human gastric cancer cells due to hypermethylation of its promoter region or hemizygous deletion (Cell, 109, 2002). To verify the genetic alterations and methylation status of the RUNX3 gene in colorectal carcinogenesis, we analysed for mutations, loss of heterozygosity (LOH), and RUNX3 gene promoter hypermethylation, in 32 colorectal cancer cell lines. RT-PCR analysis showed undetectable or low RUNX3 expression in 16 cell lines, and no mutations were found in the RUNX3 gene by PCR-SSCP analysis. Of these 16 cell lines, hypermethylation of the RUNX3 promoter was confirmed in 12. The following observations were made: (i) RUNX3 was re-expressed after 5-aza-2'-deoxycytidine treatment, (ii) the RUNX3 promoter was found to be methylated by MS-PCR, and (iii) hypermethylation of the RUNX3 promoter was confirmed by direct sequencing analysis after sodium bisulfite modification in the above 12 cell lines. RUNX3 was neither methylated nor expressed in four cell lines. Of these four, microsatellite instability (MSI) at the RUNX3 locus was found in three, SNU-61 (D1S246), SNU-769A, and SNU-769B (D1S199). This study suggests that transcriptional repression of RUNX3 is caused by promoter hypermethylation of the RUNX3 CpG island in colorectal cancer cell lines, and the results of these experiments may contribute to an understanding of the role of RUNX3 inactivation in the pathogenesis of colorectal cancers. PMID- 15273737 TI - Tumor necrosis factor alpha sensitizes malignant cells to chemotherapeutic drugs via the mitochondrial apoptosis pathway independently of caspase-8 and NF-kappaB. AB - The Hodgkin cell line HD-MyZ is resistant to apoptosis induced by tumor necrosis factor alpha (TNFalpha). In the present work, we show that pretreatment with TNFalpha sensitized the cells to apoptosis induced by antineoplastic agents and ceramide. TNFalpha pretreatment resulted in enhanced cleavage and activity of caspase-3 upon addition of etoposide, epirubicin or ceramide. No caspase-8 activation was detectable, although caspase-8 could be activated in cell-free extracts. Inhibition of caspase-8 by z-IETD-fmk did not block the sensitizing effect of TNFalpha. Furthermore, exogenous ceramide, a mediator of TNFalpha signaling, could not substitute for TNFalpha in sensitization to drug-induced apoptosis. In contrast, we observed mitochondrial changes following cotreatment of cells with TNFalpha and drugs. Mitochondrial permeability transition, cytochrome c release and subsequent processing of caspase-9 preceded the onset of apoptosis, and were enhanced by TNFalpha pretreatment. Interestingly, although transcription factor NF-kappaB protected HD-MyZ cells from drug-induced apoptosis, TNFalpha-mediated sensitization was independent of NF-kappaB, since overexpressing a dominant-negative IkappaB mutant did not alter the TNFalpha effect. Sensitization for drug-induced apoptosis by TNFalpha was abrogated by Bcl x(L). Thus, the sensitizing effect of TNFalpha is mediated by the mitochondrial pathway and involves processing of caspase-2, -3 and -9, but appears to be independent of caspase-8 processing, Bid cleavage and NF-kappaB signaling. Therefore, sensitization by TNFalpha is mediated at least in part through different pathways, as reported for TRAIL. There, sensitization occurs through a FADD/caspase-8-dependent mechanism. Regarding TNFalpha, the sensitizing effect was also observed in myeloid leukemia cells. Therefore, TNFalpha or alternate molecules activating its pathways might be useful as sensitizers for chemotherapy in hematological malignancies. PMID- 15273738 TI - INK4a/Arf is required for suppression of EGFR/DeltaEGFR(2-7)-dependent ERK activation in mouse astrocytes and glioma. AB - Amplification of the epidermal growth factor receptor (EGFR) or expression of its constitutively activated mutant, DeltaEGFR(2-7), in association with the inactivation of the INK4a/Arf gene locus is a frequent alteration in human glioblastoma. The notion of a cooperative effect between these two alterations has been demonstrated in respective mouse brain tumor models including our own. Here, we investigated underlying molecular mechanisms in early passage cortical astrocytes deficient for p16(INK4a)/p19(Arf) or p53, respectively, with or without ectopic expression of DeltaEGFR(2-7). Targeting these cells with the specific EGFR inhibitor tyrphostin AG1478 revealed that phosphorylation of ERK was only abrogated in the presence of an intact INK4a/Arf gene locus. The sensitivity to inhibit ERK phosphorylation was independent of ectopic expression of DeltaEGFR(2-7) and independent of the TP53 status. This resistance to downregulate the MAPK pathway in the absence of INK4a/Arf was confirmed in cell lines derived from our mouse glioma models with the respective initial genetic alterations. Thus, deletion of INK4a/Arf appears to keep ERK in its active, phosphorylated state insensitive to an upstream inhibitor specifically targeting EGFR/DeltaEGFR(2-7). This resistance may contribute to the cooperative tumorigenic effect selected for in human glioblastoma that may be of crucial clinical relevance for treatments specifically targeting EGFR/DeltaEGFR(2-7) in glioblastoma patients. PMID- 15273739 TI - Comparison of gene-expression profiles between diffuse- and intestinal-type gastric cancers using a genome-wide cDNA microarray. AB - Gastric cancer is the fourth leading cause of cancer-related death in the world. Two histologically distinct types of gastric carcinoma, 'intestinal' and 'diffuse', have different epidemiological and pathophysiological features that suggest different mechanisms of carcinogenesis. A number of studies have investigated intestinal-type gastric cancers at the molecular level, but little is known about mechanisms involved in the diffuse type, which has a more invasive phenotype and poorer prognosis. To clarify the mechanisms that underlie its development and/or progression, we compared the expression profiles of 20 laser microbeam-microdissected diffuse-type gastric-cancer tissues with corresponding noncancerous mucosae by means of a cDNA microarray containing 23,040 genes. We identified 153 genes that were commonly upregulated and more than 1500 that were commonly downregulated in the tumors. We also identified a number of genes related to tumor progression. Furthermore, comparison of the expression profiles of diffuse-type with those of intestinal-type gastric cancers identified 46 genes that may represent distinct molecular signatures of each histological type. The putative signature of diffuse-type cancer exhibited altered expression of genes related to cell-matrix interaction and extracellular-matrix (ECM) components, whereas that of intestinal-type cancer represented enhancement of cell growth. These data provide insight into different mechanisms underlying gastric carcinogenesis and may also serve as a starting point for identifying novel diagnostic markers and/or therapeutic targets for diffuse-type gastric cancers. PMID- 15273740 TI - AMID is a p53-inducible gene downregulated in tumors. AB - AMID, also called PRG3, is an AIF-homologous and mitochondria-associated protein that has been implicated in caspase-independent apoptosis. In this report, we demonstrated that human AMID gene promoter was activated by p53 in reporter gene assays. Chromatin immunoprecipitation experiments indicated that p53 could bind to human AMID promoter. Deletion mutagenesis indicated that human AMID promoter contains two p53-responsive elements. Furthermore, expression array analysis indicated that human AMID mRNA expression was downregulated in a majority of human tumors. Our findings point to the possibility that AMID is a p53-downstream gene involved in tumorigenesis. PMID- 15273741 TI - Interactions of EGFR and caveolin-1 in human glioblastoma cells: evidence that tyrosine phosphorylation regulates EGFR association with caveolae. AB - Epidermal growth factor receptor (EGFR) amplification and type III mutation (EGFRvIII), associated with constitutive tyrosine kinase activation and high malignancy, are commonly observed in glioblastoma tumors. The association of EGFR and EGFRvIII with caveolins was investigated in human glioblastoma cell lines, U87MG and U87MG-EGFRvIII. Caveolin-1 expression, determined by RT-PCR, real-time quantitative PCR and Western blot, was upregulated in glioblastoma cell lines (two-fold) and tumors (20-300-fold) compared to primary human astrocytes and nonmalignant brain tissue, respectively. U87MG-EGFRvIII expressed higher levels of caveolin-1 than U87MG. In contrast, the expression of caveolin-2 and -3 were downregulated in glioblastoma cells compared to astrocytes. A colocalization of EGFR, but not of EGFRvIII, with lipid rafts and caveolin-1 was observed by immunocytochemistry. Association of EGFR and EGFRvIII with caveolae, assessed in vitro by binding to caveolin scaffolding domain peptides and in vivo by immunocolocalization studies in cells and caveolae-enriched cellular fraction, was phosphorylation-dependent: ligand-induced phosphorylation of EGFR resulted in dissociation of EGFR from caveolae. In contrast, inhibition of the EGFRvIII constitutive tyrosine phosphorylation by AG1478 increased association of EGFRvIII with caveolin-1. AG1478 also increased caveolin-1 expression and reduced glioblastoma cell growth in a semi-solid agar. The evidence suggests that the phosphorylation-regulated sequestration of EGFR in caveolae may be involved in arresting constitutive or ligand-induced signaling through EGFR responsible for glial cell transformation. PMID- 15273742 TI - TNF-alpha regulates epithelial expression of MMP-9 and integrin alphavbeta6 during tumour promotion. A role for TNF-alpha in keratinocyte migration? AB - Mice deficient in TNF-alpha (TNF-alpha(-/-) mice) are resistant to skin carcinogenesis and expression of MMP-9 is inhibited in TNF-alpha(-/-) mice during skin tumour development. In the early stages of tumour promotion, MMP-9 protein initially localized to the follicular epidermis but subsequently began to accumulate in the interfollicular epidermis of wild-type but not TNF-alpha(-/-) mice. Inhibition of TNF-alpha or MMP-9 function reduced keratinocyte migration in vitro. In addition, a deficiency of TNF-alpha delayed re-epithelialization in vivo and this correlated with reduced MMP-9 expression. Collectively, these data suggest that MMP-9 regulates keratinocyte migration in a TNF-alpha-dependent manner. Expression profiling of genes that control cell adhesion and migration revealed markedly lower levels of the integrin subunits alphav and beta6 in TNF alpha(-/-) compared with wild-type keratinocytes in vitro. alphavbeta6 expression was upregulated by keratinocytes in vitro and during tumour promotion in vivo in a TNF-alpha-dependent manner. Furthermore, alphavbeta6 blockade significantly inhibited keratinocyte migration and TNF-alpha-stimulated MMP-9 expression in vitro. These data illustrate a novel TNF-alpha-dependent mechanism for the control of alphavbeta6 expression and suggest one pathway for TNF-alpha regulation of MMP-9. Increased MMP-9 and alphavbeta6 expression may stimulate epithelial cell migration during tumour formation and may be one mechanism whereby TNF-alpha acts as an endogenous tumour promoter. PMID- 15273743 TI - Self-assembled single-crystal ferromagnetic iron nanowires formed by decomposition. AB - Arrays of perpendicular ferromagnetic nanowires have recently attracted considerable interest for their potential use in many areas of advanced nanotechnology. We report a simple approach to create self-assembled nanowires of alpha-Fe through the decomposition of a suitably chosen perovskite. We illustrate the principle behind this approach using the reaction 2La(0.5)Sr(0.5)FeO(3) --> LaSrFeO(4) + Fe + O(2) that occurs during the deposition of La(0.5)Sr(0.5)FeO(3) under reducing conditions. This leads to the spontaneous formation of an array of single-crystalline alpha-Fe nanowires embedded in LaSrFeO(4) matrix, which grow perpendicular to the substrate and span the entire film thickness. The diameter and spacing of the nanowires are controlled directly by deposition temperature. The nanowires show uniaxial anisotropy normal to the film plane and magnetization close to that of bulk alpha-Fe. The high magnetization and sizable coercivity of the nanowires make them desirable for high-density data storage and other magnetic-device applications. PMID- 15273744 TI - Crystallographic alignment of high-density gallium nitride nanowire arrays. AB - Single-crystalline, one-dimensional semiconductor nanostructures are considered to be one of the critical building blocks for nanoscale optoelectronics. Elucidation of the vapour-liquid-solid growth mechanism has already enabled precise control over nanowire position and size, yet to date, no reports have demonstrated the ability to choose from different crystallographic growth directions of a nanowire array. Control over the nanowire growth direction is extremely desirable, in that anisotropic parameters such as thermal and electrical conductivity, index of refraction, piezoelectric polarization, and bandgap may be used to tune the physical properties of nanowires made from a given material. Here we demonstrate the use of metal-organic chemical vapour deposition (MOCVD) and appropriate substrate selection to control the crystallographic growth directions of high-density arrays of gallium nitride nanowires with distinct geometric and physical properties. Epitaxial growth of wurtzite gallium nitride on (100) gamma-LiAlO(2) and (111) MgO single-crystal substrates resulted in the selective growth of nanowires in the orthogonal [1?[Evec]0] and [001] directions, exhibiting triangular and hexagonal cross sections and drastically different optical emission. The MOCVD process is entirely compatible with the current GaN thin-film technology, which would lead to easy scale-up and device integration. PMID- 15273745 TI - Flow-induced properties of nanotube-filled polymer materials. AB - Carbon nanotubes (CNTs) are under intense investigation in materials science owing to their potential for modifying the electrical conductivity sigma, shear viscosity eta, and other transport properties of polymeric materials. These particles are hybrids of filler and nanoscale additives because their lengths are macroscopic whereas their cross-sectional dimensions are closer to molecular scales. The combination of extended shape, rigidity and deformability allows CNTs to be mechanically dispersed in polymer matrices in the form of disordered 'jammed' network structures. Our measurements on representative network-forming multiwall nanotube (MWNT) dispersions in polypropylene indicate that these materials exhibit extraordinary flow-induced property changes. Specifically, sigma and eta both decrease strongly with increasing shear rate, and these nanocomposites exhibit impressively large and negative normal stress differences, a rarely reported phenomenon in soft condensed matter. We illustrate the practical implications of these nonlinear transport properties by showing that MWNTs eliminate die swell in our nanocomposites, an effect crucial for their processing. PMID- 15273746 TI - Mouse model of Noonan syndrome reveals cell type- and gene dosage-dependent effects of Ptpn11 mutation. AB - Noonan syndrome is a common human autosomal dominant birth defect, characterized by short stature, facial abnormalities, heart defects and possibly increased risk of leukemia. Mutations of Ptpn11 (also known as Shp2), which encodes the protein tyrosine phosphatase Shp2, occur in approximately 50% of individuals with Noonan syndrome, but their molecular, cellular and developmental effects, and the relationship between Noonan syndrome and leukemia, are unclear. We generated mice expressing the Noonan syndrome-associated mutant D61G. When homozygous, the D61G mutant is embryonic lethal, whereas heterozygotes have decreased viability. Surviving Ptpn11(D61G/+) embryos ( approximately 50%) have short stature, craniofacial abnormalities similar to those in Noonan syndrome, and myeloproliferative disease. Severely affected Ptpn11(D61G/+) embryos ( approximately 50%) have multiple cardiac defects similar to those in mice lacking the Ras-GAP protein neurofibromin. Their endocardial cushions have increased Erk activation, but Erk hyperactivation is cell and pathway specific. Our results clarify the relationship between Noonan syndrome and leukemia and show that a single Ptpn11 gain-of-function mutation evokes all major features of Noonan syndrome by acting on multiple developmental lineages in a gene dosage-dependent and pathway-selective manner. PMID- 15273747 TI - Systemic delivery of genes to striated muscles using adeno-associated viral vectors. AB - A major obstacle limiting gene therapy for diseases of the heart and skeletal muscles is an inability to deliver genes systemically to muscles of an adult organism. Systemic gene transfer to striated muscles is hampered by the vascular endothelium, which represents a barrier to distribution of vectors via the circulation. Here we show the first evidence of widespread transduction of both cardiac and skeletal muscles in an adult mammal, after a single intravenous administration of recombinant adeno-associated virus pseudotype 6 vectors. The inclusion of vascular endothelium growth factor/vascular permeability factor, to achieve acute permeabilization of the peripheral microvasculature, enhanced tissue transduction at lower vector doses. This technique enabled widespread muscle-specific expression of a functional micro-dystrophin in the skeletal muscles of dystrophin-deficient mdx mice, which model Duchenne muscular dystrophy. We propose that these methods may be applicable for systemic delivery of a wide variety of genes to the striated muscles of adult mammals. PMID- 15273748 TI - Trans-splicing repair of CD40 ligand deficiency results in naturally regulated correction of a mouse model of hyper-IgM X-linked immunodeficiency. AB - X-linked immunodeficiency with hyper-IgM (HIGM1), characterized by failure of immunoglobulin isotype switching, is caused by mutations of the CD40 ligand (CD40L), which is normally expressed on activated CD4(+) T cells. As constitutive expression of CD40L induces lymphomas, we corrected the mutation while preserving the natural regulation of CD40L using pre-mRNA trans-splicing. Bone marrow from mice lacking CD40L was modified with a lentivirus trans-splicer encoding the normal CD40L exons 2-5 and was administered to syngenic CD40L-knockout mice. Recipient mice had corrected CD40L mRNA, antigen-specific IgG1 responses to keyhole limpet hemocyanin immunization, regulated CD4(+) T-cell CD40L expression after CD3 stimulation in primary and secondary transplanted mice, attenuation of Pneumocystis carinii pneumonia, and no evidence of lymphoproliferative disease over 1 year. Thus, HIGM1 can be corrected by CD40L trans-splicing, leading to functional correction of the genetic defect without the adverse consequences of unregulated expression of the CD40L gene. PMID- 15273749 TI - Kiss-and-run quantal secretion in frog nerve-muscle synapse. AB - Electrophysiological and optical methods were used to study exo- and endocytosis of synaptic vesicles underlying secretion of the neurotransmitter from motor nerve terminals in frog sternocutaneous muscle. Increase in extracellular concentration of K+ or sucrose produced similar increase in the frequency of miniature endplate currents recorded by extracellular microelectrode. Fluorescent microscopy revealed bright spots in nerve terminal during stimulation of secretion with high-potassium solutions in the presence of endocytosis marker FM1 43. These spots corresponded to clusters of synaptic vesicles that passed through the cycles of exo- and endocytosis. Subsequent high-potassium stimulation of exocytosis in normal Ringer solution led to disappearance of marker spots, while in hyperosmotic saline the spots were preserved. No spots were seen after stimulation of neurotransmitter secretion with sucrose in the presence of FM1-43. It is concluded that quantal secretion of the neurotransmitter in frog motor nerve endings can be realized via both complete exocytosis of synaptic vesicles with subsequent endocytosis and kiss-and-run mechanism with the formation of a temporary pore. PMID- 15273750 TI - Effect of cholinergic and adrenergic receptor blockade on arrhythmogenic activity of endothelin-1 during inhibition of nitric oxide synthesis in awake mice. AB - We studied the effects of blockade of nicotinic receptors in sympathetic and parasympathetic ganglia (hexamethonium), muscarinic receptors (atropine), and beta1-adrenoceptors (atenolol) on arrhythmogenic activity of endothelin-1 during inhibition of nitric oxide synthesis with Nomega-nitro-L-arginine in NMRI mice. Atropine reduced, while hexamethonium completely abolished the arrhythmogenic effect of endothelin-1 during nitric oxide synthase inhibition. Atenolol potentiated arrhythmogenic activity of Nomega-nitro-L-arginine, but endothelin-1 had no effect on the incidence of arrhythmias under these conditions. PMID- 15273751 TI - Mechanisms underlying the effect of E. coli endotoxin on contractile function of lymphatic vessels. AB - E. coli endotoxin decreased the amplitude and frequency of spontaneous phasic contractions in isolated bovine mesenteric lymphatic vessels. This substance in a concentration of 5 mg/liter blocked spontaneous contractions and reduced tonic tension of smooth muscle cells. The dilatory effect of endotoxin on lymphatic vessels was primarily realized via stimulation of synthesis of NO and prostacyclin by endotheliocytes. PMID- 15273752 TI - Relationship between bioelectric activity of neurons in the gigantocellular nucleus of the medulla oblongata and spontaneous movements of chick embryo. AB - Electrophysiological study revealed a correlation between changes in bioelectric activity of the reticular gigantocellular nucleus and movements of chick embryos during ontogeny (16-20 days). This relationship increased by the end of embryogenesis. The reticular gigantocellular nucleus is the major source of supraspinal influences on motor activity during ontogeny. Blockade of proprioceptive impulses with myorelaxin inhibited bioelectric activity of the regulatory gigantocellular nucleus, which attests to the activating effect of proprioception. PMID- 15273753 TI - Effect of defibrillation pulses of different shapes on biomembranes: experimental study. AB - We studied the effects of high-voltage single, double unipolar, and double bipolar electric pulses of exponential or sine shape on erythrocyte membranes. Either single or double (mono- or bipolar) pulses were used. All pulses electroporated the membranes, and the electroporation threshold did not depend on the pulse shape. Two successive pulses decreased erythrocyte number in a nonadditive way. Similar to defibrillation of the whole heart, the effect of two bipolar pulses on erythrocytes was more pronounced than the effect of two unipolar pulses. PMID- 15273754 TI - Individual resistance to cerebral ischemia and negative effect of emotional stress on the course of this disorder. AB - The ratio of low-activity and high-activity rats differed in autumn, winter, and spring litters. Initially more intensive cerebral blood flow in low-active rats and its more pronounced decrease after common carotid artery occlusion determined their higher sensitivity to cerebral ischemia (compared to high-activity animals). After 18-h immobilization stress cerebral blood flow decreased by 10 15%, which abolished the difference in the individual resistance to cerebral ischemia. Independently on emotional resistance, cerebral ischemia was not accompanied by the development of collateral blood flow in the acute period and caused death of 90% rats. PMID- 15273755 TI - Active and passive behavior of animals during the postresuscitation period. AB - Behavioral reactions (open-field test, elevated plus-maze, pain stress, and feeding behavior) were studied in various periods after clinical death caused by circulatory arrest for 10 or 15 min. We revealed two different phases of behavioral changes: active behavior directed at attaining a specific goal and passive behavior directed towards isolation of the organism from external signals and functional minimization. Active behavior determined by pathological excitation in the central nervous system increased the severity of structural damage to hippocampal CA1 neurons during the postresuscitation period. By contrast, passive behavior and minimization of functions preserved structural integrity in these neurons. PMID- 15273756 TI - Effect of locus coeruleus stimulation on ocular hypertension and pathology of pulmonary surfactant during chronic emotional stress. AB - In chronic experiments on rabbits and rats, parameters of eye homeostasis (intraocular pressure and ocular hydrodynamics) and lungs (water balance and surface-active properties of surfactant) were studied during electrical stimulation of the locus coeruleus against the background of chronic stress induced in rabbits by repeated electrical stimulation of the ventromedial nucleus and in rats by daily immobilization on a platform. Stimulation of the locus coeruleus eliminated ocular hypertension of hypothalamic origin and stress induced disturbances in surface activity, blood volume, and water balance in the lungs. PMID- 15273757 TI - Relationship between changes in rat behavior and integral biochemical indexes determined by laser correlation spectroscopy after photothrombosis of the prefrontal cortex. AB - Experiments on rats showed that Noopept improved retention and retrieval of conditioned passive avoidance response after phototrombosis of the prefrontal cortex (a procedure impairing retention of memory traces). The impairment of mnesic functions was accompanied by changes in integral biochemical indexes of the plasma determined by laser correlation spectroscopy. Treatment of behavioral disorders with Noopepet normalized biochemical indexes. PMID- 15273758 TI - Free radical processes in the liver of adult and old rats during stress. AB - We measured the content of free radical lipid peroxidation products, intensity of induced lipid peroxidation, and concentration of carbonylated proteins in the liver of adult (10-12 months) and old rats (22-25 months) subjected to 30-min immobilization. Changes in the intensity of induced lipid peroxidation in homogenates and accumulation of carbonylated proteins in subcellular liver fractions indicate that immobilization stimulates free radical generation in the liver of both adult and old rats. The intensity of this process in old rats surpassed that in adult animals. PMID- 15273759 TI - Mechanisms underlying diquertin-mediated regulation of neutrophil function in patients with non-insulin-dependent diabetes mellitus. AB - We studied the effects of dihydroquercetin (3.3.4.5.7-pentahydroxyflavanone, a new Russian patented preparation) on functional activity of polymorphonuclear neutrophils from patients with non-insulin-dependent diabetes mellitus. Flavonoids (quercetin and its derivative dihydroquercetin) dose-dependently suppressed generation of anion radicals and hypochlorous acid and production of malonic dialdehyde during oxidation of neutrophil membranes. Dihydroquercetin decreased activities of protein kinase C and myeloperoxidase in activated polymorphonuclear neutrophils and could bind transition metals (Fe2+). These properties determine the ability of dihydroquercetin to decrease in vitro functional activity of polymorphonuclear neutrophils from patients with non insulin-dependent diabetes mellitus. PMID- 15273760 TI - Possible role of the phosphoinositide pathway for signal transduction in changes in the sensitivity of delta-opiate receptors during diabetes mellitus. AB - We studied the effects of selective delta-opiate receptor agonists and antagonists on the phosphoinositide pathway in lymphocytes from healthy donors and patients with diabetes mellitus. The test compounds probably play a role in changes in the sensitivity to pharmacological substances binding to delta-opiate receptors during diabetes mellitus. PMID- 15273761 TI - Absorption and digestion of phagocytized objects by mononuclear phagocytes during rheumatoid arthritis. AB - Radioisotope study of mononuclear phagocytes from patients with rheumatoid arthritis showed impaired ingestion of bacteria in the presence of pronounced digestive activity. Excessive accumulation methylumbelliferyl phosphate beta glucuronide (product of hydrolysis catalyzed by glucuronidase released from cells) into the incubation medium was observed. This was probably related to the predominance of extracellular digestion. PMID- 15273762 TI - Effect of M4-cholinoceptor blockade on haloperidol-produced catatonic syndrome in rats. AB - We studied the relationship between the efficiency of muscarinic receptor antagonists in preventing haloperidol-induced catatonia and their activity in tests for the interaction of ligands with various subtypes of muscarinic receptors (M1-M4) in rats. Mathematical modeling showed that affinity of the ligand for M4 receptors positively affects its ability to correct extrapyramidal disorders (catatonic syndrome) produced by haloperidol, while affinity for M2 receptors had a negative effect on this characteristic. PMID- 15273763 TI - Effect of Noolit, a novel lithium preparation, on electrophysiological activity of rat cerebral cortex. AB - The effects of lithium carbonate and Noolit, a novel lithium enterosorbent, on electrophysiological activity of cerebral cortex in rats were compared. Both agents potentiated the theta-, alpha-, and beta-rhythms and modified the response to rhythmic flash stimulation from potentiation to inhibition of cerebral rhythms. Moreover, these drugs increased dispersion of all rhythms. By contrast to lithium carbonate, the effect of Noolit was milder and developed more slowly. PMID- 15273764 TI - Local mechanisms underlying the regulatory effect of Kropanol on hemopoiesis during paradoxical sleep deprivation. AB - We studied the effect of Kropanol on local regulatory mechanisms of hemopoiesis during paradoxical sleep deprivation. The regulatory effect of Kropanol on granulomonocytopoiesis was determined by increased binding capacity of stromal cells in relation to granulocyte-macrophage precursors and increased formation of mixed hemopoietic islets leading to accelerated maturation of granulocyte macrophage precursors and hyperplasia of the granulomonocytic hemopoietic stem. Stimulation of erythropoiesis with Kropanol was associated with increased formation of erythroid hemopoietic islets, accelerated differentiation of erythroid precursors, and high content of erythroid cells in the bone marrow. Kropanol increased proliferative activity of erythroid precursors. Stimulation of these processes depended on enhanced production of short-distance humoral regulators of erythro- and granulomonocytopoiesis. PMID- 15273765 TI - Effects of cholinoblockers on acetylcholine content in rat striatum in neuroleptic-induced parkinsonism. AB - Correction of neuroleptic-induced parkinsonism in rats with two central cholinoblockers atropine and pentifine (acetylene aminoalcohol synthesized at Institute of Toxicology) were studied by measuring the content of acetylcholine in the striatum. The content of the transmitter secretion was estimated from the content of bound acetylcholine fraction in homogenates of the above-mentioned compartment of the brain. The results indicate that atropine and pentifine in doses equally effectively preventing catalepsy in rats had different effects on acetylcholine secretion in the striatum. Hence, cholinolytics with different pharmacological selective effects differently interact with central muscarine receptor subtypes. PMID- 15273766 TI - Amino acid composition of Nephrophyt, a new complex plant preparation and its possible role in correction of mercuric chloride-induced acute renal failure in rats. AB - Amino acid composition of a new Russian nephroprotective drug Nephrophyt (a complex mixture of dry plant extracts) was studied for the first time in order to validate its pharmacological effects in rats with mercuric chloride-induced nephropathy. The concentration of pyrrolidone amino acids and serine in this preparation was extremely high, which is uncommon for plant raw material. These data are in good correlation with pronounced nephroprotective effect and agree with published reports on pharmacological effects of individual amino acids and their metabolites. The prospects of the use of this preparation based on the data on its amino acid composition are discussed. PMID- 15273767 TI - Variability of natural killer activity during immunotherapy. AB - In patients with multiple sclerosis, chronic active hepatitis B, and relapsing genital herpes integral parameters of variability in natural killer activity were higher than in donors. In the dynamics of immunotherapy with IFN preparations (chronic active hepatitis B) and IFN inductor ridostine (relapsing genital herpes) these parameters decreased and approached the normal values. The possibility of using variability of natural killer activity for evaluation of cell function and prediction of the efficiency of immunotherapy is discussed. PMID- 15273768 TI - Effects of secretory products of activated neutrophils on morphological composition and functional activity of peritoneal exudation cells during inflammation of staphylococcal origin. AB - Substance A5 isolated from supernatants of activated neutrophils from donors significantly increases the percentage of neutrophils and macrophages in the peritoneal exudation of mice on days 3 and 7 of staphylococcal inflammation and stimulates functional activity (lysosomal, phagocytic, and NBT-reducing) of these cells, reduced as a result of inflammation, on days 3, 7, and 14 of the inflammatory process. PMID- 15273769 TI - Alternative splicing of murine interleukin-4 mRNA. AB - We found an alternative form of mRNA with spliced second exon (IL-4delta2 mRNA) in mouse bone marrow and splenic cells. At rest, the amount of IL-4 mRNA markedly surpassed that of IL-4delta2 mRNA. Stimulation increased the content of both mRNA forms, but the alternative variant is accumulated more intensively and rapidly. We did not detect predominance of IL-4delta2 mRNA over full-length mRNA variant in the studied mouse tissues. PMID- 15273770 TI - Expression of biomolecular markers (Ki-67, PCNA, Bcl-2, BAX, BclX, VEGF) in breast tumors. AB - We carried out a retrospective immunohistochemical study of Ki-67, PCNA, Bcl-2, BAX, BclX, and VEGF expression in tumors of two groups of breast cancer patients with favorable and unfavorable course of the disease. Considerably enhanced VEGF expression was detected in tumors of patients with early relapses of breast cancer. High VEGF expression was paralleled by high level of Ki-67 and PCNA expression in tumors. It can be hypothesized that expression of VEGF, Ki-67, and PCNA in primary tumor can be used for predicting the course of breast cancer or detecting the patients at a high risk of early relapses. PMID- 15273771 TI - Antioxidant spectrum of blood serum and its peculiarities in children. AB - We compared the contribution of high- and low-molecular-weight antioxidants into total antioxidant activity of blood serum in children and adults. Ten serum samples from children aged from 3 months to 12 years and 6 serum samples from adults were fractionated by chromatography and antioxidant activity and the contents of transferrin and ceruloplasmin were measured in total serum and individual chromatographic fractions. It was found that total antioxidant activity of the serum from children measured in the system of yolk lipoproteins considerably surpassed that in adults. Moreover, in adults the major part in serum antioxidant activity is played by a fist identified high-molecular-weight fraction (600 kDa) and a 67 kDa fraction containing ceruloplasmin and transferrin. Serum antioxidant activity in children was determined only by the high-molecular-weight peak not containing ceruloplasmin and transferrin, which was probably due to significantly lower serum transferrin content in children compared to adults. PMID- 15273772 TI - Immunophenotypical characteristics of permanent cultures of lymphoid cells from Papio hamadryas and Macaca arctoides. AB - Immunophenotypical characteristics of primate cells were studied by enzyme immunoassay and flow cytofluorometry using a panel of monoclonal antibodies to human B- and T-lymphocyte antigens. Specific features of immunophenotype of cultured cells were detected. Simian lymphoid cultures consist of a mixture of B- and T-cells with mosaic antigenic structure expressing markers of B and T cell specificity. PMID- 15273773 TI - Study of local blood flow and pressure in tissues and vessels of hip joint of experimental animals. AB - Acute experiments on dogs showed that clamping of the femoral artery below origination of its deep branch playing the leading role in blood supply to the hip joint increases local blood flow in femoral head due to redistribution of arterial flow in favor of the deep femoral artery. This fact is essential for the development of surgical methods for the treatment of patients with aseptic necrosis of the femoral head. PMID- 15273774 TI - Structural organization of the genome of SARS-associated coronavirus (Strain SoD) isolated on the territory of the Russian Federation. AB - Complete nucleotide cDNA sequence (29715 nucleotides) of SARS-associated coronavirus (strain SoD) isolated for the first time in the territory of the Russian Federation was determined. Phylogenetic analysis revealed maximum similarity between strain SoD genome and Frankfurt 1 strain genome. Three nucleotide substitutions determining two amino acid substitutions were detected. PMID- 15273775 TI - Pathomorphology of vascular bed in different variants of arrhythmogenic heart development. AB - A total of 200 hearts from patients with various forms of cardiosclerosis and pronounced disorders of the heart rhythm were examined postmortem by contrast polypositional cardioventriculography, coronarography, volume and weight cardiometry, and morphometry. Left-ventricular, right-ventricular, and septal variants of arrhythmogenic heart development were distinguished. Left-ventricular variant is characterized by compensatory restructuring of the vascular bed with appreciably increased volume of vascular density mainly in the left ventricle and with the median left type of blood supply. The right-ventricular variant is characterized by signs of compensation and decompensation of circulation and intensification of the right ventricular vascularization. The septal variant is characterized by signs of vascular bed decompensation with more intense vascularization of the septum. The detected diagnostic criteria indicate appreciable structural rearrangement of the coronary bed of arrhythmogenic heart, which explains the essential shifts in stimulation, contraction, and mechanical characteristics of the heart ventricles. PMID- 15273776 TI - Morphology of reparative regeneration in organs and tissues during treatment with new generation Sulfacrylate. AB - We studied morphogenesis of reparative processes in parenchymatous and hollow organs after surgery with the use of Sulfacrylate glue composition characterized by good adhesive and bactericidal properties. The glue rapidly and effectively arrested parenchymatous bleeding, reduced the volume of necrotic tissue, and promoted wound healing without suppuration. Cicatrix was completely formed 1 month after surgery and the glue completely resorbed at this term. The bioglue reliably connected and hermetically sealed sutures when used for gluing intestinal loops and for creation of intestinal anastomoses. The use of biological glue promoted the formation of elastic fibrous tissue not distorting the intestinal lumen. PMID- 15273777 TI - Induction of NO-synthase and acetaldehyde dehydrogenase in neurons of human cerebellar cortex during chronic alcohol intoxication. AB - Induction of NO-synthase and acetaldehyde dehydrogenase in Purkinje cells, basket like neurons, and microvascular endothelium of human cerebellar cortex was detected in patients with chronic alcohol intoxication. It was hypothesized that the neuromodulating effect of NO is associated with mechanisms protecting neurons from toxic effects of ethanol and acetaldehyde. PMID- 15273778 TI - Brazil and the international association for dental research. PMID- 15273779 TI - Pediatric HIV-related oral manifestations: a five-year retrospective study. AB - The purpose of this study was to carry out a five-year retrospective descriptive follow-up of the oral manifestation frequency, systemic condition and type of medication used in HIV-infected children and adolescents after the introduction of combined antiretroviral therapy. Fifty-eight patients were examined in 2001/2002, and their previous medical and dental records (1997 to 2000) were researched from files. There was an occurrence of 7 new cases of AIDS in a sample of 19 children, while 46.5% of the entire sample (n = 58) progressed as to classification of HIV infection. No difference was noted among the frequencies of oral manifestations, categories of the immunosuppression and viral load categories. The oral manifestations in the group of children and adolescents followed up in this study remained stable, even after treatment with combined antiretroviral therapy. However, a downward trend in the frequency of oral candidiasis and parotid enlargement was noted. PMID- 15273780 TI - An infant oral health programme in Goiania-GO, Brazil: results after 3 years of establishment. AB - The objective of this study was to describe the outcomes of an infant oral health programme 3 years after implementation, a programme focused on health education for parents and caries prevention methods for children in a baby clinic. A retrospective cohort study was carried out. The setting was the Infant Oral Health Programme developed at the baby clinic of the State Department of Health, Goiania-GO, Brazil. The sample comprised 100 children who entered the programme from birth to 12 months and were followed for 2 to 3 years. Variables investigated were caries experience, caries risk, and children's behaviour in the dental clinic. The number of children with caries experience was 1 at the initial visit and 8 after the follow-up. There was a dramatic decrease in the number of children in the high risk group, from 51% at the initial visit to only 1% after 2 to 3 years. Children's behaviour in the dental clinic was according to their psychological development. It was concluded that the Infant Oral Health Programme in Goiania showed positive outcomes after 3 years of establishment. Further investigations should evaluate the cost-benefit, as well as the effectiveness of the procedures used in the programme. PMID- 15273781 TI - Effect of 0.02% NaF solution on enamel demineralization and fluoride uptake by deciduous teeth in vitro. AB - The application of 0.02% NaF solution on teeth with a cotton swab instead of brushing with fluoride dentifrice has been suggested for young children to reduce the risk of dental fluorosis, but its anticariogenic effect has not been evaluated. Thus, we studied the in vitro effect of 0.02% NaF solution on enamel demineralization and fluoride uptake in deciduous teeth; non-fluoride dentifrice and fluoride dentifrice (1.100 mug F/g) were used, respectively, as negative and positive controls. The treatment with fluoride dentifrice was more effective in reducing enamel demineralization (p < 0.05) and on fluoride uptake by the enamel (p < 0.05) than the non-fluoride dentifrice and the 0.02% NaF solution. Data suggest that the alternative use of 0.02% NaF solution instead of fluoride dentifrice should be reevaluated especially if dental caries are to be controlled. PMID- 15273782 TI - Polymerization time for a microwave-cured acrylic resin with multiple flasks. AB - This study aimed at establishing the polymerization time of a microwave-cured acrylic resin (AcronTM MC), simultaneously processing 2, 4, and 6 flasks. Required time was measured according to the parameters: monomer release in water, Knoop hardness, and porosity. Samples were made with AcronTM MC in different shapes: rectangular (32 x 10 x 2.5 mm) for monomer release and porosity; and half disc (30 mm in diameter x 4 mm in height) for Knoop hardness. There were four experimental groups (n = 24 per group): G1) one flask (control); G2) two flasks; G3) four flasks, and G4) six flasks. At first, polymerization protocol was similar for all groups (3 min/450 W). Time was then adjusted for G2, G3, and G4, based on monomer release evaluation in the control group, obtained by spectrophotometer Beckman DU-70, with emitting wave of 206 nm. Knoop hardness test was performed using a Shimadzu HMV 2000 hardness tester, and 10 indentations were performed on each specimen's surface. Porosity was assessed after specimens were immersed in black ink and the pores counted in a microscope. Results showed that the complete polymerization of the resin occurred in 4.5 min for two flasks (G2); 8.5 min for four flasks (G3); and 13 min for six flasks (G4), all with 450 W. Statistical analysis revealed that the number of flasks does not interfere with polymerization, Knoop hardness, and porosity of the resin. Results showed that polymerization of microwave-curing resin with more than one flask is a viable procedure, as long as polymerization time is adjusted. PMID- 15273783 TI - Analysis of the prevalence of different topographical characteristics of the residual ridge in mandibular free-end arches. AB - This study observed the prevalence of different types of residual ridge inclination in free-ends of mandibles and reported possible correlative factors that may affect resorption. For this purpose, periapical radiographs and individual data collected from a sample of 64 hemiarches were used. Two radiographs were taken of each free-end, and tracing was employed to determine the angles formed by the resorption configuration in the area of the 1st mandibular molar. The following conclusions were drawn: 1). the great majority of alveolar ridges were distally descending; 2). the average angle was wider for users of mandibular removable partial dentures; 3). the results obtained suggest that the type of opposing maxillary arch affects the inclination of mandibular ridges; 4). greater inclination was observed when the 2nd bicuspids of the mandible were the abutment teeth; 5). no significant correlation was established between age, sex and residual ridge resorption. PMID- 15273784 TI - Computed tomography assessment of Apert syndrome. AB - Apert syndrome, or acrocephalosyndactyly type I, is a craniofacial dysostosis, an autosomal dominant condition characterized by severe developmental disturbances of the craniofacial region including bilateral coronal synostosis associated with midface hypoplasia, exophthalmia, hypertelorism, and symmetric syndactyly of the hands and feet. The aim of this study is to assess the clinical and computed tomography imaging patterns of non-operated patients with Apert syndrome, correlating the bone abnormalities of the cranium, face and the skull base. The study population consisted of 5 patients with Apert syndrome. As part of the craniofacial assessment of the imaging center's routine, all patients underwent clinical evaluation and CT (computed tomograph) exam. Three-dimensional images were generated from helical CT scans, using an independent workstation, to evaluate the craniofacial abnormalities of the syndrome. Clinical exam determined that syndactyly of the hands and feet, pseudocleft in the midline palate and midface hypoplasia were features observed in all of the Apert patients. 3D-CT showed that some abnormalities such as bilateral coronal synostosis, calvarial midline defect and reduction in the antero-posterior dimension of the anterior, medial and posterior cranial fossae were present in all cases. In conclusion, the correlation of clinical and CT imaging findings can be useful to assess the main features observed in Apert patients, improving the criteria for examining the patient and diagnosing this condition, and contributing to the therapeutic planning and surgical follow-up. PMID- 15273785 TI - Influence of soft tissues on mandibular gray scale levels. AB - The purpose of this study was to analyze the gray levels, expressed in pixels, of the mandibular retromolar region, with regard to the influence of muscular and fat soft tissues near this region. Fifteen dry mandibles were X-rayed with the presence of soft tissue simulators. The radiographs were digitized and evaluated by Digora software. A one cm thick layer of wax was used as a simulator of the muscular soft tissue. Animal fat samples of different thicknesses - 0.5, 1.0, 1.5 and 2.0 cm - were used as a simulator of the fat soft tissue. Results showed that the fat soft tissue simulator influenced the gray level values in pixels of the mandibular retromolar region when analyzed in different thicknesses using the Digora digitized image software. PMID- 15273786 TI - Photoelastic analysis of stress distribution on parallel and angled implants after installation of fixed prostheses. AB - The longevity of implant-supported prosthetic rehabilitation depends largely on how the masticatory forces are transferred to the implants and surrounding bone. Anatomical conditions, bone morphology and aesthetics usually dictate implant placement in less than ideal positions for prosthetic rehabilitation and sometimes it is possible to find them with different inclinations. The purpose of this paper was to compare, through photoelastic analysis, the stress distribution in a fixed prosthesis with 3 parallel implants, to the stress distribution in the same prosthesis in the existence of an angled central implant. Two photoelastic resin models were made and a polariscope was used in the visualization of isochromatic fringes formed in the models when axial loads of 2 kg, 5 kg and 10 kg were applied to a unique central point of the prosthesis. The presence of inducted tensions (preloads) was observed in the models after applying torque to the retention screws. Preloads were intensified with the incidence of occlusal forces. In the parallel implants, the force dissipation followed the long axis. The angled implant had a smaller quantity of fringes and the stresses were located mostly around the apical region of the lateral implants. PMID- 15273787 TI - Morphometric characterization of sexual differences in the rat sublingual gland. AB - The presence of morphological differences in the sublingual gland of male and female adult rats was determined by morphometry. Absolute and relative glandular mass was 21% lower and 31% higher, respectively, in females than in males. The fractions of glandular volume occupied by the mixed acini, intercalated ducts and striated ducts did not differ significantly between genders; however, their absolute volume was respectively 29, 42 and 58% higher in males. Despite the differences in the volume of these morphological compartments, the number of cells did not differ significantly between genders, except for the excretory duct compartment, for which a larger number was observed in males. With respect to cell volume, 13, 33 and 47% higher volumes were observed in males for mucous acinar cells and striated and excretory duct cells, respectively, while a 38% higher volume of serous demilune cells was observed for females. The surface-to volume ratio of acini and striated ducts was respectively 16 and 35% higher in females. Based on these results, we conclude that the sublingual gland of female rats possesses smaller acini, and shorter ducts whose caliber is narrower, smaller mucous acinar and larger serous cells than the ones found in the male gland, indicating the presence of sexual dimorphism as well as suggesting sexual differences in the quality of the secreted product. PMID- 15273788 TI - Diffusion of calcitonin through the wall of the root canal. AB - The aim of this study was to evaluate the in vitro diffusion of synthetic salmon calcitonin (CT), used as an intracanal medication, to the external root surface, with or without the presence of intact root cementum. Fifty-four human central incisors were used in the experiment, and were divided into two groups of 21 (test groups) and two groups of 6 teeth (control groups). After root canal preparation, 10 mul of calcitonin was inserted within the root canal chamber. The root was sealed and made externally impermeable. Specimens were then placed in tubes with saline solution buffered with phosphates and stored at 37 degrees C. The diffusion of calcitonin was measured after 1, 4 and 7 days. To count calcitonin present at the external media (PBS), ELISA test (an antigen-antibody reaction) was used. Results showed that there was calcitonin diffusion through dentin in all of the test samples. The absence of cementum increased the diffusion of calcitonin (p=0.05). The highest counts of CT were obtained on day 7 for groups with or without cementum - showing a direct relation between time and diffusion of the medication. PMID- 15273789 TI - The influence of vinegars on exposure of dentinal tubules: a SEM evaluation. AB - Dentin hypersensitivity is a common painful condition observed in clinics. Dietary habits have been much associated with its development and persistence during and following periodontal treatment. The aim of this in vitro study was to evaluate the influence of vinegars on the removal of smear layer and exposure of dentinal tubules. Extracted human teeth were submitted to manual scaling with Gracey curettes in order to remove the cementum as well as to form a smear layer. Dentin samples with 3 mm(2) were obtained and distributed into six experimental groups: one control and five types of vinegars (alcohol, apple, rice, white wine and balsamic). Each group included two methods of vinegar application: topical and friction. After routine preparation for SEM analysis, photomicrographs were assessed by a calibrated and blind examiner using an appropriate index system. Kruskal-Wallis test indicated a significant influence of vinegars on smear layer removal. There was a statistically significant difference between groups treated with apple, white and rice vinegars and the control group (p < 0.05). Nevertheless, Mann-Whitney test indicated that removal of smear layer did not vary with the method of application (topical versus friction) for any of the tested substances. We can conclude that the contact of vinegar may remove smear layer and expose dentinal tubules, regardless of the type of application. However, balsamic vinegar was associated with less removal of smear layer after both methods of application. PMID- 15273790 TI - Clinical and biochemical evaluation of the saliva of patients with xerostomia induced by radiotherapy. AB - Clinical aspects and biochemical properties in the saliva of 21 patients prior to and following radiotherapy for head and neck cancer were evaluated (experimental group) and compared with the same properties in a control group of 21 subjects free of cancer. Salivary flow was evaluated by measuring the time necessary, in seconds, for the output of 2 ml of stimulated saliva; and the buffering capacity changes were determined using a simple colorimetric method. Total salivary protein concentration was determined by the Bradford 4 method. Amylase activity was measured by reducing sugars released from a soluble starch substrate, quantified by the dinitrosalicylic method. The electrophoretic profile was evaluated in polyacrylamide gel (12% SDS-PAGE) using samples of 5 mg of salivary protein. A statistically significant reduction (p < 0.01) of the salivary flow was observed, (162.47 s +/- 28.30 before and 568.71 s +/- 79.75 after irradiation), as well as a reduction in the salivary buffering capacity (pH 5.45 +/- 0.14 before and pH 4.40 +/- 0.15 after irradiation). No statistically significant alteration was observed in total salivary protein concentration. A statistically significant reduction (p < 0.01) of salivary alpha-amylase activity (856.6 ng/mg +/- 88.0 before and 567.0 ng/mg +/- 120.6 after irradiation) was observed. Electrophoretic profile differences in salivary protein bands were also observed after radiotherapy, mainly in the range of molecular weight of 72000 to 55000 Daltons. Clinically, patients presenting xerostomia induced by radiotherapy presented an increase in oral tissue injury. PMID- 15273791 TI - Influence of post-bleaching time intervals on dentin bond strength. AB - It has been reported that bond strength of resin to tooth structure can be reduced when the bonding procedure is carried out immediately after the bleaching treatment. This study evaluated the effect of bleaching of non-vital teeth bleaching on the shear bond strength (SBS) of composite resin/bovine dentin interface and the influence of delaying the bonding procedures for different time intervals following internal bleaching. According to a randomized block design, composite resin cylinders (Z100/Single bond - 3M) were bonded to the flattened dentin surface of two hundred and fifty-six teeth which had previously been subjected to four different treatments: SPH - sodium perborate + 30% hydrogen peroxide; SPW - sodium perborate + distilled water; CP - 37% carbamide peroxide; and CON - distilled water (control), each one followed by storage in artificial saliva for 0 (baseline), 7, 14, and 21 days after bleaching (n = 16). The bleaching agents in the pulp chambers were replaced every 7 days, over 4 weeks. The SBS test of the blocks was done using a universal testing machine. The ANOVA showed that there was no significant interaction between time and bleaching agents, and that the factor time was not statistically significant (p > 0.05). For the factor bleaching treatment, the Student's t-test showed that [CON = CP] > [SPW = SPH]. The bleaching of non-vital teeth affected the resin/dentin SBS values when sodium perborate mixed with 30% hydrogen peroxide or water was used, independently of the elapsed time following the bleaching treatment. PMID- 15273792 TI - In vitro antifungal susceptibility of Candida spp. isolates from patients with chronic periodontitis and from control patients. AB - Superinfection by Candida can be refractory to conventional periodontal treatments in specific situations, such as in immunocompromised patients. In these cases, the systemic therapy with antifungal drugs could be indicated. The aim of this study was to analyse antifungal susceptibility of Candida spp. strains isolated from chronic periodontitis patients and from control individuals. A total of 39 C. albicans isolates, 9 C. tropicalis, 2 C. glabrata and 5 Candida spp. from control individuals and 30 C. albicans, 3 C. tropicalis and 2 C. glabrata from periodontitis patients were tested. In the control group, 1 isolate of C. glabrata was resistant to ketoconazole and 1 Candida spp. was resistant to amphotericin B, ketoconazole and miconazole. Among the isolates of periodontitis group, 1 (3.33%) C. albicans isolate was resistant to flucytosine and ketoconazole. According to the obtained results, it could be concluded that fluconazole was the most effective drug against the several Candida species studied. There were not expressive differences in the susceptibility of isolates from periodontitis patients or from control individuals. PMID- 15273793 TI - Impact of oral health on quality of life among the elderly population of Joacaba, Santa Catarina, Brazil. AB - The objective of this study was to investigate the impact of oral health conditions on the quality of life of elderly people in Joacaba - SC, in Southern Brazil. A survey based on systematic sampling of clusters was carried out with 183 elderly people that belong to old age groups. The survey was conducted in order to assess the oral conditions of the participants (use of and need for prosthesis) based on the criteria from the World Health Organization publication "Oral Health Surveys, Basic Methods", 4th edition. The oral health impact profile (OHIP) was used to evaluate the impact of oral condition in the quality of life. ABIPEME (Brazilian Association of Market Research Institutes) criterion was used, together with the level of education and the number of people in the household to determine social inequalities. The participants were mostly women (82%) and the OHIP mean was 10.35. No correlation was observed between the OHIP level and formal education or between OHIP and number of residents per household. There was a correlation of 0.240 (p = 0.001) between ABIPEME and OHIP. The OHIP mean for those not using maxillary prosthesis was 12.48 and the mean for those using it was 9.81 (p = 0.399). The mean OHIP for those in need of maxillary prosthesis for those who did not need it was 13.00 and 8.88, respectively (p = 0.014). The same trend was found for the use and need for mandibular prosthesis. The conclusion was that the need for maxillary and mandibular prosthesis impacted the quality of life among the elderly population of Joacaba. PMID- 15273794 TI - Clinical and immunopathological spectrum of American cutaneous leishmaniasis with special reference to the disease in Amazonian Brazil: a review. AB - The wide variety of Leishmania species responsible for human American cutaneous leishmaniasis combined with the immune mechanisms of the host results in a large spectrum of clinical, histopathological, and immunopathological manifestations. At the middle of this spectrum are the most frequent cases of localized cutaneous leishmaniasis (LCL) caused by members of the subgenera Leishmania and Viannia, which respond well to conventional therapy. The two pathogenicity extremes of the spectrum generally recognized are represented at the hypersensitivity pole by mucocutaneous leishmaniasis (MCL) and at the hyposensitivity pole by anergic diffuse cutaneous leishmaniasis (ADCL). Following the present study on the clinical, histopathological and immunopathological features of cutaneous leishmaniasis in Amazonian Brazil, we propose the use of the term "borderline disseminated cutaneous leishmaniasis" for the disseminated form of the disease, due to parasites of the subgenera Leishmania and Viannia, which might be regarded as intermediate between LCL and the extreme pathogenicity poles MCL and ADCL. PMID- 15273795 TI - Re-infestation of houses by Triatoma dimidiata after intra-domicile insecticide application in the Yucatan peninsula, Mexico. AB - In most countries, Chagas disease transmission control remains based on domestic insecticide application. We thus evaluated the efficacy of intra-domicile cyfluthrin spraying for the control of Triatoma dimidiata, the only Chagas disease vector in the Yucatan peninsula, Mexico, and monitored potential re infestation every 15 days for up to 9 months. We found that there was a re infestation of houses by adult bugs starting 4 months after insecticide application, possibly from sylvatic/peridomicile areas. This points out the need to take into account the potential dispersal of sylvatic/peridomestic adult bugs into the domiciles as well as continuity action for an effective vector control. PMID- 15273796 TI - Population dynamics of intermediate snail hosts of Fasciola hepatica and some environmental factors in San Juan y Martinez municipality, Cuba. AB - The variation of abundances of intermediate snail hosts of Fasciola hepatica in Cuba (Fossaria cubensis and Pseudosuccinea columella) was studied during one year under natural conditions at five sampling sites in San Juan y Martinez municipality, Pinar del Rio province, Cuba. The effect of some environmental variables on the lymnaeid abundances was also studied. A canonical correspondence analysis showed that both species do not generally occur together in the same habitat and that most factors affect them in an opposite fashion, although both of them correlate positively through time to the diversity of the habitats. F. cubensis prefers the sites that are in or closer to the city whereas P. columella is more abundant in rural sites. Lymnaeid abundances are mainly affected by nitrite and nitrate concentrations as well as by the abundance of the thiarid Tarebia granifera. F. cubensis is more abundant in polluted habitats with low densities (or absence) of T. granifera whereas P. columella prefers cleaner habitats and can coexist with the thiarid, even at its higher densities. The implications of divergent preferences of the two lymnaeids for the control of fasciolosis are discussed. PMID- 15273797 TI - Epidemiology of Chagas disease in Jaguaruana, Ceara, Brazil. I. Presence of triatomines and index of Trypanosoma cruzi infection in four localities of a rural area. AB - In order to assay the triatomine infestation and domiciliation in the rural area of Jaguaruana district, state of Ceara, Brazil, we studied, from November 2000 to April 2002, 4 localities comprising 158 domiciles as a whole, with an average of 4 inhabitants/house, who are dwelling in there for more than 7 years. Most houses have tile-covered roofs and the walls built with plaster-covered bricks (57%), followed by bricks without plaster (33%), and mud walls (7.5%). A total of 3082 triatomines were captured from different locations, according to the following capture plan: (a). intradomiciles: 238 Triatoma brasiliensis, 6 T. pseudomaculata, 9 Rhodnius nasutus, and 2 Panstrongylus lutzi; (b). peridomiciles (annexes): 2069 T. brasiliensis, 223 T. pseudomaculata, 121 R. nasutus, and 1 P. lutzi; (c). wild, in carnauba palms (Copernicia prunifera): 413 R. nasutus. From the captured triatomines, 1773 (57.5%) were examined. The natural index of Trypanosoma cruzi infection ranged from 10.8% to 30.2% (average of 17%), depending on the species and the location from where the triatomines were captured. PMID- 15273798 TI - Prevalence of blood parasites in Tyrannidae (flycatchers) in the Eastern plains of Colombia. AB - Blood samples from 159 birds of the New-world family Tyrannidae (the flycatchers) from the eastern plains of Colombia, were examined for haematozoa parasites, in 1999-2000. Haematozoa were detected in six of 20 species. The overall prevalence was 10.1%. The most common parasites detected were microfilariae, followed by Trypanosoma and Plasmodium. The highest prevalence (9.6%) was found in the Ochre bellied Flycatcher (Mionectes oleaginea). Mixed infections with more than one genus of blood parasite were rare and most infections encountered were of low intensity. The results of this study suggest an important role of ecologically diverse conditions determining composition, transmission, and prevalence of a blood parasite fauna, presumably through host interaction population density. Some new host parasite relationship records are presented. PMID- 15273799 TI - Freshwater snails and Schistosomiasis mansoni in the state of Rio de Janeiro, Brazil: IV - Sul Fluminense Mesoregion. AB - In this paper, the forth of a series dealing with the survey of freshwater gastropods of the state of Rio de Janeiro, the results of collections carried out in the Sul Fluminense Mesoregion from 2000 to 2002 are presented and revealed the occurrence of 18 species: Antillorbis nordestensis; Biomphalaria glabrata; Biomphalaria peregrina; Biomphalaria straminea; Biomphalaria tenagophila; Drepanotrema anatinum; Drepanotrema cimex; Drepanotrema lucidum; Ferrissia sp.; Gundlachia ticaga; Gundlachia sp.; Heleobia sp.; Lymnaea columella; Melanoides tuberculatus; Physa acuta; Physa marmorata; Pomacea sordida and Pomacea sp. As to the snail hosts of Schistosoma mansoni the most frequent species was B. tenagophila, found in all municipalities surveyed, except Parati. Besides new records the present study extends the distribution of B. peregrina and B. straminea in the state. No specimens were found harbouring larval forms of S. mansoni although different kinds of cercariae had been observed. An account about the current schistosomiasis transmission sites in this Mesoregion is presented as well. PMID- 15273800 TI - Prevalence of human papillomavirus type 16 variants in the Federal District, Central Brazil. AB - We report the prevalence of human papillomavirus type 16 (HPV-16) variants in women with cervical lesions from the Federal District, Central Brazil. We analyzed 34 HPV-16 samples, identifying the sequence variations of E6 and L1 genes and correlating variant frequency with disease status. The most prevalent HPV-16 variant was the European (50%), followed by Asian-American (41.2%), African-1 (5.9%), and African-2 (2.9%). European and non-European variants appeared in equal frequencies among the cytological types of lesions - atypical squamous or glandular cells of undetermined significance, cytological alterations suggesting HPV infection, cervical intraepithelial neoplasias, squamous cell carcinoma, and adenocarcinoma. PMID- 15273801 TI - Pulmonary tuberculosis: evaluation of interferon-gamma levels as an immunological healing marker based on the response to the Bacillus Calmette-Guerin. AB - Tuberculosis (TB) is a disease caused by Mycobacterium tuberculosis whose interaction with the host may lead to a cell-mediated protective immune response. The presence of interferon gamma (IFN-gamma) is related to this response. With the purpose of understanding the immunological mechanisms involved in this protection, the lymphoproliferative response, IFN-gamma and other cytokines like interleukin (IL-5, IL-10), and tumor necrosis factor alpha (TNF-alpha) were evaluated before and after the use of anti-TB drugs on 30 patients with active TB disease, 24 healthy household contacts of active TB patients, with positive purified protein derivative (PPD) skin tests (induration > 10 mm), and 34 asymptomatic individuals with negative PPD skin test results (induration < 5 mm). The positive lymphoproliferative response among peripheral blood mononuclear cells of patients showed high levels of IFN-gamma, TNF-alpha, and IL-10. No significant levels of IL-5 were detected. After treatment with rifampicina, isoniazida, and pirazinamida, only the levels of IFN-gamma increased significantly (p < 0.01). These results highlight the need for further evaluation of IFN-gamma production as a healing prognostic of patients treated. PMID- 15273802 TI - Leishmania braziliensis: partial control of experimental infection by interleukin 12 p40 deficient mice. AB - Resistance to infection by Leishmania major has been associated with the development of a Th1 type response that is dependent on the presence of interleukin 12 (IL-12). In this work the involvement of this cytokine in the response to infection by L. braziliensis, a less virulent species in the mouse model, was evaluated. Our results show that while interferon (IFN-gamma) deficient (-/-) mice inoculated L. braziliensis develop severe uncontrolled lesions, chronic lesions that remained under control up to 12 weeks of infection were observed in IL-12p40 -/- mice. IL 12p40 -/- mice had fewer parasites in their lesions than IFN-gamma (-/-) mice. Lymph node cells from IL-12p40 -/- were capable of producing low but consistent levels of IFN-gamma suggestive of its involvement in parasite control. Furthermore, as opposed to previous reports on L. major-infected animals, no switch to a Th2 response was observed in IL-12p40 /- infected with L. braziliensis. PMID- 15273803 TI - The sexual behaviour of Panstrongylus megistus (Hemiptera: Reduviidae): an experimental study. AB - The factors affecting the sexual behaviour of Panstrongylus megistus were studied under laboratory conditions. A general description of mating behaviour is presented for this species. The effect of the time elapsed after the first imaginal feeding on the mating frequency, the motivation of males to mate and the rejection behaviour by females, were analyzed. We also determined the number of copulas accepted by females of this species. Finally, the possible existence of a sexual chemical signal promoting male aggregation around mating couples was evaluated. Results showed that mating frequency increased with the time elapsed since the first adult meal. Despite the number of male copulatory attempts did not change as a function of time, the rejection behaviour of females became gradually less frequent. Females rejected mating by means of body flattening on the substrate, abdominal movements, evasion or stridulation. After a single copula, females did not usually accept to mate again. Neither male nor female aggregation around mating couples was observed, suggesting the absence of a sexual assembling pheromone in P. megistus. PMID- 15273804 TI - Genomic characterization of a Brazilian TT virus isolate closely related to SEN virus-F. AB - SEN virus (SENV) is a circular, single stranded DNA virus that has been first characterized in the serum of a human immunodeficiency virus type 1 (HIV-1) infected patient. Eight genotypes of SENV (A-H) have been identified and further recognized as variants of TT virus (TTV) in the family Circoviridae. Here we describe the first genomic characterization of a SENV isolate (5-A) from South America. Using 'universal' primers, able to amplify most, if not all, TTV/SENV genotypes, a segment of > 3 kb was amplified by polymerase chain reaction from the serum of an HIV-1 infected patient. The amplicon was cloned and a 3087 nucleotide sequence was determined, that showed a high (85%) homology with the sequence of the Italian isolate SENV-F. Proteins encoded by open reading frames (ORFs) 1 to 4 consisted of 758, 129, 276, and 267 amino acids, respectively. By phylogenetic analysis, isolate 5-A was classified into TTV genotype 19 (phylogenetic group 3), together with SENV-F and TTV isolate SAa-38. PMID- 15273805 TI - Virulence markers and antimicrobial susceptibility of bacteria of the Bacteroides fragilis group isolated from stool of children with diarrhea in Sao Paulo, Brazil. AB - Bacteroides fragilis has been isolated from several human and non-human monomicrobial and mixed infections. In this study, some virulence markers and the antimicrobial susceptibility of bacteria of the B. fragilis group isolated from children's stools were evaluated. All the 64 isolates showed the following characteristics: capsulated, beta-hemolytic, hydrophilic, and serum-resistant. Only, 24 (37.5%) strains were resistant at 60 masculine C, for 30 min, and among them, 12 (18.75%) were resistant at 60 masculine C, for 60 min. Also, none strain was resistant at 100 masculine C. Four strains were able to hemagglutinate erythrocytes and D-mannose, D-galactose, D-arabinose, and D-xylose inhibited hemagglutination in 2 B. fragilis strains (p76a, p76b). The hemagglutination in the strain B. uniformis p3-2 was inhibited by D-xylose and D-galactose. The bft gene detection and the enterotoxin production were observed only in 13 EF enterotoxigenic species. Fragilysin activity was confirmed on HT-29 cells. The antimicrobial determination confirmed that both imipenem and metronidazole were efficient against B. fragilis species; all the strains were resistant to lead and nickel. Plasmids of 2.9, 4.4, 4.8, and 8.9 kb were observed in 6 tested strains. These results show the values of the species identification from clinical infections, as well as of the periodic evaluation of the resistance patterns of the B. fragilis group at Brazilian medical institutions. PMID- 15273806 TI - The in vitro cytopathology of a porcine and the simian (SA-11) strains of rotavirus. AB - Rotaviruses have been implicated as the major causal agents of acute diarrhoea in mammals and fowls. Experimental rotavirus infection have been associated to a series of sub-cellular pathologic alterations leading to cell lysis which may represent key functions in the pathogenesis of the diarrhoeic disease. The current work describes the cytopathic changes in cultured MA-104 cells infected by a simian (SA-11) and a porcine (1154) rotavirus strains. Trypan blue exclusion staining showed increased cell permeability after infection by both strains, as demonstrated by cell viability. This effect was confirmed by the leakage of infected cells evaluated by chromium release. Nuclear fragmentation was observed by acridine orange and Wright staining but specific DNA cleavage was not detected. Ultrastructural changes, such as chromatin condensation, cytoplasm vacuolisation, and loss of intercellular contact were shown in infected cells for both strains. In situ terminal deoxynucleotidyl transferase (Tunel) assay did not show positive result. In conclusion, we demonstrated that both strains of rotavirus induced necrosis as the major degenerative effect. PMID- 15273807 TI - Survey of antimicrobial susceptibility patterns of the bacteria of the Bacteroides fragilis group isolated from the intestinal tract of children. AB - The bacteria of the Bacteroides fragilis group are considered important clinical pathogens and they are the most common anaerobes isolated from human endogenous infections. In this study, the susceptibility patterns to antibiotics and metals of 114 species of the B. fragilis group isolated from children with and without diarrhea were determined. Susceptibility was assayed by using an agar dilution method with Wilkins-Chalgren agar. All B. fragilis strains were resistant to lead and nickel, but susceptible to metronidazole and imipenem. beta-lactamase production was detected by using biological and nitrocefin methods, respectively, in 50% and 90.6% of the isolates of children with diarrhea and in 60% and 90% of the isolates of children without diarrhea. Our results show an increase of antibiotics and metals resistance in this microbial group, and a periodic evaluation of the antimicrobial susceptibility is needed. In Brazil, the contamination for antibiotics or metal ions is often observed, and it is suggested an increase the antimicrobial resistance surveillance of this microbial group, mainly those isolated from children's diarrhea. PMID- 15273808 TI - Anti-leishmanial activity of alkaloidal extract from Aspidosperma ramiflorum. AB - Infections due to protozoa of the genus Leishmania are a major worldwide health problem, with high endemicity in developing countries. The drugs of choice for the treatment of leishmaniasis are the pentavalent antimonials (SbV), which present renal and cardiac toxicity. Besides, the precise chemical structure and mechanism of action of these drugs are unknown up to date. In order to find new drugs against leishmaniasis, we have been studying extracts of Brazilian trees. In the present study, we have evaluated the effectiveness of an alkaloid extract of Aspidosperma ramiflorum Muell. Arg. (Apocynaceae), against the extracellular forms promastigotes of L. (L.) amazonensis and L. (V.) braziliensis. The alkaloid extract of A. ramiflorum was much more effective against L. (L.) amazonensis (LD50 < 47 microg/ml) than L. (V.) braziliensis. Based on these in vitro results against L. (L.) amazonensis new studies should be made to find the compounds with anti-leishmanial activity. PMID- 15273809 TI - Effect of thiadiazine derivatives on intracellular amastigotes of Leishmania amazonensis. AB - Current therapy for leishmaniasis is not satisfactory. We describe the in vitro antiproliferative effects of new thiadiazine derivatives against Leishmania amazonensis. The compounds were found to be active against the amastigote form of the parasite, inhibiting parasite growing, from 10 to 89%, at a concentration of 100 ng/ml. This activity suggests that thiadiazine derivatives could be considered as potential antileishmanial compounds. PMID- 15273810 TI - Application of control measures for infections caused by multi-resistant gram negative bacteria in intensive care unit patients. AB - Multi-resistant gram-negative rods are important pathogens in intensive care units (ICU), cause high rates of mortality, and need infection control measures to avoid spread to another patients. This study was undertaken prospectively with all of the patients hospitalized at ICU, Anesthesiology of the Hospital Sao Paulo, using the ICU component of the National Nosocomial Infection Surveillance System (NNIS) methodology, between March 1, 1997 and June 30, 1998. Hospital infections occurring during the first three months after the establishment of prevention and control measures (3/1/97 to 5/31/97) were compared to those of the last three months (3/1/98 to 5/31/98). In this period, 933 NNIS patients were studied, with 139 during the first period and 211 in the second period. The overall rates of infection by multi-resistant microorganisms in the first and second periods were, respectively, urinary tract infection: 3.28/1000 patients/day; 2.5/1000 patients/day; pneumonia: 2.10/1000 patients/day; 5.0/1000 patients/day; bloodstream infection: 1.09/1000 patients/day; 2.5/1000 patients/day. A comparison between overall infection rates of both periods (Wilcoxon test) showed no statistical significance (p = 0.067). The use of intervention measures effectively decreased the hospital bloodstream infection rate (p < 0.001), which shows that control measures in ICU can contribute to preventing hospital infections. PMID- 15273811 TI - Role of enhanced detoxication in a deltamethrin-resistant population of Triatoma infestans (Hemiptera, Reduviidae) from Argentina. AB - Deltamethrin and other pyrethroids have been extensively used in Argentina since 1980, for the chemical control of Triatoma infestans Klug (Hemiptera: Reduviidae). Recently, resistance to deltamethrin was detected in field populations by the survival of bugs exposed by topical application to the diagnostic dose estimated on the CIPEIN susceptible strain. Results of the current study showed low resistant ratios (RRs) to deltamethrin for the resistant populations (RR ranged from 2.0 for San Luis colony to 7.9 for Salta colony). Biochemical studies were made on the most resistant colony (Salta) and the susceptible strain (CIPEIN), in order to establish the importance of degradative mechanisms as a cause of the detected resistance. Esterase activity was measured on 3 days old first instars through phenylthioacetate and a-naphtyl acetate activities. The results showed a significant difference in no cholinesterase esterase activity from susceptible (7.6 +/- 0,7 micro M S./i.min.) and Salta resistant colony (9.5 +/- 0.8 microM S./i.min.). Cytochrome p450 mono-oxygenase (p450) activity was measured on individual insects through ethoxycoumarine deethylase (ECOD) activity using a fluorescence microplate reader. The dependence of ECOD activity on age and body region of the nymphs, and pH and time of incubation were studied in order to optimize the measurement. As a result, comparative studies were performed on abdomens of 2 days old first instars at pH 7.2 and 4 h incubation time. ECOD activity of first nymphs was significantly lower in the susceptible colony (61.3 +/- 9.08 pg ECOD/ insect) than in the resistant one (108.1+/- 5.7 pg ECOD/ insect). These results suggest that degradative esterases (no-cholinesterase) and mono-oxygenases cytochrome p450, play an important role in the resistance to deltamethrin in Salta colony from Argentina. PMID- 15273812 TI - A high-copy T7 Escherichia coli expression vector for the production of recombinant proteins with a minimal N-terminal His-tagged fusion peptide. AB - We report here the construction of a vector derived from pET3-His and pRSET plasmids for the expression and purification of recombinant proteins in Escherichia coli based on T7 phage RNA polymerase. The resulting pAE plasmid combined the advantages of both vectors: small size (pRSET), expression of a short 6XHis tag at N-terminus (pET3-His) and a high copy number of plasmid (pRSET). The small size of the vector (2.8 kb) and the high copy number/cell (200 250 copies) facilitate the subcloning and sequencing procedures when compared to the pET system (pET3-His, 4.6 kb and 40-50 copies) and also result in high level expression of recombinant proteins (20 mg purified protein/liter of culture). In addition, the vector pAE enables the expression of a fusion protein with a minimal amino-terminal hexa-histidine affinity tag (a tag of 9 amino acids using XhoI restriction enzyme for the 5'cloning site) as in the case of pET3-His plasmid and in contrast to proteins expressed by pRSET plasmids (a tag of 36 amino acids using BamHI restriction enzyme for the 5'cloning site). Thus, although proteins expressed by pRSET plasmids also have a hexa-histidine tag, the fusion peptide is much longer and may represent a problem for some recombinant proteins. PMID- 15273813 TI - Age-related changes in radiation-induced micronuclei among healthy adults. AB - The aim of the present study was to establish the extent of in vitro radioresponse of lymphocytes among 62 healthy adults of both genders and to estimate the distribution of baseline micronuclei and radiosensitivity among individuals of the study population using the cytochalasin block micronucleus test. A younger study group consisted of 10 males (mean age, 22.4 years; range, 21-27) and 12 females (mean age, 24.8 years; range, 20-29), whereas an older study group consisted of 18 males (mean age, 35.1 years; range, 30-44) and 22 females (mean age, 38.5 years; range, 30-48). For evaluation of radiosensitivity blood samples were irradiated in vitro using 60Co gamma-ray source. The radiation dose employed was 2 Gy, the dose rate 0.45 Gy/min. The study revealed a significant gender effect on baseline micronuclei favoring females (Z = 3.25, P < 0.001), while yields of radiation-induced micronuclei did not differ significantly (Z = 0.56, P < 0.56) between genders. The distribution of baseline micronuclei among the individuals tested followed Poisson distribution in both study groups and in both genders, whereas the distribution of radiosensitivity among individuals of the older study group did not fulfill Poisson expectations (Kolmogorov-Smirnof test, P < 0.01). In contrast to a nonsignificant difference in radiosensitivity between males and females of the same age group (Z = 1.97, P < 0.56), a statistically significant difference in radiosensitivity between younger and older group for both genders was found (Z = 3.03, P < 0.03). Since the individuals tested were healthy, the observed variability in radiation response is considered to be an early effect of ageing. PMID- 15273814 TI - Patterns of intracellular cytokines in CD4 and CD8 T cells from patients with mycobacterial infections. AB - Using a short-term bulk culture protocol designed for an intracellular-staining method based on a flow cytometry approach to the frequencies of cytokine producing cells from tuberculosis and leprosy patients, we found distinct patterns of T cell subset expression. The method also reveals the profile of peak cytokine production and can provide simultaneous information about the phenotype of cytokine-producing cells, providing a reliable assay for monitoring the immunity of these patients. The immune response of Mycobacterium leprae and purified protein derivative (PPD) in vitro to a panel of mycobacteria-infected patients from an endemic area was assessed in primary mononuclear cell cultures. The kinetics and source of the cytokine pattern were measured at the single-cell level. IFN-gamma-, TNF-alpha-, IL-4- and IL-10-secreting T cells were intracytoplasmic evaluated in an attempt to identify M. leprae- and PPD-specific cells directly from the peripheral blood. The analysis by this approach indicated that TNF-alpha was the first (8 h) to be produced, followed by IFN-gamma (16 h), IL-10 (20 h) and IL-4 (24 h), and double-staining experiments confirmed that CD4+ were a greater source of TNF-alpha than of CD8+ T cells (P < 0.05). Both T cell subsets secreted similar amounts of IFN-gamma. We conclude that the protocol permits rapid evaluation of cytokine production by different T cell populations. The method can also be used to define immune status in non-infected and contact individuals. PMID- 15273815 TI - Assessing negative priming by attended distractors in a paper-and-pencil task. AB - The paper-and-pencil digit-comparison task for assessing negative priming (NP) was introduced, using a referent-size-selection procedure that was demonstrated to enhance the effect. NP is indicated by slower responses to recently ignored items, and proposed within the clinical-experimental framework as a major cognitive index of active suppression of distracting information, critical to executive functioning. The digit-comparison task requires circling digits of a list with digit-asterisk pairs (a baseline measure for digit-selection), and the larger of two digits in each pair of the unrelated (with different digits in successive digit-pairs) and related lists (in which the smaller digit subsequently became a target). A total of 56 students (18-38 years) participated in two experiments that explored practice effects across lists and demonstrated reliable NP, i.e., slowing to complete the related list relative to the unrelated list, (F(2, 44) = 52.42, P < 0.0001). A 3rd experiment examined age-related effects. In the paper-and-pencil digit-comparison task, NP was reliable for the younger (N = 8, 18-24 years) and middle-aged adults (N = 8, 31-54 years), but absent for the older group (N = 8, 68-77 years). NP was also reduced with aging in a computer-implemented digit-comparison task, and preserved in a task typically used to test location-specific NP, accounting for the dissociation between identity- and spatial-based suppression of distractors (Rao R(3, 12) = 16.02, P < 0.0002). Since the paper-and-pencil digit-comparison task can be administered easily, it can be useful for neuropsychologists seeking practical measures of NP that do not require cumbersome technical equipment. PMID- 15273816 TI - Preliminary data on the prevalence of psychiatric disorders in Brazilian male and female juvenile delinquents. AB - The aim of the present investigation was to study the prevalence of psychiatric disorders in a sample of delinquent adolescents of both genders and to compare the prevalence between genders. A total of 116 adolescents (99 males and 17 females) aged 12 to 19 on parole in the State of Rio de Janeiro were interviewed using the screening interview based on the Schedule for Affective Disorders and Schizophrenia for School-Age Children -- Present and Lifetime (KSADS-PL). Data were collected between May 2002 and January 2003. Of 373 male and 58 female adolescents present in May 2002 in the largest institution that gives assistance to adolescents on parole in the city of Rio de Janeiro, 119 subjects were assessed (three of them refused to participate). Their average age was 16.5 years with no difference between genders. The screening interview was positive for psychopathology for most of the sample, with the frequencies of the suggested more prevalent psychiatric disorders being 54% for attention deficit/hyperactivity disorder, 77% for conduct disorder, 41% for oppositional defiant disorder, 57% for anxiety disorder 57, 60% for depressive disorder 60, 63% for illicit drug abuse, and 58% for regular alcohol use. Internalizing disorders (depressive disorders, anxiety disorders and phobias) were more prevalent in the female subsample. There was no significant difference in the prevalence of illicit drug abuse between genders. There were more male than female adolescents on parole and failure to comply with the sentence was significantly more frequent in females. The high prevalence of psychopathology suggested by this study indicates the need for psychiatric treatment as part of the prevention of juvenile delinquency or as part of the sentence. However, treatment had never been available for 93% of the sample in this study. PMID- 15273817 TI - Relative contribution of expectancy and immediate arousal to the facilitatory effect of an auditory accessory stimulus. AB - An auditory stimulus speeds up a digital response to a subsequent visual stimulus. This facilitatory effect has been related to the expectancy and the immediate arousal that would be caused by the accessory stimulus. The present study examined the relative contribution of these two influences. In a first and a third experiment a simple reaction time task was used. In a second and fourth experiment a go/no-go reaction time task was used. In each of these experiments, the accessory stimulus preceded the target stimulus by 200 ms for one group of male and female volunteers (GFix). For another group of similar volunteers (GVar) the accessory stimulus preceded the target stimulus by 200 ms in 25% of the trials, by 1000 ms in 25% of the trials and was not followed by the target stimulus in 50% of the trials (Experiments 1a and 1b) or preceded the target stimulus by 200 ms in 6% of the trials and by 1000 ms in 94% of the trials (Experiments 2a and 2b). There was a facilitatory effect of the accessory stimulus for GFix in the four experiments. There was also a facilitatory effect of the accessory stimulus at the 200-ms stimulus onset asynchrony for GVar in Experiments 1a and 1b but not in Experiments 2a and 2b. The facilitatory effects observed were larger in the go/no-go task than in the simple task. Taken together, these results suggest that expectancy is much more important than immediate arousal for the improvement of performance caused by an accessory stimulus. PMID- 15273818 TI - Looking for the GAP effect in manual responses and the role of contextual influences in reaction time experiments. AB - When the offset of a visual stimulus (GAP condition) precedes the onset of a target, saccadic reaction times are reduced in relation to the condition with no offset (overlap condition) - the GAP effect. However, the existence of the GAP effect for manual responses is still controversial. In two experiments using both simple (Experiment 1, N = 18) and choice key-press procedures (Experiment 2, N = 12), we looked for the GAP effect in manual responses and investigated possible contextual influences on it. Participants were asked to respond to the imperative stimulus that would occur under different experimental contexts, created by varying the array of warning-stimulus intervals (0, 300 and 1000 ms) and conditions (GAP and overlap): i) intervals and conditions were randomized throughout the experiment; ii) conditions were run in different blocks and intervals were randomized; iii) intervals were run in different blocks and conditions were randomized. Our data showed that no GAP effect was obtained for any manipulation. The predictability of stimulus occurrence produced the strongest influence on response latencies. In Experiment 1, simple manual responses were shorter when the intervals were blocked (247 ms, P < 0.001) in relation to the other two contexts (274 and 279 ms). Despite the use of choice key-press procedures, Experiment 2 produced a similar pattern of results. A discussion addressing the critical conditions to obtain the GAP effect for distinct motor responses is presented. In short, our data stress the relevance of the temporal allocation of attention for behavioral performance. PMID- 15273819 TI - Antagonist G-mediated targeting and cytotoxicity of liposomal doxorubicin in NCI H82 variant small cell lung cancer. AB - The aim of the present study was to characterize the interactions of antagonist G (H-Arg-D-Trp-N(me)Phe-D-Trp-Leu-Met-NH 2)-targeted sterically stabilized liposomes with the human variant small cell lung cancer (SCLC) H82 cell line and to evaluate the antiproliferative activity of encapsulated doxorubicin against this cell line. Variant SCLC tumors are known to be more resistant to chemotherapy than classic SCLC tumors. The cellular association of antagonist G targeted (radiolabeled) liposomes was 20-30-fold higher than that of non-targeted liposomes. Our data suggest that a maximum of 12,000 antagonist G-targeted liposomes were internalized/cell during 1-h incubation at 37 masculine C. Confocal microscopy experiments using pyranine-containing liposomes further confirmed that receptor-mediated endocytosis occurred, specifically in the case of targeted liposomes. In any of the previously mentioned experiments, the binding and endocytosis of non-targeted liposomes have revealed to be negligible. The improved cellular association of antagonist G-targeted liposomes, relative to non-targeted liposomes, resulted in an enhanced nuclear delivery (evaluated by fluorimetry) and cytotoxicity of encapsulated doxorubicin for incubation periods as short as 2 h. For an incubation of 2 h, we report IC50 values for targeted and non-targeted liposomes containing doxorubicin of 5.7 +/- 3.7 and higher than 200 micro M doxorubicin, respectively. Based on the present data, we may infer that receptors for antagonist G were present in H82 tumor cells and could mediate the internalization of antagonist G-targeted liposomes and the intracellular delivery of their content. Antagonist G covalently coupled to liposomal drugs may be promising for the treatment of this aggressive and highly heterogeneous disease. PMID- 15273820 TI - Effects of estragole on the compound action potential of the rat sciatic nerve. AB - Estragole, a relatively nontoxic terpenoid ether, is an important constituent of many essential oils with widespread applications in folk medicine and aromatherapy and known to have potent local anesthetic activity. We investigated the effects of estragole on the compound action potential (CAP) of the rat sciatic nerve. The experiments were carried out on sciatic nerves dissected from Wistar rats. Nerves, mounted in a moist chamber, were stimulated at a frequency of 0.2 Hz, with electric pulses of 50-100-micros duration at 10-20 V, and evoked CAP were monitored on an oscilloscope and recorded on a computer. CAP control parameters were: peak-to-peak amplitude (PPA), 9.9 +/- 0.55 mV (N = 15), conduction velocity, 92.2 +/- 4.36 m/s (N = 15), chronaxy, 45.6 +/- 3.74 micros (N = 5), and rheobase, 3.9 +/- 0.78 V (N = 5). Estragole induced a dose-dependent blockade of the CAP. At 0.6 mM, estragole had no demonstrable effect. At 2.0 and 6.0 mM estragole, PPA was significantly reduced at the end of 180-min exposure of the nerve to the drug to 85.6 +/- 3.96 and 13.04 +/- 1.80% of control, respectively. At 4.0 mM, estragole significantly altered PPA, conduction velocity, chronaxy, and rheobase (P < or = 0.05, ANOVA; N = 5) to 49.3 +/- 6.21 and 77.7 +/- 3.84, 125.9 +/- 10.43 and 116.7 +/- 4.59%, of control, respectively. All of these effects developed slowly and were reversible upon a 300-min wash out. The data show that estragole dose-dependently blocks nerve excitability. PMID- 15273821 TI - Effects of autacoid inhibitors and of an antagonist on malaria infection in mice. AB - The effects of p-chlorophenylalanine, an inhibitor of serotonin synthesis, indomethacin, an inhibitor of prostaglandin synthesis, cyproheptadine, a serotonin, bradykinin and histamine antagonist, were assessed separately and in combination with chloroquine (CQ) in Vom strains of Swiss albino mice (18-22 g) of either sex infected intraperitoneally with 1 x 10(7) Plasmodium yoelii nigeriensis-induced malaria. As prophylactic, these agents reduced from 31.9 +/- 4.5 to 16.1 +/- 8.1% the level of parasitemia relative to control but had no appreciable activity as curative agents when administered subcutaneously once daily for 4 days after 72 h of parasites innoculum in vivo. However, CQ alone and the combination of these agents with CQ in curative and prophylactic treatments significantly reduced (from 50.3 +/- 5.8 to 4.9 +/- 0.75%) the level of parasitemia (P < 0.05), which was taken only once 72 h after the parasites innoculum. The prophylactic result was shown to produce better results than the curative treatment. The data indicate that inhibitors and an antagonist can reduce the parasitemia load (the extent of damage and the severity of infection) as well as enhance the effects of CQ when combined with it for malaria therapy. The study reveals that the production of autacoids in established infection renders autacoid inhibitors and an antagonist ineffective for radical cure in malarial mice; however, selective inhibition of local hormones implicated in the pathological manifestations of malaria infection by autacoid inhibitors and an antagonist may be a possible pathway to reduce the severity of infection and the associated tissue damage and to enhance the efficacy of available anti-malarials. PMID- 15273822 TI - Anti-inflammatory, antinociceptive and ulcerogenic activity of a zinc-diclofenac complex in rats. AB - We investigated the anti-inflammatory, antinociceptive and ulcerogenic activity of a zinc-diclofenac complex (5.5 or 11 mg/kg) in male Wistar rats (180-300 g, N = 6) and compared it to free diclofenac (5 or 10 mg/kg) and to the combination of diclofenac (5 or 10 mg/kg) and zinc acetate (1.68 or 3.5 mg/kg). The carrageenin induced paw edema and the cotton pellet-induced granulomatous tissue formation models were used to assess the anti-inflammatory activity, and the Hargreaves model of thermal hyperalgesia was used to assess the antinociceptive activity. To investigate the effect of orally or intraperitoneally (ip) administered drugs on cold-induced gastric lesions, single doses were administered before exposing the animals to a freezer (-18 degrees C) for 45 min in individual cages. We also evaluated the gastric lesions induced by multiple doses of the drugs. Diclofenac plus zinc complex had the same anti-inflammatory and antinociceptive effects as diclofenac alone. Gastric lesions induced by a single dose administered per os and ip were reduced in the group treated with zinc-diclofenac when compared to the groups treated with free diclofenac or diclofenac plus zinc acetate. In the multiple dose treatment, the complex induced a lower number of the most severe lesions when compared to free diclofenac and diclofenac plus zinc acetate. In conclusion, the present study demonstrates that the zinc-diclofenac complex may represent an important therapeutic alternative for the treatment of rheumatic and inflammatory conditions, as its use may be associated with a reduced incidence of gastric lesions. PMID- 15273823 TI - Protective effect of N-acetylcysteine against oxygen radical-mediated coronary artery injury. AB - The present study investigated the protective effect of N-acetylcysteine (NAC) against oxygen radical-mediated coronary artery injury. Vascular contraction and relaxation were determined in canine coronary arteries immersed in Kreb's solution (95% O2-5% CO2), incubated or not with NAC (10 mM), and exposed to free radicals (FR) generated by xanthine oxidase (100 mU/ml) plus xanthine (0.1 mM). Rings not exposed to FR or NAC were used as controls. The arteries were contracted with 2.5 microM prostaglandin F2alpha. Subsequently, concentration response curves for acetylcholine, calcium ionophore and sodium fluoride were obtained in the presence of 20 microM indomethacin. Concentration-response curves for bradykinin, calcium ionophore, sodium nitroprusside, and pinacidil were obtained in the presence of indomethacin plus Nomega-nitro-L-arginine (0.2 mM). The oxidative stress reduced the vascular contraction of arteries not exposed to NAC (3.93 +/- 3.42 g), compared to control (8.56 +/- 3.16 g) and to NAC group (9.07 +/- 4.0 g). Additionally, in arteries not exposed to NAC the endothelium dependent nitric oxide (NO)-dependent relaxation promoted by acetylcholine (1 nM to 10 microM) was also reduced (maximal relaxation of 52.1 +/- 43.2%), compared to control (100%) and NAC group (97.0 +/- 4.3%), as well as the NO/cyclooxygenase independent receptor-dependent relaxation provoked by bradykinin (1 nM to 10 microM; maximal relaxation of 20.0 +/- 21.2%), compared to control (100%) and NAC group (70.8 +/- 20.0%). The endothelium-independent relaxation elicited by sodium nitroprusside (1 nM to 1 microM) and pinacidil (1 nM to 10 microM) was not affected. In conclusion, the vascular dysfunction caused by the oxidative stress, expressed as reduction of the endothelium-dependent relaxation and of the vascular smooth muscle contraction, was prevented by NAC. PMID- 15273824 TI - Toxicity of a cyanobacterial extract containing microcystins to mouse lungs. AB - Toxic cyanobacteria in drinking water supplies can cause serious public health problems. In the present study we analyzed the time course of changes in lung histology in young and adult male Swiss mice injected intraperitoneally (ip) with a cyanobacterial extract containing the hepatotoxic microcystins. Microcystins are cyclical heptapeptides quantified by ELISA method. Ninety mice were divided into two groups. Group C received an injection of saline (300 microl, ip) and group Ci received a sublethal dose of microcystins (48.2 microg/kg, ip). Mice of the Ci group were further divided into young (4 weeks old) and adult (12 weeks old) animals. At 2 and 8 h and at 1, 2, 3, and 4 days after the injection of the toxic cyanobacterial extract, the mice were anesthetized and the trachea was occluded at end-expiration. The lungs were removed en bloc, fixed, sectioned, and stained with hematoxylin-eosin. The percentage of the area of alveolar collapse and the number of polymorphonuclear (PMN) and mononuclear cell infiltrations were determined by point counting. Alveolar collapse increased from C to all Ci groups (123 to 262%) independently of time, reaching a maximum value earlier in young than in adult animals. The amount of PMN cells increased with time of the lesion (52 to 161%). The inflammatory response also reached the highest level earlier in young than in adult mice. After 2 days, PMN levels remained unchanged in adult mice, while in young mice the maximum number was observed at day 1 and was similar at days 2, 3, and 4. We conclude that the toxins and/or other cyanobacterial compounds probably exert these effects by reaching the lung through the blood stream after ip injection. PMID- 15273825 TI - Effect of hyperbaric oxygenation on the regeneration of the liver after partial hepatectomy in rats. AB - The aim of the present study was to assess the influence of hyperbaric oxygenation (HBO) on rat liver regeneration before and after partial hepatectomy. Rats were sacrificed 54 h after 15% hepatectomy, liver and body weights were measured, and serum alanine transaminase (ALT) and aspartate transaminase (AST) activity and albumin levels were determined. The lipid peroxide level, as indicated by malondialdehyde production in the remnant liver was measured, and liver sections were analyzed by light microscopy. Five groups of 10 rats in each group were studied. The preHBO and pre-hyperbaric pressure (preHB) groups were treated before partial hepatectomy with 100% O2 and 21% O2, respectively, at 202,650 pascals, daily for 3 days (45 min/day). The control group was not treated before partial hepatectomy and recovered under normal ambient conditions after the procedure. Groups postHBO and postHB were treated after partial hepatectomy with HBO and HB, respectively, three times (45 min/day). The preHBO group presented a significant increase in the initiation of the regeneration process of the liver 54 h postoperatively. The liver/body weight ratio was 0.0618 +/- 0.0084 in the preHBO compared to 0.0517 +/- 0016 g/g in the control animals (P = 0.016). In addition, the preHBO group showed significant better liver function (evaluated by the lowest serum ALT and AST activities, P = 0.002 and P = 0.008, respectively) and showed a significant decrease in serum albumin levels compared to control (P < 0.001). Liver lipid peroxide concentration was lowest in the preHBO group (P < 0.001 vs control and postHBO group) and light microscopy revealed that the composition of liver lobules in the preHBO group was the closest to normal histological features. These results suggest that HBO pretreatment was beneficial for rat liver regeneration after partial hepatectomy. PMID- 15273826 TI - Imidazoline receptors in the heart: a novel target and a novel mechanism of action that involves atrial natriuretic peptides. AB - Chronic stimulation of sympathetic nervous activity contributes to the development and maintenance of hypertension, leading to left ventricular hypertrophy (LVH), arrhythmias and cardiac death. Moxonidine, an imidazoline antihypertensive compound that preferentially activates imidazoline receptors in brainstem rostroventrolateral medulla, suppresses sympathetic activation and reverses LVH. We have identified imidazoline receptors in the heart atria and ventricles, and shown that atrial I1-receptors are up-regulated in spontaneously hypertensive rats (SHR), and ventricular I1-receptors are up-regulated in hamster and human heart failure. Furthermore, cardiac I1-receptor binding decreased after chronic in vivo exposure to moxonidine. These studies implied that cardiac I1 receptors are involved in cardiovascular regulation. The presence of I1-receptors in the heart, the primary site of production of natriuretic peptides, atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP), cardiac hormones implicated in blood pressure control and cardioprotection, led us to propose that ANP may be involved in the actions of moxonidine. In fact, acute iv administration of moxonidine (50 to 150 microg/rat) dose-dependently decreased blood pressure, stimulated diuresis and natriuresis and increased plasma ANP and its second messenger, cGMP. Chronic SHR treatment with moxonidine (0, 60 and 120 microg kg(-1) h(-1), sc for 4 weeks) dose-dependently decreased blood pressure, resulted in reversal of LVH and decreased ventricular interleukin 1beta concentration after 4 weeks of treatment. These effects were associated with a further increase in already elevated ANP and BNP synthesis and release (after 1 week), and normalization by 4 weeks. In conclusion, cardiac imidazoline receptors and natriuretic peptides may be involved in the acute and chronic effects of moxonidine. PMID- 15273827 TI - Hemodynamic and hormonal actions of adrenomedullin. AB - Adrenomedullin, a 52-amino acid residue peptide, has numerous biological actions which are of potential importance to cardiovascular homeostasis, growth and development of cardiovascular tissues and bone, prevention of infection, and regulation of body fluid and electrolyte balance. Studies in man using intravenous infusion of the peptide have demonstrated that, at plasma levels detected after myocardial infarction or in heart failure, adrenomedullin reduces arterial pressure, increases heart rate and cardiac output, and activates the sympathetic and renin-angiotensin systems but suppresses aldosterone. The thresholds for these responses differ, being lower under some experimental circumstances for arterial pressure than for the other biological effects. Adrenomedullin administration inhibits the pressor and aldosterone-stimulating action of angiotensin II in man. By contrast, the pressor effect of norepinephrine is little altered by concomitant adrenomedullin administration. Although in the absence of a safe, specific antagonist of the actions of endogenous adrenomedullin it is difficult to be certain about the physiological and pathophysiological importance of this peptide in man, current evidence suggests that it serves to protect against cardiovascular overload and injury. Hope has been expressed that adrenomedullin or an agonist specific for adrenomedullin receptors might find a place in the treatment of cardiovascular disorders. PMID- 15273828 TI - Enhanced expression of Ang-(1-7) during pregnancy. AB - Pregnancy is a physiological condition characterized by a progressive increase of the different components of the renin-angiotensin system (RAS). The physiological consequences of the stimulated RAS in normal pregnancy are incompletely understood, and even less understood is the question of how this system may be altered and contribute to the hypertensive disorders of pregnancy. Findings from our group have provided novel insights into how the RAS may contribute to the physiological condition of pregnancy by showing that pregnancy increases the expression of both the vasodilator heptapeptide of the RAS, angiotensin-(1-7) [Ang-(1-7)], and of a newly cloned angiotensin converting enzyme (ACE) homolog, ACE2, that shows high catalytic efficiency for Ang II metabolism to Ang-(1-7). The discovery of ACE2 adds a new dimension to the complexity of the RAS by providing a new arm that may counter-regulate the activity of the vasoconstrictor component, while amplifying the vasodilator component. The studies reviewed in this article demonstrate that Ang-(1-7) increases in plasma and urine of normal pregnant women. In preeclamptic subjects we showed that plasma Ang-(1-7) was suppressed as compared to the levels found in normal pregnancy. In addition, kidney and urinary levels of Ang-(1-7) were increased in pregnant rats coinciding with the enhanced detection and expression of ACE2. These findings support the concept that in normal pregnancy enhanced ACE2 may counteract the elevation in tissue and circulating Ang II by increasing the rate of conversion to Ang-(1-7). These findings provide a basis for the physiological role of Ang-(1-7) and ACE2 during pregnancy. PMID- 15273829 TI - Reactive oxygen species and angiotensin II signaling in vascular cells -- implications in cardiovascular disease. AB - Diseases such as hypertension, atherosclerosis, hyperlipidemia, and diabetes are associated with vascular functional and structural changes including endothelial dysfunction, altered contractility and vascular remodeling. Cellular events underlying these processes involve changes in vascular smooth muscle cell (VSMC) growth, apoptosis/anoikis, cell migration, inflammation, and fibrosis. Many factors influence cellular changes, of which angiotensin II (Ang II) appears to be amongst the most important. The physiological and pathophysiological actions of Ang II are mediated primarily via the Ang II type 1 receptor. Growing evidence indicates that Ang II induces its pleiotropic vascular effects through NADPH driven generation of reactive oxygen species (ROS). ROS function as important intracellular and intercellular second messengers to modulate many downstream signaling molecules, such as protein tyrosine phosphatases, protein tyrosine kinases, transcription factors, mitogen-activated protein kinases, and ion channels. Induction of these signaling cascades leads to VSMC growth and migration, regulation of endothelial function, expression of pro-inflammatory mediators, and modification of extracellular matrix. In addition, ROS increase intracellular free Ca2+ concentration ([Ca2+]i), a major determinant of vascular reactivity. ROS influence signaling molecules by altering the intracellular redox state and by oxidative modification of proteins. In physiological conditions, these events play an important role in maintaining vascular function and integrity. Under pathological conditions ROS contribute to vascular dysfunction and remodeling through oxidative damage. The present review focuses on the biology of ROS in Ang II signaling in vascular cells and discusses how oxidative stress contributes to vascular damage in cardiovascular disease. PMID- 15273830 TI - Effects of galantamine on attention and memory in Alzheimer's disease measured by computerized neuropsychological tests: results of the Brazilian Multi-Center Galantamine Study (GAL-BRA-01). AB - OBJECTIVE: To investigate the effects of galantamine on the performance of patients with mild to moderate Alzheimer's disease (AD) in a computerized neuropsychological test battery (CNTB). METHOD: Thirty-three patients with probable AD were treated with galantamine for three months and evaluated in a prospective, open-label, multi-center study. The CNTB and the ADAS-Cog were administered at baseline and after 12 weeks. The CNTB includes reaction time tests to evaluate attention, implicit and episodic memory for faces and words. Statistical comparisons were performed between the results in week 12 versus baseline. Patients who did not reach the therapeutic doses were excluded from the efficacy analysis. RESULTS: Four patients (12.1%) were excluded from the analysis either because of treatment discontinuation (n=3) or because a therapeutic dose was not reached (n=1). The remaining 29 patients were treated with doses of 24 mg/day (n=22) and 16 mg/day (n=7). After 12 weeks, significant reductions in reaction time were seen in the test of episodic memory for faces (p=0.023) and in the test of two-choice reaction time (p=0.039) of the CNTB. CONCLUSION: Treatment with galantamine produced improvement in computerized tests of attention and episodic memory after 12 weeks, leading to statistically significant reduction in the reaction times. PMID- 15273831 TI - Effects of caffeine on visual evoked potential (P300) and neuromotor performance. AB - The stimulant effects of caffeine on cognitive performance have been widely investigated. The visual evoked potential, specially the P300 component, has been used in studies that explain the stimulant mechanisms of caffeine through neurophysiological methods. In this context, the present study aimed to investigate electrophysiological changes (P300 latency) and modification of cognitive and motor performance produced by caffeine. Fifteen healthy volunteers, 9 women and 6 men (26 +/- 5 years, 67 +/- 12.5 kg) were submitted three times to the following procedure: electroencefalographic recording, Word Color Stroop Test, and visual discrimination task. Subjects took a gelatin caffeine capsule (400 mg) or a placebo (P1 and P2), in a randomized, crossover, double-blind design. A one-factor ANOVA and Tukey post hoc test were used to compare dependent variables on the C, P1 and P2 moments. The statistical analyses indicated a non significant decrease in reaction time, Stroop execution time and latency at Cz on the caffeine moment when compared to the others. Moreover, a non-significant increase in Stroop raw score and latency at Pz could be observed. The only significant result was found at Fz. These findings suggest that the positive tendency of caffeine to improve cognitive performance is probably associated with changes in the frontal cortex, a widely recognized attention area. PMID- 15273832 TI - Plasmapheresis in the treatment of myasthenia gravis: retrospective study of 26 patients. AB - We analyzed the experience of Unicamp Clinical Hospital with plasma exchange (PE) therapy in myasthenia gravis (MG). About 17.8 % of a totality of MG patients had PE performed: 26 cases, 19 women and seven men. The mean age-onset of MG was 28 years, extremes 11 and 69. Minimum deficit observed in the group was graded IIb (O & G) or IIIa (MGFA scale). One patient had prethymectomy PE. In seven the procedures were performed due to myasthenic crisis and in 18 patients due to severe myasthenic symptoms or exacerbation of previous motor deficit. Two patients were also submitted to chronic PE considering refractoriness to other treatments. Twenty-six patients had 44 cycles of PE and 171 sessions. The mean number of sessions was 3.9 (SD +/- 1.4) each cycle; median 5, extremes 2 and 6. The mean time by session was 106,5 minutes (SD +/- 35.2); median 100.5 (extremes of 55 and 215). The mean volume of plasma exchanged in each session was 2396 ml (SD +/- 561); median 2225 (extremes 1512 and 4500). Side effects occurred: reversible hypotension (seven cases), mild tremor or paresthesias (seven cases). Infection and mortality rates due to PE were zero. All patients had immediate benefit of each PE cycle and usually they also received prednisone or other immunosuppressors. Good acceptance of the procedure was observed in 80.7% of patients. PMID- 15273833 TI - Clonidine in respiratory panic disorder subtype. AB - OBJECTIVE: Clonidine, which inhibits locus coeruleus discharge, would seem for theoretical reasons to be a good antipanic drug. Panic disorder (PD) presents a heterogeneous cluster of symptoms and a classification based on subtypes has been suggested and the respiratory symptoms group appears as a distinct subtype. METHOD: We report three cases of respiratory PD patients who were successfully treated with clonidine. RESULTS: Patients obtained panic free status, reduced anxiety levels and better functioning after clonidine administration (0.30-0.45 mg/day) for 6 weeks. CONCLUSION: Clonidine can be effective in the treatment of respiratory PD. This drug might play a role in relieving symptoms of anxiety due to noradrenergic hyperactivity in these patients. PMID- 15273834 TI - The effect of methylphenidate on oppositional defiant disorder comorbid with attention deficit/hyperactivity disorder. AB - OBJECTIVE: To assess the effect of methylphenidate on the diagnosis of oppositional-defiant disorder (ODD) comorbid with attention-deficit hyperactivity disorder (ADHD). METHOD: We conducted an open-label study in which 10 children and adolescents with a dual diagnosis of ODD and ADHD were assessed for their ODD symptoms and treated with methylphenidate. At least one month after ADHD symptoms were under control, ODD symptoms were reevaluated with the Parent form of the Children Interview for Psychiatric Syndromes (P-ChIPS). RESULTS: Nine of the 10 patients no longer fulfilled diagnostic criteria for ODD after they were treated with methylphenidate for ADHD. CONCLUSION: Methylphenidate seems to be an effective treatment for ODD, as well as for ADHD itself. The implications for the treatment of patients with ODD not comorbid with ADHD needs further investigation. PMID- 15273835 TI - Use of intrathecal morphine infusion for spasticity. AB - OBJECTIVE: To study the efficacy of intrathecal morphine infusion to treat spasticity. SETTING: Functional Neurosurgery Division of University of Sao Paulo, Brazil. METHOD: Twelve patients with clinical refractory spasticity were studied. Two females and 10 males, with median age of 34.4 years (20 to 61 ys.). The initial Ashworth scale was 4.6. They were submitted to pump implantation for intrathecal morphine infusion. RESULTS: The final Ashworth scale was 2.2. The median dose concentration of Intrathecal morphine was 0.95 mg, with a mean frequency of 1.8 times a day. Four patients developed pruritus, two patients nausea, two patients urinary retention, however all improved after morphine concentration was decreased. One patient was submitted to pump review after extrusion catheter. CONCLUSIONS: Intrathecal morphine infusion is very helpful in patients with spasticity refractory to clinical treatment, and we observed only minor complications. PMID- 15273836 TI - The conformation of the brain plays an important role in the distribution of diffuse axonal injury in fatal road traffic accident. AB - OBJECTIVE: A study was made of the brain lesions in 120 random victims of fatal road traffic accidents to determine the frequency and topographic distribution of diffuse axonal damage (DAI) in relation to the midline brain structures. METHOD: The identification of axons was carried out with a mouse antibody anti neurofilament proteins 70-, 160-, and 210-kD. RESULTS: DAI was identified in 96 (80%) brains and classified as Grade 1 in 21.9%, as Grade 2 in 51%, and as Grade 3 in 27.1% of the patients. In spite of the diffuse distribution that is characteristic of DAI, damage occurred preferentially in the interhemispheric formations (corpus callosum and fornix) and rostral portion of the brainstem, usually to one side of the midline. CONCLUSION: From a mechanical point of view, the interhemispheric formations and the rostral portion of the brainstem act as fixating structures for the cerebral hemispheres during rotational acceleration of the head. It is known that the motion of the cerebral hemispheres is delayed at the points of fixation, where greater stress would be produced, particularly on the side subjected to greater displacement. The frequent involvement by DAI of deep, center-medial brain structures, usually to one side of the midline, supports the mechanism proposed above. PMID- 15273837 TI - Neurological morbidity in vaccine-associated paralytic poliomyelitis in Brazil from 1989 up to 1995. AB - We collected 30 cases of vaccine associated paralytic poliomyelitis (VAPP) from 4081 cases of acute flaccid palsies cases notified from 1989 to 1995 to the Brazilian Ministry of Health. There were 30 VAPP cases with 56% of children younger than 1 year old, 56.7% of female. 46% of cases were reported in the Northeast. Ten P2 vaccine virus, 8 P3 and 2 P1 and associations amongst them were isolated. The clinical pattern in 60 days was: monoplegia (16), paraplegia (6), tetraplegia (5), hemiplegia (2) and triplegia (1). There was no strong relationship between fever, before or after the prodrome period, or the use of intramuscular medication to morbidity. CONCLUSION: if the anti-poliomyelitis strategy adopted in Brazil has lead to the eradication of the poliomyelitis with wild virus infection, the existence of a minimum risk of vaccine-associated poliomyelitis is a matter of concern because there will be a permanent neurological deficit. PMID- 15273838 TI - Sequelae from meningococcal meningitis in children: a critical analysis of dexamethasone therapy. AB - OBJECTIVE: To evaluate the effectiveness of dexamethasone as an adjunctive therapy to antibiotics in children with meningococcal meningitis. METHOD: A total of 81 children diagnosed with meningococcal meningitis hospitalized in sequence were studied at the University Hospital of Sao Paulo University, with the objective of evaluating the presence of sequelae in four different groups of patients, following the administration of dexamethasone: Group I - 25 patients who received the first dose at least 10 minutes before the introduction of the antibiotic therapy; Group II - 19 patients who received the corticosteroid concomitantly; Group III - 14 patients for which the dexamethasone was administered after beginning the antibiotic scheme; Group IV - 23 patients that did not receive dexamethasone. The groups were evaluated for homogeneity through the prognostic indexes and clinical and laboratory characteristics, based on the records obtained at hospitalization. RESULTS: Some degree of sequelae occurred in 16 (26.22%) of the survivors and 23 patients (28.39%) coursed with sequelae or died. The mean period of neurological attendance was 36.97 months and neurological alterations were detected in 16.17% of the patients. No significant difference was found between the four groups. There was also no statistical difference in the comparison of the neurological sequelae in the children from group IV with the children of groups I and II or even with groups I, II and III analyzed as a whole. The presence of hearing loss occurred in 11.11% of the patients, again there was no significant difference between the four groups. Psychological evaluation was performed using the WPSSI and WISC tests. A mild mental disability was detected in one patient from group I and another in group III. The overall analysis of the sequelae (neurological, auditory and intellectual level) also did not demonstrate any significant difference between the four groups. Comparing the children from groups I and II together and also groups I, II and III as a whole with the children in group IV also failed to detect a significant difference arising from the use or nonuse of the corticosteroid. CONCLUSION: Dexamethasone was not proven to be effective in decreasing the number of sequelae among patients with meningococcal meningitis. PMID- 15273839 TI - The pattern of proton magnetic resonance spectroscopy in non-neoplastic encephalic lesions. AB - The purpose of this article is show the role of proton magnetic resonance spectroscopy (MRS), associated with magnetic resonance images, in the study of non-neoplastic disorders, helping in diagnosis and better characterization of the nature of the lesion. Herein, we analyzed single voxel proton spectroscopy in eight different non-neoplastic lesions, displayed in six categories (infectious, ischaemic, demyelinating, inflammatory, malformation of development and phacomatosis). The presence or the ratios of signal intensities brain tissue metabolites observed with this technique (N-acetyl aspartate, choline, creatine, lactate and lipids) helped in their differentiation with neoplastic lesions and helped in correct diagnosis. In infectious diseases, signals of acetate, succinate and amino acids were also important. In conclusion, proton MRS is a noninvasive method, very useful as an additional technique to define the nature of non-neoplastic encephalic lesions. PMID- 15273840 TI - Magnetic resonance imaging of cavernous sinus invasion by pituitary adenoma diagnostic criteria and surgical findings. AB - This study used MRI to define preoperative imaging criteria for cavernous sinus invasion (CSI) by pituitary adenoma (PA). MR images of 103 patients with PA submitted to surgery (48 with CSI) were retrospectively reviewed. The following MR signs were studied and compared to intraoperative findings (the latter were considered the gold standard for CSI detection): presence of normal pituitary gland between the adenoma and CS, status of the CS venous compartments, CS size, CS lateral wall bulging, displacement of the intracavernous internal carotid artery (ICA) by adenoma, grade of parasellar extension (Knosp-Steiner classification) and percentage of intracavernous ICA encased by the tumor. Statistical analysis was performed using qui-square testing and sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) were obtained for each MR finding. The following signs have been found to represent accurate criteria for non-invasion of the CS: 1- normal pituitary gland interposed between the adenoma and the CS (PPV, 100%); 2- intact medial venous compartment (PPV, 100%); 3- percentage of encasement of the intracavernous ICA lower than 25% (NPV, 100%) and 4- medial intercarotid line not crossed by the tumor (NPV, 100%). Criteria for CSI were: 1- percentage of encasement of the intracavernous ICA higher than 45%; 2- occlusion of three or more CS venous compartments and 3- occlusion of the CS lateral venous compartment. The CS was very likely to be invaded if the inferior venous compartment was not detected (PPV. 92,8%), if the lateral intercarotid line was crossed (PPV. 96,1%) or if a bulging lateral dural wall of the CS was seen (PPV, 92,3%). The preoperative diagnosis of CSI by PA is extremely important since endocrinological remission is rarely obtained after microsurgery alone in patients with invasive tumors. The above mentioned MR imaging criteria may be useful in advising most of the patients preoperatively on the potential need for complimentary therapy after surgery. PMID- 15273841 TI - Bizarre behavior during intracarotid sodium amytal testing (Wada test): are they predictable? AB - The intracarotid sodium amytal test (ISAT or Wada Test) is a commonly performed procedure in the evaluation of patients with clinically refractory epilepsy candidates to epilepsy surgery. Its goal is to promote selective and temporary interruption of hemispheric functioning, seeking to define language lateralization and risk for memory compromise following surgery. Behavioral modification is expected during the procedure. Even though it may last several minutes, in most cases it is subtle and easily manageable. We report a series of patients in whom those reactions were unusually bizarre, including agitation and aggression. Apart of the obvious technical difficulties (patients required physical restraining) those behaviors potentially promote testing delay or abortion and more importantly, inaccurate data. We reviewed those cases, seeking for features that might have predicted their occurrence. Overall, reactions are rare, seen in less than 5% of the ISAT procedures. The barbiturate effect, patients' psychiatric profiles, hemisphere dominance or selectiveness of the injection were not validated as predictors. Thorough explanation, repetition and simulation may be of help in lessening the risk of those reactions. PMID- 15273842 TI - [Somatosensory evoked potential in children with evoked spikes by tapping of the feet or hands on electroencephalogram]. AB - OBJECTIVE: To study the characteristics of Somatosensory evoked potential (SEP) cortical components in children with evoked spikes (ES) on EEG. METHOD: The children were aged 7-12 years, had normal neurological examination and neuropsychomotor development, and did not present signs/symptoms of CNS lesions. Data were compared within a group of 20 "normal" children. RESULTS: The amplitude of the cortical components N75 and P98, obtained by posterior tibial nerve stimulation, was higher in ES groups as compared with the normal group. CONCLUSION: High amplitude SEP (N75 and P98) has higher values in the group of children with ES than in the normal group. The exact mechanism involved in the genesis of ES and high amplitude SEP is not clear yet. A possible mechanism would be focal cortical hyperexcitability related to functional activity. No difference was observed in SEP components in children with ES considering occurrence or not of epilepsy. Therefore, SEP does not bring elements to distinguish between the groups suffering or not epilepsy. PMID- 15273843 TI - [Quantitative electroencephalography in children with benign childhood epilepsy with centrotemporal spikes: analysis of band power]. AB - Quantitative EEG aspects are studied in children with benign childhood epilepsy with centrotempral spikes (BCET). A total of 27 children, from 7 to 11 years neurologically and intellectually normal was studied and compared to a control group of normal children. They were submitted to anamnesis, neurological examination, Raven test, digital electroencephalogram and quantitative eletroencephalogram analysis. There was a higher delta, theta, alpha and beta absolute power in most of the electrodes and of alpha and beta for some electrodes in the BCET group. Relative theta power was also higher for the BECT group in most of the electrodes. These findings suggest that in BECT there are diffuse differences form age-matched normal children including a difference in relative spectrum of electrical cerebral activity and that this may be related to a functional immaturity. PMID- 15273844 TI - [School performance in children with benign childhood epilepsy with centrotemporal spikes]. AB - Neuropsychological implications of benign childhood epilepsy with centrotemporal spikes-rolandic spikes (BECTS) have not been adequately investigated. The aim of this study was to compare the results in a school performance test of patients with BECTS and normal age-matched controls. A total of 20 children with BECTS and 20 normal controls were submitted to anamnesis, clinical evaluation, Raven test, school performance test (SPT), digital electroencephalogram and quantitative electroencephalogram analysis. Comparing with normal controls, children with BECTS showed significantly lower SPT results, especially in reading test. There was an association between the higher number of rolandic spikes and inferior performance in SPT reading test. These findings suggest that discharges may be a factor in the genesis of lower performance in reading test in children with BECTS. PMID- 15273845 TI - [Prolonged video-EEG monitoring in nonepileptic seizures: clinical semiology]. AB - The purpose of this study was to point out the effectiveness of prolonged video EEG monitoring (PVEM) in the diagnosis of nonepileptic seizures (NES) as well as to estimate its prevalence in a reference center of epilepsy (EP). A sample of 47 patients with the diagnosis of NES with spontaneous or provoked seizures was observed. A protocol with the clinical history and semiology of seizures was analyzed; Fisher's exact test and cluster analysis were used for statistical observation. The results showed a prevalence of 10% of NES; more prevalence in females (63.8%); the crises were spontaneous in 57% of the patients. The mean age was 32.5 +/- 11 years and the most frequent semiological sign was apparent sleep (82.2%). Either EP or NES was observed in 9% of the patients. There were three groups according to the cluster analysis: hypermotor NES of the extremities with tonus alteration; NES with automatism; and axial NES with eye movements. In conclusion, the study of clinical semiology of NES during the PVEM provides both this nosological entity and the differential EP diagnoses while the provocative test helps to obtain the seizures. PMID- 15273846 TI - [Oral language acquisition: relation and risk for written language]. AB - The present study relates the acquisition of oral language to the development of writing in 236 children of a private school in the city of Porto Alegre, Brazil. The objective of this research was to identify non-linguistic factors involved in phonological acquisition and to describe the relation of phonological acquisition with alterations of writing. At the age of 6 years, kindergarten students were divided into 2 groups, based on the test of Phonological Evaluation of Children. In the follow-up, at 9 years of age, students were evaluated by means of Balanced Dictation and textual production. The comparison of results from case and control groups showed statistically significant difference as to the number of mistakes made in writing, pointing to the acquisition of oral language as a predictive factor for the development of spelling. PMID- 15273847 TI - [Executive functions in children with phenylketonuria: variations as a function of phenilalanine plasm level]. AB - The present study investigates the hypothesis of a specific executive function deficit in children with Phenilketonuria (PKU) whose Phenilalanine level is between 360 and 600 mmol/l. Participants were 21 early and continuously treated 9 month-old children with PKU and 18 9-month-old controls. The children with PKU were divided into two groups on the basis of their mean phenilalanine level prior to the study: the group of children whose level was between 120 and 360 mmol/l, and the group of children whose level was between 360 and 600 mmol/l. Although the three groups did not differ with regard to performance on a test of global mental development, the PKU children with high phenilalanine performed significantly worse than both the low phenilalanine PKU children and the control children on a task that assesses executive functioning. PMID- 15273848 TI - [Importance of the clinical genetics evaluation on hydrocephalus]. AB - The aim of this study was to characterize the possibility of genetic etiology in a group of individuals with congenital hydrocephalus in which the etiology was indeterminate and to confirm that earlier diagnosed. The casuistry was composed by 16 individuals with congenital hydrocephalus. Investigation protocol included anamnesis, familial investigation, physical examination, computerized tomography or magnetic resonance image of head, vertebral column X-ray, karyotype and dysmorphological study. Results were analyzed in two groups. In Group I (3M:9F) was composed by hydrocephalus associated with unspecific signs. Group II (7 males) had findings of epectrum of L1 disease. Genetic counseling could be offered in 11 cases. These results demonstrate the great etiological heterogeneity of congenital hydrocephalus and reinforce the importance of dysmphology evaluation as an important complementary investigation. PMID- 15273849 TI - [Fetal myelomeningocele and the potential in-utero repair: follow-up of 58 fetuses]. AB - INTRODUCTION: Prenatal diagnosis of myelomeningocele (MM) allows planning its management and, recently, a possible in utero repair. OBJECTIVE: To describe the perinatal outcome of fetuses with MM, in a Fetal Medicine Unit, identifying possible candidates for the in utero surgical repair. METHOD: Retrospective and descriptive study of 58 cases of prenatally diagnosed MM, at CAISM-UNICAMP, from January 1997 to December 2001, identifying possible fetal candidates for in utero repair. RESULTS: the diagnosis mean gestational age was 29 weeks (17-39); level of lesions was above sacral region in 85%, association with hydrocephaly in 86%. Surgical complications were present in 39% of the neonates. During follow-up, 98% presented neurogenic bladder and 60% neurological/mental handicap. Twenty eight fetuses (42%) could have indication of in utero repair. CONCLUSION: MM is associated with severe and frequent poor results. Almost one third of our cases could had fetal repair as a treatment choice. PMID- 15273850 TI - [Spontaneous cervical carotid and vertebral arteries dissection: study of 48 patients]. AB - OBJECTIVE: To report a Brazilian series of spontaneous cervical arterial dissections, risk factors, warning symptoms, clinical manifestations, diagnostic tests, treatment and prognosis. METHOD: We performed the retrospective analysis of clinical and neuroradiological records (MRI, A-MRI and Angiography) of patients with this diagnosis who were evaluated in a tertiary hospital for the period of 1997-2003. RESULTS: 48 patients (24 men) with median age 37.9 years: 26 patients with unilateral internal carotid dissection (ICAD), 15 with unilateral vertebral artery dissection (VAD) and 7 with multivessel dissections. All patients presented neurological deficits. Hypertension, smoking and dyslipidemia were the main risk factors. More than 80% of patients presented at least one initial symptom, most of them temporoparietal headache. 44% of patients with VAD and only 3.4% of patients with ICAD had neck pain. The median interval between the onset of symptom and the appearance of neurological deficit was 5.4 days for ICAD and 13.5 days for VAD. Five patients with ICAD presented preceding TIA. Angiography was performed in 93% of patients. In 42% of these patients, MRI and A MRI were associated. In three patients the diagnosis was made just through cervical MRI. 75% of patients received anticoagulation. Two patients received intravenous thrombolytic therapy with no complications. Prognosis was good for all patients but two patients with bilateral ICAD died. CONCLUSION: Our results are similar to the literature, except for the low frequency of neck pain in ICAD patients and predominance of temporoparietal headache in cervical artery dissection patients. Vascular risk factors were commonly found. PMID- 15273851 TI - [Dysphagia in patients undergoing anterior cervical surgery]. AB - Dysphagia is one of the complications of anterior cervical surgery. Although common, few articles were published on this subject. Its incidence and duration varies depending on the author. We show a prospective study, analyzing the incidence, duration and quality of the dysphagia after anterior cervical surgery. PMID- 15273852 TI - [Incidence of oropharyngeal dysphagia associated with stroke in a regional hospital in Sao Paulo State - Brazil]. AB - Cerebrovascular disease is recognized as to be associated with the highest mortality rate in Brazil. Dysphagia, speech and language disturbances are common consequences of the high incidence of stroke. Dysphagia is known to occur in at least 50% of the patients with acute stroke. The study is designed to establish the incidence of stroke in a reference hospital. One hundred and two consecutive patients admitted between January 2001 and January 2002 underwent a neurological examination with dysphagia specific analysis, performed by speech/dysphagia professionals as soon as they have clinical conditions. The localization of the lesions are determined by computer tomography or magnetic resonance image of the brain. Sixty one patients underwent videofluorscopic evaluation of swallowing. There was detected oropharyngeal dysphagia in 78 patients (76.5%) if the examination was limited to the clinical evaluation. Nevertheless, if complemented by videofluorscopic evaluation, the incidence grows to 90%. The explanation for the high incidence observed in this study could be pointed to the fact that dysphagia was registered on different times of the convalescence period. The data reinforces the importance of repeated evaluations made by different professionals of the staff involved in acute stroke attendance. PMID- 15273853 TI - [Supratentorial low grade glioma in adults: an experience with 23 surgical cases]. AB - Low-grade supratentorial astrocytomas and oligodendrogliomas in adults are uncommon tumors of the central nervous system. We analyzed retrospectively 23 patients with this type of neoplasia, who were operated on between 1986 and 2002. There were no post-operative deaths. The survival rate at 5 and 10 years post surgery for the entire sample was 67 and 30% respectively, similar to other outcomes. With 14 patients we achieved a complete removal of the lesion (60.8%) and with 9 (39.2%) partial removal of the tumor. In the sub-group which underwent total resection of the tumor, 89% survived 5 years and 45% attained 10 years of survival, in contrast with the sub-group that underwent partial removal in which only 35% attained 5 years of survival and none 10 years. Due to the deleterious effects of radiation therapy, we preferred to prescribe it only in cases of tumor recurrence. PMID- 15273854 TI - [Acute treatment of migraine in emergency room: comparative study between dexametasone and haloperidol. Preliminary results]. AB - We studied the efficacy of dexamethasone (4 mg) and haloperidol (5 mg) in the treatment of migraine in the emergency room. Twenty nine patients who had diagnosis of migraine according to the International Headache Society criteria and were evaluated for a painful episode at the emergency room of Santa Casa of Sao Paulo were included. All the patients scored their pain in 10 when evaluated, even after the use of intravenous analgesia (dipyrone). Fourteen patients were treated with haloperidol and the remaining 15 received dexamethasone. The patients were asked about pain intensity at 30, 60, 90 and 120 minutes after the use of either the drugs. Both drugs were equally efficient in pain relief after two hours. Patients who were treated with haloperidol showed an important improvement (more than 50% of improve in the analogic pain scale) in the first 30 minutes. The dexamethasone treated patients only reached this grade of analgesia after 120 minutes. Although we studied a small series of patients, our data suggest that both drugs are efficient in the treatment of a refractory migraine attack. Haloperidol seemed to work quickly in pain relief. No important side effects were observed in neither groups. PMID- 15273855 TI - Cerebellar hemorrhage as a complication of temporal lobectomy for refractory medial temporal epilepsy: report of three cases. AB - Cerebellar hemorrhage is listed among the potential complications following neurosurgical procedures. In this scenario it is usually reported as a rare condition. However, it seems that epilepsy surgery patients are somewhat more prone to this kind of complication, compared to other surgical groups. Head positioning, excessive cerebral spinal fluid draining and the excision of non expanding encephalic tissue (or combinations among the three) are likely to be cause underlying remote cerebellar hemorrhage. Out of the 118 ATL/AH performed at our institution, between 1996 and 2002, we identified 3 (2.5%) patients presenting with cerebellar hemorrhage. We report on such cases and review the literature on the topic. PMID- 15273856 TI - Acquired Ondine's curse: case report. AB - We report and discuss the case of a 55-year old man who presented a history of stroke as well as chronic obstructive pulmonary disease. When admitted into the emergency room, he was diagnosed with a vertebro-basilar syndrome. A brain MRI showed a hyperintense area in the lower right brainstem laterally within the medulla, which corresponds to the area of the pathways descending from the autonomic breathing control center. During hospitalization, the patient had several episodes of prolonged apnea, mainly when asleep, having often to be "reminded" to breath. A tracheostomy was then performed with the patient under mechanical ventilation. Treatment with medroxyprogesterone, fluoxetine and acetazolamide was also started. He was discharged after 64 days breathing environmental air with no apparent episodes of apnea. He returned to the emergency room in the following day with a clinical picture of aspiration bronchopneumonia, followed by septic shock and death. CONCLUSION: the Ondine's curse is one of the posterior stroke's presentation characterized by loss of automatic breathing and for the unpredictability of clinical evolution and prognosis. Such a syndrome has rarely been reported in adults and the diagnostic criteria are not consensual in the reviewed literature. Thus any diagnostic confirmation should be flexible. There are many therapeutic symptomatic options in such cases, ranging from pharmacologic approach, use of bilevel positive airway pressure and implantation of diaphragmatic pacemaker. PMID- 15273857 TI - Ventricular arteriovenous malformation bleeding: a rare cause of headache in children. Case report. AB - Headache as a chief complaint is rare in the paediatric emergency room. Actually, very seldom cases secondary to life threatening conditions as non-traumatic subarachnoid haemorrhage have been reported. A child with severe headache and nuchal rigidity and no other abnormalities on the physical examination is reported. Magnetic resonance angiography and cerebral angiography disclosed a ventricular arteriovenous malformation in the choroid plexus, supplied by the anterior choroidal artery, classified according to Spetzler grading system as grade 3 (deep venous drainage: 1; eloquence area: 0 and size: 2). The differences in the clinical presentations of the central nervous system arteriovenous malformation between children and adults are discussed. PMID- 15273858 TI - Cavernous angioma of the cauda equina: case report. AB - We present a rare case of cavernous angioma of the cauda equina and review the eleven cases available in the literature. A 44-year-old woman presented with low back pain and sciatica associated with bowel and bladder dysfunction and motor weakness of the lower extremity. The MRI revealed an enhancing, heterogeneous and hyperintense intradural lesion compressing the cauda equina roots at the L4 level. Laminectomy at L3-L4 and total removal of the tumor were performed without additional neurological deficit. Pathology revealed a cavernous angioma. The literature, clinical presentation, technical examinations, and treatment are reviewed. PMID- 15273859 TI - Isolated superficial peroneal nerve lesion in pure neural leprosy: case report. AB - Patients with leprosy may have only nerve involvement without skin changes. These cases are known as pure neural leprosy and can be seen in 10% of leprosy patients. Most patients have mononeuritic or multiple mononeuritic neuropathy patterns. The isolated lesion of the superficial peroneal nerve is uncommonly seen. We report a patient with involvement of this nerve in which there was no thickening of superficial nerves. The performed nerve biopsy showed inflammatory infiltration, loss of fibers and presence of Mycobacterium leprae. We believe that in prevalent leprosy countries we should take in mind the possibility of isolated pure neural leprosy in some patients without skin lesion. In these cases the diagnosis of leprosy is impossible on clinical grounds and nerve biopsy is mandatory. PMID- 15273860 TI - Simvastatin-induced mononeuropathy multiplex: case report. AB - The association between the use of statins and neuromuscular disease is currently being intensely discussed. We relate a 63 years old man with possible case of statin-induced neuropathy in a patient with dislipidemia in use of simvastatina at high doses. The electrophysiologic studies disclosed findings compatible with mononeuropathy multiplex, suggested by clinical prescutation of asymmetrical numbness and weakness. More common causes of mononeuropathy multiplex were excluded and the patient improved after the discontinuation of the drug. PMID- 15273861 TI - Recurrent neuromyelitis optica with diffuse central nervous system involvement: case report. AB - Several demyelinating disorders can affect children. The differential diagnosis between these diseases is usually an arduous task. Diagnostic criteria have been proposed for some of these disorders, however most of them have not yet been clinically and prospectively validated. Here we present a case of a ten year-old boy with recurrent bilateral optic neuritis and spinal cord involvement. Clinical and cerebrospinal fluid data have fulfilled diagnostic criteria for Devic's neuromyelitis optica (NMO). The differential diagnosis with multiple sclerosis (MS) has become troublesome since not only optic nerves and spinal cord were involved. In one of the relapses a left hemiparesis with facial involvement was registered. Magnetic resonance imaging was also compatible with MS. This case illustrates that CNS demyelinating disorders can fulfill diagnostic criteria for more than one demyelinating disease, making the clinical judgment an important tool in the management of these patients. PMID- 15273862 TI - Progressive encephalomyelitis with rigidity: a paraneoplastic presentation of oat cell carcinoma of the lung. Case report. AB - Progressive encephalomyelitis with rigidity and myoclonus (PEWR) is a rare neurological disorder, characterised by muscular rigidity, painful spasms, myoclonus, and evidence of brain stem and spinal cord involvement. A 73-year-old white man was admitted with a 10-day history of painful muscle spasms and continuous muscle rigidity on his left lower limb. He had involuntary spasms on his legs and developed encephalopathy with cranial nerves signs and long tract spinal cord symptomatology. Brain CT scan and spinal MRI were normal. The CSF showed lymphocytic pleocytosis and no other abnormalities. EMG showed involuntary muscle activity with 2-6 seconds of duration, interval of 30-50 ms and a frequency of 2/second in the left lower limb. Anti-GAD antibodies were detected in the blood. We detected radiological signs of lung cancer during the follow-up, which proved to be an oat cell carcinoma. The patient died two weeks after the diagnosis of the cancer. PMID- 15273863 TI - [Progressive multifocal leukoencephalopathy confirmed by PCR for JC virus in cerebrospinal fluid: case report]. AB - A case of progressive multifocal leukoencephalopathy (PML) is presented, with literature review. PML diagnosis and its differential diagnosis are presented, with emphasis on neuroradiology, cerebrospinal fluid analysis and polymerase chain reaction studies. The prognosis of PML is usually poor, with a median survival of 1-6 months. There is yet no proven effective treatment for this condition; HAART has become the standard of care for these patients. PMID- 15273864 TI - [Isolated central nervous system angiitis and myelodysplastic syndrome: case report]. AB - Isolated central nervous system (CNS) angiitis are vasculitides of undetermined etiology in which only nervous system vessels are affected. In most cases there is no associated systemic disease. We report the case of a 67 years old man with previous hematologic diagnosis of myelodysplastic syndrome who developed an isolated CNS angiitis. PMID- 15273865 TI - [Prince Liev Nikolaievitch Michkin ("The Idiot", Fiodor Dostoevsky) and the interictal personality syndrome of temporal lobe epilepsy]. AB - Russian romancist Fiodor Dostoevsky's composition, besides its extraordinary literary value, has a special importance for neurologists and epileptologists. The writer, who suffered of epilepsy, transmitted in his texts the epileptic's universe and how the patient is perceived by the society. His novels had great influence on how epilepsy is perceived by western culture. The romance "The Idiot" has as protagonist Prince Liev Nikolaievitch Michkin, a epileptic with remarkable personality. Considering the propose made by Geschwind-Waxman (1975) of a interictal personality syndrome in temporal lobe epilepsy, this article intends to discuss the behavioral alterations in epileptic patient, from Michkin, the main character in Fiodor Dostoevsky's "The Idiot". PMID- 15273867 TI - Epidermal growth factor receptor pathway substrate 8 (Eps8) expression in maturing testis. AB - AIM: Although epidermal growth factor receptors are expressed in the testes, whether they signal through epidermal growth factor receptor pathway substrate 8 (Eps8) is unknown. Here we evaluated the expression pattern of Eps8 in the maturing testis. METHODS: The expression of Eps8 was analysed by Northern blotting, immunocytochemistry and Western blotting in primary Sertoli cell cultures and in testicular tissue of rodents. RESULTS: Eps8 is specifically expressed in gonocytes, Leydig and Sertoli cells of the neonatal rats and in Leydig and Sertoli cells of the adult rats and mice. Although gonocytes express Eps8, no signal was found in prepubertal or mature spermatogonia and the expression level of Eps8 in Sertoli cells increases with age. No regulation of Eps8 expression in primary immature rat Sertoli cells by Follicle stimulating hormone (FSH) was detected by Western blotting. CONCLUSION: Eps8 seems to be involved in the growth factor-controlled regulation of cell proliferation and differentiation in the seminiferous epithelium. Eps8 is a possible marker for gonocytes and in Sertoli cells it could be involved in crosstalk with other growth factor pathways. PMID- 15273868 TI - Potentiation of apomorphine effect on sildenafil-induced penile erection in conscious rabbits. AB - AIM: To investigate a possible potentiation effect of apomorphine (APO) on sildenafil-induced penile erection in the conscious rabbit. METHODS: Erection of male New Zealand White rabbits (3.5 - 4.0 kg, n=12) was assessed by measuring the length of the uncovered penile mucosa and the duration of erection before and after intravenous administration of agents. After injection of APO (0, 0.05, 0.1 and 0.4 mg/kg), sildenafil was administered intravenously in a dose-response manner (0.5, 1 and 5 mg/kg). In additional experiments, the effect of increasing doses of sildenafil in combination with APO on systemic blood pressure was evaluated. RESULTS: Systemic administration of sildenafil induced a dose dependent increase in the penile length. Intravenous injection of APO alone did not produce any change in the penile length, while significantly enhanced the penile erection induced by sildenafil. The co-administration of 0.1 mg/kg of APO and 1 mg/kg of sildenafil was found to be the most effective combination in producing penile erection. Intravenous administration of sildenafil caused a concentration-dependent decrease in systemic blood pressure, but no additional decrease was observed with co-administration of APO. CONCLUSION: APO enhances the penile erection induced by sildenafil in the conscious rabbit without causing an additional decrease in blood pressure. PMID- 15273869 TI - Second to fourth digit ratio (2D:4D) and testosterone in men. AB - AIM: To investigate the relationship between 2D:4D and testosterone in men attending an infertility clinic and men drawn from the general population. METHODS: Data on 2D:4D and testosterone from two samples were collected: (1) 43 men attending an infertility clinic, and (2) 51 men drawn from the general population without regard to fertility. RESULTS: In sample (1) there were negative associations between 2D:4D and testicular function, and men with lower 2D:4D in their right compared to left hand had higher testosterone levels than men with higher 2D:4D in their right compared to left hand. Sample (2) showed no significant associations between 2D:4D or side differences in 2D:4D and testosterone. CONCLUSION: Adult levels of testosterone may be related to aspects of 2D:4D in samples which contain men with compromised testicular function, but not in men from normative samples. Associations between 2D:4D and fertility associated traits probably arise from early organisational effects of testosterone rather than from activational effects of current testosterone. PMID- 15273870 TI - Expression of germ cell nuclear factor in mouse germ cells and sperm during postnatal period. AB - AIM: To assess the spatial and temporal expression of germ cell nuclear factor (GCNF) in male mouse germ cells during postnatal development and in sperm before and after capacitation. METHODS: The indirect immunofluorescence method with anti GCNF antiserum was used to investigate the GCNF expression in mice at day 8, 10, 14, 17, 20, 28, 35, 70, and 420 after birth and in sperm before and after capacitation. RESULTS: With the proceeding of spermatogenesis, GCNF was first detected in the nuclei of spermatogonia and a few early stage primary spermatocytes at day 8, which was increased gradually at day 10 to 14 inclusive. From day 17 to day 20, the GCNF was concentrated in round spermatids, while both spermatogonia and early stage primary spermatocytes became GCNF negative. From day 28 until day 420, strong GCNF expression was shown in round spermatids and pachytene spermatocytes, while spermatogonia, early primary spermatocytes and elongating spermatids were all GCNF negative. In addition, it was also found that GCNF was localized on the acrosomal cap region of spermatozoa and there was a big change in GCNF expression during capacitation, from 98 % GCNF positive before capacitation to about 20 % positive following capacitation. The localization of GCNF in caput and cauda spermatozoa was similar. CONCLUSION: GCNF may play important roles in spermatogenesis, capacitation and fertilization. PMID- 15273871 TI - Effect of tamoxifen on spermatogenesis and tubular morphology in rats. AB - AIM: To observe the effect of tamoxifen citrate on spermatogenesis and tubular morphology in rats. METHODS: The effect of tamoxifen citrate i.g. at doses of 400 and 800 mg.kg(-1).day(-1) in 0.1 mL olive oil for 30 days on seminiferous tubular morphology, seminiferous epithelial diameter (STD), epithelial height (SEH), epididymal sperm count and percent abnormal sperm were evaluated at day 1, 12 and 36 after treatment. Controls were given the vehicle. RESULTS: The higher dose resulted in tubular atrophy on day 31. The STD, SEH and sperm count were decreased and the abnormal spermatozoa increased in a dose-dependent manner with the maximal effect on day 36. CONCLUSION: Tamoxifen citrate induces tubular shrinkage and atrophy and sperm abnormality at a dose-dependent manner. PMID- 15273872 TI - Influence of selenium induced oxidative stress on spermatogenesis and lactate dehydrogenase-X in mice testis. AB - AIM: To evaluate the effect of oxidative stress on the spermatogenesis and lactate dehydrogenase-X (LDH-X) activity in mouse testis. METHODS: For creating different levels of oxidative stress in mice, three selenium (Se) level diets were fed in separate groups for 8 weeks. Group 1 animals were fed yeast-based Se deficient (0.02 ppm) diet. Group 2 and Group 3 animals were fed with the same diet supplemented with 0.2 ppm and 1 ppm Se as sodium selenite, respectively. After 8 weeks, biochemical and histopathological observations of the testis were carried out. LDH-X levels in the testis were analyzed by western immunoblot and ELISA. RESULTS: A significant decrease in testis Se level was observed in Group 1 animals, whereas it was enhanced in Group 3 as compared to Group 2. The glutathione peroxidase (GSH-Px) activity was significantly reduced in both the liver and testis in Group 1, but not in Group 2 and 3. A significant increase in the testis glutathione-S-transferase (GST) activity was observed in Group 1, whereas no significant change was seen in Groups 2 and 3. Histological analysis of testis revealed a normal structure in Group 2. A significant decrease in the germ cell population in Group 1 was observed as compared to Group 2 with the spermatids and mature sperm affected the most. Decrease in the lumen size was also observed. In the Se-excess group (Group 3), displacement of germ cell population was observed. Further, a decrease in the LDH-X level in testis was observed in Group 1. CONCLUSION: Excessive oxidative stress in the Se deficient group, as indicated by changes in the GSH-Px/GST activity, affects the spermatogenic process with a reduction in mature sperm and in turn the LDH-X level. PMID- 15273873 TI - Effect of sildenafil citrate on penile erection of rhesus macaques. AB - AIM: To examine the effect of sildenafil citrate on penile erection of male rhesus macaque. METHODS: Twenty Macaca mulatta were divided into the sildenafil treated and the control groups of 10 animals each. The penile size, the corpus cavernosal electromyogram (EMG) and the intra-corpus cavernosal pressure (ICP) were determined. RESULTS: The diameter of penis and the ICP were significantly increased and the corpus cavernosal EMG significantly reduced in the sildenafil group. CONCLUSION: Sildenafil citrate increases the penile size and ICP and reduces the corpus cavernosal EMG in male rhesus macaque. PMID- 15273874 TI - Effect of lead chloride on spermatogenesis and sperm parameters in mice. AB - AIM: To evaluate the effect of acute lead chloride exposure on testis and sperm parameters in mice. METHODS: PbCl2, 74 mg/kg, was daily administered to sexually mature male mice for 3 days and the effects on the testicular histology and ultrastructure as well as the motility and density of spermatozoa in cauda epididymis were observed. An additional group of mice were treated for 1-3 days and were allowed to recover for 32 days to determine the reversibility of lead induced changes. RESULTS: The testicular weight, seminiferous tubular diameter and sperm counts were significantly decreased following 3 days of PbCl2 treatment, but were unaffected by shorter-term exposures. The changes caused by lead are mostly reversible. CONCLUSION: Acute lead chloride exposure injures the fertility parameters of male mice and the effects are partially reversible. PMID- 15273875 TI - Seasonality in human semen quality of smokers and non-smokers: effect of temperature. AB - AIM: To analyse the possible effect of seasonal variation on semen parameters. METHODS: The participants consisted of 1,688 men attending the andrology laboratory between 1991 and 1997 for reduced fertility in the couple. Semen analysis was performed according to the WHO manual. The 84 individual months of the study period were each assigned to one of the three groups according to the average monthly outside temperature; Group A (temperature < 4.4 degrees C), Group B (4.4 degrees C - 13.3 degrees C) and Group C (>13.3 degrees C). RESULTS: When comparing the different sperm parameters, the morphology was significantly better in Group C. However, when the smokers were analysed separately, this difference disappeared and significant seasonal variations were found in sperm density, total sperm count, motility and total motile sperm; they were deteriorated in the warmer season. In non-smokers, no such negative effect of increased temperature was observed. CONCLUSION: Sperm quality is influenced by seasonal factors. Increased environmental temperature, (maybe also light exposure) has an additional negative effect on the spermatogenesis in smokers, leading to reduced sperm quality in men with borderline fertility. PMID- 15273876 TI - Antibiotic-coated medical devices: with an emphasis on inflatable penile prosthesis. AB - One of the most serious complications associated with the use of the inflatable penile prosthesis is infection. This can lead to significant morbidity for the patient, as well as significant health care costs. A number of methods have been used in attempts at minimizing the infection risk, including applying an antibiotic coating to the medical devise. This review aims to evaluate the effectiveness of these products in preventing clinically significant infections. PMID- 15273877 TI - Regulation of spermatogenesis by paracrine/autocrine testicular factors. AB - Spermatogenesis is a complex process regulated by endocrine and testicular paracrine/autocrine factors. Gonadotropins are involved in the regulation of several testicular paracrine factors, mainly of the IL-1 family and testicular hormones. Testicular cytokines and growth factors (such as IL-1, IL-6, TNF, IFN gamma, LIF and SCF) were shown to affect both the germ cell proliferation and the Leydig and Sertoli cells functions and secretion. Cytokines and growth factors are produced by immune cells and in the interstitial and seminiferous tubular compartments by various testicular cells, including Sertoli, Leydig, peritubular cells, spermatogonia, differentiated spermatogonia and even spermatozoa. Corresponding cytokine and growth factor receptors were demonstrated on some of the testicular cells. These cytokines also control the secretion of the gonadotropins and testosterone in the testis. Under pathological conditions the levels of pro-inflammatory cytokines are increased and negatively affected spermatogenesis. Thus, the expression levels and the mechanisms involved in the regulation of testicular paracrine/autocrine factors should be considered in future therapeutic strategies for male infertility. PMID- 15273878 TI - Androgenic effect of Mondia whitei roots in male rats. AB - AIM: To determine the effect of the aqueous extract of Mondia whitei (Periplocaceae) roots on testosterone production and fertility of male rats. METHODS: Adult male Wistar rats were used. In the acute study, 20 rats were randomly divided into 5 groups of 4 animals each. Four treated groups were administered orally a single dose of Mondia whitei (400 mg/kg) and the controls received a similar amount of distilled water. One group of animals were sacrificed by cervical dislocation 1, 2, 4 and 6 h after treatment, respectively. The controls were sacrificed at 6 h. Testicular testosterone was determined by radioimmunoassay. In the chronic study, 28 rats were divided at random into 4 groups of 7 animals each: Groups 1, 2 and 3 were given orally the plant extract (400 mg.kg(-1).day(-1)) for 2, 4 and 8 days, respectively. The animals of Groups 1 and 2 were sacrificed 24 hours after the last dosing. The controls (Group 4) received the same amount of distilled water for 8 days. The fertility was assessed only in Groups 3 and 4 and after that, the animals were sacrificed and the epididymal sperm density, the serum testosterone and the testicular testosterone and 17 beta-estradiol were assayed. The serum, testicular and epidydimal protein contents were also determined. RESULTS: In the acute treatment groups, the serum and testicular concentrations of testosterone remained unchanged at all the time points. Chronic treatment for 8 days induced a significant increase in the testicular weight, the serum and testicular testosterone, the testicular protein content and the sperm density (P < 0.05 0.01), but did not affect the accessory gland weights, the serum protein contents, the testicular concentration of 17beta -estradiol and the fertility compared to the controls. CONCLUSION: Mondia whitei root extract possesses an androgenic property. PMID- 15273879 TI - Congenital penile curvature: long-term results of operative treatment using the plication procedure. AB - AIM: To determine the long-term outcome, effectiveness and patient satisfaction of congenital penile curvature correction by plication of tunica albuginea. METHODS: From January 1992 to January 2002, 106 young patients underwent surgical correction of congenital penile curvature by corporeal plication. Indications for operation were difficult or impossible vaginal penetration and cosmetic problems. The technique of corporeal plication consists of placing longitudinal plication sutures of 2-zero braided polyester on the convex side of the curvature until the curvature is corrected when erection is artificially induced. Results of this procedure were obtained by retrospective chart reviews and questionnaires via mail. Long-term follow-up ranged from 11 to 132 (mean 69.3) months and data were available for 68 patients. RESULTS: Penile straightening was excellent in 62 patients (91 %) and good with less than 15 degree of residual curvature in 6 patients (9 %). Sixty-seven patients reported no change in erectile rigidity or maintenance postoperatively, while 1 described early detumescence. Shortening of the penis without functional problems was noted by 26 patients (38 %). Thirty Five patients (51 %) reported feeling palpable indurations (suture knots) on the penis. Temporary numbness of glans penis was described in 3 patients. Overall, 60 patients were very satisfied, 6 satisfied, 2 unsatisfied. CONCLUSION: Corporeal plication is an effective and durable procedure with a high rate of patient satisfaction. PMID- 15273880 TI - [International Headache Society classification of headaches. Lights and shadows]. PMID- 15273881 TI - [Spanish version of the 7 Minute screening neurocognitive battery. Normative data of an elderly population sample over 70]. AB - INTRODUCTION: To standardize the Spanish version of the 7 Minute screening neurocognitive battery (7MS) in a population sample of elderly over 70 years. METHODS: We examined 416 persons, living at home, participating in elderly the longitudinal study "Aging in Leganes", aged 71 to 99 years old (mean age: 79 +- 9.2 years; 51.7 % women; 10.6 illiterate, 25 % without formal education). In order to do so, we used an extensive clinical survey, general and neurological exam and extensive neuropsychological battery with several cognitive scales, attention, language, memory, visuomotor skill and reasoning tests, Jorm's IQCODE questionnaire, CES-D depression questionnaire and the 7MS including the Benton Orientation Test, Clock Drawing Test, Free and Cued Learning Test and Categorial Verbal Fluency. Dementia was diagnosed according to DSM-IV criteria but independently of the 7MS scores. Several methods to obtain the total score of the 7MS were analyzed and the normative parameters of the test were obtained in the subgroup of non-demented subjects. RESULTS: The easiest and most efficient method to obtain the total score of the 7MS was the sum of the z-scores of the four subtests. We present the mean values, -1 and -1.5 standard deviations, range and percentiles of the partial and total scores of the 7MS stratified by age (71-75, 76-80, 81-85 and > or = 86 years) and education (less than primary education and primary education or greater) in the subgroup of non-demented subjects. CONCLUSIONS: The normative data of the 7MS obtained in a representative sample of the general elderly population support its rigorous use in the Spanish clinical setting. PMID- 15273882 TI - [Modifications of the lipid metabolism induced by interferon beta in multiple sclerosis patients and its relationship with the disease activity]. AB - INTRODUCTION: It has been recently suggested that total cholesterol and low density lipoproteins (LDL) levels can behave as biological markers of activity in demyelinating diseases. Thus, our aim has been to describe the modifications of the plasma levels of total cholesterol and triglyceride due to treatment with interferon-beta in multiple sclerosis (MS) patients and to determine their relationship with the disease activity. PATIENTS AND METHODS: Study of the follow up of MS patients under treatment with interferon-beta. Clinical and analytical controls were performed before initiating treatment and than at 1, 3, 6, 12, 18 and 24 months of its initiation. RESULTS: Fifty six patients have been studied, 41 of them women. Mean age was 37.4 years. Fifty were relapsing- remitting forms and the rest secondary progressive forms. The mean plasma levels of triglyceride increased and total cholesterol levels diminished during the 24 months of treatment with interferon, mainly in the first 3 months. No statistically significant relationship was found between disease activity and mean plasma levels of triglyceride and total cholesterol before the beginning of the treatment and during the period of follow-up. CONCLUSIONS: Treatment with interferon-beta in the MS patients originates changes in the plasma lipid profile, but neither these changes nor the plasma lipid levels before the treatment behave as biological markers of disease activity. PMID- 15273883 TI - [Predicting factors for depression in multiple sclerosis]. AB - INTRODUCTION: We assess frequency and intensity of depression in multiple sclerosis (MS) patients, the degree at which it is detected and its relationship to the treatment with beta interferon and other clinical and paraclinical factors. METHODS: The series comprises MS patients, seen in the Demyelinating Disease Unit of a tertiary hospital, who fulfilled the following criteria: clinically defined MS (relapsing-remitting or secondary progressive), disease duration greater than two years and absence of relapses during the month prior to the study. The variables analyzed were detection and assessment of depression with the Hamilton Depression Scale, general demographic data, functional systems, EDSS, ISS, ESS, Pittsburgh Sleep Quality Index, interferon treatment, chronic fatigue and a series of analytical variables. Statistical study: both variate and multivariate analysis by logistic regression. RESULTS: 100 patients (72 female and 28 male). Mean age: 39.27 years. RR MS form, 88%, and SPMS form, 12 %. Mean evolution time, 11.2 years. Mean EDSS, 2.54. Depression was present in 44 % of the patients in our group and was not related to neurological degree of disability, disease evolution time, clinical form, interferon treatment, or to sleep disorders. However, depression was related to the presence of both chronic fatigue and ESS scores. CONCLUSIONS: Depression is common in MS patients and is associated with the presence of chronic fatigue and a worse social status. PMID- 15273884 TI - [Efficacy and safety of endovascular treatment of brain aneurysms: experience in our setting]. AB - INTRODUCTION: The objective of our study is to know the efficacy and safety of endovascular treatment in our setting. PATIENTS AND METHODS: a) prospective evaluation of patients with aneurysmal subarachnoid hemorrhage (SAH) or incidental aneurysm (Dec 1999-Dec 2002), and b) neurological, angiographic and evolutive evaluation of the endovascularly operated patients. RESULTS: A total of 79 patients were evaluated: 75 with SAH, 4 with incidental aneurysm. Arteriographies were performed in 74 (93.6%), detecting 58 aneurysms in 52 patients, 15 of the anterior communicating artery (25%) and 14 of the posterior communicating artery (24%). Fifty-two aneurysms were treated with Guglielmi Detachable Coiling (GDC) (89.6 %). Total occlusion (100 %) was obtained in 48 (82.7%) and partial occlusion in 4. Technique failure occurred in six cases. The earlier complications, after endovascular procedure (< 24 h), were one rebleeding, four angiographic spasms and ischemia and one coil displacement. During the first month, there were other cases of rebleeding and two cases of spasm and ischemia. Morbidity rate was 11.5 %. Death rate was 5.7 %. Follow-up period ranges between 6 and 42 months. After endovascular treatment 40 patients had a favourable outcome (Ranking 1-2) and 9 suffered moderate-severe incapacity (Ranking 3-5). We prescribed angiographic controls one year after the hemorrhagic event and there was only one case of aneurysmal recurrence. DISCUSSION: In our experience endovascular treatment is useful to treat ruptured and unruptured aneurysms, with similar surgical occlusion, morbidity and death rates. PMID- 15273885 TI - [Psychogenic disorders: concepts, terminology and classification]. AB - The neurological interest on functional or psychogenic disorders (mental or physical disturbances with no organic basis, generally unleashed by stressful situations) has been receiving increasingly more interest over the last few years. In this article we review concepts, terms and classifications of these disorders, very different over time and among different authors. Psychogenic disorders are divided into: a) dissociation (with memory, consciousness and self identity impairment), and b) disturbances with somatizations, divided into somatoform (unconscious), factitious (voluntary search for patient's role) and malingering (searching for material gain). Special emphasis is placed on conversion or hysteria, included in somatoform disorders. New findings in functional neuroimaging are analyzed. These new data suggest an important role of unconscious and involuntary inhibition in loss of volition (similar to hypnosis and different from malingering). Normal activity in certain brain areas (motor or sensory cortex) is blocked by other areas related to emotional integration (anterior cingular and orbitofrontal cortex). The neurologist's role is important to achieve an early diagnosis of psychogenic disturbances, particularly conversive ones. This means the use of fewer economic resources and better prognosis for the patient. PMID- 15273886 TI - [A legendary neurological disease favored the refoundation of the diocese of Palencia]. AB - INTRODUCTION: At the beginning of the XI century, the diocese of Palencia, Spain, was refounded and politically empowered following the indications of Sancho III, King of Pamplona. Beginning in the XIII century orally and then after the XV century in several written documents, there is a legend that explains this historical event by the miraculous cure of the King for an apparent neurological disease. CLINICAL DESCRIPTION: One day, when he was about 42 years old, King Sancho III was hunting near the river Carrion. Following the trail of a wild boar, he entered a ruined crypt in which the mortal remains of the martyr Saint Antolin laid. Just when he was going to kill the animal, the King abruptly suffered weakness in his right arm, which made him drop his lance. On his knees and aware of his disease, and praying sorrowfully, the King recovered the mobility of his arm and left the crypt cured. RESULTS: We analyzed the written documents and artistic representations that recall this legendary episode. We reviewed the political reasons, apart from mere religious events, that might have persuaded Sancho III to empower the land of Palencia. CONCLUSIONS: We have had the opportunity of reviewing a beautiful legend of our medieval Spain, which could correspond to one of the first ever-existing descriptions of a cardioembolic transient ischemic attack. PMID- 15273887 TI - [Image of the month. FSH producing macroadenoma: a disease that is not as benign as the histology indicates]. PMID- 15273888 TI - [Trimetazidine-induced parkinsonism]. AB - Eight patients, six women and two men, ages between 72 and 94 years (mean age 80), suffered from parkinsonism while taking trimetazidine, seven of them for a 6 to 12 months time period and the other for 9 years. The parkinsonism reverted after drug withdrawal. This is the first report of trimetazidine induced parkinsonism. It is important to know this side effect in order to avoid its administration to patients with any parkinsonism, especially those suffering dementia with Lewy bodies. PMID- 15273889 TI - [Paraparesis, hyperprolactinemia and adynamic ileus in Guillain-Barre syndrome]. AB - A case of Guillain-Barre with unusual autonomic dysfunction at its onset, that consisted of constipation and hypertension, followed by adynamic ileus and flaccid paraparesis with areflexia limited to the lower limbs, is presented. Inappropriate secretion of antidiuretic hormone and hyperprolactinemia were demonstrated, which resolved spontaneously afterwards. The adynamic ileus resolved and the paraparesis improved with gastric aspiration and intravenous immunoglobulin administration, only to worsen eighty days later. Paraparesis worsened, accompanied by hypertension, an abnormal hypotensive response to captopril and limb pain. The whole clinical picture resolved after a second course of intravenous immunoglobulin. Unusual clinical aspects of Guillain-Barre syndrome found in this case are reviewed, such as paraparesis, prolonged adynamic ileus and endocrine abnormalities, comparing them with available data from the neurological literature. Differential diagnosis with myelopathy, metabolic and paraneoplasic neuropathies, POEMS syndrome, chronic inflammatory demyelinating polyneuropathy and recurrent Guillain-Barre syndrome are commented on. Hyperprolactinemia is proposed as a marker, either of adenopituitary denervation or of the autoimmune response, in the course of this disease. PMID- 15273890 TI - [Intravenous immunoglobulin therapy in Vogt-Koyanagi-Harada syndrome]. AB - Vogt-Koyanagi-Harada (VKH) syndrome or uveomeningitic syndrome is a disease affecting several organs: eye (bilateral uveitis, exudative retinal detachments), ear (tinnitus, dysacousia), skin and hair (vitiligo, alopecia, poliosis) and the nervous system (meningism, headache, pleocytosis in cerebrospinal fluid). The etiology remains unknown but it is probably a cell-mediated autoimmune disorder in individuals genetically susceptible to antigenic components of melanocytes. We report a 25 year old patient with VKH syndrome treated with intravenous steroid therapy and cycles of intravenous immunoglobulin with good clinical response. We concluded that treatment of the VKH syndrome should be early but definitely aggressive with high doses of systemic corticosteroids and intravenous immunoglobulin, assessing the maintenance of the latter by cycles. PMID- 15273891 TI - [Sudden bilateral deafness as first manifestation of meningeal carcinomatosis]. PMID- 15273892 TI - The relationship between energy expenditure and lean tissue in monozygotic twins discordant for spinal cord injury. AB - Energy expenditure and fat-free mass (FFM), as well as the relationships between these parameters, were investigated in thirteen pairs of monozygotic twins discordant for SCI. Basal energy expenditure (BEE) and resting energy expenditure (REE) were determined by indirect calorimetry. Measurements for FFM and fat mass were obtained by dual-energy x-ray absorptiometry. Total body potassium was determined by a 4-Pi whole-body counting chamber. Values are expressed as mean standard deviation. BEE and REE of the twins with SCI were significantly less than those of the able-bodied co-twins (1387 268 vs. 1660 324 kcal/d, p < 0.005, and 1682 388 vs. 1854 376 kcal/d, p < 0.05, respectively). Regardless of the group, direct and highly significant relationships were evident between BEE or REE and FFM or TBK. In summary, twins with SCI had lower energy expenditure than their able-bodied co-twins. Regardless of paralysis, direct linear relationships existed between energy expenditure and measures of lean mass. PMID- 15273893 TI - Effect of variable loading in the determination of upper-limb anaerobic power in persons with tetraplegia. AB - This article examines the effects of levels of resistance loading during arm Wingate Anaerobic Testing (WAnT) in persons with differing levels of cervical spinal cord injury (SCI). Thirty-nine persons with motor-complete SCI tetraplegia (13 each at C5, C6, and C7) performed six bouts of arm-crank WAnT with relative loads equivalent to 1.0, 1.5, 2.0, 2.5, 3.0, and 3.5 percent of body mass (BM). Power output was determined with the use of the SMI OptoSensor 2000 (Sports Medicine Industries, Inc., St. Cloud, MN, USA) hardware and software package. Values of peak power (P(peak)) and mean power (P(mean)) were examined statistically between groups (C5, C6, and C7) and across levels of resistance loading. Resistance loads that provided the greatest values of P(mean) for the three groups were as follows: C5 = 1.0 or 1.5 percent of BM; C6 = 1.5 or 2.0 percent of BM; and C7 = 2.5, 3.0, or 3.5 percent of BM. Appropriate loading for arm WAnT is specific to the level of tetraplegia and may provide a useful assessment of upper limb power production. PMID- 15273894 TI - Effects of spinal cord injury on lower-limb passive joint moments revealed through a nonlinear viscoelastic model. AB - We developed a mathematical model to describe the lower-limb passive joint moments to investigate and compare these moments in a small sample of able-bodied volunteers and individuals with long-standing motor complete paraplegia. Isokinetic tests, which were performed on a sample of four subjects with spinal cord injuries (SCIs) and five uninjured individuals, measured the passive moments at the ankle, knee, and hip joints throughout their ranges of motion in the sagittal and coronal planes. We fitted an 11-parameter nonlinear viscoelastic model to the acquired passive moment data (mean square error ranging from 0.020 to 5.1 Nm2) to compare subject populations and to determine the influences of joint velocity and passive coupling between adjacent joints. Although the passive moment curves of the SCI and able-bodied groups exhibited many similarities in shape, a repeated measures analysis of variance (ANOVA) that compared the passive moment curves of the two groups indicated a statistically significant (p < 0.01) difference for every joint except the knee. This new model for passive joint moments should prove to be useful in examining how changes in passive properties affect bipedal function and movement. PMID- 15273895 TI - Variable-frequency-train stimulation of skeletal muscle after spinal cord injury. AB - Skeletal muscle, after spinal cord injury (SCI), becomes highly susceptible to fatigue. Variable-frequency trains (VFTs) enhance force in fatigued human skeletal muscle of able-bodied (AB) individuals. VFTs do this by taking advantage of the "catch-like" property of skeletal muscle. However, mechanisms responsible for fatigue in AB and SCI subjects may not be the same, and the efficacy of VFT stimulation after SCI is unknown. Accordingly, we tested the hypothesis that VFT stimulation would augment torque-time integral in SCI subjects. The quadriceps femoris muscle was stimulated with constant frequency trains (CFTs) (six 200 s square wave pulses separated by 70 ms) or VFTs (a train identical to the CFT, except that the first two pulses were separated by 5 ms) in SCI and AB subjects. After 180 contractions (50% duty cycle), isometric peak torque decreased 44, 56, and 67 percent, in the AB (n = 10), acute SCI (n = 10), and chronic SCI (n = 12) groups, respectively. In fatigued muscle, VFTs enhanced the torque-time integral by 18 percent in AB subjects and 6 percent in chronic SCI patients, and had no effect in acute SCI patients when compared to the corresponding CFT. The much faster rise times in SCI subjects (approximately 80 ms vs. 120 ms in AB subjects) probably contributed to the inability of VFTs to enhance torque-time integrals in SCI patients. The results suggest that the use of VFT stimulation in patients with SCI may not be as efficacious as it is in AB persons. PMID- 15273896 TI - A portable, 8-channel transcutaneous stimulator for paraplegic muscle training and mobility--a technical note. AB - This paper introduces an 8-channel transcutaneous neuromuscular stimulator, called ExoStim, which was designed and developed to provide stimulation to the lower-limb muscles of spinal cord injured individuals. The intended purposes of the ExoStim were to act as a skin-surface precursor to an implantable neuromuscular stimulator for the specific tasks of increasing paralyzed leg strength and endurance, enabling the performance of basic lower-limb functional tasks, and familiarizing patients with functional electrical stimulation training. The initial design specifications included portability (approximately 500 g), battery-powered output, constant current control (0-300 mA), 8 channels of biphasic stimulation (charge-balanced, constant current), and microprocessor control of all stimulation parameters. Various tests, including output power characteristics, environmental, mechanical, and battery life, were performed on three prototype units to validate our design specifications. Having successfully passed all tests, the ExoStim is now ready to be deployed to clinical trial sites for further evaluation with spinal cord injured subjects. PMID- 15273897 TI - Effects of neostigmine and glycopyrrolate on pulmonary resistance in spinal cord injury. AB - Preliminary findings in subjects with spinal cord injury (SCI) suggest that neostigmine administered intravenously increases colonic tone, increases colonic contractions, and facilitates bowel evacuation. Of concern are potential pulmonary side effects, including an increase in airway secretions and bronchospasm. The objectives of the study were to determine the effects of intravenously administered neostigmine or neostigmine combined with glycopyrrolate on forced oscillation indices in persons with SCI. Pulmonary resistances at 5 Hz (R5) and 20 Hz (R20) were measured with the use of an impulse oscillation system (IOS) in 11 subjects with SCI. Values were obtained before and after the intravenous administration of 2 mg of neostigmine alone and, on a separate day, before and after the administration of 2 mg of neostigmine combined with 0.4 mg of glycopyrrolate. Baseline R5 and R20 values before neostigmine correlated significantly with baseline values before neostigmine combined with glycopyrrolate. Following neostigmine, mean R5 values increased 25% and mean R20 values increased 18%. Following neostigmine combined with glycopyrrolate, mean R5 values fell 9% and mean R20 values fell 7%. In summary, baseline IOS values obtained on 2 different days were highly reproducible in this population. Neostigmine alone induced significant bronchoconstriction, whereas neostigmine combined with glycopyrrolate caused bronchodilation. PMID- 15273898 TI - Bronchodilator responses to metaproterenol sulfate among subjects with spinal cord injury. AB - A previous study using spirometric methods demonstrated that 42% of subjects with tetraplegia experienced significant bronchodilation following inhalation of metaproterenol sulfate (MS). Comparative studies involving subjects with paraplegia were not performed and none has been performed in this population using body plethysmography, a more sensitive method used to assess airway responsiveness. Stable subjects with tetraplegia (n = 5) or paraplegia (n = 5) underwent spirometry and determination of specific airway conductance (sGaw) by body plethysmography at baseline and 30 minutes after nebulization of MS (0.3 mL of a 5% solution). Among subjects with tetraplegia, inhaled MS resulted in significant increases in spirometric indices and sGaw. Among subjects with paraplegia, only sGaw increased significantly, although this increase was considerably less than that seen in subjects with tetraplegia. Our findings indicate that subjects with tetraplegia exhibit greater bronchodilation in response to inhaled MS than do subjects with paraplegia and that sGaw measurements may confer greater sensitivity for assessing bronchodilator responsiveness in tetraplegia. PMID- 15273899 TI - The effect of rear-wheel position on seating ergonomics and mobility efficiency in wheelchair users with spinal cord injuries: a pilot study. AB - This study analyzed the effect of rear-wheel position on seating comfort and mobility efficiency. Twelve randomly selected paraplegic wheelchair users participated in the study. Wheelchairs were tested in two rear-wheel positions while the users operated the wheelchair on a treadmill and while they worked on a computer. Propulsion efficiency, seating comfort, and propulsion qualities were registered at different loads during the treadmill session. During the computer session, pelvic position, estimated seating comfort, and estimated activity performance were measured. The change in rear-wheel position affected wheelchair ergonomics with respect to weight distribution (p < 0.0001) and seat inclination angle (position I = 5 and position II = 12). These changes had a significant effect on push frequency (p < 0.05) and stroke angle (p < 0.05) during wheelchair propulsion. We found no consistent effect on mechanical efficiency, estimated exertion, breathlessness, seating comfort, estimated propulsion qualities, pelvic position, or activity performance. PMID- 15273900 TI - Predicting consistency of pain over a 10-year period in persons with spinal cord injury. AB - This longitudinal study was designed to test the hypothesis that persons who consistently report pain at three (women) or four (men) measurement points across 10 years (1988 to 1998) are different both physically and psychologically from those who inconsistently or never report pain. Participants were 96 persons with spinal cord injury (SCI) living in the community who participated at every measurement point. Measures included consistency of reports of pain (i.e., reported having had problems with pain in the 12 months prior to all, some, or no measurement points); demographic and injury-related data; and measures of physical and psychological health, function, and social support. Of the 96 participants, approximately half of the men and three-fourths of the women consistently reported pain at each point. Phase 1 predictors of the consistency of pain reports for men were being less impaired, being more independent, experiencing more stress, and receiving less social support. Women consistently reporting pain had more stress at Phase 1 than women inconsistently reporting pain. Persons with SCI at risk for chronic pain should be identified and referred to a multidisciplinary pain management program. PMID- 15273901 TI - Common carotid and common femoral arterial dynamics during head-up tilt in persons with spinal cord injury. AB - We examined the effect of the level and completeness of spinal cord injury (SCI), tetraplegia and paraplegia, on common carotid arterial (CCA) and common femoral arterial (CFA) functions supine and during head-up tilt (HUT), compared with able bodied controls. Subjects (tetraplegia [n = 7], paraplegia [n = 8], and controls [n = 8]) were healthy males between the ages of 19 and 60 years. We used Doppler ultrasound to determine vessel diastolic diameters and flow velocities while supine and at 45 HUT. The results indicated that supine CCA diameter and flow were augmented in the tetraplegia group compared with the paraplegia group (p < 0.05); no other group differences were noted. However, CCA(flow) was significantly reduced from supine to 45 HUT in the tetraplegia group (p < 0.01). CFA diameter and flow were significantly reduced in the SCI groups compared with the control group, and CFA(flow) was reduced from supine to 45 HUT in the tetraplegia group. These results demonstrate that individuals with tetraplegia have increased resting CCA diameters and flows compared with individuals with paraplegia, an adaptation which may contribute to orthostatic tolerance. The significant reduction in CFA(flow) from supine to 45 HUT in the tetraplegia group may be related to the completeness of lesion rather than the level of lesion. PMID- 15273902 TI - Difficulty with evacuation after spinal cord injury: colonic motility during sleep and effects of abdominal wall stimulation. AB - Difficulty with evacuation (DWE) is common in individuals with spinal cord injury (SCI). Numerous studies have concluded that constipation, impaction, and incontinence cause significant morbidity and, collectively, constitute an important quality-of-life issue in individuals with SCI. Colonic motor activity was assessed using a solid-state manometry probe. We report here that colonic pressure activity is depressed during sleep compared to that observed in able bodied controls. In addition, pressure activity was decreased during sleep compared to pre-sleep and post-sleep. We suspect that this may contribute to delayed colon transit time after SCI. In addition, since contraction of the abdominal wall musculature plays a role in normal defecation, we assessed whether an abdominal belt with implanted electrodes would improve DWE. In this respect, we demonstrated that neuromuscular stimulation of the abdominal wall improves a number of indices of defecatory function, including time to first stool and total bowel care time. PMID- 15273903 TI - Lack of justification for routine abdominal ultrasonography in patients with chronic spinal cord injury. AB - Little evidence-based research is available to indicate which procedures should routinely be performed for screening exams in patients with spinal cord injuries (SCIs). It had been the procedure to routinely perform abdominal ultrasonography on a yearly basis at our medical center. Therefore, we conducted a retrospective study to determine whether the repetition of these procedures resulted in detection of any pathology warranting treatment that otherwise would have gone undetected. The electronic records of 174 individuals were reviewed, along with a total of 359 abdominal ultrasounds and exams. High incidences of abnormal findings were found in the liver, pancreas, spleen, gallbladder, and kidney; however, no specific interventions were noted solely on the basis of the ultrasound findings. Moreover, no added benefits could be documented through the performance of repetitive exams. We recommend that further evidence-based studies be performed to ascertain the benefits of performance of routine procedures in patients with SCIs. PMID- 15273904 TI - Continued commitment to cutting-edge research benefits patients today. PMID- 15273905 TI - James J. Peters: vision accomplished! PMID- 15273906 TI - [Anti-infective drug therapy in ophthalmology -- Part 1: bacterial infections]. AB - This review presents a rational antibacterial therapy. First, systemic versus topical administration of the drugs is discussed including substances specific for the respective routes. Secondly, ocular diseases requiring systemic therapy are presented followed by a discussion of infections suitable for topical administration. Finally, the pros and cons of a combined antibiotic-steroid therapy are discussed and examples described. PMID- 15273907 TI - [Reduction of decentration after LASIK using a modified eye tracker ring for the MEL-70 excimer laser]. AB - BACKGROUND: The aim of this study was to determine and compare the rate of eccentric laser ablation after LASIK depending on the eye tracker ring used. PATIENTS AND METHODS: All LASIK treatments were carried out using the MEL-70 flying spot excimer laser (Zeiss-Meditec, Jena). The flap was produced using a Corneal Shaper trade mark or Hansatome trade mark Microkeratome (B and L Surgical, Heidelberg). Initially we used an 11 mm eye tracker ring without hinge protector. At the end of February 2001 this ring was replaced by a 10 mm and a 9.5 mm ring with built-in hinge protector. An additional modification was introduced by us: at 1 mm separations little teeth-like spikes were engraved into the eyeward side of the ring, thus stabilising the position of the ring on the globe and allowing free liquid to flow through the spaces between each spike. The built-in calibration system of the corneal topography (TMS 3, Tomey, Erlangen) from patients with a follow-up of one month or longer was used to determine the distance between the centre of the ablation zone from the fixation point. RESULTS: In group I patients (old ring) 42 eyes were treated. In 4 eyes ablation was perfect, in 21 eyes the ablation centre was located 0.1 to 0.49 mm from the fixation point, in 11 eyes 0.51 to 0.99 mm and in 5 eyes 1.1 to 1.49 mm whereas one eye showed a decentred ablation of 1.53 mm. In group II (new ring) 42 eyes were investigated also. In 11 eyes ablation was perfect, in 20 eyes the ablation centre was located 0.1 to 0.49 mm from the fixation point, in 10 eyes 0.5 to 0.99 mm and one eye had an eccentric ablation of 1.28 mm from the fixation point. CONCLUSION: The further development of our eye tracker ring for the MEL-70 laser considerably reduced the rate of decentred ablations. An enhanced grip of the ring onto the globe reduces a slow slide during the laser procedure. PMID- 15273908 TI - [Ocular hemodynamics in normal tension glaucoma: effect of bimatoprost]. AB - BACKGROUND: Altered ocular perfusion plays a role in the pathophysiology of normal tension glaucoma. Prostaglandin-like substances are very effective in lowering intraocular pressure. Less data are available regarding the influence of these compounds on ocular perfusion. In the present study the effects of bimatoprost, which has recently been shown to increase the vascular tone of ciliary arteries in vitro, on the blood flow velocity are investigated. PATIENTS AND METHODS: n = 9 eyes from 9 normal tension glaucoma patients were subjected to color Doppler imaging (CDI) before and during a 3 - 5 week therapy with bimatoprost. RESULTS: Bimatoprost reduces intraocular pressure from 14.0 +/- 0.4 to 11.0 +/- 0.5 mmHg (n = 9; P < 0.001). Systolic as well as diastolic blood flow velocities, resistive index (RI) and pulsatility index (PI), measured by CDI, were unaltered in the presence of bimatoprost. DISCUSSION: Bimatoprost does not influence blood flow velocities in the retrobulbar vessels. The in vitro observation of increased vascular tone in the presence of bimatoprost seems not to be relevant for ocular hemodynamics. PMID- 15273909 TI - ["Doctor, how much can I really see?"]. PMID- 15273910 TI - [Strategies of visual acuity assessment]. AB - Visual acuity can be assessed by different strategies. The constant-stimulus-, the stair-case-, the Best-PEST-strategies, and the EN ISO 8596 are discussed. In cases of presumed psychogenic visual impairment and malingering it can be useful to modify these strategies striving for a determination of the (minimum) visual acuity based on statistical analysis rather than "clinical experience". The EN ISO 8596 defines visual acuity by a 60 % criterion, thus implying guesswork, if recognition of the optotypes is uncertain. Therefore, the forced-choice-procedure is mandatory. Objective psychophysical (preferential looking, induction or suppression of rhythmic eye movements) and electrophysiological methods (VEP) of visual acuity assessment can be useful in infants, in the mentally handicapped, and in patients with presumed psychogenical influence or malingering. PMID- 15273911 TI - [Central vision]. AB - The clinical assessment of vision by means of optotypes does by no means test just two-point resolution, since a correct naming of the letters or digits requires a preceding visual object recognition. Cortical lesions can massively deteriorate vision up to a "Seelenblindheit" in spite of intact optics and retina. There are different processing levels involved in the analysis which can be individually defective, leading to disorders from visual indiscrimination to agnosia or anomia. PMID- 15273912 TI - [Seeing in the dusk: physiological basis and investigation]. AB - Dark adaptation and seeing in the dusk require a complex interaction of the cone and rod system. Whereas the former provides high temporal resolution and colour vision in daylight, temporal resolution of the latter is smaller but sensitivity higher by a factor of 100 to 1,000. The two operational ranges overlap by several decades. Characteristic symptoms of disease may be derived from the systems' physiological function and should be enquired about specifically. Problems due to opacity of the optic media or reduced visual acuity should be differentiated from night vision disorders in the more specific sense. Apart from a routine ophthalmological examination a set of other tests should be used: When rods function normally the second limb of the dark adaptation curve initiates at 5 - 12 min und reaches a normal absolute threshold after 30 - 40 min; dark adapted visual fields are unrestricted apart from a physiological central scotoma, and the dark adapted electroretinogram (ERG) shows normal amplitudes and latencies for low intensity flashes. The influence of glare on mesopic contrast sensitivity may be investigated with a mesoptometer. Night vision disorders may arise from cone system dysfunction, too. Intact cone vision provides high visual acuity, normal colour vision, normal photopic visual fields, a quick regeneration within minutes during the first limb of the dark adaptation curve and normal single flash and oscillatory potentials in the light-adapted ERG. Localised defects may be detected using the multifocal ERG. Interpretation of the results must account for the age-related decay of contrast sensitivity and speed of adaptation. PMID- 15273913 TI - [Visual acuity testing in pre-school children: a comparison between the Sheridan Gardiner test and the Rader (broken wheel) test]. AB - PURPOSE: To evaluate the comparability between the well-established Sheridan Gardiner test (SGT) and a new type of visual acuity test, called the Rader test (RT = broken wheel test) in pre-school children, and to compare test durations of these infant visual acuity tests. SUBJECTS AND METHODS: The RT consists of 16 cards with visus values of 0.16, 0.2, 0.25, 0.3, 0.5, 0.6, 0.8 and 1.0. One pair of cards depicting a car is used for testing. On one of the cards the car has intact wheels, on the other the wheels are incomplete, symbolized by a Landolt ring. The child must indicate, at a viewing distance of 3 meters, which of the wheels is incomplete. The SGT consists of seven visus plates: 5/60, 5/36, 5/24, 5/18, 5/12, 5/9 and 5/6. Each level is tested with one letter and can be repeated by the presentation of a further letter (A, H, O, T, U, V, X). The examination distance is 5 meters. The child must indicate, with reference to a card depicting all seven symbols, which letter the examiner is showing. The SGT and RT were performed in a randomized cross-over sequence in 30 children (20male, 10 female) of pre-school age (from 2 years up to and including the age of 5 years, mean 3.4 years +/- 0.77 SD, median 3.0 years). In all cases, the right eye was examined first. Examination duration was assessed for each acuity test, and for each eye separately with a stopwatch. The instruction time was not considered. The possible visual acuity values of both bests were replaced by a unified scale of visual acuity levels (ranging from 1 to 10). A difference of at least two levels was considered as relevant. The results were compared by means of the sign test at a significance level of 0.05. RESULTS: In particular, for higher visual acuity levels there were considerable differences, with SGT generally showing better results than RT: in 11 of 29 children, in both eyes RT values turned out to be at least 2 lines better than those obtained with SGT. The contrary situation, i. e., favoring SGT by more than 2 lines compared to RT, never occurred. According to the sign test, these differences were significant (p < 0.001). SGT revealed also clearly better visual acuity levels in those 22 children out of the 30, who exhibited differences by 2 lines or more in at least one eye (p < 0.001). The examination procedure with RT revealed problems in making the required directional decisions, especially between 2 and 4 years of age. This might interfere with the test interpretation and lead to distortion of the RT results. Total examination duration did not differ considerably between SGT (1.6 to 5.8 minutes, median 3.0 minutes) and RT (1.6 to 9.4 minutes, median 4.6 minutes), respectively. CONCLUSIONS: The Sheridan-Gardiner test generally shows better results than the new Rader (RT = broken wheel) test in pre-school children. Problems in making the required directional decisions may interfere with RT in this age group. PMID- 15273914 TI - [Visual object and space perception battery: normal values for children from 8 to 12]. AB - BACKGROUND: Diagnostics of central visual perception is a relevant branch of developmental medicine and neuropsychological diagnostic efforts of morphological or functional lesions of the brain. However, no assessment battery for testing the central-visual perception in German-speaking children exists. PROBANDS AND METHOD: In 30 children, aged 8 - 12 years, the Visual Object and Space Perception Test Battery (VOSP) was applied. RESULTS: The group values were documented as standard values. Instructions and aims of the tests were well comprehended by the children. CONCLUSION: The VOSP is well applicable to children. Further studies examining correlation between the VOSP and clinical data are warranted. PMID- 15273917 TI - [Epidemiology of attempted suicide using drugs. An inquiry from the Dresden University Clinic]. AB - BACKGROUND AND OBJECTIVE: Little representative data of the epidemiology of attempted suicide exists in Germany. In this study the frequency of parasuicidal drug intoxication, the distribution of age and gender, as well as the kind and origin of used drugs were evaluated. Furthermore the knowledge about used drugs and possible adverse effects of a previously given medication were analysed. PATIENTS AND METHODS: Over a period of 2 years (January 1998-December 1999) 155 patients (41 males, 114 females, average age 40.5 years) with drug intoxication by attempted suicide were recruited at the University Hospital of Dresden, Germany, for further retrospective analysis. RESULTS: 74 % of these patients were women. Sedatives and hypnotics were most frequently used for parasuicide (44 %), followed by analgesics (18 %) and antidepressants (12 %). Benzodiazepines and benzodiazepine-agonists were the most commonly used drugs (32 %). Moreover, 80 % of all drugs used had been prescribed by physicians. Approximately half of the patients were well-informed about drugs taken. In 43 (47 %) of 92 patients with long-term medication an adverse effect was considered as a possible cause of the attempted suicide. CONCLUSION: Our data underline the importance of attempted suicide in view of the frequency of their use, the need of hospitalization, the required intensive care and possible relapses. Because the majority of drugs used were prescribed by physicians, before giving any medication to their possible suicidal use should be considered. PMID- 15273918 TI - [Effect of immunosuppression-induced hypogonadism on bone metabolism after heart transplantation]. AB - BACKGROUND AND OBJECTIVE: Accelerated bone loss is a well recognized complication after cardiac transplantation (HTx). The role of an immunosuppressive-induced hypogonadism, a well-known cause of osteoporosis in men and its prevention are less defined after HTx. The aim of this study was first, to evaluate the incidence of hypogonadism after HTx and its influence on bone mineral metabolism and second, to assess the effect of a testosterone replacement therapy in hypogonadal transplants. PATIENTS AND METHODS: Due to hormonal status, 88 male cardiac transplants were randomised to a normogonadal or hypogonadal group. At baseline as well as after 1 and 2 years bone mineral density (BMD g/cm (2), T score) was measured at the lumbar spine with DEXA. All patients received a basic therapy of calcium and vitamin D. The hypogonadal patients received additional testosterone. RESULTS: 21 patients (24 %) showed an age-independent hypogonadism. Hypogonadal transplants showed a significant lower BMD (p < 0.001) (BMD = 0.8070 g/cm (2), T-value = -2.6514) than normogonadal patients (BMD = 0.9882 g/cm (2), T value = -1.0568). Despite testosterone replacement hypogonadal patients showed no significant additional increase in BMD over 1 - 2 years compared with the normogonadal. CONCLUSION: Male cardiac transplants in all age groups show an high prevalence of hypogonadism (approximately 25 %) which contributes to a significant bone loss. An additional testosterone substitution did not significantly increase BMD. PMID- 15273919 TI - [Pyogenic liver abscess in chronic alcoholic pancreatitis]. AB - HISTORY AND ADMISSION FINDINGS: A 44-year-old patient was transferred for further treatment of pyogenic liver abscess and a severe attack of a chronic pancreatitis for strong upper right quadrant abdominal pain and recurring fever. INVESTIGATIONS: Laboratory results revealed a significant inflammatory constellation. Abdominal ultrasound was performed which showed a big pyogenic abscess in the right lobe of the liver. Escherichia coli and Enterococcus faecalis could be isolated from abscess aspirates. Endoscopic retrograde cholangiography (ERC) without access of the pancreatic duct showed stenosis of the Ductus hepatocholedochus which was treated with a biliary endoprothesis. DIAGNOSIS, TREATMENT AND COURSE: Antibiotic treatment and percutaneous drainage led to complete remission of the abscess. A few days after discharge the patient returned with identical clinical symptoms. Abdominal ultrasound showed recurrence of the abscess. Because of excessively high pancreatic amylase in aspirated abscess material the patient underwent endoscopic retrograde cholangiopancreaticography (ERCP). There, a pancreatico-hepatic fistula was seen, probably the result of necrosis caused by a severe acute attack of the chronic pancreatitis. After insertion of a naso-fistular drainage, continous rinse and appropriate antibiotic therapy both abscess and fistula completely disappeared without recurrence. CONCLUSION: The rare case of a pancreatic fistula should be considered when a pyogenic liver abscess follows an episode of acute pancreatitis or attack of chronic pancreatitis. Determination of pancreatic amylase in aspired abscess material can be an important step towards correct diagnosis. PMID- 15273920 TI - [Pyogenic liver abscess. State-of-the-art situation for diagnosis and therapy]. PMID- 15273921 TI - [Postmenopausal hormone therapy]. AB - The menopausal transition is a natural phase of every womans life. Several symptoms emerging during the climacteric may be treated with estrogens. Hot flushes and symptoms of genital ageing may be relieved by estrogen therapy. Progestin therapy is indicated in all women with an intact uterus to avoid endometrial pathology. Given the present risk-benefit scenario, estrogen therapy is no longer a first-line choice to prevent osteoporosis. Counselling requires integration of recent knowledge provided by the first results of the Womens Health Initiative and other significant randomized, placebo-controlled, prospective clinical studies with clinically relevant outcomes, not the least unless good-quality data regarding other estrogens and progestins, apart from conjugated equine estrogens/medroxyprogesterone acetate are lacking. Women should be enabled to decide upon estrogen therapy after provision of extensive information by their physicians. PMID- 15273922 TI - ["Medical futility": the doctor caught between the demands for and the limitations of treatment]. AB - The term medical futility is applied to justify withholding treatments in patients who are likely to gain minimal benefit. Futility refers to limiting the moral obligation to provide medical treatment on the basis of clinical efficacy. Current discussions about the meaning of medical futility revolve around two distinct but related concerns. Firstly, whether it is possible to define medical futility in a way that enables objective assessment. Secondly, who should determine when a treatment is futile. In this article we summarize the debate about futility and discuss the relevance of this concept. PMID- 15273923 TI - [Can we still afford progress in intensive care medicine? Re: the article from DMW 1-2/2004]. PMID- 15273924 TI - [Can we still afford progress in intensive care medicine? Re: the article from DMW 1-2/2004]. PMID- 15273925 TI - [Chronic headache: improvement using acupuncture. Re: the article from DMW 20/2004]. PMID- 15273926 TI - [Breast cancer during pregnancy -- a challenge for the anaesthesiologist?]. PMID- 15273927 TI - [How to establish an preoperative anaesthetic clinic]. AB - The preoperative anaesthetic clinic has established at large german hospital since 1980 because of shorter ways, waiting times and best preoperative risk management. Now there is an increasing need for the preoperative anaesthetic clinic in all german hospitals because of the introduction of DRGs for hospital billing since 2003. Organization, leadership, equipment and aspects of the personnel are discussed. Tasks of a "anaesthetic management center" can be fulfilled by the preoperative anaesthetic clinic like OP-organization, distribution of intensive care units, implementation of clinical pathways and quality management. Thus requires an adequate equipment and personnel. An model of rotation for the physician personnel is introduced. Cooperation of the OP departments is necessary. At least our preoperative anaesthetic day clinic has become an logistic centre and an speaking point for patients and the OP department. PMID- 15273928 TI - [Relationship between nutrition and ASA-classification in the elderly]. AB - OBJECTIVE: Old age and bad nourishment are risk factors for the postoperative period. In this study, the "mini nutritional assessment" (MNA) of elderly patients was evaluated before the operation and compared with their ASA classification. METHODS: 215 outpatients (age > 60 years) were included. MNA score was fixed as follows: MNA 24 - 30 = normal (MNA I); MNA 17 - 23.5 = risk of malnutrition (MNA II; MNA < 17 = undernourished (MNA III). In addition, the ASA score of all patients was registered. chi (2)-, Mann-Whitney-U- and correlation analysis were used for statistical analysis. A cut off-value of 24 was fixed for MNA and correlated with the ASA-score. RESULTS: 34.9 % of all patients were allocated to MNA II or III, but only 19.9 % to ASA III or IV. The sensitivity of the ASA-classification for evaluation of the nutritional status was 0.33, selectivity was 0.87, positive predictive value was 0.58 and negative predictive value was 0.70. CONCLUSION: ASA evaluation is not suitable for assessment of the nutritional status. With regard to typical postoperative complications, the nutritional status of patients should be assessed separately. PMID- 15273929 TI - [Cross-validation of simple questioning methods to determine patient satisfaction with anaesthesia care]. AB - INTRODUCTION: Evaluation of patient satisfaction with the perioperative care is an integral part of modern quality management. For this purpose simple questioning techniques are often used in clinical practise or reports of clinical trials. However, little research has been performed to investigate whether these tools and methods are useful and provide valid information. METHODS: Two-hundred patients undergoing elective surgical procedures were interviewed. Five different simple techniques or questions that have been used in the international and in the German literature throughout the last years were applied in random sequence together with the validated German translation of the QoR-9 questionnaire to measure quality of recovery. All analyses were performed descriptively. RESULTS: All investigated techniques could be answered without help by another person in 95 % up to 100 %. All simple dichotomous questions regarding satisfaction were insufficient to discriminate satisfied from less satisfied patients. A rating using grades known from the German school system (1 - 6) and a visual analogue scale (VAS; 0 - 100 mm) obtained ratings lower than the maximum possible values in 10 % and 11 % of the patients respectively. Furthermore, the flexibility that was provided by these tools was not utilized by the patients. The results of the QoR-9 questionnaire as a marker of postoperative recovery showed only a moderate correlation with ratings of patient satisfaction. CONCLUSION: Until now, there is no method or questioning technique in the German language that can be recommended for a quality assurance program. Thus, further research is needed to develop tools that provide valid information with adequate resolution to allow discrimination of patient satisfaction with perioperative care. PMID- 15273930 TI - [Tumorectomy in conscious patient with suspected pregnancy associated breast cancer under cervical epidural anesthesia]. AB - In this report we present a 35 year old pregnant woman (no significant disease in patient history, non smoker, primipara, gestational week 15) who had to undergo elective tumorectomy due to suspected pregnancy associated breast cancer. General anesthesia during pregnancy can potentially be harmful for the fetus (hypoxia, acidosis, premature delivery, teratogenicity). We decided to anesthetize the patient with a cervical segmental epidural block. PMID- 15273931 TI - [Perioperative coagulation disorders]. PMID- 15273934 TI - Genetic association of the R620W polymorphism of protein tyrosine phosphatase PTPN22 with human SLE. AB - We genotyped 525 independent North American white individuals with systemic lupus erythematosus (SLE) for the PTPN22 R620W polymorphism and compared the results with data generated from 1,961 white control individuals. The R620W SNP was associated with SLE (genotypic P=.00009), with estimated minor (T) allele frequencies of 12.67% in SLE cases and 8.64% in controls. A single copy of the T allele (W620) increases risk of SLE (odds ratio [OR]=1.37; 95% confidence interval [CI] 1.07-1.75), and two copies of the allele more than double this risk (OR=4.37; 95% CI 1.98-9.65). Together with recent evidence showing association of this SNP with type 1 diabetes and rheumatoid arthritis, these data provide compelling evidence that PTPN22 plays a fundamental role in regulating the immune system and the development of autoimmunity. PMID- 15273936 TI - Everyday inspiration. PMID- 15273943 TI - A systematic integrative review of infant pain assessment tools. AB - PURPOSE: To examine the issue of pain assessment in infants by acquiring all available published pain assessment tools and evaluating their reported reliability, validity, clinical utility, and feasibility. DESIGN AND METHODS: A systematic integrative review of the literature was conducted using the following databases: MEDLINE and CINAHL (through February 2004), and Health and Psychosocial Instruments, and Cochrane Systematic Reviews (through 2003). MeSH headings searched included "pain measurement," with limit of "newborn infant"; "infant newborn"; and "pain perception." SUBJECTS: Thirty-five neonatal pain assessment tools were found and evaluated using predetermined criteria. The critique consisted of a structured comparison of the classification and dimensions measured. Further, the population tested and reports of reliability, validity, clinical utility, and feasibility were reviewed. RESULTS: Of the 35 measures reviewed, 18 were unidimensional and 17 were multidimensional. Six of the multidimensional measures were published as abstracts only, were not published at all, or the original work could not be obtained. None of the existing instruments fulfilled all criteria for an ideal measure; many require further psychometric testing. CONCLUSIONS: When choosing a pain assessment tool, one must also consider the infant population and setting, and the type of pain experienced. The decision should be made after carefully considering the existing published options. Confidence that the instrument will assess pain in a reproducible way is essential, and must be demonstrated with validity and reliability testing. Using an untested instrument is not recommended, and should only occur within a research protocol, with appropriate ethics and parental approval. Because pain is a multidimensional phenomenon, well-tested multidimensional instruments may be preferable. PMID- 15273935 TI - Genomewide linkage scan for myopia susceptibility loci among Ashkenazi Jewish families shows evidence of linkage on chromosome 22q12. AB - Mild/moderate (common) myopia is a very common disorder, with both genetic and environmental influences. The environmental factors are related to near work and can be measured. There are no known genetic loci for common myopia. Our goal is to find evidence for a myopia susceptibility gene causing common myopia. Cycloplegic and manifest refraction were performed on 44 large American families of Ashkenazi Jewish descent, each with at least two affected siblings. Individuals with at least -1.00 diopter or lower in each meridian of both eyes were classified as myopic. Microsatellite genotyping with 387 markers was performed by the Center for Inherited Disease Research. Linkage analyses were conducted with parametric and nonparametric methods by use of 12 different penetrance models. The family-based association test was used for an association scan. A maximum multipoint parametric heterogeneity LOD (HLOD) score of 3.54 was observed at marker D22S685, and nonparametric linkage analyses gave consistent results, with a P value of.0002 at this marker. The parametric multipoint HLOD scores exceeded 3.0 for a 4-cM interval, and significant evidence of genetic heterogeneity was observed. This genomewide scan is the first step toward identifying a gene on chromosome 22 with an influence on common myopia. At present, we are following up our linkage results on chromosome 22 with a dense map of >1,500 single-nucleotide-polymorphism markers for fine mapping and association analyses. Identification of a susceptibility locus in this region may eventually lead to a better understanding of gene-environment interactions in the causation of this complex trait. PMID- 15273944 TI - Guide to a systematic physical assessment in the infant with suspected infection and/or sepsis. AB - This article provides the resources for the bedside caregiver to conduct a focused physical assessment of the infant with suspected sepsis. The importance of obtaining a complete history to identify associated obstetric and neonatal risk factors is emphasized. Further evaluation of the infant's clinical presentation for signs and symptoms suggestive of a systemic inflammatory response to infection is discussed, along with a step-by-step guide to a systematic physical assessment. Strategies to evaluate physical findings in context with the available diagnostic data to develop a differential diagnosis are provided. The international consensus definitions for the sepsis continuum are presented and are compared and contrasted to the definitions more commonly used in the neonatal population. The article provides tables that can serve as checklists to structure a thorough obstetric and neonatal history and to further evaluate the infant's systemic inflammatory response to infection. PMID- 15273946 TI - The effect of fluid density and volume on the accuracy of test weighing in a simulated oral feeding situation. AB - BACKGROUND: For preterm infants and infants who have difficulty with oral feeding, excessive drooling during oral feedings can result in inaccurate assessment of intake. The drooled volume is typically estimated by visual and tactile assessment of the bib. Research, however, has demonstrated that visual assessment is inaccurate. PURPOSE: The purpose of this study was to determine the accuracy of a scale that was used for the test weighing of milk that was drooled during a study of oral feeding for preterm infants. Additionally, the effect of weighing solutions with different densities on the accuracy of test weights was examined. METHOD: Descriptive, comparative design. PROCEDURES: A simulated feeding situation was performed using 3 fluids (water, Enfamil(20), and Enfamil(24)) and 3 volume ranges (1 mL to 10 mL, 11 mL to 20 mL, and 21 mL to 30 mL). Data collection sessions were conducted for each of the 3 fluids using each range of volumes, for a total of 180 test weights. The research assistant performing the test weights was blinded to the preweight of the bib and the amount of fluid being applied to the bib. RESULTS: Differences between the actual volume applied to the bib and the volume estimated by the scale were very small, with 51% of the differences equaling 0 mL and 48% of the differences between -1 mL and 1 mL. There were significant differences in errors related to both the type of fluid (F = 25.7; df = 2; P < 0.001) and volume range (F = 12.7; df = 2; P < 0.001), as well as for the interaction between the 2 factors (F = 7.02; df = 4; P < 0.001). Water had significantly less mean error than either formula, and large volumes had significantly greater mean error than either small or medium volumes. CONCLUSIONS: Test weighing is an accurate method for measuring fluids of different densities and volumes in a simulation of drooling during oral feeding. The increased error with larger volumes of higher density solutions was not clinically significant. The study supports the need to consider both the accuracy of the scale and characteristics of the fluid when test weighing is used to measure volumes of fluids. PMID- 15273947 TI - Reducing the risk for preterm birth: evidence and implications for neonatal nurses. AB - The incidence of preterm birth has been slowly rising despite advances in obstetric health care. Neonatal nurses have ongoing contact with women who have had a preterm birth and, thus, who are at high risk to have another preterm birth. This article discusses current evidence about reducing risk for preterm birth. Although preterm birth cannot always be prevented, preconception care can help identify and modify maternal risk and promote optimal health before conception. Quality prenatal care, as defined by the Institute of Medicine, consists of continuing risk assessment, health promotion, and interventions to modify medical and psychosocial risk. When these 3 components are consistently applied, they may confer some protection against preterm birth. Women at highest risk for preterm birth need to seek prenatal care from an expert in maternal fetal medicine. Women also need to learn about the common, subtle signs of preterm labor so they can recognize symptoms and quickly seek treatment. Neonatal nurses can provide critical information and anticipatory guidance to women at risk for preterm birth so they can make sound decisions about future pregnancies. PMID- 15273948 TI - Reducing the risk of preterm birth. PMID- 15273951 TI - Covariate adjustment in clinical trials with non-ignorable missing data and non compliance. AB - Estimating causal effects in psychiatric clinical trials is often complicated by treatment non-compliance and missing outcomes. While new estimators have recently been proposed to address these problems, they do not allow for inclusion of continuous covariates. We propose estimators that adjust for continuous covariates in addition to non-compliance and missing data. Using simulations, we compare mean squared errors for the new estimators with those of previously established estimators. We then illustrate our findings in a study examining the efficacy of clozapine versus haloperidol in the treatment of refractory schizophrenia. For data with continuous or binary outcomes in the presence of non compliance, non-ignorable missing data, and a covariate effect, the new estimators generally performed better than the previously established estimators. In the clozapine trial, the new estimators gave point and interval estimates similar to established estimators. We recommend the new estimators as they are unbiased even when outcomes are not missing at random and they are more efficient than established estimators in the presence of covariate effects under the widest variety of circumstances. PMID- 15273952 TI - Methods for modelling change in cluster randomization trials. AB - Randomized trials are often designed to assess an intervention's ability to change patient knowledge, behaviour or health. The study outcome will then need to be measured at least twice for each subject--prior to random assignment and following implementation of the intervention. In this paper we consider methods for modelling change when data are obtained from cluster randomization trials where the unit of allocation is a family, school or community. Attention focuses on mixed effects linear regression extensions of (i) two-sample t-tests and (ii) analysis of covariance, in both cases accounting for dependencies among cluster members. Algebraic expressions for tests of the intervention effect are derived for the special case where there are a fixed number of subjects per cluster while simulation studies are used to compare the power of these procedures in the more realistic case where there is variability in cluster size. A key conclusion is that there can be considerable gains in power when allowing for different individual-level and cluster-level associations between the baseline and follow up assessments. The discussion is illustrated using data from a school-based smoking prevention trial. PMID- 15273953 TI - Sample size for testing and estimating the difference between two paired and unpaired proportions: a 'two-step' procedure combining power and the probability of obtaining a precise estimate. AB - Clinical trials and scientific research studies are currently planned calculating sample sizes to fulfill power requirements, but the simultaneous need to obtain a satisfactorily precise effect estimate is not widely recognized. I have devised a 'two-step' iterative procedure for comparing two binomial parameters for two paired and unpaired proportions (the most frequent situations in scientific research), which takes into account power and the probability of obtaining a predetermined precision of the effect estimate. The first step provides the sample size for the power of the statistical test, the expected width of its corresponding confidence interval, and the probability of obtaining, under the alternative hypothesis, confidence intervals whose width is less than that expected. The second step iteratively increases this sample size until the probability of obtaining such confidence intervals exceeds a required threshold. PMID- 15273954 TI - Development and validation of a prognostic model for survival time data: application to prognosis of HIV positive patients treated with antiretroviral therapy. AB - The process of developing and validating a prognostic model for survival time data has been much discussed in the literature. Assessment of the performance of candidate prognostic models on data other than that used to fit the models is essential for choosing a model that will generalize well to independent data. However, there remain difficulties in current methods of measuring the accuracy of predictions of prognostic models for censored survival time data. In this paper, flexible parametric models based on the Weibull, loglogistic and lognormal distributions with spline smoothing of the baseline log cumulative hazard function are used to fit a set of candidate prognostic models across k data sets. The model that generalizes best to new data is chosen using a cross-validation scheme which fits the model on k-1 data sets and tests the predictive accuracy on the omitted data set. The procedure is repeated, omitting each data set in turn. The quality of the predictions is measured using three different methods: two commonly proposed validation methods, Harrell's concordance statistic and the Brier statistic, and a novel method using deviance differences. The results show that the deviance statistic is able to discriminate between quite similar models and can be used to choose a prognostic model that generalizes well to new data. The methods are illustrated by using a model developed to predict progression to a new AIDS event or death in HIV-1 positive patients starting antiretroviral therapy. PMID- 15273955 TI - Local influence analysis when interfacing toxicokinetic and proportional hazard models. AB - We present a local influence analysis to assigned model quantities in the context of a dose-response analysis of cancer mortality in relation to estimated absorbed dose of dioxin. The risk estimation is performed using dioxin dose as a time dependent explanatory variable in a proportional hazard model. The dioxin dose is computed using a toxicokinetic model, which depends on some factors, such as assigned constants and estimated parameters. We present a local influence analysis to assess the effects on final results of minor perturbations of toxicokinetic model factors. In the present context, there is no evidence of local influence in risk estimates. It is however possible to identify which factors are more influential. PMID- 15273956 TI - Imputing gene-treatment interactions when the genotype distribution is unknown using case-only and putative placebo analyses--a new method for the Genetics of Hypertension Associated Treatment (GenHAT) study. AB - There is a sizeable literature on methods for detecting gene-environment interaction in the framework of case-control studies, particularly with reference to the assumption of independence of genotype and exposure. In the context of a clinical trial, wherein gene-drug interactions with regard to outcomes are examined, these methods may be readily applied, as gene and drug are independent by randomization. In an active-controlled trial (experimental treatment vs standard) that has collected genotype information, gene-drug interactions can be estimated. In addition, the effect of the experimental treatment vs placebo can be imputed by using data from a historical placebo-controlled trial (standard vs placebo) if either (a) genotype information is available from the historical trial or (b) assumptions are made about the prevalence of genotype and the odds ratios of genotype and disease in the historical trial using information from other studies. Motivation for these procedures is provided by the Genetics of Hypertension Associated Treatment, a large pharmacogenetics, ancillary study of a hypertension clinical trial, and examples from published hypertension trials will be used to illustrate the methods. PMID- 15273957 TI - Estimation of infectious disease parameters from serological survey data: the impact of regular epidemics. AB - Serological surveys are a useful source of information about epidemiological parameters for infectious diseases. In particular they may be used to estimate contact rates, forces of infection, the reproduction number and the critical vaccination threshold. However, these estimation methods require the assumption that the infection is in endemic equilibrium. Such equilibria seldom exist in practice: for example, many common infections of childhood exhibit regular epidemic cycles. In this paper, we investigate whether ignoring such cycles produces biased estimates. We apply the methods to data on mumps and rubella in the U.K. prior to the introduction of the combined measles, mumps, rubella (MMR) vaccine. We conclude that past epidemics have only a marginal effect on estimates, and that standard methods that do not adjust for regular epidemics are valid. PMID- 15273958 TI - Flexible modelling of discrete failure time including time-varying smooth effects. AB - Discrete survival models have been extended in several ways. More flexible models are obtained by including time-varying coefficients and covariates which determine the hazard rate in an additive but not further specified form. In this paper, a general model is considered which comprises both types of covariate effects. An additional extension is the incorporation of smooth interaction between time and covariates. Thus, in the linear predictor smooth effects of covariates which may vary across time are allowed. It is shown how simple duration models produce artefacts which may be avoided by flexible models. For the general model which includes parametric terms, time-varying coefficients in parametric terms and time-varying smooth effects estimation procedures are derived which are based on the regularized expansion of smooth effects in basis functions. The approach is used to model the sojourn time in a psychiatric hospital. It is demonstrated how initial conditions which have non-linear influence are damped over time. PMID- 15273959 TI - Modelling relative survival using transformation methods. AB - Patient survival is often assessed as a measure of effectiveness of treatment in cancer clinical trials. The relative survival rate is a measure of patient survival corrected for the effect of other causes of death. We incorporate an additive hazards model for the overall force of mortality focusing attention on the form of the underlying disease process. Transformation models for the cause specific hazard rate describing the disease process are considered. We demonstrate a method for estimating the transformation parameter providing insight into the appropriate choice of model structure. The models include the additive and multiplicative structures as special cases. We write the models in the generalized linear model framework and demonstrate ease of fitting via existing statistical software. The methodology is applied to a Hodgkin's disease data set to assess the effect of clinical trial results on population survival. Our analysis concludes that the multiplicative structure provides a more appropriate description of the data as compared to the additive structure. PMID- 15273960 TI - The association between HLA-DPB1Glu69 and chronic beryllium disease and beryllium sensitization. AB - BACKGROUND: Several case-control studies have found an association between chronic beryllium disease (CBD) and HLA-DPB1 gene variants. However, the relationship between HLA-DPB1 and beryllium sensitization, and whether the presence of one or two HLA-DPB1(Glu69) alleles is differentially associated with CBD and beryllium sensitization have not been completely resolved. METHODS: Restriction fragment length polymorphism (RFLP) analysis was used to address these questions in a large population-based cohort consisting of 884 beryllium workers (90 with CBD, 64 beryllium sensitized). RESULTS: HLA-DPB1(Glu69) was associated with both CBD (OR = 9.4; 95% CI = 5.4, 16.6) and sensitization (OR = 3.3, 95% CI = 1.9, 5.9). Further, workers with CBD and sensitization were more likely to be homozygous HLA-DPB1(Glu69) compared to workers without disease or sensitization (P < 0.001). CONCLUSIONS: Follow-up of this cohort, scrutiny of HLA DPB1 haplotypes, and evaluation of gene-environment and gene-gene interactions will be important for fully understanding the immunogenetic nature of this occupational disease. PMID- 15273961 TI - Spray-painting and chronic airways obstruction. AB - BACKGROUND: The aim was to investigate the respiratory response of HDI-based paint aerosol within the context of the protection afforded by current exposure guidelines. METHODS: A cross-sectional study of 240 painters spraying polyurethane enamels was undertaken at four aircraft maintenance plants. Questionnaire and spirometric data were related to gravimetric measures of cumulative total and respirable paint aerosol (TPA and RPA) and estimated isocyanate in total and respirable aerosols (TIA and RIA). RESULTS: Average cumulative exposures in mg/m(3)-years +/- SD were 159.0 +/- 115.2 TPA, 19.1 +/- 13.8 RPA, 15.8 +/- 11.5 TIA, and 1.9 +/- 1.4 RIA. After adjusting for smoking and asthma symptoms, higher exposures were associated with statistically significant reduction in expiratory flowrates. Significant smoking-related reductions were also observed, without exposure interactions. CONCLUSIONS: These results suggest important respiratory effects from exposures to spray paint aerosols at levels generally in compliance with existing standards for otherwise unregulated particulates and for the isocyanate component of the paint. PMID- 15273962 TI - Lack of modulating influence of GSTM1 and GSTT1 polymorphisms on urinary biomonitoring markers in coke-oven workers. AB - BACKGROUND: Coke-oven workers (COWs) are occupationally exposed to high concentrations of polycyclic aromatic hydrocarbons (PAHs). Urinary 8-hydroxy-2 deoxyguanosine (8-OH-dG) and 1-hydroxypyrene (1-OHP) are biological markers of oxidative DNA damage and PAHs metabolism, respectively. In this study, we investigated whether polymorphisms of glutathione S-transferase (GSTM1 and GSTT1) can modulate the relationship between urinary 8-OH-dG and 1-OHP concentrations among the COWs. METHODS: This was a cross-sectional study. Between February and November of 2001, 53 topside-oven and 130 side-oven workers with the presence of GSTM1 and GSTT1 genotypes were investigated. RESULTS: Urinary 1-OHP and 8-OH-dG concentrations (mean +/- SD) in the topside-oven workers with the presence of GSTM1 were 107.2 +/- 107.9 and 15.3 +/- 9.7 ng/ml, respectively, which were not significantly different from those in the absence of GSTM1 (84.1 +/- 104.5 and 12.8 +/- 14.1 ng/ml). The similar insignificant results were also noted in the sideoven workers. For GSTT1 polymorphism, the results remained insignificant. In contrast, individual excretion of urinary 8-OH-dG and 1-OHP concentrations were still highly correlated (Spearman correlation coefficients: r = 0.43, P < 0.0001, n = 183). CONCLUSIONS: GST may not play a role in the regulation of metabolism of urinary biological markers in COWs. PMID- 15273963 TI - Impairment of cell and plasma redox state in subjects professionally exposed to chromium. AB - BACKGROUND: Chromium (Cr) is widely used in chemical, tannery, building, and metal industries. More recently, it has been demonstrated that Cr induces oxidative stress in mouse brain. Nevertheless very few data exist on in vivo oxidative damage in humans exposed to Cr. METHODS: Changes in antioxidant parameters both in plasma (acid ascorbic redox state and total antioxidant capacity) and in red blood cells (glutathione (GSH) redox state) of 40 subjects (age 37.65 +/- 7.46; M/F 20/20) professionally exposed to Cr who were recruited from metal, chemistry, and building industries were evaluated. We also evaluated the levels of lipoperoxidation (thiobarbituric acid-reactive material, TBA-RM) and thiol levels in plasma to assess the extent of oxidative stress state. To evaluate Cr exposure rate, we measured urinary-chromium (U-Cr) by an electrothermic atomization-atomic absorption spectrometry (ETA-AAS) method. RESULTS: In this study, we found that Cr exposure induced a decrease both in GSH (P < 0.0005) and GSH/oxidized glutathione (GSSG) ratio (P < 0.0001) in red blood cells from workers with respect to control subjects. Furthermore, we also demonstrated a significant decrease of plasma acid ascorbic levels (45.7 +/- 14.9 vs. 53.5 +/- 16.5 micromol/L; P < 0.05) and in total plasma antioxidant capacity (1,126.3 +/- 212.2 vs. 1,266.9 +/- 207.8 micromol/L; P < 0.05) in subjects exposed to Cr. No difference was found with regard to TBA-RM and thiol levels. CONCLUSIONS: This study demonstrated that in humans, an oxidative stress occurs for Cr exposures as low as those considered safe. This oxidative stress appears to be able to affect intracellular and plasmatic antioxidant defense. PMID- 15273964 TI - Airflow obstruction attributable to work in industry and occupation among U.S. race/ethnic groups: a study of NHANES III data. AB - OBJECTIVES: To estimate the fraction of airflow obstruction attributable to workplace exposure by U.S. race/ethnic group. METHODS: U.S. population-based third National Health and Nutrition Examination Survey (NHANES III) data on 4,086 Caucasians, 2,774 African-Americans, and 2,568 Mexican-Americans, aged 30-75, were studied. Airflow obstruction was defined as FEV1/FVC<75% and FEV1<80% predicted. Weighted prevalence, and prevalence odds ratios (OR) adjusted for the effect of age, smoking status, pack-years, body mass index, education, and socio economic status were estimated using SUDAAN software. RESULTS: Industries with the most cases of airflow obstruction attributable to workplace exposure include: armed forces; rubber, plastics, and leather manufacturing; utilities; textile mill manufacturing; health care; food products manufacturing; sales; construction; and agriculture. The fraction of cases with airflow obstruction associated with work in industry varied by race/ethnic group and was estimated as 22.2% (95% CI 9.1-33.4) among Caucasians, 23.4% (95% CI 2.2-40.0) among African Americans, and 49.6% (32.1-62.6) among Mexican-Americans. CONCLUSIONS: This study found differences in the fraction of airflow obstruction cases associated with employment pattern among major U.S. race/ethnic population groups. PMID- 15273965 TI - Self-reported dermatitis and skin cancer in California farm operators. AB - BACKGROUND: High rates of skin diseases and higher non-melanoma skin cancer rates have been reported in farmers. METHODS: Self-report of dermatitis and skin cancer was among the information collected from 1947 California farm operators, mostly men, in a telephone survey. The majority of the farmers cultivated fruits, nuts, or other field crops. RESULTS: Dermatitis was reported by 8.9% of men and 15.8% of women during the previous 12 months. In a logistic regression model, female gender (OR 2.0, 95% confidence interval 1.3-3.0) and respiratory atopy (OR 1.4, 1.01-1.90) were the only significant independent risk factors for reported dermatosis. There was significantly less reporting of skin cancer among field farmers when compared to others. Regular sunscreen use was reported significantly more often by women (42%) as compared to men (11%). CONCLUSIONS: More in-depth studies are needed to get information on the role of agrochemicals as risk factors for dermatitis and skin cancer. PMID- 15273966 TI - Evaluation of the reliability and validity of the University of South Florida Environmental Assessment Questionnaire. AB - BACKGROUND: The National Institute for Occupational Safety and Health's (NIOSH) National Occupational Research Agenda (NORA), identified "exposure assessment methods" as a priority area for research in occupational medicine. METHODS: The University of South Florida Environmental Assessment Questionnaire (EAQ) describes exposures from the workers point of view. The questionnaire was distributed to a sample of workers drawn from a population of over 3,000 workers employed by a Florida food processing plant. The analysis of the validity was based on 211 subjects and the analysis of the reliability was based on 44 subjects. RESULTS: For the category scores, the sensitivities ranged from 0.67 to 1.0, the specificities ranged from 0.52 to 0.83, the positive predictive values ranged from 0.60 to 1.0, and the negative predictive values ranged from 0.55 to 1.0. All of the weighted kappas for the subcategories were above zero. The Spearman rank-order coefficients were above 0.5 for all of the exposure categories except the mold, plant, and animal exposure category which was 0.49. The correlation coefficient for the questionnaire as a whole was 0.85. CONCLUSIONS: This study showed the EAQ as a whole to be reliable and valid. Gathering valid occupational exposure information with this method was both feasible and economical. PMID- 15273967 TI - P.W.J. Bartrip's attack on Irving J. Selikoff. PMID- 15273968 TI - Workplace policy considerations for curbing the tobacco epidemic. PMID- 15273969 TI - Smoking is an occupational hazard. AB - BACKGROUND: Even though the prevalence of tobacco smoking has declined in the general population and among white-collar workers, the prevalence of tobacco smoking among blue-collar workers remains unacceptably high. Blue-collar workers experience greater exposure to workplace toxins which can add to, or even multiply, their risk of adverse health effects from tobacco smoking. Among blue collar workers, workers in the restaurant, bar, and gaming industries are exposed to much higher levels of environmental tobacco smoke (ETS) than are office workers, and are at increased risk of cancer and cardiovascular diseases even if they are non-smokers themselves. METHODS: The literature on health risks, and the disparity between white and blue collar workers in smoking prevalence, and the literature on various tobacco control strategies provide the sources on which this review is based. CONCLUSIONS: Over the past 20 years, the accumulating scientific evidence about smoking as an occupational hazard has prompted the implementation of various educational, economic, and legal tobacco control strategies. PMID- 15273970 TI - Reducing occupation-based disparities related to tobacco: roles for occupational health and organized labor. AB - BACKGROUND: Persistent and growing occupation-based disparities related to tobacco pose a serious public health challenge. Tobacco exacts a disproportionate toll on individuals employed in working class occupations, due to higher prevalence of smoking and exposure to secondhand smoke among these workers compared to others. METHODS: We provide an overview of recent advances that may help to reduce these disparities, including research findings on a successful social contextual intervention model that integrates smoking cessation and occupational health and safety, and a new national effort to link labor unions and tobacco control organizations around their shared interest in reducing tobacco's threat to workers' health. CONCLUSIONS: Implications of these efforts for future research and action are discussed. PMID- 15273971 TI - Efficacy of tobacco dependence treatment in the context of a "smoke-free grounds" worksite policy: a case study. AB - BACKGROUND: Smoking restrictions provide opportunities to modify smoking behavior. A large insurance company implemented a smoke-free grounds policy at two of their office complexes in January, 2000. METHODS: This cohort study evaluated the impact of the smoke-free grounds policy on abstinence among 128 employees who participated in a tobacco dependence treatment program. RESULTS: The overall quit rate at 6 months was 44.5%. The larger complex showed a trend for higher quit rates compared to the smaller complex (46.5 vs. 28.6%). Post-ban participants had higher quit rates than pre-ban participants (52.4 vs. 43.0%). The probability of abstinence at 6 months follow-up was higher for post-ban compared to pre-ban participants (P = 0.03). Post-ban participants were 80% less likely to relapse than pre-ban participants. Non-quitters decreased their consumption by 6.6 cigarettes/day (39.1% decrease). CONCLUSIONS: A "smoke-free grounds" policy encourages abstinence and may play a significant role in harm reduction among continuing tobacco users. PMID- 15273972 TI - The New York City Smoke-Free Air Act: second-hand smoke as a worker health and safety issue. AB - BACKGROUND: Despite the provisions of a Smoke-Free Air Act (SFAA) enacted in 1995, more than 415,000 non-smoking New York City workers reported exposure to second-hand smoke in the workplace all or most of the time in 2002. Continued exposure to second-hand smoke in New York City prompted a renewed debate about a broader smoke-free air law. METHODS: The approach taken by the New York City Department of Health and Mental Hygiene to make the case for workplace protection from second-hand smoke, counter the opposition's arguments, and ultimately win the support of policymakers and the public for comprehensive smoke-free workplace legislation is described. RESULTS: On December 30, 2002, New York City's Mayor signed the SFAA of 2002 into law, making virtually all workplaces, including restaurants and bars, smoke-free. CONCLUSIONS: Proponents for a stronger law prevailed by defining greater protection from second-hand smoke as a matter of worker health and safety. Efforts to enact smoke-free workplace laws will inevitably encounter strong opposition, with the most common argument being that smoke-free measures will harm businesses. These challenges, however, can be effectively countered and public support for these measures is likely to increase over time by focusing the debate on worker protection from second-hand smoke exposure on the job. PMID- 15273973 TI - Predictors of smoke-free workplaces by employee characteristics: who is left unprotected? AB - BACKGROUND: Over the last decade, there has been steady progress in the adoption of workplace smoking policies in the United States. However, the benefits of a smoke-free workplace are not distributed equally among the workforce. METHODS: Data from 44,357 adults who reported that they work indoors were derived from an optional tobacco module on the 2001 Behavioral Risk Factor Surveillance System (BRFSS), administered by 25 states. Logistic regression was utilized to examine factors associated with the absence of a smoke-free workplace policy. RESULTS: Overall, 70.9% of respondents reported working under a smoke-free workplace policy. The likelihood of being protected by a smoke-free workplace policy was significantly lower among workers who were younger, male, non-white, engaged in heavy labor, earned less than 50,000 US dollars annually, had a high school education or less, smoked everyday, or worked in the South or Midwest. CONCLUSIONS: There remain considerable gaps in smoke-free workplace policy coverage. PMID- 15273974 TI - Clinical pharmacokinetic data of racemic drugs obtained by the indirect method following precolumn diastereomer formation: is the influence of racemization during chiral derivatization significant? AB - This report reviews the stereoselective data of a number of racemic drugs (n = 17) obtained from indirect chiral method (via diastereomer formation) in comparison to similar data generated by the application of a direct chiral method. While it was noted that the indirect method still continued to be used to characterize the stereoselective disposition of racemic drugs, the present review critically evaluates the issue of racemization that has the potential to skew the stereoselective data obtained from the indirect method. The review describes various remedies to counter and/or minimize the impact of racemization on the final outcome of the stereoselective analysis by the indirect method. On the basis of this review it could be concluded that the indirect method is a viable and important tool for gathering stereoselective data of racemic drugs used in medical practice as well for those racemic drugs still in discovery and developmental stages. PMID- 15273975 TI - Chromatographic determination of herbicide residues in various matrices. AB - The newest results in the use of various extraction techniques and chromatographic methods such as gas-liquid and high-performance liquid chromatography used for the assessment of herbicide residues in various matrices have been compiled and critically evaluated. Practical employments in water and soil research, environmental protection, clinical and food chemistry are presented. PMID- 15273976 TI - A new HPLC method for serum neopterin measurement and relationships with plasma thiols levels in healthy subjects. AB - Neopterin, a pyrazinopyrimidine compound, serves as a marker of cellular immune system activation, and it can be used as a prognostic predictor for certain types of diseases. We propose a new simple HPLC method to measure serum neopterin with highly sensitive fluorimetric detection. After TCA serum protein precipitation, the supernatant was diluted five times, injected into a C18 reversed-phase column and eluted at a flow rate of 1.5 mL/min by an isocratic water-acetonitrile (99:1) mobile phase. The natural fluorescence of the molecule was detected at excitation wavelength 353 nm and emission 438 nm. In these conditions the neopterin retention time was about 4 min. Our proposed method was compared with a validated chromatographic separation, and the obtained data of the serum neopterin from 35 healthy volunteers were analysed by Passing-Bablok regression and Bland-Altman test. Neopterin measurement in healthy subjects was also employed to investigate on its potential relationships with plasma thiols levels. PMID- 15273977 TI - Quantitative analysis of phytoestrogens in kudzu-root, soy and spiked serum samples by high-pressure liquid chromatography. AB - A sensitive and reliable HPLC method that allows simultaneous quantification of phytoestrogens extracted from kudzu-root and soy preparations, and serum samples has been developed. Kudzu-root and soy preparations were mixed with 5 microg flavone and 15 microg rutin (internal standards) and the phytoestrogens were extracted by using solid-phase (C18) extraction cartridges. Blank or spiked serum samples were extracted by using either C18 cartridges or trichloroacetic acid methanol extraction. The extracts were analyzed by the HPLC equipped with a reverse-phase (250 x 4 mm, C18) column and UV, diode-array or MS detector. A linear gradient of acetic acid and acetonitrile provided excellent separation of glycoside and aglycone-phytoestrogens from kudzu root and soy preparations. The C18 cartridge extraction of serum yielded excellent recovery of both glycoside- and aglycone-phytoestrogens, while the trichloroacetic acid-methanol extraction yielded excellent recovery of glycoside but poor recovery of aglycone compounds. UV and MS detectors were suitable for phytoestrogen analysis in plant and serum samples, while the diode-array detector was suitable for generating the UV absorbance curve for phytoestrogens. PMID- 15273978 TI - Analysis of the aconitine alkaloids in traditional Chinese medicines by nonaqueous capillary electrophoresis using a new recording mode. AB - The determination of three aconitine alkaloids (hypaconitine, aconitine, mesaconitine) in five traditional Chinese medicines including two Tibetan medicines, Chuanwu, Caowu, Fuzi, Aconitum Tanguticum Maxim and Aconitum Gymnandrum Maxim by non-aqueous capillary electrophoresis using a new recording mode is described. The dissociation constants of aconitine, mesaconitine and hypaconitine have also been determined by CZE and were 7.71, 6.60 and 6.25, respectively. The separation was achieved by optimizing the applied voltage, the pH and the concentration of the buffer. The electrophoretic medium was 20 mM borax-70% (v/v) methanol (pH 8.5) and an uncoated capillary (50 cm x 75 microm i.d.) was used. Detection was carried out with a UV monitor at 214 nm. The total time for separation and determination was under 13 min. PMID- 15273979 TI - Isoform-specific quantification of endothelins in HUVEC culture supernatants by on-line high-performance liquid chromatography/electrospray mass spectrometry. AB - A method for the quantitative analysis of endothelin peptides in human umbilical vein endothelial cell (HUVEC) culture supernatants is reported. The analysis is isoform-specific and employs solid-phase extraction and subsequent HPLC fractionation followed by HPLC-ESIMS analysis. The peptide vasoactive-intestinal contractor (VIC) was used as internal standard for the HPLC-ESIMS analysis. Linearity of calibration curves was from 50 fmol to 25 pmol. The limit of detection of the HPLC-ESIMS step using a buffer matrix was estimated at 50 fmol (S/N > 3). The overall limit of detection for supernatants of HUVEC was 500 fmol/mL. In HUVEC culture supernatants only ions of endothelin-1 (ET1) were observed. Basal levels were determined to be 1.8 +/- 0.3 pmol/mL. Quantitative results obtained for ET1 were in agreement with those obtained by using a standard addition method and by an ELISA method. PMID- 15273980 TI - Rapid separation of barbiturates and benzodiazepines by capillary electrochromatography with 3-(1,8-naphthalimido)propyl-modified silyl silica gel. AB - A capillary electrochromatographic (CEC) method was applied to the simultaneous separation of barbiturates (barbital, phenobarbital, secobarbital and thiopental) and benzodiazepines (nitrazepam, diazepam and triazolam). The separation was performed in a 75 microm i.d. capillary, packed with 3-(1,8-naphthalimido)propyl modified silyl silica gel (NAIP), studying the effects of buffer pH and mobile phase composition. Using an applied voltage of 20 kV and the short-end injection method (9 cm capillary effective length), the mobile phase of 1.0 mM citrate buffer (pH 5.0) containing 45% methanol provided the baseline separation of seven toxic drugs in less than 9 min. In CEC with NAIP, the benzodiazepines were separated by the combination of hydrophobic and pi-pi interactions, whereas the separation of the barbiturates was based on the hydrophobic interaction. PMID- 15273981 TI - Switching from an analogous to a stable isotopically labeled internal standard for the LC-MS/MS quantitation of the novel anticancer drug Kahalalide F significantly improves assay performance. AB - The importance of a stable isotopically labeled (SIL) internal standard for the quantitative LC-MS/MS assay for Kahalalide F in human plasma is highlighted. Similar results can be expected for other LC-MS/MS assays. Therefore, we emphasize the need for an SIL internal standard for accurate and precise LC-MS/MS assays of drugs in biological matrices. PMID- 15273982 TI - A more sensitive MS detector does not obviously lead to a more sensitive assay: experiences with ES-285. AB - In this paper the transfer of an existing method for the quantitative determination of the anticancer agent ES-285 in human plasma using liquid chromatography tandem mass spectrometry on an API 365 to an API 3000 mass spectrometer is described. The transfer appeared not to be straightforward. Problems arose resulting from carry-over and interferences. In addition, due to the expansion of the calibration range, data ought to be weighted with a different factor to increase the accuracy of the lower concentrations. After finding appropriate solutions for these problems, the lower limit of quantitation could be lowered from 10 to 1 ng/mL for ES-285 in human plasma. The usefulness and necessity of the modified assay was demonstrated by analysis of plasma samples from a patient receiving a low dosage of the drug. PMID- 15273983 TI - Silence of the fathers: early X inactivation. AB - X chromosome inactivation is the mammalian answer to the dilemma of dosage compensation between males and females. The study of this fascinating form of chromosome-wide gene regulation has yielded surprising insights into early development and cellular memory. In the past few months, three papers reported unexpected findings about the paternal X chromosome (X(p)). All three studies agree that the X(p) is imprinted to become inactive earlier than ever suspected during embryonic development. Although apparently incomplete, this early form of inactivation insures dosage compensation throughout development. Silencing of the X(p) persists in cells of extraembryonic tissues, but it is erased and followed by random X inactivation in cells of the embryo proper. These findings challenge several aspects of the current view of X inactivation during early development and may have profound impact on studies of pluripotency and epigenetics. PMID- 15273984 TI - Coming to our senses. AB - Sensory organs are specialized to receive different kinds of input from the outside world. However, common features of their development suggest that they could have a shared evolutionary origin. In a recent paper, Niwa et al. show that three Drosophila adult sensory organs all rely on the spatial signals Decapentaplegic and Wingless to specify their position, and the temporal signal ecdysone to initiate their development. The proneural gene atonal is an important site for integration of these regulatory inputs. These results suggest the existence of a primitive sensory organ precursor, which would differentiate according to the identity of its segment of origin. The authors argue that the eyeless gene controls eye disc identity, indirectly producing an eye from the sensory organ precursor within this disc. PMID- 15273985 TI - Renewed debate over postnatal oogenesis in the mammalian ovary. AB - The central dogma of female reproductive biology has long held that oogenesis ceases prior to birth in mammals. During the first half of the last century, there was much debate about whether this was the case or whether oogenesis continued in the postnatal ovary. A report in 1951 effectively put an end to this debate and laid the foundation for the dogma. A new paper by Johnson et al. (2004) resurrects the debate over whether postnatal oogenesis occurs in the mammalian ovary. If confirmed, this would have tremendous impact on issues related to female fertility and reproductive longevity. PMID- 15273986 TI - Mouse-centric comparative transcriptomics of protein coding and non-coding RNAs. AB - The largest transcriptome reported so far comprises 60,770 mouse full-length cDNA clones, and is an effective reference data set for comparative transcriptomics. The number of mouse cDNAs identified greatly exceeds the number of genes predicted from the sequenced human and mouse genomes. This is largely because of extensive alternative splicing and the presence of many non-coding RNAs (ncRNAs), which are difficult to predict from genomic sequences. Notably, ncRNAs are a major component of the transcriptomes of higher organisms, and many sense antisense pairs have been identified. The ncRNAs function in a range of regulatory mechanisms for gene expression and other biological processes. They might also have contributed to the increased functional diversification of genomes during evolution. In this review, we discuss aspects of the transcriptome of various organisms in relation to the mouse data, in order to shed light on the regulatory mechanisms and physiological significance of these abundant RNAs. PMID- 15273987 TI - Polycystins and mechanosensation in renal and nodal cilia. AB - The external surfaces of the human body, as well as its internal organs, constantly experience different kinds of mechanical stimulations. For example, tubular epithelial cells of the kidney are continuously exposed to a variety of mechanical forces, such as fluid flow shear stress within the lumen of th nephron. The majority of epithelial cells along the nephron, except intercalated cells, possess a primary cilium, an organelle projecting from the cell's apical surface into the luminal space. Despite its discovery over 100 years ago, the primary cilium's function continued to elude researchers for many decades. However, recent studies indicate that renal cilia have a sensory function. Studies on polycystic kidney disease (PKD) have identified many of the molecular players, which should help solve the mystery of how the renal cilium senses fluid flow. In this review, we will summarize the recent breakthroughs in PKD research and discuss the role(s) of the polycystin signaling complex in mediating mechanosensory function by the primary cilium of renal epithelium as well as of the embryonic node. PMID- 15273988 TI - Opposing FGF and retinoid pathways: a signalling switch that controls differentiation and patterning onset in the extending vertebrate body axis. AB - Construction of the trunk/caudal region of the vertebrate embryo involves a set of distinct molecules and processes whose relationships are just coming into focus. In addition to the subdivision of the embryo into head and trunk domains, this "caudalisation" process requires the establishment and maintenance of a stem zone. This sequentially generates caudal tissues over a long period which then undergo differentiation and patterning in the extending body axis. Here we review recent studies that show that changes in the signalling properties of the paraxial mesoderm act as a switch that controls onset of differentiation and pattern in the spinal cord. These findings identify distinct roles for different caudalising factors; in particular, Fibroblast Growth Factor (FGF) inhibits differentiation in the caudal stem zone, while Retinoic acid (RA) provided rostrally by somitic mesoderm is required for neuronal differentiation and establishment of ventral neural pattern. Furthermore, the mutual opposition of FGF and RA pathways controls not only neural differentiation but also mesoderm segmentation and might also underlie the progressive assignment of rostrocaudal identity by regulating Hox gene availability and activation. PMID- 15273989 TI - The ins and outs of lysophosphatidic acid signaling. AB - Lysophosphatidic acid (LPA) is a lipid mediator with a wide variety of biological actions, particularly as an inducer of cell proliferation, migration and survival. LPA binds to specific G-protein-coupled receptors and thereby activates multiple signal transduction pathways, including those initiated by the small GTPases Ras, Rho, and Rac. LPA signaling has been implicated in such diverse processes as wound healing, brain development, vascular remodeling and tumor progression. Knowledge of precisely how and where LPA is produced has long proved elusive. Excitingly, it has recently been discovered that LPA is generated from precursors by 'autotaxin', a once enigmatic exo-phosphodiesterase implicated in tumor cell motility. Exogenous phospholipases D can also produce LPA, which may contribute to their toxicity. Here we review recent progress in our understanding of LPA bioactivity, signaling and synthesis. PMID- 15273990 TI - The PARP superfamily. AB - Poly(ADP-ribosyl)ation is an immediate DNA-damage-dependent post-translational modification of histones and other nuclear proteins that contributes to the survival of injured proliferating cells. Poly(ADP-ribose) polymerases (PARPs) now constitute a large family of 18 proteins, encoded by different genes and displaying a conserved catalytic domain in which PARP-1 (113 kDa), the founding member, and PARP-2 (62 kDa) are so far the sole enzymes whose catalytic activity has been shown to be immediately stimulated by DNA strand breaks. A large repertoire of sequences encoding novel PARPs now extends considerably the field of poly(ADP-ribosyl)ation reactions to various aspects of the cell biology including cell proliferation and cell death. Some of these new members interact with each other, share common partners and common subcellular localizations suggesting possible fine tuning in the regulation of this post-translational modification of proteins. This review summarizes our present knowledge of this emerging superfamily, which might ultimately improve pharmacological strategies to enhance both antitumor efficacy and the treatment of a number of inflammatory and neurodegenerative disorders. A provisional nomenclature is proposed. PMID- 15273991 TI - The redox regulation of intermediary metabolism by a superoxide-aconitase rheostat. AB - In this article, we discuss a hypothesis to explain the preferential synthesis of the superoxide sensitive form of aconitase in mitochondria and the phenotype observed in manganese superoxide dismutase mutant mice, which show a gross over accumulation of stored fat in liver. The model proposes that intermediary metabolism is redox regulated by mitochondrial superoxide generated during mitochondrial respiration. This regulates the level of reducing equivalents (NADH) entering the electron transport chain (ETC) through the reversible inactivation of mitochondrial aconitase. This control mechanism has a dual function; firstly, it regulates levels of superoxide generated by the ETC and, secondly, it fine-tunes metabolism by channeling citrate either for the production of NADH for energy metabolism or diverting it for the synthesis of fats. In this setting, the mitochondrial redox state influences metabolic decisions via a superoxide-aconitase rheostat. PMID- 15273992 TI - Functional genomics studied by proteomics. AB - The human genome contains about 30,000 genes, each creating several transcripts per gene. Transcript structures and expression are studied by high-throughput transcriptomic techniques using microarrays. Generally, transcripts are not directly operating molecules, but are translated into functional proteins, post translationally modified by proteolysis, glycosylation, phosphorylation, etc., sometimes with great functional impact. Proteins need to be analyzed by proteomic techniques, less suited for high-throughput. Two-dimensional polyacrylamide gel electrophoresis (2D-PAGE), separating thousands of proteins has developed slowly over the past quarter of a century. This technique is now quite reproducible and suitable for differential proteomics, comparing normal and diseased cells/tissues revealing differentially regulated proteins. 2D-PAGE is combined with protein identification methods, currently mass spectrometry (MS), which has been significantly improved over the last decade. Other proteomic techniques studying protein-protein interactions are now either established or still being developed, such as peptide or protein arrays, phage display, and the yeast two-hybrid system. The strengths and weaknesses of these techniques are discussed. PMID- 15273993 TI - Serial analysis of gene expression (SAGE): unraveling the bioinformatics tools. AB - Serial analysis of gene expression (SAGE) is a powerful technique that can be used for global analysis of gene expression. Its chief advantage over other methods is that it does not require prior knowledge of the genes of interest and provides qualitative and quantitative data of potentially every transcribed sequence in a particular cell or tissue type. This is a technique of expression profiling, which permits simultaneous, comparative and quantitative analysis of gene-specific, 9- to 13-basepair sequences. These short sequences, called SAGE tags, are linked together for efficient sequencing. The sequencing data are then analyzed to identify each gene expressed in the cell and the levels at which each gene is expressed. The main benefit of SAGE includes the digital output and the identification of novel genes. In this review, we present an outline of the method, various bioinformatics methods for data analysis and general applications of this important technology. PMID- 15273995 TI - Bioanalysis: its past, present, and some future. AB - An overview of about 100 years of bioanalysis is here disastrously attempted. The beginning of rigorous analytical systems can perhaps be traced back to the building and testing of the analytical ultracentrifuge by Svedberg and the apparatus for moving-boundary electrophoresis of Tiselius, both systems relying on expensive and hard to operate machines. In the sixties, Porath discovered porous beads for the determination of relative molecular mass (Mr) of proteins, based on the principle of molecular sieving. Concomitantly, Svensson and his pupil Vesterberg described a revolutionary principle for fractionating proteins in a nonisocratic environment, based on generation of stable pH gradients in an electric field, a technique that went down to history as isoelectric focusing (IEF). Polyacrylamide gel electrophoresis (PAGE), with the brilliant idea of discontinuous buffers, was brought to the limelight and in 1967, sodium dodecyl sulfate (SDS)-PAGE was described, permitting easy assessment of protein purity and reasonable measurements of Mr values of denatured polypeptide chains. By the mid seventies, another explosive concept was realized: orthogonal combination of two unrelated techniques, based on surface charge and mass fractionation, namely, two-dimensional (2-D) PAGE already in the very first papers by O'Farrell elaborated to its utmost sophistication. The eighties saw the systematic growth of 2-D PAGE, accompanied by systematic efforts to develop instrumentation for large-scale production of 2-D maps and computer evaluation for 2-D map analysis, based on the sophisticated algorithms adopted by astronomers for mapping stars in the sky. Another fundamental innovation in the field of IEF was the discovery of immobilized pH gradients (IPGs) that brought the much needed reproducibility in 2 D maps while allowing exquisite resolution in very narrow pH ranges. The nineties were definitely the decade of capillary zone electrophoresis, with the concomitant concept of automation and miniaturization in electrokinetic methodologies. Also 2-D map analysis witnessed a big revival, thanks to the adoption of IPGs for the first dimension. The enormous progress of mass spectrometry resulted in first reports on the analysis of macromolecules and the building of data bases on gene and protein banks. The third millennium is, perhaps, exasperating the concept of miniaturization at all costs, while not disdaining increasingly larger maps for 2-D analysis of complex protein mixtures. PMID- 15273996 TI - Alkali-stable high-pI isoelectric membranes for isoelectric trapping separations. AB - Alkali-stable, high-pI isoelectric membranes have been synthesized from quaternary ammonium derivatives of cyclodextrins and poly(vinyl alcohol), and bifunctional cross-linkers, such as glycerol-1,3-diglycidyl ether. The new, high pI isoelectric membranes were successfully applied as cathodic membranes in isoelectric trapping separations in place of the hydrolytically more labile, polyacrylamide-based cathodic isoelectric membranes, and permitted the use of catholytes as alkaline as 1 M NaOH. The new high-pI isoelectric membranes have shown excellent mechanical stability, low electric resistance and long life times, even when subjected to electrophoresis with current densities as high as 80 mA/cm2. PMID- 15273997 TI - A continuous-flow instrument for the determination of linear polyacrylamide stability. AB - Alkaline hydrolysis of high-molecular-mass (10 MDa) linear polyacrylamide, a widely used replaceable sieving medium for the capillary electrophoresis of DNA fragments and proteins, was investigated. The release rate of ammonia, a product of amide group hydrolysis, was monitored by a high-sensitivity continuous-flow system. The experimental results show a rapid onset of hydrolysis at a temperature of 70 degrees C. While the degree of 1 h hydrolysis was evaluated to reach about 16% at 70 degrees C and pH 13, it drops to 5% at 50 degrees C and pH 12.5. PMID- 15273998 TI - Sequencing of oligosaccharides derivatized with benzylamine using electrospray ionization-quadrupole time of flight-tandem mass spectrometry. AB - Oligosaccharides were derivatized by reductive amination using benzylamine and analyzed by nanoelectrospray ionization-quadrupole time of flight-tandem mass spectrometry (nanoESI-QTOF-MS/MS) in the positive ion mode. The major signals were obtained under these conditions from the [M+H]+ ions for all benzylamine derivatized oligosaccharides. To obtain structural information from these derivatized oligosaccharides, MS/MS was applied. Protonated molecular ions underwent extensive fragmentation, even under low-energy collision-induced dissociation. MS/MS spectra of [M+H]+ ions are characterized by simple fragmentation patterns which result from cleavage of the glycosidic bonds and thus allow a straightforward interpretation. Fragmentation of the [M+H]+ ions gave predominantly B- and Y-type glycosidic fragments. A systematic study of various oligosaccharides showed that information on sugar sequence and branching could easily be obtained. Predictable and reproducible fragmentation patterns could be obtained in all cases. This derivatization procedure and mass spectrometric methodology were applied successfully to neutral and acidic glycans released from 10 microg of glycoproteins separated by gel electrophoresis. Moreover, the derivatives retain their sensitivity to exoglycosidases. Thus a series of sequential on-target exoglycosidase treatments combined with matrix assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF MS) was found to be useful for the determination of structural features of the glycans released from proteins separated by gel electrophoresis such as the monosaccharide sequence, branching pattern, and anomeric configurations of the corresponding glycosidic linkages. Our strategy can be used successfully to assign the major glycans released from proteins separated by gel electrophoresis. PMID- 15273999 TI - Matrix-assisted laser desorption/ionization-time of flight-mass spectrometry of lipopolysaccharide species separated by slab-polyacrylamide gel electrophoresis: high-resolution separation and molecular weight determination of lipooligosaccharides from Vibrio fischeri strain HMK. AB - We recently demonstrated that the combined use of lipopolysaccharide (LPS) reverse staining and high-efficiency passive elution techniques can be successfully used as a suitable interface between LPS slab-gel separation and electrospray ionization-mass spectrometry (ESI-MS) of LPS-derived oligosaccharides. Here, we extend our micropurification strategy for the analysis of O-deacylated LPS forms from Vibrio fischeri HMK after recovery from single reverse-stained LPS bands using matrix-assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS). The quantities (30-40 microg) obtained from the two gel-resolved LPS bands were sufficient to allow MALDI-TOF MS detection of O-deacylated LPS glycoforms at m/z 3767.1, 3890.1 for the high molecular-weight or at m/z 2522.5, 2645.4, 2725.7, and 2848.7 for the low molecular-weight LPS band. These LPS band heterogeneities resulted not only from variations in the oligosaccharide region of the LPS but also from two phosphorylation states of the lipid A (diphosphoryl and diphosphoryl plus a single phosphoethanolamine substitution). On the other hand, MALDI-TOF mass spectra of the separated LPS bands displayed reduced heterogeneity and increased signal-to-noise ratios as compared to spectra of the unpurified LPS. Furthermore, micropurification of LPS bands prior MALDI-TOF-MS led to a higher sensitivity of detection of less abundant low-molecular-weight LPS glycoforms. Taken together, this and our previous study on gel-micropurified LPS using ESI definitively show how one can unambiguously determine the different molecular species contained within each gel-separated LPS band, their relative abundance and oligosaccharide sequences. PMID- 15274000 TI - Which genetic marker for which conservation genetics issue? AB - Conservation genetics focuses on the effects of contemporary genetic structuring on long-term survival of a species. It helps wildlife managers protect biodiversity by identifying a series of conservation units, which include species, evolutionarily significant units (ESUs), management units (MUs), action units (AUs), and family nets (FNs). Although mitochondrial DNA (mtDNA) evolves 5 10 times faster than single-copy nuclear DNA (scnDNA), it records few traces of contemporary events. Thus, mtDNA can be used to resolve taxonomic uncertainties and ESUs. Variable number of tandem repeats (VNTRs) evolve 100-1000 times faster than scnDNA and provide a powerful tool for analyzing recent and contemporary events. VNTR analysis techniques include polymerase chain reaction (PCR)-based microsatellite assays and oligonucleotide probing. Size homoplasy problems in PCR based microsatellite assays can strongly affect the inference of recent population history. The high homozygosity in endangered species is reflected in a relatively low number and level of variability in microsatellite loci. This combined with "allelic dropout" and "misprinting" errors contributes to the generation of highly biased genetic data following analyses of natural populations. Thus, in conservation genetics, microsatellites are of limited use for identifying ESUs, MUs, and AUs. In contrast to PCR-based microsatellite analysis, oligonucleotide probing avoids errors resulting from PCR amplification. It is particularly suitable for inferring recent population history and contemporary gene flow between fragmented subpopulations. Oligonucleotide fingerprinting generates individual-specific DNA banding patterns and thus provides a highly precise tool for monitoring demography of natural populations. Hence, DNA fingerprinting is powerful for distinguishing ESUs, MUs, AUs, and FNs. The use of oligonucleotide fingerprinting and fecal DNA is opening new areas for conservation genetics. PMID- 15274001 TI - Capillary electrophoresis sequencing of small ssDNA molecules versus the Ogston regime: fitting data and interpreting parameters. AB - We study the mobility of short ssDNA fragments (approximately 30-500 bases) separated by capillary electrophoresis in entangled polymer solutions. Although this corresponds to what is commonly called the Ogston regime, the corresponding sieving concept has never been defined properly nor tested quantitatively. We consider three formulas that have been suggested to fit data in this range of ssDNA sizes, and we discuss how their free parameters are related to actual physical parameters. We test these formulas with new data obtained in our laboratory using a commercial poly-N,N-dimethylacrylamide sieving matrix. Our results show that all three formulas provide decent fits. However, the traditional Ogston equation produces fitting parameters that appear to lack physical meaning. Surprisingly, all three approaches predict that the effective pore size and fiber radius are almost equal. This is the first step towards the development of a systematic approach to optimizing sequencing systems for this size range. PMID- 15274002 TI - Separation of double-stranded DNA fragments by capillary electrophoresis: Impacts of poly(ethylene oxide), gold nanoparticles, ethidium bromide, and pH. AB - The separation of DNA by capillary electrophoresis using poly(ethylene oxide) (PEO) containing gold nanoparticles (GNPs) is presented. The impacts of PEO, GNPs, ethidium bromide (EtBr), and pH on the separation of double-stranded DNA have been carefully explored. Using a capillary dynamically coated with 5.0% poly(vinylpyrrolidone) and filled with 0.2% PEO containing 0.3 x GNPs (the viscosity less than 15 cP), we have demonstrated the separation of DNA markers V and VI within 5 min at pH 8.0 and 9.0. In terms of resolution and reproducibility, GNPs have a greater impact on the separation of DNA at pH 9.0. Resolution improvements for large DNA fragments (> 300 base pairs, bp) are greater than those for small ones in the presence of GNPs. It is important to point out that reproducibility is excellent (relative standard deviations for the migration times less than 0.5%) and thus no further dynamic coating is required in at least 20 consecutive runs in the presence of GNPs. Using 0.2% PEO (pH 9.0) containing 0.3 x GNPs, the separation of DNA fragments ranging in size from 21 to 23,130 bp was accomplished in 7 min. The results presented in this study show the advantage of PEO containing GNPs for DNA separation, including rapidity, high resolving power, excellent reproducibility, and ease of filling capillaries. PMID- 15274003 TI - A column gel electrophoresis-coupled genomic DNA subtractive hybridization technique. AB - We have developed a DNA subtractive hybridization technique especially designed for mammalian genome comparison. The core of this protocol is a newly devised denaturant-containing polyacrylamide gel formed in a glass-column. In this gel system, the following DNA manipulation steps are carried out sequentially: size separation by electrophoresis, heat-denaturation, renaturation, and recovery. In the first step, a mixture of tester and driver DNA fragments are segregated according to their size whilst keeping their double-stranded forms. This reduces the complexity of the original genomic DNA fragments and also segregates DNA fragments having closely related sequences. In the second step, fractionated DNA fragments are quickly denatured and subjected to successive subtractive hybridization in situ by controlling gel temperature in a water bath. In the third step, DNA fragments are recovered by electrophoresis towards the reverse orientation and are adsorbed onto ion-exchange beads. Two lines of experiments show that our protocol is able to highly enrich or directly extract differences among genomic DNA samples. PMID- 15274004 TI - Gene expression analysis of an integrin family of genes by systematic multiplex reverse transcription-polymerase chain reaction. AB - We have established the systematic multiplex reverse transcription-polymerase chain reaction (SM RT-PCR) system of 26 members of the integrin family of genes, and used this system to investigate their expression in 25 different kinds of human adult tissues. A hierarchical clustering analysis of the tissue expression data illustrated clustering of functionally related tissues. Although the correlation was weak, bone marrow, thymus, and spleen were clustered, potentially confirming the role of certain integrin molecules in the immune response. We also investigated changes in integrin gene expression in cancer. Several differences were observed between normal and tumor, as well as between normal and cancer cell lines. The SM RT-PCR experiments for the study of alternative splicing showed that most of the integrin genes analyzed exhibited fixed ratios of differentially spliced transcripts probably due to a tissue-independent mechanism of splicing predetermined by the nucleotide sequences around the splicing donor and acceptor sites. PMID- 15274005 TI - Refining the results of a whole-genome screen based on 4666 microsatellite markers for defining predisposition factors for multiple sclerosis. AB - Multiple sclerosis (MS) is a demyelinating disease of the central nervous system with a complex genetic background. In order to identify loci associated with the disease, we had performed a genome screen initially using 6000 microsatellite markers in pooled DNA samples of 198 MS patients and 198 controls. Here, we report on the detailed reanalysis of this set of data. Distinctive features of microsatellites genotyped in pooled DNA causing false-positive association or masking existing association were met by improved evaluation and refined correction factors in the statistical analyses. In order to assess potential errors introduced by DNA pooling and genotyping, we resurveyed the experiment in a subset of microsatellite markers using de novo-composed DNA pools. True MS associations of markers were verified via genotyping all individual DNA samples comprised in the pools. Microsatellites share characteristically superb information content but they do not lend themselves to automation in very large scale formats. Especially after DNA pooling many artifacts of individual marker systems require special attention and treatment. Therefore, in the near future comprehensive whole-genome screens may rather be performed by typing single nucleotide polymorphisms on chip-based platforms. PMID- 15274006 TI - Sensitive and simultaneous analysis of five transgenic maizes using multiplex polymerase chain reaction, capillary gel electrophoresis, and laser-induced fluorescence. AB - The benefits of using multiplex polymerase chain reaction (PCR) followed by capillary gel electrophoresis with laser-induced fluorescence (CGE-LIF) for the simultaneous detection of five transgenic maizes (Bt11, T25, MON810, GA21, and Bt176) are demonstrated. The method uses a hexaplex PCR protocol to amplify the five mentioned transgenic amplicons plus the zein gene used as reference, followed by a CGE-LIF method to analyze the six DNA fragments. CGE-LIF was demonstrated very useful and informative for optimizing multiplex PCR parameters such as time extension, PCR buffer concentration and primers concentration. The method developed is highly sensitive and allows the simultaneous detection in a single run of percentages of transgenic maize as low as 0.054% of Bt11, 0.057% of T25, 0.036% of MON810, 0.064% of GA21, and 0.018% of Bt176 in flour obtaining signals still far from the detection limit (namely, the signal/noise ratios for the corresponding DNA peaks were 41, 124, 98, 250, 252, and 473, respectively). These percentages are well below the minimum threshold marked by the European Regulation for transgenic food labeling (i.e., 0.5-0.9%). A study on the reproducibility of the multiplex PCR-CGE-LIF procedure was also performed. Thus, values of RSD lower than 0.67 and 6.80% were obtained for migration times and corrected peak areas, respectively, for the same sample and three different days (n = 12). On the other hand, the reproducibility of the whole procedure, including four different multiplex PCR amplifications, was determined to be better than 0.66 and 23.3% for migration times and corrected peak areas, respectively. Agarose gel electrophoresis (AGE) and CGE-LIF were compared in terms of resolution and sensitivity for detecting PCR products, demonstrating that CGE-LIF can solve false positives induced by artifacts from the multiplex PCR reaction that could not be addressed by AGE. Moreover, CGE-LIF provides better resolution and sensitivity. To our knowledge, these results demonstrate for the first time that multiplex PCR-CGE-LIF is a solid alternative to determine multiple genetically modified organisms in maize flours in a single run. PMID- 15274007 TI - Evaluation and validation of the ABI 3700, ABI 3100, and the MegaBACE 1000 capillary array electrophoresis instruments for use with short tandem repeat microsatellite typing in a forensic environment. AB - The demand for high-throughput DNA profiling has increased with the introduction of national DNA databases and has led to the development of automated methods of short tandem repeat (STR) profile production; however, a potential bottleneck still exists at the gel electrophoresis stage. Capillary electrophoresis sequencers capable of processing 96 samples with considerably reduced manual intervention are now available. In this paper, we compare the ABI Prism 377 slab gel sequencer currently used by the Forensic Science Service with three leading capillary array electrophoresis instruments: the ABI Prism 3700, the Amersham MegaBACE 1000 and the 16-capillary ABI Prism 3100. We describe the experiments used to evaluate and validate these platforms for forensic use with the STR multiplex Ampf/STR SGMplus [1, 2], along with comparative data from the ABI Prism 377 sequencer. PMID- 15274008 TI - Capillary electrochromatography of peptides and proteins. AB - This paper reviews recent progress in bioanalysis using capillary electrochromatography (CEC), especially in the field of separation of proteins and peptides. Fundamentals of CEC are briefly discussed. Since most of the recent developments on CEC have focused on column technology, i.e., design of new stationary phases and development of new column configurations, we describe here a variety of column architectures along with their advantages and disadvantages. Newly emerged column technologies in CEC for high speed and high efficiency separation are also discussed. Different analytical platforms of CEC such as pressure-assisted CEC or voltage-assisted micro- high-performance liquid chromatography (HPLC), CEC with different detection techniques, CEC on microchip platforms and multidimensional electrochromatography with their applications in peptide and protein analysis are presented. PMID- 15274009 TI - On-line capillary electrophoresis-mass spectrometry for the analysis of biomolecules. AB - Mass spectrometry (MS) has become a key tool for the characterization of biologically relevant molecules in the last decade. Due to the complexity of most biological samples an upstream separation is essential. Capillary electrophoresis (CE) has gained much interest due to its high separation efficiency, speed, and often complementary selectivity to liquid chromatography. We describe the state of-the-art of on-line CE-MS for the analysis of molecules of biological origin. The characterization of peptides, including the study of post-translational modifications, intact proteins, oligonucleotides, and related interaction studies are reviewed. Relevant publications are summarized in tables, including some important method parameters. Key applications are discussed with respect to the advantages and limitations of CE-MS. Coupling interfaces, preconcentration techniques, capillary coatings, and the different CE techniques, e.g., capillary zone electrophoresis, capillary isoelectric focusing, capillary gel electrophoresis, etc. are briefly discussed against the background of their bioanalytical applications. PMID- 15274010 TI - Capillary electrophoresis of biological particles: viruses, bacteria, and eukaryotic cells. AB - A review about the application of electrophoretic methods in the capillary format for the investigation of large biological assemblies like viruses, bacteria, yeast or entire mammalian cells is given. These entities are of a size ranging between some nanometers and tens of micrometers. They can form colloidal solutions or dispersions and move under the influence of an electric field. They are separated by zone electrophoresis according to their different electrophoretic velocity, and characterized by the electrophoretic mobility, which is easily determinable in free solution in capillaries or in other microdevices. As the charge of these particles, when being amphoteric, is pH dependent, isoelectric focusing can also be carried out and the capillary format is increasingly being employed for their separation and determination of pI values. Furthermore, interactions with ligands can be assessed by various modes of affinity capillary electrophoresis. Capillary zone electrophoresis has thus become a valuable tool for investigation of large macromolecular assemblies in the field of biochemistry, clinical chemistry, toxicology, and nutrition chemistry amongst many others. PMID- 15274011 TI - Capillary electrophoresis analysis of fosfomycin in biological fluids for clinical pharmacokinetic studies. AB - A feasible capillary zone electrophoresis (CZE) method with indirect UV and contactless conductivity detection was developed for the determination of fosfomycin, an antibiotic, in human plasma and microdialysis samples. Samples were collected from test persons during a clinical trial. The background electrolytes used consisted of 25 mM benzoic acid and 0.5 mM hexadecyltrimethylammonium bromide, adjusted with tris(hydroxymethyl)aminomethane solution to pH 6.95 for plasma, and to pH 8.05 for microdialysis samples. CZE separations of the anionic analyte were carried out with reversed electroosmotic flow directed towards the anode. The limit of detection was between 0.6 and 2 microg/mL, depending on the matrix and the detection method. No sample preparation was needed for microdialysis samples; for plasma samples, proteins were precipitated with methanol (1+2, v+v), and the supernatant was analyzed. The yield determined with spiked samples was about 100%, the reproducibility of the entire method, expressed by the RSD% of three independent determinations of fosfomycin in triplicate after spiking Ringer's solutions and plasma samples, respectively, was better than 8%. The method is thus well-suited for clinical studies for the determination of the antibiotic in biological fluids. PMID- 15274012 TI - Separation and investigation of structure-mobility relationships of insect oostatic peptides by capillary zone electrophoresis. AB - Capillary zone electrophoresis (CZE) has been applied to qualitative analysis, separation, and physicochemical characterization of synthetic insect oostatic peptides (IOPs) and their derivatives and fragments. Series of homologous IOPs were separated in three acidic background electrolytes (BGEs; pH 2.25, 2.30, 2.40) and an alkaline BGE (pH 8.1). Best separation was achieved in acid BGE composed of 100 mM H3PO4, 50 mM Tris, pH 2.25. The effective electrophoretic mobilities, mu(ep), of all IOPs in four BGEs were determined and several semiempirical models correlating effective mobility with charge-to-size ratio (mu(ep) versus q/Mr k) were tested to describe the migration behavior of IOP in CZE. None of models was found to be unambiguously applicable for the whole set of 20 IOPs differing in size (dipeptide - decapeptide) and charge (-2 to +0.77 elementary charges). However, a high coefficient of correlation, 0.9993, was found for the subset of homologous series of IOPs with decreasing number of proline residues at C-terminus, H-Tyr-Asp-Pro-Ala-Prox-OH, x = 6 - 0, for the dependence of mu(ep) on q/Mr k with k = 0.5 for IOPs as anions in alkaline BGE and with k = 2/3 for IOPs as cations in optimized acidic Tris-phosphate BGE. From these dependences the probable structure of IOPs in solution could be predicted. PMID- 15274013 TI - Improved capillary electrophoresis method for the determination of carbohydrate deficient transferrin in patient sera. AB - Capillary zone electrophoresis (CZE) with a dynamic double coating formed by charged polymeric reagents represents an effective tool for the separation of iron-saturated transferrin (Tf) isoforms and thus the determination of carbohydrate-deficient transferrin (CDT, sum of asialo-, monosialo- and disialo Tf in relation to total Tf) in human serum. Using the CEofix-CDT reagents, a 50 microm inner diameter (ID) capillary of 60 cm total length and the P/ACE MDQ under optimized instrumental conditions (20 kV and 30 degrees C) is demonstrated to provide outstanding assay precision for the determination of CDT in human serum. For CDT levels of 1.0% and 4.5%, precision relative standard deviation (RSD) values (n = 8) were determined to be < 3.0% and < 1.5%, respectively. During the first year of operation under routine conditions, more than 600 patient samples were analyzed in a total of 62 sets of runs. Except for selected samples of patients with severe liver diseases, interference-free Tf patterns were detected. Asialo-Tf was not detected in control sera and in patient sera with a CDT level < 1.70%, but became detectable in 89.6% of sera with > 2.3% disialo-Tf. Monosialo-Tf was only detected in two sera containing > 13.3% CDT. The optimized CZE assay was applied to confirm positive CDT results produced by an immunoassay during long-term monitoring of a patient which led to the determination of the elimination kinetics of asialo-Tf, disialo-Tf, and CDT after an episode of high alcohol consumption (estimated apparent half lifes of 4.86, 7.24, and 6.74 days, respectively). The optimized CZE assay with an upper reference limit for CDT of 1.70% represents an attractive alternative to high performance liquid chromatography (HPLC). It features simpler sample preparation, faster analysis time, and higher isoform resolution compared to the most recent HPLC approach and can thus be regarded as a new candidate of a reference method for CDT. PMID- 15274014 TI - Use of a native affinity ligand for the detection of G proteins by capillary isoelectric focusing with laser-induced fluorescence detection. AB - Affinity probe capillary isoelectric focusing (CIEF) with laser-induced fluorescence was explored for detection of Ras-like G proteins. In the assay, a fluorescent BODIPY FL GTP analogue (BGTPgammaS) and G protein were incubated resulting in formation of BGTPgammaS-G protein complex. Excess BGTPgammaS was separated from BGTPgammaS-G protein complex by CIEF using a 3-10 pH gradient and detected in whole-column imaging mode. In other cases, a single point detector was used to detect zones during the focusing step of CIEF using a 2.5-5 pH gradient. In this case, analyte peaks passed the detector in approximately 5 min at an electric field of 350 V/cm. Detection during focusing allowed for more reproducible assays at shorter times but with a sacrifice in sensitivity compared to detection during mobilization. Resolution was adequate to separate BGTPgammaS Ras and BGTPgammaS-Rab3A complexes. Formation of specific complexes was confirmed by adding GTPgammaS to samples containing BGTPgammaS-G protein. GTPgammaS competed with BGTPgammaS for G protein binding sites resulting in decreased BGTPgammaS-G protein peak heights. The concentrating effect of CIEF enabled detection limits of 30 pM. PMID- 15274015 TI - Automated carbohydrate profiling by capillary electrophoresis: a bioindustrial approach. AB - Automated, high-resolution, quantitative, high-throughput analysis of mono- and oligosaccharides, produced by enzymatic digestion of cellohexaose (model substrate) and lignocellulosic biomass, is demonstrated using high-performance capillary electrophoresis in conjunction with a single-step fluorophore labeling strategy for sensitive laser-induced fluorescence detection. Unattended batch sample processing from 96-well plates enabled reliable industrial-scale carbohydrate analysis. Excellent resolution of mono- and oligosaccharides was achieved under suppressed electroosmotic flow conditions, using either covalently or dynamically coated fused-silica capillary columns. The proposed approach readily supports the demands of bioindustrial operation environments with respect to high-throughput carbohydrate profiling. PMID- 15274016 TI - Polymer solutions and entropic-based systems for double-stranded DNA capillary electrophoresis and microchip electrophoresis. AB - We give an overview of recent development of low-viscosity polymer solutions and entropic trapping networks for double-stranded DNA (dsDNA) separations by conventional capillary electrophoresis and microchip electrophoresis. Theoretical models for describing separation mechanisms, commonly used noncross-linked polymer solutions, thermoresponsive (viscosity-adjustable) polymer solutions, and novel entropic trapping networks are included. The thermoresponsive polymer solutions can be loaded at one temperature into microchannels at lower viscosities, and used in separation at another temperature at entanglement threshold concentrations and higher viscosities. The entropic-based separations use only arrays of regular obstacles acting as size-separations and do not need viscous polymer solutions. These progresses have potential in integration to automated capillary and microfluidic chip systems, enabling better reusability of separation microchannels, much shorter DNA separation times, and higher reproducibility due to less matrix degradation. PMID- 15274017 TI - An integrated method for mutation detection using on-chip sample preparation, single-stranded conformation polymorphism, and heteroduplex analysis. AB - This work integrates rapid techniques for mutation detection by producing single stranded DNA and (renatured) double-stranded DNA on-chip, labeling these with fluorescent DNA stains and then performing two complementary methods of mutation detection-single stranded conformation polymorphism (SSCP) analysis and heteroduplex analysis (HA). This involves the denaturation of double-stranded polymerase chain reaction (PCR) product into single-stranded DNA, the mutation analysis of the single-stranded DNA by SSCP and the rehybridized double-stranded DNA by HA. These steps were performed entirely on-chip within several minutes of operation. The combination of these two mutation detection methods on-chip provides a highly sensitive method of mutation detection for either genotyping or screening. Many mutation analysis methods rely upon fluorescently labeled samples from a PCR with fluorescently labeled primers. By labeling on-chip we not only attain improved signal strength, but the method is considerably more versatile. Although we used PCR products in this work, this method could be used to analyze DNA from any source. We believe that this combination of several procedures on a single chip represents a significant step in the development of higher levels of integration upon microfluidic devices. PMID- 15274018 TI - Optimization of the electrokinetic supercharging preconcentration for high sensitivity microchip gel electrophoresis on a cross-geometry microchip. AB - We developed a novel on-line preconcentration procedure for microchip gel electrophoresis (MCGE), which enables application of electrokinetic supercharging (EKS) for highly sensitive detection of DNA fragments on a cross-geometry microchip. In comparison with conventional pinched injection using the cross microchip, the present approach allows loading a much larger amount of the sample by taking advantage of a newly developed operational mode. In order to obtain high preconcentration effect and prevent splitting of an enriched sample into subchannels, i.e., off the detector range, effects of the voltage applied on the reservoirs and the time of isotachophoretic preconcentration were examined. The optimal balance between the voltage and time was found for a high-sensitivity analysis of DNA fragments. After experimental optimization the detection limit of a 150 bp fragment was as low as 0.22 mg/L (S/N = 3) that is 10 times better than using the conventional pinched injection. PMID- 15274019 TI - Integrated on-chip derivatization and electrophoresis for the rapid analysis of biogenic amines. AB - We demonstrate the monolithic integration of a chemical reactor with a capillary electrophoresis device for the rapid and sensitive analysis of biogenic amines. Fluorescein isothiocyanate (FITC) is widely employed for the analysis of amino group containing analytes. However, the slow reaction kinetics hinders the use of this dye for on-chip labeling applications. Other alternatives are available such as o-phthaldehyde (OPA), however, the inferior photophysical properties and the UV lambdamax present difficulties when using common excitation sources leading to a disparity in sensitivity. Consequently, we present for the first time the use of dichlorotriazine fluorescein (DTAF) as a superior in situ derivatizing agent for biogenic amines in microfluidic devices. The developed microdevice employs both hydrodynamic and electroosmotic flow, facilitating the creation of a polymeric microchip to perform both precolumn derivatization and electrophoretic analysis. The favorable photophysical properties of the DTAF and its fast reaction kinetics provide detection limits down to 1 nM and total analysis times (including on-chip mixing and reaction) of <60 s. The detection limits are two orders of magnitude lower than current limits obtained with both FITC and OPA. The optimized microdevice is also employed to probe biogenic amines in real samples. PMID- 15274020 TI - Comprehensive two-dimensional chromatography and capillary electrophoresis coupled with tandem time-of-flight mass spectrometry for high-speed proteome analysis. AB - A comprehensive two-dimensional capillary liquid chromatography and capillary zone electrophoresis system coupled with tandem matrix assisted laser desorption/ionization-time of flight-time of flight-mass spectrometry (MALDI-TOF TOF-MS) proteomics analyzer is presented. Protein/peptide samples were separated by capillary high-performance liquid chromatography (cHPLC). The effluents from cHPLC (the first dimension) were continuously transferred into capillary zone electrophoresis (CZE, the second dimension) through a novel valve-free hydrodynamic sampling interface. The CZE effluents were mixed with alpha-cyano-4 hydroxycinnamic acid (CHCA) matrix sheath flow via CE-MALDI interface, and then directly deposited on the MALDI target at a 3 s time-interval for further MS analysis. The high efficiency of the overall system was demonstrated by analysis of proteins in D20 (human hepatocellular carcinoma model in nude mice with high metastatic potential) liver cancer tissue. More than 300 proteins were identified, which proved the system potential for high-throughput analysis and application in proteomics. PMID- 15274021 TI - Virtual two-dimensional gel electrophoresis of high-density lipoproteins. AB - High-density lipoproteins (HDLs) isolated by immunoaffinity chromatography and separated by immobilized pH gradient-isoelectric focusing (IPG-IEF) were examined by mass spectrometry directly, applying a new proteomics technology, virtual two dimensional (2-D) gel electrophoresis. A preliminary examination of HDL particles has revealed at least 42 unique masses for protein species with isoelectric points between pH 5.47-5.04, some of which have not been observed previously. By delivering masses of intact proteins from complex cellular mixtures in a format that correlates directly to classical 2-D gel analyses, virtual 2-D gel electrophoresis constitutes a general discovery tool to expose and monitor protein isoforms and post-translational modifications. Furthermore, its general ability to deliver ions from sub-picomole level proteins enmeshed in complex cellular mixtures potentially fulfills the need of top-down proteomics to obtain intact protein ions from microscale samples. Additional comparison of such data to 2-D gel analyses and their identified proteins may elucidate the functions of the individual apolipoprotein components and the cardioprotective effects of HDL. PMID- 15274022 TI - Aberrant expression of acute-phase reactant proteins in sera and breast lesions of patients with malignant and benign breast tumors. AB - We have analyzed unfractionated sera of newly diagnosed patients (n=10) with breast carcinoma (BC), prior to treatment, and patients (n=5) with fibrocystic disease of the breast (FDB) by two-dimensional gel electrophoresis (2-DE) and silver staining. The patients' 2-DE serum protein profiles obtained were then subjected to image analysis and compared to similar data generated from sera of normal healthy female controls (n=10) of the same range of age. The relative expression of alpha1-antichymotrypsin (ACT), clusterin, and complement factor B was significantly higher in all BC patients as compared to normal controls. However, the expression of alpha1-antitrypsin (AAT) in BC patients was apparently lower than that of the controls. Similar differential expression of ACT was detected in the FDB patients. The aberrant expression of the serum acute-phase proteins of patients with BC and FDB was confirmed by competitive enzyme-linked immunosorbent assay (ELISA). Similar altered proteins expression was also observed from immunohistochemical studies of malignant (n=5) and benign (n=5) breast lesions of the respective patients performed using antisera to the aberrantly expressed proteins. However, the malignant breast lesions were instead positively stained for AAT. The differential expression of the serum proteins was apparently abrogated when a six-month follow-up study was performed on nine of the BC patients subsequent to treatment. PMID- 15274023 TI - Mining the human cerebrospinal fluid proteome by immunodepletion and shotgun mass spectrometry. AB - We have analyzed the proteome of human cerebrospinal fluid with the help of shotgun mass spectrometry. In order to identify low-abundant proteins in these fluids, we have found it necessary to remove the abundant protein components from the mixture. Immunodepletion of the abundant proteins has allowed us to identify more than 100 proteins in cerebrospinal fluids from a patient suffering from normal pressure hydrocephalus. The identified proteins belong to a variety of different classes ranging from serum proteins to intracellular mediators that are involved in signal transduction and transcription. This work establishes a platform for future studies aimed at the comparative proteome analysis of cerebrospinal fluids from different groups of patients suffering from various psychiatric and neurological disorders. PMID- 15274024 TI - Two-step elution of human serum proteins from different glass-modified bioactive surfaces: a comparative proteomic analysis of adsorption patterns. AB - Plasma protein adsorption patterns on surfaces may give vital information to evaluate biocompatibility of biomaterials designed for direct blood-contacting applications or tissue integration. Adsorption of human serum proteins on four different types of biomaterials (glass, aminosilanized glass, hyaluronan and sulfated hyaluronan) was analyzed by two-dimensional electrophoresis. Desorption of proteins from the surfaces was first classically achieved by sodium dodecyl sulfate (SDS) elution. We introduced a second elution step (by use of isoelectric focusing (IEF) sample buffer consisting of urea, 3-[(3 cholamidopropyl)dimethylammonio]-1-propansulfonate, and dithioerythritol) which allows more stringent elution conditions and is a tool to evaluate the protein adsorption strength to biomaterials. Moreover, the two-step elution may discriminate between irreversible and reversible adsorption of plasma proteins for biomaterials, thus helping to elucidate the structure of protein multilayers which form a complex system at the surfaces. The IEF sample buffer proved not to alter the biomaterial structure and integrity. Hydrophobic bonds resulted to be the main strength driving protein adsorption onto our biomaterials. Apolipoproteins were the most important proteins interacting with the surfaces suggesting that high-density lipoprotein (HDL) particles could play a role in biocompatibility due to their beneficial effects on endothelial cells. PMID- 15274028 TI - ProtocadherinX/Y, a candidate gene-pair for schizophrenia and schizoaffective disorder: a DHPLC investigation of genomic sequence. AB - Protocadherin X and Protocadherin Y (PCDHX and PCDHY) are cell-surface adhesion molecules expressed predominantly in the brain. The PCDHX/Y gene-pair was generated by an X-Y translocation approximately 3 million years ago (MYA) that gave rise to the Homo sapiens-specific region of Xq21.3 and Yp11.2 homology. Genes within this region are expected to code for sexually dimorphic human characteristics, including, for example, cerebral asymmetry a dimension of variation that has been suggested is relevant to psychosis. We examined differences in patients with schizophrenic or schizoaffective psychosis in the genomic sequence of PCDHX and PCDHY in coding and adjacent intronic sequences using denaturing high performance liquid chromatography (DHPLC). Three coding variants were detected in PCDHX and two in PCDHY. However, neither the coding variants nor the intronic polymorphisms could be related to psychosis within families. Low sequence variation suggests selective pressure against sequence change in modern humans in contrast to the structural chromosomal and sequence changes including fixed X-Y differences that occurred in this region earlier in hominid evolution. Our findings exclude sequence variation in PCDHX/Y as relevant to the aetiology of psychosis. However, we note the unusual status of this region with respect to X-inactivation. Further investigation of the epigenetic control of PCDHX/Y in relation to psychosis is warranted. PMID- 15274029 TI - Association of the dopamine transporter gene (DAT1) core promoter polymorphism 67T variant with schizophrenia. AB - Dysfunction of the central dopaminergic neurotransmission has been suggested to play an important role in the etiology of schizophrenia. The dopamine transporter (DAT1) mediates the active reuptake of dopamine from the synapses and thereby plays a key role in the regulation of the dopaminergic neurotransmission. In this study, we sought to determine the possible association of the DAT1 gene core promoter polymorphism -67A/T with schizophrenia in a case/control study. The allele and genotype frequencies of the polymorphism were studied in 100 patients and 100 controls, which were matched on the basis of sex, age, and ethnicity. The genotype frequencies in the patients group were as follows: AA 29%; AT 59%; TT 12% versus the genotype frequencies in the control group: AA 57%; AT 38%; TT 5% [chi2 = 16.54, df = 2, OR = 2.25 (95% CI 1.46-3.45, P < or = 0.0003]. For the first time, these findings provide tentative evidence for the contribution of the DAT1 gene core promoter polymorphism to the etiopathophysiology of schizophrenia at least in the Iranian male population that we studied. Replication studies of independent samples and family-based association studies are necessary to further evaluate the significance of our findings. PMID- 15274030 TI - Evidence for association between novel polymorphisms in the PRODH gene and schizophrenia in a Chinese population. AB - Haploinsufficiency for or mutation in at least one gene from the velocardiofacial syndrome (VCFS) region at chromosome 22q11 is implicated in psychosis. Linkage disequilibrium mapping of the region in patients identified a segment containing two genes, proline dehydrogenase (PRODH) and DGCR6, as candidates [Liu et al., 2002a] and by analysis of additional polymorphisms the PRODH gene was associated with schizophrenia in adult and early onset patients. In the present study we provide additional evidence in support of genetic association between PRODH and schizophrenia in a Chinese population. We analyzed the PRODH gene in a samples of schizophrenic patients and their families from Sichuan, SW China consisting of 528 family trios and sibling pairs. We genotyped six SNPs, PRODH*1195C-->T, PRODH*1482C-->T, PRODH*1483A-->G, PRODH*1766A-->G, PRODH*1852G-->A PRODH*1945T- >C, two of which (PRODH*1483A-->G and PRODH*1852G-->A) have not been previously reported. We found association with schizophrenia for two haplotypes consisting of PRODH*1945T-->C and PRODH*1852G-->A (Global P = 0.006), and PRODH*1852G-->A and PRODH*1766A-->G (Global P = 0.01) which include one of the newly identified markers. After six-fold Bonferroni correction for multiple testing the PRODH*1945T-C/PRODH*1852G-A haplotypes remained significant. This is a sub haplotype of the PRODH haplotype previously associated with schizophrenia and it also maps to the 3' region of the gene, indicating that this is the region most likely to contain the underlying risk alleles. Overall this finding supports a role for the PRODH locus in schizophrenia. PMID- 15274031 TI - Positive association of the human frizzled 3 (FZD3) gene haplotype with schizophrenia in Chinese Han population. AB - Frizzled 3 (FZD3) gene is located on chromosome 8p21, a region that has been implicated in schizophrenia in genetic linkage studies. The FZD3 is a transmembrane receptor required for Wnt signal transduction cascades that have been thought to be involved in producing the cytoarchitectural defects observed in schizophrenia. Previous work has showed a strong association between FZD3 locus and schizophrenia in family-based study. To confirm this issue further, we investigated a genetic association between four single nucleotide polymorphisms (SNPs) located in the FZD3 gene and schizophrenia by case-control study using polymerase chain reaction (PCR)-based restriction fragment length polymorphism (RFLP) in the Chinese Han population. Our studies showed the SNPs rs2323019 and rs880481 have significant differences in both genotype and allele frequencies between control subjects and schizophrenic patients (rs2323019: Allele A > G, chi2 = 6.7277, df = 1, P = 0.0095; Genotype, chi2 = 10.6583, df = 2, P = 0.0049; rs880481: Allele A > G, chi2 = 10.3945, df = 1, P = 0.0013; Genotype, chi2 = 16.8049, df = 2, P = 0.0002). In addition, we constructed three-locus haplotypes to test their association with schizophrenia. The globe chi-squared test for haplotype analysis showed a significant association (chi2 = 66.38, df = 7, P < 0.000001). These results suggested that the FZD3 gene might be involved in the predisposition to schizophrenia. PMID- 15274032 TI - Assessment of the frequency of the 22q11 deletion in Afrikaner schizophrenic patients. AB - A hemizygous deletion of the q11 band on chromosome 22 occurs in 1 of every 5,950 live births (0.017%). The deletion is mediated by low copy repeats (LCRs) flanking this locus. Presence of the deletion is associated with variable phenotypic expression, which can include distinctive facial dysmorphologies, congenital heart disease and learning disabilities. An unusually high percentage of individuals with this deletion (25-30%) have been described to develop schizophrenia or schizoaffective disorder. In previous studies, the prevalence of the 22q11 deletion in patients with schizophrenia was found to be approximately 2% in Caucasian adults and 6% in childhood-onset cases. Both these frequencies represent a dramatic increase from the prevalence of the deletion in the general population. In this study, we investigate the occurrence of the 22q11 deletion in an independent sample of schizophrenic patients of Afrikaner origin. We first ascertained a sample of 85 patients who meet full diagnostic criteria for schizophrenia for presence of two or more of the clinical features associated with presence of the 22q11 deletion. A group of six patients (7%) met these criteria. This group was subjected to fluorescent in situ hybridization (FISH) and presence of the 22q11 deletion was confirmed for two subjects. Our study therefore confirms the previously reported rate of 2% frequency of the 22q11 deletion in adult schizophrenic patients and provides a two-stage screening protocol to identify these patients. PMID- 15274033 TI - Regulator of G-protein signaling 4 (RGS4) gene is associated with schizophrenia in Irish high density families. AB - The regulator of the G-protein signaling 4 (RGS4) gene was shown to have a different expression pattern in schizophrenia patients in a microarray study. A family-based study subsequently implicated the association of this gene with schizophrenia. We replicated the study with our sample from the Irish Study of High Density Schizophrenia Families (ISHDSF). Single marker transmission disequilibrium tests (TDT) for the four core SNPs showed modest association for SNP 18 (using a narrow diagnostic approach with FBAT P = 0.044; with PDT P = 0.0073) and a trend for SNP 4 (with FBAT P = 0.1098; with PDT P = 0.0249). For SNP 1 and 7, alleles overtransmitted to affected subjects were the same as previously reported. Haplotype analyses suggested that haplotype G-G-G for SNP1-4 18, which is the most abundant haplotype (42.3%) in the Irish families, was associated with the disease (narrow diagnosis, FBAT P = 0.0061, PDT P = 0.0498). This was the same haplotype implicated in the original study. While P values were not corrected for multiple testing because of the clear prior hypothesis, these results could be interpreted as supporting evidence for the association between RGS4 and schizophrenia. PMID- 15274034 TI - New GABAA receptor alpha5 subunit gene polymorphism that may confound genotyping. AB - We report the discovery of a new GABAA receptor alpha5 subunit gene polymorphism close to the polymorphism described by Glatt et al. (GT)5GCGTGC(GT)21. This new polymorphism is of great importance, because it means that non-denaturing acrylamide gels used to separate the different alleles of the polymorphism described by Glatt et al. cannot distinguish an allele with the sequence: (GT)4GCGTGC(GT)n from another allele with the sequence: (GT)4(GCGT)4GC(GT)(n-6). These gel fragments are separated by size, which would be the same in these two cases. An alternative would be to use an analysis method that can detect base changes, for instance, single strand conformation polymorphism (SSCP) or denaturing gradient gel electrophoresis (DGGE). PMID- 15274035 TI - Lack of association between 5HT2A receptor gene haplotype, bipolar disorder and its clinical subtypes in a West European sample. AB - Bipolar affective disorder (BPAD) is a complex psychiatric disorder with a major genetic contribution. Abnormalities in serotonergic function have been implicated in its aetiology. The 5HT2A receptor (5HT2AR) gene is a strong candidate gene for involvement in BPAD, but previous association studies have reported conflicting results. These data are difficult to interpret because most negative results were obtained with small samples. The aim of this study was to test the association between the 5HT2AR gene and BPAD in a large West European sample. We studied the 1438G/A and the His452Tyr polymorphisms, for haplotype analysis to increase both informativity and the likelihood of detecting an association between BPAD and the 5HT2AR gene. We analysed the genotype, allele and haplotype distributions of two 5HT2AR gene variants in a population of 356 BPAD patients, which we compared with 208 healthy controls. We also carried out exploratory analysis in clinical subgroups of patients defined according to personal history of mood disorders, suicidal behaviour, comorbid psychiatric disorders and family history of affective disorders. We found no difference between BPAD patients and controls for allele, genotype and haplotype distributions. Exploratory analysis in subgroups of BPAD patients showed only a marginal difference in haplotype distribution between controls and BPAD patients with antidepressant-induced mania (P = 0.018). This difference was not significant after correction for multiple testing. Our study suggests that the 5HT2AR gene is unlikely to be involved in genetic susceptibility to BPAD but should be further investigated in a pharmacogenetic study. PMID- 15274036 TI - Non-replication of the brain-derived neurotrophic factor (BDNF) association in bipolar affective disorder: a Belgian patient-control study. AB - This patient-control association study was conducted to investigate a possible association of two single nucleotide polymorphisms (SNPs), g.11757C > G and g.196G > A, in the brain-derived neurotrophic factor (BDNF) with bipolar affective disorder (BPAD). Two hundred seventy-five individuals of Belgian origin (at least two generations of Belgian ancestors) were genotyped (112 BPAD and 163 controls). No significant differences were found in the frequency of genotypes and alleles of g.196G > A (P = 0.37 and 0.94, respectively) and g.11757C > G (P = 0.49 and 0.59, respectively) between controls and BPAD patients. An haplotype analysis revealed no difference between patients and controls (P = 0.44). We failed to replicate previous findings implicating BDNF in the aetiology of BPAD. However, BDNF remains an interesting target for future genetic studies and should be tested in prospective pharmacogenetic therapeutic trials. PMID- 15274037 TI - Further evidence of a combined effect of SERTPR and TPH on SSRIs response in mood disorders. AB - We reported an independent association of the short variant of the serotonin transporter gene-linked polymorphic region (SERTPR) and tryptophan hydroxylase (TPH) genes with antidepressant response to selective serotonin reuptake inhibitors (SSRIs). The aim of the present study was to confirm the effect of the SERTPR and TPH gene variants on the SSRIs antidepressant activity in a new sample of major and bipolar depressives. Two hundred and twenty one inpatients (major depressives = 128, bipolar disorder = 93) were treated with SSRIs (fluvoxamine or paroxetine) for 6 weeks; the severity of depressive symptoms was weekly assessed with the Hamilton Rating Scale for Depression (HAMD). SERTPR and TPH variants were determined using PCR-based techniques, 220 subjects genotyped for SERTPR and 221 for TPH that were never included in previous studies. SERTPR*s/s variant association with a poor response to SSRI treatment was confirmed, even if with less significant P values (P = 0.034), independently from clinical variables; pooling the present sample with previous ones we observed a highly significant effect (P < 0.000001). TPH*A/A variants showed higher HAMD scores throughout the trial but with only a trend in the same direction of our previous study in terms of a worse response of A/A genotypes. Thus, the previous positive association was not fully replicated for TPH. The present independent replication confirms SERTPR variants as a liability factor for antidepressant efficacy while the TPH effect is not unequivocal. PMID- 15274038 TI - Lack of relationship between CO2 reactivity and serotonin transporter gene regulatory region polymorphism in panic disorder. AB - Changes in the function of the serotonergic system influence both panic phobic symptoms and carbon dioxide (CO2) reactivity in patients with panic disorder. Schmidt et al. [2000: J Abnorm Psychol 109(2):308-320] recently reported a predictive role of the genetic variants of the 5-HTTLPR on the fearful response to CO2 in healthy controls. We tested the hypothesis that the heterogeneity of CO2 reactivity in patients with panic disorder could be related to the allelic variation of the 5-HTT promoter. Ninety-five patients with panic disorder were challenged with 35% CO2. 5-HTTLPR allelic variation in each subject was determined using a PCR-based method. There were no differences for all the measures of CO2 reactivity among the genotype groups. CO2 reactivity of patients with panic disorder seems not to be influenced by the genetic variants of the 5 HTTLPR; this finding does not support a role for the serotonin transporter in the etiopathogenesis of CO2 reactivity in panic disorder. PMID- 15274039 TI - A missense polymorphism (S205L) of the low-affinity neurotrophin receptor p75NTR gene is associated with depressive disorder and attempted suicide. AB - Several lines of evidence have implicated that neurotrophins play an important role in the pathophysiology of mood disorders. This study examined whether a common missense polymorphism (S205L) of a gene encoding the p75NTR, the low affinity receptor for neurotrophins, is associated with depressive disorder in a Japanese sample of 164 patients and the same number of controls matched for age and sex. There were significant differences in the genotype distribution and allele frequency between the cases and controls. The minor allele (L205) was significantly decreased in the patients than in the controls (P < 0.05, odds ratio 0.54, 95% CI 0.31-0.94), suggesting that this allele may have a protective effect against the development of major depression. Furthermore, this association was more strongly observed in the patients with a history of attempted suicide than those without such a history. Our results suggest that the S205L polymorphism of the p75NTR gene is involved in the pathogenesis of depressive disorder and suicidal behavior. PMID- 15274040 TI - Genome-wide linkage survey for genetic loci that affect the risk of suicide attempts in families with recurrent, early-onset, major depression. AB - We previously described the results of a genome-wide linkage survey for genetic loci that influenced the development of unipolar mood disorders in 81 families identified by individuals with Recurrent, Early-Onset, Major Depressive Disorder (RE-MDD) [Zubenko et al. 2003b; Am J Med Genet (Neuropsychiatr Genet) 123B:1-18]. In the current study, we extended this linkage analysis by including the history of a suicide attempt as a covariate to identify chromosomal regions that harbor genes that influence the risk of this behavior in the context of mood disorders. This approach identified six linkage peaks with maximum multipoint DeltaLOD scores that reached genome-wide adjusted levels of significance (2p, 5q, 6q, 8p, 11q, and Xq). Four of these (2p, 6q, 8p, and Xq) exceeded the criterion for "highly-significant linkage" (genome-wide adjusted P < 0.001) recommended by Lander and Kruglyak [1995; Nat Genet 11:241-246]. The strongest evidence for linkage was observed in analyses employing affected relative pairs (ARPs) with the most severe and disabling Mood Disorders: Depression Spectrum Disorder and RE MDD. The highest DeltaLOD score that emerged from this linkage analysis, 5.08, occurred for ARPs with Depression Spectrum Disorder at D8S1145 (37.0 cM, 18.2 Mbps, P < 0.0001) at cytogenetic location 8p22-p21. Significant linkage results on Xq arose from analyses of ARPs with RE-MDD at DXS1047 (143 cM, 127.8 Mbps, DeltaLOD = 3.87, P < 0.0001), a finding that may contribute to the higher rate of suicide attempts among women than men. These findings provide evidence for suicide risk loci that are independent of susceptibility loci for Mood Disorders, and suggest that the capacity for suicide risk loci to affect the development of suicidal behavior depends on the psychiatric disorder or subtype with which they interact. PMID- 15274041 TI - The dysbindin gene in major depression: an association study. AB - The pathophysiological mechanisms, as well as the molecular loci of antidepressant drug action have not yet been established, but recent models proposed that several adaptive mechanisms in signal transduction cascades beyond the receptor and reuptake systems are involved in antidepressant action and play an important role in the etiology of affective disorders. In this context, the dysbindin gene (dystrobrevin-binding-protein 1, DTNBP1), which was recently reported to be associated with schizophrenia seems to be an interesting candidate gene for affective disorders. Dysbindin is widely expressed in the human brain and binds to the dystrophin-associated protein complex (DPC) which appears to be involved in signal transduction pathways, which have been repeatedly investigated and described as altered or disturbed in affective disorders [McLeod et al. [2003: Psychopharmacol Bull 35:24-41]; Brambilla et al. [2003: Mol Psychiatry 8:721-737]]. Therefore, we investigated whether five SNPs in the dysbindin gene could be susceptibility factors in the ethiology of major depression or for the response to antidepressant treatment in a sample of 293 patients compared to 220 healthy controls. Applying single SNP evaluation, as well as haplotype analysis we could not detect an association between the dysbindin polymorphisms and major depression or the response to antidepressant treatment. In conclusion, our results suggest that SNPs in the dysbindin gene are unlikely to play a major role in the pathophysiology of major depression or are in linkage disequilibrium (LD) with a neighboring mutation or gene. Further analysis are needed to confirm these results. PMID- 15274042 TI - Do aggressive and non-aggressive antisocial behaviors in adolescents result from the same genetic and environmental effects? AB - Antisocial behavior (ASB) in adolescents can broadly be separated into two forms; aggressive and non-aggressive. Both are heritable and it has been suggested that aggressive ASB is more heritable. The extent to which genes contribute to the correlation between the two is unknown. Structural equation modeling was applied to a population-based twin sample of 258 twins pairs aged 11-18 to estimate the heritability of each form of ASB and to estimate the extent to which the phenotypic correlation was the consequence of shared genes and environmental factors. Non-shared environment and genetic factors substantially influenced both forms of ASB. The heritability of aggressive (but not non-aggressive) ASB was significantly higher in girls than in boys. Combining both sexes, a model in which the genetic effects on aggressive and non-aggressive ASB were identical could be rejected. Our results suggest a partial genetic overlap with a specific genetic effect contributing to the variance of aggressive ASB and a stronger genetic effect on aggression in females than in males. PMID- 15274043 TI - Myelin oligodendrocyte glycoprotein (MOG) gene is associated with obsessive compulsive disorder. AB - Obsessive-compulsive disorder (OCD) is a severe neuropsychiatric disorder with a strong genetic component, and may involve autoimmune processes. Support for this latter hypothesis comes from the identification of a subgroup of children, described by the term pediatric autoimmune neuropsychiatric disorder associated with streptococcal infections (PANDAS), with onset of OCD symptoms following streptococcal infections. Genes involved in immune response therefore represent possible candidate genes for OCD, including the myelin oligodendrocyte glycoprotein (MOG) gene, which plays an important role in mediating the complement cascade in the immune system. Four polymorphisms in the MOG gene, a dinucleotide CA repeat (MOG2), a tetranucleotide TAAA repeat (MOG4), and 2 intronic single nucleotide polymorphisms, C1334T and C10991T, were investigated for the possibility of association with OCD using 160 nuclear families with an OCD proband. We examined the transmission of alleles of these four polymorphisms with the transmission disequilibrium test (TDT). A biased transmission of the 459 bp allele (allele 2: chi2 = 5.255, P = 0.022) of MOG4 was detected, while MOG2, C1334T, and C10991T showed no statistically significant bias in the transmission of alleles. The transmission of the C1334T.MOG2.C10991T.MOG4 haplotype 1.13.2.2 (chi2 = 6.426, P = 0.011) was also significant. Quantitative analysis using the family-based association test (FBAT) was significant for MOG4 in total Yale-Brown Obsessive-Compulsive Scale severity score (allele 2: z = 2.334, P = 0.020). Further investigations combining genetic, pathological, and pharmacological strategies, are warranted. PMID- 15274044 TI - Variants of the orexin2/hcrt2 receptor gene identified in patients with excessive daytime sleepiness and patients with Tourette's syndrome comorbidity. AB - The orexin-2/hypocretin-2 (OX2R) receptor gene is mutated in canine narcolepsy and disruption of the prepro-orexin/hypocretin ligand gene results in both an animal model of narcolepsy and sporadic cases of the human disease. This evidence suggests that the structure of the OX2R gene, and its homologue, the OX1R gene, both members of the G protein-coupled receptor (GPCR) family, and the gene encoding the peptide ligands, the prepro-orexin/hypocretin gene, may be variables in the etiology of sleep disorders. We report a single stranded conformational polymorphism (SSCP) analysis of the coding regions of these genes in idiopathic sleep disorder patients diagnosed with excessive daytime sleepiness (EDS) (n = 28), narcolepsy (n = 28), Tourette's syndrome/chronic vocal or motor tic disorder (n = 70), and control subjects (n = 110). Two EDS patients showed a Pro11Thr change. One Tourette's syndrome patient was found to have a Pro10Ser alteration. The Pro10Ser and Pro11Thr variants were not found in non-disease populations. Analysis of the ability of the mutant receptors to mobilize calcium compared to the wild-type receptor in response to orexin agonists indicated that they resulted in decreased potency at high (etaM) concentrations of orexin ligands. Further work is warranted to study the variability of the orexin/hypocretin system in a variety of disorders characterized by EDS. PMID- 15274045 TI - Familial symptom domains in monozygotic siblings with autism. AB - Autism is characterized by a triad of symptom domains (impaired social interaction, communication deficits, and repetitive behaviors) that vary significantly in their clinical presentation across the population. Within families with more than one affected member, however, discrepant findings exist with regard to symptom variability. Reduced intrafamily variance is of particular importance because it supports an underlying model of genetic heterogeneity in the transmission of autism, and the identification of familial clinical subtypes can be used to select more homogeneous samples for linkage analysis in the future. This study examines whether there are specific features of autism that show decreased variance within 16 families with monozygotic siblings concordant for autism. Evidence for familiality was defined as significantly decreased variance of symptom levels within monozygotic siblingships as compared to between siblingships. Using regression analysis, we demonstrated significant aggregation of symptoms within monozygotic siblingships for two of the three main symptom domains in autism: impairments in communication and social interaction showed significant familiality. Within the repetitive behavior domain, only the categories of circumscribed interests and preoccupation with part-objects showed reduced variance within siblingships. In addition, with the exception of a negative association between the social and behavior domains, partial correlation coefficients did not reveal significant associations between the levels of different symptom domains within families, suggesting that the levels of clinical features seen in autism may be a result of mainly independent genetic traits. Because of presumed genetic heterogeneity and the wide clinical variation seen in autism and other pervasive developmental disorders, selecting probands according to specific features known to show reduced variance within families may provide more homogeneous samples for genetic analysis and strengthen the power to detect the specific genes involved in autism. PMID- 15274046 TI - Mutation screening of X-chromosomal neuroligin genes: no mutations in 196 autism probands. AB - Autism, a childhood neuropsychiatric disorder with a strong genetic component, is currently the focus of considerable attention within the field of human genetics as well many other medical-related disciplines. A recent study has implicated two X-chromosomal neuroligin genes, NLGN3 and NLGN4, as having an etiological role in autism, having identified a frameshift mutation in one gene and a substitution mutation in the other, segregating in multiplex autism spectrum families (Jamain et al. [2003: Nat Genet 34:27-29]). The function of neuroligin as a trigger for synapse formation would suggest that such mutations would likely result in some form of pathological manifestation. Our own study, screening a larger sample of 196 autism probands, failed to identify any mutations that would affect the coding regions of these genes. Our findings suggest that mutations in these two genes are infrequent in autism. PMID- 15274047 TI - Genetic disorders affecting white matter in the pediatric age. AB - Pediatric white matter disorders can be distinguished into well-defined leukoencephalopathies, and undefined leukoencephalopathies. The first category may be subdivided into: (a) hypomyelinating disorders; (b) dysmyelinating disorders; (c) leukodystrophies; (d) disorders related to cystic degeneration of myelin; and (e) disorders secondary to axonal damage. The second category, representing up to 50% of leukoencephalopathies in childhood, requires a multidisciplinar approach in order to define novel homogeneous subgroups of patients, possibly representing "new genetic disorders" (such as megalencephalic leukoencepahlopathy with subcortical cysts and vanishing white matter disease that have recently been identified). In the majority of cases, pediatric white matter disorders are inherited diseases. An integrated description of the clinical, neuroimaging and pathophysiological features is crucial for categorizing myelin disorders and better understanding their genetic basis. A review of the genetic disorders affecting white matter in the pediatric age, including some novel entities, is provided. PMID- 15274048 TI - Using nonword repetition to distinguish genetic and environmental influences on early literacy development: a study of 6-year-old twins. AB - This study considered whether cognitive profile could distinguish groups of children where genes or environment played a major role in influencing reading level. Same-sex twin pairs from an epidemiological study were categorized according to parental report at 4 years of age into those with low language skills and a typically developing group. A total of 132 same-sex twin pairs from the low language group and 66 from the control group were assessed at 6 years of age, to investigate heritability of reading ability adjusted for nonverbal IQ. For pairs where both twins had normal scores on a nonword repetition test, heritability was zero, with environmental influences explaining all the variance. For pairs where one or both twins had low nonword repetition, the heritability estimate was 0.79 and the variance due to shared environment was zero. Future studies of genetics of reading development should treat those with poor nonword repetition skills as a separate subgroup. PMID- 15274049 TI - Association analysis of two candidate phospholipase genes that map to the chromosome 15q15.1-15.3 region associated with reading disability. AB - Molecular genetic studies have suggested a reading disability (RD, dyslexia) susceptibility locus on chromosome 15q. We have previously mapped this locus by association to the region surrounding D15S994. Very little is known about the neurobiological processes involved in RD, and therefore selecting positional candidate genes for analysis based upon function is difficult. Nevertheless we were able to identify two functional candidates based upon existing hypotheses. Both were phospholipase genes, phospholipase C beta 2 (PLCB2) and phospholipase A2, group IVB (cytosolic; PLA2G4B). D15S944 is located within PLCB2 and is 1.6 Mb from PLA2G4B. We examined each gene for association using a mixed direct and indirect association approach, a case (n = 164)/control (n = 174) sample, and a partially overlapping sample of 178 RD parent-proband trios from South Wales and England. Mutation analysis revealed 14 sequence variants in PLCB2 and 33 variants in PLA2G4B. All non-synonymous SNPs were genotyped as were SNPs across each gene with maximum distance between SNPs of 6 kb. Case-control analyses revealed modest evidence (0.01 < P < 0.05) for association between a single variant in PLCB2 and two variants in PLA2G4B. However, association was not confirmed in the family based sample. As the latter sample has previously generated replicated significant evidence for association between RD and markers/haplotypes surrounding D15S944, it should have sufficient power to detect association to variants in susceptibility gene itself. We conclude that neither gene accounts for the association signal we previously observed. As these are the only clear cut functional candidate genes in the region, identification of the putative susceptibility locus for RD on 15q will require more methodical non-hypothesis driven positional cloning approaches. PMID- 15274050 TI - Allelic and haplotypic association of GABRA2 with alcohol dependence. AB - Alcohol dependence is a highly prevalent disorder that is associated with serious morbidity and mortality. Because the GABAA neurotransmitter receptor is an important mediator for several behavioral effects of alcohol, genes encoding GABA related proteins are functional candidates to influence risk of alcohol dependence. Two genome-wide scans showed linkage of alcohol dependence to a region on chromosome 4p, which contains a cluster of genes encoding GABAA receptor subunits. A recent effort to fine map that region showed a haplotypic association of alcohol dependence to the gene encoding the GABAA receptor alpha-2 subunit (GABRA2). We examined 10 single nucleotide polymorphisms (SNPs) spanning the coding region of this gene in samples of European American subjects with alcohol dependence (n = 446), and controls (n = 334) screened to exclude substance use disorders. There was evidence of association to alcohol dependence for seven adjacent markers spanning 98,000 bp in the middle and 3'-portion of the GABRA2 gene (range of P-values = 0.008-0.03). When the subset of the alcohol dependent subjects excluding those with a diagnosis of cocaine or opioid dependence or major depressive episode (n = 198) was examined, the strength of the association was increased across these 7 SNPs (range of P-values = 0.002 0.007). Two common haplotypes in this region accounted for 90.8% of chromosomes. The more common haplotype was present in 55.6% of control group chromosomes versus 48.2% of alcohol-dependent subjects (P = 0.007) and 45.8% of subjects with alcohol dependence but no co-morbid drug dependence or depression (P = 0.003). These findings replicate and extend recently reported findings, which together underscore the potential contribution of polymorphic variation at the GABRA2 locus to the risk for alcohol dependence. PMID- 15274052 TI - Association study of the Epac gene and tobacco smoking and nicotine dependence. AB - Exchange protein directly activated by cAMP (Epac) has been shown to increase its expression in rat prefrontal cortex after self-administration of nicotine. [Konu et al. 2001: Brain Res 909:194-203]. We investigated the role of the human ortholog of Epac in tobacco smoking and nicotine dependence (ND). Using a case control design, we typed 5 SNPs in this gene in 688 subjects, of whom 244 were non-smokers, 215 were Low-ND smokers, and 229 were High-ND smokers. We tested allele and genotype association to smoking initiation (SI) and progression to ND. Three of the five typed SNPs showed modest allele association with progression to ND. Weak association with SI was also observed for one SNP. Considering the function of the gene in cellular signal transduction pathway, its elevated expression after nicotine self-administration, and multiple markers association with both SI and progression to ND, it is plausible to suggest that variants in Epac contribute to the liability to ND. PMID- 15274051 TI - Genomic screen for loci associated with alcohol dependence in Mission Indians. AB - Alcohol dependence is a leading cause of morbidity and mortality in Native Americans, yet biological factors underlying the disorder in this ethnic group remain elusive. This study's aims were to map susceptibility loci for DSM-III-R alcohol dependence and two narrower alcohol-related phenotypes in Mission Indian families. Each participant gave a blood sample and completed an interview using the Semi-Structured Assessment for the Genetics of Alcoholism (SSAGA) that was used to make alcohol dependence diagnoses and the narrower phenotypes of withdrawal, and drinking severity. Genotypes were determined for a panel 791 microsatellite polymorphisms. Analyses of multipoint variance component LOD scores for the dichotomous DSM-III-R phenotype revealed no peak LOD scores that exceeded 2.0 at any chromosome location. Two chromosomes, 4 and 12, had peak LOD scores that exceeded 2 for the alcohol use severity phenotype and three chromosomes 6, 15, 16 were found to have peaks with LOD scores that exceeded 2 for the withdrawal phenotype. Evidence for linkage to chromosomes 4 and 15, and 16 have been reported previously for alcohol related phenotypes whereas no evidence has as yet been reported for chromosomes 6 and 12. Combined linkage and association analysis suggest that alcohol dehydrogenase 1B gene polymorphisms are partially responsible for the linkage result on chromosome 4 in this population. These results corroborate the importance of several chromosomal regions highlighted in prior segregation studies in alcoholism and further identify new regions of the genome that may be unique to either the restricted phenotypes evaluated or this population of Mission Indians. PMID- 15274053 TI - Association analysis of the DRD4 and COMT genes in methamphetamine abuse. AB - We analyzed two polymorphisms in genes encoding proteins of the dopamine system, the Val158Met polymorphism in the catechol-O-methyltransferase gene and the 120 bp VNTR polymorphism in the promoter of the dopamine D4 receptor gene for association with methamphetamine abuse. We used a case/control design with 416 methamphetamine abusing subjects and 435 normal controls. All subjects were Han Chinese from Taiwan. We found an excess of the high activity Val158 allele in the methamphetamine abuser group, consistent with several previous reports of association of this allele with drug abuse. The 120-bp VNTR polymorphism in the promoter of the dopamine D4 receptor gene itself did not show significant association with methamphetamine abuse. However, analysis of the 120-bp VNTR polymorphism and the exon 3 VNTR in the dopamine D4 receptor as a haplotype showed significant association with methamphetamine abuse, which gave an empirical P value 0.0034 for a heterogeneity model. Moreover, there were significant interactive effects between polymorphisms in the catechol-O methyltransferase and dopamine D4 genes. The evidence of interaction between COMT 158 Val/Met and DRD4 48-bp VNTR polymorphisms (P = 0.0003, OR = 1.45, 95% CI: 1.148-1.77), and between COMT 158 Val/Met and DRD4 120 bp promoter polymorphisms (P = 0.01, OR = 1.10, 95% CI: 1.10-1.18) were significant but the latter was weak. We conclude that genetic variation in the dopamine system may encode an additive effect on risk of becoming a methamphetamine abuser. PMID- 15274054 TI - Forms of cannabis and cocaine: a twin study. AB - Cannabis and cocaine are illicit psychoactive substances that have fallen under intense scrutiny by epidemiologists and behavioral geneticists. However, most analyses have used a composite variable to represent the use of these two drugs. For example, the composite variable of cannabis use often includes use of marijuana or hashish. Similarly, cocaine use involves different preparations (crack vs. cocaine hydrochloride) and varying routes of administration (intranasal insufflation vs. smoking). While there is some epidemiological evidence for the difference in addictive potentials between crack and intranasal cocaine, genetically informative studies have not examined the relationship between the forms of cannabis or cocaine. We used data from male and female same sex twin pairs to examine the extent of genetic, shared environmental, and unique environmental overlap between (i) marijuana and hashish for cannabis use and (ii) intranasal and crack cocaine for cocaine use. Bivariate Cholesky models were fit using the structural equation modeling software Mx. Our results indicate that for both drugs, the individual drug forms show a complete overlap of genetic factors and a substantial overlap of shared environmental influences. While marijuana and hashish share a moderate proportion of their unique environment, crack and intranasal cocaine only show a modest overlap of unique environmental factors, adding some evidence for form-specific environmental factors. In conclusion, there is substantial overlap of familial factors between forms of a single drug and preference is primarily determined by unique environmental influences. These findings also reinforce the validity of composite variables in epidemiological and genetic research. PMID- 15274056 TI - Biochemotherapy for melanoma: rational therapeutics in the search for weapons of melanoma destruction. PMID- 15274057 TI - The risk of venous thromboembolic disease associated with adjuvant hormone therapy for breast carcinoma: a systematic review. AB - BACKGROUND: Tamoxifen therapy for patients with breast carcinoma is perceived as an independent risk factor for venous thromboembolic events (VTE), but the risk associated with other adjuvant therapies is less well recognized. METHODS: The authors conducted a computerized PubMed literature search for English-language articles published between January 1966 and December 2003. Studies were analyzed with regard to trial design, breast carcinoma staging, adjuvant agent, definition of VTE outcomes, method of VTE case ascertainment, and the presence of concomitant VTE risk factors. RESULTS: Accurate determination of VTE rates was impaired by the universal lack of routine assessments for asymptomatic VTE. Therefore, only the risk of symptomatic VTE could be derived. The risk of VTE was increased twofold to threefold during tamoxifen or raloxifene use for breast carcinoma chemoprevention. It remains unknown whether the risk is increased further in women with inherited hypercoagulable states. In the setting of early stage breast carcinoma, the risk of VTE is increased both with tamoxifen use and anastrozole use. Such risk appeared to be lower, albeit not negligible, with anastrozole. Significant methodologic limitations of all available studies in women with advanced-stage breast carcinoma precluded determination of the true VTE risk associated with different adjuvant hormonal agents and made it nearly impossible to compare the risk between different drugs. CONCLUSIONS: All agents used for breast carcinoma chemoprevention and adjuvant therapy appear to increase the risk of VTE. Available data were insufficient to support any assumptions that newer hormonal forms of hormone manipulation are safer than tamoxifen in women with advanced breast carcinoma. PMID- 15274058 TI - Breast cancer survivorship in a multiethnic sample: challenges in recruitment and measurement. AB - BACKGROUND: The inclusion of ethnic minorities in cancer-related studies continues to be an important concern for researchers. In this article, the authors present 1) a brief discussion of recruitment and measurement challenges in conducting multiethnic survivorship research, and 2) recruitment outcomes and sample characteristics for a health-related quality-of-life study with a multiethnic sample of breast cancer survivors (BCS). METHODS: A case-control, cross-sectional design with mixed sampling methods was used. The Contextual Model for Recruitment and Enrollment of Diverse Samples was used to guide the protocol. BCS were recruited from the California Cancer Surveillance Program, from hospital registries, and from community agencies. Participation rates, demographic factors, and medical factors were compared. The reliability of standard measures by ethnicity was assessed. RESULTS: Seven hundred three women participated, including 135 African-American women (19%), 206 Asian-American women (29%), 183 Latino-American women (26%), and 179 European-American women (26%). Participation was influenced by ethnicity, age, and site of recruitment. Overall, African Americans were least likely to participate, and European Americans most likely to participate. African Americans and Asian Americans were more likely to refuse, European Americans and Latino Americans were more likely to agree to participate, and European Americans and Asian Americans were most likely to complete the survey after consenting. Measures possessed moderate to excellent reliability (0.64-0.91). CONCLUSIONS: Despite important recruitment and measurement challenges, this study obtained acceptable participation rates and good internal consistency of the measures. The results demonstrate the utility of a culturally responsive approach to health disparities research. PMID- 15274059 TI - 'Chemobrain' in breast carcinoma?: a prologue. AB - BACKGROUND: Chemotherapy-induced cognitive dysfunction in patients with breast carcinoma has been described previously. However, those studies only assessed patients' postchemotherapy cognitive functioning and were not able to determine the relation between cognitive function and other treatments, such as surgery and radiotherapy, that often precede systemic chemotherapy. METHODS: Eighty-four women with breast carcinoma underwent a comprehensive neuropsychologic evaluation before receiving adjuvant therapy for nonmetastatic primary breast carcinoma. RESULTS: Before the start of systemic therapy, 35% of women in the current cohort exhibited cognitive impairment. Verbal learning (18%) and memory function (25%) were impaired significantly more frequently relative to normative expectations. Although the impairments were not significant in the women who were examined, nonverbal memory (17%), psychomotor processing speed and attention (13%), confrontational naming (13%), visuoconstruction (13%), and upper-extremity fine motor dexterity (12%) were impaired more frequently than was expected. Affective distress was related significantly to cognitive impairment (Pearson chi-square = 9.90; P = 0.002). Given the conservative statistical approach employed, extent of surgery, hormone replacement therapy history, and current menopausal status failed to achieve statistical significance, but these variables did exhibit provocative trends with respect to cognitive impairment. CONCLUSIONS: Cognitive impairment frequently is observed before the administration of systemic chemotherapy. Thus, investigations purporting to measure chemotherapy-induced cognitive dysfunction must employ study designs that incorporate prechemotherapy baseline assessments to accurately detect changes in cognitive function that are attributable to chemotherapy. PMID- 15274060 TI - The management of early breast carcinoma before and after the introduction of clinical practice guidelines. AB - BACKGROUND: Clinical practice guidelines for the management of breast carcinoma were introduced in Australia in October, 1995. This article describes the management of early-stage breast carcinoma both before and after the introduction of these guidelines. METHODS: All cases of early breast carcinoma that were diagnosed over the same 6-month period in 1995 and 1999 and registered with a state-based cancer registry were identified. Treating surgeons completed a survey assessing tumor characteristics and primary treatment. In 1995, 95% of 188 surgeons who were approached participated and 96% of the surveys were returned. In 1999, 92% of 159 surgeons who were approached participated and 91% of the surveys were returned. Analyses are based on 1066 cases from 1995 and 1001 cases from 1999. RESULTS: The pathologic disease stage of the patients was similar in both study years. The proportion of patients who underwent breast-conserving therapy (BCT) increased from 54% in 1995 to 69% in 1999. This increase was noted across most levels of disease characteristics but was not evident among those patients treated by the least active surgeons. The proportion of patients treated with BCT who received radiotherapy increased from 59% in 1995 to 80% in 1999. This trend was observed across most levels of tumor characteristics and surgeon caseload. The proportion of women with receptor-positive tumors who were treated with endocrine therapy increased, whereas the proportion of patients with receptor-negative tumors who received this therapy decreased from 39% in 1995 to 17% in 1999. CONCLUSIONS: The management of early breast carcinoma in the state of Victoria appeared to change between 1995 and 1999 in the direction expected if the national guidelines had been incorporated into the practice patterns of surgeons treating breast carcinoma patients. PMID- 15274061 TI - Complex of urokinase-type plasminogen activator with its type 1 inhibitor predicts poor outcome in 576 patients with lymph node-negative breast carcinoma. AB - BACKGROUND: The ability of a solid tumor to grow and metastasize has a significant dependence on protease systems, such as the plasminogen activation system. The plasminogen activation system includes the urokinase-type plasminogen activator (uPA) and plasminogen activator inhibitor type 1 (PAI-1), among other molecules. Both uPA and PAI-1 are established prognostic factors for patients with breast carcinoma. In the current study, the authors investigated whether the complex of uPA with PAI-1 is also associated with the natural course of this malignancy. METHODS: Cytosolic levels of uPA, PAI-1, and the uPA:PAI-1 complex were measured in tumor tissue from 576 patients with lymph node-negative invasive breast carcinoma using quantitative enzyme-linked immunosorbent assays. Patients did not receive adjuvant systemic therapy, and the median follow-up duration was 61 months (range, 2-187 months) after primary diagnosis. Correlations with well known clinicopathologic factors were assessed, and univariate and multivariate survival analyses were performed. RESULTS: uPA:PAI-1 complex levels were positively associated with adverse histologic grade and inversely correlated with estrogen and progesterone receptor status. On univariate analysis, increased levels of the uPA:PAI-1 complex were found to be associated with reduced recurrence-free survival (RFS) and overall survival (OS) rates. On multivariate analysis, uPA:PAI-1 complex levels were found to be an independent predictor of OS (P = 0.039), but not RFS (P = 0.240). When uPA and PAI-1 levels were not included in the multivariate analysis, uPA:PAI-1 complex levels became a significant predictor of both RFS and OS (P = 0.029 and P = 0.007, respectively). CONCLUSIONS: The results of the current study demonstrate that uPA:PAI-1 complex levels have prognostic value on univariate analysis. In addition, increased uPA:PAI-1 complex levels were significantly associated with poor OS on multivariate analysis. Increased uPA:PAI-1 complex levels were also significantly associated with reduced RFS rates after the exclusion of uPA and PAI-1 levels from the multivariate analysis model. PMID- 15274062 TI - Mammographic screening: patterns of use and estimated impact on breast carcinoma survival. AB - BACKGROUND: Although many studies support the life-saving potential of screening mammography, the actual utilization of screening and the impact of the actual pattern of screening use on the breast carcinoma death rate, remain incompletely understood. In the current report, the authors describe patterns of screening use among women who were examined at a large screening and diagnostic service and estimate the added mortality associated with missed screening mammograms. METHODS: Mammography use was assessed in a population of 72,417 women who received a total of 254,818 screening mammograms at the Massachusetts General Hospital (MGH) Avon Comprehensive Breast Center (Boston, MA) between January 1, 1985, and February 19, 2002. A computer simulation of breast carcinoma growth, spread, and detection of breast carcinoma was used to estimate the likely health consequences of various types of screening use. RESULTS: Both prompt return for annual screening and full use of screening over extended periods of time were rare, and comparison of the MGH population with other populations revealed that the low level of use observed in the MGH population was not atypical. Only 6% of women who received a mammogram in 1992 received all annual mammograms that were available over the next 10 years; the mean number of mammograms received during this period was 5.06, or 51% of the number recommended by the American Cancer Society. Computer simulation results indicate that this underutilization of screening should result in higher mortality levels. Women from traditionally underserved socioeconomic, racial, and ethnic groups, women without insurance, and women who did not speak English had lower levels of use compared with other women. Lower levels of use also were observed among women receiving their first mammogram or who in the past had not returned promptly. Women ages 55-65 years had higher levels of use than did younger or older women. Women who previously had breast carcinoma also had higher levels of screening use. Nonetheless, none of the subpopulations of women stratified by age, race, ethnicity, zip code, income,language, insurance, status, previous screening use, or medical history exhibited a widespread propensity to promptly return for annual screening over an extended period of time. CONCLUSIONS: By many measures, the current analysis is one of the most detailed descriptions of screening use to date. The authors observed a level of screening use that was disappointingly low, with potentially negative health-related consequences, among women across categories defined by racial, ethnic, socioeconomic, and geographic characteristics; insurance status; language; age; medical history; and previous screening use. Improvements in the promptness with which women return to screening appear to have the potential to lead to considerable reductions in breast carcinoma death. PMID- 15274063 TI - Feasibility of breast-conserving surgery for patients with breast carcinoma associated with nipple discharge. AB - BACKGROUND: Few studies in the literature address the surgical management of patients with breast carcinoma who present with associated nipple discharge. The purpose of the current study was to determine the feasibility of breast conserving surgery (BCS) for these patients. METHODS: The medical records of patients who presented with pathologic nipple discharge and underwent diagnostic or curative surgery between January 1990 and December 2002 were retrospectively reviewed. RESULTS: A total of 188 patients presented with nipple discharge during the study period. Of those, 47 had breast carcinoma. One patient had metachronous bilateral nipple discharge associated with malignant disease. Therefore, medical records associated with a total of 48 cases were reviewed. The median patient age was 52 years (range, 29-87 years), and the median follow-up duration was 45 months (range, 6-109 months). Twenty-nine patients had ductal carcinoma in situ (DCIS), 14 had Stage I disease, 3 had Stage II disease, and 2 had Stage III disease. Twenty-four patients were ultimately treated with mastectomy. For 16 of these patients, mastectomy was required because extensive disease was found in reexcisional segmental mastectomy specimens. Among patients with Stage 0 or I disease, the incidence of occult nipple-areola complex (NAC) involvement was 16% (3 of 19 patients). Twenty-four patients were ultimately treated with BCS with (n = 13) or without (n = 11) adjuvant radiotherapy. Local disease recurrence was noted at 14, 28, and 40 months, respectively, in 3 patients who declined adjuvant radiotherapy after BCS for DCIS. Among patients treated with BCS, comedonecrosis, multifocality, and the absence of adjuvant radiotherapy were associated with decreased local recurrence-free survival (P = 0.0005, P = 0.045, and P = 0.013, respectively). However, disease-free survival (mean +/- standard error) was similar for patients who underwent mastectomy and patients who underwent BCS (90 +/- 6 months; 95% confidence interval [CI], 78-101 months vs. 82 +/- 6 months; 95% CI, 69-94 months; P = 0.528). One patient with Stage I disease died of distant metastases at 99 months. CONCLUSIONS: Patients with breast carcinoma accompanied by nipple discharge presented primarily with early-stage breast carcinoma associated with DCIS. Occult NAC involvement was not an uncommon finding in patients with early-stage breast carcinoma. Nonetheless, BCS can be performed safely if negative margins are achieved and if appropriate adjuvant radiotherapy or systemic therapy is administered. PMID- 15274064 TI - Adenocarcinoma of the small bowel: presentation, prognostic factors, and outcome of 217 patients. AB - BACKGROUND: Primary adenocarcinoma of the small bowel is a rare neoplasm, and to the authors' knowledge, few studies to date have addressed the topic. METHODS: In the current study, the records of 217 patients with small bowel adenocarcinoma were reviewed retrospectively for the presentation, prognostic factors, treatment modalities, and outcome. RESULTS: The median age of the patients was 55 years and there were 133 (61%) males. Tumors originated in the duodenum in 113 (52%) patients, the jejunum in 54 (25%) patients, the ileum in 28 (13%) patients, and in nonspecified sites in 22 (10%) patients. Patients with proximal tumors were diagnosed for the most part using endoscopy (i.e., 46 of 108 [43%]), whereas laparotomy enabled diagnosis in 16 of 28 (57%) patients with distal tumors. Based on TNM staging, 9 (4%) patients had Stage I disease, 43 (20%) patients had Stage II disease, 86 (39%) patients had Stage III disease, and 75 (35%) patients had Stage IV disease. The liver was the most common site of metastasis in 44 (59%) patients. Cancer-directed surgery was performed in 146 (67 %) patients, including the Whipple procedure in 36 patients (17%). The median overall survival time was 20 months. The 5-year overall survival rate was 26%. Cancer-directed surgery, early-stage disease, and lymph node involvement ratio were significantly associated with overall survival by univariate analysis. However, only cancer directed surgery and lymph node involvement ratio were independent predictors of overall survival in a multivariate analysis (adjusted rate ratio = 0.14; 95% confidence interval [95% CI], 0.04-0.46; P = 0.001 and adjusted rate ratio = 0.25; 95% CI, 0.12-0.53; P < 0.001, respectively). CONCLUSIONS: Performing an oncologic surgery resulted in the best outcome in patients with nonmetastatic disease. Because cancer-directed surgery is associated with high morbidity and mortality in primary centers, these patients should be referred to a tertiary center for adequate treatment. PMID- 15274065 TI - Prostate needle biopsies: multiple variables are predictive of final tumor volume in radical prostatectomy specimens. AB - BACKGROUND: Tumor volume is one of the most powerful predictors of patient outcome in prostatic adenocarcinoma. It is uncertain as to which preoperative variables are most predictive of final tumor volume at radical prostatectomy, especially among patients who have had positive biopsies at multiple biopsy sites. The current study attempted to identify the biopsy variables that are most predictive of final tumor volume. METHODS: The authors examined prostate biopsy specimens from 151 consecutive patients with at least 2 positive biopsy sites. The following data were collected: highest percentage of adenocarcinoma at any biopsy site, percentage of adenocarcinoma at the biopsy site with the highest Gleason score, highest percentage of cores positive for adenocarcinoma at any biopsy site, percentage of positive cores with carcinoma at the site with the highest Gleason score, number of positive sites, tumor bilaterality, and percentage of biopsy sites positive for disease. All patients underwent radical prostatectomy. The prostatectomy specimens were entirely embedded and whole mounted. Tumor volume was measured using the grid method. Logarithmic transformation was applied to tumor volumes for the purposes of the analysis. RESULTS: Highest percentage of adenocarcinoma at any biopsy site (P = 0.012), percentage of adenocarcinoma at the biopsy site with the highest Gleason score (P = 0.021), number of positive biopsy sites (P = 0.026), tumor bilaterality (P = 0.008), and percentage of biopsy sites positive for disease (P = 0.0001) all were significant predictors of tumor volume on linear regression analysis. Highest percentage of cores positive for adenocarcinoma (P = 0.081) and percentage of positive cores with carcinoma at the site with the highest Gleason score (P = 0.240) were not significant predictors of tumor volume. Based on the model F statistic, percentage of biopsy sites positive for tumor, tumor bilaterality, and highest percentage of adenocarcinoma at any biopsy site were the variables that were most predictive of tumor volume. CONCLUSIONS: Highest percentage of adenocarcinoma at any biopsy site, percentage of adenocarcinoma at the biopsy site with the highest Gleason score, number of positive biopsy sites, tumor bilaterality, and percentage of biopsy sites positive for disease all are useful preoperative predictors of tumor volume in radical prostatectomy specimens. Although these preoperative biopsy parameters were significant in linear regression models, none was sufficient as a single predictor of tumor volume. PMID- 15274066 TI - Interferon-alpha-2a with or without 13-cis retinoic acid in patients with progressive, measurable metastatic renal cell carcinoma. AB - BACKGROUND: In patients with metastatic renal cell carcinoma (MRCC), interferon alpha (IFN) monotherapy leads to response rates of 5-15%, dependent on the selection of patients. In 1995, preclinical and clinical data indicated an improvement of these results if IFN was combined with 13-cis retinoic acid (CRA). METHODS: In a randomized Phase II study, patients with measurable MRCC received either subcutaneous IFN (9 MU daily; Arm A) or the same daily subcutaneous dose of IFN plus oral CRA (1 mg/kg; Arm B). A central expert panel reviewed the X-ray documentation of objective responses. RESULTS: In the 50 eligible patients from Arm A, the objective, expert-reviewed response rate was 6% (95% confidence interval [95% CI], 1.3-16.6%; 2 complete responses [CRs] and 1 partial response [PR]). A 19% response rate (95% CI, 9.4-32.0%) was stated for 53 eligible patients from Arm B (2 CRs and 8 PRs). Only one of the four CRs claimed by the clinical investigator was confirmed by the central review committee. Conversely, the expert committee deemed that 3 of 12 investigator-stated PRs were CRs. Constitutional toxicity (flu-like symptoms) and/or side effects from skin, mucosa, or eyes led to discontinuation of treatment in 22% of nonprogressing patients, more often in Arm B than in Arm A. CONCLUSIONS: The results from this randomized Phase II study support expansion of the trial into a Phase III study to evaluate the effect of IFN-CRA combination therapy on the survival of patients who undergo nephrectomy prior to IFN-based immunotherapy. The considerable interobserver variability of response evaluation (individual investigator vs. expert panel) indicates the necessity of a central review of claimed responses in future Phase II studies involving patients with MRCC. PMID- 15274067 TI - Natural history of bone complications in men with prostate carcinoma initiating androgen deprivation therapy. AB - BACKGROUND: As evidence accumulates in favor of androgen deprivation therapy (ADT) in patients with recurrent or metastatic prostate carcinoma, concern has increased regarding bone loss associated with therapeutic hypogonadism. The current study described the natural history of bone complications in men with prostate carcinoma who have initiated ADT. METHODS: Using 1992-2001 claims data from a 5% national random sample of Medicare beneficiaries, the authors identified men with prostate carcinoma who initiated ADT between 1992 and 1994. They analyzed inpatient, outpatient, and physician claims for bone complications over 7 subsequent years. They stratified the quartile of patients who survived longest into 2 cohorts: those who had received ADT for longer than and those who had received ADT for shorter than the median of 697 days. They evaluated the cumulative proportions of patients in each cohort with claims for pathologic fractures, osteoporosis/osteopenia, and nonpathologic fractures. RESULTS: In the 1992-1994 sample, 4494 men with prostate carcinoma initiated ADT. Of these, 1126 survived > 2028 days (5.5 years). During the first 3 years of evaluation, the proportion of bone events was similar for men with shorter durations of ADT and men with longer durations of ADT. However, by 7 years, more men in the longer ADT cohort (45%) had sustained at least 1 pathologic or nonpathologic fracture compared with men in the shorter ADT cohort (40%). CONCLUSIONS: In the current study, men with prostate carcinoma were found to be at risk for adverse bone effects from both the disease and the treatment. These longitudinal data revealed that fractures are common in this patient population and appear to be linked to the duration of ADT. PMID- 15274068 TI - A randomized, controlled trial of aerobic exercise for treatment-related fatigue in men receiving radical external beam radiotherapy for localized prostate carcinoma. AB - BACKGROUND: Advice to rest and take things easy if patients become fatigued during radiotherapy may be detrimental. Aerobic walking improves physical functioning and has been an intervention for chemotherapy-related fatigue. A prospective, randomized, controlled trial was performed to determine whether aerobic exercise would reduce the incidence of fatigue and prevent deterioration in physical functioning during radiotherapy for localized prostate carcinoma. METHODS: Sixty-six men were randomized before they received radical radiotherapy for localized prostate carcinoma, with 33 men randomized to an exercise group and 33 men randomized to a control group. Outcome measures were fatigue and distance walked in a modified shuttle test before and after radiotherapy. RESULTS: There were no significant between group differences noted with regard to fatigue scores at baseline (P = 0.55) or after 4 weeks of radiotherapy (P = 0.18). Men in the control group had significant increases in fatigue scores from baseline to the end of radiotherapy (P = 0.013), with no significant increases observed in the exercise group (P = 0.203). A nonsignificant reduction (2.4%) in shuttle test distance at the end of radiotherapy was observed in the control group; however, in the exercise group, there was a significant increase (13.2%) in distance walked (P = 0.0003). CONCLUSIONS: Men who followed advice to rest and take things easy if they became fatigued demonstrated a slight deterioration in physical functioning and a significant increase in fatigue at the end of radiotherapy. Home-based, moderate-intensity walking produced a significant improvement in physical functioning with no significant increase in fatigue. Improved physical functioning may be necessary to combat radiation fatigue. PMID- 15274069 TI - Monoclonal gammopathy of undetermined significance and multiple myeloma are associated with an increased incidence of venothromboembolic disease. AB - BACKGROUND: Venous thromboembolic disease (VTD) is a recently recognized complication of thalidomide in combination therapy for patients with multiple myeloma (MM). The authors assessed the frequency of VTD and its risk factors in 612 consecutive patients with plasma cell dyscrasia (PCD) who were evaluated and followed from 1991 to 2001. METHODS: In the current study, 404 patients were diagnosed with multiple myeloma (MM), 174 with monoclonal gammopathy of undetermined significance (MGUS), and 34 with other forms (excluding amyloidosis). Univariable correlates of VTD were assessed using Kaplan-Meier analysis and Cox proportional hazards analysis. RESULTS: The authors identified several univariable correlates of VTD in patients with MGUS, including a family and medical history of VTD, immobility, low serum albumin level, and high leukocyte count. Patients with MGUS with immunoglobulin (Ig) G monoclonal immunoglobulin were found to be less prone to develop VTD. In patients with MM, a family and medical history of VTD and the presence of a hypercoagulable state were factors identified in univariable analysis to be associated with an increased risk of VTD. In patients with MM, for each unit increase in serum albumin, the risk of VTD was lower. The type of the treatment regimen did not appear to correlate with the development of VTD. CONCLUSIONS: In the current study, the risk for thromboembolic diseases among patients with PCD was increased compared with the risk in the general population. Further studies are necessary to define the mechanisms involved. PMID- 15274070 TI - Risk assessment of patients with hematologic malignancies who develop fever accompanied by pulmonary infiltrates: a historical cohort study. AB - BACKGROUND: The mortality rate associated with fever accompanied by pulmonary infiltrates after chemotherapy for hematologic malignancies remains higher than the corresponding rate associated with febrile neutropenia without pulmonary infiltrates. Nonetheless, few studies have focused on the factors that predict outcome for patients with lung infiltrates. The purpose of the current study was to construct a risk model for clinical use by assessing the factors that affect outcome for patients with fever and pulmonary infiltrates. METHODS: A historical cohort of 110 patients with hematologic malignancies who developed fever and pulmonary infiltrates was examined. Using parameters for which data were available at the onset of lung infiltrates, univariate and multivariate analyses were performed to assess factors affecting outcome. After a value of one point was assigned to each significant variable, a prediction score was calculated for each patient; scores were used to generate a system for identifying patients with a low risk of death due to fever accompanied by pulmonary infiltrates. RESULTS: The crude mortality rate associated with pulmonary infiltrates was 23%; factors associated with cure included a favorable change in white blood cell counts (odds ratio [OR], 5.6; 95% confidence interval [CI], 1.7-18.9; P = 0.001), C-reactive protein levels < 10 mg/dL (OR, 4.6; 95% CI, 1.6-13.8; P = 0.001), and serum albumin levels > or = 3 g/dL (OR, 3.2; 95% CI, 1.4-7.3; P = 0.004). Low-risk patients (risk score, 2-3) and high-risk patients (risk score, 0-1) had survival rates of 95% and 46%, respectively (P < 0.0001). The risk model had a specificity of 88% and a positive predictive value of 95%. CONCLUSIONS: The risk model tested in the current study accurately predicted the survival of patients with hematologic malignancies who developed fever with pulmonary infiltrates. Once prospectively validated, the model could be used to select patients for trials involving novel diagnostic and therapeutic strategies. PMID- 15274071 TI - Oral capecitabine for the treatment of hepatocellular carcinoma, cholangiocarcinoma, and gallbladder carcinoma. AB - BACKGROUND: The goal of the current study was to evaluate the efficacy and toxicity of capecitabine in patients with nonresectable hepatobiliary carcinoma. METHODS: The authors performed a retrospective analysis of all patients with hepatocellular carcinoma (HCC), cholangiocarcinoma (CCA), or gallbladder carcinoma (GBC) who were ever treated with oral capecitabine. The medical records of 116 patients with hepatobiliary carcinoma who were treated at The University of Texas M. D. Anderson Cancer Center (Houston, TX) between July 1998 and March 1999 were reviewed. RESULTS: A total of 63 patients were treated with capecitabine (37 with HCC, 18 with CCA, 8 with GBC). Capecitabine 1000 mg/m(2) was administered twice daily for 14 days. Treatment was repeated every 21 days. Each patient received 1-15 treatment cycles. Nine patients (14%)-11% of patients with HCC, 6% of patients with CCA, and 50% of patients with GBC-had either a complete response (CR) or a partial response. A CR was radiologically confirmed in one patient with HCC and in two patients with GBC. The median survival times were 10.1 months (95% confidence interval [CI], 4.5-15.7 months) for patients with HCC, 8.1 months (95% CI, 7.4-8.9 months) for patients with CCA, and 9.9 months (95% CI, 4.4-15.4 months) for patients with GBC. The most common toxicity was hand-foot syndrome (37%). Grade 3 thrombocytopenia occurred in 8% of patients with HCC. No other significant toxicities were observed. For all patients, response to treatment was positively correlated with survival and decline in tumor markers. CONCLUSIONS: Capecitabine was found to be safe for patients with hepatobiliary carcinoma, including those with cirrhosis. The antitumor activity of single-agent capecitabine was most pronounced in patients with GBC, was modest in patients with HCC, and was poor in patients with CCA. PMID- 15274072 TI - Adapting the Lung Cancer Symptom Scale (LCSS) to mesothelioma: using the LCSS Meso conceptual model for validation. AB - BACKGROUND: The underpinning conceptual model for the Lung Cancer Symptom Scale (LCSS), an instrument used to assess health-related quality of life in patients with lung cancer, has been described elsewhere. The patient-rated scale of the LCSS was modified slightly for patients with mesothelioma (LCSS-Meso), because no other mesothelioma-specific instrument was available. METHODS: In the current methodologic study, the authors tested the conceptual model for the LCSS-Meso. Chemotherapy-naive patients with unresectable malignant pleural mesothelioma who were participating in two clinical trials of pemetrexed (ALIMTA; Eli Lilly, Indianapolis, IN) completed the scale twice before the start of therapy and once weekly during the trials. Three time points were analyzed: baseline, Day 40, and Day 82. Poisson regression was used to determine the contribution of predictive factors (i.e., symptoms) to the summary items (symptom distress, activity level, and global quality of life). RESULTS: The model was tested in 495 patients who had malignant pleural mesothelioma. More than 85% of patients reported pain, dyspnea, fatigue, and appetite loss. Pain, dyspnea, and fatigue were significant and stable predictors for all summary items; however, pain had a significant effect on global quality of life only through Day 40. Appetite loss was a significant and stable predictor of activity level and global quality of life. The explained variance for the model was 39-55%. CONCLUSIONS: Further support for the content validity of the LCSS-Meso was obtained, as nearly all patients validated that the symptoms described in the scale captured their disease experience. The only exception was the hemoptysis item, which was removed based on the current large normative data set. Support for the construct validity of the LCSS-Meso also was obtained. For both mesothelioma and lung cancer, the majority of factors within the LCSS model are relevant and have the expected amount of variability. These findings support the use of the LCSS as a sensitive instrument for serial measurement during clinical trials involving patients with lung malignancies. PMID- 15274073 TI - Pilot study of high-dose, concurrent biochemotherapy for advanced melanoma. AB - BACKGROUND: Durable complete response rates ranging from 5% to 20% have been reported in association with biochemotherapy for patients with metastatic melanoma, with response rates on the low end of this range being observed in trials that used lower doses and less intense treatment schedules. In the current study, the authors addressed the feasibility of increasing the doses of agents used in concurrent biochemotherapy. METHODS: Three patients with metastatic melanoma were enrolled at each of six concurrent biochemotherapy dose levels. The doses were as follows: dacarbazine 800 mg/m(2) or 1000 mg/m(2) (Day 1); cisplatin 25 mg/m(2) or 30 mg/m(2) (Days 1-4); vinblastine 1.6 mg/m(2) or 1.8 mg/m(2) (Days 1-5); interleukin-2 9 million units (MU) per m(2) or 12 MU/m(2) as a 24-hour continuous infusion (Days 1-4); and interferon-alpha-2b 5 MU/m(2), 10 MU/m(2), or 15 MU/m(2) (Days 1-5) and 5 MU/m(2) (Days 7, 9, and 11) administered subcutaneously. RESULTS: Of the 19 patients who were enrolled, 18 were evaluable for toxicity and response. Sixty-nine treatment courses were administered in total (median, 4 courses per patient; range, 1-6 courses per patient), with reduction of the dose of at least 1 agent being required in 7 courses (10%). Twenty-six courses were delayed by a median of 7 days (range, 3-29 days), with interferon-alpha-2b administration frequently omitted because of thrombocytopenia, most often after Day 5. Blood product support was required in 40 courses. Dose-limiting toxic effects included global encephalopathy, renal and hepatic dysfunction, pancreatitis, and ileus. There was 1 complete response, and there were 10 partial responses. The median time to disease progression was 6.9 months, and the median survival duration was 12.2 months. CONCLUSIONS: Although dose intensification can be achieved safely in patients with advanced melanoma, other strategies should be pursued to enhance the clinical activity of biochemotherapy as a response induction regimen. PMID- 15274074 TI - Association between laminin-8 and glial tumor grade, recurrence, and patient survival. AB - BACKGROUND: The authors previously sought to identify novel markers of glioma invasion and recurrence. Their research demonstrated that brain gliomas overexpressed a subset of vascular basement components, laminins, that contained the alpha4 chain. One of these laminins, laminin-8, was found to be present in highly invasive and malignant glioblastoma multiforme (GBM) (Grade 4 astrocytoma); its expression was associated with a decreased time to tumor recurrence, and it was found in vitro to promote invasion of GBM cell lines. METHODS: In the current study, the authors studied glial tumors of different grades in an attempt to correlate laminin-8 expression with tumor recurrence and patient survival. Immunohistochemistry and Western blot analysis were used to detect laminin isoforms of interest. RESULTS: Using immunohistochemistry and Western blot analysis, the authors confirmed high levels of laminin-8 expression in approximately 75% of the GBM cases examined and in their adjacent tissues, whereas astrocytomas of lower grades expressed for the most part a different isoform, laminin-9, which also was found in low amounts in normal brain tissue and benign meningiomas. Overexpression of laminin-8 in GBM was found to be associated with a statistically significant shorter time to tumor recurrence (P < 0.0002) and a decreased patient survival time (P < 0.015). CONCLUSIONS: The data suggest that laminin-8, which may facilitate tumor invasion, contributes to tumor regrowth after therapy. Laminin-8 may be used as a predictor of tumor recurrence and patient survival and as a potential molecular target for glioma therapy. PMID- 15274075 TI - The prevalence of pituitary adenomas: a systematic review. AB - BACKGROUND: Pituitary adenomas display an array of hormonal and proliferative activity. Once primarily classified according to size (microadenomas, < 1 cm; macroadenomas, > or = 1 cm), these tumors are now further classified according to immunohistochemistry and functional status. With these additional classifications in mind, the goals of the current study were to determine the prevalence of pituitary adenomas and to explore the clinical relevance of the findings. METHODS: The authors conducted a metaanalysis of all existing English-language articles in MEDLINE. They used the search string (pituitary adenoma or pituitary tumor) and prevalence and selected relevant autopsy and imaging evaluation studies for inclusion. RESULTS: The authors found an overall estimated prevalence of pituitary adenomas of 16.7% (14.4% in autopsy studies and 22.5% in radiologic studies). CONCLUSIONS: Given the high frequency of pituitary adenomas and their potential for causing clinical pathologies, the findings of the current study suggest that early diagnosis and treatment of pituitary adenomas should have far reaching benefits. PMID- 15274076 TI - Brain metastases of malignant germ cell tumors in children and adolescents. AB - BACKGROUND: Brain metastases of pediatric germ cell tumors are uncommon, and there is limited information regarding their incidence, clinical presentation, response to treatment, and influence on survival. METHODS: The authors reviewed the experience with brain metastases from pediatric germ cell tumors at St. Jude Children's Research Hospital (Memphis, TN) over a 40-year period. RESULTS: Between March 1962 and February 2002, 16 of 206 patients with germ cell tumors (7.8%) had brain metastases at the time of initial presentation (n = 2), later in the course of the illness (n = 12), or at autopsy (n = 2). Twelve of 16 patients (75%) had symptoms referable to the brain (nausea/emesis, headaches, or seizures), and 14 (88%) had pulmonary metastases at the time brain metastases were identified. Patients with brain metastases were more likely to have an extragonadal primary tumor (P = 0.013), advanced-stage disease at initial presentation (P = 0.016), and choriocarcinoma within the primary tumor (P < 0.001). The incidence of brain metastases was significantly lower in the second 2 decades of the study period (5 of 135 patients [3.7%]) than in the first 2 decades (11 of 71 patients [15.5%]; P = 0.005). Two of the 16 patients in the current study are long-term survivors. CONCLUSIONS: Brain metastases are uncommon in childhood germ cell tumors, and their incidence appears to be decreasing. In the current study, most patients with such metastases were symptomatic and had pulmonary metastases at the time brain metastases were identified. Patients with the highest risk of developing brain metastases include those with extragonadal tumors, those with high disease stage at initial presentation, and those with choriocarcinoma as a component of the primary tumor. The probability of survival is poor, although a small proportion of patients may become long-term survivors. PMID- 15274077 TI - Synovial sarcoma: a retrospective analysis of 271 patients of all ages treated at a single institution. AB - BACKGROUND: The optimal treatment strategy for synovial sarcoma (SS) is subject to debate, and different strategies have been used for pediatric and adult patients. The current retrospective analysis examined a large group of patients of all ages who were treated at a single institution over a 30-year period. METHODS: The study included 271 patients who ranged in age from 5 years to 87 years; 255 had localized disease, which was macroscopically resected in 215 cases and deemed unresectable at diagnosis in 40 cases. Chemotherapy was administered to 41% of patients, corresponding to 76% of patients age or 16 years and < 20% of older patients; 28% of patients with macroscopically resected disease received chemotherapy on an adjuvant basis. RESULTS: The 5-year event-free survival rate for the study cohort as a whole was 37%, although this rate varied with age (66%, 40%, and 31% for patients age < or = 16 years, 17-30 years, and > 30 years, respectively). Chemotherapy was used more commonly for children than for adults. Among patients with surgically resected disease, the 5-year metastasis-free survival (MFS) rate was 60% for those who were treated with chemotherapy and 48% for those who were not; the benefit associated with chemotherapy use appeared to be greatest for patients age > or = 17 years who had tumors measuring > 5 cm (MFS, 47% [chemotherapy] vs. 27% [no chemotherapy]). In the subgroup of patients with measurable disease, the rate of tumor response to chemotherapy was approximately 48%. CONCLUSIONS: Although the authors await more convincing proof of the efficacy of adjuvant chemotherapy in the treatment of adult soft tissue sarcoma, they recommend that patients with high-risk SS (tumor size > 5 cm) be the first to be considered for this type of treatment. PMID- 15274078 TI - Assessment of humoral immunity to poliomyelitis, tetanus, hepatitis B, measles, rubella, and mumps in children after chemotherapy. AB - BACKGROUND: To evaluate the effect of chemotherapy on humoral immunity to vaccine preventable disease, the authors investigated the persistence of protective antibody titers in a group of patients who were alive and well after they were treated for pediatric malignancies. METHODS: Serum antibody levels were evaluated for polio, tetanus, hepatitis B, rubella, mumps, and measles in 192 children. The terms lack of immunity and loss of immunity, respectively, were used to describe the absence of immunity in patients who were tested only after chemotherapy and in patients who were tested both before and after chemotherapy and determined to have immunity before chemotherapy. RESULTS: Overall, the absence of a protective serum antibody titer for hepatitis B, measles, mumps, rubella, tetanus, and polio was detected in 46%, 25%, 26%, 24%, 14%, and 7% of patients, respectively. On univariate analysis, loss of antibodies against rubella, mumps, and tetanus was associated significantly with younger age (P < 0.001, P = 0.02, and P = 0.001, respectively), and loss of antibodies against measles was significantly associated with younger age and female gender (P = 0.0003 and P = 0.008, respectively). The administration of 59 booster vaccinations to 51 patients who had lost > or = 1 protective antibody titer resulted in an overall response rate of 93%. CONCLUSIONS: Chemotherapy induced different rates of loss of protective antibody titers depending on the type of vaccination administered. This finding may be responsible for the failure of vaccination programs for patients who have undergone chemotherapy. The administration of a booster dose after the completion of chemotherapy is a simple and cost-effective way to restore humoral immunity against most vaccine-preventable diseases. PMID- 15274079 TI - Hepatocellular carcinoma and liver tumors in South African children: a case for increased prevalence. AB - BACKGROUND: The high regional incidence of hepatocellular carcinoma (HCC) in South Africa also may be present in children of the region, although the link to hepatitis B (HBV) appears less clear. The objective of this study was to assess the incidence and probable causes of HCC in South African children. METHODS: Data were obtained from seven participating pediatric oncology units and from the tumor registry to review hepatic tumors in children in South Africa. RESULTS: One hundred ninety-four children (ages 0-14 years) presented with malignant primary hepatic tumors (1988-2003). One hundred twelve tumors (57%) were hepatoblastoma (HB), 68 tumors (35%) were hepatocellular carcinoma (HCC) (including 9 patients with the fibrolamellar variant, 6 of which occurred in black children), 10 tumors (5%) were sarcoma of the liver, and 4 tumors were lymphoma. The ratio of HB to HCC (1.67) was markedly lower compared with other reports, suggesting a greater prevalence of HCC. Correlation with population statistics indicated an incidence of 1.066 malignant liver tumors per year per 10(6) children age < 14 years (HB, 0.61 per 10(6) children; HCC, 0.39 per 10(6)). Two-thirds of patients with HCC were positive for HBV surface antigen (HBsAg), and HCC occurred mostly in black African patients (93%). The mean age of onset was 1.47 years for HB and 10.48 years for HCC. A preponderance of males (3.5:1.0) was noted in the HBsAg-positive group that was not reflected elsewhere. Serum alpha-fetoprotein (AFP) levels were raised both in patients with HB (100%; most AFP levels were very high) and in patients with HCC (69%), although 15% of patients with HCC had low or normal AFP levels. CONCLUSIONS: It appeared from the current results that HCC is more prevalent among children in South Africa compared with the children in more developed countries, although their rates were lower that the rates noted in adults. A collaborative approach will be required to improve their diagnosis and management. PMID- 15274080 TI - The natural history of incidentally detected small renal masses. PMID- 15274082 TI - High-dose tamoxifen and sulindac as first-line treatment for desmoid tumors. PMID- 15274084 TI - Gemcitabine and docetaxel as front-line chemotherapy in patients with carcinoma of an unknown primary site. PMID- 15274086 TI - Ribosome motions modulate electrostatic properties. AB - The electrostatic properties of the 70S ribosome of Thermus thermophilus were studied qualitatively by solving the Poisson-Boltzmann (PB) equation in aqueous solution and with physiological ionic strength. The electrostatic potential was calculated for conformations of the ribosome derived by recent normal mode analysis (Tama, F., et al. Proc Natl Acad Sci USA 2003 100, 9319-9323) of the ratchet-like reorganization that occurs during translocation (Frank, J.; Agrawal, R. K. Nature 2000 406, 318-322). To solve the PB equation, effective parameters (charges and radii), applicable to a highly charged backbone model of the ribosome, were developed. Regions of positive potential were found at the binding site of the elongation factors G and Tu, as well as where the release factors bind. Large positive potential areas are especially pronounced around the L11 and L6 proteins. The region around the L1 protein is also positively charged, supporting the idea that L1 may interact with the E-site tRNA during its release from the ribosome after translocation. Functional rearrangement of the ribosome leads to electrostatic changes which may help the translocation of the tRNAs during the elongation stage. PMID- 15274087 TI - Protein structure preference, tRNA copy number, and mRNA stem/loop content. AB - From statistical analyses of protein sequences for humans and Escherichia coli we found that the messenger RNA segment of m-codons (for m=2 to 6) with average high tRNA copy number (TCN) (larger than approximately 10.5 for humans or approximately 1.95 for E. coli) preferably code for the alpha helix and that with low TCN (smaller than approximately 7.5 for humans or approximately 1.7 for E. coli) preferably code for coil. Between them there is an intermediate region without correlation to structure preference. For the beta strand the preference/ avoidance tendency is not obvious. All strong preference-modes of TCN for protein secondary structures have been deduced. The mutual interaction between two factors--protein secondary structural type and codon TCN--is tested by F distribution. A phenomenological model on the relation between structure preference and translational efficiency or accuracy is proposed. It is pointed out that the structure preference of codons is related to the distribution of mRNA stem/loop content in three TCN regions. PMID- 15274088 TI - Characterization of the conformational behavior of peptide Contryphan Vn: a theoretical study. AB - In this work we report the study of a peptide, the Contryphan Vn produced by Conus ventricosus, a vermivorous cone snail living in the temperate Mediterranean sea. This cyclic peptide of nine residues is a ring closed by a Cys-Cys (Cys: cysteine) disulfide bond containing two proline (Pro) residues and two tryptophans (Trp), one of them being a D-Trp. We present a statistical mechanical characterization of the peptide, simulated in water for about 200 ns with classical molecular dynamics (MD). In recent years there has been a growing interest in the study of the mechanics and dynamics of biological molecules, and in particular for proteins and peptides, about the relationship between collective motions and the active conformations which exert the biological function. To this aim we used the essential dynamics analysis on the MD trajectory and extracted, from the total fluctuations of the molecule, the dominant dynamical modes responsible of the principal conformational transitions. The Contryphan Vn small size allowed us to investigate in details the all-atoms dynamics and the corresponding thermodynamics in conformational space defined by the most significant intramolecular motions. PMID- 15274089 TI - Hydration of proteins: SAXS study of native and methoxy polyethyleneglycol (mPEG) modified L-asparaginase and bovine serum albumin in mPEG solutions. AB - Two mPEG-modified globular proteins [mPEG: methoxy poly(ethylene glycol)], and their native unmodified forms, were examined by small-angle x-ray scattering to evaluate the extent of their surface hydration. The effects of free and protein bound mPEG on the hydration shell were modeled with discrete electron density profiles. We show that an mPEG-depleted layer can account for the decrease in the measured radius of gyration R(g) from 34.1 to 31.1 A in native L-asparaginase, and from 32.4 to 31.0 A in native bovine serum albumin (BSA) in mPEG-containing solvents. For mPEG-modified proteins in mPEG-free solvents, we attribute the observed increase in the R(g) over that of the native proteins (approximately 3% in L-asparaginase, and 10% in BSA) to the presence of mPEG on the protein surface. The R(g) of the mPEG-modified proteins in mPEG solutions generally decrease with mPEG concentration. Relative to the corresponding unmodified protein, this decrease in R(g) is much larger in BSA (from 35.6 to 31.2 A) but much smaller (from 34.9 to 34.3 A) in L-asparaginase. From these studies, the thickness of the hydration layer around L-asparaginase and BSA is estimated to be approximately 15 A. Exclusion of polyols from the protein domain could be related to the presence of the hydration shell around the protein. PMID- 15274090 TI - Ultrabright fluorescein-labeled antibodies near silver metallic surfaces. AB - Fluorescein-labeled antibodies are widely used in clinical assays and fluorescence microscopy. The fluorescent signal per labeled antibody is limited by fluorescein self-quenching, which occurs when the antibody is heavily labeled with multiple fluoresceins. We examined immunoglobulin G (IgG) when labeled with 0.7 to about 30 fluoresceins per antibody molecule. The extent of self-quenching was decreased, and the signal increased, when the labeled antibody was in close proximity to metallic silver particles. Time-resolved measurements showed that the intensity increase was due in part to a silver-induced increase in the radiative decay rate. These results suggest the use of labeled antibodies conjugated to silver particles as ultrabright probes for imaging or analytical applications. PMID- 15274091 TI - Conformational studies on enkephalins using the MOLS technique. AB - Conformational studies of two linear enkephalin molecules, Met-enkephalin and Leu enkephalin, have been carried using the mutually orthogonal Latin squares (MOLS) technique with the ECEPP/3 force field. This technique was developed recently in our laboratory to perform an unbiased search of the conformational space of peptides and to locate low energy conformations. The present study identified all the folds predicted by other studies, and in addition picked up other energetically favorable structures. The results suggest that the peptide backbone exists as a mixture of folded and unfolded forms (approximately 50% each). The study also provides information on the distribution of the low energy conformations that we have classified on the basis of structural motifs, backbone hydrogen-bonding patterns, and root mean square deviations in atomic positions. PMID- 15274094 TI - Bilirubin influence on oxidative lung damage and surfactant surface tension properties. AB - To study the hypothesis that hyperbilirubinemia might reduce in vivo oxidative lung damage while also diminishing lung surfactant surface tension properties during acute lung injury, we performed a randomized study in a rabbit model of acute lung injury. Twenty rabbits were randomized to receive bilirubin or saline intravenously. Acute lung injury was induced by lung lavages with saline. Lung tissue oxidation was evaluated by measuring total hydroperoxide (TH), advanced oxidation protein products (AOPP), and protein carbonyls (PC) in bronchial aspirate (BA) samples. Surface surfactant activity was studied in BA samples using a capillary surfactometer. Bilirubin BA concentration increased in bilirubin-treated rabbits, while it remained undetectable in controls. A similar increase in TH, AOPP, and PC bronchial aspirate concentrations was found in both the study and control groups, while surfactant surface activity was lower in the bilirubin than in the control group. We conclude that during hyperbilirubinemia, bilirubin enters the lung tissue, where it can be detected in BA fluid. Bilirubin is not effective as an antioxidant agent and exerts a detrimental effect on lung surfactant surface tension properties. These findings may have relevance to the management of premature neonates suffering from respiratory distress syndrome and hyperbilirubinemia. PMID- 15274095 TI - Simultaneous treatment of asthma and allergic rhinitis. AB - Asthma and allergic rhinitis (AR) form a well-recognized comorbidity. This study aims at assessing the efficacy of nasally inhaled beclomethasone dipropionate (BDP) in their simultaneous treatment. A randomized controlled trial was conducted with 78 allergic rhinitis and asthma patients aged 5-17 years. Seventy five individuals completed the study. During 8 weeks, 38 subjects received BDP CFC aerosol (>or= 500 mcg/day) exclusively via nasal inhalation through a facemask attached to a plastic valved spacer. The control group (37 patients) received 200 mcg/day of aqueous intranasal beclomethasone plus oral inhalation of BDP-CFC (>or= 500 mcg/day) through a mouthpiece connected to the same spacer. Primary outcomes analyzed in order to assess the response to treatment were clinical scoring for allergic rhinitis and measurements of nasal inspiratory peak flow (NIPF). AR clinical scoring and NIPF did not differ in the two groups at admission or at nearly all follow-up visits. Nasal inhalation of beclomethasone dipropionate provides AR symptom relief while maintaining control of asthma by delivering it to the lungs. Therefore, this therapeutic strategy might be considered for patients suffering from this comorbidity, especially in low resource countries, since it is less expensive than the conventional treatment. PMID- 15274096 TI - Smoking habits in adolescents with mild to moderate asthma. AB - To study the impact of mild to moderate asthma on smoking habits in adolescents. Specifically, we tested the hypothesis that asthma does not prevent adolescents from smoking. A research questionnaire, filled by a systematic sample of military personnel upon enrollment to service in the Israeli Defense Force (IDF), was analyzed. Conscripts were asked to voluntarily fill (after obtaining a signed informed consent) a research questionnaire about their medical history, and several health related topics including smoking. This database was matched with the military medical profile of the soldier, which includes the patient asthma status. Overall, 38,047 young adults were included in this study. There was a significant increase in the rate of mild to moderate asthma, from the mid-1980's to date. During the 1980's and early 1990's, asthmatics smoked significantly less frequently (20-22%) than non-asthmatics (25-27%). In the mid- to late-1990's, the smoking rates increased relatively more in asthmatics, to the point that in the last 8 years of this study, they were found to be almost identical in both groups, at a rate of approximately 30%. The presence of asthma is not a powerful motivating agent to prevent from smoking. It is likely that smoking asthmatic teenagers are at risk for suboptimal lung growth, and as young adults, they will become at greater risk of lung function deterioration. We suggest that primary care physicians, caring for asthma in children, adolescents, and young adults, should explain the particular risks generated by tobacco smoking. PMID- 15274097 TI - Urban air pollution and asthma in children. AB - This study investigated the relationship between atmospheric pollution and emergency hospital admission for asthma among children resident in Turin in the period 1997-1999, using a case-control design. On the basis of the primary diagnosis, pediatric patients (< 15 years old) resident in Turin and admitted for asthma were defined as cases (n(1) = 1,060); age-matched patients admitted for causes other than respiratory diseases or heart diseases were defined as controls (n(2) = 25,523). Nitrogen dioxide (NO(2) in microg/m(3)) and total suspended particulates (TSP in microg/m(3)) were considered as indicators of urban air pollution; sex and age of patient, seasonality, temperature, humidity, solar radiation, and day of admission were considered as principal confounders. Statistical analyses were performed using simple and multiple logistic regression models; the association between emergency admission for asthma and exposure was shown as percentage of risk modification for a 10 microg/m(3) increment of exposure to each pollutant and relative 95% confidence interval. The number of emergency admissions for respiratory causes rose significantly with increased exposure to each pollutant: 2.8% (95% CI, 0.7-4.9%) and 1.8% (95% CI, 0.3-3.2) for a 10 microg/m(3) increment of exposure to NO(2) and TSP, respectively. A significant association was found between increased number of hospital emergency admissions for respiratory causes and exposure to principal urban pollutants in Turin. The study confirms the results reported for other Italian and European cities, using a case-control design. PMID- 15274098 TI - Relation of sweat chloride concentration to severity of lung disease in cystic fibrosis. AB - In cystic fibrosis (CF), sweat chloride concentration has been proposed as an index of CFTR function for testing systemic drugs designed to activate mutant CFTR. This suggestion arises from the assumption that greater residual CFTR function should lead to a lower sweat chloride concentration, as well as protection against severe lung disease. This logic gives rise to the hypothesis that the lower the sweat chloride concentration, the less severe the lung disease. In order to test this hypothesis, we studied 230 patients homozygous for the DeltaF508 allele, and 34 patients with at least one allele associated with pancreatic sufficiency, born since January 1, 1955, who have pulmonary function data and sweat chloride concentrations recorded in our CF center database, and no culture positive for B. cepacia. We calculated a severity index for pulmonary disease, using an approach which takes into account all available pulmonary function data as well as the patient's current age and survival status. Patients with alleles associated with pancreatic sufficiency had significantly better survival (P = 0.0083), lower sweat chloride concentration (81.4 +/- 23.8 vs. 103.2 +/- 14.2 mEq/l, P < 0.0001), slower rate of decline of FEV(1) % predicted ( 0.75 +/- 0.34 vs. -2.34 +/- 0.17% predicted per year), and a better severity index than patients homozygous for the DeltaF508 allele (median 73rd percentile vs. median 55th percentile, P = 0.0004). However, the sweat chloride concentration did not correlate with the severity index, either in the population as a whole, or in the population of patients with alleles associated with pancreatic sufficiency, who are thought to have some residual CFTR function. These data suggest that, by itself, sweat chloride concentration does not necessarily predict a milder pulmonary course in patients with cystic fibrosis. PMID- 15274099 TI - Snoring and atopic disease: a strong association. AB - The prevalence and factors associated with snoring and habitual snoring in Asian children are largely unknown. Our objectives were to evaluate the prevalence and factors associated with snoring and habitual snoring in preschool and primary school children in Singapore. A self-response questionnaire on snoring was administered to parents of children aged 4-7 years in randomly selected preschools and primary schools in Singapore. The overall response rate was 91.3% (nt = 11,114). Snoring and habitual snoring were reported in 28.1% and in 6.0% of the children, respectively. On multivariate logistic regression analysis, snoring was significantly associated with male gender, race, atopy (asthma, allergic rhinitis, or atopic dermatitis), maternal atopy (allergic rhinitis or atopic dermatitis), maternal smoking, and breastfeeding. Habitual snoring was significantly associated with obesity (odds ratio (OR), 3.75; 95% confidence interval (CI), 1.67-8.42), allergic rhinitis (OR, 2.90; 95% CI, 2.06-4.08), atopic dermatitis (OR, 1.80; 95% CI, 1.28-2.54), maternal smoking (OR, 2.22; 95% CI, 1.09-4.53), and breastfeeding (OR, 1.49; 95% CI, 1.11-1.98). Atopy was the strongest risk factor for habitual snoring, and the effect was cumulative. The odds ratio of a child with all three atopic diseases (asthma, allergic rhinitis, and atopic dermatitis) to have habitual snoring was 7.45 (95% CI, 3.48-15.97). In conclusion, snoring and habitual snoring are common in Asian children. Atopy is strongly associated with snoring and habitual snoring. We suggest that children who are significantly atopic receive additional attention during screening for snoring, habitual snoring, and other features of obstructive sleep apnea syndrome. PMID- 15274100 TI - A simple method of reducing complications of pediatric nonbronchoscopic bronchoalveolar lavage. AB - Our objective was to determine whether a simple method of maintaining positive pressure ventilation during nonbronchoscopic bronchoalveolar lavage (NB-BAL) would successfully reduce the incidence and/or severity of desaturation events. Our design was a clinical trial with historical controls. Seventy ventilated pediatric patients undergoing diagnostic NB-BAL participated. Two NB-BAL techniques were compared: 1) the "unsealed" method, where the suction catheter was passed through an open system, maintaining oxygenation but not airway pressure; and 2) the "sealed" technique, which was identical except that the catheter was passed through a diaphragm, maintaining positive pressure ventilation throughout. NB-BAL was performed on 35 patients using the "unsealed" technique and 2 years later on 35 patients using the "sealed" method. Heart rate and oxyhemoglobin saturation (SaO(2)) were recorded before, during, and after NB BAL. There was no difference between groups with regard to demographic data, oxygenation, or ventilatory requirements (P >or= 0.1). The "sealed" group experienced a median drop in SaO(2) of 6.0% (range, -6% to 44%), and the "unsealed" group a drop of 13.0% (-2% to 61%), during NB-BAL (P < 0.05). Patients with oxygenation index greater than 10 experienced the most severe desaturation events in both groups: 53.8% of patients in the "sealed" group with oxygenation index >10 desaturated to <80% vs. 91.6% in the "unsealed" group (P < 0.05). In conclusion, we describe a simple, inexpensive modification of the NB-BAL technique that reduces the incidence and severity of desaturation during NB-BAL. PMID- 15274101 TI - Reduced eosinophil pro-fibrogenic effect in severe childhood asthma compared to mild disease: an effect of corticosteroids? AB - Eosinophils play an important role in inflammation and probably in airway remodeling in asthma. We previously demonstrated that eosinophils from atopic subjects display pro-fibrogenic properties towards lung fibroblasts partially by preformed transforming growth factor-beta (TGF-beta). We hypothesized that the pro-fibrogenic potential of eosinophils is increased in children with life threatening asthma (LTA). Six children with atopic LTA clinically well-controlled by inhaled corticosteroids (ICS) and 5 children with atopic mild asthma (MA) treated only with inhaled beta(2)-agonists were investigated. The effects of their peripheral blood eosinophils on fibroblast proliferation and lattice contraction were investigated. In addition, TGF-beta(1) and IL-6 eosinophil content were also evaluated. Unexpectedly, eosinophils from LTA increased fibroblast proliferation (5.4-fold) and gel contraction (1.1-fold) significantly less than those from MA. TGF-beta(1) but not IL-6 eosinophil content in LTA was significantly lower than that in MA (2.7-fold). In vitro, addition of dexamethasone on eosinophils stimulated by mast cells resulted in a marked decrease in their TGF-beta(1) content by 1.6-fold. In conclusion, eosinophils from children with ICS-treated LTA displayed significantly less pro-fibrogenic properties than those from MA treated only with beta(2)-agonists. Our data suggest that the pro-fibrogenic effect of eosinophils might be influenced by treatment with ICS in childhood asthma. PMID- 15274102 TI - Asthma and acute chest in sickle-cell disease. AB - Our objective was to determine if physician-diagnosed asthma increases the risk of acute chest syndrome (ACS) in children with sickle-cell disease (SCD) hospitalized for pain. Our study design was a retrospective case-control study of all SCD patients, aged 2-21 years, hospitalized for pain during the interval 1999 2000. Medical records of first admissions during the interval were reviewed to determine the presence of ACS during the admission. Cases were defined as patients with ACS, and controls were patients without ACS. Independently, medical records of admissions prior to the study interval were reviewed for evidence of physician-diagnosed asthma. Sixty-three cases with ACS and 76 controls without ACS were identified. No significant differences in gender, age, and hemoglobin phenotype were found. Patients with physician-diagnosed asthma were 4.0 times (95% CI, 1.7, 9.5) more likely to develop ACS during the admission than patients without asthma. Individuals with physician-diagnosed asthma had a longer hospitalization for ACS, i.e., 5.6 days vs. 2.6 days, respectively (P = 0.01). In conclusion, our preliminary data suggest that asthma in children with SCD admitted to the hospital for pain may be a risk factor for ACS and may increase the duration of hospitalization when compared to children with SCD and without asthma. PMID- 15274103 TI - Lymphocytic inflammation in childhood bronchiolitis obliterans. AB - Childhood bronchiolitis obliterans (CBO) is an infrequent, severe disorder characterized by persistent obstructive respiratory symptoms after an acute episode of bronchiolitis. The viral etiology is most common, and adenovirus is the most frequently identified causative agent. Pathologically, the disease is characterized as constrictive type BO, with variable degrees of chronic inflammation and fibrosis in the bronchioles. The nature of the cellular infiltrate is largely unknown, and its characterization may provide better understanding of the disease and offer clues for therapy. Therefore, the aim of the present study was to characterize the inflammatory infiltrate in the bronchioles of 23 open lung biopsies of children with CBO and to compare this to the infiltrate in histologically normal airways. Our results show that CD3+ T cells were the most frequent cell type observed in CBO, with a predominance of the CD8+ T-cell subtype. When compared to the control group, there was a larger number of CD8+, CD4+, CD20+, granzyme B+, and perforin+ lymphocytes in the CBO group. Further studies are needed to address the role of different cell types in the development of CBO. PMID- 15274104 TI - Volume guarantee: stability of tidal volume and incidence of hypocarbia. AB - Excessive tidal volume (V(T)) can lead to lung injury, hypocarbia, and neurologic damage. Volume guarantee (VG) uses exhaled V(T) as the control variable to reduce the risk of volutrauma and more closely control PaCO(2). Our objective was to test the hypothesis that VG combined with assist/control (A/C) will maintain PaCO(2) and V(T) within target range more consistently than assist/control alone during the first 72 hr of life in ventilated preterm infants. Eligible infants were randomly assigned to A/C + VG or A/C alone. Data were recorded directly from the pressure and volume module of the Draeger Babylog 8000+ ventilator. Arterial blood gases were obtained every 2-6 hr, as clinically indicated. In A/C, inspiratory pressure was adjusted to achieve a V(T) of 4-6 ml/kg. In VG, the target V(T) was 5 ml/kg. Subsequent adjustments were made by the clinical team in response to arterial blood gas measurements (ABG). Proportion of breaths and PaCO(2) values outside the target range were compared by chi(2), and continuous variables by t-test. There were no differences in demographic or baseline ventilator variables between the 18 infants in the two groups. For 1,805/11,950 breaths (15.1%), V(T) was > target with A/C + VG, vs. 2,503/9,853 (25.4%) with A/C (P < 0.001). V(T) was < target for 21.7% of breaths with A/C + VG, vs. 35.7% with A/C (P < 0.001). Twenty percent of PaCO(2) values were < target, with A/C + VG vs. 36.3% with A/C, P < 0.05. The proportion of PaCO(2) values > target was similar in the two groups. Oxygenation and mean pH were not different. No complications related to mechanical ventilation were observed. In conclusion, VG significantly reduced hypocarbia and excessively large V(T). This suggests the potential to reduce pulmonary and neurologic complications of mechanical ventilation. Larger studies are needed to establish safety and demonstrate such benefits. PMID- 15274105 TI - Reference values for forced inspiratory flows in children aged 7-15 years. AB - In order to construct reference equations, we attempted to measure forced inspiratory flows, i.e., peak inspiratory flow (PIF) and maximal inspiratory flow at 50% of FVC (MIF50%FVC) in 332 healthy schoolchildren aged 7-15 years during flow-volume loop measurements, using an electronic spirometer. In 255 children (122 boys and 133 girls), the results were satisfactory. Statistical analysis revealed that the only predictive variables were sex and height. The best fit of the data was obtained with the power model (Y = A * H(B)); the coefficients of correlation between flows and height ranged from 0.66-0.77, and were slightly greater for boys. Forced inspiratory flows in children increase with height, and the variability is higher than for forced expiratory flows. Reference values for forced inspiratory flows can be useful in assessing the ability of children to generate affective inspiratory flows for choosing an inhalation device, or in resolving diagnostic problems, e.g., extrathoracic obstruction. PMID- 15274106 TI - Inhaled fluticasone dipropionate decreases levels of nitric oxide in recurrenty wheezy infants. AB - We examined the effect of inhaled fluticasone diproprionate (FDP) on symptoms, lung function (FEV(0.5)), and exhaled nitric oxide (Fe(NO)) in infants with recurrent wheeze and raised Fe(NO). Thirty-one infants aged 6-19 months (mean, 12.7 months; 12 girls) completed the study. All infants had a history of recurrent wheeze and a parental history of atopy. All children had raised Fe(NO), as determined by an offline tidal breathing technique prior to randomization. Lung function and Fe(NO) were assessed before and after 4 weeks of treatment with FDP or placebo. The parents recorded daily symptoms during the treatment period. Sixteen infants received FDP and 15 the placebo for 4 weeks. At completion of the study, infants treated with FDP had a significant reduction in Fe(NO) (35.0 ppb to 16.5 ppb) compared to those that received placebo (35.2 ppb to 30.2 ppb) (P = 0.05). Small increases in FEV(0.5) were observed in both groups, but these changes were not different between groups (P = 0.8). Symptom scores were not significantly different in either group following the intervention. We showed that a moderate dose of inhaled FDP reduces levels of Fe(NO), a potential marker of airway inflammation, even in the absence of significant changes in lung function and symptoms. PMID- 15274107 TI - Preoperative polysomnograms and infant pulmonary function tests do not predict prolonged postoperative mechanical ventilation in children following scoliosis repair. AB - Our objective was the identification of children with scoliosis at higher risk of prolonged postoperative mechanical ventilation (MV) permits improved pre- and perioperative respiratory care to reduce postoperative complications. Pulmonary function testing (PFT) predicts prolonged postoperative MV in children who can reliably perform PFT, but some children cannot perform PFT. The objective of this study was to determine if polysomnography (PSG) or infant pulmonary function testing (IPFT) could predict prolonged postoperative MV (defined as MV >3 days) in children undergoing scoliosis surgery who could not reliably perform PFT. We studied 110 patients (age range, 10.8 +/- 4.9 [SD] years) who had preoperative PSG, and 18 patients (age range, 4.0 +/- 2.9 [SD] years) who had preoperative IPFT prior to undergoing any type of scoliosis repair by the Children's Hospital of Los Angeles Division of Orthopedic Surgery from January 1990- July 2001. The following information was reviewed and correlated: preoperative PSG parameters (baseline and nadir S(aO(2) ), baseline and peak P(ETCO(2) ), and apnea hypopnea index [AHI]), preoperative IPFT parameters (respiratory system compliance [C(rs)], respiratory system resistance [R(rs)], tidal volume [V(T)], and FRC), and length of postoperative MV. Twenty-seven patients (25%) who had PSG and 5 patients (28%) who had IPFT required postoperative MV >3 days. There was no association between baseline and nadir S(aO(2) ) 100 times in the case of one protein) and arrays (20 times in the case of eight proteins) without the need for intermediate re-inking; (iii) transferring spots of proteins with good homogeneity in surface coverage to glass slides; (iv) applying this technique to surface-based immunoassays; (v) stamping that requires only sub nanomolar amounts of protein (typically approximately 3 microg in approximately 0.6 microL of solution); (vi) stamping without the need for drying of the proteins, as opposed to stamping with stamps made of poly(dimethylsiloxane); and (vii) patterning gradients of proteins by allowing two proteins to diffuse toward each other in an agarose stamp, followed by printing the protein gradients onto a surface. PMID- 15274133 TI - Preprocessing of two-dimensional gel electrophoresis images. AB - Proteomics produces a huge amount of two-dimensional gel electrophoresis images. Their analysis can yield a lot of information concerning proteins responsible for different diseases or new unidentified proteins. However, an automatic analysis of such images requires an efficient tool for reducing noise in images. This allows proper detection of the spots' borders, which is important in protein quantification (as the spots' areas are used to determine the amounts of protein present in an analyzed mixture). Also in the feature-based matching methods the detected features (spots) can be described by additional attributes, such as area or shape. In our study, a comparison of different methods of noise reduction is performed in order to find out a method best suited for reducing noise in gel images. Among the compared methods there are the classical methods of linear filtering, e.g., the mean and Gaussian filtering, the nonlinear method, i.e., median filtering, and also the methods better suited for processing of nonstationary signals, such as spatially adaptive linear filtering and filtering in the wavelet domain. The best results are obtained by filtering of gel images in the wavelet domain, using the BayesThresh method of threshold value determination. PMID- 15274134 TI - Atomic force microscopy revelation of molecular complexes in the multiprotein cytochrome P450 2B4-containing system. AB - The application of atomic force microscopy (AFM) to the identification and visualization of individual molecules and their complexes in a reconstituted monooxygenase P450 2B4 system without the phospholipid was demonstrated. The method employed in this study distinguishes the monomeric proteins from their binary complexes and, also, the binary from the ternary complexes. The AFM images of the full-length P450 2B4 system's constituent components - cytochrome P450 2B4 (2B4), NADPH-cytochrome P450 reductase and cytochrome b5 (b5), were obtained on highly-oriented pyrolitic graphite. The typical heights of the d-2B4, d flavoprotein (Fp) and d-b5 molecules were measured and found to be 2.2 +/- 0.2, 2.3 +/- 0.2 and 1.8 +/- 0.1 nm, respectively. The measured heights of the binary d-Fp/d-2B4 and d-2B4/d-b5 complexes were estimated to be 3.4 +/- 0.2 and 2.8 +/- 0.2 nm, respectively. No formation of d-Fp/d-b5 complexes was registered. The ternary d-Fp/d-2B4/d-b5 complexes were visualized and their heights were found to be roughly equal to 4.3 +/- 0.3 nm and 6.2 +/- 0.3 nm. PMID- 15274135 TI - Identification of proteins in activated human neutrophils susceptible to tyrosyl radical attack. A proteomic study using a tyrosylating fluorophore. AB - Tyrosyl radicals cross-linked to protein tyrosine residues (tyrosylated proteins) represent hallmarks of neutrophil-mediated injury at the inflammatory locus. Yet the proteins targeted by tyrosyl radicals in an intact cellular system remain to be elucidated. Here, we show that tyrosyl radicals generated by human neutrophils after activation by phorbol 12-myristate 13-acetate (PMA), interferon-gamma (IFN gamma) or TNF-alpha could act in an autocrine manner by cross-linking to endogenous proteins. We have identified the tyrosylated proteins by using a membrane-impermeable tyrosine analogue, tyramine coupled to fluorescein (TyrFluo), in combination with proteomics techniques. Confocal microscopy images indicated that initially the tyrosylated proteins were localized in patches at the cell surface to become internalized subsequently. In the neutrophil membrane associated proteome, lactoferrin was the prime target of tyrosylation. Out of three isoforms identified, an 80 kDa neutral isoform was tyrosylated more extensively than the 85 kD basic isoform, particularly after PMA activation. Although all three stimuli induced tyrosylation of the filamentous component vimentin, additional tyrosylated vimentin fragments were detected after IFN-gamma and TNF-alpha-stimulation. Moreover, upon activation the bulk of vimentin behaved as a dimer (M(r) 120 kDa) being slightly tyrosylated, yet phosphorylated at Thr-425 possibly as a requirement for its externalization. Unexpectedly, bovine catalase added to end tyrosyl radicals formation was detected as a highly tyrosylated neutrophil-associated protein. A moderate stimulus-dependent tyrosylation of ATP synthase-beta, alpha-enolase, glyceraldehyde 3-phosphate dehydrogenase, cytokeratin-10, filamin-A, and annexin-I was also observed. When the membrane-permeable probe (acetylTyrFluo) was used, protein tyrosylation was not observed indicating that the intracellular proteins were well protected against oxidative attack. This study shows that human neutrophils can modulate their proteome via a tyrosine oxidation pathway induced by pro-inflammatory mediators. PMID- 15274136 TI - Quantitative proteome profiling during the fermentation process of pleiotropic Bacillus subtilis mutants. AB - Using a combined quantitative proteomic and bioinformatic approach, we monitored the cytoplasmic proteome profile of the Gram-positive bacterium Bacillus subtilis during a fermentation process in complex medium. Proteome signatures were applied to elucidate the physiological changes occurring in the gene expression profile during growth. Furthermore, we determined the significance level of quantitative proteome changes, identified relative to the threshold of scatter in replicated samples and developed a statistically rigorous method that allowed us to determine significant fold-changes at 95% confidence between different proteomes. Different functional groups of proteins were clustered according to their pattern of significant expression changes. The largest group is induced by the exhaustion of glucose and the presence of alternative carbon and nitrogen sources. Furthermore, depletion of glucose caused the induction of the trichloroacetic acid (TCA) cycle enzymes and the downregulation of glycolytic enzymes. The onset of the transition phase may be provoked by amino acid starvation, resulting in the RelA-dependent repression of proteins involved in the translation process and in the induction of amino acid biosynthetic pathways. Comparisons between the parental strain and two subtilisin-hypersecreting strains revealed only small cytoplasmic differences in the main metabolic pathways. Instead, the overproduction of degradative enzymes in both of these mutants was reflected in the extracellular proteome. PMID- 15274137 TI - Proteomic response to amino acid starvation in Candida albicans and Saccharomyces cerevisiae. AB - Saccharomyces cerevisiae activates general amino acid control (GCN) in response to amino acid starvation. Some aspects of this response are known to be conserved in other fungi including Candida albicans, the major systemic fungal pathogen of humans. Here, we describe a proteomic comparison of the GCN responses in S. cerevisiae and C. albicans. We have used high-resolution two-dimensional (2-D) gel electrophoresis and peptide mass fingerprinting to develop a 2-D protein map of C. albicans. A total of 391 protein spots, representing 316 open reading frames, were identified. Fifty-five C. albicans and 65 S. cerevisiae proteins were identified that responded reproducibly to 3-aminotriazole (3AT) in a Gcn4p dependent fashion. The changes in the S. cerevisiae proteome correlated with the response in the S. cerevisiae transcript profile to 3AT treatment (rank correlation coefficient = 0.59; Natarajan et al., Molec. Cell. Biol. 2001, 21, 4347-4368). Significant aspects of the GCN response were conserved in C. albicans and S. cerevisiae. In both fungi, amino acid biosynthetic enzymes on multiple metabolic pathways were induced by 3AT in a Gcn4p-dependent fashion. Carbon metabolism functions were also induced. However, subtle differences were observed between these fungi. For example, purine biosynthetic enzymes were induced in S. cerevisiae, but were not significantly induced in C. albicans. These differences presumably reflect the contrasting niches of these relatively benign and pathogenic yeasts, respectively. PMID- 15274138 TI - Proteome analysis of barley seeds: Identification of major proteins from two dimensional gels (pI 4-7). AB - Germination of monocotyledonous plants involves activation and de novo synthesis of enzymes that degrade cell walls and starch and mobilize stored endosperm reserves for embryo growth. Two-dimensional (2-D) gel electrophoresis and mass spectrometry were applied to identify major water-soluble proteins in extracts of mature barley (Hordeum vulgare) seeds and to follow their fate during germination. About 1200 and 600 spots of pI 4-7 were detected on 2-D gels by silver staining and colloidal Coomassie Brilliant Blue staining, respectively. About 300 spots were selected for in-gel digestion followed by matrix-assisted laser desorption/ionization-mass spectrometry-peptide map fingerprint analysis. Database searches using measured peptide masses resulted in 198 identifications of 103 proteins in 177 spots. These include housekeeping enzymes, chaperones, defence proteins (including enzyme inhibitors), and proteins related to desiccation and oxidative stress. Sixty-four of the identifications were made using expressed sequence tags (ESTs). Numerous spots in the 2-D gel pattern changed during germination (micromalting) and an intensely stained area which contained large amounts of the serpin protein Z appeared centrally on the 2-D gel. Spots containing alpha-amylase also appeared. Identification of 22 spots after three days of germination represented 13 different database entries and 11 functions including hydrolytic enzymes, chaperones, housekeeping enzymes, and inhibitors. PMID- 15274139 TI - UDP-glucose pyrophosphorylase is upregulated in carriers of the porcine RN- mutation in the AMP-activated protein kinase. AB - The AMP-activated protein kinase (AMPK) plays a key role in the regulation of energy metabolism in eukaryotic cells acting as a metabolic sensor. In its activated form AMPK inhibits ATP consuming pathways and stimulates ATP generating pathways. A dominant mutation, denoted RN(-), in the porcine PRKAG3 gene, encoding the regulatory gamma3 subunit of AMPK, results in hyperaccumulation of glycogen in glycolytic skeletal muscle cells. To study the effects of this mutation on protein expression patterns in skeletal muscle, comparative proteome analysis of muscle samples from 12 animals (6 rn (+)/rn (+) and 6 RN(-)/rn (+)) was performed. The major finding of the proteome analysis was that the key enzyme in the synthesis of glycogen, UDP-glucose pyrophosphorylase, was significantly up regulated in RN(-) carriers. This observation was subsequently supported by studies of enzyme activity and Northern blot analysis. Furthermore, the expression patterns of enzymes related to glycolysis and the citric acid cycle were also affected. Our data suggests that hyperaccumulation of glycogen mediated by the RN(-) mutation is due to an increased synthesis of glycogen. PMID- 15274140 TI - Pleiotropic molecular effects of the pro-apoptotic dietary constituent flavone in human colon cancer cells identified by protein and mRNA expression profiling. AB - The flavonoid flavone contained in a variety of fruits and vegetables was identified as a very potent apoptosis inducer in human colonic cancer cells. In search of the molecular targets of flavone action in HT-29 cells we analyzed changes in mRNA and protein expression levels by proteomics and oligonucleotide array technologies. Proteome analysis identified several heat-shock proteins, annexins, and cytoskeletal caspase substrates as regulated by flavone and these protein classes are known to play a role in apoptosis induction and execution. Protein kinase C-beta, which serves as an ultimate marker for colon cancer development was no longer detectable in HT-29 cells exposed to flavone. Besides proteins involved in gene regulation or detoxification pathways, proteins involved in intermediary metabolism were altered by flavone exposure and this was associated with changes in the flux of energetic substrates. Oligonucleotide arrays, using chips with around 10 000 oligonucleotides spotted, revealed numerous changes in transcript levels of genes related to signaling, transcription, cancer development but also to metabolism. In conclusion, flavone has a surprisingly broad spectrum of effects on mRNA and protein expression in a human colonic cancer cell line with clusters of targets related to its apoptosis inducing activity and to cellular metabolism. PMID- 15274141 TI - Proteomics of buccal squamous cell carcinoma: the involvement of multiple pathways in tumorigenesis. AB - Squamous cell carcinoma (SCC) of the buccal mucosa is an aggressive oral cancer. It mainly occurs in Central and Southeast Asia, and is closely related to the practice of tobacco smoking and betel squid chewing. The high recurrence and low survival rates of buccal SCC require our continued efforts to understand the pathogenesis of the disease for designing better therapeutic strategies. We used proteomic technology to analyze buccal SCC tissues aiming at identifying tumor associated proteins for the utilization as biomarkers or molecular targets. With the exception of alpha B-crystallin being substantially reduced, a number of proteins were found to be significantly over-expressed in cancer tissues. These increased proteins included glycolytic enzymes, heat-shock proteins, tumor antigens, cytoskeleton proteins, enzymes involved in detoxification and anti oxidation systems, and proteins involved in mitochondrial and intracellular signaling pathways. These extensive protein variations indicate that multiple cellular pathways were involved in the process of tumorigenesis, and suggest that multiple protein molecules should be simultaneously targeted as an effective strategy to counter the disease. At least, SCC antigen, G protein, glutathione S transferase, manganese superoxide dismutase, annexins, voltage-dependent anion channel, cyclophilin A, stratifin and galectin 7 are candidates for targeted proteins. The present findings also demonstrated that rich protein information can be produced by means of proteomic analysis for a better understanding of the oncogenesis and pathogenesis in a global way, which in turn is a basis for the rational designs of diagnostic and therapeutic methods. PMID- 15274142 TI - Comprehensive proteome analysis of ovarian cancers using liquid phase separation, mass mapping and tandem mass spectrometry: a strategy for identification of candidate cancer biomarkers. AB - A two-dimensional (2-D) liquid phase separation method, liquid isoelectric focusing followed by nonporous reversed-phase high performance liquid chromatography (HPLC), was used to separate proteins from human ovarian epithelial whole cell lysates. HPLC eluent was interfaced on-line to an electrospray ionization (ESI) time of flight (TOF) mass spectrometer to obtain accurate intact protein molecular weights (Mr). 2-D protein expression maps were generated displaying protein isoelectric point (pI) versus intact protein Mr. Resulting 2-D images effectively displayed quantitative differential protein expression in ovarian cancer cells versus non-neoplastic ovarian epithelial cells. Protein peak fractions were collected from the HPLC eluent, enzymatically digested, and analyzed by matrix-assisted laser desorption/ionization (MALDI) TOF mass spectrometry (MS) peptide mass fingerprinting and by MALDI-quadrupole TOF tandem mass spectrometry peptide sequencing. Interlysate comparisons of differential protein expression between two ovarian adenocarcinoma cell lines, ES2 and MDAH-2774, and ovarian surface epithelial cells was performed. Five pI fractions from each sample were selected for comparative study and over 300 unique proteins were positively identified from the 2-D liquid expression maps using MS, which covered around 60% of proteins detected by on-line ESI-TOF-MS. This represents one of the most comprehensive proteomic analyses of ovarian cancer samples to date. Protein bands with significant up- or down-regulation in one cell line versus another as viewed in the 2-D expression maps were identified. This strategy may prove useful in identifying novel ovarian cancer marker proteins. PMID- 15274143 TI - Proteomic tools to characterize the protein fraction of Equidae milk. AB - The principal components of the protein fraction in pony mare's milk have been successfully identified and partially characterized using proteomic tools. Skimmed pony mare's milk was fractionated by either reversed phase-high performance liquid chromatography (RP-HPLC) on a C4 column or a bi-dimensional separation technique coupling RP-HPLC in the first dimension and sodium dodecyl sulfate-polyacrylamide electrophoresis (SDS-PAGE) in the second dimension (two dimensional RP-HPLC/SDS-PAGE). The fractions thus obtained were analyzed by Edman N-terminal microsequencing and mass determination, with or without tryptic digestion, on a matrix-assisted laser desorption/ionization-time of flight spectrometer. Based on the sequence and molecular mass information obtained, identifications were achieved through a protein database search using homology or pattern research algorithms. This methodological approach was shown to be rapid, efficient and reliable in identifying the principal proteins in pony mare's milk. kappa-, alpha(s1)-, alpha(s2)-, and beta-casein, lysozyme C, alpha-lactalbumin and beta-lactoglobulin I and II were thus identified. alpha(s1) and beta-caseins displayed polymorphic patterns, probably due to alternative splicing processes leading to casual exon skipping events involving exons 7 and 14 in alpha(s1) casein and exon 5 in beta-casein. Edman N-terminal microsequencing over 35 amino acid residues, for pony alpha(s1)-casein, clearly demonstrated the occurrence, in Equidae, of a splicing pattern similar to that reported in rodents, characterized by the constitutive outsplicing of exon 5. Pony mare's milk SDS-PAGE and RP-HPLC patterns were compared with those obtained for other milks (cow, goat and human), as were the relative levels of caseins and major whey proteins in these milks. Our results provide further evidence to support the notion that Equidae milk is closer to human breast milk than milk from bovine and caprine with respect to the casein and lysozyme C contents and casein/whey proteins ratio. PMID- 15274148 TI - Effect of stents in reducing restenosis in small coronary arteries: a meta analysis. AB - The ability of stents to reduce restenosis was established in larger coronary arteries. Clinical trials of stenting in smaller vessels have yielded conflicting results due in part to their sample sizes. The aim of this meta-analysis was to increase the statistical power by pooling data from these clinical trials. Trials were identified from Medline search, review of recent cardiology meetings' abstracts, and manual review of bibliographies. Studies were included if they were prospective randomized controlled trials. Endpoints examined included a dichotomized definition of angiographic restenosis, target lesion revascularization (TLR), target vessel revascularization (TVR), or any repeat revascularization. Pooling of data was performed by calculating a Mantel-Haenszel odds ratio (OR). The analysis included 2,598 patients enrolled in eight clinical trials. Stenting significantly reduced restenosis (OR = 0.62; 95% CI = 0.61 0.63). Concordantly, stenting reduced TLR (OR = 0.49), TVR (OR = 0.90), and any revascularization (OR = 0.48). This meta-analysis supports the hypothesis that stenting reduces restenosis in small coronary arteries as well as in larger coronary arteries. The apparent discordant result of individual clinical trials was due in part to underpowering related to small sample sizes. PMID- 15274149 TI - Stenting small vessels? Not a small issue. PMID- 15274150 TI - Coronary anatomy and left ventricular ejection fraction in patients with type 2 diabetes admitted for elective coronary angiography. AB - Patients with diabetes mellitus (DM) have more severe coronary artery disease and a two- to fourfold higher risk for myocardial infarction and death as compared to patients without DM. In this study, we analyzed coronary anatomy, left ventricular ejection fraction, and cardiac risk factors in patients with DM referred for coronary angiography and compared them with findings in nondiabetic patients. Coronary anatomy was assessed in a total of 6,234 patients and left ventricular ejection fraction in a subset of 4,767 (76.5%) patients. Diabetic patients (n = 641) were older (60.8 +/- 9.6 vs. 58.5 +/- 10.5 years; P < 0.0001) and had higher rates of hypertension (65% vs. 47%; P < 0.0001). Three-vessel disease (DM 44.7% vs. no DM 25.4%; P < 0.0001) and reduced left ventricular ejection fraction (DM 58.4% +/- 15.2 vs. no DM 63.9% +/- 13.2; P < 0.0001) were significantly associated with DM. After adjustment for age and other vascular risk factors, the presence of DM was associated with a higher atherosclerotic burden. We conclude that advanced coronary heart disease and left ventricular dysfunction are highly prevalent in diabetic patients, independent of age and other cardiovascular risk factors. Thus, cardiac assessment in diabetic patients should, in addition to optimal diabetic control, involve screening for left ventricular dysfunction. PMID- 15274151 TI - Acute and long-term clinical and angiographic outcome after S-Stent implantation: S-Stent multicenter safety and efficacy trial. AB - The purpose of this study is to demonstrate safety and effectiveness of the S Stent in de novo coronary lesions treated with conventional percutaneous coronary balloon angioplasty. Between January 2000 and June 2001, 120 patients were prospectively enrolled at four study centers. Patients were treated with coronary stenting in a total of 137 lesions. Procedural success was achieved in 100% of 137 attempted lesions. Clinical success was 99.8%. In-hospital mortality was 0.8%; myocardial infarction occurred in 0.8% and stent thrombosis in 0.8%. After stent implantation, the minimal lumen diameter increased from 0.92 +/- 0.43 to 2.74 +/- 0.36 mm (P < 0.0001) and the percent diameter stenosis decreased from 68.0 +/- 16.2 to 4.5 +/- 12.0 (P < 0.0001). At 6-month follow-up, the percent diameter stenosis was 33.5 +/- 21.3 and the angiographic restenosis rate was 16.5%. Target lesion revascularization was required in 12 patients (10.1%). We conclude that the use of S-Stent for coronary intervention resulted in a high procedural success rate and low angiographic restenosis at 6 months after implantation. PMID- 15274152 TI - Combined fibrinolysis using reduced-dose alteplase plus abciximab with immediate rescue angioplasty versus primary angioplasty with adjunct use of abciximab for the treatment of acute myocardial infarction: Asia-Pacific Acute Myocardial Infarction Trial (APAMIT) pilot study. AB - We conducted a randomized feasibility pilot study comparing combined fibrinolysis with immediate rescue angioplasty vs. primary angioplasty with adjunctive abciximab in patients with acute myocardial infarction (AMI). Seventy patients with ST segment elevation AMI of /= 3 years) outcomes of coronary artery stenting using Palmaz-Schatz and Multi-Link stent implantations between November 1995 and October 1999 were analyzed. There were 655 Palmaz-Schatz stent implantations in 577 lesions on 477 patients (group A) and 428 Multi-Link stent implantations in 381 lesions on 326 patients (group B). The baseline characteristics were similar in the two groups. Group B had more complex lesions, longer stenotic lesions, and larger reference vessel sizes than group A. However, both groups had a similar in hospital cardiac events. Four hundred and two patients with 488 lesions in group A and 260 patients with 307 lesions in group B underwent a 6-month follow-up coronary angiography. The restenotic rate per lesion was 16% in both groups (P = 0.872). A 3-year angiographic follow-up was performed in 262 patients of group A (301 lesions) and 139 patients of group B (162 lesions), and restenosis was noted in only 3 patients (1.36%) in group A and 5 patients (4%) in group B, in which the lesion was patent at the 6-month angiographic follow-up. Significant increase in minimal luminal diameter was noted from 2.23 +/- 0.66 mm at 6 months to 2.33 +/- 0.64 mm in group A (P < 0.01), and insignificant increase from 2.23 +/- 0.77 to 2.28 +/- 0.82 mm was noted in group B (P = 0.27). No differences were noted between the two groups in mortality, reinfarction, recurrent angina, target lesion angioplasty, or elective coronary artery bypass surgery during a follow-up period of 60 +/- 3 months. Forty-five patients (9.4%) in group A and 18 patients (5.5%) in group B received additional stenting procedures for newly developed lesions. The overall cardiac event-free survival was 66% in group A and 72% in group B (P = 0.844). In conclusion, the procedural success rate, in-hospital morbidity, 6-month angiographic results, and long-term (>/= 3 years) clinical and angiographic outcomes were similar with coronary stenting using either Palmaz Schatz or Multi-Link stent. The stented lesions were stable; however, late regression of minimal luminal diameter was noted in both groups, and progression of atherosclerotic change in the nonstented site was noted during long-term follow-up. PMID- 15274154 TI - Luminal narrowing due to intramural hematoma shift from left anterior descending coronary artery to left circumflex artery. AB - This case report demonstrates subacute luminal narrowing 20 days after balloon angioplasty in the left anterior descending coronary artery due to an intramural hematoma. Stenting was performed and resulted in side-branch compromise caused by squeezing the hematoma from the left anterior descending coronary artery into the left circumflex artery. Another stent was deployed to treat the stenosis in the left circumflex artery. PMID- 15274155 TI - Ventricular septal rupture masquerading as coronary perforation during intervention for acute myocardial infarction. AB - Ventricular septal rupture is a serious complication of acute myocardial infarction. We experienced a case of septal rupture immediately after primary angioplasty with thrombolysis, whose angiographic findings were similar to those of coronary perforation. The progression of septal rupture was delineated by the serial angiograms. PMID- 15274156 TI - Percutaneous transcatheter retrieval of retained balloon catheter in distal tortuous coronary artery: a modified double-helix approach. AB - The rapid increase in the incidence of retained components in the coronary arteries during angioplasty raises the question of the optimal approach in these cases. Management extends from simply leaving the fragment behind to aggressive surgical removal. Increasingly, percutaneous approaches are being considered. A new percutaneous two-guidewire helix method has been reported. We describe a case of successful retrieval of an angioplasty balloon using this approach. PMID- 15274157 TI - Effects of contrast media on porcine bone marrow-derived mononuclear cells and calf myoblast viability and secretion of VEGF and MCP-1. AB - We investigated the effect of contrast media on bone marrow-derived cell viability, growth factor secretion, and myoblast viability. Bone marrow was exposed to contrast media, mononuclear cells were isolated, viability was assessed by Trypan blue exclusion or cultured for 4 weeks, and conditioned medium was assayed for vascular endothelial growth factor (VEGF) and monocyte chemoattractant protein-1 (MCP-1). Skeletal myoblasts viability was assessed after exposing them to contrast media. In separate experiments, bone marrow or bone marrow-derived mononuclear cells were exposed to contrast media, cultured for 40 hr, then assessed for viability. None of the contrast media tested had any effect on bone marrow-derived cell viability. Hypaque or Hexabrix increased myoblasts viability by 8-10%. VEGF and MCP-1 concentrations in the conditioned medium increased in a time-related manner. These findings support the concept that for cell therapy, bone marrow cells or myoblasts may be mixed with contrast media and injected into ischemic myocardium without compromise in viability or function. PMID- 15274158 TI - Emergency localization of radioactive seeds lost during intracoronary brachytherapy. AB - Recently, it has been reported that brachytherapy catheters ruptured in vivo. Localization of lost beta-radiation-emitting seeds is a problem because no appropriate technique is available that is rapid and precise. We developed a technique to localize beta-emitting seeds utilizing the effect that beta radiation induces bremsstrahlung. The loss of a single radioactive source was simulated in an Alderson Phantom representing a human body. The beta-induced bremsstrahlung could be detected selectively by a gamma-camera. The position of the radioactive seed could be located within 5 min with an accuracy of +/- 0.5 cm. The result of this study suggests that in an emergency case of loss of a brachytherapy source, a commercially available gamma-camera can be a valuable tool to detect lost beta-radiation-emitting seeds rapidly and precisely. In addition, the technique minimizes the patient's as well as the surgeon's exposure to radiation and reduces the extent of surgical trauma. PMID- 15274159 TI - Current status of rotational atherectomy. AB - Despite the increasing use of percutaneous transluminal coronary angioplasty and intracoronary stent placement for the treatment of obstructive coronary artery disease, a large subset of coronary lesions cannot be adequately treated with balloon angioplasty and/or intracoronary stenting alone. Such lesions are often heavily calcified or fibrotic and undilatable with the present balloon technology and attempts to treat them with balloon angioplasty or intracoronary stent placement often lead to vessel dissection or incomplete stent deployment with resultant adverse outcomes. Rotational atherectomy remains a useful niche device for the percutaneous treatment of such complex lesions, usually as an adjunct to subsequent balloon angioplasty and/or intracoronary stent placement. In contrast to balloon angioplasty or stent placement that widen the coronary lumen by displacing atherosclerotic plaque, rotational atherectomy removes plaque by ablating the atherosclerotic material, which is dispersed into the distal coronary circulation. Other lesion subtypes amenable to treatment with this modality include ostial and branch-ostial lesions, chronic total occlusions, and in-stent restenosis. This review discusses the technique and principles of rotational atherectomy, the various treatment strategies for its use (including adjunctive pharmacotherapy), the lesion-specific applications for this device, and the complications unique to this modality. Recommendations are also made for its use in the current interventional era. PMID- 15274160 TI - Endovascular stents for treatment of coarctation of the aorta: acute results and follow-up experience. AB - Balloon angioplasty as treatment for coarctation of the aorta is increasingly performed. Endovascular stents have been proposed as a means of improving the efficacy and safety of the procedure. In this report, we describe one institution's immediate results and clinical follow-up after implantation of endovascular stents. Retrospective analysis for endovascular stent placement for coarctation of the aorta between 1993 and 2002 was made. The immediate hemodynamic results and clinical follow-up were reviewed. Thirty-two patients underwent attempted stent placement for coarctation. Twenty-three patients had postoperative recurrent coarctation and nine had native coarctation. The systolic gradient decreased from 31 to 1.8 mm Hg (P = 0.001) and the diameter was increased 8.1 to 13.5 mm (P-0.001). Mean follow-up was 1.5 years. The mean follow up gradient as assessed by sphygomomanometry was 13.1 mm Hg. Eight patients underwent 10 successful further dilations. Complications included one stent migration and one aortic dissection. The use of stents as an adjunct to balloon angioplasty in selected patients with coarctation can be performed with low complication rates and provides excellent immediate relief of obstruction with promising follow-up. Further dilation of these stents is possible. Long-term follow-up is warranted. PMID- 15274161 TI - Novel growth stent for the permanent treatment of vessel stenosis in growing children: an experimental study. AB - Stent implantation in stenotic vessels of infants and small children is problematic because there is no ideal stent model that is small enough to be easily introduced into the infant femoral vein or artery and, at the same time, large enough to be dilated during growth to adult vessel diameters. To overcome this problem, we designed a new stent, the growth stent. This growth stent is a balloon-expandable metal stent. Two longitudinal halves are connected with bioabsorbable sutures so that a circular stent is created. It was postulated that after absorption of the sutures the stent would not impede growth. Twenty of these stents were implanted in the aorta, pulmonary arteries, and inferior vena cava of piglets (average weight 6.9 kg). After 18 weeks (14-23 weeks) and a mean weight gain of 59 kg, none of the stented vessels showed any significant stenosis or pressure gradient, documented by angiography and catheter pullback. During fluoroscopy, the two halves of the stent were clearly separated in all animals. The growth stent has the potential to be nonrestrictive during vessel growth, and thus is a promising new device for the permanent treatment of stenotic vessels in infancy and childhood. PMID- 15274162 TI - Does a "split" stent make sense? PMID- 15274163 TI - Transcatheter patent foramen ovale closure for secondary prevention of paradoxical embolic events: acute results from the FORECAST registry. AB - Patients with patent foramen ovale (PFO) and paradoxical embolism are at increased risk for recurrent events. Percutaneous PFO closure is a new treatment modality aimed at secondary prevention. We report the multicenter experience of interventional closure of PFO using two different devices in 272 patients. The mean age was 51 +/- 14.2 years and 52.9% were males. The implantation procedure resulted in an initial complete shunt closure rate of 74.3% with a periprocedural complication rate of 6.6%. There were no deaths or pericardial tamponade. The mean procedure time was less than 44 min under either conscious sedation, local or general anesthesia. This is the largest report to date demonstrating the procedural safety, reliability, and feasibility of the transcatheter PFO closure technique with the STARFlex and CardioSEAL septal implants. Randomized clinical trials are currently in progress to identify patients most likely to benefit from this intervention. PMID- 15274164 TI - Closure of patent foramen ovale: The abolition of stroke and asymptotic realities. PMID- 15274165 TI - Clinical outcomes of patent foramen ovale closure for paradoxical emboli without echocardiographic guidance. AB - The objective of this study was to determine the feasibility of device closure of patent foramen ovale (PFO) for presumed paradoxical emboli without echocardiographic guidance or balloon sizing and the clinical outcome after device closure. Closure of the PFO has been proposed as an alternative to anticoagulation in patients with presumed paradoxical emboli. At present, most centers perform device closure with transesophageal echocardiographic guidance and balloon sizing of the defect. Between May 1998 and April 2002, 92 consecutive patients underwent device closure for a PFO using fluoroscopic monitoring only. Procedural success and major complications were recorded. Follow-up outcomes were recurrence rate and residual atrial shunting on transthoracic echocardiography. All patients (mean age, 45 +/- 13 years; 52% male) had successful device deployment using either the CardioSeal (n = 78) and Amplatzer (n = 14) PFO occluders with no major complications. Mean procedure time and fluoroscopy time was 27 +/- 13 and 6 +/- 4 min, respectively. One patient had a residual shunt on echocardiography at 1 year. Cumulative event-free survival for recurrence of paradoxical embolus at 1 year was 97.3% +/- 1.8%. This study provides a basis for device closure of PFO becoming a safe, day-case procedure, resulting in a low rate of residual shunting and recurrent thromboembolic events. PMID- 15274166 TI - Percutaneous closure of multiple atrial septal defects with three Amplatzer septal occluder devices. AB - We report on the successful closure of three atrial septal defects with three Amplatzer septal occluders in a 55-year-old woman. PMID- 15274167 TI - Noncardiac applications of glycoprotein IIb/IIIa inhibitors. AB - Platelet activation and aggregation have become increasingly recognized as the primary processes involved in the cascade that leads to thrombus formation in atherosclerotic vascular disease. Glycoprotein IIb/IIIa receptor inhibitors (GPI) favorably impact thrombus formation and distal embolization by inhibiting the final common pathway of platelet aggregation. Glycoprotein IIb/IIIa inhibitors have been used effectively in a wide variety of clinical scenarios including unstable angina, non-ST segment elevation myocardial infarction, ST segment elevation myocardial infarction, and low and high risk percutaneous coronary interventions with and without intracoronary stenting, however there is limited data regarding the use of these potent antiplatelet agents in the setting of extracardiac vascular disease. This article will review the non-cardiac applications of glycoprotein IIb/IIIa inhibitors in the setting of acute ischemic stroke, carotid and vertebral angioplasty and stenting, acute critical limb ischemia, and percutaneous interventions in peripheral arterial occlusive disease. PMID- 15274168 TI - Unmet needs in bipolar depression. AB - Bipolar disorders, particularly bipolar spectrum disorders, frequently go unrecognized and undiagnosed by clinicians and thus remain untreated or inappropriately treated. Although the symptoms of bipolar I disorder are widely acknowledged and recognized among clinicians, epidemiology sampling studies over the past several years have found that bipolar II disorder and bipolar spectrum disorders are likely to be more prevalent and more challenging to diagnose, particularly as depressive presentations are far more common in these groups. Bipolar disorder is associated with increased morbidity and mortality, as well as higher healthcare costs, but it is unclear how much of the consequences of bipolar disorder are unrecognized in the face of poor recognition of bipolar II and bipolar spectrum disorders. This article addresses challenges in diagnosing and treating bipolar disorder in the face of a depressive episode, and offers guidelines for recognizing and appropriately managing these patients. Studies with the newer anticonvulsant mood stabilizer lamotrigine have shown antidepressant effects in bipolar disorder, and may fill an unmet need for treatment options in patients who present with depression in the context of bipolar disorder. PMID- 15274169 TI - Cognitive-behavioral therapy with childhood anxiety disorders: functioning in adolescence. AB - We examined anxiety symptoms, anxiety-related impairment, and further treatment in adolescents who received cognitive behavioral therapy (CBT) for childhood anxiety disorders 6-7 years previously. Forty-three adolescents and their parents (14 boys, 29 girls; mean age 16.7 years) participated in structured telephone interviews. Participants (68% of initial sample of 63) did not differ in age, diagnostic profile, socioeconomic status, or initial severity from nonparticipants but more girls than boys participated. Indices based on child- and parent-reported symptoms and impairment were calculated, and within-sample comparisons by age, gender, diagnosis, and initial severity were done using t tests. Predictors of symptoms and impairment were also examined. On average, adolescents reported modest levels of anxiety-related impairment. Further treatment for anxiety had occurred in 30% (13 of 43) of patients. Stepwise regressions found female gender and diagnosis other than generalized anxiety disorder predictive of increased symptoms by parent report, and initial severity predicted adolescent-reported impairment. Adolescents showed limited internalizing symptomatology and impairment but almost one third had required further treatment. Studies comparing treated and untreated samples are needed to clarify whether CBT alters the natural history of childhood anxiety disorders and to replicate our findings regarding predictors of symptomatology and impairment. PMID- 15274170 TI - Appraisal of social concerns: a cognitive assessment instrument for social phobia. AB - The current study describes the validation of a new cognitive assessment measure for social phobia, entitled the Appraisal of Social Concerns (ASC). Item content is relevant to a range of social situations. The ASC can be used to tailor interventions to patients' idiosyncratic concerns. Data are presented from both clinical (n = 71) and non-clinical (n = 550) samples. Preliminary data indicate that the ASC has good internal consistency and test-retest reliability. The construct validity of the ASC is comparable to that of well-established measures in use with social phobics. A strength of the ASC is its sensitivity to the effect of treatment. An exploratory factor analysis yielded three factors tapping concerns about negative evaluation, observable symptoms, and social helplessness. Subscale scores were strongly correlated. Preliminary findings suggest that the ASC is a psychometrically sound, time efficient instrument that can be used for both clinical and research purposes. PMID- 15274171 TI - Post-treatment effects of exposure therapy and clomipramine in obsessive compulsive disorder. AB - We sought to determine whether adults with obsessive-compulsive disorder (OCD) who respond to intensive exposure and response (ritual) prevention (EX/RP) with or without clomipramine (CMI) fare better 12 weeks after treatment discontinuation than responders receiving CMI alone. After receiving 12 weeks of treatment (EX/RP, CMI, EX/RP+CMI, or pill placebo [PBO] in a randomized clinical trial conducted at three outpatient research centers), 46 adults with OCD who responded to treatment (18 EX/RP, 11 CMI, 15 EX/RP+CMI, 2 PBO) were followed after treatment discontinuation for 12 weeks. Patients were assessed every 4 weeks with the Yale-Brown Obsessive-Compulsive Scale, the National Institutes of Health Global Obsessive-Compulsive Scale, and the Clinical Global Impressions scale by an evaluator who was blind to original treatment assignment. The primary hypothesis was that EX/RP and EX/RP+CMI responders would be less likely to relapse 12 weeks after treatment discontinuation than responders to CMI alone. Twelve weeks after treatment discontinuation, EX/RP and EX/RP+CMI responders, compared to CMI responders, had a significantly lower relapse rate (4/33 = 12% versus 5/11 = 45%) and a significantly longer time to relapse. The CMI relapse rate was lower than previously reported. Nonetheless, responders receiving intensive EX/RP with or without CMI fared significantly better 12 weeks after treatment discontinuation than responders receiving CMI alone. PMID- 15274172 TI - Escitalopram in the treatment of generalized anxiety disorder: double-blind, placebo controlled, flexible-dose study. AB - Escitalopram has been shown in clinical trials to improve anxiety symptoms associated with depression, panic disorder, and social anxiety disorder. This study was designed to evaluate the efficacy and tolerability of escitalopram in the treatment of generalized anxiety disorder (GAD). Outpatients (18 years or older) who met DSM-IV criteria for GAD, with baseline Hamilton Rating Scale for Anxiety (HAMA) scores > or = 18, were randomly assigned to double blind treatment with escitalopram (10 mg/day for the first 4 weeks and then flexibly dosed from 10-20 mg/day) or placebo for 8 weeks, following a 1-week, single-blind, placebo lead-in period. The primary efficacy variable was the mean change from baseline in total HAMA score at Week 8. The escitalopram group (N = 158) showed a statistically significant, and clinically relevant, greater improvement at endpoint compared with placebo (N = 157) in all prospectively defined efficacy parameters. Significant improvement in HAMA total score and HAMA psychic anxiety subscale score for the escitalopram-treated group vs. the placebo-treated group was observed beginning at Week 1 and at each study visit thereafter. Mean changes from baseline to Week 8 on the HAMA total score using a last-observation-carried forward (LOCF) approach were -11.3 for escitalopram and -7.4 for placebo (P<.001). Response rates at Week 8 were 68% for escitalopram and 41% for placebo (P<.01) for completers, and 58% for escitalopram and 38% for placebo LOCF values (P<.01). Treatment with escitalopram was well tolerated, with low rates of reported adverse events and an incidence of discontinuation due to adverse events not statistically different from placebo (8.9% vs. 5.1%; P=.27). Escitalopram 10 20 mg/day is effective, safe, and well tolerated in the treatment of patients with GAD. PMID- 15274173 TI - Efficacy and tolerability of escitalopram in 12- and 24-week treatment of social anxiety disorder: randomised, double-blind, placebo-controlled, fixed-dose study. AB - Selective serotonin reuptake inhibitors are the pharmacological treatment of choice for the treatment of social anxiety disorder (SAD). The efficacy and tolerability of fixed doses of escitalopram were compared to those of placebo in the long-term treatment of generalised SAD, using paroxetine as an active reference. Patients with a DSM-IV diagnosis of SAD between 18-65 years of age were randomised to 24 weeks of double-blind treatment with placebo (n = 166), 5 mg escitalopram (n = 167), 10 mg escitalopram (n = 167), 20 mg escitalopram (n = 170), or 20 mg paroxetine (n = 169). Based on the primary efficacy parameter, Liebowitz Social Anxiety Scale (LSAS) total score at Week 12 (LOCF), a significantly superior therapeutic effect compared to placebo was seen for 5 and 20 mg escitalopram and for all doses for the OC analyses. Further improvement in LSAS scores was seen at Week 24 (OC and LOCF), with significant superiority over placebo for all doses of escitalopram, and 20 mg escitalopram was significantly superior to 20 mg paroxetine. Response to treatment (assessed by a Clinical Global Impression-Improvement score < or = 2) was significantly higher for all active treatments than for placebo at Week 12. Clinical relevance was supported by a significant decrease in all the Sheehan disability scores, and the good tolerability of escitalopram treatment. It is concluded that doses of 5-20 mg escitalopram are effective and well tolerated in the short- and long-term treatment of generalised SAD. PMID- 15274174 TI - Safety and benefits of distance-adjusted prefrontal transcranial magnetic stimulation in depressed patients 55-75 years of age: a pilot study. AB - In contrast to the effects seen in younger adults, depressed elderly subjects have shown more modest antidepressant responses to transcranial magnetic stimulation (TMS). We theorized that higher stimulation intensities in older depressed subjects with prefrontal atrophy might be needed to stimulate underlying cortex. In an open design with patients on stable baseline medications, we treated 18 treatment-resistant elderly depressed subjects (mean age 61.2 +/- 7.3) with 15 rTMS sessions over 3 weeks. We adjusted the delivered TMS intensity to account for MRI measured prefrontal atrophy. The skull to prefrontal cortex distance increased with age, whereas the skull to motor cortex distance did not. All subjects tolerated the higher doses well. The average intensity used was 114% of motor threshold (MT) with a range from 103-141% MT. There was an average 35% decline over the 3 weeks in HRSD scores. After 3 weeks of treatment, 27% (5/18) met response criteria (> 50% improvement), with four of these five also meeting criteria for remission (exit Hamilton Depression Score < 8). These initial pilot findings support the need for blinded studies using prefrontal TMS in an elderly population, testing whether TMS, delivered at stimulation intensities calculated to overcome atrophy, is more effective than TMS without adjusting for atrophy. PMID- 15274175 TI - JNK MAP kinase is involved in the humoral immune response of the greater wax moth larvae Galleria mellonella. AB - We investigated the participation of MAP kinases in the response of Galleria mellonella larvae to immune challenge. JNK MAP kinase was activated in fat body 10-15 min after LPS injection in vivo. The level of JNK activation was time- and LPS dosage-dependent. JNK MAP kinase isolated from cell-free extract of fat bodies dissected from immune stimulated larvae phosphorylated c-Jun protein in vitro. The activity of Gm JNK kinase was abolished in the presence of the JNK specific inhibitor SP600125. Our data indicate a correlation between JNK phosphorylation and induction of antimicrobial activity in the insect hemolymph after immune stimulation. Hemolymph from larvae pre-treated with JNK specific inhibitor SP600125 showed a reduced level of antibacterial activity after LPS injection. JNK inhibition by SP600125 abolished antibacterial activity of the in vitro culture of G. mellonella fat body. Finally, we also show a correlation between JNK-dependent immune response of G. mellonella larvae and elevated temperature. PMID- 15274176 TI - Bombyx mori prothoracicostatic peptide inhibits ecdysteroidogenesis in vivo. AB - Bombyx prothoracicostatic peptide (Bom-PTSP) is a brain neuropeptide that has recently been reported to have in vitro inhibitory activity to prothoracicotropic hormone (PTTH)-stimulated ecdysteroid biosynthesis in the prothoracic gland of the silkworm, Bombyx mori. In the present report, Bom-PTSP has been shown to significantly decrease hemolymph ecdysteroid titer in the fifth instar larvae when Bom-PTSP was injected into the fifth instar day 8 silkworm larvae, resulting in significant delay in spinning behavior. This is the first evidence that Bom PTSP inhibits in vivo ecdysteroidogenesis in the silkworm. PMID- 15274177 TI - Active sites in the carboxyl-terminal region of the laminin alpha chain in Drosophila neuronal cell spreading. AB - An established Drosophila neuronal cell line (BG2-c6) proved to be useful to analyze laminin-mediated cell spreading and signal transduction [Takagi et al. (2000) Biochem Biophys Res Commun 270:482-487]. Here, we report, in addition to the whole molecule, the truncated alpha chain of Drosophila laminin (containing the entire carboxyl-terminal globular domain) and two dodecapeptides corresponding to the cell-binding sites identified in the alpha1 chain of mouse laminin were also active to stimulate BG2-c6 cell spreading. Our previous study [Takagi et al. (1996) J Biol Chem 271:18074-18081] revealed that these recombinant protein and synthetic peptides promoted neurite outgrowth in the primary cell culture system prepared from Drosophila embryo. Therefore, the similar effects by these proteins and peptides suggest the presence of a common mechanism of laminin and neuronal cell interaction working in both primary and established cells. One of the two active peptides contains the sequence SIKVGV. Its murine counterpart carries the sequence SIKVAV by which the interaction of laminin and cells is mediated. Furthermore, laminin-dependent BG2-c6 cell spreading was inhibited by heparin. This observation suggests that cell surface glycoproteins participate in the interaction of laminin and BG2-c6 cells. PMID- 15274178 TI - Fatty acyl-CoA elongation in Blatella germanica integumental microsomes. AB - Insect cuticular hydrocarbons are synthesized de novo in integumental tissue through the concerted action of fatty acid synthases (FASs), fatty acyl-CoA elongases, a reductase, and a decarboxylase to produce hydrocarbons and CO2. Elongation of fatty acyl-CoAs to very long chain fatty acids was studied in the integumental microsomes of the German cockroach, Blatella germanica. Incubation of [1-14C]palmitoyl-CoA, malonyl-CoA, and NADPH resulted in the production of 18 CoA with minor amounts of C20, C22, C24, C30, and C32 labeled acyl-CoA moieties. Similar experiments with [1-14C]stearoyl-CoA rendered C20-CoA as the major product, and lesser amounts of C22 and C24-CoAs were also detected. After solubilization of the microsomal FAS, kinetic parameters were determined radiochemically or by measuring NADPH consumption. The reaction velocity was linear for up to 3 min incubation time, and with a protein concentration up to 0.025 microg/microl. The effect of the chain length on the reaction velocity was compared for palmitoyl-CoA, stearoyl-CoA, and eicosanoyl-CoA. The optimal substrate concentration was 10 microM for C16-CoA, between 8 and 12 microM for C18-CoA, and close to 3 microM for C20-CoA. In vivo hydrocarbon biosynthesis was inhibited from 55.5 to 72.5% in the presence of 1 mM trichloroacetic acid, a known inhibitor of elongation reactions. PMID- 15274179 TI - A neurohormonal role for serotonin in the control of locust oviducts. AB - Serotonin increases the frequency and amplitude of spontaneous contractions and leads to an increase in the basal tonus of the locust oviducts. These effects were dose-dependent and were seen on both the non-innnervated and innervated portion of the oviducts. Vertebrate type serotonin agonists and antagonists were used and the profile shows that the receptors on the non-innervated and innervated portion of the oviducts are more similar to 5-HT3 receptors than to either 5-HT1 or 5-HT2 receptors. No serotonin was found associated with the oviducts or the innervation to the oviducts using immunohistochemistry and HPLC coupled to electrochemical detection, suggesting a neurohormonal role for serotonin in the control of locust oviducts. PMID- 15274180 TI - Dr. Zhong Wei Chen: a microsurgery giant. PMID- 15274181 TI - Microsurgical lip replantation: evaluation of functional and aesthetic results of three cases. AB - Lip amputations are rare, and microsurgical replantation must be systematically tried to restore form and function in one step. The authors present a series of three cases. Revascularization of the amputated segment was obtained by arterial anastomosis with the corresponding labial coronary artery. No venous anastomosis was carried out, because no vein could be identified. Venous drainage was obtained by inducing bleeding and by postoperative application of leeches for 6 days. Anticoagulant therapy and antibiotherapy were used for 10 days. With this approach, two lip amputations were completely saved, and a third amputation only suffered partial necrosis. Aesthetic and functional results were evaluated as being good, with reestablishment of labial continence and recovery of protective sensitivity. PMID- 15274182 TI - Reconstruction of the hand and upper limb with free flaps based on musculocutaneous perforators. AB - Since the advent of perforator flaps, a wide variety of applications have been documented. This study focuses on free flaps based on musculocutaneous perforators, because they have not been well-described in the literature of upper limb reconstruction. They can be trimmed to be thin and pliable, and may provide large flaps with multiple components on the same pedicle to facilitate three dimensional inset of flaps. Microvascular free flaps based on musculocutaneous perforators were performed in 36 cases for reconstruction of the thumb and thenar web, palm, dorsum of the hand, wrist, and forearm. They included the anterolateral thigh perforator flap (27 cases), thoracodorsal perforator flap (5 cases), and deep inferior epigastric perforator flap (4 cases). In 2 other cases not included in this series, the thoracodorsal perforator flap could not be elevated due to anatomical variations. There was no failure in this series, but complications included: 1) hematoma in 2 cases, and 2) infection in 2 cases with flap rim necrosis which was treated by a local rotation flap and skin graft. The thin flaps facilitated secondary reconstructive procedures, and only minor effort was required for the debulking procedure of the flaps. On average, these patients required 2.3 occasions of secondary procedures for further reconstruction following coverage with a perforator flap. The perforator flaps provide medium thickness flaps for coverage of large defects in the upper limb with improved aesthetics and function. With careful dissection of the musculocutaneous perforators and primary thinning of the flaps, the use of a perforator flap is quite safe. Preservation of the muscles leads to better preservation of donor site functions. Less requirement of secondary debulking procedures is a great advantage. However, caution should be taken in the presence of wound infection. PMID- 15274183 TI - Bilateral anterolateral thigh flaps for large-volume breast reconstruction. AB - Autologous tissue reconstruction of a large breast in patients who are not candidates for a TRAM flap is a difficult problem. We present a case report of the use of bilateral free anterolateral thigh (ALT) flaps for immediate reconstruction of a unilateral large breast in a patient who had a previous abdominoplasty. Use of ALT flaps allows two or three surgical teams to work simultaneously, does not require intraoperative patient repositioning, has minimal donor-site morbidity, and can provide ample malleable soft tissue for breast reconstruction. These are advantages compared to the use of gluteal donor sites. The disadvantages include more conspicuous donor-site scarring on the anterior thighs. PMID- 15274184 TI - Role of ulnar forearm free flap in oromandibular reconstruction. AB - The ulnar forearm flap is not frequently utilized for oromandibular reconstruction. This study evaluated the usefulness of the ulnar free flap for reconstruction. A retrospective study of 32 patients was conducted. The ulnar forearm flap was combined with an osseous flap in 24 patients. Nine females and 23 males with a mean age of 58.15 years comprised our study population. Squamous cell carcinoma was the diagnosis in 93.75% of cases (56.25% T4), of which 20% were recurrent. Functional evaluation of swallowing was based on the University of Washington Questionnaire (UWQ). The mean hospital stay was 9.8 days. The external carotid (100%) was the recipient artery, and the internal jugular (74.07%) was the main recipient vein. Overall flap survival was 96.8%. One flap was lost due to unsalvageable venous thrombosis. Major local complications were seen in 9.4% of cases and included partial flap loss, hematoma, and an orocutaneous fistula. At the time of this study, 21 patients were available for functional evaluation. Speech was rated excellent and good in 33.3% of patients. Swallowing was found good in 28.6% of patients. Chewing was rated excellent and good in 47.6% of patients. Cosmetic acceptance was rated good in 71.4% of cases. The ulnar forearm is a useful free flap in oromandibular reconstruction. It is available when the radial artery is the dominant artery of the hand. Being more hidden, it may be more cosmetically accepted. It affords pliable soft tissue for lining and/or covering of oromandibular defects, and can be used as a second choice after other free-flap failures. PMID- 15274185 TI - Change of weight-bearing pattern before and after plantar reconstruction with free anterolateral thigh flap. AB - We reconstructed a large-sized defect at the weight-bearing plantar region by a free anterolateral thigh flap successfully. This is the first case report of using the anterolateral thigh flap for reconstruction of the plantar foot. Based on the preoperative and postoperative pedogram examinations, the pressure distribution on the weight-bearing area reconstructed by the transferred flap was obviously improved and demonstrated a nearly normal pattern. No previous report has compared the weight-bearing pattern before and after large plantar reconstruction with a free flap. The anterolateral thigh free flap, which provides adequate bulk and contour of the foot, and which withstands weight pressure and shearing force and has the ability to provide recovery of sensation, is considered a good alternative in covering a large weight-bearing plantar defect. PMID- 15274186 TI - Microvascular anastomosis through the tibial tunnel: a new technique in free tissue transfer to the leg. AB - Free-tissue transfer to a severely traumatized leg has a high rate of vascular complications. We present three successful cases using a new technique of microvascular anastomosis through the tibial tunnel. Because of the unavailability of anterior tibial artery due to posttraumatic vascular disease, donor vascular pedicles were passed posterior to the tibia through the tibial tunnel and anastomosed to the posterior tibial artery or its branch in an end-to end fashion. The flaps survived perfectly, without any vascular complication. This technique represents a safe route, and the shortest route, to an expected anastomosis point. Our technique is indicated especially in cases with a single vessel leg. PMID- 15274187 TI - Salvage of free flaps after venous thrombosis: case report. AB - Venous thrombosis of a free flap is a serious complication in microsurgery. Several agents with the ability to dissolve an occluding thrombus exist. Recombinant tissue plasminogen activator (rt-PA) seems the most effective. We present our experience with a procedure that was successful in elimination of the occluding thrombus in two patients. PMID- 15274188 TI - Atraumatic suction tip for microsurgery: technical note. AB - During microneurosurgery, frequent suction is essential for a successful operative course. A new self-made disposable suction tip is described which facilitates atraumatic suction, even near vital anatomical structures. The efficacy of this suction tip was confirmed in selected operative procedures. PMID- 15274189 TI - 2-Octylcynanoacrylate-assisted microvascular anastomosis in a rat model: long term biomechanical properties and histological changes. AB - The aim of this study was to establish the long-term biomechanical and histological properties of 2-octylcyanoacrylate-assisted microvascular anastomosis over conventional suture-only anastomosis in the laboratory rat model. The biomechanical and histological properties of three groups of vessels were compared: 1) vessels with 2-octylcyanoacrylate-assisted anastomoses (study group); 2) vessels with suture-only anastomoses (control group); and 3) normal unoperated vessels (sham group). In total, 144 adult rats were used, and these were studied at 1 week, 1 month, 3 months, and 6 months postanastomosis. At 6 months, the tensile strength of study vessels was significantly higher than control vessels. The stiffness of study and control vessels was similar at all time intervals. Histologically, there was no evidence that 2- octylcyanoacrylate caused toxicity to vessel walls, and there was less perivasacular foreign-body giant-cell reaction in the study group compared to the control group. Long-term follow-up showed that microvascular anastomosis with 2-octylcyanoacrylate in rat femoral arteries had superior tensile strength and similar stiffness to vessels anastomosed with sutures only, without adverse effects to surrounding tissues. PMID- 15274190 TI - Use of anti-CD40 ligand monoclonal antibody as antirejection therapy in a murine peripheral nerve allograft model. AB - Monoclonal antibody directed against CD40 ligand prevents acute allograft rejection in several models of solid-organ transplantation. This study describes the use of CD40 ligand as antirejection therapy in a mouse peripheral nerve allograft model. C3H mice received 8-mm nerve isografts (n = 2) or nerve allografts from C57BL donors. Treated animals (n = 11) received anti-CD40 ligand antibody applied to the graft and by intraperitoneal injections postoperatively. At 3 weeks, nerve histology from treated animals was comparable to isografts, whereas untreated allografts demonstrated virtually no signs of regeneration. Walking-track analysis demonstrated a trend toward improved functional recovery in treated animals. In conclusion, blockade of the CD40 pathway suppresses nerve allograft rejection in mice, and facilitates regeneration comparable to isografts. PMID- 15274191 TI - NF-kappaB p65 involves in reperfusion injury and iNOS gene regulation in skeletal muscle. AB - This study investigated the effects of inhibition of NF-kappaB activation on microcirculation and inducible NOS expression in reperfused rat cremaster muscle. The muscle from 16 rats underwent 5-h ischemia and 90-min reperfusion. Each rat received NF-kappaB inhibitor pyrrolidine dithiocarbamate (PDTC, 150 mg/kg) or phosphate-buffered saline 15 min before reperfusion. Results showed that PDTC treatment had a significant overall increase in muscle blood flow during reperfusion. Blood flow more rapidly recovered to and over baseline in the PDTC treated group than in controls, with a significant difference at 10-30 min and 70 90 min. Expression of iNOS mRNA had a 167-fold increase from normal in controls, but was significantly (P < 0.05) reduced to a 63-fold increase in PDTC-treated muscles. In addition, PDTC treatment significantly (P < 0.05) decreased a reperfusion-induced increase in activated NF-kappaB p65 and nuclear p65 protein. Our results suggest that NF-kappaB is involved in I/R injury and that inhibition of NF-kappaB p65 activation affords protection against I/R injury, perhaps via downregulating expression of iNOS transcription. PMID- 15274192 TI - Inhibition of iNOS with 1400W improves contractile function and alters nos gene and protein expression in reperfused skeletal muscle. AB - This study examined the effects of 1400W, an inhibitor of inducible nitric oxide (iNOS), on contractile function and iNOS expression in reperfused skeletal muscle. The right extensor digitorum longus (EDL) muscle of 104 rats underwent a sham operation or 3-h ischemia followed by 3-h or 24-h reperfusion (I/R). Rats received 3 mg/kg 1400W, 10 mg/kg 1400W, or water subcutaneously. Results showed that EDL contractile function in both 1400W-treated groups significantly outperformed the controls at 24-h but not at 3-h reperfusion. Although iNOS expression increased in all three I/R groups during reperfusion, a significantly smaller increase was found in 1400W-treated muscles after 3-h reperfusion, and more dramatically so after 24-h reperfusion. Our results indicate that inhibition of iNOS preserved the contractile function in reperfused skeletal muscle, perhaps via downregulating iNOS expression. Protection by 1400W at 24-h reperfusion suggests that the role of iNOS in exaggerating reperfusion injury is more prominent in the later stages of injury. PMID- 15274193 TI - Free microvascular transfer of coccygeofemoral muscle in rabbits. AB - A new experimental free flap model, the cocygeofemoral muscle flap of the rabbit, is introduced in this study. The coccygeofemoral muscle is a superficially located triangular muscle originating from the sacrum and inserting into the lateral condyle of the tibia, and it abducts the thigh. Anatomical dissection and microangiographic studies revealed that it has a major and a minor pedicle, the former originating from the caudal gluteal artery, and the latter originating from the caudal femoral artery. Its major pedicle contains the 0.7-mm artery, 1 mm vein, and the single nerve originating from the inferior gluteal nerve. In flap studies in 10 animals, the distal minor pedicle was ligated, and muscle was dissected up to its major pedicle and denervated, and the free muscle was transferred to the contralateral inguinal region via end-to-end anastomosis to the recipient vessels (i.e., the femoral artery and vein). At postoperative day 7, the transferred muscles were harvested and histologically examined. Nine of 10 muscles were viable; the coccygeofemoral muscle free flap is believed to be an easy-to-dissect and bulky flap model, having a relatively thick pedicle. It may serve as a good model in free muscle flap studies. PMID- 15274194 TI - Simple and viable in vitro perfusion model for training microvascular anastomoses. AB - In this report, we describe a novel in vitro perfused microvessel model for training microvascular anastomotic exercises. Arteries and veins with a diameter of ca. 1 mm were explanted from chicken wings. These vessels were cannulated at both ends and mounted on a platform. Preserved, expired whole blood obtained from the blood bank was continuously injected through the proximal catheter, using an automatic perfusor. This in vitro perfused microvessel model exactly simulated the viable small-animal vessels. The setting is very simply and reliably repeated; the materials used are very cheap and universally available. There are no ethical questions involved. Vessels explanted from the human placenta or omentum may be used in a similar manner to gain the "feel" of functioning human microvascular tissue. But such materials are rarer and require the approval of ethical committees. PMID- 15274195 TI - Effect of the two-wall-stitch mistake upon patency of rat femoral vein anastomosis: Preliminary observations. AB - Anastomotic patency is believed to be the most important factor in microvascular surgery. The two-wall stitch is a technical error commonly considered to cause thrombosis of the anastomosis, especially on the venous side. In order to demonstrate the real effect on vein patency of the two-wall stitch, the authors performed a standardized mistake after correct microanastomosis on the femoral vein of 15 rats, with one stitch passing through the whole thickness of the two walls of the vein. Traditional correct anastomoses on the contralateral side were used as controls. Patency was assessed at 5, 20, and 60 min and at 24 h by the milking test, and by direct section of the vessel at 24 h. The results showed no statistically significant difference between the two techniques. Histological examination confirmed the clinical judgment about the vessel's patency, and ultrastructural microscopy evidenced only mild signs of endothelial activation. In conclusion, this study indicates that the occasional two-wall stitch does not necessarily increase the risk of venous occlusion in anastomoses of the rat femoral vein. PMID- 15274196 TI - Modified technique of ureteroureterostomy in rat kidney transplantation. AB - Different strain combinations of rats are available to study immunological and transplant-related problems in the models of kidney transplantation. Although numerous modifications of surgical techniques for ureteric reconstruction are evaluated in order to reduce complications and to extend long-term survival, ureteric complications still occur frequently, especially when the difference in diameter of both donor and host ureters is disproportionate. Instead of using the current nonsplinted ureteroureterostomy (method A), a versatile and rapid technical modification (method B) was developed to perform reconstruction of ureters with disproportionate diameters. The overall incidence of ureteric complications was 80% (8/10) using method A, whereas this rate was significantly reduced to 15% (3/20) using method B (P < 0.001). Our modification proves the feasibility of nonsplinted ureteroureterostomy in a technical, highly demanding rat model of kidney transplantation with an acceptable rate of ureteric complication, considering the disproportionate difference in diameter between the host and donor ureters. PMID- 15274197 TI - Thrombotic thrombocytopenic purpura: a syndrome that keeps evolving. PMID- 15274198 TI - Immunotherapy using autologous monocyte-derived dendritic cells pulsed with leukemic cell lysates for acute myeloid leukemia relapse after autologous peripheral blood stem cell transplantation. AB - Although a second stem cell transplantation (SCT) can be used as salvage therapy in patients with relapsing leukemia after SCT, most of these patients have a poor outcome. We tried clinical vaccination using monocyte-derived dendritic cells (DCs) pulsed with leukemic lysates to treat relapsing acute myeloid leukemia (AML) after autologous SCT. To generate DCs, CD14+ cells isolated from peripheral blood stem cell products were cultured in AIM-V in the presence of GM-CSF and IL 4. Adding TNF-alpha on day 6 induced maturation of the DCs, which were harvested on day 8 or 9. The DCs were incubated with tumor lysate and KLH for 2 hr at 37 degrees C. After certifying the absence of microorganisms and endotoxins, the patients received four DC vaccinations at two- to three-week intervals. Two patients received four DC vaccinations with means of 7.8 x 10(6) and 9 x 10(6) DCs at two- to three-week intervals. The DC vaccinations were well tolerated with no apparent side effects. After the vaccinations, the patients showed immunological responses with positive delayed-type hypersensitivity skin reaction and increasing autologous T cells stimulatory capacity to the DCs; however, the BM blast percentage of the patients did not improve. The results suggest that DCs are a feasible cellular therapy for relapsing AML after autologous SCT. PMID- 15274199 TI - Improved treatment of sudden hearing loss by specific fibrinogen aphaeresis. AB - The etiology of sudden sensorineural hearing loss is still unclear and is thought to result from disturbances of microcirculation, infectious causes, or autoimmune disorders. So far standard therapy did not show clear improvement over spontaneous remission rate, which is assumed to be about 50% [Nakashima et al., Acta. Otolaryngol. Stockh. 514:14-16, 1994; Schuknecht and Donovan, Arch. Otorhinolaryngol. 243:1-15, 1986; Harris and Sharp, Laryngoscope 100:516-524, 1990; Mayot et al., Clin. Immunol. Immunopath. 68:41-45, 1993; Gussen, Ann. Otol. Rhinol. Laryngol. 85:94-100, 1976]. Elevated blood viscosity due to high fibrinogen levels is supposed to cause decreased cochlear blood flow and thus initiate sudden hearing loss. The specific lowering of fibrinogen immediately decreases plasma viscosity exactly to the desired extent and should lead to improved cochlear blood flow [Suckfull et al., Acta. Otolaryngol 119:763-766, 1999; Suckfull, Lancet 360:1811-1817, 2002; Walch et al., Laryngol. Rhino. Otol. 75:641-645, 1996; Suckfull et al., Otol. Neurotol. 23:309-311, 2002]. In a prospective uncontrolled pilot study on 36 patients with unilateral sudden onset sensorineural hearing loss (SHL) we tried to establish that 1-3 specific fibrinogen aphaereses alone improve recovery of hearing and that it is possible to lower fibrinogen to the target of 80-100 mg/dl without important side effects. Pure tone audiometry was carried out immediately before and after each aphaeresis as well as at 2 and 4 weeks and 6 months after treatment. Sixteen patients recovered spontaneously before undergoing fibrinogen adsorption. All 20 aphaeresis patients improved during immunoadsorption; in 60% of patients auditory thresholds returned to normal after the first immunoadsorption and treatment could be discontinued, in another 20% of patients complete recovery was reached after 4 weeks. The mean plasma fibrinogen concentration of the 20 patients before the first aphaeresis session was 308.1 +/- 51.5 mg/dl. Immediately after the first treatment session, the fibrinogen concentration was lowered to 100.7 +/- 25.3 mg/dl (P < 0.001). The second and third sessions also showed highly significant reductions in plasma fibrinogen. No important side effects were seen. In conclusion, specific fibrinogen adsorption is a promising new treatment modality that should be tested in a prospective, randomized controlled trial in patients with sudden hearing loss. PMID- 15274200 TI - Plasma exchange conditioning for ABO-incompatible renal transplantation. AB - The supply of deceased donor kidneys is inadequate to meet demand. To expand the pool of potential donors, ABO-incompatible transplants from living donors have been performed. We present the Mayo Clinic experience with such transplants. Enrollment was open to patients when the only available potential living kidney donor was ABO-incompatible. Conditioning consisted of plasma exchanges followed by intravenous immunoglobulin. Splenectomy was performed at the time of transplant surgery. Post-transplant immunosuppression consisted of anti-T lymphocyte antibody, tacrolimus, mycophenolate mofetil, and prednisone. Isoagglutinin titers and scores were determined before and after each plasma exchange. Transplant outcomes were determined. Twenty-six ABO-incompatible transplants were performed. No hyperacute rejection occurred. Mean patient follow up was 400 days. Patient and graft survivals at last follow-up were 92 and 85%, respectively. Antibody-mediated rejection occurred in 46% and was apparently reversed in 83% by plasma exchange and increased immunosuppression. The initial plasma exchange reduced immediate spin and AHG hemagglutination reactivity scores by 53.5 and 34.6%, respectively. Over the course of the pretransplant plasma exchanges, the immediate spin and AHG hemagglutination reactivity scores decreased by 96.4 and 68.5%, respectively. At 3 and 12 months, the immediate spin and AHG hemagglutinin reactivity scores and titers were less than those at baseline but greater than or equal to those on the day of transplantation. Despite an increase in scores and titers, antibody-mediated rejection was not present. Pre-transplant plasma exchange conditioning combined with other immunosuppressives can be used to prepare patients for ABO-incompatible kidney transplantation from living donors, but antibody-mediated rejection post transplant is a common occurrence and allograft survival may be reduced. Controlled clinical trials are needed to identify the optimum conditioning for ABO-incompatible renal transplants. PMID- 15274201 TI - Relapsed thrombotic thrombocytopenic purpura presenting as an acute cerebrovascular accident. AB - Thrombotic thrombocytopenic purpura (TTP) is an uncommon but severe disorder that classically presents with microangiopathic hemolytic anemia (MAHA), thrombocytopenia, and fluctuating neurological changes. Previously, it was impossible to make a diagnosis of TTP in the absence of thrombocytopenia or microangiopathic hemolysis (MAHA). We describe two cases of relapsing TTP that presented with acute cerebrovascular accident (CVA) without concurrent thrombocytopenia or MAHA after initial classical presentation of TTP. In both cases, the diagnosis of TTP as the cause of the CVA was attributed to severe deficiency of the von Willebrand factor cleaving protease, ADAMTS13 in plasma (11 and 12%, normal 79-127%). Each patient had a dramatic clinical improvement in response to therapeutic plasma exchange. The experience in these two cases suggests that TTP should be considered as a potential cause among patients presenting with a CVA, particularly if the patients have a history of TTP. PMID- 15274202 TI - Effect of different LDL-apheresis methods on parameters involved in atherosclerosis. AB - LDL-apheresis lowers the LDL level in patients with severe hypercholesterolemia. The present study compared three apheresis methods--DSA, DALI, and plasma exchange--for effectiveness in removal of LDL and effect on various blood parameters involved in atherogenesis. The study group included 6 patients with primary hypercholesterolemia unresponsive to maximal drug therapy. All were treated first with 4 consecutive plasma exchange sessions followed by 4 DSA sessions; in four out of six, an additional 4 sessions of DALI were then performed. Levels of lipoproteins, apoproteins, CRP, homocysteine, fibrinogen, and blood count were determined before and after each session. All 3 procedures yielded a significant reduction in total cholesterol and ApoB-containing lipoproteins, with DALI being the most effective. Also, a significant reduction in triglycerides, HDL, and ApoA1 was observed with all the methods. The reduction in HDL-C with DSA and DALI was greater than previously reported. The LDL/HDL cholesterol ratio decreased significantly with DSA and DALI and increased with plasma exchange. There was a significant decrease in CRP, fibrinogen, and platelets with all three methods, and a significant decrease in homocysteine only with DSA and DALI. All three procedures effectively reduced the concentration of various compounds involved in atherosclerosis. Plasma exchange is nonselective and cannot be recommended as the procedure of choice. PMID- 15274203 TI - L-carnitine improves pH and decreases surface phosphatidylserine expression in extended stored apheresis platelets. AB - Extension of the storage period of apheresis platelets to seven or ten days may be possible with the implementation of screening for bacteria. This, however, may impair platelet quality, and additive compounds that improve storage parameters would be desirable. Apheresis platelets were harvested using the Cobe LRS device. Part of the product was aliquoted into two CLX bags, 60 ml into each, on day 0. L carnitine (LC) to a final concentration of 5 mM was added to one container and saline to the other. pH, morphology score, and surface expression of phosphatidylserine were measured on day 1, and, in addition, hypotonic shock response (HSR) and the extent of shape change (ESC) on days 5, 10, and 13. Differences between test and controls were analyzed using paired t-tests. The addition of LC improved pH by day 5, but was more evident by days 10 and 13. By day 10, significant differences (<0.01) were observed in pH (6.54 +/- 0.3 vs. 6.75 +/- 0.3), lactate (176 +/- 31 vs. 150 +/- 24 mg %), morphology score (213 +/ 27 vs. 229 +/- 35) and ESC (7 +/- 6 vs. 11 +/- 6). Percent surface phosphatidylserine expression was less in the LC treated platelets (16 +/- 7 vs. 12 +/- 4, P<0.03). Much of the benefit observed was attributable to improved parameters in some donors. LC improves the quality of extended stored apheresis platelets. PMID- 15274204 TI - Hematopoietic progenitor cell large volume leukapheresis (LVL) on the Fenwal Amicus blood separator. AB - A technique for large volume leukapheresis (LVL) for hematopoietic progenitor cell (HPC) collection using the Fenwal Amicus is presented. It was compared to standard collections (STD) with regard to CD34+ cell yields and cross-cellular content. Optimal cycle volumes and machine settings were evaluated for LVL procedures. A total of 68 patients underwent 80 HPC collection procedures. Because of differences in CD34+ cell yields associated with peripheral white blood cell counts (WBC), the comparison was divided into groups of 20 with WBC < or =35 x 10(9)/L (< or =35 K) and those >35 x 10(9)/L (>35 K). Baseline CD34+ cell counts (peripheral count when patient started HPC collection) were used (median 18-23 cells/microl). Significantly more whole blood (corrected for anticoagulant) was processed with LVL (LVL 20 l vs. STD 13.5 l). For < or =35 K, median CD34+ x 10(6), WBC x 10(9), RBC ml, Plt x 10(11) yields/collection were 183, 21.2, 14, 0.8, respectively, for STD vs. 307, 22.1, 11, 1.0, respectively, for LVL. For >35 K, median CD34+ x 10(6), WBC x 10(9), RBC ml, Plt x 10(11) yields/collection were 189, 32.7, 15, 1.4, respectively, for STD vs. 69, 40.8, 21, 1.3, respectively, for LVL. We have described a method of LVL using the Amicus that, in patients with pre-procedure WBC < or =35 x 10(9)/L, collects more CD34+ cells than a standard procedure with acceptable cross-cellular content. This method is not recommended when pre-procedure WBC counts are >35 x 10(9)/L. PMID- 15274208 TI - Mild methods for removing organic templates from inorganic host materials. PMID- 15274209 TI - Stable cyclopentynes--made by metals!? AB - Structural limitations in small cyclic hydrocarbons are a longstanding challenge to chemistry, leading to molecules with unusual geometric parameters and reactivity. For example, incorporation of C==C==C==C or C-C[triple chemical bond]C-C arrangements, which are normally linear in open-chain compounds, into small carbocyclic rings results in enormous ring strain as a result of the angle deformation. Nevertheless, such uncommon structures have been realized by complexation of zirconocene fragments with butadiynes and butatrienes, giving five-membered metallacyclocumulenes and metallacyclopentynes. The metal atom is essential for these molecules but complicates the simplified description because of internal interactions of the metals with the multiple bonds. PMID- 15274210 TI - Nickel-catalyzed reductive cyclizations and couplings. AB - For over 50 years, nickel catalysis has been applied in cycloaddition processes. Nickel-catalyzed reductive couplings and cyclizations, however, have only recently attracted a high level of interest. This group of new reactions allows a broad range of multicomponent couplings involving two or more pi components with a main-group or transition-metal reagent. These processes allow the assembly of important organic substructures from widely available reaction components. Multiple contiguous stereocenters, polycyclic ring systems, and novel arrays of complex functionality may often be prepared from simple, achiral, acyclic precursors. With three or more reactive functional groups participating in the catalytic processes, many mechanistic questions abound, including the precise timing of bond constructions and the nature of reactive intermediates. This Review is thus aimed at providing a critical evaluation of recent progress in this rapidly developing field. PMID- 15274211 TI - Synthesis, structure, and magnetic properties of a large lanthanide-transition metal single-molecule magnet. PMID- 15274212 TI - A 3D interlocked structure from a 2D template: structural requirements for the assembly of a square-planar metal-coordinated [2]rotaxane. PMID- 15274213 TI - Identification of the 5-methylthiopentosyl substituent in Mycobacterium tuberculosis lipoarabinomannan. PMID- 15274214 TI - Triggering of RNA secondary structures by a functionalized nucleobase. PMID- 15274215 TI - Stable and selective recognition of three base pairs in the parallel triple helical DNA binding motif. PMID- 15274217 TI - Reaction of alkene-zirconocene complexes and cyclic enol ethers through new reaction pathways. PMID- 15274216 TI - Efficiency and fidelity in a click-chemistry route to triazole dendrimers by the copper(i)-catalyzed ligation of azides and alkynes. PMID- 15274218 TI - Arynes in a three-component coupling reaction: straightforward synthesis of benzoannulated iminofurans. PMID- 15274219 TI - Switchable electron-transfer processes in a mixed-valence, kinetically locked, trinuclear Ru(II) metallamacrocycle. PMID- 15274220 TI - Asymmetric catalytic coupling of organoboranes, alkynes, and imines with a removable (trialkylsilyloxy)ethyl group--direct access to enantiomerically pure primary allylic amines. PMID- 15274221 TI - Cationic planar chiral palladium P,S complexes as highly efficient catalysts in the enantioselective ring opening of oxa- and azabicyclic alkenes. PMID- 15274222 TI - Stereoselective total synthesis of (-)-borrelidin. PMID- 15274224 TI - Cross-coupling of alkyl halides with aryl grignard reagents catalyzed by a low valent iron complex. PMID- 15274223 TI - General catalytic synthesis of highly enantiomerically enriched terminal aziridines from racemic epoxides. PMID- 15274225 TI - Catalytic asymmetric intramolecular Michael reaction of aldehydes. PMID- 15274226 TI - Left-handed helical twists in "mixed beta-peptides" derived from alternating C linked carbo-beta3-amino acids and beta-hGly units. PMID- 15274227 TI - Synthesis and self-assembly of perylenediimide-oligonucleotide conjugates. PMID- 15274228 TI - Selenol nitrosation and Se-nitrososelenol homolysis: a reaction path with possible biochemical implications. PMID- 15274229 TI - Design, synthesis, and biological evaluation of a small-molecule inhibitor of the histone acetyltransferase Gcn5. PMID- 15274231 TI - [Internist's therapy of rheumatoid arthritis]. AB - Rheumatoid arthritis is the most common inflammatory joint disease and is characterized by chronic, symmetric, erosive synovitis of small joints of hands and feet. Prevalence in women is threefold higher than in man. Structural damage of the joints starts between the first and second year of the disease. Early therapeutic interventions can alter the course of rheumatoid arthritis by delaying the progression of radiographic joint destruction, which correlates with the grade of disability. Approval of new biologic antirheumatic drugs in the last few years improved the outcome of rheumatoid arthritis. PMID- 15274233 TI - Effects on boy-attracted pedosexual males of viewing boy erotica. PMID- 15274234 TI - Repairing homophobics. PMID- 15274235 TI - Avascular necrosis after surgical treatment for development dysplasia of the hip. AB - We reviewed the medical records of 101 patients with developmental dysplasia of the hip who were treated with Dega's (102 hips), or Salter's (42 hips)osteotomy preceded by open reduction and femoral intertrochanteric osteotomy. The minimal follow-up was 17 years. At the last follow-up, there were proximal fem-oral growth disturbances in 52 hips (36%). In 20 hips, the disturbances were graded as mild and in six as severe. We found significantly better clinical and radiological results in hips without avascular changes. Risk factors for the development of avascular necrosis were: involvement of the left side and surgical treatment initiated after 2 years of age without pre-operative traction and without femoral shaft shortening. We found that the incidence of avascular necrosis increased with the length of follow-up. The avascular necrosis influenced both clinical and radiological results. PMID- 15274236 TI - Total knee replacement influences both knee and hip joint kinematics during stair climbing. AB - A gait analysis system was used to evaluate the kinematics of the hip and knee during stair ascending and descending after operation with total knee replacement. Patients with 5 degrees varus/valgus alignment or less were selected randomly to receive either a flat or a concave tibial component with retention of the posterior cruciate ligament. Patients who had more than 5 degrees varus/valgus alignment and/or an extension defect of 10 degrees or more were selected randomly to receive the concave or posterior-stabilized tibial component with resection of the posterior cruciate ligament. Twenty patients and 17 controls were studied 1-2 years after the operation. Patients had abnormal kinematics during stair ascending and descend-ing. Both knee extension and flexion were reduced. Hip extension tended to decrease, and decreased hip extension moment was noted. PMID- 15274237 TI - Distractible vertebral body replacement in patients with malignant vertebral destruction or osteoporotic burst fractures. AB - We studied prospectively the clinical and radiological course of 36 patients with localised spinal instability treated with vertebrectomy. A distractible titanium cage filled with polymethylmethacrylate(PMMA) in combination with transpedicular fixation was used to restore spinal stability. Mean follow-up was 16 (8-55) months. Average correction of the segmental kyphosis was 15.2 degrees (-2-29 degrees). During follow-up, a loss of correction of 0.9 degrees (-2-15 degrees) was seen. Segmental height was increased by 7.6 (0-14) mm on average with a mean loss of 1.6 (0-2) mm. Twenty-one patients without pre-operative neurological deficits were ambulatory without orthosis at discharge. Patients presenting with neurological deficits were unchanged (n=3) or improved (n=12). PMID- 15274238 TI - Modular total shoulder system with short stem. A prospective clinical and radiological analysis. AB - Between 1994 and 2001, a short-stemmed modular shoulder prosthesis was inserted in 62 shoulders in patients with rheumatoid arthritis (RA) or osteoarthrosis (OA). We reviewed 53 patients with 60 shoulders (45 RA/15 OA) with at least 24 months follow-up. In 22 shoulders, we used a total shoulder prosthesis including a glenoid polyethylene component, whereas 38 shoulders only had a humeral component. In six shoulders, the humeral component was cemented. The average follow-up was 47 (24-99) months. There were no intraoperative complications but one wound infection and one patient with proximal migration of the humeral component. Hospital for Special Surgery Score increased from 44(19-72) to 63 (21 93) points and Shoulder Function Assessment score (SFA) from 24 (12-46) to 42 (11 66)points. The VAS score for pain at rest improved from 4.3 to 1.9. Nonprogressive radiolucent lines were seen adjacent to nine glenoid and one humeral components. Fifty-six patients were satisfied with the result. PMID- 15274239 TI - Re: International Orthopaedics, vol 27, (suppl 1) 2003. PMID- 15274240 TI - Politics dressed as science: two think tanks on environmental regulation and health. PMID- 15274241 TI - [Life picture. Karlheinz Engelhardt 29 November 1930]. PMID- 15274242 TI - [HIV/HCV double infection: combination is a clear therapy option]. PMID- 15274243 TI - [Rapidly and without pain: endovenous laser therapy for varicose veins]. PMID- 15274244 TI - [For which skin diseases would lasers be used?]. PMID- 15274245 TI - Biodegradation of poly(L-lactide). AB - The biodegradation of poly(L-lactide) (PLA) is reviewed. The important role of actinomycetes in PLA degradation is emphasized. These PLA-degrading actinomycetes belong phylogenetically to the Pseudonocardiaceae family and related genera, including Amycolatopsis, Lentzea, Streptoalloteichus, Kibdelosporangium and Saccharothrix. A PLA-degrading enzyme purified from an isolated Amycolatopsis strain-41 has substrate specificity on PLA higher than proteinase K. The application of these strains and their enzymes can be effectively used for biological treatment of plastic wastes containing PLA. PMID- 15274246 TI - Nonlethal development, validation, and application of cytochrome P4501A1 (CYP1A1) as a biomarker for contaminant exposure. PMID- 15274247 TI - [Common disorders and diseases of the nails. Anatomy, physiology, disorders, clarification and therapy]. AB - Disorders of anatomical variants of the nails are often seen in clinical practice. Their correction identification and interpretation are essential dermatological skills. The increasing emphasis on aesthetic considerations in dermatology means that even the slightest nail changes may assume significance for their patients. In order to skillfully evaluate nails, one must be familiar with the terminology and classification of nail disorders. The nail plate, nail bed and periungual tissue may also be affected. Longitudinal melanonychia is especially important as a possible clue to subungual melanoma. PMID- 15274248 TI - p-Bromophenacyl bromide prevents cumene hydroperoxide-induced mitochondrial permeability transition by inhibiting pyridine nucleotide oxidation. AB - Mitochondrial permeability transition is commonly characterized as a Ca2+ dependent non-specific increase in inner membrane permeability that results in swelling of mitochondria and their de-energization. In the present study, the effect of different inhibitors of phospholipase A2--p-bromophenacyl bromide, dibucaine, and aristolochic acid--on hydroperoxide-induced permeability transitions in rat liver mitochondria was tested. p-Bromophenacyl bromide completely prevented the hydroperoxide-induced mitochondrial permeability transition while the effects of dibucaine or aristolochic acid were negligible. Organic hydroperoxides added to mitochondria undergo reduction to corresponding alcohols by mitochondrial glutathione peroxidase. This reduction occurs at the expense of GSH which, in turn, can be reduced by glutathione reductase via oxidation of mitochondrial pyridine nucleotides. The latter is considered a prerequisite step for mitochondrial permeability transition. Among all the inhibitors tested, only p-bromophenacyl bromide completely prevented hydroperoxide-induced oxidation of mitochondrial pyridine nucleotides. Interestingly, p-bromophenacyl bromide had no affect on mitochondrial glutathione peroxidase, but reacted with mitochondrial glutathione that prevented pyridine nucleotides from being oxidized. Our data suggest that p-bromophenacyl bromide prevents hydroperoxide-induced deterioration of mitochondria via interaction with glutathione rather than through inhibition of phospholipase A2. PMID- 15274249 TI - The doctors of the Hartford Hospital 1854-1954. 1954. PMID- 15274251 TI - [An addition to the case report "Adenocarcinoma of the small intestine" by Oliverius and Wohl published in Rozhledu v chirurgii, no. 10, 2003]. PMID- 15274252 TI - [Proceedings of the Acute Coronary Syndrome Symposium--Myocardial Infarct and Unstable Angina. 4 July 2003, Zagreb]. PMID- 15274253 TI - The late consequences of scaphoid fractures. PMID- 15274254 TI - The management of slipped capital femoral epiphysis. PMID- 15274255 TI - Ortho-geriatric liaison--the missing link? PMID- 15274256 TI - Mechanical prophylaxis of deep-vein thrombosis after total hip replacement a randomised clinical trial. AB - Routine prophylaxis for venous thromboembolic disease after total hip replacement (THR) is recommended. Pneumatic compression with foot pumps seems to provide an alternative to chemical agents. However, the overall number of patients investigated in randomised clinical trials has been too small to draw evidence based conclusions. This randomised clinical trial was carried out to compare the effectiveness and safety of mechanical versus chemical prophylaxis of DVT in patients after THR. Inclusion criteria were osteoarthritis of the hip and age less than 80 years. Exclusion criteria included a history of thromboembolic disease, heart disease, and bleeding diatheses. There were 216 consecutive patients considered for inclusion in the trial who were randomised either for management with the A-V Impulse System foot pump. We excluded 16 patients who did not tolerate continuous use of the foot pump or with low-molecular-weight heparin (LMWH). Patients were monitored for DVT using serial duplex sonography at 3, 10 and 45 days after surgery. DVT was detected in three of 100 patients in the foot pump group and with six of 100 patients in the LMWH group (p < 0.05). The mean post-operative drainage was 259 ml in the foot-pump group and 328 ml in the LMWH group (p < 0.05). Patients in the foot-pump group had less swelling of the thigh (10 mm compared with 15 mm; p < 0.05). One patient developed heparin-induced thrombocytopenia. This study confirms the effectiveness and safety of mechanical prophylaxis of DVT in THR. Some patients cannot tolerate the foot pump. PMID- 15274257 TI - Total hip arthroplasty using the Wroblewski golf ball cup inserted through the posterior approach. A high rate of dislocation. AB - The Wroblewski golf ball acetabular cup was introduced by surgeons using the trochanteric osteotomy approach for revision total hip replacement (THR) in order to reduce the rate of dislocation. We have routinely used the Ogee long posterior wall (Ogee LPW) and the Wroblewski angle bore cups in THR. Although the new Wroblewski golf ball cup performed well there was a significant early rate of dislocation of 20%. Our rate of dislocation over a period of ten years using the Ogee LPW and Wroblewski angle bore cups had been 0.52%. We present our findings and an investigation as to why the new cup has such a high rate of dislocation when used with the posterior approach. We show that a relatively small change in the design of the acetabular component resulted in significant adverse clinical results. PMID- 15274258 TI - Pelvic remodelling after the Chiari osteotomy. A long-term review. AB - Survivorship analysis of 215 medial displacement pelvic osteotomies undertaken for symptomatic, incongruent dysplasia of the hip since 1966 showed that four of every five hips had not required conversion to a total hip arthroplasty. The radiological characteristics of 86 osteotomies were evaluated at a mean of 18 years (5 to 30) after surgery which was performed at the age of 15.9 +/- 9.5 years. Revision was significantly (p < 0.05) more likely in those patients operated on after the age of 25 years. The centre-edge (CE) angle increased from 2.5 +/- 13.9 degrees before to 41.8 +/- 15.0 degrees immediately after operation. The increase in CE angle was maintained at later review (38.5 +/- 16.5 degrees). Even with severe dysplasia with a CE angle less than zero a substantial improvement in the cover of the femoral head was achieved, usually by medial shift of the lower pelvic fragment. However, the head was not invariably medialised by the osteotomy and lateral movement of the ilium was noted when the position of the joint was relatively medial before operation or when the hip was arthritic. In the longer term pelvic remodelling did not reverse the medialisation produced by the osteotomy, and the cover of the femoral head was maintained. PMID- 15274259 TI - Footballer's hip a report of six cases. AB - We have reviewed a consecutive series of six professional footballers who presented with intractable hip pain which was attributed at arthroscopy, to an anterior acetabular labral tear with adjacent chondral damage. The location and type of labral tear were identical for each patient. There was no evidence of acetabular dysplasia. A variable area of chondral damage was associated with the labral tear. All unstable tissue was resected. Five returned to professional football at the highest level. Acetabular labral pathology should be included in the differential diagnosis of footballers with hip or groin pain. Arthroscopy of the hip is an appropriate method of diagnosis and treatment and minimises the length of rehabilitation required. PMID- 15274260 TI - Foot function after subtalar distraction bone-block arthrodesis. A prospective study. AB - Subtalar distraction bone-block arthrodesis for malunited calcaneal fractures was performed in 31 patients (26 men, five women), with a mean age of 38.5 years. The mean time from injury to arthrodesis was 36 months. There were no cases of nonunion. One patient had an early dislocation of the bone block requiring a repeat arthrodesis, and one had a soft-tissue infection. The mean AOFAS hindfoot score improved significantly from 23.5 before operation to 73.2 at a mean follow up of 33 months (p > 0.001). Compared with the unaffected side, the talocalcaneal height was corrected by 61.8%, the talus-first metatarsal axis by 46.5%, the talar declination angle by 38.5% and the talocalcaneal angle by 35.4%. Dynamic pedobarography revealed a return to normal of the pressure distribution during roll-over and a more energetic gait. The distribution of local transfer of load correlated well with the AOFAS score. The amount of correction of the heel height correlated with a normal pattern of pressure transfer on the heel (p < 0.05). PMID- 15274261 TI - Hallux valgus and cartilage degeneration in the first metatarsophalangeal joint. AB - This study relates the extent of cartilage lesions within the first metatarsophalangeal joint to hallux valgus. We prospectively examined 265 first metatarsophalangeal joints of 196 patients with a mean age of 54.2 years at operation for the existence of cartilage lesions. Grade I lesions were found in 41 feet (15.5%), grade II in 82 (30.9%), grade III in 51 (19.3%), grade IV in 20 (7.5%). Only 71 (26.8%) showed no cartilage lesion. Cartilage lesions were found within the metatarsosesamoid and metatarsophalangeal compartments in 66 feet (34.0%), within the metatarsophalangeal compartment in 26 (13.4%) and within the metatarsosesamoid compartment in 102 (52.6%). A statistically significant correlation was found between the grade of cartilage lesion and the hallux valgus angle, both for the changes within the metatarsophalangeal and the metatarsosesamoid joints. PMID- 15274262 TI - The posteromedial corner revisited. An anatomical description of the passive restraining structures of the medial aspect of the human knee. AB - We have reviewed the literature on the anatomy of the posteromedial peripheral ligamentous structures of the knee and found differing descriptions. Our aim was to clarify the differing descriptions with a simplified interpretation of the anatomy and its contribution to the stability of the knee. We dissected 20 fresh frozen cadaver knees and the anatomy was recorded using video and still digital photography. The anatomy was described by dividing the medial collateral ligament (MCL) complex into thirds, from anterior to posterior and into superficial and deep layers. The main passive restraining structures of the posteromedial aspect of the knee were found to be superficial MCL (parallel, longitudinal fibres), the deep MCL and the posteromedial capsule (PMC). In the posterior third, the superficial and deep layers blend. Although there are oblique fibres (capsular condensations) running posterodistally from femur to tibia, no discrete ligament was seen. In extension, the PMC appears to be an important functional unit in restraining tibial internal rotation and valgus. Our aim was to clarify and possibly simplify the anatomy of the posteromedial structures. The information would serve as the basis for future biomechanical studies to investigate the contribution of the posteromedial structures to joint stability. PMID- 15274263 TI - Alignment in total knee arthroplasty. A comparison of computer-assisted surgery with the conventional technique. AB - Restoration of neutral alignment of the leg is an important factor affecting the long-term results of total knee arthroplasty (TKA). Recent developments in computer-assisted surgery have focused on systems for improving TKA. In a prospective study two groups of 80 patients undergoing TKA had operations using either a computer-assisted navigation system or a conventional technique. Alignment of the leg and the orientation of components were determined on post operative long-leg coronal and lateral films. The mechanical axis of the leg was significantly better in the computer-assisted group (96%, within +/- 3 degrees varus/valgus) compared with the conventional group (78%, within +/- 3 degrees varus/valgus). The coronal alignment of the femoral component was also more accurate in the computer-assisted group. Computer-assisted TKA gives a better correction of alignment of the leg and orientation of the components compared with the conventional technique. Potential benefits in the long-term outcome and functional improvement require further investigation. PMID- 15274264 TI - Infection after total knee arthroplasty. AB - The aim of our study was to determine the current incidence and outcome of infected total knee arthroplasty (TKA) in our unit comparing them with our earlier audit in 1986, which had revealed infection rates of 4.4% after 471 primary TKAs and 15% after 23 revision TKAs at a mean follow-up of 2.8 years. In the interim we introduced stringent antibiotic prophylaxis, and the routine use of occlusive clothing within vertical laminar flow theatres and 0.05% chlorhexidine lavage during arthroplasty surgery. We followed up 931 primary TKAs and 69 revision TKAs for a mean of 6.5 years (5 to 8). Patients were traced by postal questionnaire, telephone interview or examination of case notes of the deceased. Nine (1%) of the patients who underwent primary TKA, and four (5.8%) of those who underwent revision TKA developed deep infection. Two of nine patients (22.2%) who developed infection after primary TKA were successfully treated without further surgery. All four of the patients who had infection after revision TKA had a poor outcome with one amputation, one chronic discharging sinus and two arthrodeses. Patients who underwent an arthrodesis had comparable Oxford knee scores to those who underwent a two-stage revision. Although infection rates have declined with the introduction of prophylactic measures, and more patients are undergoing TKA, the outcome of infected TKA has improved very little. PMID- 15274265 TI - Glenohumeral arthrodesis in upper and total brachial plexus palsy. A comparison of functional results. AB - We have compared the functional outcome after glenohumeral fusion for the sequelae of trauma to the brachial plexus between two groups of adult patients reviewed after a mean interval of 70 months. Group A (11 patients) had upper palsy with a functional hand and group B (16 patients) total palsy with a flail hand. All 27 patients had recovered active elbow flexion against resistance before shoulder fusion. Both groups showed increased functional capabilities after glenohumeral arthrodesis and a flail hand did not influence the post operative active range of movement. The strength of pectoralis major is a significant prognostic factor in terms of ultimate excursion of the hand and of shoulder strength. Glenohumeral arthrodesis improves function in patients who have recovered active elbow flexion after brachial plexus palsy even when the hand remains paralysed. PMID- 15274266 TI - Genetic influences in the aetiology of tears of the rotator cuff. Sibling risk of a full-thickness tear. AB - From a retrospective, cohort study of 205 patients diagnosed with full-thickness tears of the rotator cuff, we determined, using ultrasound, the prevalence of such tears in their 129 siblings. Using 150 spouses as controls, the relative risk of full-thickness tears in siblings versus controls was 2.42 (95% CI 1.77 to 3.31). The relative risk of symptomatic full-thickness tears in siblings versus controls was 4.65 (95% CI 2.42 to 8.63). The significantly increased risk for tears in siblings implies that genetic factors play a major role in the development of full-thickness tears of the rotator cuff. PMID- 15274267 TI - A comparison of open and percutaneous techniques in the surgical treatment of tennis elbow. AB - We conducted a prospective, randomised, controlled trial of 45 patients (47 elbows), with tennis elbow, who underwent either a formal open release or a percutaneous tenotomy. All patients had pre- and post-operative assessment using the Disability of Arm, Shoulder and Hand (DASH) scoring system. Both groups were followed up for a minimum of 12 months. Statistical analyses using the Mann Whitney U test and repeated measured ANOVA showed significant improvements for patient satisfaction (p = 0.012), time to return to work (p = 0.0001), improvements in DASH score (p = 0.001) and improvement in sporting activities (p = 0.046) in the percutaneous group. Those patients undergoing a percutaneous release returned to work on average three weeks earlier and improved significantly more quickly than those undergoing an open procedure. The percutaneous procedure is a quicker and simpler procedure to undertake and produces significantly better results. PMID- 15274268 TI - MRI and plain radiography in the assessment of displaced fractures of the waist of the carpal scaphoid. AB - We treated 50 patients with fractures of the waist of the scaphoid in a below elbow plaster cast for up to 13 weeks. Displacement of the fragments was assessed independently by two observers using MRI and radiographs performed within two weeks of injury. The MRI assessments showed that only the measurement of sagittal translation of the fragments and an overall assessment of displacement had satisfactory inter- and intra-observer reproducibility and revealed that nine of the 50 fractures were displaced. Only three of the 49 fractures with adequate follow-up failed to unite, and all were displaced with more than 1 mm of translation in the sagittal plane. If the MRI assessment of displacement of the fracture was used as the measurement of choice, assessment of displacement on the initial scaphoid series of radiographs showed a sensitivity of between 33% and 47% and a positive predictive value of between 27% and 86%. Neither observer was able correctly to identify more than 33% to 47% of the displaced fractures from the plain radiographs. Although the overall assessment of displacement and gapping and translation in the coronal plane on the plain radiographs influenced the rate of union, none of these parameters identified all three fractures which failed to unite. We conclude that the assessment of displacement of scaphoid fractures on MRI can probably be used to assess the likelihood of union although the small number of nonunions limits the power of the study. In contrast, the assessment of displacement on routine radiography is inaccurate and of less value in predicting union. PMID- 15274269 TI - Closed argon-based cryoablation of bone tumours. AB - We report our experience with a new technique for cryosurgical ablation of bone tumours which allows accurate determination of the temperature and freezing time within a cavity of any geometrical shape. Between 1997 and 2000, 58 patients diagnosed with 13 malignant and 45 aggressive benign bone tumours underwent argon based cryoablation. This technique includes removal of the tumour by curettage and filling the cavity with a gel medium into which metal probes are inserted. Argon gas is delivered through the metal probes and both time and temperature are computer-controlled. After formal reconstruction, all patients were followed for more than two years. None had skin necrosis, infection, neurapraxia or thromboembolic complication. Fractures occurred in two patients (3.4%) and the tumour recurred in two patients (3.4%). PMID- 15274270 TI - Calcium hydroxyapatite ceramic implants in bone tumour surgery. A long-term follow-up study. AB - We reviewed the results of 51 patients with benign bone tumours treated by curettage and implantation of calcium hydroxyapatite ceramic (CHA). The mean follow-up was 11.4 years (10 to 15.5). Post-operative fractures occurred in two patients and three had local recurrences; three had slightly limited movement of the adjacent joint and one had mild osteoarthritis. There were no allergic or neoplastic complications. In all cases, radiographs showed that the CHA was well incorporated into the host bone. Statistical analysis showed that absorption of the implanted CHA was greater in males (odds ratio, 6.2; 95% CI, 1.6 to 23.7) and younger patients (odds ratio, 0.6 for increase in age of 10 years; 95% CI, 0.91 to 0.99). However, the implanted CHA was not completely absorbed in any patient. We conclude that CHA is a useful and safe bone substitute for the treatment of benign bone tumours. PMID- 15274271 TI - Valgus femoral osteotomy for hinge abduction in Perthes' disease. Decision-making and outcomes. AB - We studied, clinically and radiologically, the growth and remodelling of 21 hips after valgus femoral osteotomy with both rotational and sagittal correction for hinge abduction in 21 patients (mean age, 9.7 years) with Perthes' disease. The exact type of osteotomy performed was based on the pre-operative clinical and radiological assessment and the results of intra-operative dynamic arthrography. The mean IOWA hip score was 66 (34 to 76) before surgery and 92 (80 to 100) at a mean follow-up of 7.1 years (3.0 to 15.0). Radiological measurements revealed favourable remodelling of the femoral head and improved hip joint mechanics. Valgus osteotomy, with both rotational and sagittal correction, can improve symptoms, function and remodelling of the hip in patients with Perthes' disease. PMID- 15274272 TI - Slipped capital femoral epiphysis in skeletally immature patients. AB - Fixation by a single screw is considered the current treatment of choice for a slipped capital femoral epiphysis. This approach promotes premature physeal closure. The use of a modified, standard, single, cannulated screw designed to maintain epiphyseal fixation without causing premature closure of the physis was reviewed in ten patients. The nine boys and one girl aged between 10.6 and 12.6 years with unilateral slipped capital femoral epiphysis (SCFE), were markedly skeletally immature (Tanner stage I, bone age 10 to 12.6 years). Clinical and radiological review at a mean follow-up of 44.3 months (36 to 76) showed no difference in the time to physeal closure between the involved and uninvolved side. Measurement of epiphyseal and physeal development showed continued growth and remodelling in all patients. Use of this device provided epiphyseal stability and maintained the capacity for physeal recovery and growth following treatment for both unstable and stable slipped capital femoral epiphysis. PMID- 15274273 TI - Extra-articular subtalar arthrodesis. A long-term follow-up in patients with cerebral palsy. AB - Of 23 children (35 feet) with cerebral palsy who had undergone a Grice extra articular subtalar arthrodesis for a valgus hindfoot between 1976 and 1981, we reviewed 17 (26 feet), at a mean of 20 years (17 years 3 months to 22 years 4 months) after operation. Seven were quadriplegic, eight spastic diplegic, and two hemiplegic. They were all able to walk at the time of operation. Thirteen patients (20 feet) were pleased with the Grice procedure, 13 had no pain and 15 (23 feet) were still able to walk. The clinical results were satisfactory for most feet. Radiography showed that the results had been maintained over time but 14 feet developed a mean ankle valgus of 11 degrees (6 to 18) with a compensatory hindfoot varus in 12 feet. No deformity of the talus or arthritis of adjacent joints was noted. The Grice procedure gives good long-term results in children with cerebral palsy. PMID- 15274274 TI - One in 13 'original' articles in the Journal of Bone and Joint Surgery are duplicate or fragmented publications. AB - Duplicate publication in orthopaedic journals may further an author's academic career but this is at the cost of both scientific integrity and knowledge. Multiple publications of the same work increase the workload of editorial boards, misguide the reader and affect the process of meta-analysis. We found that of 343 'original' articles published in the Journal of Bone and Joint Surgery in 1999, 26 (7.6%) had some degree of redundancy. The prevalence of duplicate publications in the orthopaedic literature appears to be less than that in other surgical specialties but it is still a matter of concern. It is the author's responsibility to notify the editor of any duality when submitting a paper for publication. PMID- 15274275 TI - Avulsed posterior edge of the tibia. Earle's or Volkmann's triangle? AB - The term Volkmann's triangle for the avulsed posterior edge of the tibia in fracture-dislocations of the ankle is incorrect. Volkmann did not publish any articles relating to the posterior edge of the tibia. Credit should go to Henry Earle, who was an outstanding British surgeon of the first half of 19th century. He described avulsion of the posterior edge of the tibia in 1828. In 1823 he also published a monograph entitled Practical observations in surgery in which he described a specially designed bed for the conservative treatment of proximal fractures of the femur. PMID- 15274276 TI - Immobilisation after radiofrequency-induced shrinkage of tendon. A histological study in rabbits. AB - Despite widespread use of radiofrequency (RF) shrinkage, there have been no animal studies on the effects of post-operative immobilisation on the histological properties of the shrunken tissue. We have therefore examined the role of post-operative immobilisation after RF shrinkage with special emphasis on the histological properties of collagenous tissue. One patellar tendon of 66 New Zealand White rabbits was shrunk. Six rabbits were killed immediately after the operation. Twenty rabbits were not immobilised, 20 were immobilised for three weeks and 20 for six weeks. Fibroblasts, collagen and vascular quality and density were evaluated on sections, stained by haematoxylin and eosin. Nine weeks after operation the histological properties were inferior to those of the contralateral control tendons. Shrunk tendons did not return to normal at any time after operation irrespective of whether the animals had been immobilised or not. All the parameters improved significantly between zero and three weeks after operation. Immobilised tendons tended to have a better and faster recovery. Careful rehabilitation is imperative after RF shrinkage. Immobilisation aids recovery of the histological properties. Our findings in this animal model support a period of immobilisation of more than three weeks. PMID- 15274277 TI - The combination of pamidronate and calcitriol reverses particle- and TNF-alpha induced altered functions of bone-marrow-derived stromal cells with osteoblastic phenotype. AB - Periprosthetic bone loss after total joint arthroplasty is a major clinical problem resulting in aseptic loosening of the implant. Among many cell types, osteoblasts play a crucial role in the development of peri-implant osteolysis. In this study, we tested the effects of calcitriol (1alpha,25-dihydroxy-vitamin-D3) and the bisphosphonate pamidronate on titanium-particle- and TNF-alpha-induced release of interleukin-6 and suppression of osteoblast-specific gene expressions in bone-marrow-derived stromal cells with an osteoblastic phenotype. We monitored the expression of procollagen alpha1[1], osteocalcin, osteonectin and alkaline phosphatase mRNAs by Northern blots and real-time reverse transcription and polymerase chain reaction analyses. The release of various cytokines was also analysed by ELISA. We found that calcitriol or pamidronate could only partially recover the altered functions of osteoblasts when added alone. Only a combination of these compounds restored all the tested functions of osteoblasts. The local delivery of these drugs may have therapeutic potential to prevent or to treat periprosthetic osteolysis and aseptic loosening of implants. PMID- 15274278 TI - Allograft impaction and cement penetration after revision hip replacement. A histomorphometric analysis in the cadaver femur. AB - We studied various aspects of graft impaction and penetration of cement in an experimental model. Cancellous bone was removed proximally and local diaphyseal lytic defects were simulated in six human cadaver femora. After impaction grafting the specimens were sectioned and prepared for histomorphometric analysis. The porosity of the graft was lowest in Gruen zone 4 (52%) and highest in Gruen zone 1 (76%). At the levels of Gruen zones 6 and 2 the entire cross section was almost filled with cement. Cement sometimes reached the endosteal surface in other Gruen zones. The mean peak impaction forces exerted with the impactors were negatively correlated with the porosity of the graft. PMID- 15274279 TI - Hip fracture in the immobile patient. PMID- 15274280 TI - Arthroscopic treatment of osteochondral lesions of the talus. PMID- 15274281 TI - Outcome after reconstruction of the anterior cruciate ligament in athletic patients. PMID- 15274282 TI - Do sunscreens prevent skin cancer? PMID- 15274283 TI - When are liver tests warranted? PMID- 15274284 TI - Role of beta-blockers up for debate. PMID- 15274285 TI - Evaluation of fever in the international traveler. Unwanted 'souvenir' can have many causes. AB - As international travel becomes more common, primary care physicians will be increasingly involved in the treatment of patients who return home with febrile illnesses. The initial laboratory evaluation is critical. This approach to diagnosis involves taking a thorough history, asking specific questions about the patient's travel itinerary and activities, and giving a careful and complete physical examination. Malaria is the most common cause of febrile illness in travelers returning from endemic areas, and prompt evaluation is essential to initiating timely treatment. Various resources are available to assist in this evaluation. PMID- 15274286 TI - The clinical picture of metabolic syndrome. An update on this complex of conditions and risk factors. AB - Metabolic syndrome has piqued the interest and concern of physicians and patients alike. The syndrome represents a commingling of several conditions and risk factors common in the United States and links accelerated cardiovascular disease with insulin resistance. As more details are uncovered about metabolic syndrome, new questions arise. Here, Dr Doelle reviews what is known and not yet known about metabolic syndrome, its etiology, and its treatment. PMID- 15274287 TI - Understanding insulin resistance. What are the clinical implications? AB - Insulin resistance is an important clinical issue in patients with other prominent components of metabolic syndrome, such as central adiposity and diabetes. However, its presence may be less evident in patients who are neither obese nor diabetic. Is measurement of insulin resistance important in clinical practice? How might its presence change management in individual patients? In this concise review, Dr Sivitz discusses the underlying mechanisms involved in insulin resistance, the issues surrounding assessment, and the implications for management in patients in whom insulin resistance is either detected or suspected. PMID- 15274288 TI - New challenges in caring for diabetic patients. Primary care physicians can get caught in the middle. PMID- 15274289 TI - Diabetic retinopathy. Control of systemic factors preserves vision. AB - Diabetic retinopathy, a retinal microangiopathy, is the leading cause of blindness for persons aged 20 to 65 years in the United States. Routine screening and early treatment are cost-effective and have been shown to help preserve sight. Primary care physicians play a key role in treatment of systemic factors that lead to poor outcomes and referral to an ophthalmologist or a retinal specialist for screening and local treatment. This article, the first of two on retinal vascular disease, provides a review of nonproliferative and proliferative diabetic retinopathy. The second article, which will appear in an upcoming issue of POSTGRADUATE MEDICINE, discusses retinal vascular disease in hypertension. PMID- 15274290 TI - Uncovering tinea incognito. Topical corticosteroids can mask typical features of ringworm. PMID- 15274291 TI - Patient notes: canker sores. PMID- 15274292 TI - Cytotoxic activity of the recombinant anti-mesothelin immunotoxin, SS1(dsFv)PE38, towards tumor cell lines established from ascites of patients with peritoneal mesotheliomas. AB - BACKGROUND: Mesothelin, a cell surface glycoprotein, is an attractive candidate for targeted therapy given its overexpression, as detected by immunohistochemistry, in mesotheliomas. The goal of this study was to evaluate mesothelin expression in fresh tumor cells obtained from ascites of patients with peritoneal mesothelioma, as well as to determine the sensitivity of these cells to an immunotoxin targeting mesothelin. MATERIALS AND METHODS: Tumor cells were evaluated for mesothelin expression by flow cytometry using the murine anti mesothelin monoclonal antibody K1. The sensitivity of these tumor cells to SS1(dsFv)PE38, an immunotoxin consisting of the anti-mesothelin Fv linked to a mutated Pseudomonas exotoxin, was evaluated using a cell proliferation assay. RESULTS: Of the 7 tumor cell lines established from ascites of 12 patients with peritoneal mesothelioma, 6 expressed mesothelin while one cell line did not. Cell lines that expressed mesothelin were very sensitive to SS1(dsFv)PE38 with IC50s ranging between 0.08-3.9 ng/ml, while the cell line that was mesothelin-negative was resistant to SS1(dsFv)PE38. CONCLUSION: High expression of mesothelin is seen on tumor cells of patients with peritoneal mesothelioma and correlates with sensitivity to SS1(dsFv)PE38. Clinical studies of SS1(dsFv)PE38 in patients with peritoneal mesotheliomas are ongoing. PMID- 15274293 TI - CD105 inhibits transforming growth factor-beta-Smad3 signalling. AB - CD105 (endoglin) is an important component of the transforming growth factor-beta (TGF-beta) receptor complex and is highly expressed in endothelial cells in tissues undergoing angiogenesis such as healing wounds, infarcts and in a wide range of tumours. In an attempt to understand the molecular mechanism by which CD105 exerts its effects on angiogenesis by modulating TGF-beta1 signalling, in this preliminary communication, CD105 transfected rat myoblasts were utilized as an in vitro model. Overexpression of CD105 in these transfectants antagonised TGF beta1-mediated inhibition of cell proliferation and reduced TGF-beta1-mediated p3TP-Lux (PAI-1 promoter) luciferase activity. It also reduced (CAGA)12-Luc luciferase activity in response to TGF-beta1. The CAGA sequence is specific for Smad3/4 binding, implying that CD105 is involved in inhibition of TGF-beta1/Smad3 signalling. Furthermore, CD105 overexpression reduced serine phosphorylation of Smad3 and inhibited subsequent nuclear translocation of Smad3. CD105 resulted in high phosphorylation of JNK1, which is able to activate c-Jun. c-Jun is known to inhibit Smad3 transcriptional activity on CAGA sites, suggesting that CD105 may also inhibit Smad3 signalling through JNK1. PMID- 15274294 TI - Charge-dependent targeting: results in six tumor cell lines. AB - BACKGROUND: Many previous studies show that cell surface sialylation of malignant cells is enhanced compared to normal tissue. The carboxyl group of the sialic acid yields, a negative surface charge of the tumor cells. This study investigates how tumor cell growth is affected when a cationic polymer is incubated with six different tumor cell lines. MATERIALS AND METHODS: Cationic dextran (CatDex) was prepared by periodate oxidation and subsequent coupling of cationic sidegroups by reductive amination. A fluorimetric cytotoxicity assay (FMCA) was used for the cell survival assay. Six different tumor cell lines (lung, breast, ovarian, prostate, colon, urinary bladder) were seeded into 96 well microtiter plates. CatDex was added at different microM concentrations and incubated for 72 h. Additionally, CatDex was fluorescence-labeled (FITC) and the interaction with the tumor cells was studied using fluorescence microscopy. The presence of sialic acid in the different cell lines was confirmed by using a FITC labeled sialic acid binding lectin. RESULTS: CatDex showed a concentration dependent growth inhibitory effect (i.e. the number of cationic side groups/dextran molecule and the molarity used). If the substitution was <20%, the growth inhibitory effect was small and difficult to reproduce. With 20-22% substitution, the growth inhibition varied between 20-95% depending on the molarity and the tumor type. Higher substitution resulted in complete cell death in all the cell lines. The fluorescent images showed intensive cell membrane interaction. CONCLUSION: Incubation with cationic dextran caused cell death in all six tumor cell lines. Our hypothesis is that CatDex binds to the anionic sialic acid residues and causes fatal disturbances in the cell membrane. However the exact mechanism remains to be elucidated. The results may indicate a new method of general interest for intra/local/regiolocal treatment of cancer. Clinical studies to explore this concept are pending. PMID- 15274295 TI - Oral 5-FU is a more effective antimetastatic agent than UFT. AB - 5-Fluorouracil (5-FU), a pyrimidine analog, is widely used to treat gastrointestinal and other cancers. In the present study, we compared the efficacy of oral 5-FU and the 5-FU prodrug, uracil plus tegafur (UFT), on liver metastasis in a highly metastatic mouse model. Genetic labeling of the tumor with green fluorescent protein (GFP) along with inexpensive video detectors, positioned external to the mouse, allowed the real-time monitoring of details of tumor growth, metastatic spread and drug response in this mouse model. 5-FU at 10 and 20 mg/kg significantly prolonged the survival time of treated animals compared with untreated controls (p=0.003 for 5-FU, 10 mg/kg; p=0.016 for 5-FU, 20 mg/kg). In contrast, UFT only showed a trend to increase survival (p=0.078). 5 FU at 10 mg/kg substantially prolonged the survival time compared to UFT (p=0.012). 5-FU (10 mg/kg) was also more effective in prolonging survival than Furtulon (5'-deoxy-5-fluorouridine, another 5-FU prodrug) (p=0.042). All control and UFT-treated animals died by day 45. In contrast, at 45 days, 5 out of 8 animals were alive in the 5-FU 10 mg/kg group, which was found to be the best treatment regimen in this study. 5-FU (10 mg/kg)-treated animals had a median survival time of 53 days compared to 26.5 in controls and 33.5 days for UFT. These results suggest the potential clinical superiority of oral 5-FU compared to UFT as an anti-metastatic agent. The data also suggest the lack of clinical need for complex and expensive prodrugs of 5-FU such as UFT. PMID- 15274296 TI - Bone sialoprotein promotes bone metastasis of a non-bone-seeking clone of human breast cancer cells. AB - Bone sialoprotein (BSP) is a major non-collagenous protein in mineralized tissues. BSP is also implied to be involved in tumor metastasis through its unique structure. Using the human breast cancer cell line MDA-231, we established both brain-seeking and bone-seeking cell clones. The brain-seeking cells (MDA4 231BR) showed no bone metastasis in an animal model. In this experiment, MDA 231BR cells were transfected with BSP cDNA and inoculated into the hearts of nude mice. All five nude mice which received BSP-transfected MDA-231BR cells developed bone metastases, while no bone lesions were observed in the control group. Histological examination revealed invasion of tumor cells into the endosteal space and erosion of the bone margin. Some animals were crippled due to large lesions. These results suggest that BSP may impart to breast cancer cells the capacity to metastasize and thus play an important role in bone metastasis of malignant tumors. PMID- 15274297 TI - Biochemical analysis and subcellular distribution of E-cadherin-catenin in adenocarcinomas of the gastro-oesophageal junction. AB - BACKGROUND: Disturbances in the expression or structure of E-cadherin-catenin, a cell-cell adhesion complex, perturb its cell adhesive function. MATERIALS AND METHODS: We studied the expression and distribution of the E-cadherin-catenin complex in 24 adenocarcinomas of the gastro-oesophageal junction (GOJ) by immunohistochemistry and Western blotting of the Triton X-100-soluble (membrane bound) and insoluble fractions (cytoskeleton bound). RESULTS: Immunohistochemistry demonstrated redistribution of E-cadherin, alpha-, beta- and gamma-catenin from the membrane to the cytoplasm in 13/24 (54%), 18/24 (75%), 16/24 (67%) and 15/24 (63%) tumours, respectively. Five tumours showed nuclear localisation of beta-catenin. Western blotting showed redistribution between the TX-100 soluble and insoluble fraction of E-cadherin and the catenins in 5/11 (45%), 4/10 (40%), 5/11 (45%) and 5/11 (45%) tumours, respectively. CONCLUSION: Loss of membrane bound E-cadherin-catenin is frequently observed in adenocarcinomas of the GOJ and this may reflect loss of function of the E cadherin-catenin complex in these cancers. PMID- 15274298 TI - Caspase-1 enhances the apoptotic response of prostate cancer cells to ionizing radiation. AB - BACKGROUND: The significance of caspase-1 in prostate cancer has recently been documented (Cancer Res 61: 1227-1232, 2001). In this study, we investigated the role of caspase-1 in radiation-induced apoptosis in order to identify the significance of this apoptotic initiator in radiation resistance. MATERIALS AND METHODS: Caspase-1 was over-expressed in DU-145 prostate cancer cells (which have weak endogenous expression of caspase-1), via transfection-mediated gene transfer. Stable transfectants were cloned and expression of caspase-1 was established at the mRNA and protein levels by RT-PCR and Western blot, respectively. Caspase-1 overexpressing clones were characterized for their apoptotic response to ionizing irradiation (0-9 Gy) on the basis of cell viability and Hoechst staining assays and profiling of expression of key apoptosis regulators, such as caspase -3 and -9. RESULTS: Caspase-1 transfectants exhibited a greater sensitivity in response to ionizing radiation than the neomycin control transfectants, as demonstrated by a dramatic loss in cell viability, that temporally correlated with apoptosis induction. Furthermore, caspase-1 overexpression resulted in a significant decrease in clonogenic survival following treatment with ionizing radiation, while the caspase-1 inhibitor, Z-YVAD.fmk, suppressed apoptosis induction in caspase-1 transfectants (p<0.008). The apoptotic effect was associated with increased expression of the pro-enzyme form of caspase-3 in both the caspase-1 transfectants and neo controls cells, with the activated caspase-3 being detected in caspase-1 transfectants only. While this activation of caspase-3 was paralleled by an elevated caspase-9 expression at 9 h post-irradiation, there was no major induction in Apaf-1 or cytochrome c release. CONCLUSION: The present study provides an initial mechanistic insight into the functional involvement of caspase-1 in changing the apoptotic threshold of prostate cancer cells to radiation. These findings will enhance the understanding of the molecular basis of prostate tumor radioresistance and may have significant clinical relevance in improving the therapeutic index of radiotherapy in prostate cancer patients. PMID- 15274299 TI - PHGPx overexpression induces an increase in COX-2 activity in colon carcinoma cells. AB - BACKGROUND: Lipid peroxidation is a constant problem that eukaryotic cells have to face. Glutathione peroxidases (GPx) are among the most effective systems that protect cells from hydroperoxide toxicity. The objective of this study was to evaluate the relationship between GPx and cyclooxygenase 2 (COX-2), implicated in cancer pathogenesis, particularly in colon cancer cells. MATERIALS AND METHODS: Phospholipid hydroperoxide glutathione peroxidase (PHGPx or GPx4), which metabolizes peroxidized phospholipids, was cloned in an expression plasmid, transfected in HT29 cl.19A colon carcinoma cells and the effects of PHGPx overexpression were measured on arachidonic acid metabolism by COX-2. Metabolites were studied by HPLC and EIA; COX-2 mRNA levels were analysed rising semi quantitative PCR. RESULTS: Prostaglandins (PGE2, PGF2alpha, 6 keto-PGF1alpha) and thromboxane (TXB2) production were increased. COX-2 mRNA levels increased in PHGPx overexpressing cells. CONCLUSION: Surprisingly, our data suggest that PHGPx overexpression noticeably increases COX-2 metabolism. PMID- 15274300 TI - Extracellular expression of cytosine deaminase results in increased 5-FU production for enhanced enzyme/prodrug therapy. AB - BACKGROUND: The cytosine deaminase/5-fluorocytosine (CD/5-FC), strategy for cancer gene therapy shows considerable promise in experimental models but, because CD is a cytosolic enzyme, intracellular production of 5-fluorouracil (5 FU) causes the demise of the transduced cells before cytotoxic concentrations of' 5-FU can be achieved within the extracellular milieu. MATERIALS AND METHODS: A soluble secreted form of CD was constructed and evaluated compared to intracellular CD in vitro and in vivo. RESULTS: The secreted form of CD temporarily spared transduced cells and enhanced accumulation of extracellular 5 FU. Cytosolic CD produced rapid inhibition of thymidylate synthase and cell death before significant extracellular concentrations of 5-FU developed. Finally, tumors expressing the secreted form of CD had an improved response to 5-FC treatment compared to tumors expressing intracellular CD. CONCLUSION: Further evaluation of extracellular expression of CD for enzyme/prodrug therapy may provide improvements in this commonly studied gene therapy strategy. PMID- 15274301 TI - Selenium compounds regulate p53 by common and distinctive mechanisms. AB - Selenium compounds show much promise in the prevention of prostate and other human cancers. Various selenium chemical forms have been shown to differ widely in their anticancer properties. The main dietary form is selenomethionine, which we showed modulated p53 activity by causing redox regulation of key p53 cysteine residues. In the current study we included other selenium chemical forms, sodium selenite and methyl-seleninic acid. All three forms are relevant selenium sources in human populations. All three forms can affect p53 activity defined as trans activation of a p53-dependent reporter gene. In addition to the reduction of cysteine sulfhydryl groups, p53 phosphorylation was also affected in cells treated with selenium compounds. Methyl-seleninic acid caused phosphorylation of one or more p53 threonine residues, but did not affect any known serine phosphorylation sites. By contrast sodium selenite caused phosphorylation of p53 serines 20, 37 and 46 known to mediate apoptosis. Selenomethionine did not cause detectable phosphorylation of p53 serines or threonines. Our data show that, although p53 modulation may be a common denominator of selenium compounds, specific mechanisms of p53 activation differ among selenium chemical forms. Post translational modifications of p53 are determinants of p53 activity and probably affect the threshold for p53-mediated functions. Different selenium chemical forms may differentially modify p53 for DNA repair or apoptosis in conjunction with a given level of endogenous or exogenous DNA damage. PMID- 15274302 TI - The ligands of peroxisome proliferator-activated receptor (PPAR) gamma inhibit growth of human esophageal carcinoma cells through induction of apoptosis and cell cycle arrest. AB - In the present study, we examined the expression of peroxisome proliferator activated receptor gamma (PPARgamma) and the growth-inhibitory effects of Troglitazone and Pioglitazone, selective ligands for PPARgamma, using a series of human esophageal carcinoma cell lines (TE-1, -3, -7, -8, -12 and -13). PPARgamma expression was detected in all six human esophageal carcinoma cell lines. The esophageal carcinoma cell line TE-13 showed marked growth inhibition in response to Troglitazone and Pioglitazone. Flow cytometry performed on TE-13 cells exposed to Troglitazone showed that the cell cycle was arrested at the G1-phase. This result was confirmed by the finding of reduced cyclin D and cyclin E expression by Western blot analysis. DNA ladder formation was also detected, as was the induction of apoptosis-related proteins. Our results suggested that Troglitazone inhibited the growth of human esophageal carcinoma cell lines via G1 arrest and apoptosis and that PPARgamma ligands should be considered as possible target molecules in the treatment of human esophageal carcinomas. PMID- 15274303 TI - Analysis of Sciellin (SCEL) as a candidate gene in esophageal squamous cell carcinoma. AB - BACKGROUND: The aim of this study was to investigate whether a candidate gene, Sciellin (SCEL), mapping to the chromosome 13q21-q31 is mutated in esophageal cancer. MATERIALS AND METHODS: The coding region and intron-exon junctions of SCEL were sequenced in 13 esophageal squamous cell cancers and matching normal esophageal samples to detect mutations. RESULTS: Three single nucleotide polymorphisms were detected in SCEL of which two were silent mutations (L640L and H654H) and one missense mutation (R366K). CONCLUSION: Single nucleotide polymorphisms were detected in both matching tumor and normal esophageal tissues but no disease-associated mutations suggesting that SCEL is not a major factor in esophageal squamous cell carcinogenesis. PMID- 15274304 TI - Correlation of circulating tumor cells with tumor size and metastatic load in a spontaneous lung metastasis model. AB - BACKGROUND: The prognostic value of circulating tumor cells remains unclear since, in principle, most tumor cells are unable to survive in the bloodstream. The aim of the study was to establish a system that can be used to investigate the metastatic process in more detail, with emphasis on circulating tumor cells. MATERIALS AND METHODS: Human colon carcinoma cells (HT29) were transplanted into severe-combined-immunodeficient (scid) mice. The metastatic load in the blood was investigated using the human-specific carcinoembryonic antigen (CEA) as target for quantitative polymerase-chain-reaction (PCR). RESULTS: A close correlation between the weight of the primary tumor and the number of circulating tumor cells was detected (r=0.7240; p<0.0001). Moreover, the number of circulating tumor cells and the actual number of spontaneous lung metastases was related (r=0.8283; p<0.0001). CONCLUSION: A tumor xenotransplantation model is presented that allows for a detailed investigation of the metastatic process in three different compartments: the primary tumor bed, the bloodstream and the target organ of metastatic residency. PMID- 15274305 TI - A synthetic peptide derived from the human eosinophil-derived neurotoxin induces apoptosis in Kaposi's sarcoma cells. AB - Several commercial preparations of human chorionic gonadotropin (hCG) have been tested as therapy for Kaposi's sarcoma (KS) in clinical trials, but with discordant outcomes. We also have found dramatic differences in the cytotoxic effects of four different commercial hCG preparations on an established KS cell line, KSIMM. A co-purified moiety (ies) present in these preparations may explain these differences. The eosinophil-derived neurotoxin ribonuclease, extended with four extra residues ((-4)EDN), has been suggested to be the putative anti-KS compound in the hCG preparations, being specifically recognized by the cells through its N terminal extension. We therefore synthesized a 16-residue peptide (MSLHV-NT12 EDN), made to resemble the active recognition sequence of (-4)EDN. MSLHV-NT12 EDN displays a dose-dependent cytotoxic effect on KSIMM (killing 50% of the cells at 9 microg/ml). The cytotoxic effect is specific for KS cells, MSLHV-NT12 EDN being harmless even at 100 microg/ml for a melanoma cell line (SK MEL-28) or for normal human fibroblasts. We also demonstrated that MSLHV-NT12 EDN induces apoptosis in KSIMM cells. In conclusion, MSLHV-NT12 EDN is a specific proapoptotic substance for KS cells, which warrants further investigation into its in vivo effects. PMID- 15274306 TI - Dydrogesterone (Duphaston) and its 20-dihydro-derivative as selective estrogen enzyme modulators in human breast cancer cell lines. Effect on sulfatase and on 17beta-hydroxysteroid dehydrogenase (17beta-HSD) activity. AB - Estradiol (E2) is one of the main factors which control the growth and evolution of breast cancer. Consequently, to block the formation of E2 inside cancer cells has been an important target in recent years. Breast cancer cells possess all the enzymatic systems (e.g. sulfatase, aromatase, 17beta-hydroxysteroid dehydrogenase [17beta-HSD]) involved in the conversion of estrogen precursors into E2. Sulfotransferase, which converts estrogen to its sulfate, is also present in this tumoral tissue. Duphaston is a synthetic progestogen with properties similar to the natural progesterone. In the present study we examined the effect of Duphaston and its 20-dihydro-metabolite on the sulfatase and 17beta-HSD activities in MCF-7 and T-47D breast cancer cells. The cells were incubated with estrone sulfate (E1S) (5x10(-9)M) in the absence or presence of Duphaston or its 20-dihydro-metabolite (5x10(-5) to 5x10(-9)M) for 24h at 37 degrees C. In another series of experiments, estrone (E1) (5x10(-9)M) was incubated with T-47D cells in the absence or presence of the two progestogens (5x10(-5) to 5x10(-9)M) for 24h at 37 degrees C. E1S, E1 and E2 were characterized by thin layer chromatography and quantified using the corresponding standard. Duphaston and its 20-dihydro metabolite, at concentrations of 5x10(-7) and 5x10(-5)M, inhibited the conversion of E1S to E2 by 14% and 63%, 65% and 74%, respectively, in MCF-7 cells; the values were 15% and 48% and 31% and 51%, respectively, in T-47D cells. In another series of experiments it was observed that, after 24-h incubation, E1 (5x10(-9)M) was converted in a great proportion to E2 in the T-47D cells and that this transformation was significantly inhibited by Duphaston and its 20-dihydro metabolite. The IC50 value, corresponding to 50% of the inhibition in the conversion of 1 to E2, was 9x10(-6)M for 20-dihydro-metabolite in this cell line. It was concluded that the progestogen Duphaston and its 20-dihydro-metabolite are potent inhibitory agents on sulfatase and 17beta-HSD activities in breast cancer cells. Duphaston is a progestogen with properties similar to the endogenous progesterone. The data open interesting perspectives to study the biological responses of these progestogens in clinical trials of patients with breast cancer. PMID- 15274307 TI - Cyclooxygenase-2 expression in human gastric tubular adenomas and carcinomas; correlation with intratumoral microvessel density and apoptotic index. AB - BACKGROUND: Cyclooxygenase (COX)-2 plays an important role in carcinogenesis in various human malignancies. This study examined the relationship among COX-2 expression, angiogenesis and apoptosis in human gastric adenoma and carcinoma. MATERIALS AND METHODS: We examined the expression of COX-2 in 30 tubular adenomas and 11 carcinomas, comparing it with intratumoral microvessel density (IMVD) and apoptotic index (AI) by immunohistochemistry and the terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-digoxygenin nick-end labeling (TUNEL) procedure. RESULTS: Immunohistochemistry demonstrated positive expression of COX-2 in 15 (50.0%) adenomas and in 50 (53.1%) carcinomas, respectively. The frequency of COX 2 expression was significantly higher in intestinal-type carcinomas than in diffuse-type, regardless of the tumor stage. The IMVD was significantly higher in the early and advanced carcinomas than in the adenomas and also higher in the COX 2-positive adenomas and carcinomas than in the negative ones. The AI was significantly higher in the adenomas than in the carcinomas and also in the COX-2 negative adenomas and intestinal-type early carcinomas than in their positive counterparts, respectively (p < 0.05). The IMVD and AI showed significant inverse correlation in both the adenomas (p=0.02, r=-0.64) and carcinomas (p=0.04, r= 0.18). CONCLUSION: COX-2 expression might be an early event in gastric tumorigenesis and provide a preferential advantage for tumor cell proliferation because of its vascular-rich microenvironment and escape from tumor cell apoptosis, especially in intestinal-type gastric carcinomas. PMID- 15274308 TI - STI571 as a potent inhibitor of growth and invasiveness of human epithelial breast cancer cells. AB - STI571, a specific tyrosine kinase inhibitor, exhibits a substantial therapeutic activity in patients with chronic myeloid leukaemia and gastrointestinal stromal tumors. In this study we examined the activity of STI571 on the growth and invasiveness of three human epithelial breast cancer cell lines of low (MCF-7) and high (ZR-75-1 and MDA-MB-231) invasive potential. Growth of all cell lines in serum-containing medium was significantly inhibited by STI571 in a dose-dependent manner, with an average IC50 of approximately 5-6 microM. Flow cytometric analysis revealed that this effect is characterized by an accumulation of all breast cancer cell types tested in the G2/M-phase of the cell cycle with a concomitant decrease of the percentage of cells in the S-phase. Interestingly, no increase in apoptosis was observed, indicating that the effect of this kinase inhibitor is cytostatic rather than cytotoxic. In addition, STI571 exerts a significant inhibition effect on the invasion of the highly invasive breast cancer cell lines ZR-75-1 and MDA-MB-231. These results encourage further preclinical investigations on the mechanisms underlying the inhibitory effects of STI571, which may be of great value in breast cancer treatment. PMID- 15274309 TI - Vitamins A, E, microelements and membrane lipid peroxidation in patients with neoplastic disease treated with calcium antagonists and antagonists of receptors H2. AB - We studied the serum levels of vitamins A, E, zinc and copper in two hundred and twenty-five subjects of both sexes. They were divided into two groups: 87 healthy subjects who served as controls and 138 patients with neoplastic disease. The patients were subdivided according to the absence (n = 79) or the presence of metastatic disease (n =59). In 59 patients with cancer, who were in therapy with scavenger drugs of free radical such as calcium antagonists and the antagonists of receptors H2, we also studied the possible effect of the same therapy on the serum levels of vitamins, on the concentrations of the microelements and on membrane lipid peroxidation. We found that membrane lipid peroxidation, evaluated from the time of in vitro formation in the blood of so-called "Heinz bodies," decreased in all patients treated with scavenger drugs. In these patients the permeability of the erythrocyte membrane was similar to the controls and the serum levels of the vitamins were equal to the levels in patients who did not receive these therapies. Zinc concentration increased while copper remained unchanged. We also studied the levels of vitamins in some organs. The results are discussed considering the role of free radicals. We underline the importance of vitamins A and E in the protection from membranous peroxidation and from free radicals and the need to consider cancer as a systemic morbid event, apart from the contingent actual location. PMID- 15274310 TI - Induction of apoptosis in breast cancer cells by Saccharomyces cerevisiae, the baker's yeast, in vitro. AB - The present study was undertaken to evaluate the effect of phagocytosis of killed yeast on the induction of apoptosis in human metastatic breast cancer cells (MCF 7 and ZR-75-1) and non-metastatic breast cancer cells (HCC70). Heat-killed Saccharomyces cerevisiae, baker's and brewer's yeast, was cultured with cancer cells at a ratio of yeast to cancer cells = 10:1, and the percent apoptotic cancer cells was determined by flow cytometry and cytospin preparation. Upon phagocytosis of yeast, breast cancer cells underwent apoptosis. Induction of apoptosis was time- and dose-dependent. Apoptosis was detected as early as 0.5 h (13%), increased to 19% at 2 h and peaked (38%) at 4 h. Metastatic cancer cells were found to be more susceptible to yeast-induced apoptosis than non-metastatic cells; 629% increase for MCF-7 as compared to cells alone, 258% for ZR-75 cells, while HCC70 cells showed a 178% increase. Phagocytosis is associated with the disruption of mitochondrial membrane potential and activation of initiator and effector caspases 8, 9 and 3. However, inhibitors of these caspases did not inhibit yeast-induced apoptosis in cancer cells, suggesting that yeast induces apoptosis in breast cancer cells by a mechanism that is independent of caspase activation. This data may have clinical implications. PMID- 15274311 TI - Pyroglutamyl-histidyl-glycine, the endogenous colon mitosis inhibitor, regulates cyclic AMP level in non-tumorigenic colonic epithelial cells. AB - BACKGROUND: We have proposed that the mitosis inhibiting peptide, pyroGlu-His-Gly (pEHG), a colon-specific negative feedback regulator of cell proliferation, works through a G protein-coupled receptor (GPCR), as do many other pyroglutamyl peptides. MATERIALS AND METHODS: Non-tumorigenic YAMC (colon mucosa of Immorto mice), IMCE (Immorto-Min mouse hybrid) and human hepatoma (HepG2) cell lines were exposed to pEHG. cAMP concentrations were measured with a protein binding assay, mRNA levels with real-time PCR and Ca2+ concentration with an inverted fluorescence microscope on Fura-2/AM-loaded cells. RESULTS: pEHG (1 nM) increased the intracellular concentration of cAMP after 5-10 min in YAMC cells, but not in HepG2 cells. No effect was seen on cytosolic Ca2+, or in the expression of the proliferation and differentiation regulatory genes c-fos, egr-1 or fosB in YAMC or IMCE cells. CONCLUSION: pEHG stimulates the second messenger cAMP, but has no effect on intracellular Ca2+ or the gene expression of c-fos, egr-1 or fosB. PMID- 15274312 TI - Characterization of the CDP-like/CTAS-1 binding site in the okadaic acid response element (OARE) of the human CDK1(p34cdc2) promoter. AB - BACKGROUND: Transcription of CDK1 is induced at the G0/G1-phase of the cell cycle and after okadaic acid treatment and we identified the Site I okadaic acid response element (OARE -944 to -763nt) enhancer in the human CDK1 promoter. MATERIALS AND METHODS: The OARE region of the CDK1 promoter was characterized for enhancer/repressor activities. RESULTS: Transient transfection of upper and lower Site I subregions suggested enhanced transcription activity was divided between both while mobility shift assays demonstrated sequence-specific protein binding to Site IA. Site IA also formed shift complexes following serum starvation/refeeding and putative transcription factor binding sites clustered in Site IA. Oligonucleotides encoding a consensus CDP transcription factor binding site effectively competed against authentic Site IA in mobility shift assays. Mutation of the CDP-like binding sequence substantially reduced competition. DNaseI footprinting revealed binding at this site. CONCLUSION: The CDP-like recognition sequence appears to comprise the OARE binding authentic CDP and/or related factors. We termed this site the CDK1 transcription activation sequence-1 (CTAS-1) enhancer of the human CDK1 promoter. PMID- 15274313 TI - Cytotoxicty and radical modulating activity of isoflavones and isoflavanones from Sophora species. AB - We investigated 2 isoflavones and 9 isoflavanones from Sophora species for their cytotoxic activity against 3 normal human cells (gingival fibroblast, pulp cell, periodontal ligament fibroblast) and 2 human tumor cell lines (squamous cell carcinoma HSC-2, submandibular gland carcinoma HSG). Compounds with 2 isoprenyl groups (one in A-ring and the other in B-ring) such as tetrapterol G [YS31] and isosophoranone [YS24], and those with alpha,alpha-dimethylallyl group at C-5' of B-ring [YS26 (secundifloran), YS27 (secundiflorol A), YS28 (secundiflorol D), YS29 (secundiflorol E)] showed relatively higher cytotoxic activity. When hydrophobicity was assessed by octanol-water partition coefficient (log P), the maximum cytotoxic activity was observed at a log P value around 4. Compounds with intermediate cytotoxic activity [YS27, genistein, YS28, YS29, YS30 (secundiflorol F)] showed relatively higher tumor specificity. All isoflavones and isoflavanones did not stimulate the nitric oxide (NO) production by mouse macrophage-like Raw 264.7 cells, but almost completely inhibited the NO production by lipopolysaccharide (LPS)-activated Raw 264.7 cells. ESR spectroscopy showed that YS26 and YS28, which are the most inhibitory for NO production, efficiently scavenged superoxide anion and NO radicals. These data suggest that the inhibition of macrophage NO production by these isoflavanones may, at least in part, be explained by their radical scavenging or reduction activity. PMID- 15274314 TI - Suppression of proline-directed protein kinase F(A) inhibits the malignant growth of human pancreatic ductal adenocarcinoma. AB - BACKGROUND: Proline-directed protein kinase F(A) (PDPK F(A)) was originally identified as a specific phosphatase activating factor, but has subsequently been demonstrated as a multisubstrate PDPK possibly involved in the regulation of diverse malignant characteristics of various types of human cancers including prostate, leukemia, bladder and colon cancers. However, the role of this PDPK in a lethal carcinoma, such as pancreatic ductal adenocarcinoma, remains to be established. MATERIALS AND METHODS: The stable antisense clones with specific suppression of overexpressed PDPK F(A) of human pancreatic ductal adenocarcinoma cells (MIA PaCa-2) were first selected and subsequently characterized for the in vitro and in vivo growth studies. RESULTS: The molecular and cellular studies revealed that the antisense clones of MIA PaCa-2 cells with specific suppression of overexpressed PDPK F(A) potentially exhibited cell growth retardation, decreased serum independence, poor clonogenic growth and loss of anchorage independent growth. The in vivo study further confirmed that the SCID mice injected with the antisense clones with low-level PDPK F(A) did not develop any detectable tumors even after 7-week observation. In sharp contrast, the parental or control-transfected clones developed very large tumors (>5 cm3) under identical conditions. CONCLUSION: The molecular, cellular and animal results taken together demonstrate that overexpressed PDPK F(A) is essential for the malignant growth of human pancreatic ductal adenocarcinoma. PMID- 15274315 TI - Nuclear methylation levels in normal and cancerous thyroid cells. AB - BACKGROUND: Most cancers show abnormal DNA methylation and a positive correlation between hypomethylation and tumour progression. PATIENTS AND METHODS: In our laboratory the extent of DNA methylation in individual nuclei in normal, cancer and non-cancer thyroid tissue samples was quantified according to a previously described method of computer-assisted semi-quantitative analysis. Cancer and non cancer samples were obtained from nine patients with different thyroid pathologies (one multinodular goitre and eight carcinomas). Quantitative analysis was performed in two sets of samples, i.e. individual nuclei from touch preparations and from tissue sections. RESULTS: In all cancer specimens a statistically significant decrease of heterochromatin methylation was consistently observed. In both sets of samples a direct correlation was consistently observed between the extent of chromatin demethylation and the degree of malignancy. CONCLUSION: Our preliminary results suggest that our method of cell-by-cell detection of intranuclear methylation abnormalities may be a useful tool in early identification of thyroid cancer lesions. PMID- 15274316 TI - N-acetyltransferase activity is involved in paclitaxel-induced N-acetylation of 2 aminofluorene in human bladder cancer cells (T24). AB - Arylamine N-acetyltransferase (NAT) plays an important role in the metabolism of 2-aminofluorene (AF) and some types of arylamine drugs and carcinogens. Our previous studies have demonstrated that paclitaxel decreases NAT activity in human bladder, blood, colon and lung cancer cells. In this study, paclitaxel was selected to test the inhibition of NAT activity (N-acetylation of AF) and NAT gene expression in a human bladder cancer cell line (T24). The NAT activity was determined by high performance liquid chromatography for measuring the levels of N-acetylation of AF. The data showed that a 24-hour paclitaxel treatment decreased the amount of N-acetylation of AF in T24 cells. The NAT enzymes were stained and analyzed by Western blotting and flow cytometry. The tests indicated that paclitaxel decreased the levels of NAT in T24 cells. The expression of the NAT gene (mRNAT NAT) was determined by polymerase chain reaction (PCR) and cDNA microarray and it was found that paclitaxel induced the down-regulation of mRNA NAT expression in T24 cells. PMID- 15274317 TI - Differential expression of the prion-like protein doppel gene (PRND) in astrocytomas: a new molecular marker potentially involved in tumor progression. AB - The expression of the prion (PRNP) and prion like-doppel (PRND) genes and the presence of the proteins prion (PrP) and doppel (Dpl) were investigated in human gliomas. The PRNP and PRND expression profiles were evaluated by real-time reverse transcription-quantitative PCR in low- and high-grade astrocytomas, in glioblastoma-derived cell lines and in non-glial tumor specimens. The presence of PrP and Dpl proteins and their cellular localization were evaluated by Western blot and immunohistochemistry. High levels of PRNP expression were found in all tumoral samples studied. Unlike the non-tumoral controls, PRND was aberrantly expressed in glioblastoma multiforme and in two glioblastoma multiforme-derived cell lines, even in the absence of the PRND gene amplification. PRND expression was directly related to malignancy of the tumor: highest in glioblastoma multiforme, lower in anaplastic astrocytoma and even lower in the low-grade astrocytoma samples. High levels of PRND were also found in non-glial malignant tumor samples, such as gastric adenocarcinoma and anaplastic meningioma. Western blot analysis confirmed the PrP and Dpl expression, displaying variability in the electrophoretic patterns. Immunohistochemical analysis revealed a diffuse cytoplasmatic Dpl distribution in different astrocytic neoplastic cells, in infiltrating lymphocytes and in blood vessel endothelial cells. Of note, Dpl reactivity was different from that of the PrP, since PrP showed typical Golgi and membrane localised staining. Our findings suggest that the PRND gene might be a useful molecular marker in astrocytoma progression and in tumor grade definition. Understanding of the mechanisms of PRND increased expression might provide insight into the regulatory pathways of glioma development. PMID- 15274318 TI - Id3-mediated enhancement of cisplatin-induced apoptosis in a sarcoma cell line MG 63. AB - The molecular mechanism(s) underlying the resistance to cis diamminedichloroplatinum (CDDP)-induced growth inhibition include DNA repair, apoptosis and cell cycle progression. Inhibitor of differentiation (Id) proteins, which belong to the group of helix-loop-helix proteins, regulate cell cycle progression, differentiation and apoptosis. We examined whether CDDP exposure modulates the expression pattern of Ids and whether ectopic expression of Ids influences CDDP-induced cell death. Cell growth was assessed by WST-8 assay kit. Reactive oxygen species (ROS) was evaluated by flow cytometry using dihydroethidium. MG-63 sarcoma cells were stimulated with CDDP for various times and Id expression was assessed by reverse transcription-polymerase chain reaction. CDDP induced a considerable transient up-regulation of Id3 mRNA, but not Id2, 1-2 h after stimulation. Enforced expression of Id3 caused the MG-63 sarcoma cells to be more sensitive to CDDP-induced growth inhibition, through generation of ROS and caspase-3 activation. Together, our results suggest that CDDP-induced cell death appears to involve Id3. PMID- 15274319 TI - High frequency of allele-specific down-regulation of HLA class I expression in lung cancer cell lines. AB - Loss or down-regulation of HLA expression has been demonstrated in various types of solid tumors and is considered to be one of the mechanisms of tumor immunoescape. The effectiveness of immunotherapy using tumor-specific antigens (TSA) largely depends on the expression of the appropriate HLA class I alleles on the tumor cells. We analyzed the allele-specific HLA class I surface expression of six lung cancer cell lines using a broad panel of allele-specific monoclonal antibodies, as well as the effect of IFN-gamma on HLA expression. Flow cytometric analysis displayed a wide range, from minimal to a high degree of expression of monomorphic HLA class I among the studied cell lines. Allele-specific loss or down-regulation of HLA also was observed in 5 out of 6 cell lines. Our results suggest that lung cancer patients considered for specific immunotherapy should be examined with respect to the expression of specific HLA class I allele binding the TSA. PMID- 15274320 TI - Metals and metal compounds in cancer treatment. AB - Metals and metal compounds have been used in medicine for several thousands of years. In this review we summarized the anti-cancer activities of the ten most active metals: arsenic, antimony, bismuth, gold, vanadium, iron, rhodium, titanium, gallium and platinum. The first reviewed metal, arsenic, presents the anomaly of displaying anti-cancer and oncogenic properties simultaneously. Some antimony derivatives, such as Sb2O3, salt (tartrate) and organic compounds, show interesting results. Bismuth directly affects Helicobacter pylori and gastric lymphoma; the effects of bismuth complexes of 6-mercaptopurine are promising. Gold(I) and (III) compounds show anti-tumour activities, although toxicity remains high. Research into the potential use of gold derivatives is still ongoing. Several derivatives of vanadium show anti-proliferative activity, but their toxicity must be overcome. Several pieces of evidence indicate that iron deprivation could be an excellent therapeutic approach; furthermore, it is synergistic with classic anti-cancer drugs. Rhodium belongs to the same group as platinum and it also presents interesting activity, but with the same nephrotoxicity. Several rhodium compounds have entered phase I clinical trials. In contrast to the platinum complexes, titanium derivatives showed no evidence of nephrotoxicity or myelotoxicity; titanocene dichloride is undergoing clinical trial. The anti-proliferative effect of gallium could be related to its competition with the iron atom; in addition a derivative appears to reverse the multidrug resistance. The last metal reviewed, platinum, has given some of the very best anti-cancer drugs. Four derivatives are used today in the clinic; their mechanism of action and of resistance are described. PMID- 15274321 TI - A novel quantitative chick embryo assay as an angiogenesis model using digital image analysis. AB - A number of pathological events in humans such as solid tumour growth and metastasis have been found to be associated with angiogenesis, the formation of new blood vessels front pre-existing vasculature. A novel approach in cancer treatment consists of blocking or delaying the progression of neovascularisation to dysplastic cells in order to stop delivery of nutrients and oxygen. Thus, agents with antiangiogenic activity may be useful to treat cancer with potentially less systemic toxicity than conventional cytotoxic therapeutics. In this study we report the development and validation of a modified quantitative and objective in vivo chick embryo assay with Digital Image Analysis to screen compounds for antiangiogenic activity. The evaluation of vascular responses was optimised to less than one minute per sample in order to make this automatic method suitable for larger experimental series. Significant activity of the known antiangiogenic compounds thalidomide and captopril was determined, which resulted in approximately 50% inhibition of the vessel area compared to the control. The stability and release of thalidomide and captopril within 48 h after application to the extraembryonic vessel system was monitored by HPLC analysis. In conclusion, this multiple target test system will be complementary to existing angiogenesis assays and may be useful to quantify the antiangiogenic activity of compounds. PMID- 15274322 TI - Ultrastructure of sarcoma 180 cells after ultrasound irradiation in the presence of sparfloxacin. AB - BACKGROUND: Sonodynamic synergistic antitumor effects have been well described to date, but detailed ultrastructural studies in this field are still lacking. MATERIALS AND METHODS: A suspension of sarcoma 180 cells was exposed to ultrasound (US) in the presence of sparfloxacin (SPFX), one of the new quinolone antibiotics, at 2 W (0.64 W/cm2) for 30 seconds. The antitumor effect was evaluated by the survival rate of cells and cell morphology by scanning and transmission electron microscopy. RESULTS: After US irradiation, the survival rate of tumor cells in the SPFX-added group was 30.9%, significantly lower than 78.7% in the control group (p = 0.0002). Most cells in the control group were spherical, but in the SPFX-added group the cells were aspherical. Cell membranes were often ruptured and numerous pores of various sizes were observed. Some cells were totally disintegrated. CONCLUSION: The most characteristic changes of the synergistic antitumor activities of US and SPFX were on the cell membrane. PMID- 15274323 TI - A microplate assay for selective measurement of growth of epithelial tumor cells in direct coculture with stromal cells. AB - Stromal cells play an important role in regulating epithelial malignancies through diffusible factors and adhesion. Modulation of the tumor-stromal cell interaction is an attractive target for new antitumor strategies. To screen for a modulator of the interaction, we have now developed a quantitative colorimetric assay for measurement of tumor cell growth in coculture with stromal cells using rhodanile blue dye. Rhodanile blue specifically stained cytokeratin-positive tumor cells in the coculture. When human prostate carcinoma cells LNCaP, PC-3 and DU-145 were cocultured with normal prostate stromal cells (PrSC) in a microplate, growth of the prostate cancer cells in the coculture was selectively measured by the rhodanile blue staining method. Using this system, we searched for a modulator of the tumor-stromal cell interaction among clinically used drugs and natural products. As a result, we found that 5-fluorouracil, bleomycin and phthoxazolin A inhibit prostate cancer cell growth more strongly in coculture with PrSC than that in monoculture. Without need to pre-label cells and transfect a marker gene, our new method is simple, rapid and thus useful for screening for modulators of the tumor-stromal cell interaction. Furthermore, our results suggest that low molecular weight compounds modulate the tumor-stromal cell interaction. PMID- 15274324 TI - The breakdown of apoptotic mechanism in the development and progression of colorectal carcinoma. AB - BACKGROUND: Fas (APO-1/CD95) is a cell surface receptor that mediates apoptosis when it reacts with Fas ligand (FasL) or Fas antibody. Alterations of Fas and FasL expression have been demonstrated in various carcinomas. MATERIALS AND METHODS: We examined the alteration of Fas and FasL expression in seventy-eight specimens of colorectal adenoma and carcinoma by immunohistochemistry and real time reverse-transcriptase polymerase chain reaction (RT-PCR). RESULTS: Our study revealed that the expression of Fas was reduced in colorectal adenoma and completely lost in some 60% of colorectal carcinomas. Fas expression was significantly down-regulated in liver metastasis compared with corresponding primary colorectal carcinoma. The expression of Fas significantly related to p53 status, tumor location and apoptosis in colorectal carcinoma. Up-regulation of FasL was not detected in colorectal adenoma, carcinoma cells and liver metastatic cancer cells. CONCLUSION: These results indicate that Fas may play an important role, not only in development but also progression, and that FasL is not always required for both development and progression in colorectal carcinomas. PMID- 15274325 TI - mRNA expression of the angiogenesis markers VEGF and CD105 (endoglin) in human breast cancer. AB - BACKGROUND: Both VEGF and CD105 (endoglin) have been identified as markers of tumor angiogenesis and prognosis in breast cancer. They have always been studied in this kind of tumor by means of immunological methods. OBJECTIVE: To study, by means of reverse-transcription polymerase chain reaction (RT-PCR), the expression of VEGF and CD105 (endoglin) at the mRNA level in a series of breast cancers and to correlate the results obtained with all available clinical and biological features of the tumors. MATERIALS AND METHODS: Fresh tumor tissue from 103 previously untreated breast cancer patients was studied for VEGF and CD105 (endoglin) expression. In addition, the following parameters were determined in all tumors: DNA ploidy by means of flow cytometry; hormone receptor (ER & PR), Ki67, c-erb-B2 and p53 expression by means of immunohistochemistry; and h-MAM (mammaglobin) expression by means of RT-PCR. Classical prognostic parameters of the tumors, such as histological and nuclear grade or axillary lymph node invasion, were also included into the statistical analysis. RESULTS: VEGF mRNA expression levels above the 25th percentile were significantly (p<0.05) associated with high proliferation (Ki67>10%) and aneuploidy of the tumors and inversely with estrogen receptor expression (p<0.01). CD105 (endoglin) mRNA expression levels above the 25th percentile only correlated significantly with nuclear grade 3 (p<0.05). The expression of both genes did not correlate with each other. CONCLUSION: VEGF mRNA expression levels seem to be directly associated with VEGF functions at the protein level, whereas this seems not to be the case for CD105 (endoglin) mRNA levels. PMID- 15274326 TI - Shikonin inhibits the growth and N-acetylation of 2-aminofluorene in Helicobacter pylori from ulcer patients. AB - N-acetyltransferase (NAT) plays an important role in the N-acetylation of arylamine carcinogens and drugs. Shikonin has been shown to induce apoptosis and inhibit angiogenesis in vivo and in vitro. In this study, shikonin elicited dose dependent bacteriostatic activity in Helicobacter pylori cultures. Arylamine N acetyltransferase (NAT) activity (N-acetylation of 2-aminofluorene) was determined in the Helicobacter pylori isolated from peptic ulcer patients. Bacterial cytosols or intact cells with or without specific concentrations of shikonin co-treatment showed different percentages of 2-aminofluorene acetylation. Shikonin also inhibited N-acetylation of 2-aminofluorene in these examined Helicobacter pylori cytosols and intact cells in a dose-dependent manner. The apparent values of Km and Vmax were decreased after co-treatment with 4 microM shikonin in the cytosol of Helicobacter pylori. PMID- 15274327 TI - Procarbacine radiolysis and experiments in vitro. AB - Steady-state radiolysis of aqueous procarbazine (PC) was studied in air-free, aerated and solutions saturated with N2O. The corresponding Gi(-PC)-values obtained at pH=7.4 were: 2.85, 5.60 and 3.45, respectively. The investigations in vitro, using E. coli (AB 1157) as a model for living systems, demonstrated that PC acts as a cytostatic in air-free as well as in aerated media. However, it shows radiation protecting ability in the presence of N2O, where OH-radicals are the predominant reactants. Similar results were observed at pH=6.2. The experimental data contribute to a better understanding of the many-sided and frequently contradictory behavior of PC. PMID- 15274328 TI - Human herpesvirus type 8 genotypes in iatrogenic, classic and AIDS-associated Kaposi's sarcoma from Greece. AB - BACKGROUND: Kaposi's sarcoma-associated herpesvirus (KSHV)/human herpesvirus 8 (HHV-8) is consistently found in almost all observed Kaposi's sarcomas (KS), whether AIDS-associated, iatrogenic or classic. To our knowledge no data are available on the genetic polymorphism of HHV-8 from Greece. We report the study of 15 renal transplant recipients with KS, 5 with AIDS-associated KS, 11 with classic KS and 60 healthy individuals from Greece. MATERIALS AND METHODS: Polymerase chain reaction (PCR) was carried out on DNA extracted from peripheral blood mononuclear cells (PBMC) or KS cutaneous biopsies, using specific primers for the HHV-8, KS330 fragment from ORF-26 (233 bp) and the highly variable region (VR1) from ORF-K1 (363 bp). RESULTS: HHV-8 DNA was detected in 30 out of 31 (97%) KS cases, regardless of their clinico-pathological subtype and in 10 out of 60 (16.7%) healthy individuals. Sequencing of the ORF26 fragment demonstrated that the 40 HHV-8 strains were of the A and C sub-types. Furthermore, sequencing of the ORF-K1 showed that these HHV-8 strains of Greek origin were of the A1, A4, C1 or C3 sub-type. CONCLUSION: Our findings imply a possible link of the C3 subtype of HHV-8 in renal transplant-related KS cases (iatrogenic KS) in Greece, a link of the A4 subtype in AIDS-associated KS cases and a potential involvement of the A1 subtype in Greek classic KS incidences, as HHV-8 strains among healthy individual tested belong to the C1, C3 or A1 subtypes. PMID- 15274329 TI - Alteration of E-cadherin expression in gastric mucosa: role of intestinal metaplasia and Helicobacter pylori infection. AB - BACKGROUND: Although E-cadherins have been involved in gastric carcinogenesis, their role in precancerous lesions, such as intestinal metaplasia, is still unclear. This study aimed to assess the role of both intestinal metaplasia and H. pylori infection on E-cadherin expression in gastric mucosa. PATIENTS AND METHODS: Twenty-one consecutive patients with intestinal metaplasia were enrolled to assess E-cadherin expression in metaplastic areas. Twenty further patients without intestinal metaplasia, with and without H. pylori, were enrolled to evaluate the role of the infection on E-cadherin expression. All patients underwent upper endoscopy and gastric biopsies were taken for histological and immunohistochemical assessment. RESULTS: A substantial reduction of E-cadherin expression in metaplastic areas was observed in 14 (67%) of the 21 patients, similarly in H. pylori-infected and uninfected patients (64% vs 71%, p=0.3). In the group without intestinal metaplasia, no reduction in E-cadherin expression was detected either in infected patients or in those without H. pylori infection. CONCLUSION: The data showed that intestinal metaplasia is associated with E cadherin down-regulation, whereas H. pylori infection does not seem to play a direct role in this process. PMID- 15274330 TI - Granulocyte/macrophage colony-stimulating factor and interleukin-4-induced dendritic cells. AB - BACKGROUND: We investigated whether GM-CSF/IL-4 is the most efficient cytokine combination for differentiating dendritic cells (DC) in terms of its ability to elicit an antitumor immune response. MATERIALS AND METHODS: Two experimental models were examined: C57BL/6 mice bearing MC38 cells and Balb/c mice bearing cachexia-inducible Colon-26 cells. After immunization with DC pulsed with whole tumor cell lysate, tumors were inoculated into the subcutis. RESULTS: C57BL/6 mice immunized with lysate-pulsed DC effectively rejected the MC38 challenge and detectable MC38-specific cytotoxic lymphocytes (CTL) were observed. However, even those groups immunized with lysate-pulsed DC exhibited no protective immunity against Colon-26 challenge in Balb/c mice. Unexpectedly, mice inoculated with lysate-unpulsed DC showed an acceleration of cachectic progression (p=0.031) compared to control mice. CONCLUSION: We speculate that GM-CSF/IL-4-induced DC promotes Th2-dominated immunity in Balb/c mice. Consideration might be given to which combination of cytokines is appropriate for the ex vivo differentiation of DC in tumor immunotherapy. PMID- 15274331 TI - Effects of clodronate on cancer growth and Ca2+ signaling of human thyroid carcinoma cell lines. AB - Clodronate, one of the halogenated bisphosphonates, was found to inhibit the cell growth of endocytic macrophages, osteoclasts and several cancer cells through diverse mechanisms. Cytosolic Ca2+ signaling had previously been suggested as an apoptotic signal to certain cancer cells. Whether clodronate has an anti-cancer effect and induces the Ca2+ signal in thyroid cancer cells remains unknown. In this study, the effects of clodronate, including growth inhibition and cytosolic Ca2+ signaling, were examined and analyzed on ARO, SW579, WRO and TT thyroid cancer cell lines. Clodronate decreased the growth of these cells in a dose dependent manner and was more effective on slow growing cells. Clodronate treatment transiently increased cytosolic Ca2+ on slow growing SW579 thyroid cancer cells but not on the fast growing ARO cells. The results from this study implied that clodronate-mediated cell growth inhibition in slow growing thyroid cancer cells might correlate with a Ca2+ signaling pathway. PMID- 15274332 TI - Chemotaxis and chemokinesis of malignant mesothelioma cells to multiple growth factors. AB - BACKGROUND: Chemotaxis is defined as directional cell movement of cells towards concentration gradients of solubilized attractants, whereas chemokinesis is defined as random cell movement in the absence of chemoattractant gradients. Since tumor cell motility plays an important role in the process of tumor invasion and metastasis, we investigated these two distinct motile behaviors in highly invasive tumor, malignant mesothelioma. MATERIALS AND METHODS: Chemotaxis and chemokinesis of mesothelioma cells were assayed using Boyden chambers fitted with filters coated with collagen type IV and different growth factors and cytokines were used as chemoattractants. RESULTS: We found that growth factors such as epidermal growth factor, transforming growth factor-alpha, amphiregulin, heparin-binding epidermal growth factor-like growth factor, beta-cellulin, insulin-like growth factor-I, insulin-like growth factor-II and stem cell factor stimulated directional (chemotactic) and/or random (chemokinetic) motility in all mesothelioma cell lines tested, whereas none of acidic fibroblast growth factor, basic fibroblast growth factor, granulocyte-macrophage colony-stimulating factor or interleukin-6 induced migration in the same mesothelioma cells. CONCLUSION: These findings provide evidence that: (i) multiple growth factors can induce chemotaxis and chemokinesis in malignant mesothelioma cell lines, and (ii) may contribute to our understanding of the highly invasive behavior of malignant mesotheliomas in vivo. PMID- 15274333 TI - Synthesis and multidrug resistance reversal activity of 1,2-disubstituted tetrahydroisoquinoline derivatives. AB - BACKGROUND: Cancer treatment often fails due to multidrug resistance (MDR) of the tumor cells. One of the major causes is overexpression of P-glycoprotein (P-gp). MATERIALS AND METHODS: By N-substitution reactions of diamine, amino acid and amino alcohol derivatives with 1-substituted tetrahydroisoquinoline skeleton, structurally diverse 1,2-disubstituted 1,2,3,4-tetrahydroisoquinolines were synthesized. The compounds were assayed as P-gp inhibitors using a standard functional assay with rhodamine (6G) on MCF-7/Adr cells. Cytotoxicity was investigated on HeLa cells using an antiproliferative assay. RESULTS: Five of the 24 compounds showed greater P-gp inhibition than the control compound verapamil with AC50 values (concentration of the compound eliciting 50% of the maximal rhodamine 6G accumulation) significantly lower than that of verapamil. CONCLUSION: Novel compounds were synthesized that showed MDR-reversal effect. One of them, (1'R*,2R*)-2-[2'-[2''-hydroxy-3''-(alpha-naphthyloxy)propyl]-6',7' dimethoxy-1',2',3',4'-tetrahydro-1'-isoquinolyl]propan-1-ol hydrochloride, showed two times higher efficacy than verapamil at 10 times lower concentrations. The outcome makes this molecule an attractive subject for further investigation and development. PMID- 15274334 TI - Changed adhesion molecule profile of Ewing tumor cell lines and xenografts under the influence of ionizing radiation. AB - BACKGROUND: Adhesion molecules are involved in cell-cell and cell-matrix interactions and may be informative to characterize intercellular mechanisms of invasion and metastasis. This study was performed to characterize radiation induced changes in the adhesion molecule profile of Ewing tumor subpopulations on a single cell level. MATERIALS AND METHODS: In the present study, two Ewing tumors were characterized in vitro 4, 24 and 72 hours after radiation with 5 Gy and in vivo in a xenograft model 4, 6 and 15 days after radiation with 30 Gy, together with non-irradiated controls, by five parameter flow cytometry. Directly fluorescence-conjugated antibodies that were directed against adhesion molecules (LFA-1 (CD11a), HCAM (CD44), VLA-2 (CD49b), ICAM-1 (CD54), NCAM (CD56), LECAM-1 (CD62L) and CD86) were used. Annexin V and 7-AAD were used to characterize radiation-induced apoptosis. RESULTS: Tumor cell subpopulations were identified by the expression of adhesion molecules, apoptotic markers and DNA content. Heterogeneous changes of the adhesion molecule profile were identified on tumor cell subpopulations after radiation. The expression of CD11a and CD62L correlated with the expression of apoptosis-associated markers. CONCLUSION: The changes of flow cytometric profile under radiation may potentially correlate with a changed metastatic potential of tumor cell subpopulations. PMID- 15274335 TI - Promoter CpG methylation of caveolin-1 in sporadic colorectal cancer. AB - BACKGROUND: Caveolin-1 has been shown to be down-regulated in human colon cancer and involved in colon tumorigenesis. We investigated the mechanism. MATERIALS AND METHODS: Cancerous and nearby non-cancerous tissues of 185 sporadic colorectal cancer samples were enrolled in this study. Methylation-specific PCR was performed to explore the mechanism of regulation of caveolin-1 gene expression. RESULTS: Aberrant promoter methylation in the caveolin-1 gene was 3.8% and 5.9% for cancerous and nearby non-cancerous tissues, respectively. All the cancerous and non-cancerous tissue contained unmethylated promoters in the caveolin-1 gene. The methylation status of caveolin-1 had no clear relationship with age, cell grade, location of tumor or lymph node metastasis. However, female gender showed statistically significant difference (p=0.045). The immunohistochemistry study demonstrated that expression of caveolin-1 correlated with aberrant promoter methylation status in sporadic colorectal cancer tissues. CONCLUSION: Our findings suggested that aberrant promoter methylation of the caveolin-1 gene may occur at the precancerous stage, regulated by gender-related factors and is associated with gene silencing of caveolin-1 in the development of colorectal cancer. PMID- 15274336 TI - Expression of different vascular endothelial markers in prostate cancer and BPH tissue: an immunohistochemical and clinical evaluation. AB - BACKGROUND: The microvessel density (MVD) is often used as a quantitative parameter for angiogenesis. The aim of this study was to evaluate the three most commonly used endothelial markers of microvessels, CD31, CD34 and vWF, in benign and malignant prostatic tissue. MATERIALS AND METHODS: Fifteen benign hyperplastic prostate and 54 prostate cancer specimens were immunohistochemically stained for CD31/CD34/vWF. The MVD obtained by each marker was quantified by a vessel count in standardized grids within the area of maximum angiogenesis. The data on MVD was interrelated and compared to tumor staging, grading and clinical follow-up. RESULTS: A significant correlation of the CD31/CD34/vWF-MVD data was observed in BPH tissue, but not in PCa. In PCa, the most sensitive marker for newly derived blood vessels was CD34. While CD34-MVD demonstrated a significant association with tumorgrading and PSA follow-up, CD31- or vWF-MVD did not. CONCLUSION: CD34 is a suitable marker for the immunohistochemical visualization of microvessels in benign and malignant prostate tissue. PMID- 15274337 TI - Retinoic acid enhances histamine content and H1 receptor expression in human neuroblastoma cell line Paju. AB - BACKGROUND: A human neuroblastoma cell line (Paju) was induced by retinoic acid (RA) to differentiate into neuron-like cells. MATERIALS AND METHODS: We studied the expression and the possible role of histamine receptors H1 and H2 in retinoic acid mediated differentiation by semiquantitative RT-PCR. We studied the effect of exogeneously added RA on the morphological change of the human neuroblastoma cell line and the differentiation was followed by vimentine, glial fibrillary acidic protein (GFAP) and neurofilament (NF) immunostaining. We monitored the change of the histidine decarboxylase (HDC) expression and the histamine content during the RA treatment by immunoblot and flow cytometry methods. RESULTS: Our data showed that H1 and H2 histamine receptors are present on Paju cells. Ten nM RA markedly increased the H1 receptor expression of these cells, while the H2 expression was unchanged. CONCLUSION: In the RA-treated Paju cells, the histamine content increased compared to the untreated cells, suggesting that neuroblastoma derived histamine is involved in the regulation of RA-induced in vitro differentiation by H1 receptors. PMID- 15274338 TI - CD44s expression, in benign, borderline and malignant tumors of ovarian surface epithelium. Correlation with p53, steroid receptor status, proliferative indices (PCNA, MIB1) and survival. AB - AIM: To investigate the role of CD44s in the biological behavior of surface epithelial ovarian tumors and its correlation with clinicopathological parameters, prognosis, p53, steroid receptor status and proliferative indices (PCNA, MIB1). MATERIALS AND METHODS: We analyzed a total of 83 patients with ovarian surface epithelial tumors, for the immunohistochemical expression of CD44s and the possible correlation with clinicopathological factors and patients' outcome. RESULTS: A statistically significant correlation was found between the expression of CD44s, which was higher in cancer cases than in benign cystadenomas (p<0.0001) and, between cancer cases, which was lower in borderline tumors, (p=0.05). No statistical correlation was found between CD44s expression and the examined markers. In overall survival analysis we did not detect a statistically significant correlation with the expression of CD44s. CONCLUSION: The current study demonstrates that CD44s may be functionally involved in the pathogenesis of epithelial ovarian lesions. PMID- 15274339 TI - Expression of L-type amino acid transporter 1 (LAT1) and 4F2 heavy chain (4F2hc) in oral squamous cell carcinoma and its precusor lesions. AB - BACKGROUND: Amino acid transporters play an important role in supplying organic nutrients to cells. The expression of L-type amino acid transporter 1 (LAT1) and its subunit 4F2 heavy chain (4F2hc) was evaluated to determine the alterations of these transporters in oral normal mucosa (ONM), oral precancerous lesion (OPL) and oral squamous cell carcinoma (OSCC). MATERIALS AND METHODS: Sections from formalin-fixed, paraffin-embedded samples of ONM, OPL or OSCC were examined using immunohistochemical staining to detect L4T1 and 4F2hc proteins. RESULTS: The LAT1 and 4F2hc expression increased progressively from ONM to hyperplastic and to dysplastic lesions and OSCC. In particular, LAT1 may be a more specific indicator of tumor progression than 4F2hc. CONCLUSION: LAT1 and 4F2hc may have an important role in the early stages of multistep oral carcinogenesis. In addition, the specific inhibition of LAT1 and 4F2hc might be a new rationale to suppress oral cancer progression. PMID- 15274340 TI - N-ethylphenyl acetamide (EPA) inhibits DNA synthesis and N-acetylation of 2 aminofluorene in human colon tumor cells (colo 205). AB - Many arylamines (including carcinogens) and hydrazine drugs are acetylated by the cytosolic N-acetyltransferase (NAT). NAT plays an important role in the first step of arylamine metabolism and has been found in immortalized human cell lines, human and laboratory animal tissues. Human colon tumor cell line (colo 205) has also been shown to acetylate arylamine and possess NAT activity. The purpose of this study was to determine whether or not N-ethylphenyl acetamide (EPA) could affect cell viability, DNA synthesis and N-acetylation of 2-aminofluorene (AF) in colo 205 cells. Acetylated and nonacetylated AF were determined by using high performance liquid chromatography. EPA displayed a dose-dependent inhibition of cytosolic and intact cells' NAT activity, inhibition of viablility and DNA synthesis. Time-course experiments showed that N-acetylation of AF measured from intact colo 205 cells was inhibited by EPA for up to 48 h. PMID- 15274341 TI - Induction of apoptosis by tocotrienol in rat hepatoma dRLh-84 cells. AB - Our aim was to evaluate the antitumor activities of tocopherol (Toc) and tocotrienol (T3) derivatives. At first, we examined the effect of these vitamin E homologues on the proliferation of rat normal hepatocyte RLN-10 and hepatoma dRLh 84 cells and found that especially T3 inhibited cell proliferation in dRLh-84 cells. Then, we examined the effect of vitamin E homologues on apoptosis induction and found that T3 induced DNA fragmentation and stimulated a rise of caspase-3 activity. In addition, T3 stimulated a rise in caspase-8 activity, while a caspase-8 inhibitor suppressed apoptosis induction by T3. We also examined the incorporation of vitamin E homologues into dRLh-84 cells. T3 was incorporated more quickly compared to Toc. These results indicated that T3 induces apoptosis in dRLh-84 cells and that caspase-8 is involved in this apoptosis induction. The difference in terms of apoptosis induction by vitamin E homologues seems to be related to their different rates of cellular incorporation. PMID- 15274342 TI - A simple model to simulate cellular changes in the T cell system following HIV-1 infection. AB - A new mathematical model is presented to simulate various changes of cell pools in the T cell immune system with validation procedures imitating viral infections. The present paper focuses on changes during the course of an acute progressive HIV-1 infection. Parameters are optimized by a direct search method and the stability of the model is studied. Mathematical modeling supports the hypothesis that the differentiation blockade is one major reason for the depletion of CD4 cells and the proliferation of CD38 cells in HIV-1 infection. The model appears to be a useful basis for further simulating disturbances of the T cell immune system in other viral infections as well as to elucidate the pathogenesis of various immunological diseases including the development of malignant lymphomas. PMID- 15274343 TI - Stealth liposomal 5-fluorouracil with or without degradable starch microspheres for hepatic arterial infusion in the treatment of liver metastases. An animal study in VX-2 liver tumor-bearing rabbits. AB - PURPOSE: Regional application of cytostatics in liver metastases leads to increased concentrations in tumor tissue. Flow retardation by temporary occlusion and drug targeting via liposome encapsulation (PEG liposomes) will further increase tumor concentrations. MATERIALS AND METHODS: Liver tumor-bearing rabbits were submitted to i.v. or i.a. therapy with or without liposome-encapsulated 5 fluorouracil (5-FU). I.a. groups were additionally treated with or without degradable starch microspheres. Tumor concentrations were calculated by HPLC as area under the curve (AUC). RESULTS: A comparison with i.v.-applied 5-FU yielded the following increasing concentrations: 5-FU-PEG liposomes i.v. 6-fold, 5-FU i.a. 20-fold, 5-FU i.a. + DSM 226-fold, 5-FU-PEG liposomes i.a. 319-fold, 5-FU PEG liposomes i.a. + DSM 2203-fold. CONCLUSION: The intratumoral concentration of 5-FU was increased up to 2203 times the intravenous dose by combination of regional application via the hepatic artery with temporary embolization by degradable starch microspheres and drug targeting by liposome encapsulated 5-FU. PMID- 15274344 TI - Isolation and characterization of a new cell line from a renal carcinoma bone metastasis. AB - BACKGROUND: The use of cell lines isolated from metastases should enable the assessment of peculiar aspects of bone resorbing cytokine expression, as well as angiogenetic activity of renal carcinoma bone metastases. MATERIALS AND METHODS: A cell line (CRBM-1990) was isolated from a renal cell carcinoma bone metastasis and compared with the ACHN and Caki-1 established lines. The expression of osteolytic cytokine and angiogenetic growth factors mRNAs, as well as the effect on migration and proliferation of a bovine bone cell line (BBE) were determined. RESULTS: There were no significant differences between the three lines in IL-6, TGF-beta, VEGF-A, VEGF-B, VEGF-C and FGF-2 mRNAs expression. VEGF-D, PIGF, or RANK-L-specific mRNA were not expressed. CRBM-1990-, Caki-1- and ACHN-conditioned media significantly stimulated the migration and proliferation of BBE. CONCLUSION: CRBM-1990 expressed IL-6-specific mRNA, but not RANK-L, expressed angiogenetic growth factors and induced migration and proliferation of bone endothelial cells at a non-significantly different level when compared with Caki 1 and ACHN. PMID- 15274345 TI - Selected cyclic dipeptides inhibit cancer cell growth and induce apoptosis in HT 29 colon cancer cells. AB - BACKGROUND: An increasing number of cyclic dipeptides (CDPs), particularly those containing proline, have been shown to exhibit important biological activity. MATERIALS AND METHODS: We investigated the potential of seven proline-based CDPs to inhibit cancer cell growth in HT-29, HeLa and MCF-7 cell lines. We also tested whether any of the CDPs were able to induce apoptosis in HT-29 cells. RESULTS: The SRB assay showed that only cyclo(Phe-Pro) (10 mM) exhibited more than 50% growth inhibition (p<0.01). The MTT assay was used to demonstrate a dose dependent (0.008-10 mM) growth inhibition by cyclo(Phe-Pro). Hoechst 33342 staining showed that 5 mM cyclo(Phe-Pro) induced chromatin condensation in 18.3+/ 2.8% (p<0.01) of HT-29 cells after 72 hours. Furthermore, annexin V binding revealed phosphatidylserine externalisation in cyclo(Phe-Pro)-treated HT-29 cells. CONCLUSION: Our findings demonstrate that cyclo(Phe-Pro) inhibits the growth of HT-29, MCF-7 and HeLa cells and induces apoptosis in HT-29 colon cancer cells, suggesting a potential antitumour activity. PMID- 15274346 TI - Comparative genomic hybridization in cartilaginous tumors. AB - Genetic aberrations in cartilaginous tumors have not yet been well characterized. We analyzed the molecular-chromosomal aberrations in 10 chondrosarcomas (four Grade-3 tumors, four Grade-2 tumors and two Grade-1 tumors) and in three benign cartilaginous tumors (two enchondromas and one chondromyxoid fibroma). Genomic imbalances were detected in 9 out of 10 cases of chondrosarcomas. The median number of changes was 7.0 per tumor (range 0-23) and the gain-to-loss ratio was 1:1.4. The most frequent gains involved 7q, 5p, or 21q and the most frequent losses were 17p, 13q, 16p, or 22q. The three benign cartilaginous tumors each had two (0 gains and two losses), six (one gain and five losses) and eight (one gain and seven losses) chromosomal aberrations. Both of the gains occurred on 13q21 and losses were frequently observed on chromosomes 19 and 22q in all three cases. Loss of chromosomes 16p, 17p, 22q, or 19 loss were common in both chondrosarcomas and benign cartilaginous tumors. However, aberrations from chromosomes 2 to 11, 14, 15, 18, or 21 were detected only in chondrosarcomas. Therefore, although the number of aberrations between benign and malignant cartilaginous tumors appears to be similar, these two entities may be differentiated by determining which chromosomes are affected. PMID- 15274347 TI - Luteinizing hormone-releasing hormone (LHRH) inhibits apoptosis induced by cytotoxic agent and UV-light but not apoptosis mediated through CD95 in human ovarian and endometrial cancer cells. AB - Luteinizing hormone-releasing hormone (LHRH) and its receptor are frequently expressed in human ovarian and endometrial cancers and are part of an autocrine mechanism of growth control. We have previously shown that the LHRH analog Triptorelin induces activation of nucleus factor kappa B (NFkappaB) and reduces apoptosis induced by doxorubicin in human ovarian cancer cells EFO-21 and EFO-27. The present study was performed to investigate the anti-apoptotic effects of LHRH analogs on apoptosis induced by doxorubicin, UV-light and ligation of CD95 in human endometrial and ovarian cancer cells. We further investigated the interaction of the LHRH system with the apoptotic pathway focusing on the effector-protease caspase 3. Doxorubicin (100 nM) induced apoptosis in the LHRH receptor-positive human endometrial cancer cell line Ishikawa and in the human ovarian cancer cell lines EFO-21 and NIH:OVCAR-3. Pretreatment for 24 h with native LHRH, the LHRH agonist Triptorelin or the LHRH antagonist Cetrorelix (100 nM) significantly reduced apoptosis induced by doxorubicin in these cells. In EFO 21 cells pretreatment with 100 nM Triptorelin also reduced UV-light-induced apoptosis from 76% to 62.7% (p<0.01). EFO-21 cells express CD95. Cross-linking of CD95 with monoclonal antibody anti-APO-1 (500 ng/ml) increased apoptosis from spontaneous rate to 10.3% to 38.3% in EFO-21 cells (p<0.001). Pre-treatment with Triptorelin did not reduce CD95-mediated apoptosis in these cells. LHRH analogs protect human endometrial and ovarian cancer cells from DNA-replication-dependent cytotoxic agent and UV-light-induced apoptosis, but not from CD95-mediated apoptosis. PMID- 15274348 TI - New evaluation of plasma DNA microsatellite analysis in patients with TCC of the urinary bladder. AB - BACKGROUND: To determine the diagnostic value of plasma DNA microsatellite analysis in patients with transitional cell carcinoma (TCC) of the urinary bladder, by redefining plasma LOH from the equivalent analysis in controls. The method was further tested for MSI (microsatellite instability) and compared with tissue DNA analysis. MATERIALS AND METHODS: Sixteen microsatellites were amplified in leukocyte, plasma and tissue DNA from 40 patients and 28 controls, and analysed in a laser-based, capillary electrophoresis system. Plasma LOH was determined from the controls' cut-off values. RESULTS: The difference between plasma LOH frequency in patients (25% (10/40)) and controls (14% (4/28)) was not significant. Nevertheless, it occurred significantly more often in low rather than high-grade tumors (p=0.03) and controls (p=0.04). Plasma MSI was dependent upon the number of PCR cycles. Tissue LOH was present in 78% (31/40) of the patients and in none of the controls. Tissue MSI was uncommon. CONCLUSION: The results of plasma DNA microsatellite analysis in TCC need cautious interpretation. PMID- 15274349 TI - Role of protein kinase C-dependent signaling pathways in the antiangiogenic properties of nafoxidine. AB - We analyzed the effect of nafoxidine on the earlier biological processes of angiogenesis and explored the role of different signaling pathways involved in the in vitro response of endothelial cells (HUVEC). Nafoxidine significantly inhibited adhesion, spreading, migration and invasion of HUVEC at concentrations ranging from 1 to 2.5 microM. Endothelial cord formation on Matrigel was inhibited by nafoxidine and cotreatment with phorbol-12-myristate-13-acetate (PMA) clearly prevented the antiangiogenic effect of the antiestrogen. On the contrary, cotreatment with the PKC inhibitor bisindolylmaleimide potentiated inhibition of cord formation. PMA also inhibited the nafoxidine-induced secretion of metalloproteinase-2 and tissue inhibitor of metalloproteinases-1 in HUVEC monolayers. Cotreatment with the phosphodiesterase inhibitor 3-isobutyl-1 methylxanthine and the cAMP analog N6,2'-o-dibutyryladenosine 3',5'-cyclic monophosphate prevented the inhibition of endothelial cord formation induced by nafoxidine. Our work presents evidence about the signaling pathways involved in the antiangiogenic effect of nafoxidine, suggesting that PKC-dependent signaling pathways are essential in angiogenesis during endothelial cord formation. PMID- 15274350 TI - Topotecan-induced alterations in the amount and stability of human DNA topoisomerase I in solid tumor cell lines. AB - BACKGROUND: Human DNA topoisomerase I (topo 1) is an essential nuclear enzyme involved in vital cellular processes and the sole target of antitumor drugs of the camptothecin (CPT) family. The CPT derivative topotecan (Tpt, Hycamtin) is currently used in clinic, its effectiveness varying considerably for different types of cancer. The purpose of this study was to compare time- and dose dependent cellular responses to Tpt in terms of alterations in the amount and stability of topo 1 in lung adenocarcinoma (A-549), ovarian adenocarcinoma (CaOv 3), colorectal adenocarcinoma (HT-29) and breast adenocarcinoma (MCF-7) cell lines. MATERIALS AND METHODS: Western blot analysis of the time-dependent redistribution of a full-size topo 1 and its proteolytical fragments was performed after Tpt treatment for 1 h at concentrations 10-fold or 100-fold higher than the Tpt IC50 for the respective cell lines. RESULTS: Tpt treatment of the CaOv-3 cell line produced a substantial time-dependent decrease in the amount of topo 1 immunoprotein. Conversely, the MCF7 cell line did not exhibit a topo 1 associated response to the Tpt treatment. Strong but different time- and dose dependent topo 1 down-regulation effects were observed in the HT-29 and A-549 cell lines. CONCLUSION: The data obtained indicate that Tpt-induced time- and dose-dependent effects on the amount and stability of topo 1 are involved in the mechanisms of Tpt activity against different solid tumor cell lines. PMID- 15274351 TI - Mitochondrial DNA mutations in light-associated skin tumors. AB - Mutations of mitochondrial DNA (mtDNA) have been proposed to be involved in carcinogenesis. In this study, we applied the polymerase chain reaction techniques to investigate the frequency of occurrence and proportion of mtDNA with length mutations (deletions and tandem duplications) in light-associated skin tumors (actinic keratosis, AK; basal cell carcinoma, BCC; and squamous cell carcinoma, SCC) in aged individuals. We demonstrated the existence of multiple mtDNA deletions and tandem duplications in tissues of AK, BCC, SCC and normal skin. We showed that the frequencies of occurrence of the 4,977 bp and 7,436 bp deletions and tandem duplications (200 bp and 260 bp) of mtDNA in light associated skin tumors were not significantly different from those of sun-exposed normal skin (p>0.05), but higher than those of non-exposed normal skin (p<0.05). In addition, we found that the proportion of the 4,977 bp-deleted mtDNA in the skin of the same individual was also affected by skin pathologies. The proportion of 4,977 bp-deleted mtDNA in relatively rapid growing tumor cells in SCC was lower than that of normal skin cells. We suggest that the existence of these length mutations of mtDNA in normal, precancerous or cancerous human skin may be attributed to the stochastic effect of photo-damage. However, it is unclear whether the mutations of mtDNA have a direct bearing on carcinogenesis in skin. Future investigation is warranted to elucidate the causal relationship between mtDNA mutations and skin cancers and to address the pathophysiological role of mtDNA mutations in skin cancer development. PMID- 15274352 TI - Antiangiogenic potency of various chemotherapeutic drugs for metronomic chemotherapy. AB - From previous preclinical findings continuous low dose (metronomic) chemotherapy is thought to inhibit tumor angiogenesis. This suggests that activated endothelial cells may be more sensitive to chemotherapeutic drugs than tumor cells. Therefore, we assessed the IC50 for several relevant chemotherapeutic drugs in different endothelial and tumor cell lines to identify optimal compounds to be used for metronomic therapy in a murine renal cell carcinoma model. Adriamycin, idarubicin, 5-fluorouracil, paclitaxel and etoposide were chosen for our studies because of their oral availability in patients or previous reports on metronomic potential. IC50s were determined by BrdU cell growth assay after short time as well as long term exposure of the following cell lines: human endothelial cells (HdmVEC/HUVEC), human breast cancer (Mcf-7), melanoma (Skmel), liver cancer (Huh7/Alexander), lung cancer (A549/LXFL), colon cancer (Dld) and murine renal cell carcinoma (RENCA). In addition, FACS analysis was performed to determine the effect on cell cycle. In vivo, doses of 2x12 mg/kg, 2x1.2 mg/kg and 10x0.24 mg/kg adriamycin were applied to 12 RENCA mice each and antitumor as well as antiangiogenic effects were assessed 21 days after tumor cell application. Independent of the exposure time, all chemotherapeutic drugs were more active against the endothelial cell lines. IC50s were significantly lower in endothelial cells (4.02E-06 to 6.16E-14 M) as compared to tumor cells (7.44E-02 to 1.9E-11 M). Cell cycle analysis of all chemotherapeutic drugs revealed a G1-arrest in endothelial cells. Adriamycin applied in metronomic doses of 10x0.24 mg/kg showed significant antiangiogenic activity whereas, in contrast, the application of 2x12 mg/kg significantly increased the vessel density in primary tumors. In summary, all chemotherapeutic agents were more active against endothelial cells in comparison to tumor cells. The hypothesis of an antiangiogenic active metronomic therapy could be confirmed in vivo by the use of adriamycin in RENCA. PMID- 15274353 TI - Histological changes pertinent to local tumor progression in hereditary nonpolyposis colorectal cancer (HNPCC). A preliminary report. AB - While the genetic profiles of hereditary colorectal tumors are being unravelled, the mechanisms implicated in their local progression remain to be deciphered. In this work histological features occurring at the invading tumor front were investigated in ten hereditary non-polyposis colorectal cancers (HNPCC). Of eight moderately-differentiated (i.e. gland-forming) adenocarcinomas, six had dilated glands with a thin layer of tumor cells and all eight had dilated glands in which a group of cells was lacking, i.e. with a glandular pore. It was apparent that the thin glandular epithelium was a stage preceding pore formation. In glands with pores, the contents of the neoplastic glands--rich in proteolytic enzymes- were extruded directly into the extracellular matrix (ECM), leading to the local destruction of the juxtaposed matrix. It was assumed that to restore the continuity of the glands new cancer cells would grow from the tip of the free borders of the pore into the damaged ECM, thus guaranteeing a stepwise, but everlasting, tumor progression in untreated patients. PMID- 15274354 TI - Selective induction of G2/M arrest and apoptosis in HL-60 by a potent anticancer agent, HMJ-38. AB - We previously reported that HMJ-38 was the most potent 2-phenyl-4-quinozolinone derivative in inhibiting tubulin polymerization and showed significant cytotoxicity against several human tumor cell lines. In this work, we studied its cytotoxic effect on HL-60 leukemia cells and the underlying mechanisms. We first investigated the effects of HMJ-38 on viability, cell cycle and induction of apoptosis in HL-60 and normal human peripheral blood mononuclear cells (PBMC). After 24-hour treatment with HMJ-38, a dose- and time-dependent decrease in the viability of HL-60 cells was observed and the approximate IC50 was 4.48 microM. The cytotoxic effect of HMJ-38 on PBMC was less significant than that on HL-60 cells, either with 24 or 48 hours of treatment. Cell cycle analysis showed that HMJ-38 induced significant G2/M arrest and apoptosis in HL-60 cells. The HMJ-38 induced G2/M arrest occurred before the onset of apoptosis. Within 24 hours of treatment, HMJ-38 influenced the CDK/cyclin B activity by increasing Chk1, Wee1 and p21 and decreasing Cdc25C protein levels. The HMJ-38-induced apoptosis was further confirmed by morphological assessment and DNA fragmentation assay. Induction of apoptosis in HMJ-38-treated HL-60 cells was accompanied by an apparent increase of cytosolic cytochrome c, down-regulation of Bcl-2, up regulation of Bax and cleavage of pro-caspase-9, -3 and poly(ADP)ribosylpolymerase (PARP). The results of the significant reduction of caspase activities and apoptosis by caspase inhibitors indicated that the HMJ-38 induced apoptosis was mainly mediated by activation of caspases-9 and -3. HMJ-38 also activated ERK in HL-60 cells. Pre-incubating cells with ERK inhibitors (U0126 and PD98059) attenuated the HMJ-38-induced ERK activation and apoptosis. Nevertheless, cells remained arrested in G2/M. These results suggest that HMJ-38 is a potent anticancer drug and it shows a remarkable action on cell cycle before commitment for apoptosis is reached. PMID- 15274355 TI - Retinoids and cancer: antitumor effect of ATRA and of a new derivative of retinoic acid, IIF, on colon carcinoma cell lines CaCo-2 and HT-29. AB - Vitamin A and its metabolic forms, like all-trans retinoic acid (ATRA), are used with promising results in the treatment of many tumors. Two major problems in the clinical use of retinoids are that the doses needed for successful treatment are often toxic, leading to "hypervitaminosis A syndrome" and that patients often develop drug resistance. In order to find compounds that can overcome these problems, many new derivatives of retinoids have been synthesized and tested. Here we present a study on the effect of a new derivative of retinoic acid, IIF (pat. WIPO W0 00/17143), on growth and differentiation of two colon carcinomna cell lines, CaCo-2 and HT-29, with different degrees of tumorigenicity, the second one being more undifferentiated. The effect of IIF was compared with that of ATRA, whose antitumoral action on colon cancer cells and other tumoral cells is widely described in the literature. Besides exerting a strong antiproliferative effect, even higher than that of ATRA, IIF induced cellular differentiation, as demonstrated by the appearance of morphological (domes and microvilli formation) and biochemical (alkaline phosphatase induction) markers. Therefore, these findings indicate the new retinoid IIF as a possible candidate in the treatment of colon cancer. PMID- 15274356 TI - Intravenous chemotherapy with cisplatin for regional lymph node metastases of auricular VX2 carcinoma. AB - INTRODUCTION: The VX2 carcinoma is well established as a useful model for studies on treatment of primary tumors of various locations including the rabbit's auricle; however, limited experience exists on the treatment modalities of lymph node (LN) metastases. In this investigation we studied the frequency and extent of lymphogenic metastatic spread of auricular VX2-carcinomas and their response to systemic chemotherapy. MATERIALS AND METHODS: Induction of a right-sided auricular VX2-carcinoma in 17 healthy New Zealand white rabbits was followed by ablation of the right auricle and intravenous application of 1 mg/kg KG CDDP (cisdiamminedichloroplatinum (II)) dissolved in 2 ml NaCl via the left-sided auricular vein in 10 rabbits (group 1), while 7 rabbits (group 2) remained untreated. After a 24-day follow-up period, animals of both groups were sacrificed and the regional draining LN as well as the lungs were isolated and examined histopathologically. RESULTS: Group 1. Following intravenous cisplatin therapy (ICT), 6/10 animals showed no vital tumor cells within LN metastases of the first draining LN station, while 4/10 animals had necrotic LN metastases limited to the parotideal LN. Group 2. All 7 animals showed necrotic LN metastases of the first and second draining LN station as well as pulmonary metastases. CONCLUSION: The auricular VX2-carcinoma, characterized by frequent lymphogenic metastatic spread and response of LN metastases to ICT, offers an excellent animal model for further studies on the optimised treatment of lymphogenic metastatic spread in HNSCC. PMID- 15274357 TI - ProteinChip Array analysis of microdissected colorectal carcinoma and associated tumor stroma shows specific protein bands in the 3.4 to 3.6 kDa range. AB - Multiple pathways of carcinogenesis have been associated with colorectal carcinomas, including the adenoma-carcinoma sequence. The non polyposis coli gene has also been implicated in the pathogenesis of these tumors. Identification of the epithelial-mesenchymal interaction may help in understanding the pathways of invasion and may lead to the development of new, non-invasive tools for the diagnosis and prognosis of colon carcinomas. A ProteinChip Array technology (SELDI=Surface Enhanced Laser Desorption Ionization) has been developed enabling analysis and profiling of complex protein mixtures from a few cells. This study describes the protein analysis of approximately 500-1000 freshly obtained cells from normal and malignant colonic epithelium and its associated stroma by SELDI TOF-MS (Surface Enhanced Laser Desorption Ionization Time-of-Flight Mass Spectrometry). Pure cell populations of normal and malignant epithelium as well as stroma (without tumor cells) were selected by microdissection from 9 patients. A pattern of 3 peptides of 3.48, 3.55 and 3.6 kDa, which were increased in the colon tumor epithelium and stroma compared to associated normal colon and stroma in all 9 patients, was observed. Coupling microdissection with SELDI represents a powerful tool to identify cell and tumor specific proteins and to understand molecular events underlying the invasive event in colorectal carcinomas. The presence of certain proteins in invasive carcinomas may lead to the development of non invasive biomarkers for the identification or detection of recurrence of colorectal malignancies. PMID- 15274358 TI - Expression of bFGF, VEGF and c-met and their correlation with microvessel density and progression in prostate carcinoma. AB - BACKGROUND: Previously, we found angiogenesis measured as microvessel density (MVD) to be associated with both pathological stage and clinical outcome after radical prostatectomy (RP). In addition, we have shown that Vascular Endothelial Growth Factor (VEGF) is one of the important inducers of angiogenesis in prostate cancer (PC). The aim of this study was to investigate the expression of additional angiogenic factors, namely basic Fibroblast Growth Factor (hFGF) and the c-met receptor of Hepatocyte Growth Factor/Scatter Factor (HGF/SF) in PC. MATERIALS AND METHODS: Ninety-eight paraffin-embedded RP specimens and 20 adjacent normal prostatic tissues were evaluated for factor VIII staining and microvessel counting. Expression of VEGF (n=55), bFGF (n=65) and c-met (n=66) was studied by immunohistochemistry. Results were correlated with pathological grade and stage, MVD and clinical outcome. RESULTS: While adjacent benign tissue in RP specimens generally showed low MVD, VEGF-, bFGF- and c-met-expression, this was different in PC. All angiogenesis inducers were associated with stage while c-met as well as VEGF expression were associated with grade. Tumor progression was associated with grade and MVD. There was a clear correlation between VEGF and c met expression and MVD. CONCLUSION: VEGF and c-met expression increase with tumor stage and grade, while bFGF expression increases only with tumor stage. In addition to VEGF, c-met seems to be important and clinically relevant to the induction of angiogenesis in PC. Both VEGF and c-met appear to influence tumor progression, mainly through their effect on MVD. PMID- 15274359 TI - Combinatorial chemoprevention: efficacy of lovostatin and exisulind on the formation and progression of aberrant crypt foci. AB - BACKGROUND: There are several advantages to combinatorial chemoprevention strategies over monotherapeutic approaches. Both the HMG-CoA reductase inhibitor (HRI) lovastatin (LOV) and the selective apoptotic antineoplastic drug (SAAND) exisulind (EXS) have shown remarkable chemopreventive effects in previous studies, in cell lines and limited studies in rodents. Here, experiments were designed to assess the potential use of these two compounds in combinatorial chemoprevention therapy, using two bio-assays in which inhibition of the carcinogen-induced preneoplastic lesions, aberrant crypt foci (ACF), was used to quantitate efficacy. MATERIALS AND METHODS: ACF were induced by the carcinogen azoxymethane (AOM) in F344 rats by two sequential weekly i.p. injections at a dose of 15 mg/kg. F344 rats were fed seven experimental diets containing LOV @ 50 parts per million (ppm), EXS @ 100, 250 and 1000 ppm and combination diets containing EXS at 100, 250 and 1000 ppm, each combined with LOV @ 50 ppm. Quantification of ACF number and type (singlet, doublet, triplet and four or more) was performed on whole mounts of rat colons stained with 1.0% methylene blue. RESULTS: During the initiation protocol, administration of LOV @ 50 ppm alone and the combination of LOV @ 50 ppm with EXS @ 1000 ppm significantly decreased the mean number of ACF when compared to the positive control by 49% and 47%, respectively; however EXS @ 250 ppm displayed tumor promoting effects by significantly increasing the mean number of ACF by 64%. The post-initiation protocol administration of EXS @ 100, 250 and 1000 ppm and the combinations of LOV @ 50 ppm with EXS @ 100 and 250 ppm significantly increased the mean number of ACF when compared to the positive control by 44%, 48%, 55%, 49% and 40%, respectively. CONCLUSION: LOV shows greater promise than EXS in fulfilling the role as a supplemental chemopreventive agent in combinatorial chemopreventive strategies for cancers such as colon cancer. EXS did not augment this activity, failing to enhance chemopreventive therapy in this animal model. PMID- 15274360 TI - A continuous model studying T cell differentiation and lymphomagenesis and its distinction with discrete models. AB - The development of T-lymphocytes (T cells) constitute one of the basic and most vital processes in immunology. Conventional mathematical models, being based on the systems theory, fail to sufficiently distinguish the constituents of thymocytes and are thus of limited significance. On the basis of some well thought-out definitions and concepts, a continuous model was designed to describe T cell maturation in the thymus. A partial differential equation was first derived through the analysis of an infinitesimal element of the flow of thymocytes. A computation scheme was designed to determine the growth field in the thymus based upon available experimental data. The corresponding algorithm proved quite simple. A numerical example is given that focuses on the DN stage of the T cell development. The model opens a window for investigating the thymic microenvironment. The potential of the model in studying lymphomagenesis is discussed. PMID- 15274361 TI - Mevastatin-induced apoptosis and growth suppression in U266 myeloma cells. AB - Statins have been used successfully in the treatment of hypercholesterinaemia. Moreover, in vitro studies have shown that statins can trigger apoptosis in a variety of tumor cell lines. In the present study we analysed the effect of mevastatin--a novel inhibitor of HMG-COA reductase, the rate-limiting enzyme of the mevalonate pathway--on U266 human myeloma cells. Apoptosis induced by mevastatin was associated with increased caspase activity and depolarisation of the mitochondrial membrane. Expression of Bcl-2 mRNA and protein was down regulated, with no change in Bax or Bcl-XL protein production. The mitochondrial program was supported by caspase-8 and cleaved-Bid activity. None of the antibodies neutralizing the death-ligand/death-receptor pathway--TRAIL-R2Fc, anti TNF-alpha, anti-FASL(NOK-1)--influenced the mevastatin-induced apoptosis. Mevastatin also stimulated shedding of syndecan-1 from the surface of myeloma cells. The apoptosis inducing effect of mevastatin could be considered as a potential participant in a complex antitumor protocol. PMID- 15274362 TI - Profile of cytokines produced in tumor tissue after administration of cyclophosphamide in a combination therapy with tumor necrosis factor. AB - BACKGROUND: Recent studies have shown that locoregional administration of tumor necrosis factor (TNF) or production of endogenous TNF in tumor tissue with a chemotherapeutic agent showed considerable antitumor effect without severe toxicities. To analyze the immunological role of chemotherapeutic agents in locoregional TNF therapy, the effect of cyclophosphamide (CY) on cytokine production after ONO-4007 treatment was examined. MATERIALS AND METHODS: Seven days after inoculation of Meth A fibrosarcoma in BALB/c mice, CY (100-150 mg/kg) was injected intraperitoneally and, after another 7 days, ONO-4007 (30 mg/kg) was injected intravenously. A competitive PCR quantitative analysis of the induced cytokine messenger RNA (mRNA) was performed for IL-12 (p40), TGF-beta, TNF-alpha, IL-1beta, IL-10 and IL-6. RESULTS: The antitumor effect of ONO-4007 was enhanced by preadministration of CY. CY significantly enhanced IL-12 (p40) mRNA compared to that by ONO-4007 alone. No increase in TGF-beta mRNA was observed in either treatment. CONCLUSION: The preadministration of CY enhances IL-12 production in tumor tissues and shifts the cytokine profile to the dominant Th1 type. PMID- 15274363 TI - Presence of human papillomavirus in tonsillar cancer is a favourable prognostic factor for clinical outcome. AB - The purpose of this article is to review the current knowledge on the status and significance of human papillomavirus (HPV) in tonsillar cancer. Current data in scientific reports and data from the Karolinska Hospital and Karolinska Institute, Sweden, demonstrate that approximately half of all tonsillar cancer is HPV-positive. Moreover, patients with HPV-positive cancer have a lower risk of relapse and longer survival compared to patients with HPV-negative tonsillar cancer. The favourable outcome for patients harbouring HPV-positive tonsillar cancer cannot be attributed to increased radiosensitivity, since there is no significant difference in sensitivity to radiotherapy between HPV-positive and negative tonsillar cancer. However, HPV-positive cancer exhibits less genetic instability i.e. shows a lower degree of aneuploidy and a tendency to have fewer chromosomal aberrations, when compared to HPV-negative tonsillar cancer. PMID- 15274364 TI - Combined treatment with histamine dihydrochloride, interleukin-2 and interferon alpha in patients with metastatic melanoma. AB - BACKGROUND: Histamine inhibits phagocyte-derived production of reactive oxygen species and improves the anti-tumour efficiency of interleukin-2 (IL-2) and interferon-alpha (IFN-alpha) in vitro and in tumour-bearing animals. PATIENTS AND METHODS: In a phase-II study, twenty-seven patients with stage IV melanoma received subcutanous injections of histamine dihydrochloride (histamine) 1.0 mg and IL-2 2.4 MIU/m2 twice daily (BID) days 1-5 and 8-12. IFN-alpha 3 MIU once daily was administered throughout a cycle (days 1-28; n=14). Alternatively, bolus doses of IL-2 10 MIU/m2 BID days 1 and 2 and histamine days 1-28 (n=13) were administered. The aim was to study efficiency (survival and tumour response), toxicity and histamine pharmacokinetics. RESULTS: The median survival time was 11.3 (2.5-45) months. One patient achieved a complete response and 3 patients had partial responses. The compounds were safely self-administered with low toxicity. Plasma histamine concentrations significantly increased after an injection of histamine over 10 minutes (3 +/- 1 vs. 63 +/- 27 nmol/l). CONCLUSION: Histamine, IL-2 and IFN-alpha treatment is safe, well-tolerated and tumour responses were observed. The putative efficiency of histamine as an adjunct to cytokine therapy in metastatic melanoma needs to be confirmed in later randomized trials. PMID- 15274365 TI - Phase I study of combination therapy with S-1 and docetaxel (TXT) for advanced or recurrent gastric cancer. AB - BACKGROUND: S-1, an oral fluorouracil antitumor drug, and docetaxel have both been identified as effective agents for the treatment of gastric cancer. The two drugs have incompletely overlapping principal toxicities, which constitute the rationale for evaluating the effects of a combination of S-1 and docetaxel in this phase I study. The aim of this phase I study was to determine the maximum tolerated dose (MTD) and the recommended dose of docetaxel with a fixed dose of S 1 in patients with advanced or recurrent gastric cancer. PATIENTS AND METHODS: The pharmacokinetics of both drugs were evaluated on Day 1 of treatment. Patients with a performance status (PS) of 0 to 2 received docetaxel at the starting dose of 40 mg/m2 by i.v. infusion over 1 hour on Day 1 and S-1 at the fill dose of 80 mg/m2 daily for two weeks every three weeks. Nine patients were treated with increasing dose levels of docetaxel as follows: (docetaxel/S-1, mg/m2): 40/80 (Level 1), 50/80 (Level 2) and 60/80 (Level 3) and all the cases were found to be assessable for drug safety, while 7 were assessable for response. Colony stimulating factor (CSF) was not used in this study. The adverse effects of the treatment were analyzed according to NCI-CTC version 2, and the response was assessed according to the Japanese Classification of Gastric Cancer, 13th Ed. RESULTS: The MTD was reached at the 50/80 mg/m2 dose level in three patients out of six, who experienced a dose-limiting toxicity (DLT). The DLTs were neutropenia and allergic reactions. No hematological or non-hematological adverse effects (nore severe than Grade 2) were observed in any of the Level 1 patients. However among the Level 2 patients, 50% developed neutropenia (more severe than Grade 2), 33% developed loss of appetite, 17% developed diarrhea, 33% developed stomatitis and 17% developed allergic reactions. On the other hand, partial response was achieved in 5 (71.4%) of the 7 patients with evaluable lesions. The pharmacokinetics of docetaxel were not altered as compared to that in the historical controls by the administration of S-1. These results indicate that the recommended doses of the two drugs in the combination therapy would be 40 mg/m2 for docetaxel and 80 mg/m2 for S-1. CONCLUSION: The drug combination showed a good safety profile, with neutropenia being a common but manageable adverse reaction. Moreover, the responses observed in the study suggest that the drug combination shows a high degree of efficacy in patients with advanced and or recurrent gastric cancer. PMID- 15274366 TI - Neoangiogenesis in patients with gastric carcinoma in relation to the expression of vascular endothelial growth factor and thymidine phosphorylase. AB - BACKGROUND: We investigated the prognostic significance of microvessel density and the relationship between the expression of vascular endothelial growth factor (VEGF) and thymidine phosphorylase (TP), and angiogenesis in patients with gastric carcinoma. MATERIALS AND METHODS: The expression of VEGF and TP, and the microvessel density were examined by immunohistochemistry in patients with gastric carcinoma invading the serosa. RESULTS: The prognosis of patients with low microvessel density in the cancerous tissue was significantly better than that of patients with high microvessel density. A multivariate analysis showed that microvessel density, lymph node metastasis and tumor size were independent prognostic indicators. VEGF was expressed in tumor cells and TP was expressed in both tumor cells and infiltrating cells. VEGF expression in tumor cells and TP expression in infiltrating cells significantly correlated with microvessel density. However, microvessel density was not correlated with TP expression in tumor cells. Combined analysis based on VEGF expression in tumor cells and TP expression in infiltrating cells revealed that microvessel density was the highest in VEGF-positive and TP-positive tumors and the lowest in VEGF-negative and TP-negative tumors. Microvessel density is an independent prognostic indicator in patients with gastric carcinoma invading the serosa. CONCLUSION: VEGF expression in tumor cells and TP expression in infiltrating cells may indicate the microvessel density. PMID- 15274367 TI - Autologous natural killer cell therapy for human recurrent malignant glioma. AB - BACKGROUND: Natural killer (NK) cells are highly efficient in the cellular immune response against malignant tumors without restriction of major histocompatibility complex. However clinical studies using autologous NK cells have been reported in only a very limited number of cases, due to the fact that selective NK expansion is difficult to achieve in this patient population. Here, we report the results of adoptive immunotherapy in patients with recurrent malignant gliomas using autologous NK cells that were expanded ex vivo by a novel method. PATIENTS AND METHODS: Peripheral blood mononuclear cells (PBMCs) were prepared from patients with malignant gliomas, and were co-cultured with an irradiated human feeder cell line (HFWT) in RHAM-alpha medium supplemented with 5% autologous plasma and interleukin-2. The resulting NK cell-rich effector cells were injected into 9 patients (16 courses) with recurrent malignant glioma (6 cases of WHO grade-3 glioma and 3 cases of grade-4 glioma). RESULTS: The mean frequency of NK cells among lymphocytes was 82.2 +/- 10.5%. A combination of focal and intravenous injections was peformed in 10 courses. Intravenous injection alone was performed in 6 courses. Further, intravenous injection of low-dose interferon beta (6x10(6) IU/week) was performed as an adjuvant therapy in all courses to achieve maximum benefit for enrolled patients. Clinical evaluation demonstrated 3 PR, 2 MR, 4 NC and 7 PD in a total of 16 courses of treatment. Severe neurological toxicity was not observed in any of the patients. CONCLUSION: It was demonstrated that NK cell rich effector cells were expanded ex vivo from PBMCs in all nine cases of recurrent malignant glioma and that NK cell therapy was safe and partially effective in patients with recurrent malignant gliomas. PMID- 15274368 TI - Gefitinib as salvage therapy in pretreated patients with advanced non-small cell lung cancer. Data from a compassionate use program. AB - PURPOSE: To evaluate the tolerabiliy and activity of gefitinib, an epidermal growth factor receptor tyrosine kinase inhibitor, in patients with pretreated advanced non-small cell lung cancer (NSCLC). MATERIALS AND METHODS: From July 2001 to August 2003, we analysed, retrospectively, the data of 125 previously treated advanced NSCLC patients receiving gefitinib 250 mg, orally once daily in a compassionate use program. RESULTS: Main toxicity was (% of patients): grade 1 2 skin changes in 7 (5.0%) and 8 (6.4%) patients, respectively. Grade 1 diarrhea in 16 (12.8%) patients. Grade 1 and 2 hypertransaminasemia in 1 (0.8%) patient, respectively. Grade 2 onychopathy in 1 (0.8%) patient and epistaxis in 1 (0.8%) case. There were 1 complete response (0.8%), 6 partial responses (4.8%) and 22 (17.6%) stable disease with an overall control of disease in 23.2% of cases. Median progression-free survival and overall survival were 8.8 and 21.5 weeks, respectively. CONCLUSION: Gefitinib is active, feasible and well-tolerated in pretreated patients affected by advanced NSCLC. PMID- 15274369 TI - A randomized consent design trial of neoadjuvant chemotherapy with tegafur plus uracil (UFT) for gastric cancer--a single institute study. AB - OBJECTIVE: Various forms of neoadjuvant chemotherapy (NAC) have been applied in the treatment of gastric cancer. The present study was designed to assess the survival benefits of NAC with UFT (tegafur plus uracil) for gastric cancer, as a randomized consent trial as described by Zelen. PATIENTS AND METHODS: The present study included 295 patients with resectable gastic cancer between 1991 and 1999. After the patients had been pre-randomized into two groups, a control (no-NAC) group (n=120) and a treatment group (n=175), the treatment group patients were then further stratified into two groups, namely those who wished to join the control group and those who wished to receive NAC with UFT (NAC-UFT group). Patient outcome was surveyed in January 2003. RESULTS: Randomization did not necessarily result in an appropriate registration of the patients, and ultimately 193 patients were included in the control group and 102 patients received NAC UFT. The NAC-UFT was well tolerated by the patients and side-effects were not severe. However, the NAC-UFT group included the patients with significantly higher stages of cancers than the control group. The survival benefit of NAC-UFT was seen in stage 2 or 3 patients, and multivariate analysis also revealed that NAC-UFT was a significant prognostic variable, as were pT, pN, M and the level of nodal dissection, but patient age, gentler and histological grade were not significant variables. CONCLUSION: NAC-UFT may be beneficial in the improvement of survival rate after gastric cancer surgery and this treatment modality is worthy of further study with a larger patient sample size. PMID- 15274370 TI - Acetabular osteosarcoma treated by irradiation-vascularized hybrid bone graft. AB - BACKGROUND: Periacetabular reconstruction after malignant bone tumor resection to preserve limb function is extremely challenging. The optimal reconstruction method has not yet received consensus and the functional outcomes still remain unsatisfactory. CASE REPORT: A thirteen-year-old girl who was suffering from acetabular osteosarcoma was treated with wide excision of the tumor, followed by periacetabular reconstruction with an autogenous extracorporeal irradiated osteoarticular graft combined with a vascularized fibula graft. Incorporation of the irradiated pelvic bone was achieved despite infection and limb function turned out to be acceptable. CONCLUSION: This is the first report on an irradiation-vascularized hybrid autograft to reconstruct massive bone loss in primary limb-sparing surgery. PMID- 15274371 TI - Histological findings in a human autogenous pasteurized bone graft. AB - BACKGROUND: Autogenous bone graft after pasteurization is one of the most valuable procedures for reconstruction of large bone defects following excision of malignant musculoskeletal tumors. To date, there have been no documented histological reports on pasteurized bone grafts, apart from short-term histological results. CASE REPORT: We treated a 27-year-old male with a periosteal chondrosarcoma of the tibia by wide excision and reimplantation of the large pasteurized bone. Biopsy specimens harvested from the pasteurized bone over 3 years after reimplantation were evaluated histologically. The graft cortices remained totally necrotic with empty osseous lacuna, whereas the architecture of the acellular cortical bones was still maintained without microfractures. Deposited seams of woven bone existed focally on the surface of the acellular trabeculae. CONCLUSION: Our medium-term histological outcome suggests the limitations of incorporating a pasteurized hone graft, but also advocates its role as a useful temporary material for the reconstruction of massive bone defects. PMID- 15274372 TI - Kaposi's sarcoma of the tongue associated with median rhomboid glossitis in a non AIDS patient. A case report. AB - Kaposi's sarcoma (KS) of the tongue is extremely rare in immunocompetent patients. We report a case of KS of the tongue associated with a median rhomboid glossitis. The main clinical, pathological and immunohistochemical features allowed the differential diagnosis. PMID- 15274373 TI - Long-term survival over 28 years of a patient with metastatic adrenal cortical carcinoma--case report. AB - Adrenal cortical carcinoma (ACC) is a rare and highly malignant tumour with up to 70% of the patients diagnosed at an advanced clinical stage, up to 40% presenting with metastases. Even after complete surgical excision, up to 80% of the patients show locoregional recurrence or metastases. We report a case of a 62-year-old woman with a non-functional ACC of the left adrenal gland (T2N0M0 classified as stage II). After the initial resection, 3 operations for metastasis of the contralateral adrenal gland and 4 operations for metastasis of the lungs were carried out, allowing survival for more than 28 years with a good quality of life. This case report emphasises the need for careful clinical and radiographic follow-up. A repeat surgical approach should be adopted whenever possible providing long-term survival over decades. PMID- 15274374 TI - Primary choriocarcinoma of the lung. AB - Primary choriocarcinoma (PCC) of the lung is extremely rare, with about 25 cases previously reported. We describe here a patient with PCC of the lung and review previously described cases to establish the clinical characteristics and most appropriate management of this disease. Survival for patients treated with both surgery and chemotherapy was longer than for patients treated with surgery alone, chemotherapy alone, or optional supportive care. Multivariate analysis of prognostic factors revealed that treatment with both surgery and chemotherapy had independent prognostic significance. Our analysis including previous cases suggests that resection followed by adjuvant chemotherapy is the most effective treatment. PMID- 15274375 TI - Validation of the limited-sampling models for carboplatin AUC in combination chemotherapy with taxanes. AB - We previously developed limited-sampling models (LSM) for estimation of the area under the concentration curve (AUC) of free carboplatin (Anticancer Res 17: 4571 4576, 1997). The one-point model was: AUC (mg/ml x min) = 0.93 x C3h + 0.47 and the two-point model was: AUC (mg/ml x min) = 0.16 x C1h + 2.26 x C8h + 0.75, in which C3h, C1h and C8h represented the plasma concentration, sampled at 3 h, 1 h and 8 h after the start of a 1-h carboplatin infusion, respectively. In this study, we tried to confirm the utility of our LSMs in combination chemotherapy with taxanes. We obtained 29 data series from 29 patients who received carboplatin in combination with docetaxel (n = 16) or paclitaxel (n = 13). The actual AUC was strongly correlated with the AUC calculated from both one- and two point models (one-point model; the mean prediction error (MPE) = -4.44 +/- 2.15%, the root mean squared error (RMSE) = 12.22 +/- 0.84%, two-point model; MPE = 1.89 +/- 1.66%, RMSE = 8.99 +/- 0.81%). Our LSMs proved to be unbiased and precise and are also useful for calculating carboplatin AUC in combination with taxanes. PMID- 15274376 TI - A retrospective study focusing on clinical predictive factors in 126 patients with oesophageal carcinoma. AB - In Sweden, approximately 400 patients are diagnosed each year with oesophageal carcinoma. Despite the introduction of different treatment schedules, only modest improvements in survival have been accomplished. To be able to select patients in whom a more favourable outcome of radiation/-chemotherapeutic treatment could be expected, the present study reviewed the charts from 126 consecutive patients with oesophageal carcinoma. All patients were treated at the Department of Oncology, Uppsala University Hospital, Sweden, between 1990 and 2000. The charts were reviewed with focus on known and potential prognostic factors. Performance status, smoking habits, swallowing function, localisation of the tumour, leucocytes and albumin levels at first admittance, and stage of the disease were prognostic factors. However, performance status and stage of the disease only remained as significant independent prognostic factors in the multivariate analysis (both with p-values < 0.001). The results imply that further characterisation of tumour biology in oesophageal carcinoma is needed to find additional predictive factors for survival and future treatment strategies. PMID- 15274377 TI - Significance of T stadium and grading as prognostic factors in transitional cell carcinoma of the ureter. AB - BACKGROUND: Surgery is nowadays the standard treatment for carcinoma of the ureter, even if adjuvant therapies can modify the prognosis in selected patients. Because of the small number of patients in the literature series, the significance of prognostic factors that can be used in clinical practice is still controversial, as is the choice of the most suitable surgical and adjuvant treatment. PATIENTS AND METHODS: We considered 27 consecutive patients (Ta-T2 N0 M0) who underwent radical surgery (nephroureterectomy with bladder cuff excision and lymphoadenectomy) for transitional cell carcinoma of the ureter, from 1982 through 1992. Seven patients (25.9%) had Ta tumors, 7 patients (25.9%) had T1 tumors and 13 patients (48.2%) had T2 tumors. In 4 cases (14.8%) the tumor was well-differentiated (G1), in 14 cases (51.8%) it was mildly-differentiated (G2), and in 9 cases (33.4%) it was poorly-differentiated. RESULTS: Thirteen of the 14 patients affected by Ta-T1 tumors were alive 10 years after surgery (one patient lost at follow-up); in the T2 tumor group the 5-year survival rate was 84.6% and 10-year survival rate was 69.2%. According to grading, the 10-year survival rate was 100% for G1 tumors, the 3, 5 and 10-year survival rates were, respectively, 100%, 92.8% G3 tumors. CONCLUSION: Data from our study show the significance of the T stage and grading as prognostic factors. PMID- 15274378 TI - Bcl-2 protein expression correlates with better prognosis in patients with advanced non-small cell lung cancer. AB - BACKGROUND: Regulation of apoptosis is an important mechanism during the development of tumors including non-small cell lung cancer (NSCLC). The aim of this study was to assess the relationship between p53 and bcl-2 expression and various clinicopathological features and survival in patients with NSCLC. MATERIALS AND METHODS: Histological specimens obtained from 120 patients with stage I-III NSCLC were examined immunohistochemically for p53 and bcl-2. RESULTS: Positive immunostaining for p53 was observed in 50 and for bcl-2 in 35 patients. Tumors with lymph node metastasis were significantly more likely to be bcl-2 positive. However, there was no correlation between p53 immunostaining and clinicopathological parameters. Cox proportional hazard multiple regression analyses identified gender, N status and bcl-2 expression as independent prognostic factors. When advanced stage tumors or tumors with lymph node metastasis were analyzed, a more favorable survival was noted in patients with bcl-2-positive tumors than those with bcl-2-negative tumors. CONCLUSION: Bcl-2 protein expression correlates with better prognosis in patients with advanced NSCLC. PMID- 15274379 TI - Preoperative chemolipiodolization of the whole liver for hepatocellular carcinoma. AB - BACKGROUND: Preoperative transarterial chemoembolization is not routinely recommended before hepatectomy for resectable hepatocellular carcinoma. This study evaluated the effect of preoperative whole-liver chemolipiodolization. PATIENTS AND METHODS: A retrospective comparison of background characteristics, operative results and long-term outcome was performed between 36 patients with chemolipiodolization confined to the tumor (selective group) and 23 patients with chemolipiodolization also involving the noncancerous liver (whole-liver group). RESULTS: There were no serious side-effects in either group and the operative outcome did not differ between the two groups. Tumor diameter was significantly greater in the selective group, but other pathological characteristics were comparable. The 5-year disease-free and overall survival rates for the selective and whole-liver groups were 11.9% and 33.0% (p=0.0191) and 44.9% and 73.2% (p=0.0121), respectively. CONCLUSION: These results indicate that preoperative whole-liver chemolipiodolization reduces postoperative recurrence and prolongs survival in patients undergoing resection of hepatocellular carcinoma. PMID- 15274380 TI - Oxaliplatin fractionated over two days with bimonthly leucovorin and 5 fluorouracil in metastatic colorectal cancer. AB - PURPOSE: To evaluate the efficacy and safety of oxaliplatin (L-OHP) fractionated over two days with bimonthly 5-fluorouracil (5-FU) and leucovorin (LV), as first line treatment of metastatic colorectal cancer (MCC) patients. PATIENTS AND METHODS: Fifty-four patients with inoperable MCC (median age, 60 years) were entered into the study. Outpatient treatment consisted of L-OHP 50 mg/m2 and LV 200 mg/m2 administered in a 2-hour i.v. infusion, followed by 5-FU 400 mg/m2 bolus and 5-FU 600 mg/m2 in a 22-hour continuous infusion, on days 1 and 2 every 2 weeks. RESULTS: A total of 488 courses of chemotherapy were administered. Responses were as follows: 3 complete responses (5.6%) and 24 partial responses (44.4%) giving an overall response rate of 50% (95% CI: 36% to 64%). Median time to progression and overall survival were 10.3 and 19.2 months, respectively. Grade 3-4 neutropoenia, leucopoenia, thrombocythopoenia and anaemia occurred in 33%, 9%, 2% and 2% of patients, respectively, while peripheral neuropathy occurred in 10% of patients. CONCLUSION: The fractionated bimonthly schedule of L OHP plus de Gramont gave a less than anticipated neurological toxicity profile, while maintaining expected efficacy. PMID- 15274381 TI - The prognostic value of both neuron-specific enolase (NSE) and Cyfra21-1 in small cell lung cancer. AB - PURPOSE: The aim of this study was to verify the prognostic significance of multiple tumour markers in small cell lung cancer (SCLC). PATIENTS AND METHODS: We examined seven tumour markers [carcinoembryonic antigen (CEA), cancer antigen 19-9 (CA 19-9), squamous cell carcinoma antigen (SCC), neuron-specific enolase (NSE), cancer antigen 125 (CA125), cytokeratin 19 fragment (Cyfra21-1) and ProGastrin-releasing peptide (ProGRP)] in 57 small cell lung cancer (SCLC) patients. RESULTS: Univariate analysis showed that NSE and Cyfra21-1 were independent negative prognostic factors along with gender, therapy and lactate dehydrogenase (LDH). Multivariate analysis showed that both NSE and Cyfra21-1 retained their significance as prognostic factors along with therapy and the respective hazard ratios were 3.918 (p=0.0122) and 2.617 (p=0.0318) among the seven tumour markers. The group with both NSE and Cyfra21-1 positive had a worse prognosis than the only NSE-positive group, with the respective hazard ratios being 10.245 (p=0.0004) and 3.913 (p=0.0123). CONCLUSION: The group with both of the markers NSE and Cyfra21-1 positive had a worse prognosis than the only NSE positive group. PMID- 15274382 TI - Tolerability of adjuvant high-dose interferon alfa-2b: 1 month versus 1 year--a Hellenic Cooperative Oncology Group study. AB - BACKGROUND: High-dose interferon alfa-2b (IFN-alpha2b) as adjuvant therapy for melanoma is associated with substantial dose-limiting toxicity. It has been suggested that the 1-month intravenous (i.v.) induction regimen may be sufficient to reduce the risk of relapse and death. PATIENTS AND METHODS: The Hellenic Cooperative Oncology Group is conducting a multicenter, randomized trial of 1 month i.v. induction versus 1 year of adjuvant IFN-alpha2b therapy in patients with stage IIB/III melanoma. Adverse events reported by the first 200 patients to complete therapy are described. RESULTS: Both induction and maintenance regimens were well tolerated. The most common toxicities were flu-like and gastrointestinal symptoms, neutropenia, liver toxicity, and neurologic toxicity. The incidence of grade 3/4 toxicity was low and occurred mainly during the induction phase in both arms. Dose was reduced in 31% of patients during induction. Only 2% of patients discontinued. Dose was reduced in 8% of patients during maintenance and only 5% of patients discontinued. CONCLUSION: Intravenous induction with 15 MIU/m2/day IFN-alpha2b is well tolerated. Efficacy results from this trial are eagerly anticipated. PMID- 15274383 TI - CYFRA 21-1 and CEA are independent prognostic factors in 153 operated stage I NSCLC patients. AB - Currently, no further therapy in addition to surgery is recommended in completely resected NSCLC stage I patients. However, the 5-year survival rate at this stage has been reported to be approximately 60%, i.e. 40% of patients had a lower survival rate. The aim of the study was to identify those patients at increased risk by using the tumor markers CYFRA21-1 and CEA as prognostic factors. One hundred and fifty-three stage I NSCLC patients, who were treated exclusively by surgery between 1996 and 1998, entered this retrospective study. It was shown, by multivariate analysis, that elevated CYFRA 21-1 (>3.3ng/ml) and CEA (>9.8ng/ml) levels were associated with a worse outcome in 21.3% and 13.1% of the patients under study, respectively. The corresponding 3-year survival rates were found to be 60.2% for increased CYFRA 21-1 levels (p=0.029) and approximately 40% for increased CEA levels (p=0.022), compared to a rate of 78.4% and 79.0% in case of normal marker levels, respectively. The relative risk (95% confidence interval) was found to be 2.156 (1.08-4.29) for elevated CYFRA 21-1 and 2.707 (1.15-6.36) for elevated CEA. The detection rate for the identification of patients with worse outcome increased when a combination of both markers was used. Thereby, it was possible to identify 32% of patients where one or both markers were elevated. The 3-year survival rate was 55.7% in this group compared to that of 82.5% in those patients where both markers were in the normal range (p=0.0014). In order to consider the degree of marker elevation that is thought to reflect tumor burden, we introduced a tumor marker index (TMI) corresponding to the geometric mean of normalized CYFRA21-1 and CEA levels (marker value divided by diagnostic cut-off). Thereby, we were able to identify 3 groups of patients at different risk levels: the first group (22.7%) had a 3-year survival rate of 96.7%, the second group (42.6%) had one of 77.2% and the third group (34.7%) had one of only 55.7%. In conclusion, elevated CYFRA 21-1 and CEA levels were able to identify a group of curatively operated NSCLC patients who were at high risk of early death. Those patients may benefit from more aggressive treatment approaches. The group of patients with a 3-year survival rate of 96.7% probably does not need further treatment. PMID- 15274384 TI - Epoetin beta (NeoRecormon) corrects anaemia in patients with hormone-refractory prostate cancer and bone metastases. AB - BACKGROUND: Severe anaemia is common in patients with metastatic, hormone refractory prostate cancer (HRPC). PATIENTS AND METHODS: We evaluated the efficacy of epoetin beta in correcting anaemia and maintaining haemoglobin (Hb) levels in this group of patients. Patients with HRPC, bone metastases and anaemia (Hb < 12 g/dl) were included. Epoetin beta, 30,000 IU per week in three divided doses, was administered subcutaneously, with iron supplementation when needed. If Hb increased by < 1 g/dl during the first 4 weeks of therapy the epoetin dose was increased (increments of 5,000 IU per dose) at fortnightly intervals to a maximum of 60,000 IU per week. Patients with haematopoietic response (Hb increase > or = 2 g/dl from baseline or Hb level > or = 12 g/dl without blood transfusions) went on to receive epoetin beta 10,000 IU once weekly for up to 24 weeks. RESULTS: All 29 evaluable patients demonstrated a haematopoietic response to epoetin beta treatment. None of the patients required blood transfusions. All patients showed improvements in quality of life (assessed using the EORTC QLQ-C30 questionnaire). Hb levels were maintained for the remainder of the trial. Epoetin beta was very well tolerated. CONCLUSION: Epoetin beta therapy resulted in a rapid and sustained improvement in Hb levels in patients with HRPC metastatic to bone. PMID- 15274385 TI - Epirubicin plus docetaxel in metastatic breast cancer: escalating dose does not improve efficacy. A phase II study. AB - BACKGROUND: The combination of anthracyclines and docetaxel have demonstrated a significant activity in metastatic breast cancer (MBC) as first-line chemotherapy. In a previous multicenter phase I study, we recommended two schedules of epirubicin-docetaxel combination for MBC: 1) epirubicin 75 mg/m2, docetaxel 80 mg/m2 every 3 weeks without G-CSF; 2) epirubicin 90 mg/m2 plus docetaxel 90 mg/m2 every 3 weeks, with G-CSF support. PATIENTS AND METHODS: Twenty-five advanced breast cancer patients were treated with epirubicin 90 mg/m2 plus docetaxel 90 mg/m2 every 3 weeks, with prophylactic G-CSF. RESULTS: The main toxicity was grade 3-4 neutropenia (41% of cycles) despite the use of G-CSF; febrile neutropenia was observed in 14% of cycles necessitating a dose reduction of both drugs in 30% of patients. Response was observed in 79% of patients: 21% complete responses and 58% partial responses. The median response duration was 10 months (range: 3-16). The median time to progression was 11 months. The overall 3 year survival was 49.7%. CONCLUSION: The antitumor activity observed in this series was comparable with that seen in other studies of taxane/anthracycline combinations. The degree of myelosuppression was severe, even though G-CSF was administered as a prophylactic. We recommend a lower dose of both drugs as reported by other authors. PMID- 15274386 TI - 5FU and oxaliplatin-containing chemotherapy in two dihydropyrimidine dehydrogenase-deficient patients. AB - Patients with a germline mutation leading to a deficiency of the dihydropyrimidine dehydrogenase (DPD) enzyme are at risk from developing severe toxicity on the administration of 5FU-containing chemotherapy. We report on the implications of this inborn genetic error in two patients who received 5FU and oxaliplatin. A possible co-medication effect of oxaliplatin is considered, as are the consequences of screening for DPD deficiency. PMID- 15274387 TI - Correlation of serum VEGF levels with clinical stage, therapy efficacy, tumor metastasis and patient survival in ovarian cancer. AB - Vascular endothelial growth factor (VEGF) has been shown to play an important role in tumor growth and progression. However, the clinical implications of VEGF expression in ovarian tumors are not fully understood. We therefore investigated the serum level of VEGF in patients with ovarian tumors and explored the potential use of VEGF as a tumor marker for diagnosis, treatment and prognosis of human ovarian cancer. The serum VEGF (sVEGF) levels in 120 patients with ovarian carcinoma, 25 patients with benign ovarian tumor and 90 healthy female blood donors were measured by an enzyme-linked immunosorbent assay in this study. We also determined the levels of sVEGF in patients with epithelial ovarian cancer before and after surgery. Our results showed that: (i) ovarian cancer patients had significantly higher levels of sVEGF compared to those of patients with benign ovarian tumor or those of healthy individuals. As a cut-off at 100 pg/ml, the sensitivity and specificity of sVEGF levels for diagnosing ovarian carcinoma were 77.1% and 87%, respectively. (ii) sVEGF levels were markedly elevated in patients with advanced stage or poorly-differentiated ovarian cancer, as well as in those with more ascites (>500 ml), as compared to patients with early stage and well-differentiated ovarian cancer, or those with less ascites (<500 ml). However, there was no significant difference in sVEGF levels among different pathological subtypes of ovarian carcinoma. (iii) The post-operative sVEGF levels were significantly lower than the pre-operative sVEGF levels. (iv) We measured significantly higher levels of sVEGF in patients with metastasis as compared to patients lacking metastasis. Lastly (v) the average survival-time in patients with higher levels of sVEGF (>100 pg/ml) was 28 months, while the average survival-time in patients with lower levels of sVEGF (<100 pg/ml) was 35 months, indicating that the elevations in sVEGF level are correlated with patient survival and tumor metastasis in ovarian carcinoma. These data suggest that VEGF may be a useful serological biomarker for clinical diagnosis and prognosis of ovarian cancer, for follow-up of ovarian tumor metastasis and for monitoring the efficacy of therapy in patients with ovarian carcinomas. PMID- 15274388 TI - The prognostic value of plasma soluble urokinase plasminogen activator receptor (suPAR) levels in stage III ovarian cancer patients. AB - The level of the urokinase plasminogen activator receptor (uPAR) is elevated in tumor tissue from several forms of cancer. uPAR is shed from the cell surface and the soluble form, soluble urokinase plasminogen activator receptor (suPAR), has been detected in several body fluids. High plasma levels of suPAR in patients with colorectal cancer and high serum levels of suPAR in patients with recurrent metastatic breast cancer have been associated with poor prognosis. In patients with ovarian cancer (OC) it has been shown that the level of suPAR is very high in ascites and cystic fluid and that high serum levels of suPAR were associated with shorter survival of the patients. We evaluated suPAR preoperatively in plasma from primary OC stage III patients and tested for association with prognosis. The prognostic significance of suPAR was also compared to two biochemical markers; cancer antigen 125 (CA125) and tetranectin (TN). No significant differences were found between patients who died of OC compared to patients still alive regarding median plasma suPAR levels (p=0.62) and median serum CA125 levels (p=0.26). In contrast, a significant difference was found between dead and alive OC patients for the median serum TN level (p<0.0001). Dividing the patients into two groups, corresponding to preoperative plasma suPAR levels below or equal to 2.0 ng/ml and higher than 2.0 ng/ml, no significant difference in survival was found between the two groups (p=0.49). When different cut-off levels of plasma suPAR were considered (2.74 ng/ml, 3.25 ng/ml and 4.18 ng/ml), no significant differences in survival could be detected (p=0.58, p=0.68 and p=0.05). Multivariate Cox regression analysis showed that the only independent prognostic factors were radicality after primary surgery (RH=5.34; 95% CI, 2.34-12.20; p<0.0001) and preoperative serum TN (RH=0.69, 95% CI, 0.57 0.82; p<0.0001), whereas plasma suPAR (4.18 ng/ml), age, histological type of tumour and serum CA 125 had no independent prognostic value. In conclusion, preoperative plasma suPAR level was of no prognostic value in this cohort of Danish stage III OC patients. PMID- 15274389 TI - Prognostic significance of serum HER2 and CA 15-3 at the time of diagnosis of metastatic breast cancer. AB - BACKGROUND: Serum HER2 testing allows the determination of real-time HER2 status during clinical course. The aim of this investigation was: (1) to study the prognostic significance of serum HER2 at the time of first diagnosis of metastatic breast cancer and (2) to evaluate its relationship to CA15-3 which is a surrogate marker for tumor load. MATERIALS AND METHODS: Serum samples of 120 breast cancer patients were assayed for HER2 and CA15-3 at the onset of metastatic disease. RESULTS: Forty-seven out of 120 (39%) metastatic breast cancer patients had elevated serum HER2 levels. The positivity rate of CA15-3 was 51%. The median survival after relapse (SAR) for HER2-positive patients was shorter (10 months, 95%-CI: 6-14 months) compared to the SAR of HER2-negative patients (19 months, 95%-CI:15-23 months) (p<0.01). The median survival of patients with increased CA15-3 was 13 months (95%-CI: 9-17 months) compared to 18 months (95%-CI: 15-21 months) for patients with normal CA15-3 concentrations (p<0.05). In the multivariate analysis serum HER2 was an independent prognostic marker for SAR even when adjusted for tumor load measured by CA15-3 levels. CONCLUSION: Serum HER2 is a strong independent prognostic factor for survival after relapse in metastatic breast cancer even when adjusted for tumor load. Therefore, the prognostic significance of serum HER2 may not only be related to the tumor load but also to the biological behavior of the tumor. PMID- 15274390 TI - Peroxisome proliferator activated receptor gamma immunohistochemical expression in human papillary thyroid carcinoma tissues. Possible relationship to lymph node metastasis. AB - BACKGROUND: Peroxisome proliferator activated receptor gamma (PPARgamma) involvement in thyroid tumorigenesis has recently been studied, especially in follicular neoplasms. Conflicting results concerning the regulation of this receptor in human papillary carcinoma have been reported. Therefore, we quantitatively assessed PPARgamma immunohistochemical expression in papillary carcinoma in comparison with other types of thyroid tumors and we evaluated its relationship with clinical criteria of aggressiveness. MATERIALS AND METHODS: Immunohistochemistry (IHC) was performed on 56 human thyroid papillary carcinomas (PTC), 9 follicular carcinomas (FTC), 20 follicular adenomas (FA) and 18 Hurthle cells adenomas. PTC were divided into subgroups according to some aggressiveness criteria: tumor size, capsular invasion, lymph node metastasis. Immunostaining was semi-quantitatively analyzed using image analysis software. RESULTS: Strong nuclear PPARgamma expression was detected in a large number of PTC (42%), similar to that found in FTC (44%) or FA (63%). Only Hurthle cell adenoma showed a significantly lower proportion of PPARgamma-positive immunoreactivity (11%, p<0.05). Cases of PTC-associated lymph node metastasis showed a higher percentage of PPARgamma-positivity than other case categories (63% vs. 20%), a result which was also noticed when comparing large PTC with infracentimentric tumors (60% vs. 39%, p<0.05). CONCLUSION: These results, combined with recently published data, suggest that the intense PPARgamma immunostaining revealed in PTC could be related to high wtPPARgamma gene levels. Moreover, they corroborate a strong relationship between PPARgamma expression and tumor progression. PPARgamma IHC evaluation is not a valuable differential diagnostic tool for thyroid tumors but it could be a reliable marker of papillary carcinoma aggressiveness and a potential predictor for an eventual therapy by PPARgamma agonists. PMID- 15274391 TI - 99m-Tc-MDP-Scintimammography in the evaluation of breast masses or tumor angiogenesis. AB - BACKGROUND: Vascular endothelial growth factor (VEGF) is a prognostic factor in breast cancer. One of the mechanisms of extra-skeletal uptake of Tc-99m methylenediphosphonate (MDP) is suggested to be tumor vascularity. We studied the correlation between MDP uptake and VEGF and compared the diagnostic accuracy of mammography versus MDP-scintimammography (MDP-S). MATERIALS AND METHODS: Fifty four patients with suspicion of breast cancer were evaluated. Breast images were collected 5-10 minutes after injection of Tc-99m-MDP prior to biopsy. Tissue slides were stained using a rabbit-polyclonal anti-VEGF. RESULTS: MDP-S showed a diagnostic accuracy of 83.3% in BIRADS category 4 lesions. Four out of 23 benign lesions were false-positive. The tumor to background (TM/BG) ratio of early images of MDP-S was correlated with VEGF staining (p: 0.014) and with tumor size (p: 0.006). CONCLUSION: Early images of Tc-99m-MDP-S may satisfactorily identify cancers with increased neovascularization. MDP-S seems to be an accurate imaging modality, especially in BIRADS category 4 lesions. PMID- 15274392 TI - Expression of betacellulin, heparin-binding epidermal growth factor and epiregulin in human malignant fibrous histiocytoma. AB - BACKGROUND: Heparin-binding epidermal growth factor (HB-EGF), betacellulin (BTC) and epiregulin (EPR) are members of the EGF system and involved in the cell growth of various epithelial malignancies. There have been no reports on the HB EGF, BTC and EPR expression in mesenchymal malignancies of fibrohistiocytic origin including malignant fibrous histiocytoma (MFH). MATERIALS AND METHODS: We investigated the expression of HB-EGF, BTC, EPR and EGF-receptor (EGF-R) in 43 human MFH tissue samples using immunohistochemical techniques. RESULTS: Positive immuno-reactivity for HB-EGF, BTC, EPR and EGF-R was identified in 28 (65%), 7 (16%), 43 (100%) and 36 (84%) out of the 43 MFH cases analyzed, respectively. Coexpression of HB-EGF/BTC, BTC/EPR and HB-EGF/EPR was observed in 6 (14%), 7 (16%) and 28 (65%) of the MFHs, respectively. Coexpression of HB-EGF/EGF-R, BTC/EGF-R and EPR/EGF-R was observed in 25 (58%), 6 (14%) and 36 (84%) of the MFHs, respectively. CONCLUSION: These results revealed that HB-EGF, BTC and EPR are expressed not only by epithelial tumor cells, but also by MFH cells. It is suggested that HB-EGF and EPR might be more important tumor growth regulators of MFH through autocrine or paracrine pathways, when compared with BTC. PMID- 15274393 TI - Decrease in specific micronutrient intake in colorectal cancer patients with tumors presenting Ki-ras mutation. AB - BACKGROUND: The diversity of the Mediterranean diet and the heterogeneity of acquired genetic alterations in colorectal cancer (CRC) led us to examine the possible association between dietary factors and mutations, such as Ki-ras mutations, in genes implicated in the pathogenesis of these neoplasms. PATIENTS AND METHODS: The study was based on 246 cases and 296 controls. For the molecular study only 117 patients with Ki-ras tumor expression were included. Dietary patterns were assessed using a semi-quantitative food frequency questionnaire. RESULTS: Patients with Ki-ras mutations in codon 12 (K12) consumed significantly less vitamin A (p=0.02), B1 (p=0.01), D (p=0.02) and iron (p=0.03) than controls, whereas patients without these mutations had similar intakes of these nutrients to controls. The consumption of fiber, folate, vitamin E and potassium was lower in the two subgroups of patients (K12-positive or -negative) than in controls. Mutation in codon 13 was not associated with any nutrient deficit. CONCLUSION: These results support previous findings that certain micronutrients protect against colorectal neoplasia and emphasize the importance of considering the different molecular forms of CRC as etiologically distinct diseases. PMID- 15274394 TI - Single nucleotide polymorphisms in MMP1 and MMP3 gene promoters as risk factor in head and neck squamous cell carcinoma. AB - Matrix metalloproteinase (MMP) 1 and 3 genes play an important role in initiating tumor growth and promoting cell spread. Since MMP1 and MMP3 transcription levels can be modulated by promoter polymorphism, we investigated the impact of different genotypes on the occurrence of head and neck cancer in a Caucasian case control study. DNA was extracted from 126 male head and neck cancer patients and from 249 male hospitalized-based controls. Genotyping was carried out using PCR multiplex allowing the co-amplification of MMAP1-1607 bp and MMP3-1171 bp polymorphisms. PCR products were separated on a capillary electrophoresis. The MMP1-2G and the MMP3-6A allele frequencies were significantly lower in cases than in controls. In particular homozygous 2G/2G individuals were at lower risk of cancer than the 1G/1G carriers (OR= 0.37 95%CI [0.19-0.71], p=0.003). MMP1 and MMP3 polymorphisms were in moderate linkage disequilibrium in cases and controls (D'=0.41 and D'=0.46). Haplotype frequencies distribution derived from these 2 polymorphisms was significantly different between cases and controls (p=0.01). The haplotype analysis suggested an implication of both MMP1 and MMP3 polymorphisms in the head and neck squamous cell carcinoma susceptibility. Indeed, the presence of the MMP1-2G and MMP3-6A alleles seemed to be associated with decreased risk of head and neck squamous cell carcinoma but mainly when they were carried by the same haplotype. By comparison to the 1G-5A haplotype, the 2G 6A haplotype was associated with a lower risk of head and neck squamous cell carcinoma (OR=0.52 95%CI [0.34-0.80], p=0.003). PMID- 15274395 TI - Circulating soluble E-cadherin levels are of prognostic significance in patients with multiple myeloma. AB - BACKGROUND: The epithelial transmembrane molecule E-cadherin (E-Cad) is the prime mediator of epithelial cell-cell adhesion, through homotypic interactions. It also participates in the maintenance of cytoskeletal structure and cell-cell signalling, while there are no published reports of expression of E-Cad in non epithelial tissues. We examined whether the circulating levels of soluble E-Cad in newly diagnosed patients with multiple myeloma (MM) are of prognostic significance. PATIENTS AND METHODS: We used an ELISA method to determine the levels of circulating soluble E-cadherin (sE-Cad) in 21 newly diagnosed patients with MM and in 29 healthy volunteers, as a control group. RESULTS: MM patients demonstrated increased circulating levels of sE-Cad, compared with controls (p<0.0001). Increased circulating sE-Cad levels correlated with LDH levels at diagnosis (p<0.001) and poor prognosis. Multivariate analysis demonstrated that sE-Cad levels are an independent prognostic factor of survival (p<0.0207). CONCLUSION: Our data suggest that adhesion molecules play a role in the pathogenesis of MM, establish sE-Cad as an independent marker of survival and, finally, provide evidence of non-epithelial production of E-Cad in MM patients. PMID- 15274396 TI - Possible mechanism pertinent to mucosal invasion in sporadic colonic adenomas. AB - Colorectal adenomas are foci of dysplastic mucosa that may antedate the development of a colorectal cancer. In this work we investigated 62 colonic polyps. A close examination revealed that in some adenomatous glands facing the muscularis mucosa a group of dysplastic cells were missing. Those glandular defects were connoted as glandular pores. Many glands with pores were dilated and had retained mucin, inflammatory cells and/or necrotic material. Those products were often released through the pores into the surrounding lamina propria. Glandular pores were recorded in 25% (3/12) of the tubular adenomas, in 33% (2/6) of the serrated adenomas, in 50% (4/8) of the tubulo-villous adenomas and in 67% (14/21) of the villous adenomas. None of the 14 hyperplastic polyps had glandular pores. While cell locomotion is considered to be the most important parameter accountable for the local progression of tumors, the present results may offer an alternative view to the cell-migration theory (as the sole pathway of invasion). The release of proteolytic secretions through glandular pores in some colonic adenomas disrupt the surrounding matrix, a mechanism that would facilitate neoplastic cell penetration into the lamina propria. PMID- 15274397 TI - Efficacy of infusional biomodulated 5-fluorouracil in metastatic colorectal cancer after raltitrexed failure. AB - BACKGROUND: Treatment of metastatic colorectal cancer (CRC) with raltitrexed results in an objective response rate (OR) and overall survival (OS) comparable to bolus 5-fluorouracil (5-FU). A phase III trial found raltitrexed to be inferior to continuous infusion of 5-FU (ci5-FU). PATIENTS AND METHODS: This phase II trial studied the activity and toxicity of methotrexate-modulated ci5-FU (ci5-FU/MTX) after failure of raltitrexed in patients with metastatic CRC. RESULTS: Of 32 patients who received raltitrexed, 27 were evaluable for response. An OR was observed in 19%. Grade 3/4 toxicity occurred in 47% of patients. Eighteen patients received second-line ci5-FU/MTX. One complete response (CR) and five stable diseases (SD) were observed. CR and SD were observed in patients with raltitrexed-resistant disease. Toxicity to ci5-FU/MTX was mild and predictable, even in patients with severe toxicity under raltitrexed. CONCLUSION: ci5-FU/MTX is ineffective after failure of raltitrexed in patients with metastatic CRC. However, response to ci5-FU/MTX in patients with raltitrexed-resistant disease indicates incomplete cross-resistance. PMID- 15274398 TI - Pharmacokinetics of cisplatin in semi-closed hyperthermic peritoneal perfusion (HPP) for treatment of peritoneal carcinomatosis. AB - BACKGROUND: This study investigates the pharmacokinetics and toxicity of cisplatin, admininistered by a new semi-closed hyperthermic peritoneal perfusion (HPP) technique to patients with peritoneal carcinomatosis. MATERIALS AND METHODS: After surgical cytoreduction, 12 patients were given cisplatin 100 mg/m2 (CDDP), introduced into the HPP circuit for 60 min at 41.7 degrees C and 1200 ml/min flow rate. Perfusate and blood samples were obtained during/after perfusion, plus normal and tumor tissues samples before/after perfusion. RESULTS: Total and ultrafiltrate (UF) CDDP had similar patterns: monophasic in peritoneum, biphasic in plasma. At the end of perfusion, total/UF platinum (Pt) concentrations in the peritoneum decreased by 63.4%-64.9%. Total/UF Pt concentrations and AUCtot in perfusate were higher than plasmatic ones. Pt concentrations in tumor specimens were higher than in normal tissues. CONCLUSION: Cisplatin administered by semi-closed HPP evidenced pharmacological advantages: higher and direct drug exposure of the tumor within the peritoneal cavity, limited systemic absorption and mild toxicity. PMID- 15274399 TI - Alterations of the K-ras and p53 genes and microsatellite instability in sporadic colorectal carcinomas. AB - BACKGROUND: The international guidelines for the evaluation of microsatellite instability (MSI) in colorectal cancer were defined in 1997 by the National Cancer Institute (NCI). Here, the relationship between MSI, cancer-associated genes and their clinicopathological variables were revaluated using these guidelines. PATIENTS AND METHODS: Mutations of K-ras at exon 1 and p53 at exons 5, 6, 7 and 8 were analyzed in 43 cases of sporadic colorectal carcinoma. MSI was analyzed using the 5 markers recommended by the NCI reference panel. RESULTS: The proportion of p53 mutations in the MSI-H cases (0 out of 5; 0%) was lower than that of non-MSI-H cases (23 out of 38; 60.5%) (p=0.0117). The proportion of p53 mutations in microsatellite stable (MSS) cases (21 out of 34; 61.8%) was higher than that of non-MSS cases (2 out of 9; 22.2%) (p=0.0366). The proportion of K ras mutations in MSI-H tumors (1 out of 5; 20.0%) was less frequent than in non MSI-H tumors (19 out of 38; 50.0%) (p=0.2115). CONCLUSION: p53 mutations in MSI-H tumors were much less common than in non-MSI-H tumors. This result suggested that alterations of the p53 gene are not closely associated with carcinogenesis in MSI H carcinomas. PMID- 15274400 TI - Thrombocytosis in gynecologic malignancies. AB - Recent studies have addressed the prevalence and prognostic impact of thrombocytosis in various gynecologic and non-gynecologic malignancies. Thrombocytosis appears to be of prognostic value in certain patients with gynecologic malignancies. In this survey we review the published data and attempt to analyze the prognostic implications of thrombocytosis in patients with gynecologic malignancies. PMID- 15274401 TI - Tumor histology and stage but not p53, Her2-neu or cathepsin-D expression are independent prognostic factors in breast cancer patients. AB - BACKGROUND: Several factors are currently employed for prognosis assessment and treatment determination in breast cancer. An array of molecular parameters, such as p53, Her2-neu (c-erbB 2) and Cathepsin-D, are also examined to improve clinical patient management. We have conducted a statistically powerful study of the prognostic value of conventional factors and of the investigational factors p53, Her2-neu and Cathepsin-D in patients with invasive breast carcinoma, in order to compare their significance. Our analysis was extended to determine the associations of p53 and Her2-neu with risk of death and relapse among patients with and without lymph node metastases. MATERIALS AND METHODS: In a set of 125 primary breast tumors, p53 and Her2-neu expression were immunohistochemically evaluated. Cathepsin-D, estrogen and progesterone receptor concentrations were determined in cytosols by a standard immunoradiometric assay. RESULTS: Over a mean of 62 months, 49 patients (39%) had a relapse and 29 patients (23%) died. Overexpression of p53, Her2-neu and Cathepsin-D was observed in 31%, 46% and 88% of cases, respectively. Overall survival was associated with histology (hazard ratio 0.04, 95% confidence interval: 0.01, 0.49 for lobular tumors) and stage (hazard ratio 5.94, 95% confidence interval: 1.30, 27.15 for stage III samples). Disease-free survival was also related to histology (hazard ratio 0.23, 95% confidence interval: 0.08, 0.73 for lobular tumors) and stage (hazard ratio 4.27, 95% confidence interval: 1.36, 13.36 for stage III tumors). Patients with both negative nodal status and Her2-neu overexpression tended to display an elevated risk of death. CONCLUSION: Our results support the prognostic power of tumor histology and stage and emphasize the need for further studies on the prognostic impact of p53. Her2-neu and Cathepsin-D in breast cancer. Additionally, our analysis indicates that deregulation of Her2-neu might characterize a subgroup of node-negative patients with poor prognosis who could benefit from an aggressive adjuvant therapy. PMID- 15274402 TI - Demographic and clinical factors as determinants of serum levels of prostate specific antigen and its derivatives. AB - BACKGROUND: To characterise the association between demographic and clinical factors and levels of total prostate specific antigen (tPSA) and its molecular derivatives complexed PSA (cPSA), free PSA (fPSA) and the ratio of free to total PSA (%fPSA)] in New Zealand Maori, Pacific Islanders and Europeans, in order to determine whether reported ethnic differences in PSA can be explained by lifestyle and social factors. MATERIALS AND METHODS: Demographic and clinical factors were examined in relation to tPSA, fPSA and cPSA levels, in 1405 Maori, Pacific Island and New Zealand European men with no clinical evidence of prostate cancer, in the Wellington region of New Zealand. Any associations between levels of PSA and PSA derivatives and body mass index, smoking status, family cancer history, non-steroidal anti-inflammatory/vitamin supplement usage, number of sexual partners, age at first intercourse, previous vasectomy, marital/partnership status, educational level and socioeconomic status were investigated by backwards stepwise regression analysis, correcting for age, ethnicity and urinary symptoms. RESULTS: Not being married/partnered was associated with increased tPSA, fPSA and cPSA. tPSA and cPSA decreased with regular non-steroidal anti-inflammatory use. cPSA was decreased in subjects with a first degree relative with any form of cancer. tPSA and fPSA were decreased if the body mass index was > 34. fPSA and %fPSA were decreased in current and former smokers. CONCLUSION: Demographic and clinical factors appear to have a significant effect on levels of PSA and its various derivatives and may account for previously observed ethnic differences. It is important that these associations are taken into account when comparing individual PSA results with standard reference ranges. PMID- 15274403 TI - Prognostic value of pelvic lymphadenectomy in surgical treatment of apparent stage I endometrial cancer. AB - BACKGROUND: The role of pelvic lymphadenectomy in early endometrial carcinoma is still being debated. MATERIALS AND METHODS: We retrospectively analyzed a total of 131 patients with FIGO stage I endometrial cancer undergoing surgery without (Group 1) or with (Group 2) pelvic lymphadenectomy. Kaplan-Meier and Cox analyses were used to calculate crude and adjusted survival rates. Moreover, the overlap of pre- and post-surgical staging was analyzed. RESULTS: Overall survival rate at 5 years was 90.1%. The difference in crude survival rates of the two groups is not statistically significant (p-value= 0.3777, log rank test). Five patients of Group 2 presented positive pelvic nodes. Therefore our results showed a pre surgical understaging, referring to nodal involvement, in 9.1% of cases (5/55). CONCLUSION: Pelvic lymphadenectomy is a useful procedure for prognostic and staging purposes, but does not improve survival in FIGO stage I endometrial carcinoma. PMID- 15274404 TI - Cardiotoxicity in advanced non-small cell lung cancer patients treated with platinum and non-platinum based combinations as first-line treatment. AB - BACKGROUND: One of the major dose-limiting toxicities of anthracyclines is cardiotoxicity due to irreversible cardiomyopathy. Whether cisplatin-based treatment induces caridiotoxicity in the short term, especially in non-small cell lung cancer (NSCLC) patients with cardiovascular comorbidity, has not been studied previously. The aim of this study was to evaluate cardiotoxicity in advanced NSCLC patients receiving cisplatin-gemcitabine (CG) or epirubicin gemcitabine (EG) as first-line treatment. PATIENTS AND METHODS: Patients were randomised to receive gemcitabine 1125 mg/m2 (days 1 and 8) plus either cisplatin 80 mg/m2 (day 2) or epirubicin 100 mg/m2 (day 1) every 3 weeks for a maximum of 5 cycles. Patients had to have a left ventricular ejection fraction (LVEF) > 45%, measured by multiple gated acquisition (MUGA) scan. A second MUGA scan was performed 12 weeks after the end of treatment. RESULTS: Sixty-nine patients were included. The mean total dose of cisplatin was 349 mg/m2 and of epirubicin 452 mg/m2. The mean difference in decline in LVEF from baseline was 2% in the CG arm versus 6% in the EG arm (p=0.016). Clinically evident cardiac failure was not observed during 12 months follow-up. No correlation was found with total drug doses administered. In patients with a history of cardiac disease a trend towards a higher decrease in LVEF was observed. CONCLUSION: Although in the EG arm the LVEF significantly declined and in the CG arm a trend for LVEF to decline was observed, the risk of cardiac failure is limited in advanced NSCLC patients. PMID- 15274405 TI - Combination chemotherapy with docetaxel and doxifluridine showed a beneficial outcome in advanced or recurrent breast cancer patients with longer disease-free interval. AB - Fundamental studies have confirmed that combination chemotherapy with docetaxel and doxifluridine (a capecitabine metabolite) is very useful in the treatment of breast cancer. This study investigated the usefulness and tolerability of a combination chemotherapy consisting of docetaxel administration on day 8 of doxifluridine therapy in 40 advanced/recurrent breast cancer patients. The overall response rate was 41.0% in 39 eligible patients. The median time to progression (TTP) for all patients was 295 days. Many responders had lung metastasis, soft tissue metastasis or a good performance status, whereas the clinical response showed no correlations with the estrogen receptor status or prior treatment with an anthracycline. The most common hematological toxicities were leukopenia and neutropenia, but dose reduction or delay of administration of either drug was unnecessary. CONCLUSION: The good response rate and long TTP of this doxifluridine plus docetaxel regimen indicate its potential as a first- or second-line treatment for advanced/recurrent breast cancer patients. PMID- 15274406 TI - Oxaliplatin hepatic arterial infusion chemotherapy for hepatic metastases from colorectal cancer: a phase I-II clinical study. AB - Oxaliplatin is a new drug active in the treatment of advanced colorectal cancer. Hepatic arterial infusion chemotherapy is under evaluation because of the high target dose and low general toxicity. Twelve patients with liver metastases from colorectal cancer were enrolled, all pretreated with evidence of progressive disease: three after a partial remission induced by oxaliplatin, folinic acid and 5-FU, three patients after a partial remission induced by irinotecan, folinic acid and 5-FU and six patients after failing a 5-FU and folinic acid regimen. They received hepatic arterial infusion chemotherapy with oxaliplatin as 30-min infusion on an outpatient basis every 3 weeks. Dose-limiting toxicity was observed at 175 mg/m2/cycle and consisted of obliteration of the hepatic artery in one patient, abdominal pain requiring morphine in one patient and severe hypotension requiring plasma expander in a third. Following phase 1, all patients received 150 mg/m2 for six cycles. We reported four cases of partial remission (33%) lasting 24, 15, 12 and 10+ weeks, respectively, 2 stabilisation of disease (17%) lasting more than 12 weeks and six progressions (50%). Six patients (50%) presented CEA reduction of > 30% and five patients (41%) showed an increase of > 8% of body weight. The median survival was 13 months (range 6-19). Oxaliplatin did not present significant toxicity for liver parenchyma and biliary tree. We advise that further studies be undertaken with oxaliplatin 150 mg/m2. PMID- 15274407 TI - Ten-year disease-free survival of a small cell lung cancer patient with brain metastasis treated with chemoradiotherapy. AB - We report the first case of 10-year disease-free survival in a patient with small cell lung cancer (SCLC) with brain metastasis. A 63-year-old man was found to have SCLC with brain metastasis and underwent chemoradiotherapy. Radiation therapy was delivered to the brain, lungs, mediastinum and supraclavicular fossa. Chemotherapy regimen mainly consisted of etoposide-plus-cisplatin. The patient has remained alive for more than 10 years after the diagnosis of SCLC with brain metastasis with no relapses. PMID- 15274408 TI - Prognostic impact of tissue inhibitor of matrix metalloproteinase-1 in plasma of patients with colorectal cancer. AB - BACKGROUND: Tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) in plasma has been reported to be related to disease progression in patients with colorectal cancer. However, the prognostic significance of plasma TIMP-1 has not been clarified. PATIENTS AND METHODS: Concentrations of TIMP-1 protein were measured by enzyme-linked immunosorbent assay in plasma samples of 87 preoperative patients who subsequently underwent resection, and prognosis was compared. The cut-off value of plasma TIMP-1 was defined as 170 ng/ml. RESULTS: When clinicopathological factors between patients with positive and those with negative plasma TIMP-1 were analyzed, significant differences were observed in lymph node metastasis, serosal invasion, curability and Dukes' classification. Univariate analysis of these factors demonstrated that depth of invasion, metastases to lymph nodes, peritoneum, liver and distant organ, lymphatic and vessel invasions, curability. Dukes' classification and plasma TIMP-1 concentration were significant. By multivariate analysis excluding patients with distant or peritoneal metastases, histological type, lymphatic invasions, lymph node metastasis and plasma TIMP-1 were retained in the final model. CONCLUSION: These results suggested that plasma TIMP-1 may be a useful prognostic marker for survival in patients with colorectal carcinoma. PMID- 15274409 TI - Gemcitabine and continuous infusion of 5-fluorouracil in locally advanced and metastatic pancreatic cancer: a phase I-II study. AB - BACKGROUND: Gemcitabine has been recently recognized as standard treatment in advanced pancreatic cancer. To potentiate its single-agent activity we conducted a phase I-II study with the primary objective of establishing the maximum tolererated dose (MTD) of gemcitabine and continuous infusion 5-FU in patients with locally advanced or metastatic pancreatic cancer. PATIENTS AND METHODS: Fifteen patients received a fired dose of 5-FU 200 mg/mq protracted infusion for six months. Gemcitabine was administered weekly for three out of four weeks for six cycles at escalating doses of 800 mg/mq to 1100 mg/mq. RESULTS: MTD was established at 1000 mg/mq of gem citabine. Of the 11 evaluable patients, 7 patients had stable disease, 1 had partial response and 3 had progressive disease. Of the 14 patients evaluable at follow-up, median time to progression was 5 months. Median survival was 10 months. CONCLUSION: This study confirms the good tolerability of the combination, of gemcitabine with 5-FU. PMID- 15274410 TI - Serrated adenomas of the cardia. AB - Adenomas of the stomach are histologically classified into tubular and villous adenomas, mixed (tubulovillous) and papillary phenotypes being also recognized. They are localized in the antrum and the body. In a previous work we reported a case of serrated adenoma of the cardia having, at the time of diagnosis, an invasive growth. In this communication we report five newly identified cases of serrated adenomas of the cardia. The five neoplasias were characterized by serrated indentations furnished with dysplastic epithelium. Five out of six serrated adenomas--including the previously reported case--had, in addition, an invasive or a suspected invasive carcinoma. The latter case had at the time of diagnosis lung and liver metastasis. Metastases were also demonstrated in two additional cases. In similarity with other adenoma phenotypes, serrated adenomas of the cardia have a propensity to evolve into invasive carcinoma, a phenomenon that appears to be size-dependent. PMID- 15274411 TI - Instruments for quality of life assessment in patients with gastrointestinal cancer. AB - Health-related quality of life (HRQoL) is a multi-dimensional concept, encompassing all aspects of patient health and used widely as an outcome measure in clinical trials. In this review, the current status of HRQoL assessment in clinical studies of gastrointestinal cancer is examined and the various instruments proposed for this purpose are considered and compared. The cancer specific questionnaires, among them the Spitzer Quality of Life Index, the Rotterdam Symptom Check List, the Functional Living Index-Cancer, the Functional Assessment of Cancer Therapy General (FACT-G) and the Quality of Life Questionnaire of the European Organization for Research and Treatment in Cancer (EORTC), provide essential information about particular concerns of cancer patients and are most sensitive in detecting changes over time. The domain specific questionnaires, among them the Multidimensional Fatigue Inventory, the McGill Pain Questionnaire, the Hospital Anxiety and Depression Scale and the Anorectal Sphincter-Conservative Treatment Questionnaire, are designed to assess one specific domain of quality of life. The core-module cancer-specific questionnaires combine a core questionnaire for use in any type of cancer with a module questionnaire which assesses specific issues in cancer patient subgroups. Such core-module instruments have been evaluated for colorectal, pancreatic, hepatobiliary, oesophageal and gastric cancer. The most valid and standardized instruments for HRQoL assessment in cancer patients are the EORTC and the FACT questionnaires, which are widely used in Europe and around the world. Data provided by these specific instruments complement clinical outcomes and may help to evaluate the costs and benefits of different treatment options, thus being essential to further improvement of treatment and care of cancer patients. PMID- 15274412 TI - CD3 zeta expression of regional lymph node and peripheral blood lymphocytes in gastric cancer. AB - BACKGROUND AND AIM: Impaired expression of the CD3 zeta chain in T cells associated with T cell anergy has been reported in cancer patients. However, few studies have investigated CD3 zeta expression in regional lymph node lymphocytes (LAL) in cancer patients. This study aims to confirm CD3 zeta expression levels by lymph node and peripheral blood lymphocytes (PBL) in gastric cancer patients and to discuss the clinical implications of intranodal or peripheral blood expression of CD3 zeta in gastric cancer. PATIENTS AND METHODS: Twenty-two gastric cancer patients were enrolled. Macroscopically non-metastatic compartment 1 LNL (C1LNL), compartment 2 LNL (C2LNL) and PBL were surgically obtained. Two color flow cytometry was then used to quantify the levels of CD3 zeta expression in C1LNL, C2LNL and PBL. RESULTS: Impaired CD3 zeta expression was confirmed in 9.5% of C1LNL, 8.9% of C2LNL and 4.2% of PBL. There was a significant difference in CD3 zeta expression levels between C1LNL and PBL (p<0.01). CD3 zeta expression in PBL was significantly correlated with depth of invasion but not nodal involvement. Distinct differences between the respective lymph node compartments were not identified. CONCLUSION: Immunological paralysis following CD3 zeta impairment may occur more frequently in LNL than in PBL in gastric cancer. Identifying such patients during the perioperative period using flow cytometric methods will increase the efficacy of cytokine therapy aiming to normalize CD3 zeta expression levels. PMID- 15274413 TI - Primitive neuroectodermal tumor of the head and neck: incidence, diagnosis, and management. AB - Primitive neuroectodermal tumors are in the Ewing's sarcoma family of tumors and are composed of small round cells. Because of their rare occurrence, optimal therapy is challenging, particularly if they occur in the head and neck. Diagnosis is based on history, immunostaining with at least 2 neural markers, ultrastructural examination, and evidence of an abnormal t(11;22)(q24;q12) translocation as the hallmark for the Ewing's sarcoma family. The prognosis in general is poor because of overt metastasis at the time of diagnosis. Of 27 reported patients with primitive neuroectodermal tumors of the head and neck, 23 were less than 20 years of age. Most patients presented with a tumor in the nasal cavity, paranasal sinuses, or neck. Symptoms developed rapidly (3.6 months, on average), and a lethal outcome occurred in 9 patients. This highly malignant tumor requires an aggressive combination of radical resection, chemotherapy, and radiotherapy. A close follow-up with regular radiographic examination for at least 5 years is mandatory. PMID- 15274414 TI - Budesonide-dependent modulation of expression of macrophage migration inhibitory factor in a polyposis model: evidence for differential regulation in surface and glandular epithelia. AB - Macrophage migration inhibitory factor (MIF) is a counterregulatory lymphokine for glucocorticoid action within the immune system. To provide further insights into the way expression of pleiotropically acting MIF is modulated by glucocorticoids, we investigated the influence of the glucocorticoid budesonide on the level of expression of MIF in a model of human nasal polyposis by quantitative immunohistochemical analysis. Ten nasal polyps obtained from surgical resection were maintained for 24 hours in the presence of 3 budesonide concentrations: 10, 50, and 250 ng/mL. As quantitatively demonstrated by computer assisted microscopy, 50 ng/mL induced an increase in MIF expression in the surface epithelium and a decrease in MIF expression in the glandular epithelium. At the 250 ng/mL dose, the inverse effect was induced. Evidently, surface and glandular epithelia react nonuniformly to the glucocorticoid regarding MIF presence, adding dependence on the cell type to the regulatory network. PMID- 15274415 TI - Immunohistochemical localization of histamine receptor subtypes in human inferior turbinates. AB - Histamine is an important chemical mediator in allergic rhinitis and plays an important role in eliciting the nasal symptoms of the disorder. However, the immunohistochemical localization of histamine receptor subtypes (H1R, H2R, H3R, and H4R) in human nasal mucosa is unknown. There are also no prior studies of H3R and H4R in human nasal mucosa. The objective of this study was to examine the distribution of histamine receptor subtypes in the human inferior turbinates by an immunohistochemical method. H1R was localized primarily in the epithelium, vessels, and nerves. H2R was localized primarily in the epithelium and the glands. H3R and H4R were clearly distributed on the nerves. In addition, H1R, H3R, and H4R were clearly localized on the same nerves. This result indicates that H1R, H3R, and H4R adjoin and regulate each other in the same nerves. All histamine receptor subtypes may play some role in patients with allergic rhinitis. PMID- 15274416 TI - Fasciitis ossificans of the paranasal sinus. AB - We report a case of fasciitis ossificans of the nasal cavity and paranasal sinus in an infant who was surgically treated in our department. Fasciitis ossificans is a rare benign reactive lesion and a variant of the more commonly known entity nodular fasciitis. We present the radiographic appearance and discuss the surgical resection, which was performed by a paranasal section, as well as the histologic and immunohistochemical results. To the best of our knowledge, no other case of fasciitis ossificans of the paranasal sinus has yet been reported. It is important to publish cases such as this, because their recognition as benign entities can prevent aggressive surgical procedures. PMID- 15274417 TI - Diagnosis and therapeutic management of iatrogenic parotid sialocele. AB - Salivary gland sialoceles are relatively common and may be a complication of trauma with a penetrating salivary gland injury or may be a complication of salivary gland surgery. The development of new diagnostic tools such as magnetic resonance sialography and endoscopic techniques has led to further improvements in the clinical and diagnostic assessment of this condition, and botulinum toxin therapy has recently been described in the management of parotid sialoceles. We here report the case of a 41-year-old patient with an unusually complicated parotid sialocele following an unsuccessful attempt to remove a stone located in the distal third of Stensen's duct. Magnetic resonance sialography and sialoendoscopy were used in order to obtain an adequate diagnostic assessment. The patient underwent extracorporeal lithotripsy that led to partial symptom regression. After the development of a parotid abscess, he received antibiotics and a botulinum toxin type A injection that induced spontaneous drainage and disappearance of the symptoms. Magnetic resonance sialography and sialoendoscopy are promising new diagnostic techniques for better noninvasive management of iatrogenic sialoceles. PMID- 15274418 TI - Use of a "hands-free" tracheostoma valve in patients with laryngectomy and tracheoesophageal puncture. AB - The goals of the study were 1) to investigate the relationship between tracheal air pressure and the tracheostoma valve seal in patients with laryngectomy and tracheoesophageal puncture (TEP) and 2) to assess whether tracheal pressure contributes to the duration of the valve seal. Ten patients with TEP after laryngectomy underwent training with an outer Blom-Singer tracheostoma valve and a 30-day trial with this "hands-free" speaking valve. We found that tracheal pressure was not a significant predictor variable in assessing the duration of the outer tracheostoma valve seal achieved by the patients with TEP. Factors other than tracheal pressure appeared to affect the duration of the tracheostoma valve seal. Mastery of use of the tracheostoma valve, the subject's neck anatomy, and the amount of time, training, and experience that was given to each subject were significant predictors of the valve seal. Tracheal pressure measurements might be of use in deciding between a standard or a low-pressure outer tracheostoma valve for an individual patient. PMID- 15274419 TI - Differential changes in peripheral and central components of the brain stem auditory evoked potentials during the neonatal period in term infants after perinatal hypoxia-ischemia. AB - To identify any differences in dynamic changes between peripheral and central hearing after perinatal hypoxia-ischemia, we studied 80 term infants during the neonatal period by serially recording brain stem auditory evoked potentials (BAEPs) at 60 dB normal hearing level. All BAEP wave latencies and the I-V interval increased significantly on day I (analysis of variance, all p < .001). Thereafter, the wave I latency decreased gradually with some variation. The wave V latency and the I-V interval increased further on day 3 and then decreased progressively. On day 30, neither the latencies nor the I-V interval differed significantly from those of normal controls, but the wave V latency and the I-V interval still tended to increase slightly. These results suggest that hearing is impaired shortly after hypoxia-ischemia. Peripheral hearing gradually recovers after day 1, whereas central impairment progresses during the first 3 days and then starts to recover. We conclude that peripheral impairment recovers sooner than central impairment after perinatal hypoxia-ischemia. PMID- 15274420 TI - Clinical and experimental investigations of spontaneous impedance changes of the middle ear. AB - The major symptoms of glomus tympanicum tumors are pulsatile tinnitus and spontaneous impedance changes (SICs) of the middle ear. On the other hand, SICs often occur even in the absence of pathological findings. The aim of this study was to analyze the occurrence of SICs in patients and healthy volunteers. We retrospectively evaluated 184 patients with SICs and/or complaints of periodic tinnitus. Most of them (n = 134) showed pulse-synchronous SICs. Pathological findings were recorded in only 66 patients. Binaural SICs were registered significantly (p = .03) more frequently in patients with arterial hypertension (63% versus 18%). Because of the positive correlation between arterial hypertension and the occurrence of SICs, the influence of increasing blood pressure (systolic blood pressure > 160 mm Hg after physical activity) on the occurrence of SICs was investigated in a prospective trial in healthy test subjects (n = 42). In 17 of them, pulse-synchronous SICs occurred for the first time or were registered at a lower sensitivity level after an increase in blood pressure. In summary, only half of the patients with pulse-synchronous SICs showed pathological findings. A significant correlation between high blood pressure and binaural pulse-synchronous SICs was demonstrated in patients with arterial hypertension and healthy volunteers after physical activity. PMID- 15274421 TI - Acoustic neuroma ingrowth in the cochlear nerve: does it influence the clinical presentation? AB - We examined the clinical presentation in patients with a histologically proven ingrowth of the cochlear nerve by acoustic neuroma to see whether this differs from what is known from large acoustic neuroma series. In total, 85 acoustic neuromas had an en bloc dissection to study histologically the relation between the cochlear nerve and the acoustic neuroma. In 21 of these 85 specimens, there was histologic proof of invasion of the cochlear nerve by the tumor. For 13 of these 21 tumors, sufficient clinical data could be retrieved to describe the clinical presentation in these patients. We collected clinical data such as age, sex, presenting symptoms, duration of symptoms, tone audiograms, tumor size measurements and volumetric calculations, and latency interval data I-V of brain stem evoked response audiometry and calculated whether there was any correlation among those data. We also compared these clinical data with the data from some large acoustic neuroma series. No clear difference could be shown between the clinical presentation of acoustic neuroma patients with cochlear nerve ingrowth and the clinical presentations in large acoustic neuroma series. This outcome favors the theory that the hearing impairment in acoustic neuroma patients is mainly the result of compression on the vessels of the cochlea and/or on the cochlear nerve. PMID- 15274422 TI - Longitudinal phenotypic analysis in patients with connexin 26 (GJB2) (DFNB1) and connexin 30 (GJB6) mutations. AB - In 15 Belgian subjects with prelingual sensorineural hearing impairment, the connexin 26 (GJB2) gene and the connexin 30 (GJB6) gene were analyzed for the presence of the 35delG mutation and the delta(GJB6-D13S1830) deletion first described by del Castillo et al in 2002. Seven patients were found to be homozygous for the 35delG mutation; 7 were combined heterozygotes for the 35delG mutation and the GJB6 deletion. In 11 subjects, phenotype and genotype were correlated. Significant, transient progression, in the range of 1.7 to 2.7 dB/y, was only found in 2 patients in the first part of the second decade of life. Hearing impairment was otherwise stable, with mean thresholds of 75, 90, and 100 dB at 0.125, 0.25, and 0.5 kHz, respectively, and 100 dB or higher at 1 to 4 kHz. There was no significant difference in hearing impairment between the patients with the homozygous 35delG mutation in GJB2 and those who are heterozygous for both the 35delG mutation and the deletion encompassing part of GJB6. PMID- 15274423 TI - Combined laryngocele. PMID- 15274424 TI - Comparison of current perception threshold between each side in unilateral complex regional pain syndrome patients does not measure the patient's pain. AB - The current perception threshold (CPT) test has been developed as one of the neuroselective sensory nerve conduction threshold tests. The score of the CPT of the affected side subtracted from the score of the CPT of the unaffected side in complex regional pain syndrome (CRPS) is expected to show pain objectively. The purpose of this study is to examine first whether the CPT of the affected side is generally lower than that of the unaffected side, and second, whether the greater score shows the more intense pain. The CPT of each side in 25 patients with unilateral CRPS type I was measured and compared. For the 2000 Hz stimulus, the CPT of the affected side was 2677 +/- 262 microAmp (mean +/- standard error) and the CPT of the unaffected side was 2194 +/- 247 microAmp (p = 0.0149). For the 250 Hz stimulus, the CPT was 876 +/- 117 microAmp and 721 +/- 73 microAmp respectively (p > 0.05). For the 5 Hz stimulus, the CPT was 730 +/- 105 microAmp and 448 +/- 56 microAmp respectively (p = 0.0018). In 2000 Hz, 250 Hz, and 5 Hz stimuli, the CPT of the affected side was higher than that of the unaffected side. This shows that generally the affected side is less sensitive than the unaffected side in terms of current perception. The score of the CPT of the affected side subtracted from the score of the CPT of the unaffected side in CRPS does not measure the patient's pain. PMID- 15274425 TI - Studies on eleven kidney transplants from non-heart-beating donors. AB - This study was performed to analyze postoperative courses and complications, retrospectively, following transplants from non-heart-beating donors and to examine the correlation between early graft function and clinical parameters. We experienced 11 cases of kidney transplants from non-heart-beating donors during the period from April 1995 to May 2003. Warm ischemic time was less than 30 min in all cases, and total ischemic time ranged from 8.4 hours to 27.9 hours. Rejection reactions occurred in seven cases, two of which were vascular rejections. Infectious disease complications included CMV in two cases, interstitial pneumonia in one case and fungal infection in one case. One patient died from interstitial pneumonia, and three patients had to be restarted on dialysis due to loss of function of the grafted kidney. The remaining seven patients all made full recoveries. All of the 16 patients who underwent living related kidney transplantations during the same period made full recoveries. Both the donor's gender and the latest creatinine level of the donor influenced the posttransplant dialysis period. The posttransplant dialysis period significantly influenced the creatinine level one month after transplant. These results suggest that patients who undergo kidney transplants from non-heart-beating donors have higher rates of complications than patients who undergo living related kidney transplantation. It is important that, in cases where the donor's creatinine level is high, especially when the donor is male, the kidney is carefully retrieved and transported to the recipent hospital to shorten the ischemic period as much as possible. PMID- 15274426 TI - Comparative benefit of preemptively applied thiopental for propofol injection pain: the advantage over lidocaine. AB - Propofol is one of the most frequently applied intravenous anesthetics for the induction of general anesthesia. However, pain on injection of this agent is a considerable problem in daily anesthesia practice because of its severity. Administration of lidocaine prior to propofol injection is a standard technique for reducing the pain on injection. However, this method provides insufficient pain relief. To evaluate whether pretreatment with an ultra-short acting barbiturate, thiopental, is more effective than with lidocaine, a randomized and single-blinded trial was conducted. Patients (20-65 years old, n = 137) were allocated into six groups, and applied with physiological saline, thiopental (25, 50, 75, or 100 mg), or lidocaine (40 mg) at 30 second prior to propofol injection (1 mg/kg, 1200 ml/h). The patient was interviewed about the degree of pain just after propofol was totally injected. Both thiopental (> or =25 mg) and lidocaine decreased the severity of pain in comparison with physiological saline as evaluated by a six-graded pain score. Lidocaine failed to influence the incidence of pain (from 86% to 55%), although thiopental significantly decreased it to 40% (25 mg), 21% (50 mg), 12% (50 mg), and 0% (100 mg), respectively. Thiopental (> or =50 mg) decreased both the severity and incidence of pain more effectively than lidocaine. A Hill plot analysis of these data, after rearrangement by patient's body weight, estimated that the half-effective dose (ED50) and the ED99 of this drug to block pain on injection of propofol were 0.6 and 1.4 mg/kg, respectively. PMID- 15274427 TI - Pharmacodynamic study of polyethylene glycol conjugated bovine hemoglobin (PEG bHb) in rats. AB - The purpose of this study was to determine the pharmacodynamics of polyethylene glycol (PEG) conjugation on stroma-free bovine hemoglobin, hemorrhage shock, and exchange transfusion rat models were used. In both rat models, blood pressure increased in 6% PEG-bHb treated animals was significantly higher than dextran 70, even than whole blood (isovolume of hemorrhage) transfusion. Six percent PEG-BHb could ameliorate the micro-circulation of the experimental animals markedly, including reducing the blood viscosity and improving the blood flow. This effect of PEG-bHb was superior to autogenous blood which only recovered 50% blood flow, and there's no effect on blood flow when used isovolumic dextran 70. Tissue oxygenations of rats were evaluated by the oxygen dependent quenching of phosphorescence using an Oxyspot phosphorimeter, and the results showed that capability of oxygen-delivery of PEG-bHb was close to autogenous blood and superior to dextran 70. Based on these effects, the survival rates of animals treated with PEG-bHb were close to that of the whole blood transfusion. And data suggested that half of the hemorrhage transfusion is the reasonable therapeutic dosage. In conclusion, PEG-bHb is an effective blood substitute with powerful tissue oxygenation and blood volume expansion. PMID- 15274428 TI - Blood substitute resuscitation as a treatment modality for moderate hypovolemia. AB - Blood substitute resuscitation as a treatment modality for moderate hypovolemia (approximately 40% blood loss) in a canine model has been evaluated using Oxyglobin (Biopure Hemoglobin Glutamer-200/ Bovine; a hemoglobin-based oxygen carrier) and Hespan (6% hetastarch; a nonoxygen-carrier) as resuscitants. Autologous (shed) blood served as control. Nine dogs were studied--after splenectomy, each dog was hemorrhaged (32-36 mL/kg; MAP = approximately 50 mmHg) and randomly assigned to the three resuscitation groups. Microvascular, systemic function and oxygenation characteristics were monitored and/or measured simultaneously in prehemorrhagic (baseline), posthemorrhagic and postresuscitation phases for correlation-real-time microvascular changes in the bulbar conjunctiva were noninvasively measured via computer-assisted intravital microscopy and systemic function and oxygenation changes were monitored and/or measured via instrumentation and devices incorporated into our bioengineering station in an operating room setting. Blood chemistry was also studied for relevant measurements. Prehemorrhagic microvascular characteristics were similar in all animals (venular diameter = 41 +/- 12 microm, A:V ratio = approximately 1:2, red-cell velocity = 0.5 +/- 0.3 mm/s). All animals also showed similar prehemorrhagic systemic function and oxygenation measurements comparable to a previous study and were consistent with normal measurements in dogs. At the completion of hemorrhaging to achieve moderate hypovolemia (approximately 40% blood loss with MAP at approximately 50 mmHg), all nine animals showed similar significant (P < 0.01) posthemorrhagic microvascular changes, including approximately 17% decrease in diameter (34 +/- 7 microm), A:V ratio = variable, and approximately 80% increase in velocity (0.9 +/- 0.5 mm/s). All animals also showed similar significant (P < 0.01) posthemorrhagic systemic function and oxygenation changes, with decreases in Hct, aHb(total), MPAP, MAP, SAP, DAP, CO, SVI, CaO2, and CvO2 and increases in HR and lactic acidosis. Shed blood (control) resuscitation restored posthemorrhagic microvascular changes close to prehemorrhagic values (diameter = 39 +/- 6 microm, A:V ratio = approximately 1:2, velocity = 0.6 +/- 0.4 mm/s). Oxyglobin and Hespan restored microvascular changes in similar manner close to prehemorrhagic values (Oxyglobin: diameter = 38 +/- 3 microm, A:V ratio = approximately 1:2, velocity = 0.6 +/- 0.4 mm/s; Hespan: diameter = 38 +/- 7 microm, A:V ratio = 1:2, velocity = 0.5 +/- 0.4 mm/s). After resuscitation, shed blood (control) restored all systemic function and oxygenation changes close to prehemorrhagic values. However, both Oxyglobin and Hespan resuscitation restored systemic function changes, but not oxygenation changes, to prehemorrhagic values. This was an interesting finding because of the different oxygen-carrying capability of Oxyglobin (oxygen-carrying) and Hespan (nonoxygen-carrying). The result suggests that either volume replenishment alone (and not oxygen-carrying capability) is needed to treat moderate hypovolemia or oxygenation measurements obtained by standard methods (oximetry, blood chemistry) may not reflect tissue oxygenation levels. PMID- 15274429 TI - Purification of hemoglobin by ion exchange chromatography in flow-through mode with PEG as an escort. AB - Development of hemoglobin-based blood substitutes requires production of highly purified hemoglobin. Process of hemoglobin purification by ion exchange chromatography in flow-through mode was researched and optimized. Three kinds of media including, QMA Spherosil LS (Biosepra, France) and Q Sepharose Big Beads (Amersham Bioscience, Sweden), and an anion exchange membrane column, Mustang Q (PALL, USA) were investigated and compared. Adding polyethylene glycol (PEG) as an escort in ion exchange chromatography improved the purity and recovery, and the recovery in the chromatography was increased from 75 to 95%. The mechanism of PEG effects on chromatography was discussed. The optimal chromatography step, in combination with hypotonic dilution hemolyzing and membrane separation, formed an integrated hemoglobin purification process. The total recovery in the process was 87.6%. The activity of hemoglobin was well preserved: P50 23.2 mmHg, and Hill coefficient 2.31. The product appeared as a single band in SDS-PAGE, and GF-HPLC showed only one peak. The purity of the prepared hemoglobin was more than 99.9%. The optimized process is time saving and suitable for large-scale preparation of hemoglobin to provide materials for further preparation of blood substitutes. PMID- 15274430 TI - Study of bovine hemoglobin dissociation by multiangle laser light-scattering method. AB - Hemoglobin (Hb) is an oligomeric protein, composed of four monomeric subunits. Hb molecule may undergo dissociation from a single native tetramer to two dimmers, which is called hemoglobin dissociation. In this article the dissociation of bovine Hb is studied by measurment of the average MW of the samples using the multiangle laser light-scattering method. Advanced multiangle laser light scattering technique is a powerful method to determine the absolute molecular weights of the protein in solution. Two different methods, microbatch multiangle light-scattering (MALS) and on-line size-exclusion high-performance liquid chromatography light scattering with refractive index detector, are used to measure the average molecular weight of bovine Hb in different concentration respectively. The results of the two methods are agreed well. From the results, it can be concluded that the average molecular weigh of bovine Hb will be about 54 kDa when the bovine Hb concentration is more than 1.5 mg/mL, and will be about 36 kDa when the concentration is less than 0.03 mg/mL. The other conclusion, which can be derived from these results, is that the dissociation of bovine Hb is related with the pH and the tetramer appears to be more stable in the pH range of 6-9. PMID- 15274431 TI - Hemoglobin polymerized with a naturally occurring crosslinking agent as a blood substitute: in vitro and in vivo studies. AB - A naturally occurring crosslinking agent, genipin, extracted from the fruits of Gardenia jasminoides ELLIS was used by our group to chemically modified biomolecules. Genipin and its related iridoid glucosides have been widely used as an antiphlogistic and cholagogue in herbal medicine. Our previous study showed that the cytotoxicity of genipin is significantly lower than glutaraldehyde. The study was to investigate the feasibility of using genipin to polymerize hemoglobin as a blood substitute. The results indicated that the rate of hemoglobin polymerization by glutaraldehyde was significantly faster than that by genipin and it readily produced polymers with molecular masses greater than 500,000 Da. It was found that the maximum degree of hemoglobin polymerization by genipin was approximately 40% if over-polymerization is to be prevented. With increasing the reaction temperature, hemoglobin concentration, and genipin-to hemoglobin molar ratio, the duration taken to achieve the maximum degree of hemoglobin polymerization by genipin became significantly shorter. The P50 value of the unmodified hemoglobin was 9 mmHg, while that of the genipin-polymerized PLP-hemoglobin increased to 21 mmHg. It was found in a rat model that the genipin polymerized PLP-hemoglobin resulted in a longer circulation time than the unmodified hemoglobin. In conclusion, the results of the study indicated that the genipin-polymerized hemoglobin solution has a lower oxygen affinity and a longer vascular retention time than the unmodified hemoglobin solution. PMID- 15274432 TI - Blood group B degrading activity of Ruminococcus gnavus alpha-galactosidase. AB - Ruminococcus gnavus is a Gram positive, nonspore-forming obligate anaerobe normally found in the human alimentary tract. In culture, this organism constitutively produces a 1-3 alpha-galactosidase. We fractionated and characterized this enzyme demonstrating hydrolysis of the B epitope on erythrocyte membranes and seroconversion to H epitope (blood type O). Since the enzyme yield was low, cell suspension studies could not be performed. Instead, hydrolysis of the B membrane epitope was studied with an ELISA. A highly purified enzyme product was analyzed for characteristics such as pH, ionic strength, and temperature optimum. Activity in red cell preservative solutions and in the presence of type B plasma was also demonstrated. Ruminococcus gnavus a 1-3 alpha galactosidase has potential application in the enzymatic conversion of type B to O packed red blood cell units. PMID- 15274433 TI - Encapsulation of brewers yeast in chitosan coated carrageenan microspheres by emulsification/thermal gelation. AB - Brewers yeast was encapsulated in kappa-carrageenan microspheres using an emulsification-thermal gelation approach. Due to heat sensitivity of the yeast at temperatures in excess of 36 degrees C, mixtures of low and high gelation temperature carrageenans were tested to obtain a blend yielding a gelation temperature under 40 degrees C. A 20:80 dispersion of 2% carrageenan sol containing cells, in warm canola oil, produced microspheres upon cooling, with a mean diameter of 450 microm and narrow size dispersion (span of 1.2). Application of a chitosan membrane coat to minimize cell release, increased the mean microsphere diameter to 700 microm, due to the coat thickness and swelling of the microspheres. This diameter was designed so as to minimize mass transfer limitations. Batch fermentations were carried out in a 3 L reactor on a commercial wort medium. Cell loading was 10(7) cells mL(-1) microspheres, and cell "burst" release was observed upon inoculation into fresh medium, whether microspheres were coated or not. The kinetics of intra- and extracapsular cell growth were determined. Increased concentrations of extracapsular free cells could be accounted for by growth in the wort medium, and by ongoing release from the gel microspheres, whether coated or not. Cell release from chitosan-coated carrageenan microspheres was less than that from uncoated microspheres, likely due to retention by the membrane coat. Growth kinetics and alpha-amino nitrogen consumption of encapsulated yeast were higher than that of free cells, and differences in alcohol and ester profiles were also observed, likely due to modified metabolism of the encapsulated yeast. PMID- 15274434 TI - Polyhemoglobin-tyrosinase, an oxygen carrier with murine B16F10 melanoma suppression properties: a preliminary report. AB - Melanoma now represents the fifth most common cancer in North America and it has increased dramatically in the past decade. One of the approaches shows that lowering of tyrosine level can inhibit the growth of melanoma in cell culture and in mice bearing B16BL6 melanoma. However, human cannot tolerate the tyrosine restricted diets for lowering tyrosine due to nausea, vomiting, and severe body weight loss. We therefore prepare a novel soluble polyhemoglobin-tyrosinase complex. Our studies show that this preparation can lower systemic tyrosine level in normal animals. This preparation also prevents the native tyrosinase from having adverse effects and from rapid removal after injection. In cell culture study, we find that this preparation inhibits the growth of murine B16F10 melanoma culture. Furthermore, in animal studies we observe that daily intravenous injection of this polyhemoglobin-tyrosinase preparation significantly delays the growth of B16F10 melanoma in mice, without causing adverse effects or changes in the growth of the treated animals. PMID- 15274435 TI - Chitosan adsorbents carrying amino acids for selective removal of low density lipoprotein. AB - Chitosan beads carrying various amino acids (a total of 12 kinds) were synthesized through quite simple procedures for selective removal of low density lipoprotein (LDL). Macroporous chitosan beads were prepared by the phase inversion method, to which the amino acids were then coupled respectively, via either ethyleneglycol diglycidylether (EGDE) or epichlorohydrin (ECH). Among the amino acids used, in vitro tests proved L-Trp to be the best ligand for binding LDL. The adsorbent, which was prepared by coupling L-Trp to the chitosan beads via EGDE, demonstrated satisfactory adsorption performance for selective removal of LDL in human plasma. PMID- 15274436 TI - Immobilized jerusalem artichoke (Helianthus tuberosus) tissue electrode for phenol detection. AB - A tissue based biosensor for the determination of phenol was developed by using Jerusalem artichoke (Helianthus tuberosus) in combination with a dissolved oxygen (DO) probe. The tissue electrode response depends linearly on phenol concentration between 0.002 and 0.0101 microM in 10 min response time. Maximum electrode response was found in phosphate buffer at pH 8.0 and 35 degrees C. The reproducibility of the enzyme electrode was also tested by using standard phenol solutions (0.005 microM). The standard deviation (SD) and variation coefficient (cv) were calculated as +/- 1.4 x 10(-4) microM and 3.1%, respectively. PMID- 15274437 TI - A smart bioconjugate of trypsin with alginate. AB - Alginate is a polymer of guluronic acid and mannuronic acid residues and is an inexpensive, nontoxic polysaccharide of marine origin. Trypsin was immobilized noncovalently on alginate with 100% retention of activity. The enzyme did not leach off the polymer even in the presence of 0.01 M HCl and Triton X-100 (0.2% vv(-1)). The V(max)/K(m) values did not change significantly on immobilization. There was 22% loss of activity in first cycle of pH change and after that the conjugate could be reused upto 4 precipitation cycles without any further loss of activity. This smart bioconjugate was also found to have better operational stability in the presence of casein than free enzyme. Fluorescence studies were carried out to probe structural changes upon immobilization. PMID- 15274438 TI - Analysis of the insect os-d-like gene family. AB - Insect OS-D-like proteins, also known as chemosensory (CSP) or sensory appendage proteins (SAP), are broadly expressed in various insect tissues, where they are thought to bind short to medium chain length fatty acids and their derivatives. Although their specific function remains uncertain, OS-D-like members have been isolated from sensory organs (including the sensillum lymph in some cases), and a role in olfaction similar to that of the insect odorant binding proteins (OBP) has been suggested for some. We have identified 15 new OS-D-like sequences: four from cDNA clones described herein and 11 from sequence databases. The os-d-like genes from the Anopheles gambiae, Apis mellifera, Drosophila melanogaster, and Drosophila pseudoobscura genomes typically have single, small introns with a conserved splice site. Together with all family members entered on GenBank, a total of 70 OS-D-like proteins, representing the insect orders Diptera, Dictyoptera, Hymenoptera, Lepidoptera, Orthoptera, and Phasmatodea, were analyzed. A neighbor joining distance phenogram identified several protein similarity classes that were characterized by highly conserved sequence motifs, including (A) N-terminal YTTKYDN(V/I)(N/D)(L/V)DEIL, (B) central DGKELKXX(I/L)PDAL, and (C) C-terminal KYDP. In contrast, three similarity classes were characterized by their diversion from these conserved motifs. The functional importance of conserved amino acid residues is discussed in relation to the crystal and NMR structures of MbraCSPA6. PMID- 15274439 TI - Orientation and feeding responses of the pollen beetle, Meligethes aeneus, to candytuft, Iberis amara. AB - The pollen beetle, Meligethes aeneus, which is an important pest of oilseed rape, Brassica napus, and turnip rape, B. rapa var. campestris, does not oviposit in all species of the Brassicaceae. The relationship between M. aeneus and candytuft, Iberis amara (Brassicacae), was investigated as part of chemical ecological studies into the development of control methods employing non-host derived repellents. In choice and nonchoice feeding tests, M. aeneus completely rejected I. amara. However, in a field experiment using traps baited with flowering racemes of I. amara and B. napus, M. aeneus was attracted to both species. Gas chromatographic (GC) and GC-electroantennogram (GC-EAG) analyses indicated that the profiles of the floral volatiles of the two species are different. At least 12 compounds among the I. amara floral volatiles were detected by the M. aeneus antenna, and, of these, hexanoic acid, (E)-4,8-dimethyl 1,3,7-nonatriene and alpha-cedrene were not found among B. napus flower volatiles. Since M. aeneus is stimulated by floral volatiles to approach I. amara, but rejects it near, or at, the plant surface, I. amara does not produce repellents that could be used to manipulate M. aeneus. However, it may contain feeding deterrent(s) that could be used in "push-pull" control techniques or in the development of resistant brassicaceous crops. PMID- 15274440 TI - Glandular trichome extracts from Medicago sativa deter settling by the potato leafhopper Empoasca fabae. AB - Extracts of glandular trichomes from Medicago sativa were tested for their ability to disrupt the settling behavior of the potato leafhopper, Empoasca fabae. Erect and procumbent glandular trichomes were mechanically isolated from stem sections of resistant genotype "G98A," and nonglandular trichomes were collected from susceptible cultivar "Ranger." Isolated trichomes were extracted with chloroform, acetone, and ethanol, and the resulting crude extracts were applied to the surface of a sachet containing an artificial diet. Leafhoppers were offered a two-way choice between crude trichome extracts from G98A and Ranger. All three of G98A solvent extracts caused various degrees of diet rejection, resulting in the crude Ranger trichome extracts being preferred over G98A extracts. Overall, the fewest leafhoppers settled on the ethanolic extracts. Additional bioassays documented a dose response associated with G98A ethanolic extracts when compared with Ranger trichome extracts and a solvent control. No difference in preference behavior was detected between Ranger trichome extracts and a solvent control. Gas chromatography-mass spectrometry analysis revealed a homologous series of nonvolatile fatty acid amides C(n)H(2n+1)NO (n = 19-23) unique to G98A glandular trichome extracts. PMID- 15274441 TI - Identification of volatile synomones, induced by Nezara viridula feeding and oviposition on bean spp., that attract the egg parasitoid Trissolcus basalis. AB - Bean plants (Vicia faba L. and Phaseolus vulgaris L.) damaged by feeding activity of Nezara viridula (L.) (Heteroptera: Pentatomidae), and onto which an egg mass had been laid, produced volatiles that attracted the egg parasitoid Trissolcus basalis (Wollaston) (Hymenoptera: Scelionidae). Extracts of volatiles of broad bean and French bean plants induced by adults of N. viridula as a result of their feeding activity, oviposition activity, and feeding and oviposition activity combined were analyzed by gas chromatography-mass spectrometry (GC-MS), and tested in Y-tube olfactometer bioassays as attractants for T. basalis females. In extracts from undamaged leguminous plants, green-leaf volatiles were absent or scarcely detected, and monoterpenes and sesquiterpenes were present at trace levels. No significant differences were detected in the profiles of volatiles of undamaged plants, and undamaged plants on which bugs were allowed only to lay eggs. In contrast, feeding and oviposition by adults of N. viridula induced in both leguminous plants a significant increase in terpenoids such as linalool, (E) beta-caryophyllene, (E,E)-4,8,12-trimethyl-1,3,7,11-tridecatetraene, and (3E)-4,8 dimethyl-1,3,7-nonatriene, which was induced only in French bean plants. Quantitative comparisons revealed increased levels of (E)-beta-caryophyllene in extracts from feeding-damaged plants with N. viridula egg masses compared to feeding-damaged plants without egg masses. In Y-tube olfactometer bioassays, T. basalis females were attracted by extracts of both leguminous plants only when N. viridula adults were allowed to feed and oviposit upon them. Fractionation of extracts of volatiles from broad bean plants with N. viridula feeding damage and egg masses yielded two fractions. but only the fraction containing (E)-beta caryophyllene was attractive to the egg parasitoid. These findings indicate that N. viridula feeding and oviposition induce leguminous plants to produce blends of volatiles that are characterized by increased amounts of (E)-beta-caryophyllene, and these blends attract female T. basalis. The role of (E)-beta-caryophyllene as a potential synomone for T. basalis is discussed. PMID- 15274442 TI - Laboratory and field responses of the mosquito, Culex quinquefasciatus, to plant derived Culex spp. oviposition pheromone and the oviposition cue skatole. AB - Laboratory and field studies were conducted on the oviposition behavior of the pathogen-vectoring mosquito, Culex quinquefasciatus, in response to the oviposition pheromone 6-acetoxy-5-hexadecanolide, produced from a renewable plant resource, Kochia scoparia (Chenopodiaceae) (plant-derived pheromone, PDP), and via an established synthetic route (synthetic oviposition pheromone, SOP). Responses to the oviposition cue skatole (3-methylindole), presented individually and in combination with the plant-derived and synthetic oviposition pheromone, were also studied. Both laboratory and field assays showed that PDP and SOP were equally attractive. Synergistic effects were observed with one combination of PDP and skatole combinations in laboratory assays. Synergy was also observed under field conditions. SOP and skatole combinations showed additive effects in laboratory assays, but were not tested in field bioassays. Although synergism has been previously demonstrated with combinations of SOP and polluted waters, the work presented here is the first example of synergy between a specific oviposition attractant and the oviposition pheromone. Furthermore, the efficacy of mosquito pheromone produced from a cheap, renewable botanical source has been demonstrated. PMID- 15274443 TI - Volatiles associated with preferred and nonpreferred hosts of the Nantucket pine tip moth, Rhyacionia frustrana. AB - Ovipositing female Nantucket pine tip moth, Rhyacionia frustrana, prefer loblolly pine, Pinus taeda L., to slash pine, Pinus elliottii Engelm, except during the first spring following planting of seedlings. Host discrimination by R. frustrana increases as seedlings develop, suggesting that changes in the chemical composition of seedlings may mediate the moth's host preferences. Volatile compounds from slash and loblolly pine seedlings were collected using solid-phase microextraction (SPME) during the first year following planting. Four collection periods coincided with adult emergence and oviposition during each of four annual generations of R. frustrana in the Georgia Coastal Plain. Infestation of slash pine peaked during the second tip moth generation and was similar to the loblolly pine infestation level. By the fourth tip moth generation, slash pine infestation levels had declined and diverged considerably from those of loblolly pine. Significant differences in relative quantities of beta-pinene, alpha phellandrene, limonene, beta-phellandrene, bornyl acetate, beta-caryophyllene, and an unidentified sesquiterpene occurred between slash and loblolly pine during the fourth generation. However, no strong correlation was observed between any individual compound and host damage that could readily explain the temporal changes in R. frustrana host preference. Gas chromatographic electroantennographic detection (GC-EAD) analyses of standards identified 19 different seedling-associated compounds that elicited antennal responses from R. frustrana females, indicating that a blend of terpenoids may mediate host discrimination. PMID- 15274444 TI - Potential for use of synthetic sex pheromone for mating disruption of the olive pyralid moth, Euzophera pinguis. AB - The potential for pheromone-based mating disruption of the olive pyralid moth (OPM), Euzophera pinguis, in olive groves was investigated during the second flight period in small-plot trials in 2002. The female of this species emits a blend of (9Z,12E)-tetradecadien-1-ol and (9Z,12E)-tetradecadienyl acetate, which were synthesized for field tests. Mating disruption efficacy in 0.8-ha trials was evaluated using two parameters: reduction of male capture in pheromone traps and reduction of infestation in infestation-prone sites. White rubber septa containing 10 mg of pheromone blend as disruptant were applied at a density of 50 septa/ha for each treatment. Mean catches of E. pinguis males in pheromone traps were greatly reduced (> 95%) in pheromone-treated plots relative to similar traps placed in control plots. In addition, significant reductions were recorded (35 40%) in the oviposition and infestation levels during pheromone treatment. The total amount of pheromone blend released from disruption dispensers during the field trials was estimated to average 5.4 mg/ha/day, over 56 days. PMID- 15274445 TI - The effect of exogenous jasmonic acid on induced resistance and productivity in amaranth (Amaranthus hypochondriacus) is influenced by environmental conditions. AB - Amaranthus hypochondriacus is a C4 pseudocereal crop capable of producing reasonable grain yields in adverse environmental conditions that limit cereal performance. It accumulates trypsin inhibitors and alpha-amylase inhibitors in seeds and leaves that are considered to act as insect feeding deterrents. Foliar trypsin and alpha-amylase inhibitors also accumulate by treatment with exogenous jasmonic acid (JA) in controlled laboratory conditions. Three field experiments were performed in successive years to test if two nonphytotoxic dosages of JA were capable of inducing inhibitor activity in A. hypochondriacus in agronomical settings, and if this induced response reduced insect herbivory and insect abundance in foliage and seed heads. The performance of JA-treated plants was compared to insecticide-treated plants and untreated controls. The effect of exogenous JA on the foliar levels of six additional putatively defence proteins was also evaluated. Possible adverse effects of JA induction on productivity were evaluated by measuring grain yield, seed protein content, and germination efficiency. The results present a complex pattern and were not consistent from year to year. To some extent, the yearly variability observed could have been consequence of growth under drought versus nondrought conditions. In a drought year, JA-treated plants had lower levels of insect herbivory-derived damage in apical leaves and panicle than control plants, whereas in nondrought years, there was an inconsistent effect on aphids, with no effect on lepidopteran larvae. JA treatments reduced the size of the insect community in seed heads. The effect varied with year. Exogenous JA did not adversely affect productivity, and in the absence of drought stress, the higher dosage enhanced grain yield. Induction of defensive proteins by JA, although sporadic, was more effective in nondrought conditions. The patterns of foliar protein accumulation observed suggest that they may be part of a constitutive, rather than inducible, chemical defense mechanism that is developmentally regulated and critically dependent on the environment. The results emphasize the difficulties that are often encountered when evaluating the performance of chemical elicitors of induced resistance in field settings. PMID- 15274446 TI - Genetic analysis of benzoquinone production in Tribolium confusum. AB - Many species of tenebrionid beetles produce and secrete benzoquinones from specialized prothoracic and postabdominal glands. Tribolium confusum produces two compounds methyl-1,4-benzoquinone (MBQ) and ethyl-1,4-benzoquinone (EBQ). These compounds are hypothesized to function as external defense compounds, killing microbes and deterring predators, and their ability to evolve by natural selection depends on both selection and the genetic vs. environmental contribution to phenotypic variation. We crossed a strain of T. confusum that produces high quantities of benzoquinones, b-Pakistan, with a low-producing strain, b-+, and measured both the internal and external quantities of MBQ and EBQ for the two extreme strains and their F1 progeny. Internal amounts show a clear pattern of inheritance, with at least 50% of the phenotypic variation attributed to genotype. Additive and dominance coefficients for internal amounts indicate that the trait is additive with no significant dominance. In contrast, external quantities show little pattern of inheritance. The role of genetics and environment in determining quantities of secretory defensive compounds is important to elucidating the ecology and evolutionary potential of chemical defenses. PMID- 15274447 TI - The chemistry of pollination in selected Brazilian Maxillariinae orchids: floral rewards and fragrance. AB - We report the chemical composition of the floral rewards and the fragrance of 10 Maxillariinae (Orchidaceae) species. The species that offer rewards (labellar secretions) are usually scentless, the rewards being collected by bees. Chemical analyses revealed that the major chemical class of compounds present in the labellar secretions are triterpenoids. The rewardless Maxillariinae flowers were usually scented, and chemical analyses of their volatiles revealed that they were composed of mono and sesquiterpenoids. PMID- 15274448 TI - Humic substances affect the activity of chlorophyllase. AB - Three humic substances--humic acid, fulvic acid, and humin--were isolated from soils located in the northern and southern forests of the Yuanyang Lake Nature Preserve in northern Taiwan's Ilan County. Aqueous extracts of fresh wet soil and of three humic substances, at concentrations of 0.125, 0.25, and 0.5 mg/ml, were investigated for their effects on the activities of chlorophyllase a and b. Aqueous extracts of forest soils at the northern and southern bank, dominated by the pure vegetation of Formosan False cypress (Chamcaecyparis formosensis Matsum), stimulate both chlorophyllase a and b activities, while those of the southern bank, dominated by a Taiwanese Miscanthus (Miscanthus transmorrisonensis Hayata), inhibits such activities. All three humic substances, despite their soil sources, stimulate the activities of both chlorophyllase a and b. Fulvic acid stimulates more chlorophyllase a activity than either humic acid or humin. Humic acid stimulates more activity of chlorophyllase b than either fulvic acid or humin. Humin exhibited the least effect on chlorophyllase a and b. It is suggested that humic substances in the soil may accelerate the chlorophyll degradation of litter in the ecosystem and that chlorophyllase a and b may be different enzymes. PMID- 15274449 TI - Allelochemicals of Polygonella myriophylla: chemistry and soil degradation. AB - Gallic acid and hydroquinone have been identified as the major allelochemicals of the known allelopathic plant Polygonella myriophylla. Both of these compounds occur in the foliage as glycosides. Quercetin and rhamnetin were identified as the major flavonoid constituents, but in much lower concentration. The behavior of gallic acid, hydroquinone, the hydroquinone glycoside arbutin, and benzoquinone in sterile and nonsterile soil from beneath Polygonella was investigated. Sterilization effectively stabilized arbutin, hydroquinone, and gallic acid. Concentrations of benzoquinone rapidly diminished in sterilized soil, and the compound was almost completely gone after 7 days. In nonsterile soils, all four compounds degraded rapidly. The order of persistence was hydroquinone > benzoquinone > gallic acid > arbutin. Persistence was rate dependent. Arbutin degraded to hydroquinone, and benzoquinone formed as a degradation product of hydroquinone. Hydroquinone was also observed as a degradation product of benzoquinone. Benzoquinone degrades rapidly by nonmicrobial oxidative processes. These results support the hypothesis that microbial and nonmicrobial oxidative transformations of soil allelochemicals are crucial in mediating the allelopathic effects of Polygonella myriophylla. PMID- 15274450 TI - [Breast cancer: how to incite women to participate in screening programmes]. PMID- 15274451 TI - [Why antiretroviral therapy must not begin to early?]. PMID- 15274452 TI - [Breast cancer: personalised care]. PMID- 15274453 TI - [Breast cancer: generalised screening in France from now on]. AB - With the first randomised screening trials for breast cancer having proved in the 1980s the efficacity of screening, a more ambitious programme has been put in place in France from 1989 to 2001. Based on existing radiological structures, it consists of one image per breast, 2 readings, every 3 years. The management is departmental. It is rapidly completed by quality assurance programmes of the radiology companies. The evaluation of the screening programme has been undertaken by the VS since 1998 and uses the European indicators of efficacity. The results, heterogenous at the beginning, later improved: from 1990 to 1998, the percentage of small size invasive cancers or those equal to 10 mm increased from 30% to 35%, and the second reading has permitted, according to the departments, the detection of 10% to 25% of supplementary cancers. In 1999, the screening was modified: 2 images per breast, 2 readings [corrected] The protocol published in 2001 involves a clinical examination by a radiologist, 2 (or 3) images per breast, 2 readings if the examination is normal [corrected] The second reading is centralised and made by a specialised radiologist [corrected] The place for ultrasound and numerical mammography hasn't yet been defined. Since the beginning of 2004, the generalisation of the screening programme has been effective in France. PMID- 15274454 TI - [Breast cancer: atlas]. PMID- 15274455 TI - [Positron emission tomography combined with computed tomography overturns the initial treatment plan]. PMID- 15274456 TI - [Notification of the diagnosis of breast cancer]. PMID- 15274457 TI - [Multidisciplinary approach in breast cancer care]. AB - With the improvement of imaging, pathology, surgery, and medical treatment, breast carcinoma care has moved toward a multidisciplinary approach to co ordinate expertise of multiple medical specialists and to achieve an optimal management. In this multidisciplinary team, the patient must keep a "reference physician" who may interact with her on behalf of the whole team and the general practitioner. PMID- 15274458 TI - [Breast-conserving therapy in breast cancer]. AB - The combination of lumpectomy, axillary node treatment and radiotherapy of the breast is the base of breast-conserving therapy. This combination of surgery and radiotherapy is now accepted as a standard treatment option for unifocal, non inflammatory lesion less than 3 cm. The widespread use of mammography to detect infraclinic breast carcinoma leads to a significant increase in the proportion of breast conserving treatment. Neoadjuvant therapeutics (chemotherapy, radiotherapy and hormonotherapy) can extend the standard indication to breast carcinoma larger than 3 cm. The standard definition is also modified by sentinel node biopsy, oncoplastic techniques and stereotactic surgery with satisfactory cosmetic results. The risk of local recurrence, particularly the margins, must be evaluated whatever the surgical treatment optimizing oncologic management. PMID- 15274459 TI - [Medical treatments for breast cancer]. AB - Breast carcinoma is no longer considered as a local disease. So, surgery and radiotherapy could be insufficient to control undetectable distant micro metastasis. Naturally, systemic medical procedures have been extended from metastatic stages to adjuvant and neo-adjuvant settings. The double sensitivity to both hormonal and chemical agents give to clinicians a large number of putative choices. Recent discovery of membrane receptor-targeted monoclonal antibodies enriched our armentarium. PMID- 15274460 TI - [Therapeutic implications of advances in breast cancer biology]. AB - The polygenic and multifactor genetic basis of breast cancer confers to each tumour a different phenotype and clinical outcome. A therapeutic stake is to better determine this heterogeneity by using more reliable prognostic factors and to develop molecular therapies targeting the tumour cells selectively. The study of molecular alterations in breast cancer allowed considerable therapeutic progress by use of the hormonal receptors and the ERBB2 receptor. Today, new high throughput technologies such as DNA microarrays allow measuring the activity of thousands of genes in a sample simultaneously. The awaited repercussions are multiple. Expression profiling of breast tumours allows the identification of new sub-groups of tumour in groups a priori identical, but with different outcome. This stratification should make it possible to better tailor the treatment and boost the discovery of new therapeutic targets PMID- 15274461 TI - [Patient checklist. Prevention of lymphedema of the upper limb]. PMID- 15274462 TI - [Residents and pharmaceutical industry]. PMID- 15274463 TI - [Mood disorders. Bipolar psychosis]. PMID- 15274464 TI - [Cardiac murmurs in children]. PMID- 15274465 TI - [Intracerebral tumours]. PMID- 15274466 TI - [Principal complications of pregnancy: gestational diabetes]. PMID- 15274467 TI - [Sport and health. Sport aptitude in children and adults. Nutritional needs in sportsmen]. PMID- 15274468 TI - [Normal grief and complicated grief]. PMID- 15274469 TI - [What paleopathology reveals about the fate of individuals with malformations in greco-roman antiquity]. PMID- 15274470 TI - Immediate traumatic thumb loss reconstruction: case reports. AB - In the article we present and evaluate our successful immediate attempts of traumatic thumb loss reconstruction in the last fourteen years: early passive stump elongation, immediate pollicization and replantation. Our experience confirms the known fact that in spite of unfavorable oblique transarticular thumb amputation with interphalangeal joint destruction, considerable bone shortening and interphalangeal joint fusion, successful replantation offered us better cosmetic and partially functional outcome than the one we have obtained ealier with other methods of immediate thumb reconstruction. PMID- 15274471 TI - Breast asymmetry: a new vision of this malformation. AB - Breast asymmetry is defined as a difference of form, position or volume of the breast and is a pathology affecting over half of the female population (1, 4). Fortunately although it is very frequent, surgical correction is seldom necessary. The authors present the results of the surgical treatment of 73 patients affected with breast asymmetry and they describe the employment of a new instrumental method (SCAN-3D) that may be used both within the correction program and during the follow-up of the surgical treatment. The system involves the use of a laser to scan the surface to be acquired and by the subsequent analysis of the image produced by the overlap of itself with the area under examination, by utilising specific software, we retrieve information regarding both the morphologic and volumetric connections in all three dimensions. The employment of new research methods (Three-dimensional scanning) is an important aid to the aesthetic surgeon, as it allows the study of the mammary glands morpho-volumetric component in relation to the rest of the thorax. Moreover, in the future, the custom production of breast implants, made to measure, and perfectly adaptable to the deformity of each patient may be possible. PMID- 15274472 TI - Use of polylactic acid in face lipodystrophy in HIV positive patients undergoing treatment with antiretroviral drugs (HAART). AB - OBJECTIVES: The recent use of antiretroviral drugs in people with HIV infection produced a drastic reduction in mortality and a remarkable improvement in the quality of life for these patients. However the aesthetic and psychological consequences that come from the reorganization of the adipose tissue induced by these drugs (facial wasting, buffalo hump) may reduce this state of wellbeing. METHODS: Our group, in cooperation with the 3rd Division of Infective Diseases of the University of Rome "La Sapienza", decided to evaluate the efficacy of polylactic acid (PLA) in combating these problems in HIV positive patients undergoing treatment with antiretroviral drugs in an evaluating study of 4 cases. The evaluation of the obtained results was performed by follow-up at one, three, six and twelve months from the first infiltration and considering the clinical evaluation, the photographical documentation and the patient's judgement. RESULTS: In all cases we obtained an improvement of the local condition with the restoration of a more booming aspect. The change was noted both within the family and the work environment. In the first three cases the results were considered as optimal by both the operators and the patients. In the fourth case the result was evaluated as discreet (a remarkable loss of weight of the patient in the months following the treatment should be taken into consideration). CONCLUSIONS: We think a new study with ultrasonography of dermal thickness before, during and after the treatment is necesary. This study will give a more reliable evaluation of the product efficacy. PMID- 15274473 TI - Mucosal malignant melanoma in upper aerodigestive tract: report of two cases. AB - Primary malignant melanoma has been seen in virtually all sites and organ systems where neural crest cells migrate. Upper aero-digestive tract is the unusual site. We presented two cases of mucosal malignant melanoma. A 59-year-old female patient had been suffering from nasal airway obstruction and headache for three months, and was admitted to our clinic. There was an upper cervical lymphadenopathy 5 cm in diameter ipsilaterally. Computerized tomography presented a nasal mass originating from the left maxillary sinus, and extending into nasal cavity and the left lateral wall of nasopharynx. Left total maxillectomy and radical neck dissection was performed. Postoperative radiotherapy was applied. A 68-year-old male patient suffering from nasal airway obstruction, hypernasality, dysphagia, and oral bleeding was admitted to our clinic. Oral examination revealed the tumoral mass protruding downward through oropharynx. The second patient referred to medical oncology. There was no recurrence in the first patient 18 months later. And the second patient died one year later. We believe that nasal mucosal malignant melanoma was rare, and carefully evaluated since it may be amelanotic lesion. Nasopharyngeal mucosal malignant melanoma is very rare, and has worse prognosis. PMID- 15274474 TI - Total antioxidant capacity of serum and prognostic indices in patients with burn trauma. AB - Burn trauma leads to increased production of reactive oxygen species and compromises the antioxidant systems. The aim of present study was to assess the total antioxidant capacity of plasma (TAC) in burn patients and evaluate its usefulness in clinical practice through analysis of association between TAC and indices of patient prognosis. We investigated TAC in 48 adults and 27 children with severe burn trauma and in 26 healthy controls. TAC was measured on the admission and every week thereafter until discharge or death of patients. The prognosis of the patients was evaluated by the Baux Index and the Abbreviated Burn Severity Index (ABSI). TAC was significantly decreased in both groups with burn trauma as compared with the controls and the decrease was a long lasting event. Significant indirect negative correlation was found between BSAB and TAC values measured in the later phase of injury. In adults an indirect correlation was found between ABSI and the lowest TAC measured on the 7th day or later. Correlations between TAC values and Baux Index were absent both in adults and in children. PMID- 15274475 TI - Fifty years of the Cleft Centre in Bratislava. AB - In our paper, we would like to present and analyse the activities in the Cleft Centre in Bratislava between 1952 and 2001. We document the gradual development of the complex treatment and the important changes in the usage of the operative techniques and in the primary and secondary operations timing, as well as the application of the new trends in the field of plastic surgery, maxillofacial surgery, anaesthesiology, orthodontics, speech therapy, paediatrics, human genetics and psychology. PMID- 15274476 TI - Genetics of metal resistance in acidophilic prokaryotes of acidic mine environments. AB - Acidophilic bacteria inhabiting acidic mine regions cause natural leaching of sulphidic ores. They are now exploited in industrial operations for leaching of metals and beneficiation of low-grade and recalcitrant ores. Recent trends emphasize application of thermoacidophiles and genetic engineering of ore leaching bacteria for greater success in this area. This requires an in-depth understanding on the molecular genetics of these bacteria and construction of cloning vectors for them. Metal resistance is considered as the most suitable phenotypic trait for cloning vectors of bio-mining chemolithoautotrophic (viz. Acidithiobacillus ferrooxidans) and heterotrophic (Acidiphilium and Acidocella species) bacteria of mine environments. These bacteria take part in ore-leaching either directly or indirectly, exhibit low to high level of resistance/tolerance to various metals under different conditions. Majority of these bacteria contain one or more plasmids--the genetic elements that usually carry metal resistant genes. But none of the At. ferrooxidans plasmids has been definitely proved to harbour metal-resistant genes which have mostly been found in the chromosome of this bacterium. Plasmids of acidophilic heterotrophs of the genera Acidiphilium and Acidocella, on the other hand, carry metal resistant genes. While genes bestowing arsenic resistance in Acidiphilium multivorum are similar to those analyzed from other sources, the metal (Cd and Zn)-resistance conferring cloned plasmid DNA fragments from Acidiphilium symbioticum KM2 and Acidocella GS19h strains were found to have no sequence similarity with the reported Cd- and Zn resistant genes. Such observations indicate some novel aspects of metal resistance in acidophilic bacteria. PMID- 15274477 TI - Goat serum: an alternative to fetal bovine serum in biomedical research. AB - Serum is frequently added to the defined basal medium as a source of certain nutritional and macromolecular growth factors essential for cell growth. Although a number of synthetic media have been prepared serum continues to be used in cell culture by many investigators. The best supplementation to a basal medium is fetal bovine serum (FBS) that is most frequently used for all types of cell cultures. During last four decades National Institute of Virology, Pune, has been working on isolation and identification of viruses from clinical specimens, employing tissue culture. Initially FBS was used for this purpose. However, due to its prohibitive cost and uncertain supply an alternative was sought. Commercially available sera from newborn calf, sheep, horse, human and serum obtained from goat blood (available from local abattoir) were tried. Goat serum (GS) was found to be suitable for most of the cell lines and primary cultures. Primary cultures from guinea pig embryo, monkey kidney, chick embryo, mouse peritoneal macrophages, and established cell lines were prepared and grown in growth media supplemented with GS. These cultures were studied for their morphology and growth in comparison with cultures grown in FBS containing media, and were used for mass cultivation of cells, quantitation and susceptibility of various virus strains, studies on effects of different nutrients and natural substances on cellular metabolism and virus replication, epitope analysis of various strains of Japanese encephalitis (JE) virus, strain differentiation studies, studies on antibody dependent plaque enhancement, assay of murine migration inhibition factor. Monoclonal antibodies against JE virus adapted to GS were characterised for their retention of functionalities. The results were comparable to those of cell cultures grown in FBS containing media. Similar results on chromosome studies were obtained from patient's whole blood cultures prepared in GS and FBS containing growth media. Organ cultures from mammalian, reptile and avian hosts; successfully grown in GS supplemented growth media, were used for different virological studies. Growth media supplemented with GS were used for in vitro cultivation of malarial parasites. Thus since the last three decades many scientists are using GS in place of FBS, in various fields of biomedical research. The present article reviews an account of the same. PMID- 15274478 TI - Differential depression of spinal synaptic transmission in vitro by different hypoxic insults. AB - The effects of hypoxia (O2-free), aglycemia (glucose-free), ischemia (O2- and glucose-free) and chemical anoxia (by 3-nitropropionic acid; 3-NPA) were evaluated on the synaptic transmission in vitro. Stimulation of a dorsal root in hemisected spinal cord from neonatal rat, evoked monosynaptic (MSR) and polysynaptic reflexes (PSR) in the segmental ventral root. In all the hypoxic conditions, the reflexes were depressed in a time-dependent manner. Hypoxia took longer time (> 240 min) to abolish the reflexes where as, aglycemia and ischemia abolished them within 35 min. Recovery after wash was complete in hypoxia, 60-70% in aglycemia and 20-25% in ischemia. The time required for 50% depression of reflexes (T-50) was also in the same order (100, 23 and 13 min). The elimination of O2 in hypoxic or ischemic solution by N2 bubbling abolished the reflexes within 16 min. The T-50 values in both the conditions were between 5-8 min. Superfusion of 3-NPA (an irreversible inhibitor of succinate dehydrogenase) depressed the reflexes. The abolition time and T-50 values were shorter with the increasing concentrations of 3-NPA. The present results reveal that the energy production in hypoxic condition with normal glucose level can sustain the synaptic activity for a longer time while the glucose deficiency even in normoxic conditions drastically impair the synaptic activity. Further, aglycemia depressed the reflexes almost in a similar time as seen with ischemia. PMID- 15274479 TI - Ca2+ -free medium enhances the magnitude of slow peak in compound action potential of frog sciatic nerve in vitro. AB - The present investigation was carried out to know the effect of Ca2+ on different peaks of compound action potential (CAP) representing the fibers having different conduction velocity. CAP was recorded from a thin bundle of nerve fibers obtained from desheathed frog sciatic nerve. Suction electrodes were used for stimulating and recording purposes. In Ca2+ -free amphibian Ringer, two distinct peaks (Peak I and Peak-II) were observed. The threshold, conduction velocity (CV), amplitude and duration of Peak-I were 0.32 +/- 0.02 V, 56 +/- 3.0 m/sec, 2.1 +/- 0.2 mV and 0.75 +/- 0.1 ms, respectively. The Peak-II exhibited ten times greater threshold, eight times slower CV, three times lower amplitude and four times greater duration as compared to Peak-I. Addition of 2 mM Ca2+ in the bathing medium did not alter CAP parameters of Peak-I excepting 25% reduction in CV. But, in Peak-II there was 70-75% reduction in area and amplitude. The concentration-attenuation relation of Peak-II to various concentrations of Ca2+ was nonlinear and 50% depression occurred at 0.35 mM of Ca2+. Washing with Ca2+ -free solution with or without Mg2+ (2 mM)/verapamil (10 microM) could not reverse the Ca2+ -induced changes in Peak-II. Washing with Ca2+ -free solution containing EDTA restored 70% of the response. The results indicate that Ca2+ differentially influence fast and slow conducting fibers as the activity of slow conducting fibers is greatly suppressed by external calcium. PMID- 15274480 TI - Effect of Aloe vera (L.) Burm. fil. leaf gel and pulp extracts on kidney in type II diabetic rat models. AB - Significant degenerative changes were observed in the kidney tissue of untreated neonatal streptozotocin (n0STZ)-induced type-II diabetic rats. These degenerative changes were diminished in the kidney tissue of diabetic animals given glibenclamide and Aloe leaf gel and pulp extracts. Kidney lipid peroxidation levels were increased in diabetic rats compared to healthy rats; these levels were higher in rats treated with glibenclamide than in those which received Aloe extracts. Serum urea and creatinine levels were higher in diabetic rats in comparison to healthy rats. The administration of Aloe gel extract and glibenclamide decreased serum urea and creatinine levels in comparison to diabetic controls. Only A. vera leaf gel extract showed improvement both in histological and biochemical parameters suggesting a protective effect of A. vera on mild damage caused by type-II diabetes on kidney tissue. PMID- 15274481 TI - Influence of coconut kernel protein on lipid metabolism in alcohol fed rats. AB - Male albino rats were given ethanol (3.76 g/kg body weight/day) to induce hyperlipidemia. The rats showed increased concentration of cholesterol and triglycerides in the serum and tissues. Inclusion of coconut protein and L arginine into ethanol fed rats produced lower levels of total cholesterol, LDL+ VLDL cholesterol, triglycerides and atherogenic index in the serum. Concentration of tissue cholesterol and triglycerides was also lower in these groups. Administration of coconut protein and L-arginine in the ethanol fed rats caused decreased activity of HMG-CoA reductase in the liver and increased activity of lipoprotein lipase in the heart. The activities of malic enzyme and glucose-6 phosphate dehydrogenase were also lower in these groups. Feeding coconut protein and L-arginine in ethanol treated rats showed increased concentration of hepatic bile acids and fecal excretion of neutral sterols and bile acids. All these effects were comparable in rats fed coconut protein and those fed L-arginine. These observations indicate that the major factor responsible for the hypolipidemic effect of coconut protein is due to the high content of L-arginine. PMID- 15274482 TI - Evaluation of dietary essentiality of vitamins for Penaeus monodon (Fabricius). AB - The effect of exclusion of individual water-soluble (thiamine, riboflavin, pyridoxine, cyanocobalamin, pantothenic acid, folic acid, niacin, biotin, choline, inositol, ascorbic acid) and fat-soluble vitamins (vit. A, D, K and E) in semi-purified diets on growth and survival of juvenile shrimp, P. monodon was studied in the laboratory for 8 weeks. Diets lacking riboflavin and vitamin K did not affect growth and survival of shrimp. However, deletion of inositol and choline resulted in poor growth. Maximum growth was observed in the control diet (C1) which was supplemented with all vitamins. Diet deficient in ascorbic acid, biotin, folic acid, niacin, thiamine and alpha-tocopherol resulted in poor appetite and poorer feed conversion efficiency. All treatments except the control (C1) resulted in histological changes in the digestive gland cells. Detachment or destruction of the epithelial cells was observed in all treatments lacking individual vitamins but more severely in the treatment without a vitamin supplement followed by inositol, choline and ascorbic acid deficient diets. PMID- 15274483 TI - Partial suppressive effect of melatonin on indomethacin-induced renal injury in rat. AB - Intramuscular injection of a single high dose of indomethacin (20 mg/kg) in fasted rats produced renal injury. The results showed increases in the level of lipid peroxidation and cholesterol, and activity of acid phosphatase and alkaline phosphatase in the kidney. Also, the renal contents of both reduced glutathione and activity of total adenosine triphosphatase were decreased by the toxicant. In serum, indomethacin increased activity of lactate dehydrogenase and acid phosphatase, and levels of creatinine and inorganic phosphorus. Paradoxically, administration of melatonin (0.75 mg/rat/day) alone for 7 days decreased significantly the activity of lipid peroxidation and acid phosphatase, and increased, but not significantly, the level of reduced glutathione in the kidney. Also, serum level of creatinine tended to decrease, but not significantly. Pretreatment with melatonin prevented the increase by subsequently administered indomethacin in the renal activity of lipid peroxidation and acid phosphatase. However, this pretreatment regimen partially suppressed the adverse changes in the remaining analyzed cytotoxic parameters induced by indomethacin in both serum and kidney. These results indicate that oral administration of melatonin at a low dose level exerted moderate antioxidant action, thereby it protected against some of the renal detrimental effects produced by indomethacin. PMID- 15274484 TI - Standardization of the method for estimation of ethambutol in pharmaceutical preparations and biological fluid. AB - A simple column chromatographic method for determination of ethambutol (EMB) in pharmaceutical preparations containing EMB in combination with other anti-TB drugs is presented. The method involved extraction of EMB into an organic solvent, followed by basification and column chromatographic separation on Amberlite CG 50 (100-200 mesh) and elution with suitable eluants and estimation at a wavelength of 270 nm. The assay was linear from 25 to 400 microg/ml. The relative standard deviations of intra and inter day assays were lower than 5%. Ethambutol was recovered from human urine quantitatively and stable for a period of at least one week in urine stored at -20 degrees C. PMID- 15274485 TI - Effect of precocene on development of ovarian follicles in flesh fly, Sarcophaga ruficornis F. AB - Administration of precocene II (6,7-dimethoxy-2, 2-dimethyl chromene) to freshly emerged virgin female flies of S. ruficornis adversely affected the development and differentiation of ovarian follicles leading to a number of morphological abnormalities. Precocene treatment resulted into suppression of development of egg chamber, differentiation of follicular epithelium, degeneration of nurse cells, growth of oocyte and uptake of yolk granules by oocytes. The results suggest that precocene induced effects are due to deficiency of juvenile hormone. PMID- 15274486 TI - Effect of potassium channel modulators on toxicity of Cleistanthus collinus. AB - The study was conducted to determine the effects of boiled extract of Cleistanthus collinus on rats by observing ECG changes and electrolyte levels in serum and urine. Influence of minoxidil and glibenclamide on Cleistanthus collinus induced toxicity was determined. ED50 for arrhythmia, changes in contractility and heart rate were recorded using the isolated frog heart. Cleistanthus at low doses caused transient tachycardia and increase in contractility and at high dose caused arrhythmia and cardiac arrest in rat. LD50 was found to be 1690 mg/kg. Minoxidil potentiated cardiac toxicity, whereas glibenclamide did not produce any significant change. High concentration of potassium in Cleistanthus extract hindered comparison of its levels. There was excretion of sodium even in the presence of hyponatraemia. Cleistanthus at low dose caused transient tachycardia and increase in contractility and at high dose caused arrhythmia and cardiac arrest in isolated frog heart. ED50 for arrhythmia was found to be 1406 mg/kg. Acute toxicity was mainly due to depressive cardiac activity of Cleistanthus. It also caused renal failure. Potassium channel modulators did not have important role in acute cardiac toxicity treatment. Probably in chronic toxicity, electrolyte level changes are involved and potassium channel modulators might have a role. PMID- 15274487 TI - Effect of nitric oxide on H+ -efflux in presence of various nutrients in Candida albicans. AB - In the present study tentative link has been established between H+ -efflux and effect of NO in presence of various nutrients (glucose, 2-deoxy-D-glucose, xylose, proline, glutamic acid and lysine) in C. albicans using sodium nitroprusside (SNP) as a potent source of NO. It was observed that there was a decreasing trend in pH with time, in control, while SNP treated cells showed an initial decline in pH for 10-15 min, followed by an increase in pH up to 30 min. In presence of glucose there was an enhancement in H+ -efflux by 9-fold whereas proline, glutamic acid and lysine showed enhancement by 3, 6 and 1.5-fold respectively. Similar trends in increase in pH after 15 min in SNP treated cells of Candida was observed in presence of all nutrients used. It was demonstrated for the first time that H+ -ATPase of C. albicans was affected by NO. PMID- 15274488 TI - Bioactivity of non-edible oil seed extracts and purified extracts against Helicoverpa armigera (Hubner). AB - Extracts and purified extracts of seeds of two plant species, Madhuca latifolia and Calophyllum inophyllum when evaluated against the 2nd instar larvae of Helicoverpa armigera reared on synthetic diet, exhibited high larval mortality, prolongation of developmental period, morphological deformities and highly significant reduction in adult emergence. The reduction in larval weights in the treatments was also highly significant. PMID- 15274489 TI - Chemical nature, ligand denticity and quantification of fungal siderophores. AB - Thirtyfive siderophore producing fungi were categorized for their hydroxamate, catecholate or carboxylate nature by chemical and bioassays. Out of 35 fungi, 30 were hydroxamates and 5 showed carboxylate nature. However, none of the fungi produced catecholate type of siderophores. Eighteen out of 29 fungi were trihydroxamate and the rest 11 fungi were dihydroxamates. Twenty-three fungi were hexadentate and 6 were tetradentate in nature. Quantification of siderophores using standard compounds deferrioxamine mesylate and rhizoferrin revealed that Phanerochaete chrysosporium produced maximum among the hydroxamate producing fungi and Mycotypha africana resulted maximum among the carboxylate producing fungi. PMID- 15274490 TI - Production of 2-hydroxy-4-methoxybenzaldehyde in roots of tissue culture raised and acclimatized plants of Decalepis hamiltonii Wight & Arn., an endangered shrub endemic to Southern India and evaluation of its performance vis-a-vis plants from natural habitat. AB - Axillary buds obtained from field grown plants of D. hamiltonii were used to initiate multiple shoots on Murashige and Skoog's medium (MS) supplemented with 2 mg L(-1) 6-benzyl aminopurine (BA) and 0.5 mg L(-1) indole-3-acetic acid (IAA). Profuse rooting was achieved when the actively growing shoots were cultured on MS medium supplemented with 1.0 mg l(-1) indole-3-butyric acid (IBA). Regenerated plants were grown successfully in the plains, in contrast to wild growth in high altitudes and rocky crevices of hilly regions. Roots of different sizes from one year-old tissue culture raised field grown plants had the same profile of 2 hydroxy-4-methoxybenzaldehyde as that of wild plants. A maximum of 0.14% and 0.12% 2-hydroxy-4-methoxybenzaldehyde was produced in roots of one year old tissue culture derived plants and greenhouse grown plants respectively. PMID- 15274491 TI - Culture filtrate of Lasiodiplodia theobromae restricts the development of natural resistance in Brassica nigra plants. AB - Culture filtrate of Lasiodiplodia theobromae increased respiration rate, phenylalanine ammonia lyase activity, and levels of hydrogen peroxide, lipid peroxides and salicylic acid in B. nigra plants. Salicylic acid (SA) level increased for 1 hr of interaction and reduced later. Development of systemic acquired resistance (SAR) was found restricted in plants infected with L. theobromae due to deficiency of SA, which is a major signal for development of SAR. Exogenously supplied SA did develop resistance and plant death was delayed. It was hypothesized that deficiency of SA could be due to jasmonic acid produced by fungus that inhibits SA biosynthesis. PMID- 15274492 TI - Evaluation of immunomodulatory activity of Suvarnamalini vasant, a generic Ayurvedic herbomineral formulation. AB - Suvarnamalini vasant-a generic Ayurvedic herbomineral preparation was studied for its immunomodulatory activity by 1) evaluating it's effect on phagocytic function of polymorphonuclear white blood cells of rats and 2) it's effect in E.coli induced peritonitis in albino mice. Pretreatment of rats with Suvarnamalini vasant improved the phagocytic function of polymorphonuclear white blood cells and also protected mice against E.coli-induced peritonitis. The results indicate the potential of Suvarnamalini vasant as an immunomodulator. PMID- 15274493 TI - Sex differences in oxidative stress induced by benzene in rats. AB - Role of sex differences on oxidative stress induced by benzene has been studied in liver, kidney and lungs of rat. It was observed that benzene administration enhanced lipid peroxidation in liver, kidney and lungs of rat, nevertheless, significant variations were recorded in male and female rats. Decrease of GSH and CYTP(450)2E1 was higher in female rats than male rats except lungs. The results suggest that oxidative stress induced by benzene is higher in female rats. PMID- 15274494 TI - In vivo enhancement of nucleopolyhedrovirus of oriental Armyworm, Mythimna separata using spindles from Helicoverpa armigera entomopoxvirus. AB - When the third instar larvae of M. separata were exposed to eight varying concentrations of polyhedral occlusion bodies (POB's) of nucleopolyhedrovirus of M. separata (MsNPV) ranging from 2.6 x 10(-1) to 2.6 x 10(8) POB's/ml, the percent mortality and incubation period ranged from 16-100% and 14 to 9 days respectively. On the other hand when the same third instar larvae of M. separata were exposed to only five varying concentration of POB's of MsNPV ranging from 2.6 x 10(2) to 2.6 x 10(6), POB's/ml along with a constant dose of entomopox viral spindles from Helicoverpa armigera, the per cent mortality ranged from 63 to 100% with reduction in incubation period from 7 to 4 days respectively. The enhancement index (log10) of the virus was 2.76 or reduction of more than 500 times in LC50. The ability and the mechanism of the spindles from H. armigera entomopoxvirus to enhance the infectivity of MsNPV has been discussed. PMID- 15274495 TI - Some comments on the nature and use of the concept of psyche in psychoanalysis and psychodynamic psychotherapy. AB - The term psyche is central to the field of mental health and dysfunction. And yet the term is seldom talked about directly, and is seldom elaborated by the professionals who function under its rubric. This article elaborates this term, and sets forth a definition for the purposes of psychology and psychodynamics as science in the sense of the establishment of psychological knowledge. Psyche is defined in this article as a hypothetical construct, as outlined in the classic article in the theory of psychological science by MacCorquodale and Meehl (1948). Psyche is to be distinguished from the concept of mind, which is identified here as a strictly phenomenological (subjective) concept. Use of psyche in this way allows it to be utilized in descriptions of its dynamics (psychodynamics), as a central concept in a science which utilizes phenomenological (subjective rather than objective) data. PMID- 15274496 TI - Orthodox Judaism and psychoanalysis: toward dialogue and reconciliation. AB - The authors examine the conflicted relationship between Orthodox Judaism and psychoanalysis. Orthodox Jewish thinkers about psychology have responded to psychoanalysis as incompatible with the practice of Orthodox Judaism. On the other hand, those psychoanalytic writers who have examined the beliefs and practices of Orthodox Jews have tended to treat these issues in a reductionistic fashion. However, the authors find possibilities for reconciliation and dialogue in the work of Aaron Rabinowitz and Moshe Halevi Spero. PMID- 15274497 TI - Psychoanalytic supportive psychotherapy of a terrified communist: report of a 37 year treatment. AB - This is a case report of a 37-year treatment of a victim of the Holocaust. It illustrates the use of psychoanalytic based supportive psychotherapy to enable a severely damaged person to pull himself together and live a respectable life. The case is of special interest because a careful evaluation of the patient's ego functioning was important all through the treatment, and revealed an unusual set of defences. He was able to deal with his rage by taking an extreme Maoist Communist political position, ranting and raving about it, including his hate of America, etc., but at the same time he encapsulated this and focused it in the psychotherapy sessions. In contrast to many such fanatical individuals, he was able to keep himself from acting out these political feelings in any way or to engage in any destructive behavior. This seemed to form an intermediate position that enabled him to function and adapt to our society, get married, find long term respectable employment, and finally retire with savings and a pension. Understanding these ego operations empathically made it possible for the author to tolerate his views without either challenging them or agreeing with them and enabled him to come regularly and utilize supportive psychotherapy. PMID- 15274498 TI - Women's internal reality: birthplace or graveyard. AB - This article is an exploration of the etiology of female subjectivity and its relationship to unconscious and conscious images of mother and mother's body. Within the analytic process in dream and phantasy internal aspects of daughter's subjective beliefs and feelings about her body, sexuality, pregnancy, maternal capacity, and mental strength emerge. An analysis of their transformations into language, conscious beliefs, and cultural biases are juxtaposed with the development of women's subjectivity. The concept of the "illness of the signifier" is shown to be the outcome of the painful difficulties of realizing the meaning of the maternal presence as they are constructed internally and culturally. PMID- 15274499 TI - Post-war syndromes: illustrating the impact of the social psyche on notions of risk, responsibility, reason, and remedy. AB - The 20th century offered many examples of post-war syndromes such as Da Costa's syndrome, irritable heart, shell shock, effort syndrome, medical evacuation syndrome, post-traumatic stress disorder, and Gulf War syndrome. These post-war syndromes occur under conditions of substantial medical and scientific uncertainty, conditions that potentially magnify the impact of social context on clinical care for these syndromes. This article reviews the social circumstances surrounding four post-war syndromes. The case is made that social context has significantly impacted professional and lay perceptions of causal mediators, relevant risk factors, defining symptoms, and appropriate therapies for these syndromes. Furthermore, it is argued that social context influences what parties are held responsible for post-war syndromes, and what clinical disciplines are ultimately deemed appropriate to provide legitimate post-war illness care. PMID- 15274500 TI - Using dreams to assess clinical change during treatment. AB - This article describes several studies that examine the relationship between the manifest content of selected dreams reported by patients and their clinical progress during psychoanalytic and psychodynamically oriented treatment. There are a number of elements that dreaming and psychotherapy have in common: affect regulation; conflict resolution; problem-solving; self-awareness; mastery and adaptation. Four different studies examined the relationship between the manifest content of selected dreams and clinical progress during treatment. In each study, the ratings of manifest content and clinical progress by independent observers were rank-ordered and compared. In three of the four studies there was a significant correlation between the rankings of manifest content and the rankings of clinical progress. This finding suggests that the manifest content of dreams can be used as an independent variable to assess clinical progress during psychoanalytic and psychodynamically oriented treatment. PMID- 15274501 TI - Elements of dynamics II: psychodynamic prescribing. AB - Scientific pharmacotherapy, as it should be, is based upon the results of aggregates of patient responses to treatments of specific target conditions for drugs. Although enduring personality traits are increasingly included as targets for pharmacotherapy, in the real world of psychiatric practice transference issues and the patient's character or personality traits play a larger role in the selection, dosage, tolerability, and outcome of treatment than is generally recognized or admitted, other than to attribute undifferentiated placebo effects. Transference and countertransference issues from the marketplace are described, as well as operational parameters of character, character-based decision making, and specific suggestions for medicating according to character types. A new field of psychodynamic pharmacotherapy is encouraged along with attention to culturally informed prescribing. PMID- 15274502 TI - Interview criteria for assessing allegations of sexual abuse in children and adults. PMID- 15274503 TI - Psychotherapy of schizophrenia. PMID- 15274504 TI - Intrinsic opposition between the scientific nature of psychoanalytic inquiry and the nature of religious belief. PMID- 15274505 TI - Case management for uninsured cuts length of stay, readmissions. PMID- 15274506 TI - Communities must tackle care for the uninsured. PMID- 15274507 TI - System links providers throughout the continuum. PMID- 15274508 TI - Electronic patient records result in efficiencies. PMID- 15274509 TI - CM redesign: how one organization tackled the process. PMID- 15274510 TI - Self-pay focus overhauls financial counseling. PMID- 15274511 TI - Report card provides staff feedback at a glance. PMID- 15274512 TI - The Ottorino Rossi Award 2004. PMID- 15274513 TI - Rational basis of rehabilitation following cerebral lesions: a review of the concept of cerebral plasticity. AB - Improvement and recovery of function after a cerebral lesion imply a reorganization of the central nervous system. This capacity of the nervous system to adapt its structural organization is referred to as cerebral plasticity. This paper summarizes the biochemical and histological basis of cerebral plasticity, reviews the modalities of brain reorganization following amputations and cerebral lesions, and concludes with a brief review of the most salient aspects of rehabilitation of memory, language, executive functions and dementia. PMID- 15274514 TI - Cluster headache: the history of the Cluster Club and a review of recent clinical research. AB - In September 2003, a scientific meeting was held in Rome to revive the International Cluster Headache Research Group (or "Cluster Club") tradition. This group of specialists was originally formed in the late 1970s by Ottar Sjaastad in order to promote research ideas, and to generate papers and other important information in this field. Its meetings, the last of which had taken place in 1994, had been informal events at which there was ample time for lively discussion. The last decade of the 20th century brought a significant increase in clinical and experimental research into cluster headache (CH), and this review summarizes some of the results of this research. The male preponderance of CH has been shown to be progressively decreasing over the years. Revised clinical criteria and a modern classification have been presented. First-degree relatives of probands with CH have been shown to have an increased risk of suffering from CH compared with the general population. Genetic analysis suggests that an autosomal dominant gene plays a role in some families. Functional neuroimaging has contributed to a better understanding of the pathophysiology of the condition. Positron emission tomography during provoked attacks has shown activation of the ipsilateral inferior posterior hypothalamus and it has been suggested that CH might be a functional neurovascular disorder of pacemaker or circadian regions in the hypothalamic grey matter. Subcutaneously administered sumatriptan has emerged as a highly effective acute treatment, but, in our opinion, the emphasis should be on attack prevention. Deep brain stimulation of the inferior posterior hypothalamic grey matter seems to be very promising as a novel treatment targeting the presumed central origin of pain attacks. PMID- 15274515 TI - A questionnaire on sleep and mental disorders in Parkinson's disease (QSMDPD): development and application of a new screening tool. AB - Psychiatric, cognitive and sleep disorders are the most frequent and disabling non-motor complications of Parkinson's disease (PD). To improve the description of sleep and mental disorders in PD patients, we set out to develop a simple and reliable data collection tool (questionnaire) for the screening of large samples of PD patients. The first draft of the questionnaire was administered to a consecutive series of 120 PD patients from the outpatient department of our unit, who were instructed to fill it in with the help of their caregivers. Subsequent drafts of the questionnaire were evaluated together with the patients and their caregivers, until a final, satisfactory version was obtained. This final version was named the Questionnaire on Sleep and Mental Disorders in PD (QSMDPD). This questionnaire--we used the Italian version, named Questionario sui Disturbi del Sonno e Mentali nella Malattia di Parkinson, ODSMMP--consists of 119 questions with multiple-choice answers. The QSMDPD was mailed or handed to 400 PD patients followed at our unit's outpatient department. Three hundred and twenty (80%) were returned to us. A review of these completed questionnaires, conducted by a neurologist together with the patients, showed 90% of them (289) to be complete and to provide reliable data. This high compliance suggests that the QSMDPD is a promising tool for collecting data on sleep and mental disorders in large samples of PD patients. A short version will be administered as a follow-up tool. PMID- 15274516 TI - Relationship between clinical variables and cognitive performances in migraineurs with and without aura. AB - Conflicting data on cognitive defects in migraine could be explained by differences in the clinical variables of the populations studied. We investigated 21 patients with migraine with aura and 24 with migraine without aura, diagnosed according to the International Headache Society criteria. The patients were submitted to a comprehensive battery of neuropsychological tests and grouped according to attack frequency and side of pain. Attack frequency was not associated with significant differences in any of the tasks, while location of pain was found to be significantly related to poorer performance on both the immediate and delayed recall of Rey Complex Figure in migraineurs both with and without aura, and a significant relationship between side of pain and number of clusters in the second trial of California Verbal Learning Test was found only in migraine with aura patients. The finding of worse performances in patients with right-sided pain seems to support a right hemisphere dysfunction hypothesis. PMID- 15274517 TI - Impairment of executive functions in a patient with a focal lesion in the anterior cingulate cortex. Evidence from neuropsychological assessment. AB - Patients with naturally occurring lesions involving the anterior cingulate cortex are rare and there thus exist very few reports of focal lesions in this area. We report a longitudinal study of a new case of selective anterior cingulate damage due to the presence of an angiocavernoma at the junction of the anterior third with the middle third of the right gyrus cinguli. Before surgery, the results of several, different tests suggested a significant impairment of executive functions, including deficits in planning, monitoring of ongoing behavior, and strategy shifting, as well as an exaggerated susceptibility to retroactive interference. Most of these symptoms disappeared completely or almost completely after the surgical removal of the angiocavernoma, although exaggerated susceptibility to interference was found to persist four months after surgery. PMID- 15274518 TI - Integrative and facilitatory processes in premotor pathology and their application in training. AB - The role of the neurologist in rehabilitation goes beyond diagnosing the patient's disease and related functional disorders. This task, like dynamic diagnosis or the longitudinal evaluation of disease course and of the effects of treatments administered, can be carried out through consultations. In reality, the neurologist's direct intervention is required and should be oriented in three complementary directions. The role of the neurologist should be: 1) to define therapeutic interventions, both integrative and facilitatory training of the still preserved functions; 2) to identify patients in whom effective intervention is possible; 3) to make provision for methods and instruments designed, in both the exploratory and the therapeutic stages, to maintain the efficiency of, and to improve, the task-independent central functions. Motor pathology, whether of central origin secondary to pyramidal system damage, or of peripheral origin due to motor unit damage, falls within the field of task-specific executive processes. In the first instance, in which it is the first motor neuron that is affected, the most feasible intervention, in both an extensive and an intensive sense, is positional feedback and electrical stimulation (PFST) to prevent disuse of the short-range integrative and facilitatory functions, at the level of the praxic engram centres. In the second instance, in which it is the second motor neuron that is affected, the intervention is based mainly on classic physiokinesitherapy, in its various forms. The sector of interest to this symposium is that of premotor pathology, and its three main systems: initiation of movement, impairment of which gives rise to akinetic and dyskinetic syndromes; coordination of movement with inhibition of the antagonist muscles, impairment of which gives rise to primary ataxic syndromes: and spatial and temporal programming, impairment of which leads to various forms of apraxia. Furthermore, these three systems are associated, even at a preliminary level, with a structural resource and with two general mechanisms: mnesic memory stores and their coding and decoding function, impairment of which gives rise to phrenasthenic and dementia syndromes; vigilance mechanisms, impairment of which leads to confusional states; and purposeful attention mechanisms, whose impairment leads to dissociative mental states. Having identified the patients in whom integrative and facilitatory functions should be investigated, it remains to define the anatomical-functional substrate of these functions. PMID- 15274519 TI - Patient outcomes and measures: special considerations. PMID- 15274520 TI - Evaluation of the reliability and validity of nursing outcomes classification patient outcomes and measures. AB - One hundred sixty-nine of the Nursing Outcomes Classification (NOC) patient outcomes were tested for interrater reliability, criterion validity, and sensitivity. In 10 field sites, ranging from hospitals to home care, pairs of nurses rated the outcome measures for 5 to 130 patients. Inter-class correlations were greater than or equal to 0.70 for 63 outcomes. Pearson's correlations with criterion measures were greater than or equal to 0.60 for 40 outcomes and from 0.39 to 0.60 for 43 additional ones. Change scores for 99 outcomes ranged from 0 to 2.0 from first to second and second to third rating. Most NOC measures demonstrated good inter-rater reliability, substantial criterion validity, and sensitivity to change. More testing and thorough training of nurses using NOC outcomes are needed. PMID- 15274521 TI - Evaluation of the sensitivity and use of the nursing outcomes classification. AB - The two purposes of the study were evaluation of the sensitivity of Nursing Outcomes Classification (NOC) outcomes to change in patient condition and description of the outcomes and interventions nurses select for patients with a specific nursing diagnosis. Data were collected in 10 clinical sites representing the care continuum. Sensitivity was evaluated by comparing paired serial ratings of the means of NOC outcomes at baseline and subsequent intervals, including discharge. Descriptive statistics were used to illustrate linkages between the outcomes and interventions selected for each diagnosis. There were 4,233 paired serial ratings on 166 NOC outcomes of which 804 included at least two follow-up ratings. The average change ranged from 0.00 to 1.47 on a 5-point scale. The 10 most frequently selected diagnoses varied across sites, as did outcomes and interventions selected for each diagnosis. The findings provide evidence of the sensitivity of NOC in the aggregate and the value of standardized terminology for studying nursing practice. PMID- 15274522 TI - Assessing the reliability, validity, and sensitivity of nursing outcomes classification in home care settings. AB - The purpose of this study is to provide evidence of the validity, inter-rater reliability, and sensitivity of 36 clinically useful of Nursing Outcomes Classification (NOC) results for Home Care (HC) settings. The results of inter rater reliability, criterion-related validity, and sensitivity evaluations of 36 NOC outcomes were compiled from a 10-site regional evaluation of the NOC. Findings of HC and all sites data were contrasted. More than 90% of the inter rater reliability scores on the 36 NOC outcome label ratings were within one point, with a substantial number of absolute agreements. All but six of the criterion measures were significantly associated with the corresponding NOC outcomes, and most mean change scores were zero or positive. This was impressive evidence of the adequacy of these measures for reliable and valid use in practice. PMID- 15274523 TI - Toward integrating a common nursing data set in home care to facilitate monitoring outcomes across settings. AB - The purpose of our research is to identify a realistic subset of North American Nursing Diagnosis Association (NANDA), Nursing Outcome Classification (NOC), and Nursing Interventions Classification (NIC) terms specific to the home care (HC) setting. A subset of 89 NOC outcomes were identified for study in HC through a baseline survey. Three research assistants then observed the care of 258 patients to whom the 89 NOC outcomes applied and recorded the associated NANDA and NIC terms. Follow-up surveys and focus groups were conducted with the nurses and research assistants. There were 81 different NANDA and 226 NIC labels used to describe study patients' care. Only 36 of the 89 NOC labels studied were deemed clinically useful for HC. We found that expert opinion about terminology usage before actual experience under practice conditions is unreliable. PMID- 15274524 TI - Testing the nursing outcomes classification in three clinical units in a community hospital. AB - The testing of the Nursing Outcomes Classification (NOC) was the focus of a 4 year study to evaluate the use of the outcomes and measurement scales developed by the Iowa Outcomes Project, a research team at the University of Iowa. Three units in a Midwest community hospital collected data as part of the larger (ten) clinical site study to test the reliability, validity, and sensitivity of the NOC. This article focuses on the results of sensitivity testing obtained in a birth center, behavioral health center, and an oncology unit in a midwestern community hospital. Methods used in this study focused on change scores from initial assessment to post-treatment status for the outcomes studied in each unit. Average baseline ratings, average follow-up ratings, average change scores, and range of change are reported. Thirty-five outcomes are reported for the behavioral health unit, 21 outcomes are reported for the Birth Center, and 8 outcomes for the Oncology Unit. The overall average baseline for the behavioral health unit was 1.89 with an average follow-up rating of 3.22. For the Birth Center, the average baseline rating was 3.23 with an average follow-up score of 3.88. For the Oncology Unit, the average baseline score was 3.01 with an average follow-up rating of 3.12. The results of this study suggest that the NOC outcomes are able to identify change in some outcome ratings through time and in a direction expected for the populations studied in these three specialty units. PMID- 15274525 TI - Establishing competency in the use of North American Nursing Diagnosis Association, Nursing Outcomes Classification, and Nursing Interventions Classification terminology. AB - The purpose of this study was to evaluate a 16-hour intervention designed to build clinician competency in the use of North American Nursing Diagnosis Association (NANDA), Nursing Outcome Classification (NOC), and Nursing Interventions Classification (NIC) (hereinafter: N3) among nurses with limited N3 knowledge. Each of 19 pairs of nurses independently selected N3 terms and rated the outcomes applicable to an actual patient for a specified time. A pair-through discussion then created a single consensus patient profile of the applicable terms. Before discussion, pairs agreed on 46% of the NANDA diagnoses, 30% of the NOC outcomes, and 20% of the NIC interventions selected. Eighty-nine percent of NOC label pair ratings were within 1 point. Building competency in N3 requires consistent use in written and oral communication with peers across time. Inter rater reliabilities (IRRs) for NOC label ratings support previous findings. PMID- 15274526 TI - Prevalence and moderators of terror-related post-traumatic stress disorder symptoms in Israeli citizens. AB - BACKGROUND: A major psychological sequel of terrorist attacks is post-traumatic stress disorder. The relation between certain psychological factors specific to terrorist attacks (e.g., perceived control attributed to oneself/to the military, anticipated duration of terrorism) and PTSD symptoms have not been examined. OBJECTIVE: To examine the prevalence, correlates and moderators of PTSD-like symptoms following terrorist attacks in Israel. METHODS: Soon after a long wave of terrorist attacks in Israel in 2002, a convenience sample of 149 Israelis from five cities was assessed for terrorist attack exposure, perceived control, control attributed to the govemment/military, anticipated duration of the terrorism wave (predictability), and frequency of listening to the news. PTSD like symptoms were assessed with a brief self-report scale. RESULTS: We found that 15.4% of the sample was directly exposed to a terrorist attack and 36.5% knew someone dose who had been exposed to an attack. "Clinically significant" PTSD-like symptoms were reported by 10.1% of the sample. Correlates of PTSD-like symptoms were: perceived control in men, government control, and education in women (all inversely correlated to PTSD symptoms), and news-listening frequency in women (positively correlated to PTSD symptoms). PTSD-like symptoms were attenuated by the ability to predict the duration of the terrorism wave only in citizens exposed to an attack, and by perceived government control only among citizens listening infrequently to the news. CONCLUSIONS: This study revealed that approximately 10% of Israelis in our sample had relatively frequent PTSD like symptoms. Correlates of PTSD-like symptoms differed between men and women, and moderator effects were found. These findings reveal additional moderators that may have implications for treating PTSD following terrorist attacks. PMID- 15274527 TI - Hyperthermia combined with radiation therapy in the treatment of local recurrent breast cancer. AB - BACKGROUND: Hyperthermia combined with radiation therapy was shown to be more effective in local recurrent breast cancer than radiotherapy alone, but its use is limited due to technical difficulties, stringent reimbursement policies and because it is time consuming. OBJECTIVES: To report our experience with a simple and convenient XRT + HT delivery system. METHODS: XRT was delivered through either electron or photon beams (total dose 30-40 Gy in previously irradiated fields or 50-70 Gy in non-irradiated fields). Hyperthermia was delivered by a dedicated HT device operating at 915 MHz. The heating session lasted 45 minutes. The maximal tumor surface temperature was set at 45 degrees C and modified according to patient comfort. No intratumoral (invasive) thermometry was used. At least two HT sessions were scheduled to each HT field during the entire XRT treatment period. Tumor response was evaluated every 3 months after completion of treatment. The overall survival was measured from XRT + HT initiation until the last follow-up. RESULTS: Fifteen women underwent 114 HT treatments delivered through 28 HT fields. Twenty-four HT fields (15 patients) were previously irradiated. There was complete infield response in 10 fields (6 patients), partial response in 8 fields (4 patients), no response or progressive disease in 4 fields (3 patients), and no parameters in 6 fields (5 patients). Eighteen fields (64%) had complete or partial response. Seven patients had outfield recurrence despite wide XRT + HT fields. Ulceration was the only major side effect (three patients, three fields). CONCLUSIONS: The combined HT+XRT delivery system, with no invasive thermometry, is a simple and effective method for treating local recurrent breast cancer. PMID- 15274528 TI - Does acetylcholinesterase inhibition affect catecholamine secretion by adrenomedullary cells? AB - BACKGROUND: Splanchnic nerve stimulation evokes adrenomedullary catecholamine secretion via acetylcholine release and occupation of nicotinic cholinergic receptors on chromaffin cells. OBJECTIVES: To assess whether among cultured adrenomedullary cells there exists a population that tonically secretes acetylcholine. If so, then blockade of enzymatic breakdown of acetylcholine by addition of a cholinesterase inhibitor to the medium would increase occupation of nicotinic receptors by endogenous acetylcholine and thereby induce catecholamine release. METHODS: Primary cultures of bovine adrenomedullary cells in 24-well plates (1 million cells per well) were incubated after 48-72 hours with fresh incubation medium (control), medium with added secretagogues (nicotine, angiotensin II, or K+) or the acetylcholinesterase inhibitor, edrophonium (10(-7) to 10(-3) M), for 1-20 minutes. Fractional release rates of epinephrine, norepinephrine and dopamine were compared to a control. We also examined whether coincubation with edrophonium enhanced the effects of the secretagogues. All experiments were performed in quadruplicate and repeated three times. RESULTS: Nicotine, angiotensin II, and K+ each elicited time-related release of epinephrine, norepinephrine and dopamine by up to fourfold compared to the control. At all tested concentrations, edrophonium had no such effect. Co incubation with edrophonium also failed to augment the secretory responses to nicotine, angiotensin II, or K+. CONCLUSION: Bovine adrenomedullary cells in primary culture do not include a population of tonically active cholinergic cells. PMID- 15274529 TI - Persistent anemia in otherwise asymptomatic severe aortic stenosis: a possible indication for valve replacement? AB - BACKGROUND: The indication for aortic valve replacement in patients with significant aortic stenosis is symptomatology. Aortic stenosis may be associated with bleeding from colonic angiodysplasia, resulting in anemia. Persistent anemia in such patients, despite lack of an identifiable source of bleeding, is not considered an indication for valve replacement. OBJECTIVES: To report our experience with two elderly female patients who suffered from severe asymptomatic aortic stenosis, low levels of large von Willebrand factor multimer (10% and 5% respectively) and persistent anemia requiring multiple blood transfusions. METHODS: Both patients underwent an intensive work-up, but a source of bleeding could not be identified. Aortic valve replacement was performed in both patients. RESULTS: Aortic valve replacement abolished the need for further blood transfusions during a follow-up period of 20 months with normalization of the vWF multimer level (20% and 30% respectively). CONCLUSION: We suggest that aortic valve replacement be considered in selected patients with severe, otherwise asymptomatic aortic stenosis, who suffer from persistent anemia requiring multiple blood transfusions, lack an identifiable source of bleeding and have low levels of large vWF multimers. PMID- 15274530 TI - Differences in infant mortality rates between Jews and Arabs in Israel, 1975 2000. AB - BACKGROUND: The infant mortality rate is a health status indicator. OBJECTIVES: To analyze the differences in infant mortality rates between Jews and Arabs in Israel between 1975 and 2000. METHODS: Data were used from the Central Bureau of Statistics and the Department of Mother, Child and Adolescent Health in the Ministry of Health. RESULTS: The IMR in 2000 was 8.6 per 1,000 live births in the Israeli Arab population as compared to 4.0 in the Jewish population. Between 1970 and 2000 the IMR decreased by 78% among Moslems, 82% among Druze, and 88% among Christians, as compared to 79% in the Jewish population. In 2000, in the Arab population, 40% of all infant deaths were caused by congenital malformations and 29% by prematurity, compared to 23% and 53%, respectively, in the Jewish population. Between 1970 and 2000 the rate of congenital malformations declined in both the Arab and Jewish populations. In the 1970s the rate was 1.4 times higher in the Arab community than in the Jewish community, and in 2000 it was 3.7 times higher. CONCLUSION: As in the Jewish population, the IMR in the Arab community has decreased over the years, although it is still much higher than that in the Jewish community. Much remains to be done to reduce the incidence of congenital malformations among Arabs, since this is the main cause of the high IMR in this population. PMID- 15274531 TI - Changes over years in gross motor function of 3-8 year old children with cerebral palsy: using the Gross Motor Function Measure (GMFM-88). AB - BACKGROUND: In the developing child the nervous system undergoes a maturation process. The development and organization of any motor ability is the naturally adopted preference among the possibilities and constraints. The motor behavior of children with cerebral palsy is a personal automatic preference based on such constraints. One of the clinical measures designed for measuring the function of children with CP is the Gross Motor Function Measure. Motor development curves for children with CP have been established based on the GMFM instrument and Gross Motor Function Classification System. OBJECTIVES: To examine the change over time in gross motor function for children with CP attending a special education school for handicapped children in Israel. METHOD: We conducted a retrospective review of the medical records of children at various ages and with varying degrees of severity who were being treated by a multidisciplinary team. The study population comprised 106 children aged 3-8 years with CP who were attending the school of special education at Assaf Harofeh Medical Center. The GMFM-88 test was performed annually for the study children over a 7 year period (1995-2001). RESULTS: During the study period the GMFM scores improved significantly. The rate of improvement and top achievements over the years differed according to the severity of the motor impairment. The gross motor development reached a plateau at the age of 6-7 years. CONCLUSIONS: The changes in gross motor development of the study population were similar to the profile of changes in the developmental process of children who develop normally. The nature of the curves of gross motor change for the children with CP should be borne in mind when designing individual treatment goals and strategies for a child. PMID- 15274532 TI - Gadolinium-based magnetic resonance imaging contrast agents in interventional radiology. AB - Gadolinium-based agents are widely used in magnetic resonance imaging as contrast agents. These agents are radio-opaque enough for diagnostic imaging of the vascular tree by using digitally subtracted images as well as for imaging of the biliary system and the urinary tract. The recommended doses for gadolinium do not impair renal function or cause adverse reactions in patients with iodine sensitivity; thus patients with such conditions can safely undergo diagnostic angiography, either by MRI angiography or by catheterization using gadolinium as contrast agent, for diagnostic and therapeutic purposes. PMID- 15274533 TI - Chemotherapy in gastric cancer: a brief chronicle with emphasis on recent developments. PMID- 15274534 TI - Immunologic aspects of protein degradation by the ubiquitin-proteasome system. AB - The ubiquitin-proteasome pathway has a central role in selective degradation of intracellular proteins. Among the key proteins degraded by the system are those involved in the control of inflammation, cell cycle regulation and gene expression. With numerous important cellular pathways affected, derangements in the ubiquitin system were shown to result in a variety of human diseases including malignancies, neurodegenerative diseases and hereditary syndromes, and proteasome inhibition was implicated as a potential treatment for cancer and inflammatory conditions. Two proteasome inhibitors are currently under clinical evaluation for multiple myeloma and acute ischemic stroke. The ubiquitin system also has an important function in the immune and inflammatory response. It is involved in antigen processing and presentation to cytotoxic T cells, and the activation of nuclear factor-kappa B--the central transcription factor of the immune system. Since the proteasome is the central source of antigenic peptides that are presented to the immune system, some viruses, such as the Epstein-Barr virus, developed escape mechanisms that manipulate the ubiquitin-proteasome system in order to persist in the infected host. Understanding the mechanisms underlying the production of viral antigens by the ubiquitin-proteasome system may have therapeutic applications such as future development of vaccines. PMID- 15274535 TI - The ripple effect of the toll of terror. PMID- 15274536 TI - Can it be TB? PMID- 15274537 TI - Vaccination may be associated with autoimmune diseases. PMID- 15274538 TI - Vaccination and autoimmune diseases: the argument against. PMID- 15274539 TI - Spinal tuberculosis with paraplegia in pregnancy. PMID- 15274540 TI - Cervical Pott's disease presenting as a retropharyngeal abscess. PMID- 15274541 TI - Who is the owner of this glove? PMID- 15274542 TI - Acute subclavian and brachial artery thrombosis as a complication of the nephrotic syndrome. PMID- 15274543 TI - Transection of the ulnar nerve as a complication of two-portal endoscopic carpal tunnel release. PMID- 15274544 TI - Treatment of hypertension by renal artery stenting. PMID- 15274545 TI - The 24th European Workshop for Rheumatology Research. Berlin, 26-29 February 2004. PMID- 15274546 TI - Experimental validation of an affinity energy distribution calculated with the expectation maximization method. AB - The difference between the average energies of the high-energy modes of the adsorption energy distributions of (S)-alprenolol and (R)-alprenolol on a chiral stationary phase calculated by the expectation maximization method agree well with the difference between the adsorption energies of these two compounds measured by isothermal titration calorimetry. PMID- 15274547 TI - Epitaxial relationships between uric acid crystals and mineral surfaces: a factor in urinary stone formation. AB - Uric acid (C5H4N4O3) is one of the final products of purine metabolism. Its concentration balance is maintained in the kidneys, but compromised kidney function can result in its crystallization either in the renal tract or in the interstitial fluid of joints. In physiological deposits, crystalline uric acid is most frequently found either in a protonated state (anhydrous or dihydrate phases) or as a deprotonated urate ion (sodium or ammonium salts). Often these precipitates are found in association with a number of mineral phases (e.g., calcium oxalates, calcium phosphates, and magnesium phosphates). Their frequent and common coexistence suggests that synergistic relationships between these crystalline phases may exist. A comprehensive list of different heterogeneous uric acid/uric acid and uric acid/mineral interfaces that are epitaxially matched was generated with the lattice-matching program EpiCalc. Two hundred twenty-five coincident epitaxial matches and four commensurate epitaxial matches were identified using this screening procedure. PMID- 15274548 TI - Liquid crystalline gel with refractive index gradient of curdlan. AB - Curdlan dissolved in aqueous sodium hydroxide was dialyzed to aqueous calcium chloride to form a gel. Transparent and turbid concentric layers observed in the gel cross section perpendicular to the long axis of the dialysis tube were identified as liquid crystalline gels with refractive index gradient and amorphous gels, respectively. The thickness of each layer was proportional to the diameter of the dialysis tube, and the gelation proceeded in proportion to the root of time. The unique pattern formation was attributed to the change of curdlan conformation and calcium-induced cross-linking resulting from a diffusion of calcium cations and hydroxide anions through the dialysis tube. It is suggested that the orderedness of the curdlan molecules decreases by the increase of the curvature of the concentric liquid crystal layers as the layer comes toward the center of the dialysis tube. PMID- 15274549 TI - Patterned dual pH-responsive core-shell hydrogels with controllable swelling kinetics and volumes. AB - Dual pH-responsive core-shell hydrogels containing both a vinyl pyridine component and a 2-dimethylaminoethyl methacrylate component were prepared using an in situ photopolymerization process. Complementary photomasks were utilized to prepare hydrogels with core/shell volume ratios of 2:1, 1:1, and 1:2. Depending on the location of each polymer component, dramatically different swelling profiles were achieved. Selective swelling of the shell followed by the core components allowed the hydrogel to expand with the usual kinetics; however, by switching the location of each polymer component and swelling the core first, swelling rates decreased by over 1 order of magnitude and were dependent on the shell component's volume. The ability to pattern core/shell volumes also provided the ability to fabricate hydrogels that possess a constant maximum diameter but different cutoff points between its first and its second transition volumes. These materials may be of interest for controlled release applications. PMID- 15274550 TI - Evaporation of liquids and solutions in confined geometry. AB - We describe a drying setup where a liquid droplet is confined between two circular glass plates and allowed to evaporate. Optical imaging of the shrinking droplet provides the temporal evolution of the solvent volume. For a pure liquid, we measure the diffusion coefficient of the evaporating gas with a good accuracy. For a solution, the slowing down of evaporation by progessive concentration of the solute yields a direct measurement of the solvent activity. PMID- 15274551 TI - IR reflection spectra of monolayer films sandwiched between two high refractive index materials. AB - Monolayer films adsorbed on substrates with high refractive indices such as metals or semiconductors yield strongly enhanced infrared reflection spectra when they are contacted with a transparent, high refractive index ambient medium and are probed with p-polarized light at high incidence angles. The sensitivity increase arises from the enhancement of the perpendicular electric field within a thin, low refractive index layer sandwiched between two high refractive index materials and gives rise to signal intensity gains up to 2 orders of magnitude in combination with an essentially exclusive detection of only perpendicular surface vibrations. Experimental spectra of ordered monolayer films of octadecanethiol on gold and of octadecylsiloxane on silicon in this sandwich configuration yield enhancement factors between 15 (on Si) and 30 (on gold) compared to conventional grazing incidence external reflection spectra and are governed by a common, simple surface selection rule, which allows immediate quantitative evaluation and comparison of the film structures on different substrates. PMID- 15274552 TI - Synthesis of amorphous silicon colloids by trisilane thermolysis in high temperature supercritical solvents. AB - Colloidal submicrometer-diameter amorphous silicon (a-Si) particles are synthesized with >90% yield by thermal decomposition of trisilane (Si3H8) in supercritical hexane at temperatures ranging from 400 to 500 degrees C and pressures up to 345 bar. A range of synthetic conditions was explored to optimize the quality of the product. Under the appropriate synthetic conditions, the colloids are spherical and unagglomerated. The colloids can be produced with average diameters ranging from 50 to 500 nm by manipulating the precursor concentration, temperature, and pressure. Relatively narrow particle size distributions, as measured by transmission electron microscopy (TEM), with standard deviations about the mean as low as approximately +/-10% could be obtained in some cases. We explored the thermal annealing of the amorphous silicon particles after isolation from the reactor and found that crystallization to diamond structure silicon occurred at temperatures as low as 650 degrees C. The amorphous and crystalline materials were characterized by X-ray diffraction and high resolution scanning and transmission electron microscopy. PMID- 15274553 TI - Surface friction of hydrogels with well-defined polyelectrolyte brushes. AB - Hydrogels of poly(2-hydroxyethyl methacrylate) (PHEMA) with well-defined polyelectrolyte brushes of poly(sodium 4-styrenesulfonate) (PNaSS) of various molecular weights were synthesized, keeping the distance between the polymer brushes constant at ca. 20 nm. The effect of polyelectrolyte brush length on the sliding friction against a glass plate, an electrorepulsive solid substrate, was investigated in water in a velocity range of 7.5 x 10(-5) to 7.5 x 10(-2) m/s. It is found that the presence of polymer brush can dramatically reduce the friction when the polymer brushes are short. With an increase in the length of the polymer brush, this drag reduction effect only works at a low sliding velocity, and the gel with long polymer brushes even shows a higher friction than that of a normal network gel at a high sliding velocity. The strong polymer length and sliding velocity dependence indicate a dynamic mechanism of the polymer brush effect. PMID- 15274554 TI - Magnetic beads as interfacial nanoprobes. AB - We use paramagnetic beads to probe strongly localized magnetic fields from one dimensional nanomagnets. Using a polarization microscope in reflection mode, we find that light reflected from beads exhibits intensity fluctuations which may help us understand Brownian motion near interfaces. We estimate the height fluctuations and femtonewton forces acting on the beads. PMID- 15274555 TI - A two-state model for selective solubilization of benzene-limonene mixtures in sodium dihexyl sulfosuccinate microemulsions. AB - When surfactants are used to solubilize oil, the oil to be solubilized is often a mixture of components with differing properties, for example, solubilization of drug molecules in microemulsion formulations, remediation of organic polluted aquifers using surfactants, and so forth. Previous research has demonstrated that selective solubilization of one organic component over the other may occur if the organic components are dissimilar. In this research, we investigated selective solubilization from benzene-limonene mixtures in Winsor type I and III microemulsion systems containing water, sodium di-n-hexyl sulfosuccinate, and NaCl. The effect of the oil phase composition and the electrolyte concentration on the selectivity was studied. It was found that the selectivity toward benzene was highest at low electrolyte and benzene concentrations, decreasing as the electrolyte or benzene concentration increased. The results are discussed on the basis of the two-state solubilization theory and by correlating the curvature of the surfactant film in the microemulsion with changes of the electrolyte concentration and the oil phase composition. A simple mathematical model is developed for the selectivity, which combines the two-state solubilization theory and the net-average curvature model of microemulsion solubilization to yield close agreement with the experimental data. PMID- 15274556 TI - First emulsion polymerization of styrene with sodium borohydride: evidence of the generation of radical intermediates by sodium borohydride in H2O. AB - Emulsion polymerization of styrene with sodium borohydride (NaBH4) in an aqueous sodium dodecyl sulfate (SDS) solution was successfully accomplished for the first time. Polystyrene with a high molecular weight (M(w) > 2 000 000) and a broad molecular weight distribution (MWD approximately 3.5) was obtained in a conversion of less than 30%. Several pieces of evidence that the polymerization proceeded through radical intermediates were observed. Variations in the concentration of NaBH4 showed a critical range in said concentration, i.e., a borderline that determined whether the main reaction was directed to either a polymerization or a competed reaction with variations in the NaBH4 level. Kinetic studies on the emulsion polymerization of styrene with NaBH4 performed at 50, 55, and 60 degrees C showed that the initiator had an approximately 50-min induction period. A plot of -ln(1 - X), where X is the fractional conversion, as a function of time resulted in a linear relationship, showing that the present initiator system followed first-order kinetics with respect to monomer concentration. The Arrhenius plot between ln k vs 1/T gave a good linear relation, and the overall activation energy was observed to be about 37.5 kcal/mol. The employment of CH3I with NaBH4 significantly increased conversion (>95%) and provided polystyrene with a well-controlled Mw and MWD (<2.3). PMID- 15274557 TI - Doxorubicin-poly(acrylic acid) complexes: interaction with liposomes. AB - Complexation of antitumor drug, doxorubicin (DOX), with poly(acrylic acid) (PAA) in buffer solutions was examined. The DOX-to-PAA binding was governed by electrostatic and stacking interactions resulting in a complex of characteristic composition with a PAA/DOX = 1.6 molar ratio. Sizes of the complex particles were found to lie in 600-900-nm range. However, the particles were able to interact with small neutral egg yolk lecithin liposomes (80-100 nm in diameter), a ternary DOX/PAA/liposome complex being formed. The observations and conclusions we made may be useful for interpreting biological effects of polymer-based bioactive constructs. PMID- 15274558 TI - One-component gels based on peptidic dendrimers: dendritic effects on materials properties. AB - This paper describes the gelation of symmetric dendrimers based on building blocks constructed from L-lysine. These dendrimers form gel-phase materials in nonpolar organic solvents. The thermal properties and concentration dependence of the gelation were investigated, and it was found that there was a clear dendritic effect on the behavior of the soft materials formed, with higher generation dendrimers giving rise to more thermally stable gels. Variable temperature 1H NMR studies indicated that this behavior was probably a consequence of more extensive interdendrimer hydrogen bonding occurring between the peptidic groups in the higher generation dendrimers. The supramolecular aggregates were found to have a fibrillar structure, with the dimensions and alignment of the fibers being dependent on dendritic generation. Circular dichroism measurements confirmed that these fibers possessed chiral organization of the peptidic groups on the supramolecular (nano) scale, assigned as helicity. This paper indicates that dendritic functionalization provides a useful way of tuning gel-phase materials properties, with clear dendritic effects on gel formation being quantified for the first time, hence illustrating the way dendritic functionalization can play a positive role in the formation of highly functional organic materials with desirable properties. PMID- 15274560 TI - Phase behavior and nano-emulsion formation by the phase inversion temperature method. AB - Formation of oil-in-water nano-emulsions has been studied in the water/C12E4/isohexadecane system by the phase inversion temperature emulsification method. Emulsification started at the corresponding hydrophilic lipophilic balance temperature, and then the samples were quickly cooled to 25 degrees C. The influence of phase behavior on nano-emulsion droplet size and stability has been studied. Droplet size was determined by dynamic light scattering, and nano-emulsion stability was assessed, measuring the variation of droplet size as a function of time. The results obtained showed that the smallest droplet sizes were produced in samples where the emulsification started in a bicontinuous microemulsion (D) phase region or in a two-phase region consisting of a microemulsion (D) and a liquid crystalline phase (L(alpha)). Although the breakdown process of nano-emulsions could be attributed to the oil transference from the smaller to the bigger droplets, the increase in instability found with the increase in surfactant concentration may be related to the higher surfactant excess, favoring the oil micellar transport between the emulsion droplets. PMID- 15274559 TI - Acceleration of nucleophilic attack on an organophosphorothioate neurotoxin, fenitrothion, by reactive counterion cationic micelles. Regioselectivity as a probe of substrate orientation within the micelle. AB - 31P NMR and UV-vis spectrometric evidence has revealed an unexpected regioselectivity in the reaction of fenitrothion, 1, an organophosphorus pesticide, with the cetyltrimethylammonium (CTA) surfactants CTAOH and CTAMINA, that incorporate the reactive counterions OH- and MINA- (the anti-pyruvaldehyde 1 oximate anion). While both micellar solutions accelerate decomposition of 1 compared to aqueous OH- alone, CTAMINA produced the largest rate enhancement (ca. 10(5)) at a pH (8.39) appropriate for environmental applications. In the absence of surfactant, reaction proceeds solely via the SN2(P) pathway. In the presence of surfactant but below the critical micelle concentration (cmc), a competitive SN2(C) pathway was observed in addition to SN2(P). Above the cmc, however, the CTAOH reaction again proceeded solely via the SN2(P) pathway while both pathways were operative with CTAMINA. The changes in reactivity and mechanistic pathway are discussed in terms of premicellar and micellar influences on rates and regioselectivity. A proposal that would account for the observed regioselectivity in the micellar system is that the aromatic ring and aliphatic side-chains of 1 are oriented toward the micellar interior, while the P=S moiety faces the aqueous pseudophase. PMID- 15274561 TI - Effect of polymer charge density and ionic strength on the formation of complexes between sodium arylamido-2-methyl-1-propane-sulfonate-co-acrylamide gels and cetylpyridinium chloride. AB - The effects of sodium chloride on the composition and structure of polyelectrolyte gel-surfactant complexes (PSCs) formed by the sodium salt of acrylamide-2-methyl-1-propane-sulfonic acid-co-acrylamide gels and cetylpyridinium chloride have been studied. At a low ionic strength of the solution, the composition of all the complexes is close to stoichiometric by charge. In the presence of 0.3 M sodium chloride, the composition of the complexes formed by the gel with 99 mol % charged groups is close to stoichiometric, while for the gel with 33 mol % charged monomer units, a nonstoichiometric complex with a high excess of the surfactant is formed. Further decrease of the charge density up to 10 mol % leads to partial or complete dissociation of the PSCs. The study of PSCs by the method of small-angle X-ray scattering (SAXS) shows that the complexes formed by the gels with high and intermediate charge densities are highly ordered. The decrease of the charge density of the swollen networks at first leads to a change in symmetry of the ordered domains in the PSCs and then to their disordering. The formation of nonstoichiometric PSCs at a high enough concentration of salt is explained by the effect of fitting, when the packing of the surfactant and polymer components in the PSCs is improved due to the inclusion of extra surfactant molecules together with their counterions in the ordered domains. PMID- 15274562 TI - Clay-vesicle interactions: fluorescence measurements and structural implications for slow release formulations of herbicides. AB - Clay-vesicle systems exhibit a potential for environmental applications, such as herbicide formulations for reduced leaching. Clay-vesicle interactions were addressed by combining adsorption and XRD measurements with fluorescence studies for didodecyldimethylammonium bromide (DDAB), dioctadecyldimethylammonium bromide (DDOB), and montmorillonite. XRD and adsorption data indicated that the adsorbing vesicles were transformed after 3 days into paraffinic and bilayer structures. Fluorescence studies revealed that adsorption was almost complete within 5 min for a loading below the cation exchange capacity (CEC). Aggregation and sedimentation of clay-surfactant particles occurred within several minutes. Fluorescent measurements of supernatants indicated decomposition of vesicles at a high clay/surfactant ratio due to rapidly adsorbing cationic monomers. The kinetics of energy transfer between vesicles labeled by NBD-PE (1,2-dipalmitoyl sn-glycero-3-phosphoethanolamine-N-(7-nitro-2-1,3-benzoxadiazol-4-yl)) and montmorillonite labeled by rhodamine-B follows that of aggregation of surfactant clay particles and structural changes of the vesicles at times of minutes to hours. Experiments following the reduction of NBD fluorescence by addition of dithionite indicate faster permeabilization of DDOB than DDAB vesicles, which was confirmed by leakage experiments. The faster permeabilization of DDOB vesicles in the presence of clay was correlated with their inferior suitability for the preparation of clay-based formulations of anionic herbicides for slow release. PMID- 15274563 TI - Spontaneously formed nonequilibrium vesicles of cetyltrimethylammonium bromide and sodium octyl sulfate in aqueous dispersions. AB - It is well-known that vesicles form in mixtures of cationic and anionic surfactants. We have investigated mixtures of cetyltrimethylammonium bromide (CTAB) and sodium octyl sulfate (SOS) with the latter in excess over a long time, about 500 days. We have followed the growth of the aggregates by light scattering and checked the morphologies by cryogenic transmission electron microscopy (cryoTEM). All samples showed a monotonic growth with decreasing rate (the change of size was about linear on a logarithmic time scale). In series of samples with weight ratio 30:70 of CTAB/SOS and total surfactant concentration between 0.5 and 3 wt %, the size increased with the surfactant concentration up to 2 wt % and decreased thereafter; cryoTEM examination revealed that the samples contained a majority of open bilayer structures at the highest concentrations. Part of the sample at 2 wt % was diluted to 0.5 wt % after 60 days. The size measured after dilution was slightly smaller than before but well above that found in the directly prepared 0.5 wt % sample, and the particle size in the three samples continued to grow in parallel. Structures other than unilamellar vesicles were observed also in samples at 2 wt % total surfactant concentration at CTAB/SOS ratios close to the borders of the vesicle lobe in the (quasi) ternary phase diagram as published (Yatcilla, M. T.; Herrington, K. L.; Brasher, L. L.; Kaler, E. W.; Chiruvolu, S.; Zasadzinski, J. A. J. Phys. Chem. 1996, 100, 5874). The results clearly show that the spontaneous vesicle populations do not represent equilibrium populations. They also suggest that the vesicle lobes in the phase diagram mainly represent areas where a lamellar phase is easily dispersed in the form of vesicles in an aqueous solution. PMID- 15274564 TI - Mixed vesicle formation on a ternary surfactant system: didodecyldimethylammonium bromide/dodecylethyldimethylammonium bromide/water. AB - The formation of mixed aggregates has been investigated on a ternary system consisting of two cationic surfactants with similar polar heads and two and/or one 12 carbon atom hydrophobic tail, respectively, didodecyldimethylammonium bromide and dodecylethyldimethylammonium bromide and water. The study has been carried out by means of conductivity, zeta potential, and cryogenic transmission electronic microscopy (cryo-TEM) experiments on the very diluted region. A variety of mixed aggregates, microaggregates, vesicles, and micelles has been found, depending on system composition and total surfactant concentration. Mixed critical microaggregate concentration and mixed critical vesicle concentration have been determined from conductivity data. Furthermore, zeta potential and cryo TEM experiments allow for the characterization of the aggregates/solution interface and of the shape and size of the aggregates. This experimental evidence has also been analyzed in terms of the theoretical packing parameter, P. PMID- 15274565 TI - Aqueous synthesis of alkanethiolate-protected Ag nanoparticles using Bunte salts. AB - The one-pot synthesis of monolayer-protected metal nanoparticles derived from sodium S-dodecylthiosulfate (Bunte salt) in aqueous solution is described. Silver nanoparticles, which were produced by the borohydride reduction of silver nitrate in H2O, were stabilized by the adsorption of S-dodecylthiosulfate followed by the removal of the SO3- moiety. Temporary stabilization of silver sols by the adsorption of borohydride and borate prevented aggregation of silver nanoparticles in H2O. The syntheses of other metal nanoparticles, including gold, copper, and palladium particles in H2O, were less successful. Gold and copper particles were completely aggregated and precipitated out immediately after the addition of NaBH4, yielding only insoluble clusters. Stable and soluble palladium nanoparticle could be prepared, but the presence of Pd-thiolate complex was also observed. These nanoparticles were characterized using 1H NMR, UV-vis spectroscopy, FT-IR spectroscopy, and transmission electron microscopy. PMID- 15274566 TI - Anion effect on mediated electron transfer through ferrocene-terminated self assembled monolayers. AB - Inorganic anions strongly influence the electron transfer rate from the ascorbate to the ferrocene-terminated self-assembled monolayer (SAM) composed of 9 mercaptononyl-5'-ferrocenylpentanoate (Fc(CH2)4COO(CH2)9SH, MNFcP). At the 1 M concentration level of the supporting anion (sodium salt electrolyte), a more than 10-fold increase in the electrocatalytic oxidation rate constant of the ascorbate is observed in the following sequence: PF6-, ClO4-, BF4-, NO3-, Cl-, SO4(2-), NH2SO3- (sulfamate), and F-. The sequence corresponds to the direction of increasing hydration energy of the corresponding anion, suggesting that highly hydrated ions promote electrocatalytic electron transfer to the ferrocene terminated SAMs, while poorly hydrated ions inhibit it. Fourier transform surface enhanced Raman spectroscopy (FT-SERS), in combination with cyclic voltammetry, indicates the formation of surface ion pairs between the ferricinium cation (Fc+) and low hydration energy anions, while, on the contrary, no ion pairs were observed in the electrolytes dominated by the high hydration energy anions. Though it is evident that the ion-pairing ability of hydrophobic anions is directly responsible for the electrocatalytic electron transfer inhibition, an estimate of the free, ion-unpaired Fc+ surface concentration shows that it cannot be directly related to the electron transfer rate. This suggests that the principal reason of the anion-induced electron transfer rate modulation might be related to the molecular level changes of the physical and chemical properties as well as the structure of the self-assembled monolayer. PMID- 15274567 TI - Electrocapillarity behavior of Au111 in SO4(2-) and F-. AB - We present the first set of results measuring the change in interfacial free energy and surface stress for Au(111) electrodes in an electrolyte containing a nonspecifically adsorbing anion and compare this behavior to that in an electrolyte containing an anion known to undergo specific adsorption. Generally, we find that the surface stress is more sensitive to changes in electrode potential and adsorption then the interfacial free energy. The results obtained in fluoride electrolytes are compared to the predictions of a thermodynamic analysis. PMID- 15274568 TI - Microstructured Au/TiO2 model catalyst systems. AB - The preparation of microstructured Au/TiO2 model catalysts as a first step toward micrometer-scale parallel studies on model catalysts and toward studies of mesoscopic effects in catalytic reactions was investigated by atomic force microscopy and X-ray photoelectron spectroscopy. The model systems, which consist of micrometer-size active areas covered with Au nanoparticles that are separated by similarly sized inactive areas free of Au particles, are fabricated by combining optical lithography methods for microstructuring and ultrahigh vacuum evaporation for Au nanoparticle deposition and by applying suitable cleaning steps. It is demonstrated that practically perfect microstructures with Au nanoparticles of catalytically relevant sizes (2-3-nm diameter) on a clean TiO2 substrate can be produced this way and that the processing steps do not affect the deposited Au nanoparticles, neither in size nor in lateral distribution. PMID- 15274569 TI - Evidence for gravity's influence on molecules at a solid-solution interface. AB - Yoshimoto et al. [Anal. Chem. 2002, 74, 4306-4309] reported that a quartz crystal microbalance or QCM changed its response to sucrose solutions according to its angle of immersion. The effect was tentatively attributed to gravity-caused stress on the viscous interface between the oscillator and the bulk solution. The present work reports results from QCM experiments carried out so that any effect of gravity on the interfacial region would be magnified. This permitted use of a lower-frequency, less-sensitive QCM. Molecules of DNA were tethered to a functionalized QCM surface and then extended in steps, via sandwich hybridization, to produce DNA of uniform and known length. This feature allowed both the effect of QCM immersion angle and the relationship between frequency and molecular length to be investigated simultaneously. Comparison of acoustic wave damping at 0 degrees and 180 degrees immersion angles offers compelling evidence that the interfacial region expands when the active face of the QCM is down and contracts when it is up. This is apparently a consequence of the interfacial region being more dense than the bulk solution. The results are consistent with (a) slow gravity-driven movement of molecules away from a down-facing QCM, (b) rapid hybridization-driven movement away from an up-facing QCM, and (c) a QCM frequency response that decreases according to a simple exponential function of the tethered molecules' radius of gyration. PMID- 15274570 TI - Characterization of pore structure in a nanoporous low-dielectric-constant thin film by neutron porosimetry and X-ray porosimetry. AB - A small-angle neutron scattering (SANS) porosimetry technique is presented for characterization of pore structure in nanoporous thin films. The technique is applied to characterize a spin-on organosilicate low dielectric constant (low-k) material with a random pore structure. Porosimetry experiments are conducted using a "contrast match" solvent (a mixture of toluene-d8 and toluene-h8) having the same neutron scattering length density as that of the nanoporous film matrix. The film is exposed to contrast match toluene vapor in a carrier gas (air), and pores fill with liquid by capillary condensation. The partial pressure of the solvent vapor is increased stepwise from 0 (pure air) to P0 (saturated solvent vapor) and then decreased stepwise to 0 (pure air). As the solvent partial pressure increases, pores fill with liquid solvent progressively from smallest to largest. SANS measurements quantify the average size of the empty pores (those not filled with contrast match solvent). Analogous porosimetry experiments using specular X-ray reflectivity (SXR) quantify the volume fraction of solvent adsorbed at each step. Combining SXR and SANS data yields information about the pore size distribution and illustrates the size dependence of the filling process. For comparison, the pore size distribution is also calculated by application of the classical Kelvin equation to the SXR data. PMID- 15274571 TI - Adsorption equilibrium of binary methane/ethane mixtures in BPL activated carbon: isotherms and calorimetric heats of adsorption. AB - The adsorption of pure methane and ethane in BPL activated carbon has been measured at temperatures between 264 and 373 K and at pressures up to 3.3 MPa with a bench-scale high-pressure open-flow apparatus. The same apparatus was used to measure the adsorption of binary methane/ethane mixtures in BPL at 301.4 K and at pressures up to 2.6 MPa. Thermodynamic consistency tests demonstrate that the data are thermodynamically consistent. In contrast to two sets of data previously published, we found that the adsorption of binary methane/ethane in BPL behaves ideally (in the sense of obeying ideal adsorbed solution theory, IAST) throughout the pressure and gas-phase composition range studied. A Tian-Calvet type microcalorimeter was used to measure low-pressure isotherms, the isosteric heats of adsorption of pure methane and ethane in BPL activated carbon, and the individual heats of adsorption in binary mixtures, at 297 K and at pressures up to 100 kPa. The mixture heats of adsorption were consistent with IAST. PMID- 15274572 TI - Theoretical contact angles on a nano-heterogeneous surface composed of parallel apolar and polar strips. AB - Neumann-Good's parallel strip model (J. Colloid Interface Sci. 1972, 38, 341) was used to analyze the contact angle hysteresis for a liquid on a heterogeneous surface composed of alternatively aligned horizontal apolar (theta = 70 degrees ) and polar (theta = 0 degree ) strips. The critical size of the strip width, below which the contact angle hysteresis disappears, was determined on the basis of the analysis of the activation energy for wetting to be from 6 to 12 nm. This calculated value of the critical strip size is 1 order of magnitude smaller than that of 0.1 microm, which has been commonly considered as the limit of heterogeneity size causing the appearance of the contact angle hysteresis. PMID- 15274573 TI - Contact angle hysteresis: study by dynamic cycling contact angle measurements and variable angle spectroscopic ellipsometry on polyimide. AB - The phenomenon of contact angle hysteresis was studied on smooth films of polyimide, a polymer type used in the microelectronic industry, by dynamic cycling contact angle measurements based on axisymmetric drop shape analysis profile in combination with variable angle spectroscopic ellipsometry (VASE). It was found that both advancing and receding contact angles became smaller with increasing the number of cycles and are, therefore, not a property of the dry solid alone. The changes of the wetting behavior during these dynamic cycling contact angle measurements are attributed mainly to swelling and/or liquid retention. To reveal the water-induced changes of the polymer film, the polyimide surface was studied before and after the contact with a water droplet by VASE. Both the experimental ellipsometric spectrum for Delta and that for Psi as well as the corresponding simulations show characteristic shifts due to the contact with water. The so-called effective medium approximation was applied to recover information about the thickness and effective optical constants of the polymer layer from the ellipsometrically measured values of Delta and Psi. On the basis of these results, the swelling and retention behavior of the polyimide films in contact with water droplets were discussed. PMID- 15274574 TI - Adsorption of cationic cellulose derivative/anionic surfactant complexes onto solid surfaces. II. Hydrophobized silica surfaces. AB - The effect of the anionic surfactant SDS (sodium dodecyl sulfate) on the adsorption behavior of cationic hydroxyethyl cellulose (Polymer JR-400) and hydrophobically modified cationic cellulose (Quatrisoft LM-200) at hydrophobized silica has been investigated by null ellipsometry and compared with the previous data for adsorption onto hydrophilic silica surfaces. The adsorbed amount of LM 200 is found to be considerably larger than the adsorbed amount of JR-400 at both surfaces. Both polymers had higher affinity toward hydrophobized silica than to silica. The effect of SDS on polymer adsorption was studied under two different conditions: adsorption of polymer/SDS complexes from premixed solutions and addition of SDS to preadsorbed polymer layers. Association of the surfactant to the polymer seems to control the interfacial behavior, which depends on the surfactant concentration. For the JR-400/SDS complex, the adsorbed amount on hydrophobized silica started to increase progressively from much lower SDS concentrations, while the adsorbed amount on silica increased sharply only slightly below the phase separation region. For the LM-200/SDS complex, the adsorbed amounts increased progressively from very low SDS concentrations at both surfaces, and no large difference in the adsorption behavior was observed between two surfaces below the phase separation region. The complex desorbed from the surface at high SDS concentrations above the critical micelle concentration. The reversibility of the adsorption of polymer/SDS complexes upon rinsing was also investigated. When the premixed polymer/SDS solutions at high SDS concentrations (>5 mM) were diluted by adding water, the adsorbed amount increased due to the precipitation of the complex. The effect of the rinsing process on the adsorbed layer was determined by the hydrophobicity of the polymer and the surface. PMID- 15274575 TI - Onset of cohesion in cement paste. AB - It is generally agreed that the cohesion of cement paste occurs through the formation of a network of nanoparticles of a calcium-silicate-hydrate ("C-S-H"). However, the mechanism by which these particles develop this cohesion has not been established. Here we propose a dielectric continuum model which includes all ionic interactions within a dispersion of C-S-H particles. It takes into account all co-ions and counterions explicitly (with pure Coulomb interactions between ions and between ions and the surfaces) and makes no further assumptions concerning their hydration or their interactions with the surface sites. At high surface charge densities, the model shows that the surface charge of C-S-H particles is overcompensated by Ca2+ ions, giving a reversal of the apparent particle charge. Also, at high surface charge densities, the model predicts that the correlations of ions located around neighboring particles causes an attraction between the particle surfaces. This attraction has a range of approximately 3 nm and a magnitude of 1 nN, values that are in good agreement with recent AFM experiments. These predictions are stable with respect to small changes in surface-surface separation, hydrated ion radius, and dielectric constant of the solution. The model also describes the effect of changes in cement composition through the introduction of other ions, either monovalent (Na) or multivalent (aluminum or iron hydroxide). PMID- 15274576 TI - Polymer grafting via ATRP initiated from macroinitiator synthesized on surface. AB - Macromolecular anchoring layer approach was used for preparation of an effective macroinitiator for the synthesis of grafted polymer layers by atom transfer radical polymerization (ATRP) initiated from the surface. For the initial surface modification, a thin layer of poly(glycidyl methacrylate) (PGMA) was deposited on the surface of a silicon wafer. The ATRP macroinitiator was synthesized on the substrate surface by the reaction between epoxy groups of PGMA and carboxy functionality of bromoacetic acid (BAA). Variation of the time and temperature of the BAA deposition as well as PGMA layer thickness allowed control over the amount of BAA attached to the surface. The PGMA anchoring layer allowed the achievement of initiator surface density significantly higher than that reported for a self-assembled monolayer of ATRP initiators. Polymer brushes were synthesized on the PGMA/BAA-modified substrates by ATRP. Different surface concentrations of BAA were used in our grafting experiments to acquire knowledge about the relationship between the amount of initiator anchored to the surface through PGMA and the rate of the grafted layer formation. The increase in the surface density of the initiating moieties led to the increase in the grafting rate. However, a cutoff initiator concentration beyond which no increase of the thickness of the grafted layer was observed. From comparison between the surface densities of the initiator and the attached polymer it was determined that the efficiency of the initiation from the surface was on the level of 5-15%. PMID- 15274577 TI - A method for the estimation of pore anisotropy in porous solids. AB - In this work a method for the estimation of pore anisotropy, b, in porous solids is suggested. The methodology is based on the pore size distribution and the surface area distribution, both calculated from trivial N2 adsorption-desorption isotherms. The materials used for testing the method were six MCM-Alx solids in which the ordered pore structure (for x = 0) was gradually destroyed by the introduction of Al atoms (x = 5, 10, 15, 20, 50) into the solids. Additionally, four silicas having random porosity were examined, in which the surface of the parent material SiO2 (pure silica) was gradually functionalized with organosilicate groups of various lengths (triple bond Si-H, triple bond Si-CH2OH, triple bond Si-(CH2)3OH) in order to block a variable amount of pores. As pore anisotropy, the ratio bi = Li/Di is defined where Li and Di are the length and the diameter of each group of pores i filled at a particular partial pressure (Pi/P0). The ratio of the surface area Si over the pore volume Vi, at each particular pressure (Pi/P0), is then expressed as Si3/Vi2 = 16 pi Nibi = 16 pi lambdai, where Nibi is the number of pores having anisotropy bi which are filled at each pressure (Pi/P0) and lambdai is the total anisotropy of all the pores Ni belonging to the group i of pores. Then plot of lambdai vs (Pi/P0) provides a clear picture of the variation of the total pore anisotropy lambdai as the partial pressure (Pi/P0) increases. For the functionalized silicas there appears a continuous drop of lambdai as partial pressure (Pi/P0) increases, a fact indicating that both Ni) and bi are continuously diminished. In contrast, for the MCM-Alx materials a sudden kink of lambdai appears at the partial pressure where the well-defined mesopores are filled up, a fact indicating that at this point Ni and/or bi is large. The kink disappears as the ordered porosity is destroyed by increasing the x doping in MCM-Alx. The pore anisotropy bi of each group i of pores is then estimated using the expression (Si3/Vi2) = 8 pi NiriSi and plotting log(lambdai) vs log ri. From those plots, the values of si can be found and therefore the values of bi = 0.5riSi are next defined. In the MCM-Alx materials the maximum pore anisotropy b is very high (bi approximately 250) for x = 0. Then as mesoporosity is destroyed by increasing x, the maximum b values drop gradually to b approximately 11 (x = 5), b approximately 8 (x = 10), and b approximately 3 (x = 15). For x = 20 and x = 50, the maximum b obtains values equal to unity. The same phenomena, although less profound, are also observed for the functionalized silicas, where the anisotropy b is altered by the process of functionalization and from bi approximately 0.5 for the nonfunctionalized or bi approximately 0.9 for the solid functionalized with Si-H groups drops to b = 0.3 and b = 0.2 for the solid functionalized with triple bond Si-(CH2)OH and triple bond Si-(CH2)3OH, respectively. A correlation factor F is suggested in cases where the pore model departs from the cylindrical geometry. PMID- 15274578 TI - Effect of molecular weight on synthesis and surface morphology of high-density poly(ethylene glycol) grafted layers. AB - This work describes studying the permanent grafting of carboxylic acid end functionalized poly(ethylene glycol) methyl ether (PEG) chains of different molecular weights from the melt onto a surface employing poly(glycidyl methacrylate) ultrathin film as an anchoring layer. The grafting led to the synthesis of the complete PEG brushes possessing exceptionally high grafting density. The maximum thickness of the attached PEG films was strongly dependent on the length of the polymer chains being grafted. The maximum grafting efficiency was close to the critical entanglement molecular weight region for PEG. All grafted PEG layers were in the "brush regime", since the distance between grafting sites for the layers was lower than the end-to-end distance for the anchored macromolecules. Scanning probe microscopy revealed that the grafting process led to complete PEG layers with surface smoothness on a nanometric scale. Practically all samples were partly or fully covered with crystalline domains that disappeared when samples were scanned under water. Due to the PEG hydrophilic nature, the surface with the grafted layer exhibited a low (up to 21 degrees ) water contact angle. PMID- 15274579 TI - pH-dependence of pesticide adsorption by wheat-residue-derived black carbon. AB - The potential of black carbon as an adsorbent for pesticides in soils may be strongly influenced by the properties of the adsorbent and pesticides and by the environmental conditions. This study evaluated the effect of pH on the adsorption of diuron, bromoxynil, and ametryne by a wheat (Triticum aestivum L.) residue derived black carbon (WC) as compared to a commercial activated carbon (AC). The pH drift method indicated that WC had a point of zero charge of 4.2, much lower than that of 7.8 for AC. The density of oxygen-containing surface functional groups, measured by the Boehm titration, on WC was 5.4 times higher than that on AC, resulting in a pesticide adsorption by WC being 30-50% of that by AC, due to the blockage of WC surface by the waters associated with the functional groups. A small decrease (5.5%/unit pH) in diuron adsorption by WC with increase in pH resulted from increased deprotonation of surface functional groups at higher pH values. A much larger decrease (14-21%/unit pH) in bromoxynil adsorption by WC with increase in pH resulted from the deprotonation of both the adsorbate and surface functional groups of the adsorbent. The deprotonation reduced the adsorptive interaction between bromoxynil and the neutral carbon surface and increased the electrical repulsion between the negatively charged WC surface and bromoxynil anions. Deprotonation of ametryne with increase in pH over the low pH range increased its fraction of molecular form and thus adsorption on WC by 15%/unit pH. Further increase in pH resulted in a 20%/unit pH decrease in ametryne adsorption by WC due primarily to the development of a negative charge on the surface of WC. The pH-dependent adsorption of pesticides by black carbon may significantly influence their environmental fate in soils. PMID- 15274580 TI - Dynamics of the interaction forces at the silver/solution interface during amine adsorption. AB - The forces of interaction between a silver-coated particle and a flat silver surface in an aqueous medium were measured in the presence of a series of organic amines of varying concentrations. Atomic force microscopy (AFM) was used to quantify the replacement rate of adsorbed citrate molecules on the silver surfaces by a variety of amines, under conditions where the time scale of the amine adsorption was significantly slower than the time scale of the AFM measurements. The decay length of the electrostatic double-layer interaction between the silver surfaces was found to be time independent; thus, the change in surface change density (determined from the interaction forces) was used to quantify the replacement rate of adsorbed citrate by amine. In the absence of amine, the interaction force between the silver surfaces exhibited evidence of a multilayer structure of adsorbed citrate molecules on each silver surface. Upon addition of the amine, a decrease in the interaction force was always observed, where the dynamics of the force were dependent on both concentration and the molecular structure of the amine. These results are discussed with respect to formation of colloidal particles in synthesis routes where particle aggregation has a significant impact on the control of particle morphology and size. PMID- 15274581 TI - Adsorption of enantiomeric poly(lactide)s on surface-grafted poly(L-lactide). AB - We show that resistance of densely grafted polymer layers to adsorption of chemically identical free chains, which is known to be caused by entropic expulsion of free chains from the grafted layer, can be suppressed using the grafted and free chains of opposite stereoconfiguration. Specifically, we study adsorption of poly(L-lactide) (PLLA) and its enantiomer poly(D-lactide) (PDLA) onto layers of surface-grafted PLLA in acetonitrile and chloroform by infrared spectroscopy (IR). The grafted layers with thicknesses ranging from 7 to 35 nm are produced by ring-opening polymerization of L-lactide from hydroxyl end-groups of a self-assembled monolayer on gold. The IR data indicate that adsorption on the bare gold surface is the same for the L- and D-form of the polymer. However, covering the gold with the surface-grafted PLLA produces a significant decline in the adsorption of free PLLA and, by contrast, a strong enhancement in the adsorption of free PDLA. In addition, the IR data indicate that the adsorbed PDLA chains are stereocomplexed with the grafted PLLA chains. Thus, entropic expulsion of free chains from the grafted layer, which is responsible for the resistance of surface-grafted PLLA to adsorption of free PLLA, is suppressed in the case of free PDLA by stereocomplexation between the grafted and free chains. PMID- 15274582 TI - Synthesis and nanostructure of strong polyelectrolyte brushes in amphiphilic diblock copolymer monolayers on a water surface. AB - We synthesized an ionic amphiphilic diblock copolymer, poly(hydrogenated isoprene)-b-poly(styrenesulfonic acid) (PIp-h2-b-PSS), by living anionic polymerization, and the nanostructure of its monolayer spread on a water surface was directly investigated by the in situ X-ray reflectivity technique. The monolayer of the diblock copolymer on a water surface had a smooth hydrophobic PIp-h2 layer on water and a "carpet"/polymer brush double layer in a hydrophilic sodium polystyrene sulfonate (PSSNa) layer under the water. The surface pressure dependence and PSSNa chain length dependence of the PIp-h2 layer thickness and the brush nanostructure were quantitatively studied. The effect of salt concentration in the subphase was also investigated in aqueous solutions containing 0-2 M NaCl. The salt effect on monolayer structure occurred at around 0.2 M. The thickness of the PSS brush layer decreased at salt concentrations above 0.2 M, while no structural change was observed below 0.2 M. This critical salt concentration is thought to be related to the balance of ionic concentrations inside the brush and in bulk solution. PMID- 15274583 TI - Spider-web amphiphiles as artificial lipid clusters: design, synthesis, and accommodation of lipid components at the air-water interface. AB - As a novel category of two-dimensional lipid clusters, dendrimers having an amphiphilic structure in every unit were synthesized and labeled "spider-web amphiphiles". Amphiphilic units based on a Lys-Lys-Glu tripeptide with hydrophobic tails at the C-terminal and a polar head at the N-terminal are dendrically connected through stepwise peptide coupling. This structural design allowed us to separately introduce the polar head and hydrophobic tails. Accordingly, we demonstrated the synthesis of the spider-web amphiphile series in three combinations: acetyl head/C16 chain, acetyl head/C18 chain, and ammonium head/C16 chain. All the spider-web amphiphiles were synthesized in satisfactory yields, and characterized by 1H NMR, MALDI-TOFMS, GPC, and elemental analyses. Surface pressure (pi)-molecular area (A) isotherms showed the formation of expanded monolayers except for the C18-chain amphiphile at 10 degrees C, for which the molecular area in the condensed phase is consistent with the cross sectional area assigned for all the alkyl chains. In all the spider-web amphiphiles, the molecular areas at a given pressure in the expanded phase increased in proportion to the number of units, indicating that alkyl chains freely fill the inner space of the dendritic core. The mixing of octadecanoic acid with the spider-web amphiphiles at the air-water interface induced condensation of the molecular area. From the molecular area analysis, the inclusion of the octadecanoic acid bears a stoichiometric characteristic; i.e., the number of captured octadecanoic acids in the spider-web amphiphile roughly agrees with the number of branching points in the spider-web amphiphile. PMID- 15274584 TI - Capillary bridges in electric fields. AB - We analyzed the morphology of droplets of conductive liquids placed between two parallel plate electrodes as a function of the two control parameters electrode separation and applied voltage. Both electrodes were covered by thin insulating layers, as in conventional electrowetting experiments. Depending on the values of the control parameters, three different states of the system were found: stationary capillary bridges, stationary separated droplets, and periodic self excited oscillations between both morphologies, which appear only above a certain threshold voltage. In the two stationary states, the morphology of the liquid is modified by the electric fields due to electrowetting and due to mutual electrostatic attraction, respectively. We determined a complete phase diagram within the two-dimensional phase space given by the control parameters. We discuss a model based on the interfacial and electrostatic contributions to the free energy. Numerical solutions of the model are in quantitative agreement with the phase boundaries found in the experiments. The dynamics in the oscillatory state are governed by electric charge relaxation and by contact angle hysteresis. PMID- 15274585 TI - Interfacial electrical properties of DNA-modified diamond thin films: intrinsic response and hybridization-induced field effects. AB - We have investigated the frequency-dependent interfacial electrical properties of nanocrystalline diamond films that were covalently linked to DNA oligonucleotides and how these properties are changed upon exposure to complementary and noncomplementary DNA oligonucleotides. Frequency-dependent electrical measurements at the open-circuit potential show significant changes in impedance at frequencies of >10(4) Hz when DNA-modified diamond films are exposed to complementary DNA, with only minimal changes when exposed to noncomplementary DNA molecules. Measurements as a function of potential show that at 10(5) Hz, the impedance is dominated by the space-charge region of the diamond film. DNA molecules hybridizing at the interface induce a field effect in the diamond space charge layer, altering the impedance of the diamond film. By identifying a range of impedances where the impedance is dominated by the diamond space-charge layer, we show that it possible to directly observe DNA hybridization, in real time and without additional labels, via simple measurement of the interfacial impedance. PMID- 15274586 TI - Rheology of binary colloidal structures assembled via specific biological cross linking. AB - The selectivity and range of energies offered by specific biological interactions serve as valuable tools for engineering the assembly of colloidal particles into novel materials. In this investigation, high affinity biological interactions between biotin-coated "A" particles (RA = 0.475 microm) and streptavidin-coated "B" particles (RB = 2.75 microm) drive the self-assembly of a series of binary colloidal structures, from colloidal micelles (a large B particle coated by smaller A particles) to elongated chain microstructures (alternating A and B particles), as the relative number of small (A) to large (B) particles (2 < or = NA/NB < or = 200) is decreased at a low total volume fraction (10(-4) < or = phiT < or = 10(-3)). At a significantly higher total volume fraction (phiT > or = 10( 1)) and a low number ratio (NA/NB = 2), the rheological behavior of volume filling particle networks connected by streptavidin-biotin bonds is characterized. The apparent viscosity (eta) as a function of the shear rate gamma, measured for networks at phiT = 0.1 and 0.2, exhibits shear-rate-dependent flow behavior, and both the apparent viscosity and the extent of shear thinning increase upon an increase of a factor of 2 in the total volume fraction. Micrographs taken before and after shearing show a structural breakdown of the flocculated binary particle network into smaller flocs, and ultimately a fluidlike suspension, with increasing shear rate. Rheological measurements provide further proof that suspension microstructure is governed by specific biomolecular interactions, as control experiments in which the streptavidin molecules on particles were blocked displayed Newtonian flow behavior. This investigation represents the first attempt at measuring the rheology of colloidal suspensions where assembly is driven by biomolecular cross-linking. PMID- 15274587 TI - Membrane transport of a polyacid-tied doxorubicin. AB - A model, based on fluorescence data, is developed for the poly(acrylic acid) assisted transport of doxorubicin, a cationic antitumor drug, through a bilayer membrane. Accordingly, the doxorubicin binds to the poly(acrylic acid) via electrostatic polymer-drug interactions plus drug-drug stacking within the complex. This complex associates with neutral egg lecithin vesicles by means of hydrophobic attraction between the doxorubicin and the vesicle bilayers. In the process, the doxorubicin "destacks", providing a fluorescence change that can be monitored. Finally, the doxorubicin enters the vesicle interior which has been imparted with an acidic pH to protonate the doxorubicin and thus, in a second stage, yield an additional fluorescence change that can also be detected. A portion of the poly(acrylic acid), now devoid of doxorubicin, then leaves the outer vesicle surface and enters to external solution. PMID- 15274588 TI - Silica nanoparticle size influences the structure and enzymatic activity of adsorbed lysozyme. AB - Adsorption of chicken egg lysozyme on silica nanoparticles of various diameters has been studied. Special attention has been paid to the effect of nanoparticle size on the structure and function of the adsorbed protein molecules. Both adsorption patterns and protein structure and function are strongly dependent on the size of the nanoparticles. Formation of molecular complexes is observed for adsorption onto 4-nm silica. True adsorptive behavior is evident on 20- and 100 nm particles, with the former resulting in monolayer adsorption and the latter yielding multilayer adsorption. A decrease in the solution pH results in a decrease in lysozyme adsorption. A change of protein structure upon adsorption is observed, as characterized by a loss in alpha-helix content, and this is strongly dependent on the size of the nanoparticle and the solution pH. Generally, greater loss of alpha helicity was observed for the lysozyme adsorbed onto larger nanoparticles under otherwise similar conditions. The activity of lysozyme adsorbed onto silica nanoparticles is lower than that of the free protein, and the fraction of activity lost correlates well with the decrease in alpha-helix content. These results indicate that the size of the nanoparticle, perhaps because of the contributions of surface curvature, influences adsorbed protein structure and function. PMID- 15274589 TI - Dendrimer-functionalized self-assembled monolayers as a surface plasmon resonance sensor surface. AB - We report here a multistep route for the immobilization of DNA and proteins on chemically modified gold substrates using fourth-generation NH(2)-terminated poly(amidoamine) dendrimers supported by an underlying amino undecanethiol (AUT) self-assembled monolayer (SAM). Bioactive ultrathin organic films were prepared via layer-by-layer self-assembly methods and characterized by fluorescence microscopy, variable angle spectroscopic ellipsometry, atomic force microscopy (AFM), X-ray photoelectron spectroscopy (XPS), and attenuated total internal reflection Fourier transform infrared spectroscopy (ATR-FTIR). The thickness of the AUT SAM base layer on the gold substrates was determined to be 1.3 nm from ellipsometry. Fluorescence microscopy and AFM measurements, in combination with analyses of the XPS/ATR-FTIR spectra, confirmed the presence of the dendrimer/biopolymer molecules on the multilayer sensor surfaces. Model proteins, including streptavidin and rabbit immunoglobulin proteins, were covalently attached to the dendrimer layer using linear cross-linking reagents. Through surface plasmon resonance measurements, we found that sensor surfaces containing a dendrimer layer displayed an increased protein immobilization capacity, compared to AUT SAM sensor surfaces without dendrimer molecules. Other SPR studies also revealed that the dendrimer-based surfaces are useful for the sensitive and specific detection of DNA-DNA interactions. Significantly, the multicomponent films displayed a high level of stability during repeated regeneration and hybridization cycles, and the kinetics of the DNA-DNA hybridization process did not appear to be influenced by surface mass transport limiting effects. PMID- 15274590 TI - Imaging of affinity microcontact printed proteins by using liquid crystals. AB - This paper reports the design of surfaces on which thermotropic liquid crystals can be used to image affinity microcontact printed proteins. The surfaces comprise gold films deposited onto silica substrates at an oblique angle of incidence and then functionalized with a monolayer formed from 2 mercaptoethylamine. Ellipsometric measurements confirm the transfer of anti biotin IgG to these surfaces from affinity stamps functionalized with biotinylated bovine serum albumin (BSA), while control experiments performed using anti-goat IgG confirmed the specificity of the IgG capture on the stamp. On these surfaces, anti-biotin IgG caused nematic phases of 4-cyano-4' pentylbiphenyl (5CB, Delta epsilon = epsilon(parallel) - epsilon(perpendicular) > 0) to assume orientations that were parallel to the surfaces (planar anchoring) but with azimuthal orientations that were distinct from those assumed by the liquid crystals on the amine-terminated surfaces not supporting IgGs. Following incubation of these samples for >8 h at 36 degrees C, we observed that the amine terminated regions of the surface not supporting IgG cause 5CB to undergo a transition from planar to perpendicular (homeotropic). Because N-(4 methoxybenzylidene)-4-butylaniline (MBBA) (Delta epsilon < 0) does not undergo a similar transition in orientation, this transition is consistent with the effects of an electrical double layer formed at the amine-terminated surface on the liquid crystal. Following the transition to homeotropic anchoring, the liquid crystals provide high optical contrast between regions of the surface supporting and not supporting IgG. We conclude that amine-terminated surfaces (I) uniformly align liquid crystals when not supporting proteins and (II) have sufficiently high surface free energy to capture proteins delivered to the surface from an affinity stamp, and thus they form the basis of a useful class of surfaces on which affinity microcontact printed proteins can be imaged using liquid crystals. PMID- 15274591 TI - Biological synthesis of strontium carbonate crystals using the fungus Fusarium oxysporum. AB - The total biological synthesis of SrCO3 crystals of needlelike morphology arranged into higher order quasi-linear superstructures by challenging microorganisms such as fungi with aqueous Sr2+ ions is described. We term this procedure "total biological synthesis" since the source of carbonate ions that react with aqueous Sr2+ ions is the fungus itself. We believe that secretion of proteins during growth of the fungus Fusarium oxysporum is responsible for modulating the morphology of strontianite crystals and directing their hierarchical assembly into higher order superstructures. PMID- 15274592 TI - Adsorption of oligonucleotides on PMMA/PNIPAM core-shell latexes: polarity of the PNIPAM shell probed by fluorescence. AB - The adsorption of a rhodamine X labeled oligonucleotide composed of 25-mers of thymine (dT(25)-ROX) onto the thermosensitive shell of PMMA/PNIPAM core-shell latex particles was studied at 22 and 40 degrees C, below and above the T(VPT) (volume phase transition temperature) of the PNIPAM shell, respectively. The experimental binding isotherms were well fitted with the cooperative Hill model. The Hill coefficient is lower than 1 at both temperatures showing that the adsorption is anticooperative. The polarity of the shell was probed by both the lifetimes and solvatochromic shifts of the zwitterionic form of rhodamine X. For temperatures below the shell T(VPT) has a polarity similar to that of water, while for temperatures above the transition the polarity is equivalent to that of a water/dioxane mixture with 30% (v/v) water. PMID- 15274593 TI - Influence of calcium on the self-assembly of partially hydrolyzed alpha lactalbumin. AB - Self-assembly of alpha-lactalbumin after partial hydrolysis by a protease from Bacillus licheniformis can result in nanotubular structures, which show many similarities to microtubules. Calcium plays a crucial role in this process. The objective of this investigation was to study the role of calcium in more detail. The kinetics of the hydrolysis step and the self-assembly step were monitored by respectively liquid chromatography-mass spectrometry and dynamic light scattering. The microstructure of the gels finally formed was investigated by transmission electron microscopy. This investigation demonstrates that calcium accelerated the kinetics of the self-assembly, but it had no effect on the hydrolysis kinetics. As a result of the accelerated self-assembly kinetics at a high calcium concentration, the time of gelation decreased as well. A minimum concentration of calcium needed to obtain the tubular alpha-lactalbumin structures was determined. Below R = 1.5 (mole calcium/mole alpha-lactalbumin), turbid gels with randomlike structure were obtained. Between R = 1.5 and R = 6, translucent gels with a fine stranded network of tubules were formed, while higher calcium concentrations had a negative effect on the tubule formation, resulting in amorphous structures. The optimum calcium concentration for alpha lactalbumin nanotube formation seemed to be around R = 3. PMID- 15274594 TI - Surface-grafted viologen for precipitation of silver nanoparticles and their combined bactericidal activities. AB - A viologen, N-hexyl-N'-(4-vinylbenzyl)-4,4'-bipyridinium dinitrate (HVVN), was synthesized and subsequently graft-copolymerized on poly(ethylene terephthalate) (PET) films. Silver nanoparticles can be deposited on the surface of the HVVN-PET film through photoinduced reduction of the silver ions in salt solution. The size and distribution of the silver nanoparticles can be varied by changing the reaction time. The pyridinium groups of the HVVN graft-copolymerized on the surface of the substrate possess bactericidal effects on Escherichia coli, and this antibacterial effect can be very significantly enhanced by the incorporation of silver nanoparticles on the HVVN-PET film. The dual functionalities of HVVN and silver remain stable after prolonged immersion in phosphate buffer solution and after aging in a weathering chamber. PMID- 15274595 TI - Beneficial role of cetyltrimethylammonium bromide in the enhancement of photovoltaic properties of dye-sensitized rutile TiO2 solar cells. AB - A new approach involving the introduction of the common cationic surfactant cetyltrimethylammonium bromide (CTAB) for modifying a rutile TiO2 film during its formation from hydrolyzed TiCl4 solution has been adopted, intending to improve the photoelectrochemical properties of the pertinent dye-sensitized solar cell. CTAB-routed films were found to consist of smaller clusters of near-spherical TiO2 particles, compared with larger clusters of long rod-shaped particles in the absence of CTAB. As a consequence, the photocurrent and photovoltage of the cell fabricated by using CTAB have increased significantly, leading to a conversion efficiency increase, compared with those of the cell prepared without CTAB. On the basis of FE-SEM, BET, and XRD analyses, the increases are attributed to decreased particle size, improved interparticle connectivity, and enhanced crystallinity of the CTAB-promoted TiO2 particles and decreased void volume in the film. Faster growth of the TiO2 film was another beneficial effect of CTAB. A mechanism is proposed for the beneficial role of CTAB during the film formation. PMID- 15274596 TI - Isothermal titration calorimetry of supramolecular polymers. AB - A method to characterize the self-association of supramolecular polymers by isothermal titration calorimetry (ITC) has been designed. Association constants in the range 10(4)-10(6) dm(3) mol(-1) have been successfully determined from the heat exchange occurring when a supramolecular polymer solution is injected into a calorimetric cell containing pure solvent. Very good agreement with literature values has been obtained. Compared to other techniques (such as NMR or Fourier transform infrared spectroscopy), the use of ITC presents several advantages: (i) the enthalpy of association is obtained together with the association constant from the same experiment, (ii) the measurements can be performed in almost any solvent, and (iii) systems with higher association constants can be characterized. PMID- 15274597 TI - Hexanethiolate monolayer protected 38 gold atom cluster. AB - The nucleation-growth-passivation Brust reaction has been modified so as to enrich the product in useful quantities of a 38-atom gold nanoparticle coated with a hexanethiolate monolayer. Two modifications are described, using -78 degrees C reduction temperature and a hyperexcess of thiol. Compositional evidence is presented that establishes the product as a Au38(C6)24 hexanethiolate monolayer protected cluster (MPC), based on transmission electron microscopy, laser ionization-desorption mass spectrometry, thermogravimetric analysis, and elemental analysis. Reverse phase HPLC confirms the relatively good monodispersity of the MPC products, but high-resolution double-column HPLC reveals that the MPCs are a mixture of closely related but chromatographically distinct products. Voltammetry, low energy spectrophotometry, and spectroelectrochemistry reveal, respectively, a 1.6 eV electrochemical energy gap between the first oxidation and the first reduction, an optical HOMO-LUMO energy absorbance edge at 1.3 eV, and a bleaching of optical absorbance near the 1.3 eV band edge that accompanies electrochemical oxidation of the nanoparticle. PMID- 15274598 TI - Improved surface chemistries, thin film deposition techniques, and stamp designs for nanotransfer printing. AB - Nanotransfer printing represents an additive approach for patterning thin layers of solid materials with nanometer resolution. The surface chemistries, thin film deposition techniques, and stamp designs are all important for the proper operation of this method. This paper presents some details concerning processing procedures and other considerations needed for patterning two- and three dimensional nanostructures with low density of defects and minimal distortions. PMID- 15274599 TI - Growth of gold nanoparticle arrays in TiO2 mesoporous matrixes. AB - An interconnected Au nanoparticle arrangement is obtained by electrodeposition from Au(III) soluble complexes within the pore system of block-copolymer templated mesoporous titania films. The resulting Au@TiO2 nanocomposites (5 nm Au particles, 5.5 nm amorphous titania walls) have the electrochemical behavior of a gold electrode of high surface area. The attenuation of Au surface plasmon due to -OH electroadsorption and the existence of mixed localized states in these Au@TiO2 nanocomposites are observed by in situ spectroelectrochemistry. PMID- 15274600 TI - Influence of the polydispersity of polymeric surfactants on the enantioselectivity of chiral compounds in micellar electrokinetic chromatography. AB - Poly(sodium undecenoyl-L-leucinate) (poly-L-SUL) was fractionated by the use of different molecular weight cutoff (MWCO) filters to narrow the polydispersity of the macromolecular sizes of the polymeric surfactant. The resulting polymeric surfactant fractions were characterized by the use of three techniques: (1) pulsed field gradient nuclear magnetic resonance (PFG-NMR) was used to determine the hydrodynamic radii, (2) analytical ultracentrifugation (AUC) was used to determine the molecular weights, and (3) steady-state fluorescence was used to determine the polarity of the nonfractionated and fractionated polymeric surfactants. From the data acquired from PFG-NMR, AUC, and fluorescence, it was noted that the hydrodynamic radii and molecular weight of the fractionated poly-L SUL increased, while the polarity decreased with the increase in the size of the MWCO filter. However, a similarity in physical properties was observed between the nonfractionated and 10-30K fractionated poly-L-SUL except for the hydrodynamic radius and diffusion coefficients. The influence of different macromolecular sizes of poly-L-SUL on the chiral separation of phenylthiohydantion (PTH)-amino acids and coumarinic derivatives, as test analytes, was elucidated by the use of micellar electrokinetic chromatography (MEKC). The size of polymeric surfactants as a prerequisite for chiral separation was demonstrated by comparing the separation properties of fractionated versus nonfractionated polymeric surfactants. Fractionated poly-L-SUL resulted in enhanced resolution and separation efficiency of the test analytes as compared to the case of the nonfractionated poly-L-SUL. This observation indicates that minimizing polydispersity of polymeric surfactants may be important for some chiral separation applications. PMID- 15274601 TI - Synthesis of zerovalent nanophase metal particles stabilized with poly(ethylene glycol). AB - Concurrent sonolysis of iron pentacarbonyl and poly(ethylene glycol)-400 (PEG 400) in hexadecane solvent proceeds via zero-order kinetics and results in Fe nanoparticles encapsulated in PEG-400 (Fe-PEG). The transmission electron microscopy images show Fe-PEG consisting of <3 nm Fe particles that are evenly dispersed in the PEG matrix. Mossbauer and X-ray absorption fine structure/X-ray absorption near-edge structure data reveal an ordered PEG assembly that helps protect the zerovalent Fe core. The Fe nanoparticles in Fe-PEG are superparamagnetic with a magnetization value of 45 emu/g-Fe at 10 KOe. The rheology of the synthesized material shows an unusual increase in viscosity with temperature that is likely due to lower critical saturation temperature phase segregation over 40 degrees C. The low-temperature mobility of the PEG-400 moiety in Fe-PEG would allow facile ligation of the Fe0 core with biologically and chemically active groups. PMID- 15274602 TI - Effect of ethanol on the interaction between poly(vinylpyrrolidone) and sodium dodecyl sulfate. AB - The effect of ethanol on the interaction between the anionic surfactant sodium dodecyl sulfate (SDS) and the nonionic polymer poly(vinylpyrrolidone) (PVP) has been investigated using a range of techniques including surface tension, fluorescence, electron paramagnetic resonance (EPR), small-angle neutron scattering (SANS), and viscosity. Surface tension and fluorescence studies show that the critical micelle concentration (cmc) of the surfactant decreases to a minimum value around 15 wt % ethanol; that is, it follows the cosurfactant effect. However, in the presence of PVP, the onset of the interaction, denoted cmc(1), between the surfactant and the polymer is considerably less dependent on ethanol concentration. The saturation point, cmc(2), however, reflects the behavior of the cmc in that it decreases upon addition of ethanol. This results in a decrease in the amount of surfactant bound to the polymer [C(bound) = cmc(2) - cmc] at saturation. The viscosity of simple PVP solutions depends on ethanol concentration, but since SANS studies show that ethanol has no effect on the polymer conformation, the changes observed in the viscosity reflect the viscosity of the background solvent. There are significant increases in bulk viscosity when the surfactant is added, and these have been correlated with the polymer conformation extracted from an analysis of the SANS data and with the amount of polymer adsorbed at the micelle surface. Competition between ethanol and PVP to occupy the surfactant headgroup region exists; at low ethanol concentration, the PVP displaces the ethanol and the PVP/SDS complex resembles that formed in the absence of the ethanol. At higher ethanol contents, the polymer does not bind to the ethanol-rich micelle surface. PMID- 15274603 TI - Surface modification studies of edge-oriented molybdenum sulfide nanosheets. AB - We have synthesized edge-oriented MoS2 nanosheets by the evaporation of a single source precursor based on Mo(IV)-tetrakis(diethylaminodithiocarbomato). The surface chemistry of the MoS2 nanosheets has been studied in order to evaluate the chemical reactivities of the basal planes and edges. By irradiating the MoS2 nanosheet with a scanning infrared laser, micron-scale lithographical structures can be created due to laser-induced oxidation of MoS2 to form nanocrystalline MoO3. Preferential reactivities of the MoS2 basal edges in an electrochemical environment and during vapor phase deposition have been demonstrated. Functionalization of the basal plane with 1-pyrene acetic acid allows the immobilization of DNA and immunoglobins on the MoS2 basal plane. PMID- 15274604 TI - Transport properties and distribution of water molecules confined in hydrophobic nanopores and nanoslits. AB - The transport properties, including the diffusivity and viscosity, of water confined in hydrophobic nanopores and nanoslits were studied by molecular dynamics simulations. The results show that the diffusion coefficient in nanopores and nanoslits is markedly lower than that in the bulk. But the viscosity is much larger than that in bulk. The parallel diffusion coefficient is obviously larger than the perpendicular ones. The diffusion coefficient in the channel pore is ever less than that in the slit pore at the same pore width, but the viscosity is larger. The temperature and density affect significantly the diffusivity and viscosity in nanopores and nanoslits. Lower density water exhibits some special characteristics on density profiles in nanopores and nanoslits at lower temperatures, and the density profiles show a change from homogeneous to inhomogeneous as the pore width is reduced. Even clusters occurred in micropores. PMID- 15274605 TI - Nanosphere lithography: fabrication of large-area Ag nanoparticle arrays by convective self-assembly and their characterization by scanning UV-visible extinction spectroscopy. AB - This work employs UV-visible extinction spectroscopy as a new spectral mapping technique to characterize self-assembled polystyrene microsphere samples produced by convective self-assembly (CSA). This spectroscopic technique was successfully used to analyze the periodic particle arrays produced by the polystyrene template, yielding a detailed characterization of each sample. The CSA-prepared samples proved to be more uniform across a sample as well as more reproducible than previous sample preparation techniques. For the first time, a detailed characterization and quantitative evaluation of the entire sample has been performed by spectroscopic mapping. PMID- 15274606 TI - Anisotropic thermal expansion behavior of thin films of polymethylsilsesquioxane, a spin-on-glass dielectric for high-performance integrated circuits. AB - Thin films of poly(methylsilsesquioxane) (PMSSQ) are candidates for use as interdielectric layers in advanced semiconductor devices with multilayer structures. We prepared thin films of PMSSQ with thicknesses in the range 25.0 1151.0 nm by spin-casting its soluble precursor onto Si and GaAs substrates with native oxide layers and then drying and curing the films under a nitrogen atmosphere at temperatures in the range 250-400 degrees C. The out-of-plane thermal expansion coefficient alpha(perpendicular) of each film was measured over the temperature range 25-200 degrees C using spectroscopic ellipsometry and synchrotron X-ray reflectivity, while the in-plane thermal expansion coefficient alpha(parallel) of each film was determined over the temperature range 25-400 degrees C by residual stress analysis. PMSSQ films cured at higher temperatures exhibited reduced thermal expansion, which is attributed to the denser molecular packing and higher degree of cross-linking that arises at higher temperatures. Surprisingly however, all the PMSSQ films were found to exhibit very strong anisotropic thermal expansion; alpha(perpendicular) and alpha(parallel) of the films were in the ranges 140-329 ppm/ degrees C and 12-29 ppm/ degrees C respectively, depending on the curing temperature. This is the first time that cured PMSSQ thin films have been shown to exhibit anisotropic thermal expansion behavior. This anisotropic thermal expansion of the PMSSQ thin films might be due to the anisotropy of cross-link density in the films, which arises because of a combination of factors: the preferential orientation of methyl groups toward the upper film surface and the preferential network formation in the film plane that occurs during curing of the confined film. In addition, the film electron densities were determined using synchrotron X-ray reflectivity measurements and the film biaxial moduli were obtained using residual stress analysis. PMID- 15274607 TI - Comparison of nanoparticle size and electrophoretic mobility measurements using a carbon-nanotube-based coulter counter, dynamic light scattering, transmission electron microscopy, and phase analysis light scattering. AB - The precision and accuracy of measurements of the diameter and electrophoretic mobility (mu) of polymeric nanoparticles is compared using four different analytical approaches: carbon-nanotube-based Coulter counting, dynamic light scattering (DLS), transmission electron microscopy (TEM), and phase analysis light scattering (PALS). Carbon-nanotube-based Coulter counters (CNCCs) use a 132 nm diameter channel to simultaneously determine the diameter (28-90 nm) and mu value for individual nanoparticles. These measurements are made without calibration of the CNCC and without labeling the sample. Moreover, because CNCCs measure the properties of individual particles, they provide true averages and polydispersities that are not convoluted into the intrinsic instrumental response function of the CNCC. CNCCs can be used to measure the size of individual nanoparticles dispersed in aqueous solutions, which contrasts with the TEM measured size of individual dehydrated particles and the ensemble size averages of dispersed particles provided by DLS. CNCCs provide more precise values of mu than PALS. PMID- 15274608 TI - Water-based ferrofluids from FexPt1-x nanoparticles synthesized in organic media. AB - This paper describes an effective method to transfer oleic acid/oleylamine-capped colloidal FePt nanoparticles dispersed in hexane into water, using tetramethylammonium hydroxide (TMAOH) as a phase transfer agent. FexPt1-x nanoparticles with different compositions (x = 0.32, 0.40, 0.48, 0.60, 0.66, 0.69) in the size range of 2-4 nm were synthesized by a high-temperature organometallic route with oleic acid and oleylamine as stabilizers. The surface of such nanoparticles was modified through removal of the organic, hydrophobic layer and adsorption of TMAOH, which provides the nanoparticles with sufficient surface charge so that an electrostatic double layer builds up, and the FePt nanoparticles can be fully redispersed in aqueous solution, even with high concentrations. The water-dispersible FePt nanoparticles were characterized by transmission electron microscopy, electrophoretic mobility, X-ray diffraction, and Fourier transform infrared spectroscopy. PMID- 15274609 TI - Electrochemical methods for the preparation of gold-coated TiO2 nanoparticles with variable coverages. AB - We report here the first electrochemical methods to prepare elemental Au0-coated TiO2 nanoparticles with controllable coverages. First, Au substrates were cycled in a deoxygenated aqueous solution containing 0.1 N HCl and 1 mM TiO2 nanoparticles from -0.28 to +1.22 V versus Ag/AgCl at 500 mV/s with different numbers of scans. The durations at the cathodic and anodic vertexes were 10 and 5 s, respectively. After this process, positively charged Au-coated TiO2 nanoparticles were formed in the solutions. Then a cathodic overpotential of 0.6 V from the open circuit potential of ca. 0.82 V versus Ag/AgCl was applied under sonication to synthesize elemental Au0-coated TiO2 nanoparticles. The coverage of Au shells in the elemental Au-coated TiO2 nanoparticles is varied from 10% to 95% by increasing the number of scans from 10 to 50 in preparing the positively charged Au-coated TiO2 nanoparticles. The extremely high coverage of 95% in this study is notable, as compared with other methods to prepare Au-coated TiO2 nanoparticles. PMID- 15274610 TI - Amphiphilic polysaccharides: useful tools for the preparation of nanoparticles with controlled surface characteristics. AB - Polymeric surfactants obtained by hydrophobic modification of dextran are used as stabilizers for oil-in-water emulsions. The kinetics of interfacial tension decrease is studied as a function of polymer structural characteristics (degree of hydrophobic substitution) and at various polymer concentrations. Several hydrocarbon oils, either aliphatic (octane, decane, dodecane, and hexadecane) or aromatic (styrene), are tested. Kinetics exhibits the same general trends no matter which oil or polymer is considered. The emulsifying properties of the polymeric surfactants are illustrated by the preparation of oil-in-water emulsions. The droplet size at the preparation is correlated to the amount of oil and to the polymer concentration in the aqueous phase. For low polymer/oil ratios, it is shown that the droplet size is limited by the initial amount of polymer. On the contrary, for high polymer/oil ratios, the droplet size seems to level down, indicating that other parameters become predominant. Emulsion aging occurs by Ostwald ripening, and it is demonstrated that the theoretical equation of Lifshitz, Slyozov, and Wagner (LSW) correctly describes the experimental results. The nature of the oil has important effects on emulsion aging, as described by the LSW equation. The aging of emulsions containing oil mixtures is quantitatively described on the basis of the results with pure oils. The influence of polymer chemical structure can be conveniently correlated to interfacial tension results through the LSW equation. On the contrary, the influence of oil volume fraction seems to be overestimated by the usual correction factor, k(phi). The effect of temperature on emulsion aging is finally examined. Miniemulsions stabilized with dextran derivatives are used for the radical polymerization of styrene. Following this procedure, polysaccharide covered polystyrene nanoparticles are prepared and characterized (size and surface coverage). The size of the particles is directly correlated to that of the initial droplets for styrene volume fractions around 10%. On the contrary, for initial styrene volume fractions around 20%, particles exhibit a larger size than the initial droplets, indicating that coalescence processes take place during polymerization. The amount of dextran at the surface of the particles is determined and compared to the adsorbed amounts resulting from emulsion preparation. PMID- 15274612 TI - Three-dimensional structure of an independently folded extracellular domain of human amyloid-beta precursor protein. AB - Cleavage of amyloid-beta precursor protein (APP) by site-specific proteases generates amyloid-beta peptides (Abetas), which are thought to induce Alzheimer's disease. We have identified an independently folded extracellular domain of human APP localized proximal to the Abeta sequence, and determined the three dimensional structure of this domain by NMR spectroscopy. The domain is composed of four alpha-helices, three of which form a tight antiparallel bundle, and constitutes the C-terminal half of the central extracellular region of APP that has been implicated in the regulation of APP cleavage. Sequence comparisons demonstrate that the domain is highly conserved among all members of the APP family, including invertebrate homologues, suggesting an important role for this region in the biological function of APP. The identification of this domain and the availability of its atomic structure will facilitate analysis of APP function and of the role of the extracellular region in the regulation of APP cleavage. PMID- 15274613 TI - Structural stability and unfolding properties of thermostable bacterial alpha amylases: a comparative study of homologous enzymes. AB - In a comparative investigation on two thermostable alpha-amylases [Bacillus amyloliquefaciens (BAA), T(m) = 86 degrees C and Bacillus licheniformis (BLA), T(m) = 101 degrees C], we studied thermal and guanidine hydrochloride (GndHCl) induced unfolding using fluorescence and CD spectroscopy, as well as dynamic light scattering. Depletion of calcium from specific ion-binding sites in the protein structures reduces the melting temperature tremendously for both alpha amylases. The reduction is nearly the same for both enzymes, namely, in the order of 50 degrees C. Thus, the difference in thermostability between BLA and BAA (DeltaT(m) approximately 15 degrees C) is related to intrinsic properties of the respective protein structures themselves and is not related to the strength of ion binding. The thermal unfolding of both proteins is characterized by a full disappearance of secondary structure elements and by a concurrent expansion of the 3D structure. GndHCl-induced unfolding also yields a fully vanishing secondary structure but with more expanded 3D structures. Both alpha-amylases remain much more compact upon thermal unfolding as compared to the fully unfolded state induced by chemical denaturants. Such rather compact thermal unfolded structures lower the conformational entropy change during the unfolding transition, which principally can contribute to an increased thermal stability. Structural flexibilities of both enzymes, as measured with tryptophan fluorescence quenching, are almost identical for both enzymes in the native states, as well as in the unfolded states. Furthermore, we do not observe any difference in the temperature dependence of the structural flexibilities between BLA and BAA. These results indicate that conformational dynamics on the time scale of our studies seem not to be related to thermal stability or to thermal adaptation. PMID- 15274614 TI - Partial B-to-A DNA transition upon minor groove binding of protein Sac7d monitored by Raman spectroscopy. AB - Members of the Sso7d/Sac7d protein family and other related proteins are believed to play an important role in DNA packaging and maintenance in archeons. Sso7d/Sac7d are small, abundant, basic, and nonspecific DNA-binding proteins of the hyperthermophilic archeon Sulfolobus. Structures of several complexes of Sso7d/Sac7d with DNA octamers are known. These structures are characterized by sequence unspecific minor groove binding of the proteins and sharp kinking of the double helix. Corresponding Raman vibrational signatures have been identified in this study. A Raman spectroscopic analysis of Sac7d binding to the oligonucleotide decamer d(GAGGCGCCTC)(2) reveals large conformational perturbations in the DNA structure upon complex formation. Perturbed Raman bands are associated with the vibrational modes of the sugar phosphate backbone and frequency shifts of bands assigned to nucleoside vibrations. Large changes in the DNA backbone and partial B- to A-form DNA transitions are indicated that are closely associated with C2'-endo/anti to C3'-endo/anti conversion of the deoxyadenosyl moiety upon Sac7d binding. The major spectral feature of Sac7d binding is kinking of the DNA. Raman markers of minor groove binding do not largely contribute to spectral differences; however, clear indications for minor groove binding come from G-N2 and G-N3 signals that are supported by Trp24 features. Trp24 is the only tryptophan present in Sac7d and binds to guanine N3, as has been demonstrated clearly in X-ray structures of Sac7d-DNA complexes. No changes of the Sac7d secondary structure have been detected upon DNA binding. PMID- 15274615 TI - Fibrillation of carrier protein albebetin and its biologically active constructs. Multiple oligomeric intermediates and pathways. AB - We showed that the genetically engineered carrier-protein albebetin and its biologically active constructs with interferon-alpha(2) octapeptide LKEKKYSP or differentiation factor hexapeptide TGENHR are inherently highly amyloidogenic at physiological pH. The kinetics of fibrillation were monitored by thioflavine-T (ThT) binding and the morphological changes by atomic force microscopy. Fibrillation proceeds via multiple pathways and includes a hierarchy of amyloid structures ranging from oligomers to protofilaments and fibrils. Comparative height and volume microscopic measurements allowed us to identify two distinct types of oligomeric intermediates: pivotal oligomers ca. 1.2 nm in height comprised of 10-12 monomers and on-pathway amyloid-competent oligomers ca. 2 nm in height constituted of 26-30 molecules. The former assemble into chains and rings with "bead-on-string morphology", in which a "bead" corresponds to an individual oligomer. Once formed, the rings and chains remain in solution simultaneously with fibrils. The latter give rise to protofilaments and fibrils, and their formation is concomitant with an increasing level of ThT binding. The amyloid nature of filamentous structures was confirmed by a pronounced ThT and Congo red binding and beta-sheet-rich far-UV circular dichroism. We suggest that transformation of the pivotal oligomers into the amyloid-prone ones is a limiting stage in amyloid assembly. Peptides, either fused to albebetin or added into solution, and an increased ionic strength promote fibrillation of albebetin (net charge of -12) by counterbalancing critical electrostatic repulsions. This finding demonstrates that the fibrillation of newly designed polypeptide-based products can produce multimeric amyloid species with a potentially "new" functionality, raising questions about their safety. PMID- 15274616 TI - Spectroscopic and X-ray crystallographic characterization of bestatin bound to the aminopeptidase from Aeromonas (Vibrio) proteolytica. AB - Binding of the competitive, slow-binding inhibitor bestatin ([(2S,3R)-3-amino-2 hydroxy-4-phenylbutanoy]-leucine) to the aminopeptidase from Aeromonas proteolytica (AAP) was examined by both spectroscopic and crystallographic methods. Electronic absorption spectra of the catalytically competent [Co_(AAP)], [CoCo(AAP)], and [ZnCo(AAP)] enzymes recorded in the presence of bestatin revealed that both of the divalent metal ions in AAP are involved in binding bestatin. The electron paramagnetic resonance (EPR) spectrum of the [CoCo(AAP)] bestatin complex exhibited no observable perpendicular- or parallel-mode signal. These data indicate that the two Co(II) ions in AAP are antiferromagnetically coupled yielding an S = 0 ground state and suggest that a single oxygen atom bridges between the two divalent metal ions. The EPR data obtained for [CoZn(AAP)] and [ZnCo(AAP)] confirm that bestatin interacts with both metal ions. The X-ray crystal structure of the [ZnZn(AAP)]-bestatin complex was solved to 2.0 A resolution. Both side chains of bestatin occupy a well-defined hydrophobic pocket that is adjacent to the dinuclear Zn(II) active site. The amino acid residues ligated to the dizinc(II) cluster in AAP are identical to those in the native structure with only minor perturbations in bond length. The alkoxide oxygen of bestatin bridges between the two Zn(II) ions in the active site, displacing the bridging water molecule observed in the native [ZnZn(AAP)] structure. The M-M distances observed in the AAP-bestatin complex and native AAP are identical (3.5 A) with alkoxide oxygen atom distances of 2.1 and 1.9 A from Zn1 and Zn2, respectively. Interestingly, the backbone carbonyl oxygen atom of bestatin is coordinated to Znl at a distance of 2.3 A. In addition, the NH(2) group of bestatin, which mimics the N-terminal amine group of an incoming peptide, binds to Zn2 with a bond distance of 2.3 A. A combination of the spectroscopic and X-ray crystallographic data presented herein with the previously reported mechanistic data for AAP has provided additional insight into the substrate-binding step of peptide hydrolysis as well as insight into important small molecule features for inhibitor design. PMID- 15274617 TI - Structure-activity relationship of crustacean molt-inhibiting hormone from the kuruma prawn Marsupenaeus japonicus. AB - In crustaceans, molt-inhibiting hormone (MIH) controls molting by suppressing the synthesis and/or secretion of molting hormone. In our previous study, which determined the solution structure of MIH by NMR, we hypothesized that the peptide's functional site spanned the region encompassing the N-terminal alpha helix and a portion of the C-terminus, both of which are located sterically close to each other [Katayama et al. (2003) J. Biol. Chem. 278, 9620-9623]. To confirm this hypothesis, various mutants of MIH were prepared and their molt-inhibiting activities were assessed. All peptides mutated at the putative functional site exhibited circular dichroism spectra similar to the natural MIH, suggesting that the mutants retained their natural conformation regardless of the mutations. As expected, a majority of the mutants, except for Delta12 (a deletion mutant of Gly(12)) and Delta75-77 (a deletion mutant of the last three residues of the C terminus), were less active than the natural MIH. In particular, I72G exhibited no molt-inhibiting activity even at 200 nM, while N13A and S71Y exhibited low activity at the same concentration. In contrast, the natural and recombinant MIHs exhibited full inhibitory activity at 20 nM. All these results indicate that the functional site of MIH is located in the region containing the C-terminal ends of the N- and C-terminal alpha-helices, and that Asn(13), Ser(71), and Ile(72) are especially significant for conferring molt-inhibiting activity. Furthermore, these findings agree with the results and the proposed hypothesis presented in previous studies on the structure-activity relationship of MIH and its related peptides. PMID- 15274618 TI - FTIR spectroscopy of the K photointermediate of Neurospora rhodopsin: structural changes of the retinal, protein, and water molecules after photoisomerization. AB - Neurospora rhodopsin (NR, also known as NOP-1) is the first rhodopsin of the haloarchaeal type found in eucaryotes. NR demonstrates a very high degree of conservation of the amino acids that constitute the proton-conducting pathway in bacteriorhodopsin (BR), a light-driven proton pump of archaea. Nevertheless, NR does not appear to pump protons, suggesting the absence of the reprotonation switch that is necessary for the active transport. The photocycle of NR is much slower than that of BR, similar to the case of pharaonis phoborhodopsin (ppR), an archaeal photosensory protein. The functional and photochemical differences between NR and BR should be explained in the structural context. In this paper, we studied the structural changes of NR following retinal photoisomerization by means of low-temperature Fourier transform infrared (FTIR) spectroscopy and compared the obtained spectra with those for BR. For the spectroscopic analysis, we established the light-adaptation procedure for NR reconstituted into 1,2 dimyristoyl-sn-glycero- 3-phosphocholine/1,2-dimyristoyl-sn-glycero-3-phosphate (DMPC/DMPA) liposomes, which takes approximately 2 orders of magnitudes longer than in BR. The structure of the retinal chromophore and the hydrogen-bonding strength of the Schiff base in NR are similar to those in BR. Unique spectral features are observed for the S-H stretching vibrations of cysteine and amide-I vibrations for NR before and after retinal isomerization. In NR, there are no spectral changes assignable to the amide bands of alpha helices. The most prominent difference between NR and BR was seen for the water O-D stretching vibrations (measured in D(2)O). Unlike for haloarchaeal rhodopsins such as BR and ppR, no O-D stretches of water under strong hydrogen-bonded conditions (<2400 cm( 1)) were observed in the NR(K) minus NR difference spectra. This suggests a unique hydrogen-bonded network of the Schiff base region, which may be responsible for the lack of the reprotonation switch in NR. PMID- 15274619 TI - NMR and molecular modeling studies of the interaction between wheat germ agglutinin and the beta-D-GlcpNAc-(1-->6)-alpha-D-Manp epitope present in glycoproteins of tumor cells. AB - The beta-D-GlcpNAc-(1-->6)-alpha-D-Manp disaccharide is a constituent of highly branched cell-surface glycoconjugates that are malignancy markers. The conformational preference of the disaccharide beta-D-GlcpNAc-(1-->6)-alpha-D-Manp OMe in solution has been studied by molecular modeling and NMR spectroscopy including 1D (1)H,(1)H T-ROESY experiments and analysis of (3)J(H,H) of the hydroxymethyl group being part of the glycosidic linkage of the disaccharide, which revealed the relative populations of the omega torsion angle as gt = 0.60, gg = 0.35, and tg = 0.05. Good agreement was obtained between the effective proton-proton distances from the experiment and those obtained by molecular modeling when the flexibility at the omega torsion angle was taken into account. Molecular modeling of the disaccharide in the binding sites of the lectin wheat germ agglutinin indicates that several conformations could be adopted in the bound state. (1)H NMR and transfer NOESY experiments confirmed that binding took place, and trans-glycosidic proton-proton interactions indicated that a conformational preference was present in the bound state, as observed by the relative change of the NOEs from H1' to H6(pro-R) and H6(pro-S). STD NMR experiments showed that binding occurred in the region of the N-acetyl group of the terminal sugar residue. In addition, the O-methyl group received saturation transfer because of the proximity to the protein. (1)H,(1)H NOEs indicated that the two methyl groups were close in space, as observed in only one of the predicted bound conformations. Experimental and theoretical data therefore agree that one conformation with a gt conformation of the hydroxymethyl group and a negative sign for the psi torsion angle is indeed selected by the lectin upon binding. PMID- 15274620 TI - Kinetic studies of Thermobifida fusca Cel9A active site mutant enzymes. AB - Thermobifida fusca Cel9A-90, an unusual family 9 enzyme, is a processive endoglucanase containing a catalytic domain closely linked to a family 3c cellulose binding domain (Cel9A-68) followed by a fibronectin III-like domain and a family 2 cellulose binding domain. To study its catalytic mechanism, 12 mutant genes with changes in five conserved residues of Cel9A-68 were constructed, cloned, and expressed in Escherichia coli. The purified mutant enzymes were assayed for their activities on (carboxymethyl)cellulose, phosphoric acid-swollen cellulose, bacterial microcrystalline cellulose, and 2,4-dinitrophenyl beta-D cellobioside. They were also tested for ligand binding, enzyme processivity, and thermostability. The results clearly show that E424 functions as the catalytic acid, D55 and D58 are both required for catalytic base activity, and Y206 plays an important role in binding, catalysis, and processivity, while Y318 plays an important role in binding of crystalline cellulose substrates and is required for processivity. Several amino acids located in a loop at the end of the catalytic cleft (T245-L251) were deleted from Cel9A-68, and this enzyme showed slightly improved filter paper activity and binding to BMCC but otherwise behaved like the wild-type enzyme. The FnIII-like domain was deleted from Cel9A-90, reducing BMCC activity to 43% of the wild type. PMID- 15274621 TI - Thermodynamic evaluation of a covalently bonded transition state analogue inhibitor: inhibition of beta-lactamases by phosphonates. AB - Serine beta-lactamases are inhibited by phosphonate monoesters in a reaction that involves phosphonylation of the active site serine residue. This reaction is much more rapid than the hydrolysis of these inhibitors in solution under the same conditions. The beta-lactamase active site therefore must have the ability to stabilize not only the anionic tetrahedral transition states of the acyl transfer reactions of substrates but also the pentacoordinated transition state(s) of phosphyl transfer reactions. A series of p-nitrophenyl arylphosphonates have been synthesized and the rate constants for their inhibition of the class C beta lactamase of Enterobacter cloacae P99 determined. There is no direct correlation between these rate constants and the dissociation constants of analogous aryl boronic acids, where the latter are believed to generate good tetrahedral transition state analogue structures. Thus, the mode of stabilization of pentacoordinated phosphorus transition states by the beta-lactamase active site is qualitatively different from that of tetrahedral transition states. Molecular modeling suggests that the difference arises from different positioning of the side chain and of one of the oxygen ligands. In principle, the quality of the stable tetrahedral phosphonate complex as a transition state analogue structure can be assessed from the effect of its formation on the stability of the protein. Phosphonylation of the P99 beta-lactamase, however, had little effect on the stability of the protein, as measured both by thermal and guanidine hydrochloride denaturation. Consideration of the results of similar experiments with the Staphylococcus aureus PC1 beta-lactamase, where considerable stabilization is observed in thermal melting and, to a lesser degree, in formation of the molten globule in guanidine hydrochloride, but not in the complete unfolding transition in guanidine, suggests that results from the method may be strongly influenced by the interactions of the ligand with its environment in the unfolded state of the protein. Thus, quantitative estimates of the quality of a covalently bonded transition state analogue cannot generally be achieved by this method. PMID- 15274622 TI - Crystal structures of human glutaryl-CoA dehydrogenase with and without an alternate substrate: structural bases of dehydrogenation and decarboxylation reactions. AB - Acyl-CoA dehydrogenases (ACDs) are a family of flavoenzymes that metabolize fatty acids and some amino acids. Of nine known ACDs, glutaryl-CoA dehydrogenase (GCD) is unique: in addition to the alpha,beta-dehydrogenation reaction, common to all ACDs, GCD catalyzes decarboxylation of glutaryl-CoA to produce CO(2) and crotonyl CoA. Crystal structures of GCD and its complex with 4-nitrobutyryl-CoA have been determined to 2.1 and 2.6 A, respectively. The overall polypeptide folds are the same and similar to the structures of other family members. The active site of the unliganded structure is filled with water molecules that are displaced when enzyme binds the substrate. The structure strongly suggests that the mechanism of dehydrogenation is the same as in other ACDs. The substrate binds at the re side of the FAD ring. Glu370 abstracts the C2 pro-R proton, which is acidified by the polarization of the thiolester carbonyl oxygen through hydrogen bonding to the 2' OH of FAD and the amide nitrogen of Glu370. The C3 pro-R proton is transferred to the N(5) atom of FAD. The structures indicate a plausible mechanism for the decarboxylation reaction. The carbonyl polarization initiates decarboxylation, and Arg94 stabilizes the transient crotonyl-CoA anion. Protonation of the crotonyl-CoA anion occurs by a 1,3-prototropic shift catalyzed by the conjugated acid of the general base, Glu370. A tight hydrogen-bonding network involving gamma-carboxylate of the enzyme-bound glutaconyl-CoA, with Tyr369, Glu87, Arg94, Ser95, and Thr170, optimizes orientation of the gamma-carboxylate for decarboxylation. Some pathogenic mutations are explained by the structure. The mutations affect protein folding, stability, and/or substrate binding, resulting in inefficient/inactive enzyme. PMID- 15274623 TI - Multiple substrate binding states and chiral recognition in cofactor-independent glutamate racemase: a molecular dynamics study. AB - Glutamate racemase (MurI) catalyzes the racemization of glutamate; two cysteine residues serve as catalytic acid and base. On the basis of the crystal structure of MurI from the hyperthermophilic bacterium Aquifex pyrophilus, we performed molecular dynamics (MD) simulations of six different systems to investigate stereochemistry, substrate ligation, and active site protonation state. The catalytic competence of individual systems was assessed by the abundance of reactive conformers. Only systems in which Cys70 is poised to deprotonate d-Glu were found to be catalytically competent (idem Cys178/l-Glu), in agreement with the experimentally observed stereochemistry of Lactobacillus fermentii MurI [Tanner, M. E. et al. (1993) Biochemistry 32, 3998-4006]. Only systems in which the alpha-amino group of l/d-Glu and the imidazole moiety of His are deprotonated are catalytically competent. The active site of MurI displays an unusual flexibility in substrate ligation, and several transitions between stable binding patterns were observed. In catalytically competent binding states, the conserved threonine residues 72, 114, and 117 ligate the alpha-carboxylate of Glu and the Asn71 amides ligate the alpha-amino group of Glu, whereas the delta-carboxylate of Glu is steered by electrostatic repulsion from the Asp7 and Glu147 side chain carboxylates. A network of hydrogen bonds controls the positioning of each thiol/thiolate. In what we term substrate flipping, Glu suddenly rotates into a binding pattern that resembles the post-racemization state of the other enantiomer, i.e., each enantiomer can be bound in two distinct states. Substrate flipping and unfavorable substrate binding successively trigger dissociation of the substrate, accompanied by an opening of the active site channel. We explain how the weak binding of Glu contributes to catalysis and suggest a mechanism by which binding mismatches are propagated into an opening of the active site. PMID- 15274624 TI - Tracing sequence diversity change of RNA-cleaving deoxyribozymes under increasing selection pressure during in vitro selection. AB - In vitro selection has been used extensively over the past 10 years to create functionally diverse DNA enzymes. The majority of in vitro selection experiments to date have focused on the outcome rather than the process itself, a process that remains to be fully elucidated. In vitro selection techniques rely on the probability that some DNA molecules in a random-sequence library will fold into an appropriate tertiary structure and catalyze a desired reaction. Thus, sufficient sequence diversity in the DNA pool (and hence more catalytic DNA sequences) is a prerequisite for the successful isolation of efficient deoxyribozymes. The catalytic sequence diversity established by in vitro selection is governed largely by the choice of selection pressures, one of which is the length of the reaction time. The objective of this study was to evaluate the sequence diversity change of a pool of RNA-cleaving deoxyribozymes as a function of the reaction time. Seventeen rounds of in vitro selection were performed, and the reaction time was progressively decreased from 5 h to 5 s. A representative population from each time class was subsequently cloned and sequenced. A decline in sequence diversity was observed with decreasing reaction time, and the relationship appears to be logarithmic. In contrast, a control selection performed with a constant reaction time during each round led to a linear and comparatively very slow decrease in sequence diversity. This study provides the first methodical examination of the change in catalytic sequence diversity that occurs through the course of a deoxyribozyme selection experiment. Moreover, it represents a first step toward fully understanding the intricate pathway that lies between the beginning and end of an in vitro selection experiment. PMID- 15274625 TI - Structural basis for catalytic differences between alpha class human glutathione transferases hGSTA1-1 and hGSTA2-2 for glutathione conjugation of environmental carcinogen benzo[a]pyrene-7,8-diol-9,10-epoxide. AB - The ultimate diol epoxide carcinogens derived from polycyclic aromatic hydrocarbons, such as benzo[a]pyrene (BP), are metabolized primarily by glutathione (GSH) conjugation reaction catalyzed by GSH transferases (GSTs). In human liver and probably lung, the alpha class GSTs are likely to be responsible for the majority of this reaction because of their high abundance. The catalytic efficiency for GSH conjugation of the carcinogenic (+)-anti-benzo[a]pyrene-7,8 diol-9,10-epoxide [(+)-anti-BPDE] is more than 5-fold higher for hGSTA1-1 than for hGSTA2-2. Here, we demonstrate that mutation of isoleucine-11 of hGSTA2-2, a residue located in the hydrophobic substrate-binding site (H-site) of the enzyme, to alanine (which is present in the same position in hGSTA1-1) results in about a 7-fold increase in catalytic efficiency for (+)-anti-BPDE-GSH conjugation. Thus, a single amino acid substitution is sufficient to convert hGSTA2-2 to a protein that matches hGSTA1-1 in its catalytic efficiency. The increased catalytic efficiency of hGSTA2/I11A is accompanied by greater enantioselectivity for the carcinogenic (+)-anti-BPDE over (-)-anti-BPDE. Further remodeling of the H-site of hGSTA2-2 to resemble that of hGSTA1-1 (S9F, I11A, F110V, and S215A mutations, SIFS mutant) results in an enzyme whose catalytic efficiency is approximately 13.5-fold higher than that of the wild-type hGSTA2-2, and about 2.5-fold higher than that of the wild-type hGSTA1-1. The increased activity upon mutations can be rationalized by the interactions of the amino acid side chains with the substrate and the orientation of the substrate in the active site, as visualized by molecular modeling. Interestingly, the catalytic efficiency of hGSTA2-2 toward ( )-anti-BPDE was increased to a level close to that of hGSTA1-1 upon F110V, not I11A, mutation. Similar to (+)-anti-BPDE, however, the SIFS mutant was the most efficient enzyme for GSH conjugation of (-)-anti-BPDE. PMID- 15274626 TI - Consequences of nonlytic membrane perturbation to the translocation of the cell penetrating peptide pep-1 in lipidic vesicles. AB - The action of the cell penetrating pep-1 at the molecular level is not clearly understood. The ability of the peptide to induce (1) vesicle aggregation, (2) lipidic fusion, (3) anionic lipid segregation, (4) pore or other lytic structure formation, (5) asymmetric lipidic flip-flop, and (6) peptide translocation across the bilayers in large unilamellar vesicles was studied using photophysical methodologies mainly related to fluorescence spectroscopy. Neflometry and turbidimetry techniques show that clustering of vesicles occurs in the presence of the peptide in a concentration- and anionic lipid content-dependent manner. Results from Forster resonance energy transfer-based methodologies prove lipidic fusion and anionic lipid segregation, but no evidence for pores or other lytic structures was found. Asymmetric lipid flip-flop was not detected either. A specific method related to the quenching of the rhodamine-labeled lipids by pep-1 was developed to study the eventual translocation of the peptide. Translocation does not occur in symmetrical neutral and negatively charged vesicles, except when a valinomycin-induced transmembrane potential exists. Our work strongly suggests that the main driving force for peptide translocation is charge asymmetry between the outer and inner leaflet of biological membranes and reveals that pep-1 is able to perturb membranes without being cytotoxic. This nonlytic perturbation is probably mandatory for translocation to occur. PMID- 15274627 TI - Botulinum neurotoxin A changes conformation upon binding to ganglioside GT1b. AB - In this work, the kinetics of the binding of botulinum neurotoxin A (BoNT/A) to ganglioside GT1b were studied using surface plasmon resonance (SPR). The neurotoxin bound polysialylated gangliosides, and that binding was affected by the ionic strength of the buffer. Although the level of binding was decreased at higher ionic strengths, it could be easily observed in Tris buffer, containing 150 mM NaCl. Data analysis revealed that the binding of BoNT/A to a GT1b containing phospholipid monolayer did not fit a traditional 1:1 model. Subsequent studies, in which the time of contact between BoNT/A and GT1b was varied, indicated that the BoNT/A-GT1b complex became more stable over time, as evidenced by its reduced rate of dissociation. Circular dichroism indicated that when BoNT/A was incubated with GT1b, it underwent a conformational change that resulted in an increase in alpha-helix content and a decrease in beta-sheet content. Therefore, the SPR kinetic data were fit to a conformational change model and kinetic rate constants determined. The apparent K(D) values obtained for the binding of BoNT/A to ganglioside GT1b ranged from 2.83 x 10(-7) to 1.86 x 10(-7) M, depending on the ionic strength of the buffer. PMID- 15274628 TI - Drug targeting of HIV-1 RNA.DNA hybrid structures: thermodynamics of recognition and impact on reverse transcriptase-mediated ribonuclease H activity and viral replication. AB - RNA degradation via the ribonuclease H (RNase H) activity of human immunodeficiency virus type I (HIV-1) reverse transcriptase (RT) is a critical component of the reverse transcription process. In this connection, mutations of RT that inactivate RNase H activity result in noninfectious virus particles. Thus, interfering with the RNase H activity of RT represents a potential vehicle for the inhibition of HIV-1 replication. Here, we demonstrate an approach for inhibiting the RNase H activity of HIV-1 RT by targeting its RNA.DNA hybrid substrates. Specifically, we show that the binding of the 4,5-disubstituted 2 deoxystreptamine aminoglycosides, neomycin, paromomycin, and ribostamycin, to two different chimeric RNA-DNA duplexes, which mimic two distinct intermediates in the reverse transcription process, inhibits specific RT-mediated RNase H cleavage, with this inhibition being competitive in nature. UV melting and isothermal titration calorimetry studies reveal a correlation between the relative binding affinities of the three drugs for each of the chimeric RNA-DNA host duplexes and the relative extents to which the drugs inhibit RT-mediated RNase H cleavage of the duplexes. Significantly, this correlation also extends to the relative efficacies with which the drugs inhibit HIV-1 replication. In the aggregate, our results highlight a potential strategy for AIDS chemotherapy that should not be compromised by the unusual genetic diversity of HIV-1. PMID- 15274629 TI - Characterization of the human mitochondrial methionyl-tRNA synthetase. AB - Human mitochondrial methionyl-tRNA synthetase (human mtMetRS) has been identified from the human EST database. The cDNA encodes a 593 amino acid protein with an 18 amino acid mitochondrial import signal sequence. Sequence analysis indicates that this protein contains the consensus motifs characteristic of a class I aminoacyl tRNA synthetase but lacks the Zn(2+) binding motif and C-terminal dimerization region found in MetRSs from various organisms. The mature form of human mtMetRS has been cloned and expressed in Escherichia coli. Gel filtration experiments indicate that this protein functions as a monomer with an apparent molecular mass of 67 kDa. The kinetic parameters for activation of methionine have been determined for the purified enzyme. The K(M) and k(cat) for aminoacylation of E. coli initiator tRNA(f)(Met) are reported. The kinetics of aminoacylation of an in vitro transcript of human mitochondrial tRNA(Met) (mtRNA(Met)) have been determined. To address the effects of the modification of mtRNA on recognition of the mitochondrial tRNA by human mtMetRS, the kinetics of aminoacylation of native bovine mtRNA(Met) and of an in vitro transcript of the bovine mtRNA(Met) have also been investigated. PMID- 15274630 TI - Mapping of factor XIII solvent accessibility as a function of activation state using chemical modification methods. AB - The transglutaminase Factor XIII (FXIII) catalyzes the formation of covalent cross-links between adjacent noncovalently associated fibrin chains in blood coagulation. The resulting covalently cross-linked hard clot is much more mechanically stable and resistant to proteolytic degradation. FXIII is activated by the serine protease thrombin in the presence of calcium ions. Protein modification experiments involving the labeling of cysteine and lysine side chains of the enzyme were performed before and after activation of the enzyme in an effort to gain further insight into structural changes occurring during the activation of FXIII. The experiments revealed differences in the labeling patterns of nonactivated and activated FXIII. These differences result from the exposure or sequestration of specific cysteine or lysine residues when the enzyme is activated, either physiologically with thrombin or nonproteolytically by exposure to calcium. Of note is the acetylation of Lys 73 and Lys 221 upon activation. Both of these residues lie within possible substrate recognition regions of FXIII. The active site Cys 314 is consistently alkylated in the activated enzyme, as is Cys 409, located near the dimer interface. Within the beta-barrel 2 domain of FXIII, Cys 695 becomes alkylated in activated FXIII. Within the same domain, an acetylated Lys (677 or 678), which is observed in the zymogen, cannot be found in the activated enzyme. The results provide a more extensive view of FXIII activation than has been previously available. PMID- 15274631 TI - Recognition of sphingomyelin by lysenin and lysenin-related proteins. AB - Lysenin is a sphingomyelin (SM)-specific toxin isolated from the coelomic fluid of the earthworm Eisenia foetida. Lysenin comprises a family of proteins together with lysenin-related protein 1 (LRP-1, lysenin 2) and LRP-2 (lysenin 3). In the present study, we characterized LRP-1 and LRP-2 together with lysenin using maltose-binding-protein-tagged recombinant proteins. LRP-2 specifically bound SM and induced hemolysis like lysenin. In contrast the binding and hemolytic activities of LRP-1 were 10 times less than those of lysenin and LRP-2. Lysenin and LRP-2 share 30 common sites of aromatic amino acids. Among them, only one position, phenylalanine 210, is substituted for isoleucine in LRP-1. The activity of LRP-1 was dramatically increased by introducing a single amino acid substitution of isoleucine 210 to phenylalanine, suggesting the importance of this aromatic amino acid in biological activities of lysenin and LRPs. The importance of aromatic amino acids was further indicated by a systematic tryptophan to alanine mutation of lysenin. Lysenin contains six tryptophan residues of which five are conserved in LRP-1 and -2. We showed that the conserved tryptophans but not the nonconserved one were required both in the recognition of SM and in the hemolytic activity of lysenin. Our results suggest the importance of tryptophan in the toxin function likely due to a direct recognition of SM or in maintaining the protein structure. PMID- 15274632 TI - Purification and characterization of the human gamma-secretase complex. AB - Gamma-secretase is a member of an unusual class of proteases with intramembrane catalytic sites. This enzyme cleaves many type I membrane proteins, including the amyloid beta-protein (Abeta) precursor (APP) and the Notch receptor. Biochemical and genetic studies have identified four membrane proteins as components of gamma secretase: heterodimeric presenilin (PS) composed of its N- and C-terminal fragments (PS-NTF/CTF), a mature glycosylated form of nicastrin (NCT), Aph-1, and Pen-2. Recent data from studies in Drosophila, mammalian, and yeast cells suggest that PS, NCT, Aph-1, and Pen-2 are necessary and sufficient to reconstitute gamma secretase activity. However, many unresolved issues, in particular the possibility of other structural or regulatory components, would be resolved by actually purifying the enzyme. Here, we report a detailed, multistep purification procedure for active gamma-secretase and an initial characterization of the purified protease. Extensive mass spectrometry of the purified proteins strongly suggests that PS-NTF/CTF, mNCT, Aph-1, and Pen-2 are the components of active gamma-secretase. Using the purified gamma-secretase, we describe factors that modulate the production of specific Abeta species: (1) phosphatidylcholine and sphingomyelin dramatically improve activity without changing cleavage specificity within an APP substrate; (2) increasing CHAPSO concentrations from 0.1 to 0.25% yields a approximately 100% increase in Abeta42 production; (3) exposure of an APP-based recombinant substrate to 0.5% SDS modulates cleavage specificity from a disease-mimicking pattern (high Abeta42/43) to a physiological pattern (high Abeta40); and (4) sulindac sulfide directly and preferentially decreases Abeta42 cleavage within the purified complex. Taken together, our results define a procedure for purifying active gamma-secretase and suggest that the lipid mediated conformation of both enzyme and substrate regulate the production of the potentially neurotoxic Abeta42 and Abeta43 peptides. PMID- 15274633 TI - Thermodynamic mechanism and consequences of the polyproline II (PII) structural bias in the denatured states of proteins. AB - A quantitative characterization of the structure and energy of the denatured states of proteins represents the cornerstone to a molecular-level understanding of both protein stability and fold specificity. Recent studies have revealed a significant bias in unstructured peptides toward the polyproline II (P(II)) conformation, even when no prolines are present in the sequence. This indicates that the P(II) conformation is a dominant component of the denatured states of proteins, although a quantitative description of the component enthalpy and entropy functions associated with this conformation (i.e., the thermodynamic mechanism) has thus far proven elusive. An experimental system has been designed that, when analyzed with high-precision isothermal titration calorimetry, provides direct access to the residue-specific thermodynamics of the P(II) structure formation in disordered proteins and peptides. Here, it is shown that the P(II) bias is driven by a favorable and significant enthalpy (Deltah) of -1.7 kcal mol(-1) residue(-1), which is partially offset by an unfavorable entropy (TDeltas) of -0.7 kcal mol(-1) residue(-1), relative to the ensemble of disordered conformations of the molecule. In addition to impacting dramatically the interpretation of thermal denaturation experiments, these experimental values form the framework of a quantitative energetic description of the denatured states of proteins. PMID- 15274634 TI - Solid-state NMR investigation of the selective perturbation of lipid bilayers by the cyclic antimicrobial peptide RTD-1. AB - RTD-1 is a cyclic beta-hairpin antimicrobial peptide isolated from rhesus macaque leukocytes. Using (31)P, (2)H, (13)C, and (15)N solid-state NMR, we investigated the interaction of RTD-1 with lipid bilayers of different compositions. (31)P and (2)H NMR of uniaxially oriented membranes provided valuable information about how RTD-1 affects the static and dynamic disorder of the bilayer. Toward phosphatidylcholine (PC) bilayers, RTD-1 causes moderate orientational disorder independent of the bilayer thickness, suggesting that RTD-1 binds to the surface of PC bilayers without perturbing its hydrophobic core. Addition of cholesterol to the POPC membrane does not affect the orientational disorder. In contrast, binding of RTD-1 to anionic bilayers containing PC and phosphatidylglycerol lipids induces much greater orientational disorder without affecting the dynamic disorder of the membrane. These correlate with the selectivity of RTD-1 for anionic bacterial membranes as opposed to cholesterol-rich zwitterionic mammalian membranes. Line shape simulations indicate that RTD-1 induces the formation of micrometer-diameter lipid cylinders in anionic membranes. The curvature stress induced by RTD-1 may underlie the antimicrobial activity of RTD-1. (13)C and (15)N anisotropic chemical shifts of RTD-1 in oriented PC bilayers indicate that the peptide adopts a distribution of orientations relative to the magnetic field. This is most likely due to a small fraction of lipid cylinders that change the RTD-1 orientation with respect to the magnetic field. Membrane-bound RTD-1 exhibits narrow line widths in magic-angle spinning spectra, but the sideband intensities indicate rigid-limit anisotropies. These suggest that RTD-1 has a well-defined secondary structure and is likely aggregated in the membrane. These structural and dynamical features of RTD-1 differ significantly from those of PG 1, a related beta-hairpin antimicrobial peptide. PMID- 15274635 TI - Influence of minor groove substituents on the structure of DNA Holliday junctions. AB - The inosine-containing sequence d(CCIGTACm(5)CGG) is shown to crystallize as a four-stranded DNA junction. This structure is nearly identical to the antiparallel junction formed by the parent d(CCGGTACm(5)()CGG) sequence [Vargason, J. M., and Ho, P. S. (2002) J. Biol. Chem. 277, 21041-21049] in terms of its conformational geometry, and inter- and intramolecular interactions within the DNA and between the DNA and solvent, even though the 2-amino group in the minor groove of the important G(3).m(5)C(8) base pair of the junction core trinucleotide (italicized) has been removed. In contrast, the analogous 2,6 diaminopurine sequence d(CCDGTACTGG) crystallizes as resolved duplex DNAs, just like its parent sequence d(CCAGTACTGG) [Hays, F. A., Vargason, J. M., and Ho, P. S. (2003) Biochemistry 42, 9586-9597]. These results demonstrate that it is not the presence or absence of the 2-amino group in the minor groove of the R(3).Y(8) base pair that specifies whether a sequence forms a junction, but the positions of the extracyclic amino and keto groups in the major groove. Finally, the study shows that the arms of the junction can accommodate perturbations to the B-DNA conformation of the stacked duplex arms associated with the loss of the 2-amino substituent, and that two hydrogen bonding interactions from the C(7) and Y(8) pyrimidine nucleotides to phosphate oxygens of the junction crossover specify the geometry of the Holliday junction. PMID- 15274636 TI - Design and synthesis of de novo cytochromes c. AB - Natural c-type cytochromes are characterized by the consensus Cys-X-X-Cys-His heme-binding motif (where X is any amino acid) by which the heme is covalently attached to protein by the addition of the sulfhydryl groups of two cysteine residues to the vinyl groups of the heme. In this work, the consensus sequence was used for the heme-binding site of a designed four-helix bundle, and the apoproteins with either a histidine residue or a methionine residue positioned at the sixth coordination site were synthesized and reacted with iron protoporphyrin IX (protoheme) under mild reducing conditions in vitro. These polypeptides bound one heme per helix-loop-helix monomer via a single thioether bond and formed four helix bundle dimers in the holo forms as designed. They exhibited visible absorption spectra characteristic of c-type cytochromes, in which the absorption bands shifted to lower wavelengths in comparison with the b-type heme binding intermediates of the same proteins. Unexpectedly, the designed cytochromes c with bis-His-coordinated heme iron exhibited oxidation-reduction potentials similar to those of their b-type intermediates, which have no thioether bond. Furthermore, the cytochrome c with His and Met residues as the axial ligands exhibited redox potentials increased by only 15-30 mV in comparison with the cytochrome with the bis-His coordination. These results indicate that highly positive redox potentials of natural cytochromes c are not only due to the heme covalent structure, including the Met ligation, but also due to noncovalent and hydrophobic environments surrounding the heme. The covalent attachment of heme to the polypeptide in natural cytochromes c may contribute to their higher redox potentials by reducing the thermodynamic stability of the oxidized forms relatively against that of the reduced forms without the loss of heme. PMID- 15274637 TI - Different conformational changes within the F-helix occur during serpin folding, polymerization, and proteinase inhibition. AB - The intrinsic metastability of the serpin native state is the thermodynamic driving force for both proteinase inhibition and the formation of inactive polymers. A number of mechanisms has been proposed to explain how both these conformational changes are achieved. However, one aspect that has received little attention is the movement of the F-helix, which physically impedes both these events. We have applied a protein engineering approach to investigate the conformational changes of this helix during proteinase inhibition, serpin folding, and polymerization. We systematically mutated two highly conserved hydrophobic residues on the F-helix, V161 and I157, and in addition, removed a hydrogen bond between D149 and the first turn of the helix. Our data demonstrate that while all three interactions are important for the stability and folding of the molecule, their contribution during inhibition and polymerization differ. The presence of I157 is crucial to all conformational changes as its loss results in inactivation of the serpin and rapid polymerization. The replacement of D149 does not affect activity but significantly increases the polymerization rate. The interactions formed by V161 play an important role only in maintaining the native conformation. Taken together, these data suggest that the F-helix undergoes a reversible conformational change in both its N- and C-termini during proteinase inhibition only the C-terminus undergoes changes during polymerization, but there is a global change required for folding. PMID- 15274638 TI - Thermodynamic properties of nucleotide reductase reactions. AB - Recent thermodynamic measurements have made it possible to calculate the apparent equilibrium constants of the ribonucleoside diphosphate reductase reaction and the ribonucleoside triphosphate reductase reaction with various reducing agents. Third law heat capacity measurements on crystals of d-ribose and other calorimetric measurements make it possible to calculate Delta(f)G degrees for D ribose and two species of D-ribose 5-phosphate. The experimental value of the apparent equilibrium constant K' for the deoxyribose-phosphate aldolase reaction makes it possible to calculate the standard Gibbs energies of formation Delta(f)G degrees for two protonation states of 2'-deoxy-D-ribose 5-phosphate. This shows that Delta(f)G degrees (2'-deoxy-D-ribose 5-phosphate(2)(-)) - Delta(f)G degrees (D-ribose 5-phosphate(2)(-)) = 147.86 kJ mol(-1) at 298.15 K and zero ionic strength in dilute aqueous solutions. This difference between reduced and oxidized forms is expected to apply to D-ribose, D-ribose 1-phosphate, ribonucleosides, and ribonucleotides in general. This expectation is supported by two other enzyme-catalyzed reactions for which apparent equilibrium constants have been determined. The availability of Delta(f)G degrees values for the species of 2'-deoxy-D-ribose and its derivatives makes it possible to calculate standard transformed Gibbs energies of formation of these reactants, apparent equilibrium constants for their reactions, changes in the binding of hydrogen ions in these reactions, and standard apparent reduction potentials of the half reactions involved as a function of pH and ionic strength at 298.15 K. The apparent equilibrium constant for ADP + thioredoxin(red) = 2'-deoxyADP + H(2)O + thioredoxin(ox) is 1.4 x 10(11) at 298.15 K, pH 7, and 0.25 M ionic strength. PMID- 15274639 TI - Domain-domain interactions in the aminoglycoside antibiotic resistance enzyme AAC(6')-APH(2''). AB - The most common determinant of aminoglycoside antibiotic resistance in Gram positive bacterial pathogens, such as Staphylococcus aureus, is a modifying enzyme, AAC(6')-APH(2' '), capable of acetylating and phosphorylating a wide range of antibiotics. This enzyme is unique in that it is composed of two separable modification domains, and although a number of studies have been conducted on the acetyltransferase and phosphotransferase activities in isolation, little is known about the role and impact of domain interactions on antibiotic resistance. Kinetic analysis and in vivo assessment of a number of N- and C-terminal truncated proteins have demonstrated that the two domains operate independently and do not accentuate one another's resistance activity. However, the two domains are structurally integrated, and mutational analysis has demonstrated that a predicted connecting alpha-helix is especially critical for maintaining proper structure and function of both activities. AAC(6')-APH(2' ') detoxifies a staggering array of aminoglycosides, where one or both activities make important contributions depending on the antibiotic. Thus, to overcome antibiotic resistance associated with AAC(6')-APH(2' '), aminoglycosides resistant to modification and/or inhibitors against both activities must be employed. Domain-domain interactions in AAC(6')-APH(2' ') offer a unique target for inhibitor strategies, as we show that their disruption simultaneously inhibits both activities >90%. PMID- 15274640 TI - Post-translational modifications of aquaporin 0 (AQP0) in the normal human lens: spatial and temporal occurrence. AB - Because of the lack of protein turnover in fiber cells of the ocular lens, Aquaporin 0 (AQP0), the most abundant membrane protein in the lens, undergoes extensive post-translational modification with fiber cell age. To map the distribution of modified forms of AQP0 within the lens, normal human lenses ranging in age from 34 to 38 were concentrically dissected into several cortical and nuclear sections. Membrane proteins still embedded in the membranes were digested with trypsin, and the resulting C-terminal peptides of AQP0 were analyzed by HPLC tandem mass spectrometry, permitting the identification of modifications and estimation of their abundance. Consistent with earlier reports, the major phosphorylation site was Ser 235, and the major sites of backbone cleavage occurred at residues 246 and 259. New findings suggest that cleavage at these sites may be a result of nonenzymatic truncation at asparagine residues. In addition, this approach revealed previously undetected sites of truncation at residues 249, 260, 261, and 262; phosphorylation at Ser 231 and to a lower extent at Ser 229; and racemization/isomerization of l-Asp 243 to d-Asp and d-iso-Asp. The spatial distribution of C-terminally modified AQP0 within the lens indicated an increase in truncation and racemization/isomerization with fiber cell age, whereas the level of Ser 235 phosphorylation increased from the outer to inner cortex but decreased in the nucleus. Furthermore, the remarkably similar pattern and distribution of truncation products from lenses from three donors suggest specific temporal mechanisms for the modification of AQP0. PMID- 15274641 TI - A disubstituted succinamide is a potent sodium channel blocker with efficacy in a rat pain model. AB - Sodium channel blockers are used clinically to treat a number of neuropathic pain conditions, but more potent and selective agents should improve on the therapeutic index of currently used drugs. In a high-throughput functional assay, a novel sodium channel (Na(V)) blocker, N-[[2'-(aminosulfonyl)biphenyl-4 yl]methyl]-N'-(2,2'-bithien-5-ylmethyl)succinamide (BPBTS), was discovered. BPBTS is 2 orders of magnitude more potent than anticonvulsant and antiarrhythmic sodium channel blockers currently used to treat neuropathic pain. Resembling block by these agents, block of Na(V)1.2, Na(V)1.5, and Na(V)1.7 by BPBTS was found to be voltage- and use-dependent. BPBTS appeared to bind preferentially to open and inactivated states and caused a dose-dependent hyperpolarizing shift in the steady-state availability curves for all sodium channel subtypes tested. The affinity of BPBTS for the resting and inactivated states of Na(V)1.2 was 1.2 and 0.14 microM, respectively. BPBTS blocked Na(V)1.7 and Na(V)1.2 with similar potency, whereas block of Na(V)1.5 was slightly more potent. The slow tetrodotoxin-resistant Na(+) current in small-diameter DRG neurons was also potently blocked by BPBTS. [(3)H]BPBTS bound with high affinity to a single class of sites present in rat brain synaptosomal membranes (K(d) = 6.1 nM), and in membranes derived from HEK cells stably expressing Na(V)1.5 (K(d) = 0.9 nM). BPBTS dose-dependently attenuated nociceptive behavior in the formalin test, a rat model of tonic pain. On the basis of these findings, BPBTS represents a structurally novel and potent sodium channel blocker that may be used as a template for the development of analgesic agents. PMID- 15274642 TI - Substrate specificity and kinetic mechanism of the Sir2 family of NAD+-dependent histone/protein deacetylases. AB - The Silent information regulator 2 (Sir2) family of enzymes consists of NAD(+) dependent histone/protein deacetylases that tightly couple the hydrolysis of NAD(+) and the deacetylation of an acetylated substrate to form nicotinamide, the deacetylated product, and the novel metabolite O-acetyl-ADP-ribose (OAADPR). In this paper, we analyzed the substrate specificity of the yeast Sir2 (ySir2), the yeast HST2, and the human SIRT2 homologues toward various monoacetylated histone H3 and H4 peptides, determined the basic kinetic mechanism, and resolved individual chemical steps of the Sir2 reaction. Using steady-state kinetic analysis, we have shown that ySir2, HST2, and SIRT2 exhibit varying catalytic efficiencies and display a preference among the monoacetylated peptide substrates. Bisubstrate kinetic analysis indicates that Sir2 enzymes follow a sequential mechanism, where both the acetylated substrate and NAD(+) must bind to form a ternary complex, prior to any catalytic step. Using rapid-kinetic analysis, we have shown that after ternary complex formation, nicotinamide cleavage occurs first, followed by the transfer of the acetyl group from the donor substrate to the ADP-ribose portion of NAD(+) to form OAADPr and the deacetylated product. Product and dead-end inhibition analyses revealed that nicotinamide is the first product released followed by random release of OAADPr and the deacetylated product. PMID- 15274643 TI - Role of peptide sequence and neighboring residue glycosylation on the substrate specificity of the uridine 5'-diphosphate-alpha-N-acetylgalactosamine:polypeptide N-acetylgalactosaminyl transferases T1 and T2: kinetic modeling of the porcine and canine submaxillary gland mucin tandem repeats. AB - A large family of uridine 5'-diphosphate (UDP)-alpha-N-acetylgalactosamine (GalNAc):polypeptide N-acetylgalactosaminyl transferases (ppGalNAc Ts) initiates mucin-type O-glycan biosynthesis at serine and threonine. The peptide substrate specificities of individual family members are not well characterized or understood, leaving an inability to rationally predict or comprehend sites of O glycosylation. Recently, a kinetic modeling approach demonstrated neighboring residue glycosylation as a major factor modulating the O-glycosylation of the porcine submaxillary gland mucin 81 residue tandem repeat by ppGalNAc T1 and T2 [Gerken et al. (2002) J. Biol. Chem. 277, 49850-49862]. To confirm the general applicability of this model and its parameters, the ppGalNAc T1 and T2 glycosylation kinetics of the 80+ residue tandem repeat from the canine submaxillary gland mucin was obtained and characterized. To reproduce the glycosylation patterns of both mucins (comprising 50+ serine/threonine residues), specific effects of neighboring peptide sequence, in addition to the previously described effects of neighboring residue glycosylation, were required of the model. Differences in specificity of the two transferases were defined by their sensitivities to neighboring proline and nonglycosylated hydroxyamino acid residues, from which a ppGalNAc T2 motif was identified. Importantly, the model can approximate the previously reported ppGalNAc T2 glycosylation kinetics of the IgA1 hinge domain peptide [Iwasaki, et al. (2003) J. Biol. Chem. 278, 5613-5621], further validating both the approach and the ppGalNAc T2 positional weighting parameters. The characterization of ppGalNAc transferase specificity by this approach may prove useful for the search for isoform-specific substrates, the creation of isoform-specific inhibitors, and the prediction of mucin-type O glycosylation sites. PMID- 15274644 TI - Acidic C-tail of HMGB1 is required for its target binding to nucleosome linker DNA and transcription stimulation. AB - HMGB1, a nonhistone chromosomal protein in higher eukaryotic nuclei, consists of two DNA binding motifs called HMG boxes and an acidic C-tail comprising a continuous array of 30 acidic amino acid residues. In the preceding study, we showed that the acidic C-tail of HMGB1 is required for transcription stimulation accompanied by chromatin decondensation in cultured cells. However, details of the involvement of the acidic C-tail in transcription stimulation were not clear. To clarify the mechanism of transcription stimulation by the acidic C-tail, we assessed the effect of the acidic C-tail on the transcription stimulation and nucleosome binding. Transcription stimulation assays using acidic C-tail deletion mutants showed that the five amino acid residues at the C-terminal end of HMGB1, a DDDDE sequence, are essential for the stimulation. The DDDDE sequence was also required for the preferential binding of HMGB1 to nucleosome linker DNA, which is a cognate HMGB1 binding site in chromatin. Cross-linking and far-Western experiments demonstrated that the DDDDE sequence interacts with the core histone H3 N-tail. These results strongly suggest that the interaction between the DDDDE sequence of HMGB1 and the H3 N-tail is a key factor for the transcription stimulation by HMGB1 as well as the preferential binding of HMGB1 to chromatin. PMID- 15274645 TI - Redox characterization of Geobacter sulfurreducens cytochrome c7: physiological relevance of the conserved residue F15 probed by site-specific mutagenesis. AB - The complete genome sequence of the delta-proteobacterium Geobacter sulfurreducens reveals a large abundance of multiheme cytochromes. Cytochrome c(7), isolated from this metal ion-reducing bacterium, is a triheme periplasmic electron-transfer protein with M(r) 9.6 kDa. This protein is involved in metal ion-reducing pathways and shares 56% sequence identity with a triheme cytochrome isolated from the closely related delta-proteobacterium Desulfuromonas acetoxidans (Dac(7)). In this work, two-dimensional NMR was used to monitor the heme core and the general folding in solution of the G. sulfurreducens triheme cytochrome c(7) (PpcA). NMR signals obtained for the three hemes of PpcA at different stages of oxidation were cross-assigned to the crystal structure [Pokkuluri, P. R., Londer, Y. Y., Duke, N. E. C., Long, W. C., and Schiffer, M. (2004) Biochemistry 43, 849-859] using the complete network of chemical exchange connectivities, and the order in which each heme becomes oxidized was determined at pH 6.0 and 8.2. Redox titrations followed by visible spectroscopy were also performed in order to monitor the macroscopic redox behavior of PpcA. The results obtained showed that PpcA and Dac(7) have different redox properties: (i) the order in which each heme becomes oxidized is different; (ii) the reduction potentials of the heme groups and the global redox behavior of PpcA are pH dependent (redox-Bohr effect) in the physiological pH range, which is not observed with Dac(7). The differences observed in the redox behavior of PpcA and Dac(7) may account for the different functions of these proteins and constitute an excellent example of how homologous proteins can perform different physiological functions. The redox titrations followed by visible spectroscopy of PpcA and two mutants of the conserved residue F15 (PpcAF15Y and PpcAF15W) lead to the conclusion that F15 modulates the redox behavior of PpcA, thus having an important physiological role. PMID- 15274646 TI - Thermodynamic stability of DNA tandem mismatches. AB - The thermodynamics of nine hairpin DNAs were evaluated using UV-monitored melting curves and differential scanning calorimetry (DSC). Each DNA has the same five base loop and a stem with 8-10 base pairs. Five of the DNAs have a tandem mismatch in the stem, while four have all base pairs. The tandem mismatches examined (ga/ga, aa/gc, ca/gc, ta/ac, and tc/tc) spanned the range of stability observed for this motif in a previous study of 28 tandem mismatches. UV-monitored melting curves were obtained in 1.0 M Na(+), 0.1 M Na(+), and 0.1 M Na(+) with 5 mM Mg(2+). DSC studies were conducted in 0.1 M Na(+). Transition T(m) values were unchanged over a 50-fold range of strand concentration. Model-independent enthalpy changes (DeltaH degrees ) evaluated by DSC were in good agreement (+/ 8%) with enthalpy values determined by van't Hoff analyses of the melting curves in 0.1 M Na(+). The average heat capacity change (DeltaC(p)) associated with the hairpin to single strands transitions was estimated from plots of DeltaH degrees and DeltaS degrees with T(m) and ln T(m), respectively, and from profiles of DSC curves. The average DeltaC(p) values (113 +/- 9 and 42 +/- 27 cal x K(-1) x mol( 1) of bp), were in the range of values reported in previous studies. Consideration of DeltaC(p) produced large changes in DeltaH degrees and DeltaS degrees extrapolated from the transition region to 37 degrees C and smaller but significant changes to free energies. The loop free energy of the five tandem mismatches at 37 degrees C varied over a range of approximately 4 kcal x mol(-1) for each solvent. PMID- 15274647 TI - Characterization of liposomal tacrolimus in lung surfactant-like phospholipids and evaluation of its immunosuppressive activity. AB - Tacrolimus (FK506) is a hydrophobic immunosuppressive agent that rapidly penetrates the plasmatic membrane and inhibits the signal transduction cascade of T lymphocytes. The objective of this study was the characterization of liposomal FK506 with surfactant-like phospholipids to be administered intratracheally after lung transplantation or in inflammatory lung diseases. We evaluated the optimal incorporation of FK506 in dipalmitoylphosphatidylcholine (DPPC) and DPPC/1 palmitoyl-2-oleoylphosphatidylglycerol (POPG) monolayers and bilayers and the effects of FK506 on the physical properties of DPPC and DPPC/POPG (8:2 w/w) vesicles. In addition, we assessed the immunosuppressive effects of surfactant like phospholipid vesicles containing different amounts of FK506 on T-cell proliferation and interleukin 2 production. From surface pressure measurements of FK506/DPPC and FK506/DPPC/POPG mixed monolayers, we determined that FK506 was embedded into these monolayers up to an FK506 concentration of about 0.4 mol %. Beyond this concentration, FK506 was not quantitatively incorporated into the monolayer, suggesting possible concentration-dependent aggregation of tacrolimus. The incorporation of FK506 into DPPC monolayers, at concentrations or =377 ng/mL), significantly less often than in 417 subjects with IL-8 below 377 ng/mL (84.7%; P < 0.0001). Anaerobic pathogens were detected in 83.3% of high IL-8 subjects, significantly more often than in normal IL-8 subjects (43.9%; P < 0.0001). By multivariate analysis, cervical IL-8 was significantly high only in subjects without Lactobacillus species; they showed a significantly higher prematurity rate than Lactobacillus-positive subjects. CONCLUSIONS: Absence of vaginal Lactobacilli was associated with increased cervical IL-8 and increased risk of premature delivery. PMID- 15274650 TI - Endometrial and peritoneal macrophages: expression of activation and adhesion molecules. AB - PROBLEM: Macrophages are highly individualized in tissues and their activities are a reflection of systemic and local environmental signals. The expression of activation (CD69, CD71) and adhesion (CD54) molecules on the surface of CD14+ endometrial macrophages at various phases of the menstrual cycle was compared with the cell surface receptors of peritoneal fluid macrophages. METHOD OF STUDY: Two-colour-flow cytometry was used to determine the peritoneal and endometrial macrophage phenotype. RESULTS: Endometrium macrophages expressed a lower level of CD69+ and CD54+ macrophages than peritoneal macrophages. However, CD71 receptors displayed similar expression in both macrophage populations, endometrium and peritoneal, except during the proliferative phase. CONCLUSION: These findings demonstrate the differences between macrophages from endometrium and peritoneal fluid with regard to CD69, CD71 and CD54 expression. In addition, increased numbers of endometrial macrophages in the late secretory phase of the menstrual cycle suggest that they may play a role in menstruation. PMID- 15274651 TI - Effect of hypoxia on urocortin production in human gestational trophoblasts in vitro. AB - PROBLEM: Urocortin is produced by the placenta throughout pregnancy but its regulation remains unknown. The effect of hypoxia on placental urocortin production is not known. The aim of this study was to determine the effect of in vitro hypoxia on human trophoblastic urocortin production. METHOD OF STUDY: Placental explants and primary cultures were incubated in anaerobe hypoxic bags for 24 h in a humidified incubator. Urocortin peptide secretion and mRNA (messenger RNA) production was determined by enzyme-linked immunosorbent assay and reverse transcription-polymerase chain reaction, respectively. Morphological and functional integrity was verified by immunohistochemical analysis of urocortin expression. Vascular endothelial growth factor expression was used to verify the generation of cellular hypoxia in our in vitro system. RESULTS: Hypoxia did not affect urocortin secretion or mRNA expression in explant and single-cell cultures. Production was greater from first trimester than term explants and from single-cell primary cultures more than from explant cultures. CONCLUSIONS: Hypoxia does not influence human placental urocortin secretion or mRNA expression in vitro. PMID- 15274652 TI - The effect of inflammatory cytokines on secretion of macrophage colony stimulating factor and monocyte chemoattractant protein-1 in human granulosa cells. AB - PROBLEM: In order to investigate the role of macrophage colony-stimulating factor (M-CSF) and monocyte chemoattractant protein -1 (MCP-1) in human ovulation, we studied the regulation of M-CSF and MCP-1 in cultured human granulosa cells. METHOD OF STUDY: Immortalized granulosa cells (GC1a) were cultured in serum-free medium, and incubated with interleukin (IL)-1alpha, IL-1 receptor antagonist (ra) and tumor necrosis factor (TNF)-alpha. The supernatants were collected, and M-CSF and MCP-1 were measured by ELISA. RESULTS: The levels of M-CSF and MCP-1 were increased after treatment with IL-1alpha (1 nm) and TNF-alpha (1 nm) in a time dependent manner. The levels of M-CSF and MCP-1 were significantly increased after treatment with IL-1alpha and TNF-alpha in a dose-dependent manner. However, the levels of M-CSF and MCP-1 were significantly decreased by treatment with IL 1alpha (1 nm) and/or increasing concentrations of IL-1 ra. CONCLUSIONS: Our data indicated that M-CSF and MCP-1 were regulated by IL-1alpha and TNF-alpha. It was suggested that M-CSF and MCP-1 may play an important role in human preovulatory processes. PMID- 15274653 TI - Life-threatening bleeding, pregnancy and lupus anticoagulant: success after steroid and anticoagulant therapy. AB - PROBLEM: The clinical manifestations associated with the presence of the lupus anticoagulant (LAC) are usually thrombotic and/or obstetric complications, specially miscarriages and/or repeated fetal deaths. The bleeding episodes in patients with LAC with no other coagulation disorders are exceptional, especially during pregnancy, intra- and postpartum. METHOD OF STUDY: Here we present two cases of patients with classification criteria of primary antiphospholipid antibody syndrome, a history of recurrent complicated pregnancies and massive bleeding during abortion and postpartum. We studied anticardiolipin (IgG/IgM), beta2glicoprotein-1 (IgG) and antimitochondrial (type 5) antibodies, LAC, rapid plasma reagin, coagulation test, clotting factors and, placental vascular flow from the second trimester of their last pregnancies. RESULTS: LAC was repeatedly detected in both cases. The prothrombin time and clotting factors including factors II and XIII were normal. Intrauterine growth restriction was also observed in both patients. Pre-eclampsia was detected in patient 2. We tried enoxaparin (60-80 mg/day) and prednisone (1 mg/kg/day) therapy and no maternal, hemorrhagic or thrombotic episodes were observed. CONCLUSIONS: Exceptionally, LAC may be associated with recurrent, life-threatening bleeding, even in patients with no prothrombin deficiency. The close follow-up and early anticoagulant and steroid therapy seems to be able to control thrombotic and hemorrhagic complications during the pregnancy and postpartum. PMID- 15274654 TI - 11beta-hydroxysteroid dehydrogenase inhibitor carbenoxolone stimulates chorionic gonadotropin secretion from human term cytotrophoblast cells differentiated in vitro. AB - PROBLEM: To investigate the effect of altering local glucocorticoid concentration on human chorionic gonadotropin (hCG) production by cultured placental trophoblast cells. METHOD OF STUDY: Human placental trophoblasts were isolated from fresh placentas. Cytotrophoblasts were purified and placed into 24-well multiplates. For cultivation Dulbecco's modified Eagle's medium (DMEM) with 15 mm HEPES and 15% FBS was used. 11beta-Hydroxysteroid dehydrogenase (11beta-HSD) activity and its inhibition by carbenoxolone (CE) were measured in cultured cells. Cultures were exposed to CE for 16-20 hr. Overnight production of hCG was measured by radioimmunoassay in control and treated cells. RESULTS: The 11beta HSD activity in these cultures was inhibited by nm concentrations of CE, the apparent Ki being 2.5 nm. Inhibition of 11beta-HSD activity with 0.1 nm CE resulted in 1.5-fold increase in the production of hCG. CONCLUSIONS: Increasing local glucocorticoid concentration by the inhibition of 11beta-HSD results in higher hCG secretion, which in turn enhance cell differentiation. PMID- 15274655 TI - Serum P-selectin level during controlled ovarian hyperstimulation--a preliminary report. AB - OBJECTIVE: To measure levels of serum P-selectin in patients undergoing controlled ovarian hyperstimulation (COH) cycles and to determine their possible correlation to COH variables. SETTING: Large university-based infertility and in vitro fertilization unit. PATIENTS: Fourteen consecutive patients undergoing our routine COH protocol for unexplained infertility. INTERVENTIONS AND MAIN OUTCOME MEASURES: Blood was drawn three times during the COH cycle: (1) day 2 or 3 of the menstrual cycle, before gonadotropin treatment (Day-0); (2) day of or prior to human chorionic gonadotropin (hCG) administration (Day-hCG); and (3) day of ovulation (Day-OVU). Serum levels of sex steroids and P-selectin were compared among the three time points. P-selectin was measured with a commercial quantitative sandwich immunoassay technique. To reduce interpatient variability, the percent difference between the Day-0 (non-stimulated, basal) level and the Day-hCG and Day-OVU levels was calculated. RESULTS: P-selectin level on Day-hCG was significantly higher than on Day-0 (P < 0.05) and non-significantly higher than on Day-OVU (P < 0.12). No significant correlations were observed between serum P-selectin and patient age, amount of gonadotropins used, or estradiol or progesterone level. CONCLUSION: The increase in serum P-selectin level during COH until peak estradiol suggests that COH may potentiate a state of platelet activation which is substantially attenuated after hCG administration. PMID- 15274656 TI - Interleukin-4 gene polymorphism is not involved in the risk of recurrent pregnancy loss. AB - PROBLEM: Enhanced secretion of type-2 T-helper (Th2) cytokine is a characteristic feature in normal physiological pregnancy. A study has demonstrated defective production of interleukine-4 (IL-4) and other Th2 cytokine in women with recurrent pregnancy loss (RPL). Several studies have suggested that IL-4 variable number of tandem repeat (VNTR) gene polymorphism is probably associated with different IL-4 production. METHODS OF STUDY: The IL-4 VNTR genotypes were assessed in 109 Japanese women with RPL and 210 ethnically matched women experiencing at least one live birth and no spontaneous abortion. RESULTS: No significant differences in IL-4 VNTR genotype frequencies were found between the RPL and the control [B1B1 genotype (reference); B1/B2 and B2/B2 genotypes, odds ratio, 0.91; 95% confidence interval, 0.58-1.45]. CONCLUSION: The present study suggests that the IL-4 VNTR allele is not a major genetic regulator in RPL. PMID- 15274657 TI - Structural and functional heterogeneity in the CD200R family of immunoregulatory molecules and their expression at the feto-maternal interface. AB - PROBLEM: We have shown that CD200Fc, a chimeric molecule including the extracellular domain of CD200 and a murine immunoglobulin (Ig)G2a Fc region, regulates immune responses and prevents T helper (Th)1 cytokine-triggered spontaneous abortions in mice. CD200 is expressed on a subpopulation of uterine decidua cells and on trophoblast, both in the mouse and human. The receptor(s) for CD200, CD200R(s), was not previously well-characterized. METHODS: 5'-rapid amplification of cDNA ends (RACE), cDNA and genomic DNA clone analysis were used to identify a family of CD200Rs on mouse chromosome 16, juxtaposed to the CD200 gene, named CD200R1, R2, R3, and R4. Northern blot and reverse transcriptase polymerase chain reaction (RT-PCR) analysis was used to detect expression of different CD200R subtypes in different organs. Rabbit polyclonal and rat monoclonal antibodies (mAbs) to CD200R isoforms was used for fluorescence activated cell sorter (FACS) analysis, to test for immunomodulatory effects on allogeneic mixed-lymphocyte responses in vitro, and for immunohistochemistry. RESULTS: The CD200Fc was able to interact physically with each of the CD200Rs expressed on the cell surface. Northern blot and RT-PCR analyses indicated distinct patterns of CD200R isoform mRNA expression in different tissues and FACS analyses confirmed unique cell- and tissue-specific expression of the different CD200Rs. mAbs directed against the different isoforms modified the development of in vitro alloimmune responses. The addition of anti-CD200R1/R4 elicited immunomodulatory responses in vitro comparable to findings with CD200Fc, but different from the effects of anti-CD200R2-3. CONCLUSIONS: These data provide evidence for a family of CD200R molecules in the mouse genome and defines the existence of previously unrecognized diversity in the CD200/CD200R immunomodulatory gene member family. Although this gene member family is clustered in the genome, the different CD200Rs and CD200 exhibit distinct expression patterns and functional properties. Restricted CD200R isoform expression at the feto-maternal interface suggests CD200:CD200R interactions may serve important function(s) determining the successful outcome of pregnancy. PMID- 15274658 TI - Molecular cloning, expression of testicular transcript abundant in germ cells and immunobiological effects of the recombinant protein. AB - PROBLEM: It has been well documented that antisperm antibodies can be causative factors for infertility. In this report we have identified a protein on human sperm referred as human sperm-associated protein (HSAP) using serum of an immunoinfertile woman; it is thus a sperm-specific protein--a candidate molecule for control of fertility. METHOD OF STUDY: An immunoinfertile woman serum showing head-head sperm agglutination and acrosomal localization, reacted with human sperm protein of apparent molecular weight of 48 kDa on Western blot. Anti-48 kDa antiserum was raised in rabbit by eluting 48 kDa protein and was used to screen the human testis cDNA expression library. A putative positive hsap cDNA clone was obtained, sequenced and subjected to tissue specificities studies by Northern blotting. The cell type-specific expression was done using in situ RNA hybridization studies. To obtain recombinant HSAP (r-HSAP), hsap cDNA was cloned in pET 22b(+) expression vector. r-HSAP was expressed as polyhistidine fusion protein in Escherichia coli and purified. Rabbits were immunized with the purified r-HSAP, which led to generation of antibodies. In order to evaluate in vitro immunocontraceptive potential, the anti-r-HSAP antibodies were characterized by agglutination assay, zona-free hamster egg penetration assay, indirect immunofluorescence (IIF) assay, and by flow cytometry analysis. RESULTS: We have cloned a human testis gene encoding a protein (HSAP) of 328 amino acids. Antibodies against the purified recombinant protein specifically recognized approximately 40 kDa r-HSAP, and a cognate 48 kDa protein band in human sperm extract in Western blot procedure. The anti-r-HSAP antibodies localized acrosomal compartment, inhibited sperm binding/attachment in zona-free hamster penetration assay and revealed surface binding with human live sperm by flow cytometry. The cDNA sequence has been submitted to EMBL and has been given the accession number Y16676. CONCLUSION: This study has put in evidence that novel sperm-specific r HSAP has role in sperm function and may have application in the development of a contraceptive vaccine. The availability of the recombinant protein will facilitate studies on the assessment of its potential as a contraceptive immunogen. PMID- 15274659 TI - Risk factors and mechanisms of preterm delivery in Malawi. AB - PROBLEM: We examined risk factors and mechanisms of preterm delivery (PTD) in malaria-exposed pregnant women in Blantyre, Malawi. METHOD OF STUDY: The human immunodeficiency virus (HIV), malaria, syphilis, and anemia were assessed in a cross-sectional study of 572 pregnant women. In a nested case-control study, chorioamnionitis (CAM) was examined; tumor necrosis factor (TNF)-alpha, interleukin (IL)-6, IL-8, macrophage inflammatory protein (MIP)-1alpha, monocyte chemotactic protein (MCP)-1, transforming growth factor (TGF)-beta, cortisol, and corticotropin-releasing hormone were measured in placental, maternal and/or cord blood. RESULTS: HIV, infrequent antenatal clinic attendance, low-maternal weight, no intermittent preventive malaria therapy (IPT), and CAM were associated with PTD, while malaria was not. Of the 18 compartmental cytokine measurements, elevations in placental and/or cord IL-6 and IL-8 were associated with both CAM and PTD. In contrast, there was no overlap between the cytokines affected by malaria and those associated with PTD. CONCLUSIONS: The HIV and CAM were the major infections associated with PTD in this study. CAM, but not malaria, causes PTD via its effect on proinflammatory cytokines. PMID- 15274661 TI - Review article: the evolving role of liver biopsy. AB - Liver biopsy is traditionally the 'gold standard' for the evaluation of liver diseases. There are several situations in which its role is being challenged. In hepatitis C, liver biopsy helps assess prognosis and treatment candidacy. An important exception is genotype 2 or 3 because treatment is more likely to succeed and therapy is relatively short in duration. For hepatitis B, liver biopsy gives some prognostic information, but serologic tests and hepatic biochemical tests are the primary determinants of treatment candidacy. Non alcoholic fatty liver disease can be accurately diagnosed without a liver biopsy and, furthermore, there are no specific therapies available. The role of liver biopsy to assess methotrexate-associated hepatotoxicity remains controversial. Finally, patients with focal liver lesions usually do not require biopsy and, in the case of hepatocellular carcinoma, biopsy carries a risk of needle-track seeding. In short, the need for liver biopsy depends on the specific situation and should be performed when there is sufficient uncertainty about diagnosis, severity of disease, prognosis, and treatment decisions. PMID- 15274662 TI - Review article: helicobacter pylori and molecular events in precancerous gastric lesions. AB - Gastric cancer can be divided into intestinal type and diffuse type that differ substantially in epidemiology and pathogenesis. The most important aetiological factor associated both with intestinal and diffuse gastric cancer, is Helicobacter pylori. Exposure of gastric epithelial cells to H. pylori results in an inflammatory reaction with the production of reactive oxygen species and nitric oxide that, in turn, deaminates DNA causing mutations. The complex interplay between H. pylori strain, inflammation and host characteristics may directly promote diffuse type gastric cancer or induce a cascade of morphological events, i.e. atrophy, intestinal metaplasia and dysplasia, finally leading to intestinal type gastric cancer. Two mechanisms, genetic and epigenetic have been held to play a role in the molecular alterations underlying gastric carcinogenesis. The former, comprising changes in the DNA sequence, is irreversible; the latter, involving DNA methylation, is potentially reversible by eliminating the triggering agents. If H. pylori is eradicated before development of stable mutations, the risk of gastric cancer will likely be prevented. Thus, eradication of H. pylori might immediately reduce the risk of diffuse type gastric cancer, whereas prevention of intestinal type gastric cancer may be less effective if patients are treated later in the evolution of the carcinogenic process. PMID- 15274663 TI - Review article: hepatic hydrothorax. AB - Patients with cirrhosis and portal hypertension often have abnormal extracellular fluid volume regulation, resulting in accumulation of fluid as ascites, oedema or pleural effusion. These complications carry a poor prognosis with nearly half of the patients with ascites dying in the ensuing 2-3 years. In contrast to what happens in the abdominal cavity where large amounts of fluid (5-8 L) accumulate with the patient only experiencing only mild symptoms, in the thoracic cavity smaller amounts of fluid (1-2 L) cause severe symptoms such as shortness of breath, cough and hypoxaemia. Hepatic hydrothorax is defined as a pleural effusion, usually >500 mL, in patients with cirrhosis without cardiopulmonary disease. The pathophysiology involves the direct movement of ascitic fluid from the peritoneal cavity into the pleural space through diaphragmatic defects. The estimated prevalence among cirrhotic patients is 5-10%. The effusion, which is a transudate, most commonly occurs in the right hemithorax. The mainstay of therapy is similar to that of portal hypertensive ascites and includes sodium restriction and administration of diuretics. Refractory hydrothorax can be managed with transjugular intrahepatic portosystemic shunt in selected cases. Pleurodesis is not routinely recommended. Suitable patients with hepatic hydrothorax should be considered candidates for liver transplantation. PMID- 15274664 TI - Four years of treatment with lamivudine: clinical and virological evaluations in HBe antigen-negative chronic hepatitis B. AB - AIM: To evaluate the clinical and virological impact of the prolonged use of lamivudine in 94 patients with HBe antigen-negative chronic hepatitis B. METHODS: Initial virological and biochemical responses were obtained in 84 (89%) and in 83 (88%) patients respectively. RESULTS: The virological response peaked within the first 12 months, but diminished to 39% at 48 months because of drug resistance. Overall a virological breakthrough developed in 44 patients (52.4%). After virological breakthrough, the actuarial probability of maintaining biochemical remission diminished to 15% at 24 months and 0% at 29 months. There was no response in 10.6%. Polymerase gene mutations were observed in 82.5% of virological breakthroughs but also in 75% of the non-responders. Overall 7.4% of patients developed a hepatocellular carcinoma. CONCLUSION: Almost 90% of patients responded initially to lamivudine but the emergence of drug resistance progressively reduced the rate of virological remission to 39% at the fourth year of therapy. YMDD mutants explained the 75% of lamivudine resistances and were also selected very early in non-responders. Although the biochemical response is invariably lost within 29 months of the YMDD mutant's duration, the clinical outcome was benign despite severe postvirological breakthrough hepatitic flares in about 12% of cases. PMID- 15274665 TI - Increased thrombin generation and circulating levels of tumour necrosis factor alpha in patients with chronic Helicobacter pylori-positive gastritis. AB - BACKGROUND: Conflicting data have been reported concerning the relationship between Helicobacter pylori infection and coronary heart disease. AIM: To evaluate clotting system activation and plasma levels of tumour necrosis factor alpha, a procoagulant cytokine, in patients with H. pylori-positive and -negative gastritis. METHODS: Three groups of patients were identified: 38 with H. pylori positive gastritis, 18 with H. pylori-negative gastritis, and 40 H. pylori negative controls with normal gastric mucosa. Plasma levels of prothrombin fragment 1 + 2 (F1 + 2) and tumour necrosis factor-alpha were assayed. Patients were also controlled after 2 and 6 months following standard H. pylori eradication treatment. RESULTS: At baseline, fragment 1 + 2 and tumour necrosis factor-alpha levels in H. pylori-positive patients were significantly higher than those in H. pylori-negative patients with gastritis (P < 0.05 and P < 0.01, respectively). After H. pylori eradication, fragment 1 + 2 and tumour necrosis factor-alpha levels showed a significant decrease at 2 months (P = 0.03 and P = 0.02, respectively) and a further reduction at 6 months, reaching levels observed in H. pylori-negative patients and controls. CONCLUSIONS: The increase thrombin generation rate and the correlation of plasma fragment 1 + 2 and tumour necrosis factor-alpha levels in H. pylori-positive patients suggest a role for inflammation in mediating the relationship between H. pylori infection and activation of the clotting system. PMID- 15274666 TI - Helicobacter pylori eradication in children and adolescents by a once daily 6-day treatment with or without a proton pump inhibitor in a double-blind randomized trial. AB - AIM: To evaluate two simplified Helicobacter pylori eradication treatment alternatives for children and adolescents. METHODS: Study subjects were identified by enzyme-linked immunosorbent assay and immunoblot in a family screening project. Helicobacter pylori infected 10-21 year olds were offered treatment, individuals with abdominal pain underwent upper endoscopy and those with peptic ulcers were excluded. Participants were randomized to either azithromycin 500 mg daily and tinidazole 500 mg two tablets daily in combination with lansoprasole 30 mg daily for 6 days (ATL-group) or with placebo (ATP-group). Urea Breath Test was performed at inclusion and after a minimum of 6 weeks after end of therapy. RESULTS: In total, 131 individuals were randomized, of whom 31 (24%) had undergone upper endoscopy. Full compliance was achieved in 93% (122 of 131). The intention-to-treat eradication rate was 67% (44 of 66) and 58% (38 of 65) for the ATL- and the ATP-group, respectively. CONCLUSION: The double-blind randomized clinical trial did not identify a simplified, successful once daily H. pylori treatment for children and adolescents. Thus, twice daily proton pump inhibitor (PPI)-based triple therapies for 7 days remain as the choice of treatment in children. Further, powerful and controlled studies are needed to elucidate the best treatment strategies for H. pylori eradication in this age group. PMID- 15274667 TI - Tumour necrosis factor-alpha receptor 1 and 2 polymorphisms in inflammatory bowel disease and their association with response to infliximab. AB - BACKGROUND: The role of tumour necrosis factor-alpha in the pathogenesis of inflammatory bowel disorders is well-known and is underscored by the effectiveness of antitumour necrosis factor-alpha treatment. Tumour necrosis factor-alpha exerts its effect by binding TNFR1 and TNFR2, which genes map to inflammatory bowel disorders susceptibility loci. AIMS AND METHODS: Since TNFR1 and TNFR2 are good candidate genes for inflammatory bowel disorders, we studied the functional TNFR2T587G and the TNFR1A36G mutation in 344 Crohn's disease and 152 ulcerative colitis patients and investigated the relation with disease phenotypes. An association with response to infliximab was evaluated in 166 Crohn's disease patients. RESULTS: The TNFR2 587G allele was more frequent in ulcerative colitis compared with controls (P = 0.03). Both single nucleotide polymorphisms were negatively associated with smoking at diagnosis in Crohn's disease (TNFR1A36G odds ratio: 0.614, 95% confidence interval: 0.452, 0.99 and TNFR2T587G odds ratio: 0.572, 95% confidence interval: 0.820, 0.875). There was a positive association between pancolitis and the TNFR1A36G polymorphism in ulcerative colitis (odds ratio: 5.341, 95% confidence interval: 1.484, 19.39). The biological response to infliximab was lower in patients carrying TNFR1 36G (odds ratio: 0.47, 95% confidence interval: 0.234, 0.946). CONCLUSION: The TNFR2 587G allele was more frequent in ulcerative colitis. Both single nucleotide polymorphisms were negatively associated with smoking in Crohn's disease. A relation between TNFR1A36G and pancolitis was found in ulcerative colitis. There was no clear effect of the polymorphisms on infliximab response although, the TNFR1 minor was associated with a lower response to infliximab. PMID- 15274668 TI - Health-related quality of life of patients with gastrointestinal symptoms. AB - BACKGROUND: Little is known about the variation in health-related quality of life among patients with different presentations of gastrointestinal symptoms. AIM: To study the association between health-related quality of life and presentations of gastrointestinal symptoms. METHODS: Health-related quality of life and demographic information was obtained from 873 patients referred to the hospital for endoscopy, using a questionnaire. RESULTS: A total of 436 patients (50%) reported predominantly upper gastrointestinal symptoms, 344 (39%) predominantly lower symptoms, and 93 (11%) patients reported both upper and lower symptoms. Patients with mild, moderate and severe symptoms, reported mean scores on a 100 point visual analogue scale (95% CI) of 90 (79-100), 75 (64-86) and 64 (53-76), respectively (P < 0.001). Mean visual analogue scale scores (95% CI) almost linearly declined from 81 (77-85) to 49 (46-52) for those with one to those with more than eight symptoms. Patients who reported upper gastrointestinal symptoms and in particular epigastric pain, bloating and vomiting had significantly impaired health status in comparison to patients without these symptoms (P < 0.05). CONCLUSIONS: Severity of gastrointestinal symptoms is the most important factor in affecting health status, followed by the numbers and type of gastrointestinal symptoms. PMID- 15274669 TI - Economic analysis of strategies in the prevention of non-steroidal anti inflammatory drug-induced complications in the gastrointestinal tract. AB - BACKGROUND: It is unclear what the best therapeutic approach is in patients who require non-steroidal anti-inflammatory drugs. In clinical practice, choice of prescriptions are often based on drug costs. AIM: To evaluate costs per upper gastrointestinal bleeding avoided with different prevention strategies. METHODS: Two major strategies have been considered (coxibs vs. non-steroidal anti inflammatory drugs plus generic/brand gastroprotective agent). The number of patients needed to treat to prevent a bleeding event, the cost of the drug and duration of treatment were used to estimate costs. RESULTS: Based on hospitalization costs of a bleeding event, no therapeutic strategy is cost effective in patients without risk factors. All strategies (including omeprazole + coxib) are cost-effective in patients with bleeding ulcer history. With other risk factors, all strategies are cost-effective but prevention of events is twice as expensive in patients <75 years of age. No strategy shows superiority unless the cheapest generics are prescribed or a 50% reduction in the incidence of lower gastrointestinal complications with coxibs is confirmed. CONCLUSIONS: Current prevention strategies to reduce serious non-steroidal anti-inflammatory drug associated gastrointestinal events are only cost-effective in patients with risk factors. No strategy shows superiority, but coxib strategy would be more cost effective if it were associated with a reduction of events of the lower gastrointestinal tract. PMID- 15274670 TI - Effect of the motilin agonist KC 11458 on gastric emptying in diabetic gastroparesis. AB - BACKGROUND: KC 11458, a motilin agonist without antibiotic properties, accelerates gastric emptying in animals and healthy humans. AIM: To evaluate the acute effects of KC 11458 on gastric emptying in diabetic gastroparesis. METHODS: Twenty-nine patients (6 type 1 and 23 type 2) with gastroparesis underwent assessments of: (i) gastric emptying of a solid/liquid meal using scintigraphy, (ii) glycaemic control (blood glucose at 0, 30, 60, 90 and 120 min during the gastric emptying measurement) and (iii) upper gastrointestinal and 'meal-related' symptoms (questionnaire), at baseline and after treatment with KC 11458 in a dose of 8 mg t.d.s., or placebo for 8 days. RESULTS: KC 11458 had no statistically significant or clinically relevant effect on gastric emptying of either the solid intragastric retention at 100 min (T100) (P = 0.87) or liquid 50% emptying time (T50) (P = 0.17) components of the meal. KC 11458 slightly worsened (P = 0.04) upper gastrointestinal symptoms when compared with placebo. The magnitude of the change in solid gastric emptying correlated with the change in the blood glucose concentration (r = 0.49; P < 0.05). CONCLUSIONS: KC 11458, in a dose of 8 mg t.d.s. for 8 days, does not accelerate gastric emptying in patients with diabetic gastroparesis. The absence of efficacy may relate to an effect of hyperglycaemia. PMID- 15274671 TI - Prevalence of irritable bowel syndrome according to different diagnostic criteria in a non-selected adult population. AB - BACKGROUND: Prevalence of irritable bowel syndrome shows great variation among epidemiological studies, which may be due to different diagnostic criteria. AIM: To assess prevalence of irritable bowel syndrome according to various diagnostic criteria and to study differences in symptom severity, psychopathology, and use of health care resources between subjects fulfilling different diagnostic criteria. METHODS: A questionnaire was mailed to 5000 randomly selected adults. Presence of irritable bowel syndrome was assessed by four diagnostic criteria: Manning 2 (at least two Manning symptoms), Manning 3 (at least three Manning symptoms), Rome I and Rome II. RESULTS: Response rate was 73%. Prevalence of irritable bowel syndrome by Manning 2, Manning 3, Rome I and Rome II criteria was 16.2%, 9.7%, 5.6%, and 5.1% respectively. Of those fulfilling Rome II criteria, 97% fulfilled Manning 2. Severe or very severe abdominal pain was reported by 27 30% of Manning-positive subjects, and 44% of Rome-positives. Prevalence of depression in Manning 2, and Rome II groups was 30.6 and 39.3%. CONCLUSIONS: Prevalence of irritable bowel syndrome by Rome II criteria is considerably lower than by Manning criteria. Subjects fulfilling Rome criteria form a subgroup of Manning-positive subjects with more severe abdominal symptoms, more psychopathology, and more frequent use of the health care system. PMID- 15274672 TI - In vivo evaluation of a novel pH- and time-based multiunit colonic drug delivery system. AB - BACKGROUND: Targeted drug delivery to the colon is important for topical treatment of inflammatory bowel diseases. Established targeting systems predominantly focus on either pH- or time-dependent release, or bacterial degradation. AIM: To perform a three-phase, crossover design trial evaluating a novel combined pH- and time-based multiunit delivery system. METHODS: Twelve healthy male volunteers each received 200 mg of caffeine as either uncoated immediate release tablets, coated pellets with pH-dependent rapid release (EUDRAGIT FS 30D), and pellets with pH- and time-based release (inner layer EUDRAGIT RL/RS 30D; outer layer EUDRAGIT FS 30D). Orocecal transit time was measured using lactose-[13C]ureide. Serum concentrations of caffeine were measured by high-performance liquid chromatography. RESULTS: In contrast to the uncoated tablet, both coated systems reached the ileocecal region almost at the same time (3.19 +/- 0.71 and 3.33 +/- 0.81 h). Serum caffeine profiles were significantly prolonged for the pH and time delivery system compared with the pH only based system (median tmax 12.0 vs. 5.5 h; P < 0.001). This was further reflected by a lower Cmax value and a lower area under the curve within 24 h after application. CONCLUSION: Compared with the conventional delivery systems, drug release from the new dosage form may offer a new dimension for the oral treatment of mid to distal ulcerative colitis. PMID- 15274673 TI - A new questionnaire for constipation and faecal incontinence. AB - BACKGROUND: The prevalence, severity and risk factors of faecal incontinence in women in the community are incompletely characterized. AIM: To develop and validate a self-report questionnaire (faecal incontinence and constipation assessment) to address these issues. METHOD: Eighty-three women completed the instrument; 20 randomly selected patients answered the faecal incontinence and constipation assessment again 6 weeks later. A gastroenterologist also completed the faecal incontinence and constipation assessment in all 83 subjects after a detailed clinical assessment. Concurrent validity was evaluated by comparing the patient's self-report to a doctor interview for every question. Reproducibility was evaluated by a test-retest approach for every question. The severity of faecal incontinence was rated by incorporating the frequency and type of faecal incontinence, rectal urgency and use of sanitary devices. RESULTS: The questionnaire was well-understood. Reproducibility [median kappa statistic, 0.80 (interquartile range: 0.66-0.90)]; and concurrent validity [0.59 (0.47-0.67)] were acceptable. For the index question on faecal incontinence, the kappa for reproducibility and concurrent validity was 0.90 and 0.95, respectively. The faecal incontinence severity score was also valid (kappa = 0.5). CONCLUSION: The faecal incontinence and constipation assessment has excellent reproducibility and reasonable validity for assessing the presence, risk factors and severity of faecal incontinence and associated bowel disorders in women when compared against clinical assessment. PMID- 15274674 TI - Reproducibility of a symptom response to omeprazole therapy in functional dyspepsia evaluated by a random-starting-day trial design. AB - BACKGROUND: Satisfactory treatment options for functional dyspepsia are lacking. Single subject trial designs may identify subgroups of patients with a uniform response to therapy. AIM: To test reproducibility of response in a new random starting-day trial design developed to identify acid-related symptoms in functional dyspepsia. METHODS: One hundred and nineteen patients with functional dyspepsia completed a 12-day, double-blind random-starting-day trial with an initial placebo run-in followed by switch to omeprazole on a randomized and blinded day (between days 5 and 9) with active treatment continuing for the rest of the trial. Response was defined as a sustained > or =50% reduction of a daily symptom-score within 3 days of active treatment. Fifty-nine patients repeated the random-starting-day trial at relapse of symptoms. RESULTS: After exclusion of placebo responders, 14% (15 of 106) were classified as responders in the first and 20% (10 of 50) in the subsequent random-starting-day trial series. Sixty eight per cent (40 of 59) of the patients reproduced their initial response with a chance-corrected agreement of 0.29. Comparing response patterns using different symptom rating-scales showed good correlation (kappa 0.60). CONCLUSION: Reproducibility of response in a random-starting-day trial was imperfect, mainly because of the low response rates and strict response criteria. Lack of symptom stability impairs the value of the random-starting-day trial and only patients with frequent and stable symptoms should be evaluated in this design. PMID- 15274676 TI - Efficacy and safety of DALI-LDL-apheresis in two patients treated with the angiotensin II-receptor 1 antagonist losartan. AB - Direct adsorption of lipids (DALI) is the first low-density lipoprotein (LDL) apheresis technique capable of adsorbing LDL and lipoprotein (a) directly from whole blood. The adsorber consists of negatively charged polyacrylate ligands linked to a Eupergit matrix. Negatively charged ligands give rise to activation of bradykinin, which is rapidly degraded by the angiotensin converting enzyme (ACE). Thus, angiotensin converting enzyme inhibitors are contraindicated in DALI LDL-apheresis. This is the first paper to describe the efficacy and safety of DALI-LDL-apheresis in patients treated with 50 mg of the angiotensin II-receptor 1 antagonist (ARA) losartan. Two hypercholesterolemic patients were treated for 79 patient months with weekly or biweekly DALI sessions (N = 221 sessions). Approximately 1.4 patient blood volumes were treated per session. Acute reductions of LDL-cholesterol (63%) and lipoprotein (a) (62%) exceeded 60% and laboratory safety parameters remained in the apheresis typical range. Mean bradykinin plasma levels peaked in the efferent line post-adsorber at 1000 mL of treated blood volume; 467 fmol/mL (N = 6 sessions) in the ARA-treated patients and 671 fmol/mL (N = 9 sessions) in a control group of three DALI patients without ARA medication (P = 0.69, n.s.). Clinically, the DALI sessions for the ARA-treated patients were completely uneventful and blood pressure was not significantly different in the two groups. In summary, according to this retrospective pilot study, DALI-LDL-apheresis was shown for the first time to be safe and effective in patients on ARA medication. PMID- 15274677 TI - Autosomal recessive hypercholesterolemia in three sisters with phenotypic homozygous familial hypercholesterolemia: diagnostic and therapeutic procedures. AB - Familial hypercholesterolemia is an autosomal-dominant inherited disorder caused by mutations in the low-density lipoprotein (LDL) receptor gene. The homozygous form is characterized by high-serum LDL cholesterol concentrations, xanthoma formation and premature atherosclerosis. Recently, another molecular defect that also results in severely elevated LDL cholesterol levels was identified: autosomal recessive hypercholesterolemia. This inherited disorder is caused by a mutation in a putative LDL receptor adaptor protein. In our lipid clinic, three sisters with phenotypic homozygous hypercholesterolemia were recently diagnosed as having autosomal recessive hypercholesterolemia. They presented in 1990 with massive tuberous xanthomas at the knees, thighs, elbows and buttocks. LDL receptor and apolipoprotein B gene defects were excluded through mutation analysis. From 1992 onward they underwent LDL-apheresis on a weekly basis. To date the clinical outcome is very satisfying with no evidence of coronary heart disease or aortic valve lesions and almost complete regression of xanthomatosis. PMID- 15274678 TI - Immunoadsorption in lupus myocarditis. AB - Reduction of pathological autoantibodies may be useful in the treatment of systemic lupus erythematosus (SLE). On the other hand clinically manifested myocarditis in SLE, though uncommon, may be life-threatening and its pathogenesis has been ascribed to autoimmunity. The aim of this study is to present a rare case of a patient with severe lupus myocarditis, where immunoadsorption (IA) was evaluated as rescue therapy. A case of SLE with initial manifestation of myocarditis is reported in a 29-year-old male who presented with arthritis, fever, lymphadenopathy, joint swelling and morning stiffness. Laboratory evaluation revealed increased antinuclear antibody (ANA), slightly decreased complement and positive anticoagulant panel. From the above clinical and laboratory features, criteria of SLE seemed applicable. During his hospitalization, the patient developed pericardial effusion and cardiogenic shock. Although pericardiotomy was performed and was treated with immunosuppressive agents, plasmapheresis and supported with current medications, his clinical condition remained critical with an ejection fraction of 20%. At this point it was decided to receive IA onto staphylococcal protein A. After 6 sessions with IA and concomitant immunosuppression, the patient responded well, his condition improved and was dismissed with an ejection fraction of 50%. Fulminant lupus myocarditis is a severe and rare situation lacking a satisfying specific therapy available today. In our presented case, IA in addition to immunosuppressive therapy was beneficial. Considering the benefits of our case and the current knowledge, it might be useful to clarify the open question in scale pilot studies. PMID- 15274679 TI - Polymyxin B-immobilized fiber hemoperfusion after emergency surgery in patients with chronic renal failure. AB - The purpose of this study was to evaluate the effect of direct hemoperfusion using a Polymyxin B (PMX) immobilized fiber column in septic patients with chronic renal failure after emergency surgery. Twenty-four renal failure patients, including 19 dialysis patients, with sepsis or septic shock were treated with direct hemoperfusion after emergency surgery. The 24 consecutive patients included nine with necrotic enterocolitis, six with colonic perforation due to diverticulitis, three with ruptured suture after colectomy, one with duodenal perforation, four with blood access infection, and one with an infected abdominal aortic aneurysm. The acute physiology and chronic health evaluation II score ranged from 13 to 26 (19 +/- 3). After completion of the first and the second hemoperfusion, mean blood pressure was significantly elevated from 69 +/- 12 mm Hg to 89 +/- 15 mm Hg and from 78 +/- 14 mm Hg to 95 +/- 13 mm Hg, respectively (P < 0.01). In addition, the catecholamine dosage needed to maintain the circulation could be decreased markedly after the treatment. The blood concentration of endotoxin in patients with Gram-negative sepsis, before and after the treatment, significantly decreased from 36 +/- 19 pg/mL to 19 +/- 19 pg/mL (P < 0.05). PMX was effective in patients with Gram-positive sepsis as well as Gram-negative sepsis. The 28-day mortality rate in patients who had emergency abdominal surgery was 10% (2/20), whereas that in patients with dialysis access infection was 50% (2/4). There was a significant difference in the Sequential Organ Failure Assessment (SOFA) score of all patients before and after treatment using PMX (9.2 +/- 3.3 vs. 7.5 +/- 3.5, P < 0.05). Furthermore, the SOFA score of survivors decreased significantly after PMX treatment (8.4 +/- 3.5 vs. 6.7 +/- 2.6, P < 0.01). Our results suggest that the early application of PMX may prevent multiple organ failure and improve survival in patients with chronic renal failure and sepsis/septic shock after emergency abdominal surgery, regardless of the type of pathogenic bacteria involved. PMID- 15274680 TI - Clinical evaluation of PMX-DHP for hypercytokinemia caused by septic multiple organ failure. AB - Endotoxin-adsorbing fibers have been applied to treat septic shock patients. The limitations of endotoxin hemoadsorption therapy (PMX-DHP) and the optimal time to start PMX-DHP were examined in patients with septic multiple organ failure with hypercytokinemia (interleukin-6 = 1000 pg/mL). Subjects were separated into those who survived more than 28 days after the start of PMX-DHP therapy (S group) and those who did not (N-S group). Severity of symptoms and background factors, blood biochemical parameters, hemodynamic parameters, PaO(2)/FiO(2), pathogens, endotoxin, cytokines, and vascular endothelial cell function-related markers were examined before and after PMX-DHP. Number of days from onset of shock (or symptom development) to PMX-DHP initiation was longer in the N-S group than in the S group. These results suggest that PMX-DHP could save more lives in patients with septic multiple organ failure with IL-6 = 1000 pg/mL when applied early after the onset of shock. PMID- 15274681 TI - Effect of post-transplant double filtration plasmapheresis on recurrent focal and segmental glomerulosclerosis in renal transplant recipients. AB - In the present study, we reviewed the effect of post-transplant double filtration plasmapheresis (DFPP) on recurrent focal segmental glomerulosclerosis (FSGS) in the transplanted kidney allograft. Sixteen patients with post-transplant recurrent FSGS were enrolled in this study. Out of 16 patients with recurrent FSGS after transplantation, five did not receive DFPP and lost their grafts, while 11 did receive DFPP and four of these patients lost their grafts. Seven patients were able to maintain normal renal function for an average observation period of 57.1 +/- 40.7 months (range 7-125 months). In five patients who had a significant reduction in urinary protein after DFPP, the urinary protein level decreased from 26.60 +/- 23.05 g/day (range 3.34-62.6 g/day) to 2.95 +/- 3.42 g/day (range 0.02-8.64 g/day) and renal function was maintained. The beneficial effects of DFPP on graft outcome were more likely to occur if the patients experienced a marked drop in urinary excretion. Thus, post-transplant DFPP appears to be effective for reducing urinary protein levels and improving long term graft survival. With the small numbers in this trial, however, none of the findings were statistically significant. We recommend the use of post-transplant DFPP to prevent the progression of recurrent FSGS. PMID- 15274682 TI - Measurement of translymphatic fluid absorption using technetium-99m human serum albumin diethylenetriamine pentaacetic acid in continuous ambulatory peritoneal dialysis patients. AB - We have established a new method of measuring translymphatic fluid absorption (TLA) using technetium-99m ((99m)Tc) human serum albumin diethylenetriamine pentaacetic acid ((99m)Tc-HSAD) that can be used commonly in clinical practice. This new method was applied in 13 continuous ambulatory peritoneal dialysis patients (11 males and two females) who had various peritoneal permeability and capacities for peritoneal transport of water. (99m)Tc-HSAD 740MBq was injected in 2 L of peritoneal dialysis fluid with 2.5% glucose, mixed well, and administered intraperitoneally. The fluid was drained extraperitoneally after 4 h and TLA was determined by the in vivo loss of (99m)Tc-HSAD. TLA was 1.41 +/- 1.11 mL/min (mean +/- SD; range, 0.27-3.69 mL/min). The estimated reduction rate by TLA in trans-peritoneally removed fluid ranged from 14.2 to 84.4%, indicating that TLA could have an extremely significant negative effect in some cases on total drainage volume. The present study, using new tracer (99m)Tc-HSAD, could confirm a large individual difference in TLA, indicating TLA as an important contributing factor for fluid-removal failure in continuous ambulatory peritoneal dialysis patients. PMID- 15274683 TI - Lowering of oxidative stress in hemodialysis patients by dialysate cleaning: in relation to arteriosclerosis. AB - The objective of this study was to investigate changes in oxidative stress associated with the cleaning of the dialysate. Thirty-six dialysis patients were studied. Changes in soluble CD-14 (sCD-14), malondialdehyde-low-density lipoprotein (MDA-LDL), and oxidized-LDL (Ox-LDL) were monitored for 1 year before and 1 year after dialysate cleaning. The mean endotoxin (ET) level in the dialysate had previously been confirmed to decrease from 39.0 EU/L to an undetectable level after the cleaning. The mean levels of sCD-14, MDA-LDL, and Ox LDL decreased significantly after the cleaning (sCD-14, P < 0.0001; MDA-LDL, P < 0.001; Ox-LDL, P < 0.001). One year after the cleaning, six cases still showed high levels of MDA-LDL and Ox-LDL. Cardiovascular events occurred in four of those six cases within 2.8 years after the cleaning. These four patients suffered from strong oxidative stress during dialysis, even after the cleaning. We therefore concluded that high levels of MDA-LDL and Ox-LDL are improved in dialysis patients by cleaning of the dialysate. These results indicate that even a dialysate containing 50 EU/L or less ET may stimulate monocytes and cause oxidative stress. They also suggest that even low levels of ET may aggravate arteriosclerosis in dialysis patients. Thus, in order to prevent cardiovascular events in dialysis patients, it is necessary to purify the dialysate. PMID- 15274684 TI - Comparison of the effects of angiotensin receptor antagonist, angiotensin converting enzyme inhibitor, and their combination on regression of left ventricular hypertrophy of diabetes type 2 patients on recent onset hemodialysis therapy. AB - Left ventricular hypertrophy (LVH) is frequently found at the initiation of dialysis therapy of diabetic and hypertensive patients, and is highly predictive of future cardiac morbidity and mortality. In patients with hypertension and LVH, both an angiotensin converting enzyme (ACE) inhibitor and an angiotensin type 1 receptor (AT1) antagonist regress LVH. However, it remains controversial whether dual blockade of the renin-angiotensin system will regress LVH in these patients using a combination of ACE inhibitor and AT1 antagonist. Thirty-three type II diabetic patients with end-stage renal disease who had just entered into hemodialysis therapy and were diagnosed as having LVH evaluated by echocardiography were selected from three dialysis units staffed by the faculty of Saitama Medical School, Saitama, Japan between 1999 and 2001. The study was carried out for 1 year. All patients were assigned randomly to three groups with equal number: group I, an ACE inhibitor, enalapril 10 mg daily; group II, an AT1 antagonist, losartan 100 mg daily; group III, combination of enalapril 10 mg and losartan 100 mg daily. All antihypertensive drugs were given 30 min after the cessation of dialysis therapy. LVH was evaluated by echocardiography before the start of administration of drugs, at 6 months and 12 months after the start of drug therapy. Systolic blood pressure levels less than 140 mmHg were the target for the three groups. Using repeated measures analysis of variance, applied to those with four echocardiograms, there were progressive decreases over time in left ventricular mass index, posterior wall thickness and interventricular septum thickness. There were no significant differences in regression of LVH as well as blood pressure control between enalapril and losartan groups; however, dual blockade induced an additional 28% reduction in left ventricular mass index compared with any type of monotherapy. Both ACE inhibitors and AT1 antagonists benefit the regression of LVH in diabetic patients who start dialysis therapy. Moreover, combination therapy with ACE inhibitors and AT1 antagonists would provide more beneficial effects on LVH in these patients than monotherapy. PMID- 15274685 TI - Clinical usefulness of a new hepatitis C virus RNA extraction method using specific capture probe and magnetic particle in hemodialysis patients. AB - Hemodialysis patients are a high-risk group for hepatitis C virus (HCV) infection. Assessment of HCV infection using HCV-RNA assay among dialysis patients is important for the issue of safety and environmental protection. However, polymerase chain reaction (PCR)-based methods are unsuitable for analyzing samples from dialysis patients because the conventional centrifugal extraction method fails to eliminate heparin, a potent inhibitor of PCR. In this study, we evaluated the usefulness of a HCV-RNA extraction method using probes and magnetic particles for hemodialysis patients in comparison with the centrifugal method. The study population consisted of 17 HCV antibody-positive patients undergoing hemodialysis. These 17 patients consisted of 12 HCV carrier patients and five patients with past HCV infection. One hundred and two samples from these patients were measured using the centrifugal and magnetic methods. Moreover, we prepared five standards that included theoretically 5 KIU/mL of HCV. One was made from non-HD patient's serum and the other four were from hemodialysis patients' serum. These standards were measured using the two methods. False-negative results were not observed with the magnetic method, but were observed in five out of 102 samples with the centrifugal method. Studies using standard samples revealed that accurate HCV-RNA measurement is achieved using the magnetic method. In conclusion, the present study showed that this magnetic extraction method is a highly reproducible and reliable assay to obtain correct information about the presence of the infective virus itself in the hemodialysis setting. Precise identification of HCV-RNA using this specific method is considered to be useful in preventing HCV infection in hemodialysis units. PMID- 15274686 TI - Increased serum osteoprotegerin level in older and diabetic hemodialysis patients. AB - Osteoprotegerin is a circulating osteoclastogenesis inhibitory factor and serum osteoprotegerin levels are elevated in hemodialysis patients. This study investigated whether osteoprotegerin levels correlated with various clinical parameters in hemodialysis patients. The subjects were 45 men and 37 women aged from 27 to 94 years (mean = 60.4 +/- 13.9 years), and the duration of dialysis was 9-277 months (mean = 89.5 +/- 64.7 months). Serum osteoprotegerin levels were measured by enzyme-linked immunosorbent assay. Data were analyzed by stepwise multiple regression analysis. The mean osteoprotegerin level of the hemodialysis patients was 303 +/- 210 pg/mL, which was higher than in age-matched healthy controls. Osteoprotegerin levels increased with age, a longer duration of dialysis, and the presence of diabetes. Skeletal resistance to parathyroid hormone might be increased by aging, a long dialysis period, and diabetes, perhaps explaining why adynamic bone disease is more common in older or diabetic patients. PMID- 15274687 TI - Kidney disease quality of life of Japanese dialysis patients who desire administration of sildenafil and the treatment of erectile dysfunction using sildenafil. AB - Erectile dysfunction (ED) is common among patients on dialysis therapy. In the present study, we attempted administration of sildenafil to Japanese patients undergoing dialysis. In order to diagnose ED before prescribing sildenafil, we assessed the hemodialysis patients who desired sildenafil by using the five items version of the International Index of Erectile Function (IIEF-5). In addition, the characteristics of the quality of life in Japanese hemodialysis patients who desired sildenafil were assessed using the kidney disease quality of life (KDQOL). To all 37 male subjects (mean age of 53.8 +/- 10.4 years) attending the Outpatient Hemodialysis Unit at Atsugi Clinic (Atsugi City, Japan), it was explained by their primary doctor that the treatment of ED with sildenafil was possible. As a result, 10 patients (27.0%) desired the treatment. For eight patients, ED was diagnosed by IIEF-5 prior to prescription of sildenafil. The mean IIEF-5 scores were 6.13 +/- 4.67 points. Sildenafil was prescribed to five patients (three diabetic, two non-diabetic) and sexual function was improved in three cases. The main adverse effect was found to be ventricular arrhythmia in one case. As for KDQOL, the group desiring sildenafil showed significantly high values in Dialysis Staff Encouragement and Patient Satisfaction. Among the other nine dialysis patients (five diabetic, four non-diabetic; mean age of 58.1 +/- 8.9 years) who visited the ED department of Ishida Hospital (Asahikawa City, Japan), sildenafil was effective for all non-diabetic patients (100%) and for only one diabetic patient (20%). Among all 14 patients at Atsugi Clinic and Ishida Hospital, sildenafil efficacy rates were 83.3% for non-diabetic patients and 37.5% for diabetic patients. Non-diabetic patients without the side-effects were all responders for the sildenafil treatment. The patients who relied on the dialysis staff and were more satisfied with the general treatment in the dialysis institute desired the administration of sildenafil under the present circumstances wherein the dialysis population had few experiences of sildenafil treatment. Diabetic status is thought to be a negative factor for the response of sildenafil treatment in dialysis patients. PMID- 15274688 TI - Thyroid storm-induced multiple organ failure relieved quickly by plasma exchange therapy. AB - We report a 54-year-old female patient in whom thyroid storm was improved dramatically by plasma exchange. The patient presented with tachycardia, high fever and pulmonary congestion, in addition to left hemiparalysis and dysarthria. Serum thyroid hormone concentrations were markedly increased and computed tomography showed a fresh cerebral infarct, suggesting that she had thyroid storm precipitated by cerebral infarction. As there was no remarkable improvement even after 24 h of conventional therapy, plasma exchange was carried out using fresh frozen plasma. Consequently, her critical condition improved quickly. The half life of thyroid hormones is so long that quick improvement is not always achieved even by sufficient doses of antithyroid drugs. Thus, plasma exchange in combination with conventional therapy appears to be effective in relieving the life-threatening state in our patient with thyroid storm precipitated by acute cerebral infarction. PMID- 15274689 TI - Adipocyte lipolysis, hormonal and metabolic changes in ethanol-drinking rats. AB - The effect of ethanol drinking on some hormonal and metabolic changes in the rat and on lipolysis in isolated adipocytes was tested. Male growing Wistar rats divided into two groups were used in the experiment. Ten percent ethanol solution as the only drinking fluid for 2 weeks depressed body weight gain. The diminution of blood insulin with simultaneous increase in leptin concentration found in these rats suggest that the physiological regulation of leptin secretion is disturbed by ethanol. Liver triglycerides content was substantially augmented due to ethanol ingestion. Adipocytes were isolated from both groups of rats by collagenase digestion and the lipolytic activity of these cells was compared. Isolated cells (10(6)/ml) were incubated for 90 min in Krebs-Ringer buffer (pH 7.4, 37 degrees C) containing 3 mm glucose and different lipolytic modulators: adrenaline (1 microm), insulin (1 nm), dibutyryl-cAMP (1 mm) and DPCPX (a selective antagonist of adenosine A1 receptor, 1 microM). To determine basal lipolysis cells were incubated without lipolytic agents. Lipolysis was determined by the amount of glycerol released from cells to the incubation medium. Basal and adrenaline-induced lipolysis was depressed in adipocytes of ethanol-drinking rats. The antilipolytic activity of insulin was the same in both groups of isolated cells. Lipolysis induced by dibutyryl-cAMP was only slightly reduced due to ethanol consumption, whereas triglycerides breakdown evoked by adenosine A1 receptor antagonism was unchanged. Results obtained in vitro indicate that subchronic ethanol drinking attenuates basal and stimulated lipolysis in adipocytes, however, the antilipolytic effect of insulin and the adenosine pathway are unchanged. PMID- 15274690 TI - Consequences of eicosapentaenoic acid (n-3) and arachidonic acid (n-6) supplementation on mast cell mediators. AB - Mast cells are important players in the pathogenesis of atopic diseases. These cells release immediate-phase and late-phase mediators of inflammation. Fatty acids are incorporated in cellular membranes and therefore seem to influence mediator production and release. A study was conducted to assess the effects of eicosapentaenoic acid (EPA, 20:5n-3) and arachidonic acid (AA, 20:4n-6) on mast cell mediators in a canine mastocytoma cell line (C2). Cells were cultured in a basic medium (Dulbecco's modified Eagle's medium/HAM's F12 1 : 1, DEH), DEH supplemented with 14.0 microm EPA (DEH-EPA) or 14 microm AA (DEH-AA). The DEH-AA cultured cells had increased spontaneous and mastoparan-stimulated PGE2 production and histamine release. Furthermore, the tryptase activity was increased. The DEH-EPA cultured cells rendered elevated levels of PGE2 and histamine release compared with DEH only after stimulation. These levels were significantly lower in comparison to DEH-AA. The increased PGE2 production of C2 cultured in DEH-AA is the consequence of the AA enrichment, because AA is the precursor of PGE2. However, the different effects by AA and EPA on mast cell mediators possibly reflect the higher susceptibility of long chain polyunsaturated fatty acids (PUFA) to undergo lipid peroxidation, because it is known that altered cellular redox state influences mediator production and release. PMID- 15274691 TI - Diurnal rhythm in heat production and oxidation of carbohydrate and fat in pigs during feeding, starvation and re-feeding. AB - Diurnal rhythm in heat production (HE), oxidation of carbohydrate (OXCHO) and fat (OXF) was calculated from daily measurements of gas exchange in 12 pigs [20-40 kg live weight, (LW)] during 6 days of near ad libitum feeding, followed by 4 days of starvation and 4 days of re-feeding. All measurements, divided in five times intervals from 12.00 to 8.00, showed the highest values of HE, reflecting the animals' energy requirements, between 12.00 and 16.00 gradually declining to the lowest values between 4.00 and 8.00. The values measured in the interval 4.00 8.00 were considered as a basal metabolic rate (BMR), being in all measurements 25% lower than during 12.00-4.00. The lowest BMR was measured on the fourth day of starvation (21.7 kJ/h.kg(0.75)). By transition from feeding to starvation, OXCHO declined gradually, but was for 16 h able to cover the energy requirement with no contribution from OXF. The decline in OXCHO proceeded for 40 h and reached zero between 4.00 and 8.00 on the first day of starvation with the energy requirement being covered by OXF. The HE during starvation was 25-30% lower than during feeding caused by absence of feed-induced thermogenesis and by the transition from OXCHO to OXF. Immediately after re-feeding dietary carbohydrates were oxidized, however, there was still a substantial OXF, proceeding until the next feeding. From the second day of re-feeding the contribution of substrates to the total HE was re-established with no OXF and the same level of HE as during feeding. PMID- 15274692 TI - Lipid concentrations of fillets, liver, plasma and lipoproteins of African catfish, Clarias gariepinus (Burchell 1822), fed diets with varying protein concentrations. AB - This study investigated the effect of the dietary protein concentration on lipid concentrations in fillet and liver and concentrations of lipids in plasma and lipoproteins in African catfish. Two experiments were carried out, in which African catfish were fed diets with various protein concentrations. In experiment 1, semisynthetic diets with various concentrations of casein (350, 450 or 550 g protein/kg) were used. In experiment 2, diets were based on a commercial trout diet supplemented with various amounts of casein or carbohydrates, resulting in protein concentrations between 282 and 545 g/kg diet. In both experiments, the dietary protein concentration had a significant effect on growth, feed conversion ratio and carcass composition. Maximum of body weight gains and feed efficiency ratios were reached in both experiments at the highest dietary protein concentrations. Increasing the dietary protein concentration continuously increased masses of fillets and reduced masses of the liver and adipose tissue in the abdominal cavity. Fish fed the diets with the highest protein concentrations had the lowest concentration of total lipids, triglycerides and cholesterol in the fillets, the highest percentage of polyunsaturated fatty acids (PUFA) in total lipids of fillets and the lowest concentrations of saturated fatty acids (SFA). Fish fed the diets with the highest protein concentration also had the lowest concentrations of triglycerides in the liver, the highest percentages of PUFA in liver total lipids and the lowest percentages of SFA. Moreover, fish fed diets with high protein concentrations (501 and 545 g/kg) had significantly lower concentrations of triglycerides, cholesterol and phospholipids in plasma than fish fed diets with lower protein concentrations. In conclusion, this study shows that the dietary protein concentration does not only influence growth, feed efficiency and carcass composition in African catfish, but also influences their lipid metabolism and lipid concentrations of liver and fillet. PMID- 15274693 TI - Effect of glycine and vitamin supplementation on sulphur amino acid utilization by growing cattle. AB - Effects of glycine (Gly) and B-vitamins on sulphur amino acid (AA) utilization were studied in growing steers maintained under conditions where methionine (Met) was first limiting. Conditions were generated by limit feeding a diet low in ruminally non-degraded protein and abomasally infusing an AA mixture limiting in Met. Retained N tended (p = 0.07) to improve when steers received 10 mg folate, 10 mg vitamin B6, and 0.10 mg vitamin B12 daily. Hepatic vitamin B12 (p = 0.08) and folate (p = 0.05) concentrations increased with vitamin supplementation. In another trial, factorial treatments were 2 or 5 g/day L-Met and 0 or 50 g/day Gly infused abomasally. Retained N increased (p < 0.05) in response to Met, and responses were numerically larger in the presence of supplemental Gly. In a different trial, factorial treatments were 0 or 2.4 g/day L-cysteine (Cys) and 0 or 40 g/day Gly. Retained N was not affected by Cys in the absence of Gly, but was increased by Cys when Gly was supplemented (interaction, p = 0.01). B-vitamin status may affect sparing of Met by Cys. Supplemental Gly improved responses to supplemental Met and Cys. PMID- 15274694 TI - Relationship between the in vitro-estimated utilizable crude protein and the Cornell Net Carbohydrate and Protein System crude protein fractions in feeds for ruminants. AB - The objectives of this experiment were to study the regression relationship between the in vitro-estimated utilizable crude protein (uCP) and the Cornell Net Carbohydrate and Protein System (CNCPS) crude protein fractions, i.e. PA, PB1, PB2, PB3 and PC of feeds for ruminants, and the possibility of using CNCPS crude protein fractions for the estimation of uCP of feeds for ruminants. The CP, the non-protein nitrogen (NPN), the soluble cru de protein (SCP), the neutral detergent insoluble crude protein (NDICP) and the acid detergent insoluble crude protein (ADICP) of 30 single feeds were determined and the CP fractions, i.e. PA, PB1, PB2, PB3 and PC, were then calculated based on CNCPS. The uCP of the 30 feeds was estimated using the in vitro incubation technique of Zhao and Lebzien (2000). It was found that there was a significant multiple regression relationship between the uCP [g/kg dry matter (DM)] and the CNCPS crude protein fractions, i.e. PA (% DM), PB1 (% DM), PB2 (% DM), PB3 (% DM) and PC (% DM) of the feed samples. uCP = (9.95 +/-2.73)PA + (2.92 +/- 1.36)PB(1) + (7.24 +/- 0.86)PB(2) + (8.20 +/- 3.33)PB(3) + (17.67 +/- 3.79) PC + (63.26 +/- 18.02), r2 = 0.90, n = 30, p < 0.0001. It was suggested that the multiple regression relationship shown above could be used for the estimation of the uCP of feeds based on the CNCPS crude protein fractions PA, PB1, PB2, PB3 and PC of feeds for ruminants. PMID- 15274695 TI - The safety and effectiveness of single-pass erbium:YAG laser in the treatment of mild to moderate photodamage. AB - BACKGROUND: There are several treatment modalities for mild to moderate photodamage. The demand for effective treatments with minimal side effects has increased. OBJECTIVE: The objective of this study was to determine the effectiveness of short-pulsed erbium:YAG laser in treating mild to moderate photodamage. METHODS: Twenty patients were treated with the short-pulse erbium:YAG laser on the face and neck. One pass was given over the entire face with two to three passes over the perioral and periorbital regions. Patients were evaluated for improvement of pigmentary irregularities, skin texture, and fine wrinkles. All side effects were recorded at follow-up visits. Two lasers were used, Sciton and Cynosure CO3, under local anesthesia. RESULTS: There was a 58% reduction in pigment irregularities and a 54% improvement in skin texture. There was minimal improvement in fine wrinkles with one pass. Two and three passes resulted in a 50% reduction in wrinkles. The procedure was minimally painful. Side effects included 3 to 5 days of erythema and edema. Patients returned to work within 3 days on average. There were no infections. Patient satisfaction with the procedure was rated as very good. There was no difference in efficacy or adverse effects with either laser. CONCLUSIONS: One-pass short-pulse erbium:YAG laser in treating photodamage of the face and neck is safe and effective. There are minimal side effects and patients heal within 3 to 5 days. PMID- 15274696 TI - 5-aminolevulinic acid photodynamic therapy: where we have been and where we are going. AB - BACKGROUND: Photodynamic therapy, utilizing the topical administration of 20% 5 aminolevulinic acid, has generated a great deal of interest in the dermatology community over the past several years. OBJECTIVE: The purpose of this article is to review the history of photodynamic therapy in dermatology and to review recent new advances with this technology that will increase its appeal to all dermatologists. METHODS: A literature review and results of new clinical trials with regards to photorejuvenation and acne vulgaris treatments with 5 aminolevulinic acid photodynamic therapy are presented. RESULTS: Short-contact, full-face 5-aminolevulinic acid photodynamic therapy treatments with a variety of lasers and light sources have shown to be successful in treating all facets of photorejuvenation and the associated actinic keratoses as well as disorders of sebaceous glands, including acne vulgaris. The treatments are relatively pain free, efficacious, and safe. They are also making already available laser/light source therapies work better for acne vulgaris and photorejuvenation. CONCLUSIONS: The use of 5-aminolevulinic acid photodynamic therapy with short contact, full-face broad-application therapy is now able to bridge the world of medical and cosmetic dermatologic surgery. This therapy is available for all dermatologists to utilize in the care of their patients. PMID- 15274697 TI - Intense pulsed light treatment of photoaged facial skin. AB - BACKGROUND: It has been reported that intense pulsed light is efficacious for rejuvenation of photoaged skin, specifically the improvement of appearance of telangiectases and solar lentigines. OBJECTIVE: The objective was to define the treatment variables for photodamaged facial skin using a newer intense pulsed light system. METHODS: Twenty-three female subjects received three treatments using double-stacked pulses with fluences of 24 and 30 J/cm2. Response to treatment was evaluated using digital photography. Three signs of photoaging were evaluated: surface texture/roughness, mottled hyperpigmentation, and erythema/telangiectases. RESULTS: There was a shift in clinical grading from more to less severe on all three measures of photoaging. CONCLUSION: Intense pulsed light therapy was efficacious in ameliorating the clinical signs of photoaging. The device was well tolerated with minimal side effects. PMID- 15274698 TI - Controversies in perioperative management of blood thinners in dermatologic surgery: continue or discontinue? AB - BACKGROUND: The use of blood thinners has increased dramatically in recent years among the general, and especially among the elderly, population. When these patients need to undergo cutaneous surgery, the surgeon encounters the obvious problem of whether to stop these medications before surgery. OBJECTIVE: The objective was to evaluate the risks and benefits associated with the continuation of blood thinners perioperatively in cutaneous and Mohs micrographic surgery. METHODS: The study comprises two parts: a search of the literature in English that examined articles that related to the perioperative use of blood thinners in dermatologic surgery and a presentation of data of continuous warfarin therapy in patients who underwent Mohs surgery in our practice. RESULTS: A total of 15 articles were published in the literature until October 2003. One article showed an increase in complications in patients treated with warfarin, but not with aspirin. All other articles showed no increase in complications during the perioperative period. Data from our practice showed that of a total of 2790 patients, 68 were operated on while taking warfarin (2.4%). Intraoperative bleeding was easily controlled and postoperative bleeding was not recorded in any of the patients. CONCLUSION: Continuous treatment with blood thinners perioperatively in patients undergoing Mohs and cutaneous surgery is not associated with an increase in surgical complications. Discontinuation of these medications may increase the risk of cerebral and cardiovascular complications. PMID- 15274699 TI - Clinical classification of bioengineered skin use and its correlation with healing of diabetic and venous ulcers. AB - BACKGROUND: Chronic wounds are being treated with bioengineering skin constructs. Yet, there is no standard way of assessing these wounds. We developed a classification system to evaluate wounds after construct application. The classification system evaluates the early clinical effect of bioengineered skin and early construct appearance giving a total score named the skin substitute score. OBJECTIVE: Apply classification system to both venous and diabetic foot ulcers and determine whether classification system has validity and predictability for healing. METHODS: Evaluated serial photographs in 83 and 78 patients with diabetic foot ulcers and in 84 and 83 patients with venous ulcer on Days 7 and 14, respectively, treated with a bilayered bioengineered skin construct. Applied the classification system and determined the percentages of healed patients. RESULTS: There was a significant correlation between better skin substitute score and complete wound closure for both venous ulcers p=0.002 on Day 7 and p=0.01 on Day 14) and diabetic foot ulcers p=0.0005 on Day 7 and p<0.0001 on Day 14). CONCLUSION: Optimal clinical effect was associated with complete wound closure. As the clinical effect becomes less than optimal continued clinical persistence of the construct becomes important. This classification system seems to have validity in predicting complete wound closure in wounds treated with a bilayered bioengineered skin construct. PMID- 15274700 TI - Topical immunomodulation in dermatology: potential of toll-like receptor agonists. AB - BACKGROUND: Topical immunomodulators include both immunostimulatory and immunosuppressive agents. If successful, topical immunotherapy may represent an important improvement in the therapy of inflammatory dermatoses, viral infections, and cancers of the skin and genital mucosa. Topical immunotherapy using obligate contact sensitizers such as diphencyprone or dinitrochlorobenzene has been used against viral (e.g., common warts) and autoimmune diseases (e.g., alopecia areata). RESULTS: Newer agents such imidazoquinolines (imiquimod and resiquimod) act by cytokine secretion from monocytes/macrophages (interferon alpha, interleukin-12, tumor-necrosis factor-alpha). The locally generated immune milieu leads to a Th1-dominance and cell-mediated immunity that have been clinically used to treat viral infections such as human papillomavirus, herpes simplex virus, and mollusca. Although these agents improve antigen presentation by dendritic cells, they also act on B cells leading to the synthesis of antibodies such as IgG2a. We have also introduced this treatment against cancerous lesions including initial squamous cell and basal cell carcinoma in immunocompetent and immunosuppressed patients. We provide examples of successful treatment of squamous cell cancer using topical imiquimod. CONCLUSION: The available and additional Toll-like receptor agonists will help to improve the specific dermatologic therapy. Topical immunotherapy with both immunostimulatory and immunosuppressive agents bears potential for effective and patient friendly treatment of inflammatory, infectious, and cancerous skin diseases. Long-term evaluation will define the tolerability and safety profile of these novel topical agents. PMID- 15274701 TI - Treatment of ischemic ulcers of the lower limbs with alprostadil (prostaglandin E1). AB - BACKGROUND: Hemodynamic, hemorheologic, and metabolic changes are main determinants in the genesis of ischemic leg ulcers. Because prostaglandin E1 (alprostadil) could successfully counteract these changes, it has been intravenously used in the treatment of this disease. OBJECTIVE: The aim of this study was to evaluate the efficacy of alprostadil in the treatment of ischemic ulcers and to compare subcutaneous with intravenous administration. METHODS: Eighty patients were enrolled. Twenty-five were treated by injecting low doses of alprostadil around ischemic ulcers of the leg and saline solution intravenously and 25 were treated with intravenous alprostadil and local injections of saline solution; the control group was composed of 30 patients who received saline solution around the ulcers and intravenously. RESULTS: All patients showed a statistically significant improvement in ulcer diameter, pain, and transcutaneous oxygen pressure compared to the control group. No relevant differences in the clinical outcome in the two treated groups were found, but patients treated with subcutaneous alprostadil experienced no side effects and showed higher values of transcutaneous oxygen pressure. CONCLUSIONS: Both intravenous and local subcutaneous alprostadil are useful in the treatment of ischemic leg ulcers, but subcutaneous administration is less expensive and easier to perform. PMID- 15274702 TI - Dermoscopic features of mucosal melanosis. AB - BACKGROUND: Melanosis (lentiginosis, labial melanotic macula) is a benign pigmented lesion of mucosa characterized by pigmentation of basal keratinocytes with melanocytic normal or slightly increased in number. Melanosis, particularly when occurring on genitalia, can clinically mimic mucosal melanoma thus creating concern in both the patient and the physician. OBJECTIVE: In this study dermoscopic features from a series of clinically equivocal (n=11) or clinically typical (n=10) mucosal melanosis were analyzed. METHODS: All the women consecutively seen at the Vulva Clinic of the Department of Obstetrics and Gynecology, University of Florence, Italy, from May 1, 2002 to June 30, 2002, were examined. RESULTS: Three major dermoscopic patterns were identified: (1) a "structureless" pattern, predominantly found in clinically equivocal vulvar melanosis, with a blue hue, associated with the presence of melanophages in the upper dermis, present in the majority of these lesions; (2) a "parallel pattern," often found in clinically typical melanotyc macules of the lips and penis; and (3) a "reticular-like" pattern associated with clinically equivocal melanosis occurring at peculiar sites such as the areola (all the three cases occurred at that site) or, rarely, on the lip. CONCLUSIONS: Dermoscopy can play a role in the noninvasive classification of mucosal melanosis. The risk of misclassification with melanoma is probably dependent on dermoscopy pattern shown by the lesion. Prospective studies including early melanomas are needed to establish diagnostic performance of dermoscopy in pigmented lesions of the mucosa. PMID- 15274703 TI - Verrucous carcinoma of the skin: long-term follow-up results following surgical therapy. AB - BACKGROUND: Owing to the benign appearance and slow growth of verrucous carcinoma of the skin, its diagnosis and therapeutic management still pose problems. OBJECTIVE: This study was undertaken to point out clinical and histopathologic features of verrucous carcinoma of the skin and to provide diagnostic and therapeutic guidelines on the basis of the long-term results from 20 patients. METHODS: A retrospective study of the long-term results of 20 patients treated surgically for verrucous carcinoma of the skin is presented. In 16 cases, a wide resection with histopathologic examination of the margins was possible. Two tumors were shaved; 1 case required below-knee amputation and 1 patient refused primary amputation. In April 1999, 9 of the 10 surviving patients underwent physical examination, ultrasonography of the regional lymph nodes and the abdomen, and chest X-ray. The protocols of autopsies or postmortem examinations of the deceased patients were consulted. RESULTS: The deceased patients achieved an average tumor-free survival period of 86.1 months; eight of the surviving patients had an average tumor-free follow-up of 127.4 months. Two patients suffered recurrences. CONCLUSION: Curative treatment can be achieved by timely and complete resection of verrucous carcinoma of the skin, even in advanced cases. PMID- 15274704 TI - The retroangular flap used in the surgery of nasal tip defects. AB - BACKGROUND: The treatment of large, deep, postsurgical nasal tip defects presents a serious problem for skin surgeons and is generally dealt with by using complex flaps or skin grafts. OBJECTIVE: The objective was to present the retroangular flap, a procedure that has only recently been described in literature. METHODS: The retroangular flap technique is based on inverting the blood flow in the angular artery, a vessel that runs along the nasal groove just below the surface of the skin. RESULTS: The retroangular flap is a single-step, easily performed, procedure that is usually carried out under local anesthesia. CONCLUSION: This technique can be used for the treatment of nasal tip defects and is a useful resource for skin surgeons. PMID- 15274705 TI - Dermoscopic features of mucinous carcinoma of the skin. AB - BACKGROUND: Dermoscopic features of nonpigmented skin lesions are seldom reported; dermoscopy might be useful in speculating pathologic features in the upper dermis. OBJECTIVE: The objective was to identify additional dermoscopic criteria. METHODS: Dermoscopy of the mucinous carcinoma of the skin occurring on the cheek of a 69-year-old man was performed. RESULTS: We have shown characteristic dermoscopic features of whitish network and light-brown globules and they correspond to the pathologic findings of fibrous septum and mucinous deposition, respectively. DISCUSSION: Dermoscopic examination seemed useful as an adjunct to the diagnosis of this rare nonpigmented malignant neoplasm. PMID- 15274706 TI - Utilization of the 1320-nm Nd:YAG laser for the reduction of photoaging of the hands. AB - BACKGROUND: Nonablative laser resurfacing has been shown to improve the appearance photoaged skin. Clinical improvement has been associated with dermal collagen remodeling. OBJECTIVE: The objective was to determine the efficacy of a 1320-nm Nd:YAG laser for the treatment of photoaging hands. METHODS: Seven patients with photoaged hands received six monthly treatments with a 1320-nm Nd:YAG laser. Improvement in skin smoothness was evaluated by objective and patient assessment using a 6-point improvement scale: 1=no improvement and 6=80% to 100% improvement. RESULTS: Mild to moderate improvement was achieved as determined by both objective and patient assessment. Mean improvement by objective assessment was 2.4 points. Objective improvement was noted in four of seven patients, and these patients demonstrated a mean improvement score of 3.5 points. The mean improvement by patient assessment was 3.1 points. CONCLUSION: This case series demonstrates that the 1320-nm Nd:YAG laser with cryogen cooling can be effective for rejuvenation of photoaged hands. PMID- 15274707 TI - Incidental idiopathic calcinosis cutis in a rhytidectomy patient. AB - Calcinosis cutis, deposition of insoluble calcium salts in cutaneous tissues, is an uncommon disorder. This condition can be classified as metastatic, dystrophic, idiopathic, and iatrogenic based on the pathogenesis of the deposition. Whereas dystrophic calcinosis cutis is a fairly common condition, idiopathic cases are very rare. Distinct cutaneous anatomic areas, the vulva, scrotum, penis, and breast, have been reported to develop this disorder. Nevertheless, our case who had idiopathic calcinosis cutis in her neck may be a proof that this condition is not confined to genitals or the breasts and can be seen all over the skin. PMID- 15274708 TI - Successful carbon dioxide laser therapy for refractory anogenital lichen sclerosus. AB - BACKGROUND: Lichen sclerosus is a chronic inflammatory dermatitis that often occurs in the anogenital area and presents a therapeutic challenge. Traditional medical management includes potent topical corticosteroids and is marked by variable results. OBJECTIVE: The objective was to describe the successful use of carbon dioxide laser ablation therapy in two women with refractory anogenital lichen sclerosus. METHODS: A case is reported and the literature is reviewed. RESULTS: Two women with medically recalcitrant anogenital lichen sclerosus were successfully treated with the carbon dioxide laser. Both patients tolerated the procedure well and had excellent surgical outcomes. CONCLUSION: Lichen sclerosus recalcitrant to medical therapy presents a therapeutic challenge This may be successfully treated with the carbon dioxide laser with excellent surgical results and minimal risk. PMID- 15274709 TI - Nd:YAG laser treatment of recalcitrant folliculitis decalvans. AB - BACKGROUND: The scarring follicular disorders pose challenging therapeutic dilemmas. OBJECTIVE: Hair removal lasers have recently been shown to be efficacious in the management of these disorders. METHODS: We present a young, Fitzpatrick skin type VI African-American patient with recalcitrant folliculitis decalvans, whom we treated with the neodymium:yttrium aluminum garnet (Nd:YAG) laser. RESULTS: A remission of folliculitis decalvans was successfully achieved using the Nd:YAG for laser depilation. CONCLUSION: Based on the optical properties of light in skin, the Nd:YAG laser is the best for laser depilation in dark individuals. PMID- 15274710 TI - Multifocal basal cell carcinoma developing in a facial port wine stain treated with argon and pulsed dye laser: a possible role for previous radiotherapy. AB - BACKGROUND: Basal cell carcinoma occurring in port wine stain is extremely rare but has been reported after radiotherapy for port wine stain. OBJECTIVE: The objective was to present 55-year-old man with a facial port wine stain who had multiple treatment sessions with both the argon laser and the pulsed dye laser and subsequently developed a recurrent multifocal basal cell carcinoma and highlight the treatment carried out and its effects on the outcome. METHODS: We describe the history of the patient's port wine stain treatment, the development of skin cancer within it, and the different modalities of therapy carried out. RESULTS: The patient developed basal cell carcinoma in his port wine stain. The possibility of a causal link between laser treatment and this skin cancer was considered but it was discovered that he had had radiotherapy treatment of his birthmark at age 9. The basal cell carcinoma was successfully treated. CONCLUSION: This case of a clinically indistinct multifocal basal cell carcinoma arising within facial port wine stain was most likely due to previous radiotherapy treatment as a child, rather than laser treatment. Careful treatment history should identify such patients who should be followed-up for development of skin cancer. PMID- 15274711 TI - Treatment of Acanthosis nigricans of the axillae using a long-pulsed (5-msec) alexandrite laser. AB - BACKGROUND: Acanthosis nigricans of the axillae is a common cutaneous disorder that is difficult to treat. OBJECTIVE: The objective was to assess the efficacy and safety of a long-pulsed alexandrite laser (5 msec) in the treatment of acanthosis nigricans of the axillae. METHODS: A single axilla was treated using the long-pulsed alexandrite laser. Ten sessions were required, at fluences of 16 to 23 J/cm2 using either 10- or 12.5-mm spot sizes. The untreated axilla served as a control. RESULTS: Greater than 95% clearance was achieved after seven sessions. There was no recurrence after 2 years. The untreated axilla was unchanged. CONCLUSIONS: The long-pulsed alexandrite laser can effectively and safely treat acanthosis nigricans of the axillae. PMID- 15274712 TI - Successful treatment of a chronic atrophic dog-bite scar with the 1450-nm diode laser. AB - BACKGROUND: Atrophic scars can be revised with surgical methods and more recently with newer ablative and nonablative laser techniques. Nonablative laser technology offers the advantage of improving the appearance of atrophic scars without the risks associated with traditional surgery or ablative lasers. METHODS: A case of a large linear atrophic dog-bite scar on the chin of greater than 2-year duration treated for three sessions at 4- to 6-week intervals with the 1450-nm diode laser is presented. RESULTS: Fifty to seventy-five percent improvement in the appearance of the scar resulted after three treatments with the 1450-nm diode laser. No adverse effects were noted from the treatment. The patient subjective rating of scar improvement was more than 50%. CONCLUSION: The 1450-nm diode laser may provide a noninvasive and effective alternative for the treatment of atrophic traumatic scars. This method appears to be potentially effective even for treatment of very mature scars and warrants additional studies. PMID- 15274713 TI - Sebaceous carcinoma of the eyelid: a review of 14 cases. AB - BACKGROUND: Sebaceous carcinoma of the eyelid is a rare tumor. Treatment can be complicated by noncontiguous spread of the tumor. OBJECTIVE: Review a series of patients with sebaceous carcinoma to illustrate clinical presentations, treatments, and outcomes. METHODS: We retrospectively reviewed medical records of patients with sebaceous carcinoma treated at Mayo Clinic (Rochester, MN). RESULTS: Fourteen patients had sufficient follow-up data available for review. Mean follow-up was 57 months (range 18-134 months). Treatment included wide local excision with frozen and permanent section control (9 patients, 64%), Mohs micrographic surgery (2 patients, 14%), external beam radiation (2 patients, 14%), and exenteration and total parotidectomy with cervical lymph node dissection (1 patient, 7%). Two patients (14%) had local recurrence of the tumor after wide local excision, and 1 patient (7%) had tumor recurrence after Mohs micrographic surgery. CONCLUSION: Treatment should be chosen on the basis of the extent of the tumor and the specific needs of the patient. The mainstay of treatment of tumors without orbital involvement has been wide local excision, with the margins checked in both permanent and frozen sections, in combination with conjunctival map biopsies when warranted. Mohs micrographic surgery is an alternative that may provide tissue conservation and lower recurrence rates. Recurrence rates between treatments are difficult to assess because of the small number of cases reported in the literature. In cases with orbital involvement, exenteration may be warranted. Radiation may be useful when surgery cannot be tolerated. PMID- 15274714 TI - Mohs micrographic surgery for angiolymphoid hyperplasia with eosinophilia. AB - BACKGROUND: Angiolymphoid hyperplasia with eosinophilia is a benign vascular proliferation that typically presents on the head and neck. Multiple treatment modalities have been proposed for angiolymphoid hyperplasia with eosinophilia, each with limited success or undesirable side effects. At this time, standard surgical excision is considered the treatment of choice but carries recurrence rates of 33% to 50%. OBJECTIVE: The objective was to present a case of angiolymphoid hyperplasia with eosinophilia successfully extirpated using Mohs micrographic surgery. METHODS: A 52-year-old woman presented with an ill-defined solitary plaque of angiolymphoid hyperplasia with eosinophilia within her left conchal bowl that enlarged despite conservative therapy with intralesional and topical corticosteroids. Mohs micrographic surgery using the fresh tissue technique and standard hematoxylin and eosin staining was performed. The characteristic histologic features of angiolymphoid hyperplasia with eosinophilia were readily identifiable on frozen sections and complete extirpation required two stages of micrographically controlled resection. RESULTS: Resection of angiolymphoid hyperplasia with eosinophilia of the conchal bowl with complete resolution of symptoms and no evidence of clinical recurrence 8 months after surgery. CONCLUSION: Given the high recurrence rates reported for standard excision, Mohs micrographic surgery with complete margin examination should be considered as a treatment option for angiolymphoid hyperplasia with eosinophilia, particularly for lesions with ill-defined margins or in locations where tissue sparing is desirable. PMID- 15274715 TI - Pilomatrix carcinoma of the back treated by mohs micrographic surgery. AB - BACKGROUND: Pilomatrix carcinoma (synonyms, matrical carcinoma or malignant pilomatrixoma) is a rare malignant neoplasm derived from the hair matrix first described in 1980. This neoplasm can exhibit local aggressive behavior and distant metastasis. Most pilomatrix carcinomas occur on the head and neck of elderly individuals with a predilection for males (M:F 5:1). Pilomatrix carcinoma is often clinically misdiagnosed as a sebaceous cyst and histologic difficulty can occur in differentiating this entity from the benign entity pilomatrixoma. OBJECTIVE: The objective was to describe a case of pilomatrix carcinoma encountered in a Mohs micrographic surgery practice. We present the first case of this lesion treated by Mohs surgery. METHODS: A case report and literature review are presented. CONCLUSION: Pilomatrix carcinoma is a rare malignant variant of pilomatrixoma. Given the rarity of this lesion there are no well-defined standards for surgical management. Wide local excision has been recommended given the high rate of reoccurrence. Mohs micrographic surgery may provide optimal treatment of this neoplasm given the ability to have 100% margin control. PMID- 15274716 TI - Inherited accessory nail of the fifth toe cured by surgical matricectomy. AB - BACKGROUND: The inherited accessory nail of the fifth toe is a common condition in the Chinese population. OBJECTIVE: The objective was to demonstrate three lesions in two cases of inherited accessory nail of the fifth toe successfully treated with surgical matricectomy. METHODS: Under local anesthesia and use of tourniquet, the proximal nail fold was incised and the matrix of the accessory nail was exposed and then excised by scalpel surgery. The skin defect left after removal of the lesion was repaired with a rotation flap. RESULTS: Histopathologic examination of the surgical specimens revealed that the matrices of the accessory nails were completely extirpated. No recurrence was found 2 years after operation. CONCLUSION: The inherited accessory nail of the fifth toe was cured by surgical matricectomy. PMID- 15274717 TI - The running bolster suture for full thickness skin grafts. PMID- 15274718 TI - Re: giant leg ulcer in Wegener's granulomatosis treated with plasmapheresis and skin graft. PMID- 15274719 TI - Nipple piercing: it is wiser to avoid in patients with hyperprolactinemia. PMID- 15274720 TI - A simple and useful suggestion for immediate postliposuction patient care: baby powder. PMID- 15274721 TI - Reconstruction of a large surgical defect of the nose. PMID- 15274722 TI - Epoxy-based production of wind turbine rotor blades: occupational dermatoses. AB - Occupational dermatoses were investigated in a factory producing rotor blades for wind turbines by an epoxy-based process. In a blinded study design, 603 workers were first interviewed and thereafter clinically examined. Based on a history of work-related skin disease, clinical findings of dermatitis, or both, 325 (53.9%) of the workers were patch tested with a specially profiled occupational patch test series and the European standard patch-test series. Calculated on all investigated workers, 17.1% of the workers were diagnosed with occupational dermatoses caused by work. Occupational allergic contact dermatitis was found in 10.9% of the workers. The estimated frequency of irritant contact dermatitis caused by work was 6.1%. Dermatitis on the hands was associated with contact allergy to epoxy resin (P = 0.017). The number of days on leave before the clinical examination was negatively associated with the presence of dermatitis (P = 0.001). Among workers employed 7-12 months, the frequency of occupational contact allergy was higher than that among workers employed for 5 g kg(-1) in mice). The extract was investigated for antihypoxic activity in two models of experimental hypoxia--haemic and circulatory. Antihypoxic activity was especially pronounced in the model of circulatory hypoxia. This effect may be attributed, at least in part, to the presence of flavonoids in the extract. PMID- 15274759 TI - The antidepressant activity of Hypericum perforatum L. measured by two experimental methods on mice. AB - The pharmacological approach to the treatment of depression includes a long-term employment of antidepressants, either in the form of monotherapy or as a combination of several antidepressants with various mechanisms of action. Hypericum perforatum L. (St. John's wort) is the only natural antidepressant. Several constituents of the extract, such as hypericin and hyperforin, seem to be important for this effect. H. perforatum is considered to be an effective alternative to other therapeutic agents in the treatment of mild to moderate depression. The paper describes the investigation of the antidepressant effect of H. perforatum (doses 7, 35 and 70 mg kg(-1) b. m.) on mice using the forced swimming and tail-suspension methods. As an indicator of the antidepressant effect, it was shown that the immobility time of animals in the forced-swimming and tail-suspension experiments was shorter, i.e. the activity of the animals was higher. With single doses of extract suspension increasing from 7 over 35 to 70 mg kg(-1) the antidepressant effect increased in proportion by 10.1%, 25.8% and 38.6% in the swimming method, and by 12.7%, 16.5% and 24.5% in the tail suspension method compared to controls. H. perforatum extract displays dose dependent antidepressant effect at a dose as low as 7 mg kg(-1). Both models have proved to be equally valuable for demonstration of substances with a potential antidepressant effect. PMID- 15274760 TI - [Pharmacoeconomics and cardiovascular disease]. PMID- 15274761 TI - [Poor scientific behavior in communication of biomedical results]. AB - INTRODUCTION: The generically known as poor scientific behavior exists in different degrees with regard to biomedical communications and publications. From authentic fraud in the data up to the called "tricks" for curriculum fattening. MATERIAL AND METHODS: The objective of this work is to review the works presented in the XVIII Congress of the Andalusian Society of Internal Medicine (Marbella, October 2001), comparing them with the abstracts books of six more scientific meetings and congresses both regional and national. RESULTS: Of the 183 works evaluated in this review, 22 (12.02%) were doubled and 13 (7.10%) fragmented, in other words, a total of 35 (19.33%). The groups that presented communications were 36, and 17 of them (47.22%) carried out one or both fraudulent tactics. CONCLUSIONS: Despite the little bibliographic reference, in our environment exists the duplication and the fragmentation of works presented, in general as a system for curriculum increase. We advocate a higher clarity and commitment in the relationship between editors and authors, and an upsurge of the aspects of punishable deontological regulation on the one hand and of the ethics or a priori commitment on the other one. PMID- 15274762 TI - [Analysis of the effectiveness and costs of amlodipine on the hospitalizations for cardiovascular events in patients with ischemic cardiopathy]. AB - OBJECTIVE: Evaluation of the effect of amlodipine on the hospitalizations for cardiovascular events (CVE) and their associated costs in patients with ischemic cardiopathy. METHODS: Data from the multicenter, randomized, double-blind, placebo-controlled PREVENT (Prospective Randomized Evaluation of the Vascular Effects of Norvasc Trial) clinical trial were utilized. A tree-type model of decision was used in order to analyze the incremental costs expected from the treatment with amlodipine with regard to placebo. Hospitalization costs were estimated with regard to the DRG weights of the American Medicare adapted for the costs of average stay available in our environment. RESULTS: Amlodipine reduced significantly the incidence of CVE that required hospitalization in contrast to placebo; 0.60 +/- 1.16 versus 0.77 +/- 1.31 (average +/- SD), p < 0.05. The expected direct expenses due to hospitalizations were higher in the placebo group than in the amlodipine group (saving of 205.76 Euro/patient). Total cost for patient in the amlodipine group was 1,723.52 Euro while in the placebo group was 1,929.28 Euro. When the relation cost/price shifted in the sensitivity analysis from 1.20 to 0.66 (cost of every hospitalization ranged between + 20% and -34%), the saving fluctuated from 330.56 Euro to 0. Accordingly, the breakeven point of the cost/price relation it is 0.66, and above this the treatment with amlodipine still generates savings in regard to its cost. CONCLUSIONS: Amlodipine is cost effective in the treatment of the patients with ischemic cardiopathy, being able to reduce the hospital costs related to ischemic episodes in this type of patients. PMID- 15274763 TI - [Relationship between cholesterol and fibrinogen in two populations of different geographical location of Catalonia]. AB - OBJECTIVES: Verification of the following in two different geographical location populations (seashore and mountain) with cardiovascular symptomatology: 1) the prevalence of hyperfibrinogenemia and possible correlation with cholesterolemia; 2) the differences between both populations in the profiles of these parameters. DESIGN: Cross-sectional descriptive study. CONTEXT: Primary Care. PARTICIPANTS: Three hundred and seventy five patients who went to two hospitals between May 1995 and July 1998. In the seashore center 256 patients and in the mountain center 119 patients. MAIN MEASUREMENTS: Clinical history and analytical parameters. Fibrinogen (FBG), indirect prothrombin time, enzymatic cholesterol (CLT) Utachi 717. The patients were sorted out into 4 groups: 1) high FBG > 300 mg/dl and high CLT > 240 mg/dl; 2) high FBG > 300 and low CLT < 240; 3) low FBG < 300 and high CLT > 240, and 4) low FBG < 300 and low CLT < 240. RESULTS: Levels of FBG: homogeneous between groups 1 and 2 (high) and 3 and 4 (low), and different between upper and lower groups. Cholesterol showed the same behavior. Group 1 with a similar number of patients in mountain and seashore (40% and 41%). Group 2 with 42.8% of patients from mountain and 26.9% from seashore. Groups 3 and 4 are presented with lower percentages. We did not find correlation between the levels of FBG and those of CLT. CONCLUSIONS: Predominance of patients with high FBG and normal CLT (group 2) in the mountain cohort, in contrast with a higher prevalence of normal FBG and high CLT (group 3) in the seashore cohort. In participants with normal levels or with high risk the variations of the FBG were not dependent nor related to those of CLT. PMID- 15274764 TI - [Malnutrition evaluation in obese patients of both sexes on a very low caloric content diet]. AB - We have evaluated the effectiveness of a very low caloric content diet (VLCD) during 6 weeks in patients with severe obesity (grades II and III). Twenty-seven men and 61 women were selected for evaluation of anthropometric (weight, body mass index [BMI], waist, hip, C/c, fatty weight and intra-abdominal fatty area) and biochemical (creatinine-height index [CHI], albumin, transferrin, retinol binding protein [RBP], prealbumin, C3, and lymphocytes count) malnutrition parameters, at the beginning and after 6 weeks of treatment with VLCD. In men we found a significant decrease of weight, BMI, waist, hip, fatty weight, and intra abdominal fatty area. In women the decrease of weight, BMI, hip, and fatty weight was also significant. We found baseline malnutrition in 7.4% of men and in 14.7% of women, and after the treatment in 22.2% of men and in 34.4% of women (p < 0.05). With regard to the biochemical parameters of protein malnutrition, only men showed significant decrease in the CHI and only women showed significant decrease in transferrin, RBP, prealbumin, and C3. In conclusion, we can state that different types of VLCD are effective for weight loss in severe obese subjects. However, within a period of follow-up of 6 weeks we have detected the presence of protein malnutrition, especially in women, being in these patients affected the visceral compartment while in men the muscular compartment is affected. PMID- 15274765 TI - [Variability of HIV-1 viral load in patients cared along 4 years]. AB - BASIS: Analysis of the variations of HIV-1 viral load (VL) in a cohort of patients. MATERIAL AND METHODS: A retrospective study was designed for the calculation and analysis of the differences between two consecutive measurements of VL in a cohort of 1,336 patients along a 48 months follow-up. RESULTS: At the beginning of the follow-up period the highest proportion of patients with decreases of VL (54.2% in their first measurement, at 0-75 days) as well as the least proportion of patients both without changes (30.7%) and with increases of their VL (15.1%), were registered. The proportion of patients with decreases was declining along the study period. More than half of the patients did not experience significant variations in the measurements carried out. CONCLUSIONS: The significant decreases of VL appeared in our series at the beginning of the follow-up period, and a growing proportion of individuals showed elevations of the VL along the period studied. PMID- 15274766 TI - [Herpes simplex 1 virus Mollaret meningitis]. AB - Mollaret meningitis is a rare disease characterized by recurrent and self-limited episodes of aseptic meningitis. It is considered a disease with benign etiology and it has been related to viral infections: Epstein-Barr virus and herpes simplex virus (HSV), type 2 most frequently. Herpes simplex 1 virus has been rarely isolated in the cerebrospinal fluid in patients with Mollaret meningitis; to this end culture, expansion with PCR, and inmunoblot has been utilized. In this article a case of Mollaret meningitis related to type 1 HSV is described. The interest is the demonstration of the herpes simplex virus type 1 within the cytoplasm of the Mollaret cells with a immunohistochemical technique (ABC peroxidase method) using monoclonal antibodies anti HSV-1 in order to support the diagnosis, being this the first case described in the literature. PMID- 15274767 TI - [Clinical-therapeutic approach in oral leukoplasia]. PMID- 15274768 TI - [Buruli's ulcer: an emerging disease]. PMID- 15274769 TI - [Tobacco dishabituation: therapeutic attitude in patients in preparation phase]. PMID- 15274770 TI - [A patient with HIV infection and tuberculosis with cervical adenopathy worsening after initiating the treatment]. PMID- 15274771 TI - [Diarrhea with hypokalemia]. PMID- 15274772 TI - [62-year-old male with fever and lumbar pain]. PMID- 15274773 TI - [Bacillary angiomatosis or Kaposi's sarcoma]. PMID- 15274774 TI - [Genital aphthous ulcers in a case of foreign dengue]. PMID- 15274775 TI - [Ceftriaxone-induced thrombocytopenia]. PMID- 15274776 TI - [Tuberculin test and treatment of latent tuberculous infection]. PMID- 15274777 TI - [The interconsultation of surgical services: a mission for the internist]. PMID- 15274778 TI - [The internist's role in the medical evaluation of surgical patients]. AB - OBJECTIVE: To describe the characteristics of the consultations carried out by surgical services to an Internal Medicine service and to determine what factors influence the prognosis of these patients. METHODS: A prospective study of the consultations carried out by the surgical services of a 540-bed hospital to an Internal Medicine service. Analyzed variables were: age, sex, service of reference, reason for consultation, medical and admission diagnoses done during the admission, and clinical evolution. RESULTS: In the study 453 interconsultations were included, corresponding to 0.96 new interconsultations by working day and to 4.05 interconsultations per every 100 admissions in surgical services during the period study. The reasons for the more common consultations were dyspnea, fever, electrolytic and metabolic disorders, assessment of multiple conditions and acute confusional syndrome. Two or more diagnoses were carried out in 257 patients (56.7%). The average number of visits carried out by patient was 3.9 +/- 3.9. The average hospital stay in the study group was 28 +/- 33.05 days, while the average hospital stay of patients admitted in the surgical services during the same period was 11.6 days. Fifty patients (11%) had died at the time of the "medical discharge", and this percentage amounted to 20.5% (93 cases) upon considering the end of the hospital admission, compared with the global mortality of 3.7% registered during that period in the surgical services. The number of medical diagnoses and the age were independent predictors of mortality in the multivariate analysis. CONCLUSIONS: The interconsultations of the surgical services to an Internal Medicine service imply an important workload. The patients are complex from the medical standpoint. The average hospital stay and mortality of these patients are different from that of the patients cared in surgical services. PMID- 15274779 TI - [Evolution of serological characteristics in 26 patients with tularemia three years after the outbreak]. AB - INTRODUCTION: Tularemia was an unknown zoonosis in our region until the first quarter of 1998. The outbreak that took place on those dates has made possible to study some scantly known characteristics of the disease, as are the determinants for the persistence of a high antibody titer. MATERIAL AND METHODS: Between March and May, 2001, a clinical-analytical control was carried out in 26 patients who showed tularemia in the first quarter of 1998. RESULTS: The clinical forms during the acute phase were: ulceroglandular (46.2% of patients), nodal (26.9%), typhoid (15.4%) and pharyngeal (11.5%). Antibiotics most prescribed were streptomycin and doxycycline. Nine patients showed sequelae three years later. The arithmetical mean of antibody titer declined from 1/1011 in 1998 to 1/187 in 2001. 76.92% of the patients maintained an antibody titer equal or higher than 1/160 three years after the acute phase. A significant relation between the initial antibody titer and the antibody titer registered in the late phase was observed. A relationship between final antibody titer and prescribed antibiotic, presence of sequelae, gender, or age was not demonstrated. CONCLUSIONS: The antibody titer in the presence of Francisella tularensis is high in an important percentage of the patients along years. We should reconsider the standard diagnostic criteria to our community, accepting as a certainty diagnosis exclusively the presence of a demonstrative culture or the existence of seroconversion. PMID- 15274780 TI - [Influence of the diet on the development of colorectal cancer in a population of Madrid]. AB - OBJECTIVE: Although the genetic component in colorectal carcinoma (CRC) is well established, some environmental factors, mainly dietary, can favor its development. The objective of this study is to evaluate the relationship between daily consumption of specific food groups and development of CRC. METHODS: We carried out a case-control study in an area of Madrid; 196 patients with diagnosis of CRC with confirmed histology and registered to May 1998 in the Community of Madrid tumor register were included, and they were compared with 196 controls matched by age, sex, and geographical area. All of them filled out a questionnaire with information on diet, substance abuse, physical activity, drugs, and family history of CRC. RESULTS: The logistic regression analysis showed a weakly positive association with meat (OR: 1.02; CI: 1.01-1.04), viscera (OR: 1.12; CI: 1.02-1.23), and sausage (OR: 1.07; CI: 1.03-1.1) consumption, and a modest inverse association with consumption of yogurt (OR: 0.97; CI: 0.95 0.98), tomato (OR: 0.99; CI: 0.98-0.99), strawberries and cherries (OR: 0.97; CI: 0.95-0.99), oranges, grapefruits, and natural fruit juices (OR: 0.99; CI: 0.98 1). CONCLUSIONS: These results are an additional empirical evidence that must be confirmed through prospective studies. PMID- 15274781 TI - [A prospective study on patients with heart failure admitted to the internal medicine department]. AB - CONTEXT: Heart failure (HF) is one of the most important causes of morbidity and mortality and represents one of the most frequent causes of rehospitalization. PATIENTS AND METHOD: Prospective study on patients admitted because of HF. A data collection questionnaire was completed: cardiology history, HF etiology, reason for admission, previous treatment, treatment during and after the admission, hospital stay and complementary explorations carried out. A biweekly telephone monitoring was carried out after the discharge. RESULTS: 62 patients admitted because of HF with an average age of 73 +/- 11 years. The etiology of the HF was: unknown (54%), hypertensive (21.5%), valvular (15.4%), ischemic (7.3%), alcohol (1.6%). All patients received diuretic treatment, 25% angiotensin-converting enzyme inhibitors (ACEI) and 6% oral anticoagulants. 51.6% showed atrial fibrillation. Echocardiogram was carried out in 10% of the patients. The average hospital stay was 6.2 +/- 3.2 days, and it was significantly higher when an echocardiogram was carried out (5.7 +/- 0.4 versus 7.55 +/- 0.9 days; p < 0.001). 6-month incidence of rehospitalization and mortality was 26% and 20% respectively. CONCLUSIONS: HF implied one fourth of admissions in a population of elderly patients with multiple conditions. The etiology of the HF was not established in more than half of the patients. A suboptimal utilization of the available treatments is demonstrated. HF is associated to a high incidence of rehospitalization and mortality. PMID- 15274782 TI - [Mechanisms of disease in Mycoplasma pneumoniae infection. Clinical manifestations and complications]. AB - CONTEXT: Mycoplasma pneumonia has been related to several conditions. With this study we have tried to establish the condition more frequently associated and their physiopathological mechanism. METHOD: One-hundred and five patients admitted to Puerta de Hierro Hospital between March 1996 and July 2001, with IgM positive serology to Mycoplasma were evaluated. Thirty four cases were selected upon the basis of two criteria: patients which positivity was confirmed by seroconversion, elevation of the antibody titer, or confirmation by complement fixation were included, and patients with some intercurrent condition or with other diagnoses were ruled out. RESULTS: 26 patients (77%) showed a respiratory infection, and 20 of them showed a pneumonia. Two types of complications were observed. The first type were the complications due to an invasion of the tissue by Mycoplasma, with 5 cases of pleuropericarditis and three cases of pleuritis; in 5 of these there was simultaneously a respiratory process. The second type were the complications mediated by an autoimmune mechanism, with two cases of reactive arthritis, one case of vasculitis with cutaneous predominance, one case of urticaria a frigore, three cases of lymphocytic meningitis, one case of disseminated encephalitis, one patient with Guillain-Barre syndrome and one case of Adie's tonic pupil. Six of these patients showed a respiratory infection on the previous days with an average delay of 10 days between the beginning of the respiratory symptomatology and the appearance of one of these diseases. CONCLUSIONS: Mycoplasma pneumoniae can give rise to disease through a mechanism of direct invasion and through autoimmunity mechanisms. The second group of complications is seen by more frequency in young women. PMID- 15274783 TI - [Bile ducts cysts in the adult: diagnostic and therapeutic strategy]. PMID- 15274784 TI - [Non surgical pneumoperitoneum]. PMID- 15274785 TI - [Slow resolution pneumonias]. PMID- 15274786 TI - [Patient with double lung lesion and history of renal adenocarcinoma]. PMID- 15274787 TI - [Disseminated histoplasmosis in two brothers with AIDS]. PMID- 15274788 TI - [Evanescent liver nodules]. PMID- 15274789 TI - [Methimazole hepatitis]. PMID- 15274790 TI - [Corynebacterium pseudotuberculosis lymphadenitis in a young patient]. PMID- 15274791 TI - [Therapeutic criteria in the acute coronary syndrome]. PMID- 15274793 TI - [Prospective evaluation of the hyperbaric-tolerance test for the diagnosis of gestational hypertension and preeclampsia]. AB - BACKGROUND AND OBJECTIVE: Recent studies have tried to overcome poor results from isolated blood pressure values in detecting hypertension in pregnancy by relying on ambulatory monitoring. Low sensitivity of the 24-hour mean has led many authors to conclude that ambulatory monitoring is not a valid approach in pregnancy. Against this background, we have evaluated prospectively the sensitivity and specificity in the diagnosis of gestational hypertension of the tolerance-hyperbaric test. This is a combined approach consisting of establishing tolerance intervals for the circadian variability of blood pressure as a function of gestational age, and then computing the hyperbaric index (area of blood pressure excess above the upper limit of the interval) by comparison of any patient's blood pressure profile with those limits. PATIENTS AND METHOD: We studied 328 women who provided a 2014 blood pressure series. They were sampled for 48 hours once every 4 weeks from the first obstetric visit (mostly within the first trimester of gestation) until delivery. The hyperbaric index of each blood pressure series was calculated by taking into account the reference circadian tolerance limits established from a 497 series previously sampled from an independent reference group of 189 normotensive pregnant women. RESULTS: Sensitivity of the tolerance-hyperbaric test was 91% for women sampled during the first trimester of gestation, and increased up to 99% in the third trimester. Specificity was above 99% in all trimesters. Positive and negative predictive values were above 96% in all trimesters. Moreover, the hyperbaric index provided an early identification of subsequent gestational hypertension and preeclampsia, on the average of 23 weeks prior to the clinical confirmation of the disease. CONCLUSIONS: The tolerance-hyperbaric test represents a reproducible, stable, noninvasive, and high sensitivity test for the very early identification of subsequent gestational hypertension and preeclampsia, which can also be used as a guide for establishing prophylactic or therapeutic interventions. PMID- 15274794 TI - [Myocardial Infarction Registry in Asturias: the RIMAS project]. AB - BACKGROUND AND OBJECTIVE: Spanish hospital registers of myocardial infarction (MI) are not uniform. The RIMAS project is trying to know the real situation of myocardial infarction in Asturias and to observe possible differences among hospitals and with other registers. PATIENTS AND METHOD: It was a cohorts study using a hospital registry of patients with MI. All cases arriving alive to all public and private-public Asturian hospitals during 1998 were included. Demographic data, cardiovascular risk factors, delays, evolution, treatments and techniques used, were all registered. RESULTS: 875 cases were registered with a coverage rate of 77%. The average age was 66.5 years (45.6% older than 70 years) and women represented 29.1%. Sixty three per cent of the patients had tobacco consumption, 43% had arterial hypertension, and 22.3% were diabetics. The extrahospital delay was 135 min and thrombolysis delay was 180 min. Thrombolytic therapy was administered to 34.1% of patients and 4% were treated with primary angioplasty. Intrahospital mortality was 14.4%. At discharge, antiagregant therapy was administrated to 94%, betablockers to 43.2%, ACE inhibitors to 33.3% and hypolipemiants to 25% of treated patients. CONCLUSIONS: People attended in Asturias with a MI are older and there is a higher percentage of women. There are delays which include the start of thrombolytic therapy. However, there are significant differences with regard to the adhesion to clinical practice guidelines between different hospitals. PMID- 15274795 TI - [Capillary blood sample for type 2 diabetes screening: fingertip versus forearm]. AB - BACKGROUND AND OBJECTIVE: In Spain, around 50% of diabetic mellitus cases remain undiagnosed. Capillary blood puncture improves the diabetes screening accessibility. New test strips used in the Blood Glucose Monitoring System allow fingertip and forearm punctures yet they create the dilemma on what is the best anatomic place. PATIENTS AND METHOD: We compared fingertip and forearm capillary blood glucose determinations. RESULTS: In a sample of 107 patients, a 110 mg/dl cut-off point for the fingertip and 125 mg/dl for the forearm were more sensitive. Patient's preferences were as follows: 67% indifferent, 28% forearm and 5% fingertip. CONCLUSIONS: In this study, the best anatomic site for puncture depended on the cut-off point used in diabetes screening. PMID- 15274796 TI - [Screening for type 2 diabetes mellitus: more than looking for undiagnosed diabetes]. PMID- 15274797 TI - [Depression: a priority in public health]. PMID- 15274798 TI - [Criteria and guidelines for combined therapy of type 2 diabetes. 2004 consensus document]. PMID- 15274799 TI - [Informatization of emergency services]. PMID- 15274800 TI - [Another model of internal medicine consulting in primary health care]. PMID- 15274801 TI - [Use of albumin dialysis with the MARS device to treat a case acute liver failure after partial hepatic resection]. PMID- 15274802 TI - [Low serum thyrotropin associated with intravenous dopamine infusion]. PMID- 15274803 TI - [Risk percentiles: a new adapted method for evaluating vascular risk. ERVPA study]. AB - BACKGROUND AND OBJECTIVE: Various cardiovascular scoring systems estimating the risk in the Spanish population are currently used. The aim of our study was to create graphs and charts of populational percentiles to assess the total burden of risk of a population and to place each subject individually in the context of their own population setting by using cardiovascular risk charts. SUBJECTS AND METHOD: A cross-sectional study of 514 individuals of both sexes from a general population setting was carried out. Cardiovascular risk was calculated according to the SCORE project equations and charts and graphs of cardiovascular risk were formulated. RESULTS: In primary prevention, 17.11% of men and 22.36% of women aged 20-79 showed a high cardiovascular risk. The percentages vary from 0% in persons of both sexes aged 20-39 to 91.2% of men and 97.9% of women in their 70s. Risk medians range in men from 0.11% (age 20-39) to 8.86% (age 70-79) and in women from 0.02% (age 20-39) to 15.20% (age 70-79). Graphs of cardiovascular risk percentiles were formulated for both sexes; graphs can be used to situate each subject individually in the context of their own population setting and to project the risk to any age. CONCLUSIONS: Nearly a fifth of the population has a high cardiovascular risk. More than 90% of men and women in their 70s have a high cardiovascular risk. PMID- 15274804 TI - [Nicotine replacement therapy versus gradual smoking withdrawal in smoking cessation]. AB - BACKGROUND AND OBJECTIVE: The aim was to study the effectiveness of the gradual nicotine withdrawal without nicotine replacement therapy (NRT) as a method of smoking cessation during a three years follow-up period in the cognitive behavioural context in the clinical work of the Clinic Hospital Unit. PATIENTS AND METHOD: Two groups of patients with similar sociodemographic and clinical characteristics and the same degree of nicotine dependence according to the Fagestromtest were compared. We included 111 patients treated for smoking cessation in the Unit Hospital: 84 patients, under the cognitive-behavioural model, were treated with NRT and 27 patients, under the same model, were treated only with gradual nicotine withdrawal without NRT. RESULTS: There was not a significant difference between the effectiveness of the NRT (65.3%) and that of the gradual nicotine withdrawal without the NRT (51.8%) on smoking cessation(X2 =1.612; p = 0.204) at the first year or during a 3 years follow-up period (NRT: 41% and gradual nicotine withdrawal: 36%; X2: 0.124; p = 0. 725). CONCLUSIONS: The gradual nicotine withdrawal without NRT scheme shows the same degree of effectiveness at the first year and during a three years follow-up period as that of the nicotine replacement therapy scheme on smoking cessation. PMID- 15274805 TI - [Iron deficiency and iron overload in an adult population in Catalonia, Spain]. AB - BACKGROUND AND OBJECTIVE: To ascertain the frequency of iron deficiency and iron overload in an adult Catalan population. SUBJECTS AND METHOD: Multiple iron measurements, including transferrin saturation (TS) and serum ferritin (SF), were performed in a representative sample of 1,296 adults. RESULTS: The prevalence of iron deficiency was 5.6% (95% CI, 4.4 to 6.9)(SF below 12 microg/l), and 9.3% (95% CI, 7.7 to 10.9) had an iron overload(SF above 300 microg/l in men and SF above 200 microg/l in women). Iron deficiency was especially frequent in women 50 years old or younger (14.8%; 95% CI, 11.4 to 18.1),while in men of the same age it was 1.1% (95% CI, 0.1 to 2.1), yet 11.7% (95% CI, 8.7 to 14.7) had iron overload. In the population over 50 years there was an iron deficiency in 0.9% (95% CI, 0.0 to 1.8), and an iron overload in 15.1% (95% CI, 11.7 to 18.4). 1.6% (95% CI, 0.9-2.3) of all population and 3.9% (1.4-6.4) of men older than 50 years had an SF above 500 microg/l. CONCLUSIONS: Iron overload is more prevalent than iron deficiency in Catalonia, particularly in men and people over 50 years. The causes and effects of the disorder should be investigated in order to carry out corrective measures in the future. PMID- 15274806 TI - [Looking for the cardiovascular risk]. PMID- 15274807 TI - [Smoking: an avoidable risk]. PMID- 15274808 TI - [Uratic gout]. PMID- 15274809 TI - [Relevance of chronic kidney disease and cardiovascular prognosis in arterial hypertension]. AB - Recently published guidelines recognize the relevance of the finding of chronic kidney disease in the stratification of risk of the hypertensive patient. Determination of the presence of microalbuminuria and estimation of glomerular filtration rate are mandatory in order to ensure an adequate evaluation of global cardiovascular risks in the hypertensive patient. The presence of subtle elevations of serum creatinine concentrations and/or proteinuria are also potent predictors of a poor cardiovascular prognosis. Clustering of associated risk factors seems to justify the elevated cardiovascular risk observed in patients with essential hypertension and mild alterations of renal function. PMID- 15274810 TI - [Advances in the therapy of cardiac failure]. PMID- 15274811 TI - [Characteristics of women with breast cancer included in RIMCAN registry 1991 2002]. PMID- 15274812 TI - [Torsade de pointes secondary to amiodarone overdose]. PMID- 15274813 TI - [Dissatisfaction of residents]. PMID- 15274815 TI - [Bosentan, hope or reality for the treatment of pulmonary arterial hypertension]. PMID- 15274817 TI - [Syndrome of inappropriate antidiuretic hormone secretion induced by enalapril]. PMID- 15274843 TI - [New insights on pulmonary arterial hypertension]. AB - The 3rd World Symposium on Pulmonary Arterial Hypertension has been a forum for the presentation and discussion of overviews on several aspects of this devastating disease, including pathology and pathobiology, genetics, epidemiology, nomenclature and classification, diagnosis and assessment, medical treatments, interventional and surgical treatments and future directions. This editorial will provide a brief overview of the new findings emerging in this field including new pathologic and pathobiologic concepts, changes in the clinical classification and in the diagnostic definitions. In addition, new treatment strategies and future perspectives will be discussed. PMID- 15274844 TI - [Drug-eluting stents: efficacy, effectiveness, efficiency and evidence]. PMID- 15274845 TI - [Is the prognostic significance of presyncope the same as for syncope?]. PMID- 15274846 TI - [Antiproliferative drug-eluting stents: systematic review of the benefits and estimate of economic impact]. AB - INTRODUCTION AND OBJECTIVES: Antiproliferative drug-coated stents are a possible solution for post-angioplasty coronary restenosis. Here we analyze their efficacy, effectiveness and safety, and estimate the economic impact of their use in Spain. MATERIAL AND METHOD: Systematic review (meta-analysis) of the scientific evidence available up to January 2004, and analysis of hospital costs within a 1-year time horizon. RESULTS: We identified 12 published studies (5 clinical series and 7 RCTs) comparing coated stents (sirolimus or paclitaxel) with conventional stents in patient with de novo single lesions < 30 mm in 2.5 3.5 mm vessels. In nearly all cases the rates of angiographic restenosis and major adverse cardiac events were lower in the coated stent group after 6-12 months. Meta-analysis showed a 69% decrease in revascularization rate (RR=0.31; 95%CI, 0.19-0.51). For every 1000 patients with de novo lesions, the use of a coated stent involved an additional average cost of Euro 818718. The estimated neutral price of a new stent was Euro 1448 at a market price per unit of Euro 2000. CONCLUSIONS: At 12-month follow-up, sirolimus- or paclitaxel-eluting stents were effective and safe in patients with de novo lesions and low or medium risk of restenosis. At current market prices, the widespread use of these stents would involve an increase in health care expenditure for the different sensitivity scenarios we evaluated. More studies are needed to specify the type of patients and lesions likely to obtain the greatest clinical benefit. PMID- 15274847 TI - [Prognosis of presyncope in patients with structural heart disease]. AB - INTRODUCTION AND OBJECTIVES: Few data are available on the prognosis of presyncope in patients with structural heart disease. The aim of this study was to compare the clinical characteristics and long-term prognosis of patients with structural heart disease admitted for presyncope or syncope in the cardiology department of a tertiary hospital. METHODS: We reviewed the medical records of 449 patients (65% men, mean age 66.8 [13.1] years) with structural heart disease admitted because of syncope (n = 272) or presyncope (n = 177) during the period from 1992 to 1998. Clinical and demographic variables were analyzed and the final diagnosis was classified according to European Society of Cardiology criteria. The follow-up (available in 97.1% of patients) consisted of a personal interview with the patient or a review of the medical records and an interview with the relatives of the patients who had died. RESULTS: Both groups had similar demographic and clinical characteristics, except for the presence of atrial fibrillation on admission, which was more common in the presyncope group. Previous syncopal episodes were more frequent in patients admitted for syncope. The mechanism of the episode was considered arrhythmic in 25.7% of the patients with syncope and 22.0% of those in the presyncope group (P=.37). After a mean follow-up of 57.4 [30.5 months the survival curves were similar for both groups and no significant differences were found regarding the causes of death or the rate of sudden death. CONCLUSIONS: The clinical characteristics and the long-term prognosis in patients with structural heart disease admitted to a cardiology department for presyncope are similar to those of patients admitted for syncope. This suggests that the approach to diagnosis and risk stratification should be similar in both groups of patients. PMID- 15274848 TI - [Localization and quantification of myocardium at risk by myocardial perfusion SPECT during coronary artery occlusion]. AB - INTRODUCTION AND OBJECTIVES: The aim of this study was to analyze the distribution and extent of the myocardium at risk using polar maps obtained with myocardial perfusion SPECT. Myocardial perfusion of territories irrigated by the left anterior descending (LAD), right coronary (RCA) and left circumflex artery (CX) was studied with the help of a technetium-radiolabeled tracer during occlusion of the vessels in the course of percutaneous coronary angioplasty. PATIENTS AND METHOD: We studied 50 patients (24 LAD, 15 RCA and 11 CX). The 99mTc compound was injected immediately after inflation of the balloon, and the artery was occluded for approximately 90 seconds. Tomographic images were acquired, and polar maps showing the extent of the ischemic region (uptake < 50% of maximum) were generated. RESULTS: Mean percentage extent of the ischemic territory was 49.8 +/- 10.3% (minimum 35%, maximum 67%), for the proximal LAD, 39.8 (8.3%) (minimum 20%, maximum 51%) for the mid LAD, 20.3 (7.6%) (minimum 8.3%, maximum 35%) for the RCA, and 21.3 (10.8%) (minimum 10.2%, maximum 30%) for the CX. CONCLUSIONS: The contours and extent of the jeopardized myocardial territory found during coronary occlusion allowed us to generate polar maps that illustrated actual coronary risk. The distribution and extent of the areas at risk differed from those in polar maps generated by most current applications used with myocardial perfusion SPECT. PMID- 15274849 TI - [Effectiveness of a quality improvement intervention in reducing cardiovascular risk in hypertensive patients]. AB - INTRODUCTION AND OBJECTIVES: To evaluate the effect of a quality improvement intervention on the reduction of cardiovascular risk in patients with hypertension. PATIENTS AND METHOD: Quasi-experimental study involving two primary care centres. One centre was assigned to implement a quality improvement intervention (n = 482 patients), while at the other center "usual care" procedures were followed (control group, n = 360 patients). The quality improvement intervention consisted of a combined program designed for the medical staff and comprising audit, feedback, training sessions and implementation of clinical practice guidelines during 6 months. The main outcome measures were blood pressure, lipid levels, diabetes, smoking and cardiovascular risk. These values were compared before the intervention and after one year. RESULTS: The baseline characteristics of the patients were similar in both groups. Absolute cardiovascular risk decreased from 15.85% to 14.36% (P< .05) in the intervention group, and no significant change was observed in the control group (15.17% to 15.76%). The intervention led to a 2.07% decrease in cardiovascular risk (95%CI, 1.21-2.93; P< .05). The percentage of patients with high cardiovascular risk (> 20% at 10 years) decreased in the intervention group from 30% to 25%, and increased in the control group from 28% to 30%. Relative cardiovascular risk decreased from 2.03 to 1.75 (P< .05) in the intervention group, and from 1.98 to 1.92 (P> .05) in the control group. The intervention thus led to a 0.25 decrease in relative risk (95%CI: 0.14-0.35). CONCLUSIONS: Absolute and relative cardiovascular risk in patients with hypertension was reduced by a quality improvement intervention. The percentage of patients with high cardiovascular risk was also reduced. PMID- 15274850 TI - [Ambulatory blood pressure monitoring to study white coat syndrome in patients with hypertension seen in primary care]. AB - INTRODUCTION AND OBJECTIVES: The white coat phenomenon is said to occur when the difference between systolic/diastolic blood pressure measured during visits to the doctor's office and in ambulatory recordings is greater than 20/10. These absolute differences, known as the white coat effect, may lead to normotensive patients being classified as having white coat hypertension (WCH). We used ambulatory blood pressure monitoring (ABPM) to monitor the prevalence and response (white coat effect, white coat hypertension or white coat phenomenon) in patients during pharmacological treatment for grade 1 or 2 hypertension, and 4 weeks after treatment was suspended under medical supervision. PATIENTS AND METHOD: Ambulatory blood pressure monitoring was used in 70 patients with hypertension that was well controlled with treatment. Blood pressure was recorded during treatment (phase 1) and 4 weeks after treatment was stopped (phase 2). RESULTS: 18 (26%) of the 70 patients did not participate in phase 2 because when medication was withdrawn, their blood pressure values became unacceptable and it was necessary to restart treatment. The white coat effect was significantly stronger in phase 1, and the prevalence of white coat phenomenon and white coat hypertension did not differ significantly between phases. At the end of phase 2 the prevalence of white coat hypertension was 33%. CONCLUSIONS: Withdrawal of antihypertensive medication in patients with well controlled grade 1 or grade 2 hypertension did not significantly modify the prevalence of white coat phenomenon or white coat hypertension. The white coat effect was greater while patients were on pharmacological treatment. One third of our patients were considered to have been mistakenly diagnosed as having hypertension. PMID- 15274851 TI - [Diabetes mellitus in cardiology practice in Spain. Survey by the Working Group on Heart and Diabetes regarding the importance of diabetes mellitus in relation with other cardiovascular diseases]. AB - INTRODUCTION AND OBJECTIVES: Diabetes mellitus has been defined as a cardiovascular disease of metabolic origin. This article reports the results of a survey of cardiologists regarding their knowledge about this disease and their management of patients with diabetes in daily practice. METHODS: A survey was mailed to all 1840 cardiologists who were members of the Spanish Society of Cardiology, and responses were returned by 348 (18.9%). The survey consisted of 16 questions dealing with organizational and practical aspects of cardiological management for patients with diabetes. RESULTS: The creation of a Working Group on the Heart and Diabetes was judged necessary by 90.2% of the responders. Almost two thirds of the members felt their knowledge of diabetes and its treatment was inadequate, and 82.5% declared that cardiologists should be better able to treat patients with diabetes, since between 15% and 30% of the patients they see have this disease. With respect to secondary prevention, 65.8% of the cardiologists felt that medical care is much better for coronary patients than for patients with diabetes. Among the latter, angiotensin-inhibiting drugs, statins and aspirin are used for secondary prevention. CONCLUSIONS: Because of gaps in the cardiologist's knowledge of how to manage diabetes, the high prevalence of diabetes and its unquestionable impact on the cardiovascular system additional clinical training and educational efforts are needed. One potentially useful measure is the creation of a specific Working Group on the Heart and Diabetes. PMID- 15274852 TI - [Natural history and therapeutic management of acute aortic syndrome]. AB - Acute aortic syndrome, an acute lesion of the aortic wall involving the media, comprises aortic dissection, intramural hematoma and penetrating ulcer. Recent advances in imaging techniques have helped us to understand the natural history and dynamics of this medical condition. Despite considerable advances in surgical treatment, the current high mortality rate in the acute phase can still be reduced by early clinical suspicion and improved surgical expertise. The advent and incorporation of endovascular treatment has opened up new perspectives in the management of this disease, and may improve the long-term prognosis. This article aims to review our current understanding of the natural history and therapeutic management of acute aortic syndrome. PMID- 15274853 TI - [Cardiac resynchronization for heart failure: background, methods, indications and results]. AB - Heart failure is one of the most prevalent diseases in industrialized countries. Although the prognosis of patients with heart failure is still poor, in recent decades new therapies have been investigated in order to improve quality of life and survival. However, up to 30% of the patients with advanced heart failure present disturbances in intraventricular conduction, and this produces asynchrony of ventricular contractility, leading to further deterioration in heart function. Cardiac resynchronization therapy can improve the synchrony of ventricular contractility. Numerous studies have demonstrated the benefits of biventricular stimulation therapy for improving hemodynamic parameters, quality of life, 6 minute walking test performance and functional class in patients with heart failure, ventricular systolic dysfunction and disturbances in intraventricular conduction. Some studies have demonstrated longer survival times in patients treated with cardiac resynchronization plus a defibrillator. Nonetheless, many questions about the benefits of heart resynchronization therapy, site of stimulation and best type of device (pacemaker or defibrillator) remain unresolved. PMID- 15274854 TI - [Magnetic resonance angiography for Crawford type II dissecting thoracoabdominal aortic aneurysm]. PMID- 15274855 TI - [Stenting in primary pulmonary hypertension with compression of the left main coronary artery]. AB - Primary pulmonary hypertension is often associated with angina-like chest pain of uncertain etiology. Left main coronary artery compression by the pulmonary artery is a treatable cause of angina and should be considered in these patients. We describe a patient presenting with primary pulmonary hypertension, clinical angina and extrinsic compression of the left main coronary artery by the pulmonary artery, who was treated with direct stenting. PMID- 15274856 TI - [Closure of an iatrogenic coronary artery fistula with a PTFE-coated stent]. AB - Acquired coronary-cameral fistula is an uncommon disorder. We describe a 50-year old man with rheumatic valvular disease who required emergency mitral and aortic valve replacement due to Staphylococcus aureus acute infective endocarditis. He underwent further surgical interventions due to bleeding and prosthetic dehiscence. During follow-up, a continuous parasternal murmur was noted. Echocardiography showed continuous coronary fistula flow from the left anterior descending artery to the right ventricle. Elective closure of the ostium was achieved with direct implantation of a 3.5 x 16 mm PTFE-coated stent (Jostent Coronary System Graft, Jomed, Germany). PMID- 15274857 TI - [Percutaneous coronary intervention for iatrogenic occlusion of the circumflex artery after mitral anuloplasty]. AB - We describe a patient with obstruction of the dominant circumflex artery after surgical repair of the mitral valve, repaired successfully with percutaneous coronary intervention during the immediate postoperative period. We discuss the etiology, prevention and management of this complication with special emphasis on percutaneous intervention. PMID- 15274858 TI - [Prevalence and clinical course of patients in Spain with acute myocardial infarction and severely depressed ejection fraction who meet the criteria for automatic defibrillator implantation]. AB - The Multicenter Automatic Defibrillator Implantation Trial (MADIT)-II has broadened the indications for cardioverter defibrillator implantation. We present a retrospective study designed to estimate the number of patients in Spain eligible for an implantable defibrillator according to the MADIT-II criteria. From January 1999 to October 2002, 758 consecutive patients were admitted to our center with the diagnosis of acute myocardial infarction. Sixty-seven had a left ventricular ejection fraction < or = 30% (mean, 23[5]) and were not eligible for revascularization. Excluding patients older than 80 years and patients with marked co-morbidity, 47 patients met the MADIT-II criteria for an implantable defibrillator. After a mean follow-up of 18 months, there were 20 deaths, 6 of which were considered sudden. In conclusion, application of the MADIT-II criteria for defibrillator implantation may benefit 6% of the patients with myocardial infarction in Spain. This proportion translates as 4110 defibrillator implantations. PMID- 15274859 TI - [Brucella myopericarditis]. PMID- 15274861 TI - [2003, asthma awareness year]. PMID- 15274862 TI - [Evaluation of the influence of medical education on the smoking attitudes of future doctors]. AB - OBJECTIVE: To evaluate whether medical education influences the prevalence, awareness of, and attitudes to smoking of medical students in the first 3 years of their degree. MATERIAL AND METHODS: In this descriptive longitudinal questionnaire-based study of university students, data for medical and veterinary students in their first 3 years were compared. RESULTS: Of the 226 registered first-year medical students, 181 (80%) returned the questionnaire. Of the 180 first-year veterinary students, 161 (89%) replied to the questionnaire. In the third year, 151 questionnaires from medical students and 139 from veterinary students were returned. There were no differences in age or sex between the 2 groups, and both had more women. The prevalence of smokers increased between the first year and the beginning of the third (from 20% to 31% among medical students and from 28% to 32% among veterinary students). An improvement in the awareness of smoking as a risk factor was observed for medical students, but no change in attitude was observed. CONCLUSIONS: The influence of medical education on this health concern is limited as there is no reduction in the prevalence of smoking. Medical education also seems unable to change attitudes to smoking. More students relate smoking to risk factors for certain diseases, showing greater awareness of the health problems caused by smoking. PMID- 15274863 TI - [Smoking cessation in a population-based cohort study]. AB - OBJECTIVE: To study the incidence rates and the determinants of smoking cessation in a population-based cohort. MATERIAL AND METHODS: We used data from the Cornella Health Interview Survey Follow-up Study. Subjects who declared they were daily smokers at baseline (1994) and had complete follow-up, with information on smoking status in 2002, entered into analysis. We calculated incidence rates and the relative risks of cessation (with 95% confidence intervals) using the Cox model. RESULTS: Out of 353 daily smokers, 100 quit smoking during the follow-up period (cumulative incidence of 28.3%). The incidence rate of cessation was higher among men (42.34 per 1000 person-years) than among women (24.97 per 1000 person-years), with a relative risk of cessation of 1.69 (95% confidence interval, 1.02-2.79) for men. Age and level of education were associated with a higher relative risk of quitting in men. CONCLUSIONS: The main determinants for smoking cessation are sociodemographic (sex, age, and level of education). PMID- 15274864 TI - [Acromegaly and sleep apnea]. AB - OBJECTIVE: Acromegaly is often associated with sleep apnea-hypopnea syndrome (SAHS). The purpose of this study was to understand the prevalence of SAHS in patients with acromegaly and define the characteristics of acromegalic patients with and without SAHS. PATIENTS AND METHODS: The study enrolled 17 patients (11 women and 6 men) residing in the province of Ourense, Spain, who were diagnosed with acromegaly. All patients underwent overnight polysomnography in a sleep laboratory. In addition, growth hormone and insulin-like growth factor 1 levels were assessed. Sixteen of the patients underwent cephalometric study. RESULTS: The average age of the patients was 58 years (95% confidence interval [CI], 52 63). The average body mass index was 31 (95% CI, 29-34) and average neck circumference was 41 (95% CI, 39-43). Ten patients (58.8%) had an apnea-hypopnea index (AHI) greater than 10. Nine had obstructive apnea and one had central apnea. Seven (5 with an AHI>10 and 2 with an AHI<10) reported excessive daytime sleepiness with Epworth scores greater than 10 (41.2%). Five patients (29.4%) were diagnosed with SAHS (AHI>10 and Epworth>10). No correlation was found between an AHI greater than 10 and hormonal activity (P=.082). The mean growth hormone level for patients with an AHI greater than 10 was 4.8 (95% CI, 0.5-9) and the mean for those with an AHI less than 10 was 12 (95% CI, 2-27). Fifty percent of the patients were treated with a somatostatin analog and half of those treated exhibited apnea (P=.302). No cephalometric differences related to the presence of apneas were found. CONCLUSIONS: We found a high prevalence of sleep apneas (58.8%) and SAHS (29.4%), and central apneas were rare. We found no correlation between hormone activity level and the presence of SAHS. The incidence of SAHS was the same in somatostatin analog treated and untreated patients. Cephalometric variables did not distinguish between acromegalic patients with and without SAHS. PMID- 15274865 TI - [Compensatory sweating after upper thoracic sympathectomy. Prospective study of 123 cases]. AB - OBJECTIVE: The most unpleasant consequence of upper thoracic sympathectomy is compensatory sweating (CS). De-pending on the series, the incidence of CS ranges from 24% to 85%. The aim of this study was to determine the relation between CS and the following factors: distribution of hyperhidrosis, procedure performed (unilateral, synchronic bilateral, or sequential bilateral), and number of sympathetic ganglia eliminated. In addition, the degree of patient satisfaction was recorded as objectively as possible. PATIENTS AND METHODS: Prospective study of 123 patients who underwent upper thoracic sympathectomy for palmar and/or axillary hyperhidrosis between 1 January, 1996 and 1 June, 2002 at our unit. All patients completed a questionnaire on symptoms 8 weeks before and after surgery to deter-mine postoperative changes in distribution of the hyperhidrosis and the overall degree of satisfaction on a scale of 0 to 4. RESULTS: The sensation of CS was reported by 86.1% of the patients. When asked to relate this sensation to changes in sweating intensity in specific parts of the body, 46.54% reported CS and 48.37% no change. The trunk was the only region where statistically significant increases in CS occurred; in the feet, a decrease in sweating was noted. No differences in CS were observed with respect to the type of surgery or the number of sympathetic ganglia eliminated. The overall results were considered very satisfactory or quite satisfactory by 84.55% of the patients, while 4.88% were very dissatisfied. CONCLUSIONS: Although CS is a side effect of upper thoracic sympathectomy, not all patients are affected by it. Significant CS occurs mainly in the back, chest, and abdomen. Neither the type of intervention nor the number of ganglia eliminated has an effect on CS. This side effect notwithstanding, overall satisfaction with the treatment is very satisfactory given that the palmar hyperhidrosis is eliminated. PMID- 15274866 TI - [Update to the Latin American Thoracic Association (ALAT) recommendations on community acquired pneumonia]. PMID- 15274868 TI - [Comparison of arterial blood sample kits]. AB - Most inaccuracies in the analysis of gases and electrolytes in arterial blood samples are due to preanalytic factors, among which is the type of equipment used for blood collection. Our objective was to compare arterial blood gas sample kits used under clinical conditions and to evaluate the impact of delay in estimation on variability in results. In 2 types of study we compared 5 kits (Radiometer's Pico 70, Becton Dickinson's Preset, SIMS Portex's Pro-Vent, SIMS-Concord's Pulsator, and Marquest's Quick ABG). In the first study kitsyringe assignment was randomized for collecting arterial blood samples from 160 consecutive patients to evaluate practical aspects of using them and the presence of bubbles in the samples taken. The second study evaluated the effects of delays of 30 and 60 minutes in estimation and of the type of heparin used in 54 blood samples. The kits which produced the fewest bubbles, gave samples with the greatest stability, and had the least impact on ion concentration were Radiometer's Pico 70 and SIMS Portex's Pro-vent. PMID- 15274869 TI - [Large cell lymphoepithelioma-like carcinoma of the lung]. AB - Lymphoepithelioma-like carcinoma of the lung is a rare tumor that is considered a subtype of undifferentiated large cell carcinoma with abundant invasion by lymphocytes. Although initially described as a tumor occurring in the nasopharynx, this type of carcinoma has since been seen in many other organs. We report the case of a 59-year-old male smoker diagnosed with lymphoepithelioma like carcinoma. PMID- 15274870 TI - [Cryptogenic organizing pneumonia and mediastinal lymphadenopathy]. PMID- 15274871 TI - [Bronchiectasis due to Flavimonas oryzihabitans in an immunocompetent patient]. PMID- 15274872 TI - [What is best for lung cancer patients?]. PMID- 15274874 TI - [Congenital toxoplasmosis. A disease with too many questions]. PMID- 15274875 TI - [Prenatal corticosteroid and early surfactant therapy in infants born at < or = 30 weeks gestation]. AB - BACKGROUND: Prenatal corticosteroid (PNC) exposure and postnatal surfactant therapy improve outcome in very low birth weight infants (VLBWI). However, the efficacy of PNC in the prevention of chronic lung disease is debated. OBJECTIVE: To study the effects of PNC exposure on outcome in VLBWI born at < or = 30 weeks' gestation. PATIENTS AND METHODS: We performed a multicenter, longitudinal study. The Spanish Surfactant Group database (n 5 1,275) was searched and 211 VLBWI born at < or = 30 weeks who received early surfactant therapy (< or = 30 min) were identified. Perinatal events, neonatal management and rates of mortality and complications were evaluated. Data on the subgroup of infants who received PNC (157, 74.4 %) were compared with data on 54 infants who did not receive this therapy. RESULTS: Mean (+/- SD) birth weight and gestational age were 944 (226) g and 27 (1.8) weeks. Surfactant was given at 16 +/- 13 min (61 % < or = 15 min). A total of 124 infants (58.8 %) developed respiratory distress syndrome. No differences were found in birth weight, gestational age, or Apgar score at 1 and 5 min. However the age at first surfactant dose was lower in infants exposed to PNC. PNC-exposed infants required fewer doses of surfactant, were extubated earlier (58.9 vs. 161 h) and needed a lower FiO2 at 48 h (0.28 vs. 0.35). Moreover, neonatal mortality (15.9 vs. 27.8 %), the incidence of intraventricular hemorrhage (25.2 vs. 50 %), ductus arteriosus (40.3 vs. 63.5 %) and necrotizing enterocolitis (9 vs. 19.2 %) were lower in infants receiving PNC. However, the incidence of chronic lung disease was similar in both groups. CONCLUSIONS: PNC exposure of VLBWI born at < or = 30 weeks receiving early surfactant therapy reduced mortality and the incidence of certain complications but did not decrease the incidence of chronic lung disease. PMID- 15274876 TI - [Obstructive lung disease after allogenic stem cell transplantation in children]. AB - INTRODUCTION: Bronchiolitis obliterans is recognized as a life-threatening pulmonary complication that can develop 3 months after bone marrow transplantation. OBJECTIVE: To determine the incidence and clinical progression of obstructive lung disease (OLD) in a population of children who had undergone allogenic hematopoietic stem cell transplantation (HSCT). PATIENTS AND METHODS: We examined a sequential sample of 110 patients who received allogeneic HSCT between January 1992 and June 2002. The incidence of OLD in the 77 children who survived for more than 100 days after transplantation was analyzed. The diagnosis of OLD was based on clinical findings with no evidence of infection, pulmonary function test (FEV1/FVC less than 80 % and FEV1 less than 80 % of predicted value) and computed tomography scan. RESULTS: Eight patients (10.4 %) developed OLD at a median time of onset of 184 days after allogenic HSCT (range: 100-1735 days). All patients with OLD had respiratory symptoms. In six out of eight patients airflow obstruction was diagnosed within 1 year of transplantation. All patients showed chronic graft-versus-host disease (GVHD) (p < 0.01). The incidence of OLD in the 23 patients with chronic GVHD was 34.8 %. Two patients (25 %) had a complete response to intensified treatment of chronic GVHD with immunosuppressant therapy. FEV1 declined rapidly in three patients (37.5 %) who died of respiratory failure. Two patients (25 %) had partial reversal but pulmonary function continued below normal values. In one patient (12.5 %) severe obstructive disease was stable. CONCLUSIONS: The time of onset and form of progression of OLD after HSCT may vary. OLD is strongly associated with chronic GVHD and its incidence depends on the number of patients with chronic GVHD. PMID- 15274877 TI - [Graves' disease in children: management and review of 20 patients]. AB - INTRODUCTION: Hyperthyroidism is a rare condition among children and the most common cause is Graves' disease. The best therapy for these patients continues to be debated. PATIENTS AND METHODS: The medical records of 20 patients with Graves' hyperthyroidism who were treated between 1989 and 2003 were reviewed. Clinical symptoms, thyroid function, thyroid autoantibodies, thyroid imaging tests, first line therapy, disease or treatment-induced complications and the need for a secondary treatment option, as well as outcomes, were analyzed. RESULTS: Age at diagnosis ranged from 5 to 16 years and there were more girls than boys (3:1). The most frequent symptom was hyperactivity (58 %). The most frequent sign was goiter (79 %). Thyroid-stimulating immunoglobulin antibodies were found in 90 % of the patients, at the beginning or during the course of the disease. All of the patients received antithyroid medication as first line therapy, but remission was achieved in just one patient. Surgical thyroidectomy was required in three patients, and two patients were treated with radioiodine. CONCLUSION: Because few children achieve remission with medical therapy, other types of treatment (surgery or radioiodine) are often required. Although antithyroid drugs are considered the first choice for treatment in Europe, some authors advocate radioiodine as the treatment of choice. PMID- 15274878 TI - [Study of the clinical and epidemiological characteristics of respiratory infections caused by adenovirus in a pediatric population (1997-2003)]. AB - INTRODUCTION: Infections due to adenoviruses are highly prevalent in pediatric patients. Because the clinical manifestations of the respiratory infections caused by adenoviruses are indistinguishable from those caused by other respiratory viruses, virological methods are required to establish their etiology. We present a retrospective study of the clinical and virological characteristics of patients with isolation of adenovirus in respiratory samples. MATERIAL AND METHODS: From 1997 to 2003 we analyzed 5,746 respiratory samples from pediatric patients (< 15 years old), of which 2,122 (36.9 %) were considered positive. The adenoviruses were isolated in the Hep-2 cell line culture by the shell vial method. RESULTS: Adenovirus was isolated in 100 clinical samples (4.7 % of all positive samples and 1.7 % of all samples studied) in a group of pediatric patients with a mean age of 14 months. The clinical diagnoses of patients were bronchiolitis (61 %), pneumonia (10 %), pertussis-like syndrome (16 %) and asthmatic crisis (11 %). Adenovirus infections mainly presented between December and March. Seventy-two percent of patients had a history of other viral respiratory tract infections and/or bronchial asthma. None of the patients had clinical conjunctivitis and only five patients had diarrhea due to adenoviruses. Seventy percent of the patients received artificial feeding and 30 % were breast fed. Ninety percent of the patients were hospitalized and treatment mainly consisted of bronchodilator agents and antibiotics. CONCLUSIONS: Respiratory tract infections caused by adenoviruses mainly affected patients aged less than 14 months, in the first four months of the year, and with clinical manifestations of bronchiolitis or pneumonia without conjunctivitis. Clinically, these infections are difficult to differentiate from other viral respiratory infections. PMID- 15274879 TI - [Antimicrobial treatment of bronchial infections in hospital emergency rooms]. AB - OBJECTIVE: To analyze antimicrobial prescribing habits in children diagnosed with bronchial infections in hospital emergency rooms. METHODS: A descriptive study was performed in a random sample of children diagnosed with acute bronchitis and bronchiolitis in the emergency rooms of 11 Spanish hospitals. Information about the type of bronchial infection diagnosed and the antimicrobial treatment prescribed was gathered. The appropriateness of antibiotic prescriptions was assessed by comparing clinical practice in the use of antibiotics for bronchial infections with consensus guidelines developed for this study. RESULTS: A total of 731 children were selected. The diagnosis was acute bronchitis in 531 (73 %) and bronchiolitis in 200 (27 %). Antimicrobial treatment was prescribed to 234 children (32 %; 95 % CI: 29-35 %). The most commonly prescribed antimicrobials were the aminopenicillins in 138 children (19 %; 95 % CI 16-22 %), cephalosporins in 54 (7 %; 95 % CI 5-9 %) and macrolides in 45 (6 % 95 % CI 4-8 %). The prescribed treatment was inappropriate in 26 % (95 % CI 23-29 %) of patients [31.5 % (95 % CI 27-35 %) of cases of acute bronchitis and 11.5 % (95 % CI 95 % 7 16 %) of cases of bronchiolitis]. Wide variability was observed in the inappropriate use of antimicrobial agents among the different hospitals, both in acute bronchitis (14-80 %) and in bronchiolitis (0-71 %). CONCLUSION: Inappropriate antimicrobial treatment is prescribed to a considerable proportion of the children with bronchial infections attended in hospital emergency rooms, although there is wide variability among different hospitals. Programs to improve the quality of antimicrobial prescription should be developed. These should combine regulatory and educational measures directed at health professionals and the general public. PMID- 15274880 TI - [Streptococcus pneumoniae and Haemophilus influenzae serotype B invasive disease. A 12-year retrospective study]. AB - BACKGROUND: Streptococcus pneumoniae and Haemophilus influenzae serotype b (Hib) are two of the most common causes of invasive bacterial disease in children. The introduction of a successful Hib vaccine and the emergence of penicillin resistant pneumococci have changed the epidemiology of these bacteria over the last few years. OBJECTIVES: To evaluate the incidence of S. pneumoniae and Hib invasive disease in children (0-15 years) admitted to our hospital during a 12 year period (January 1991-December 2002). METHODS: We retrospectively reviewed the medical records of 101 children with isolation of S. pneumoniae and Hib in blood, cerebrospinal fluid (CSF) or synovial fluid samples during the study period. RESULTS: SP was isolated in 81 samples (72 in blood and nine in CSF) from 73 children (72.6 % were aged < 2 years). The annual incidence per 100,000 children aged 0-2 years was 53.76 cases (95 % CI: 40.7-70.9). The most common clinical diagnosis was bacteremia (67 %). The percentage of penicillin-resistant strains was 44 %. Hib was isolated in 38 samples (26 in blood, 10 in CSF and two in synovial fluid) from 28 children (82.1 % were aged < 2 years). The most common clinical diagnosis was meningitis (35.7 %). The annual incidence per 100,000 children aged between 0-5 years dropped from 21.52 cases (95 % CI: 14.4-32) to 1.7 cases (95 % CI: 0.1-10.8) after systematic vaccination. The percentage of ampicillin-resistant strains was 63.1 %. CONCLUSIONS: The incidence of invasive pneumococcal disease and the increased antimicrobial resistance rates of pneumococcus are two major reasons to support the routine use of pneumococcal conjugate vaccination in children. Systematic Hib vaccination since October 1998 has dramatically reduced the incidence of this disease. PMID- 15274881 TI - [Patient demand and management in a hospital pediatric emergency setting]. AB - OBJECTIVE: To describe patient demand and management in a hospital pediatric emergency setting. PATIENTS AND METHOD: We analyzed the number of episodes registered in our pediatric emergency unit between 1995 and 2002 and performed a retrospective, random survey of 540 episodes in children aged less than 14 years admitted between 2001-10-1 and 2002-09-30. Epidemiological details, physical findings, complementary tests, the treatment administered, admissions, and unscheduled return visits were analyzed. RESULTS: Between 1995-1-01 and 2002-31 12, a total of 337,842 episodes were registered in our emergency unit, requiring 11,767 (3.48 %) admissions to a ward or the pediatric intensive care unit (PICU). The number of episodes/ year increased from 38,659 in 1995 to 51,933 in 2002 (Delta = 34.3 %). The mean age of the sample (n = 540) was 3.5 6 3.2 years (54.6 % were younger than 3 years) and 306 (56 %) were boys. Nearly one-third of the children presented with processes of recent onset (less than 6 hours). The most frequent chief complaints were fever in 160 children (29.6 %), respiratory distress in 78 (14.4 %) and vomiting-diarrhea in 65 (12 %). Complementary investigations were performed in 176 patients (32.6 %), mainly radiologic tests (115; 21.2 %). One hundred fifty-three (28.3 %) received treatment in our emergency unit, mainly antipyretics and bronchodilators. Sixty-five percent stayed less than 1 hour in the emergency unit. The most frequent diagnoses were fever without source in 66 patients (12.2 %), diarrhea/acute gastroenteritis in 40 (7.4 %), asthma in 35 (6.5 %), and croup in 27 (5 %). Sixteen children (3 %) were admitted to a ward and three (0.6 %) were admitted to the PICU. Home drug treatment was recommended in 359 patients (71.8 %). Unscheduled return visits were registered in 59 patients (10.9 %), and five were admitted. CONCLUSIONS: The number of patients attended in our pediatric emergency unit gradually increased between 1995 and 2002, mostly due to young children with fever or respiratory distress. In nearly half of the patients, a thorough anamnesis and physical examination were sufficient for evaluation. Because many of the processes were of recent onset, continuous observation, whether at home or in hospital, was useful to adequately monitor these children. PMID- 15274882 TI - [Joint hypermobility: prevalence and relationship with musculoskeletal pain]. AB - OBJECTIVES: (1) To determine the prevalence of joint hypermobility in children aged 4-14 years old without organic disease of the locomotor system; (2) to compare the prevalence of hypermobility in children with and without arthralgia and (3) to analyze the influence of certain variables on the development of arthralgia. PATIENTS AND METHODS: We performed an observational study in a sample of children aged 4-14 years old living in Area 4 of the Community of Madrid (Spain). Joint hypermobility was evaluated using a goniometer. Hypermobility was defined using Beighton's criteria. RESULTS: A total of 222 subjects were analyzed: 176 in the primary care setting and 46 in the emergency department of a referral hospital. Of the 222 children, 43 reported arthralgia. The prevalence of hypermobility (> or = 4 criteria) was 55 % (123/222), reaching 71 % (49/69) in children aged less than 8 years. No significant differences were found in the prevalence of hypermobility in children with and without arthralgia (65 % and 53 % respectively). Of the variables analyzed (age, sex, country of origin, primary care/emergency department setting) only differences in the absolute number of Beighton criteria present in children with and without arthralgia (4.34 +/- 2.47 and 3.48 +/- 2.35, p = 0.03) were detected, which disappeared when at least four criteria (definition of hypermobility) were required. CONCLUSIONS: Fifty-five percent of the population studied and 71 % of those younger than 8 years old met the criteria for joint hypermobility. In the sample analyzed, the presence of joint hypermobility did not seem to favor the development of arthralgias. PMID- 15274883 TI - [Semiautomatic defibrillation in children]. AB - The main survival factor in cardiac arrest secondary to ventricular fibrillation (VF) is the interval between collapse and defibrillation; consequently, this treatment constitutes one of the most important links in the survival chain in adults. Although VF is a rare cause of out-of-hospital cardiac arrest in children, its detection and treatment is essential because in the pediatric cardiac arrest scenario, VF is the dysrhythmia with the best prognosis. Automated external defibrillators (AED) are simple devices that allow cardiac rhythm to be analyzed; they can also determine whether it is shockable or not with high sensitivity and specificity in adults and children. Currently available evidence has prompted the recommendation of AED use in children older than 1 year without signs of circulation, mainly in the pre-hospital setting and ideally with a dose limiting device. PMID- 15274884 TI - [Identifying children at risk for cardiorespiratory arrest]. AB - Cardiorespiratory arrest in children with severe disease does not usually present suddenly or unexpectedly but is often the result of a progressive deterioration of respiratory and/or circulatory function. Before failure of these functions occurs, there is a series of clinical signs that serve as a warning. Health professionals should not only evaluate clinical signs of respiratory and/or circulatory insufficiency but should also be able to identify these warning signs as early as possible, preferably in the compensation phase, given that the possibility that this process can be reversed by therapeutic measures decreases as the process progresses. PMID- 15274885 TI - [Transverse myelitis in immunocompetent children]. AB - Acute transverse myelitis is an acute inflammatory medullar disease characterized by acute or subacute motor, sensory and autonomic dysfunction. The incidence is low and is estimated at 1-4 cases/10(6) inhabitants per year. In Spain, the disorder is exceptional and most reported cases have occurred in immunodepressed patients. We describe two new cases of transverse myelitis in immunocompetent children and review the etiopathogenesis, diagnosis and outcome of this disorder. PMID- 15274886 TI - [Osteoarthropathy in three siblings with Wilson's disease]. AB - Osteoarthropathies are one of the less usual manifestations of Wilson's disease. They appear in different forms such as osteoporosis (the most frequent), inflammatory changes in small joints, osteomalacia, osteoarthritis in younger ages, spine osteochondritis, fractures and heterotopic ossification. This article describes the different osteoarthropathies in three children: two brothers and one sister with Wilson's disease that first manifested in early childhood with severe neurological signs. After drug treatment and an intense rehabilitation program, the clinical signs stabilized. During the course of the disease, all three children presented fractures of the ulna and radius after low energy trauma, several heterotopic ossifications, some of which were asymptomatic, and inflammatory processes such as hip synovitis and reduction in mineral bone density. The etiology of this kind of manifestation is not yet clear, although the most widely accepted explanation is alteration in calcium and phosphate metabolism. PMID- 15274887 TI - [Tuberculosis in an area with high incidence and a large percentage of immigrants]. PMID- 15274889 TI - [Meningococcal arthritis as the form of presentation of unsuspected invasive disease]. PMID- 15274890 TI - [Bacteremia due to Neisseria subflava biovar flava]. PMID- 15274891 TI - [Septic arthritis due to Kingella kingae: difficulties of diagnosis]. PMID- 15274892 TI - [Simultaneous corticosteroid-sensitive nephrotic syndrome and Hashimoto's thyroiditis]. PMID- 15274893 TI - [Transient third nerve palsy. A case of ophthalmoplegic migraine?]. PMID- 15274894 TI - [Celiac disease, cystic fibrosis and dilated cardiomyopathy]. PMID- 15274895 TI - [Diagnosis of 9p- syndrome at birth. A new case]. PMID- 15274896 TI - [What is your diagnosis? Infraclavicular mass. Myositis ossificans non traumatic]. PMID- 15274897 TI - [The double face of technology: computerisation and the confidentiality of clinical records]. PMID- 15274898 TI - [Domestic violence as a factor associated to women's health problems]. AB - OBJECTIVE: To study the impact of intimate partner violence (IPV) on women's physical and psychological health. DESIGN: Cross-sectional study. SETTING: Primary care centers in 3 Andalusian provinces. PATIENTS: A total of 425 women, aged 18 to 65 years, were recruited following the same randomisation process in 6 primary care centers. MEASUREMENTS: A self-administered structured questionnaire for this study was used to gather the information. As well as sociodemographic variables, the instrument included questions about IPV, physical health indicators (chronic disease and type, lifetime surgeries, days in bed), psychological health (psychological morbidity, use of tranquilizers, antidepressants, pain killers, alcohol and recreational drugs), self-perceived health and social support. RESULTS: Of 425 women, 31.5% ever experienced any type of partner violence. Women experiencing IPV were more likely to suffer a chronic disease. IPV was significantly associated with a number of adverse health outcomes, including spending more than 7 days in bed in the last three months (ORa=2.96; CI 95%, 1.00-8.76), psychological morbidity (ORa=2.68; CI 95%, 1.60 4.49) and worse self-perceived health (ORa=1.89; CI 95%, 1.04-3.43), after controlling for potential confounding variables. CONCLUSION: This study shows that ever experiencing IPV is associated with a worse psychological and self perceived health. Physical injuries are not the only "evidence" of the presence of IPV. Primary health care professionals are in a privileged position to help women who are abused by their partners. PMID- 15274899 TI - [Domestic violence: do we know how violence affects the health of its victims?]. PMID- 15274900 TI - [Consumption of antibiotics and their possible relationship with bacterial resistance in the "Costa de Ponent" health region: analysis of evolution through the initial and end periods of the last decade]. AB - OBJECTIVE: To analyse antibiotic consumption (ABC) and its trend in primary care in an area with a high rate of bacterial resistance during the periods 1993-96 and 2000-02. DESIGN: Comparative, retrospective analysis. SETTING: Costa de Ponent Health Region. Population in 1993-96 period was 1,158,098 inhabitants; in 2000-02 period, 1,188,007 inhabitants. PARTICIPANTS: Those patients in the Area who took AB during the periods under study. MEASUREMENTS: Data obtained from the appropriate records of the Catalan Health Service. Measurement of ABC was the Defined Daily Dose (DDD)/1000 inhabitants/day (DID) [criteria of the WHO Collaborating Centre for Drugs Statistics Methodology]. Trends were quantified as percentage increase or drop, comparing the 1993-96 and 2000-02 periods. The U Mann-Whitney test was used. RESULTS: Overall consumption was high, with a light fall from 17.049 DID during the first period to 15.826 DID in the second (-7.1%). Amoxycillin increased from 4.878 DID (57.6%) to 6.19 DID (67.6%) (+26.8%), but amoxycillin-clavulanic did not rise. The percentages of macrolides, quinolones, cephalosporins, sulphonamides, and tetracyclines were higher in the first period. The variations in the consumption of clarithromycin, azithromycin, cefuroxime, cefixime, and ciprofloxacin between the two periods was the most notable point. CONCLUSIONS: Although overall ABC was high and in accord with the high rate of bacterial resistance in our area, a satisfactory evolutionary trend was found. The fall in consumption in the second period was not significant, but an appropriate modification in its profile was noted: domination of aminopenicillins and drop in macrolides, cephalosporins and fluorquinolones. PMID- 15274901 TI - [Study of the prevalence of urinary incontinence in women from 18 to 65 and its influence on their quality of life]. AB - OBJECTIVE: To find the prevalence of urinary incontinence (IU) in women of working age and its impact on their quality of life (QL). DESIGN: Descriptive, cross-sectional study. SETTING: Urban health district in Ourense. PARTICIPANTS: 1000 women between 18 and 65 were selected at random from a total of 8443. MAIN MEASUREMENTS: The prevalence of UI was evaluated by means of a questionnaire designed for the study and sent by mail. In addition, the Incontinence Impact Questionnaire translated into Spanish was used. This was to assess QL in women with UI. RESULTS: The reply rate was 43.6%, with 20% prevalence of UI. Mean age of the women studied was 43.4 (95% CI, 42.1-44.6); and for those with UI, 50.3 (95% CI, 48.4-52.4). 56.8% of women with UI had not been to the doctor. Mean score of impact on QL (from 0 to 90) was 12.7 points. 87.7% of those affected scored less than 30 points; and 1.5% were above 60 points. CONCLUSIONS: High prevalence of UI was found in women of working age, with low rate of medical consultation, probably because for most women it only lightly affected their QL. PMID- 15274902 TI - [Computerized medical records and confidentiality]. PMID- 15274903 TI - [Legibility: a fundamental factor for understanding a text]. PMID- 15274904 TI - ["Keta" (ketamine): from medication to drug abuse. Bio-psycho-social clinical profile of users and some therapy proposals]. PMID- 15274905 TI - [Depression and epilepsy]. PMID- 15274906 TI - [How much do medicines that are thrown away cost?]. PMID- 15274907 TI - [Electrocardiographic changes and arterial ischemia in migraine transformed by ergotamine consumption]. PMID- 15274908 TI - [Densitometry in primary care, more than ever]. PMID- 15274909 TI - More insights into structural plasticity of actin binding proteins. PMID- 15274910 TI - Protein-protein interfaces are special. PMID- 15274911 TI - The protein surface is a moving target. PMID- 15274913 TI - The linked conservation of structure and function in a family of high diversity: the monomeric cupredoxins. AB - The monomeric cupredoxins are a highly divergent family of copper binding electron transport proteins that function in photosynthesis and respiration. To determine how function and structure are conserved in the context of large sequence differences, we have carried out a detailed analysis of the cupredoxins of known structure and their sequence homologs. The common structure of the cupredoxins is formed by a sandwich of two beta sheets which support a copper binding site. The structure of the deeply buried core is intimately coupled to the binding site on the surface of the protein; in each protein the conserved regions form one continuous substructure that extends from the surface active site and through the center of the molecule. Residues around the active site are conserved for functional reasons, while those deeper in the structure will be conserved for structural reasons. Together the two sets support each other. PMID- 15274914 TI - Structure and mechanism of GDP-mannose glycosyl hydrolase, a Nudix enzyme that cleaves at carbon instead of phosphorus. AB - GDP-mannose glycosyl hydrolase (GDPMH) catalyzes the hydrolysis of GDP-mannose and GDP-glucose to GDP and sugar by substitution with inversion at C1 of the sugar. The enzyme has a modified Nudix motif and requires one divalent cation for activity. The 1.3 A X-ray structure of the GDPMH-Mg(2+)-GDP complex, together with kinetic, mutational, and NMR data, suggests a mechanism for the GDPMH reaction. Several residues and the divalent cation strongly promote the departure of the GDP leaving group, supporting a dissociative mechanism. Comparison of the GDPMH structure with that of a typical Nudix hydrolase suggests how sequence changes result in the switch of catalytic activity from P-O bond cleavage to C-O bond cleavage. Changes in the Nudix motif result in loss of binding of at least one Mg(2+) ion, and shortening of a loop by 6 residues shifts the catalytic base by approximately 10 A. PMID- 15274915 TI - Chitobiose phosphorylase from Vibrio proteolyticus, a member of glycosyl transferase family 36, has a clan GH-L-like (alpha/alpha)(6) barrel fold. AB - Vibrio proteolyticus chitobiose phosphorylase (ChBP) belongs to glycosyl transferase family 36 (GT-36), and catalyzes the reversible phosphorolysis of chitobiose into alpha-GlcNAc-1-phosphate and GlcNAc with inversion of the anomeric configuration. As the first known structures of a GT-36 enzyme, we determined the crystal structure of ChBP in a ternary complex with GlcNAc and SO(4). It is also the first structures of an inverting phosphorolytic enzyme in a complex with a sugar and a sulfate ion, and reveals a pseudo-ternary complex structure of enzyme-sugar-phosphate. ChBP comprises a beta sandwich domain and an (alpha/alpha)(6) barrel domain, constituting a distinctive structure among GT families. Instead, it shows significant structural similarity with glycoside hydrolase (GH) enzymes, glucoamylases (GH-15), and maltose phosphorylase (GH-65) in clan GH-L. The structural similarity reported here, together with distant sequence similarities between ChBP and GHs, led to the reclassification of family GT-36 into a novel GH family, namely GH-94. PMID- 15274916 TI - The first crystal structure of the novel class of fructose-1,6-bisphosphatase present in thermophilic archaea. AB - As the first structure of the novel class of fructose-1,6-bisphosphatase (FBPase) present in thermophilic archaea, we solved the crystal structure of the ST0318 gene product (St-Fbp) of Sulfolobus tokodaii strain 7. The St-Fbp structure comprises a homooctamer of the 422 point-group. The protein folds as a four-layer alpha-beta-beta-alpha sandwich with a novel topology, which is completely different from the sugar phosphatase fold. The structure contains an unhydrolyzed FBP molecule in the open-keto form, as well as four hexacoordinated magnesium ions around the 1-phosphoryl group of FBP. The arrangement of the catalytic side chains and metal ligands is consistent with the three-metal ion assisted catalysis proposed for conventional FBPases. The structure provides an insight into the structural basis of the strict substrate specificity of St-Fbp. PMID- 15274917 TI - Structural basis for the mechanism of Ca(2+) activation of the di-heme cytochrome c peroxidase from Pseudomonas nautica 617. AB - Cytochrome c peroxidase (CCP) catalyses the reduction of H(2)O(2) to H(2)O, an important step in the cellular detoxification process. The crystal structure of the di-heme CCP from Pseudomonas nautica 617 was obtained in two different conformations in a redox state with the electron transfer heme reduced. Form IN, obtained at pH 4.0, does not contain Ca(2+) and was refined at 2.2 A resolution. This inactive form presents a closed conformation where the peroxidatic heme adopts a six-ligand coordination, hindering the peroxidatic reaction from taking place. Form OUT is Ca(2+) dependent and was crystallized at pH 5.3 and refined at 2.4 A resolution. This active form shows an open conformation, with release of the distal histidine (His71) ligand, providing peroxide access to the active site. This is the first time that the active and inactive states are reported for a di-heme peroxidase. PMID- 15274918 TI - Crystal structure of the targeting endonuclease of the human LINE-1 retrotransposon. AB - The human L1 endonuclease (L1-EN) is encoded by the non-LTR retrotransposon LINE 1 (L1). L1 is responsible for more than 1.5 million retrotransposition events in the history of the human genome, contributing more than a quarter to human genomic DNA (L1 and Alu elements). L1-EN is related to the well-understood human DNA repair endonuclease APE1, and its nicking specificity is a major determinant for retrotransposon integration site selection. The crystal structure of human L1 endonuclease is the first of a retrotransposon-encoded protein and a prototype for retrotransposon-encoded endonucleases involved in target-primed reverse transcription. Structure-based endonuclease alignments reveal a conserved threonine in addition to previously identified invariant residues and suggest that DNA recognition proceeds via the accommodation of an extrahelical nucleotide within a pocket of the enzyme. The present analysis will help to refine phylogenetic and functional relationships among metal-dependent phosphohydrolases and provides a basis for manipulating non-LTR retrotransposon integration site selection. PMID- 15274919 TI - The structure of the N-terminal domain of the product of the lissencephaly gene Lis1 and its functional implications. AB - Mutations in the Lis1 gene result in lissencephaly (smooth brain), a debilitating developmental syndrome caused by the impaired ability of postmitotic neurons to migrate to their correct destination in the cerebral cortex. Sequence similarities suggest that the LIS1 protein contains a C-terminal seven-blade beta propeller domain, while the structure of the N-terminal fragment includes the LisH (Lis-homology) motif, a pattern found in over 100 eukaryotic proteins with a hitherto unknown function. We present the 1.75 A resolution crystal structure of the N-terminal domain of mouse LIS1, and we show that the LisH motif is a novel, thermodynamically very stable dimerization domain. The structure explains the molecular basis of a low severity form of lissencephaly. PMID- 15274920 TI - Structure of the actin crosslinking core of fimbrin. AB - Filamentous actin is organized into bundles and orthogonal networks by the fimbrin/alpha-actinin superfamily of F-actin crosslinking proteins. The crystal structure of the Arabidopsis thaliana and Schizosaccharomyces pombe fimbrin cores provides the first description of a functional F-actin crosslinking protein and highlights the compact and distinctly asymmetric organization of the fimbrin molecule, in which the two actin binding domains present distinct surfaces to solvent. The mapping of functionally important residues onto the structure affords new insights into the binding process and provides additional constraints which must be accommodated by models for F-actin binding and crosslinking. Most strikingly, this work provides unique insight into the mechanistic features of conditional-lethal mutants and their extragenic suppressors, which highlight conformational and dynamic properties required for fimbrin function. These results underscore the power of jointly considering structural and genetic suppressor data for obtaining unexpected and biologically relevant mechanistic information. PMID- 15274921 TI - The three-dimensional structure and X-ray sequence reveal that trichomaglin is a novel S-like ribonuclease. AB - Trichomaglin is a protein isolated from root tuber of the plant Maganlin (Trichosanthes Lepiniate, Cucurbitaceae). The crystal structure of trichomaglin has been determined by multiple-isomorphous replacement and refined at 2.2 A resolution. The X-ray sequence was established, based on electron density combined with the experimentally determined N-terminal sequence, and the sequence information derived from mass spectroscopic analysis. X-ray sequence-based homolog search and the three-dimensional structure reveal that trichomaglin is a novel S-like RNase, which was confirmed by biological assay. Trichomaglin molecule contains an additional beta sheet in the HV(b) region, compared with the known plant RNase structures. Fourteen cystein residues form seven disulfide bridges, more than those in the other known structures of S- and S-like RNases. His43 and His105 are expected to be the catalytic acid and base, respectively. Four hydrosulfate ions are bound in the active site pocket, three of them mimicking the substrate binding sites. PMID- 15274922 TI - Protein-protein interactions; coupling of structurally conserved residues and of hot spots across interfaces. Implications for docking. AB - Hot spot residues contribute dominantly to protein-protein interactions. Statistically, conserved residues correlate with hot spots, and their occurrence can distinguish between binding sites and the remainder of the protein surface. The hot spot and conservation analyses have been carried out on one side of the interface. Here, we show that both experimental hot spots and conserved residues tend to couple across two-chain interfaces. Intriguingly, the local packing density around both hot spots and conserved residues is higher than expected. We further observe a correlation between local packing density and experimental deltadeltaG. Favorable conserved pairs include Gly coupled with aromatics, charged and polar residues, as well as aromatic residue coupling. Remarkably, charged residue couples are underrepresented. Overall, protein-protein interactions appear to consist of regions of high and low packing density, with the hot spots organized in the former. The high local packing density in binding interfaces is reminiscent of protein cores. PMID- 15274923 TI - Chromophore conformation and the evolution of tertiary structural changes in photoactive yellow protein. AB - We use time-resolved crystallography to observe the structural progression of a bacterial blue light photoreceptor throughout its photocycle. Data were collected from 10 ns to 100 ms after photoactivation of the E46Q mutant of photoactive yellow protein. Refinement of transient chromophore conformations shows that the spectroscopically distinct intermediates are formed via progressive disruption of the hydrogen bond network to the chromophore. Although structural change occurs within a few nanoseconds on and around the chromophore, it takes milliseconds for a distinct pattern of tertiary structural change to fully progress through the entire molecule, thus generating the putative signaling state. Remarkably, the coupling between the chromophore conformation and the tertiary structure of this small protein is not tight: there are leads and lags between changes in the conformation of the chromophore and the protein tertiary structure. PMID- 15274924 TI - HhaI DNA methyltransferase uses the protruding Gln237 for active flipping of its target cytosine. AB - Access to a nucleotide by its rotation out of the DNA helix (base flipping) is used by numerous DNA modification and repair enzymes. Despite extensive studies of the paradigm HhaI methyltransferase, initial events leading to base flipping remained elusive. Here we demonstrate that the replacement of the target C:G pair with the 2-aminopurine:T pair in the DNA or shortening of the side chain of Gln237 in the protein severely perturb base flipping, but retain specific DNA binding. Kinetic analyses and molecular modeling suggest that a steric interaction between the protruding side chain of Gln237 and the target cytosine in B-DNA reduces the energy barrier for flipping by 3 kcal/mol. Subsequent stabilization of an open state by further 4 kcal/mol is achieved through specific hydrogen bonding of the side chain to the orphan guanine. Gln237 thus plays a key role in actively opening the target C:G pair by a "push-and-bind" mechanism. PMID- 15274925 TI - RIalpha subunit of PKA: a cAMP-free structure reveals a hydrophobic capping mechanism for docking cAMP into site B. AB - In eukaryotes the primary target for cAMP, a ubiquitous second messenger, is cAMP dependent protein kinase (PKA). Understanding how binding and release of cAMP changes the cAMP binding domains and then triggers long-range allosteric responses is an important challenge. This conformational switching requires structure solutions of cAMP binding domains in cAMP-bound and cAMP-free states. We describe for the first time a crystal structure of the cAMP binding domains of PKA type Ialpha regulatory subunit where site A is occupied by cGMP and site B is unoccupied. The structure reveals that the carboxyl terminus of domain B serves as a hydrophobic cap, locking the cyclic nucleotide via its adenine ring into the beta-barrel. In the absence of cAMP, the "cap" is released via an extension of the C-terminal helix. This simple hinge mechanism for binding and release of cAMP also provides a mechanism for allosteric communication between sites A and B. PMID- 15274926 TI - The structure of MSK1 reveals a novel autoinhibitory conformation for a dual kinase protein. AB - Mitogen and stress-activated kinase-1 (MSK1) is a serine/threonine protein kinase that is activated by either p38 or p42ERK MAPKs in response to stress or mitogenic extracellular stimuli. MSK1 belongs to a family of protein kinases that contain two distinct kinase domains in one polypeptide chain. We report the 1.8 A crystal structure of the N-terminal kinase domain of MSK1. The crystal structure reveals a unique inactive conformation with the ATP binding site blocked by the nucleotide binding loop. This inactive conformation is stabilized by the formation of a new three-stranded beta sheet on the N lobe of the kinase domain. The three beta strands come from residues at the N terminus of the kinase domain, what would be the alphaB helix in the active conformation, and the activation loop. The new three-stranded beta sheet occupies a position equivalent to the N terminus of the alphaC helix in active protein kinases. PMID- 15274927 TI - Crystal structure of the DH/PH fragment of Dbs without bound GTPase. AB - Dbl proteins are guanine nucleotide exchange factors for Rho GTPases, containing adjacent Dbl homology (DH) and pleckstrin homology (PH) domains. This domain architecture is virtually invariant and typically required for full exchange potential. Several structures of DH/PH fragments bound to GTPases implicate the PH domain in nucleotide exchange. To more fully understand the functional linkage between DH and PH domains, we have determined the crystal structure of the DH/PH fragment of Dbs without bound GTPase. This structure is generally similar to previously determined structures of Dbs bound to GTPases albeit with greater apparent mobility between the DH and PH domains. These comparisons suggest that the DH and PH domains of Dbs are spatially primed for binding GTPases and small alterations in intradomain conformations that may be elicited by subtle biological responses, such as altered phosphoinositide levels, are sufficient to enhance exchange by facilitating interactions between the PH domain and GTPases. PMID- 15274928 TI - The crystal structure of the carboxy-terminal dimerization domain of htpG, the Escherichia coli Hsp90, reveals a potential substrate binding site. AB - Hsp90 is a ubiquitous, well-conserved molecular chaperone involved in the folding and stabilization of diverse proteins. Beyond its capacity for general protein folding, Hsp90 influences a wide array of cellular signaling pathways that underlie key biological and disease processes. It has been proposed that Hsp90 functions as a molecular clamp, dimerizing through its carboxy-terminal domain and utilizing ATP binding and hydrolysis to drive large conformational changes including transient dimerization of the amino-terminal and middle domains. We have determined the 2.6 A X-ray crystal structure of the carboxy-terminal domain of htpG, the Escherichia coli Hsp90. This structure reveals a novel fold and that dimerization is dependent upon the formation of a four-helix bundle. Remarkably, proximal to the helical dimerization motif, each monomer projects a short helix into solvent. The location, flexibility, and amphipathic character of this helix suggests that it may play a role in substrate binding and hence chaperone activity. PMID- 15274929 TI - E. coli trp repressor forms a domain-swapped array in aqueous alcohol. AB - The E. coli trp repressor (trpR) homodimer recognizes its palindromic DNA binding site through a pair of flexible helix-turn-helix (HTH) motifs displayed on an intertwined helical core. Flexible N-terminal arms mediate association between dimers bound to tandem DNA sites. The 2.5 A X-ray structure of trpR crystallized in 30% (v/v) isopropanol reveals a substantial conformational rearrangement of HTH motifs and N-terminal arms, with the protein appearing in the unusual form of an ordered 3D domain-swapped supramolecular array. Small angle X-ray scattering measurements show that the self-association properties of trpR in solution are fundamentally altered by isopropanol. PMID- 15274930 TI - A computational model of transmembrane integrin clustering. AB - The presented work describes a structural model for integrin homooligomerization, focusing on the transmembrane domains. The two noncovalently linked integrin subunits, alpha and beta, were previously shown to homodimerize or homotrimerize, respectively. Our work is based on published mutational work that induced homotrimerization of beta3 integrins. The mutations provided structural restraints for the creation of a structural model of the beta3 homotrimer by a computational search of the conformational space of homomeric interactions of the beta3 integrin. Additionally, we explored possible conformations of the alphaIIb integrin homodimer, for which no unique solution was found. Two possible models of signal transduction, involving two different alphaIIb conformations, are discussed. One of the possible homodimeric alphaIIb conformations is GpA like, which is in line with experimental evidence. Based on our here-presented structural models and on recent experiments, we will argue that most probably the heteromeric alpha/beta transmembrane complex separates in the course of clustering. PMID- 15274931 TI - Lack of agreement on colour description between clinicians examining childhood bruising. AB - INTRODUCTION: In child protection cases clinicians are often asked to describe and age bruises. This paper looks at both intra- and inter-observer variability in the description of childhood bruising. METHODS: Fifty-eight bruises on 44 children were described by three observers, the bruises were then photographed and the same observers described the bruises at a later date. The descriptions were compared and classified in terms of complete, partial, or no agreement, both between observers and between the in vivo and photographic descriptions. RESULTS: Complete agreement on colour description between two observers in vivo occurred in 27% of descriptions in vivo and 24% of photographs. Only 31% of descriptions completely agreed with the later description of a photograph of the same bruise. CONCLUSIONS: This marked variability in colour description, severely questions the practice of estimating the age of bruises especially from clinical photographs as evidence in child protection proceedings. PMID- 15274932 TI - Methicillin resistant Staphylococcus aureus (MRSA) in an injecting heroin user: implications for hygiene in police station custody suites. AB - A 32 year old male was seen as a detainee, with multiple injection site abscesses. He was followed up in general practice, subsequent to debridement of injection site abscesses on both deltoid regions. Follow up swabs grew Methicillin resistant Staphylococcus aureus (MRSA) from both debridement areas. This case report confirms that MRSA can occur in heroin injection site ulcers in the UK. Such a finding in San Francisco is not unusual. Special care should be taken of post debridement ulcers found in a heroin injecting patient/detainee. MRSA could contaminate the custody area and all police authorities need to have an MRSA policy. This case report has implications for custody, suites in police stations, examination rooms and cell hygiene. PMID- 15274933 TI - Seroprevalence of HIV, HBV and HCV in forensic autopsies, of presumed low risk, in Tehran, the capital of Iran. AB - The purpose of this study is to determine whether autopsies of bodies believed to be from low risk groups, are safe or not. A research study was undertaken to identify the seroprevalence of HIV, HBV and HCV in a presumed low risk forensic autopsy population in Tehran, the capital of Iran. One hundred and seventy three blood samples were collected from cases autopsied at the Tehran Legal Medicine Organization between September 2000 and October 2001. Eight serum samples were positive for HbsAg; 7 were positive for anti-HCV; None was positive for anti-HIV 1, 2. The authors advise that all forensic specimens-even those from bodies presumed to be of low risk, should be treated as potentially infectious and appropriate precautions should be taken when performing autopsies. PMID- 15274934 TI - Torture by excision and ingestion of the ear helix. AB - The forensic medical documentation of torture is important to the legal disposition of cases of alleged crimes against humanity perpetrated on civilian populations. Most injuries that have been attributed to torture are not 'aietologically specific' of torture, and take the form of blunt impact trauma and thermal injuries that are consistent with, but not diagnostic of, alleged torture. In this case report, excision of the helix of the pinna and subsequent forced ingestion of the amputated segment is described in the context of torture. PMID- 15274935 TI - Interrupted aortic arch with ventricular septal defect: an unsuspected fatal congenital anomaly in forensic pathology. AB - Interrupted aortic arch (IAA) is a relatively rare cardiovascular anomaly worldwide and is very rarely encountered in forensic pathology. A case of clinically unsuspected fatal IAA in a 5-day old neonate is described. A full term female baby was declared dead on arrival at an emergency hospital few hours after having been discharged from a maternity hospital following delivery. There were no noticeable signs and symptoms reported during the short postnatal period. Antenatal medical history was unremarkable and there were no abnormalities noticed at birth. Autopsy revealed an unsuspected IAA arch with ventricular septal defect (VSD) as cause of sudden unexpected death. Although amenable to surgical intervention, IAA may not produce obvious clinical symptoms as was the case for this baby. Such sudden unexpected deaths raise medico-legal questions as to the manner and the precise cause of death. The importance of a full forensic autopsy to clarify such a sudden death was re-emphasized. PMID- 15274936 TI - Death from atlanto-occipital dislocation in a boy due to personal water craft collision. AB - Injuries associated with use of personal water craft (PWC) typically include fractures, lacerations, or head injuries. Deaths are rare. We recently encountered an unusual case of a collision between two PWCs in which a 9-year-old boy was struck on the left side, causing the vehicle to rock and him to twist upward and to the right. There was rapid extension, flexion, and rotation of his head. At postmortem examination, it was established that the boy died from atlanto-occipital dislocation and brainstem transection. The risk of atlanto occipital dislocation is high in children under 10. The tectoral ligament which stabilizes the upper cervical spine can be torn or ruptured by hyperextension flexion injury. Some atlanto-occipital dislocations in children can be stabilized and recovery is possible, but in this case of complete ligament rupture and brainstem transection, death was immediate. PMID- 15274937 TI - Why people don't like doctors--medical practice in a dysfunctional society. PMID- 15274938 TI - Advice to FME's. PMID- 15274940 TI - Re: Intracerebral haematoma: traumatic or non-traumatic. PMID- 15274942 TI - Re: European Committee for the Prevention of Torture. PMID- 15274944 TI - Suprapelvic and pelvic phleboliths--a reliable radiographic marker for positive identification. AB - Identification of unknown human remains is often achieved by radiographic comparisons. Among the markers compared in antemortem and postmortem radiographs is the presence of concordant pelvic and supra-pelvic phleboliths which are calcified intravenous blood clots. An illustrative case is presented, where an unidentified decomposed cadaver of an elderly male was positively identified with the aid of medical data provided by the Israel National Police. Two missing persons' records were suggested as possible matches and the concordant presence of phleboliths in one of the cases was useful for the identification. The reliability of phleboliths for radiographic positive identification is discussed along with the etiology and frequency of this phenomenon. PMID- 15274945 TI - Investigating the presence of a common drug of abuse (benzhexol) in hair; the Jordanian experience. AB - The analysis of hair is now accepted as a recognised alternative method for the determination of drug misuse. It has several advantages over the biological fluids; blood and urine, including collection of information regarding long-term drug use and determination of compliance with treatment programmes. In Jordan, the abuse of Artane (benzhexol hydrochloride) has been recognised as the most commonly abused drug among Jordanian youths. Hair samples were collected from nine patients (Male 25-55 years, M=39.11, SD=10.53, CV=26.93%). Samples were analysed for the presence of benzhexol and the toxicological analysis revealed the presence of benzhexol in all samples and its concentration ranged from 0.104 to 7.81 ng/mg hair. Solid phase extraction and GC-MS on selective ion storage (SIS) were used for extraction and detection of the drug with papaverine as external standard. The mass detector was operated at selective ion storage (SIS) to monitor the m/z of 98 and 218 for benzhexol and m/z 339 and 324 for the papaverine. The retention times of benzhexol and papaverine were 6.77 and 12.48 min, respectively. The method was linear in the range of 0.5 to 40 ng/mg hair, with a mean coefficient of determination (R2=0.9982). The limit of detection was 0.04 ng/mg. The intra- and inter-day variations were 3.85% and 3.35%. Recovery was found to be above 90%. PMID- 15274946 TI - Drugs, driving and sobriety tests--a review of recent developments. PMID- 15274947 TI - Pathological findings in overlaying. AB - A 2-month-old girl is reported who was found in cardiorespiratory arrest beneath her unconscious mother. Full autopsy examination failed to reveal any features which would give an indication of the nature of the terminal event. This case confirms the non-specificity of autopsy findings in an infant who suffocated underneath an adult--a situation mimicking classical 'overlaying'. Given that there may be no specific histopathological findings when an infant suffocates underneath an adult, a diagnosis should not be formulated if a full death scene description is not available and no specific lesions or diseases are found. The manner of death should be designated as 'undetermined'. PMID- 15274948 TI - Delayed presentation of carotid dissection, cerebral ischemia, and infarction following blunt trauma: two cases. AB - Carotid artery dissection followed by cerebral infarction as a result of blunt trauma can occur in a number of forensically relevant situations. We describe two such cases. In the first case, a 19-year-old female was involved in a road traffic accident, when her car crashed into the rear of another car. Initially, the young woman presented a minor head injury without loss of consciousness and minor bruising to the left side of the neck. After 48 h, she had developed confusion, speech difficulties, right facial nerve paralysis, and right hemiplegia. CT scan and carotid angiography showed cerebral ischemia with infarction in the territory of the middle left cerebral artery and complete dissection of the left carotid artery. In the second case, a 33-year-old male with depression attempted to hang himself. The rope gave way and he fell down. He had also taken a paracetamol, and a non-steroidal anti-inflammatory drug overdose. He did not lose consciousness but appeared withdrawn and depressed. Approximately 6 h later, his conscious state deteriorated. A CT scan revealed thrombosis of the left internal carotid artery, extending to the middle cerebral artery. The patient died. Both cases reinforce the need for full neurological assessment and review of any individual subject to blunt trauma to the neck, whether accidental or deliberate or where the history is incomplete. In the forensic setting, in particular, RTAs, suspension by the neck, strangulation, and garotting are all instances when examination and assessment must be thorough--and clear advice given--in the absence of any immediate signs or symptoms--that any new symptoms or signs require immediate and thorough neurological investigation. There should be low threshold for prolonged neurological observation or further neurovascular investigations such as ultrasound, CT or MRI scan or angiography, to minimize the risk of developing potentially fatal or incapacitating sequelae. PMID- 15274949 TI - Caveat Emptor: Death involving the use of 4-methoxyamphetamine. PMID- 15274950 TI - Controversies concerning human tissue retention and implications for the forensic practitioner. PMID- 15274951 TI - Epilepsy and the law: an Indian perspective. AB - All over the world epileptic patients have traditionally been discriminated legally in matters affecting their everyday existence. India is no exception. In the current paper certain important issues are explored, namely, epilepsy in relation to marriage, criminality, driving and socio-economic crimes. PMID- 15274952 TI - A case of delayed shock due to dissection of the hyperplastic coronary artery after balloon angioplasty and stenting. AB - A 42-year-old obese woman with a history of liver cirrhosis and diabetes mellitus was admitted because of chest pain. Coronary balloon angioplasty and stenting were performed on the left anterior descending artery (LAD), which was 90% stenotic. She developed moderate shock about 6h later, and about 15 h after the procedure, she died from excessive bleeding from the right femoral artery because of removal of the catheter sheath by herself. Autopsy disclosed haemorrhagic cardiac tamponade and extensive haemorrhage into the epicardial adipose tissue, however, neither coronary perforation nor myocardial rupture was recognized. Histological examination of the dilated coronary segment revealed extensive dissection with an eccentric intimal thickening and the disruption of the adventitia. It was thought to be the origin of the haemorrhage. Although coronary dissection is a well-known complication during cardiac catheterization procedures, this report describes a rare fatal case with delayed onset of shock due to coronary dissection caused by balloon angioplasty and stenting in the presence of eccentric hyperplasia of the vessel wall. PMID- 15274953 TI - Sudden, unexpected death of a cannabis bodypacker, due to perforation of the rectum. AB - A forty year old cannabis bodypacker was found dead in his flat in November 2000, two days after arriving back from a trip to Northern India. On his return he had complained to his family of feeling unwell, although he had refused to let them in or accept medical help. At post-mortem he was found to have 55 packages of cannabis resin in the large intestine, wrapped in cellophane. Subsequent search of the flat by the police revealed the presence of a further 133 similar packages in the fridge, suggesting that he had concealed 188 packages in total. The cause of death was given as peritonitis due to perforation of the distal large intestine caused by swallowing the packages. PMID- 15274954 TI - Detecting organic toxins in possible fatal poisonings--a diagnostic problem. AB - Two fatal cases are reported where there was strong circumstantial evidence of plant toxin ingestion. In only one case however was a low urine level of hyoscine detected (in keeping with a history of Datura sp. consumption). Fatal cases of plant toxin ingestion may be a problem for the laboratory given the wide range and rarity of certain plant poisons, and the limitation of standard screening. Identification of plant materials at the scene of suspected poisoning may be crucial in directing toxicological investigations. PMID- 15274955 TI - Obsessive compulsive disorder secondary to head injury. AB - Obsessive-compulsive disorder (OCD) is a rare sequel of head injury. There are few reports on the subject. This article presents the case of a young patient who developed OCD several years after head injury. In the light of the present state of knowledge, the authors discuss the relationship between OCDS and the brain lesions observed, their localizations and their physiopathological mechanisms, which may explain the occurrence of the disorder. PMID- 15274956 TI - Sexual crimes: different perspectives. AB - In this paper an attempt has been made to emphasize certain facts pertinent to countries in which the legal definition of sexual crimes includes consented heterosexual relations between unmarried adults (fornication). Relevant Articles of one such statute, namely the Libyan penal code, have been cited and discussed. Within that legal definition the presentation of the involved parties would obviously differ from the usual presentation in the Western world where the presence of a victim is almost always essential. Delayed presentation, being the rule rather than the exception, sometimes leaves the examiner no choice but to reflect on clinical examination alone. Although important clues may be obtained, especially from examining the hymen of the alleged virgins, it should only be considered with caution. Without the support of laboratory evidence, the forensic medical examiner's opinion is neither complete nor infallible. Making careful inferences based solely on the morphology of the hymen is sometimes essential, but excessive speculations without discernible grounds should not be a basis for a medico-legal opinion. Guidelines to that effect are also included for the forensic medical examiner to consider. PMID- 15274957 TI - Suicide in Dubai, United Arab Emirates. AB - The annual rate of suicide in Dubai between 1992 and 2000 was 6.2/100,000. There was a non- significant peak incidence in November while July recorded the lowest incidence. Fridays showed the lowest incidence with no significant difference between Muslims and non-Muslims. Victims were predominantly male expatriates with no significant difference between Muslims and non-Muslims. However, a significant increase in the number of non-Indian female expatriates was recorded. The majority of victims were of the age group 21-40 with a significant increase in teenaged females and teenaged citizens. Hanging was the commonest method for committing suicide. Females and non-Indian expatriates resorted significantly to jumping from a high and self-poisoning. The age of the victim had no effect on the choice of the method used to commit suicide. The majority of incidents took place in the victim's own house. A suicide note was left behind in 5% of cases. History of psychological illness or trauma was available in 9.7% of cases. In these cases, depressive illness and recent unemployment were the major triggers for suicidal impulse. A total of 27.7% of screened victims tested positive for alcohol and were predominantly males with no significant difference between Muslims and non-Muslims. PMID- 15274958 TI - Modern forensic medicine and the medico-legal system in Iran. AB - The history of forensic medicine as it is practised today in Iran is described. Modern forensic medicine was not introduced into Iran until the 1850s. At present, there are two organizations in Iran involved in the forensic medicine field: 1) the Universities of Medical Sciences, 2) the Legal Medicine Organization (L.M.O.), and its Medico-Legal Centres (M.L.Cs) in the provinces and cities. These organizations are closely related to each other. The L.M.O. is an official body of the Judiciary, which is situated in Tehran. The L.M.O. and the M.L.Cs are responsible for providing expert witness reports on technical and scientific subjects related to forensic sciences when required by the courts, Judges and the administration. Forensic medicine specialization is achieved after a 3-year full training residency. In Iran, medico-legal autopsies are performed for the investigation of suspicious deaths and crimes. Death investigation in Iran is described. PMID- 15274959 TI - An unusual case of homicide by use of repeated administration of organophosphate insecticides. AB - We present an unusual murder case by use of repeated administration of organophosphate insecticides. A 49-year-old woman suffering from mental retardation, epileptic fits and acromegaly was poisoned by her husband. At first, her death was considered as a 'sudden and unexpected' natural death. Abdominal abscesses of pancreatic origin found at autopsy were compatible with repeated administration of pesticides with anticholinergic action. In her medical history at least one episode consistent with an organophosphate intoxication was retrieved. Thorough inquiry revealed that the victim had ingested phosphamidon and/or omethoate orally. Organophosphate intoxication should be considered when unexplained neurological symptoms are associated with pancreatic disturbances. PMID- 15274960 TI - Adolescent suicide due to inhalation of insect spray. AB - A 17-year-old female who had been expressing suicidal ideation following domestic problems was found dead in her room next to a towel and an empty can of insect spray. There were no suspicious circumstances and it appeared that the deceased had inhaled insect spray using the towel to maintain the concentration of vapour around her nose and mouth. There was no history of recreational inhalation of volatile substances or other illicit drug use. Inhalation of insect spray is a rare method of suicide in keeping with the suggestion that young suicide victims utilize unusual methods of self-destruction determined to some degree by availability and accessibility. PMID- 15274961 TI - The validity of self-reported substance misuse amongst detained persons in police custody. PMID- 15274962 TI - Accidental injury by a grass strimmer: an unusual fatality. AB - The body of a 61-year-old woman was found in the morning lying near a wire fence at her farm where she was cutting grass using a motor-powered strimmer. Autopsy revealed a laceration of the heart apex, haemopericardium and left haemothorax caused by a U-shaped wire projectile which was found lodged in the left thoracic cavity. The mechanistic analysis of the injuries and manner of death, however, were conclusive after a scene visit following the autopsy. The farm was surrounded by a wire fence to prevent wild pigs from entering in. The wire projectile was cut by unshielded blade of the strimmer and struck her chest. This case represents an unusual accidental fatality and indicates the need for improved user education and safety when using such implements. This case also reemphasizes scene investigation as an important part of medicolegal autopsy for proper determination of mechanisms of injuries and precise manner of death. We hope this report will increase awareness and alert the general public to the dangers of the grass strimmer operated without proper safety precautions. PMID- 15274963 TI - The frog and the scorpion: sado-masochistic injury. AB - Interpreting the lesions of victims of sexual violence is one of the most difficult and most controversial areas of forensic medicine. The case we report involves the care of a victim of sado-masochism. It identifies the difficulties of both the forensic and legal management of sexual violence in an unusual context. PMID- 15274964 TI - Sigmoid volvulus and unexpected death in the elderly. AB - An 84-year-old female who was found unexpectedly dead at her home was shown at autopsy to have a massively dilated sigmoid colon with twisting of the lower colon on an unusually long mesentery. The volvulus had caused mechanical obstruction of the bowel with critical reduction of the blood supply resulting in intestinal infarction. Death was due to ischaemic necrosis of the lower large intestine with significant fluid and electrolyte sequestration within the bowel lumen, associated with disseminated sepsis. Gastrointestinal disorders that may result in unexpected death are uncommon in adults and may present atypically in the elderly. The diagnosis of unexpected death due to an infarcted sigmoid volvulus may not be established until an autopsy has been undertaken. PMID- 15274965 TI - Forensic web watch. AB - A search for forensic odontology sites on the Internet revealed thousands of 'hits', of which many constituted dentists offering a private service on their homepages. Several organisations and associations were identified following extensive sifting through these addresses and, despite its growing acceptance in the forensic community, the resources available were superficial and of only moderate educational content. Although many of the sites provided non-specialist information and points of contact for local experts only one was commended for its design and depth. The need for improvement in this field is therefore recommended. PMID- 15274966 TI - The effect of controlled drugs on the unborn child and fetus. AB - Illicit substance abuse is more prevalent than thought in women of a child bearing age and its incidence is increasing. Although maternal factors, such as poor socioeconomic status, diet, smoking, alcohol and infection, have detrimental effects on the fetuses of drug-abusing mothers, harm is increased due to the pharmacological activity of the drugs themselves. This article reviews the pharmacophysiological interactions between mother and fetus, describes the general effects of substance abuse during each trimester and details the deleterious effects on the fetus of the more commonly abused controlled drugs. PMID- 15274967 TI - Sudden death during long distance air travel in an Hb S/C disease patient. AB - Sickle cell disease is an inherited disorder and individuals who are homozygous for the sickle cell gene (HbS/S) show the clinical manifestations of the disease. The individuals who are heterozygous for the sickle cell gene (HbA/S) are referred to as sickle cell trait. In these people, under normal circumstances, symptoms are usually absent or mild. However, thorough investigation of the latter condition is also important, because sickling could occur under certain situations, such as prolonged hypoxia. The level of haemoglobin S(HbS), the ratio of HbS to haemoglobin A (HbA) and the presence of variants such as haemoglobin C (HbC) can alter the entire course of the condition. An unexpected sudden death in a 41-year-old Nigerian, who was apparently in good health and was on a long duration flight, is presented. According to available evidence he was previously diagnosed to be suffering from sickle cell trait. Based on medical advice oxygen was supplied to him throughout the flight. Two hours prior to landing at the international airport in Kuala Lumpur, Malaysia he suddenly became breathless and died shortly after. Autopsy revealed that the immediate cause of death was pulmonary thrombo-embolism originating from calf vein thrombosis. It was also established that the thrombus in the calf vein was not pre-existing. Histology revealed that there was extensive and generalized sickling. Haemoglobin electrophoresis on the postmortem sample of blood confirmed that the deceased had Hb S/C disease and not sickle cell trait. The presence of HbC together with the long hours of flight and associated inactivity had probably complicated the case. Various aspects of the sickle-cell condition are highlighted. Allegations of negligence were made against the airline and the doctor who cleared the deceased in Nigeria (the deceased was employed in a well-known multinational company) for long distance non-stop air travel. Various medico-legal issues pertaining to the cause and mode of death, the importance of an accurate diagnosis of the precise sickling disorder and possible negligence on the part of various agencies are discussed. PMID- 15274968 TI - Sudden and unexpected infant death due to occult lymphoblastic leukaemia. AB - A 16-week-old boy was found moribund in his crib. At autopsy widespread infiltration of multiple organs by lymphoblastic leukaemia (pre-B cell type) was found. Death was attributed to extensive myocardial infiltration by leukaemic cells. There was no associated intracranial haemorrhage. Home video footage taken on the day before death revealed a non-distressed and relatively normal-appearing infant. This case demonstrates an extremely rare cause of unexpected infant death and emphasizes the need for full autopsy examination in such cases. It also confirms that certain infants may have minimal or no symptoms of illness, even in the presence of severe, systemic, life-threatening disease. PMID- 15274969 TI - Intracerebral haematoma: traumatic or non-traumatic. AB - A 35-year-old woman, a chronic alcoholic, died from an intracerebral haematoma 10 hr after she fell downstairs. Some subcutaneous bleeding was seen on the head and face, but there were no new skull fractures and surface contusions of the brain. She appeared to have few predisposing conditions for non-traumatic cerebral haemorrhage. In addition, the haematoma was mainly located "lateral" to the basal ganglia, not where hypertensive bleeding most commonly occurs, and subdural and haemorrhage in the corpus callosum was found with subdural/and subarachnoid haemorrhage. We concluded that on falling a shearing strain from a rotating force produced the intracerebral haemorrhage, but without skull fractures and surface contusions of the brain. She had been admitted to a neurosurgical hospital just 11 months before this incident because of an epidural haemorrhage with left temporal bone fracture. Mild thrombocytopenia was found during that hospitalization. In this report, this abnormality was thought to have some relation to the formation of the huge haematoma occurring after the intracerebral bleeding started. PMID- 15274970 TI - Death on the needle syndrome. AB - A toxicological analysis is mandatory in the investigation of sudden deaths, which in drug abusers may be due to many factors. In some instances the toxicology may be essentially negative and requires an explanation for the cause of death. The case report illustrates a challenge for forensic practitioners when 'death on the needle' may be the mode of death with an anaphylactoid reaction. PMID- 15274971 TI - Forensic web watch. AB - The chosen subject for this month's review is toxicology and covers sites touching upon prescription and illicit drugs, analytical techniques and poisonous plants. It highlights a common problem to the user of the Internet. As more and more people log on and put their web site on for public access, searching for a single, comprehensive, all-encompassing single site becomes almost impossible. Many sites are repetitive or purely personal adverts. Unless you are recommended a site or you are prepared to wade your way through all the junk, one will never find the 'El Dorado' you are seeking. PMID- 15274972 TI - A novel approach to forensic investigation: three-dimensional kinematic and kinetic motion analysis. AB - Following a domestic incident, a five-year-old child was pronounced dead on arrival at hospital. The subsequent post-mortem examination found that the cause of death was peritonitis as a result of a rupture of the duodenum. During the police interview, the main suspect alleged that the injury occurred whilst playing a game which involved standing on the child's abdomen and chest. A reconstruction of the game was performed using 3-dimensional kinematic and kinetic motion analysis to investigate the feasibility of the statement as described by the defendant. Presented is a summary of the investigation together with a description of kinematic and kinetic methods employed. PMID- 15274973 TI - Drugs driving--standardized field sobriety tests: a survey of police surgeons in Strathclyde. AB - OBJECTIVE: To determine the opinion of police surgeons within Strathclyde Police to the proposed introduction of Standardized Field Sobriety Tests (SFSTs) in the assessment of suspect drivers. METHOD: 25 police surgeons who attended a full day training programme in respect of 'Drugs and Driving' received a questionnaire relating to the tests. Following analysis of the responses, an identical questionnaire was posted to all 101 registered police surgeons in Strathclyde, resulting in a 45% response and results compared. RESULTS: Of the conference attendees, 54% of doctors were satisfied with the tests, while 46% expressed reservations. Each test was considered separately, however the Walk and Turn Test and the One Leg Stand Test caused the highest levels of concern from at least 50% of the doctors. POSTAL RESPONSES: 52% were satisfied with the tests, however 48% expressed concerns. Again, the Walk and Turn Test and the One Leg Stand Test were by far the tests causing most concern. CONCLUSIONS: a significant percentage of police surgeons in Strathclyde have expressed concerns regarding the SFSTs. Irrespective of length of experience and postgraduate qualification, the tests most contentious are the Walk and Turn Test and the One Leg Stand Test. PMID- 15274974 TI - A 51-year retrospective study of the trends of height, weight and body mass index at the time of death in those aged 16-103. AB - The study investigates the trends in survival age, height, weight and body mass index (BMI) in both genders for those aged 16-103 at the time of death. It also investigates the trend in the proportion of the population classified as obese using data extracted from 37,206 autopsy reports spanning the period 1947-1997. The data was analysed using the computerised statistical software SPSS with tables and figures produced of mean values per investigation value per sex per 10 year age group per decade. The data confirmed reported trends that the population is getting older, taller and heavier with the mean BMI increasing such that the proportion of individuals who can be classified as obese is increasing. These trends will potentially have an impact on healthcare as more people will have an increased risk of associated morbidity and mortality, as well as an implication on welfare and housing for the increasingly elderly population. PMID- 15274975 TI - Suspicious death related to gamma-hydroxybutyrate (GHB) toxicity. AB - I present a case report of a death following the use of gamma-hydroxybutyrate (GHB) in combination with alcohol; this was the third death reported in the UK, with, to date, at least 27 deaths in America. GHB is also known as one of those used in drug rape; in particular, as it is colourless and odourless, it is very easy to add to drinks; although it has a short half-life of 20 min, the effect is prolonged with the alcohol used to disguise it and it has a marked amnesic effect, making it harder for the victim to present for help and forensic investigation. There is currently a campaign to consider re-classification of gamma-hydroxybutyrate as a Schedule1 drug, whereby possession will become a criminal offence. PMID- 15274976 TI - A case report demonstrating the value of chest X-rays in comparative identification. AB - Comparative radiological identification using bone criteria has mainly been based on cranial characteristics, in particular sinus patterns or bone changes after surgery or trauma. However, the thoracic skeleton, of which antemortem images are frequently available, also provides useful information. We report a case in which ante- and postmortem chest X-rays enabled positive identification of the victim of a road traffic accident. PMID- 15274977 TI - Carbon-monoxide poisoning, behavioural changes and suicide: an unusual industrial accident. AB - We report a case of CO intoxication caused by a motor vehicle's faulty heating system. A truck driver experienced severe mental deterioration, behavioural changes and delirium after acute CO intoxication and committed suicide 15 months later. This report examines the pathogenetic mechanism of CO, the immediate and delayed consequences of CO intoxication, diagnostic difficulties and current treatment options. The medical-legal aspects of the case are discussed. PMID- 15274978 TI - Forensic web watch. AB - The chosen subject for this month's review is toxicology and covers sites touching upon prescription and illicit drugs, analytical techniques and poisonous plants. It highlights a common problem to the user of the Internet. As more and more people log on and put their web site on for public access, searching for a single, comprehensive, all-encompassing single site becomes almost impossible. Many sites are repetitive or purely personal adverts. Unless you are recommended a site or you are prepared to wade your way through all the junk, one will never find the 'El Dorado' you are seeking. PMID- 15274979 TI - Wilm's tumour presenting as acute sexual assault. AB - A 3-year-old presented with severe apparent perineal bleeding. The parents description of a precipitating accident did not provide an adequate explanation for the bleeding. Examination under anaesthesia with police presence revealed no genital injury. Palpation of the abdomen revealed a large renal mass. It was found that the child had a Stage 4 Wilm's Tumour. PMID- 15274980 TI - Substance misuse and the legal system in England and Wales. PMID- 15274981 TI - What is the definition of a poisoning? AB - New insights in medicine and acceptable treatments necessitates an adjustment of the existing definition of clinical or forensic poisoning to: 'An individual's medical or social unacceptable condition as a consequence of being under influence of an exogenous substance in a dose too high for the person concerned'. For medical and legal purposes it is important to know how the victim became poisoned. In general, there are three ways of causing medical poisoning: accidental poisoning, including iatrogenic poisoning, experimental and intentional poisoning. Nowadays iatrogenic intoxication, poisoning caused by the Munchhausen's syndrome (by proxy) and experimental poisoning (designer drugs) have a major place in contemporary toxicology. Although some toxicologists use the word 'intoxication' only overdoses with central effects, in this article 'intoxication' and 'poisoning' are considered to be synonymous. PMID- 15274982 TI - Continuing education programme--Victorian Institute of Forensic Medicine and Department of Forensic Medicine, Monash University. PMID- 15274983 TI - Continuing education programme--Victorian Institute of Forensic Medicine and Department of Forensic Medicine, Monash University. PMID- 15274984 TI - A renaissance in forensic medicine in 2000? PMID- 15274985 TI - Forensic studies of a stabbed infant bottlenose dolphin. AB - A 4-6-week-old male bottlenose dolphin (Tursiops truncatus) was found freshly dead on a beach. He belonged to a nearby dolphin group that was known to have considerable curiosity about human activity and to frequently approach pleasure craft. An autopsy revealed an otherwise healthy animal with death due to a stab wound that had passed completely through the body, incising the aorta. Careful assessment of the wounds indicated that the blade of the weapon had been at least 190 mm in length and 22 mm in width, with a single edge. This report demonstrates that standard forensic techniques may be extremely useful in determining the cause of death in animals, in documenting injuries to assist in the investigation of such cases, and in providing facts to aid in the successful prosecution of those guilty of killing or injuring animals. PMID- 15274986 TI - Medico-legal implications of acute subdural haematoma in boxing. AB - A 16-year-old male high school student with a history of fever (38-39 degrees C) for 4 days lost consciousness following the end of a three round boxing match. He was transferred to a neurosurgical unit located 48 km away 1 h 36 min after injury. Clinical assessment at the unit revealed a comatose patient with a left acute subdural haematoma, but because of advanced brain oedema surgical management was deemed futile. At autopsy an acute subdural haematoma and a severe brain oedema were confirmed. Evaluation of physical conditions before the fight should have been more rigorous in our patient. The observations in our case support the published literature that boxing matches should be held only where neurosurgical expertise is readily available. PMID- 15274987 TI - Continuing education programme--Victorian Institute of Forensic Medicine and Department of Forensic Medicine, Monash University. PMID- 15274988 TI - Sudden cardiac death during sport and recreational activities in Israel. AB - Sudden unexpected death of healthy individuals during strenuous physical activity is rarely encountered. In the present study, 36 cases of death during physical exercise over the past 19 years were examined at the Israel National Center of Forensic Medicine. The victims' ages ranged from 13-62 years; none of them had a pre-existing history of a cardio-vascular condition. Atherosclerotic cardiovascular disease (ASCVD) was the cause of death in most of the older participants in sport activities (35 years of age and over), while younger individuals died mainly as a result of diseases of the myocardium. More than half of the deceased had been found fit to participate in active sports following basic medical examination. We conclude that more detailed medical screening tests should be utilized to detect potentially hazardous cardiovascular diseases in all those engaged in strenuous physical activity. PMID- 15274989 TI - Forensic web watch 4. AB - Finding dedicated sites on the World Wide Web (WWW) touching upon issues related to the autopsy which could be of use to forensic practitioners is, as with other areas of forensic medicine and science, a time-consuming task. Unfortunately, one has to wade through lists related to 'Alien autopsy' sites and even 'Furby autopsy' sites that are generated by the most commonly used web search engines, which have been featured in earlier web reviews. Numerous sites containing large archives of autopsy photographs are available on the web. However, many of these sites represent the darker side of the WWW as they are often presented purely for titillation. Unfortunately, one can equate these sites to the modern-day version of the Victorian 'freak show'--Typically, these sites ask for your Visa card number to view their contents, and several have links to pornography sites; one even links to a Satanist site. Luckily a few of these sites do now require age confirmation codes. As many of these sites show autopsy photographs from real cases one has to ask how these were obtained and who is placing them on the WWW. This review does not list any of these sites for obvious reasons, but it does draw the reader's attention to sites touching upon issues related to autopsies which forensic practitioners may wish to visit or use. PMID- 15274990 TI - Continuing education programme--Victorian Institute of Forensic Medicine and Department of Forensic Medicine, Monash University. PMID- 15274991 TI - Fatal haemorrhage following male ritual circumcision. AB - Lethal complications following ritual circumcision are extremely rare, the most common being sepsis. We present here a case of fatal haemorrhage from a tiny incision of the glans, following a 'home' circumcision of a 6-week-old baby. The post-mortem examination disclosed idiopathic neonatal hepatitis. It is suggested that the previously undiagnosed hepatic condition was responsible for the fatal haemorrhage. PMID- 15274992 TI - Needle phobia. PMID- 15274993 TI - Forensic web watch 3. AB - Since the publication of the first 'Forensic Web Watch' article a new search engine has become available, free of charge at the time of writing, to surfers of the Internet (Net). Fast Search claims to seek out sites of interest for the user from 'all the Web, all the time trade mark ' as opposed to parts of the Net as is more common with other search machines. It is easy to use, extremely fast but as it searches so much more of the Net, the end result is considerably larger. This, in turn, may lead to a longer time to seek out useful information as opposed to the obscure. Having said this, it is recommended to add to your search engine bookmarks. A search for sites on issues related to 'Police Surgeons' will yield limited information, as each country will have a different person filling this role, all referred to by different terms. The one common feature, however, to all such groups as well as forensic pathologists and scientists, is that they will work with, or in some cases for, the police services of their respective country. Thus, in this article we will look at sites related to the police which may have useful information related to their work, specific cases of interest and research and development which may effect our practice. PMID- 15274994 TI - Re: Stark MM, Wells D. Drug-mediated sexual assault. PMID- 15274995 TI - Re: Fitness for interview following alcohol consumption. PMID- 15274996 TI - Traditional Chinese drugs and traditional medicine in Japan. PMID- 15274997 TI - Clients from ethnic and local communities in Greater Manchester attending St. Mary's Sexual Assault Referral Centre. AB - INTRODUCTION: St. Mary's Centre conducts forensic examinations on behalf of Greater Manchester Police. The sexual assault rate (and reporting to police) across ethnic communities is of some debate. An audit was conducted to assess recording of clients' ethnicity, and representation of clients from local and ethnic communities of Greater Manchester County. METHODOLOGY: Performances of recording ethnicity, area of residence, and assault type relevant to all new clients in 2001 (n=653) were measured against standards. Figures relating to local and ethnic populations were also compared to 1991 Census data as a guide. RESULTS: Recording clients' ethnicity and assault type were below required standards, recording area of residence was above. Overall ethnic minority representation was above general population figures, but differences existed within communities. Manchester city resident clients were over-represented compared to other county boroughs. CONCLUSIONS: Factors affecting record-keeping included: client's awareness of the assault; if the client was a police referral; if an examination was conducted. The high number of black clients accounted for the 'over-representation' of ethnic minorities, masking under-representation of other ethnic communities. The 'over-representation' of Manchester city residents is considered to be a function of the Centre's Manchester location, and the city's greater opportunities for late-night socialising. PMID- 15274998 TI - Wound and injury awareness amongst students and doctors. AB - INTRODUCTION: Concern has been raised in the medical press regarding the inability of doctors to describe wounds and injuries of medico-legal significance correctly. METHODS: Medical students and doctors of all grades, from the Department of Surgery at a London teaching-hospital affiliated unit were shown a series of photographs illustrating wounds and injuries expected to be encountered in emergency medical practice. They were asked to identify the type of wound or injury as well as the manner in which that injury may have been caused. RESULT: The results indicated that students and doctors of all grades in this unit were not confident at identifying wounds and injuries, using the correct terminology. The term 'laceration' was widely used to describe incised wounds, and vice versa. Gun shot wounds were poorly identified as such, and were only correctly described by those who clearly 'knew' what they were looking at. CONCLUSION: In order to provide doctors with the skills with which they need to correctly describe wounds and injuries encountered in an emergency setting, this author proposes the introduction of specific teaching in wound identification at undergraduate and postgraduate levels. PMID- 15274999 TI - The BCG scar after percutaneous multiple puncture vaccination may help establish the nationalities of unidentified cadavers. AB - BCG vaccination using the multiple puncture device (the Heaf gun) has been used in Japan and in the United Kingdom. The appearance of the BCG scars therefore differs from that caused by the conventional intradermal BCG vaccination, which is used throughout the world. The aim of this study was to investigate the usefulness of examining BCG scars to identify the nationalities of unidentified cadavers. We investigated the BCG vaccination program not only in Japan, but also in other countries, along with the relation between the BCG scar and nationality. The results showed that the countries which domestically make and use the Heaf gun are Japan (where it is the only method that has been in use since 1968), the United Kingdom (where it has been used, in part, since 1982), and South Africa (where it has been used, in part, since 1972). In addition, for the past 10 years, the Japanese Heaf gun method has been partially applied in the Republic of Korea and in Brazil. The Heaf gun scar can be clearly distinguished from the intradermal scar, and is visible throughout a person's life when good technique is used to administer the vaccination. If the Heaf gun scar is found on the left upper arm of an unidentified Asian cadaver, it is sure to be that of a Japanese. The findings of this present study indicate that the Heaf gun scar can be examined to identify the nationalities of unidentified cadavers. PMID- 15275000 TI - Wounding patterns and human performance in knife attacks: optimising the protection provided by knife-resistant body armour. AB - BACKGROUND: Stab attacks generate high loads, and to defeat them, armour needs to be of a certain thickness and stiffness. Slash attacks produce much lower loads and armour designed to defeat them can be far lighter and more flexible. METHODS AND SUBJECTS: Phase 1: Human performance in slash attacks: 87 randomly selected students at the Royal Military College of Science were asked to make one slash attack with an instrumented blade on a vertically mounted target. No instructions on how to slash the target were given. The direction, contact forces and velocity of each attack were recorded. Phase 2: Clinical experience with edged weapon attacks: The location and severity of all penetrating injuries in patients attending the Glasgow Royal Infirmary between 1993 and 1996 were charted on anatomical figures. RESULTS: Phase 1: Two types of human slash behaviour were evident: a 'chop and drag' blow and a 'sweep motion' type of attack. 'Chop and drag' attacks had higher peak forces and velocities than sweep attacks. Shoulder to waist blows (diagonal) accounted for 82% of attacks, 71% of attackers used a long diagonal slash with an average cut length of 34 cm and 11% used short diagonal attacks with an average cut length of 25 cm. Only 18% of attackers slashed across the body (short horizontal); the average measured cut length of this type was 28 cm. The maximum peak force for the total sample population was 212 N; the maximum velocity was 14.88 m s(-1). The 95 percentile force for the total sample population was 181 N and the velocity was 9.89 m s(-1). Phase 2: 431 of the 500 patients had been wounded with edged weapons. The average number of wounds sustained by victims in knife assaults was 2.4. The distribution of wounds by frequency and severity are presented. CONCLUSIONS: Anti-slash protection is required for the arms, neck, shoulders, and thighs. The clinical experience of knife-attack victims provides information on the relative vulnerabilities of different regions of the body. It is anticipated that designing a tunic-type of Police uniform that is inherently stab and slash resistant will eventually replace the current obvious and often bulky extra protective vest. Attempts at making a combined garment will need to be guided by ergonomic considerations and field testing. A similar anatomical regional risk model might also be appropriate in the design of anti-ballistic armour and combined anti-ballistic and knife resistant armour. PMID- 15275001 TI - The Southwark Police Surgeon Study 1978-1997: alcohol. AB - This paper takes routine clinical data from one Forensic Medical Examiner's (FME) practice over 20 years in the London Borough of Southwark. The relationship of alcohol to gender, categories of call, and patterns of variation over time are addressed. Alcohol related calls are found to be significantly associated with males who are ill or injured, but not with victims or perpetrators of violence. There has also been significant increases in alcohol related calls over the study period. There is no evidence to suggest that was influenced by the introduction of the Police and Criminal Evidence Act (PACE). PMID- 15275002 TI - Periligature injuries in hanging. AB - Hanging is a common mode of suicide while ligature strangulation is a common way of homicide. Ligature marks (patterned abrasion caused by ligature material) can be of great significance to the forensic pathologist in determining the cause and manner of death. Ligature material usually produces a prominent mark over the bight area, which is opposite to the knot and an inverted 'V' pattern at the site of the knot. Apart from ligature mark, sometimes findings such as rope burns & nail marks may be seen around the ligature mark and can be termed as 'periligature injuries'. Nail marks over the neck are usually suggestive of throttling. But they can also be self inflicted by the victim while trying to extricate himself/herself from the strangling grip of hanging or ligature strangulation. Such injuries, when present may mislead the forensic pathologist in drawing conclusion as to whether it is due to hanging or manual strangulation. Rope burn is caused by friction of rope against skin leading to blister formation. Thus it is an antemortem feature. We present two cases of hanging with periligature injuries. PMID- 15275003 TI - Fatal pulmonary thromboembolism following physical torture. AB - Although perpetrators of torture usually use common methods to torture their victim, occasionally they devise novel techniques. Doctors are often called upon to examine victims of torture. This case report is intended to highlight 'the helicopter technique' (also called the, 'parrots perch' in Latin America in 1970s and 1980s) and 'chicken kebab' in Iran (more recently) and the complication of venous thromboembolism. This is a report of a victim who was arrested by the police and subsequently tortured. He died 2 weeks later from pulmonary embolism. The history, physical examination, treatment and autopsy findings of this case are discussed. PMID- 15275004 TI - Artefact in forensic medicine: scrotal mummification. AB - Misinterpretation of postmortem artefacts in forensic medical practice may result in a misdiagnosis of criminal activity. Forensic physicians at the scene can liaise with forensic pathologists and visit the mortuary in the company of the crime scene investigators or scenes-of-crime officers to ensure all relevant facts are identified. In this case a postmortem change created by partial dessication of the scrotal skin was misinterpreted as a suspicious injury. PMID- 15275005 TI - Clinical forensic medicine--management of crime victims from trauma to trial. AB - The loss of human life and function due to violence constitutes a phenomenon that affects millions of patients annually. Society demands an investigation of trauma associated with criminal activity. No longer is it acceptable for health care professionals to operate in isolation of forensic philosophies and principles. It is assumed that the individuals responsible for the performance of the examination of forensic victims have the necessary basic education, experience, and skills. Health care professionals involved in the initial response to these victims, in the emergency department, are faced with unique problems, as social changes require continual reevaluation of standards and professional responsibility. The role of forensic medicine has been expressly designed to provide solutions to some of the most urgent concerns in our society. Forensic medicine focuses on the areas in which medicine and human behavior interface with the law. Existing problems are great and multifaceted and call for new solutions. The application of forensic science to contemporary medical practice reveals a wider role in the investigation of crime and the legal process that contributes to public health and safety. The responsibility of the forensic medicine is to provide continuity of care from the health care institution or the crime scene to courts of law...from trauma to trial. PMID- 15275006 TI - The Laming Inquiry. PMID- 15275007 TI - Coffee may not be good for your health. PMID- 15275008 TI - Forensic web watch--forensic podiatry. AB - A search for forensic podiatry sites on the Internet revealed thousands of 'hits', of which very few were of any educational merit. Following extensive sifting of these addresses, it was found that only a few of the associations for human identification included any information on forensic podiatry methods. The search was also made difficult by many websites failing to make the distinction between studies of footwear prints and forensic podiatry: the study of barefoot impressions. At present, the volume and quality of information on the World Wide Web does not reflect the potential importance of podiatry in forensic investigations. The need for improvement in the quality and available information this field is therefore recommended. PMID- 15275009 TI - Spectrum of unnatural fatalities in the Chandigarh zone of north-west India--a 25 year autopsy study from a tertiary care hospital. AB - A 25 year (1977-2002) autopsy study of 5933 unnatural fatalities from a tertiary care hospital of north-west India revealed an abrupt rise in unnatural deaths (3050; 51.4%) since 1997. 84.2% subjects were between the age group of 16 and 45. Accidental deaths (79.3%) constituted the majority of unnatural fatalities followed by suicidal (13.9%) and homicidal (6%) deaths. Road traffic accidents (RTAs), burn, poisoning, accidental falls from height and firearm injury were responsible for 94.5% of the total unnatural deaths. A male preponderance (73.4%) was seen in all causes of deaths except for burns where females (61%) outnumbered males (39%). The incidence of fatalities due to poisoning and burns had increased from 5.7% and 22.6% to 12% and 24.3%, respectively, whereas due to fire arms and machinery accidents decreased from 4.5% and 1.2% to 1.2% and 0.4%, respectively. The proportion of mortality due to road traffic accidents (50.3%) and accidental fall from height (6.9%) remained almost static. Two-wheeler occupants (motor cycles, etc. 33.3%) were the main victims in road traffic accidents. Pouring of kerosene oil (36% dowry death), malfunctioning and bursting of kerosene oil stove (43.5%) were the most common factors in burn deaths. Between 1977 and 1987 barbiturates (33.3%), organophosphates (23.8%) and copper sulphate (14.3%) and in 1987-1997 organophosphates (45%) and aluminium phosphide (26.5%) were the major fatal poisons. Since 1992 aluminium phosphide (80%), a fumigant pesticide used for wheat preservation was the most common poison. The incidence of suicidal deaths increased from 10.9% (1987-1992) to 15.7% (1997-2002) with a peak incidence of 18.2% in 1992-1997, when this pesticide with no effective antidote was made freely available in the market. PMID- 15275010 TI - A study on the incidence of suicide by hanging in the sub-region of Transkei, South Africa. AB - BACKGROUND: Transkei is a former black homeland in South Africa. It is a historical place and well known for spearheading the freedom fight of South Africa because most of the African National Congress leaders are from this region. It was therefore deprived of development by the apartheid government and is now a poverty stricken area. Transkei is characterized by the lack of infrastructure, and hence a, high rate of unemployment. The majority of individuals are dependent on either the income of migrant mineworkers or subsistence farming at home. OBJECTIVE: To estimate the incidence of suicides in the sub-region of Transkei. METHOD: This is a retrospective record review of deaths due to hanging during the period of January 1993-December 2000 at the Umtata General Hospital. All medico-legal autopsies were recorded in a register at the mortuary. The names, addresses, age, and causes of deaths are found recorded in the register. The catchment areas of these autopsies are Umtata and Nqgeleni magisterial areas which have a population of about 300,000. All autopsy records were collected and analyzed manually. RESULTS: There has been an increasing trend of hanging since 1993, with an overall suicidal death rate increasing from 23.7 per 100,000 in 1993 to 38.6 per 100,000 in 2000. It is observed to be higher in November 32 (13%) and least in September 12 (5%). Over half the deaths 32 (51%) due to hanging were young adults (16-30 years) and 8 (13%) adolescents less than 15 years of age. Twelve (19%) deaths were 31-45 year olds and 5 (7%) were over 61 years. CONCLUSION: There is increasing incidence of deaths due to hanging in Transkei. Nearly two-thirds of them were young adults less than 30 years. RECOMMENDATION: Suicidal tendency in terms of young adults is emerging as an important mental health issue that needs to be addressed. There is need of a well-planned prospective study to be carried out in Transkei. PMID- 15275011 TI - Firearm fatalities. A preliminary study report from Iran. AB - During the one-year period from March 2002 to March 2003 there were 89 firearm fatalities investigated by the Legal Medicine Organization of Iran in Tehran. We determined the characteristics of these 89 firearm deaths which comprised 0.83% of all postmortem examinations. Of these, 60.7% were homicides, 30.3% suicides, 4.5% accidental, and 4.5% unclassifiable. Most victims were young male. Military rifles were responsible for almost all suicides, whereas in homicide handguns were the preferred weapons. The most common site of entrance wounds in suicides were the under the chin (37%) and chest (25.9%). In homicide group, 42.6% of entrance wounds were located in the head. The suicidal gunshots were fired from contact/near contact range in 84.6% while this was the case in only two cases of the homicides. All but one of the suicides were committed at the garrisons or police stations. The unique pattern of suicide that was found in this series was not similar to that reported in earlier studies. We believe our pattern of firearm fatalities must be in great part due to the Iran's strict gun control regulations and cultural background. PMID- 15275012 TI - Self inflicted gunshot wound caused by a home-made gun--medico-legal and ballistic examination. AB - The reconstruction of the mechanism of a fatal gunshot wound inflicted with a zip gun is presented. The joint investigation of medico-legal team and firearms experts teams resulted in an explanation of the unusual distribution of gunpowder encountered around the entry wound. Although the use of crude home-made weapons associated with various criminal activities and suicide is widespread, the forensic literature on this subject is sparse especially concerning tests conducted on the weapon. PMID- 15275013 TI - A fatal case of amlodipine poisoning. AB - A fatal overdose of amlodipine, a calcium channel blocker, in a 50-year-old man is described. Biological samples obtained at autopsy were screened for common drugs and narcotics. The amlodipine determination was made by HPLC with diode array detection and a post-mortem blood concentration of 2.3 mg/kg was determined. The only other drug detected was a blood alcohol concentration of 0.008%. The presence of amlodipine was confirmed in other tissues and in the stomach content. The overdose is assumed to be an accumulation of amlodipine due to the long half-life of this drug. PMID- 15275014 TI - Investigation of suspicious deaths and deaths related to violence--a Malaysian perspective. PMID- 15275015 TI - Medicolegal impact of the new hurt laws in Pakistan. AB - The penal laws in Pakistan went through sweeping reforms in 1979 with intent to bring them in line with the Islamic Jurisprudence. The introduction of these laws repealed the definitions as well as punishments of various types of injuries. Besides that these laws also redefined and reclassified the crime of murder. The laws required that the injuries be identified and documented by an authorized physician. The new classification of injuries and deaths, lack of proper forensic training of the emergency room physicians, performance of medicolegal work by doctors at the earliest stages of their careers and theoretical methods of undergraduate teaching in forensic medicine are matters of concern as they adversely affect the quality of medicolegal work performed in the country. This article gives an overview of the medicolegal system of Pakistan. It also reviews the current laws, their impact on the medical and legal systems of the country and offers some recommendations to correct the existing situation of forensic training and work standards. PMID- 15275016 TI - To legalize physician-assisted suicide or not?--A dilemma. PMID- 15275017 TI - Alcohol-induced hypoglycaemia presenting as drug intoxication. PMID- 15275018 TI - Postinflammatory hyperpigmentation following torture. AB - Hyperpigmentation after torture in darker skinned patients has regularly been noted, although its pathophysiology, and thus its forensic importance, has not previously been documented. Hyperpigmentation is not well described in the dermatological literature. It is the result of inflammation. Any inflammation can cause hyperpigmentation, and the shape of the resulting lesion can closely follow the contours of the site of original inflammatory response. This can be important in correlating the lesion with the alleged cause. It also helps to establish the differential diagnosis of the lesion, which also assists in assessing the degree of consistency between the lesion and the alleged cause. Patterns of hyperpigmentation can therefore, be helpful in assessing allegations of torture months or years after the event. PMID- 15275019 TI - Is post-mortem toxicology quackery? PMID- 15275020 TI - Re: Allegations of rape. PMID- 15275021 TI - Re: Drugs, driving and sobriety tests--authors' response. PMID- 15275022 TI - Forensic web watch--medicolegal aspects of paediatric pathology. AB - The trials over the deaths of Matthew Eappen and Victoria Climbie have highlighted the importance of forensic evidence in cases of suspected child abuse. The debate as to whether bruises, fractures or head injuries have been sustained as a result of previous trauma or non-accidental injury is central to these, and other, cases. A variety of subjects are encountered in forensic paediatric pathology, including Shaken Baby Syndrome, non-accidental injury, retinal haemorrhage, skeletal injury, Sudden Infant Death, sexual abuse and Munchausen Syndrome by Proxy. The coverage of these areas on the internet was assessed using two search engines (Google and the meta-search engine Mamma) and revealed patchy coverage. The majority of sites uncovered were, unsurprisingly, aimed at the layperson concerned by such issues; however, several sites containing useful information for the professional are available. PMID- 15275023 TI - A study on the prevalence of HIV-seropositivity among rape survivals in Transkei, South Africa. AB - BACKGROUND: South Africa has the highest incidence of rape, including child rape, in the world. The country has about 5-million individuals infected with HIV/AIDS. HIV/AIDS is becoming a life-threatening consequence of rape. It is therefore important to provide anti-retroviral drugs and some provinces have already begun to do so. OBJECTIVE: To estimate the prevalence of HIV-seropositivity in the victims of rape in Transkei. METHODS: All the victims of rape who attended Sinawe Rape Crisis Center in Umtata General Hospital during daytime from Monday to Friday between November 2000 and May 2002 were included in the study. All were tested for VDRL and HIV. RESULTS: A total of 243 victims were examined. 22 (9%) were seropositive for HIV. Two blood results were not available. One hundred and sixty six (68.3%) were less than 20 years old, 57 (23.4%) were less than 10 years, and 12 (4.9%) were less than 5 years of age. The highest HIV-positivity (2.8%) was found among the adolescents (15-19 years). No children of less than 5 years were infected with HIV. Only 5 (2.2%) returned for the second HIV test, and one (0.4%) seroconverted after 3 months. CONCLUSION: There were 219 (90%) rape victims who were HIV negative at the time of the incident. Serious consideration must be given to cover rape victims with anti-retroviral agents to prevent them contracting HIV infection. PMID- 15275024 TI - Determinants of suicide in the Transkei sub-region of South Africa. AB - BACKGROUND: South Africa has a history of traumatized citizens and is a society in transition. Suicidal behavior among the black population group in South Africa appears to be on the increase. Under the post-apartheid dispensation they are undergoing a lot of stresses especially in relation to their as yet unmet expectations and demands from the government. OBJECTIVE: To identify the possible determinants of suicide in the Transkei sub-region of South Africa. METHODS: Interviews with relatives of suicide victims, and analysis of victims' demographic, socioeconomic, and psychosocial data. RESULTS: Rural people (> 90%) were much more likely to commit suicide than urban dwellers (< 10%). Suicide notes were left by 13% of the victims. Hanging was the method of choice in 57%, gunshot in 30%, and poisoning in 13%. Among those who died by hanging and by gunshot injuries, males far outnumbered the females (82% and 89%, respectively). By contrast, females constituted the greater proportion of deaths by poisoning (75%). Apparent precipitating factors included economic hardship (87%), alcohol abuse (23%), health related issues (17%), marital problems (13%), and social disputes (10%). The uneducated (70%) and unemployed (64%) used hanging as the method of choice. The highly educated 5 (17%) were all employed, and used a firearm in committing suicide. Family disputes 5 (17%) and separation of parents from teenage children were recorded in 17% of the cases. CONCLUSION: Financial hardship was the main underlying reason, identified in 87% victims of suicide. To break this vicious cycle of unemployment, alcohol abuse, and poor health, a comprehensive poverty alleviation program along with community education could be an important step towards reducing suicides in the Transkei sub-region of South Africa. PMID- 15275025 TI - A simple field test to detect elevated concentrations of carboxyhemoglobin in autopsy blood. AB - Rapid, semi-quantitative measurement of carboxyhemoglobin (COHb) in close temporal proximity to a post-mortem examination can prove to be extremely useful, and provide potentially life-saving information. Other inhabitants of a dwelling with faulty heating or operators of older or poorly maintained motor vehicles with faulty exhaust systems could be quickly informed of potentially dangerous situations. We describe a method for assessing the COHb concentration which will confirm a negative result in 30-40 min and rule out significant levels in less than 2 h, utilizing common reagents and requiring no instrumentation. The method depends on a reaction between formaldehyde and hemoglobin, with associated color change from red to brown. PMID- 15275026 TI - Postoperative bilateral vertebral artery dissection: a case report. AB - With the improvement of medical imaging and surgical techniques, surgery on cervical vertebral is more frequent. Some cases of complications of this type of surgery have been described. We report a case of postoperative bilateral vertebral artery dissection. It concerns a 58 year-old woman who suffered from a left cervico-brachial C6 neuralgia with paresthesiae of the thumb. She underwent discectomy at C5-C6 and C6-C7 followed by setting up intersomatic cages. In subsequent days, an irreversible coma developed. Supra-aortic echographic study revealed bilateral vertebral artery thrombosis. CT scan revealed ischemic lesions of the brain stem and cerebellum. Cerebral death was declared five days after the operation. Autopsy was performed to determine whether death was the consequence of the intervention. The cause of death was determined to be ischemic brain injury of the brain stem and cerebellum resulting from bilateral traumatic occlusion of the vertebral arteries caused by the surgery. PMID- 15275027 TI - Cetacean cafe coronary. AB - The carcass of a young adult male Indian Ocean Bottlenose dolphin (Tursiops cf aduncus) was found floating in an estuary near Adelaide, South Australia. An autopsy revealed that death had been caused by obstruction of the upper aerodigestive tract by a 660 mm Cobbler Carpetshark (Sutorectus tentaculatus). Similar airway obstruction in humans while feeding has been termed cafe coronary syndrome. Although death may have merely resulted from over-enthusiastic feeding, the possibility of neurological impairment was considered, and limited toxicological analyses of tissues was undertaken. No increase in organochlorine pesticides was found, however the possibility of heavy metal poisoning was not excluded. Formal neuropathology was unable to be undertaken. When sudden death in other mammal species mimics cases that are found in humans, similar underlying mechanisms may be present. PMID- 15275028 TI - Flunitrazepam used in a case of poisoning. AB - A male was found in a drowsy condition after being given a cup of tea and a tablet by a neighbour. An ambulance was called, he was taken to hospital and attended the police station on release. A blood sample taken 9h after the incident contained eight times the normal therapeutic level for Flunitrazepam. No motive could be proved and the neighbour was charged with Administering Poison under the Offences against the Person Act of 1861. PMID- 15275030 TI - 'Apparent life threatening events' in sleeping infants: is gastroesophageal reflux ever to blame? PMID- 15275029 TI - Solvent abuse-related toluene leukoencephalopathy. AB - The clinical and neuropathological features of a man with solvent vapour-abuse leukoencephalopathy due to toluene is described. The findings suggest that the clinical spectrum of toluene leukencepathalopathy includes dementia with prominent prefrontal dysfunction. In addition, there was evidence of ongoing white matter damage in the form of persistence of PAS-positive macrophages among the demyelinated fibres in the cerebrum and cerebellum, despite the absolute abstinence from toluene for years prior to death. PMID- 15275032 TI - Case 1--child abuse. PMID- 15275033 TI - Case 2--fitness for interview. PMID- 15275034 TI - Tramadol-benzodiazepines and buprenorphine-benzodiazepines: two potentially fatal cocktails? PMID- 15275036 TI - Re: Letter to the editor: Regarding lack of agreement on colour description in bruising. PMID- 15275035 TI - Re: Drugs, driving and sobriety tests. PMID- 15275037 TI - Forensic web watch. AB - When one thinks of print identification techniques one automatically considers fingerprints. Although finger prints have been in use now for over 100 years there is in fact an older type of identification technique related to prints left at scenes of crime and the anatomy of human body parts. This is the world of ear prints. This short web review considers web sites related to ear print identification particularly the continuing controversy as to whether or not an ear print is unique. PMID- 15275038 TI - Assailant technique in knife slash attacks. AB - Most knife assault victims attending hospital have slash-type wounds, mainly to the face, with fewer over the upper limb and trunk. Only 11% have multiple wounds. Sixty seven male soldiers were asked to slash a vertical human-sized target with a blade. The method of slashing was recorded for each. Approximately, half used multiple strikes, most attacked at the height of the upper torso. This differs from patterns of knife injury seen in clinical practice. The mechanics of fighting in which the victims fend off strikes or disengage is the likely explanation for these differences. PMID- 15275039 TI - Rapid detection of sperm: comparison of two methods. AB - Sperm detection can be an important factor in confirming sexual assault in cases of rape. This paper compares two biochemical methods used in forensic medicine: the first detects the presence of zinc, the second detects acid phosphatase activity. The population studied was composed of 174 consenting women seen at the Male Infertility Center in Toulouse, France. The date of their last sexual intercourse was known accurately. Cytology was the reference test to confirm the presence of sperm in the vaginal samples. We studied the sensitivity, specificity, and positive and negative predictive value of the two biochemical methods. Acid phosphatase detection was the most valuable technique, but its use is limited in time. The zinc test gave disappointing results in our study and does not seem to be a useful reference method for the forensic physician. PMID- 15275040 TI - Use of a pupillometer to assess change in pupillary size post-cannabis. PMID- 15275041 TI - Certification of deaths at Umtata General Hospital, South Africa. AB - BACKGROUND: Audited information on the causes of death is one of the basic components of a country's health information system. Data are usually derived from death certificates. There are many shortcomings in the routine collection of such information. A critical analysis of Umtata General Hospital's (UGH) death certification was done establish how these shortcomings might influence reporting. Such an audit is simple, cheap and useful for monitoring the quality of mortality information. It can be used in health planning and management. OBJECTIVES: To determine the accuracy of death certification by doctors in Umtata General Hospital. METHOD: The study was carried out at UGH. Three hundred and four (304) death notification forms collected over a period of 1 year from January 1999 to December 1999 were studied. These forms were obtained from different clinical departments. RESULTS: The majority of deaths (78.9%) were certified as cardiorespiratory failure. In the remaining (21.1%) cases the causes of death were stated as due to esophageal carcinoma, lobar pneumonia, severe shock, diarrhea, and unknown. CONCLUSION: Doctors at Umtata General Hospital are not experienced in death certification. Cardiorespiratory failure is neither a cause of death nor a mechanism of death. RECOMMENDATIONS: There is need for continuing education on death certification by doctors at UGH. PMID- 15275042 TI - Artefact in forensic medicine: pseudo-scalloped gunshot wound. AB - The forensic pathologist uses a well known rule of thumb for estimating the range or distance between the discharged firearm and the target from entry wound characteristics. In this case, the range was overestimated because pseudo scalloping of a shotgun entry wound was misdiagnosed as lacerations caused by pellet scatter. An unrecognised postmortem artefact which resembles a gunshot injury may be mistaken for a genuine wound feature. PMID- 15275043 TI - Literature review of death certification procedures--international aspects. PMID- 15275045 TI - Sex, pain and parasites. PMID- 15275044 TI - The pathophysiology of cocaine abuse. AB - Cocaine is a naturally occurring alkaloid that increases dopamine concentrations in the reward centers of the brain. There has been a marked increase in cocaine abuse over the last two decades. A neuropsychological stimulant, cocaine also reduces somnolence, increases alertness and improves concentration. However, cocaine abuse has many pathophysiological consequences. These fall broadly into four groups: pathology associated with a drug abusing lifestyle, pathology that occurs whilst intoxicated with (but not directly due to) the drug, pathology associated with drug administration and pathology resulting from pharmacological action of the drug. This review provides a detailed description of the physiological, pharmacological, and pathological effects of cocaine, and highlights the forensic and medicolegal implications of cocaine abuse. PMID- 15275046 TI - Cell-mediated immunity in experimental Trypanosoma cruzi infection. AB - Infection of humans with the protozoan Trypanosoma cruzi leads to Chagas disease, or American trypanosomiasis, a disease that affects nearly 20 million people, and constitutes one of the largest socioeconomic burdens in Latin America. Much of the present knowledge on pathogenic mechanisms underlying T. cruzi infection comes from experimental murine models. Here, George A. DosReis reviews recent findings about the features of host cell-mediated immunity against the parasite and possible mechanisms leading to chronic infection. PMID- 15275047 TI - LD1 amplifications in Leishmania. AB - The genome of Leishmania is quite plastic. Chromosomal rearrangements and DNA amplifications are common events in all the species of the genus. Gene amplification occurs both as a mechanism of drug resistance and in the absence of drug pressure. The best known spontaneous amplification in Leishmania is the so called LD1 family of amplicons. In the past few years there have been great advances in our knowledge of LD1 elements; here, Manuel Segovia and Gines Ortiz review all the available data. PMID- 15275048 TI - Zoonotic transmission of Cryptosporidium parvum: Implications for water-borne cryptosporidiosis. AB - The emergence of Cryptosporidium parvum-associated cryptosporidiosis as a worldwide zoonosis has stimulated interest in the modes of pathogen transmission. Here, Thaddeus Graczyk, Ronald Fayer and Michael Cranfield discuss the complex epidemiology of C. parvum, emphasizing the crosstransmission potential of the pathogen, mechanical vectors involved in water-borne transmission of the oocysts, and factors contributing to contamination of pristine waters with Cryptosporidium. They also outline the public health importance of proper interpretation of positive detection of Cryptosporidium oocysts at water treatment facilities and identify means by which watersheds can be protected from Cryptosporidium contamination. PMID- 15275049 TI - Hemoglobin: Food for thought in vectors and parasites. AB - Several reports indicate that hemoglobin can serve as a source of peptides involved in regulatory functions in mammals, including humans. Here, Rosane Charlab and Eloi Garcia discuss the potential role of hemoglobin-derived peptides as regulatory molecules in blood-sucking vectors and protozoan parasites. PMID- 15275050 TI - A hypothesis about the chronicity of malaria infection. AB - It is generally accepted that malaria evolves as a chronic blood infection by escaping the immune responses directed against a series of antigens that express variable epitopes and/or by selecting parasite populations with distinct polymorphic antigens. However, exacting in vitro studies, performed with clinically well-defined biological material, have correlated the state of protection of African adults (in whom low-grade infection persists) with an indirect defence mechanism where the antibodies are effective owing to their ability to cooperate with blood monocytes. Further studies showed that the antibody bridges the parasite (at the merozoite stage) with a monocyte and triggers the release of mediators which have a parasitistatic, reversible and non antigen-specific effect. The fact that the parasite directly triggers the antiparasite effect leads Pierre Druilhe and Jean-Louis Perignon to formulate here an alternative hypothesis for the chronicity of malaria infection, which would rely on conserved antigenic targets and, in contrast with direct mechanisms, would not select emerging mutated parasites. The above two mechanisms are discussed in the context of their fitness with clinical and parasitological observations. It is proposed that they are not mutually exclusive but, rather, may come into play successively as patients gradually evolve from high-grade symptomatic to low-grade asymptomatic parasitic infection. PMID- 15275051 TI - Presence of the parasitophorous duct in Plasmodium falciparum and P. vivax parasitized Saimiri monkey red blood cells. AB - Although the exchange of metabolites between the intraerythrocytic malaria parasite and the external medium has been studied extensively, the transport of molecules across the erythrocyte cytoplasmic membrane and cytoplasm and the parasitophorous vacuolar membrane needs to be investigated more fully to be completely understood. Recently, the concept of the parasitophorous duct, establishing a continuity between the environment and the vacuolar space surrounding the intraerythrocytic parasite, has been suggested to provide an explanation of how macromolecules can cross two membranes in a cell devoid of an endocytic system. This concept is highly controversial and has been suspected to be an in vitro artefact. In this article, Bruno Pouvelle and Jurg Gysin present evidence of the existence of the parasitophorous duct in Saimiri monkey Plasmodium falciparum- and P. vivax-infected erythrocytes, with a series of ex vivo experiments showing stage and species dependent variations of the characteristics of this structure. PMID- 15275052 TI - More on Chagas disease cardiomyopathy. PMID- 15275054 TI - Why does Plasmodium have a pre-erythrocytic cycle? PMID- 15275056 TI - Protein particle vaccines against malaria. AB - Many viral coat proteins retain the ability to assemble into virus-like particles when produced as recombinant proteins. These small particles are highly immunogenic, and in many cases can be used to carry epitopes or antigens from other pathogens. Most particle-forming proteins can tolerate only small additions or alterations to their sequence, but Hepatitis B virus surface antigen (HBsAg) and the yeast-derived Ty particle are exceptionel in their ability to form particles with long N- or C-terminal extensions. Both have been used to produce hybrid particles carrying Plasmodium sequences. These have been shown to be highly immunogenic in animal studies and also in human phase I trials, in the case of HBsAg. Recently, six out of seven human volunteers were protected against sporozoite challenge by a recombinant HBsAg particle vaccine, the most encouraging result to date for any pre-erythrocytic malaria vaccine. Here, Sarah Gilbert and Adrian Hill review the prospects for the future development of protein particle vaccines against malaria. PMID- 15275057 TI - Signal transduction in malaria parasites. AB - Over the past few years, several reports have been published about the characterization of Plasmodium genes that are thought, on the basis of sequence homology with eukaryotic genes of known function, to be involved in the regulation of growth and differentiation of the parasite. Taken together with phenomenological observations on the regulation of developmental stages in the malaria life cycle, these data form the basis of an informative, albeit incomplete, picture of signal transtruction in Plasmodium. Christian Doerig here reviews Plasmodium elements that are presumably part of major regulatory pathways conserved in eukaryotes, and addresses the problem of how to pursue such studies beyond the stage of gene identification. PMID- 15275058 TI - Mapping malaria risk in Africa: What can satellite data contribute? AB - Recent developments in the access of remotely sensed vegetation and weather data, and their analysis along with other data sources within a geographical information system (GIS), have opened up new possibilities for African health services and research institutes in malaria stratification, monitoring and early warning. Madeleine Thomson, Stephen Connor, Paul Milligan and Stephane Flasse review the current situation and outline the way ahead. PMID- 15275059 TI - Malaria mortality and transmission rates in Africa. PMID- 15275060 TI - Trypanosomes with multicoloured coats. PMID- 15275061 TI - Environmental parasitology: What can parasites tell us about human impacts on the environment? AB - There are a variety of ways that environmental changes affect parasites, suggesting that information on parasites can indicate anthropogenic impacts. Parasitism may increase if the impact reduces host resistance or increases the density of intermediate or definitive hosts. Parasitism may decrease if definitive or intermediate host density declines or parasites suffer higher mortality directly (eg. from toxic effects on parasites) or indirectly (infected hosts suffer differentially high mortality). Although these scenarios are opposing, they can provide a rich set of predictions once we understand the true associations between each parasite and impact. In this review, Kevin Lafferty discusses how parasite ecologists have used and can use parasites to assess environmental quality. PMID- 15275062 TI - The epidemiology and control of cattle schistosomiasis. AB - Schistosomiasis remains a major health problem in much of the developing world. Despite decades of research, many fundamental questions on the dynamics of infection and immunity development remain unanswered. Schistosomiasis is also a common parasitic infection in cattle, and studies on livestock exposed to their own species of schistosome may help in understanding some aspects of the host parasite relationship. Here, Jan De Bont and Jozef Vercruysse review the current knowledge on the epidemiology and control of cattle schistosomiasis. PMID- 15275063 TI - Current views on the population structure of plasmodium falciparum: Implications for control. AB - In recent years there has been a considerable debate on the population genetic structure of malaria parasites. Work on this subject has been revolutionized by the advent of the polymerase chain reaction (PCR) technique, which has made it feasible to study the genetic diversity of parasites in small samples of infected blood, allowing extensive surveys of natural parasite populations to be made. In addition, the technique can be applied to the mosquito stages of the malaria parasite, allowing direct assessments to be trade of the frequency of crossing between parasite clones in Nature. Studies on Plasmodium falcjparum in a wide range of malaria-endemic regions are now revealing the relationship between parasite population structure and malaria epidemiology. In this article, Hamza Babiker and David Walliker review recent work in this field, and discuss how such knowledge might be used to advise on the future deployment of control measures such as antimalarial drugs and possible malaria vaccines. PMID- 15275064 TI - Borrelia burgdorferi genes selectively expressed in ticks and mammals. AB - Lyme disease is a tick-borne infection caused by the spirochete Borrelia burgdorferi. Recent studies have focused on how the Lyme disease bacterium overcomes the challenges faced by an organism that depends on a vector-borne life style. These studies indicate that the spirochete expresses different surface proteins at different stages of its life. Here, Aravinda de Silva and Erol Fikrig review the evidence for differential gene expression and discuss the implications of these findings for the Lyme disease vaccine that is currently being tested in human trials. PMID- 15275065 TI - Heteroduplex analysis in medical entomology: A rapid and sensitive sequence-based tool for population and phylogenetic studies. AB - Classical studies on the insect vectors of parasitic diseases have relied on morphological, biochemical and cytological characters to classify vector species. These are often inadequate for a detailed dissection of the dynamics of the parasite-vector relationship. Molecular techniques have the potential to provide more reliable data on various aspects of vector biology, including population genetics, molecular phylogenetics, and molecular differentiation and classification of closely related species. However, most molecular techniques are expensive, cumbersome or difficult to apply to the analysis of a large number of samples. Here, Jianming Tang and Tom Unnasch discuss the practical advantages of heteroduplex analysis for the study of medically important vectors, using the black flies that serve as vectors for the filarial parasite Onchocerca volvulus as an example. This technique is simple, rapid, inexpensive and capable of detecting minor differences among DNA sequences. PMID- 15275066 TI - Clinical case definition of Schistosoma mansoni. PMID- 15275067 TI - Malaria toxins revisited. PMID- 15275069 TI - What is a parasite strain? PMID- 15275071 TI - Is the high incidence of malaria in alpha-thalassaemic children evidence against the 'malaria hypothesis'? PMID- 15275072 TI - Pghl expression and drug resistance in Saccharomyces cerevisiae. PMID- 15275073 TI - The relative significance of mechanisms of antigenic variation in African trypanosomes. AB - The large number of genes involved in antigenic variation in African trypanosomes has been the focus of a wide literature that describes an almost bewildering array of mechanisms for their differential activation. To the outsider searching for an underlying strategy for antigenic variation, this can appear as a rather disordered and confusing picture. Here, David Barry argues that an understanding of which mechanisms are significant, which ones are primarily inconsequential and which ones perhaps even arise from overdependence on laboratory models, might be achieved by turning attention to trypanosomes that have not undergone adaptation in laboratory conditions. Application of such an approach has led to a proposal for a main mechanism for antigenic variation. PMID- 15275074 TI - Drosophila cellular immunity against parasitoids. AB - Insect host-parasite interrelations involve co-adaptations of considerable complexity. Against endoparasites, immune competent insect hosts initiate a hemocyte-mediated response that quickly destroys the intruders and envelops them in multilayered, melanotic capsules. In this review, Yves Carton and Anthony Nappi focus on recent studies of the cytological, biochemical and genetic mechanisms involved in the cellular immune response of Drosophila against wasp parasitoids. PMID- 15275075 TI - Plasmodium vevax and P. falciparum: Biological interactions and the possibility of cross-species immunity. AB - The question of whether infection of humans with one species of malaria parasite alters the course of infection with another has been largely ignored because no such interaction was found during studies of induced malaria in patients with neurosyphilis. However, in animal model systems some degree of cross-species interaction is the rule rather than the exception. Furthermore, recent epidemiological observations in Vanuatu in the South Pacific have suggested a biological interaction between the dominant species, Plasmodium vivax, and P. falciparum. Kathryn Maitland, Tom Williams and Chris Newbold here speculate on the basis of these observations and other published findings that infection with P. vivax may result in the development of immunity sufficient to ameliorate the clinical course of subsequent infections with the potentially lethal parasite P. falciparum. PMID- 15275076 TI - The acquisition of purines by trypanosomatids. AB - Parasites of the family Trypanosomatidae have an absolute requirement for purines, yet lack the intracellular machinery to synthesize their own purine ring de novo. As a result, the enzymes devoted to the transport and metabolism of purines are extremely important to the parasite. Here, Claudia Cohn and Michael Gottlieb emphasize the value of understanding purine salvage for the development of trypanocidal drugs, and discuss the putative transporters devoted to purine uptake. PMID- 15275077 TI - Leishmania vaccines: old and new. AB - The approach to the development of a Leishmania vaccine has undergone a revolution since its early beginnings with the ancient practice of leishmazation: the inoculation of infectious parasites from an active lesion in order to produce a self-healing lesion in a healthy individual. Controlled infection has been followed by injection of killed parasites and has now progressed to subunit and naked DNA vaccines. Emanuela Handman here discusses the current studies and the future prospects for a Leishmania vaccine with a focus on cutaneous leishmaniasis. Unfortunately, what J.F. Williams said about antiparasite vaccines in 1987 (Ref. 1) is still true in 1997: 'the reasons for optimism are less evident than the reasons for enthusiasm'. PMID- 15275078 TI - Purine salvage enzymes of parasites as targets for structure-based inhibitor design. AB - Nearly 30 years have passed since purine salvage enzymes were first proposed as targets of drugs in the chemotherapeutic treatment of diseases caused by parasites. The rationale behind a structure-based approach to the design of chemotherapeutic agents involves the use of information about substrate preference and the three-dimensional structure of a target enzyme to design potent selective inhibitors of that enzyme. This approach is outlined here by Syd Craig and Ann Eakin, as it applies to the possible design of inhibitors of a purine salvage enzyme, the hypoxanthine phosphoribosyltransferase. PMID- 15275079 TI - On malaria and the cell cycle. PMID- 15275081 TI - Altered membrane phospholipid asymmetry in Plasmodium falciparum-infected erythrocytes. PMID- 15275082 TI - The epidemiology of heartwater: Establishment and maintenance of endemic stability--two comments. PMID- 15275085 TI - Evolutionary origins of trichomonad hydrogenosomes. PMID- 15275086 TI - Combination chemotherapy for Plasmodium falciparum malaria. PMID- 15275087 TI - Multi-gene vaccination against malaria: A multistage, multi-immune response approach. AB - An ideal malaria vaccine will induce immune responses against each stage of the Plasmodium spp life cycle. During its complicated life cycle, the parasite exists extracellularly in the host's bloodstream, within cells that express major histocompatibility complex (MHC) molecules (hepatocytes), within cells that do not express MHC molecules (erythrocytes) and within the mosquito vector. Different arms of the immune system are required to attack the parasite at the different stages. Therefore, a multistage vaccine must be a multi-immune response vaccine. In addition, given the unique antigenicities of the different stages of the life cycle, implicit in this definition is that the vaccine be multivalent. Here, Denise Doolan and Stephen Hoffman present the rationale for developing a multistage, multivalent, multi-immune response malaria vaccine and explain why, among currently available technologies, DNA vaccines may offer the best prospect for success. PMID- 15275088 TI - Amoebapores. AB - The enormous cytolytic potential of Entamoeba histolytica appeals to parasitologists and immunologists because it kills target cells in a contact dependent reaction resembling that of cytotoxic lymphocytes. In this review, Matthias Leippe summarizes what is currently known about a family of pore-forming peptides termed 'amoebapores', to which the cytolytic effect has been attributed, and describes the structural and functional properties of these potent factors, as well as their structure-activity relationships. Finally, a comparison is made with effector molecules of the mammalian defensive system. PMID- 15275089 TI - Survival strategies of Entamoeba histolytica: Modulation of cell-mediated immune responses. AB - Tissue invasion and disease associated with the protozoan Entamoeba histolytica has long been connected with suppression of host cellular immunity. Dampening of the host's defences may facilitate survival of amoebae in extraintestinal sites and development of the characteristic amoebic abscesses. In recent years, several studies have begun to clarify, at the cellular level, the specific effects E. histolytica has on immune cell accessory and effector cell functions. Here, Darren Campbell and Kris Chadee discuss the parasite's multiple modulatory effects on macrophages and T cells and how this manipulation of immune defences may enable the parasite to remain viable in the host. They suggest the putative amoebic molecules involved and potential modulation by the cytokines: interleukins IL-4 and IL-10 and transforming growth factor-beta. PMID- 15275090 TI - The role of extracellular cysteine proteinases in pathogenesis of Entamoeba histolytica invasion. AB - The extracellular cysteine proteinases of Entamoeba histolytica have been implicated as important virulence factors in the pathogenesis of amebiasis and play a key role in tissue invasion and disruption of host defenses. These proteinases have attracted considerable interest as targets for novel therapeutic agents and as vaccine candidates. Here, Xuchu Que and Sharon Reed highlight some of the more recent findings, focusing in particular on functional and structural features of the extracellular cysteine proteinases of E. histolytica. PMID- 15275091 TI - Annual economic loss caused by Taenia saginata asiatica taeniasis in East Asia. AB - Taeniasis is an important medical and economic problem in many countries in East Asia, especially in the mountainous and remote areas where the inhabitants are fond of eating raw or undercooked meat. P.C. Fan here discusses how sociocultural factors and local customs can contribute to the transmission of taeniasis and evaluates the economic losses caused by the disease. PMID- 15275092 TI - An alternative approach to evaluating the intraspecific genetic variability of parasites. AB - Analysis of DNA polymorphisms provides important information for the molecular characterization of parasite strains and clones. Because we still know little about the genomes of parasites, such analysis has to rely on methods applicable to any eukaryotic genome, such as DNA fingerprinting with multilocal minisatellite probes and the polymerase chain reaction (PCR)-based random amplified polymorphic DNA technique (RAPD). However, DNA fingerprinting is cumbersome and needs large amounts of parasite DNA, and RAPD can exhibit low reproducibility and spurious bands, both of which appear to be related to the low stringency of the PCR procedure. Riva Oliveira, Andrea Macedo, Egler Chiari and Sergio Pena here evaluate the applicability to parasites of a technique described two years ago called simple sequence repeat-anchored PCR amplification (SSR-PCR), in which a single primer is needed [the (CA)(8)RY primer] and highstringency conditions are applied. PMID- 15275095 TI - Parateny: When the wrong way can be right. PMID- 15275093 TI - Non-systemic infection in Rhipicephalus appendiculatus ticks. PMID- 15275096 TI - Parasites and cancer. PMID- 15275098 TI - Drug-resistant scrub typhus: Paradigm and paradox. PMID- 15275097 TI - Sterol biosynthesis inhibitors: potential chemotherapeutics against Chagas disease. PMID- 15275099 TI - Why so few transmission stages? Reproductive restraint by malaria parasites. AB - Vast numbers of malaria parasites exist in a population: perhaps 10(10) in just one vertebrate host. Yet gametocytes, the only stage capable of transmission, usually constitute just a few percent or even less of the circulating parasites. Why? Parasite fitness should be intimately linked with transmission probability and infectiousness rises with gametocyte density. Here, Louise Taylor and Andrew Read propose several testable hypotheses that might explain why natural selection has not favoured variants producing more transmission stages. PMID- 15275100 TI - Chagas disease vector control in Central America. AB - As the Southern Cone Initiative proceeds steadily towards eradication of Triatoma infestans, there is increasing interest in applying similar approaches to control Chagas disease vectors in Mexico, Central America and countries of the Andean Pact. Here, Chris Schofield and Jean-Pierre Dujardin discuss the technical feasibility of such an approach. PMID- 15275101 TI - Atypical IgG subclass antibody responses to Plasmodium falciparum asexual stage antigens. AB - The ability of Plasmodium falciparum to induce long-term immunity in the absence of continual restimulation has often been questioned. Recently it has been shown that, while a high proportion of individuals living in areas of high malaria endemicity have antibodies to merozoite surface antigen 2 (MSA2; MSP2) of P. falciparum, these antibodies are primarily of the IgG3 subclass. In this article, Antonio Ferrante and Christine Rzepczyk discuss how such atypical antibody responses may in part explain why immunity to malaria has been widely perceived to be short-lived. PMID- 15275102 TI - Anthelmintic resistance in human helminths: Learning from the problems with worm control in livestock. AB - During the past decade, the prevalence of anthelmintic resistance in some economically important helminths of sheep, goats and horses has increased dramatically. In some regions of Australia, South America and South Africa, anthelmintic resistance has become a serious threat to the survival of the sheep industry. Mass treatment programmes and exclusive reliance on anthelmintics for worm control in livestock are amongst the most important reasons for the development of anthelmintic resistance. In this article, Stanny Geerts, Gerald Coles and Bruno Gryseels draw the attention to a number of errors that have occurred in the control of helminths in livestock and that should be avoided in the control of worms in humans. PMID- 15275103 TI - Schistosomiasis control in the People's Republic of China. AB - Schistosomes, snail-transmitted trematodes (blood flukes), cause a major parasitic disease that ranks second only to malaria in terms of human suffering in the tropics. Schistosoma japonicum has occupied its ecological niche in China for thousands of years; through natural selection it has evolved survival mechanisms that make it difficult (if not impossible) to eradicate. As discussed here by Allen Ross, Li Yuesheng, Adrian Sleigh and Don McManus, vaccination, in combination with current control strategies, may significantly reduce the morbidity of this disease and ultimately improve the quality of life for those living adjacent to endemic zones. This article provides a special focus in Hunan province and examines the potential impact of the Three Gorges Super Dam Project on schistosomiasis control. PMID- 15275105 TI - Autoimmunity in Chagas disease: the mouse model. PMID- 15275107 TI - Defining resistance in Schistosoma. PMID- 15275109 TI - Why does Quinine still work after 350 years of use? PMID- 15275108 TI - Schistosome resistance to praziquantel: Fact or fiction? PMID- 15275110 TI - Malaria transmission and vector control. PMID- 15275111 TI - The pathogenesis of chorioretinal disease in onchocerciasis. AB - The pathogenesis of onchocercal chorioretinopathy is poorly understood. In this article, Philip Cooper, Ronald Guderian, Roberto Proano and David Taylor discuss the important clinical, histological and epidemiological features of the resulting lesions that cause blindness, and review the numerous mechanisins that have been put forward to explain its pathogenesis. The pathogenesis of anterior segment disease, particulary sclerosing keratitis, has been reviewed in depth previously(1) and will not be discussed here. PMID- 15275112 TI - Recent developments in transgenic insect technology. AB - In this short review, David O'Brochta and Peter Atkinson examine recent progress in the development of transgenic insect technology. To date, only Drosophila melanogaster and a few closely related species can be routinely transformed; transformation is far from routine in all other insects. The key bottleneck that has impeded progress has been the identification of transposable elements or viruses that are mobile in target species such as the mosquito, Anopheles gambiae. These mobile genetic elements will serve as platforms upon which effective gene vectors, genetagging agents and enhancer traps will be designed and constructed. Significant progress has been made on a number of research fronts. PMID- 15275113 TI - Antigens of trichinella spiralis. AB - Following infection with Trichinella spiralis, the host elicits a strong immune response that causes rapid expulsion of parasites, a reduction in reproductive capacity of the remaining parasites a reduction in the number of larvae recovered from host muscles, and impairment of the mobility of worms in the intestines. The need to identify the antigens that evoke such responses is twofold: (1) to develop immunodiagnostic tools; and (2) to understand the basis of protective immunity. In this review, Yuzo Takahashi describes the antigenic profiles of T. spiralis, emphasizing immunocytochemical findings. PMID- 15275114 TI - Quantitative aspects of the relationship between the sickle-cell gene and malaria. AB - The relationship between resistance to Plasmodium falciparum infection and the frequency and distribution of the sickle-cell gene in populations exposed to endemic malaria transmission is reducible to clear and quantifiable terms. In this review, Trevor Jones examines the prediction of gene frequency changes under selective pressure, the selective advantage to the heterozygote (balanced polymorphism) that the sickle-cell gene provides to individuals in areas with malaria transmission, and the relationship between sickle-cell gene frequency and malaria, as measured by, for example, sporozoite rate and basic reproduction rate. He seeks to clarify what one can infer about malaria transmission from an analysis of the distribution and inheritance patterns of the sickle-cell gene and sickle-cell disease and under what circumstances these inferences should be made. PMID- 15275115 TI - Geographic information systems and the distribution of Schistosoma mansoni in the Nile delta. AB - New computer-based sensor technology and geographic methods have led to emerging interest in use of satellite environmental assessment tools for design of disease control programs, especially for those that are vector borne. The long-range goal of work reported here by John Malone and colleagues on behalf of this Egyptian Ministry of Health-USAID Schistosomiasis Research Project team (Box 1) is to utilize data from sensor systems on board earth-observing satellites to develop more-sensitive disease-prediction and -control models. If successful, methods developed may provide a potentially vital capability for use by disease control program managers, particularly in less-developed countries, where mapping resources are not well advanced. Longer term, broader basic questions on the interaction of environment and disease in anticipation of predicted global climate change may be addressed. These studies focused on the lower Nile river basin of Egypt. The specific objective was to link data on environmental requirements for propagation and transmission of schistosomiasis with parameters measurable from space. PMID- 15275116 TI - Extracellular matrix: A tool for defining the extracorporeal function of parasite proteases. AB - The significance of cysteine protease activity present in excretory/secretory products of the feeding stages of Haemonchus contortus is discussed here by Marcia Rhoads and Raymond Fetterer. Based, in part, on the in vitro degradation and uptake of extracellular matrix components by live parasites, they argue that the cysteine proteases have an essential extracorporeal function in the digestion of host tissues. They also outline the merits of the extracellular matrix model, which mimics the in vivo structure of connective tissue and basement membranes, in analyzing host-parasite interactions and (possibly) parasite developmental processes. PMID- 15275117 TI - Focus on the effect of bednets on malaria morbidity and mortality. PMID- 15275118 TI - Concerns on long-term efficacy of an insecticide-treated bednet programme on child mortality. PMID- 15275119 TI - A call for integrated approaches to controlling malaria. PMID- 15275121 TI - The need for assays predictive of protection in development of malaria bloodstage vaccines. PMID- 15275122 TI - HLA class I-associated resistance to severe malaria: a parasitological re assessment. PMID- 15275123 TI - Probing the nematode surface. AB - The surface of parasitic nematodes has been well studied with respect to its structural and immunological properties, but little is known about its biophysical nature and the role this plays in the host-parasite relationship. In this article, Clare Roberts and Jay Modha highlight some biophysical features of nematode surfaces and discuss their recent findings regarding mechanisms controlling surface-associated biophysical phenomena observed in parasitic nematodes during infection or culture in medium simulating the mammalian host environment. The nematode surface is distinct from the plasma membrane, nevertheless some parallel features exist and are described. PMID- 15275124 TI - Rab GTPases and the unusual secretory pathway of plasmodium. AB - A detailed analysis of some of the unusual features of secretory protein trafficking in Plasmodium has been hindered by the paucity of markers available for identifying the different compartments of the parasite's secretory apparatus. Gary Ward, Lew Tilney and Gordon Langsley here outline what is currently known about the secretory pathways of Plasmodium falciparum, and discuss how the recent description of a family of parasite rab genes is being used to generate a set of compartment-specific markers. They illustrate this point by describing studies with PfRab6, an established Golgi marker in other eukaryotic cells, which argue in favor of a functional Golgi in Plasmodium spp. PMID- 15275125 TI - A role for the enteric nervous system in the response to helminth infections. AB - The enteric nervous system (ENS) in the gut contains a particularly high concentration of nerve cells, and effectively functions as an independent 'minibrain'. Interactions between nerve, endocrine, immune and other cell types allow the sophisticated regulation of normal gut physiology. They can also bring about a co-ordinated response to parasitic infection, possibly leading to expulsion of the parasite. In this review, Derek McKay and Ian Fairweather will consider, in brief, data pertaining to changes in the ENS following intestinal helminth infections and speculate on the role that these alterations may have in the expulsion of the parasite burden and the putative ability of the parasite to modulate these events. PMID- 15275126 TI - Chondroitin sulphate A as an adherence receptor for Plasmodium falciparum infected erythrocytes. AB - Until recently, the sequestration of erythrocytes infected with Plasmodium falciparum has been thought to be due to one of a number of protein-protein interactions. In this article, Stephen Rogerson and Graham Brown summarize the emerging evidence that, in vitro, infected erythrocytes can also adhere to the glycosaminoglycan chondroitin sulphate A (CSA) expressed on the surface of cells and immobilized on plastic. In vivo, binding of infected erythrocytes to CSA could be crucial to the development of malarial infection of the placenta, and possibly to sequestration in the lung and brain. The consequences of this may include maternal morbidity and mortality, low birth weight in the infant, pulmonary oedema and cerebral malaria. They discuss the need to characterize the molecular basis of this interaction, and to investigate the possible therapeutic role of CSA in malaria. Chondroitin sulphates are nontoxic compounds already in use for other diseases in humans. Vaccines based on inhibiting this receptor ligand interaction could also be appropriate. PMID- 15275127 TI - Modulating ongoing Th2-cell responses in experimental leishmaniasis. AB - Dramatically polarized T helper (Th)-cell responses are seen in experimental murine infections with Leishmania major. Resistant mice develop a Th1-cell type response and heal the primary lesion, while susceptible mice develop non protective Th2-cell responses, and the disease eventually proves fatal. Deservedly, much effort has gone into determining factors that influence the development of these T-cell subsets early in infection; however, little is known about how the polarity of established responses can be permanently modified. In this article, Gary Nabors reviews his findings on modifying ongoing Th2-cell responses in susceptible mice, and discusses therapies that have proven effective involving reducing the level of infection using a conventional antileishmanial drug, combined with agents that push the response towards the Th1 pole. PMID- 15275128 TI - A simple method for quantifying Leishmania in tissues of infected animals. AB - In experimental animals infected with Leishmania major, the size of cutaneous lesions of the parasite often does not correlate with the number of parasites within the lesion. Indeed, cutaneous lesions can heal, but still contain parasites. Thus, the ability to determine parasite burden in infected animals becomes important, especially when assessing vaccines that are intended to induce sterilizing immunity. Here, Hermenio Lima, Julie Bleyenberg and Richard Titus describe a simple technique for enumerating Leishmania in infected tissue. It is hoped that this technique will allow all researchers working with Leishmania (especially those in countries where leishmaniasis is endemic) to determine parasite burden easily in infected animals. PMID- 15275129 TI - Super or not so super: Other considerations on T-cell immunity to Toxoplasma gondii. PMID- 15275131 TI - What are the microsporidia? PMID- 15275132 TI - The efficacy of antifolate antimalarial combinations in Africa: a predictive model based on pharmacodynamic and pharmacokinetic analyses. AB - At present, effective treatment for non-severe malaria is the most important malaria control strategy in Africa. Pyrimethamine-sulfadoxine (PSD) is rapidly becoming the first-line treatment in areas of chloroquine resistance, although the parasite chemoresistance factors that dispose towards clinical failure with PSD are still unclear. Here, Bill Watkins and colleagues analyse the relationship between the pharmacokinetic properties of two treatment combinations (PSD and chlorproguanil-dapsone) in vivo and the respective in vitro isobolograms for parasites with specific drug-resistance patterns. From this relationship, they develop a hypothesis that may explain clinical drug failure and differential efficacy between treatments. The deductions can be tested in field studies to validate or refute the model. PMID- 15275133 TI - The role of host-derived heat-shock protein in immunity against Toxoplasma gondii infection. AB - Heat-shock proteins (HSPs) are evolutionarily highly conserved polypeptides synthesized by cells to preserve cellular functions under a variety of stressful conditions, including infections. In infections, both host cells and pathogens express HSPs, although the role of these molecules in the host-pathogen relationship is elusive. Here, Hajime Hisaeda and Kunisuke Himeno show that a correlation exists between the 65 kDa HSP molecule (HSP65) and protection against Toxoplasma gondii infection, suggesting that this protein contributes to the host defense system. These findings may help in the understanding of the complicated host-parasite relationship. PMID- 15275134 TI - Studies with artificial extrachromosomal elements in trypanosomatids: could specificity in the initiation of DNA replication be linked to that in transcription? AB - Historically, artificial replicons have served as useful models for the definition of regulatory elements involved in chromosomal replication and transmission in yeast and DNA replication in bacteria. Here, Pradeep Patnaik examines what we have learnt so far from the replicative behaviour of various artificial extrachromosomal elements available for trypanosomatids. He highlights the involvement of transcription regulatory elements in virtually every eukaryotic origin of replication analysed in detail and, by drawing upon the extensive literature supporting a close association between DNA replication and transcription, he speculates that the nature and organization of origins of replication on a chromosome also may hold clues to the manner by which an organism regulates gene expression. PMID- 15275135 TI - Global mapping of lymphatic filariasis. AB - Disease maps are becoming increasingly important in infectious disease epidemiology and control. For lymphatic filariasis, the development of such maps has been hampered in the past by the lack of data on the geographical distribution of levels of infection or disease. Here, Edwin Michael and Don Bundy present an atlas for this parasitic disease derived from a recently compiled geographical database. Focusing on mapping and analysis of case prevalence data at the global and regional levels, the authors show how mapping the geographical distribution is integral not only to assessing spatial patterns in the infection and disease distribution but also to stratifying endemic areas by infection and/or disease rate. PMID- 15275136 TI - Antibody-mediated effects on parasite behavior: Evidence of a novel mechanism of immunity against a parasitic protist. AB - The parasitic ciliate Ichthyophthirius multifiliis is well known in commercial aquaculture as the etiological agent of 'white spot', a disease that afflicts a wide range of fresh-water fish. While Ichthyophthirius is highly pathogenic, animals exposed to controlled infections develop a strong acquired resistance to the parasite. Recent studies suggest host resistance involves a novel mechanism of humoral immunity affecting parasite behavior. Rather than being killed, parasites are forced to exit fish prematurely in response to antibody binding. The target antigens involved in this process are a class of highly abundant glycosylphosphatidyl-inositol-anchored coat proteins referred to as immobilization antigens, or i-antigens. Here, Theodore Clark and Harry Dickerson describe this phenomenon and offer a number of hypotheses that could account for the forced exit. PMID- 15275137 TI - Live attenuated vaccines against avian coccidiosis: Success with precocious and egg-adapted lines of Eimeria. AB - Avian coccidiosis, caused by parasites of the genus Eimeria, is a major disease of intensively reared poultry. Anticoccidial drugs, and vaccines based on live wild-type (virulent) parasites, have aided the management of coccidiosis, but the development of vaccines based on live attenuated parasites has been a significant advance. Here, Martin Shirley and Petr Bedrnik explain the basis of these recent vaccines and discuss their use. PMID- 15275138 TI - Microbial control of mosquitoes: management of the upper rhine mosquito population as a model programme. AB - For more than a decade, Bacillus thuringiensis israelensis (Bti) and B. sphaericus have been used successfully as biological control agents against mosquitoes in Germany. Over 1000 km(2) of breeding areas have been treated with Bti, resulting in a yearly reduction of the mosquito population by more than 90%, without evidence of any harmful impact on the environment. Here, Norbert Becker describes the control strategy and operational use of Bti and B. sphaericus in Germany. PMID- 15275139 TI - Is cryptosporidium clonal? PMID- 15275141 TI - Ross centenary: a perspective from a different angle. PMID- 15275140 TI - Nematodes in human muscle. PMID- 15275142 TI - A unified nomenclature for filarial genes. PMID- 15275143 TI - Onchocerciasis control: Moving towards the millennium. AB - The recognition of onchocerciasis as a major public health problem in the savanna belts of West Africa resulted in the establishment of the Onchocerciasis Control Programme (OCP) in 1974. Control was initially based on vector control by weekly larviciding. The OCP is now in transition towards its final phase in which repeated treatment with ivermectin, a safe and effective microfilaricide, is incorporated with vector control, or in certain circumstances is used alone. Ivermectin distribution hingeing on sustainable community systems is the basis of a new programme in endemic African countries outside the OCP and in the Americas. David Molyneux and John Davies describe the latest trends and developments related to onchocerciasis control. PMID- 15275144 TI - Twenty-two years of blackfly control in the onchocerciasis control programme in West Africa. AB - Twenty-two years after the launch of the Onchocerciasis Control Programme in West Africa (OCP), Jean-Marc Hougard and colleagues critically review the vector control strategy adopted. They go on to identify the few hydrological basins where transmission of the infection remains difficult to control, to analyse the causes and to propose appropriate corrective measures on a case-by-case basis. Most of these measures, which are mainly based on ivermectin chemotherapy, will continue to be applied after the end of the OCP in 2002, under the control of the countries concerned. PMID- 15275145 TI - Global eradication of Guinea worm. AB - Little more than a decade ago, it was estimated that over three million cases of dracunculiasis occurred worldwide. Since then, the numbers have fallen dramatically, thanks to the water supply initiatives of the 1980s and, more recently, the national guinea worm eradication programmes implemented in a score of endemic countries. Herve Peries and Sandy Cairncross discuss how eradication will require the containment of cases in the remaining endemic areas, together with the simultaneous strengthening of surveillance to permit the certification of eradication. This aim requires existing strategies to be adapted to maintain their efficacy and also to improve their sustainability and cost-effectiveness. Sudan with its civil war, and more than a hundred thousand reported cases, remains a major obstacle to rapid achievement of the goal. PMID- 15275146 TI - The global burden of intestinal nematode infections--fifty years on. AB - Fifty years after Stoll published his 'This Wormy World' article, the global prevalence of infections with intestinal nematodes remains virtually unchanged. The main species involved are Ascaris lumbricoides, Trichuris trichiura and hookworms, and there are now approximately one billion infections with each of these, worldwide. Given these large numbers, Man-Suen Chan here focuses on attempting to quantify the disease burden caused by these infections, using a recently formulated method of calculating disability-adjusted life years (DALYs). Using a mathematical model, it is estimated that approximately 70% of this burden can be prevented in high-prevalence communities by treating schoolchildren alone. Programmes targeted at schoolchildren have been shown to be extremely cost effective, and hence this provides a realistic approach for combating these infections in the future. PMID- 15275147 TI - Control of schistosomiasis--a global picture. AB - The control of schistosomiasis has been a challenging task for most endemic countries. Thus, despite the concerted efforts to date, schistosomiasis remains a major public health concern, second only to malaria in the tropics and subtropics. In this review, Lorenzo Savioli and colleagues highlight changes in schistosomiasis prevalence and distribution over the past decades, discuss the success and limitations of the various control strategies, and present possible control initiatives for the future. PMID- 15275148 TI - Mycoplasma contamination of Plasmodium cultures--a case of parasite parasitism. PMID- 15275149 TI - New tools for diagnosis and monitoring of bancroftian filariasis parasitism: the Polynesian experience. AB - Bancroftian filariasis is endemic in French Polynesia and control programs with diethylcarbamazine, started in the 1950s, led to a sharp reduction of the microfilaria prevalence. Consequently, the control program was interrupted in 1982. Ten years later, however, the incidence of the parasitism again reached pre control levels (20-30% microfilaremia in some islands), indicating that the adult worms (for which no diagnostic tool was available) had persisted. Apart from research on chemotherapy strategies, the Institut Malarde has been actively involved in developing and evaluating more-powerful diagnostic tools than the unique detection of microfilariae by blood smear examination. These include: (1) the detection of adult worm circulating antigens in humans, and (2) the detection of Wuchereria bancrofti larvae in mosquitoes, using DNA probes. In this paper, Luc Nicolas reviews the available diagnostic tools to detect W. bancrofti and their implementation in epidemiological areas, based on the Polynesian experience. PMID- 15275150 TI - Sex, strains and virulence. AB - When are populations of infectious agents likely to evolve into distinct strains? Are they likely to differ in their virulence? Will genetically distinct strains or clones remain stable long enough to be useful as epidemiological markers? Sexual recombination can break down the genetic associations that define a strain structure, but if sex is rare or inbreeding is common, can strains persist? In this paper Ian Hastings and Bruce Wedgwood-Oppenheim show how some simple population genetic theory may provide a basis for addressing these questions. PMID- 15275151 TI - Tick-host immunology: Significant advances and challenging opportunities. AB - Immunological interactions at the tick-host interface involve innate and acquired host defenses against infestation and immunomodulatory countermeasures by the tick. The cellular and molecular immunological bases of these host-parasite relationships are being defined. Acquired resistance to tick infestation involves humoral and cellular immunoregulatory and effector pathways. Ticks respond by suppressing antibody production, complement, and cytokine elaboration by both antigen-presenting cells and specific T-cell subsets. Tick-borne disease-causing agents probably exploit tick suppression of host defenses during transmission and initiation of infection. Because of the public health importance of ticks and Pick-borne diseases, it is crucial that we understand these interactions and exploit them in novel immunological control strategies. Here, Stephen Wikel and Douglas Bergman discuss recent advances in understanding tick-host immunology and propose future studies. PMID- 15275152 TI - A role for matrix metalloproteinases in the pathology and attenuation of Theileria annulata infections. AB - Upon infection with Theileria annulata, bovine leukocytes are induced to express eight novel metalloproteinase activities. In this article, Rachel Adamson and Roger Hall suggest that these enzymes are virulence factors and their presence may explain some of the features of the pathology of the disease. Specifically, they discuss the possibility that the metastatic properties of infected cells, the 'cigarette burn' ulcers and the cachexia characteristic of tropical theileriosis are associated with metalloproteinase expression. Furthermore, they propose that loss of metalloproteinase activity during the generation of a vaccine line could explain the attenuated phenotype. PMID- 15275153 TI - Speculation on possible life cycles for the clonal lineages in the genus toxoplasma. AB - Recent evidence suggests that the strains currently classified in the genus Toxoplasma, ie. within the species Toxoplasma gondii, may actually comprise at least two clonal lineages correlated with their virulence in mice. Here, Alan Johnson reviews these data in the context of evolution and speciation within the genus, and raises hypotheses on how the virulent lineage may undergo an asexual life cycle in nature, similar to that found for the very closely related coccidian, Neospora camnum. The putative vertical transmission life cycle of this mouse virulent lineage of T. gondii could involve passage to the foetus late in pregnancy, or transmission in milk to the neonate after birth. PMID- 15275154 TI - Satellite DNA sequences as taxonomic markers in nematodes of agronomic interest. AB - The success of alternative crop protection practices against plant-parasitic nematodes using host resistance genes depends fundamentally upon identification of the species and pathotypes effectively controlled by these genes. In the same way, biological control of insects by entomopathogenic nematodes will work only if the nematode strains used are indeed active against the pests to be eliminated. For these applications, the accurate interspecific and/or intraspecific identification of nematodes is thus of outstanding importance. Here, Eric Grenier, Philippe Castagnone-Sereno and Pierre Abad discuss the recent use of satellite DNA sequences in nematode taxonomic diagnostics. PMID- 15275155 TI - The ICT Filariasis Test: A rapid-format antigen test for diagnosis of bancroftian filariasis. AB - Antigen testing is now recognized as the method of choice for detection of Wuchereria bancrofti infections. Unlike tests that detect microfilariae, antigen tests can be performed with blood collected during the day or night. However, existing enzyme-linked immunosorbent assay (ELISA) tests for filarial antigenemia are difficult to perform in the field, and this has limited their use in endemic countries. In this article, Gary Weil, Patrick Lammie and Niggi Weiss review their experience with a new rapid-format filarial antigen test. They found that the ICT card test was very easy to perform and that it was comparable with ELISA for the detection of filarial antigen in sera from people with microfilaremia. The introduction now of an antigen test suitable for use in the field is especially timely, in that it may facilitate implementation of new strategies proposed by the World Health Organization for control and elimination of lymphatic filariasis. PMID- 15275156 TI - A simple method for counting cells in tissue sections. PMID- 15275157 TI - A Fibronectin-like molecule expressed by Eimeria tenella as a potential coccidial vaccine. PMID- 15275158 TI - A proposal for karyotype nomenclature in Trypanosoma brucei. PMID- 15275159 TI - Plasmodium cdc2-related kinases: Do they regulate stage differentiation? PMID- 15275160 TI - The pentose phosphate pathway and parasitic protozoa. AB - The pentose phosphate pathway plays a crucial role in the host-parasite relationship. It maintains a pool of NADPH, which serves to protect against oxidant stress and which generates carbohydrate intermediates used in nucleotide and other biosynthetic pathways. Deficiency in the first enzyme of the pathway, glucose-6-phosphate dehydrogenase, protects human erythrocytes from infection with Plasmodium falciparum for reasons that remain obscure. Loss of the third enzyme of the pathway, 6-phosphogluconate de-hydrogenase, is toxic, suggesting this enzyme might be a target for chemotherapy. Mike Barrett here summarizes the roles of the pentose phosphate pathway in various parasitic protozoa. PMID- 15275161 TI - The Trypanosoma cruzi genome initiative. AB - An initiative was launched in 1994 by the Special Programme for Research and Training in Tropical Diseases (TDR) of the WHO to analyse the genomes of the parasites Filaria, Schistosoma, Leishmania, Trypanosoma brucei and Trypanosoma cruzi. Five networks were established through wide publicity, holding meetings of key laboratories and developing proposals which were then reviewed by the Steering Committee of Strategic Research for financial support. The aim of the Programme was to use the platform of these networks to: (1) train scientists from tropical disease-endemic countries; (2) transfer technology and share material and expertise, thereby reducing costs and increasing efficiency; and (3) provide an information system that is accessible globally as soon as the results become available. The initial target was to produce a low-resolution genome map for each of the parasites, but it soon became evident that by using rapidly developing technologies, it might be feasible to complete DNA-sequence analysis for some of the parasites in the next decade, as discussed here by Alberto Carlos Frasch and colleagues, with particular focus on the T. cruzi genome initiative. PMID- 15275162 TI - Programmed cell death in trypanosomatids. AB - It has generally been assumed that apoptosis and other forms of programmed cell death evolved to regulate growth and development in multicellular organisms. However, recent work has shown that some parasitic protozoa have evolved a cell suicide pathway analogous to the process described as apoptosis in metazoa. In this review, Susan Welburn, Marcello Barcinski and Gwyn Williams discuss the possible implications of a cell suicide pathway in the vector-borne Trypanosomatids. PMID- 15275163 TI - Natural selection on polymorphic malaria antigens and the search for a vaccine. AB - Most protein antigens identified as malaria vaccine candidates are polymorphic in natural parasite populations. Current opinion is that a vaccine must be based on conserved regions of antigens, and if naturally acquired immune responses to these regions are only partially protective in humans, then the vaccine must create what is lacking in Nature. An alternative view is that a successful vaccine might need to be based on multiple allelic forms of an antigen. David Conway here shows that, far from being too pessimistic or impractical, this view offers positive ways to identify targets of protective immunity. PMID- 15275164 TI - Signal transduction mechanisms in Trypanosoma cruzi. AB - Trypanosoma cruzi, the etiological agent of Chagas disease, is an adequate model for studies on the evolution of signal transduction pathways. These pathways involve molecular entities such as membrane receptors, transduction G proteins, protein kinases and second messengers (Ca(2+), cyclic AMP, cyclic GMP, nitric oxide). In this article, Mirtha M. Flawia, Maria T. Tellez-Inon and Hector N. Torres describe the studies performed on T. cruzi transduction pathways and their role in the control of metacyclogenesis and cell motility. PMID- 15275165 TI - Generalized linear modelling for parasitologists. AB - Typically, the distribution of macroparasites over their host population is highly aggregated and empirically best described by the negative binomial distribution. For parasitologists, this poses a statistical provlem, which is often tackled by log-transforming the parasite data prior to analysis by parametric tests. Here, Ken Wilson and Bryan Grenfell show that this method is particularly prone to type I errors, and highlight a much more powerful and flexible alternative: generalized linear modelling. PMID- 15275167 TI - Schistosomiasis, fibrosis and esophageal varices. PMID- 15275168 TI - Scenario for survival. PMID- 15275169 TI - The importance of updating epidemiological data. PMID- 15275170 TI - Bovine Neospora and Neospora caninum: One and the same. PMID- 15275171 TI - Helminth immunogenetics: Why bother? AB - The importance of host genotype as a determinant of protective responses against helminth infection is well established. In contrast, there have been relatively few investigations of the role of helminth genotype, despite the importance accorded to the genetics of other disease-causing organisms. Here, Andrew Read and Mark Viney discuss the reasons for this oversight. They argue that it is not for any compelling empirical reason: there is at least as much evidence that worm genetics affects host protective responsiveness as there is that it does not. PMID- 15275172 TI - Nematode neuropeptides: Localization, isolation and functions. AB - Historically, peptidergic substances (in the form of neurosecretions) were linked to moulting in nematodes. More recently, there has been a renewal of interest in nematode neurobiology, initially triggered by studies demonstrating the localization of peptide immunoreactivities to the nervous system. Here, David Brownlee, Ian Fairweather, Lindy Holden-Dye and Robert Walker will review progress on the isolation of nematode neuropeptides and efforts to unravel their physiological actions and inactivation mechanisms. Future avenues for research are suggested and the potential exploitation of peptidergic pathways in future therapeutic strategies highlighted. PMID- 15275173 TI - The Pharmacology of Nematode FMRFamide-related Peptides. AB - FMRFamide-related peptides (FaRPs) are the largest known family of invertebrate neuropeptides. Immunocytochemical screens of nematode tissues using antisera raised to these peptides have localized extensive FaRP-immunostaining to their nervous systems. Although 21 FaRPs have been isolated and sequenced from extracts of free-living and parasitic nematodes, available evidence indicates that other FaRPs await discovery. While our knowledge of the pharmacology of these native nematode neuropeptides is extremely limited, reports on their physiological activity in nematodes are ever increasing. All the nematode FaRPs examined so far have been found to have potent and varied actions on nematode neuromuscular activity. It is only through the extensive pharmacological and physiological assessment of the tissue, cell and receptor interactions of these peptidic messengers that an understanding of their activity on nematode neuromusculature will be possible. In this review, Aaron Maule and colleagues examine the current understanding of the pharmacology of nematode FaRPs. PMID- 15275174 TI - Organelle DNAs: The bit players in malaria parasite DNA replication. AB - The replication mechanics of the extrachromosomal DNAs of the malaria parasite are beginning to be anravelled. At 6 kb, the mitochondrial genome is the smallest known and, unlike higher eukaryotes, its multiple copies per cell occur as polydisperse linear concatemers. Here, Don Williamson, Peter Preiser and Iain Wilson discuss recent evidence that this DNA replicates by a process akin to those of certain bacteriophages, which make use of extensive recombination coupled with rolling circles. The parasite's second extrachromosomal DNA, a 35 kb circular molecule thought to be a plastid remnant inherited from a remote photoautotroph, probably replicates in a more familiar fashion from conventional origins or D loops. Improved understanding of both organelle's replicative mechanisms could give new leads to malaria chemotherapy. PMID- 15275175 TI - A Toxoplasma gondii Superantigen: Biological effects and implications for the host-parasite interaction. AB - Superantigens exert their biological effects by activating large families of T cells, based on expression of the variable beta chain of the T-cell receptor. As a result, the reactive cells proliferate, secrete high levels of inflammatory cytokines, and ultimately die or become anergic to further stimulation. It is now becoming clear that the intracellular protozoan Toxoplasma gondii has many of these same superantigenic properties. As discussed here by Eric Denkers, this activity may play a key role in the induction of cell-mediated immunity to the parasite, and may prove to be responsible for much of the pathology associated with the clinical manifestations of toxoplasmosis. PMID- 15275176 TI - Parasite richness/sampling effort/host range: the fancy three-piece jigsaw puzzle. AB - In this article, Jean-Francois Guegan and Clive Kennedy propose an alternative explanation for the confounding effects of host geographical range and sampling effort on parasite species richness using pathway analysis procedure. They suggest that much of the species richness revealed by sampling effort is also a reflection of host range. Thus, the total contribution of host range logically incorporates a contribution from sampling effort. The implications of indirect effects of host range on richness estimates have not previously been discussed, and the authors here attempt to redress the balance. The contribution of host range to richness, as derived from control of sampling effort on richness estimates, therefore, is a mathematical expression that does not take into account the cause-and-effect nature of things. PMID- 15275177 TI - Identification and quantification of haemozoin: Some additional facts. PMID- 15275179 TI - Schools for health: Focus on health, education and the school-age child. AB - Mortality in children under five years old has been dramatically reduced through successful programmes of immunization and control of diarrhoeal diseases. UNICEF estimates that some 90% of children in developing regions now survive to reach school age. These survivors face new and continuing threats to their health, which can affect their physical development and may also prevent them taking full advantage of their only opportunity for formal education. The physical and mental growth of the 1000 million school-age children today will influence how the world is shaped for coming generations. Yet the health problems of this age group have received little attention. Recognizing the importance of this age group, a workshop funded by the Edna McConnell Clark Foundation was held 10-13 November 1994 in Fort Mitchell, Kentucky, USA, to review what is known about the health of school-age children, what is or can be done to improve their health, and what steps must be taken to find ways to improve the health and educational achievement of this important segment of the world's population. Don Bundy and Helen Guyatt here report on the workshop, which had three major conclusions: (1) the school-age children of developing countries face health problems that remain neglected and poorly understood; (2) an important research need is to develop simpler means of monitoring the health status of school-age children and evaluating the impact of public health interventions in this age group; and (3) two strategies are available to address this public health problem. The first is to develop further and test programmes that appear, from available evidence and pilot studies, to offer effective means of improving the health of this age group at reasonable cost, and to be sustainable; and the second is, over a longer term, to develop the capacity within countries to assess the health problems of school age children and devise cost-effective strategies to address these problems. This report attempts, in brief form, to survey what is known about the health status of school-age children, to discuss the possible benefits to health and learning that accrue from health interventions, and to suggest some avenues currently available for both research and application. PMID- 15275180 TI - Animal model for the pathogenesis of reactive amyloidosis. AB - The pathogenesis of amyloidosis is not well understood. Here, Zafer Ali-Khan, Weihua Li and Sic L. Chan present a metazoan parasite mouse model of reactive amyloidosis, review the relationship between chronic inflammation and multiorgan AA amyloidosis and postulate how ubiquitin might function in the processing of serum amyloid A and in AA amyloid formation in the endosomes-lysosomes of activated murine reticuloendothetial cells. PMID- 15275181 TI - The paraflagellar rod of kinetoplastida: solved and unsolved questions. AB - The flagellum of almost every member of the Kinetoplastida contains, next to its canonical 'nine-plus-two' axoneme, structure, a unique, complex and highly organized lattice-like structure called the paraflagellar rod or paraxial rod. Here, Philippe Bastin, Keith Matthews and Keith Gull summarize the latest findings on its structure, the nature of its protein components and their corresponding genes. They also consider the possible functions of this intriguing organelle. PMID- 15275182 TI - Rhoptry organelles of the apicomplexa: Their role in host cell invasion and intracellular survival. AB - Members of the phylum Apicomplexa are obligate intracellular parasites that invade erythrocytes, lymphocytes, macrophages or cells of the alimentary canal in various vertebrate species. Organelles within the apical complex of invasive stages facilitate host cell invasion. Parasites in this phylum cause some of the most debilitating diseases of medical and veterinary importance. These include malaria, toxoplasmosis, babesiosis, theileriosis (East Coast fever), and coccidiosis in poultry and livestock. In recent years, opportunistic infections caused by Cryptosporidium parvum, and recrudescent Toxoplasma gondii infections in AIDS patients have prompted intensified efforts in understanding the biology of these parasites. In this review, Tobili Sam-Yellowe examines the unifying and variant molecular features of rhoptry proteins, and addresses the role of multigene families in organelle function: the biogenesis of the rhoptries will also be examined, in an attempt to understand the sequence of events leading to successful packaging, modification and processing of proteins within the organelle. PMID- 15275183 TI - Schistosome resistance to praziquantel: Fact or artifact? AB - Praziquantel is the current drug of choice for human schistosomiasis. Recent reports from laboratory and field studies concerning reduced praziquantel efficacy against Schistosoma mansoni have generated some controversy. The prevailing question is whether the emergence of strains of schistosome resistant to praziquantel is a fact, or an artifact resulting from erroneous field or laboratory experimentation. In this article, Padraic Fallon, Liang-feng Tao, Magdi Ismail and James Bennett examine the available evidence for schistosome resistance to praziquantel. Contributory factors to the schistosomicidal activity of praziquantel, which may interfere with evaluation of drug efficacy or resistance, are also considered. PMID- 15275184 TI - Iscoms in parasitological research. AB - During the history of vaccine development, a number of adjuvants and adjuvant formulations have been tested and evaluated for their ability to increase the immunogenicity of different antigens. In this review, Anna Lunden, Karin Lovgren Bengtsson, Anders Sjolander and Arvid Uggla focus on iscoms (immune stimulating complexes), their characteristics and applications to different types of parasitic antigens. PMID- 15275185 TI - Marine parasites as biological tags of cephalopod hosts. AB - The use of marine parasites as non-intrusive natural tags of their hosts was first broadly applied in fisheries science in the 1940s. Both micro- and macroparasites have been used to assess the status of current stocks of several commercially exploited species of marine animals. Here, Santiago Pascual and Eric Hochberg offer a brief comment on marine parasite tags as a stock assessment methodology, with special reference to cephalopod hosts. PMID- 15275186 TI - Parasites and fire. PMID- 15275187 TI - Matchbox technology for irradiation hazards. PMID- 15275189 TI - Leishmania and the pathogenesis of HIV infection. AB - Acquired immunodeficiency syndrome (AIDS) and leishmaniasis overlap in several parts of the world, and microorganisms responsible for these human diseases infect and replicate within the macrophage. Therefore, the opportunity that the pathogenesis of the human immunodeficiency virus (HIV) and Leishmania infections could be modulated within dually infected individual is optimized. The most prominent clinical feature of AIDS is the development of debilitating secondary infections induced by several opportunistic microorganisms, including protozoa. Michel Tremblay, Martin Olivier and Richard Bernier here focus on the recently reported information on the putative cofactor role that the intracellular pathogen of the genus Leishmania may play in the pathogenesis of HIV infection. PMID- 15275190 TI - Experimental onchocercal keratitis. AB - In individuals with onchocerciasis, severe visual impairment (river blindness) occurs as a result of corneal inflammation induced by antigens released from dead and dying Onchocerca volvulus microfilariae. To characterize the underlying immune response, animal models have been developed that partially reproduce many of the clinical features of human onchocercal keratitis, Eric Pearlman here discusses how these studies have identified systemic and local immune responses associated with keratitis, including T helper-cell subset and cytokine responses. PMID- 15275191 TI - Lymphatic filariasis: what mice can tell us. AB - Human lymphatic filariasis is a major tropical disease in which clinical manifestations range from asymptomatic microfilaraemia to chronic pathology. Investigative immunological research into this disease is hampered by the fact that mice are refractory to the full developmental cycle of this parasitic nematode. However, studies using either single-stage infections or immunocompromised mice have greatly added to our knowledge of the filarial induced immune pathways leading to protective immunity, pathology and immunological tolerance. In this review, Rachel Lawrence discusses the recent advances in our understanding of the immunology of lymphatic filariasis using the mouse model. PMID- 15275192 TI - Artificial feeding of ixodid ticks. AB - Ixodid ticks are economically important as they cause direct damage to livestock and are vectors of several pathogens that cause diseases in humans and animals. Some of the important tick-borne pathogens of livestock are Theileria parva, T. annulata, Babesia bigemina, B. bovis, Anaplasma marginale and Cowdria ruminantium. These pathogens are responsible for causing enormous losses in livestock. Identification of factors that influence transmission and development of these pathogens in ticks will greatly facilitate development of rational strategies for control of tick-borne diseases. This research has been hampered by the lack of suitable artificial feeding methods. In this paper, Sam Waladde, Aian Young and Subhash Morzaria review recent developments in the artificial feeding of ixodid ticks and evaluate how this method can potentially be exploited. They use an example the transmission of an important livestock pathogen, T. parva, by Rhipicephalus appendiculatus. PMID- 15275193 TI - Genomic repetitive DNA elements of Trypanosoma cruzi. AB - Repetitive DNA sequences are interspersed throughout the genomes of mammals and other higher eukaryotes, and represent a substantial portion of the genome. Although it has been generally assumed that the redundant DNA is present only in the complex genomes of high order organisms, over the past few years a number of repetitive DNA sequences have been also detected in the protozoan parasite Trypanosoma cruzi. A compilation of the repetitive DNA sequences found in the T. cruzi genome is here presented by Jose Maria Requena, Manuel Carlos Lopez and Carlos Alonso, who also speculate on their possible origin and functional implications regarding retrotransposition and gene regulation. PMID- 15275194 TI - Host-like sequences in the schistosome genome. AB - A series of recent papers has indicated that widespread genomic rearrangements take place in the genome of schistosomes during the life cycle of the parasite. These results have been controversial since genomic rearrangements are not common in eukaryotes, probably because excessive genome plasticity would carry a heavy evolutionary price. Here, Karen Clough, Alec Drew and Paul Brindley present data that ostensibly support the concept of widespread genomic rearrangements, but for which they suggest a different interpretation. They conclude that artefactual contamination of schistosome genome preparations with host DNA can probably explain the Southern hybridization results which led to the original hypothesis of developmental, genomic rearrangements. PMID- 15275195 TI - Water-borne Cryptosporidium: a perspective from the USA. AB - In the USA, Cryptosporidium parvum has attracted considerable interest due to recent outbreaks of cryptosporidiosis attributed to water supplies contaminated with oocysts. Because of its unprecedented magnitude, the outbreak which occurred in Milwaukee in April 1993 was widely publicized and studied. Information collected during this and other recent outbreaks is being evaluated by regulatory agencies and the water industry with the aim of designing cost-effective measures that will reduce the risk of future water-borne outbreaks, as discussed here by Giovanni Widmer, Margaretha Carraway and Saul Tzipori. PMID- 15275196 TI - Malaria toxins: TNF-mediated phenomena. PMID- 15275198 TI - Protein motifs in filarial chitinases: An alternative view. PMID- 15275197 TI - More WWW addresses for parasitologists. PMID- 15275199 TI - Annexin V binding to mouse erythrocytes following infection with Plasmodium parasites. PMID- 15275200 TI - The epidemiology of morbidity of schistosomiasis. AB - The epidemiology of schistosomiasis is changing because treatment of chronically infected individuals is often followed by reinfection. As a major goal of schistosomiasis control is the reduction of morbidity, direct assessment of disease is essential because infection status is a relatively poor indication of morbidity. Introduction of ultrasonography to the study of schistosomiasis and the increased appreciation of the effects of schistosomiasis on growth and development in children have greatly enhanced our understanding of schistosome induced morbidity in endemic communities. Peter Wiest here reviews the changes in the assessment of schistosomiasis-induced morbidity. PMID- 15275201 TI - The pathogenesis of cryptosporidiosis. AB - Human infection with the protozoan parasite Cryptosporidium parvum has recently emerged as a global public health problem. Although infection is unrelenting in patients classically regarded as immunocompromised, a tantalizing observation is that infection with this parasite results in both acute self-limited as well as chronic diarrhea in young children. Recent data have begun to elucidate multiple potential mechanisms by which parasitism of the intestinal epithelium may yield an intestinal secretory response. However, a central issue for future studies is to understand how Cryptosporidium infection in young children results in such a broad spectrum of clinical presentation. An answer to this question is likely to result through a dual understanding of how systemic or enteric immunity impacts on intestinal secretory responses and how intra-cellular parasitism alters intestinal epithelial cell function and signals the submucosal intestinal compartment. The virulence factors of Cryptosporidium mediating these events need to be identified. Douglas Clark and Cynthia Sears here review the current understanding of the pathogenesis of intestinal secretion in response to Cryptosporidium infection, and discuss key questions requiring additional study. PMID- 15275202 TI - Topoisomerases in kinetoplastids. AB - Topoisomerases are enzymes that mediate topological changes in DNA that are essential for nucleic acid biosynthesis and for cell survival. The kinetoplastid protozoa, which include pathogenic trypanosomes and Leishmania, have yielded an interesting variety of purified topoisomerase activities as well as several topoisomerase genes. In these parasites, topoisomerases are involved in the metabolism of both nuclear and mitochondrial (kinetoplast) DNA. In this review, Christian Burri, Armette Bodley and Theresa Shapiro summarize what is known about topoisomerases in kinetoplastids, and consider the intriguing possibility that these enzymes may act as valuable antiparasite drug targets. PMID- 15275203 TI - Mating behaviour in Schistosomes: are paired worms always faithful? AB - Previously assumed to be monogamous, the mating system of schistosomes has been the subject of some debate since recent findings have shown that change of mate can occur among these parasites. Here, Louis-Albert Tchuem Tchuente, Vaughan R. Southgate, Claude Combes and Joseph Jourdane review progress made in the understanding of the mating behaviour of schistosomes, and highlight the importance of mating systems in the dynamics of natural transmission of schistosomiasis. PMID- 15275204 TI - Combating malaria morbidity and mortality by reducing transmission. AB - Jean-Francois Trape and Christophe Rogier present epidemiological data and an analysis of the relationship between transmission, morbidity and mortality from malaria which suggest that any intervention aiming to reduce transmission will not, on a long-term basis, reduce the burden of malaria in the majority of epidemiological contexts observed in tropical Africa. PMID- 15275205 TI - Control of lymphatic filariasis by annual single-dose diethylcarbamazine treatments. AB - It has long been stressed that diethylcarbamazine citrate must be given at a total dosage of 72 mg per kilogram of body weight in 12 divided doses of 6 mg kg( 1) to obtain maximum effect against Wuchereria bancrofti. However, recent studies revealed that only a single dose at 6 mg kg(-1) could reduce microfilaria (Mf) counts by 90%, and that the effect would persist for 12-18 months. The annual repeat of the single-dose mass treatment was shown to be effective in reducing Mf prevalence and density in large-scale, long-term field trials. The scheme is simple and economic, and could be sustainable in many endemic areas, where health manpower and resources are often not sufficient. Annual single-dose mass treatments can be an effective weapon against human lymphatic filariasis, as discussed here by Eisaku Kimura and Jona Mataika. PMID- 15275206 TI - In vitro systems in pneumocystis research. AB - Most groups involved in Pneumocystis research need large quantities of well preserved, viable Pneumocystis organisms free of host cell contamination. Biological, biochemical, immunological, genetic or other studies on Pneumocystis usually involve the separation of Pneumocystis from lung tissue as well as elimination of host cell debris from parasite extracts. In other investigations, such as transmission, infectivity, life cycle, biochemical, in vitro culture or drug-screening studies, viable and infectious Pneumocystis organisms are urgently required. However, there is no generally accepted methodology for obtaining Pneumocystis from experimental hosts or from human clinical samples; methods are still far from reaching standardization, as discussed here by the members of the European Concerted Action (ECA) on Pneumocystis carinii, which is co-ordinated by Eduardo Dei-Cas and Jean-Charles Cailliez. PMID- 15275207 TI - Eosinophil and mast cell staining in sheep infected by Oestrus ovis larvae. PMID- 15275208 TI - HDL particles as the trypanosome-killing factor in human serum: An exclusive or inconclusive role? PMID- 15275209 TI - Human immune response to MSP-1. PMID- 15275210 TI - Mosquitocidal toxins, genes and bacteria: the hit squad. AB - Certain entomopathogenic species of bacilli and Clostridium produce one or more toxins that kill mosquito larvae even at concentrations in the picomolar range. Altogether, 19 distinct genes are known that encode mosquitocidal toxins, which vary in their potency, species specificity and mode of action. Unlike chemical insecticides, mosquitocidal bacilli used as larvicides are safe for animals and the environment, and do not affect non-pest insects. Mosquitocidal bacteria are effective to varying degrees against Culex, Anopheles and Aedes mosquito larvae, but their rapid sedimentation from the larval feeding zone, UV-light sensitivity and narrow host range have hampered their development. New genetic engineering approaches are being investigated that could overcome these limitations and allow stable expression of broad host range combinations of toxins in UV-resistant, buoyant recombinant bacteria, as discussed here by Alan Porter. PMID- 15275211 TI - Malaria and onchocerciasis: On HLA and related matters. AB - In recent years, associations of particular factors of the human leukocyte antigen (HLA) system with two major infectious diseases of tropical countries have been recognized: common West African HLA antigens are associated with protection from severe Plasmodium falciparum malaria, and HLA-D alleles are associated with generalized disease, localized disease and putative immunity in Onchocerca volvulus infection. Here, Christian Meyer and Peter Kremsner summarize current information on the involvement of HLA factors in P. falciparum malaria and O. volvulus infection, and briefly report on HLA-related immunological characteristics of various conditions in these infectious diseases. PMID- 15275212 TI - Amphids in Strongyloides stercoralis and other parasitic nematodes. AB - In this review, Francis Ashton and Gerhard Schad examine the ultrastructure of the amphids of several animal parasitic nematodes. These structures are the main chemosensory organs of these worms and probably play an important role in host finding behavior and the control of development. Reconstructions made from serial micrographs of the neurons in the amphids of the threadworm Strongyloides stercoralis are shown. These stereo images permit three-dimensional visualization of these complex sense organs. The association between each amphidial neuron and its cell body has not been made previously for a parasitic nematode; however, this has been done for the free-living nematode Caenorhabditis elegans, which served as a model for these studies. Recognition of the cell bodies will provide a point of departure for laser microbeam ablation studies to determine individual neuronal function. PMID- 15275213 TI - Control of Babesia equi parasitemia. AB - Infection of horses with the hemoprotozoan Babesia equi has been reported in southern Florida, US Virgin Islands, part of Asia, Russia, India, the Middle East, Europe, Africa, Australia, South America, Central America, Mexico, Philippine Islands and some Caribbean islands. The restrictions placed on the international movement of infected horses has refocused attention on potential methods to control or eliminate infection. Don Knowles here discusses the primary chemotherapeutic compounds that have been used; the current knowledge concerning immune responses that potentially contribute to control of the parasite, and the development of infection of severe combined immuno-deficient foals as a model to dissect potential mechanisms of immunological control. PMID- 15275214 TI - Recent advances in DNA-mediated gene transfer of Entamoeba histolytica. AB - During the past few years, the introduction of DNA-mediated gene transfer into parasite research has permitted subtle studies on fundamental aspects of parasite biology. In this paper, Egbert Tannich describes the recent breakthrough of successful Entamoeba histolytica transfection, and the subsequent developments in this field. PMID- 15275215 TI - gammadelta T cells in malaria infections. AB - The association of a pronounced gammadelta T-cell response with Plasmodium infections is intriguing. The ability of parasite material to activate gammadelta T cells in vitro, and the localization of these cells in vivo in the red pulp of the spleen, suggests that these cells could play a role in the killing of bloodstage malaria parasites. However, the magnitude, the response and the predominance of inflammatory cytokines secreted by these cells may also indicate a role in the pathology of malaria infections. In this article, Jean Langhorne reveiws the current status of gammadelta T cells in malaria in the context of what is known about the function and specificity of gammadelta T cells in general. PMID- 15275216 TI - A mouse model for cerebral babesiosis. AB - Clinical symptoms and pathology observed in the cattle infected with Babesia bovis are quite similar to those of human cerebral malaria. Mechanisms involved in the pathogenesis of cerebral babesiosis, however, are still poorly understood because of the lack of a suitable experimental animal model. In this report, Masayoshi Tsuji and his colleagues describe B. bovis infection in severe combined immunodeficiency (SCID) mice, whose circulating red blood cells (RBCs) have been substituted with bovine RBCs (Bo-RBC-SCID mice). The infected mice not only develop a substantial level of parasitemia, but also show nerve symptoms and pathology similar to those observed in infected cattle. PMID- 15275218 TI - Low malaria mortality among children and high rates of Plasmodium falciparum inoculation: A Congolese reality in the 1980s. PMID- 15275219 TI - Nitric oxide in malaria: Indicator of disease severity and infection control. PMID- 15275220 TI - The pig as a unique host model for Schistosoma japonicum infection. PMID- 15275221 TI - The ATP-binding cassette (ABC) gene family of Plasmodium falciparum. AB - Multidrug resistance (MDR) in mammalian tumour cells is mediated by P glycoproteins. The apparent similarities between MDR and the chloroquine resistance phenotype (CQR) in Plasmodium falciparum suggests that homologous proteins may be involved. In mammals, P-glycoproteins are encoded by mdr genes that are a subset of a super-family characterized by ATP-binding cassettes (ABC). Three genes, pfmdr1, pfmdr2 and pfef3-rl, have been identified in P. falciparum that have homology to the ABC transporter gene family. Each protein encoded by these genes has a distinct structure, suggesting functional differences between the three. Justin Rubio and Alan Cowman here discuss the structure and possible function of the ABC proteins from P. falciparum and evidence that the protein encoded by the pfmdr1 gene can influence quinoline-containing antimalarial drug resistance phenotypes. PMID- 15275222 TI - The immune response to Giardia. AB - The flagellate Giardia duodenalis has been considered for many years to be a commensal living in the lumen of the small intestine of its host. It is only 25 years ago that it was accepted that Giardia is a significant pathogen of humans. Knowledge that Giardia can elicit an immune response that would probably contribute to the onset or absence of symptoms is not much older. The use of animal models to study the disease in the laboratory, together with the production of the whole life cycle in a test tube, have contributed greatly to our present knowledge of the immune responses to Giardia and of antigens that are specific to the trophozoite or cyst stages. In this review, Gaetan Faubert focuses on studies published since the last review in Parasitology Today in 1988, and examines the roles played by the humoral and cell-mediated immune responses in the control of the infection. It also covers the immunodiagnostic assays that have been recently developed on the basis of advances in our knowledge of the antigens of Giardia. PMID- 15275223 TI - Modulation of host cell intracellular Ca2+. AB - During the course of evolution, protozoan parasites have developed strategies to subvert the immune response of their host in order to multiply, reproduce and survive. One of these inherited strategies is their capacity to modulate the host cell transductional mechanisms in their favor. Alteration of host cells Ca(2-) homeostasis following interaction and/or invasion by protozoan parasites such as Leishmania donovani, Trypanosoma cruzi, Plasmodium falciparum or Entamoeba histolytica has been reported. There is direct evidence that such disturbances are responsible for pathogenesis observed during parasitic infections. This homeostatic imbalance of Ca(2+) in the host cell is an early inducible event whose underlying mechanisms needs further investigation, as discussed here by Martin Olivier. PMID- 15275224 TI - DNA replication in the malaria parasite. AB - Malaria is increasing as a global problem. Many of the drugs that were effective earlier in this century are now becoming obsolete as the parasite develops resistance to them and, despite earlier hopes, an affordable and effective vaccine remains elusive. It is hoped that a deeper understanding of the parasite's cell and molecular biology will give us a resource for the future and help us to achieve effective control. One aspect of parasite metabolism that has been the subject of recent studies is DNA replication: its timing during parasite development, the enzymes involved and the genes encoding them. In this review John White and Brian Kilbey report on the present status of these studies. PMID- 15275225 TI - Currently available molluscicides. AB - Views about the importance of the role of molluscicides in the integrated control of human schistosomiasis have passed through cyclical changes over the past 15 years. For a time, it was hoped that chemotherapy alone would achieve significant morbidity control; it has since become clear that molluscicides cannot be easily excluded from the anti-schistosome armoury. In this review, Sheena Perrett and Phil Whitfield summarize the evidence for this conclusion and provide an overview of currently available synthetic molluscicides and those natural product molluscicides currently under active investigation. PMID- 15275226 TI - Limburger cheese as an attractant for the malaria mosquito Anopheles gambiae s.s. AB - In the process of bloodfeeding, female Anopheles can transmit malaria parasites to humans. At night, while searching for blood, these insects respond to visual, physical and chemical properties of humans. Current research concentrates on the identification of kairomones, which guide mosquitoes to humans. Earlier observations on the biting behaviour of Anopheles gambiae s.s. on humans have now resulted in the discovery of a remarkable attractant for this important malaria vector, and it is thought that this will accelerate the development of odour baited traps for malaria mosquito surveillance and control in sub-Saharan Africa, as discussed here by Bart Knols and Ruurd De Jong. PMID- 15275227 TI - Haemozoin: identification and quantification. AB - Haemozoin (malaria pigment) is a haem polymer resulting from the breakdown of haemoglobin by Plasmodium spp. This refractory substance has been the focus of many studies and of much debate, mainly because of its role in the pharmacological activity of certain antimalarials. Haemozoin is also important because its presence in tissues serves as an indicator of malaria infections, and may itself be a mediator of malaria pathogenesis. In this article, Amy Sullivan and Steven Meshnick review the structure and synthesis of haemozoin, and then focus on methods of haemozoin identification in tissue. This latter aspect has implications for the study of haemozoin both as an indicator of malaria infection and as a possible mediator of malaria pathogenesis. PMID- 15275229 TI - Effectively controlling strongyloidiasis. PMID- 15275232 TI - Immunity to schistosomes in humans and other animals. PMID- 15275230 TI - Digestion of haemoglobin by schistosomes. PMID- 15275233 TI - Toxoplasma gondii: The best terrestrial biological weapon against extraterrestrial invaders? PMID- 15275234 TI - Paleopathology on schistosomiasis in Egyptian mummies. PMID- 15275235 TI - A test of the malaria strain theory. PMID- 15275236 TI - How do bednets influence the transmissibility of Plasmodium falciparum? PMID- 15275237 TI - FISH digital imaging microscopy in mosquito genomics. AB - The yellow fever mosquito, Aedes aegypti, transmits pathogens that affect both humans and livestock, and has been the focus of extensive research to identify genetic loci that may be useful in control strategies. Fluorescence in situ hybridization (FISH) and digital imaging microscopy have provided a rapid mechanism to populate the physical map with probes derived from genetic markers, cDNAs and recombinant genomic libraries. When the physical and genetic linkage maps are aligned, map-based cloning will allow the rapid isolation of target genomic sequences. The strategy of FISH mapping and the results of initial hybridization studies are reviewed here by Martin Ferguson, Susan Brown and Dennis Knudson. An Ae. aegypti-specific genomic database, which collates data from mapping studies, sequences, references and other relevant information, is also discussed. PMID- 15275238 TI - The evolution of parasitic diseases. AB - Parasites are characterized by their fitness-reducing effect on their hosts. Studying the evolution of parasitic diseases is an attempt to understand these negative effects as an adaptation of the parasite, the host, both or neither. Dieter Ebert and E. Allen Herre here discuss how the underlying concepts are general and are applicable for all types of disease-producing organisms, broadly defined here as parasites. The evolutionary processes that lead to the maintenance of the harmful effects are believed to be characterized by genetic correlations with other fitness components of the parasite. Depending on the shape of these correlations, any level of virulence can evolve. PMID- 15275239 TI - Glomerulopathy associated with parasitic infections. AB - Numerous infectious diseases, among them several parasitic infectious, have been shown to be associated with glomerular disease, although the exact pathogenetic mechanisms have not yet been elucidated. In this article, Marie-Louise van Velthuysen reviews the work published on glomerulopathy associated with the most important parasitic infections, ie. malaria, schistosomiasis, leishmaniasis and irypanosomiasis. PMID- 15275240 TI - Karyotype variability in Trypanosoma cruzi. AB - Like many other protozoam parasites, Trypanosoma cruzi (the causative agent of Chagas disease) has a plastic genome. Chromosome size polymorphisms occur in different strains of T. cruzi as well as among clones originating from the same strain, Despite this polymorphism, major interchromosomal rearrangements appear to be rare since several linkage groups of chromosomal markers are well conserved among different T. cruzi strains. In addition, some correlation has been found between karyotype variability and classification by multilocus enzyme electrophoresis. In this review, Jan Henriksson, Lena Aslund and Ulf Petterson discuss the genomic variability and suggest that amplication of repetitive sequences or members of gene families make a major contribution to the chromosomal size variation PMID- 15275241 TI - Worm burdens in schistosome infections. AB - Schistosomiasis, caused by fluke worms of Schistosoma spp, is one of the most common tropical diseases. Despite decades of research and progress towards the control of the disease, many aspects of the dynamics of infection and immunity remain unresolved. There is, in fact, not even an approximate measure of how many worms are harboured by infected humans. Epidemiological, mathematical and biomedical arguments indicate that individual worm burdens in endemic areas number hundreds to thousands of adult schistosomes, instead of the few to dozens generally assumed on the basis of available autopsy data. As Bruno Gryseels and Sake de Vlas here discuss, this hypothesis has important consequences for research and control, as many constants in schistosomiasis research have to be reconsidered. PMID- 15275242 TI - Schistosomes and serpins: a complex business. AB - The view of the schistosome host-parasitic relationship has changed in the past two decades. Previously, it was thought the parasite simply defended itself in the face of a hostile host environment. However, it is now realized that the host parasite interaction is much more of a dynamic interplay, where the parasite is able to exploit host homeostatic mechanisms for survival, maturity and transmission. Here, Jay Modha, Clare Roberts and John Kusel discuss the recent identification of serine protease inhibitors (serpins) on the schistosome surface and suggest how their properties might be exploited by the parasite. PMID- 15275243 TI - Phosphatidylserine expression on the surface of malaria-parasitized erythrocytes. PMID- 15275245 TI - Considering Taenia asiatica at species level. PMID- 15275247 TI - Rare codon usage in Escherichia coli and the expression of potentially toxic genes. PMID- 15275249 TI - Schistosome asparaginyl endopeptidase SM32 in hemoglobin digestion. PMID- 15275251 TI - The VSG-procyclin switch. PMID- 15275250 TI - Trade-offs in parasitology, evolution and behavior. PMID- 15275254 TI - The origins, dynamics and generation of Trypanosoma brucei rhodesiense epidemics in East Africa. AB - The history of sleeping sickness in East Africa has provoked controversy not only about the origins and spread of the disease, but also the identity of the causative organisms involved. Molecular methodology(1) has shed new light on the genetic makeup of the organisms involved in recent epidemics. Here, Geoff Hide, Andrew Tait, Ian Maudlin and Susan Welburn discuss these new data in relation to previous theories about the origins of epidemics in East Africa which emphasized the importance of the introduction of new strains. PMID- 15275255 TI - Lectin-parasite interactions. AB - Lectins are proteins that bind specifically to carbohydrate residues and are widely distributed in Nature. All parasites have such residues which vary in their configurations. Here, Jake Jacobson and Ron Doyle review the application of lectins in defining the developmental stages of parasites and the characterization, localization and structural composition of parasite glycoconjugates. The lectins of some parasites and lectin-mediated host-parasite interaction are also discussed. PMID- 15275256 TI - The role of Ca2+ in the process of cell invasion by intracellular parasites. AB - In order to replicate, many parasites must invade host cells. Changes in the intracellular Ca(2+) concentration ([Ca(2+)](i)) of different parasites and tissue culture cells during their interaction have been studied. An increase in cytosolic Ca(2+) in Trypanosoma cruzi trypomastigotes occurs after association of the parasites with host cells. Ca(2+) mobilization in the host cells also takes place upon contact with T. cruzi trypomastigotes, Leishmania donovani amastigotes or Plasmodium falciparum merozoites. When Ca(2+) transients are prevented by intracellular Ca(2+) chelators, a decrease in parasite association to host cells is observed. This reveals the importance of [Ca(2+)](i) in the process of parasite-host cell interaction, as discussed here by Roberto Docampo and Silvia Moreno. PMID- 15275257 TI - Evolutionary biology of parasitic platyhelminths: The role of molecular phylogenetics. AB - As our appreciation of the diversity within the flatworms has grown, so too has our curiosity about the ways in which these varied creatures are related to one another. In particular, the parasitic groups (trematodes, cestodes and monogeneans have been the focus of enquiry. Until recently, morphology, anatomy and life histories have provided the raw data for building hypotheses on relationships. Now, ultrastructural evidence, and most recently, molecular data from nucleic acid sequences, have been brought to bear on the topic. Here, David Blair, Andres Campos, Michael Cummings and Juan Pedro Laclette discuss the ways in which molecular data, in particular, are helping us recognize the various lineages of flatworms. PMID- 15275258 TI - Hookworms in the Americas: An alternative to trans-Pacific contact. AB - Recent New World archaeoparasitological discoveries of hookworm in pre-European contexts have revived a long-standing debate about the origin of hookworms in the Americas. Historically, the climatic conditions of Beringia encountered by migrating people have been considered too harsh for tropical hookworms to survive, suggesting that hookworms must have been introduced into South America by 'storm-tossed' fisherman or explorers from Asia. Here, John Hawdon and Susan Johnston review the history of hookworms in the Americas, and propose an alternative to their trans-Pacific introduction based on the unique natural history of one of the human hookworms. PMID- 15275259 TI - Transmission dynamics of Plasmodium falciparum. AB - Recent models of malaria have been developed by Gupta and her co-workers. A frequent assumption used to illustrate these models is that levels of malaria are controlled by lifelong strain-specific immunity. In this article, Allan Saul examines the predictions this model makes about the equilibrium values of parasite prevalence and the dynamics of an epidemic following the introduction of a new strain. He reaches the conclusion that the stability of malaria makes long term strain-specific immunity highly improbable, thus rendering models requiring lifelong strain-specific immunity unlikely to be of relevance in most epidemiological contexts. PMID- 15275260 TI - Second-generation antimalarial endoperoxides. AB - Artemisinin, derived from a Chinese herbal remedy, is a potent peroxide containing antimalarial. New types of peroxides, derived from this structure, as well as other naturally occurring antimalarial peroxides, have been synthesized and found to have potent antimalarial activities. Studies on the activities, modes of action, and toxicities of these compounds are discussed here by Steven Meshnick and colleagues. PMID- 15275264 TI - Sequestration or proliferation? PMID- 15275262 TI - The Plasmodium digestive vacuole and drug choice. PMID- 15275265 TI - Metabolic compartmentation in African trypanosomes. AB - Differences between host and parasite energy metabolism are eagerly sought after as potential targets for antiparasite chemotherapy. In Kinetoplastia, the first seven steps of glycolysis are compartmented inside glycosomes, organelles that are related to the peroxisomes of higher eukaryotes. This arrangement is unique in the living world. In this review, Christine Clayton and Paul Michels discuss the implications of this unusual metabolic compartmentation for the regulation of trypanosome energy metabolism, and describe how an adequate supply of energy is maintained in different species and life cycle stages. PMID- 15275266 TI - Co-feeding ticks: Epidemiological significance for tick-borne pathogen transmission. AB - Until recently, the transmission of tick-borne pathogens via vertebrates was thought to depend on the development of a systemic infection in the vertebrate hosts. Pathogen transmission has now been shown to occur between infected and uninfected ticks co-feeding in time or space in the absence of a systemic infection, originally for viruses, but now also for bacteria. The epidemiological consequences of this new non-systemic transmission pathway necessitate a major reassessment of the components and dynamics of tick-borne pathogen enzootic cycles. Here Sarah Randolph, Lise Gern and Pat Nuttall show that a much wider range of natural hosts than was previously recognized may contribute significantly to the transmission of tick-borne diseases, and compare quantitatively the relative contributions made by the systemic and non-systemic transmission pathways. PMID- 15275267 TI - Immunodeficient mice as hosts for hemoparasitic infections. AB - Thiruchandurai Rajan, Julie Moore and Leonard Shultz here review the evolution of technology in murine xeno-lymphohemopoietic chimeras, produced by engraftment with xenogeneic (fetal or adult) progenitor cells or mature lymphohemopoietic tissues into immunodeficient mice, and their use as hosts for hemoprotozoan parasites. Particular attention is paid to the development of chimeras that house xenogeneic peripheral red blood cells (xeno-RBC). These chimeras are potentially invaluable models for hemoprotozoan parasites, such as Babesia and Plasmodium. There are, however, daunting limitations that have to be overcome before these models can become universally acceptable systems for the study of these parasitic agents. PMID- 15275268 TI - Species of Echinococcus: pattern and process. AB - Although classification and nomenclature within the cestode genus Echinococcus has, historically, been controversial, the past 20-30 years have provided a period of relative stability. Recent calls for taxonomic revision in the genus have therefore created something of a stir. In this article, Alan Lymbery and Andrew Thompson describe the reasons for the new controversy, and suggest that the problem can only be resolved by agreement on an appropriate species concept and on operational procedures for implementing that concept. PMID- 15275269 TI - Selective metabolic labelling of intracellular parasite proteins using ricin. AB - The characterization of protein synthesis by eukaryotic intracellular parasites is inherently difficult because it may represent only a small fraction of host cell protein synthesis. Here, Mark Carrington, Yael Shochat and Anne Gurnett describe a method for overcoming this problem through the use of the toxin ricin to inhibit host cell protein synthesis specifically. PMID- 15275270 TI - De novo biosynthesis of heme in Plasmodium falciparum. PMID- 15275271 TI - Quantification of malaria transmissibility. PMID- 15275272 TI - Chronic chagasic tissue lesions in the absence of Trypanosoma cruzi: a proposed mechanism. PMID- 15275273 TI - In chronic Chagas heart disease, don't forget the parasite. PMID- 15275274 TI - Stage-specific gene expression in lymphatic filarial nematodes. AB - Lymphatic filarial nematodes remain a significant cause of morbidity throughout much of the tropics. One approach to the development of rational control methods is an improved understanding of the basic biology of these organisms in relation to the mechanisms used to complete their life cycles. In this article, Eileen Devaney, Sam Martin and Fiona Thompson review new approaches to defining stage specific molecules in filarial nematodes, and discuss their recent work on the isolation and characterization of stage-regulated cDNAs from Brugia pahangi. PMID- 15275275 TI - Nematode sperm. AB - Parasitic nematode infections remain a major public health problem in many parts of the world. Because most of the current strategies aimed at controlling parasitic nematode infections have met with only limited success, it may be time to consider alternative approaches. An aspect of nematode biology that has drawn little attention as a target for control is the reproductive process. Although there are numerous facets of the overall reproductive biology of nematodes that hold potential as targets for intervention, Alan Scott here focuses on the male reproductive system, and outlines some of the known unique processes and characteristics of sperm formation and sperm function that could be exploited to block fertilization. PMID- 15275276 TI - Anticoagulants in vector arthropods. AB - Arthropod-borne diseases cause significant morbidity and mortality worldwide. Mosquitoes alone may account for as many as three million deaths annually via the transmission of malaria. Because these diseases are transmitted to humans and to other vertebrates as a result of the ability of arthropods to feed on blood, the study of the biochemical mechanisms and adaptations that arthropods have evolved to facilitate hematophagy may provide insight into how this feeding behavior contributes to the transmission of disease. In this review, Kenneth Stark and Anthony James examine the diversity of arthropod anticoagulants and their role in hematophagy and potential implications for parasite transmission. PMID- 15275277 TI - The key components of resistance to Ostertagia circumcincta in lambs. AB - One of the most remarkable features of parasitic infections in general, and Ostertagia circumcincta infection of sheep in particular, is the extensive variation among hosts in resistance to infection, as assessed by parasite burdens and production of eggs or infective larvae. Here Mike Stear, Michael Park and Stephen Bishop describe the factors that account for the variation among animals within a flock, including dam, sire, sex, date of birth and history of exposure to infection. There are no detectable genetic effects in lambs less than three months old. Genetic control of an acquired response develops in two stages: first, a reduction in the average egg production per worm, which is associated with the development of a parasite-specific local IgA response; and second, control of worm burden, which is associated with the production of globule leukocytes in the abomasal mucosa. PMID- 15275278 TI - Malaria and the cell cycle. AB - The current model of cell cycle control features a succession of active cyclin CDK (cyclin-dependent kinase) complexes, where accumulation of each successive cyclin leads to activation of its associated kinase. Cell fusion experiments have shown that nuclei sharing common cytoplasm progress through the cell cycle in synchrony. During schizogony of Plasmodium falciparum, nuclear division occurs asynchronously, and thus cannot be regulated by synthesis and accumulation of cyclins in the cytoplasm. We suggest that schizonts must have a ready pool of cyclins for activating all stages of the cycle, and that the cell cycle is regulated independently in each nucleus. PMID- 15275279 TI - Exploitation of the complement system by Leishmania promastigotes. AB - Complement has long been acknowledged as an important component of host defense to extracellular pathogens. David Mosser and Andrew Brittingham here describe how Leishmania spp, intracellular pathogens of mononuclear phagocytes, exploit the opsonic and chemotactic effects of complement to initiate infection in the mammalian host. PMID- 15275280 TI - Severe adverse reaction risks during mass treatment with ivermectin in loiasis endemic areas. AB - The control of onchocerciasis remains a priority in sub-Saharan Africa. Jean Philippe Chippaux, Michel Boussinesq, Jacques Gardon, Nathalie Gardon-Wendel and Jean-Christophe Ernould here outline studies concerning possible severe adverse reactions to ivermectin used to control onchocerciasis in areas where loiasis is also endemic, and discuss precautions that may be advisable during the implementation of such programmes. PMID- 15275281 TI - Enhanced detection of Plasmodium vivax liver stages by cytocentrifugation. AB - Malaria parasites circulating in the blood or developing in the mosquito host can easily be seen and studied. We know much less about malaria parasites in the liver not only because of their location, but also because there are so few of them. Hepatic parasites are most often grown within hepatoma cells in culture, stained and visualized on slides under direct microscopy. In this report, Chitraporn Karnasuta and George Watt investigate the potential of cytocentrifugation as a tool for improving the detection of liver-stage malaria parasites. PMID- 15275282 TI - On hookworms in the Americas and trans-Pacific contact. PMID- 15275284 TI - The behavioral ecology of host-parasite interactions: An interdisciplinary challenge. PMID- 15275285 TI - Approaching helminth biology from the molecular direction. PMID- 15275287 TI - Models for malaria: Nature knows best. AB - Despite our increasing knowledge of the immunology of malaria in humans and in experimental systems, many questions remain unanswered. Given that this research has not yet led to a successful bloodstage vaccine, perhaps we should look again more carefully, in the light of new knowledge, at how many natural malaria infections are successfully controlled. Of these, the simian malarias are probably the most interesting, as described here by Geoff Butcher. PMID- 15275288 TI - Sex steroids, pregnancy-associated hormones and immunity to parasitic infection. AB - A wealth of evidence has accumulated that illustrates the ability of sex associated hormones to influence directly a variety of diverse immunological functions. Thus, it is not surprising that differences have also been noted between the sexes in their relative susceptibility to parasitic infections. Furthermore, during pregnancy, much of the observed maternal immunomodulation, essential for fetal survival, has been attributed to changes in the levels of steroid hormones. These pregnancy-induced alterations in immune function can also have profound effects on the course of parasitic infection. In this article, Craig Roberts, Abhay Satoskar and James Alexander review the immunological basis for differences in the relative susceptibilities of males, non-pregnant females and pregnant females to parasitic infection, particularly leislumaniasis and toxoplasmosis. They also discuss the role of the major sex- and pregnancy associated hormones in mediating these effects. PMID- 15275289 TI - Tick salivary prostaglandins: Presence, origin and significance. AB - Prostaglandins (PGs) are oxygenated metabolites of polyunsaturated fatty acids, most notably arachidonic acid, that act as 'local hormones', regulating a plethora of physiological processes in mammals and other vertebrates. For a long time, PGs were reported only in higher vertebrates, but more recently they have been reported in lower organisms such as bacteria, yeasts and protozoa, and much information is now available on PGs in insects. Prostaglandins are increasingly reported to exist at the host-parasite interface and are thought to aid the parasite by modulating the inflammatory and immune response. Ticks secrete saliva containing extremely high concentrations of PGs into the host, and in this article Alan Bowman, Jack Dillwith and John Sauer provide a synopsis of the information, to date, on the presence, synthesis and proposed roles for these tick salivary PGs. PMID- 15275290 TI - Autoimmunity in chagas disease cardiomyopathy: Fulfilling the criteria at last? AB - Here, Jorge Kalil and Edecio Cunha-Neto review the recent evidence for autoimmunity in chronic Chagas cardiomyopathy (CCC) involving molecularly defined antigens and immunopathological mechanisms. They also discuss the criteria for assignment of CCC as an organ-specific autoimmune disease. PMID- 15275291 TI - Strategies for the prevention of antimalarial drug resistance: rationale for combination chemotherapy for malaria. AB - Among the several 'tropical' diseases that affect humans, malarin poses special control problems due to the increasing population at risk from the disease, the difficulties in eradicating the mosquito vector in the tropics and the emergence and spread of parasite resistance to commonly used antimalarial drugs. There is both clinical experience and experimental evidence that, however effective when first introduced, the lifespan of drugs is inevitably curtailed by the emergence of resistant parasites. Resistance is the most important factor in determining the useful lifespan of antimalarial drugs. In this review, Nick White and Piero Olliaro discuss the rationale for combination chemotherapy. PMID- 15275292 TI - The epidemiology of heartwater: Establishment and maintenance of endemic stability. AB - Although heartwater (Cowdria ruminantium infection) is one of the most economically important tick-borne diseases of sub-Saharan Africa, its epidemiology he's remained poorly understood until recently. New data, suggesting that heartwater is present in an endemically stable state in much of sub-Saharan Africa and demonstrating vertical transmission of Cowdria ruminantium in the field, have altered previously accepted views on heartwater epidemiology. In this paper, Sharon Deem and colleagues present an overview of the epidemiology of heartwater based on recent studies, discuss the factors that make endemic stability possible, make recommendations for future directions in research, and provide a foundation for the development of epidemiological models. PMID- 15275293 TI - Differentiation of Entamoeba histolytica and Entamoeba dispar infections. AB - In this article, Terry jackson and Jonathan Ravdin briefly review the latest information on monoclonal antibody-based ELISAs that use antigen capture as a tool in the differential detection of human infection with Entamoeba histolytica and E. dispar. Current technology of culture and isoenzyme analysis is not widely available, is cumbersome and too time-consuming. A further potential benefit of antigen detection tests is that they can be used to monitor the efficacy of therapy; this is a shortcoming of serological tests owing to the persistence of the antibody response after successful treatment. PMID- 15275294 TI - Reliability of malariological measurements. PMID- 15275295 TI - Estimating the force of malaria infection. PMID- 15275297 TI - Diagnosis of Giardia infections. PMID- 15275299 TI - Viable Leishmania infantum in urine and semen in experimentally infected dogs. PMID- 15275300 TI - An immunological hypothesis to explain the enhanced susceptibility to malaria during pregnancy. PMID- 15275302 TI - Progress towards an amebiasis vaccine. AB - Amebiasis (infection by Entamoeba histolytica) remains a major health problem in much of the developing world. Morbidity and mortality from amebic dysentery and amebic liver abscess have persisted despite the availability of effective anti amebic therapy, suggesting a need for alternative measures of disease control. Through the application of recombinant DNA technology, several E. histolytica antigens have now been expressed in prokaryotic systems and tested in animal models as vaccines to prevent invasive amebiasis. In this review, Sam Stanley Jr discusses why a vaccine for amebiasis may be feasible, and describes the recent development of several promising recombinant E. histolytica antigen-based parenteral and oral vaccine candidates. PMID- 15275303 TI - Praziquantel: an urgent and exciting challenge. AB - The anthelmintic drug praziquantel has proved useful in the treatment of schistosomiasis. The precise mechanism by which praziquantel kills the parasites has yet to be elucidated. Here, John Kusel and colleagues review the current theories on praziquantel action and suggest future avenues for research, which becomes urgent in the light of some reports of drug resistance. PMID- 15275304 TI - Plasmodium vivax: Merozoites, invasion of reticulocytes and considerations for malaria vaccine development. AB - Several Plasmodium vivax merozoite proteins have been characterized over the past few years, including two that bind specifically to reticulocytes. Here, Mare Galinski and John Barnwell examine P. vivax merozoites and constituent molecules that are involved in host cell selection and invasion, and that also are viewed as malaria vaccine candidates. They also discuss how knowledge of the reticulocyte-binding proteins furthers the development of a conceptual framework for malaria merozoite invasion at the molecular level, not only for P. vivax, but for all species of the parasite. PMID- 15275305 TI - Developmental differentiation between tachyzoites and bradyzoites of Toxoplasma gondii. AB - An important event in the pathogenesis of toxoplasmosis is the interconversion between the bradyzoite and the tachyzoite stage of Toxoplasma gondii within the intermediate host. The factors that influence either cyst formation (bradyzoites) or reactivation (tachyzoites) are unknown. Uwe Gross, Wolfgang Bohne, Martine Soete and Jean Francois Dubremetz here describe current knowledge about the mechanisms that might lead to the induction of stage differentiation of this protozoan parasite. PMID- 15275306 TI - Spliced leader RNA trans-splicing in metazoa. AB - Spliced leader trans-splicing is a form of RNA processing originally described and studied in parasitic kinetoplastida. This mechanism of gene expression also occurs in parasitic and free-living metazoa. In this review, Dick Davis describes current knowledge of the distribution, substrates, specificity and functional significance of trans-splicing in metazoa. PMID- 15275307 TI - More on sleeping sickness in Uganda. PMID- 15275311 TI - Protein motifs in filarial chitinases. PMID- 15275309 TI - The 'Negro lethargy' in Uganda. PMID- 15275312 TI - The lipophosphoglycan-associated molecules of Leishmania. PMID- 15275313 TI - Protective immune mechanisms against the metacestode of Echinococcus multilocularis. AB - Infection with the larval stage of the fox tapeworm Echinococcus multilocularis results in a life-threatening hepatic disease concerning humans and intermediate rodent hosts. Immunoepidemiological surveys provided information that a large proportion of infected individuals may demonstrate either constitutional resistance to early post-oncospheral development of the parasite or late resistance to disease by exhibiting an intrahepatic died-out parasite lesion. Similar events have been found in secondary infections of laboratory rodents. Dissection of humoral and cell-mediated immune responses in susceptible versus resistant individuals provides insight into immunological pathways associated with the different outcome of infection. Survival strategy of the metacestode obviously focuses on the crucial role played by the parasite laminated layer. This layer protects the metacestode from host effector mechanisms which can potentially kill the proliferating germinative compartments in case of resistant hosts. Bruno Gottstein and Richard Felleisen here discuss the need to search for more parameters discriminating between the different immune pathways in order to find out (immunogenetic?) predispositions responsible for the respective phenomena. PMID- 15275314 TI - Eicosanoid production by parasites: from pathogenesis to immunomodulation? AB - In this review, Adam Belley and Kris Chadee discuss eicosanoid production by various parasites and propose roles they may play in pathogenesis and immunomodulation. The commonality between parasites is prostaglandin production and, therefore, special attention is given to the cyclooxygenase pathway, highlighting the enzymes and functions of prostaglandins. PMID- 15275315 TI - Regulation of immunity to malaria: valuable lessons learned from murine models. AB - A major advance in immunology has been the establishment of a framework for analysing how certain immune responses dominate following exposure to a particular pathogen or antigen. CD4(+) T helper (Th) cells can be separated into two major subsets which mediate qualitatively distinct cell-mediated (Th1) and humoral (Th2) immune responses. Immunity to most pathogens can be broadly categorized into a predominant protective response of either type. A characteristic of murine malarias is that primary infections with asexual erythrocytic parasites (the pathogenic stage of the malaria life cycle) generate a host protective immune response with a broad spectrum of Th1- and Th2-type CD4(+) T-cell involvement and so can be examined as models of the interaction of Th1 and Th2 cells during an immune response to an infectious agent. Andrew Taylor Robinson here describes recent events in the dissection of the mechanisms responsible for the generation of protective immunity to Plasmodium chabaudi chabaudi and other experimental malarias in mice. PMID- 15275316 TI - Evolution of parasite life history traits: myths and reality. AB - Parasitism has evolved independently several times in many different animal lineages. Observations made on distantly related parasites have revealed a variety of adaptations to parasitism, including changes in physiology, morphology, and life history traits. These observations have led parasitologists to formulate general rules about the evolution of parasites, rules that define a common evolutionary path presumably followed by all parasitic organisms. Robert Poulin uses recent evidence to question the generality of these rules and to show that parasite evolution may take different roads. The selective pressures acting on parasites are diverse and may guide their evolution in any direction, just as they have shaped a wide variety of free-living organisms. PMID- 15275317 TI - AGA/AGG codon usage in parasites: implications for gene expression in Escherichia coli. PMID- 15275318 TI - Naturally acquired immunity to Plasmodium falciparum. PMID- 15275319 TI - Are trees real? PMID- 15275320 TI - Trypanosoma brucei brucei and high-density lipoproteins: old and new thoughts on the identity and mechanism of the trypanocidal factor in human serum. AB - Nature has provided humans with a surprising means of protection against the African trypanosome Trypanosoma brucei brucei There is consensus, in that this singular trypanocidal factor is serum high-density lipoproteins (HDL). which the trypanosomes engulf through a physiological, receptor-mediated pathway for delivery to acidic intracellular vesicles. There is also controversy, however, in that the active particles and their essential cytotoxic elements are disputed, in part reflecting the ill-defined mechanism by which the parasites are finally killed. Here Patrick Lorenz, Bruno Betschart and Jim Owen discuss the possibilities for resolving these discrepancies and speculate on the prospects of exploiting this unexpected property of human HDL for protecting livestock. PMID- 15275321 TI - Oviposition pheromones in insect vectors. AB - Oviposition aggregation pheromones occur in a range of insect groups including Diptera, where they mediate oviposition in four different families of disease vectors. In this paper, Philip McCall and Mary Cameron discuss the selection pressures favouring oviposition pheromones and speculate on their potential applications in disease monitoring and control. PMID- 15275322 TI - Animal parasites, politics and agricultural research. PMID- 15275323 TI - The cysteine protease of Trypanosoma cruzi as a model for antiparasite drug design. AB - By combining new technology in molecular biology, X-ray crystallography, computer graphics and biochemistry, structure-based drug design provides a parallel and cost-effective strategy for identification of new antiparasite chemotherapy. James McKerrow, Mary McGrath and Juan Engel here discuss an example of the amplication of this strategy is its use in targeting the major cysteine protease in Trypanosoma cruzi. Tools from molecular biology helped overcome the obstacle of limited parasite material to allow production of reagent quantities of enzyme for inhibitor screening. Computer graphics analysis and X-ray crystallography are allowing rapid identification of new inhibitors based on either leads already identified or compounds selected by computer graphics screening of chemical databases. PMID- 15275324 TI - Cytoadhesion and falciparum malaria: going with the flow. AB - Sequestration of parasitized red blood cells in the cerebral vasculature is the predisposing event to the development of cerebral malaria during infection with Plasmodium falciparum. The adhesive interaction between these cells and receptors on the endothelial cell (cytoadhesion) occurs in the dynamic environment of the microcirculation, but most studies have neglected this factor and have concentrated on measuring adhesion in static (no flow) assays. Such studies ignore the markedly different rheological properties of parasitized red blood cells that become apparent when adhesion is examined under dynamic, flow conditions that resemble those of the circulation in vivo. Here, Brian Cooke and Ross Coppel review a number of novel aspects of cytoadhesion that have been identified using flow-based assays, and discuss their relevance to the pathophysiology, investigation and clinical management of falciparum malaria. PMID- 15275325 TI - The irradiated cercariae vaccine model: looking on the bright side of radiation. AB - Schistosomes infect between 200 and 300 million people at any one time. A major strategy to reduce the impact of schistosomiasis on human health is the development of a defined antigen vaccine. Protective immunity induced in mice by irradiated cercariae may serve as a model for the development of a vaccine. In such vaccinated mice, worm burdens resulting from challenge infection can be reduced by more than 90% compared to non-vaccinated mice. During the past three decades, the irradiated-carcariae vaccine model has been dissected in the detail in order to determine factors that may be relevant to vaccination, such as the participating immune compartments, the site and kinetics of the immune response, and the antigens recognized. In this review, Dania Richter, Donald A. Harn and Franz-Rainer Matuschka highlight the research on the vaccine model, focusing on the murine model using gamma-irradiated cercariae of Schistosoma mansoni. PMID- 15275326 TI - The plasmodium digestive vacuole: metabolic headquarters and choice drug target. AB - The Plasmodium digestive (food) vacuole is an acidic proteolytic compartment central to the metabolism of the parasite. Here haemoglobin is degraded, haem is polymerized, amino acid are transported, oxygen radicals are detoxified, drugs are accumulated, acidification is maintained and free iron may be generated. Despite these crucial roles in parasite development, a number of questions about the digestive vacuole and the haemoglobin ingestion pathway remain unanswered; in consequence, a number of attractive drug targets remain to be exploited. Piero Olliaro and Daniel Goldberg here review the morphology, metabolism and pharmacological disruption of this specialized organelle. PMID- 15275327 TI - Straw poll on nomenclature of parasitic diseases. PMID- 15275328 TI - Malaria: phosphatidylserine expression is not increased on the surface of parasitized erythrocytes. PMID- 15275329 TI - Digestion of haemoglobin by schistosomes: 35 years on. AB - Adult schistosomes obtain nutrients by digesting haemoglobin, which they obtain from ingested host red blood cells. Here John Dalton, Angela Smith, Karen Clough and Paul Brindley argue that a cathepsin L proteinase recently identified in their laboratory as the predominant cysteine proteinase activity of Schistosoma mansoni may play the leading role in haemoglobin digestion. They discuss the importance of elucidating the roles of both cathepsin B and cathepsin L in the digestion of haemoglobin, since both should be considered important targets to which novel schistosomicidal agents could be directed. PMID- 15275330 TI - The 1901 uganda sleeping sickness epidemic revisited: a case of mistaken identity? AB - The great sleeping sickness epidemic that occurred in Busoga at the turn of the century was caused by a trypanosome identified by Bruce as Trypanosoma gambiense. A study of trypanosomes from the recent epidemic in southeast Uganda has shed new light on the origins of the disease in Busoga. Thorsten Koerner, Peter de Raadt and Ian Maudlin suggest that the epidemic of the turn of the century was of T. p. rhodesiense sleeping sickness, brought about then, as now by social upheaval. PMID- 15275331 TI - Sampling effort and parasite species richness. AB - Comparative studies of parasite species richness among host taxa can be confounded by uneven sampling effort. Sampling ceases to be a confounding factor when extrapolation methods are used to estimate true species richness. Here, Bruno Walther and colleagues review examples of sampling bias and the use of extrapolation methods for circumventing it. They also discuss the confounding effects of phylogenetic association of estimates of species richness. PMID- 15275333 TI - Co-stimulatory activity of Leishmania-infected macrophages. PMID- 15275332 TI - FISH techniques for constructing physical maps on schistosome chromosomes. AB - In an effort to provide useful information about parasites important in tropical diseases, the WHO has initiated genome mapping projects for a number of parasites. One goal of this effort is to establish physical maps of the genomes of the targeted parasites. Multicellular parasites (helminths) contain various numbers of chromosomes, some large, that condense during the cell cycle. Here Hirohisa Hirai and Phil LoVerde present details of fluorescence in situ hybridization as a means to localize genes and DNA fragments to schistosome chromosomes. Although the techniques presented are for schistosome chromosomes, they are applicable to any system where the chromosomes condense at metaphase. PMID- 15275334 TI - Immunobiology of experimental cutaneous leishmaniasis. AB - The study of the murine model of infection with Leishmania major is providing important insights into the understanding of the complex interactions between the host and intracellular pathogens. Using this model system, basic research is actively leading to the identification of host factors promoting or circumventing the development of immunity to L. major. Here, Genevieve Milon, Giuseppe Del Giudice and Jacques A. Louis review recent results related to the characterization of immunological host factors determining resistance and susceptibility to this parasite, and try to identify areas where further research is required for a better understanding of the complex events triggered by intracellular parasites within their hosts. Extrapolation to the human leishmaniases of the rapid advances made in this murine model of infection, should pave the way to the rational design of future immunoprophylactic and immunotherapeutic measures. PMID- 15275335 TI - Mefloquine resistance in Plasmodium falciparum. AB - Mefloquine resistance in Plasmodium falciparum, the most dangerous of the four pathogenic malaria parasites of humans, is established in several endemic regions of the world. After a promising start, resistance has developed to disturbing extents in some areas, whereas in many regions it remains an effective drug. In this article, Frank Mockenhaupt reviews the factors that are likely to influence the development of mefloquine resistance, its possible mechanism and its geographical spread. PMID- 15275337 TI - Co-stimulatory activity of Leishmania-infected macrophages: reply. PMID- 15275338 TI - Transglutaminase activity in the microfilarial sheath. PMID- 15275340 TI - Leishmania, LPG and the Sandfly connection. PMID- 15275341 TI - Haemozoin and the mode of action of blood schizontocides: more controversy. PMID- 15275344 TI - Malarial toxins and the regulation of parasite density. AB - For over a century it has been recognized that many of the clinical symptoms of malaria are caused by toxins released by rupturing schizonts, but it is only in the past few years that the underlying mechanisms have begun to be understood. Dominic Kwiatkowski here focuses on the toxins that cause malaria fever by stimulating host cells to produce tumour necrosis factor a (TNF) and other pyrogenic cytokines. Both TNF and fever have antiparasite properties, and it is proposed that the release of these toxins plays an important role in the regulation of parasite density within the host. Cerebral malaria is related to excessive TNF production. Recent data indicate that this can be the consequence of genetic variation in the host's propensity to produce TNF. PMID- 15275345 TI - EBA-175: an erythrocyte-binding ligand of Plasmodium falciparum. PMID- 15275346 TI - Mechanisms of stage-regulated gene expression in kinetoplastida. AB - During their life cycle, trypanosomatid parasites of mammals encounter substantially different environments in their hosts and insect vectors, to which they must adapt by undergoing a series of differentiation processes. At the molecular level, these processes must be the direct result of an elaborate series of changes in stage-regulated expression of a wide range of gene products. How are these changes accomplished? In this review, Sheila Graham discusses some recent advances in understanding the mechanisms of gene expression in trypanosomatids, and examines some clues to some intriguingly complex means of regulating life cycle stage-specific gene expression. PMID- 15275347 TI - Wolbachia pipientis: symbiont or parasite? PMID- 15275348 TI - Human Plasmodium liver stages in SCID mice: a feasible model? AB - In a recent issue of Parasitology Today, Stanley and Virgin have stressed the potential of B- and T-cell deficient mice, among which severe combined immunodeficiency (SCID) mice are most frequently used, as models for the study of parasites. One of the most tantalizing prospects has been in the development of liver stages (LS) of human Plasmodium. PMID- 15275349 TI - A pocket guide to host-parasite models. AB - Mathematical models have been used to describe the population dynamics of a wide range of host-parasite interactions. Mick Roberts here discusses mathematical models for the dynamics of helminth endoparasites of non-human mammalian hosts, paying particular attention to the density-dependent factors that regulate the parasite populations, and the interaction between parasite and wild or feral animal host populations. PMID- 15275350 TI - Malaria during pregnancy: a priority area of malaria research and control. AB - More than 2000 million people live in areas where malaria transmission occurs and are therefore at risk of being infected. It follows that 1000 million people are exposed to the risks of malaria when pregnant. Although the special features of malaria during pregnancy have been recognized for nearly a century(1), it is only recently that it is being considered as a priority for malaria research and control, as discussed here by Clara Menendez. PMID- 15275352 TI - Clinical disease and pathogenesis in malaria. AB - This is the report of a meeting held in Ahungalla, Sri Lanka, 16-19 January 1994, under the sponsorship of the Rockefeller Foundation, Health Sciences Division. The meeting was initiated jointly by the Rockefeller Foundation and the TDR Special Programme of the World Health Organization in order to bring together scientists with a wide spectrum of experience relating to malarial disease and pathogenesis. The objective was to generate interdisciplinary discussion ranging from the clinical pictures of malarial infections and their impact in different parts of the world, to current investigations on mechanisms of pathogenesis and clinical immunity and the genetic determinants in human and parasite populations affecting the nature of the disease. PMID- 15275353 TI - Is the broad range of hosts and geographical distribution of Trypanosoma evansi attributable to the loss of maxicircle kinetoplast DNA? PMID- 15275356 TI - The cytoplasmic ribosomal RNAs of Plasmodium spp. AB - Plasmodium spp maintain several structurally distinct sets of ribosomal RNA genes whose expression is developmentally regulated. This feature sets them apart from all other eukaryotes studied to date. In this review, Thomas McCutchan, Jun Li, Glenn McConkey, John Rogers and Andy Waters give an account of the progress in our understanding of this unusual phenomenon as it relates to the biology of the parasite. They also outline an interesting turnabout in scientific direction involving the use of the parasite as an important new model for the study of the eukaryotic ribosome. PMID- 15275357 TI - Plasmodium berghei: the application of cultivation and purification techniques to molecular studies of malaria parasites. AB - Species of malaria parasites that infect rodents provide models for the study of the biology of malaria parasites that infect humans. In this article, Chris Janse and Andy Waters describe some of the recent advances in the cultivation and purification methodology of one of these species, Plasmodium berghei. The improvement of these techniques, and the increasing knowledge about the molecular biology of P. berghei enhance the value of this particular rodent model for the investigation of many aspects of the biology of Plasmodium. PMID- 15275359 TI - How ticks make a living. PMID- 15275358 TI - Immunological strategies for control of insect disease vectors: a critical assessment. AB - Insect-borne diseases are one of the major causes of disease and death in the tropical world. Conventional methods of disease control have proven insufficient and there is pressing need for devising new strategies. One approach that has been explored by several laboratories is to compromise vector fecundity and survivorship through the immunization of vertebrate hosts with the vector's internal organs (concealed antigens). Here, Marcelo Jacobs-Lorena and Francisco J.A. Lemos critically review the results obtained to date by use of this approach. It appears that the published work is less rigorous than would be desirable and the results are contradictory. In contrast to the successes obtained with a similar immunization strategy implemented for tick-borne diseases, it is not yet clear whether or not such an approach can be applied to insect-borne diseases. PMID- 15275360 TI - Influence of maternal infection on offspring resistance towards parasites. AB - Immunoglobulins, parasite circulating antigens, immune cells, cytokines and other cell-related products can be transferred from infected mothers to their young. They can combine their effects to interact with the invading parasites, as well as to induce a long-term modulation of the offspring's capacity to mount an immune response to subsequent exposure to parasites. The protective effect of maternally derived antibodies may be limited by the selective transfer of immunoglobulin isotypes. Maternal antibodies may also prevent the priming of specific cells in offspring or inhibit the progeny's antibody production by interacting with B-cell receptors or with the idiotypic repertoire. The potentially beneficial priming effect of transferred parasitic antigens may be altered by the Th2-cell-biased foetal environment and such antigens may also induce deletion or anergy of T- and B-cell clones in offspring. Therefore, besides protective effects, maternal infection may downregulate the offspring's immune response. If such hyporesponsiveness may be clearly harmful (in increasing the risk or in worsening congenital or postnatally acquired infections in offspring), it can also be beneficial (in limiting the pathogenesis of some infections). Here, Yves Carlier and Carine Truyens review the rationale of these complex foeto-maternal relationships in parasitic diseases. PMID- 15275361 TI - Immune evasion by Babesia bovis and Plasmodium falciparum: cliff-dwellers of the parasite world. AB - Erythrocyte-dwelling parasites, such as Babesia bovis and Plasmodium falciparum, are not accessible to the host immune system during most of their asexual reproductive cycle because they are intracellular. While intracellular, the host immune response must be directed toward the surface of the infected erythrocyte. Immune individuals mount protective antibody and cell-mediated responses which eliminate most of the parasites, yet some survive to establish chronic infections. In this review, David Allred discusses some of the mechanisms used by these parasites to evade individual immune mechanisms targeting the infected erythrocyte to survive in the hostile environment of an effective immune response. PMID- 15275362 TI - Host age as a determinant of naturally acquired immunity to Plasmodium falciparum. AB - The usual course of infection by Plasmodium falciparum among adults who lack a history of exposure to endemic malaria is fulminant. The infection in adults living with hyper- to holoendemic malaria is chronic and benign. Naturally acquired immunity to falciparum malaria is the basis of this difference. Confusion surrounds an essential question regarding this process: What is its rate of onset? Opinions vary because of disagreement over the relationships between exposure to infection, antigenic polymorphism and naturally acquired immunity. In this review, Kevin Baird discusses these relationships against a backdrop of host age as a determinant of naturally acquired immunity to falciparum malaria. PMID- 15275363 TI - What makes a malaria host? PMID- 15275366 TI - Super-multiple describership. PMID- 15275364 TI - Offensive secretory SODs? PMID- 15275367 TI - Nucleic acids: vaccines of the future. AB - The recent successful immunization of experimental animals using nucleic acids has provided a revolutionary new approach in vaccinology. In this article, Gary Waine and Don McManus examine the potential of nucleic acid vaccines for their effectiveness not only against infectious and parasitic organisms exhibiting an intracellular phase during their life cycle, but also against parasitic helminths, whose life cycle stages are either predominantly or completely extracellular. PMID- 15275370 TI - Convergent evolution of tropomyosin epitopes. PMID- 15275371 TI - Chitinase as a vaccine. PMID- 15275372 TI - Parasite subversion of the host cell endocytic network. PMID- 15275373 TI - T-cell activation and the balance of antibody isotypes in human lymphatic filariasis. AB - Human filarial infection presents a spectrum of clinical states with two major poles: asymptomatic microfilaraemia and amicrofilaraemic chronic disease. Microfilaremia is associated with a Th1-type tolerance, and maximal IgG4 antibodies, while elephantiasis patients react across a broad range of immune parameters. In this review, Rick Maizels and his colleagues discuss recent advances in the immunology of human filariasis and present a summary of their latest studies in an endemic area of Indonesia. PMID- 15275374 TI - Worm control and anthelmintic resistance: adventures with a model. AB - There are three common questions asked of parasitologists about anthelmintic resistance. Does it matter? How do you prevent it? Can you help me (it's here!)? In short, the respective answers are yes, read on the read on. Elizabeth Barnes, Robert Dobson and Ian Barger examine these issues in the context of nematode parasite control in grazing sheep. With the aid of a model, they examine some important factors that influence drug resistance and how farm management decisions and worm genetics modify these factors. They also explore the likely impact of new technologies on drug resistance and how efficient they need to be to sustain good worm control. PMID- 15275375 TI - Mechanisms of resistance to Onchocerca infection in blackflies. AB - In this article Peter Ham, Hans Hagen, Andrea Baxter and Jorg Grunewald focus on the susceptibility of blackflies to parasitic filarial infection (particularly Onchocera spp, most of the vectors of which belong to the genus Simulium). They outline what is known about, as well as speculating on, the various defence mechanisms of these insects. Investigations have involved the use of natural and surrogate vectors of bovine onchocerciasis as models for the human vector parasite relationship. PMID- 15275376 TI - Rejoinder: the use of PCR in the field. PMID- 15275377 TI - Parasites in Vietnam. PMID- 15275378 TI - Parasitophorous duct? Still more questions than answers. PMID- 15275381 TI - Trypanosomatid shuttle vectors: new tools for the functional dissection of parasite genomes. AB - In the past five years, gene-transfer systems have been established for each of the medically important trypanosomatids: Leishmania sp, Trypanosoma brucei and T. cruzi. Transformation can be mediated by integration, which occurs exclusively by homologous recombination, or by episomal shuttle vectors. In this article, John Kelly will focus on recent progress in the development and applications of trypanosomatid shuttle vectors, ie. vectors which are maintained extrachromosomally and which are capable of autonomous replication in both trypanosomatid and bacterial hosts. PMID- 15275382 TI - Oesophagostomum infections in humans. AB - Oesophagostomum spp are normally found as nematode parasites of ruminants, pig and monkeys. Occasionally humans are involved. In the past decade it became clear that, in some parts of Africa, humans are adequate final hosts. In those areas, prevalences of infection are high and morbidity is significant. The presence of lumen-dwelling adult worms, which do not seem to cause a great deal of pathology, can be demonstrated through coproculture. The presence of immature worms, encapsulated in nodules and responsible for pathology, on the other hand, is more difficult to confirm. It is not known what factors limit the distribution of endemic human oesophagostomiasis to a small focus in West Africa. The relationship between the 'helminthomas' described a long time ago in Uganda and the human Oesophagostomum infections in West Africa is unclear and it remains a mystery how humans get infected so effectively by ingesting L3 larvae. In this overview, Ton Polderman and Coby Blotkamp give an account of what is known and what is still to be elucidated in human Oesophagostomum infections. PMID- 15275383 TI - Vaccines against babesiosis using soluble parasite antigens. AB - Babesiosis in cattle and dogs causes severe economical and emotional loss. Although effective chemotherapeutic treatment of infected animals is available, the prevention of babesiosis by vaccination would be preferable. Attenuated parasite lines of Babesia bovis have been used successfully to control tropical babesiosis in cattle. However, among other drawbacks associated with live vaccines, such vaccines bear the risk of variable infectivity and morbidity requiring veterinary surveillance. Soluble parasite antigens derived from different Babesia species have proven to induce immune responses that do not necessarily affect the parasite, per se, but reduce the manifestations of clinical disease upon infection. In this review, Theo Schetters and Sonia Montenegro-James present an overview of the results obtained with vaccines based on soluble parasite antigens and their characterization, and discuss the possible immune effector mechanisms of such vaccines. PMID- 15275386 TI - Murine models of cerebral malaria: a qualified defence. PMID- 15275384 TI - Malaria toxins: hypoglycaemia and TNF production are induced by different components. PMID- 15275387 TI - Enhanced production of reactive nitrogen intermediates in human and murine malaria. PMID- 15275388 TI - Enhanced production of reactive nitrogen intermediates in human and murine malaria: reply. PMID- 15275389 TI - Will reducing Plasmodium falciparum malaria transmission alter malaria mortality among African children? PMID- 15275390 TI - Is reduction of transmission desirable for malaria control? PMID- 15275392 TI - Mast cell and eosinophil staining. PMID- 15275393 TI - Parthenogenesis in schistosomatidae. AB - Among which species, in what situations and how, does parthenogenesis occur in the biology of reproduction of schistosomes? Here, Jose Jourdane, Daniele Imbert Establet and Louis Albert Tchuem Tchuente review the literature on parthenogenesis in schistosomes, and debate the evolutionary aspects of this type of reproduction. PMID- 15275394 TI - Echinococcus multilocularis in germany: increased awareness or spreading of a parasite? AB - The 'small fox tapeworm' Echinococcus multilocularis has recently become a matter of intense interest in Germany. A long-term increase of its prevalence in foxes has been noted in the well-known endemic areas in Southern Germany, and reports on the occurrence of the parasite in other parts of the country suggest that the parasite is actually much more widespread than previously thought. As nearly all of the relevant studies are published in the German language in a veterinary journal and in the hunting press, accessibility to the information is limited. Richard Lucius and Brigit Bilger here describe the situation, and discuss the possible reasons and consequences. PMID- 15275395 TI - Cryptosporidium and cryptosporidiosis. AB - The tiny, iodine- and chlorine-resistant protozoan oocysts of Cryptosporidium parvum, long recognized by veterinarians, have become increasingly noted as a cause of watery diarrhea in developed and developing countries throughout the world. For immunocompromised patients, particularly those with AIDS, this diarrhea can be severe and life-threatening. Clovis Martins and Richard Guerrant here discuss the increasing recognition of this important pathogen in immunocompetent patients as well, and outline new challenges to improved water treatment, immunologic and antiparasite chemotherapy. PMID- 15275396 TI - Home improvements: malaria and the red blood cell. AB - In real-estate agent's terms, the red blood cell is a renovator's dream. The mature human erythrocyte has no internal organelles, no protein synthesis machinery and no infrastructure for protein trafficking. The malaria parasite invades this empty shell and effectively converts the erythrocyte back into a fully functional eukaryotic cell. In this article, Michael Foley and Leann Tilley examine the Plasmodium falciparum proteins that interact with the membrane skeleton at different stages of the infection and speculate on the roles of these proteins in the remodelling process. PMID- 15275397 TI - Human immunity to schistosomes. PMID- 15275398 TI - The ins and outs of editing RNA in kinetoplastids. AB - Over 30 million people in tropical regions suffer from Chagas disease, African sleeping sickness or leishmaniasis. The causative agents of these diseases, flagellated protozoa collectively known as kinetoplastids, represent an ancient lineage of eukaryotes. These unusual organisms carry out a large number of unique biochemical processes, one striking example being the sequence editing of mitochondrial messenger RNAs. In this review, Scott Seiwert focuses on recent studies that examine the reaction mechanism, molecular machinery and evolutionary history of this unusual RNA processing reaction. PMID- 15275399 TI - Polydnaviruses: potent mediators of host insect immune dysfunction. AB - Endoparasitic insects are used as biological control agents to kill many species of insect pest. One key to the success of parasitoids that develop in the hemocoel of their host is their ability to knock out the host's immune system, inducing a decline in the responsiveness of a variety of cellular and humoral components so that parasitoid eggs are not encapsulated. In many species parasitized by braconid and ichneumonid wasps, host immunosuppression appears to be mediated by polydnaviruses (PDVs) injected by the female parasitoid into the host hemocoel. The viruses exhibit a complex and intimate genetic relationship with the wasp, since viral sequences are integrated within the wasp's chromosomal DNA. Here Mark Lavine and Nancy Beckage summarize the current evidence for mechanisms of virally induced host immunosuppression in parasitized insects, as well as the roles of other factors including wasp ovarian proteins and venom components, in suppressing hemocyte-mediated and humoral immune responses. Interestingly, in some species, the PDV-induced host immunosuppression appears transitory, with older parasitoid larvae probably exploiting other mechanisms to protect themselves from the host's immune system during the final stages of parasitism. During the final stages of parasitism, the parasitoids likely exploit other mechanisms of immunoevasion via antigen masking, antigen mimicry, or production of active inhibitors of the hemocyte-mediated encapsulation response as well as inhibiting melanization. PMID- 15275400 TI - The sticky secrets of sequestration. AB - Sequestration, the adherence of infected erythrocytes containing the more mature stages of parasite development (trophozoites and schizonts) to the endothelial cells lining the capillaries and post-capillary venules, is characteristic of Plasmodium falciparum infections. In this review, Irwin Sherman and his colleagues discuss recent advances in the characterization of the adhesive molecules on the surface of malaria-infected erythrocytes and the receptors on the endothelium to which they bind. PMID- 15275401 TI - Parasite cyclophilins and antiparasite activity of cyclosporin A. AB - Cyclosporin A (CsA) was initially developed as an immunosuppressive drug. In the past several years, it has been shown to possess antiparasite activity independent of the immune system. It is not known how the drug exerts these antiparasite effects, or why it is stage and/or species specific. The answers may lie in the enzymatic function of cyclophilins. The cyclophilins are a growing family of proteins that exhibit peptidyl-prolyl cis-trans isomerase (PPiase) activity and bid CsA to varying degrees. PPiases have been shown to play a role in the folding of many essential proteins. Antony Page, Sanjay Kumar and Clotilde Carlow here review parasite cyclophilins and their association with CsA. The possible biological function of parasite cyclophilins and their potential role in future drug discovery are also discussed. PMID- 15275402 TI - Human immunity to schistosomes: reply. PMID- 15275403 TI - Cysticercosis in Africa. PMID- 15275405 TI - Programmed T-cell death in experimental chagas disease. AB - In mature T cells, programmed cell death is thought to serve a regulatory function by limiting both the duration and amplitude of immune responses. Programmed cell death might also be involved in immuno-pathogenesis of certain infectious diseases: recent evidence suggests that programmed T-cell death plays an important role in immune suppression during viral infections. In this article, George DosReis, Maria Evangelina Fonseca and Marcela Lopes review their findings on programmed T-cell death in experimental infection induced by the protozoan parasite Trypanosoma cruzi, the causative agent of Chagas disease. They also discuss the differential behavior of CD4(+) and CD8(+) T-cell subsets regarding programmed cell death, and same possible pathogenic aspects of host-parasite interaction, where abnormal or exaggerated programmed T-cell death could be involved. PMID- 15275406 TI - Leishmania major infection: the overture. AB - Local infection of mice with Leishmania major results in either healing or death depending on the preferential action of Th1 or Th2 T helper cells, respectively. Although the parasite-induced T-cell responses and their consequences for the disease are well understood, relatively little is known about the initial events that kindle the adaptive immune response. Werner Salbach and Tamas Laskay here discuss how differences in parasites spreading from the site of infection to different immune organs during the first 10-24 hours and, in consequence, the 'where and when' of the first encounter of Leishmania with the cells of the immune system may well be the starting point for the development of resistance or susceptibility. PMID- 15275407 TI - What are T-cell subpopulations really doing in chagas disease? PMID- 15275408 TI - The role of T cells in Trypanosoma cruzi infections. PMID- 15275409 TI - Confirmation of the conspecificity of two American vectors of the agent of lyme disease. PMID- 15275410 TI - Applications of molecular marker analysis to mosquito vector competence. AB - A rapidly expanding cadre of molecular biology techniques is being developed for human and plant genetics, including development of the technology to identify large numbers of genetic markers and to evaluate these markers relative to phenotypic observations. In this review, David Severson discusses applications of these techniques for the analysis of mosquito vector competence. PMID- 15275411 TI - The assembly of kinetoplast DNA. AB - The unusual structure of the kinetoplast DNA (kDNA) of trypanosomatids requires unique replication mechanisms. Deciphering the mechanisms that regulate the network assembly has been a challenge for many years. A better understanding of these processes was facilitated by recent studies on the fine structure of resting and replicating kDNA networks. In this review, Joseph Shlomai discusses our current view of the structural and mechanistic aspects of the assembly of kinetoplast DNA. PMID- 15275412 TI - Energy generation in parasitic helminths. AB - Although parasitic helminths are a very heterogeneous group of organisms, they share many interesting properties in their energy metabolism. In certain stages of their life cycle, they all have a large capacity for anaerobic functioning. In other stages, an aerobic energy metabolism prevails. Parasites have to adapt to different environments in which the availability of oxygen and food varies widely. These variations in their external conditions strongly influence their energy metabolism. Here, Louis Tielens presents an introduction to the current ideas on the bioenergetics of parasitic helminths, focusing on the differences in energy metabolism between various stages (free-living and parasitic), and paying special attention to the mechanisms involved in the transitions between the different methods of energy generation. PMID- 15275413 TI - Cellular response of protozoan parasites to host-derived cytokines. AB - Cytokines are extracellular signalling molecules, produced by different cell types and displaying a wide range of activities such as the induction or inhibition of target cell survival, proliferation and differentiation. When directly interacting with different parasites, cytokines exert similar activities, acting as growth factors and, in one of the examples given here, also enhancing parasite survival. The importance of this interaction in the natural history of parasitic diseases as well as the selective forces maintaining functional cytokine 'receptors' in protozoan parasites is discussed in this review by Marcello Barcinski and Maria Elisabete Costa-Moreira. PMID- 15275414 TI - Confirmation of the conspecificity of two American vectors of the agent of lyme disease: reply. PMID- 15275415 TI - Dilatations in mosquito ovarioles. PMID- 15275416 TI - Milestone legacies. PMID- 15275417 TI - Parasites and zoos. PMID- 15275418 TI - Immunization against Toxoplasma gondii. AB - The emergence of Toxoplasma gondii as a major opportunistic organism in immunocompromised individuals and the steady increase in economic losses due to animal toxoplasmosis have fuelled the interest in vaccine development. In this review, Fausto Araujo addresses aspects of vaccination against T. gondii with regard to the problem posed by this parasite to pregnant women and to severely immunocompromised individuals. PMID- 15275419 TI - Trypanosomiasis and trypanotolerance in cattle: a role for congopain? AB - A Trypanosoma congolense cysteine protease (congopain) elicits a high IgG response in trypanotolerant but not in trypanosusceptible cattle during primary infections. As discussed here by Edith Authie, this observation suggests that congopain, like other parasite cysteine proteases, may play a role in pathogenicity and that more efficient immune responses to congopain may contribute to trypanotolerance. PMID- 15275420 TI - Localized cellular responses to Ichthyophthirius multifiliis: protection or pathogenesis? AB - The ciliate Ichthyophthirius multifiliis is an economically important parasite in aquaculture as well as in the ornamental fish trade. The parasite has been well studied and details of life cycle, host response and pathogenesis have been characterized. Parasite development within host epithelial tissues initiates localized leukocytic infiltrations, although the relationship between these responses and host resistance is uncertain, and whether or not leukocyte responses play a role in protective immunity is unclear. Here Frank Cross outlines the character of localized cellular responses during I. multifiliis infection and discusses the contribution of such responses to protective immunity and pathogenesis. PMID- 15275421 TI - Mechanisms of survival of mice during acute and chronic Toxoplasma gondii infection. PMID- 15275422 TI - Dense-granule organelles of Toxoplasma gondii: their role in the host-parasite relationship. AB - The dense-granule organelles of Toxoplasma gondii are secretory vesicles that play a major role in the structural modifications of the parasitophorous vacuole. In this article, Marie-France Cesbron-Delauw reviews work on a prominent set of molecules recently characterized as components of the dense granules, and discusses various aspects on their post-translational trafficking and delivery within the host cell. PMID- 15275423 TI - Coenzyme Q homologs in parasitic protozoa as targets for chemotherapeutic attack. AB - The central role of coenzyme Q (ubiquinone) in cellular energy metabolism is well established. Recent work has implicated this molecule in a wide range of other cellular functions, including roles in growth control, plasma membrane oxidase and as a cellular antioxidant. In this review, Jayne Ellis presents an overview of the current knowledge of this important cellular component in species of parasitic protozoa, discusses current therapies using its analogs and proposes its potential roles in these organisms. PMID- 15275424 TI - Age and 'strain-transcending' immunity to Plasmodium falciparum. PMID- 15275427 TI - Milestone knockabout. PMID- 15275428 TI - The role of Kupffer cells in the clearance of malaria sporozoites from the circulation. AB - In the past decade, one of the most intriguing subjects in understanding the mechanism of malaria infection has been explanation of the role of Kupffer cells. These liver cells, which play an important part in the body's defense against infection, seemed to have on essential supportive role in the homing o f sporozoites. Do Kupffer cells favor the establishment of primary malaria infection? Extensive research has revealed much, but still not everything we need to know about the sporozoite-Kupffer cell affair. PMID- 15275429 TI - Proteinase inhibitors in Ascarida. AB - Ascaris suum and A. lumbricoides are intestinal parasites that survive in a hostile hydrolytic environment They contain low-molecular-weight proteins that can inactivate most of the proteinases present in the worm's surroundings. Jeffrey Hawley, Mark Martzen and Robert Peanasky suggest that host specificity of these nematodes may be associated with the efficiency of their inhibitors. PMID- 15275430 TI - Th2-like CD8 T cells: their role in protection against infectious diseases. AB - The expression of cytolytic activity and production of interferon gamma (IFN gamma) by CD8(+) T cells is thought to play a fundamental role in protection against infection by viruses and intracellular parasites. Francois Erard and Graham Le Gros have recently shown that CD8(+) T cells activated in the presence of interleukin 4 (IL-4) can switch development to a CD8(-)CD4(-)Th2-like phenotype that is not cytolytic and that does not produce IFN-gamma. Here they speculate on whether this IL-4-induced switch is used by the host to make a more effective response against parasite invasion, or i f it is a host mechanism used by the parasite to evade protective CD8(+) T-cell responses. PMID- 15275431 TI - Human T-cell responses to malaria: mostly forgotten or committed to memory? AB - T cells have been implicated in both malaria immunity and malaria disease and factors controlling the maintenance of T-cell responses over time may alter the clinical outcome o f infection. Michael Good and Janine Bilsborough have compared the T-cell responses to epitopes from the Plasmodium vivax and P. falciparum circumsporozoite proteins and here discuss the issue of T-cell memory as it applies to malaria. PMID- 15275432 TI - P-glycoproteins in nematodes. AB - The P-glycoprotein (Pgp), a member of a group of integral membrane proteins that contain the ATP-binding cassette, is widely represented in the animal kingdom. A family of Pgp homologues has recently been described in nematodes. Pgps have been implicated in drug resistance in Plasmodium and in other parasitic protozoa, so the interest of porasitologists now focuses on whether or not they are involved in anthelmintic resistance. Their role in normal cellular functions is also discussed in this article by Nick Songster. PMID- 15275433 TI - The role of chemokines in Schistosoma mansoni granuloma formation. AB - The eradication of parasitic diseases, such as schistosomiasis, has been the focus of investigations worldwide for many decades. However, attempts to control their continual spread have, at best, been met with limited success. In the face of these results, it is important to attempt to understand and thus to control the pathology of these widespread diseases. In this review, Nicholas Lukacs, Steven Kunkel, Robert Strieter and Stephen Chensue focus on a family of cytokines that play a pertinent role for leukocyte recruitment in chronic inflammation. PMID- 15275434 TI - MVR-PCR analysis of hypervariable DNA sequence variation. AB - Techniques for accurate marking of infectious microbial agents circulating in populations would be very useful to epidemiologists. In this article, David Arnot, Cally Roper and Ali Sultan review recent progress in transferring MVR-PCR DNA finger-printing techniques from human forensic medicine to parasitology. PMID- 15275435 TI - The malaria/G6PD hypothesis revisited. PMID- 15275436 TI - cdc2-related protein kinases and cell cycle control in trypanosomatids. AB - The molecular mechanisms that control the cell cycle have been studied extensively in yeast and higher eukaryotes. Investigations have centred on the cyclin-dependent kinase family of serine/threonine protein kinases, the best characterized of which is cdc2, a key regulatory element in the control of mitosis. Cell cycle control plays an important role in trypanosomes and Leishmania, not only in cellular proliferation, but also in the developmental system that controls the transfer of the parasite between hosts. In this review, Jeremy Mottram compares the family of trypanosome cdc2-related kinases with that of yeast and the higher eukaryotes. PMID- 15275437 TI - Transgenic mice and the study of cytokine function in infection. AB - Most information about the involvement of the different cytokines in immunity to infection has been obtained by the administration to infected animals of recombinant molecules or of antibodies against them. Now another approach to the study of cytokine function, in vivo, is available in the form of transgenic mice that express a transgene encoding a particular cytokine, and of 'knockout' animals, in which a cytokine gene, or a gene for its receptor (which usually comes to the same thing), have been rendered inactive by targeted disruption. While a few of these lines of mice have been analysed for their response to infection by protozoan parasites or worms, more have been tested for their ability to withstand intracellular infections by bacteria or viruses. In this review, Janice Taverne outlines those described to date in which the immune (or immunopathological mechanisms concerned may be relevant to parasitic diseases. PMID- 15275438 TI - The malaria/G6PD hypothesis revisited: reply. PMID- 15275439 TI - Pathogenicity or virulence? PMID- 15275441 TI - Leishmania infantum tropism: strain genotype or host immune status? AB - In apparently immunocompetent patients, Leishmania infantum provokes a spectrum of disease, ranging from simple skin lesion to severe visceral leishmaniasis, that is determined mainly by the protozoan genotype. In HIV-positive individuals, leishmanial infection results almost exclusively in visceral disease. In this review, Luigi Grodoni and Marina Gromiccia discuss the role o f the intrinsic virulence of L. infantum strains and the immune condition of the host, and focus on recently described mechanisms of immunological control of leishmanial infection. PMID- 15275442 TI - Insects: handle with care. PMID- 15275443 TI - T-cell responses to the trypanosome variant surface glycoprotein: a new paradigm? AB - During the past decade, extensive knowledge has been gained with respect to the cellular and molecular mechanisms associated with variant surface glycoprotein (VSG) gene switching in trypanosomes. However, comparatively little is known about the cellular and molecular factors that regulate the host B-cell response to VSG determinants during infection. Here, John Mansfield reflects on the nature of this response. PMID- 15275444 TI - Is phagocytosis related to virulence in Entamoeba histolytica Schaudinn, 1903? AB - Phagocytosis has been suggested as a marker of pathogenicity and virulence in Entamoeba histolytica. Irmgard Montfort and Ruy Perez-Tamayo here review the evidence for this suggestion. PMID- 15275445 TI - Asian (Taiwan) Taenia: species or strain? AB - Recent studies on the epidemoiological pattern of taenisis in Southeast Asia have indicated the existence of a third form of human Taenia which is distinguishable from Taenia saginata and T. solium. Don McManus and Josephine Bowles here review how new genetic evidence supports earlier conclusions that the Asian Taenia is a distinct entity but is closely related T. saginata, and suggests its taxonomic classification as a subspecies or strain of T. saginata is more appropriate than formal designation as a new species. PMID- 15275446 TI - Compound exocytosis and cumulative degranulation by eosinophils and their role in parasite killing. AB - The killing of metazoan parasitic larvae by eosinophils occurs following cell adhesion and the secretion o f their cytotoxic proteins onto the surface o f these targets. In eosinophils, as in mast cells and neutrophils, stimulus secretion coupling is mediated by GTP-binding proteins. In this article, Susanne Scepek Redwon Moqbel and Manfred Lindou summarize recent results indicating that the granule-fusion events activated by GTP-binding proteins lead to compound exocytosis and cumulative fusion. They propose that these exocytotic processes, following contact with opsonized larvae, may direct secretion to a restricted space defined by the site o f contact Such a focused release may be essential for effective targeting in parasite killing, thus preventing uncontrolled random diffusion of the secreted cytotoxic proteins with the possible undesirable consequences of damage to intact host tissue. PMID- 15275447 TI - Impregnated bednets and the dose-severity relationship in malaria. AB - The finding that insecticide-impregnated bednets decrease symptomatic malaria and death more than infection has been interpreted as providing evidence that dose influences seventy o f disease in malaria. The complications o f this argument in areas where much o f the population has a background parasitaemia have already been considered. In this article, Judith Glynn, Jo Lines and David Bradley explore whether or not the results of impregnated bednet trials can help to determine the existence of a dose-severity relationship for malaria. PMID- 15275448 TI - The in vitro model of tissue cyst formation in Toxoplasma gondii. AB - Fundamental advances in the development and design of drugs for toxoplasmosis will only follow a more-thorough understanding o f the basic biology of Toxoplasma gondii tissue cyst formation. Tim McHugh, Richard Holliman and Philip Butcher here describe the in vitro model of tissue cyst formation in T. gondii. PMID- 15275449 TI - There is no ducking the duct. PMID- 15275450 TI - Biology of mammalian Isospora. AB - Isospora species are coccidial parasites that can cause serious disease in humans and pigs. Disease is observed less frequently in non-human primates, dogs or cats. Isospora species do not produce disease in horses, domestic ruminants or domestic poultry, and reports of isosporan oocysts in the feces of these hosts probably represents pseudoparasites that originated in feed or water contaminated with wild bird feces. David Lindsay and Byron Blagburn here summarize what is known about the biology of the Isospora species of domestic animals and non-human primates. PMID- 15275451 TI - Blood monocytes: differing effector role in experimental visceral versus cutaneous leishmaniasis. AB - Irrespective of the tissue infected or the strain involved, all Leishmania species selectively parasitize and replicate within the resident tissue macrophage. Henry Murray here discusses the role of a second mononuclear phagocyte, the blood monocyte, which is also attracted to leishmanial lesions, but which appears to play quite different roles in experimental visceral versus cutaneous infection: in visceral disease, the monocyte is a critical host defense effector cell, in cutaneous disease, it may, paradoxically, serve to perpetuate intracellular infection. PMID- 15275452 TI - There is no ducking the duct: reply. PMID- 15275455 TI - PFGE: improved conditions for rapid and high-resolution separation of Plasmodium falciparum chromosomes. PMID- 15275456 TI - Endotoxins and their significance for murine trypanosomiasis. AB - The study of endotoxins is complicated by their heterogenous nature, their multiple effects and the complex methodologies required for their identification. In this brief review, Vic Pentreath summarizes how the substances have been implicated in the pathogenesis of several diseases caused by parasitic protozoa, and how the parasites (eg. Plasmodium falciparum and Trypanosoma cruzi) may themselves contain endotoxin-like materials. Recent studies have shown that, during T. b. brucei infection in mice, serum endotoxin levels become markedly elevated and that endotoxin-like substances are also present in the purified parasite extracts. PMID- 15275457 TI - Cellular immune response in Pneumocystis carinii infection. AB - Pneumocystis carinii is an important pulmonary pathogen causing disease in immunocompromised individuals. The majority o f conditions predisposing to Pneumocystis pneumonia are associated with profound defects in cellular immunity. Although our understanding o f the host response to the organism is still limited, advances in antigen preparation and the availability o f animal models have permitted an improved understanding of some aspects o f the cell-mediated immune response to Pneumocystis. In this review, George Smulian and Sue Theus will highlight recent advances in our knowledge regarding the role of macrophages, T cells and cytokines in the response to the organism. PMID- 15275458 TI - The role of eosinophils in Angiostrongylus cantonensis infection. AB - Angiostrongylus cantonensis is the causative agent of human eosinophilic meningoencephalitis in the Pacific Islands and Southeast Asia. Prominent eosinophilia in the cerebrospinal fluid (CSF) of the patients has been used as one of the diagnostic criteria for the disease but the role(s) of the CSF eosinophils has remained to be elucidated. In this article, Kentaro Yoshimura, Hiroko Sugaya and Kazuto Ishido discuss the involvement of CSF eosinophils in the killing of intracranial worms and the damage of the central nervous system of the hosts, and consider why eosinophils in A. cantonensis infection play a more important role in nonpermissive hosts (including humans) than in the permissive rat host. PMID- 15275459 TI - The role of IL-5 in the immune responses to nematodes in rodents. AB - In this review, Masataka Korenoga and Isoo Toda discuss the role of interleukin 5 (IL-5) in immune responses caused by parasitic infections. Recent studies have shown that eosinophilia in parasitic infections is dependent on IL-5. Although eosinophils have been thought to be potent effector cells (from results of experiments, in vitro), their role in protective immunity to parasites is controversial. PMID- 15275460 TI - Selection for varroatosis resistance in honeybees. AB - The parasitic mite Varroa jacobsoni is a major problem for beekeeping worldwide. It can be controlled efficiently with a variety of ocaracides. However, Robin F.A. Moritz argues that, owing to the risk of honey contamination and the costs involved with continuous treatment of honeybee (Apis mellifera L.) colonies, there is a pressing need to find alternative ways of varroatosis control. A variety of physiological and behavioural traits of the honeybee are known to control efficiently the development and spread of V. jacobsoni infestation. Breeding of a varroatosis-resistant honeybee seems possible and selection could offer swift results if one capitalizes on the male haploid population structure of the honeybee. PMID- 15275461 TI - A universally applicable diagnostic approach to filarial and other infections. AB - Antibody-based assays for the diagnosis of filarial and other infections cannot reliably distinguish between past and current infection, nor can they be used to assess the efficacy of chemotherapy. In this article, Thomas Nutmon, Peter Zimmerman, Joseph Kubofcik and Donna Kostyu discuss how the detection of parasite specific PCR products using on ELISA-based assay may overcome these problems. PMID- 15275462 TI - Trans-sialidases in the insect-vector stages of African and American trypanosomes. PMID- 15275463 TI - Conservation of heat-shock proteins in Trypanosoma cruzi. AB - Heat shock is an integral part of the life cycle of Trypanosoma cruzi. Here, Edson Rondinelli reviews the parasite's response to stress. PMID- 15275464 TI - Chromosome rearrangements in Giardia lamblia. AB - Recent studies have shown that the genome of Giardia lamblia is plastic. Clinical isolates exhibit extensive karyotypic heterogeneity and chromosome rearrangements occur frequently, in vitro. In this review, Sylvie Le Blancq looks at genome organization and the impact of DNA rearrangement events. PMID- 15275465 TI - Trans-sialidases in the insect-vector stages of African and American trypanosomes: reply. PMID- 15275466 TI - Nematophagous fungi and free-living nematodes. PMID- 15275468 TI - What is the function of MSP-I on the malaria merozoite? AB - In the absence o f any clear enzymatic activity, attempts to define the role of merozoite surface protein-I have focused mainly on analysis of its structure, on its interaction with the immune system and on binding assays. But how does our knowledge of the structure o f this protein contribute to functional studies? Are there data to suggest a role in the evasion of effective host immune responses? Binding studies have used the intact protein or various fragments and peptides, but do such approaches provide a reliable indicator of function? In this article, Tony Holder and Mike Blackman review these areas. PMID- 15275469 TI - Attachment and invasion of host cells by Toxoplasma gondii. AB - Recent studies indicate that Toxoplasma gondii attachment is mediated via a parasite ligand-host cell receptor interaction. Lloyd Kosper and Jose Mineo here survey factors involved in the attachment to and penetration and invasion of host cells by T. gondii. PMID- 15275470 TI - Molluscan defence mechanisms: immunity or population biology? AB - In recent studies of associations between host and parasite, there has been a tendency to consider the internal defence mechanisms of the mollusc as pivotal in the relationship. However, evolution is a population-based phenomenon, and it is by considering populations rather than individuals that the success of these associations can best be appreciated. Among susceptible host species within the biocoenosis, truly resistant strains appear to be remarkably rare. Clive Shiff here examines mechanisms of defence or avoidance among molluscan hosts of tremotodes from both the internal and ecological perspective. PMID- 15275471 TI - Interleukin 12 and the regulation of CD4+ T-cell subset responses during murine Leishmaniasis. AB - Interleukin 12 is unique among cytokines in that it is capable of protecting genetically susceptible mice against progressive infection with Leishmania major. Because of the probable causal relationships between CD4(+) T-cell differentiation, cytokine production and disease outcome, this cytokine may prove useful as a component of cytokine-bosed therapies and Thl-selective vaccines, as discussed here by Frederick Heinzel. PMID- 15275472 TI - Extraction of intraerythrocytic malarial parasites by phagocytic cells. AB - Phagocytosis is an intricate process adopted by some unicellular organisms as a feeding behaviour. It has developed in the tissues of multicellular organisms, both vertebrates and invertebrates, as a defence system to confine and eliminate foreign matter and, in this manner, protect the host against infection. During evolutionary development, phagocytic cells have evolved to show greater specificity. Lakshmi Kumaratilake, Antonio Ferrante, Jaliya Kumaratilake and Anthony Allison here describe a unique mechanism used by phogocytic leukocytes to engulf intra-erythrocytic malarial parasites. PMID- 15275473 TI - Immunoepidemiology of human schistosomes: taking the theory into the field. AB - Much is known about human immune responses to schistosome infection, but it has proved difficult to determine the impact of these responses on schistosome epidemiology in the field. In this paper, Mark Woolhouse compares epidemiological patterns from field data with the predictions o f simple mathematical models of different immunological processes. The comparison gives some indications as to which types of immune response may be important, and of their strength and duration. The results are consistent with a significant impact of a process similar to 'concomitant' immunity, with a possible role for anti-fecundity effects. PMID- 15275474 TI - A plea for consistency. PMID- 15275476 TI - Effect of chemoprophylaxis on immunity to gastrointestinal nematodes in cattle. AB - Control of gastrointestinal nematodes in first-season grazing cattle is largely based on either strategic anthelmintic treatment or use of anthelmintic-release devices. Control is evolving towards more efficacious anthelmintics and delivery systems, almost annihilating parasite contact. Here, Jozef Vercruysse, Hans Hilderson and Edwin Claerebout focus on the inverse relationship between the intensive use of modern anthelmintics and the build-up of immunity to gastrointestinal nematodes. PMID- 15275477 TI - The inducible antibacterial peptides of insects. AB - Insects respond to bacterial challenge by the rapid and transient synthesis of a large number of potent antibacterial peptides that are active against many different bacteria. Two families of inducible antibacterial peptides are well characterized: the cecropins and the insect defensins. A rapidly increasing number of proline- and glycine-rich peptides are reported from various insect species together with cecropins and insect defensins. In this review, Stephane Cociancich, Philippe Bulet, Charles Hetru and Jules A. Hoffmann give an update of our current information on the induced antibacterial peptides. PMID- 15275478 TI - GPI-anchor synthesis. AB - The glycosyl phosphatidylinositol (GPI) anchor of membrane proteins is widely distributed in eukaryotes and parasitic protozoa. The structure and biosynthetic pathway of its core have been elucidated and appear to be conserved in various species. Some of the genes involved in mammalian GPI-anchor biosynthesis have recently been isolated using GPI-anchor-deficient mutant cell lines and expression cloning methods. One of these genes proved to be responsible for a GPI anchor deficiency known as paroxysmal nocturnal hemoglobinuria. Since the core of the GPI anchor is variously modified in different species and since there may be other differences between its biosynthetic pathway in parasites and their hosts, this pathway could be a target for chemotherapy. In this review, Taroh Kinoshita and Junji Takeda focus on the GPI-anchor biosynthetic pathway and the genes involved in it. PMID- 15275479 TI - A plea for consistency: reply. PMID- 15275481 TI - Milestones in parasitology (1). PMID- 15275480 TI - Milestones in parasitology (2). PMID- 15275482 TI - Use of helper T cells to identify potential vaccine antigens of Babesia bovis. AB - Babesia bovis is an economically important hemoprotozoon parasite o f cattle that is prevalent in many, tropical and subtropical regions o f the world. Effective vaccines against this tick-transmitted parasite consist o f live organisms attenuated by passage through splenectomized calves. However, the nature o f acquired resistance to challenge infection with heterologous isolates of this parasite has not been clearly defined. Unsuccessful attempts to select protective antigens have relied upon the use of antibodies to identify immunodominant proteins. In this review, Wendy Brown and Allison Rice-Ficht discuss the limitations of this approach and the rationale behind using helper T cells to select potential vaccine antigens. PMID- 15275483 TI - Conserved proteins as immunogens: glutathione S-transferase of Schistosoma. AB - Although some immunogenic proteins of parasites evolve rapidly and seem to have no function other than the eliciting o f an immune response, others are proteins that are ordinarily conserved because of functional constraints. The 26 kDa and 28 kDa glutathione S-transferases (GSTs) of Schistosoma would seem to fall in the latter category. However, in the generally more conserved N-terminal portion o f the molecule, the 28 kDa GSTs have evolved over twice as fast at nor-synonymous nucleotide sites as have the 26 kDa GSTs. It is possible that this accelerated rate o f evolution results from natural selection by the host immune system, as discussed here by Austin Hughes. PMID- 15275484 TI - The existence of lipophosphoglycanlike molecules in Trichomonads. AB - The trichomonods, Trichomonas vaginalis and Tritrichomonas foetus, appear to express novel lipophosphoglycon (LPG) like glycoconjugates on their cell surface, which are structurally distinct from Leishmania LPGs'. In this article, Bibhuti Singh discusses the structural and cellular aspects o f these molecules, and speculates on their biological significance. PMID- 15275485 TI - Borrelia burgdorferi and the macrophage: routine annihilation but occasional haven? AB - Borrelia burgdorferi, the agent for Lyme disease, has a typical pattern of bacterial interaction with phagocytes: attachment, stimulation o f release o f inflammatory mediators and, in most cases, ingestion and killing. Spirochetes are killed extracellulorly by antibody plus complement via the classical pathway, as well as by phagocytes through apparently nonoxidative means. Yet rare persistent spirochetes (mutants?) have been identified both in patients' tissues and in cells grown in vitro. Ruth Montgomery and Stephen Malawista here ask: are some Borrelia wolves in sheeps' clothing, evading macrophage anti-microbial action? PMID- 15275486 TI - Comparative ecology and epidemiology of lyme disease and tick-borne encephalitis in the former Soviet Union. AB - The recently defined range of human Lyme disease in Russia coincides with that of the intensively studied agent of tick-borne encephalitis (TBE). Although identical vectors transmit both infections during the same seasons and in the same locations, more than twice as many people suffer from Lyme disease as from TBE. Edward Korenberg here describes characteristic features of vectors with respect to their seasonal activity, abundance, rates of infection by TBE and Lyme disease agents, as well as incidence of human-tick contact, and how these factors determine the incidence of TBE and Lyme disease in different regions. PMID- 15275487 TI - Leishmaniasis and AIDS co-infection: the Spanish example. AB - There are an estimated 300 instances of Leishmania/HIV co-infection, of which 200 have occurred in Spain. Jorge Alvar here asks: is there an epidemiological or immunological basis for this high proportion? PMID- 15275488 TI - Gene transfer and antisense nucleic acid techniques. AB - Attempts to suppress a harmful genetic trait by antisense means, or to restore a normal phenotype by gene transfer, attract much publicity. This is especially the case where clinical trials incorporating such methodologies have been initiated, such as antisense oligonucleotide therapies for some types of leukaemia, antisense gene-transfer therapy for a form of lung cancer, and gene-transfer therapies for adenosine deaminase deficiency, severe combined immunodeficiency disease, and various forms of cancer including brain tumours and melanoma. However, translation of laboratory success into treatment or control of disease is unlikely to be straightforward. Here, Nick Miller and Richard Vile summarize the rationale, problems and potential of such techniques as applied to parasitic disease. PMID- 15275489 TI - Variability and its implications for host-parasite interactions. AB - Variability in host-parasite interactions has considerable impact on the ecology and evolution of parasites and on the epidemiology of disease. The nature of the impact depends largely on the level of ecological organization where variability occurs: variability of parasites within their individual hosts, variability of host individuals within populations, or variability of hosts and parasites among populations. In this review, Paul Schmid-Hempel and Jacob Koella give some examples of variability at each of these levels, with particular emphasis on microparasites (defined broadly as viruses, bacteria and protozoa), consider the maintenance of the variability, and describe the implications of variability for the epidemiology of disease and the ecology of host parasite associations. In particular, they describe how variability at each level of ecological organization can affect the perception of AIDS and the evolution of virulence. PMID- 15275490 TI - How Plasmodium secures nutrients: new targets for drugs? PMID- 15275491 TI - Vaccination strategies against Tritrichomonas foetus. AB - Immunoprophyloxis for bovine trichomoniosis has been a priority because of the high prevalence o f the disease, the considerable economic loss due to the infection and the lack of approved chemotherapeutic agents. The commercial availability of first-generation vaccines provides hope not only for even more effective immunization regimens far this disease, but also for other protozoal infections and for sexually transmitted diseases (STDs) caused by a wide variety of infectious agents. At present, efficacious vaccines for protozoal diseases and for STDs are rare. Since information gained on immunization against Tritrichomonas foetus may have broad significance for control of these two classes of infection,it is important to explore the biological basis of protection against this protozoal infection of the reproductive tract In this paper, Lynette Corbeil reviews data on host-parasite relationships in bovine trichomoniasis as a basis for developing vaccine strategies. PMID- 15275492 TI - Intermediary metabolism of Trypanosoma cruzi. AB - In this article, Julio Urbino discusses the characteristics o f the intermediary metabolism of Trypanosoma cruzi (the causative agent of Chagas disease), which are responsible for the unusual capacity of this parasite to use carbohydrates or amino acids as carbon and energy sources without drastic changes in its catabolic enzyme levels(1-3). Many, but not all, o f the metabolic capabilities of this organism are shared with Leishmania and the procyclic form o f the African trypanosomes, and the reviewer presents a metabolic model which is also consistent with the information available on these other parasites(2,4). PMID- 15275493 TI - Germ-line determination in Caenorhabditis and Ascaris: will a helicase begin to unravel the mystery? AB - How cell lineages are established during development in higher eukaryotes is being addressed by geneticists and by developmental and molecular biologists. In Drosophila melanogaster, a gene corresponding to a germ-line-specific RNA helicase, vasa, has been shown to be a component o f the posteriorly localized germ granules o f the developing embryo. A putative RNA helicase, glh-I r which appears germ-line specific in its expression, has recently been reported from the free-living nematode Caenorhabditis elegans. Parasitologists studying the nematode Ascaris lumbricoides var. suum have found it to be a useful complement to Caenorhabditis. Deborah Roussell, Michael Gruidl and Karen Bennett predict that Ascaris will be valuable in determining the role played by germ-line helicases in development. PMID- 15275494 TI - Organization of chromosomes in Plasmodium falciparum: a model for generating karyotypic diversity. AB - Chromosomes of the human malaria parasite, Plasmodium falciparum, are compartmentalized into conserved and polymorphic domains. The conserved domains are transcribed and localized within the central region o f a chromosome, whereas the polymorphic domains are transcriptionally silent and situated at chromosome ends. Unlike the central chromosomal domains, chromosome ends contain repetitive sequence elements. In this review, Michael Lonzer, Derik de Bruin, Samuel Wertheimer and Jeffrey Ravetch propose that the chromosome ends play a functional role in generating genetic diversity by promoting meiotic and mitotic recombination and chromosomal rearrangement events. PMID- 15275495 TI - Use of PCR in the field. AB - Polymerase chain reaction (PCR) represents a powerful new technology with a variety of field applications. While most of these are still experimental, implementation of PCR-based detection of Onchocerca volvulus in black flies, and subspecific differentiation strongly suggest that potential problems can be overcome. Because of high sensitivity and specificity, PCR provides in some cases the only means to address central parasitological questions, and may well become the 'gold standard' by which other diagnostic techniques are measured. In spite of these advantages, routine implementation of PCR,at present,requires transportation of samples to a central facility for processing, and personnel whose technical competence is high. In addition, reagents are expensive. Robert Barker here weighs up these considerations with regard to the potential utility of PCR assays for some applications. PMID- 15275496 TI - Eradication of the screwworm from Libya using the sterile insect technique. AB - The introduction in 1988 of the New World screwworm into Libya presented a serious threat to the livestock and wildlife sectors o f the African continent and the Mediterranean region. In this article, Moises Vargas-Teran, Brian S. Hursey and Edward P. Cunningham describe the action taken to determine the extent of the problem, to prevent the spread of the infestation and to eradicate the fly from the region using the sterile insect technique. PMID- 15275497 TI - Pathogenicity, virulence and Entamoeba histolytica. PMID- 15275498 TI - What can GPI do for you? AB - Various functions for glycosylphosphatidylinositol (GPI) protein anchors have been described in mammalian and protozoan systems. These data suggest that some functions are common to higher and lower eukaryotes, whereas others may represent adaptations that are specifically advantageous to either unicellular or metazoan organisms. In this article, Mike Ferguson discusses the current theories of GPI function that have relevance to protozoan parasites and their mammalian hosts. PMID- 15275499 TI - Invariant surface proteins in bloodstream forms of Trypanosoma brucei. AB - Antigenic variation of the glycoprotein forming the coat of African trypanosomes has been a dominant field of investigation for many years. The extravagant potential of these parasites to change their surface coat has destroyed hopes for a vaccine based on the variant surface glycoprotein. Recently, there has been a rising interest in the characterization of surface proteins that are not subject to antigenic variation. In this review, Peter Overath, Maliha Chaudhri, Dietmar Steverding and Karl Ziegelbauer summarize the present evidence for the occurrence, cellular localization and function of invariant surface proteins in Trypanosoma brucei. PMID- 15275501 TI - Taxonomic uncertainty and Blastocystis (Protista:Sarcodina). PMID- 15275500 TI - Mariner: its prospects as a DNA vector for the genetic manipulation of medically important insects. AB - Originally described in Drosophila mauritiana, the mariner transposable element has very recently been identified in 63 other insect species, representing ten insect orders, and including the malaria-vector mosquito, Anopheles gambiae. Ann Warren and Julian Crampton here discuss how transposable elements can be exploited as valuable research tools for the molecular characterization of genomes and as DNA vectors for genome manipulation and the 'creation' of transgenic organisms. PMID- 15275502 TI - Clinical immunity to Plasmodium falciparum. PMID- 15275504 TI - Medicinal plants in the fight against leishmaniasis. AB - Despite the tremendous progress mode in the understanding o f the molecular biology of Leishmania and the clinical possibilities presented by some experimental chemotherapeutic agents, no new drugs have been developed for the treatment of leishmaniasis since the introduction of the pentovalent antimoniols more than 50 years ago. As reviewed here by Maurice M. Iwu, Joan E. Jackson and Brion G. Schuster, recognition of the current extensive use of herbal therapy in Leishmania-endemic regions has renewed interest in evaluation of plant remedies used in traditional medicine as sources of potential antileishmanials. PMID- 15275505 TI - The species specificity of Bulinus-Schistosoma interactions. AB - Despite certain inherent drawbacks, advances have been made recently in furthering our understanding of Bulinus-Schistosoma interactions. By highlighting these, Terry Preston and Vaughon Southgate hope to stimulate further research into this neglected but important area. PMID- 15275506 TI - Rosetting. AB - Why do some individuals get severe falciparum malaria while others don't? Rosetting (the binding of uninfected erythrocytes to Plasmodium falciparum infected erythrocytes), together with endothelial cytoadherence, has been shown to play a crucial role in the obstruction of the microvosculoture in P. falciparum malaria. Here, Mats Wahlgren, Victor Fernandez, Carin Scholonder and Johan Carlson review the literature surrounding rosetting. PMID- 15275507 TI - SPf66 malaria vaccine trial: an update. PMID- 15275508 TI - Axenic culture of African trypanosome bloodstream forms. AB - In this article, Hiroyuki Hirumi and Kazuko Hirumi review recent technical developments of in vitro systems that support the growth of bloodstream forms of African trypanosomes in the absence of mammalian feeder layer cells, which were required in earlier methods. PMID- 15275509 TI - Antigenic diversity and the transmission dynamics of Plasmodium falciparum: the clonality/sexuality debate revisited. PMID- 15275510 TI - Nearly right or precisely wrong? Natural versus laboratory studies of vector borne diseases. AB - Recent studies that compare experimental vector-borne disease systems incorporating elements of natural pathogen-vector-host interactions with model systems using unnatural associations have highlighted quantitative, and even qualitative, differences in the results. Here, Sarah Randolph and Pat Nuttall argue that the use of mathematical models to explore epidemiological processes and patterns depends on accurate parameter values obtained from natural systems. PMID- 15275511 TI - Pathogenicity of trypanosomatids on insects. AB - More and more effects of trypanosomatids on insects have been recognized in the past few years. Here, Gunter A. Schaub reviews such effects, classifying the flagellates according to the intensity of the effects on the insect host into pathogenic, subpathogenic and apathogenic trypanosomatids. He emphasizes that subpathogenic trypanosomatids which cause only minor effects under optimal conditions might act synergistically with natural stressors, thereby being an important regulatory factor in insect populations. PMID- 15275513 TI - Old Uncle Tom Cobbley et al. PMID- 15275512 TI - Standardized nomenclature of parasitic diseases. PMID- 15275514 TI - Induction of signal transduction pathways in lymphocytes infected by Theileria parva. PMID- 15275515 TI - Cycles within cycles: the interplay between differentiation and cell division in Trypanosoma brucei. AB - The life cycle o f the African trypanosome is divided between the mammal and the tsetse. Those life cycle stages which traverse between these two hosts appear to be pre-adopted for survival in their new habitat They are also non-dividing. Here, Keith Matthews and Keith Gull discuss how and why trypanosomes might enmesh the control o f their cell cycle with their regulation o f the transition between different life cycle forms. PMID- 15275516 TI - A strain theory of malaria transmission. AB - Does the fact that the risk o f getting malaria is high in most endemic areas mean that it will be impossible to control through vaccination? Not if malaria is composed of several mildly transmissible strains, and what we are measuring as the high risk is the probability of being infected by any one of the several strains circulating independently within the same area. In this article, Sunetra Gupta and Karen Day discuss a strain theory of malaria transmission that fits both recent serological and molecular observations and more conventional epidemiological data on age distributions of infection and disease. Their analyses suggest that the transmissibility of malaria has been grossly overestimated, and that the control of malaria through vaccination may be far easier than previously assumed. PMID- 15275517 TI - Superoxide dismutase. AB - Eric James here discusses the molecular forms of superoxide dismutase and looks at its potential role in the pathogenesis of parasitic infections. PMID- 15275518 TI - Parasites and T helper cell development: some insights. AB - Five years after the initial observations implicating the T helper (Th)-cell dichotomy (Th1/Th2) as the focal point in the immunoregulation of murine infection with Leishmania major, investigation has shifted to the factors that govern the differentiation of a specific immune response from its pre-immune of undifferentiated state. In this article, Steven Reiner focuses on the most recent advances concerning the lineage commitment of mature Th-cell populations, showing how new techniques [such as polymerase chain reaction (PCR) and transgenic mice] have allowed for a more-careful dissection of the early evolution of an immune response. PMID- 15275519 TI - Maximum likelihood for parasitologists. AB - In quantitative biology, observed data are fitted to a model that captures the essence of the system under investigation in order to obtain estimates of the parameters of the model, as well as their standard errors and interactions. The fitting is best done by the method of maximum likelihood, though least-squares fits are often used as an approximation because the calculations are perceived to be simpler. Here Brian Williams and Chris Dye argue that the method of maximum likelihood is generally preferable to least squares giving the best estimates of the parameters for data with any given error distribution, and the calculations are no more difficult than for least-squares fitting. They offer a relatively simple explanation of the methods and describe its implementation using examples from leishmaniasis epidemiology. PMID- 15275520 TI - The ParaSight-F test: a simple rapid manual dipstick test to detect Plasmodium falciparum infection. AB - A rapid diagnostic for Plasmodium falciparum based on an antigen capture has been incorporated in a simple, easily interpreted dipstick by Becton Dickinson Advanced Diagnostics. In this article Clive Shiff, Japhet Minjas and Zul Premji discuss its evaluation in rural Tanzania and the implications of such a test in handling malaria cases under field conditions. PMID- 15275521 TI - Molecular biology of natural resistance-associated macrophage protein. PMID- 15275522 TI - The pathogenesis of human cerebral malaria. PMID- 15275523 TI - Economic losses caused by foodborne parasitic diseases. AB - Fragmentary data indicate that zoonotic parasites cause human illnesses with medical costs and productivity and disability losses totalling billions of dollars annually. Food is an important vehicle for some of these parasitic diseases. The cost to public health is not reflected in the priorities given to these parasitic diseases in either research or public health planning. In this article, Tanya Roberts, Darwin Murrell and Suzanne Marks discuss the cost of toxoplasmosis, taeniasis, cysticercosis, trichinellosis and other foodborne parasitic diseases. PMID- 15275524 TI - Plant-cyst nematode and plant-root-knot nematode interactions. AB - Root-knot nematodes and cyst nematodes are obligate plant parasites that cause extensive damage to the agriculture of both temperate and tropical countries. In this review, Andreas Niebel, Godelieve Gheysen and Marc Van Montagu describe how, in the past decade, the use of molecular techniques has provided new insights in the complex interactions between these sedentary plant-parasitic nematodes and their infected host plants. They give an account of the progress in our understanding of both the parasite and the host during compatible and incompatible interactions. They also outline the importance of a new model host system. Arabidopsis thaliana. PMID- 15275525 TI - The role of salivary vasodilators in bloodfeeding and parasite transmission. AB - In this paper, Donald Champagne reviews the salivary vasodilators, points to effects of similar compounds that may be shared by the insect substances, and discusses the potential significance of these effects with regard to parasite transmission. PMID- 15275526 TI - Beef as a source of trichinellosis. PMID- 15275527 TI - More on endotoxins and their significance for murine trypanosomiasis. PMID- 15275528 TI - Support for the duct. PMID- 15275530 TI - Giardia infection in farm animals. AB - Giardia infection is prevalent in farm animals from an early age. Despite the wide occurrence of the infection and recent understanding of its epidemiology, many aspects of giardiasis of farm animals remain unclear. In this article, Lihua Xiao discusses the prevalence, patterns, sources and clinical importance of Giardia infection in a variety of farm animals. PMID- 15275531 TI - What is clinical malaria? Finding case definitions for field research in highly endemic areas. AB - In non-endemic areas, the diagnosis of clinical malaria may be made on the basis of fever and a positive blood film. However, in areas of high endemicity, asymptomatic parasitaemia is very common: to assume that a child who presents with fever and parasitaemia is ill from malaria will result in overdiagnosis. In this article, Jo Schellenberg, Tom Smith, Pedro Alonso and Richard Hayes discuss the relationship between fever and parasite density in such areas, and show how the proportion of fevers due to malaria (the attributable fraction) can be estimated and used to evaluate case definitions for use in field trials. PMID- 15275532 TI - The regulation of procyclin expression in Trypanosoma bruceli: making or breaking the rules? AB - The identification of procyclins as stage-specific coat proteins of procyclic forms of Trypanosoma brucei has not only provided a convenient molecular marker for the differentiation of bloodstream-form trypanosomes into procyclic forms, but has also allowed some important insights into gene regulation in trypanosomes. Here, Adrian Hehl and Isabel Roditi summarize what has been learnt in the past few years about the control mechanisms that may contribute to the stage-specific expression of procyclins. PMID- 15275533 TI - Dispersion and bias: can we trust geometric means? AB - Geometric means are frequently used to estimate the intensity of parasite infection within a population. In this article, Tony Fulford uses Schistosoma mansoni field data to illustrate that such estimates are biased and, more importantly, that the degree of bias can vary markedly with host age. Similar problems plague the interpretation of prevalence. The cause can be traced back to age-dependent differences in the dispersion of parasites among hosts. PMID- 15275534 TI - Plant microtubule inhibitors against trypanosomatids. AB - Trypanosomatid protozoa are etiologic agents of several prevalent tropical diseases. Tubulins constitute 10% of the total proteins of these organisms. In addition, they are conserved within the Trypanosomatidae family but are different from that of the mammalian hosts. Since current chemotherapy has severe side effects, new compounds are urgently needed. The microtubular system provides a target for selective chemotherapy. Plant microtubule inhibitors, trifluralin and its analogues, inhibits Leishmania and Trypanosoma brucei, and Marion Chan and Dunne Fong here discuss the biosafety and potential for development of drug resistance to these compounds. PMID- 15275536 TI - Prospects for malaria control through the genetic manipulation of its vectors. PMID- 15275537 TI - The case for malaria control by genetic manipulation of its vectors. PMID- 15275538 TI - Why entomological antimalaria research should not focus on transgenic mosquitoes. PMID- 15275539 TI - Human immunity to schistosomes: concomitant immunity? PMID- 15275540 TI - Human immunity to Schistosomes: some questions. PMID- 15275541 TI - Human resistance to Schistosoma infections: age or experience? PMID- 15275542 TI - A rationale for antiparasite drug discovery. PMID- 15275543 TI - The contribution of empiricism to antiparasite drug discovery. PMID- 15275544 TI - Rational drug design in parasitology. PMID- 15275545 TI - Ducts, channels and transporters in Plasmodium-infected erythrocytes. PMID- 15275547 TI - The parasitophorous duct pathway: new opportunities for antimalarial drug and vaccine development. PMID- 15275546 TI - Nutrient transport pathways in Plasmodium-infected erythrocytes: what and where are they? PMID- 15275548 TI - Drugs or vaccines? PMID- 15275549 TI - Drugs. PMID- 15275550 TI - Vaccines or drugs: complementarity is crucial. PMID- 15275551 TI - Cerebral malaria: mediators, mechanical obstruction or more? PMID- 15275552 TI - The cytokine theory of human cerebral malaria. PMID- 15275553 TI - Cerebral malaria: the sequestration hypothesis. PMID- 15275554 TI - How plasmodium secures nutrients: new targets for drugs? PMID- 15275555 TI - Immunizing against toxic malarial antigens. PMID- 15275556 TI - Pore formation and cytolysis by Entamoeba histolytica. PMID- 15275557 TI - CD8+ T cell-coccidia interactions. AB - Host responses to coccidian parasites involve many facets of the immune system, including antigen-specific as well as antigen-nonspecific components. Hyun Lillehoj and James Trout here review the evidence that cell-mediated responses are probably the main line of defense against coccidial infection. PMID- 15275558 TI - Does helminth infection affect mental processing and educational achievement? AB - In this article, Catherine Nokes and Donald Bundy reexamine the evidence linking intestinal helminth infection to impaired cognitive function and educational outcomes. They consider first the evidence that implies a connection between intellectual dysfunction and the sequelae of infection, then the significance of correlations between infection and poor mental status, and finally the evidence from case-control and double-blind intervention studies. The article is not intended as a comprehensive summary of all the research on helminth infection and mental function - indeed the majority of research undertaken in the early part of this century is not included - but rather as a thought provoking article to highlight the difficulties with interpreting existing data and to stimulate new interest in this field. PMID- 15275559 TI - How do dilatations form in mosquito ovarioles? AB - Ovariolar dilatations have been used to estimate parity status in mosquitoes for over 40 years. The theoretical basis for this parity-diagnostic method, which was established for mosquitoes by Polovodova 1, has been challenged by a number of Russian researchers. The findings of this new school of researchers have elicited little response from those currently using the method outside Russia, except for a recent paper by Hoc and Charlwood2. This has stimulated Andrew Fox and Reinhart Brust to review the formation of ovariolar dilatations here, and compare what they term the Old and the New Schools. Although the basic application of Polovodova's method is unaffected by the premise of either School, a clear understanding of ovarian dynamics is needed to understand the method and its limits. PMID- 15275560 TI - Immunizing against toxic malarial antigens: reply. PMID- 15275561 TI - How Plasmodium secures nutrients: new targets for drugs?: reply. PMID- 15275562 TI - Green roots of malaria. PMID- 15275563 TI - Recurrent cutaneous leishmaniasis: a role for persistent parasites? AB - Leishmaniosis is, with increasing frequency, reported as an opportunistic infection of immunosuppressed individuals. Re-activation of persistent parasites may be responsible for the disease in a number of these patients. Here, Toni Aebischer reviews some of the evidence for the implication of persistent Leishmania infections in recurrent disease with the emphasis on cutaneous leishmoniasis in humans and in the mouse model. The data suggest that parasite persistence is a common feature in Leishmania infections. The availability of on excellent laboratory model provides on opportunity to study this phenomenon in detail. The findings of these analyses are likely to be important for the identification of people at risk of developing recurrent disease and for the assessment of new therapies for relapsing leishmaniasis and might also have implications for the design of a future anti-Leishmania vaccine. PMID- 15275564 TI - Spatial patterns in the abundance and distribution of parasites of freshwater fish. AB - It is not yet known whether the abundance and distribution of freshwater fish parasites form spatial patterns. Here, Rita Hartvigsen and Odd Halvorsen review the main approaches to the study of spatial patterns of these parasites, and propose an alternative approach based on meta-population theory and landscape ecology. PMID- 15275565 TI - The taxonomy of the bovine Theileria spp. AB - The classification of the benign species of Theileria of cattle is very confusing Representatives of this group of parasites appear worldwide, and are known as T. sergenti in Japan, T. buffeli in Australia and T. orientalis elsewhere. Consequently, these parasites are frequently referred to as the T. sergenti/buffeli/orientalis group. Kozo Fujisaki, Shin-ichiro Kawazu and Tsugihiko Kamio here argue that the Australian T. buffeli and British T. orientalis belong to one and the same species, and that the Japanese T. sergenti is a separate species. PMID- 15275566 TI - Ephemerality and reproductive senescence in avian filarioids. AB - Adult Eulimdana spp (Nemotoda, Filarioidea) of shorebirds (Charodrii formes) produce long-lived, skin-inhabiting microfilariae and then die and are resorbed. The host is thereafter refractory to infection. Pelecitus fulicaeatrae of coots (Fulica americana) also produces long lived, skin-inhabitingmicro filariae, but adult worms become reproductively senescent. Ephemerality of adults in Eulimdana spp and reproductive senescence in P. fulicaeatrae ensure that the skin will not become saturated with microfilariae, large numbers of which would probably be harmful to mallophogan vectors that remain on the host and are continually exposed to micro filariae while feeding. Here Roy C. Anderson and Cheryl M. Bartlett discuss the significance of ephemerality and senescence in the life histories o f avian f filarioids transmitted by permanent ectoparasites. PMID- 15275567 TI - Expression of Caenorhabditis elegans mRNA in n Xenopus oocytes: a model system to study the mechanism of action of avermectins. AB - It has recently been shown that Xenopus oocytes injected with mRNA from the free living nematode Caenorhabditis elegans express avermectin-sensitive chloride channels. Joseph Arena here reviews what is known about the mechanism of action of avermectin and how these recent results relate to the mechanism in nematodes. PMID- 15275568 TI - Visceral leishmaniasis in Rio de Janeiro. AB - Visceral leishmoniosis has recently become established in the peri-urban areas of the city of Rio de Janeiro. Mauro Marzochi, Keyla Marzochi and Raimundo Carvalho here discuss its incidence, and consider the factors involved in its establishment and propagation, as well as environmental, human, social, economic and historical factors. PMID- 15275569 TI - Development of a sporozoite vaccine. PMID- 15275570 TI - Candidate antigens for an asexual blood-stage vaccine. PMID- 15275571 TI - Transmission blocking vaccines. PMID- 15275572 TI - Towards a malaria vaccine - discussion. PMID- 15275573 TI - Cryptosporidium: notes on epidemiology and pathogenesis. PMID- 15275574 TI - Immunology of acquired resistance to ticks. PMID- 15275575 TI - Abdominal angiostrongyliasis: a problem of public health. PMID- 15275576 TI - Heartwater invades the Caribbean. PMID- 15275577 TI - Airport malaria and jumbo vector control. PMID- 15275578 TI - Ultrasound - improving the image of tropical medicine. PMID- 15275581 TI - Tsetse eradication plans for southern Africa. PMID- 15275579 TI - Appropriate disease control. PMID- 15275582 TI - Immunity in theileriosis. AB - The theilerioses can be separated on the basis of their principal pathogenic features, into a lymphoproliferative group caused by Theileria parva and T. annulata in cattle, and T. hirci in goats and sheep, and a haemoproliferative group caused by T. sergenti and T. mutans both in cattle. In the former group, proliferation of parasites within lymphoid cells followed by lymphodestruction are the main pathogenic features; whereas in the latter group, invasion and destruction of erythrocytes, causing anaemia, are more important. In addition, a number of other theilerial parasites which cause mild or inapparent infections, are found in domestic livestock. This review focuses on T. parva, the causative agent of East Coast fever (ECF) in cattle in East and Central Africa, because it is the most pathogenic species and the immunology of ECF has been more intensively studied than that of the other theilerioses. PMID- 15275583 TI - Clinical presentations of Leishmania braziliensis braziliensis. AB - Leishmania braziliensis braziliensis (Lbb) is probably the most serious, leishmanial infection of the New World. Although epidemiological information is incomplete, its distribution is believed to extend from Belize in Central America to northern Argentina, involving all countries east of the Andes. Lbb causes a variety of clinical lesions and is one of the most difficult forms of leishmaniasis to treat. We still do not know how many patients suffer from mucosal disease requiring treatment. In this review of 10 years field experience in an endemic area of Brazil, Philip Marsden shows that parasitologists have much to contribute, especially in improving diagnostic methods, developing better animal models, and providing new drugs suitable for routine treatment on a large scale. PMID- 15275584 TI - Cryopreservation of helminths. PMID- 15275585 TI - Fight the mite, and ditch the itch. PMID- 15275586 TI - Endosymbionts of trypanosomatidae. PMID- 15275588 TI - Impregnated fabrics against malaria mosquitoes. PMID- 15275587 TI - Colloidal gold probes for parasite antigens. PMID- 15275589 TI - What do people think of bednets? PMID- 15275590 TI - Sexually acquired parasitic infections in homosexual men. PMID- 15275591 TI - Developmental hormones in parasitic helminths. PMID- 15275592 TI - The epidemiology of Giardiasis. AB - Giardiasis is now known to occur not only endemically, but also as an epidemic disease spread primarily via drinking water. Giardia from different animals, although morphologically indistinguishable, vary in host specificity. This raises the possibility that lower animals may harbour representatives of this genus which are transmissible to humans. In this article, Ernest Meyer discusses our present understanding of the epidemiology of Giardia, related to problems of speciation in this genus. PMID- 15275593 TI - Catching-out the tsetse fly. PMID- 15275594 TI - Bat trypanosome models for Trypanosoma cruzi. PMID- 15275595 TI - Schistosome reproduction: The Johns Hopkins University International Conference: 1-3rd May 1985. PMID- 15275596 TI - Serodiagnostic tests for West African Trypanosomiasis. PMID- 15275597 TI - Effects of water velocity on snails and cercariae. PMID- 15275599 TI - Novel antimalarial drugs with potent broad spectrum anti-protozoal activity. PMID- 15275600 TI - Cestode neurotransmitters. PMID- 15275601 TI - The population biology of Ostertagia ostertagi. PMID- 15275602 TI - Determining the age of an insect. PMID- 15275603 TI - How do lipids affect African trypanosomes? PMID- 15275604 TI - Induction of sexual differentiation in malaria. PMID- 15275606 TI - Mosquito coils protect against bites. PMID- 15275605 TI - DNA probes for vector identification. PMID- 15275607 TI - Toxoplasmosis - a neglected disease. PMID- 15275608 TI - Toxoplasmosis in developing countries. PMID- 15275609 TI - Chronic ascariasis and malnutrition. AB - Malnutrition is the most widespread ill of mankind. Its global distribution coincides indistinguishably with that of the common roundworm. Ascaris lumbricoides is common - infecting about one-quarter of the world's population. Acute clinical ascariasis sometimes requires hospital treatment, but chronic infections contribute to long-term malnutrition. This alone should justify greater expenditure on the treatment and control of Ascaris. PMID- 15275610 TI - Diagnosis and treatment of lymphatic filariasis. AB - The lymphatic filariases, Wuchereria bancrofti, Brugia malayi, and B. timori, infect nearly 100 million people throughout the tropics, but mainly in Africa and southeast Asia. Over 900 million people live in endemic, areas at risk to the infection. The filarial parasites reproduce slowly, whereas their mosquito vectors are quickly-reproducing opportunists. Thus, although vector control can reduce the risk of transmission, the parasite itself would seem a more vulnerable target for prolonged attack. In this article, Felix Partono discusses the clinical diagnosis of f lariasis and argues that the disease can be effectively controlled by attacking the parasites in infected communities, using diethyl carbamazine (DEC) as the drug of choice. PMID- 15275611 TI - Inheritance of susceptibility to trypanosomes in tsetse flies. PMID- 15275612 TI - T-lymphocytes recognise Leishmania glycoconjugates. PMID- 15275613 TI - VIIth Ecdysone workshop. PMID- 15275614 TI - Molecular karyotyping. PMID- 15275615 TI - Low-cost housing and parasite vectors. PMID- 15275616 TI - Is there autoimmunity in Chagas disease? PMID- 15275617 TI - Autoimmune phenomena in chronic chagasic cardiopathy. PMID- 15275618 TI - Ivermectin: an update. AB - Ivermecan was introduced as an antiparasitic agent in 1981. It is now registered for animal-health use in 35 countries and is being evaluated for possible use in man. This review summarises its antiparasitic efficacy and apparent mode of action. Additional information is given in previous review articles. PMID- 15275619 TI - Genetic control of immunity to helminth infections. PMID- 15275620 TI - Pathogenic free-living amoebae. PMID- 15275621 TI - Lice, damned lice, and statistics. PMID- 15275622 TI - Advances in trypanosome culture. PMID- 15275623 TI - Forecasting the vector-borne disease implications of water development. PMID- 15275624 TI - Expanded polystyrene for mosquito control. PMID- 15275625 TI - Centered endovascular irradiation to prevent postangioplasty restenosis of arteriovenous fistula in hemodialysis patients; Results of a feasibility study. AB - PURPOSE: To report follow-up results of a prospective trial on centered endovascular gamma-irradiation (CEGI) after percutaneous transluminal angioplasty (PTA) for stenosis of arteriovenous fistula in hemodialysis patients. METHODS AND MATERIALS: Eight patients receiving PTA for recurrent (n = 4) or de novo arteriovenous fistula stenoses were treated with CEGI with iridium-192 (14 Gy). Angiography was performed after 6 and 12 months or if problems reoccurred during hemodialysis. Parameters of hemodialysis and duplex sonography were determined the day before and after PTA and after 1, 3, 6, 9, and 12 months. RESULTS: CEGI was performed successfully and without complications in seven patients. In six patients, restenosis occurred 6-52 weeks (mean 20.8 +/- 17.9 weeks) after PTA and required PTA. Parameters of hemodialysis and duplex sonography deteriorated during follow-up. CONCLUSIONS: Centered endovascular gamma-irradiation with iridium 192 immediately after PTA of fistula stenoses was a safe and feasible method but did not prevent restenosis. PMID- 15275626 TI - Intracoronary beta brachytherapy as a treatment option for high-risk refractory in-stent restenosis; Compassionate use. AB - BACKGROUND: Vascular (VBT) has clearly been shown in multiple clinical trials to decrease restenosis rates for in-stent restenosis (ISR). However, patients enrolled in these randomized clinical trials represent a select group, and the efficacy of VBT in patients with ISR who were excluded from these controlled trials due to more complex coronary anatomy requires further investigation. This study sought to define the angiographic and clinical profile and outcomes of these high-risk patients with ISR who were excluded from the randomized clinical trials and who received VBTusing Strontium-90 (Sr-90) using the Novoste Beta-Cath System through a Compassionate Use Protocol (CUP). METHODS: The study was designed as a single center, prospective, open label registry trial evaluating the use of VBT on complex instent restenotic lesions in patients who were excluded from the START and START 40 trials. In general, these patients included those with saphenous vein graft (SVG) lesions, long lesions (>35 mm), and patients with a history of more than three prior interventions. VBT using Sr-90 was delivered using the Novoste Beta-Cath System after successful angioplasty. The predetermined primary endpoint was freedom from target vessel revascularization (TVR) at 8 months, one and two years. The secondary endpoint was a composite of death, myocardial infarction (MI) and TVR at 8 months, one year, and two years. RESULTS: Between September 4, 1998 and December 6, 2000, 32 patients were treated with VBT under the UCP protocol. The mean duration of follow up was 15.3 +/- 8.3 months. There were 9 major cardiac events at eight months including one death, one acute myocardial infarction and 7 TVR. Excluding the one patient who died, 33 lesions were available for follow-up. The rate of TVR in this high-risk patient population was 21.1% (n = 7/33 lesions). The method of revascularization included one bypass surgery and 6 repeat percutaneous coronary interventions. CONCLUSIONS: This trial demonstrates that utilization of the Beta-Cath System using Sr-90 for the treatment of ISR in a patient population excluded from the randomized clinical trials due to unfavorable lesions characteristics is feasible appears to be associated TVR rates that compare favorably with the event rates of patients enrolled in other trials enrolling lower-risk groups. PMID- 15275627 TI - Impact of intravascular ultrasound-guided direct stenting on clinical outcome of patients treated for native coronary disease. AB - BACKGROUND: Despite the fact that in animal models direct stenting (DS) reduces the vessel injury, in clinical practice this treatment strategy did not reduce late restenosis as compared to conventional strategy with balloon predilatation (PD). However, the influence of DS was not evaluated when stent expansion is optimized by intravascular ultrasound (IVUS) assessment. METHODS: We analyzed the in-hospital and 1-year outcomes of patients at Washington Hospital Center who were treated with percutaneous coronary interventions and stent implantation when percutaneous intervention was guided by IVUS. Only patients treated for single de novo lesions were included. RESULTS: In 1386 patients, 251 (18.1%) were treated with DS and 1135 (71.9%) were treated with PD. Pre- and postprocedure characteristics by angiography and IVUS were similar in both groups. Postprocedure non-Q-wave myocardial infarction (MI) occurred in 4.9% of the DS group and in 12.5% of the PD group (P = .005). At 1-year follow-up, target lesion revascularization (TLR) rate was 4.9% in the DS group and 14.8% in the PD group (P = .005). DS strategy (odds ratio = .46, confidence interval = .25-.85, P = .013) was independently correlated to lower risk for revascularization in multivariate analysis. CONCLUSION: When DS is implemented by IVUS assessment, it is associated with low in-hospital and long-term events. PMID- 15275628 TI - Serotonin and thromboxane A2 stimulate platelet-derived microparticle-induced smooth muscle cell proliferation. AB - INTRODUCTION: At the sites of vascular injury, activated and aggregating platelets release small vesiculated structures called platelet microparticles (PMPs). Apart from PMPs they also release several vasoactive mediators including serotonin and thromboxane A2 (TXA2). PMPs, serotonin, and TXA2 have been shown to stimulate vascular smooth muscle cell (VSMC) proliferation. Thus, this study is designed to examine the interaction between PMPs and serotonin or TXA2 in inducing rabbit VSMC proliferation. METHODS: Growth-arrested rabbit SMCs were incubated in serum-free medium with different concentrations of PMPs with or without serotonin or TXA2. VSMC proliferation was examined by increase in incorporation of [3H]thymidine into DNA and by increase in cell number. RESULTS: PMPs stimulated DNA synthesis in a dose-dependent manner; up to an added concentration of 30 microg/ml (1489 +/- 90%) they stimulated SMC proliferation in a logarithmic fashion. Serotonin at 50 microM (345 +/- 21%) and TXA2 at 7.5 microM (900 +/- 36%) had their maximal effect. When added together, PMPs (10 microg/ml) and serotonin (5 microM), synergistically induced DNA synthesis (581 +/- 36% and 211 +/- 11% when added alone and 1201 +/- 95% when added together), whereas PMPs (10 microg/ml) and TXA2 (5 microM) additively induced DNA synthesis (581 +/- 36% and 781 +/- 56% when added alone and 1262 +/- 115% when added together). These increases in DNA synthesis were paralleled by increase in cell number. CONCLUSION: PMPs, serotonin, and TXA2 are mitogenic to SMC, and function as amplification factors to each other, suggesting that inhibition of neointimal proliferation after vascular injury may require the combined use of multiple growth factor inhibitors to simultaneously block several critical cellular activation pathways. PMID- 15275629 TI - Distal endothelial function and vascular morphology after catheter-based radiation in pig coronary arteries. AB - PURPOSE: Endovascular irradiation inhibits neointimal hyperplasia in ballooned and stented arteries but impacts both diseased and adjacent normal tissue. Little is known about the effects of irradiation on downstream vasculature. In this study, we investigated vascular function and structure of pig coronary arteries distal to sites of endoluminal irradiation. MATERIALS AND METHODS: Vasomotor responses of distal arteries to contraction of KCl and PGF2alpha and endothelium dependent (substance P and A23187) and -independent (sodium nitroprusside) relaxation were studied in naive, sham-treated, irradiated, stented, and stented plus irradiated vessels. Light and scanning electron microscopy were used to assess vascular morphology. RESULTS: Relaxations to substance P and A23187 at 1 month post treatment were significantly decreased in the irradiated group, whereas contractile response to PGF2alpha was significantly increased. Hemorrhage, mural thrombus, and inflammation were present at the upstream irradiated site; inflammatory cells were also present adherent to the endothelium in the distal segments. CONCLUSIONS: Distal vasomotor function reflects an influence from the nature of a proximal intervention. The effect of irradiation on downstream conduit arteries to increase the threshold of contractility and suppress endothelium-dependent relaxation may be related to the presence of inflammatory cells at both the upstream-instrumented site as well as the distal location. PMID- 15275630 TI - Prevention of intimal hyperplasia; Initial results with low-dose beta-radiation applied by an external vascular polymer wrap. AB - OBJECTIVE: Endovascular brachytherapy, delivered by a variety of catheter-based devices, has proven clinically effective for the inhibition of neointimal hyperplasia (NIH) after coronary and peripheral balloon/stent angioplasty. No platform, however, has been developed to deliver low-dose radiation in concert with vascular surgical operations. The purpose of this study was to evaluate the vascular response following balloon injury to the rabbit carotid artery, with and without topical low-dose 45Ca, applied by an external vascular "wrap". METHODS: Twelve rabbit carotid arteries were subjected to balloon injury by embolectomy catheter. The common carotid artery was then "wrapped" circumferentially with a biostable polyurethane membrane (Nanoskin Secant Medical, Perkasie, PA), without radiation (n = 6), or with radiation (n = 6) (45Ca approximately 50 microCi). The animals were sacrificed at 4 weeks for histologic assessment of the treated vessels. RESULTS: The 45Ca wrap inhibited NIH evidenced by trends towards reduction of intimal area (0.46 +/- 0.19 control carotid vs. 0.35 +/- 0.15 (45)Ca treated carotid arteries; P = .11), maximal intimal thickness (0.21 +/- 0.08 vs. 0.16 +/- 0.05; P = .12), average intimal thickness (0.12 +/- 0.06 vs. 0.08 +/- 0.03; P = .08), marginally significant reduction in percent area stenosis (33 +/- 15% vs. 21 +/- 9%; P = .06) and marked neointima suppression in areas immediately adjacent to 45Ca wrap remnants. Medial necrosis (P = .003), however, was observed slightly more for 45Ca-treated carotid arteries versus control arteries. CONCLUSION: Low-dose 45Ca beta-radiation labeled onto a polyurethane membrane appears to inhibit NIH in an animal model. PMID- 15275631 TI - Radiation-induced coronary artery disease in Hodgkin's disease. AB - PURPOSE: Secondary malignant tumours and cardiovascular complications are of great importance among the late complications after treatment for Hodgkin's disease (HD) because they may significantly reduce the patients' life expectancy. We report on coronary artery disease (CAD) after treatment for HD. METHODS AND MATERIALS: We present the case of two female patients with HD who received mediastinal irradiation after which complete remission was achieved. In 12 and 19 years after the onset of the disease, control examinations revealed ischaemic heart disease, which was confirmed by coronary arteriography and solved by stent implantation in one of the patients and bypass surgery in the other one. CONCLUSIONS: Ischaemic heart disease was due to early CAD, which was associated with mediastinal irradiation accompanied by hypothyroidism, hyperlipidaemia and, possibly, early menopause, all posing an increased risk for the cardiologic disease. Our cases may serve as a warning example to carefully plan the management (low dose, involved field irradiation) of newly diagnosed patients as well as the cardiologic follow-up of patients with HD. PMID- 15275632 TI - Peroxisome proliferator-activated receptor gamma; Its role in atherosclerosis and restenosis. AB - Cellular proliferation and migration are fundamental processes that contribute to the injury response in major blood vessels. The resultant pathologies are atherosclerosis and restenosis. As we begin to understand the cellular changes associated with vascular injury, it is critical to determine whether the inhibition of growth and movement of cells in the vasculature could serve as a novel therapeutic strategy to prevent atherosclerosis and restenosis. PMID- 15275633 TI - Noninvasive visualisation of coronary atherosclerosis with multislice computed tomography. AB - Computed Tomography (Electron Beam Tomography: EBCT and multislice computed tomography: MSCT) have recently emerged as non-invasive diagnostic modalities that can quantify coronary calcium which is not only as indicator of coronary risk, but also permits assessment of the coronary lumen. The 16-slice MS-CT, the most recent CT-scanner has a very high resolution, which allows non-invasive assessment of coronary plaques. This has led to a stimulus for further research to assess the role of MSCT coronary plaque imaging in the identification of high risk coronary plaques. PMID- 15275634 TI - Plasmacytoid DCs and cancer: a new role for an enigmatic cell. PMID- 15275636 TI - Large-scale screening for genes involved in T-cell signaling: do we know all the players now? PMID- 15275637 TI - Signaling crosstalk between NF-kappaB and JNK. PMID- 15275638 TI - Is immunosenescence infectious? PMID- 15275639 TI - Concurrency in leukocyte vascular recognition: developing the tools for a predictive computer model. PMID- 15275640 TI - Tissue targeting of T cells by DCs and microenvironments. PMID- 15275641 TI - Epstein-Barr virus LMP2A: regulating cellular ubiquitination processes for maintenance of viral latency? PMID- 15275642 TI - The immune response under stress: the role of HSP-derived peptides. PMID- 15275643 TI - SOCS: role in inflammation, allergy and homeostasis. PMID- 15275644 TI - Extravascular T-cell recruitment requires initiation begun by Valpha14+ NKT cells and B-1 B cells. PMID- 15275645 TI - uPA and uPAR in fibrinolysis, immunity and pathology. PMID- 15275646 TI - Daily and annual variations of free fatty acid, glycerol and leptin plasma concentrations in goats (Capra hircus) under different photoperiods. AB - The aim of the study was to differentiate the impact of lighting conditions and feeding times on the regulation of lipid metabolism of goats under different photoperiods throughout the year. Seven Finnish landrace goats were kept under artificial lighting that simulated the annual changes of photoperiod at 60 degrees N (the longest light period 18 h, the shortest 6 h). Ambient temperature and feeding regime were kept constant. Blood samples were collected six times a year at 2-h intervals for 2 days, first in light/dark (LD) conditions and then after 3 days in constant darkness (DD). Significant daily variations were detected in the concentrations of plasma free fatty acids (FFA) and glycerol throughout the year. The nocturnal decrease and morning rise of FFA levels were related to the photoperiod, while the trough levels of glycerol were associated with the concentrate meal times. In DD conditions, FFA and glycerol rhythms were unstable. A significant seasonal variation was detected in the overall FFA and glycerol levels suggesting decreased lipogenesis in winter, increased lipolysis in spring and high lipogenesis in summer and fall. There was no significant daily rhythm in serum leptin levels, nor did the profiles in LD and DD conditions differ. The leptin level was slightly lower in early fall than in the other seasons, paralleling a small decrease of body mass in the goats after the grazing season. The daily or annual variations of FFA and glycerol levels were not clearly related to leptin concentrations. The results suggest that lipid metabolism of goats is regulated by light even in constant temperature and feeding conditions; however, no significant contribution of leptin levels could be shown. PMID- 15275647 TI - Effects of acute temperature changes on aerial and aquatic gas exchange, pulmonary ventilation and blood gas status in the South American lungfish, Lepidosiren paradoxa. AB - Lungfish (Dipnoi) are probably sister group relative to all land vertebrates (Tetrapoda). The South American lungfish, Lepidosiren paradoxa, depends markedly on pulmonary gas exchange. In this context, we report on temperature effects on aquatic and pulmonary respiration, ventilation and blood gases at 15, 25 and 35 degrees C. Lung ventilation increased from 0.5 (15 degrees C) to 8.1 ml BTPS kg( 1) min(-1) (35 degrees C), while pulmonary O(2)-uptake increased from 0.06 (15 degrees C) to 0.73 ml STPD kg(-1) min(-1) (35 degrees C). Meanwhile aquatic O(2) uptake remained about the same ( approximately 0.01 ml STPD kg(-1) min(-1)) at all temperatures. Concomitantly, the pulmonary gas exchange ratio (R(E)) rose from 0.11 (15 degrees C) to 0.62 (35 degrees C), because a larger fraction of total CO(2) output became eliminated by the lung. Accordingly, PaCO(2) rose from 13 (15 degrees C) to 37 mm Hg (35 degrees C), leading to a significant decrease of pHa at higher temperature (pHa=7.58-15 degrees C; 7.33-35 degrees C). The acid base status of L. paradoxa was characterized by a generally low pH (7.4-7.5), high bicarbonate level (20-25 mM) and PaO(2) ( approximately 80 mm Hg). The increased dependence on the lung at higher temperature parallels data for amphibians. Further, the effects of bimodal gas exchange on temperature-dependent acid-base regulation closely resemble those of anuran amphibians. PMID- 15275648 TI - Impact of gender on basal and insulin-like growth factor I-regulated nitric oxide synthase activity in adult rat left ventricular myocytes. AB - Cardiovascular morbidity and mortality are far less in pre-menopausal women compared to age-matched men. Ovarian hormones are believed to be mainly responsible for this "female advantage" in cardiovascular function although the underlying mechanism has not been fully elucidated. A gender difference exists in vascular nitric oxide (NO) synthesis, which may play a key role in ventricular function and cardiac remodeling. This study was designed to compare NO production, basal NO synthase (NOS) expression and activity, as well as insulin like growth factor I (IGF-1)-induced response on NOS activity in left ventricular myocytes from age-matched adult male and female Sprague-Dawley rats. NO production and protein expression of NOS, IGF-1 receptor (IGF-1R) and IGF binding protein-3 (IGFBP-3) were measured by Griess assay and Western blot analysis, respectively. NOS activity was evaluated by conversion of (3)H-arginine to (3)H citrulline. Basal NO production, endothelial NOS expression and NOS activity were both significantly higher in female left ventricular myocytes than their male counterparts. However, protein expression of inducible and neuronal NOS as well as IGFBP-3 was comparable between the two genders. IGF-1R expression was less in female than male group. IGF-1 (10(-10)-10(-6) m) induced a concentration dependent inhibition of NOS activity in male myocytes with a maximal inhibition of 22.2%. However, the IGF-1-induced inhibition in NOS activity was not present in left ventricular myocytes from female rats. These data revealed a gender difference in myocardial basal NO levels, endothelial NOS expression, basal NOS activity and IGF-1-induced inhibition on NOS activity, which may contribute to the gender-related difference of cardiac function. PMID- 15275649 TI - DNA modification in chick heart and cerebrum. AB - Heart muscle cells and cerebral neurons are known to lose the ability to proliferate and are called terminally differentiated cells. They are generated in appropriate numbers during embryogenesis and retained throughout adult life without turnover. We are interested in such a long-lived DNA. We isolated DNA from chick heart and cerebrum and compared it with DNA from other organs after incubation with DNase I. Single-strand breaks were assessed using a reaction system composed of DNA and Escherichia coli DNA polymerase. The DNA of both organs was relatively resistant to DNase I, and DNA modification occurred during embryogenesis. CIMS (chemical ionization mass spectrometry) indicated that the molecular mass of the deoxynucleoside of both DNAs was larger than that of the corresponding canonical deoxyribonucleoside by m/z 28 (or 30 for the protonated form). The difference between these deoxynucleosides is based on a difference in sugar constituents. Cerebral deoxynucleotides were analyzed by (13)C NMR. An extra signal near 173 ppm was observed, which was assigned to the amide carbonyl. We propose a model of the deoxynucleoside where a carbonyl residue exists between the base and the 2-deoxyribose moiety of the canonical deoxyribonucleoside. PMID- 15275650 TI - The ontogeny of physiological response to temperature in early stage spiny lobster (Jasus edwardsii) larvae. AB - The physiological response to temperature, in terms of oxygen consumption, nitrogen excretion and feed intake was examined in Jasus edwardsii larvae at mid stages I-III. From stage I to stage III, the mass-specific oxygen consumption increased in a sigmoid pattern over the temperature range of 10-22 degrees C. The Q(10) value declined significantly from 14-18 to 18-22 degrees C range, indicating a reduced temperature dependence of larval metabolism at higher temperatures. At all stages, feed intake increased with increasing temperature but reached a plateau at the higher temperatures for stages I and II larvae. In contrast, nitrogen excretion increased linearly over this temperature range for all larval stages. Therefore, higher temperatures ( approximately 22 degrees C) may cause an energetic imbalance and reduce growth potential in early stage larvae. While the convection requirement index (quotient of feed intake and oxygen consumption) indicated an equivalent metabolic feeding efficiency from 14 to 22 degrees C, a consistent decline of the O/N ratio above 16-18 degrees C from stage I to stage III suggested that exposure to elevated temperatures may result in an increase in the amount of protein being diverted from growth to catabolic processes. Based on these results, a temperature of 18 degrees C is recommended for the culture of early stage J. edwardsii larvae. PMID- 15275651 TI - The effect of captivity and diet on KLH isoform ratios in Megathura crenulata. AB - Aquaculture of the giant keyhole limpet, Megathura crenulata, may provide a reliable long-term supply of keyhole limpet hemocyanin (KLH) for many promising biomedical applications. However, previous studies have reported a complete loss of the KLH1 isoform under certain cultivation conditions. We examined whether captivity per se and diet caused a significant change in the isoform profile of M. crenulata. Although there was a trend toward a decreasing percentage of KLH1 in some animals, in general isoform profiles were not significantly affected by captivity or dietary limitations. Further, the percentage of KLH1 significantly increased for limpets with previously low levels of KLH1 when fed a supplemental mixed diet. Our results indicate that normal isoform profiles can be maintained in limpets held in captivity even when fed insufficient diets, and that these conditions do not cause a complete loss of either KLH isoform. Notably, the enhancement of abnormally low levels of KLH1 suggests that variability in isoform profiles could potentially be minimized through diet. While there is a need for further research on the factors responsible for the variability of KLH, overall, these results support the premise that culture of M. crenulata may provide a sustainable source of this biomedically important product. PMID- 15275652 TI - Immunohistochemical distribution of neuropeptide Y in the mesencephalon and rhombencephalon of carp, Cyprinus carpio L. (Cyprinidae: Teleostei). AB - The localization of neuropeptide Y (NPY)-immunoreactive elements was investigated in the mesencephalon and rhombencephalon of carp, Cyprinus carpio, by using antisera raised against porcine NPY and the immunoperoxidase technique. Concurrently, to identify the distribution of NPY-immunoreactivity, we developed an atlas of the studied areas based on Nissl-stained sections. The NPY immunoreactive (NPY-ir) elements were located in many zones of the mesencephalon and rhombencephalon. In the mesencephalon, positive fibers were the most abundant elements while neurons were scarce. The rhombencephalon rostral part was characterized by a low to moderate fiber density, distributed in the ventro medial and ventro-lateral region. Differently the caudal part of the rhombencephalon exhibited several NPY-ir elements. In particular, a high density of immunoreactivity was located in the gustatory area at the level of the nucleus (n.) originis nervi glossopharyngei, in the n. nervi vagi, and in the vagal lobe. The latter can be considered a valid neuroanatomical model for the study of gustatory signal processing in vertebrates. Our results regarding the primary gustatory centers give neuroanatomical support to the view that NPY may act as a neurotransmitter and/or a neuromodulator in a wide neural network for feeding behavior control. PMID- 15275653 TI - Blood profile of pigeons (Columba livia) during growth and breeding. AB - Changes in the blood profile of domestic pigeons (Columba livia) were studied during growth and breeding cycle. Counts of erythrocytes and leucocytes, and values of mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH), blood volume, plasma volume (BV), width of erythrocytes, and length, width and volume of erythrocyte nuclei of squabs almost reached adult values by the 4th week of age. During courtship and mating, while the level of plasma glucose increased, those of albumin, potassium, cholesterol, calcium and uric acid decreased. At nest-building, plasma albumin and plasma calcium increased significantly. The initial phase of incubation showed an elevation in plasma calcium and a decline in cholesterol and sodium, whereas mid-phase of incubation indicated a marked rise in cholesterol and uric acid. Terminal phase of incubation had significantly low plasma protein level. During feeding and brooding period, a significant rise in sodium, protein and glucose levels and a fall in calcium were observed. Following egg-laying, there was a significant rise in calcium and a drop in protein, haemoglobin, cholesterol, sodium and MCH values. Concomitant with the phenomenal rate of growth of squabs, their haematological indices neared adult values by the 4th week of age and during breeding activity significant changes in blood values occurred. PMID- 15275654 TI - Pacemaker activity in hydra is modulated by glycine receptor ligands. AB - In the mammalian central nervous system, the neurotransmitter, glycine, acts both on an inhibitory, strychnine-sensitive receptor (GlyR) and an excitatory, strychnine-insensitive site at the NMDA receptor. Here we present electrophysiological evidence that the strychnine-sensitive glycine agonists, glycine and taurine, and the antagonist, strychnine, affect the endodermal rhythmic potential (RP) system and that the ectodermal contraction burst (CB) pacemaker system is modulated by glycine and strychnine in hydra. The RP and CB pacemaker systems are responsible for the respective elongation and contraction of hydra's body column. Activity of the CB system, quantified by the rate of contraction bursts (CBs), the number of pulses per contraction burst (P/CB), and the duration of bursts, was decreased by glycine. Glycine, coadministered with the strychnine-insensitive glycine site blocker, indole-2-carboxylic acid (I2CA), decreased RPs but not CBs or P/CB. The effect was mimicked by taurine. Strychnine increased the duration of RP production, and decreased CB duration. The effect of glycine with I2CA was counteracted by strychnine. The results support the idea that a vertebrate-like GlyR may be involved in modulating activity of the endodermal RP system and suggest that a glycine site on an NMDA receptor may be involved in the CB system. PMID- 15275655 TI - Pericardial and pericardioperitoneal canal relationships to cardiac function in the white sturgeon (Acipenser transmontanus). AB - Sturgeons are primitive bony fishes and their hearts have structural features found in other primitive fishes. Sturgeons have a pericardioperitoneal canal (PPC), a one-way conduit into the peritoneum. A PPC also occurs in elasmobranchs (sharks and rays) and studies with that group demonstrate that pericardial pressure and pericardial fluid loss via the PPC affect stroke volume. A study of white sturgeon (Acipenser transmontanus) heart function was conducted to test for a comparable PPC and pericardial effects. White sturgeon-elasmobranch heart function similarities include biphasic ventricular filling, a comparable operational pericardial pressure (-0.03 kPa), and a strongly negative pressure ( 0.2 to -0.6 kPa) with complete pericardial fluid withdrawal. Differences include the white sturgeon's relatively smaller atrium and ventricle but a larger conus arteriosus. Although white sturgeon heart size is also smaller, its pericardial volume is disproportionately less (2.4 to 2.7 vs. 3.5 to 5.4 ml kg(-1) in elasmobranchs), meaning it has less scope for increasing stroke volume upon PPC fluid release. These differences may reflect the phylogenetic progression from the less complex operation of the elasmobranch heart, which lacks sympathetic innervation and has a mechanically mediated (PPC) stroke volume, to the condition in the more derived bony fishes which have sympathetic and parasympathetic regulation of both stroke volume and heart rate. PMID- 15275656 TI - Absorption of alpha-ketoglutarate by the gastrointestinal tract of pigs. AB - Only a small percentage of alpha-ketoglutarate (AKG) administered lumenally to pigs appears in the portal circulation. This has been attributed to mucosal metabolism, and possibly by limited absorption. Although transporters for di- and tricarboxylic acids, which includes the sodium-dependent transporter NaDC-1, have been detected in the small intestine, correlations with functional assays are lacking. Therefore, intact tissues from three regions of the small intestine, stomach, and colon of weaned pigs were used to measure rates of AKG absorption. Western analysis was used to detect NaDC-1 in the three regions of small intestine. Rates of AKG absorption were highest in the small intestine, lowest in the colon, and intermediate in the stomach. Immunoreactive NaDC-1 was detected in the small intestine and this coincided with a component of AKG absorption that was inhibited by AKG and succinate. In contrast, absorption of AKG was inhibitable by unlabeled AKG, but not succinate, in the stomach, and by neither in the colon. Feeding studies indicated that the amounts of AKG that might be included in practical diets for pigs would not (1) upregulate rates of AKG absorption or (2) exceed estimated capacities of the small intestine to absorb AKG. The present findings indicate that the efficacy of AKG as an alternative metabolic fuel for enterocytes to spare dietary amino acids is not limited by absorption. PMID- 15275657 TI - Temperature-dependent enhancement of cell proliferation and mRNA expression for type I collagen and HSP70 in primary cultured goldfish cells. AB - Goldfish (Carasius auratus) primary culture cells derived from caudal fin were incubated over a temperature range of 20-35 degrees C. The population doubling time of cells cultured at 20, 25, 30 and 35 degrees C were 34, 29, 17 and 14 h, respectively. Interestingly, cDNA-representational difference analysis revealed type I collagen alpha chain (colalpha(I)) as a candidate for a warm temperature specific gene. mRNA levels of colalpha(I) increased with an increase of incubation temperature and days of culture. Furthermore, the cell growth rate and colalpha(I) mRNA levels were rapidly changed following temperature shifts. To examine the effects of culture temperature shift on the cellular physiological states, mRNA levels of HSP70 were additionally investigated. HSP70 mRNA levels in the cells cultured at 30 and 35 degrees C were again 2-3 times higher than those at 20 and 25 degrees C. When the culture temperature was shifted from 20 to 35 degrees C, HSP70 mRNA levels were rapidly increased within 1 h. Subsequently, mRNA levels of the 35 degrees C-treated cells decreased, but remained doubled compared with those of the 20 degrees C-treated cells, even 4 h following the temperature shift. When the culture temperature was lowered from 35 to 20 degrees C, HSP70 mRNA levels decreased to about 70% of the original levels in 4 h. These results indicate that goldfish cells cultured at different temperatures easily develop temperature-associated steady physiological states within 4 h of temperature shifts. PMID- 15275658 TI - Differential gene expression during wing morph differentiation of the ectoparasitoid Melittobia digitata (Hym., Eulophidae). AB - Melittobia digitata is an ectoparasitoid of solitary bees and wasps that displays a trade-off between reproduction and dispersion through the development of two wing morphs (long and short wing morphs (LWM and SWM)). The morph differentiation of this species is an exceptional adaptation to maximize host exploitation and habitat colonization, and an understanding of the mechanisms underlying this developmental process will shed light on how nutrients or environmental elicitors alter regulatory pathways leading to physiological and metabolic changes resulting in such drastic developmental rearrangements. Here we describe the differential gene expression between SWM and LWM larvae of M. digitata in order to unravel the molecular mechanisms controlling the morph differentiation in this minute parasitoid and pinpoint the pathways involved in the regulation of this developmental process. The suppression subtractive hybridization (SSH) methodology was used to isolate differentially expressed genes using mRNA populations collected soon after morph development commitment. Dot blot analysis of 384 clones from a forward SSH library identified approximately 200 differentially expressed clones, including those transcripts present in very low abundance. Further DNA sequence analysis of a sub-population of 42 clones revealed 31 putatively unique transcripts, from which 5 were further analyzed by Northern blot analysis and semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR). The complete cDNA of one of these transcripts, a putative metalloprotease, was fully sequenced and is described. The role of the putative differentially expressed genes during the wing morph differentiation of M. digitata is discussed. PMID- 15275659 TI - Temperature acclimation modulates the oxygen binding properties of the Atlantic cod (Gadus morhua L.) genotypes-HbI*1/1, HbI*1/2, and HbI*2/2-by changing the concentrations of their major hemoglobin components (results from growth studies at different temperatures). AB - The influence of long-term acclimation temperatures in Atlantic cod (Gadus morhua) was studied by growth experiments carried out over a total of 272 individuals. The attention focused on the structural and functional modulation of the five electrophoretically distinguishable genotypes of cod hemoglobin (HbI*1/1, HbI*1/2, HbI*2/2, HbI*1/2b, and HbI*2/2b) and on the correlation with body length/weight. The main results can be summarized as follows. (1) Acclimation to lower (4 and 8 degrees C) and higher (12 and 15 degrees C) temperatures favors the expression of, respectively, more anodic and more cathodic hemoglobin components. (2) The optimal O(2) transporting features are observed at 12 degrees C, as well as a saturation-dependent temperature dependence of O(2) binding, which furthermore is strongly dependent upon the acclimation background. (3) The optimal growth condition for the three main genotypes (HbI*1/1, HbI*1/2, and HbI*2/2) is associated with T=12 degrees C. The overall results are consistent with the idea that environmental temperatures constitute a primary factor in the aggregation of individuals physiologically more than genetically homogeneous. This is fully confirmed by careful statistical analysis carried out over a subset of individuals for which the full set of structural (isoelectric focusing), functional (O(2) binding), and growth data was available. PMID- 15275660 TI - Aposymbiotic Plesiastrea versipora continues to produce cell-signalling molecules that regulate the carbon metabolism of symbiotic algae. AB - The scleractinian coral Plesiastrea versipora produces cell signals that regulate the carbon metabolism of its symbiotic algae. Host release factor (HRF) stimulates the release of photosynthate, and photosynthesis inhibiting factor (PIF) partially inhibits carbon fixation in freshly isolated symbiotic algae. Naturally occurring aposymbiotic specimens of P. versipora are rare in Port Jackson, Australia, but one that was collected contained HRF and PIF. Artificially produced aposymbiotic corals of P. versipora that had been kept in the dark for up to 23 months continued to produce both HRF and PIF in the absence of photosynthetically active algae. Aposymbiotic P. versipora from which most of the tissue had been removed, regenerated when they were kept in the dark and fed; the regenerated tissue also contained HRF and PIF. These results suggest that the presence of symbiotic algae is not required for the production of HRF and PIF in P. versipora. We suggest that these cell signals may have evolved in response to symbiosis with Symbiodinium sp. but are now always expressed in the coral P. versipora. PMID- 15275661 TI - BAC end sequences and a physical map reveal transposable element content and clustering patterns in the genome of Magnaporthe grisea. AB - Transposable elements (TEs) are viewed as major contributors to the evolution of fungal genomes. Genomic resources such as BAC libraries are an underutilized resource for studying genome-wide TE distribution. Using the BAC end sequences and physical map that are available for the rice blast fungus, Magnaporthe grisea, we describe a likelihood ratio test designed to identify clustering of TEs in the genome. A significant variation in the distribution of three TEs, MAGGY, MGL, and Pot2 was observed among the fingerprint contigs of the physical map. We utilized a draft sequence of M. grisea chromosome 7 to validate our results and found a similar pattern of clustering. By examining individual BAC end sequences, we found evidence for 11 unique integrations of MAGGY or MGL into Pot2 but no evidence for the reciprocal integration of Pot2 into another TE. This suggests that: (a) the presence of Pot2 in the genome predates that of the other TEs, (b) Pot2 was less transpositionally active than other TEs, or (c) that MAGGY and MGL have integration site preference for Pot2. High transition/transversion mutation ratios as well as bias in transition site context was observed in MAGGY and MGL elements, but not in Pot2 elements. These features are consistent with the effects of a Repeat-Induced Point (RIP) mutation-like process occurring in MAGGY and MGL elements. This study illustrates the general utility of a physical map and BAC end sequences for the study of genome-wide repetitive DNA content and organization. PMID- 15275662 TI - The glyceraldehyde-3-phosphate dehydrogenase homologue is differentially regulated in phases of Paracoccidioides brasiliensis: molecular and phylogenetic analysis. AB - Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) plays important roles in various cellular processes. Here we report the sequence and analysis of a novel developmentally regulated gene and cDNA (Pbgadph), encoding a GAPDH homologue (PbGAPDH), of the pathogenic dimorphic fungus Paracoccidioides brasiliensis. We have analyzed the protein, the cDNA and genomic sequences to provide insights into the structure, function, and potential regulation of PbGAPDH. That Pbgapdh encodes PbGAPDH was demonstrated by micro-sequencing of the native protein homologue isolated from the fungus proteome. The deduced amino acid sequence of Pbgapdh showed identity to those of from other species (88-76%). Phylogenetic analysis indicated that GAPDH could be useful for the determination of evolutionary relationships. Expression of the Pbgapdh gene and the cognate protein were developmentally regulated in phases of P. brasiliensis, with a higher expression in the yeast parasitic phase and was induced during the transition from mycelium to yeast and decreased during the reverse process, transition from yeast to mycelium. PMID- 15275663 TI - Evidence for subdivision of the root-endophyte Phialocephala fortinii into cryptic species and recombination within species. AB - The genetic structure of the root-endophyte Phialocephala fortinii was analyzed in three study sites using 11 single-copy RFLP probes. A total of 541 strains isolated from surface-sterilized, fine roots (diameter 0.5-3 mm) of Norway spruce (Picea abies) were examined. The average gene diversity (H) was high in all three study sites. Cluster analysis showed that up to four well-separated clusters of multi-locus haplotypes were present within the sites. Significant population subdivision was detected among these clusters, suggesting that groups of multi locus haplotypes were reproductively isolated and that P. fortinii is a species complex composed of several cryptic species. This hypothesis was supported by ISSR-PCR which showed clusters consistent with those of the multi-locus haplotypes identified by RFLP analysis. In contrast, ITS sequence analysis did not allow to separate the species as clearly. The index of association (IA) did not deviate significantly from zero within any cryptic species, suggesting that recombination occurs within these species. Cryptic species occurred sympatrically. Thalli of two cryptic species were detected in the same 5-mm-long root segment in one instance. No significant differentiation was observed among populations of the same cryptic species in forest stands located approximately 5 km from each other. This finding is consistent with significant gene flow over this spatial scale. In addition, several isolates with both identical multi-locus haplotype and identical ISSR fingerprint were found at each study site indicating genotype flow or a recent common history between study sites. PMID- 15275664 TI - Photomorphogenesis in the hypogeous fungus Tuber borchii: isolation and characterization of Tbwc-1, the homologue of the blue-light photoreceptor of Neurospora crassa. AB - Truffles form a group of plant-symbiotic Ascomycetes whose hypogeous life cycle is poorly understood. Here we present initial evidence for the influence of light on Tuber borchii mycelial growth and the identification and cloning of a gene, Tbwc-1, homologous to a blue-light photoreceptor of Neurospora crassa. Blue-light irradiation of T. borchii colonies inhibits their apical growth. It also alters apical growth in N. crassa. In Neurospora, the response is controlled by a nuclear photoreceptor, NcWC-1 (White Collar-1), which consists of a sensor domain (LOV) and a transcriptional factor moiety. We isolated a gene (Tbwc-1) whose deduced amino acid sequence shows a high similarity and colinearity of domains with NcWC-1, except for the polyglutamine regions. As previously found in Neurospora, Tbwc-1 mRNA is under light control and its steady state level increases upon irradiation. In silico analysis of the TbWC-1 sensor domain (LOV) supports the hypothesis that TbWC-1 is a photoreceptor, while the absence of the two polyglutamine regions involved in transcriptional activation in Neurospora suggests that this function in Tuber could be lost. PMID- 15275665 TI - New information on the mechanism of forcible ascospore discharge from Ascobolus immersus. AB - Many ascomycete fungi spurt their spores from asci pressurized by osmosis. This paper explores the details of this process in the coprophilous species Ascobolus immersus, through a combination of biomechanical and biochemical experiments, and mathematical modeling. A. immersus forms large asci that expel 8 spores as a single, mucilage-embedded projectile. Measurements of ascus turgor using a microprobe attached to a strain gauge showed a pressure of 0.3 MPa or 3 atm. Analysis of ascus sap using GC/MS identified glycerol as a major osmolyte, accounting for 0.1 MPa of the osmotic pressure within the ascus sap. A mathematical model indicated that a pressure of 0.2 MPa would be sufficient to propel the cluster of ascospores over the distance measured in previous studies. The difference between the measured and predicted pressures is ascribed to loss of pressure as the spores are forced through the tip of the open ascus. PMID- 15275666 TI - Penicillium chrysogenum Pex5p mediates differential sorting of PTS1 proteins to microbodies of the methylotrophic yeast Hansenula polymorpha. AB - We have isolated the Penicillium chrysogenum pex5 gene encoding the receptor for microbody matrix proteins containing a type 1 peroxisomal targeting signal (PTS1). Pc-pex5 contains 2 introns and encodes a protein of approximately 75 kDa. P. chrysogenum pex5 disruptants appear to be highly unstable, show poor growth, and are unable to sporulate asexually. Furthermore, pex5 cells mislocalize a fluorescent PTS1 reporter protein to the cytosol. Pc-pex5 was expressed in a PEX5 null mutant of the yeast Hansenula polymorpha. Detailed analysis demonstrated that the PTS1 proteins dihydroxyacetone synthase and catalase were almost fully imported into microbodies. Surprisingly, alcohol oxidase, which also depends on Pex5p for import into microbodies, remained mainly in the cytosol. Thus, P. chrysogenum Pex5p has a different specificity of cargo recognition than its H. polymorpha counterpart. This was also suggested by the observation that Pc-Pex5p sorted a reporter protein fused to various functional PTS1 signals with different efficiencies. PMID- 15275667 TI - The influence of genotypic variation on metabolite diversity in populations of two endophytic fungal species. AB - The relationship between metabolite production and genotypic diversity in two endophytic fungi was investigated. We selected populations of Cylindrocarpon destructans and Heliscus lugdunensis from the roots of a single tree. A total of 49 isolates of both species were selected and classified by simple genotypic tests (random amplified polymorphic DNA analysis and rDNA-ITS sequencing). In a blind test, the ability of these fungi to produce natural products was tested by ethyl acetate extraction of hyphae and culture filtrates, followed by high performance liquid chromatography analysis (HPLC). A positive relationship was found between genotype classification and the pattern of natural products produced by a given isolate. To test the robustness of this correlation, a discriminate selection procedure was carried out by collecting fungal isolates from a second site and selecting a sub-set of the population, on the basis of genotypic variability. This sub-set of fungal isolates produced greater numbers of unique metabolites than those selected indiscriminately. PMID- 15275668 TI - Addressing multiple behavioral health risks in primary care. Broadening the focus of health behavior change research and practice. PMID- 15275669 TI - Addressing multiple behavioral risk factors in primary care. A synthesis of current knowledge and stakeholder dialogue sessions. AB - BACKGROUND: Addressing behavioral risk factors in primary care has become a pressing concern due to the increasing burden of behavioral risk factors on disease, healthcare costs, and public health. Risk factors considered include smoking, risky drinking, sedentary lifestyle, and unhealthy diet-singly or in combination. The already burdened primary care system needs a practical approach to efficiently and effectively address any combination of multiple risk factors. Multiple perspectives and broad insight are urgently needed to gain a deeper understanding of the interacting scientific, systems, and policy issues associated with multiple risk factor interventions (MRFIs). PURPOSE: This paper synthesizes findings from literature reviews, epidemiologic analyses, and structured interactive dialogue sessions, and includes a set of recommendations designed to stimulate further action. METHODS: Several papers were produced to document current knowledge, research evidence, and salient issues related to multiple risk factor assessment and intervention. Structured interactive dialogue sessions were then conducted with clinician, health system, and health policy leaders regarding what advantage or energy would be liberated by a multiple risk factor approach (rather than separate single risk factor approaches), and how to build a policy framework or constituency for MRFIs. This information is synthesized in this paper. RESULTS: There is a clear need to address MRFIs among multiple stakeholders, including patients, purchasers, payers, clinicians, health system leaders, and policy-level stakeholders. MRFIs need to bring with them a compelling value proposition for all stakeholders, and a vision of practical and systematic ways to make it a reality in already-pressed primary care practices. Involving stakeholders in dialogue aimed at helping them see the world through each other's eyes helps overcome discouragement and generates energy for jointly designing new approaches. Recommendations for further action include the creation of multistakeholder dialogue, creation of a policy agenda, development of a translation or integration agenda that connects researchers and practitioners in a two-way exchange, initiation of a series of demonstration projects around MRFIs, and support for research on multiple (rather than only single) risk factor interventions. CONCLUSIONS: The need to address multiple behavioral risk factors in primary care is increasingly urgent. Whereas stakeholders by themselves may be willing to address multiple risk factors, they agree that it can only be done successfully with a collaborative approach. Findings based on evidence reviews, hypotheses generation, and stakeholder dialogue provide guidance for appropriate further action that, based on what is known already, can be initiated right away. PMID- 15275670 TI - Prevalence of multiple chronic disease risk factors. 2001 National Health Interview Survey. AB - BACKGROUND: Four common factors--cigarette smoking, risky drinking of alcoholic beverages, physical inactivity, and overweight--contribute substantially to chronic disease prevalence. METHODS: We used data from the 2001 National Health Interview Survey to provide an up-to-date picture of multiple risk factor prevalence and clustering in the U.S. population. We conducted a multinomial logit analysis to examine the independent association between each covariate and the dependent ordinal risk factor variable with three levels (none or one risk factor, two risk factors, and three or four risk factors). RESULTS: Seventeen percent of the sample of 29,183 subjects had three or more risk factors. For the entire sample, the mean number of risk factors was 1.68 (95% confidence interval [CI]=1.66-1.70). Many demographic and health factors were significantly associated with the mean number of risk factors including gender, age, ethnic/racial categories, education, martial status, presence of chronic diseases, level of mental distress, country of birth, and presence and type of health insurance. Using the risk factor score as the ordinal dependent variable, adjusted odds for having a risk score of three or four versus zero or one were as follows: men aged <65, 2.49 (95% CI=2.29-2.72); education attainment of high school graduate or less, 3.24 (95% CI=2.86-3.67); and individuals with high levels of mental distress, 2.06 (95% CI=1.65-2.58). CONCLUSIONS: Our analyses confirm earlier reports of the high prevalence of multiple, clustered behavioral risk factors and underline the challenge this presents for primary care and public health systems. PMID- 15275671 TI - Meeting recommendations for multiple healthy lifestyle factors. Prevalence, clustering, and predictors among adolescent, adult, and senior health plan members. AB - BACKGROUND: Whereas much is known about single lifestyle-related health risk factor prevalence and covariates, more research is needed to elucidate the interactions among multiple healthy lifestyle factors and variables that may predict adherence to these factors. Such data may guide both clinical and health policy decision making and person-centered approaches to population health improvement. METHODS: We document the prevalence and cluster patterns of multiple healthy lifestyle factors among a random sample of adolescents (n =616), adults (n =585), and seniors (n =685) from a large Midwestern health plan. Modifiable, lifestyle-related health factors assessed included physical activity, nonsmoking, high-quality diet, and healthy weight for all subjects; adults and seniors were also asked about their alcohol consumption. Second, we sought to identify characteristics associated with the likelihood of meeting recommendations for healthy lifestyle factors. The healthy lifestyle factors sum score was categorized into three levels, that is, 0 to 2, 3, or 4 to 5 healthy lifestyle factors (4 for adolescents), and we used ordinal logistic regression to estimate the odds of meeting each of these criteria from several demographic characteristics and disease states. RESULTS: Overall, only 14.5% of adolescent, adult, and senior health plan members meet recommended guidelines for four common healthy lifestyle factors. Only 10.8% of adults and 12.8% of seniors met all five behavior-related factors. For adolescents, only being nondepressed was associated with an increased likelihood to be in adherence to multiple healthy lifestyle factors (odds ratio [OR]=2.15; p <0.05). For adults, being in the 50- to 64-year old cohort (OR=1.46, p<0.05), having a college degree (OR=1.65; p <0.05), and having no chronic disease (OR=1.92; p <0.05) were all associated with an increased likelihood to be in adherence to multiple healthy lifestyle factors. For seniors, having a college degree (OR=1.61; p <0.05), was the only variable associated with an increased likelihood to be in adherence to multiple healthy lifestyle factors. CONCLUSIONS: A small proportion of health plan members meet multiple recommended healthy lifestyle guidelines at once. This analysis identifies population subgroups of specific interest and importance based on adherence to multiple healthy lifestyle factors, and predictors for increased likelihood to be in adherence to multiple healthy lifestyle factors. It presents a potentially useful summary measure based on person-centered measures of healthy lifestyle factors. Clinicians may derive meaningful information from analyses that address adherence to multiple healthy lifestyle factors. Health systems administrators may use this information to influence health policy and resource allocation decisions. Further studies are needed to assess the usefulness of this comprehensive lifestyle-related health measure as a metric of progress toward public health goals, or as a clinical metric that conveys information on future health status and directs interventions at the individual level. PMID- 15275672 TI - Physician screening for multiple behavioral health risk factors. AB - BACKGROUND: Screening rates in primary care for single behavioral health risk factors are widely documented. However, such risk factors cluster in individuals and populations. This article examines the number and types of behavioral risk factors that U.S. adults reported, and reported having been screened for in their last routine medical checkup. METHODS: The sample consisted of 16,818 adults from the 1998 National Health Interview Survey who reported having a routine checkup in the past year. Respondents completed questions regarding four behavioral risk factors (physical inactivity, overweight, cigarette smoking, risky drinking), and provider screening for behaviors related to these risk factors. RESULTS: Half of the sample (52.0%) reported having two or more of the four risk factors, and more than half (59.4%) were screened for two or more risk behaviors during their last routine checkup, although 28.6% reported being screened for none of them. Respondents reporting at least one risk factor were screened for an average of 57.7% of their own risk factors. Women, adults with lower levels of income and education, and those aged 65 and older, reported being screened for fewer of their risk factors. CONCLUSIONS: While guidelines for risk factor screening and intervention typically focus on single behavioral risk factors, most primary care patients present with, and are screened for, more than one. Behavioral risk factor screening tools and interventions must be expanded to cover multiple risks. Additionally, efforts are needed to reduce the substantial missed opportunities for screening, and to eliminate demographic disparities in screening practices and accuracy. PMID- 15275673 TI - Assessing multiple risk behaviors in primary care. Screening issues and related concepts. AB - The concept of behavioral risk refers to health behaviors that increase the likelihood of a variety of illness conditions. With increased scientific research, it has become clear that this concept is useful in understanding the linkage between behavior and health. This paper reviews scientific, conceptual, and practical issues related to the identification of health risk behaviors in primary care. It includes both a literature review and an analysis of the feasibility of screening and health risk appraisal from a public health perspective, giving special attention to four behavioral risk factors: cigarette smoking, alcohol misuse, physical inactivity, and unhealthy diet. The review indicates that there are a wide variety of acceptable screening tests that can be used for population screening programs, and a large number of health risk appraisal instruments to employ in medical and work settings where preventive health services are available. Given the variety of available assessment procedures, the choice of a given instrument will depend on the target population, the purpose of the program, the time available for assessment, and a number of other practical considerations, such as cost. Multiple risk factor screening is feasible, but there is no single instrument or procedure that is optimal for all risk factors or populations. Based on the results of this review, the specific test or combination of tests is less important than the use of screening to make both patients and healthcare providers more aware of the critical importance of monitoring behavioral risk factors on a routine basis. We conclude that while further research and development work needs to be done, sufficient progress has been made to warrant a more ambitious effort that would bring behavioral risk factor screening into the mainstream of preventive medicine and public health. PMID- 15275674 TI - Coronary heart disease multiple risk factor reduction. Providers' perspectives. AB - BACKGROUND: Although primary care physicians understand the importance of preventive services for patients with multiple risk factors (MRF) for coronary heart disease, physician intervention is limited. This study investigated (1) physicians' views of challenges faced in managing patients with MRF; (2) the counseling and management methods they utilize; and (3) possible strategies to enhance MRF intervention in the primary care setting. METHODS: Two focus groups were conducted with primary care physicians from varying settings to gain insight into these issues noted above. Each group was co-facilitated by a physician and a behavioral scientist using a previously developed semistructured interview guide. The group discussions were tape recorded and subsequently transcribed. Transcripts were analyzed using the constant comparative method for analysis. RESULTS: Physicians are challenged by knowledge limitations (contribution of individual risk factors to overall risk); limited support (guidelines, materials, and staff); and logistic difficulties (organizational issues, time limitations). Their approach to MRF management tends to be highly individualized with an initial preference for lifestyle change interventions rather than prescription of medications with some qualifying circumstances. Physicians favored a serial rather than a parallel approach to MRF intervention, starting with behaviors that the patient perceives as a priority. Proposed solutions to current challenges emphasize physician education and the development of innovative approaches that include physician assistance and a team approach. CONCLUSIONS: Physicians are aware of and sensitive to the complexity of MRF management for their patients and themselves. However, future MRF interventions will require nonphysician staff involvement and increased systems support. PMID- 15275675 TI - Multiple behavioral risk factor interventions in primary care. Summary of research evidence. AB - BACKGROUND: An important barrier to the delivery of health behavior change interventions in primary care settings is the lack of an integrated screening and intervention approach that can cut across multiple risk factors and help clinicians and patients to address these risks in an efficient and productive manner. METHODS: We review the evidence for interventions that separately address lack of physical activity, an unhealthy diet, obesity, cigarette smoking, and risky/harmful alcohol use, and evidence for interventions that address multiple behavioral risks drawn primarily from the cardiovascular and diabetes literature. RESULTS: There is evidence for the efficacy of interventions to reduce smoking and risky/harmful alcohol use in unselected patients, and evidence for the efficacy of medium- to high-intensity dietary counseling by specially trained clinicians in high-risk patients. There is fair to good evidence for moderate, sustained weight loss in obese patients receiving high-intensity counseling, but insufficient evidence regarding weight loss interventions in nonobese adults. Evidence for the efficacy of physical activity interventions is limited. Large gaps remain in our knowledge about the efficacy of interventions to address multiple behavioral risk factors in primary care. CONCLUSIONS: We derive several principles and strategies for delivering behavioral risk factor interventions in primary care from the research literature. These principles can be linked to the "5A's" construct (assess, advise, agree, assist, and arrange-follow up) to provide a unifying conceptual framework for describing, delivering, and evaluating health behavioral counseling interventions in primary healthcare settings. We also provide recommendations for future research. PMID- 15275676 TI - Interactive behavior change technology. A partial solution to the competing demands of primary care. AB - BACKGROUND: Primary care practices are faced with the challenge of having too much to do in too little time. As a result, behavioral counseling is often overlooked, especially for patients with multiple health behaviors in need of change. METHODS: This paper describes recent examples of the application of interactive behavior change technologies (IBCTs) to deliver health behavior change counseling before, during, and after the office visit to inform and enhance patient-clinician interactions around these issues. The 5A's framework (assess, advise, agree, assist, arrange follow-up) is used to consider how interactive technology can be used to implement behavior change counseling more consistently. RESULTS: A variety of IBCTs, including the Internet, clinic-based CD-ROMs, and interactive voice-response telephone calls have been shown to be feasible and potentially valuable adjuncts to clinic-based behavioral counseling. These technologies can both increase the effectiveness of behavioral counseling and extend the reach of these services to patients with barriers to face-to-face interactions. CONCLUSIONS: If appropriately developed with the context of primary care in mind and integrated as part of a systems approach to intervention, IBCT can be a feasible and appropriate aid for primary care. Recommendations are made for the types of IBCT aids and research that are needed to realize this potential. PMID- 15275677 TI - Translating what we have learned into practice. Principles and hypotheses for interventions addressing multiple behaviors in primary care. AB - BACKGROUND: The evidence base regarding what works in practice for helping patients change multiple risk behaviors is less developed than is the more basic literature on behavior change. Still, there is enough consistency of findings to present testable hypotheses for clinicians and administrators to evaluate and guide practice until more definitive evidence is available. METHODS: The behavior change principles known as the 5A's outline a sequence of support activities (assess, advise, agree, assist, arrange) that are effective for helping patients to change various health behaviors. These same principles also apply at the clinic level for designing activities to support behavior change. RESULTS: Successful practices promoting sustainable changes in multiple behaviors are patient centered, tailored, proactive, population based, culturally proficient, multilevel, and ongoing. Often a stepped-care model can be used to provide increasingly intensive (and costly) interventions for patients who are not successful at earlier intervention levels. CONCLUSIONS: Contextual factors are influential in determining success at both the patient and the office practice level. Therefore, greater attention should be paid to creating supportive family, healthcare system, and community resources and policies. We enumerate 15 hypotheses to be tested for improving patient-clinician interactions and for medical office change. PMID- 15275678 TI - Multiple risk factors interventions. Are we up to the challenge? PMID- 15275679 TI - Beyond efficacy testing redux. PMID- 15275680 TI - Toward a public policy agenda for addressing multiple health risk behaviors in primary care. PMID- 15275682 TI - Nerve tumors of the hand and upper extremity. AB - Tumors of peripheral nerve origin are usually slow growing and minimally symptomatic, making differentiation from other soft tissue neoplasms difficult. Yet failure to recognize a nerve tumor may result in irreversible loss of neurologic function. This article provides current information on the history, pathologic identification, and treatment of upper extremity nerve tumors. Other neoplastic and tumor-like lesions that occur within the peripheral nerve are also considered. PMID- 15275683 TI - Benign tumors of fibrous tissue and adipose tissue in the hand. AB - This article presents the current understanding of soft tissue hand tumors and the best options for treating them. The majority of soft tissue hand tumors are benign. Discussion includes hand tumors of fibrous and adipose tissue origin, determining the diagnostic and therapeutic pathways for these tumors, and controversial treatment issues. PMID- 15275684 TI - Ganglion cysts and other tumor related conditions of the hand and wrist. AB - Most regard ganglion, giant cell tumor of tendon sheath and epidermal inclusion cysts as tumor-like conditions as opposed to true neoplasms. Ganglion cysts are the most common lesion of the hand and wrist, accounting for 50% to 70% of all masses identified. The majority of ganglion cysts can be treated nonoperatively but when surgery is performed a low recurrence rate can be anticipated. Giant cell tumor of the tendon sheath hand epidermoid cysts are also common hand lesions that require surgical excision in most instances. Of the three, giant cell tumor of tendon sheath have the most notable recurrence rates. This article reviews the clinical presentations of these lesions as well as their proposed pathophysiology. PMID- 15275685 TI - Vascular tumors. AB - Proliferative conditions involving blood vessels and masses that mimic vascular tumors can present to hand surgeons. Patient age at presentation can range from neonates with congenital tumors to adults with malignancies, such as angiosarcoma. This article reviews three basic categories of vascular tumors in the upper extremity: benign, congenital,and malignant. PMID- 15275686 TI - Aneurysmal bone cyst and giant cell tumor of bone of the hand and distal radius. AB - Aneurysmal bone cyst and giant cell tumor of bone have historically been considered benign lesions that can behave in locally aggressive fashion. The clinical and pathologic findings share some common characteristics. When considering a radiographic differential diagnosis, these entities are usually considered together. There are some very distinct differences in potential aggressiveness and in response to treatment. This report summarizes current knowledge regarding these lesions when they occur in the hand and outlines current treatment methods. PMID- 15275687 TI - Pigmented skin lesions of the upper extremity. AB - This article discusses pigmented lesions of the upper extremities, including nevi, melanoma, and Merkel cell carcinoma. The diagnosis and work-up is emphasized, with attention also given to the techniques of sentinel node biopsy. An overview of the latest treatment strategies is provided. PMID- 15275688 TI - Non-melanotic skin tumors of the upper extremity. AB - Most non-melanotic skin cancers involving the upper extremities are squamous cell and basal cell carcinomas. Ultraviolet sun light, fair complexion, and advancing age are important risk factors. Because of their high prevalence, the hand surgeon must be familiar with the diagnosis and clinical management of non melanotic skin tumors. Early detection ultimately decreases the morbidity associated with the locally destructive nature of these tumors. Following diagnosis, the appropriate steps for management include wide surgical excision with pathologic conformation of normal margins and lymph node dissection for clinically positive nodes PMID- 15275689 TI - Bone-forming tumors of the upper extremity and Ewing's sarcoma. AB - This article presents an overview of bone-forming tumors that occur in the upper extremity. Osteoid osteoma, osteoblastoma, osteosarcoma, and Ewing's sarcoma are covered. Each tumor type is described, and suggestions are made for diagnostic workup and differential diagnosis. Locations in the upper extremity where each tumor typically occurs are given. Preferred treatment regimens and incidence of recurrence are also presented. PMID- 15275690 TI - Benign and malignant cartilage tumors of the hand. AB - Primary bone tumors of the hand and wrist are unusual but if present are frequently chondrogenic in origin. Enchondroma is the most common primary bone tumor of the hand and can be treated by curettage and bone grafting. Chondrosarcoma is rare and most likely the result of malignant degeneration of a pre-existing lesion. Wide en bloc resection is appropriate for chondrosarcoma and typically involves ray resection or partial hand amputation. PMID- 15275691 TI - Soft tissue sarcoma of the upper extremity. AB - Soft tissue sarcomas of the upper extremities are rare and hand surgeons typically encounter only one or two undiagnosed soft tissue sarcomas during their careers. It is incumbent on the physician to review repeatedly the characteristics of these tumors and remain suspicious, because these lesions typically are misdiagnosed and treatment is delayed. The most common soft tissue sarcomas of the upper extremity are the epithelioid sarcoma, synovial cell sarcoma, and malignant fibrous histiocytoma. Limb salvage surgery is the treatment of choice for soft tissue sarcomas to preserve upper extremity function. Following wide tumor resection, adjuvant therapies such as chemotherapy, external beam radiation therapy, and brachytherapy may lessen local recurrence rates, but their effect on overall survival remains unclear. PMID- 15275692 TI - The hand in metastatic disease and acral manifestations of paraneoplastic syndromes. AB - Metastatic tumors to the hand and wrist are rare, accounting for approximately 0.1% of all metastatic lesions to the skeleton. The biochemically mediated pathways of bone metastases, the location of the hand at the distal extremity, and the small amount of marrow in the bones of the hand and wrist account for the low prevalence of acrometastases. More rarely, hand dermatologic and soft tissue changes of paraneoplastic syndromes herald an occult malignancy. PMID- 15275694 TI - Founders of child neurology in Japan--Masataka Arima. PMID- 15275695 TI - The classification of cortical dysplasias through molecular genetics. AB - Recent genetic insight into the mechanisms of human brain malformation have allowed one to consider a classification of these disorders by the genetic disruption. In this article an attempt is made to classify human cortical dysplasias by the known genetic disruptions or insults that lead to them. The discussion of malformation is within the context of the embryologic processes that have thought to have gone awry. Human disorders of segmentation, cell proliferation, telencephalic cleavage, differentiation, and neuronal migration are discussed. As this is a rapidly changing area, the reader is encouraged to check online databases for updates on the genetic insights that have been gained since the publication of this article. PMID- 15275696 TI - Sphingolipidoses in Turkey. AB - During the last 5 years 2057 children under the age of 5 with various neurologic symptoms with the suspected diagnosis of lysosomal storage diseases were referred to our hospital from different universities and state hospitals. We were able to separate sphingolipidoses by lysosomal enzyme screening. A total of 300 patients (15%) with sphingolipidoses were diagnosed; there were deficiencies of arylsulfatase A [metachromatic leukodystrophy (MLD)] in 93 (31%), hexosaminidase [Sandhoff disease (SHD)] in 62 (20.7%), hexosaminidase A [Tay-Sachs disease (TSD)] in 15 (5%), beta-galactosidase (GM1 gangliosidosis) in 35 (11.7%), alpha galactosidase (Fabry disease) in one (0.3%) cerebroside beta-galactosidase (Krabbe disease) in 65 (21.7%) and glucosylceramidase (Gaucher disease) in 29 (9.6%). SHD (20.7%), MLD (31%) and Krabbe disease (21.7%) were common. Prenatal enzymatic diagnosis was made in 70 at risk pregnancies, 64 for TSD and SHD, three for MLD and three for GM1 gangliosidosis by using chorionic villus biopsy in 54, cord blood samples in 12 and cultured amniotic fluid cells in four. Seventeen fetuses were found to be affected. We have calculated the relative frequency and minimum incidence of sphingolipidoses in Turkey. The combined incidence of sphingolipidoses is 4.615 per 100,000 live births. The calculated incidences are 1.43, 0.95, 1, 0.23, 0.54, 0.45, 0.015 per 100,000 live births for MLD, SHD, Krabbe, Gaucher, TSD, GM1 gangliosidosis and Fabry diseases, respectively. The real incidence, which covers all subtypes of this group of diseases, should be greater than this number. The results suggested that, as a group, sphingolipidoses are relatively common and represent an important health problem in Turkey and some rare autosomal recessive diseases of Turkey are due to 'founder effect' created by consanguineous marriages. PMID- 15275697 TI - Analysis of childhood absence epilepsy using haplotype-based haplotype relative risk and transmission disequilibrium test. AB - The authors performed haplotype-based haplotype relative risk (HHRR) and transmission disequilibrium test (TDT) analysis of childhood absence epilepsy in 30 trios families, using gene typing technology based on microsatellite polymorphic marker. The five microsatellite DNA markers (D8S554, D8S1753, D8S534, D8S1100, D8S1783) used in the study are on chromosome 8q24. HHRR shows D8S554(4) (chi2 = 5.939, P < 0.05), D8S1100(3) (chi2 = 5.081, P < 0.05), D8S1783(6) (chi2 = 4.308, P < 0.05), TDT shows D8S554(4) (chi2 = 4.455, P < 0.05), D8S1783(6) (chi2 = 4, P < 0.05), some signs of association and disequilibrium between these loci and CAE. A suspected association of childhood absence epilepsy in the Chinese population to chromosome 8q24 has been proposed. In addition, it is hypothesized that the CAE gene might have a genetic heterogeneity in the population from a different race. PMID- 15275698 TI - Melatonin in wake-sleep disorders in children, adolescents and young adults with mental retardation with or without epilepsy: a double-blind, cross-over, placebo controlled trial. AB - The aim of the present study was to verify the clinical efficacy of melatonin (MLT) in children, adolescents and young adults with wake-sleep disorder and mental retardation, most of them on chronic anticonvulsant therapy for epileptic seizures, by means of a randomized, double-blind, placebo-controlled cross-over trial. Twenty-five patients (16 males, nine females), aged from 3.6 to 26 years (mean 10.5 years), all affected with mental retardation mostly with epileptic seizures, were randomized to oral synthetic fast-release MLT or placebo. Melatonin was initiated at the daily dose of 3 mg, at nocturnal bedtime. In case of inefficacy, MLT dose could be titrated up to 9 mg the following 2 weeks at increments of 3 mg/week, unless the patient was unable to tolerate it. The analysis of all the sleep logs disclosed a significant treatment effect of melatonin on sleep latency (P = 0.019). Melatonin was well tolerated in all patients and no side effects were reported. In conclusion, our study supports the efficacy of MLT in young patients with mental disabilities and epileptic seizures in improving the wake-sleep disorders such as time to fall asleep. Overall, MLT appeared to influence the seizure frequency poorly, though there may be occasional seizure worsening or improving. Such a dual effect requires further studies in young epileptic patients. PMID- 15275699 TI - Spontaneous improvement of intractable epileptic seizures following acute viral infections. AB - In general, epileptic seizures become more serious following infections. However, transient and permanent improvement of epileptic seizures has been observed following acute viral infections, without a recent change in anti-epileptic therapy. Questionnaires were sent to 73 institutions, throughout Japan, where pediatric neurologists care for children with epilepsy to characterize this phenomenon through clinician survey. Completed surveys were received from 11 institutions, and 21 cases were selected for the study. The age of the patients were 6 months to 17 years. The West syndrome or epilepsy subsequent to West syndrome cases were 16 out of 21. Two cases of symptomatic generalized epilepsy and one case each of symptomatic partial epilepsy, continuous spike-waves of slow sleep and severe myoclonic epilepsy in infancy were also reported. These seizures disappeared within 2 weeks subsequent to viral infections such as, exanthema subitum, rotavirus colitis, measles and mumps. The disappearance of intractable epileptic seizures following acute viral infections might be related to the inflammatory processes or the increased levels of antibodies after viral infections. PMID- 15275700 TI - Non-invasive screening of fragile X syndrome A using urine and hair roots. AB - The diagnosis of fragile X A syndrome (FRAXA) during childhood depends largely on DNA-based diagnostic tests due to the lack of the specific clinical features. To determine a non-invasive screening method for fragile X syndrome, we studied the method of DNA-based diagnosis using urine or hair roots instead of routinely used peripheral blood cells. The amplification of repeat-containing alleles of FMR-1 by PCR using Pfu polymerase was applied on DNA extracted from urine sediments or hair roots of 50 and 28 normal individuals, respectively. Consistent amplification of repeat-containing DNA fragments of normal size to ethidium visible quantities were obtained in 92% (46/50) of urine samples and 100% (28/28) of hair roots. No bands of normal size or abnormal or artificial smears were detected in two male FRAXA patients. No female samples were examined in the present study because the separation of two alleles was unsatisfactory on agarose gels with DNA from blood samples. Our results indicate that the use of hair roots in a DNA-based test constitutes a rapid, simple and less-invasive screen to diagnose males with FRAXA. PMID- 15275701 TI - Lymphoblastoid cell lines of Rett syndrome patients exposed to oxidative-stress induced apoptosis. AB - Despite the identification of mutations in the methyl CpG binding protein 2 gene, the pathogenesis of Rett syndrome (RS) is still unknown. In order to clarify the role of apoptosis in this disorder, we studied lymphoblastoid cell lines in five classical RS patients and five controls, incubated with 2-deoxy-d-ribose (dRib), a reducing sugar that induces apoptosis in human cells, through oxidative damage. The apoptotic response was detected by flow cytometric analysis and agarose gel electrophoresis. The cells of RS patients showed a lower percentage of apoptosis in a routine condition than those of controls did, whereas, in the presence of dRib, the percentage of apoptotic cells in RS patients increased with time and reached the same percentage of those of controls at 72 h. The data observed here suggest that RS may have a low susceptibility or an increased resistance to the apoptotic cell death, which may be corrected only in the presence of a strong apoptotic stimulus. PMID- 15275702 TI - Postural rhythmic muscle bursting activity in Angelman syndrome. AB - Postural impairment is one of the most consistent features of Angelman syndrome. Using multiple-channel electromyography, we studied a lower limb and an upper limb isometric postural task in 14 patients with Angelman syndrome and 18 unimpaired control subjects. Both tasks were associated with synchronous bursts of activity at frequencies of 6-8 s(-1) in all recorded muscles in all patients with Angelman syndrome and none of the control subjects. This pattern was not altered by extra-loading. Electroencephalogram recorded during the upper limb task showed no change in relation to the task. Burst-locked back-averaging of the electroencephalogram showed no spiking before or during the bursts. Various physiological and pathological rhythmic muscle activities have been proposed to be a manifestation of oscillations in the central nervous system and it has been suggested that such oscillations may have a role in the processing of motor commands. The mechanism of the rhythmic muscle bursting activity associated with maintaining posture in patients with Angelman syndrome is not clear, although it could be consistent with cerebellar Purkinje cell dysfunction, either as a pathological feature or as an adaptive process to overcome deficits in motor coordination. PMID- 15275703 TI - Spontaneous recurrent seizure following status epilepticus enhances dentate gyrus neurogenesis. AB - It is known that evoked seizures can increase neurogenesis in the dentate gyrus in adult rats. Whether spontaneous seizures occurring after status epilepticus (SE) also results in alterations in neurogenesis is not known. Here, we measured neurogenesis in rats with and without spontaneous seizures following SE. Lithium pilocarpine was used to induce seizures in postnatal (P) day 20 rats. Spontaneous seizure frequency was assessed 2 months using video monitoring. Rats then received bromodeoxyuridine to label dividing DNA and were sacrificed 24 h later. Animals with spontaneous seizures (n = 9) had a modest increase in neurogenesis compared to animals with SE (n = 6) and no spontaneous seizures and control rats (n = 10). These findings demonstrate that the hippocampus is capable of generating new neurons weeks following SE and further that recurrent seizures enhance the production of new neurons. These alterations in neurogenesis may contribute to ongoing pathological changes week and months following SE. PMID- 15275704 TI - Neuron-specific enolase as a marker of the severity and outcome of hypoxic ischemic encephalopathy. AB - The aim of this study was to evaluate serum concentrations of neuron-specific enolase (NSE) as a marker of the severity of hypoxic ischemic encephalopathy (HIE) and to elucidate the relation among the concentrations of NSE, grade of HIE and short-term outcome. Forty-three asphyxiated full-term newborn infants who developed symptoms and signs of HIE (Group 1) and 29 full-term newborn infants with meconium-stained amniotic fluid but with normal physical examination (Group 2) were studied with serial neurological examination, Denver developmental screening test (DDST), electroencephalogram and computerized cerebral tomography (CT) for neurological follow-up. Thirty healthy infants were selected as the control group. In the patient groups, two blood samples were taken to measure NSE levels, one between 4 and 48 h and the other 5-7 days after birth. Serum NSE levels were significantly higher in infants with HIE compared to those infants in Group 2 and control group. The mean serum concentrations of the second samples decreased in all groups studied but they were significantly higher in Group 1 compared to those in Group 2. Serum NSE concentrations of initial samples were significantly higher in patients with stage III HIE than in those with stages II and I. The sensitivity and specificity values of serum NSE as a predictor of HIE of moderate or severe degree (cut-off value 40.0 microg/l) were 79 and 70%, respectively, and as a predictor of poor outcome (cut-off value 45.4 microg/l) were calculated as 84 and 70%, respectively. The predictive capacity of serum NSE concentrations for poor outcome seems to be better than predicting HIE of moderate or severe degree. However, earlier and/or CSF samples may be required to establish serum NSE as an early marker for the application of neuroprotective strategies. PMID- 15275705 TI - Ictal MEG in two children with partial seizures. AB - We report on the successful identification of epileptic foci in two children with partial epilepsy using ictal magnetoencephalography (MEG). Case 1 is a 12-year old male suffering with simple partial seizures with leftwards nystagmus. Ictal SPECT revealed a hyperperfusion area in the right lateral occipital area, and MRI revealed cortical dysplasia in the same area. Interictal EEG dipoles were concentrated in the right mesial occipital lobe. Both interictal and ictal MEG dipoles were concentrated in the right mesial occipital lobe, which corresponded well with neuroimaging data and his clinical features. Case 2 is a 5-year-old female suffering with simple partial seizures with left-side facial twitching. Interictal EEG dipoles were located in her left motor area, the pre-sylvian fissure, close to the location of the interictal MEG-estimated dipoles. Ictal EEGs showed no remarkable changes associated with her clinical manifestations. However, ictal MEG showed high-voltage slow waves over her left hemisphere, and ictal MEG iso-contour maps revealed a clear dipolar pattern, which suggested that the MEG dipole was located in the area of the sylvian fissure. Ictal SPECT revealed hyperperfusion areas around the left sylvian fissure. CONCLUSION: Ictal MEG is useful for determining the precise location of epileptic focus in patients with motionless seizures, including children. PMID- 15275706 TI - Unusual compulsive motor activity during treatment with clothiapine in a mentally retarded adolescent. AB - Atypical antipsychotic agents, specifically those with a high hyposerotonergic activity such as clozapine and clothiapine, have been associated with de novo obsessive-compulsive symptoms. We report the case of a 16-year-old adolescent male with severe mental impairment and disruptive behaviour who developed a compulsive head and body turning disorder on clothiapine. Such a symptom had to be distinguished from epileptic partial seizures; it promptly disappeared with the drug discontinuation. PMID- 15275707 TI - Hemiconvulsion-hemiplegia syndrome and primary human herpesvirus 7 infection. AB - We report a case of hemiconvulsion-hemiplegia (HH) syndrome. An 18-month-old female infant had a hemiconvulsion followed by left hemiplegia. Magnetic resonance imaging immediately after the onset of hemiplegia showed high intensity in the right hemisphere in diffusion-weighted images (DWI), while T1- and T2 weighted images were normal. Single photon emission computed tomography showed hypoperfusion of the right hemisphere in the acute phase. Virological analyses proved primary human herpesvirus 7 (HHV-7) infection. DWI are useful for the early evaluation of HH syndrome. Vascular disorders due to HHV-7 infection may have been related to the development of HH syndrome in this patient. PMID- 15275708 TI - How and why does radioimmunotherapy work? PMID- 15275709 TI - Mirror, mirror on the wall--which is the greatest predictive assay of them all? PMID- 15275710 TI - Radiobiology of radioimmunotherapy: targeting CD20 B-cell antigen in non Hodgkin's lymphoma. AB - The radiobiology of radioimmunotherapy is an important determinant of both the toxicity and the efficacy associated with the treatment of B-cell non-Hodgkin's lymphoma with radiolabeled anti-CD20 monoclonal antibodies. The properties of the target, CD20, and the mechanisms of action of both the monoclonal antibodies and the associated exponentially decreasing low-dose-rate radiotherapy are described. The radiation dose and dose-rate effects are discussed and related to both the tumor responses and normal organ toxicity. Finally, the use of either unlabeled or radiolabeled anti-CD20 monoclonal antibodies as a component of combined modality therapy (including the sequential or concurrent use of sensitizers) and future directions of the field are discussed. PMID- 15275711 TI - Phase I clinical evaluation of near-simultaneous computed tomographic image guided stereotactic body radiotherapy for spinal metastases. AB - PURPOSE: To evaluate in a Phase I study the safety, feasibility, and patient positioning accuracy of treating patients with intensity-modulated, near simultaneous, computed tomographic (CT) image-guided stereotactic body radiotherapy (SBRT). PATIENTS AND METHODS: Fifteen consecutive patients with metastatic spinal disease who met protocol eligibility criteria were entered into a Phase I clinical trial. Each patient received five treatments of intensity modulated, near-simultaneous CT image-guided SBRT, for a total of 75 treatments with 90 isocenter setups during the course of the study. Patients uniformly received 30 Gy (if possible) of radiotherapy in 5 fractions to the clinical target volume. The total dose was constrained by limiting the spinal cord to a maximum dose of 10 Gy. To verify correct daily patient positioning before each treatment and to determine the daily treatment setup error after radiation delivery, axial CT scans were taken before and immediately after each treatment without moving the patient from the treatment position, for comparison with the planning CT scan. Toxicity was measured using the Common Toxicity Criteria, the Late Effects of Normal Tissue scoring system and a neurologic function scale. Follow-up was conducted 4 weeks after completion of SBRT, and then 2, 3, 6, 9, 12, and every 6 months thereafter. RESULTS: The procedure was technically feasible to perform in all patients. No neurologic toxicity was observed in any patient. The median follow-up time was 9 months (range 6-16). The Clopper-Pearson upper bound on the probability of paralysis with 95% confidence is no greater than 0.181. The positional setup error was determined to be within 1 mm of planning isocenter. CONCLUSIONS: This Phase I study shows that intensity modulated, near simultaneous, CT image-guided SBRT is a feasible, and highly precise technique for the noninvasive treatment of spinal metastases. Although no paralysis has developed in the 15 patients treated, continued monitoring for spinal cord toxicity is warranted, as larger numbers of patients will be needed to more precisely define the upper bound on the probability of spinal cord myelopathy. PMID- 15275712 TI - Prediction of outcome in head-and-neck cancer patients using the standardized uptake value of 2-[18F]fluoro-2-deoxy-D-glucose. AB - PURPOSE: Tumor uptake of 2-[(18)F] fluoro-2-deoxy-D-glucose (FDG) may relate to outcome in cancer patients. Pretreatment FDG uptake was evaluated as a predictor of local control (LC) and disease-free survival (DFS) in patients with head-and neck cancer managed primarily either by radiotherapy (RT) or surgery. PATIENTS AND METHODS: Tumor FDG uptake using the Standardized Uptake Value (SUV) was measured in 120 patients studied prospectively using positron emission tomography (PET). Treatment consisted of either radical RT with or without chemotherapy (73 patients) or radical surgery with or without postoperative RT (47 patients). Median follow-up of the surviving patients was 48 months. RESULTS: The median SUV was higher in 46 patients who failed treatment than in the remaining controlled patients (5.8 vs. 3.6, p = 0.002). In monovariate analysis, patients with tumors having high FDG uptake (SUV > median, 4.76) had poorer LC (p = 0.003) and DFS (p = 0.005). This difference was also observed when the RT and surgery groups were analyzed separately. In the multivariate analysis T-category (p = 0.005) and SUV (p = 0.046) remained independent adverse factors for LC, whereas N-category (p = 0.004), T-category (p = 0.02) and SUV (p = 0.05) were independent determinants of DFS. CONCLUSION: These results suggest that pretreatment tumor FDG uptake represents an independent prognostic factor in patients with head-and-neck cancers, whatever the primary treatment modality. Tumors having high FDG uptake are at greater risk of failure and should be considered for more aggressive multimodality therapy. PMID- 15275713 TI - A population-based atlas and clinical target volume for the head-and-neck lymph nodes. AB - PURPOSE: To develop a population-based three-dimensional lymph node target volume of the head and neck. METHODS AND MATERIALS: The T2 weighted axial magnetic resonance imaging (MRI) images of 35 patients with known head and neck cancer were reviewed. An experienced head and neck radiologist marked each lymph node (LN) electronically. The images were distributed to one of 12 axial levels of the head and neck with each level representing a distinct portion of the neck based on external contours and the presence of anatomic structures. The LNs were marked with five different symbols to distinguish the superior/inferior extent of each LN within each level. With the categorization of each image into a different superior/inferior level of the neck, the registration of the images was limited to two dimensions. Nonlinear transformation accounted for inter-patient differences although no local warping was used. The co-registration used recognizable anatomic landmarks (vertebral body, mandible, maxilla, clivus as well as the sternocleidomastoid muscle, external skin contour, spinal cord) to match the patient anatomy. RESULTS: In total, 503 images were co-registered with the baseline images. The majority of the co-registrations were of good quality; 361, 122, and 20 image co-registrations scored as global, limited, and poor co registrations respectively. One thousand and fifty seven LNs were marked, with 122 LNs marked as submandibular and submental LN. Among the levels A, B, C, D, E, F, G, H, 22, 44, 206, 199, 196, 175, 63, 35 LN were marked respectively. Noteworthy anatomic variation was observed among the different nodal groups that are summarized in the representative baseline images. CONCLUSIONS: Image registration of a series of head and neck images generates a valuable population based lymph node map that can be used to guide the three-dimensional delineation of the elective lymph node target volume. Significant variation in the lymph node location was seen in all LN groups. The medial border of the internal jugular vein can be used as an important landmark structure in delineating the jugular LN clinical target volume and, for that reason, intravenous contrast is recommended to improve visualization. The location of the submandibular LN appear to be limited to the space anterior and lateral to the submandibular gland and are found mostly along the inferior edge of the mandible. The location of the retropharyngeal LN does vary but their location does not appear to vary with any other recognizable axial structure. The lymph node map provides another collaborating piece of evidence in defining the head and neck LN clinical target volume. PMID- 15275714 TI - Concurrent chemoradiotherapy in locoregionally recurrent nasopharyngeal carcinoma. AB - PURPOSE: To analyze the results of concurrent chemoradiotherapy in patients with locoregional recurrent nasopharyngeal carcinoma. METHODS AND MATERIALS: We performed a retrospective analysis of 35 patients with locoregional recurrent nasopharyngeal carcinoma referred to our department between March 1994 and November 2002. Most patients were male (77%), Chinese (97%), and had undifferentiated carcinoma (89%). Most had extensive locally recurrent Stage rT3 T4 disease (66%) with a median age at recurrence of 49 years (range, 35-69 years). A repeat course of radiotherapy was given concurrently with cisplatin, with cisplatin/5-fluorouracil as consolidation treatment. Significant morbidities were present, including cranial nerve palsies due to extensive recurrent local disease before treatment of the recurrence. RESULTS: The response rate to concurrent chemoradiotherapy was 58% (29% complete response and 29% partial response). The 5-year progression-free and overall survival rate, calculated using the Kaplan-Meier method, was 15% and 26%, respectively. Only 3 patients developed systemic metastases. Grade 3-4 acute toxicities included emesis (9%) and neutropenia (14%), and Grade 3-4 late toxicities consisted of temporal lobe necrosis (3%), cranial neuropathy (6%), and endocrine abnormalities (14%). CONCLUSION: Concurrent chemoradiotherapy is feasible in a selected group of patients with locoregional recurrent NPC, but the risk of major late toxicities is significant. PMID- 15275715 TI - Radiotherapy of squamous cell carcinoma of the nasal vestibule. AB - PURPOSE: To evaluate the results of primary radiotherapy (RT) for squamous cell carcinoma of the nasal vestibule. METHODS AND MATERIALS: A total of 56 patients with Stage T1 and T2 tumors (Wang classification) were treated with external beam RT (EBRT) with or without a boost using endocavitary brachytherapy. Of these, 32 were treated with EBRT and an additional boost with intermediate-dose-rate brachytherapy, 15 with EBRT and a boost with high-dose-rate brachytherapy, and 9 with EBRT alone. RESULTS: The local control rate at 2 years was 80%. Most cases could be successfully salvaged with surgery, resulting in an ultimate local control rate of 95%. No statistically significant differences were noted among the different treatment approaches. Of the 56 patients, 12% developed lymph node metastases. CONCLUSION: Primary RT is an adequate treatment for Stage T1 and T2 squamous cell carcinoma of the nasal vestibule with excellent cosmetic results. The radiation technique used depends on the primary tumor extension and the experience of the treating radiation oncologist. In the case of N0 disease, elective treatment of the regional lymph nodes is not recommended. PMID- 15275716 TI - Brachytherapy with 192Ir as treatment of carcinoma of the tarsal structure of the eyelid. AB - PURPOSE: Some eyelid tumors present a great challenge for surgical therapy because of the cosmetic and functional impairment. Interstitial radiation with (192)Ir is an optimal alternative treatment modality. The aim of this study was to evaluate the local tumor control and cosmetic results in patients with eyelid tumors treated by interstitial (192)Ir wires. METHODS AND MATERIALS: Twenty-four previously untreated carcinomas involving the eyelid tarsal structure in 23 patients were treated with (192)Ir wire implantation. The tumor location was in the lower eyelid in 22 cases and in the upper eyelid in 2. The mean tumor size was 1.33 cm. Of the 24 tumors, 79.2% were basal cell carcinoma, 16.7% were squamous cell carcinoma, and 4.2% were adenocarcinoma. The total radiation dose was 4000 cGy, delivered to 2-mm depth (mean dose rate, 73 cGy/h). RESULTS: The mean follow-up was 43 months. Local control was obtained in 22 (91.6%) of 24 tumors. Good functional results were achieved in all patients. CONCLUSION: (192)Ir interstitial brachytherapy in a braided silk filament appears to be an excellent method to treat carcinomas involving the eyelid tarsal structure for tumor control and functional and cosmetic results. PMID- 15275717 TI - Efficacy of I131 ablation therapy using different doses as determined by postoperative thyroid scan uptake in patients with differentiated thyroid cancer. AB - PURPOSE: The optimal dose of I(131) for ablation of functioning residual thyroid tissue after surgery is controversial. The current study was conducted to determine the optimal dose of I(131) for remnant postoperative ablation. A review of the literature is included. METHODS AND MATERIALS: A total of 238 patients with papillary and follicular carcinoma were treated with I(131) for ablation of a postoperative thyroid remnant. The I(131) dose was based on the 24-h percentage of neck uptake in the postoperative thyroid scans. Patients with < 5% uptake received a median of 85 mCi; 6-10% uptake, a median of 80 mCi; 11-15% uptake, a median of 60 mCi; 16-20% uptake, a median of 50 mCi; and > or =21% uptake, a median of 30 mCi. The ablation results were compared with the prognostic factors. RESULTS: Complete ablation was observed in 40 (92%) of 43 patients receiving 85 mCi, in 31 (94%) of 33 who received 80 mCi, in 39 (95%) of 41 who received 60 mCi, in 51 (93%) of 55 who received 50 mCi, in 37 (94%) of 39 who received 30 mCi, and in 18 (96%) of 19 who received 30 mCi. The overall successful ablation rate was 94% (95% confidence interval, 89-100%). CONCLUSION: Our findings suggest that patients with differentiated thyroid cancer can be treated with doses of I(131) according to the percentage of neck uptake of postoperative total body scan, with high complete ablation rates, without exposing patients to higher dose levels of I(131). PMID- 15275718 TI - Her2/neu-positive disease does not increase risk of locoregional recurrence for patients treated with neoadjuvant doxorubicin-based chemotherapy, mastectomy, and radiotherapy. AB - PURPOSE: Preclinical data suggest that overexpression of Her2/neu confers cellular radioresistance. We retrospectively studied whether Her2/neu-positive disease was associated with locoregional recurrence (LRR) after postmastectomy radiotherapy (RT) for breast cancer. METHODS AND MATERIALS: Data from 337 patients treated in four institutional prospective clinical trials neoadjuvant doxorubicin-based chemotherapy, mastectomy, and RT were reviewed. The trials were conducted between 1989 and 2000. Of the 337 patients, 108 (32%) had tumors that were tested for Her2/neu, with positivity defined by 3+ immunohistochemistry staining or gene amplification detected by fluorescence in situ hybridization. RT was delivered to the chest wall and draining lymphatics (median dose, 50 Gy) followed by a chest wall boost (median dose, 10 Gy). RESULTS: Thirty-two patients had Her2/neu-positive disease and 76 patients had Her2/neu-negative disease. The Her2/neu-positive tumors were associated with a greater rate of estrogen receptor negative disease (p = 0.03), the presence of supraclavicular disease at diagnosis (p = 0.027), and a greater number of positive lymph nodes after chemotherapy (p = 0.026). Despite these adverse features, the actuarial overall LRR rate was roughly equivalent for the patients with Her2/neu-positive tumors vs. those with Her2/neu-negative tumors (5-year rate 17.5% vs. 13.9%, respectively; 10-year rate 17.5% vs. 18.9%, respectively; p = 0.757). On Cox regression analysis of LRR adjusted for N stage and estrogen receptor status, the hazard ratio for Her2/neu positivity was 0.89 (95% confidence interval, 0.31-2.59; p = 0.83). CONCLUSION: Her2/neu overexpression does not appear to predispose to LRR after neoadjuvant doxorubicin-based chemotherapy, mastectomy, and RT. PMID- 15275719 TI - Impact of mean rectal dose on late rectal bleeding after conformal radiotherapy for prostate cancer: dose-volume effect. AB - PURPOSE: To identify the clinical and dosimetric factors predictive of a greater risk of Grade 2 or worse late rectal bleeding in patients with localized prostate cancer treated with three-dimensional conformal radiotherapy in a prospective dose-escalation study. METHODS AND MATERIALS: We performed a retrospective analysis of the clinical records and dose-volume histograms of 107 patients with Stage T1c-T3 prostate cancer treated at our institution with three-dimensional conformal radiotherapy who had a minimal follow-up of 1 year. Of the 107 patients, 21 were treated at dose level 1 (70.0 Gy), 57 at dose level 2 (72.0 Gy), and 29 at dose level 3 (75.6 Gy). The mean International Commission on Radiation Units and Measurements reference dose was 76.5 Gy (range, 69.8-82.6 Gy). RESULTS: The 4-year actuarial incidence of Grade 2 or worse late rectal bleeding was 7.7% +/- 2.5%. The results of the multivariate analysis indicated that the mean rectal dose (rectal D(mean); p = 0.003) and the percentage of rectum receiving >60 Gy (Vr(60); p = 0.002) correlated with Grade 2 or worse rectal bleeding. The receiver operating characteristic curve analysis showed that this logistic regression model using both Vr(60) and rectal D(mean) had good reliability to predict the risk of late rectal bleeding. The area under the curve for Vr(60) and rectal D(mean) was 0.889 and 0.892, respectively. CONCLUSION: The results of the present study provide clear evidence of a dose-volume effect and the importance of intermediate doses (60.0 Gy) on the risk of rectal bleeding at this prescription dose level. On the basis of these results, new constraints have been implemented in our institution to keep the risk of developing Grade 2 rectal bleeding reasonably low (rectal D(mean) 50.0 Gy and Vr(60) 42%). PMID- 15275720 TI - Combined modality treatment in the management of high-risk prostate cancer. AB - PURPOSE: The efficacy of a multimodality protocol using neoadjuvant and concomitant hormonal therapy, brachytherapy, and three-dimensional conformal external beam radiotherapy (RT) in high-risk prostate cancer was evaluated using biochemical outcomes and posttreatment biopsy results. METHODS AND MATERIALS: Between February 1994 and November 1999, 132 high-risk patients were treated with combined hormonal therapy (9 months), permanent radioactive seed brachytherapy, and external beam RT, with follow-up ranging from 36 to 88 months (median, 50 months). The eligibility criteria were any of the following: Gleason score 8-10, initial prostate-specific antigen (PSA) level >20 ng/mL, clinical Stage T2c-T3, or positive seminal vesicle biopsy, or two or more of the following: Gleason score 7, PSA level >10-20 ng/mL, or Stage T2b. Twenty percent of patients had a positive seminal vesicle biopsy before therapy. Negative laparoscopic pelvic lymph node dissections were performed in 44% of patients. RESULTS: The actuarial overall freedom from PSA failure rate was 86% at 5 years. The freedom from PSA failure rate at 5 years was 97% for those with a Gleason score of < or =6 (35 of 36), 85% for a Gleason score of 7 (50 of 59), and 76% for a Gleason score of 8-10 (28 of 37; p = 0.03). A trend was noted toward worse outcomes in seminal vesicle biopsy-positive patients, with a 5-year freedom from PSA failure rate of 74% vs. 89% for all other patients (p = 0.06). Posttreatment prostate biopsies were performed in 47 patients and were negative in 96% at the first biopsy and 100% at the last biopsy. CONCLUSION: Trimodality therapy with androgen suppression, brachytherapy, and external beam RT for high-risk prostate cancer results in excellent biochemical and pathologically confirmed local control. PMID- 15275721 TI - Acute urinary retention after transperineal template-guided prostate biopsy. AB - PURPOSE: Urinary retention occurs in 5%-36% of patients with prostate cancer after implantation of radioactive seeds for brachytherapy. We used transperineal biopsy as a model to determine the influence of needle trauma on urinary retention. METHODS AND MATERIAL: We retrospectively reviewed medical records of 157 men with high risk of prostate cancer who underwent systematic ultrasound guided biopsy of the prostate with the transperineal template technique and an 18 gauge automated biopsy device. RESULTS: Eighteen of 157 patients (11.5%; 95% confidence interval, 6.9%-17.5%) had urinary retention within 48 hours after biopsy. Median age was 68.5 years in patients with retention vs. 67.0 years in patients without (p = 0.319); median calculated prostate volume, 76.5 vs. 51.5 mL (p = 0.015); and median number of biopsy cores, 22.0 vs. 20.0 (p = 0.038). Age distribution differed between groups (p = 0.047), with more younger men in the no retention group. On multivariate analysis, only number of biopsy cores significantly predicted urinary retention (p = 0.003). Four patients required transurethral resection; 1 had an indwelling catheter until radical prostatectomy; and 13 were catheter-free within 1-5 days. CONCLUSIONS: Needle trauma alone may cause urinary retention in men undergoing transperineal procedures. The number of needle incursions and prostate size are predictors of postprocedure urinary retention. PMID- 15275722 TI - Androgen deprivation-induced changes in prostate anatomy predict urinary morbidity after permanent interstitial brachytherapy. AB - PURPOSE: To evaluate the cytoreductive consequences of neoadjuvant androgen deprivation therapy on International Prostate Symptom Score (IPSS) normalization, catheter dependency, and the need for surgical intervention secondary to bladder outlet obstruction after permanent interstitial brachytherapy. METHODS AND MATERIALS: A total of 116 patients (median follow-up, 30 months) with preandrogen and postandrogen deprivation therapy ultrasound studies and no history of preimplant transurethral resection of the prostate were evaluated. Androgen deprivation-induced changes in prostate volume, transition zone (TZ) volume, and urethral location were correlated with IPSS resolution, catheter dependency, and the need for postimplant surgical intervention. Prostate gland and TZ dimensions and volumes were measured by prolate ellipsoid calculation from the static ultrasound images. The urethral location was determined by identification of a urinary catheter. Additional clinical, treatment, and dosimetric parameters evaluated included patient age, pretreatment prostate-specific antigen, Gleason score, clinical T stage, preimplant IPSS, pre- and postandrogen deprivation ultrasound studies, treatment planning volume, supplemental external beam RT, isotope, total implant activity, Day 0 maximal dose received by 90% of the prostate gland, Day 0 percentage of prostate volume receiving 100%, 150%, and 200% of the prescribed minimal peripheral dose, and urethral dose. RESULTS: For hormonally manipulated patients, the prostate volume at implantation did not have a statistical influence on the percentage of patients returning to IPSS baseline, the time for IPSS normalization, the incidence of catheter dependency, the catheter-dependency time, or the need for postimplant surgical intervention. However, when compared with the hormone-naive cohort, hormonally manipulated patients were more likely to undergo postimplant surgical intervention (5.2% vs. 0.3%, p = 0.001). Greater androgen deprivation-induced reductions in prostate and TZ volumes, along with movement of the urethra closer to the posterior border of the prostate gland, resulted in a decreased incidence of postimplant urinary morbidity. Using Cox regression analysis, the time to IPSS resolution was best predicted by the percentage of TZ volume reduction. Stepwise linear regression analysis demonstrated that the catheter-dependency time was best predicted by the prehormonal therapy prostate volume, posthormonal therapy TZ volume, and the change in the urethral position; prolonged catheter dependency by the percentage of TZ volume reduction, prehormonal therapy TZ index, and the change in the urethral position; and the need for postimplant surgical intervention by the posthormonal therapy TZ index and the change in the urethral location. CONCLUSION: After neoadjuvant androgen deprivation therapy for volume reduction, some brachytherapy-related urinary morbidity parameters are highly related to the preandrogen deprivation prostate volume, variants in the TZ volume, and changes in the urethral location. PMID- 15275723 TI - The radiation doses to erectile tissues defined with magnetic resonance imaging after intensity-modulated radiation therapy or iodine-125 brachytherapy. AB - PURPOSE: To report penile bulb (PB) and corporal bodies (CB) doses during intensity-modulated radiation therapy (IMRT) and permanent (125)I prostate implant alone (BT) for favorable, early stage, clinically localized prostate cancer using computed tomography (CT) and magnetic resonance imaging (MRI) to provide a basis for comparison as the initial report of a comprehensive project to develop erectile tissues sparing techniques. METHODS AND MATERIAL: Prostate, PB and CB volumes were defined by a fused CT/MRI simulation study performed before treatment in 29 IMRT patients and verification study performed 30 days postimplant in 15 BT patients. The median prescribed prostate dose for the IMRT and BT groups was 74 Gy and 145 Gy, respectively. Dose volume histograms (DVHs) were generated to determine the dose characteristics for the PB, CB, and prostate for each patient. D(90), V(100), and V(50) were used, where D(i) was defined as the dose that covers i% of the prostate volume and V(i) is the fractional volume of the prostate that receives i% of the prescribed dose. The Wilcoxon rank sum test was used to evaluate significance between the groups. RESULTS: The median PB D(90), V(100), and V(50) values were 17.5 Gy, 0%, and 31.9% for the IMRT group; and 52.5 Gy, 21.5%, and 89.7% for the BT group. The median CB D(90), V(100), and V(50) values were 7.3 Gy, 0%, and 0.9% for the IMRT group; and 26.9 Gy, 2.4%, and 20.1% for the BT group. The differences between the IMRT vs. BT V(100) values, but not V(50), were statistically significant for the PB (p = 0.001) and CB (p = 0.001). CONCLUSIONS: Radiation dose to the PB and CB is low with IMRT or BT. Magnetic resonance imaging is superior to CT for the imaging of erectile tissues. Intensity-modulated radiation therapy may offer further reductions in the doses received by the PB and CB; however, at what cost to prostate coverage and normal tissue sparing will be the subject of a follow-up study. PMID- 15275724 TI - Correlating the degree of needle trauma during prostate brachytherapy and the development of acute urinary toxicity. AB - PURPOSE: To determine if there is an association between the degree of prostate trauma during prostate brachytherapy and development of acute urinary toxicity. METHODS AND MATERIALS: In a consecutive prospective cohort of permanent (125)I prostate brachytherapy patients, the number of times each needle was repositioned was tracked, and the dosimetry plans were used to determine the number of times needles within 1 cm of the urethra were manipulated. Additionally, prostate volume, total number of needles, number of needles/prostate volume, and the number of periurethral needle manipulations/prostate volume were determined. The need for catheterization beyond 24 hours and the Radiation Therapy Oncology Group (RTOG) urinary toxicity score at 4 weeks were recorded. The independent samples t test was used to search for a correlation between these parameters and the recorded toxicity scores. RESULTS: Twenty-eight consecutive implant patients were evaluated in the study. Median (range) values were as follows: prostate volume 35 cc ( range, 15-51 cc), number of needles per patient 32 (range, 21-41), number of needle manipulations per patient 94.5 ( range, 55-147), and number of periurethral needle manipulations 42 (range, 17-65). The only significant association between urinary toxicity and these variables was for the number of periurethral needle manipulations (p = 0.025). CONCLUSIONS: These data provide evidence that needle prostate trauma during brachytherapy contributes to acute urinary toxicity. PMID- 15275725 TI - Factors predicting for urinary incontinence after prostate brachytherapy. AB - PURPOSE: To define risk factors that predict for urinary incontinence after (125)I prostate brachytherapy. METHODS AND MATERIALS: Urinary incontinence after (125)I prostate brachytherapy was evaluated using a patient self-assessment questionnaire based on the NCI Common Toxicity Criteria (version 2). Grade 0 is defined as no incontinence; Grade 1 incontinence occurs with coughing, sneezing, or laughing; Grade 2 is spontaneous incontinence with some control; and Grade 3 is no control. One hundred fifty-three patients received monotherapy (145 Gy) (125)I implants between October 1996 and December 2001, and 112 (75%) responded to our survey. Median follow-up was 47 months (range, 14-74 months). Patient characteristics included a preimplant prostate-specific antigen < or =10, Gleason score < or =6, and stage < or =T2b. CT-based postimplant dosimetry was analyzed approximately 30 days after the procedure, and dose-volume histograms of the prostate and the prostatic urethra were generated based on contoured volumes. Dosimetric parameters evaluated as predictive factors for incontinence included the prostate volume; total activity implanted; number of needles; number of seeds; seed activity; urethral D(5), D(10), D(25), D(50), D(75), and D(90) doses; prostate D(90) doses; and prostate V(100), V(200), and V(300). Clinical parameters evaluated included age, Gleason score, prostate-specific antigen, preimplant International Prostate Symptom Score (I-PSS), and length of follow-up. RESULTS: Urethral D(10) dose and preimplant I-PSS predicted for urinary incontinence on multivariate analysis (p = 0.002 and p = 0.003, respectively). Twenty-eight patients reported Grade 1 incontinence (26%), and 5 patients reported Grade 2 (5%). Patients with Grade 1 and 2 incontinence were analyzed together, because of the small number of patients who experienced Grade 2. No patients reported Grade 3 incontinence. Mean urethral D(10) was 314 +/- 78 Gy in patients with Grade 0 compared with 394 +/- 147 Gy in patients with Grades 1, 2 incontinence (p = 0.002). The incidence of incontinence doubled as the urethral D(10) dose increased above 450 Gy. Patients with Grade 0 had a mean preimplant I PSS score of 6.6 +/- 4.5 compared with 10.0 +/- 6.4 for Grades 1, 2 (p = 0.003). A significant increase in the incidence of incontinence was noted when the preimplant I-PSS was greater than 15. No relationship was noted between incontinence and prostate volume, total activity implanted, or the number of needles used (p = 0.83, p = 0.89, p = 0.36, respectively). CONCLUSION: Urethral D(10) dose and preimplant I-PSS are predictive for patients at higher risk of urinary incontinence. To decrease the risk of this complication, an effort should be made to keep the urethral D(10) dose as close to the prescribed dose as possible, and the preimplant I-PSS should be thoroughly evaluated in an attempt to select patients with scores less than 15. PMID- 15275726 TI - Assessment of i-125 prostate implants by tumor bioeffect. AB - PURPOSE: A method of prostate implant dose distribution assessment using a bioeffect model that incorporates a distribution of tumor cell densities is demonstrated. This method provides both a quantitative method of describing implant quality and spatial information related to the location of underdosed regions of the prostate. This model, unlike any other, takes into account the likelihood of finding cancer cells in the underdosed region. METHODS AND MATERIAL: The prostate volumes of 5 patients were divided into multiple subsections and a unique cell density was assigned to each subsection. The assigned cell density was a function of probability of finding tumor foci in that subsection. The tumor control probability (TCP) for each subsection was then calculated to identify the location of any significantly underdosed part of the prostate. In addition, a single TCP value for the entire prostate was calculated to score the overall quality of the implant. RESULTS: Adequately dosed subsections scored TCP values greater than 0.80. The TCP for underdosed regions fell dramatically particularly in subsections at higher risk of containing tumor cells. CONCLUSIONS: Despite uncertainties in radiobiological parameters used to calculate the TCP and the distribution of cancer foci through the prostate, the bioeffect model was found to be useful in identifying regions of underdosed prostate that may be at risk of local recurrence due to inadequate dose. Unlike the isodose distribution, the model has the potential to demonstrate that small volumes of tissue underdosed in regions most likely to contain higher numbers of tumor cells may be more significant than larger volumes irradiated to a lower dose but with a lower probability of containing cancer cells. PMID- 15275728 TI - Phase III randomized trial comparing LDR and HDR brachytherapy in treatment of cervical carcinoma. AB - PURPOSE: Intracavitary brachytherapy plays an important role in the treatment of cervical carcinoma. Previous results have shown controversy between the effect of dose rate on tumor control and the occurrence of complications. We performed a prospective randomized clinical trial to compare the clinical outcomes between low-dose-rate (LDR) and high-dose-rate (HDR) intracavitary brachytherapy for treatment of invasive uterine cervical carcinoma. METHODS AND MATERIALS: A total of 237 patients with previously untreated invasive carcinoma of the uterine cervix treated at King Chulalongkorn Memorial Hospital were randomized between June 1995 and December 2001. Excluding ineligible, incomplete treatment, and incomplete data patients, 109 and 112 patients were in the LDR and HDR groups, respectively. All patients were treated with external beam radiotherapy and LDR or HDR intracavitary brachytherapy using the Chulalongkorn treatment schedule. RESULTS: The median follow-up for the LDR and HDR groups was 40.2 and 37.2 months, respectively. The actuarial 3-year overall and relapse-free survival rate for all patients was 69.6% and 70%, respectively. The 3-year overall survival rate in the LDR and HDR groups was 70.9% and 68.4% (p = 0.75) and the 3-year pelvic control rate was 89.1% and 86.4% (p = 0.51), respectively. The 3-year relapse-free survival rate in both groups was 69.9% (p = 0.35). Most recurrences were distant metastases, especially in Stage IIB and IIIB patients. Grade 3 and 4 complications were found in 2.8% and 7.1% of the LDR and HDR groups (p = 0.23). CONCLUSION: Comparable outcomes were demonstrated between LDR and HDR intracavitary brachytherapy. Concerning patient convenience, the lower number of medical personnel needed, and decreased radiation to health care workers, HDR intracavitary brachytherapy is an alternative to conventional LDR brachytherapy. The high number of distant failure suggests that other modalities such as systemic concurrent or adjuvant chemotherapy might lower this high recurrence, especially in Stage IIB and IIIB. PMID- 15275729 TI - Pathologic stage I-II endometrial carcinoma in the elderly: radiotherapy indications and outcome. AB - PURPOSE: To assess the indications for, and the outcome of, adjuvant radiotherapy (RT) in elderly patients with pathologic Stage I-II endometrial carcinoma. METHODS AND MATERIALS: Between 1980 and 2001, 79 elderly (age > or =75 years) patients with pathologic Stage I-II endometrial carcinoma were seen at our institution. All underwent total abdominal hysterectomy and bilateral salpingo oophorectomy with assessment of peritoneal cytology. Pelvic and paraaortic lymph node sampling was performed in 39 and 29 women, respectively. Patients with Stage IA and IB Grade 1-2 were designated as low risk; those with Stage IB Grade 3 and IC-IIB were designated as high risk. Thirty-four received adjuvant RT consisting of whole pelvic (n = 23) or vaginal brachytherapy (n = 3), or both (n = 8). Actuarial disease-free survival, cause-specific survival, and pelvic recurrence free survival analyses were performed by the Kaplan-Meier method. RESULTS: Adverse factors were common, including deep (>50%) myometrial invasion (47%), Grade 3 disease (28%), cervical involvement (15%), and unfavorable histologic features (15%). Overall, 46 patients (58%) had high-risk disease. Primarily because of concerns over toxicity, RT was administered in only 73%, 55%, and 67% of patients with deep myometrial invasion, Grade 3 disease, and cervical involvement, respectively. Thirty-one high-risk patients (67%) received adjuvant RT. At a median follow-up of 33.5 months, 19 patients had relapsed, for a 5-year actuarial disease-free survival rate of 67.7%. Ten patients (12%) had recurrence in the pelvis, 9 of whom had been patients treated with surgery alone. The 5-year pelvic recurrence-free survival rate of patients treated with and without RT was 97% and 73.1%, respectively (p = 0.02). The corresponding rates in the high-risk patients were 97% and 47% (p = 0.0001). High-risk patients treated with RT also had better 5-year actuarial disease-free survival (p = 0.0001) and cause-specific survival (p = 0.003) than those treated with surgery alone. RT was well tolerated, with all patients receiving their treatment as planned. Only 1 patient developed significant late toxicity. CONCLUSIONS: Adverse features are common in pathologic Stage I-II elderly endometrial carcinoma patients, and pelvic recurrence is high after surgery alone. Given the improvement in outcome and low incidence of toxicity, our results support the use of adjuvant RT in elderly pathologic Stage I-II patients with high-risk disease. PMID- 15275727 TI - MRI-guided HDR prostate brachytherapy in standard 1.5T scanner. AB - PURPOSE: Magnetic resonance imaging (MRI) provides superior visualization of the prostate and surrounding anatomy, making it the modality of choice for imaging the prostate gland. This pilot study was performed to determine the feasibility and dosimetric quality achieved when placing high-dose-rate prostate brachytherapy catheters under MRI guidance in a standard "closed-bore" 1.5T scanner. METHODS AND MATERIALS: Patients with intermediate-risk and high-risk localized prostate cancer received MRI-guided high-dose-rate brachytherapy boosts before and after a course of external beam radiotherapy. Using a custom visualization and targeting program, the brachytherapy catheters were placed and adjusted under MRI guidance until satisfactory implant geometry was achieved. Inverse treatment planning was performed using high-resolution T(2)-weighted MRI. RESULTS: Ten brachytherapy procedures were performed on 5 patients. The median percentage of volume receiving 100% of prescribed minimal peripheral dose (V(100)) achieved was 94% (mean, 92%; 95% confidence interval, 89-95%). The urethral V(125) ranged from 0% to 18% (median, 5%), and the rectal V(75) ranged from 0% to 3.1% (median, 0.3%). In all cases, lesions highly suspicious for malignancy could be visualized on the procedural MRI, and extracapsular disease was identified in 2 patients. CONCLUSION: High-dose-rate prostate brachytherapy in a standard 1.5T MRI scanner is feasible and achieves favorable dosimetry within a reasonable period with high-quality image guidance. Although the procedure was well tolerated in the acute setting, additional follow-up is required to determine the long-term safety and efficacy of this approach. PMID- 15275730 TI - The role of abdominal-pelvic radiotherapy in the management of uterine papillary serous carcinoma. AB - PURPOSE: To evaluate the role of abdominal-pelvic radiotherapy (APR) as adjuvant treatment for uterine papillary serous carcinoma (UPSC). METHODS AND MATERIAL: The medical records database at the Toronto-Sunnybrook Regional Cancer Centre identified 121 patients with the diagnosis of UPSC between 1980 and 2001. Fifty nine patients received APR as adjuvant treatment. A retrospective chart review was done to evaluate recurrence rates, sites of failure, and treatment toxicity. RESULTS: Of 59 patients who received APR, 30 had advanced-stage disease (Stage III or IV). Eleven had complete surgical staging. Median follow-up was 71 months. Twenty-five of 59 (42%) recurred, with a median time to relapse of 50 months. Five-year disease-free survival was 43%, and 5-year overall survival was 45%. Of the 25 who recurred, only 3 experienced a sole failure outside the irradiated volume. Thirteen women had their treatment interrupted or discontinued because of toxicity. CONCLUSIONS: This single-institution study reveals that there is a high recurrence rate despite APR, especially among patients with advanced stage disease, and the majority of recurrences continue to be within the irradiated volume. The role of APR remains undefined in early disease but its effectiveness is questionable in advanced disease. Innovative strategies are needed to improve outcome in these patients. PMID- 15275731 TI - Evaluation of a radiotherapy protocol based on INT0116 for completely resected gastric adenocarcinoma. AB - PURPOSE: With the results of the INT0116 study, adjuvant radiochemotherapy has become the standard treatment after complete resection of gastric adenocarcinoma. However, the implementation of radiotherapy (RT) remains a concern. In response, consensus guidelines on RT technique have been published. Our objective was to measure the inter- and intraclinician variability in RT field delineation using conventional two- (2D) and three-dimensional (3D) techniques. METHODS AND MATERIALS: Between 1999 and 2003, five radiation oncologists (ROs) treated 45 patients with completely resected, gastric adenocarcinoma using postoperative radiochemotherapy (INT0116). Two cases were included in this study (Patient 1 had cardia and Patient 2 had antral disease). Standardized vignettes (with surgical and pathologic findings) and preoperative and postoperative imaging for each case were developed. Each RO designed AP-PA fields for each patient (2D planning) on two separate occasions. This was repeated using a 3D planning technique. RESULTS: Patient 1 had a mean field area of 250.2 cm(2) (SD 12.0) and 227.9 cm(2) (SD 26.5) using 2D and 3D planning, respectively (p = 0.03). The mean clinical target volume (CTV) volume was 468.3 cm(3) (SD 65.9). Patient 1 had a significantly greater inter- than intra-RO variation for the field area designed with 3D planning; however, no difference occurred with 2D planning or CTV contouring. Patient 2 had a mean field area of 234.8 cm(2) (SD 33.1) and 226.8 cm(2) (SD 19.3) using 2D and 3D planning, respectively (p = 0.5). The mean CTV was 729.4 cm(3) (SD 67.3). For Patient 2, the inter-RO variability was significantly greater than the intra-RO variability for the field area using both 2D and 3D planning, and no difference was seen for the CTV. Composite beam's-eye-view plots revealed that the superior, inferior, and right lateral borders proved to be most contentious. CONCLUSION: Despite published guidelines and a departmental protocol, significant variations in the RT field areas were seen among ROs for both 2D and 3D planning. However, in general, CTV contouring was reproducible. Because 3D-RT hinges on accurate target identification, caution should be exercised before migrating to 3D planning for postoperative gastric cancer. PMID- 15275732 TI - 5-fluorouracil-based chemoradiation in unresectable pancreatic carcinoma: Phase I II dose-escalation study. AB - PURPOSE: A Phase I-II dose-escalation study was performed to evaluate the possible impact of the dose on response, toxicity, pain relief, and outcome in patients with unresectable pancreatic carcinoma. METHODS AND MATERIALS: A total of 50 patients entered the study. The external beam radiotherapy (RT) dose was 39.6 Gy in the first 15 patients, 50.4 Gy in the next 15 patients, and 59.4 Gy in the remaining 20 patients, at five 1.8-Gy fractions weekly. During external beam RT, patients received concurrent continuous infusion of 5-fluorouracil (1000 mg/m(2) on Days 1-4 and 21-24). Patients were evaluated for toxic reactions, local disease control, survival, and pain relief. RESULTS: No treatment-related deaths occurred from acute toxicity. Four patients required a temporary treatment interruption because of acute hematologic (2 patients) or GI (2 patients) toxicity, not correlated with the delivered RT dose. Three patients (6%) developed late toxicity (duodenal ulcer in 2 and duodenal stenosis in 1). All patients who developed late toxicity had received a dose of 59.4 Gy. At univariate analysis, only the RT dose correlated significantly with the incidence of late toxicity (at 2 years, 39.6-50.4 Gy resulted in 0% and 59.4 Gy resulted in 58.2%; p = 0.023). At multivariate analysis, the RT dose also showed a trend with the incidence of late side effects (p = 0.052). Overall, 6 patients had a partial response (12%) and 44 (88%) had no change. The overall response rate was 8.0% (95% confidence interval, 1.5-20.5%). The rate of response was not different in the three groups. In-field locoregional disease progression was seen in 7 patients (14.0%). Distant relapse was documented in 34 patients (68.0%). None of analyzed variables, in particular, the RT dose delivered, showed a statistically significant correlation with objective response, local control, incidence of metastasis, disease-free survival, or overall incidence of pain symptoms after therapy. The whole group median survival was 9 months. The actuarial survival rate at 1, 2, and 3 years was 31.3%, 2.8%, and 0.0%, respectively. None of analyzed parameters correlated significantly with survival at univariate or multivariate analysis. CONCLUSION: In a Phase I-II study, the association of high RT doses with the incidence of severe toxicity in the treatment of unresectable pancreatic carcinoma was confirmed. Furthermore, this dose-escalation study did not document a clearcut correlation, using 5-fluorouracil-based chemoradiation, between the radiation dose and clinical outcome. PMID- 15275733 TI - Adjuvant therapy in pancreatic cancer: Phase I trial of radiation dose escalation with concurrent full-dose gemcitabine. AB - PURPOSE: To determine the maximal tolerated dose of radiation delivered to the primary tumor bed, in combination with full-dose gemcitabine (1000 mg/m(2) weekly x 3), after resection of pancreatic cancer. METHODS AND MATERIALS: Patients with resected pancreatic carcinoma and poor prognostic features, a positive resection margin, or involved lymph nodes were eligible. Radiotherapy (RT) was directed at the preoperative tumor volume with a conformal technique. Regional lymph node basins were not included. The initial starting radiation dose was 24 Gy in 1.6-Gy fractions. Escalation was achieved by increasing the fraction size in 0.2-Gy increments, keeping the duration of RT to 3 weeks. Gemcitabine was given i.v. for 30-40 min at a dose of 1000 mg/m(2) before RT on Days 1, 8, and 15 of a 28-day cycle. After completion of RT and chemotherapy, an additional cycle of gemcitabine was delivered. RESULTS: Between November 1997 and October 2001, 32 patients were entered: 30 after Whipple resection (positive margins in 2, positive nodes in 22, and both in 6), 1 after distal pancreatectomy, and 1 after incomplete resection of a tumor involving the body (both patients with positive margins and nodes). Treatment was well tolerated. Of the 32 patients, 27 completed all protocol therapy and 29 maintained their pretreatment weight within 5%. Five patients experienced dose-limiting toxicity, four with Grade 3 vomiting requiring hospitalization and one fatal toxicity secondary to pneumonia/sepsis. At the final radiation dose level (42 Gy), 2 patients experienced GI dose limiting toxicity. At the 39-Gy-dose level, 5 of 6 patients were without dose limiting toxicity. Isolated local or regional progression was documented in 1 patient. Distant progression was documented in 26 of 32 patients (6 with concurrent local or regional progression). The median survival was 16.5 months (95% confidence interval 12.3-19.9) CONCLUSION: The results of our study indicate that the maximal tolerated radiation dose, administered using conformal techniques targeted to the tumor bed, is 39 Gy. In this high-risk population, data on locoregional control suggest that the reduction in radiation dose and field size minimizes toxicity and does not result in excess failures at these sites. PMID- 15275734 TI - Results of the first prospective study of carbon ion radiotherapy for hepatocellular carcinoma with liver cirrhosis. AB - PURPOSE: To evaluate the toxicity and antitumor effect of carbon ion radiotherapy for hepatocellular carcinoma within a Phase I-II trial. METHODS AND MATERIALS: Between June 1995 and February 1997, 24 patients with histopathologically proven hepatocellular carcinoma were treated to 15 fractions within 5 weeks in a step wise dose-escalation study. The disease stage was Stage II in 10, IIIA in 6, and IVA in 8 patients. The Common Toxicity Criteria, Radiation Therapy Oncology Group/European Organization for the Research and Treatment of Cancer criteria, and Child-Pugh score were used to evaluate toxicity. The antitumor effect was evaluated by the tumor response, cumulative local control, and survival rates. RESULTS: During a median follow-up of 71 months (range, 63-83 months), no severe adverse effects and no treatment-related deaths occurred. The Child-Pugh score did not increase by >2 points after the start of therapy. In 78% and 75% of all patients, the score did not increase by >1 point in the early and late phase, respectively. The overall tumor response rate was 71%. The local control and overall survival rate was 92% and 92%, 81% and 50%, and 81% and 25% at 1, 3, and 5 years, respectively. CONCLUSION: Carbon ion radiotherapy appears safe and effective for patients with hepatocellular carcinoma. Additional clinical studies using a larger subject group are required to confirm the therapeutic efficacy. PMID- 15275735 TI - A population-based study of the prevalence and influence of gifts to radiation oncologists from pharmaceutical companies and medical equipment manufacturers. AB - PURPOSE: Hospital-based physicians are responsible for the purchase of expensive equipment. Little is known about the influence of gift giving on their behavior. We wanted to ascertain the prevalence of gift giving from the pharmaceutical industry and medical equipment manufacturers to radiation oncologists and determine whether or not the size of accepted gifts influences their opinions regarding gifts. METHODS AND MATERIALS: A population-based survey of hospital based physicians conducted between 2002 and 2003. The study population consisted of all radiation oncologists who were members of the American Society of Therapeutic Radiology and Oncology between 2000 and 2001. A random number generator was used to identify 20% of the population. This group was invited by e mail and conventional mail to complete a Likert scale questionnaire. Those asked to complete the questionnaire electronically were directed to a specially designed web site. RESULTS: Of 640 individuals who were asked to participate, 241 (38%) completed the questionnaire. 96% admitted accepting gifts. The most commonly accepted low value gifts were: pen or pencil (78%), drug samples for patient's use (70%), meal (66%), and a note pad (59%). The most commonly accepted high value gifts were trips to "equipment-users meetings" (15%), honoraria for speaking at a conference (10%), and participation in a conference call (9%). Only 5% of radiation oncologists agreed with the statement "my prescribing practices are affected" by gifts; however, 33% agreed with the statement "I believe that other physicians prescribing practices are affected." Similarly, although only 4% felt that their recommendations concerning purchases of medical equipment are affected by gifts, 19% felt that other physicians would be influenced. A test of the hypothesis that physicians believe that their conduct is less affected than those of their colleagues (i.e., "I am not influenced by gifts but someone else is" was strongly affirmed by a correlation statistic) (p < 0.0001). Of the radiation oncologists surveyed, 74% felt that they should be free to accept gifts of small value, 31% felt they should be free to accept meals or gifts of any type, 16% felt that residency programs should ban free meals provided by companies, 13% felt professional associations should discourage companies from hosting parties at the annual meeting, 17% felt that gift giving should stop, and 66% agreed that clinical information provided by companies provides a useful continuing medical education service. Those who accepted larger gifts were far more likely to disagree with statements such as "professional societies should actively discourage companies from hosting parties and providing free meals and giving gifts to physicians attending the annual meeting" (p = 0.0003) and "the practice of gift giving by companies should stop" (p = 0.0017); they were slightly more likely to agree with statements such as "clinical information provided to radiation oncologists by companies provides a useful continuing medical education service." CONCLUSIONS: To our knowledge, this study represents the first large-scale population based study of a hospital-based specialty and gift giving. This study demonstrates that: (1) Gift giving in radiation oncology is endemic. (2) Although each physician is likely to consider himself or herself immune from being influenced by gift giving, he or she is suspicious that the "next person" is influenced. (3) There is a correlation between the willingness of individual physician to accept gifts of high value and their sympathy toward this practice. PMID- 15275736 TI - Recovery from sublethal damage during intermittent exposures in cultured tumor cells: implications for dose modification in radiosurgery and IMRT. AB - PURPOSE: In stereotactic irradiation using a linear accelerator and intensity modulated radiation therapy (IMRT), radiation is administered intermittently, and 30 min or longer is often required in one treatment session. The purpose of the present study was to determine how different the dose delivered with such intermissions is from that delivered continuously and to estimate dose-modifying factors. METHODS AND MATERIALS: Murine EMT6 and SCCVII cells in culture were used. First, two doses of 4 Gy were given with an interruption of 15 min to 6 h or 1-10 min, or without interruption. Next, five fractions of 1.6 Gy were given with interfraction intervals of 1 to 5 min each. Doses of 6.5-8 Gy were also given without interruption to estimate dose-modifying factors. Cell survival was determined by a colony assay. Furthermore, a total dose of 2 Gy was given in 5 or 10 even fractions at intervals of 1-5 min or 30 s-3 min each, respectively, and the results were compared with those obtained after 1.6-2 Gy delivered continuously by using a cytokinesis-block micronucleus assay. RESULTS: In the two fraction experiments, a significant increase in cell survival resulting from recovery from sublethal damage (SLDR) was observed when the interruption time was 2 min or longer in EMT6 cells and 3 min or longer in SCCVII. With a 5-min interval, cell survival increased by 13% in EMT6 and by 18% in SCCVII. In the five-fraction experiments, SLDR was evident when the interfraction interval was 2 min or longer in both 8-Gy and 2-Gy dose experiments. In the 10-fraction experiment using a dose of 2 Gy, SLDR was evident when the interfraction interval was 1 min or longer. In the 5-fraction and 10-fraction experiments, the dose modifying factors were between 1.08 and 1.16 when the total time for irradiation was between 20 and 30 min. CONCLUSIONS: The effects of stereotactic radiosurgery and IMRT that require considerably long beam interruption (e.g., 8 min or longer in total) may be less than those of the same dose administered continuously. In treatments that take 20 min or longer, dose modification appears necessary based on biologically estimated dose-modifying factors. PMID- 15275737 TI - Cluster models of dose-volume effects. AB - PURPOSE: Describe cluster models, normal-tissue complication probability models in which both the number and the spatial location of radiation-sterilized functional subunits play a role in defining complication probability. METHODS AND MATERIALS: Computer simulation was used to determine the maximum size cluster of sterilized subunits associated with a given dose distribution. Complications were associated with large clusters. RESULTS: Cluster models showed a volume effect, as increasing effect for constant dose when the volume increased or constant effect when the dose was reduced with increasing volume. Cluster models gave similar results to existing models when tissues were irradiated uniformly. With inhomogeneous dose distributions, on the other hand, different spatial distributions of "hot spots" may lead to different predictions of complication probability by cluster models. The result was that a higher complication probability resulted when hot spots were contiguous (clustered) than when they were dispersed, even if both situations are characterized by the same dose-volume histogram. A potential advantage of cluster models is to provide an easy, internally consistent way to predict complications arising from the inhomogeneous dose distributions that sometimes arise with intensity-modulated radiotherapy. CONCLUSION: Cluster models offer a new way to quantify complication probability in treatment situations in which a wide variety of hot-spot distributions occur. PMID- 15275738 TI - X-irradiation to human malignant glioma cells enhances the cytotoxicity of autologous killer lymphocytes under specific conditions. AB - PURPOSE: To clarify the effect of combining x-irradiation and human killer lymphocytes against autologous malignant glioma cells, we analyzed not only the alteration of surface antigen expression in irradiated tumor cells, but also the cytotoxic effects of human killer lymphocytes on the autologous tumor cells with and without x-irradiation. METHODS AND MATERIALS: Six malignant glioma cell-lines (MG 1-6) established from each patient with a malignant glioma in our institute, and U87MG, were used as materials. They were irradiated by 0-50 Gy of X-rays, and the alternations of their human histocompatability leukocyte antigen (HLA), HLA ABC, HLA-DR, -DP, -DQ, and FAS expressions were examined. Then, three sets of autologous natural killer (NK) cells, and autologous tumor-specific T lymphocytes (ATTL) were induced from the peripheral blood mononuclear cells (PBMCs) of three patients, and in vitro cytotoxic effects of these killer cells on the irradiated autologous tumor cells were analyzed. RESULTS: Irradiation-enhanced HLA-DR, -DP, DQ, and FAS expression in glioma cell lines with low p53 expression. However, there was no correlation between HLA-ABC expression and X-ray dose. After irradiation of the tumor cells, cytotoxicity was enhanced in four of six effectors; in particular, it was significantly elevated in two killer lymphocytes. It was speculated that the enhancing effect was influenced not only by the p53 status of the tumor, but also by the types of killer lymphocytes; the alteration of cytotoxicity in NK cells on irradiated tumor cells may be compensatory for alteration in ATTLs. CONCLUSION: It was indicated that irradiation of malignant tumor cells enhanced killer cell-mediated cytotoxicity in autologous models under specific conditions. These basic data should contribute to clinical trials using local radiotherapy and systemic adoptive immunotherapy with killer lymphocytes. PMID- 15275739 TI - Simultaneous integrated boost for breast cancer using IMRT: a radiobiological and treatment planning study. AB - PURPOSE: The purpose of this work is to explore the possibility of using intensity-modulated radiation therapy (IMRT) to deliver the boost dose to the tumor bed simultaneously with the whole-breast IMRT to reduce the radiation treatment time by 1-2 weeks. METHODS AND MATERIALS: The biologically effective dose (BED) for different treatments was calculated using the linear-quadratic (LQ) model with parameters previously derived for breast cancer from clinical data (alpha/beta = 10Gy, alpha = 0.3Gy(-1)). A potential doubling time of 15 days (from in vitro measurements) for breast cancer and a generic alpha/beta ratio of 3 Gy for normal tissues were used. A series of regimens that use IMRT as initial treatment and an IMRT simultaneous integrated boost (SIB) were derived using biologic equivalence to conventional schedules. Possible treatment plans with IMRT SIB to the tumor bed were generated for 2 selected breast patients, 1 with a shallow tumor and 1 with a deep-seated tumor. Plans with a simultaneous integrated electron boost were also generated for comparison. Dosimetric merits of these plans were evaluated based on dose volume histograms. RESULTS: A commonly used conventional treatment of 45 Gy (1.8 Gy x 25) to the whole breast and then a boost of 20 Gy (2 Gy x 10) is biologically equivalent to an alternative plan of 1.8 Gy x 25 to the whole breast with a 2.4 Gy x 25 SIB to the tumor bed. The new regime reduces treatment time from 7 to 5 weeks. For the patient with a deep-seated tumor, the IMRT plans reduce the volume of the breast that receives high doses (compared with the conventional photon boost plan) and provides good coverage of the target volumes. CONCLUSION: It is biologically and dosimetrically feasible to reduce the overall treatment time for breast radiotherapy by using an IMRT simultaneous integrated boost. For selected patient groups, IMRT plans with a new regimen can be equal to or better than conventional plans. PMID- 15275740 TI - Improved treatment of pelvis and inguinal nodes using modified segmental boost technique: dosimetric evaluation. AB - PURPOSE: To describe a novel, yet simple, modified segmental boost technique (MSBT) and to compare the dosimetry of our method with that of other traditional methods of treatment for the pelvis and inguinal nodes. METHODS AND MATERIALS: We developed a radiotherapy technique that uses linear accelerators with multileaf collimators to treat the pelvis and sequentially boost the inguinal regions, while minimizing "hot spots" across the match-line. This was achieved by angling the gantry for the inguinal fields so that their medial borders aligned with the divergence of the posterior pelvic field. Film dosimetry was performed to compare the MSBT with the traditional segmental boost technique, partial transmission block, and photon/electron combination techniques. These treatment techniques were scored on the basis of the dose homogeneity index, defined as the ratio of match-line maximum dose to the average dose at a given depth in the groin treatment area. RESULTS: The values of the dose homogeneity index were the same (1.04) for MSBT and partial transmission block, and 1.21, 1.39, and 1.18 for the segmental boost technique, photon pelvis with electron tags, and photon pelvis with electron boost, respectively. CONCLUSION: The MSBT proved to be technically simple while optimizing dose homogeneity compared with the other techniques and allows for maximum use of the features of modern linear accelerators. PMID- 15275741 TI - An immobilization system for claustrophobic patients in head-and-neck intensity modulated radiation therapy. AB - PURPOSE: To evaluate the effectiveness of an immobilization treatment system used for claustrophobic patients in head-and-neck intensity-modulated radiation therapy (IMRT). METHODS AND MATERIALS: Instead of the thermoplastic facemask, the Vac Fix (S & S Par Scientific, Odense, Denmark) mold is used for immobilization of claustrophobic patients at the University of Florida in head-and-neck IMRT. The immobilization procedure combines the use of commercial stereotactic infrared (IR) ExacTrac camera system (BrainLAB, Inc., Westchester, IL) for patient setup and monitoring. The Vac Fix mold is placed on the headrest and folded up as needed to provide support before the mold is hardened. For the camera system, a frame referred to as a "tattoo-free immobilization accessory" is fabricated, on which the IR markers can be placed. A patient-specific dental impression is made with the bite tray. The movement of the markers, connected through the dental impression of the patient, accurately represents the overall patient motion. Patient movement is continuously monitored and repositioning is performed whenever patient movement exceeds the predefined tolerance limit. Monitored patient movements are recorded at a certain frequency. Recorded data are analyzed and compared with those of patients immobilized with the thermoplastic facemask plus the camera system that is the standard immobilization system in our clinic. RESULTS: For three patients treated with the Vac Fix mold plus the camera system, on average, the histogram-based uncertainties, U(95)(5), U(95)(20), and mean displacement, R(mean) (mm) were 1.03, 1.08, and 0.60, respectively. These values are close to those obtained with the mask plus the camera system. The Vac Fix mold plus the camera system often requires more beam interruptions because of repositioning than the mask plus the camera system (on average, the Vac Fix mold plus the camera system required repositioning 7.7 times and the mask plus the camera system required repositioning 1.8 times during 20 treatments). CONCLUSION: The Vac Fix mold immobilization procedure plus the camera monitoring system has been set up for patients who are claustrophobic or cannot tolerate a mask during head-and-neck IMRT. Although this system causes more frequent beam delivery interruptions, it is as effective as the mask plus the camera system in immobilizing patients within the tolerance limit. PMID- 15275742 TI - Dose verification in clinical IMRT prostate incidents. AB - PURPOSE: In view of the need for dose-validation procedures on each individual intensity-modulated radiation therapy (IMRT) plan, dose-verification measurements by film, by ionization chamber, and by polymer gel-MRI dosimetry were performed for a prostate-treatment plan configuration. Treatment planning system (TPS) calculations were evaluated against dose measurements. METHODS AND MATERIALS: Intensity-modulated radiation therapy (IMRT) treatments were planned on a commercial TPS. Kodak EDR-2 films were used for the verification of two dimensional (2D) dose distributions at 1 coronal and 5 axial planes in a water equivalent phantom. Full three-dimensional (3D) dose distributions were measured by use of a novel polymer gel formulation and a 3D magnetic resonance imaging (MRI) readout technique. Calculations were compared against measurements by means of isocontour maps, gamma-index maps (3% dose difference, 3-mm distance to agreement) and dose-volume histograms. RESULTS: A good agreement was found between film measurements and TPS predictions for points within the 60% isocontour, for all the examined plans (gamma-index <1 for 96% of pixels). Three dimensional dose distributions obtained with the polymer gel-MRI method were adequately matched with corresponding TPS calculations, for measurements in a gel phantom covering the planning-target volume (PTV). CONCLUSIONS: Measured 2D and 3D dose distributions suggest that, for the investigated prostate IMRT plan configuration, TPS calculations provide clinically acceptable accuracy. PMID- 15275743 TI - MLC leaf width impact on the clinical dose distribution: a Monte Carlo approach. AB - PURPOSE: The influence of the multileaf collimator (MLC) leaf width on the dose distribution in patients treated with conformal radiotherapy and intensity modulated radiotherapy has been analyzed. This study was based on the Monte Carlo simulation with the beams generated by a linac with the double-focused MLC. MATERIALS AND METHODS: The transmission through the leaves and the exact shape of the penumbra regions are difficult to model by treatment planning system algorithms. An accurate assessment of the dose variations due to the leaf width change can be achieved by means of Monte Carlo simulation. The BEAM/EGS4 code was used at the Hospital of the Virgen Macarena to model a Siemens PRIMUS linac, featuring an MLC with a leaf width projecting 1 cm at the isocenter. Based on this real model, a virtual head was designed while allowing for a variation of the leaf width projection. Both the real linac and the virtual linac, with leaves projecting 0.5 cm, were used to obtain the dose distributions for several treatments. A few disease sites, including the prostate, head and neck, and endometrium, were selected for the design of the conformal and intensity modulated radiotherapy treatments with a forward planning algorithm sensitive to the different shapes of the volumes of interest. Isodose curves, differential matrix, gamma function, and the dose-volume histograms (DVHs) corresponding to both MLC models were obtained for all cases. The tumor control probability and the normal tissue complication probability were derived for those cases studied featuring the greatest differences between results for both MLCs. RESULTS: The impact on the DVHs of changing leaf width projections at the isocenter from 1.0 cm to 0.5 cm was low. Radiobiologic models showed slightly better tumor control probability/normal tissue complication probability values using the virtual MLC with a leaf width projecting 0.5 cm at isocenter in those cases presenting greater differences in the DVHs. CONCLUSIONS: The impact on the clinical dose distribution due to the MLC leaf width change is low based on the design and conditions used in this study. PMID- 15275745 TI - Human and animal experimental models of acute and chronic muscle pain: intramuscular algesic injection. PMID- 15275746 TI - Complex regional pain syndrome-like symptoms during herpes zoster. AB - Complex Regional Pain Syndrome (CRPS) associated with herpes zoster (HZ) was first reported by Sudeck in 1901 (Sudeck, 1901) and is recognized clinically. However, only 13 cases have been published in the literature, and nothing is known about the incidence, prevalence, or natural history (Chester, 1992; Foster et al., 1989; Grosslight et al., 1986; Ketz and Schliack,1968; Kishimoto et al., 1995; Querol and Cisneros, 2001; Sudeck, 1901; Visitsunthorn and Prete, 1981). The aim of the present study was to determine the prevalence of CRPS-like symptoms in a prospectively gathered cohort of subjects with HZ and to follow the natural history of their pain and sensory disturbance during the first 6 months after onset of HZ. Subjects were evaluated at four time points after HZ: 2-6 weeks, 6-8 weeks, 3 months, and 6 months. Only subjects aged 50 or older with pain VAS ratings of >/=20/100 at 2-6 weeks were eligible. The first (screening) visit included a neurological and physical examination that was updated at each subsequent visit. Assessments included ratings of pain intensity, allodynia severity, and rash severity. The neurological exam included determination of presence or absence of the following CRPS-like symptoms: (1) increased sweating, (2) color changes, (3) skin temperature changes, (4) weakness of the affected area based on physical exam, (5) edema, and (6) extension of CRPS-like symptoms outside the affected dermatome. For subjects with HZ in dermatomes that can include the limbs (C4-T2 and L1-S2), extremity involvement was considered present if allodynia or rash extended beyond the neck of the humerus (upper extremity), the inguinal ligament (anterior lower extremity), or gluteal sulcus (posterior lower extremity). Involvement of the extremity was considered proximal if neither HZ rash nor allodynia extended past the elbow (upper extremity) or knee (lower extremity). Of the first 75 subjects recruited, 25 had HZ outbreaks in dermatomes that extended into the extremities (C4-T2 and L1-S2). In this group, 8 subjects had no extremity involvement, 8 had proximal extremity involvement, and 9 had distal extremity involvement. Subjects with distal extremity HZ reported more pain across the four visits (p < 0.05). At 3 months, more subjects with distal extremity involvement met criteria for PHN (8 out of 9, 89%), while only 4 out of 8 (50%) with proximal involvement and 2 out of 8 (25%) of subjects without extremity involvement met criteria for PHN (Chi-square test: p < 0.05). Only 25 out of the remaining 50 (50%) subjects with outbreaks in dermatomes that do not include the extremities met criteria for PHN at 3 months (Chi-square test: p < 0.05). Six months after onset of HZ, 6 out of 9 subjects with distal extremity involvement met PHN criteria compared with 2 out of 8 (25%) with proximal involvement and 2 out of 8 (25%) without extremity involvement (Chi-square test: p = 0.12). Fifteen out of 50 (30%) subjects with outbreaks in dermatomes that do not include the extremities met criteria for PHN (Chi-square test: p < 0.05). No subject had all six CRPS-like symptoms. Of the 17 subjects with extremity involvement, 9 subjects had '0-2 CRPS-like symptoms' and 8 had '3-5 CRPS-like symptoms'. None of the eight subjects without extremity involvement had any CRPS like symptoms. Of the 50 subjects with HZ outside the extremity, only one had abdominal weakness. Pain ratings were higher in subjects with '3-5 CRPS-like symptoms'. More subjects with '3-5 CRPS-like symptoms' met criteria for PHN at 3 months (7 out of 8, 88%), compared to 5 out of 9 (55%) of subjects with '0-2 CRPS like symptoms' (p = 0.07). At 6 months, 2 out of 9 (22%) of subjects with '0-2 CRPS-like symptoms' met criteria for PHN, compared with 6 out of 8 (75%) of subjects with '3-5 CRPS-like symptoms' (Chi-square test: p < 0.03). Two case reports are presented. In summary, the occurrence of CRPS-like symptoms is common in subjects with HZ outbreaks affecting the extremity, particularly if the distal extremity is involved. It is uncertain if the pathophysiology underlying the CRPS like symptoms observed in this study is similar to that of CRPS from other causes, or if it is relatively specific to HZ. Development of PHN is common in subjects who have experienced CRPS-like symptoms. More aggressive preventive treatments may be justified in this high-risk subset of HZ subjects to prevent development of PHN. Prospective randomized controlled studies are needed to determine which subjects are most likely to benefit and when treatment should begin. PMID- 15275747 TI - Pro-nociceptive role of peripheral galanin in inflammatory pain. AB - We investigated the peripheral function of galanin (GAL) in capsaicin (CAP) induced inflammatory pain. Intraplantar GAL (0.1 ng/microl) alone does not produce nociceptive behaviors. However, ipsilateral but not contralateral GAL at low doses (0.1 ng/microl) significantly increases CAP-evoked nociceptive behaviors approximately twofold. This effect is attributed to activation of peripheral GAL receptor 2 (GalR2) because a selective GalR2 agonist (AR-M1896) mimics the pro-nociceptive actions of GAL. Recording from nociceptors confirms that GAL does not modify activity of nociceptors but markedly enhances CAP induced excitation of these fibers. CAP produces a discharge rate of 0.15+/-0.05 impulses/s which increases to 0.54+/-0.17 impulses/s following CAP+GAL. Immunohistochemical studies indicate GalR2 are highly expressed (65.8%) in L5 dorsal root ganglion (DRG) cells. Also, 44.5% GalR2-positive DRG neurons label for the capsaicin receptor (vanilloid receptor 1, VR1) while 61.7% of VR1 positive DRG neurons label for GalR2; 28.1% of total DRG neurons are double labeled supporting the hypothesis that GAL-induced effects are mediated by GalR2 on capsaicin-sensitive primary afferents. Furthermore, 68.0% unmyelinated and 23.1% myelinated digital nerve axons label for GalR2, indicating the receptor is transported out to the periphery. Immunostaining for GAL peptide in digital nerves labels 46.4% unmyelinated and 27.1% myelinated axons, suggesting that afferents are a major source of ligand for peripheral GalR2. These results suggest that peripheral GAL has an excitatory role in inflammatory pain, likely mediated by peripheral GalR2 and that GAL can modulate VR1 function. PMID- 15275749 TI - Pre-operative and post-operative effect of a pain management programme prior to total hip replacement: a randomized controlled trial. AB - Patients may wait some time for total hip replacement with conservative management of pain and disability, but no attempts to rehabilitate them. This study randomised 40 patients accepted for and awaiting total hip replacement to a brief rehabilitative psychologically based pain management programme (PMP) or to a control group with no intervention. Patients were assessed before randomisation, 3 months after the PMP or equivalent waiting time, and again one year later after total hip replacement. Assessments included pain, impact of pain (Arthritis Impact Scale: AIMS), mobility (timed walk), sleep and analgesic consumption. The patients who had learned pain management reported lower pain intensity and pain distress and less sleep disturbance than waiting list controls, but no improvement in function or analgesic consumption. Six patients opted to delay, but this did not differ between groups. Post-hip replacement both groups improved in pain and some aspects of activity (AIMS) with greater improvement in the PMP group for physical activity and total AIMS scores, suggesting that some techniques had continued to be of use post-surgically. Rehabilitative pain management may be useful to patients pre-operatively in managing everyday pain, but not to the extent that they opt to delay surgery; it may also improve their function after hip replacement. PMID- 15275748 TI - Antihyperalgesic effects of cizolirtine in diabetic rats: behavioral and biochemical studies. AB - Although clinically well controlled at the metabolic level, type I diabetes resulting from an insufficient insulin secretion remains the cause of severe complications. In particular, diabetes can be associated with neuropathic pain which fails to be treated by classical analgesics. In this study, we investigated the efficacy of a novel non opioid analgesic, cizolirtine, to reduce mechanical hyperalgesia associated with streptozotocin (STZ)-induced diabetes, in the rat. Cizolirtine was compared to paroxetine, an antidepressant drug with proven efficacy to relieve painful diabetic neuropathy. Under acute conditions, cizolirtine (30 and 80 mg/kgi.p.) significantly increased paw withdrawal and vocalization thresholds in the paw pressure test in diabetic rats displaying mechanical hyperalgesia. The antihyperalgesic effects of cizolirtine persisted under chronic treatment conditions, since pre-diabetes thresholds were recovered after a two week-treatment with the drug (3 mg/kg/day, s.c.). In this respect, cizolirtine was as efficient as paroxetine (5 mg/kg per day, s.c.) which, however, was inactive under acute treatment conditions. Measurements of the spinal release of calcitonin gene-related peptide (CGRP) through intrathecal perfusion under halothane-anesthesia showed that acute administration of cizolirtine (80 mg/kg, i.p.) significantly diminished (-36%) the peptide outflow in diabetic rats suffering from neuropathic pain. This effect as well as the antihyperalgesic effect of cizolirtine were prevented by the alpha(2) adrenoreceptor antagonist idazoxan (2 mg/kg, i.p.). These data suggest that the antihyperalgesic effect of cizolirtine in diabetic rats suffering from neuropathic pain implies an alpha(2)-adrenoceptor-dependent presynaptic inhibition of CGRP-containing primary afferent fibers. PMID- 15275750 TI - P300-amplitudes in upper limb amputees with and without phantom limb pain in a visual oddball paradigm. AB - The aim of the study was to investigate to what extent cortical hyper-reactivity to visual stimuli is present in upper limb amputees. Five amputees with phantom limb pain (PLP), five amputees without PLP (Non-PLP) and 10 healthy controls (HC) were investigated using a visual oddball paradigm. Two hundred visual stimuli were presented with target stimuli occurring at a probability of 25% and standard stimuli at a probability of 75%. Event-related potentials were recorded from nine scalp positions (F3, F4, Fz, C3, C4, Cz, P3, P4, Pz). The PLP-patients had significantly higher P300-amplitudes to both types of stimuli compared to the non PLP-patients. The HC were not significantly different from both amputee groups. P300-amplitude to targets at frontal sites in the hemisphere contralateral to the amputation was higher in the PLP patients. P300-latencies to target stimuli differed only at frontal sites with PLP-patients showing significantly longer latencies than non-PLP-patients. To standard stimuli, however, they showed significantly shorter latencies at central and parietal scalp positions. The HC had significantly shorter latencies than both amputee groups. The size of the P300-amplitude was positively correlated with the intensity of PLP. These findings suggest a higher magnitude of non-specific cortical excitability in amputees with PLP and a reduced excitability in amputees without PLP. This extends previous findings of differences in cortical excitability in PLP and non PLP patients in the sensorimotor domain. PMID- 15275752 TI - Attenuation of mechanical allodynia by clinically utilized drugs in a rat chemotherapy-induced neuropathic pain model. AB - Chemotherapy-induced peripheral neuropathy is a common, dose-limiting side effect of cancer chemotherapeutic agents, including the vinca alkaloids such as vincristine. The resulting symptoms, which frequently include moderate to severe pain, can often be disabling. The current study utilized a vincristine-induced neuropathic pain animal model [Pain 93 (2001) 69], in which rats were surgically implanted with mini-osmotic pumps set to deliver vincristine sulfate (30 microg kg(-1)day(-1), i.v.), to examine the time course of progression of various pain modalities and to compare the dose-response effects of clinically utilized drugs on mechanical allodynia to further validate the relevance of this model to clinical pathology. Vincristine infusion resulted in significant cold allodynia after 1 week post-infusion, however mechanical and thermal nociception showed little to no effect. In contrast, marked mechanical allodynia occurred by 1 week of vincristine infusion and returned nearly to pre-infusion levels by the 4th week after infusion pump implantation. ED(50) values (micromol/kg, p.o.) were determined in the mechanical allodynia assay for lamotrigine (82), dextromethorphan (94), gabapentin (400), acetaminophen (1100) and carbamazepine (3600); however, aspirin and ibuprofen had no effects up to 300 and 1000 micromol/kg, respectively. Additionally, ED(50) values (micromol/kg, i.p.) were determined in the mechanical allodynia assay for clonidine (0.35) and morphine (0.62), but desipramine and celecoxib had no effects up to 66 and 260 micromol/kg, respectively. Findings from the current, preclinical study further validate this model as clinically relevant for chemotherapy-induced pain. The surprisingly good effects observed with acetaminophen warrant further investigation of its mechanism(s) of action in neuropathic pain. PMID- 15275751 TI - A randomized clinical trial of the effectiveness of a scheduled oral analgesic dosing regimen for the management of postoperative pain in children following tonsillectomy. AB - The purpose of this study was to determine whether around-the-clock (i.e. ATC) dosing of acetaminophen with codeine, with or without nurse coaching, compared to standard care with as needed (i.e. PRN) dosing: reduced children's reports of pain intensity with and without swallowing; increased pain relief, and increased analgesic consumption. Eighty children, 6-15 years, undergoing tonsillectomy were randomized to one of three treatment groups to receive acetaminophen with codeine (120 mg/12 mg/5 ml) for 3 days after surgery: PRN group (N = 28)-every 4 h PRN, with standard postoperative instructions, without nurse coaching; ATC group (N = 26)-every 4 h ATC, with standard postoperative instructions, without nurse coaching; and ATC+coaching group (N = 26)-every 4 h ATC, with standard postoperative instructions and nurse coaching. In all three groups, significant decreases were found over time in pain intensity scores at rest (P < 0.001) and with swallowing (P < 0.001). However, mean pain scores at rest and with swallowing were >3/10 until the fourth evening after tonsillectomy. Children in both ATC dosing groups received significantly greater amounts of acetaminophen and codeine than children in the PRN group (P < 0.003). No significant differences were found in the amount of analgesic administered between the ATC dosing groups with and without nurse coaching. No significant differences were found in the amount of nausea and vomiting among the three groups. Scheduled dosing of acetaminophen with codeine did not provide adequate pain relief for children following tonsillectomy. Nurse coaching does not increase parent's adherence with an ATC dosing schedule. PMID- 15275753 TI - Mustard oil has differential effects on the response of trigeminal caudalis neurons to heat and acidity. AB - Topical application of mustard oil (allyl isothiocyanate) to the skin or injection into joints induces hyperalgesia, allodynia, and neuroinflammation. However, when applied to the oral or nasal mucosa, mustard oil evokes a desensitizing pattern of irritation. Presently we investigated the responses of neurons in superficial laminae of trigeminal subnucleus caudalis (Vc) to noxious thermal (53 degrees C) and chemical (pentanoic acid; 200 mM) stimuli prior to and following lingual mustard oil application. A low concentration of mustard oil (0.125%) applied by constant flow (0.5 ml/min; 15 min), initially excited Vc neurons followed by partial desensitization. Responses to noxious heat were unchanged following mustard oil. A high concentration of mustard oil (1.25%) initially excited Vc neurons followed quickly (within 20 s) by nearly complete desensitization. The desensitization was transient since reapplication of mustard oil approximately 20 min later elicited a comparable response that also rapidly desensitized. Mustard oil also transiently cross-desensitized Vc responses to pentanoic acid (to 52%), in striking contrast to noxious heat-evoked responses which were significantly sensitized to approximately 160% of pre-mustard oil levels. The data suggest that the effect of mustard oil on subsequent lingual nociceptive responses is concentration dependent, transient, and modality specific. PMID- 15275754 TI - Sex differences in temporal characteristics of descending inhibitory control: an evaluation using repeated bilateral experimental induction of muscle pain. AB - Little is known about sex differences in the temporal pattern of descending inhibitory mechanisms, such as descending noxious inhibitory control (DNIC). Sex differences in temporal characteristics of DNIC were investigated by measuring pressure pain thresholds (PPTs) over time in the trapezius muscles (local pain areas) and the posterolateral neck muscles (referred pain areas) following repeated bilateral injection of hypertonic versus isotonic saline into both trapezius muscles. Ten females and 11 males received two consecutive bilateral injections, with 15 min interval, of either 5.8% hypertonic saline (0.5 ml in each side for each bilateral injection) or isotonic saline as a control in a randomized manner. Following hypertonic saline injection, the maximal pain intensities of the first and second bilateral injections were significantly higher in females than in males. The PPTs in the trapezius muscles were significantly lower in females than in males. Significantly higher PPTs (hypoalgesia) in men than in women were shown 15 min after the first bilateral injection, and 7.5 and 15 min after the second bilateral injection in the referred pain areas. Importantly, the second bilateral injection failed to further increase the PPTs for both sexes. These results showed that there were sex differences in temporal characteristics of descending inhibition with long lasting hypoalgesia in men than in women. Repeated noxious muscular stimuli may inhibit further build-up of DNIC, which may reflect a mechanism of plasticity of the descending inhibitory systems following recurrent nociceptive barrage for both sexes. PMID- 15275755 TI - Peripheral P2X receptors and nociception: interactions with biogenic amine systems. AB - ATP is implicated in peripheral nociception following activation of P2X, and particularly P2X(3) receptors. The present study examined interactions between alphabeta-methylene-ATP (a P2X(3) receptor ligand) and 5-hydroxytryptamine (5 HT), noradrenaline (NA) and histamine, following local administration into the hindpaw, on spontaneous pain behaviors and thermal hyperalgesia in Sprague-Dawley rats. The interaction with NA was further explored using systemic 6 hydroxydopamine (6-OHDA) and locally administered indomethacin. alphabeta methylene-ATP produced no spontaneous pain behaviors. Coadministration of 5-HT with alphabeta-methylene-ATP mildly augmented flinching behaviors, while histamine had no such effect. Coadministration of NA with alphabeta-methylene-ATP produced a pronounced expression of flinching and biting/licking behaviors. alphabeta-Methylene-ATP, given alone, produced thermal hyperalgesia, and this was markedly augmented by both 5-HT and NA, but not histamine. 6-OHDA (neurotoxin for sympathetic neurons) and indomethacin (cyclooxygenase inhibitor) reduced the augmenting effect of NA on alphabeta-methylene-ATP-induced thermal hyperalgesia, but had no effect on spontaneous pain behaviors produced by the alphabeta methylene-ATP/NA combination. Effects of alphabeta-methylene-ATP, NA and their combination were also examined in Long Evans and Wistar rats. In both strains, alphabeta-methylene-ATP and NA both individually led to significant intrinsic flinching behaviors, and the effect of their combination was even more pronounced than in Sprague-Dawley rats. These results provide evidence for: (a) a strong enhancement by NA and 5-HT of nociception produced by peripheral P2X receptors in Sprague-Dawley rats, (b) an indirect action of NA, via sympathetic efferents and prostanoids, with thermal hyperalgesia, and (c) a greater expression of spontaneous pain behaviors with alphabeta-methylene-ATP and NA alone, and with their combination, in Wistar and Long Evans rats compared to Sprague-Dawley rats. PMID- 15275756 TI - Neuroelectric source imaging of steady-state movement-related cortical potentials in human upper extremity amputees with and without phantom limb pain. AB - Whereas several studies reported a close relationship between changes in the somatotopic organization of primary somatosensory cortex and phantom limb pain, the relationship between alterations in the motor cortex and amputation-related phenomena has not yet been explored in detail. This study used steady-state movement-related cortical potentials (MRCPs) combined with neuroelectric source imaging to assess the relationship of changes in motor cortex and amputation related phenomena such as painful and non-painful phantom and residual limb sensations, telescoping, and prosthesis use. Eight upper limb amputees were investigated. A significant positive relationship between reorganization of the motor cortex (distance of the MRCP source location from the mirrored source for hand movement) and phantom limb pain was found. Non-painful phantom sensations as well as painful and non-painful residual limb sensations were unrelated to motor cortical reorganization. A higher amount of motor reorganization was associated with less daily prosthesis use, which also tended to be related to more severe phantom limb pain. These results extend previous findings of a positive relationship between somatosensory reorganization and phantom limb pain to the motor domain and suggest a potential positive effect of prosthesis use on phantom limb pain and cortical reorganization. PMID- 15275757 TI - Pain-related catastrophizing: a daily process study. AB - Little is known about the extent to which individuals vary versus remain stable in their pain-related catastrophizing, or to which catastrophizing is associated with pain and related problems on a daily basis. We used daily electronic interviews to examine the: (1) reliability and validity of a brief daily catastrophizing measure; (2) stability of catastrophizing; (3) patient characteristics associated with catastrophizing; (4) associations between catastrophizing and concurrent and subsequent outcomes (pain, activity interference, jaw use limitations, and negative mood), between and within patients; and (5) associations between pain and subsequent catastrophizing. One hundred patients with chronic temporomandibular disorder pain completed electronic interviews three times a day for 2 weeks [mean (SD) number of interviews=46 (15)]. The catastrophizing scale had high internal consistency (Cronbach's alpha = 0.95) and validity (r = 0.65 with the Coping Strategy Questionnaire Catastrophizing scale), and catastrophizing was stable (ICC=0.72) over time. Younger age and greater baseline depression, pain, and disability predicted greater daily catastrophizing. Daily catastrophizing was associated significantly with concurrent outcomes, between- and within-subjects (P < 0.001); however, associations with same-day subsequent outcomes were greatly attenuated after adjusting for prior outcome levels. Similarly, daily pain was associated significantly with subsequent catastrophizing, but this association was no longer statistically significant after adjusting for prior catastrophizing. The data indicate that catastrophizing is stable over short periods of time in the absence of substantial change in pain, and that within patients, times of greater catastrophizing are associated with worse pain, disability, and mood. PMID- 15275758 TI - Block of NMDA and non-NMDA receptor activation results in reduced background and evoked activity of central amygdala neurons in a model of arthritic pain. AB - The latero-capsular division of the central nucleus of the amygdala (CeA) is now defined as the 'nociceptive amygdala' because of its high content of neurons activated exclusively or preferentially by noxious stimuli. Multireceptive (MR) neurons that respond to innocuous and, more strongly, to noxious stimuli become sensitized in arthritis pain. This form of nociceptive plasticity involves presynaptic group I metabotropic glutamate receptors, which increase glutamate release. Here we address the role of N-methyl-D-aspartate (NMDA) and non-NMDA receptors. Extracellular single-unit recordings were made from 25 CeA neurons in anesthetized rats. The neurons' responses to graded brief (15 s) mechanical stimuli, background activity, receptive field size and threshold were measured before and after the induction of kaolin/carrageenan arthritis in one knee and before and during drug applications into the CeA by microdialysis. All neurons examined received excitatory input from the knee(s) and were MR neurons. A selective NMDA receptor antagonist (AP5) inhibited responses to noxious stimuli more potently in the arthritic pain state (n = 6) than under control conditions before arthritis (n = 8) AP5 also inhibited the enhanced background activity and increased responses to normally innocuous stimuli in arthritis, but had no significant effects on these parameters under control conditions. A selective non NMDA receptor antagonist (NBQX) inhibited background activity and evoked responses under normal control conditions (n = 6) and in arthritis (n = 8) These data suggest that activation of both NMDA and non-NMDA receptors contributes to pain-related sensitization of amygdala neurons. PMID- 15275759 TI - Self-mutilation in young children following brachial plexus birth injury. AB - Brachial plexus injury in adults commonly produces persistent pain. Pediatric textbooks and case series suggest that perinatal brachial plexus injury is very rarely associated with pain, though this is difficult to determine in preverbal infants. Some of these young children self-mutilate the affected extremity, which may or may not reflect pain. This study was designed to characterize the clinical presentation and course of self-mutilation following perinatal brachial plexus injury. In this retrospective chart review, 280 patients were identified as having a perinatal brachial plexus injury from 1990-2002. Self-mutilation behavior was defined as excessive mouthing of or biting of any part of the affected limb, and/or loss of any parts of the affected limb secondary to biting and infection. Case reports were generated which described the severity of the primary injury, the types of surgical interventions, the duration and temporal relationship of behavior with surgical interventions and the nature of the self mutilation behavior. Eleven patients demonstrated self-mutilating behavior, yielding a cumulative incidence of 3.9%. The median age of onset of this behavior was 17 (IQR=11-21) months, the median onset of the behavior was 8 (IQR) months after surgery and the median duration of this behavior was 6 (IQR=4-7) months. The incidence of self-mutilation among children who had undergone surgery was 6.8% (9 of 133 children) compared to the 1.4% (2 of 147 children) for non surgical patients (P<0.05). Seven of 24 children (29.1%) who underwent brachial plexus dissection demonstrated self-mutilation, which was significantly different from the incidence of self-mutilation in children who did not have surgery (P < 0.001). Self-mutilation behavior in our population occurred more frequently in children following brachial plexus microsurgery. The reasons for this association are unclear, but may be related to either the surgery or the severity of the initial injury or both. PMID- 15275760 TI - Variation in reporting of pain and other subjective health complaints in a working population and limitations of single sample measurements. AB - Measuring health complaints by administrating a single report is common. Our aim was to assess variation in pain and other subjective complaints over an extended period, whether a single-sample produces representative data, and determine associations between complaints. Health-complaint reports were collected from postal workers at monthly intervals over a period of 32-34 consecutive months (1997-2000). We computed six compound complaint-severity indices of 30 complaint severity scores (intensity score x duration score, scale 0-9). In 67% of the scores, the complaints exhibited larger deviation from a reference (12 consecutive reports in the last 24 months of the study period) when using one report from the respective reference period compared with the mean of two consecutive reports. Four consecutive samples were needed to obtain agreement for 95% of the data when the criterion of accepted deviation from the reference was set to +/-1.0. Neither inspection of graphs nor statistical tests revealed any seasonal pattern or trend on either a group or individual level. The musculoskeletal and psychological complaint-severity indices correlated strongly (rs > 0.66). Correlations between the different somatic indices were generally weak or moderate (rs < 0.55). The initial report produced higher complaint ratings than subsequent reports did. Due to large intra-individual complaint variability and higher complaint-severity level exhibited on the initial report compared to those that followed, measuring subjective health with a single-sample approach does not produce data representativeness for average complaints over a period. More than two samples should be collected when the purpose is to reveal changes in health. PMID- 15275761 TI - An integrated undergraduate pain curriculum, based on IASP curricula, for six health science faculties. AB - Pain education, especially for undergraduates, has been identified as important to changing problematic pain practices, yet, no published data were found describing an integrated, interprofessional pain curriculum for undergraduate students. Therefore, this project aimed to develop, implement, and evaluate an integrated pain curriculum, based on the International Association for the Study of Pain curricula [http://www.iasp-pain.org/curropen.html], for 540 students from six Health Science Faculties/Departments. Over an 18-month period, the University of Toronto Centre for the Study of Pain's Interfaculty Pain Education Committee developed a 20-h undergraduate pain curriculum to be delivered during a 1-week period. Students from Dentistry, Medicine, Nursing, Pharmacy, Physical Therapy, and Occupational Therapy participated as part of their 2nd or 3rd year program. Teaching strategies included large and small groups, Standardized Patients, and 63 facilitators. Evaluation methods included: (a) pre- and post-tests of the Pain Knowledge and Beliefs Questionnaire (PKBQ) and (b) Daily Content and Process Questionnaire (DCPQ) to obtain feedback about process, content, and format across the curriculum's 5 days. A significant improvement in pain knowledge and beliefs was demonstrated (t = 181.28, P < 0.001), although non-responders were problematic at the post-test. DCPQ overall ratings of 'exceeding or meeting expectations' ranged from 74 to 92%. Ratings were highest for the patient-related content and panel, and the small-group discussions with Standardized Patients. Overall evaluations were positive, and statistically significant changes were demonstrated in students' pain knowledge and beliefs. This unique and valuable learning opportunity will be repeated with some modifications next year. PMID- 15275762 TI - Acidic pH and capsaicin activate mechanosensitive group IV muscle receptors in the rat. AB - Strenuous exercise of muscle as well as inflammation and ischaemia are associated with tissue acidosis. However, so far, nothing is known about the sensitivity to hydrogen ions of slowly conducting muscle afferent units. The study investigated if acid-sensing ion channels, e.g. the polymodal acid/capsaicin-sensitive vanilloid receptors, were present on unmyelinated (group IV) muscle afferent units of the gastrocnemius-soleus muscle of the rat. Intramuscular injections of acidic phosphate buffer pH 6 excited 56.0% of the group IV units. A similar proportion (54.29%) was activated by capsaicin (655 microM). Tests of the same unit with both adenosine triphosphate at neutral pH (ATP, 7.6 mM; pH 7.4) and acidic phosphate demonstrated that most acid-sensitive units were also excited by ATP at neutral pH. The data show that (1) a high proportion of group IV muscle receptors are responsive to an increased extracellular hydrogen ion concentration, and (2) a subpopulation of these units are sensitive to both acidic pH and ATP. PMID- 15275763 TI - Nociceptive facilitating neurons in the rostral ventromedial medulla. AB - The role of the periaqueductal gray-rostral ventromedial medulla (RVM) system in descending inhibition of nociception has been studied for over 30 years. The neural basis for this antinociceptive action is reasonably well understood, with strong evidence that activation of a class of RVM neurons termed 'off-cells' exerts a net inhibitory effect on nociception. However, it has recently become clear that this system can facilitate, as well as inhibit pain. Although the mechanisms underlying the facilitation of nociception have not been conclusively identified, indirect evidence points to activation of a class of neurons termed 'on-cells' as mediating descending facilitation. Here we used focal infusion of the tridecapeptide neurotensin within the RVM in lightly anesthetized rats to activate on-cells selectively. Neurotensin has been shown in awake animals to produce a dose-related, bi-directional effect on nociception when applied within the RVM, with hyperalgesia at low doses, and analgesia at higher doses. Using a combination of single cell recording and behavioral testing, we now show that on cells are activated selectively by low-dose neurotensin, and that the activation of on-cells by neurotensin results in enhanced nociceptive responding, as measured by the paw withdrawal reflex. Furthermore, higher neurotensin doses recruit off-cells in addition to on-cells, producing behavioral antinociception. Selective activation of on-cells is thus sufficient to produce hyperalgesia, confirming the role of these neurons in facilitating nociception. Activation of on-cells likely contributes to enhanced sensitivity to noxious stimulation or reduced sensitivity to analgesic drugs in a variety of conditions. PMID- 15275764 TI - State-dependent block of voltage-gated Na+ channels by amitriptyline via the local anesthetic receptor and its implication for neuropathic pain. AB - Amitriptyline is a tricyclic antidepressant, which also alleviates various pain syndromes at its therapeutic plasma concentration (0.36-0.90 microM). Accumulated evidence suggests that such efficacy may be due to block of voltage-gated Na(+) channels. The Na(+) channel alpha-subunit protein consists of four homologous domains (D1-D4), each with six transmembrane segments (S1-S6). The aims of this study were to locate the amitriptyline receptor in the Na(+) channel alpha subunit and to compare the amitriptyline affinity in open, inactivated, and resting states of the Na(+) channel. Wild-type and mutant rat skeletal muscle alpha-subunit Na(+) channels were expressed in human embryonic kidney cells and assayed under whole-cell voltage clamp conditions. Our results indicate that the amitriptyline receptor overlaps with the local anesthetic receptor to a great extent in Na(+) channels. Residues N434 (at D1-S6), L1280 (D3-S6), and F1579 (D4 S6) may jointly form parts of the amitriptyline/local anesthetic receptor, with residue L1280 being most critical for amitriptyline binding. Open-channel block by amitriptyline was assessed in inactivation-deficient Na(+) channels and compared with the resting- and inactivated-channel block in wild-type channels. The open-channel block by amitriptyline has the highest affinity, with a 50% inhibitory concentration (IC(50)) of 0.26 microM. The inactivated-channel block by amitriptyline had a weaker affinity (0.51 microM), whereas the resting-channel displayed the weakest affinity (33 microM). We hypothesize that selective block of both persistent late openings and the inactivated state of neuronal Na(+) channel isoforms by amitriptyline also occurs at its therapeutic concentration and likely contributes to its efficacy in pain syndromes. PMID- 15275765 TI - Gabapentin for the prevention of postoperative pain after vaginal hysterectomy. AB - Gabapentin alleviates and/or prevents acute nociceptive and inflammatory pain both in animals and volunteers, especially when given before trauma. Gabapentin might also reduce postoperative pain. To test the hypothesis that gabapentin reduces the postoperative need for additional pain treatment (postoperative opioid sparing effect of gabapentin in humans), we gave 1200 mg of gabapentin or 15 mg of oxazepam (active placebo) 2.5 h prior to induction of anaesthesia to patients undergoing elective vaginal hysterectomy in an active placebo controlled, double blind, randomised study. Gabapentin reduced the need for additional postoperative pain treatment (PCA boluses of 50 microg of fentanyl) by 40% during the first 20 postoperative hours. During the first 2 postoperative hours pain scores at rest and worst pain score (VAS 0-100 mm) were significantly higher in the active placebo group compared to the gabapentin-treated patients. Additionally, pretreatment with gabapentin reduced the degree of postoperative nausea and incidence of vomiting/retching possibly either due to the diminished need for postoperative pain treatment with opioids or because of an anti-emetic effect of gabapentin itself. No preoperative differences between the two groups were encountered with respect to the side effects of the premedication. However, 15 mg oxazepam was more effective in relieving preoperative anxiety than 1200 mg gabapentin. PMID- 15275766 TI - Relative analgesic potency of fentanyl and sufentanil during intermediate-term infusions in patients after long-term opioid treatment for chronic pain. AB - Sufentanil, a potent mu-opioid agonist, historically has not been been given systemically to treat chronic pain. An implantable, fixed-rate osmotic pump that delivers sufentanil subcutaneously is being developed for this purpose. In that transdermal fentanyl may be a useful intermediary to estimate the appropriate sufentanil dose before implant, accurate information is needed about the relative analgesic potency of sufentanil and fentanyl during continuous infusion. To determine this relative potency, we administered these drugs to opioid-treated chronic pain patients using a target-controlled infusion (TCI). Sixty-three patients with stable chronic pain and daily oral opioid requirements equivalent to 100-1000 mg of morphine received TCI of fentanyl and sufentanil, each for a minimum of 16 h. Drug administration was double-blind and the order of administration was randomly assigned. Target concentration was changed until the patient reported that analgesia was adequate (defined as a pain level equal to or better than baseline). Seven patients did not complete the infusion and protocol violations invalidated data for 15 patients. For the remaining 41 patients, target concentrations associated with adequate analgesia were achieved for both sufentanil and fentanyl. The median value for the equianalgesic concentration ratio (steady-state fentanyl infusion to steady-state sufentanil infusion) was 7.5; mean potency ratio was 7.44 (95% confidence interval 6.8-8.2). During titrated, intermediate-term infusions in patients previously treated with opioids for chronic pain, sufentanil is approximately 7.5 times as potent as fentanyl. PMID- 15275767 TI - Differentiating sensory and affective-sensory pain descriptions in patients undergoing magnetic resonance imaging for persistent low back pain. AB - The study design is a cross-sectional survey with psychometric analysis. The objective is to determine the validity of a modified version of the Short-Form McGill Pain Questionnaire (SF-MPQ). The SF-MPQ has been widely used to differentiate between reports of sensory and affective pain. The validity of this instrument to reflect independence between these constructs remains unclear. The SF-MPQ, the Roland-Morris Questionnaire (RM) and a measure of current pain intensity were completed by 373 patients undergoing lumbar magnetic resonance imaging (MRI). Four hypothesized factor structures for the SF-MPQ (three 2-factor and one 1-factor solution) were tested using confirmatory factor analysis. A modified 2-factor solution (MSF-MPQ) containing 3 items labeled sensory and 5 items labeled affective-sensory had the best degree of fit. Correlations between factors were substantially lower for the modified 2-factor solution (0.48) than for previously described 2-factor solutions (0.88 and 0.92) indicating a higher degree of independence between these factors. Correlations with measures of pain intensity and the RM were significant, but slightly lower, for the subscales of the modified 2-factor solution (0.26-0.40) than for the subscales of the previously described 2-factor solutions (0.34-0.45). The MSF-MPQ can be used as a brief tool to differentiate the language used to describe pain in patients who are undergoing lumbar MRI. The evidence indicates that this clinical tool can be used to categorize how these patients describe their pain and potentially may be very valuable in determining the optimal course of treatment. PMID- 15275768 TI - Sensitization to bradykinin B1 and B2 receptor activation in UV-B irradiated human skin. AB - Bradykinin B1 and B2 receptors contribute to nociceptor sensitization under inflammatory conditions. Here, we examined the vascular inflammatory responses and nociceptive effects resulting from activation of B1 and B2 receptors in healthy and UV-B irradiated skin in human volunteers. The B1 receptor agonist des Arg(10)-Kallidin (10(-6)-10(-3)M) and the B2 receptor agonist bradykinin (10(-9) 10(-4)M) were administered by dermal microdialysis to the ventral thigh. UV-B irradiation was performed 24 h prior to the experiment with the threefold minimum erythemal dose. Pain sensation perceived during the stimulation with the bradykinin receptor agonists was estimated on a numeric scale. Local and axon reflex-induced vasodilations were recorded by laser Doppler imaging. For protein extravasation, total protein content in the dialysate was assessed as a measure of increased endothelial permeability. In normal skin, both B1 and B2 receptor activation dose-dependently evoked pain, vasodilatation and protein extravasation. In UV-B irradiated skin, pain sensation and axon reflex vasodilatation were enhanced by both B1 and B2 agonists, whereas local vasodilatation was increased only following B1 receptor activation. The UV-B irradiation did not enhance B1 and B2 receptor-induced protein extravasation indicating a differential sensitization of the neuronal, but not the vascular response. PMID- 15275769 TI - Opiate anti-nociception is attenuated following lesion of large dopamine neurons of the periaqueductal grey: critical role for D1 (not D2) dopamine receptors. AB - The periaqueductal grey (PAG) area is involved in pain modulation as well as in opiate-induced anti-nociceptive effects. The PAG possess dopamine neurons, and it is likely that this dopaminergic network participates in anti-nociception. The objective was to further study the morphology of the PAG dopaminergic network, along with its role in nociception and opiate-induced analgesia in rats, following either dopamine depletion with the toxin 6-hydroxydopamine or local injection of dopaminergic antagonists. Nociceptive responses were studied through the tail-immersion (spinal reflex) and the hot-plate tests (integrated supraspinal response), establishing a cut-off time to further minimize animal suffering. Heroin and morphine were employed as opiates. Histological data indicated that the dopaminergic network of the PAG is composed of two types of neurons: small rounded cells, and large multipolar neurons. Following dopamine depletion of the PAG, large neurons (not small ones) were selectively affected by the toxin (61.9% dopamine cell loss, 80.7% reduction of in vitro dopaminergic peak), and opiate-induced analgesia in the hot-plate test (not the tail-immersion test) was reliably attenuated in lesioned rats (P < 0.01). After infusions of dopaminergic ligands into the PAG, D(1) (not D(2)) receptor antagonism attenuated opiate-induced analgesia in a dose-dependent manner in the hot-plate test. The present study provides evidence that large neurons of the dopaminergic network of the PAG participate in supraspinal (not spinal) nociceptive responses after opiates through the involvement of D(1) dopamine receptors. This dopaminergic system should be included as another network within the PAG involved in opiate induced anti-nociception. PMID- 15275770 TI - Why cooling is beneficial: non-linear temperature-dependency of stimulated iCGRP release from isolated rat skin. AB - The capsaicin receptor in nociceptive neurons is a target for the sensitizing actions of algogenic inflammatory mediators. Capsaicin and potential endogenous ligands are thought not to gate this heat-activated ion channel but to sensitize it so profoundly that even room temperature can open it. We investigated the temperature dependency of capsaicin-induced CGRP release from nociceptive nerve fibers in isolated rat skin over a range of ambient temperatures using different agonist concentrations (10(-7)-10(-5)M) and KCl (60 mM) for control. Ambient temperature (4-40 degrees C) showed no significant influence on the basal iCGRP outflow. The supramaximal capsaicin concentration of 10(-6)M as a stimulus evoked a response that was not significantly diminished by temperatures decreasing from 40 to 24 degrees C but lost 65% of its amplitude between 24 and 14 degrees C (Q(10) approximately 6.7). Such a collapse of the response occurred between 40 and 32 degrees C at lower capsaicin concentration (10(-7)M). The concentration response curves showed a rightward shift upon cooling from 40 to 24 degrees C and a major loss of slope and maximum effect at 14 degrees C which formally describes a noncompetitive antagonism. KCl-induced iCGRP release showed a much more linear temperature dependency (Q(10) approximately 2.4 between 24 and 14 degrees C). Significant capsaicin responses even at 8 degrees C suggest a contribution of noxious-cold sensitive neurons known to coexpress CGRP and the capsaicin receptor. The heat-activated ion channels (TRPV1-4) are thought to play a significant role in inflammatory pain which is effectively relieved by cooling. The present results contribute to understanding this phenomenon. PMID- 15275771 TI - Distraction from chronic pain during a pain-inducing activity is associated with greater post-activity pain. AB - The aim of this study was to investigate the effects of distraction from pain during and after a pain-inducing lifting task in a sample of chronic low back pain (CLBP) patients. Fifty-two CLBP patients (25 males, 27 females; mean age=46.30 years) performed a pain-inducing lifting task twice, once alone and once with a simultaneous cognitive distraction task. The results revealed that (1) distraction had no effect upon self-reported pain during the lifting task, (2) distraction had a paradoxical effect of more pain immediately after the lifting task, (3) both pain-related fear and pain catastrophizing did not moderate the effects of distraction on pain, but (4) catastrophic thinking about pain during the lifting task was related to more vigilance to pain and less engagement in the distracting task. Further investigation of the catastrophizing data showed that the effect of catastrophizing about pain during the lifting task on the cognitive distraction task was mediated by the amount of attention paid to pain. Clinical implications of these findings are discussed. PMID- 15275772 TI - Pain-related fear and daily functioning in patients with osteoarthritis. AB - There is growing evidence supporting the relationship between pain-related fear and functional disability in chronic musculoskeletal pain conditions. In osteoarthritis (OA) patients the role of pain-related fear and avoidance has received little research attention so far. The present study investigates the degree to which pain-related fear, measured with the Tampa Scale for Kinesiophobia (TSK), influences daily functioning in OA patients. The purpose of the present paper was twofold: (1) to investigate the factor structure of the TSK in a sample of OA patients by means of confirmatory factor analysis; and (2) to investigate the role of pain-related fear in OA compared to other factors, such as radiological findings and level of pain intensity. The results show that TSK consists of two factors, called 'activity avoidance' and 'somatic focus', which is in line with other studies in low back pain and fibromyalgia. Furthermore, pain-related fear occurred to a considerable extent in this sample of osteoarthritis patients and was negatively associated with daily functioning. Level of pain and level of pain-related fear were significantly associated with functional limitations. Radiological findings were not significant predictors and when compared to pain-related fear they were not significant. These findings underscore the importance of pain-related fear in daily functioning of OA patients. Therefore, treatment strategies aiming at reduction of pain-related fear in OA patients need to be developed and investigated. PMID- 15275773 TI - Evidence for an exclusive antinociceptive effect of nociceptin/orphanin FQ, an endogenous ligand for the ORL1 receptor, in two animal models of neuropathic pain. AB - Nociceptin/orphanin FQ (noci/OFQ), the endogenous ligand for the orphan ORL1 (opioid receptor-like1), has been shown to be anti- or pronociceptive and modify morphine analgesia in rats after central administration. We comparatively examined the effect of noci/OFQ on hyperalgesia and morphine analgesia in two experimental models of neuropathic pain: diabetic (D) and mononeuropathic (MN) rats. Noci/OFQ, when intrathecally (i.t.) injected (0.1, 0.3, or 1, to 10 microg/rat) was ineffective in normal rats, but reduced and suppressed mechanical hyperalgesia (paw-pressure test) in D and MN rats, respectively. This spinal inhibitory effect was suppressed by naloxone (10 microg/rat, i.t.) in both models. Combinations of systemic morphine with spinal noci/OFQ resulted in a strong potentiation of analgesia in D rats. In MN rats, an isobolographic analysis showed that the morphine+noci/OFQ association (i.t.) suppressed mechanical hyperalgesia in a superadditive manner. In summary, the present findings reveal that spinal noci/OFQ produces a differential antinociception in diabetic and traumatic neuropathic pain according to the etiology of neuropathy, an effect possibly mediated by opioid receptors. Moreover, noci/OFQ combined with morphine produces antinociceptive synergy in experimental neuropathy, opening new opportunities in the treatment of neuropathic pain. PMID- 15275774 TI - Antinociceptive mechanism of L-DOPA. AB - The mechanism of L-DOPA for antinociception was investigated. Nociceptive behaviors in mice after an intrathecal (i.t.) administration of substance P were evaluated. L-DOPA (i.t.) dose-dependently attenuated the substance P-induced nociceptive behaviors. Co-administration of benserazide (i.t.), a DOPA decarboxylase inhibitor, abolished the antinociceptive effect of L-DOPA. The L DOPA-induced antinociception was antagonized by sulpiride, a D2 blocker, but not by SCH 23390, a D1 blocker. These results suggest that L-DOPA relieves pain after conversion to dopamine, with the dopamine sedating pain transmission by way of the dopamine D2 receptor. PMID- 15275775 TI - Diurnal changes of tonic nociceptive responses in mice: evidence for a proalgesic role of melatonin. AB - Diurnal variations in tonic pain reactions have been described in mice tested in Spring, but the underlying mechanisms are still unknown. We tested the potential role of melatonin, a key hormone in the control of neuro-endocrine circadian rhythms. The experiments were performed in male CBA/J mice housed under controlled temperature, humidity, and light (12/12 dark/light cycle) conditions, during the Light (7-10a.m.) or Dark (7-10p.m.) phases of the diurnal cycle. In a first group of experiments, animals were either pretreated with i.p. saline (controls) or with the melatonin receptor antagonist, luzindole (30 mg/kg), before the s.c. injection of a dilute formalin solution into a hindpaw. In control animals, pain-related behavioral reactions (licking and flinching) were higher in the evening (Dark) than in the morning (Light), both during the first (0-10 min) and the second (11-55 min) phase of the response to s.c. formalin. In animals pre-treated with luzindole, no diurnal changes occurred, pain reactions in the Dark being similar to those of the Light Control group. In a second group of experiments, artificial pinealectomy, obtained by exposing animals to continuous light for 48 h, also reduced pain reactions in the evening to levels comparable to those in the morning. Receptor autoradiography showed lower binding availability at spinal cord level in mice sacrificed during the Dark, as expected from the circadian pattern of melatonin secretion. A further significant decrease of melatonin receptor binding was induced by noxious stimulation. These results suggest a proalgesic role of endogenous melatonin in tonic pain. PMID- 15275776 TI - The 5-HT3 receptor facilitates at-level mechanical allodynia following spinal cord injury. AB - Spinal cord injury (SCI) results in the development of mechanical allodynia immediately rostral to the lesion site, within the dermatome border of normal sensation and sensory loss (at-level mechanical allodynia). We propose that an observed threefold increase in serotonergic fibre immunoreactivity within spinal segments corresponding to these allodynic dermatomes facilitates the maintenance of chronic neuropathic pain via activation of the 5-HT(3) receptor (5-HT(3)-R). Serotonin (5-HT), the non-selective 5-HT(1)/5-HT(2) receptor antagonist, methysergide, the 5-HT(3)-R agonist, m-chlorophenylbiguanide (m-CPBG) or the 5 HT(3)-R antagonist, ondansetron were intrathecally administered five weeks following SCI in rats. Ondansetron produced a robust, long-term reduction of at level mechanical allodynia, while m-CPBG exacerbated allodynia. Exogenous 5-HT transiently reduced at-level mechanical allodynia. This effect was opposed by methysergide, which enhanced mechanical allodynia. Co-administration of 5-HT and ondansetron produced a short-lasting partial summation of effects, further decreasing mechanical allodynia while co-administration of methysergide attenuated the anti-allodynic effect of ondansetron. Depletion of spinal 5-HT via 5,7-dihydroxytryptamine (5,7-DHT) resulted in decreased at-level mechanical allodynia. The reduction of allodynia by ondansetron was lost following 5,7-DHT administration, suggesting that reduced allodynia following intrathecal ondansetron is via blockade of 5-HT-induced excitation of the 5-HT(3)-R. These results suggest that increased 5-HT fibre density immediately rostral to the SCI lesion site could have transient effects to reduce mechanical allodynia via actions at 5-HT(1) and/or 5-HT(2) receptors. However, the more long-lasting effects of this enhanced serotonergic input may facilitate chronic, at-level allodynia via the 5-HT(3)-R. PMID- 15275777 TI - Mechanical allodynia and thermal hyperalgesia upon acute opioid withdrawal in the neonatal rat. AB - Upon withdrawal from opioids many patients experience a heightened sensitivity to stimuli and an exaggerated pain response. We present evidence that neonatal rats exhibit allodynia and hyperalgesia on acute opiate withdrawal. Postnatal 7 and 21 day rats were used to approximately model a full term human infant and a human child, respectively. The opiate antagonist naloxone was used to precipitate withdrawal at 30 or 120 min after a single acute administration of morphine. Alternatively, rats were allowed to undergo spontaneous withdrawal. Behavioral manifestations of withdrawal syndrome were not observed when naloxone was administered at 30 min post-morphine, but were present when withdrawal was precipitated at 120 min. Spontaneous and precipitated withdrawal from a single acute administration of morphine produced mechanical allodynia and thermal hyperalgesia in postnatal day 7 rats and mechanical allodynia in postnatal day 21 rats. A higher dose of morphine was required to produce mechanical allodynia in postnatal day 21 versus 7 rats but this increase was independent of the analgesic efficacy of morphine at these two ages. The present work illustrates the need to examine the phenomenon of hypersensitivity upon opioid withdrawal in the human pediatric population. PMID- 15275778 TI - Exaggerated nociceptive responses on morphine withdrawal: roles of protein kinase C epsilon and gamma. AB - On withdrawal from opioids many patients experience a heightened sensitivity to stimuli and an exaggerated pain response. The phenomenon has been little studied in infants. We present evidence that in postnatal day 7 rats an exaggerated nociceptive ventral root response of spinal cords in vitro and withdrawal associated thermal hyperalgesia in vivo are dependent on protein kinase C (PKC), and we document the roles of PKC and gamma isozymes. In vitro, the slow ventral root potential (sVRP) is a nociceptive-related response in spinal cord that is depressed by morphine and recovers to levels significantly above control on administration of naloxone. A broad-spectrum PKC antagonist, GF109213X, blocked withdrawal hyperresponsiveness of the sVRP whereas an antagonist specific to Ca(++)-dependent isozymes, Go69076, did not. Consistent with this finding, a specific peptide inhibitor of calcium-independent PKC, but not an inhibitor of calcium-dependent PKC gamma, blocked withdrawal hyperresponsiveness of the sVRP. Similarly, in vivo in 7-day-old rat pups, inhibition of PKC, but not PKC gamma, prevented thermal hyperalgesia precipitated by naloxone at 30 min post-morphine. In contrast, thermal hyperalgesia during spontaneous withdrawal was inhibited by both PKC and gamma inhibitors. The consistency between the in vivo and in vitro findings with respect to naloxone-precipitated withdrawal provides further evidence that the sVRP reflects nociceptive neurotransmission. In addition the difference between naloxone-precipitated and spontaneous withdrawal in vivo suggests that in postnatal day 7 rats, morphine exposure produces an early phase of primary afferent sensitization dependent upon PKC translocation, followed by a later phase involving spinal sensitization mediated by PKC gamma. PMID- 15275779 TI - The effects of axotomy on neurons and satellite glial cells in mouse trigeminal ganglion. AB - Damage to peripheral nerves induces ectopic firing in sensory neurons, which can contribute to neuropathic pain. As most of the information on this topic is on dorsal root ganglia we decided to examine the influence of infra-orbital nerve section on cells of murine trigeminal ganglia. We characterized the electrophysiological properties of neurons with intracellular electrodes. Changes in the coupling of satellite glial cells (SGCs) were monitored by intracelluar injection of the fluorescent dye Lucifer yellow. Electrophysiology of SGCs was studied with the patch-clamp technique. Six to eight days after axotomy, the percentage of neurons that fire spontaneously increased from 1.6 to 12.8%, the membrane depolarized from -51.1 to -45.5 mV, the percentage of cells with spontaneous potential oscillations increased from 19 to 37%, the membrane input resistance decreased from 44.4 to 39.5 MOmega, and the threshold for firing an action potential decreased from 0.61 to 0.42 nA. These changes are consistent with increased neuronal excitability. SGCs were mutually coupled around a given neuron in 21% of the cases, and to SGCs around neighboring neurons in only 4.8% of the cases. After axotomy these values increased to 37.1 and 25.8%, respectively. After axotomy the membrane resistance of SGCs decreased from 101 MOmega in controls to 40 MOmega, possibly due to increased coupling among these cells. We conclude that axotomy affects both neurons and SGCs in the trigeminal ganglion. The increased neuronal excitability and ectopic firing may play a major role in neuropathic pain. PMID- 15275781 TI - Electronic pain questionnaires: a randomized, crossover comparison with paper questionnaires for chronic pain assessment. AB - Electronic questionnaires for pain assessment are becoming increasingly popular. There have been no published reports to establish the equivalence or psychometric properties of common pain questionnaires administered via desktop computers. This study compared responses to paper (P) and touch screen electronic (E) versions of the Short-Form McGill Pain Questionnaire (SF-MPQ) and Pain Disability Index (PDI), while examining the role of computer anxiety and experience, and evaluating patient acceptance. In a randomized, crossover design 189 chronic pain patients completed P and E versions of the SF-MPQ and PDI, and self-ratings of anxiety, experience, relative ease and preference. Psychometric properties were highly similar for P and E questionnaires. For the SF-MPQ, 60% or more of subjects gave equivalent responses on individual descriptors and PPI scale, with 80% rating within +/-1 point for an 11-point VAS. Correlations for the SF-MPQ scales ranged from 0.68 to 0.84. For the PDI, 60% or more of subjects responded within +/-1 point on individual questions, and the total score correlation was 0.67. Comparison of mean difference scores revealed no significant differences between modes for any of the questionnaire items or scores. Anxiety and experience scores showed no significant associations through correlations and high/low comparisons. Although nearly half of subjects reported no computer training, anxiety ratings were low, and considerably more subjects rated the E questionnaires as easier and preferred. Findings are consistent with test-retest reliability data, and support the validity and acceptance of electronic versions of the SF-MPQ and PDI. PMID- 15275780 TI - Peri-sciatic proinflammatory cytokines, reactive oxygen species, and complement induce mirror-image neuropathic pain in rats. AB - In inflammatory neuropathy, immune activation near intact peripheral nerves induces mechanical allodynia. The identity of the peripheral immune product(s) that lead to these changes in pain behavior is unknown. The present series of studies utilized the sciatic inflammatory neuropathy (SIN) model to examine this question. Here, inflammatory neuropathy is created by injecting an immune activator (zymosan) around one sciatic nerve via an indwelling catheter. Our prior studies demonstrated that peri-sciatic zymosan activated macrophages and neutrophils to release proinflammatory cytokines and reactive oxygen species (ROS). In addition, zymosan is a classical activator of the complement cascade. Thus the present series of experiments examined whether any of these inflammatory mediators are involved in the initial induction of SIN-induced ipsilateral or bilateral allodynias. Peri-sciatic injection of selective inhibitors/antagonists revealed that a number of immune products are early mediators of the resultant allodynias, including proinflammatory cytokines (tumor necrosis factor, interleukin-1, and interleukin-6), ROS, and complement. Thus these immune-derived substances can markedly alter sensory nerve function at mid-axon. PMID- 15275782 TI - Amplitudes of laser evoked potential recorded from primary somatosensory, parasylvian and medial frontal cortex are graded with stimulus intensity. AB - Intensity encoding of painful stimuli in many brain regions has been suggested by imaging studies which cannot measure electrical activity of the brain directly. We have now examined the effect of laser stimulus intensity (three energy levels) on laser evoked potentials (LEPs) recorded directly from the human primary somatosensory (SI), parasylvian, and medial frontal cortical surfaces through subdural electrodes implanted for surgical treatment of medically intractable epilepsy. LEP N2* (early exogenous/stimulus-related potential) and LEP P2** (later endogenous potential) amplitudes were significantly related to the laser energy levels in all regions, although differences between regions were not significant. Both LEP peaks were also significantly correlated with the pain intensity evoked by the laser stimulus, excepting N2* over the parasylvian region. Peak latencies of both LEP peaks were independent of laser energy levels. N2* and P2** amplitudes of the maxima in all regions showed significant positive linear correlations with laser energy, excepting N2* over the parasylvian region. The lack of correlation of parasylvian cortical N2* with laser energy and pain intensity may be due to the unique anatomy of this region, or the small sample, rather than the lack of activation by the laser. Differences in thresholds of the energy correlation with amplitudes were not significant between regions. These results suggest that both exogenous in endogenous potentials evoked by painful stimuli, and recorded over SI, parasylvian, and medial frontal cortex of awake humans, encode the intensity of painful stimuli and correlate with the pain evoked by painful stimuli. PMID- 15275783 TI - The human histocompatibility leukocyte antigen (HLA) haplotype is associated with the onset of postherpetic neuralgia after herpes zoster. AB - In some herpes zoster patients, pain persists for more than 3 months or more after healing of vesicular eruptions; this condition is termed postherpetic neuralgia (PHN). We have recently reported the association of the human histocompatibility leukocyte antigens (HLA) haplotype, HLA-A*3303-B*4403 DRB1*1302 with PHN patients; however, it has not been determined whether the haplotype is also associated with herpes zoster that did not develop subsequent PHN. To distinguish whether the haplotype is associated with herpes zoster or the development of PHN, we examined if herpes zoster patients without subsequently PHN are also associated with the HLA haplotype or not. Herpes zoster patients were followed up for more than 6 months, and HLA alleles and haplotypes were compared among the PHN patients (n = 52) the herpes zoster patients who did not develop PHN (n = 42) and healthy controls (n = 125). The frequencies of the risk haplotype in the PHN patients, in the healthy controls and in the herpes zoster patients without subsequent PHN were 16.3, 5.2 and 4.8%, respectively. While the frequency of the risk haplotype was significantly higher in the PHN patients than in the healthy controls (P = 0.0006) no difference was observed between the herpes zoster patients without subsequent PHN and the healthy controls. No significant association was found between the duration of symptoms or the site of herpes zoster and the HLA alleles and the haplotype. These results suggest that the HLA-A*3303-B*4403-DRB1*1302 haplotype plays an important role in the development of PHN after herpes zoster, but not in the onset of herpes zoster. PMID- 15275784 TI - Determinants of prescribing meperidine compared to morphine in hospitalized patients. AB - Morphine is a preferred narcotic since meperidine forms toxic metabolites. Determinants of meperidine use have been poorly described. The objective of this study is to explore factors associated with the ordering of meperidine versus morphine. Retrospective chart review of adult patients, randomly selected based on orders for morphine or meperidine. 1552 orders were written for 670 patients. Of these, 36% were for meperidine. In multivariable analysis, the ordering of meperidine was associated with the following variables in decreasing order of importance: physician specialty, total doses received, hospital location, patient race, age and insurance, and physician gender. More orders for meperidine were written for those receiving fewer doses. Though meperidine has little role in the routine management of hospital pain, we found it continues to be used frequently. Importantly, meperidine is ordered more frequently for patients who receive shorter courses of narcotics. Our study suggests that interventions targeted at more appropriate use of meperidine rather than complete elimination might be more acceptable to physicians while minimizing the risk of toxicity. PMID- 15275785 TI - Contributions of the anterior cingulate cortex and amygdala to pain- and fear conditioned place avoidance in rats. AB - The pain experience includes a sensory-discriminative and an affective-emotional component. The sensory component of pain has been extensively studied, while data about the negative affective component of pain are quite limited. The anterior cingulate cortex (ACC), and amygdala are thought to be key neural substrates underlying emotional responses. Using formalin-induced conditioned place avoidance (F-CPA) and electric foot-shock conditioned place avoidance (S-CPA) models, the present study observed the effects of bilateral excitotoxic (quinolinic acid 200 nmol/microl) lesions of the ACC and amygdala on pain and fear induced negative emotion, as well as on sensory component of pain. In the place-conditioning paradigm, both intraplantar (i.pl.) injection of formalin and electric foot-shock produced conditioned place avoidance. Excitotoxin-induced lesion of either the ACC or amygdala significantly reduced the magnitude of F CPA. However, the decrease in the magnitude of S-CPA occurred only in the amygdala, but not ACC lesioned animals. Neither ACC nor amygdala lesion significantly changed formalin-induced acute nociceptive behaviors. These results suggest that the amygdala is involved in both pain- and fear-related negative emotion, and the ACC might play a critical role in the expression of pain-related negative emotion. PMID- 15275786 TI - Pain behavior and nerve electrophysiology in the CCI model of neuropathic pain. AB - Experimental painful peripheral neuropathy induced by chronic constriction injury (CCI) of the sciatic nerve results in cutaneous thermal and mechanical allodynia of the hind limb. Our histological studies indicate that the major pathology in the CCI model is a loss of large diameter myelinated fibers distal to the site of injury. Electrophysiological recordings from axons central to the lesion that respond to electrical stimulation distal to it, revealed severe fiber loss, reflected by a decrease (P < 0.05) from 5.2+/-6.8 to 0.5+/-0.1 axons/microfilament 5-9 days post operatively (dpo). At 12-15th dpo some recovery was seen, i.e. 1.5+/-0.28 axons/microfilament in the CCI group. The ratio of A- to C-axons in the control group remained constant throughout the experiment. A distinct area in the paw served by the injured nerve was selected to study the response of axons in each microfilament to mechanical stimulation with von Frey monofilaments. In the control group, 91%+/-0.6 of the microfilaments had at least one axon with a receptive field in this area. This decreased to 17%+/-2.9 in the CCI group 5-9 dpo, but had partially recovered to 44+/-4.2% by 12-15-dpo. Our conclusion is that in the CCI model there is an equal reduction in the number of A and C axons conducting past the lesion site, thus preserving a constant ratio between the two fiber populations. This is true despite the apparent preservation of C-fibers observed in previous histological studies. PMID- 15275787 TI - The prevalence of pain and pain interference in a general population of older adults: cross-sectional findings from the North Staffordshire Osteoarthritis Project (NorStOP). AB - Although pain is experienced at all ages, there is uncertainty about the pattern of its occurrence in older people. We have investigated the prevalence of three aspects of self-reported pain-occurrence of any recent pain, number and location of pain sites, and interference with daily life-to determine their association with age in older people. A cross-sectional postal survey of all adults aged 50 years and over registered with three general practices (n = 11230) in North Staffordshire using self-complete questionnaires was conducted. Respondents' gender, age, employment status, socio-economic classification, and general health status were gathered to characterise the population under study. The location of any recent pain (past 4 weeks) was recorded on a full-body manikin and pain interference was based on a single question. Completed questionnaires were received from 7878 respondents (adjusted response of 71.3%). The 4-week prevalence of any pain was 72.4%; similar across 10-year age-groups, and higher in females than males. In those with pain the median number of painful areas (from 44) was 6, and 12.5% of the responding population were classified as having widespread pain, both figures similar across age-groups. Most regional pains showed a decline in prevalence in the older age-groups, the exceptions being the lower limb regions (hip, knee, foot). Pain that interfered with daily activities was reported by 3002 (38.1%) respondents overall. There was a clear age-related rise in this prevalence with age up to and including the oldest group. Within each regional pain subgroup, the proportion of people who also reported pain interference rose with age. Our study has provided evidence that increasing age in the elderly population is not associated with any change in the overall prevalence of pain, although, as previous studies have suggested, the pattern of pain prevalence in different body regions does change with age. More importantly the extent to which pain interferes with everyday life increases incrementally with age up to the oldest age-group in the community-dwelling general population. PMID- 15275788 TI - Differential expression of central metabotropic glutamate receptor (mGluR) subtypes in a clinical model of post-surgical pain. AB - Tissue damage during surgery can induce 'central sensitization' and the development of pain and hyperalgesia post-operatively. Metabotropic glutamate receptors (mGluRs) contribute to nociception, inflammatory pain and hyperalgesia. This study characterized the temporal expression of group I (mGluR(1), mGluR(5)) and II (mGluR(2), mGluR(3)) mGluRs in spinal cord following abdominal surgery. Lumbar spinal cord was recovered from adult sheep euthanased 5 h, 1, 2, 3 and 6 days after undergoing a midline laparotomy, and processed for mGluR mRNA (real time PCR, in situ hybridization) and protein (Western blotting). mGluR(5) mRNA was up-regulated 5 h and 1 day post-surgery in laminae I-II of the spinal cord dorsal horn. mGluR(5) protein was increased 1 day post-surgery. A delayed induction of mGluR(2) and mGluR(3) mRNAs and mGluR(2/3) protein occurred in spinal cord 3 days after surgery. By 6 days, mGluR(2) mRNA levels had returned to normal, however, mGluR(3) mRNA and mGluR(2/3) protein remained elevated. No change was detected in mGluR(1). These results demonstrate that mGluRs are differentially regulated following surgery and support a link between mGluR mediated activity and post-surgical pain. PMID- 15275789 TI - The Cheshire Foot Pain and Disability Survey: a population survey assessing prevalence and associations. AB - Previous foot studies have consistently reported high prevalence estimates in self-reported foot disorders. Few population studies, however, have attempted to assess the impact of foot problems in terms of pain and disability so that the burden associated with foot pain is unknown. A cross-sectional postal survey was conducted on a random community sample of 4780 individuals with 3417 (84%) responding. Subjects reporting foot pain in the past month, current pain and marking one item on the Manchester Foot Pain and Disability Index were classified as having disabling foot pain. Those with disabling foot pain and a random sample with no symptoms (matched for age and gender) received a follow-up interview and standardised foot examination. Self-reported nail problems, corns and callosities, swollen feet, foot deformities and recent foot injuries were found to be associated with disabling foot pain. Foot disability was also associated with pain in the shoulder, axial skeleton, hip/upper leg and knee along with other indicators of poor general health. Clinician diagnosed foot problems associated with disabling foot pain were swollen feet, knee and foot joint tenderness. Only 36% of persons with disabling foot pain received professional foot treatment in the 6 months preceding the survey. The results showed that 323/3417 (9.5%) reported symptoms of disabling foot pain and that this condition is likely to be multi-factorial in origin. Further work is necessary to understand more about the extent and type of unmet need and on how patients presenting with symptoms of disabling foot pain should best be managed. PMID- 15275790 TI - Chronic fentanyl or buprenorphine infusion in the mouse: similar analgesic profile but different effects on immune responses. AB - It is known that morphine has a negative impact on the immune responses. The potent opioids fentanyl and buprenorphine have recently become available as transdermal preparation for the treatment of chronic pain. We analyze the effect of fentanyl and buprenorphine on splenic cellular immune responses in the mouse. The parameters evaluated were lymphoproliferation, natural killer cell activity and interleukin-2 and interferon-gamma production. Drugs were administered acutely at the equianalgesic doses of 0.25 mg/kg for fentanyl and 5 mg/kg for buprenorphine, or delivered continuously with osmotic pumps for 24 h, 3 and 7 days at the rate of 7.5 microg/h per mouse (fentanyl) and 12.5 microg/h per mouse (buprenorphine). After acute administration, a significant decrease of lymphoproliferation is observed in fentanyl-treated animals only. After 24 h of fentanyl administration all the parameters were significantly reduced. After 3 days of fentanyl infusion NK activity had returned to normal values, while all the other parameters were still significantly reduced. In 7 day fentanyl-treated animals immunological tolerance had developed, since no differences with controls were present. In contrast no immune alterations were ever present in buprenorphine-treated animals. No tolerance to the antinociceptive effect of drugs had yet developed. After 1 week of infusion with fentanyl and buprenorphine, new pumps were implanted releasing double amounts of drugs. Neither fentanyl nor buprenorphine-treated animals showed altered immune responses at any time considered. These results indicate that fentanyl and buprenorphine exert different immune effects. Opioid-induced immunosuppression is less relevant in chronic administration than in acute or short-time administration. PMID- 15275791 TI - Cold and heat pain assessment of the human oesophagus after experimental sensitisation with acid. AB - The aim of the present study was to investigate the effect of thermal stimulation of the oesophagus before and after sensitisation with acid. In 17 healthy subjects a stimulation bag was used to re-circulate water at 5 and 60 degrees C for up to 90 s in the lower part of the oesophagus. The area under the temperature curve was used to assess the caloric load. The thermal stimuli were repeated after perfusion of the oesophagus with acid. The evoked pain intensity and referred pain areas (at the pain threshold) were assessed. At baseline the subjects were able to tolerate less caloric load (42%) for the heat compared to the cold stimuli (P = 0.007). The heat stimuli resulted in an increased referred pain area as compared with the cold stimuli (P = 0.03). Following acid perfusion there was a selective sensitisation to the heat pain stimuli as only 36% of the initial caloric load was tolerated (P = 0.012) whereas the sensation to the cold stimuli was unchanged. After acid perfusion, the referred pain area to the heat pain stimulation increased 49% (P = 0.04) but was not changed to cold stimulation (P = 0.82). After sensitisation the words used to describe the sensations to heat pain stimuli shifted from a warmth quality towards a more burning quality in most subjects. This multi-modal sensory testing study showed that acid sensitises the oesophagus to heat but not to cold pain. This may account for the modality specific symptoms and hypersensitivity reported in patients suffering from, e.g. gastro-oesophageal reflux disease. PMID- 15275792 TI - The effects of early or late neurolytic sympathetic plexus block on the management of abdominal or pelvic cancer pain. AB - Neurolytic sympathetic plexus block (NSPB) has been proposed to prevent the development of pain and improve the quality of life of patients with cancer, thus questioning the WHO protocol that proposes the use of invasive methods only as a final resort. This study evaluates the pain relief, opioid consumption and quality of life provided by the use of NSPB in two different phases of cancer pain and compares them with that provided by pharmacological therapy only. Sixty patients with abdominal or pelvic cancer pain were divided into three groups and observed for 8 weeks. In group I, neurolytic celiac (NCPB) or superior hypogastric plexus block (SHPB), or lumbar sympathetic ganglion chain block (LSGCB) was performed with alcohol in patients using NSAID and a weak oral opioid or morphine (dose/=4. In group II, NCPB, SHPB or LSGCB were performed on patients using NSAID and morphine (dose>/=90 mg/day) and reporting VAS>/=4. The patients of group III received pharmacological therapy only. The patients of groups I and II had a significant reduction of pain (P < 0.004), opioid consumption (P < 0.02) and a better quality of life (P < 0.006) than those of group III, but no significant differences between groups I and II were seen in these aspects. Opioid-related adverse effects were significantly greater in group III (P < 0.05). The occasional neurolysis-related complications were transitory. The results suggest NSPB for the management of cancer pain should be considered earlier in the disease. PMID- 15275793 TI - The calpain inhibitor MDL 28170 prevents inflammation-induced neurofilament light chain breakdown in the spinal cord and reduces thermal hyperalgesia. AB - Since long-term hyperexcitability of nociceptive neurons in the spinal cord has been suggested to be caused and maintained by changes of protein expression we assessed protein patterns in lumbar spinal cord during a zymosan induced paw inflammation employing two-dimensional (2D) gel electrophoresis. 2D PAGE revealed a time-dependent breakdown of scaffolding proteins one of which was neurofilament light chain (NFL) protein, which has been previously found to be important for axonal architecture and transport. Nociception induced breakdown of NFL in the spinal cord and dorsal root ganglias was prevented by pretreatment of the animals with a single dose of the specific inhibitor of the protease calpain (MDL-28170) which has been shown to be the primary protease involved in neurofilament degradation in neurodegenerative diseases. Treatment with the calpain inhibitor also provided anti-inflammatory and anti-hyperalgesic effects in the zymosan induced paw inflammation model irrespective of whether the drug was administered systemically (i.p.) or delivered onto the lumbar spinal cord. This suggests that the activation of calpain is involved in the sensitization of nociceptive neurons what is partly due to neurofilament breakdown but cleavage of other calpain substrates may also be involved. Our results indicate that inhibition of pathological calpain activity may present an interesting novel drug target in the treatment of pain and inflammation. PMID- 15275794 TI - Prostaglandin E2 in the midbrain periaqueductal gray produces hyperalgesia and activates pain-modulating circuitry in the rostral ventromedial medulla. AB - Recent years have seen significant advances in our understanding of the peripheral and spinal mechanisms through which prostaglandins contribute to nociceptive sensitization. By contrast, the possibility of a supraspinal contribution of these compounds to facilitated pain states has received relatively little attention. One possible mechanism through which prostaglandins could act supraspinally to facilitate nociception would be by recruitment of descending facilitation from brainstem pain-modulating systems. The rostral ventromedial medulla (RVM) is now known to contribute to enhanced responding in a variety of inflammatory and nerve injury models. Its major supraspinal input, the midbrain periaqueductal gray (PAG), expresses prostanoid receptors and synthetic enzymes. The aim of the present study was to determine whether direct application of prostaglandin E(2) (PGE(2)) within the ventrolateral PAG is sufficient to produce hyperalgesia, and whether any hyperalgesia could be mediated by recruiting nociceptive modulating neurons in the RVM. We determined the effects of focal application of PGE(2) in the PAG on paw withdrawal latency and activity of identified nociceptive modulating neurons in the RVM of lightly anesthetized rats. Microinjection of PGE(2) (50 fg in 200 nl) into the PAG produced a significant decrease in paw withdrawal latency. The PGE(2) microinjection activated on-cells, RVM neurons thought to facilitate nociception, and suppressed the firing of off-cells, RVM neurons believed to have an inhibitory effect on nociception. These data demonstrate a prostaglandin-sensitive descending facilitation from the PAG, and suggest that this is mediated by on- and off-cells in the RVM. PMID- 15275795 TI - Neck pain in adolescence. A 4-year follow-up of pain-free preadolescents. AB - The main aim of this study was to explore the occurrence and changes of neck pain in pain-free preadolescents. The evaluation was performed at 1- and 4-year follow ups. Of the pain-free preadolescents, 366 (71.9%) completed structured pain questionnaires at 1 and 4 years. The occurrence of neck pain at least once a month was 21.3 and 43.4% and at least once a week was 6.3 and 19.4%. Sex difference was found only at the 4-year follow-up, when subjects were 13-16-year old. Neck pain was then more common among girls than boys (P < 0.001). The intensity of pain increased with the frequency of pain (P < 0.001). Of those with neck pain, 28% had used painkillers. The proportion increased with the frequency of neck pain (P = 0.054). Neck pain occurred more often with some other musculoskeletal pain than as a single pain. The frequency of neck pain correlated with the frequency of headache (r = 0.39 [95% confidence interval (CI), 0.30 0.47]) and with the disability (r = 0.26 [95% CI, 0.16-0.35]). This study strengthens the results of the previous cross-sectional studies that occurrence of neck pain increases with age, and that neck pain becomes more common among girls than boys in adolescence. Among preadolescents who were originally pain free, there was only a small proportion who reported frequent neck pain at both 1 and 4 years. It also showed that the frequency of neck pain reflects the intensity of pain fairly well. PMID- 15275796 TI - Site 1 sodium channel blockers prolong the duration of sciatic nerve blockade from tricyclic antidepressants. AB - Many recent reports in the literature address the local anesthetics efficacy of tricyclic antidepressants (TCAs). Here we investigated whether nerve block from TCAs is prolonged by site 1 sodium channel blockers such as tetrodotoxin and saxitoxin, which are known to prolong block from conventional local anesthetics. Tetrodotoxin and saxitoxin greatly prolonged block from TCAs. For example, the median duration of thermal nociceptive blocks for 10 mM amitriptyline, nortriptyline and doxepin were 0, 0, and 124 min; co-injection with 20 microM TTX (median block duration=0), yielded blocks lasting 404, 325, and 697 min, respectively. Co-injection of 12 microM saxitoxin (median block duration=0) with 10 mM amitriptyline resulted in a thermal nociceptive block duration of 373 min. Co-injection of 7.7 mM bupivacaine and 7.7 mM amiptriptyline did not result in block prolongation. Systemic (subcutaneous) delivery of tetrodotoxin or amitriptyline did not result in prolongation of block from the other class of drug injected at the sciatic nerve. In TCA-containing formulations, motor blockade was consistently longer than thermal nociceptive block; motor blockade was also prolonged by tetrodotoxin and saxitoxin. In summary site 1 sodium channel blockers prolong the duration of TCAs via a locally mediated mechanism. PMID- 15275797 TI - Imagery reduces children's post-operative pain. AB - This un-blinded experimental study investigated the effectiveness of imagery, in addition to routine analgesics, in reducing tonsillectomy and/or adenoidectomy pain and anxiety after ambulatory surgery (AS) and at home. Seventy-three children, aged 7-12, were recruited from five AS settings. Thirty-six children randomly assigned to the treatment group watched a professionally developed videotape on the use of imagery and then listened to a 30-min audio tape of imagery approximately 1 week prior to surgery (T1). They listened to only the audio tape 1-4 h after surgery (T2), and 22-27 h after discharge from AS (T3). The 37 children in the attention-control group received standard care. Pain and anxiety were measured at each time-point in both groups. Measures of sensory pain were the Oucher and amount of analgesics used in AS and home; affective pain was measured with the Facial Affective Scale (FAS). Anxiety was measured using the State Trait Anxiety Inventory for Children (STAIC). When controlling for trait anxiety and opioid and non-opioid intake 1-4 h before the pain measures, MANCOVA showed significantly lower pain and anxiety in the treatment group at T2, but not at T3. When controlling for trait anxiety, a two-way RM MANCOVA indicated no significant group differences in combined opioid and non-opioid use between the groups, or between times. Appropriately trained health care providers should use imagery to reduce post-operative pain following tonsillectomy and/or adenoidectomy in AS. Teaching parents about adequate home administration of analgesics may increase the effectiveness of imagery at home. PMID- 15275798 TI - Familial aggregation of depression in fibromyalgia: a community-based test of alternate hypotheses. AB - Numerous studies report that fibromyalgia (FM), a syndrome characterized by widespread pain and generalized tender points, is comorbid with major depressive disorder (MDD). The current study tests two alternate explanations for their comorbidity using a family study methodology. The first is that FM is a depression spectrum disorder. The second is that depression is a consequence of living with FM. We recruited potential probands by initially screening by telephone for FM and MDD among women in the NY/NJ metropolitan area, randomly selecting telephone numbers from a list of households with women. Eligible women were invited for second stage physical examinations for FM diagnosis and psychiatric interviews for MDD diagnosis. All available adult, first-degree relatives received psychiatric interviews. Relatives of probands were divided into four groups on the basis of the probands' FM and MDD diagnoses (FM+/MDD+ (n = 156), FM+/MDD- (n = 51), FM-/MDD+ (n = 351) and FM-/MDD- (n = 101)). Results indicated that rates of MDD in the relatives of probands with FM but without personal histories of MDD were virtually identical to rates of MDD in relatives of probands with MDD themselves. This outcome is consistent with the hypothesis that FM is a depression spectrum disorder, in which FM and MDD are characterized by shared, familially mediated risk factors. The implications of these findings for a stress-vulnerability model of FM are discussed. PMID- 15275799 TI - Symptoms and signs in patients with suspected neuropathic pain. AB - The study sought to determine if symptoms and signs cluster differentially in groups of patients with increasing evidence of neuropathic pain (NP). We prospectively looked at symptoms and signs in 214 patients with suspected chronic NP of moderate to severe intensity. According to a set of clinical criteria the patients were a priori classified as having the so-called 'Definite NP' (n = 91), 'Possible NP' (n = 71), or 'Unlikely NP' (n = 52). A recording of symptoms including pain descriptors, intensity of five categories of pain, Short Form McGill Pain Questionnaire, and Major Depression Inventory were done. Sensory tests including repetitive pinprick stimulation, examination for cold-evoked pain by an acetone drop and brush-evoked pain were carried out in the maximal pain area and in a control area. High intensity of superficial ongoing pain, and touch or cold provoked pain was associated with chronic pain classified as definite or possible neuropathic. Intensity of deep ongoing pain, and 'paroxysms' was similar in the three groups. Brush-evoked pain was more frequent in definite NP. The McGill Pain Questionnaire and the used pain descriptors could not distinguish between the three clinical categories. Although certain symptoms (touch or cold provoked pain) and signs (brush-evoked allodynia) are more prominent in patients with definite or possible NP, we found considerable overlap with the clinical presentation of patients with unlikely NP. PMID- 15275800 TI - Differential nociceptive deficits in patients with borderline personality disorder and self-injurious behavior: laser-evoked potentials, spatial discrimination of noxious stimuli, and pain ratings. AB - Approximately 70-80% of women meeting criteria for borderline personality disorder (BPD) report attenuated pain perception or analgesia during non suicidal, intentional self-mutilation. The aim of this study was to use laser evoked potentials (LEPs) and psychophysical methods to differentiate the factors that may underlie this analgesic state. Ten unmedicated female patients with BPD (according to DSM-IV) and 14 healthy female control subjects were investigated using brief radiant heat pulses generated by a thulium laser and five-channel LEP recording. Heat pulses were applied as part of a spatial discrimination task (two levels of difficulty) and during a mental arithmetic task. BPD patients had significantly higher heat pain thresholds (23%) and lower pain ratings (67%) than control subjects. Nevertheless, LEP amplitudes were either normal (N1, P2, P3) or moderately enhanced in BPD patients (N2). LEP latencies and task performance did not differ between patients and control subjects. The P3 amplitudes, the vertex potential (N2-P2), and the N1, which is generated near the secondary somatosensory cortex, were significantly reduced during distraction by mental arithmetic in both groups. In addition, P3 amplitudes reflected task difficulty. This study confirms previous findings of attenuated pain perception in BPD. Normal nociceptive discrimination task performance, normal LEPs, and normal P3 potentials indicate that this attenuation is neither related to a general impairment of the sensory-discriminative component of pain, nor to hyperactive descending inhibition, nor to attention deficits. These findings suggest that hypoalgesia in BPD may primarily be due to altered intracortical processing similar to certain meditative states. PMID- 15275801 TI - The analgesic effects that underlie patient satisfaction with treatment. AB - Patient satisfaction and global ratings of study medications are increasingly used as secondary outcome measures in pain clinical trials. However, little is known about the factors that underlie and contribute to these ratings. 191 patients who participated in a randomized trial of parenteral parecoxib sodium followed by oral valdecoxib for pain following laparoscopic cholecystectomy versus standard care rated their satisfaction with the overall performance of the study medications (postoperative days 1 and 7) and also provided global evaluation of the analgesics on postoperative day 7. Analyses indicated that treatment regimen, age, worst pain experienced, pain interference with functioning, morphine equivalent dose taken, and number of opioid-related symptoms (e.g. nausea, fatigue) were all associated with satisfaction with the overall performance of the study medications at day 1. Controlling for all of the predictors, pain interference and morphine equivalent dose use made independent contributions to the prediction of the day 1 global rating. These results were replicated in the prediction of day 7 ratings, except that at day 7, treatment regimen also made a significant independent contribution to the prediction of satisfaction. These findings indicate that the study participants considered more than one factor when estimating their satisfaction with the study medications, and that the changes produced by the treatment (e.g. decreased pain, opioid related symptoms) mediated, in part, the effects of treatment on treatment satisfaction. PMID- 15275802 TI - Comparison of two psychometric scaling methods for ratings of acute musculoskeletal pain. AB - Psychometric theory allows interindividual comparisons by scaling differences among subjects with respect to some psychological attribute. The most widely used psychometric scaling method is classical test theory. The properties of classical test theory pain scores are limited to the observed range of pain scores in a given sample, to the specific conditions in a given sample that are the source of pain (e.g. surgery vs. cancer), and to a particular pain survey. The reliability and meaning of classical test theory scores differ for subjects who have higher or lower amounts of pain, have different painful conditions, or are given a different pain survey. Thus, classical test theory pain scores cannot be used to compare dissimilar patient groups or painful conditions, especially if different pain surveys are used. A different psychometric scaling method, item response theory, produces scores with properties that are not limited to an observed score range, specific conditions, or a particular pain survey, and may thus be better for making such comparisons. To compare the two psychometric methods, data were obtained from 335 subjects who rated their clinical pain using 15 words. The psychometric scaling methods were compared using standardized residuals (data fit), standard errors of measurement (score precision), and a graphical plot of predicted scores against scale scores (bias). The item response theory scores demonstrated better data fit and less bias than did the classical test theory scores. In addition to superior psychometric properties, item response theory scores have several other important theoretical and practical advantages. PMID- 15275803 TI - Complex regional pain syndrome as a stress response. AB - A man in his 50's with a prior traumatic brain injury and multiple psychiatric disorders developed acute pain and swelling in his left leg distal to the mid shin. These symptoms arose during an exacerbation of his post-traumatic stress disorder (PTSD). Among his traumatic memories, he reported having witnessed the combat injury and death of a friend who had lost his left leg distal to the mid shin. A diagnosis of conversion disorder was technically excluded because the findings met criteria for Complex Regional Pain Syndrome (CRPS) type I. Based on recent research into the neurobiology of CRPS, PTSD and conversion disorder, we propose a supraspinal mechanism which could explain how emotional stress can produce both symptoms and signs. PMID- 15275804 TI - More on the clinical and scientific relevance of 'symptom amplification' and psychological factors in pain. PMID- 15275807 TI - Precision averaging. PMID- 15275809 TI - Reverse thermo-responsive poly(ethylene oxide) and poly(propylene oxide) multiblock copolymers. AB - Aiming at developing new reverse thermo-responsive polymers, poly(ethylene oxide) poly(propylene oxide) multiblock copolymers were synthesized by covalently binding the two components using carbonyl chloride and diacyl chlorides as the coupling molecules. The appropriate selection of the various components allowed the generation of systems displaying much enhanced rheological properties. For example, 15 wt% aqueous solutions of an alternating poly(ether-carbonate) comprising PEO6000 and PPO3000 segments, achieved a viscosity of 140,000 Pas, while the commercially available Pluronic F127 displayed 5,000 Pas only. Furthermore, the structure of the chain extender played a key role in determining the sol-gel transition. While poly(ether-ester)s containing therephtaloyl (para) and isophtaloyl (metha) coupling units failed to gel at any concentration, a 15 wt% aqueous solution of the polymer chain-extended with phtaloyl chloride (ortho) gelled at 43 degrees C. The water solutions were also studied by dynamic light scattering and a clear influence of the PEO/PPO ratio on the aggregate size was observed. By incorporating short aliphatic oligoesters into the backbone, prior to the chain extension stage, reverse thermal gelation-displaying biodegradable poly(ether-ester-carbonate)s, were generated. PMID- 15275810 TI - Crosslinked hyaluronic acid hydrogels: a strategy to functionalize and pattern. AB - The physiological activity of hyaluronic acid (HA) polymers and oligomers makes it a promising material for a variety of applications. The development of HA hydrogel scaffolds with improved mechanical stability against degradation and biochemical functionality may enhance their application to tissue engineering. In this report, a crosslinking strategy targeting the alcohol groups via a poly(ethylene glycol) diepoxide crosslinker was investigated for the generation of degradable HA hydrogels. To provide support for cell adhesion in vitro, collagen was incorporated into the HA solution prior to the crosslinking process. The hydrogels have a continuous exterior and a porous interior, with pore diameters ranging from 6 to 9 microm. HA and HA-collagen hydrogels degrade in the presence of hyaluronidase and collagenase enzymes, indicating that the chemical modification does not prevent biodegradation. Complete degradation of the hydrogels occurred within 14 days in hyaluronidase (100 U/ml) and 3 days in collagenase (66 U/ml). Pattern transfer was employed to introduce a surface topography onto the hydrogel, which was able to orient cell growth. Furthermore, the hydrogels could be functionalized with the biomolecule neutravidin by incorporation of biotin along the HA backbone. This biotinylation approach may allow attachment of bioactive molecules that are conjugated to avidin. PMID- 15275811 TI - The influence of surface topography of ceramic abutments on the attachment and proliferation of human oral fibroblasts. AB - As different implant abutments are introduced to obtain a sufficient soft tissue barrier, the aim of this study was to determine the effect of three different surface modifications of densely sintered high-purity aluminium oxide on morphology, attachment and proliferation of human gingival fibroblasts. Fibroblasts were cultured on pressed aluminium oxide, milled, and then sintered to full density (1), on pressed, densely sintered (2), and on pressed, densely sintered and then polished surfaces (3). The different surfaces were analyzed using a confocal laser scanner, an atomic force microscope and a scanning electron microscope. The cell profile areas were measured using a semiautomatic interactive image analyzer and the figures were expressed as percent of attachment. The polished specimens had the smoothest surfaces and the roughest were the milled surfaces in terms of height deviation. No difference was found in the spacing between the peaks on the polished surfaces compared to the milled surfaces. Fibroblasts on the milled ceramic appeared to follow the direction of the fine irregularities on the surface. The analyses showed the polished surfaces had significantly higher percentages of initial cell attachment than the other surfaces (P < 0.05). After 3 days of cell culture, significantly more cells were attached to the milled and sintered surfaces than to the polished one, possibly indicating higher proliferation capacity on those types of surfaces. PMID- 15275812 TI - Control of focal adhesion dynamics by material surface characteristics. AB - The mechanisms of cell adhesion to the extracellular matrix (ECM) which are of fundamental importance for function, survival, and growth of cells involve the formation of focal adhesions to facilitate integrin signaling. Recently, it became evident that focal adhesions are not stable but move to enable cell migration and ECM formation. We examined the number, size, and dynamic behavior of focal adhesions in living MG-63 osteoblastic cells, which were cultured on titanium surfaces with different roughnesses and on stainless steel (SS). As a marker for focal adhesions we used GFP-tagged vinculin, a cytoskeletal protein. Focal adhesions were smaller on titanium and on SS than on collagen-coated glass coverslips. The corundum-blasted rough surface of titanium induced the smallest adhesions. On all the surfaces that we have tested, we observed a mobility of focal adhesions. On collagen-coated coverslips focal adhesions moved with a speed of 60 nm/min. The speed was reduced on titanium and still more restricted on SS. The topography did not affect the mobility of focal adhesions. We conclude that on the material surfaces that we have studied a reduced mobility of focal adhesions may strengthen the linkages between cell and ECM but impair the ability to dynamically organize and remodel the ECM. The results may have a great impact in the functional evaluation of tailored biomaterial surfaces for the application in tissue engineering. PMID- 15275813 TI - Hydrolysis of tetracalcium phosphate under a near-constant-composition condition- effects of pH and particle size. AB - Tetracalcium phosphate (TTCP) is a component of a number of calcium phosphate cements used clinically for bone defect repairs. The strength, phase composition, and solubility of the set cement are highly dependent on the reactions of the cement components during setting. This study investigated hydrolysis reactions of TTCP under solution compositions chosen to mimic the compositions of the cement liquid during setting. The study utilized a pseudo-constant-composition technique that allowed both the rate and stoichiometry of the reaction to be determined while the reaction proceeded under a specific, constantly held solution pH, thereby keeping a constant calcium-to-phosphate ratio in solution. The hydrolysis experiments were conducted using either a fine (median particle size 3.5 microm) or coarse (median particle size 13.2 microm) TTCP powder at pH 7, 8 and 10. Low crystalline calcium (Ca)-deficient hydroxyapatite (HA) was the product in all experiments. Both the solution pH and TTCP particle size produced significant effects on all aspects of the hydrolysis reaction. At a given pH, the fine TTCP produced a HA product with a greater Ca deficiency than did the coarse TTCP. For a given particle size, the Ca deficiency generally decreased with increasing pH. Hydrolysis reaction rate generally decreased with increasing pH or TTCP particle size. At pH 7 and 8, the solution was undersaturated with respect to TTCP and supersaturated with respect to HA, suggesting that the reaction rate was limited by TTCP dissolution. In contrast, at pH 10, the solution was approximately saturated with respect to TTCP and highly supersaturated with respect to HA, suggesting that HA formation was the rate-determining step of the reaction. The findings provided useful insights into the setting reaction mechanisms of TTCP containing calcium phosphate cements. PMID- 15275814 TI - Cytoprotection of PEG-modified adult porcine pancreatic islets for improved xenotransplantation. AB - Functional poly(ethylene glycol) (PEG) derivatives, including monosuccinimidyl PEG (MSPEG) with molecular weight (MW) of 2000 (2 kDa) as well as 5 kDa and disuccinimidyl PEG (DSPEG) with MW of 3 and 6 kDa, were synthesized and characterized. They were used to modify the surface of adult porcine islets for cytoprotection. The islets were isolated, purified and modified with functional PEG. Untreated porcine islets were used as control. An in vitro human antibody/complement-mediated cytotoxicity test based on the release of intracellular lactate dehydrogenase was used to evaluate cytotoxicity of human serum to the modified islets. In vitro cell viability was assessed using membrane integrity straining and islet metabolism in culture. In vitro islet functionality was evaluated by glucose-stimulated insulin release of islets in static incubation with human serum. In vivo islet functionality was evaluated by monitoring non-fasting blood glucose level in streptozotocin-induced diabetic (SCID) immunocompromized mice after intraportal transplantation of porcine islets. Results show that all the PEG derivatives used in the study showed significant in vitro and in vivo cytoprotections against cytotoxic effects elicited by human serum and diabetic SCID mice, respectively, to porcine islets. DSPEG derivatives combined with human albumin exhibited a better cytoprotection, as compared to MSPEG ones, due to the capacity of the succinimidyl groups to selectively react with amino groups of the albumin under physiological conditions. The effects of both MW and concentration of the PEG derivatives on cytoprotection were significant. It appears that this novel biotechnology will be an attractive approach for improved xenotransplantation of islets. PMID- 15275815 TI - Quantitative methods for analysis of integrin binding and focal adhesion formation on biomaterial surfaces. AB - Integrin binding and focal adhesion assembly are critical to cellular responses to biomaterial surfaces in biomedical and biotechnological applications. While immunostaining techniques to study focal adhesion assembly are well established, a crucial need remains for quantitative methods for analyzing adhesive structures. We present simple yet robust approaches to quantify integrin binding and focal adhesion assembly on biomaterial surfaces. Integrin binding to fibronectin and a RGD-containing synthetic peptide was quantified by sequentially cross-linking integrin-ligand complexes via a water-soluble homo-bifunctional cross-linker, extracting bulk cellular components in detergent, and detecting bound integrins by ELISA. Focal adhesion components (vinculin, talin, alpha actinin) localized to adhesion plaques were isolated from bulk cytoskeletal and cytoplasmic components by mechanical rupture at a plane close to the basal cell surface and quantified by Western blotting. These approaches represent simple and efficient methodologies to analyze structure-function relationships in cell material interactions. PMID- 15275816 TI - Bacterial cellulose as a potential scaffold for tissue engineering of cartilage. AB - Tissue constructs for cartilage with native mechanical properties have not been described to date. To address this need the bacterial cellulose (BC) secreted by Gluconacetobacter xylinus (= Acetobacter xylinum) was explored as a novel scaffold material due to its unusual material properties and degradability. Native and chemically modified BC materials were evaluated using bovine chondrocytes. The results indicate that unmodified BC supports chondrocyte proliferation at levels of approximately 50% of the collagen type II substrate while providing significant advantages in terms of mechanical properties. Compared to tissue culture plastic and calcium alginate, unmodified BC showed significantly higher levels of chondrocyte growth. Chemical sulfation and phosphorylation of the BC, performed to mimic the glucosaminoglycans of native cartilage, did not enhance chondrocyte growth while the porosity of the material did affect chondrocyte viability. The BC did not induce significant activation of proinflammatory cytokine production during in vitro macrophage screening. Hence, unmodified BC was further explored using human chondrocytes. TEM analysis and RNA expression of the collagen II from human chondrocytes indicated that unmodified BC supports proliferation of chondrocytes. In addition, ingrowth of chondrocytes into the scaffold was verified by TEM. The results suggest the potential for this biomaterial as a scaffold for tissue engineering of cartilage. PMID- 15275817 TI - The effect of pore size on cell adhesion in collagen-GAG scaffolds. AB - The biological activity of scaffolds used in tissue engineering applications hypothetically depends on the density of available ligands, scaffold sites at which specific cell binding occurs. Ligand density is characterized by the composition of the scaffold, which defines the surface density of ligands, and by the specific surface area of the scaffold, which defines the total surface of the structure exposed to the cells. It has been previously shown that collagen glycosaminoglycan (CG) scaffolds used for studies of skin regeneration were inactive when the mean pore size was either lower than 20 microm or higher than 120 microm (Proc. Natl. Acad. Sci., USA 86(3) (1989) 933). To study the relationship between cell attachment and viability in scaffolds and the scaffold structure, CG scaffolds with a constant composition and solid volume fraction (0.005), but with four different pore sizes corresponding to four levels of specific surface area were manufactured using a lyophilization technique. MC3T3 E1 mouse clonal osteogenic cells were seeded onto the four scaffold types and maintained in culture. At the experimental end point (24 or 48 h), the remaining viable cells were counted to determine the percent cell attachment. A significant difference in viable cell attachment was observed in scaffolds with different mean pore sizes after 24 and 48 h; however, there was no significant change in cell attachment between 24 and 48 h for any group. The fraction of viable cells attached to the CG scaffold decreased with increasing mean pore size, increasing linearly (R2 = 0.95, 0.91 at 24 and 48 h, respectively) with the specific surface area of the scaffold. The strong correlation between the scaffold specific surface area and cell attachment indicates that cell attachment and viability are primarily influenced by scaffold specific surface area over this range (95.9 150.5 microm) of pore sizes for MC3T3 cells. PMID- 15275818 TI - Biaxial mechanical properties of muscle-derived cell seeded small intestinal submucosa for bladder wall reconstitution. AB - Bladder wall replacement remains a challenging problem for urological surgery due to leakage, infection, stone formation, and extensive time needed for tissue regeneration. To explore the feasibility of producing a more functional biomaterial for bladder reconstitution, we incorporated muscle-derived cells (MDC) into small intestinal submucosa (SIS) scaffolds. MDC were harvested from mice hindleg muscle, transfected with a plasmid encoding for beta-galactosidase, and placed into single-layer SIS cell culture inserts. Twenty-five MDC and/or SIS specimens were incubated at 37 degrees C for either 10 or 20 days. After harvesting, mechanical properties were characterized using biaxial testing, and the areal strain under 1 MPa peak stress used to quantify tissue compliance. Histological results indicated that MDC migrated throughout the SIS after 20 days. The mean (+/-SE) areal strain of the 0 day control group was 0.182 +/- 0.027 (n=5). After 10 days incubation, the mean (+/-SE) areal strain in MDC/SIS was 0.247 +/- 0.014 (n=5) compared to 10 day control SIS 0.200 +/- 0.024 (n=6). After 20 days incubation, the mean areal strain of MDC/SIS was 0.255 +/- 0.019 (n=5) compared to control SIS 0.170 +/- 0.025 (n=5). Both 10 and 20 days seeded groups were significantly different (p=0.027) than that of incubated SIS alone, but were not different from each other. These results suggest that MDC growth was supported by SIS and that initial remodeling of the SIS ECM had occurred within the first 10 days of incubation, but may have slowed once the MDC had grown to confluence within the SIS. PMID- 15275820 TI - Hypothesis: cannabinoid therapy for the treatment of gliomas? AB - Gliomas, in particular glioblastoma multiforme or grade IV astrocytoma, are the most frequent class of malignant primary brain tumours and one of the most aggressive forms of cancer. Current therapeutic strategies for the treatment of glioblastoma multiforme are usually ineffective or just palliative. During the last few years, several studies have shown that cannabinoids-the active components of the plant Cannabis sativa and their derivatives--slow the growth of different types of tumours, including gliomas, in laboratory animals. Cannabinoids induce apoptosis of glioma cells in culture via sustained ceramide accumulation, extracellular signal-regulated kinase activation and Akt inhibition. In addition, cannabinoid treatment inhibits angiogenesis of gliomas in vivo. Remarkably, cannabinoids kill glioma cells selectively and can protect non-transformed glial cells from death. These and other findings reviewed here might set the basis for a potential use of cannabinoids in the management of gliomas. PMID- 15275819 TI - Locally delivered nanoencapsulated tyrphostin (AGL-2043) reduces neointima formation in balloon-injured rat carotid and stented porcine coronary arteries. AB - Local delivery of antiproliferative drugs encapsulated in biodegradable nanoparticles (NP) has shown promise as an experimental strategy for preventing restenosis development. A novel PDGFRbeta-specific tyrphostin, AGL-2043, was formulated in polylactide-based nanoparticles and was administered intraluminally to the wall of balloon-injured rat carotid and stented pig coronary arteries. The disposition and elimination kinetics within the vessel wall, as well as the antirestenotic potential of the novel drug and delivery system, were evaluated. The efficacy and the local drug elimination kinetics were affected by the size of the NP and the drug-carrier binding mode. Despite similar arterial drug levels 90 min after delivery in rats, small NP were more efficacious in comparison to large NP (90 and 160 nm, respectively). AGL-2043 selectively inhibited vascular SMC in a dose-dependent manner. The antiproliferative effect of nanoencapsulated tyrphostin was considerably higher than that of surface-adsorbed drug. In the pig model, intramural delivery of AGL-2043 resulted in reduced in-stent neointima formation in the coronary arteries over control despite similar degrees of wall injury. The results of this study suggest that locally delivered tyrphostin AGL 2043 formulated in biodegradable NP may be applicable for antirestenotic therapy independent of stent design or type of injury. PMID- 15275821 TI - Heterosynaptic co-activation of glutamatergic and dopaminergic afferents is required to induce persistent long-term potentiation. AB - The persistence of protein synthesis-dependent long-term potentiation (late-LTP) is thought to require heterosynaptic activation of both glutamate and neuromodulatory receptors in the hippocampus. The present series of experiments contrasts two alternative accounts of heterosynaptic activation. The original version of the synaptic-tag hypothesis of the variable persistence of LTP implied that neuromodulatory and glutamatergic activation could occur independently, albeit within a critical time-window; an alternative view is that there needs to be simultaneous co-activation of both receptors to trigger the up-regulation of relevant protein synthesis (Neuron 34 (2002) 235). Our findings include a replication, over 6 h post-LTP-induction, of earlier findings showing heterosynaptic influences on LTP persistence. Specifically, 'strong' tetanisation with multiple trains of stimulation of one input pathway in a conventional hippocampal slice preparation induces a D1/D5 receptor-dependent form of late-LTP that enables 'weak' tetanic stimulation to induce late-LTP on an independent pathway. However, we also observed that when the first pathway was tetanised in the presence of AP5, not only was no LTP observed on that pathway, but there was also no rescue of late-LTP on the second pathway. Thus, it appears that DA receptors must be co-activated with NMDA receptors in a common pool of neurons to enable LTP persistence, although late-LTP can still be induced by selective activation of glutamatergic synapses if this occurs at time periods shortly before or shortly after this essential coactivation. PMID- 15275822 TI - Regulation of transmitter release by high-affinity group III mGluRs in the supraoptic nucleus of the rat hypothalamus. AB - We analyzed the subtypes of group III metabotropic glutamate receptors (mGluRs) modulating inhibitory and excitatory transmission in the rat supraoptic nucleus. Bath application of the agonist l-AP4 at 200 microM, a concentration that activates all group III mGluR subtypes, inhibited the frequency but not the amplitude of miniature inhibitory and excitatory postsynaptic currents, indicating a presynaptic site of action. l-AP4 at low concentrations (10 microM), as well as ACPT-1 (50 microM), a specific mGluR III agonist, inhibited transmission at GABAergic and glutamatergic synapses to the same extent as 200 microM l-AP4. Because the potency of l-AP4 and ACPT-1 is much higher on mGluR4 and mGluR8 than on mGluR7, these results are consistent with the presence of high affinity group III mGluRs regulating transmitter release in this nucleus. In agreement with these findings, DCPG (30 microM), a selective mGluR8 agonist, induced a significant depression of inhibitory and excitatory synaptic currents. Group III mGluRs such as mGluR8, because of their high affinity for glutamate, are particularly well suited to detect small changes in the concentration of this excitatory amino acid in the extracellular space. Their presence, therefore, may favor the negative feedback control exerted by glutamate on its own release as well as the intersynaptic crosstalk mediated by glutamate spillover on adjacent synapses. PMID- 15275823 TI - Anxiolytic-like effects of MTEP, a potent and selective mGlu5 receptor agonist does not involve GABA(A) signaling. AB - Several lines of evidence suggest a crucial involvement of glutamate in the mechanism of action of anxiolytic drugs including the involvement of group I metabotropic glutamate (mGlu) receptors. Given the recent discovery of a selective and brain penetrable mGlu5 receptor antagonists, the effect of 3-[(2 methyl-1,3-thiazol-4-yl)ethynyl]-pyridine (MTEP), i.e. the most potent mGlu5 antagonist, was evaluated in established models of anxiety after single or repeated administration. We also studied if the anxiolytic effect of MTEP is mediated by mechanism involving the GABA-benzodiazepine (BZD) receptor complex. Experiments were performed on male Wistar rats or male Albino Swiss mice. The anxiolytic-like effects of MTEP were tested in the conflict drinking test and the elevated plus-maze test in rats as well as in the four-plate test in mice. MTEP (0.3-3.0 mg/kg) induced anxiolytic-like effects in the conflict drinking test (after single and repeated administration) and in the elevated plus-maze test in rats. In the four-plate test in mice, it exerted anxiolytic activity at a dose of 20 mg/kg. MTEP had no effect on the locomotor activity of animals. The anxiolytic like effect of MTEP was not changed by BZD antagonist flumazenil. Moreover, a synergistic interaction between non-effective doses of MTEP and diazepam was observed in the conflict drinking test. These data suggest that selective mGlu5 receptor antagonists mediated anxiolysis is not dependent on GABA-ergic system and that these agents may play a role in the therapy of anxiety. PMID- 15275824 TI - Behavioural effects of the novel AMPA/GluR5 selective receptor antagonist NS1209 after systemic administration in animal models of experimental pain. AB - The effects of systemic administration of the novel AMPA/GluR5 selective receptor antagonist NS1209 in animal models of experimental pain have been tested and compared with the AMPA receptor antagonist NBQX and the opiate morphine. In the mouse hot plate test, NS1209 (3-30 mg/kg, s.c. and i.p.) and morphine (3-30 mg/kg, s.c.) significantly increased the nociceptive response latency, whereas NBQX (3-30 mg/kg, i.p.) was ineffective. In the rat formalin test, a model of persistent pain, NS1209 (3 and 6 mg/kg, i.p.) and morphine (0.5 and 3 mg/kg, s.c.) produced dose-dependent reductions in second phase nociceptive behaviours, although NBQX (10 and 20 mg/kg, i.p.) was without effect. In the chronic constriction injury model of neuropathic pain, NS1209 (3 and 6 mg/kg, i.p.), NBQX (10 and 20 mg/kg, i.p.) and morphine (3 and 6 mg/kg, s.c.) all reduced mechanical allodynia and hyperalgesia responses to von Frey hair and pin prick stimulation of the injured hindpaw. NS1209 and morphine also reduced cold hypersensitivity in response to ethyl chloride stimulation of the injured hindpaw. At the doses associated with anti-nociceptive actions, no effects on motor performance as determined by the rotarod test were observed for any of the drugs tested. Thus, systemic administration of NS1209 at non-ataxic doses has marked analgesic actions comparable to those of morphine in a range of animal models of experimental pain. PMID- 15275825 TI - Characterisation of the effects of ATPA, a GLU(K5) kainate receptor agonist, on GABAergic synaptic transmission in the CA1 region of rat hippocampal slices. AB - Kainate receptors are implicated in a variety of physiological and pathological processes in the CNS. Previously we demonstrated that (RS)-2-amino-3-(3-hydroxy-5 tert-butylisoxazol-4-yl)propanoic acid (ATPA), a selective agonist for the GLU(K5) subtype of kainate receptor, depresses monosynaptically evoked inhibitory postsynaptic potentials (IPSPs) in the CA1 region of the rat hippocampus. In the current study, we provide a more detailed characterisation of this effect. Firstly, our data demonstrate a rank order of potency of domoate>kainate>ATPA>alpha-amino-3-(3-hydroxy-5-methyl-4-isoxalolyl)propionic acid Secondly, we confirm that the effects of ATPA are not mediated indirectly via the activation of gamma-aminobutyric acid receptors (i.e. either GABA(A) or GABA(B)). Thirdly, we show that the small increase in conductance induced by ATPA is insufficient to account for the depression of monosynaptic inhibition. Fourthly, we show that the effects of ATPA on IPSPs are antagonised by the GLU(K5)-selective antagonist (3S, 4aR, 6S, 8aR)-6-(4-carboxyphenyl)methyl 1,2,3,4,4a,5,6,7,8,8a-decahydroisoquinoline-3-carboxylic acid (LY382884). However, LY382884 is less potent as an antagonist of the effects of ATPA on IPSPs compared to its depressant effect on EPSPs. PMID- 15275826 TI - Lipid emulsions reduce NMDA-evoked currents. AB - Membrane currents conducted by the NMDA receptor channels were investigated in cultured cortical neurons and TsA cells transfected with NR1-1a/NR2A subunits of the NMDA receptor. The whole-cell recording technique was used. Current transients evoked by bath application of NMDA for 5 s were characterized by a fast peak and a slow decay to 46.1 +/-15.5% of the peak level at the end. When NMDA was applied in combination with various lipid emulsions (Intralipid, ClinOleic, Lipofundin or Abbolipid, the NMDA-induced currents were reduced, although this reduction did not affect the fast peak, it did affect the decay phase. The amount of reduction depended on the concentration of the lipids (in the case of Abbolipid diluted at 1:40, the current at the end of the 5-s drug application was approximately 2/3 of control). When Abbolipid was applied 40 s before NMDA, peak and late current were reduced to approximately 2/3. The effect of current reduction was the same at either of the two chosen membrane potentials (-80 and +40 mV) which indicates that the effect was not mediated by contamination of the emulsions with Mg(2+). The current reduction produced by Abbolipid was about the same in native neuronal cells and in TsA cells expressing the NR1-1a/NR2A subunits. The current-reducing effect of the lipid emulsions may add to the anesthetic, analgesic and neuroprotective effects seen with hypnotics administered by way of lipid carriers. PMID- 15275827 TI - Distinct properties of carbachol- and DHPG-induced network oscillations in hippocampal slices. AB - The aim of this study was to compare and contrast the properties of gamma oscillations induced by activation of muscarinic acetylcholine or metabotropic glutamate receptors in the CA3 region of rat hippocampal slices. Both carbachol and the group I metabotropic glutamate receptor agonist, (RS)-3,5 dihydroxyphenylglycine (DHPG), induced network oscillations in the gamma frequency range (30-100 Hz). The M1 muscarinic receptor antagonist, pirenzepine, blocked carbachol-, but enhanced DHPG-induced oscillations, whereas LY 341495, an antagonist at metabotropic glutamate receptors, abolished DHPG-, but left carbachol-induced oscillations unchanged. There were significant differences in the peak frequency, maximal power, and spectral width of the two oscillations. Pharmacological experiments showed that both types of oscillation depend on fast excitatory and inhibitory synaptic transmission. Interestingly, activation of neurokinin-1 receptors by substance P fragment or enhancement of inhibitory synaptic currents by the benzodiazepine ligand, zolpidem, boosted DHPG-, but reduced the power of carbachol-induced oscillations. These results suggest that, although carbachol and DHPG might activate similar conductances in individual pyramidal cells, the oscillations they induce in slices involve different network mechanisms, most likely by recruiting distinct types of GABAergic interneuron. PMID- 15275828 TI - The interaction of cannabinoids and opioids on pentylenetetrazole-induced seizure threshold in mice. AB - Cannabinoid and opioid receptor agonists show functional interactions in a number of their physiological effects. Regarding the seizure-modulating properties of both classes of receptors, the present study examined the possibility of a functional interaction between these receptors. We used acute systemic administration of cannabinoid selective CB(1) receptor agonist (ACPA) and antagonist (AM251) and opioid receptor agonist (morphine) and antagonists (naltrexone and norbinaltorphimine) in a model of clonic seizure induced by pentylenetetrazole (PTZ). Acute administration of ACPA (1.5-2 mg/kg) increased the PTZ-induced seizure threshold. In contrast, AM251 (0.5-2 mg/kg) dose dependently decreased the seizure threshold. Low dose of AM251 (0.5 mg/kg), which did not alter seizure threshold by itself, reversed the anticonvulsant effect of ACPA (2 mg/kg), showing a CB(1) receptor-mediated mechanism. Naltrexone (1 or 10 mg/kg) but not specific kappa-opioid receptor antagonist norbinaltorphimine (5 mg/kg) completely reversed the anticonvulsant effect of ACPA (2 mg/kg). Moreover, the combination of the lower doses of AM251 (0.5 mg/kg) and naltrexone (0.3 mg/kg) had an additive effect in blocking the anticonvulsant effect of ACPA. In accordance with previous reports, morphine exerted biphasic effects on clonic seizure threshold with anticonvulsant effect at lower (0.5-1 mg/kg) and proconvulsant effect at a higher (30 mg/kg) doses. The pretreatment with AM251 blocked the anticonvulsant effect of morphine at 1 mg/kg, while pretreatment with ACPA (1 mg/kg) potentiated the anticonvulsant effect of morphine at 0.5 mg/kg. The proconvulsant effect of morphine at 30 mg/kg was also inhibited by AM251 (2 mg/kg). A similar interaction between cannabinoids and opioids was also detected on their anticonvulsant effects against the generalized tonic-clonic model of seizure. In conclusion, cannabinoids and opioids show functional interactions on modulation of seizure susceptibility. PMID- 15275829 TI - The alpha3 and beta4 nicotinic acetylcholine receptor subunits are necessary for nicotine-induced seizures and hypolocomotion in mice. AB - Binding of nicotine to nicotinic acetylcholine receptors (nAChRs) elicits a series of dose-dependent behaviors that go from altered exploration, sedation, and tremors, to seizures and death. nAChRs are pentameric ion channels usually composed of alpha and beta subunits. A gene cluster comprises the alpha3, alpha5 and beta4 subunits, which coassemble to form functional receptors. We examined the role of the beta4 subunits in nicotine-induced seizures and hypolocomotion in beta4 homozygous null (beta4 -/-) and alpha3 heterozygous (+/-) mice. beta4 -/- mice were less sensitive to the effects of nicotine both at low doses, measured as decreased exploration in an open field, and at high doses, measured as sensitivity to nicotine-induced seizures. Using in situ hybridization probes for the alpha3 and alpha5 subunits, we showed that alpha5 mRNA levels are unchanged, whereas alpha3 mRNA levels are selectively decreased in the mitral cell layer of the olfactory bulb, and the inferior and the superior colliculus of beta4 -/- brains. alpha3 +/- mice were partially resistant to nicotine-induced seizures when compared to wild-type littermates. mRNA levels for the alpha5 and the beta4 subunits were unchanged in alpha3 +/- brains. Together, these results suggest that the beta4 and the alpha3 subunits are mediators of nicotine-induced seizures and hypolocomotion. PMID- 15275830 TI - Co-administration of memantine has no effect on the in vitro or ex vivo determined acetylcholinesterase inhibition of rivastigmine in the rat brain. AB - Rivastigmine, a cholinesterase inhibitor, is successfully used for the symptomatic therapy of Alzheimer's disease (AD) in the clinic. The drug has a very low potential for drug-drug interactions, as has been demonstrated within large clinical trials. Memantine, recently approved by the FDA for the treatment of moderate to severe AD, acts as a low affinity, non-competitive NMDA antagonist, on a completely different neurotransmitter system, the glutamatergic system. Given the different sites of action, the possibility to combine a cholinergic with a glutamatergic intervention as potentially superior AD therapy has recently been proposed. In vitro studies have demonstrated that memantine, when added to reversible AChE inhibitors, such as tacrine, donepezil or galantamine, did not interfere with the inhibitory action of any of these drugs. The results from the present study provide evidence that rivastigmine as a pseudo irreversible (or slow-reversible) AChE inhibitor shares this property described for reversible inhibitors, since memantine (1-100 microM), irrespective of whether given prior to or after rivastigmine did not influence rivastigmine's AChE inhibition in vitro. A similar observation was also made under in vivo conditions (ex vivo measurements): following a 21 day chronic, oral administration of 6 micromol/kg rivastigmine alone or of a combination of rivastigmine plus memantine (6 micromol/kg p.o. of either of the two compounds), an identical degree of AChE inhibition was observed. The concentrations of rivastigmine, its metabolite NAP 226-90 and memantine were measured in the brain of the same animals. Following an equimolar oral dose (6 micromol/kg) of both compounds, the brain level of memantine exceeded that of rivastigmine + metabolite, by a factor of around 30, when measured 2 h after the final dosing, irrespective of the duration of treatment (acute, for 3 or 21 days). This indicates that neither of the two drugs showed accumulation but also, and more importantly, that memantine does not modulate the prime therapeutic action of rivastigmine (AChE inhibition) in vitro or in vivo. Clinical trials using a combination of both drugs will provide a final proof of whether a combination therapy would lead to an increased efficacy in AD patients. PMID- 15275831 TI - Striatal adenosine A(2A) receptor blockade increases extracellular dopamine release following l-DOPA administration in intact and dopamine-denervated rats. AB - The influence of the selective adenosine A(2A) receptor antagonist ZM 241385 on exogenous l-DOPA-derived dopamine (DA) release in intact and dopamine-denervated rats was studied using an in vivo microdialysis in freely moving animals. Local infusion of l-DOPA (2.5 microM) produced a marked increase in striatal extracellular DA level in intact and malonate-lesioned rats. Intrastriatal perfusion of ZM 241385 (50-100 microM) had no effect on basal extracellular DA level, but enhanced dose-dependently the l-DOPA-induced DA release in intact and malonate-lesioned animals. A non-selective adenosine A(2A) receptor antagonist DMPX (100 microM), similarly to ZM 241385, accelerated conversion of l-DOPA in intact and malonate-denervated rats. This effect was not produced by the adenosine A(1) receptor antagonist, CPX (10-50 microM). However, ZM 241385 did not affect the l-DOPA-induced DA release in rats pretreated with reserpine (5 mg/kg i.p.) and alpha-methyl-p-tyrosine (AMPT, 300 mg/kg i.p.). Obtained results indicate that blockade of striatal adenosine A(2A) receptors increases the l-DOPA derived DA release possibly by indirect mechanism exerted on DA terminals, an effect dependent on striatal tyrosine hydroxylase activity. Selective antagonists of adenosine A(2A) receptors may exert a beneficial effect at early stages of Parkinson's disease by enhancing the therapeutic efficacy of l-DOPA applied exogenously. PMID- 15275832 TI - Adenosine acting via A1 receptors, controls the transition to status epilepticus like behaviour in an in vitro model of epilepsy. AB - Adenosine has powerful inhibitory effects in the central nervous system. In this study, we aim to understand how adenosine controls the progression of seizure like events (SLEs) in a seizure-prone region of the brain, the entorhinal cortex. We chose to use a low Mg(2+) model of epilepsy in an in vitro slice preparation where, in the entorhinal cortex, SLEs progress into a type of epileptiform activity called late recurrent discharges (LRDs) that bear resemblance to status epilepticus. Adenosine, acting via its A1 receptor, exerted powerful inhibitory effects to prevent the spontaneous progression to LRDs while the potent A1 receptor antagonist, DPCPX, accelerated the progression in a concentration dependent manner. The spontaneous progression from SLEs to LRDs was associated with a decline in total cellular ATP levels and studies with metabolic inhibitors indicated a key role for the production of endogenous adenosine from ATP. We therefore hypothesise that when ATP becomes rate limiting, extracellular adenosine levels fall, the normal inhibitory brake is removed and the progression from SLEs to LRDs or status epilepticus-like activity can ensue. Moreover, under these conditions, inhibition of the adenine nucleotide salvage pathways reversed the status epilepticus-like activity. Our findings suggest a powerful role for adenosine for the control of the progression to status epilepticus-like activity in an epilepsy model that is refractory to most anti-epileptic drugs. On this basis, manipulation of adenine nucleotide metabolism may represent a potential therapeutic approach for the treatment of status epilepticus. PMID- 15275833 TI - Potency of catecholamines and other L-tyrosine derivatives at the cloned mouse adrenergic receptors. AB - The adrenergic system is a neuromodulatory system whose endogenous ligands are considered to be the catecholamines norepinephrine (NE) and epinephrine (E). Evidence suggests that the catecholamine dopamine (DA) may also activate adrenergic signaling. Further, tyramine (TA) and octopamine (OA) are other monoamines that can be produced in catecholaminergic cells when tyrosine hydroxylase activity is low or absent, as in some genetic mouse models of adrenergic function. Here, we systematically examine the ability of these L tyrosine-derived monoamines to activate all 10 known isoforms of the cloned mouse adrenergic receptors expressed in Chinese hamster ovary cells. In comparison to NE or E, DA is nearly as efficacious in this system but is from 1 to 4 orders of magnitude less potent. In comparison to DA, OA has roughly equivalent potency but is usually only a partial agonist. TA is either very weak or lacks agonism. Of note, all three mouse alpha(1) receptors increase cAMP, in contrast to results reported for human alpha(1d) receptors. In addition, a 12-amino acid hemagglutinin epitope tag added to the N-terminus of alpha(2) receptors selectively enhances the potency of NE approximately 10- to 100-fold, indicating that caution should be applied when interpreting physiological results from experiments using modified receptors. PMID- 15275834 TI - Flufenamic acid is a pH-dependent antagonist of TRPM2 channels. AB - Like a number of other TRP channels, TRPM2 is a Ca(2+)-permeable non-selective cation channel, the activity of which is regulated by intracellular and extracellular Ca(2+). A unique feature of TRPM2 is its activation by ADP-ribose and chemical species that arise during oxidative stress, for example, NAD(+) and H(2)O(2). These properties have lead to proposals that this channel may play a role in the cell death produced by pathological redox states. The lack of known antagonists of this channel have made these hypotheses difficult to test. Here, we demonstrate, using patch clamp electrophysiology, that the non-steroidal anti inflammatory compound flufenamic acid (FFA) inhibits recombinant human TRPM2 (hTRPM2) as well as currents activated by intracellular ADP-ribose in the CRI-G1 rat insulinoma cell line. All concentrations tested in a range from 50 to 1000 microM produced complete inhibition of the TRPM2-mediated current. Following FFA removal, a small (typically 10-15%) component of current was rapidly recovered (time constant approximately 3 s), considerably longer periods in the absence of FFA produced no further current recovery. Reapplication of FFA re-antagonised the recovered current and subsequent FFA washout produced recovery of only a small percentage of the reblocked current. Decreasing extracellular pH accelerated FFA inhibition of TRPM2. Additional experiments indicated hTRPM2 activation was required for FFA antagonism to occur and that the generation of irreversible antagonism was preceded by a reversible component of block. FFA inhibition could not be induced by intracellular application of FFA. ADP-ribose activated currents in the rat insulinoma cell line CRI-G1 were also antagonised by FFA with concentration- and pH-dependent kinetics. In contrast to the observations made with hTRPM2, antagonism of ADP-ribose activated currents in CRI-G1 cells could be fully reversed following FFA removal. These experiments suggest that FFA may be a useful tool antagonist for studies of TRPM2 function. PMID- 15275835 TI - Neonatal dexamethasone treatment affects social behaviour of rats in later life. AB - Synthetic glucocorticoids, like dexamethasone (DEX), have been frequently administered to premature infants to prevent chronic lung disease. Major concern has arisen about the long-term neurodevelopmental sequelae of this DEX treatment. In the present study, we found that neonatal DEX treatment in rats, using a treatment protocol resembling the one used in the clinical situation, increased social play behaviour in juvenile life. Furthermore, neonatal DEX treatment increased sexual motivation and intromission behaviour in the bi-level chamber, decreased submissive behaviour during an aggressive encounter, and impaired social memory in adulthood. These changes in social behaviour are not due to a general behavioural impairment since anxiety behaviour in the elevated plus maze and exploratory activity in the open-field were not affected in DEX rats. In addition, DEX rats showed no alteration in the total duration of social interest or social activity during a social interaction test. These effects of neonatal DEX treatment on behaviour later in life likely result from neurodevelopmental actions of the hormone since we found no differences in received maternal care between DEX and SAL treated pups. Together these results indicate that neonatal treatment with DEX selectively alters aspects of the behavioural response to social challenges. Thus, neonatal DEX treatment may lead to inappropriate interactions with conspecifics later in life. These data therefore warrant investigation of lasting and potentially adverse effects of treatment of human neonates with DEX on social functioning. PMID- 15275836 TI - Do microcracks decrease or increase fatigue resistance in cortical bone? AB - Fatigue of cortical bone produces microcracks; it has been hypothesized that these cracks are analogous to those occurring in engineered composite materials and constitute a similar mechanism for fatigue resistance. However, the numbers of these linear microcracks increase substantially with age, suggesting that they contribute to increased fracture incidence among the elderly. To test these opposing hypotheses, we fatigued 20 beams of femoral cortical bone from elderly men and women in load-controlled four point bending having initial strain ranges of 3000 or 5000 microstrain. Loading was stopped at fracture or 10(6) cycles, whichever occurred first, and microcrack density and length were measured in the loaded region and in a control region that was not loaded. We studied the dependence of fatigue life and induced microdamage on initial microdamage, cortical region, subject gender and age, and several other variables. When the effect of modulus variability was controlled, longer fatigue life was associated with higher rather than lower initial crack density, particularly in the medial cortex. The increase in crack density following fatigue loading was greater in specimens from older individuals and those initially having longer microcracks. Crack density increased as much in specimens fatigued short of the failure point as in those that fractured, and microcracks were, on average, shorter in specimens with greater numbers of resorption spaces, a measure of remodeling rate. PMID- 15275837 TI - Effects of shortening on stretch-induced force enhancement in single skeletal muscle fibers. AB - The main purpose of this study was to evaluate the effects of shortening on the stretch-induced force enhancement in single muscle fibers, and indirectly test the hypothesis that force enhancement may be associated with the engagement of a passive element upon activation. Fibers were placed on the descending limb of the force-length relationship, and stretch and shortening contractions were performed. Fibers underwent two sets of shortening-stretch cycles. First, fibers were shortened by a fixed amplitude and speed (10% fiber length, and at 40% fiber length/s), and then were stretched (10% fiber length, and at 40% fiber length/s) immediately following shortening, or 500 or 1000 ms following the shortening. Second, fibers were shortened by varying amounts (5%, 10% and 15% fiber length) and at a constant speed (40% fiber length/s) immediately preceding a given fiber stretch (10% fiber length, and at 40% fiber length/s). When stretching was immediately preceded by shortening, force enhancement was decreased proportionally with the shortening magnitude. When intervals were introduced between shortening and stretch, the effects of shortening on the stretch-induced force enhancement became less prominent. We concluded that, in contrast to published suggestions, shortening affects the stretch-induced force enhancement in an amplitude-dependent manner in single fibers, as it does in whole muscles, but this effect is diminished by increasing the time period between the shortening and stretch phases. PMID- 15275838 TI - A three-dimensional digital image correlation technique for strain measurements in microstructures. AB - A three-dimensional digital image correlation technique is presented for strain measurements in open-cell structures such as trabecular bone. The technique uses high-resolution computed tomography images for displacement measurements in the solid structure. In order to determine the local strain-state within single trabeculae, a tetrahedronization method is used to fill the solid structure with tetrahedrae. Displacements are calculated at the nodes of the tetrahedrae. The displacement data is subsequently converted to a deformation tensor in each of the tetrahedral element centers with a least-squares estimation method. Because the trabeculae are represented by a mesh, it is possible to deform this mesh according to the deformation tensor and, at the same time, visualize the calculated local strain in the deformed mesh with a finite element post processing tool. In this way, the deformation of a single trabecula from an aluminum foam sample was determined and validated with rendered images of the three-dimensional sample. A precision analysis showed that a rigid translation or rotation does not affect the accuracy. Typical values for the standard deviation in the displacement and strain components are 2.0 microm and 0.01, respectively. Presently, the precision limits the technique to strain measurements beyond the yield strain. PMID- 15275839 TI - Gastrocnemius and soleus lengths in cerebral palsy equinus gait--differences between children with and without static contracture and effects of gastrocnemius recession. AB - Equinus gait is one of the most common abnormalities in children with cerebral palsy. Although it is generally assumed that the calf muscles are abnormally short in equinus gait, no studies have been done to confirm that the muscles are short and that this shortness contributes to the equinus. This study used musculoskeletal modeling combined with computerized gait analysis to examine medial gastrocnemius (MGAS), lateral gastrocnemius (LGAS), and soleus (SOL) musculotendinous lengths during equinus gait in children with cerebral palsy. All three muscles were abnormally short during equinus gait whether or not the children had equinus contractures (P < or = 0.005). Children with static contractures had shorter maximum static MGAS and LGAS lengths than children with dynamic equinus (P < or = 0.002). The children with static contractures had ratios of peak dynamic length to maximum static length close to 1.0 for MGAS and LGAS (1.005 +/- 0.015) but lower ratios for SOL (0.984 +/- 0.024). For the children with static contracture, these ratios did not change significantly after gastrocnemius recession (P > or = 0.14) because both static and dynamic lengths increased postoperatively (P < or = 0.04). These results support the current clinical understanding of the role of calf "tightness" in equinus gait, including the appropriateness and effectiveness of gastrocnemius recession for children with equinus contracture. PMID- 15275840 TI - Effect of the "squat protective response" on impact velocity during backward falls. AB - Risk for injury during a fall depends on the position and velocity of the body segments at the moment of impact. One technique for reducing impact velocity is to absorb energy in the lower extremity muscles during descent, as occurs during squatting or sitting. However, the protective value of this response may depend on the time during descent when the response is initiated. We tested this hypothesis by conducting backward falling experiments with young women (n = 23; aged 21-29 years), who fell onto a soft gymnasium mattress after being suddenly releasing from an inclined position. In trials where subjects were released from a 5 degrees lean, average impact velocities were reduced by 18% when squatting was utilized as opposed to inhibited. Furthermore, increases in the release angle caused an increase in average impact velocity of 8% between lean angles of 2 degrees and 5 degrees, and 7% between lean angles of 5 degrees and 12 degrees. This was due to declines in peak extensor torques and peak flexion rotations, and corresponding reductions in both joint work and potential energy at impact. These results suggest that squatting during descent reduces impact severity, but the effectiveness of the response depends on the stage during descent when it is initiated, diminishing in benefit as the fall progresses and the state of imbalance grows increasingly severe. PMID- 15275841 TI - Are in vivo and in situ brain tissues mechanically similar? AB - Brain tissue mechanical properties have been well-characterized in vitro, and were found to be inhomogeneous, nonlinear anisotropic and influenced by neurological development and postmortem time interval prior to testing. However, brain in vivo is a vascularized tissue, and there is a paucity of information regarding the effect of perfusion on brain mechanical properties. Furthermore, mechanical properties are often extracted from preconditioned tissue, and it remains unclear if these properties are representative of non-preconditioned tissue. We present non-preconditioned (NPC) and preconditioned (PC) relaxation responses of porcine brain (N = 10) obtained in vivo, in situ and in vitro, at anterior, mid and posterior regions of the cerebral cortex during 4mm indentations at either 3 or 1 mm/s. Material property characteristics showed no dependency on the site tested, thus revealing that cortical gray matter on the parietal and frontal lobes can be considered homogenous. In most cases, preconditioning decreased the shear moduli, with a more pronounced effect in the dead (in situ and in vitro) brain. For most conditions, it was found that only the long-term time constant of relaxation (tau > 20 s) significantly decreased from in vivo to in situ modes (p < 0.02), and perfusion had no effect on any other property. These findings support the concept that perfusion does not affect the stiffness of living cortical tissue. PMID- 15275842 TI - Effect of the pole--human body interaction on pole vaulting performance. AB - The purposes of this study were: (a) to examine the interactions between the athlete and the pole and the possibility for the athlete to take advantage of the pole's elasticity by means of muscular work and (b) to develop performance criteria during the interaction between the athlete and the pole in pole vaulting. Six athletes performed 4-11 trials each, at 90% of their respective personal best performance. All trials were recorded using four synchronized, genlocked video cameras operating at 50 Hz. The ground reaction forces exerted on the bottom of the pole were measured using a planting box fixed on a force plate (1000 Hz). The interaction between athlete and pole may be split into two parts. During the first part, energy is transferred into the pole and the total energy of the athlete decreases. The difference between the energy decrease of the athlete and the pole energy is an indicator of the energy produced by the athletes by means of muscular work (criterion 1). During the second part of the interaction, energy is transferred back to the athlete and the total energy of the athlete increases. The difference between the returned pole energy and the amount of energy increase of the athlete defines criterion 2. In general, the function of the pole during the interaction is: (a) store part of the kinetic energy that the athlete achieved during the run up as strain energy and convert this strain energy into potential energy of the athlete, (b) allow the active system (athlete) to produce muscular work to increase the total energy potential. PMID- 15275843 TI - Three-dimensional load measurements in an external fixator. AB - On the basis of a six-degree-of-freedom adjustable fracture reduction hexapod external fixator, a system which can be used for measuring axial and shear forces as well as torsion and bending moments in the fixator in vivo was developed. In a pilot study on 9 patients (7 fresh fractures and 2 osteotomies of the tibia), the load in the fixator during the healing process was measured after 2, 4, 8 and 12 weeks and at fixator removal. The measured values enabled both the type of fracture to be determined as well as the monitoring of the healing process. In well-reduced type A3 fractures small axial (direction of the bone axis) forces were found in the fixator. A2, B2 and C3 fractures showed distinct axial forces, which decreased during the healing process, according to an increasing load transfer over the bone. Bending moments in the fixator showed good correspondence with the clinical healing process, except in the case of a C3 fracture. A combination of bending moment and axial force proved to be particularly suitable to assess fracture healing. In transverse fractures, the well-known resorption phenomenon of bone in the fracture gap at approximately 4 weeks was detected by the system. Compared with other external fixator load measurements in vivo, the hexapod offers the advantage of being able to measure all forces and moments in the fixator separately and with a relatively simple mechanical arrangement. In our opinion, it will be possible to control fracture healing using this system, thereby minimizing radiation exposure from radiographs. Furthermore, the measurement system is a step towards the development of external fixator systems that enable automatic adjustments of the callus mechanical situation ("automatic dynamization") and inform the patients about the optimal weight bearing of their extremity ("intelligent fixator"). PMID- 15275844 TI - Load transfer mechanics between trans-tibial prosthetic socket and residual limb- dynamic effects. AB - The effects of inertial loads on the interface stresses between trans-tibial residual limb and prosthetic socket were investigated. The motion of the limb and prosthesis was monitored using a Vicon motion analysis system and the ground reaction force was measured by a force platform. Equivalent loads at the knee joint during walking were calculated in two cases with and without consideration of the material inertia. A 3D nonlinear finite element (FE) model based on the actual geometry of residual limb, internal bones and socket liner was developed to study the mechanical interaction between socket and residual limb during walking. To simulate the friction/slip boundary conditions between the skin and liner, automated surface-to-surface contact was used. The prediction results indicated that interface pressure and shear stress had the similar double-peaked waveform shape in stance phase. The average difference in interface stresses between the two cases with and without consideration of inertial forces was 8.4% in stance phase and 20.1% in swing phase. The maximum difference during stance phase is up to 19%. This suggests that it is preferable to consider the material inertia effect in a fully dynamic FE model. PMID- 15275845 TI - Heel-shoe interactions and the durability of EVA foam running-shoe midsoles. AB - A finite element analysis (FEA) was made of the stress distribution in the heelpad and a running shoe midsole, using heelpad properties deduced from published force-deflection data, and measured foam properties. The heelpad has a lower initial shear modulus than the foam (100 vs. 1050 kPa), but a higher bulk modulus. The heelpad is more non-linear, with a higher Ogden strain energy function exponent than the foam (30 vs. 4). Measurements of plantar pressure distribution in running shoes confirmed the FEA. The peak plantar pressure increased on average by 100% after 500 km run. Scanning electron microscopy shows that structural damage (wrinkling of faces and some holes) occurred in the foam after 750 km run. Fatigue of the foam reduces heelstrike cushioning, and is a possible cause of running injuries. PMID- 15275846 TI - Functional significance of the coupling between head and jaw movements. AB - When humans open or close the jaw they also move the head. Unintentionally, it rotates backwards when the jaw opens and returns upon jaw closure. We hypothesized that this mutual movement coupling is related to the muscles in the floor of the mouth. A biomechanical model was applied to comprehend the functional significance of this movement coupling. As the jaw opened the jaw opening muscles shortened and became less forceful. Meanwhile they had to stretch the jaw closing muscles. The simulations showed that a simultaneous head extension facilitated jaw opening. A possible functional significance for the coupling between head and jaw movements is that it can extend jaw gape. Head extension can contribute to a wider jaw gape by on the one hand a reduced shortening of the jaw opening muscles and on the other hand by a reorientation of these muscles so that they obtain a more favorable position for jaw opening. PMID- 15275847 TI - A method to combine numerical optimization and EMG data for the estimation of joint moments under dynamic conditions. AB - To solve the problem of muscle redundancy at the level of opposing muscle groups, an alternative method to inverse dynamics must be employed. Considering the advantages of existing alternatives, the present study was aimed to compute knee joint moments under dynamic conditions using electromyographic (EMG) signals combined with non-linear constrained optimization in a single routine. The associated mathematical problems accounted for muscle behavior in an attempt to obtain accurate predictions of the resultant moment as well as physiologically realistic estimates of agonist and antagonist moments. The experiment protocol comprised (1) isometric trials to determine the most effective EMG processing for the prediction of the resultant moment and (2) stepping-in-place trials for the calculation of joint moments from processed EMG under dynamic conditions. Quantitative comparisons of the model predictions with the output of a biological based model, showed that the proposed method (1) produced the most accurate estimates of the resultant moment and (2) avoided possible inconsistencies by enforcing appropriate constraints. As a possible solution for solving the redundancy problem under dynamic conditions, the proposed optimization formulation also led to realistic predictions of agonist and antagonist moments. PMID- 15275848 TI - Analysis of human mandibular mechanics based on screw theory and in vivo data. AB - In this paper the mechanics of human mandibular function is described in terms of the associated screws. The two distinct, yet related features of jaw mechanics, involving the motion itself as well as the forces, are both functions of the anatomical constraints, namely the contact areas that exist within the temporomandibular joint, and the forces of the muscles and tendons that allow motion to occur. The relationships that exist between these two aspects of jaw motion are identified in this paper showing that muscle forces can be uniquely represented in terms of the action screw. This new approach to analyzing the mechanics of jaw-motion also incorporates the previously studied motion screw or helical axis. A consistent dynamic model is formulated where the action screw is used to represent the action of the closing muscle forces while the moment arms of the muscle forces are determined about the motion screw representing mandibular kinematics. The action screw formulation is verified using in vivo motion data and MR image information for a single asymptomatic subject. The results confirm the feasibility of the method and its application in dental research. A general increase in the mechanical advantage of most muscles, in the distance between action and motion screws as well as in the expended energy towards the end of the jaw-closing phase was observed. Asymmetries in the distribution of muscle force magnitudes appeared to influence the resultant force and moment of the action screw but had little effect on its spatial location. The method presented is intended to facilitate understanding of mandibular function and dysfunction. PMID- 15275849 TI - Contribution of inter-site variations in architecture to trabecular bone apparent yield strains. AB - Apparent yield strains for trabecular bone are uniform within an anatomic site but can vary across site. The overall goal of this study was to characterize the contribution of inter-site differences in trabecular architecture to corresponding variations in apparent yield strains. High-resolution, small deformation finite element analyses were used to compute apparent compressive and tensile yield strains in four sites (n = 7 specimens per site): human proximal tibia, greater trochanter, femoral neck, and bovine proximal tibia. These sites display differences in compressive, but not tensile, apparent yield strains. Inter-site differences in architecture were captured implicitly in the model geometries, and these differences were isolated as the sole source of variability across sites by using identical tissue properties in all models. Thus, the effects inter-site variations in architecture on yield strain could be assessed by comparing computed yield strains across site. No inter-site differences in computed yield strains were found for either loading mode (p > 0.19), indicating that, within the context of small deformations, inter-site variations in architecture do not affect apparent yield strains. However, results of ancillary analyses designed to test the validity of the small deformation assumption strongly suggested that the propensity to undergo large deformations constitutes an important contribution of architecture to inter-site variations in apparent compressive yield strains. Large deformations substantially reduced apparent compressive, but not tensile, yield strains. These findings indicate the importance of incorporating large deformation capabilities in computational analyses of trabecular bone. This may be critical when investigating the biomechanical consequences of trabecular thinning and loss. PMID- 15275850 TI - Comparison of three methods to estimate the center of mass during balance assessment. AB - Evaluation of postural control is generally based on the interpretation of the center of pressure (COP) and the center of mass (COM) time series. The purpose of this study is to compare three methods to estimate the COM which are based on different biomechanical considerations. These methods are: (1) the kinematic method; (2) the zero-point-to-zero-point double integration technique (GLP) and (3) the COP low-pass filter method (LPF). The COP and COM time series have been determined using an experimental setup with a force plate and a 3D kinematic system on six healthy young adult subjects during four different 30 s standing tasks: (a) quiet standing; (b) one leg standing; (c) voluntary oscillation about the ankles and (d) voluntary oscillation about the ankles and hips. To test the difference between the COM trajectories, the root mean square (RMS) differences between each method (three comparisons) were calculated. The RMS differences between kinematic-LPF and GLP-LPF are significantly larger than kinematic-GLP. Our results show that the GLP method is comparable to the kinematic method. Both agree with the unified theory of balance during upright stance. The GLP method is attractive in the clinical perspective because it requires only a force plate to determine the COP-COM variable, which has been demonstrated to have a high reliability. PMID- 15275851 TI - Use of pressure insoles to calculate the complete ground reaction forces. AB - A method to calculate the complete ground reaction force (GRF) components from the vertical GRF measured with pressure insoles is presented and validated. With this approach it is possible to measure several consecutive steps without any constraint on foot placement and compute a standard inverse dynamics analysis with the estimated GRF. PMID- 15275852 TI - Anisotropic viscoelastic properties of cortical bone. AB - Relaxation Young's modulus of cortical bone was investigated for two different directions with respect to the longitudinal axis of bone (bone axis, BA): the modulus parallel (P) and normal (N) to the BA. The relaxation modulus was analyzed by fitting to the empirical equation previously proposed for cortical bones, i.e., a linear combination of two Kohlraush-Williams-Watts (KWW) functions (Iyo et al., 2003. Biorheology, submitted): E(t)=E0 (A1 exp[-(t/tau1)beta]+(1-A1) exp[-(t/tau2)gamma]), [0 < A1, beta, gamma < 1], where E0 is the initial modulus value E0. Tau1 and tau2(>>tau1) are characteristic times of the relaxation, A1 is the fractional contribution of the fast relaxation (KWW1 process) to the whole relaxation process, and beta and gamma are parameters describing the shape of the relaxation modulus. In both P and N samples, the relaxation modulus was described well by the empirical equation. The KWW1 process of a P sample almost completely coincided with that of an N sample. In the slow process (KWW2 process), there was a difference between the relaxation modulus of a P sample and that of an N sample. The results indicate that the KWW1 process in the empirical equation represents the relaxation in the collagen matrix in bone and that the KWW2 process is related to a higher-order structure of bone that is responsible for the anisotropic mechanical properties of bone. PMID- 15275853 TI - Segment-interaction analysis of the stance limb in sprint running. AB - A high angular velocity of the thigh of the stance limb, generated by hip extensor musculature, is commonly thought to be a performance-determining factor in sprint running. However, the thigh segment is a component of a linked system (i.e., the lower limb), therefore, it is unlikely that the kinematics of the thigh will be due exclusively to the resultant joint moment (RJM) at the hip. The purpose of this study was to quantify, by means of segment-interaction analysis, the determinants of sagittal plane kinematics of the lower limb segments during the stance phase of sprint running. Video and ground reaction force data were collected from four male athletes performing maximal-effort sprints. The analysis revealed that during the first-third of the stance phase, a hip extension moment was the major determinant of the increasing angular velocity of the thigh. However, during the mid-third of stance, hip and knee extension moments and segment interaction effects all contributed to the thigh attaining its peak angular velocity. Extension moments at the ankle, and to a lesser extent the knee, were attributed with preventing the 'collapse' of the shank under the effects of the interactive moment due to ground reaction force. The angular acceleration of the foot was determined almost completely by the RJM at the ankle and the interactive moment due to ground reaction force. Further research is required to determine if similar results exit for a wide range of athletes and for other stages of a sprint race (e.g. early acceleration, maximal velocity, and deceleration phases). PMID- 15275854 TI - Moment arms and musculotendon lengths estimation for a three-dimensional lower limb model. AB - This paper presents a set of polynomial expressions that can be used as regression equations to estimate length and three-dimensional moment arms of 43 lower-limb musculotendon actuators. These equations allow one to find, at a low computational cost, the musculotendon geometric parameters required for numerical simulation of large musculoskeletal models. Nominal values for these biomechanical parameters were established using a public-domain musculoskeletal model of the lower limb (IEEE Trans. Biomed. Eng. 37 (1990) 757). To fit these nominal values, regression equations with different levels of complexity were generated, based on the number of generalized coordinates of the joints spanned by each musculotendon actuator. Least squares fitting was used to identify regression equation coefficients. The goodness of the fit and confidence intervals were assessed, and the best fitting equations selected. PMID- 15275855 TI - Transforming growth factor beta (TGFbeta) signalling in palatal growth, apoptosis and epithelial mesenchymal transformation (EMT). AB - Formation of the medial edge epithelial (MEE) seam by fusing the palatal shelves is a crucial step of palate development. The opposing shelves adhere to each other at first by adherens junctions, then by desmosomes in the MEE. The MEE seam disappears by epithelial mesenchymal transformation (EMT), which creates confluence of connective tissue across the palate. Cleft palate has a mutifactorial etiology that often includes failure of adherence of apposing individual palatal shelves and/or EMT of the MEE. In this review, we first discuss TGFbeta biology, including functions of TGFbeta isoforms, receptors, down stream transcription factors, endosomes, and signalling pathways. Different isoforms of the TGFbeta family play important roles in regulating various aspects of palate development. TGFbeta1 and TGFbeta2 are involved in growth, but it is TGFbeta3 that regulates MEE transformation to mesenchyme to bring about palatal confluence. Its absence results in cleft palate. Understanding of TGFbeta family signalling is thus essential for development of therapeutic strategies. Because TGFbeta3 and its downstream target, LEF1, play the major role in epithelial transformation, it is important to identify the signalling pathways they use for palatal EMT. Here, we will discuss in detail the mechanisms of palatal seam disappearance in response to TGFbeta3 signalling, including the roles, if any, of growth and apoptosis, as well as EMT in successful MEE adherence and seam formation. We also review recent evidence that TGFbeta3 uses Smad2 and 4 during palatal EMT, rather than beta-Catenin, to activate LEF1. TGFbeta1 has been reported to use non-Smad signalling using RhoA or MAPKinases in vitro, but these are not involved in activation of palatal EMT in situ. A major aim of this review is to document the genetic mechanisms that TGFbeta uses to bring about palatal EMT and to compare these with EMT mechanisms used elsewhere. PMID- 15275856 TI - Soluble guanylyl cyclase is localised in the acinar cells and participates in amylase secretion in rat parotid gland. AB - It is well known that the muscarinic cholinergic agonists, carbachol and methacholine, enhance nitric oxide synthase (NOS) activity, and also stimulate salivary secretion. In the present study, we investigated whether salivary secretion by muscarinic cholinergic stimulation is mediated through the NO/cGMP signaling pathway in rat salivary glands. Since NO activates soluble guanylyl cyclase (sGC) and cGMP may function as a mediator, the localisation of sGC was investigated in the salivary glands. sGC was localized in both the acinar and duct cells of the rat parotid and sublingual glands, and localized only in the acinar cells of the submandibular glands. S-Nitroso-glutathione (NO generator; GSNO) and YC-1 (NO-independent sGC activator) stimulated sGC in the cytosol to synthesise cGMP. The combination of GSNO and YC-1 stimulated sGC synergistically. Carbachol, GSNO and YC-1 enhanced amylase release from the rat parotid glands. Amylase release stimulated by carbachol and GSNO was inhibited by addition of the sGC inhibitor, ODQ, and cGMP-dependent protein kinase inhibitor, KT-5823. These results indicate that amylase release may be mediated through the NO/cGMP signaling pathway. PMID- 15275857 TI - Glycosylations in demilunar and central acinar cells of the submandibular salivary gland of ferret investigated by lectin histochemistry. AB - 'Resting' submandibular salivary glands obtained post-mortem from mature ferrets of both sexes were examined here. The binding patterns of labelled lectins applied to paraffin sections of tissue slivers fixed in an aldehyde-HgCl2 mixture and the effects of pretreatment procedures on the results were assessed lightmicroscopically. Lectins with affinity for terminal GalNAc residues (DBA, SBA) bound preferentially to demilunar acinar cells which were also strongly reactive with Fuc-directed UEA I. In contrast, lectins with affinity for neuraminic acid (SNA, WGA) bound to central acinar cells where consistent binding of DBA and SNA occurred only after neuraminidase digestion, and variation in the binding of UEA I was seen. The reactivities corresponded with the distribution of secretory granules, but staining in Golgi-like areas occurred in central acinar cells with PNA lectin. The results suggest that glycosylations are more advanced in central than demilunar acinar cells of the ferret submandibular gland. Possibly demilunar and central acinar cells reflect phenotypic changes of a single secretory cell, the 'central' acinar phenotype being influenced by incorporation of neuraminic acid in glycoprotein side chains and by increased Golgi activity. PMID- 15275858 TI - Environmental influences on the trace element content of teeth--implications for disease and nutritional status. AB - The aim of this study was to compare the trace element content of children's primary teeth from Uganda and the UK. The Ugandan teeth were from children living in an area where endomyocardial fibrosis (EMF), a cardiac disease, is prevalent. The latter has been putatively linked to insufficient magnesium intake and excess cerium exposure. Primary teeth were collected from 21 Ugandan and 27 UK children. The crowns and roots of the teeth were separated and the former digested and analysed for several major and trace elements by inductively coupled plasma mass spectrometry (ICP-MS) and atomic emission spectrometry (ICP-AES). In addition, the enamel and dentine of eight UK and seven Ugandan primary teeth were isolated via density separation and analysed as above. The data were assessed using non parametric statistical tests. The Ugandan teeth contained significantly (P < 0.05) greater concentrations of strontium, barium, cerium, lanthanum, praseodymium and significantly less zinc than the UK teeth. No significant difference in the concentrations of aluminium, calcium, copper, magnesium, lead and uranium were found. Analysis of enamel and dentine demonstrated that the former was enriched with several elements including cerium. It is concluded, that the environment, influences the trace element content of primary teeth and this may be useful for monitoring nutritional status. With respect to a geochemical cause for EMF, there is no positive evidence that EMF in Uganda is associated with reduced magnesium and increased cerium uptake in primary teeth. This does not, however, exclude cerium from playing a role in the aetiology of EMF. PMID- 15275859 TI - A cluster analysis model for caries risk assessment. AB - Cluster analysis was applied to determine, the natural grouping of individuals, among sixty 8-10-year-old children, and to identify the most significant set of markers for risk assessment. The risk clusters were obtained with initial clinical and bacteriological measurements including dmf + DMFS, active caries, mutans streptococci and lactobacilli counts in plaque or saliva on two media, and Snyder's test results. The morbidity clusters were constructed with the final clinical indexes and incidence after 18 months (dependent variables). A risk cluster was identified that included the following significant initial variables; dmf + DMFS, active caries, counts of mutans streptococci from plaque on TSY20B and lactobacilli in saliva, and Snyder's test results. This set of markers identified 86% of the children at high risk who developed high morbidity, as well as 94% of children in the low-risk cluster who developed low or no caries. The results of this investigation provide the basis to develop a system for caries risk assessment. PMID- 15275860 TI - Direct detection of cell surface interactive forces of sessile, fimbriated and non-fimbriated Actinomyces spp. using atomic force microscopy. AB - Actinomyces species are predominant early colonizers of the oral cavity and prime mediators of inter-bacterial adhesion and coaggregation. Previous workers have evaluated the adhesion of Actinomyces spp. by quantitative assessment of sessile, as opposed to planktonic cells attached to substrates, but did not quantify the cell surface interactive forces. Therefore we used atomic force microscopy to directly detect the interactive force between an approaching silicon tip and sessile Actinomyces spp. adhering to a substrate, at nanonewton (nN) range force levels. A total of eight strains each belonging to fimbriated and non-fimbriated Actinomyces species were employed, namely A. bovis, A. gerencseriae, A. israelii, A. meyeri, A. naeslundii genospecies 1 and 2, A. odontolyticus and A. viscosus. The sterile mica discs, used as the adhesion substrate, were immersed in mono species bacterial suspensions for five days to obtain a thin bacterial biofilm. Interactive forces were measured using a silicon nitride cantilever attached to a Nanoscope IIIA atomic force microscope. The interactive forces between the approaching silicon nitride tip and bacterial biofilm surfaces were randomly quantified at three different locations on each cell; namely, the cell surface proper, the periphery of the cell and the substrate and, the interface between two cells. When the interactive forces at these locations of the same species were compared, significantly higher force levels at the cell-cell interface than the other two locations were noted with A. gerencseriae (P < 0.001), A. viscosus (P < 0.01) and A. israelii (P < 0.05). When the interactive forces of different Actinomyces spp. at an identical location were compared, fimbriated A. naeslundii genospecies 2 showed the greatest interactive force at the cell surface proper ( 32.6 +/- 8.7 nN, P < 0.01). A. naeslundii genospecies 1, 2 and A. viscosus demonstrated greater interactive force at the cell-mica periphery than the other five species (P < 0.05); A. viscosus (-34.6 +/- 10.5 nN) displayed greater interactive force at the cell-cell interface than the others (P < 0.01), except for A. gerencseriae (P > 0.05). These data indicate that fimbriated Actinomyces spp., including A. naeslundii genospecies 1, 2 and A. viscosus exert higher cell surface interactive forces than those devoid of fimbriae and, such variable force levels may modulate their adhesion and coaggregation during biofilm formation. PMID- 15275861 TI - Immuno-localization of COX-1 and COX-2 in the rat molar periodontal tissue after topical application of lipopolysaccharide. AB - Up-regulation of prostaglandin E2 (PGE2) production in the periodontal tissue is considered to be important for periodontal tissue destruction. The purpose of the study was to demonstrate the dynamic changes of immuno-localization of cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) in rat periodontal tissue after topical application of lipopolysaccharide (LPS: 5 mg/ml in physiological saline) from Escherichia coli into the rat molar gingival sulcus. In the normal periodontal tissue, small numbers of junctional epithelium (JE) cells and numerous osteocytes embedded in alveolar bone constitutively expressed COX-1. The COX-1 expression was not effected by LPS application. JE cells, especially in the coronal portion of JE also expressed COX-2. LPS application induced the JE cells with consequent transient expression of COX-2 with a peak at day 1. These findings suggest that JE cells may play a critical role in first defense line against LPS challenge and PGE2 from JE cells may be responsible for the initiation of periodontal inflammation. In the deep periodontal tissue, cementoblasts and osteoblasts showed constitutive expression of COX-2, which may be induced by continuous cyclic tension force due to occlusal pressure. LPS application caused a transient up-regulation of COX-2 expression in periodontal ligament fibroblasts, cementoblasts and osteoblasts. It is suggested that the inducible production of PGE2 via COX-2 by these cells may be associated with connective tissue destruction and alveolar bone resorption. PMID- 15275862 TI - Subperiosteal hydroxyapatite implants in rats submitted to ethanol ingestion. AB - It has been reported that excessive alcohol consumption can contribute to failures on the osteointegration process in the site of implantation. Hydroxyapatite blocks were implanted under the periosteum of the femur and skull of 40 rats divided into two groups of 20 animals, one of them received 25% ethanol diluted in water and the other did not. Bone formation close to the hydroxyapatite implant was observed in the femur of all animals 2 weeks after surgery, however the bone volume was lower in ethanol-treated animals. It was observed in the skulls of the ethanol-treated animals a delay in new bone formation process, as a lower bone volume, too. After 4 weeks of the implantation, just one ethanol-treated animal showed no new bone formation in the femur, while no bone formation was observed in the skulls of two other rats. On the 8th and 16th weeks, bone formation was observed in both femur and skull from both groups, although always with less volume in ethanol-treated rats. We concluded that ethanol consumption did not impair osteointegration of ceramic implants, but it might have reduced the osteogenic capacity of periosteal cells in the femur and parietal bone of the rats. PMID- 15275863 TI - Bone healing in osteoporotic female rats following intra-alveolar grafting of bioactive glass. AB - We have investigated the effect of ovariectomy combined with a low Ca diet on bone healing following the implantation of bioactive glass into extraction sockets, in rats. Ovariectomized rats received a low Ca diet from the day of surgery until sacrifice while sham-operated animals were fed a standard laboratory chow. Two weeks after surgery the upper incisors were extracted and the alveolar sockets in both groups were partially filled with a particulate bioglass (PerioGlas). The animals were killed 1, 2, 3 and 9 weeks after tooth extraction and the relative volume fraction of the healing components (bone trabeculae, connective tissue and coagulum remnants) was estimated in histological paraffin sections by a histometric differential point-counting method. The bioglass particles persisted inside the socket for all the experimental periods and, as bone repair proceeded, they were progressively enclosed in newly formed bone trabeculae which in some cases established a close contact with their surface. The volume fraction of neoformed bone trabeculae relative to the volume fraction of connective tissue and coagulum remnants was greater in the sockets of ovariectomized animals implanted with bioglass than in those of the overiectomized non-implanted groups. PMID- 15275864 TI - Nitric oxide function in the skin. AB - Endogenously produced nitric oxide (NO) has a remarkably diverse range of biological functions, including a role in neurotransmission, smooth muscle relaxation, and the response to immunogens. Over the last 10 years, it has become clear that this extraordinary molecular messenger also plays a vital role in the skin, orchestrating normal regulatory processes and underlying some of the pathophysiological ones. We thought it pertinent to review the current literature concerning the possible function of NO in normal skin, its clinical and pathological significance, and the potential for therapeutic advances. The keratinocytes, which make up the bulk of the epidermis, constitutively express the neuronal isoform of NO synthase (NOS1), whereas the fibroblasts in the dermis and other cell types in the skin express the endothelial isoform (NOS3). Under certain conditions, virtually all skin cells appear to be capable of expressing the inducible NOS isoform (NOS2). The expression of NOS2 is also strongly implicated in psoriasis and other inflammatory skin conditions. Constitutive, low level NO production in the skin seems to play a role in the maintenance of barrier function and in determining blood flow rate in the microvasculature. Higher levels of NOS activity, stimulated by ultraviolet (UV) light or skin wounding, initiate other more complex reactions that require the orchestration of various cell types in a variety of spatially and temporally coordinated sets of responses. The NO liberated following UV irradiation plays a significant role in initiating melanogenesis, erythema, and immunosuppression. New evidence suggests that it may also be involved in protecting the keratinocytes against UV-induced apoptosis. The enhanced NOS activity in skin wounding (reviewed recently in this journal [Nitric oxide 7 (2002) 1]) appears to be important in guiding the infiltrating white blood cells and initiating the inflammation. In response to both insults, UV irradiation and skin wounding, the activation of constitutive NOS proceeds and overlaps with the expression of NOS2. Thus, at a macro-level, at least three different rates of NO production can occur in the skin, which seem to play an important part in organizing the skin's unique adaptability and function. PMID- 15275865 TI - Protection of lung tissue against ischemia/reperfusion injury by preconditioning with inhaled nitric oxide in an in situ pig model of normothermic pulmonary ischemia. AB - Topical administration of nitric oxide (NO) by inhalation is currently used as therapy in various pulmonary diseases, but preconditioning with NO to ameliorate lung ischemia/reperfusion (I/R) injury has not been fully evaluated. In this study, we investigated the effects of NO inhalation on functional pulmonary parameters using an in situ porcine model of normothermic pulmonary ischemia. After left lateral thoracotomy, left lung ischemia was maintained for 90 min, followed by a 5h reperfusion period (group I, n = 7). In group II (n = 6), I/R was preceded by inhalation of NO (10 min, 15 ppm). Animals in group III (n = 7) underwent sham surgery without NO inhalation or ischemia. In order to evaluate the effects of NO preconditioning, lung functional and hemodynamic parameters were measured, and the zymosan-stimulated release of reactive oxygen species in arterial blood was determined. Animals in group I developed significant pulmonary I/R injury, including pulmonary hypertension, a decreased pO(2) level in pulmonary venous blood of the ischemic lung, and a significant increase of the stimulated release of reactive oxygen species. All these effects were prevented, or the onset (release of reactive oxygen species) was delayed, by NO inhalation. These results indicate that preconditioning by NO inhalation before lung ischemia is protective against I/R injury in the porcine lung. PMID- 15275866 TI - An in situ evidence for autocrine function of NO in the vasculature. AB - The concept of endothelium derived relaxing factor (EDRF) implies that nitric oxide (NO) generated by NO synthase in the endothelium diffuses to the underlying vascular smooth muscle cells (VSMC) modulating thereby vascular tone. VSMC were regarded as passive recipients of NO from endothelial cells. However, this paradigm of a paracrine function of NO became currently subject to considerable debate. To address this issue, we examined the localization of enzymes engaged in l-arginine-NO-cGMP signaling in the rat blood vessels. Employing multiple immunocytochemical labeling complemented with signal amplification, electron microscopy, Western blotting, and RT-PCR, we found that NO synthase was differentially expressed in blood vessels depending on the blood vessel type. Moreover, the expression pattern of NO synthase in VSMC showed striking parallels with arginase and soluble guanylyl cyclase. Our findings challenge the commonly accepted view that the expression of NO synthase is restricted to vascular endothelial cells and lends further support to an alternative mechanism, by which constitutive local NOS expression in VSMC may modulate vascular functions in an endothelium-independent manner. Moreover, the co-expression of enzymes engaged in l-arginine-NO-cGMP signaling (NO synthase, arginase, and soluble guanylyl cyclase) in VSMC is indicative of an autocrine fashion of NO signaling in the vasculature in addition to the paracrine role of NO generated in the endothelium. PMID- 15275867 TI - Nitric oxide activates the expression of IRAK-M via the release of TNF-alpha in human monocytes. AB - The activation of interleukin receptor associated kinases (IRAK) is an important event in several inflammatory processes. However, exposing monocytes to a nitric oxide (NO) donor inhibits the activity of IRAK-1 and its molecular interaction with TNF receptor associated factor-6 (TRAF6). Despite the fact that NO is known to regulate many events in the immune and vascular system, the mechanism that underlies this inhibition remains unknown. We have recently demonstrated that IRAK-M inhibits the TLR/IRAK pathway during endotoxin tolerance and thus, we hypothesized that IRAK-M may be involved in the inhibition of IRAK-1 activity in the presence of NO. Hence, we have analyzed the expression of IRAK-M in human monocytes following exposure to a NO donor (GSNO) and we have observed that GSNO was capable of inducing IRAK-M mRNA and protein expression 8 and 20 h after stimulation, respectively. It is known that NO induces the expression of TNF alpha in monocytes and we found that exposure to TNF-alpha induced IRAK-M mRNA expression in human monocytes within 2 h of stimulation. Furthermore, the expression of IRAK-M induced by GSNO was inhibited by the presence of a blocking antibody raised against TNF-alpha. Thus, our data indicate that stimulation of human monocytes with a NO donor results in a clear induction of IRAK-M and this is dependent on the release of TNF-alpha by this kind of cells. PMID- 15275868 TI - Bound NO in human red blood cells: fact or artifact? AB - There has been considerable debate over the nature and chemistry of the interaction between nitric oxide (NO) and red blood cells (RBCs), in particular whether hemoglobin consumes or conserves NO bioactivity. Given the vast range of nitrosation levels reported for human RBCs in the literature, we sought to investigate whether there was a common denominator that could account for such discrepancies across different methodologies and reaction conditions and if such a pathway may exist in physiology. Here, we show that there are marked differences in reactivity toward NO between human and rat hemoglobin, which offers a mechanistic explanation for why basal levels of NO-adducts in primate RBCs are considerably lower than those in rodents. We further demonstrate that the inadvertent introduction of trace amounts of nitrite and incomplete thiol alkylation lead to rapid heme and thiol nitros(yl)ation, with generation of nitrosylhemoglobin (NOHb) and S-nitrosohemoglobin (SNOHb), while neither species is detectable in human RBCs at physiological nitrite concentrations. Thus, caution should be exercised in interpreting experimental results on SNOHb/NOHb levels that were obtained in the absence of knowledge about the degree of nitrite contamination, in particular when a physiological role for such species is implicated. PMID- 15275869 TI - Pulmonary 15NO uptake in interstitial lung disease. AB - Because lung nitric oxide (NO) diffusing capacity (DL) represents alveolar capillary gas diffusion, we queried as to whether disturbances of pulmonary gas exchange in interstitial lung disease (ILD) are appropriately reflected by using NO. In this pilot study, we applied the (15)N-labeled stable isotope (15)NO (relative abundance 0.37% of total NO) in order to ignore the endogenous NO production. In 10 ILD-outpatients, we measured DL (15)NO by performing the single breath method. Lung function parameters as well as arterial oxygen partial pressure (PaO(2)) were also tested. Values of DL (15)NO ranged within 50-151 ml (15)NO/(mmHg min). Ratios of DL (15)NO/reference were between 43 and 108% of predicted data as taken from our previous work on healthy volunteers [Eur. J. Physiol. 446 (2003) 256]. We found a significant reduction of DL (15)NO/reference in five patients. Additionally, values of PaO(2) were significantly correlated to ratios of DL (15)NO/reference (adjusted R2 +/-SEE=0.407+/-8.051). In conclusion, (15)NO represents an appropriate indicator gas for reflecting an ILD-induced impairment of alveolar-capillary gas exchange. PMID- 15275870 TI - Portosystemic shunts in children: a 15-year experience. AB - BACKGROUND: The role of portosystemic shunt (PSS) in children with portal hypertension has changed because of acceptance of liver transplantation and endoscopic hemostasis. We report our experience with PSS, mainly the distal splenorenal shunt, to define its role in the management of variceal bleeding. STUDY DESIGN: From 1987 to 2002, 20 children with variceal bleeding after endoscopic therapy underwent PSS. Patient and database records were reviewed. RESULTS: There were 14 boys and 6 girls; mean age was 11 years (range 3 to 18 years). Seventeen distal splenorenal and three mesocaval venous interposition shunts were performed. There was no operative mortality, 19 patients were alive at a median followup of 31 months (range 4 to 168 months) without evidence of recurrent gastrointestinal bleeding. One patient underwent transplantation 2 years after PSS and 1 patient died of hepatic failure while awaiting transplantation. The cause of portal hypertension was portal vein thrombosis (n = 13), biliary atresia (n = 3), congenital hepatic fibrosis (n = 2), hepatitis C cirrhosis (n = 1), and Budd-Chiari syndrome (n = 1). Eighteen children were Child Turcotte-Pugh class A and the remaining two were class B. One patient had two episodes of hematemesis after PSS. Two patients had worsening ascites. One patient had mild encephalopathy and one patient had shunt stenosis requiring angioplasty. CONCLUSIONS: PSS is a safe and durable therapy for pediatric patients with portal hypertension. Liver transplantation should be reserved for children with poor synthetic function associated with variceal bleeding. PSS may also serve as a bridge to transplantation in patients with preserved hepatic function. PSS, in particular the distal splenorenal shunt, has produced excellent results. This experience challenges the need for alternative forms of portal decompression. PMID- 15275871 TI - Delayed hemorrhage after pancreaticoduodenectomy. AB - BACKGROUND: Postoperative hemorrhage, particularly delayed hemorrhage after pancreaticoduodenectomy, is a serious complication and one of the most common causes of mortality after pancreaticoduodenectomy. STUDY DESIGN: The medical records of 500 patients who underwent pancreaticoduodenectomy between October 1994 and December 2002 were analyzed with regard to postoperative hemorrhagic complications. Delayed hemorrhage was defined as bleeding at the operation site after 5 or more postoperative days. RESULTS: Delayed hemorrhage occurred in 22 patients (4.4%), with a median time of 13 days (range 7 to 32 days) after pancreaticoduodenectomy, and developed more frequently (9/77 versus 13/423, p = 0.003) in patients with preceding intraabdominal complications such as pancreatic fistula, bile fistula, and intraabdominal abscess. In 17 of these 22 patients, angiography and laparotomy revealed bleeding foci at 14 arterial and 3 anastomotic sites. In nine patients, hemorrhage developed from pseudoaneurysms of the major arteries around the pancreaticojejunostomy. Hemostatis was attempted by transcatheter arterial embolization in 14 patients and with laparotomy in 4 patients. Four of 14 patients who received transcatheter arterial embolization eventually required laparotomy. Overall, 4 of the 22 delayed hemorrhage patients died (18.2%) of complications related to massive bleeding or transcatheter arterial embolization. CONCLUSIONS: Delayed hemorrhage after pancreaticoduodenectomy is associated with a high mortality. Intraabdominal complications after pancreaticoduodenectomy should be evaluated properly and guidelines for the diagnosis and treatment of delayed hemorrhage should be established in advance. Clinicians must be alert to the possibility of pseudoaneurysm hemorrhage. PMID- 15275872 TI - Management of failed biliary repairs for major bile duct injuries after laparoscopic cholecystectomy. AB - BACKGROUND: Many bile injuries are managed without referral to tertiary centers. Management of patients referred for a primary repair, or after a failed repair, was reviewed to compare outcomes. STUDY DESIGN: Retrospective review of data collected in prospective database. RESULTS: A total of 133 patients had been treated over 12 years ending in December 2002. Forty-six (35%) were treated for failed earlier repairs and 40 (30%) had their primary surgical repair at our institution. Patients with a failed repair were referred at a longer interval (165 versus 9 days, p < 0.001), were more often diagnosed intraoperatively (28 [61%] versus 13 [33%], p = 0.009), and presented with biliary obstruction (41 [89%] versus 13 [33%], p < 0.001). Of the failed repairs, 26 patients (56%) had an earlier biliary-enteric anastomosis and 20 had primary end-to-end repair. One third of failed repairs was successfully treated with stenting and was significantly more successful after a biliary-enteric anastomosis. Surgical revision of failed repairs was required in 27 patients (59%) and was more likely in earlier primary repairs. At a mean followup of 64 months, recurrent biliary strictures occurred in 5 patients (6%). CONCLUSIONS: Management of a failed major bile duct repair requires multiple modalities, but eventually the majority of repairs require surgical revision. Good results can be expected for all surgical biliary repairs at tertiary centers. PMID- 15275873 TI - Conservative management of pancreatic fistula after pancreaticoduodenectomy with pancreaticogastrostomy. AB - BACKGROUND: Pancreatic fistula (PF), which is a major complication of pancreaticoduodenectomy (PD), can be treated conservatively or by reoperation. The aim of this study was to evaluate conservative management of PF, which was attempted whenever possible as a first-intention treatment in a large series of PD. STUDY DESIGN: From 1990 to 2000, 242 patients underwent PD with pancreaticogastrostomy. PF was observed in 31 (13%) and was defined by an amylase rich surgical drainage fluid (above fivefold serum amylase) after postoperative day 5, or by presence on CT scan of a fluid collection located close to the anastomosis or containing amylase-rich fluid, or by operative findings in case of reoperation. Conservative management included total parenteral nutrition, nasogastric suction, imaging-guided percutaneous drainage of collection when necessary, and somatostatin or its analogues. RESULTS: PF was symptomatic in 20 patients (65%). Amylase level on surgical drainage fluid was elevated in 23 patients (74%). Four patients (13%), including two with hemorrhage and two with intraabdominal collection not accessible by percutaneous approach, were not considered for conservative management and underwent early reoperation. Conservative management was successful in the 27 patients (100%) in whom it was attempted, including the 10 who required percutaneous drainage. The only death (3%) occurred after massive hemorrhage complicating misdiagnosed PF. Mean hospital stay was 36 +/- 12 days (range 18 to 71) after successful conservative management. CONCLUSIONS: Conservative management of PF complicating PD is feasible and successful in above 85% of patients. PMID- 15275874 TI - Surgical procedures and histopathologic findings for patients with xanthogranulomatous cholecystitis. AB - BACKGROUND: Xanthogranulomatous cholecystitis (XGC) is an unusual and destructive inflammatory process of the gallbladder. Laparoscopic cholecystectomy (LC) may be contraindicated in XGC because of a high incidence of complications and coexistent malignancy. In this study, we examined the management of LC in patients with XGC. STUDY DESIGN: LC was attempted on 1,408 consecutive patients, including 27 (1.9%) patients with histopathologically diagnosed XGC. All patients underwent preoperative spiral computed tomography after IV infusion cholangiography and intraoperative cholangiography. We examined the correlation between the inflammatory grade of XGC and the difficulty of LC. RESULTS: LC was completed in 22 (81%) of the 27 patients diagnosed with XGC. Two patients with common bile duct injuries (partial lacerations) were confirmed by laparoscopic cholangiography, and injuries were simply closed using a laparoscopic technique. An intraoperative frozen-section examination revealed gallbladder carcinomas in two patients, and additional hepatectomies were performed in these patients after LC. Five patients (19%) with XGC required open operation. All of the laparoscopic failures were attributable to dense fibrotic adhesions in Calot's triangle and in the surrounding tissues. Histopathologically, nine patients had a xanthogranuloma with severe fibrotic reaction in the gallbladder wall, and four of these patients were treated by open operation. CONCLUSIONS: Although XGC has a relatively high conversion rate to open cholecystectomy, we conclude that patients with XGC should be considered for LC after an adequate patient selection, a clear visualization of anatomic structures and landmarks, and an intraoperative frozen section examination. PMID- 15275876 TI - Mortality prediction of head Abbreviated Injury Score and Glasgow Coma Scale: analysis of 7,764 head injuries. AB - BACKGROUND: We assessed the prognostic value and limitations of Glasgow Coma Scale (GCS) and head Abbreviated Injury Score (AIS) and correlated head AIS with GCS. STUDY DESIGN: We studied 7,764 patients with head injuries. Bivariate analysis was performed to examine the relationship of GCS, head AIS, age, gender, and mechanism of injury with mortality. Stepwise logistic regression analysis was used to identify the independent risk factors associated with mortality. RESULTS: The overall mortality in the group of head injury patients with no other major extracranial injuries and no hypotension on admission was 9.3%. Logistic regression analysis identified head AIS, GCS, age, and mechanism of injury as significant independent risk factors of death. The prognostic value of GCS and head AIS was significantly affected by the mechanism of injury and the age of the patient. Patients with similar GCS or head AIS but different mechanisms of injury or ages had significantly different outcomes. The adjusted odds ratio of death in penetrating trauma was 5.2 (3.9, 7.0), p < 0.0001, and in the age group > or = 55 years the adjusted odds ratio was 3.4 (2.6, 4.6), p < 0.0001. There was no correlation between head AIS and GCS (correlation coefficient -0.31). CONCLUSIONS: Mechanism of injury and age have a major effect in the predictive value of GCS and head AIS. There is no good correlation between GCS and head AIS. PMID- 15275875 TI - Is emergency department resuscitative thoracotomy futile care for the critically injured patient requiring prehospital cardiopulmonary resuscitation? AB - BACKGROUND: Documented prehospital asystole justifies termination of resuscitation, but recently it has been proposed to extend this policy to patients in the field with pulseless electrical activity. Consequently, we questioned whether resuscitative thoracotomy is warranted in the critically injured patient who fails to respond to prehospital CPR. STUDY DESIGN: A prospective database of all emergency department resuscitative thoracotomies (EDT) performed at our Level I trauma center has been maintained since January 1977. These registry data were augmented by a review of prehospital paramedic records for all survivors of EDT to verify length of CPR. RESULTS: During the 26 year study period, 959 patients underwent EDT. Of the 62 patients who survived to leave the hospital, 26 (42%) required prehospital CPR. The injury mechanism in these 26 patients was stab wounds in 18 (69%), gunshot wounds in 4 (15%), and blunt trauma in 4 (15%). The duration of prehospital CPR ranged from 3 to 15 minutes and in 7 patients CPR exceeded 10 minutes. Five survivors had asystole documented at the time of EDT; four of these patients had good functional outcomes at discharge. Each of these patients had pericardial tamponade from ventricular stab wounds. Patients with blunt trauma had uniformly dismal neurologic outcomes. CONCLUSIONS: EDT after prehospital CPR can be used to salvage select critically injured patients. Based on these data, we propose that resuscitative thoracotomy is futile care in patients with blunt trauma requiring prehospital CPR longer than 5 minutes, and in patients with penetrating trauma with more than 15 minutes of prehospital CPR. EDT is warranted in those patients with penetrating trauma with less than 15 minutes of prehospital CPR, and should be performed despite documented asystole on arrival if pericardial tamponade is the proximate event. PMID- 15275877 TI - Patterns of Internet use: bariatric versus colorectal patients in a private institution. AB - BACKGROUND: The frequency of Internet use for self-directed medical care in different patient populations is increasing. We evaluated Internet use by patients presenting for bariatric surgery. STUDY DESIGN: Surveys were completed by 136 patients (109 women, 22 men) presenting to a private academic clinic for bariatric surgery. Data collected included age, gender, education level, household income, and pattern of Internet use. Comparisons were made with a group of 135 patients who visited a colorectal surgery clinic in the same institution. RESULTS: Bariatric patients who used the Internet were more likely than colorectal patients to inform themselves about their medical problem (76% versus 49%, p < 0.001) and tended to use the Internet more overall (85% versus 78%, p = ns). Use of the Internet to research bariatric surgery was associated with education level (p = 0.002) and household income (p = 0.01), but not with age or gender. Bariatric patients were more likely than colorectal patients to search our institution's Web site (40% versus 17%, p < 0.001) and to use the Internet to find out about their surgeon (47% versus 31%, p = 0.01). Only 9% of bariatric patients used a chat room. Ninety-six percent of bariatric patients found the information on the Internet easy to understand and 58% described it as very helpful. CONCLUSIONS: Bariatric patients are especially likely to use the Internet to gain information about their medical condition, possibly reflecting their limited mobility. This represents an educational opportunity for the surgical community. PMID- 15275878 TI - Relationship between sestamibi uptake, parathyroid hormone assay, and nuclear morphology in primary hyperparathyroidism. AB - BACKGROUND: Technetium-99m-sestamibi scanning and the intraoperative intact parathyroid hormone (iPTH) assay have permitted development of focused parathyroid techniques for treatment of primary hyperparathyroidism. The purpose of this study was to assess if any pre- and intraoperative factors (degree of sestamibi uptake and iPTH levels) were associated with postoperative results (resected gland weight and parathyroid nuclear morphology). STUDY DESIGN: We performed a retrospective analysis of 101 consecutive patients who underwent preoperative sestamibi scintigraphy and a targeted parathyroid exploration for primary hyperparathyroidism. Sestamibi uptake was graded visually on a 4-point scale of 0 (no uptake or false-negative result) to 3 (high uptake) and compared with respect to iPTH levels, gland morphology, and specimen weight. A Kruskal Wallis test and a Pearson test were used for the statistical analysis. RESULTS: Degree of sestamibi uptake was associated with gland weight (median weight of 250 mg, 340 mg, 655 mg, 1,400 mg for grades 0, 1, 2, and 3, respectively, p < 0.001). The uptake of sestamibi was also associated with preoperative PTH levels (median PTH levels of 113, 151, 129, and 170 pg/mL for grades 0, 1, 2, and 3, respectively, p = 0.02), but not with the other parameters. The weight of resected gland(s) was associated with preoperative PTH levels (p = 0.02), and with some of the morphologic nuclear data (mean surface area (p = 0.02), maximum diameter (p = 0.01), and perimeter of the nuclei (p = 0.01). CONCLUSIONS: These data suggest that false-negative sestamibi scanning (visual score of 0) might result when hypersecreting parathyroid glands are small. Preoperative PTH level might be useful to estimate the amount of hypersecreting tissue to be resected. PMID- 15275879 TI - Critical involvement of stress-activated mitogen-activated protein kinases in the regulation of intracellular adhesion molecule-1 in serosal fibroblasts isolated from patients with Crohn's disease. AB - BACKGROUND: Stricture formation in Crohn's disease occurs as a result of persistent fibroblast activation. Chronic inflammation seen in patients with Crohn's disease leads to enhanced adhesion molecule expression in fibroblasts. Stress-activated mitogen-activated protein kinases are critical signaling pathways that control expression of intracellular adhesion molecule-1 (ICAM-1) in inflammation. The purpose of this study was to investigate the involvement of stress-activated mitogen-activated protein kinases in the regulation of ICAM-1 expression by tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL 1beta) in serosal fibroblasts isolated from patients with Crohn's disease. STUDY DESIGN: Fibroblasts were isolated from serosal biopsies of strictures in patients with Crohn's disease and normal colon in patients with colorectal carcinoma. Cell surface and whole cell ICAM-1 expression were evaluated by flow cytometry and Western blot analysis, respectively. Cells were stimulated with TNF-alpha and IL 1beta. To determine the mitogen-activated protein kinase signaling pathway required for ICAM-1 induction, cells were pretreated with inhibitors to Jun N terminal kinase, p38 kinase, and p42/44 kinase. RESULTS: Baseline ICAM-1 expression was higher (p < 0.001) in fibroblasts isolated from strictures in patients with Crohn's disease (3.2 +/- 0.3) as compared with nonstrictured Crohn's fibroblasts (2.1 +/- 0.3) and control fibroblasts (1.6 +/- 0.1). TNF alpha and IL-1beta increased ICAM-1 expression in both control and Crohn's disease. Pretreatment of fibroblasts with the Jun N-terminal kinase inhibitor dimethylaminopurine abolished TNF-alpha- and IL-1beta-stimulated ICAM-1 expression. CONCLUSIONS: Serosal fibroblasts isolated from strictures of patients with Crohn's disease demonstrate enhanced expression of ICAM-1. TNF-alpha and IL 1beta upregulate ICAM-1 expression in serosal fibroblasts through a Jun N terminal kinase signaling pathway. Specific inhibition of inflammatory signaling pathways could provide novel therapeutic targets for treatment of Crohn's disease. PMID- 15275880 TI - Minimally invasive video-assisted thyroidectomy: five years of experience. AB - BACKGROUND: In the last decade, development of videolaparoscopic surgery allowed several operations to be performed with minimally invasive techniques, making them less invasive and painful. Neck surgery was also involved in this effort, in spite of the skepticism shown by some authors. STUDY DESIGN: Minimally invasive video-assisted thyroidectomy was developed in 1998, and since then, about 600 operations have been performed. Access was the same as was previously described for parathyroidectomy; it was based on a small central incision (1.5 cm) and on external retraction without neck insufflation. RESULTS: From July 1998 to October 2003, 579 patients were selected from 5,450 for minimally invasive video-assisted thyroidectomy. The operation consisted of a total thyroidectomy in 312 patients and lobectomy in 267 patients. Mean operative time was 41 +/- 19.5 minutes (range 15 to 120 minutes) for lobectomy and 51.6 +/- 18.8 minutes (range 30 to 140 minutes) for total thyroidectomy. Postoperative hospital stay was 24 hours (overnight discharge) for all patients. Complications were postoperative bleeding (0.1%), recurrent nerve palsy (1.3%), and definitive hypoparathyroidism (0.2%). CONCLUSIONS: After 5 years of experience using this approach for various indications, we achieved a good esthetic result with an operative time comparable to that of conventional open surgery. Minimally invasive video-assisted thyroidectomy was found to be a safe operation, with advantages over traditional procedures represented by better cosmetic outcomes and postoperative course, as demonstrated by visual analogue scales and statistically analyzed numeric scales. PMID- 15275882 TI - Statement of support of motorcycle helmet laws. PMID- 15275881 TI - Inducible nuclear factor-kappaB activation contributes to chemotherapy resistance in gastric cancer. AB - BACKGROUND: Activation of nuclear factor-kappaB (NF-kappaB) inhibits chemotherapy induced apoptosis in some cancer cell lines. Inhibition of NF-kappaB by adenoviral delivery of an IkappaBalpha superrepressor (Ad.IkappaBalpha-SR) should potentiate 5-fluorouracil (5-FU) and irinotecan chemotherapy in gastric cancer cells. STUDY DESIGN: NCI-N87 and AGS human gastric cancer cells were studied. Chemotherapy-induced NF-kappaB activation was assessed using a luciferase reporter assay. Inhibition of NF-kappaB was assessed by luciferase reporter assay and by electrophoretic mobility shift assay. Cells were pretreated for 1 hour with Ad.IkappaBalpha (25 MOI) and incubated with 5-FU or the active metabolite of irinotecan (SN-38). Cell growth was assessed by cell proliferation assay and induction of apoptosis was determined by flow cytometry and caspase 3/7 assay. RESULTS: 5-FU and SN-38 significantly induced NF-kappaB activation as measured by luciferase reporter assay (p < 0.001). Ad.IkappaBalpha-SR treatment inhibited NF kappaB binding as demonstrated by electrophoretic mobility shift assay and by luciferase reporter assay. In AGS cells, pretreatment with Ad.IkappaBalpha-SR followed by 5-FU (0.005 mmol/L) or SN-38 (10 ng/mL) led to increased growth inhibition of 13% and 59%, respectively (p < 0.001). Similarly, growth inhibition in NCI cells was significantly increased by pretreatment with Ad.IkappaBalpha followed by 5-FU (0.001 mmol/L) or SN-38 (0.5 ng/mL) (p < 0.001). In both cell lines, Ad.IkappaBalpha-SR enhanced apoptosis by both flow cytometry and caspase 3/7 assay as compared with chemotherapy alone. CONCLUSIONS: NF-kappaB is activated in human gastric cancer in response to chemotherapy and may result in inducible chemoresistance. Inhibition of NF-kappaB by Ad.IkappaBalpha-SR enhances the antitumor effects of chemotherapy and has potential as a novel antineoplastic strategy. PMID- 15275883 TI - Motorcycle helmet laws: every surgeon's responsibility. PMID- 15275884 TI - What's new in general thoracic surgery. PMID- 15275885 TI - What's new in pediatric surgery. PMID- 15275886 TI - Vitalin: the rationale for a hypothetical hormone. PMID- 15275887 TI - Training surgeons to do evidence-based surgery: a collaborative approach. AB - BACKGROUND: Three of the Accreditation Council for Graduate Medical Education general competencies contain specific wording indicating that trainees must learn how to locate, appraise, and integrate the best information from the literature into their patient care practices. What is less clear is how to best translate evidence-based concepts into the workday of the resident, fellow, or attending surgeon. In this article we describe our use of the assignment-based training program we developed to ensure that our trainees can actually do what is required to practice evidence-based operations. STUDY DESIGN: Our collaborative program draws on the expertise of an attending surgeon, a medical librarian, and a research coordinator. The curriculum is designed so that all residents in our program develop and refine their evidence-based surgery skills in a context relating to their clinical practice. They are given a practice-related clinical question and are asked to demonstrate their competence in finding the best available evidence to answer it. This involves restating the question as a well formulated clinical question, doing a focused literature search, critically appraising the results to find the best evidence, and integrating the information into practice, if appropriate. Search assignments are evaluated using a structured form and additional training is designed based on the results. Another question is then assigned to assess improvement. RESULTS: Residents' performance on a first assignment showed specific weaknesses in use of textwords and limiters. Performance was strongly related to a resident's ability to obtain the best evidence in answer to a clinical question (p = 0.011). Substantial improvement was shown on a second assignment after additional training. CONCLUSIONS: Our hands-on, performance-based program allows us to document trainees' progress in developing skills that will allow them to efficiently locate the best evidence available to inform their patient care decisions. PMID- 15275888 TI - Incentive systems for academic productivity in a department of surgery. PMID- 15275889 TI - Parathyroid carcinoma in secondary and tertiary hyperparathyroidism. PMID- 15275891 TI - Palliation as a core surgical principle: part 2. PMID- 15275892 TI - Giant superior mesenteric artery aneurysm. PMID- 15275893 TI - Adrenocorticotropic hormone-secreting pancreatic islet cell carcinoma. PMID- 15275894 TI - Anatomic visualization with ultrasound-assisted intracranial image guidance in neurosurgery: a report of 30 patients. PMID- 15275895 TI - Temporary bowel diversion using the Bogota bag (Hadera stoma): technical details. PMID- 15275896 TI - Risk of stroke in endarterectomy group of SAPPHIRE trial. PMID- 15275897 TI - Placental umbilical cord whole blood transfusion. PMID- 15275900 TI - Detection of very high correlation in the alpha band between temporal regions of the human brain using MEG. AB - It is generally believed that alpha band (8-12 Hz) electric and magnetic activity in the area of the left and right temporal regions in the human brain are at best poorly correlated. There are no previous reports of very high alpha band correlation between left and right temporal regions by magnetoencephalography (MEG) or electroencephalography (EEG). We present whole head magnetoencephalography (MEG) results that demonstrate that, for temporal channels in the majority of healthy subjects tested, the alpha band signals are highly to very highly correlated and are antiparallel in direction. A correlation as high as -0.97 was found for a limited time in one subject. We suggest that the correlation found may be the consequence of strong direct or indirect coupling between homologue areas in left and right temporal regions rather than a common source. The correlation may provide a valuable index of loss of connectivity in the brain due to disease as well providing valuable insight to brain function and deserves further investigation. PMID- 15275901 TI - Temporal dynamics of alpha and beta rhythms in human SI and SII after galvanic median nerve stimulation. A MEG study. AB - In this MEG study, we investigated cortical alpha/sigma and beta ERD/ERS induced by median nerve stimulation to extend previous evidence on different resonant and oscillatory behavior of SI and SII (NeuroImage 13 [2001] 662). Here, we tested whether simple somatosensory stimulation could induce a distinctive sequence of alpha/sigma and beta ERD/ERS over SII compared to SI. We found that for both alpha/sigma (around 10 Hz) and beta (around 20 Hz) rhythms, the latencies of ERD and ERS were larger in bilateral SII than in contralateral SI. In addition, the peak amplitude of alpha/sigma and beta ERS was smaller in bilateral SII than in contralateral SI. These results indicate a delayed and prolonged activation of SII responses, reflecting a protracted information elaboration possibly related to SII higher order role in the processing of somatosensory information. This temporal dynamics of alpha/sigma and beta rhythms may be related to a sequential activation scheme of SI and SII during the somatosensory information processes. Future studies should evaluate in SII the possible different functional significance of alpha/sigma with respect to beta rhythms during somatosensory processing. PMID- 15275902 TI - Dissociating the spatio-temporal characteristics of cortical neuronal activity associated with human volitional swallowing in the healthy adult brain. AB - Human swallowing represents a complex highly coordinated sensorimotor function whose functional neuroanatomy remains incompletely understood. Specifically, previous studies have failed to delineate the temporo-spatial sequence of those cerebral loci active during the differing phases of swallowing. We therefore sought to define the temporal characteristics of cortical activity associated with human swallowing behaviour using a novel application of magnetoencephalography (MEG). In healthy volunteers (n = 8, aged 28-45), 151 channel whole cortex MEG was recorded during the conditions of oral water infusion, volitional wet swallowing (5 ml bolus), tongue thrust or rest. Each condition lasted for 5 s and was repeated 20 times. Synthetic aperture magnetometry (SAM) analysis was performed on each active epoch and compared to rest. Temporal sequencing of brain activations utilised time-frequency wavelet plots of regions selected using virtual electrodes. Following SAM analysis, water infusion preferentially activated the caudolateral sensorimotor cortex, whereas during volitional swallowing and tongue movement, the superior sensorimotor cortex was more strongly active. Time-frequency wavelet analysis indicated that sensory input from the tongue simultaneously activated caudolateral sensorimotor and primary gustatory cortex, which appeared to prime the superior sensory and motor cortical areas, involved in the volitional phase of swallowing. Our data support the existence of a temporal synchrony across the whole cortical swallowing network, with sensory input from the tongue being critical. Thus, the ability to non-invasively image this network, with intra-individual and high temporal resolution, provides new insights into the brain processing of human swallowing. PMID- 15275903 TI - Recognition memory for studied words is determined by cortical activation differences at encoding but not during retrieval. AB - Prior work has shown that when responses to incidentally encoded words are sorted, subsequently remembered words elicit greater left prefrontal BOLD signal change relative to forgotten words. Similarly, low-frequency words elicit greater activation than high-frequency words in the same left prefrontal regions, contributing to their better subsequent memorability. This study examined the relative contribution of encoding and retrieval processes to the correct recognition of target words. A mixture of high- and low-frequency words was incidentally encoded. Scanning was performed at encoding as well as during retrieval. During encoding, greater activation in the left prefrontal and anterior cingulate regions predicted a higher proportion of hits for low frequency words. However, data acquired during recognition showed that word frequency did not modulate activation in any of the areas tracking successful recognition. This result demonstrates that under some circumstances, the recognition of studied words is determined purely by processes that are active during encoding. In contrast to the finding for hits, activation associated with correctly rejected foils was modulated by word frequency, being higher for high frequency words in the left lateral parietal and anterior prefrontal regions. These findings were replicated in two further experiments, one in which the number of test items at recognition was doubled and another where encoding strength for high-frequency words was varied (once vs. 10 times). These results indicate that word frequency modulates activity in the left lateral parietal and anterior prefrontal regions contingent on whether the item involved is correctly recognized as a target or a foil. This observation is consistent with a dual process account of episodic memory. PMID- 15275904 TI - The functional anatomy of inspection time: an event-related fMRI study. AB - Twenty healthy young adults underwent functional magnetic resonance imaging (fMRI) of the brain while performing a visual inspection time task. Inspection time is a forced-choice, two-alternative visual backward-masking task in which the subject is briefly shown two parallel vertical lines of markedly different lengths and must decide which is longer. As stimulus duration decreases, performance declines to chance levels. Individual differences in inspection time correlate with higher cognitive functions. An event-related design was used. The hemodynamic (blood oxygenation level-dependent; BOLD) response was computed as both a function of the eight levels of stimulus duration, from 6 ms (where performance is almost at chance) to 150 ms (where performance is nearly perfect), and a function of the behavioral responses. Random effects analysis showed that the difficulty of the visual discrimination was related to bilateral activation in the inferior fronto-opercular cortex, superior/medial frontal gyrus, and anterior cingulate gyrus, and bilateral deactivation in the posterior cingulate gyrus and precuneus. Examination of the time courses of BOLD responses showed that activation was related specifically to the more difficult, briefer stimuli and that deactivation was found across most stimulus levels. Functional connectivity suggested the existence of two networks. One comprised the fronto opercular area, intrasylvian area, medial frontal gyrus, and the anterior cingulate cortex (ACC), possibly associated with processing of visually degraded percepts. A posterior network of sensory-related and associative regions might subserve processing of a visual discrimination task that has high processing demands and combines several fundamental cognitive domains. fMRI can thus reveal information about the neural correlates of mental events which occur over very short durations. PMID- 15275905 TI - Hand movement distribution in the motor cortex: the influence of a concurrent task and motor imagery. AB - The aim of this work was to study the relevance of the primary motor cortex (M1) for motor functions different to the simple execution of motor orders. The M1 activity during the performance with individual fingers of a simple motor task (tonic flexion), a motor task that includes a complex motor computation but not motor execution (motor imagery), and a motor task that involves both the computation and execution of movements (phasic movement) was evaluated by functional magnetic resonance imaging (fMRI). The possible influence of other cortical tasks on the M1 activation induced by finger movements was assessed by evaluating the effect of a distracting concurrent task (numeric calculation). Data show that both the dimension of the area activated and the intensity of response were higher during motor planning than during motor execution. There is a mosaic-like distribution for motor-planning M1 functions, with the movement of individual fingers being controlled from several M1 loci. The concurrent mental task induces a rapid functional reconfiguration of M1, adding M1 subsets to motor programming but excluding others. Present data support the involvement of the M1 in more than just simple motor execution, showing broader and more intense modifications during motor tasks not accompanied by movements (motor imagery) than during the execution of simple motor acts (tonic flexion). PMID- 15275906 TI - Semiautomatic brain region extraction: a method of parcellating brain regions from structural magnetic resonance images. AB - Structural MR imaging has become essential to the evaluation of regional brain changes in both healthy aging and disease-related processes. Several methods have been developed to measure structure size and regional brain volumes, but many of these methods involve substantial manual tracing and/or landmark identification. We present a new technique, semiautomatic brain region extraction (SABRE), for the rapid and reliable parcellation of cortical and subcortical brain regions. We combine the SABRE parcellation with tissue compartment segmentation [NeuroImage 17 (2002) 1087] to produce measures of gray matter (GM), white matter (WM), ventricular CSF, and sulcal CSF for 26 brain regions. Because SABRE restricts user input to a few easily identified landmarks, inter-rater reliability is high for all volumes, with all coefficients between 0.91 and 0.99. To assess construct validity, we contrasted SABRE-derived volumetric data from healthy young and older adults. Results from the SABRE parcellation and tissue segmentation showed significant differences in multiple brain regions in keeping with regional atrophy described in the literature by researchers using lengthy manual tracing methods. Our findings show that SABRE is a reliable semiautomatic method for assessing regional tissue volumes that provides significant timesavings over purely manual methods, yet maintains information about individual cortical landmarks. PMID- 15275907 TI - Age differences in neural correlates of route encoding and route recognition. AB - Spatial memory deficits are core features of aging-related changes in cognitive abilities. The neural correlates of these deficits are largely unknown. In the present study, we investigated the neural underpinnings of age-related differences in spatial memory by functional MRI using a navigational memory task with route encoding and route recognition conditions. We investigated 20 healthy young (18-29 years old) and 20 healthy old adults (53-78 years old) in a random effects analysis. Old subjects showed slightly poorer performance than young subjects. Compared to the control condition, route encoding and route recognition showed activation of the dorsal and ventral visual processing streams and the frontal eye fields in both groups of subjects. Compared to old adults, young subjects showed during route encoding stronger activations in the dorsal and the ventral visual processing stream (supramarginal gyrus and posterior fusiform/parahippocampal areas). In addition, young subjects showed weaker anterior parahippocampal activity during route recognition compared to the old group. In contrast, old compared to young subjects showed less suppressed activity in the left perisylvian region and the anterior cingulate cortex during route encoding. Our findings suggest that age-related navigational memory deficits might be caused by less effective route encoding based on reduced posterior fusiform/parahippocampal and parietal functionality combined with diminished inhibition of perisylvian and anterior cingulate cortices correlated with less effective suppression of task-irrelevant information. In contrast, age differences in neural correlates of route recognition seem to be rather subtle. Old subjects might show a diminished familiarity signal during route recognition in the anterior parahippocampal region. PMID- 15275908 TI - Preparatory deployment of attention to motion activates higher-order motion processing brain regions. AB - We used event-related fMRI to test the hypothesis that preparatory attention modulations occur in higher-order motion-processing regions when subjects deploy attention to internally driven representations in a complex motion-processing task. Using a cued attention-to-motion task, we found preparatory increases in fMRI activity in visual motion regions in the absence of visual motion stimulation. The cue, a brief enlargement of the fixation cross, directed subjects to prepare for a complex motion discrimination task. This preparation activated higher-order and lower-order motion regions. The motion regions activated included temporal regions consistent with V5/MT+, occipital regions consistent with V3+, parietal-occipital junction regions, ventral and dorsal intraparietal sulcus, superior temporal sulcus (STS), posterior insular cortex (PIC), and a region of BA 39/40 superior to V5/MT+ involving the angular gyrus and supramarginal gyrus (A-SM). Consistent with our hypothesis that these motion sensory activations are under top-down control, we also found activation of an extensive frontal network during the cue period, including anterior cingulate and multiple prefrontal regions. These results support the hypothesis that anticipatory deployment of attention to internally driven representations is achieved via top-down modulation of activity in task-relevant processing areas. PMID- 15275909 TI - Quantification of [18F]diprenorphine kinetics in the human brain with compartmental and non-compartmental modeling approaches. AB - 6-O-(2-[(18)F]fluoroethyl)-6-O-desmethyldiprenorphine ([(18)F]FDPN) is a nonselective opiate ligand that binds to postsynaptic micro, kappa and delta opiate receptors. Due to the longer half-life of F-18, compared to C-11, labeling DPN with F-18 allows for alternative experimental protocols and potentially the evaluation of endogenous opioid release. The applicability of this compound to assorted experimental protocols motivated the evaluation of [(18)F]FDPN kinetics with compartmental and non-compartmental models. The results indicate that a two tissue compartmental model best characterizes the data obtained following a bolus injection of [(18)F]FDPN (120-min scanning protocol). Estimates of distribution volume (DV) were robust, being highly correlated for the one-tissue compartmental model as well as the invasive Logan model and the basis function method. Furthermore, the DV estimates were also stable under a shortened protocol of 60 min, showing a significant correlation with the full protocol. The binding potential (BP) values showed more variability between methods and in some cases were more sensitive to protocol length. In conclusion, this evaluation of [(18)F]FDPN kinetics illustrates that DV values can be estimated robustly using compartmental modeling, the basis function method or the invasive Logan modeling approach on a volume of interest level. BP values were also found to correlate with DV values; however, these results should be interpreted with the understanding that specific binding in the reference region (occipital region) may exist. PMID- 15275910 TI - Ballistocardiogram artifact reduction in the simultaneous acquisition of auditory ERPS and fMRI. AB - Electroencephalography (EEG) and functional magnetic resonance imaging (fMRI) are now being combined to analyze brain function. Confounding the EEG signal acquired in the MR environment is a ballistocardiogram artifact (BA), which is predominantly caused by cardiac-related body movement. The objective of this study was to develop and evaluate a method for reducing these MR-induced artifacts to retrieve small auditory event-related potentials (ERPs) from EEG recorded during fMRI. An algorithm for BA reduction was developed that relies on timing information obtained from simultaneous electrocardiogram (ECG) recordings and subsequent creation of an adaptive BA template. The BA template is formed by median-filtering 10 consecutive BA events in the EEG signal. The continuously updated template is then subtracted from each BA in the EEG. The auditory ERPs are obtained through signal averaging of the remaining EEG signal. Experimental and simulated ERP data were estimated to assess effectiveness of the BA reduction. Simulation showed that the algorithm reduced BA without significantly altering the morphology of a signal periodically inserted in the EEG. Auditory ERP data, obtained in a 1.5-T scanner during a passive auditory oddball paradigm and processed with the BA reduction algorithm, were comparable to data recorded in a mock scanner outside the magnetic field with the same experimental paradigm. It is concluded that through adequate reduction of the BA, relatively small auditory ERPs can be acquired in the MR environment. PMID- 15275911 TI - Age-associated changes of cerebral glucose metabolic activity in both male and female deaf children: parametric analysis using objective volume of interest and voxel-based mapping. AB - Quantitative analysis of brain activity in the brains of children requires the establishment of age-associated norms. We investigated regional differences in age-associated changes in fluorodeoxyglucose (FDG) uptake in the developmental brains. From 87 (44 male and 43 female) deaf children from the age of 1 to 15, brain FDG positron emission tomography (PET) images were examined after spatial normalization, smoothing, and global normalization to identify brain regions showing a correlation between FDG uptake and age. Using population-based probabilistic volume of interests (VOIs), an objective VOI analysis was performed where normalized relative FDG uptake was measured and their correlations with age were examined in both genders. For the voxel-based analyses, the correlations with age were examined in a general linear model using statistical parametric mapping (SPM99). Both methods revealed that FDG uptake linearly increases with age both in the bilateral inferior prefrontal/orbitofrontal gyri and the right dorsomedial frontal gyrus and decreases in the inferior temporal gyrus and internal capsule white matter. Male children showed age-associated increases of FDG uptake in the right dorsomedial frontal gyrus, and female children in the left dorsolateral prefrontal cortex and thalamus. These changes in FDG uptake in various brain regions may suggest changes in synaptic density or regional activity resulting from normal maturation or deaf-induced adaptation. Caution should be exercised in interpreting the differences in the brain of child patients when compared with adult control's or with a different gender. Further research will be needed to examine if gender difference is manifested in the development rate of behavioral/cognitive functions in association with the age associated changes of the right medial frontal (male) or the left dorsolateral prefrontal cortices. PMID- 15275912 TI - An electrophysiological dissociation of retrieval mode and retrieval orientation. AB - The relationship between two classes of retrieval process-retrieval orientation and retrieval mode-was investigated by recording event-related potentials (ERPs) during the period in which participants were preparing to retrieve information from memory. Participants were cued on a trial-by-trial basis to complete either a semantic retrieval task or one of two episodic retrieval tasks (remember spatial location or remember encoding task). ERPs were recorded time-locked to the cues indicating which task to complete. There were commonalities between the ways in which ERPs evoked by the two episodic retrieval cues diverged from those evoked by the semantic cue. These shared differences are a likely correlate of processes related to retrieval mode. Critically, there were also reliable differences between the ERPs evoked by the two episodic cues. This novel finding is consistent with the view that participants adopted task-specific retrieval sets-retrieval orientations-which varied according to the kinds of episodic information that were to be retrieved. PMID- 15275913 TI - Anatomy and time course of discrimination and categorization processes in vision: an fMRI study. AB - Studies investigating the cerebral areas involved in visual processes generally oppose either different tasks or different stimulus types. This work addresses, by fMRI, the interaction between the type of task (discrimination vs. categorization) and the type of stimulus (Latin letters, well-known geometrical figures, and Korean letters). Behavioral data revealed that the two tasks did not differ in term of percentage of errors or correct responses, but a delay of 185 ms was observed for the categorization task in comparison with the discrimination task. All conditions activated a common neural network that includes both striate and extrastriate areas, especially the fusiform gyri, the precunei, the insulae, and the dorsolateral frontal cortex. In addition, interaction analysis revealed that the right insula was sensitive to both tasks and stimuli, and that stimulus type induced several significant signal variations for the categorization task in right frontal cortex, the right middle occipital gyrus, the right cuneus, and the left and right fusiform gyri, whereas for the discrimination task, significant signal variations were observed in the right occipito-parietal junction only. Finally, analyzing the latency of the BOLD signal also revealed a differential neural dynamics according to tasks but not to stimulus type. These temporal differences suggest a parallel hemisphere processing in the discrimination task vs. a cooperative interhemisphere processing in the categorization task that may reflect the observed differences in reaction time. PMID- 15275914 TI - Dissociable concurrent activity of lateral and medial frontal lobe during negative feedback processing. AB - External feedback on results of one's behavior guides flexible adaptation to changing environments. It has been suggested that the lateral and medial parts of the frontal lobe are responsible for cognitive and emotional functions, respectively. In the present fMRI study, multiple mental components evoked by the presentation of negative feedback were dissociated along the cognitive-emotional axis in set-shifting paradigms. The double dissociation of the concurrent feedback-related activity was observed in the right frontal lobe: the lateral frontal lobe was associated with the inferential component, whereas the medial frontal lobe was associated with the emotional component. However, among the multiple right lateral frontal regions, a region of interest (ROI) analysis indicated that the inferential component was not dominant in the region near the inferior frontal junction. The medial frontal activations were observed ventral and anterior to the presupplementary motor area, and dorsal and posterior to the anterior cingulate cortex. The double dissociation in the right frontal lobe suggests that the lateral and medial frontal lobe cooperatively but differentially contributes to the negative feedback processing, demonstrating the lateral-medial dichotomy of the frontal lobe functions suggested by previous neuropsychological studies. At the same time, the functional heterogeneity in the lateral and medial frontal lobe demands modifications of the traditional view of the functional organization of the frontal lobe. PMID- 15275915 TI - Parametric design and correlational analyses help integrating fMRI and electrophysiological data during face processing. AB - Face perception is typically associated with activation in the inferior occipital, superior temporal (STG), and fusiform gyri (FG) and with an occipitotemporal electrophysiological component peaking around 170 ms on the scalp, the N170. However, the relationship between the N170 and the multiple face sensitive activations observed in neuroimaging is unclear. It has been recently shown that the amplitude of the N170 component monotonically decreases as gaussian noise is added to a picture of a face [Jemel et al., 2003]. To help clarify the sources of the N170 without a priori assumptions regarding their number and locations, ERPs and fMRI were recorded in five subjects in the same experiment, in separate sessions. We used a parametric paradigm in which the amplitude of the N170 was modulated by varying the level of noise in a picture, and identified regions where the percent signal change in fMRI correlated with the ERP data. N170 signals were observed for pictures of both cars and faces but were stronger for faces. A monotonic decrease with added noise was observed for the N170 at right hemisphere sites but was less clear on the left and occipital central sites. Correlations between fMRI signal and N170 amplitudes for faces were highly significant (P < 0.001) in bilateral fusiform gyrus and superior temporal gyrus. For cars, the strongest correlations were observed in the parahippocampal region and in the STG (P < 0.005). Besides contributing to clarify the spatiotemporal course of face processing, this study illustrates how ERP information may be used synergistically in fMRI analyses. Parametric designs may be developed further to provide some timing information on fMRI activity and help identify the generators of ERP signals. PMID- 15275916 TI - Distributed self in episodic memory: neural correlates of successful retrieval of self-encoded positive and negative personality traits. AB - Words processed with reference to the self are generally better remembered than words processed in semantic terms. An account of this phenomenon, labeled the Self Reference Effect (SRE), is that the self promotes elaboration and organization of encoded information. Although a few neuroimaging studies associated self-referential encoding with activations of the medial prefrontal cortex, no previous study has investigated the neural correlates of remembering emotional words encoded in an SRE paradigm. The main goal of this study was to define with fMRI the neural correlates of the successful retrieval of negative and positive personality traits encoded in a self-referential mode. Functional MRI scans were acquired for 11 subjects as they recognized positive and negative emotional personality traits adjectives encoded in a self-referential condition, a semantic condition and in a phonemic condition. The correct recognition of self encoded personality traits engaged dorso-medial prefrontal cortex and lateral prefrontal regions, premotor cortex, parietal and occipital cortex, caudate and cerebellum. The specific recognition of self-encoded negative personality traits involved greater neural activation in the right extra-striate region than the recognition of positive personality traits. Our fMRI findings suggest that specific processes may operate at both encoding and retrieval to subserve the SRE. Unlike self-encoding, the retrieval of personality traits is modulated by the valence of the stimuli with greater activation for negative words. Our results indicate that personally relevant words may signal important emotional clues and support the notion of a widely distributed set of brain regions involved in maintaining the concepts of self. PMID- 15275917 TI - Dissociating linguistic and nonlinguistic gestural communication in the brain. AB - Gestures of the face, arms, and hands are components of signed languages used by Deaf people. Signaling codes, such as the racecourse betting code known as Tic Tac, are also made up of such gestures. Tic Tac lacks the phonological structure of British Sign Language (BSL) but is similar in terms of its visual and articulatory components. Using fMRI, we compared the neural correlates of viewing a gestural language (BSL) and a manual-brachial code (Tic Tac) relative to a low level baseline task. We compared three groups: Deaf native signers, hearing native signers, and hearing nonsigners. None of the participants had any knowledge of Tic Tac. All three groups activated an extensive frontal-posterior network in response to both types of stimuli. Superior temporal cortex, including the planum temporale, was activated bilaterally in response to both types of gesture in all groups, irrespective of hearing status. The engagement of these traditionally auditory processing regions was greater in Deaf than hearing participants. These data suggest that the planum temporale may be responsive to visual movement in both deaf and hearing people, yet when hearing is absent early in development, the visual processing role of this region is enhanced. Greater activation for BSL than Tic Tac was observed in signers, but not in nonsigners, in the left posterior superior temporal sulcus and gyrus, extending into the supramarginal gyrus. This suggests that the left posterior perisylvian cortex is of fundamental importance to language processing, regardless of the modality in which it is conveyed. PMID- 15275918 TI - Optimizing the experimental design for ankle dorsiflexion fMRI. AB - Compared to motor studies of the upper limb, few experiments have sought a relationship between blood oxygen level-dependent (BOLD) functional magnetic resonance imaging (fMRI) sensorimotor signals and the resulting lower limb output. In Experiment 1, using an fMRI simulator system, we determined the optimized experimental protocol based on two design types and four behavioral movement types during ankle dorsiflexion. Experiment 2 involved testing the BOLD sensitivity at 1.5 T during ankle movements. Subjects performed large- and small amplitude dorsiflexion movement types using an event-related design, with the intent of contrasting spatial and temporal features of the BOLD signal. In both experiments, the subject's behavior was guided by visual biofeedback of their ankle flexion angle, using an MR-compatible fiberoptic tape. From Experiment 1, we found electromyography (EMG) difference voltage ratio of approximately 2:1 for large (40 degrees ) and small (15 degrees ) dorsiflexion, 0.13 mV and 0.07 mV, respectively. In Experimental 2, we found the peak BOLD % signal changes of 1.04% and 0.89%, for large (40 degrees ) and small (15 degrees ) dorsiflexion, respectively. In addition, graded dorsiflexion produced graded BOLD signals in the primary sensorimotor and supplementary motor areas in 10 of 12 healthy young subjects, attesting to the feasibility of lower-limb fMRI at 1.5 T. This study provides insight into the cortical network involved in dorsiflexion using an experimental paradigm that is likely to translate effectively to hemiparetic stroke subjects. PMID- 15275919 TI - Social and emotional attachment in the neural representation of faces. AB - To dissociate the role of visual familiarity from the role of social and emotional factors in recognizing familiar individuals, we measured neural activity using functional magnetic resonance imaging (fMRI) while subjects viewed (1) faces of personally familiar individuals (i.e. friends and family), (2) faces of famous individuals, and (3) faces of strangers. Personally familiar faces evoked a stronger response than did famous familiar faces and unfamiliar faces in areas that have been associated with 'theory of mind', and a weaker response in the amygdala. These response modulations may reflect the spontaneous activation of social knowledge about the personality and attitudes of close friends and relatives and the less guarded attitude one has around these people. These results suggest that familiarity causes changes in neural response that extend beyond a visual memory for a face. PMID- 15275920 TI - Sex-specific, postpuberty changes in mouse brain structures revealed by three dimensional magnetic resonance microscopy. AB - Sexual dimorphism of brain structures has been reported in some species. We report that sex-dependent developmental structure changes exist in the C57Bl/6(J) mouse, a common model for the genetic analysis of brain function. High resolution, three-dimensional (3D) magnetic resonance microscopy (MRM) images were obtained in intact brains of male and female adult and peripubertal mice. The lateral and third ventricles, hippocampus, amygdala, striatum, and total brain were reconstructed in 3D. As observed in humans, there was overall cerebral growth from peripuberty to adulthood in both sexes. After correcting for the increased brain size, the hippocampus and amygdala were disproportionately larger in adult compared to peripubertal mice. Several sexual dimorphisms were also observed. The lateral ventricles were larger, while the amygdala (the left side in particular) was smaller in females compared to males. Lateral and third ventricles were reduced over time in males only, exhibiting a sex-specific developmental profile. The striatal size was uniform among the groups studied. The surface area of the segmented structures was assayed. Possible shape distortions were detected for the lateral ventricles, hippocampus, and overall brain structure based on a lack of covariance between the surface area and volumetric measurements. Although many sexually dimorphic changes are reported perinatally, our results suggest that there are additional sex-specific transformations that occur around puberty and persist in adulthood. PMID- 15275921 TI - Enabling the sharing of neuroimaging data through well-defined intermediate levels of visibility. AB - The sharing of neuroimagery data offers great benefits to science, however, data owners sharing their data face substantial custodial responsibilities, such as ensuring data sets are correctly interpreted in their new shared context, protecting the identity and privacy of human research participants, and safeguarding the understood order of use. Given choices of sharing widely or not at all, the result will often be no sharing, due to the inability of data owners to control their exposure to the risks associated with data sharing. In this context, data sharing is enabled by providing data owners with well-defined intermediate levels of data visibility, progressing incrementally toward public visibility. In this paper, we define a novel and general data sharing model, Structured Sharing Communities (SSC), meeting this requirement. Arbitrary visibility levels representing collaborative agreements, consortium memberships, research organizations, and other affiliations are structured into a policy space through explicit paths of permissible information flow. Operations enable users and applications to manage the visibility of data and enforce access permissions and restrictions. We show how a policy space can be implemented in realistic neuroinformatic architectures with acceptable assurance of correctness, and briefly describe an open source implementation effort. PMID- 15275922 TI - Local landmark-based mapping of human auditory cortex. AB - Mammalian sensory cortex is functionally partitioned into cortical fields that are specialized for different processing operations. In theory, averaging functional and anatomical images across subjects can reveal both the average anatomy and the mean functional organization of sensory regions. However, this averaging process must overcome at least two obstacles: (1) the relative locations and sizes of cortical sensory areas vary in different subjects so that across-subject averaging introduces spatial smearing; (2) the relative locations and sizes of cortical areas vary between hemispheres, making it difficult to compare activations between hemispheres or to combine activations across hemispheres. These difficulties are particularly acute for small cortical regions such as auditory cortex. In whole-brain averaging procedures, considerable intersubject variance in the location and orientation of auditory cortex is introduced by variance of the size and shape of structures outside auditory cortex. Here, we compared these global methods with local landmark-based methods (LLMs) that use warping based on local anatomical landmarks. In comparison to maps made with global methods, LLMs produced anatomical maps of auditory cortex with clearer gyral and sulcal structure, and produce functional maps with improved resolution. These results suggest that LLMs have significant advantages over global mapping procedures in studying the details of auditory cortex organization. PMID- 15275923 TI - A method for the analysis of the geometrical relationship between white matter pathology and the vascular architecture of the brain. AB - A novel method for the visual and quantitative analysis of the geometrical relationship between the vascular architecture of the brain and white matter pathology is presented. The cerebro vascular system is implicated in the pathogenesis of many diseases of the cerebral white matter, for example, stroke, microcerebrovascular disease, and multiple sclerosis (MS). In our work, white matter lesions and vessels are depicted using magnetic resonance imaging (MRI) and extracted using image analysis techniques. We focus on measuring distance relationships between white matter lesions and vessels, and distribution of lesions with respect to vessel caliber. Vascular distance maps are generated by computing for each voxel the Euclidean distance to the closest vessel. Analogously, radius maps assign the radius of the closest vessel to each voxel in the image volume. The distance and radius maps are used to analyze the distribution of lesions with respect to the vessels' locations and their calibers. The method was applied to three MS patients to demonstrate its functionality and feasibility. Preliminary findings indicate that larger MS lesions tend to be farther from detected vessels and that the caliber of the vessels nearest to larger lesions tends to be smaller, suggesting a possible role of relative hypoperfusion or hypoxia in lesion formation. PMID- 15275924 TI - Neuroimaging studies of shifting attention: a meta-analysis. AB - This paper reports a meta-analysis of neuroimaging studies of attention shifting and executive processes in working memory. We analyzed peak activation coordinates from 31 fMRI and PET studies of five types of shifting using kernel based methods [NeuroImage 19 (2003) 513]. Analyses collapsing across different types of shifting gave more consistent results overall than analysis within individual types, suggesting a commonality across types of shifting. These areas shared substantial, significant overlap with regions derived from kernel-based analyses of reported peaks for executive processes in working memory (WM). The results suggest that there is a common set of brain regions active in diverse executive control operations, including medial prefrontal, superior and inferior parietal, medial parietal, and premotor cortices. However, within several of these regions, different types of switching produced spatially discriminable activation foci. Precise locations of meta analysis-derived regions from both attention shifting and working memory are defined electronically and may be used as regions of interest in future studies. PMID- 15275925 TI - The neural correlates of theory of mind within interpersonal interactions. AB - Tasks that engage a theory of mind seem to activate a consistent set of brain areas. In this study, we sought to determine whether two different interactive tasks, both of which involve receiving consequential feedback from social partners that can be used to infer intent, similarly engaged the putative theory of mind neural network. Participants were scanned using fMRI as they played the Ultimatum Game (UG) and the Prisoner's Dilemma Game (PDG) with both alleged human and computer partners who were outside the scanner. We observed a remarkable degree of overlap in brain areas that activated to partner decisions in the two games, including commonly observed theory of mind areas, as well as several brain areas that have not been reported previously and may relate to immersion of participants in real social interactions that have personally meaningful consequences. Although computer partners elicited activation in some of the same areas activated by human partners, most of these activations were stronger for human partners. PMID- 15275926 TI - The hippocampal region is involved in successful recognition of both remote and recent famous faces. AB - There is currently a debate regarding the precise role of medial temporal regions in memory, in particular regarding the time scale of their involvement in conscious recollection of information stored in long-term memory. Using event related fMRI, we have attempted to contribute to this debate by identifying brain regions associated with the successful recognition of famous faces from two different periods: "Old" faces of people who became famous in the 1960s-1970s and "Recent" faces of people who became famous in the 1990s. We demonstrate that the hippocampus is involved in the successful recognition of famous faces from both periods and does not appear to distinguish between these two periods. We also highlight a network of brain regions, including the left prefrontal cortex, the retrosplenial cortex, the temporo-parietal junction, the caudate and the right cerebellum, which is activated in association with successful recognition of famous faces. Finally, an analysis of the results obtained during a post hoc episodic recognition task shows the specific involvement of anterior hippocampus in the successful encoding of the unfamiliar faces, which were presented during the fame decision task, suggesting a functional distinction between anterior and posterior parts of the hippocampus, the former being specifically involved in successful episodic encoding and the latter being associated with successful retrieval of semantic information. PMID- 15275927 TI - Sex and age dependencies of cerebral blood volume changes during cognitive activation: a multichannel near-infrared spectroscopy study. AB - In this study, we measured the change in cerebral hemoglobin concentrations during a cognitive task using multichannel near-infrared spectroscopy (NIRS), and investigated the relationship between regional cerebral blood volume and sex, age, and task performance. Thirty-nine healthy volunteers (24 males and 15 females; mean age, 33.0 years) participated after giving their informed consent and performed a word fluency task. The relative oxy-hemoglobin concentration ([oxy-Hb]) was measured using frontal and temporal probes with two sets of 24 channel NIRS machines. The effects of sex, age, and task performance on [oxy-Hb] changes were analyzed using analysis of covariance: with sex, age, and task performance as independent variables, and [oxy-Hb] changes as dependent variables, and years of education as covariates. The effects on [oxy-Hb] increase were significant in many channels in the frontal and temporal probes for sex, that is the most prominent effect, and in a few frontal channels for age: [oxy Hb] increases were larger in males than in females, and in the young than in the middle-aged. The effects on [oxy-Hb] increase were not significant for task performance, but [oxy-Hb] increases in subjects with low performance tended to be larger than those in subjects with high performance. The results demonstrated that multichannel NIRS could detect cerebral activation during cognitive tasks and clarify sex- and age-dependent differences in such cerebral activation. Sex- and age-dependent differences in cerebral activation, as demonstrated in the present study, should be considered when interpreting cerebral blood volume, cerebral blood flow, and cerebral glucose metabolism data. PMID- 15275928 TI - Brain activation patterns during imagined stance and locomotion in functional magnetic resonance imaging. AB - Posture and gait are sensorimotor actions that involve peripheral, spinal, and supraspinal structures. To investigate brain activity during stance and locomotion, 13 healthy subjects were asked to stand, walk, run, and lie down; subsequently, they were trained to imagine standing, walking, running, and lying [imagined lying as rest condition in functional magnetic resonance imaging (fMRI)]. Separate and distinct activation/deactivation patterns were found for the three imagined conditions: (1) standing imagery was associated with activation in the thalamus, basal ganglia, and cerebellar vermis; (2) walking imagery was associated with activation in the parahippocampal and fusiform gyri (areas involved in visuospatial navigation), occipital visual areas, and in the cerebellum; (3) running imagery caused a predominantly cerebellar activation in the vermis and adjacent hemispheres (six times larger than during imagination of walking or standing), but activations in the parahippocampal and fusiform gyri were smaller than during walking. Deactivations were found for walking and running, but not for standing imagery. They were located in the vestibular (posterior insula, superior temporal gyrus, supramarginal gyrus) and somatosensory (postcentral gyrus) cortex with right-hemispheric dominance. These findings support the concept of a hierarchical organization of posture and locomotion. Automated locomotion, for example, running, is based on spinal generators whose pace is driven by the cerebellar locomotor region. Deactivation in the vestibular and somatosensory cortex prevents adverse interactions with the optimized spinal pattern and sensory signals; this confirms earlier findings of a multisensory inhibition during unhindered locomotion. During slow walking, spatial navigation, mediated by the parahippocampal cortex, becomes more important. Postural control during standing involves a low intensity cerebellar activity and sensorimotor control via the thalamus and basal ganglia. PMID- 15275929 TI - Correction for intracranial volume in analysis of whole brain atrophy in multiple sclerosis: the proportion vs. residual method. AB - Two techniques that correct (normalize) regional and whole brain volumes according to head size-the proportion method (tissue-to-intracranial volume ratio) and the residual method (regression-based predicted brain tissue volumes) are used pervasively in neuroimaging research, but have received little critical evaluation or direct comparison. Using a quantitatively derived MRI data set of patients with multiple sclerosis (n = 18) and age-/sex-matched normal controls (n = 18), we introduced various types of error into estimates of intracranial volume (ICV) and absolute parenchymal volume (APV) to observe how this error affected the final outcome of normalized brain measures and their ability to detect group differences, as computed by a proportion (brain parenchymal fraction [BPF]) and residual method (predicted parenchymal volume [PPV]). The results indicated that systemic error in ICV and APV values considerably affected BPF means based on the proportion method, except with dependent-related systematic APV error, but essentially did not change statistical power associated with group differences in BPF. Random error altered BPF means to a much smaller extent, but was associated with moderate reductions in statistical power. On the other hand, PPV estimates based on the residual method were unaffected by these same ICV and APV errors, except with dependent-related systematic APV error, and were not associated with reductions in statistical power. Our findings suggest that head size correction of brain regions with the residual method generally may provide advantages over the proportion method. PMID- 15275930 TI - Haemodynamic responses to sensory stimulation are enhanced following acute cocaine administration. AB - Cocaine enhances neural activity in response to sensory stimulation, an effect that may play a role in the development of drug craving. However, cocaine-induced sensory enhancement may be difficult to study in humans using neuroimaging if the global increases in baseline haemodynamic parameters, which cocaine produces, interfere with the ability of enhanced sensory-related neural activity to lead to enhanced haemodynamic responses. To investigate the effect of cocaine-induced baseline haemodynamic changes on sensory-related haemodynamic (and electrophysiological) responses, field potential (FP) and haemodynamic responses (obtained using optical imaging spectroscopy and laser-Doppler flowmetry) in the barrel cortex of the anaesthetised rat were measured during mechanical whisker stimulation following cocaine (0.5 mg/kg) or saline administration. During cocaine infusion, the relationship between blood flow and volume transiently decoupled. Following this, cocaine caused large baseline increases in blood flow (133%) and volume (33%), which peaked after approximately 6 min and approached normal levels again after 25 min. During the peak baseline increases, FP responses to whisker stimulation were similar to saline whereas several haemodynamic response parameters were slightly reduced. After the peak, significant increases in FP responses were observed, accompanied by significantly enhanced haemodynamic responses, even though the haemodynamic baselines remained elevated. Hence, the haemodynamic response to sensory stimulation is transiently reduced in the presence of large increases in baseline but, after the baseline peak, enhanced neural responses are faithfully accompanied by enhanced haemodynamic responses. The findings suggest that any cocaine-induced enhancement of sensory-related neural activity in humans is likely to be detectable by neuroimaging. PMID- 15275931 TI - Mapping hippocampal and ventricular change in Alzheimer disease. AB - We developed an anatomical mapping technique to detect hippocampal and ventricular changes in Alzheimer disease (AD). The resulting maps are sensitive to longitudinal changes in brain structure as the disease progresses. An anatomical surface modeling approach was combined with surface-based statistics to visualize the region and rate of atrophy in serial MRI scans and isolate where these changes link with cognitive decline. Sixty-two [corrected] high-resolution MRI scans were acquired from 12 AD patients (mean [corrected] age +/- SE at first scan: 68.7 +/- 1.7 [corrected] years) and 14 matched controls (age: 71.4 +/- 0.9 years) [corrected] each scanned twice (1.9 +/- 0.2 [corrected] years apart, when all subjects are pooled [corrected] 3D parametric mesh models of the hippocampus and temporal horns were created in sequential scans and averaged across subjects to identify systematic patterns of atrophy. As an index of radial atrophy, 3D distance fields were generated relating each anatomical surface point to a medial curve threading down the medial axis of each structure. Hippocampal atrophic rates and ventricular expansion were assessed statistically using surface-based permutation testing and were faster in AD than in controls. Using color-coded maps and video sequences, these changes were visualized as they progressed anatomically over time. Additional maps localized regions where atrophic changes linked with cognitive decline. Temporal horn expansion maps were more sensitive to AD progression than maps of hippocampal atrophy, but both maps correlated with clinical deterioration. These quantitative, dynamic visualizations of hippocampal atrophy and ventricular expansion rates in aging and AD may provide a promising measure to track AD progression in drug trials. PMID- 15275932 TI - Diffusion tensor imaging detects early Wallerian degeneration of the pyramidal tract after ischemic stroke. AB - We used diffusion tensor imaging (DTI) to assess Wallerian degeneration of the pyramidal tract within the first 2 weeks after ischemic stroke, and correlated the extent of Wallerian degeneration with the motor deficit. Nine patients with middle cerebral artery stroke were examined 2-16 days after stroke by DTI and T2 weighted MRI. We measured fractional anisotropy (FA), averaged diffusivity (Dav), eigenvalues of the diffusion tensor and T2-weighted signal in the cerebral peduncle and compared these values between the affected and the unaffected side and between patients and six controls. FA was significantly reduced on the affected side compared to the unaffected side and compared to the control group. The largest eigenvalue was reduced, whereas the smallest eigenvalue was elevated on the affected side. There was no significant difference in T2-weighted signal and Dav. The decrease of anisotropy correlated positively with the motor deficit at the time of DTI study and 90 days after stroke. The reduction of anisotropy mirrors the disintegration of axonal structures, as it occurs in the early phase of Wallerian degeneration. DTI detects changes of water diffusion related to beginning pyramidal tract degeneration within the first 2 weeks after stroke that are not yet visible in conventional T2-weighted or orientationally averaged diffusion weighted MRI. We demonstrated for the first time a correlation of early DTI findings of pyramidal tract damage with the motor deficit. DTI can help prognosing recovery of motor function after stroke within the early subacute phase. PMID- 15275933 TI - Feedforward and feedback processes in motor control. AB - In this study, we utilized functional magnetic resonance imaging (fMRI) to examine which brain regions contribute to feedback and feedforward motor control processes. Several studies have investigated the contributions of cortical and subcortical brain regions to motor performance by independently varying factors such as movement rate, force, and speed, and observing the neural responses. Such studies have contributed greatly to our understanding of neural coding of movement variables. Under natural movement conditions, however, these factors interact in a complex manner to produce differing performance levels. In the current investigation, we induced performance changes in a less constrained way, by having subjects move a joystick to hit targets of differing sizes on an LCD screen. These parametric changes in target size resulted in the well-known speed accuracy tradeoff effect, allowing us to examine the brain regions responsive to global shifts in motor performance levels. That is, movements made to larger targets relied more on feedforward control whereas movements made to smaller targets relied more on feedback control. Using functional MRI, we identified two sets of brain regions in which activation was modulated with task difficulty. Areas exhibiting activation that was positively correlated with increasing target size included primary motor cortex, premotor cortex, and the basal ganglia, regions that are typically classified as playing a role in force control and movement planning. Brain regions whose activation was negatively correlated with increasing target size included the ipsilateral sensorimotor cortex, multiple cerebellar regions, and the thalamus. These areas contributed to motor performance under higher levels of task difficulty. The results elucidate cortical and subcortical brain regions that are responsive to global shifts in motor performance, reflecting changes along the continuum of feedforward and feedback motor control. PMID- 15275934 TI - The relationship between visually evoked cerebral blood flow velocity responses and visual-evoked potentials. AB - A noninvasive assessment of neurovascular coupling would be of great importance. For this reason, we simultaneously studied graded responses of visually evoked cerebral blood flow (CBF) velocity responses (VEFR) and visual-evoked potentials (VEP) to visual contrasts. The records were made from 30 healthy volunteers aged 38.0 +/- 9.6 years. The stimulus was a black-and-white checkerboard with visual contrasts (VC) of 1%, 10%, and 100%. The VEFR were measured in the posterior cerebral artery using transcranial Doppler, and the VEP were recoded from the scalp from occipital leads. To test the relationship between the VEFR and the VEP, a linear regression analysis was performed. We found that the VEFR at 100% VC were 36% higher than those at 10% VC (P < 0.01). The VEFR at 10% VC were 81% higher than those at 1% VC (P < 0.01). The VEP at 100% VC were 76% higher than those at 10% VC (P < 0.01). The VEP at 10% VC were 184% higher than those at 1% VC (P < 0.01). The linear regression showed a significant, moderate association between the VEP and the VEFR (r = 0.66, P < 0.01). The analysis of the regression slopes (b = 0.48 in older subjects vs. b = 0.58 in younger subjects) between two different age subgroups (P < 0.01) did not show any significant difference (P = 0.035). We concluded that a simultaneous recording of VEFR and VEP to graded visual contrasts could allow an assessment of neurovascular coupling. PMID- 15275935 TI - Deformable registration of cortical structures via hybrid volumetric and surface warping. AB - Registration of cortical structures across individuals is a very important step for quantitative analysis of the human brain cortex. This paper presents a method for deformable registration of cortical structures across individuals, using hybrid volumetric and surface warping. In the first step, a feature-based volumetric registration algorithm is used to warp a model cortical surface to the individual's space. This step greatly reduces the variation between the model and individual, thus providing a good initialization for the next step of surface warping. In the second step, a surface registration method, based on matching geometric attributes, warps the model surface to the individual. Point correspondences are also established at this step. The attribute vector, as the morphological signature of surface, was designed to be as distinctive as possible, so that each vertex on the model surface can find its correspondence on the individual surface. Experimental results on both synthesized and real brain data demonstrate the performance of the proposed method in the registration of cortical structures across individuals. PMID- 15275936 TI - Correlation of functional activation in the rat spinal cord with neuronal activation detected by immunohistochemistry. AB - The relationship between neuronal activity in the rat cervical and lumbar spinal cord was examined using functional magnetic resonance imaging (fMRI) and immunohistochemistry. Neuronal activity determined by c-fos staining was greatest between L4 and L6, and C5 to C7 spinal cord segments during noxious electrical stimulation of the rat hindpaw and forepaw, respectively. Areas of activity determined by fMRI are consistent with spinal cord physiology, and are predominantly found in regions of the spinal cord associated with pain, namely the dorsal horn. Activity in the ventral region of the cord was also observed, as expected. Combined results from repeated experiments demonstrated consistent areas of activity in response to stimulation, and show a high degree of reproducibility. Good correspondence was observed between functional MRI and sites of neuronal activity determined by c-fos labeling. PMID- 15275937 TI - Lexical learning of the English language: a PET study in healthy French subjects. AB - To investigate the neural correlates of word learning in adults, 10 right-handed French subjects who had learned English without mastering it performed an English and a French naming task during two PET sessions, one before (PET1) and the second after (PET2) a 4-week lexical training in English. Behavioral performance was collected during the two PET exams and 2 months after (T3). At T2, performance on English naming increased in all subjects; this improvement persisted at T3, with no correlation between English performance at T2 and T3. Cerebral activation during French naming mainly showed a left frontal temporal network. The pattern specifically associated with English lexical learning included, in addition to the anterior cingulate cortex involved in attentional processing and BAs 4/6 reflecting speech output, the right cerebellum and the left insular cortex that are linked to speech gesture learning, and the right medial temporal regions, likely to reflect the involvement of episodic memory during verbal learning. Correlations between English T2/T1 performance and English T2/T1 rCBF changes reinforced the hypothesis of intervention of episodic memory since they interested right frontal, hippocampal, and lateral temporal regions. 'Predictive' correlations between English T3/T2 performance and English T2/T1 rCBF changes showed, in good reminders, increased activities in the left posterior superior temporal sulcus and middle temporal cortex probably related to efficient semantic storage of learned words. PMID- 15275938 TI - Visual word recognition: the first half second. AB - We used magnetoencephalography (MEG) to map the spatiotemporal evolution of cortical activity for visual word recognition. We show that for five-letter words, activity in the left hemisphere (LH) fusiform gyrus expands systematically in both the posterior-anterior and medial-lateral directions over the course of the first 500 ms after stimulus presentation. Contrary to what would be expected from cognitive models and hemodynamic studies, the component of this activity that spatially coincides with the visual word form area (VWFA) is not active until around 200 ms post-stimulus, and critically, this activity is preceded by and co-active with activity in parts of the inferior frontal gyrus (IFG, BA44/6). The spread of activity in the VWFA for words does not appear in isolation but is co-active in parallel with spread of activity in anterior middle temporal gyrus (aMTG, BA 21 and 38), posterior middle temporal gyrus (pMTG, BA37/39), and IFG. PMID- 15275939 TI - Diffusion tensor MRI visualizes decreased subcortical fiber connectivity in focal cortical dysplasia. AB - Diffusion tensor imaging (DTI) was applied to 12 patients with focal cortical dysplasia (FCD) in frontal or occipital cortex. Fiber tractography was obtained from seeding points in superior longitudinal fasciculus or posterior corona radiata. Mean fractional anisotropy of fiber bundles around the affected cortex was decreased in comparison to the contralateral hemisphere with statistical significance (paired t test, P = 0.0274). On visual analysis, tractography depicted decreased volume of fiber bundles connected to the dysplastic cortex invariably even in those with a normal T2 signal intensity of underlying white matter adjacent to FCD. DTI has high potential to be applied to localize the FCD and to provide a better understanding of the pathological changes in the white matter. PMID- 15275940 TI - Panic attacks, depression and anxiety symptoms, and substance use behaviors during late adolescence. AB - This study examines panic attacks and substance use in a sample of incoming college freshman (n = 399 ) using questionnaires. Panickers (n = 47 ) were significantly more likely than nonpanickers (n = 290) to report having ever used sedatives, stimulants, opiods, and other drugs, but not tobacco, alcohol, cocaine, or hallucinogens. Gender and race did not substantially moderate the associations between substance use and panic attacks. Sedative, stimulant, opiod, and other drug use was not associated with panic attack frequency or the occurrence of unexpected attacks. The relationships of anxiety and depression with substance use were larger for panickers than nonpanickers. These results are consistent with the idea that self-medication and symptom exacerbation play a role in the development of co-occurring substance use disorders and mood and anxiety disorders. PMID- 15275941 TI - The relationship between trait vulnerability and anxiety and depressive diagnoses at long-term follow-up of Generalized Anxiety Disorder. AB - The current study examined the relationship between measures of trait vulnerability and long-term outcome in 83 patients diagnosed and treated for Generalized Anxiety Disorder (GAD) 8-14 years previously. Diagnostic status was assessed by structured interview, and trait affect, trait anxiety and trait depression were measured by the Positive and Negative Affect Scale (PANAS), the State-Trait Anxiety Inventory (STAI-T) and the Personal Style Inventory (PSI), respectively. Trait measures were all highly inter-correlated, and patients with diagnoses of GAD, social phobia and depressive disorders at long-term follow-up recorded very poor scores on all three scales. Trait anxiety recorded pre treatment was also related to both anxiety and depression at long-term follow-up. However, trait depression showed no significant association with panic disorder. Increased numbers of comorbid diagnoses were strongly related to high levels of both trait anxiety and negative affect (NA). The findings suggest that patients reporting high trait anxiety or NA may suffer from a chronic course of disorder and higher levels of comorbidity over the longer term. PMID- 15275942 TI - Attentional bias to threat and emotional response to biological challenge. AB - Attentional bias towards threat reliably correlates with clinical anxiety status as well as elevated trait anxiety. Although such findings have led many to posit a potential causative or predictive role of threat-biased attentional processes on anxiety problems, little informative research exists. The present investigation was designed to address the role of threat-biased attentional processes on emotional/fearful responding. Eighty-seven participants provided baseline measures of anxiety vulnerability (i.e., anxiety sensitivity; unmasked/masked emotional Stroop task indices) and then underwent biological challenge procedures (inhalations of 20% carbon dioxide (CO2)-enriched air). Following challenge, participants completed measures of emotional response. Regression analyses indicated that both unmasked and masked attentional bias indices significantly predicted emotional responding above and beyond anxiety sensitivity. Exploratory analyses also revealed a gender effect, with prediction of emotional response largely attributable to females. These findings support attentional bias towards threat as a relatively independent factor predictive of emotional responding. PMID- 15275943 TI - The Internet: home to a severe population of individuals with social anxiety disorder? AB - The current study sought to understand better the psychological characteristics of socially anxious individuals who seek information on the internet about social anxiety disorder and its treatment. Participants were 434 individuals who responded to an internet-based survey linked to the website of an anxiety specialty clinic. Using established cut-off scores, 92% of the sample met criteria for social anxiety disorder. Internet survey respondents who met these criteria reported greater severity of and impairment due to social anxiety than a treatment-seeking sample of persons with social anxiety disorder. Nevertheless, only about one-third of these internet respondents reported having received psychotherapy, and a similar percentage reported having received pharmacotherapy. Those with the most severe social interaction anxiety and who spent the most time interacting on the internet endorsed positive effects of internet use. However, a significant number of negative effects also were endorsed. PMID- 15275944 TI - Post-event processing and the retrieval of autobiographical memories in socially anxious individuals. AB - Individuals with social anxiety often report considerable ruminative thoughts following ambiguous social events (post-event processing). The purpose of this study was to determine whether post-event processing affects retrieval of autobiographical memories rated as negative, anxious and shameful in a sample of socially anxious individuals and controls. Results indicated that, compared to controls, socially anxious individuals recalled memories that were rated as significantly more negative and shameful regardless of the type of post-event processing engaged in. Unexpectedly, after negative post-event processing socially anxious individuals recalled memories that although anxious and shameful, were rated as significantly more calming than after other types of post event processing. The results imply that post-event processing may have some adaptive benefit that could explain why it persists in socially anxious individuals. PMID- 15275945 TI - Psychometric evaluation of the Social Anxiety Scale for Adolescents and the Social Phobia and Anxiety Inventory for Children: construct validity and normative data. AB - This study evaluated the psychometric properties of the Social Anxiety Scale for Adolescents (SAS-A) and Social Phobia and Anxiety Inventory for Children (SPAI-C) in a sample of 1147 adolescents aged 13-17 years. The fit indices of confirmatory factor analyses were comparable to those obtained in prior studies and supported the hypothesized models of the SAS-A and SPAI-C. The internal consistency was good and 12-month test-retest reliability modest for both measures. A significant, positive correlation was found between the SAS-A and SPAI-C, showing that these measures assess related, but relatively independent constructs of social anxiety and phobia. These findings support the use of the SAS-A and SPAI-C with adolescents. PMID- 15275946 TI - The measurement and impact of childhood teasing in a sample of young adults. AB - This study examined the psychometric properties of the Teasing Questionnaire Revised (TQ-R) and the relationships among recalled childhood teasing and current psychosocial distress in 414 undergraduate students. Participants were administered the TQ-R, Beck Depression Inventory-II, State-Trait Anxiety Inventory-Trait Version, Brief Fear of Negative Evaluation Scale, and UCLA Loneliness Scale. Confirmatory factor analysis supported a five-factor model assessing teasing related to performance, academic issues, social behavior, family background, and appearance. Internal consistency of the TQ-R and its factors was acceptable, and intercorrelations among subscales were moderate, suggesting that the factors measure related but conceptually distinct teasing experiences. Defining Pearson product-moment correlations with a magnitude of greater than.25 as conceptually meaningful, we found that the TQ-R Total score was meaningfully related to depressive symptoms, anxiety, fear of negative evaluation, and loneliness. Being teased in the Performance and Social domains as a child was moderately related to current psychopathology. Implications of these findings for clinical practice and future research are discussed. PMID- 15275947 TI - Gender differences in Childhood Anxiety Sensitivity Index (CASI) dimensions. AB - We examined the hierarchical structure of the Childhood Anxiety Sensitivity Index (CASI) as a function of gender and examined the occurrence of gender differences in anxiety sensitivity (AS) dimensions in a large nonclinical sample of children and adolescents (N = 1698). Separate principal components analyses (PCAs) on the 18 CASI items for the total sample, boys, and girls revealed similar lower-order three-factor structures for all groups. The three factors reflected Physical, Social/Control, and Psychological Concerns. PCAs on the lower-order factor scores revealed similar unidimensional higher-order solutions for all groups. Girls scored higher than boys on the Physical and, to a lesser extent, Social/Control Concerns factors; girls scored higher on the Physical Concerns factor relative to their scores on the Social/Control and Psychological Concerns factors; and boys scored higher on the Social/Control and Psychological Concerns factors relative to their scores on the Physical Concerns factor. Girls also scored higher than boys on the higher-order factor representing the Global AS construct. The present study provides additional support for the theoretical hierarchical structure of AS and suggests that there is a difference in the manifestation of AS between girls and boys. PMID- 15275948 TI - A naturalistic examination of positive expectations, time course, and disgust in the origins and reduction of spider and insect distress. AB - We used a naturalistic method to examine the causes of changes in individuals' reactions to, and feelings about, spiders and insects. In this descriptive retrospective study, 50 college students who reported substantial changes in their attitudes toward spiders and/or insects (in the absence of professional treatment) underwent telephone interviews about the change process. We found that individuals frequently describe the role of positive experiences and expectations in positive change and some individuals report sudden changes. Further, descriptions of the important role of disgust in the change process were common. We discuss the implications of our findings for understanding the etiology and treatment of spider and insect distress and make a case for the usefulness of naturalistic methods in expanding scientific knowledge. PMID- 15275949 TI - The neglected relationship between social interaction anxiety and hedonic deficits: differentiation from depressive symptoms. AB - Depressive symptoms are associated with both the presence of negative subjective experiences and relative absence of positive subjective experiences. A similar affective profile of high negative affect and low positive affect (PA) has been associated with excessive social anxiety (SA). This initial cross-sectional study evaluated the incremental effects of social interaction anxiety on hedonic deficits beyond the effects of depressive and anxiety (i.e., physiological arousal, worry) symptoms. From a sample of 97 college students, a factor analysis on self-report measures of hedonic functioning derived two domains: Positive Subjective Experiences and Curiosity. Social interaction anxiety was uniquely, negatively related to Positive Subjective Experiences and Curiosity after removing variance attributable to various depressive and anxiety symptoms. In contrast, anxious arousal and nonspecific anxiety had near-zero relationships with both domains, and depressive symptoms were negatively related to Positive Subjective Experiences. These data provide some evidence for an association between social interaction anxiety and hedonic deficits that is not attributable to covariance with other internalizing conditions. PMID- 15275950 TI - A novel method for induction of salt appetite in rats. AB - A number of procedures exist for the experimental induction of sodium appetite. With the exception of a low sodium diet, nearly all of these methods are invasive, requiring injections, surgery, or both. In addition to stimulating intake of concentrated salt, some of them produce substantial side-effects like reduced food intake and weight gain. The present experiment was designed to evaluate the efficacy of a novel non-invasive method to induce a salt appetite. We investigated the effects of ingesting 100 microM amiloride, a diuretic and natriuretic compound, on urine quantity, electrolyte balance, 0.5 1M sodium chloride (NaCl) intake, water intake, and Na(+)-free chow intake, and weight gain in rats. A water ingestion only group served as control. Consumption of amiloride in mixture with water produced greater loss of urinary Na(+) and intake of 0.51 M NaCl compared with controls. This treatment was without effect on food intake and only modestly influenced weight gain. These results demonstrate a rapid and non invasive method for the induction of salt appetite free of unwanted side-effects. PMID- 15275951 TI - Investigation of an inducible nitric oxide synthase gene (NOS2A) polymorphism in a multiple sclerosis population. AB - Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system (CNS) affecting most commonly the Caucasian population. Nitric oxide (NO) is a biological signaling and effector molecule and is especially important during inflammation. Inducible nitric oxide synthase (iNOS) is one of the three enzymes responsible for generating NO. It has been reported that there is an excessive production of NO in MS concordant with an increased expression of iNOS in MS lesions. This study investigated the role of a bi-allelic tetranucleotide polymorphism located in the promoter region of the human iNOS (NOS2A) gene in MS susceptibility. A group of MS patients (n = 101) were genotyped and compared to an age- and sex-matched group of healthy controls (n = 101). The MS group was subdivided into three subtypes, namely relapsing-remitting MS (RR-MS), secondary progressive MS (SP-MS) and primary-progressive MS (PP-MS). Results of a chi squared analysis and a Fisher's exact test revealed that allele and genotype distributions between cases and controls were not significantly different for the total population (chi(2) = 3.4, P(genotype) = 0.15; chi(2) = 3.4, P(allele) = 0.082) and for each subtype of MS (P > 0.05). This suggests that there is no direct association of this iNOS gene variant with MS susceptibility. PMID- 15275952 TI - An experimental method for measuring the mean length of cerebellar parallel fibers: validation and derivation of a correction factor by computational simulation and probability analysis. AB - The length of cerebellar parallel fibers is important for information integration by the Purkinje cells. Based on the Copernican principle for analyzing the length of stochastic events, we have recently devised a stochastic method to estimate the mean length of parallel fibers within a given cerebellar region. The purpose of the present report is to provide validation of this methodology via computational simulations. We create virtual parallel fibers with known lengths and program each step of our stochastic method for computational simulation. We then compare the observed mean length obtained from our computational simulation with the known mean length of the virtual parallel fibers. In particular, we investigate the effect of cutting parallel fibers into segments during histological sectioning. Our computational results reveal an over-estimation factor ranging from 1.0 (no correction is necessary) to 2.0 as the parallel fiber segmentation becomes increasingly severe. Based on probability theory considerations, we have confirmed the existence of this over-estimation. We have further determined the cause of this over-estimation to be an artificial consequence of one of the sampling steps in our stochastic method. These results provide validation of our methodology, as well as a correction factor, which can be derived directly from the experimentally measured parameters and used to obtain the true mean length of parallel fibers. Potential applications of the stochastic method include a comparative analysis of the length of parallel fibers as an approach to gain clues about cerebellar circuit principles and function. In addition, the stochastic method may also find promising applications in other functionally important axonal systems in the brain. PMID- 15275953 TI - Impaired motor performance in mice lacking neurosteroid vitamin D receptors. AB - Vitamin D is a neuroactive seco-steroid and its importance to the nervous system is receiving increasing recognition. Since numerous data link vitamin D dysfunctions to various neurological and behavioural disorders, we studied whether genetic ablation of vitamin D receptors (VDR) may be associated with motor impairments in mice subjected to several behavioural tests. The data obtained in the vertical screen and swim tests show that VDR genetic ablation produces severe motor impairment (shorter screen retention and poor swimming) in mutant mice compared to wild-type and heterozygous control animals. These impairments appear to be unrelated to visual, vestibular and activity/emotionality parameters of mice, and are likely associated with disturbed calcium homeostasis. This study confirms the important role of the vitamin D system in motor functions and suggests that animal genetic models targeting the vitamin D/VDR system may be a useful tool to study vitamin D related motor/behavioural disorders. PMID- 15275954 TI - Cue valence representation studied by Fos immunocytochemistry after acquisition of a discrimination learning task. AB - The piriform cortex (PCx) and related structures such as hippocampus and frontal cortex could play an important role in olfactory memory. We investigated their involvement in learning the biological value of an odor cue, i.e. predicting reward or non-reward in a two-odor discrimination task. Rats were sacrificed after stimulation by either rewarded or non-rewarded odor and Fos immunocytochemistry was performed. The different experimental groups of rats did not show strongly differentiated Fos expression pattern in either the PCx or the hippocampus. A few differences were noted in frontal areas. In the ventro-lateral orbital cortex, rats, ramdomly rewarded during the conditionning had a higher Fos level in comparison with other groups. In infralimbic cortex, rats, which learned the reward value of the olfactory cue and were water-reinforced the day of sacrifice, showed a higher Fos expression. Data are discussed in view of the olfactory learning paradigm and of the accuracy of the control groups used in the present experimental design. The behavioural conditions leading to Fos expression are further discussed since Fos is a marker of learning-induced plasticity as well as a general activity marker which can be activated by a wide range of stimuli not directly linked to memory. PMID- 15275955 TI - Delayed non-matching to position performance in aged hybrid Fischer 344 x brown Norway rats: a longitudinal study. AB - In this study, the effects of aging on the performance in a delayed non-matching to position (DNMTP) task were investigated longitudinally in hybrid Fischer 344 x Brown Norway rats. The rats were first trained to perform the task. Subsequently, their performance was assessed monthly from 28 to 34 months of age. The measures of responding on the DNMTP schedule did not decrease in the course of the study. After the last DNMTP test, choline acetyltransferase (ChAT) activity and glial fibrillary acidic protein (GFAP) content were measured in frontal cortex and hippocampus. We found that higher levels of GFAP in the frontal cortex, but not hippocampus, were associated with a poorer performance in the DNMTP task. Our findings support the notion that repeated testing prevents the age-related decline in cognitive functions that has been reported in cross-sectional studies. Pathology of the frontal cortex seems to predict a faster rate of forgetting in aging rats. PMID- 15275956 TI - Increased urinary F(2)-isoprostanes levels in the patients with Alzheimer's disease. AB - Isoprostanes, novel markers of oxidative injury, are generated by the free radical-mediated peroxidation of arachidonic acid (AA). They are thought to be important in the pathogenesis of neurodegenerative diseases such as Alzheimer's disease (AD). Using gas chromatography-mass spectrometry (GC-MS), we investigated the alteration of urinary F(2)-isoprostanes in AD patients compared to that of healthy subjects. Our results show that the levels of urinary F(2)-isoprostanes, sum of the prostaglandin F(2 alpha) isomer; prostaglandin F(2 alpha) (PGF(2 alpha)), prostaglandin F(2 beta) (PGF(2 beta)), and 8-isoprostaglandin F(2 alpha) (8-isoPGF(2 alpha)), significantly increased in AD patients (P < 0.05). The concentration of urinary 8-isoPGF(2 alpha), one of the biomarkers of oxidative stress, was not significantly different between 34 AD patients and 20 age-matched controls (P > 0.05). The PGF(2 alpha), formed by endoperoxide reductase from PGH(2), was significantly increased in AD patients, when compared with the levels of the normal controls (P < 0.05). The PGF(2 alpha), an enzymatic product of arachidonic acid, may affect the pathogenesis of AD. In addition, urinary F(2) isoprostanes can play an important role as a biomarker in AD. PMID- 15275957 TI - Indomethacin attenuates hyperthermia produced by anterior coronal lateral hypothalamic knife cuts. AB - Electrolytic lesions of the lateral hypothalamus (LH) and coronal knife cuts of fibers anterior to the LH produce an elevation in core body temperature, or hyperthermia. Prostaglandin has been shown to mediate hyperthermia produced by electrolytic LH lesions. The present study characterizes the time course and the role of prostaglandin in mediating knife-cut-induced hyperthermia. Results show that the prostaglandin synthesis inhibitor indomethacin significantly attenuates hyperthermia produced by the knife cuts, suggesting that prostaglandin is involved in mediating this temperature increase. A disruption of axonal fibers that project from the LH to the preoptic area is postulated to be responsible for the temperature increase. There was no effect of knife cuts on food intake and body weight loss, which were also measured, suggesting that this fiber system is not involved in feeding behavior. PMID- 15275958 TI - Decreases in the expression of CGRP and galanin mRNA in central and peripheral neurons related to the control of blood pressure following experimental hypertension in rats. AB - Calcitonin gene-related peptide (alpha CGRP) and galanin (GAL) are peptides known to participate in central mechanisms of blood pressure control. Nonetheless, variations in the synthesis of the peptides in response to a hypertensive challenge are not well described, specially using a model, which allows acute and chronic analyses. In this study, we have employed in situ hybridization to analyse changes in mRNA expression of alpha CGRP and GAL in the nucleus tractus solitarii (NTS), hypothalamic paraventricular nucleus (PVN) as well as petrosal and nodose ganglia after aortic coarctation-induced hypertension in rats. Acute (2h) and chronic (3 and 7 days) analyses were performed in order to evaluate the involvement of both peptides in different periods of hypertension. The analysis of relative mRNA levels showed significant differences between sham-operated and aortic coarcted hypertensive rats. alpha CGRP mRNA expression was decreased 2h (40%) and 3 days (42%) in nodose and petrosal ganglia, respectively, after coarctation. No changes in CGRP mRNA signal were seen in the NTS and PVN in the analysed periods. GAL mRNA expression was decreased in the NTS (19%) and PVN (55%), 3 and 7 days, respectively, after coarctation-induced hypertension. No changes in GAL mRNA expression were observed in petrosal and nodose ganglia following aortic coarctation. Data suggest that alpha CGRP and GAL may participate in the mechanisms involved in the establishment/maintenance of hypertension induced by aortic coarctation. Acute changes might be involved with the adaptation to the hypertensive state, while changes at the chronic phase might be related to counteraction of hypertension. PMID- 15275959 TI - Opposite morphological responses of partially denervated cortical serotonergic and noradrenergic axons to repeated stress in adult rats. AB - We examined plastic changes in serotonin (5-HT) axons following repeated stress in the adult rat brain, and compared stress-induced changes between 5-HT and noradrenaline (NA) axons. We locally injected the specific neurotoxin to 5-HT axons or to NA axons into the frontal cortex to cause partial denervation. The animals were mildly restrained from 1 day after the neurotoxin injection and this stress was repeated daily for 20 min during the first 2 days and for 40 min during the next 11 days. On the fourteenth day after injection, the brains were removed to visualize 5-HT and NA axons by immunohistochemistry. Repeated stress did not significantly alter the denervation area of 5-HT or NA axons, but the density of 5-HT axons was increased whereas that of NA axons was decreased in cortical regions outside the denervation site. In addition, the expression of brain-derived neurotrophic factor (BDNF) was increased in cortical regions where the 5-HT axon density was increased in response to stress. These results suggest that repeated stress causes opposite changes in the morphology of partially denervated 5-HT and NA axons in the cerebral cortex. The stress-induced increase in BDNF expression may contribute to 5-HT axon sprouting following repeated stress. PMID- 15275960 TI - Regulation by equilibrative nucleoside transporter of adenosine outward currents in adult rat spinal dorsal horn neurons. AB - A current response induced by superfusing adenosine was examined in substantia gelatinosa (SG) neurons of adult rat spinal cord slices by using the whole-cell patch-clamp technique. In 78% of the neurons examined, adenosine induced an outward current at -70 mV [18.8 +/- 1.1 pA (n = 98) at 1mM] in a dose-dependent manner (EC(50) = 177 microM). A similar current was induced by A(1) agonist N(6) cyclopentyladenosine (1 microM), whereas A(1) antagonist 8-cyclopentyl-1,3 dipropylxanthine (1 microM) reversed the adenosine action. The adenosine current reversed its polarity at a potential being close to the equilibrium potential for K(+), and was attenuated by Ba(2+) (100 microM) and 4-aminopyridine (5mM) but not tetraethylammonium (5mM). The adenosine current was enhanced in duration by equilibrative nucleoside-transport (rENT1) inhibitor S-(4-nitrobenzyl)-6 thioinosine (1 microM) and adenosine deaminase (ADA) inhibitor erythro-9-(2 hydroxy-3-nonyl) adenine (1 microM), and slowed in falling phase by adenosine kinase (AK) inhibitor iodotubercidine (1 microM). We conclude that a Ba(2+)- and 4-aminopyridine-sensitive K(+) channel in SG neurons is opened via the activation of A(1) receptors by adenosine whose level is possibly regulated by rENT1, adenosine deaminase and adenosine kinase. Considering that intrathecally administered adenosine analogues produce antinociception, the regulatory systems of adenosine may serve as targets for antinociceptive drugs. PMID- 15275961 TI - Differential effects of NMDA and AMPA/kainate receptor antagonists on superoxide production and MnSOD activity in rat brain following intrahippocampal injection. AB - The involvement of NMDA and AMPA/kainate receptors in the induction of superoxide radical production in the rat brain was examined after injection of kainate, non NMDA receptor agonist, kainate plus 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), selective AMPA/kainate receptor antagonist, or kainate plus 2-amino-5 phosphonopentanoic acid (APV), selective NMDA receptor antagonist. Competitive glutamate receptor antagonists were injected with kainate unilaterally into the CA3 region of the rat hippocampus. We investigated superoxide production and mitochondrial MnSOD activity after injection. The measurements took place at different times (5, 15 min, 2, 48 h and 7 days) in the ipsi- and contralateral hippocampus, forebrain cortex, striatum, and cerebellum homogenates. Used glutamate antagonists APV and CNQX both expressed sufficient neuroprotection in sense of decreasing superoxide production and increasing MnSOD levels, but with differential effect in mechanisms and time dynamics. Our findings suggest that NMDA and AMPA/kainate receptors are differentially involved in superoxide production. Following intrahippocampal antagonists injection they, also, interpose different neuroprotection effect on the induction of MnSOD activity in distinct brain regions affected by the injury, which are functionally connected via afferents and efferents. It suggests that MnSOD protects the cells in these regions from superoxide-induced damage and therefore may limit the retrograde and anterograde spread of neurotoxicity. PMID- 15275962 TI - Amplification of T-cell responses by neutrophils: relevance to allograft immunity. AB - Because rejection of allografts is primarily caused by T and B lymphocyte responses to allogenic histocompatibility molecules, the role of innate immunity in organ transplant rejection is often overlooked. However, the very first damages to vascularized organ allografts are caused by ischemia-reperfusion, an inflammatory reaction involving activation of vascular endothelial cells and release of neutrophil chemoattractants. Herein, we review experimental observations suggesting that the early neutrophil influx in organ transplants favors T cell-mediated rejection. PMID- 15275963 TI - CD45: direct and indirect government of immune regulation. AB - The protein tyrosine phosphatase (PTP) CD45 is abundantly expressed on all nucleated hematopoietic cells and is critical for classical antigen receptor signalling indicated by the arrested development of B and T cells in mice deficient for CD45. Despite its clear role in positive regulation of signalling through the activation of the Src family of tyrosine kinases, many reports have shown CD45 to also negatively regulate this process. Given such a dichotomy in CD45 function and a poor understanding of the mechanism underlying the phenotype of CD45(-/-) lymphocytes, we considered it timely to review the existing data and attempt to determine whether aspects of the CD45(-/-) phenotype result from excessive positive or negative kinase activity and the target molecules that may mediate such effects. PMID- 15275964 TI - Human neutrophil-derived CAP37 inhibits lipopolysaccharide-induced activation in murine peritoneal macrophages. AB - Human cationic antimicrobial protein, CAP37, is released from neutrophil granules during infection. CAP37 attracts monocytes, binds Gram-negative endotoxin (lipopolysaccharide, LPS), is bactericidal for a range of Gram-negative bacteria, and reduces mortality in murine polymicrobial sepsis. Here, we report that recombinant CAP37 specifically targets murine peritoneal macrophages. Under steady-state conditions, the bulk of cell-associated CAP37 was localized at the plasma membrane. However, a fraction of CAP37 gained access to the endocytic system, but did not accumulate in recycling endosomes or in the trans-Golgi network (TGN). Instead, CAP37 was internalized by fluid phase endocytosis and accumulated in a prelysosomal compartment. Macrophages that were preexposed to CAP37 exhibited diminished LPS responsiveness, as determined by analysis of c-Jun phosphorylation. Further examination showed that pretreatment with CAP37 reduced the ability of macrophages to bind LPS. Taken together, these observations demonstrate that prolonged exposure to CAP37 desensitizes macrophages to LPS and suggest that this protein plays a novel anti-inflammatory role in polymicrobial sepsis. PMID- 15275965 TI - Soluble interleukin-6 receptor (sIL-6R) in cerebrospinal fluid of patients with inflammatory and non inflammatory neurological diseases. AB - IL-6 acts on target cells via the ligand-binding protein interleukin-6 receptor (IL-6R) and the affinity-converting and signal-transducing glycoprotein 130 (gp130). Soluble interleukin-6 receptor (sIL-6R) has an agonistic role because the soluble complex (IL-6/sIL-6R) can activate cells that do not express IL-6R and an antagonistic role as it enhances the inhibitory activity of sgp130. Soluble forms of both receptors, sIL-6R and sgp130, regulate the action of IL-6. sIL-6R was measured by a sensitive enzyme-linked immunosorbent assay in paired sera and cerebrospinal fluid (CSF) from 46 patients with inflammatory neurological diseases (IND), 45 patients with relapsing-remitting multiple sclerosis (RR-MS), 13 patients with primary progressive multiple sclerosis (PP MS), 17 patients with other non inflammatory neurological diseases (NIND) and 13 mentally healthy individuals--healthy controls (HC). Patients with RR-MS had CSF sIL-6R levels comparable to those from patients with IND, but higher than patients with NIND and HC. A positive correlation between the CSF/serum albumin (QAlb) and CSF sIL-6R levels was observed in IND but not in RR-MS patients indicating that CSF sIL-6R levels in IND patients could be influenced by serum sIL-6R and blood brain barrier (BBB) permeability properties. RR-MS patients had higher values of [CSF/serum sIL-6R:CSF/serum albumin] (sIL-6R index) than IND patients suggesting that in multiple sclerosis (MS), the increase in CSF sIL-6R could be due to intrathecal synthesis of sIL-6R. The finding of increased CSF sIL 6R concentrations (>979 pg/ml) with sIL-6R index (>4.66), in correlation with positive oligoclonal bands in RR-MS patients, suggests that values of sIL-6R index > 4.66 indicate intrathecal increase of sIL-6R and might be used as an indicator of neuroimmunoregulatory and inflammatory processes in the central nervous system (CNS). PMID- 15275966 TI - IL-12p40-overexpressing immature dendritic cells induce T cell hyporesponsiveness in vitro but accelerate allograft rejection in vivo: role of NK cell activation and interferon-gamma production. AB - Infusion of genetically modified dendritic cells (DC) expressing immunosuppressive molecules is a potential therapy for organ rejection. IL-12p70, a cytokine produced mainly by DC and macrophages, consists of two subunits, p40 and p35. IL-12p70 is an activator of T cells, while the IL-12p40 subunit serves as a natural antagonist for IL-12p70 action. The primary aim of this study was to evaluate the effect of IL-12p40 gene-modification on both the T-cell stimulatory activity of immature DC (imDC) and their ability to prolong cardiac allograft survival. IL-12p40 gene-modified imDC (DC-p40) exhibited a phenotype characteristic of imDC and displayed impaired T-cell allostimulatory ability in vitro. However, to our surprise, for murine vascularized heterotopic heart transplantation (HHT), administration of donor-derived DC-p40 7 days prior to transplantation did not prolong allograft survival but instead significantly exacerbated cardiac allograft rejection. Further study showed that DC-p40 augmented NK cell activity both in vitro and in vivo and enhanced interferon gamma (IFN-gamma) production in vivo, which might be due to the increased IL-23 production by DC-p40. Our data suggested that although IL-12p40 gene-modified immature DC can induce T cell hyporesponsiveness in vitro, their ability to activate NK cells and induce IFN-gamma production counterbalances this, exacerbating cardiac allograft rejection. The unexpected effects of DC-p40 limit their value in promoting allograft survival in vivo and likely reflect the complexity of IL-12p40 biology. PMID- 15275967 TI - Cytokine induction in human cord blood lymphocytes after pulsing with UV inactivated influenza viruses. AB - Mitogenic activity of UV-inactivated influenza viruses in cord blood lymphocytes (CBL), as measured by cytokine release, was investigated. Using prototype viruses of subtype H3N2 (A/Aichi/68), H2N2 (A/Japan/57), and H1N1 (A/Puerto Rico/34) for influenza A virus, and B/Lee/40 for influenza B virus, the results indicated that both Th1 and Th2 cytokines were induced. Stimulation indices were significantly higher for IFNgamma, IL-4 and IL-10 by influenza A viruses than by influenza B virus. Stimulation indices for IL-2 and IL-6 were lower, as these two cytokines were spontaneously released by cord blood lymphocytes in culture. Alignment of the amino acid sequences of the HA for the viruses used in this study indicated that influenza B virus lacked sequence homology to the antigenic sites identified for influenza A virus. Therefore, the antigenic sites may play a role in the mitogenic property, and cord blood lymphocytes could provide a system to compare this property for clinical isolates of influenza virus. PMID- 15275968 TI - Role of interaction between Ly49 inhibitory receptors and cognate MHC I molecules in IL2-induced development of NK cells in murine bone marrow cell cultures. AB - Murine bone marrow (BM) cell preparations lack mature cytotoxic natural killer (NK) cells, but NK cells may be induced in these cell preparations by culturing with interleukin-2 (IL2). Present study was aimed at studying the role of interactions between Ly49 molecules and major histocompatibility complex (MHC) class I molecules during IL2-induced development of mature NK cells in BM cell cultures. Addition of monoclonal antibodies (mabs) specific to class I MHC molecules of H-2b haplotype, to block any interaction of MHC I molecules with their receptors, was found to inhibit NK cell development. Mouse NK cells express several types of Ly49 molecules including Ly49C, which is an inhibitory receptor specific to MHC I molecules of H-2b haplotype. Blocking Ly49-MHC I interaction by using anti-Ly49C mab inhibited the development of cytotoxic NK cells. Addition of anti-Ly49A (no specificity for H-2b MHC I molecules) or anti-Ly49D (activating receptor specific for MHC I molecules of many H-2 haplotypes including H-2b) mabs, however, had no effect on IL2-induced NK cell development in BM cells. Mabs specific to Ly49C molecule and MHC I molecules of H-2b haplotype inhibited the development of mature NK cells from highly purified NK precursor cell population. These results indicate that specific interaction between inhibitory self-reactive Ly49 molecules and MHC I molecules may be crucial for NK cell development. We propose a model in which Ly49-MHC I interaction may have a permissive role in allowing development of only such NK cell clones that expresses at least one self reactive inhibitory Ly49 molecule so that lysis of autologous healthy cells by mature NK cells may be avoided. PMID- 15275969 TI - The expression of B7-H1 on keratinocytes in chronic inflammatory mucocutaneous disease and its regulatory role. AB - PD-1 and its ligands, B7-H1/PD-L1 and B7-DC/PD-L2, have been identified recently as CD28-B7 family molecules that are implicated in immune regulation. Lichen planus (LP) is a T cell-mediated chronic inflammatory mucocutaneous disease. We investigated the expression and function of PD-1 and its two ligands in LP. Immunohistochemical examination revealed the abundant expression of PD-1 and B7 H1 in infiltrating T cells and macrophages, and lower-level expression of B7-DC on macrophages in the subepithelium. Interestingly, substantial expression of B7 H1 on keratinocytes (KCs) was found close to the numerous T cell infiltrates in the subepithelium. Unstimulated cultured KCs expressed both B7-H1 and B7-DC, and their expression was upregulated by proinflammatory cytokines, particularly IFN gamma. The T-cell proliferative responses and IFN-gamma production that were induced by IFN-gamma-treated KCs were augmented preferentially by anti-B7-H1 mAb, but not by anti-B7-DC mAb. These results indicate the regulatory role of B7-H1 on KCs in the interactions with T cells. Our results suggest that the induction of B7-H1 on KCs may play an important role in tolerance induction in the inflamed oral mucosa and skin. PMID- 15275970 TI - Effects of Helicobacter pylori infection on gastric inflammation and local cytokine production in histamine-deficient (histidine decarboxylase knock-out) mice. AB - Helicobacter pylori infection in humans causes gastritis. The infection elicits a complex immune response in which the activation of mast cells and histamine release is of particular importance. Histamine further promotes the immune response and stimulates gastric acid secretion. The inflammatory effects of H. pylori can be studied in intragastrically infected mice. The aim of this study was to compare the local cytokine responses of histamine-deficient, histidine decarboxylase knock-out (HDC KO) and wild-type (WT) mice following H. pylori infection. METHODS: H. pylori was administered intragastrically to HDC KO and WT mice. The animals were infected three times in a 1-week-period and were sacrificed 8 weeks after the first intervention. The local TNF-alpha, IL-6 and IL 10 cytokine levels in gastric mucosal specimens were determined by ELISA. Gastric mucosa sections were also analysed for histological signs of inflammation. To investigate the antibody response following H. pylori infection, the total anti H. pylori IgG and the ratio of IgG1/IgG2a isotypes were determined in the serum by ELISA. RESULTS: H. pylori induced considerable cytokine production in the infected groups. The TNF-alpha and IL-6 levels were significantly higher in the WT mice than in the HDC KO mice, whereas the IL-10 levels did not differ between the groups. Anti-H. pylori IgG was detected only in the infected groups and the titre was higher in the WT mice. A higher IgG1/IgG2a ratio was observed in the H. pylori infected HDC KO group. Histological analysis revealed that the grades of inflammation were less severe in the infected HDC KO animals. CONCLUSIONS: The results suggest that H. pylori induces lower TNF-alpha and IL-6 secretion in the gastric mucosa in the HDC KO mice than in the WT animals, while the levels of induction of IL-10 were similar. The imbalance between Th1/Th2 is less pronounced in the HDC KO mice, which might explain the milder inflammation in the gastric mucosa. These results provide further information on the role of histamine in the pathomechanism of H. pylori-induced gastritis. PMID- 15275971 TI - The heat stable antigen (CD24) is not required for the generation of CD4+ effector and memory T cells by dendritic cells in vivo. AB - Previous work has established that CD24 is a costimulatory molecule for T-cell clonal expansion. Studies using CD24 -/- mice demonstrated that CD24 plays a critical role in the CD28-independent immune response against virus and soluble antigens. The role of CD24 on dendritic cells (DCs) has not been reported. Here, we compare the CD24(+/+) and CD24(-/-) DCs in the induction of initial clonal expansion and elicitation of memory CD4(+) T cells in vivo. Our results demonstrate that the CD24 expressed on DCs is essential neither for the induction of initial T-cell clonal expansion nor for elicitation of memory activity of primed T cells in vivo. PMID- 15275972 TI - Expression of membrane-bound and soluble receptor activator of NF-kappaB ligand (RANKL) in human T cells. AB - The receptor activator of NF-kappaB ligand (RANKL) and its receptor RANK are critical regulators for immune responses as well as bone remodeling. RANKL is a type II transmembrane protein that has two forms-a membrane-anchored protein and a secreted protein. In this report, we demonstrate for the first time the kinetical expression of two forms of RANKL in human T cells using two monoclonal antibodies (mAbs) against human RANKL, which we newly derived. Freshly isolated T cells rarely expressed mRANKL, while the activation of T cells induced a substantial but minimal level of mRANKL as well as the accumulation of considerable amounts of sRANKL. The addition of the metalloprotease inhibitor KB R8301 efficiently suppressed the release of sRANKL from activated T cells or RANKL-transfectants, and reciprocally enhanced the mRANKL expression. The membrane form of RANKL was also expressed on the infiltrating T cells in the rheumatoid synovial fluid and in the gingival tissues of patients with periodontitis. Our results demonstrate that the expression of mRANKL on T cells is strictly limited, and the majority of RANKL protein produced by T cells may be active in the soluble form after shedding. The mAbs that were derived in this study may be useful for investigating the regulation and function of RANKL in immune responses and bone remodeling. PMID- 15275973 TI - The inner side of T cell lipid rafts. AB - A key question in understanding the functional role of lipid rafts is whether lipid microdomains at the plasma membrane outer leaflet are coupled to lipid microdomains at the inner leaflet. By using a cyan-fluorescent protein (CFP) targeted to inner plasma membrane rafts of Jurkat T cells, we found that raft domains at the outer and inner leaflets are physically coupled and that this coupling requires cholesterol. Interestingly, TCR/CD3 cross-linking induces co capping of the raft bilayer independently of cholesterol or signaling events, indicating that cholesterol-extracting drugs are unable to destroy TCR-lipid rafts interaction. PMID- 15275974 TI - Recombinant hydrophilic human gp100: uptake by dendritic cells and stimulation of autologous CD8+ lymphocytes from melanoma patients. AB - Native gp100, a glycoprotein highly expressed in the majority of melanomas, contains several immunogenic peptides that are recognized by cytotoxic lymphocytes (CTLs) in the context of major histocompatibility complex (MHC) class I molecules. The objective of this study was to evaluate the ability of dendritic cells (DCs) from melanoma patients to take up gp100 protein and stimulate specific autologous CTL. The gp100 used in this study was a recombinant molecule with diminished hydrophobicity, HR-gp100, produced in Escherichia coli bacteria and in Pichia pastoris yeast. Stimulation of CD8+ T cells from melanoma patients with HR-gp100-loaded DC was visualized by confocal microscopy using stained target cells, and was quantitatively measured by the production of IFN-gamma using an ELISPOT assay. The results showed that HR-gp100 protein, produced either in bacteria or in yeast, when loaded on DC from melanoma patients, stimulated autologous CD8+ lymphocytes. By direct visualization, these lymphocytes were found in close contact with dead melanoma cells, and to contain membrane material transferred from stained melanoma cells; in cultures containing control lymphocytes stimulated with unloaded DC, no melanoma cell killing was observed. In ELISPOT assays, increased number of IFN-gamma-producing CD8+ T lymphocytes from patients, but not from healthy controls, were measured upon stimulation with HR-gp100-loaded DC. HR-gp100 could represent a useful tool to load DC with multiple immunogenic epitopes/antigen-derived epitopes for the immunotherapy of melanoma. PMID- 15275975 TI - Peptides eluted from HLA-B27 of human splenocytes and blood cells reveal a similar but partially different profile compared to in vitro grown cell lines. AB - The sequences and profiles of peptides which bind to HLA-B*2705 splenocytes and peripheral blood cells were compared with those previously published from in vitro long-term cell cultures. B*2705 peptide profile analysed by solid-phase Edman degradation and 15 individual peptide sequences determined by LC-MS/MS were partially similar to those defined from in vitro long-term cell cultures. Arg at P2 was found in 11 of 15 sequenced peptides (73.3%). This value is lower in comparison with other published data. Two sequences were matching to unknown proteins, which displayed similarity with myosin. These are first data on peptide sequences isolated directly from HLA-B27 molecules without prior in vitro propagation of the cells. PMID- 15275978 TI - Diagnosis and management of hyperbilirubinemia in the term neonate: for a safer first week. AB - New data support restructuring the approach toward diagnosis and management of hyperbilirubenia in the term neonate to make it more physician-friendly and gain wider implementation. The authors advocate clear criteria for patient safety, preventive approaches, and timely interventions. Structural changes to facilitate a system-based approach should include predischarge bilirubin management; follow up bilirubin management; and lactational support and nutritional management. The authors advocate total serum bilirubin screening and a scoring system based on clinical risk factors as predischarge screening strategies; we should screen all babies for hyperbilirubinemia and for targeted follow-up based on an hour specific total serum bilirubin measured for risk assessment. We should also provide focused universal education emphasizing adequate lactational nutrition, to decrease severe hyperbilirubinemia and thus prevent kernicterus. PMID- 15275979 TI - Skin lesions in the neonate. AB - Neonatal skin provides physical protection and assists in fluid balance,immunosurveillance, and thermoregulation; thus, playing a vital role in the newborn's transition from an aqueous to an air-dominant environment. Worried parents often seek medical attention from their child's physician regarding skin lesions. Thus,a working knowledge of both normal and abnormal cutaneous lesions of the neonate is required to properly address these issues. In this article, the authors briefly discuss transient benign lesions,pustular and vesicular infections, "birthmarks" (vascular and pigmentary lesions), common congenital abnormalities, select blistering disorders, and various other skin conditions. PMID- 15275980 TI - Herpesviridae infections in newborns: varicella zoster virus, herpes simplex virus, and cytomegalovirus. AB - Varicella zoster virus (VZV), herpes simplex virus (HSV) and cytomegalovirus (CMV) are all members of the Herpesviridae family.Humans are the only source of infection for these double stranded DNA viruses. Infants may acquire these infections in utero, peripartum, or postnatally, resulting in a variety of clinical syndromes, ranging from asymptomatic infection to severe infection,with high mortality rates and significant long-term morbidity. This article presents the epidemiology, clinical characteristics, treatment,and prevention strategies for VZV, HSV, and CMV infections in infants. PMID- 15275981 TI - Management of the infant born to a mother with HIV infection. AB - In countries with adequate resources, rates of perinatal mother-to-child transmission (MTCT) of HIV can be as low as 2% or lower. To achieve this low rate of MTCT of HIV requires identification of women with HIV infection early in pregnancy, treatment of the pregnant woman with appropriate combination antiretroviral therapy, special interventions in maternal management during labor and delivery, and appropriate care of the newborn infant. Although many of the steps in preventing HIV MTCT fall to obstetrical care providers, practitioners focused on care of the newborn also play an important role in the prevention of perinatal HIV MTCT, follow-up to identify or exclude HIV infection in the infant, and ongoing care for children and families affected by HIV. PMID- 15275982 TI - Diagnosis and management of bacterial infections in the neonate. AB - Perinatally acquired bacterial neonatal sepsis is a low-incidence,high-risk disease. Although incidence of the most common etiology,group B Streptococcus, has been reduced by prophylactic strategies,neonatal sepsis has not been eradicated, and vigilance must remain high. Accurate diagnosis is difficult: signs and symptoms are hard to distinguish from other causes of neonatal distress, and definitive diagnostic tests are not available. The clinician must make a judgment call, considering the perinatal history, the constellation of signs and symptoms, and the results of existing diagnostic tests,before neonatal sepsis can diagnosed or excluded. With diagnosis,knowledge of the specific disease states and clinical algorithms for management aid in formulating a plan of treatment with antimicrobial agents and supportive care. PMID- 15275983 TI - Neonatal seizures. AB - Neonatal seizures typically indicate significant underlying disease.They are poorly classified, under-recognized, and often difficult to treat. Recognition of etiology is often helpful in prognosis and treatment; the most common is hypoxic ischemic encephalopathy. Patients generally have a poor prognosis, with most developing a severe encephalopathy and epilepsy. Studies suggest that neonatal seizures and their etiology have a significant impact on the developing brain; it is critical to recognize seizures early and initiate immediate antiepileptic therapy. Continuous computerized simultaneous video electroencephalograph monitoring is imperative;at-risk infants will frequently have electrographic seizures without clinical manifestations. Although there are antiepileptic therapies for neonatal seizures, they are ineffective in over 35% of cases. The goal of research should be the development of more effective therapies for neonatal seizures, regardless of etiology. PMID- 15275984 TI - The neonate with an abdominal mass. AB - Abdominal masses in neonates reflect a wide spectrum of diseases,from lesions that can cause significant morbidity and mortality,to conditions readily corrected surgically, to entities which maybe safely observed. It is incumbent upon the infant's physician to determine the nature of the mass in a timely, safe, and cost-effective manner. PMID- 15275985 TI - An approach to diagnosis and management of cyanosis and tachypnea in term infants. AB - Tachypnea and cyanosis in the newborn are frequently encountered problems in the nursery. The incidence of respiratory distress ranges from 2.9% to 7.6%, and 4.3% of newborns may require supplemental oxygen therapy. In this article, the pathophysiology,approach to the diagnosis, and management of clinical conditions are discussed. The physiologic mechanisms involved in the pathogenesis of these problems and the clinical assessment and early stabilization of an infant before transport to a tertiary care center, if necessary, are also discussed. PMID- 15275986 TI - Differential diagnosis and approach to a heart murmur in term infants. AB - This article reviews the differential diagnosis of and approach to a heart murmur in the term infant. After addressing important aspects of the history, physical examination, and cardiac examination,the most common structural cardiac lesions generating a heart murmur in the term infant are discussed. PMID- 15275987 TI - Differential diagnosis and management of the fetus and newborn with an irregular or abnormal heart rate. AB - This article separately discusses the differential diagnosis and management of irregular or abnormal heart rate in both the fetus and the newborn. Conditions covered include ectopic beats, tachyarrythmias,atrial flutter, bradyarrythmias, tachycardia, congenital atrioventricular block, long QT syndrome, and bradycardias,among others. PMID- 15275988 TI - Urologic problems of the neonate. AB - Maternal sonography is integrated into routine prenatal care, and numerous fetal anomalies are detected, with genitourinary anomalies the most common. This has profoundly influenced the presentation and natural history of neonatal urologic problems. This article is divided into two sections. The first addresses clinical questions raised by prenatally detected anomalies, abdominal masses, urosepsis, urinary retention, scrotal masses, and abnormal external genitalia. The second discusses evaluation and management of specific anomalies,including controversies, and optimum timing for tertiary care center referral. The selected anomalies include renal anomalies, tumors, the exstrophy complex, urogenital anomalies, ambiguous genitalia, posterior urethral valves, scrotal anomalies, spina bifida, and common urologic syndromes such as the triad syndrome. Readers are referred to urologic texts for additional information. PMID- 15275989 TI - Differential diagnosis and management of polycythemia. AB - One percent to 5% of all newborns in the United States are polycythemic.As the venous hematocrit rises above 65%, the thickness or viscosity of whole blood also increases, potentially compromising blood flow to a variety of organs. Fortunately, relatively few infants who have neonatal polycythemia or hyperviscosity develop complications attributable to their thick blood; however, controversy and the need for continued research envelop the issue of which infants are at risk and need to be treated. This article reviews the differential diagnosis, clinical presentation, and treatment of neonatal polycythemia. PMID- 15275991 TI - Differential diagnosis and management of thrombocytopenia in childhood. AB - The purpose of this article is to provide the reader with a firm knowledge of the major causes of thrombocytopenia and their treatments, and to form a broad differential diagnosis, so that it will be clearer when to consider a rare etiology. The various etiologies are presented by known disease entities, grouped by age,and described as they would occur and be considered in a realistic clinical setting. A brief categorization of causes of thrombocytopenia by mechanism, notably abnormal platelet production, platelet destruction, or sequestration, is included. With each disease process, the pathophysiology as it is currently known is described and discussed. PMID- 15275990 TI - Differential diagnosis and management of anemia in the newborn. AB - Neonatal anemia is a condition with a diverse etiologic spectrum.Therefore, in order to form a focused differential diagnosis, it is important for the caregiver to have some knowledge of the more common causes of low hemoglobin and hematocrit concentrations in the neonate. Proper history taking, physical examination, and interpretation of diagnostic tests can narrow this focus and aid in establishing an accurate diagnosis and in directing the appropriate therapeutic interventions. PMID- 15275992 TI - Endocrine disorders in the neonate. AB - There is a significant amount of knowledge that has been gained in recent years in the study of endocrine disorders in the newborn. The explosion of genetic data shedding light on the origins of endocrine disease has expanded the level of diagnostic evaluation and management of these infants. This article provides a general review of endocrine disorders as they present in a newborn. PMID- 15275993 TI - Management of birth injuries. AB - Birth injuries are a significant cause of neonatal morbidity and mortality. Although they are frequently associated with traumatic delivery, birth injuries often occur in normal spontaneous deliveries in the absence of any risk factors. This article discusses the diagnosis and management of the most common birth injuries encountered by health care providers caring for newborns. PMID- 15275994 TI - Common issues and concerns in the newborn nursery, part II. PMID- 15275995 TI - A four-dimensional simulation model of tumour response to radiotherapy in vivo: parametric validation considering radiosensitivity, genetic profile and fractionation. AB - The aim of this paper is to present the current state of a four-dimensional simulation model of solid tumour growth and response to radiotherapy developed by our group. The most prominent points of the algorithms describing the fundamental biological phenomena involved are outlined. A specific application of the model to a selected clinical case of glioblastoma multiforme is described and comparative studies are performed, using various exploratory values of the model parameters. Qualitative agreement with clinical observations has been achieved. Special emphasis is laid on the variability of radiosensitivity parameters throughout the cell cycle and on the influence of the genetic profile of the tumour on its radiosensitivity. The results of the simulation are three dimensionally reconstructed. A valuable tool for getting insight into the biology of tumour growth and response to radiotherapy and at the same time an advanced patient specific decision support system is expected to emerge after the completion of the necessary extensive clinical evaluation. PMID- 15275996 TI - A cellular growth model for root tips. AB - Growth of the root tip is modeled using a one-dimensional string of cells. Each cell is characterized by three distinct phases, division, elongation-only or maturity. In this model two hypothetical phytohormones, one produced at the root tip and the other at the shoot, determine the behavior of the cell, and therefore the growth of the entire tip. While the division rate is taken to be a step function of the string coordinate, the growth rate of each cell is assumed to be piecewise linear and composed of linear functions of cell length. Thereafter, suitable operators for the calculation of the velocity and relative growth rate distributions are given. The results of the model are finally compared to measurements of Arabidopsis thaliana, Nicotiana tabacum and Pisum sativum roots. PMID- 15275997 TI - The prevalence of male homosexuality: the effect of fraternal birth order and variations in family size. AB - The prevalence of male homosexuality probably varies over time and across societies. One reason for this variation may be the joint effect of two factors: (1) variations in fertility rate or family size; and (2) the fraternal birth order effect, the finding that the odds of male homosexuality increases with each additional older brother. Because of these effects, the rate of male homosexuality may be relatively high (at least in terms of sexual attraction if not behavior) in societies that have a high fertility rate, but this rate has probably declined somewhat in some, particularly western, societies. Thus, even if accurately measured in one country at one time, the rate of male homosexuality is subject to change and is not generalizable over time or across societies. PMID- 15275998 TI - A note on the discrepancy between the predicted and observed speed of the propagated action potential in the squid giant axon. AB - The Hodgkin-Huxley model for the ionic currents in the membrane of the squid giant axon has become the standard model for the electrophysiological behaviour of many excitable cells. A strong test of the model predicted a travelling wave speed of 18.76 m/s for the propagated action potential in an axon with a reported speed of 21.2 m/s. This discrepancy between prediction and observation was considered satisfactory when the model was proposed 50 years ago, appears not to have been re-evaluated, but is unsatisfactory for a mature and important model. The separate and combined influences of measurement error and biological variability on the discrepancy between prediction and observation are quantified, as is the effect of using of a one-dimensional model to represent a three dimensional axon. The main tool in this investigation is the use of simulation to study the behaviour of the Hodgkin-Huxley membrane model. These studies show that measurement error in combination with biological variability cannot account for the discrepancy between prediction and observation. Also, calculation shows that the one-dimensional description of the behaviour of the axon is adequate. Further calculation shows that the travelling wave description of the propagated action potential is valid only for sufficiently long axons. In shorter axons the propagated action potential is predicted to travel faster than the travelling wave; consequently under suitable experimental conditions the discrepancy between prediction and observation may be negligible. PMID- 15275999 TI - Biomechanical analysis of Oligochaeta crawling. AB - Hydrostatic skeletons, such as that of Oligochaeta and Hirudinea, allow the locomotion of animals with soft segmented bodies. In this paper crawling of Oligochaeta, and in particular that of earthworm (Lumbricus terrestris), is analyzed from a biomechanical point of view, starting from the experimental kinematic description of deformations coupled with a simple friction model. The analysis is able to predict crawling velocity with an error of about 15% with respect to the experimental measured values. Also muscular stress during locomotion is evaluated and found to be compatible with biological values. PMID- 15276000 TI - Spread of two linked social norms on complex interaction networks. AB - In this paper, we study the spread of social norms, such as rules and customs that are components of human cultures. We consider the spread of two social norms, which are linked through individual behaviors. Spreading social norms depend not only on the social network structure, but also on the learning system. We consider four social network structures: (1) complete mixing, in which each individual interacts with the others at random, (2) lattice, in which each individual interacts with its neighbors with some probability and with the others at random, (3) power-law network, in which a few influential people have more social contacts than the others, and (4) random graph network, in which the number of contacts follows a Poisson distribution. Using the lattice model, we also investigate the effect of the small-world phenomenon on the dynamics of social norms. In our models, each individual learns a social norm by trial and error (individual learning) and also imitates the other's social norm (social learning). We investigate how social network structure and learning systems affect the spread of two linked social norms. Our main results are: (1) Social learning does not lead to coexistence of social norms. Individual learning produces coexistence, and the dynamics of coexistence depend on which social norms are learned individually. (2) Social norms spread fastest in the power-law network model, followed by the random graph model, the complete mixing model, the two-dimensional lattice model and the one-dimensional lattice. (3) We see a "small world effect" in the one-dimensional model, but not in two dimensions. PMID- 15276001 TI - Forest gap dynamics and the Ising model. AB - The vegetation height in forest ecosystems is spatially heterogeneous. Canopy gaps (sites with low vegetation) are formed by treefalls, and they recover to canopy sites (with high vegetation) either by growth of small trees or by branch extension of surrounding trees. The dynamics of canopy gaps have been studied using a spatial Markov chain with nearest neighbor interaction. (1) If the canopy recovery rate is constant and if the gap formation rate for a site increases exponentially with the number of neighboring gap sites, the equilibrium distribution is the same as the one generated by the Ising model in statistical mechanics. Here, we extend the equivalence to the situation in which both the gap formation and canopy recovery depend on the neighborhood, as shown in recent forest data. (2) We develop a statistical test of whether a given spatial pattern is a random sample from the Ising model. The test is based on the conditional probability of configurations on a partial lattice. We apply the method to vegetation height data from the Ogawa forest reserve, Japan, measured on a 5x5 m grid in 1976, 1981, 1986, and 1991. The spatial pattern of the original forest data deviates significantly from the Ising model. We examine whether a larger sampling distance or the removal of the effects of the topography can reduce this deviation. PMID- 15276002 TI - How and why are uniformly polarization-sensitive retinae subject to polarization related artefacts? Correction of some errors in the theory of polarization induced false colours. AB - If the photoreceptors of a colour vision system are polarization sensitive, the system detects polarization-induced false colours. Based on the functional similarities between polarization vision and colour vision, earlier it was believed that a uniformly polarization-sensitive (insect) retina (UPSR)-in which receptors of all spectral types have the same polarization sensitivity ratio and microvilli direction-cannot detect polarization-induced false colours. Here we show that, contrary to this belief, a colour vision based on a UPSR is subject to polarization-related artefacts, because both the degree and the angle of polarization of light reflected from natural surfaces depend on wavelength. Our second goal is to correct certain errors in the theory of polarizational false colours. The quantitative estimation of the influence of polarization sensitivity on colour vision was recently motivated by the suggestion that certain Papilio butterflies detect such false colours. The theoretical basis of this subject is to calculate the colour loci in the colour space of a visual system from the quantum catches of polarization-sensitive receptors of different spectral types. Horvath et al. (J. Exp. Biol. 205 (2002) 3281) gave the first exact mathematical and receptor-physiological derivation of formulae for these calculations. Here we prove that the two formulae given earlier by others are inappropriate or erroneous. This, however, does not influence the validity of the experimental data and the principal conclusions drawn about the colour vision and polarization sensitivity in Papilio butterflies. PMID- 15276003 TI - A predictive model of moment-angle characteristics in human skeletal muscle: application and validation in muscles across the ankle joint. AB - In the present work, a generic model for the prediction of moment-angle characteristics in individual human skeletal muscles is presented. The model's prediction is based on the equation M = V x Lo(-1)sigma c cos phi x d, where M, V, and Lo are the moment-generating potential of the muscle, the muscle volume and the optimal muscle fibre length, respectively, and sigma, phi and d are the stress-generating potential of the muscle fibres, their pennation angle and the tendon moment arm length, respectively, at any given joint angle. The input parameters V, Lo, sigma, phi and d can be measured or derived mechanistically. This eliminates the common problem of the necessity to estimate one or more of the input parameters in the model by fitting its outcome to experimental results often inappropriate for the function modelled. The model's output was validated by comparisons with the moment-angle characteristics of the gastrocnemius (GS) and tibialis anterior (TA) muscles in six men, determined experimentally using voluntary contractions at several combinations of ankle and knee joint angles for the GS muscle and electrical stimulation for the TA muscle. Although the model predicted realistically the pattern of moment-angle relationship in both muscles, it consistently overestimated the GS muscle M and consistently underestimated the TA muscle M, with the difference gradually increasing from dorsiflexion to plantarflexion in both cases. The average difference between predicted and measured M was 14% for the GS muscle and 10% for the TA muscle. Approximating the muscle fibres as a single sarcomere in both muscles and failing to achieve complete TA muscle activation by electrical stimulation may largely explain the differences between theory and experiment. PMID- 15276004 TI - Intra-membrane ligand diffusion and cell shape modulate juxtacrine patterning. AB - A key problem in developmental biology is how pattern and planar polarity are transmitted in epithelial structures. Examples include Drosophila neuronal differentiation, ommatidia formation in the compound eye, and wing hair polarization. A key component for the generation of such patterns is direct cell cell signalling by transmembrane ligands, called juxtacrine signalling. Previous models for this mode of communication have considered homogeneous distributions in the cell membrane, and the role of polarity has been largely ignored. In this paper we determine the role of inhomogeneous protein and receptor distributions in juxtacrine signalling. We explicitly include individual membrane segments, diffusive transport of proteins and receptors between these segments, and production terms with a combination of local and global responses to ligand binding. Our analysis shows that intra-membrane ligand transport is vital for the generation of long wavelength patterns. Moreover, with no ligand transport, there is no pattern formation for lateral induction, a process in which receptor activation up-regulates ligand production. Biased production of ligand also modulates patterning bifurcations and predicted wavelengths. In addition, biased ligand and receptor trafficking can lead to regular polarity across a lattice, in which each cell has the same orientation-directly analogous to patterns of hairs in the Drosophila wing. We confirm the trends in pattern wavelengths previously observed for patterns with cellular homogeneity-lateral inhibition tends to give short-range patterns, while lateral induction can give patterns with much longer wavelengths. Moreover, the original model can be recovered if intra-membrane bound receptor diffusion is included and rapid equilibriation between the sides is considered. Finally, we consider the role of irregular cell shapes and waves in such networks, including wave propagation past clones of non-signalling cells. PMID- 15276005 TI - On the cross-regulation of protein tyrosine phosphatases and receptor tyrosine kinases in intracellular signaling. AB - Intracellular signaling proteins are very often regulated by site-specific phosphorylation. For example, growth factor receptors in eukaryotic cells contain intrinsic tyrosine kinase activity and use inter- and intra-molecular interactions to recruit and orient potential protein substrates for phosphorylation. Equally important in determining the magnitude and kinetics of such a response is protein dephosphorylation, catalysed by phosphatase enzymes. A growing body of evidence indicates that certain protein tyrosine phosphatases (PTPs), like tyrosine kinases, are affected by intermolecular interactions that alter the specific activity or localization of their catalytic domains. Using a detailed kinetic modeling framework, we theoretically explore the regulation of PTPs through their association with receptor tyrosine kinases, as noted for the Src homology 2-domain-containing PTPs, SHP-1 and -2. Receptor-PTP binding, in turn, is expected to influence the phosphorylation pattern of those receptors and proteins they associate with, and we show how PTPs might serve to co- or counter regulate parallel pathways in a signaling network. PMID- 15276006 TI - Computational prediction of conserved operons and phylogenetic footprinting of transcription regulatory elements in the metal-reducing bacterial family Geobacteraceae. AB - Members of the family Geobacteraceae are an important group of microorganisms from the delta subdivision of Proteobacteria that couple the oxidation of organic compounds to metal reduction. In order to uncover transcription regulatory interactions in these organisms, we used computational methods to identify conserved operons and putative cis-regulatory transcription elements. We identified 26 putative operons with gene order and function conserved among two species of Geobacteraceae, Geobacter sulfurreducens and Geobacter metallireducens. Most of these operons were also conserved in Desulfovibrio vulgaris, an additional metal reducing organism from family Desulfovibrionaceae of the delta subdivision of Proteobacteria. The predicted conserved operons were investigated for the presence of transcription factor binding sites by two different methods, (i) comparison of non-coding regions in conserved operons, and (ii) neural network promoter prediction. Predicted motifs were screened to identify most likely transcription factor binding sites and ribosome-binding sites. We provide information on motifs in Geobacteraceae similar to known transcription factor binding sites in Escherichia coli, conserved motifs in other bacterial species, putative palindromic sites, and predicted ribosome-binding sites. These predictions will aid in further elucidation of regulatory networks of gene interactions in Geobacteraceae. PMID- 15276007 TI - Murine physiology: measuring the phenotype. AB - The phenotype is the observable and measurable characteristic (or biochemical trait) of an organism that results from the interaction of the environment with the expression of gene alleles (i.e. the genotype). Accurate phenotyping demands that findings are not missed, anticipated, or biased; a mixture of assays that characterize distinct, but interrelated aspects of the cardiovascular phenotype are often necessary to meet these requirements. Such stringent conditions are crucial because it is phenotypic analysis that ultimately determines the utility of genetically altered and spontaneous mutant mice for biomedical research. PMID- 15276008 TI - Scaffolding and docking proteins of the heart, an introduction. PMID- 15276009 TI - Scaffold proteins and assembly of multiprotein signaling complexes. AB - Intracellular signaling involves assembly and regulation of multiprotein complexes. These complexes are functional units of signal transduction and are a means by which protein networks carry out tasks within the cell. One mechanism to influence the components, the subcellular localization, and the activity of these complexes, involves scaffold proteins. Scaffold proteins facilitate signal transduction by tethering molecules together and serving as molecular backbones for signaling complex assembly. Recent studies, particularly in the field of signaling kinases, have considerably advanced our understanding of the role that scaffold proteins play within multiprotein complexes in cardiac and other cell types. PMID- 15276010 TI - Interaction of Gbetagamma with RACK1 and other WD40 repeat proteins. AB - Heterotrimeric G-proteins, composed of Galpha and Gbetagamma subunits, transmit numerous and diverse extracellular stimuli via a large family of heptahelical cell-surface receptors to various intracellular effector molecules. The Gbetagamma subunit plays a central role in G-protein signaling. The Gbeta subunit belongs to a large family of WD40 repeat proteins, which adopt a circular beta bladed propeller structure. This unique structural feature confers interactions of Gbetagamma with a variety of proteins to play diverse functions. Intriguingly, we recently found that Gbetagamma can interact with three other WD40 repeat proteins, receptor for activated C kinase 1 (RACK1), dynein intermediate chain-1A and a protein of unknown function. This raises the following questions: are interactions among WD40 proteins a common theme and does the formation of a WD40 WD40 repeat protein complex constitute a protein scaffold for integrating signals from different cellular processes. We are beginning to address these issues by studying the interaction between Gbetagamma and RACK1. Here we will describe the molecular mechanism underlying this interaction and the implications of the interaction on the signal transduction of G-protein and RACK1. PMID- 15276012 TI - Macromolecular complexes regulating cardiac ryanodine receptor function. AB - The cardiac ryanodine receptor (RyR) is the sarcoplasmic reticulum (SR) Ca release channel which is centrally involved in the myocyte excitation-contraction (E-C) coupling process and certain cardiac arrhythmias, and even contributes to pacemaker activity in the heart. The RyR is also the center of a massive macromolecular complex which includes numerous regulatory proteins which can modulate RyR function. This complex includes proteins that interact with the cytoplasmic part of the RyR directly or indirectly (e.g. calmodulin (CaM), FK-506 binding proteins, protein kinase A, Ca-CaM-dependent protein kinase, phosphatases 1 and 2A, mAKAP, spinophilin, PR130, sorcin, triadin, junctin, calsequestrin and Homer). Information is evolving in terms of understanding both the physical/molecular nature of the protein-protein interactions between RyR and these other proteins. There is also a slowly developing picture as to how this complex of proteins may be involved in the functional modulation of the RyR. This RyR complex exists in physical proximity to regulatory complexes associated with sarcolemmal Ca channels, which have some similar components. These complexes, and their relative independence emphasizes the importance of thinking about other aspects of very local molecular signaling, analogous to the local control of SR Ca-release at the heart of current (E-C) coupling theory. PMID- 15276011 TI - beta(2)-Adrenergic receptor signaling complexes in cardiomyocyte caveolae/lipid rafts. AB - The traditional notion that catecholamine actions are mediated by the predominant beta(1)-adrenergic receptor (beta(1)-AR) subtype linked to the activation of adenylyl cyclase and the accumulation of cyclic adenosine 3',5'-monophosphate (cAMP) in cardiomyocytes has been challenged by recent studies showing that cardiomyocytes co-express pharmacologically distinct beta(2)-AR subtypes that activate a more broad range of downstream effectors. While beta(1)- and beta(2) ARs exert largely functionally equivalent cellular actions in heterologous expression systems, signaling by endogenous beta-AR subtypes in highly differentiated cells such as cardiomyocytes can be strikingly different. There is growing evidence that certain features of the signaling phenotypes for beta-AR subtypes in cardiomyocytes are inconsistent with traditional models that attribute signaling specificity to high-affinity protein-protein interactions between receptors, G-proteins, and effectors freely mobile on surface membranes. This chapter summarizes recent studies that focus on membrane microdomains (such as caveolae or lipid rafts) as sites that differentially localizing individual beta-AR subtypes as well as the downstream signaling machinery that generates, propagates, and downregulates the cAMP-protein kinase A signaling pathway. To the extent that this mechanism calibrates beta-AR responses in cardiomyocytes, it would be expected to be pertinent to the pathogenesis of heart failure, where chronic/persistent beta-AR signaling contributes to ventricular remodeling and impacts on long-term survival. PMID- 15276013 TI - Heart failure: how important is cellular sequestration? The role of the renin angiotensin-aldosterone system. AB - Heart failure, which is classically described as a pathologic process characterized by a decline of heart contractility, involves a complex set of alterations like the release of inflammatory cytokines, endothelin, angiotensin II and aldosterone. These abnormalities cause appreciable changes at cellular and molecular levels with consequent impairment of cell coupling, impulse propagation as well as morphologic alterations. In the present review the question whether cellular sequestration elicited by impairment of cell coupling and interstitial fibrosis plays a role on the development of the disease is discussed and the role of the plasma and cardiac renin-angiotensin-aldosterone systems on the process of cellular sequestration is described. The possible role of an intracellular renin angiotensin system on intercellular signaling is also discussed. The beneficial role of the angiotensin-converting enzyme inhibitors is related to the improvement of cellular synchronization which seems related to different factors like increment of cell coupling, hyperpolarization of cell membrane and morphologic remodeling including a decrease of interstitial fibrosis and ventricular hypertrophy. PMID- 15276014 TI - Cardiac ion channel expression and regulation: the role of innervation. AB - The expression and function of numerous cardiac ion channels change with development and disease. Whereas multiple regulatory processes and molecular mechanisms are certainly involved, one factor, sympathetic innervation, contributes to many of the developmental changes and is suggested to play a role in pathology. The onset of cardiac sympathetic innervation of the mammalian ventricle during early post-natal life has been associated with functional alterations in several ionic currents, including Na(+), L-type Ca(2+), pacemaker, inward rectifier and transient outward K(+) currents. The neural signaling molecule is not the same in each case, with evidence pointing to contributions from sustained activation of myocardial neuropeptide Y receptors, alpha adrenergic receptors and beta-adrenergic receptors, as well as additional, but as yet unidentified, targets. Knowledge of the mechanisms by which innervation regulates ion channel expression and function during normal development may aid efforts to reverse remodel the diseased heart and to target pharmacologic agents to remodeled channels. PMID- 15276015 TI - The role of phosphoinositide-3 kinase and PTEN in cardiovascular physiology and disease. AB - Phosphoinositide-3 kinases (PI3Ks) are a family of evolutionary conserved lipid kinases that mediate many cellular responses in both physiologic and pathophysiologic states. Class I PI3K can be activated by either receptor tyrosine kinase (RTK)/cytokine receptor activation (class I(A)) or G-protein coupled receptors (GPCR) (class I(B)). Once activated PI3Ks generate phosphatidylinositols (PtdIns) (3,4,5)P(3) leading to the recruitment and activation of Akt/protein kinase B (PKB), PDK1 and monomeric G-proteins (e.g. Rac GTPases), which then activate a range of downstream targets including glycogen synthase kinase-3beta (GSK-3beta), mammalian target of rapamycin (mTOR), p70S6 kinase, endothelial nitric oxide synthase (eNOS) and several anti-apoptotic effectors. Class I(A) (PI3Kalpha, beta and delta) and class I(B) (PI3Kgamma) PI3Ks mediate distinct phenotypes in the heart and under negative control by the 3'-lipid phosphatase, phosphatase and tensin homolog on chromosome ten (PTEN) which dephosphorylate PtdIns(3,4,5)P(3) into PtdIns(4,5)P(2). PI3Kalpha, gamma and PTEN are expressed in cardiomyocytes, fibroblasts, endothelial cells and vascular smooth muscle cells where they modulate cell survival/apoptosis, hypertrophy, contractility, metabolism and mechanotransduction. Several transgenic and knockout models support a fundamental role of PI3K/PTEN signaling in the regulation of myocardial contractility and hypertrophy. Consequently the PI3K/PTEN signaling pathways are involved in a wide variety of diseases including cardiac hypertrophy, heart failure, preconditioning and hypertension. In this review, we discuss the biochemistry and molecular biology of PI3K (class I isoforms) and PTEN and their critical role in cardiovascular physiology and diseases. PMID- 15276016 TI - Ions and channels: new answers to old questions. PMID- 15276017 TI - Rapid upregulation of CTGF in cardiac myocytes by hypertrophic stimuli: implication for cardiac fibrosis and hypertrophy. PMID- 15276018 TI - Early reperfusion levels of Na(+) and Ca(2+) are strongly associated with postischemic functional recovery but are disassociated from K(ATP) channel induced cardioprotection. AB - We previously demonstrated that pinacidil does not affect Na(+)(i) accumulation, cellular energy depletion, or acidosis during myocardial ischemia, but dramatically improves the cationic/energetic status during reperfusion. We investigated the role of this latter effect in K(ATP) channel-induced cardioprotection. Employing (23)Na and (31)P nuclear magnetic resonance spectroscopy with perfused rat hearts, reperfusion Na(+)(i) was altered with brief infusions of ouabain and/or RbCl to transiently decrease or increase Na(+)/K(+) ATPase activity. The increases and decreases in functional recovery (%LVDP-R) with pinacidil or ouabain, respectively, were largely unaltered by each other's presence. Early reperfusion Na(+)(i) and cellular energy were greatly altered by ouabain and indicated linear relationships with %LVDP-R. Pinacidil shifted these relationships to higher %LVDP-R. Increasing early reperfusion Na(+)(i) decreased %LVDP-R but did not diminish pinacidil's capacity to improve %LVDP-R. Approximately 75% and 45% of the pinacidil-induced improvements in %LVDP R, could be disassociated from early reperfusion Na(+)(i) and cellular energy, respectively. Both pinacidil and RbCl infusion blunted ouabain's elevation of reperfusion Na(+)(i), but RbCl did not improve %LVDP-R. Atomic absorption tissue Ca(2+) measurements indicated that pinacidil reduced late reperfusion Ca(2+) uptake, but did not reduce early reperfusion Ca(2+), and its beneficial effects were resistant to ouabain-induced early reperfusion Ca(2+) increases. In conclusion, K(ATP) channel-induced cardioprotection does not require moderation of Na(+)(i) accumulation, cellular energy depletion, or acidosis during ischemia. K(ATP) channel-induced cardioprotection is largely independent of the accelerated reperfusion Na(+)(i) recovery it induces and does not require early reperfusion reductions of tissue Ca(2+). A larger role for early reperfusion cellular energy cannot be excluded. PMID- 15276019 TI - Pigment epithelium-derived factor inhibits TNF-alpha-induced interleukin-6 expression in endothelial cells by suppressing NADPH oxidase-mediated reactive oxygen species generation. AB - Pigment epithelium-derived factor (PEDF) has recently been shown to be involved in the pathogenesis of proliferative diabetic retinopathy. Atherosclerosis is an inflammatory-fibroproliferative disease as well. Oxidative stress plays a major role in retinopathy and atherosclerosis. Accordingly, we investigated effects of PEDF on reactive oxygen species (ROS) generation, NF-kappaB activation and interleukin (IL)-6 expression in TNF-alpha-exposed HUVEC. TNF-alpha significantly increased intracellular ROS generation, which was completely blocked by PEDF or diphenylene iodonium, an inhibitor of NADPH oxidase. Further, PEDF completely prevented the TNF-alpha-induced increase in NADPH oxidase activity. PEDF or an antioxidant, N-acetylcysteine, significantly inhibited the TNF-alpha-induced NF kappaB activation. PEDF inhibited TNF-alpha-induced expression of IL-6 at both mRNA and protein levels. Moreover, TNF-alpha downregulated PEDF mRNA levels. Ligand blot analysis revealed that HUVEC possessed a membrane protein with binding affinity for PEDF. The results demonstrated that PEDF inhibited TNF-alpha induced NF-kappaB activation and subsequent IL-6 overexpression in HUVEC by suppressing NADPH oxidase-mediated ROS generation. Our present study suggests that PEDF may play an important role in the development and progression of atherosclerosis. PMID- 15276020 TI - Gender modulation of Ca(2+) uptake in cardiac mitochondria. AB - BACKGROUND: Mitochondrial calcium overload is an important factor in defining ischemia/reperfusion injury. Since pre-menopausal women are relatively protected from ischemia and heart disease, we tested the hypothesis that gender differences alter Ca(2+) handling in rat cardiac mitochondria. METHODS: Using cardiac mitochondria isolated from male, female, and ovariectomized Sprague-Dawley rats, we measured mitochondrial calcium transport, redox state, and membrane potential (Deltapsi(m)) during exposure to a calcium bolus. Redox state was modulated using either succinate (S) or succinate and pyruvate (SP) as substrates. RESULTS: Net Ca(2+) uptake rates were significantly lower in female than male mitochondria using SP, substrate conditions that resulted in a lower redox state (NADH/NAD(+)). Inhibition of the mitochondrial transition pore (MTP) using cyclosporin A showed significantly lower net Ca(2+) uptake in both substrate solutions when mitochondria from female and ovariectomized animals were compared to males, a finding consistent with gender modulation of the mitochondrial uniporter. Blockade of the Ca(2+) uniporter by ruthenium red abolished gender or substrate solution differences in calcium release. While there were no significant differences in resting Deltapsi(m), or Deltapsi(m) following Ca(2+) addition, 80% of female samples recovered from Ca(2+)-induced depolarization compared to 57% and 43% of male and ovariectomized animals, respectively. CONCLUSIONS: Mitochondria from female hearts have lower Ca(2+) uptake rates under physiologic substrate solutions (succinate/pyruvate) and are able to appropriately maintain DeltaYm under conditions of high [Ca(2+)]. These differences are consistent with gender modulation of the Ca(2+) uniporter and may be a mechanism by which female myocardium suffers less injury with ischemia/reperfusion. PMID- 15276021 TI - Arginase pathway in human endothelial cells in pathophysiological conditions. AB - Objective. - Arginase is a nitric oxide synthase-alternative pathway for l arginine breakdown leading to biosynthesis of urea and l-ornithine. Arginase pathway is inducible by inflammatory molecules-such as cytokines and bacterial endotoxin-in macrophages and smooth muscle cells. The presence of an arginase pathway in human endothelial cells and its possible modulation by inflammation are unknown. Methods. - We have: (i) characterised arginase pathway in terms of activity, isoform type and gene expression in a primary human umbilical vein endothelial cells (HUVEC) line; (ii) evaluated arginase functional role in cell proliferation with the aid of l-norvaline, an arginase inhibitor and (iii) determined the effects of tumour necrosis factor-alpha and endotoxin on arginase pathway. Results. - HUVEC showed a baseline arginase activity and expression of both arginase isoforms (arginase I and II (A-I and A-II, respectively)) which resulted in l-norvaline-inhibitable cellular polyamine synthesis. The baseline arginase activity is important for HUVEC proliferation as cell cycle analysis and nuclear factor Ki-67 immunostaining revealed. Following incubation with inflammatory molecules, arginase activity increased but HUVEC cell cycling decreased. Conclusions. - A-I and A-II are constitutively expressed in HUVEC where they take part to the regulation of cell cycling. Although arginase activity is positively modulated by inflammatory molecules, it is insufficient to counteract the overall cell cycling inhibiting effects of inflammation. PMID- 15276022 TI - Salutary effects of attenuation of angiotensin II on coronary perivascular fibrosis associated with insulin resistance and obesity. AB - Obesity and insulin resistance confer increased risk for accelerated coronary disease and cardiomyopathic phenomena. We have previously shown that inhibition of angiotensin-converting enzyme (ACE) prevents coronary perimicrovascular fibrosis in genetically obese mice that develop insulin resistance. This study was performed to elucidate mechanism(s) implicated and to determine the effects of attenuation of angiotensin II (Ang) II. Genetically obese ob/ob mice were given ACE inhibitor (temocapril) or Ang II type 1 (AT(1)) receptor blocker (olmesartan) from 10 to 20 weeks. Cardiac expressions of plasminogen activator inhibitor (PAI)-1, the major physiologic inhibitor of fibrinolysis, and transforming growth factor (TGF)-beta(1), a prototypic profibrotic molecule, were determined and extent of perivascular coronary fibrosis was measured. Twenty-week old obese mice exhibited increased plasma levels of PAI-1 and TGF-beta(1) compared with the values in lean counterpart. Perivascular coronary fibrosis in arterioles and small arteries was evident in obese mice that also showed increased left ventricular collagen as measured by hydroxyproline assay. Immunohistochemistry confirmed the deposition of perivascular type 1 collagen. Markedly increased PAI-1 and TGF-beta were seen immunohistochemically in coronary vascular wall and confirmed by western blotting. When obese mice were treated with temocapril or olmesartan from 10 to 20 weeks, both were equally effective and prevented increases in perivascular fibrosis, plasma PAI-1 and TGF-beta(1), left ventricular collagen and mural immunoreactivity for PAI-1, TGF-beta and collagen type 1. The c-Jun NH(2)-terminal kinase (JNK) activity was elevated in the left ventricle of obese mice (western) and blocked by temocapril and olmesartan. Ang II-mediated upregulation of PAI-1 and TGF-beta(1) with collagen deposition may explain the mechanism of perivascular fibrosis in obese mice. ACE inhibition and blockade of AT(1) receptor may prevent coronary perivascular fibrosis and collagen deposition even before development of overt diabetes. JNK activation may be a mediator of obesity-related cardiac dysfunction and a potential therapeutic target. PMID- 15276023 TI - Inflammatory stimuli upregulate Rho-kinase in human coronary vascular smooth muscle cells. AB - Recent studies have demonstrated that upregulated Rho-kinase plays an important role in the pathogenesis of arteriosclerosis and vasospasm in both animals and humans. However, little is known about the molecular mechanism(s) involved in the Rho-kinase upregulation. Since inflammatory mechanisms have been implicated in the pathogenesis of arteriosclerosis and vasospasm, we examined whether inflammatory stimuli upregulate Rho-kinase in vitro and in vivo. In cultured human coronary vascular smooth muscle cells (hcVSMC), inflammatory stimuli, such as angiotensin II and interleukin-1beta, increased Rho-kinase expression (at both mRNA and protein levels) and function (as evaluated by the extent of the phosphorylation of the ERM (the ezrin/radixin/moesin) family, substrates of Rho kinase) in a time- and concentration-dependent manner. The expression of Rho kinase was inhibited by blockades of protein kinase C (PKC) (by either GF109253 or prolonged treatment with phorbol myristate acetate for 24 h) and an adenovirus mediated gene transfer of dominant-active Ikappa-B, suggesting an involvement of PKC and NF-kappaB in the intracellular signal transduction pathway for the Rho kinase expression. Furthermore, coronary vascular lesion formation (characterized by medial thickening and perivascular fibrosis) induced by a long-term administration of angiotensin II was markedly suppressed in NF-kappaB(-/-) mice with reduced expression and activity of Rho-kinase in vivo. These results indicate that the expression and function of Rho-kinase are upregulated by inflammatory stimuli (e.g. angiotensin II and IL-1beta) in hcVSMC with an involvement of PKC and NF-kappaB both in vitro and in vivo. PMID- 15276024 TI - Gender differences in hypertrophy, insulin resistance and ischemic injury in the aging type 2 diabetic rat heart. AB - Aging and diabetes in women increase their susceptibility to myocardial ischemic injury, but the cellular mechanisms involved are not understood. Consequently, we studied the influence of gender on cardiac insulin resistance and ischemic injury in the aging of Goto-Kakizaki (GK) rat, a model of type 2 diabetes. Male and female GK rats had heart/body weight ratios 29% (P < 0.0001) and 53% (P < 0.0001) higher, respectively, than their sex-matched controls, with the female GK rat hearts significantly more hypertrophied than the male (P < 0.001). Glucose transporter (GLUT) 1 protein levels were the same in all hearts, but GLUT4 protein levels were 28% lower (P < 0.01) in all GK rat hearts compared with their sex-matched controls. In isolated, perfused hearts, insulin-stimulated (3)H glucose uptake rates were decreased by 23% (P < 0.05) and 40% (P < 0.05) in male and female GK rat hearts, respectively, compared with their controls, with the female significantly more insulin resistant than the male GK rat hearts (P < 0.05). Protein kinase B protein levels and insulin-stimulated phosphorylation were the same in all hearts. During low-flow ischemia, glucose uptake was 59% lower (P < 0.001) in female, but the same as controls in male, GK rat hearts. Consequently, recovery of contractile function during reperfusion was 30% lower (P < 0.05) in female, but the same as controls in male GK rat hearts. We conclude that the aging female type 2 diabetic rat heart has increased insulin resistance and greater susceptibility to ischemic injury, than non-diabetic or male type 2 diabetic rat hearts. PMID- 15276025 TI - Blockade of macrophage migration inhibitory factor ameliorates experimental autoimmune myocarditis. AB - Macrophage migration inhibitory factor (MIF) is a cytokine that plays a critical role in the regulation of macrophage effector functions and T-cell activation. However, its role in the pathogenesis of experimental autoimmune myocarditis (EAM) has remained unresolved. In this study, we studied the role of the MIF in EAM. We investigated the expression of MIF in EAM using enzyme-linked immunosorbent assay, Northern blotting, and immunohistochemistry. Moreover, a neutralizing antibody (Ab) to MIF was injected intraperitoneally from day 0 to 20 (experiment 1), or from day 13 to 19 (experiment 2), after the immunization. Disease severity was estimated by the macroscopic and microscopic findings for the heart, heart weight to body weight ratio, and cellular and humoral immune responses on day 21. Enhanced MIF protein and mRNA expression in the heart tissue and an elevated serum MIF concentration were confirmed in EAM. In experiment 1, the anti-MIF Ab treatment markedly inhibited the onset of EAM. Moreover, a significant reduction in disease severity was also achieved even after the delayed anti-MIF Ab treatment in experiment 2. Furthermore, we demonstrated that MIF blockade decreased the expression of VCAM-1, TNF-alpha, and IL-1beta and the migration of T-cells and macrophages in the EAM heart. These results demonstrate an important role of MIF in the pathogenesis of EAM and suggest that MIF blockade may be a promising new strategy for the treatment of myocarditis. PMID- 15276026 TI - NHE-1 and NBC during pseudo-ischemia/reperfusion in rabbit ventricular myocytes. AB - Despite many studies into the pathophysiology of cardiac ischemia-reperfusion injury, a number of key details are as yet undisclosed. These include the timing and magnitude of the changes in both Na(+)/H(+) exchange (NHE-1) and Na(+) -- HCO(3)(-) -cotransport (NBC) transport rates. We fluorimetrically measured H(i)(+) fluxes (J(NHE-1) and J(NBC)) and Na(i)(+) fluxes in single contracting rabbit ventricular myocytes subjected to metabolic inhibition, pseudo-ischemia (i.e. metabolic inhibition and extracellular acidosis of 6.4), and pseudo reperfusion. Metabolic inhibition and pseudo-ischemia inhibited NHE-1 by 43 +/- 3.1% and 91 +/- 3.6%, and NBC by 66 +/- 5.4% and 100%, respectively. Inhibition was due to both an acidic shift of the pH(i) at which NHE-1 and NBC become quiescent (set-point pH(i)) and a reduction of the steepness of the pH(i) -- H(i)(+) flux profiles. NHE-1 and NBC did not contribute to Na(i)(+) loading during metabolic inhibition (Na(i)(+) 18 +/- 1.7 mM) or pseudo-ischemia (Na(i)(+) 21 +/- 1.7 mM), because pH(i) acidified less than set-point pH(i)'s. Upon pseudo reperfusion NBC recovered to 54 +/- 7.3% but NHE-1 to 193 +/- 11% of aerobic control flux, and set-point pH(i)'s returned to near neutral values. Metabolic inhibition and reperfusion caused an acid load of 18 +/- 3.2 mM H(+) 94% of which were extruded by the hyperactive NHE-1. At pseudo-reperfusion Na(i)(+) rose sharply to 31 +/- 5.8 mM within 1.5 min and that coincided with hypercontracture. Cariporide not only prevented the Na(i)(+) transient, but also inhibited pH(i) recovery and prevented hypercontracture. Our results are consistent with the view that NHE-1 is active during metabolic inhibition if, like in whole hearts, pH(i) is driven more acidic than NHE-1 set-point pH(i). Furthermore, either an acidic pH(i) or absence of additional Na(i)(+) loading during reperfusion, or both, limit ischemia-reperfusion injury. PMID- 15276027 TI - Protection from the effects of metabolic inhibition and reperfusion in contracting isolated ventricular myocytes via protein kinase C activation. AB - The protective effects of the PKC activator Phorbol 12-myristate 13-acetate (PMA) were investigated in electrically field stimulated (EFS) rat isolated ventricular myocytes following 7 min of metabolic inhibition induced by cyanide, iodoacetic acid and substrate removal, followed by reperfusion. PMA reduced reperfusion damage and increased functional recovery (response to EFS) following 10 min reperfusion from 20.0 +/- 10.7% of control myocytes to 90.0 +/- 7.2% following 5 min PMA pre-treatment (p<0.001). PMA significantly increased the time from the onset of MI before the myocytes failed to respond to EFS from 135 +/- 19s in control cells to 200 +/- 14s in PMA pre-treated cells (p<0.05). Additionally, there was an increase in the time to rigor with PMA pre-treated cells entering rigor 255 +/- 17s after MI compared to 174 +/-15s in control cell (p<0.05), indicating a delay in ATP depletion. During MI PMA pre-treated cells showed a significantly smaller increase in [Ca(2+)]i compared to control myocytes. Following reperfusion the majority of PMA pre-treated myocytes recovered calcium transients in response to EFS and diastolic Ca(2+) levels not significantly different to those seen prior to metabolic inhibition. Activation of PKC is thought to involve translocation to the particulate fraction. Our results demonstrate the presence of PKC-alpha, beta, gamma, delta, iota, lambda/zeta in rat ventricular myocytes, all of which translocate to the membrane in response to PMA. PMID- 15276028 TI - Loss-of-function mutations of the K(+) channel gene KCNJ2 constitute a rare cause of long QT syndrome. AB - Mutations of the KCNJ2 gene encoding the potassium channel Kir2.1 were previously shown to cause Andersen's syndrome (AS), a multisystem disease manifesting with developmental abnormalities, cardiac arrhythmias and periodic paralyses. We conducted a search for KCNJ2 mutations among 188 unrelated patients suspected to have long QT syndrome (LQTS). The screening was performed by denaturing high performance liquid chromatography (dHPLC) and DNA sequencing. Two novel mutations of the KCNJ2 gene were detected: a missense threonine to alanine mutation (T75A) in the N-terminal region (family 1) and an in-frame deletion of two amino acids (DeltaFQ163-164) in the M2 transmembrane region (family 2). In addition, a previously described silent polymorphism C1146T was detected. In family 1, some of the affected family members had a history of periodic muscle weakness characteristic of AS, but no dysmorphic features. The mean QTc interval of the affected members were 444 +/- 24 ms (family 1, n=7) and 456 +/- 8 ms (family 2, n=2). The mutations affect functionally important regions of the KCNJ2 channel protein: upon injection of the Xenopus oocytes with the wild type and mutant KCNJ2 constructs, the channel proteins were correctly synthesized and localized to the cell surface, but no measurable inward K(+) current could be detected for the mutant KCNJ2 constructs. In conclusion, we report two novel loss-of-function mutations of the KCNJ2 channel, affecting different domains of the channel protein. Mutations of the KCNJ2 gene should be considered in genetic subclassification of LQTS patients, even in the absence of overt manifestations of AS. PMID- 15276029 TI - Phenylephrine and endothelin-1 upregulate connective tissue growth factor in neonatal rat cardiac myocytes. AB - Cardiac hypertrophy is associated with hypertrophic growth of cardiac myocytes and increased fibrosis. Much is known of the stimuli which promote myocyte hypertrophy and the changes associated with the response, but the links between the two are largely unknown. Using subtractive hybridization, we identified three genes which are acutely (<1 h) upregulated in neonatal rat ventricular myocytes exposed to the alpha-adrenergic agonist, phenylephrine. One represented connective tissue growth factor (CTGF) which is implicated in fibrosis and promotes hypertrophy in other cells. We further examined the expression of CTGF mRNA and protein in cardiac myocytes using quantitative PCR and immunoblotting, confirming that phenylephrine increased CTGF mRNA (maximal within 1 h) and protein (increased over 4 - 24 h). Endothelin-1 promoted a greater, though transient, increase in CTGF mRNA, but the increase in CTGF protein was sustained over 8 h. Neither agonist increased CTGF mRNA in cardiac non-myocytes. By increasing the expression of CTGF in cardiac myocytes, hypertrophic agonists such as phenylephrine and endothelin-1 may promote fibrosis. CTGF may also propagate the hypertrophic response initiated by these agonists. PMID- 15276030 TI - Threonine-17 phosphorylation of phospholamban: a key determinant of frequency dependent increase of cardiac contractility. AB - Multiple studies have shown that phospholamban (PLN) plays a key role in regulation of frequency-dependent increase of cardiac contraction, a hallmark of the contractile reserve in myocardium. However, the mechanisms underlying this relationship remain elusive. Phosphorylation of PLN occurs on residues: serine-16 (Ser(16)) and threonine-17 (Thr(17)) in vivo. In isolated wild-type cardiomyocytes, we found that increases of stimulation frequency from 0.5 to 5 Hz were associated with increased Thr(17) phosphorylation of PLN and cardiac contractility. To further delineate the role of PLN phosphorylation in the frequency-dependent increases of cardiac function, three transgenic mouse models, expressing wild-type, Ser16Ala (S16A), or Thr17Ala (T17A) mutant PLN in the null background were generated. Transgenic lines expressing similar levels of wild type or mutant PLN were selected and isolated cardiomyocytes were paced from 0.5 to 5 Hz. Upon increases in pacing frequency, the fractional shortening (FS) and rates of contraction (+dL/dt) and relaxation (-dL/dt) increased in wild-type and S16A mutant PLN cardiomyocytes. In contrast, in myocytes expressing the T17A mutant PLN, there were no increases in FS and +/-dL/dt upon increasing the frequency of stimulation. The time to 50% peak shortening (TTP(50)) and to 50% relaxation (TTR(50)) were also abbreviated to a much higher extent (two-fold) in wild-type and S16A mutant compared to T17A mutant PLN cardiomyocytes. These results indicate that Thr(17) phosphorylation of PLN is the major contributor to frequency-dependent increases of contractile and relaxation parameters in mouse cardiomyocytes, although some increases in these parameters occur even in the absence of PLN phosphorylation. Thus, the positive force-frequency relationship in cardiomyocytes is mechanistically and mainly related to PLN phosphorylation. PMID- 15276031 TI - Adsorption of heavy metal ions on soils and soils constituents. AB - The article focuses on adsorption of heavy metal ions on soils and soils constituents such as clay minerals, metal (hydr)oxides, and soil organic matter. Empirical and mechanistic model approaches for heavy metal adsorption and parameter determination in such models have been reviewed. Sorption mechanisms in soils, the influence of surface functional groups and surface complexation as well as parameters influencing adsorption are discussed. The individual adsorption behavior of Cd, Cr, Pb, Cu, Mn, Zn and Co on soils and soil constituents is reviewed. PMID- 15276032 TI - Adsorption of Co(II), Ni(II), Cu(II), and Zn(II) on hexagonal templated zirconia obtained thorough a sol-gel process: the effects of nanostructure on adsorption features. AB - Using zirconium tetrabutoxide, diaminedecane, and diamineoctane as precursors, a templated hexagonal zirconia matrix is synthesized and characterized by X-ray diffractometry and scanning electron microscopy. The adsorption capacity of such a matrix toward Co(II), Ni(II), Cu(II), and Zn(II) from aqueous solutions is studied. The adsorption affinity of the synthesized hexagonal templated zirconia toward the cations is Cu(II)>Zn(II) >>Ni(II)>Co(II). It is also verified that the adsorption of the cations follows a Langmuir and not a Freundlich isotherm. All obtained isotherms are of type I, according to the IUPAC classification. The observed adsorption affinity sequence can be explained by taking into account the velocity constant for the substitution of water molecules into the cation coordination spheres, as well as the Irving-Williams series. PMID- 15276033 TI - ToF-SIMS depth profiling analysis of the uptake of Ba2+ and Co2+ ions by natural kaolinite clay. AB - The sorption behavior of Ba(2+) and Co(2+) ions on a natural clay sample rich in kaolinite was studied using time-of-flight secondary ion mass spectrometry (ToF SIMS). Depth profiling at 10-A steps was performed up to a 70-A matrix depth of the clay prior to and following sorption. The results showed that Co(2+) is sorbed in slightly larger quantities than Ba(2+), with significant numbers of ions fixed on the outermost surface of the clay. Depletion of the ions K(+), Mg(2+), and Ca(2+) from the clay lattice was observed to accompany enrichment with Co(2+) and Ba(2+) ions. The data obtained using X-ray powder diffraction (XRPD) and scanning electron microscopy (SEM) indicated insignificant structural and morphological changes in the lattice of the clay upon sorption of both Ba(2+) and Co(2+) ions. Analysis using energy dispersive X-ray spectroscopy (EDS) showed that the average atomic percentage (+/-S.D.) of Ba and Co on kaolinite surface were 0.49 +/- 0.11 and 0.61 +/- 0.19 , respectively, indicating a limited uptake capacity of natural kaolinite for both ions. PMID- 15276034 TI - Adsorption of naphthalene on zeolite from aqueous solution. AB - Polynuclear aromatic hydrocarbons (PAHs), which are environmental hormones and carcinogens, are viewed as the priority pollutants to deal with by many countries. Most PAHs are hydrophobic with high boiling and melting points and high electrochemical stability, but with low water solubility. Compared with other PAH species, naphthalene has less toxicity and is easily found in the environment. Thus, naphthalene is usually adopted as a model compound to examine the environmental and health aspects of PAHs. This study attempted to use an adsorption process to remove naphthalene from a water environment. The adsorption equilibrium of naphthalene on zeolite from water-butanol solution, which is a surfactant-enriched scrubbing liquid, was successfully evaluated by Langmuir, Freundlich, and linear isotherms. Among the tested kinetics models in this study (e.g., pseudo-first-order, pseudo-second-order, and Elovich rate equations), the pseudo-second-order equation successfully predicted the adsorption. PMID- 15276035 TI - Nitrogen adsorption characterization of aligned multiwalled carbon nanotubes and their acid modification. AB - Aligned multiwalled carbon nanotube (CNT) arrays were synthesized by using an iron-based sol-gel catalyst and acetylene as the precursor. These CNTs show high purity, uniform diameters and pore-wall thickness. Low temperature nitrogen adsorption was employed to characterize the structural and surface properties of the as-synthesized sample and that modified with boiling concentrated nitric acid. The adsorption characteristics of the as-synthesized and modified CNTs were thoroughly investigated. High-resolution comparative alpha(s)-plot showed that the nitrogen adsorption on CNTs takes place via a multistage mechanism closely related to their structures. It was also found that the acid modification significantly increased the adsorption energy and enhanced the adsorption capacity under low pressures. High-resolution comparative method provided valuable insights about the surface and pore structures of CNTs. PMID- 15276036 TI - Comparative adsorption studies of indigo carmine dye on chitin and chitosan. AB - The adsorption of indigo carmine dye onto chitin and chitosan from aqueous solutions was followed in a batch system. The ability of these materials to adsorb indigo carmine dye from aqueous solution was followed through a series of adsorption isotherms adjusted to a modified Langmuir equation. The maximum number of moles adsorbed was 1.24 +/- 0.16 x 10(-5) and 1.54 +/- 0.03 x 10(-4) mol g(-1) for chitin and chitosan, respectively. The same interactions were calorimetrically followed and the thermodynamic data showed exothermic enthalpic values of -40.12 +/- 3.52 and -29.25 +/- 1.93 kJ mol(-1) for chitin and chitosan, respectively. Gibbs free energies for the two adsorption processes of indigo carmine dye presented a positive value for chitin and a negative one for chitosan, reflecting that dye/surface interactions are thermodynamic favorable for chitosan and nonspontaneous for chitin at 298.15 K. The interaction processes were accompanied by an increase of entropy value for chitosan (90 +/- 6 J mol( 1)K(-1)) and a decrease for chitin (-145 +/- 13 J mol(-1)K(-1)). Thus, dye/chitosan interaction showed favorable enthalpic and entropic processes, reflecting thermodynamic stability of the formed complex, while dye/chitin interaction showed an exothermic enthalpic value and a highly nonfavorable entropic effect, resulting in a nonspontaneous thermodynamic system. PMID- 15276037 TI - Studies on manganese-nodule leached residues; 1. Physicochemical characterization and its adsorption behavior toward Ni(2+) in aqueous system. AB - Physicochemical characterization of manganese-nodule leached residues was carried out by chemical analyses, XRD, TG-DTA, surface area measurement, and FTIR techniques. The material is very fine-grained (<75 microm), is cryptocrystalline to amorphous in nature, and contains mainly of delta-MnO(2), quartz (alpha SiO(2)), and zeolite/feldspar minerals. Physically adsorbed sulfates in the leached residue are removed by repeated water washing and the washed sample shows an appreciable increase in surface area. This is indicated by the absence of 1387 and 1099 cm(-1) peaks in the IR spectrum of the washed sample. The adsorption behavior of the washed sample toward Ni(2+) was recorded as a function of time, pH, temperature, and concentrations of adsorbent and adsorbate. PMID- 15276038 TI - Modeling the adsorption and precipitation processes of Cu(II) on humin. AB - Humins (HU) are the most insoluble fraction of humic substances. Chemically, they can be considered as humic macromolecules bonded to the mineral matter of soil. The HU have a marked colloidal character and they are extremely important in retention of pollutants in soils. The aim of this work is to combine adsorption data with spectroscopic techniques in order to study the adsorption and precipitation processes of Cu(II) on HU. Analysis of sorption isotherms by means of several single-adsorption-process-based models makes it possible to obtain the speciation diagrams of Cu(II) species on HU surfaces. Further, FTIR (which provides information about the changes in the surface groups of the HU) and DRX (which shows the formation of possible crystalline phases on the HU surface) were used to determine the specific interactions of Cu(II) cations with the surface reactive groups of HU. The shape of the isotherms at constant pH varies with pH from L1-type (pH 2-4) to L3-type (pH 5-6) and S-type (pH 8), which indicates a change in the retention mechanism. When pH is 2 the retention of Cu(II), as [Cu(H(2)O)(6)](2+), is the preferred retention mechanism. The retained quantity of Cu(II) as [Cu(OH)(H(2)O)(5)](+) increases with pH. Starting from pH 4 the Cu(II) begins its precipitation, which is the preferred retention mechanism at pH 8. The presence of HU has a great influence on the precipitation process of Cu(II), giving rise to botalackite, which reveals epitaxial growth of crystals. PMID- 15276039 TI - Interaction forces between polyethylene oxide-polypropylene oxide ABA copolymers adsorbed to hydrophobic surfaces. AB - Block and graft copolymers are frequently used as stabilizing agents in colloidal dispersions. One common material is the range of polymers known as "Pluronics," which is a BASF trade name for ABA block copolymers composed of a propylene oxide anchoring block (B block) and two ethylene oxide buoy or stabilizing blocks (A block); the equivalent ICI (Uneqima) trade name is Synperonic. In the work presented here the interactions between adsorbed layers of these materials immersed in 10(-2) M sodium sulfate solutions are presented. The block copolymers investigated had an approximately fixed molecular weight of around 3250 Da for the anchoring B block, whilst the molecular weight of the stabilizing polyethylene oxide chains varies around 800-6500 Da. Hydrophobic glass surfaces were used as the test substrate. It was found that in the absence of polymer a long ranged attractive interaction is observed, typical for the interaction between hydrophobic surfaces in aqueous media, but that in the presence of the polymers a repulsion was observed. The repulsion became longer ranged as the molecular weight of the ethylene oxide chain increased. On separation of the surfaces, the interaction was slightly longer ranged, suggesting that the two polymer layers intertwine and stretch each other on separation. This effect was more noticeable for the higher molecular weight polymers. The compression data were well described using a scaling analysis for the interaction between polymer brushes. PMID- 15276040 TI - Fluorescent labels to study thermal transitions in epoxy/silica composites. AB - The analysis of the fluorescent response from the dansyl moiety as a function of temperature has been used to estimate the thermal transitions in silica particle/epoxy-based composite materials. Silica particles were surface-coated with 3-aminopropyltriethoxysilane (APTES) and 3-aminopropylmethyldiethoxysilane (APDES). 5-Dimethylamino-1-naphtalenesulfonyl chloride (DNS) and 5 dimethylaminonaphthalene-1-(2-aminoehyl)) sulfonamide (DNS-EDA) were selectively attached to the silanized silica particles and to the epoxy matrix respectively. The fluorescence results, interpreted in terms of the model of interpenetrating polymer networks, suggest that (i) independent of the silane coating, the interfacial region is slightly more rigid and heterogeneous than the epoxy bulk and (ii) the interface generated with APTES seems to be more flexible than that obtained with APDES. PMID- 15276041 TI - Investigation of surface properties of amino acids: polarity scale for amino acids as a means to predict surface exposed residues in films of proteins. AB - It is of great interest and importance to study how different amino acid residues contribute to and affect the properties of proteins coated as films on solid surface. This work shows that the solid/liquid interfacial energy of surface localized amino acid films and their Gibbs energies of transfer at the air/solution interface have the potential to be used as a rapid and simple method for studying the surface properties of proteins. Based on these results, a new polarity scale for amino acids has been proposed. This scale is compared with existing hydropathy scales in a benchmark test using some proteins with solved 3D structure. The proteins were characterized in terms of surface-exposed residues with a computer program, Graphical Representation and Analysis of Surface Properties (GRASP). It was also shown that each amino acid contribution is relative to the total protein surface and the other residues on the surface. PMID- 15276042 TI - Time-resolved SAXS study of the effect of a double hydrophilic block-copolymer on the formation of CaCO3 from a supersaturated salt solution. AB - The effect of a double hydrophilic block-copolymer additive (made of polyaspartic acid and polyethyleneglycol, pAsp(10)-b-PEG(110)) on the initial formation of calcium carbonate from a supersaturated salt solution has been studied in situ by means of time-resolved synchrotron small-angle X-ray scattering (SAXS). A stopped flow cell was used for rapidly mixing the 20 mM aqueous reactant solutions of calcium chloride and sodium carbonate. In reference measurements without polymer additive the very rapid formation of primary, overall spherical CaCO(3) particles with a radius of ca. 19 nm and a size polydispersity of ca. 26% was observed within the first 10 ms after mixing. A subsequent, very rapid aggregation of these primary particles was evidenced by a distinct upturn of the SAXS intensity at smallest angles. During the aggregation process the size of the primary particles remained unchanged. From an analysis of the absolute scattering intensity the mass density of these particles was determined to 1.9 g/cm(3). From this rather low density it is concluded that those precursor particles are amorphous, which has been confirmed by simultaneous wide-angle X-ray diffraction measurements. Upon adding 200 pm of the block-copolymer no influence on the size, the size polydispersity and morphology of the primary particles, nor on the kinetics of their formation and growth, was found. On the other hand, the subsequent aggregation and precipitation process is considerably slowed down by the additive and smaller aggregates result. The crystalline morphology of the sediment was studied in situ by WAXS ca. 50 min after mixing the reactants. Several diffraction rings could be detected, which indicate that a transformation of the metastable, amorphous precursor particles to randomly oriented vaterite nanocrystallites has taken place. In addition, a few isolated Bragg spots of high intensity were detected, which are attributed to individual, oriented calcite microcrystals that nucleated at the wall of the capillary. PMID- 15276043 TI - A highly efficient phase transfer method for preparing alkylamine-stabilized Ru, Pt, and Au nanoparticles. AB - A highly efficient phase-transfer method was developed to prepare alkylamine stabilized nanoparticles of several noble metals. This method involved first mixing the metal hydrosols and an ethanol solution of dodecylamine and then extracting the dodecylamine-stabilized metal nanoparticles into toluene. The efficiency of this phase-transfer method was nearly 100%. Alkylamine-stabilized Ru, Pt, and Au nanoparticles 3.45, 4.33, and 7.89 nm in diameter, respectively, could be prepared this way. The self-assembly of dodecylamine-stabilized Pt and Au particles was also detected by transmission electron microscopy (TEM). PMID- 15276044 TI - A novel one-step chemical method for preparation of copper nanofluids. AB - This paper presents a novel one-step method for preparing of copper nanofluids by reducing CuSO(4).5H(2)O with NaH(2)PO(2).H(2)O in ethylene glycol under microwave irradiation. Nonagglomerated and stably suspended Cu nanofluids are obtained. The influences of CuSO(4) concentration, addition of NaH(2)PO(2), and microwave irradiation on the reaction rate and the properties of Cu nanofluids were investigated by transmission electron microscopy, infrared analysis, and sedimentation measurements. It is found to be a fast, efficient one-step chemical method to prepare Cu nanofluids. PMID- 15276045 TI - Microwave synthesis and characterization of Co-ferrite nanoparticles. AB - Stable CoFe(2)O(4) nanoparticles have been obtained by co-precipitation using a microwave heating system. Transmission electron microscopy images analysis shows an agglomeration of particles with an average size of about 5 nm, and X-ray diffraction reveals the presence of a pure ferrite nanocrystalline phase. X-ray photoelectron spectroscopy and thermal gravimetric analysis show the presence of organic matter in the range of about 16 wt%. The magnetic response in DC fields is typical for an assembly of single-domain particles. The measured saturation magnetization is slightly larger than the bulk value, probably due to the presence of small amounts of Co and Fe. AC magnetization data indicate the presence of magnetic interactions between the nanoparticles. PMID- 15276046 TI - Temperature-induced gelation of concentrated silicon carbide suspensions. AB - Due to the steric barrier provided by the adsorption of the dispersant hypermer KD1 (a polyester/polyamine condensation polymer), stable and low-viscosity suspensions of SiC, Y(2)O(3), and Al(2)O(3) powder mixtures could be prepared in methyl ethyl ketone (MEK)/ethanol (E) solvent with solids loading as high as 60 vol%. The solvency of the dispersant in MEK/E decreased dramatically on cooling. Steady shear viscosity and oscillatory measurements were performed as a function of temperature for suspensions with different solids loading. The viscosity and elastic modulus of suspension increased with decreasing temperature and became more sensitive with the increase of solids loading. The suspensions with solids loading higher than 40 vol% could be solidified with decreasing temperature, but gelation temperature and gelation stiffness decreased with decreasing solids loading. The 60 vol% solid-loaded suspension was a stable and free-flowing fluid at 20 degrees C and gradually transformed to a very highly viscous and elastic system upon cooling to about 13 degrees C. Complete solidification occurred when the temperature was decreased to 5 degrees C. The gelation mechanism was mainly based on the collapse of the adsorbed layer as the temperature decreases, which induced incipient flocculation and formed a stiff network. The gelled body was further strengthened by separation of the dispersant from the suspension. PMID- 15276047 TI - Structure of organoclays--an X-ray diffraction and thermogravimetric analysis study. AB - X-ray diffraction has been used to study the changes in the surface properties of a montmorillonitic clay through the changes in the basal spacings of montmorillonite (SWy-2) and surfactant-intercalated organoclays. Variation in the d-spacing was found to be a step function of the surfactant concentration. High resolution thermogravimetric analysis (HRTG) shows that the thermal decomposition of SWy-2-MMTs modified with the surfactant octadecyltrimethylammonium bromide takes place in four steps. A mass-loss step is observed at room temperature and is attributed to dehydration of adsorption water. A second mass-loss step is observed over the temperature range 87.9 to 135.5 degrees C and is also attributed to dehydration of water hydrating metal cations such as Na+. The third mass loss occurs from 178.9 to 384.5 degrees C and is assigned to a loss of surfactant. The fourth mass-loss step is ascribed to the loss of OH units through dehydroxylation over the temperature range 556.0 to 636.4 degrees C. A model is proposed in which, up to 0.4 CEC, a surfactant monolayer is formed between the montmorillonitic clay layers; up to 0.8 CEC, a lateral-bilayer arrangement is formed; and above 1.5 CEC, a pseudotrimolecular layer is formed, with excess surfactant adsorbed on the clay surface. PMID- 15276048 TI - Concentration-dependent surface-enhanced Raman scattering of 2-benzoylpyridine adsorbed on colloidal silver particles. AB - Surface-enhanced Raman scattering (SERS) of 2-benzoylpyridine (2-BP) adsorbed on silver hydrosols has been investigated. It has been observed that with a small change in the adsorbate concentration, the SER spectra of 2-BP show significant change in their features, indicating different orientational changes of the different part of the flexible molecule on the colloidal silver surface with adsorbate concentration. The time dependence of the SER spectra of the molecule has been explained in terms of aggregation of colloidal silver particles and co adsorption and replacement kinetics of the adsorbed solute and solvent molecules on the silver surface. The broad long-wavelength band in the absorption spectra of the silver sol due to solute-induced coagulation of colloidal silver particles is found to be red-shifted with the increase in adsorbate concentration. The surface-enhanced Raman excitation profiles indicate that the resonance of the Raman excitation radiation with the new aggregation band contributes more to the SERS intensity than that with the original sol band. PMID- 15276049 TI - Effect of surface and interlayer structure on the fluorescence of rhodamine B montmorillonite: modeling and experiment. AB - The surface and interlayer structure of rhodamine B (RhB)-montmorillonite for various guest concentrations has been studied using a combination of X-ray powder diffraction and molecular modeling (molecular mechanics and molecular dynamics) in the Cerius(2) modeling environment. The joint effect of surface and interlayer structure on the fluorescence spectrum has been observed and discussed in relation to the position and orientation of RhB(+) cations with respect to the silicate layer. Structural analysis showed that the surface and interlayer structures are different as to the arrangement of RhB(+) cations, and both of them strongly depend on the guest concentration in the intercalation solution and on the method of preparation. The repeated intercalation of montmorillonite by rhodamine B used in the present work allowed obtaining RhB-montmorillonite in the maximum degree of ion exchange for every sample. PMID- 15276050 TI - Encapsulation of Fe(III) and Cu(II) complexes in NaY zeolite. AB - The use of nonporphyrin complexes encapsulated in zeolites as catalysts for oxidation reactions has been improved in the past decades by the discovery of increasing numbers of nonheme monoxygenases. The zeolite lattice can change the oxidative chemistry of the metallocomplexes, resulting in a catalytic effect different from those observed in homogeneous reactions. We report the encapsulation of iron and copper metallocomplexes with the ligand (2 hydroxybenzyl)(2-methylpyridyl)amine, Hbpa, and iron complexes with the ligand N,N'-bis(2-hydroxybenzyl)-N,N'-bis(2-methylpyridyl) ethylenediamine, H(2)bbpen. The zeolite-encapsulated metallocomplexes were prepared by diffusion of the ligands through the pores of the zeolites, already exchanged with the respective metal. The syntheses were performed in methanol and toluene solutions. Elemental analysis of solids with the Hbpa ligand have indicated better complexation for synthesis in toluene, where 74% of the iron atoms were coordinated by the ligand, against 37% for the synthesis in methanol. For the immobilization with the H(2)bbpen ligand in toluene it was observed that 46% of the iron atoms are coordinated, showing that the diffusion of the small ligand Hbpa through the zeolite cage was facilitated. The EPR spectra of the solids show signals at g = 2.0, which was attributed to an Fe-Fe interaction from the noncoordinated atoms, and g = 4.3 attributed to iron (III) in a rhombic geometry. PMID- 15276051 TI - Silylation and surface properties of chemically grafted hydrophobic silica. AB - A commercial mesoporous silica (Grace Davison) was chemically grafted with trimethylsilyl chloride (TMSCl) and hexamethyldisilanaze (HMDS). The silylation process brought about some reduction in the specific BET area, the pore volume, and the pore sizes of the samples. Thermogravimetric studies of the silylated samples revealed that the grafting process is kinetically controlled at short reaction times. In the kinetic regime, increasing concentrations of the silylant agent up to 2 wt% in the solvent led to an increase of the extent of the silylated surface, although this limitation disappeared at higher concentrations. Silylation was confirmed by diffuse reflectance infrared Fourier transform (DRIFTS), (29)Si CP-MAS NMR, and photoelectron (XPS) spectroscopic techniques. Solid-state (29)Si MAS-NMR spectra of the silylated samples revealed the presence of -SiCH(3) groups (9.5 ppm) together with two resonances, Q3 (approximately equal to -104 ppm) and Q4 (approximately equal to -114 ppm), coming from siloxane [Qn approximately Si(OSi)n(OH)(4-n), n approximately 2-4] groups, the Q3 signal decreasing upon silylation. The DRIFT spectra of the silylated samples exhibited two well defined bands at 2970 and 2907 cm(-1), due to stretching vibration modes of the C-H bonds in surface -CH(3) groups formed during the silylation process, and also the disappearance of the band at 3740 cm(-1). This observation indicates the complete removal of terminal and geminal hydroxyl groups by grafting with the silylating agent. Similarly, high-resolution photoelectron spectra of the Si2p core levels showed a high binding-energy component (103.5 eV) in all the samples, coming from the Si coordinated with oxide anions in SiO(2), together with a second component at 102.1 eV, which is the fingerprint of Si coordinated by oxide anions and an organic group. Finally, the samples were ranked according to their hydrophobicity, as determined from the temperature-programmed desorption profiles of adsorbed water and 2-methylbutane. PMID- 15276052 TI - Structure analysis of intercalated layer silicates: combination of molecular simulations and experiment. AB - Na(+)-montmorillonite type Wyoming, cloisite Na(+) from Southern Clay Products, Inc., was intercalated (i) with octadecylammonium cations and subsequently intercalated with octadecylamine molecules, (ii) with dodecylamine molecules, and (iii) with octylamine molecules to determine the applicability of these intercalates for nanocomposite materials on the base of polymer/clay. The structures were determined on the basis of a combination of results from X-ray diffraction and molecular simulations. The calculated values of basal spacings are in good agreement with experimental basal spacings when experimental samples were prepared. The interlayer space of intercalated montmorillonite shows a monolayer or bilayer arrangement of alkyl chains in dependence on the concentration of the intercalation solution. The values of the total sublimation energy, interaction energy, and exfoliation energy were calculated for all investigated samples. Low values of exfoliation energies lead to better exfoliation of intercalated silicate layers and this material appears suitable for use as a precursor for polymer/clay nanocomposites. The values of exfoliation energy for the investigated samples show that montmorillonite intercalated with dodecylamine or octadecylamine molecules is suitable for exfoliation of silicate layers. PMID- 15276053 TI - Preparation and electrochemical characterization of cation- and anion exchange/polyaniline composite membranes. AB - Composite membranes were prepared by chemical polymerization of a thin layer of polyaniline (PANI) in the presence of a high oxidant concentration on a single face of a sulfonated cation-exchange membrane (CEM) and quaternary aminated anion exchange membrane (AEM). IR and SEM studies for both types of membranes confirmed PANI loading on the ion-exchange membranes. PANI composite ion-exchange membranes were characterized as a function of the polymerization time by ion-exchange capacity, coating density, and membrane conductance measurements. Membrane potential measurements were performed in various electrolyte solutions in order to observe the selectivity of these membranes for different types of counterions. Membrane potential data in conjunction with membrane conductance data was interpreted on the basis of frictional considerations between membrane matrix and solute. Electrodialysis experiments, using PANI composite ion-exchange membranes with 4 h polymerization time, were performed in single and mixed electrolyte solutions for observing electromigration of solute across PANI composite ion exchange membranes. Relative dialytic rates of Na(2)SO(4), CaCl(2), and CuCl(2) were estimated with reference to NaCl on the basis of electrodialysis experiments and it was concluded that it is possible to separate different electrolytes using PANI composite ion-exchange membranes. PMID- 15276054 TI - Micropatterned self-assembled film based on temperature-responsive poly(N isopropylacrylamide-co-acrylic acid). AB - Micropatterned cross-linked film making up a temperature-responsive component has been fabricated through the following two steps: layer-by-layer electrostatic assembly of photosensitive nitrodiazoresin (NDR) and a thermosensitive copolymer of poly(N-isopropylacrylamide-co-acrylic acid) (P(NIPA-AA)), and subsequent selective exposure to UV light through a photomask followed with development in sodium dodecyl sulfate (SDS) aqueous solution. The irradiated regions of the film are retained due to the formed covalently linked structure, whereas the unirradiated parts of the film are removed fully from the substrate in SDS solution. The well-defined micropatterns were characterized with field-emission scanning electron spectroscopy (FE-SEM) and atomic force microscopy (AFM). PMID- 15276055 TI - Simultaneous electroosmotic and permeation flows through a Nafion membrane. 1. Aqueous electrolyte solutions. AB - The volume flow through a Nafion membrane originated by the simultaneous action of an electric potential difference and a pressure difference has been measured using aqueous KCl solutions under different experimental conditions. The behavior has been analyzed when both gradients act in the same and in the opposite sense. The results indicate that the simultaneous action of the pressure and potential differences originates a total flow different from the sum of the electroosmotic and permeation flows due to each force acting separately. The application of irreversible thermodynamics, which includes second-order terms, allowed the determination of the phenomenological coefficients. Moreover, from these values, the equivalent pore radius was estimated on the assumption that the membrane is a porous medium filled with an internal solution. PMID- 15276056 TI - Free energy balance for three fluid phases in a capillary of arbitrarily shaped cross-section: capillary entry pressures and layers of the intermediate-wetting phase. AB - In this work we derive rigorously the free energy balance for three fluid phases in a straight capillary of arbitrarily shaped cross-section. This balance is then used to derive the general equation for the capillary entry pressures of all possible two-phase and three-phase displacements. Moreover, the equation provides the criterion determining the existence of layers of the intermediate-wetting phase separating the wetting and non-wetting phases in the corners or cavities of a capillary, by also treating the spreading of such layers as a capillary displacement. For a number of combinations of interfacial tensions and contact angles, illustrating all the different relevant situations, we calculate the criteria for spreading of such a layer in the corner of a capillary with polygonal cross-section. In a capillary with a cross-section in the shape of an isosceles triangle of varying corner size, these criteria are used to determine the unique capillary entry pressures for piston-like displacement from alternative solutions of the general equation. These solutions relate to displacements in the presence or absence of layers in the various differently sized corners. PMID- 15276057 TI - Thermodynamic study of adsorption of homologous anionic and cationic surfactants. AB - The simplified form of an integral adsorption isotherm based on Butler's equation was applied to describe surface behavior of a series of anionic (sodium alkylsulfonates) and cationic (alkylpyridinium halides) surfactants. This theory allows for the calculation of the free energy of adsorption (Delta G jk) value corresponding to the ability of a particular surfactant to undergo adsorption. The obtained results indicate that the value of Delta G jk depends linearly on the length of the hydrocarbon chain as well as on the kind and concentration of the added inorganic electrolyte. Moreover, it has been found that in the case of surfactants, which have the same length of the alkyl chain and adsorb from solutions containing the same inorganic electrolyte, the charge of hydrophilic group has insignificant influence on the value of Delta G jk. PMID- 15276058 TI - Physicochemical investigation of acrylamide solubilization in sodium bis(2 ethylhexyl)sulfosuccinate and lecithin reversed micelles. AB - The state of acrylamide confined within dry sodium bis(2 ethylhexyl)sulfosuccinate (AOT) and lecithin reversed micelles dispersed in CCl(4) has been investigated by FTIR and (1)H NMR spectroscopy. Measurements have been performed at 25 degrees C as a function of the acrylamide-to-surfactant molar ratio (R) at a fixed surfactant concentration (0.1 mol kg(-1)). The analysis of experimental data, corroborated by the results of SAXS measurements, is consistent with the hypothesis that acrylamide is quite uniformly distributed among reversed micelles mainly located in proximity to the surfactant head-group region and that its presence induces significant unidimensional growth of micellar aggregates. Moreover, the confinement of acrylamide within reversed micelles involves some changes of the typical H-bonded structure of pure solid acrylamide attributable to the establishment of system-specific acrylamide/surfactant head group interactions. Preliminary experiments showed that, by exposure to X-rays, the polymerization of acrylamide can be induced in the confined space of dry AOT and lecithin reversed micelles. PMID- 15276059 TI - Interactions between poly(ethylene oxide) and fatty acids sodium salts studied by surface tension measurements. AB - This work focuses onto the interactions between poly(ethylene oxide) (PEO) and fatty acids, in order to set their potential role of contaminants for PEO-based retention systems. Surface tension measurements were used to investigate PEO fatty acid systems and they made it possible to clearly point out the interactions between the polymer and the sodium octadecylcarboxylates with different degrees of unsaturation. The observed interaction seems to be dependent on the fatty acids' solubility, the increase of which leads to less pronounced phenomena, which are, in contrast, emphasized by the increase in PEO chain length. PMID- 15276060 TI - Favorable interactions of amine- and ester-terminated PAMAM with cationic surfactants: photophysical and transport studies. AB - Conductivity (kappa), turbidity (tau), and fluorescence (I1/I3) studies of hexadecyltrimethylammonium bromide (HTAB), hexadecylpyridinium bromide (HPyBr), and hexadecylpyridinium chloride (HPyCl) in aqueous poly(amido amine) (PAMAM) dendrimers of generations 0 to 2.5 G have been carried out. The complexation of surfactant monomers with the PAMAM surface groups is demonstrated by the critical aggregation concentration (cac), which is two to three orders of magnitude less than the micellization of cationic surfactants in aqueous PAMAM and denoted by critical micelle concentration (cmc*). In the presence of aqueous amine terminated PAMAM, the cmc* value for each surfactant was much lower than the cmc in pure water, while they remain close to each other in the presence of aqueous ester-terminated PAMAM for each surfactant. The fluorescence studies demonstrated that both amine- and ester-terminated PAMAM interact with the cationic surfactants, though the mode of interaction varied due to the different nature of surface groups. PMID- 15276061 TI - Use of self-assembled surfactant systems as media for a substitution reaction. AB - The reaction between 3-bromo-1-propanol and phenol and a series of phenols carrying substituents in 4-position was studied in micellar media and in microemulsions based on either a cationic or a nonionic surfactant. The reactivity and the yield were evaluated and compared to those obtained in a microhomogeneous medium, methanol-water. It was found that the micellar system based on the cationic surfactant dodecyltrimethylammonium bromide was particularly efficient as reaction medium in giving the highest yields. The high reactivity obtained in this system was attributed to formation of a pi-cation complex between the benzene ring of the phenol and the headgroup of the cationic surfactant, resulting in exposure of the phenolate oxygen to the aqueous subvolume where the substrate, 3-bromo-1-propanol, was situated. A highly distorted NMR spectrum of the cationic surfactant gave evidence for this hypothesis. PMID- 15276062 TI - Phase behavior of poly(oxyethylene) cholesteryl ether/novel alkanolamide/water systems. AB - The phase behavior and microstructure of mixed nonionic surfactant systems containing poly(oxyethylene) cholesteryl ether (ChEOn, n=15 and 10), a new alkanolamide-type foam booster, dodecanoyl N -methylethanolamide (NMEA-12), and water, were investigated at 25 degrees C by means of visual observation and small angle X-ray scattering. In the ChEO(15)/water binary system, aqueous micellar (W(m)), discontinuous cubic liquid crystal (I(1)), hexagonal (H(1)), rectangular ribbon (R(1)), lamellar (L(alpha)), and solid (S) phases are successively formed with increasing surfactant concentration. Although the R(1) phase is an intermediate phase formed in a very narrow composition range in conventional surfactant systems, its domain is unusually wider than that of H(1), which may be attributed to the packing constraint caused by the bulky cholesteric group in the lipophilic core of the aggregate. Upon addition of lipophilic NMEA-12 to the ChEO(15)/water binary system, the interfacial curvature of the aggregates decreases, and the micellar or liquid crystal phases formed in the binary system transform to the reverse micellar (O(m)) phase via the L(alpha) phase existing over a wide concentration range. The SAXS results establish an epitaxial relationship between the (11) plane of the R(1) phase and the (10) plane of the L(alpha) phase. The ChEO(10)/NMEA-12/water system shows a phase diagram of similar general appearance, except that the W(m) to R(1) phase transformation occurs via an optically anisotropic liquid crystal phase of unknown structure and the R(1) to L(alpha) phase transition occurs through a narrow intermediate defected lamellar (L(alpha)(H)) phase. The variation in the aggregate size and shape and the unit cell of the R(1) phase formed in ChEOn/NMEA-12/water systems is also discussed. PMID- 15276063 TI - Effect of low-molecular-weight organic anions on electrokinetic properties of variable charge soils. AB - It is known that some inorganic anions can be adsorbed by variable-charge soils specifically, resulting in the lowering of the zeta potential of the clay particle. Reasoning similarly, organic anions should also have such an effect. In this article, the effect of the anions of five low-molecular-weight (LMW) organic acids existing widely in soils on the zeta potentials of two variable-charge soils was examined. The results showed that the presence of organic anions led to a decrease in zeta potential. The effect of different anions on zeta potential followed the order oxalate>citrate>malate>maleate>acetate. The effect increased with the increase in anion concentration and decreased with the increase in pH. The extent of the effect on different soils was apparently related to their iron oxide content. The presence of organic anions also led to a decrease in the isoelectric point (IEP) of the soil. The IEPs of two soils in organic anion systems followed the order acetate>maleate>malate>citrate. No IEP was detected for the oxalate system. PMID- 15276064 TI - Analysis of the electrophoretic mobility and viscosity of dilute Ludox solutions in terms of a spherical gel layer model. AB - A spherical gel layer model of colloidal particles is used to analyze the electrophoretic mobility and viscosity of a dilute suspension of the silica sol, Ludox, reported previously by Laven and Stein (J. Laven, H.N. Stein, J. Colloid Interface Sci. 238 (2001) 8-15). The colloid is modeled as a sphere with a solid inner core surrounded by a diffuse gel layer of uniform thickness and comprising a specific fraction, f, of the colloidal particle's mass. The gel layer is accessible to solvent and ions, but the gel layer retards the flow of solvent, which is assumed to obey the Brinkman equation. The colloidal charge is assumed to be spherically symmetric, but its disposition on the surface of the core particle and gel layer (alpha = fraction of charge in the gel layer) is left as an adjustable parameter. Experiments at pH 5.7 and 8.7 over a KCl concentration range of 0.3 to 80 mM are examined. At high salt and/or low pH, the thickness of the gel layer is estimated to be 1.0 to 1.6 nm depending on the assumed fraction of silica present in the gel layer. At low salt (0.3 mM) and high pH, where the net absolute charge of Ludox is large, the thickness of the gel layer is estimated to be 3.7 to 4.1 nm. Thus, the thickness of the gel layer appears to increase with decreasing salt at high pH. The net charge required to simultaneously match experimental and model mobilities and viscosities is sensitive to the choice of f and alpha. Nonetheless, for reasonable choices of these parameters (f = 0.13 and alpha approximately equal to 1.0), the estimated net absolute charges of Ludox from present modeling are in good agreement with the titration charges of Bolt (G.H. Bolt, J. Phys. Chem. 61 (1957) 1166-1169), and Milonjic (S.K. Milonjic, Colloids Surf. 23 (1987) 301-311) over the entire salt concentration range at pH 8.7. At pH 5.7, however, the estimated net absolute charge from current modeling exceeds the Bolt values by about 50%. PMID- 15276065 TI - Response to "Comment on 'Removal of copper from aqueous solution by aminated and protonated mesoporous aluminas: kinetics and equilibrium'". PMID- 15276066 TI - Phenyl substitution of furamidine markedly potentiates its anti-parasitic activity against Trypanosoma cruzi and Leishmania amazonensis. AB - Furamidine (DB75) and related unfused aromatic diamidines have proven useful for the treatment of parasitic infections. These compounds were primarily developed to combat infections by Pneumocystis carinii and African trypanosomes but they are also active against other parasites. Here we have investigated the in vitro effects of DB75 and its phenyl-substituted analog DB569 on two kinetoplastid haemoflagellates Trypanosomatidae: Trypanosoma cruzi and Leishmania (L) amazonensis. The phenyl-amidine compound DB569 has equivalent DNA binding properties compared to DB75 but it was selected on the basis of its distinct tumor cell distribution properties. We found that DB569 is significantly more potent than DB75 at reducing the proliferation of the parasites, using either isolated parasites in cultures or with cardiomyocyte and macrophage host cells. DB569 is effective towards the intracellular forms of T. cruzi (IC(50) in the low micromolar range) and it exhibits trypanocidal dose-dependent effects against trypomastigote forms of T. cruzi parasites obtained from the Y strain and Dm28c clone, which belong to two different biodemes. Fluorescence microscopy experiments indicated that both diamidines were mostly localized in the nucleus of the mammalian host cells and within the nuclei and kinetoplast of the parasites. Electron microscopy studies showed that the treatment of the parasites with DB75 and DB569 induces important alterations of the parasite nucleus and kinetoplast, at sites where their DNA target is localized. Altogether, the data suggest that the phenyl-substituted furamidine analogue DB569 is a potential new candidate for the treatment of the Chagas' disease and Leishmaniasis. PMID- 15276067 TI - Detection of an alternatively spliced form of deoxycytidine kinase mRNA in the 2' 2'-difluorodeoxycytidine (gemcitabine)-resistant human ovarian cancer cell line AG6000. AB - Gemcitabine (2'-2'-difluorodeoxycytidine (dFdC)) is a deoxycytidine analogue that is effective against solid tumors, including lung cancer and ovarian cancer. dFdC requires the phosphorylation by deoxycytidine kinase (dCK) as a primary step in its activation. Deficiency of dCK is associated with resistance against this compound both in vitro in cancer cell lines and in clinical practice in acute myeloid leukemia and solid tumors. The human ovarian cancer cell line AG6000 is 100,000-fold resistant against dFdC compared to its parent cell line A2780. This cell line proved to be dCK deficient in enzyme activity assays and by Western blot analysis, but by RT-PCR, a normal and a truncated dCK mRNA was found. Sequencing revealed that exon 3 was deleted from the dCK cDNA, resulting in a 74 aa-long open-reading frame due to the generation of a premature stop codon. No gross genomic alteration was observed at the dCK locus, suggesting the involvement of post-transcription mechanisms. Transient transfection experiments indicated that the truncated dCK transcripts are not translated to protein. To study the functional role of the truncated dCK transcripts, both A2780 cells and AG6000 cells were stably transfected with human and rat dCK. The results indicated that over-expression of full-length dCK genes in AG6000 failed to completely reverse the sensitivity to dFdC or other drugs. PMID- 15276068 TI - Differential effects of 9-cis retinoic acid on expression of CC chemokine receptors in human monocytes. AB - 9-cis Retinoic acid (9-CRA) is a lipophilic molecule that binds to the retinoid X receptor (RXR). Although retinoic acid (RA) has been known to regulate neutrophil differentiation, a specific role for 9-CRA in chemokine-mediated cellular processes remains obscure. We investigated the effects of 9-CRA on expression of CC chemokine receptors (CCRs) in human monocytic THP-1 cells and peripheral blood monocytes. RNase protection assay was performed to examine the mRNA levels of CCRs in 9-CRA-treated THP-1 cells. mRNA expression of CCR1 and CCR2 was induced in both a dose and time dependent manner. CCR1 and CCR2 mRNA expression began to increase from 6h after a 100nM 9-CRA treatment and reached a maximal level at 12h. Surface expression of CCRs was monitored by flow cytometry. CCR1 and CCR2 surface expression increased in 9-CRA-treated THP-1 cells, but not in untreated cells. Calcium mobilization and chemotactic activity were determined to examine the effect of 9-CRA on cell movement. The intracellular Ca(2+) concentration and the chemotactic activity increased in 9-CRA-treated cells in response to the CCR1 dependent chemokines Lkn-1, MIP-1alpha, and RANTES, and the CCR2-specific chemokine MCP-1. Increased surface expression of CCR1 and the Ca(2+) influx due to 9-CRA were confirmed in peripheral blood monocytes. Taken together, 9-CRA increases the expression levels of mRNA and protein of both CCR1 and CCR2, and the cell migration ability in THP-1 cells and peripheral blood monocytes, indicating that 9-CRA may regulate inflammatory processes through an increased response to CCR1- and CCR2-dependent chemokines. PMID- 15276069 TI - The flavones luteolin and apigenin inhibit in vitro antigen-specific proliferation and interferon-gamma production by murine and human autoimmune T cells. AB - Plant-derived flavonoids are inhibitors of various intracellular processes, notably phosphorylation pathways, and potential inhibitors of cellular autoimmunity. In this study, the inhibiting effects of various flavonoids on antigen-specific proliferation and interferon-gamma (IFN-gamma) production by human and murine autoreactive T cells were evaluated in vitro. T-cell responses were evaluated for the human autoantigen alpha B-crystallin, a candidate autoantigen in multiple sclerosis, and for the murine encephalitogen proteolipid protein peptide PLP (139-151). The flavones apigenin and luteolin were found to be strong inhibitors of both murine and human T-cell responses while fisitin, quercitin, morin and hesperitin, members of the subclasses of flavonoles and flavanones, were ineffective. Antigen-specific IFN-gamma production was reduced more effectively by flavones than T-cell proliferation, suggesting that the intracellular pathway for IFN-gamma production in T cells is particularly sensitive to flavone inhibition. These results indicate that flavones but not flavanoles or flavanones are effective inhibitors of the potentially pathogenic function of autoreactive T cells. The effects of flavones were the same for human and murine autoreactive T cells, stressing the usefulness of animal models of autoimmunity for further studies on the effects of flavonones on autoimmune diseases. PMID- 15276070 TI - Modulation of eosinophil migration from bone marrow to lungs of allergic rats by nitric oxide. AB - Chronic blockade of nitric oxide (NO) synthesis attenuates the eosinophil infiltration into airways of allergic rats. This study was designed to investigate whether the inhibition of eosinophil influx to the lung of allergic rats reflects modifications in the pattern of cell mobilization from the bone marrow to peripheral blood and/or to lung. Male Wistar rats were treated with N(omega)-nitro-l-arginine methyl ester (l-NAME; 20mg/rat per day) for 4 weeks and sensitized with ovalbumin (OVA). In control rats, the pulmonary OVA-challenge promoted an early (24h) increase in the bone marrow eosinophil population that normalized at 48 h after OVA-challenge, at which time the eosinophils disappeared from the blood and reached the lungs in mass. In l-NAME-treated rats, an accumulation of eosinophils in bone marrow was observed at 24 and 48 h post-OVA challenge. No variation in this cell type number was observed in peripheral blood and bronchoalveolar lavage throughout the time-course studied. In control rats, the adhesion of bone marrow eosinophils to fibronectin-covered wells was significantly increased at 24h after OVA-challenge, whereas in l-NAME-treated rats the increased adhesion was detected at 48 h. A 32% decrease in the expression of inducible nitric oxide synthase (iNOS) (but not endothelial nitric oxide synthase; eNOS) in eosinophils from l-NAME-treated rats was observed. The levels of IgE, IgG(1) and IgG(2a) were not affected by the l-NAME treatment. Our findings suggest that inhibition of NO synthesis upregulates the binding of eosinophils to extracellular matrix proteins such as fibronectin, producing a delayed efflux of eosinophils from bone marrow to peripheral blood and lungs. PMID- 15276071 TI - Sulfated polymannuroguluronate, a novel anti-acquired immune deficiency syndrome (AIDS) drug candidate, targeting CD4 in lymphocytes. AB - Sulfated polymannuroguluronate (SPMG), a marine sulfated polysaccharide, has entered the Phase II clinical trial in China as the first anti-acquired immune deficiency syndrome (AIDS) drug candidate obtained from marine organisms. To determine the binding site(s) (receptors) of SPMG in lymphocytes mediating its anti-AIDS activities, fluorescein-5-isothiocyanate (FITC)-labeled SPMG was used to investigate SPMG binding to lymphocytes. Flow cytometry (FCM) and fluorescence microscopy analysis showed that the SPMG binds to lymphocytes in a rapid, specific, reversible, and saturable fashion. Several SPMG binding proteins were purified by affinity chromatography from lymphocyte membrane preparations. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting analysis revealed that a 55kDa lymphocyte membrane protein is CD4. To characterize the SPMG and CD4 interaction, inhibition assay and surface plasmon resonance (SPR) assay were carried out. SPMG bound to CD4 in a multivalent fashion with specificity. The binding of SPMG to human lymphocyte CD4 was competitively inhibited by human soluble CD4 (hsCD4). Likewise, the binding between hsCD4 and immobilized SPMG was blocked by excess free SPMG. These results indicate that CD4 is one of the specific SPMG binding sites (receptors) in lymphocytes. The interaction between SPMG and CD4 may provide a mechanistic explanation of the immunopotentiating and anti-AIDS activities of SPMG in human immunodeficiency virus (HIV) infected individuals. PMID- 15276072 TI - Cyclic nucleotide-dependent phosphodiesterases (PDEI) inhibition by muscarinic antagonists in bovine tracheal smooth muscle. AB - In bovine tracheal smooth muscle (TSM) strips, muscarinic antagonists (atropine, 4-DAMP, AFDX-116 and methoctramine) were able to increase simultaneously and a similar fashion the intracellular levels of cyclic nucleotides, with a cAMP/cGMP ratio higher than 2.0. These original pharmacological responses were time-and dose-dependent, exhibiting maximal values at 15 min, with a pEC(50) of 7.4 +/- 0.2 for atropine and 4-DAMP. These effects on cAMP and cGMP levels were similar to the ones obtained with isobutyl-methylxantine (IBMX, 10 microM), a non selective cyclic nucleotide phosphodiesterase (PDE) inhibitor, suggesting the involvement of PDEs in these muscarinic antagonist responses. Neither, rolipram (10 microM), a specific PDEIV inhibitor, nor zaprinast (10 microM), a PDEV inhibitor, exhibited this "atropine-like" responses. Instead, atropine enhanced the increments of cAMP levels induced by rolipram and cGMP levels by zaprinast. However, vinpocetine (20 microM), a non-calmodulin dependent PDEIC inhibitor was able to mimic these muscarinic antagonist responses in intact smooth muscle strips. In addition, in cell free systems, muscarinic antagonists inhibited the membrane-bound PDEIC activity whereas soluble (cytosol) PDEIC activity was not affected by these muscarinic drugs. These results indicate that muscarinic antagonists acting possibly as inverse agonists on M(2)/M(3)mAChRs anchored to sarcolemma membranes can initiate a new signal transducing cascade leading to the PDEIC inhibition, which produced a simultaneous rise in both cAMP and cGMP intracellular levels in tracheal smooth muscle. PMID- 15276073 TI - Differential response of MG132 cytotoxicity against small cell lung cancer cells to changes in cellular GSH contents. AB - The effect of the depletion or oxidation of cellular GSH on cytotoxicity of MG132 was assessed. Viability loss and decrease in GSH contents in small cell lung cancer (SCLC) cells treated with MG132 was attenuated by caspase inhibitors (z IETD.fmk, z-LEHD.fmk and z-DQMD.fmk). Thiol compounds (N-acetylcysteine and N-(2 mercaptopropionyl)glycine) and free radical scavengers reduced MG132-induced cell death. Antioxidants, including N-acetylcysteine, inhibited the MG132-induced nuclear damage, loss in mitochondrial transmembrane potential, cytosolic accumulation of cytochrome c and caspase-3 activation. Depletion of GSH due to buthionine sulfoxime did not affect the cell viability loss, ROS formation and GSH depletion due to MG132 in SCLC cells. A thiol oxidant monochloramine, p chloromercuribenzoate and N-ethylmaleiamide also did not affect cytotoxicity of MG132. The results suggest that the toxicity of MG132 on SCLC cells is mediated by activation of caspase-8, -9 and -3. Removal of free radicals and recovery of GSH contents may attenuate MG132-induced apoptotic cell death. Nevertheless, depletion or oxidation of cellular GSH may not affect toxicity of MG132. PMID- 15276074 TI - Pulses of external ATP aid to the synchronization of pancreatic beta-cells by generating premature Ca(2+) oscillations. AB - Pancreatic beta-cells respond to glucose stimulation with increase of the cytoplasmic Ca(2+) concentration ([Ca(2+)](i)), manifested as membrane-derived slow oscillations sometimes superimposed with transients of intracellular origin. The effect of external ATP on the oscillatory Ca(2+) signal for pulsatile insulin release was studied by digital imaging of fura-2 loaded beta-cells and small aggregates isolated from islets of ob/ob-mice. Addition of ATP (0.01-100 microM) to media containing 20mM glucose temporarily synchronized the [Ca(2+)](i) rhythmicity in the absence of cell contact by eliciting premature oscillations. External ATP triggered premature [Ca(2+)](i) oscillations also when the sarcoendoplasmic reticulum Ca(2+)-ATPase was inhibited with 50 microM cyclopiazonic acid and phospholipase C inhibited with 10 microM U-73122. The effect of ATP was mimicked by other activators of cytoplasmic phospholipase A(2) (10nM acetylcholine, 0.1-1 micro M of the C-terminal octapeptide of cholecystokinin and 2 microg/ml melittin) and suppressed by an inhibitor of the enzyme (50 microM p-amylcinnamoylanthranilic acid). Premature oscillations generated by pulses of ATP sometimes triggered subsequent oscillations. However, prolonged exposure to high concentrations of the nucleotide (10-100 microM) had a suppressive action on the beta-cell rhythmicity. The early effects of ATP included generation of transients induced by inositol (1,4,5) trisphosphate and superimposed on the premature oscillation or on an ordinary oscillation induced by glucose. The results support the idea that purinergic activation of phospholipase A(2) has a co-ordinating effect on the beta-cell rhythmicity by triggering premature [Ca(2+)](i) oscillations mediated by closure of ATP sensitive K(+) channels. PMID- 15276075 TI - Agonist binding and activation of the rat beta(1)-adrenergic receptor: role of Trp(134(3.28)), Ser(190(4.57)) and Tyr(356(7.43)). AB - We investigated the role of Trp(134(3.28)), Ser(190(4.57)) and Tyr(356(7.43)) in agonist binding to, and activation of, the rat beta(1)-adrenergic receptor by comparing pK(i)s and functional responses of W134A, S190A and Y356F mutant receptors to wild type, all stably expressed in CHO cells. All three mutations significantly (P < 0.05) reduced adenylyl cyclase intrinsic activity (IA) compared to wild type in response to stimulation with both (-)-isoprenaline (53 88%) and (-)-RO363 (46-61%), and there was no significant correlation either between IA or pD(2) and pK(i) (P > 0.4), suggesting that changes in pK(i) were not sufficient to explain the fall in adenylyl cyclase activity. The most pronounced reduction in affinity (126-fold, P < 0.01) was displayed by xamoterol for the Y356F mutation, suggesting that xamoterol is able to directly interact with Tyr(356(7.43)). For the other agonists, the change in pK(i) values for the mutant receptors ranged from a 20-fold decrease to a 2-fold increase compared to the wild type. In a three-dimensional model of the rat beta(1)-adrenergic receptor, Trp(134(3.28)) and Tyr(356(7.43)) form part of a hydrophobic binding pocket involving residues in transmembrane helices 1, 2, 3 and 7. Our results suggest that Trp(134(3.28)) and Tyr(356(7.43)), together with Trp(353(7.40)), are able to interact via pi-pi interactions to stabilize the extracellular ends of transmembrane helices 3 and 7. Ser(190(4.57)) appears to be involved in a hydrogen bonding network, which maintains the spatial relationship between transmembrane helices 3 and 4. These interhelical interactions suggest that the three mutated residues stabilize the active receptor state by maintaining the proper packing of their respective transmembrane helix within the helix bundle, facilitating the appropriate movement and rotation of the transmembrane regions during the activation process. PMID- 15276076 TI - Peripheral-type benzodiazepine receptor density and in vitro tumorigenicity of glioma cell lines. AB - The peripheral-type benzodiazepine receptor is found primarily on the outer mitochondrial membrane and consists of three subunits: the 18kDa isoquinoline binding protein, the 32kDa voltage-dependent anion channel, and the 30kDa adenine nucleotide transporter. The current study evaluates the potential importance of peripheral-type benzodiazepine receptor expression in glioma cell tumorigenicity. While previous studies have suggested that peripheral-type benzodiazepine receptor-binding may be relatively increased in tumor tissue and cells, so far, little is known about the relationships between peripheral-type benzodiazepine receptor density and factors underlying tumorigenicity. In the present study, we found in glioma cell lines (C6, U87MG, and T98G), that peripheral-type benzodiazepine receptor ligand-binding density is relatively high for C6 and low for T98G, while U87MG displays intermediate levels. Cell growth of these cell lines in soft agar indicated that high levels of peripheral-type benzodiazepine receptor-binding were associated with increased colony size, indicative of their ability to establish anchorage independent cell proliferation. Potential causes for differences in tumorigenicity between these cell lines were suggested by various cell death and proliferation assays. Cell death, including apoptosis, appeared to be low in C6, and high in T98G, while U87MG displayed intermediate levels in this respect. Cell proliferation appeared to be high in C6, low in T98G, and intermediate in U87MG. In conclusion, our study suggests that relatively high peripheral-type benzodiazepine receptor-binding density is associated with enhanced tumorigenicity and cell proliferation rate. In particular, apoptosis appears to be an important tumorigenic determinant in these glioma cell lines. Moreover, application of PBR-specific ligands indicated that PBR indeed are functionally involved in apoptosis in glioma cells. PMID- 15276077 TI - Differential regulation of zinc efflux transporters ZnT-1, ZnT-5 and ZnT-7 gene expression by zinc levels: a real-time RT-PCR study. AB - Intracellular zinc levels are strictly regulated by zinc channels and zinc binding proteins to maintain cellular zinc-dependent functions. We demonstrated a correlation between extracellular zinc concentration and intracellular exchangeable zinc levels using the fluorescent zinc-specific probes zinquin and zinpyr-1. The effect of extracellular zinc status on the regulation of the two trans-Golgi network directed zinc transporters ZnT-5 and ZnT-7 was next studied by real-time RT-PCR in zinc supplemented or depleted HeLa cells. While sub-toxic extracellular zinc addition strongly induced the efflux transporter ZnT-1 gene expression, consistent with its activation by the transcription factor MTF-1, treated HeLa cells did not display any change in ZnT-5 and ZnT-7 mRNA levels compared to control cells. In contrast, zinc depletion induced by non-toxic doses of the zinc chelator TPEN (N,N,N',N' tetrakis-(2 pyridylmethyl) ethylene diamine) resulted in a up to eight-fold induction of transporters ZnT-5 and ZnT-7 mRNA levels, providing the first evidence of a transcriptional control of these two zinc efflux transporters by zinc deficiency in cultured cells. PMID- 15276078 TI - Enantioselectivity of ribonucleotide reductase: a first study using stereoisomers of pyrimidine 2'-azido-2'-deoxynucleosides. AB - In this paper, the enantioselectivity of ribonucleotide reductase (RNR, EC 1.17.4.1), a pivotal enzyme involved in DNA biosynthesis, was studied using the beta-d and beta-l stereoisomers of 2'-azido-2'-deoxynucleosides of uracil and cytosine. The corresponding 5'-diphosphate derivatives in the d-configuration have been extensively studied as mechanism-based inhibitors of the enzyme. The original l-enantiomers were synthesized and evaluated in vitro. In cell culture experiments, only the cytosine derivative with a d-configuration was found cytostatic and able to deplete dNTP pools in response to RNR inhibition. In the case of the uracil enantiomeric pair, this result correlates with an inefficient intracellular monophosphorylation as demonstrated in testing their substrate properties against human uridine-cytidine kinase 1. Regarding cytosine analogues, human deoxycytidine kinase was found to be able to phosphorylate both enantiomers with comparable efficiency but only the d-stereoisomer was active in human cell culture. The interaction of the beta-d and beta-l stereoisomers of 2'-azido-2' deoxyuridine 5'-diphosphate with purified Escherichia coli RNR was also examined. Inactivation of the enzyme was only observed in the presence of the d stereoisomer, demonstrating that RNR exhibits enantiospecificity with respect to the natural configuration of the sugar moiety, as far as 2'-azido-2' deoxynucleotides are concerned. PMID- 15276079 TI - Inhibition of lipopolysaccharide-induced inducible nitric oxide synthase expression by a novel compound, mercaptopyrazine, through suppression of nuclear factor-kappaB binding to DNA. AB - Macrophage cells in response to cytokines and endotoxins produced a large amount of nitric oxide (NO) by expression of inducible nitric oxide synthase (iNOS), resulting in acute or chronic inflammatory disorders including septic hypotension and atherosclerosis. In the present study, we investigated the effect and the mechanism of mercaptopyrazine (MP) in the induction of iNOS and NO production as a culminating factor for several inflammatory disorders. Pretreatment of MP alleviated the mortality of endotoxemic mice receiving a lethal bolus of lipopolysaccharide (LPS), which was associated with the reduced levels of serum nitrite/nitrate and IL-1beta. In RAW264.7 mouse macrophage cells, MP (300microM) inhibited both protein and mRNA levels of iNOS stimulated by LPS/interferon-gamma (IFNgamma) up to 50%. The nuclear factor-kappa B (NF-kappaB)-driven transactivation was also suppressed by MP to the same degree. Treatment of MP reduced the binding of NF-kappaB to the oligonucleotides containing NF-kappaB consensus sequence, while it did not affect the translocation of NF-kappaB to nuclear. Suppression of NF-kappaB activity by MP was completely reversed by a reducing agent, dithiothreitol, implying that MP might oxidize the sulfhydryl group(s) of DNA binding domain of NF-kappaB. In conclusion, MP would be one of interesting candidates or chemical moieties of iNOS expression inhibitor via specific suppression of NF-kappaB binding to DNA, and be useful as a chemopreventive agent or a therapeutic against iNOS-associated inflammatory diseases. PMID- 15276080 TI - Participation of cyclin D1 deregulation in TNP-470-mediated cytostatic effect: involvement of senescence. AB - Inhibition of angiogenesis is becoming one promising, alternative approach to stop tumor from growth and spreading to distant organs. TNP-470, an analog of fumagillin, possesses potent anti-angiogenic effects with minimal toxicity in animal tumor models and is now in the phase III of human cancer trial. Although TNP-470 induced endothelial cell cycle arrest at G1 phase via p53 and p21(Cip1), the underlying mechanism of the cytostatic effect of TNP-470 on endothelial cells remains limited. We have found that TNP-470 did not only induce p53 and p21(Cip1) but also cyclin D1 in the basic fibroblast growth factors (bFGF)-treated endothelial cells. The TNP-470-mediated increase of cyclin D1 protein was due to the enhanced expression of mRNA. The induced cyclin D1 formed a complex with cyclin-dependent kinase4 (CDK4) and p21(Cip1). The ability of cyclin D1 associated CDK4 to phosphorylate retinoblastoma (Rb) protein was, however, reduced in the same cells. TNP-470 also significantly increased senescence associated-beta-galactosidase activity (SA-gal), hallmark of cells undergoing senescence. Interestingly, the effect of increased cyclin D1 protein mimicked by overexpression of cyclin D1 increased the sensitivity of human umbilical vein endothelial cells (HUVECs) to TNP-470. In summary, the cytostatic effect of TNP 470 on endothelial cells is in part mediated by induction of senescence and cyclin D1 is a key molecule participating in this event. PMID- 15276081 TI - Inhibition of ossification in vivo and differentiation of osteoblasts in vitro by tributyltin. AB - Tributyltin is ubiquitous in the environment and an endocrine disruptor for many wildlife species. However, minimal information is available regarding the effect of this chemical on bone formation. When tributyltin chloride (TBT) (1mg/kg body weight) was administered subcutaneously to pregnant mice at 10, 12, and 14 days post coitus (dpc), fetuses at 17.5 days post coitus revealed the inhibition of calcification of supraoccipital bone. In contrast, 1mg/kg body weight monobutyltin trichloride (MBT) did not affect the fetal skeleton. Therefore, we examined the effects of TBT and its metabolites (dibutyltin dichloride, DBT, and MBT) on bone metabolism using rat calvarial osteoblast-like cells (ROB cells). The viability of ROB cells was not affected by the exposure of the cells to 10( 10) to 10(-7)M TBT. However, TBT reduced the activity of alkaline phosphatase (ALPase) and the rate of deposition of calcium of ROB cells. In addition, the expression levels of mRNA for ALPase and osteocalcin, which are markers of osteoblastic differentiation, were depressed by the treatment with TBT. TBT inhibited ALPase activity and the deposition of calcium to a greater extent than did DBT. MBT had no effect on the osteoblast differentiation of ROB cells. Tributyltin is known to inhibit the activity of aromatase. However, the aromatase inhibitor aminoglutethimide did not reproduce the inhibitory effects of TBT on osteoblast differentiation. Our findings indicate that TBT might have critical effects on the formation of bone both in vivo and in vitro although its action mechanism is not clarified. PMID- 15276082 TI - Characterization of hydroxyl radical formation by microsomal enzymes using a water-soluble trap, terephthalate. AB - Using terephthalic acid as a water-soluble trap, we characterized hydroxyl radicals (HO?) formation by liver microsomal enzymes from isoniazid-treated rats. We found that HO? formation was entirely dependent on intact microsomal enzymes, the presence of NADPH, and iron complexed with EDTA. In contrast to the other radical traps, we found no evidence that terephthalate is a substrate for cytochrome P450. Cumene hydroperoxide, an artificial supporter of cytochrome P450 catalyzed oxidation, failed to maintain HO(.-) formation. HO(.-) formation in liver microsomes was inhibited by the HO(.-) radical scavengers: dimethyl sulfoxide (DMSO), mannitol, and citrulline. It was abolished by catalase, but not superoxide dismutase (SOD), indicating that hydrogen peroxide was the sole precursor of the HO(.-). Therefore, the generation of hydroxyl radicals by microsomal enzymes appears to be dependent on two processes: (1) the rate of hydrogen peroxide production; and (2) the availability of iron ions or other transition metals for Fenton type reactions. PMID- 15276083 TI - Effect of the histone deacetylase inhibitor trichostatin A on spontaneous apoptosis in various types of adult rat hepatocyte cultures. AB - Acetylation and deacetylation of histones, catalysed by histone acetyl transferases and histone deacetylases (HDAC), respectively, are known to be involved in gene expression regulation. Here, the effect on the activity and expression of several apoptosis-related proteins of trichostatin A (TSA), a well known HDAC inhibitor, were studied in short-term (conventional monolayer) and long-term cultured (collagen I gel sandwich cultures and co-cultures) adult rat hepatocytes. No significant effects of TSA on the caspase-3-like activity were seen in rat hepatocytes cultured in a sandwich configuration or in a co-culture with rat liver epithelial cells of primitive biliary origin. In both culture models, the basal level of apoptosis was found to be much lower than in control monolayer cultures. In the latter system, it was found that, after 4 days of culture, TSA decreased the levels of caspase-3 (both proform and p17 fragment) and of the pro-apoptotic protein Bid. No effect of TSA was found on the expression of Bax. As expected, a TSA-mediated increase of acetylated histones H3 and H4 was observed in all culture systems examined. In addition, in the presence of TSA, increased albumin secretion and cytochrome P450 1A1/2 and 2B1-dependent enzyme activities were found in conventional cultures after 7 days. In conclusion, TSA delayed the occurrence of apoptosis and loss of liver specific functions in conventional hepatocyte monolayers. In contrast, in hepatocyte culture models in which spontaneous apoptosis is already minimised through the addition of either extracellular matrix components (sandwich cultures) or non parenchymal liver cells (co-cultures), TSA did not have any additional anti apoptotic effect. PMID- 15276084 TI - Comparative pharmacology of human dopamine D(2)-like receptor stable cell lines coupled to calcium flux through Galpha(qo5). AB - The goal of this study was to develop a new approach to study the pharmacology of the dopamine D(4) receptor that could be used in comparative studies with dopamine D(2) and D(3) receptors. Stable HEK-293 cell lines co-expressing recombinant human D(2L), D(3) or D(4) receptors along with Galpha(qo5) cDNA were prepared. Dopamine induced a robust, transient calcium signal in these cell lines with EC(50)s for D(2L), D(3) and D(4) of 18.0, 11.9 and 2.2 nM, respectively. Reported D(4)-selective agonists CP226269 and PD168077 were potent, partial D(4) agonists exhibiting 31-1700-fold selectivity for D(4) over D(3) or D(2). Non selective D(2)-like agonists apomorphine and quinpirole showed full efficacy but did not discriminate across the three receptors. D(3)-selective agonists 7 hydroxy-DPAT and PD128907 were potent but non-selective D(2)-like agonists. The reported D(3) partial agonist BP-897 exhibited minimal agonist activity at D(3) but was a potent D(3) antagonist and a partial D(4) agonist. Other D(2)-like antagonists, haloperidol, clozapine, and domperidone showed concentration dependent inhibition of dopamine responses at all three receptors with K(i) ranging from 0.05 to 48.3 nM. The D(3) selective antagonist S33084 and D(4) selective antagonist L-745870 were highly selective for D(3) and D(4) receptors with K(b) of 0.7 and 0.1 nM, respectively. Stable co-expression of D(2)-like receptors with chimeric Galpha(qo5) proteins in HEK-293 cells is an efficient method to study receptor activation in a common cellular background and an efficient method for direct comparison of ligand affinity and efficacy across human D(2L), D(3) and D(4) receptors. PMID- 15276085 TI - Involvement of cytochrome P450 1A2 in the biotransformation of trans-resveratrol in human liver microsomes. AB - This study was aimed at identifying the isoform(s) of human liver cytochrome P450 (CYP) involved in the hepatic biotransformation of trans-resveratrol (trans 3,5,4'-trihydroxystilbene). Trans-resveratrol metabolism was found to yield two major metabolites, piceatannol (3,5,3',4'-tetrahydroxystilbene) and another tetrahydroxystilbene named M1. Trans-resveratrol was hydroxylated to give piceatannol and M1 with apparent K(m) of 21 and 31 microM, respectively. Metabolic rates were in the range 14-101 pmol min(-1) mg(-1) protein for piceatannol and 29-161 pmol min(-1) mg(-1) protein for M1 in the 13 human liver microsomes tested. Using microsomal preparations from different human liver samples, piceatannol and M1 formation significantly correlated with ethoxy resorufin-O-deethylation (r(2) = 0.84 and 0.88, respectively), phenacetin-O deethylation (r(2) = 0.92 and 0.94) and immuno-quantified CYP1A2 (r(2) = 0.85 and 0.90). Formation of these metabolites was markedly inhibited by alpha naphthoflavone and furafylline, two inhibitors of CYP1A2. Antibodies raised against CYP1A2 also inhibited the biotransformation of trans-resveratrol. In addition, the metabolism of trans-resveratrol into these two metabolites was catalyzed by recombinant human CYP1A1, CYP1A2 and CYP1B1. Our results provide evidence that in human liver, CYP1A2 plays a major role in the metabolism of trans-resveratrol into piceatannol and tetrahydroxystilbene M1. PMID- 15276086 TI - Rapid induction of P-glycoprotein expression by high permeability compounds in colonic cells in vitro: a possible source of transporter mediated drug interactions? AB - P-glycoprotein (PGP) substrates with high membrane permeability, such as propranolol and verapamil, are considered to be essentially "transparent" to PGP since the transporter does not significantly limit their absorption or elimination. However, the question of whether such compounds can modulate PGP expression in epithelial cells following short-term exposure, with potential consequences for drug interactions, has not been addressed. LS180 colonic epithelial cells were exposed to propranolol or verapamil at concentrations (50 300 microM) consistent with those likely to be present in the gut lumen during oral dosing. Both compounds stimulated four to six-fold increases in MDR1 mRNA and PGP protein expression measured by quantitative real-time PCR and immunoblotting, respectively. These changes were accompanied by an induction in transporter activity measured by rhodamine 123 efflux. In contrast, metoprolol, a compound with similar permeability but no affinity for PGP had no effect on PGP expression. The induction of PGP by propranolol and verapamil was rapid with significant increases occurring within 3h with maximal stimulation after 6h exposure. Rifampicin, shown to cause clinical drug interactions via a PXR mediated increase in PGP expression, exhibited a very similar time-course and extent of induction. In conclusion, verapamil and propranolol, whose trans epithelial permeability are unaffected by PGP, appear to be effective inducers of PGP expression in gut epithelial cells in vitro. While the in vivo significance of these observations is unknown, this questions whether high permeability, "PGP transparent" compounds, currently favoured in drug selection strategies, should be evaluated in terms of their potential for transporter-mediated drug interactions. PMID- 15276087 TI - Effect of acetaminophen on expression and activity of rat liver multidrug resistance-associated protein 2 and P-glycoprotein. AB - We evaluated the effect of acetaminophen (APAP), given as a single, 1g/kg body weight dose, on expression and activity of rat liver multidrug resistance associated protein 2 (Mrp2) and P-glycoprotein (P-gp), two major canalicular drug transporters. The studies were performed 24h after administration of the drug. APAP induced an increase in plasma membrane content of Mrp2 detected by western blotting, consistent with increased detection of the protein at the canalicular level by immunoflourescence microscopy. In vivo biliary excretion of dinitrophenyl-S-glutathione, a well known Mrp2 substrate, was slightly but significantly increased by APAP, agreeing well with upregulation of the transporter. Basal biliary excretion of oxidized glutathione, an endogenous Mrp2 substrate, was also increased by APAP, likely indicating increased hepatic synthesis as a result of APAP-induced oxidative stress followed by accelerated canalicular secretion mediated by Mrp2. APAP also increased the expression of P gp detected by western blotting and immunofluorescence microscopy as well as the in vivo biliary secretory rate of digoxin, a model P-gp substrate. Because specific APAP-conjugated metabolites are Mrp2 substrates, we postulate that induction of Mrp2 by APAP may represent an adaptive mechanism to accelerate liver disposition of the drug. In addition, increased Mrp2-mediated elimination of oxidized glutathione may be essential in maintaining the redox equilibrium in the hepatocyte under conditions of APAP-induced oxidative stress. PMID- 15276088 TI - Efficacy and safety of two unfractionated heparin dosing strategies with tenecteplase in acute myocardial infarction (results from Assessment of the Safety and Efficacy of a New Thrombolytic Regimens 2 and 3). AB - We investigated the effect of smaller dose, weight-adjusted heparin with earlier monitoring of activated partial thromboplastin time on the incidence of ischemic and hemorrhagic complications in patients with ST-elevation myocardial infarction treated with full-dose tenecteplase. We compared the outcomes of patients enrolled in the Second Assessment of the Safety and Efficacy of a New Thrombolytic Regimen (ASSENT-2; n = 8,461) who received heparin stratified by weight (patients weighing >67 kg received a 5,000-U bolus plus infusion at 1,000 U/hour; those weighing < or =67 kg received a 4,000-U bolus plus infusion at 800 U/hour) with patients in ASSENT-3 who received weight-adjusted heparin (60-U/kg bolus, maximum 4,000 U/hour, followed by a 12-U/kg/hour infusion, maximum 1,000 U/hour). Compared with patients in ASSENT-2, those in ASSENT-3 had similar rates of 30-day mortality, recurrent infarction, and intracranial hemorrhage, less major bleeding (2.2% vs 4.7%, p <0.001), and less refractory ischemia (6.5% vs 8.6%, p <0.001). After adjustment for baseline characteristics, patients in ASSENT-3 had similar rates of 30-day mortality (odds ratio [OR] 0.96, 95% confidence interval [CI] 0.77 to 1.19) and intracranial hemorrhage (OR 1.02, 95% CI 0.61 to 1.69) but less major bleeding (OR 0.49, 95% CI 0.35 to 0.67) than did patients in ASSENT-2. These findings support the use of smaller dose, weight adjusted heparin in patients with ST-elevation myocardial infarction treated with tenecteplase. PMID- 15276089 TI - Quantification of myocardial infarct size and transmurality by contrast-enhanced magnetic resonance imaging in men. AB - Contrast-enhanced magnetic resonance (ce-MR) imaging allows precise delineation of infarct transmurality. An issue of debate is whether data analysis should be performed visually or quantitatively. Accordingly, a head-to-head comparison was performed between visual and quantitative analyses of infarct transmurality on ce MR imaging. In addition, infarct transmurality was related to the severity of wall motion abnormalities at rest. In 27 patients with long-term ischemic left ventricular (LV) dysfunction (LV ejection fraction 33 +/- 8%) and previous infarction, cine MR imaging (to assess regional wall motion) and ce-MR imaging were performed. Using a 17-segment model, each segment was assigned a wall motion score (from normokinesia to dyskinesia), and segmental infarct transmurality was visually assessed on a 5-point scale (0 = no infarction, 1 = transmurality < or =25% of LV wall thickness, 2 = transmurality 26% to 50%, 3 = transmurality 51% to 75%, and 4 = transmurality 76% to 100%). Quantification of transmurality was performed with threshold analysis; myocardium showing signal intensity above the threshold was considered scar tissue, and percent transmurality was calculated automatically. Wall motion was abnormal in 56% of the 459 segments, and 55% of segments showed hyperenhancement (indicating scar tissue). The agreement between visual and quantitative analyses was excellent: 90% of segments (kappa 0.86) were categorized similarly by visual and quantitative analyses. Infarct transmurality paralleled the severity of contractile dysfunction; 96% of normal or mildly hypokinetic segments had infarct transmurality < or =25%, whereas 93% of akinetic and dyskinetic segments had transmurality >50% on visual analysis. In conclusion, visual analysis of ce-MR imaging studies may be sufficient for assessment of transmurality of infarction. PMID- 15276090 TI - Differential prognostic significance of peri-infarction versus remote myocardial ischemia on stress technetium-99m sestamibi tomography in patients with healed myocardial infarction. AB - Peri-infarction and remote myocardial ischemia involve different myocardial substrates, but their differential clinical implications have not been previously studied. We assessed the differential prognostic significance of peri-infarction and remote ischemia during long-term follow-up in patients with healed myocardial infarction. We studied 345 patients (59 +/- 12 years old; 282 men) with previous myocardial infarction who demonstrated reversible perfusion abnormalities on exercise or dobutamine stress technetium-99m sestamibi tomography. Follow-up events for 5.5 +/- 2.6 years were 60 deaths (17%; 40 cardiac deaths) and 25 reinfarctions (7%). Reversible perfusion abnormalities were detected in the remote region in 129 patients (37%), the peri-infarction region in 142 patients (41%), and in both regions in 74 patients (21%). The annual rates of cardiac death in these groups were 1.2%, 2.8%, and 2.9%, respectively (p <0.01). The annual rates of reinfarction were 1%, 1.5%, and 0.9%, respectively (p = NS). In a multivariate analysis model, independent predictors of cardiac death were history of heart failure (risk ratio [RR] 2.8, 95% confidence interval [95% CI] 1.2 to 7), diabetes mellitus (RR 4.1, 95% CI 1.9 to 8.9), summed score at rest (RR 1.4, 95% CI 1.1 to 3.1), and peri-infarction ischemia (RR 2.6, 95% CI 1.1 to 6.1). Predictors of reinfarction were age (RR 1.03, 95% CI 1.01 to 1.07) and diabetes mellitus (RR 3.3, 95% CI 1.2 to 9.1). Peri-infarction ischemia assessed by stress technetium-99m sestamibi tomography is associated with a greater risk of cardiac death than is remote ischemia. The risk of reinfarction is not related to the location of ischemia. PMID- 15276091 TI - Brachial artery reactivity in asymptomatic patients with type 2 diabetes mellitus and microalbuminuria (from the Detection of Ischemia in Asymptomatic Diabetics brachial artery reactivity study). AB - Microalbuminuria is a novel atherosclerotic risk factor in patients with type 2 diabetes mellitus (DM) and predicts future cardiovascular events. Endothelial dysfunction and systemic inflammation have been proposed as common links between microalbuminuria and cardiovascular disease. However, no study has assessed the relation between microalbuminuria and vascular dysfunction as measured by brachial artery reactivity (BAR) in DM. We evaluated 143 patients (85 men; mean age 60.0 +/- 6.7 years) with DM (mean duration 8.2 +/- 7.4 years) enrolled in the Detection of Ischemia in Asymptomatic Diabetics study. Subjects were categorized as those with microalbuminuria (ratio of urinary albumin to creatinine 30 to 299 microg/mg creatinine, n = 28) and those with normoalbuminuria (ratio of urinary albumin to creatinine 0 to 29.9 microg/mg creatinine, n = 115). High-resolution ultrasound BAR testing was used to measure endothelium-dependent and endothelium independent vasodilations. C-reactive protein was measured as a marker of systemic inflammation. Patients with microalbuminuria and normoalbuminuria had similar baseline characteristics, with the exception that those with microalbuminuria had a longer duration of DM (p = 0.03). Endothelium-dependent vasodilation at 1 minute (p = 0.01) and endothelium-independent vasodilation at 3 minutes (p = 0.007) were significantly less in patients with microalbuminuria. In addition, 96% of patients with microalbuminuria and 76% of those with normoalbuminuria had impaired endothelium-dependent vasodilation (<8%, p = 0.01). Microalbuminuria was an independent predictor of endothelium-dependent vasodilation in the entire cohort (p = 0.045) and after excluding patients on hormone replacement therapy (p = 0.01). Levels of C-reactive protein were significantly higher in patients with microalbuminuria than in those with normoalbuminuria (p = 0.02). We conclude that in DM the presence of microalbuminuria is associated with impaired endothelium-dependent and endothelium-independent vasodilations of the brachial artery and a higher degree of systemic inflammation. In addition, microalbuminuria is an independent predictor of endothelial dysfunction in asymptomatic patients with DM, especially in the absence of hormone replacement therapy. PMID- 15276092 TI - Impact of chronic kidney disease on prognosis of patients with diabetes mellitus treated with percutaneous coronary intervention. AB - Chronic kidney disease (CKD) is a frequent complication of diabetes mellitus. However, the role of CKD in outcomes of patients with diabetes who have undergone percutaneous coronary intervention (PCI) has not been studied specifically. Therefore, we investigated the impact of CKD on prognosis of patients with diabetes who underwent PCI. Of 1,575 diabetic patients who underwent PCI, 1,046 (66%) had preserved renal function, 492 (31%) had CKD (baseline serum creatinine >1.5 mg/dl or estimated glomerular filtration rate <60 ml/min/1.73 m(2)) without dialysis, and 37 (2.3%) were dependent on dialysis. Patients with CKD versus those without CKD had more in-hospital complications, including mortality (2.6% vs 0.5%, respectively; p <0.0001), neurologic events (3.1% vs 0.6%, p = 0.0001), and gastrointestinal bleeding (2.9% vs 0.9%, p = 0.01). Contrast-induced nephropathy after PCI (increase > or =25% and/or > or =0.5 mg/dl of serum creatinine before PCI vs 48 hours after PCI) was found in 15% of patients without CKD versus 27% of those with CKD, and de novo dialysis was instituted in 0.1% versus 3.1%, respectively. Contrast-induced nephropathy was independently predicted (all p <0.0001) by peri-PCI hypotension (odds ratio [OR] 2.62), insulin treatment (OR 1.84), and volume of contrast medium (OR 1.30). The 1-year mortality rate was strikingly higher (all p <0.0001) in patients with CKD who did not receive dialysis (16%) and those on dialysis (44%) compared with the group with preserved renal function (5%). Contrast-induced nephropathy was among the independent predictors of a 1-year mortality rate (OR 2.75, p <0.001). PMID- 15276093 TI - Safety and compliance with once-daily niacin extended-release/lovastatin as initial therapy in the Impact of Medical Subspecialty on Patient Compliance to Treatment (IMPACT) study. AB - Niacin extended-release/lovastatin is a new combination product approved for treatment of primary hypercholesterolemia and mixed dyslipidemia. This open labeled, multicenter study evaluated the safety of bedtime niacin extended release/lovastatin when dosed as initial therapy and patient compliance to treatment in various clinical practice settings. A total of 4,499 patients with dyslipidemia requiring drug intervention was enrolled at 1,081 sites. Patients were treated with 1 tablet (500 mg of niacin extended-release/20 mg of lovastatin) once nightly for 4 weeks and then 2 tablets for 8 weeks. Patients also received dietary counseling, educational materials, and reminders to call a toll-free number that provided further education about dyslipidemia and niacin extended-release/lovastatin. Primary end points were study compliance, increases in liver transaminases to >3 times the upper limit of normal, and clinical myopathy. Final study status was available for 4,217 patients (94%). Compliance to niacin extended-release/lovastatin was 77%, with 3,245 patients completing the study. Patients in the southeast and those enrolled by endocrinologists had the lowest compliance and highest adverse event rates. Flushing was the most common adverse event, reported by 18% of patients and leading to discontinuation by 6%. Incidence of increased aspartate aminotransferase and/or alanine aminotransferase >3 times the upper limit of normal was <0.3%. An increase of creatine phosphokinase to >5 times the upper limit of normal occurred in 0.24% of patients, and no cases of drug-induced myopathy were observed. Niacin extended release/lovastatin 1,000/40 mg, dosed as initial therapy, was associated with good compliance and safety and had very low incidences of increased liver and muscle enzymes. PMID- 15276094 TI - Temporal aspects of improved survival with the implanted defibrillator (MADIT II). AB - The present study retrospectively explored the reasons for delay in the onset of survival benefit from the implanted cardioverter defibrillator (ICD) in the Second Multicenter Automatic Defibrillator Implantation Trial. The cumulative probability of cause-specific death over time was estimated by the Kaplan-Meier method and by proportional hazards regression analysis. Early cardiac death survival curves were similar by treatment assignment in the 0- to 12-month period (p = 0.76). Late cardiac death survival curves by treatment assignment in the >12 to 52-month follow-up period were divergent with a lower probability of late cardiac death in the ICD arm compared with conventional therapy group (p <0.001). The time-specific hazard ratios of ICD to conventional therapy for cardiac death in the early and late periods were significantly different from each other (nominal p = 0.004). There was a significant decrease in sudden cardiac death with ICD therapy in the early (p = 0.012) and late (p <0.001) groups. In the early period, the rate of nonsudden cardiac death was significantly higher in the ICD group than in the conventional therapy group (p = 0.003). Rates of late nonsudden cardiac death were similar in the 2 treatment arms (p = 0.11). PMID- 15276095 TI - Differentiation of constrictive pericarditis from restrictive cardiomyopathy using mitral annular velocity by tissue Doppler echocardiography. AB - This study evaluated the diagnostic role of early diastolic mitral annular velocity (E') by tissue Doppler echocardiography for differentiating constrictive pericarditis from restrictive cardiomyopathy (primary restrictive cardiomyopathy and cardiac amyloidosis). The study group consisted of 75 patients (53 men, 22 women; mean age 62 years, range 27 to 87). Of these, 23 patients had surgically confirmed constrictive pericarditis, 38 had biopsy-proved systemic amyloidosis and typical echocardiographic features of cardiac involvement, and 14 had primary restrictive cardiomyopathy. Standard mitral inflow characteristics were measured. Tissue Doppler echocardiography was used to measure E' at the septal annulus. E' was significantly higher in patients with constrictive pericarditis than in those with primary restrictive cardiomyopathy or cardiac amyloidosis (12.3 vs 5.1 cm/second, p <0.001). An E' cut-off value > or =8 cm/second resulted in 95% sensitivity and 96% specificity for the diagnosis of constrictive pericarditis. There was no overlap of E' between patients who had constrictive pericarditis and those who had cardiac amyloidosis. In a subgroup analysis of restrictive cardiomyopathy, E' of patients who had cardiac amyloidosis was significantly lower than that of patients who had primary restrictive cardiomyopathy (4.6 vs 6.3 cm/second, p <0.001). Thus, E' velocity can distinguish between constrictive pericarditis and restrictive cardiomyopathy with a specific cut-off value in patients with clinical and echocardiographic evidence of diastolic heart failure. PMID- 15276096 TI - Steven Evan Nissen, MD: a conversation with the editor. Interview by William Clifford Roberts. PMID- 15276097 TI - Comparison of the extent and severity of coronary artery disease in patients with acute myocardial infarction with and without microalbuminuria. AB - The extent and severity of coronary atherosclerosis were compared between 43 microalbuminuric and 87 normoalbuminuric nondiabetic patients with acute myocardial infarction. Patients with microalbuminuria had significantly greater scores of the severity (Gensini score) and extent (Hamsten score) of coronary artery disease (p <0.001). PMID- 15276098 TI - Clinically unrecognized Q-wave myocardial infarction in patients with diabetes mellitus, systemic hypertension, and nephropathy. AB - During the Irbesartan Diabetic Nephropathy Trial, 1,387 participants with type 2 diabetes mellitus, hypertension, and nephropathy underwent serial electrocardiograms for the identification of Q-wave myocardial infarction (MI). During a mean follow-up of 2.5 years, 14 of 99 first nonfatal MIs in this group were clinically unrecognized, accounting for 14% of all first nonfatal MIs. PMID- 15276099 TI - Usefulness of myocardial contrast echocardiography in predicting global left ventricular functional recovery after anterior wall acute myocardial infarction. AB - Although multiple recent studies have shown that myocardial contrast echocardiography (MCE) reliably differentiates between regional stunning and necrosis after acute myocardial infarction (AMI), prognosis is more closely related to measures of global left ventricular systolic function. One hundred fifteen patients underwent baseline wall motion assessment and MCE 2 days after admission and follow-up echocardiography a mean of 69 days later. Good agreement was found between perfusion score index and follow-up wall motion score index, indicating that MCE performed early after anterior wall AMI may be clinically useful in routine post-AMI risk stratification. PMID- 15276100 TI - Clinical characteristics and thrombolysis in myocardial infarction frame counts in women with transient left ventricular apical ballooning syndrome. AB - The characteristics of 16 women with transient left ventricular (LV) apical ballooning syndrome in a United States population are presented. Additionally, Thrombolysis In Myocardial Infarction (TIMI) frame counts were evaluated during the acute period. Patients generally presented with anterior ST-elevation acute coronary syndrome in the absence of obstructive coronary disease. All patients had LV apical wall motion abnormalities. An acute emotional or physiologic stressor preceded most cases. TIMI frame counts were abnormal in all patients and often abnormal in all 3 major coronary vessels, suggesting that the diffuse impairment of coronary microcirculatory function may play a role in the pathogenesis of the syndrome. PMID- 15276101 TI - Exercise echocardiography with addition of atropine. AB - Exercise echocardiography with the addition of atropine was performed in 31 patients who had a very small probability of achieving submaximal heart rate. Conclusive tests were obtained in 77% of the patients with this protocol. PMID- 15276102 TI - Epidemiology and association of vascular and valvular calcium quantified by multidetector computed tomography in elderly asymptomatic subjects. AB - The epidemiology of and association between vascular and valvular calcium as quantified by multidetector computed tomography (MDCT) were studied in 416 elderly subjects with no history of coronary artery disease. Coronary calcium (CC), descending thoracic aortic calcium (DTAC), aortic valve calcium (AVC), and mitral valve calcium (MVC) were present in 282 (68%), 214 (51%), 152 (37%), and 68 (16%) subjects, respectively. Multiple logistic regression analysis showed that after adjusting for age and gender, subjects with AVC (odds ratio [OR] 2.3), MVC (OR 2.81), and DTAC (OR 2.79) were independently and significantly more likely to have CC. Further evidence is provided for the notion that calcifications in those regions are associated and that MDCT can be used as a tool for the global assessment of vascular and valvular calcium. PMID- 15276103 TI - Effectiveness of sirolimus-eluting stent implantation for treatment of in-stent restenosis after brachytherapy failure. AB - The impact of the use of sirolimus-eluting stents (SESs) in the treatment of in stent restenosis in previously irradiated sites has not been adequately evaluated. Fifteen consecutive patients who underwent percutaneous coronary interventions using SESs in lesion sites previously intervened with intracoronary radiation therapy were identified. All stents were implanted successfully, and there were no major in-hospital complications. At 30-day follow-up, there was 1 case of subacute thrombosis that led to target lesion revascularization (TLR). At 6 months, 2 patients underwent TLR because of recurrent angina with angiographic restenosis, and 1 patient underwent target vessel revascularization distally to the SES site; no other major adverse cardiac events occurred at long-term follow up (mean 17 +/- 8 months). PMID- 15276104 TI - Assessment of biventricular remodeling by magnetic resonance imaging after successful primary stenting for acute myocardial infarction. AB - Inferior acute myocardial infarction (AMI) is associated with a better outcome compared with anterior AMI, even in the presence of comparable infarct size. Whether left ventricular remodeling, a major predictor of poor outcome, and right ventricular (RV) remodeling depend on the site of an AMI remains unknown. Biventricular volumes were assessed by magnetic resonance imaging 7 +/- 2 days and 3.4 +/- 0.3 months after successful primary stenting in 51 consecutive patients with inferior or anterior AMI. This study documents RV involvement and biventricular reverse remodeling in patients with inferior AMI in the absence of RV infarction, as opposed to those with anterior AMI who show progressive biventricular remodeling. PMID- 15276105 TI - Effect of clopidogrel pretreatment on periprocedural rise in C-reactive protein after percutaneous coronary intervention. AB - This study sought to determine the effect of clopidogrel pretreatment on the increase in C-reactive protein (CRP) after percutaneous coronary intervention. Clopidogrel pretreatment attenuated the periprocedural increase in CRP by 65% and was independently associated with an attenuation in the CRP increase in a multivariate model. PMID- 15276106 TI - Comparison of femoral bleeding complications after coronary angiography versus percutaneous coronary intervention. AB - Vascular complications arising after coronary angiography (CA) and percutaneous coronary intervention (PCI) may be under-reported. Access site complications were studied after consecutive CA and PCI in the investigators' center (an urban university hospital) from October to December 2002. Three hundred eleven consecutive procedures (237 CA studies and 74 PCIs) were included, of which 309 (99%) involved femoral arterial access. Seventy percent of all arterial punctures were closed by manual pressure, 28% by C-pressure clamps, and 2% with closure devices. Femoral hematomas occurred in 22% and 41% of CA studies and PCIs, respectively. Hematoma >5 cm occurred in 6% and 11% of CA studies and PCIs, respectively. Nineteen patients (23%) with access site complications had prolonged hospital admission. Increased body mass index and hematoma development within the catheter laboratory were predictors of prolonged admission. PMID- 15276107 TI - Effect of atorvastatin on the expression of CD40 ligand and P-selectin on platelets in patients with hypercholesterolemia. AB - The effects of atorvastatin on levels of the CD40 ligand (CD40L or CD154) and P selectin on platelets were investigated in patients with hypercholesterolemia. The major finding was that short-term atorvastatin treatment (8 weeks) in a group of hypercholesterolemic patients resulted in significant suppression of CD40L and P-selectin expression. Moreover, there was a significant correlation between the magnitude of CD40L downregulation and that of very low-density lipoprotein cholesterol, the ratio of low-density lipoprotein cholesterol to high-density lipoprotein cholesterol, and triglycerides. In hypercholesterolemic patients, in addition to its effects on decreasing cholesterol, atorvastatin can intervene in the interaction of CD40-CD40L and the expression of P-selectin on platelets. Thus, interference of CD40-CD40L can be recognized as an integral part of the anti-inflammatory activity of atorvastatin in hypercholesterolemia. PMID- 15276108 TI - Comparison of low risk and higher risk profiles in middle age to frequency and quantity of coronary artery calcium years later. AB - Little is known about the relation of having favorable levels of all major cardiovascular risk factors (low risk [LR]) earlier in life to coronary artery calcium (CAC) later in life. From 2002 to 2003, CAC was compared in participants aged >60 years who were LR (n = 42) with those not LR (n = 39) at baseline (from 1967 to 1973). Despite adverse changes in risk factors, the prevalence of measurable CAC and mean CAC scores were less for LR participants than for non-LR participants (60% vs 77%, p = 0.09, and 217 vs 443, p = 0.05, respectively). PMID- 15276109 TI - Effect of body mass index on defibrillation thresholds for internal cardioversion in patients with atrial fibrillation. AB - This study reports that patients with large body mass indexes (BMIs) and long duration atrial fibrillation (AF) have greater defibrillation thresholds than patients with normal BMIs and AF of short duration. BMI is an independent predictor of defibrillation thresholds in patients with longer duration AF in whom external cardioversion has failed. PMID- 15276110 TI - Age- and sex-related atrial electrophysiologic and structural changes. AB - Age and gender dependence of the 3 major variables--left atrial (LA) dimension, effective refractory period (ERP), and atrial conduction--that govern the development of atrial fibrillation (AF) were examined in 76 women and 39 men without structural heart disease. A significant positive correlation was found between age and LA dimension, mainly because of a strongly positive correlation in women not taking hormone replacement therapy but not in women on hormone replacement therapy. Men had a significantly greater average LA dimension than women. Neither ERP nor atrial conduction estimated by normalized P-wave duration demonstrated any significant correlation with either age or gender. It was concluded that LA size is greater in the elderly and in men, which may increase their risk for AF. PMID- 15276111 TI - Effectiveness of early implantation of cardioverter defibrillator for postoperative ventricular tachyarrhythmia. AB - The effectiveness of implantable cardioverter defibrillators (ICDs) implanted in the early postoperative period after cardiac surgery for ventricular tachyarrhythmias is unknown, because all of the major trials excluded this patient population. Thus, a 10-year retrospective study was conducted of patients who had ICDs implanted for de novo postoperative ventricular tachyarrhythmias during the index admission for cardiac surgery. There was a high rate of early recurrence of ventricular tachyarrhythmia treated by defibrillators, and this finding questions the exclusion of this important patient population from large trials. PMID- 15276112 TI - Efficacy of distinct energy delivery protocols comparing two biphasic defibrillators for cardiac arrest. AB - Limited data have been published on the use of external defibrillators that deliver impedance compensated biphasic (ICB) waveforms in patients. We compared 2 ICB defibrillators, the Heartstream XL (150-150-150 J protocol) and Heartsine Samaritan (100-150-200 J protocol) in 78 consecutive patients in cardiac arrest. The performance of the 2 devices over the first 2 shocks was statistically equivalent. By the third shock, the Heartsine Samaritan had significantly better performance in removing ventricular fibrillation (p = 0.029). Energy selection for ICB waveforms requires further validation. PMID- 15276113 TI - A likely explanation for the J-curve of blood pressure cardiovascular risk. AB - We prospectively tested in the combined original and offspring Framingham cohorts the hypothesis that the increase in cardiovascular disease (CVD) incidence at low diastolic blood pressure (BP) is largely confined to subjects with increased systolic BP and hence an increased pulse pressure. The 10-year risk of 951 nonfatal CVD events and 204 CVD deaths was estimated at diastolic pressures of <80, 80 to 90, and > or =90 mm Hg, according to concomitant systolic BP. An increasing tendency for a J-curve relation of CVD incidence to diastolic BP was observed with successive increments in accompanying systolic BP. In both genders, a statistically significant excess of CVD events was observed at a diastolic BP of <80 mm Hg only when accompanied by a systolic BP of >140 mm Hg that persisted after adjustment for age and associated CVD risk factors. Patients with this condition of isolated systolic hypertension have been shown to benefit from antihypertensive treatment. PMID- 15276114 TI - Insulin-like growth factor-I gene polymorphism and risk of heart failure (the Rotterdam Study). AB - We studied 4,963 participants of the population-based Rotterdam Study and found that a genetically determined chronic exposure to low insulin-like growth factor I (IGF-I) levels is associated with an increased risk for heart failure in elderly patients. PMID- 15276115 TI - Age-related prevalence of cardiac valvular abnormalities warranting infectious endocarditis prophylaxis. AB - The goal of our study was to determine the prevalence of older patients with cardiac valvular abnormalities warranting endocarditis prophylaxis. We performed a retrospective analysis of 1,000 randomly selected echocardiograms (inpatients and outpatients) from our tertiary care institution. We found that the prevalence of valvular abnormalities increased significantly with age, and that 50% of patients > or =60 years of age warranted endocarditis prophylaxis using current guidelines. With the aging population of the United States and the negative consequences of widespread antibiotic prophylaxis, further investigation is needed to identify patients who are truly at risk for infectious endocarditis. PMID- 15276116 TI - Comparison of echocardiographic features of noncompaction of the left ventricle in adults versus idiopathic dilated cardiomyopathy in adults. AB - Noncompaction of left ventricular myocardium (NCLV), or "spongy myocardium," in adults represents an arrest in endomyocardial morphogenesis and occurs as an isolated cardiomyopathy. Because NCLV can be readily mistaken for idiopathic dilated cardiomyopathy, echocardiographic features other than the structural features of the myocardial wall need to be carefully defined for distinguishing the 2 conditions. This study was therefore designed to characterize the echocardiographic features that could be useful for differentiating NCLV from idiopathic dilated cardiomyopathy. PMID- 15276117 TI - Endothelial cell function in patients with Down's syndrome. AB - Patients with Down's syndrome show an increased pulmonary vascular reactivity that could be due to an impaired vascular endothelial function, which is possibly related to increased oxidative stress. In 8 patients with Down's syndrome and 9 euploid patients of similar age, endothelium-dependent and -independent vasodilation was studied, measuring brachial flow velocity with an intravascular Doppler flow wire. Patients with Down's syndrome showed a significant impairment of endothelial function versus controls. In presence of the antioxidant vitamin C, endothelium-dependent vasodilation in the patients with Down's syndrome was only slightly, but not significantly, improved. PMID- 15276118 TI - Comparison of left ventricular function by tissue Doppler imaging in patients with diabetes mellitus without systemic hypertension versus diabetes mellitus with systemic hypertension. AB - Because diabetes mellitus substantially increases the risk of development of heart failure, we sought to establish early alterations in left ventricular systolic and diastolic function in patients with diabetes mellitus with and without coexisting systemic hypertension. We studied 134 subjects using echocardiography comprising standard 2-dimensional and conventional Doppler as well as tissue Doppler imaging. Our study demonstrated the early appearance of both left ventricular systolic and diastolic dysfunction in diabetic patients at rest and the contributory effects of diabetes to myocardial impairment produced by hypertension, as well as the high usefulness of tissue Doppler imaging in detection and quantitation of myocardial dysfunction in diabetics. This method was superior to other echocardiographic techniques and plasma brain natriuretic peptide evaluation. PMID- 15276119 TI - Comparison of aortic dissection in patients with and without Marfan's syndrome (results from the International Registry of Aortic Dissection). AB - Among 1,049 patients diagnosed with aortic dissection in a multinational registry, patients with Marfan's syndrome were typically younger and had unique presentations. Despite their younger age at presentation, patients with Marfan's syndrome and aortic dissection had a high mortality rate, similar to patients without Marfan's syndrome, an older patient cohort. Our data support the importance of aneurysm surveillance and prophylactic surgical intervention for patients with Marfan's syndrome to potentially reduce the risk of mortality. PMID- 15276120 TI - Time trends in warfarin-associated hemorrhage. AB - The annual incidence of warfarin-related bleeding at Brigham and Women's Hospital increased from 0.97/1,000 patient admissions in the first time period (January 1995 to October 1998) to 1.19/1,000 patient admissions in the second time period (November 1998 to August 2002) of this study. The proportion of patients with major and intracranial bleeding increased from 20.2% and 1.9%, respectively, in the first time period, to 33.3% and 7.8%, respectively, in the second. PMID- 15276121 TI - Usefulness of a hand-carried cardiac ultrasound device for bedside examination of pericardial effusion in patients after cardiac surgery. AB - To identify the incidence of pericardial effusion in patients after cardiac surgery using a hand-carried cardiac ultrasound device, 200 patients were assessed on postoperative day 3. If a pericardial effusion was found, patients were monitored for 3 consecutive days with a hand-carried cardiac ultrasound device. Within 72 hours after surgery, 43 patients (21.5%) had developed an effusion, of whom 2 patients had cardiac tamponade and 41 patients (21%) had a small pericardial effusion. No difference was found in the incidence of effusion based on the type of cardiac surgery. Of patients with a small pericardial effusion on day 3 after surgery, an additional 2 of 41 (5%) developed cardiac tamponade. PMID- 15276122 TI - Utility and diagnostic accuracy of hand-carried ultrasound for emergency room evaluation of chest pain. AB - This study examined the utility and accuracy of immediate hand-carried echocardiography in patients presenting to the emergency room with chest pain and a normal or nondiagnostic electrocardiogram. Hand-carried echocardiography was highly concordant (kappa = 0.8) with troponin T tests as well as the discharge diagnosis of acute coronary syndrome, had a 100% sensitivity for the detection of acute coronary syndrome, 93% specificity, and 71% and 100% positive and negative predictive values, respectively. PMID- 15276123 TI - Salvage of nondiagnostic transthoracic echocardiograms on patients in intensive care units with intravenous ultrasound contrast. AB - Transthoracic echocardiography (TTE) is often technically difficult on patients in intensive care units. Contrast echocardiography can improve left ventricular wall visualization and the assessment of regional and global left ventricular wall motion. Our study undertook to determine what proportion of nondiagnostic TTE studies on patients in intensive care units could be salvaged (i.e., converted to diagnostic studies) with contrast. Ninety-two patients with nondiagnostic TTEs had a repeat study after contrast. Using predefined criteria, 51% of studies were salvaged with contrast. Female gender emerged as the only factor associated with less likelihood of salvaging a study. PMID- 15276124 TI - Two levels of block within a pulmonary vein. AB - We present a 37-year-old woman with drug refractory atrial fibrillation referred for a pulmonary vein (PV) isolation procedure who manifested a tachycardia with 2:1 intra-PV block within a dissociated PV. During ablation in the right superior PV, the surface rhythm became normal sinus, with persistent atrial fibrillation within the PV. Shortly thereafter, an atrial tachycardia with a cycle length of 190 ms and 2:1 distal to proximal exit block was observed within the isolated PV. Conduction block within a PV electrically isolated from the left atrium is a newly observed phenomenon that may have implications to the electrophysiologic properties of the PV muscular sleeves. PMID- 15276125 TI - Significant statistical heterogeneity in a meta-analysis of the usefulness of acetylcysteine for prevention of contrast nephropathy. PMID- 15276127 TI - Genes and genetics in respiratory control. AB - The genetic approach to respiratory control is opening up new paths for research into developmental respiratory control disorders. Despite the identification of numerous genes involved in respiratory control, none of the genetically engineered mice developed to date fully replicate the human respiratory phenotype of human developmental respiratory disorders. However, combining studies in humans and studies in mouse models has proved useful in identifying candidate genes for human developmental respiratory control disorders and providing pathogenic information. In clinical practice, the development of databases that incorporate clinical phenotypes and genetic samples from patients would facilitate further genetic studies. International multicentre studies would advance the area of respiratory control research. PMID- 15276128 TI - Developmental and metabolic implications of the hypoxic ventilatory response. AB - This review explores the evidence to support the leading hypothesis that the metabolic response to hypoxia early in life provides the pathophysiological basis for the metabolic syndrome. Hypoxia is a frequent occurrence during early development and induces a state of energy depletion that triggers a wide range of 'metabolic' responses to preserve homeostasis. Recent interest in the sequelae of energy depletion through hypoxic mechanisms has grown, particularly because of demonstrated links with ensuing metabolic abnormalities and increased risk for future cardiovascular disease. The 'metabolic syndrome' refers to the combination of obesity, hyperinsulinaemia, dyslipidaemia and hypertension in adults. The metabolic responses to energy depletion during early development provide explanations for some of the mechanisms that ultimately lead to serological features of metabolic dysfunction in children with sleep-disordered breathing. Thus, the acute compensatory response of energy conservation to hypoxia during early development at the cellular, serological and whole organism levels suggests that the metabolic abnormalities that develop later in life may in fact originate very early in life; in other words, constitute early life antecedents of adult disease. Evidence regarding the circumstances under which responses to hypoxia become maladaptive will be discussed, with a focus on chronic conditions and those associated with intermittent respiratory dysfunction such as sleep disordered breathing. PMID- 15276129 TI - Congenital central hypoventilation syndrome: not just another rare disorder. AB - Congenital central hypoventilation syndrome (CCHS) is a rare syndrome, present from birth, and is defined as the failure of automatic control of breathing. Patients have absent or negligible ventilatory sensitivity to hypercapnia and hypoxaemia during sleep and wakefulness. Therefore, especially while asleep, children with CCHS experience progressive hypercapnia and hypoxaemia. They lack arousal responses and sensations of dyspnoea to the endogenous challenges of isolated hypercapnia and hypoxaemia and to the combined stimulus of hypercapnia and hypoxaemia. Patients with CCHS do not exhibit signs of respiratory distress when challenged with hypercarbia or hypoxia. The diagnosis is one of exclusion, ruling out any primary pulmonary, cardiac, metabolic or neurologic cause for central hypoventilation. CCHS is associated with other manifestations of autonomic nervous system dysfunction, including Hirschsprung's disease. All patients with CCHS require lifelong ventilatory support during sleep but some will be able to maintain adequate ventilation without assistance while awake once past infancy. However, some CCHS patients require ventilatory support for 24h/day. Modalities of home mechanical-assisted ventilation include positive pressure ventilation via tracheostomy, non-invasive positive pressure ventilation (bi-level ventilation), negative pressure ventilation and diaphragmatic pacers. Supplemental oxygen alone is inadequate treatment. With early diagnosis and adequate ventilatory support, these children can have good outcomes and lead productive lives. PMID- 15276130 TI - Respiratory control and arousal in sleeping infants. AB - Control of the cardiovascular and respiratory systems undergoes rapid maturation during infancy. Sleep is at a lifetime maximum during this period and has a marked influence on cardiorespiratory function. The mechanisms leading to sudden infant death syndrome (SIDS) may include a failure in the neural integration of the cardiovascular and respiratory systems, with a concomitant failure to arouse from sleep. Studies have shown that sleep states exert a marked influence on respiratory control and arousability. Infants are more arousable in active sleep compared with quiet sleep from both somatosensory and respiratory stimuli. Post natal and gestational age at birth also have a marked influence on arousability. Arousability is depressed by the major risk factors for SIDS (prone sleeping, maternal smoking, prematurity and recent infection) and is increased by factors that decrease the risk for SIDS (e.g. use of dummies, breastfeeding). PMID- 15276131 TI - Difficult asthma in the pre-school child. AB - The most important aspect of dealing with a pre-school child suspected of having difficult asthma, is to ensure that the diagnosis is correct, in order to avoid the inappropriate use of therapies such as inhaled corticosteroids. After exclusion of other diagnoses, if a pre-school child is thought to have asthma, difficult or otherwise, the corollary is, what sort of asthma? Is it a syndrome with airway inflammation susceptible to treatment, or one in which there is no inflammation and time alone will result in resolution of symptoms? Probably the most common mistake in this age group is to fail to recognise the latter and institute ever more aggressive and useless therapies. An approach to excluding other diagnoses, appropriate investigations to elicit the presence of airway inflammation and suggestions for subsequent management have been detailed in this review. PMID- 15276132 TI - Improving asthma outcomes in harder-to-reach populations: challenges for clinical and community interventions. AB - The burden of asthma differs from country to country and within populations. The factors that influence this variation include asthma prevalence and severity, aspects of healthcare services (such as accessibility, quality and utilisation) and social demographic factors (such as income inequality, cultural and linguistic diversity and indigenous populations). The identification of individuals and populations that are 'harder to reach', 'special' or at greater risk of poor asthma outcomes therefore depends on how the burden of asthma and its management are measured. Meeting the challenge of educating harder-to-reach populations with asthma is the focus of this article. In clinical settings, communication is the mainstay of engaging and building trust to influence behavioural change in individuals; community-based interventions provide valuable opportunities for targeting harder-to-reach populations. However, they require active community consultation and participation and innovative approaches to service delivery, in addition to being theoretically driven and systematically developed. PMID- 15276133 TI - Psychological approaches to the management of respiratory symptoms in children and adolescents. AB - Acute respiratory symptoms often demand immediate medical attention and can be a frightening experience for parents and children alike. Once immediate medical concerns have been dealt with, the physician may be left with a complex clinical picture of uncertain aetiology. Vocal cord dysfunction, hyperventilation, sighing dyspnoea and psychogenic cough are four well-described clinical syndromes where the aetiology may not be clear. Indeed for a busy clinician, it may also be unclear how to manage these patients and when to refer for more detailed psychological or psychiatric management. This paper will discuss the management of complex cases where a range of factors may influence outcome. A framework is presented that can be used to aid formulation and intervention. It is argued that the challenge to the medical practitioner is to combine a medical assessment with careful attention to the personal perceptions and experiences of patients' symptoms. In paediatric and adolescent medicine, particular attention needs to be paid to the wider context of the family, as well as to communication within the consulting room. A clinical example is presented that highlights common dilemmas and illustrates the use of solution-focused brief therapy and motivational interview techniques. PMID- 15276134 TI - Tuberculosis in HIV-infected children. AB - Tuberculosis (TB) is the most common opportunistic infection in human immunodeficiency virus (HIV)-infected people worldwide. HIV-positive children are at risk of diagnostic error as well as delayed diagnosis of TB because of overlapping clinical and radiographic features with other lung diseases. Acute pneumonias and chronic lung diseases such as bronchiectasis and lymphocytic interstitial pneumonitis are difficult to distinguish from TB. TB manifestations are more severe in HIV-positive children and progression to death is more rapid than in HIV-negative children. The response to standard short-course therapy in HIV-positive children is not as good as in HIV-negative children due to lower cure rates and higher mortality. TB hastens the progression of HIV disease by increasing viral replication and reducing CD4 counts further. Although Bacille Calmette-Guerin vaccination could lead to disseminated Mycobacterium bovis disease in the presence of immunosuppression, this has been rarely reported. More studies are required to assess the role of newer diagnostic tests, TB preventive therapy and co-administration of anti-retroviral therapy in the control of TB among HIV-infected children. PMID- 15276135 TI - Novel approaches in conventional mechanical ventilation for paediatric acute lung injury. AB - Acute lung injury remains a major cause of morbidity and mortality in paediatric intensive care units. Research over the past decade has altered our understanding of the pathophysiology of acute lung injury and the effects of mechanical ventilation on the lung. As a result, approaches to conventional mechanical ventilation of the injured lung are now largely centred around preservation of adequate gas exchange while protecting the lung from further ventilator-induced lung injury. Current techniques for accomplishing these goals include adjusting the ventilator based on the measurement and interpretation of pressure-volume curves, limitation of inspiratory tidal volumes, use of elevated levels of positive end-expiratory pressure, recruiting manoeuvres and prone positioning. The currently available data regarding the efficacy and appropriate use of these techniques are reviewed. PMID- 15276136 TI - Paediatric pulmonary vascular disease. AB - Pulmonary vascular disease comprises any congenital or acquired pathology of the intrinsic pulmonary vessels with the unique feature of pulmonary arteries carrying unsaturated blood and pulmonary veins carrying oxygenated blood. Pulmonary hypertension (PH) ensues when pulmonary vascular disease affects at least 50% of the capillary resistance vessels, i.e. pulmonary pre-acinar and intra-acinar arteries (so-called pre-capillary PH), or when pressure in the pulmonary venous system distal to the capillaries rises above a mean of 15 mmHg (so-called post-capillary PH). PH is defined by a mean pulmonary arterial pressure above 25 mmHg at rest. Vasoconstriction, remodelling and thrombosis of small pulmonary arteries lead to an increase in pulmonary vascular resistance. The consequence is failure of the afterload-intolerant right ventricle. In this review, pulmonary vascular disease in children will be addressed according to the 2003 World Health Organisation (Venice) classification of PH. PMID- 15276137 TI - An overview of paediatric lung transplantation. AB - Paediatric lung transplantation is indicated in selected children with end-stage lung disease that is not amenable to conventional medical or surgical therapy. The indications and complications differ from adult lung transplant patients. Due to the long waiting times for suitable cadaveric lungs, other types of lung transplantation, such as living donor lobar and split-lung procedures, have been utilised in paediatric patients. Unlike adult candidates, cystic fibrosis and primary pulmonary hypertension are the primary indications. Most recipients are in the adolescent age group. Complications that occur with greater frequency in paediatric lung recipients include somatic growth and graft function, post transplant lymphoproliferative disease and medical non-adherence. While long-term outcome remains similar between adult and paediatric lung transplant recipients, there is a lower risk of bronchiolitis obliterans in very young recipients and in those who receive living donor lobar lung transplantation. Research into these clinical problems is hampered by the fact that only a small number of paediatric transplants are performed at each centre. Hence, improvement in outcome for these children will be dependent on developing methods to produce better tolerance, understanding the mechanisms/treatment of bronchiolitis obliterans and multi centre studies that focus on the problems that primarily affect the paediatric lung transplant recipient. PMID- 15276138 TI - Mechanisms of rhinovirus-induced asthma. AB - Several epidemiological studies using sensitive detection methodologies have confirmed that the majority of acute asthma exacerbations follow upper respiratory tract infections--common colds. Most of these colds are due to human rhinoviruses (RVs). RVs are able to reach and replicate in epithelial cells of the lower airways and can activate these cells to produce pro-inflammatory mediators. Under some circumstances, RVs can also become cytotoxic to the epithelium. Atopic asthmatic individuals produce less interferon-gamma and more interleukin-10 than normal subjects in response to RV infection. Symptom severity as well as viral shedding after experimental RV infection, is inversely correlated with 'atopic' status, expressed as the interferon-gamma to interleukin 5 ratio. Expression of co-stimulatory molecules on immune cells is also affected in atopic asthmatics, suggesting an aberrant immune response to RV that may lead to suboptimal viral clearance and viral persistence. Some of the above effects can be reversed in vitro by corticosteroids, second-generation antihistamines or anti-oxidants; however, the optimal strategy for treating acute asthma exacerbations requires further research at both mechanistic and clinical levels. PMID- 15276139 TI - Abnormal quantitative EEG scores identify patients with complicated idiopathic generalised epilepsy. AB - OBJECTIVE: To investigate the relationship between quantitative EEG (QEEG) scores and "complicating factors" (psychopathology, true pharmacoresistance, neurological symptoms) in idiopathic generalised epilepsy (IGE). METHODS: 35 newly referred, newly diagnosed, unmedicated IGE patients were collected in a prospective and random manner. Standard neuro-psychiatric and EEG examination was done. The patients were treated and controlled at regular visits. After 2 years of follow-up, clinical data were summarised and were compared to QEEG results. Clinical target items were neurologic and psychiatric abnormalities, proven pharmacoresistance. Patients with at least one of these items were labelled "complicated", whereas patients without these additional handicap were labelled as "uncomplicated". The 12 QEEG target variables were: Z-transformed absolute power values for three (anterior, central, posterior) brain regions and four frequency bands (1.5-3.5; 3.5-7.5; 7.5-12.5; 12.5-25.0 Hz). QEEG scores outside the +/- 2.5 Z range were accepted as abnormal. The overall QEEG result was classified as normal (0-2 abnormal scores), or pathological (3 or more abnormal scores). Clinical and QEEG results were correlated. RESULTS: All patients with psychopathology showed 4-8 positive pathological scores (power excess not confined to a single cortical region or frequency band). The two patients with pure pharmacoresistance showed pathological negative values (delta power deficit) all over the scalp. Statistically significant (P < 0.001) association was found between patients with uncomplicated IGE and normal QEEG, and between complicated IGE and pathological QEEG. Patients with neurological items had normal QEEG. CONCLUSION: Higher degree of cortical dysfunction (as assessed in the clinical setting) is reflected by higher degree of QEEG abnormalities. QEEG analysis can differentiate between IGE patients with or without psychopathology. Forecasting psychopathology may be the practical application of the findings. PMID- 15276140 TI - The relationship between illness severity, sociodemographic factors, general self concept, and illness-specific attitude in Swedish adolescents with epilepsy. AB - PURPOSE: The aim of this study was to describe the relationship between the epilepsy condition (illness severity), sociodemographic factors, general self concept, and illness-specific attitude in adolescents with uncomplicated epilepsy. METHODS: Adolescents, aged 13-22, fulfilling criteria registered in four Swedish hospitals, answered questionnaires (n = 149). The instruments "I think I am" and "Sense of coherence" measured the patients' general self-concept. The "Child Attitude Toward Illness Scale" measured illness-specific attitude. A summary score (index) calculated from seizure frequency, seizure type, and antiepileptic drug (AED) with side effects measured "Illness Severity". RESULTS: Illness severity was significantly related to the participants' general self concept, as well as to their attitude toward their condition; i.e. higher illness severity scores were correlated with lower sense of coherence (SOC), poorer self esteem, and a more negative attitude towards the epilepsy condition. Females had more severe illness according to the Illness Severity Index, with almost 80% found in the moderate and high severity groups as compared to 63% of males in the moderate/high severity groups. CONCLUSIONS: It was concluded that the severity of the epilepsy condition was related to the adolescents' general self-concept and illness-specific attitude, but further research is needed to understand the causality of the relationship. The brief assessment of illness severity, constructed and used in this study should be addressed and developed further. PMID- 15276141 TI - Epilepsy and destructive brain insults in early life: a topographical classification on the basis of MRI findings. AB - Destructive insults of early development can lead to a wide variety of lesional patterns and are a well known cause of epilepsy. The aim of this study is to present a topographic magnetic resonance imaging (MRI) classification of these lesions in adult patients with epilepsy. Thirty-three consecutive patients were divided in three groups according to the topographic distribution of their lesion on MRI: hemispheric (H, n = 7); main arterial territory (AT, n = 18); arterial borderzone (Bdz, n = 8). We analyzed clinical, MRI and magnetic resonance angiography (MRA) data. Status epilepticus (SE) during childhood was more common in group H (7/7) than in the groups AT (1/18) and Bdz (0/8) (P < 0.001). MRA pattern of impaired flow signal in the distal segments of all three major arteries in the affected hemisphere was present in 85.7% of group H patients, and was exclusive to this group. 88.8% (16/18) of patients from group AT presented congenital motor deficit, in contrast to 37.5% (3/8) of group Bdz, and in none of group H (P < 0.001). All patients with Bdz lesions had antecedent of fetal distress, in contrast to 1/7 from group H and 5/18 of group AT (P = 0.001). The MRAs of patients with Bdz lesions were often normal except in those with larger lesions. Our data suggest that in adult patients with epilepsy due to precocious destructive brain insults, a MRI topographical classification distributes them in relatively homogenous clinical groups. PMID- 15276142 TI - Vagus nerve stimulation for 1 year in 269 patients on unchanged antiepileptic drugs. AB - PURPOSE: To study, in patients on unchanged antiepileptic drugs (AEDs): (1) seizure rates after 3 and 12 months of vagus nerve stimulation (VNS); (2) effects of VNS parameters; (3) patient characteristics versus VNS responsiveness. METHODS: We located in the VNS registry 269 patients treated for 1 year with no changes in AEDs. Seizure rates were calculated at 3 and 12 months. We analyzed: (1) 3 months versus 12 months seizure rates; (2) effects of changing duty cycles between 3 and 12 months; (3) effects of output current; (4) seizure rate changes associated with patient characteristics. RESULTS: Seizure rates improved between 3 months (median = -45%) and 12 months (median = -58%) (P < 0.0001). There were no differences between patients who stayed on standard or rapid cycling, or changed from standard to rapid. Stimulation parameters did not affect seizure rates. VNS responsiveness was associated with older age (P = 0.016), longer duration epilepsy (P = 0.033), and syndromes other than Lennox-Gastaut (P = 0.003). CONCLUSIONS: This was an analysis of treatment outcomes, not a prospective clinical trial. Nonetheless, our results suggest: (1) seizure rates decline with increasing VNS duration; (2) this decline occurs without AED changes; (3) this decline is not due to changes in stimulation parameters; (4) patient characteristics predictive of VNS responsiveness remain elusive. PMID- 15276143 TI - Determination of language dominance with near-infrared spectroscopy: comparison with the intracarotid amobarbital procedure. AB - PURPOSE: To determine the efficacy of near-infrared spectroscopy (NIRS) in identifying the hemisphere associated with language by measuring changes in bilateral lateral inferior frontal blood flow during a word generation task in epilepsy surgery patients and healthy volunteers. METHODS: Sixteen patients who underwent the intracarotid amobarbital procedure (IAP) and eight healthy right handed/right-footed individuals were tested with NIRS during a word generation paradigm. RESULTS: Increases in lateral inferior frontal total hemoglobin concentrations agreed with the IAP in 11 of 16 patients, including 2 of 3 with right hemisphere (atypical) speech dominance (P = 0.13). NIRS revealed increases in lateral inferior frontal total hemoglobin concentrations congruent with language dominance predicted by handedness/footedness in 18 of 24 subjects, including 100% of healthy right-handed subjects (P = 0.02). CONCLUSION: NIRS is a simple, non-invasive, safe modality for measuring cerebral blood flow. Further research and development of testing procedures and instrumentation are needed before routine implementation in pre-surgical testing. PMID- 15276144 TI - The localizing and lateralizing value of ictal/postictal coughing in patients with focal epilepsies. AB - Postictal coughing has so far been reported to indicate a temporal origin of focal epilepsy. A trend towards non-dominant hemisphere lateralization and mesial temporal localization has been suggested. However, postictal coughing has also been reported in a few patients with extratemporal epilepsies. We have retrospectively evaluated the localizing and lateralizing value of ictal/postictal coughing in 197 patients with temporal and extratemporal epilepsy who received presurgical video-EEG long-term recordings from 1999 to 2001. There was no statistical significant difference in percentage of coughing patients in both groups. However, only patients belonging to the temporal group presented with coughing as a regular element of seizure semiology (simple partial and complex partial seizures) whereas in the extratemporal group coughing occurred more sporadically. Within the temporal group a statistically significant tendency to left-sided seizure onset and a statistically not significant preponderance of mesial seizure onset was observed. Additional vegetative signs were observed only in about half of the patients. These results suggest that coughing occurs in both temporal and extratemporal lobe epilepsy and may only be indicative of temporal lobe seizure onset if representing a regular semiologic element. Coughing may be due to two different mechanisms, one dependent and the other independent from additional vegetative symptoms. PMID- 15276145 TI - Color vision in epileptic adolescents treated with valproate and carbamazepine. AB - OBJECTIVE: The aims of our study were to evaluate whether deficits in color vision exist in epileptic adolescents, to study if monotherapy with valproic acid (VPA) and carbamazepine (CBZ) can affect color vision, and to determine the possible relationship between abnormal color vision tests and AEDs dosage and their serum concentrations. PATIENTS: We examined 45 epileptic patients before the beginning of therapy and after 1 year of VPA or CBZ monotherapy and 40 sex- and age-matched healthy controls. METHODS: Color vision was evaluated with Farnsworth Munsell 100 (FM100) hue test and achromatic and short-wavelength automated perimetry (SWAP). STATISTICAL ANALYSIS: To evaluate intergroup differences we used ANOVA with Scheffe's post hoc test, when appropriate. Repeated measures ANOVA was used to evaluate the intragroup modifications of total error score (TES) and perimetric threshold during the follow-up. Pearson's correlation test was performed to correlate chromatic sense and perimetric data and AEDs dosage and serum concentrations. RESULTS: Before the beginning of therapy, there were no differences in central color vision and SWAP between controls and epileptic patients. After 1 year, patients treated with VPA or CBZ showed a deficit in FM100 hue test and SWAP parameters while no significant deficit was found in achromatic perimetry. In particular, with the FM100 hue test a higher number of errors was found in both groups of patients (CBZ patients: 166.00 +/- 27.72 TES; VPA patients: 151.19 +/- 44.09, P < 0.001) in comparison with controls (controls: 109.29 +/- 24.73) and baseline values (CBZ patients: 110.65 +/- 22.9; VPA patients 107.43 +/- 21.70). With SWAP patients of both groups showed significant variation of foveal threshold (controls: 21.07 +/- 2.01 dB; CBZ patients: 19.35 +/- 1.32, P < 0.001; VPA patients: 18.88 +/- 1.89, P < 0.001), full-field mean threshold perimetric sensitivity (controls: 18.50 +/- 1.24 dB; CBZ patients: 16.60 +/- 1.47, P < 0.001; VPA patients: 16.23 +/- 1.55, P < 0.001) and mean threshold perimetric sensitivity of the three evaluated subareas of the visual field (area 1 controls: 21.01 +/- 1.15; CBZ patients: 19.45 +/- 1.74, P = 0.001; VPA patients: 18.25 +/- 1.61, P < 0.001; area 2 controls: 18.40 +/- 1.43; CBZ patients: 16.07 +/- 1.58, P +/- 0.001; VPA patients: 16.13 +/- 1.46, P = 0.001; area 3 controls: 17.20 +/- 1.49; CBZ patients: 14.28 +/- 1.51, P < 0.001; VPA patients: 14.31 +/- 2.90, P = 0.001). CONCLUSIONS: Our study demonstrates that treatment with VPA or CBZ can affect significantly both central and paracentral color vision after a short treatment period. PMID- 15276146 TI - Effects of lateralisation and gender on temporal lobe ictal behaviour associated with hippocampal sclerosis. AB - INTRODUCTION: Information derived from animal models and neuroradiological studies in humans indicates that males and females exhibit differences in the functional and anatomical organisation of the brain. This study aimed to evaluate the effect of gender in ictal behaviour considering lateralisation in a group with homogeneous pathology. METHODS: Patients with hippocampal seizures who underwent temporal lobectomy and who were seizure-free during one year of follow up were selected. Surgery was performed on the right side in 27 patients and on the left side in 21. Videotape recordings of the patients were reviewed in order to investigate ictal behaviour. There were 42 seizure episodes in 20 males and 40 in 21 females. For auras, 48 patients' data were reviewed. Ictal behaviour was evaluated taking into consideration the lateralisation of seizures and gender differences. RESULTS: Ictal vocalisation was significantly higher in females with right temporal lobe epilepsy (RTLE) (P < 0.05). Forced head deviation was significantly higher in males with left temporal lobe epilepsy (LTLE) (P < 0.03) and in females with RTLE (P < 0.0001). Unforced head deviation was significantly higher in males with RTLE (P < 0.002). Ipsilateral eye deviation was significantly higher in RTLE, with no differences between males and females. Postictal coughing was significantly higher in RTLE, again with no differences between males and females (P < 0.03). With regard to automatisms, posturing and nose wiping, there was no difference between right and left temporal lobe seizures or between genders. CONCLUSIONS: To the best of our knowledge, this study is the first demonstrating differences in ictal behaviour between females and males, thus showing that gender is related to different functional and anatomical organisations of the human brain. PMID- 15276147 TI - Epilepsy in the United Kingdom: seizure frequency and severity, anti-epileptic drug utilization and impact on life in 1652 people with epilepsy. AB - OBJECTIVES: To describe the clinical characteristics of epilepsy in a representative sample of the UK population, including seizure frequency and severity; overall severity of epilepsy; patterns of anti-epileptic drug (AED) use; and the impact of epilepsy on patients' lives. Secondly, to determine if these characteristics differ according to age. METHOD: A large, geographically comprehensive survey of people with epilepsy by means of a postal questionnaire distributed by general practitioners to 3455 unselected patients receiving AEDs for epilepsy, regardless of age or type of epilepsy and including all regions of the UK. Data were collected on age and gender; age of onset of seizures; seizure frequency and severity; AED use and adverse effect levels; and impact on life of epilepsy. Sub-analyses were performed with stratification by epilepsy severity and age-group. RESULTS: There were 1652 completed replies. The mean age was 44.2 years; there were 47.2% males, 48.5% females (4.4% not recorded). The mean age at first seizure, 25.1 years, and the mean duration of epilepsy, 19.7 years, were comparable with previous studies. In the preceding one year, 51.7% of patients had no seizures; 7.9% one seizure, 17.2% 2-9 seizures and 23.2% 10 or more. Sixty four percent of patients had epilepsy classified as mild and 32% severe. There was a marked and significant decrement of seizure frequency with increasing age. The most commonly used AEDs were carbamazepine (37.4%), valproate (35.7%), phenytoin (29.4%), phenobarbitone or primidone (14.2%) and lamotrigine (10.3%). Monotherapy was used in 68% of patients. Patients taking multiple AEDs reported significantly higher levels of adverse effects and worse seizure control. The major impacts of epilepsy on life were work and school difficulties, driving prohibition, psychological and social life. The impacts listed varied with the epilepsy severity and age. CONCLUSIONS: Seizures remain uncontrolled in up to half of all people with epilepsy in the UK with significant impact on work, family and social life. Previously, there has been a deficiency of data on the characteristics of epilepsy in older people, although it is recognized that the condition is of increasing epidemiological importance in this age group. We have found clear differences in the clinical characteristics of epilepsy in older people, particularly that seizure frequency appears to decline with increasing age. PMID- 15276148 TI - Early antiepileptic drug reduction following anterior temporal lobectomy for medically intractable complex partial epilepsy. AB - PURPOSE: To determine the safety, in our practice, of allowing patient preference to influence the timing of antiepileptic drug (AED) reduction, once they became seizure-free after anterior temporal lobectomy (ATL). METHODS: Thirty patients underwent anterior temporal lobectomy for medically intractable complex partial epilepsy at Loma Linda University Medical Center between December 1st 1991 and November 30th 2001. Timing of AED reduction in seizure-free patients was based on patient request. A review of patient records noted seizure status, duration from surgery to AED reduction, AED side effects, seizure recurrence and whether control was regained. RESULTS: Twenty-four (80%) of the 30 patients became seizure-free on their preoperative AEDs after initial ATL; three additional patients after a second operation. AEDs were not reduced in the reoperated patients, the three patients who did not become seizure-free, and in two patients who asked to increase AEDs to control auras. Thus, AEDs were reduced in 22 of the 27 seizure-free patients. Patients were followed an average of 3.4 +/- 2.7 (mean +/- standard deviation) years. AED reduction was initiated 4.6 +/- 7.2 months (range 0-27 months) after surgery. Polytherapy use decreased from 54% preoperatively to 18% at last follow up. Seizures recurred in six patients (27% of 22); three became seizure-free after AED adjustments. CONCLUSIONS: In our practice, using an individualized approach to AED reduction following successful epilepsy surgery resulted in early reduction in AEDs. Our data suggest that early AED reduction can be performed safely and without undue risk of seizure recurrence. PMID- 15276149 TI - The sociodemographic findings, beliefs and behaviours of the patients admitted to Kocaeli University, Faculty of Medicine, Epilepsy Section. AB - Using a standard questionnaire, 121 consequent epileptic patients have been evaluated sociodemographically and according to their beliefs and behaviours about their illness. Monthly income per person was between 26 and 62.5 U.S. dollars (USD) for 3/4 of the patients. About half of the patients were concealing their illness. About one third of the patients' education was hindered because of their illness. When the patients who concealed their illness were compared with the ones who did not, it became clear that the education of the ones who did not conceal the illness was far more hindered. "Carrying an amulet" ratio was greater among the patients who had no social security when compared to the patients with social security. We concluded that prejudice against epileptic patients and superstitions in Turkish population are still valid. Physician should consider the expense of antiepileptics for both patients and the country when prescribing them. All parts of the society should be informed about epilepsy but this again depends on the country's budget. PMID- 15276150 TI - Orgasmic aura--a report of seven cases. AB - We report on seven patients who experienced an orgasmic aura at the start of their seizures. The patients (five women, two men) were aged 36-58. Three of seven patients described the exact nature of their auras only many years after their appearance, when the epilepsy diagnostic procedure became more intensive due to drug resistance. Moreover, one patient even refused any new therapeutical options due to the reportedly positive role of the orgasmic aura in her life. All of our patients had temporal lobe epilepsy. The clinical picture, EEG, MRI or SPECT findings suggested a right temporal epileptic focus in six patients, while in one patient the epileptogenic region was localised in the left temporal lobe. In the latter case, the left hemisphere was speech-dominant, while in the other cases no Wada tests were done. Our results confirm that orgasmic aura could be considered as an ictal lateralising sign to the right hemisphere, however, it has no 100% lateralising value. PMID- 15276151 TI - Oligodendrocyte maturation is inhibited by bone morphogenetic protein. AB - Mature oligodendrocytes myelinate axons in the CNS. The development of the myelin sheath is dependent on the proper maturation of oligodendrocytes from precursors cells, a spatially restricted process that is regulated by inductive and repressive cues. Several members of the bone morphogenetic protein family (BMP2 and 4) have been implicated as repressors of oligodendrocyte development in vitro by shifting oligodendrocyte precursors into the astrocyte lineage. We now report on a second role of BMPs in oligodendrocyte development, regulation of myelin protein expression in immature oligodendrocytes. Purified immature rodent oligodendrocytes treated with BMP4 maintained galactocerebroside (GalC) expression, whereas the expression of three key myelin proteins, proteolipid protein (PLP), myelin basic protein (MBP), and 2'-3'-cyclic nucleotide 3' phosphodiesterase (CNP), was severely decreased. Paradoxically, BMP-treated oligodendrocytes show increased process extension and complexity, normally a feature of oligodendrocyte maturation. We also investigated whether BMP4 could inhibit myelin protein expression in an E 12.5 mouse explant culture of cervical spinal cord and hindbrain that maintains the in vivo cellular relationships and architecture. Beads soaked in BMP protein implanted into these explants inhibited the expression of myelin proteins, proteolipid protein, and myelin-associated glycoprotein (MAG), in the local area surrounding the bead. Since these explants also contained precursors cells, expression of galactocerebroside and O4, an oligodendrocyte marker, were also decreased by BMP treatment but to a much lesser degree than the myelin markers. Together, these data indicate that BMPs have multiple roles in oligodendrocyte development. At earlier stages, they affect cell lineage decisions and at later stages, they inhibit cell specialization. PMID- 15276152 TI - A potassium channel-linked mechanism of glial cell swelling in the postischemic retina. AB - The cellular mechanisms underlying glial cell swelling, a central cause of edema formation in the brain and retina, are not yet known. Here, we show that glial cells in the postischemic rat retina, but not in control retina, swell upon hypotonic stress. Swelling of control cells could be evoked when their K(+) channels were blocked. After transient ischemia, glial cells strongly downregulated their K(+) conductance and their prominent Kir4.1 protein expression at blood vessels and the vitreous body. In contrast, the expression of the aquaporin-4 (AQP4) (water channel) protein was only slightly altered after ischemia. Activation of D(2) dopaminergic receptors prevents the hypotonic glial cell swelling. The present results elucidate the coupling of transmembraneous water fluxes to K(+) currents in glial cells and reveal the role of altered K(+) channel expression in the development of cytotoxic edema. We propose a mechanism of postischemic glial cell swelling where a downregulation of their K(+) conductance prevents the emission of intracellularly accumulated K(+) ions, resulting in osmotically driven water fluxes from the blood into the glial cells via aquaporins. Inhibition of these water fluxes may be beneficial to prevent ischemia-evoked glial cell swelling. PMID- 15276153 TI - Theoretical consideration of olfactory axon projection with an activity-dependent neural network model. AB - Olfactory sensory neurons (OSNs) that express a given odorant receptor (OR) target their axons onto a few specific glomeruli in the olfactory bulb. Although the odorant receptor plays an indispensable role in olfactory axon targeting, the mechanisms underlying this guidance are largely unknown. In particular, there is much controversy regarding the involvement of activity-dependent mechanism in the targeting process. In this study, we developed an activity-dependent self organization model of the glomerular layer in the olfactory bulb and simulated the targeting of olfactory axons onto the layer. Our model successfully constructed discrete glomeruli that received olfactory axons expressing a common odorant receptor through odorant-evoked neural activities. Furthermore, our model explained the perplexing experimental results that have been reported in olfactory axon targeting. For example, dispersion of olfactory axons in knockout mice for the odorant receptor gene was reasonably reproduced in the simulation. The segregated projection of the axons that express the same odorant receptor transcribed from the different alleles was also successfully simulated if the genetically modified allele was assumed to express a smaller amount of the receptor protein. The activity-dependent model even explained the inconsistent effects of disruption of the activity-evoking ion channel on axons expressing different odorant receptors, although some of these results were regarded as evidence for activity-independency of the olfactory targeting. Taken together, the activity-dependent targeting of olfactory axons seems to be a simple probable mechanism that can provide a unified explanation of glomerular formation. PMID- 15276154 TI - The neuronal glycine transporter 2 interacts with the PDZ domain protein syntenin 1. AB - The glycine transporter subtype 2 (GlyT2) is localized at glycinergic axon terminals where it mediates the re-uptake of glycine from the extracellular space. In this study, we used the yeast two-hybrid system to search for proteins that interact with the cytoplasmic carboxy terminal tail region of GlyT2. Screening of a rat brain cDNA library identified the PDZ domain protein syntenin 1 as an intracellular binding partner of GlyT2. In pull-down experiments, the interaction between GlyT2 and syntenin-1 was found to involve the C-terminal amino acid residues of GlyT2 and the PDZ2 domain of syntenin-1. Syntenin-1 is widely expressed in brain and co-localizes with GlyT2 in brainstem sections. Furthermore, syntenin-1 binds syntaxin 1A, which is known to regulate the plasma membrane insertion of GlyT2. Thus, syntenin-1 may be an in vivo binding partner of GlyT2 that regulates its trafficking and/or presynaptic localization in glycinergic neurons. PMID- 15276155 TI - Alpha-protocadherins are presynaptic and axonal in nicotinic pathways. AB - The protocadherin families pcdh-alpha, beta, and gamma have been proposed to mediate synaptic specificity via homophilic interactions. Here we report isolation of two pcdh-alpha family members from chick. We find pcdh-alpha mRNA in multiple regions of chick CNS including cerebellum, tectum, olfactory bulb, and forebrain, and in the autonomic nervous system. Immunoblots identify major components of 120 and 140 kDa both in brain and ciliary ganglion extracts. Immunohistochemistry reveals pcdh-alphas in axons and perisynaptically in preganglionic terminals, adjacent to transmitter release sites. Pcdh-alphas appear to be absent from postsynaptic sites: They are nonoverlapping with postsynaptic receptor clusters in the ganglion and are rapidly lost after ganglionic denervation. Similar pcdh-alpha patterns are found in motor axons and at neuromuscular junctions of birds and mammals, and persist into adulthood. The results indicate that pcdh-alphas are widely expressed in nicotinic cholinergic pathways and may engage in heterophilic interactions at synapses and on axons. PMID- 15276156 TI - Invulnerability of retinal ganglion cells to NMDA excitotoxicity. AB - NMDA excitotoxicity has been proposed to mediate the death of retinal ganglion cells (RGCs) in glaucoma and ischemia. Here, we reexamine the effects of glutamate and NMDA on rat RGCs in vitro and in situ. We show that highly purified RGCs express NR1 and NR2 receptor subunits by Western blotting and immunostaining, and functional NMDA receptor channels by whole-cell patch-clamp recording. Nevertheless, high concentrations of glutamate or NMDA failed to induce the death of purified RGCs, even after prolonged exposure for 24 h. RGCs co-cultured together with ephrins, astrocytes, or mixed retinal cells were similarly invulnerable to glutamate and NMDA, though their NMDA currents were 4 fold larger. In contrast, even a short exposure to glutamate or NMDA induced the rapid and profound excitotoxic death of most hippocampal neurons in culture. To determine whether RGCs in an intact retina are vulnerable to excitotoxicity, we retrogradely labeled RGCs in vivo using fluorogold and exposed acutely isolated intact retinas to high concentrations of glutamate or NMDA. This produced a substantial and rapid loss of amacrine cells; however, RGCs were not affected. Nonetheless, RGCs expressed NMDA currents in situ that were larger than those reported for amacrine cells. Interestingly, the NMDA receptors expressed by RGCs were extrasynaptically localized both in vitro and in situ. These results indicate that RGCs in vitro and in situ are relatively invulnerable to glutamate and NMDA excitotoxicity compared to amacrine cells, and indicate that important, as yet unidentified, determinants downstream of NMDA receptors control vulnerability to excitotoxicity. PMID- 15276157 TI - Synergistic effects of neuregulin and agrin on muscle acetylcholine receptor expression. AB - The proper function of neuromuscular junctions requires an extremely high density of acetylcholine receptors (AChRs) that may be achieved from neuron-derived factors including agrin and neuregulin. Here, we show that neuregulin-1 and agrin co-localize at neuromuscular junctions in vivo and form complexes when co transfected into COS-7 cells. When these COS-7 cells are cultured with myotubes, synergistic effects are observed for AChR clustering, membrane insertion of new AChRs, and induction of AChR mRNA. Even a muscle form of agrin that lacks intrinsic clustering activities by itself, significantly enhances neuregulin induced clustering and insertion of AChRs. While the heparin-binding (A) domain of agrin is required for agrin localization in the extracellular matrix adjacent to AChR clusters, the heparan sulfate-containing domain of agrin is needed for the synergistic effects and co-localization with neuregulin-1. These results suggest that matrix interactions between exogenously supplied agrin and neuregulin-1 on the muscle surface provide a localized source of signaling factors needed to produce high densities of AChRs at neuromuscular junctions. PMID- 15276160 TI - Modification of Cytochrome c by 4-hydroxy- 2-nonenal: evidence for histidine, lysine, and arginine-aldehyde adducts. AB - 4-Hydroxy-2-nonenal (4HNE), a major secondary product of lipid peroxidation, has been associated with a number of disease states involving oxidative stress. Despite the recognized importance of post-translational modification of proteins by products such as 4HNE, little is known of the modification of cytochrome c by this reagent and its analysis by mass spectrometry. The purpose of this study was to investigate the chemical interaction of 4HNE and cytochrome c, a protein essential to cellular respiration, under in vitro conditions. Isoelectric focusing of native and 4HNE-modified cytochrome c using immobilized pH gradient (IpG) strips showed a decrease in the pI of the 4HNE-modified protein suggesting modification of charged amino acids. Reaction of 4HNE with cytochrome c resulted in increases in molecular weight consistent with the addition of four 4HNE residues as determined by matrix-assisted laser desorption time-of-flight mass spectrometry (MALDI-TOF MS). Samples of both native and 4HNE-modified cytochrome c were enzymatically digested and subjected to peptide mass fingerprinting using MALDI-TOF MS. Analysis of these samples using LC-electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) provided sequence information that was used to determine specific residues to which the aldehyde adducted. Taken together, the data indicated that H33, K87, and R38 were modified by 4HNE. Mapping these results onto the X-ray crystal structure of native cytochrome c suggest that 4HNE adduction to cytochrome c could have significant effects on tertiary structure, electron transport, and ultimately, mitochondrial dysfunction. PMID- 15276161 TI - A SIFT ion-molecule study of some reactions in Titan's atmosphere. reactions of N(+), N(2)(+), and HCN(+) with CH(4), C(2)H(2), and C(2)H(4). AB - The results of a study of the ion-molecule reactions of N(+), N(2)(+), and HCN(+) with methane, acetylene, and ethylene are reported. These studies were performed using the FA-SIFT at the University of Canterbury. The reactions studied here are important to understanding the ion chemistry in Titan's atmosphere. N(+) and N(2)(+) are the primary ions formed by photo-ionization and electron impact in Titan's ionosphere and drive Titan's ion chemistry. It is therefore very important to know how these ions react with the principal trace neutral species in Titan's atmosphere: Methane, acetylene, and ethylene. While these reactions have been studied before the product channels have been difficult to define as several potential isobaric products make a definitive answer difficult. Mass overlap causes difficulties in making unambiguous species assignments in these systems. Two discriminators have been used in this study to resolve the mass overlap problem. They are deuterium labeling and also the differences in reactivities of each isobar with various neutral reactants. Several differences have been found from the products in previous work. The HCN(+) ion is important in both Titan's atmosphere and in the laboratory. PMID- 15276162 TI - MALDI-MS analysis of peptides modified with photolabile arylazido groups. AB - The ability of MALDI-MS to analyze photolabile arylazido peptide derivatives was investigated. Peptides containing UV-labile p-azidobenzoyl groups were subjected to MALDI-MS analysis in a variety of matrices. As standard MALDI-MS employs a UV laser (337 nm), we investigated conditions that would allow detection of the intact molecule ions for these light-sensitive peptides. When using alpha-cyano-4 hydroxycinnamic acid (ACHC) or 2,5 dihydroxybenzoic acid (DHB) as the matrix, photoinduced degradation products were prevalent. In contrast, when employing the matrix sinapinic acid, the intact molecule ion corresponding with the azido peptide was the predominant signal. The protection of photolabile azido derivatives correlates with the UV absorbance properties of the matrix employed, i.e., sinapinic acid, which exhibits a strong absorbance near 337 nm, most efficiently protects the azido derivative from photodegradation. PMID- 15276163 TI - Phenyl- and cyclopentylimino derivatization for double bond location in unsaturated C(37)-C(40) alkenones by GC-MS. AB - A method for the identification of double bond locations in polyunsaturated long chain alkenones adapted to nanogram amounts as currently analyzed by gas chromatography coupled to mass spectrometry (GC-MS) has been developed. The method is based on interpretation of the electron impact mass spectra of the imino derivatives of the carbonyl groups using either cyclopentyl or phenyl substitutents. Other complementary derivatization methods such as elaidization and hydrogenation have also been used for structural characterization of these compounds. This application has led to the identification of a novel homologous series of di-, tri-, and tetraunsaturated ketones with carbon number chain lengths between 37 and 40 in coastal hypersaline sediments. The novel series identified shows a distribution in which the double bond position between different homologs is established by reference to the distance from the carbonyl group whereas the previously known alkenones were constituted by unsaturated homologs with double bonds located at defined distances of the terminal methyl. This difference points out to a dissimilar, but still unknown, biogenic precursor of these novel alkenones. PMID- 15276164 TI - A new matching algorithm for high resolution mass spectra. AB - We present a new matching algorithm designed to compare high-resolution spectra. Whereas existing methods are bound to compare fixed intervals of ion masses, the accurate mass spectrum (AMS) distance method presented here is independent of any alignment. Based on the Jeffreys-Matusitas (JM) distance, a difference between observed peaks across pairs of spectra can be calculated, and used to find a unique correspondence between the peaks. The method takes into account that there may be differences in resolution of the spectra. The algorithm is used for indexing in a database containing 80 accurate mass spectra from an analysis of extracts of 80 isolates representing the nine closely related species in the Penicillium series Viridicata. Using this algorithm we can obtain a retrieval performance of approximately 97-98% that is comparable with the best of the existing methods (e.g., the dot-product distance). Furthermore, the presented method is independent of any variable alignment procedures or binning. PMID- 15276165 TI - Characterization of noncovalent complexes of antimalarial agents of the artemisinin-type and FE(III)-heme by electrospray mass spectrometry and collisional activation tandem mass spectrometry. AB - In this study, we demonstrate, using electrospray ionization mass spectrometry (ESI-MS) and collision-induced dissociation tandem mass spectrometry (ESI MS/CID/MS), that stable noncovalent complexes can be formed between Fe(III)-heme and antimalarial agents, i.e., quinine, artemisinin, and the artemisinin derivatives, dihydroartemisinin, alpha- and beta-artemether, and beta-arteether. Differences in the binding behavior of the examined drugs with Fe(III)-heme and the stability of the drug-heme complexes are demonstrated. The results show that all tested antimalarial agents form a drug-heme complex with a 1:1 stoichiometry but that quinine also results in a second complex with the heme dimer. ESI-MS performed on mixtures of pairs of various antimalarial agents with heme indicate that quinine binds preferentially to Fe(III)-heme, while ESI-MS/CID/MS shows that the quinine-heme complex is nearly two times more stable than the complexes formed between heme and artemisinin or its derivatives. Moreover, it is found that dihydroartemisinin, the active metabolite of the artemisinin-type drugs in vivo, results in a Na(+)-containing heme-drug complex, which is as stable as the heme-quinine complex. The efficiency of drug-heme binding of artemisinin derivatives is generally lower and the decomposition under CID higher compared with quinine, but these parameters are within the same order of magnitude. These results suggest that the efficiency of antimalarial agents of the artemisinin type to form noncovalent complexes with Fe(III)-heme is comparable with that of the traditional antimalarial agent, quinine. Our study illustrates that electrospray ionization mass spectrometry and collision-induced dissociation tandem mass spectrometry are suitable tools to probe noncovalent interactions between heme and antimalarial agents. The results obtained provide insights into the underlying molecular modes of action of the traditional antimalarial agent quinine and of the antimalarials of the artemisinin-type which are currently used to treat severe or multidrug-resistant malaria. PMID- 15276166 TI - A preconcentrator coupled to a GC/FTMS: advantages of self-chemical ionization, mass measurement accuracy, and high mass resolving power for GC applications. AB - Coupling of a cryogenic preconcentrator (PC) to a gas chromatograph/Fourier transform ion cyclotron resonance mass spectrometer (GC/FT-ICR MS) is reported. To demonstrate the analytical capabilities of the PC/GC/FT-ICR MS, headspace samples containing volatile organic compounds (VOCs) emitted from detached pine tree twigs were analyzed. Sub-ppm mass measurement accuracy (MMA) for highly resolved (m/Deltam(50%) > 150 k) terpene ions was achieved. Direct PC/GC/FT-ICR MS analyses revealed that detached twigs from pine trees emit acetone, camphor, and four detectable hydrocarbon isomers with C(10)H(16) empirical formula. The unknown analytes were identified based on accurate mass measurement and their mass spectral appearances. Authentic samples were used to confirm initially unknown identifications. Self-chemical-ionization (SCI) reactions furnished an additional dimension for rapid isomer differentiation of GC eluents in real time. PMID- 15276167 TI - Automated orthogonal control system for electrospray ionization. AB - Low-flow electrospray ionization is typically a purely electrostatic method, used without supporting sheath-gas nebulization. Complex spray morphology results from a large number of possible spray emission modes. Spray morphology may assume the optimal Taylor cone-jet spray mode under equilibrium conditions. When coupling to nanobore gradient elution chromatography, however, stability of the Taylor cone jet spray mode is compromised by the gradient of mobile phase physiochemical properties. The common spray modes for aqueous/organic mobile phases were characterized using orthogonal (strobed illumination) transmitted light and (continuous illumination) scattered light imaging. Correlation of image sets from these complementary illumination methods provides the basis for spray mode identification using qualitative and quantitative image analysis. An automated feedback-controlled electrospray source was developed on a computer capable of controlling electrospray potential using an image-processing based algorithm for spray mode identification. The implementation of the feedback loop results in a system that is both self-starting and self-tuning for a specific spray mode or modes. Thus, changes in mobile phase composition and/or flow rate are compensated in real-time and the source is maintained in the cone-jet or pulsed cone-jet spray modes. PMID- 15276168 TI - Quantitation of lanosterol and its major metabolite FF-MAS in an inhibition assay of CYP51 by azoles with atmospheric pressure photoionization based LC-MS/MS. AB - Azoles affect the steroid balance in all biological systems and may therefore be called endocrine disrupters. Lanosterol 14alpha-demethylase (CYP51) is an enzyme inhibited by azoles. Only few data have been reported showing their inhibitory potency since an assay in an in vitro system is not available so far. In the present work an inhibition assay using human recombinant CYP51, coexpressed with human P450 oxido-reductase by the baculovirus/insect cell expression system, and LC-MS/MS as analytical method is described. Atmospheric pressure photoionization (APPI) and atmospheric pressure chemical ionization (APCI) sources were used with a triple quadrupole mass spectrometer to compare quantitation of lanosterol (substrate) and 4,4-dimethyl-5alpha-cholesta-8,14,24-triene-3beta-ol (FF-MAS) (product of CYP51) with d(6)-2,2,3,4,4,6-cholesterol (d(6)-cholesterol) as internal standard. Optimization of analytical parameters resulted in a LC-APPI MS/MS method with a LOQ of 10 pg on column for FF-MAS. The sensitivity of the method (LOD 0.5 ng/ml) makes it possible to analyze supernatants of inhibition experiments after precipitation of proteins by isopropanol without any sample enrichment. The coefficient of variation of the analytical method was <20% (n = 5) for FF-MAS, lanosterol and d(6)-cholesterol. The external calibration curve was linear from 1 to 10,000 ng/ml with R(2) >/= 0.999 and an accuracy of 94-115%. Compared with APCI, APPI provides a ten- to 500-fold increase in sensitivity for the analytes in this study. IC(50) values of epoxiconazole and miconazole-two widely used azole fungicides used in agriculture and in human medicine, respectively-were 1.95 microM and 0.057 microM. PMID- 15276169 TI - Rapid identification of stress-related fingerprint from whole bacterial cells of Bifidobacterium lactis using matrix assisted laser desorption/ionization mass spectrometry. AB - Whole cells of Bifidobacterium lactis were analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS). Characteristic and reproducible mass spectra were obtained in the mass range from 6 to 19 kDa. After several days of bacterial cell storage at 4 degrees C (D0, D2, and D6), only minor signal differences were observed. Under identical and reproducible conditions, fourteen relevant diagnostic ions were identified. Moreover, control- and stress-related fingerprints were rapidly obtained using MALDI-TOFMS by comparison of protein patterns obtained from non-stressed (control) versus stressed cells (addition of bile salts during growth). After quantitative validation of the MALDI-MS data by a statistical approach, two and eight signals were assigned as control- and stress-specific ions, respectively. This work provides the evidence that MALDI-TOFMS can be used for the identification of stress-related fingerprint of B. lactis bacterial cells and could have a high potential for the assessment of the physiological status of the cells. PMID- 15276170 TI - Product analysis of caffeic acid oxidation by on-line electrochemistry/electrospray ionization mass spectrometry. AB - On-line electrochemistry/electrospray ionization mass spectrometry (EC/ESI-MS) was developed using a microflow electrolytic cell. This technique was applied to electrochemical oxidation of caffeic acid (CAF) which is known to be a highly antioxidative agent. Effects of electrolytic potentials on ion intensities of product ions and on electrolytic currents were examined at different pHs. Dimer products were detected at electrolytic potentials of E = 0.7 V (vs. Ag/AgCl) and trimer products at 1.0 V at pH 9. Dimer products were distinguished from hydrogen bonded complexes by MS/MS experiments. Hydrogen/deuterium exchange experiments determined the number of hydroxyl and carboxyl groups in the Dimers formed by electrolysis. The mechanism of oxidative polymerization of CAF is discussed with speculation as to the structure of the dimer product. PMID- 15276171 TI - Probing three-dimensional structure of bovine serum albumin by chemical cross linking and mass spectrometry. AB - Serum albumin is the principal transporter of fatty acids that are otherwise insoluble in circulating plasma. While the crystal structure of human serum albumin (HSA) as well as its binding with fatty acids has been characterized, the three dimensional structure of bovine serum albumin (BSA) has not been determined although both albumins share 76% sequence homology. In this study we used mass spectrometry coupled with chemical cross-linking, to probe the tertiary structure of BSA. BSA was modified with lysine specific cross-linkers, bis(sulfosuccinimidyl) suberate (BS(3)), disuccinimidyl suberate (DSS) or disuccinimidyl glutarate (DSG), digested with trypsin and analyzed by tandem mass spectrometry. With O-18 labeling during the digestion, through-space cross-linked peptides were readily identified in mass spectra by a characteristic 8 Da shift. From the cross-linked peptides identified in this study, we found that 12 pairs of lysine residues were separated within 20 A, while 5 pairs were spaced between 20 and 24 A. The spatial distance constraints generated from five K-K pairs in BSA were consistent with the corresponding distance obtained from the crystal structure of HSA, although only six equivalent K-K pairs could be compared. According to our data, the distance between K235 of IIA and K374 of IIB domain in BSA was farther by 7-11 A than that expected from the crystal structure of HSA, suggesting structural differences between BSA and HSA in this region. The distance constraints obtained for lysine residues using various cross-linkers should be valuable in assisting the determination of the 3-D structure of BSA. PMID- 15276172 TI - Tandem time-of-flight mass spectrometer for photodissociation of biopolymer ions generated by matrix-assisted laser desorption ionization (MALDI-TOF-PD-TOF) using a linear-plus-quadratic potential reflectron. AB - A tandem time-of-flight mass spectrometer for the study of photodissociation of biopolymer ions generated by matrix-assisted laser desorption ionization was designed and constructed. A reflectron with linear and quadratic (LPQ) potential components was used. Characteristics of the LPQ reflectron and its utility as the second stage analyzer of the tandem mass spectrometer were investigated. Performance of the instrument was tested by observing photodissociation of [M + H](+) from angiotensin II, a prototype polypeptide. Quality of the photodissociation tandem mass spectrum was almost comparable to that of the post source decay spectrum. Monoisotopic selection of the parent ion was possible, which was achieved through the ion beam-laser beam synchronization. General theoretical considerations needed for a successful photodissociation of large biopolymer ions are also presented. PMID- 15276173 TI - Clinical and research validity of hadrontherapy with ion beams. AB - Clinical results obtained with hadrontherapy have been extremely positive for various tumours, with percentages of local control and survival higher than those ascribed to conventional radiotherapy. Most clinical data obtained with charged particles are related to protontherapy but the implementation of carbon ion therapy has demonstrated to be of great interest in the last decade. These results, accompanied by the new performances in accelerator technology and calculation systems of the delivered doses, have determined over the past years an increased interest for the development of hadrontherapy, with the construction of new centres provided with equipment entirely dedicated to clinical activity (LLUMC, Loma Linda and NPTC, Boston in USA, HIMAC-NIRS, Chiba, PROBEAT, Tsukuba, and Hyogo Beam Medical Centres in Japan). A revision of the clinical indications specifically focused on ion therapy is presented as well as the results obtained in the different centres. With hadrontherapy, it is finally possible to increase the spectrum of treatments allowing preserving the organ and its functionality, with positive impacts from a social and economic point of view. PMID- 15276174 TI - Acute respiratory failure in the cancer patient: the role of non-invasive mechanical ventilation. AB - The most common cause of ICU admission in patients affected by a hematologic or solid cancer is acute respiratory failure, often associated with a respiratory infection. The prognosis of these critically ill patients is disappointingly low especially if they require endotracheal intubation. In the last 10 years, non invasive mechanical ventilation (NIV), delivered through a face or nose mask, has been increasingly used as an alternative to invasive ventilation. There is good evidence that, compared to the standard medical therapy alone or with invasive mechanical ventilation, NIV may improve survival and reduce the rate of infectious complications in patients affected by hematologic cancers. Patients with a solid tumor and "reversible" acute respiratory failure are also likely to benefit from NIV, while the use of NIV in palliative care of terminally ill patients still needs to be elucidated. The success of NIV is strictly dependent on its "early" use and on the experience of the staff involved. PMID- 15276175 TI - Quality assurance of surgery in gastric and rectal cancer. AB - Multimodality and quality controlled treatment result in improved treatment outcome in patients with solid tumours. Quality assurance focuses on identifying and reducing variations in treatment strategy. Treatment outcome is subsequently improved through the introduction of programs that reduce treatment variations to an acceptable level and implement standardised treatment. In chemotherapy and radiotherapy, such programmes have been introduced successfully. In surgery however, there has been little attention for quality assurance so far. Surgery is the mainstay in the treatment of patients with gastric and rectal cancer. In gastric cancer, the extent of surgery is continuously being debated. In Japan, extended lymph node dissection is favoured whereas in the West this type of surgery is not routinely performed with two large European trials concluding that there is no survival benefit from regional lymph node clearance. Post-operative chemoradiation is part of the standard treatment in the United States, although its role in combination with adequate surgery has not been established yet. These global differences in treatment policy clearly relate to the extent and quality of surgical treatment. As for gastric cancer, surgical treatment of rectal cancer patients determines patient's prognosis to a large extent. With the introduction of total mesorectal excision, local control and survival have improved substantially. Most rectal cancer patients receive adjuvant treatment, either pre or post-operatively. The efficacy of many adjuvant treatment regimens has been investigated in combination with conventional suboptimal surgery. Traditional indications of adjuvant treatment might have to be re-examined, considering the substantial changes in surgical practise. Quality assurance programs enable the introduction of standardised and quality controlled surgery. Promising adjuvant regimens should be investigated in combination with optimal surgery. PMID- 15276176 TI - Rectal cancer. AB - Rectal cancer is an important tumour from an epidemiological point of view and represents the benchmark for an optimal use of integrated treatments (surgery, radiotherapy and chemotherapy) in the oncological practice. Performing radio chemotherapy (best if preoperatively), medical and radiation oncologists are now able to increase survival, to decrease the occurrence of pelvic recurrence and to ameliorate the quality of life of patients. Updated recommendations for the management of these patients are here reported. PMID- 15276177 TI - Colon cancer. AB - Colon cancer is one of the leading tumours in the world and is considered among the big killers, together with lung, prostate and breast cancer. In the recent years very important advances occurred in the field of treatment of this frequent disease: adjuvant chemotherapy was demonstrated to be effective, chiefly in stage III patients, and surgery was optimized in order to achieve the best results with a low morbidity. Several new target-oriented drugs are under evaluation and some of them (cetuximab and bevacizumab) have already exhibited a good activity/efficacy, mainly in combination with chemotherapy. The development of updated recommendations for the best management of these patients is crucial in order to obtain the best results, not only in clinical research but also in everyday practice. This report summarizes the most important achievements in this field and provides the readers useful suggestions for their professional practice. PMID- 15276178 TI - Crystal structure of p19--a universal suppressor of RNA silencing. AB - RNA silencing in plants has an antiviral role and, consequently, plant viruses encode counter-defensive suppressor proteins that block this process. The recently reported crystal structure of two Tombusvirus suppressor proteins reveals a novel RNA-binding structure and illustrates precisely how the silencing mechanism is blocked. These suppressor protein structures, combined with molecular analyses of their effects in animal and plant cells, are informative about RNA silencing mechanisms. They also suggest various ways that Tombusvirus suppressors can be used to investigate RNA silencing in plants and animals. PMID- 15276179 TI - Stop the killer: how to inhibit the anthrax lethal factor metalloprotease. AB - Inhalation of anthrax spores rapidly develops into a deadly bacteraemia and toxaemia. Anthrax toxins include the lethal factor (LF), a mitogen-activated protein kinase (MAPK)-kinase-specific metalloprotease, which acts in the cell cytosol and plays a major part in anthrax pathogenesis. Recently, screening methods have led to the discovery of LF inhibitors that are membrane permeable. This will pave the way for design of novel anthrax therapeutics that are capable of inhibiting the metalloprotease activity of LF in vivo. PMID- 15276180 TI - MT1-MMP: an enzyme with multidimensional regulation. AB - The activity of membrane-type 1 matrix metalloproteinase (MT1-MMP) is a double edged sword--it is crucial for both physiological processes and disease progression. MT1-MMP modifies various cellular functions and it is, sthus, regulated precisely as a proteinase and as a membrane protein. Recent studies have further revealed that the function of MT1-MMP is modified and regulated by O glycosylation, interaction with CD44, internalization and recycling. Such multidimensional mechanisms enable MT1-MMP to be regulated spatially and temporally, and are essential for its proper functioning on the cell surface. PMID- 15276181 TI - Vitamin K epoxide reductase: homology, active site and catalytic mechanism. AB - Vitamin K epoxide reductase (VKOR) recycles reduced vitamin K, which is used subsequently as a co-factor in the gamma-carboxylation of glutamic acid residues in blood coagulation enzymes. VKORC1, a subunit of the VKOR complex, has recently been shown to possess this activity. Here, we show that VKORC1 is a member of a large family of predicted enzymes that are present in vertebrates, Drosophila, plants, bacteria and archaea. Four cysteine residues and one residue, which is either serine or threonine, are identified as likely active-site residues. In some plant and bacterial homologues the VKORC1 homologous domain is fused with domains of the thioredoxin family of oxidoreductases. These might reduce disulfide bonds of VKORC1-like enzymes as a prerequisite for their catalytic activities. PMID- 15276182 TI - The history of deciphering the genetic code: setting the record straight. PMID- 15276183 TI - MD-2: the Toll 'gatekeeper' in endotoxin signalling. AB - Lipopolysaccharide (LPS) from the outer cell wall of Gram-negative bacteria is a potent stimulator of the mammalian innate immune system. The Toll-like receptor 4 (TLR4) pathway triggers the inflammatory responses induced by LPS in a process that requires the interaction of LPS-bound myeloid differentiation-2 (MD-2) with TLR4. Here we propose two possible mechanisms for LPS recognition and signalling that take into account both the structural information available for TLR4 and MD 2, and the determinants of endotoxicity, namely, the acylation and phosphorylation patterns of LPS. In our first model, LPS induces the association of two TLR4-MD-2 heterodimers by binding to two different molecules of MD-2 through the acyl chains of lipid A. In our second model, the binding of LPS to a single TLR4-MD-2 complex facilitates the recruitment of a second TLR4-MD-2 heterodimer. These models contrast with the activation of Drosophila Toll, where the receptor is crosslinked by a dimeric protein ligand. PMID- 15276184 TI - The kinetic mechanism of kinesin. AB - The chemical kinetic mechanism of kinesin (K) is considered by using a consensus scheme incorporating biochemically defined open, closed and trapped states. In the absence of microtubules, the dominant species is a trapped K*ADP state, which is defined by its ultra-slow release of ADP (off rate, k(off) approximately 0.002 s(-1)) and weak microtubule binding (dissociation constant, K(d) approximately 10 20 microM). Once bound, this trapped state equilibrates with a strongly binding open state that rapidly releases ADP (k(off) approximately 300 s(-1)). After ADP release, Mg*ATP binds (on rate, k(on) approximately 2 microM(-1)s(-1)) driving formation of a closed state that is defined by hydrolysis competence and by strong binding to microtubules. Hydrolysis (k(hyd) approximately 100-300 s(-1)) and phosphate release (k(off)>100 s(-1)) both occur in this microtubule-bound closed state. Phosphate release acts as a gate that controls reversion to the trapped K*ADP state, which detaches from the microtubule, completing the cycle. PMID- 15276185 TI - Emerging functions of BRCA2 in DNA recombination. AB - Mutations in the breast cancer susceptibility protein BRCA2 cause inherited susceptibility to breast, ovarian and other cancers. There is now compelling experimental evidence that a major biological function of BRCA2 is the maintenance of chromosome structure stability in dividing cells by regulation of steps in recombination between homologous DNA strands. Recent experimental findings shape the current models for BRCA2 function, and structural and biochemical advances shed new light on the interactions between BRCA2, the RAD51 recombinase and single-stranded DNA during DNA recombination. PMID- 15276186 TI - Mechanism of the nuclear receptor molecular switch. AB - Nuclear receptors are central to the regulation of development, endocrine signalling and metabolism. The transcriptional activity of many receptors is controlled through the binding of small, fat-soluble molecules to the ligand binding domain. In most cases, ligand binding turns the receptors into potent activators of transcription. This switch involves the exchange of co-regulator proteins that mediate transcriptional regulation. Structural and biochemical studies have together revealed the mechanism of action of this ligand-induced molecular switch, in which changes in the dynamic behaviour of the receptor play a key role. This remarkable dynamic mechanism has facilitated the evolution of a family of nuclear receptors with highly diverse ligand recognition and signalling properties. PMID- 15276187 TI - Jumping to rafts: gatekeeper role of bilayer elasticity. AB - Two of the physiologically important processes that take place in biological membranes are the partitioning of water-soluble proteins into the membrane and the sequestering of specific transmembrane proteins into membrane microdomains or 'rafts'. Although these two processes often involve different classes of protein, recent biophysical studies indicate that they both strongly depend on the structural and elastic properties of the membrane bilayer. That is, both the partitioning of peptides into membranes and the distribution of transmembrane peptides in the plane of the membrane are modulated by physical properties of the lipid bilayer that are controlled by cholesterol content and the composition of the phospholipid hydrocarbon chain. PMID- 15276188 TI - Psychostimulants: new concepts for palliative care from the modafinil experience? PMID- 15276189 TI - Subanesthetic ketamine: an essential adjuvant for intractable cancer pain. PMID- 15276190 TI - Olanzapine-induced delirium in a terminally ill cancer patient. PMID- 15276191 TI - Evaluating palliative care: bereaved family members' evaluations of patients' pain, anxiety and depression. AB - Palliative care surveys often rely on bereaved family members to act as proxies to provide information on patient care at the end of life, after the patient's death. However, when comparing bereaved family members' assessments with those of the patients, agreement is found to be better for symptoms that are more concrete and observable than subjective aspects such as psychological symptoms and pain. To date, little is known about how proxies actually evaluate these types of symptoms. The present study used retrospective verbal protocol analysis to elucidate the thought processes of 30 bereaved relatives during their evaluations of patients' pain, anxiety and depression. The qualitative analysis raised awareness of the difficulties experienced by proxies when discerning the presence of symptoms. It also provided insights into the cues and strategies used when making decisions, contributing to a fuller understanding of how proxies distinguish symptoms. Recommendations are made to improve the design of retrospective palliative care surveys. PMID- 15276192 TI - Prediction of survival in terminal cancer patients in Taiwan: constructing a prognostic scale. AB - We prospectively identified prognostic factors and developed a prognostic scale in 356 Taiwanese terminal cancer patients (training set). Demographic data, severity of symptoms/signs, and survival were statistically analyzed to create the scale, which was tested in another 184 patients (testing set). In the training set, liver and lung metastases, functional performance status, weight loss, edema, cognitive impairment, tiredness, and ascites were independently associated with shorter survival (multivariate analysis). The scale ranged from 0.0 (no altered variables) to 8.5 (maximal alteration for all variables). When scores were < 3.5, 2-week survival was predicted with 0.72 and 0.61 accuracy for the training and testing sets, respectively. With scores < 6.0, 1-week survival was predicted with 0.72 and 0.66 accuracy, respectively. This scale, which includes lung and liver metastases and severity of symptoms/signs, may help in identifying the stage of dying and its corresponding symptoms/signs and also in improving survival prediction in terminal cancer patients. PMID- 15276193 TI - Dyspnea and its correlates in taiwanese patients with terminal cancer. AB - This study prospectively assessed dyspnea and related bio-psycho-social-spiritual factors--including severity, cause, psychological distress, and fear of death- that were possibly related to dyspnea in 125 terminal cancer patients at admission and two days before their death. At admission, 74 patients had dyspnea, which improved but later worsened. Causes included cachexia, anemia, pleural effusion, and lymphangitis. Quality of life, anxiety, depression, and fear of death improved after admission; anxiety was correlated with dyspnea before death (r = 0.211, P < 0.05, univariate analysis). Lung infection (odds ratio = 2.29, 95% confidence interval = 0.68-3.90; multiple regression), airway obstruction (2.27, 1.41-3.13), acidemia (1.82, 0.72-2.98), and pericardial effusion (1.38, 0.44-2.32) were independent correlates of dyspnea severity at admission (42.8% of explained variance). Before death, airway obstruction, esophageal cancer, pericardial effusion, lung infection, and mediastinal mass were independent correlates of severity (42.7% of explained variance). Comprehensive care, including improved psychospiritual status, can help in controlling dyspnea and enhancing patients' quality of life. PMID- 15276194 TI - Symptom experience in Korean adults with lung cancer. AB - This study aimed to examine how symptoms vary in relation to demographic characteristics (age and sex), stage of disease, histology of lung cancer, and treatment type in Korean adults with lung cancer. Symptoms were measured with the Symptom Distress Scale. A total 106 patients with a mean age of 60.9 (SD = 10.38) years participated. The results indicated that 1) overall symptom distress was more severe (mean 32.74, SD 10.75) compared to the studies reported in Western countries, and 2) among the variables, only the stage of lung cancer showed a significant relationship with total symptom distress (P < 0.05). In analyses of the individual symptoms, bowel-related symptoms showed significant relationships with sex, age, and type of treatment. The results highlight the importance of symptom management as well as the need to tailor clinical interventions according to related factors in order to maximize effective symptom management in Korean patients with lung cancer. PMID- 15276195 TI - Comparative efficacy and safety of skeletal muscle relaxants for spasticity and musculoskeletal conditions: a systematic review. AB - Skeletal muscle relaxants are a heterogeneous group of medications used to treat two different types of underlying conditions: spasticity from upper motor neuron syndromes and muscular pain or spasms from peripheral musculoskeletal conditions. Although widely used for these indications, there appear to be gaps in our understanding of the comparative efficacy and safety of different skeletal muscle relaxants. This systematic review summarizes and assesses the evidence for the comparative efficacy and safety of skeletal muscle relaxants for spasticity and musculoskeletal conditions. Randomized trials (for comparative efficacy and adverse events) and observational studies (for adverse events only) that included oral medications classified as skeletal muscle relaxants by the FDA were sought using electronic databases, reference lists, and pharmaceutical company submissions. Searches were performed through January 2003. The validity of each included study was assessed using a data abstraction form and predefined criteria. An overall grade was allocated for the body of evidence for each key question. A total of 101 randomized trials were included in this review. No randomized trial was rated good quality, and there was little evidence of rigorous adverse event assessment in included trials or observational studies. There is fair evidence that baclofen, tizanidine, and dantrolene are effective compared to placebo in patients with spasticity (primarily multiple sclerosis). There is fair evidence that baclofen and tizanidine are roughly equivalent for efficacy in patients with spasticity, but insufficient evidence to determine the efficacy of dantrolene compared to baclofen or tizanidine. There is fair evidence that although the overall rate of adverse effects between tizanidine and baclofen is similar, tizanidine is associated with more dry mouth and baclofen with more weakness. There is fair evidence that cyclobenzaprine, carisoprodol, orphenadrine, and tizanidine are effective compared to placebo in patients with musculoskeletal conditions (primarily acute back or neck pain). Cyclobenzaprine has been evaluated in the most clinical trials and has consistently been found to be effective. There is very limited or inconsistent data regarding the effectiveness of metaxalone, methocarbamol, chlorzoxazone, baclofen, or dantrolene compared to placebo in patients with musculoskeletal conditions. There is insufficient evidence to determine the relative efficacy or safety of cyclobenzaprine, carisoprodol, orphenadrine, tizanidine, metaxalone, methocarbamol, and chlorzoxazone. Dantrolene, and to a lesser degree chlorzoxazone, have been associated with rare serious hepatotoxicity. PMID- 15276196 TI - A reassessment of trends in the medical use and abuse of opioid analgesics and implications for diversion control: 1997-2002. AB - This study updates a previous analysis of trends in medical use and abuse of opioid analgesics, and provides data from 1997 through 2002. Two research questions were evaluated: 1) What are the trends in the medical use and abuse of frequently prescribed opioid analgesics used to treat severe pain, including fentanyl, hydromorphone, meperidine, morphine, and oxycodone? 2) What is the abuse trend for opioid analgesics as a class compared to trends in the abuse of other drug classes? Results demonstrated marked increases in medical use and abuse of four of the five studied opioid analgesics. In 2002, opioid analgesics accounted for 9.85% of all drug abuse, up from 5.75% in 1997. Increase in medical use of opioids is a general indicator of progress in providing pain relief. Increases in abuse of opioids is a growing public health problem and should be addressed by identifying the causes and sources of diversion, without interfering with legitimate medical practice and patient care. PMID- 15276197 TI - Molecular events associated with reactive oxygen species and cell cycle progression in mammalian cells. AB - Cell cycle progression is regulated by a wide variety of external factors, amongst them are growth factors and extracellular matrix factors. During the last decades evidence has been obtained that reactive oxygen species (ROS) may also play an important role in cell cycle progression. ROS may be generated by external and internal factors. In this overview we describe briefly the generation of ROS and their effects on processes that have been demonstrated to play an essential role in cell cycle progression, including such systems as signal transduction cascades, protein ubiquitination and degradation, and the cytoskeleton. These different effects of ROS influence cell cycle progression dependent upon the amount and duration of ROS exposure. Activation of growth factor stimulated signaling cascades by low levels of ROS result in increased cell cycle progression, or, in case of prolonged exposure, to a differentiation like growth arrest. From many studies it seems clear that the cyclin kinase inhibitor protein p21 plays a prominent role, leading to cell cycle arrest at higher but not directly lethal levels of ROS. Dependent upon the nature of p21 induction, the cell cycle arrest may be transient, coupled to repair processes, or permanent. At high concentrations of ROS all of the above processes are activated, in combination with enhanced damage to the building blocks of the cell, leading to apoptosis or even necrosis. PMID- 15276198 TI - Cellular control of gene expression by T-type cyclin/CDK9 complexes. AB - The family of Cyclin-Dependent Kinases (CDKs) can be subdivided into two major functional groups based on their roles in cell cycle and/or transcriptional control. This review is centered on CDK9, which is activated by T-type cyclins and cyclin K generating distinct Positive-Transcription Elongation Factors termed P-TEFb. P-TEFb positively regulates transcriptional elongation by phosphorylating the C-terminal domain (CTD) of RNA polymerase II (RNA pol II), as well as negative elongation factors, which block elongation by RNA pol II shortly after the initiation of transcription. Work over the past few years has led to a dramatic increase in our understanding of how productive transcriptional elongation occurs. This review will briefly describe the mechanisms regulating the activity of T-type cyclin/CDK9 complexes and discuss how these complexes regulate gene expression. For further information, the reader is directed to excellent existing reviews on transcriptional elongation and HIV transcription. PMID- 15276199 TI - Identification, expression, modeled structure and serological characterization of Plasmodium vivax histone 2B. AB - Histones play important role in DNA packaging, replication and gene expression. Here, we describe the isolation and characterization of histone 2B (PvH2B) gene from the most common but non-cultivable human malaria parasite Plasmodium vivax. The isolated cDNA clone of PvH2B was allowed to express in Escherichia coli and the recombinant protein was purified by affinity chromatography. The expressed PvH2B protein showed DNA-binding properties on the South-Western analysis and the confocal microscopy localized it in the parasite nucleus. This gene is actively expressed during blood stages of the parasite and all P. vivax patients produced antibodies against the protein. The mRNA of PvH2B was found to contain a poly(A) tail at its 3' end, unlike abundant mRNA of human H2B. The encoded polypeptide is 118 amino acid long contains a nuclear targeting site, a signature motif of H2B and showed 74% homology to its host molecule. The structure of PvH2B showed that it has certain differences from that of its host at critical functional sites (viz acetylation, methylation, trypsin cleavage, DNA-binding and inter-histone interaction) which are required for general gene expression and DNA packaging. The distinctive structural features of P. vivax H2B described here may help in designing the specific antimalarial drugs. PMID- 15276200 TI - Cloning and characterization of a novel endoplasmic reticulum localized G-patch domain protein, IER3IP1. AB - The endoplasmic reticulum (ER) is the site of protein synthesis, folding, post translational modifications and export. The ER membrane and its lumen contain various chaperones and enzymes that are involved in every aspect of the ER function. In this report, we identified a novel endoplasmic reticulum protein (immediate early response 3 interacting protein 1, IER3IP1) during the large scale partial sequencing of a liver cDNA library. The full-length 1304 bp IER3IP1 cDNA has a predicted open reading frame (ORF), which encodes an 82 amino-acid protein possessing a G-patch domain. This domain is found in several RNA associated proteins and has been suggested to be involved in RNA binding. IER3IP1 gene was mapped to the chromosome 18q12 by radiation hybrid analysis. In northern blot hybridization, it was shown that IER3IP1 gene has a high expression in heart, skeletal muscle and kidney, a moderate expression in liver and brain and a low expression in placenta, lung and peripheral blood leukocyte. With the presence of transmembrane domain at the C-terminal, the translated IER3IP1 protein was localized to endoplasmic reticulum of HepG2 cells and was confirmed by co-localization with ER specific marker. PMID- 15276201 TI - Five POTE paralogs and their splice variants are expressed in human prostate and encode proteins of different lengths. AB - POTE is a new gene that contains ankyrin and spectrin domains and is expressed in prostate, testis, ovary, and placenta. In humans, 10 highly homologous variants of the gene are dispersed among eight chromosomes. POTE paralogs are detected in primates but not in other species. Using prostate RNA, we characterized cDNAs from five paralogs and their splice variants. The proteins encoded by the POTE paralogs and their variants range from 80 to 32 kDa. Transfection of POTE constructs into 293T cells shows that the POTE protein, like spectrin, is localized on the inner aspect of the plasma membrane. We also detect a noncoding transcript expressed on the opposite strand from POTE on chromosome 14 or 22. We speculate that POTE has an important signaling function in the reproductive system. PMID- 15276202 TI - Bitis gabonica (Gaboon viper) snake venom gland: toward a catalog for the full length transcripts (cDNA) and proteins. AB - The venom gland of the snake Bitis gabonica (Gaboon viper) was used for the first time to construct a unidirectional cDNA phage library followed by high-throughput sequencing and bioinformatic analysis. Hundreds of cDNAs were obtained and clustered into contigs. We found mostly novel full-length cDNA coding for metalloproteases (P-II and P-III classes), Lys49-phospholipase A2, serine proteases with essential mutations in the active site, Kunitz protease inhibitors, several C-type lectins, bradykinin-potentiating peptide, vascular endothelial growth factor, nucleotidases and nucleases, nerve growth factor, and L-amino acid oxidases. Two new members of the recently described short coding region family of disintegrin, displaying RGD and MLD motifs are reported. In addition, we have identified for the first time a cytokine-like molecule and a multi-Kunitz protease inhibitor in snake venoms. The CLUSTAL alignment and the unrooted cladograms for selected families of B. gabonica venom proteins are also presented. A significant number of sequences were devoid of database matches, suggesting that their biologic function remains to be identified. This paper also reports the N-terminus of the 15 most abundant venom proteins and the sequences matching their corresponding transcripts. The electronic version of this manuscript, available on request, contains spreadsheets with hyperlinks to FASTA formatted files for each contig and the best match to the GenBank and Conserved Domain Databases, in addition to CLUSTAL alignments of each contig. We have thus generated a comprehensive catalog of the B. gabonica venom gland, containing for each secreted protein: (i) the predicted molecular weight, (ii) the predicted isoelectric point, (iii) the accession number, and (iv) the putative function. The role of these molecules is discussed in the context of the envenomation caused by the Gaboon viper. PMID- 15276203 TI - The SV40 T antigen nuclear localization sequence enhances nuclear import of vector DNA in embryos of a crustacean (Litopenaeus schmitti). AB - A genetic transformation system for penaeid shrimp could provide a powerful technique for the improvement of different production traits of importance for a sustainable aquaculture. The development of a successful transformation system depends on the ability to efficiently introduce exogenous DNA into the target species. The ability of the nuclear localization signal (NLS) peptide of the SV40 T antigen to facilitate nuclear import and transient gene expression is known from vertebrate systems and for the first time, is shown here to be efficient in a crustacean species, i.e. the shrimp Litopenaeus schmitti. Electroporation was used to introduce the pCMV-lacZ plasmid that contains the human cytomegalovirus promoter/enhancer (CMV) fused to the beta-galactosidase (lacZ) coding region, into L. schmitti zygotes. Supercoiled DNA was used at 50 or 500 ng/microl naked or bound to NLS peptide. The hatching rate of electroporated zygotes was around 60% for all groups, except from the pCMV-lacZ:NLS group at 500 ng/microl (43%). Based on Southern blot analyses of polymerase chain reaction (PCR) products the gene transfer frequency was 2-fold higher using DNA:NLS complexes than with naked DNA (23.8% vs. 11.5%, with 50 ng/microl of plasmid DNA, 44.3% vs. 28.8% with 500 ng/microl). The beta-galactosidase activity assay indicated that nuclear uptake is faster for the DNA:NLS complexes than for naked DNA. The beta-galactosidase activity was always higher in the DNA:NLS groups than in the naked DNA groups. To our knowledge, this is the first report on the use of an NLS peptide to improve gene transfer and nuclear uptake in crustaceans. PMID- 15276204 TI - Expression of proprotein convertase 2 mRNA in the ovarian follicles of the medaka, Oryzias latipes. AB - Proprotein convertases (PCs) are enzymes responsible for processing the precursors of many bioactive peptides in vertebrates and invertebrates. In the present study, a cDNA for proprotein convertase 2 (PC2) was cloned for the first time from a fish. The clone, which was isolated from the ovary of the medaka, Oryzias latipes, by a combination of RT-PCR cloning and 5'- and 3'-rapid amplification of cDNA ends, codes for a protein of 641 amino acid residues highly homologous to other vertebrate PC2. The medaka preproPC2 consists of a signal sequence, a propeptide with sites for autocatalytic activation, a Kex2-like catalytic domain, and a P-domain. The catalytic triad residues (Asp-169, His-210, and Ser-386) were all conserved. Northern blot analysis revealed that PC2 was expressed in the brain, ovary, and kidney of the fish. The size of PC2 mRNA expressed in the ovary was 2.3 kb, whereas those of the brain and kidney were 2.8 kb. This size difference was attributed to the lack of an approximately 300-bp nucleotide sequence just before the poly(A)+ tail of the ovarian PC2 mRNA. Ovarian expression of the PC2 gene was found in the medaka but not in the mouse, and therefore further analysis was conducted for the fish ovary. The greatest expression of PC2 mRNA in the oocytes of small growing follicles in the mature medaka was demonstrated by Northern blotting, RT-PCR and in situ hybridization analysis. These results suggest that PC2 may play a role in the processing of proproteins and/or pro-hormones expressed in the growing oocytes. PMID- 15276205 TI - Negative and positive effects of an IAP-LTR on nearby Pcdaalpha gene expression in the central nervous system and neuroblastoma cell lines. AB - Intracisternal A-particles (IAPs) are defective retrovirions encoded by members of a large family of endogenous proviral elements in the murine genome. An intact IAP element was found in the protocadherin alpha (Pcdhalpha) gene cluster of five laboratory mouse strains. However, IAP insertion was not detected in three wild mouse strains we investigated. This IAP insertion caused the disruption of one variable exon of laboratory mouse and down-regulated expression of the Pcdhalpha v8 exon, which is located just downstream of the IAP in the brain following the methylation of 5' regulatory region of Pcdhalpha v8. In contrast, the Pcdhalpha v8 exon was highly expressed in mouse neuroblastoma cell lines. This suggested that the IAP insertion activates the expression of the nearby Pcdhalpha v8 exon in these cell lines. In fact, the Pcdhalpha v8 exon expression was driven by the IAP-long terminal repeat (LTR) following the de-methylation of 5' regulatory region of Pcdhalpha v8. To investigate the promoter activity of the IAP, we constructed an IAP-LTR-ECFP reporter gene and introduced it into neuroblastoma, melanoma, lymphoma, and plasmacytoma cell lines. Interestingly, ECFP-positive cells were observed only in the neuroblastoma cell lines. Moreover, there were no differences in the promoter activities of the IAP-LTR whether it was in the sense or complimentary orientation. Thus, this IAP-LTR has negative and positive regulation on near by gene expression in the brain and neuroblastoma cell lines. PMID- 15276206 TI - Structural and expression analysis of the porcine FUS2 gene. AB - The putative porcine tumor suppressor gene FUS2 or N-acetyltransferase (Nat6) assigned to SSC13q21 spans 864-basepairs (bp) of genomic DNA, consisting of a single exon encoding a protein of 288 amino acids (aa), with 73% identity to the human and 74% to the mouse protein. Similar to man and mouse, the gene possesses an N-acetyltransferase domain, but the cell attachment motif arginine-glycine aspartate (RGD) is exclusively found in the pig gene. Expression studies of the gene in several organs by RT-amplification and by quantitative polymerase chain reaction (Q-PCR) showed that FUS2 is widely expressed in porcine tissues. A point mutation was detected at position 836 of the coding sequence (G to A) leading to an amino acid substitution from cystein (C) to tyrosine (Y) at position 278 of the protein. Genes of the tumor suppressor gene (TSG) cluster act together to suppress tumor growth through their functional activation of tumor suppressing pathways. Studies in humans have proven that mutations in N-acetyltransferase genes are associated with some kind of cancers. Knowledge of structure and function of the respective porcine genes and proteins is important. Pigs-in particular minipigs-will be the non-rodent biomodels for human oncology and cancer therapy in the future. PMID- 15276207 TI - A genetic, non-transcriptional assay for nuclear receptor ligand binding in yeast. AB - We describe the development of a genetic, non-transcriptional assay for the detection of ligand binding to nuclear receptors based on the ligand-dependent reconstitution of the defective Ras/cAMP viability pathway of the Saccharomyces cerevisiae strain CDC25-2. We have characterized the assay using the estrogen receptor (ER) alpha as an example and found it to be extremely sensitive, stringent, rapid and selective. We applied this assay to different ligands and ligand-binding domains (LBDs) and analyzed co-stimulation with 17beta-estradiol (E2) and the synthetic ligand 4-hydroxytamoxifen (4-OHT) in vivo. This simple and inexpensive assay may be useful for the study of steroid hormone receptor (SHR) actions at the plasma membrane and for the analysis of ligand binding in vivo. Furthermore, it may allow for the selection of novel ligands and ligand-binding domains and has significant potential for application in compound screening. PMID- 15276208 TI - DNase I hypersensitive sites and histone acetylation status in the chicken Ig beta locus. AB - DNase I hypersensitive sites (DHSs) and histone acetylation status were examined in the Ig-beta locus of chicken B lymphocyte-derived DT40 cells and liver-derived LMH cells. Twelve DT40-specific DHSs were identified: one in the Ig-beta promoter, one in the first intron of the Ig-beta gene, three in the sodium channel gene located upstream of the Ig-beta gene, two between the sodium channel gene and the Ig-beta gene, four between the Ig-beta gene and a downstream growth hormone (GH) gene, and one in the downstream region of the GH gene. Transient transfection studies show that the DHS in the intron of Ig-beta gene enhances the activity of the Ig-beta promoter fourfold. A 1.6 kb DNA fragment, which includes two DHSs, from the sodium channel gene enhanced promoter activity threefold. The transcription enhancing ability of the intron DHS was dependent on orientation, but was not promoter specific. Electrophoretic mobility shift assays (EMSA) demonstrated that an Ets protein family member binds to the intron DHS. In DT40 cells, a distinguished acetylation of H3 and H4 histones was found at the Ig-beta promoter, in addition to the enhanced acetylation of both histones at DT40 specific DHSs. PMID- 15276209 TI - DdAlix, an Alix/AIP1 homolog in Dictyostelium discoideum, is required for multicellular development under low Ca2+ conditions. AB - Apoptosis-linked gene 2 (ALG-2) interacting protein X (Alix), also called AIP1, is a widely conserved protein in eukaryotes. Alix and its homologs are involved in various phenomena such as apoptosis, regulation of cell adhesion, protein sorting, adaptation to stress conditions, and budding of human immunodeficiency virus (HIV). To investigate the role of Alix in development, we identified an Alix homolog in the cellular slime mold Dictyostelium discoideum and disrupted the gene by homologous recombination. The growth of DdAlix deletion mutant (alx-) cells was significantly impaired in the presence of 5 mM Li+. On an agar plate, alx- cells underwent normal development and formed fruiting bodies indistinguishable from those formed by wild-type cells. However, alx- cells could not form fruiting bodies in the presence of 5 mM Li+. Similar results were obtained when cells were developed in the presence of 3,4,5-trimethoxybenzoic acid 8-(diethylamino)octyl ester (TMB-8), which is an antagonist of intracellular Ca2+ store. Furthermore, when the extracellular free Ca2+ was reduced to 10 nM, the ability of alx- cells, but not that of wild-type cells, to form fruiting bodies was impaired. The results indicate that DdAlix is essential for development under low Ca2+ conditions and suggest that DdAlix is involved in Ca2+ signaling during development. PMID- 15276210 TI - Regulation of alternative SRC promoter usage in HepG2 hepatocellular carcinoma cells. AB - Alternative promoters allow for increased spatial and temporal diversity in expression patterns for a single gene. The human SRC gene, encoding the non receptor c-Src tyrosine kinase, is regulated by two alternative promoters separated by approximately 1 kb. The distal SRC1alpha promoter is tissue restricted, while expression of the proximal SRC1A promoter appears to be ubiquitous. A barrier to elucidating the mechanisms of SRC transcriptional regulation has been the finding that the individual strengths of the SRC promoters in isolation do not match their relative strength of use seen in vivo. For example, in HepG2 hepatocellular carcinoma cells, SRC1A is significantly stronger in isolation than SRC1alpha, despite SRC1alpha being the predominant promoter used in this cell line. Previously, we have shown that HepG2 cells, as well as various colon cancer cell lines, display activated SRC transcription, which is linked to the elevated c-Src expression and activity necessary for growth and survival of these cells. These findings thus highlight the importance of understanding the mechanisms of SRC transcriptional regulation in human cancer. We hypothesize the discrepancy between individual SRC promoter strength and relative usage in vivo stems from a lack of linked promoter context. Therefore, we have developed and validated a novel dual SRC promoter reporter strategy to allow the simultaneous mechanistic study of both SRC promoters in their natural linked context. This approach has yielded evidence that SRC activation proceeds through genomic element(s) outside the promoter region in HepG2 cells. Therefore, we performed a preliminary study of DNaseI hypersensitive (DH) site composition within the SRC locus. This approach identified a HepG2 specific DH site that displayed activating potential towards the SRC1alpha promoter. These results thus provide important insight to the mechanism of SRC transcriptional activation in liver cancer cells. PMID- 15276211 TI - Zebrafish cellular nucleic acid-binding protein: gene structure and developmental behaviour. AB - Here we analyse the structural organisation and expression of the zebrafish cellular nucleic acid-binding protein (zCNBP) gene and protein. The gene is organised in five exons and four introns. A noteworthy feature of the gene is the absence of a predicted promoter region. The coding region encodes a 163-amino acid polypeptide with the highly conserved general structural organisation of seven CCHC Zn knuckle domains and an RGG box between the first and the second Zn knuckles. Although theoretical alternative splicing is possible, only one form of zCNBP is actually detected. This form is able to bind to single-stranded DNA and RNA probes in vitro. The analysis of zCNBP developmental expression shows a high amount of CNBP-mRNA in ovary and during the first developmental stages. CNBP-mRNA levels decrease while early development progresses until the midblastula transition (MBT) stage and increases again thereafter. The protein is localised in the cytoplasm of blastomeres whereas it is mainly nuclear in developmental stages after the MBT. These findings suggest that CNBP is a strikingly conserved single-stranded nucleic acid-binding protein which might interact with maternal mRNA during its storage in the embryo cell cytoplasm. It becomes nuclear once MBT takes place possibly in order to modulate zygotic transcription and/or to associate with newly synthesised transcripts. PMID- 15276212 TI - Functional analysis of the rat cytochrome c oxidase subunit 6A1 promoter in primary neurons. AB - Cytochrome c oxidase (COX) is a multimeric enzyme consisting of 13 subunits that are encoded in both mitochondrial and nuclear genomes. We analyzed the promoter of the rat gene encoding the liver isoform of COX subunit VIa. Using transiently transfected primary neuronal cultures as a model system, we found that the basal promoter activity of this gene is localized to a region between positions -244 and +58 relative to the transcriptional start site. This region contains putative binding sites for the transcription factors Sp1, NRF-1, and NRF-2. Two of the NRF 2 sites in this basal promoter are organized in a tandem repeat. A deletion that disrupted this tandem repeat reduced transcription to approximately 25% of the basal level. Additional small deletion series and point mutation experiments confirmed the presence of two functional NRF-2 sites arranged in a tandem repeat, as well as a NRF-1 site and an Sp1 site. In vivo binding of NRF-2 to the rCOX6A1 promoter was confirmed with chromatin immunoprecipitation assay (ChIP). We conclude that Sp1, NRF-1, and NRF-2 are important in activating transcription of the rat COX6A1 gene. PMID- 15276213 TI - Isolation and characterization of two serpentine membrane proteins containing glycerophosphodiester phosphodiesterase, GDE2 and GDE6. AB - Serpentine membrane protein with a glycerophosphodiester phosphodiesterase (GP PDE) motif, GDE3, is involved in morphological change of cells and accelerates the program of osteoblast differentiation, suggesting that mammalian GP-PDEs play an important role in the regulation of cytoskeletal modification. Here, we isolated two cDNAs encoding serpentine membrane proteins, GDE2 and GDE6, containing GP-PDE motif from mouse cDNA libraries. The deduced sequence of GDE2 contains 607 amino acids with seven putative transmembrane regions. GDE6 was composed of 633 amino acids also with seven putative transmembrane regions. In amino acid sequences, GDE2 and GDE6 are 43.7% and 34.3% identical to GDE3, respectively. Although GDE3 mRNA is highly expressed in bone tissue and spleen, GDE2 mRNA was expressed in a variety of mouse tissues including lung and heart, while the GDE6 transcript was particularly abundant in spermatocytes of mouse testis. Immunohistochemical analyses using anti-GDE2 antibody showed that GDE2 protein is expressed in the epithelial cell layer of mouse lung. These results suggest that GP-PDEs are differentially expressed in mouse tissues, and might have distinct roles. PMID- 15276214 TI - Molecular cloning, sequence and expression pattern analysis of the mouse orthologue of the leukemia-associated guanine nucleotide exchange factor. AB - Guanine nucleotide exchange factors (GEFs) of the Dbl family activate Rho proteins and thereby participate in diverse signaling pathways involving this family of small GTPases. However, specific role of Dbl-GEFs in developmental processes, particularly cell differentiation, remains largely unexplored. Recently, a novel member of the Dbl family, leukemia-associated Rho GEF factor (LARG), has been reported to be a fusion partner with mixed-lineage leukemia (MLL) protein in the acute myeloid leukemia, suggesting potential involvement of LARG in regulation of hematopoiesis. In this study, we describe the isolation of the cDNA copy and analysis of genomic structure and expression profile of the mouse orthologue of the human LARG gene. The mouse LARG (mLARG) gene contains 42 exons. The mLARG cDNA is 10,040 bp long and contains a 4631-bp open reading frame (ORF). The predicted mLARG protein shares an overall 89% identity with its human counterpart and contains the same functional domains, namely, Dbl homology (DH), pleckstrin homology (PH), PSD-95, Dlg and ZO-1/2 (PDZ) and regulator of G protein signaling (RGS) domains, as well as two putative signals of nuclear localization. The mLARG message is expressed in all tissues studied, with comparably higher expression levels observed in brain, lung, testis, liver and heart. Using amplified cDNA samples from sorted hematopoietic cells, we showed that mLARG is highly expressed in hematopoietic stem cell fractions, as well as in immature erythroid cells. These results demonstrate high similarity between mouse and human LARG proteins and genes and provide further support to the hypothesis of the potential involvement of LARG in regulation of early stages of hematopoiesis. PMID- 15276215 TI - Sequencing and analysis of the large virulence plasmid pLVPK of Klebsiella pneumoniae CG43. AB - We have determined the entire DNA sequence of pLVPK, which is a 219-kb virulence plasmid harbored in a bacteremic isolate of Klebsiella pneumoniae. A total of 251 open reading frames (ORFs) were annotated, of which 37% have homologous genes of known function, 31% match the hypothetical genes in the GenBank database, and the remaining 32% are novel sequences. The obvious virulence-associated genes carried by the plasmid are the capsular polysaccharide synthesis regulator rmpA and its homolog rmpA2, and multiple iron-acquisition systems, including iucABCDiutA and iroBCDN siderophore gene clusters, Mesorhizobium loti fepBC ABC-type transporter, and Escherichia coli fecIRA, which encodes a Fur-dependent regulatory system for iron uptake. In addition, several gene clusters homologous with copper, silver, lead, and tellurite resistance genes of other bacteria were also identified. Identification of a replication origin consisting of a repA gene lying in between two sets of iterons suggests that the replication of pLVPK is iteron-controlled and the iterons are the binding sites for the repA to initiate replication and maintain copy number of the plasmid. Genes homologous with E. coli sopA/sopB and parA/parB with nearby direct DNA repeats were also identified indicating the presence of an F plasmid-like partitioning system. Finally, the presence of 13 insertion sequences located mostly at the boundaries of the aforementioned gene clusters suggests that pLVPK was derived from a sequential assembly of various horizontally acquired DNA fragments. PMID- 15276216 TI - Isolation of a novel USF2 isoform: repressor of cathepsin B expression. AB - We previously demonstrated that upstream stimulatory factor 1 (USF1) and USF2 regulate transcription of cathepsin B. Here, we have cloned a novel transcript variant of USF2 from a human DU145 prostate cancer cell line by reverse transcription-polymerase chain reaction (RT-PCR). This new transcript variant, designated USF2c, results from alternative splicing of the primary USF2 transcript using a cryptic splicing acceptor site within exon 6. As a consequence, USF2c is missing exons 4, 5, and part of exon 6. USF2c can be transcribed and translated to a protein of approximately 29 kDa in vitro, and the resulting USF2c protein can bind as a homodimer to the E-box of the cathepsin B promoter. USF2c is expressed in two other prostate cancer cell lines (LNCaP, PC3), and U87 human glioblastoma cells as are USF2a and USF2b, two previously identified isoforms of USF2. Cotransfection experiments in DU145 and U87 cells demonstrate that USF2c can down-regulate expression of cathepsin B. These results suggest that USF2c regulates expression of cathepsin B by binding to the E box element in the cathepsin B promoter as a repressor. PMID- 15276217 TI - Choosing a behavioral therapy platform for pharmacotherapy of substance users. AB - Behavioral therapy platforms have become virtual requirements in pharmacotherapy trials due to their utility in reducing noise variability, preventing differential medication adherence and protocol attrition, enhancing statistical power and addressing ethical issues in placebo-controlled trials. Selecting an appropriate behavioral platform for a particular trial requires study-specific tailoring, taking into account both the stage of development of the medication being evaluated, as well as the specific strengths and weaknesses of a broad array of available empirically supported behavioral therapies and the range of their possible targets (e.g., enhancing medication adherence, preventing attrition, addressing co-morbid problems, fostering abstinence, and targeting specific weaknesses of the pharmacologic agent). Choosing a suitable behavioral platform also requires consideration of the characteristics of the population to be treated, stage of scientific knowledge regarding the medication's effects, appropriate balance of internal and external validity, and consideration of potential ceiling effects. Available manualized behavioral treatments are reviewed, noting their strengths and limitations as behavioral therapy platforms for pharmacotherapy trials and as potential concomitant therapies in clinical practice. PMID- 15276218 TI - Residual neuropsychological effects of illicit 3,4-methylenedioxymethamphetamine (MDMA) in individuals with minimal exposure to other drugs. AB - BACKGROUND: A substantial literature suggests that users of illicit 3,4 methylenedioxymethamphetamine (MDMA or "ecstasy") display residual cognitive deficits. Most MDMA users, however, use other illicit drugs as well, so it is difficult to be certain that these deficits are due to MDMA, as opposed to other drug use or additional confounding factors. METHODS: We administered a battery of neuropsychological tests to 23 young MDMA users who reported minimal exposure to any other drugs, including alcohol, and to 16 comparison individuals equally involved with the rave subculture, but reporting no MDMA use. We compared the groups by regression analyses adjusting for numerous potentially confounding variables. To test for a possible dose-response effect, we also performed a median split of 12 moderate MDMA users (22-50 lifetime uses) and 11 heavy users (60-450 uses), and compared these subgroups with non-users. RESULTS: MDMA users as a whole performed worse than non-users on most test measures, but these comparisons rarely reached statistical significance. This picture changed markedly in the subgroup analysis: although moderate users displayed virtually no differences from non-users on any measures, the heavy users displayed significant deficits on many measures, particularly those associated with mental processing speed and impulsivity. These differences did not appear explainable by differences in family-of-origin variables, verbal IQ, levels of depression, or time since last MDMA use. CONCLUSIONS: The presence of residual cognitive deficits, even among unusually "pure" frequent users of illicit MDMA, analyzed with adjustment for confounding variables, augments the evidence that MDMA itself, rather than some associated factor, is responsible for the deficits observed. PMID- 15276219 TI - Have Halpern et al. (2004) detected 'residual neuropsychological effects' of MDMA? Not likely. PMID- 15276221 TI - The interplay between help-seeking and alcohol-related outcomes: divergent processes for professional treatment and self-help groups. AB - This study examined the influence of self-selection, as reflected in alcohol related functioning, on the duration of professional treatment and Alcoholics Anonymous (AA), and the influence of social causation, as reflected in the duration of treatment and AA, on alcohol-related outcomes. A sample of alcoholic individuals was surveyed at baseline and 1, 3, and 8 years later. At each point, participants completed an inventory that assessed participation in treatment and AA since the last assessment and alcohol-related functioning. There were divergent processes of self-selection and social causation with respect to the duration of participation in professional treatment and AA. Individuals with more severe alcohol-related problems obtained longer episodes of professional treatment, but this self-selection process was much less evident for AA. Longer participation in professional treatment in the first year predicted better alcohol-related outcomes; however, the duration of subsequent treatment was not associated with better subsequent outcomes. In contrast, longer participation in AA consistently predicted better subsequent alcohol-related outcomes. These findings are consistent with a need-based model of professional treatment, in which more treatment is selected by and allocated to individuals with more severe problems, and an egalitarian model of self-help, in which need factors play little or no role in continued participation. PMID- 15276222 TI - Mate similarity for substance dependence and antisocial personality disorder symptoms among parents of patients and controls. AB - Substance dependence (SD) and antisocial personality disorder (ASPD) are highly comorbid and aggregate in families. Mating assortment may be an important process contributing to this familial aggregation. HYPOTHESIS: Symptom counts of substance dependence, antisocial personality disorder, and retrospectively assessed conduct disorder (CD) will be correlated significantly among parents of youth in treatment for substance use and conduct problems and, separately, among parents of community controls. METHODS: We examined SD, ASPD, and CD among 151 pairs of parents of adolescents in treatment for substance use and conduct problems, and in 206 pairs of parents of control subjects. RESULTS: For average dependence symptoms (ADS) (the sum of across-drug substance dependence symptoms divided by the number of substance categories meeting minimum threshold use) mother-father correlations were 0.40 for patients and 0.28 for controls. Mother- father correlations for ASPD symptom count were 0.33 for patients and 0.26 for controls and for CD symptom count were 0.31 for patients (all P < 0.01) and 0.10 for controls (P = 0.14). CONCLUSIONS: Spousal correlations for ADS and ASPD, suggest substantial non-random mating. Results support gender differences in homogamy for SD. Behavior genetic studies of these disorders need to account for assortment to avoid biases in estimates of genetic and environmental effects. PMID- 15276223 TI - Association between injection practices and duration of injection among recently initiated injection drug users. AB - BACKGROUND: Earlier studies suggest higher infection risk among recently initiated injection drug users (IDUs) than more experienced users. Whether IDUs' risky injection practices rise progressively with duration of injection or frequency of practices is higher near initiation and then taper remains an open question. METHODS: Recently initiated IDUs were street recruited and interviewed between 1997 and 1999 as part of a multisite cohort study in five US urban cities. Recent risky injection practices (injecting with others and injecting on average more now) were examined across three cross-sections defined by duration of injection: 0-1 year, 2-3 years, and 4-6 years. RESULTS: The IDU groups of <2 years duration (n = 691) and 2-3 years duration (n = 697) had higher odds than the 4-6 year group (n = 520) of reporting injecting with others (Odds Ratio, OR = 1.52, and OR = 1.47, respectively) and injecting on average more now (OR = 1.44 and OR = 1.44, respectively). The associations remained after multivariate adjustment for demographic variables. CONCLUSIONS: These data on recently initiated IDUs suggest that risky injection practices were more frequent earlier than later within the first 6 years of initiation, emphasizing that outreach prevention needs to identify and intervene with IDUs early. PMID- 15276224 TI - Marijuana and cocaine use among female African-American welfare recipients. AB - A key issue that came to the forefront during the welfare reform debate in the United States during the 1990s concerned the relationship between welfare receipt and drug use and abuse. This paper examines the relationship between persistent welfare assistance, welfare background, and marijuana and cocaine use among African-American women. We hypothesize that women who have received welfare assistance for a period of 5 years or more will be more likely to use drugs compared to those who have never received welfare assistance or who have received it for a shorter duration. Data for this analysis comes from a longitudinal study of African-Americans living in a Chicago community followed from first grade (N = 1242) to age 32. Multinomial logistic regression analyses were performed to examine the relationship between years of welfare receipt and three categories of marijuana and cocaine use (never, past, and current) among female respondents (N = 496). Results indicate an increased risk of past-year cocaine and marijuana use for women who reported receiving welfare benefits for 5 years or more. Growing up in a family that received welfare did not significantly predict adult drug use, but did significantly predict an adult welfare experience. Implications of results are discussed. PMID- 15276225 TI - Short-term distribution of nicotine in the rat lung. AB - Nicotine is a dibasic amine with a pK(a) of 8.0. At physiological pH roughly 1/4 of the compound is nonionized and able to cross membranes, most notably the alveolar membranes of the lung. Many models of nicotine addiction assume that the time it takes nicotine to pass from inspired air to the blood stream is negligible, resulting in a large peak or bolus in nicotine blood levels following each puff from a cigarette. However, the results of several previous studies have suggested that the lung may act as a short-term depot for nicotine. This was directly investigated in the present study. In anesthetized rats with open-chest and ventilated lungs, 0.4 mg [(3)H] ]nicotine in 50 microl was rapidly injected into the right ventricle of the heart and blood was sampled from the left ventricle. It was found that the [[(3)H]nicotine left the lungs at a significantly slower rate than [(14)C]dextran, a compound which remains in the plasma compartment (3.11% versus 7.71% injected/s for [(3)H]nicotine and [(14)C]dextran, respectively). In similar experiments, lung, heart and brain tissue were obtained at 5s intervals. Significant [(3)H]nicotine remained in the lung throughout the 40 s period, with lung tissue nicotine greater than brain at all time points. These results indicate that the lung may act as a short-term depot for nicotine, delaying and depressing its appearance in the systemic arterial circulation. PMID- 15276226 TI - Interactions between low concentrations of ethanol and nicotine on firing rate of ventral tegmental dopamine neurones. AB - This study investigated interactions between ethanol and nicotine on dopamine sensitive neurone firing in the ventral tegmental area (VTA), recorded in midbrain slices. No changes in spontaneous activity of the neurones were seen with nicotine at 10, 25, or 100 nM; at 250 nM there was a small significant increase in firing rate. Ethanol, applied alone, caused a significant increase in firing rate at 40 mM and at 60 nM but not at 20 mM. Combinations of 10 or 25 nM nicotine with 20 or 40 mM ethanol did not result in increased firing rates compared with either drug alone. However, nicotine 100 nM plus ethanol 60 mM significantly increased the rate of spontaneous firing compared with that after either drug alone at these concentrations. In contrast, nicotine at 250 nM plus ethanol at 60 mM did not increase firing rate, compared with each drug alone. Ketanserin, 2 microM, prevented the potentiating effect of nicotine 100 nM plus ethanol 60 mM. The results show synergism between ethanol and nicotine at specific concentrations that are likely to be present in the brain during the behavioural effects of these drugs, but the interaction is complex and may involve multiple drug actions. PMID- 15276227 TI - Heterosexual relationships among heroin users in Italy. AB - This study investigated how many stable partners of drug users (DUs) had a history of drug use or were current DUs. Of 589 DUs interviewed, 41% reported that they had a partner with current or previous experience of drug addiction. A strong gender difference emerged: 77% of female DUs reported a stable relationship with partners with a history of addiction, versus only 30% for male DUs. Partners with a history of drug dependence are more likely to be: male, older, with a lower educational level and a lower rate of stable employment than partners without a history of drug addiction. Logistic regression analysis indicated that the characteristics of heroin users who have current partners with histories of drug use include: female gender, older age, living with a partner, lengthy duration of the relationship and HIV positive status. Fewer subjects are married if the partner has a history of addiction, and there is an association between lengthy drug use and partner without drug addiction history. The high percentage (59%) of subjects who were in stable relationships with partners without histories of heroin addiction and the relatively long duration of these relationships, raises the issue of possible transmission of blood-borne viruses from the DUs to their sexual partners. The study does suggest the need for consideration of sexual partnerships and gender differences in providing drug abuse treatment for heroin users. PMID- 15276228 TI - Chronic opiate treatment enhances both cocaine-reinforced and cocaine-seeking behaviors following opiate withdrawal. AB - After chronic exposure to psychostimulants or opiates, self-administration or conditioned place preference with either class is increased (sensitized). Cross sensitization of conditioned place preference, i.e., enhancement of psychostimulant-induced preferences after exposure to opiates, has also been described, but increases in cocaine self-administration after morphine pretreatment have not been reported. The present study evaluated effects of chronic morphine treatment on cocaine reinforcement. Opiate dependence was established in Wistar rats by administration of morphine as a constant infusion that was gradually increased to a dose of 50mg/kg per day over a 1-week period. Immediately after discontinuation of chronic morphine treatment, animals were allowed to acquire cocaine self-administration under a simple fixed-ratio schedule (FR-1), and were subsequently advanced to a progressive ratio schedule. Acquisition of cocaine self-administration under the FR-1 did not differ in saline- and morphine-pretreated animals. For cocaine self-administration under a progressive ratio schedule measured at 5 or more days after the onset of opiate withdrawal, chronic pretreatment with morphine increased the number of ratios completed, augmented final response requirements, and produced a more stable pattern of cocaine self-administration. Responding was also increased in morphine pretreated animals during an initial extinction session. These results show that chronic opiate treatment can enhance both cocaine-reinforced and cocaine-seeking behaviors following opiate withdrawal. A similar effect may occur in human patients who discontinue methadone or other forms of replacement therapy for opiate abuse, and may contribute to relapse involving use of cocaine or other psychostimulants. PMID- 15276229 TI - Adjustment of the vestibulo-ocular reflex gain as a function of perceived target distance in humans. AB - The human vestibulo-ocular reflex (VOR) was investigated during active head movements in yaw while subjects were asked to view targets located at 20, 30, 40, 60, 90, and 120 cm distance aligned with eye level. Binocular video cameras were used to study conjugate eye movements and binocular convergence. Perceived target distance was determined during head oscillation by having the subjects move a cursor to the remembered position of the previously seen targets. The changes in VOR gain with viewing distance were found to be more closely related to perceived target distance than to actual target distance or fixation distance. This result suggests that the adjustment of VOR gain with viewing distance is under stronger cognitive control than would be expected of a simple motor reflex. PMID- 15276230 TI - Novel splice variants increase molecular diversity of aprataxin, the gene responsible for early-onset ataxia with ocular motor apraxia and hypoalbuminemia. AB - Early-onset ataxia with ocular motor apraxia and hypoalbuminemia (EAOH) is one of the most common forms of autosomal recessive cerebellar ataxia. We identified six new alternative transcripts produced by the aprataxin gene responsible for EAOH. Total eight transcripts encoded truncated proteins that were located within the nucleus or cytoplasm and showed different binding abilities to wild-type (WT) aprataxin. Thus, the alternative splicing increases the molecular diversity of aprataxin and the expression profiles of these transcripts in various tissues may be related to the tissue-specific phenotypes. PMID- 15276231 TI - No association between polymorphisms in the lectin-like oxidised low density lipoprotein receptor (ORL1) gene on chromosome 12 and Alzheimer's disease in a UK cohort. AB - Genome scans in sporadic Alzheimer's disease (AD) have revealed possible susceptibility loci on chromosome 12. Recently, two studies were published investigating the +1071 and +1073 polymorphisms in the lectin-like oxidised low density lipoprotein receptor (OLR1) gene with AD, a gene that lies within the area of chromosome 12 linkage. OLR1 is a good candidate gene, due to its function in lipid metabolism pathways, other components of which have been previously implicated as risk factors for AD. We undertook an association study in our UK cohort of 356 AD patients and 358 matched controls, using the same polymorphisms and performing the same sub-group and haplotype analysis as previously described. We found no association with AD in our case-control group as a whole, or when stratified into those with early (<65 years) or late (>65 years) onset. When the group was split into APOE 4 bearers and 4 non-bearers, we could not confirm the associations described in the original study. Similarly, no significant differences were observed between AD patients and controls, in terms of their haplotype distributions. Therefore, in this present study, we find no evidence for the involvement of these ORL1 polymorphisms in increasing susceptibility to AD. PMID- 15276232 TI - Effects of serotonin-dopamine antagonists on prepulse inhibition and neurotransmitter contents in the rat cortex. AB - Perospirone is a serotonin-dopamine antagonist (SDA) recently developed in Japan as an atypical antipsychotic to be used in the treatment of schizophrenia. The amines and amino acids in the cortex are assumed to play an important role in the cognitive dysfunction of schizophrenia. To investigate the acute effect of perospirone on cognition, we compared perospirone to risperidone and haloperidol by assessing their influence on prepulse inhibition (PPI). Moreover, we studied the effects of these drugs on amine and amino acid contents in the rat cortex. Perospirone had a significant influence: PPI, dopamine turnover and glycine contents increased statistically and serotonin decreased statistically in comparison to control levels. Our results suggest that, of the three antipsychotic drugs, only perospirone promotes cognition, and this ability is associated with increase in dopamine turnover, reduction in serotonin turnover and increase in glycine contents. PMID- 15276233 TI - Assessment of Nurr1 nucleotide variations in familial Parkinson's disease. AB - Parkinson's disease (PD) is characterised by the death of dopaminergic neurons of the substantia nigra. As Nurr1 seems to regulate the development and maintenance of these neurons, we evaluated its potential role in Parkinson's disease using genetic methods. We genotyped two polymorphisms and screened a case-control sample for the presence/absence of two mutations recently described in exon 1. Our results failed to replicate the association initially observed and none of the mutations were present in our familial Parkinson's disease cases. These observations suggest that this gene is unlikely to play a major effect in French familial Parkinson disease. PMID- 15276234 TI - No association between the promoter variants of tumor necrosis factor alpha (TNF alpha) and schizophrenia in Chinese Han population. AB - An increasing amount of evidence suggests that the pathophysiology of schizophrenia is associated with activation of the immune system. Four studies have established an association of -308G/A polymorphism of tumor necrosis factor alpha (TNF-alpha), a cytokine involved in inflammatory processes, with schizophrenia [Mol. Psychiatry 6 (2001) 79; Mol. Psychiatry 8 (2003) 718; Schizophr. Res. 65 (2003) 19; Biol. Psychiatry 54 (2003) 1205]. In the present study, however, no significant positive association has been found between any individual SNP or haplotype constituted of the five promoter polymorphisms ( 1031T/C, -863C/A, -857C/T, -308G/A and -238G/A) in the human TNF-alpha gene and schizophrenia (314 Chinese Han schizophrenic patients and 340 healthy control). A meta-analysis we did in this work, which is based on previous nine studies plus our own unpublished data including a total of 2399 schizophrenic patients (sporadic cases 2099, familial cases >505) and more than 3261 controls, failed to show significant difference of -308G/A distribution between patients and controls in both the whole sample and the pooled Asian sample. By contraries, the significant results in the pooled Caucasian sample imply an ethnic heterogeneity in -308G/A variation in the TNF-alpha gene in schizophrenia. PMID- 15276235 TI - Expression of nestin in ballooned neurons in patients with Creutzfeldt-Jakob disease. AB - Nestin is an intermediate filament protein and mainly expressed during the development of the central nervous system. Recently, we reported that nestin is present in the cytoplasm, dendrites, and torpedoes of Purkinje cells in patients from Creutzfeldt-Jakob disease as the pathological change advances. In this study, we focused on the temporal and frontal lobes of the same patients in order to examine the ballooned neurons using anti-human nestin antibody. Although the ballooned neurons showed cytoplasmic immunoreactivity against nestin, the level of nestin immunoreactivity differed in each ballooned neuron. These findings could suggest that ballooned neurons are being reactivated to promote survival, although the role of nestin is not clear. PMID- 15276236 TI - Recurrence quantification analysis of sleep electoencephalogram in sleep apnea syndrome in humans. AB - The aim of this study is to elucidate whether the results of recurrence quantification analysis (RQA) of sleep EEGs in sleep apnea syndrome are valuable for analyzing sleep EEGs in sleep apnea syndrome. We investigated the ability of RQA to discriminate sleep stages and to characterize the different behaviors of sleep EEGs in sleep apnea syndrome. RQA was applied to EEG signals during sleep stages 1, 2, slow wave sleep (SWS), REM and the stage 'awake.' The sleep EEG signals were obtained from the MIT-BIH polysomnographic database. To examine the differences in the RQA measures for all sleep stages, one-way analysis of variance (ANOVA) and post hoc analysis were performed. From the results, all sleep stages could be distinctly discriminated by means of the RQA measure of %RATIO. We observed that stage 1 and REM had fewer recurrences, and that stage 2 was more autocorrelated than the other stages. The different dynamic behaviors of wakefulness and sleep EEG were also observed. Of significant interest was the observation that RQA was able to distinguish stage 1 from REM. In conclusion, we suggest that the information obtained from RQA of sleep EEGs in sleep apnea syndrome is valuable for its analysis, and that RQA constitutes a useful tool for analyzing sleep EEGs in subjects with sleep apnea syndrome. PMID- 15276237 TI - Interamygdaloid connection of basolateral nucleus through the anterior commissure in the rat. AB - Basolateral amygdaloid nucleus interconnections through the anterior commissure (ac) are well visualized using wheat germ agglutinin-horseradish peroxidase tracer (WGA-HRP) in control animals. On the other hand, few neurons in the contralateral basolateral amygdaloid nucleus were labeled when the WGA-HRP was injected after a complete transection of the anterior commissure. These results indicate that the basolateral nucleus projects through the anterior commissure to the homologous amygdaloid nucleus in the contralateral hemisphere. PMID- 15276238 TI - Case-control and family-based association studies between the neuregulin 1 (Arg38Gln) polymorphism and schizophrenia. AB - Genetic variations in the neuregulin 1 (NRG1), a critical gene in neuronal development, have been reported to be associated with schizophrenia in several reports. Association has been reported between a non-synonymous NRG1 polymorphism (Arg38Gln) and schizophrenia in a Chinese family-based association study; however, this finding is not yet confirmed by other research findings analyzed using independent sample. To replicate this finding and assess the association between age at onset of schizophrenia and the NRG1 Arg38Gln polymorphism, we investigated the prevalence of this polymorphism in a Chinese population (228 schizophrenic disorder patients and 269 controls). We were unable, however, to demonstrate a significant association between the NRG1 Arg38Gln polymorphism and schizophrenia (P = 0.869 for genotype and P = 0.597 for allelic frequencies) or age at onset (P = 0.940). Our family-based association study (15 schizophrenic bios and 221 schizophrenic trios) demonstrated 38Gln was transmitted in excess by the parent to the affected offspring (P = 0.052). However, this result contrasts with a previous finding in Chinese that 38Arg was transmitted in excess by the parent to the affected offspring. On the basis of the contrast between the findings of other study and our family-based study and the negative findings of our case-control association study, we conclude that NRG1 Arg38Gln polymorphism is not likely to play a major role in the pathogenesis of schizophrenia in Chinese populations. PMID- 15276239 TI - Application of a novel Active Allothetic Place Avoidance task (AAPA) in testing a pharmacological model of psychosis in rats: comparison with the Morris Water Maze. AB - Administration of a non-competitive NMDA antagonist dizocilpine (MK-801) was proposed to be an animal model of psychosis. NMDA-receptor blockade is accompanied by increased locomotion, behavioral deficits, and other changes resembling psychotic symptoms. However, the role of NMDA-receptors in organizing brain representations is not understood yet. We tested the effect of NMDA receptor blockade by systemic administration of dizocilpine at two different doses (0.1 or 0.2 mg/kg) in a recently designed Active Allothetic Place Avoidance (AAPA), a task which requires rats to separate spatial stimuli from two continuously dissociated subsets. The effect of dizocilpine on learning in the AAPA task was compared with its effect on acquisition of the reference memory version of the Morris Water Maze task. Both doses impaired performance in the Morris Water Maze task, whereas only the higher dose impaired performance in the AAPA task. The Morris Water Maze appears to be more sensitive to dizocilpine induced behavioral deficit than the AAPA task. These findings support the notion that these two tasks are differentially dependent on the NMDA-receptor function. PMID- 15276240 TI - Minocycline suppresses hypoxic activation of rodent microglia in culture. AB - Hypoxia is one of the important physiological stimuli that are often associated with a variety of pathological states such as ischemia, respiratory diseases, and tumorigenesis. In the central nervous system, hypoxia that is accompanied by cerebral ischemia not only causes neuronal cell injury, but may also induce pathological microglial activation. We have previously shown that hypoxia induces inflammatory activation of cultured microglia, and the hypoxic induction of nitric oxide production in microglia is mediated through p38 mitogen-activated protein kinase pathway. Now, we present evidence that minocycline, a tetracycline derivative, suppresses the hypoxic activation of cultured microglia by inhibiting p38 mitogen-activated protein kinase pathway. The drug markedly inhibited hypoxia induced production of inflammatory mediators such as nitric oxide, TNFalpha, and IL-1beta as well as iNOS protein expression. The signal transduction pathway that leads to the activation of p38 mitogen-activated protein kinase was the molecular target of minocycline. Thus, the known neuroprotective effects of minocycline in animal models of cerebral ischemia may be partly due to its direct actions on brain microglia. PMID- 15276241 TI - Wheel running use in dopamine D2L receptor knockout mice. AB - The present study investigated the role of dopamine and the opioid system in the acquisition and maintenance of wheel running use (WR), employing D2L receptor deficient (D2L-/-) mice as a model system. The daily administration of 1mg/kg of naloxone virtually abolished the acquisition of WR in D2L-/-, but did not modify the number of wheel turns after the consolidation of this behavior. These findings imply that both dopamine and opioid reward systems are necessary for WR consolidation, and they also suggest that reward systems in WR could vary according to the motivational stage of the animal. PMID- 15276242 TI - Age-related and regional differences in secretin and secretin receptor mRNA levels in the rat brain. AB - In the present study expression levels of secretin and secretin receptor mRNAs in several brain regions of rats ranging in age from postnatal days 7 to 60 were investigated by quantitative real-time PCR. Expression of secretin and secretin receptor was detected in the central amygdala, hippocampus, area postrema, nucleus of the tractus solitary and cerebellum. The cerebellum expressed secretin receptor at significantly higher levels than that found in other brain regions within all the ages examined. In contrast, secretin mRNA was significantly higher in the nucleus of the tractus solitary than in the other four brain regions examined in postnatal day-21, -30 and -60 rats. Within most brain regions, both secretin and secretin receptor mRNAs were more abundant in postnatal day-7 and 14 rats as compared to postnatal day-21, -30 and -60 rats. Thus, secretin and its receptor are widely expressed in rat brain and the expression of both genes is developmentally regulated during the first few weeks following birth. PMID- 15276243 TI - Choline acetyltransferase G +4 A polymorphism confers a risk for Alzheimer's disease in concert with Apolipoprotein E epsilon4. AB - To examine whether the single nucleotide polymorphism at position +4 of the choline acetyltransferase (ChAT) confers a risk for Alzheimer's disease (AD), we determined the ChAT and the Apolipoprotein (APOE) genotypes of the 246 AD patients and 561 non-demented controls. The ChAT AA genotype was found to confer the risk for AD in concert with the APOE epsilon4 by stochastic search variable selection (SSVS) approach. The odds of the ChAT AA for AD were 3.25 (95% CI = 1.17-9.03). The mean ages-at-onset of AD were lower in those carrying the ChAT AA than those carrying the ChAT AG or ChAT GG, regardless to the occurrence of the APOE epsilon4. The ChAT AA is a novel genetic risk factor AD, and the SSVS is a useful approach for analyzing association with multiple candidate genes simultaneously. PMID- 15276244 TI - NGF-mediated sensitization of the excitability of rat sensory neurons is prevented by a blocking antibody to the p75 neurotrophin receptor. AB - Nerve growth factor (NGF) can play a causal role in the initiation of hyperalgesia. Recent work demonstrates that NGF can act directly on nociceptive sensory neurons to augment their sensitivity to a variety of stimuli. Based on the existing literature, it is not clear whether this sensitization is mediated by the high-affinity TrkA receptor or the low-affinity p75 neurotrophin receptor. We examined whether a blocking antibody to the p75 neurotrophin receptor can prevent the NGF-induced enhancement of excitability in capsaicin-sensitive small diameter sensory neurons that have been isolated from the adult rat. In this report, pretreatment with the p75 blocking antibody completely prevents the NGF induced increase in the number of action potentials evoked by a ramp of depolarizing current as well as the suppression of a delayed rectifier-type of potassium current(s) in these neurons. Although the sensitization by NGF was blocked, the antibody had no effect on the capacity of ceramide, a putative downstream signaling molecule, to either enhance the excitability or inhibit the potassium current. These results indicate that NGF can increase the excitability of nociceptive sensory neurons through activation of the p75 neurotrophin receptor and its consequent liberation of ceramide from neuronal sphingomyelins. PMID- 15276245 TI - TRPV2-immunoreactive intrinsic neurons in the rat intestine. AB - Transient receptor potential channel vanilloid subfamily 2 (TRPV2) was shown to receive noxious thermal stimuli (>52 degrees C), and to be expressed in fine myelinated afferent neurons. The mRNA and the immunoreactivity have also been detected in several peripheral tissues. We examined the expression of TRPV2 in the rat intestine. An analysis by transcriptase-polymerase chain reaction (RT PCR) demonstrated TRPV2 gene expression in the intestine. Many TRPV2-positive neurons were observed in the myenteric plexus by immunohistochemistry. Some of these neurons were positive for calbindin D-28K (CaBP), which is present in intrinsic afferent neurons. TRPV2 immunoreactivity was also observed in nodose ganglion neurons (vagal afferents). These findings suggest that TRPV2 is expressed not only in sensory ganglion neurons, but also in enteric neurons, including primary afferent neurons. PMID- 15276246 TI - Neuropeptide Y up-regulation in cerebrocortical neurons after Borna Disease Virus infection is unrelated to brain inflammation in rats. AB - Neuropeptides participate in the pathophysiology of cerebral inflammatory diseases. We analyzed the involvement of neuropeptide Y (NPY) in rat brain infected with Borna Disease Virus (BDV). NPY expressing cerebrocortical neurons were increased during the acute stage of BDV-induced encephalitis. The increase was resistant to immunosuppression by systemic dexamethasone, which greatly reduced inflammatory reactions in the brain. This indicates that the increase of cerebrocortical NPY expression is not causally related to inflammation. As cerebral NPY is known to be increased during experimental seizures and to have anticonvulsive actions, we propose that NPY up-regulated during BDV encephalitis limits seizures known to be associated with Borna Disease. PMID- 15276247 TI - Changes in AMPA receptor phosphorylation in the rostral ventromedial medulla after inflammatory hyperalgesia in rats. AB - To study the glutamatergic mechanisms underlying changes in excitability in the brain stem pain modulatory circuitry after injury, we examined GluR1 serine 831 phosphorylation in the rostral ventromedial medulla (RVM) after complete Freund's adjuvant-induced hindpaw inflammation. Western blots indicated a rapid and prolonged (30 min and 7 days post-inflammation) increase in phosphoserine 831 GluR1 protein levels in the RVM. The upregulated GluR1 phosphorylation was blocked by pretreatment, but not by post-treatment, with the local anesthetic, lidocaine, at the site of inflammation. The upregulation of phosphoserine 831 GluR1 was attenuated by pretreatment with chelerythrine, a selective PKC inhibitor, KN-93, a selective CaMKII inhibitor, and two NMDA receptor antagonists, MK-801 and APV. These findings provide new evidence linking in vivo AMPA receptor phosphorylation in the RVM pain modulatory circuitry to the enhanced descending pain modulation after inflammation. PMID- 15276248 TI - Anticipatory modulation of neck muscle reflex responses induced by mechanical perturbations of the human forehead. AB - The aim of this study was to test whether anticipation of upcoming head blow stimuli, which elicit reflex responses in the neck muscle, makes the reflex responses greater or not. In nine healthy subjects the reflex responses were elicited in the sternocleidomastoid (SCM) muscle in the eyes-open (EO) and eyes closed (EC) conditions, which corresponded to the predictable and unpredictable conditions, respectively. The subjects were instructed not to resist the perturbations after the impact. The results demonstrated that the reflex response of the SCM muscle was significantly smaller in the predictable EO condition than in the unpredictable EC condition (P < 0.05), although no significant differences were observed in either the background EMG activities or the head accelerations. Further, this effect of anticipation was observed only in the later reflex EMG component, which most probably mediated the stretch reflex pathway. In contrast, no significant difference was observed in the early component, which was presumed to be the vestibular-collic reflex. The reduced stretch reflex response was suggested to be functionally relevant to the task requirement, i.e., to let the neck extension movement occur, and not to resist after the impact of the head blow. It was concluded that the anticipation has an effect on reducing the stretch reflex responses in the neck muscle, but does not have any effect on the presumed vestibular-collic reflex under the present experimental paradigm. It is suggested that the gain of the stretch reflex pathway is modulated by anticipatory information of upcoming mechanical event. PMID- 15276249 TI - Analgesic tolerance and cross-tolerance to i.c.v. endomorphin-1, endomorphin-2, and morphine in mice. AB - The present study examined the development of analgesic tolerance to endomorphin 1 (EM1), endomorphin-2 (EM2), and morphine, and cross-tolerance among these drugs. Male Swiss Webster mice were injected i.c.v. with EM1, EM2, morphine, or vehicle once daily for 5 days, and tested for analgesia in the tail flick test. To determine the extent of cross-tolerance, on the sixth day mice from each of the above groups received i.c.v. injections of EM1, EM2, morphine, or vehicle before analgesic testing. The development of tolerance to EM1 and EM2 closely resembled that of morphine. Complete, symmetrical cross-tolerance was observed between all drugs in the study. These results demonstrate a time-course and extent of tolerance similar to morphine, and support a common mechanism of action through the mu-opioid receptor. PMID- 15276250 TI - Expression of c-fos in the nucleus of the solitary tract following electroacupuncture at facial acupoints and gastric distension in rats. AB - Clinical practice has shown that acupuncture at facial acupoints has curative effects on some visceral diseases (especially gastrointestinal diseases). However, the physiological basis has not been clarified yet. In the present study, expression of c-fos in the nucleus of the solitary tract (NTS) of rats following gastric distension and electroacupuncture (EA) at Yangbai (GB14) and Sibai (ST2) as well as Jiache (ST6) acupoints was observed by using immunohistochemistry technique. After EA at the three facial acupoints, c-fos immunoreactive (c-fos-IR) neurons were mainly distributed in the medial (mNTS) and intermediate subnucleus of the NTS, and a few were scatteredly distributed in the dorsalmedial and commissural subnucleus of the NTS. Furthermore, there is difference in the number of c-fos-IR neurons in the mNTS following EA at the three facial acupoints. The number in the EA at ST2 and GB14 group is the highest and the lowest, respectively. Gastric distension induces obviously the expression of c-fos, which is mainly confined in the mNTS. The results suggest that the noxious visceral and somatic afferent information from the stomach and face may converge in the mNTS, which may be involved in the effect of EA at facial acupoints on the gastrointestinal pain. PMID- 15276251 TI - Mouse spinal cord compression injury is ameliorated by intrathecal cationic manganese(III) porphyrin catalytic antioxidant therapy. AB - This study evaluated the effects of the cationic manganese(III) tetrakis(N,N' diethylimidazolium-2-yl)porphyrin catalytic antioxidant Mn(III)TDE-2-ImP5+ (AEOL 10150) on outcome from spinal cord compression (SCC) in the mouse. C57BL/6J mice were subjected to 60 min thoracic SCC after discontinuation of halothane anesthesia. In Experiment 1, mice were given intravenous Mn(III)TDE-2-ImP5+ (0.5 mg/kg bolus followed by 1 mg kg(-1) h(-1) for 24 h), methylprednisolone (30 mg/kg bolus followed by 5.4 mg kg(-1) h(-1) for 24 h), or vehicle (n = 25 per group). In Experiment 2, mice were given intrathecal Mn(III)TDE-2-ImP5+ (2.5 or 5.0 microg/kg) or vehicle (n = 18 per group). In both experiments, treatment began 5 min post-SCC onset. Rotarod performance was measured on post-SCC days 3, 7, 14, and 21. On post-SCC day 21, the spinal cord was histologically examined and a total damage score was calculated. Neither intravenous Mn(III)TDE-2-ImP5+ nor methylprednisolone altered rotarod performance (accelerated rate P = 0.11, fixed rate P = 0.11) or mean +/- S.D. total damage score (Mn(III)TDE-2-ImP5+ = 21 +/- 9, methylprednisolone = 24 +/- 8, vehicle = 22 +/- 10; P = 0.47; shams = 0). Intrathecal Mn(III)TDE-2-ImP5+ (both 2.5 and 5.0 microg) given at SCC-onset improved rotarod performance (P = 0.05) and total damage score (2.5 microg = 19 +/- 10, P = 0.04; 5.0 microg =19 +/- 8, P = 0.03) versus vehicle (26 +/- 10). These studies demonstrate sustained benefit from manganese(III) porphyrin catalytic antioxidant therapy after SCC. However, efficacy was dependent upon route of administration suggesting that bioavailability is critical in defining efficacy. PMID- 15276252 TI - Thermal analgesia induced by 30-min exposure to 1 microT burst-firing magnetic fields is strongly enhanced in a dose-dependent manner by the alpha2 agonist clonidine in rats. AB - Most of the research concerning analgesia following brief exposures to physiologically patterned weak magnetic fields has focused upon their morphine related properties. However, the alpha-adrenergic system interacts with morphine induced analgesia. In the present study we found that prazosin, phenylephrine, and yohimbine did not augment the robust analgesia to thermal stimuli in rats evoked by whole-body exposures to a 1 microT, burst-firing magnetic field presented once every 4s for 30 min. However, the alpha2 agonist clonidine enhanced the field-induced analgesia in a dose-dependent manner that reflected a receptor-saturation response. Potentiation between the field and clonidine was evident at 0.2 mg/kg and approached asymptote at 1 mg/kg. The combination of the effects from exposure to the magnetic field and the clonidine explained more than 75% of the variance in the change in nociceptive thresholds from baseline levels. The possibility that properly patterned weak magnetic fields could be a powerful adjunct to pharmacological treatments of pain is considered. PMID- 15276253 TI - [Plastic surgery, cleft lip and palate and the Third World]. PMID- 15276254 TI - [Caustic burns. Clinical study of 24 patients with sulfuric acid burns in Cambodia]. AB - Over a period of 12 years, 24 burns have been treated by a Cambodgian and French team (Doctors of the World). This experience has revealed some characteristics of this population: (1) a majority of young women (2/3), (2) victims of assault (20 cases). These notions are found in the medical literature particularly South East Asia. All cases are of a third degree. Every surgeon, every anesthetist should apply emergency measures which are recalled here. The aims is to close all the burns surface through a skin graft within a month. However the patients are treated 6 months to 2 years after the burn and have, at this stage, terrible sequella (22 patients). Treating them requires the coordinated efforts of a multidisciplinary specialized team. The challenge is to save the eyes, the mouth, the nose and give back a "human" face to these young burns who are often rejected because of their handicap. Enabling them to smile again is a strong commitment from the surgeon and from the patient who has to undergo multiple operations and a long treatment over the years. PMID- 15276255 TI - ["Operation Smile" missions in Mongolia]. AB - We report our experience of plastic surgery of the face, limbs and burns in Mongolia. This country is three times as huge as France and has got 2,4 millions of inhabitants of whom almost one million live in the capital city Oulan-Baator, where sanitary and weather conditions are very tough for a large part of the population. The main observed and treated pathologies were sequellae of nerves and vessels lacerations, sequellae of complex hand injuries, sequellae of leg traumas, face, neck and limb burns, lips and palate clefts and pressure shores. The training of Mongol surgeons is far from good in the fields of skin coverage, digits reconstruction, treatment of nerve and vascular injuries and management of burns. Pedicled radial antebrachial, posterior interosseous, inguinal, sural, saphenous and lower limb muscular flaps were performed and taught. Microsurgery is still starting up. Thus we performed the first second toe transfer to reconstruct an amputated thumb. Some clinical cases are presented. This French Mongolian cooperation should continue in order to match between both countries the knowledge and use of standard techniques in plastic surgery. PMID- 15276256 TI - [Palatovelopharyngoplasty at the same time. Our experience in Philippines]. AB - After many surgical charity missions in the same hospital in the Philippines, we have evaluated the child's phonation being operated of cleft palate. We had bad results so we decided to do in the same time an Orticochea pharyngoplasty and the closure of the palate. These patients were evaluated too, the results were improved. PMID- 15276257 TI - [Buruli disease and plastic surgery]. AB - The Buruli ulcer is a skin infection with Mycobacterium ulcerans which progresses silently. This infection affects mostly women and children who live near stagnant waters. Buruli ulcer is disease that has terrible consequences if not promptly diagnosed and treated. It destroys progressively skin tissues and consequently leaves very important scars. There is no efficient medical treatment. This presentation proposes to take care efficiently of the Buruli ulcer by simple plastic surgery techniques. We relate our experience of a mission in Benin, in the context of the national programme of struggle against the Buruli ulcer. PMID- 15276258 TI - [Myanmar mission]. AB - The authors report the accomplishment of humanitarian missions in plastic surgery performed by a small team in town practice in Yangon, about their 3 years experience in Myanmar with 300 consultations and 120 surgery cases. They underline the interest of this type of mission and provide us their reflexion about team training, the type of relation with the country where the mission is conducted and the type of right team. PMID- 15276259 TI - [Interplast in India. Review of 14 years]. AB - Interplast teams conducted two-week (or less) camps in India over a 14 year period. The majority of the teams were mixed and consisted of German and Dutch plastic surgeons and nurses. In five different villages and cities, 1015 patients were operated: 41% concerned cleft lip and palate and 32% post-burn contractures. The work is rewarding and is considered by some the best holiday imaginable. The teams intend to continue their project in the future. PMID- 15276260 TI - [Noma. Proposal for a surgical treatment]. AB - The authors present their experience of surgical treatment of noma in situation of surgical camp. The strategy is focused on two objectives: treatment of tissue loss itself and treatment of the trismus. After having presented the means at disposal, going from local flaps, quickly exceeded, to distant flaps, they decided upon the indications by taking in account the NOILTULP classification. Thus, the authors present their experience of fascia temporalis skin grafted for oral lining in combination with the submental or Backamjian flap for external coverage. The treatment of the trismus is less codified requiring osteotomies in stages 3 and 4. They finally insist first on the prevention of this disease, very accessible to antibiotics at the initial stage of stomatitis and second on simple means very efficient to prevent the trismus which impairs heavily the functional outcome of the reconstructions. PMID- 15276261 TI - [GESNOMA (Geneva Study Group on Noma): state-of-the-art medical research for humanitarian purposes]. AB - Noma is a devastating gangrenous disease that leads to severe tissue destructions in the face. It is seen almost exclusively in children living in less developed countries. The exact prevalence of the disease is unknown and the cause remains unknown too. Risk factors are: malnutrition, a compromised immune system, poor oral hygiene and a lesion of the gingival mucosal barrier, as well as an unidentified bacterial factor. Herpes viruses might also contribute. Studies of the buccal flora in acute phases of noma and comparison with control children do not exist. Our study takes place in Niger. For each child (cases and controls) we take samples of gingival fluid, saliva, blood and mouth mucosal swabs. The samples are analysed in Geneva in different laboratories. We control the serologies for Herpes viruses and measles. We also perform a nutritional assessment and the mucosal swabs are cultivated for the presence of viruses. The gingival flora is investigated by microarrays. These microarrays are instrumental to test for the presence of thousands of different bacteria in each clinical sample. This method allows a qualitative and quantitative description of the oral flora in noma-children and control cases. This is the first large scale study on the etiology of noma which uses new technical approaches for humanitarian purposes. PMID- 15276262 TI - [Plastic surgery under challenging conditions: the concept of the five F's]. AB - This work was done essentially for the benefit of the surgeon who wishes to practice plastic surgery under challenging conditions. The questions he asks himself are most often aimed at what type of pathology he will encounter during his mission, on the operating techniques he will have to adopt and the surgical principles he will have to observe in this type of environment. This presentation tries to come up with an answer, among others, by offering a concept which could be adapted to surgical techniques under challenging conditions. The study, retrospectively, of 100 patients operated during a mission led us to suggest six techniques of reconstructive surgery. We have observed the conditions in which we can apply these different techniques and we have paid a lot of attention and reflection on the environment of our therapy. Before going on a plastic surgery mission we strongly advise the surgeon to bear in mind, in his choice of a surgical technique, the following concepts: the ultimate goal to be reached, the feasibility of the surgical practice, the safety of the technique, the familiarity of the surgical procedure and the easiness to teach of the method. PMID- 15276263 TI - [Humanitarian plastic surgery in question]. AB - Humanitarian plastic surgery has become very fashionable and more and more surgeons are attracted by this type of commitment. The authors remind here of the necessary conditions and limitations of these actions. The communicative action according to J. Habermas, which means a true partnership with the local health care specialists should be the only valid engagement. PMID- 15276264 TI - [Humanitarian surgical mission. Coming home]. AB - Every mission questions our motivation, our skills, and our efficiency through the discovery of a disconcerting world. Three points should be considered to keep a lucid commitment: to evaluate what we have done, which requires modesty; to testify to the courage, the beauty, the violence we have seen; to seek a meaning to the mission beyond the surgical frame: questioning our selves; isn't there a mission going farther the mission? PMID- 15276265 TI - [Can craniofacial surgery be exported?]. PMID- 15276266 TI - [Humanitarian action...and then? An interview of Jean-Christophe Rufin, ex vice president of Medecins sans Frontieres, president of Action contre la faim, Goncourt prize 2001]. AB - First dedicated to emergency situations, then more involved in long term duration development, humanitarian action is concerned by a crisis which appears absolutely necessary after 30 years of growing up. Many factors may contribute to that situation: structural professionalisation, official financial pressures, political influences, more ideology for less ideal behaviour, competition with rising of new ideas like alter mondialisation etc. It seems interesting in such a situation to get some advice about humanitarian future from a personality who has recognised responsibilities in both action and thoughts consideration. PMID- 15276268 TI - Levels of PAHs in soil and vegetation samples from Tarragona County, Spain. AB - The levels of 16 polycyclic aromatic hydrocarbons (PAHs) were determined in 24 soil and 12 wild chard samples collected in Tarragona County (Catalonia, Spain), an area with an important number of chemical and petrochemical industries. Samples were also collected in urban/residential zones and in presumably unpolluted sites (control samples). In soils, the sum of the 16 PAHs ranged between 1002 and 112 ng/g (dry weight) for samples collected near chemical industries and unpolluted sites, respectively. With the exception of acenaphthylene, acenaphthene, anthracene and benzo[k]fluoranthene, no significant differences in the levels of the remaining PAHs were found among the different zones of sample collection. In chard samples, the highest value (sum of 16 PAHs) was observed in the residential area, followed by the industrial and the unpolluted zones, with concentrations of 179, 58 and 28 ng/g (dry weight), respectively. In general terms, the current PAH concentrations in soil and vegetation are lower than the levels reported in a number of investigations from different regions and countries. They are also below the maximum PAH concentrations allowed by the Catalan legislation for different uses of soil. PMID- 15276269 TI - Determining the background levels of bismuth in tissues of wild game birds: a first step in addressing the environmental consequences of using bismuth shotshells. AB - Bismuth shotshells have been approved as a "nontoxic" alternative to lead in North America. Approval was based on a limited number of studies; even background levels of bismuth in wildfowl were unknown. We report on the concentration of bismuth (and lead) in muscle and liver tissues of wildfowl (Anas platyrhynchos, Anas acuta, Anas crecca, Branta canadensis, Chen caerulescens) harvested with lead shotshell. Average liver-bismuth levels detected in the present study (e.g., teal, 0.05 microg/g dw; mallard, 0.09 microg/g dw) suggest analytical error in other studies examining the effects of bismuth in birds. Significant positive relationships between bismuth- and lead-tissue levels for muscle when all species were combined (and for B. canadensis and C. caerulescens separately) can be explained by noting that bismuth is a contaminant of lead. Thus, more research is recommended to confirm the appropriateness of bismuth as a "nontoxic" shot alternative. PMID- 15276270 TI - Phytoextraction of heavy metals by canola (Brassica napus) and radish (Raphanus sativus) grown on multicontaminated soil. AB - Phytoextraction can provide an effective in situ technique for removing heavy metals from polluted soils. The experiment reported in this paper was undertaken to study the basic potential of phytoextraction of Brassica napus (canola) and Raphanus sativus (radish) grown on a multi-metal contaminated soil in the framework of a pot-experiment. Chlorophyll contents and gas exchanges were measured during the experiment; the heavy metal phytoextraction efficiency of canola and radish were also determined and the phytoextraction coefficient for each metal calculated. Data indicated that both species are moderately tolerant to heavy metals and that radish is more so than canola. These species showed relatively low phytoremediation potential of multicontaminated soils. They could possibly be used with success in marginally polluted soils where their growth would not be impaired and the extraction of heavy metals could be maintained at satisfying levels. PMID- 15276271 TI - Accumulation of lead, zinc, copper and cadmium by 12 wetland plant species thriving in metal-contaminated sites in China. AB - The concentrations of lead, zinc, copper and cadmium accumulated by 12 emergent rooted wetland plant species including different populations of Leersia hexandra, Juncus effusus and Equisetum ramosisti were investigated in field conditions of China. The results showed that metal accumulation by wetland plants differed among species, populations and tissues. Populations grown in substrata with elevated metals contained significantly higher metals in plants. Metals accumulated by wetland plants were mostly distributed in root tissues, suggesting that an exclusion strategy for metal tolerance widely exists in them. That some species/populations could accumulate relatively high metal concentrations (far above the toxic concentration to plants) in their shoots indicates that internal detoxification metal tolerance mechanism(s) are also included. The factors affecting metal accumulation by wetland plants include metal concentrations, pH, and nutrient status in substrata. Mostly concentrations of Pb and Cu in both aboveground and underground tissues of the plants were significantly positively related to their total and/or DTPA-extractable fractions in substrata while negatively to soil N and P, respectively. The potential use of these wetland plants in phytoremediation is also discussed. PMID- 15276272 TI - Factors influencing liver PCB concentrations in sparrowhawks (Accipiter nisus), kestrels (Falco tinnunculus) and herons (Ardea cinerea) in Britain. AB - Large scale temporal and spatial changes in the exposure of terrestrial vertebrates to PCBs have been monitored in the UK by measuring liver residues in sparrowhawks (Accipiter nisus), kestrels (Falco tinnunculus) and grey herons (Ardea cinerea) from throughout the country. Residues in the three species are typically characterised by large intra- and inter-specific variation. Data for 306 sparrowhawks, 186 kestrels and 47 herons collected between 1992 and 1997 as part of a national Predatory Bird Monitoring Scheme were examined to determine how much of this variation was explained by body condition, age and sex, rather than other factors. In all three species, body condition was the single most important factor and accounted for up to 49% of the variation in PCB liver residues; starved birds had the highest liver concentrations. Age and sex were also significant but of lesser importance. Adult sparrowhawks and kestrels had liver PCB residues that were 2 to 10-fold higher than in first-year birds. Sex did not affect residue magnitude in a consistent manner. PCB concentrations in the liver were higher in males than females in both first-year and adult kestrels and in first-year sparrowhawks, but adult female sparrowhawks had similar PCB residues to adult males. Liver residues also varied seasonally. PCB concentrations in first-year sparrowhawks increased during the first year following fledging and a similar pattern was detected in adult female sparrowhawks following egg laying. When these physiological factors were taken into account, it was evident that while kestrels with high fat scores had significantly lower PCB concentrations than either sparrowhawks or herons, liver residues were similar in all three species when birds were in a starved condition. Overall during 1992-1997, the combined influence of body condition, age and sex explained more of the variation in liver PCB concentrations than species differences or other factors, such as geographical variation in exposure. PMID- 15276273 TI - Oribatid mite communities and metal bioaccumulation in oribatid species (Acari, Oribatida) along the heavy metal gradient in forest ecosystems. AB - The responses of oribatid communities to heavy metal contamination were studied. Concentration of cadmium, copper and zinc in nine oribatid species along a gradient of heavy metal pollution was measured. Oribatid mites were sampled seasonally during two years in five forests located at different distances from the zinc smelter in the Olkusz District, southern Poland. The most numerous and diverse oribatid communities were found in the forest with moderate concentrations of heavy metals. Analysis by atomic absorption spectrophotometry revealed large differences in metal body burdens among species. All studied oribatid species appeared to be accumulators of copper with Oppiella nova, Nothrus silvestris and Adoristes ovatus characterized by the highest bioaccumulation factors. Most species poorly accumulate cadmium and zinc. The accumulation of heavy metals in the body of oribatids was not strictly determined by their body size or the trophic level at which they operate. PMID- 15276274 TI - Microbial acidification and pH effects on trace element release from sewage sludge. AB - Leaching of sludge-borne trace elements has been observed in experimental and field studies. The role of microbial processes in the mobilization of trace elements from wastewater sludge is poorly defined. Our objectives were to determine trace element mobilization from sludge subjected to treatments representing microbial acidification, direct chemical acidification and no acidification, and to determine the readsorption potential of mobilized elements using calcareous sand. Triplicate columns (10-cm diameter) for incubation and leaching of sludge had a top layer of digested dewatered sludge (either untreated, acidified with H2SO4, or limed with CaCO3; all mixed with glass beads to prevent ponding) and a lower glass bead support bed. Glass beads in the sludge layer, support layer or both were replaced by calcareous sand in four treatments used for testing the readsorption potential of mobilized elements. Eight sequential 8-day incubation and leaching cycles were operated, each consisting of 7.6 d of incubation at 28 degrees C followed by 8 h of leaching with synthetic acid rain applied at 0.25 cm/h. Leachates were analyzed for trace elements, nitrate and pH, and sludge layer microbial respiration was measured. The largest trace element, nitrate and S losses occurred in treatments with the greatest pH depression and greatest microbial respiration rates. Cumulative leaching losses from both microbial acidification and direct acidification treatments were > 90% of Zn and 64-80% of Cu and Ni. Preventing acidification with sludge layer lime or sand restricted leaching for all trace elements except Mo. Results suggested that the primary microbial role in the rapid leaching of trace elements was acidification, with results from direct acidification being nearly identical to microbial acidification. Microbial activity in the presence of materials that prevented acidification mobilized far lower concentrations of trace elements, with the exception of Mo. Trace elements mobilized by acidification were readsorbed by calcareous sand when present. PMID- 15276275 TI - Fluxes of trichloroacetic acid through a conifer forest canopy. AB - Controlled-dosing experiments with conifer seedlings have demonstrated an above ground route of uptake for trichloroacetic acid (TCA) from aqueous solution into the canopy, in addition to uptake from the soil. The aim of this work was to investigate the loss of TCA to the canopy in a mature conifer forest exposed only to environmental concentrations of TCA by analysing above- and below-canopy fluxes of TCA and within-canopy instantaneous reservoir of TCA. Concentrations and fluxes of TCA were quantified for one year in dry deposition, rainwater, cloudwater, throughfall, stemflow and litterfall in a 37-year-old Sitka spruce and larch plantation in SW Scotland. Above-canopy TCA deposition was dominated by rainfall (86%), compared with cloudwater (13%) and dry deposition (1%). On average only 66% of the TCA deposition passed through the canopy in throughfall and stemflow (95% and 5%, respectively), compared with 47% of the wet precipitation depth. Consequently, throughfall concentration of TCA was, on average, approximately 1.4 x rainwater concentration. There was no significant difference in below-canopy fluxes between Sitka spruce and larch, or at a forest edge site. Annual TCA deposited from the canopy in litterfall was only approximately 1-2% of above-canopy deposition. On average, approximately 800 microg m(-2) of deposited TCA was lost to the canopy per year, compared with estimates of above-ground TCA storage of approximately 400 and approximately 300 microg m(-2) for Sitka spruce and larch, respectively. Taking into account likely uncertainties in these values ( approximately +/- 50%), these data yield an estimate for the half-life of within-canopy elimination of TCA in the range 50 200 days, assuming steady-state conditions and that all TCA lost to the canopy is transferred into the canopy material, rather than degraded externally. The observations provide strong indication that an above-ground route is important for uptake of TCA specifically of atmospheric origin into mature forest canopies, as has been shown for seedlings (in addition to uptake from soil via transpiration), and that annualized within-canopy elimination is similar to that in controlled-dosing experiments. PMID- 15276276 TI - Growth of Phragmites australis (Cav.) Trin ex. Steudel in mine water treatment wetlands: effects of metal and nutrient uptake. AB - The abandoned mine of Shilbottle Colliery, Northumberland, UK is an example of acidic spoil heap discharge that contains elevated levels of many metals. Aerobic wetlands planted with the common reed, Phragmites australis, were constructed at the site to treat surface runoff from the spoil heap. The presence of a perched water table within the spoil heap resulted in the lower wetlands receiving acidic metal contaminated water from within the spoil heap while the upper wetland receives alkaline, uncontaminated surface runoff from the revegetated spoil. This unique situation enabled the comparison of metal uptake and growth of plants used in treatment schemes in two cognate wetlands. Results indicated a significant difference in plant growth between the two wetlands in terms of shoot height and seed production. Analyses of metal and nutrient concentrations within plant tissues provided the basis for three hypotheses to explain these differences: (i) the toxic effects of high levels of metals in shoot tissues, (ii) the inhibition of Ca (an essential nutrient) uptake by the presence of metals and H+ ions, and (iii) low concentrations of bioavailable nitrogen sources resulting in nitrogen deficiency. This has important implications for the engineering of constructed wetlands in terms of the potential success of plant establishment and vegetation development. PMID- 15276277 TI - Shade and flow effects on ammonia retention in macrophyte-rich streams: implications for water quality. AB - Controlled releases of NH4-N and conservative tracers (Br- and Cl-) to five reaches of four streams with contrasting macrophyte communities have shown differing retentions, largely as a result of the way plants interact with stream flow and velocity. First-order constants (k) were 1.0-4.8 d(-1) and retention of NH4-N was 6-71% of amounts added to each reach. Distance travelled before a 50% reduction in concentration was achieved were 40-450 m in three streams under low flow conditions, and 2400-3800 m at higher flows. Retention (%) of NH4-N can be approximated by a simple function of travel time and k, highlighting the importance of the relationship between macrophytes and stream velocity on nutrient processing. This finding has significant management implications, particularly with respect to restoration of riparian shade. Small streams with predominantly marginal emergent plants are likely to have improved retention of NH4-N as a result of shading or other means of reducing plant biomass. Streams dominated by submerged macrophytes will have impaired NH4-N retention if plant biomass is reduced because of reduced contact times between NH4-N molecules and reactive sites. In these conditions water resource managers should utilise riparian shading in concert with unshaded vegetated reaches to achieve a balance between enhanced in-stream habitat and nutrient processing capacity. PMID- 15276278 TI - Characterization of contaminants in snapping turtles (Chelydra serpentina) from Canadian Lake Erie Areas of Concern: St. Clair River, Detroit River, and Wheatley Harbour. AB - PCBs, organochlorine pesticides and dioxins/furans in snapping turtle eggs and plasma (Chelydra serpentina) were evaluated at three Areas of Concern (AOCs) on Lake Erie and its connecting channels (St. Clair River, Detroit River, and Wheatley Harbour), as well as two inland reference sites (Algonquin Provincial Park and Tiny Marsh) in 2001-2002. Eggs from the Detroit River and Wheatley Harbour AOCs had the highest levels of p,p'-DDE (24.4 and 57.9 ng/g) and sum PCBs (928.6 and 491.0 ng/g) wet weight, respectively. Contaminant levels in eggs from St. Clair River AOC were generally higher than those from Algonquin Park, but similar to those from Tiny Marsh. Dioxins appeared highest from the Detroit River. The PCB congener pattern in eggs suggested that turtles from the Detroit River and Wheatley Harbour AOCs were exposed to Aroclor 1260. TEQs of sum PCBs in eggs from all AOCs and p,p'-DDE levels in eggs from the Wheatley Harbour and the Detroit River AOCs exceeded the Canadian Environmental Quality Guidelines. Furthermore, sum PCBs in eggs from Detroit River and Wheatley Harbour exceeded partial restriction guidelines for consumption. Although estimated PCB body burdens in muscle tissue of females were well below consumption guidelines, estimated residues in liver and adipose were above guidelines for most sites. PMID- 15276279 TI - Arsenic hyperaccumulation by Pteris vittata from arsenic contaminated soils and the effect of liming and phosphate fertilisation. AB - Pot experiments were carried out to investigate the potential of phytoremediation with the arsenic hyperaccumulator Pteris vittata in a range of soils contaminated with As and other heavy metals, and the influence of phosphate and lime additions on As hyperaccumulation by P. vittata. The fern was grown in 5 soils collected from Cornwall (England) containing 67-4550 mg As kg(-1) and different levels of metals. All soils showed a similar distribution pattern of As in different fractions in a sequential extraction, with more than 60% of the total As being associated with the fraction thought to represent amorphous and poorly crystalline hydrous oxides of Fe and Al. The concentration of As in the fronds ranged from 84 to 3600 mg kg(-1), with 0.9-3.1% of the total soil As being taken up by P. vittata. In one soil which contained 5500 mg Cu kg(-1) and 1242 mg Zn kg(-1), P. vittata suffered from phytotoxicity and accumulated little As (0.002% of total). In a separate experiment, neither phosphate addition (50mg P kg(-1) soil) nor liming (4.6 g CaCO3 kg(-1) soil) was found to affect the As concentration in the fronds of P. vittata, even though phosphate addition increased the As concentration in the soil pore water. Between 4 and 7% of the total soil As was taken up by P. vittata in 4 cuttings in this experiment. The results indicate that P. vittata can hyperaccumulate As from naturally contaminated soils, but may be suitable for phytoremediation only in the moderately contaminated soils. PMID- 15276280 TI - Perchlorate in water, soil, vegetation, and rodents collected from the Las Vegas Wash, Nevada, USA. AB - Water, soil, vegetation, and rodents were collected from three areas along the Las Vegas Wash, a watershed heavily contaminated with perchlorate. Perchlorate was detected at elevated concentrations in water, soil, and vegetation, but was not frequently detected in rodent liver or kidney tissues. Broadleaf weeds contained the highest concentrations of perchlorate among all plant types examined. Perchlorate in rodent tissues and vegetation was correlated with perchlorate concentrations in soil as expected, however rodent residues were not highly correlated with plant perchlorate concentrations. This indicates that soil may be a greater source, or a more constant source of perchlorate exposure in rodents than vegetation. PMID- 15276281 TI - Cumulative impact of 40 years of industrial sulfur emissions on a forest soil in west-central Alberta (Canada). AB - The impact of 40 years of sulfur (S) emissions from a sour gas processing plant in Alberta (Canada) on soil development, soil S pools, soil acidification, and stand nutrition at a pine (Pinus contorta x Pinus banksiana) ecosystem was assessed by comparing ecologically analogous areas subjected to different S deposition levels. Sulfur isotope ratios showed that most deposited S was derived from the sour gas processing plant. The soil subjected to the highest S deposition contained 25.9 kmol S ha(-1) (uppermost 60 cm) compared to 12.5 kmol S ha(-1) or less at the analogues receiving low S deposition. The increase in soil S pools was caused by accumulation of organic S in the forest floor and accumulation of inorganic sulfate in the mineral soil. High S inputs resulted in topsoil acidification, depletion of exchangeable soil Ca2+ and Mg2+ pools by 50%, podzolization, and deterioration of N nutrition of the pine trees. PMID- 15276282 TI - White poplar (Populus alba) as a biomonitor of trace elements in contaminated riparian forests. AB - Trees can be used to monitor the level of pollution of trace elements in the soil and atmosphere. In this paper, we surveyed the content of eight trace elements (As, Cd, Cu, Fe, Mn, Ni, Pb and Zn) in leaves and stems of white poplar (Populus alba) trees. We selected 25 trees in the riparian forest of the Guadiamar River (S. Spain), one year after this area was contaminated by a mine spill, and 10 trees in non-affected sites. The spill-affected soils had significantly higher levels of available cadmium (mean of 1.25 mg kg(-1)), zinc (117 mg kg(-1)), lead (63.3 mg kg(-1)), copper (58.0 mg kg(-1)) and arsenic (1.70 mg kg(-1)), than non affected sites. The concentration of trace element in poplar leaves was positively and significantly correlated with the soil availability for cadmium and zinc, and to a lesser extent for arsenic (log-log relationship). Thus, poplar leaves could be used as biomonitors for soil pollution of Cd and Zn, and moderately for As. PMID- 15276283 TI - Over one hundred years of trace metal fluxes in the sediments of the Pearl River Estuary, South China. AB - The rapid economic development in the Pearl River Delta (PRD) region in South China in the last three decades has had a significant impact on the local environment. Estuarine sediment is a major sink for contaminants and nutrients in the surrounding ecosystem. The accumulation of trace metals in sediments may cause serious environmental problems in the aquatic system. Thirty sediment cores were collected in the Pearl River Estuary (PRE) in 2000 for a study on trace metal pollution in this region. Heavy metal concentrations and Pb isotopic compositions in the four 210Pb-dated sediment cores were determined to assess the fluxes in metal deposits over the last one hundred years. The concentrations of Cu, Pb and Zn in the surface sediment layers were generally elevated when compared with the sub-surface layers. There has been a significant increase in inputs of Cu, Pb and Zn in the PRE since the 1970s. The results also showed that different sampling locations in the estuary received slightly different types of inputs. Pb isotopic composition data indicated that the increased Pb in the recent sediments was of anthropogenic origin. The results of trace metal influxes showed that about 30% of total Pb and 15% of total Zn in the sediments in the 1990s were from anthropogenic sources. The combination of trace metal analysis, Pb isotopic composition and 210Pb dating in an estuary can provide vital information on the long-term accumulation of metals in sediments. PMID- 15276284 TI - Spatial distribution of atmospheric PAHs and PCNs along a north-south Atlantic transect. AB - Ship-board air samples collected between The Netherlands and South Africa in January-February 2001 were analysed for polycyclic aromatic hydrocarbons (PAHs) and polychlorinated naphthalenes (PCNs). The highest PAH concentrations occurred in the European samples, and in samples close to West Africa and South Africa. Consistently low PAH concentrations were measured in the southern hemisphere open ocean samples (190-680 pg/m3). The highest PCN concentrations occurred in the European samples, but high values were also detected off the West African coast, and in the sample taken closest to South Africa. Data are presented for diurnal cycles taken in the remote South Atlantic. The day:night ratios of phenanthrene, 1-methylphenanthrene and fluoranthene were typically approximately 1.5-2.5:1. The mechanism(s) causing this observation is/are not understood at present, but dynamic environmental process(es) is/are implicated. PMID- 15276285 TI - [Infection and MALT lymphoma. New developments?]. PMID- 15276286 TI - [Detection of Alzheimer's disease in general medicine: preliminary results of a Sentinelles general practitioner's network survey]. AB - CONTEXT: The availability of cholinesterase inhibitors has reinforced the need for early detection of Alzheimer's disease in which the general practitioner plays a key role. This study seeks to appraise the diagnostic procedures of Alzheimer's disease in general medicine. METHOD: A postal survey of 1176 general practitioners belonging to the French Sentinelles network. RESULTS: Response rate: 43%. The surveyed doctors have seen 1.5 new cases in 2002, and were in charge of the follow-up of four patients. Reasons leading to consultation were: memory impairment (84%), memory impairment together with decline in daily functioning and disorientation (42%). Seventy-six per cent of the Sentinelles used the MMSE, 91% referred the patient to a specialist. Fifty-four per cent announced the diagnosis to the patient; 94% to the family. Twenty-six per cent of the surveyed doctors systematically used the DSM-IV criteria and 77% considered early diagnosis valuable. CONCLUSION: The results of this survey show that Alzheimer's disease is still under-diagnosed in general medicine in spite of the general practitioner's favourable opinion as to early diagnosis of the disease. Compared to his European colleagues, French general practitioner's attitude is characterized by frequent referral to a specialist service, follow-up of diagnostic criteria and frequency of disclosure of diagnosis to the family. PMID- 15276287 TI - [Update of pernicious anemia. A retrospective study of 49 cases]. AB - PURPOSE: The aim of this study was to describe the present clinical characteristics of the pernicious anemia (PA). METHOD: It is a retrospective (1996-2002) multicenter (five departments of internal medicine) study of 49 patients presenting an established cobalamin deficiency related to PA. RESULTS: The median age of the patients was 74 years (25-93), the female/male ratio 2:9. Several autoimmune disorders were noted in 35% of the patients. Various clinical manifestations, mainly neurological, cutaneous and thrombotic, were found in 65.4% of the patients, at least one hematological abnormalities in 100%. Average serum vitamin B12 and homocystein levels were with 73 pg/ml (20-1960) and 42.9 micromol/l (7, 8-124). Anti-intrinsic factor or anti-parietal gastric cells antibodies were found in 87.5% and 62% of the patients (at least one antibody, in 96%) abnormal Schilling's test results in 86%. All the followed patients were successful treated with intramuscular (n = 27) or oral crystalline cyanocobalamin (n = 5). CONCLUSIONS: PA was associated with several autoimmune disorders; PA may be responsible of various clinical manifestations or biological abnormalities; and oral crystalline cyanocobalamin treatment may be successful. PMID- 15276288 TI - [Thymus function and autoimmunity]. AB - PURPOSE: Thymus is the site of T-cell development and is essential for the induction of self-tolerance, by deletion of autoreactive T lymphocytes (negative selection) and by generation of regulatory T cells. Defect of the selection mechanism of both types of lymphocytes lead to autoimmune diseases. CURRENT KNOWLEDGE AND KEY POINTS: Elimination of potentially self-reactive T cells in the thymus requires the intrathymic expression of ubiquitous and "tissue-specific" antigens. Some thymic antigen expressions are dependent on AIRE expression. Mutations in the AIRE gene that are associated with the absence of autoantigen expression in the thymus, defects in the peptide presentation or in apoptosis can allow autoreactive T cells to escape negative selection, and are associated with autoimmune diseases. Recent data are now available concerning the thymic selection of autoreactive regulatory T cells. The Foxp3 gene was recently shown to be predominantly expressed in regulatory T cells and could be a more specific marker of regulatory T cells than phenotypic markers. FUTURE PROSPECTS AND PROJECTS: Animal models show that regulatory T cells injection or intrathymic inoculation of antigen lead to immunological tolerance in autoimmunity and transplantation. These novel strategies could be used in human. PMID- 15276289 TI - [MALT gastric lymphomas]. AB - INTRODUCTION: The stomach is the most common site involved in primary gastrointestinal lymphoma. Gastric lymphoma originates from the mucosa-associated lymphoid tissue so called MALT. It comprises a group of distinctive clinicopathological entities which are important to consider for clinical management. CURRENT KNOWLEDGE AND KEY POINTS: In recent years, new diagnostic tools and new treatment strategies have improved the overall prognosis. One of the most exciting recent discoveries is the hypothesis that an infection by a bacterium, Helicobacter pylori has a decisive role in gastric lymphoma. FUTURE PROSPECTS AND PROJECTS: Recent advances, essentially due to molecular biology and cytogenetic studies may emerge with the understanding of pathogenesis and new prognostic factors of these different types of gastric lymphomas. It is the aim of our oncoming studies together with the evaluation of the new therapeutic options such as radiotherapy and monoclonal antibodies in prospective studies. PMID- 15276290 TI - [Appropriate drug prescribing in the elderly]. AB - PURPOSE: The frequency of chronic illness and the expenditure of medications increase with the older age. If drug-prescribing is very often beneficial for the patients, elderly subjects are particularly exposed to the side-effects of medications, and to their consequences. Although the age in itself do not generally forbid a medication, it can modify some of the objectives and the modality of the treatment. CURRENT KNOWLEDGE AND KEY POINTS: Four main factors explain the growing frequency of the iatrogenic pathology in the old age: polymedication, pharmacological modifications that occur with the ageing process, lack of coordination between different prescribers, and physical and psycho social deficiencies which are the keys for identifying the "frail elderlies". FUTURE PROSPECTS AND PROJECTS: In regard to the frequent polypathological conditions of the oldest patients, one does not have to consider a sum of sicknesses, but a global situation, and one needs to replace a condition in a general context, and to establish priorities in the goals of the treatment. Appropriate drug-prescribing also needs to take account of the latest medical recommendations concerning numbers of medications, and a more frequent involving of elderly subjects in clinical trials. PMID- 15276291 TI - [Pulmonary plasmacytoma: about two new cases and review of the literature]. AB - INTRODUCTION: Extramedullary plasmacytoma is an uncommon plasma cell malignancy mainly located to the upper aerodigestive tract. Primary pulmonary plasmacytoma is extremely rare. EXEGESIS: We report two new cases of primary pulmonary plasmacytoma and then proceed to a review of the literature concerning 35 similar cases previously described. CONCLUSION: Complete or partial responses were obtained in 24 cases (65%). Five patients (14%) have developed multiple myeloma within 3 years following plasmacytoma diagnostic. In spite of sustained responses with radiotherapy or chemotherapy, surgical resection while feasible remains the first therapeutic option. PMID- 15276292 TI - [Primary non-Hodgkin lymphoma of the liver in HIV infected patient: case-report]. AB - INTRODUCTION: Primary liver non-Hodgkin's lymphoma, are extremely rare in HIV infected patient. Most of them are diffuse large-cell lymphoma with B cell type. EXEGESIS: We report here the case of a 34-year-old HIV-infected patient, admitted for jaundice and fever since 15 days. Abdominal computerised tomography showed numerous hypodense lesions on all liver segments. The various biological, microbiological and morphological examinations (ultrasound, MRI with intravenous contrast agent specific for the liver) initially suggested a tumoral origin. The liver biopsy concluded to a large B-cell lymphoma. A chemotherapy (CHOP) with anti-CD20 monoclonal antibody (rituximab) was initiated without discontinuing antiretroviral therapy. CONCLUSION: This case-report does emphasize on the numerous presentations of primary liver lymphoma in HIV-Infected patient, and we illustrate the interest of MRI using a new intravenous contrast agent critical for differential diagnosis. PMID- 15276293 TI - [A patient with adrenal insufficiency and morphea]. PMID- 15276294 TI - [An uncommon case of hydroxyurea-induced fever]. PMID- 15276295 TI - [Juvenile idiopathic polyarthritis in monozygotic twins with congenital C4 deficiency]. PMID- 15276296 TI - [Possible confusion between Buerger disease and Hansen disease in tropical area: report of a case]. PMID- 15276297 TI - [Severe acute respiratory syndrome: one case of indirect transmission by Coronavirus]. PMID- 15276298 TI - Benzo[b]acronycine derivatives: a novel class of antitumor agents. AB - A hypothesis of bioactivation of the antitumor alkaloid acronycine by transformation of the 1,2-double bond into the corresponding epoxide in vivo and the suggestion that acronycine could interact with DNA, led to develop 1,2 dihydroxy-1,2-dihydrobenzo[b]acronycine diesters (1,2-dihydroxy-6-methoxy-3,3,14 trimethyl-1,2,3,14-tetrahydro-7H-benzo[b]pyrano[3,2-h]acridin-7-one diesters) as new anticancer drug candidates. Compared to acronycine these compounds were markedly more potent, both in terms of cytotoxicity and antitumor activity. The biological activity of these compounds was strongly related with their ability to give covalent adducts with purified as well as genomic DNA. Formation of those adducts involves alkylation of the exocyclic N-2 amino groups of guanines exposed in the minor groove of double helical DNA by the carbocation produced by the elimination of the acyloxy leaving group at position 1 of the drug. A transesterification process of the ester group from position 2 to position 1 accounted for the intense activity of cis-1-hydroxy-2-acyloxy-1,2 dihydrobenzo[b]acronycine derivatives. Cis-1,2-diacetoxy-1,2 dihydrobenzo[b]acronycine, which displays a particularly impressive broad antitumor spectrum, is currently developed by Servier Laboratories under the code S23906-1. PMID- 15276299 TI - Novel cephalosporin derivatives possessing a substituted cinnamoyl moiety at the 7 beta-position. Synthesis, structural characterization and antibacterial activity of 3-acetoxymethyl cephalosporin derivatives. AB - Twenty 3-acetoxymethyl cephalosporin derivatives, with various cinnamoyl (3 phenyl-2-propenoyl) substituted groups at the 7beta-position, were synthesized and evaluated for antibacterial activity in vitro. Some of these cephalosporin derivatives showed good selective activity against Gram-positive bacteria. Although substitution on the aromatic ring of cinnamoyl moiety generally reduced antimicrobial activity against Staphylococcus sp. and Enterococcus sp., a hydroxy group at the para position, and particularly ortho, para di-chloro substitution, improved the activity against methicillin resistant strains of Staphylococcus aureus (MRSA). Substitution on the double bond alpha position of the cinnamoyl moiety also affected the antimicrobial activity. A cyano group attached to this position increased activity against both negative coagulase Staphylococcus and Enterococcus sp. and extended the antibacterial spectrum towards Gram-negative bacteria. PMID- 15276300 TI - A comparative study of the hydrolysis pathways of substituted aryl phosphoramidate versus aryl thiophosphoramidate derivatives of stavudine. AB - A comparative study of aryl phosphoramidate and aryl thiophosphoramidate derivatives of 2',3'-didehydro-2',3'-dideoxythymidine (d4T) was performed. The study focused on the nature of the substituents and the influence of a thiophosphoramidate in the structure of these derivatives. The rate of alkaline hydrolysis of these two types of d4T derivatives indicated that replacement of oxygen with sulfur decreases the rate of hydrolysis by twofold. Additionally, the activation energy (E(a)) for the sulfur analogs is comparatively higher than that of the oxygen analogs. Notably, an intermediate was formed in the hydrolysis reaction of the sulfur analogs of d4T that was absent in the case of the oxygen analog, and the tentative structure of the intermediate was proposed based on LC/mass spectroscopy data. Using both HPLC and (31)P-NMR techniques, we identified the hydrolysis product of the phosphoramidate derivatives and were able to show in in vitro studies that porcine liver esterase can hydrolyze the methyl ester portion of the phosphoramidate derivatives. Aryl phosphoramidate derivatives of d4T were 1000-fold more active than the corresponding aryl thiophosphoramidate derivatives, indicating that the energy of activation of hydrolysis of these phosphoramidate derivatives plays a significant role in their biological potency. PMID- 15276301 TI - Synthesis and antileishmanial activity of (1,3-benzothiazol-2-yl) amino-9-(10H) acridinone derivatives. AB - (1,3-Benzothiazol-2-yl) amino-9-(10H)-acridinone derivatives were synthesized via a procedure based on the Ullman reaction and were assessed for their in vitro antileishmanial and anti-HIV activities. Two derivatives, 4-(6-nitro-benzothiazol 2-ylamino)-10H-acridin-9-one and 1-(6-amino-benzothiazol-2-ylamino)-10H-acridin-9 one, revealed a selective antileishmanial activity, mainly due to amastigote specific toxicity. Results suggested that:the addition of a benzothiazole group on a parent amino-9-(10H)-acridinone ring could enhance antileishmanial abilities, the presence of a 6-amino-benzothiazole group on position 2 amino chain or a 6-nitro-benzothiazole group on position 4 amino chain was essential for specific anti-amastigote properties. PMID- 15276302 TI - Studies on the synthesis and characterization of four trans planaramineplatinum(II) complexes of the form trans-PtL(NH3)CL2 where L = 2 hydroxypyridine, 3-hydroxypyridine, imidazole, and imidazo(1,2-alpha)pyridine. AB - Four trans-planaramineplatinum(II) complexes code named YH9, YH10, YH11 and YH12, each of the form trans-PtL(NH(3))Cl(2) where L = 2-hydroxypyridine and 3 hydroxypyridine, imidazole, and imidazo(1,2-alpha)pyridine for YH9, YH10, YH11 and YH12, respectively. All of the compounds have significant anticancer activity against human cancer cell lines. YH12 is found to be significantly more active than cisplatin against cisplatin-resistant ovary cell line A2780(cisR). PMID- 15276303 TI - Synthesis and antibacterial activity of 2-(4-substituted phenyl)-3(2H) isothiazolones. AB - Several new and known 2-(4-substituted phenyl)-3(2H)-isothiazolone derivatives with or without chloro substituent at C-5 position were synthesized and their in vitro antibacterial activity against selected Gram-negative and Gram-positive bacteria were evaluated using agar dilution method. Most of compounds exhibited moderate to high activities against tested microorganisms, and in comparison with the reference drugs some compounds showed comparable or higher activities. In contrast to results of the previous studies, some 5-chloro derivatives showed lower or comparable activities against some tested microorganism, in comparison with analogues without C-5 substitution. In general, most of the compounds bearing electron withdrawing group at 4-position of the phenyl ring were more active against Gram-positive and most of those having piperazine derivatives were more active against Gram-negative bacteria. PMID- 15276304 TI - Synthesis, crystal structure and cytotoxicity of new oxaliplatin analogues indicating that improvement of anticancer activity is still possible. AB - Oxaliplatin, (trans-R,R-cyclohexane-1,2-diamine)oxalatoplatinum(II), has recently been approved for combination chemotherapy of metastatic colorectal cancer. Oxaliplatin is significantly more active than its trans-S,S isomer and the mixture of both enantiomers. New oxaliplatin analogues, (SP-4-3)-(4-methyl-trans cyclohexane-1,2-diamine)oxalatoplatinum(II) and (SP-4-3)-(4-ethyl-trans cyclohexane-1,2-diamine)oxalatoplatinum(II), have been synthesized, and their cytotoxicity has been tested in comparison to oxaliplatin, its corresponding trans-S,S isomer, and the mixture of both enantiomers. In comparison to oxaliplatin, even the trans-R,R/trans-S,S mixture of the 4-methyl and 4-ethyl substituted oxaliplatin analogues have shown an equivalent cytotoxicity in ovarian cancer cells (CH1) and superior antiproliferative properties in colon cancer cells (SW480) in the case of a predominantly equatorial position of the substituent at position 4 of the trans-cyclohexane-1,2-diamine ligand, whereas an axial substitution results in decreased cytotoxic potency. PMID- 15276305 TI - Potential prodrugs of non-steroidal anti-inflammatory agents for targeted drug delivery to the CNS. AB - Recently non-steroidal anti-inflammatory drugs (NSAIDs) have been proposed to prevent or to cure Alzheimer's disease. In this respect, we synthesized new potential prodrugs of several NSAIDs in order to increase their access to the brain. The carboxylic group of NSAIDs was attached to the 1,4-dihydro-1 methylpyridine-3-carboxylate moiety, which acts as a carrier, via an amino alcohol bridge, according to the chemical delivery approach developed by Bodor. The lipophilicity of potential prodrugs was evaluated both via traditional experimental parameters, such as partition coefficient and chromatographic R(m) value, and by predictive computational methods. From experimental parameters, all prodrugs were more lipophilic when compared to their corresponding parent compounds and consequently a better blood brain barrier (BBB) penetration is hypothesised. Prodrug lipophilicity was correlated with a calculated log P value according to Kowwin's method. The correlation between experimental Rm0 and calculated log P, determined by PLS analysis, was good for all compounds with the exception of compound 7i. In addition the BBB permeation profile of our synthesized compounds was predicted using the BBB VolSurf model and seven of the synthesized prodrugs resulted in good candidates for BBB penetration. PMID- 15276306 TI - 3-Chloro-2-methylphenyl-substituted semicarbazones: synthesis and anticonvulsant activity. AB - A series of 3-chloro-2-methylphenyl substituted semicarbazones (3-33) was synthesized and evaluated for anticonvulsant and CNS activities. After intraperitoneal injection to mice or rats, the semicarbazone derivatives were examined in the maximal electroshock seizure (MES), subcutaneous pentylenetetrazole (scPTZ), and subcutaneous strychnine (scSTY)-induced seizure and neurotoxicity screens. The aryl urea (1) and the semicarbazide (2) showed anticonvulsant activity in the MES and scPTZ screens with acute neurotoxicity, whereas the semicarbazone derivatives showed good anticonvulsant potency in the scSTY screen with moderate activity against MES and scPTZ screens. Compound 21 exhibited anticonvulsant potency against all the three screens with lesser neurotoxicity. Some titled compounds exhibited lesser CNS depression and neurotoxicity compared to phenytoin or carbamazepine as was evident from the CNS studies. PMID- 15276307 TI - Determinants of caesarean section in Egypt: evidence from the demographic and health survey. AB - This paper examines the impact of near birth complications and socio-demographic, healthcare and spatial characteristics of caesarean section in Egypt, using data on 4032 births from the 2000 Egypt Demographic and Health Survey. The hospital caesarean section rate was 22% in Egypt. Fever/vaginal discharge around delivery, birth weight, mother's age and education, birth order, residence and antenatal visits were important determinants of caesarean section. Variations by place of delivery were evident, although complications were more significant determinants of caesarean section in public settings and demographic characteristics were more important in private facilities. Unexpectedly, long labour and bleeding around delivery were not associated with caesarean section, particularly, in private hospitals. In view of the high and rising caesarean section rate in Egypt, monitoring the quality of maternity services in Egypt is imperative. An investigation of the forces sustaining the differential in determinants by place of delivery is needed. PMID- 15276308 TI - Opposition to unpopular research results: Finnish professional reactions to the WHI findings. AB - PURPOSE: Preventive hormone therapy (HT) has been popular. This article describes what happened, when unexpected results that speak against HT were published. METHODS: The article describes what happened in Finland after a large US randomised study. Women's Health Initiative (WHI) showed preventive HT to be harmful. The data used in the present study came from statistics, surveys, participatory observations, and written material. RESULTS: Use of HT increased in Finland in the 1990s, and long-term therapy became common. The opinions of Finnish physicians on HT were positive. After the WHI results were published in July 2002, they received wide publicity, and recommendations to reduce HT use were made in the United States. In Finland, the initial critical comments on HT were soon replaced by criticism of the WHI study in both the professional and the lay press. The criticism concerned the type of the drug used (not used in Finland), the population ("not healthy, too old"), and outcome measures; the results were said not to apply to Finland. The value of a trial in contrast to the non-experimental evidence was rarely stressed. Odd information supplied and the uniformity of the formulations used by gynaecologists and medical journalists suggested that these professionals had been trained by the drug firms preparing competing HT drugs. CONCLUSIONS: This case shows that when a well-done trial results in a negative assessment of a widely used drug, those benefiting from its continued use may aggressively resist, even despite weak evidence for their position. PMID- 15276309 TI - Estimation of the impact of providing outpatients with information about SARS infection control on their intention of outpatient visit. AB - To examine the effect of provision of information about the infection control in the specific infection disease treatment unit in a city hospital on the outpatient's intention of outpatient service use, respondents who underwent outpatient medical care at the hospital (N = 821) were asked whether or not they intended to continue the outpatient visit at the hospital if a severe acute respiratory syndrome (SARS) patient was admitted to the unit. Although 56% of respondents replied that they could continue to visit the department if a SARS patient was admitted to the unit in the hospital before they read the information, the proportion of those who intended to continue outpatient care significantly increased by 15% after they read it. The logistic regression analyses revealed that respondents who had frequently visited the outpatient department (P < 0.001), those who felt relieved by reading the information about the unit (P < 0.001), and those who did not worry about nosocomial SARS infection inside the hospital (P < 0.001) were significantly more likely to reply that they would continue outpatient visits. We estimated that admission of a SARS patient to the unit would result in a 20% decrease in the cumulative total number of outpatients in the hospital during a 180-day interval after admission of a SARS patient to the unit, and the cumulative total number of outpatients increased by 7% after they read the information. This study suggests that providing outpatients with appropriate information about SARS infection control in the hospital had a statistically significant and substantial impact on the outpatients' intention to continue outpatient visits at the hospital. PMID- 15276310 TI - Economic benefit from clinical practice guideline compliance in stroke patient management. AB - BACKGROUND AND PURPOSE: In a previous study we showed that compliance with evidence-based guidelines improves the health outcome of stroke patients in terms of both survival and residual disability. In this analysis, we shall investigate the impact of such guidelines on healthcare costs during the acute/sub-acute hospitalisation phase. METHOD: we considered the direct costs from the hospital's point of view, where funding is provided by the National Healthcare System. We did not consider production loss or intangible costs related to the decreased quality of life. Data was collected on both costs and guideline compliance prospectively, and the relationship between them was studied through a multivariate statistical model. RESULTS: Patients treated according to guidelines result in lower costs; on average they have a shorter length of stay in hospital (10.8 versus 12.9 days), leading to a significant difference in the consumption of hospital resources. On a level of statistical analysis, guideline compliance is a significant independent indicator of cost, together with the patient's initial disability and neurological deficit. PMID- 15276311 TI - Trading patients. Lessons from Scandinavia. AB - The next few decades will bring about more trade in services, among them health care. This paper describes a recent project on cross-border trading of patients initiated by the Norwegian parliament (The Patient Bridge). This health policy reform met some resistance among hospital physicians. However, patients were willing to participate if being properly informed and supported by local health care workers. The Patient Bridge turned out to be a relatively expensive project partly because of the transaction costs involved (transportation and escort) and partly because of high treatment costs. Excessive treatment costs were a result of insufficient cost-consciousness in the purchasing organization. The Patient Bridge revealed large price differentials not only between Norwegian and foreign hospitals, but also between hospitals abroad, even within the same country. This finding points to the possibilities of reaching mutual gains from trading patients across borders. PMID- 15276312 TI - China's evolving health care market: how doctors feel and what they think. AB - This paper reports on a questionnaire survey and 12 focus groups conducted among doctors in three provinces of China, namely Guangdong, Shanxi, and Sichuan. The survey (N = 720) and focus group participants were drawn from both rural and urban areas, as well as public and private sectors, in equal numbers The aim was to gauge how Chinese doctors feel about themselves and what they think of the Chinese health care system. We found low satisfaction levels with own income (8%), job (27%), skill (30%), and other important aspects of their professional life. The health care system received only 32% approval rating. Quality of care and patient safety issues were major concerns, especially in the growing but poorly regulated private sector. The public sector came under criticism for its high fees and bad service quality. The feedback point to the need for an appropriate regulatory framework to guide the development of China's evolving health care market. A revitalized medical profession that is fully engaged in the reform process could also significantly impact the success of ongoing health care reform efforts. PMID- 15276313 TI - Measuring financial protection in health in the United States. AB - One rationale for health insurance coverage is to provide financial protection against catastrophic health expenditures. This article defines a lack of financial protection as household spending on health care when: (1) out-of-pocket (OOP) health expenditures exceed 10% of family income; (2) out-of-pocket expenditures exceed an absolute level of 2000 US dollars per family member on an annual basis; and (3) combined out-of-pocket and prepaid health expenditures exceed 40% of family income. The article explores how the likelihood of households in the United States surpassing these thresholds varies by income level, extent of insurance coverage, and the number of chronic conditions. The results show clearly that there is a lack of financial protection for health services for a wide segment of the US population-particularly so for poor families and those with multiple chronic conditions. The results are placed in an international context. Similar studies in other countries would allow for more in depth comparisons of financial protection than are currently possible. PMID- 15276314 TI - Women, men and public health-how the choice of normative theory affects resource allocation. AB - Women live longer than men in almost all countries, but men are more privileged in terms of power, influence, resources and probably morbidity. This investigation aims at illustrating how the choice of normative framework affects judgements about the fairness in these sex differences, and about desired societal change. The selected theories are welfare economics, health sector extra welfarism, justice as fairness and feminist justice. By means of five Swedish proposals aiming at improving the population's health or "sex equity", facts and values are applied to resource allocation. Although we do not claim a specific ethical foundation, it seems to us that the feminist criterion has great potential in public health policy. The overall conclusion is that the normative framework must be explicitly discussed and stated in issues of women's and men's health. PMID- 15276315 TI - Implementation of the project 'Support and Consultation on Euthanasia in The Netherlands' (SCEN). AB - OBJECTIVE: In the project 'Support and Consultation on Euthanasia in The Netherlands' (SCEN), general practitioners (GPs) receive training in formal consultation and in giving expert advice to colleagues who have questions about euthanasia and physician-assisted suicide (EAS). This study describes the way in which this project was implemented in The Netherlands and how it was received by GPs. METHOD: In the period from April 2000 to December 2002, all GPs in the districts in which SCEN had been implemented received a written post-test questionnaire a year and a half after the start of the project. Registration forms were also filled in by the SCEN physicians and the GPs who contacted SCEN. The post-test questionnaire was returned by 60% of the GPs (n = 3614), and registration forms were returned by 86% (n = 3337) of the GPs who contacted SCEN. RESULTS: The gradual nation-wide implementation of SCEN was completed within the 4-year study period. Almost all GPs were familiar with the project (99%) and most GPs knew what they could contact SCEN for. Most GPs felt supported by the presence of SCEN, and most GPs also had a positive attitude towards consultation and SCEN. GPs who had received an explicit request for EAS, or had performed EAS, often consulted SCEN physicians (71 and 85%, respectively). Reasons for contacting SCEN were: independence (60%), expertise (58%), and accessibility/availability of a consultant (45%). Reasons for not contacting SCEN were: enough other possibilities for counselling and consultation (48%) only at the beginning of the decision-making process (36%), and sufficient knowledge about EAS (22%). Most GPs intended to use SCEN in the future (96%). CONCLUSION: The implementation of SCEN has been successful, according to the four steps for successful implementation: awareness, attitude, use and future use. In this respect, linking up with existing networks and good organisation may play an important role. Furthermore, GPs consider it important to have a facility like SCEN which they can contact concerning EAS. PMID- 15276316 TI - Gender and the utilisation of health services in the Ashanti Region, Ghana. AB - The survey seeks to structure a model for gender-based health services utilisation for the Ashanti Region of Ghana, and in addition, recommend intervention measures to ensure gender equity in the utilisation of health services. A sample size of 650 covered over 3108 houses, and the main research instruments were the questionnaire and formal interview. A multiple regression model is used for the analysis of the relationship between the complex independent variables and utilisation by gender. Results show that although females have a greater need for health services than males, they do not utilise health services as much. Secondly, whereas quality of service, health status, service cost and education have greater effect on male utilisation than females, distance and income have higher impact on female utilisation. It is recommended that, to ensure equity in health care utilisation, females be empowered through increased access to formal education and sustainable income opportunities. The introduction of a national health insurance scheme is also recommended to ensure adequate access by both sexes. PMID- 15276317 TI - A study of malaria care provider choice in Ghana. AB - Improved understanding of the factors that influence malaria care seeking behaviour is necessary in order to enhance the effectiveness of current malaria control strategies. This paper empirically examines the factors that affect household choice of malaria treatment options in Ghana. The treatment options considered were choice of a public provider of health care, a private provider, purchase of drugs from a drug store, or self-medication. The results indicate that treatment and time costs are significant factors affecting the choice of health care provider. Education and household size also play an important role in malaria care seeking behaviour. The demand for malaria care is inelastic with respect to costs, and the magnitudes of the elasticities suggest that malaria care is a necessity. The policy implications are addressed. PMID- 15276318 TI - Impact of the European Union enlargement on health professionals and health care systems. AB - As the European family enlarges, the admission of new human resources in the health services will have an impact on the European market and health care system. Under the umbrella of the European Union (EU) equality, the educational quality barriers (e.g. PLAB test in UK, DIKATSA test in Greece) will be abolished. The overproduction of health professionals and their heterogeneous regional and per specialty distribution will lead to medical unemployment and demotion of the medical profession. Medical and political authorities and decision makers of the EU need to reform the European Health System, supervise, and assess the quality of medical education, harmonize the individual National Health System policies, and follow the World Health Organization (regional office for Europe) guidelines on health policy. An agreed, structured European Health Policy might moderate the vibrations of the forthcoming EU enlargement. PMID- 15276319 TI - Fatty acyl CoA-mediated inhibition of endoplasmic reticulum assembly. AB - The protein machinery that mediates homotypic fusion of mammalian endoplasmic reticulum (ER) membranes is becoming increasing well defined. However, little is known of how acylation of constituent membrane components might impact upon this event. This is particularly important as acylation has been shown to promote both fusion and fission of heterotypic membranes. Using a previously characterised cell-free ER fusion assay, I show here that incubation of membranes in the presence of either palmitoyl CoA or myristoyl CoA potently inhibits assembly. Furthermore, inhibition does not occur when membranes are incubated in the constituent palmitate or CoA moieties alone. These findings suggest that not only do palmitoyl CoA and myristoyl CoA inhibit ER assembly, but that they might instead be functioning to actively facilitate ER membrane fission. PMID- 15276320 TI - Sorting of lipoproteins to the outer membrane in E. coli. AB - Escherichia coli lipoproteins are anchored to the periplasmic surface of the inner or outer membrane depending on the sorting signal. An ATP-binding cassette (ABC) transporter, LolCDE, releases outer membrane-specific lipoproteins from the inner membrane, causing the formation of a complex between the released lipoproteins and the periplasmic molecular chaperone LolA. When this complex interacts with outer membrane receptor LolB, the lipoproteins are transferred from LolA to LolB and then localized to the outer membrane. The structures of LolA and LolB are remarkably similar to each other. Both have a hydrophobic cavity consisting of an unclosed beta-barrel and an alpha-helical lid. Structural differences between the two proteins reveal the molecular mechanisms underlying the energy-independent transfer of lipoproteins from LolA to LolB. Strong inner membrane retention of lipoproteins occurs with Asp at position 2 and a few limited residues at position 3. The inner membrane retention signal functions as a Lol avoidance signal and inhibits the recognition of lipoproteins by LolCDE, thereby causing their retention in the inner membrane. The positive charge of phosphatidylethanolamine and the negative charge of Asp at position 2 are essential for Lol avoidance. The Lol avoidance signal is speculated to cause the formation of a tight lipoprotein-phosphatidylethanolamine complex that has five acyl chains and therefore cannot be recognized by LolCDE. PMID- 15276321 TI - Involvement of mitochondrial and death receptor pathways in tributyltin-induced apoptosis in rat hepatocytes. AB - Tri-n-butyltin (TBT), a biocide, is known for its immunotoxicity and hepatotoxicity and is a well-characterised mitochondrial toxin. This report investigates the mechanisms involved in induction of apoptosis by TBT in primary cultures of rat hepatocytes. Release of cytochrome c from mitochondria into the cytosol was apparent after 15 min of exposure to 2.5 microM TBT. In addition, activity of initiator caspase-9 increased after 30 min, representing activation of the mitochondrial pathway in hepatocytes. The death receptor pathway was also activated by TBT, as indicated by recruitment of the adaptor protein FADD from the cytosol to the membrane as soon as 15 min after treatment. In addition, levels of the pro-apoptotic protein Bid decreased in the cytosol, while there was an increase in levels of the cleaved form tBid, in TBT-treated hepatocytes. Activity of initiator caspase-8 increased after 30 min. The principal effector caspase-3 was activated following 30 min of treatment with TBT. Activation was confirmed by immunodetection of a 17-kDa cleaved fragment. Apoptotic substrates such as Poly(ADP-ribose) polymerase and DNA fragmentation factor-45 are cleaved by caspase-3 to ensure the dismantlement of the cell. Cleavage of Poly(ADP ribose) polymerase into a 85-kDa fragment appeared after 30 min of TBT treatment. DNA fragmentation factor-45 disappeared in TBT-exposed rat hepatocytes. This is the first detailed study reporting the involvement of initiator and effector caspases, cleavage of their intracellular substrates and activation of both death receptor and mitochondrial pathways in TBT-induced apoptosis in rat hepatocytes. The comprehension of molecular events of apoptosis is important for the evaluation of the risk to humans and animals. PMID- 15276322 TI - The spectrin family member Syne-1 functions in retrograde transport from Golgi to ER. AB - To address the function of the Golgi- and nuclear envelope-localized spectrin family member synaptic nuclear envelope protein-1 (Syne-1), we expressed two separate recombinant fragments derived from the central portion of the molecule. Both of these fragments were predicted to act as dominant negative inhibitors of Syne-1 function at the Golgi. One of the fragments was previously shown to bind the Golgi complex. The other fragment was found to form microtubule-associated puncta that sequester endogenous Syne-1. Expression of either fragment resulted in a cell type-specific alteration in the structure of the Golgi complex, which appeared to collapse into a compact juxtanuclear structure in some cell types but not others. These fragments were expressed in cultured cells and their effects on Golgi function were examined. Expression of both dominant negative Syne-1 fragments blocked recycling of the endoplasmic reticulum (ER) resident protein disulfide isomerase (PDI), which accumulated in the Golgi complex. In addition, we found that fragment expression altered the distribution of the KDEL receptor and the COP-I coat protein beta-COP, two proteins known to be involved in regulating the retrograde pathway. We conclude that these results indicate a role for Syne-1 in facilitating retrograde vesicular trafficking from the Golgi to the ER. PMID- 15276323 TI - Overexpression of antioxidant enzyme peroxiredoxin 5 protects human tendon cells against apoptosis and loss of cellular function during oxidative stress. AB - Oxidative stress and apoptosis are implicated in tendon degeneration. Peroxiredoxin 5 (PRDX5) is a novel thioredoxin peroxidase recently identified in mammals, participating directly in eliminating hydrogen peroxide (H(2)O(2)) and neutralizing other reactive oxygen species (ROS). We have previously reported that PRDX5 is upregulated in degenerative human tendon. However, the effects of this upregulation on human tendon cell function remain unknown, in particular, with regards to oxidative stress conditions. Here we report that exposure of human tendon cells to 50 microM H(2)O(2) for 24 h (in vitro oxidative stress) caused a significant increase in the percentage of apoptotic cells (P<0.05) as assessed by flow cytometric analysis of Annexin V binding, accompanied by increased PRXD5 mRNA and protein expression. Overexpression of PRDX5 in human tendon cells via transfection inhibited H(2)O(2)-induced tendon cell apoptosis by 46% (P<0.05), and prevented the decrease in tendon cell collagen synthesis which occurs under H(2)O(2) challenge, although the decrease in collagen synthesis was small. Results from our study indicate that the antioxidant enzyme PRDX5 plays a protective role in human tendon cells against oxidative stress by reducing apoptosis and maintaining collagen synthesis. PMID- 15276325 TI - Overexpression of catalase or Bcl-2 alters glucose and energy metabolism concomitant with dexamethasone resistance. AB - Glucocorticoids induce apoptosis in lymphocytes by causing the release of cytochrome c into the cytosol; however, the events in the signaling phase between translocation of the steroid-receptor complex to the nucleus and the release of cytochrome c have not been elucidated. Previously, we found that, in response to steroid treatment, WEHI7.2 mouse thymic lymphoma cells overexpressing catalase (CAT38) show delayed apoptosis (delayed cytochrome c release) compared to the parental cells, while Bcl-2 overexpressing cells (Hb12) are protected from steroid-induced apoptosis. In lymphocytes, glucocorticoid treatment decreases glucose uptake. Both glucose deprivation and the attendant ATP drop are known inducers of apoptosis. Therefore, we used (31)P and (1)H NMR spectroscopy to compare metabolic profiles of WEHI7.2, CAT38 and Hb12 cells in the presence and absence of dexamethasone to determine: (1) whether glucocorticoid effects on glucose metabolism contribute to the mechanism of steroid-induced apoptosis; and (2) whether catalase or Bcl-2 overexpression altered metabolism thereby providing a mechanism of steroid resistance. Loss of mitochondrial hexokinase activity was correlated to the induction of apoptosis in WEHI7.2 and CAT38 cells. CAT38 and Hb12 cells have an altered basal metabolism which includes increases in hexokinase activity, lactate production when subcultured into new medium, use of mitochondria for ATP production and potentially increased glutaminolysis. These data suggest that: (1) glucocorticoid effects on glucose metabolism may contribute to the mechanism of steroid-induced lymphocyte apoptosis; and (2) the altered metabolism seen in catalase and Bcl-2 overexpressing cells may contribute to both the steroid resistance and increased tumorigenicity of these variants. PMID- 15276324 TI - Oxaline, a fungal alkaloid, arrests the cell cycle in M phase by inhibition of tubulin polymerization. AB - Oxaline and neoxaline, fungal alkaloids, were found to inhibit cell proliferation and to induce cell cycle arrest at the G(2)/M phase in Jurkat cells. CBP501 (a peptide corresponding to amino acids 211-221 of Cdc25C phosphatase), which inhibits the G(2) checkpoint, did not affect the G(2)/M arrest caused by oxaline, suggesting that oxaline causes M phase arrest but not G(2) phase arrest. The Cdc2 phosphorylation level of oxaline-treated cell lysate was lower than that of the control cells, indicating that oxaline arrests the M phase. Oxaline disrupted cytoplasmic microtubule assembly in 3T3 cells. Furthermore, oxaline inhibited polymerization of microtubule protein and purified tubulin dose-dependently in vitro. In a binding competition assay, oxaline inhibited the binding of [(3)H]colchicine to tubulin, but not that of [(3)H]vinblastine. These results indicate that oxaline inhibits tubulin polymerization, resulting in cell cycle arrest at the M phase. PMID- 15276326 TI - Pro-apoptotic signaling pathway activated by echistatin in GD25 cells. AB - Disintegrins, low molecular weight RGD-containing polypeptides isolated from snake venoms, have seen use as integrin antagonists in the field of tumor biology and angiogenesis. In this study, we investigated the molecular mechanism by which the disintegrin echistatin affects cell adhesion and signaling resulting in an apoptotic response in the GD25 cell system. Wild-type GD25 cells, which lack expression of the beta(1) family of integrin, and stable transfectants expressing the A isoform of beta(1) integrin subunit were used. Nanomolar concentrations of echistatin detached fibronectin- and vitronectin-adherent GD25 cells from immobilized substratum. However, prior to inducing detachment of adherent cells, echistatin caused apoptosis as measured by caspase-3 activation. Either cell detachment or apoptotic response induced by echistatin were more pronounced on fibronectin-adherent GD25 cells than on vitronectin-adherent ones. GD25 cell exposure to echistatin caused a reduction of tyrosine phosphorylation levels of pp125(FAK), whereas it didn't affect either Shc tyrosine phosphorylation levels or Shc-Grb2 functional association. The down-regulation of pp125(FAK) preceded apoptosis and cell detachment induced by echistatin. Our results indicate that pp125(FAK) and not Shc pathway is involved in echistatin-induced apoptotic response in the GD25 cell system. PMID- 15276327 TI - Preliminary kinetics and metabolism of 2,4,6-trinitrotoluene and its reduced metabolites in an aquatic oligochaete. AB - We examined the toxicokinetics and metabolism of 2,4,6-trinitirotoluene (TNT) and four of its major reduced metabolites (2-amino-4,6-dinitrotoluene (2ADNT), 4 amino-2,6-dinitrotoluene (4ADNT), and 2,4-diamino-6-nitrotoluene (2,4DANT)) in the freshwater, aquatic oligochaete Tubifex tubifex exposed to spiked, reconstituted water. In uptake experiments with each compound, steady state concentrations were reached within 1h, and all absorbed compounds were completely eliminated in 0-3 h. The appearance of 2ADNT and 4ADNT (from metabolism of absorbed TNT) was five times slower, reaching 95% of steady state in 14.2-16.1h. Approximately, 82% of absorbed TNT was metabolized to ADNTs; metabolism to 4ADNT was favored over 2ADNT by a factor of 3. No further metabolism of ADNTs to DANTs was detected. After a loss of 29-50% of metabolically-generated ADNTs during the first hour of the TNT depuration experiment, Tubifex ADNT concentrations remained constant throughout the 53h depuration period. This suggests differences between the toxicokinetics of absorbed ADNTs and the toxicokinetics of metabolically generated ADNTs. Experiments using radiolabeled (14C) TNT revealed that only 15 47% of 14C-TNT equivalents were identified as TNT, 2ADNT, or 4ADNT, indicating significant metabolism and/or binding to biomolecules. Of unidentified 14C-TNT equivalents, 28-38% remained unextractable. Both unextractable and extractable unidentified substances increased throughout the 54 h 14C-TNT uptake experiment. The unidentified portions of the radiolabel were not eliminated during a 53-h depuration experiment. Bioconcentration factors (BCFs) for HPLC-detectable compounds were found to be linearly related to log K(OW) (r2 = 0.9963). BCFs for 2ADNT, 4ADNT, and 2,4DANT were 10.22, 12.41, and 2.75, respectively. The BCF for TNT was 12.25, based on a molar sum of total TNT and its metabolites (SigmaTNT + 2ADNT + 4ADNT), and 2.53 based on TNT only. Compound hydrophobicity predicted the toxicokinetics and bioconcentration of compounds absorbed from water, however, the large discrepancy between the toxicokinetics of absorbed versus metabolically generated ADNTs and the bioconcentration and toxicokinetics of the unidentified substances warrant further investigation. PMID- 15276328 TI - Expression of metallothionein gene during embryonic and early larval development in zebrafish. AB - Metallothionein (Mt) has been considered as a molecular marker of metal pollution in aquatic ecosystems. Less is known about the expression of mt gene during embryogenesis. Here, we report the cloning, sequencing, and the expression pattern of mt gene during developmental stages in zebrafish. The zebrafish embryogenesis when takes place in a medium containing a dosage of 1000 microM zinc resulted in high mortality, indicating the deleterious effect of zinc on development. The zebrafish mt gene consists of three exons encoding 60 amino acids with 20 conserved cysteine residues. RT-PCR result indicates the maternal contribution of Mt transcripts. Using digoxigenin (DIG)-labeled anti-sense RNA probe, whole-mount in situ hybridization was performed to observe the expression pattern of zebrafish mt gene during embryonic and early larval stages. Stronger as well as ubiquitous expression of mt gene during early embryonic stages narrowed to specific expression after hatching. The mt promoter region contains seven copies of putative metal-responsive elements (MREs), which are shown to be important for the high level activity by deletion analysis. The expression of mt gene during embryogenesis implies its significant role on development. PMID- 15276329 TI - Effects of the brominated flame retardants hexabromocyclododecane (HBCDD), and tetrabromobisphenol A (TBBPA), on hepatic enzymes and other biomarkers in juvenile rainbow trout and feral eelpout. AB - Brominated flame retardants (BFRs) leak out in the environment, including the aquatic one. Despite this, sublethal effects of these chemicals are poorly investigated in fish. In this study, a screening of selected biomarkers in juvenile rainbow trout (Oncorhynchus mykiss) and feral eelpout (Zoarces viviparus) was performed after exposure to hexabromocyclododecane (HBCDD) and tetrabromobisphenol A (TBBPA). Rainbow trout was injected intraperitoneally (i.p.) with HBCDD or TBBPA. Two out of four short-term experiments with HBCDD showed an increase in the activity of catalase. A 40% increase in liver somatic index (LSI) could be observed after 28 days. HBCDD did also seem to have an inhibitory effect on CYP1A's activity (ethoxyresorufin-O-deethylase (EROD)). A putative peroxisome proliferating activity of the compound was investigated without giving a definite answer. HBCDD did not seem to be estrogenic or genotoxic. TBBPA increased the activity of glutathione reductase (GR) after 4, 14 and 28 days in rainbow trout suggesting a possible role of this compound in inducing oxidative stress. The compound did not seem to be estrogenic. TBBPA seemed to compete with the artificial substrate ethoxyresorufin in vitro, during the EROD assay. In eelpout, only one 5 days in vivo experiment was performed. Neither of the compounds gave rise to any effect in this fish. This was the first screening of sublethal effects of the two chemicals in fish, using high doses. Our results indicate that there is a need for further studies of long-term, low dose effects of these two widely used flame retardants. PMID- 15276330 TI - Cadmium-induced apoptosis through the mitochondrial pathway in rainbow trout hepatocytes: involvement of oxidative stress. AB - Cadmium (Cd) induces oxidative stress and apoptosis in trout hepatocytes. We therefore investigated the involvement of the mitochondrial pathway in the initiation of apoptosis and the possible role of oxidative stress in that process. This study demonstrates that hepatocyte exposure to Cd (2, 5 and 10 microM) triggers significant caspase-3, but also caspase-8 and -9 activation in a dose-dependent manner. Western-blot analysis of hepatocyte mitochondrial and cytosolic fractions revealed that cytochrome c (Cyt c) was released in the cytosol in a dose-dependent manner, whereas the pro-apoptotic protein Bax was redistributed to mitochondria after 24 and 48 h exposure. We also found that the expression of anti-apoptotic protein Bcl-xL, known to be regulated under mild oxidative stress to protect cells from apoptosis, did not change after 3 and 6 h exposure to Cd, then increased after 24 and 48 h exposure to 10 microM Cd. In the second part of this work, two antioxidant agents, 2,2,6,6-tetramethylpiperidinyl 1-oxyl (TEMPO) (100 microM) and N-acetylcysteine (NAC, 100 microM) were used to determine the involvement of reactive oxygen species (ROS) in Cd-induced apoptosis. Simultaneously exposing trout hepatocytes to Cd and TEMPO or NAC significantly reduced caspase-3 activation after 48 h and had a suppressive effect on caspase-8 and -9 also, mostly after 24 h. Lastly, the presence of either one of these antioxidants in the treatment medium also attenuated Cd induced Cyt c release in cytosol and the level of Bax in the mitochondria after 24 and 48 h, while high Bcl-xL expression was observed. Taken together, these data clearly evidenced the key role of mitochondria in the cascade of events leading to trout hepatocyte apoptosis in response to Cd and the relationship that exists between oxidative stress and cell death. PMID- 15276331 TI - Effects of beta-naphthoflavone, phenobarbital and dichlobenil on the drug metabolizing system of liver and nasal mucosa of Italian water frogs. AB - In this study, we have examined the presence and inducibility of phase I and II drug-metabolizing enzymes in the liver and nasal mucosa of Italian water frogs of control and pretreated with beta-naphthoflavone, phenobarbital and dichlobenil by using typical substrates for these enzymes along with polyclonal antibodies mainly raised against mammalian enzymes. The CYP content and various monooxygenase and phase II enzyme activities in the liver of this frog were found similar, when reported, to those of largely aquatic and semiaquatic frogs. The treatment with beta-naphthoflavone resulted in an induction in the liver of a CYP1A and the induction was manifested by (a) immunoblot analysis using anti-rat CYP1A1, (b) an increase of CYP1A-mediated methoxyresorufin-O-demethylase and ethoxyresorufin-O-deethylase activities. The treatments with both phenobarbital and dichlobenil did not produce in the liver any effect on the assayed enzymes. When the nasal mucosa of water frogs was analyzed, various monooxygenase and phase II enzymatic activities, generally comparable to those of liver, were determined. However, by using antibodies anti-three GST different classes, we found a different reactivity into the cytosol of the two tissues indicating a differential tissue susceptibility to toxic effects of xenobiotics. In the nasal mucosa, a protein immunorelated to CYP2A and monooxygenase activities (i.e. ethoxycoumarin-O-deethylase and coumarin-7-hydroxylase) linked in mammals to this isoform have also been found. The treatment of water frogs with the herbicide dichlobenil decreased both the above-mentioned activities and the immunoreactive CYP2A apoprotein. The pretreatment with metyrapone, a CYP inhibitor, protected the CYP2A apoprotein and its linked activities from toxic effect of dichlobenil indicating a key role of this enzyme in the bioactivation of this herbicide. The findings of the present work suggest that the hepatic CYP1A induction and the nasal CYP2A-like inhibition profiles might provide two potential biomarkers of the Italian water frogs exposure to environmental and aquatic pollutants. PMID- 15276332 TI - Effects of short-term copper exposure on gill structure, metallothionein and hypoxia-inducible factor-1alpha (HIF-1alpha) levels in rainbow trout (Oncorhynchus mykiss). AB - Rainbow trout (Oncorhynchus mykiss) were exposed to 1.65 microM of waterborne copper for 24 h. Fish were then transferred to metal-free water. Metallothionein mRNA induction in rainbow trout liver and gill tissue, hypoxia-inducible factor-1 (HIF-1alpha) accumulation in gill tissue and arithmetic mean thickness of gill epithelium (Har) were determined at 4 and 24h of exposure as well as 48 h after transfer to metal-free water. The arithmetic mean distance from water to blood was significantly elevated after both 4 and 24 h of exposure (Har was 4.67 and 4.66 microm, respectively in exposed fish, compared to 3.81 and 3.62 microm for the corresponding control fish). During the 48 h recovery Har returned towards the control values; the recovery value of 4.21 microm was significantly lower than values during exposures. There was also a significant increase in gill metallothionein mRNA levels after the 4 h exposure with MT/GAPDH ratio of 1.288 versus the control value of 0.988. In liver, metallothionein induction was not observed. HIF-1alpha protein showed an increased accumulation in gills after 4 h, with the HIF-1alpha/alpha-tubulin ratio of 0.562 being significantly higher than the 24 h exposure value of 0.232. These results suggest that exposure to copper for four hours causes hypoxia in the gill epithelium, which is adequate for the activation of HIF-1alpha. PMID- 15276333 TI - Potential drug (oxytetracycline and oxolinic acid) pollution from Mediterranean sparid fish farms. AB - The potential for input of two common antibacterial agents in Mediterranean fish farms, oxytetracycline (OTC) and oxolinic acid (OA), was estimated from measurements of these drugs in the faecal excretions of two important farmed sparids, gilthead sea bream, Sparus aurata and sharpsnout sea bream Diplodus puntazzo. Oxolinic acid was found to be well absorbed by gilthead sea bream (92%) and sharpsnout sea bream (88%) while the absorption of OTC was found to be considerably lower in both species (27 and 40%, respectively). These data were integrated with production records for sparids, drug dosage regimes and treatment frequency information to calculate potential annual drug release to the aquatic environment from Greek fish farms. These calculations suggest potentially significant quantities of unmetabolised OTC can be passed unabsorbed through the body of treated sparids and excreted via the faeces into the local marine environment. The situation with OA was much less pronounced. It was estimated that potentially more than 1900 kg of OTC and more than 50 kg of OA may be released via faecal excretion into the environment by sparid farms per year. Further drug may also be released via uneaten medicated feed, leached drugs and other routes of fish elimination (renal excretion, branchial secretions). Drug pollution of the marine environment in the vicinity of fish farms can have adverse ecological effects, including development of resistant bacterial populations and exposure with potential drug accumulation in aquatic fauna and flora. PMID- 15276334 TI - Differential susceptibility of horizontal and vertical swimming activity to cadmium exposure in a gammaridean amphipod (Gammarus lawrencianus). AB - In this study two indices of swimming behavior (horizontal and vertical swimming activity) in a gammaridean amphipod (Gammarus lawrencianus) were examined for their sensitivity to Cd exposure. G. lawrencianus were exposed for 72 h to a variety of Cd concentrations [background (approximately 12), 62, 125, 250 and 500 microg l(-1)] at 20 ppt. Subsequent to exposure, video surveillance of survivors held within grooved rings or clear boxes was used to assess horizontal swimming activity (percentage of time mobile) and vertical swimming activity (number of surfacings), respectively. Results show that control amphipods were quite active, being mobile approximately 61% of the time, with horizontal swimming activity decreasing (P < 0.01) to approximately 0.3% between a Cd exposure concentration of 125 and 500 microg l(-1). Vertical swimming activity in amphipods was also impacted by Cd exposure (P < 0.001), with the greatest decrease occurring between background (approximately 12 microg l(-1)) and 62 microg l(-1) (60 versus approximately 26 surfacings, respectively), which is approximately four-fold lower than the estimated 72 h LC50 (250 microg l(-1)) for G. lawrencianus. Based on fluid dynamic considerations, it is speculated that of the two behaviors, vertical swimming activity is more sensitive to Cd exposure because of the presumed greater energetic costs associated with producing enough thrust to attain the lift required to make a vertical ascent into the water. PMID- 15276335 TI - Regarding "Namesake biography: Dwight David Eisenhower Army Medical Center". PMID- 15276336 TI - Controversies in laparoscopic surgery for colorectal cancer. PMID- 15276337 TI - Postoperative ileus. PMID- 15276338 TI - Restorative proctocolectomy: one stage or two? PMID- 15276339 TI - Adenoviral gene therapy for pancreatic adenocarcinoma. PMID- 15276340 TI - Current management of diverticulitis. PMID- 15276341 TI - Laparoscopic colectomy for diverticulitis. PMID- 15276342 TI - Abdominal apoplexy: a case study of the spontaneous rupture of the gastroepiploic artery. AB - This is a case report of abdominal apoplexy (AA) or spontaneous rupture of a visceral vessel, without associated aneurysmal dilation of the vessel. Spontaneous rupture of the left gastroepiploic artery (LGEA) resulting in a hemoperitoneum is discussed. The clinical presentation of left lower quadrant abdominal pain, along with the histologic findings of medial degeneration of the LGEA, makes this case an unusual one. The incidence, origin, associated predisposing medical conditions, clinical presentation, and treatment of abdominal apoplexy are discussed. PMID- 15276343 TI - Analgesic administration prior to surgical evaluation for acute appendicitis. AB - BACKGROUND: The attitudes of surgeons and nonsurgeons regarding the administration of pain medicine prior to arriving at a surgical diagnosis are changing. It is common practice to administer narcotic analgesics prior to a general surgeon's evaluation. Several studies have advocated the safety of this practice in the emergency department. Many of these studies are flawed by inclusion of many patients who did not have a surgical illness. Our study examined the practice of narcotic administration in patients determined to have appendicitis who underwent operation. METHODS: We retrospectively reviewed 75 consecutive appendectomies. Emergency department records and in-patient charts were reviewed to assess differences in 2 groups of patients: those who received narcotic pain medicine and those who did not. Specific outcome parameters were reviewed such as time in hospital, time to the operating room, and complication rate. We also created a scoring system for the physical examination to attempt to quantify a difference between the groups. FINDINGS: Overall, 75 patient charts were reviewed. Nine patients were excluded. There was no statistically significant difference in the 2 groups in regard to time in hospital, time to operation, complication rate, perforation rate, or negative appendectomy rate. The physical examination scoring system did show a difference between those who got pain medicine and those who did not, but failed to show a difference between examiners after pain medicine was given. CONCLUSIONS: There does not appear to be a difference in hospital stay, time to the operating room, complication rate, negative appendectomy rate, or perforation rate in patients who received pain medicine prior to a surgeon's evaluation and those who did not in this retrospective review. PMID- 15276344 TI - Bilateral reexpansion pulmonary edema after treatment of spontaneous pneumothorax. PMID- 15276345 TI - Iatrogenic intestinal injury concomitant to iatrogenic bile duct injury: the second component. AB - OBJECTIVE: Bile duct injuries have a frequency of 0.1% to 0.3% even in the most experienced centers. Complex biliary lesions usually require a bilioenteric anastomosis, achieving good long-term results in 80% to 90% of the cases. Besides injuries to the abdominal contents during laparoscopy (by trocars or electrocautery), intestinal complications associated with reconstruction attempts can be observed. We analyzed the concomitant intestinal complications in 251 patients with iatrogenic biliary injuries reconstructed over this 12-year period. METHODS: A retrospective review of patients with biliary tract reconstruction after iatrogenic injury in a tertiary academic health-care center was done. All patients with concomitant intestinal injury were included; type of operation and postoperative outcome were analyzed. RESULTS: Among 251 patients, 35 cases had a concomitant intestinal injury. The most common site of fistulization was the duodenum (18 cases, 50%); 9 cases were associated with long-term subhepatic drains (more than three weeks), and the other 9 cases were associated with a dehiscent hepatoduodenostomy. Faulty Roux-en-Y reconstruction was observed in 5 cases. In 5 cases, fistulization of the jejunum and ileum, secondary to drain placement, was documented, as well as 3 cases with colonic injuries. Two patients had a dehisced Roux-en-Y anastomosis. One had a bilioenteric omega type ileal anastomosis, and 1 had a hepatoileal anastomosis without omega reconstruction. Primary repair of the duodenum with resection of the affected intestinal or colonic segment was done at the same time of biliary repair without related morbidity. CONCLUSIONS: Concomitant gastrointestinal injures were found with an incidence of 15% in our series. The most common site of fistulization is the duodenum. In half of the patients, it was secondary to a dehiscent hepatoduodenostomy, whereas in the other, it was caused by long-term subhepatic drains. Besides faulty Roux-en-Y reconstruction, fistulization was related with long-term drains. Primary repair and resection of the affected segment of jejunum, ileum, and colon can be done during the same operative stage of biliary reconstruction, without significant correlated mortality. PMID- 15276346 TI - Intestinal lymphoma causing intussusception in HIV(+) patient: a rare presentation. AB - Ten-percent of all malignancies affecting the HIV(+) patient population are lymphomas. Lymphoma involving the gastrointestinal tract may be more common than anticipated in this select group of patients. Because this patient population is frequently seen by the surgeon for abdominal complaints, the diagnosis of enteric lymphoma should be entertained and the general surgeons should be aware of its frequency. We report a case of intussusception caused by enteric lymphoma in an HIV(+) patient. PMID- 15276347 TI - Invasive pancreatic cancer presenting as gastrointestinal hemorrhage--a case report. AB - Upper gastrointestinal hemorrhage is not uncommonly seen by the surgical practitioner. We present a case of a patient who presented with melena and syncope, who was subsequently found to have invasive metastatic pancreatic cancer as his source of bleed. PMID- 15276348 TI - Are you alive in there? PMID- 15276349 TI - Looking forward to the unknown. PMID- 15276350 TI - Medical education: a forgotten item in the health care budget. PMID- 15276351 TI - Surgeon of one: future perspectives of an army surgeon. PMID- 15276353 TI - Victor Paul Satinsky--renaissance doc. PMID- 15276354 TI - In vivo evaluation of tetrahedral amorphous carbon. AB - The in vivo behavior and tissue reaction to tetrahedral amorphous carbon (ta-C) has been evaluated for periods of up to 6 months in SV129 mice. Two sample types were tested--silicon die coated with ta-C (n = 53) and micromachined particles (n = 40). The coated samples were compared to uncoated silicon die (n = 22). Die samples were implanted subcutaneously, and tissue reaction and capsule formation were evaluated at various time points. Micromachined particles of 1, 3, 10, and 30 microm were injected adjacent to the sciatic nerve, and tissue samples were examined histologically at various time points (4 days-6 months). Tissue reaction to ta-C was mild and was localized to the area of the injection or implantation. Samples with a higher ratio of 3-fold bonding appeared to shed material during the experiments; this was not observed on samples with a higher level of 4-fold bonding, nor on uncoated silicon die. The results strongly suggest that films with greater 4-fold bonding character (more diamond-like) are more resistant to in vivo fragmentation than films with higher 3-fold character (more graphitic). PMID- 15276355 TI - The influence of the phosphorus content on the bioactivity of sol-gel glass ceramics. AB - The aim of this work was to study the influence of the phosphorus on the crystallization and bioactivity of glass-ceramics obtained from sol-gel glasses. For this purpose two sol-gel glasses with a similar composition but one of them containing P2O5 (70% SiO2; 30% CaO and 70% SiO2; 26% CaO; 4% P2O5, mol%) were prepared. Pieces of these glasses were treated at temperatures ranging between 700 degrees C and 1400 degrees C for 3 h. The obtained materials were characterized by XRD, FTIR, SEM-EDS and the biaxial flexural strength was determined in samples heated at 1100 degrees C. In addition, an in vitro bioactivity study in simulated body fluid (SBF) was carried out. The results showed that phosphorus plays an important role in the crystallization of the glasses: it induced the crystallization of calcium phosphate phases, the stabilization of the wollastonite phase at high temperature as well as the crystallization of SiO2 phases at low temperatures. Moreover, the presence of phosphorus produced a heterogeneous distribution of defects in the pieces and, therefore, the flexural strength of samples containing this element decreased. Finally, glass-ceramics obtained from glasses containing phosphorus showed the fastest formation rate of the apatite layer when soaked in SBF. PMID- 15276356 TI - In vitro cytocompatibility of MG63 cells on chitosan-organosiloxane hybrid membranes. AB - Chitosan-silicate hybrids were synthesized using gamma glycidoxypropyltrimethoxysilane (GPSM) as the agent for cross-linking the chitosan chains. CaCl2 was introduced in the hybrids in expectation that it would improve cell adhesion and differentiation of the hybrid surfaces. Fourier transform infrared (FT-IR) spectroscopy and 29Si CP-MAS NMR spectroscopy were used to analyze the structures of the hybrids. Cytocompatibility of the hybrids was investigated in terms of proliferation of an osteoblastic cell line, MG63. The adhesion and proliferation of the osteoblastic cells cultured on the surface of a chitosan-GPSM hybrid without calcium were similar to those on a control culture plate, and were better than those on a chitosan membrane. The ALP activity of the cells cultured on this hybrid was higher than that on the chitosan membrane. Contrary to expectations, the incorporation of calcium ions into the hybrids did not improve cell attachment and proliferation on their surfaces. PMID- 15276357 TI - Multiphoton autofluorescence imaging of intratissue elastic fibers. AB - Multiphoton induced blue/green autofluorescence by near infrared femtosecond laser pulses has been used to selectively image intratissue elastic fibers in native and tissue engineered (TE) viable heart valves without any invasive tissue removal, embedding, fixation, and staining. Elastic fibers could be clearly distinguished from collagenous structures which emit ultraviolet/violet radiation when excited with intense ultrashort pulses due to second harmonic generation. Deep-tissue three-dimensional imaging of elastic fibers with submicron spatial resolution was performed by optical sectioning of heart valves using a multiphoton laser scanning microscope in connection with a tunable 80 MHz femtosecond laser source. The technology was used to diagnose extracellular matrix structures and cell resettlement of TE heart valves prior implantation. This novel non-invasive method opens the general possibility of high-resolution in situ imaging of elastic fibers, collagen structures and intracellular organelles in living intact tissues without staining. PMID- 15276358 TI - Antibacterial activity of polymeric substrate with surface grafted viologen moieties. AB - An asymmetric viologen, N-hexyl-N'-(4-vinylbenzyl)-4,4'-bipyridinium bromide chloride (HVV), was synthesized and graft copolymerized with commercial PET films. The surface graft concentration of HVV on the PET film is easily controlled by varying the monomer concentration used in the UV-induced graft copolymerization process. The HVV surface functionalized PET film functions as a smart window whose transmittance is reduced upon exposure to light. Concomitantly, the film possesses antibacterial activity, as shown by its bactericidal effect on Escherichia coli (E. coli). The antibacterial activity depends on the concentration of pyridinium groups on the surface and a surface concentration of 25 nmol/cm2 on PET has been shown to be highly effective in killing the bacteria. PMID- 15276359 TI - Improvement in biocompatibility of ZrO2-Al2O3 nano-composite by addition of HA. AB - The biocompatibility of zirconia-alumina (ZA) nano-composites in load-bearing applications such as dental/orthopedic implants was significantly enhanced by the addition of bioactive HA. The ZA matrix was composed of nano-composite powder obtained from the Pechini process and had higher flexural strength than conventionally mixed zirconia-alumina composite. Because the ZA nano-composite powder effectively decreased the contact area between HA and zirconia for their reaction during the sintering process, the HA-added ZA nano-composites contained biphasic calcium phosphates (BCP) of HA/TCP and had higher flexural strength than conventionally mixed ZA-HA composite. From the in vitro test with osteoblastic cell-lines, the proliferation and the differentiation (as expressed by the alkaline phosphatase activity) of the cellular response on the HA-added ZA nano composites gradually increased as the amount of HA added increased. From the mechanical and biological evaluations of the HA-added ZA nano-composites, 30HA (30 vol% HA + 70 vol% ZA) was found to be the optimal composition for load bearing biological applications. PMID- 15276360 TI - In vivo cytokine secretion and NF-kappaB activation around titanium and copper implants. AB - The early biological response at titanium (Ti), copper (Cu)-coated Ti and sham sites was evaluated in an in vivo rat model. Material surface chemical and topographical properties were characterized using Auger electron spectroscopy, energy dispersive X-ray spectroscopy and interferometry, respectively. The number of leukocytes, cell types and cell viability (release of lactate dehydrogenase) were determined in the implant-interface exudate. The contents of activated nuclear transcription factor NF-kappaB, interleukin-6 (IL-6) and interleukin-10 (IL-10) were determined by enzyme linked immunosorbent assay. An increase in the number of leukocytes, in particular, polymorphonuclear leukocytes, was observed between 12 and 48 h around Cu. A marked decrease of exudate cell viability was found around Cu after 48 h. The total amounts of activated NF-kappaB after 12 h was highest in Ti exudates whereas after 48 h the highest amount of NF-kappaB was detected around Cu. The levels of cytokine IL-6 were consistently high around Cu at both time periods. No differences in IL-10 contents were detected, irrespective of material/sham and time. The results show that materials with different toxicity grades (titanium with low and copper with high toxicity) exhibit early differences in the activation of NF-kappaB, extracellular expression and secretion of mediators, causing major differences in inflammatory cell accumulation and death in vivo. PMID- 15276361 TI - Bladder acellular matrix as a substrate for studying in vitro bladder smooth muscle-urothelial cell interactions. AB - The objective of this study was to evaluate the ability of bladder acellular matrix (BAM) to support the individual and combined growth of primary porcine bladder smooth muscle (SMC) and urothelial (UEC) cells. An in vitro co-culture system was devised to evaluate the effect of UEC on (i) SMC-mediated contraction of BAM discs, and (ii) SMC invasiveness into BAM. Cells were seeded onto BAM discs under 4 different culture conditions. Constructs were incubated for 1, 7, 14 and 28 days. Samples were then harvested for evaluation of matrix contraction. Immunohistochemistry (IHC) was utilized to examine cellular organization within the samples and conditioned media supernatants analyzed for net gelatinase activity. BAM contraction was significantly increased with co-culture. The same side co-culture configuration lead to a greater reduction in surface area than opposite side co-culture. IHC revealed enhanced SMC infiltration into BAM when co culture was utilized. A significant increase in net gelatinase activity was also observed with the co-culture configuration. Enhanced infiltration and contractile ability of bladder SMCs with UEC co-culture may, in part, be due to an increase in gelatinase activity. The influence of bladder UECs on SMC behaviour in vitro indicates that BAM may contain some key inductive factors that serve to promote important bladder cell-cell and cell-matrix interactions. PMID- 15276362 TI - Effect of functionalization of multilayered polyelectrolyte films on motoneuron growth. AB - We studied in vitro cell-substrate interaction of motoneurons with functionalized polylectrolyte films. Thin polylectrolyte films were built on glass by alternating polycations, poly(ethylene-imine) PEI, poly(L-lysine) PLL, or poly(allylamine hydrochloride) PAH, and polyanions, poly(sodium-4 styrenesulfonate) PSS or poly(L-glutamic acid) (PGA). These architectures were functionalized with Brain Derived Neurotrophic Factor (BDNF) or Semaphorin 3A (Sema3A). We used Optical Waveguide Lightmode Spectroscopy (OWLS) and Atomic Force Microscopy (AFM) to characterize the architectures. The viability of motoneurons was estimated by the acid phosphatase method, and morphometrical measures were performed to analyse the influence of different architectures on cell morphology. Motoneurons appeared to adhere and spread on all the architectures tested and preferentially on PSS ending films. The viability of motoneurons on polyelectrolyte multilayers was higher compared to polyelectrolyte monolayers. BDNF and Sema3A embedded in the films remained active and thereby create functionalized nanofilms. PMID- 15276363 TI - A multifunctional bioreactor for three-dimensional cell (co)-culture. AB - Investigation of cell abilities to growth, proliferation and (de)-differentiation in a three-dimensional distribution is an important issue in biotechnological research. Here, we report the development of a new bioreactor for three dimensional cell culture, which allows for co-cultivation of various cell types with different culture conditions in spatial separation. Preliminary results of neonatal rat cardiomyocyte cultivation are shown. Isolated neonatal rat cardiomyocytes were cultured in spatial separated bioreactor compartments in recirculating medium on a biodegradable fibrin matrix for 2 weeks. Glucose, lactate, and lactate dehydrogenase (LDH), pO2, pCO2, and pH levels were monitored in the recirculated medium, daily. Morphological characterization of matrix and cells was assessed by hematoxylin and eosin staining, and MF-20 co-immunostaining with 4',6-diamidino-2-phenylindole (DAPI). Cell viability was determined by LIVE/DEAD staining before cultivation and on day 3, 7, and 14. The optimized seeding density in the matrix was 2.0 x 10(7) cells retaining cellular proportions over the cell culture period. The bioreactor allows the maintenance of physiologic culture conditions with aerobic cell metabolism (low release of lactate, LDH), a high oxygen tension (pO2-183.7 +/- 18.4 mmHg) and physiological pH values (7.4 +/- 0.02) and a constant level of pCO2 (43.1 +/- 2.9) throughout the experimental course. The cell viability was sufficient after 2 weeks with 82 +/- 6.7% living cells. No significant differences were found between spatial separated bioreactor compartments. Our novel multifunctional bioreactor allows for a three-dimensional culture of cells with spatial separation of the co cultured cell groups. In preliminary experiments, it provided favorable conditions for the three-dimensional cultivation of cardiomyocytes. PMID- 15276364 TI - Osteoblast alignment, elongation and migration on grooved polystyrene surfaces patterned by Langmuir-Blodgett lithography. AB - Topographically patterned surfaces are known to influence cellular behavior in a controllable manner. However, the relatively large surface areas (several cm2) required for many biomaterial applications are beyond the practical limits of traditional lithography. Langmuir-Blodgett lithography, a recently developed method, was used to fabricate regularly spaced grooves of different depths (50 and 150 nm) with a periodicity of 500 nm over several square centimeter on silicon surfaces. These topographies were transferred into polystyrene surfaces by means of nanoimprinting. Primary osteoblasts were cultured on the patterned polymer surfaces. They were observed to align, elongate and migrate parallel to the grooves. The combination of Langmuir-Blodgett lithography with nanoimprinting enables the fabrication of large, nanostructured surface areas on a wide spectrum of different biomaterials. Osteoblasts show a significant anisotropic behavior to these surfaces, which can enhance cell settlement on the surface or be used to direct tissue generation on the biomaterial interface. PMID- 15276365 TI - Rheological characterisation of primary and binary interactive bioadhesive gels composed of cellulose derivatives designed as ophthalmic viscosurgical devices. AB - In this study the formulation and rheological characterisation of novel candidate ophthalmic viscosurgical devices (OVD) based on binary interactive polymer gels is described. Primary systems containing either hydroxyethylcellulose (HEC) or sodium carboxymethylcellulose (NaCMC) or binary interactive gels composed of HEC and NaCMC were manufactured. Rheological characterisation was performed using texture profile analysis and oscillatory rheometry. All formulations exhibited pseudoplastic flow. Systems composed of HEC or HEC and NaCMC behaved as gels (G' > G") over the range of oscillatory frequencies whereas systems composed of NaCMC were primarily elastoviscous. Increasing the polymer concentration in all systems increased the compressional rheological properties (hardness, compressibility), zero frequency viscosity (derived from the Cross model) and the viscoelastic properties (G', G" and eta'). Rheological synergy was observed in the binary gels and was indicative of interaction between the parent polymers. Importantly, the range of rheological properties offered by the binary mixtures was greater than those exhibited by the primary systems. The binary systems described in this study possessed viscoelastic properties and steady-state viscosities that were similar to commercially available systems and would therefore be appropriate for the maintenance of the ocular space. The acceptable compressional rheological and pseudoplastic properties of these systems would facilitate administration into the eye using a syringe. Additionally and uniquely, the excellent adhesive properties of the binary interactive gels would suggest an ability to interact with the corneal endothelium that would offer protection during phacoemulsification. Based on the described rheological properties it is suggested that binary gels composed of mass ratios of HEC to NaCMC of either 3.6: 2.4 or 2.4: 3.6 would be acceptable as OVD and would uniquely offer duality of function. PMID- 15276366 TI - Is it churlish to criticise Bush over his spending on AIDS? PMID- 15276367 TI - The practicalities of keeping clean. PMID- 15276368 TI - Lessons from MATCH for future randomised trials in secondary prevention of stroke. PMID- 15276369 TI - Teaching safe sex in English schools. PMID- 15276370 TI - Insights into the autoimmune nature of aplastic anaemia. PMID- 15276371 TI - Meningococcal conjugate vaccine in the UK: an update. PMID- 15276372 TI - The McMaster-Lancet health and peace conferences. PMID- 15276373 TI - Targeting energy metabolism in Huntington's disease. PMID- 15276374 TI - Hyperventilation during cardiac arrest. PMID- 15276375 TI - Cannabis for medical purposes: cultivating science, weeding out the fiction. PMID- 15276376 TI - Keywords in the history of medicine: vitamin. PMID- 15276378 TI - XV International AIDS Conference: an unkept promise of access for all. PMID- 15276379 TI - IGF-1, IGFBP-3, and cancer risk. PMID- 15276380 TI - IGF-1, IGFBP-3, and cancer risk. PMID- 15276383 TI - Disclosure of non-paternity. PMID- 15276384 TI - Tale of the dural tail. PMID- 15276386 TI - Anniversary of rubella epidemic. PMID- 15276387 TI - Why is measles still endemic in Japan? PMID- 15276388 TI - First-line and second-line antiretroviral therapy. PMID- 15276391 TI - Clinical trials register. PMID- 15276389 TI - First-line and second-line antiretroviral therapy. PMID- 15276392 TI - Aspirin and clopidogrel compared with clopidogrel alone after recent ischaemic stroke or transient ischaemic attack in high-risk patients (MATCH): randomised, double-blind, placebo-controlled trial. AB - BACKGROUND: Clopidogrel was superior to aspirin in patients with previous manifestations of atherothrombotic disease in the CAPRIE study and its benefit was amplified in some high-risk subgroups of patients. We aimed to assess whether addition of aspirin to clopidogrel could have a greater benefit than clopidogrel alone in prevention of vascular events with potentially higher bleeding risk. METHODS: We did a randomised, double-blind, placebo-controlled trial to compare aspirin (75 mg/day) with placebo in 7599 high-risk patients with recent ischaemic stroke or transient ischaemic attack and at least one additional vascular risk factor who were already receiving clopidogrel 75 mg/day. Duration of treatment and follow-up was 18 months. The primary endpoint was a composite of ischaemic stroke, myocardial infarction, vascular death, or rehospitalisation for acute ischaemia (including rehospitalisation for transient ischaemic attack, angina pectoris, or worsening of peripheral arterial disease). Analysis was by intention to treat, using logrank test and a Cox's proportional-hazards model. FINDINGS: 596 (15.7%) patients reached the primary endpoint in the group receiving aspirin and clopidogrel compared with 636 (16.7%) in the clopidogrel alone group (relative risk reduction 6.4%, [95% CI -4.6 to 16.3]; absolute risk reduction 1% [-0.6 to 2.7]). Life-threatening bleedings were higher in the group receiving aspirin and clopidogrel versus clopidogrel alone (96 [2.6%] vs 49 [1.3%]; absolute risk increase 1.3% [95% CI 0.6 to 1.9]). Major bleedings were also increased in the group receiving aspirin and clopidogrel but no difference was recorded in mortality. INTERPRETATION: Adding aspirin to clopidogrel in high-risk patients with recent ischaemic stroke or transient ischaemic attack is associated with a non-significant difference in reducing major vascular events. However, the risk of life-threatening or major bleeding is increased by the addition of aspirin. PMID- 15276393 TI - Pupil-led sex education in England (RIPPLE study): cluster-randomised intervention trial. AB - BACKGROUND: Improvement of sex education in schools is a key part of the UK government's strategy to reduce teenage pregnancy in England. We examined the effectiveness of one form of peer-led sex education in a school-based randomised trial of over 8000 pupils. METHODS: 29 schools were randomised to either peer-led sex education (intervention) or to continue their usual teacher-led sex education (control). In intervention schools, peer educators aged 16-17 years delivered three sessions of sex education to 13-14 year-old pupils from the same schools. Primary outcome was unprotected (without condom) first heterosexual intercourse by age 16 years. Analysis was by intention to treat. FINDINGS: By age 16 years, significantly fewer girls reported intercourse in the peer-led arm than in the control arm, but proportions were similar for boys. The proportions of pupils reporting unprotected first sex did not differ for girls (8.4% intervention vs 8.3% control) or for boys (6.2% vs 4.7%). Stratified estimates of the difference between arms were -0.4% (95% CI -3.7% to 2.8%, p=0.79) for girls and -1.4% (-4.4% to 1.6%, p=0.36) for boys. At follow-up (mean age 16.0 years [SD 0.32]), girls in the intervention arm reported fewer unintended pregnancies, although the difference was borderline (2.3% vs 3.3%, p=0.07). Girls and boys were more satisfied with peer-led than teacher-led sex education, but 57% of girls and 32% of boys wanted sex education in single-sex groups. INTERPRETATION: Peer-led sex education was effective in some ways, but broader strategies are needed to improve young people's sexual health. The role of single-sex sessions should be investigated further. PMID- 15276394 TI - Long-term effects of darusentan on left-ventricular remodelling and clinical outcomes in the EndothelinA Receptor Antagonist Trial in Heart Failure (EARTH): randomised, double-blind, placebo-controlled trial. AB - BACKGROUND: Endothelin-receptor blockade provides haemodynamic benefit in experimental and clinical heart failure. We aimed to measure the effects of long term endothelin-blockade on left-ventricular (LV) remodelling and clinical outcomes in patients with chronic heart failure. METHODS: 642 patients with chronic heart failure were assigned the oral endothelin(A)-antagonist darusentan at 10, 25, 50, 100, or 300 mg daily or placebo for 24 weeks in addition to standard therapy in a randomised, double-blind, placebo-controlled trial. In the 50-300 mg groups, darusentan was uptitrated over 6 weeks. Primary endpoint was change in LV end-systolic volume (LVESV) at 24 weeks from baseline, measured by MRI. All patients for whom assessable MRI scans were available at baseline and follow-up were included in the analysis. FINDINGS: Darusentan was well tolerated. LVESV could be assessed in 485 (76%) patients with paired MRI data at baseline and 6 months. The change in LVESV was not significantly different from that with placebo at any dose (mean difference from placebo 1.27 mL [95% CI -9.9 to 12.4] with 10 mg dose, -1.84 mL [-13.0 to 9.3] with 25 mg, -5.68 mL [-16.9 to 5.6] with 50 mg, -4.05 mL [-15.5 to 7.4] with 100 mg, and -4.34 mL [-15.7 to 7.0] with 300 mg). Heart failure worsened in 71 (11.1%) patients, and 30 (4.7%) died during the study with no difference between groups. INTERPRETATION: Endothelin(A) blockade with darusentan did not improve cardiac remodelling or clinical symptoms or outcomes in patients with chronic heart failure receiving an angiotensin converting-enzyme inhibitor, beta blocker, or aldosterone antagonist. Thus, endothelin(A) blockade is unlikely to be useful as an add-on treatment in such patients. PMID- 15276395 TI - In-vivo dominant immune responses in aplastic anaemia: molecular tracking of putatively pathogenetic T-cell clones by TCR beta-CDR3 sequencing. AB - BACKGROUND: Aplastic anaemia is a bone-marrow-failure syndrome characterised by low blood-cell counts and fatty bone marrow. In most cases, no obvious aetiological factor can be identified. However, clinical responses to immunosuppression strongly suggest an immune pathophysiology. METHODS: To test the hypothesis that aplastic anaemia results from antigen-specific lymphocyte attack against haemopoietic tissue, we analysed effector immunity, seeking especially dominant specific T-cell responses. Blood samples from 54 patients with aplastic anaemia were subjected to flow cytometry to define T-cell-receptor Vbeta-chain usage and expansion of particular Vbeta subsets. We measured the size distribution of the complementarity-determining region 3 (CDR3) for expanded Vbeta subsets, then cloned and sequenced skewed, oligoclonal, or monoclonal peaks. FINDINGS: Expanded Vbeta subsets were identified in almost all the patients. Over-represented Vbeta subsets from CD8-positive cells showed oligoclonal or monoclonal CDR3 size patterns. The CDR3 sequence repertoire in aplastic anaemia showed much redundancy compared with healthy donors. We identified patient-specific putative pathogenetic clonotypes that were not detectable in controls. In selected patients who were assessed longitudinally, these clonotypes were quantitatively related to disease activity. Selective killing of autologous haemopoietic progenitors by the Vbeta-specific lymphocyte population was shown in one patient. These apparently pathogenetic CDR3 sequences showed homology between individuals, suggesting a role for a "semi-public" immune response in the pathophysiology of aplastic anaemia. INTERPRETATION: In-vivo dominant clonal immune response can be identified in many patients with aplastic anaemia, which is evidence for an underlying antigen-driven immune process. Longitudinal tracking by molecular techniques could inform individual clinical decisions and the development of new treatments in autoimmune diseases. RELEVANCE TO PRACTICE: Although the target of the aberrant immune response is the haemopoietic stem cell, the triggering antigens remain unknown. We combined cell phenotypic, molecular biology, and functional analyses to study the effector arm of immunity in an attempt to establish an immune pathophysiology. Clinical application of such a model could broadly extend to other autoimmune diseases. PMID- 15276396 TI - Effectiveness of meningococcal serogroup C conjugate vaccine 4 years after introduction. AB - The meningococcal serogroup C conjugate (MCC) vaccine programme in England has successfully controlled the incidence of serogroup C disease, as a result of high short-term vaccine effectiveness and substantial herd immunity. However, the long term effectiveness of the vaccine remains unknown. We assessed surveillance data from the 4 years since introduction of the programme. Vaccine effectiveness remained high in children vaccinated in the catch-up campaign (aged 5 months to 18 years). However, for children vaccinated in the routine infant immunisation programme, the effectiveness of the MCC vaccine fell to low levels after only 1 year. The number of individuals in these cohorts remains low, but alternative routine immunisation schedules should be considered to ensure high levels of protection are sustained. PMID- 15276397 TI - Caesarean scar endometrioma. PMID- 15276398 TI - Osteomyelitis. AB - Bone and joint infections are painful for patients and frustrating for both them and their doctors. The high success rates of antimicrobial therapy in most infectious diseases have not yet been achieved in bone and joint infections owing to the physiological and anatomical characteristics of bone. The key to successful management is early diagnosis, including bone sampling for microbiological and pathological examination to allow targeted and long-lasting antimicrobial therapy. The various types of osteomyelitis require differing medical and surgical therapeutic strategies. These types include, in order of decreasing frequency: osteomyelitis secondary to a contiguous focus of infection (after trauma, surgery, or insertion of a joint prosthesis); that secondary to vascular insufficiency (in diabetic foot infections); or that of haematogenous origin. Chronic osteomyelitis is associated with avascular necrosis of bone and formation of sequestrum (dead bone), and surgical debridement is necessary for cure in addition to antibiotic therapy. By contrast, acute osteomyelitis can respond to antibiotics alone. Generally, a multidisciplinary approach is required for success, involving expertise in orthopaedic surgery, infectious diseases, and plastic surgery, as well as vascular surgery, particularly for complex cases with soft-tissue loss. PMID- 15276399 TI - "Neglected" diseases but unrecognised successes--challenges and opportunities for infectious disease control. PMID- 15276400 TI - Peace through health: key concepts. PMID- 15276401 TI - Intuition, evidence, and safety. PMID- 15276402 TI - The world through tinted glasses. PMID- 15276403 TI - Chronic health effects in people exposed to arsenic via the drinking water: dose response relationships in review. AB - Chronic arsenic (As) poisoning has become a worldwide public health issue. Most human As exposure occurs from consumption of drinking water containing high amounts of inorganic As (iAs). In this paper, epidemiological studies conducted on the dose-response relationships between iAs exposure via the drinking water and related adverse health effects are reviewed. Before the review, the methods for evaluation of the individual As exposure are summarized and classified into two types, that is, the methods depending on As concentration of the drinking water and the methods depending on biological monitoring for As exposure; certain methods may be applied as optimum As exposure indexes to study dose-response relationship based on various As exposure situation. Chronic effects of iAs exposure via drinking water include skin lesions, neurological effects, hypertension, peripheral vascular disease, cardiovascular disease, respiratory disease, diabetes mellitus, and malignancies including skin cancer. The skin is quite sensitive to arsenic, and skin lesions are some of the most common and earliest nonmalignant effects related to chronic As exposure. The increase of prevalence in the skin lesions has been observed even at the exposure levels in the range of 0.005-0.01 mg/l As in drinking waters. Skin, lung, bladder, kidney, liver, and uterus are considered as sites As-induced malignancies, and the skin is though to be perhaps the most sensitive site. Prospective studies in large area of endemic As poisoning, like Bangladesh or China, where the rate of malignancies is expected to increase within the next several decades, will help to clarify the dose-response relationship between As exposure levels and adverse health effects with enhanced accuracy. PMID- 15276404 TI - Arsenic toxicity at low doses: epidemiological and mode of action considerations. AB - Current approaches to risk assessment typically assume a linear dose-response for mutagenic compounds that directly interact with DNA or when the carcinogenic mechanism is unknown. Because the mode of action of arsenic-induced carcinogenesis is not well established, recent dose-response assessments for arsenic have assumed linearity at low doses despite evidence that arsenic is not a direct-acting mutagen. Several modes of action, including generation of oxidative stress, perturbation of DNA methylation patterns, inhibition of DNA repair, and modulation of signal transduction pathways, have been proposed to characterize arsenic's toxicity. It is probable that these mechanisms do not act in isolation, but overlap, and contribute to the complex nature of arsenic induced carcinogenesis. All of the proposed mechanisms are likely to be nonlinear at low does. Furthermore, studies of populations outside the US exposed to arsenic in drinking water show increases in cancer only at relatively high concentrations, that is, concentrations in drinking water of several hundred micrograms per liter (microg/l). Studies in the US of populations exposed to average concentrations in drinking water up to about 190 microg/l do not provide evidence of increased cancer. Consideration of arsenic's plausible mechanisms and evidence from epidemiological studies support the use of nonlinear methods, either via biologically based modeling or use of a margin-of-exposure analysis, to characterize arsenic risks. PMID- 15276405 TI - Arsenic contamination and arsenicosis in China. AB - Arsenicosis is a serious environmental chemical disease in China mainly caused by drinking water from pump wells contaminated by high levels of arsenic. Chronic exposure of humans to high concentrations of arsenic in drinking water is associated with skin lesions, peripheral vascular disease, hypertension, blackfoot disease, and high risk of cancers. Lead by the Ministry of Health of China, we carried out a research about arsenicosis in China recently. Areas contaminated with arsenic from drinking water are determined by 10% pump well water sample method while areas from burning coal are determined by existing data. Two epidemic areas of Shanxi Province and Inner Mongolia are investigated for the distribution of pump wells containing high arsenic. Well water in all the investigated villages of Shanxi Province showed polluted by high arsenic, and the average rate of unsafe pump well water is 52%. In Inner Mongolia, the high percentage of pump wells containing elevated arsenic is found only in a few villages. The average rate of unsafe pump well water is 11%. From our research, we find that new endemic areas are continuously emerging in China. Up to now, epidemic areas of arsenicosis mainly involve eight provinces and 37 counties in China. In the affected areas, the discovery of wells and coal with high levels of arsenic is continuing sporadically, and a similar scattered distribution pattern of patients is also being observed. PMID- 15276406 TI - Water intake in an Asian population living in arsenic-contaminated area. AB - Exposure evaluation is an indispensable step for the risk assessment of chronic arsenic toxicity. The amount of water intake, which consists of the base of exposure calculation, has been lacking for the arsenic-affected populations in the developing countries. Thus, the purposes of the present study were (1) to estimate the water intake in such population, and (2) to estimate the relative importance of water versus food as the source of arsenic exposure. Adult males and females (n = 19 for each) were selected from two rural Bangladeshi communities that entirely depended on tubewells for their water supply. Their water intake was measured by two methods, a 24-h self-report and an interview with frequent visits. Results of the two methods generally agreed with each other in terms of correlation and the absolute intakes. Mean water intake obtained by the self-report method was found to be around 3 l/day with substantial individual variation (the maximum = 6 l/day), no sex difference, and a significant between community difference. The calculation for total arsenic exposure demonstrated that there was no sex difference in arsenic exposure except when the exposure was mainly from food and thus relatively low. Although these results need to be further confirmed under various environmental settings, these results suggested that (1) the sex difference in the manifestation of arsenic toxicity previously observed in this area should be related with factors other than exposure level and that (2) the risks associated with low arsenic concentrations of groundwater should be carefully interpreted because food may be providing additional burden of arsenic. PMID- 15276407 TI - Nitric oxide and superoxide anion production in monocytes from children exposed to arsenic and lead in region Lagunera, Mexico. AB - We evaluated in Mexican children environmentally exposed to arsenic and lead monocyte nitric oxide (NO) and superoxide anion production in response to direct activation with interferon-gamma (IFN-gamma) + lipopolysaccharide (LPS). The integrity of Th1-regulated cellular immune response when monocytes were indirectly activated was also evaluated. Most children lived near a primary lead smelter. Lead and arsenic contamination in soil and dust by far exceeded background levels. As levels in water were between 10 and 30 ppb. Most children (93%) had urinary arsenic (AsU) concentrations above 50 microg/l (range 16.75 465.75) and 65% had lead blood levels (PbB) above 10 microg/dl (range 3.47 49.19). Multivariate analyses showed that NO production in monocytes activated indirectly was negatively associated with both PbB and AsU. Superoxide production in directly activated monocytes was negatively associated with AsU but positively associated with PbB. The models including the interaction term for AsU and PbB suggested the possibility of a negative interaction for NO production and a positive interaction for superoxide. There were indications of differential gender-based associations, NO production in indirectly activated monocytes obtained from girls was negatively associated with AsU but not with PbB. Superoxide production was positively associated with PbB in both directly and indirectly activated monocytes from boys but the latter was negatively associated with AsU. These effects are consistent with immune system abnormalities observed in human populations exposed to Pb or As. Further studies in larger populations are required to characterize As and Pb interactions and the mechanism(s) underlying the observed effects. PMID- 15276408 TI - Evaluation of DNA damage in patients with arsenic poisoning: urinary 8 hydroxydeoxyguanine. AB - The relationship between arsenic exposure and DNA damage in patients with acute or chronic arsenic poisoning was analyzed. Urinary 8-hydroxydeoxyguanine (8-OHdG) concentrations were measured as an indication of oxidative DNA damage. A remarkable increase in 8-OHdG in the urine was observed in 60% of 52 patients with acute arsenic poisoning from the accidental oral intake of the arsenic trioxide. This was two- to threefold higher than levels in normal healthy subjects (n = 248). There was a clear relationship between arsenic concentrations in urine after acute poisoning and elevated levels of 8-OHdG. Levels of urinary 8 OHdG returned to normal within 180 days after the acute arsenic poisoning event. In patients chronically poisoned by the consumption of well water with elevated levels of arsenate [As(V)], elevated 8-OHdG concentrations in urine were also observed. A significant correlation between the 8-OHdG levels and arsenic levels in the urine was observed in 82 patients with chronic poisoning. Thus, evidence of oxidative DNA damage occurred in acute arsenic poisoning by arsenite [As(III)] and in chronic arsenic poisoning by As(V). In chronic poisoning patients provided low-arsenic drinking water, evidence of DNA damage subsided between 9 months and 1 year after the high levels of arsenic intake were reduced. The initial level of arsenic exposure appeared to dictate the length of this recovery period. These data indicate that some aspects of chronic and acute arsenic poisoning may be reversible with the cessation of exposure. This knowledge may contribute to our understanding of the risk elevation from arsenic carcinogenesis and perhaps be used in a prospective fashion to assess individual risk. PMID- 15276409 TI - Arsenic speciation in human urine: are we all the same? AB - We studied the arsenic speciation in human urine samples by using high performance liquid chromatography-inductively coupled plasma-mass spectrometry (HPLC-ICP-MS). We investigated the arsenic speciation in the urine collected from nine volunteers during a 3-day period after a meal of blue mussels, Mytilus edulis. We also studied the effect of cooking on the arsenic speciation. Arsenobetaine and dimethylarsinic acid (DMAA) were the major arsenic metabolites found in the urine samples. Significant amounts of unknown metabolites were also detected. The excretion patterns of arsenic from individuals were generally similar except for two subjects. One of whom excreted high amounts of arsenobetaine even though no arsenic-rich food was eaten for 3 days before the experiment. The results reveal that we need a better understanding of the metabolism of arsenic compounds by human. PMID- 15276410 TI - Speciation of arsenic in biological samples. AB - Speciation of arsenicals in biological samples is an essential tool to gain insight into its distribution in tissues and its species-specific toxicity to target organs. Biological samples (urine, hair, fingernail) examined in the present study were collected from 41 people of West Bengal, India, who were drinking arsenic (As)-contaminated water, whereas 25 blood and urine samples were collected from a population who stopped drinking As contaminated water 2 years before the blood collection. Speciation of arsenicals in urine, water-methanol extract of freeze-dried red blood cells (RBCs), trichloroacetic acid treated plasma, and water extract of hair and fingernail was carried out by high performance liquid chromatography (HPLC)-inductively coupled argon plasma mass spectrometry (ICP MS). Urine contained arsenobetaine (AsB, 1.0%), arsenite (iAs(III), 11.3), arsenate (iAs(V), 10.1), monomethylarsonous acid (MMA(III), 6.6), monomethylarsonic acid (MMA(V), 10.5), dimethylarsinous acid (DMA(III), 13.0), and dimethylarsinic acid (DMA(V), 47.5); fingernail contained iAs(III) (62.4%), iAs(V) (20.2), MMA(V) (5.7), DMA(III) (8.9), and DMA(V) (2.8); hair contained iAs(III) (58.9%), iAs(V) (34.8), MMA(V) (2.9), and DMA(V) (3.4); RBCs contained AsB (22.5%) and DMA(V) (77.5); and blood plasma contained AsB (16.7%), iAs(III) (21.1), MMA(V) (27.1), and DMA(V) (35.1). MMA(III), DMA(III), and iAs(V) were not found in any plasma and RBCs samples, but urine contained all of them. Arsenic in urine, fingernails, and hair are positively correlated with water As, suggesting that any of these measurements could be considered as a biomarker to As exposure. Status of urine and exogenous contamination of hair urgently need speciation of As in these samples, but speciation of As in nail is related to its total As (tAs) concentration. Therefore, total As concentrations of nails could be considered as biomarker to As exposure in the endemic areas. PMID- 15276411 TI - Elucidating the pathway for arsenic methylation. AB - Although biomethylation of arsenic has been studied for more than a century, unequivocal demonstration of the methylation of inorganic arsenic by humans occurred only about 30 years ago. Because methylation of inorganic arsenic activates it to more reactive and toxic forms, elucidating the pathway for the methylation of this metalloid is a topic of considerable importance. Understanding arsenic metabolism is of public health concern as millions of people chronically consume drinking water that contains high concentrations of inorganic arsenic. Hence, the focus of our research has been to elucidate the molecular basis of the steps in the pathway that leads from inorganic arsenic to methylated and dimethylated arsenicals. Here we describe a new S adenosylmethionine (AdoMet)-dependent methyltransferase from rat liver cytosol that catalyzes the conversion of arsenite to methylated and dimethylated species. This 42-kDa protein has sequence motifs common to many non-nucleic acid methyltransferases and is closely related to methyltransferases of previously unknown function that have been identified by conceptual translations of cyt19 genes of mouse and human genomes. Hence, we designate rat liver arsenic methyltransferase as cyt19 and suggest that orthologous cyt19 genes encode an arsenic methyltransferase in the mouse and human genomes. Our studies with recombinant rat cyt19 find that, in the presence of an exogenous or a physiological reductant, this protein can catalyze the entire sequence of reactions that convert arsenite to methylated metabolites. A scheme linking cyt19 and thioredoxin-thioredoxin reductase in the methylation and reduction of arsenicals is proposed. PMID- 15276412 TI - A review of the enzymology of arsenic metabolism and a new potential role of hydrogen peroxide in the detoxication of the trivalent arsenic species. AB - This laboratory has studied the enzymology involved in the biotransformation of inorganic arsenic to dimethylarsinous acid (DMA(III)) and in human studies established that monomethylarsonous acid (MMA(III)) and DMA(III) appear in urine of people chronically exposed to arsenic. It appears that only two proteins are required for inorganic arsenic biotransformation in the human, namely, monomethylarsonic acid (MMA(V)) reductase and arsenic methyltransferase. MMA(V) reductase and the unique glutathione transferase omega (hGST-O) are identical proteins. Arsenicals with a +3 oxidation state are more toxic than the +5 species. While methylation of arsenite, MMA(III), and DMA(III) produces less toxic +5 oxidation arsenic species containing an additional methyl group such as MMA(V), dimethylarsinic acid (DMA(V)), and TMAO, a new mechanism involving hydrogen peroxide for detoxifying arsenite, MMA(III), and DMA(III) is proposed based on in vitro experiments. PMID- 15276413 TI - Distributions and chemical forms of arsenic after intravenous administration of dimethylarsinic and monomethylarsonic acids to rats. AB - The observed toxicity of arsenic is highly dependent on animal species and differences in metabolism. Rats are one of the most tolerant species, and the metabolic pathway is quite different in some aspects from those of other mammals. The distinct metabolic pathway including the preferential accumulation in red blood cells (RBCs) has been explained, whereby allowing an effective use of rats as an animal model for the arsenic metabolism. In the present study, distributions of arsenic among organs/tissues/body fluids and their chemical forms were studied after intravenous injection of arsenic in the forms of dimethylarsinic (DMA(V)) and monomethylarsonic acids (MMA(V)) to rats. DMA(V) and MMA(V) were mostly excreted into urine immediately after the injection as the intact forms, and both forms were taken up less effectively by organs/tissues than arsenite. The methylated arsenics distributed in organs/tissues were excreted directly into urine and excreted before being redistributed in RBCs. DMA(V) and MMA(V) taken up by the liver were transformed to metabolites not yet identified, accumulated transiently in the liver, and then they disappeared from the liver. The unidentified metabolites were assumed to be transformed from dimethylarsinic acid (DMA(III)) following the consecutive metabolic reactions [MMA(V) --> monomethylarsonous acid (MMA(III)) --> DMA(V) --> DMA(III)]. The unidentified metabolites were excreted not into the bile but into the bloodstream. Injections of DMA(V) and MMA(V) induced a biliary excretion of arsenic but only at 0.2-0.3% of the dose, the arsenic in the bile being their intact free forms. PMID- 15276414 TI - Microbial metabolite of dimethylarsinic acid is highly toxic and genotoxic. AB - Dimethylarsinic acid [DMA, (CH(3))(2)AsO(OH)] causes cancer in the urinary bladder of rats. However, its mechanism of cancer or the ultimate carcinogenic form is not yet known. Rats administered dimethylarsinic acid excrete three unknown arsenic compounds (termed M-1, M-2, and M-3) in urine or feces, and these compounds are presumed to be produced by intestinal bacteria. Escherichia coli A3 6 isolated from a rat yielded two unknown arsenic compounds (M-2 and M-3) from dimethylarsinic acid and M-1 from trimethylarsine oxide (TMAO) in the presence of cysteine (Cys). Contents of M-2 and M-3 varied with cysteine concentration. The cytotoxicity and genotoxicity of the bacteria-free solution of dimethylarsinic acid or trimethylarsine oxide metabolized by E. coli A3-6 were studied using V79 cells. Dimethylarsinic acid (1 mM) metabolized by E. coli A3-6 in the presence of cysteine (1 mM) was highly cytotoxic (50% survival reduction concentration; 2.1 microM As) in V79 cells, and the toxic substance appeared to be M-2. The metabolite solution (at 2.5-10 microM total As) induced c-mitosis and tetraploids, and caused mitotic arrest, since it increased mitotic cells at the cytotoxic dose. The metabolite solution also significantly increased sister chromatid exchange (SCE) and chromosomal aberrations, most of which were chromatid gaps and chromatid breaks. A3-6 converted 96.1% of trimethylarsine oxide to M-1 in the presence of cysteine. This metabolite solution did not exhibit cytotoxicity or genotoxicity. The reported M-2 concentration in urine of rats administered levels of DMA via drinking water known to cause bladder tumors was sufficient to exhibit cytotoxic and genotoxic effects in urinary bladder. Thus, we hypothesize that intestinal bacteria play an important role in carcinogenicity of dimethylarsinic acid. PMID- 15276415 TI - Role of glutathione in dimethylarsinic acid-induced apoptosis. AB - Inorganic arsenicals are clearly toxicants and carcinogens in humans. In mammals, including humans, inorganic arsenicals often undergo methylation, forming compounds such as dimethylarsinic acid (DMAs(V)). Recent evidence indicates that DMAs(V) is a complete carcinogen in rodents although evidence for inorganic arsenicals as carcinogens in rodents remains equivocal. Thus, we studied the molecular mechanisms of in vitro cytolethality of DMAs(V) using a rat liver epithelial cell line (TRL 1215). DMAs(V) selectively induced apoptosis in TRL 1215 cells; its LC(50) value after 48 h exposure was 4.5 mM. The addition of a glutathione synthase inhibitor, L-buthionine-[S,R]-sulfoximine (BSO), actually decreased DMAs(V)-induced apoptosis. DMAs(V) exposure temporarily decreased cellular reduced glutathione (GSH) levels and enhanced cellular glutathione S transferase (GST) activity from 6 h after the exposure when the cells were still alive. Also, DMAs(V) exposure activated cellular caspase 3 activity with a peak at 18 h after the exposure when apoptosis began, and BSO treatment completely inhibited this enzyme activity. The additions of inhibitors of caspase 3, caspase 8, and caspase 9 significantly reduced DMAs(V)-induced apoptosis. Taken together, these data indicate that cellular GSH was required for DMAs(V)-induced apoptosis to occur, and activation of cellular caspases after conjugation of DMAs(V) with cellular GSH appears to be of mechanistic significance. Further research will be required to determine the role of intracellular GSH and methylation in the toxicity of arsenicals in chronic arsenic poisoning or in cases where arsenicals are used as chemotherapeutics. PMID- 15276416 TI - Understanding arsenic carcinogenicity by the use of animal models. AB - Although numerous epidemiological studies have indicated that human arsenic exposure is associated with increased incidences of bladder, liver, skin, and lung cancers, limited attempts have been made to understand mechanisms of carcinogenicity using animal models. Dimethylarsinic acid (DMA), an organic arsenic compound, is a major metabolite of ingested inorganic arsenics in mammals. Recent in vitro studies have proven DMA to be a potent clastogenic agent, capable of inducing DNA damage including double strand breaks and cross link formation. In our attempts to clarify DMA carcinogenicity, we have recently shown carcinogenic effects of DMA and its related metabolites using various experimental protocols in rats and mice: (1) a multi-organ promotion bioassay in rats; (2) a two-stage promotion bioassay by DMA of rat urinary bladder and liver carcinogenesis; (3) a 2-year carcinogenicity test of DMA in rats; (4) studies on the effects of DMA on lung carcinogenesis in rats; (5) promotion of skin carcinogenesis by DMA in keratin (K6)/ornithine decarboxylase (ODC) transgenic mice; (6) carcinogenicity of DMA in p53(+/-) knockout and Mmh/8-OXOG-DNA glycolase (OGG1) mutant mice; (7) promoting effects of DMA and related organic arsenicals in rat liver; (8) promoting effects of DMA and related organic arsenicals in a rat multi-organ carcinogenesis test; and (9) 2-year carcinogenicity tests of monomethylarsonic acid (MMA) and trimethylarsine oxide (TMAO) in rats. The results revealed that the adverse effects of arsenic occurred either by promoting and initiating carcinogenesis. These data, as covered in the present review, suggest that several mechanisms may be involved in arsenic carcinogenesis. PMID- 15276417 TI - Animal models for arsenic carcinogenesis: inorganic arsenic is a transplacental carcinogen in mice. AB - Inorganic arsenic is a known human carcinogen causing tumors of the skin, urinary bladder, lung, liver, kidney, and possibly other organs. However, the animal models for inorganic arsenic carcinogenesis have been limited and development has been problematic. Gestation is often a period of high sensitivity to carcinogenesis so we investigated inorganic arsenite as a transplacental carcinogen in mice. Pregnant C3H mice were exposed to sodium arsenite (0, 42.5, and 85 ppm as arsenic) in the drinking water for a brief period during gestation (from gestation day 8 to 18), with no further arsenic exposure or other treatments. The offsprings were monitored up to 90 weeks. Transplacental inorganic arsenic exposure produced a dose-dependent induction of tumors in the liver, adrenal, lung, and ovary in the offsprings after they had reached adulthood. This included hepatocellular carcinoma (HCC), a tumor associated with arsenic exposure in humans. These tumors occurred when mice became adults in the absence of any other treatments and well after arsenic exposure had ended. Genomic analysis of liver tumors and tumor-surrounding tissues revealed several patterns of aberrant gene expression associated with transplacental arsenic carcinogenesis. This animal model demonstrated that inorganic arsenic could act as a "complete" transplacental carcinogen in mice. In addition, other important animal models for inorganic arsenic as a skin tumor co-promoter or as a co carcinogen are discussed. The development of these animal models should advance our understanding of the mechanisms of inorganic arsenic carcinogenesis. PMID- 15276418 TI - The role of active arsenic species produced by metabolic reduction of dimethylarsinic acid in genotoxicity and tumorigenesis. AB - In recent research of arsenic carcinogenesis, many researchers have directed their attention to methylated metabolites of inorganic arsenics. Because of its high cytotoxicity and genotoxicity, trivalent dimethylated arsenic, which can be produced by the metabolic reduction of dimethylarsinic acid (DMA), has attracted considerable attention from the standpoint of arsenic carcinogenesis. In the present paper, we examined trivalent dimethylated arsenic and its further metabolites for their chemical properties and biological behavior such as genotoxicity and tumorigenicity. Our in vitro and in vivo experiments suggested that the formation of cis-thymine glycol in DNA was induced via the production of dimethylated arsenic peroxide by the reaction of trivalent dimethylated arsenic with molecular oxygen, but not via the production of common reactive oxygen species (ROS; superoxide, hydrogen peroxide, hydroxyl radical, etc.). Thus, dimethylated arsenic peroxide may be the main species responsible for the tumor promotion in skin tumorigenesis induced by exposure to DMA. Free radical species, such as dimethylarsenic radical [(CH(3))(2)As.] and dimethylarsenic peroxy radical [(CH(3))(2)AsOO.], that are produced by the reaction of molecular oxygen and dimethylarsine [(CH(3))(2)AsH], which is probably a further reductive metabolite of trivalent dimethylated arsenic, may be main agents for initiation in mouse lung tumorigenesis. PMID- 15276419 TI - Evidence that arsenite acts as a cocarcinogen in skin cancer. AB - Inorganic arsenic (arsenite and arsenate) in drinking water has been associated with skin cancers in several countries such as Taiwan, Chile, Argentina, Bangladesh, and Mexico. This association has not been established in the United States. In addition, inorganic arsenic alone in drinking water does not cause skin cancers in animals. We recently showed that concentrations as low as 1.25 mg/l sodium arsenite were able to enhance the tumorigenicity of solar UV irradiation in mice. The tumors were almost all squamous cell carcinomas (SCCs). These data suggest that arsenic in drinking water may need a carcinogenic partner, such as sunlight, in the induction of skin cancers. Arsenite may enhance tumorigenicity via effects on DNA repair and DNA damage-induced cell cycle effects, leading to genomic instability. Others have found that dimethlyarsinic acid (DMA), a metabolite of arsenite, can induce bladder cancers at high concentrations in drinking water. In those experiments, skin cancers were not produced. Taken together, these data suggest that arsenite (or possibly an earlier metabolite), and not DMA, is responsible for the skin cancers, but a second genotoxic agent may be a requirement. The differences between the US and the other arsenic-exposed populations with regard to skin cancers might be explained by the lower levels of arsenic in the US, less sun exposure, better nutrition, or perhaps genetic susceptibility differences. PMID- 15276420 TI - Toxicity of indium arsenide, gallium arsenide, and aluminium gallium arsenide. AB - Gallium arsenide (GaAs), indium arsenide (InAs), and aluminium gallium arsenide (AlGaAs) are semiconductor applications. Although the increased use of these materials has raised concerns about occupational exposure to them, there is little information regarding the adverse health effects to workers arising from exposure to these particles. However, available data indicate these semiconductor materials can be toxic in animals. Although acute and chronic toxicity of the lung, reproductive organs, and kidney are associated with exposure to these semiconductor materials, in particular, chronic toxicity should pay much attention owing to low solubility of these materials. Between InAs, GaAs, and AlGaAs, InAs was the most toxic material to the lung followed by GaAs and AlGaAs when given intratracheally. This was probably due to difference in the toxicity of the counter-element of arsenic in semiconductor materials, such as indium, gallium, or aluminium, and not arsenic itself. It appeared that indium, gallium, or aluminium was toxic when released from the particles, though the physical character of the particles also contributes to toxic effect. Although there is no evidence of the carcinogenicity of InAs or AlGaAs, GaAs and InP, which are semiconductor materials, showed the clear evidence of carcinogenic potential. It is necessary to pay much greater attention to the human exposure of semiconductor materials. PMID- 15276421 TI - Effects of arsenite on UROtsa cells: low-level arsenite causes accumulation of ubiquitinated proteins that is enhanced by reduction in cellular glutathione levels. AB - Chronic arsenic exposure increases risk for the development of diabetes, vascular disease, and cancers of the skin, lung, kidney, and bladder. This study investigates the effects of arsenite [As(III)] on human urothelial cells (UROtsa). As(III) toxicity was determined by exposing confluent UROtsa cells to As(III) (0.5-200 microM). Depleting cellular glutathione levels with buthionine sulfoximine (BSO) potentiated the toxicity of As(III). Cell viability was assessed with the (4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. UROtsa cell ability to biotransform As(III) was determined by dosing cells with environmentally relevant concentrations of As(III) followed by HPLC/ICP-MS analysis of cell media and lysate. Both pentavalent and trivalent monomethylated products were detected. Although cytotoxicity was observed at high doses of As(III) (approximately 100 microM) in UROtsa cells, perturbations of a variety of molecular processes occurred at much lower doses. Exposure to low level As(III) (0.5-25 microM) causes an accumulation of ubiquitin (Ub)-conjugated proteins. This effect is enhanced when cellular glutathione levels have been reduced with BSO treatment. Because As(III) has many effects on UROtsa cells, a greater understanding of how As(III) is affecting cellular proteins in a target tissue will lead to a better understanding of the mechanism of toxicity and pathogenesis for low-level As(III). PMID- 15276422 TI - Arsenic and urinary bladder cell proliferation. AB - Epidemiologic studies have demonstrated that a close association exists between the elevated levels of arsenic in drinking water and the incidence of certain cancers, including transitional cell carcinomas of the urinary bladder. We have employed in vitro and in vivo models to examine the effects of sodium arsenite on the urinary bladder epithelium. Mice exposed to 0.01% sodium arsenite in drinking water demonstrated hyperproliferation of the bladder uroepithelium within 4 weeks after initiating treatment. This occurred in the absence of amorphous precipitates and was accompanied by the accumulation of trivalent arsenite (iAs(3+)), and to a lesser extent dimethylarsenic (DMA), arsenate (iAs(5+)), and monomethylarsenic (MMA) in bladder tissue. In contrast to the bladder, urinary secretion was primarily in the form of DMA and MMA. Arsenic-induced cell proliferation in the bladder epithelium was correlated with activation of the MAP kinase pathway, leading to extracellular signal-regulated kinase (ERK) kinase activity, AP-1 activation, and expression of AP-1-associated genes involved in cell proliferation. Activation of the MAP kinase pathway involved both epidermal growth factor (EGF) receptor-dependent and -independent events, the latter involving Src activation. Studies summarized in this review suggest that arsenic accumulates in urinary bladder epithelium causing activation of specific signaling pathways that lead to chronic increased cell proliferation. This may play a non-epigenetic role in carcinogenesis by increasing the proliferation of initiated cells or increasing the mutational rate. PMID- 15276423 TI - Inhibition of insulin-dependent glucose uptake by trivalent arsenicals: possible mechanism of arsenic-induced diabetes. AB - Chronic exposures to inorganic arsenic (iAs) have been associated with increased incidence of noninsulin (type-2)-dependent diabetes mellitus. Although mechanisms by which iAs induces diabetes have not been identified, the clinical symptoms of the disease indicate that iAs or its metabolites interfere with insulin stimulated signal transduction pathway or with critical steps in glucose metabolism. We have examined effects of iAs and methylated arsenicals that contain trivalent or pentavalent arsenic on glucose uptake by 3T3-L1 adipocytes. Treatment with inorganic and methylated pentavalent arsenicals (up to 1 mM) had little or no effect on either basal or insulin-stimulated glucose uptake. In contrast, trivalent arsenicals, arsenite (iAs(III)), methylarsine oxide (MAs(III)O), and iododimethylarsine (DMAs(III)O) inhibited insulin-stimulated glucose uptake in a concentration-dependent manner. Subtoxic concentrations of iAs(III) (20 microM), MAs(III)O (1 microM), or DMAs(III)I (2 microM) decreased insulin-stimulated glucose uptake by 35-45%. Basal glucose uptake was significantly inhibited only by cytotoxic concentrations of iAs(III) or MAs(III)O. Examination of the components of the insulin-stimulated signal transduction pathway showed that all trivalent arsenicals suppressed expression and possibly phosphorylation of protein kinase B (PKB/Akt). The concentration of an insulin-responsive glucose transporter (GLUT4) was significantly lower in the membrane region of 3T3-L1 adipocytes treated with trivalent arsenicals as compared with untreated cells. These results suggest that trivalent arsenicals inhibit insulin-stimulated glucose uptake by interfering with the PKB/Akt dependent mobilization of GLUT4 transporters in adipocytes. This mechanism may be, in part, responsible for the development of type-2 diabetes in individuals chronically exposed to iAs. PMID- 15276424 TI - Arsenic-induced alterations in the contact hypersensitivity response in Balb/c mice. AB - Previous studies in our laboratory indicate that arsenic alters secretion of growth promoting and inflammatory cytokines in the skin that can regulate the migration and maturation of Langerhans cells (LC) during allergic contact dermatitis. Therefore, we hypothesized that arsenic may modulate hypersensitivity responses to cutaneous sensitizing agents by altering cytokine production, LC migration, and T-cell proliferation. To investigate this hypothesis, we examined the induction and elicitation phases of dermal sensitization. Mice exposed to 50 mg/l arsenic in the drinking water for 4 weeks demonstrated a reduction in lymph node cell (LNC) proliferation and ear swelling following sensitization with 2,4 dinitrofluorobenzene (DNFB), compared to control mice. LC and T-cell populations in the draining lymph nodes of DNFB-sensitized mice were evaluated by fluorescence-activated cell sorting; activated LC were reduced in cervical lymph nodes, suggesting that LC migration may be altered following arsenic exposure. Lymphocytes from arsenic-treated animals sensitized with fluorescein isothiocyanate (FITC) exhibited reduced proliferative responses following T-cell mitogen stimulation in vitro; however, lymphocyte proliferation from nonsensitized, arsenic-treated mice was comparable to controls. Arsenic exposure also reduced the number of thioglycollate-induced peritoneal macrophages and circulating neutrophils. These studies demonstrate that repeated, prolonged exposure to nontoxic concentrations of sodium arsenite alters immune cell populations and results in functional changes in immune responses, specifically attenuation of contact hypersensitivity. PMID- 15276425 TI - Arsenic and atherosclerosis. AB - Epidemiological studies have demonstrated a correlation between environmental or occupational arsenic exposure and a risk of vascular diseases related to atherosclerosis. Studies summarized in this review suggest that arsenic induces endothelial dysfunction, including inflammatory and coagulating activity as well as impairs nitric oxide (NO) balance. This may provide the pathophysiological basis for atherogenic potential of arsenic. Consistent with these data, arsenic accelerates atherosclerosis in apolipoprotein E (ApoE) deficient mice, a model of human atherosclerosis. PMID- 15276426 TI - Molecular basis for arsenic-induced alteration in nitric oxide production and oxidative stress: implication of endothelial dysfunction. AB - Accumulated epidemiological studies have suggested that prolonged exposure of humans to arsenic in drinking water is associated with vascular diseases. The exact mechanism of how this occurs currently unknown. Nitric oxide (NO), formed by endothelial NO synthase (eNOS), plays a crucial role in the vascular system. Decreased availability of biologically active NO in the endothelium is implicated in the pathophysiology of several vascular diseases and inhibition of eNOS by arsenic is one of the proposed mechanism s for arsenic-induced vascular diseases. In addition, during exposure to arsenic, overproduction of reactive oxygen species (ROS) can occur, resulting in oxidative stress, which is another major risk factor for vascular dysfunction. The molecular basis for decreased NO levels and increased oxidative stress during arsenic exposure is poorly understood. In this article, evidence for arsenic-mediated alteration in NO production and oxidative stress is reviewed. The results of a cross-sectional study in an endemic area of chronic arsenic poisoning and experimental animal studies to elucidate a potential mechanism for the impairment of NO formation and oxidative stress caused by prolonged exposure to arsenate in the drinking water are also reviewed. PMID- 15276427 TI - The accumulation and toxicity of methylated arsenicals in endothelial cells: important roles of thiol compounds. AB - Excess intake of arsenic is known to cause vascular diseases as well as skin lesions and cancer in humans. Recent reports suggest that trivalent methylated arsenicals, which are intermediate metabolites in the methylation process of inorganic arsenic, are responsible for the toxicity and carcinogenicity of environmental arsenic. We investigated acute toxicity and accumulation of monomethylarsonic acid (MMA(V)), dimethylarsinic acid (DMA(V)), trimethylarsine oxide (TMAO), and monomethylarsonous acid diglutathione (MMA(III) (GS)(2)) in rat heart microvessel endothelial (RHMVE) cells. MMA(V) (LC(50) = 36.6 mM) and DMA(V) (LC(50) = 2.54 mM) were less toxic than inorganic arsenicals (cf. LC(50) values for inorganic arsenite (iAs(III)), and inorganic arsenate (iAs(V)) was reported to be 36 and 220 microM, respectively, in RHMVE cells. TMAO was essentially not toxic. However, MMA(III) (GS)(2) was highly toxic (LC(50) = 4.1 microM). The order of cellular arsenic accumulation of those four organic arsenic compounds was MMA(III) (GS)(2) >> MMA(V) > DMA(V) > TMAO. MMA(III) (GS)(2) was efficiently taken up by the cells and cellular arsenic content increased with the concentration of MMA(III) (GS)(2) in culture medium. N-acetyl-l-cysteine (NAC) reduced cellular arsenic content in DMA(V)-exposed cells and also decreased the cytotoxicity of DMA(V), whereas it changed neither cellular arsenic content nor the viability in MMA(V)-exposed cells. mRNA levels of heme oxygenase-1 (HO-1) were decreased by NAC in DMA(V)-exposed, but MMA(V)-exposed cells. Buthionine sulfoximine (BSO), a cellular glutathione (GSH) depleting agent, enhanced the cytotoxicity of MMA(V). However, BSO reduced, rather than enhanced, the cytotoxicity of DMA(V). These results suggest that intracellular GSH modulated the toxic effects of arsenic in opposite ways for MMA(V) and DMA(V). Even though intracellular GSH decreased the cytotoxicity of MMA(V), extracellularly added GSH enhanced the cytotoxicity of MMA(V). The use of high-performance liquid chromatography (HPLC)-inductively coupled plasma mass spectrometric analyses suggested that a small amount of MMA(V) was converted to MMA(III) (GS)(2) in the presence of GSH. These results suggest that MMA(III) (GS)(2) is highly toxic compared to other arsenic compounds because of faster accumulation of this species by cells, in addition to having the toxic nature of methylated trivalent organic arsenics. PMID- 15276428 TI - Proteomics: empowering systems biology in plants. PMID- 15276429 TI - Rice proteomics: recent developments and analysis of nuclear proteins. AB - Rice is the most important cereal crop in Asia, and is considered as a model cereal plant for genetic and molecular studies. An immense progress has been made in rice genome sequence analysis during the last decade. This prompted the researcher to identify the functions, modifications, and regulations of every encoded protein. Proteome analysis provides information to predict the translation and relative concentration of gene products, including the extent of modification, none of which can be accurately predicted from the nucleic acid sequence alone. During the last couple of years, considerable researches were conducted to analyze rice proteome, and only recently a remarkable progress has been made to systematically analyze and characterize the functional role of various tissues and organelles in rice. In this review, the rice proteomic research has been compiled and also presented a comprehensive analysis of rice nuclear proteins. In rice nucleus, 549 proteins were resolved using 2D-PAGE. Among them, 257 proteins were systematically analyzed by Edman sequencing and mass spectrometry and identified 190 proteins following database searching (http://gene64.dna.affrc.go.jp/RPD/main.html). The identified proteins were sorted into different functional categories. In these data, the proteins involved in signaling and gene regulations dominated others, reflecting the role of nucleus in gene expression and regulation. PMID- 15276430 TI - Proteomic approach to characterize the supramolecular organization of photosystems in higher plants. AB - A project to investigate the supramolecular structure of photosystems was initiated, which is based on protein solubilizations by digitonin, protein separations by Blue native (BN)-polyacrylamide gel electrophoresis (PAGE) and protein identifications by mass spectrometry (MS). Under the conditions applied, nine photosystem supercomplexes could be described for chloroplasts of Arabidopsis, which have apparent molecular masses between 600 and 3200 kDa on BN gels. Identities of the supercomplexes were determined on the basis of their subunit compositions as documented by 2D BN/SDS-PAGE and BN/BN-PAGE. Two supercomplexes of 1060 and approximately 1600 kDa represent dimeric and trimeric forms of photosystem I (PSI), which include tightly bound LHCI proteins. Compared to monomeric PSI, these protein complexes are of low abundance. In contrast, photosystem II mainly forms part of dominant supercomplexes of 850, 1000, 1050 and 1300 kDa. According to our interpretation, these supercomplexes contain dimeric PSII, 1-4 LHCII trimers and additionally monomeric LHCII proteins. The 1300-kDa PSII supercomplex (containing four LHCII trimers) is partially converted into the 1000-kDa PSII supercomplex (containing two LHCII trimers) in the presence of dodecylmaltoside on 2D BN/BN gels. Analyses of peptides of the trypsinated 1300-kDa PSII supercomplex by mass spectrometry allowed to identify known subunits of the PSII core complex and additionally LHCII proteins encoded by eight different genes in Arabidopsis. Further application of this experimental approach will allow new insights into the supermolecular organization of photosystems in plants. PMID- 15276431 TI - The hydrophobic proteome of mitochondrial membranes from Arabidopsis cell suspensions. AB - The development of mitochondria and the integration of their function within a plant cell rely on the presence of a complex biochemical machinery located within their limiting membranes. The aim of the present work was: (1) to enhance our understanding of the biochemical machinery of mitochondrial membranes and (2) to test the versatility of the procedure developed for the identification of the hydrophobic proteome of the chloroplast envelope [Molecular and Cellular Proteomics 2 (2003) 325-345]. A proteomic analysis was performed, to provide the most exhaustive view of the protein repertoire of these membranes. For this purpose, highly purified mitochondria were prepared from Arabidopsis cultured cells and membrane proteins were extracted. To get a more exhaustive array of membrane proteins from Arabidopsis mitochondria, from the most to the less hydrophobic ones, various extraction procedures (chloroform/methanol extraction, alkaline or saline treatments) were applied. LC-MS/MS analyses were then performed on each membrane subfraction, leading to the identification of more than 110 proteins. The identification of these proteins is discussed with respect to their mitochondrial localization, their physicochemical properties and their implications in the metabolism of mitochondria. In order to provide a new overview of the biochemical machinery of the plant mitochondria, proteins identified during this work were compared to the lists of proteins identified during previous proteomic analyses performed on plant and algae mitochondria (Arabidopsis, pea, Chlamydomonas, rice, etc.). A total of 502 proteins are listed. About 40% of the 114 proteins identified during this work were not identified during previous proteomic studies performed on mitochondria. PMID- 15276432 TI - Proteomics of Medicago sativa cell walls. AB - A method for the sequential extraction and profiling by two-dimensional gel electrophoresis (2-DE) of Medicago sativa (alfalfa) stem cell wall proteins is described. Protein extraction included freezing, grinding in a sodium acetate buffer, separation by filtration of cell walls from cytosolic contents, and extensive washing. Cell wall proteins were then extracted sequentially with a solution containing 200 mM CaCl2 and 50 mM sodium acetate, followed by extraction with 3.0 M LiCl and 50 mM sodium acetate. Cell wall proteins from both the CaCl2 and LiCl fractions were profiled by 2-DE. Approximately 150 protein spots were extracted from these two gels, digested with trypsin, and analyzed using nanoscale HPLC coupled to a hybrid quadrupole time-of-flight (Q-tof) tandem mass spectrometer (LC/MS/MS). More than 100 proteins were identified and used in conjunction with the 2-DE profiles to generate proteomic reference maps for cell walls of this important legume. Identified proteins include classical cell wall proteins as well as proteins traditionally considered as non-secreted. Two unique extracellular proteins were also identified. PMID- 15276433 TI - Sub-cellular proteomic analysis of a Medicago truncatula root microsomal fraction. AB - Since the last decade, Medicago truncatula has emerged as one of the model plants particularly investigated in the field of plant-microbe interactions. Several genetic and molecular approaches including proteomics have been developed to increase knowledge about this plant species. To complement the proteomic data, which have mainly focused on the total root proteins from M. truncatula, we carried out a sub-cellular approach to gain access to the total membrane associated proteins. Following the setting up of the purification process, microsomal proteins were separated on 2-DE. Ninety-six out of the 440 well resolved proteins were identified by MALDI-TOF peptide mass fingerprinting. A high percent (83%) of successful protein identification was obtained when using M. truncatula clustered EST database for queries. During the purification process, the enrichment in membrane-associated proteins was monitored on 2-D gels. The membrane location of microsomal proteins was further confirmed using PMF identification. This study reports a fractionation process for characterizing microsomal root proteins of M. truncatula, which could be an interesting tool for investigating the molecular mechanisms involved in root symbioses. PMID- 15276434 TI - High-throughput peptide mass fingerprinting of soybean seed proteins: automated workflow and utility of UniGene expressed sequence tag databases for protein identification. AB - Identification of anonymous proteins from two-dimensional (2-D) gels by peptide mass fingerprinting is one area of proteomics that can greatly benefit from a simple, automated workflow to minimize sample contamination and facilitate high throughput sample processing. In this investigation we outline a workflow employing robotic automation at each step subsequent to 2-D gel electrophoresis. As proof-of-concept, 96 protein spots from a 2-D gel were analyzed using this approach. Whole protein (1 mg) from mature, dry soybean (Glycine max [L.] Merr.) cv. Jefferson seed was resolved by high resolution 2-D gel electrophoresis. Approximately 150 proteins were observed after staining with Coomassie Blue. The rather low number of detected proteins was due to the fact that the dynamic range of protein expression was greater than 100-fold. The most abundant proteins were seed storage proteins which in total represented over 60% of soybean seed protein. Using peptide mass fingerprinting 44 protein spots were identified. Identification of soybean proteins was greatly aided by the use of annotated, contiguous Expressed Sequence Tag (EST) databases which are available for public access (UniGene, ftp.ncbi.nih.gov/repository/UniGene/). Searches were orders of magnitude faster when compared to searches of unannotated EST databases and resulted in a higher frequency of valid, high-scoring matches. Some abundant, non seed storage proteins identified in this investigation include an isoelectric series of sucrose binding proteins, alcohol dehydrogenase and seed maturation proteins. This survey of anonymous seed proteins will serve as the basis for future comparative analysis of seed-filling in soybean as well as comparisons with other soybean varieties. PMID- 15276435 TI - Proteomics of calcium-signaling components in plants. AB - Calcium functions as a versatile messenger in mediating responses to hormones, biotic/abiotic stress signals and a variety of developmental cues in plants. The Ca(2+)-signaling circuit consists of three major "nodes"--generation of a Ca(2+) signature in response to a signal, recognition of the signature by Ca2+ sensors and transduction of the signature message to targets that participate in producing signal-specific responses. Molecular genetic and protein-protein interaction approaches together with bioinformatic analysis of the Arabidopsis genome have resulted in identification of a large number of proteins at each "node"--approximately 80 at Ca2+ signature, approximately 400 sensors and approximately 200 targets--that form a myriad of Ca2+ signaling networks in a "mix and match" fashion. In parallel, biochemical, cell biological, genetic and transgenic approaches have unraveled functions and regulatory mechanisms of a few of these components. The emerging paradigm from these studies is that plants have many unique Ca2+ signaling proteins. The presence of a large number of proteins, including several families, at each "node" and potential interaction of several targets by a sensor or vice versa are likely to generate highly complex networks that regulate Ca(2+)-mediated processes. Therefore, there is a great demand for high-throughput technologies for identification of signaling networks in the "Ca(2+)-signaling-grid" and their roles in cellular processes. Here we discuss the current status of Ca2+ signaling components, their known functions and potential of emerging high-throughput genomic and proteomic technologies in unraveling complex Ca2+ circuitry. PMID- 15276436 TI - Identification of barley CK2alpha targets by using the protein microarray technology. AB - We have successfully established a novel protein microarray-based kinase assay, which we applied to identify target proteins of the barley protein kinase CK2alpha. As a source of recombinant barley proteins we cloned cDNAs specific for filial tissues of developing barley seeds into an E. coli expression vector. By using robot technology, 21,500 library clones were arrayed in microtiter plates and gridded onto high-density filters. Protein expressing clones were detected using an anti-RGS-His6 antibody and rearrayed into a sublibrary of 4100 clones. All of these clones were sequenced from the 5'-end and the sequences were analysed by homology searches against protein databases. Based on these results we selected 768 clones expressing different barley proteins for protein purification. The purified proteins were robotically arrayed onto FAST slides. The generated protein microarrays were incubated with an expression library derived barley CK2alpha in the presence of [gamma-33P]ATP, and signals were detected by X-ray film or phosphor imager. We were able to demonstrate the power of the protein microarray technology by identification of 21 potential targets out of 768 proteins including such well-known substrates of CK2alpha as high mobility group proteins and calreticulin. PMID- 15276437 TI - Proteomics-based sequence analysis of plant gene expression--the chloroplast transcription apparatus. AB - The chloroplast transcription apparatus has turned out to be more complex than anticipated, with core polypeptides surrounded by multiple accessory proteins of diverse, and in part unexpected, functions. At least two different RNA-binding proteins and several redox-responsive proteins are components of the major chloroplast RNA polymerase termed PEP-A. One of the key-regulatory factors has been identified as a Ser/Thr-specific protein kinase that is sensitive to SH group modification by glutathione and by this means is able to modulate transcription. The cloned plastid transcription kinase from mustard (Sinapis alba L.) has been assigned as a member of the (mostly nucleo-cytosolic) CK2 family and hence has been termed cpCK2. Despite its apparent role in mustard chloroplast transcription, until recently no data have been available for other plant species. Using the web database resources, we find evidence for an evolutionarily conserved role of this redox-sensitive plastid transcription factor. PMID- 15276438 TI - Proteomics of curcurbit phloem exudate reveals a network of defence proteins. AB - Many different proteins can be separated from the sap of mature sieve tubes of different plant species. To date, only a limited number of those have been identified and functionally characterised. Due to sieve tubes inability of transcription and translation, the proteins are most probably synthesised in the intimately connected companion cells and transported into the sieve elements through plasmodesmata. The specific protein composition of phloem sap suggests an important role of these proteins not only for sieve tube maintenance, but also for whole plant physiology and development. Here we describe a comprehensive analysis of the phloem protein composition employing one- and high-resolution two dimensional gel electrophoresis and partial sequencing by mass spectrometry. In this study more than 300 partial sequences generated by hybrid mass spectrometry were used to identify a total of 45 different proteins from the phloem exudates of cucumber (Cucumis sativus L. cv. Hoffmanns Giganta) and pumpkin (Cucurbita maxima Duch. cv. Gelber Zentner) plants. In addition to previously described phloem proteins, it was possible to localise proteins with high similarity to an acyl-CoA binding protein, a glyoxalase, a malate dehydrogenase, a rhodanese-like protein, a drought-induced protein, and a beta-glucosidase. The results indicate that the majority of the so far identified proteins are involved in stress and defence reactions. PMID- 15276439 TI - Specific changes in the Arabidopsis proteome in response to bacterial challenge: differentiating basal and R-gene mediated resistance. AB - Alterations in the proteome of Arabidopsis thaliana leaves during early responses to challenge by Pseudomonas syringae pv. tomato DC3000 (DC3000) were analysed using two-dimensional (2D) gel electrophoresis. Protein changes characteristic of the establishment of basal resistance and R-gene mediated resistance were examined by comparing responses to DC3000, a hrp mutant and DC3000 expressing avrRpm1 respectively. The abundance of selected transcripts was also analysed in GeneChip experiments. Here we present data from the soluble fraction of leaf protein, highlighting changes in two antioxidant enzyme groups; the glutathione S transferases (GSTs F2, F6, F7 and F8) and peroxiredoxins (PrxA, B and IIE). Members of both enzyme groups showed signs of specific post-translational modifications, represented by multiple spots on gels. We suggest that oxidation of specific residues is responsible for some of the spot shifts. All forms of the GST proteins identified here increased following inoculation with bacteria. GSTF8 showed particularly dynamic responses to pathogen challenge, the corresponding transcript was significantly up-regulated by 2 h after inoculation, and the protein showed post-translational modifications specific to an incompatible interaction. Differential changes were observed with the peroxiredoxin proteins; PrxIIE and to a lesser extent PrxB, no change was observed with PrxA, but a truncated form PrxA-L was greatly reduced in abundance following bacterial challenges. Our data suggest that bacterial challenge generally induces Prxs and the antioxidants GSTs, however individual members of these families may be specifically modified dependent upon the virulence of the DC3000 strain and outcome of the interaction. Finally, proteomic and transcriptomic data derived from the same inoculation system are compared and the advantages offered by 2D gel analysis discussed in light of our results. PMID- 15276440 TI - A proteomic approach to studying plant response to crenate broomrape (Orobanche crenata) in pea (Pisum sativum). AB - Crenate broomrape (Orobanche crenata) is a parasitic plant that threatens legume production in Mediterranean areas. Pea (Pisum sativum) is severely affected, and only moderate levels of genetic resistance have so far been identified. In the present work we selected the most resistant accession available (Ps 624) and compared it with a susceptible (Messire) cultivar. Experiments were performed by using pot and Petri dish bioassays, showing little differences in the percentage of broomrape seed germination induced by both genotypes, but a significant hamper in the number of successfully installed tubercles and their developmental stage in the Ps 624 compared to Messire. The protein profile of healthy and infected P. sativum root tissue were analysed by two-dimensional electrophoresis. Approximately 500 individual protein spots could be detected on silver stained gels. At least 22 different protein spots differentiated control, non-infected, Messire and Ps 624 accessions. Some of them were identified by MALDI-TOF mass spectrometry and database searching as cysteine proteinase, beta-1,3-glucanase, endochitinase, profucosidase, and ABA-responsive protein. Both qualitative and quantitative differences have been found among infected and non-infected root extracts. Thus, in the infected susceptible Messire genotype 34 spots were decreased, one increased and three newly detected, while in Ps 624, 15 spots were increased, three decreased and one newly detected. In response to the inoculation, proteins that correspond to enzymes of the carbohydrate metabolism (fructokinase, fructose-bisphosphate aldolase), nitrogen metabolism (ferredoxin NADP reductase) and mitochondrial electronic chain transport (alternative oxidase 2) decreased in the susceptible check, while proteins that correspond to enzymes of the nitrogen assimilation pathway (glutamine synthetase) or typical pathogen defence, PR proteins, including beta-1,3-glucanase and peroxidases, increased in Ps 624. Results are discussed in terms of changes in the carbohydrate and nitrogen metabolism an induction of defence proteins in response to broomrape parasitism. PMID- 15276441 TI - A proteomic analysis of plant programmed cell death. AB - Programmed cell death (PCD) is an active cellular suicide that occurs in animals and plants throughout development and in response to both abiotic and biotic stresses. In contrast to animals, little is known about the molecular machinery that regulates plant PCD. We have previously identified transcriptomic changes associated with heat- and senescence-induced PCD in an Arabidopsis cell suspension culture [Plant J. 30 (2002) 431]. However, since plant PCD is also likely to involve elements that are regulated post-transcriptionally, we have undertaken a proteomic analysis in the Arabidopsis system. We identified 11 proteins that increased in abundance relative to total protein in both treatments despite extensive degradation of other proteins. We argue that some of these proteins are maintained during PCD and may therefore have specific functions in the PCD pathway. The increased abundance of several antioxidant proteins as well as a measured increase in free Fe2+ content of the cells indicates an oxidative stress in this system. Several mitochondrial proteins were identified, confirming the importance of this organelle during PCD. We also identified an extracellular glycoprotein that may function in the transmission of a 'death signal' from cell to cell. Putative roles for the identified proteins are presented. PMID- 15276442 TI - Identification of oxidised proteins in the matrix of rice leaf mitochondria by immunoprecipitation and two-dimensional liquid chromatography-tandem mass spectrometry. AB - Highly purified mitochondria were isolated from green 7-day-old rice leaves. The mitochondria were sonicated and the matrix fraction isolated as the 100,000g supernatant. Part of the matrix fraction was left untreated while the other part was subjected to a mild oxidative treatment (0.5 mM H2O2+0.2 mM CuSO4 for 10 min at room temperature). The oxidised proteins in both samples were tagged with dinitrophenylhydrazine (DNP), which forms a covalent bond with carbonyl groups. The DNP-tagged proteins were immunoprecipitated using anti-DNP antibodies and digested with trypsin. The mixture of peptides was analysed by nano-HPLC coupled online to an ESI-Quad-TOF mass spectrometer. The peptides were separated by stepwise ion exchange chromatography followed by reverse phase chromatography (2D LC), and analysed by MS/MS. Proteins were identified by un-interpreted fragment ion database searches. Using this approach we identified 20 oxidised proteins in the control sample and a further 32 in the oxidised sample. Western blots of 2D gels of the same samples prior to immunoprecipitation verified that the oxidation treatment increases protein oxidation also for specific proteins. Likewise Western blots showed that neither the isolation of mitochondria nor their subfractionation introduced carbonyl groups. We therefore conclude that a number of proteins are oxidised in the matrix of rice leaf mitochondria in vivo and further identify a group of proteins that are particularly susceptible to mild oxidation in vitro. PMID- 15276443 TI - Proteomic analysis of small heat shock protein isoforms in barley shoots. AB - The analysis of stress-responsiveness in plants is an important route to the discovery of genes conferring stress tolerance and their use in breeding programs. High temperature is one of the environmental stress factors that can affect the growth and quality characteristics of barley (Hordeum vulgare). In this study a proteomic analysis (2D-PAGE, MS) was used to detect the effects of heat shock on the protein pattern of an abiotic stress-tolerant (Mandolina) and an abiotic stress-susceptible (Jubilant) barley cultivar. Evaluation of two dimensional gels revealed several proteins to be differentially expressed as a result of heat stress in both cultivars. The protein spots of interest were, after an in-gel tryptic digestion, further investigated by mass spectrometry. For the analysis of the peptide mixture, we both used a matrix-assisted laser desorption/ionization (MALDI) tandem time of flight mass spectrometer (TOF/TOF) and an automated nano-HPLC system coupled to an electrospray ionization quadrupole linear ion trap (Q-TRAP) instrument. The hyphenation of the latter techniques proved to be a powerful technique as shown by the identification of six isoforms of a 16.9 kDa sHSP in one single spot. We observed that S adenosylmethionine synthetase (SAM-S) was differentially expressed between the two cultivars. Recent results refer to the role of SAM-S as being involved in abiotic stress tolerance. Furthermore, comparison of the heat shock treated samples also revealed several small heat shock proteins (sHSP), of which distinct isoforms could be characterised. PMID- 15276445 TI - Plant proteome analysis by mass spectrometry: principles, problems, pitfalls and recent developments. AB - The genome of several species has now been elucidated; these genomes indicate the proteomic potential of the cell. While identification of genomes has been, and continues to be, a technically and intellectually demanding process, the identification of the proteome contains inherently greater difficulties. The proteome of each living cell is dynamic, altering in response to the individual cell's metabolic state and reception of intracellular and extracellular signal molecules, and many of the proteins which are expressed will be post translationally altered. Thus if the purpose of the proteome analysis is to aid the understanding of protein function and interaction, then it is identification of the proteins in their final state which is required: for this mass spectrometric identification of individual proteins, indicating site and nature of modifications, is essential. Here we review the principles of the methodologies involved in such analyses, give some indication of current achievements in plant proteomics, and indicate imminent and prospective technical developments. PMID- 15276446 TI - Technical aspects of functional proteomics in plants. AB - Since the completion of genome sequences of several organisms, attention has been focused to determine the function and functional network of proteins by proteome analysis. This analysis is achieved by separation and identification of proteins, determination of their function and functional network, and construction of an appropriate database. Many improvements in separation and identification of proteins, such as two-dimensional electrophoresis, nano-liquid chromatography and mass spectrometry, have rapidly been achieved. Some new techniques which include top-down mass spectrometry and tandem affinity purification have emerged. These techniques have provided the possibility of high-throughput analysis of function and functional network of proteins in plants. However, to cope with the huge information emerging from proteome analyses, more sophisticated techniques and software are essential. The development and adaptation of such techniques will ease analyses of protein profiling, identification of post-translational modifications and protein-protein interaction, which are vital for elucidation of the protein functions. PMID- 15276447 TI - Proteomic capacity of recent fluorescent dyes for protein staining. AB - Staining of two-dimensional gel constitutes a crucial step in comparative proteome analysis with respect to both the number of proteins analysed, the accuracy of spot quantification and reproducibility. In this work, we compared the efficiency of recent fluorophores to stain Arabidopsis total protein extract: Sypro Ruby (SR), Deep Purple (DP) and 5-hexadecanoylamino-fluorescein (C16-F). In addition, classical visible dyes, colloidal Coomassie blue (CCB) and silver nitrate (SN), were also included. High quality images were obtained for the three fluorescent dyes, DP giving the cleaner background, whereas spikes were observed with SR and a rough background with C16-F. On the other hand, saturation occurred for abundant spots with SR and DP. For a same protein load the number of detected spots ranged between 250 for CCB and 800 for SR in the sequence SR > DP approximately SN > C16-F > CCB. These differences were shown to rely mainly on the sensitivity between dyes leading to the detection of additional spots belonging to classes of lower abundance. Analysis of the distribution of variation coefficients for spots from replicates showed differences in the staining reproducibility between dyes that ranged in the order SR > C16-F > DP > SN > CCB. The implications of these results for the selection of a convenient stain are discussed according to specific objectives as well as practical aspects. PMID- 15276448 TI - Subtle modification of isotope ratio proteomics; an integrated strategy for expression proteomics. AB - Use of minor modification of isotope ratio to code samples for expression proteomics is being investigated. Alteration of (13)C abundance to approximately 2% yields a measurable effect on peptide isotopic distribution and inferred isotope ratio. Elevation of (13)C abundance to 4% leads to extension of isotopic distribution and background peaks across every unit of the mass range. Assessment of isotope ratio measurement variability suggests substantial contributions from natural measurement variability. A better understanding of this variable will allow assessment of the contribution of sequence dependence. Both variables must be understood before meaningful mixing experiments for relative expression proteomics are performed. Subtle modification of isotope ratio ( approximately 1 2% increase in (13)C) had no effect upon either the ability of data-dependent acquisition software or database searching software to trigger tandem mass spectrometry or match MSMS data to peptide sequences. More severe modification of isotope ratio caused a significant drop in performance of both functionalities. Development of software for deconvolution of isotope ratio concomitant with protein identification using LC-MSMS, or any other proteomics strategy, is underway (Isosolv). The identified peptide sequence is then be used to provide elemental composition for accurate isotope ratio decoding and the potential to control for specific amino acid biases should these prove significant. It is suggested that subtle modification of isotope ratio proteomics (SMIRP) offers a convenient approach to in vivo isotope coding of plants and might ultimately be extended to mammals including humans. PMID- 15276449 TI - Untangling multi-gene families in plants by integrating proteomics into functional genomics. AB - The classification and study of gene families is emerging as a constructive tool for fast tracking the elucidation of gene function. A multitude of technologies can be employed to undertake this task including comparative genomics, gene expression studies, sub-cellular localisation studies and proteomic analysis. Here we focus on the growing role of proteomics in untangling gene families in model plant species. Proteomics can specifically identify the products of closely related genes, can determine their abundance, and coupled to affinity chromatography and sub-cellular fractionation studies, it can even provide location within cells and functional assessment of specific proteins. Furthermore global gene expression analysis can then be used to place a specific family member in the context of a cohort of co-expressed genes. In model plants with established reverse genetic resources, such as catalogued T-DNA insertion lines, this gene specific information can also be readily used for a wider assessment of specific protein function or its capacity for compensation through assessing whole plant phenotypes. In combination, these resources can explore partitioning of function between members and assess the level of redundancy within gene families. PMID- 15276450 TI - Multivariate approaches in plant science. AB - The objective of proteomics is to get an overview of the proteins expressed at a given point in time in a given tissue and to identify the connection to the biochemical status of that tissue. Therefore sample throughput and analysis time are important issues in proteomics. The concept of proteomics is to encircle the identity of proteins of interest. However, the overall relation between proteins must also be explained. Classical proteomics consist of separation and characterization, based on two-dimensional electrophoresis, trypsin digestion, mass spectrometry and database searching. Characterization includes labor intensive work in order to manage, handle and analyze data. The field of classical proteomics should therefore be extended to also include handling of large datasets in an objective way. The separation obtained by two-dimensional electrophoresis and mass spectrometry gives rise to huge amount of data. We present a multivariate approach to the handling of data in proteomics with the advantage that protein patterns can be spotted at an early stage and consequently the proteins selected for sequencing can be selected intelligently. These methods can also be applied to other data generating protein analysis methods like mass spectrometry and near infrared spectroscopy and examples of application to these techniques are also presented. Multivariate data analysis can unravel complicated data structures and may thereby relieve the characterization phase in classical proteomics. Traditionally statistical methods are not suitable for analysis of the huge amounts of data, where the number of variables exceed the number of objects. Multivariate data analysis, on the other hand, may uncover the hidden structures present in these data. This study takes its starting point in the field of classical proteomics and shows how multivariate data analysis can lead to faster ways of finding interesting proteins. Multivariate analysis has shown interesting results as a supplement to classical proteomics and added a new dimension to the field of proteomics. PMID- 15276451 TI - Genome-scale, biochemical annotation method based on the wheat germ cell-free protein synthesis system. AB - Since the complete genomic DNA sequencing of various species, attention has turned to the structural properties, and functional characteristics of proteins. Current cell-free protein expression systems from eukaryotes are capable of synthesizing proteins with high speed and accuracy; however, the yields are low due to their instability over time. This report reviews the high-throughput, genome-scale biochemical annotation method based on the cell-free system prepared from wheat embryos. We first briefly reviewed our highly efficient and robust wheat germ cell-free protein synthesis system, and then showed an application of the system for materialization and characterization of genetic information taking a cDNA library of protein kinase from Arabidopsis thaliana as an example. The procedure consists of: (1) fusion of the gene-of-interest to a purification-tag, amplified by the split-primer PCR method; (2) transcription and purification of mRNA; (3) cell-free protein synthesis in the bilayer system using 96-well titer plate; (4) affinity purification and activity measurement. We took 439 cDNAs encoding kinases among 1064 genes annotated so far, and they were translated in parallel into protein. Subsequent assay revealed 207 products having autophosphorylation activity. Furthermore, seven proteins out of 26 calcium dependent protein kinase genes tested did phosphorylate a synthetic peptide substrate in the presence of calcium ion, demonstrating that the translation products, retained their substrate specificity. The information on biochemical function of gene products accumulated should revolutionize our understanding of biology and fundamentally alter the practice of medicine and influence other industries as well. PMID- 15276452 TI - The Arabidopsis phenylalanine ammonia lyase gene family: kinetic characterization of the four PAL isoforms. AB - In Arabidopsis thaliana, four genes have been annotated as provisionally encoding PAL. In this study, recombinant native AtPAL1, 2, and 4 were demonstrated to be catalytically competent for l-phenylalanine deamination, whereas AtPAL3, obtained as a N-terminal His-tagged protein, was of very low activity and only detectable at high substrate concentrations. All four PALs displayed similar pH optima, but not temperature optima; AtPAL3 had a lower temperature optimum than the other three isoforms. AtPAL1, 2 and 4 had similar K(m) values (64-71 microM) for l-Phe, with AtPAL2 apparently being slightly more catalytically efficacious due to decreased K(m) and higher k(cat) values, relative to the others. As anticipated, PAL activities with l-tyrosine were either low (AtPAL1, 2, and 4) or undetectable (AtPAL3), thereby establishing that l-Phe is the true physiological substrate. This detailed knowledge of the kinetic and functional properties of the various PAL isoforms now provides the necessary biochemical foundation required for the systematic investigation and dissection of the organization of the PAL metabolic network/gene circuitry involved in numerous aspects of phenylpropanoid metabolism in A. thaliana spanning various cell types, tissues and organs. PMID- 15276453 TI - Proteome analysis differentiates between two highly homologues germin-like proteins in Arabidopsis thaliana ecotypes Col-0 and Ws-2. AB - A proteome approach based on 2-D gel electrophoresis (2-DE) was used to compare the protein patterns of the Arabidopsis ecotypes Col-0 and Ws-2. In leaf extracts a pair of protein spots were found to be diagnostic for each of the lines. Both pairs of spots were identified as closely related germin-like proteins differing in only one amino acid by using peptide mass finger printing of tryptic digests and by gaining additional data from post-source decay spectra in the MALDI-TOF analysis. Western blot analysis after separation of protein extracts by 2-DE confirmed results from Coomassie blue-stained gels and revealed additional immunoreactive spots for both ecotypes most likely representing dimers of the spots first identified. Western blot analysis and mass spectrometrical identification of the corresponding weakly stained protein in Coomassie blue stained gels of the ecotype Col-0 also demonstrated for the first time the occurrence of AtGER3 protein in root extracts. Our results demonstrate the capacity of proteome analysis to analyse and distinguish closely related members of large protein families. PMID- 15276454 TI - Investigation of cationic peanut peroxidase glycans by electrospray ionization mass spectrometry. AB - Cationic peanut peroxidase (CP) was isolated from peanut (Arachis hypogaea) cell suspension culture medium. CP is a glycoprotein with three N-linked glycan sites at Asn60, Asn144, and Asn185. ESI-MS of the intact purified protein reveals the microheterogeneity of the glycans. Tryptic digestion of CP gave a near complete sequence coverage by ESI-MS. The glycopeptides from the tryptic digestion were separated by RP HPLC identified by ESI-MS and the structure of the glycan chains determined by ESI-MS/MS. The glycans are large structures of up to 16 sugars, but most of their non-reducing ends have been modified giving a mixture of shorter chains at each site. Good agreement was found with the one glycan previously analyzed by (1)H NMR. This work is the basis for the future studies on the role of the glycans on stability and folding of CP and is another example of a detailed structural characterization of complex glycoproteins by mass spectrometry. PMID- 15276455 TI - Mapping of the Physcomitrella patens proteome. AB - The moss Physcomitrella patens is unique among land plants due to the high rate of homologous recombination in its nuclear DNA. The feasibility of gene targeting makes Physcomitrella an unrivalled model organism in the field of plant functional genomics. To further extend the potentialities of this seed-less plant we aimed at exploring the P. patens proteome. Experimental conditions had to be adopted to meet the special requirements connected to the investigations of this moss. Here we describe the identification of 306 proteins from the protonema of Physcomitrella. Proteins were separated by two dimensional electrophoresis, excised form the gel and analysed by means of mass spectrometry. This reference map will lay the basis for further profound studies in the field of Physcomitrella proteomics. PMID- 15276456 TI - A two-dimensional proteome map of maize endosperm. AB - We have established a proteome reference map for maize (Zea mays L.) endosperm by means of two-dimensional gel electrophoresis and protein identification with LC MS/MS analysis. This investigation focussed on proteins in major spots in a 4-7 pI range and 10-100 kDa M(r) range. Among the 632 protein spots processed, 496 were identified by matching against the NCBInr and ZMtuc-tus databases (using the SEQUEST software). Forty-two per cent of the proteins were identified against maize sequences, 23% against rice sequences and 21% against Arabidopsis sequences. Identified proteins were not only cytoplasmic but also nuclear, mitochondrial or amyloplastic. Metabolic processes, protein destination, protein synthesis, cell rescue, defense, cell death and ageing are the most abundant functional categories, comprising almost half of the 632 proteins analyzed in our study. This proteome map constitutes a powerful tool for physiological studies and is the first step for investigating the maize endosperm development. PMID- 15276457 TI - Environmental and transgene expression effects on the barley seed proteome. AB - The barley (Hordeum vulgare) cultivar Golden Promise is no longer widely used for malting, but is amenable to transformation and is therefore a valuable experimental cultivar. Its characteristics include high salt tolerance, however it is also susceptible to several fungal pathogens. Proteome analysis was used to describe the water-soluble protein fraction of Golden Promise seeds in comparison with the modern malting cultivar Barke. Using 2D-gel electrophoresis to visualise several hundred proteins in the pH ranges 4-7 and 6-11, 16 protein spots were found to differ between the two cultivars. Eleven of these were identified by mass spectrometric peptide mass mapping, including an abundant chitinase implicated in defence against fungal pathogens and a small heat-shock protein. To enable a comparison with transgenic seed protein patterns, differences in spot patterns between field and greenhouse-grown seeds were analysed. Four spots were observed to be increased in intensity in the proteome of greenhouse-grown seeds, three of which may be related to nitrogen availability during grain filling and total protein content of the seeds, since they also increased in field grown seeds supplied with extra nitrogen. Finally, the fate of transgene products in barley seeds was followed. Spots containing two green fluorescent protein constructs and the herbicide resistance marker phosphinothricin acetyltransferase were observed in 2D-gel patterns of transgenic seeds and identified by mass spectrometry. Phosphinothricin acetyltransferase was observed in three spots differing in pI suggesting that post-translational modification of the transgene product had occurred. PMID- 15276458 TI - Thioredoxin targets of developing wheat seeds identified by complementary proteomic approaches. AB - The role of thioredoxin in wheat starchy endosperm was investigated utilizing two proteomic approaches. Thioredoxin targets were isolated from total KCl-soluble extracts of endosperm and flour and separated by 2-DE following (1) reduction of the extract by the NADP/thioredoxin system and labeling the newly generated sulfhydryl (SH) groups with monobromobimane (mBBr), and, in parallel, (2) trapping covalently interacting proteins on an affinity column prepared with mutant thioredoxin h in which one of the active site cysteines was replaced by serine. The two procedures were complementary: of the total targets, one-third were observed with both procedures and one-third were unique to each. Altogether 68 potential targets were identified; almost all containing conserved cysteines. In addition to confirming known interacting proteins, we identified 40 potential thioredoxin targets not previously described in seeds. A comparison of the results obtained with young endosperm (isolated 10 days after flowering) to those with mature endosperm (isolated 36 days after flowering) revealed a unique set of proteins functional in processes characteristic of each developmental stage. Flour contained 36 thioredoxin targets, most of which have been found in the isolated developing endosperm. PMID- 15276459 TI - Cell-specific protein profiling in Arabidopsis thaliana trichomes: identification of trichome-located proteins involved in sulfur metabolism and detoxification. AB - Metabolite, protein, and transcript analysis at the cellular level gives unparalleled insight into the complex roles tissues play in the plant system. However, while capillary electrophoresis and PCR amplification strategies make the profiling of metabolites and transcripts in specific cell types possible, the profiling of proteins in small samples represents a bottleneck. Here for the first time protein profiling has been achieved in a specific plant cell type: The application of specific cell sampling and shotgun peptide sequencing (nano LC/MS/MS) resulted in the identification of 63 unique proteins from pooled Arabidopsis trichome cells. A complete S-adenosylmethionine pathway cluster, two S-adenosylmethionine synthase isoforms, a glutathione S-conjugate translocator and other proteins involved in sulfur metabolism and detoxification are shown to be present in these cells, in agreement with previous work done at the level of trichome transcript analysis. The technology described here brings the simultaneous identification and localization of physiologically relevant cellular proteins within reach. PMID- 15276460 TI - Impact of sewage sludges on Medicago truncatula symbiotic proteome. AB - The effects of sewage sludges were investigated on the symbiotic interactions between the model plant Medicago truncatula and the arbuscular mycorrhizal fungus Glomus mosseae or the rhizobial bacteria Sinorhizobium meliloti. By comparison to a control sludge showing positive effects on plant growth and root symbioses, sludges enriched with polycylic aromatic hydrocarbons or heavy metals were deleterious. Symbiosis-related proteins were detected and identified by two dimensional electrophoresis and matrix-assisted laser desorption ionization mass spectrometry, and image analysis was used to study the effects of sewage sludges on M. truncatula symbiotic proteome. PMID- 15276461 TI - Synchronization and integration of multiple hypertrophic pathways in the heart. PMID- 15276462 TI - Adaptive and maladaptive hypertrophic pathways: points of convergence and divergence. AB - Myocardial hypertrophy is a response of cardiac muscle to altered conditions of haemodynamic overload caused by a large number of physiological and pathological conditions. Traditionally, it has been considered a beneficial mechanism. However, sustained hypertrophy has been associated with a significant increase in the risk of cardiovascular disease and mortality. Actually, many researchers are trying to understand whether left ventricular hypertrophy is a 'good' mechanism to stimulate or a 'bad' process to prevent. In this review we investigate the most common biochemical signaling pathways involved in the hypertrophic response to identify the precise role, either 'adaptive' or 'maladaptive', of each molecular pathway. Delinealing intracellular signaling pathways involved in the different aspects of cardiac hypertrophy will permit future improvements in the signaling that controls beneficial growth. PMID- 15276463 TI - Molecular and mechanical synergy: cross-talk between integrins and growth factor receptors. AB - The full repertoire of molecules and mechanisms which lead to cardiac hypertrophy are poorly understood. Many studies over the last several decades have shown how various growth factors are involved in the hypertrophic response. It has also been intuitively obvious that mechanical mechanisms which impose hemodynamic loads on the working myocardium must also be involved in this process. Integrins are cell adhesion receptors that are potent bi-directional signaling molecules. They are cellular mechanoreceptors in many cells and are clearly one of the molecules which orchestrate mechano-biochemical coupling in the heart. In recent years they too have been shown to be involved in the hypertrophic response pathway. This review will detail background information on integrins in general, discuss integrins in the myocardium and illustrate how integrin and growth factor signaling pathways might combinatorially function in the heart. PMID- 15276464 TI - Network integration of the adrenergic system in cardiac hypertrophy. AB - Adrenergic receptors play a pivotal role in regulating cardiac function in response to a constantly changing environment. Altered alpha and beta adrenergic receptor signaling in vivo is associated with cardiac hypertrophy and failure. This review focuses on the different roles of adrenergic receptors in regulating cardiac function under normal and pathological conditions. Understanding the signaling mechanisms of these receptors in the context of the heart is likely to provide a better therapeutic approach towards treatment of heart failure. PMID- 15276465 TI - Ras, PI3-kinase and mTOR signaling in cardiac hypertrophy. AB - Cardiac hypertrophy involves increased mass (growth) of the heart and a cardinal feature of this condition is increased rates of protein synthesis. Several signaling pathways have been implicated in cardiac hypertrophy including the phosphatidylinositol 3-kinase (PI3K) and Ras/Raf/MEK/Erk pathways. PI3K lies upstream of the mammalian target of rapamycin (mTOR), an important positive regulator of protein synthesis and cell growth. However, recent data suggest that, in response to certain hypertrophic agents, signaling via Ras and MEK/Erk, as well as mTOR, is required for activation of protein synthesis, indicating new connections between these key signaling pathways. PMID- 15276466 TI - Cross-talk of opioid peptide receptor and beta-adrenergic receptor signalling in the heart. AB - Opioid peptide receptor (OPR) and beta-adrenergic receptor (beta-AR) are well established members of G-protein-coupled receptor (GPCR) superfamily and are involved in regulating cardiac contractility, energy metabolism, myocyte survival or death. OPRs are typical Gi/Go-coupled receptors and activated by opioid peptides derived from the endorphin, dynorphin and enkephalin families, whereas beta-AR stimulated by catecholamines is the model system for Gs-coupled receptors. While it is widely accepted that beta-AR stimulation serves as the most powerful means to increase cardiac output in response to stress or exercise, we have only begun to appreciate functional roles of OPR stimulation in regulating cardiovascular performance. Cardiovascular regulatory effects of endogenous opioids were initially considered to originate from the central nervous system and involved the pre-synaptic co-release of norepinephrine with enkephalin from sympathetic neuronal terminals in the heart. However, opioid peptides of myocardial origin have been shown to play important roles in local regulation of the heart. Notably, OPR stimulation not only inhibits cardiac excitation-contraction coupling, but also protects the heart against hypoxic and ischemic injury via activation of Gi-mediated signalling pathways. Further, OPRs functionally and physically cross-talk with beta-ARs via multiple hierarchical mechanisms, including heterodimerization of these receptors, counterbalance of functional opposing G protein signalling, and interface at downstream signalling events. As a result, the beta-AR-mediated positive inotropic effect and increase in cAMP are markedly attenuated by OPR activation in isolated cardiomyocytes as well as sympathectomized intact rat hearts. This brief review will focus on the interaction between beta-AR and OPR and its potential physiological and pathophysiological relevance in the heart. PMID- 15276467 TI - TGF-beta1 and angiotensin networking in cardiac remodeling. AB - The renin-angiotensin system (RAS) and transforming growth factor-beta1 (TGF beta1) play a pivotal role in the development of cardiac hypertrophy and heart failure. Recent studies indicate that angiotensin II (Ang II) and TGF-beta1 do not act independently from one another but rather act as part of a signalling network in order to promote cardiac remodeling, which is a key determinant of clinical outcome in heart disease. This review focuses on recent advances in the understanding, how Ang II and TGF-beta1 are connected in the pathogenesis of cardiac hypertrophy and dysfunction. Increasing evidence suggests that at least some of the Ang II-induced effects on cardiac structure are mediated via indirect actions. Ang II upregulates TGF-beta1 expression via activation of the angiotensin type 1 (AT1) receptor in cardiac myocytes and fibroblasts, and induction of this cytokine is absolutely required for Ang II-induced cardiac hypertrophy in vivo. TGF-beta induces the proliferation of cardiac fibroblasts and their phenotypic conversion to myofibroblasts, the deposition of extracellular matrix (ECM) proteins such as collagen, fibronectin, and proteoglycans, and hypertrophic growth of cardiomyocytes, and thereby mediates Ang II-induced structural remodeling of the ventricular wall in an auto /paracrine manner. Downstream mediators of cardiac Ang II/TGF-beta1 networking include Smad proteins, TGFbeta-activated kinase-1 (TAK1), and induction of hypertrophic responsiveness to beta-adrenergic stimulation in cardiac myocytes. PMID- 15276468 TI - Cross-regulation between the renin-angiotensin system and inflammatory mediators in cardiac hypertrophy and failure. AB - One of the major conceptual advances in our understanding of the pathogenesis of heart failure has been the insight that heart failure may progress as the result of the sustained overexpression of biologically active "neurohormones", such as norepinephrine and angiotensin II, which by virtue of their deleterious effects are sufficient to contribute to disease progression by provoking worsening left ventricular (LV) remodeling and progressive LV dysfunction. Recently, a second class of biologically active molecules, termed cytokines, has also been identified in the setting of heart failure. Analogous to the situation with neurohormones, the overexpression of cytokines is sufficient to contribute to disease progression in heart failure phenotype. Although important interactions between proinflammatory cytokines and the adrenergic system have been recognized in the heart for over a decade, the nature of the important interactions between proinflammatory cytokines and the renin-angiotensin system has become apparent only recently. Accordingly, in the present review, we will discuss the evidence which suggests that there is a functionally significant cross-talk between neurohormonal and inflammatory cytokine signaling in cardiac hypertrophy and failure. PMID- 15276469 TI - Interactions between the sympathetic nervous system and the cardiac natriuretic peptide system. AB - The sympathetic nervous system (SNS) and the cardiac natriuretic peptide system (NPS) are fundamentally important neurohumoral systems for cardiovascular regulation. Their mutual interactions have been subject to numerous experimental and human studies. In the current manuscript, results from in vivo and in vitro studies will be reviewed with a focus on sympathetic outflow on the control of natriuretic peptide release. PMID- 15276470 TI - Interference of antihypertrophic molecules and signaling pathways with the Ca2+ calcineurin-NFAT cascade in cardiac myocytes. AB - Cardiac hypertrophy occurs in a number of disease states associated with chronic increases in cardiac work load. Although cardiac hypertrophy may initially represent an adaptive response of the myocardium, ultimately, it often progresses to ventricular dilatation and heart failure. Much investigation has focused on the signaling pathways controlling cardiac hypertrophy at the level of the single cardiac myocyte. One prohypertrophic pathway that has received much attention involves the ubiquitously expressed Ca2+/calmodulin-activated phosphatase calcineurin. Upon activation by Ca2+, calcineurin dephosphorylates nuclear factor of activated T cell (NFAT) transcription factors, leading to their nuclear translocation. As common in complex biological systems, cardiac hypertrophy is controlled simultaneously by stimulatory (prohypertrophic) and counter-regulatory (antihypertrophic) pathways. Given the potent prohypertrophic effects of the Ca2+ calcineurin-NFAT pathway in cardiac myocytes, it is not surprising that the activity of this pathway is tightly controlled at multiple levels. Inhibitory mechanisms upstream (nitric oxide (NO), cGMP, cGMP-dependent protein kinase type I (PKG I), heme oxygenase-1 (HO-1), biliverdin, carbon monoxide (CO)) and downstream from calcineurin (glycogen synthase kinase-3 (GSK3), c-Jun N-terminal kinases (JNKs), p38 mitogen-activated protein kinase (MAPKs)) have been described. Moreover, several inhibitors directly target calcineurin enzymatic activity (cyclosporine A (CsA), tacrolimus (FK506), calcineurin-binding protein-1 (Cabin-1)/calcineurin-inhibitory protein (Cain), A-kinase-anchoring protein-79 (AKAP79), calcineurin B homology protein (CHP), MCIPs, VIVIT). Considering the dominant role of the calcineurin pathway in cardiac hypertrophy and failure, calcineurin-inhibitory strategies may lead to the identification of novel therapeutic approaches for patients with cardiac disease. PMID- 15276471 TI - Fibroblast growth factor 2 isoforms and cardiac hypertrophy. AB - Fibroblast growth factor 2 (FGF-2), a multifunctional polypeptide that affects cell growth and differentiation and becomes upregulated by stress, is expressed as AUG-initiated 18 kDa FGF-2 or CUG-initiated 21-34 kDa (hi-FGF-2) isoforms. Animal models have provided strong evidence that FGF-2 is essential for the manifestation of overload- and angiotensin-induced cardiac hypertrophy. Nevertheless, studies to-date have not discriminated between the activities of 18 kDa FGF-2 and hi-FGF-2. Our recent work has pointed to a potent pro-hypertrophic effect of added hi-FGF-2, and a pro-apoptotic effect of sustained intracrine hi FGF-2 signaling. In the future, it will be important to differentiate between the activities of the different FGF-2 isoforms in the context of adaptive and maladaptive myocardial hypertrophy and heart failure. Based on all available evidence, we propose that while the 18-kDa FGF-2 is a component of an adaptive trophic response, a switch to hi-FGF-2 accumulation would exacerbate hypertrophy and contribute to cell death, thus driving the myocardium towards a maladaptive phenotype. PMID- 15276473 TI - Role of Ca2+/calmodulin-dependent protein kinase II in cardiac hypertrophy and heart failure. AB - Ca2+/calmodulin-dependent protein kinase II (CaMKII), a critical transducer of Ca2+ signaling, is a multifunctional protein kinase which can phosphorylate a wide range of substrates and regulate numerous cellular functions. The delta isoforms of CaMKII predominate in the heart and two splice variants of CaMKIIdelta, deltaB and deltaC, have been demonstrated to be present in the adult mammalian myocardium. The deltaB isoform contains a nuclear localization signal (NLS) that is absent from deltaC, and consequently, the two isoforms have different subcellular localization. Recent work from our laboratory and others has implicated CaMKII in the development of cardiac hypertrophy and heart failure. The specific roles of these CaMKII isoforms in regulating cardiac function appear to be determined by their subcellular localization. The nuclear deltaB isoform plays a key role in hypertrophic gene expression, whereas the cytoplasmic deltaC isoform can affect excitation-contraction (E-C) coupling through phosphorylation of Ca2+ regulatory proteins and may also transduce signals leading to apoptosis. In addition, the nuclear deltaB and the cytoplasmic deltaC isoforms of CaMKII are differentially regulated in pressure overload induced cardiac hypertrophy. This review focuses on evidence that CaMKII plays an essential role in transcriptional activation associated with cardiac hypertrophy, as well as the aberrant Ca2+ handling and apoptosis that may contribute to heart failure. The hypothesis that CaMKII isoform selective activation, localization and substrate phosphorylation lead to specificity in the resultant signaling pathways is discussed. PMID- 15276472 TI - Calcineurin-NFAT signaling regulates the cardiac hypertrophic response in coordination with the MAPKs. AB - Prolonged cardiac hypertrophy of pathologic etiology is associated with arrhythmia, sudden death, decompensation, and dilated cardiomyopathy. In an attempt to understand the mechanisms that underlie the hypertrophic response, extensive investigation has centered on a characterization of the molecular pathways that initiate or maintain the pathologic growth of individual cardiac myocytes. While a large number of signal transduction cascades have been identified as critical regulators of cardiac hypertrophy, here the scientific evidence implicating the protein phosphatase calcineurin (PP2B) and the mitogen activated protein kinases (MAPK) as co-regulators of reactive hypertrophy will be discussed. Gain- and loss-of-function studies in genetically altered mice and in cultured cardiomyocytes have demonstrated the necessity and sufficiency of calcineurin to regulate pathologic cardiac hypertrophy. However, using similar approaches, the hypertrophic regulatory role attributed to various branches of the MAPK signaling pathway has been less conclusive, although a loose consensus suggests that the c-Jun N-terminal kinases (JNK) and p38 kinases function as mediators of dilated cardiomyopathy, while extracellular signal-regulated kinases (ERKs) function as regulators of hypertrophy. More recently, the actions of calcineurin and MAPK signaling pathways have been shown to be co-dependent such that unitary activation of calcineurin in myocytes leads to up-regulation in ERK and JNK signaling, but down-regulation in p38 signaling. Conversely, unitary activation of JNK or p38 in cardiac myocytes leads to down-regulation of calcineurin effectiveness by directly antagonizing nuclear factor of activated T cells (NFAT) nuclear occupancy. Thus, an emerging paradigm suggests that calcineurin-NFAT and MAPK signaling pathways are inter-dependent and together orchestrate the cardiac hypertrophic response. PMID- 15276474 TI - Myocyte hypertrophy and apoptosis: a balancing act. AB - In response to a variety of extrinsic and intrinsic stimuli that impose increased biomechanical stress the heart responds by enlarging the individual myofibers. Even though myocardial hypertrophy can normalize wall tension, it instigates an unfavorable outcome and threatens affected patients with sudden death or progression to overt heart failure, suggesting that in most instances hypertrophy is a maladaptive process. Increasing evidence suggests that several of the signaling cascades controlling myocyte growth in the adult heart also function to enhance survival of the myocyte population in response to pleiotropic death stimuli. In this review, we summarize recent insights into hypertrophic signaling pathways and their ability to control the balance between myocyte life and death. As modulation of myocardial growth by antagonizing intracellular signaling pathways is increasingly recognized as a potentially auspicious approach to prevent and treat heart failure, the design of such therapies should respect the dichotomous action of pathways that dictate a balance between myocyte hypertrophy, survival and death. PMID- 15276475 TI - Negative regulators of cardiac hypertrophy. AB - Throughout the past decade, much effort has been spent on deciphering the signaling pathways positively mediating cardiac hypertrophy. Recently, several endogenous molecules in the heart have been shown to negatively regulate cardiac hypertrophy. One group of these molecules is constitutively active at baseline, while molecules belonging to the second group serve as negative feedback regulators, which are activated in response to pathologic insults. Studies upon the negative regulators of cardiac hypertrophy may allow us to develop novel strategies to treat heart failure by mimicking the naturally preferred mechanisms to maintain homeostasis. In addition, the search for molecular targets of these negative regulators may allow us to identify novel positive mediators of hypertrophy. The aim of this article is to provide a brief overview of these newly identified negative regulators of cardiac hypertrophy. PMID- 15276476 TI - Gender modulates cardiac phenotype development in genetically modified mice. AB - Recent research using genetically modified mice has revealed significant sex differences in cardiac phenotypes. In the majority of strains, females display a lower mortality, less severe hypertrophy, better preserved function and mitigated cardiac pathology compared with male counterparts. Thus, gender is an independent determinant for the development of cardiac phenotype in murine models. While there is strong evidence for estrogen as a cardiac protector, emerging evidence indicates adverse actions of testicular hormones that might be responsible in part for the sex differences. Studies using mouse models have also revealed novel information on signalling mechanisms mediating the sex difference. PMID- 15276477 TI - Human cardiac inwardly rectifying current IKir2.2 is upregulated by activation of protein kinase A. AB - OBJECTIVE: The cardiac inwardly rectifying potassium current IK1 and its molecular correlates Kir2.1 and Kir2.2 play an important role in cardiac repolarisation and in the pathogenesis of hereditary long-QT syndrome (LQTS-7). Protein kinases A (PKA) and C (PKC) are key enzymes in adrenergic signal transduction, inducing arrhythmias in heart disease. This study investigated the regulation of Kir2.2 (KCNJ12) by PKA. METHODS: Cloned Kir2.2 channels were expressed heterologously in Xenopus oocytes and currents were measured with the double-electrode voltage-clamp technique. RESULTS: After activation of PKA by forskolin (100 micromol/l) or Ro-20-1724 (100 micromol/l), wild type currents at 120 mV were increased by 93.7% and 79.0%, respectively. Coapplication of the PKA inhibitor KT-5720 (2.5 micromol/l) attenuated this effect. No significant changes were apparent after mutation of the single PKA consensus site S430. In addition, removal of all four PKC consensus sites in Kir2.2 induced a phorbolester-mediated current increase which could be suppressed by PKA inhibitors H-89 (50 micromol/l) and KT-5720 (2.5 micromol/l). CONCLUSIONS: This study demonstrates antagonistic effects of PKA and PKC in the regulation of Kir2.2. Phosphorylation by PKC has been shown to cause an inhibition of Kir2.2 currents, whereas activation of PKA leads to current upregulation. PMID- 15276478 TI - Atrial natriuretic peptide modulates the hyperpolarization-activated current (If) in human atrial myocytes. AB - OBJECTIVES: The relationship between atrial stretching and changes in cell excitability is well documented. Once stretched, human atrial myocytes (HuAM) release atrial natriuretic peptide (hANP). Receptors for hANP (NPR) are coupled to a guanylyl cyclase (GC) activity, and are present on HuAM, but the electrophysiological effects of hANP are largely unknown. We investigated the effect of hANP on If, the hyperpolarization-activated current present in HuAM, and the underlying intracellular pathway. METHODS: HuAM were isolated from atrial appendages and utilized for patch-clamp recording. RESULTS: hANP caused a significant and concentration dependent shift of the midpoint activation potential (DeltaVh) toward less negative potentials of 6.9 +/- 1.0 mV at 0.1 nM; 13.0 +/- 2.6 mV at 1 nM and 15.3 +/- 2.2 mV at 10 nM (p < 0.001 for all); a parallel increase of If rate of activation occurred. The effect of hANP was completely blocked by isatin, a potent antagonist of NPR (p < 0.01 vs. hANP). In the presence of the inhibitors of guanylyl cyclase (ODQ and LY83583), hANP caused a significantly smaller DeltaVh (p < 0.01 vs. hANP for both). 8Br-cGMP mimicked the effect of hANP, both in the presence and absence of KT5823, a selective inhibitor of Protein kinase G. Pretreatment with pertussis toxin (PTX) did not change the effect of hANP, thus excluding a major role for the coupling of NPR with the Gi-Proteins system. Pretreating cells with cyclopentyladenosine (CPA), an A1-adenosine receptor agonist, completely blocked hANP effect. Adding hANP to maximal serotonin concentration produced an additive response. CONCLUSIONS: Our data demonstrate for the first time that ANP is able to increase If, likely through a modulation of intracellular cGMP and cAMP levels. This effect could have implications in the relationship between stretch and arrhythmogenesis in the human atrium. PMID- 15276479 TI - Contractile arrest reveals calcium-dependent stimulation of SERCA2a mRNA expression in cultured ventricular cardiomyocytes. AB - OBJECTIVE: Downregulation of sarco-endoplasmic reticulum calcium ATPase 2a (SERCA2a) expression is a critical marker of pathological myocardial hypertrophy. The effects of calcium-dependent signaling and of contractile activity on the regulation of myocardial SERCA2a expression remain unclear. The present study dissociates effects of calcium-dependent signaling through calcineurin (CN) and calmodulin dependent protein kinase-II (CAMK-II), from effects of contractile activity in spontaneously contracting rat neonatal ventricular cardiomyocytes (NVCM) using 2,3-butanedione monoxime (BDM), which arrests contractions but maintains calcium fluxes. METHODS: SERCA2a mRNA expression was analysed using Northern hybridisation in spontaneously contracting NVCM (control) and in NVCM treated with either BDM, L-type Ca2+-channel blocker (verapamil), CN-blocker (cyclosporin A; CsA), CAMK-II blocker (KN-93), or combinations thereof. Transient transfection of the CN-dependent transcription factor nuclear factor of activated T-lymphocytes (NFATc), coupled to GFP, was used to detect NFAT nuclear translocation. The effects of CN/CAMK-II-dependent signaling were further dissected into effects of the transcription factors NFATc4 and myocyte enhancer factor 2c (MEF2c) on the activity of various SERCA2a promoter fragments using transient transfection assays. RESULTS: Treatment with BDM induced a 2.5-fold rise in SERCA2a mRNA, which was abolished by addition of verapamil and was reduced by addition of CsA (-40%) and KN-93 (-20%). NFAT nuclear translocation was similar in control and BDM-treated NVCM. SERCA2a promoter activity was stimulated by NFATc4 and MEF2c, but only when both factors were co-transfected. CONCLUSION: Following contractile arrest with BDM, upregulation of SERCA2a mRNA expression by CN/CAMK-II signaling becomes evident. This upregulation is likely the result of synergistic stimulation of SERCA2a promoter activity by NFATc4 and MEF2c. Contractile activity opposes this upregulation through distinct and independent pathways. PMID- 15276480 TI - Heme oxygenase-1 inhibition of MAP kinases, calcineurin/NFAT signaling, and hypertrophy in cardiac myocytes. AB - OBJECTIVE: Heme oxygenases (HO) are the rate-limiting enzymes in heme degradation, catalyzing the breakdown of heme to equimolar quantities of biliverdin (BV), carbon monoxide (CO), and ferrous iron. The inducible HO isoform, HO-1, confers protection against ischemia/reperfusion (I/R)-injury in the heart. We hypothesized that HO-1 and its catalytic by-products constitute an antihypertrophic signaling module in cardiac myocytes. METHODS AND RESULTS: The G protein-coupled receptor (GPCR) agonist endothelin-1 (ET-1) (30 nmol/l) stimulated a robust hypertrophic response in cardiac myocytes isolated from 1- to 3-day-old Sprague-Dawley rats, with increases in cell surface area (planimetry), sarcomere assembly (confocal laser scanning microscopy), and prepro-atrial natriuretic peptide (ANP) mRNA expression. Adenoviral overexpression of HO-1, but not beta-galactosidase, significantly inhibited ET-1 induced cardiac myocyte hypertrophy. The antihypertrophic effects of HO-1 were mimicked by BV (10 micromol/l) and the CO-releasing molecule [Ru(CO)3Cl2]2 (10 micromol/l), strongly suggesting a critical involvement of BV and CO in the antihypertrophic effects of HO-1. Both BV and CO suppressed extracellular signal-regulated kinases (ERK1/ERK2) and p38 mitogen-activated protein kinase (MAPK) activation by ET-1 stimulation. Moreover, BV and CO inhibited the prohypertrophic calcineurin/NFAT pathway. This inhibition occurred upstream from calcineurin because BV and CO inhibited NFAT activation in response to ET-1 stimulation but not in response to adenoviral expression of a constitutively active calcineurin mutant. Upstream inhibition of the calcineurin/NFAT pathway by CO occurred independent from cGMP and cGMP-dependent protein kinase type I (PKG I). CONCLUSIONS: Heme oxygenase-1 and its catalytic by-products, BV and CO, constitute a novel antihypertrophic signaling pathway in cardiac myocytes. Biliverdin and CO inhibition of MAPKs and calcineurin/NFAT signaling provides a mechanistic framework how heme degradation products may promote their antihypertrophic effects. PMID- 15276481 TI - Inhibition of Ca2+-dependent PKC isoforms unmasks ERK-dependent hypertrophic growth evoked by phenylephrine in adult ventricular cardiomyocytes. AB - OBJECTIVE: The duration of extracellular signal-regulated kinase (ERK) activation and the ERK-dependency of hypertrophic growth differ between stimulation of alpha adrenoceptors or angiotensin II receptors. As both receptor systems activate different protein kinase C (PKC) isoforms, we hypothesized that PKC isoforms contribute to the specific effect of alpha-adrenoceptor stimulation. METHODS: Isolated adult ventricular cardiomyocytes from rats were used. Different PKC isoforms were inhibited either pharmacologically by six different PKC inhibitors or specifically downregulated by antisense oligonucleotides. ERK activation was determined by phosphorylation relative to total ERK. The rate of protein synthesis was determined by 14C-phenylalanine incorporation. RESULTS: The hypertrophic response of phenylephrine was inhibited in a concentration-dependent fashion by three different inhibitors of Ca2+-independent PKC isoforms (Go6983, rottlerin, Go6850), but not by three distinct PKC inhibitors directed preferentially against Ca2+-dependent PKC isoforms (Ro32-0432, HBDDE, Go6976). Antisense oligonucleotides directed against PKC-alpha, -delta, or -epsilon downregulated their specific isoforms. Their corresponding sense oligonucleotides did not affect PKC isoform expression. The phenylephrine-induced increase in protein synthesis was blocked by antisense oligonucleotides directed against PKC delta or PKC-epsilon but not PKC-alpha, confirming the pharmacological experiments. Inhibition of Ca2+-dependent PKC isoforms by HBDDE or Go6976 converted a transient activation of ERK by phenylephrine into a sustained response. Under these conditions, phenylephrine increased protein synthesis in an ERK-dependent way. CONCLUSION: Inhibition of Ca2+-dependent PKC isoforms converts the ERK-independent effect of phenylephrine on protein synthesis into an ERK dependent induction of protein synthesis. We conclude that co-activation of Ca2+ dependent PKC isoforms by phenylephrine contributes to the specific effect on adult ventricular cardiomyocytes from rat. PMID- 15276483 TI - Quo vadimus? PMID- 15276482 TI - Effects of alpha1-adrenergic stimulation on normal and hypertrophied mouse hearts. Relation to caveolin-3 expression. AB - BACKGROUND: Modulation of the transduction efficiency through G-protein coupled receptors, caused by external stimulation, is essential in designing antihypertrophic treatment strategies in the dysfunctional heart. We compared protein-kinase C (PKC)-dependent regulation of positive inotropic effect via alpha1-adrenoreceptor (ADR)/Gq protein in hyperdynamic versus hypertrophied myocardium. METHODS: Inotropic (work performing isolated heart) and cellular effects of alpha1-adrenoreceptor stimulation were studied in nontransgenic (Ntg) and transgenic (Tg) mice with cardiac specific overexpression of L-type voltage dependent calcium channels (L-type VDCC). RESULTS: Transgenic hyperdynamic and hypertrophic myocardium (due to overexpression of the L-type VDCC alpha1 subunit) were characterized by a lack of positive inotropic effect (PIE) to alpha1-ADR stimulation with phenylephrine (PE), as compared to a positive response in Ntg hearts. This was partially restored by PKC inhibition with chelerythrine and staurosporine only at the hyperdynamic stage. The inability of PKC inhibition to increase positive inotropy was associated with markedly decreased cardiac specific caveolin-3 expression, and no changes in Galphaq, PLC-beta1, caveolin-1 and alpha1-adrenoreceptor expression. CONCLUSION: In the hyperdynamic myocardium, PKC activation may be one of the switches responsible for an impaired alpha1 adrenergic positive inotropic response. In the hypertrophied myocardium, the interruption of the transduction from Galphaq-protein coupled receptors to downstream effectors may be due to the down-regulation of caveolin-3 expression. PMID- 15276484 TI - Pneumonectomy for nontuberculous mycobacterial infections. AB - BACKGROUND: Pneumonectomy is considered in the treatment of nontuberculous mycobacterial infections when an entire lung is affected. However, this procedure carries high morbidity. We report on our experience in using pneumonectomy for treating patients with nontuberculous mycobacterial infections. METHODS: Between 1983 and 2002, 53 patients infected with nontuberculous mycobacteria underwent 55 pulmonary resections. Of these patients, 11 (3 men, 8 women) underwent pneumonectomy (5 right, 6 left). Median age was 57 years (range, 43 to 69 years). Mycobacterium avium complex disease occurred in 10 patients and Mycobacterium abscessus disease in 1. Indications for pneumonectomy included multiple cavities in one lung and destruction of an entire lung. The bronchial stump was covered with a latissimus dorsi muscle flap in 7 patients and with an intercostal pedicle flap in 2. RESULTS: Operating time ranged from 142 to 477 minutes (median, 360 minutes). The median intraoperative blood loss was 555 mL (range, 130 to 1,245 mL). There was no operative mortality. Bronchopleural fistula occurred in 3 patients. All fistulas were observed after right pneumonectomy, and were treated by reclosure of the bronchus. Empyema occurred in 1 patient, who was treated with irrigation. All patients achieved sputum-negative status after surgery. Two late deaths occurred. One patient died of respiratory failure 11 months after surgery. A second patient, the only patient who had recurrent disease, died of respiratory failure 4 years postoperatively. CONCLUSIONS: Despite bronchial stump protection, right pneumonectomy carries a risk for bronchopleural fistula. Nonetheless, pneumonectomy can result in high cure rates in patients with nontuberculous mycobacterial infections. PMID- 15276485 TI - Transcervical-subxiphoid-videothoracoscopic "maximal" thymectomy--operative technique and early results. AB - BACKGROUND: The operative technique of a transcervical-subxiphoid videothoracoscopic "maximal" thymectomy without sternotomy is described and the early results of the follow-up of patients operated on are analyzed. METHODS: One hundred "maximal" transcervical-subxiphoid-videothoracoscopic thymectomies were performed for nonthymomatous myasthenia gravis during a recent 32-month period (from September 1, 2000 to May 8, 2003). Patient characteristics, complications, pathologic findings, and the results of follow-up were analyzed. RESULTS: The study group included 83 women and 17 men. The mean age was 29.8 years (range, 10 69 years). The mean preoperative duration of myasthenia was 2.73 years (range, 3 months to 17 years). The preoperative Osserman score was I-III, 27 patients were taking steroids preoperatively. Eleven operations were performed by two teams working simultaneously and 89 operations were performed by one surgeon including four combined thymectomy-thyroid operations in patients with myasthenia and thyroid nodules. The mean operative time for two-team approach thymectomies was 159.09 minutes (range, 140-170 minutes) and the mean operative time for the thymectomy performed by one surgeon was 199.41 minutes (range, 150-270 minutes) (p = 0.0004). There was a 15.0% (15 out of 100) postoperative morbidity and no mortality. Foci of ectopic thymic tissue were found in 71.0% of the patients and were most prevalent in the perithymic fat (37.0%) and in the aorta-pulmonary window (33.0%). The mean weight of the specimen was 78.4 g (range, 14.5-253.0 g). In 48 patients followed-up for 12 months, the improvement rate was 83.3%, the no improvement rate was 14.6%, and 1 patient died during the follow-up period. Complete remission rates were 18.8% and 32.0% after 1 and 2 years of follow-up, respectively. CONCLUSIONS: We conclude that the "maximal" transcervical subxiphoid-videothoracoscopic thymectomy is a safe operative technique, avoiding a sternotomy, performed partly in an open fashion with the extensiveness comparable with the transsternal extended and "maximal" thymectomies. The two team approach helps to reduce the operative time. However, because of the limited time of follow-up it is too early for the final assessment of the long-term results of this method in the treatment of myasthenia gravis. PMID- 15276487 TI - Atypical thymoma: a report of seven patients. AB - BACKGROUND: Most thymic neoplasms fall under the designation of thymoma, consisting of well-differentiated epithelial cells, resembling normal thymus. At the opposite spectrum are thymic carcinomas; the cell of origin while similar is malignant. Recently a third category of thymic neoplasms, atypical thymomas, has been recognized representing thymic neoplasms manifesting atypia although without overt cytomorphologic criteria of malignancy. METHODS: Seven patients with a diagnosis of atypical thymoma were encountered over a 6-year period from the patient files of the cardiothoracic division of The Ohio State Medical Center. RESULTS: In all patients there was gross or light microscopic invasive disease with involvement of the capsule, phrenic nerve, diaphragm, chest wall, and lung. Surgical extirpation/de-bulking along with radiation therapy in six and chemotherapy in one led to complete disease regression. Intrathoracic recurrences developed in 4 involving lung, pleura, chest wall and diaphragm. All patients are well. CONCLUSIONS: Atypical thymomas are locally aggressive tumors with a high incidence of intrathoracic recurrence; extrathoracic spread is not seen. Our study corroborates other reports that death attributable to atypical thymoma is uncommon. PMID- 15276488 TI - Axillary thoracotomy versus videothoracoscopy for the treatment of primary spontaneous pneumothorax. AB - BACKGROUND: A prospective, randomized study was carried out on patients with primary spontaneous pneumothorax, with the aim of determining if video-assisted thoracoscopy is superior to axillary thoracotomy in the surgical treatment of this condition. METHODS: Patients were randomly assigned to two groups; video assisted thoracoscopy (group A; n = 46) and axillary thoracotomy (group B; n = 44). All fit the established criteria for surgical indication (relapse or persistent air leakage after pleural drainage). In all cases the treatment consisted of apical segmentectomy of the blebs or dystrophic complex and pleural mechanical abrasion. The study evaluated the following factors: postoperative blood loss, respiratory function (maximum inspiratory and expiratory pressures, forced expiratory volume in the first second and forced vital capacity), postoperative pain (analog visual scale), supplementary doses of analgesics, postoperative complications, hospital stay, and resumption of normal activity. Relapses were evaluated for the minimum period of time of two years. RESULTS: No significant differences were found in any of the factors studied in either group. CONCLUSIONS: Video-assisted thoracoscopy and axillary thoracotomy offer similar results in the surgical treatment of primary spontaneous pneumothorax. The rate of complication is low and the level of pain is acceptable without long-term sequelae. PMID- 15276489 TI - Repair of the pectus deformity: results of the Ravitch approach in the current era. AB - BACKGROUND: Recent publications have advocated a minimally invasive approach to repair of the pectus deformity. Efforts to evaluate this new approach have been hampered by lack of comparative information regarding outcomes of the standard Ravitch approach. We use a modified Ravitch procedure, and present our series as a basis for comparison. METHODS: Records of 69 consecutive patients undergoing repair of the pectus deformity were retrospectively reviewed. Modifications included a minimal incision and a new technique to address sternal angulation. A patient satisfaction survey evaluated the patients' perception of the outcome. RESULTS: We found one wound infection (1.4%). Five patients (7.2%) had a seroma, and were treated as outpatients. Because the minimally invasive approach is used for pectus excavatum, we divided our series into excavatum and carinatum subsets. The subset of 44 pectus excavatum patients had a mean postoperative length of stay (LOS) of 2.9 days. The median patient satisfaction score was 4 on a scale of 1 to 5, at an average of 4.75 years after repair. The subset of 25 pectus carinatum patients had a mean LOS of 2.4 days and a median patient satisfaction score of 5. CONCLUSIONS: The modified Ravitch procedure yields excellent results with low morbidity, hospital LOS, and cost, combined with high patient satisfaction. These current data will be useful for comparison as newer techniques for pectus repair continue to evolve. PMID- 15276490 TI - Severity of compensatory sweating after thoracoscopic sympathectomy. AB - BACKGROUND: Compensatory sweating is a well-known side effect after sympathectomy for hyperhidrosis. It is often claimed to correlate with the extent of sympathectomy, but results from the literature are conflicting, and few have actually considered differences in the intensity of compensatory sweating. METHODS: A total of 158 patients underwent thoracoscopic sympathectomy for primary hyperhidrosis or blushing, or both. Sympathectomy was performed bilaterally at Th2 for facial hyperhidrosis/blushing (n = 49), Th2-3 for palmar hyperhidrosis (n = 62), and Th2-4 for axillary hyperhidrosis (n = 47). RESULTS: Follow-up by questionnaire was possible in 94% of patients after a median of 26 months. Compensatory sweating occurred in 89% of patients and was so severe in 35% that they often had to change their clothes during the day. The frequency of compensatory sweating was not significantly different among the three groups, but severity was significantly higher after Th2-4 sympathectomy for axillary hyperhidrosis (p = 0.04). Gustatory sweating occurred in 38% of patients, and 16% of patients regretted the operation. CONCLUSIONS: Compensatory and gustatory sweating were remarkably frequent side effects after thoracoscopic sympathectomy for primary hyperhidrosis. We found no significant difference between the level of sympathectomy and the occurrence of compensatory sweating. However, it appears that this is the first study to demonstrate that severe sweating is significantly more frequent after Th2-4 sympathectomy for axillary hyperhidrosis. We encourage informing patients thoroughly about these side effects before surgery. PMID- 15276491 TI - The radiologic appearance of intercostal muscle flap. AB - BACKGROUND: The intercostal muscle flap (ICMF) is commonly used in airway and esophageal surgery to reinforce an anastomosis or site of closure. These flaps undergo heterotopic ossification that may result in stenosis of adjacent airways or the esophagus. We evaluated the computer tomography (CT) scan, technetium-99m methylene diphosphonate bone scan and positron emission tomography with 2-[18F] fluoro-2-deoxy-D-glucose (FDG-PET) findings of ICMF and the frequency of airway or esophageal stenosis. METHODS: A retrospective review was made of the radiologic records of 23 patients (9 women, 14 men) who underwent ICMF. The CT scans were obtained a mean of 36 months (range, 1 week to 58 months) after surgery and the size, morphology, and density of the ICMFs were recorded. Correlative bone scan in 13 patients and FDG-PET scans in 11 patients were reviewed. RESULTS: A discontinuous, thin, linear calcified stripe or parallel stripes (mean thickness, 4 mm; mean density, 430 Houndsfield unit [HU]) were present in all patients on CT. The flap contained fat density (mean, -59 HU) in 18 patients and soft tissue density (mean, 41 HU) in 8 patients and measured about 1 cm in thickness. The appearance of ICMF is characteristic when the ossification extends from the posterolateral chest wall to an adjacent bronchial stump. There was no increased uptake on bone scan or FDG-PET scan. None of the patients had airway or esophageal stenosis. CONCLUSIONS: The ICMF manifests on CT as a thin, linear calcified stripe or parallel stripes with central fat or soft tissue density. Airway stenosis due to ICMF is likely quite rare. We did not detect any airway stenosis. PMID- 15276492 TI - Interleukin-4 receptor cytotoxin as therapy for human malignant pleural mesothelioma xenografts. AB - BACKGROUND: Malignant pleural mesothelioma (MPM) is an uncommon but highly fatal neoplasm for which only limited treatment is available. METHODS: Immunohistochemical analysis was used to determine the expression of interleukin 4 receptors (IL-4R) on mesothelioma cell lines and resected mesothelioma tumors. Radioreceptor binding assays were used to show that these IL-4R were high affinity receptors. Previously, we had shown that a chimeric protein composed of a circularly permuted IL-4 molecule fused to a truncated form of Pseudomonas exotoxin A, IL-4(38-37)-PE38KDEL, could be used to kill IL-4R-bearing tumor cells in vitro. The toxicity of this molecule to mesothelioma cell lines was tested using a protein synthesis inhibition assay. A human mesothelioma xenograft model was then developed to assess the efficacy of this molecule in vivo. RESULTS: All MPM cell lines tested were found to express high-affinity cell-surface IL-4R. Immunohistochemical analysis of resected mesothelioma tumor specimens from 13 patients revealed that all tumors expressed moderate-to-high levels of IL-4R. Coculture of malignant mesothelioma cell lines with IL-4(38-37)-PE38KDEL resulted in a dose-dependent inhibition of tumor cell protein synthesis through an interaction with cell-surface IL-4R. In a nude mouse xenograft model of human MPM, intratumoral administration of IL-4(38-37)-PE38KDEL mediated a dose dependent decrease in tumor volume and a dose-dependent increase in survival. CONCLUSIONS: The chimeric protein, IL-4(38-37)-PE38KDEL, has potent antitumor effects against MPM both in vitro and in vivo. PMID- 15276493 TI - Expansion of chondrocytes in a three-dimensional matrix for tracheal tissue engineering. AB - BACKGROUND: The generation of autologous tracheal implants by tissue-engineering techniques is a promising concept for otherwise untreatable patients. A functional cartilaginous backbone represents a prerequisite for any bioartificial tracheal graft. The aim of this study was to define suitable cell types and culture conditions for the generation of tracheal cartilage. METHODS: We obtained tracheal, costal, and auricular cartilage from porcine donor animals (n = 10). The chondrocytes were cultured two-dimensionally in cell flasks or mixed with a liquid collagen solution forming a three-dimensional culture system. Labeling with carboxy fluorescein diacetate succinimidyl ester (CFDA SE) and biochemical reduction of formazan served to determine cell viability and proliferation. The extracellular matrix produced by the chondrocytes was characterized by Western blot. RESULTS: The CFDA SE labeling proved viability and the MTT assays documented a proliferation of the chondrocytes over time in vitro. While the chondrocytes in the three-dimensional cell culture system produced hyaline cartilage composed of collagen II, the two-dimensional culture conditions resulted in nonspecific collagen synthesis. CONCLUSIONS: Chondrocytes grown in a three-dimensional matrix can effectively proliferate and produce cartilage and are viable for more than 2 weeks. Costal chondrocytes are suitable for tracheal cartilage tissue engineering. PMID- 15276495 TI - Integrin-dependent protein tyrosine phosphorylation is a key regulatory event in collagen-IV-mediated adhesion and proliferation of human lung tumor cell line, Calu-1. AB - BACKGROUND: The clinical phenomenon of lung cancer metastasis to specific target organs is believed to be a direct interaction between tumor cells and extracellular matrix components. During invasion, tumor cells attach to the basement membrane protein, collagen type IV, degrade it, migrate through blood vessels, and adhere to extracellular matrix proteins. METHODS: Four nonsmall-cell lung cancer cells were tested for adhesion, proliferation, migration and morphologic alterations on collagen type IV matrix by immunoprecipitation, Western blotting, phase contrast and time lapse video microscopy. RESULTS: Collagen type IV promoted Calu-1 cell adhesion (< 15 minutes) and motility (< 6 hours) without any significant effect on proliferation. beta(1)-integrin is essential for collagen type IV adhesion and 8 to 10 fold higher expression of beta1-integrin on the surface of Calu-1 cells was identified. A protein tyrosine phosphatase inhibitor, peroxyvanadate, caused 50% inhibition in the adhesion process within 5 minutes but exposure to 60 micromol/L genistein for 72 hours, a protein tyrosine kinase inhibitor, drastically inhibits Calu-1 cell proliferation (> 70%). We examined the influence of beta1-integrin, peroxyvanadate and genistein on the spreading morphogenesis of Calu-1 cells on Collagen type IV. Use of either 1 microg of anti beta1-integrin inhibitory antibody (P5D2), 100 micromol/L peroxyvanadate or 60 micromol/L genistein had dramatic influence on the spreading of Calu-1 cells. Finally, Focal adhesion kinase was identified as a phosphoprotein target. CONCLUSIONS: We have characterized an in vitro model of matrix-specific binding of a lung cancer cell line, Calu-1 to Coll IV. Calu-1 cells use primarily a beta1-integrin mediated intracellular tyrosine phosphorylation phenomenon as the key regulatory mechanism for its binding advantage to Coll IV matrix. PMID- 15276496 TI - Adjunctive transmyocardial revascularization: five-year follow-up of a prospective, randomized trial. AB - BACKGROUND: In a prospective, randomized trial involving 263 patients who would be incompletely revascularized by coronary artery bypass grafting (CABG) alone, CABG plus transmyocardial revascularization (CABG/TMR) provided an early mortality benefit with similar angina relief compared with CABG alone at 1 year. We evaluated the long-term outcome of patients randomized to CABG/TMR or CABG alone. METHODS: Thirteen centers that enrolled 83% (218/263) of the patients in the original trial participated in this longitudinal study. Between 1996 and 1998, these centers randomized 218 patients who would be incompletely revascularized by CABG alone because of diffusely diseased target vessels to either holmium:yttrium-aluminum-garnet (holmium:YAG) CABG/TMR (n = 110) or CABG alone (n = 108). Baseline demographics and operative characteristics were similar between groups. Follow-up (mean 5.0 +/- 1.7 years) included survival and blinded angina class assessment. RESULTS: At this 5-year follow-up both groups experienced significant angina improvement from baseline, however, the CABG/TMR group had a lower mean angina score (0.4 +/- 0.7 vs 0.7 +/- 1.1, p = 0.05), a significantly lower proportion of patients with severe angina (class III/IV: 0% [0/68] vs 10% [6/60], p = 0.009), and a trend towards greater number of angina free patients (78% [53/68] vs 63% [38/60], p = 0.08), compared with CABG alone patients. Kaplan-Meier survival at 6 years was similar between CABG/TMR and CABG alone patients (76% vs 80%, p = 0.90). CONCLUSIONS: Five-year follow-up of prospectively randomized patients who would be incompletely revascularized because of diffuse coronary artery disease indicates that the addition of TMR to conventional CABG provides superior angina relief compared to CABG alone. PMID- 15276497 TI - Survival in patients with peripheral vascular disease after percutaneous coronary intervention and coronary artery bypass graft surgery. AB - BACKGROUND: Patients with peripheral vascular disease (PVD) undergoing coronary revascularization have high rates of adverse outcomes. Whether there are important differences in outcomes for surgical versus percutaneous coronary revascularization is unknown. The objective of this study was to compare survival in patients with PVD who underwent percutaneous coronary intervention (PCI) or coronary artery bypass grafting (CABG) surgery for multivessel coronary artery disease. METHODS: In-hospital data were collected on 1,305 consecutive patients undergoing coronary revascularization (PCI, n = 341; CABG, n = 964) in northern New England from 1994 to 1996. Patient records were linked to the National Death Index to assess survival out to 3 years (mean 1.2 years). Logistic and Cox proportional hazards regression were used to calculate risk-adjusted odds ratios and hazard ratios. RESULTS: Compared with CABG patients, those undergoing PCI were more often women, had more renal failure, more prior coronary revascularizations, were more likely to have two-vessel coronary artery disease and were more likely to undergo the procedure emergently. They were less likely to have a history of heart failure. After adjusting for differences in baseline characteristics, patients undergoing CABG had better intermediate survival than did PCI patients (hazard ratio 0.68; 95% confidence interval, 0.46 to 1.00; p = 0.05). CONCLUSIONS: Patients with multivessel coronary artery disease and PVD undergoing CABG surgery have better intermediate survival out to 3 years than similar patients undergoing PCI. This information may be useful in counseling patients with PVD requiring coronary revascularization. PMID- 15276499 TI - Comparison of bilateral thoracic artery grafting with percutaneous coronary interventions in diabetic patients. AB - BACKGROUND: This study compares the outcome of percutaneous coronary interventions (PCI) with bilateral internal thoracic grafting (BITA) in diabetic patients. METHODS: From May 1996 to December 1999, 802 consecutive diabetic patients underwent myocardial revascularization: 363 by PCI and 439 by BITA. The two groups were similar; however, left main disease (28% versus 3.3%), ejection fraction less than 0.35 (14.5% versus 5.5%), and chronic obstructive lung disease (8.4% versus 3%) were more prevalent in the BITA group, and prior percutaneous transluminal coronary angioplasty, in the PCI group (16.8% versus 10.5%). RESULTS: The number of coronary vessels treated per patient was higher in the BITA group (3.4 versus 1.2; p < 0.001). Thirty-day mortality was similar: 3.4% in the BITA group and 2.8% in the PCI group. Late follow-up (3 to 6.5 years) showed decreased return of angina (11% versus 64%; p < 0.001), fewer reinterventions (2.7% versus 55%; p < 0.001), and increased cardiovascular event-free survival (80% versus 30%; p < 0.001) in the BITA group. Six-year survival of BITA and PCI patients was 85.5% and 81.2%, respectively (not significant). However, survival of the subgroups of patients with left main or three-vessel coronary artery disease was significantly better with BITA (86% versus 76%; p = 0.003). CONCLUSIONS: Despite higher risk profile of diabetic patients treated surgically by BITA, their late outcome is better than that of patients treated by PCI. The results of this study support referring diabetics with single-vessel or double vessel disease to PCI and those with three-vessel and left main coronary artery disease to surgery. PMID- 15276501 TI - Midterm results of coronary artery bypass graft surgery with internal thoracic artery under low free-flow conditions. AB - BACKGROUND: We use the left internal thoracic artery (LITA) even when flow is very low. In this study, we investigated midterm outcome of coronary artery bypass graft surgery with low free-flow LITA. METHODS: One hundred patients undergoing coronary artery bypass graft surgery using LITA with a free flow of less than 20 mL/min were reviewed. The mean follow-up duration was 47.4 months, ranging from 1 to 65 months. Graft angiography was performed postoperatively. The diameter of the LITA was assessed angiographically. Cumulative graft patency, cardiac-related event-free rate, and actuarial survival rate were calculated by the Kaplan-Meier method. RESULTS: An early postoperative Doppler study showed that the diastolic-to-systolic ratio in the LITA was 1.76 +/- 0.33. A 1-month postoperative angiography revealed LITA string sign in 2 patients. One had a percutaneous coronary intervention, whereas string sign was not detected in the second patient, and LITA patency showed a marked improvement in 1-year postoperative angiogram. One month postoperatively the LITA diameter was 1.6 +/- 0.4 mm, and significantly enlarged in the second angiogram (1.9 +/- 0.4 mm, p = 0.0003). There was a significant correlation between the diameter of the LITA and the left anterior descending coronary artery (r = 0.889, p = 0.0001). The cumulative graft patency rate at 1 and 4 years was 99.0% and 94.3%, respectively. The cardiac-related event-free rate at 1 and 5 years was 97.0% and 93.3%, respectively. The actuarial survival rate at 5 years was 97.1%. CONCLUSIONS: Even with a very low LITA free flow, graft function improves with LITA growth, if there was no mechanical damage that impedes recovery. PMID- 15276502 TI - Competitive flow in arterial composite grafts and effect of graft arrangement in Off-Pump coronary revascularization. AB - BACKGROUND: We sought to investigate the incidence of competitive flow in arterial composite grafts and to delineate the effect of the location of moderately stenotic branch, the extent of the revascularized territories and the arrangement of in situ and free arterial grafts in off-pump coronary artery bypass grafting (OPCAB). METHODS: Three hundred eighteen patients who underwent OPCAB with aorta no-touch technique using the composite graft with totally arterial materials between December 2000 and March 2003 were studied. A total of 362 composite grafts were used. We reviewed their coronary angiography before and early after operation. Competitive flow was defined as the phenomenon that at least one of the distal anastomotic sites of the composite graft was not opacified in in situ graft angiography, but clearly opacified in native coronary angiography. The number of distal anastomoses was 3.47 +/- 0.93 per patient and 2.87 +/- 0.81 per composite graft. RESULTS: Early patency rate of the distal anastomotic sites of composite grafts was 98.7%. Competitive flow was found in 53/362 (14.6%) composite grafts, and graft occlusion occurred in 13/362 (3.6%) composite grafts. In the multivariate analysis of 362 composite grafts, 75% stenosis in right coronary artery (RCA) territory (p < 0.0001) and the number of distal anastomoses (p = 0.004) were significant predictors of competitive flow and graft occlusion. Multivariate analysis of 318 patients demonstrated that 75% stenosis in RCA territory (p < 0.0001) and the total number of distal anastomoses (p = 0.003) were statistically significant predictors of competitive flow and graft occlusion. The use of more than two in situ grafts and the shape of composite graft (branched or straight) did not have significant correlation with the outcome. CONCLUSIONS: Coronary artery revascularization using composite arterial grafts provided satisfactory early patency rates with an acceptable incidence of competitive flow. Because the implication of competitive flow in an arterial composite graft may differ from that in conventional bypass grafts unpredictably, long-term follow-up is mandatory. PMID- 15276503 TI - Impact of multivessel coronary artery disease on outcome after isolated minimally invasive bypass grafting of the left anterior descending artery. AB - BACKGROUND: The outcome in patients treated surgically for coronary artery disease is known to be influenced by the extent of the disease. Whether this factor also has an effect in patients undergoing isolated minimally invasive revascularization of the left anterior descending (LAD) artery using the internal thoracic artery (ITA) (MIDCAB) has not been looked at. Thus, this study sought to evaluate the impact of multivessel disease (MVD) on midterm outcome after MIDCAB. METHODS: From 1996 to 1999, 411 patients received a MIDCAB at our institution and were now followed up. Isolated disease of the LAD (SVD -single vessel disease) was presented in 262 patients (63.7%) and 149 patients (36.3%) had MVD at the time of operation. The reasons for apparent incomplete revascularization in patients with MVD were very small target vessels (< 1.0-mm diameter), stenoses of less than 50%, distal localization of the stenoses, long-term patency after angioplasty, or an extensive risk for sternotomy and(or) cardiopulmonary bypass. The midterm outcome was evaluated by questionnaires sent to the patients and their physicians. RESULTS: The mean follow-up was 29.4 +/- 11.1 months. The incidence of myocardial infarction was significantly higher in MVD as compared to SVD patients (8.1% vs 1.9%, p = 0.04). Patients with MVD had significantly more subsequent percutaneous transluminal coronary angioplasty (10.7% vs 5.3%, p = 0.049) and a similar number of repeat surgical revascularizations as compared to SVD patients. Patients with MVD had a significantly higher total 3-year mortality as compared to SVD patients by Kaplan-Meier estimate (8.7% vs 3.1%, relative risk [RR] = 2.56, p = 0.011). The 3-year cardiac mortality was significantly higher in patients with MVD as compared to SVD (4.0% vs 0.4%, RR = 9.48, p = 0.0054). After adjustment of baseline characteristics by Cox regression analysis, the 3-year risk of cardiac death was significantly higher in the MVD groups (RR = 2.2, confidence interval [CI] 95%: 1.8 to 4.65, p = 0.029). CONCLUSIONS: Patients with isolated disease of the LAD appear to benefit from ITA grafting in the form of a MIDCAB procedure. Here, it should be an approach of choice. The results show that MVD is an independent risk factor for outcome in patients undergoing a MIDCAB procedure. Nevertheless, the midterm morbidity and mortality in MVD patients after a MIDCAB procedure where the LAD is the only target vessel for interventional or surgical treatment is acceptable despite a higher morbidity than in SVD patients. PMID- 15276504 TI - Bradykinin preconditioning in coronary artery bypass grafting. AB - BACKGROUND: Experimental studies have shown that activation of bradykinin B2 receptor is one of the most important triggers of ischemic preconditioning. However, the effect of exogenous administration of bradykinin in cardiac surgery is not yet known. The present prospective randomized study was designed to investigate the effect of bradykinin pretreatment in patients undergoing elective coronary artery bypass surgery. METHODS: Forty-one patients with multiple-vessel coronary artery disease and stable angina, admitted for the first time for elective coronary artery bypass surgery, were randomized into control or bradykinin (BK) groups. Patients in the BK group received bradykinin infusion for 7 minutes (total dose 25 microg) before the initiation of cardiopulmonary bypass. Perioperative cardiac specific troponin I (cTnI) and creatine kinase cardiac isoenzyme (CKMB) release and hemodynamics were recorded. RESULTS: Bradykinin infusion caused acute decrease of blood pressure in most of the cases and the mean minimum mean blood pressure during bradykinin infusion was 72.7% of the original mean blood pressure (MBP) level (74.7 +/- 7.9 vs 54.4 +/- 12.1 mm Hg, p < 0.01). There were no differences in baseline levels of cTnI and CKMB between the groups. The postoperative cTnI levels were lower than 10 ng/mL in most patients in both groups (18 in the BK group and 15 in the control group). There was no difference in cTnI between the groups. However, patients who received bradykinin released significantly less CKMB than did the controls postoperatively (6 hours, BK, 22.1 +/- 9.5 vs control, 23.6 +/- 12.7 U/L; 12 hours, BK, 19.4 +/- 12.4 vs control, 28.7 +/- 23.8 U/L; 24 hours, BK, 21.5 +/- 14.7 vs control, 35.5 +/- 28.9 U/L; 48 hours, BK, 14.4 +/- 7.5 vs control, 23.5 +/- 13.6 U/L; analysis of variance [ANOVA] for repeated measurement, p = 0.036). Maximum CKMB was also lower in the BK group (22.4 +/- 14.4 vs 37.7 +/- 27.5 U/L, p = 0.044). There was no significant difference between the groups in any of the hemodynamic variables. CONCLUSIONS: Exogenous bradykinin infusion showed weak cardioprotective effect in the low-risk patients undergoing coronary artery bypass surgery but the dose used in the study caused acute decrease of systemic blood pressure. PMID- 15276505 TI - Combined off-pump coronary artery bypass surgery and pulmonary resection. AB - BACKGROUND: Combined cardiac surgery and pulmonary resection using cardiopulmonary bypass (CPB) has been described previously. There are a few reports of combined procedures done without using CPB. Off-pump coronary artery bypass grafting (OPCABG) eliminates organ dysfunction and suppression of immune system related to extracorporeal circulation. METHODS: Six patients underwent combined OPCABG and lung resection during a 4-year period. Follow-up ranging from 9 months to 3 years is available for these patients. RESULTS: Malignant pathology was the diagnosis in 5 patients and 1 patient was diagnosed with advanced chronic obstructive pulmonary disease. Right upper lobectomy was performed in 3 patients, left upper lobectomy was performed in 1 patient, right upper and middle bilobectomy was performed in 1 patient, and bilateral lung volume reduction was performed in one patient. Prolonged air leak occurred in 1 patient postoperatively and another patient experienced small right-sided residual pleural space that was resolved at 6 weeks follow-up. There were no operative deaths but there were 2 late deaths. Evidence of recurrence for angina or malignancy upon follow-up was not detected. CONCLUSIONS: A combined procedure is a safe approach in patients diagnosed with concomitant coronary artery and pulmonary disease. Avoidance of CPB may decrease the incidence of postoperative complications. PMID- 15276506 TI - Intraoperative angiography leads to graft revision in coronary artery bypass surgery. AB - BACKGROUND: Graft anastomosis quality in coronary artery bypass surgery can be assessed by intraoperative angiography. The aim of the present study was to quantify the on-table revision rate initiated by intraoperative angiography. METHODS: Intraoperative angiography was carried out in 186 patients undergoing coronary artery bypass surgery, with a total of 427 grafts. The operation was performed on-pump in 34%, off-pump through a sternotomy in 49%, and as a minimally invasive direct coronary bypass grafting (MIDCAB) procedure in 17%. The angiography was performed intraoperatively while the patients were still in general anesthesia, with the possibility for on-table revision. Follow-up angiography was carried out after a mean of 346 days. RESULTS: Eighteen of 427 grafts (4.2%) were revised due to the findings at intraoperative angiography. Revision rate after on-pump surgery was 1.1%, after off-pump through a sternotomy 6.4%, and after MIDCAB 6.5%. In 6 patients the lesions were located at the distal anastomoses and in 12 patients in the conduit. All but one was successfully revised, and at 1-year follow-up all these 17 grafts were patent. CONCLUSIONS: Intraoperative angiography saves a potential number of grafts that otherwise could have been occluded. An increased implementation of intraoperative quality assessment in coronary artery bypass surgery can lead to improved outcome. PMID- 15276508 TI - A prospective randomized study to evaluate stress response during beating-heart and conventional coronary revascularization. AB - BACKGROUND: Cardiopulmonary bypass (CPB) is associated with a systemic stress hormonal response, which can lead to changes in hemodynamics and organ perfusion. We examined perioperative stress hormone release in low-risk patients undergoing coronary artery bypass grafting with and without cardiopulmonary bypass. METHODS: Fifty-two patients undergoing primary coronary artery bypass grafting by the same surgeon were randomly assigned into either on-pump (n = 26) or off-pump (n = 26) groups. The on-pump coronary artery bypass grafting group underwent mildly hypothermic (35 degrees C) pulsatile cardiopulmonary bypass with arterial line filtration. Arterial blood samples were collected preoperatively, at the end of operation, and at 1, 6, and 24 hours postoperatively. Plasma levels of vasopressin and cortisol were measured using radioimmunoassay. Anesthetic management was standardized. RESULTS: Both groups had similar demographic makeup and extent of revascularization (on-pump coronary artery bypass grafting, 2.8 +/- 1.0 grafts versus off-pump coronary artery bypass grafting, 2.4 +/- 0.9 grafts; p = 0.20). No mortality or major morbidity was observed and there were no crossovers. The cardiopulmonary bypass and aortic cross-clamp times in the on pump coronary artery bypass grafting group were 63 +/- 24 and 33 +/- 11 minutes, respectively. In both groups there was a similar and significant rise in cortisol and vasopressin levels in the early postoperative phase, with a partial recovery toward baseline values observed at 24 hours postoperatively. Repeated measures analysis of covariance showed no significant difference between the groups with time for both hormones (cortisol, p = 0.40; vasopressin, p = 0.30). CONCLUSIONS: Despite the avoidance of cardiopulmonary bypass, off-pump coronary artery bypass grafting surgery triggers a systemic stress hormone response that is comparable to conventional surgical revascularization. The neurohormonal environment during beating-heart surgery should be further explored. PMID- 15276509 TI - Neurocognitive deficit following coronary artery bypass grafting: a prospective study of surgical patients and nonsurgical controls. AB - BACKGROUND: To objectively measure long-term neurocognitive deficit in patients undergoing coronary artery bypass grafting and compare the findings with nonsurgical controls. METHODS: We prospectively measured neurocognitive function in patients undergoing coronary artery bypass grafting (CABG) with cardiopulmonary bypass (n = 104; mean age 64.1 years old; EuroSCORE 2.7 [means]). A cohort of age- and sex-matched patients (n = 80; mean age 63.4 years old) served as nonsurgical controls. After CABG, neurocognitive function was serially reevaluated at 7-day (n = 104), 4-month (n = 100), and 3-year follow-up (n = 88). Neurocognitive function was objectively measured by means of cognitive P300 evoked potentials. Additionally, standard psychometric tests were performed (Trailmaking Test A, Mini Mental State Examination). RESULTS: As compared to preoperative measures (364 +/- 36 ms), cognitive P300 evoked potentials were prolonged (=impaired) at 7-day (381 +/- 36 ms; p = 0.001), 4-month (378 +/- 31 ms; p = 0.08), and 3-year follow-up (379 +/- 35 ms; p = 0.002), respectively. Trailmaking Test A was abnormal, as compared to preoperative, at 3-year follow-up (p < 0.001). Before the operation, surgical patients were fully comparable in P300 measures to nonsurgical controls (363 +/- 32 ms; p = 0.362). Most importantly, throughout the entire postoperative follow-up cognitive measures in surgical patients were prolonged (=impaired) as compared with controls (7-day p = 0.001; 4-month p = 0.002 and 3-year p = 0.003, respectively). In stepwise multivariate regression analysis, neurocognitive deficit at 4-month follow-up (p < 0.001), age (p = 0.012), and persistent atrial fibrillation (p = 0.024) were predictive for long-term neurocognitive deficit at 3-year follow-up. CONCLUSIONS: As shown by means of objective measures, and in comparison to nonsurgical controls, coronary artery bypass grafting with cardiopulmonary bypass grafting causes long-term neurocognitive deficit. PMID- 15276511 TI - Preliminary report on the interaction of apolipoprotein E polymorphism with aortic atherosclerosis and acute nephropathy after CABG. AB - BACKGROUND: Renal dysfunction is a serious complication of cardiac surgery that is highly associated with short- and long-term adverse outcome. While the apolipoprotein E (APOE) epsilon4 allele has been linked to the occurrence of both postcardiac surgery acute renal injury (epsilon4 favorable) and ascending aortic arteriosclerosis (epsilon4 unfavorable), the role of epsilon4 in the relationship between these two conditions is unknown. We hypothesized that patients with and without the epsilon4 allele (E4/non-E4) would have different associations between atheroma burden and postoperative renal dysfunction. METHODS: Ascending, arch, and descending aorta atheromatous burden and APOE status were evaluated for 130 coronary bypass patients. Multivariable analyses were performed for aortic regions to assess the relationship of atheroma burden and APOE epsilon4 status with peak in-hospital postoperative serum creatinine. All p < 0.05 were considered significant. RESULTS: We found an interaction between E4 status (E4/non-E4; 24/106) and atheroma burden, with a much greater predicted peak in hospital postoperative serum creatinine for increases in ascending aorta atheroma load for non-E4 patients versus E4 patients (beta coefficient -0.13; p = 0.002). We also confirmed the association between ascending aorta atheroma and peak creatinine (beta coefficient 0.11; p = 0.0008), after controlling for E4 status, preoperative creatinine, and the E4-atheroma interaction. CONCLUSIONS: Equivalent ascending aortic atheroma burden is associated with a greater susceptibility to postoperative renal injury among patients undergoing cardiac operation who lack the APOE epsilon4 allele. Findings may be attributable to APOE-related differences in inflammation, susceptibility to atheroma detachment (eg, during operative aortic manipulation), or renal vulnerability to embolic injury. PMID- 15276514 TI - Long-term neurologic hand complications after radial artery harvesting using conventional cold and harmonic scalpel techniques. AB - BACKGROUND: The purpose of this study was to determine the incidence of neurologic hand complications after radial artery harvesting and to compare the harmonic scalpel versus conventional cold scalpel technique. METHODS: From 1995 to 2000, 786 radial arteries were harvested from 782 patients for coronary artery bypass grafting. From 1995 to 1997, the conventional cold scalpel technique was used (422 patients), and from 1998 to 2000, the harmonic scalpel was used (360 patients). Mean follow-up was 4.2 +/- 2.1 years and was 90% complete. Symptoms included thumb weakness or numbness, tingling, or pain in the hand. RESULTS: The incidence of neurologic hand complications was similar with both techniques (11.2% +/- 3.5% cold, 11.0% +/- 3.6% harmonic, p > 0.95), and in 19% (13 of 67 with symptoms) there was complete resolution within 1 year. Symptoms persisted long-term in 9.0% +/- 3.2% cold scalpel and 9.0% +/- 3.3% harmonic scalpel patients (p > 0.81), but were considered a "constant and significant source of discomfort" in only 0.6% +/- 0.9% cold scalpel and 1.4% +/- 1.3% harmonic scalpel patients (p > 0.41). CONCLUSIONS: The incidence of adverse neurologic outcomes causing significant long-term discomfort in the hand was low using either the cold scalpel or harmonic scalpel technique. However, a significant number of patients had neurologic hand symptoms in both groups, and this should be included when discussing operative risks with the patient. PMID- 15276512 TI - Reexploration for bleeding after coronary artery bypass surgery: risk factors, outcomes, and the effect of time delay. AB - BACKGROUND: We aimed to identify risk factors for reexploration for bleeding after surgical revascularization in our practice. We also looked at the impact of resternotomy and the effect of time delay on mortality and other in-hospital outcomes. METHODS: In all, 2,898 consecutive patients undergoing coronary artery bypass grafting between April 1999 and March 2002 were retrospectively analyzed from our cardiac surgery registry. Multivariate logistic regression analysis was used to identify risk factors for reexploration for bleeding. To assess the effect of preoperative aspirin and heparin, reexploration patients were propensity matched with unique patients not requiring reexploration. We carried out a casenote review to ascertain the timing and causes for bleeding in patients undergoing resternotomy. RESULTS: Eighty-nine patients (3.1%) underwent reexploration for bleeding. Multivariate analysis revealed smaller body mass index (p = 0.003), nonelective surgery (p = 0.022), 5 or more distal anastomoses (p = 0.035), and increased age (p = 0.041) to have increased risks. Propensity matched analysis showed that preoperative use of aspirin (p = 0.004) and heparin (p = 0.001) were associated with increased risk in the on-pump coronary surgery group only. Patients requiring resternotomy had a significantly greater need for inotropic agents (p < 0.001), and longer intensive care unit stay (p < 0.001) and postoperative stay (p < 0.001) than their propensity-matched controls. However, there was no significant difference in the mortality rate. Adverse outcomes were significantly higher when patients waited more than 12 hours after return to the intensive care unit for resternotomy. CONCLUSIONS: Risk factors for reexploration for bleeding after coronary artery bypass grafting include older age, smaller body mass index, nonelective cases, and 5 or more distal anastomoses. Preoperative aspirin and heparin were risk factors for the on-pump coronary artery surgery group. Patients needing reexploration are at higher risk of complications if the time to reexploration is prolonged. Policies that promote early return to the operating theater for reexploration should be encouraged. PMID- 15276515 TI - Harvest of the radial artery for coronary artery surgery preserves maximal blood flow of the forearm. AB - BACKGROUND: Use of the radial artery as a conduit for coronary artery surgery has increased dramatically. It has been assumed that blood flow to the forearm will not be compromised by its removal. METHODS: Sixteen patients who had the left radial artery harvested for coronary surgery at least 3 months earlier were studied. The right radial artery was not harvested. The radial, ulnar, and brachial artery diameters and flows were measured using pulsed wave Doppler with a 15-MHz linear array transducer. Measurements were performed at rest, with the right radial artery compressed, and after ischemia with forearm exercise. RESULTS: At rest, the (mean +/- SE) diameter of the left ulnar artery was consistently greater than the right (2.4 +/- 0.09 versus 2.1 +/- 0.09 mm, p = 0.001) as was flow (74 +/- 9.9 versus 48 +/- 8.5 mL/min, p = 0.005). There was no difference between diameters or flows in the brachial arteries. After compression of the radial artery, flow increased in the right ulnar artery from 39 +/- 8.0 to 72 +/- 17.6 mL/min (p = 0.019) without an increase in ulnar artery size and was not different from the left ulnar artery flow at rest (p = 0.440). After ischemic forearm exercise, flow increased in the two brachial arteries almost equally (left, 348 +/- 50; right, 371 +/- 63 mL/min). CONCLUSIONS: Blood flow to the forearm and hand is not compromised by harvest of the radial artery. PMID- 15276516 TI - Chronic ultrastructural effects of temporary intraluminal shunts in a porcine off pump model. AB - BACKGROUND: Temporary intraluminal shunts (TILS) are routinely used in off-pump revascularization to facilitate the anastomosis while maintaining myocardial blood supply. Whereas tourniquet-occlusion can cause vessel wall trauma, potentially adverse chronic effects of TILS on the coronary intima have not been evaluated yet. This chronic large animal study investigated ultrastructural effects of TILS on the vessel wall. METHODS: Four groups of acute and chronic pigs with either tourniquet-occlusion (TOUR) or TILS (40 kg; acute, n = 12; chronic, n = 20) were analyzed. Animals underwent median sternotomy, heparin (150 U/kg) administration, and left anterior descending coronary artery exposure. In groups with TOUR the left anterior descending coronary artery was temporarily occluded (10 minutes) with a tourniquet. In groups with TILS a silicone shunt (1.5 mm diameter, 12 mm length) was placed in the left anterior descending coronary artery more than 10 minutes and then removed, and the insertion was repaired. Thirty minutes after reperfusion all acute animals were sacrificed whereas chronic animals were extubated, maintained for 3 months, and then sacrificed. The left anterior descending coronary artery regions of occlusion or placement of the TILS silicone bulbs were examined histopathologically by scanning and transmission electron microscopy by a blinded pathologist. RESULTS: In both acute and chronic investigations animals in the TILS group exhibited significantly less morphologic damage than animals in the TOUR group. In the acute phase significantly more loss of cell junction (p = 0.037), loss of endothelium (p = 0.032), and intimal edema (p = 0.037) in the TOUR group than in the TILS group was observed. Three months later, characteristic features with a changed pattern were detected: vacuolization of the cell (p = 0.03), loss of cell junction (p = 0.042), and removal of basal membrane (p = 0.046) as well as extensive loss of endothelium (p = 0.003) in the TOUR group compared with the TILS group. CONCLUSIONS: Intimal lesions occur with both maneuvers early and late. However, animals in the TOUR group exhibited injuries significantly more often and more severely. Therefore, acute and chronic intimal integrity of the coronary vessel may be better preserved using TILS and may thus have a positive impact on the extent of de novo stenosis and long-term prognosis of the revascularized region. PMID- 15276517 TI - Operative outcome of simultaneous carotid and valvular surgery. AB - BACKGROUND: Operative outcome of simultaneous carotid endarterectomy and valvular surgery has not been clarified. We retrospectively reviewed short-term and long term outcomes after carotid endarterectomy combined with valvular replacement. METHODS: Seventy-nine patients (50 men and 29 women. mean age, 68.9 +/- 6.9 years; range, 53.3 to 78.7 years) underwent carotid endarterectomy combined with valve replacement from February 1985 to April 2002. Indication of carotid endarterectomy was more than 75% carotid stenosis with or without ulceration. Thirteen patients had history of stroke. Endarterectomy was performed under mild hypothermia with cardiopulmonary bypass in all cases. Positions of replaced valves were aortic in 64 patients, mitral in 10, and mitral and aortic in 5 patients. RESULTS: There were 8 early deaths (10.1%). Early neurologic complications occurred in 8 patients (10.1%); two late events were observed. Double valve replacement was an independent risk factor for early death (p = 0.039; odds ratio = 25.6). For early stroke we found no statistically significant risk factor. Myocardial infarction (p = 0.022; odds ratio = 3.0) and age more than 70 years (p = 0.03; odds ratio = 2.5) were independent risk factors for premature death; we found no independent risk factor for late stroke. Permanent impairment or death as a stroke consequence was seen in 5 patients, 3 of them had ipsilateral strokes, 2 had contralateral strokes. CONCLUSIONS: Endarterectomy can be safely performed combined with aortic valve surgery. Concomitant mitral or double valve replacement cannot be judged reliably because of the small number of patients, but they might be a high risk. PMID- 15276519 TI - Six-year monitoring of the donor-specific immune response to cryopreserved aortic allograft valves: implications with valve dysfunction. AB - BACKGROUND: The immune rejection has been anticipated as one of the major causes of allograft aortic valve (AAV) degeneration. The purpose of this study was to prospectively serially measure the magnitude and evolution of the recipient anti HLA class I antibody response up to 6 years from AAV implant and to correlate serologic data with valve performance by means of a concurrent echocardiographic survey. METHODS: Cryopreserved AAVs were obtained from multiorgan HLA-typed donors. Nineteen patients younger than 50 years (mean age, 43.3 +/- 8 years) were prospectively studied. After successful surgery, all AAV recipient underwent at 3 and 6 months and each year postoperatively (mean follow-up, 71.9 months) concomitant serum sample collection and two-dimensional transthoracic echocardiography. The presence of anti-HLA antibodies was tested against a panel of lymphocytes obtained from 30 blood donors. RESULTS: Progressive structural valve deterioration was seen in 6 patients (31.5%) of whom 4 (21%) were reoperated. All pretransplant recipients sera were panel-reactive antibody negative. Seventeen patients (89.4%) demonstrated significant panel-reactive antibody levels, which peaked at 6 months postoperatively, declined from 6 to 24 months, and slowly decreased afterward. In 14 of 19 cases (73.6%) donor-specific HLA antibodies were identified. A strong immunization (6-year persistence of panel-reactive antibody > 70% and peak panel-reactive antibody > 80%) was detected in 31.5% and 36.8% of recipients, respectively. Strong immunization was found to be significantly associated with progressive structural deterioration. CONCLUSIONS: The immune reaction after cryopreserved AAV implantation is a peculiar long-lasting response occurring in the majority of recipients younger than 50 years of age. An association between a sustained and pronounced immunization and an aggressive AAV degeneration was observed. PMID- 15276520 TI - Anterior leaflet augmentation for ischemic mitral regurgitation. AB - BACKGROUND: Mitral valve repair improves survival and quality of life in patients with ischemic mitral regurgitation (MR). Although many repair methods exist for this condition, the ideal approach remains unknown. The purpose of this study is to describe a simple technique for repair of ischemic MR that addresses the pathophysiology of tethered leaflets and to report its early results. METHODS: The technique consists of pericardial patch enlargement of the anterior mitral leaflet and placement of a flexible annuloplasty band. Candidates for the repair had ischemic cardiomyopathy and echocardiographic evidence of moderate or severe Carpentier type IIIb MR. Patients were followed with serial echocardiography. RESULTS: Between January 2002 and November 2003, 25 adult patients underwent anterior leaflet augmentation for ischemic MR. Mean age was 64.8 +/- 10.6 years, and mean left ventricular ejection fraction was 0.36 +/- 0.14. Preoperative MR by transesophageal echocardiography was severe in 84% of patients and moderate in 16%. Annuloplasty band sizes were 27 mm to 31 mm (mean, 28.4 +/- 1.1 mm). Concomitant coronary artery bypass grafting was performed in all patients. Transesophageal echocardiography immediately after repair revealed MR to be none or trace in 80% of patients and mild in 20%. No intraoperative conversion to valve replacement was performed. In follow-up, 2 patients have experienced moderate MR and are being treated medically, and no patients have mitral stenosis. At 2 years, actuarial freedom from moderate or greater MR is 81%. CONCLUSIONS: For patients with ischemic MR, anterior leaflet augmentation is a simple and reproducible method of valve repair that addresses the pathophysiology of tethered leaflets. Early results in a small number of patients have been encouraging. PMID- 15276522 TI - Off-pump epicardial tissue sealing--a novel method for atrioventricular disruption complicating mitral valve procedures. AB - BACKGROUND: Atrioventricular disruption (AVD) is a rare (1%-2%) but fatal complication after mitral valve procedures; the intraoperative mortality is more than 50% despite the current standard procedure of surgical closure of the defect. We compared the outcome of 9 patients with intraoperative AV disruption, 4 being surgically treated on-pump and 5 receiving epicardial tissue sealing off pump. METHODS: Between March 1998 and May 2002 a total of 9 patients presented with AV disruption intraoperative. The first 4 patients were treated with surgical repair on-pump by reconstruction of the defects with patch or buttressed suture. The second series of 5 patients were treated with a biodegradable collagen system with fibrinogen-based coating off-pump. Three to six layers were placed over the bleeding site with manual pressure for 30-60 minutes on the beating heart until bleeding was stopped. Cell saved blood was retransfused. RESULTS: In the on-pump surgical repair group 3 patients (75%) died within the first day after repair either because of persistent bleeding or cardiac tamponade. One patient survived at 30 days and 1 year. In the off-pump tissue sealing group 30 days and 1 year survival was 100%. Postoperative echocardiography showed normal left ventricular (LV) function with no regional wall motion abnormalities. CONCLUSIONS: Our data show that epicardial tissue sealing off-pump results in successful termination of bleeding from AVD and considerably improves survival when compared with the standard procedure. Because of this tremendous improvement in patient survival we now consider this technique as standard therapy for AV disruption in our center. PMID- 15276524 TI - Three-dimensional echocardiography for planning of mitral valve surgery: current applicability? AB - BACKGROUND: Two-dimensional transesophageal echocardiographic (2D TEE) assessment of the mitral valve requires mental integration of a limited number of 2D imaging planes. Structural display in three dimensions from any perspective may be of advantage to the surgeon for better judgment and planning. METHODS: Feasibility, accuracy, and limitations of preoperative three-dimensional transesophageal echocardiography (3D TEE) was assessed in 51 patients with mitral valve disease. The width of the anterior mitral valve was measured with either method and compared with the operative finding. Three-dimensional dynamic sequences of the reconstructed mitral valve were shown preoperatively to the surgeon and later compared with the intraoperative finding. RESULTS: The quality of the 3D reconstruction was graded as good in 25 patients (49.0%), fair in 16 patients (31.4%), and poor in 10 patients (19.6%) where atrial fibrillation did not allow ECG gating. Thirty-nine patients had successful mitral valve repair and twelve patients required valve replacement. Based on intraoperative findings, sensitivity for the diagnosis of mitral valve prolapse using 2D TEE and 3D TEE was 97.7% and 92.9% (p = ns) respectively and specificity was 100% by both methods. Sensitivity for the diagnosis of rupture of chordae tendineae using 2D TEE and 3D TEE was 92.3% and 30.8% respectively (p < 0.05) and specificity was 100% by both methods. CONCLUSIONS: Dynamic 3D echocardiography is feasible and can provide good insight into valvular motion and allows adequate preoperative planning when reconstruction is being considered. However dynamic 3D reconstruction is currently limited by the quality of the original 2D echo cross sectional images which can be adversely affected by minimal patient movements, breathing, or cardiac arrhythmia, thus limiting accuracy of the 3D TEE significantly compared with 2D TEE. PMID- 15276525 TI - Impact of intraoperative transesophageal echocardiography in patients undergoing valve replacement. AB - BACKGROUND: The role of intraoperative transesophageal echocardiography (IOTEE) in valve replacement surgery is not well established. The aim of this study was to explore the impact of immediate postpump IOTEE in valve replacement surgery at a single tertiary medical center. METHODS: The departmental database was screened for valve replacement operations (mechanical or bioprosthetic valves) performed during a 55-month period that were succeeded by immediate postpump IOTEE. Data was gathered regarding the impact of IOTEE on the immediate postoperative course. RESULTS: The study group included 417 patients (44.8% male, 55.2% female, age 65.2 +/- 13.9 years). Prepump IOTEE was performed in 352 patients (84.4%). A single valve was replaced in 336 patients (80.6%) and two or more valves were replaced in 81 patients (19.4%). Overall 501 valves were inserted: mitral, 237 (131 mechanical, 106 biological); aortic, 221 (89 mechanical, 132 biological); tricuspid, 43 (2 mechanical, 41 biological). Unexpected pathologic echocardiographic findings on postpump IOTEE necessitated immediate surgical correction in 15 patients (3.6%): perivalvular leak in 8 patients (4 mitral, 4 aortic), immobilized leaflet in 4 patients (3 mitral, 1 tricuspid), coronary obstruction by an aortic bioprosthesis in 2 patients, and incompetent xenograft in 1 patient. Prolonged removal of air was necessary in 45 patients (10.8%). In 47 patients (11.3%) the postpump IOTEE contributed to the evaluation of difficult weaning from the bypass pump and to its appropriate therapeutic management (volume expansion, inotropic agents, vasodilators, or mechanical assistance). CONCLUSIONS: Immediate postpump IOTEE is an important diagnostic and therapeutic role in valve replacement surgery and should be widely implemented. PMID- 15276527 TI - Surgery for acute type A aortic dissection: is advanced age a contraindication? AB - BACKGROUND: With the general increase in human lifespan, cardiac surgeons are faced with treating an increasing number of elderly patients. The purpose of this study was to demonstrate early and late results of surgery for aortic dissection in patients older than 70 years of age compared with those younger than 70 years and to clarify the clinical problems related to this subset of patients. METHODS: Between 1976 and 2001, 315 patients underwent emergency operation for acute type A dissection: 245 were younger than 70 years (group 1) and 70 patients were 70 years of age and older (group 2). Early and late outcomes of both groups were compared. RESULTS: The hospital mortality rates were 20.5% in group 1 and 17.6% in group 2 (p = 0.751). The mean extracorporeal circulation time was 192.6 +/- 65.2 minutes and 185.7 +/- 58.4 minutes in groups 1 and 2, respectively (p = 0.42). The mean cross-clamp time was 116.3 +/- 45.8 minutes and 100 +/- 36.7 minutes in groups 1 and 2, respectively (p = 0.009). Actuarial survival rates were 77.1% after a mean follow-up time of 259 +/- 9 months for patients of group 1 and 80% after 77 +/- 5 months for patients of group 2, without any statistically significant difference (p = 0.619). CONCLUSIONS: No significant differences were observed in the 30-day mortality and actuarial survival between the two groups. Therefore we believe that surgery for type A acute aortic dissection in patients 70 years of age or older can be performed with acceptable risk of death and satisfactory results. PMID- 15276528 TI - Neurocognitive functions after aortic arch repair with right brachial artery perfusion. AB - BACKGROUND: Satisfactory neurologic outcome following aortic arch repair through right brachial artery perfusion is well established. However, how neurocognitive functions are affected following selective cerebral perfusion, still needs to be elucidated. METHODS: In a period between April 2002 and March 2003, 22 patients (19 male, 3 female, with a mean age of 46.8 +/- 12; range: 26 to 70 years old), underwent aortic arch repair using right brachial artery low flow (8 to 10 mL x kg(-1) x min(-1)) selective antegrade cerebral perfusion under moderate hypothermia (26 degrees C). There were 6 Stanford type-A dissections and 16 ascending aortic aneurysms. All patients were evaluated preoperatively and postoperatively (at seventh day and second month) for neurocognitive functions. RESULTS: There was no operative mortality. The average cardiopulmonary bypass time was 115.0 +/- 24.2 minutes and the average antegrade cerebral perfusion time was 29.8 +/- 7.1 minutes (19 to 38 minutes). No major neurologic deficit was observed in the postoperative period. In terms of neurocognitive test results, between the preoperative and postoperative assessments for both hemispheric cognitive functions no deterioration was detected. CONCLUSIONS: The low-flow selective antegrade cerebral perfusion technique through the right brachial artery may safely be used for the great majority of patients undergoing aortic arch repair without causing deteriorations in neurocognitive functions. PMID- 15276529 TI - Nonneurologic morbidity and profound hypothermia in aortic surgery. AB - BACKGROUND: Use of profoundly hypothermic cardiopulmonary bypass may increase the risk of postoperative bleeding and lung and renal dysfunction. The aim of this study was to analyze postoperative blood loss and indices of pulmonary and renal dysfunction in patients undergoing proximal aortic surgery with and without the use of profound hypothermia to determine risk factors for nonneurologic morbidity. METHODS: Risk factors for blood loss, transfusion requirement, and pulmonary and renal dysfunction were studied in 116 patients undergoing thoracic aortic surgery with profoundly or moderately hypothermic cardiopulmonary bypass. RESULTS: Overall mortality was 8.6%. Mean (+/- standard deviation) cardiopulmonary bypass times were 191 +/- 53 minutes (profoundly hypothermic group) and 131 +/- 48 minutes (moderately hypothermic group; p < 0.0001). The incidence of blood loss more than 1 L or resternotomy for bleeding was 25% (29 patients). Fifteen patients (12.9%) experienced postoperative pulmonary dysfunction, and 25 patients (21.6%) had postoperative renal dysfunction. Forty one patients (35.3%) had a prolonged intensive therapy unit length of stay. Multivariate analysis demonstrated that prolonged cardiopulmonary bypass time was the only predictor of postoperative hemorrhage and resternotomy for bleeding (p = 0.03). Increased intensive therapy unit length of stay was predicted by total arch replacement (p = 0.01) and low 6-hour ratio of partial pressure of arterial oxygen to inspired fraction of oxygen (p = 0.05). Increased preoperative creatinine (p = 0.002) and emergency status (p = 0.015) predicted postoperative renal dysfunction. Low 6-hour ratio of partial pressure of arterial oxygen to inspired fraction of oxygen was predicted by increased preoperative creatinine (p = 0.03) and prolonged cardiopulmonary bypass time (p = 0.03). CONCLUSIONS: Profound hypothermia may cause a coagulopathy, but procedure extent is the primary determinant of postoperative bleeding. Profoundly hypothermic cardiopulmonary bypass does not appear to be a risk factor for renal or early pulmonary dysfunction or intensive therapy unit length of stay. PMID- 15276530 TI - MCI-186 reduces oxidative cellular damage and increases DNA repair function in the rabbit spinal cord after transient ischemia. AB - BACKGROUND: Paraplegia is a serious complication of operations on the thoracic and thoracoabdominal aorta. To investigate the mechanism by which motor neurons are damaged during these operations, we have reported a rabbit model of spinal cord ischemia. We also tested whether a free radical scavenger MCI-186 that is useful for treating ischemic damage in the brain can protect against ischemic spinal cord damage. METHODS: Fifteen minutes of ischemia was induced, then MCI 186 or vehicle was injected intravenously. Cell damage was analyzed by observing the function of the lower limbs and by counting the number of motor neurons. To investigate the mechanism by which MCI-186 prevents ischemic spinal cord damage, we observed the immunoreactivity of 8-hydroxy-2'-deoxyguanosine as an oxidative DNA damage marker and redox effector as a DNA repair marker. RESULTS: In sham control, 8-hydroxy-2'-deoxyguanosine was not observed, and the nuclear expression of redox effector was observed. In vehicle injection group (group I), the nuclear expression of 8-hydroxy-2'-deoxyguanosine was observed at 1 and 2 days after reperfusion. The nuclear expression of redox effector was observed at 8 hours and 1 day, and disappeared at 2 days after transient ischemia. In MCI-186 injection group (group M), the nuclear expression of 8-hydroxy-2'-deoxyguanosine was not observed, and redox effector was observed at 8 hours and 1 and 2 days. CONCLUSIONS: These results suggest that redox effector decreased in motor neurons after transient ischemia and this reduction preceded oxidative DNA damage. MCI 186 works as a radical scavenger and reduced oxidative DNA damage, so redox effector did not disappear. MCI-186 could be a strong candidate for a use as a therapeutic agent in the treatment of ischemic spinal cord injury. PMID- 15276531 TI - Simplified treatment of postoperative mediastinitis. AB - BACKGROUND: Wound infection after median sternotomy for cardiac or thoracic surgery is a serious complication. A variety of treatment plans have been advocated, and there is lack of agreement regarding the best treatment method. We present our results in patients with mediastinitis who have been treated in a simple, consistent manner. METHODS: We reviewed our experience with 40 consecutive patients with mediastinitis who were treated between January 1995 and May 2003 with a single-stage treatment consisting of sternal and soft tissue debridement and wound closure over mediastinal tubes with continuous irrigation and drainage. Tubes were placed posterior to the sternum in all patients and were irrigated continuously for at least 7 days with antibiotic or antibacterial solution. Systemic antibiotics were selected based on culture and sensitivity data and were administered for 2 to 6 weeks. RESULTS: All patients with mediastinitis treated in this manner survived. Of the 40 patients, 38 achieved complete healing of the wound without further operative intervention or major complication. One patient had recurrent infection and required sternal resection and advancement of muscle flaps. One patient had a residual localized focus of chondritis and underwent limited resection of cartilage. CONCLUSIONS: In this series of patients with postoperative mediastinitis, a simplified approach consisting of wound debridement, reclosure over drains, and anterior mediastinal irrigation has been an effective treatment. The results we have achieved suggest that this technique may be a suitable option for treating this condition. PMID- 15276532 TI - Substernal epicardial echocardiography: a recommended examination sequence and clinical evaluation in patients undergoing cardiac surgery. AB - BACKGROUND: Substernal epicardial echocardiography is a novel echocardiography window, utilizing a modified mediastinal drain incorporating a sleeve for the insertion of a transesophageal echocardiography probe. METHODS: Forty-six patients undergoing cardiac surgery from two institutions were evaluated, and an examination sequence was developed. RESULTS: An 11-view examination is presented as a consensus between the two institutions. In clinical usage, there were no major complications attributable to use of the device. Minor air leaks occurred in 6 patients, and 2 cases of sternal wound infection occurring in a cluster of infections are reported, but causation was not attributed to use of the device. There were no significant differences in measurements of the aortic valve area, pulmonary artery diameter, left ventricular outflow tract dimension, or the sinotubular junction between substernal and transesophageal examinations. All 16 wall-motion segments were well visualized in most patients with substernal epicardial echocardiography. CONCLUSIONS: Substernal epicardial echocardiography is a safe device for use in the postoperative environment. PMID- 15276534 TI - Donor heart preservation with pinacidil: the role of the mitochondrial K ATP channel. AB - BACKGROUND: Pinacidil solutions have been shown to have significant cardioprotective effects. Pinacidil activates both sarcolemmal and mitochondrial potassium-adenosine triphosphate (K(ATP)) channels. This study was undertaken to compare pinacidil solution with University of Wisconsin (UW) solution and to determine if the protective effect of pinacidil involved mitochondrial or sarcolemmal K(ATP) channels. METHODS: Thirty-two rabbit hearts received one of four preservation solutions in a Langendorff apparatus: (1) UW; (2) a solution containing 0.5 mmol/L pinacidil; (3) pinacidil with Hoechst-Marion-Roussel 1098 (HMR-1098), a sarcolemmal channel blocker; and (4) pinacidil with 5 hydroxydecanote, a mitochondrial channel blocker. Left ventricular pressure volume curves were generated by an intraventricular balloon. All hearts were placed in cold storage for 8 hours, followed by 60 minutes of reperfusion. RESULTS: Postischemic developed pressure was better preserved by pinacidil than by UW. This cardioprotective effect was eliminated by 5-hydroxydecanote and diminished by HMR-1098. Diastolic compliance was better preserved by pinacidil when compared with UW. This protection was abolished by the addition of 5 hydroxydecanote and moderately decreased by HMR-1098. CONCLUSIONS: Our results support the superiority of pinacidil over UW after 8 hours of storage. The cardioprotective role of pinacidil is mediated primarily by the mitochondrial K(ATP) channel. PMID- 15276535 TI - Brief pressure overload of the left ventricle preconditions rabbit myocardium against infarction. AB - BACKGROUND: Several nonischemic stimuli have been shown to precondition myocardium. We investigated cardioprotective effects and underlying mechanisms of brief pressure overload of the left ventricle in this study. METHODS: Brief pressure overload of the left ventricle was achieved by two 10-minute partial snaring of the ascending aorta so that systolic left ventricular pressure was raised 50% above the baseline value. Ischemic preconditioning was elicited by two 10-minute coronary artery occlusions. Ten minutes after different pretreatments, myocardial infarction was induced by a 60-minute coronary artery occlusion followed by 3-hour reperfusion. Area at risk and myocardial infarct was determined by blue dye injection and triphenyl tetrazolium chloride staining. RESULTS: The myocardial infarct size, expressed as percentage of area at risk, was significantly reduced in the pressure overload group (15.9% +/- 2.9%, p < 0.001, n = 9) as well as in the ischemic preconditioning group (14.9% +/- 1.9%, p < 0.001, n = 9) versus the control group (30.0% +/- 6.9%, n = 10). Pretreatment with a blocker of stretch-activated ion channels (gadolinium, 40 micromol/kg, intravenous) abolished the protection induced by pressure overload and ischemic preconditioning. Gadolinium itself did not alter the extent of infarct. There was no significant difference in hemodynamics, area at risk, and mortality among all groups of animals. CONCLUSIONS: Brief pressure overload of the left ventricle by partial snaring of the ascending aorta preconditioned rabbit myocardium against infarction. The underlying mechanism might be related to activation of stretch activated ion channels. PMID- 15276536 TI - Effect of leukocyte depletion on endothelial cell activation and transendothelial migration of leukocytes during cardiopulmonary bypass. AB - BACKGROUND: Although leukocyte depletion from systemic circulation during cardiopulmonary bypass (CPB) has been studied, the effect of leukocyte depletion on the leukocyte-endothelial cascade remains poorly understood. So far, there has been no published work on the effects of leukocyte filters during cardiac operations from the viewpoint of endothelial activation and transendothelial neutrophil migration. METHODS: Thirty-two patients undergoing elective heart operations were randomly allocated to a leukocyte-depletion (LD) group or a control group. Blood samples were collected at seven time points: before sternotomy, at 30 minutes and at 60 minutes of CPB, at 5 minutes after coronary reperfusion, at the end of CPB, and at 2 hours and 24 hours after the cessation of CPB. The plasma concentrations of P-selectin, intercellular adhesion molecule 1 (ICAM-1), interleukin-8, and platelet-endothelial cell adhesion molecule-1 (PECAM-1) were measured using enzyme-linked immunosorbent assays. Plasma malondialdehyde (MDA) concentration was determined by measurement of thiobarbituric acid-reactive substances in plasma. In addition, blood samples collected at intervals before and after operation were used for arterial blood gases. RESULTS: Our studies show significant increases of plasma levels of P selectin, ICAM-1, interleukin-8, PECAM-1, and MDA during and after CPB in the control group. Interestingly, a significant decrease of plasma levels of P selectin, ICAM-1, interleukin-8, PECAM-1, and MDA, and better preservation of lung function could be found in the LD group compared with the control group. CONCLUSIONS: Our results demonstrate a rationale for using a leukocyte filter in patients undergoing cardiac surgery to attenuate the endothelial-mediated component of the CPB-induced inflammatory response by reducing endothelial activation and neutrophil transmigration. PMID- 15276538 TI - Cardiac retransplantation in children. AB - BACKGROUND: Experience with pediatric cardiac retransplantation is limited. Outcomes should be inspected to insure proper use of donor hearts. METHODS: Of 152 pediatric heart transplantations, we performed 20 retransplants in 17 children (3 had a second retransplant). The retransplant children were older than the primary transplant children (11.1 +/- 4.4 years versus 7.1 +/- 6.0 years; p = 0.005). Excluding 1 early retransplant, the interval from primary transplant to retransplant was 5.5 +/- 3.3 years (range, 1.1 to 11.1). The retransplant patients were clinically more ill than the primary transplant patients (United Network for Organ Sharing status I, 75% versus 63%; mechanical circulatory support or dialysis, 20% versus 3.8%). RESULTS: Donor ischemia time (188 versus 165 minutes) and cardiopulmonary bypass time (127 versus 127 minutes) were not significantly different for the retransplant patients. Excluding 1 retransplant patient who required a tracheostomy, days on the ventilator (2.7 versus 2.7), days on inotropic support (3.0 versus 3.2), intensive care unit days (7.2 versus 6.7), and hospital days (15.9 versus 13.8) were similar in the retransplant group. Freedom from rejection at 90 days and 1 year was not different in the retransplant patients. Actuarial patient survival in the patients undergoing first retransplant was similar to the primary transplant patients at 30 days (95% versus 94.7%), 1 year (94.1% versus 80.7%), and 3 years (78.4% versus 73.1%). Two of 3 children receiving a third transplant died within 1 year of redo retransplantation. CONCLUSIONS: Cardiac retransplantation can be performed in children with results comparable with those for primary transplantation despite increased clinical acuity. These early results suggest that cardiac retransplantation in children is a reasonable therapeutic option. Children with repeat retransplantation do not fare as well. PMID- 15276539 TI - Mid-term results for double inlet left ventricle and similar morphologies: timing of Damus-Kaye-Stansel. AB - BACKGROUND: Patients with double inlet left ventricle/l-transposition and similar morphologies have their systemic outflow traverse a bulboventricular foramen (BVF), which has a propensity to narrow over time. A Norwood procedure may be performed as the initial palliation. We prefer aortic arch repair and pulmonary artery banding, delaying Damus-Kaye-Stansel (DKS) or BVF resection until the second palliation. The aims of this study were to compare our results with those reported for Norwood strategy and examine the development of systemic outflow obstruction. METHODS: Retrospective study of patients with double inlet left ventricle, L-TGA or similar morphology presenting between 1990 and 2000. Follow up with clinical assessment, echocardiography and catheter studies. RESULTS: Twenty-five patients had initial palliation with pulmonary artery banding with repair of any associated arch obstruction. Twelve patients had DKS performed as part of their second stage procedure, and 3 had DKS performed later for recurrent stenosis after prior enlargement of BVF. Six patients had BVF resection without later restenosis and 4 patients did not develop BVF stenosis. There was one early death (4%) and two late (8%). Fontan completion was achieved in 20 of the 22 survivors. There were no cases of DKS obstruction, no pulmonary valve had more than mild regurgitation. CONCLUSIONS: Our approach achieves low operative mortality and morbidity and compares favorably with reported results for Norwood palliation. The significant rate of systemic outflow obstruction in those who did not undergo DKS at the second stage confirms the utility of early DKS in children with this morphology. PMID- 15276541 TI - Postoperative assessment of the univentricular repair by dynamic radionuclide studies. AB - BACKGROUND: The purpose of this investigation was to determine the role of radionuclide studies in evaluating postoperative Fontan hemodynamics and to quantify its diagnostic accuracy. METHODS: One hundred five patients (105), aged 11 months to 35 years old, who had undergone univentricular repair, underwent first-pass and multigated acquisition scan 1 month to 10 years after univentricular repair. Forty-five patients with evidence of Fontan failure underwent radionuclide studies using Technetium-99 m as well as cardiac catheterization (group 1). The remaining sixty randomly selected patients with excellent functional status received radionuclide studies alone (group 2). The receiver operating characteristic curve analysis was done to quantify the diagnostic accuracy of the first-pass study. RESULTS: There was paradoxical filling of the right lung after femoral injection in all cases of tunnel or conduit obstruction. A first-pass transit time of 16 to 25 seconds (mean +/- standard deviation [SD] = 18.82 +/- 2.69) was always associated with Fontan failure and high right atrial pressure (range = 20 to 24 mm Hg, mean +/- SD = 22.02 +/- 1.58). A first-pass transit time of 16 seconds was associated with a sensitivity of 100% and a specificity of 93.33%. The predictive accuracy of a positive or negative result was 91.8% and 100% respectively. The area measured under the receiver operating characteristic curve indicates that 99.41% (SE +/- 0.0035) of the time, the value of first-pass time is higher for the Fontan failure group (group 1) compared to the normal group (group 2; p = 0.000). CONCLUSIONS: Our data indicate that Fontan circuit can be reliably evaluated for both anatomic and functional flaws by radionuclide studies; radionuclide first pass time may be used to predict the chances of Fontan failure postoperatively as well as its presence; and in the presence of atrial fibrillation with fast ventricular rate, analysis using first-pass radionuclide may be impossible and gated equilibrium radionuclide angiocardiography may be the preferred method. Inspection of the systemic ventricular time-activity curve is of crucial importance in this regard. PMID- 15276542 TI - Better surgical prognosis for patients with complete atrioventricular septal defect and Down's syndrome. AB - BACKGROUND: Several studies have shown that Down's syndrome is not a risk factor for biventricular repair of complete atrioventricular septal defects. However, few data are available about the comprehensive outcome of all the cardiac surgical procedures in patients with trisomy 21, including palliative surgery. METHODS: This is a retrospective study of 206 consecutive patients who underwent cardiac surgery from January 1992 to January 2002. Data about mortality and morbidity were analyzed and the impact of Down's syndrome was evaluated. RESULTS: Overall mortality was 7.7%. Actuarial survival was 94% among patients with Down's syndrome versus 86% of the group with normal karyotype (p = 0.12). The presence of unbalanced ventricles was the only independent risk factor affecting survival at multivariate analysis (p < 0.0001). The need for a Norwood type surgery was more frequent among non-Down patients (12.0% vs 1.5%, p = 0.02) as was the prevalence of pulmonary artery banding operations (22.9% vs 9.3%, p = 0.04). Cumulative mortality after palliation was higher in non-Down patients (44% vs 2.9%, p = 0.0001). Freedom from reoperation was lower in the group with normal chromosomes in respect to patients with Down's syndrome (81.4% vs 94.6%, p = 0.04), due to the higher prevalence of anomalies of the mitral valve (4.9% vs 1.8%, p = 0.03) or left ventricular outflow tract (7.3% vs 0%, p = 0.01). CONCLUSIONS: Down patients showed a decreased risk for biventricular repair and lower mortality and morbidity in cases of complex cardiac malformations requiring complex palliative operations. PMID- 15276544 TI - Novel treatment strategy for leg and sternal wound complications after coronary artery bypass graft surgery: bioengineered Apligraf. AB - PURPOSE: To demonstrate that bioengineered Apligraf improves time to wound healing in sternal and leg wound complications after coronary artery bypass surgery. DESCRIPTION: Between 1998 and 2001, 1,550 patients underwent coronary artery bypass surgery utilizing saphenous vein. In 45 (2.9%) of 1,550 patients, leg wound complications developed (group A); and in 15 (0.9%) of 1,550 patients, sternal wound complications developed (group B). Apligraf was utilized as the primary treatment for 30 (66%) of 45 leg wounds and for 9 (60%) of 15 sternal wounds. Traditional wound care included debridement and daily wet-to-dry dressings. EVALUATION: Time to wound healing ranged from 26 to 72 days (mean, 46) for Apligraf group A and from 34 to 120 days (mean, 84) for traditional wound care group A. The time to wound healing ranged from 21 to 80 days (mean, 39) for Apligraf group B, and from 36 to 110 days (mean, 62) for traditional care group B. Apligraf treatment was simpler, with less time and resource utilization than traditional wound care. CONCLUSIONS: Apligraf significantly improves time to wound healing in patients with leg and sternal wound complications and offers an attractive new treatment alternative to traditional wound care. PMID- 15276546 TI - Ninety-degree anterior cardiac displacement in off-pump coronary artery bypass grafting: the Starfish cardiac positioner preserves stroke volume and arterial pressure. AB - PURPOSE: In off-pump coronary surgery through sternotomy, exposure of posterior circumflex branches causes circulatory deterioration in both patients and pigs. We assessed cardiac pump function when displacing the pig heart anteriorly with a suction cardiac positioner. DESCRIPTION: Six pigs (+/-80 kg) underwent sternotomy for hemodynamic instrumentation using catheter-tipped manometers and paced at 80 beats/min. Ultrasound flow probes were placed around the aorta and proximal coronary arteries. The heart was retracted anteriorly to 90 degrees with the Starfish cardiac positioner attached to the apex by means of suction (-400 mm Hg). Retraction was guided by cardiac output monitoring. EVALUATION: Anterior displacement to 90 degrees facilitated full exposure of posterior arteries. Stroke volume and mean arterial pressure decreased to 94% +/- 13% (mean +/- SD, p = 0.135) and 95% +/- 13% (p = 0.09) of control values, respectively. Right and left ventricular end-diastolic pressure increased to 129% +/- 37% (p = 0.009) and to 128% +/- 57% (p = 0.235), respectively. Coronary flow remained unchanged. Additional 15-degree head-down positioning increased stroke volume to 113% +/- 17% (p = 0.015) and mean arterial pressure to 113% +/- 25% (p = 0.087) at the expense of further increased right and left ventricular end-diastolic pressure (186% +/- 63%, p < 0.001 and 157% +/- 49%, p < 0.001, respectively). CONCLUSIONS: When lifting the porcine heart ninety degrees anteriorly, the Starfish cardiac positioner facilitated exposure of posterior branches and, when guided by cardiac output, preserved stroke volume and arterial pressure. PMID- 15276548 TI - Surgical revision of an uncommonly dislocated self-expanding Amplatzer septal occluder device. AB - Open heart surgery is the standard procedure for closure of ostium secundum atrial septal defects. Recently, percutaneous transcatheter procedures emerged as therapeutic alternatives for closure of both atrial septal defects and patent foramen ovale. Unfortunately, however, such percutaneous procedures may require surgical intervention for early or late complications. We report a case with emergent surgery for dislocation of the Amplatzer septal occluder into the aortic arch diagnosed 30 days after percutaneous closure of an atrial septal defect. PMID- 15276551 TI - Long-term left internal mammary artery graft patency for coronary artery disease associated with pseudoxanthoma elasticum. AB - Pseudoxanthoma elasticum (PXE) is a rare, inherited connective tissue disorder with numerous systemic manifestations that include premature coronary artery disease. Coronary artery bypass grafting (CABG) is known to be beneficial in patients with PXE-related coronary artery disease. In these patients, however, the suitability of arterial conduits, including the internal mammary artery, has been controversial. We present a patient with PXE-related coronary artery disease who has had long-term patency of a left internal mammary artery (LIMA) graft after an off-pump CABG procedure in which LIMA and bilateral radial artery conduits were used. PMID- 15276550 TI - Late bleeding from right internal mammary artery after HeartMate left ventricular assist device implantation. AB - Postoperative bleeding is one of the major complications after implantation of left ventricular assist devices. We experienced 5 unusual cases, which had bleeding from the right internal mammary artery between 5 and 69 days after implantation of a HeartMate (Thoratec Corporation, Pleasanton, CA) device. It was evident that the outflow graft had eroded through the vessel. Sudden decreases in device flow, hypotension, bleeding from the driveline or chest tube sites, and a drop in hematocrit were the initial manifestations. Chest roentgenogram and transthoracic echocardiography were effective in identifying hemothorax and cardiac tamponade. Four out of 5 patients survived to heart transplantation and were discharged from the hospital. When identified and treated appropriately, this complication does not impair patient outcome. PMID- 15276549 TI - Pulmonary homograft endocarditis after Ross procedure. AB - We report the case of a 36-year-old patient who experienced an isolated acute pulmonary homograft endocarditis two years after a Ross procedure for aortic valve infective endocarditis. PMID- 15276552 TI - Rupture of a giant coronary artery aneurysm due to Kawasaki disease. AB - Coronary artery aneurysm requiring surgery is rare. We report a case of a ruptured giant coronary artery aneurysm due to Kawasaki vasculitis which presented with cardiac arrest and was successfully treated by emergency coronary artery bypass grafting. The controversies surrounding the management of this disease are also discussed. PMID- 15276553 TI - Jehovah's Witnesses requiring complex urgent cardiothoracic surgery. AB - Blood-sparing surgical technique and perioperative medical management allows complex surgery in Jehovah's Witnesses. The authors review four cases of urgent cardiothoracic surgery performed on Jehovah's Witnesses. No lasting sequelae associated with either bleeding or end-organ ischemia were noted. Cardiopulmonary bypass management, meticulous hemostatic operative technique, and pharmacologic support of hemostasis and erythropoiesis permit this complex surgery. PMID- 15276554 TI - Regression of severe pulmonary arteriovenous malformations after Fontan revision and "hepatic factor" rerouting. AB - Although previously described in patients undergoing staged palliation for univentricular heart disease, the mechanism by which hepatic venous flow prevents development of pulmonary arteriovenous malformations is still not completely understood. We present a case in which successful H-type Fontan revision with rerouting of hepatic venous flow through a hemiazygous vein successfully reversed the progression of severe left pulmonary arteriovenous malformations. PMID- 15276555 TI - Recurrent acute rheumatic fever: a forgotten diagnosis? AB - The incidence of acute rheumatic fever has seen a dramatic decline over the last 15 to 20 years in most developed countries and treatment of this disease has changed little since. The ease of travel and immigration and the cosmopolitan nature of many cities mean that occasionally the disease will come to the attention of clinicians not familiar with its presentation, resulting in delayed diagnosis and treatment. We present a case of recurrent acute rheumatic fever in a patient who was initially thought to be suffering from acute bacterial endocarditis on her previously diseased rheumatic aortic valve. This culminated in her undergoing urgent aortic valve replacement during a phase of the illness that should have been treated with high dose anti-inflammatory medication. Therefore, clinicians should be aware of this condition and include it in their differential diagnosis of the febrile patient with a previous history of rheumatic fever. We briefly discuss the diagnostic dilemma of patients suffering from this condition and in differentiating it from acute endocarditis. PMID- 15276557 TI - Autologous salvaged blood transfusion in spontaneous hemopneumothorax. AB - Spontaneous hemopneumothorax (SHP) is a rare clinical entity, and an emergent operation due to continuous bleeding or hypovolemic shock is at times necessary. Although allogeneic blood transfusions are urgently required for significant blood loss, autologous blood transfusions can also be considered in patients with SHP. We herein report two cases of successful autologous blood transfusions using blood in the pleural space, decreasing or obviating the need for allogeneic blood transfusion. PMID- 15276556 TI - Successful treatment of esophageal cancer with transhiatal esophagectomy after heart transplantation. AB - A 55-year-old heart transplant recipient with reflux esophagitis presented for routine endoscopic surveillance of an area of Barrett's metaplasia initially seen 3 years previously. Esophagogastroduodenoscopy revealed adenocarcinoma at 33 cm from the incisors. The preoperative clinical stage was T1N0M0 by endoscopic ultrasound. Transhiatal esophagectomy was performed with R0 resection of the cancer, and the patient recovered uneventfully. Pathologic examination confirmed esophageal adenocarcinoma (T1N0M0) in Barrett's mucosa. The patient is doing well, and has no evidence of disease after 18 months. PMID- 15276558 TI - Pneumothorax after left pneumonectomy: implantation of an intrapleural prosthesis. AB - Postpneumonectomy syndrome is defined as an airway obstruction due to mediastinal shift and rotation after pneumonectomy. A patient who had undergone a left pneumonectomy for bronchial carcinoma 13 years before presented with tension pneumothorax of her remaining lung. Although all factors relevant to the development of postpneumonectomy syndrome were ascertained, the patient had a pneumothorax rather than an airway obstruction. This pneumothorax was treated surgically. The goal of this operation was to reduce the right pleural cavity volume by implanting an intrapleural prosthesis in the pneumonectomy cavity. This treatment is identical to that used for postpneumonectomy syndrome, which allows the right lung to be rejoined with the thoracic wall. PMID- 15276559 TI - Lung cancer and skeletal muscle metastases. AB - Skeletal muscle metastases from lung cancer are rare, and the optimal treatment strategy is unknown. Three cases of skeletal muscle metastases from lung cancer are described. In 2 patients surgical biopsy of muscle swelling disclosed the presence of the lung tumor; the first patient underwent lung resection to remove the primary lesion, the second was not operable because of the metastatic extension of the disease. In the third patient muscle metastasis was observed and excised after lung resection. Adenocarcinoma, squamous cell, and small cell carcinoma were the histologic types diagnosed. Various regimens of radiotherapy and chemotherapy were adopted. Survival times were 3, 6, and 30 months. PMID- 15276560 TI - Spontaneous pneumomediastinum. AB - Pneumomediastinum is the presence of air in the mediastinum. Spontaneous pneumomediastinum (SPM) is an infrequent, benign, and self-limiting condition that predominantly affects young males and pregnant females. It is important to distinguish pneumomediastinum symptoms from similar clinical findings that require immediate treatment, such as cardiac tamponade, angina pectoris, dissecting aortic aneurysm, mediastinitis, and pulmonary embolism. This report describes 2 cases of SPM managed at University Hospital Hamburg-Eppendorf during the period 2000 to 2001. Spontaneous pneumomediastinum should be considered whenever there are anamnestic data for retrosternal chest pain that radiates to the neck or back accompanied by dysphagia, dysphonia, dyspnea, and a positive Hamman's sign. PMID- 15276561 TI - Peripheral tumors of the intercostal nerves. AB - Fewer than 10% of primary neural tumors of the chest originate peripherally from intercostal nerves; most neural tumors of the chest arise in the mediastinum. Most patients with primary tumors of the intercostal nerve are asymptomatic. We report a case of neurilemmoma arising from an intercostal nerve in a woman seen for severe pain in the chest wall. Resecting the tumor relieved the pain. Recent medical literature describing peripheral tumors of thoracic nerves is reviewed. PMID- 15276562 TI - Extraskeletal Ewing sarcoma of the diaphragm presenting with hemothorax. AB - Ewing sarcoma is a relatively uncommon malignant bone neoplasm that usually occurs in children and young adults and involves the major long bones, pelvis, and ribs. Primary diaphragmatic Ewing sarcoma is extremely rare. To the best of our knowledge, only three cases of primary Ewing sarcoma of the diaphragm have been reported. A 12-year-old girl presented spontaneous occurrences of the right hemothorax. After drainage, a roentgenogram film, computed tomography, ultrasonography, and magnetic resonance image showed a giant mass on the right diaphragm. Primary diaphragmatic tumor was resected totally by right posterolateral thoracotomy, and histologically, an extraskeletal Ewing sarcoma was identified. The patient received adjuvant radio-chemotherapy, and there was no evidence of disease 10 months after the operation. Although extremely rare, extraskeletal Ewing sarcoma should be kept in mind in the differential diagnosis of diaphragmatic soft tissue tumors. PMID- 15276563 TI - Giant neurofibroma of the chest wall. PMID- 15276564 TI - Three-dimensional demonstration of the artery of Adamkiewicz by multidetector-row computed tomography. PMID- 15276565 TI - An alternative method of neck flexion after tracheal resection. AB - Tracheal resections for benign and malignant disease are well described. The addition of release procedures, including suprathyroid and suprahyoid laryngeal release, has increased the capability of extended tracheal resection and primary reconstruction. Constant neck flexion by a suture between the skin of the point of the chin and midline of the chest over the manubrium is also widely considered paramount to successful tracheal resections. We designed a straightforward alternative to this method for patient comfort and compliance. PMID- 15276566 TI - Interrupted distal anastomosis: the interrupted "porcupine" technique. AB - The distal coronary artery bypass graft anastomosis created by an interrupted technique using nitinol clips is likely superior to that created with continuous suture because surgeons place clips with optimal visualization, and the anastomosis exhibits optimal compliance and cannot become a "purse-string" once constructed. Skillful use of the clips allows the surgeon to work in the ever more cramped quarters of the minithoracotomy or minimally invasive incision. Anastomosing vessels without knot tying is a valuable practice in the application of remote and robotic surgeries. PMID- 15276567 TI - Exposure of the proximal descending thoracic aorta in the first stage of the elephant trunk procedure. AB - My colleagues and I propose a simple and reproducible technique to achieve optimal exposure and mobilization of the distal aortic arch and proximal descending thoracic aorta in the first stage of the elephant trunk procedure. The technique uses division of the ligamentum arteriosum and a series of circumferential pledgeted traction stitches on the segment of aorta selected for the distal anastomosis. PMID- 15276568 TI - Total extracardiac cavopulmonary connection: an alternative technique of fenestration. AB - Fenestrating an extracardiac conduit used for total cavopulmonary connection normally requires an additional incision on the right-sided atrium and is time consuming. Herein we describe an alternative technique that may be used to facilitate this process, which consists of creating the fenestration by using part of the atrial incision resulting from the disconnection of the inferior vena cava from the right atrium. The advantages of this technique are avoidance of an extra incision and suture line on the atrium, and the ease of construction. This may be especially useful in patients with heterotaxy syndromes with mesocardia or dextrocardia, in whom the atrial mass is displaced posteriorly and can be difficult to reach. Closure of the fenestration can be easily performed at a later stage in the cardiac catheterization laboratory by using a septal occluding device. PMID- 15276569 TI - Role of statin therapy in the coronary bypass patient. AB - Statins have been proven to prevent or delay ischemic events in patients at risk for atherosclerotic coronary disease. Increasing evidence suggests that statin therapy is also beneficial to patients undergoing coronary revascularization. In this review statin therapy will be shown to improve vein graft patency, minimize recurrent ischemic events, and decrease the need for repeat revascularization procedures in patients who have undergone coronary artery bypass grafting. PMID- 15276570 TI - Alejandro Posadas, Argentinian pioneer: thoracic surgery in the Western world in his time. PMID- 15276571 TI - Drawbacks to videothoracoscopic management of solitary pulmonary nodules. PMID- 15276574 TI - Is surgery for T4 non-small-cell lung cancer worthwhile? PMID- 15276575 TI - Outcomes in nonagenerians after open heart operation. PMID- 15276576 TI - Risk factors for stroke after cardiac operations. PMID- 15276579 TI - Is stenting an option? PMID- 15276580 TI - Surgical approach to ascending aorta in patients with bicuspid aortic valve. PMID- 15276581 TI - Retained cuff of retrograde cardioplegia cannula in the coronary sinus. PMID- 15276582 TI - Airway obstruction complicating esophageal stent placement in two post pneumonectomy patients. AB - Expandable metallic stents have been used effectively to treat multiple nonsurgical esophageal conditions. Here we describe two cases in postpneumonectomy patients in which expandable esophageal stent placement resulted in respiratory compromise requiring reintervention due to airway compression. PMID- 15276583 TI - First robotic endoscopic epicardial isolation of the pulmonary veins with microwave energy in a patient in chronic atrial fibrillation. AB - The pulmonary veins have been demonstrated to play an important role in generating atrial fibrillation. We report the first successful endoscopic epicardial isolation of the pulmonary veins in a patient with permanent atrial fibrillation, along with a 1-year follow-up. The procedure consisted of making a conduction block around the pulmonary veins with a flexible microwave energy delivery probe. The probe was placed endoscopically on the left atrial epicardium with the aid of robotic instruments. PMID- 15276584 TI - Coronary artery to the left atrial fistula after resection of atrial appendages. AB - We report the case of a fistula formation between the left circumflex coronary artery draining into the left atrium as a complication of radiofrequency cardio ablation and resection of the atrial appendages. This complication was diagnosed with the use of transesophageal echocardiography and was subsequently confirmed on coronary angiography. PMID- 15276585 TI - Antegrade endovascular repair of a coarctation-associated aneurysm through an upper hemi-sternotomy. AB - Late aneurysm formation is a well-described complication after surgical correction of aortic coarctation. Endovascular repair of such aneurysms avoids the morbidity of conventional reoperative thoracic surgery. We describe a unique case of antegrade endovascular repair of a distal coarctation-associated aneurysm with vascular access acquired through the aortic arch by an upper hemi sternotomy. PMID- 15276586 TI - Open-heart surgery in pediatric patients with end-stage liver disease. AB - Little is known about the safety of pediatric cardiac surgery in children with end-stage liver disease. We reviewed our experience with 4 patients with biliary atresia or Alagille's syndrome who underwent repair of ventricular septal defect and tricuspid regurgitation, atrioventricular canal, subaortic stenosis, or supravalvular aortic stenosis. One patient died on postoperative day 2. All other patients survived to discharge. At follow-up, 1 patient died at home awaiting liver transplantation and the remaining patients are doing well. One patient received a successful liver transplant. Pediatric cardiac surgery in children with end-stage liver disease can be done safely, albeit with a higher mortality. PMID- 15276587 TI - Mediastinal cystic teratoma. PMID- 15276588 TI - Spontaneous coronary artery dissection complicating midterm pregnancy. PMID- 15276589 TI - Triangular plication of the anterior mitral leaflet: a new operative technique. AB - This study describes the technique of triangular plication in patients with mitral valve incompetence that is due to segmental anterior leaflet prolapse. A nonabsorbable suture plicates the prolapsed leaflet area towards the ventricular aspect in a triangular fashion by decreasing the suture width towards the leaflet base. Because no leaflet tissue is resected, this technique allows for the intraoperative correction of an imperfect plication. Triangular plication was successful in all except one patient. In this patient, a failed repair was corrected with mitral valve replacement. Freedom from mitral valve incompetence of more than grade 0-I was 100% at 12 months and 86% at 36 months postoperatively. PMID- 15276590 TI - Multivessel distal sutureless off-pump coronary artery bypass grafting procedure using magnetic connectors. AB - Proximal anastomotic devices for beating heart coronary artery bypass grafting (CABG) have been developed to avoid ascending aortic manipulation. Distal anastomotic devices may become an extremely useful tool to assist in enabling minimally invasive (robotic) multivessel CABG. As a transition phase toward this ultimate goal we have been using a distal anastomotic device for the left internal mammary artery-left anterior descending artery (LIMA-LAD) anastomosis. In addition we recently performed two off-pump coronary artery bypass procedures that were distally completely sutureless. PMID- 15276591 TI - Prognostic value of the admission electrocardiogram in patients with unstable angina/non-ST-segment elevation myocardial infarction treated with very early revascularization. AB - PURPOSE: The goals of this study were to determine if very early revascularization might ameliorate the adverse prognosis associated with ST segment depression in patients with unstable angina/non-ST-segment elevation myocardial infarction. METHODS: In this prospective cohort study, 1450 consecutive patients with unstable angina/non-ST-segment elevation myocardial infarction were stratified by the presence of ST-segment depression, T-wave inversion, or no changes on the admission electrocardiogram (ECG). All patients underwent coronary angiography and, if appropriate, revascularization within 24 hours after admission. The primary endpoint was all-cause mortality. RESULTS: During up to 59 months of follow-up, the in-hospital mortality rate was 2.1% (19/895) in patients with no ECG changes, 4% (6/136) in those with ST-segment depression, and 0.2% (1/419) in those with T-wave inversion. The cumulative death rate at 36 months was 8.0% (n = 49) in patients with no ECG changes, 19.9% (n = 18) in patients with ST-segment depression, and 5.1% (n = 13) in patients with T wave inversion (P = 0.0001 by log-rank). After adjustment for potential cofounders, ST-segment depression (hazard ratio [HR] = 2.2; 95% confidence interval [CI]: 1.1 to 4.6) and T-wave inversion (HR = 0.44; 95% CI: 0.20 to 0.96) were associated with long-term mortality. CONCLUSION: ST-segment depression and T wave inversion on the admission ECG were important predictors of outcome in patients with unstable angina/non-ST-segment elevation myocardial infarction undergoing very early revascularization. In contrast to the considerable mortality seen in patients with ST-segment depression, T-wave inversion was associated with a more favorable outcome. PMID- 15276592 TI - Effects of weekend admission and hospital teaching status on in-hospital mortality. AB - PURPOSE: The effect of reduced hospital staffing during weekends on in-hospital mortality is not known. We compared mortality rates between patients admitted on weekends and weekdays and whether weekend-weekday variation in rates differed between patients admitted to teaching and nonteaching hospitals in California. METHODS: The sample comprised patients admitted to hospitals from the emergency department with any of 50 common diagnoses (N = 641,860). Mortality between patients admitted on weekends and those admitted on weekdays (the "weekend effect") was compared. The magnitude of the weekend effect was also compared among patients admitted to major teaching, minor teaching, and nonteaching hospitals. RESULTS: The adjusted odds of death for patients admitted on weekends when compared with weekdays was 1.03 (95% confidence interval [CI]: 1.01 to 1.06; P = 0.0050). Three diagnoses (cancer of the ovary/uterus, duodenal ulcer, and cardiovascular symptoms) were associated with a statistically significant weekend effect. None of the 50 diagnoses demonstrated a statistically significant reduction in mortality for weekend admissions as compared with weekday admissions. Mortality was similar among patients admitted to major (odds ratio [OR] = 1.06; 95% CI: 0.94 to 1.19) and minor (OR = 1.03; 95% CI: 0.97 to 1.09) teaching hospitals, compared with nonteaching hospitals. However, the weekend effect was larger in major teaching hospitals compared with nonteaching hospitals (OR =1.13 vs. 1.03, P = 0.03) and minor teaching hospitals (OR = 1.05, P = 0.11). CONCLUSION: Patients admitted to hospitals on weekends experienced slightly higher risk-adjusted mortality than did patients admitted on weekdays. While overall mortality was similar for patients admitted to all hospital categories, the weekend effect was larger in major teaching hospitals and is cause for concern. PMID- 15276593 TI - Association of the ER22/23EK polymorphism in the glucocorticoid receptor gene with survival and C-reactive protein levels in elderly men. AB - PURPOSE: We recently demonstrated that a polymorphism in codons 22 and 23 of the glucocorticoid receptor gene is associated with relative glucocorticoid resistance, greater insulin sensitivity, and lower total and low-density lipoprotein cholesterol levels. In the present study, we investigated whether the ER22/23EK polymorphism is associated with survival, cholesterol levels, and two predictors of mortality: serum C-reactive protein and interleukin 6 levels. METHODS: We studied 402 men (mean [+/- SD] age, 77.8 +/- 3.6 years). C-reactive protein was measured by a highly sensitive method using a latex-enhanced immunoephelometric assay. Interleukin 6 was determined by a commercially available immulite assay. RESULTS: After a follow-up of 4 years, 73 (19%) of 381 noncarriers died, while none of the 21 ER22/23EK carriers had died (P = 0.03). C reactive protein levels were about 50% lower in ER22/23EK carriers (P = 0.01). There were no differences in interleukin 6 levels. CONCLUSION: Carriers of the ER22/23EK polymorphism have better survival than noncarriers, as well as lower C reactive protein levels. PMID- 15276594 TI - Differences in treatment outcome for hepatitis C among ethnic groups. AB - BACKGROUND: Studies of interferon-based therapies for hepatitis C virus (HCV) infected patients have documented variable response rates according to ethnicity. However, these studies enrolled low numbers of ethnic minorities. METHODS: Data from two multicenter trials of combination therapy for hepatitis C were analyzed to determine predictors of treatment success. The first trial was a randomized study comparing interferon administered three times weekly with daily administration. Patients in both interferon groups received weight-based ribavirin. The second trial was an observational study of daily interferon and ribavirin. Only treatment-naive patients were included in the analysis. Ethnicity (used as a nonspecific term to include race) was determined by patient self report. Sustained virologic response was defined as negative HCV RNA by polymerase chain reaction at 24 weeks after completion of therapy. RESULTS: A total of 661 patients (390 from the randomized trial and 271 from the observational trial) were available for analysis. Sustained virologic response was highest among Asians (61% [22/36]), followed by whites (39% [193/496]), Hispanics (23% [18/79]), and African Americans (14% [7/50]). In a multiple logistic regression model that adjusted for other factors known to affect treatment outcome, including hepatitis C genotype, Asians continued to be more likely to respond to treatment, whereas Hispanics and African Americans were less likely, as compared with whites. CONCLUSION: Sustained response rates to interferon and ribavirin therapy differ among ethnic groups. Ethnicity appears to be associated with treatment outcomes, even in a model that adjusts for other factors that influence response to therapy. PMID- 15276595 TI - Serum vitamin A concentration and the risk of hip fracture among women 50 to 74 years old in the United States: a prospective analysis of the NHANES I follow-up study. AB - BACKGROUND: Recent studies on the association between vitamin A and fracture risk have focused on samples with high vitamin A intake. We analyzed a cohort that was more representative of the overall U.S. population to test the hypothesis that both high and low serum vitamin A concentrations increase the risk of hip fracture. METHODS: We utilized data on 2799 women who were 50 to 74 years of age from the first National Health and Nutrition Examination Survey Epidemiologic Follow-up Study. There were 172 incident hip fractures during the 22-year follow up period. Using Cox regression analysis, we analyzed the relation between baseline serum vitamin A (retinol and retinyl esters) concentration, as a continuous variable and by quintiles, and hip fracture risk. RESULTS: While there was no linear relation between serum vitamin A concentration and the risk of hip fracture in the multivariate analysis (hazard ratio [HR] per SD increase = 1.0; 95% confidence interval [CI]: 0.9 to 1.2), analysis by quintiles revealed a U shaped relation between serum vitamin A concentration and hip fracture. Fracture risk was significantly higher among subjects in the lowest (HR = 1.9; 95% CI: 1.1 to 3.3) and highest (HR = 2.1; 95% CI: 1.2 to 3.6) quintiles compared with those in the middle quintiles. CONCLUSION: Both low and high serum vitamin A concentrations may be associated with an increased risk of hip fracture. PMID- 15276596 TI - Waiting for urgent procedures on the weekend among emergently hospitalized patients. AB - PURPOSE: Many hospital departments tend to have lower staffing levels on weekends. We evaluated the use of selected urgent procedures for emergently hospitalized patients and measured the time until procedure based upon the day of hospital admission. METHODS: We analyzed all acute care admissions from all 190 emergency departments in Ontario, Canada, between 1988 and 1997. We selected patients (n = 126,754) who underwent one of six prespecified procedures as their most responsible procedure: fiberoptic bronchoscopy, esophageal gastroduodenoscopy, magnetic resonance imaging, echocardiography, ventilation perfusion scanning, or coronary angiography. We noted each patient's day of procedure and day of hospital admission. For waits of less than 8 days, we analyzed the time to procedure based upon the day of admission. RESULTS: Only 5% (n = 5903) of the urgent procedures were performed on the weekend. Of the six selected procedures, coronary angiography showed the most skewed pattern of performance (1.5% performed on the weekend) and esophageal gastroduodenoscopy showed the least skewed pattern (8% performed on the weekend). Patients admitted on Fridays or Saturdays had the longest waits for procedures. For all six procedures, patients with relatively longer waits had relatively longer total in hospital stays (P <0.001 for each). CONCLUSION: Relatively few urgent procedures are performed in emergently hospitalized patients on the weekend, suggesting that greater attention to weekend care might result in more timely interventions and shorter lengths of stay. PMID- 15276597 TI - Abdominal computed tomography in the evaluation of patients with asymptomatic iron deficiency anemia: a prospective study. PMID- 15276598 TI - Should U.S. hospitals go 24/7? PMID- 15276599 TI - Effect of ammonia on the immune response of Taiwan abalone Haliotis diversicolor supertexta and its susceptibility to Vibrio parahaemolyticus. AB - Taiwan abalone Haliotis diversicolor supertexta held in 30 parts/per thousand seawater and 26 degrees C were injected with TSB-grown Vibrio parahaemolyticus (1.6 x 10(5)cfu abalone(-1)), and then placed in water containing different concentrations of ammonia-N (un-ionized plus ionized ammonia) at 0.01 mg l(-1) (control), 1.12, 3.22, 5.24 and 10.18 mg l(-1). Mortality of abalone increased directly with ambient ammonia-N concentration. After 12 h, the mortality of V. parahaemolyticus-injected abalone held in 3.22 mg l(-1) ammonia-N was significantly higher than those placed in 1.12 mg l(-1) ammonia-N and the control solution. In another experiment, the abalone which had been exposed to control, 1.08, 3.16, 5.37 and 10.34 mg l(-1) ammonia-N for 24, 72 and 120 h were examined for THC (total haemocyte count), phenoloxidase activity, respiratory burst (release of superoxide anion), and phagocytic activity and clearance efficiency to V. parahaemolyticus. The abalone when exposed to 3.16 mg l(-1) ammonia-N had decreased THC after 72 h, and decreased phenoloxidase activity, phagocytic activity and clearance efficiency after 24 h. However, the abalone when exposed to 3.16 mg l(-1) ammonia-N had increased respiratory burst after 24 h. The immune parameters except superoxide anion seemed to be suppressed in a dose-dependent fashion after 24 h. It is concluded that ammonia caused a depression in immune parameters and an increase in mortality of H. diversicolor supertexta from V. parahaemolyticus infection. PMID- 15276600 TI - Effect of turpentine oil on C-reactive protein (CRP) production in rainbow trout (Oncorhynchus mykiss). AB - The effect of turpentine oil on C-reactive protein (CRP) production was studied in rainbow trout (Oncorhynchus mykiss). Serum CRP concentration was estimated by sandwich enzyme-linked immunosorbent assay using anti-rainbow trout CRP monoclonal antibody (mAb) AC4 and polyclonal antibody. Intracellular CRP was demonstrated by flow cytometry using anti-trout CRP mAb. Hepatocytes, head kidney macrophages, spleen lymphocytes and peripheral blood lymphocytes showed reaction against AC4, but RTG-2 fibroblastic line cells, derived from rainbow trout gonad did not. This is the first report on the detection of intracellular CRP in fish. CRP levels decreased significantly 1 day after intramuscular injection of turpentine oil and remained low for 14 days. Significant decreases in the expression of CRP in hepatocytes, head kidney macrophages and spleen lymphocytes after injection of turpentine oil were found. The reduction of serum CRP concentration after turpentine oil injection may be attributed to decreases in intracellular CRP synthesis. PMID- 15276601 TI - Monoclonal antibodies recognising serum immunoglobulins and surface immunoglobulin-positive cells of puffer fish, torafugu (Takifugu rubripes). AB - Immunoglobulin of the torafugu, Takifugu rubripes, was purified by a combination of precipitation by low ionic strength dialysis and gel filtration. The Ig was used to immunise mice for the production of monoclonal antibody (MAb). Supernatants of hybridoma cultures were screened by enzyme-linked immunosorbent assay using purified-torafugu Ig-coated plates, and two stable hybridomas producing MAbs against torafugu Ig were obtained. Sodium dodecyl sulphate polyacrylamide gel electrophoresis under reducing conditions and Western blotting indicated that one MAb (16F3) was specific for the deglycosylated heavy chain of torafugu, and the other MAb (4H5) did not bind to the reduced Ig, suggesting that 4H5 recognised the higher-order structure of Ig. Under non-reduced conditions, both MAbs recognised mainly a 750 kDa band and also minor bands of 672, 410 and 205 kDa. MAb 16F3- and 4H5-primed magnetic beads (Dynabeads) adsorbed 84.9+/-3.3% and 63.6+/-4.4% of the torafugu Ig, respectively. The Ig adsorbed by MAb 16F3 primed Dynabeads was reactive to 4H5 on immunoblotting, and vice versa, indicating that the epitopes for both MAbs are held on the same Ig molecule. Both of these MAbs cross-reacted extensively with the Ig of other Takifugu species, but not with other genus. The MAbs were used to identify surface Ig-positive lymphocytes in the spleen, pronephros, peripheral blood and thymocytes of torafugu by flow cytometry. Flow cytometric analysis of the cells in the lymphocyte-enriched fraction revealed that 50.2+/-6.9% in the PBL, 11.8+/-1.7% in the mesonephros, 13.3+/-2.1% in the pronephros, 42.5+/-4.3% in the spleen and 3.2+/-0.6% in thymus were reactive to 4H5 or 16F3. PMID- 15276602 TI - Flow cytometric analysis of crayfish haemocytes activated by lipopolysaccharides. AB - Lipopolysaccharides (LPS) from Gram-negative bacteria are strong stimulators of white river crayfish, Procambarus zonangulus, haemocytes in vitro. Following haemocyte treatment with LPS and with LPS from rough mutant R5 (LPS Rc) from Salmonella minnesota, flow cytometric analysis revealed a conspicuous and reproducible decrease in cell size as compared to control haemocytes. These LPS molecules also caused a reduction in haemocyte viability as assessed by flow cytometry with the fluorescent dyes calcein-AM and ethidium homodimer. The onset of cell size reduction was gradual and occurred prior to cell death. Haemocytes treated with LPS from S. minnesota without the Lipid A moiety (detoxified LPS) decreased in size without a reduction of viability. The action of LPS on crayfish haemocytes appeared to be related to the activation of the prophenoloxidase system because phenoloxidase (PO)-specific activity in the supernatants from control and detoxified LPS-treated cells was significantly lower than that from LPS and LPS-Rc treated cells (P0.05) different, except for the total haemocyte number at 18 degrees C was significantly (P<0.05) higher than in crayfish at 4 degrees C. The percentage of granular cells in crayfish held at 4 degrees C was the highest compared to crayfish maintained at other temperatures. The phenoloxidase activities in haemocyte lysate supernatant (HLS) of crayfish at all temperature groups remained similar. The amount of proPO-mRNAs in haemocytes was much higher than the amount of LGBP-m RNAs in all the experimental groups. However, there was no change in the level of pro PO-mRNA at the tested temperatures. Interestingly, the level of LGBP-mRNA of crayfish kept at 22 degrees C was much lower than in those held at lower temperatures. Proliferation of the haematopoietic tissues was higher at high temperatures which may support replication of WSSV, and explain the high mortality of crayfish with WSSV infection at high temperature. Based on these studies it is concluded that crayfish might act as a carrier of WSSV at low water temperature and could develop white spot disease if the water temperature is increased. PMID- 15276607 TI - Immunogenic antigens of the eel pathogen Vibrio vulnificus serovar E. AB - The immunogenic antigens of Vibrio vulnificus serovar E were investigated in the eel. Fish were vaccinated by immersion with Vulnivaccine (V), revaccinated 2 years later by intraperitoneal injection (RV) and bath infected 15 days post revaccination (RVI). The specific immune response in serum was followed in all groups, and selected sera were used for immunostaining of surface (SA) and extracellular antigens (ECA). Bacteria were grown in iron-rich (TSB and MSWYE) and iron-poor media (TSB and MSWYE plus human transferrin (TSB-T and MSWYE-T)) as well as eel serum (ES), and their SA and ECA were extracted and electrophoretically analysed. Cells grown in MSWYE-T and ES presented the same antigenic profiles, which suggests that iron-restriction is the main growth limiting factor in vivo. The electrophoretic pattern of SA, but not that of ECA, varied with iron-availability in the growth medium. Further, SA extracted from bacteria grown in iron restriction were strongly immunogenic for eels, especially after vaccination and infection. Among the immunogenic antigens over expressed in iron-restriction, three outer membrane proteins of around 70-80 kDa, including the putative receptor for vulnibactin, together with the rapid and slow migrating forms of the lipopolysaccharide (LPS), were identified. The response was not so evident in the case of capsule, which was not clearly stained with any of the eel sera. With respect to ECA, two proteins, identified as the V. vulnificus protease (Vvp) and the major outer membrane protein (OMP), probably liberated to the medium after cell death, were recognised by RV and, more strongly, by RVI sera. The specific antibodies against the mentioned OMPs, LPS bands and the Vvp may contribute to the protection of vaccinated eels against infection, giving a reasonable explanation for the high effectiveness of Vulnivaccine. PMID- 15276608 TI - Production of recombinant C5a from rainbow trout (Oncorhynchus mykiss): role in leucocyte chemotaxis and respiratory burst. AB - Activation of the complement system can lead to the formation of the membrane attack complex, in which the component C5 is cleaved into C5a and C5b fragments. The C5a anaphylatoxin is a very potent pro-inflammatory molecule that induces chemotaxis and respiratory burst processes in a variety of mammalian leucocytes. While C5a has been well studied in mammals, little is known about the structure and function of C5a in teleost fish or other non-mammalian species. In the present study, we have produced and purified recombinant rainbow trout C5a (rtC5a), and we have shown that it plays an important role in inducing leucocyte migration as well as in triggering the respiratory burst of peripheral blood (PBLs) and head kidney leucocytes (HKLs). When the carboxy-terminal Arg was removed from rtC5a, its ability to induce cell migration and superoxide production remained intact. Interestingly, we show that leucocytes migrating towards rtC5a attached to the plate with a well-spread circular morphology, whereas those migrating towards activated trout serum displayed more irregular and dendritic-like shapes. Our data suggest that the basic mechanisms of action of the C5a anaphylotoxin have remained conserved for more than 300 million years. PMID- 15276609 TI - The Toll of herpes simplex virus infection. AB - Herpes simplex virus (HSV) infections provoke an inflammatory cytokine response, but the innate pathogen-sensing mechanisms that transduce the signal for this response are poorly understood. Recent findings have revealed that Toll-like receptor (TLR) 2 initiates the inflammatory process, and surprisingly that the response the TLR triggers might be overzealous in its attempt to counter the attack by the virus. Other recent findings suggest complexity in the array of TLRs that are triggered by HSV and the cell types they activate. Here we discuss the new revelations about these guardians against HSV infection and the consequences of the alarms raised in the host that they are assigned to protect. PMID- 15276610 TI - EBV-associated nasopharyngeal carcinomas: from epidemiology to virus-targeting strategies. AB - Nasopharyngeal carcinoma is a human malignancy consistently associated with the Epstein-Barr virus. Exposure to non-viral carcinogens and genetic predisposition are other crucial etiologic factors. Tumor development appears to require the expression of a small subset of transforming viral RNAs and proteins with concomitant silencing of most other viral genes. Impairment of the interactions of viral proteins with cellular partners or disruption of viral latency might prove to be useful for novel therapeutic strategies. PMID- 15276611 TI - Morphological and functional asymmetry in alpha-proteobacteria. AB - The release of an increasing number of complete bacterial genomic sequences allows the evolutionary analysis of processes such as regulatory networks. CtrA is a response regulator of the OmpR subfamily, belonging to a complex regulatory network in the dimorphic bacterium Caulobacter crescentus. It coordinates the cell cycle with an asymmetric division, which is part of the adaptation of Caulobacter to poor-nutrient environments. CtrA is only found in alpha proteobacteria, a group of bacteria encompassing genera with very distinct lifestyles, including host-associated bacteria. Analyses of CtrA regulatory networks and morphological examinations of some alpha-proteobacteria are presented. Our observations suggest that the core of the CtrA regulation network is conserved and that alpha-proteobacteria divide asymmetrically. We propose that the two daughter cells might be differentiated bacteria, each one displaying specific functions. PMID- 15276612 TI - Bacterial linguistic communication and social intelligence. AB - Bacteria have developed intricate communication capabilities (e.g. quorum sensing, chemotactic signaling and plasmid exchange) to cooperatively self organize into highly structured colonies with elevated environmental adaptability. We propose that bacteria use their intracellular flexibility, involving signal transduction networks and genomic plasticity, to collectively maintain linguistic communication: self and shared interpretations of chemical cues, exchange of chemical messages (semantic) and dialogues (pragmatic). Meaning based communication permits colonial identity, intentional behavior (e.g. pheromone-based courtship for mating), purposeful alteration of colony structure (e.g. formation of fruiting bodies), decision-making (e.g. to sporulate) and the recognition and identification of other colonies - features we might begin to associate with a bacterial social intelligence. Such a social intelligence, should it exist, would require going beyond communication to encompass unknown additional intracellular processes to generate inheritable colonial memory and commonly shared genomic context. PMID- 15276613 TI - Multilocus sequence typing--what is resolved? AB - Nucleotide sequence-based methods for bacterial typing (multilocus sequence typing; MLST) allow rapid and global comparisons between results from different laboratories. Combining this advantage with the reduced cost of high throughput sequencing, increasing automation and the amenability of sequence data for evolutionary analysis, it seems inevitable that sequence-based typing will eventually predominate over gel-based methods such as pulsed-field gel electrophoresis (PFGE) for most bacterial species. The increasing availability of multiple genome sequences for single pathogenic species, and the recent development of many new MLST schemes, means that a re-examination of the utility of multilocus sequencing, and in particular the choice of gene loci, is now appropriate. PMID- 15276614 TI - Staphylococcus aureus: superbug, super genome? AB - Staphylococcus aureus is a common cause of infection in both hospitals and the community, and it is becoming increasingly virulent and resistant to antibiotics. The recent sequencing of seven strains of S. aureus provides unprecedented information about its genome diversity. Subtle differences in core (stable) regions of the genome have been exploited by multi-locus sequence typing (MLST) to understand S. aureus population structure. Dramatic differences in the carriage and spread of accessory genes, including those involved in virulence and resistance, contribute to the emergence of new strains with healthcare implications. Understanding the differences between S. aureus genomes and the controls that govern these changes is helping to improve our knowledge of S. aureus pathogenicity and to predict the evolution of super-superbugs. PMID- 15276615 TI - Unravelling rhizosphere-microbial interactions: opportunities and limitations. AB - The rhizosphere is a biologically active zone of the soil around plant roots that contains soil-borne microbes including bacteria and fungi. Plant-microbe interactions in the rhizosphere can be beneficial to the plant, the microbes or to neither of them. One of the major difficulties that plant biologists and microbiologists face when studying these interactions is that many groups of microbes that inhabit this zone are not cultivable in the laboratory. Recent developments in molecular biology methods are shedding some light on rhizospheric microbial diversity. This review discusses recent findings and future challenges in the study of plant-microbe interactions in the rhizosphere. PMID- 15276616 TI - The role of Paneth cells and their antimicrobial peptides in innate host defense. AB - The intestinal epithelium is the largest surface area that is exposed to various pathogens in the environment, however, in contrast to the colon the number of bacteria that colonize the small intestine is extremely low. Paneth cells, one of four major epithelial cell lineages in the small intestine, reside at the base of the crypts and have apically oriented secretory granules. These granules contain high levels of antimicrobial peptides that belong to the alpha-defensin family. Paneth cells secrete these microbicidal granules that contain alpha-defensins when exposed ex vivo to bacteria or their antigens, and recent evidence reveals that antimicrobial peptides, particularly alpha-defensins, that are present in Paneth cells contribute to intestinal innate host defense. PMID- 15276617 TI - Rapid yeast estrogen bioassays stably expressing human estrogen receptors alpha and beta, and green fluorescent protein: a comparison of different compounds with both receptor types. AB - Previously, we described the construction of a rapid yeast bioassay stably expressing human estrogen receptor (hERalpha) and yeast enhanced green fluorescent protein (yEGFP) in response to estrogens. In the present study, the properties of this assay were further studied by testing a series of estrogenic compounds. Furthermore, a similar assay was developed based on the stable expression of human estrogen receptor beta (hERbeta). When exposed to 17beta estradiol, the maximum transcriptional activity of the ERbeta cytosensor was only about 40% of the activity observed with ERalpha, but the concentration where half maximal activation is reached (EC50), was about five times lower. The relative estrogenic potencies (REP), defined as the ratio between the EC50 of 17beta estradiol and the EC50 of the compound, of the synthetic hormones dienestrol, hexestrol and especially mestranol were higher with ER, while DES was slightly more potent with ERbeta. The gestagens progesterone and medroxyprogesterone acetate showed no response, whereas the androgen testosterone showed a very weak response. The anabolic agent, 19-nortestosterone showed a clear dose-related response with estrogen receptor but not beta. The phytoestrogens coumestrol, genistein, genistin, daidzein, daidzin and naringenin were relatively more potent with ERbeta. Ranking of the estrogenic potency with ER was: 17beta-estradiol >> 8 prenylnaringenin > coumestrol > zearalenone >> genistein >> genistin > naringenin. The ranking with the ERbeta was: 17beta-estradiol >> coumestrol > genistein > zearalenone > 8-prenylnaringen >> daidzein > naringenin > genistin >> daidzin. The hop estrogen 8-prenylnaringenin is relatively more potent with ERalpha. These data show that the newly developed bioassays are valuable tools for the rapid and high-throughput screening for estrogenic activity. PMID- 15276618 TI - Use of artificial androgen receptor coactivators to alter myoblast proliferation. AB - Skeletal muscle has long been thought to be a target tissue for androgens, eliciting their effect through the androgen receptor. In order to better understand androgen receptor action, a series of mutated androgen receptors were developed and their degree of specificity and cellular responses determined. Specificity, as measured by a reporter assay using HeLa cells, indicated that mutation of the ligand-binding domain or the AR (mutation H865Y), in combination with the p65 transactivating domain, resulted in an increased response to androgens as well as decreased specificity. Transfection of the mutant AR into mouse and rat myoblast cell lines resulted in an increase in expression of the reporter gene consistent with the data from HeLa cells. Overexpression of the wild type or mutant AR into myoblasts and treatment with testosterone induced both greater proliferation and faster differentiation of the cells compared to those expressing endogenous AR. Additionally, when treated with estrogen, these cells were able to proliferate and differentiate to similar levels as cells treated with testosterone. The ability of the mutated AR to act as an artificial coactivator to up-regulate androgen responsive genes is a useful tool for understanding the interaction of androgens and muscle growth. PMID- 15276619 TI - Identification of estrogen-responsive genes in the GH3 cell line by cDNA microarray analysis. AB - To identify estrogen-responsive genes in somatolactotrophic cells of the pituitary gland, a rat pituitary cell line GH3 was subjected to cDNA microarray analysis. GH3 cells respond to estrogen by growth as well as prolactin synthesis. RNAs extracted from GH3 cells treated with 17beta-estradiol (E2) at 10(-9) M for 24 h were compared with the control samples. The effect of an antiestrogen ICI182780 was also examined. The array analysis indicated 26 genes to be up regulated and only seven genes down-regulated by E2. Fourteen genes were further examined by real-time RT-PCR quantification and 10 were confirmed to be regulated by the hormone in a dose-dependent manner. Expression and regulation of these genes were then examined in the anterior pituitary glands of female F344 rats ovariectomized and/or treated with E2 and 8 out of 10 were again found to be up regulated. Interestingly, two of the most estrogen-responsive genes in GH3 cells were strongly dependent on E2 in vivo. #1 was identified as calbindin-D9k mRNA, with 80- and 118-fold induction over the ovariectomized controls at 3 and 24 h, respectively, after E2 administration. #2 was found to be parvalbumin mRNA, with 30-fold increase at 24 h. Third was c-myc mRNA, with 4.5 times induction at 24 h. The levels were maintained after one month of chronic E2 treatment. Identification of these estrogen-responsive genes should contribute to understating of estrogen actions in the pituitary gland. PMID- 15276620 TI - Phospholipase C-delta1 rescues intracellular Ca2+ overload in ischemic heart and hypoxic neonatal cardiomyocytes. AB - Ischemia and simulated ischemic conditions cause intracellular Ca2+ overload in the myocardium. The relationship between ischemia injury and Ca2+ overload has not been fully characterized. The aim of the present study was to investigate the expression and characteristics of PLC isozymes in myocardial infarction-induced cardiac remodeling and heart failure. In normal rat heart tissue, PLC-delta1 (about 44 ng/mg of heart tissue) was most abundant isozymes compared to PLC gamma1 (6.8 ng/mg) and PLC-beta1 (0.4 ng/mg). In ischemic heart and hypoxic neonatal cardiomyocytes, PLC-delta1, but not PLC-beta1 and PLC-gamma1, was selectively degraded, a response that could be inhibited by the calpain inhibitor, calpastatin, and by the caspase inhibitor, zVAD-fmk. Overexpression of the PLC-delta1 in hypoxic neonatal cardiomyocytes rescued intracellular Ca2+ overload by ischemic conditions. In the border zone and scar region of infarcted myocardium, and in hypoxic neonatal cardiomyocytes, the selective degradation of PLC-delta1 by the calcium sensitive proteases may play important roles in intracellular Ca2+ regulations under the ischemic conditions. It is suggested that PLC isozyme-changes may contribute to the alterations in calcium homeostasis in myocardial ischemia. PMID- 15276621 TI - Effect of the relation between neural cholinergic action and nitric oxide on ovarian steroidogenesis in prepubertal rats. AB - The coeliac ganglion and the ovary are related by the superior ovarian nerve, which penetrates into the ovary by the hilium and innervates mainly the ovarian stroma. On the other hand, it is known that the gaseous neurotransmitter nitric oxide (NO) and the two isoforms of its synthesis enzyme, the nitric oxide synthetase (NOS), are present in the ovary. Both innervation and NO participate in ovarian steroidogenesis. Therefore, the purposes of this work were (a) to standardize an in vitro coeliac ganglion-superior ovarian nerve-ovary integrated system in prepubertal rats; (b) to determine the presence of NO in the ovary and analyze the ganglionic cholinergic effect on the ovarian release of androstenedione, progesterone and NO; and (c) to assess the steroids/NO relationship. The system was incubated in buffer solution for 120 min, with the ganglion and ovary located in different compartments and linked by the superior ovarian nerve. From the results obtained, it is concluded that the system is viable and functional. The presence of basal NO is stimulated by the cholinergic action, while the release of the steroids is inhibited, which might indicate that the ganglionic cholinergic effect is probably mediated by NO. To our knowledge, this work constitutes the first study of the relationship between the neural cholinergic action and NO on the ovarian steroidogenesis of prepubertal rats. PMID- 15276622 TI - Estrogen-like activity of licorice root constituents: glabridin and glabrene, in vascular tissues in vitro and in vivo. AB - Post-menopausal women have higher incidence of heart diseases compared to pre menopausal women, suggesting a protective role for estrogen. The recently Women's Health Initiative (WHI) randomized controlled trial concluded that the overall heart risk exceeded benefits from use of combined estrogen and progestin as hormone replacement therapy for an average of five years among healthy postmenopausal US women. Therefore, there is an urgent need for new agents with tissue-selective activity with no deleterious effects. In the present study, we tested the effects on vascular tissues in vitro and in vivo of two natural compounds derived from licorice root: glabridin, the major isoflavan, and glabrene, an isoflavene, both demonstrated estrogen-like activities. Similar to estradiol-17beta (E2), glabridin (gla) stimulated DNA synthesis in human endothelial cells (ECV-304; E304) and had a bi-phasic effect on proliferation of human vascular smooth muscle cells (VSMC). Raloxifene inhibited gla as well as E2 activities. In animal studies, both intact females or after ovariectomy, gla similar to E2 stimulated the specific activity of creatine kinase (CK) in aorta (Ao) and in left ventricle of the heart (Lv). Glabrene (glb), on the other hand, had only the stimulatory effect on DNA synthesis in vascular cells, with no inhibition by raloxifene, suggesting a different mechanism of action. To further elucidate the mechanism of action of glb, cells were pre-incubated with glb and then exposed to either E2 or to gla; the DNA stimulation at low doses was unchanged but there was abolishment of the inhibition of VSMC cell proliferation at high doses as well as inhibition of CK stimulation by both E2 and by gla. We conclude that glb behaved differently than E2 or gla, but similarly to raloxifene, being a partial agonist/antagonist of E2. Glabridin, on the other hand, demonstrated only estrogenic activity. Therefore, we suggest the use of glb with or without E2 as a new agent for modulation of vascular injury and atherogenesis for the prevention of cardiovascular diseases in post-menopausal women. PMID- 15276623 TI - Partitioning of 5alpha-dihydrotestosterone and 5alpha-androstane-3alpha, 17beta diol activated pathways for stimulating human prostate cancer LNCaP cell proliferation. AB - The growth and development of the prostate gland are regulated by androgens. Despite our understanding of molecular actions of 5alpha-dihydrotestosterone (5alpha-DHT) in the prostate through the trans-activation of the androgen receptor (AR), comprehensive analysis of androgen responsive genes (ARGs) has just been started. Moreover, expression changes induced by the androgen effects of 5alpha-androstane-3alpha,17beta-diol (3alpha-diol), a metabolite of 5alpha-DHT through the action of 3alpha-hydroxysteroid dehydrogenases (3alpha-HSDs), remain undefined. We demonstrated that both 5alpha-DHT and 3alpha-diol stimulated similar levels of androgen sensitive human prostate cancer LNCaP cell proliferation. However, consistent with the fact that 3alpha-diol has low affinity toward the AR, 3alpha-diol did not elicit the same levels of AR trans activation activity as that of 5alpha-DHT. Since LNCaP cells respond to androgen stimulation by transcriptionally activating the AR downstream genes, gene expression patterns between 0 and 48 h following 3alpha-diol and 5alpha-DHT stimulation were analyzed using cDNA-based membrane arrays to define the temporal regulation of ARGs. Array analysis identified 217 and 219 androgen-modulated genes in at least one time point following 3alpha-diol and 5alpha-DHTstimulation, respectively, including key regulators of cell proliferation. Only a subset of these genes (143) was regulated by both androgens. These data suggest that 3alpha diol exerts androgenic effects independent of the action of 5alpha-DHT in steroid target tissues. Accordingly, 3alpha-diol might activate cell proliferation cascades independent of AR pathway in the prostate. PMID- 15276624 TI - Analysis of the putative regulatory region of the gastric inhibitory polypeptide receptor gene in food-dependent Cushing's syndrome. AB - Gastric inhibitory polypeptide (GIP)-dependent Cushing's syndrome (CS) results from the ectopic expression of non-mutated GIP receptor (hGIPR) in the adrenal cortex. We evaluated whether mutations or polymorphisms in the regulatory region of the GIPR gene could lead to this aberrant expression. We studied 9.0kb upstream and 1.3kb downstream of the GIPR gene putative promoter (pProm) by sequencing leukocyte DNA from controls and from adrenal tissues of GIP- and non GIP-dependent CS patients. The putative proximal promoter region (800 bp) and the first exon and intron of the hGIPR gene were sequenced on adrenal DNA from nine GIP-dependent CS, as well as on leukocyte DNA of nine normal controls. Three variations found in this region were found in all patients and controls; at position -4/-5, an insertion of a T was seen in four out of nine patients and in five out of nine controls. Transient transfection studies conducted in rat GC and mouse Y1 cells showed that the TT allele confers loss of 40% in the promoter activity. The analysis of the 8-kb distal pProm region revealed eight distal single nucleotide polymorphisms (SNPs) without probable association with the disease, since frequencies in patients and controls were very similar. In conclusion, mutations or SNPs in the regulatory region of the GIPR gene are unlikely to underlie GIP-dependent CS. PMID- 15276625 TI - Release of ovarian progesterone during the rat oestrous cycle by ganglionic cholinergic influence: the role of norepinephrine. AB - The coeliac ganglion neurons, whose axons constitute the superior ovarian nerve (SON), contain cholinergic receptors. The aim of this work was to study the effect of cholinergic agents added to the coeliac ganglion on the release of ovarian progesterone in the coeliac ganglion-SON-ovary in vitro system. We also analyzed the release of norepinephrine in the ovarian compartment and its possible relationship with the release of progesterone. After the addition of cholinergic agents in the ganglion compartment, progesterone release was determined by radioimmuneassay (RIA) and norepinephrine by catecholamine assay (HPLC). The release of progesterone and norepinephrine in the ovary compartment was studied during period of 180 min in pre-oestrus (PE), oestrus (E), dioestrus day 1 (D1) and dioestrus day 2 (D2) rats. The most relevant results concerning the action of acetylcholine were found on PE and dioestrus. On PE, the pre ovulatory peak of progesterone, which is known to respond to the endocrine action, was not modified by neural effect of acetylcholine in our scheme. On the other hand, the progesterone peak occurs in the afternoon of D1, which has been described as independent of the gonadotrophic action but was inhibited by neural effect of acetylcholine in our experimental scheme. This action on D1 was accompanied by a decrease of norepinephrine release in the ovary compartment. We conclude that the action of cholinergic agents varies according to the oestrous cycle stage and constitutes one of the factors governing the secretory activity of the ovarian steroids, in this case, progesterone. PMID- 15276626 TI - Purification and characterization of 7beta-hydroxysteroid dehydrogenase from rabbit liver microsomes. AB - 7beta-Hydroxysteroid dehydrogenase (7beta-HSD), a specific enzyme active in the metabolization of 7beta-hydroxycholesterol, was purified about 300-fold from male rabbit liver microsomes using ion exchange, hydroxylapatite, 2'5'ADP Sepharose 4B, and high-performance liquid chromatography on the basis of its catalytic activity. The specific activity of the purified enzyme was 276 nmol/min/mg protein. The molecular weight of the purified enzyme was 34,000. The preferred coenzyme was beta-NADP+. The optimum pH for oxidation was around 7.7 in potassium phosphate buffer, and 11.0 in glycine-NaOH buffer. The purified enzyme catalyzed the synthesis of not only 7beta-hydroxycholesterol but also corticosterone and hydrocortisone. Enzyme activities toward these three substrates accompanied all purification steps of 7beta-HSD. The amino acid sequence of the N-terminal of the purified enzyme showed that 7beta-HSD had sequence similarity to rabbit type I 11beta-hydroxysteroid dehydrogenase (11beta-HSD), indicating that 7beta-HSD may belong to the rabbit type I 11beta-HSD family and may play the same role in the metabolism of 11-hydroxysteroids and 7-hydroxysterols. PMID- 15276627 TI - Determination of protein adsorption by comparative drop and bubble profile analysis tensiometry. AB - Proteins adsorb at a liquid interface at very low bulk concentrations which is significantly decreased due to the adsorbed amount. Depending on the surface area A to solution bulk volume V ratio this loss in protein can be significant. Using experimental methods with large differences in the A/V ratio the adsorbed amount can be determined from the measured quantities, for example the surface tension. For beta-casein and bovine and human serum albumin data from literature obtained by the Wilhelmy technique and own data measured by drop profile analysis tensiometry are used. The adsorbed amounts measured directly from ellipsometry agree very well with those determined by the proposed method. PMID- 15276628 TI - Lipid composition-dependent incorporation of multiple membrane proteins into liposomes. AB - Membrane proteins from bacteria Pasteurella multocida were used as a model for studying its incorporation into liposomes. An important step to achieve efficient high yield protein incorporation in proteoliposomes is the study of the more suitable lipid composition. To this end, we compared the amount of total protein, reconstituted by co-solubilization methods, into liposomes of phospholipids with different polar head groups and acyl chain lengths. The liposomes and proteoliposomes were characterised by isopycnic centrifugation in sucrose gradient and by dynamic light scattering. Experimental and theoretical results were compared considering the effects exerted through the hydrocarbon chain length, volume, and optimal cross-sectional area of the phospholipid (combined in the geometrical critical packing parameter, lipid-protein matching), critical spontaneous radius of curvature of the bilayer vesicle, phase transition temperature of the lipid and ratio of lipid-protein molecules present in the vesicles. The highest incorporation of multiple proteins was found with dipalmitoylphosphatidylcholine (DPPC), reaching a yield of 93% compared to the lower relative amounts incorporated in proteoliposomes of the other lipids. The incorporation of multiple proteins induces a proportional enhancement of vesicular dimension, since DPPC-proteoliposomes have an average diameter of 1850A, compared to the 1430A for pure DPPC vesicles. PMID- 15276629 TI - Interfacial and foaming properties of prolylenglycol alginates. Effect of degree of esterification and molecular weight. AB - In the present work we have studied the characteristics of propylene glycol alginates (PGA) adsorption at the air-water interface and the viscoelastic properties of the films in relation to its foaming properties. To evaluate the effect of the degree of PGA esterification and viscosity, different commercial samples were studied--Kelcoloid O (KO), Kelcoloid LVF (KLVF) and Manucol ester (MAN). The temperature (20 degrees C) and pH (7.0) were maintained constant. For time-dependent surface pressure measurements and surface dilatational properties of adsorbed PGA at the air-water interface an automatic drop tensiometer was used. The foam was generated by whipping and then the foam capacity and stability was determined. The results reveal a significant interfacial activity for PGA due to the hydrophobic character of the propylene glycol groups. The kinetics of adsorption at the air-water interface can be monitored by the diffusion and penetration of PGA at the interface. The adsorbed PGA film showed a high viscoelasticity. The surface dilatational modulus depends on the PGA and its concentration in the aqueous phase. Foam capacity of PGA solutions increased in the order KO > MAN > KLVF, which followed the increase in surface pressure and the decrease in the viscosities of PGA solutions. The stability of PGA foams monitored by the drainage rate and collapse time follows the order MAN > KLVF > KO. The foam stability depends on the combined effect of molecular weight/degree of esterification of PGA, solution viscosity and viscoelasticity of the adsorbed PGA film. PMID- 15276630 TI - Attenuated total reflectance infrared studies of liposome adsorption at the solid liquid interface. AB - Interfacial interactions between liposomes and the solid-liquid interface (i.e. a ZnSe internal reflection element, modified to mimic a biological surface) were studied by Fourier transform infrared (FTIR) spectroscopy in attenuated total reflectance (ATR) mode. Both conventional liposomes, containing lecithin and cholesterol and Stealth liposomes containing poly(ethylene)glycol (PEG)5000- or PEG2000-lipids were investigated. IR bands due to the liposome components were observed to increase with time and enabled the liposome adsorption kinetics and thermodynamics to be quantified. The liposome solution conditions, surface properties and compositions have all been shown to influence liposome adsorption. Free energies of adsorption were determined to be in the range from -10.0 to 11.0 kJ mol(-1) and slightly reduced by PEG incorporation. The adsorption rate constant is decreased with increased solution pH and decreased ionic strength; this reflects the importance of electrostatics in controlling liposome adsorption. Increasing the level and molecular weight of PEG incorporation in the liposomes significantly reduced both the rate and extent of liposome adsorption; steric hindrance is considered to play a key role. Findings from this research will improve the understanding of liposome interaction during drug delivery, give insight into the actions of liposomes in the body and may form the basis for improved liposome formulations. PMID- 15276631 TI - A reversible adsorption-desorption interface of DNA based on nano-sized zirconia and its application. AB - It is essential for the information storage in DNA-based bio-chips to construct a reversible exchange interface of DNA. Here, a highly reproducible and reversible adsorption-desorption interface of DNA based on the nano-sized zirconia in different pH solution was successfully fabricated. The results showed that DNA can be adsorbed onto the nano-sized zirconia from its solution, and can desorb from the nanoparticles in 0.10 M KOH solution. When the matrix with nanoparticles returns to the DNA solution again, DNA can be re-adsorbed onto them as initial state. Moreover, the interaction of DNA with non-electroactive molecules, 2,2' bipyridine, has been studied by electrochemistry method in the aid of probe Co(phen)(3)(3+). The experiments showed that when 2,2'-bipyridine was added into the test solution, the voltammetric peak currents of Co(phen)(3)(3+) decreased; and the decrease value of peak current against the concentration of 2,2' bipyridine has a good Langmuir relationship, by which the equilibrium constant of interaction between 2,2'-bipyridine and DNA was estimated to be 1.57 x 10(4)M( 1). PMID- 15276632 TI - Monoolein cubic phases containing hydrogen peroxide. AB - Monoolein (MO) cubic phases were prepared by hydrating MO using distilled water or 12wt.% H(2)O(2) solution so that the content of aqueous phase in the cubic phase is 30wt.%. The thermal transition of the isotropic cubic phase to reversed hexagonal phase was observed on a polarizing photomicroscope and the transition temperature was found to be around 65 degrees C on a differential scanning calorimeter (DSC). Small-angle X-ray scattering (SAXS) patterns indicated the cubic phases had diamond surfaces. The cubic phase released H(2)O(2) into an aqueous phase in a saturation manner so that approx. 50% of total loaded H(2)O(2) release in the first 10 h and thereafter relatively slow was observed over 40 h. The cubic phase was stable at 45 degrees C for 56 days before it broke down into an oily phase and an aqueous phase in 70 days. According to (1)H NMR spectrum, glycerol moiety and -CH(2)=CH(2)- of the oily phase were detected less in number than those of intact MO. Therefore, the hydrolysis and the oxidation of MO would be responsible for the breakdown of the cubic phase. The tensile adhesive forces of the cubic phases were higher than a skin-adhesive patch prepared using polyacrylate. The cubic phase containing H(2)O(2) could be used as a topical disinfected gel for a wounded skin. PMID- 15276633 TI - Poly(ethylene glycol) enhances the surface activity of a pulmonary surfactant. AB - The primary role of lung surfactant is to reduce surface tension at the air liquid interface of alveoli during respiration. Axisymmetric drop shape analysis (ADSA) was used to study the effect of poly(ethylene glycol) (PEG) on the rate of surface film formation of a bovine lipid extract surfactant (BLES), a therapeutic lung surfactant preparation. PEG of molecular weights 3,350; 8,000; 10,000; 35,000; and 300,000 in combination with a BLES mixture of 0.5 mg/mL was studied. The adsorption rate of BLES alone at 0.5 mg/mL was much slower than that of a natural lung surfactant at the same concentration; more than 200 s are required to reach the equilibrium surface tension of 25 mJ/m(2). PEG, while not surface active itself, enhances the adsorption of BLES to an extent depending on its concentration and molecular weight. These findings suggest that depletion attraction induced by higher molecular weight PEG (in the range of 8,000 to 35,000) may be responsible for increasing the adsorption rate of BLES at low concentration. The results provide a basis for using PEG as an additive to BLES to reduce its required concentration in clinical treatment, thus reducing the cost for surfactant replacement therapy. PMID- 15276634 TI - A novel process for inking the stamp with biomacromolecule solution used in reactive microcontact printing. AB - This paper describes a novel process for inking the stamp with biomacromolecule solution used in reactive microcontact printing. The stamp was first coated with biomacromolecule solution such as bovine serum albumin (BSA) solution for 20 min, and then dried by soft nitrogen flow. After cooling the stamp below the dew point for 1 min and incubated at room temperature for 10 s, a thin layer of condensed water was formed on the stamp surface. Then, an aldehyde functionalized glass slide was pressed immediately onto the stamp for certain time, yielding the covalently patterning of the biomacromolecules on the aldehyde-containing surface. A notable feature of this process is that the biomacromolecule solution can be inked onto the stamp at a controlled state, neither too "dry" nor too "wet". As a result, the covalently grafting reaction can occur at a comparable speed with those in solution, while avoiding the contamination caused by dispersal of excessive solvent. PMID- 15276635 TI - Heteronuclear polarization transfer by symmetry-based recoupling sequences in solid-state NMR. AB - We demonstrate a new set of methods for transferring spin polarization between different nuclear isotopes in magic-angle-spinning solid-state NMR. The technique employs symmetry-based recoupling sequences on one irradiation channel and a simple sequence of between one and three strong radiofrequency pulses on the second channel. A phase shift of the recoupling sequences is applied at the same time as a pi/2 pulse on the second channel. The trajectory of the transferred polarization may be used to estimate heteronuclear distances. The method is particularly attractive for nuclei with low gyromagnetic ratios or for those experiencing strong anisotropic spin interactions, where conventional Hartmann Hahn cross-polarization is difficult to apply. We demonstrate the method on 1H 13C, 1H-15N and 19F-109Ag systems. PMID- 15276636 TI - A low-resolution non-invasive NMR characterization of ancient paper. AB - The use of a portable NMR device allows a non-invasive investigation of the paper in order to assess the state of conservation of books and documents of historical or artistic interest. The NMR investigation has been found mainly on relaxation measurements whose results seem compatible with different relaxation rates in crystalline and amorphous cellulose domains. By a simplified physical picture based on spin-diffusion it appears possible to detect the alteration of the crystalline/amorphous cellulose balance and therefore to get information on some deterioration processes of paper. The use of a portable NMR device shows great potentiality because of its safe and simple in situ approach to Cultural Heritage documents. In this work, we present a research carried out on a 17th century manuscript. PMID- 15276637 TI - Tracer diffusion measurements in solid lithium: a test case for the comparison between NMR in static and pulsed magnetic field gradients after upgrading a standard solid state NMR spectrometer. AB - This paper reports on the upgrading of a standard solid state NMR spectrometer, which has been used in combination with a field variable 7 T cryomagnet, to a low cost combined SFG and PFG NMR spectrometer. Both methods are applied to solid lithium as a simple test case. The results show that under the given conditions SFG NMR and PFG NMR can provide tracer diffusion coefficients for 7 Li diffusion down to about 10(-14) and 10(-13) m2/s, respectively. SFG and PFG NMR are complementary methods. The paper demonstrates advantages and disadvantages of each method with a concrete example and why it is desirable to be able to apply both methods to the same sample. PMID- 15276638 TI - MAS-NMR at very high temperatures. AB - We report MAS-NMR experiments at temperatures of approx. 1200 K using a CO(2) laser as the heating device. An internal NMR thermometer based on the (7)Li T1 data of Li(0.24)La(0.54)TiO(3) is used for temperature calibration. Using this setup, temperatures as high as 1191 K could be reached under MAS conditions as confirmed by the melting of Li(2)B(4)O(7) at 1191 K which could be followed by (7)Li-MAS-NMR. PMID- 15276639 TI - A lattice model for the simulation of one and two dimensional 129Xe exchange spectra produced by translational diffusion. AB - Xenon-129 spectra in some heterogeneous polymer systems consist of two resonances which collapse to a single resonance as a function of temperature. Two different resonances arise from spatially separated, distinct sorption environments and spectral collapse occurs when xenon atoms diffuse from one environment to the other at a sufficiently fast rate. This exchange mechanism involves a distribution of time constants and a two domain lattice model is used to generate a realistic distribution of correlation times resulting from diffusion in a heterogeneous matrix. The distribution of correlation times is inhomogeneous in the sense that different xenon atoms would exchange between the two domains or environments with a variety of time constants and the resulting spectrum is a superposition of spectra associated with each of the time constants. To demonstrate the nature of exchange according to this model, diffusion out of a sphere is simulated which corresponds to a progressive saturation experiment used to determine the diffusion constant of xenon in polystyrene. Then the model is used to demonstrate the difference between homogeneous and heterogeneous spectral collapse in one- and two-dimensional examples. Lastly, the simulation model is used to interpret one- and two- dimensional xenon-129 line shape changes for xenon sorbed into poly(2,6-dimethyl-1,4-phenylene oxide) as a function of temperature. Two broad resonances are observed at low temperatures in this polymer corresponding to xenon-129 sorbed in high free volume and low free volume domains. Exchange between the two main resonances collapses the spectrum to a single peak at higher temperatures. Both the collapse in one dimension and exchange in two dimensions as a function of mixing time can be simulated using the distribution from the lattice model. An average domain size of 70 nm is estimated by combining the simulation of the exchange experiment with the results of a one-dimensional progressive saturation experiment. The size of the sites sorbing individual xenon atoms has been reported from positron annihilation lifetime spectroscopy as 1.4 nm for the high free volume sites and 0.3 nm for the low free volume sites. The domain size is more than an order of magnitude larger than the individual sorption site indicating that domains consist of many sites as assumed in the lattice model description. PMID- 15276640 TI - High resolution 31P NMR study of octacalcium phosphate. AB - We have assigned the (31)P high-resolution spectrum of octacalcium phosphate by (31)P double quantum and HETCOR spectroscopy. The (31)P peaks at -0.2, 2.0, 3.3 and 3.7 ppm are assigned to P5/P6, P3, P2/P4 and P1, respectively. Our data reveal that substantial amount of the PO(4)(3-) groups at the P2 and P4 sites have been transformed to HPO(4)(2-) in our octacalcium phosphate sample. PMID- 15276641 TI - Solid-state 17O nuclear magnetic resonance spectroscopy without isotopic enrichment: direct detection of bridging oxygen in radiation damaged zircon. AB - Protocols are presented for obtaining natural abundance (17)O magic angle spinning and static NMR spectra in the solid state. Rotor-assisted population transfer (RAPT), Carr-Purcell-Meiboom-Gill (CPMG) echo trains and cross polarisation (CP) are all used to obtain spectra of sites with large as well as small electric field gradients in proton and non-proton containing inorganic materials. Spectra are of sufficient quality to obtain the typical NMR parameters by standard fitting of the spectra. The protocol is then applied to identifying the changes that accompany radioactive decay in zircon (ZrSiO(4)) where enrichment is impossible. The (17)O NMR spectra of a partially metamict zircon sample clearly show evidence of bridging oxygens being produced as a consequence of radiation damage. The spectra have been acquired at moderate magnetic fields over periods typically of 60 h (1 weekend) and it is concluded that a 'routine' overnight (17)O experiment of 15 h at high field (e.g. 21 T) may well be possible. PMID- 15276642 TI - Interaction of ACE2 and integrin beta1 in failing human heart. AB - ACE2 purified from failing human heart was found to form a complex with integrin beta1 by immunoprecipitation, Western blotting, activity assay, and ESI tandem mass spectroscopy. The ACE2/integrin complex showed a Km of 6.8 microM and a Vmax of 2.13 pmol/min/microl purified enzyme. Activity was optimal at pH 7.5 with Ang II substrate. PMID- 15276643 TI - Cytosolic carbonic anhydrase activity in chronic myeloid disorders with different clinical phenotype. AB - Carbonic anhydrase family (CAs) plays an important role in the extracellular acidification and several studies suggest a possible involvement of such enzymes in the increased tumor progression due to the acidic extracellular pH. We measured the activities of carbonic anhydrase I and II isoforms in a group of patients affected by four specific chronic haematological diseases, sharing a common origin but characterized by a different neoplastic evolution: agnogenic myeloid metaplasia (AMM), essential thrombocythemia (ET), chronic myeloid leukemia (CML) and polycythemia vera (PV) in order to understand the correlation between CAs activities and neoplastic outcome. In comparison to controls, our data demonstrate an increase of CAI and CAII activities in all our patients with a specific increase of the CAI activity in the group of the diseases with major malignancy (CML and AMM). These results suggest a possible role of such isozymes in the progression of the myeloid disorders and CAs specific inhibitors should be useful in slowing the progression of the disease. PMID- 15276644 TI - Oxidant stress in type I autoimmune hepatitis: the link between necroinflammation and fibrogenesis? AB - Autoimmune hepatitis (AIH) is a chronic liver disease of unknown aetiology characterized by circulating autoantibodies, hyperglobulinaemia and interface hepatitis. The mechanisms of progression from initial autoimmune attack to fibrosis and cirrhosis are unclear but oxidant stress may be involved. Markers of lipid peroxidation, antioxidant status, hepatic fibrogenesis and liver function were measured in blood and urine in 35 controls and in 33 patients with type-1 AIH; histology was assessed in 18 patients. In AIH, markers of lipid peroxidation were significantly elevated (8-isoprostane in both plasma and urine P < 0.001; plasma malondialdehyde P = 0.017). Total antioxidant capacity in protein-free serum and total glutathione in both whole blood and plasma were significantly reduced (P = 0.007, P = 0.037, P < 0.001, respectively). The antioxidants selenium, vitamin A and vitamin E were significantly decreased (P = 0.007, P < 0.001, P = 0.025, respectively); vitamin C was unchanged. Urinary 8-isoprostane correlated positively with interface hepatitis and necroinflammatory score and with hepatic fibrogenesis (type III procollagen peptide). Interface hepatitis correlated negatively with vitamin A and whole blood total glutathione. Oxidant stress, as reflected in blood and urine by a wide range of pro- and antioxidant markers, is a significant feature of AIH and provides a probable mechanism linking hepatic necroinflammation to fibrogenesis and disease progression. PMID- 15276645 TI - The effect of finger millet feeding on the early responses during the process of wound healing in diabetic rats. AB - In the present study, the role of finger millet feeding on skin antioxidant status, nerve growth factor (NGF) production and wound healing parameters in healing impaired early diabetic rats is reported. Hyperglycemic rats received food containing 50 g/100 g finger millet (FM). Non-diabetic controls and diabetic controls received balanced nutritive diet. Full-thickness excision skin wounds were made after 2 weeks prior feeding of finger millet diet. The rate of wound contraction, and the levels of collagen, hexosamine and uronic acid in the granulation tissue were determined. The skin antioxidant status and lipid peroxide concentration were also monitored during the study. In hyperglycemic rats fed with finger millet diet, the healing process was hastened with an increased rate of wound contraction. Skin levels of glutathione (GSH), ascorbic acid and alpha-tocopherol in alloxan-induced diabetic rat were lower as compared to non-diabetics. Altered activities of superoxide dismutase (SOD) and catalase (CAT) were also recorded in diabetics. Interestingly, thiobarbituric acid reactive substances (TBARS) were elevated in the wound tissues of all the groups, when compared to normal (unwounded) skin tissues. However, in diabetic rats the TBARS levels of both normal and wounded skin tissues were significantly elevated (P < 0.001) when compared with control (non-diabetic) and diabetics fed with FM. Impaired production of NGF, determined by ELISA, in diabetic rats was improved upon FM feeding and further confirmed by immunocytochemical observations reflects the increased expression of NGF in hyperglycemic rats supplemented with FM enriched diet. Histological and electron microscopical evaluations revealed the epithelialization, increased synthesis of collagen, activation of fibroblasts and mast cells in FM-fed animals. Thus, increased levels of oxidative stress markers accompanied by decreased levels of antioxidants play a vital role in delaying wound healing in diabetic rats. However, FM feeding to the diabetic animals, for 4 weeks, controlled the glucose levels and improved the antioxidant status, which hastened the dermal wound healing process. PMID- 15276646 TI - Involvement of caspase-10 in advanced glycation end-product-induced apoptosis of bovine retinal pericytes in culture. AB - Apoptosis appears to be the death mechanism of pericyte loss observed in diabetic retinopathy. We have previously shown that advanced glycation end-products (AGE MGX) induce apoptosis of retinal pericytes in culture associated with diacylglycerol (DAG)/ceramide production. In the present study, we investigated possible caspase involvement in this process. Bovine retinal pericytes (BRP) were cultured with AGE-MGX and apoptosis examined after annexin V staining. Effects of peptidic inhibitors of caspases were determined on DAG/ceramide production and apoptosis. Pan-caspase inhibitor z-VAD-fmk (50 microM) was able to inhibit both DAG/ceramide production and apoptosis, whereas caspase-3-like inhibitor z-DEVD fmk (50 microM) or caspase-9 inhibitor z-LEHD-fmk (50 microM) was only active on apoptosis. This differential effect strongly suggests involvement of initiator caspase(s) upstream and effector caspase(s) downstream DAG/ceramide production in AGE-mediated apoptosis. Pericyte treatment with caspase-8 inhibitor z-IETD-fmk (50 microM) did not protect cells against AGE-induced apoptosis and we failed to detect caspase-8 in pericytes by immunoblotting assay. Interestingly, one inhibitor of caspase-10 and related caspases z-AEVD-fmk (50 microM) inhibited both AGE-MGX-induced apoptosis and DAG/ceramide formation in pericytes. Cleavage of caspase-10 precursor into its active subunits was demonstrated by immunoblotting assay in pericytes incubated with AGE-MGX. These results strongly suggest that caspase-10, but not caspase-8, might be involved in the early phase of AGE-induced pericyte apoptosis, in contrast to caspase-9 and -3-like enzymes involved after DAG/ceramide production. This finding may provide new therapeutic perspectives for early treatment in diabetic retinopathy. PMID- 15276647 TI - Increased gut-derived norepinephrine release in sepsis: up-regulation of intestinal tyrosine hydroxylase. AB - Studies have shown that increased gut-derived norepinephrine (NE) release plays an important role in producing hepatocellular dysfunction at the early stage of sepsis. Although the gut has been demonstrated to be the major source of NE in sepsis, it remains unknown whether the increased NE is associated with up regulation of intestinal NE biosynthesis enzymes such as tyrosine hydroxylase (TH) and dopamine beta-hydroxylase (DBH). To determine this, adult male rats were subjected to sepsis by cecal ligation and puncture (CLP) followed by fluid resuscitation. Small intestinal samples were harvested at 2 h (i.e., early sepsis) or 20 h (late sepsis) after CLP or sham-operation. Protein levels of TH and DBH were determined by Western blot analysis and immunohistochemistry. Their gene expression was assessed by RT-PCR technique. The results indicate that intestinal TH protein levels increased significantly at 2 and 20 h after CLP, while DBH was not altered under such conditions. Immunohistochemical examination shows that both TH and DBH were located in intestinal sympathetic nerve fibers and TH staining was markedly increased in septic animals. TH gene expression increased significantly at 2 h but not at 20 h after CLP, while DBH gene expression was not altered in sepsis. Thus, the increased TH gene and protein expression appears to be responsible for the increased gut-derived NE in sepsis. PMID- 15276648 TI - N-octyl-beta-valienamine up-regulates activity of F213I mutant beta-glucosidase in cultured cells: a potential chemical chaperone therapy for Gaucher disease. AB - Gaucher disease (GD) is the most common form of sphingolipidosis and is caused by a defect of beta-glucosidase (beta-Glu). A carbohydrate mimic N-octyl-beta valienamine (NOV) is an inhibitor of beta-Glu. When applied to cultured GD fibroblasts with F213I beta-Glu mutation, NOV increased the protein level of the mutant enzyme and up-regulated cellular enzyme activity. The maximum effect of NOV was observed in F213I homozygous cells in which NOV treatment at 30 microM for 4 days caused a approximately 6-fold increase in the enzyme activity, up to approximately 80% of the activity in control cells. NOV was not effective in cells with other beta-Glu mutations, N370S, L444P, 84CG and RecNciI. Immunofluorescence and cell fractionation showed localization of the F213I mutant enzyme in the lysosomes of NOV-treated cells. Consistent with this, NOV restored clearance of 14C-labeled glucosylceramide in F213I homozygous cells. F213I mutant beta-Glu rapidly lost its activity at neutral pH in vitro and this pH-dependent loss of activity was attenuated by NOV. These results suggest that NOV works as a chemical chaperone to accelerate transport and maturation of F213I mutant beta Glu and may suggest a therapeutic value of this compound for GD. PMID- 15276649 TI - Inactivation and intracellular retention of the human I183N mutated melanocortin 3 receptor associated with obesity. AB - Melanocortins are known to be involved in the regulation of feeding behavior. These hormones mediate their effects through G protein-coupled receptors (GPCRs) by stimulating adenylate cyclase. The melanocortin 3 receptor (MC3R) in the melanocortin receptor (MCR) family has been identified as a neural receptor subtype mainly expressed in the brain in mammals. Until now, only one heterozygous mutation (I183N) has been identified in the coding region of this receptor in two obese patients of the same family. In this study, we reported the functional characterization of the I183N mutated MC3R compared with that of the wild-type MC3R after transfection in HEK293 cells. Our results showed that the I183N mutation totally abolished the activity of the mutated receptor to generate intracellular cAMP. Furthermore, confocal microscopy observation revealed that the mutation induced an intracellular retention of the mutated receptor. Moreover, we demonstrated for the first time by co-transfection studies that the mutated receptor could reduce the wild-type receptor activity through a dominant negative effect. PMID- 15276650 TI - Mouse liver PMP70 and ALDP: homomeric interactions prevail in vivo. AB - ALDP, ALDPR, PMP70 and PMP70R are half ATP-binding cassette (ABC) transporters of the mammalian peroxisomal membrane. By analogy with other members of this family, it is assumed that peroxisomal ABC transporters must dimerize to become functional units. However, not much is known regarding the type of dimers (i.e., homodimers and/or heterodimers) that are formed in vivo under normal expression conditions. In this work, we have characterized the quaternary structure of mouse liver PMP70 and ALDP. The PMP70 protein complex was purified to apparent homogeneity using a two-step purification protocol. The ALDP-containing protein complex was characterized by preparative immunoprecipitation experiments. In both cases, no evidence for the existence of heteromeric interactions or for the presence of accessory proteins in these ABC transporter protein complexes could be obtained. Our data indicate that the majority (if not all) of mouse liver PMP70 and ALDP are homomeric proteins. PMID- 15276651 TI - Apolipoprotein E isoprotein-specific interactions with tissue plasminogen activator. AB - Apolipoprotein E (Apo E) is an important genetic risk factor for multiple neurological, vascular and cardiovascular diseases. Previously, we reported Apo E isoprotein-specific modulation of tissue plasminogen activator (tPA) using an in vitro blood-clotting assay. Here, we studied the conformational changes of Apo E2, E3 and E4 in the presence of tPA and vice versa using circular dichroism (CD) and dual polarization interferometry (DPI). We report isoprotein and state specific intermolecular interactions between the Apo E isoforms and tPA. Apo E2 interaction with immobilized tPA leads to significant conformational changes which are not observed with Apo E3 or E4. Additionally, tPA induces changes in helicity of lipidated Apo E2 whereas no detectable changes were observed in Apo E3 or E4. The Tukey's test for interaction indicated a significant (P < 0.001) interaction between tPA and Apo E2 in the lipidated environment. These results may be important regarding the mechanism by which Apo E has isoprotein-specific effects on many biological processes and diseases involving blood clotting, proteolysis and perfusion. PMID- 15276652 TI - Drastic reduction in the luminal Ca2+ -binding proteins calsequestrin and sarcalumenin in dystrophin-deficient cardiac muscle. AB - Luminal Ca2+ -binding proteins play a central role in mediating between Ca2+ uptake and Ca2+ -release during the excitation-contraction-relaxation cycle in muscle fibres. In the most commonly inherited neuromuscular disorder, Duchenne muscular dystrophy (DMD), the reduced expression of key Ca2+ -binding proteins causes abnormal Ca2+ -buffering in the sarcoplasmic reticulum (SR) of skeletal muscle. The heart is also affected in dystrophinopathies, as manifested by the pathological replacement of cardiac fibres by connective and fatty tissue. We therefore investigated whether similar changes occur in the abundance of luminal Ca2+ -regulatory elements in dystrophin-deficient cardiac fibres. Two-dimensional immunoblotting of total cardiac extracts was employed to unequivocally determine potential changes in the expression levels of SR components. Interestingly, the expression of the histidine-rich Ca2+ -binding protein was increased in the dystrophic heart. In contrast, the major Ca2+ -reservoir protein of the terminal cisternae, calsequestrin (CSQ), and the Ca2+ -shuttle and ion-binding protein of the longitudinal tubules, sarcalumenin, were drastically reduced in cardiac mdx fibres. This result agrees with the recently reported decrease in the Ca2+ release channel and Ca2+ -ATPase in the mdx heart. Abnormal Ca2+ -handling appears to play a major role in the molecular pathogenesis of the cardiac involvement in X-linked muscular dystrophy. PMID- 15276653 TI - Cisplatin up-regulates the in vivo biosynthesis and degradation of renal polyamines and c-Myc expression. AB - Time-dependent changes in polyamine metabolism and c-Myc expression are reported in kidney of mice treated with cisplatin, a widely used anticancer drug. We show that cisplatin significantly induces the expression of two enzymes critical to proper homeostasis of cellular polyamines, ornithine decarboxylase (ODC) and spermidine/spermine N1-acetyltransferase (SSAT). We also document the cross-talk between signalling pathway(s) induced by cisplatin injury to renal tubules and the testosterone/androgen receptor pathway. Their interaction results in a decrease in testosterone-induced ODC activity and ODC mRNA level, and in differential modulation of SSAT expression. Moreover, cisplatin and antifolate CB 3717, another nephrotoxic drug examined, severalfold up-regulate expression of c Myc mRNA, albeit with different kinetics. However, cisplatin, contrary to CB 3717, does not induce renal hepatocyte growth factor (HGF)/c-Met expression being without effect on HGF mRNA level and significantly down-regulating c-Met transmembrane receptor message. In conclusion, these in vivo studies document significant cisplatin-induced modulation of polyamine biosynthesis/degradation and up-regulation of c-Myc expression, and suggest that c-Myc transcription factor is involved in the induction of ODC in kidney injured with antifolate, but not with cisplatin. PMID- 15276654 TI - Statin blocks Rho/Rho-kinase signalling and disrupts the actin cytoskeleton: relationship to enhancement of LPS-mediated nitric oxide synthesis in vascular smooth muscle cells. AB - We previously demonstrated statins to enhance cytokine-mediated nitric oxide (NO) synthesis in vascular smooth muscle cells (VSMC). To clarify the mechanism by which this occurs, we evaluated the effects of fluvastatin in lipopolysaccharide (LPS)-stimulated VSMC. NO production induced by LPS was dose-dependently enhanced by fluvastatin, as were iNOS mRNA levels and iNOS protein expression. Exogenous mevalonate and geranylgeranylpyrophosphate (GGPP) dampened the stimulatory effect of fluvastatin. A pull-down assay demonstrated fluvastatin to decrease levels of GTP-bound Rho A. Moreover, a Rho-kinase inhibitor, Y-27632, was observed to enhance LPS-induced NO production. We recently demonstrated that disrupting F actin formation dramatically potentiates the ability of LPS to induce iNOS mRNA and protein expression. In the present study, staining of F-actin with nitrobenzoxadiazole (NBD)-phallacidin demonstrated that fluvastatin significantly impairs F-actin stress fiber formation. In light of these results, the ability of statins to increase NO production is due, at least in part, to their ability to block the biosynthesis of mevalonate, thereby preventing isoprenoid biosynthesis. This inhibits Rho/Rho-kinase signalling and, in turn, disrupts the actin cytoskeleton. Further analysis of the signalling pathway by which the actin cytoskeleton affects iNOS expression might yield new insight into mechanisms of regulation of NO production. PMID- 15276655 TI - Rapid tranquilization: new approaches in the emergency treatment of behavioral disturbances. AB - Psychiatric emergencies are often accompanied by behavioral disturbances that interfere with normal assessment and call for immediate intervention. Different pharmacological treatment regimens have been used for this purpose. Most of these regimens are based upon common clinical practice and have limited evidence base. Recently, a major publication by experts in the field of emergency psychiatry has covered this topic and the therapeutic armamentarium has been extended with the atypical antipsychotics. However, research is still hampered by different methodological limitations: unclear definition of the agitated state and therapeutic goal, idiosyncratic measurement, small sample sizes. The perspective of the patient and the interaction between the emergency care setting and treatment regimen also need further attention. All these important, but often neglected issues are covered in a selective review of the literature. PMID- 15276656 TI - Identification of somatic and anxiety symptoms which contribute to the detection of depression in primary health care. AB - Somatic symptoms and anxiety symptoms are often disregarded in the detection of depression in primary care. The present investigation examined to what extent somatic and anxiety symptoms recorded with the Composite International Diagnostic Interview-Primary Health Care Version (CIDI-PHC) can improve the detection of depression as compared to the General Health Questionnaire-12-item version alone. Data from the World Health Organization study on Psychological Problems in General Health Care were used. The study sample consisted of primary care attenders from 15 centres from all over the world who underwent a psychiatric examination with the CIDI-PHC. Medically unexplained somatic symptoms (back pain, feelings of heaviness/lightness in parts of the body, periods of bodily weakness, seizures/convulsions, permanent tiredness, exhaustion after a minimum of effort) and-to a smaller extent-diverse anxiety symptoms (e.g. feelings of anxiousness/nervousness, feelings of tension, difficulties relaxing) significantly contributed to the detection of depression in a logistic regression analysis. The results confirm the observation that in primary care somatic symptoms play an important role in the manifestation of depressive disorders. The items investigated herein could prove beneficial for future depression screening instruments to improve the detection of depressive disorders in primary care. PMID- 15276657 TI - One-year course of the first vs. multiple episodes of depression--Polish naturalistic study. AB - PURPOSE: The aim of the study was to compare a clinical course and treatment results of depression occurring as the first depressive episode, the second depressive episode or the third or further depressive episode. The study was 1 year, prospective, naturalistic observation made by Polish psychiatrists. METHODS: One-hundred and seventy-nine patients with the first depressive episode (group I), 170 patients with the second episode (group II) and 183 patients with the third or further episode of depression (group III) were compared. The main analysed variable was remission, defined as the score of < or =7 points on 17 item Hamilton depression rating scale (HDRS), after 6 and 12 months of observation. RESULTS: The groups of patients studied did not initially differ as to age, proportion of gender and intensity of depression. The percentages of remission after 6 months of observation in groups I-III were: 49%, 41% and 32%, and after 12 months 69%, 60% and 50%, respectively. CONCLUSIONS: The results obtained indicate that the course of subsequent depressive episodes is less favourable compared to the first depressive episode. The percentages of remission obtained in individual groups studied may have implications regarding duration of pharmacological treatment of depressive episode. PMID- 15276658 TI - Prevalence of depression as measured by the CBDI in a predominantly adolescent school population in Turkey. AB - OBJECTIVES: This cross-sectional randomised study involving a predominantly adolescent school population (ranging from the 6th to the 11th grades) was conducted to determine the prevalence and clinical characteristics of depression in adolescents in the city of Mersin, Turkey. SUBJECTS AND METHODS: A stratified sample of 4256 students was selected as representative of the city's adolescent school population. The students were divided into two groups according to diagnosis (Group I: depressive adolescents; Group II: adolescents without depression). The age range varied between 10 and 20 years, with a mean of 14.53 years (S.D. = 1.89), i.e. a mean of 14.73 years (S.D. = 1.79) for Group I, and 15.5 years (S.D. = 1.9) for Group II. Data was obtained via two structured questionnaires designed to determine the presence and clinical characteristics of depression both in adolescents and in their parents. In addition, the Child Beck Depression Inventory (CBDI) was administered to all students participating in the study. After quality control of data, the study sample was reduced to 4143 adolescents. The mean age of the students was 11.23 +/- 6.44 years, and the ratio of boys/girls was 1.19:1. RESULTS: The prevalence of depression according to the CBDI (cut-off point: 19) was found to be 12.55% in this study group, with a significantly higher prevalence of depression in girls than in boys. Binary regression analysis demonstrated that the most important factors involved in the onset of depression in adolescents were having problems with parents, staying down a grade, and humiliation at school, and that the most common symptoms were feelings of worthlessness/guilt, sadness, emptiness, irritability and somatic disorders. CONCLUSION: This cross-sectional prospective randomised school-based study has examined sociodemographic and clinical characteristics of adolescents with depression in a student population. It was found that in this study group there was a relatively high level of depressive symptoms, with a clear predominance of females over males. Other clinical characteristics of adolescents with depression have been discussed in the context of previous investigations. PMID- 15276659 TI - Social adjustment in generalised anxiety disorder: a long-term placebo-controlled study of venlafaxine extended release. AB - The objective of this analysis was to evaluate the short- (8 weeks) and long-term (24 weeks) efficacy of three fixed doses of venlafaxine extended release (ER) and placebo on the social adjustment of patients with generalised anxiety disorder (GAD). We analysed data from 544 outpatients who participated in a 24-week, double-blind, multicentre, parallel-group, placebo-controlled study conducted at 55 centres in five countries. All patients meet the DSM-IV criteria for GAD and were randomly assigned to receive venlafaxine ER 37.5, 75, and 150 mg or matched placebo administered orally once daily. Social adjustment was measured using the Social Adjustment Scale-Self Report, which explores social adaptation in the areas of work, social and leisure, extended family, primary relationship (marital), parental, and family unit. At baseline, the GAD patients had a high level of social dysfunction. Venlafaxine ER showed a dose-related improvement in social impairment during short-term treatment and in sustaining this improvement over the long-term. In the most severely socially impaired subgroup, placebo remission rates on the HAM-A were low, and the magnitude of the venlafaxine placebo difference on the mean HAM-A total score was high, reaching more than 7 points. The benefits of venlafaxine ER treatment of GAD extend beyond that of improvement of anxiety symptoms to a significant improvement in the impairment of functioning that is associated with the illness. PMID- 15276660 TI - Predictors of work disabilities in patients with panic disorder with agoraphobia. AB - OBJECTIVE: The aim of this study was to ascertain predictors of work insufficiency in patients with panic disorder (PD) with agoraphobia (AG). METHOD: Linear regression was used to identify predictors of work insufficiency in a sample of 72 consecutive outpatients with PD with AG. Intensity of work insufficiency was ascertained from modified National Institute of Mental Health Panic Questionnaire (NIMH PQ). That represented dependent variable. Independent variables were demographic data, duration of illness, presence of comorbid current major depression episode, presence of any personality disorder and scores on the Panic and Agoraphobia Scale (PAS) subscales: panic attacks, AG (avoidance behavior), anticipatory anxiety and worries about health. RESULTS: Patients reported severe work insufficiency. The best predict variable for the work insufficiency in patients with PD with AG was high score on the PAS dimension of AG. CONCLUSION: Patients generally reported severe effects of PD with AG on work efficacy and the results suggested that the impaired work efficacy was the most associated with avoidance behavior. These results recommend that the treatment of PD with AG patients should be related to decreasing avoidance behavior in order to establish adequate work performance in patients. PMID- 15276661 TI - Time-on-task effect in trait anhedonia. AB - The capacity to sustain attention was explored in a sample of anhedonic subjects according to the Chapman physical anhedonia scale. Sustained attention was determined by studying task-induced changes over the duration of the Eriksen response competition task [Percept. Psychophys. 16 (1974) 143]. Anhedonic subjects had longer reaction times (RTs), but missed no more targets than control subjects. Anhedonic subject RTs got longer with time-on-task (TOT) and displayed greater intra-subject variability. These results confirm those of a previous study indicating that anhedonic subjects may have developed a more conservative response strategy [Psychophysiology 37 (2000) 711] and suggest that this strategy may result in a more rapid decrease in energetical resources. Moreover, the greater intra-subject variability demonstrates the importance of assessing performance over time and its relationship to the variability of responses in the cognitive performance of anhedonic subjects. PMID- 15276662 TI - Obsessive-compulsive disorder and body dysmorphic disorder: a comparison of clinical features. AB - Body dysmorphic disorder (BDD) is currently classified as a somatoform disorder in DSM-IV, but has been long noted to have some important similarities with obsessive-compulsive disorder (OCD). In addition, BDD and OCD have been often reported to be comorbid with each other. In the present study, we compared demographic characteristics, clinical features and psychiatric comorbidity in patients with OCD, BDD or comorbid BDD-OCD (34 subjects with BDD, 79 with OCD and 24 with BDD-OCD). We also compared the pattern of body dysmorphic concerns and associated behaviors in BDD patients with or without OCD comorbidity. In our sample, BDD and OCD groups showed similar sex ratio. Both groups with BDD and BDD OCD were significantly younger, and experienced the onset of their disorder at a significantly younger age than subjects with OCD. The two BDD groups were also less likely to be married, and more likely to be unemployed and to have achieved lower level degree, than OCD subjects even when controlling for age. The three groups were significantly different in the presence of comorbid bulimia, alcohol related and substance-use disorders, BDD-OCD patients showing the highest rate and OCD the lowest. BDD-OCD reported more comorbid bipolar II disorder and social phobia than in the other two groups, while generalized anxiety disorder was observed more frequently in OCD patients. Patients with BDD and BDD-OCD were similar as regards the presence of repetitive BDD-related behaviors, such as mirror-checking or camouflaging. Both groups also did show a similar pattern of distribution as regards the localization of the supposed physical defects in specific areas of the body. The only significant difference concerned the localization in the face, that was more frequent in the BDD group. Our results do not contradict the proposed possible conceptualization of BDD as an OCD spectrum disorder. However, BDD does not appear to be a simple clinical variant of OCD and it seems to be also related to social phobia, mood, eating and impulse control disorders. The co-presence of BDD and OCD features appears to possibly individuate a particularly severe form of the syndrome, with a greater load of psychopathology and functional impairment and a more frequent occurrence of other comorbid mental disorders. PMID- 15276663 TI - Comorbid psychiatric diagnoses in kleptomania and pathological gambling: a preliminary comparison study. AB - Kleptomania and pathological gambling (PG) are currently classified in the DSM IV as impulse control disorders. Impulse control disorders are characterized by an overwhelming temptation to perform an act that is harmful to the person or others. The patient usually feels a sense of tension before committing the act and then experiences pleasure or relief while in the process of performing the act. Kleptomania and PG are often associated with other comorbid psychiatric diagnoses. Forty-four pathological gamblers and 19 kleptomanics were included in this study. All enrolled patients underwent a complete diagnostic psychiatric evaluation and were examined for symptoms of depression and anxiety using the Hamilton depression rating scale and the Hamilton anxiety rating scale, respectively. In addition, the patients completed self-report questionnaires about their demographic status and addictive behavior. The comorbid lifetime diagnoses found at a high prevalence among our kleptomanic patients included 47% with affective disorders (9/19) and 37% with anxiety disorders (7/19). The comorbid lifetime diagnoses found at a high prevalence in our sample of pathological gamblers included 27% with affective disorders (12/44), 21% with alcohol abuse (9/44), and 7% with a history of substance abuse (3/44). A larger study is needed to confirm these preliminary results. PMID- 15276664 TI - Attention deficit hyperactivity disorder (ADHD) in a 19th century children's book. AB - Attention deficit hyperactivity disorder (ADHD) is a major mental disorder in children. Presently, its pathogenesis and treatment as well as its role in adult psychiatry are subjects of heated debate. As early as 1846, the typical symptoms of ADHD were described by Heinrich Hoffmann, a physician who later founded the first mental hospital in Frankfurt. Interestingly, his description was published in a children's book entitled "Struwwelpeter" which he had designed for his 3 year-old son Carl Philipp. The symptomatology is impressively depicted in the colourfully illustrated story of "Zappel-Philipp" ("Fidgety Philip"), probably the first written mention of ADHD by a medical professional. This clearly shows that the diagnosis of ADHD is not an "invention" of modern times. PMID- 15276665 TI - Dimensions of mania: differences between mixed and pure episodes. AB - OBJECTIVE: The presence of at least five dimensions in mania has recently been established. This study extends previous findings by comparing the dimensions of pure vs. mixed mania. MATERIALS AND METHOD: One hundred and three inpatients with bipolar I disorder, manic or mixed (DSM IV), were assessed with SCID-I, YMRS and HDRS-21. The five-factor solution found after applying factorial analysis with Varimax rotation was compared between manic and mixed patients. RESULTS: There were differences between pure mania and mixed states on factor 1 (depression) and factor 3 (hedonism). There was a tendency to present higher values on factor 5 (activation) in the pure manic group. No differences were found in factor 2 (dysphoria) and factor 4 (psychosis). DISCUSSION: Hedonism and activation dimensions are present to a lesser degree in mixed states. Although the principal difference between mixed and pure bipolar disorder is the existence of depressive symptoms, the depressive dimension is strongly present in patients with pure mania. CONCLUSIONS: There is need to search for core depressive symptoms in all patients suffering from mania and to evaluate their outcome in clinical trials. PMID- 15276666 TI - "Bread madness" revisited: screening for specific celiac antibodies among schizophrenia patients. AB - PURPOSE: A possible association between gluten consumption and schizophrenia has been reported. The objective was to compare patients with chronic schizophrenia and matched controls for sociodemographic variables, prevalence of celiac specific anti-endomysial antibodies and disease-related variables. SUBJECTS AND METHODS: The study group was comprised of 50 consecutive patients diagnosed with schizophrenia, 18 years of age and older attending the out-patient clinic of the Mental Health Center in Beer-Sheva, Israel. The control group was comprised of mentally normal volunteers who came to primary care clinics for blood tests unrelated to gastrointestinal tract complaints and who were not diagnosed with celiac disease. Known celiac patients and those who refused to participate, did non-speak Hebrew or were incoherent were excluded from the study. All participants in both groups underwent a blood test for anti-endomysial IgA antibody and completed a questionnaire. RESULTS: Each group was comprised of 50 participants. There were no significant differences between the groups in gender, BMI or country of birth. The mean age of the study group was significantly higher than the controls. All tests for anti-endomysial antibody in both groups were negative. DISCUSSION AND CONCLUSIONS: In contrast to previous reports, we found no evidence for celiac disease in patients with chronic schizophrenia as manifested by the presence of serum IgA anti-endomysial antibodies. It is unlikely that there is an association between gluten sensitivity and schizophrenia. PMID- 15276667 TI - Prevalence of induced mania in patients treated with milnacipran: a comparison with paroxetine. PMID- 15276668 TI - Rapid improvement of severe anorexia nervosa during treatment with ethyl eicosapentaenoate and micronutrients. PMID- 15276669 TI - Remission of polydipsia as antipsychotic effect of clozapine. PMID- 15276670 TI - Carbamazepine may trigger new-onset epileptic seizures in an individual with autism spectrum disorders: a case report. AB - We herein report a case of new-onset epileptic seizures induced by carbamazepine in an individual with autism spectrum disorders (ASD). We clinicians should bear in mind the possibility that epileptic seizures may possibly be either precipitated or exacerbated by carbamazepine especially in individuals with ASD. PMID- 15276671 TI - Response of the auditory nerve to sinusoidal electrical stimulation: effects of high-rate pulse trains. AB - Electrical stimulation of the auditory nerve produces highly synchronized responses. As a consequence, electrical stimulation may result in a narrow dynamic range of hearing and poor temporal representation of an input signal. The electrically evoked compound action potential (ECAP) is an electrophysiologic response used for neural assessment in individuals with auditory prostheses. Because the ECAP arises from the activity of a population of auditory nerve fibers, within- and across-fiber synchrony should be evident in the responses. Due to its clinical relevance and reflection of neural response properties, the ECAP is used in the present study to examine changes in neural synchrony. Empirical and modeled single-fiber data indicate that stimulation with electrical pulses of a sufficiently high rate may induce stochastic neural response behaviors. This study investigated the effects of adding high-rate conditioning pulses (5000 pps) on the ECAP in response to 100 Hz electrical sinusoids. The results showed that high-rate conditioning pulses increased response amplitudes at low sinusoidal levels and decreased the amplitudes at high sinusoidal levels, indicating a decrease in the slope of the ECAP growth functions to sinusoidal stimuli. The results are consistent with a hypothesis that high-rate conditioning pulses increase single-fiber relative spread (RS) in response to sinusoidal stimuli, and the effect is highly dependent on the level of the high-rate conditioning pulses. PMID- 15276672 TI - CNTFRalpha and CNTF expressions in the auditory brainstem: light and electron microscopy study. AB - CNTF receptor alpha (CNTFRalpha) is involved in the development, the maintenance and the regeneration of a variety of brain structures. However, its in vivo distribution has not been determined in the auditory system. CNTFRalpha expression was studied in developing and adult rat brainstem auditory nuclei using immunohistochemistry. At birth, the CNTFRalpha immunolabeling was clearly present in somata of the external nucleus of the inferior colliculus but was diffuse throughout brainstem auditory nuclei. The labeling was present in most brainstem auditory nuclei by post-natal day (PND) 6. The intensity of the staining subsequently increased to its highest level at PND21 and decreased to an adult-like appearance by the fourth post-natal week. In adult, CNTFRalpha labeling occurred in most neurons of the cochlear nucleus (CN), the lateral superior olive (LSO), the medial superior olive (MSO), and the medial nucleus of the trapezoid body (MNTB). CNTFRalpha labeling first appeared in the central nucleus of the inferior colliculus (IC) by the end of the fourth week. There was a general increase in the expression of CNTFRalpha that begins prior to the onset of hearing and reaches its highest level after this important developmental stage. Ultrastructural analysis in the adult ventral CN revealed the presence of CNTFR in post-synaptic sites. The presence of CNTF has been investigated in the adult using both Western blot and immunohistochemistry. Western blot showed the presence of CNTF in both peripheral and central auditory structures. The CNTF label was generally localized to the somatic compartment, in axons and as puncta surrounding neuronal cell bodies and dendrites. Differential CNTF labeling was observed between the different auditory nuclei. CNTF staining is present in neurons of the CN, the MNTB and the LSO, while it is restricted to axons and puncta surrounding neuronal somata in the IC. The clear presence of CNTFRalpha at post-synaptic terminals and that of its ligand the CNTF in axons and puncta surrounding neuronal cell bodies suggest an anterograde mode of action for CNTF in the central auditory system. PMID- 15276673 TI - Cochlear microphonic changes after noise exposure and gentamicin administration during sleep and waking. AB - These experiments were designed to investigate the effect of noise, sleep, and gentamicin on the cochlear microphonic (CM) of the guinea pigs. Are the changes observed due to intrinsic cochlear phenomena or to efferent system actions? To answer this question, noise exposure together with efferent system blockade by gentamicin administration was performed. In the normal (non-treated) animal, noise exposure decreased both variability and amplitude of the tone evoked CM in about the first 10 min while the physiological modulation of slow wave sleep increasing the CM is not present. Following administration of gentamicin, noise no longer affect the CM in about the first 10 min, although it produces amplitude and variability increments. The influence of slow wave sleep on the CM is not altered. Thus, gentamicin does not block the CM sleep/wakefulness related shifts. The data were discussed in terms of the influence of gentamicin on the olivo cochlear bundle. It was hypothesized that the effects of noise on the CM is a result of both peripheral and central influences. PMID- 15276674 TI - Pitch is determined by naturally occurring periodic sounds. AB - The phenomenology of pitch has been difficult to rationalize and remains the subject of much debate. Here we test the hypothesis that audition generates pitch percepts by relating inherently ambiguous sound stimuli to their probable sources in the human auditory environment. A database of speech sounds, the principal source of periodic sound energy for human listeners, was compiled and the dominant periodicity of each speech sound determined. A set of synthetic test stimuli were used to assess whether the major pitch phenomena described in the literature could be explained by the probabilistic relationship between the stimuli and their probable sources (i.e., speech sounds). The phenomena tested included the perception of the missing fundamental, the pitch-shift of the residue, spectral dominance and the perception of pitch strength. In each case, the conditional probability distribution of speech sound periodicities accurately predicted the pitches normally heard in response to the test stimuli. We conclude from these findings that pitch entails an auditory process that relates inevitably ambiguous sound stimuli to their probable natural sources. PMID- 15276675 TI - Age-dependent changes in the lateral superior olive of the gerbil (Meriones unguiculatus). AB - Data from humans and animal models provide evidence for an age-dependent impairment in the ability to localize sound. The lateral superior olive (LSO) in the ascending auditory pathway is one important center involved in processing of binaural auditory stimuli. To identify potential age-dependent changes we characterized the LSO in young (< 15 months) and old (> or =3 years) gerbils with a special emphasis on the expression of GABA- and glycine-like immuno-reactivity. The dimensions of the LSO, as well as the number and density of glycine- and GABA immuno-reactive neurons, were not significantly different between young and old gerbils. The size of glycine- and GABA-immuno-reactive neurons was significantly reduced in the high-frequency (medial) limb of the LSO. Over all, age-dependent changes in the LSO of the gerbil were small. PMID- 15276676 TI - Hearing in workers exposed to low-dose radiation for a long period. AB - The aim of the present study was to evaluate changes in hearing thresholds with standard and high frequency audiometry in workers exposed to low-dose ionizing radiation for a long period. A total of 57 (49 male and 8 female) technical staff working in radiology-related jobs who were exposed to occupational radiation were included in the study. The control group consisted of 32 (27 male and 5 female) volunteer subjects with normal hearing. The symptoms like tinnitus, vertigo, weakness and lack of appetite were evaluated. A standard ascending/descending method was applied to the subjects of the study and the control groups in order to determine their hearing thresholds at eleven different frequencies between 250 and 16,000 Hz. In the study group, the working duration of subjects ranged from 4 to 23 years, and the percentage of tinnitus, weakness, vertigo and lack of appetite were 47%, 28%, 24% and 17%, respectively. It was observed that pure tone hearing thresholds were markedly increased for 4,000, 6,000, 8,000, 14,000 and 16,000 Hz frequencies in the study group compared to the control group (p<0.01). Levels of static compliance and middle ear pressures of the study group were similar to the control group. Tinnitus, vertigo and hearing loss in high frequencies were observed in the subjects exposed to the radiation for a long period. Subjects under high risk should be evaluated periodically. We suggest that the use of standard and high frequency audiometry together could be beneficial in the evaluation of these subjects. PMID- 15276677 TI - Molecular mechanisms underlying ectopic otoconia-like particles in the endolymphatic sac of embryonic mice. AB - Otoconin-90, the principal otoconial matrix protein, provided a tool to investigate the molecular mechanism of otoconial morphogenesis. The endolymphatic sac of the embryonic chick and guinea pig contain otoconia. Here, we show that the embryonic mouse transiently expresses ectopic otoconia in the endolymphatic sac. Massive precipitate of otoconin-90-positive material is detectable in the lumen of the endolymphatic sac between embryonic day 14.5 and 17.5 with frequent accretion into more heavily staining otoconia-like particles. Otoconin-90 was also localized at the surface and the interior of epithelial cells lining the endolymphatic sac as well as incorporated into free floating cells. In contrast, in situ hybridization failed to detect mRNA in the endolymphatic duct and sac, even though the adjacent nonsensory vestibular structures are heavily stained. Because of ample expression of otoconin-90 protein in the absence of the corresponding mRNA, we conclude that the luminal otoconin-90 is imported via longitudinal flow from the vestibular compartments, where both mRNA and protein are strongly expressed. Because of absence of mRNA, the expression of the corresponding protein by the epithelia lining the endolymphatic sac can only be explained by a resorptive process, as previously proposed on the basis of the movement of luminal macromolecules. The data do not support the previous hypothesis that the transient expression of otoconia-like particles of the endolymphatic sac represents a vestigial phenomenon from the amphibian stage, since amphibia express ample mRNA encoding otoconin-22 in the endolymphatic sac system. PMID- 15276678 TI - PET imaging of the 40 Hz auditory steady state response. AB - The auditory steady state response (aSSR) is an oscillatory electrical potential recorded from the scalp induced by amplitude-modulated (AM) or click/tone burst stimuli. Its clinical utility has been limited by uncertainty regarding the specific areas of the brain involved in its generation. To identify the generators of the aSSR, 15O-water PET imaging was used to locate the regions of the brain activated by a steady 1 kHz pure tone, the same tone amplitude modulated (AM) at 40 Hz and the specific regions of the brain responsive to the AM component of the stimulus relative to the continuous tone. The continuous tone produced four clusters of activation. The boundaries of these activated clusters extended to include regions in left primary auditory cortex, right non-primary auditory cortex, left thalamus, and left cingulate. The AM tone produced three clusters of activation. The boundaries of these activated clusters extended to include primary auditory cortex bilaterally, left medial geniculate and right middle frontal gyrus. Two regions were specifically responsive to the AM component of the stimulus. These activated clusters extended to include the right anterior cingulate near frontal cortex and right auditory cortex. We conclude that cortical sites, including areas outside primary auditory cortex, are involved in generating the aSSR. There was an unexpected difference between morning and afternoon session scans that may reflect a pre- versus post-prandial state. These results support the hypothesis that a distributed resonating circuit mediates the generation of the aSSR. PMID- 15276679 TI - Lack of association between Connexin 31 (GJB3) alterations and sensorineural deafness in Austria. AB - Mutations in the gap junction protein beta 3 (GJB3) gene encoding Connexin 31 (Cx31) are known to cause autosomal inherited sensorineural deafness, erythrokeratodermia and neuropathy. The role of Cx31 mutations has not been described in familial cases of non-syndromic hearing impairment (NSHI) in central European populations. To identify mutations in the Austrian population, highly selected familial (n=24) and sporadic (n=21) cases of isolated NSHI were screened by analysis of the complete coding sequence of Cx31, after exclusion of a common Cx26 causing deafness. Three different variations occurring in a total of 37% of all cases were identified. A C94T (R32W) missense mutation was seen in 4.4% of cases and two silent alterations C357T and C798T were detected in 8.9% and 24.4% of cases exclusively in a heterozygous pattern. No correlation between Cx31 alterations and deafness was found. To investigate the role of heterozygous Cx31 variations for a possibly combination allelic disease inheritance with Cx26 mutations as shown for Connexin 30 and Connexin 26, patients with Cx26 variations were tested. Our data suggest that Cx31 alterations are common but have no or a low genetic relevance in the Austrian hearing impaired population with or without Cx26 alterations. PMID- 15276680 TI - Evaluating cochlear function and the effects of noise exposure in the B6.CAST+Ahl mouse with distortion product otoacoustic emissions. AB - Cochlear function and susceptibility to noise over-exposure were examined in the congenic mouse strain B6.CAST+Ahl (B6.CAST) and compared to these same features in the CAST/Ei (CAST) and C57BL/6J (C57) parental strains. For both types of comparisons, the primary measure was the distortion-product otoacoustic emissions (DPOAE) at 2f1-2f2. Our assumption was that the B6.CAST mouse was corrected for the early onset age-related hearing loss (AHL) exhibited by one of its parental strains (C57) by the age-resistant properties of its other parental strain (CAST), and thus would exhibit neither AHL nor susceptibility to noise overstimulation effects. With respect to cochlear function, for 2.5-month mice, there was a tendency for DPOAEs to be slightly lower for mid-frequency primary tones for both C57 and B6.CAST mice, while the former mice showed clear AHL effects at the highest test frequency. However, by 5 months of age, the B6.CAST mice, like the CAST mice, displayed robust DPOAE levels that were significantly larger than DPOAE levels for the C57 mice, which were essentially absent for frequencies above about 30 kHz. To investigate the role of the Ahl gene in the susceptibility of the cochlea to the effects of noise over-exposure, two distinct paradigms consisting of temporary (TTS: 1-min, 105-dB SPL, 10-kHz pure tone) and permanent (PTS: 1-h, 105-dB SPL, 10-kHz octave band noise) threshold-shift protocols were used. The brief TTS exposure produced reversible reductions in DPOAEs that for both the B6.CAST and CAST mice recovered to within a few dB of their baseline levels by 3 min post-exposure. In contrast, the C57 mice recovered somewhat slower and, by 5 min post-exposure, emission levels were still 5 dB or more below their corresponding pre-exposure values. At 3 months of age, the TTS mice along with another group of naive subjects representing the same three mouse strains were exposed to the PTS paradigm. By 4 days post-exposure, for B6.CAST and CAST mice, DPOAE levels had recovered to their pre-exposure control levels. However, DPOAEs for the C57 mice at most of the measurable frequencies were at least 10-30 dB lower than their counterpart baseline levels. Together these data suggest that the Ahl allele in the C57 strain contributes to both the early onset AHL exhibited by these mice as well as their susceptibility to both TTS and PTS over-exposures. PMID- 15276681 TI - The response of the apical turn of cochlea modeled with a tuned amplifier with negative feedback. AB - In an earlier study [Hear. Res. 149 (2000) 55] velocity amplitudes of the outer Hensen's cell (HC) and basilar membrane (BM) were measured before, and at different times, after, sacrificing the animal. The velocity amplitude changed in a way that was characteristic of a negative feedback amplifier. A simple negative feedback amplifier model was proposed to explain the magnitude of the HC and BM velocity changes at CF. In the experiment tuning changed as well, both at the HC and BM. The model has now been extended to include tuning changes. The model response is compared with the experimental observations. The model is able to account quantitatively for the following experimental observations: (i) At the HC the tuning broadens and velocity decreases slowly after sacrifice. (ii) At the BM tuning sharpens and velocity increases at a faster rate. (iii) The velocity increase at BM is much larger than the decrease at HC. PMID- 15276682 TI - Distortion product otoacoustic emissions show exceptional resistance to noise exposure in MOLF/Ei mice. AB - Baseline distortion-product otoacoustic emissions (DPOAEs) at several primary tone levels were compared between naive 2- to 3-month old inbred CBA/CaJ (CBA) and wild-derived MOLF/Ei (MOLF) mice. Only minor DPOAE differences were noted between the two strains and these differences were not systematic across frequency or test levels. These emission findings were consistent with earlier results on auditory brainstem response thresholds reported by others [Zheng et al., Hear. Res. 130 (1999) 94-107] thus suggesting that both CBA and MOLF strains have normal hearing. Subsequent episodes of over-exposure to a 105-dB SPL, octave band noise centered at 10 kHz for 8 h revealed that MOLF DPOAEs were exceptionally resistant to the adverse aftereffects of excessive noise exposure as compared to CBA mice. Unlike the noise-exposure resistant inbred 129/SvEvTac strain, which has reduced baseline DPOAE levels especially at high frequencies, MOLF mice have normal DPOAEs making the interpretation of noise-exposure effects more straightforward. PMID- 15276683 TI - Factors affecting psychophysical tuning curves for normally hearing subjects. AB - These experiments were conducted to clarify the influence of beats and combination products on psychophysical tuning curves (PTCs) for normally hearing subjects. PTCs for 1- and 4-kHz sinusoidal signals were determined using as maskers a sinusoidal tone and 80-, 160-, and 320-Hz wide bands of noise. PTCs obtained using the sinusoidal masker showed distinct irregularities, particularly for masker frequencies close to the signal frequency. The PTCs determined for the noise maskers were more regular. The broader the masker, the more regular were the shapes of the PTCs. To reduce the detectability of beats produced by the interaction of the signal and masker, a pair of low-frequency tones, called "Modulation detection interference (MDI) tones", was used to introduce beats at the same rate. The MDI tones reduced the threshold level of the sinusoidal masker by up to 20 dB for frequencies within 300 Hz of the signal frequency; a similar but smaller effect was found when an 80-Hz wide masker was used. Adding a lowpass filtered (LF) noise to the sinusoidal or narrowband noise masker did not affect the low-frequency sides of the PTCs, suggesting no influence of combination products. The LF noise did affect the high-frequency sides of the PTCs, but this can be attributed to it reducing off-frequency listening. To achieve a PTC whose shape around the tip is minimally affected by beats, we propose using a noise masker with a bandwidth approximately equal to the bandwidth of the auditory filter for which the PTC is measured. PMID- 15276684 TI - Isoflurane increases amplitude and incidence of evoked and spontaneous otoacoustic emissions. AB - The volatile anesthetic isoflurane was tested for its effect on cochlear function by means of measuring distortion product otoacoustic emissions (DPOAE) and spontaneous otoacoustic emissions (SOAE) in the mustached bat (Pteronotus parnellii parnellii). Averaged growth functions of DPOAE and spontaneous otoacoustic emissions were assessed and compared between the control group (no isoflurane application) and the isoflurane group (application of isoflurane at vaporizer settings sof about 1.5-2%). Isoflurane significantly increases the DPOAE amplitude, e.g. at a primary tone level l2 of 40 dB SPL by 10.7 dB. Additionally, the incidence of SOAEs was highly increased during application of isoflurane. The sound-evoked efferent effect on the generation of otoacoustic emissions was significantly reduced in the isoflurane group. We suggest that isoflurane might affect the postsynaptic action of acetylcholine (ACh) released by the efferent terminals of outer hair cells (OHCs). This could lead to the observed decrease of efferent suppression and to a disinhibition of cochlear amplification. PMID- 15276685 TI - Clinical response to antidepressant treatment and 3-methoxy-4-hydroxyphenylglycol levels: mini review. AB - Plasma 3-methoxy-4-hydroxyphenylglycol (MHPG) may provide valuable information regarding central noradrenergic activity. In this article, we mainly reviewed about the associations between plasma MHPG levels and responses to antidepressant treatment. There exists heterogeneity of depression with regards to plasma levels of MHPG; in other words, depressed patients might be dichotomized into one group characterized by anxiety and/or perceptions of powerlessness with high plasma MHPG levels and another group characterized by psychomotor retardation with low plasma MHPG levels. In addition, it is possible that patients with lower pretreatment MHPG levels might respond to drugs that affect both noradrenergic neurons and serotonergic neurons or predominantly noradrenergic neurons. On the other hand, patients with higher pretreatment MHPG levels might respond to drugs that affect predominantly serotonergic neurons or GABAergic neurons. It is possible to predict the responses to antidepressant drugs by means of plasma MHPG levels in depressed patients. PMID- 15276686 TI - Changes in craving for a cigarette and arterial nicotine plasma concentrations in abstinent smokers. AB - Although the relationship between nicotine and changes in heart rate and blood pressure has been demonstrated, the relationship between nicotine and subjective effects such as decreased craving, relaxation, sickness, and decreased nervousness, is less well delineated. In this study, arterial nicotine levels were drawn in 21 smokers who smoked two average nicotine (AN) cigarettes and one low nicotine (LN) cigarette. Craving for a cigarette, relaxation, sickness, and decreased nervousness were rated on a visual analog scale (VAS) before and after smoking each cigarette. None of these subjective measures except craving for a cigarette was changed significantly by smoking. The change in craving was significantly correlated with the area under the plasma nicotine concentration versus time curve (r = -0.57, p = 0.01) calculated from the arterial nicotine samples drawn up to 20 min after the initiation of smoking the first AN cigarette. Although well-documented behavioral manipulations, such as smoking denicotinized cigarettes, reduce craving, increases in plasma arterial nicotine concentrations after smoking the first cigarette of the day also reduce craving. Both the psychology and pharmacology of nicotine/tobacco smoking are involved in craving reduction. PMID- 15276687 TI - Growth hormone releasing hormone reverses endotoxin-induced localized inflammatory hyperalgesia without reducing the upregulated cytokines, nerve growth factor and gelatinase activity. AB - During inflammatory processes, the hypothalamic-pituitary axis is activated which can subsequently result in analgesia. For example, hypothalamic corticotrophin releasing hormone (CRH) that is released during such activation has been attributed with analgesic actions. It is believed that the somatotrophic axis is also activated during inflammation. The aim of this study was to determine the analgesic actions of growth hormone-releasing hormone (GHRH), in a rat model of localized inflammatory hyperalgesia, induced by intraplantar (i.pl.) endotoxin (ET) injections. Pretreatment with intraperitoneal (i.p.) injections of GHRH (2, 5, 10 microg kg(-1)) 30 min before i.pl. ET injection (1.25 microg in 50 microl saline) prevented, in a dose-dependent manner, both mechanical hyperalgesia determined by the paw pressure (PP) test and thermal hyperalgesia determined by the hot plate (HP) and paw immersion (PI) tests. Pretreatment with GHRH had no significant effect on the elevated levels of the inflammatory mediators, interleukin (IL)-1beta, tumor necrosis factor (TNF)-alpha, IL-6 and nerve growth factor (NGF) due to i.pl. ET injection. No significant effect was obtained by pretreatment with GHRH, on the increased expression of gelatinase B due to ET injection. In conclusion, GHRH reverses inflammatory hyperalgesia in the rat without affecting the upregulated inflammatory mediators and these actions may be clinically important. PMID- 15276688 TI - Extracellular serotonin, dopamine and glutamate levels are elevated in the hypothalamus in a serotonin syndrome animal model induced by tranylcypromine and fluoxetine. AB - Serotonin (5-HT) syndrome is a potentially fatal condition associated with various combinations of serotonergic drugs. The present study was undertaken to demonstrate that nervous systems other than the 5-HT system also participate in the pathophysiology of 5-HT syndrome. Concentrations of 5-HT, dopamine (DA) and glutamate in the hypothalamus were measured in two different 5-HT syndrome animal models using a microdialysis technique. The first model was induced by tranylcypromine, a nonselective monoamine oxidase (MAO) inhibitor (3.5 mg/kg) and fluoxetine, a selective serotonin reuptake inhibitor (SSRI) (10 mg/kg). The second model was induced by clorgyline, an MAO-A inhibitor (1.2 mg/kg) and 5 hydroxy-L-tryptophan, a precursor of 5-HT (5-HTP) (80 mg/kg). In the first model, the levels of 5-HT and DA increased by 40-fold and 44-fold, respectively, compared with the preadministration levels. In the second model, the concentrations of 5-HT increased by up to 140-fold, whereas DA levels increased by only 10-fold, of the preadministration levels. Although the level of glutamate in the second model barely changed, a delayed increase in the glutamate level was observed in the first model. These findings suggest that not only hyperactivity of the 5-HT system, but also hyperactivity of the DA system, are present in 5-HT syndrome, and that the glutamatergic system is influenced in some 5-HT syndrome cases in which the DA concentration markedly increases. PMID- 15276689 TI - Isobolographic analysis of interaction between cyclooxygenase inhibitors and tramadol in acetic acid-induced writhing in mice. AB - Non-steroidal anti-inflammatory drugs (NSAIDs) and opioids are the most commonly used analgesics in the management of acute and chronic pain. Combined use of NSAIDs and opioids has been indicated for achieving better analgesia with reduced side effects. The present study was aimed at evaluating the combination of different NSAIDs, which inhibit cyclooxygenase (COX) enzymes and tramadol against acetic acid-induced writhing in mice. The expected beneficial effect of combination regimen was analyzed by isobolographic analysis. The oral and intrathecally administered tramadol, a mu-opioid and naproxen, a nonselective COX inhibitor produced dose-dependent antinociception, however, rofecoxib, a selective COX-2 inhibitor lacked analgesic efficacy in writhing test. Isobolographic analysis showed synergistic or supra-additive interactions for the combinations of naproxen and tramadol after oral and intrathecal administration. However, similar interaction was not observed when tramadol was combined with rofecoxib. Pretreatment with naloxone partially reversed the antinociceptive effect of tramadol per se and its combination with naproxen without modifying the per se effect of NSAID. The results demonstrated marked synergistic interaction between naproxen and tramadol and such interaction involved opioid as well as non opioid mechanisms of tramadol and inhibition of COX-1 but not COX-2 by naproxen. PMID- 15276690 TI - Parahippocampal gyrus in first episode psychotic disorders: a structural magnetic resonance imaging study. AB - Neuropathological abnormalities in schizophrenia have been demonstrated in the parahippocampal gyrus (PHG). Only a few studies on first-episode neuroleptic naive schizophrenia patients have been done using in vivo neuroimaging techniques. The authors examined the PHG morphology using structural MRI in neuroleptic-naive subjects with first episode psychoses. Volumetric measurements of PHG and intracranial volume (ICV) were obtained on subjects with schizophrenia and schizoaffective disorders (SCZ; n = 33), nonschizophrenia psychotic disorders (NSCZ; n = 11) and matched healthy subjects (HS; n = 43). The subjects were rated on the Brief Psychiatric Rating Scale (BPRS). Group differences and clinical correlations of ICV-adjusted PHG volumes were examined. Left PHG was significantly different across the groups consisting of SCZ, NSCZ and HS. PHG was larger in NSCZ compared to SCZ but not relative to HS. Bilaterally, PHG was no different between SCZ and HS. In pooled psychotic patients, the PHG volume negatively correlated with total positive symptom, delusion and conceptual disorganization scores on BPRS. Patients with delusions had relatively smaller PHG compared to nondelusional subjects. Observed differences in PHG volume in first-episode neuroleptic-naive patients suggest that these observations are not confounded by illness chronicity or medication effects. Significant association of PHG volume with psychotic symptoms suggests that PHG pathology plays an important role in the etiopathology of psychosis and its symptoms. PMID- 15276691 TI - Effect of switching to atypical antipsychotics on memory in patients with chronic schizophrenia. AB - While the usefulness of atypical antipsychotics for improving cognitive function has been proven, the specific effects of these drugs are still unknown. The objective of this study was to investigate changes of the immediate memory and verbal working memory in patients with chronic schizophrenia after switching to one of four atypical antipsychotic agents and cessation of anticholinergic therapy. The subjects included 77 schizophrenic patients who were treated primarily with typical antipsychotics. Treatment was randomly switched to one of four atypical antipsychotics (olanzapine, perospirone, quetiapine, or risperidone) over a 4-week period, and then the drug was continued for another 4 weeks while the patient was taken off anticholinergics. The immediate memory, verbal working memory, and symptoms were evaluated. Significant improvement of immediate memory was only seen with olanzapine and risperidone. Improvement was also seen after switching to perospirone, but immediate memory worsened after treatment with this anticholinergic drug was discontinued. Deterioration was seen after switching to quetiapine, but immediate memory improved and reached the previous level after treatment with anticholinergic drugs was discontinued. Significant improvement of the verbal working memory was only seen during risperidone administration. The findings suggested that switching chronic schizophrenic patients to atypical antipsychotics can improve both the immediate memory and the verbal working memory when risperidone is used, while improvement of immediate memory can be expected with olanzapine. From the viewpoint of improving the memory, quetiapine should not be administered concomitantly with anticholinergic drugs, and caution should be exercised when discontinuing anticholinergic drugs during treatment with perospirone. PMID- 15276692 TI - Effects of the hyperbaric oxygen treatment on the Na+,K+ -ATPase and superoxide dismutase activities in the optic nerves of global cerebral ischemia-exposed rats. AB - The effects of hyperbaric oxygen (HBO) treatment on the Na+,K+ -ATPase and superoxide dismutase (SOD) activities were examined in the optic nerves of the rats exposed to global cerebral ischemia. Animals were exposed to global cerebral ischemia of 20-min duration and were either sacrificed or exposed to the first HBO treatment immediately, 0.5, 1, 2, 6, 24, 48, 72 or 168 h after ischemic procedure (for Na+,K+ -ATPase activities measurement) or 2, 24, 48 or 168 h after ischemia (for SOD activities measurement). HBO procedure was repeated for 7 consecutive days. It was found that global cerebral ischemia induced a statistically significant decrease in the Na+,K+ -ATPase activity of the optic nerves, starting from 0.5 to 168 h of reperfusion. Maximal enzymatic inhibition was registered 24 h after the ischemic damage. The decline in the Na+,K+ -ATPase activity was prevented in the animals exposed to HBO treatment within the first 6 h of reperfusion. The results of the presented experiments demonstrated also a statistically significant increase in the SOD activity after 24, 48 and 168 h of reperfusion in the optic nerves of non-HBO-treated ischemic animals as well as in the ischemic animals treated with HBO. Our results indicate that global cerebral ischemia induced a significant alterations in the Na+,K+ -ATPase and SOD activities in the optic nerves during different periods of reperfusion. HBO treatment, started within the first 6 h of reperfusion, prevented ischemia induced changes in the Na+,K+ -ATPase activity, while the level of the SOD activity in the ischemic animals was not changed after HBO administration. PMID- 15276694 TI - Sex differences on spontaneous alternation in prepubertal rats: implications for an animal model of obsessive-compulsive disorder. AB - Sex differences in the prevalence of obsessive-compulsive disorder (OCD) in prepubertal children have been described. Deficits on spontaneous alternation behavior (SAB) have been proposed as an animal model of OCD. OBJECTIVES: To explore possible sex differences in the ontogeny of SAB and in the effect of the 5-HT1A agonist, 8-OH-hydroxy-2 (di-n-propylamino)-tetralin (8-OH-DPAT) in an animal model of OCD. METHODS: The ontogeny of SAB and the perseveration produced by 8-OH-DPAT were compared between male and female prepubertal rats. RESULTS: Males alternated their arm choose from postnatal day 32 onwards, while females perseverated in the chosen arm until postnatal day 38. The mean number of repetitive choices remained close to 1 in males from postnatal day 23 onwards, but females showed a mean number of repetitive choices higher than 1.5 until the end of the test. The 8-OH-DPAT (0.125, 0.5 and 2.0 mg/kg, 15 min) produced perseveration in males but not in females. CONCLUSIONS: These data show important sex differences in the ontogeny of SAB and the effect of 8-OH-DPAT in a model of OCD. Such differences could be relevant for the sex differences in the prevalence of childhood OCD. PMID- 15276693 TI - The dopaminergic stabiliser ACR16 counteracts the behavioural primitivization induced by the NMDA receptor antagonist MK-801 in mice: implications for cognition. AB - The Carlsson research group has developed a series of compounds capable of stabilising the dopamine system without inducing the deleterious hypodopaminergia that encumbers the currently used antipsychotic drugs. In the present study one of these dopaminergic stabilisers, ACR16, was tested in a mouse model for cognitive deficits of schizophrenia and autism. Since we believe that hypoglutamatergia is a key element in both schizophrenia and autism we used mice rendered hypoglutamatergic by treatment with the N-methyl-D-aspartate (NMDA) antagonist MK-801. MK-801 causes both hyperactivity and a behavioural primitivization. ACR16 attenuated the MK-801-induced hyperactivity and, in addition, caused a marked improvement of behavioural quality with a movement pattern approaching that of control animals. Since we believe that the impoverishment of the behavioural repertoire caused by MK-801 may correspond to the cognitive deficits seen in schizophrenia and autism, these results suggest that ACR16 may improve cognitive status in these disorders. PMID- 15276695 TI - Hypothalamic superoxide dismutase, xanthine oxidase, nitric oxide, and malondialdehyde in rats fed with fish omega-3 fatty acids. AB - Phospholipids located in the cellular membrane play a critical role in the fluid mosaic model of membrane structure and membrane function. Evidence is mounting for the role of abnormal phospholipid metabolism in some neuropsychiatric disorders including schizophrenia. As an important essential fatty acid (EFA), omega-3 (omega-3) fatty acid series are found in large amounts in fish oil. The aim of this experimental study was to assess the changes of some of the oxidant and antioxidant parameters in the hypothalamus of rats fed with omega-3 EFA diet (0.4 g/kg/day) for 30 days. Eight control rats and nine rats fed with omega-3 were decapitated under ether anesthesia, and hypothalamus was removed immediately. Malondialdehyde (MDA) and nitric oxide (NO) levels as well as superoxide dismutase (SOD) and xanthine oxidase (XO) enzyme activities in the hypothalamus were measured. SOD activity was significantly decreased in omega-3 EFA treated group compared to control group (p < 0.014). Tissue MDA and NO levels were also decreased in omega-3 EFA treated group compared to control rats (p < 0.0001). Xanthine oxidase activity was found to be increased in omega-3 EFA treated rats when compared to the control group (p < 0.0001). Taken together, this preliminary animal study provides strong support for a therapeutic effect of omega-3 EFA in some neuropsychiatric disorders in which reactive oxygen species (ROS) are recently accused to be an important physiopathogenetic factor. PMID- 15276696 TI - Stress-induced hyperthermia in the mouse: c-fos expression, corticosterone and temperature changes. AB - In mammals, stress exposure is frequently associated with an elevated body temperature ['emotional fever', stress-induced hyperthermia (SIH)]. Rectal measurement of body core temperature of the mouse induces a rise of 1-1.5 degrees C over a 10- to 15-min time interval. This phenomenon has been exploited to design a specific test for measuring stress-induced hyperthermia: the singly housed SIH paradigm in mice. In the present experiments, changes in body temperature and corticosterone levels were studied 10, 30, 60, 90 and 120 min after the first insertion of the rectal probe. In addition, changes in patterns of neural activation, as observed after immunostaining for Fos-immunoreactivity (Fos-IR), were studied in the brains of animals perfused at times 0, 60 or 120 min. Our results show that SIH and corticosterone levels have their peak values between 10 and 30 min and are no longer different from control values after 60 min. Patterns of Fos-IR have been studied in 11 brain areas, of which 2 brain areas (anterodorsal preoptic and periolivary nuclei) showed a continuing rise in Fos-IR after 60 and 120 min, while six nuclei, mostly hypothalamic and septal, showed a peak induction of Fos-IR after 60 min. In three brain areas, no consistent changes in Fos-IR could be observed. The authors conclude that the changes observed in the patterns of Fos-IR, after application of the singly housed SIH-test in mice, reflect the effects of both the stressor application and the ensuing thermoregulatory responses. The role of each activated brain area in either one of these effects is discussed in view of data available from the literature. PMID- 15276697 TI - Low serum levels of brain-derived neurotrophic factor in patients with schizophrenia do not elevate after antipsychotic treatment. AB - Brain-derived neurotrophic factor (BDNF) has been suggested to be involved in the etiology of schizophrenia. There is a line of evidence that disruption of neurotrophins could play a role in the etiology of schizophrenia, and antipsychotics show their effect by altering levels of neurotrophins. The aim of this study was to evaluate the effect of antipsychotics on serum BDNF levels and their relationship with the symptoms in patients with schizophrenia. Twenty-two schizophrenia patients were enrolled in the study. The control group consisted of 22 age- and sex-matched physically and mentally healthy volunteers (7 male, 15 female). Serum BDNF levels and the positive and negative syndrome scale (PANSS) scores were recorded at baseline and after 6 weeks of treatment. Serum BDNF levels were also recorded in the control group. Schizophrenia patients who failed to meet 30% improvement in PANSS score were excluded from the study. The baseline serum BDNF levels of schizophrenia patients were lower than those of controls (t = 4.56; df = 21; p < 0.001). There was no correlation between serum BDNF levels and PANSS scores in patients with schizophrenia (p > 0.05). Although PANSS (for positive symptoms p < 0.001, for negative symptoms p < 0.001) and general psychopathology (t = 20.9; df = 22; p < 0.001) scores improved significantly after 6 weeks of antipsychotic treatment; there was no change in BDNF levels in patients' serum (p > 0.05). Our results support the view that BDNF would be associated with schizophrenia. However, we could not conclude that treatment with antipsychotics alters serum BDNF levels in patients with schizophrenia. PMID- 15276698 TI - Neuronal expression of cyclooxygenase-2, a pro-inflammatory protein, in the hippocampus of patients with schizophrenia. AB - Several types of evidence suggesting that the inflammatory response system is associated with pathophysiology of schizophrenia have been accumulated. Recently, a prospective double-blind study demonstrated that supplementary treatment with celecoxib, a selective cyclooxygenase-2 (COX-2) inhibitor, produced significantly greater improvement in scores on the Positive and Negative Syndrome Scale (PANSS) and on all subscales during the acute phase in patients with schizophrenia compared with risperidone alone therapy. The therapeutic effect of celecoxib on the psychopathology of schizophrenia is speculated to be based on COX activity inhibition; however, the detailed pharmacological mechanisms are unclear. To clarify whether or not COX-2 expression is altered in schizophrenia, we examined neuronal COX-2 expression in the hippocampus from cases of schizophrenia (n = 17), normal controls (n = 22), and cases of Alzheimer's disease (AD) as a positive control (n = 17). Quantitative immunohistochemical analysis demonstrated that neuronal COX-2 expression was significantly up-regulated in each CA1-4 region in Alzheimer's disease compared with controls, and that the mean COX-2 immunointensity in CA1-4 was significantly correlated with Abeta load in cases of Alzheimer's disease. In contrast, COX-2 expression was not up-regulated in any subdivision of the hippocampus in the schizophrenia group. These results suggest that celecoxib may affect the pathophysiology of schizophrenia through COX-2 independent actions rather than by inhibiting activity of up-regulated COX-2 protein. PMID- 15276699 TI - Mechanism of action of A-85380 in an animal model of depression. AB - A role for neuronal nicotinic receptor (NNR) activation in animal models of depression has been established. In order to determine the mechanism by which NNR ligands exert their antidepressant effects, experiments using different NNR receptor antagonists in both the mouse and the rat forced swim test (RFST) were performed. In the mouse forced swim test (MFST), A-85380 (0.62 micromol/kg = 0.14 mg/kg, i.p.), an NNR agonist, increased swim distance when administered 15 min prior to test. This effect was blocked by pre-treatment with mecamylamine (1.5 micromol/kg = 0.3 mg/kg, i.p.), suggesting that an NNR mechanism is involved. Further, chlorisondamine at a non-central nervous system (CNS) penetrating dose (1.6 micromol/kg = 1 mg/kg, i.p.) did not antagonize A-85380 in this model, thus implicating central rather than peripheral nicotinic receptors. Dihydro-beta erythroidine (DHbetaE, 0.3 micromol/kg = 0.1 mg/kg, i.p.) pre-treatment also blocked this effect, indicating that the alpha4beta2 receptor subtype may be involved in A-85380-induced antidepressant effects. Finally, methiothepin (0.33 micromol/kg = 0.14 mg/kg, i.p.) pre-treatment antagonized this effect, suggesting serotonergic involvement. In the rat modified forced swim test, sub-acute administration of A-85380 (0.62 micromol/kg, i.p.) increased swimming behavior and decreased immobility. Climbing behavior was unaffected. In contrast, desipramine treatment (33 micromol/kg = 10 mg/kg, i.p.) resulted in an increase in climbing behavior with no effect on swimming. This behavioral profile has been shown to be more typical of serotonergic rather than noradrenergic antidepressants, suggesting that A-85380 exerts its effects via NNR activation of serotonergic systems. PMID- 15276700 TI - Phosphatidylserine: an antidepressive or a cognitive enhancer? AB - The aim of the present study was to evaluate the putative antidepressive and cognitive enhancer effects of phosphatidylserine (BC-PS). The antidepressive effect of BC-PS (50, 100 or 200 mg/kg), compared to saline or imipramine (IMI; 25 mg/kg), was studied in the forced swimming test in rats. These drugs were administered 1 and 8 h after training and 1 h before the test. BC-PS (50, 100 and 200 mg/kg)-treated rats exhibited a significant decrease in immobility time (IT) in the test session (performed 24 h after training) when compared to control rats. Moreover, the IMI-treated group showed a significant reduction in IT in comparison to control rats. The cognitive enhancer effect of BC-PS (50, 100 and 200 mg/kg) was studied in the three versions of the water maze task: spatial working memory version, spatial reference memory version, and cued version. There was no significant difference between the BC-PS-treated groups and control animals in these memory tasks. Taken together, the present results are suggestive of an antidepressive effect of BC-PS in the forced swimming test in rats but not of a cognitive enhancer effect of the drug in the water maze test. PMID- 15276701 TI - BanI polymorphism of the cytosolic phospholipase A2 gene may confer susceptibility to the development of schizophrenia. AB - Membrane phospholipid abnormalities have been proposed to be involved in the pathogenesis of schizophrenia. Cytosolic phospholipase A2 (cPLA2) plays a major role in the metabolism of fatty acids but is also found in abnormalities in patients with schizophrenia. This study examined the association between the cPLA2 gene BanI polymorphism and schizophrenia. Ninety-seven Korean schizophrenia patients and 117 healthy controls participated in this study. Genotyping was performed by using polymerase chain reaction (PCR)-based methods. Genotype and allele distributions were significantly different between the schizophrenia patients and controls. In particular, the A2 allele was associated with an increased risk of schizophrenia (p = 0.003; odds ratio (OR) = 1.799; confidence interval (CI) = 1.192-2.716). However, the polymorphism was not different when the patient group was subdivided by the presence or absence of family history and by positive and negative subgroups according to the positive and negative syndrome scale (PANSS) score on schizophrenia. The results of this study replicated those of previous findings from Western countries and indicates the need for further studies on the potential role of the cPLA2 gene polymorphism in the susceptibility to schizophrenia. PMID- 15276702 TI - Antipsychotic treatment of psychosis associated with multiple sclerosis. AB - This case report deals with the antipsychotic treatment in multiple sclerosis (MS). Psychiatric symptoms are a frequent event in patients with MS. However, there are only few systematic studies of antipsychotic treatment in MS patients. Most of the studies are related to clozapine due to the lack of neurological, particularly extrapyramidal side effects (EPS). Therefore, experiences with other atypical drugs are requested. This paper discusses the pros and cons of different atypical drugs using the example of one patient, who showed adverse effects after treatment with quetiapine and olanzapine. Risperidone was not administered with respect to possible EPS. However, ziprasidone was tolerated well and appeared to be effective. PMID- 15276704 TI - Comments on "A new view of diabetic Retinopathy" by A.J. Barber. PMID- 15276705 TI - Deoxyribozymes: cleaving a path to clinical trials. AB - Deoxyribozymes (DNAzymes) comprise an exciting class of nucleic acid molecules that are capable of specific cleavage of target mRNA. Recent reports attest to the potential of this class of molecules in cell culture and preclinical studies, where DNAzymes exhibit the ability to silence disease-associated genes (mainly those associated with cardiovascular disease and cancer). Rigorous testing in preclinical studies is now required before this relatively new entity enters Phase I clinical trials. PMID- 15276706 TI - Are synaptic MAGUK proteins involved in chronic pain? AB - Chronic pain, particularly neuropathic pain, is notoriously difficult to treat. NMDA receptor antagonists are effective in reducing pain hypersensitivity in animal models and clinical settings but are associated with an unacceptable level of side-effects. Recent studies of the role of a family of synaptic membrane associated guanylate kinase proteins in chronic pain provide new insights into central mechanisms of chronic pain that could result in new biochemical targets for its treatment. PMID- 15276707 TI - The antibody-capture [(35)S]GTPgammaS scintillation proximity assay: a powerful emerging technique for analysis of GPCR pharmacology. PMID- 15276708 TI - How antipsychotics become anti-"psychotic"--from dopamine to salience to psychosis. AB - The relationship between dopamine, psychosis and antipsychotics has been challenged by the suggestion that there is a delay, of weeks, between the onset of dopamine receptor blockade and improvement in psychosis. However, recent data show that there is no significant delay. In light of these new findings, it is proposed that dopamine, through its role in reward prediction and motivational salience, provides a link to psychosis. Psychosis results from aberrant reward prediction and aberrant attribution of salience that is caused by disordered dopamine transmission. Antipsychotics become anti-"psychotic" by blocking dopamine transmission and attenuating the motivational salience of the symptoms, leading to the common statement from patients that symptoms "don't bother me as much anymore". This attenuation of salience also impacts on normal motivational drives, providing an explanation for why antipsychotics might induce iatrogenic negative symptoms and dysphoria, often leading to non-compliance by patients. The implications of this framework for relapse and other clinical phenomena, animal models and future studies are discussed. PMID- 15276709 TI - A common vaccine for fighting neurodegenerative disorders: recharging immunity for homeostasis. AB - Neurodegenerative conditions share common primary risk factors and mediators of disease progression. Because many degenerative disorders are age related, deteriorating immunity in aging patients might impose additional risk. Adaptive (T-cell-mediated) immunity is a defense mechanism that instructs microglia to fight off and clear away self-derived enemies. Such adaptive immunity can be boosted, without risking the development of autoimmune disease, by injecting weak agonists of self-antigens or by weakening the suppressive CD4+CD25+ regulatory T cells. If widely cross-reactive, the agonist might effectively counteract a variety of neurodegenerative disorders. Boosting of relevant T cells by vaccination could thus "recharge" a deteriorating immune system that has to contend with an increasing number of risk factors. PMID- 15276710 TI - Historical review: a brief history and personal retrospective of seven transmembrane receptors. AB - Pharmacologists have studied receptors for more than a century but a molecular understanding of their properties has emerged only during the past 30-35 years. In this article, I provide a personal retrospective of how developments and discoveries primarily during the 1970s and 1980s led to current concepts about the largest group of receptors, the superfamily of seven-transmembrane (7TM) receptors [also known as G-protein-coupled receptors (GPCRs)]. Significant technical advances such as the development of methods for radioligand binding, solubilization and purification of the beta(2)-adrenoceptor and other adrenoceptors led to the cloning of receptor genes and the discovery of their 7TM architecture and homology with rhodopsin. A universal mechanism of receptor regulation by G-protein-coupled receptor kinases (GRKs) and arrestins, originally discovered as a means of "desensitizing" G-protein-mediated second-messenger generation, was subsequently found to mediate both receptor endocytosis and activation of a growing list of signaling pathways such as those involving mitogen-activated protein kinases. Numerous opportunities for novel therapeutics should emerge from current and future research on 7TM receptor biology. PMID- 15276711 TI - Importance of P-glycoprotein at blood-tissue barriers. AB - P-glycoprotein is the product of the ABCB1 [also known as multidrug resistance 1 (MDR1)] gene. It translocates a broad variety of xenobiotics out of cells. P glycoprotein was first described in tumor cells that were resistant to various anticancer agents as a result of P-glycoprotein overexpression. P-glycoprotein is not only expressed in tumor cells but also in a broad variety of normal tissues with excretory function (small intestine, liver and kidney) and at blood-tissue barriers (blood-brain barrier, blood-testis barrier and placenta). In particular, following the generation of P-glycoprotein-deficient mice it became clear that this efflux transporter limits the absorption of orally administered drugs, promotes drug elimination into bile and urine, and protects various tissues (e.g. brain, testis and fetus) from potentially toxic xenobiotics. In humans, a considerable interindividual variability in P-glycoprotein tissue expression is observed, and current research is focused on the potential role of ABCB1 polymorphisms and haplotypes that affect P-glycoprotein tissue expression, plasma concentrations of drugs, the frequency of adverse drug reactions and treatment outcome. PMID- 15276712 TI - PDE4 inhibition: a novel approach for the treatment of inflammatory bowel disease. AB - Inflammation is a hallmark of inflammatory bowel disease (IBD), and elevation of cAMP levels can inhibit the pro-inflammatory and tissue-destructive properties of leukocytes. Phosphodiesterase 4 (PDE4) is the predominant enzyme that metabolizes cAMP in inflammatory cells, and the anti-inflammatory and immunomodulatory potential of PDE4 inhibitors in human leukocytes, endothelium and epithelium is well documented. Although PDE4 inhibitors have been investigated as treatments for several inflammatory diseases, this has focused mainly on asthma and chronic obstructive disease (COPD). Historically, their clinical utility has been limited by nausea and emesis. However, the PDE4 inhibitors cilomilast and roflumilast have recently shown efficacy in asthma and COPD, with a reduced propensity to cause nausea and emesis. In this review, we summarize for the first time the evidence that PDE4 inhibitors might have therapeutic benefit in IBD, and discuss mechanisms of action beyond the inhibition of inflammatory cells. PMID- 15276713 TI - CAR: detailing new models. AB - Functional analysis has broadened our understanding of the physiological roles of the two related nuclear receptors pregnane X receptor (PXR; NR1I2) and constitutive androstane receptor (CAR; NR1I3). Initial research focused on the role of these two receptors in xenobiotic detoxification and, more recently, additional functional roles for CAR have been identified. Specifically, CAR activity has been shown to ameliorate the effects of hyperbilirubinemia, caloric restriction and toxic bile acids. Thus, the physiological role of CAR has broadened to include responses to metabolic and nutritional stress. These data highlight potential new opportunities in targeting CAR for drug discovery. PMID- 15276714 TI - Techniques: the application of accelerator mass spectrometry to pharmacology and toxicology. AB - The exquisite sensitivity of accelerator mass spectrometry (AMS) is being used in biomedical applications to quantitate many isotopes, including 14C, 3H, 41Ca and 27Al, at attomole (10(-18)) concentrations. This enables compounds and metabolites to be measured in human urine and plasma after administration of low pharmacologically or toxicologically relevant doses of labelled chemicals and drugs. The detection of modified proteins or DNA in target organs after dosing with potential carcinogens has also been achieved in many studies. Advances aimed at increasing sample throughput and expanding applications by coupling AMS instruments directly to chromatographic separation systems are currently underway. PMID- 15276715 TI - Environmental impact of diuron transformation: a review. AB - Diuron is a biologically active pollutant present in soil, water and sediments. A synthesis of literature data on its physicochemical properties, partitioning behaviour, abiotic and biotic transformations, toxicological and ecotoxicological impacts has been here performed. Data have shown that diuron is generally persistent in soil, water and groundwater. It is also slightly toxic to mammals and birds as well as moderately toxic to aquatic invertebrates. However, its principal product of biodegradation, 3,4-dichloroaniline exhibits a higher toxicity and is also persistent in soil, water and groundwater. Thus, diuron indirectly possesses a significant amount of toxicity and could be a potential poisoning pesticide contaminant of groundwater. Unfortunately, groundwater contamination will still persist despite the progressive suppression of diuron (Directive 200/60/CE). Therefore, determining the main factors influencing its degradation and its ecotoxicological effects on the environment and health could provide a basis for further development of bioremediation processes. PMID- 15276716 TI - Analysis of heavy metal distribution in superficial estuarine sediments (estuary of Bilbao, Basque Country) by open-focused microwave-assisted extraction and ICP OES. AB - Open-focused microwave-assisted extraction and ICP-OES determination of As, Cd, Cr, Cu, Fe, Mn, Ni, Pb and Zn from surface sediments of the estuary of Bilbao (Basque Country, North of Spain) was carried out. All the samples were collected at three different tributaries of the estuary (Asua, Galindo and Nerbioi Ibaizabal) every two months during 1999. The digestion procedure was proposed from the conclusions of a fractionated factorial design, and the precision and accuracy of the method was verified using a certified reference sediment (RTC008 050). The results of the analysis were statistically treated by means of principal component analysis and correlation analysis. The principal component analysis of sediment data (32 samples x 9 metals) indicated different patterns of contamination regarding the tributary and sampling station. The two main patterns observed were a steady increment of the metal concentration along all the campaigns in the samples collected in the Galindo River and a seasonal variation in the Nerbioi-Ibaizabal River, with higher metallic content during summertime and lower content during wintertime. PMID- 15276717 TI - Biosorption of cadmium by Myriophyllum spicatum L. and Myriophyllum triphyllum orchard. AB - The aim of this study was to characterize the biological treatment of heavy metal contaminated water employing Myriophyllum species, namely M. spicatum L. and M. triphyllum. Both species were found to be capable of removing cadmium (Cd) from water; the latter significantly outperformed. Myriophyllum species were treated with 0, 2, 4, 6, 8, 16 mgl(-1) cadmium solutions for 24, 48, 72, 96 h, respectively. Cd uptake of both species was the lowest at 2 mgl(-1) and the highest at 16 mgl(-1). Concentration related cadmium stress on both species exhibit significant difference on pigment levels (8-16 mgl(-1)). These findings contribute to the fact that submerged aquatic plants can be used for the removal of heavy metals. PMID- 15276718 TI - Nitration and hydroxylation of benzene in the presence of nitrite/nitrous acid in aqueous solution. AB - This paper studies the nitration and hydroxylation of benzene in the presence of nitrite/nitrous acid in aqueous solution, both in the dark upon addition of hydrogen peroxide and under 360 nm irradiation. In both cases the detected transformation intermediates were phenol (P), nitrobenzene (NB), 2-nitrophenol (2NP) and 4-nitrophenol (4NP). P and NB directly form from benzene, and the initial formation rate of P is at least an order of magnitude higher than that of NB. In our experiments nitrophenols arise from P nitration, as can be inferred by their time evolution and isomer ratio (2NP:4NP = 60:40, 3NP below detection limit). Nitrophenols may also form upon hydroxylation of NB, but in a different ratio (2NP:3NP:4NP = 45:30:25). The detection of 3NP is thus a marker for the hydroxylation of NB, since this isomer is not formed in P nitration processes. The formation rates of P and NB increase with decreasing pH, both in the presence of HNO(2) + H(2)O(2) in the dark (which produce HOONO) and in the presence of NO(2)(-)/HNO(2) under irradiation. In the former case the pH dependence reflects the formation rate of HOONO. In the case of the irradiation experiments the pH effect can be accounted for by the higher molar absorbivity and photolysis quantum yield of nitrous acid when compared with nitrite. Interestingly, benzene does not react with HNO(2) alone in the dark. An important feature of benzene nitration in the presence of NO(2)(-)/HNO(2) under irradiation is that the process is not inhibited by the addition of hydroxyl scavengers, differently from the case of phenol nitration. This finding indicates that nitrite irradiation might lead to the nitration of certain aromatic compounds in natural waters even in the presence of natural hydroxyl scavenging agents, which are usually thought to limit the environmental role of many photochemical processes. PMID- 15276719 TI - Polycyclic aromatic hydrocarbons, black carbon, and molecular markers in soils of Switzerland. AB - Polycyclic aromatic hydrocarbons (PAH) were analysed in 23 soil samples (0-10 cm layer) from the Swiss soil monitoring network (NABO) together with total organic carbon (TOC) and black carbon (BC) concentration, as well as some PAH source diagnostic ratios and molecular markers. The concentrations of the sum of 16 EPA priority PAHs ranged from 50 to 619 microg/kg dw. Concentrations increased from arable, permanent and pasture grassland, forest, to urban soils and were 21-89% lower than median numbers reported in the literature for similar Swiss and European soils. NABO soils contained BC in concentrations from 0.4 to 1.8 mg/g dw, except for two sites with markedly higher levels. These numbers corresponded to 1-6% of TOC and were comparable to the limited published BC data in soil and sediments obtained with comparable analytical methods. The various PAH ratios and molecular markers pointed to a domination of pyrogenically formed PAHs in Swiss soils. In concert, the gathered data suggest the following major findings: (1) gas phase PAHs (naphthalene to fluorene) were long-range transported, cold condensated at higher altitudes, and approaching equilibrium with soil organic matter (OM); (2) (partially) particle-bound PAHs (phenanthrene to benzo[ghi]perylene) were mostly deposited regionally in urban areas, and not equilibrated with soil OM; (3) Diesel combustion appeared to be a major emission source of PAH and BC in urban areas; and (4) wood combustion might have contributed significantly to PAH burdens in some soils of remote/alpine (forest) sites. PMID- 15276720 TI - Photodegradation of new amino acid derivatives suitable for chelating agents in pulp bleaching applications. AB - The purpose of this study was to evaluate photodegradabilities of the following new low-nitrogen chelating agents: N-bis[(carboxymethoxy)ethyl]glycine (compound 1), N-bis[(1,2-dicarboxyethoxy)ethyl]glycine (compound 2) and N-bis[(1,2 dicarboxyethoxy)ethyl]aspartic acid (compound 3). At first photodegradation of these chelating agents as uncomplexed Na-compound 1-3 and Cu(II) complexes were tested, both in lake and distilled water, by exposing them to near-UV region radiation at the range of 315-400 nm. Uncomplexed Na-compounds 2 and 3 were selected to sunlight exposure experiments carried out in lake and distilled water. Compound 3 was also tested in sunlight as Cu and Ca complexes in both solutions. Photodegradation of Na(6)-compound 3 in distilled water was studied by exposing it to radiation at the wavelength of 253.7 nm. Photodegradation products were analysed by means of GC-MS (gas chromatography with mass selective detector). The results demonstrated that compound 1 was quite photostable even as Cu complex while compounds 2 and 3 were found to be photodegradable. Over 90% reduction of compound 3 was achieved during one week and 80% reduction of compound 2 in two weeks' time when they were added as Na salt to lake water and exposed to sunlight. Compound 3 as Cu complex degraded totally in the sunlight in less than one week. In the case of compound 3, the degradation rate decreased depending on the counter cation in the order Cu > Na > or = Ca. The study demonstrated that photodegradation of Na(6)-compound 3 does not result in total mineralization of the compound. A photodegradation pathway for Na(6)-compound 3 is proposed. PMID- 15276721 TI - Pollution survey of polycyclic aromatic hydrocarbons in surface water of Hangzhou, China. AB - The concentrations of 10 polycyclic aromatic hydrocarbons (PAHs) were simultaneously measured for five times (July and November 1999-2002) in four water bodies of Hangzhou, China. To investigate possible sources of PAH contamination, sediments, soils, runoff water and atmospheric particles of the region were also analyzed for their PAH contents. The maximum levels of PAHs in the water bodies (34.4-67.7 microg/l) were found in July, while significantly lower PAH concentrations (4.7-15.3 microg/l) were measured in November. The contamination is substantial and it may have resulted in acute toxic effects on aquatic organisms. The measured PAH concentrations in sediments and soils (224 4222 ng/g), runoff water (8.3 microg/l) and air particles (2.3 microg/m(3)) are discussed in relation to concentrations and patterns found in the surface water bodies. Comparison of PAH levels in sediments and soils led to the conclusion that the erosion of soil material does not contribute significantly to the contamination of sediments. The atmospheric PAH deposition to water bodies in the city area of Hangzhou was estimated to be 530 tons/a, while the contribution of surface runoff water was estimated to be 30.7 tons/a. The ratios of selected PAH were then used to illuminate the possible origin of PAHs in the examined samples (petrogenic, pyrogenic). PMID- 15276722 TI - Routine, automated determination of inorganic and total mercury in multimedia using cold vapour atomic absorption spectrometry. AB - A routine, automated analytical method for simultaneous determination of total and inorganic mercury by cold vapour atomic absorption spectrometry in multimedia is described. Excellent accuracy and precision results were obtained with human hair certified reference materials namely, BCR-397 and IAEA-086. The reproducibility relative standard deviation for total mercury was 4% and 22%, respectively. The limit of detection for total and inorganic mercury was 0.2 microg/g hair. The described method has been successfully applied in determination of total and inorganic mercury as well as organic mercury in human hair, urine and fish tissue samples. PMID- 15276723 TI - Complexation of arsenate with humic substance in water extract of compost. AB - The interactions of environmental toxicants with organic substances affect the speciation and dynamics, and subsequent toxicity, mobility, and fate of toxicants in the environment. For the purpose of understanding the complexation of As(V) with humic substances, arsenate-containing solutions with As concentrations from 1 to 8 mgl(-1) were prepared to react with the water extract of compost (WEC). All the reaction systems including the control were incubated for 48 h at 25 degrees C. The complexation of As(V) with humic substance was examined by dialysis and ion exchange techniques. From 30% to 51% of added As(V) reacted with organic substance in WEC to form an As-metal-organic complex. This was verified as a hydrophobic organic fraction after separation of As-metal-organic complex fraction from the hydrophilic fraction by XAD-8 resin. The complex substance was also identified as a humic substance by the method of proton binding formation function determination. This suggests that cations, such as Ca and Mg, and especially Fe, Al, and Mn act in cation bridging in the complexation of As(V) with humic substance. The role of metals in the complexation of As(V) with humic substance in terrestrial and especially aquatic environments thus merits close attention. PMID- 15276724 TI - Determination of 4-tert-octylphenol, 4-nonylphenol and bisphenol A in surface waters from the Haihe River in Tianjin by gas chromatography-mass spectrometry with selected ion monitoring. AB - The estrogenic pollutants 4-tert-octylphenol (OP), 4-nonylphenol (NP) and bisphenol A (BPA) were determined in surface water samples from the Haihe River, Tianjin, China. The analytes were extracted and concentrated from 300 ml acidified water samples by liquid-liquid extractions using dichloromethane, derivatized with trifluoroacetic anhydride, and quantified by gas chromatography mass spectrometry (GC-MS) with selected ion monitoring (SIM). Among the samples collected from 14 sampling sites, only one sample was found to have a relatively high concentration of BPA (8.30 microgl(-1)) and NP (0.55 microgl(-1)). The concentrations of OP, NP and BPA in the other samples were in the range of 18.0 20.2, 106-296 and 19.1-106 ngl(-1), respectively. Recoveries for OP, NP and BPA in the spiked water samples were all over 80%. PMID- 15276725 TI - Photolysis of spinosyns in seawater, stream water and various aqueous solutions. AB - Spinosad, a reduced-risk insecticide, contains primarily two active compounds, spinosyns A and D that are fermentation products of bacterium Saccharopolyspora spinosa. It is currently used to control fruit flies in Hawaii, USA. In this study, we investigated photodegradation of spinosyns A and D, respectively, in seawater, stream, tap and distilled-deionized waters under various light sources. Photodegradation of the two chemicals was also studied in various aqueous solutions prepared with phosphate buffer at different pH or chemical sensitizers. Two major photolytic products from spinosyn A were detected as spinosyn B and hydroxylated spinosyn A. Spinosyn D was similarly hydroxylated and N demethylated. Spinosyns A and D were photodegraded rapidly under sunlight in Hawaii, USA. The half-life of spinosyns A and D in stream water was 1.1 and 1.0 h, respectively, and was a half of that in distilled-deionized water, 2.2 and 2.0 h, respectively. Photodegradation of spinosyns A and D followed an order of increasing rate constants in distilled-deionized, seawater, stream and tap water under 300 nm artificial light, and was enhanced approximately 8- and 17-fold, respectively, in acetone-sensitized solution as compared to in distilled deionized water. Photolysis rates of spinosyns A and D in isopropanol- or humic acid-fortified water did not differ much as compared with those accordingly in distilled-deionized water. Spinosyns A and D photodegraded slower in acidic aqueous solution than in basic aqueous solution. PMID- 15276726 TI - Batch washing of cadmium from soil and sludge by a mixture of Na2S2O5 and Na2EDTA. AB - Washing of cadmium contaminated soil and sludge using a mixture of 0.1 M Na(2)S(2)O(5) and 0.01 M Na(2)EDTA was investigated in the batch mode. Initial Cd concentration in samples was 500 mg kg(-1). The sequential extraction was conducted to study of what form that Cd was removed. SPSS program version 9.01 was performed to determine what soil parameter had the greatest influence on the washing. The organic matter in soil was found to be the main factor for the washing. Soil with low organic matter would have high percentage of removing Cd. When adding more washing solution, the Cd removal efficiency was lower. The highest removal efficiency was between 67.83% and 97.3% when using a 1 g:2.5 ml soil to washing solution ratio. The predominant form of the removed Cd was exchangeable form. By contrast for the sludge, the highest Cd removal efficiency was 17.13% when using sludge in washing solution at the ratio of 1 g:7.5 ml. Most of washed Cd was in reducible form. PMID- 15276727 TI - Investigation on the influence of methanol on adsorption and leaching of pesticides with soil column liquid chromatography. AB - The effect of methanol of low concentration on adsorption and leaching of atrazine and tebuconazole was studied in this paper. The adsorption coefficients and the retardation factors (R(m)) of pesticides on EUROSOIL 3# log-linearly decreased as volumetric fraction of methanol (f(c)) was increased in the binary solvent mixtures of methanol and water. These data are consistent with solvophobic theory formerly outlined for describing the adsorption and transport of hydrophobic organic chemicals from mixed solvents. Nevertheless, the adsorption of these pesticides in soil-water system slightly increased when the soil was pre-washed with methanol in comparison with that pre-washed with water (pure water system). Furthermore, their adsorption coefficients were still higher in binary solvent systems with methanol of very low concentrations, i.e. f(c) < 0.03 for atrazine and f(c) < 0.01 for tebuconazole, than those in pure water system. The adsorption coefficients (log K(w)) of atrazine and tebuconazole predicted by solvophobic theory were 0.5792 and 1.6525, respectively, and their experimental log K(w) were 0.3701 and 1.6275 in pure water system. Obviously, the predicted log K(w) of the two pesticides was higher than the experimental log K(w) in pure water system. The predicted K(w) and the retardation factor (R(w)) in pure water system by solvophobic theory are thus possibly inaccurate. PMID- 15276728 TI - Pharmaceuticals in the aquatic environment--a comparison of risk assessment strategies. AB - The growing concern over the release of pharmaceutically active compounds and personal care products into the environment has prompted the introduction of risk assessment guidelines in both the European Union by the European Medicines Evaluation Agency (EMEA) and in the United States by the Food and Drug Administration (FDA), details of which are presented herein. Both employ a similar tiered system that compares the predicted environmental concentrations (PEC) with the worst-case no effect concentrations estimated from standard toxicity assays. These approaches are compared and contrasted. Results demonstrate room for improvement in areas such as the use of threshold values to trigger investigations, chronic and mechanism specific toxicity screening and mixture toxicity for which possible solutions are proposed. PMID- 15276729 TI - Study on the formation and transport of ozone in relation to the air quality management and vegetation protection in Tenerife (Canary Islands). AB - An experimental study on the formation and transport of ozone in ambient air was performed in Tenerife (Canary Islands) in order to investigate the processes affecting ozone levels and air quality. The special features of Tenerife (prevalence of the trade wind pattern (NE), orography and the specific location of the local ozone sources) permit to quantify the role of the 'long-range transport from northern latitudes' versus the 'formation and transport of ozone downwind of the main urban areas' of Tenerife. Levels of O(3), NO(2) and O(X) were monitored in different types of environments to achieve this purpose. The results showed that: (1) upwind of the urban areas ozone is mainly transported from the ocean by trade winds, (2) local ozone titration (by NO) and ozone replenishment from the ocean are the main causes of ozone variations in urban and suburban areas, and (3) photochemical ozone production occurs downwind of the urban areas. Photochemical production causes daylight O(3) and O(X) levels downwind of urban areas to be frequently (60% and 35% days/year, respectively) higher than upwind of the urban sites (O(3) and O(X) excess frequently in the range 5-20 ppbv). Due to the above processes, different daily ozone cycles occur in short distances (<30 km), with maximum O(3) levels during daylight or night depending on the site. Ozone phytotoxicity was assessed by calculating the AOT40 index upwind and downwind of the main urban areas. The critical value for the 5 day-AOT40 index was simultaneously exceeded at the two sites (few times/year) during long-range transport events. During the additional exceedances of the critical value downwind of the urban area, relatively high 5-day-AOT40 values were recorded upwind of the urban site. Thus, long-range transport from northern latitudes may produce relatively high 5-days-AOT40 levels in the oceanic boundary layer. These results are important for the protection of the large number of endemic plants in the Canaries. The conceptual model discussed in this study may be qualitatively applied to other islands which possess features similar to those of Tenerife. PMID- 15276730 TI - Modelling the dynamics of pentachlorophenol bioavailability in column experiments. AB - The aqueous-phase concentration of an organic pollutant found in a subsurface environment is often assumed to be its bioavailable concentration. However, the aqueous-phase concentration does not adequately reflect the dynamics of contaminant availability to microbes in flow-through systems. This paper assesses the effects of interacting processes such as sorption, biodegradation, and transport on contaminant bioavailability, and the fraction of the bioavailable contaminant that is taken up by microbes. The evolution of the bioavailable and uptake fractions is studied in two ways. Firstly, column experiments are conducted in which the introduced contaminant (pentachlorophenol, PCP) can flow through the columns, be consumed by microorganisms, or be sorbed by a solid matrix. Secondly, a phenomenological model (Flow/Sink/Reservoir model) that illustrates the dynamic nature of bioavailability and quantifies the uptake fraction is developed, based on a flow balance. Results show that after 60 h of sorption-limited bioavailability, the microorganisms induce desorption, so that the sorbed pool becomes bioavailable and bioavailability is limited by the PCP injection rate. A conclusion is drawn that the aqueous-phase concentration is a poor indicator of contaminant bioavailability to microbes. PMID- 15276731 TI - Routine monitoring of antibiotics in water and wastewater with a radioimmunoassay technique. AB - Antibiotics are one of a group of pharmaceutical compounds that have been found in lakes and streams throughout the world and the occurrence of these compounds in the environment has raised concerns regarding the toxicity to aquatic organisms and the emergence of strains of antibiotic-resistant bacteria. The objective of this study was to assess the use of a relatively simple coupled solid-phase extraction (SPE)/radioimmunoassay (RIA) method for screening and/or monitoring tetracycline (TC) and sulfonamide (SA) compounds in water. Cross reactivity of TCs and SAs was used to determine the specificity of the assays. The results indicate that TC, oxytetracycline (OTC), chlortetracycline (CTC) of the investigated 5 TCs, and sulfamethazine (SMT), sulfamethoxazole (SMX), sulfadimethoxane (SDM) and sulfathiazole (STZ) of the investigated 6 SAs in water matrix cross-react to a similar degree within each family in RIA and SPE/RIA. Water samples were collected across a watershed in northern Colorado in addition to the influent and effluent of a wastewater treatment plant. SPE/RIA analysis of these samples was compared with SPE/liquid chromatography (LC)-mass spectrometry (MS) quantification of 5 TC and 6 SA compounds. Results of the study indicate that SPE/RIA can be an effective technique for monitoring antibiotic compounds in waters suspected to be contaminated with these compounds. The coupled method provides a sufficiently low detection limit (0.05 microg/L) to screen large sample sets at environmentally relevant concentrations. The method provides a semi-quantitative composite measurement of similar compounds in an antibiotic family without complex and expensive analytical equipment. PMID- 15276732 TI - Anaerobic digestion of olive mill wastewaters in biofilm reactors packed with granular activated carbon and "Manville" silica beads. AB - Anaerobic digestion is one of the most promising technologies for disposing olive mill wastewaters (OMWs). The process is generally carried out in the conventional contact bioreactors, which however are often unable to efficiently remove OMW phenolic compounds, that therefore occur in the effluents. The possibility of mitigating this problem by employing an anaerobic OMW-digesting microbial consortium passively immobilized in column reactors packed with granular activated carbon (GAC) or "Manville" silica beads (SB) was here investigated. Under batch conditions, both GAC- and SB-packed-bed biofilm reactors exhibited OMW COD and phenolic compound removal efficiencies markedly higher (from 60% to 250%) than those attained in a parallel anaerobic dispersed growth reactor developed with the same inoculum; GAC-reactor exhibited COD and phenolic compound depletion yields higher by 62% and 78%, respectively, than those achieved with the identically configured SB-biofilm reactor. Both biofilm reactors also mediated an extensive OMW remediation under continuous conditions, where GAC reactor was much more effective than the corresponding SB-one, and showed a tolerance to high and variable organic loads along with a volumetric productivity in terms of COD and phenolic compound removal significantly higher than those averagely displayed by most of the conventional and packed-bed laboratory-scale reactors previously proposed for the OMW digestion. PMID- 15276733 TI - Nitrification modelling in biofilms under inhibitory conditions. AB - A biofilm model has been developed for simulating nitrification in biofilms under inhibitory conditions. Nitrification inhibition has been modelled using uncompetitive inhibition kinetics. Inhibition kinetic experiments were performed by varying the bulk concentrations of inhibitory compound, aniline. Two sets of results were obtained with a nitrifying biofilm that was unacclimated to aniline and another which was acclimated to aniline. Fitting of the nitrification inhibition biofilm model to the experimental results yielded the nitrification inhibition constant, Ki, for aniline. Both the experiments yielded a value of about 3mg/L for Ki, which was similar to that obtained during nitrification inhibition experiments with suspended growth process carried out in an earlier study. The nitrification inhibition biofilm model is general and can be applied to nitrification inhibition with other toxic compounds. PMID- 15276734 TI - Bioavailability of wastewater-derived organic nitrogen to the alga Selenastrum Capricornutum. AB - Recent attempts to control cultural eutrophication in nitrogen-limited systems have focused on the simultaneous control of all forms of nitrogen with the underlying assumption that inorganic and organic nitrogen are equally bioavailable. To assess the validity of this assumption, algal growth bioassays were conducted on denitrified wastewater effluent samples, in the presence and absence of bacteria isolated from an effluent-receiving surface water. Bioassay results indicated that wastewater-derived dissolved organic nitrogen (DON) is not bioavailable to the algae Selenastrum Capricornutum in the absence of bacteria. However, approximately half of the wastewater-derived organic nitrogen was available to the algae in the presence of bacteria during a 2-week incubation. These results suggest that while it is inappropriate to assume that wastewater derived DON cannot cause cultural eutrophication, it will not cause as much eutrophication as inorganic nitrogen. Additional research is needed to develop methods of minimizing the discharge of bioavailable forms of wastewater-derived organic nitrogen by wastewater treatment plants. PMID- 15276735 TI - Use of synthetic zeolites for arsenate removal from pollutant water. AB - Arsenic is known to be a hazardous contaminant in drinking water that causes arsenical dermatitis and skin cancer. In the present work, the potential use of a variety of synthetic zeolites for removal of arsenic from water below the current and proposed EPA MCL has been examined at room temperature. Experiments have been conducted to examine the extent of arsenic removal as a function of pH. The effect of initial arsenic concentration and liquid to solid ratio in the batch reactions has also been studied. Zeolite NH4+/Y (NY6) showed significant arsenate removal capacity over a wide initial pH range of 2-12. Zeolite NY6 achieved this performance by buffering the initial pH to within a range of 3.5 < pH < 7 where uptake of arsenate onto aluminol surface groups is at a maximum. The high aluminum content of NY6 (i.e. low Si/Al ratio) was an important factor governing the improved performance of this zeolite relative to other tested zeolites with higher Si/Al ratio. The pH buffering capacity of NY6 could lead to savings in cost and process time for industrial effluent treatment due to avoidance of a pH pre-conditioning step prior to arsenate removal. PMID- 15276736 TI - Bioleaching of heavy metals from contaminated sediment by indigenous sulfur oxidizing bacteria in an air-lift bioreactor: effects of sulfur concentration. AB - The effects of sulfur concentration on the bioleaching of heavy metals from the sediment by indigenous sulfur-oxidizing bacteria were investigated in an air-lift reactor. Increasing the sulfur concentration from 0.5 to 5 g/l enhanced the rates of pH reduction, sulfate production and metal solubilization. A Michaelis-Menten type equation was used to explain the relationships between sulfur concentration, sulfate production and metal solubilization in the bioleaching process. After 8 days of bioleaching, 97-99% of Cu, 96-98% of Zn, 62-68% of Mn, 73-87% of Ni and 31-50% of Pb were solubilized from the sediment, respectively. The efficiency of metal solubilization was found to be related to the speciation of metal in the sediment. From economical consideration, the recommended sulfur dosage for the bioleaching of metals from the sediment is 3g/l. PMID- 15276737 TI - Predicting the effect of livestock inputs of E. coli on microbiological compliance of bathing waters. AB - Three alternative approaches to predicting delivery of faecal indicators from livestock sources to surface water in the catchment of the River Irvine, Ayrshire, Scotland, are described. These are a soil transport model which assumes all E. coli are transported through the soil, a regression model using observed E. coli concentrations in surface waters, and a distributed catchment model (PAMIMO). Each of these is linked to a simple group of equations describing inputs of E. coli from livestock to land, transport and inactivation in the river Irvine and mixing and inactivation in the sea. The models predict E. coli content of bathing water for Irvine beach. The regression model gives the best predictions of bathing water quality. The low values predicted by the soil transport model suggests that preventing surface runoff of faecal indicators from livestock would provide an adequate solution to the problem of bathing water contamination. PMID- 15276738 TI - Drinking water denitrification using a membrane bioreactor. AB - A membrane bioreactor (MBR) was investigated for denitrification of nitrate (NO3( )) contaminated drinking water. In the MBR, NO3(-) contaminated water flows through the lumen of tubular microporous membranes and NO3(-) diffuses through the membrane pores. Denitrification takes place on the shell side of the membranes, creating a driving force for mass transfer. The microporous membranes provide a high NO3(-) permeability, while separating the treated water from the microbial process, reducing carryover of organic carbon and sloughed biomass to the product water. Specific objectives of this research were to develop a model for NO3(-) mass transfer in the MBR, investigate the effect of shell and lumen velocity on NO3(-) mass transfer and investigate the effects of NO3(-) and organic carbon loading on denitrification rate and product water quality. A mathematical model of NO3(-) mass transfer was developed, which fit abiotic mass transfer data well. Correlations of dimensionless parameters were found to underestimate the overall NO3(-) mass transfer coefficient by 30-45%. The MBR achieved over 99% NO3(-) removal at an influent concentration of 200 mg NO3(-) NL(-1). The average NO3- flux to the biomass was 6.1g NO3(-)-Nm(-2)d(-1). Low effluent turbidity was achieved; however, approximately 8% of the added methanol partitioned into the product water. PMID- 15276739 TI - Rapid loss of estrogenicity of steroid estrogens by UVA photolysis and photocatalysis over an immobilised titanium dioxide catalyst. AB - The presence of low levels of natural and synthetic steroid estrogens in the aquatic environment, and their biological effects on aquatic organisms, are presently issues of concern. In this study, we investigated the temporal removal of estrogenic activity of several potent and environmentally relevant steroid estrogens by photocatalysis over an immobilised titanium dioxide (TiO2) catalyst. We used a recombinant yeast assay to measure estrogenic activity, which provided detection limits within the reactor of 53 ng/l for 17beta-estradiol and 17alpha ethinylestradiol, and 100 ng/l for estrone. Pseudo-first-order kinetic data showed that photocatalysis over titanium dioxide was equally effective at removing the estrogenic activity of all three steroid substrates in aqueous solutions (initial concentrations of 10 microg/l) with a 50% reduction in estrogenicity within 10 min. In control experiments without TiO2 catalyst, the rate of UVA photolysis of the steroid substrates varied, but was most effective with 17alpha-ethinylestradiol followed by estrone, and was least effective with 17beta-estradiol (0.42, 0.2 and < 0.1 times the rate achieved with photocatalysis, respectively). The application of photocatalysis for the removal of steroid compounds within STW effluent released into the aquatic environment is discussed. PMID- 15276740 TI - Spatial and temporal variability of priority volatile organic compounds in the Scheldt estuary. AB - The occurrence and spatial-temporal variability of 25 volatile organic compounds (VOCs) was studied from May 1998 to November 2000 in the Scheldt estuary. Target VOCs were selected from lists of priority pollutants to the North Sea, and included chlorinated short-chain hydrocarbons (CHCs), monocyclic aromatic hydrocarbons (MAHs) and chlorinated monocyclic aromatic hydrocarbons (CMAHs). Samples were taken between Vlissingen and Temse over a 95 km trajectory, and analysed by purge-and-trap and high resolution gas chromatography-mass spectrometry. Data were attended by analyses deemed 'in control' by a rigorous quality assurance/quality control program, as proposed by Quality Assurance of Information for Marine Environmental Monitoring in Europe (QUASIMEME). CHCs were among the most commonly found VOCs in the estuary, ranging from a few ng l(-1) to several microg l(-1). Most analytes were traceable to a single input source in the upper part of the estuary, which is densely populated and highly industrialized. By contrast, the occurrence of MAHs resulted from contributions of a wide spectrum of sources. The CMAHs were less abundant in the water column, and were mainly detected as chlorobenzene and 1,4-dichlorobenzene. Concentrations of several priority VOCs have markedly decreased since 1995 owing to emission reduction efforts within international frameworks for the protection of the North Sea. PMID- 15276741 TI - Liming as an advanced treatment for sludge sanitisation: helminth eggs elimination--Ascaris eggs as model. AB - The presence of helminth eggs (Ascaris eggs) in sewage sludge may constitute a sanitary risk when used as agricultural fertiliser. Sanitisation of sewage sludge can be achieved by treatment with quick lime, a process that destroys sludge pathogens in two ways: pH increase and temperature rise. Among the pathogens of epidemiological relevance, Ascaris eggs are the most resistant to liming, and, hence, may serve as indicators of hygienic quality of biosolids. This research aims at defining, between 50 degrees C and 60 degrees C, the time required in the case of limed sludge to obtain a product with a negligible level of viable Ascaris eggs. To achieve this objective, investigations on inactivation kinetics of Ascaris eggs were conducted in the following products: contaminated milk of lime; naturally contaminated sludge treated with slaked lime and heat; naturally contaminated sludge treated with quick lime; and sludge treated at full scale with quick lime. For the inactivation kinetics where a negligible level of Ascaris eggs was reached, the inactivation threshold was determined. Depending on the experimental situation, the inactivation threshold period was found to fluctuate between 5 and 75 min at 55 degrees C and between 1 and 8 min at 60 degrees C. PMID- 15276742 TI - Chromatographic separation of chlorophenoxy acid herbicides and their radiolytic degradation products in water samples. AB - HPLC procedure for simultaneous determination of chlorophenoxy acid herbicides and their radiolytic degradation products in waters is described with the use of octadecylsilica column and spectrophotometric detection at 280 nm. The satisfactory separation was achieved with a mobile phase of pH 2.5 consisting of 43.7 mM acetic acid with 40% (v/v) acetonitrile. Limit of detection values for herbicides and phenol derivatives were in the range of 19-41 microg/l and 10-60 microg/l, respectively. The developed method was applied for monitoring the effectiveness of radiolytic degradation of herbicides. Studies of products of gamma-radiolysis of 2,4-dichlorophenol have shown that the efficiency of this process is affected by the presence of naturally occurring scavengers of gamma radiation such as carbonates or nitrates. PMID- 15276743 TI - The estimation of NMVOC emissions from an urban-scale wastewater treatment plant. AB - The emissions of 19 different non-methane volatile organic compounds (NMVOCs) from the sewage treatment plant of the province of Eskisehir in Turkey were estimated. The estimations were based on the modified surface-renewal model suggested by EPA. The estimated total annual amounts of the pollutants emitted (from the plant's primary and secondary clarifier units and their weirs, as well as the aerated biological treatment unit) varied between a range of 0.00024 t (1,3-dichlorobenzene) and 0.1646 t (tetrachloroethylene). The corresponding flux data ranged from 9.98 x 10(-10)g cm(-2) h(-1) (1,3-dichlorobenzene) to 8450 x 10( 10)g cm(-2) h(-1) (tetrachloroethylene). Resulting total hourly NMVOC emission rate (0.041 kg h(-1)) was found not to exceed the current national standards. This work may be considered as a regional background for a possible contribution to the national and international emission inventory study on NMVOCs. PMID- 15276744 TI - Evaluating the effect of dissolved oxygen on ammonia-oxidizing bacterial communities in activated sludge. AB - The effect of dissolved oxygen (DO) on the communities of ammonia-oxidizing bacteria (AOB) in activated sludge was evaluated in lab-scale and full-scale reactors using the amoA gene as the basis for phylogenetic comparisons. Under controlled laboratory conditions, two chemostats seeded with activated sludge from the same source were operated with high-DO (8.5 mg/L) and low-DO (0.24 and 0.12 mg/L) concentrations for a period of 300 days. At the end of the operation period, the chemostats had enriched AOB communities that belonged to the Nitrosomonas europaea lineage, but were differentiable based on phylogenetic and kinetic analyses. The low-DO chemostat harbored the growth of two different groups within this lineage, differentiable by the amoA sequence comparison and by terminal fragment signatures. The difference in oxygen affinity between high-DO and low-DO enrichments was demonstrated by evaluating the growth kinetics as a function of oxygen concentration. The low-DO enrichment had a higher growth rate at DO concentrations below 4.7 mg/L, but the growth rate significantly decreased at higher DO concentrations, for which the high-DO enrichment experienced higher growth rates. In addition, the dynamic changes in AOB populations in two parallel trains within one full-scale treatment plant were evaluated in response to a significant reduction of DO in one of the treatment trains. Only the train operated with DO concentrations below 1mg/L favored the establishment of a population of AOB related to the N. europaea lineage. PMID- 15276745 TI - Application of multi-wavelength fluorometry for on-line monitoring of an anaerobic digestion process. AB - This work examined the use of multi-wavelength fluorometry for on-line monitoring of an anaerobic digestion process. Experiments were carried out in a laboratory scale anaerobic digestor fed with either synthetic or agricultural (cheese factory) wastewater. An in-line fiber optic probe installed in the external recirculation loop of the reactor was used to acquire fluorescence spectra with an interval of 5-10 min. The spectra were compared with analytical measurements taken at the same time to develop regression models, which were then used to predict concentrations of chemical oxygen demand, volatile fatty acids, and other key process parameters. A comparison of partial least squares (PLS), nonlinear principal components regression, and step-wise regression models on an independent set of data showed that the PLS model gave the best prediction accuracy. PMID- 15276746 TI - Removal of non-biodegradable organic matter from landfill leachates by adsorption. AB - Leachates produced at the La Zoreda landfill in Asturias, Spain, were recirculated through a simulated landfill pilot plant. Prior to recirculation, three loads of different amounts of Municipal Solid Waste (MSW) were added to the plant, forming in this way consecutive layers. When anaerobic digestion was almost completed, the leachates from the landfill were recirculated. After recirculation, a new load of MSW was added and two new recirculations were carried out. The organic load of the three landfill leachates recirculated through the anaerobic pilot plant decreased from initial values of 5108, 3782 and 2560 mg/l to values of between 1500 and 1600 mg/l. Despite achieving reductions in the organic load of the leachate, a residual organic load still remained that was composed of non-biodegradable organic constituents such as humic substances. Similar values of the chemical oxygen demand (COD) were obtained when the landfill leachate was treated by a pressurised anoxic-aerobic process followed by ultrafiltration. After recirculation through the pilot plant, physico-chemical treatment was carried out to reduce the COD of the leachate. The pH of the leachate was decreased to a value of 1.5 to precipitate the humic fraction, obtaining a reduction in COD of about 13.5%. The supernatant liquid was treated with activated carbon and different resins, XAD-8, XAD-4 and IR-120. Activated carbon presented the highest adsorption capacities, obtaining COD values for the treated leachate in the order of 200mg/l. Similar results were obtained when treating with activated carbon, the leachate from the biological treatment plant at the La Zoreda landfill; in this case without decreasing the pH. PMID- 15276747 TI - The use of magnetic bed conditioning and pH control to enhance filtration by natural titanomagnetite. AB - The fine particle size and magnetic properties of natural titanomagnetite (TM) iron sand offer options for novel granular water filtration systems. The isoelectric points (IEPs) of a variety of natural and synthetic TMs were determined to be in the range from 3.60+/-0.06 to 4.01+/-0.06. TM beds (125-150 microm particle size) expanded by up to 28% were successfully conditioned with vertical fields of approximately 0.018 T and produced up to a three-fold increase in hydraulic conductivity. Filtration studies showed that the filtration efficiency decreased with bed expansion but that this reduction was more than compensated for by reducing the pH to below the IEP of TM. Backwash performance was improved by magnetic conditioning which allowed higher interstitial flow velocities at a given bed expansion. PMID- 15276748 TI - Simultaneous biological removal of nitrogen, carbon and sulfur by denitrification. AB - Refinery wastewaters may contain aromatic compounds and high concentrations of sulfide and ammonium which must be removed before discharging into water bodies. In this work, biological denitrification was used to eliminate carbon, nitrogen and sulfur in an anaerobic continuous stirred tank reactor of 1.3 L and a hydraulic retention time of 2 d. Acetate and nitrate at a C/N ratio of 1.45 were fed at loading rates of 0.29 kg C/m3 d and 0.2 kg N/m3 d, respectively. Under steady-state denitrifying conditions, the carbon and nitrogen removal efficiencies were higher than 90%. Also, under these conditions, sulfide (S(2-)) was fed to the reactor at several sulfide loading rates (0.042-0.294 kg S(2-)/m3 d). The high nitrate removal efficiency of the denitrification process was maintained along the whole process, whereas the carbon removal was 65% even at sulfide loading rates of 0.294 kg S(2-)/m3 d. The sulfide removal increased up to approximately 99% via partial oxidation to insoluble elemental sulfur (S0) that accumulated inside the reactor. These results indicated that denitrification is a feasible process for the simultaneous removal of nitrogen, carbon and sulfur from effluents of the petroleum industry. PMID- 15276749 TI - Development of cultures capable of reducing perchlorate and nitrate in high salt solutions. AB - An ion exchange process with biological perchlorate and nitrate destruction and reuse of spent regenerant brine has been proposed as an efficient and environmentally sound method to treat perchlorate-contaminated groundwater. A culture capable of reducing perchlorate and nitrate in spent ion exchange regenerant brine containing at least 30 g/L NaCl is needed for this to be feasible. A batch culture inoculated from activated sludge failed to acclimate to more than 15 g/L NaCl whether nitrate was present or not. A mixed culture inoculated from marine sediment was capable of simultaneously reducing 100mg/L perchlorate and denitrifying 500 mg/L nitrate within 5 h in a synthetic medium in the presence of 30 g/L NaCl. The growth conditions to maintain this culture in a healthy state required the addition of trace metals, Na2S, and phosphate. A second culture capable of removing 100 mg/L perchlorate from synthetic medium containing 60 g/L NaCl within 24 h was also developed. PMID- 15276750 TI - Testing a surface tension-based model to predict the salting out of polycyclic aromatic hydrocarbons in model environmental solutions. AB - Molar-based Setschenow constants (Ks) for six alkali and alkaline earth metal based inorganic salts were determined at 20 degrees C to evaluate their influence on the solubilities, and thus the aqueous activity coefficients, of three polycyclic aromatic hydrocarbons (PAHs). The six salts tested exhibited a wide range of Ks values, varying from 0.105 +/- 0.009 M(-1) (for NaClO4 and pyrene) to 1.29 +/- 0.17 M(-1) (for K2SO4 and perylene). In general, salting out effects with these electrolytes were observed in the order Ca2+ > Na+ > K+ and SO4(2-) > Cl- > ClO4-, consistent with previous reports. However, the expected salting out trend of perylene > pyrene > naphthalene was only observed with K2SO4. In CaCl2 solutions, the Ks value of pyrene was significantly lower than that of naphthalene. For NaCl, KCl and NaClO4, pyrene Ks values were found to be lower than, but not significantly different from, those of naphthalene. Setschenow constants for all six salts were predicted using a semi-empirical, thermodynamically-based equation that relates the standard free energy change associated with transferring solutes from water to a salt solution to the difference in surface tensions between the two solutions. With this equation, predicted Ks values were in reasonable agreement with observed Ks values (generally within +/- 50%). Lack of better agreement between predicted and observed values likely reflects the inability of the simple surface tension model to account for all interactions among the cations, anions, PAH molecules and water molecules in the respective systems. PMID- 15276751 TI - Integrated real-time control strategy for nitrogen removal in swine wastewater treatment using sequencing batch reactors. AB - A new integrated real-time control system was designed and operated with fluctuating influent loads for swine wastewater treatment. The system was operated with automatic addition control of an external carbon source, using real time control technology, which utilized the oxidation-reduction potential (ORP) and the pH as parameters to control the anoxic phase and oxic phase, respectively. The fluctuations in swine wastewater concentration are extreme; an influent with a low C/N ratio is deficient in organic carbon, and a low carbon source level can limit the overall biological denitrification process. Consequently, a sufficient organic source must be provided for proper denitrification. The feasibility of using swine waste as an external carbon source for enhanced biological nitrogen removal was investigated. The real-time control made it possible to optimize the quantity of swine waste added as the load fluctuated from cycle to cycle. The average removal efficiencies achieved for TOC and nitrogen were over 94% and 96%, respectively, using the integrated real-time control strategy. PMID- 15276753 TI - Evaluating trends in biofilm density using the UMCCA model. AB - We present a series of modeling cases that illustrate the trends described by the unique features of the unified multiple-component cellular automaton (UMCCA) model for a heterogeneous, two-dimensional biofilm. The outputs of the UMCCA model show five general trends. (1) The concentration profiles for the two soluble microbial products are opposite the profile for original substrate. (2) The top of the biofilm is dominated by active biomass and EPS, while the bottom is dominated by residual inert biomass. Within the top layers, active biomass has a much higher concentration than EPS. (3) The top of all biofilm is quite "fluffy," while the bottom is dense. (4) The peak of the composite density does not correspond to the peak of active biomass. (5) All biomass types show considerable local heterogeneity. The series of cases also indicate what conditions lead to particular characteristics observed in some biofilms. Biofilm clusters are promoted by substrate limitation, a high detachment rate, or strong consolidation. A high biofilm density is associated with an old biofilm, which is favored by a low substrate concentration, a high detachment rate, and strong consolidation. Old biofilms also can develop low-density pockets near the substratum, a possible cause of sloughing. Local heterogeneity is generally related to the same factors that cause a high density. We also solved the UMCCA model for conditions similar to the experiments of Bishop et al. (Water Sci. Technol. 31(1) (1995) 143), who measured the total biomass density in layers from the substratum. The model outputs captured all the major trends in the experimental data: the overall thickness and density of biofilms increase with time, and the total biomass density is 5-10 times greater near the substratum than near the top of the biofilm. Furthermore, the model indicates that the residual inert biomass becomes denser toward the substratum, a trend observed experimentally; the UMCCA model suggests that this trend is due to the combined effects of consolidation and inert biomass having a larger maximum density. PMID- 15276752 TI - Modeling the development of biofilm density including active bacteria, inert biomass, and extracellular polymeric substances. AB - We present the unified multi-component cellular automaton (UMCCA) model, which predicts quantitatively the development of the biofilm's composite density for three biofilm components: active bacteria, inert or dead biomass, and extracellular polymeric substances. The model also describes the concentrations of three soluble organic components (soluble substrate and two types of soluble microbial products) and oxygen. The UMCCA model is a hybrid discrete-differential mathematical model and introduces the novel feature of biofilm consolidation. Our hypothesis is that the fluid over the biofilm creates pressures and vibrations that cause the biofilm to consolidate, or pack itself to a higher density over time. Each biofilm compartment in the model output consolidates to a different degree that depends on the age of its biomass. The UMCCA model also adds a cellular automaton algorithm that identifies the path of least resistance and directly moves excess biomass along that path, thereby ensuring that the excess biomass is distributed efficiently. A companion paper illustrates the trends that the UMCCA model is able to represent and shows a comparison with experimental results. PMID- 15276755 TI - Enhanced in situ denitrification for a municipal well. AB - In 37% of small community systems in Nebraska at least one sample exceeded the drinking water standard for nitrate of 10 mg N L(-1) during the period from 1982 to 1998 (US Bureau of Reclamation, US Department of the Interior, 1999). In this experiment a daisy well system was designed to promote denitrification in the radial capture zone of a municipal well with nitrate-N levels> 10mg L(-1), and thereby bring the nitrate concentration into compliance. The remediation design consisted of eight 15 cm diameter outer perimeter reduction wells and eight 5 cm diameter inner perimeter oxidation wells which are located roughly 18 and 9 m, respectively, from the municipal well, which serves as the extraction well. Endemic microbes are stimulated by pulsing separate injections of acetate-C and nitrate contaminated water (C:N = 1.2) to enhance denitrification in the capture zone. Water was extracted from the municipal well at 6.6 L s(-1) (liters per second). A 45% nitrate reduction occurred in municipal well samples when the total acetate-C input was increased by lengthening the acetate pulse from 1.0 to 1.5 h (C:N=1.8). Nitrate concentration stabilized at about 6.3 mg NO3-N L(-1) for two weeks during alternating acetate pulse lengths. The in situ denitrification process was sustained for three months without evidence of clogging. Results from this experiment indicated that the extracted water was in compliance with respect to nitrate, nitrite, trihalomethanes, turbidity, and total and fecal coliforms; however, the total plate count exceeded the maximum permissible limit (500 cfu/mL). PMID- 15276754 TI - The role of zeolites in wastewater treatment of printing inks. AB - The adsorption of residual organic pollutants from flocculated printing ink wastewater onto several synthetic zeolites was investigated as a finishing method for additional reduction of TOC. The nonselective removal of total organic content was studied. The amount of adsorbed organics was largest for ZSM-5 and NH4-Beta while the other zeolites studied showed lower efficiency, suggesting that adsorption is independent of pore structure. The adsorption rates of organic pollutants were fast. Although the TOC removal increases with increasing amount of zeolite, because of the necessity of additional filtration to lower turbidity to required levels, 5.0 g/L of zeolite was found to be optimum. The 88% reduction of TOC obtained with a single flocculation treatment was improved with the combination of flocculation and adsorption with ZSM-5 which resulted in the overall TOC efficiency of 95%. The addition of zeolites in decantated supernatant water, obtained after flocculation, was also studied in order to assess the effect of floc on zeolite capacity. A decrease in adsorption capacity occurred only if a coagulant concentration less than optimal was applied. Removal efficiency then decreased by around 10%. It was concluded that flocculation followed by adsorption with zeolites is an effective treatment method for this kind of wastewater. PMID- 15276756 TI - Aerobic granulation with industrial wastewater in sequencing batch reactors. AB - Granular sludge formation was promoted in two laboratory scale sequencing batch reactors (SBRs), R1 and R2 fed with industrial wastewater produced in a laboratory for analysis of dairy products. Both reactors were operated under similar conditions during most of the experimental period. However, an anoxic phase between 10 and 30 min was included at the beginning of every cycle of operation of R1, but not in R2. Organic and nitrogen loading rates applied to both systems were high, up to 7 g COD/(L d) and 0.7 g N/(L d). Nitrogen removal efficiency was 70% in both units even considering that R2 was operated always under aerobic conditions. Granules with similar morphology were developed in both systems. Size distribution was comprehended between 0.25 and 4.0 mm for both systems. The presence of TSS in the effluent of the SBRs was strongly affected by either the length of the withdrawal period or by the particulated COD to biomass ratio (CODp/VSS) applied to the systems. The lower concentrations of TSS in the effluent were attained when the systems were operated with a CODp/VSS ratio lower than 0.12 g COD/g VSS. There was a strong reduction of the average TSS content in the effluent from 450 to 200 and 150 mg TSS/L when the length of the withdrawal period was diminished sequentially from 3 to 1 and 0.5 min, respectively. This was caused by a more intensive washout of small suspended biomass aggregates that took place when the length of this period was shortened. PMID- 15276757 TI - A non-biological surrogate for sequential disinfection processes. AB - An evaluation of Fluorescent YG-microspheres (Polysciences Inc.) was performed to simulate Cryptosporidium parvum (C. parvum) oocysts inactivation in treatment systems that utilize multiple disinfectants. Experiments were conducted in batch reactors that included an ozone primary stage and a secondary free chlorine treatment stage. A flow cytometer was used to track changes in the fluorescence intensity distribution due to exposure to the chemical disinfectant. Microsphere 'survival ratios' (N/No) were calibrated by selecting an appropriate fluorescence intensity threshold to replicate the inactivation of different C. parvum oocysts strains. Results showed that fluorescent microspheres displayed synergistic effects in the presence of two sequential disinfectants. In addition, microsphere structural tests showed that the polystyrene surface was damaged due to exposure to ozone. This polystyrene damage enhanced the diffusion of the secondary disinfectant into the microsphere, where dye was degraded in the opened polymer layer. As a result, YG-fluorescent microspheres is a promising non-biological technique that is capable of producing similar synergistic behavior as with C. parvum oocysts exposed to ozone followed by chlorine. PMID- 15276758 TI - Stir bar contamination: a method to establish and maintain constant water concentrations of poorly water-soluble chemicals in bioconcentration experiments. AB - A novel experimental system to establish and maintain constant dissolved concentrations of poorly water-soluble compounds for bioconcentration experiments with algae was developed. Although still recommended in the literature as a "non adsorptive" material, a commercially available Teflon stir bar that was preloaded with polychlorinated biphenyls (PCBs) proved to be an effective donor for partitioning controlled delivery. When placed in a bioconcentration test chamber, the PCB concentrations in water and in the headspace remained constant for up to 8 days. Henry's law constants determined in experiments using water only were in good accordance with values found in the literature, indicating that the chemical was truly dissolved. When algae (Emiliana huxleyi, class prymnesiophyceae) were added to the chamber, the water and air concentrations varied initially but stabilised within several hours, and smooth uptake curves were obtained for the algae. This indicates that the contamination system compensates for chemical loss from the water column. In addition, the Henry's law constants and the headspace measurements were used to estimate what fraction of the PCB in filtered water was associated with dissolved organic carbon, opening the opportunity to constrain one of the major artefacts in bioconcentration experiments. PMID- 15276759 TI - Investigating phosphorus interactions with bed sediments in a fluvial environment using a recirculating flume and intact soil cores. AB - Phosphorus uptake by bed sediments in surface drains can reduce phosphorus exports from irrigated land. This paper reports on an investigation into the effects of velocity and water depth on phosphorus uptake by bed sediments, which consisted of eight sequential flow events conducted in a recirculating flume as well as a concurrent experiment using sediment cores. For the heavy clay bed sediment discussed in this paper, velocity and depth of water column had no significant effect on net phosphorus uptake and the rates of phosphorus uptake in either the cores or the recirculating flume. The most significant factor affecting phosphorus uptake was the experiment number which represented the sequential nature of experiments within the flume and increasing phosphorus saturation of the surface sediments. Of the kinetic equations used to describe phosphorus uptake (Elovich, boundary layer and diffusion) the Elovich equation provided the best representation of the results, both in terms of the adj-R2 values and the absence of systematic errors in the residuals. Results suggest that intact soil cores may be used to parameterise rate equations such as the Elovich equation for use in process-based mathematical models of phosphorus transport in fluvial systems. PMID- 15276760 TI - Irreversible membrane fouling during ultrafiltration of surface water. AB - For more efficient use of membranes, the control of irreversible membrane fouling, which can be defined as fouling requiring chemical reagents to be mitigated, is of importance. In this study, irreversible fouling caused by constituents in surface water was investigated, based on a long-term pilot scale study. The membrane employed was a low-pressure hydrophobic ultrafiltration (UF) membrane made of polysulfone and having a molecular weight cutoff of 750,000 Da. Various chemical reagents were examined to overcome the irreversible fouling that had developed through 5 months of continuous filtration. Among the tested cleaning reagents, alkaline (NaOH) and oxidizing reagent (NaClO) showed good performance in the restoration of membrane permeability, which implied that organic matter played an important role in the development of the irreversible fouling in this study. Chemical analysis, adsorptive fractionation methods, fluorescence excitation-emission matrix (EEM) and Fourie-transformed infra-red (FTIR) spectra analysis were applied to elucidate which fraction of organic matter caused the irreversible fouling. All of the analysis indicated that polysaccharide-like organic matter was responsible for the evolution of the irreversible fouling. In addition to organic matter, presumably iron and manganese also contributed to the irreversible fouling to some extent. PMID- 15276761 TI - Dynamics and influencing factors of heterotrophic bacterial utilization of acetate in constructed wetlands treating woodwaste leachate. AB - 14C-acetate was used as a tracer of substrates for heterotrophic bacteria to investigate the dynamics and influencing factors of bacterial activity in surface flow wetlands treating woodwaste leachate. Epiphytic assimilation and mineralization of acetate increased considerably over three weeks of microbial colonization. No significant longitudinal variation in acetate uptake was found. The temporal variation of bacterial activity was neither correlated to water temperature nor acetic acid concentration. Mineralization percentage varied significantly with acetate uptake rate. Acetate uptake rates of bacterioplankton (408+/-258 microg L(-1) h(-1)), epiphyton (67.7+/-41.7 mg m(-2) h(-1)) and sedimentary bacteria (26.7+/-25.6 microg g(-1) h(-1)) were influenced by the concentrations of organic substrates and inorganic nutrients. Sedimentary bacteria contributed to the majority (55-73%) of the total heterotrophic acetate uptake, while epiphytic bacteria played only a minor role (1-3%). The acetate mineralization percentage of sediment (16%) was much lower than that of water (55%) and epiphyton (64%). Addition of ammonium nitrate fertilizer increased acetate uptake rates significantly, especially for sediment. No remarkable changes in mineralization percentage and the relative importance of water, sediment and epiphyton were found due to fertilization. PMID- 15276762 TI - Neural networks provide superior description of Giardia lamblia inactivation by free chlorine. AB - Use of conventional models to describe data on microbial inactivation during disinfection suffers from limitations with respect to flexibility and direct quantitative incorporation of water quality variables. This paper develops an approach to analysis of such data using neural networks (NNs). Using the data on free chlorine inactivation of Giardia lamblia previously reported, it was found that the use of an NN with a single hidden layer and four hidden neurons provided a superior (better) fit to the data with a reduced number of model parameters when compared to the fitting of this data using a conventional approach. Therefore, the use of NN models should be considered in the future assessment of microbial inactivation during disinfection. Incorporation of additional facets of the disinfection process, such a disinfectant decay, needs to be considered in subsequent development of this approach. PMID- 15276763 TI - Treatment of biorefractory organic compounds in wool scour effluent by hydroxyl radical oxidation. AB - Wool scouring effluent that had been treated with chemical flocculation and aerobic biological treatment (Sirolan CFB effluent) was tertiary treated by hydroxyl radical oxidation to remove residual organic compounds. These compounds impart a high chemical oxygen demand of 500-3000 mg/L and dark colour. However, a H2O2/UV process was found to effectively treat the majority of residual compounds, with up to 75% COD, 85% total organic carbon, and 100% removal of colour (T(480 nm)) achieved. This was despite the effluent being strongly absorbing in the UV region, with a film thickness of 0.21 mm reducing T(254 nm) by 50%. Treatment was unaffected by pH over the range 3-9. H2O2/UV treatment increased the biodegradability of the effluent (5-day biochemical oxygen demand increased from < 10 to 86 mg/L), but a combined chemical and biological process did not increase maximum COD removal or overall process efficiency. The tertiary treated effluent had a final COD in the range 125-750 mg/L, equating to a total COD removal from raw wool scour effluent of approximately 97.5%. This degree of treatment is sufficient for discharge in many, but not all, circumstances. PMID- 15276764 TI - Comments on "effect of extended idle conditions on structure and activity of granular activated sludge" by Zhu and Wilderer. PMID- 15276766 TI - Comment on "Degradation of monomethylmercury chloride by hydroxyl radicals in simulated natural waters". PMID- 15276768 TI - Fatal meningitis in a calf caused by Mannheimia varigena. AB - Mannheimia varigena was identified as the etiologic agent of meningitis in a young Belgian White Blue heifer calf. Species identification of the bacterium was done by phenotyping and molecularly confirmed by tDNA-PCR. Standard bacteriological examination might fail to differentiate species belonging to the genus Mannheimia. PMID- 15276769 TI - Detection of bacterial DNA in synovial fluid from horses with infectious synovitis. AB - Standard culturing techniques are often unrewarding in confirming diagnosis of synovial infection in the equine patient. Several human studies report the use of sensitive polymerase chain reaction (PCR) techniques for the detection of bacterial involvement in acute synovitis. However, successful extraction of bacterial DNA directly from clinical samples from horses without prior culture has not been reported yet. The goal of this study was to develop a sensitive and reliable method for molecular detection and identification of bacterial species in synovial fluid from horses with infectious synovitis. Synovial fluid samples from 6 horses with culture confirmed synovial infection were used for broad range 16S rRNA gene PCR. Synovial aspirates of 2 healthy horses were used as negative controls. Following extraction and purification of synovial fluid DNA, all samples were processed by touchdown PCR. Amplicons were detected by reverse line blot hybridisation and visualised with chemiluminescence. Pathogen-specific detection of 16S rRNA gene sequences was successful in all 6 synovial fluid samples. No bacterial DNA was detected in the aspirates from the negative control horses using touchdown PCR followed by 25 additional cycles of amplification. The identity of the pathogens was confirmed by DNA sequencing of the amplicons. It can be concluded that broad range 16S rRNA gene PCR followed by reverse line blot hybridisation is a promising technique for detection of bacterial DNA in synovial fluid samples. Further research should aim at the detection of bacterial DNA in synovial fluid samples suspected of infection but having negative culture results. When the 16S PCR proves to be reliable and more sensitive than standard culturing techniques, it may become a powerful tool in the diagnosis of synovial infection. PMID- 15276770 TI - Prevalence of exposure and infection of Lawsonia intracellularis among slaughter age pigs. AB - The extent of clinical or subclinical infection associated with Lawsonia intracellularis within Dutch pig herds was uncertain. A case-control study of slaughter age pigs was used to study natural infection within Dutch herds and to compare diagnostic methods. From six case herds where clinical disease had been identified recently, and six disease-free herds, 40 pigs of slaughter-age were examined postmortem. The diagnostic methods used were: serology, gross examination, Haematoxylin and Eosin stain (HE), Warthin-Starry silver stain, Lawsonia-specific indirect immunoperoxidase of the ileum, and PCR of ileum mucosa and colon contents. There were 59% seropositive pigs in case herds and 26% seropositive pigs in control herds. Using immunohistochemistry, 57% of case herds and 46% of control herds were bacteria positive in the ileum mucosa. It was concluded that a majority of Dutch herds contain L. intracellularis infected finisher pigs. In some herds this is associated with clinical outbreaks of acute haemorrhagic enteropathy but in other herds no clinical disease is apparent. Many seropositive pigs in herds without clinical disease had evidence of Lawsonia antigen in sites other than the apical cytoplasm of proliferating epithelial cells, particularly the supranuclear region. It was uncertain whether to classify these pigs as having "recovered" from an infection or whether they have a sub clinical or chronic form of the disease. We concluded that PCR examination of faeces and serology probably provide more specific results than gross examinations at slaughter, and that a monoclonal antibody-based examination of ileum mucosa should be the accepted screening method for this infection. PMID- 15276771 TI - Characterization of native and recombinant bovine pregnancy-associated glycoproteins. AB - Pregnancy-associated glycoproteins (PAGs) are products of the ruminant placenta that belong to the aspartic proteinase family. Extensive glycosylation may account for the size and heterogeneity of their molecules. To assess this we investigated the effect of glycosidase and tunicamycin treatments on native (n) and mammalian-cell generated recombinant (r) bovine PAGs. Native PAG came from explant culture conditioned medium (150 days pregnancy) while rPAG was obtained by transfection of HEK 293 cells with the bPAG-1 gene employing the PRcRSV expression vector. The undigested nPAG gave a homogenous band at 67 kDa after one dimensional SDS-PAGE, silver staining and Western blotting, but rPAG gave dual bands at 54 and 52 kDa. PNGase F digestion of nPAG gave five bands ranging from 60 to 37 kDa and digestion of rPAG gave three bands ranging from 54 to 37 kDa. On two-dimensional electrophoresis, the undigested pI ranges of n- and rPAGs were 4.7-5.6 and 7.3-8.8, respectively. The digested isoforms of n- and rPAGs had pI ranges from 5.1 to 8.5 and 7.9-8.5, respectively. Tunicamycin treatment had no effect on the mobility of nPAG but it had a pronounced time-dependant effect on the mobility of rPAG. Our findings indicate that both n- and rPAGs have principally N-linked oligosacharides. PMID- 15276772 TI - The cytokines of bovine mammary gland: prospects for diagnosis and therapy. AB - The lack of efficacy of conventional strategies for the maintenance of healthy udders in domestic cattle has prompted studies on the use of cytokines for this purpose. The adjuvant use of recombinant bovine cytokines, such as IL-2, IFN gamma and TNF-alpha, in normal mammary gland, mobilizes innate and acquired immunity. However, stimulated immunity does not prevent or eradicate infection, particularly in the case of Staphylococcus aureus mastitis. Cytokines do, however, improve the bactericidal efficiency of certain antibiotics. The subtle and sensitive changes in the cytokine network of normal and mastitic bovine mammary gland may encourage the use of cytokines in the diagnosis and prognosis of udder health. Numerous studies support this hypothesis, and detection and monitoring of cytokines could become an important alternative management for udder health. The use of cytokines in the immunotherapy, diagnosis and prognosis of mastitis will grow with knowledge of the cytokine network in bovine mammary glands and the development of efficient cytokine diagnostic techniques. PMID- 15276773 TI - Quantification of MDR-1 gene expression in canine tissues by real-time reverse transcription quantitative polymerase chain reaction. AB - MDR-1 gene product mediated multidrug resistance is thought to play a major role in the outcome of chemotherapy in some canine tumors, especially malignant lymphoma. In the present study, MDR-1 RNA expression in normal lymph node and liver tissue as well as in tumor biopsies from 23 dogs with lymphomas and two dogs with liver tumors was measured by real-time RT-quantitative PCR. MDR-1 gene expression was detected in all samples analyzed. Comparably high MDR-1 RNA levels were measured in all normal liver tissues, one of the lymphomas and a cholangiocarcinoma. MDR-1 expression levels in canine lymphomas were found to vary over a wide range with most tumors expressing relative low levels. Interestingly, gastrointestinal lymphomas expressed higher MDR-1 RNA levels than multicentric lymphomas (p = 0.03). In conclusion, real-time RT-quantitative PCR appears to be a suitable method for sensitive and quantitative determination of MDR-1 gene expression in canine normal and neoplastic tissues. PMID- 15276774 TI - A method for in vitro evaluation of protein hydrolysates for potential inclusion in veterinary diets. AB - A candidate chicken-protein hydrolysate was subjected to high-performance size exclusion chromatography to characterize its molecular weight profile. An inhibition ELISA assay was developed to assess the residual antigenicity, using canine serum IgG produced by dogs sensitized to the intact protein. Finally, the hydrolysate was compared to the intact protein through electrophoresis and immunoblotting. The chicken hydrolysate had a suitable molecular weight profile with 96.9% reduced to less than 10 kDa peptides. The inhibition ELISA demonstrated a residual antigenic mass of 1.5% compared with the intact protein. Immunoblotting demonstrated a strong immunoreactive band at 68-70 kDa consistent with chicken serum albumin in the intact protein, which was absent in the hydrolysate. These results demonstrate the suitability of the chicken hydrolysate for use in a protein hydrolysate diet, and provide a basis for the future comparisons of the peptide components of hydrolyzed protein diets so that veterinarians may make more informed decisions in their dietary prescriptions. PMID- 15276775 TI - Green tea flavan-3-ols and oligomeric proanthocyanidins inhibit the motility of infective larvae of Teladorsagia circumcincta and Trichostrongylus colubriformis in vitro. AB - The effects of a hot water infusion and an aqueous acetone extract of green tea (Camellia sinensis) on the motility of infective larvae of the sheep nematodes Teladorsagia circumcincta and Trichostrongylus colubriformis were investigated under in vitro conditions. The infusion and extract dose-dependently inactivated the infective larvae as assessed by the larval migration inhibition (LMI) assay. To determine the components responsible for the inhibitory activity, the hot water infusion and aqueous acetone extract of green tea were fractionated on Sephadex LH-20 and the green tea extract fractions (GTE-I-VIII) characterised by mass spectrometry. The larvae were exposed to increasing concentrations of these GTE fractions. Fractions containing epigallocatechin gallate (EGCG) and proanthocyanidin oligomers were most effective. GTE fractions were more effective against T. circumcincta than T. colubriformis larvae as assessed by the LMI assay. PMID- 15276776 TI - Pharmacokinetics of progesterone in dromedary camels (Camelus dromedarim). AB - The effect of ovariectomy or administration of progesterone (P4) on the disposition kinetics of P4 was determined in dromedary camels. The disappearance of P4 from peripheral circulation of the camel following ovariectomy or after a single intravenous (I.V.) injection of 1 mg kg(-1) body weight followed a bioexponential curve. Both curves were parallel indicating that the disappearance of injected P4 behaved similarly to endogenous P4. The mean (+/- SD) half-life calculated from the slower component of this decline, was 26.0 +/- 2.0 min after I.V. injection of P4 and 28.0 +/- 2.1 min after ovariectomy. The apparent volume of distribution (1370 ml kg(-1)) exceeded total body water suggesting extensive tissue penetration. PMID- 15276777 TI - Effect of matrix depleting agents on the expression of chondrocyte metabolism by equine chondrocytes. AB - This study was carried out to investigate the effect of two enzymes (collagenase and chondroitinase) and two cytokines/metabolites (interleukin-1beta and retinoic acid) of known catabolic activity on the expression of cartilage metabolism/phenotype in equine articular cartilage. Articular cartilage explants from 11 horses (5-13 years old) were treated for 48 h and assayed for total sulphated glycosaminoglycan (GAG), the incorporation of 35S-sulphate, collagen degradation and mRNA expression of the proteoglycans collagen II, collagen IIA, collagen III, collagen IX, collagen X, collagen XI and glyceraldehyde-3-phosphate (GAPDH). Purified collagenase and retinoic acid were responsible for increased GAG loss from the tissues. Chondroitinase, responsible for catalysing the elimination of glucuronate residues from chondroitin A, B and C (Chondroitinase ABC) and retinoic acid treatment induced an inhibition of proteoglycan synthesis, whereas collagenase treatment did not. Collagenase activity was correlated with increased appearance of the CB11B epitope and type II collagen denaturation. By RT-PCR there was evidence of expression of altered collagen type IIA in purified collagenase treated tissues. PMID- 15276778 TI - Pulmonary artery wedge pressure measurement in healthy warmblood horses and in warmblood horses with mitral valve insufficiencies of various degrees during standardised treadmill exercise. AB - In 12 healthy warmblood horses and 10 horses with mitral valve insufficiencies (MVI) of various degrees heart rate and pulmonary artery wedge pressure (PWP) was measured at rest and during standardised exercise on a high speed treadmill. There was a significant increase in PWP with each change in speed of the treadmill (p < 0.01). The PWP of horses with mild mitral valve regurgitation under working conditions was not significantly different compared to the healthy horses. The horses with moderate mitral valve regurgitation showed a significant higher pulmonary artery wedge pressure at rest and during exercise compared to the healthy horses (p < 0.01) at rest and during treadmill velocity. The tendencies were seen that mild mitral valve regurgitation results only in mild hemodynamic changes during exercise, while moderate MVI have an important influence on haemodynamics. PMID- 15276779 TI - Genome sequence analysis of 10 Dutch porcine circovirus type 2 (PCV-2) isolates from a PMWS case-control study. AB - The factors responsible for the emergence of post-weaning multisystemic wasting syndrome (PMWS) as an epidemic disease with significant impact upon the pig industry are not all known. Porcine circovirus type 2 (PCV-2) has been shown to be necessary but not sufficient for the expression of PMWS. Retrospective serological and molecular surveys have shown that PCV-2 was widespread and was maintained with only occasional reports of sporadic PMWS in the 30 year period prior to the recent emergence of the epidemic form of the syndrome. However, the recent spread of the disease in Europe and elsewhere has pointed to the transmission of a novel pathogen. One explanation to reconcile this paradox is that PWMS is caused by a unique PCV-2 variant that is being spread through pig populations. To test this hypothesis, complete genomes (1767 bp) of 10 Dutch PCV 2 isolates from 4 PMWS affected premises and 6 farms without PMWS were sequenced. Phylogenetic analysis showed that these sequences were grouped together although they differed on 77 nucleotide positions relative to each other (95.6-100% identity between the 10 isolates). None of these nucleotide changes identified impacted upon transcriptional elements or other important recognised features of the genome of the PCV-2. Amino acid changes were recorded on 4 positions in ORF1 and on 16 positions in ORF2 but, importantly, no consistent pattern was evident between PCV-2 isolates from affected and control pigs. These data provide further evidence to suggest that factor(s) in addition to PCV-2 are necessary in the development of PMWS. PMID- 15276780 TI - Animal models for the study of the effect of prolonged stress on lactation in rats. AB - The suppressive effect of stress on lactation is well acknowledged. Animal models used to study this problem have used primarily acute stressors. As such, they are not adequate for the study of the effect of prolonged stress on lactation. This study aimed at developing a paradigm(s) in rats for the study of the effect on lactation of prolonged exposure to a stressful situation. Two models were examined. The first consisted of a separation paradigm whereby dams had 1, 2, 4, or 6 contacts per day with their pups, but only for a total of 4 h per day. The second model implicated the introduction of a male intruder into the home cage during the 12-h light period. Both protocols were implemented for four consecutive days beginning on Days 10/11 postpartum. Milk release was measured on Day 13/14 postpartum. For both models, milk release was significantly reduced following the preexposure period. During the assessment of milk release, maternal behavior of the experimental and control animals was similar at all times. It is advanced that these two rat models of stress are appropriate for the study of lactation following a prolonged exposure to stress. PMID- 15276781 TI - Intracerebroventricular administration of chicken motilin does not induce hyperphagia in meat-type chicks. AB - The effect of chicken motilin on food intake was investigated in meat-type chicks under ad libitum feeding, refeeding, and fasting conditions. We found that the intracerebroventricular injection of chicken motilin (0.1 and 0.2 microg) tended to increase food intake under ad libitum feeding and refeeding conditions at 60 min postinjection, but the differences were not significant (P>.05). On the other hand, central administration of chicken motilin (0.2 and 0.4 microg) showed a tendency to suppress feeding of fasted chicks as well as the result of high dose (5.0 microg) under ad libitum feeding conditions. Therefore, the results presented here suggest that central motilin alone does not induce hyperphagia in meat-type chicks. PMID- 15276782 TI - Behavioral and physiological adaptation to repeated chair restraint in rhesus macaques. AB - Physical restraint is a commonly used procedure when working closely with nonhuman primates. Nonhuman primates show rapid behavioral changes when learning the restraint procedure, and these changes have been taken to reflect behavioral and physiological habituation to the procedure. This study examined the behavioral and adrenocortical responses to repeated physical restraint in a large sample of adult male rhesus monkeys. Subjects showed a decline in behavioral agitation and cortisol concentrations across seven consecutive days of restraint. The changes in adrenocortical responsiveness were also coincident with an increased sensitivity to dexamethasone and a change in early morning basal cortisol secretion. The subjects were restrained for a single session 6 months later, and while the reduction in behavioral agitation was still present, the majority of changes in adrenocortical responsiveness were no longer present. These data show that behavior is not necessarily an indicator of underlying physiological processes and that the reduction of hypothalamic-pituitary-adrenal (HPA) activity with repeated restraint is due to physiological adaptation to high glucocorticoid concentrations and not to psychological habituation to the restraint procedures. PMID- 15276783 TI - The anxiogenic video-recorded Stroop Color-Word Test: psychological and physiological alterations and effects of diazepam. AB - From among the few human experimental models that can be used to predict the clinical activity of new anxiolytic drugs, the video-recorded Stroop Color-Word Test (VRSCWT), which uses subjective scales to evaluate anxious states, is notable for its simplicity. However, considering that the choice of treatment for anxiety disorders is heavily dependent on the level of somatic symptomatology, a quantitative evaluation of the physiological alterations elicited by the anxiogenic situation of the VRSCWT would also be of great interest. In the present study, 36 healthy male and female volunteers were submitted to either the VRSCWT or to a nonanxiogenic test. The results showed that, as well as a sensation of anxiety, the VRSCWT elicited increases in heart rate and gastrocnemius tension. Subsequently, a further 48 healthy men and women were randomly assigned to three treatments: placebo, 5 and 10 mg of diazepam, and were submitted to the VRSCWT. The results showed that in men, diazepam blocked the feeling of anxiety elicited by the test, although it did not prevent the physiological alterations, while in women, there was no response to the anxiolytic action of the drug. Taken as a whole, these results suggest that the VRSCWT is an efficient method of inducing anxiety experimentally. It is able to elicit observable psychological and physiological alterations and can detect the blocking, by an anxiolytic, of the feelings of anxiety in healthy men. Furthermore, the results suggest that the neural pathways for the control of the psychological and physiological manifestations of anxiety may be separate. This study also draws attention to the fact that gender is an important variable in the evaluation of anxiolytic drugs. PMID- 15276784 TI - Ethanol intake and motor sensitization: the role of brain catalase activity in mice with different genotypes. AB - The C57BL/6J strain of inbred mice shows a characteristic pattern of ethanol induced behaviors: very weak acute locomotor stimulation, a lack of locomotor sensitizing effect of ethanol, and a high level of ethanol intake. This strain has relatively low levels of activity of the ethanol metabolizing enzyme catalase, and it has been proposed that brain catalase plays a role in the modulation of some behavioral effects of ethanol. In the first study of the present paper, we investigated the effects of pharmacological manipulations of brain catalase activity on C57BL/6J mice in acute ethanol-induced locomotion and ethanol intake. Results indicated that the reduction in motor activity produced by ethanol was reversed by pretreatment with catalase potentiators and it was enhanced by catalase inhibitors. In addition, ethanol intake was highly correlated with brain catalase activity in mice treated with a catalase potentiator. In the second study, F1 hybrid mice (SWXB6) from the outbred Swiss Webster mice and the inbred C57BL/6J mice were used. Basal brain catalase activity levels of F1 mice were intermediate between to those of the two progenitor genotypes. That profile of catalase activity was parallel to the acute ethanol-induced locomotion and to repeated-ethanol-induced motor sensitization effects observed across the three types of mice. These data suggest that brain catalase activity modifications in the C57BL/6J strain change the pattern of several ethanol-related behaviors in this inbred mouse. PMID- 15276785 TI - Hindbrain catecholamine neurons mediate consummatory responses to glucoprivation. AB - Previous work using the retrogradely transported immunotoxin, saporin (SAP) conjugated to a monoclonal antibody against dopamine-beta-hydroxylase (DBH; DSAP), to selectively lesion norepinephrine (NE) and epinephrine (E) neurons projecting to the medial hypothalamus, demonstrated the essential role of these neurons for appetitive ingestive responses to glucoprivation. Here, we again utilized this lesion to assess the importance of these same neurons for the consummatory phase of glucoprivic feeding. To test consummatory responses, milk was infused intraorally through a chronic cheek fistula until rejected. Appetitive responses were tested in the same rats using pelleted food. Feeding responses to insulin-induced hypoglycemia, 2-deoxy-D-glucose (2DG)-induced blockade of glucose utilization, mercaptoacetate (MA)-induced blockade of fatty acid oxidation, 0.9% saline, and 18-h food deprivation were assessed. Unlike unconjugated SAP controls, the DSAP rats did not increase their food intake in response to glucoprivic challenges in either the pelleted food or the intraoral feeding tests. However, the DSAP rats did not differ from SAPs in their ingestive responses to food deprivation and blockade of fatty acid oxidation. The selective impairment of glucoprivic feeding responses indicates that DSAP did not impair the underlying circuitry required for either appetitive or consummatory ingestive responding but eliminated the mechanism for control of this circuitry specifically by glucoprivation. Results suggest that both appetitive and consummatory responses to glucoprivation are controlled and coordinated by multilevel terminations of the same catecholamine neurons. PMID- 15276786 TI - Estradiol-mediated increases in the anorexia induced by intraperitoneal injection of bacterial lipopolysaccharide in female rats. AB - Lipopolysaccharide (LPS) derived from the cell walls of gram-negative bacteria causes a robust acute phase response (APR) that includes fever, anorexia, and many other elements. Because immune system function, including some models of illness anorexia, is sexually differentiated, we investigated the sexual differentiation of the anorexia induced by intraperitoneal LPS injections in rats. Cycling female Long-Evans rats tested either during diestrus or estrus ate less following 6.25 microg/kg LPS than did intact males. Following 12.5 microg/kg LPS, females in estrus ate less than either females during diestrus or males. Similarly, a more pronounced anorexia occurred following 12.5, 25, and 50 microg/kg LPS in ovariectomized females that received cyclic estradiol treatment and were tested on the day modeling estrus than in untreated ovariectomized rats. LPS also increased the length of the rats' ovarian cycles, usually by a day, especially when injected during diestrus. As in male rats, when LPS injections were repeated in the same rats, both estradiol-treated and untreated rats failed to display any significant anorexia. The inhibitory effects of LPS on eating in intact and ovariectomized rats were expressed solely as decreases in spontaneous meal frequency, without significant alteration of spontaneous meal size. These data indicate that anorexia following peripheral LPS administration is sexually differentiated and that estradiol is sufficient to produce this response. The mechanism of the pathophysiological effect of estradiol on meal frequency appears to be different from the physiological effect of estradiol on food intake because the latter is expressed solely as a change in meal size. PMID- 15276787 TI - Subdiaphragmatic vagal deafferentation fails to block the anorectic effect of hydroxycitrate. AB - We investigated the neural mediation of the feeding suppression through orally administered hydroxycitrate (HCA) in male rats that were fed a high-glucose diet (about 48% glucose). Ten-day ad libitum food intake and body weight regain after previous body weight loss (13% of initial body weight) due to restrictive feeding were measured in rats with sham deafferentation (SHAM; n = 6), subdiaphragmatic vagal deafferentation (SDA; n = 7), and SDA plus celiac-superior mesenteric ganglionectomy (SDA/CGX; n = 9). HCA suppressed the 10-day cumulative food intake in all surgical groups and body weight regain in SDA and SDA/CGX groups. Independent of HCA, SDA and SDA/CGX rats consumed less food and gained less weight compared to SHAM rats. These results demonstrate that all vagal afferents from below the diaphragm and vagal efferents of the dorsal trunk as well as splanchnic nerves (afferents and efferents) are not necessary for the feeding suppressive effect of HCA in this animal model. Vagal afferents, however, appear to play a role in the control of intake when a high-glucose diet is consumed after a period of restrictive feeding. PMID- 15276788 TI - Modifications of a field method for fecal steroid analysis in baboons. AB - By extracting steroid metabolites from feces, researchers can track endocrine activity noninvasively in free-ranging animals. Sample preservation is a critical component of such methods because steroid metabolites rapidly decompose. Here, we describe a method for preservation, field extraction, and radioimmunoassay of steroid metabolites (estradiol, progesterone, glucocorticoids, and testosterone) from the feces of wild female baboons (Papio spp.). This method is a modification of that developed by Stavisky [Socioendocrinology: noninvasive techniques for monitoring reproductive function in captive and free-ranging primates. PhD, Emory University, 1994.], which employs reversed-phase octadecylsilane cartridges to extract steroids from feces. In addition to providing physiological validation for this method, we examine variation in steroid concentration across different (1) collection times (morning vs. afternoon), (2) methanol extraction treatments (homogenized vs. hand-mixed), and (3) solid-phase extraction times (2 vs. 10 h after collection). We then examine the stability of sample storage at ambient and subzero temperatures to determine whether storage time significantly alters steroid concentrations. Our results show that hormone concentrations do not differ between morning and afternoon samples, homogenization yields significantly higher fecal steroid concentrations, and fecal steroids are stable in a methanol/acetone solution for up to 10 h. When stored at ambient temperatures, only glucocorticoid metabolites had some degradation over a period of up to 40 days. However, when stored at -10 degrees C, no significant steroid changes were observed for up to 400 days. This method is particularly suited for behavioral research because it permits delays between sample collection and sample processing, thus allowing behavioral observations to continue. PMID- 15276789 TI - A simple method for short-term controlled anesthesia in newborn mice. AB - In this study, we describe a simple and inexpensive method for inducing short term anesthesia and rapid recovery in newborn mice. Litters of Swiss mice pups were randomly allocated to testing on postnatal days 2, 5, and 8. Anesthesia was induced by placing the pup in a syringe and adding a volume of isoflurane saturated gas that produced an estimated level of 32% isoflurane. Exposure to isoflurane lasted 30 s. All the pups survived the anesthesia. At all study ages, this method abolished the nociceptive response to tail clamp without inducing mortality, thus showing effective anesthesia. Recovery from anesthesia was assessed immediately after isoflurane exposure, based on two nonnoxious behavioral tests: the defensive response to a drop of water (10 tests, 1 min apart) and 10 min later the righting reflex, i.e., the time to recovery of the prone position (five tests, 10 min apart). The water drop test scores increased during the recovery phase toward the control values in all age groups. Treatment and time had no significant effect on righting reflex scores. The initial volume in the syringe, the volume of added isoflurane-saturated gas, and the duration of exposure may be adjusted according to postnatal age and specific strains or species (e.g., rats). This method is well suited to behavioral or physiological phenotype studies in developing mice, in which noxious procedures must precede functional testing, making rapid recovery from anesthesia a key requirement. PMID- 15276790 TI - Daily activity patterns of a nocturnal and a diurnal rodent in a seminatural environment. AB - The entrainment of circadian rhythms by light-dark (LD) cycles has been extensively investigated in laboratory studies. In almost all of these studies, organisms have not been allowed to modulate their exposure to the LD cycle. In the present study, the rhythm of running-wheel activity was investigated in nocturnal (domestic mice) and diurnal (Nile grass rats) rodents provided with light-tight nest boxes and maintained under long and short photoperiods. Photoperiod length had a significant effect on the duration of the daily active phase (alpha), on the phase angle of entrainment (psi), and on diurnality or nocturnality in both species. The availability of a nest box had a modest effect only on the variability of activity onsets. Neither in the nocturnal nor in the diurnal species was there any evidence of entrainment by frequency demultiplication or of entrainment without photic stimulation at either dawn or dusk. These results indicate that at least in the species studied, the ability of rodents to modulate their exposure to the LD cycle does not have a major effect on photic entrainment. PMID- 15276791 TI - Cardiovascular effects of morning nutrition in preadolescents. AB - The cardiovascular response to eating has been extensively investigated in adults, but comparable data in children are lacking. In this investigation, heart rate and heart-rate variability were evaluated in preadolescents during resting periods in the morning initially while participants maintained overnight fasting, and again after the participants either ate a standardized breakfast or continued fasting. Relative to the initial fasting period, heart rate (HR) increased slightly in fed participants and decreased significantly in those who continued to fast. These effects were associated with significant increases in low- (LF: 0.04-0.15 Hz; primarily sympathetic influences) and high-frequency (HF:0.15-0.5 Hz; parasympathetic influences) spectral components in fasting participants and with nonsignificant decreases in both components in fed participants. Although these HF changes are consistent with the observed heart-rate variations (i.e., increases and decreases in parasympathetic influence associated with decreased and increased HR, respectively), the LF increase with the slowing, and decrease with the acceleration of HR run counter to expected sympathetic effects on HR. The net effect of these modulations was unchanged sympathovagal balance (LF/HF) for fasting participants but a significant decrease for fed participants across recording periods. The results indicate that the continuation of overnight fasting is associated with a significant increase in parasympathetic activity that is attenuated by eating breakfast. Furthermore, the findings suggest that the parasympathetic contribution to the LF spectral component is significantly enhanced in preadolescents, and, consequently, the LF/HF ratio-generally considered to reflect sympathovagal balance-does not segregate sympathetic and parasympathetic influences in children. PMID- 15276792 TI - Taste avoidance induced by wheel running: effects of backward pairings and robustness of conditioned taste aversion. AB - Rats repeatedly exposed to a distinctive novel solution (conditioned stimulus, CS) followed by the opportunity to run in a wheel subsequently drink less of this solution. Investigations on this phenomenon indicate that wheel running is an effective unconditioned stimulus (US) for establishing conditioned taste aversion (CTA) when using a forward conditioning procedure (i.e., the US-wheel running follows the CS-taste). However, other studies show that wheel running produces reliable preference for a distinctive place when pairings are backward (i.e., the CS-location follows the US-wheel running). One possibility to account for these results is that rewarding aftereffects of wheel running conditioned preference to the CS. The main objective of the present study was to assess the effects of backward conditioning using wheel running as the US and a distinctive taste as the CS. In a between-groups design, two experimental groups [i.e., forward (FC) and backward conditioning (BC)] and two control groups [CS-taste alone (TA) and CS-US unpaired (UNP)] were compared. Results from this experiment indicated that there is less suppression of drinking when a CS-taste followed a bout of wheel running. In fact, rats in the BC group drank more of the paired solution than all the other groups. PMID- 15276793 TI - Dose-dependent cocaine place conditioning and D1 dopamine antagonist effects in male Japanese quail. AB - The dopamine D1 receptor subtype has been implicated in drug reward processes in mammals. Two experiments investigated whether dose-dependent differences in cocaine conditioned place preference (CPP) would be obtained in an avian species and whether these cocaine effects were mediated by the dopamine D1 antagonist R(+/-)-SCH23390. In Experiment 1, male birds were given intraperitoneal injections of 1, 3, 10, or 30 mg/kg of cocaine hydrochloride, paired with a chamber that contained distinct visual cues. On alternate days, they received saline paired with a chamber containing different visual cues. A CPP test was given after four pairings of cocaine with the distinct chamber. In Experiment 2, 0.0025, 0.025, or 0.25 mg/kg of SCH 23390 or saline was administered 15 min prior to cocaine (3 mg/kg) and placement into the least preferred chamber. CPP was observed at 1, 3, and 10 mg/kg doses of cocaine but not at 30 mg/kg or saline. All doses of SCH 23390 attenuated cocaine-induced CPP. The findings suggest that cocaine administration results in a dose-dependent CPP, and that similar with mammals, it may be mediated by D1 receptors in an avian species. Thus, the avian species may be a beneficial comparative model for drug studies, especially those involving visual cue mechanisms of drug reward. PMID- 15276794 TI - How habitual caffeine consumption and dose influence flavour preference conditioning with caffeine. AB - This study investigated the effects of both habitual caffeine use and dose administered in determining the ability of caffeine to reinforce conditioned changes in flavour preference. Thirty overnight-withdrawn moderate caffeine consumers and 30 non or low-dose caffeine (non/low) consumers evaluated five novel-flavoured fruit teas. Subsequently, their median-rated tea was used in four ensuing conditioning sessions. Either placebo, 1 or 2 mg/kg of caffeine (n=10 consumers, 10 non/low consumers in each condition), was added to the target tea, and all five teas were reevaluated at a final tasting. Pleasantness ratings over the four conditioning sessions indicated that non/low consumers' liking increased for the noncaffeinated fruit tea with no change for the tea containing either 1 or 2 mg/kg of caffeine. Among consumers, pleasantness ratings tended to decrease for the noncaffeinated fruit tea but increased significantly at the 1-mg dose and showed a tendency to increase at the 2-mg dose. Similar effects were shown in the evaluations made before and after conditioning, with no change in the nonexposed drinks. These results show that 1.0 mg/kg of caffeine reinforces changes in flavour pleasantness in acutely withdrawn habitual consumers but not in nonconsumers or nondependent low-caffeine consumers, further endorsing the negative-reinforcement theory of conditioning with caffeine. PMID- 15276795 TI - Odor discrimination assessment with an automated olfactometric method in a prosimian primate, Microcebus murinus. AB - The present study was aimed at adapting an automated olfactometer designed for use with rodents to a nocturnal lemur Microcebus murinus. This apparatus allows rigorous control of odor stimuli. We show that M. murinus could remain quiet and attentive for about 20 min in the test chamber, allowing daily sessions of 40 consecutive trials. This allowed us to train M. murinus subjects to learn a go/no go discrimination procedure using fruity-odor cues. Each of seven subjects reached or exceeded a criterion of 33 correct responses in a block of 40 trials in a range of 4-14 training sessions. When trained on an odor reversal task, performance initially dropped sharply, followed by rapid acquisition of the new task. These outcomes demonstrate that, like rodents, M. murinus can be trained using operant conditioning in an automated olfactometer. This species should prove useful for investigating cognitive capacities and neurodegenerative disease in a primitive primate model. PMID- 15276796 TI - Licking rate adaptations to increased mandibular weight in the adult rat. AB - In this experiment, chronic mandibular loading was used to study adaptation in licking rate. Twenty-four 115-day-old rats were randomly divided into two groups. The experimental group received 1.791 +/- 0.083 g submandibular gold implants, and the control group received 0.179 +/- 0.009 g submandibular acrylic implants. The animals were videotaped while lapping on two separate occasions preoperatively, and once every week postoperatively for 12 weeks. The videotapes were used to obtain licking rates for each animal at each taping session. The findings showed that licking rate decreased significantly after surgery for both groups; however, the decrease was similar for both the experimental and control groups. This indicates that licking rate was affected by the experimental design, but not specifically by the weight of the gold implant. PMID- 15276797 TI - Blunted cortisol rhythm is associated with learning impairment in aged hamsters. AB - In mammals, the cognitive decline that accompanies unsuccessful aging is associated with circadian rhythm dysfunction and increased levels of circulating glucocorticoids. The possible causal relations among these factors are not known. To test for primary effects of circadian clock dysfunction versus increased glucocorticoid levels as contributors to age-related learning impairment, we measured cortisol and wheel-running rhythms along with context learning in aged hamsters (15-18 months). At this age, locomotor rhythms of learners and nonlearners were found to be indistinguishable. However, plasma cortisol levels were lower, and the amplitude of the cortisol rhythm was reduced in the impaired animals. These data suggest that age-related cognitive decline may be related more to a loss of hormone rhythmicity than to a loss of behavioral rhythmicity or an increase in hormone level. PMID- 15276798 TI - Effects of nonsaponin fraction of red ginseng on learning deficits in aged rats. AB - Previously we reported that oral application of red ginseng significantly ameliorated learning deficits in aged rats and young rats with hippocampal lesions. In the present study, we investigated the effects of the nonsaponin fraction of red ginseng on learning deficits in aged rats in behavioral studies and those on long-term potentiation (LTP) in the hippocampal CA3 subfield in young rats in electrophysiological studies. In the behavioral studies, three groups of rats [aged rats with and without oral administration of the nonsaponin fraction of red ginseng and young rats] were tested with the three types of spatial-learning task [distance movement task (DMT), random-reward place search task (RRPST), and place-learning task (PLT)] in a circular open field. The results in the DMT and RRPST indicated that motivational and motor activity was not significantly different among the three groups of rats. However, performance of the aged rats without nonsaponin was significantly impaired in the PLT when compared with the young rats. Treatment with nonsaponin significantly ameliorated deficits in place-navigation learning in the aged rats in the PLT. In the electrophysiological studies, effects of nonsaponin on the LTP in the CA3 subfield of the hippocampal slices were investigated in vitro. Pretreatment with nonsaponin significantly augmented the increase in population spike amplitudes in the CA3 subfield after LTP induction. These results suggest that the nonsaponin fraction of red ginseng contains important substances to improve learning and memory in aged rats and that this amelioration by nonsaponin might be attributed partly to augmentation of LTP in the CA3 subfield. PMID- 15276799 TI - Anxiety and aggression associated with the fermentation of carbohydrates in the hindgut of rats. AB - Lactic acid accumulation in the caecum and colon resulting from the fermentation of carbohydrates can lead to deleterious effects in ruminant and monogastric animals, including humans. In the present study, we examined the behavioural effects of two types of commonly consumed foods: soluble and fermentable carbohydrates (FCs). Thirty-six male Wistar rats were fed either a commercial rat and mouse chow, a soluble carbohydrate (SC)-based diet or an FC-based diet. Social interaction, anxiety, aggression and locomotor activity were examined by employing a social interaction test and a light/dark emergence test, while physical parameters of hindgut fermentation were examined after sacrifice, either 3 or 21 h after feeding. Results showed that anxiety (spending less time in the light compartment during the light/dark emergence test) and aggression (increased fighting during the social interaction test) were increased following raised concentrations of fermentation end products, such as lactic acid and volatile fatty acids (VFAs) in the caecum of rats. These associations occurred regardless of dopamine and 5-HT concentrations in the prefrontal cortex (PFC) and provide evidence supporting a general effect of FCs on behaviour. Possible mechanisms of action along with similarities between a rat and human model of acidosis are discussed. PMID- 15276800 TI - Evaluation of functional asymmetry in rats with dose-dependent lesions of dopaminergic nigrostriatal system using elevated body swing test. AB - Although drug-induced rotational behavior has conventionally been used for the assessment of functional asymmetry in 6-hydroxydopamine (OHDA)-lesioned rats, a pure behavioral test that can evaluate animals with dose-dependent lesions of the dopaminergic nigrostriatal system in a drug-free state may better reflect a more natural response following lesion. In this study, elevated body swing test (EBST) was used for evaluation of rats with varying lesions of the dopaminergic nigrostriatal system. For this purpose, rats received intrastriatal injection of 8 (L1), 12 (L2), 16 (L3), and 20 (L4) microg of the neurotoxin 6-OHDA (dissolved in 5 microl of saline-ascorbate). Apomorphine-induced rotational and drug-free elevated body swing behaviors were evaluated before and at different time points after the experiment. The results showed that although there is a significant trend for contralateral rotations and drug-free swings in lesioned rats as compared to sham-operated group, however, there were no significant differences between the L1 and L2 and between the L3 and L4 groups regarding EBST. Therefore, EBST may be valuable for behavioral analysis of rats with mild and/or severe damage of the dopaminergic nigrostriatal system. PMID- 15276801 TI - Differential effects on somatic and reflex development by chronic clomipramine treatment. AB - The developmental effect of altered 5-HT and NE levels is a subject that requires more attention, especially when considering the increased demand for antidepressive dual reuptake inhibitors. Serotonin and norepinephrine are bioamines that differentially influence the nervous tissue growth. This study investigated the somatic maturation and the ontogeny of reflexes in neonate rats treated from the 1st to the 21st postnatal day (PND) with clomipramine (20 mg/kg sc, daily), a potent monoamine reuptake inhibitor. Indicators of both general body growth (body weight, axis of the head and body lengths) and physical maturation (ear unfolding, auditory conduit opening, eruption of the lower incisors and eye opening) were appraised. Ontogeny of motor and sensory reflexes (righting, free-fall righting--acceleration, negative geotaxis, cliff avoidance, auditory startle response and vibrissa placing) was also observed. The results demonstrated that chronic neonatal treatment with clomipramine alters the somatic growth. However, it did not interfere with the onset time of many physical features and reflexes. These results provide insights into the consequences of dual transmitter during early development. PMID- 15276802 TI - Gait dynamics in trisomic mice: quantitative neurological traits of Down syndrome. AB - The segmentally trisomic mouse Ts65Dn is a model of Down syndrome (DS). Gait abnormalities are almost universal in persons with DS. We applied a noninvasive imaging method to quantitatively compare the gait dynamics of Ts65Dn mice (n=10) to their euploid littermates (controls) (n=10). The braking duration of the hind limbs in Ts65Dn mice was prolonged compared to that in control mice (60+/-3 ms vs. 49+/-2 ms, P<.05) at a slow walking speed (18 cm/s). Stride length and stride frequency of forelimbs and hind limbs were comparable between Ts65Dn mice and control mice. Stride dynamics were significantly different in Ts65Dn mice at a faster walking speed (36 cm/s). Stride length was shorter in Ts65Dn mice (5.9+/ 0.1 vs. 6.3+/-0.3 cm, P<.05), and stride frequency was higher in Ts65Dn compared to control mice (5.9+/-0.1 vs. 5.3+/-0.1 strides/s, P<.05). Hind limb swing duration was prolonged in Ts65Dn mice compared to control mice (93+/-3 vs. 76+/-3 ms, P<.05). Propulsion of the forelimbs contributed to a significantly larger percentage of stride duration in Ts65Dn mice than in control mice at the faster walking speed. Indices of gait dynamics in Ts65Dn mice correspond to previously reported findings in children with DS. The methods used in the present study provide quantitative markers for genotype and phenotype relationship studies in DS. This technique may provide opportunities for testing the efficacy of therapies for motor dysfunction in persons with DS. PMID- 15276803 TI - Dissociation of ethanol and saccharin preference in fosB knockout mice. AB - The Fos family of transcription factors may play a key role in various forms of brain plasticity. Among different genes coding Fos proteins is the fosB gene. Protein products of the fosB gene are thought to be critically involved in neural adaptations produced by chronic treatment with drugs of abuse. fosB gene transcription leads to accumulation of full-length FosB as well as its truncated form, deltafosB. Stable isoforms of deltafosB called chronic FRAs accumulate in the brain after chronic administration of various drugs of abuse. The purpose of the present study was to evaluate the role of the fosB gene in two-bottle choice ethanol self-administration. For this aim, ethanol (2-8% v/v) intake and preference was assessed in fosB mutant (n=17) and wild-type (WT) mice (n=16). For comparison, consumption of saccharin (0.05-0.8% w/v) and quinine (15-960 microM) solutions was assessed in the same animals. Ethanol preference in both groups varied from around 50% for the lowest to 20% for the highest ethanol concentration. Neither ethanol intake (g/kg) nor preference differed between the two genotypes. In contrast, saccharin preference, but not intake, was higher in the fosB mutants. Only slight and inconsistent between-group differences were observed in terms of quinine preference. The present results suggest that permanent elimination of fosB gene products does not alter ethanol intake but may enhance preference for sweet solutions in mice. PMID- 15276804 TI - Amount of ingested custard dessert as affected by its color, odor, and texture. AB - The effects of nonoral sensations, such as visual texture and odor, on the size of the first bite were investigated in a series of studies using specially constructed food delivery cups with lower, from which custards were ingested ("ingested custard"), and upper, from which a custard was viewed and/or smelled ("upper custard") compartments. Ingested and upper custards were either the same or different. Bite size was defined as the weight of custard sucked out of the lower compartment during a single suck through an 11-mm diameter straw. The results from the first study indicated that the recognition of oral qualities of custards via vision or olfaction determined the size of the first bite. When this recognition was favorable, e.g., when the upper custard was known to be creamy, a relatively large bite was taken, irrespective of the custard that was actually ingested. When this recognition was unfavorable, a relatively small bite was taken. The second study showed that when recognition was prevented by modifications of the upper custard's color, odor, or visual texture, bite size was determined by the oral qualities of the ingested custard. This was confirmed in a third study, where the oral characteristics of the ingested custard were varied by adding a flavorant (benzaldehyde) and/or by using nose clips to eliminate retronasal smelling. Bite sizes decreased significantly when these variations reduced creaminess. Odor and visual texture characteristics of the upper custard significantly affected the perception of creaminess and other attributes related to the food's viscosity, melting, and thickness. PMID- 15276805 TI - Increased grooming behavior in mice lacking vitamin D receptors. AB - Vitamin D is a neuroactive secosteroid with several important functions in the nervous system. Many human and animal findings link alterations in the vitamin D system to various neurological and behavioral disorders. Since grooming is an important element of animal behavior, here we studied whether genetic ablation of vitamin D receptors (VDR) in mice may be associated with altered grooming behaviors. Overall, VDR knockout (VDRko) mice presented longer duration and higher frequency of grooming when tested in the actimeter, open field, elevated plus maze, and horizontal rod tests. Increased grooming did not, however, correlate with unaltered general activity level (actimeter test), anxiety-like behaviors (hole board and elevated plus maze tests), and emotional reactivity index (defecation boli). In general, our results confirm the role of vitamin D and VDR in the regulation of behavior, including grooming, and suggest that increased grooming behavioral phenotype may be associated with genetic ablation of VDR in mutant mice. PMID- 15276806 TI - Acid taste thresholds assessed by conditioned taste aversion and two-bottle preference in rats. AB - The conditioned taste aversion (CTA) threshold for either citric acid (CA) or HCl solutions and the two-bottle taste preference (TBP) threshold were determined in rats that are familiarized to the odor of conditioning solutions or that are naive. The CTA method appeared to be more sensitive than the TBP test, particularly when rats were not familiarized to the odor of the conditioning solution. The CTA threshold for HCl-conditioned rats and familiarized to the odor of conditioning solution lies between 1.00 and 2.00 mmol; in unconditioned rats, it lies between 4.00 and 5.00 mmol. In CA-conditioned and odor-familiarized rats, the threshold lies between 0.09 and 0.20 mmol; in unconditioned rats, it lies between 7.00 and 10.00 mmol. In rats not familiarized to the odor of the conditioning solution, the threshold for HCl-conditioned rats lies between 0.90 and 1.00 mmol; in unconditioned rats, it lies between 2.00 and 3.00 mmol. In CA conditioned rats, the CTA threshold lies between 0.03 and 0.05 mmol; in unconditioned rats, it lies between 4.00 and 7.00 mmol. The two-bottle test is less sensitive than the CTA method. The TBP threshold lies between HCl 4.00 and 5.00 mmol, and between CA 4.00 and 7.00 mmol. The odor of a solution may potentiate the ability of rats to detect the concentration of CA and HCl solutions. PMID- 15276807 TI - Age at group formation alters behavior and physiology in male but not female CD-1 mice. AB - In the laboratory environment, rodents are usually housed in unisexual groups, which are assembled after weaning. Housing of unfamiliar subjects has been described, however, as a stressful social setting for rodents and other mammals. Aim of the present study was to evaluate whether the age at which house mice are grouped might affect their behavior and physiology. Male or female unisexual groups were formed at different ages: at weaning, i.e., before puberty (JUV); at adolescence, i.e., after puberty (AD); and controls were raised with siblings since birth (CON). Results show that age at group formation induced several behavioral and physiological alterations in males but not in females. Specifically, when compared to controls, JUV males showed higher aggression, smaller preputial gland, and a marked reduction of neophobia in the free exploratory paradigm. Fewer changes occurred in the AD males, which showed reduced neophobia in the free exploratory paradigm and, when adults, a reduction in body weight. Females were not affected by the experimental treatment. Surprisingly, the basal corticosterone assessed at the nadir was lower for both males and females JUV and AD respect to CON. In conclusion, it is clear that mixing groups at different ages has profound effects on mouse behavior and physiology. PMID- 15276808 TI - Associations between taste genetics, oral sensation and alcohol intake. AB - Alcohol produces a range of oral sensations, some of which have been shown to vary with the perceived bitterness of 6-n-propylthiouracil (PROP), one marker for genetic variation in taste. Some studies report that offspring of alcoholics are most likely to be PROP nontasters [Physiol. Behav. 51 (1992) 1261; Physiol. Behav. 64 (1998) 147], yet others report the offspring as more responsive to sodium chloride (NaCl) and citric acid, which appears to contradict the taste genetic hypothesis. We predicted alcohol sensation and intake from measures of taste genetics (PROP bitterness and number of fungiform papilla), NaCl and citric acid intensity, and spatial taste pattern in 40 females and 43 males. Subjects used the general Labeled Magnitude Scale (gLMS) [Chem. Senses 18 (1993) 683; J. Food Qual. Pref. 14 (2002) 125] as an intensity and hedonic scale. Those who tasted PROP as most bitter or had highest numbers of fungiform papilla reported greatest oral burn from an alcohol probe; those who tasted least PROP bitterness consumed alcoholic beverages most frequently. Although higher NaCl and citric acid ratings associated with more frequent consumption of alcoholic beverages, the findings could be explained by lower intensity of tastants on the tongue tip (chorda tympani nerve) relative to whole mouth perception. In multiple regression analyses, PROP bitterness and the spatial pattern of taste perception were independent contributors to the prediction of alcohol intake. In summary, the results support that variation in oral sensation associates with alcohol intake. Those who taste PROP as least bitter and have low chorda tympani relative to whole mouth taste intensity appear to have fewest oral sensory hindrances to the consumption of alcoholic beverages. PMID- 15276809 TI - Influence of pellet size on rat's hoarding behavior. AB - The body weight threshold for hoarding behavior of rats is routinely used as a means to discern an animal's body weight regulation. We explored whether the size of food pellets would modify hoarding and the hoarding threshold. In Experiment 1, we offered the rats either large (ca. 5 g) or small (ca. 2 g) food pellets on alternate days while keeping their body weights within a narrow range when they were not in the hoarding sessions. The hoarding threshold was not influenced, by food size (312+/-32 g small and 298+/-13 g large pellets, N.S.). On the other hand, the relationship between hoarded food and body mass significantly differed between small and large pellets (ratio of 2.7). Because such a ratio was similar to that of the respective pellet weights, this suggests that the more the rat is deprived of food, the more willing it is, in a predetermined manner, to move about in search of food. Experiment 2 verified this hypothesis: instead of weighing the food hoarded, we counted the number of pellets hoarded. The slopes of the regression lines were similar in both cases, when the pellets were counted and when the hoarded food was weighed. Results showed no significant differences between these two approaches, suggesting that the weight of hoarded food is a good indicator of the number of trips from home to food in the hoarding experiment. PMID- 15276810 TI - Food restriction in pregnant rat-like hamsters (Cricetulus triton) affects endocrine, immune function and odor attractiveness of male offspring. AB - We studied the effect of intrauterine food restriction (FR) on the immune function, endocrine status and attractiveness of scents of male rat-like hamsters, Cricetulus triton. Work was conducted on field-caught parents from the North China plain and their laboratory-born progeny. Restricted pregnant dams were fed 70% of the mean daily intake of hamsters with free access to food. FR caused a marked and protracted weight reduction of the body, adrenal, testes and epididymides in males. During the refeeding period, the spleen and thymus, but not the adrenal weight of the malnourished offspring caught up with that of the control after about 60 days. The present results demonstrated that estrous females preferred the odors of control males to that of FR males. Males whose mothers were food restricted during gestation had lower testosterone concentrations, immune responses and reproductive organ mass but had higher circulating cortisol than did the males in the control group. Thus, the effect of maternal FR may be an important cause in population regulation in the rat-like hamster. The testosterone level was positively correlated with immune function in rat-like hamsters, but the lower immunity was not suppressed by higher level of testosterone, as previously suggested. We also found a negative relationship between cortisol and immune function in the rat-like hamster. PMID- 15276811 TI - Responses of PROP taster groups to variations in sensory qualities within foods and beverages. AB - Despite increasing evidence that variations in sensitivity to the bitterness of 6 n-propylthiouracil (PROP) are also reflected in responses to other tastes in solution, there has been little research examining the impact of PROP sensitivity on responses to sensory qualities in foods or beverages. The present studies examined responses of PROP taster groups to systematic variations in tastes and oral irritation in different foods and beverages. In Experiment 1, PROP groups were asked to discriminate variations in bitterness, sweetness, or sourness within two foods (yogurt and cream cheese) and a beverage (orange juice). In most cases, tasters and especially supertasters (STs) were able to discriminate smaller variations in tastant concentration than PROP nontasters (NTs). Differences were most evident with variations in bitterness and sourness. In Experiment 2, PROP taster groups rated the sweetness, sourness, and oral irritation in carbonated fruit drinks that systematically varied in citric acid (CA) and CO2 concentrations. Ratings of sourness and irritation were highest for STs and lowest for NTs, although there were no group differences for sweetness ratings. These data are some of the first to show PROP taster group differences in tastes and irritation within foods and provide a basis for reported differences of PROP groups in their hedonic responses to foods. PMID- 15276812 TI - The role of CCK2 receptors in energy homeostasis: insights from the CCK2 receptor deficient mouse. AB - The present study explored the contribution of type 2 cholecystokinin (CCK) receptors in energy regulation. A total of 78 CCK2 receptor-deficient mice and 80 wild-type controls were acclimated to a 12:12 light-dark cycle at 30 +/- 1 degrees C. Using a computer-monitored biotelemetry system, circadian patterns of body temperature, food intake, and activity were monitored for 4 days. Body weight and water consumption were manually recorded during this period. Results indicate that CCK2 receptor invalidation produces elevated body temperature during both the photophase and scotophase (by 0.38 and 0.12 degrees C, respectively), increased body weight (29.3 +/- 0.2 vs. 26.8 +/- 0.2 g) and water consumption (4.1 +/- 0.1 vs. 3.2 +/- 0.1 ml), and decreased scotophase locomotor activity (WT: 7.0 +/- 0.2 vs. KO: 6.1 +/- 0.2 counts/min). These findings suggest an important role for CCK2 receptors in processes underlying energy regulation during basal and possibly pathological states. PMID- 15276813 TI - Effects of distraction and stress on delayed matching-to-place performance in aged rats. AB - The performance of male Long-Evans rats on the delayed match-to-place (DMP) version of the Morris water maze was assessed in two separate experiments; the first compared young (4 months) with middle-aged (16 months) rats, whereas the second compared middle-aged (14 months) with old (26 months) rats. Old rats continued to use a short-term memory strategy on the DMP task, but their performance on both search and recall trials was impaired relative to that of middle-aged animals. Rats of all ages habituated rapidly to visual distraction and the performance of old rats was not affected by exposure to a mild predator stress in the form of cat urine. The performance of the middle-aged rats did not differ significantly from that of young rats, even when they were challenged on recall trials by visual distraction or by exposure to predator odour. These results do not provide strong support for the prediction that visual distraction and psychological stress would interact with age in affecting spatial short-term memory in Long-Evans rats. PMID- 15276814 TI - Effect of a beta-3 agonist on food intake in two strains of rats that differ in susceptibility to obesity. AB - CONTEXT: Beta-3 agonists acutely reduce food intake, but the mechanism is not well understood. OBJECTIVE: To evaluate the effect of a beta3 agonist on food intake in two strains of rats that differ in their sensitivity to becoming obese while eating a high-fat (HF) diet. METHODS: Male Osborne-Mendel (OM) and S5B/Pl (S5B) rats were treated with a beta3-adrenergic agonist (CL 316,243) at 8 weeks of age, after an adaptation to either an HF (56% fat energy) or a low-fat (LF; 10% fat energy) diet that was equicaloric for protein (24% energy). Ad-lib-fed rats were injected intraperitoneally with CL 316,243, at doses of 0.03, 0.1, 0.3, 1.0 or 3.0 mg/kg, or with vehicle at the beginning of the dark cycle. Food intake was measured at 1, 3, 6 and 24 h after injections. RESULTS: The beta3 agonist CL 316,243 significantly decreased food intake at all timepoints in both strains of rats eating both diets. However, this inhibition of food intake was significantly greater in the S5B rat. CL 316,243 significantly decreased serum leptin and serum glucose in both the OM and the S5B rats, and again, the inhibition was greater in the S5B rat. Whereas CL 316,243 increased serum insulin levels in the OM rat, it decreased them in the S5B rat on an LF diet. In a second experiment, chow-fed rats were implanted with vascular ports into the jugular vein and allowed to recover. When CL 316,243 was injected into the animals that were fasted overnight, rats of both strains significantly increased their serum insulin at 30 min, but the increase was much more pronounced in the S5B rat. Serum glucose was decreased significantly at both the 30- and 60-min timepoints in the OM rat and at 30 min in the S5B rat. CONCLUSION: These experiments demonstrate that a beta3 agonist (CL 316,243) has a much greater effect in a strain of rats that resist fat-induced obesity. PMID- 15276815 TI - Perinatal hypothyroidism effects on step-through passive avoidance task in rats. AB - Previous studies have documented a decrease in the ability of neonatal hypothyroid animals to learn and habituate to maze tests, and an increase in spontaneous activity. However, there is little information about the effects of perinatal (i.e., prenatal and postnatal) hypothyroidism on behaviour. The present study was designed to assess whether perinatal hypothyroidism in rats induces alteration on acquisition and/or short- and long-term retention of a learned response in male Wistar rats. Perinatal hypothyroidism was induced by prolonged (E9-P21) exposure of pregnant and lactating dams to methimazole (administered orally in drinking water, 0.2 mg/ml). Cognitive function was tested at 50 days by means of a step-through passive avoidance task. The effects of perinatal hypothyroidism on the retention of the passive avoidance response are long lasting being, however, highly dependent on the retention after the original training. Our results showed that methimazole-treated rats performed more poorly when retention was tested at long-term (24 h and 7 days) retention interval. Instead, methimazole-treated rats showed longer retrieval latencies than the control ones did when retention was tested at short term (1 h). PMID- 15276816 TI - Effects of postnatal social isolation on hormonal and immune responses of pigs to an acute endotoxin challenge. AB - Social stress during early postnatal life often results in long-term effects on neuroendocrine and immune adaptation mechanisms. Therefore, the aim of the present study was to determine the influence of a 2-h daily social isolation from Day 3 to Day 11 on the acute and long-term proinflammatory and neuroendocrine responses of piglets challenged with the bacterial endotoxin lipopolysaccharide (LPS; 100 microg/kg body weight). Peripheral LPS administration significantly increased plasma concentrations of tumor necrosis factor-alpha (TNF-alpha), ACTH and cortisol in isolated and control pigs. However, the activity of the hypothalamic-pituitary-adrenal (HPA) axis after LPS stimulation was not significantly affected by isolation treatment, whereas the prior social isolation diminished the plasma TNF-alpha response to LPS 1 day as well as 45 days after the isolation period. The hippocampal TNF-alpha concentration in response to LPS was also reduced in priorly isolated pigs compared to control animals. Furthermore, the significant increase of TNF-alpha in the spleen caused by LPS was associated with a dramatic decrease in glucocorticoid receptor (GR) binding. The GR binding in hippocampus was increased in isolated pigs and was significantly decreased after LPS injection. In addition, the repeated isolation stressor was shown to increase hippocampal levels of interleukin-1beta (IL 1beta). The present results indicate that repeated social isolation of neonatal pigs may cause long-term effects on proinflammatory regulation at the periphery and in the brain following immune challenge with particular importance of TNF alpha in mediating these interactions. PMID- 15276817 TI - Effect of selection for behavior on pituitary-adrenal axis and proopiomelanocortin gene expression in silver foxes (Vulpes vulpes). AB - Silver foxes from a commercial population (farm bred or unselected for behavior control) and from populations selected for tame behavior and enhanced aggressiveness towards man have been investigated. Plasma cortisol and adrenocorticotropic hormone (ACTH) levels, pituitary ACTH levels, POMC gene expression in the anterior pituitary, and corticotropin-releasing factor (CRF) gene expression in the hypothalamus were assessed. The results indicate that the males from the tame-behavior group have lower plasma cortisol and ACTH levels and POMC gene expression in the anterior pituitary in response to capture and handling in comparison with unselected ones. Foxes from the aggressive behavior group also have lower POMC expression, although plasma cortisol and ACTH levels remain the same as in unselected ones. The three groups of animals show no significant changes in the ACTH level in the pituitary and CRF expression in the hypothalamus. PMID- 15276818 TI - The effects of neurotoxins on web-geometry and web-building behaviour in Araneus diadematus Cl. AB - The process of orb weaving and the resultant orb web constitute a good example of a complex behavioural pattern that is still governed by a relatively simple set of rules. We used the orb spider Araneus diadematus as a model organism to study the effect of the three neurotoxins (scopolamine, amphetamine, and caffeine) on the spider's behaviour. Scopolamine was given at two concentrations, with the lower one showing no effects but the higher one reducing web-building frequency; there also appeared to be a weak effect on web geometry. Amphetamine and caffeine, on the other hand, both resulted in significant changes in both building frequency and web geometry, compared to the controls. Amphetamine webs retained their size but showed an increase in spiral spacing and radius irregularity, as well as a decrease in building efficiency. Caffeine led to a general decrease in size and a slight increase in spiral spacing, as well as radius irregularity. Furthermore, caffeine caused webs to be rounder. Our observations suggest that these neurotoxins disturb different parts of the web building programme presumably by affecting different actions in the spider's CNS. PMID- 15276819 TI - Release of peppermint flavour compounds from chewing gum: effect of oral functions. AB - During chewing, the oral cavity functions like a bellow, forcing volatile flavour compounds into the exhaling air to the nasal compartment. Accordingly, we hypothesised that flavour release from chewing gum is predominantly governed by chewing frequency (CF), although other oral functions, like masseter muscle activity (MMA), chewing force (CFO), and saliva flow rate (SFR), may also play a role. In 10 healthy young males, the retronasal expired air of menthol and menthone from peppermint-flavoured (2%) chewing gum was determined as functions of CF, SFR, MMA, and CFO. The experimental setup comprised three separate series of a 4-min chewing period. These series differed only with respect to CF, i.e., habitual frequency, and 60 and 88 strokes/min. Results showed that more than 50% of the released menthol and menthone could be retrieved in the expired air and saliva. After 2-min of chewing, the concentration of flavour compounds in the expired air depended primarily on MMA and CF, becoming higher with increased MMA and CF. The concentration of flavour compounds in saliva depended primarily on SFR and the duration of the chewing task, becoming lower with high SFR and prolonged chewing duration. An increased volume of saliva in the mouth seemed to keep more flavour compounds in the aqueous phase, thereby diminishing the release via the retronasal route. In conclusion, flavour release to the retronasal compartment was dependent on MMA and CF and influenced by the volume of saliva present in the mouth. PMID- 15276820 TI - Running-wheel activity and body composition in golden hamsters (Mesocricetus auratus). AB - Running wheels are frequently used in behavioural and physiological experiments. The function of wheel-running activity in laboratory animals is controversial. In the present long-term study, the influence of this activity was evaluated in male golden hamsters over a period of 52 weeks. Four months after the start of the experiment, hamsters with access to running wheels were significantly heavier than those without these wheels. In addition, food consumption nearly doubled. The absolute values of fat-free mass (FFM), total body water (TBW) and crude fat mass (CFM) increased. However, in contrast to these absolute differences, the relative values were never different and general body composition was therefore unaffected by running-wheel activity. Different organ masses were established for absolute values of kidneys, testes and epididymis; possible effects on reproduction are discussed. The present data indicating improved physical condition leads to the assumption that a running wheel is a useful enrichment, enhancing animal welfare in the golden hamster. PMID- 15276821 TI - Seasonal changes in metabolic and temperature responses to cold air in humans. AB - The metabolic and temperature response to mild cold were investigated in summer and winter in a moderate oceanic climate. Subjects were 10 women and 10 men, aged 19-36 years and BMI 17-32 kg/m2. Metabolic rate (MR) and body temperatures were measured continuously in a climate chamber with an ambient temperature of 22 degrees C for 1 h and subsequently 3 h of 15 degrees C. The average metabolic response during cold exposure, measured as the increase in kJ/min over time, was significantly higher in winter (11.5%) compared to summer (7.0%, P < .05). The temperature response was comparable in both seasons. The metabolic response in winter was significantly related to the response in summer (r2 = .47, P < .001). Total heat production during cold exposure was inversely related to the temperature response in both seasons (summer, r2 = .39, P < .01; winter r2 = .32, P < .05). In conclusion, the observed higher metabolic response in winter compared to summer indicates cold adaptation. The magnitude of the cold response varies, but the relative contribution of metabolic and temperature response was subject specific and consistent throughout the seasons, which can have implications for energy balance and body composition. PMID- 15276822 TI - Tolerance to unfamiliar conspecifics varies with social organization in female African mole-rats. AB - Intolerance to familiar conspecifics characterises solitary mole-rats and distinguishes them from social ones. However, no study has compared the patterns of tolerance to unfamiliar conspecifics. Theoretically, both solitary and social species should react similarly and show intolerance to unfamiliar same-sex conspecifics, unless the evolution of grouping has favoured higher tolerance to conspecifics among the social species. Our study compares tolerance to unfamiliar conspecifics in four African mole-rat species exhibiting varying degrees of sociality. Dyadic encounters between female unfamiliar conspecifics were performed in a neutral arena, and the assessment of social tolerance was based on both behavioural observations of amicable contact behaviour as opposed to aggression and avoidance behaviour and the assessment of a stress response to the encounter measured as an increase in plasma cortisol concentrations. Our results show that the two highly social species and the solitary one presented similar high levels of agonistic behaviours during encounters with unfamiliar conspecifics. Nevertheless, all three social species displayed social tolerance and did not show a stress arousal during encounters with unfamiliar conspecifics, a pattern that contrasted significantly with that evidenced in the solitary species. The results suggest that physiological and behavioural characteristics allow a higher tolerance to unfamiliar conspecifics in the social as opposed to the solitary mole-rat, and the adaptive value of these characteristics are discussed. Finally, we discuss why constraints on social tolerance may be an important limiting factor to take into account in theories concerning the evolution of grouping. PMID- 15276823 TI - The chronobiology of the Natal mole-rat, Cryptomys hottentotus natalensis. AB - The Natal mole-rat, Cryptomys hottentotus natalensis, rarely, if ever, is exposed to external light cues because it occurs in completely sealed tunnel systems. As a result, their classical visual system is regressed, and therefore, their circadian system is expected proportionally to be expanded. Locomotor activity was investigated under a number of different photic regimes. Nine of the 12 mole rats exhibited endogenous circadian rhythms of locomotor activity under constant darkness, with a mean free run period of 24.13 h (range 23.93-24.13 h), with these animals entrained to a light-dark cycle (12 L:12 D). Because C. hottentotus natalensis are able to entrain their locomotor activity to an external light source, light must reach the suprachiasmatic nucleus (SCN), suggesting a functional circadian clock. A clear day-night rhythm of melatonin secretion in animals housed under a neutral photoperiod (12 L:12 D) was observed, with higher melatonin concentrations in the dark compared with the light phase. The rhythm was maintained after the animals were transferred to either continuous light (LL) or dark (DD), suggesting that the endogenous rhythm was maintained under acute exposure to light and dark. However, under DD, the rhythm appeared to shift slightly, potentially as a result of the rhythm free running. These results show that C. hottentotus natalensis has endogenous rhythms of both locomotor activity and melatonin secretion, which are modulated by light. PMID- 15276824 TI - Enhancement revisited: the effects of multiple depletions on sodium intake in rats vary with strain, substrain, and gender. AB - Repeated sodium depletion enhances spontaneous sodium intake in animals and humans, and may be a significant determinant of lifelong sodium intake. In rats, even a single sodium depletion is reported to enhance both need-induced and spontaneous sodium intakes enduringly. Both types of increases are reported to plateau after two to three depletions, and to be greater in females. Here, in two strains of rats, Wistar (W) and Sprague-Dawley (SD), and in two laboratories using different W substrains, in Israel and France, we studied the influence of repeated sodium depletions on need-induced and spontaneous sodium intake. Need induced intake in W increased incrementally, but in SD, need-induced intake occurred fully at the first depletion. In both cases, the response was not related to changes in the diuretic efficacy of furosemide. In all strains, depletions enhanced spontaneous sodium intake, but the enhancement dissipated within days following each depletion, and only attained significance over the whole 7-week experiment in male SD. Enhancement was not greater in females, despite their greater spontaneous sodium intakes. Finally, in rats given a choice of NaCl and CaCl2, depletions enhanced only NaCl intake, both need-induced and spontaneous, attesting to the specificity of the appetite in both its forms. Our findings show that enhancement of need-induced and spontaneous sodium intake resulting from repeated sodium depletions depends upon the gender and strain of rats, and it does not result from an altered diuretic response. Persistent enhancement of spontaneous sodium intake is not a ubiquitous phenomenon. PMID- 15276825 TI - Amygdaloid kindled seizures induce weight gain that reflects left hemisphere dominance in rats. AB - The present study sought to determine the effects of kindled seizures generated from the left and right amygdala upon weight gain in rat. Seventy-five female Sprague-Dawley rats were implanted with electrodes in basal amygdala of the left and right hemispheres. A kindling paradigm was employed in which electrical stimulation was applied once per day for 30 days after Stage 5 seizures. An electrode was implanted into the basal amygdala of the control rats but no stimulation was applied. All rats were weighed daily during the course of the experiment and changes in weight during this period were recorded for all rats. The results demonstrated that kindling from either the left or right amygdala induced significant increases in weight gain relative to the control rats. However, kindling from the left basal amygdala induced increases in body weight that were four times greater than control rats and two times greater than the rats kindled from the right side of the basal amygdala. Likewise, serum leptin levels, which were highly correlated with weight gain, also showed significantly greater increases in left amygdaloid kindled rats relative to rats kindled from the right amygdala and control rats. These findings demonstrate that basal amygdaloid kindling induces significant increases in weight gain and that the magnitude of these effects is linked to the dominance of the left hemisphere. PMID- 15276826 TI - A family-based approach reveals the function of residues in the nuclear receptor ligand-binding domain. AB - Literature studies, 3D structure data, and a series of sequence analysis techniques were combined to reveal important residues in the structure and function of the ligand-binding domain of nuclear hormone receptors. A structure based multiple sequence alignment allowed for the seamless combination of data from many different studies on different receptors into one single functional model. It was recently shown that a combined analysis of sequence entropy and variability can divide residues in five classes; (1) the main function or active site, (2) support for the main function, (3) signal transduction, (4) modulator or ligand binding and (5) the rest. Mutation data extracted from the literature and intermolecular contacts observed in nuclear receptor structures were analyzed in view of this classification and showed that the main function or active site residues of the nuclear receptor ligand-binding domain are involved in cofactor recruitment. Furthermore, the sequence entropy-variability analysis identified the presence of signal transduction residues that are located between the ligand, cofactor and dimer sites, suggesting communication between these regulatory binding sites. Experimental and computational results agreed well for most residues for which mutation data and intermolecular contact data were available. This allows us to predict the role of the residues for which no functional data is available yet. This study illustrates the power of family-based approaches towards the analysis of protein function, and it points out the problems and possibilities presented by the massive amounts of data that are becoming available in the "omics era". The results shed light on the nuclear receptor family that is involved in processes ranging from cancer to infertility, and that is one of the more important targets in the pharmaceutical industry. PMID- 15276827 TI - Scanning force microscopy of DNA translocation by the Type III restriction enzyme EcoP15I. AB - Type III restriction enzymes are multifunctional heterooligomeric enzymes that cleave DNA at a fixed position downstream of a non-symmetric recognition site. For effective DNA cleavage these restriction enzymes need the presence of two unmethylated, inversely oriented recognition sites in the DNA molecule. DNA cleavage was proposed to result from ATP-dependent DNA translocation, which is expected to induce DNA loop formation, and collision of two enzyme-DNA complexes. We used scanning force microscopy to visualise the protein interaction with linear DNA molecules containing two EcoP15I recognition sites in inverse orientation. In the presence of the cofactors ATP and Mg(2+), EcoP15I molecules were shown to bind specifically to the recognition sites and to form DNA loop structures. One of the origins of the protein-clipped DNA loops was shown to be located at an EcoP15I recognition site, the other origin had an unspecific position in between the two EcoP15I recognition sites. The data demonstrate for the first time DNA translocation by the Type III restriction enzyme EcoP15I using scanning force microscopy. Moreover, our study revealed differences in the DNA translocation processes mediated by Type I and Type III restriction enzymes. PMID- 15276829 TI - Avian reovirus morphogenesis occurs within viral factories and begins with the selective recruitment of sigmaNS and lambdaA to microNS inclusions. AB - We have recently shown that the avian reovirus non-structural protein microNS forms cytoplasmic inclusions in transfected cells and recruits sigmaNS to these structures. In the present study we further demonstrate that microNS mediates the association of the major core protein lambdaA, but not of sigmaA or sigmaC, with inclusions, indicating that the recruitment of viral proteins into avian reovirus factories has specificity. Thus, some proteins appear to be initially recruited to factories by association with microNS, whereas others are recruited subsequently through interaction with as-yet-unknown factors. We next used metabolic pulse-chase radiolabeling combined with cell fractionation and antibody immunoprecipitation to study the recruitment of newly synthesized viral polypeptides into viral factories and virus particles. The results of this combined approach revealed that avian reovirus morphogenesis is a complex and temporally controlled process that takes place exclusively within globular viral factories that are not microtubule-associated. Our findings further suggest that cores are assembled within the first 30 minutes after the synthesis of their polypeptide components, and that reovirion morphogenesis is completed over the next 30 minutes by the subsequent addition of outer capsid proteins. PMID- 15276828 TI - Interactions between the Rhodobacter sphaeroides ECF sigma factor, sigma(E), and its anti-sigma factor, ChrR. AB - Rhodobacter sphaeroides sigma(E) is a member of the extra cytoplasmic function sigma factor (ECF) family, whose members have been shown to regulate gene expression in response to a variety of signals. The functions of ECF family members are commonly regulated by a specific, reversible interaction with a cognate anti-sigma factor. In R.sphaeroides, sigma(E) activity is inhibited by ChrR, a member of a newly discovered family of zinc containing anti-sigma factors. We used gel filtration chromatography to gain insight into the mechanism by which ChrR inhibits sigma(E) activity. We found that formation of the sigma(E):ChrR complex inhibits the ability of sigma(E) to form a stable complex with core RNA polymerase. Since the sigma(E):ChrR complex inhibits the ability of the sigma factor to bind RNA polymerase, we sought to identify amino acid substitutions in sigma(E) that altered the sensitivity of this sigma factor to inhibition by ChrR. This analysis identified single amino acid changes in conserved region 2.1 of sigma(E) that either increased or decreased the sensitivity of sigma(E) for inhibition by ChrR. Many of the amino acid residues that alter the sensitivity of sigma(E) to ChrR are located within regions known to be important for interacting with core RNA polymerase in other members of the sigma(70) superfamily. Our results suggest a model where solvent-exposed residues with region 2.1 of sigma(E) interact with ChrR to sterically occlude this sigma factor from binding core RNA polymerase and to inhibit target gene expression. PMID- 15276830 TI - A conserved element in the yeast RNase MRP RNA subunit can participate in a long range base-pairing interaction. AB - RNase MRP is a ribonucleoprotein endoribonuclease involved in eukaryotic pre-rRNA processing. The enzyme possesses a putatively catalytic RNA subunit, structurally related to that of RNase P. A thorough structure analysis of Saccharomyces cerevisiae MRP RNA, entailing enzymatic and chemical probing, mutagenesis and thermal melting, identifies a previously unrecognised stem that occupies a position equivalent to the P7 stem of RNase P. Inclusion of this P7-like stem confers on yeast MRP RNA a greater degree of similarity to the core RNase P RNA structure than that described previously and better delimits domain 2, the proposed specificity domain. The additional stem is created by participation of a conserved sequence element (ymCR-II) in a long-range base-pairing interaction. There is potential for this base-pairing throughout the known yeast MRP RNA sequences. Formation of a P7-like stem is not required, however, for the pre-rRNA processing or essential function of RNase MRP. Mutants that can base-pair are nonetheless detrimental to RNase MRP function, indicating that the stem will form in vivo but that only the wild-type pairing is accommodated. Although the alternative MRP RNA structure described is clearly not part of the active RNase MRP enzyme, it would be the more stable structure in the absence of protein subunits and the probability that it represents a valid intermediate species in the process of yeast RNase MRP assembly is discussed. PMID- 15276831 TI - Terbium-mediated footprinting probes a catalytic conformational switch in the antigenomic hepatitis delta virus ribozyme. AB - The two forms of the hepatitis delta virus ribozyme are derived from the genomic and antigenomic RNA strands of the human hepatitis delta virus (HDV), where they serve a crucial role in pathogen replication by catalyzing site-specific self cleavage reactions. The HDV ribozyme requires divalent metal ions for formation of its tertiary structure, consisting of a tight double-nested pseudoknot, and for efficient self- (or cis-) cleavage. Comparison of recently solved crystal structures of the cleavage precursor and 3' product indicates that a significant conformational switch is required for catalysis by the genomic HDV ribozyme. Here, we have used the lanthanide metal ion terbium(III) to footprint the precursor and product solution structures of the cis-acting antigenomic HDV ribozyme. Inhibitory Tb(3+) binds with high affinity to similar sites on RNA as Mg(2+) and subsequently promotes slow backbone scission. We find subtle, yet significant differences in the terbium(III) footprinting pattern between the precursor and product forms of the antigenomic HDV ribozyme, consistent with differences in conformation as observed in the crystal structures of the genomic ribozyme. In addition, UV melting profiles provide evidence for a less tight tertiary structure in the precursor. In both the precursor and product we observe high-affinity terbium(III) binding sites in joining sequence J4/2 (Tb(1/2) approximately 4 microM) and loop L3, which are key structural components forming the catalytic core of the HDV ribozyme, as well as in several single-stranded regions such as J1/2 and the L4 tetraloop (Tb(1/2) approximately 50 microM). Sensitized luminescence spectroscopy confirms that there are at least two affinity classes of Tb(3+) binding sites. Our results thus demonstrate that a significant conformational change accompanies catalysis in the antigenomic HDV ribozyme in solution, similar to the catalytic conformational switch observed in crystals of the genomic form, and that structural and perhaps catalytic metal ions bind close to the catalytic core. PMID- 15276832 TI - The biochemical requirements of DNA polymerase V-mediated translesion synthesis revisited. AB - In addition to replicative DNA polymerases, cells contain specialized DNA polymerases involved in processes such as lesion tolerance, mutagenesis and immunoglobulin diversity. In Escherichia coli, DNA polymerase V (Pol V), encoded by the umuDC locus, is involved in translesion synthesis (TLS) and mutagenesis. Genetic studies have established that mutagenesis requires both UmuC and a proteolytic product of UmuD (UmuD'). In addition, RecA protein and the replication processivity factor, the beta-clamp, were genetically found to be essential co-factors for mutagenesis. Here, we have reconstituted Pol V-mediated bypass of three common replication-blocking lesions, namely the two major UV induced lesions and a guanine adduct formed by a chemical carcinogen (G-AAF) under conditions that fulfil these in vivo requirements. Two co-factors are essential for efficient Pol V-mediated lesion bypass: (i) a DNA substrate onto which the beta-clamp is stably loaded; and (ii) an extended single-stranded RecA/ATP filament assembled downstream from the lesion site. For efficient bypass, Pol V needs to interact simultaneously with the beta-clamp and the 3' tip of the RecA filament. Formation of an extended RecA/ATP filament and stable loading of the beta-clamp are best achieved on long single-stranded circular DNA templates. In contrast to previously published data, the single-stranded DNA binding protein (SSB) is not absolutely required for Pol V-mediated lesion bypass provided ATP, instead of ATPgammaS, activates the RecA filament. Further discrepancies with the existing literature are explainable by the use of either inadequate DNA substrates or a UmuC fusion protein instead of native Pol V. PMID- 15276833 TI - Modulation of DNA conformations through the formation of alternative high-order HU-DNA complexes. AB - HU is an abundant, highly conserved protein associated with the bacterial chromosome. It belongs to a small class of proteins that includes the eukaryotic proteins TBP, SRY, HMG-I and LEF-I, which bind to DNA non-specifically at the minor groove. HU plays important roles as an accessory architectural factor in a variety of bacterial cellular processes such as DNA compaction, replication, transposition, recombination and gene regulation. In an attempt to unravel the role this protein plays in shaping nucleoid structure, we have carried out fluorescence resonance energy transfer measurements of HU-DNA oligonucleotide complexes, both at the ensemble and single-pair levels. Our results provide direct experimental evidence for concerted DNA bending by HU, and the abrogation of this effect at HU to DNA ratios above about one HU dimer per 10-12 bp. These findings support a model in which a number of HU molecules form an ordered helical scaffold with DNA lying in the periphery. The abrogation of these nucleosome-like structures for high HU to DNA ratios suggests a unique role for HU in the dynamic modulation of bacterial nucleoid structure. PMID- 15276834 TI - Downstream DNA selectively affects a paused conformation of human RNA polymerase II. AB - Transcriptional pausing by human RNA polymerase II (RNAPII) in the HIV-1 LTR is caused principally by a weak RNA:DNA hybrid that allows rearrangement of reactive or catalytic groups in the enzyme's active site. This rearrangement creates a transiently paused state called the unactivated intermediate that can backtrack into a more long-lived paused species. We report that three different regions of the not-yet-transcribed DNA just downstream of the pause site affect the duration of the HIV-1 pause, and also can influence pause formation. Downstream DNA in at least one region, a T-tract from +5 to +8, increases pause duration by specifically affecting the unactivated intermediate, without corresponding effects on the active or backtracked states. We suggest this effect depends on RNAPII-modulated DNA plasticity and speculate it is mediated by the "trigger loop" thought to participate in RNAP's catalytic cycle. These findings provide a new framework for understanding downstream DNA effects on RNAP. PMID- 15276835 TI - Stopped-flow and mutational analysis of base flipping by the Escherichia coli Dam DNA-(adenine-N6)-methyltransferase. AB - By stopped-flow kinetics using 2-aminopurine as a probe to detect base flipping, we show here that base flipping by the Escherichia coli Dam DNA-(adenine-N6) methyltransferase (MTase) is a biphasic process: target base flipping is very fast (k(flip)>240 s(-1)), but binding of the flipped base into the active site pocket of the enzyme is slow (k=0.1-2 s(-1)). Whereas base flipping occurs in the absence of S-adenosyl-l-methionine (AdoMet), binding of the target base in the active site pocket requires AdoMet. Our data suggest that the tyrosine residue in the DPPY motif conserved in the active site of DNA-(adenine-N6)-MTases stacks to the flipped target base. Substitution of the aspartic acid residue of the DPPY motif by alanine abolished base flipping, suggesting that this residue contacts and stabilizes the flipped base. The exchange of Ser188 located in a loop next to the active center by alanine led to a seven- to eightfold reduction of k(flip), which was also reduced with substrates having altered GATC recognition sites and in the absence of AdoMet. These findings provide evidence that the enzyme actively initiates base flipping by stabilizing the transition state of the process. Reduced rates of base flipping in substrates containing the target base in a non-canonical sequence demonstrate that DNA recognition by the MTase starts before base flipping. DNA recognition, cofactor binding and base flipping are correlated and efficient base flipping takes place only if the enzyme has bound to a cognate target site and AdoMet is available. PMID- 15276836 TI - Crystal structure of the reaction complex of 3-deoxy-D-arabino-heptulosonate-7 phosphate synthase from Thermotoga maritima refines the catalytic mechanism and indicates a new mechanism of allosteric regulation. AB - 3-Deoxy-d-arabino-heptulosonate-7-phosphate synthase (DAHPS) catalyzes the first reaction of the aromatic biosynthetic pathway in bacteria, fungi, and plants, the condensation of phosphoenolpyruvate (PEP) and d-erythrose-4-phosphate (E4P) with the formation of DAHP. Crystals of DAHPS from Thermotoga maritima (DAHPS(Tm)) were grown in the presence of PEP and metal cofactor, Cd(2+), and then soaked with E4P at 4 degrees C where the catalytic activity of the enzyme is negligible. The crystal structure of the "frozen" reaction complex was determined at 2.2A resolution. The subunit of the DAHPS(Tm) homotetramer consists of an N-terminal ferredoxin-like (FL) domain and a (beta/alpha)(8)-barrel domain. The active site located at the C-end of the barrel contains Cd(2+), PEP, and E4P, the latter bound in a non-productive conformation. The productive conformation of E4P is suggested and a catalytic mechanism of DAHPS is proposed. The active site of DAHPS(Tm) is nearly identical to the active sites of the other two known DAHPS structures from Escherichia coli (DAHPS(Ec)) and Saccharomyces cerevisiae (DAHPS(Sc)). However, the secondary, tertiary, and quaternary structures of DAHPS(Tm) are more similar to the functionally related enzyme, 3-deoxy-d-manno octulosonate-8-phosphate synthase (KDOPS) from E.coli and Aquiflex aeolicus, than to DAHPS(Ec) and DAHPS(Sc). Although DAHPS(Tm) is feedback-regulated by tyrosine and phenylalanine, it lacks the extra barrel segments that are required for feedback inhibition in DAHPS(Ec) and DAHPS(Sc). A sequence similarity search revealed that DAHPSs of phylogenetic family Ibeta possess a FL domain like DAHPS(Tm) while those of family Ialpha have extra barrel segments similar to those of DAHPS(Ec) and DAHPS(Sc). This indicates that the mechanism of feedback regulation in DAHPS(Tm) and other family Ibeta enzymes is different from that of family Ialpha enzymes, most likely being mediated by the FL domain. PMID- 15276837 TI - The elongation and contraction of actin bundles are induced by double-headed myosins in a motor concentration-dependent manner. AB - Many types of myosin have been found and characterized to date, and already nearly 20 classes have been identified. However, these myosin motors can be classified more simply into two groups according to their head-structure, i.e. double- or single-headed myosins. Why do some myosin motors possess a double headed structure? One obvious possible reason would be that the two heads improve the motor's processivity and sliding performance. Previously, to investigate the possibility that the double-headed myosins simultaneously interact with parallel arrayed two actin filaments in the presence of Mg-ATP, we developed an in vitro assay system using actin bundles formed by inert polymers. Using that system, we show here that skeletal muscle heavy meromyosin (HMM), a double-headed myosin derivative, but not subfragment-1 (S-1), a single-headed one, was able to contract or elongate actin bundles in a concentration-dependent manner. Similar elongation or contraction of actin bundles can also be induced by other double headed myosin species isolated in the native state from Dictyostelium, from green algae Chara or from chicken brain. The results of this study confirm that double headed myosin motors can induce sliding movements among neighboring actin filaments. The double-headed structure of myosins may also be important for generating tension or elongation in actin bundles or gels, and for organizing polarity-sorted actin networks, not just for improving their motor processivity or activity. PMID- 15276839 TI - Domain organization and function of Salmonella FliK, a flagellar hook-length control protein. AB - Salmonella hook-length control protein FliK, which consists of 405 amino acid residues, switches substrate specificity of the type III flagellar protein export apparatus from rod/ hook-type to filament-type by causing a conformational change in the cytoplasmic domain of FlhB (FlhB(C)) upon completion of the hook assembly. An N-terminal region of FliK contains an export signal, and a highly conserved C terminal region consisting of amino acid residues 265-405 (FliK((265-405))) is directly involved in the switching of FlhB(C). Here, we have investigated the structural properties of FliK. Gel filtration chromatography, multi-angle light scattering and analytical ultracentrifugation showed that FliK is monomeric in solution and has an elongated shape. Limited proteolysis showed that FliK consists of two domains, the N-terminal (FliK(N)) and C-terminal domains (FliK(C)), and that the first 203 and the last 35 amino acid residues are partially unfolded and subjected to proteolysis. Both FliK(N) and FliK(C) are more globular than full-length FliK, suggesting that these domains are connected in tandem. Overproduced His-FliK((199-405)) failed to switch export specificity of the export apparatus. Affinity blotting revealed that FlhB(C) binds to FliK and FliK((1-147)), but not to FliK((265-405)). Based on these results, we propose that FliK(N) within the central channel of the hook-basal body during the export of FliK is the sensor and transmitter of hook completion information and that the binding interaction of FliK(C) to FlhB(C) is structurally regulated by FliK(N) so as to occur only when the hook has reached a preset length. The conformational flexibility of FliK(C) may play a role in interfering with switching at an inappropriate point of flagellar assembly. PMID- 15276838 TI - A highly conserved interspecies V(H) in the human genome. AB - Idiotype conservation between human and mouse antibodies has been observed in association with various infectious and autoimmune diseases. We have isolated a human anti-idiotypic antibody to a mouse monoclonal anti-IgE antibody (BSW17) suggesting a conserved interspecies idiotype associated with an anti-IgE response. To find the homologue of BSW17 in the human genome we applied the guided selection strategy. Combining V(H) of BSW17 with a human V(L) repertoire resulted in three light chains. The three V(L) chains were then combined with a human V(H) repertoire resulting in three clones specific for human IgE. Surprisingly, one clone, Hu41, had the same epitope specificity and functional in vitro activity as BSW17 and V(H) complementarity-determining regions identical with that of BSW17. Real-time PCR analysis confirmed the presence of the Hu41 V(H) sequence in the human genome. These data document the first example of the isolation of a human antibody where high sequence similarity to the original murine V(H) sequence is associated with common antigen and epitope specificity. PMID- 15276840 TI - Crystal structure of the Mycobacterium tuberculosis dUTPase: insights into the catalytic mechanism. AB - The structure of Mycobacterium tuberculosis dUTP nucleotidohydrolase (dUTPase) has been determined at 1.3 Angstrom resolution in complex with magnesium ion and the non-hydrolyzable substrate analog, alpha,beta-imido dUTP. dUTPase is an enzyme essential for depleting potentially toxic concentrations of dUTP in the cell. Given the importance of its biological role, it has been proposed that inhibiting M.tuberculosis dUTPase might be an effective means to treat tuberculosis infection in humans. The crystal structure presented here offers some insight into the potential for designing a specific inhibitor of the M.tuberculosis dUTPase enzyme. The structure also offers new insights into the mechanism of dUTP hydrolysis by providing an accurate representation of the enzyme-substrate complex in which both the metal ion and dUTP analog are included. The structure suggests that inclusion of a magnesium ion is important for stabilizing the position of the alpha-phosphorus for an in-line nucleophilic attack. In the absence of magnesium, the alpha-phosphate of dUTP can have either of the two positions which differ by 4.5 Angstrom. A transiently ordered C terminal loop further assists catalysis by shielding the general base, Asp83, from solvent thus elevating its pK(a) so that it might in turn activate a tightly bound water molecule for nucleophilic attack. The metal ion coordinates alpha, beta, and gamma phosphate groups with tridentate geometry identical with that observed in the crystal structure of DNA polymerase beta complexed with magnesium and dNTP analog, revealing some common features in catalytic mechanism. PMID- 15276841 TI - Crystal structure of EMS16 in complex with the integrin alpha2-I domain. AB - Snake venoms contain a number of heterodimeric C-type lectin-like proteins (CLPs) that interact specifically with components of the haemostatic system. EMS16 from the venom of Echis multisquamatus binds to the collagen receptor, integrin alpha2beta1, also known as glycoprotein (GP) Ia/IIa, and specifically inhibits collagen binding. Here we report the crystal structure of EMS16 in complex with recombinant integrin alpha2-I domain that plays a central role in collagen binding. The structure of the complex at 1.9 Angstrom resolution reveals that the collagen-binding site of the alpha2-I domain is covered completely by the bound EMS16. This blockage by EMS16 appears to spatially inhibit collagen binding to the alpha2-I domain. The bound alpha2-I domain adopts a closed conformation, which is seen in the absence of ligand, suggesting that EMS16 stabilizes a closed conformation corresponding to the less active structure of the alpha2-I domain. EMS16 does not directly bind to the manganese ion and residues of the metal ion dependent adhesion site (MIDAS) of the alpha2-I domain, suggesting that EMS16 may have the potential to bind specifically to the alpha2-I domain in a metal ion independent fashion. PMID- 15276843 TI - Probing hydration of monovalent cations condensed around polymeric nucleic acids. AB - We report the relative molar sound velocity increments, [U], partial molar volumes, V(o), and partial molar adiabatic compressibilities, K(S)(o), of the Li(+), Na(+), K(+), Rb(+), Cs(+), NH(4)(+), and N(CH(3))(4)(+) salts of poly(dAdT)poly(dAdT), poly(dGdC)poly(dGdC), poly(dIdC)poly(dIdC), poly(rA)poly(rU), poly(rG)poly(rC), poly(rI)poly(rC), and poly(rU) at 25 degrees C. When analyzing these data, we take into account the Donnan membrane equilibrium effect. Comparison between the values of [U], V(o), and K(S)(o) exhibited by the nucleic acid salts and respective chlorides (LiCl, NaCl, KCl, RbCl, CsCl, NH(4)Cl, and N(CH(3))(4)Cl) yields information about the state of counterion hydration in the vicinity of each nucleic acid structure studied here. Our analysis reveals that the poly(dGdC)poly(dGdC), poly(dIdC)poly(dIdC), and poly(rI)poly(rC) duplexes and single-stranded poly(rU) do not significantly influence the hydration properties of their condensed counterions. In the vicinity of these polymers, counterions retain their full hydration shells (within +/-15%). By contrast, counterions condensed around the poly(dAdT)poly(dAdT), poly(rA)poly(rU), and poly(rG)poly(rC) duplexes are significantly dehydrated and retain, respectively, only 65(+/-18)%, 34(+/-21)%, and 33(+/-9)% of their original hydration shells. Taken together, the volumetric data reported here provide important new information that ultimately may help us understand the central role that hydration and counterions play in modulating the conformational preferences of nucleic acids and the energetics of DNA recognition events. PMID- 15276842 TI - How insulin binds: the B-chain alpha-helix contacts the L1 beta-helix of the insulin receptor. AB - Binding of insulin to the insulin receptor plays a central role in the hormonal control of metabolism. Here, we investigate possible contact sites between the receptor and the conserved non-polar surface of the B-chain. Evidence is presented that two contiguous sites in an alpha-helix, Val(B12) and Tyr(B16), contact the receptor. Chemical synthesis is exploited to obtain non-standard substitutions in an engineered monomer (DKP-insulin). Substitution of Tyr(B16) by an isosteric photo-activatable derivative (para-azido-phenylalanine) enables efficient cross-linking to the receptor. Such cross-linking is specific and maps to the L1 beta-helix of the alpha-subunit. Because substitution of Val(B12) by larger side-chains markedly impairs receptor binding, cross-linking studies at B12 were not undertaken. Structure-function relationships are instead probed by side-chains of similar or smaller volume: respective substitution of Val(B12) by alanine, threonine, and alpha-aminobutyric acid leads to activities of 1(+/ 0.1)%, 13(+/-6)%, and 14(+/-5)% (relative to DKP-insulin) without disproportionate changes in negative cooperativity. NMR structures are essentially identical with native insulin. The absence of transmitted structural changes suggests that the low activities of B12 analogues reflect local perturbation of a "high-affinity" hormone-receptor contact. By contrast, because position B16 tolerates alanine substitution (relative activity 34(+/-10)%), the contribution of this neighboring interaction is smaller. Together, our results support a model in which the B-chain alpha-helix, functioning as an essential recognition element, docks against the L1 beta-helix of the insulin receptor. PMID- 15276844 TI - X-ray and thermodynamic studies of staphylococcal nuclease variants I92E and I92K: insights into polarity of the protein interior. AB - We have used crystallography and thermodynamic analysis to study nuclease variants I92E and I92K, in which an ionizable side-chain is placed in the hydrophobic core of nuclease. We find that the energetic cost of burying ionizable groups is rather modest. The X-ray determinations show water molecules solvating the buried glutamic acid under cryo conditions, but not at room temperature. The lysine side-chain does not appear solvated in either case. Guanidine hydrochloride (GnHCl) denaturation of I92E and I92K, done as a function of pH and monitored by tryptophan fluorescence, showed that I92E and I92K are folded in the pH range pH 3.5-9.0 and pH 5.5-9.5, respectively. The stability of the parental protein is independent of pH over a broad range. In contrast, the stabilities of I92E and I92K exhibit a pH dependence, which is quantitatively explained by thermodynamic analysis: the PK(a) value of the buried K92 is 5.6, while that of the buried E92 is 8.65. The free energy difference between burying the uncharged and charged forms of the groups is modest, about 6 kcal/mol. We also found that epsilon(app) for I92K and I92E is in the range approximately 10 12, instead of 2-4 commonly used to represent the protein interior. Side-chains 92E and 92K were uncharged under the conditions of the X-ray experiment. Both are buried completely inside the well-defined hydrophobic core of the variant proteins without forming salt-bridges or hydrogen bonds to other functional groups of the proteins. Under cryo conditions 92E shows a chain of four water molecules, which hydrate one oxygen atom of the carboxyl group of the glutamic acid. Two other water molecules, which are present in the wild-type at all temperatures, are also connected to the water ring observed inside the hydrophobic core. The ready burial of water with an uncharged E92 raises the possibility that solvent excursions into the interior also take place in the wild type protein, but in a random, dynamic way not detectable by crystallography. Such transient excursions could increase the average polarity, and thus epsilon(app), of the protein interior. PMID- 15276845 TI - Modulation of S6 fibrillation by unfolding rates and gatekeeper residues. AB - We present a protein engineering analysis of the fibrillation of a protein from a thermophilic organism, the 101 residue S6 from Thermus thermophilus. When agitated, S6 fibrillates at pH 2.0 in 0.4 M NaCl. Under these solvent conditions, S6 has native-like secondary structure and also unfolds and refolds cooperatively. However, its tertiary structure appears to be more plastic than at neutral pH, and some regions of the protein may be partially unstructured. At 42 degrees C, there is a lag phase of several days after which fibrillation takes place over several hours. Data from the fibrillation behaviour of a comprehensive series of single and double mutants of S6 suggests that several factors control the onset of fibrillation. Firstly, there appears to be a contiguous region of "gatekeeper" residues that inhibit fibrillation, since their truncation significantly reduces the duration of the lag phase. This region overlaps extensively with the partially unstructured region of the protein, suggesting that residues with enhanced flexibility and solvent-accessibility are important for the initiation of fibrillation. Secondly, longer lag phases correlate with faster rates of unfolding. We interpret this to mean that kinetic stability also controls fibrillation but in the sense that the quasi-native state, rather than the denatured state, is the species that participates in nucleation. This implies that fibrillation can also occur from a quasi-native state as opposed to an ensemble of highly fluctuating structures, and highlights the delicate balance between flexibility and structure required to form organized assemblies of polypeptide chains. PMID- 15276846 TI - Localized nature of the transition-state structure in goat alpha-lactalbumin folding. AB - To investigate whether the structure partially formed in the molten globule folding intermediate of goat alpha-lactalbumin is further organized in the transition state of folding, we constructed a number of mutant proteins and performed Phi-value analysis on them. For this purpose, we measured the equilibrium unfolding transitions and kinetic refolding and unfolding reactions of the mutants using equilibrium and stopped-flow kinetic circular dichroism techniques. The results show that the mutants with mutations located in the A helix (V8A, L12A), the B-helix (V27A), the beta-domain (L52A, W60A), the C-helix (K93A, L96A), the C-D loop (Y103F), the D-helix (L105A, L110A), and the C terminal 3(10)-helix (W118F), have low Phi-values, less than 0.2. On the other hand, D87N, which is located on the Ca(2+)-binding site, has a high Phi-value, 0.91, indicating that tight packing of the side-chain around Asp87 occurs in the transition state. One beta-domain mutant (I55V) and three C-helix mutants (I89V, V90A, and I95V) demonstrated intermediate Phi-values, between 0.4 and 0.7. These results indicate that the folding nucleus in the transition state of goat alpha LA is not extensively distributed over the alpha-domain of the protein, but very localized in a region that contains the Ca(2+)-binding site and the interface between the C-helix and the beta-domain. This is apparently in contrast with the fact that the molten globule state of alpha-lactalbumin has a partially formed structure inside the alpha-domain. It is concluded that the specific docking of the alpha and beta-domains at a domain interface is necessary for this protein to organize its native structure from the molten globule intermediate. PMID- 15276847 TI - Iron binding and oxidation kinetics in frataxin CyaY of Escherichia coli. AB - Friedreich's ataxia is associated with a deficiency in frataxin, a conserved mitochondrial protein of unknown function. Here, we investigate the iron binding and oxidation chemistry of Escherichia coli frataxin (CyaY), a homologue of human frataxin, with the aim of better understanding the functional properties of this protein. Anaerobic isothermal titration calorimetry (ITC) demonstrates that at least two ferrous ions bind specifically but relatively weakly per CyaY monomer (K(d) approximately 4 microM). Such weak binding is consistent with the hypothesis that the protein functions as an iron chaperone. The bound Fe(II) is oxidized slowly by O(2). However, oxidation occurs rapidly and completely with H(2)O(2) through a non-enzymatic process with a stoichiometry of two Fe(II)/H(2)O(2), indicating complete reduction of H(2)O(2) to H(2)O. In accord with this stoichiometry, electron paramagnetic resonance (EPR) spin trapping experiments indicate that iron catalyzed production of hydroxyl radical from Fenton chemistry is greatly attenuated in the presence of CyaY. The Fe(III) produced from oxidation of Fe(II) by H(2)O(2) binds to the protein with a stoichiometry of six Fe(III)/CyaY monomer as independently measured by kinetic, UV-visible, fluorescence, iron analysis and pH-stat titrations. However, as many as 25-26 Fe(III)/monomer can bind to the protein, exhibiting UV absorption properties similar to those of hydrolyzed polynuclear Fe(III) species. Analytical ultracentrifugation measurements indicate that a tetramer is formed when Fe(II) is added anaerobically to the protein; multiple protein aggregates are formed upon oxidation of the bound Fe(II). The observed iron oxidation and binding properties of frataxin CyaY may afford the mitochondria protection against iron induced oxidative damage. PMID- 15276848 TI - Empirical analysis of protein insertions and deletions determining parameters for the correct placement of gaps in protein sequence alignments. AB - To understand how protein segments are inserted and deleted during divergent evolution, a set of pairwise alignments contained exactly one gap, and therefore arising from the first insertion-deletion (indel) event in the time separating the homologs, was examined. The alignments showed that "structure breaking" amino acids (PGDNS) were preferred within and flanking gapped regions, as are two residues with hydrophilic side-chains (QE) that frequently occur at the surface of protein folds. Conversely, hydrophobic residues (FMILYVW) occur infrequently within and flanking the gapped region. These preferences are modestly different in protein pairs separated by an episode of adaptive evolution, than in pairs diverging under strong functional constraints. Surprisingly, regions near an indel have not evolved more rapidly than the sequence pair overall, showing no evidence that an indel event must be compensated by local amino acid replacement. The gap-lengths are best approximated by a Zipfian distribution, with the probability of a gap of length L decreasing as a function of L(-1.8). These features are largely independent of the length of the gap and the extent of divergence (measured by both silent and non-silent sequence changes) separating the two proteins. Surprisingly, amino acid repeats were discovered in more than a third of the polypeptide segments in and around the gap. These correspond to repeats in the DNA sequence. This suggests that a signature of the mechanism by which indels occur in the DNA sequence remains in the encoded protein sequences. These data suggest specific tools to score gap placement in an alignment. They also suggest tools that distinguish true indels from gaps created by mistaken gene finding, including under-predicted and over-predicted introns. By providing mechanisms to identify errors, the tools will enhance the value of genome sequence databases in support of integrated paleogenomics strategies used to extract functional information in a post-genomic environment. PMID- 15276849 TI - Spontaneous lacquer crack lesions in the retinopathy, globe enlarged (rge) chick. AB - Lacquer crack lesion (LCL), a complication of myopia in human patients, is characterized by loss of retinal pigment epithelium and breaks in Bruch's membrane. This report describes comparable lesions in the "retinopathy, globe enlarged" (rge) chick. Twenty-six birds, (nine rge/rge [affected], 12 rge/+ [carriers] and five +/+ [normal]), were examined ophthalmoscopically from hatching up to 336 days of age. Ophthalmoscopically detected fundus lesions were investigated by light and transmission electron microscopy. Pale, linear fundus lesions were detected in both eyes of seven of the nine rge/rge chicks, from as early as 45 days of age. Histological and ultrastructural examination of four of the affected rge/rge chicks revealed areas of ruptured Bruch's membrane, with focal absence of retinal pigment epithelium. Fibroblasts covered the interface of the abnormal Bruch's membrane and choriocapillaris. There was disorganization of overlying photoreceptor outer and inner segments, and thinning of inner and outer nuclear layers, while the rest of the inner retina appeared unaltered. The lesions present in rge chicks showed histological changes similar to those of LCLs described in human patients with pathological myopia. The formation of LCLs in rge chicks is probably due to stretching of Bruch's membrane secondary to abnormal globe enlargement, resulting in linear rupture of the membrane and associated changes. The rge chick may prove a useful model for human LCL formation. PMID- 15276850 TI - A retrospective study of 286 cases of neurological disorders of the cat. AB - Archive central nervous tissue from 286 cats with neurological disorders was reviewed for histological evidence of feline spongiform encephalopathy (FSE), which may have occurred before it was first recognized in 1990. The following six categories of disease were identified: congenital; degenerative; inflammatory; neoplastic; FSE; lesion-free. The largest category (inflammatory) contained 92 cats, of which 47 were considered to be consistent with infection by feline infectious peritonitis (FIP) virus. Six cats showed evidence of more than one disease process; thus, one cat with FIP also had toxocara infection of the lateral ventricles and five cats with FSE also showed perivascular cuffing suggestive of concurrent viral infection. In only two cases did the diagnosis on review differ significantly from the original interpretation. There was no evidence of FSE before the original case was recognized in April 1990. PMID- 15276851 TI - Expression of monocyte chemoattractant protein-1 and macrophage inflammatory protein-1 in porcine circovirus 2-induced granulomatous inflammation. AB - The expression of monocyte chemoattractant protein-1 (MCP-1) and macrophage inflammatory protein-1 (MIP-1) in granulomatous inflammation of lymph nodes was determined in 1-day-old piglets inoculated intranasally with porcine circovirus 2 (PCV2). The methods used were reverse transcription-polymerase chain reaction (RT PCR), granulomatous lesion scoring, and in-situ hybridization. Maximum MCP-1 expression and MIP-1 expression were observed at 17 and 21 days post-inoculation (dpi), respectively. Granulomatous lesions of maximum severity were observed somewhat later (28 dpi). A close cell to cell relation between PCV2 and the two cytokines in serial sections from lymph nodes suggested that their induction was a direct response to PCV2. The results provide evidence of the ability of MCP-1 and MIP-1 to induce granulomatous inflammation in vivo. PMID- 15276852 TI - Lectin binding patterns and immunohistochemical antigen detection in the genitalia of Tritrichomonas foetus-infected heifers. AB - Heifers inoculated intra-vaginally with Tritrichomonas foetus were examined after long-term infection (70 days) and short-term infection (20 days) by lectin histochemical, immunohistochemical and cultural techniques. The organism was recovered from the genital tract and T. foetus antigens were detected immunohistochemically in the lumina of uterine glands and cytoplasm of vaginal subepithelial macrophages. An increase of galactosylated residues (galactose and N-acetyl galactose), binding to PNA, was observed in the genital epithelium (vagina, uterus and oviduct) from infected animals. In the oviductal epithelium of short- but not long-term infected heifers, mannose (binding to Con A) was detected, suggesting that the persistent presence of T. foetus and its virulence factors or inflammatory processes result in a change in the glycoproteins of the epithelial surface. The findings have implications for the adhesion of T. foetus to cells and for the pathogenesis of bovine trichomonosis. PMID- 15276853 TI - Characteristic and non-characteristic pathological findings in peste des petits ruminants (PPR) of sheep in the Ege district of Turkey. AB - This report describes the pathological and immunohistochemical findings in naturally infected lambs from three outbreaks of peste des petits ruminants in Mugla and Aydin provinces of the Ege district of Turkey. At necropsy, ulcerative stomatitis, catarrhal or fibrinous bronchopneumonia, and acute catarrhal enteritis were observed. Histopathologically, syncytial cells containing inclusion bodies were seen in the tongue and in the buccal, labial and soft palate mucosae. In pneumonic lungs, syncytial cells were present in the alveolar lumina, and cytoplasmic inclusion bodies in epithelial cells of the bronchi and bronchioli. Immunohistochemically, viral antigen was strongly labelled in the lung, oral tissues and small intestine. PMID- 15276854 TI - Placental pathology associated with fetal death in cattle inoculated with Neospora caninum by two different routes in early pregnancy. AB - Pregnant cattle were inoculated with N. caninum strain NC-1 tachyzoites intravenously (iv) (group 1, n = 8) or subcutaneously (sc) (group 2, n = 8) at 70 days' gestation. Control animals (group 3; n = 8) received uninfected Vero cells iv. Two animals from each group were killed at 14, 28, 42 and 56 days post inoculation (dpi). Fetal mortality was 100% and 50%, respectively, in groups 1 and 2 from 28 dpi. In group 1 foci of degenerative fetal placental villi were observed at 14 dpi, with clusters of N. caninum tachyzoites in the affected mesenchyme. There was also inflammation of maternal septal tissues, with necrotic cell debris and serum exudate at the interstitium. At 28 dpi pregnancy had ended and the fetal cotyledons had become detached from the maternal caruncles. Immunohistochemically, particulate N. caninum antigen was detected in the cotyledons. At 42 and 56 dpi, fetal tissues had disappeared, the caruncles were greatly reduced in size, and the uterine epithelium had been largely restored. In group 2, lesions were either severe or absent ("all or nothing" response). In one animal carrying a dead fetus at 28 dpi, placentitis was much more severe than that seen in group 1 at 14 dpi. Lesions contained neutrophils, eosinophils and N. caninum antigen. In animals carrying dead fetuses at 42 and 56 dpi, fetal remains were found and the cotyledons contained N. caninum antigen. Antigen was also detected in fetal tissues. No significant pathological changes were detected in group 2 animals carrying live fetuses or any animal in group 3. Thus, N. caninum administered iv or sc in early pregnancy resulted in rapid fetal death, with parasite-associated lesions in the placenta and fetus. Of the two inoculation routes, the intravenous induced the more acute placental lesions and greater mortality. PMID- 15276855 TI - Non-functional C-cell adenoma in aged horses. AB - Thyroid tumours occur in older horses, and most such tumours have been considered to be of follicular epithelial origin. However, their immunohistochemical characterization has not been investigated. The purpose of this study was to confirm a suspicion that most of these tumours are in fact parafollicular cell (C cell)-derived adenomas, and to evaluate their pathogenesis and functional state. Thyroid glands from 38 horses aged 10-29 years were evaluated, all tissue samples being examined histologically, immunohistochemically and ultrastructurally. Nodular tumour masses were found in the thyroids of 12 of 38 horses older than 10 years (31.6%), and in nine of 12 horses older than 20 years (75.0%), regardless of sex or breed. Nodular lesions were composed of solid proliferations of polygonal cells with eosinophilic granular cytoplasm. Immunohistochemically, tumour cells were positive for calcitonin and neuron specific enolase, but negative for thyroglobulin. Ultrastructurally, few if any secretion granules were found in tumour cells. On the basis of these results it was concluded that the nodular lesions were C-cell adenomas, not follicular adenomas. It was suspected that the C-cell adenomas were non-functional and unlikely to lead to calcitonin hypersecretion-related diseases. PMID- 15276856 TI - Development of pericardial mesothelioma in golden retrievers with a long-term history of idiopathic haemorrhagic pericardial effusion. AB - This report describes the development of pericardial mesothelioma in five golden retrievers with a long-term history of idiopathic haemorrhagic pericardial effusion (IHPE). These five dogs were treated with repeated pericardiocentesis for recurrent episodes of pericardial fluid accumulation; other than IHPE, all potential causes of this fluid accumulation were ruled out by the results of diagnostic imaging and cytology and bacterial or fungal culture of fluid obtained during pericardiocentesis. In three dogs that eventually underwent pericardiectomy, neoplastic lesions were not detected in any organs or tissues within the thoracic cavity during the surgical procedure, and the surgical biopsies were consistent with IHPE. In one of the three dogs, however, cytology of recurrent thoracic effusion revealed clusters of neoplastic mesothelial cells from 1 month after surgical intervention until death. The clinical course of the disease ranged from 30 to 54 months between the first visit and death, and on post-mortem examination pericardial mesothelioma was diagnosed in all five dogs. The clinical observations, together with the breed and age of the affected animals, suggested that the five dogs initially suffered from IHPE, which was then followed by the development of pericardial mesothelioma. It is possible that IHPE is associated with the development of pericardial mesothelioma in golden retrievers through a chronic inflammatory process. PMID- 15276857 TI - Pathology and viral distribution in fatal Usutu virus infections of birds from the 2001 and 2002 outbreaks in Austria. AB - In the summer of 2001, Usutu virus (USUV) was isolated for the first time in Europe, from an episode of mass mortality in Eurasian blackbirds (Turdus merula). In the present study, 40 of the birds (representing three species), confirmed as cases of USUV infection, were examined by four methods (histopathology, immunohistochemistry [IHC], in-situ hybridization [ISH] and reverse-transcriptase polymerase chain reaction [RT-PCR]). The major macroscopical finding was hepatosplenomegaly; histologically, neuronal necrosis, myocardial lesions, and coagulative necrosis of the liver and spleen were observed. IHC with cross reactive polyclonal antibodies to West Nile virus detected viral antigen predominantly in brain neurons (40/40 birds; 100%), myocardial fibres (25/32; 78%), cells of the splenic capsule (29/33; 88%), renal glomeruli (22/35; 63%), tunica muscularis of intestines (17/22; 77%), proventricular glands (16/19; 84%), lungs (18/33; 55%) and hepatic Kupffer cells (7/38; 18%). ISH with an USUV specific oligonucleotide probe demonstrated viral nucleic acid predominantly in brain neurons (40/40; 100%), myocardial fibres (24/33; 73%), splenic macrophages (12/34; 35%), renal tubular cells (19/36; 53%), tunica muscularis of intestines (13/32; 41%), proventricular glands (19/22; 86%), lungs (7/34; 21%) and hepatic Kupffer cells (12/38; 32%). All of 33 birds tested additionally by USUV-specific RT-PCR gave positive results. PMID- 15276858 TI - Compensated overexpression of procollagens alpha 1(I) and alpha 1(III) following perilla mint ketone-induced acute pulmonary damage in horses. AB - Interstitial lung disease with chronic fibrosis is a frequent cause of reduced performance in horses. The aim of this study was to establish a model of acute alveolar damage and interstitial lung disease in horses that could be used to monitor the histopathological lesions and changes in expression levels of genes relevant to pulmonary fibrosis. Six adult horses were given a single intravenous injection (6 mg per kg body weight) of perilla mint ketone (PMK). Transthoracic lung biopsy samples (1 x 0.2 x 0.2 cm) were collected before and after (days 1, 4, 8, 11, 15, 18, 22, 25 and 29) the administration of PMK. Light and electron microscopy revealed severe acute alveolar damage (days 1 to 4), proliferation of type II pneumocytes (days 4 to 11) and finally complete healing at about day 18. However, unexpectedly severe clinical signs necessitated euthanasia in two horses on days 9 and 11. The expression levels of the collagen genes COL1AI and COL3AI as well as transforming growth factor (TGF)-beta were examined in the biopsy samples by reverse transcription-real time quantitative polymerase chain reaction. COL1AI and COL3AI gene expressions were upregulated (3- and 17-fold, respectively) between days 1 and 29 in all six horses, whereas TGF-beta was upregulated in two horses (2- and 4-fold, respectively), between days 4 and 18. Although the gene expression analyses indicated a strong activation of the pro fibrotic pathway, no interstitial fibrosis was seen in any horse. A complete necropsy performed on day 60 revealed complete recovery of the lungs of the four surviving horses, with no evidence of fibrosis. Unidentified compensatory mechanisms may have prevented pulmonary fibrosis, despite strong upregulation of pro-fibrotic genes. PMID- 15276859 TI - Monoclonal antibody-based immunohistochemical diagnosis of Malaysian Nipah virus infection in pigs. AB - Formalin-fixed, paraffin wax-embedded tissues of three Malaysian farm pigs naturally infected with Nipah virus were used to investigate the value of anti Nipah virus mouse monoclonal antibodies (Mabs) and rabbit polyclonal antibody for immunohistochemical diagnosis. Mabs 11F6 and 12A5 gave intense immunolabelling in lung tissue that had been fixed in 10% neutral buffered formalin for about 4 years, whereas the reactivity of Mabs 13A5 and 18C4 and polyclonal antibody was reduced significantly by long-term formalin fixation. Immunohistochemical examination of Malaysian farm pig samples with Mab 11F6 confirmed the affinity of Nipah virus for respiratory epithelium, renal glomerular and tubular epithelium, meningeal arachnoidal cells, and systemic vascular endothelium and smooth muscle. In addition, Nipah virus antigens were identified in laryngeal epithelial cells, Schwann cells of peripheral nerve fascicles in the spleen, and endothelial cells in the atrioventricular valve. The study demonstrated the value of Mabs 11F6 and 12A5 for the immunohistochemical diagnosis of Nipah virus infection in pigs. PMID- 15276860 TI - Canine lymphomas: a morphological and immunohistochemical study of 55 cases, with observations on p53 immunoexpression. AB - The purpose was to determine the immunophenotype of canine lymphomas (CLs) classified according to the WHO nomenclature for domestic animals, and to relate these findings to the immunoexpression of p53 protein. Lymphomas were immunophenotyped with antibodies to CD79a, CD3, and p53 protein, suitable for paraffin wax-embedded tissue sections. Of 55 cases, 40 (72.7%) were of the B-cell phenotype, 12 (21.8%) of the T-cell phenotype, and three (5.4%) were non-B-non-T lymphomas. Of the 40 B-cell lymphomas, 31 were of the large B-cell type, six were lymphoplasmacytic, one lymphocytic, one follicular (grade II) and one unclassified low-grade. One of the peripheral T-cell lymphomas was morphologically consistent with anaplastic large cell lymphoma. Immunolabelling for p53 occurred in 24 B-cell and seven T-cell lymphomas. Lymphomas with high grade histology showed a significantly increased frequency of p53 positivity (P = 0.01). Positivity for p53 (more than 10% positive cells) tended to be associated with the T-cell phenotype (P = 0.06). Mean patient age was significantly higher in p53-positive cases (P = 0.02). These data are comparable with findings in human lymphomas. PMID- 15276861 TI - Immunohistochemical diagnosis of mouse hepatitis virus and mycoplasma pulmonis infection with murine antiserum. AB - This study established a modified alkaline phosphatase-labelled avidin-biotin complex (ABC-AP) method for diagnosis of mouse hepatitis virus (MHV) and Mycoplasma pulmonis infection from formalin-fixed, paraffin wax-embedded sections, murine antibody-positive serum being used as the primary reagent. With this method, MHV antigen in cAnNCrj.Cg-Foxn1(nu)/Foxn1(nu) mice and M. pulmonis antigen in Wistar rats were immunolabelled in tissue sections. MHV antigen was clearly detected in samples of liver, stomach, caecal and colonic mucosa, and spleen. M. pulmonis antigen was demonstrated on the luminal surface of bronchiolar epithelial cells. This method may prove useful in diagnosis when commercial antisera are unavailable or when immunosuppression prevents serological diagnosis. PMID- 15276862 TI - Characterization of macrophages and occurrence of T cells in intestinal lesions of subclinical paratuberculosis in goats. AB - The granulomatous lesions of subclinical paratuberculosis of goats were examined with emphasis on phenotypic characteristics of macrophages and the presence of different subpopulations of T cells. The macrophages in the granulomatous lesions were morphologically homogeneous in histological sections but showed varying expression of the macrophage marker CD68 (a glycoprotein found mainly in late endosomal and lysosomal membranes) and varying acid phosphatase activity. The lesional macrophages showed decreased expression of complement receptor 3 and major histocompatibility complex proteins, which are markers associated with phagocytosis and antigen-presentation, respectively. The granulomas showed low proliferation activity as measured by the proliferation-associated protein Ki-67, indicating that most cells were recruited to the lesions. Few apoptotic cells were demonstrated by the TUNEL technique, suggesting a low cell turnover in the lesions. CD4(+) T cells constituted the main T-cell population among the CD68(+) macrophages in the granulomatous lesions, and few CD8(+) T cells and gamma delta T cells were observed within the lesions, suggesting the limited ability of these cells to influence the granulomatous lesions in caprine subclinical paratuberculosis. Both WC1(+) and WC1(-) gamma delta T cells were present in the small intestinal wall, but the latter were the more numerous. No difference in the numbers of these cells was observed between the subclinically infected animals and control animals. PMID- 15276863 TI - Chromosome aberrations in cattle with chronic enzootic haematuria. AB - Chromosomal aberrations were investigated in 56 cattle with chronic enzootic haematuria (CEH) raised on pastures giving access to bracken fern. Of these animals, 27 were slaughtered and showed neoplastic lesions of the urinary bladder. Tumour tissue from 11 of the 27 cattle contained bovine papillomavirus type 2 (BPV-2) DNA. Increased numbers of chromosomal aberrations were seen in all animals with CEH, as compared with 30 control cattle that had had no access to bracken fern. The highest clastogenic effect was observed in cattle with urinary bladder cancer and evidence of BPV-2 DNA, suggesting that BPV-2 and bracken fern act synergistically in the production of chromosomal instability. In 19 of 20 animals with CEH, two bracken fern toxic compounds (quercitin and ptaquiloside) were demonstrated in urine, serum and milk. PMID- 15276864 TI - Foci of altered hepatocytes in a fox squirrel (Sciurus niger) with haemochromatosis. AB - Numerous minute milky white foci were distributed throughout the dark brown liver in an adult male fox squirrel. Histologically, the hepatic focal lesions were composed of large eosinophilic granular hepatocytes, which were mostly positive for glutathione S-transferase mu antigen and proliferating cell nuclear antigen. Electron microscopy demonstrated an increased number of mitochondria. These features corresponded to those in the eosinophilic type of foci of altered hepatocytes. Berlin blue stain showed severe haemosiderin deposition in hepatocytes, except in the focal lesions. Since the fox squirrel is known to be liable to develop congenital porphyria, it is suggested that the hepatic anomalies described may be closely associated with the development of porphyria. PMID- 15276865 TI - Immunohistochemical expression of galectin-3 in canine mammary tumours. AB - Galectin-3, an endogenous beta-galactoside-binding protein, plays a significant role in cell growth, cell activation, and cell to extracellular matrix interactions. To define the role of galectin-3 in canine mammary tumours, its expression was determined in mammary samples by immunohistochemical methods. In adjacent normal mammary glands, distant from the neoplasm, mammary epithelial cells showed little or no expression of galectin-3. In the majority of adenomas, the neoplastic cells showed moderate to strong galectin-3 immunoreactivity, but in the majority of adenocarcinomas such immunoreactivity was weak. These results suggest that the progression of canine mammary tumours is associated with low galectin-3 expression. PMID- 15276866 TI - Meningeal carcinomatosis in two cats. AB - Multifocal to diffuse meningeal infiltration by neoplastic epithelial cells was observed in two aged cats with neurological signs and a history of surgical ablation of the auricular pinnae because of the presence of squamous cell carcinoma. In both cats, examination of the external ear canals revealed neoplastic lesions consistent with squamous cell carcinoma, but no changes of the tympanic bullae were noted. In one cat, post-mortem examination revealed marked thickening of the dura mater and infiltration of the arachnoid layer by cytokeratin-positive, neoplastic epithelial cells. In the other cat, no macroscopic brain lesions were noted, but multifocal dissemination of neoplastic epithelial cells to the leptomeninges was observed histologically. Several pathways by which neoplastic cells can reach the meninges have been suggested and haematogenous dissemination was considered most likely in these cats. Squamous cell carcinoma is the most common cutaneous malignant neoplasm in cats and meningeal carcinomatosis can be considered a rare complication. PMID- 15276867 TI - Inhibition of proliferation and induction of apoptosis by 2-benzoyl-3-phenyl-6,7 dichloroquinoxaline 1,4-dioxide in adult T-cell leukemia cells. AB - Human T-cell lymphotrophic virus type-1 (HTLV-1) is a retrovirus which causes adult T-cell leukemia (ATL), an aggressive malignancy of activated T-cells. So far, there is no proven therapy for this disease. The compound 2-benzoyl-3-phenyl 6,7-dichloro quinoxaline 1,4-dioxide (DCQ) has been shown to exhibit a wide range of antibacterial activities and to induce antiproliferation and apoptosis of human colon cancer cell lines. In the present study, we investigated the in vitro effects of DCQ in HTLV-1 positive (C91-PL and HuT-102) and negative (CEM and Jurkat) malignant T-cells. The results indicate that DCQ induced growth inhibition in all four cell lines examined in a dose-dependent manner. The inhibitory effect was mainly due to the induction of apoptosis which was verified by flow cytometry analyses and ELISA-based apoptosis assays. The role of transforming growth factor (TGF) in mediating the antiproliferative and apoptotic effects of DCQ in ATL cells was investigated. Interestingly, in three of the four cell lines used, DCQ increased the TGF-beta1 transcript levels and decreased TGF alpha mRNA, but did not induce changes in TGF-beta2 expression. DCQ treatment also induced an upregulation of p53 and p21 protein levels, key mediators of cell cycle arrest and apoptosis. The anti-apoptotic Bcl-2alpha protein level was found to be reduced. These findings indicate that DCQ inhibits the growth of ATL cell lines, at least in part, by inducing apoptosis mediated by the modulation of TGF expression, the upregulation in p53 and p21 proteins and downregulation in Bcl 2alpha expression. The present findings suggest that DCQ merits further investigation as a potential therapeutic agent for this incurable disease. PMID- 15276868 TI - Copper induces histone hypoacetylation through directly inhibiting histone acetyltransferase activity. AB - The abnormal accumulation of Cu2+ is closely correlated with the incidence of different diseases, such as Alzheimer's disease and Wilson disease. To study in vivo functions of Cu2+ will lead to a better understanding of the nature of these diseases. In the present study, effect of Cu2+ on histone acetylation was investigated in human hepatoma cells. Exposure of cells to Cu2+ resulted in a significant decrease of histone acetylation, as indicated by the decrease of the overall histone acetylation and the decrease of histone H3 and H4 acetylation. Since histone acetyltransferase (HAT) and histone deacetylase (HDAC) are the enzymes controlled the state of histone acetylation in vivo, we tested their contribution to the inhibition of Cu2+ on histone acetylation. One hundred nanomolar trichostatin A, the specific inhibitor of HDAC, did not attenuate the inhibitory effect of Cu2+ on histone acetylation. Combined with that Cu2+ showed no effect on the in vitro activity of HDAC, these results led to the conclusion that it is HAT, but not HDAC that is involved in Cu2+ -induced histone hypoacetylation. This conclusion was confirmed by the facts that (1) Cu2+ significantly inhibited the in vitro activity of HAT, (2) Cu2+ -treated cells possessed a lower HAT activity than control cells, and (3) 50 or 100 microM bathocuproine disulfonate, a chelator of Cu2+, significantly attenuated the inhibition of Cu2+ on HAT activity and histone acetylation in the similar pattern. Combined with that Cu2+ showed no or obvious cytotoxicity at 100 or 200 microM in human hepatoma cells, and the previous study that Cu2+ inhibits the histone H4 acetylation of yeast cells at nontoxic or toxic levels, the data presented here suggest that inhibiting histone acetylation is probably one general in vivo function of Cu2+, where HAT is its molecular target. PMID- 15276869 TI - The antioxidant and antifibrogenic effects of the glycosaminoglycans hyaluronic acid and chondroitin-4-sulphate in a subchronic rat model of carbon tetrachloride induced liver fibrogenesis. AB - Hepatic fibrosis involves the interplay of many factors including reactive oxygen species. Recent reports described antioxidant properties of glycosaminoglycans (GAGs). Since several findings have shown that hyaluronic acid (HYA) and chondroitin-4-sulphate (C4S) may act as antioxidant molecules, the aim of this research was to evaluate the antioxidant effects of HYA and C4S treatment in a rat model of liver fibrosis. The effect on tissue inhibitors of metalloproteinases (TIMPs) was also studied. Liver fibrosis was induced in rats by eight intraperitoneal injections of CCl4, twice a week for 6 weeks. HYA or C4S alone (25 mg/kg) or HYA and C4S in combination (12.5 + 12.5 mg/kg) were administered daily by the same route during the 6 weeks. At the end of the 6-week treatment period (24 h after the last dose of GAGs), the following parameters were evaluated: (1) serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities, as index of hepatic cell disruption; (2) hepatic conjugated dienes (CD), as index of lipid peroxidation; (3) hepatic TIMPs activity and expression; (4) hepatic superoxide dismutase (SOD) and glutathione peroxidase (GPx) activity, as index of endogenous defences; (5) hepatic hydroxyproline, as index of collagen deposition. CCl4-induced liver fibrosis enhanced lipid peroxidation and TIMPs activation, increased ALT and AST, depleted antioxidants SOD and GPx, and caused collagen deposition in liver tissue. Treatment with GAGs, especially when in combination, successfully reduced ALT and AST rise, lipid peroxidation by evaluating conjugated dienes, TIMPs activation and mRNA expression, partially restored SOD and GPx activities, and limited collagen deposition in the hepatic tissue. The data obtained showed that these molecules were able to limit hepatic injury induced by chronic CCl4 intoxication and especially limited liver fibrosis. They also confirm that HYA and C4S may exert antioxidant mechanism, while reduction of TIMPs expression suggests that GAGs may influence MMPs and TIMPs imbalance in liver fibrosis. PMID- 15276870 TI - Protective effect of fosfomycin on gentamicin-induced lipid peroxidation of rat renal tissue. AB - Fosfomycin is clinically recognized to reduce the aminoglycoside antibiotics induced nephrotoxicity. However, little has been clarified why fosfomycin protects the kidney from the aminoglycosides-induced nephrotoxicity. Gentamicin, a typical aminoglycoside, is reported to cause lipid peroxidation. We focused on lipid peroxidation induced by gentamicin as a mechanism for the aminoglycosides induced nephrotoxicity. The aim of this study is to investigate the effect of fosfomycin on the gentamicin-induced lipid peroxidation. In rat renal cortex mitochondria, fosfomycin was shown to depress the gentamicin-induced lipid peroxidation, which was evaluated by formation of thiobarbituric acid reactive substances (TBARS). Interestingly, this effect was observed in rat renal cortex mitochondria, but not in rat liver microsomes. However, fosfomycin did not affect lipid peroxidation of arachidonic acid caused by gentamicin with iron. Fosfomycin inhibited the gentamicin-induced iron release from rat renal cortex mitochondria. These results indicated that fosfomycin inhibited the gentamicin-induced lipid peroxidation by depressing the iron release from mitochondria. This may possibly be one mechanism for the protection of fosfomycin against the gentamicin-induced nephrotoxicity. PMID- 15276871 TI - Protection of tamoxifen against oxidation of mitochondrial thiols and NAD(P)H underlying the permeability transition induced by prooxidants. AB - The effects of tamoxifen (TAM) were studied on the mitochondrial permeability transition (MPT) induced by the prooxidant tert-butyl hydroperoxide (t-BuOOH) or the thiol cross-linker phenylarsine oxide (PhAsO), in the presence of Ca2+, in order to clarify the mechanisms involved in the MPT inhibition by this drug. The combination of Ca2+ with t-BuOOH or PhAsO induces mitochondrial swelling and depolarization of membrane potential (deltapsi). These events are inhibited by cyclosporine A (CyA), suggesting the inhibition of the MPT. The pre-incubation of mitochondria with TAM also prevents those events and induces a time-dependent reversal of deltapsi depolarization following MPT induction, similarly to CyA. Moreover, TAM inhibits the Ca2+ release and the oxidation of NAD(P)H and protein thiol (-SH) groups promoted by t-BuOOH plus Ca2+. On the other hand, the MPT induced by PhAsO plus Ca2+ does not induce -SH groups oxidation, supporting the notion that MPT induction by this compound is not mediated by the oxidation of specific membrane proteins groups. However, TAM also inhibits the PhAsO induced MPT, suggesting that this drug may inhibit this phenomenon by inhibiting PhAsO binding to -SH vicinal groups, implicated in the MPT induction. These data indicate that the MPT inhibition by TAM may be related to its antioxidant capacity in preventing the oxidation of NAD(P)H and -SH groups or by blocking these groups, since the oxidation of these groups increases the sensitivity of mitochondria to the MPT induction. Additionally, they suggest an MPT-independent pathway for TAM-induced apoptosis and a potential ER-independent mechanism for the effectiveness of this drug in the cancer therapy and prevention. PMID- 15276872 TI - An investigation of the genotoxic effects of N-nitrosomorpholine in mammalian cells. AB - N-nitrosomorpholine (NMOR) is a well-known hepatocarcinogen. Since this compound is representative of the group of indirect-acting N-nitrosamines, its metabolic activation should be essential. However, the mechanism of NMOR-induced carcinogenesis is still not completely clear. In this paper we tried to further our understanding of the genotoxic effects of NMOR. The central aim of this study was to elucidate to what extent NMOR requires metabolic activation. For evaluation of the mutagenicity of NMOR, V79 cells were used either in the presence or absence of the microsomal S9 fraction in the mutation assay and formation of reactive oxygen/nitrogen species (ROS/RNS) in Caco-2 cells treated with NMOR was measured by a fluorescent assay. A very weak rise of 6-thioguanine resistant mutations was observed in both NMOR-treated model cells, V79/-S9 and V79/+S9. A significant difference between the level of mutations in V79/-S9 and V79/+S9 cells was recorded on the 7th day of expression only. Data obtained by the fluorescent assay confirmed that NMOR caused generation of ROS/RNS. In summary, the presented results showed that NMOR might induce DNA damage not only indirectly by its activation by drug-metabolizing enzymes but also via direct formation of ROS/RNS. PMID- 15276873 TI - Immunocytochemical localization of GABABR1 receptor subunits in the basolateral amygdala. AB - Gamma-aminobutyric acid B (GABAB) receptors (GBRs) are G-protein-coupled receptors that mediate a slow, prolonged form of inhibition in the basolateral amygdala (ABL) and other brain areas. Recent studies indicate that this receptor is a heterodimer consisting of GABABR1 (GBR1) and GABABR2 subunits. In the present investigation, antibodies to the GABABR1 subunit were used to study the neuronal localization of GBRs in the rat ABL. GBR immunoreactivity was mainly found in spine-sparse interneurons and astrocytes at the light microscopic level. Very few pyramidal neurons exhibited perikaryal staining. Dual-labeling immunofluorescence analysis indicated that each of the four main subpopulations of interneurons exhibited GBR immunoreactivity. Virtually 100% of large CCK+ neurons in the basolateral and lateral nuclei were GBR+. In the basolateral nucleus 72% of somatostatin (SOM), 73% of parvalbumin (PV) and 25% of VIP positive interneurons were GBR+. In the lateral nucleus 50% of somatostatin, 30% of parvalbumin and 27% of VIP positive interneurons were GBR+. Electron microscopic (EM) analysis revealed that most of the light neuropil staining seen at the light microscopic level was due to the staining of dendritic shafts and spines, most of which probably belonged to spiny pyramidal cells. Very few axon terminals (Ats) were GBR+. In summary, this investigation demonstrates that the distal dendrites of pyramidal cells, and varying percentages of each of the four main subpopulations of interneurons in the ABL, express GBRs. Because previous studies suggest that GBR-mediated inhibition modulates NMDA-dependent EPSPs in the ABL, these receptors may play an important role in neuronal plasticity related to emotional learning. PMID- 15276874 TI - Pharmacological characterisation of the rat brachial plexus avulsion model of neuropathic pain. AB - Recently, our laboratory has proposed the avulsion of rat brachial plexus as a new and reliable model for the study of neuropathic pain. In this model, the neuropathy can be detected even at distant sites from the injury, both in ipsilateral and contralateral hindpaws. The purpose of this study was to pharmacologically characterise this behavioural model of persistent peripheral neuropathic pain by assessing the effects of several analgesic drugs currently used in clinical practice. For this purpose, the effects of these drugs on the mechanical and cold allodynia were analysed 20-40 days after rat brachial plexus avulsion. Injection of saline, administered by the same route as the other drugs, did not significantly affect the nociceptive threshold either in sham-operated or in neuropathic rats. However, administration of the opioid analgesic morphine (5 mg/kg, s.c.), the alpha2 adrenoceptor agonist clonidine (300 microg/kg, i.p.), the NMDA receptor antagonist ketamine (25 mg/kg, i.p.) or the anticonvulsant drug gabapentin (70 mg/kg, p.o.) consistently reduced both mechanical and cold allodynia following avulsion of rat brachial plexus. The administration of the selective COX-2 inhibitor celecoxib (10 mg/kg, p.o.) blocked mechanical allodynia, but not cold allodynia, whereas the sodium channel blocker lidocaine (40 mg/kg, i.p.) attenuated only cold allodynia. The non-steroidal anti inflammatory drug diclofenac (100 mg/kg, i.p.), the steroidal anti-inflammatory dexamethasone (1.5 mg/kg, i.p.) and the antidepressant imipramine (10 mg/kg, i.p.) all failed to significantly attenuate both mechanical and cold allodynia in the rats following avulsion of brachial plexus. These findings suggest that avulsion-associated mechanical and cold allodynia, two classic signs of persistent neuropathic pain, were consistently prevented by several analgesics currently available in clinical practice, namely morphine, clonidine, ketamine and gabapentin, and to a lesser extent by celecoxib and lidocaine. Therefore, this new proposed model of persistent nociception seems to be suitable for the study of the underlying mechanisms involved in neuropathic pain and for the identification of potential clinically relevant drugs to treat this aspect of peripheral neuropathy. PMID- 15276875 TI - Brain Na+,K+-ATPase isozyme activity and protein expression in ouabain-induced hypertension. AB - In normotensive rats, chronic infusion of exogenous ouabain causes hypertension involving central mechanisms. To determine whether ouabain-induced hypertension is associated with specific changes in brain Na+,K+-ATPase activity and expression, we assessed brain Na+,K+-ATPase isozyme activity and protein expression in rats treated with ouabain (50 microg/day s.c. or 10 microg/day i.c.v. for 14 days). Resting mean arterial pressure (MAP) was higher in s.c.- and i.c.v.-ouabain-treated animals vs. control (124+/-2 vs. 105+/-2 and 130+/-2 vs. 109+/-2, respectively, p<0.01). Ouabain infused s.c. or i.c.v. for 14 days had no effect on Na+,K+-ATPase isozyme activity in hypothalamic, pontine/medullary or cortical microsomes. However, the percent increase in total Na+,K+-ATPase activity produced in vitro by antibody Fab fragments that bind ouabain with high affinity (Digibind) was two-fold greater in s.c.- and i.c.v.-ouabain-treated rats vs. control, but only in hypothalamic microsomes. Thus, ouabain infused s.c. or i.c.v. does appear to directly inhibit Na+,K+-ATPase activity in the hypothalamus. On the other hand, in the hypothalamus, s.c.- and i.c.v.-ouabain infusions tended to increase alpha3 (by 30-44%), but had no effect on alpha1 or alpha2 Na+,K+-ATPase isozyme protein expression. In addition, ouabain was found to partially dissociate from the Na+,K+-ATPase enzyme following sample processing. Thus, the inability to detect a decrease in enzyme activity in the hypothalamus in response to ouabain may be due, in part, to an increase in enzyme expression and the dissociation of ouabain during sample processing. PMID- 15276876 TI - Cocaine and kappa-opioid withdrawal in Planaria blocked by D-, but not L-, glucose. AB - Planarians (Dugesia dorotocephala) that were exposed for 1 h to cocaine (80 microM) or to the kappa-selective opioid receptor agonist U-50,488H (1 microM) displayed an abstinence-induced withdrawal syndrome, indicative of the development of physical dependence, when they were tested in cocaine- (or U 50,488H-) free water, but not when they were tested in cocaine- (or U-50,488H-) containing water. The withdrawal was manifested as a significant (P<0.05) decrease in the rate of planarian spontaneous locomotor activity over a 5-min observation period, using a recently designed metric. Co-exposure of the planarians to D-glucose (1 microM) or to 2-deoxy-D-glucose (2-DG, 1 microM), but not to L-glucose (1 microM), significantly attenuated (P<0.05) the development of physical dependence, shown by an attenuated withdrawal syndrome, from cocaine and U-50,488H. These results suggest that either D-glucose and 2-deoxy-D-glucose compete with a common cocaine and kappa-opioid transport mechanism or that the development of physical dependence (or the inhibition of abstinence-induced withdrawal) in planarians requires energy supplied from glucose metabolism. PMID- 15276877 TI - Neuroprotective effects of olanzapine on methamphetamine-induced neurotoxicity are associated with an inhibition of hyperthermia and prevention of Bcl-2 decrease in rats. AB - It is hypothesized that atypical antipsychotic drugs have neuroprotective effects which may be one of the mechanisms in treatment of schizophrenia. We investigated the neuroprotective effects of olanzapine (OLA), an atypical antipsychotic drug, on methamphetamine (METH)-induced neurotoxicity in rats. After pretreatment with OLA (2 mg/kg/day) by intraperitoneal injection for 2 weeks, rats were administered METH (7.5 mg/kg, four times at 2-h intervals) by subcutaneous injection while their body temperature was monitored. The rats were sacrificed 24 h after the last injection of METH for immunohistochemistry. METH-induced 24 h mortality was effectively reduced and METH-induced decrease of tyrosine hydroxylase immunoreactivity in caudate putamen (CPu) was significantly attenuated by OLA chronic pretreatment. Furthermore, we showed that the above neuroprotective potential of OLA might be associated with its attenuating effects on METH-induced hyperthermia and with its preventative actions on METH-induced decrease of Bcl-2, an anti-apoptotic gene product, in the CPu. Our results suggest that OLA may be a neuroprotective agent and that its neuroprotective potential may contribute to its therapeutic effects in treatment of schizophrenia. PMID- 15276878 TI - Upregulation of corticotropin-releasing hormone gene expression in the paraventricular nucleus of cyclophosphamide-induced cystitis in male rats. AB - We examined the effects of cyclophosphamide (CP)-induced cystitis on the expression of corticotropin-releasing hormone (CRH) mRNA in the paraventricular nucleus (PVN) and the serum levels of adrenocorticotropic hormone (ACTH) using in situ hybridization histochemistry and radioimmunoassay. In addition, the expression of AVP heteronuclear (hn) RNA and neuronal nitric oxide synthase (nNOS) mRNA was also examined in the PVN of a CP-induced cystitis model. We found that the levels of CRH mRNA were significantly increased in the PVN at 2 h after intraperitoneal administration of CP compared to those in saline-treated rats. The CRH mRNA levels in the PVN peaked at 12 h after CP administration and the levels were still significantly higher than those in saline-treated group at 24 h after CP administration. The serum ACTH levels in CP-treated group were also significantly higher compared to those in saline-treated group at any of the time points examined. Unlike previous findings showing upregulation of nNOS mRNA and AVP hnRNA under somatic nociceptive states, the levels of nNOS mRNA and AVP hnRNA were unchanged in the PVN following CP-induced cystitis, visceral nociceptive stimulation. These results suggest that visceral nociceptive stimulation as well as somatic nociceptive stimulation may activate the hypothalamo-pituitary axis but the hypothalamic neuroendocrine responses produced by visceral nociceptive stimulation may be different from those produced by somatic nociceptive stimulation. PMID- 15276879 TI - Fluoxetine inhibits A-type potassium currents in primary cultured rat hippocampal neurons. AB - The effects of fluoxetine (Prozac) on the transient A-currents (IA) in primary cultured hippocampal neurons were examined using the whole-cell patch clamp technique. Fluoxetine did not significantly decrease the peak amplitude of whole cell K+ currents, but it accelerated the decay rate of inactivation, and thus decreased the current amplitude at the end of the pulse. For further analysis, IA and delayed rectifier K+ currents (IDR) were isolated from total K+ currents. Fluoxetine decreased IA (the integral of the outward current) in a concentration dependent manner with an IC50 of 5.54 microM. Norfluoxetine, the major active metabolite of fluoxetine, was a more potent inhibitor of IA than was fluoxetine, with an IC50 of 0.90 microM. Fluoxetine (3 microM) inhibited IA in a voltage dependent manner over the whole range of membrane potentials tested. Analysis of the time dependence of inhibition gave estimates of 34.72 microM(-1) s(-1) and 116.39 s(-1) for the rate constants of association and dissociation, respectively. The resulting apparent Kd was 3.35 microM, similar to the IC50 value obtained from the concentration-response curve. In current clamp configuration, fluoxetine (3 microM) induced depolarization of resting membrane potential and reduced the rate of action potential. Our results indicate that fluoxetine produces a concentration- and voltage-dependent inhibition of IA, and that this effect could affect the excitability of hippocampal neurons. PMID- 15276880 TI - Proprioceptive control of human wrist extensor motor units during an attention demanding task. AB - The responsiveness of the tonically firing single motor units (SMU) to Ia afferent volleys elicited by either mechanical (T-reflex) or electrical nerve stimulation (H-reflex) was tested in the extensor carpi radialis muscle (ECR) while the subjects were maintaining a steady wrist extension force using visual feedback set either at low or high gain. The aim was to determine whether the proprioceptive control of tonic motoneuronal activity depends on the level of attentiveness required by the behavioural context. The response probability of the SMUs to tendon taps was significantly higher (p<0.0001) and that to electrical nerve stimulation was lower (p<0.001) during the more demanding task. Since these changes in SMU responsiveness were not accompanied by any differences in either the motor unit firing patterns or the mean levels of EMG muscle activity, it can be concluded that there were no attention-related changes in the net excitatory drive to the ECR motoneurons. These results are consistent with the idea that fusimotor sensitization of the muscle spindle may have occurred in the more demanding task: an increase in the mechanical sensitivity of the muscle spindles would certainly account for both the T-reflex facilitation and the H reflex depression observed. The attention-demanding task therefore seemed to involve an independent fusimotor drive activation process. The results of this study suggest that an adaptation of the fusimotor system occurs in humans, depending on the levels of attention and accuracy required to perform the ongoing motor task, as previously reported to occur in animals. PMID- 15276881 TI - RU486 blocks fasting-induced decrease of neuronal nitric oxide synthase in the rat paraventricular nucleus. AB - It has been reported that food deprivation decreases expression of neuronal nitric oxide synthase (nNOS) in the hypothalamic paraventricular nucleus (PVN). Food deprivation produces autonomic changes and the PVN nitric oxide has been suggested to be involved in regulation of autonomic functions. In order to understand the molecular mechanism by which food deprivation decreases nNOS expression in the PVN, we examined if plasma glucocorticoids, which reported to be elevated during food deprivation, mediates the fasting-induced down-regulation of the PVN-nNOS. Male Sprague-Dawley rats underwent 48 h of food deprivation, but not water deprivation, with/without subcutaneous RU486, glucocorticoid receptor antagonist, and the brain tissues were processed for immunohistochemistry with specific antibodies against nNOS. Immunoreactivity of phosphorylated cAMP response element-binding protein (pCREB) was also examined in the PVN sections, because nNOS promoter carries cAMP response element (CRE). Food deprivation significantly decreased both nNOS and pCREB immunoreactivity (-ir) in the medial parvocellular PVN, and RU486 blocked this reduction. In the posterior magnocellular PVN, nNOS-ir, but not pCREB-ir, was decreased by food deprivation, and RU486 exerted no effect. These results suggest that glucocorticoid receptor may mediate the fasting-induced down-regulation of nNOS in the parvocellular PVN, but not in the magnocellular PVN. PMID- 15276882 TI - 5-HT2A/2C receptor and 5-HT transporter densities in mice prone or resistant to chronic high-fat diet-induced obesity: a quantitative autoradiography study. AB - The present study examined the density of 5-HT2A/2C receptors and 5-HT transporters in the brains of chronic high-fat diet-induced obese (cDIO) and obese-resistant (cDR) mice. Thirty-five male mice were used in this study. Twenty eight mice were fed with a high-fat diet (40% of calories from fat) for 6 weeks and then classified as the cDIO (n=8) or cDR (n=8) mice according to the highest and lowest body weight gainers. Seven mice were placed on a low-fat diet (LF: 10% of calories from fat) and were used as controls. After 20 weeks of feeding, the sum of epididymal, perirenal, omental and inguinal fat masses was 9.3+/-0.3 g in the cDIO group versus 3.1+/-0.5 g in the cDR (p<0.005) and 1.5+/-0.1 g in the LF (p<0.001) groups. Using quantitative autoradiography techniques, the binding site densities of 5-HT2A/2C receptors and 5-HT transporters were measured in multiple brain sections of mice from the three groups. Most regions did not differ between groups but, importantly, the cDIO mice had a significantly higher 5-HT2A/2C binding density in the anterior olfactory nucleus and ventromedial hypothalamic nucleus (VMH) compared to the cDR and LF mice (+39% and +47%, p=0.003 and 0.045, respectively), whereas the latter two groups did not differ. The density of 5 HT2A/2C receptors in the VMH was associated with total amount of fat mass (r=0.617, p=0.032). On the other hand, the cDR mice had significantly lower 5-HT transporter binding than the cDIO and LF mice, respectively, in the nucleus accumbens (-44%, -38%, both p<0.02), central nucleus of the amygdaloid nucleus ( 40%, -44%, p=0.003 and 0.009), and olfactory tubercle nucleus (-42%, -42%, both p=0.03). In conclusion, this study has demonstrated differentially regulated levels of the 5-HT2A/2C receptor and 5-HT transporter in specific brain regions of the cDIO and cDR mice. It provides neural anatomical bases by which genetic variability in 5-HT2A/2C receptors and 5-HT transporter may influence satiety and sensory aspects of energy balance. PMID- 15276883 TI - Increased BBB permeability by parasympathetic sphenopalatine ganglion stimulation in dogs. AB - The blood-brain barrier (BBB) is a major obstacle for movement of large molecules to and from the brain. Stimulation of the sphenopalatine ganglion (SPG), the major source of parasympathetic innervation to brain vasculature, is known to vasodilate brain vessels, and has recently been shown to also increase the permeability of the BBB in the rat. In this work, we studied the effect of SPG stimulation on BBB permeability in larger animals--Beagle dogs. Left SPG was exposed by lateral approach in five Beagle dogs, and stimulated at 10 Hz. FITC labeled 10 kDa dextran was continuously infused to the left atrium during stimulation, and cerebral angiography was periodically obtained via the vertebral artery. Three control dogs received labeled dextran, without SPG exposure or stimulation. Brains were perfused with saline thoroughly at the end of stimulation, and samples from various regions were taken for fluorescence reading of tissue homogenates. Cerebral vasodilatation was evidenced in all but one dog, whose fluorescence results were consequently excluded from analysis, assuming that its SPG had been damaged by surgery. Fluorescence was significantly higher in the four stimulated compared to the three non-stimulated animals; e.g. mean FITC-dextran concentration in the anterior brain regions was 0.98+/-0.12 ug (mean+/-S.D.) FITC/g brain for experimental animals, and 0.40+/-0.02 for controls (p<0.01). No effect was seen in the pons and cerebellum (0.68+/-0.22 vs. 0.60+/ 0.03, NS) whose vascular innervation is supplied by the otic rather than the SPG ganglion. SPG stimulation appears to be an effective way to increase BBB permeability, allowing introduction of large molecules to the brain. This could be a therapeutic method for a wide variety of brain disorders, including tumors and neurodegenerative diseases. PMID- 15276884 TI - Extracellular level of basolateral amygdalar dopamine responding to reversal of appetitive-conditioned discrimination in young and old rats. AB - Young and old rats, aged 3 and 24 months old, respectively, were conditioned to press a lever under lamp-on conditions for reward acquisition and lamp-off for no reward using a variable interval reinforcement schedule that averaged 15 s; i.e., the minimal requirement was 4 responses/min. Over a 30-day period, young and old groups showed increased response to lamp-on from 22 to 35/min and from 10 to 23/min, respectively, and shortened response to lamp-off after initial training. Response to lamp-on as a percentage of total response to lamp-on and -off (the discrimination ratio) was over 80%. For the next 30 days, reversal learning was imposed to reinforce discrimination of the lamp-off state. Young rats showed a steadily increasing discrimination ratio from 40% to 70%, and old rats from 30% to 60%. In comparison with the initial training, young rats showed a total response increase from 50% to 60%, while old rats showed an approximately 5% decrease without loss of reward-obtaining efficiency. In vivo microdialysis during reversal revealed that young rats had higher dopamine transmission in the basolateral amygdala than old rats. The dopamine level was positively correlated with the number of responses to state of reward in young rats and negatively with the numbers of both NRF and RF responses to lamp-on and -off states in old rats. These results suggest that in reversal discrimination, basolateral amygdalar dopamine efflux correlates with the manner of age-related conditioned response rather than the ability to learn. PMID- 15276885 TI - Responses of feline medial medullary reticular formation neurons with projections to the C5-C6 ventral horn to vestibular stimulation. AB - Prior studies have shown that the vestibular system contributes to adjusting respiratory muscle activity during changes in posture, and have suggested that portions of the medial medullary reticular formation (MRF) participate in generating vestibulo-respiratory responses. However, there was previously no direct evidence to demonstrate that cells in the MRF relay vestibular signals monosynaptically to respiratory motoneurons. The present study tested the hypothesis that the firing of MRF neurons whose axons could be antidromically activated from the vicinity of diaphragm motoneurons was modulated by whole-body rotations in vertical planes that stimulated vestibular receptors, as well as by electrical current pulses delivered to the vestibular nerve. In total, 171 MRF neurons that projected to the C5-C6 ventral horn were studied; they had a conduction velocity of 34+/-15 (standard deviation) m/sec. Most (135/171 or 79%) of these MRF neurons lacked spontaneous firing. Of the subpopulation of units with spontaneous discharges, only 3 of 20 cells responded to vertical rotations up to 10 degrees in amplitude, whereas the activity of 8 of 14 neurons was affected by electrical stimulation of the vestibular nerve. These data support the hypothesis that the MRF participates in generating vestibulo-respiratory responses, but also suggest that some neurons in this region have other functions. PMID- 15276886 TI - Effect of lambda-carrageenan-induced inflammatory pain on brain uptake of codeine and antinociception. AB - This study investigated the potential clinical implications of lambda-carrageenan induced inflammatory pain on brain uptake of a commonly used analgesic, codeine, in relation to the fundamental properties of the blood-brain barrier (BBB) correlated to its antinociceptive profile over a 168-h time course. BBB uptake of [14C]sucrose (a membrane impermeant marker) and [3H]codeine were investigated using an in situ brain perfusion model in the rat. Results demonstrated a significantly increased brain uptake of [14C]sucrose at 1, 3, 6 and 48 h (139+/ 9%, 166+/-19%, 138+/-13% and 146+/-7% compared with control, respectively) and [3H]codeine at 3 and 48 h (179+/-6% and 179+/-12% compared with control, respectively). Capillary depletion analyses ensured that increased radioisotope associated with the brain was due to increased uptake rather than trapping in the cerebral vasculature. Antinociception studies using a radiant-heat tail flick analgesia method demonstrated that lambda-carrageenan-induced inflammatory pain enhanced the in vivo antinociceptive profile of i.p.-administered codeine (7 mg/kg) at 3 and 48 h (144+/-11% and 155+/-9% compared with control, respectively). This study demonstrated that brain uptake and antinociception of codeine are increased during lambda-carrageenan-induced inflammatory pain, suggesting that the presence of inflammatory pain may be an important consideration in therapeutic drug dosing, potential adverse effects and/or neurotoxicity. PMID- 15276887 TI - Injection of orexin A into the diagonal band of Broca induces sympathetic and hyperthermic reactions. AB - This experiment tested the effect of an injection of orexin A into the diagonal band of Broca on the sympathetic activity and body temperature. Concentration of glycerol into white fat of lumbar region, firing rates of sympathetic nerves to interscapular brown adipose tissue (IBAT), IBAT and colonic temperatures, and heart rate were monitored in urethane-anesthetized male Sprague-Dawley rats for 30 min before and 150 min after injections of orexin A (0.4 and 0.7 nmol) into the diagonal band of Broca. The same variables were monitored in control rats with an injection of saline. The results show that orexin A increases glycerol concentration, sympathetic firing rate, IBAT and colonic temperatures, and heart rate. The saline injection did not induce any modification. These findings suggest that the diagonal band of Broca is a cerebral structure involved in the induction of the hyperthermia due to orexin A. PMID- 15276888 TI - Antinociceptive effects of bethanechol or dimethylphenylpiperazinium in models of phasic or incisional pain in rats. AB - The mechanism by which muscarinic or nicotinic agonists produce antinociception has been the subject of several studies. In the present investigation, we used intrathecal administration of drugs to rats to show that muscarinic or nicotinic agonists such as bethanechol (BCh) and dimethylphenylpiperazinium (DM), respectively, dose-dependently increased the tail flick latency and reduced the pain produced by a surgical incision performed on the plantar aspect of a hind paw. The effects of BCh in both tests were inhibited by the previous intrathecal administration of atropine, but not mecamylamine (muscarinic and nicotinic antagonists, respectively). Mecamylamine significantly reduced the effects of DM in both tests. Atropine significantly reduced the effect of DM in the tail flick test and inhibited the effect of DM against the incisional pain. Intrathecal hemicholinium-3 (HC-3), a reversible inhibitor of choline transporter, did not change the effect of BCh in the tail flick test but produced a non-significant reduction of the effect of BCh against incisional pain. In contrast, HC-3 produced a non-significant reduction of the effect of DM in the tail flick test but fully inhibited the effect of DM against incisional pain. Therefore, the BCh induced antinociception depends on a direct activation of muscarinic receptors, whereas DM-induced antinociception results in drug interaction with nicotinic receptors to activate the further release of acetylcholine from intrinsic spinal cholinergic terminals. The acetylcholine released by DM in turn induces antinociception via activation of muscarinic receptors. PMID- 15276890 TI - Influence of type of curriculum on students' perceptions of the medical course: a compilation of results from the Cognitive Behavior Survey, Attitudes Toward Social Issues In Medicine survey, and Learning Environment Questionnaire. AB - BACKGROUND: This study administered selected scales from the Cognitive Behavior Survey, Attitudes Toward Social Issues in Medicine survey, and the Learning Environment Questionnaire. Data were gathered from other medical schools to put results into context. PURPOSE: To present results on these scales from other medical schools and to compare the effects of type of curriculum on the results. METHODS: Articles that had cited the articles that originally presented the scales used were searched for. Results were organized by type of curriculum and effect sizes were calculated where possible. RESULTS: Within the limits of small numbers of studies so far, problem-based learning (PBL) curricula appear to have much more positive effects on students' reported cognitive behaviors than do hybrid curricula. Both have substantial positive effects on students' perceptions of the learning environment (PBL medium to large; hybrid small to medium). Neither appears to have much effect on students' attitudes toward social issues in medicine. CONCLUSION: These data provide a starting point for further study of some of the effects of curricular interventions. PMID- 15276889 TI - AM404 enhances the spontaneous release of L-glutamate in a manner sensitive to capsazepine in adult rat substantia gelatinosa neurones. AB - In 84% of substantia gelatinosa (SG) neurones examined in adult rat spinal cord slices, an anandamide transport inhibitor, AM404, increased the frequency of spontaneous excitatory postsynaptic currents in a manner similar to that of capsaicin. AM404 was without actions in the presence of a vanilloid TRPV1 receptor antagonist, capsazepine. We conclude that AM404 enhances the spontaneous release of L-glutamate by activating TRPV1 receptors in the SG. PMID- 15276891 TI - Attitudes of Chinese medical students toward the global minimum essential requirements established by the Institute for International Medical Education. AB - BACKGROUND: The Institute for International Medical Education has published "Global Minimum Essential Requirements (GMERs) in Medical Education." PURPOSE: This study examined attitudes of a sample of Chinese medical students toward the GMERs. METHODS: Matriculating and graduating West China School of Medicine Sichuan University medical students were administered parallel surveys during the 2001 to 2002 academic years. RESULTS: Both cohorts produced similar response profiles. The majority in both groups rated the 7 GMER domains as either important or very important for their medical education. Matriculating students rated professional values, attitudes, behavior, and ethics as most important, whereas graduating students valued clinical skills highest. Population health and health systems received the lowest importance ratings from both groups. Please note that this study was conducted before the SARS outbreak. As a result of the SARS experience, attitudes toward population health and health systems might have changed. CONCLUSION: Although medical students ascribe importance to the GMERs, efforts are needed to increase the perceived importance of the population health and health systems domain. PMID- 15276892 TI - Evaluation of the effect of a computerized training simulator (ANAKIN) on the retention of neonatal resuscitation skills. AB - BACKGROUND: Neonatal resuscitation knowledge and skills deteriorate after initial training. PURPOSE: To evaluate the effectiveness of a computerized simulator system (ANAKIN) as a means for boosting neonatal resuscitation knowledge, skills, and self-reported confidence beliefs. METHOD: A randomized pretest-posttest control group study design involving 60 3rd-year medical students. At a 4-month, post-training interval, experimental group was exposed to ANAKIN and control group to a training video. Both groups assessed at an 8-month, post-neonatal resuscitation training interval. RESULTS: Knowledge level for both groups decreased significantly at 4- and 8-month, post-training intervals despite booster exposure. Confidence level for both study groups increased significantly following booster exposure. However, no significant difference between study group skill levels at 8 months and no significant relation between neonatal resuscitation knowledge, confidence, or skills. CONCLUSION: Computerized simulator system was as effective as video for maintaining resuscitation skills of medical students, and students were very satisfied with experience of remote computer simulation training. PMID- 15276893 TI - Effect of gender, age, and relevant course work on attitudes toward empathy, patient spirituality, and physician wellness. AB - BACKGROUND: The emphasis in medical education on viewing the patient as a whole person addresses current concerns about the negative impact of standard physician training that may lead to impaired patient-physician relationships. PURPOSES: To assess self-ratings of empathy, spirituality, wellness, and tolerance in a sample of medical students and practitioners to explore differences by gender, age, and training. METHODS: A survey was created that assesses empathy, spirituality, wellness, and tolerance in the medical setting. Surveys were completed anonymously by medical students and practitioners from the medical school. RESULTS: The youngest groups scored highest on empathy and wellness and lowest on tolerance. Participation in medical school wellness sessions correlated with higher empathy and wellness scores; participation in both empathy and spirituality sessions correlated with higher empathy scores. CONCLUSION: Exposure to educational activities in empathy, philosophical values and meaning, and wellness during medical school may increase empathy and wellness in medical practice. PMID- 15276894 TI - The physical examination of patients with abdominal pain: the long-term effect of adding standardized patients and small-group feedback to a lecture presentation. AB - BACKGROUND: One of the most effective methods for teaching physical diagnosis may be standardized patient instructors. PURPOSE: To determine if a lecture plus standardized patient instructors with small-group sessions is more effective than a lecture alone for teaching the evaluation of patients with abdominal pain. METHODS: Control (class of 2001) and intervention (class of 2002) groups both attended a lecture on the abdominal examination. The intervention group then underwent an exercise with standardized patient instructors and a review session with surgical faculty. An evaluation 18 months later used standardized patient instructors to complete evaluations assessing history-taking and physical examination skills. RESULTS: The intervention group performed significantly better than the control group on both the history and the physical examination subscales. CONCLUSION: It is possible to have an important, measurable, and lasting effect on physical examination skills by adding standardized patient instructors and small-group discussion to a lecture presentation. PMID- 15276895 TI - Teaching the musculoskeletal examination: are patient educators as effective as rheumatology faculty? AB - BACKGROUND: Effective education of clinical skills is essential if doctors are to meet the needs of patients with rheumatic disease, but shrinking faculty numbers has made clinical teaching difficult. A solution to this problem is to utilize patient educators. PURPOSE: This study evaluates the teaching effectiveness of patient educators compared to rheumatology faculty using the musculoskeletal (MSK) examination. METHOD: Sixty-two 2nd-year medical students were randomized to receive instruction from patient educators or faculty. Tutorial groups received instructions during three, 3-hr sessions. Clinical skills were evaluated by a 9 station objective structured clinical examination. Students completed a tutor evaluation form to assess their level of satisfaction with the process. RESULTS: Faculty-taught students received a higher overall mark (66.5% vs. 62.1%,) and fewer failed than patient educator-taught students (5 vs. 0, p = 0.02). Students rated faculty educators higher than patient educators (4.13 vs. 3.58 on a 5-point Likert scale). CONCLUSION: Rheumatology faculty appear to be more effective teachers of the MSK physical exam than patient educators. PMID- 15276896 TI - A practical method for increasing scholarly activity in an academic family medicine department. AB - BACKGROUND: Providing a stimulating environment designed to increase scholarly activity is becoming increasingly difficult in today's academic medicine departments. This article was written to communicate methods and results of a practical program designed to increase scholarly activities in an academic medicine department. DESCRIPTION: A straightforward program was developed and implemented with the objective of increasing scholarly activity in an academic medicine department. Concomitant departmental changes in scholarly activities including publications, lecturing, and research were determined over a 3-year period. EVALUATION: At Year 3 post program initiative, there was a significant increase in the number of faculty members publishing (from 15.8%-73.7%). The number of articles submitted to peer-reviewed journals increased from 5 to 44, and the number accepted increased from 8 to 10. The number of national lectures given by faculty increased from 1 to 17, whereas the number of local lectures did not increase. CONCLUSION: A straightforward program can be effectively used to increase scholastic endeavors such as writing and lecturing in an academic setting. PMID- 15276897 TI - Comprehensive assessment of professional competence: the Rochester experiment. AB - BACKGROUND: A required 2-week comprehensive assessment (CA) for 2nd-year medical students that integrates basic science, clinical skills, information management, and professionalism was implemented. DESCRIPTION: The CA links standardized patients (SPs) with computer-based exercises, a teamwork exercise, and peer assessments; and culminates in student-generated learning plans. EVALUATION: Scores assigned by SPs showed acceptable interrater reliability. Factor analyses defined meaningful subscales of the peer assessment and communication rating scales. Ratings of communication skills were correlated with information gathering, patient counseling, and peer assessments; these, in turn, were strongly correlated with the written exercises. Students found the CA fair, with some variability in opinion of the peer and written exercises. Useful learning plans and positive curricular changes were undertaken in response to the CA results. CONCLUSION: A CA that integrates multiple domains of professional competence is feasible, useful to students, and fosters reflection and change. Preliminary data suggest that this format is reliable and valid. PMID- 15276898 TI - Developing an ethics curriculum for an internal medicine residency program: use of a needs assessment. AB - BACKGROUND: Residency programs are required to teach and evaluate trainees in the area of professionalism and medical ethics. Prior to developing a curriculum in this area, residents and fellows were surveyed to assess learning needs. DESCRIPTION: A case-based survey was developed based on published curricula. Residents and fellows were asked to describe their comfort level in 11 clinical scenarios on a Likert-type scale ranging from 1 (not at all comfortable) to 10 (extremely comfortable). EVALUATION: 151 surveys were returned for an overall response rate of 73%. Comfort levels ranged from a low of 3.1 to a high of 8.5 on the 10-point scale. Despite additional years of clinical training, fellows only reported an increased comfort level in 1 case. CONCLUSION: Learning needs exist in residents and fellows in the area of medical ethics. Use of a needs assessment was instrumental in planning and designing an ethics curriculum. PMID- 15276900 TI - Using developmental, cognitive, and neuroscience approaches to understand executive control in young children. AB - The 7 articles in this special issue address the nature of executive control in young children. Executive control is framed in a developmental context, where the unique aspects of cognition in this age range are considered. The set of articles demonstrates the multidisciplinary approaches to study cognition in young children that includes application of cognitive, neuroscience, and developmental paradigms in typically developing youngsters, as well as those affected by clinical conditions, such as traumatic brain injury, exposure to low levels of lead in the environment, and prematurity. Although much work remains to be done, these study results are illustrative of the dynamic work in this exciting developmental period. PMID- 15276901 TI - The Object Classification Task for Children (OCTC): a measure of concept generation and mental flexibility in early childhood. AB - In this study, the development of concept generation and mental flexibility was investigated in 84 Australian children between 3 and 7 years of age, using the Object Classification Task for Children (OCTC), a newly developed executive function test for use with young children. On this task, which was adapted from the Concept Generation Test (Levine, Stuss, & Milberg, 1995) and the Concept Generation Test for Children (Jacobs, Anderson, & Harvey, 2001), children were asked to categorize 6 plastic toys according to 3 predetermined groupings (i.e., color, size, and function). The test included 3 performance levels, each providing increasing levels of structure for the child. Findings from the OCTC show meaningful age-related changes in performance across age groups, with older children being less dependent on additional structure to complete the task, in comparison to younger children. Furthermore, findings from this study suggest that the ability to generate concepts emerges between 3 and 4 years of age, continuing to develop beyond the age of 7 years. A developmental spurt in cognitive flexibility was observed around 4 to 5 years of age, with refinement of this capacity occurring between 5 and 7 years of age. Results suggest that the OCTC is a useful measure of conceptual reasoning skills in early childhood. PMID- 15276902 TI - Executive function in preschool children: examination through everyday behavior. AB - Clinical assessment of executive function in preschool-age children is challenging given limited availability of standardized tasks and preschoolers' variable ability to participate in lengthy formal evaluation procedures. Given the benefits of ecological validity of measuring behavior by rating scales, the Behavior Rating Inventory of Executive Function (Gioia, Isquith, Guy, & Kenworthy, 2000) was modified for use with children ages 2 through 5 years to assess executive functions in an everyday context. The scale development process, based on samples of 460 parents and 302 teachers, yielded a single 63-item measure with 5 related, but nonoverlapping, scales, with good internal consistency and temporal stability. Exploratory factor analyses identified 3 consistent factors: Emergent Metacognition, Flexibility, and Inhibitory Self Control across parent and teacher samples. In a second study with a mixed sample of preschool children with various developmental disorders, parents and teachers rated these preschool children as having greater executive difficulties in most domains than matched controls. Such rating-scale methodology may be a useful complementary tool by which to reliably assess executive functions in preschool children via everyday behaviors in the natural environment. PMID- 15276903 TI - Executive functioning in preschoolers: reducing the inhibitory demands of the dimensional change card sort task. AB - To investigate the role of inhibitory control in preschoolers' ability to switch sets, 3 conditions of the Dimensional Change Card Sort task ( Zelazo, Reznick, & Pinon, 1995) were tested. In Condition B (novel response options, standard stimuli) action inhibition was reduced, but the need for attentional inhibition was maintained. In Condition C (novel stimuli, standard response options) demands on both action and attentional inhibition were reduced. Performance in these was compared to that in the standard condition (A). Rule complexity was comparable across conditions. All 21 children who passed preswitch (average age 37 months) were tested on all postswitch conditions, order counterbalanced. Although reducing demands on action inhibition (Condition B) did not significantly improve performance, when demands on both action and attentional inhibition were reduced (Condition C) almost all children (95%) successfully switched sets (even children only 2 1/2 years old). Inadequate inhibition (of attention alone or both attention and action) appears sufficient to account for virtually all errors by preschoolers on this card sorting task. PMID- 15276904 TI - Using path analysis to understand executive function organization in preschool children. AB - There continues to be no consensus definition of executive functions. One way to understand different executive function components is to study abilities at their emergence, that is, early in development, and use advanced statistical methods to understand the interrelations among executive processes. However, to fully determine the constructs of interest, these methods often require complete data on a large battery of tasks, which are difficult to obtain with young children. Path analysis is an alternative statistical technique that requires only a single measure of each construct, yet still allows researchers to investigate complex relations among measures, to compare nested models, and to compare model fit across groups. Therefore, 117 preschool children (ages 2 years 8 months to 6 years 0 months) completed several executive function tasks. Path analysis was used to determine the relations between complex problem solving and working memory, inhibition, and set shifting processes. The best-fitting model included paths from working memory and inhibition to problem solving, and a correlation between working memory and inhibition. Interestingly, in younger children, inhibition was the strongest predictor of problem solving, whereas working memory contributed more strongly in older children. Suggestions for useful statistical methods to investigate the relations among executive functions in children are discussed. PMID- 15276905 TI - The contribution of executive functions to emergent mathematic skills in preschool children. AB - Mathematical ability is related to both activation of the prefrontal cortex in neuroimaging studies of adults and to executive functions in school-age children. The purpose of this study was to determine whether executive functions were related to emergent mathematical proficiency in preschool children. Preschool children (N = 96) were administered an executive function battery that was reduced empirically to working memory (WM), inhibitory control (IC), and shifting abilities by calculating composite scores derived from principal component analysis. Both WM and IC predicted early arithmetic competency, with the observed relations robust after controlling statistically for child age, maternal education, and child vocabulary. Only IC accounted for unique variance in mathematical skills, after the contribution of other executive functions were controlled statistically as well. Specific executive functions are related to emergent mathematical proficiency in this age range. Longitudinal studies using structural equation modeling are necessary to better characterize these ontogenetic relations. PMID- 15276906 TI - Executive functions following traumatic brain injury in young children: a preliminary analysis. AB - To examine executive processes in young children with traumatic brain injury (TBI), we evaluated performance of 44 children who sustained moderate-to-severe TBI prior to age 6 and to 39 comparison children on delayed response (DR), stationary boxes, and spatial reversal (SR) tasks. The tasks have different requirements for holding mental representations in working memory (WM) over a delay, inhibiting prepotent responses, and shifting response set. Age at the time of testing was divided into 10- to 35- and 36- to 85-month ranges. In relation to the community comparison group, children with moderate-to-severe TBI scored significantly lower on indexes of WM/inhibitory control (IC) on DR and stationary boxes tasks. On the latter task, the Age x Group interaction indicated that performance efficiency was significantly reduced in the older children with TBI relative to the older comparison group; performance was similar in younger children irrespective of injury status. The TBI and comparison groups did not differ on the SR task, suggesting that shifting response set was not significantly altered by TBI. In both the TBI and comparison groups, performance improved with age on the DR and stationary boxes tasks. Age at testing was not significantly related to scores on the SR task. The rate of acquisition of working memory (WM) and IC increases steeply during preschool years, but the abilities involved in shifting response set show less increase across age groups (Espy, Kaufmann, & Glisky, 2001; Luciana & Nelson, 1998). The findings of our study are consistent with the rapid development hypothesis, which predicts that skills in a rapid stage of development will be vulnerable to disruption by brain injury. PMID- 15276907 TI - Impaired neuropsychological functioning in lead-exposed children. AB - Neuropsychological functions were assessed in 174 children participating in a longitudinal study of low-level lead exposure. At age 5 1/2 years, children were administered the Working Memory and Planning Battery of the Cambridge Neuropsychological Testing Automated Battery. Measures of sociodemographic characteristics of the family, prenatal and perinatal risk, quality of caregiving and crowding in the home, and maternal and child intelligence were used as covariates to test the hypothesis that children with higher lifetime average blood lead concentrations would perform more poorly on tests of working memory, attentional flexibility, and planning and problem solving. The lifetime average blood lead level in this sample was 7.2 micrograms per deciliter (mug/dL; range: 0-20 mug/dL). Children with greater exposure performed more poorly on tests of executive processes. In both bivariate and multivariate analyses, children with higher lifetime average blood lead concentrations showed impaired performance on the tests of spatial working memory, spatial memory span, intradimensional and extradimensional shifts, and an analog of the Tower of London task. Many of the significant associations remained after controlling for children's intelligence test scores, in addition to the other covariates. These findings indicate that the effects of pediatric lead exposure are not restricted to global indexes of general intellectual functioning, and executive processes may be at particular risk of lead-induced neurotoxicity. PMID- 15276908 TI - Analytical quality goals derived from the total deviation from patients' homeostatic set points, with a margin for analytical errors. AB - The deviation of test results from patients' homeostatic set points in steady state conditions may complicate interpretation of the results and the comparison of results with clinical decision limits. In this study the total deviation from the homeostatic set point is defined as the maximum absolute deviation for 95% of measurements, and we present analytical quality requirements that prevent analytical error from increasing this deviation to more than about 12% above the value caused by biology alone. These quality requirements are: 1) The stable systematic error should be approximately 0, and 2) a systematic error that will be detected by the control program with 90% probability, should not be larger than half the value of the combined analytical and intra-individual standard deviation. As a result, when the most common control rules are used, the analytical standard deviation may be up to 0.15 times the intra-individual standard deviation. Analytical improvements beyond these requirements have little impact on the interpretability of measurement results. PMID- 15276909 TI - Biosynthesis of 32P-labelled hydroxocobalamin and a study of its behaviour in rats. AB - Using Propionibacterium freudenreichii and 32P-ATP, batches of 32P-labelled cobalamin (Cbl) were biosynthesized with a maximum specific activity of 61 microCi/mg, i.e. about 100 times higher than previously reported. Pharmacological doses mixed with 57Co-Cbl were injected subcutaneously in the form of hydroxo-Cbl into rats subsequently killed 5-20 days later. The two labelled Cbls were distributed in approximately the same way, the highest concentration being found in kidney (typical for rats) and about one-fifth of that in liver. These findings tallied with previous observations with radioactive cyano-Cbl and microbiological assay. In all injected rats, the 57Co/32P ratio was lower in liver than in kidney. Drugs eradicating the intestinal flora had no influence. In rats receiving the vitamin orally, the ratio was higher in liver than in kidney. All of our findings could be due to formation of a cobinamide-like compound lacking phosphorus. It is concluded that we have produced radiophosphorus-labelled Cbl that enables studies in vivo. PMID- 15276910 TI - C18 hydroxy fatty acids as markers of lipid peroxidation ex vivo and in vivo. AB - Different C18 monohydroxy fatty acids (OHFAs) were evaluated for their usefulness as markers of plasma lipid peroxidation (unsaturated fatty acid oxidation) ex vivo and in vivo. First, plasma samples (n = 5) were exposed for 3 h to different radical fluxes ex vivo. The formation of OHFAs was assessed by using varying concentrations of Cu2+ ions and AAPH (2,2'-azobis(2-amidinopropane) hydrochloride) as radical flux initiators. Secondly, a cross-sectional study was carried out in 47 middle-aged men. In this study, plasma concentrations of different in vivo OHFAs were compared with other indices of lipid peroxidation. Under mild oxidation conditions (heparin plasma containing 4.2 or 8.3 mM AAPH), concentrations of all the measured OHFAs (8, 9, 10, 11, 12, 13, 15 and 16-OH acids) increased in an identical manner, but under highly oxidative conditions (heparin plasma containing 83 mM AAPH or 4.2 to 8.3 mM CuSO4) mainly 9 and 13 OHFAs were formed. In the cross-sectional study, plasma 11 and 13-OHFA levels were associated statistically significantly with plasma free F2alpha isoprostanes, recognized index of in vivo lipid peroxidation (r = 0.305, p = 0.037 and r = 0.308, p = 0.035, respectively). In addition, 16-OHFA levels correlated with the ratio of electronegatively charged LDL to total LDL (r = 0.335, p = 0.021). With respect to the other OHFAs, 15-OHFA had no correlation with either other OHFAs or the reference substances used. In addition, occasionally there were contamination problems in the assessment of 12-OHFA. It is concluded that all of the measured C18 OHFAs can be used as indicators of plasma lipid peroxidation under mild oxidation conditions, though the 12 and 15 OHFAs may need to be used with some caution. Under high oxidation conditions, 9 and 13-OHFAs seem to be the most useful indices because of their high formation capacity. PMID- 15276911 TI - Clinical evaluation of the measurement of serum procalcitonin: comparative study of procalcitonin and serum amyloid A protein in patients with high and low concentrations of serum C-reactive protein. AB - Levels of C-reactive protein (CRP) and serum amyloid A protein (SAA) in blood are increased as acute phase proteins in patients with inflammatory conditions. Most of the currently used inflammatory markers, such as erythrocyte sedimentation rate and CRP or SAA levels, are non-specific parameters. By contrast, procalcitonin (PCT) has been reported to be selectively induced by severe infection in systemic inflammatory response syndrome (SIRS) and also in sepsis or multiorgan dysfunction syndrome. PCT expression is induced only slightly, if at all, by viral infections, autoimmune disorders, neoplastic disorders and trauma arising from surgical intervention. Serum PCT and SAA levels were compared in 93 patients with a CRP concentration higher than 100 mg/L and in 26 patients with a CRP concentration lower than 1.5 mg/L. In patients with high levels of CRP, all patients with sepsis and severe bacterial infection showed a significantly increased PCT concentration of more than 1.0 microg/L and it was possible to differentiate between the patients with neoplastic disorders and those with other inflammatory diseases. In patients with low levels of CRP, the PCT concentration was less than 0.3 microg/L and an increased PCT level was not seen in patients with autoimmune disorders or viral and fungal infections. These results suggest that determining the serum PCT level may be useful in the differential diagnosis of severe infection. PMID- 15276914 TI - Strategies developed by bacteria and virus for protection from the human complement system. AB - The complement system is an important part of innate immunity providing immediate protection against pathogens without a need for previous exposure. Its importance is clearly shown by the fact that patients lacking complement components suffer from fulminant and recurring infections. Complement is an explosive cascade, and in order to control it there are inhibitors present on every human cell and also circulating in blood. However, many infectious agents have developed strategies to prevent clearance and destruction by complement. Some pathogens simply hijack the host's complement inhibitors, while others are able to produce their own homologues of human inhibitors. Knowledge of these mechanisms on a molecular level may aid development of vaccines and novel therapeutic strategies that would be more specific than the use of antibiotics that, apart from causing resistance problems, also affect the normal flora, the outcome of which could be devastating. In this study the structural requirements and functional consequences of interactions between the major soluble inhibitor of complement C4b-binding protein and Neisseria gonorrhoeae, Bordetella pertussis, Streptococcus pyogenes, Escherichia coli K1, Moraxella catarrhalis and Candida albicans are described. Furthermore, a novel inhibitor produced by Kaposi's sarcoma-associated herpesvirus is identified and characterized in detail: KCP. It is shown that KCP inhibits classical C3-convertase and presents activated complement factors C4b and C3b for destruction by a serine proteinase, factor I. Using molecular modelling and site-directed mutagenesis, it was possible to localize sites on the surface of KCP required for complement inhibition and it is concluded that KCP uses molecular mechanisms identical to human inhibitors. PMID- 15276915 TI - ProBNP-derived peptides in cardiac disease. AB - The natriuretic peptides constitute a family of structurally related peptides that regulate fluid homeostasis, vascular tonus and growth. After the discovery of an endocrine component of the heart almost 25 years ago, the cardiac natriuretic peptides have now been fully accepted as useful markers in diverse aspects of cardiology including as diagnostic, therapeutic and prognostic markers of cardiac disease. In humans, atrial natriuretic peptide (ANP) and B-type natriuretic peptide (BNP) are mainly synthesized and secreted by the failing heart, whereas the related C-type natriuretic peptide (CNP) appears to be a local regulatory peptide secreted by the vascular endothelium. Accordingly, CNP is not a cardiac peptide. With the recent implementation of sensitive and specific immunoassays, increased plasma concentrations of proBNP-derived peptides have now been associated with several cardiac conditions, where the major application today seems related to ventricular dysfunction. Recently, focus has also turned to ischemic heart disease, since myocardial hypoxia increases the local BNP gene expression. This review recapitulates the established clinical applications of measuring proBNP-derived peptides in plasma. Furthermore, the evidence of increased cardiac BNP expression in ischemic heart disease will be emphasized. In turn, plasma measurement of proBNP-derived peptides may still hold new possibilities in screening for coronary artery disease. PMID- 15276916 TI - Quantitative PCR--new diagnostic tool for quantifying specific mRNA and DNA molecules: HER2/neu DNA quantification with LightCycler real-time PCR in comparison with immunohistochemistry and fluorescence in situ hybridization. AB - Previously, polymerase chain reaction (PCR) technology has been hampered by its inability to generate quantitative results, a drawback inherent to the high degree of amplification taking place in the reaction. Recently, PCR techniques have been described with the potential of quantifying the amount of mRNA or DNA in biological samples. In this study quantitative PCR was used to investigate the role of the EGF (epidermal growth factor) system in cancer both for measurements of mRNA concentrations and for measurements of the number of copies of specific genes. It is shown that the mRNA expression of a subset of ligands from the EGF system is increased in bladder cancer. Furthermore, measurement of the mRNA concentration gives important information such as the expression of these ligands correlated to the survival of the patients. In addition to the alterations at the mRNA level, changes also can occur at the DNA level in the EGF system. Thus, it has been demonstrated that the number of genes coding for the human epidermal growth factor receptor 2 (HER2) is increased in a number of breast tumors. It is now possible to treat breast cancer patients with a humanized antibody reacting with HER2, and the treatment is considered to be justified if the tumor displays an increased amount of HER2. For this reason there is a need for techniques suitable for HER2 measurements. A LightCycler real-time PCR method used for HER2/neu DNA quantification was evaluated and the results compared with those obtained by immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH). Tumor biopsies were collected from 112 patients diagnosed with early breast cancer from January 1990 to March 1994. The samples were analyzed for HER2 DNA amplification by real-time PCR on LightCycler and by FISH and for HER2 protein expression by IHC. Inter-assay variation for HER2 measured by LightCycler was 10% (x =3.1; n=17). Amplification > or = 2 was observed in 19% of the patients. Concordance rates between real-time PCR and the other methods were 91% (IHC) and 92% (FISH). The correlation between real-time PCR and FISH was highly significant (p < 0.001). The "LightCycler-HER2/neu DNA quantification kit" produces results with a high level of reproducibility and its ease of use allows rapid screening for amplification of HER2. In this paper useful information is given on how real-time PCR compares with FISH and IHC. The data show that results obtained for amplification of HER2 by real-time PCR on the LightCycler instrument are comparable to results obtained by IHC and FISH. PMID- 15276918 TI - Worldwide trends in alcohol and drug impaired driving. AB - Improved laws, enhanced enforcement, and public awareness brought about by citizens' concern, during the 1980s led to dramatic declines in drinking and driving in the industrialized world. The declines included about 50% in Great Britain, 28% in The Netherlands, 28% in Canada, 32% in Australia, 39% in France, 37% in Germany, and 26% in the United States. Some of these declines may be due in part to lifestyle changes, demographic shifts, and economic conditions. In most countries the declines reversed in the early 1990s and drinking and driving began to increase. By the middle of that decade the increases stabilized and the rates of drinking and driving again began to decline. These decreases were much less dramatic than those in the 1980s. Approaching the end of the 1990s and early in the new century, the record has been mixed. Some countries (France and Germany (until 2002)) continued to reduce drinking and driving while in other countries (Canada, the Netherlands, Great Britain, and the United States), there was stagnation and in some cases small increases or even large increase as was the case in Sweden. Complacency and attention to other issues in recent years have been difficult to overcome in some countries. Harmonization of traffic safety laws in the European Union has strengthened laws in some countries but threatens existing strong policies in others. It may be that the major gains have already been made and that additional progress will require a much greater level of scientific knowledge, use of new and emerging technologies, and political and social commitment to put in place proven countermeasures. PMID- 15276919 TI - Effects of alcohol and other drugs on driver performance. AB - In the past century we have learned that driving performance is impaired by alcohol even in low dosage, and that many other drugs are also linked to impairment. This paper is a summary of some of the more relevant studies in the past fifty years--an overview of our knowledge and unanswered questions. There is no evidence of a threshold blood alcohol (BAC) below which impairment does not occur, and there is no defined category of drivers who will not be impaired by alcohol. Alcohol increases not only the probability of collision, but also the probability of poor clinical outcome for injuries sustained when impaired by alcohol. This review samples the results of the myriad studies that have been performed during the last half century as experiments have moved from examination of simple sensory, perceptual and motor behaviours to more complex measures of cognitive functioning such as divided attention and mental workload. These more sophisticated studies show that significant impairment occurs at very low BACs (< 0.02 gm/100 ml). However, much remains to be determined regarding the more emotional aspects of behaviour, such as judgment, aggression and risk taking. Considering that the majority of alcohol related accidents occur at night, there is a need for increased examination on the role of fatigue, circadian cycles and sleep loss. The study of the effects of drugs other than alcohol is more complex because of the number of substances of potential interest, the difficulties estimating drug levels and the complexity of the drug/subject interactions. The drugs of current concern are marijuana, the benzodiazepines, other psychoactive medications, the stimulants and the narcotics. No one test or group of tests currently meets the need for detecting and documenting impairment, either in the laboratory or at the roadside. PMID- 15276920 TI - Preventing impaired driving using alcohol policy. AB - Considerable progress has been made in the reduction of impaired driving crashes during the last two decades. Much of this progress is attributable to strengthening laws against impaired driving along with vigorous enforcement efforts aimed at deterring impaired driving. In addition, many useful strategies can also be applied that focus on the control of alcohol availability, use, and promotion. Alcohol policies include controls on the price of alcohol, the location, density, and opening hours of sales outlets, controls on the social availability of alcohol, and on the promotion and advertising of alcohol. Enforcement of these policies is an important aspect of their effectiveness. These strategies have been shown to be effective or promising in reducing impaired driving as well as other consequences related to alcohol use and misuse. PMID- 15276921 TI - Preventing alcohol-related traffic injury: a health promotion approach. AB - The conditions that give rise to drinking and driving are complex, with multiple and interrelated causes. Prevention efforts benefit from an approach that relies on the combination of multiple interventions. Health promotion provides a useful framework for conceptualizing and implementing actions to reduce drinking and driving since it involves a combination of educational, behavioral, environmental, and policy approaches. This review draws on data from a range of settings to characterize the effectiveness of various interventions embedded within the health promotion approach. Interventions considered part of the health promotion approach include: (1) economic interventions (2) organizational interventions, (3) policy interventions, and (4) health education interventions, including the use of media, school and community education, and public awareness programs. Effective health promotion strengthens the skills and capabilities of individuals to take action and the capacity of groups or communities to act collectively to exert control over the determinants of alcohol-impaired driving. There is strong evidence for the effectiveness of some components of health promotion, including economic and retailer interventions, alcohol taxation, reducing alcohol availability, legal and legislative strategies, and strategies addressing the servers of alcohol. There is also evidence for the effectiveness of sobriety checkpoints, lower BAC laws, minimum legal drinking age laws, and supportive media promotion programs. Other interventions with moderate evidence of effectiveness include restricting alcohol advertising and promotion, and actions involving counter advertising. Health education interventions alone that have insufficient evidence for effectiveness include passive server training programs, school drug and alcohol education programs, community mobilization efforts, and health warnings. Because each intervention builds on the strengths of every other one, ecological approaches to reducing alcohol-impaired driving using all four components of the health promotion model are likely to be the most effective. Settings such as schools, workplaces, cities, and communities offer practical opportunities to implement alcohol-impaired driving prevention programs within this framework. PMID- 15276922 TI - Sobriety checkpoints: evidence of effectiveness is strong, but use is limited. AB - There is substantial and consistent evidence from research that highly publicized, highly visible, and frequent sobriety checkpoints in the United States reduce impaired driving fatal crashes by 18% to 24%. Although checkpoints are not conducted in 13 states for legal or policy reasons, there is strong evidence that if conducted appropriately, checkpoints would save lives in the other states. However, a recent survey of checkpoint use has demonstrated that despite the efforts of the U.S. Department of Transportation to encourage checkpoint use through publications, providing funds for equipment, and for officer overtime expenses, only about a dozen of the 37 states that conduct checkpoints do so on a weekly basis. The survey found that lack of local police resources and funding, lack of support by task forces and citizen activists, and the perception that checkpoints are not productive or cost effective are the main reasons for their infrequent use. This article discusses each of these problems and suggests a method for local communities to implement checkpoints without depending on state or federal funds. Low-staffing sobriety checkpoints conducted by as few as three to five officers have been shown to be just as effective as checkpoints conducted by 15 or more officers. A modified sobriety checkpoint program using passive alcohol sensors ("PASpoints") can be implemented by small- to moderate-sized communities in the United States to deter impaired driving. If implemented in a majority of communities, this strategy has a potential level of effectiveness similar to the high level achieved by several Australian states in their random breath-test (RBT) programs. The PASpoint system calls for a small group of three to five officers on traffic patrol duty to converge on a preset site and conduct a mini-checkpoint, returning to their standard patrol duties within two hours. Within this framework, the PASpoint operation would become a standard driving under the influence (DUI) enforcement technique regularly used within the community's jurisdiction. As a standard traffic enforcement activity, the cost would be covered by the normal enforcement budget. PMID- 15276923 TI - Underage drinking: frequency, consequences, and interventions. AB - OBJECTIVES: To examine the frequency of underage drinking, driving after drinking and alcohol-related crashes, trends in these behaviors, and promising interventions. METHODS: We examined drinking and drinking- and-driving behaviors reported in the United States in the 2001 U.S. National Household Survey of Drug Abuse, the Centers for Disease Control and Prevention 2001 Youth Risk Behavior Survey, the 1992 National Longitudinal Alcohol Epidemiologic Study, and the 1999 National Survey of Drinking and Driving conducted for the National Highway Traffic Administration. We also examined the 1999 European School Survey Project on Alcohol and Other Drugs. Alcohol-related fatal crashes were examined from the U.S. Fatality Analysis Reporting System. Evaluation of interventions to reduce teenage drinking and driving after drinking were reviewed. RESULTS: In the United States, 19% of youth ages 12-20 consumed five or more drinks on an occasion in the past 30 days. Although European nations have lower legal drinking ages (16 18) than in the United States (21), similar proportions engage in underage drinking. In two-thirds of European countries, a greater percentage of 15-16 year olds drank five or more drinks on an occasion in the past month than in the United States. In both the United States and Europe, the earlier people begin to drink, the greater the likelihood of developing alcohol dependence and other alcohol-related problems, including alcohol-related crash involvement, during adolescence and adult years. During the past 20 years alcohol-related traffic deaths among people younger than 21 have been cut in half in the United States, but progress has halted since 1995 and the problem is still large. Interventions shown by research to reduce alcohol-related crashes among youth include raising the legal drinking age to 21, zero tolerance laws, and some interventions that are family, school, or community based. CONCLUSIONS: Despite research showing that a variety of interventions can reduce underage drinking and alcohol-related crash fatalities, the frequency of these behaviors remains high and the average age of drinking initiation is declining in the United States. Efforts are needed to enhance publicized enforcement of underage drinking laws. Comprehensive community interventions that include enforcement of these laws also are needed. PMID- 15276924 TI - The prevention of young driver's DWI (driving while intoxicated) and RWDI (riding with a driver under influence) in Europe: a social-sequential model. AB - Inspired by the "critical incident technique" of McKnight et al. (1995) who analyzed 12,000 drivers' decisions leading to (or not to) DWI, and identified the influence of social, motivational, and economic factors, we have developed a social-sequential model of young drivers' DWI (driving while intoxicated) and RWDI (riding with a driver under influence) prevention. DWI or RWDI may be analyzed as a four-stage process: (1) the decision to drink and to associate leisure activities with drinking; (2) the management of alcohol consumption during the evening; (3) the decision to drive after drinking; and (4) the behavioral adaptation, once the decision to drive is taken (disinhibition of risk taking or risk compensation). At each of these four stages, preventive action can reflect the intervention of two types of actors: (1) formal social control of behavior is influenced by professionals involved in accident prevention, and (2) informal social control of behavior is influenced by the proximal environment of the subject. PMID- 15276925 TI - Drugs and driving. AB - The authors present a global overview on the issue of drugs and driving covering four major areas: (1) Epidemiology and Prevalence--which reviews epidemiological research, summarizes available information, discusses the methodological shortcomings of extant studies, and makes recommendations for future research to better define prevalence and epidemiology; (2) Effects of Medicinal and Illegal Drugs on Driving Performance--focuses on the six classes of drugs most often found in impaired and injured drivers, draws conclusions regarding the risk of these drugs to traffic safety and discusses the need for additional research; (3) Toxicological Issues--discusses ways to identify drug users via behavioral testing and analytical techniques, reviews the approaches used by different countries, screening and confirmation techniques, alternative specimens (e.g., urine, oral fluid, sweat), and how rapid roadside testing could be coupled with behavioral and laboratory testing in an effective approach to identifying and prosecuting drugged drivers; (4) Driving Under the Influence of Drugs [DUID] Laws -provides an overview of DUID laws in the United States and Europe, discusses the basic tenets of these laws, the various types of DUID statutes, the reasons why many existing laws hinder the prosecution of drugged drivers and the rationale for developing per se legislation as a strategy to more effectively manage the drugged driver problem. PMID- 15276926 TI - Epidemiology of alcohol and other drug use among motor vehicle crash victims admitted to a trauma center. AB - The objectives of this research were to (1) determine the incidence and prevalence of alcohol and other drug use among motor vehicle crash (MVC) victims admitted to a regional Level-I trauma center, and (2) to examine the utility of using a rapid point-of-collection (POC) drug-testing device to identify MVC patients with drug involvement. Blood and urine specimens were routinely collected per clinical protocol for each MVC victim at the time of admission. Blood alcohol concentration (BAC) levels were determined per standard clinical protocol. Clinical urine specimens were routinely split so that a POC drug testing device for the detection of commonly abused drugs (Marijuana, Cocaine, Amphetamines, Methamphetamines, and Opiates) could be compared to that of the standard hospital laboratory analysis of each urine specimen (which also included Barbiturates and Benzodiazepines). In the six-month period of this study, nearly two-thirds of trauma center admissions were victims of motor vehicle crashes. During this time, blood and urine was collected from 322 MVC victims. Toxicology results indicated that 59.3% of MVC victims tested positive for either commonly abused drugs or alcohol. More patients tested positive for drug use than tested positive for alcohol, with 33.5% testing positive for drug use only, 15.8% testing positive for alcohol use only, and 9.9% testing positive for both drugs and alcohol. Less than half (45.2%) of the substance-abusing patients in this study would have been identified by an alcohol test alone. After alcohol, marijuana and benzodiazepines were the most frequently detected drugs. Point of collection (POC) test results correlated well with laboratory results and provide important information to initiate rapid intervention/treatment for substance use problems among injured patients. PMID- 15276927 TI - Hard core drinking drivers. AB - The term "hard core" has been used extensively over the past 15 years to identify persons who drink and drive regularly, typically at high blood alcohol levels. This article discusses how the term arose and clarifies what it means, both as a concept and in practice. It describes the characteristics of hard core drinking drivers and estimates their contribution to drinking driver trips, arrests, and crashes. It summarizes current knowledge and recommendations on the most effective means to affect their behavior and reduce their drinking and driving. PMID- 15276928 TI - Effects of enhanced sanctions for high-BAC DWI offenders on case dispositions and rates of recidivism. AB - Implemented January 1, 1998, Minnesota's high-BAC law mandates more severe administrative pre-conviction penalties and more severe post-conviction penalties for offenses with BACs > or = 0.20%. Most notably, the law provides for the administrative impoundment of the license plate of first-time DWI offenders with BACs > or = 0.20. During the three years after the law took effect, a large majority of first-time and repeat offenders with BACs > or = 0.20% did, in fact, receive high-BAC administrative dispositions and/or high-BAC court convictions, which carried more severe penalties. For example, in 1998 85.6% of first offenders with BACs > or = 0.20% received a high-BAC administrative disposition and/or a high-BAC court conviction; 65.0% received both high-BAC administrative and high-BAC court dispositions. The proportion of high-BAC first-time offenders who received the statutory high-BAC dispositions declined from 1998 to 1999 and 2000. Based on survival analysis, the one-year recidivism rate among first offenders arrested in 1998 with BACs > or = 0.20% was significantly lower than for offenders with BACs 0.17-0.19% (who also had relatively high BACs but were not subject to enhanced sanctions), after controlling for age and gender. There were similar, but not significant, results for first offenders arrested in 1999. PMID- 15276929 TI - The emergency care setting for screening and intervention for alcohol use problems among injured and high-risk drivers: a review. AB - Each year thousands of people are treated in emergency departments and trauma centers for alcohol-related injuries, including those sustained in drinking driving crashes. Emergency departments and trauma centers provide an opportunity to screen for alcohol use problems and intervene with injured or high-risk drivers to reduce future alcohol-related traffic and injury risk. Recently physicians have expressed interest in exploring screening and intervention for alcohol use problems in these venues as a means of improving clinical care. This article reviews the literature that has examined screening and brief interventions in acute care settings to reduce future alcohol consumption and alcohol-related injury. The methodological and practical issues inherent in conducting these studies as well as in actual practice are discussed. The chaotic environment of acute care, the large numbers of patients required to be screened to obtain an adequate study sample, and high attrition rates make study in these settings difficult at best and are methodological problems that should be addressed in future research. A basic question that has not been adequately answered by research to date is whether reduction in alcohol consumption will translate to reduced alcohol-related harm, such as driving while impaired, or injurious or fatal crashes. Long-term studies that assess records-based outcomes in addition to alcohol-consumption levels are needed. PMID- 15276930 TI - Controlling impaired driving through vehicle programs: an overview. AB - The growing recognition of the problem presented by illicit vehicle operation by those whose license has been suspended for driving while intoxicated (DWI) has led to the increasing use of vehicle sanctions. These sanctions include vehicle impoundment and forfeiture, vehicle registration cancellation, and vehicle interlocks as penalties for DWI and driving while suspended (DWS). This article reviews the current information available on the use and effectiveness of vehicle sanctions for reducing offender recidivism. In the United States, 14 states have impoundment laws that are widely used as sanctions for both DWI and DWS, with the length of the impoundment increasing with the number of previous offenses. These laws have been shown to reduce recidivism while the vehicle is in custody and, to a lesser extent, even after the vehicle has been released. Vehicle impoundment is also widely used in Canada and New Zealand. Although a larger number of U.S. states have laws providing for vehicle forfeiture for DWI or DWS, this sanction tends to be limited to multiple offenders and therefore impacts fewer drivers. Cancellation of the vehicle registration and the confiscation of the vehicle plates are increasing in popularity because the vehicle tags are the property of the state, rather than the vehicle owner. Vehicle alcohol interlocks have proven to be an effective method for reducing DWI offender recidivism while they are on the car, but appear to produce only limited post-treatment behavior change. Interlocks are widely used in the United States and Canada and are beginning to be implemented in Europe and Australia. The issues that arise in implementing vehicle sanction programs are discussed and the actions taken by states to deal with them are described. PMID- 15276931 TI - Alcohol ignition interlock programs. AB - The alcohol ignition interlock is an in-vehicle DWI control device that prevents a car from starting until the operator provides a breath alcohol concentration (BAC) test below a set level, usually .02% (20 mg/dl) to .04% (40 mg/dl). The first interlock program was begun as a pilot test in California 18 years ago; today all but a few US states, and Canadian provinces have interlock enabling legislation. Sweden has recently implemented a nationwide interlock program. Other nations of the European Union and as well as several Australian states are testing it on a small scale or through pilot research. This article describes the interlock device and reviews the development and current status of interlock programs including their public safety benefit and the public practice impediments to more widespread adoption of these DWI control devices. Included in this review are (1) a discussion of the technological breakthroughs and certification standards that gave rise to the design features of equipment that is in widespread use today; (2) a commentary on the growing level of adoption of interlocks by governments despite the judicial and legislative practices that prevent more widespread use of them; (3) a brief overview of the extant literature documenting a high degree of interlock efficacy while installed, and the rapid loss of their preventative effect on repeat DWI once they are removed from the vehicles; (4) a discussion of the representativeness of subjects in the current research studies; (5) a discussion of research innovations, including motivational intervention efforts that may extend the controlling effect of the interlock, and data mining research that has uncovered ways to use the stored interlock data record of BAC tests in order to predict high risk drivers; and (6) a discussion of communication barriers and conceptual rigidities that may be preventing the alcohol ignition interlock from taking a more prominent role in the arsenal of tools used to control DWI. Whether interlock programs can help public policymakers achieve their expressed goals of substantially reducing the level of impaired driving will remain uncertain until procedural barriers and intransigent judiciary practices can be overcome that provide for more systematic routine use of interlock programs. Despite strong effectiveness evidence in all studies to date, the real potential of this technology to reduce the road toll cannot be estimated until they are more widely adopted. PMID- 15276932 TI - Emerging technological approaches for controlling the hard core DUI offender in the U.S. AB - Drivers convicted of impaired driving are substantially overrepresented in alcohol-related fatal crashes. Because many such offenders continue to drive with suspended operators' licenses, monitoring their postconviction driving is a significant problem for the criminal justice system. Technology for tracking the location and drinking of such offenders is a rapidly developing field, which promises to provide methods for monitoring offenders on a 24/7 basis. The status of traditional monitoring methods is reviewed and contrasted with the new technologies that are being implemented. Although those technologies offer considerable promise, they have not yet been evaluated in programs for impaired driving offenders. Eight issues related to the probability of rapid implementation of the new technologies are discussed. PMID- 15276933 TI - The International Consortium on Mental Health Policy and Services: objectives, design and project implementation. AB - The concept of the burden of disease, introduced and estimated for a broad range of diseases in the World Bank report of 1993 illustrated that mental and neurological disorders not only entail a higher burden than cancer, but are responsible, in developed and developing countries, for more than 15% of the total burden of all diseases. As a consequence, over the past decade, mental disorders have ranked increasingly highly on the international agenda for health. However, the fact that mental health and nervous system disorders are now high on the international health agenda is by no means a guarantee that the fate of patients suffering from these disorders in developing countries will improve. In most developing countries the treatment gap for mental and neurological disorders is still unacceptably high. To address this problem, an international network of collaborating institutions in low-income countries has been set up. The establishment and the achievements of this network--the International Consortium on Mental Health Policy and Services--are reported. Sixteen institutions in developing countries collaborate (supported by a small number of scientific resource centres in industrialized nations) in projects on applied mental health systems research. Over a two-year period, the network produced the key elements of a national mental health policy; provided tools and methods for assessing a country's current mental health status (context, needs and demands, programmes, services and care and outcomes); established a global network of expertise, i.e., institutions and experts, for use by countries wishing to reform their mental health policy, services and care; and generated guidelines and examples for upgrading mental health policy with due regard to the existing mental health delivery system and demographic, cultural and economic factors. PMID- 15276934 TI - The mental health policy template: domains and elements for mental health policy formulation. AB - Mental disorders are a major and rising cause of disease burden in all countries. Even when resources are available, many countries do not have the policy and planning frameworks in place to identify and deliver effective interventions. The World Health Organization (WHO) and the World Bank have emphasized the need for ready access to the basic tools for mental health policy formulation, implementation and sustained development. The Analytical Studies on Mental Health Policy and Service Project, undertaken in 1999-2001 by the International Consortium for Mental Health Services and funded by the Global Forum for Health Research aims to address this need through the development of a template for mental health policy formulation. A mental health policy template has been developed based on an inventory of the key elements of a successful mental health policy. These elements have been validated against a review of international literature, a study of existing mental health policies and the results of extensive consultations with experts in the six WHO regions of the world. The Mental Health Policy Template has been revised and its applicability will be tested in a number of developing countries during 2001-2002. The Mental Health Policy Template and the work of the Consortium for Mental Health Services will be presented and the future role of the template in mental health policy development and reform in developing countries will be discussed. PMID- 15276935 TI - The appropriateness and use of focus group methodology across international mental health communities. AB - The ability to interpret collected data across international mental health communities often proves to be difficult. The following paper reports on the use and appropriateness of focus group methodology in helping to clarify issues that could help substantiate data collection and comparison across different cultures and regions. Field tests of the focus group methodology were undertaken in different regions and this paper describes an overview of the final field test in Sofia, Bulgaria. The findings and experiences with utilizing this methodology were incorporated in subsequent data collections. PMID- 15276936 TI - The mental health country profile: background, design and use of a systematic method of appraisal. AB - This article describes the construction and use of a systematic structured method of mental health country situation appraisal, in order to help meet the need for conceptual tools to assist planners and policy makers develop and audit policy and implementation strategies. The tool encompasses the key domains of context, needs, resources, provisions and outcomes, and provides a framework for synthesizing key qualitative and quantitative information, flagging up gaps in knowledge, and for reviewing existing policies. It serves as an enabling tool to alert and inform policy makers, professionals and other key stakeholders about important issues which need to be considered in mental health policy development. It provides detailed country specific information in a systematic format, to facilitate global sharing of experiences of mental health reform and strategies between policy makers and other stakeholders. Lastly, it is designed to be a capacity building tool for local stakeholders to enhance situation appraisal, and multisectorial policy development and implementation. PMID- 15276937 TI - Kenya mental health country profile. AB - The Kenya country profile is a description of Kenya covering the demographic, economic, cultural, religious, and health aspects including mental health in the country today. Like any other developing countries, Kenya is faced today with major challenges in terms of poverty, economic decline, and lack of adequate resources to meet the health needs and demands, including the mental health of the population. The situational analysis is described in the country profile with a snapshot of the approach in terms of objectives to address the way forward for Kenya. PMID- 15276938 TI - Uganda mental health country profile. AB - With the help of the International Consortium for Mental Health Policy and Services, data on country mental health services was gathered through a descriptive, cross sectional study. The study population included policymakers, health providers and consumers of health services. Data was collected at national level from relevant sectors including the Ministry of Health, Butabika National Referral Mental Hospital and four rural districts. The districts were purposively selected because of existing consumer groups. Quantitative data was collected by interviewer-administered questionnaire and record reviews at hospitals and district headquarters. It was observed that the country has inadequate numbers of mental health professionals with poor mental health funding. Such factors, compounded with inappropriate cultural beliefs, are major obstacles to the delivery of mental health services. There is however an attempt by the Government to improve mental health services. The current health policy is an opportunity to improve access to mental health care. Currently there is improved pre-service and in-service training for mental health workers with ongoing rehabilitation and remodelling of the mental health infrastructure in the country. The burden of mental disorders in Uganda is high in a country that is poorly resourced. The majority of the population is rural and still harbours negative cultural beliefs. There is a need to increase advocacy for mental health and develop capacity for professional mental and general health workers to be supported by appropriate policies, facilities and finances. PMID- 15276939 TI - Zambia mental health country profile. AB - This country profile for Zambia was compiled between 1998 and 2002. The objectives of the exercise were to first of all avail policymakers, other key decision makers and leaders in Zambia, information about mental health in Zambia in order to assist policy and services development. Secondly, to facilitate comparative analyses of mental health services between countries. The work involved formation of a core group of experts who coordinated the collection of information from the various organizations in Zambia. The information was later shared to a broad spectrum of stakeholders for consensus. A series of focus group discussions (FGDs) supplemented the information collected. There are various factors that contribute to mental health in Zambia. It is clear from the Zambian perspective that social, demographic, economic, political, environmental, cultural and religious influences affect the mental health of the people. With a population of 10.3 million and annual growth rate of 2.9%, Zambia is one of the most urbanized countries in sub-Saharan Africa. Poverty levels stood at 72.9% in 1998. In terms of unemployment, the most urbanized provinces, Lusaka (the capital city), and the copper-belt are the most affected. The gross domestic product (GDP) is US$3.09 billion dollars while per capita income is US$300. The total budget allocation for health in the year 2002 was 15% while the proportion of the GDP per capita expenditure for health was 5.6%. The HIV/AIDS prevalence rates stand at 20% among the reproductive age group 15-49 years. Political instability and wars in neighbouring states has resulted in an influx of refugees. Environmental factors affecting the country include natural and man-made disasters such as floods and drought, mine accidents, and deforestation. To a large extent in Zambia, people who are mentally ill are stigmatized, feared, scorned at, humiliated and condemned. However, caring for mental ill health in old age is positively perceived. It is traditionally the duty and responsibility of the extended family to look after the aged. Gender based violence (GBV) is another issue. Women, who are totally dependent on their spouses economically, are forced by circumstances to continue living in abusive relationships to the detriment of their mental well-being. In Zambia, the family is considered sacrosanct and the affairs of the family members, private. It is within this context that GBV is regarded as a family affair and therefore a private affair, yet spouse beating has led to depression and in some cases death. In terms of psychiatric services, there are close to 560 beds for psychiatric patients across the country. Common mental disorders found in Zambia are acute psychotic episodes, schizophrenia, affective disorders, alcohol related problems and organic brain syndromes. About 70-80% of people with mental health problems consult traditional health practitioners before they seek help from conventional health practitioners. Over time the number of frontline mental health workers and professional staff has been declining. This is due to the 'brain drain', retirement, death and low output from training institutions. For practicing psychiatrists, only one is available for the whole country. Other key mental health workers such as psychologists, social workers and occupational therapists are also in short supply. All in all, the mental health services situation in Zambia could be described as critical, requiring urgent attention. PMID- 15276940 TI - Chile mental health country profile. AB - This paper describes main facts about Chile starting with key socio-demographic, socio-economic, political, environmental, epidemiological, social support and social pathology aspects that characterize the context in which current mental and neurological policy and programmes have been put in place since 2000, as part of the National Health Plan and Health Sector Strategy Plan. The 'National Plan for Mental Health and Psychiatry', using a community psychiatry approach, has been partially implemented for people covered by the Public Health Insurance, which comprises 62% of the Chilean population (people with lower income). This paper also describes: the management, population needs and demands, financial resources, human resources in primary care, mental health specialist care and community-based care, physical capital, social capital, provision and processes, and outcomes of the plan. Strengths are analyzed, like the health reform, including its values and principles, the active participation of consumer and family groups as well as mental health NGOs, access to mental health services through primary care, quality assurance of the mental health services delivered to the population and progressive development of a culture of respect for human rights, including those of people with mental illnesses. Finally, difficulties for the advance of mental health care are also enumerated: the low priority still given to mental health compared with physical health by the country's leaders, the insufficient emphasis on mental health in both undergraduate and postgraduate professional training, the strong stigma and discrimination associated with mental illness in the general population and the advocacy by some mental health professionals of the traditional model of care (role of the psychiatric hospital). PMID- 15276942 TI - Bulgaria mental health country profile. AB - The mental health profile of Bulgaria has been compiled and following analysis of both the factual findings and the process of data collection a report has been prepared. The subject of discussion in the paper concerns several major findings: the discrepancy between what the policy documents state and the actual situation in mental health; the organizational culture, which alienates; and the peculiarities of the process of change and how it is driven under political pressure from outside the country. Analysis extends to encompass the influence of the general health reform on the mental health sector, the deficits of the leadership and how they impact on the effectiveness of the system, and the interdependence between the country's economy and the health sector. A conclusion is made about the need to consolidate the public health approach using the lever of international collaboration in the field of mental health. PMID- 15276941 TI - Pakistan mental health country profile. AB - The Republic of Pakistan is a South East Asian country with a population of over 140.7 million. Its population is fast growing and the majority (70%) live in rural areas with a feudal or tribal value system. The economy is dependent on agriculture and 35% of the population live below the poverty line. Islam is the main religion and 'mental illnesses' are stigmatized and widely perceived to have supernatural causes. The traditional healers along with psychiatric services are the main mental health service providers. The number of trained mental health professionals is small as compared to the population demands and specialist services are virtually non-existent. Lack of data on prevalence of various mental illnesses and monitory constraints are the major hurdles in the development of mental health services. A number of innovative programmes to develop indigenous models of care like the 'Community Mental Health Programme' and 'Schools Mental Health Programme' have been developed. These programmes have been found effective in reducing stigma and increase awareness of mental illness amongst the adults and children living in rural areas. Efforts by the government and mental health professionals have led to the implementation of a 'National Mental Health Policy' and 'Mental Health Act' in 2001. These aim at integrating mental health services with the existing health services, improving mental health care delivery and safeguarding the rights of mentally ill people. A favourable political will and the help of international institutions like the World Health Organization are required to achieve these aims. PMID- 15276943 TI - Georgia mental health country profile. AB - This article is on the mental health of the population of Georgia describing context and mental health needs, extrinsic and intrinsic influences on mental health in the country, and health and social services for people with mental health problems and mental illnesses. The mental health profile of Georgia has been compiled with the help of an instrument developed by the International Consortium on Mental Health. Georgia is one of the former republics of the USSR with a population of about 4.5 million people. Political, social and economic changes have led to wide scale social stress in Georgia in recent years and to related psychosocial and behavioural problems. Difficult economic conditions and internal displacement are thought to be key contributory factors to the three fold increase in suicide experienced during the decade following independence. Georgia has produced a written mental health policy. However, its implementation is hampered by the lack of resources, training and community awareness. The Government has passed a Mental Health Act, but this is also ineffective at present due to a lack of staff, financing and appropriate facilities. PMID- 15276944 TI - Lithuania mental health country profile. AB - As a part of international mental health policy, programmes and services project, the 'country profile' instrument was used for assessment of mental health policy and services in the Republic of Lithuania. Analysis of contextual factors revealed high levels of social pathology (including violence, suicide and other self-destructive behaviour) with stigmatizing approaches by the general population to mentally disturbed persons and other vulnerable groups. Analysis of existing data about resources invested in the mental health care system raises questions for policymakers about the effectiveness of this traditional way of investment. The largest proportion of physical and human capital is concentrated in psychiatric institutions, with large numbers of beds, psychiatrists and increasing funding for medications, while other components of care--such as housing, psychosocial and vocational rehabilitation, community-based child mental health services--are not being developed. Statistical accounts keep the tradition of presenting processes as outcomes, while modern assessment of outcomes of services, programmes and policies are lacking. The findings from this country profile may be very useful in the development of modern mental health policies in the countries of Eastern and Central Europe, which have been deprived for decades from the opportunity to introduce evidence-based mental health policies and services. PMID- 15276945 TI - India mental health country profile. AB - India, the second most populated country of the world with a population of 1.027 billion, is a country of contrasts. It is characterized as one of the world's largest industrial nations, yet most of the negative characteristics of poor and developing countries define India too. The population is predominantly rural, and 36% of people still live below poverty line. There is a continuous migration of rural people into urban slums creating major health and economic problems. India is one of the pioneer countries in health services planning with a focus on primary health care. Improvement in the health status of the population has been one of the major thrust areas for social development programmes in the country. However, only a small percentage of the total annual budget is spent on health. Mental health is part of the general health services, and carries no separate budget. The National Mental Health Programme serves practically as the mental health policy. Recently, there was an eight-fold increase in budget allocation for the National Mental Health Programme for the Tenth Five-Year Plan (2002 2007). India is a multicultural traditional society where people visit religious and traditional healers for general and mental health related problems. However, wherever modern health services are available, people do come forward. India has a number of public policy and judicial enactments, which may impact on mental health. These have tried to address the issues of stigma attached to the mental illnesses and the rights of mentally ill people in society. A large number of epidemiological surveys done in India on mental disorders have demonstrated the prevalence of mental morbidity in rural and urban areas of the country; these rates are comparable to global rates. Although India is well placed as far as trained manpower in general health services is concerned, the mental health trained personnel are quite limited, and these are mostly based in urban areas. Considering this, development of mental health services has been linked with general health services and primary health care. Training opportunities for various kinds of mental health personnel are gradually increasing in various academic institutions in the country and recently, there has been a major initiative in the growth of private psychiatric services to fill a vacuum that the public mental health services have been slow to address. A number of non governmental organizations have also initiated activities related to rehabilitation programmes, human rights of mentally ill people, and school mental health programmes. Despite all these efforts and progress, a lot has still to be done towards all aspects of mental health care in India in respect of training, research, and provision of clinical services to promote mental health in all sections of society. PMID- 15276946 TI - Nepal mental health country profile. AB - The Kingdom of Nepal is situated in the heart of Asia, between its two big neighbours China and India. Nepal is home to several ethnic groups. The majority of the 23 million population reside in the countryside. Although figures on many of the health and socio-economic indicators are non-existing, some existing ones show gradual improvement over the years. However the figures for illiteracy and infant mortality are still one of the highest in the world. As per GDP, and population living below the poverty line and per capita income, Nepal still remains one of the poorest countries in the world. Despite this, it provides shelter to thousands of Bhutanese refugees in its land. Frequent natural disasters and recent violent conflicts in Nepal have further added hardship to life. Less than 3% of the national budget is allocated to the health sector. Mental health receives insignificant attention. The Government spends about 1% of the health budget on mental health. There is no mental health act and the National Mental Health Policy formulated in 1997 is yet to be fully operational. Mental ill health is not much talked about because of the stigma attached. The roles of the legal and insurance systems are almost negligible. The financial burden rests upon the family. The traditional/religious healing methods still remain actively practiced, specifically in the field of mental health. The service, comprising little more than two-dozen psychiatrists along with a few psychiatric nurses and clinical psychologists (mainly practicing in modern health care facilities) has started showing its impact--however this is limited to specific urban areas. The majority of the modern health care facilities across the country are devoid of a mental health facility. The main contextual challenges for mental health in Nepal are the provision of adequate manpower, spreading the services across the country, increasing public awareness and formulating and implementing an adequate policy. PMID- 15276947 TI - Thailand mental health country profile. AB - Thailand, a constitutional monarchy, has undergone a rapid shift in its demography and economy in last two decades. This has put a great burden on the health services, including mental health care of the country. The current emphasis of the Ministry of Public Health is to change its role from health care provider to policymaker and regulator of standards, and to provide technical support to health facilities under its jurisdiction as well as in the private sector. The Department of Mental Health, established in 1994, has laid down a mental health policy that aims to promote mental health care within the community with the help of people's participation in health programmes. Focus has been placed on developing suitable and efficient technology by seeking cooperation both within and outside the Ministry of Public Health. Consequently, the Department of Mental Health has been receiving increasing budgetary allocations. Since there is a paucity of trained manpower, the emphasis is being laid on the utilization of general health care for mental health care. Some of the specific interventions are community services, prison services, psychiatric rehabilitation, and use of media in mental health operations. There have been active efforts towards international cooperation for developing technologies for specific programmes. Private and non-governmental organizations are supported and encouraged to provide mental health care to the marginalized sections of society. Efforts have also been made by the Department of Mental Health to inspect and raise the efficiency of its operations to result in quality service. PMID- 15276948 TI - Philippines mental health country profile. AB - The Philippines is one of the world's most heavily populated countries. Even though democracy was restored in 1986 after years of occupation and dictatorship, a high level of poverty still exists and malnutrition and communicable diseases continue to be the main cause of morbidity. For almost 50 years people with mental disorders have been treated in a mental hospital setting. The National Mental Health Program aims to establish psychiatric wards in university and private hospitals and encourage community-based mental health care. PMID- 15276949 TI - Malaysia mental health country profile. AB - Malaysia is a tropical country in the heart of south east Asia with a population of 24 million people of diverse ethnic, cultural and religious backgrounds living in harmony in 330,000 km(2) of land on the Asian mainland and Borneo. Malaysia, which lies on the crossroads of trade between east and west Asia, has an ancient history as a centre of trading attracting commerce between Europe, west Asia, India and China. It has had influences from major powers that dominated the region throughout its history. Today the country, after independence in 1957, has embarked on an ambitious development project to make it a developed country by 2020. In this effort the economy has changed from one producing raw material to one manufacturing consumer goods and services and the colonial health system has been overhauled and social systems strengthened to provide better services for its people. The per capita income, which was under 1,000 US dollars at independence, has now passed 4,000 US dollars and continues to grow, with the economy largely based on strong exports that amount to over 100 billion US dollars. The mental health system that was based on institutional care in four mental hospitals at independence from British colonial rule in 1957 with no Malaysian psychiatrists is today largely based on over 30 general hospital psychiatric units spread throughout the country. With three local postgraduate training programmes in psychiatry and 12 undergraduate departments of psychiatry in the country--all started after independence--there is now a healthy development of mental health services. This is being supplemented by a newly established primary care mental health service that covers community mental health by integrating mental health into primary health care. Mental health care at the level of psychiatrists rests with about 140 psychiatrists most of whom had undertaken a four-year masters course in postgraduate psychiatry in Malaysia since 1973. However, there continues to be severe shortages of other professionals such as clinical psychologists and social workers in mental health services. There are a few specialists, and specialized services in child, adolescent, forensic, rehabilitative, liaison or research fields of mental health. In the area of services for women and children, as well as the disabled in the community, there are strong efforts to improve the care and provide services that are in keeping with a caring society. New legislation on these are being passed every year and the setting up of a Ministry for Women's Affairs is one such move in recent years. Mental health in Malaysia has been slow in developing but has in the past decade seen important strides to bring it on par with other branches of medicine. PMID- 15276951 TI - Neurologic aspects of traumatic brain injury. AB - Traumatic brain injury is a common neurologic condition that can have a significant emotional and financial burden. Neurologic injury is classified on the basis of initial clinical status by the Glasgow Coma Scale, and also by the type and location of head injury. Complications in the management of these patients are reviewed, ranging from intracranial pressure management and stroke to post-traumatic epilepsy. In addition, predictive prognostic variables that can be used to predict outcome based on a patient's presentation at the time of a head trauma are discussed. Finally, interventions such as induced hypothermia that can be undertaken to try to optimize outcome, are discussed along with current data in support of or against such techniques. PMID- 15276952 TI - Post concussion syndrome. AB - Individuals sustaining mild traumatic brain injuries often report a constellation of physical, cognitive, and emotional/behavioral symptoms referred to as post concussion symptoms (PCS). The most commonly reported post concussion symptoms are headache, dizziness, decreased concentration, memory problems, irritability, fatigue, visual disturbances, sensitivity to noise, judgment problems, depression, and anxiety. Although these PCS often resolve within one month, in some individuals PCS can persist from months to years following injury and may even be permanent and cause disability. When this cluster of PCS is persistent in nature, it is often called the post concussion syndrome or persistent PCS. Both physiological and psychological etiologies have been suggested as causes for persistent post concussion symptoms and this has led to much controversy and debate in the literature. Most investigators now believe that a variety of pre morbid, injury-related, and post-morbid neuropathological and psychological factors contribute to the development and continuation of these symptoms in those sustaining mild traumatic brain injury (MTBI). PMID- 15276953 TI - Mood disorders following traumatic brain injury. AB - Mood disorders are a frequent complication of traumatic brain injury that exerts a deleterious effect on the recovery process and psychosocial outcome of brain injured patients. Prior psychiatric history and impaired social support have been consistently reported as risk factors for developing mood disorders after traumatic brain injury (TBI). In addition, biological factors such as the involvement of the prefrontal cortex and probably other limbic and paralimbic structures may play a significant role in the complex pathophysiology of these disorders. Preliminary studies have suggested that selective serotonin reuptake inhibitors such as sertraline, mood stabilizers such as sodium valproate, as well as stimulants and ECT may be useful in treating these disorders. Mood disorders occurring after TBI are clearly an area of neuropsychiatry in which further research in etiology as well as treatment is needed. PMID- 15276954 TI - Psychosis following traumatic brain injury. AB - Psychosis is a relatively infrequent but potentially serious and debilitating consequence of traumatic brain injury (TBI), and one about which there is considerable scientific uncertainty and disagreement. There are several substantial clinical, epidemiological, and neurobiological differences between the post-traumatic psychoses and the primary psychotic disorders. The recognition of these differences may facilitate identification and treatment of patients whose psychosis is most appropriately regarded as post-traumatic. In the service of assisting psychiatrists and other mental health clinicians in the diagnosis and treatment of persons with post-traumatic psychoses, this article will review post-traumatic psychosis, including definitions relevant to describing the clinical syndrome, as well as epidemiologic, neurobiological, and neurogenetic factors attendant to it. An approach to evaluation and treatment will then be offered, emphasizing identification of the syndrome of post-traumatic psychosis, consideration of the differential diagnosis of this condition, and careful selection and administration of treatment interventions. PMID- 15276955 TI - A quantitative review of the effects of traumatic brain injury on cognitive functioning. AB - Changes in cognitive functioning often result from traumatic brain injury (TBI) and predict other important aspects of psychosocial recovery. Despite this pivotal role, no quantitative review of cognitive functioning across the spectrum of TBI severity has been reported. We therefore conducted a meta-analysis of 39 mostly cross-sectional studies of the cognitive effects of mild head injury (MHI) and moderate-severe TBI from the acute phase through long-term follow-up. The studies reported 48 comparisons of patients (n = 1716) and control subjects (n = 1164). Averaged across all follow-up periods, the effect of moderate-severe TBI (weighted mean Cohen's d = -0.74) was more than three times the effect of MHI (weighted mean d = -0.24) on overall cognitive functioning. Further, the natural logarithm of the follow-up interval correlated very strongly with estimates of d among patients with MHI, but less so among those with moderate-severe TBI. In short, findings from published research suggest that overall cognitive functioning recovers most rapidly during the first few weeks following MHI, and essentially returns to baseline within 1-3 months. Cognitive functioning also improves during the first two years after moderate-severe TBI, but remains markedly impaired even among patients tested > 2 years post-injury. PMID- 15276956 TI - Obsessive-compulsive disorder after traumatic brain injury. AB - Traumatic brain injury ( TBI) neuropsychiatric sequelae are a significant cause of morbidity in TBI victims. Among the recognized sequelae are anxiety, obsessions, compulsions and obsessive-compulsive disorder (OCD). This review addresses the emergence of OCD and OCD symptoms after TBI with an emphasis on neural circuits that underlie OCD symptom expression that may be affected by the injury. Current studies suggest that post-TBI emergent psychopathology, including OCD, is influenced by underlying sub-clinical diathesis, brain injury lesions sites, environmental stressors and the rehabilitation process. Pre-morbid status can be obtained by structured psychiatric interviews, and TBI brain lesions can be defined with advanced neuroimaging techniques. This information along with the management of family and environmental stressors and the enhanced clinical identification of symptoms of anxiety and OCD can be used in the rehabilitation process to improve prognosis after TBI. PMID- 15276957 TI - Pharmacological management of the psychiatric aspects of traumatic brain injury. AB - Traumatic brain injury (TBI) is a significant public health problem that affects millions of people annually in the USA. Neuropsychiatric symptoms such as cognitive deficits, depression, mania, anxiety, psychosis, apathy, and sleep disturbance are common after TBI. An extensive array of pharmacological options are available to treat a wide range of neuropsychiatric sequelae of TBI, yet there have been few controlled clinical trials to assess the effects of pharmacotherapy in TBI patients. Treatment of the neuropsychiatric disturbances associated with TBI should result in decreased handicap, improved quality of life, and decreased societal impact. There is a dire need for large, randomized, double blind, placebo-controlled trials that include a broad range of cognitive and behavioral outcome measures. PMID- 15276958 TI - Non-pharmacological management of psychiatric disturbances after traumatic brain injury. AB - Persons who suffer traumatic brain injury (TBI) often demonstrate a variety of psychiatric and neuropsychiatric disturbances. Some of those disturbances may be managed by non-pharmacological methods. The methods draw heavily on established principles of psychotherapy and behavioral modification. However, the unique problems imposed by neurocognitive deficits must be factored into any form of non pharmacological intervention with this patient group. A simple model consolidates information about the important ingredients in the non-pharmacological management of psychiatric disturbances in TBI patients. PMID- 15276960 TI - Eating disorders. AB - Eating disorders rank among the most debilitating psychiatric disturbances that affect young women. Knowledge has increased in recent years about the two major eating disorders, anorexia nervosa (AN) and bulimia nervosa (BN); however, much remains unknown. This review article will provide an overview of the epidemiology, proposed risk factors and clinical features of AN, and BN, as well as current recommendations for evaluation and treatment of these disorders. PMID- 15276961 TI - Psychiatric disorders during pregnancy. AB - Treating women with psychiatric disorders during pregnancy is a challenge for numerous reasons. Balancing the risks and benefits of symptoms and treatments is particularly important during pregnancy because both medication and maternal illness may have adverse effects on the fetus. Communication of options in the management of psychiatric disorders in pregnancy is vital to optimal treatment. One barrier to effective communication has been a paucity of research from which clinicians can draw information, particularly in the area of pharmacological treatment. However, emerging evidence points to the low risk of many psychotropic medications during pregnancy. Uncertainty must not prevent frank risk-benefit discussions from occurring between treating physicians and their pregnant patients. Psychiatrists can prepare themselves for management decisions by reviewing the current literature. PMID- 15276962 TI - Postpartum mood disorders. AB - Depression is a common disorder in women of childbearing age. Many women experience depressive symptoms during the postpartum period, ranging from mild postpartum blues to significant mood disorders such as postpartum depression and postpartum psychosis. The 'baby blues' are extremely common, affecting 30-75% of new mothers. This form of postpartum mood change is self-limited and requires no specific treatment other than education and support. While less common, occurring in 10-15% of births, postpartum depression has the potential for significant impact on both the health of the mother and baby. Unfortunately, affective illness in women frequently goes unrecognized and untreated. While there are effective pharmacological treatments for postpartum depression, the treatments for postpartum depression are often not utilized due to concerns about lactation. Postpartum psychosis is extremely rare, affecting one to two women per 1000 births; each case represents a true psychiatric emergency. Identifying and treating postpartum affective illness in women is critical to the health of both mother and infant. This paper reviews the literature on the diagnosis and treatment of mood disorders in the postpartum period: postpartum blues, postpartum depression and postpartum psychosis. PMID- 15276963 TI - The impact of maternal depression on familial relationships. AB - Depression is one of the most prevalent psychiatric illnesses. It is particularly common in women of childbearing age. It is recurrent and tends to have a chronic course and is often comorbid in nature. It is important to view depression within its social context, as it is a disease, which impacts not only the individual but also the wider community. Evidence abounds as to the negative impact of maternal depression on children, husbands/partners, and family. Children of depressed women show deficits in social, psychological, and cognitive domains and are at increased risk for depression themselves and other psychiatric illness such as conduct disorder. They are also at an increased risk for child abuse. The mechanisms by which maternal depression may lead to child psychopathology including genetics, poor parenting, modelling, and environment are explored. Many children with depressed mothers cope well and escape negative effects; consequently the concept of resilience is elucidated. Research shows that a significant percentage of men become depressed when their wives/partners are depressed particularly if they have postnatal depression. There is an increase in marital discord and conflict within families of depressed women, all of which can have a deleterious effect on children. Children with two depressed parents are at an elevated risk of a negative outcome as compared to those with only one depressed parent. Finally the various interventions, management, and recommendations are examined. PMID- 15276964 TI - Alcohol misuse by women. AB - Alcohol misuse among women is an important and growing problem. There is epidemiological and metabolic evidence that risk factors for and consequences of alcohol misuse are significantly different for women than for men. Understanding these differences is imperative if effective preventative and treatment interventions are to be undertaken. This article reviews the epidemiology of alcohol misuse by women, effects of alcohol misuse on women, fetuses, and relationships, and assessment and treatment strategies. We then suggest directions for future research in this field. PMID- 15276965 TI - Psychiatric disorders in older women. AB - With increased longevity and an aging 'baby boomer' population, the numbers of women in older age groups in the United States will be increasing significantly over the coming years. Older women with schizophrenia and mood disorders that began in early adult life continue to need psychiatric treatment, although treatment considerations may require modification with age. In addition, late onset depressive disorders are more common among older women than older men and may undermine physical and psychosocial well-being if inadequately treated. Late onset schizophrenia and bipolar disorder are less common, but also affect older women more often than older men. Moreover, psychiatric concerns related to Alzheimer's disease and care giving are pertinent to this older population. Clinicians need to be knowledgeable about differences in epidemiology and clinical presentation that distinguish psychiatric disorders in older women from both men and from younger cohorts. PMID- 15276966 TI - The role of estrogen in mood disorders in women. AB - Major depression is twice as common in women as men and depressive episodes appear to be more common in women with bipolar disorder. There is accumulating evidence that, in at least some women, reproductive-related hormonal changes may play a role in increasing the risk of depressive symptoms premenstrually, postpartum and in the perimenopausal period. In this review, the evidence for the role of hormonal fluctuations, specifically estrogen, in triggering depressive symptoms in a subgroup of women is summarized. In addition, the potential role of estrogen in triggering depressive symptoms via its effects on the serotonergic system, brain-derived neurotrophic factor and Protein Kinase C is reviewed. PMID- 15276976 TI - Walter T. Johnson (1892 to 1937): pioneer of coccidiosis research in the fowl. AB - Walter T. Johnson (1892 to 1937), veterinarian at Western Washington Agricultural Experiment Station (WWAES), Puyallup, Washington, USA, and subsequently poultry pathologist and professor of veterinary medicine at Oregon State University Agricultural Experiment Station (OSAES), Corvallis, Oregon, USA, made many important contributions to our understanding of the disease coccidiosis. His pioneering work included the first description of Eimeria necatrix and Eimeria praecox from the chicken and identification of four other species of Eimeria from the fowl. He demonstrated the relationship between numbers of oocysts ingested and severity of infection, and described the phenomenon of host specificity and the significance of immunity. Contrary to widespread opinion, he considered that coccidia were not involved in blackhead disease and other pathological conditions reported from the fowl. His views on control were ahead of his time and he anticipated the possibility of vaccinating birds by infecting them with live oocysts. In addition to his studies of coccidiosis, Johnson introduced a vaccine for fowlpox and ran a pullorum-testing laboratory. He produced numerous articles of an advisory nature on a diverse range of topics concerned with poultry and cattle. Much of Johnson's research was published in bulletins of the WWAES and OSAES that are not widely available and consequently have often been overlooked by the scientific community. Following his premature death, the director of OSAES claimed, "more information on the parasitic disease coccidiosis has been discovered at the Oregon Station than at almost any other place. The Oregon Station is probably the only institution in the world where six known species of coccidia of the chicken are available in pure culture" (Anonymous, 1938a). This is a claim that few institutions can match today. PMID- 15276977 TI - Colibacillosis in caged layer hens: characteristics of the disease and the aetiological agent. AB - In Europe, outbreaks of acute mortality in layer flocks due to colisepticaemia have frequently been observed since the mid-1990s. The aims of this study were to describe the disease, to identify the serotypes of the avian pathogenic Escherichia coli (APEC) present in these outbreaks, and to detect the presence of F11 fimbriae and flagella in the isolates. For this purpose, 20 flocks with APEC associated increased mortality and 20 control flocks matched for age were examined. Weekly mortality rates in the colibacillosis-affected flocks reached 1.71%, versus 0.30% in the control flocks. The maximum cumulative mortality over an entire colibacillosis outbreak reached 9.19%. The disease was often flock and hen house associated, with recurrent outbreaks within one round and in successive rounds in the same house. Disease was usually acute without clinical symptoms. Peritonitis with yolk material deposited in the peritoneal cavity and polyserositis were the main lesions at necropsy. O78 strains were isolated in 15 of the 20 colibacillosis flocks, and in only one of the control flocks. The majority of strains from the control flocks could not be serotyped by the 28 O antisera used. In general, F11 fimbriae and flagella were present in the majority of the strains. F11 fimbriae were significantly more often found in O78 isolates than in the other serotypes, and are thus more often present in isolates from colibacillosis flocks. Strains positive for F11, and for F11 and flagella, were more frequently present in heart and liver of the colibacillosis-affected flocks. PMID- 15276978 TI - Quantification of infectious bursal disease viral proteins 2 and 3 in inactivated vaccines as an indicator of serological response and measure of potency. AB - Viral protein 2 and viral protein 3 (VP2 and VP3) were quantified in a series of inactivated infectious bursal disease oil emulsion vaccines using enzyme-linked immunosorbent assay, and the dependence of the serological response on vaccine antigen content was studied. Large differences in antigen content, up to 50-fold, were found between vaccines. Neutralizing antibody titres at 3 to 6 weeks after vaccination varied from 3 log2 to 16 log2. None of the vaccines induced an antibody titre equal to that of the reference serum used as an indicator of sufficient potency in the European Pharmacopoeia. Neutralizing antibody titres after vaccination correlated highly with the VP2 content of the vaccines. A significant correlation was also found between the VP3 content and the antibody response. Our data illustrate that the antigen content of inactivated infectious bursal disease vaccines is a reliable indicator of the protective serological response after vaccination, and consequently could be used as a measure of vaccine potency. This holds true for both VP2, the antigen that induces neutralizing antibodies, as well as for VP3, which does not induce neutralizing antibodies. PMID- 15276979 TI - Observations on Salmonella contamination of eggs from infected commercial laying flocks where vaccination for Salmonella enterica serovar Enteritidis had been used. AB - Eggs were collected monthly from 12 cage-layer flocks on four farms where Salmonella Enteritidis was present in vaccinated flocks despite vaccination with an S. Enteritidis bacterin. Where possible, hens were also taken for culture at the end of the laying period, and faecal and environmental samples were taken from the laying houses before and after cleaning and disinfection. Twenty-four batches of six egg shells from the 13 652 tested (0.18% [0.11 to 0.26 CI(95)] single egg equivalent) were positive for S. Enteritidis and 54 (0.40% [0.30 to 0.52 CI(95)] single egg equivalent) for other serovars. Six batches of 13 640 (0.04% [0.02 to 0.10 CI95] single egg equivalent) egg contents, bulked in six egg pools, contained S. Enteritidis and three batches contained other serovars. In addition three further batches contained S. Enteritidis in both contents and shells, and two other batches contained other serovars in both. The total level of contamination by S. Enteritidis of both contents and shells found in vaccinated flocks was therefore 33 batches/13 682 eggs(0.24% [0.17 to 0.34 CI(95)] single egg equivalent). The total of contamination for any Salmonella serovar was 92 batches/13 682 eggs (0.68% [0.55 to 0.84 CI(95)] single egg equivalent). These results contrast with the findings of testing of eggs from three unvaccinated flocks prior to this study where 21 batches of egg shells from a total of 2101 eggs (1.0% [0.63 to 1.56 CI(95)] single egg equivalent) and six batches of contents from 2051 eggs (0.29% [0.11 to 0.64 CI95] single egg equivalent) were contaminated with S. Enteritidis. S. Enteritidis was found in 67/699 (9.6%) of vaccinated spent hens and 64/562 (11.4%) of bulked fresh faecal samples taken from laying houses. Failure to adequately clean and disinfect laying houses and to control mice appeared to be a common feature on the farms. PMID- 15276980 TI - In vivo studies of Gallibacterium anatis infection in chickens. AB - The aim of the present study was to investigate the pathology in normal or immunosuppressed chickens followed intravenous or intraperitoneal inoculation with a well-characterized strain of Gallibacterium anatis. Two groups of 30 15 week-old commercial brown laying chickens were used, having been screened and found negative for Gallibacterium organisms. One group was treated with 5 fluorouracil to promote heterophil depletion, while the other was saline treated. Ten days later 15 chickens from each group were inoculated either intravenously or intraperitoneally with 3.3 x 10(7) colony-forming units of G. anatis strain 12656-12. Subsets of chickens were sacrificed at 3, 12 or 24 h post-infection and examined for lesions. Livers and spleens were examined by culture and by fluorescent in situ hybridization. Intravenously infected birds showed severe septicaemic lesions in both the normal and immunosuppressed birds. Mortality was recorded only in the latter, with an overall rate of 73%. The intraperitoneally infected chickens of normal immune status showed various degrees of localized purulent peritonitis at the inoculation site, but in the immunosuppressed birds the entire peritoneum tended to be involved along with the abdominal organs. This was similar to previous descriptions of natural infections and may represent a useful infection model for detailed analysis of Gallibacterium virulence factors and pathogenesis. PMID- 15276981 TI - Susceptibility of the domestic duck (Anas platyrhynchos) to experimental infection with Toxoplasma gondii oocysts. AB - A total of 28 domestic ducks were divided into seven groups of four ducks. Six groups were inoculated per os with 10(1), 10(2), 10(3), 10(4), 10(5) and 10(5.7) oocysts Toxoplasma gondii oocysts (K21 strain, which is avirulent for mice), and the remaining group was used as a control. Antibodies to T. gondii were detected in all ducks by the indirect fluorescence antibody test first on day 7 post inoculation (p.i.). Antibody titres were found in the range of 1:20 to 1:640 depending on the infectious dose of the oocysts. From day 14 p.i. antibody titres increased to 1:80 to 1:20 480. Between days 14 and 28 p.i. (end of the experiment), antibody titres decreased in 14 ducks, remained the same in seven ducks, and continued to increase in three ducks. Bioassay in mice revealed T. gondii in the breast and leg muscles and the heart (100%, n=47), brain (91%, n=22), liver (54%, n=13) and stomach (46%, n=24). The infected ducks showed no clinical signs; however, the results of bioassay indicate that, compared with some gallinaceous birds, domestic ducks were relatively susceptible to T. gondii infection. PMID- 15276982 TI - Increased resistance of vaccinated turkeys to experimental infection with an H7N3 low-pathogenicity avian influenza virus. AB - A trial was performed to establish whether turkeys vaccinated against avian influenza with a vaccine containing a strain with a heterologous neuraminidase to the challenge virus required a higher infectious dose to develop infection than naive birds. Birds were vaccinated with a commercially available, inactivated oil emulsion product containing the strain A/ty/Italy/99/(H7N1) and challenged with different dilutions of a LPAI isolate A/ty/Italy/8000/02(H7N3) obtained during the 2002 to 2003 Italian epidemic. Groups of 10 vaccinated and 10 unvaccinated birds were infected experimentally with a virus suspension containing 10(2), 10(4) and 10(6) median embryo infective dose (EID50)/0.1 ml. Infected birds were observed daily with tracheal and cloacal swabs collected at regular intervals for antigen detection, virus isolation and real-time reverse transcription-polymerase chain reaction. Pre-infection and post-infection serology was also performed. The results of the experiment indicate that infection is achieved in naive birds with 10(4) EID50, while vaccinated birds are resistant at this challenge dose. Vaccinated and unvaccinated birds were susceptible to infection with 10(6) EID50, although the duration and/or the number of birds shedding was reduced in the vaccinated group. The data presented indicate that heterologous vaccination in the framework of a 'Differentiating Infected from Vaccinated Animals' strategy can be a valid tool to support eradication measures in areas with high densities of susceptible animals. . PMID- 15276983 TI - Phylogenetic analysis of fowl adenoviruses. AB - The sequences of the L1 loop of the hexon protein from representative fowl adenovirus (FAdV) strains of the different European and American collections were determined and compared. This study highlighted the lack of consensus in the numbering of the individual serotypes between the American and the European classifications. An identification system is proposed based on restriction fragment length polymorphism of the hexonA/hexonB polymerase chain reaction product. In addition, new insights into the relationships among FAdV strains are presented and discussed on the basis of phylogenetic analysis of the L1 loops sequences. Six clusters of strains that are supported by high bootstrap values were identified. Three of them are clearly independent, forming groups A, B and C, whereas the three others are clustered in a single 'supergroup', denominated D. Interestingly, the Japanese strain TR22 that is presently classified as European type 5 (species B) could not be assigned to any of the aforementioned clusters and might therefore constitute the sole representative of a seventh cluster. PMID- 15276984 TI - Rapid characterization of avian reoviruses using phylogenetic analysis, reverse transcription-polymerase chain reaction and restriction enzyme fragment length polymorphism. AB - A reverse transcription-polymerase chain reaction is described, which amplified the full-length sigmaC-encoding and sigmaNS-encoding genes of avian reovirus (ARV). DNA fragments of 1022 and 1152 base pairs were amplified among ARV isolates, respectively, indicating that there were no apparent deletions or insertions in these regions. Fragments amplified from vaccine strains and field isolates were digested with five different restriction enzymes Bcn I, Hae III, Taq I, Dde I, and Hinc II, respectively. Restriction fragment profiles observed on polyacrylamide gels showed heterogeneity between vaccine and Taiwanese isolates. All ARV isolates tested showed different restriction enzyme cleavage patterns and could be clearly distinguished. The strain-typing based on the cleavage sites in the sigmaC-encoding gene of ARV showed that viruses could be classified into four distinct groups. A phylogenetic tree based on the nucleotide sequences of the sigmaC-encoding gene revealed that Taiwanese ARV isolates were classified into four distinct groups, indicating that the genotyping is consistent with typing based on restriction enzyme fragment length polymorphism of the sigmaC-encoding gene of ARV. The results suggested that polymerase chain reaction followed by restriction enzyme analysis provided a simple and rapid approach for characterization of ARV isolates. Also, it is possible to determine whether a new variant strain has been introduced into a flock or a given virus strain has spread from one flock to another. PMID- 15276985 TI - Reversion of molecularly engineered, partially attenuated, very virulent infectious bursal disease virus during infection of commercial chickens. AB - A molecularly cloned, tissue culture-adapted infectious bursal disease virus (BD 3tc) was generated from a very virulent strain by the reverse genetics approach following site-directed mutagenesis (Q253H and A284T in VP2). The pathogenicity of BD-3tc was tested in commercial chickens. The wild-type strain (BD-3wt) and the molecularly cloned parental strain (BD-3mc) were included for comparison. The subclinical course of the disease, with delayed and milder pathological lesions followed by quick follicular regeneration in the bursa of Fabricius in BD-3tc inoculated birds, suggested that these amino acid substitutions made BD-3tc partially attenuated. However, severe bursa atrophy was observed at 14 days after inoculation. Reverse transcription-polymerase chain reaction coupled with restriction enzyme analysis revealed that both point mutations in BD-3tc had reverted 14 days after inoculation. Further investigations demonstrated that the codon for amino acid at position 284 had already reverted to the wild-type phenotype (T284A) 3 days after inoculation. PMID- 15276986 TI - Characterization of four very virulent Argentinian strains of Marek's disease virus and the influence of one of those isolates on synergism between Marek's disease vaccine viruses. AB - Isolates of Marek's disease virus (MDV) from vaccinated flocks in Argentina were characterized as very virulent (vv) and very virulent plus (vv+) strains. Experimental infection with these viruses caused a high incidence of Marek's disease in both resistant N-2a line and susceptible P-2a line birds. Vaccine viruses from each of the three Marek's disease viral serotypes were evaluated alone and in various combinations for protection against challenge with a vvMDV called NULP-1. Vaccination of P-2a birds with HVT did not protect satisfactorily against any of the vv and vv+MDV strains isolated. However, CVI988/Rispens vaccine alone or combined with serotype 2 and/or serotype 3 vaccine strains enhanced protection significantly against NULP-1. Serotype 2 plus serotype 3 vaccines also provided significant protection when challenged with this strain. This is one the first reports of the occurrence of vvMDV and vv+MDV in Argentina and Latin America. It is also a preliminary evaluation of the synergistic protective effect of different vaccine viruses with local MDV strains. However, further studies are needed to evaluate the real role of these and other Marek's disease isolates in 'vaccination failures' and the influence of serotype and virus strain on synergism between Marek's disease vaccine viruses. PMID- 15276987 TI - A granulomatous conjunctivitis associated with Morexella phenylpyruvica in an ostrich (Struthio camelus). AB - The aim of study was to evaluate a case of granulomatous conjunctivitis, clinically and pathologically, in the right eye of a 2-year-old, female ostrich. A mass measuring 5 cm x 3 cm x 4 cm was removed surgically from the eye of the ostrich. Morexella phenylpyruvica was recovered from the mass. On histopathological examination, hyperplasia or squamous metaplasia in some area of conjunctival palpebra, and a granulomatous inflammation in the submucosa were observed. The lesion was described as a granulomatous conjunctivitis caused by M. phenylpyruvica. The lesion was located in the lower eyelid conjunctiva and was not only restricted to the gl. lacrimalis, but also present in the connective tissue. After excision of the mass, the ostrich was treated with topical and systemic antibiotics and corticosteroid. The ostrich recovered fully and the function of the eye appeared to be normal. PMID- 15276988 TI - Hafnia alvei infection in pullets in Italy. AB - This paper describes an outbreak of disease caused by Hafnia alvei in pullets. Cloudy swelling and the fatty degeneration of liver associated with splenic lymphocytic depletion were the most preminent lesions. The organism was identified by biochemical tests and by a Hafnia-specific bacteriophage test. Laying hens and pullets were infected experimentally with the organism by the oral and intraperitoneal route, and the clinical and pathological effects were similar to those observed in naturally infected subjects. The use of more sensitive diagnostic tests is suggested to avoid the possibility of a misdiagnosis due to similarities between this organism and Salmonella species. PMID- 15276989 TI - Dogs as potential carriers of infectious bursal disease virus. AB - In this study, the possibility that dogs could eventually be carriers of infectious bursal disease virus (IBDV) after having eaten (voluntarily or accidentally) IBDV-infected chicks has been evaluated. A single Beagle dog was fed chicks infected by a very virulent IBDV strain (vvIBDV). Afterwards, the presence and viability of IBDV in the faeces was assessed. Viable vvIBDV was detected in the dog's faeces for 2 days after the initial ingestion, which indicates excretion of vvIBDV. Comparison by molecular techniques of the administered and excreted virus using reverse transcription-polymerase Chain reaction and enzymatic digestion confirmed that the initial virus maintained the same characteristics after being excreted. We believe that this study could be of great interest for a better understanding of the epidemiology of IBD disease on farms where dogs live close to avian facilities. PMID- 15276990 TI - Aerosolization of Mycoplasma synoviae compared with Mycoplasma gallisepticum and Enterococcus faecalis. AB - In order to study the airborne transmission of an arthropathic strain of Mycoplasma synoviae, preliminary aerosol experiments were performed. They were conducted in duplicate in an empty isolator (1.3 m3) to assess the yield and viability of M. synoviae with time compared with Mycoplasma gallisepticum and Enterococcus faecalis. After aerosol generation air samples were taken with two different devices using gelatine or cellulose nitrate filters. There was no difference between the devices, but cellulose nitrate filters yielded very low bacterial counts. The aerosolized dose per isolator for M. synoviae was 3.4 x 10(10) colony-forming units (cfu), for M. gallisepticum was 2.6 x 10(10) cfu and for E. faecalis was 3 x 10(10) cfu. Immediately after aerosolization, concentrations of about 10(6) to 10(7) cfu/m3, 10(7) to 10(8) cfu/m3 and 10(8) to 10(9) cfu/m3 air of M. synoviae, M. gallisepticum and E. faecalis were found, respectively. At 25 min M. synoviae concentrations dropped below the detection level (<4 x 10(4) cfu), while 10(5) to 10(6) and 10(8) to 10(9) cfu were found for M. gallisepticum and E. faecalis, respectively. The average M. synoviae concentration during the experiment was estimated at 10(2) to 10(3) cfu/l. The M. gallisepticum and E. faecalis aerosol generated an average of approximately 10(3) to 10(4) cfu/l air and 10(5) to 10(6) cfu/l air, respectively. Thus mycoplasma and E. faecalis aerosols were successfully generated despite considerable initial loss as measured by culture. The loss was greater in the mycoplasma aerosols, especially those of M. synoviae. PMID- 15276991 TI - Newcastle disease in geese: natural occurrence and experimental infection. AB - A novel disease entity in domestic geese with unusual epidemiology and pathological changes has emerged in some regions of China since the late 1990s. Velogenic Newcastle disease (ND) viruses were isolated from visceral organs and cloacal swabs of diseased birds in the field. Gross lesions of diseased geese were characterized by extensive necrotic foci in intestinal mucosa, spleen and pancreas. Histological examination revealed multisystemic distribution of lesions. Similar disease was reproduced experimentally in geese not only with two field isolates of goose origin, but also with ND viruses belonging to genotypes VI, VII, VIII and IX. These led us to define the novel disease as clinical ND in geese. Additionally, the goose-originated ND virus isolates could be transmitted from geese to specific pathogen free chickens, illustrating the potential threat they posed to the chicken industry. This is the first detailed report of the natural outbreaks and experimental reproduction of ND in geese, providing evidence that geese could play an important role in the epidemiology of ND. PMID- 15276992 TI - Heat inactivation of Newcastle disease virus (strain Herts 33/56) in artificially infected chicken meat homogenate. AB - Heat inactivation curves were constructed for Newcastle disease virus strain Herts 33/56 in artificially infected meat homogenate at 60 degrees C, 65 degrees C, 70 degrees C, 74 degrees C and 80 degrees C. For the four higher temperatures the time taken to reduce the infectivity by 90% (1 log10) at the specified temperature (Dt) were estimated as: D65=120 sec, D70=82 sec, D74=40 sec and D80 29=sec. PMID- 15276993 TI - Coronary artery rupture in male commercial turkeys. AB - Mortalities of 3% in a 1-week period and 2.5% in a 4-week period occurred in two flocks of 13-week-old to 16-week-old male turkeys, respectively. Eleven of 18 birds submitted to the laboratory for necropsy had large amounts of clotted blood in the pericardial sac and transverse bands of haemorrhage at the base of the left heart. Three other birds had clotted blood in the abdominal cavity due to rupture of the aorta. Histopathology of the hearts with haemorrhage at the base of the left heart revealed medial degeneration, necrosis and rupture of the coronary arteries. Special stains revealed decrease in elastic fibres and increase in fibrous connective tissue in the coronary arteries. Fourteen of 16 birds had a low copper concentration (<5 mg/kg wet weight) in the liver; the zinc concentration was high in one bird. Levels of heavy metals including copper and selenium in the feed were within normal ranges for poultry. No mycotoxins were detected in the feed. Tests for bacterial and viral pathogens showed no significant findings. The average weight of these turkeys at market at 16 weeks was 12.5 kg, which was considered 1.4 to 2.3 kg higher than normal. It is well known that male turkeys are prone to hypertensive angiopathy. Therefore, it is probable that higher body weights in the turkeys in conjunction with the hypertensive angiopathy and low levels of copper may have predisposed the birds for coronary artery rupture. Genetic diseases such as connective tissue disorders of the elastin and/or collagen were also considered as possible causes. PMID- 15276994 TI - Methods for evaluating and developing commercial chicken strains free of endogenous subgroup E avian leukosis virus. AB - The genome of nearly all chickens contains various DNA proviral insertions of retroviruses of subgroup E avian leukosis virus (ALVE). However, the elimination or control of ALVE gene expression is desirable to improve productivity, to improve resistance to avian leukosis virus (ALV)-induced tumours, and to develop safer live virus vaccines in chick embryos and cultured chick cells. Restriction fragment length polymorphism and polymerase chain reaction methods are used to define the presence of ALVE genes; and the expression of ALVE in chicken plasma or on cells, and the susceptibility of cells to ALVE is determined by flow cytometry using a specific (R2) antibody. ADOL line 0 chickens have been selected to be free of ALVE genes, while being resistant (i.e. lack receptors to ALVE), but susceptible to exogenous ALV (i.e. ALVA, ALVB, ALVC and ALVJ). To develop improved line 0-type chickens, ADOL line 0 was outcrossed to a commercial line that had one ALVE gene and evidence for ALVE resistance. Rous sarcoma virus (RSV) challenge was used to confirm resistance of F1 chickens to ALVE, and susceptibility of F2 breeders to ALVA and ALVB using test chicks produced by matings to line 7(2). Selected F2 breeders were resistant to ALVE, but susceptible to exogenous ALVA, ALVB, ALVC and ALVJ, based on challenge tests of progeny chick cells using an enzyme-linked immunosorbent assay. The new line, 0(1), has evidence for improved egg size, productivity, fertility and hatchability. Similar procedures may be used for development of productive ALVE free chicken lines with preferred ALV susceptibility traits. PMID- 15276995 TI - Epizootic occurrence of haemorrhagic nephritis enteritis virus infection of geese. AB - Recent outbreaks of haemorrhagic nephritis enteritis in geese flocks of 3 to 10 weeks in age in Hungary were investigated. Mortality varied between 4% and 67%. Affected birds generally died suddenly. Occasional clinical signs included tremors of the head and neck, subcutaneous haemorrhages and excretion of faeces containing partly digested blood. At necropsy the most frequent findings were a turgid wall and reddish mucosa of the intestines and reddish discolouration of the swollen kidneys, but oedema and haemorrhages of the subcutaneous connective tissue, hydropericardium and ascites were also seen. In subacute cases, visceral gout was frequently observed. Histological examination revealed zonal necrosis of the tubular epithelial cells with haemorrhages in the kidney. Other histological findings were serous hepatitis with fatty infiltration, necrotizing haemorrhagic enteritis and haemorrhages in the different organs including the brain. Experimental geese infected parenterally with crude liver and spleen homogenates prepared from diseased birds died after 8 to 20 days without premonitory signs, and had typical gross and histological lesions. Attempts to isolate cytopathic virus on different tissue cultures failed. The presence of polyomavirus was proven by polymerase chain reaction. Five isolates were further investigated by analysing their complete VP1 gene sequence. All tested strains were very closely related to each other on the basis of the nucleotide sequence, and they were identical at the deduced amino acid level. PMID- 15276996 TI - Salmonella gallinarum gyrA mutations associated with fluoroquinolone resistance. AB - Salmonella enterica subsp. enterica serovar Gallinarum (S. gallinarum) is the causative organism of fowl typhoid, and an outbreak of fowl typhoid in Korea was confirmed in 1992. The aim of this study was to investigate possible changes in fluoroquinolone susceptibility among S. gallinarum isolates from 1995 to 2001, and to analyse mutations of the gyrA gene in fluoroquinolone-resistant isolates. Among 258 S. gallinarum isolates tested by the disk diffusion method, isolates from 1995 (n=18) were susceptible to all fluoroquinolones tested, whereas a number of isolates from 2001 (n=46) showed reduced susceptibility to enrofloxacin (6.5%), ciprofloxacin (10.9%), norfloxacin (52.5%) and ofloxacin (82.6%). The minimum inhibitory concentration range of enrofloxacin, ciprofloxacin, norfloxacin, ofloxacin and danofloxacin increased from < or =0.06 approximately 0.25 microg/ml in 1995 to 2 approximately 8 microg/ml in 2001. When amino acid changes in the gyrA were analysed by DNA sequencing, 22.5% and 14.7% among 258 isolates had a mutation at the Ser-83 and Asp-87 codons, respectively, and the prevalence of these mutants increased from 5.6% in 1995 to 89.1% in 2001. These mutants contained a change from Ser to Phe or Tyr at codon 83, or a change from Asp to Gly, Tyr or Asn at codon 87, and showed a range of minimum inhibitory concentrations of enrofloxacin from 0.5 to 8 microg/ml, ciprofloxacin from 0.25 to 4 microg/ml, norfloxacin from 2 to 32 microg/ml, ofloxacin from 0.5 to 4 microg/ml, and danofloxacin from 0.5 to 4 microg/ml. These results suggested an important association between the gyrA mutations and fluoroquinolone resistance of S. gallinarum. PMID- 15276997 TI - A molecular epidemiological investigation of isolates of the variant avian paramyxovirus type 1 virus (PPMV-1) responsible for the 1978 to present panzootic in pigeons. AB - A sequence of 375 nucleotides, which included the region encoding the cleavage activation site and signal peptide of the fusion protein gene, was determined for 178 isolates of the pigeon variant strain of Newcastle disease virus (PPMV-1). These were compared with the sequences of 47 similar isolates published by GenBank, which included 30 isolates from pigeons and 17 representatives from each sublineage of avian paramyxovirus type 1. The resulting alignment was analysed phylogenetically using maximum likelihood and the results are presented as unrooted phylogenetic trees. By phylogenetic analysis all the PPMV-1 isolates except one were placed in lineage 4b (VIb). Within this lineage there was considerable genetic heterogeneity, which appears to be predominantly influenced by the date of isolation and, to a lesser extent, geographical origins of the isolates. There were two large distinguishable groups, 4bi and 4bii. The earliest isolate available, PIQPI78442, isolated in 1978 in Iraq, was situated at the node from which the two groups diverge. PMID- 15277000 TI - Stress/aging: endocrine profiles/reproductive dysfunction in men. AB - Extensive investigations have been carried out on the biology/physiology of stress [13, 18, 20, 25, 35, 48, 49, 53, 55], pathophysiology of aging [41], sexual dysfunction [7, 27] and adrenal insufficiency [4, 31]. PMID- 15277001 TI - Effects of semen characteristics on IUI combined with mild ovarian stimulation. AB - To determine the influence of sperm parameters inseminated on the outcome of intrauterine insemination (IUI) in patients undergoing ovarian stimulation with clomiphen citrat (CC) or human menopausal gonadotropin (HMG) therapy, a retrospective review was performed for 2 years on data from the IUI program. 190 couples underwent a total of 268 IUI cycles in which CC or HMG was used for ovulation induction. The initial sperm concentration (mil/ml), motility (percent), preprocessing total motile sperm (TMS) count (million), fast motile sperm (percent) and postprocessing sperm concentration (mil/ml), motility (percent), TMS count, fast motile sperm (percent), sperm morphology, hypoosmotic swelling (HOS) scores, semen leuocytes, and bacteria were analyzed. 268 inseminations were followed by a pregnancy rate of 12% and couple pregnancy rate of 17%. On multivariable logistic regression analysis, total motile sperm (TMS) count, percent motility, and percent of fast motile sperm were independent prognostic factors of fertility. The impact of the preprocessing and postprocessing sperm parameters on pregnancy outcome after IUI was evaluated. There was a trend toward an increasing percent of conception with increasing TMS count, motility, and percent of fast motile sperm. The TMS count, motility and percent of fast motile sperm independently predict success with IUI. Patients with original sperm motility > or = 30% had a higher cumulative pregnancy rate (74%) than patient with motility < 30% (p < 0.005). Pregnancy rate increased 4 times with motility of > or = 30%. PMID- 15277002 TI - Is routine hormonal measurement necessary in initial evaluation of men with erectile dysfunction? AB - To prospectively compare serum hormone levels and the incidence of hormonal pathologies between men with and without erectile dysfunction, and investigate risk factors that might predict hormonal pathologies in men complaining of erectile dysfunction. The study included 262 men with erectile dysfunction and 53 healthy men with no erectile dysfunction as a control group. All men enrolled in the study were evaluated with a detailed history, physical examination, international index of erectile function (IIEF-5), and serum hormone measurement. Hypotestosteronemia was considered as serum total testosterone value of < 3 ng/mL, and hyperprolactinemia was considered as serum prolactin level of > 18 ng/mL. Serum hormone levels and the incidence of hormonal abnormalities were compared between the two groups. In addition, risk factors for hormonal abnormalities were investigated. There were no significant differences in the mean serum FSH (p = 0.212), LH (p = 0.623), testosterone (p = 0.332) and prolactin values (p = 0.351) between the men with and without erectile dysfunction. Hypotestosteronemia was detected in 29 (11%) of the erectile dysfunction group and in 2 (3.7%) of the control group, revealing no significant difference (p = 0.104). Hyperprolactinemia was detected in 25 (9.5%) of the erectile dysfunction group and in 2 (3.7%) of the control group, revealing no significant difference (p = 0.171). To investigate risk factors that might predict hormonal pathologies, there were no significant differences in the patient age, duration of the sexual dysfunction, smoking history and duration, the presence of chronic disease and the type of erectile dysfunction. Our findings suggest that hormonal measurement should not be routinely performed in the initial evaluation of men presenting with erectile dysfunction, and may be necessary based only on the findings obtained with a careful history and physical examination. PMID- 15277003 TI - Clinical efficacy and safety of sildenafil in elderly patients with erectile dysfunction. AB - Erectile dysfunction (ED) is a common medical disorder affecting elderly men. Sildenafil citrate has been shown to be an effective and well-tolerated oral agent for treating ED in the general population of adult men with ED of broad spectrum etiology. Elderly men are more likely to have concomitant medical problems than the general population of men with ED. In this study, we examined the efficacy and safety of sildenafil administration in elderly patients with ED. Forty-four elderly men with ED (> or = 60 years old) of broad-spectrum etiology were treated with 25 mg or 50 mg doses of sildenafil citrate. Age ranged from 60 to 78 years (65 +/- 4.5; means +/- S.D.). Mean follow-up period was 12.3 +/- 6.5 months, with a range of 1 to 25 months. Primary efficacy assessments were performed using the International Index of Erectile Function 5 (IIEF5) before their first dose of sildenafil and after at least 4 weeks of therapy. Serum testosterone was measured before treatment. The mean IIEF5 among all patients increased from 8.5 +/- 3.9 to 20 +/- 4.2 after sildenafil use (P < 0.0001). In patients younger than 70 years, the IIEF5 score increased from 9.5 +/- 5.0 to 17 +/- 4.3 while in patients 70 years and older, the score increased from 8.2 +/- 3.6 to 21 +/- 3.9, a near normalization. The rate of improvement in younger men was higher than in older men. Serum testosterone before treatment was similar in the two groups. The most commonly experienced adverse events were flushing and dyspepsia, which occurred in 6.8% and 2.3%, respectively. No patients discontinued sildenafil treatment due to adverse events. In conclusion, oral sildenafil is efficacious and well tolerated by elderly men with ED, even among those older than 70 years. PMID- 15277004 TI - Chromosomal abnormalities and polymorphisms in Mexican infertile men. AB - A cross-sectional study was conducted to estimate the prevalence of chromosome abnormalities and normal variable chromosome features (polymorphisms) in infertile men from northeastern Mexico. Karyotyping was carried out in 326 men with diagnosis of infertility. The sperm counts showed 204 patients with oligozoospermia, 87 with azoospermia and 35 normozoospermia. Five patients with oligozoospemia and two with azoospermia presented chromosome abnormalities. Nonzoospermic men did not show chromosomal abnormalities. Polymorphisms of heterochromatin and satellite length showed a significant increased in oligozoospermic and azoospermic men with respect to normozoospermic men, respectively. This study reports the prevalence of chromosome abnormalities, polymorphisms of heterochromatin length, and polymorphisms in satellites in Mexican infertile men. The prevalence in this study was similar to other studies in world literature. PMID- 15277005 TI - Induction of spermatogenesis in idiopathic hypogonadotropic hypogonadism with gonadotropins in older men. AB - We investigated the treatment results in 6 azoospermic idiopathic hypogonadotropic hypogonadism (IHH) cases that remained untreated 41-47 years of age. Medical history, physical examination, hormone profile measurements, peripheral blood karyotype, skull X-ray and/or magnetic resonance imaging were performed. Patients received 1,000 to 5,000 IU hCG, 2-3 times per week, and 75 to 150 IU hMG, 2-3 times per week for 24 months. Serum testosterone levels were assessed every month for maximum 6 months to evaluate optimal dose of treatment and then every 3 months thereafter. Sperm counts were assessed every 3 months. Testosterone level increased from 2.7 +/- 0.9 mIU/L to 22 +/- 7.04 mIU/L with treatment; testicular volume increased by 4.6 ml during the treatment. Sperm were detected in the ejaculate in 3 out of 6 patients on the 22nd, 18th, and 15th month of treatment. 3 patients underwent testicular biopsy; histopathology revealed tubular hyalinization. Spermatogenesis in older men with IHH was restored by exogenous gonadotropins. PMID- 15277006 TI - Fluorometric study of rabbit sperm head membrane phospholipid asymmetry during capacitation and acrosome reaction using Annexin-V FITC. AB - This study was conducted to evaluate phosphatidylserine translocation in head plasma membrane of Percoll-gradient purified of rabbit cauda epididymal sperm during capacitation and acrosome reaction (AR) using Annexin-V. Propidium iodide was used as control to reject dead or dying cells. The presence and distribution of Annexin-V binding sites were analyzed using flow fluorocytometry and confocal microscopy. After 6 h of incubation of sperm in capacitation medium, the number of cells positively stained with Annexin-V showed a small but significant increment. The Annexin-V binding sites produced during capacitation were found mainly in the post-acrosomal region of the sperm head plasma membrane. After AR induction with progesterone, the localization of phosphatidylserine was changed and the Annexin-V binding sites were found almost only in the acrosomal region, but with higher number of binding sites in the equatorial area. On the contrary, after AR induction with A23187, phosphatidylserine translocation, although predominant over the acrosomal region, was also observed in the post-acrosomal region. Plasma membrane destabilization during capacitation and AR may be important for sperm-oocyte fusion. PMID- 15277007 TI - Expression of p50 C-terminal Src kinase (Csk) in mouse testis. AB - C-terminal Src Kinase (Csk) is a cytoplasmic tyrosine kinase that phosphorylates a critical tyrosine residue in each of the Src family kinases to inhibit their activities. To investigate the possible regulation of spermatogenesis by Src-Csk loop, the postnatal changes in the expression of Csk were examined in mouse testes. Semiquantitative RT-PCR analysis revealed that Csk mRNA increased during neonatal development and peaked at 2 weeks of age. Following the decrease during pubertal development, Csk expression re-increased in adult testes. In Western blot, immature testes showed higher expression of Csk protein than the pubertal or adult testes. In immature testis, Csk immunoreactivity was largely found in the Sertoli cell and there was no visible difference in the Csk immunoreactivity among the seminiferous tubules. In adult testis, however, a differential Csk immunoreactivity was found among the seminiferous tubules. Intense signal was found in the adluminal cytoplasm of the Sertoli cells bearing the post-meiotic differentiating germ cells, suggesting that Csk may participate in the remodeling of seminiferous tubule during late phase of spermatogenesis. Csk immunoreactivity was also found in the Leydig cells, suggesting the possible regulation of Leydig cell function. Src-Csk loop may participate in the differentiation of the seminiferous epithelia and Leydig cells in mouse testis. PMID- 15277008 TI - Erectile function after brachytherapy with external beam radiation for prostate cancer. AB - The effect of therapeutic modalities on sexual potency is an important consideration for patients choosing a treatment for prostate cancer. We assessed erectile function after iridium-192 (1r-192) high-dose rate (HDR) brachytherapy with external beam radiation therapy (EBRT), and examined the efficacy of sildenafil after this treatment. Forty-two prostate cancer patients (T1c to T3bN0M0) were treated with 22Gy HDR brachytherapy with 36.8Gy EBRT without neoadjuvant hormone therapy. Erectile function was assessed using a 5-item version of the International Index of Erectile Function questionnaire (IIEF-5), pre, 3 and 12 months after treatment, Potency was defined as an IIEF-5 score > or = 11. Ten patients with potency before HDR brachytherapy with EBRT with or without neoadjuvant hormone therapy requested Sildenafil 3 months after treatment. The mean IIEF-5 score of all patients was 10.5 +/- 8.5, 4.5 +/- 5.3 (p < 0.001), and 3.8 +/- 4.7 (p < 0.001), pre, 3 and 12 months after treatment, respectively. Seventeen (40.4%) patients were potent before treatment. The mean IIEF-5 score of those patients was 15.8 +/- 3.2, 9.6 +/- 5.1 (p = 0.04), and 11.3 +/- 6.1 (p = 0.06), pre, 3 and 12 months after treatment, respectively. Ten of 17 (58.8%) patients maintained their potency 12 months after treatment. In 10 patients with potency before treatment who were treated with sildenafil, the mean IIEF-5 score increased from 6.2 +/- 3.5 at 3 months to 13.6 +/- 5.1 (p < 0.001) at 12 months after treatment. Eight of 10 (80%) patients treated with sildenafil had recovered 12 months after treatment. HDR brachytherapy with EBRT can be performed with favorable results for maintaining potency. PMID- 15277009 TI - Detection and bioimaging of nitric oxide in bovine oocytes and sperm cells. AB - Mammalian gametes contain constitutive nitric oxide synthases (NOS) to synthesize nitric oxide (NO). The detection and bioimaging of NO in bovine gametes is important to determine the regulatory roles of NO during the different events of fertilization. Diaminofluoresceins, new fluorescence indicators for NO, were applied to detect the release of NO from bovine gametes. These compounds yield green fluorescent triazolofluoresceins, which provide the advantages of high specificity, sensitivity, and simplicity for the detection of NO. In this study, we mapped the expression of NOS in the bovine sperm and ova. NOS activity in sperm first appeared in the acrosome, then 60 min later in the head, middle piece, cytoplasmic droplet, and tail. Cow ova had high NO activity in the cytoplasm and in the surrounding corona cells, but not in the zona pellucida. These results show that for bovine gametes, the synthesis NO by the NOS system presents clear patterns of time and spatial distribution that may be important for the different events of fertilization. PMID- 15277010 TI - Correlation of sperm parameters with apoptosis assessed by dual fluorescence DNA integrity assay. AB - Failed fertilization after intracytoplasmic sperm injection or miscarriages occurs in cases involving apoptotic and necrotic sperm. Identifying normal sperm is important for successful assisted reproductive technologies (ART) procedures. The study was conducted to correlate sperm parameters with intact sperm with normal DNA assessed by the dual stain assay in 118 separate individuals. The results showed differences in percent DNA intact sperm in individuals with normal W.H.O. sperm features (62 +/- 1.1; mean +/- S.E.M.) compared with oligoasthenoteratozoospermia patients (38 +/- 5.3). Individuals whose sperm had fertilizing capacity had higher percentages of intact DNA (60 +/- 1.3 versus 47 +/- 2.4). The percentages of intact DNA sperm were significantly correlated to total motility in semen (R = 0.7), post-wash motility (R = 0.6), rapid progression (R = 0.6), intact acrosome (R = 0.5), and strict morphology (R = 0.5). There were no correlations with the remaining parameters. The dual stain assay identified sperm with normal physiology and fertilizing capacity. The dual stain assay measures DNA integrity and is a promising method to select normal sperm for ART. PMID- 15277012 TI - Quantitative structure-activity relationships within a homologous series of 7 alkoxyresorufins exhibiting activity towards CYP1A and CYP2B enzymes: molecular modelling studies on key members of the resorufin series with CYP2C5-derived models of human CYP1A1, CYP1A2, CYP2B6 and CYP3A4. AB - 1. The results of quantitative structure-activity relationships (QSARs) within a homologous series of 7-n-alkoxyresorufins are reported. They are consistent with homology modelling of the relevant P450s involved in their metabolism. 2. QSARs were generated for activities involving CYP1A and CYP2B enzymes with structural descriptors relating to compound planarity and other shape parameters, together with certain features of the n-alkoxyresorufin electronic structure, especially electron densities and superdelocalizabilities. 3. A quadratic relationship between compound lipophilicity and binding to CYP2B enzymes was apparent, and which indicated maximal interaction for 7-pentoxyresorufin. Such indications help to explain enzyme selectivity in terms of optimal alkyl chain length for fitting within the relevant active site region. 4. Calculation of the binding affinities for methoxy-, ethoxy-, pentoxy- and benzyloxy-resorufins towards either CYP1A2 or CYP2B6 enzymes, depending on the 7-alkoxyresorufin agree favourably with experimental values obtained from K(m) determinations. PMID- 15277013 TI - Molecular modelling of CYP2A enzymes: application to metabolism of the tobacco specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK). AB - 1. Tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) is a lung carcinogen in a variety of animal models and a putative human lung carcinogen. Its tumorigenic potential is unmasked via cytochrome P450 (CYP) mediated hydroxylation of the carbon atoms adjacent to the nitroso moiety (i.e. alpha-hydroxylation). Therefore, elucidation of enzyme-substrate interactions that facilitate alpha-hydroxylation is important to gain insight into the tumorigenic mechanism of NNK and to develop potent inhibitors of this detrimental reaction. 2. Molecular models of CYP2A enzymes from mice, rats and humans that are catalysts of NNK bioactivation were constructed and used, in conjunction with docking experiments, to identify active-site residues that make important substrate contacts. 3. Docking studies revealed that hydrophobic residues at positions 117, 209, 365 and 481, among others, play critical roles in orienting NNK in the active site to effect alpha-hydroxylation. These molecular models were then used to rationalize the stereo- and regioselectivity, as well as the efficiency, of CYP2A-mediated NNK metabolism. PMID- 15277014 TI - In vitro assessment of intestinal permeability and hepatic metabolism of 4' bromoflavone, a promising cancer chemopreventive agent. AB - 1. The intestinal permeability and hepatic metabolism of the investigational cancer chemoprevention agent 4'-bromoflavone were investigated in vitro using human intestinal Caco-2 cell monolayers, human liver microsomes and human hepatocytes. Liquid chromatography-mass spectrometry and tandem mass spectrometry were used for quantitative analysis in support of the Caco-2 cell studies and for the characterization of metabolites of 4'-bromoflavone. 2. The Caco-2 cell model indicated that 4'-bromoflavone would be absorbed by the intestine at a moderate rate by means of direction-independent, passive diffusion. There was no indication of active transport or efflux. 3. Three monohydroxylated metabolites and one monohydroxylated, hydrated metabolite of 4'-bromoflavone were detected at relatively low levels in the human liver microsomal and hepatocyte incubations. The structures of these metabolites were confirmed by comparison with synthetic standards. Hydroxylation occurred on the A-ring of 4'-bromoflavone but not on the B-ring, probably due to deactivation of the B-ring by bromine. No phase II metabolites were detected following incubation of 4'-bromoflavone in these in vitro systems. 4. In conclusion, these studies predict that 4'-bromoflavone should show moderate oral bioavailability, and that it would probably be excreted as unchanged compound and monohydroxylated metabolites. The results might be helpful in the design of clinical trials and in the interpretation of pharmacokinetic studies of 4'-bromoflavone. PMID- 15277015 TI - Homology modelling of CYP3A4 from the CYP2C5 crystallographic template: analysis of typical CYP3A4 substrate interactions. AB - 1. The results of homology modelling of cytochrome P4503A4 (CYP3A4), which is a human enzyme of major importance for the Phase 1 metabolism of drug substrates, from the CYP2C5 crystal structure is reported. 2. The overall homology between the two protein sequences was generally good (46%) with 24% of amino acid residues being identical and a 22% similarity between matched pairs in the CYP3A4 and CYP2C5 aligned sequences, thus indicating that CYP2C5 represents a viable template for modelling CYP3A4 by homology. 3. The CYP3A4 model appears to show consistency with the reported findings from the extensive site-directed mutagenesis studies already published. 4. Typical CYP3A4 substrates, such as midazolam, testosterone, nifedipine and verapamil, are shown to fit the putative active site of the enzyme structure in a manner consistent with their known positions of metabolism. PMID- 15277016 TI - Prediction of clinical pharmacokinetic parameters of linezolid using animal data by allometric scaling: applicability for the development of novel oxazolidinones. AB - 1. Allometric scaling has previously been used as an effective tool for the prediction of human pharmacokinetic data. The pharmacokinetic data for linezolid, a novel oxazolidinone to treat Gram-positive pathogens, in mice, rats and dogs were subjected to simple allometric scaling. Generated allometric equations for parameters such as clearance (CL), volume of distribution (Vss) and elimination rate constant (K10) were used to predict human pharmacokinetic parameters including elimination half-lives. In addition, the human plasma concentration time curve was simulated using a one-compartmental model. 2. Application of simple allometry (Y = aWb) for animal parameters such as CL, Vss, and K10 showed excellent allometric fit (r > or = 0.98). The allometric equations for CL, Vss, and K10 were -0.5465W(0.6595), -0.1369W(0.9246), and -0.4117W(-0.3139), respectively. The confidence in predictability of CL and Vss parameters was particularly high since the allometric exponents of CL and Vss almost approached the suggested values of 0.75 and 1.00, respectively. 3. Animal pharmacokinetic parameters generated in the present authors' laboratories for linezolid were in close agreement with reported literature values. The predicted human values for CL (4.68 l h(-1)), Vss (37.07 litres), and K10 (0.10 h(-1)) were within the range observed for linezolid in the literature (CL = 4-10.5 l h(-1); Vss = 21-53 litres; K10 = 0.09-0.3 h(-1)). The human half-life (t(1/2)) predicted using allometry (6.9 h) was similar to reported values in humans of 5 h. In summary, the retrospective analysis for linezolid suggests that allometric scaling can be used as a prospective tool for predicting human pharmacokinetic parameters of novel oxazolidinones. PMID- 15277017 TI - Pharmacokinetics of two major hydroxylated polychlorinated biphenyl metabolites with specific retention in rat blood. AB - 1. Hydroxylated metabolites of polychlorinated biphenyls (OH-PCBs) are, depending on their structure, strongly retained in mammalian, fish and bird blood. This is due to strong, though reversible, binding to the thyroxine binding and transporting protein transthyretin. 2,3,3',4',5-Pentachloro-4-biphenylol (4-OH-CB 107) and 2,2',3,4',5,5',6-heptachloro-4-biphenylol (4-OH-CB 187) are two of five major OH-PCB congeners in human plasma. 2. The relative amounts of OH-PCB congeners vary between species and also between human populations, in spite of similar PCB congener patterns, and may depend on different pharmacokinetic parameters of the OH-PCBs. In the present study, the pharmacokinetic parameters of 4-OH-CB 107 and 4-OH-CB 187 were determined in the rat after a single intravenous dose of 1 micromol kg(-1). Plasma samples were analysed by gas chromatography/mass spectrometry. 3. 4-OH-CB 107 had a half-life of 3.8 days; 4 OH-CB 187 had a half-life of 15 days. Volumes of distribution were 0.07 and 0.11 l kg(-1), respectively; clearances (ml h(-1)) were 0.67 and 0.22, respectively; and the areas under the curve were estimated as approximately 1500 and 4450 nmol h ml(-1). 4. The pharmacokinetic parameters thus determined help to explain the observed differences in the relative amounts of OH-PCBs in humans and other mammals exposed to environmental PCBs. PMID- 15277018 TI - Metabolism of the antineoplastic and immunosuppressive drug 2-CdA (Leustatin) in animals and humans. AB - 1. The in vivo metabolism of the antineoplastic and immunosuppressive drug 2-CdA (Leustatin) was investigated in mice, monkeys and humans after a single subcutaneous dose of cladribine 60 mg kg(-1) to eight male and eight female mice and 10 mg kg(-1) to one male and one female monkey, and an intravenous infusion dose of cladribine 22-45 mg(-1) per subject to 12 male patients. 2. Plasma (1 h), red blood cells (1 h) and faecal samples (0-24 h) were obtained from mice and monkeys, and urine samples (0-24 h) were obtained from these species and humans. 3. Unchanged cladribine (urine: 47% of the sample in human; 60% of the sample in mouse; 73% of the sample in monkey) and 10 metabolites, consisting of four phase I metabolites (M1-3, M7) and six phase II metabolites -- five glucuronides (M4, M6, M8-10) and one sulfate (M5) -- were profiled, characterized and tentatively identified in plasma, red blood cells, and faecal and urine samples on the basis of API ionspray-mass spectrometry (MS) and MS/MS data. 4. Metabolites were formed via the following three metabolic pathways: oxidative cleavage at the adenosine and deoxyribose linkage (A); oxidation at adenosine/deoxyribose (B); and conjugation (C). 5. Pathways A and B appear to be major steps, forming four oxidative/cleavage metabolites (M1-3, M7) (each 3-20% of the sample). 6. Pathway C along or in conjunction with pathways A and B produced cladribine glucuronide, cladribine sulfate and four glucuronides of oxidative/cleavage metabolites in minor/trace quantities (each < or = 5% of the sample). 7. In addition, the in vitro metabolism of cladribine was conducted using rat and human liver microsomal fractions in the presence of an beta-nicotinamide adenine dinucleotide phosphate generating system. Unchanged cladribine (> or = 90% of the sample) plus three minor metabolites, M1-3 (each < 8% of the sample), were profiled and tentatively identified by thin-layer chromatography and MS data. 8. Cladribine is not extensively metabolized in vitro and in vivo in all species. However, humans appear to metabolize cladribine to a greater extent than other animals. PMID- 15277019 TI - A feasibility study of interstitial hyperthermia plus external beam radiotherapy in glioblastoma multiforme using the Multi ELectrode Current Source (MECS) system. AB - PURPOSE: Thermoradiotherapy has been shown in several randomized trials to increase local control compared to radiotherapy alone. The first randomized study of interstitial hyperthermia in glioblastoma multiforme showed a survival benefit for hyperthermia, though small. Improvement of the heating technique could lead to improved results. The purpose of this feasibility study is to present the clinical and thermal data of application of an improved interstitial hyperthermia system. METHODS AND MATERIALS: Six patients with a glioblastoma multiforme were treated with interstitial hyperthermia using the Multi Electrode Current Source Interstitial Hyperthermia (MECS-IHT) system. The MECS-IHT system has the capability of spatial monitoring of temperature and individually steering of heating electrodes. Three sessions were given aiming at a steady state temperature of 42 degrees C for 1 h, with an interval of 3-4 days, during an external irradiation scheme of 60 Gy in 6 weeks. Hyperthermia was delivered with a mean of 10 catheters, 18 heating electrodes and 38 thermal probes per patient. RESULTS: Sub-optimal temperatures were encountered in the first two patients leading to adjustments in technique thereafter with subsequent improvement of thermal data. With a catheter spacing of 11-12 mm, measurements yielded a mean T(90), T(50) and T(10) of 39.9, 43.7 and 45.2 degrees C, respectively, over three sessions in the last patient. The power per electrode to reach this temperature distribution varied from 25-100% of full power in each of the last four patients. Thermal data were reproducible over the three sessions. Acute toxicity was minimal. CONCLUSIONS: Despite the spatial steering capabilities of the MECS-IHT system, a large temperature heterogeneity was encountered. The heterogeneity was the reason to limit the catheter spacing to 11-12 mm, thus making only small tumour volumes feasible for interstitial heating. PMID- 15277020 TI - Effect of 8-MHz radiofrequency-capacitive regional hyperthermia with strong superficial cooling for unresectable or recurrent colorectal cancer. AB - A well-known disadvantage of a radiofrequency-capacitive device for deep-seated tumours is preferential heating of the subcutaneous fat tissue. The authors previously developed the hyperthermia with their own external cooling unit and achieved strong superficial cooling, and reported its usefulness for the reduction of the preferential heating. The purpose of the present study was to evaluate the effect of hyperthermia with strong superficial cooling on the treatment results for unresectable or recurrent colorectal cancers. From 1986 to 2002, 44 patients with primary unresectable or locally recurrent colorectal cancer treated with thermoradiotherapy were analysed retrospectively. The patients with obesity as a subcutaneous fat thickness more than 3 cm, a high age or other serious complications did not undergo therapy. The results were compared between 17 cases with strong superficial cooling treated after 1997 (Group A) and 27 cases without strong superficial cooling treated before 1996 (Group B). Significant differences in thermometry data of T(max), T(ave) and T(min) were noted between Groups A (45.3, 44.4 and 43.6 degrees C, respectively) and B (42.9, 42.0 and 41.1 degrees C, respectively) (p<0.01). Complete response plus partial response rates were better for Group A than for Group B (59 versus 26%, p = 0.05). Multivariate analysis by logistic regression to evaluate the effects of certain factors on complete response plus partial response was strongly correlated with strong superficial cooling (p<0.05). The median survival times for overall survival were 24.3 months for Group A and 17.1 months for Group B (p<0.05). Eight-megahertz radiofrequency-capacitive regional hyperthermia with strong superficial cooling is potentially useful for improving treatment results in unresectable or recurrent colorectal cancers. PMID- 15277021 TI - Effect of calcitonin gene related peptide vs sodium nitroprusside to increase temperature in spontaneous canine tumours during local hyperthermia. AB - The objectives of this study were to compare the effects of two vasodilators, sodium nitroprusside (SNP) and calcitonin gene-related peptide (CGRP) on mean arterial pressure (MAP), heart rate (HR) and temperatures in tumour and surrounding normal tissue during local hyperthermia treatment. Eleven tumour bearing pet dogs with spontaneous soft tissue sarcomas were given SNP intravenously during local hyperthermia. The drug infusion rate was adjusted to maintain a 20% decrease in MAP. The median (95% CI) increase in the temperature distribution descriptors T(90) and T(50) was 0.2 degrees C (0.0-0.4 degrees C, p = 0.02) and 0.4 degrees C (0.1-0.7 degrees C, p = 0.02), respectively, in tumour. Normal subcutaneous tissue temperatures were mildly increased but remained below the threshold for thermal injury. The effects of CGRP were investigated in six tumour-bearing dogs following a protocol similar to that used for SNP. The median (interquartile (IQ) range) decrease in mean arterial pressure was 19% (15-26%) after CGRP administration and a significant increase was seen in tumour but not normal subcutaneous tissue temperatures. The median (95% CI) increase in the temperature distribution descriptors T(90) and T(50) was 0.5 degrees C (0.1-1.6 degrees C, p = 0.03) and 0.8 degrees C (0.1-1.6 degrees C, p = 0.13), respectively. Administration of SNP or CGRP did not result in local or systemic toxicity in tumour-bearing dogs. However, the magnitude of increase in tumour temperatures was not sufficient to improve the likelihood of increased response rates. Therefore, there is little justification for translation of this approach to human trials using conventional local hyperthermia. PMID- 15277022 TI - Hsp27 anti-sense oligonucleotides sensitize the microtubular cytoskeleton of Chinese hamster ovary cells grown at low pH to 42 degrees C-induced reorganization. AB - Chinese hamster ovary (CHO) cells maintained in vitro at pH 6.7 were used to model cells in the acidic environment of tumours. CHO cells grown at pH 6.7 develop thermotolerance during 42 degrees C heating at pH 6.7 and their cytoskeletal systems are resistant to 42 degrees C-induced perinuclear collapse. Hsp27 levels are elevated in cells grown at pH 6.7 and are further induced during 42 degrees C heating, while Hsp70 levels remain low or undetectable, suggesting that Hsp27 is responsible for some of the novel characteristics of these cells. An anti-sense oligonucleotide strategy was used to test the importance of Hsp27 by lowering heat-induced levels of the protein. The response of the microtubular cytoskeleton to heat was used as an endpoint to assess the effectiveness of the anti-sense strategy. Treatment with anti-sense oligonucleotides prevented the heat-induced increase of Hsp27 levels measured immediately following heat. Treatment with anti-sense oligonucleotides also sensitized the cytoskeleton of cells grown at low pH to heat-induced perinuclear collapse. However, cytoskeletal collapse was not evident in cells grown at pH 6.7 and treated with 4-nt mismatch oligonucleotides or in control cells maintained and heated at pH 6.7. The cytoskeleton collapsed around the nucleus in cells cultured and heated at pH 7.3. These results confirm that over-expression of Hsp27 confers heat protection to the microtubular cytoskeleton in CHO cells grown at low pH. PMID- 15277023 TI - Cytokine induction during exertional hyperthermia is abolished by core temperature clamping: neuroendocrine regulatory mechanisms. AB - The immunomodulatory effects of physiological temperature change remain poorly understood and inter-relationships between changes in core temperature, stress hormones and cytokines during exertional hyperthermia are not well established. This experimental study was designed to examine how cytokine (tumour necrosis factor (TNF)-alpha, interleukin (IL)-6, IL-12 and IL-1ra (receptor antagonist)) and hormone (epinephrine (Epi), norepinephrine (NE), growth hormone (GH) and cortisol (CORT)) responses are modified when the exercise-induced rise in core temperature is attenuated or exacerbated by immersion in a water bath. Ten men ((mean +/- SD) age: 26.9 +/- 5.7 years; height 1.75 +/- 0.07 m; body mass 76.0 +/ 10.9 kg; O(2 peak): 48.0 +/- 12.4 mL kg(-1) min(-1)) completed two 40-min cycle ergometer exercise trials at 65% O(2 peak) while immersed to mid-chest. Rectal temperature (T(re)) peaked at 39.1 +/- 0.03 and 37.5 +/- 0.13 degrees C during the hot (39 degrees C) and cold (18 degrees C) conditions, respectively. Blood samples were collected before, during (20- and 40-min) and after (30- and 120 min) exercise. Increases in circulating NE (>350%), Epi (>500%), GH (>900%), IL 12 (>150%) and TNF-alpha (>90%) were greatest after 40-min exercise in the heat. Substantial elevations of CORT (80%), IL-1ra (150%) and IL-6 (>400%) did not occur until after exercise was complete. Core temperature clamping decreased the rise in circulating stress hormone concentrations and abolished increases in plasma cytokine concentrations. These findings suggest that exercise-associated elevations of T(re) mediate increases of circulating stress hormones, which subsequently contribute to induction of circulating cytokine release. PMID- 15277024 TI - Changes in the dielectric properties of rat prostate ex vivo at 915 MHz during heating. AB - Changes in the tissue dielectric properties at 915 MHz of rat prostate tissue due to heating at temperatures in the range 45-75 degrees C were measured. The changes were found to be reversible with temperature and independent of the time of heating. The dielectric properties at 23 +/- 1 degrees C were measured as epsilon' = 62.8 +/- 2.7 and sigma = 1.17 +/- 0.07 S/m, while the linear temperature coefficients for reversible changes were 1.10 +/- 0.11%/ degree C for conductivity and -0.31 +/- 0.05% /degree C for relative permittivity. These properties and their temperature coefficients can be utilized in microwave treatment planning programmes to provide insight into the effects of dielectric changes that arise during microwave thermal therapy of prostate cancer. PMID- 15277025 TI - Development of a phantom compatible for MRI and hyperthermia using carrageenan gel--relationship between dielectric properties and NaCl concentration. AB - A phantom has previously been developed containing carrageenan, agarose and gadolinium chloride (called CAG phantom) for MRI with 1.5 T. T(1) and T(2) relaxation times of this phantom are independently changeable by varying concentrations of relaxation-time modifiers to simulate relaxation times of the various types of human tissues. The CAG phantom has a T(1) value of 202-1904 ms and a T(2) value of 38-423 ms, when the GdCl(3) concentration is varied from 0 140 micromol/kg and the agarose concentration is varied from 0-1.6%. A new phantom has now been developed (called CAGN phantom), made by adding an electric conductive agent, NaCl, to the CAG phantom for use in the areas of MRI and hyperthermia research. Dielectric properties of the CAGN phantom were measured and the results of experiments were expressed by the Cole-Cole equation in the frequency range of 5-130 MHz. The relationship between the conductivity of the CAGN phantom and the concentration of NaCl was expressed by a linear function in the frequency range of 1-130 MHz. The linear function involves a parameter of frequency and, when the frequency is 10 MHz, the conductivity of the CAGN phantom can be changed from 0.27-1.26 Sm(-1) by increasing the NaCl concentration from 0 0.7%. The CAGN phantom developed can be employed in basic experiments for non invasive temperature measurement using MRI and as a loading phantom for MRI with up to 3 T. PMID- 15277026 TI - Infrared thermographic SAR measurements of interstitial hyperthermia applicators: errors due to thermal conduction and convection. AB - Thermal conduction and convection were examined as sources of error in thermographically measured SAR patterns of an interstitial microwave hyperthermia applicator. Measurements were performed in a layered block of muscle-equivalent phantom material using an infrared thermographic technique with varying heating duration. There was a 52.7% reduction in maximum SAR and 75.5% increase in 50% iso-SAR contour area for a 60-s heating duration relative to a 10-s heating duration. A finite element model of heat transfer in an homogeneous medium was used to model conductive and convective heat transfer during the thermographic measurement. Thermal conduction artefacts were found to significantly distort thermographically measured SAR patterns. Convective cooling, which occurs when phantom layers are exposed for thermal image acquisition, was found to significantly affect the magnitude, but not the spatial distribution, of thermographically measured SAR patterns. Results from this investigation suggest that the thermal diffusion artefacts can be minimized if the duration of the applied power pulse is restricted to 10 s or less. PMID- 15277028 TI - Interferon-gamma expression in the inner ear of rats following secondary immune reaction in the endolymphatic sac. AB - Recently, we demonstrated increased intercellular adhesion molecule-1 (ICAM-1) expression in the inner ear of systemically pre-sensitized rats after antigen [keyhole limpet hemocyanin (KLH)] challenge into the endolymphatic sac (ES), in good correlation with the cellular infiltration. Interferon-gamma (IFN-gamma) is an important cytokine that upregulates the expression of ICAM-1. Here, we report upregulation of IFN-gamma expression in the inner ear of systemically pre sensitized rats after antigen (KLH) challenge into the ES. Immunoreactivity for IFN-gamma was detected in the spiral ligament, suprastrial region, spiral modiolar veins, spiral collecting venules, surface membrane of the perilymphatic compartment and perilymphatic space of immunized, but not control, rats. IFN gamma expression was detected at 1.5 h post-challenge, peaked at 6 h and gradually returned to baseline levels after 7 days. Interestingly, the time kinetics of IFN-gamma expression were in good correlation with those of ICAM-1. These observations demonstrate that antigen challenge into the ES induces IFN gamma expression, which can then upregulate ICAM-1 expression and induce cell infiltration, suggesting that IFN-gamma may play a crucial role in immune mediated inner ear diseases. PMID- 15277029 TI - Changes in aquaporin expression in the inner ear of the rat after i.p. injection of steroids. AB - The aquaporins (AQPs) are a family of small transmembrane water transporters. It has recently been revealed that they play a role in regulating homeostasis in the inner ear fluids. Steroid therapy is usually administered to patients with inner ear disorders; however, the mechanism of steroid effects has not been clearly determined. To elucidate the points of action of steroids in the inner ear, we recently examined the distributions of AQP isoform mRNAs in the rat inner ear and identified AQP1-6 mRNAs in the rat cochlea and AQP1, 3, 4, 5 and 6 mRNAs in the rat endolymphatic sac by means of reverse transcriptase polymerase chain reaction (PCR). In this study, we investigated changes in expression of AQP mRNAs in the rat inner ear after i.p. injections of steroids using real-time quantitative PCR and found that AQP3 mRNA in the endolymphatic sac was significantly upregulated in both dose- and time-dependent manners. This result suggests that steroids may effect water homeostasis in the rat inner ear via AQPs. PMID- 15277030 TI - Novel method for homogeneous gene transfer to the inner ear. AB - Viral vectors are widely used in gene therapy due to their efficiency. In this paper we describe a novel method for transfecting the whole inner ear of a guinea pig using adenoviral vectors. Very small perforations are made in both the cochlea and lateral semicircular canal, into which 50 microl of adenoviral suspension (8.9x10(8) plaque forming unit (PFU)/ml) is gently infused. Any excess suspension flows out through the perforation in the semicircular canal and therefore makes no contact with the central nervous system. Our method can therefore be utilized to perform homogeneous gene transfer and may eliminate any effects on other organs, such as the contralateral ear. PMID- 15277032 TI - Inactivating and non-inactivating potassium currents in isolated inner hair cells from guinea pig cochlea. AB - Using conventional whole-cell voltage-clamp recordings, we examined the 4 aminopyridine (4-AP)- and tetraethylammonium (TEA)-sensitive K(+) currents in the cochlear inner hair cells (IHCs) of guinea pigs. 4-AP-sensitive currents were activated slowly and sustained the same current level, whereas TEA-sensitive currents were activated rapidly, followed by inactivation. The inactivation time course of TEA-sensitive currents was voltage-dependent, becoming faster at more depolarized levels. The inactivation of TEA-sensitive currents almost recovered within 5 ms. 4-AP- and TEA-sensitive K(+) currents coexisted in the same IHC. PMID- 15277031 TI - Actions of subtype-specific purinergic ligands on rat spiral ganglion neurons. AB - In a previous study we showed that, in rat spiral ganglion neurons (SGNs), the adenosine 5'-triphosphate (ATP)-evoked currents were a combination of the activation of ionotropic receptors (the first fast current) and the activation of metabotropic receptors which secondarily opened non-selective cation channels. These two conductances imply the involvement of different receptor subtypes. In the present study, we tested three subtype-specific purinergic ligands: alpha,beta-methylene ATP (a;pha,beta-meATP) for P2X receptors, uridine 5' triphosphate (UTP) for P2Y receptors and 2'-3'-O-(4-benzoylbenzoyl) ATP (Bz-ATP) for P2Z (P2X(7)) receptors. Application of 100 microM alpha,beta-meATP did not trigger any significant change in membrane conductance, while the SGNs were responsive to ATP. Pressure application of UTP (100 microM, 1 s) evoked an inward current averaging 344+/-169 pA at a holding potential of -50 mV. The conductance developed after a latency averaging 1.5+/-0.6 s, took 4-6 s to peak and reversed slowly within 15-30 s. The current-voltage curve reversed near 0 mV, suggesting a non-selective cation conductance, like the second component of the ATP conductance. Bz-ATP evoked an inward current which developed without latency, was sustained during ligand application and was rapidly inactivated at the end of application: the same characteristics as the first component of the ATP-evoked current. The Bz-ATP conductance reversed around -10 mV, indicating also a non selective cation conductance. These results suggest that, in SGNs, ATP acts via two different receptor subtypes, ionotropic P2Z receptors and metabotropic P2Y receptors, and that these two receptor subtypes can assume different physiological roles. PMID- 15277033 TI - Effect of masker frequency on N1m amplitude in forward masking. AB - The effect of frequency on N1m has been investigated by various methods. However, it has not yet been measured using forward masking. In this study, the frequency specificity of N1m was investigated using forward masking. Although the masker frequency had some influence on N1m amplitudes, the results suggested that the frequency specificity of N1m was worse than that of a single-neuron or psychological tuning curve. This is probably because N1m includes various components, both frequency-specific and non-specific, some of which may be less affected by masking. Thus, our results agree with those of previous studies using intervening tones that suggested widespread neural representation in the auditory cortex. PMID- 15277034 TI - Presence and connections of auditory neurons in the rostrodorsal and rostrolateral parts of the thalamic reticular nucleus. AB - OBJECTIVE: Auditory neurons have been identified in the caudoventral part of the thalamic reticular nucleus (TRN). We examined the acoustic input to single cells in the rostrodorsal part of the TRN. MATERIAL AND METHODS: In alpha-chloralose anesthetized cats, we extracellularly recorded the responses of single neurons in the rostral TRN to acoustic and light stimuli. Next, to examine efferent projections of auditory neurons in the rostral TRN, we injected wheat germ agglutinin conjugated to horseradish peroxidase (WGA-HRP) into other thalamic nuclei where auditory neurons were detected, including the lateral posterior nucleus (LP), the lateral medial and suprageniculate nuclei and the centromedian nucleus. Finally, intracortical microstimulation of the LP was performed to demonstrate antidromic activation of the auditory neurons in the rostral TRN. RESULTS: In the rostral TRN, 2 types of response to auditory stimuli were observed: brief, short-latency bursts (13-20 ms; mean 16.5 ms) and longer bursts with a long latency (38.8-50 ms; mean 44.8 ms). Injection of WGA-HRP into the medial LP labeled cells only in the rostrodorsal TRN, while extending the injection to the other nuclei labeled cells in the rostrodorsal and rostrolateral parts of the nucleus. Auditory neurons in the rostral TRN were activated antidromically by microstimulation of auditory neurons in the LP, with a latency of 1.2 ms. CONCLUSIONS: These results strongly suggest that auditory neurons in the rostrodorsal TRN project to auditory neurons in the LP. The rostral auditory TRN may be involved in transmission of auditory information via the non-specific association system of the thalamus. PMID- 15277035 TI - Optical mapping of neural responses and their gamma-aminobutyric acid-ergic inhibitory effects in the auditory brainstem of early postnatal mice. AB - gamma-Aminobutyric acid (GABA)ergic neurons play important tropic and modulatory roles in the auditory pathway, especially in the early stage between postnatal Days 0 and 5. The effects of GABA and GABAa receptor antagonist were observed in this experimental study. Numerous histological and electrophysiological studies have been performed on the contribution of GABA to the auditory pathway; however, the spatio-temporal patterns of excitatory propagation and the relationships between GABA receptor and excitatory propagation have yet to be reported. Using an optical recording technique and a voltage-sensitive dye, the spatio-temporal patterns of excitatory propagation were observed in the auditory brainstem slices of early postnatal mice. A bath containing 50 microM GABA was applied, which largely inhibited the excitatory activities along the vestibulocochlear pathway. Bicuculline methiodide (BMI), a competitive antagonist against GABAa receptor, partially reversed the effects of GABA on the optical signals. Bath application of BMI alone helped to facilitate the depolarization course and its effect was apparent as an enlargement of the depolarized region from the cochlear nucleus and vestibular nucleus to some adjacent brainstem nuclei, as well as enhancing the amplitude of changes in the optical signals. The experimental results seem to suggest that GABAa receptors are widely distributed in an early postnatal auditory brainstem. GABA exhibited a greater modulating effect in the adjacent brainstem nuclei, which are involved in complex information processes, than that observed in the modulating primary auditory pathway. In the present experiment, significant GABAergic contributions to the optical recordings in the auditory brainstem were observed. PMID- 15277036 TI - Vestibular function and vasopressin. AB - OBJECTIVE: To investigate the relation between the vestibular system and vasopressin (AVP). MATERIAL AND METHODS: We examined the effects of electrical and caloric vestibular stimulation on plasma AVP levels in anesthetized rats. Plasma AVP levels of patients with intractable Meniere's disease who were subjected to endolymphatic drainage and steroid instillation surgery (EDSS) or intratympanic gentamicin (GM) injection were measured before and after these interventions. RESULTS: Electrical vestibular stimulation increased plasma AVP levels in a current intensity-dependent manner. Plasma AVP levels were also increased by caloric stimulation with cold water. Plasma AVP levels decreased rapidly after EDSS, and were maintained at a low level even 6-12 months following EDSS or intratympanic GM injection. CONCLUSIONS: Vestibular activation or inhibition-induced imbalance of intervestibular activities increased plasma AVP levels in rats. Therefore, vestibular disorder would seem to increase plasma AVP and thus worsen endolymphatic hydrops. EDSS rapidly decreased plasma AVP and would seem to reduce hydrops. Inhibition of vertigo spells by EDSS or intratympanic GM injection would reduce a possible stress response, resulting in a decrease in plasma AVP levels a long time after the treatments. This resultant decrease in AVP would beneficially inhibit the formation and/or maintenance of hydrops and thus prevent vertigo spells. PMID- 15277037 TI - Acoustic stimulation promotes the expression of inducible nitric oxide synthase in the vestibule of guinea pigs. AB - Loud acoustic stimulation is known to cause inner ear disturbance. We examined immunohistochemically the vestibule of 12 guinea pigs after acoustic stimulation. The animals were divided into two equal groups: a control group and an acoustic stimulation group. The temporal bones were fixed by means of a cardiac infusion of fixative and immunohistochemically stained for inducible nitric oxide synthase (iNOS). The temporal bones in the control group did not show any iNOS. In the acoustic stimulation group, immunoreactivity for iNOS was detected in the supporting cells and sensory cells of the sensory epithelium, in the dark cell areas and in the vestibular ganglion cells. These findings suggest that free radicals are involved in the pathogenesis of noise-induced inner ear damage. Furthermore, free radicals may cause vestibular damage, as is seen in noise induced inner ear damage. PMID- 15277038 TI - Effect of a glutamate blocker, ipenoxazone hydrochloride on the hypoxia-induced firing in the medial vestibular nucleus. AB - To elucidate the effectiveness of the drug in the treatment of vertebrobasilar insufficiency (VBI), we performed an electrophysiological study to examine the effects of ipenoxazone hydrochloride, a glutamate blocker, on hypoxia-induced firing in the medial vestibular nucleus (MVN) neuron, using alpha-chloralose anesthetized cats. The single neuronal activity of the MVN was recorded extracellularly with a glass-insulated silver wire microelectrode attached along a seven-barrel micropipette. The firing rate of MVN neurons showed a transient increase [hypoxic depolarization (HD)] during 5% O(2) inhalation, followed by a gradual decrease and disappearance. HD and the time to disappearance of firing induced by hypoxia were inhibited by iontophoretic application of ipenoxazone hydrochloride. These results suggest that ipenoxazone hydrochloride protects against hypoxic neuronal dysfunction, and may be an effective drug for vertigo caused by VBI. PMID- 15277039 TI - The terminal of the sympathetic nerve fibers in the facial nerve. AB - OBJECTIVE: The terminal of the sympathetic nerve fibers of the rat facial nerve in the temporal bone region was investigated. MATERIAL AND METHODS: We used tyrosine hydroxylase (TH) and the synaptophysin antibody as markers of the sympathetic nerve fiber and the membrane of the synaptic vesicle, respectively. Using immunohistochemistry, we determined whether and where the synapse exists in the facial nerve of the Sprague-Dawley rat. RESULTS: TH-immunoreactive fibers were confirmed as being present in both the epineurium and the nerve fascicle of the facial nerve. A synaptophysin immunoreaction was found in the facial nerve in a region of the temporal bone. These reaction products looked like varicosities. Most TH-positive fibers in the facial nerve disappeared after superior cervical ganglionectomy. CONCLUSIONS: As the synaptophysin immunoreaction indicates the existence of a synapse, we speculate that the sympathetic fibers affect the facial nerve in the region of the temporal bone. Further studies may be needed to elucidate the function of the sympathetic system in the facial nerve. PMID- 15277040 TI - Potential oscillation elicited by i.v. olfaction and its applicability as an objective clinical olfaction test. AB - OBJECTIVE: Alinamin has long been applied in Japan for testing i.v. olfaction and to diagnose olfactory disorders. The test is subjective, each subject being asked about the presence or absence of olfaction. The credibility of the answers is highly questionable in some cases; as a result, the reliability of the test is poor. Recent studies demonstrated an induced electric potential in the scalp during i.v. olfactory testing. Some patients complain of the pain of the injection during i.v. olfactory testing; therefore, the effect of this pain must be considered with respect to measurement of the i.v. olfaction-elicited potential (IVOP). MATERIAL AND METHODS: This investigation involved 179 subjects with various olfaction levels. Each subject received an Alinamin injection; the elicited potential amplitude was compared before and after the injection and the increasing ratio (IR) was computed. Gender, age, level of olfactory disorder, the presence or absence of olfaction and the presence or absence of the pain of injection were considered as factors affecting IR. RESULTS: IR showed significant increases in groups characterized by the presence of olfaction as well as in groups reporting pain of injection. The test subjects were further divided into four groups based on their olfaction and pain of injection patterns as follows: Group A, no smell and no pain; Group B, smell and no pain; Group C, no smell and pain; and Group D, smell and pain. Subjects exhibiting no recognizable olfaction or pain of injection (Group A) revealed no increase in IVOP following injection. Subjects with either recognizable olfaction or pain of injection (Groups B and C) exhibited a slight increase in IVOP following injection. Subjects with both noticeable olfaction and pain of injection (Group D) demonstrated a significant increase in IVOP following the injection with a very high value of IR (>2). Furthermore, there were significant differences between the four groups in terms of IR level, with the exception of Groups B and C. CONCLUSIONS: Olfaction is largely involved with the generation of IVOP. However, pain resulting from injection of Alinamin is considered to be a significant factor. IVOP showed significant effectiveness for diagnosing olfactory disorders in cases who did not experience pain of injection. PMID- 15277041 TI - Suitability of the odor stick identification test for the Japanese in patients suffering from olfactory disturbance. AB - We studied the suitability of the Odor Stick Identification Test for the Japanese (OSIT-J) in patients suffering from olfactory disturbance. In 120 patients with olfactory disturbance (age range 12-85 years) there were statistically significant correlations between the odor identification rate on the OSIT-J, the results of the Japanese standardized olfactory test (T&T olfactometry) and subjective symptom scores. In every patient treated for olfactory disturbance, the OSIT-J reflected the grade of recovery from the olfactory disturbance as determined by means of T&T olfactometry. The odor identification rate on the OSIT J also correlated significantly with the results of the i.v. Alinamin test. Regarding the rate of correct recognition of odors on the OSIT-J, menthol and curry odors were recognized with a high rate and orange and wood odors with a low rate. Although the OSIT-J includes 13 kinds of odorants, the number of odorants used can be reduced to a minimum of 5 as the results obtained with this reduced form of the OSIT-J also correlated with the results of T&T olfactometry and the subjective symptom scores as well as with the results obtained with the 13 odorant OSIT-J. We conclude that the OSIT-J is suitable not only as a screening test for olfactory disturbance but also for practical use in clinical otorhinolaryngology. PMID- 15277042 TI - Olfactory impairment and Parkinson's disease-like symptoms observed in the common marmoset following administration of 1-methyl-4-phenyl-1,2,3,6 tetrahydropyridine. AB - OBJECTIVE: Although olfactory disturbance appears to occur in the early stages of Parkinson's disease (PD) in humans, little is known about its mechanism. The aim of this study was to make a PD model using injection of 1-methyl-4-phenyl-1,2,3,6 tetrahydropyridine (MPTP) in the common marmoset and to discover the mechanism of olfactory disturbance in this animal. MATERIAL AND METHODS: Olfactory disturbance induced by MPTP in the common marmoset was observed by behavioral, biochemical and immunohistochemical means. RESULTS: Administration of MPTP caused common marmosets to enter an akinetic state within a few days and to show signs of impaired olfactory function. Biochemical study showed a decrease in dopamine levels, especially in tissue samples from the caudate nucleus and putamen. Immunohistochemical analysis revealed a lack of tyrosine hydroxylase immunoreactivity, not only in the substantia nigra, caudate nucleus and putamen but also in the olfactory tubercle. CONCLUSION: These results demonstrate that MPTP causes both PD-like symptoms and olfactory disturbance in the common marmoset. The olfactory disturbance observed in these animals may be due to the lack of dopamine in the olfactory tubercle. PMID- 15277043 TI - Expression of neurokinin a receptor mRNA in human nasal mucosa. AB - In airway tissues, it has been suggested that tachykinins act as the transmitter for afferent sensory nerves which respond to various irritants and may be involved in airway allergic reactions. Three classes of tachykinin receptor have been recognized, denoted NK1, NK2 and NK3, which exhibit preferential affinity for substance P, neurokinin A and neurokinin B, respectively. We used molecular probes to study the gene expression and distribution of NK2 receptor in human nasal mucosa. Total RNA was isolated from human nasal mucosa and NK2 receptor mRNA was detected in these tissues using reverse transcriptase polymerase chain reaction (RT-PCR). For an in situ hybridization study of human nasal mucosa, we utilized the PCR directly to incorporate a T7 RNA polymerase promoter sequence onto the NK2 receptor cDNA, and these PCR products were used as the DNA template for producing digoxigenin-labeled antisense and sense RNA probes. These studies revealed that NK2 receptor mRNA was expressed in blood vessels. The results suggest a primary role for neurokinin A in the form of vascular responses in the upper respiratory tract. PMID- 15277044 TI - Quantitative analysis of expression of NeuroD, GAP43 and receptor tyrosine kinase B in developing mouse olfactory neuroepithelium. AB - OBJECTIVE: The mammalian olfactory neuroepithelium (OE), which harbors olfactory receptor neurons (ORNs), has the unusual characteristic of continuous neurogenesis throughout its lifetime. This unique feature provides an excellent model for neuronal differentiation. Recently, we found dual-phase expression of NeuroD, a member of the basic helix-loop-helix transcription factor family, in the developing mouse OE, suggesting multiple roles of NeuroD during the development of mammalian ORNs. MATERIAL AND METHODS: In order to better understand the molecular mechanism of the development of ORNs, we performed quantitative analysis of expression of NeuroD, GAP43 and receptor tyrosine kinase B (TrkB), as well as 5-bromo-2'-deoxyuridine (BrdU)-labeled cells, in the developing mouse OE from gestational Day 10 to postnatal Day 28. RESULTS: During the embryonic period, NeuroD expression is mostly confined to the basal compartment. During the neonatal period, NeuroD expression is detected in two compartments: the middle compartment and the basal compartment. GAP43-expressing cells were located between these two NeuroD-positive layers. TrkB-expressing cells were located above the NeuroD-positive layer in the middle compartment. As the mice grew, the numbers of NeuroD-expressing cells and BrdU-labeled cells in the basal compartment significantly decreased, while the number of NeuroD expressing cells in the middle compartment gradually increased. The number of TrkB-expressing cells dramatically increased. The number of GAP43-expressing cells also gradually increased. However, the relative proportion of GAP43 cells decreased as the OE developed. CONCLUSION: NeuroD is a useful molecular marker for studying olfactory neurogenesis. PMID- 15277045 TI - Near-infrared spectroscopy of the adult human olfactory cortex. AB - OBJECTIVE: Near-infrared spectroscopy (NIRS) is a non-invasive method for investigating activation of the human cortex. The applicability of NIRS to the olfactory cortex was investigated. MATERIAL AND METHODS: The relative oxy- and deoxy-hemoglobin levels of the orbito-frontal cortex during olfactory stimulation in healthy subjects were measured using NIRS. RESULTS: When perfumed strips containing the odorants beta-phenyl ethyl alcohol, iso-valeric acid and gamma undecalactone were presented, the oxy-hemoglobin level increased but the deoxy hemoglobin level did not change. The increase in the oxy-hemoglobin level was observed bilaterally. A placebo perfumed strip did not elicit a change in the hemoglobin level. It was also observed that the odorant intensity affected the oxy-hemoglobin level. Although the orbito-frontal cortices seemed to be activated bilaterally during olfaction, the right cortex was activated to a greater extent than the left. CONCLUSION: NIRS appears to be an adequate method for investigating the human olfactory cortex. PMID- 15277046 TI - Immunohistochemical localization of eotaxin immunoreactivity in nasal polyps. AB - Eotaxin is a C-C chemokine that acts to selectively induce local accumulation of eosinophils and basophils. Eotaxin is also believed to be involved in the infiltration of eosinophils in the nasal polyps of patients with chronic sinusitis. However, only a few studies on eotaxin in nasal polyps have been performed. In this study, we investigated the localization of eotaxin in human nasal polyps and the identification of eotaxin-positive cells using immunohistochemistry. The distribution of eotaxin immunoreactivity in the nasal polyps of patients with chronic sinusitis was found to almost coincide with the presence of eosinophils. Eotaxin immunoreactivity was also detected in some vascular endothelial cells. These findings suggest that eotaxin is produced by eosinophils and vascular endothelial cells in nasal polyps and is involved in the accumulation of eosinophils in nasal polyps. PMID- 15277047 TI - Bone-constructing cells from ethmoid bone may have multilineage differentiation potential: preliminary report. AB - OBJECTIVE: In order to investigate multilineage differentiation in human cultured cells from ethmoid bone, we conducted a morphological study to examine adipogenic and chondrogenic differentiation. MATERIAL AND METHODS: After reaching confluence, cells underwent terminal adipogenic differentiation by treatment with 100 microM indomethacin, 0.5 mM 1-methyl-3-isobutylxanthine, 1 microM dexamethasone (DEX), 10 microg/ml insulin and 0.3% dimethylsulfoxide in a medium supplemented with 10% fetal bovine serum. Chondrogenic differentiation was attempted by centrifuging a pelleted micromass using transforming growth factor beta3 (TGF-beta3), DEX, ascorbic acid (AA), pyruvate acid, proline, glucose and (ITS)-plus. RESULTS: The cultured cells displayed adipocyte but not chondrogenic lineage under these conditions. Considering the possibility that some differentiation potential may be lost with in vitro culture but maintained using another chondrogenic differentiation medium containing TGF-beta1, it is possible that cultured cells may have multilineage potential, including chondrogenic differentiation ability. CONCLUSIONS: These morphological abilities of human cultured cells may indicate the possibility of the existence of mesenchymal stem cells in sinus bone. If mesenchymal stem cells exist in ethmoid bone, they may play an important role in future research on the regulation mechanisms of human bone tissue. PMID- 15277048 TI - Clinical study of flavor disturbance. AB - OBJECTIVE: We have observed that, in cases of smell loss, patients often complain of taste loss as well even though they actually have normal gustatory acuity according to gustatory tests; we have defined such symptoms as "flavor disturbance". The clinical features of flavor disturbance are reported in this paper. MATERIAL AND METHODS: A total of 297 patients (99 males, 198 females; mean age 55.5 years) were treated for olfactory disturbance at the hospital of Hyogo College of Medicine between July 1995 and August 2001. Sixty-six out of 297 patients (22.5%) also experienced taste disturbance, and 49 of these 66 cases were evaluated by means of smell and taste tests. These 49 patients who complained of taste and smell loss were classified into two groups according to the results of their smell and taste tests. Patients who only complained of olfactory disturbance were also reviewed. RESULTS: There was no relationship between the severity of olfactory disturbance and the degree of flavor disturbance. The incidence of flavor disturbance was high in patients with sudden olfactory disturbance after upper respiratory tract infection or head trauma and low in those with slowly progressive olfactory disturbance. The symptoms of flavor disturbance improved regardless of whether smell was improved or not. CONCLUSIONS: The patients with flavor disturbance tended to misrecognize that they had taste loss because of sudden smell loss, and there were more of these cases than we expected. When patients with smell and taste loss are treated, flavor disturbance should also be considered. PMID- 15277049 TI - Taste function in elderly patients with unilateral middle ear disease. AB - OBJECTIVE: It is generally said that taste function deteriorates naturally with age. Taste function after middle ear surgery in elderly patients has not been clarified. MATERIAL AND METHODS: The taste function of the chorda tympani nerve on the normal and diseased sides was examined before and after middle ear surgery using electrogustometry in 79 patients aged>60 years, and the findings were compared with those in 228 young and middle-aged patients. RESULTS: The threshold of electrogustometry on the normal side increased significantly with increasing age (p<0.0001). The rate of thresholds that were off the scale was highest in the>70 years age group. The preoperative threshold on the diseased side increased significantly with increasing age in patients with chronic otitis media (p=0.0029) and cholesteatoma (p<0.0001). In patients with chronic otitis media, the postoperative threshold of the>60 years age group tended to be higher than that of the<60 years age group. CONCLUSIONS: These findings suggest that the taste function of the chorda tympani nerve deteriorated on the diseased side as much as on the normal side in elderly patients. Therefore, in most cases, we do not have to pay as much attention to the chorda tymapani nerve when performing surgery in elderly patients compared to young and middle-aged patients. PMID- 15277050 TI - Abnormalities of the electrogastrogram in globus pharyngeus. AB - There appear to be multiple indicators for the presence of globus pharyngeus. We have assumed that patients with abnormal gastric motility can be included among those suffering from globus pharyngeus. Gastric motility can be examined non invasively using the electrogastrogram (EGG). In this study, 32 patients (22 females, 10 males; mean age 55+/-13 years) with symptoms of globus pharyngeus consented to EGG recording. The EGGs of asymptomatic healthy adults (6 females, 10 males; mean age 28+/-5 years) were obtained as controls. No significant statistical difference was found in any of the measurement parameters between the two groups. However, we found that 9.4% of patients with globus pharyngeus exhibited abnormal gastric myoelectrical activity as measured by the EGG. This suggests that a small proportion of patients who complain of globus pharyngeus have abnormal gastric motility. PMID- 15277051 TI - Clinical and ultracytochemical investigation of sialadenosis. AB - OBJECTIVE: The records of 65 patients with sialadenosis treated between 1989 and 2002 at Tokyo Women's Medical University Hospital were studied clinically and ultracytochemically, focusing on the secretory granules of the acinar cells. MATERIAL AND METHODS: Biopsies of parotid glands were performed in 18 patients. The obtained parotid tissues were investigated electron microscopically and ultracytochemically. For the ultracytochemistry of carbohydrates, the periodic acid-methenamine silver method (PAM) and lectin-gold technology were used. An immunohistochemical study using anti-alpha-amylase antibody was also undertaken. RESULTS: Light microscopically, the acinar cells of the parotid glands of sialadenosis patients were pale and swollen. Ultrastructurally, most secretory granules of the acinar cells were homogeneous and could be classified into two types: electron-lucent and electron-dense. The former type was seen mainly in patients with anorexia nervosa and bulimia, while the latter type was predominantly found in patients with hypertension or diabetes mellitus. Both types of granule reacted uniformly with PAM. Acinar cells from normal parotid glands contained spherical dense cores which showed little reaction. The peripheral rims of the granules were strongly stained. Lectin-gold binding in the granules showed a similar pattern to PAM staining. Immunohistochemically, the acinar cells of the sialadenosis patients reacted with anti-alpha-amylase antibody. The dark granular type showed more intense reaction than the light granular type. CONCLUSION: These findings suggest that amylase and glycoconjugates are present in the secretory granules in sialadenosis although the granule structure is abnormal, being enlarged by disturbance of normal protein synthesis and release at the cellular level and by dense-core formation in the granules. PMID- 15277054 TI - Analysis of catechin content of commercial green tea products. AB - Tea (Camellia sinensis) contains numerous polyphenolic flavonoid-derived compounds known as catechins, which have shown interesting protective activity against cancer and cardiovascular disease. Numerous products based on tea are commercially available, many of which claim to contain specific amounts of the bioactive catechins. The catechin content of seven commercial green tea products (encapsulated extracts or tea bags) was quantified by HPLC and, where possible, compared to that claimed on the label. Wide variability was observed in the catechin content between green tea products, even those that appear outwardly similar to consumers. Measured catechin content ranged from 9% to 48% of label claims; all values were significantly lower than those claims (P < 0.05). These results continue to demonstrate the problems that exist with quality control in the dietary supplement and herbal medicine industry and, there, for consumers of nutraceuticals. PMID- 15277055 TI - In vivo effects of black cohosh and genistein on estrogenic activity and lipid peroxidation in Japanese Medaka (Oryzias latipes). AB - This study was designed to assay the estrogenic activities and the antioxidant potential of ethanol extracts from the herbal dietary supplement black cohosh (Cimicifuga racemosa) relative to the natural phytoestrogen genistein. The in vivo mechanisms of action of these two natural products have not been completely elucidated, and Japanese medaka (Oryzias latipes) provides a useful organism for initial in vivo screening of natural products. While both genistein and estradiol altered ovarian and testicular steroid release and decreased circulating testosterone levels in males, neither black cohosh total extract (75-30,000 ng/fish), cimiracemoside A, 25-O-methyl-cimigenoside, actein, nor 26-deoxy-actein caused any differences in estrogenic activity compared to control fish. To assess antioxidant potential, animals were treated with natural products then challenged with 2-acetylaminofluorene (2-AAF) to induce lipid peroxidation (LPO) in the liver. Neither the total ethanol extracts from black cohosh nor its individual components showed an inhibitory effect in 2-AAF induced LPO. However, genistein manifested potent antioxidative activity in the LPO assay, with similar potency to a high dose of a-tocopherol. In contrast to genistein, black cohosh did not exhibit traditional estrogenic effects nor significant in vivo anti-oxidant potential in this fish model system. PMID- 15277059 TI - Marshmallow (Althaea officinalis L.) monograph. PMID- 15277057 TI - The natural product education exchange. PMID- 15277060 TI - Good herb, bad herb... PMID- 15277061 TI - Investigation of quality in ephedrine-containing dietary supplements. AB - Ephedra alkaloids in 47 dietary supplements were measured to examine variability within and between products as well as for comparison of actual constituents with label claims of manufactured products. Samples were analyzed for (-)-ephedrine, (+)-pseudoephedrine, (-)-methylephedrine, (+)-methylpseudoephedrine,(-) norephedrine, and (+)-norpseudoephedrine without derivatization using gas chromatographic mass spectrometry (GC/MS). Samples were then screened for pharmaceutically derived chiral contaminants. The resulting data demonstrated that label claims matched total ephedra alkaloid content to within about 25% for most of the supplements, and no products purporting to be ephedrine-free contained any ephedra alkaloids. Furthermore, chiral analysis of these dietary supplements revealed only naturally occurring ephedra alkaloids. (-)-Ephedrine and (+)-pseudoephedrine accounted for > 95% of the alkaloid content in each of the ephedra-containing products. Examination of identical supplements originating from different lots revealed consistency in total alkaloid content between lots but large variations in specific individual alkaloid content. PMID- 15277062 TI - Hawthorn evokes a potent anti-hyperglycemic capacity in streptozotocin-induced diabetic rats. AB - The hypoglycaemic effect of an aqueous extract of hawthorn leaves (Crataegus oxyacantha) was investigated in normal and streptozotocin (STZ) diabetic rats. After a single dose or 9 daily doses, oral administration of the aqueous extract produced a significant and dose-dependent decrease on blood glucose levels in STZ diabetic rats (P < 0.001), but had no effect on blood glucose levels in normal rats. No changes were observed in basal plasma insulin concentrations after treatment in normal or STZ diabetic rats. In addition, the acute toxicity study of the extract was investigated in mice. The results obtained showed that the aqueous extract had a high LD50 value (13.5 g/kg) in mice. We conclude that an aqueous extract of hawthorn leaves exhibits a potent anti-hyperglycemic activity in STZ rats, but not in normal rats, without affecting basal plasma insulin concentrations. PMID- 15277063 TI - Evaluation of the biological activities of crude extracts from patagonian prosopis seeds and some of their active principles. AB - Extracts of different polarities from three species and three varieties of the genera Prosopis: P. alpataco, P. denudans var. denudans, P. denudans var. patagonica, and P. denudans var. stenocarpa, were screened in order to evaluate their antibacterial, antifungal, antifeedant, antihelminthic, molluscicidal and toxic activities. The extractions of the plant materials were carried out successively with petroleum ether, dichloromethane, ethyl acetate, methanol and water. All petroleum ether extracts showed antibacterial activity. The dichloromethane extract of P. alpataco showed antibacterial and antifungal activities. Methanol and aqueous extracts of P. denudans var. denudans and P. denudans var. patagonica showed antifungal activities and a slight response to the toxicity test. Fatty acids and a group of pentacyclic triterpenes were identified as responsible for antibacterial activities in some of the active extracts. PMID- 15277064 TI - University of Pittsburgh. PMID- 15277065 TI - Butterbur for allergic rhinitis. PMID- 15277068 TI - Alfalfa (Medicago sativa L.):a clinical decision support tool. PMID- 15277072 TI - A randomized, controlled study of Kan Jang versus amantadine in the treatment of influenza in Volgograd. AB - Two randomized, parallel-group clinical studies with a verum and a control group were performed to investigate the effect of a standardized extract (SHA-10) of Andrographis panaiculata (N.) fixed combination Kan Jang in the treatment of diagnosed influenza viral infection. The pilot study was performed on 540 patients with 71 Kan Jang-treated patients with the second phase conducted enrolling 66 patients. The differences in the duration of sick leave and frequency of post-influenza complications indicate that the Kan Jang phytopreparation not only contributes to quicker recovery, but also reduces the risk of post-influenza complications. Kan Jang was well tolerated by patients. PMID- 15277073 TI - Anti-amyloidogenic effect of Allium sativum in Alzheimer's transgenic model Tg2576. AB - High levels of cholesterol are implicated in potentiating Alzheimer's disease (AD). Therefore, the use of cholesterol-lowering agents such as statins has attracted considerable interest in treating AD. However, statins stimulate inflammatory response, which may aggravate AD-pathology. Although garlic (Allium sativum) is historically known for its hypocholesterolemic effects in relation to cardiovascular functions, no reports indicate its use in treating AD. Current study tested the feasibility of using dietary garlic on the reduction of amyloid burden in a transgenic mouse model of AD that overexpresses the human amyloid precursor protein 695 carrying Swedish double mutation (K670N/M671L) (Tg2576). Animals were treated with aged garlic extract (40 mg/kg/d/4 wks). Cerebral levels of sAPPalpha, sAbeta40, sAbeta42 were analyzed by sandwich ELISA. Results show 64% reduction of sAPPalpha, and approximately 21-fold elevation of Abeta40 and Abeta42 in untreated Tgs compared to wild type and littermate controls. Dietary garlic increased sAPPalpha by 25% and decreased Abeta40 and Abeta42 by 31% and 32%, respectively, compared to untreated Tgs. These results suggest a simple and non-invasive dietary therapy for reducing risk of AD in probable cases and reducing preexisting amyloid burden in clinically diagnosed AD cases. PMID- 15277075 TI - "Indian" echinacea. PMID- 15277074 TI - Curricula development. PMID- 15277076 TI - Chaparral monograph:a clinical decision support tool. AB - An evidence-based systematic review including scientific evidence, expert opinion, folkloric precedent, history, pharmacology, kinetics/ dynamics, interactions, adverse effects, toxicology, and dosing. PMID- 15277078 TI - Migraine therapy:seamlessness between plant-derived and synthetic drugs. AB - Migraine has been endemic for many centuries. Seamlessness between plant-derived and synthetic migraine therapies is evident in three major aspects: (A) The latter are molecular adaptations of the former, (B) Combinations of herbal and non-herbal migraine therapies (e.g., one for prophylaxis and the other for acute therapy) are used by many patients, and (C) All migraine treatments, whether plant-derived or not, are capable of clinical trial using the same endpoints. At least two, newly discovered plant-derived migraine therapies are currently under research using standard clinical trial methodology. Suggestions are provided for more flexible clinical trial designs that might particularly suit research into herbal remedies. The future of migraine therapy is likely to continue to exhibit an intimate, symbiotic relationship between plant-derived and synthetic drugs. PMID- 15277079 TI - Hypoglycemic effect of aqueous extract of Ammi visnaga in normal and streptozotocin-induced diabetic rats. AB - The effect of the aqueous extract of Ammi visnaga (Apiaceae) on blood glucose levels was investigated in fasting normal and streptozotocin-induced diabetic rats after single and repeated oral administration. The aqueous extract of Ammi visnaga (AV) at a dose of 20 mg/kg significantly reduced blood glucose in normal rats six hours after a single oral administration (P < 0.005) and nine days after repeated oral administration (P < 0.05). This hypoglycemic effect is more pronounced in streptozotocin (STZ) diabetic rats (P < 0.001). Acute toxicity (LD50) and general behavioural effects of an aqueous extract of AV fruits was studied in mice. The LD50 of intraperitoneal (i.p.) and oral administration was 3.6 and 10.1 g/kg, respectively. These findings suggest that the aqueous extract of AV possess significant hypoglycemic effect in both normal and STZ diabetic rats and support, therefore, its claimed clinical use by the Moroccan population. PMID- 15277080 TI - A study of Russian phytomedicine and commonly used herbal remedies. AB - Immigrants into this country often bring their own cultural beliefs and medical system with them. Since many parts of the world use phytomedicine as the sole source of pharmaceuticals, this means that their traditional form of therapy includes the use of herbal products. It has been observed that many immigrants from the former Soviet Union are used to using phytomedicine and actively seek out familiar herbs when they come to the United States (U.S.). This paper provides a description of the most commonly used Russian phytomedicinals with an explanation of indications for use and Latin and English translations of each plant's name. This makes available insights into the use of herbals in the former Soviet Union and aids in bridging the cultural and language gap between herbal medicine in the U.S. and in Russia. PMID- 15277081 TI - Evaluation of a mucoactive herbal drug, Radix Ophiopogonis, in a pathogenic quail model. AB - We investigated the effect of Radix Ophiopogonis on airway mucociliary clearance and mucus secretion in anesthetized quails. The oral administration of 10 g/kg of Radix Ophiopogonis significantly increased tracheal mucociliary transport velocity (MTV). Moreover, either 10 g/kg or 3 g/kg of Radix Ophiopogonis markedly attenuated the human neutrophil elastase (HNE)-induced decrease in MTV. Furthermore, we found that 10 g/kg of Radix Ophiopogonis significantly abolished the HNE-induced increases in fucose and protein contents of tracheal lavage, whereas Radix Ophiopogonis at the same dose only significantly decreased the protein content in the control group. These results suggest that Radix Ophiopogonis improves airway mucociliary clearance and that the improvement may, at least in part, be ascribed to the amelioration of airway mucus secretion. PMID- 15277083 TI - Kava monograph: a clinical decision support tool. AB - The proposed uses, dosing parameters, adverse effects, toxicology, interactions and mechanism of action of kava is systematically reviewed in monograph format. PMID- 15277085 TI - Characterizing adverse events reported to the California Poison Control System on herbal remedies and dietary supplements: a pilot study. AB - OBJECTIVE: The purpose of this study was to (1) characterize the population who uses and reports adverse events to the California Poison Control System (CPCS) related to herbal remedies (HR) and dietary supplements (DS), (2) to assess whether adverse drug reactions (ADRs) occur within the usual recommended dosing, and (3) to describe the nature of adverse events. METHODS: A retrospective search of HR and DS related calls was conducted between January 1997 and June 1998. Data collection included the demographics of callers, types of exposures, substances involved, amounts ingested and severity. RESULTS: Of the 918 HR and DS calls received, 259 (28.2%) were drug information queries, 599 (65.3%) were exposures, and 60 (6.5%) were excluded. Exposures occurred most often in pediatric patients 324 (54.1%), but exposures resulting in ADRs occurred most often in adults at recommended doses. There were 233 ADRs, of which 67 (28.8%) occurred in children. The most common products involved in ADRs were zinc (38.2%), echinacea (7.7%), chromium picolinate (6.4%) and witch hazel (6%). Severity of ADRs required no treatment or was minor in a majority of cases, but may have contributed to hospitalization in three. CONCLUSION: Children were involved in a majority of exposure calls to the CPCS involving HR and DS. Adults, however, were more likely to develop an ADR from these products due to intentional exposure. Overall, the ADRs caused by HR and DS were mild, could be managed at home and rarely resulted in a severe outcome or other sequelae. Products involved in ADRs were consistent with the most common HR and DS products used by adult U.S. consumers. PMID- 15277086 TI - Neem in human and plant disease therapy. AB - As a therapeutic agent, neem is one of the most popular trees in traditional medicinal systems and is increasingly becoming important in herbal alternative therapy. The tree itself is considered a "village pharmacy" because of the well established fact that every part of the tree has an application in curing human diseases. The tree has been a constant source of novel and structurally unique phytochemicals that can constitute the basis for the development of novel pharmaco-therapeutic agents against various human diseases. Being a prototype for the development of safer drugs and ecofriendly, pro-human health agrochemical agents against a vast variety of plant diseases, the tree always remains in the center of safe herbal drug and pesticide development in the service of mankind. PMID- 15277087 TI - Fatty acid content of selected seed oils. AB - Fatty acid content of selected seed oils from world-wide edible fruits, Ceratonia ciliqua (carob) from Caesalpiniaceae family, Diospyros kaki (persimmon) from Ebenaceae family, Zizyphus jujuba (jujube) from Rhamnaceae family, and Persea gratissima (avocado pear) from Lauraceae family, were determined by capillary gas chromatography- mass spectrometry (GC-MS) to find new natural sources for essential fatty acids. Among the seed oils analyzed, Ceratonia ciliqua has been found to have the highest essential fatty acid content. PMID- 15277088 TI - A new compound from Forsythia suspensa (Thunb.) Vahl with antiviral effect on RSV. AB - A new compound, 2-(1,4-dihydroxy cyclohexanyl )-acetic acid isolated from the seeds of Forsythia suspensa (Thunb.) Vahl, has been assessed for potent antiviral effect on RSV for the first time in vitro by cell morphology methods. Its structure was elucidated on the basis of spectroscopic evidence (IR, MS, 1H-NMR, 13C-NMR). We have trivially named it Rengynic acid. PMID- 15277090 TI - Red clover (Trifolium pratense) monograph: a clinical decision support tool. AB - Red clover, a legume resembling soy is used by man as a phytoestrogen. Other uses include asthma, pertussis, cancer and gout. The authors systematically review this herb in terms of pharmacology, efficacy, safety, side effects, standardization, dosing, toxicology as well as other parameters. PMID- 15277091 TI - PC-SPES for the treatment of prostate cancer? AB - A 73-year-old man recently attended his local USToo International prostate cancer support group and during the meeting several members were discussing using PC SPES for treatment of their prostate cancer. The patient's history is significant for prostate cancer and as a result he had a radical prostatectomy. Two years ago, he relapsed with bone metastases and he was subsequently treated with combined anti-androgen therapy. Over the last 6 months his symptoms and prostate specific antigen have increased, despite anti-androgen withdrawal and a regimen of cancer chemotherapy. The patient is aware that his prostate cancer is incurable, but he wants to know if PC-SPES can help. PMID- 15277092 TI - Alkaloids of Thalictrum angustifolium. AB - Fractionation and chromatography of the ethanolic extract of the roots of Thalictrum angustifolium L. (Ranunculaceae) afforded five alkaloids: noroxyhydrastinine (1), O-methylthalicberine (2), berberine iodide (3), jatrorrhizine iodide (4), magnoflorine iodide (5). In addition, one simple aromatic compound, methyl-4-hydroxybenzoate (6), was also isolated. These compounds were identified via comparison of their spectral data with authentic compounds or spectra available within our laboratory. This is the first reported isolation of these compounds from this species. PMID- 15277093 TI - The clinical utility of milk thistle (Silybum marianum) in cirrhosis of the liver. AB - Milk thistle (Silybum marianum) is a flowering herb utilized for its potentially protective effects on the liver. Although the mechanism of action is not fully understood, one explanation may be that it concentrates in the hepatocytes and competes with toxins for hepatocyte binding and penetration. Preliminary clinical evaluations of milk thistle for cirrhosis of the liver indicate potential benefits in healthier patients with alcoholic cirrhosis. However, major flaws in many of the studies make it difficult to draw solid conclusions. Milk thistle appears to be relatively safe, even with long-term use. PMID- 15277094 TI - Effects of garlic extract supplementation on blood lipid and antioxidant parameters and atherosclerotic plaque formation process in cholesterol-fed rabbits. AB - OBJECTIVE: Possible effects of garlic extract supplementation on blood oxidant/antioxidant status, blood lipid profile and coronary plaque formation process were investigated in cholesterol-fed rabbits. METHODS: Thirty-one male rabbits of New Zealand strain were used. Twenty-two animals were given cholesterol for 4 months. Seven of them were sacrificed to investigate plaque formation and to measure blood parameters. Seven of the remaining 15 animals were fed on normal laboratory diet and others normal diet plus garlic extract for additional 3 months. Blood antioxidant and lipid parameters were measured and histological examination was made. RESULTS: Total, HDL and LDL cholesterol levels were found to be significantly higher in the Cholesterol Group relative to controls. In the histological investigation, a dense atherosclerotic plaque formation was observed in the aortas of this group. In the Normal Diet Group, total, HDL and LDL cholesterol levels were higher relative to the control group. No significant differences were observed between plaque surface areas of the Cholesterol and Normal Diet Groups. In the Extract Group however, there were differences with regard to all the analysis parameters. Total, HDL and LDL cholesterol levels were found to be decreased in this group. There was significant reduction in the plaque surface area in the aortas of this group. Blood antioxidant potential (AOP) was higher than the other groups but, malondialdehyde (MDA) level and, value of susceptibility to oxidation (SO) were lower in the Extract Group relative to the other groups. There were however no significant differences between MDA and SO values of the Control and Extract Groups. CONCLUSIONS: Our results demonstrate that cholesterol supplementation leads to dense plaque formation in the aortas of the rabbits. Garlic extract supplementation ameliorates blood lipid profile and, increases antioxidant potential. Extract treatment can significantly reduce plaque surface area in the aorta. Our results suggest that increased blood antioxidant potential due to extract supplementation might be one of the factors leading to this end. PMID- 15277095 TI - A review of the status of Western herbal medicine in Australia. AB - Western herbal medicine is the most widely used form of herbal medicine in Australia although Ayurvedic and Chinese herbal medicines are becoming better known. The agricultural production and manufacture of locally grown herbs is, with some exceptions, relatively underdeveloped, as is the research and development of indigenous flora. However, the use of herbal medicine is increasingly becoming mainstream with retail sales of herbal products in Australia estimated to be 200 million dollars. Concurrent with the increase in popularity of herbal medicine with health consumers have come advances in herbal medicine education and regulation. Although small by European standards, research into herbal medicines is increasing, mainly through industry and University-based initiatives. PMID- 15277096 TI - The inhibitory effects of pure flavonoids on in vitro protein glycosylation. AB - Non-enzymatic glycosylation of proteins is the major cause of diabetic complications, such as cardiovascular disorders, retinopathy, nephropathy and neuropathy. It seems that protein glycosylation can be inhibited effectively by antioxidants. Several flavonoids, such as rutin, kaempferol, quercetin, apigenin, naringin, morin and biochanin A were selected to determine their antioxidant effects on in vitro insulin, hemoglobin and albumin glycosylation. The optimal glucose concentration and incubation time were obtained for each protein. Then, the inhibition percentage of protein glycosylation was measured in the presence of three different concentrations (0.5, 5, 10 microg/ml) of each flavonoids by a colorimetric method. The results demonstrated that biochanin A, the best inhibitor of insulin and hemoglobin glycosylation, inhibits their glycosylation 100% and 60%, respectively. Glycosylation of albumin was inhibited 100% by both biochanin A and apigenin. Therefore, it seems probable that plants containing flavonoids may have preventive effects in diabetic complications. PMID- 15277097 TI - Possible subdural hematoma associated with Ginkgo biloba. AB - A 78-year-old Caucasian man developed headache, confusion and progressive right sided weakness following a fall resulting in ecchymosis over the left eye orbit five days prior to admission. A subdural hematoma was diagnosed upon CAT scan. Upon evacuation, a very large left frontoparietal subdural hematoma with the appearance of mixed elements of hemorrhage and older fluid was noted. The finding was chronic subdural hematoma described as "crankcase oil." The patient's only prior medications were lisinopril 20 mg daily and ginkgo biloba 50 mg three times a day. Ginkgo biloba is advocated to augment cerebral blood flow to enhance memory and improve dementia. One of Ginkgo biloba's components is ginkgolide B, a potent inhibitor of platelet activating factor essential for induction of arachidonate-independent platelet aggregation. We believe Ginkgo biloba either caused or predisposed this patient to subdural hematoma and is of concern given ginkgo's widespread use with minimal or no monitoring. PMID- 15277099 TI - Shark cartilage monograph: a clinical decision support tool. AB - The use, adverse effects, mechanism of action, pharmacology, interactions and supporting evidence of shark cartilage is presented. Dosage, toxicology and safety parameters are also discussed. PMID- 15277101 TI - Cancer therapy with mistletoe:hope or hype? PMID- 15277102 TI - Synergism between lectins and vesicles of Viscum album L.- detection by biochemical and immunological methods. AB - Mistletoe (Viscum album L.) extracts, used in cancer therapy, contain several antitumor and immunologically active ingredients of which the cytotoxic mistletoe lectins and the immunoactive vesicles of chloroplast membranes are particularly important. We have investigated interactions between vesicles and lectins with respect to the question of synergistic or antagonistic effects. First we used biochemical methods. Lectin binding to vesicles was dependent on the pH-value and ionic strength of the buffers used. The strongest interaction was observed at low pH-values and at low ionic strength. Using immunological methods, we found that the combination of lectins and vesicles showed a strong amplifying synergistic effect on the stimulation of lymphocyte proliferation. We found an antagonistic effect in terms of cytotoxicity. In summary, these results demonstrate a significant influence of vesicles on all commonly used methods of determination of mistletoe lectins. PMID- 15277103 TI - Herb-drug interactions and confounding in clinical trials. AB - Herb-drug interactions are being recognized for their potential to reduce the efficacy of, or cause additive side effects with, conventional medications. Herb use, particularly when undisclosed to investigators, may be a potential unforeseen confounding variable in clinical trials. The use of herbal products by patients enrolled in clinical trials should be determined and considered as a confounding variable. PMID- 15277104 TI - An assessment of herbal therapy use, adherence and utilization of pharmacy services in HIV clinics. AB - OBJECTIVE: To assess herbal therapy use, adherence to antiretroviral therapy (ART) and pharmacy service utilization in two HIV clinics using a prospective questionnaire-based assessment. RESULTS: Seventy-six patients completed the questionnaire. Twenty-six patients (34%) reported using at least one herbal therapy; 14 (54%) reported this to their provider. Providers correctly predicted herbal therapy use in 10 (38%) patients reporting herbal therapy use. Seventy three patients (96%) reported a high level of adherence (> 90%), while only 37% had a viral load < 80 copies/ml. Clinic and community-based pharmacy services were underutilized. CONCLUSIONS: Herbal therapy use was common, under-reported and difficult for providers to predict. Unreported herbal therapy use could lead to virologic failure as a result of unknown drug-herb interactions. Consultative pharmacy services in the clinic and retail pharmacies are underutilized. PMID- 15277105 TI - The use of a silymarin/phospholipid compound as a fetoprotectant from ethanol induced behavioral deficits. AB - We have replicated an earlier study in which silymarin/phytosome appeared to prevent deficits in social memory function in male rats exposed in utero to ethanol (EtOH).1 Female rats were included in the current study as well as a second behavioral test, the radial arm maze. Pregnant Sprague-Dawley rats were provided with liquid diets containing 35% ethanol derived calories (EDC). The silybin/phospholipid compound (SI) was co-administered with EtOH to the experimental group. The offspring were tested at age 90 days on the social recognition task and at 75 days on the radial arm maze. Female EtOH-exposed offspring performed more poorly on the radial arm maze than did female EtOH/SI offspring and the offspring of female controls. Male EtOH-exposed offspring were less able to form social memories than the male EtOH/SI offspring and the offspring of male controls. PMID- 15277106 TI - GS-02 for dysthymic disorder: results of a preliminary, open study. AB - In an open clinical trial, 15 patients diagnosed with DSM-IV dysthymic disorder were treated with GS-02, a herbal formulation containing extracts of four Indian herbs: Ashvatha, Kapikachu, Dhanvayasa, and Bhuriphali. Twelve patients completed the study. The medication was very well-tolerated. Among treatment completers, three (25%) patients showed no response, two (16.7%) showed partial response, and seven (58.3%) showed good response; these response rates are similar to what can be expected from an allopathic antidepressant trial. In an intent- to-treat analysis, significant improvement was observed on Hamilton depression ratings as well as on global measures. The results of this preliminary study encourage further clinical investigation of the GS-02 formulation. PMID- 15277108 TI - Resources for information on herbal medicinals and dietary supplements. PMID- 15277107 TI - Cardiovascular adverse reactions associated with Guarana: is there a causal effect? AB - Herbal supplements have been used as adjuncts to medical therapy for many years by various cultures. Many consumers believe that because herbal supplements are natural products, they are somewhat safer or more effective than traditional prescribed medications. This is also one reason that alternative medicine is growing and gaining more popularity. On the other hand, adverse reactions to herbal supplements or their interactions with patients' current medications are no different than pharmaceutical medicines. We report a case of premature ventricular contraction associated with two herbal supplements. These products contained multiple different herbs and both included large doses of guarana. Guarana, which is found in some supplements marketed in U.S., contains a substantial amount of caffeine. Although the exact cause of tachycardia in our report is not proven, a large amount of caffeine consumption is thought to be a possible causal effect. The purpose of this report is to remind health care professionals to evaluate and educate patients on the use of herbal products and any potential adverse reactions, drug interactions, or possible toxicities. PMID- 15277109 TI - Horse chestnut:a multidisciplinary clinical review. AB - Horse chestnut seed extract (HCSE) is widely used in Europe for the management of chronic venous insufficiency (CVI). Although traditionally recommended for a variety of medical conditions, CVI is the only indication for which there is strong supportive scientific evidence. Review of the literature reveals 14 randomized controlled trials, of which seven are methodologically of high quality, albeit limited by small sample sizes and short durations. These studies support the superiority of HCSE over placebo, and suggest equivalence to compression stockings and to oral oxerutins. In the future, a longer and adequately powered randomized trial is warranted to compare HCSE to standard of care, and to further assess safety and long-term efficacy. There are no data to suggest that horse chestnut flower, raw seed, branch bark, or leaf are effective for any indication, and it is recommended that these products not be used, as they are known to be toxic when ingested. PMID- 15277117 TI - Oral ingestion of Ginkgo biloba extract reduces thiobarbituric acid reacting (TBAR) substances in washed platelets of healthy subjects. AB - Thiobarbituric acid reacting (TBAR) substances were measured in washed platelets before and after ingesting 120 mg of standardized Gingko biloba extract daily for 3 months, in both normocholesterolemic (total cholesterol, 160 +/- 27 mg/dl; age 40 +/- 13 years; n = 18) and hypercholesterolemic subjects (total cholesterol, 229 +/- 35; age, 45 +/- 8 years, n = 12). Gingko biloba extract significantly reduced cellular content of TBAR substances; 42 +/- 21 vs. 28 +/- 16 pmol/10(7 platelets (p < 0.0025) and 50 +/- 17 vs. 29 +/- 13 pmol/10(7) platelets (P < 0.004), for the normo- and hypercholemic subjects, respectively. In conclusion, Gingko biloba extract is a potent antioxidant for both groups, reducing TBAR substances possibly by inhibiting platelet COX-1 isoform activity. PMID- 15277118 TI - Herbal healing in pregnancy: women's experiences. AB - Many women use home remedies to maintain their health during pregnancy. Here, pregnant women's perspectives on herbal medicine are exposed in a small (n = 27) non-random sample of pregnant women in British Columbia, Canada, and follow-up interviews with six mentors from the community. While many of the women were cautious about using herbs during pregnancy, they considered them to be safer-as a general rule-than pharmaceutical drugs. Herbal tonics were widely used, and simple home remedies were usually the first line of defence against common health complaints. In choosing to self-medicate with herbs, the women said they were guided by prior knowledge (32%), trusted sources of advice (56%), and intuition (12%). A reliance on prior knowledge was not strongly correlated with the woman's age (r = -0.27) or the number of pregnancies she had experienced (r = 0.21). Trusted sources of advice included books, friends, family members, maternity care providers, herbalists, herb shops, and internet. The majority of herbal advice (69%) was received by word-of-mouth. The women's mentors were an important source of herbal self-care information. PMID- 15277119 TI - The effect of a herbal water-extract on histamine release from mast cells and on allergic asthma. AB - A water extract of a mixture of eight herbs (chamomile, saffron, anise, fennel, caraway, licorice, cardomom and black seed) was tested for its inhibitory effect on histamine released from rat peritoneal mast cells stimulated either by compound 48/80 or be IgE/anti-IgE. The effect of the herb extract was compared to that of the flavonoid quercetin. The herbal water-extract inhibited histamine released from chemically- and immunologically-induced cells by 81% and 85%, respectively; quercetin treated cells were inhibited by 95% and 97%, respectively. The clinical results showed significant improvements of sleep discomfort, cough frequency and cough intensity in addition to increased percentages of FEV1/FVC in patients suffering from allergic asthma, who used the herbal tea compared to those who used the placebo tea. PMID- 15277120 TI - Cost evaluation of herbal medicine. PMID- 15277122 TI - Black cohosh and menopausal symptoms. PMID- 15277124 TI - Wild yam (Dioscoreaceae). PMID- 15277125 TI - Finding ways to close the gap. PMID- 15277127 TI - The academy movement: a structural approach to reinvigorating the educational mission. AB - Despite its fundamental importance, the educational mission of most medical schools receives far less recognition and support than do the missions of research and patient care. This disparity is based, in part, on the predominance of discipline-based departments, which focus on the more sustainable enterprises of research and patient care. Where departmental teaching is emphasized, it tends to center on trainees directly associated with the department-leaving medical students unsupported. The authors argue that the ongoing erosion of the educational mission will never be reversed unless there are changes in the underlying structure of medical schools. Academies of medical educators are developing at a number of medical schools to advance the school-wide mission of education. The authors describe and compare key features of such organizations at eight medical schools, identified through an informal survey of the Society of Directors of Research in Medical Education, along with direct contacts with specific schools. Although these entities are relatively new, initial assessments suggest that they have already had a major impact on the recognition of teaching efforts by the faculty, fueled curricular reform, promoted educational scholarship, and garnered new resources to support teaching. The academy movement, as a structural approach to change, shows promise for reinvigorating the educational mission of academic medicine. PMID- 15277126 TI - Medical education as a process management problem. AB - With complaints that new doctors are less prepared for residency and practice than expected, are burdened with debt, and then take even longer to complete their specialty training, the authors ask whether medical education can be designed more effectively. Curriculum redesign and pedagogical reform efforts to date address fragments of medical education-the content of particular courses or clerkships or the way in which the courses or clerkships are conducted. However, these reforms do not typically address the relationships among the various elements, that is, in what order skill sets should be sequenced, how communication should occur between disciplines, and by what mechanisms skills or knowledge should be mastered and assessed by the end of one phase so students are prepared adequately for the next. In failing to address these systems issues, current reform efforts may forgo some opportunities to convey and properly insure greater mastery of knowledge and skills in less time, at less cost. A case study of a typical student's third- and fourth-year clerkships illustrates how focusing only on educational elements leads to the exclusion of opportunities to systemically facilitate the relationships among them. This situation is contrasted with how other demanding, high-tech, knowledge-intensive industries with outstanding operations have learned to achieve superlative performance by managing and designing both the elements and the interactions among them within complex work and learning systems. The authors' exploratory research offers suggestions for medical education reform and frames additional opportunities for further discussion. PMID- 15277128 TI - Integrating communication training into a required family medicine clerkship. AB - Persistent evidence suggests that the communication skills of practicing physicians do not achieve desired goals of enhancing patient satisfaction, strengthening health outcomes and decreasing malpractice litigation. Stronger communication skills training during the clinical years of medical education might make use of an underutilized window of opportunity-students' clinical years to instill basic and important skills. The authors describe the implementation of a novel curriculum to teach patient-centered communication skills during a required third-year, six-week family medicine clerkship. Curriculum development and implementation across 24 training sites in a five-state region are detailed. A faculty development effort and strategies for embedding the curriculum within a diverse collection of training sites are presented. Student and preceptor feedback are summarized and the lessons learned from the curriculum development and implementation process are discussed. PMID- 15277129 TI - A medical school for international health run by international partners. AB - In early 1996, the Ben Gurion University Faculty of Health Sciences (BGU), Beer Sheva, Israel, in collaboration with Columbia University Medical Center (CUMC), New York City, United States, decided to found a second medical school within BGU, the Medical School for International Health (MSIH), to prepare students to work both in medicine and in cross-cultural and international health and medicine (IHM). Methods used to establish and jointly run MSIH include (1) defining clearly the tasks of each university according to how it can best contribute to the new school; (2) establishing an organizational structure in each university for accomplishing these tasks; (3) establishing clear communication between the two organizational structures; (4) defining outcomes to measure success; and (5) developing methods for addressing management problems. CUMC's functions were admission, public relations, and the fourth-year elective program. BGU's functions were developing and running an innovative curriculum, including a four year required track in IHM, evaluating students, taking the lead in helping students' with their personal problems, and managing financial aid. The first students were admitted in 1998. Variables reflecting MSIH's success include scores on the United States Medical Licensing Examination, residency placement, the attrition rate, and success in preparing students in IHM (e.g., success in learning cross-cultural medicine and the percentage of students who work in IHM). MSIH is running well and has solved its inter-university management problems. Its 85 graduates matched at very good to excellent U.S. hospitals and have learned and maintained enthusiasm for the IHM curriculum. PMID- 15277131 TI - Medicine and the arts. A piece of monologue. PMID- 15277130 TI - An innovative, longitudinal program to teach residents about end-of-life care. AB - At the University of California, Irvine Medical Center, an end-of-life curriculum was implemented in 2000 for an internal medicine residency utilizing a longitudinal approach that allowed residents to follow patients through their entire hospice experience. An elective home hospice rotation was developed for which third-year residents served as primary care physicians for patients at the end of life over a one-year period. Residents were supervised by faculty who were hospice medical directors. They also learned through case vignettes, quarterly meetings, textbook reading, and personal projects. From July 2000 to June 2002, residents demonstrated positive attitudes towards hospice care and recommended the rotation highly (mean 8.86 on a scale of 1-10). The rotation grew in popularity from six initial residents to ten residents the next year, and has since become a mandatory rotation for all senior residents. A 360-degree evaluation uniformly indicated positive resident performance from the hospice team (mean scores 7.56-8.69 on a 1-9 scale), family (mean scores 9.3-9.7 on a 1 10 scale) and faculty (mean scores 7.29-7.72 on a 1-9 scale). Residents were also pleased with the level of teaching (mean 8.86 on a scale of 1-10) and felt that the patient care load was "just right." Their knowledge improved by 8% (p =.0175). In conclusion, a longitudinal hospice rotation was implemented that fulfilled curricular goals without undue burden on the residents or residency program. PMID- 15277133 TI - End-of-life care in the curriculum: a national study of medical education deans. AB - PURPOSE: To describe attitudes and practices of end-of-life care teaching in the undergraduate medical curriculum in the United States as reported by administrative leadership and identify opportunities for improvement. METHOD: A telephone survey of associate deans for medical education or curricular affairs at a random sample of 62 accredited U.S. medical schools was conducted in 2002. RESULTS: Fifty-one deans participated (82% response rate). Most (84%) described end-of-life care education as "very important" and supported incorporating more end-of-life care teaching into the undergraduate curriculum. Sixty-seven percent reported that insufficient time is currently given to palliative care in their curriculum. Although a majority opposed required courses (59%) or clerkships (70%) that focused on end-of-life care, they did unanimously endorse integrating teaching end-of-life care into existing courses or clerkships. Key barriers to incorporating more end-of-life care into the curriculum included lack of time in the curriculum, lack of faculty expertise, and absence of a faculty leader. CONCLUSION: Associate deans for medical education or curricular affairs in the United States support integrating end-of-life care content into existing courses and clerkships throughout the undergraduate medical curriculum. Successful integration will require institutional investment in faculty development, including both the development of faculty leaders to drive change efforts, and the education of all faculty who teach students and exert influence as role models and mentors. The strong support for end-of-life care education expressed by academic leaders in this study, combined with the high level of interest expressed in the authors' 2001 national survey of students, provide evidence of the potential for meaningful change in the undergraduate medical curriculum. PMID- 15277134 TI - Interdisciplinary education: evaluation of a palliative care training intervention for pre-professionals. AB - PURPOSE: Medical education inadequately prepares students for interdisciplinary collaboration, an essential component of palliative care and numerous other areas of clinical practice. This study developed and evaluated an innovative interdisciplinary educational program in palliative care designed to promote interdisciplinary exchange and understanding. METHOD: The study used a quasi experimental longitudinal design. Thirty-three medical students (third and fourth year) and 38 social work students (second year of masters degree) were recruited. The intervention group students (21 medical and 24 social work students) participated in a series of four training sessions over four weeks while the control group students received written materials after the study. The curriculum and teaching methods were based on theories of professional socialization and experiential learning. The intervention included experiential methods to promote interdisciplinary interaction to foster communication, exchange of perspectives, and the building of mutual trust and respect. Both groups completed assessments of perceived role understanding, a primary component of effective interdisciplinary teamwork, in palliative care. Self-administered surveys were completed at baseline, intervention completion, and three months later. The intervention group also completed an anonymous evaluation about the interdisciplinary education. RESULTS: The intervention group demonstrated a significant increase in perceived role understanding compared with the control group. Three-month follow-up data suggested that intervention group subjects maintained gains in perceived role understanding. CONCLUSION: An interdisciplinary educational intervention improves role understanding early in the process of professional socialization in a pilot program. Further implementation of interdisciplinary education should evaluate the effect on subsequent interdisciplinary practice and the quality of patient care. PMID- 15277135 TI - Improving knowledge in palliative medicine with a required hospice rotation for third-year medical students. AB - PURPOSE: The Liaison Committee for Medical Education requires accredited U.S. and Canadian medical schools to teach end-of-life care. The purpose of this study was to evaluate a new required curriculum in palliative medicine for third-year medical students. METHOD: Beginning in July 2001, a required four-day (32 hour) curriculum was piloted as part of an ambulatory month in the 12-week medicine clerkship. Students spent Day 1 in the classroom learning core concepts regarding hospice, palliative care, and symptom management. A two-hour session with a standardized patient to break bad news was included. Students spent Days 2 and 3 making home visits or participating in inpatient care. Day 4 was spent in the classroom reviewing cases they had seen with interdisciplinary faculty, making presentations on assigned topics, and discussing professional self-care. Students completed a self-awareness project. Educational outcomes were measured with the students' completion of five pre- and postcourse assessment instruments: (1) self assessment of competency, (2) attitudes, (3) concerns, (4) a 50-item, multiple choice knowledge test, and (5) an assessment of elements of the course. RESULTS: Analysis of 127 paired evaluations showed significant improvements in three instruments: 56% improvement in competence (p <.0001), 29% reduction in concern (p <.0001), and 23% improvement in knowledge (p <.0001). There were no significant changes attitudes (p =.35). CONCLUSION: This 32-hour required curriculum in palliative medicine for third-year medical students improved knowledge. They came to the course with appropriate attitudes that did not change. PMID- 15277136 TI - Documentation systems for educators seeking academic promotion in U.S. medical schools. AB - PURPOSE: To explore the state and use of teaching portfolios in promotion and tenure in U.S. medical schools. METHOD: A two-phase qualitative study using a Web based search procedure and telephone interviews was conducted. The first phase assessed the penetration of teaching portfolio-like systems in U.S. medical schools using a keyword search of medical school Web sites. The second phase examined the current use of teaching portfolios in 16 U.S. medical schools that reported their use in a survey in 1992. The individual designated as having primary responsibility for faculty appointments/promotions was contacted to participate in a 30-60 minute interview. RESULTS: The Phase 1 search of U.S. medical schools' Web sites revealed that 76 medical schools have Web-based access to information on documenting educational activities for promotion. A total of 16 of 17 medical schools responded to Phase 2. All 16 continued to use a portfolio like system in 2003. Two documentation categories, honors/awards and philosophy/personal statement regarding education, were included by six more of these schools than used these categories in 1992. Dissemination of work to colleagues is now a key inclusion at 15 of the Phase 2 schools. The most common type of evidence used to document education was learner and/or peer ratings with infrequent use of outcome measures and internal/external review. CONCLUSIONS: The number of medical schools whose promotion packets include portfolio-like documentation associated with a faculty member's excellence in education has increased by more than 400% in just over ten years. Among early-responder schools the types of documentation categories have increased, but students' ratings of teaching remain the primary evidence used to document the quality or outcomes of the educational efforts reported. PMID- 15277137 TI - Evaluation of a teaching workshop for residents at the University of Saskatchewan: a pilot study. AB - PURPOSE: The University of Saskatchewan College of Medicine in Saskatoon, Canada, has been running a two-day workshop for teachers since 1993 to which both faculty and residents had been invited. Although the design of the workshop is consistent with principles of adult learning, it was important to determine if the workshop effectively helped residents to acquire and then use key teaching skills in real world situations where they were called on to organize and make presentations. METHOD: This study, conducted in 1998 and 1999 with residents only, used a randomized controlled experiment with third-party ratings of before and after videotaped teaching sessions done in actual performance settings. There were eight residents each in the control and intervention groups. RESULTS: The intervention group made statistically significant and positive changes in two key areas taught in the workshop and showed slight improvement in a third. The changes made by the residents in the intervention group were in presenting the opening "set" (41.4% absolute improvement) and the use of instructional objectives (11.5%). The "body" of their teaching sessions increased slightly (9.3%). The control group held relatively stable. CONCLUSION: While this study demonstrates that the workshop likely made a difference in the teaching performance of the intervention group, the small sample size (eight in each group) and the presence of confounding variables suggest that further research should be conducted. PMID- 15277138 TI - Effectiveness of a brief workshop designed to improve teaching performance at the University of Alberta. AB - PURPOSE: To determine whether a two-day teaching enhancement workshop at the University of Alberta improved participants' teaching performance as rated by students. METHOD: Workshop participants (academic staff or residents) were asked to assess the value of the workshop. In addition, students were asked to rate instructors' teaching abilities before and after the instructors participated in the workshops, by completing a five-statement questionnaire routinely used to assess instruction at the University of Alberta. For control purposes, ratings were also obtained for a group of instructors who had not taken the workshop, over a similar time period. The authors used data from 1993-2002. RESULTS: The participants uniformly regarded the workshops as helpful. Both faculty and residents regarded the short teaching exercise as the most important component of the program. Of the instructional sections, the presentations on objectives and on structure (set, body, closure) were rated most highly by both groups. The students' mean ratings for the instructors after the workshop were significantly increased, while ratings for those who had not taken the workshop were unchanged CONCLUSION: Short teaching-enhancement workshops are regarded by the participants as helpful in improving their instructional skills. This view is supported by a significant increase in students' ratings of the instructors after they had taken the workshop. PMID- 15277139 TI - Validity of scores generated by a web-based multimedia simulated patient case software: a pilot study. AB - PURPOSE: The value of multimedia simulated patient cases (MSPCs) in medical education remains unclear. The authors conducted a pilot study to assess the validity of automated scores of diagnostic reasoning ability provided by DxR Clinician, a widely available Web-based MSPC software. METHOD: In 2002-03, all 89 students enrolled in a required third-year primary care clerkship at the University of California, Davis, School of Medicine were assigned to complete four MSPCs. The authors determined the degree of correlation between the Clinical Reasoning Score (CRS) and Level of Diagnostic Performance (LDP) generated by the MSPC software and subscale scores from a validated measure of diagnostic reasoning sophistication, the Diagnostic Thinking Inventory (DTI). RESULTS: Of 356 completed case events, instructor override of automated scoring was required in 206 (58%) to obtain an accurate LDP and CRS. Mean DTI subscale scores improved significantly from the beginning to the end of the year (p <.0001, Wilcoxon signed rank test). However, there were no significant correlations between CRS or LDP scores on any of the four cases and either of the two DTI subscale scores. CONCLUSION: Automated diagnostic reasoning scores generated by one widely available MSPC software appear to lack criterion validity. The validity of automated diagnostic reasoning scores generated by MSPCs should be established before such cases can be confidently employed as educational tools. PMID- 15277140 TI - Is folic acid the answer? PMID- 15277141 TI - Postprandial glycemia, glycemic index, and the prevention of type 2 diabetes. PMID- 15277142 TI - Yogurt and gut function. AB - In recent years, numerous studies have been published on the health effects of yogurt and the bacterial cultures used in the production of yogurt. In the United States, these lactic acid-producing bacteria (LAB) include Lactobacillus and Streptococcus species. The benefits of yogurt and LAB on gastrointestinal health have been investigated in animal models and, occasionally, in human subjects. Some studies using yogurt, individual LAB species, or both showed promising health benefits for certain gastrointestinal conditions, including lactose intolerance, constipation, diarrheal diseases, colon cancer, inflammatory bowel disease, Helicobacter pylori infection, and allergies. Patients with any of these conditions could possibly benefit from the consumption of yogurt. The benefits of yogurt consumption to gastrointestinal function are most likely due to effects mediated through the gut microflora, bowel transit, and enhancement of gastrointestinal innate and adaptive immune responses. Although substantial evidence currently exists to support a beneficial effect of yogurt consumption on gastrointestinal health, there is inconsistency in reported results, which may be due to differences in the strains of LAB used, in routes of administration, or in investigational procedures or to the lack of objective definition of "gut health." Further well-designed, controlled human studies of adequate duration are needed to confirm or extend these findings. PMID- 15277143 TI - Weight management through lifestyle modification for the prevention and management of type 2 diabetes: rationale and strategies. A statement of the American Diabetes Association, the North American Association for the Study of Obesity, and the American Society for Clinical Nutrition. AB - Overweight and obesity are important risk factors for type 2 diabetes. The marked increase in the prevalence of overweight and obesity is presumably responsible for the recent increase in the prevalence of type 2 diabetes. Lifestyle modification aimed at reducing energy intake and increasing physical activity is the principal therapy for overweight and obese patients with type 2 diabetes. Even moderate weight loss in combination with increased activity can improve insulin sensitivity and glycemic control in patients with type 2 diabetes and prevent the development of type 2 diabetes in high-risk persons (ie, those with impaired glucose tolerance). The American Diabetes Association, the North American Association for the Study of Obesity, and the American Society for Clinical Nutrition have joined together to issue this statement on the use of lifestyle modification in the prevention and management of type 2 diabetes. PMID- 15277144 TI - Should calcium and vitamin D be added to the current enrichment program for cereal-grain products? AB - Mean dietary intakes of calcium and vitamin D in the US adult population are far below the adequate intake (AI) values recommended by the Food and Nutrition Board, Institute of Medicine of the National Academy of Sciences, and thus substantial segments of the American population have inadequate intakes and elevated risks of osteoporosis and colon cancer. The current Code of Federal Regulations, Title 21, sets standards for the optional addition of moderate amounts of calcium and vitamin D in the enrichment of cereal-grain products, a provision that is essentially not used. We propose that the addition of calcium and vitamin D to currently enriched cereal-grain products be mandated in the United States: this would result in an increase in mean daily dietary intakes in the United States of approximately 400 mg Ca and > or =50 IU (or possibly >200 IU) vitamin D. The benefits would be a significant reduction in the incidences of osteoporosis and colon cancer over time and overall improvement in health, with little risk and a modest financial cost because of the ability to capitalize on existing technology. We suggest a full scientific review of cereal-grain enrichment with calcium and vitamin D. PMID- 15277146 TI - Effects of cis-9,trans-11 conjugated linoleic acid supplementation on insulin sensitivity, lipid peroxidation, and proinflammatory markers in obese men. AB - BACKGROUND: We recently showed that trans-10,cis-12 (t10,c12) conjugated linoleic acid (CLA) causes insulin resistance in obese men. However, metabolic effects of the c9,t11 CLA isomer are still unknown in obese men. Because c9,t11 CLA is the predominant CLA isomer in foods and is included in dietary weight-loss products, it is important to conduct randomized controlled studies that use c9,t11 CLA preparations. OBJECTIVE: We investigated the effects of c9,t11 CLA supplementation on insulin sensitivity, body composition, and lipid peroxidation in a group at high risk for cardiovascular disease. DESIGN: In a randomized, double-blind, placebo-controlled study, 25 abdominally obese men received 3 g c9,t11 CLA/d or placebo (olive oil). Before and after 3 mo of supplementation, we assessed insulin sensitivity (hyperinsulinemic euglycemic clamp), lipid metabolism, body composition, and urinary 8-iso-prostaglandin F(2alpha) (a major F(2)-isoprostane) and 15-keto-dihydro-prostaglandin F(2alpha), markers of in vivo oxidative stress and inflammation, respectively. RESULTS: All subjects completed the study. Compared with placebo, c9,t11 CLA decreased insulin sensitivity by 15% (P < 0.05) and increased 8-iso-prostaglandin F(2alpha) and 15-keto-dihydro prostaglandin F(2alpha) excretion by 50% (P < 0.01) and 15% (P < 0.05), respectively. The decreased insulin sensitivity was independent of changes in serum lipids, glycemia, body mass index, and body fat but was abolished after adjustment for changes in 8-iso-prostaglandin F(2alpha) concentrations. There were no differences between groups in body composition. CONCLUSIONS: A CLA preparation containing the purified c9,t11 CLA isomer increased insulin resistance and lipid peroxidation compared with placebo in obese men. Because c9,t11 CLA occurs in commercial supplements as well as in the diet, the present results should be confirmed in larger studies that also include women. PMID- 15277145 TI - Visceral adipose tissue: relations between single-slice areas and total volume. AB - BACKGROUND: Visceral adipose tissue (VAT), which is linked with the metabolic consequences of obesity, is usually characterized by measuring VAT area at the L4 L5 vertebral interspace. However, the location of the slice with the strongest relation to VAT volume is not established. OBJECTIVE: We sought to investigate the relations between cross-sectional VAT areas at different anatomic locations and VAT volume in a large, diverse sample of healthy subjects. DESIGN: VAT volume was derived from slice areas taken at 5-cm intervals from magnetic resonance images in 121 healthy men [x +/- SD age: 41.9 +/- 15.8 y; body mass index (BMI; in kg/m(2)): 26.0 +/- 3.2; VAT: 2.7 +/- 1.8 L] and 198 healthy women (age: 48.1 +/- 18.7 y; BMI: 27.0 +/- 5.4; VAT: 1.7 +/- 1.2 L). Regression models were developed to identify the best single slice for estimating VAT volume. RESULTS: The VAT area 10 cm above L4-L5 (A(+10)) in men (R(2) = 0.932, P < 0.001) and 5 cm above L4-L5 (A(+5)) in women (R(2) = 0.945, P < 0.001) had the highest correlation with abdominal VAT. R(2) increased by only 3.8% in men and 0.5% in women with adjustment for age, race, scanning position, BMI, and waist circumference. Studies using A(+10) in men and A(+5) in women will require 14% and 9% fewer subjects, respectively, than those using slices at L4-L5 and will have equivalent power. CONCLUSION: Measurement of slice areas at A(+10) in men and A(+5) in women provides greater power for the detection of VAT volume differences than does measurement at L4-L5. PMID- 15277147 TI - Postprandial lipid and carbohydrate responses after the ingestion of a casein enriched mixed meal. AB - BACKGROUND: Postprandial lipemia is markedly modulated when carbohydrates are added to a fatty meal. The effect of added protein is less known, however, and the data are controversial. OBJECTIVE: We investigated the effects of casein added to various fat-rich meals in the absence and presence of oligosaccharides. DESIGN: Four different test meals were given to 24 healthy volunteers: 1) fat alone, consisting of 3 g cream/kg body wt; 2) fat plus 75 g oligosaccharides; 3) fat plus 50 g sodium caseinate; and 4) a combination of all 3 components. Blood samples were taken before the meals and 1, 2, 3, 4, 5, 6, 7, and 8 h thereafter. The variables measured were serum free fatty acids, arginine, glucose, insulin, and C-peptide as well as triacylglycerol in serum, in chylomicrons, and in VLDL. Gastric emptying was monitored with the use of a (13)C breath test. RESULTS: Addition of oligosaccharides resulted in the known delay and reduction in postprandial lipemia. Casein caused additional effects: chylomicrons were further reduced and delayed, independently of gastric emptying. C-peptide and insulin, as expressed by their areas under the curves, were raised not only during the early response to the glucose load but also in the postabsorptive state. Concentrations of free fatty acids, which were markedly suppressed by 24% after oligosaccharides alone, were lowered a further 20% after the addition of casein. CONCLUSIONS: Casein added to a fatty meal lowers free fatty acids markedly in the postprandial and postabsorption phases, probably via its insulinotropic activity. Postprandial lipemia is also moderately reduced. PMID- 15277148 TI - Serum cholesterol concentrations are associated with visuomotor speed in men: findings from the third National Health and Nutrition Examination Survey, 1988 1994. AB - BACKGROUND: Current international recommendations advise aggressive treatment of relative hypercholesterolemia despite an incomplete understanding of any neurobehavioral effects of low or lowered serum cholesterol. OBJECTIVE: The objective was to examine the relation between serum cholesterol concentrations and performance in immediate memory, visuomotor speed, and coding speed tests. DESIGN: The participants were 4110 adults aged 20-59 y who completed a set of neurobehavioral tests and had blood specimens collected as a part of the third National Health and Nutrition Examination Survey, 1988-1994. RESULTS: After adjustment for sociodemographic variables, serum trace elements and vitamins, dietary energy intake, and risk factors for cardiovascular disease, we found inverse linear associations of serum total cholesterol and non-HDL cholesterol with visuomotor speed in men. The least-squares mean (+/- SE) visuomotor speeds were 231.6 +/- 2.6, 224.0 +/- 2.2, and 218.9 +/- 2.5 ms, respectively, for men with serum total cholesterol concentrations below the 25th, between the 25th and the 75th, and at or above the 75th percentile (P for trend < 0.001) and were 231.7 +/- 2.7, 225.8 +/- 2.4, and 214.1 +/- 2.3 ms, respectively, for men with a non-HDL-cholesterol concentration below the 25th, between the 25th and the 75th, and at or above the 75th percentile (P for trend < 0.001). No significant associations were observed between memory or coding speed and the selected serum cholesterol measures in men, and the scores of the 3 neurobehavioral tests were unrelated to serum cholesterol in women. CONCLUSION: Low serum total cholesterol and non-HDL cholesterol are associated with slow visuomotor speed in young and middle-aged men. PMID- 15277149 TI - Appetite and inflammation, nutrition, anemia, and clinical outcome in hemodialysis patients. AB - BACKGROUND: Malnutrition-inflammation complex syndrome, an outcome predictor in maintenance hemodialysis (MHD) patients, may be related to anorexia. OBJECTIVES: We examined whether subjectively reported appetite is associated with adverse conditions and increased morbidity and mortality in MHD patients. DESIGN: A cohort of 331 MHD outpatients was asked to rate their recent appetite status on a scale from 1 to 4 (very good, good, fair, and poor appetite, respectively). Anemia indexes and nutritional and inflammatory markers-including serum concentrations of C-reactive protein, tumor necrosis factor alpha, and interleukin 6-were measured. The malnutrition-inflammation score was used to evaluate the malnutrition-inflammation complex syndrome, and the SF36 questionnaire was used to assess quality of life (QoL). Mortality and hospitalization were followed prospectively for up to 12 mo. RESULTS: Patients were aged 54.5 +/- 14.4 y. Diminished appetite (fair to poor) was reported by 124 patients (38%). Hemoglobin, protein intake, and QoL scores were progressively lower, whereas markers of inflammation, malnutrition-inflammation scores, and the required erythropoietin dose were higher across the worsening categories of appetite. The adjusted odds ratios of diminished versus normal appetite for increased serum tumor necrosis factor alpha and C-reactive protein concentrations were significant. Significant associations between a poor appetite and an increased rate of hospitalization and mortality were observed. The hazard ratio of death for diminished appetite was 4.74 (95% CI: 1.85, 12.16; P = 0.001). CONCLUSION: Diminished appetite (anorexia) is associated with higher concentrations of proinflammatory cytokines and higher levels of erythropoietin hyporesponsiveness and poor clinical outcome, including a 4-fold increase in mortality, greater hospitalization rates, and a poor QoL in MHD patients. Appetite status may yield significant insight into the clinical status of dialysis patients. PMID- 15277150 TI - Independent and additive effects of energy restriction and exercise on glucose and insulin concentrations in sedentary overweight men. AB - BACKGROUND: Overweight and inactivity are associated with impaired glucose tolerance, reduced insulin sensitivity, and diabetes. Few controlled trials have assessed the independent and combined effects of energy restriction and exercise on the prevention of these conditions. OBJECTIVE: The objective was to evaluate the independent and additive effects of 16 wk of energy restriction and exercise on glucose and insulin concentrations. DESIGN: Sixty nonsmoking, overweight, sedentary men aged 20-50 y were randomly assigned to either maintain or restrict their energy intake (4186-6279 kJ/d). Within each of these arms, the subjects were further randomly assigned to either a light-intensity (control) or a vigorous-intensity exercise program for 30 min 3 times/wk. RESULTS: Fifty-one subjects completed the study. Maximal oxygen uptake increased ( approximately 24%; P < 0.001) with vigorous but not with light exercise. Significant weight loss was observed with energy restriction (x: 10.12 kg; 95% CI: 8.02, 12.22 kg; P < 0.001) but not with exercise. Vigorous exercise reduced fasting glucose and glucose and insulin areas under the curve (AUCs) by 13% (P = 0.01) and 20% (P = 0.02), respectively. Exercise effects were independent of weight change. Energy restriction resulted in a 40% reduction in the insulin AUC (P = 0.01). Vigorous exercise and energy restriction were additive in reducing the insulin AUC. CONCLUSIONS: Energy restriction and vigorous exercise independently and additively reduce glucose and insulin concentrations in response to an oral glucose-tolerance test. Both of these lifestyle interventions provide a potent strategy that should be an integral part of any program to reduce the risk of impaired glucose tolerance, insulin resistance, and diabetes in overweight and sedentary persons. PMID- 15277152 TI - Association of body size with outcomes among patients beginning dialysis. AB - BACKGROUND: Although obesity confers an increased risk of mortality in the general population, observational reports on the dialysis population have suggested that obesity is associated with improved survival. These reports have generally not examined extremely high values of body mass index (BMI; in kg/m(2)), survival >1 y, or alternative measures of adiposity. OBJECTIVE: We sought to clarify the relation between body size and outcomes among a large cohort of patients beginning dialysis. DESIGN: Data on 418 055 patients beginning dialysis between 1 April 1995 and 1 November 2000 were analyzed by using US Renal Data System data. BMI was divided into 8 categories in increments of 3 units, ranging from < 19 to > or =37, and the relation between survival and BMI was examined by using proportional hazards regression with adjustment for demographic, laboratory, and comorbidity data. RESULTS: High BMI was associated with increased survival in this cohort, even at extremely high BMI, after adjustment, and over a 2-y average follow-up time. This was true for whites, African Americans, and Hispanics but not for Asians. High BMI was also associated with a reduced risk of hospitalization and a lower rate of mortality in all mortality categories. Alternative estimates of adiposity, including the Benn index and estimated fat mass, yielded similar results, and adjustments for lean body mass did not substantially alter the findings. CONCLUSIONS: High BMI is not associated with increased mortality among patients beginning dialysis. This finding does not appear to be a function of lean body mass and, although modified by certain patient characteristics, it is a robust finding. PMID- 15277151 TI - Serum folate and homocysteine and the incidence of acute coronary events: the Kuopio Ischaemic Heart Disease Risk Factor Study. AB - BACKGROUND: Several, but not all, prospective studies have shown that low folate intakes, low circulating folate concentrations, or high plasma total homocysteine (tHcy) concentrations are associated with an increased risk of coronary artery disease (CAD). OBJECTIVE: We examined the relations of both serum folate and serum tHcy concentrations with acute coronary events in middle-aged men from eastern Finland who had no CAD at baseline. DESIGN: In a population-based prospective cohort study, 1027 men aged 46-64 y were examined in 1991-1993 as part of the Kuopio Ischaemic Heart Disease Risk Factor Study. During an average follow-up of 7.7 y (7900 person-years of follow-up), 114 acute coronary events were observed in 61 men who had no previous history of CAD (n = 810). RESULTS: In a Cox model, compared with men whose serum folate concentrations were in the lowest tertile, those whose concentrations were in the highest tertile had a risk factor-adjusted relative risk of acute coronary events of 0.35 (95% CI: 0.17, 0.73; P = 0.005). Serum tHcy concentrations were not significantly associated with the risk of acute coronary events (for the highest tertile compared with the lowest, adjusted relative risk = 1.03; 95% CI: 0.57, 1.87; P = 0.932). CONCLUSIONS: The results of this prospective cohort study do not support the hypothesis that a high circulating tHcy concentration is a risk factor for acute coronary events in a male population free of prior heart disease. However, they do suggest that moderate-to-high serum folate concentrations are associated with a greatly reduced incidence of acute coronary events. PMID- 15277153 TI - Adjusting body cell mass for size in women of differing nutritional status. AB - BACKGROUND: Body cell mass (BCM) may be estimated in clinical practice to assess functional nutritional status, eg, in patients with anorexia nervosa. Interpretation of the data, especially in younger patients who are still growing, requires appropriate adjustment for size. Previous investigations of this general issue have addressed chemical rather than functional components of body composition and have not considered patients at the extremes of nutritional status, in whom the ability to make longitudinal comparisons is of particular importance. OBJECTIVE: Our objective was to determine the power by which height should be raised to adjust BCM for height in women of differing nutritional status. DESIGN: BCM was estimated by (40)K counting in 58 healthy women, 33 healthy female adolescents, and 75 female adolescents with anorexia nervosa. The relation between BCM and height was explored in each group by using log-log regression analysis. RESULTS: The powers by which height should be raised to adjust BCM were 1.73, 1.73, and 2.07 in the women, healthy female adolescents, and anorexic female adolescents, respectively. A simplified version of the index, BCM/height(2), was appropriate for all 3 categories and was negligibly correlated with height. CONCLUSIONS: In normal-weight women, the relation between height and BCM is consistent with that reported previously between height and fat-free mass. Although the consistency of the relation between BCM and fat-free mass decreases with increasing weight loss, the relation between height and BCM is not significantly different between normal-weight and underweight women. The index BCM/height(2) is easy to calculate and applicable to both healthy and underweight women. This information may be helpful in interpreting body-composition data in clinical practice. PMID- 15277155 TI - Glycemic index, glycemic load, and dietary fiber intake and incidence of type 2 diabetes in younger and middle-aged women. AB - BACKGROUND: Increasing evidence suggests an important role of carbohydrate quality in the development of type 2 diabetes. OBJECTIVE: Our objective was to prospectively examine the association between glycemic index, glycemic load, and dietary fiber and the risk of type 2 diabetes in a large cohort of young women. DESIGN: In 1991, 91249 women completed a semiquantitative food-frequency questionnaire that assessed dietary intake. The women were followed for 8 y for the development of incident type 2 diabetes, and dietary information was updated in 1995. RESULTS: We identified 741 incident cases of confirmed type 2 diabetes during 8 y (716 300 person-years) of follow-up. After adjustment for age, body mass index, family history of diabetes, and other potential confounders, glycemic index was significantly associated with an increased risk of diabetes (multivariate relative risks for quintiles 1-5, respectively: 1, 1.15, 1.07, 1.27, and 1.59; 95% CI: 1.21, 2.10; P for trend = 0.001). Conversely, cereal fiber intake was associated with a decreased risk of diabetes (multivariate relative risks for quintiles 1-5, respectively: 1, 0.85, 0.87, 0.82, and 0.64; 95% CI: 0.48, 0.86; P for trend = 0.004). Glycemic load was not significantly associated with risk in the overall cohort (multivariate relative risks for quintiles 1-5, respectively: 1, 1.31, 1.20, 1.14, and 1.33; 95% CI: 0.92, 1.91; P for trend = 0.21). CONCLUSIONS: A diet high in rapidly absorbed carbohydrates and low in cereal fiber is associated with an increased risk of type 2 diabetes. PMID- 15277154 TI - No difference in body weight decrease between a low-glycemic-index and a high glycemic-index diet but reduced LDL cholesterol after 10-wk ad libitum intake of the low-glycemic-index diet. AB - BACKGROUND: The role of glycemic index (GI) in appetite and body-weight regulation is still not clear. OBJECTIVE: The objective of the study was to investigate the long-term effects of a low-fat, high-carbohydrate diet with either low glycemic index (LGI) or high glycemic index (HGI) on ad libitum energy intake, body weight, and composition, as well as on risk factors for type 2 diabetes and ischemic heart disease in overweight healthy subjects. DESIGN: The study was a 10-wk parallel, randomized, intervention trial with 2 matched groups. The LGI or HGI test foods, given as replacements for the subjects' usual carbohydrate-rich foods, were equal in total energy, energy density, dietary fiber, and macronutrient composition. Subjects were 45 (LGI diet: n = 23; HGI diet: n = 22) healthy overweight [body mass index (in kg/m(2)): 27.6 +/- 0.2] women aged 20-40 y. RESULTS: Energy intake, mean (+/- SEM) body weight (LGI diet: -1.9 +/- 0.5 kg; HGI diet: -1.3 +/- 0.3 kg), and fat mass (LGI diet: -1.0 +/- 0.4 kg; HGI diet: -0.4 +/- 0.3 kg) decreased over time, but the differences between groups were not significant. No significant differences were observed between groups in fasting serum insulin, homeostasis model assessment for relative insulin resistance, homeostasis model assessment for beta cell function, triacylglycerol, nonesterified fatty acids, or HDL cholesterol. However, a 10% decrease in LDL cholesterol (P < 0.05) and a tendency to a larger decrease in total cholesterol (P = 0.06) were observed with consumption of the LGI diet as compared with the HGI diet. CONCLUSIONS: This study does not support the contention that low-fat LGI diets are more beneficial than HGI diets with regard to appetite or body-weight regulation as evaluated over 10 wk. However, it confirms previous findings of a beneficial effect of LGI diets on risk factors for ischemic heart disease. PMID- 15277156 TI - Postabsorptive and insulin-stimulated energy and protein metabolism in patients with myotonic dystrophy type 1. AB - BACKGROUND: Exaggerated insulin resistance was described as the major metabolic abnormality in myotonic dystrophy type 1 (DM1). We reported recently that the severity of the impairment in insulin-stimulated glucose metabolism in these patients was overestimated. OBJECTIVE: The aim was to dissect out insulin action with respect to whole-body energy homeostasis and glucose, protein, and lipid metabolism in patients with DM1 to assess the relevance of insulin resistance to the heterogeneous clinical manifestations of this syndrome. DESIGN: Ten nondiabetic patients with DM1 and 10 matched healthy control subjects were studied by means of 1) dual-energy X-ray absorptiometry; 2) a euglycemic hyperinsulinemic clamp (40 mU. m(-2). min(-1)) combined with a primed, continuous infusion of [6,6-d(2)]glucose and [1-(13)C]leucine; 3) indirect calorimetry; and 4) localized (1)H magnetic resonance spectroscopy of the calf muscles. RESULTS: Patients with DM1 had less lean body mass, greater fat mass, and greater intramyocellular lipid contents than did healthy control subjects. Energy expenditure and glucose and lipid metabolism did not differ significantly between the groups. In contrast, markers of proteolysis were higher in DM1 patients in the postabsorptive and insulin-stimulated conditions and were associated with lower plasma concentrations of insulin-like growth factor 1 (P < 0.03) and higher plasma concentrations of tumor necrosis factor alpha receptor 2 (P = 0.04). CONCLUSIONS: Despite greater body fat and intramyocellular lipid contents in patients with DM1, insulin sensitivity was not significantly different between patients and control subjects. In contrast, the loss of lean body mass in patients with DM1 was associated with abnormal postabsorptive and insulin stimulated regulation of protein breakdown. Lower plasma insulin-like growth factor 1 concentrations and higher tumor necrosis factor system activity might be involved in the muscle wasting of DM1. PMID- 15277158 TI - Antioxidant deficiency in cystic fibrosis: when is the right time to take action? AB - BACKGROUND: Little is known about age- and disease-related changes in prooxidant and antioxidant systems in patients with cystic fibrosis (CF). OBJECTIVE: We investigated changes in antioxidant concentrations and oxidative stress in plasma, buccal mucosal cells (BMCs), and breath condensate in patients with CF in relation to age and disease progression. DESIGN: We recruited 22 patients with CF as well as 35 healthy control subjects and conducted a cross-sectional study by dividing the participants into 4 age groups (<6 y, 6-11 y, 12-17 y, > or =18 y). We collected fasting blood samples, BMCs, and breath condensate. Carotenoids, alpha-tocopherol, vitamin C, protein carbonyls, thiobarbituric acid-reactive substances, and F(2)alpha-isoprostane were assessed. RESULTS: In patients with CF, plasma vitamin C concentrations, plasma and BMC alpha-tocopherol concentrations, and forced expiratory volume in 1 s (percentage predicted) decreased significantly with age. Plasma beta-carotene, beta-cryptoxanthin, and total lycopene were significantly lower in patients than in control subjects in all age groups. Furthermore, alpha-tocopherol and vitamin C plasma concentrations as well as alpha-tocopherol concentrations in BMCs were significantly lower in CF patients > or =18 y old, whereas all indicators of oxidative stress assessed were significantly higher than those same indicators in control subjects. CONCLUSIONS: Adult patients with CF in particular showed distinct vitamin deficits and elevated indicators of oxidative stress in plasma, BMCs, and breath condensate along with a progression of clinical status. We suggest that early in life dietary habits should be improved and that innovative supplementation strategies should be applied to optimize the antioxidant status of patients with CF. PMID- 15277157 TI - Prediction equations for resting energy expenditure in overweight and normal weight black and white children. AB - BACKGROUND: Accurate estimation of children's resting energy expenditure (REE) is important for planning dietary therapy. OBJECTIVE: Our objective was to compare the utility of 5 REE prediction equations in a diverse sample of young children. DESIGN: REE was obtained in 502 black and white girls and boys aged 6-11 y by using indirect calorimetry at 4 US sites. Measured REE and REE predicted from the equations were compared. RESULTS: None of the equations provided both accurate and unbiased estimates of REE. Two new sets of sex-specific equations including race as a factor were generated and evaluated. One set used easily measured variables-females: REE = 0.046 x weight - 4.492 x 1/height(2) - 0.151 x race + 5.841; males: REE = 0.037 x weight - 4.67 x 1/height(2) - 0.159 x race + 6.792 and accounted for 72% and 69%, respectively, of REE variance. The other set used body-composition variables-females: REE = 0.101 x fat-free mass + 0.025 x fat mass + 0.293 x height(3) - 0.185 x race + 1.643; males: REE = 0.078 x fat-free mass + 0.026 x fat mass - 2.646 x 1/height(2) - 0.244 x race + 4.8-and accounted for 75% and 71%, respectively, of REE variance. When split by race and adiposity, the small bias generated could be corrected to within 0.25 MJ (60 kcal) of the mean measured value. CONCLUSION: Sex-specific equations must take race into account to predict REE adequately in children. PMID- 15277159 TI - Metabolic adaptations to low zinc intakes in premenarcheal girls. AB - BACKGROUND: Zinc deficiency is increasingly recognized as an important cause of mortality and morbidity. Children in developing countries are at especially high risk because of relatively low zinc intakes and poor bioavailability. OBJECTIVE: We assessed the effect of 2-wk adaptation to low zinc intake (4 mg/d) on fractional zinc absorption, endogenous fecal zinc excretion, and urinary zinc excretion. DESIGN: Sixteen healthy 9-14-y-old girls were studied twice in random order after 2-wk adaptation to diets providing either 12 mg/d (high) or 4 mg/d (low) zinc. Fractional zinc absorption and endogenous fecal zinc excretion were measured with use of established stable isotope techniques. RESULTS: Plasma zinc was not significantly lower during the low dietary intake period (1.06 +/- 0.18 mg/L) than during the high dietary intake period (1.14 +/- 0.23 mg/L, P = 0.30). Endogenous fecal zinc excretion was significantly lower during the low intake period (1.08 +/- 0.62 mg/d) than during the high intake period (1.82 +/- 0.95 mg/d, P < 0.026), but there was no significant change in fractional zinc absorption (30.6% +/- 12.4% compared with 26.6% +/- 9.0%, P = 0.32) or urinary zinc excretion (0.68 +/- 0.35 mg/d compared with 0.59 +/- 0.24 mg/d, P = 0.30). Approximate zinc balance was significantly lower during the low-intake period than during the high-intake period (P = 0.007) and significantly (P < 0.0001) less than zero. CONCLUSION: Short-term zinc restriction in premenarcheal girls leads to a significant decrease in endogenous fecal zinc excretion, which was inadequate to restore normal zinc balance. PMID- 15277160 TI - Supplementation with cactus pear (Opuntia ficus-indica) fruit decreases oxidative stress in healthy humans: a comparative study with vitamin C. AB - BACKGROUND: Cactus pear (Opuntia ficus-indica) fruit contains vitamin C and characteristic betalain pigments, the radical-scavenging properties and antioxidant activities of which have been shown in vitro. OBJECTIVE: We investigated the effects of short-term supplementation with cactus pear fruit compared with vitamin C alone on total-body oxidative status in healthy humans. DESIGN: In a randomized, crossover, double-treatment study, 18 healthy volunteers received either 250 g fresh fruit pulp or 75 mg vitamin C twice daily for 2 wk, with a 6-wk washout period between the treatments. Before (baseline) and after each treatment, 8-epi-prostaglandin F(2alpha) (8-epi-PGF(2alpha)) and malondialdehyde in plasma, the ratio of reduced to oxidized glutathione (GSH:GSSG) in erythrocytes, and lipid hydroperoxides in LDL were measured as biomarkers of oxidative stress; plasma Trolox-equivalent antioxidant activity (TEAC) and vitamins A, E, and C were evaluated as indexes of antioxidant status. RESULTS: Both treatments caused comparable increases compared with baseline in plasma concentrations of vitamin E and vitamin C (P < 0.05); vitamin A and TEAC did not change significantly. After supplementation with cactus pear fruit, 8-epi PGF(2)alpha and malondialdehyde decreased by approximately 30% and 75%, respectively; GSH:GSSG shifted toward a higher value (P < 0.05); and LDL hydroperoxides were reduced by almost one-half. Supplementation with vitamin C did not significantly affect any marker of oxidative stress. CONCLUSIONS: Consumption of cactus pear fruit positively affects the body's redox balance, decreases oxidative damage to lipids, and improves antioxidant status in healthy humans. Supplementation with vitamin C at a comparable dosage enhances overall antioxidant defense but does not significantly affect body oxidative stress. Components of cactus pear fruit other than antioxidant vitamins may play a role in the observed effects. PMID- 15277161 TI - Carotenoid bioavailability is higher from salads ingested with full-fat than with fat-reduced salad dressings as measured with electrochemical detection. AB - BACKGROUND: The amount of dietary fat required for optimal bioavailability of carotenoids in plant matrices is not clearly defined. OBJECTIVE: The objective was to quantify the appearance of carotenoids in plasma chylomicrons after subjects ingested fresh vegetable salads with fat-free, reduced-fat, or full-fat salad dressings. DESIGN: The subjects (n = 7) each consumed 3 salads consisting of equivalent amounts of spinach, romaine lettuce, cherry tomatoes, and carrots with salad dressings containing 0, 6, or 28 g canola oil. The salads were consumed in random order separated by washout periods of > or =2 wk. Blood samples were collected hourly from 0 to 12 h. Chylomicrons were isolated by ultracentrifugation, and carotenoid absorption was analyzed by HPLC with coulometric array detection. RESULTS: After ingestion of the salads with fat-free salad dressing, the appearance of alpha-carotene, beta-carotene, and lycopene in chylomicrons was negligible. After ingestion of the salads with reduced-fat salad dressing, the appearance of the carotenoids in plasma chylomicrons increased relative to that after ingestion of the salads with fat-free salad dressing (P < 0.04). Similarly, the appearance of the carotenoids in plasma chylomicrons was higher after the ingestion of salads with full-fat than with reduced-fat salad dressing (P < 0.02). CONCLUSIONS: High-sensitivity HPLC with coulometric array detection enabled us to quantify the intestinal absorption of carotenoids ingested from a single vegetable salad. Essentially no absorption of carotenoids was observed when salads with fat-free salad dressing were consumed. A substantially greater absorption of carotenoids was observed when salads were consumed with full-fat than with reduced-fat salad dressing. PMID- 15277162 TI - Calcium from milk or calcium-fortified foods does not inhibit nonheme-iron absorption from a whole diet consumed over a 4-d period. AB - BACKGROUND: Single-meal studies have indicated that calcium inhibits iron absorption in humans. However, numerous dietary factors influence iron absorption, and the effect of calcium may not be as pronounced when calcium is served as part of a whole diet. OBJECTIVE: We investigated the effect of 3 sources of calcium served with the 3 main meals on nonheme-iron absorption from a 4-d diet. DESIGN: (59)Fe absorption was estimated from whole-body retention measurements in 14 women aged 21-34 y, each of whom consumed four 4-d diets in a randomized crossover design. The diets differed in the source of calcium as follows: a basic diet (BD) with a low content of calcium (224 mg Ca/d), a BD with a glass of milk served at each meal (826 mg Ca/d), a BD with calcium lactate (802 mg Ca/d), and a BD with a milk mineral isolate containing calcium (801 mg Ca/d). The 2 latter calcium sources were added to selected foods of the BD (rye bread, white bread, chocolate cake, and orange juice), and these foods were consumed with the 3 meals. All diets provided 13.2 mg Fe/d. RESULTS: No significant differences in nonheme-iron absorption were found between the BD and the BD supplemented with milk, calcium lactate, or the milk mineral isolate [7.4% (95% CI: 5.3%, 10.5%), 5.2% (3.5%, 7.9%), 6.7% (5.0%, 8.9%), and 5.1% (3.2%, 7.9%), respectively; P = 0.34]. CONCLUSION: Consumption of a glass of milk with the 3 main meals or of an equivalent amount of calcium from fortified foods does not decrease nonheme-iron absorption from a 4-d diet. PMID- 15277163 TI - Copper, selenium, zinc, and thiamine balances during continuous venovenous hemodiafiltration in critically ill patients. AB - BACKGROUND: Acute renal failure is a serious complication in critically ill patients and frequently requires renal replacement therapy, which alters trace element and vitamin metabolism. OBJECTIVE: The objective was to study trace element balances during continuous renal replacement therapy (CRRT) in intensive care patients. DESIGN: In a prospective randomized crossover trial, patients with acute renal failure received CRRT with either sodium bicarbonate (Bic) or sodium lactate (Lac) as a buffering agent over 2 consecutive 24-h periods. Copper, selenium, zinc, and thiamine were measured with highly sensitive analytic methods in plasma, replacement solutions, and effluent during 8-h periods. Balances were calculated as the difference between fluids administered and effluent losses and were compared with the recommended intakes (RI) from parenteral nutrition. RESULTS: Nineteen sessions were conducted in 11 patients aged 65 +/- 10 y. Baseline plasma concentrations of copper were normal, whereas those of selenium and zinc were below reference ranges; glutathione peroxidase was in the lower range of normal. The replacement solutions contained no detectable copper, 0.01 micromol Se/L (Bic and Lac), and 1.42 (Bic) and 0.85 (Lac) micromol Zn/L. Micronutrients were detectable in all effluents, and losses were stable in each patient; no significant differences were found between the Bic and Lac groups. The 24-h balances were negative for selenium (-0.97 micromol, or 2 times the daily RI), copper (-6.54 micromol, or 0.3 times the daily RI), and thiamine ( 4.12 mg, or 1.5 times the RI) and modestly positive for zinc (20.7 micromol, or 0.2 times the RI). CONCLUSIONS: CRRT results in significant losses and negative balances of selenium, copper, and thiamine, which contribute to low plasma concentrations. Prolonged CRRT is likely to result in selenium and thiamine depletion despite supplementation at recommended amounts. PMID- 15277164 TI - Calcium homeostasis during pregnancy and lactation in Brazilian women with low calcium intakes: a longitudinal study. AB - BACKGROUND: Physiologic adjustments in calcium homeostasis during pregnancy and lactation in women with marginal calcium intakes have not been described. OBJECTIVE: The objective was to examine longitudinal changes in various aspects of calcium homeostasis during pregnancy and lactation in 9 healthy Brazilian women who habitually consumed approximately 500 mg Ca/d. DESIGN: Calcium homeostasis was assessed at 3 time points: 10-12 (early pregnancy, EP) and 34-36 (late pregnancy, LP) wk of pregnancy and 7-8 wk postpartum (early lactation, EL). At each time point, the following variables were measured: dietary calcium intake with a 3-d weighed food record, 24-h urinary calcium excretion (UCa), intestinal calcium absorption (%CaAbs) via administration of stable calcium isotopes with a breakfast meal, serum 1,25-dihydroxyvitamin D, parathyroid hormone (PTH), insulin like growth factor I (IGF-I), and biochemical markers of bone turnover. RESULTS: Dietary calcium did not change during the study. %CaAbs increased from 69.7 +/- 5.4% ( +/- SEM) during EP to 87.6 +/- 4.5% during LP (P < 0.05) and returned to 65.1 +/- 6.2% during EL. Compared with EP, UCa decreased 22% during LP and 68% during EL (P < 0.05). The net mean change in calcium retention was 212 mg/d during LP and 182 mg/d during EL. Several significant associations were found between the main outcome variables (%CaAbs, UCa, and markers of bone turnover) and serum hormones, especially IGF-I and PTH. CONCLUSIONS: Calcium homeostasis appears to be attained by a more efficient intestinal calcium absorption during pregnancy and by renal calcium conservation during both pregnancy and lactation. IGF-I and PTH seem to play major roles in the adjustment of calcium metabolism during pregnancy and lactation. PMID- 15277165 TI - Lactation, weaning, and calcium supplementation: effects on body composition in postpartum women. AB - BACKGROUND: Concern that long-term weight retention after pregnancy contributes to obesity underscores the need to identify factors that facilitate postpartum weight loss. Lactation is believed to facilitate postpartum weight loss and fat loss. Calcium intake also has been hypothesized to promote weight loss and fat loss. OBJECTIVE: We addressed the following questions: 1) whether lactation enhances loss of fat mass, and 2) whether loss of fat mass during lactation and after weaning is greater in women receiving calcium supplementation than in women receiving placebo. DESIGN: We used data from 87 lactating and 81 nonlactating women enrolled in a randomized, double-blind, calcium supplementation trial from 2 wk to 6 mo postpartum and data from 76 previously lactating and 82 nonlactating women enrolled in a parallel trial from 6 to 12 mo postpartum. Body fat and lean masses were measured by using dual-energy X-ray absorptiometry. RESULTS: Nonlactating women lost whole-body, arm, and leg fat at a faster rate than did lactating women between 2 wk and 6 mo postpartum (lactation group x time effect, P < or = 0.01). Fat mass of the trunk, arms, and legs decreased between 6 and 12 mo postpartum regardless of previous lactation status (time effect, P < or = 0.001). Calcium supplementation did not affect postpartum fat loss. CONCLUSIONS: Body-composition changes occur differently in nonlactating and lactating women during the first 6 mo postpartum and occur at some sites until 12 mo postpartum regardless of previous lactation status. Clinicians should use caution when advising lactating mothers about expected rates of postpartum fat loss. Calcium supplementation (1 g/d) does not promote postpartum weight loss or fat loss. PMID- 15277166 TI - Sex differences in resting energy expenditure and their relation to insulin resistance in children (EarlyBird 13). AB - BACKGROUND: Insulin resistance is believed to be the process underlying type 2 diabetes and premature cardiovascular disease. We have established that a relation between body mass and insulin resistance calculated by homeostasis model assessment (HOMA-IR) exists by 5 y of age in contemporary UK children. Resting energy expenditure (REE) is variable among individuals and is one of many factors controlling body mass. OBJECTIVE: The objective was to investigate the relations between REE, body mass, and HOMA-IR in young children. DESIGN: EarlyBird is a nonintervention prospective cohort study of 307 healthy 5-y-olds that asks the question: Which children develop insulin resistance and why? REE by indirect calorimetry and HOMA-IR were measured in addition to total body mass, fat-free mass (FFM) by bioimpedance, body mass index (BMI; in kg/m(2)), and skinfold thickness when the mean age of the cohort was 5.9 +/- 0.2 y. RESULTS: Whereas the BMI of the boys was lower than that of the girls (x +/- SD: boys, 15.9 +/- 1.9; girls, 16.5 +/- 1.9; P = 0.03), their REE was higher by 6% (x +/- SD: 4724 +/- 615 compared with 4469 +/- 531 kJ/d; P = 0.002). This difference persisted after adjustment for FFM and other anthropometric variables (P = 0.04). In boys, there was a weak, although significant, inverse correlation between REE and HOMA-IR, independent of fat mass and FFM (boys: r = -0.21, P = 0.03; girls: r = 0.12, P = 0.34). CONCLUSION: There is a sex difference in REE at 6 y of age that cannot be explained by body composition. The difference appears to be intrinsic, and its contribution to sex differences in adiposity and HOMA-IR in children merits further exploration. PMID- 15277167 TI - Prematurity and reduced body fatness at 8-12 y of age. AB - BACKGROUND: Increasing evidence suggests that body composition, an important factor influencing morbidity and mortality in adult life, might be programmed by early growth and nutrition. Children born preterm remain shorter and lighter than their term-born peers during childhood, but it is unclear whether the size difference is associated with altered body composition. OBJECTIVE: We tested the hypothesis that both fat mass (FM) and fat-free mass (FFM) are proportionately lower in children born preterm than in children born at term. DESIGN: A total of 497 children born preterm and 95 children born at term were studied at 8-12 y of age. Body composition was determined with the use of skinfold thicknesses and dual-energy X-ray absorptiometry (in 200 preterm and 95 term children). FM and FFM were normalized for height to give fat mass index (FMI) and fat-free mass index (FFMI), respectively. RESULTS: Children born preterm were significantly lighter than those born at term and had lower FM and FFM. However, FMI was significantly lower in preterm children, whereas FFMI was not. FMI was also significantly lower in boys than in girls and in children with higher activity levels. Additional data available for the preterm group showed no association between birth weight, gestational age, or neonatal diet and later FMI or FFMI. CONCLUSIONS: The smaller size of children born preterm than of children born at term is associated with lower FM but not FFM when normalized for height. We hypothesize that this could result in a reduction in the risk of obesity and related diseases during adult life. PMID- 15277168 TI - Fifty-year trends in serial body mass index during adolescence in girls: the Fels Longitudinal Study. AB - BACKGROUND: A decline in the age at menarche was recently reported for US girls. Although it is possible that this recent drop stems from the concurrent increase in childhood obesity, few longitudinal studies of growth and development have been undertaken to specifically address the temporal relation between growth, adiposity, and the age at menarche. OBJECTIVE: The objective was to simultaneously examine the effects of birth cohort (secular trend) and rate of maturation (age at menarche) on the timing and pattern of increases in body mass index (BMI) during adolescence in girls. DESIGN: We applied mixed-effects polynomial models to serial BMI data, spanning from 6 y before menarche to 6 y after menarche, obtained from 211 girls enrolled in the Fels Longitudinal Study. We examined the effects of birth cohort (defined as girls born 1929-1946, 1947 1964, and 1965-1983) and age at menarche (defined as < or =11.9 y, 12.0-13.1 y, and > or =13.2 y) on the magnitude and velocity of BMI during adolescence. RESULTS: BMI and BMI velocity in girls born after 1965 were significantly greater than those of girls of earlier birth cohorts, despite stability in the mean age at menarche. Although girls with early menarche tended to have significantly higher BMIs than did girls with average or later menarche, these differences did not emerge until after menarche. CONCLUSION: These data suggest that increases in relative weight are a consequence, rather than a determinant, of the age at menarche and that secular changes in BMI and in the mean age at menarche could be independent phenomena. PMID- 15277169 TI - Animal protein intake, serum insulin-like growth factor I, and growth in healthy 2.5-y-old Danish children. AB - BACKGROUND: Studies from developing countries indicate that intake of animal protein, especially of milk, is associated with greater velocity of linear growth in childhood. Whether the same association exists in industrialized countries, where protein intake is high, is not clear. OBJECTIVE: Our objective was to examine associations between protein intake, serum insulin-like growth factor I (sIGF-I) concentrations, and height in healthy children. DESIGN: We analyzed the associations between protein intake, sIGF-I concentrations, and height in 2.5-y old children. Diet (7-d record) and sIGF-I (radioimmunoassay) data were available from 90 children (54 boys). RESULTS: The 10th, 50th, and 90th percentiles of protein intake were 2.4, 2.9, and 4.0 g. kg(-1). d(-1), respectively; 63% was animal protein. In multiple linear regressions with adjustment for sex and weight, height (cm) was positively associated with intakes of animal protein (g/d) [0.10 +/- 0.038 (b +/- SE); P = 0.01] and milk (0.0047 +/- 0.002; P = 0.007), but not with those of vegetable protein or meat. The sIGF-I concentration was significantly associated with intakes of animal protein (1.4 +/- 0.53; P = 0.01) and milk (0.049 +/- 0.024; P = 0.045), but not with those of vegetable protein or meat. sIGF-I concentrations were positively associated with height (0.019 +/- 0.008; P = 0.02). CONCLUSION: Milk intake was positively associated with sIGF-I concentrations and height. An increase in milk intake from 200 to 600 mL/d corresponded to a 30% increase in circulating IGF-I. This suggests that milk compounds have a stimulating effect on sIGF-I concentrations and, thereby, on growth. PMID- 15277170 TI - Birth weight predicts response to vaccination in adults born in an urban slum in Lahore, Pakistan. AB - BACKGROUND: Substantial evidence exists linking small size at birth to later-life susceptibility to chronic disease. Evidence is also emerging that some components of immune function may be programmed in early life. However, this evidence is limited and requires confirmation. OBJECTIVE: We investigated the association between size at birth and response to vaccination in a cohort of 257 adults (mean age: 29.4 y; 146 men) born in an urban slum in Lahore, Pakistan, during 1964 1978. DESIGN: A single dose of Vi polysaccharide vaccine for Salmonella typhi and 2 doses of rabies vaccine were given to each subject. Antibody titers were measured in prevaccination serum samples (Vi) and in postvaccination samples (Vi and rabies). RESULTS: The mean birth weight of the subjects was 3.24 kg; 14% of the subjects had low birth weights (<2.5 kg). Vaccine responses were not consistently associated with contemporary variables (month of study, sex, current age, or indicators of wealth). Response to typhoid vaccination was positively related to birth weight (anti-Vi immunoglobulin G: r = 0.138, P = 0.031; anti-Vi immunoglobulin M: r = 0.197, P = 0.034). Response to the rabies vaccine was not significantly associated with birth weight. CONCLUSIONS: These findings add to a growing body of evidence suggesting that components of the immune system may be permanently programmed by events in early life. The contrasting effects on typhoid and rabies responses suggest that antibody generation to polysaccharide antigens, which have greater B cell involvement, is compromised by fetal growth retardation. PMID- 15277171 TI - Randomized controlled trial of nutritional supplementation in patients with newly diagnosed tuberculosis and wasting. AB - BACKGROUND: Nutritional support is often recommended as part of the treatment of tuberculosis, but it has never been properly tested. OBJECTIVE: We assessed the effects of early nutritional intervention on lean mass and physical function in patients with tuberculosis and wasting. DESIGN: Patients who started antituberculous therapy within the previous 2 wk were randomly assigned to receive standard nutritional counseling (control group) or nutritional counseling to increase their intake through diet and high-energy supplements (nutritional supplement group) for 6 wk. Body composition was measured by dual-energy X-ray absorptiometry, and physical function was assessed by maximum grip strength. RESULTS: Patients in the nutritional supplement group (n = 19) had a significantly greater increase in body weight (2.57 +/- 1.78 compared with 0.84 +/- 0.89 kg, P = 0.001), total lean mass (1.17 +/- 0.93 compared with 0.04 +/- 1.26 kg, P = 0.006), and grip strength (2.79 +/- 3.11 compared with -0.65 +/- 4.48 kg, P = 0.016) than did the control subjects (n = 17) at week 6. During subsequent follow-up, the increase in body weight remained greater in the nutritional supplement group, but this increase was due mainly to a greater gain in fat mass in the nutritional supplement group than in the control group. CONCLUSIONS: Early intervention to increase nutritional intake increases lean mass and physical function. This adjunct to tuberculosis therapy could confer socioeconomic and survival benefits that deserve investigation in large-scale trials. Nutritional intervention after the initial phase of treatment could be less beneficial because it mainly increases fat. PMID- 15277172 TI - Weight-loss intention in the well-functioning, community-dwelling elderly: associations with diet quality, physical activity, and weight change. AB - BACKGROUND: Many older adults desire to lose weight, yet the proportion with a health-related weight-loss indication, weight-loss strategies, and success is unknown. OBJECTIVE: We examined the associations of reported intention to lose weight with health-related indications for weight loss, diet quality, physical activity, and weight-loss success in well-functioning older adults. DESIGN: This prospective, community-based cohort included 2708 elderly persons aged 70-79 y at baseline. We determined indication for weight loss by using the modified National Institutes of Health guidelines, diet quality by using the Healthy Eating Index, and weight-loss intention and physical activity by using questionnaires. Measured weight change over 1 y was assessed. RESULTS: Twenty-seven percent of participants reported an intention to lose weight, and 67% of those participants had an indication for weight loss. Participants who reported a weight-loss intention were heavier than those who did not, had more depressive symptoms, and were more likely to be dissatisfied with their weight, regardless of weight-loss indication. Participants with an intention to lose weight reported better eating behaviors and a more active lifestyle than did participants without a weight-loss intention, independent of other health conditions. No significant difference in actual weight loss was found between participants intending and not intending to lose weight, regardless of indication for weight loss. CONCLUSIONS: Despite being associated with healthier behaviors, the intention to lose weight did not predict greater weight loss in this well-functioning elderly cohort. More attention needs to be focused on the necessity and efficacy of specific strategies for weight loss in older adults. PMID- 15277173 TI - Anthropometric assessment of 10-y changes in body composition in the elderly. AB - BACKGROUND: An increased central distribution of fat with advancing age is associated with chronic metabolic and cardiovascular abnormalities. Little is known about the magnitude or pattern of fat distribution and its association with healthy aging. OBJECTIVE: This study describes approximately 10-y changes in body composition at 11 anthropometric sites in elderly persons and the metabolic and physical activity factors associated with these changes. DESIGN: Skinfold thicknesses, girths, body fat by hydrodensitometry, physical activity by questionnaire, and metabolic variables were examined twice, 9.4 +/- 1.4 y apart, in 54 men and 75 women aged 60.4 +/- 7.8 y at baseline. RESULTS: Subcutaneous fat declined (-17.2%; P < 0.001), whereas total fat mass increased (7.2%; P < 0.05). Waist and hip circumference changes were the best anthropometric predictors of total fat mass change (r(2) = 0.40-0.65, P < 0.0001). Thigh girth change was more strongly associated with fat-free mass change (r(2) = 0.22, P < 0.01) than with fat mass change (r(2) = 0.07, P < 0.05) in women. An increase in physical activity was associated with an attenuation of thigh girth decline in men and women (F ratio = 5.13, P < 0.007). Traditional metabolic markers of visceral adiposity (triacylglycerol, glucose, and total cholesterol) were not significantly related to the change in waist circumference. CONCLUSIONS: Skinfold thicknesses cannot be used to assess changes in body fat mass because of age related fat redistribution. Higher levels of physical activity can attenuate the decline in appendicular lean tissue expected over 10 y. Waist and thigh girths, rather than skinfold thicknesses, should be considered for use in longitudinal studies in the elderly because the changes in these girths capture increased abdominal adiposity and sarcopenia, respectively. PMID- 15277174 TI - Plasma amino acid concentrations in patients with amnestic mild cognitive impairment or Alzheimer disease. AB - BACKGROUND: Plasma concentrations of several amino acids may affect the availability of important neurotransmitter precursors in the brain. Abnormalities in the plasma amino acid profile have been reported in elderly persons with cognitive impairment, but no data exist for the prodromal phase of Alzheimer disease (AD), which is characterized by amnestic mild cognitive impairment (aMCI). OBJECTIVE: The objective was to investigate whether the plasma amino acid profiles of elderly patients with aMCI or AD are abnormal. DESIGN: The plasma amino acid profile was assessed in 29 cognitively normal control subjects (age: 86.7 +/- 5.9 y), 21 patients with aMCI (age: 84.9 +/- 7.0 y), and 51 patients with AD (age: 86.7 +/- 5.4 y). The participants were from the University of Bologna Research Center for Physiopathology of Aging, Italy. RESULTS: Higher plasma concentrations of the aromatic amino acid phenylalanine were found in the aMCI (68 micromol/L; 95% CI: 63, 73) and AD (62 micromol/L; 95% CI: 59, 65) patients than in the control subjects (54 micromol/L; 95% CI: 48, 61; P < 0.05). The ratio of arginine to other basic amino acids was also higher in the aMCI (0.31 +/- 0.04) and AD (0.27 +/- 0.08) patients than in the control subjects (0.21 +/- 0.05; P < 0.05). Adjustment for differences in body composition, serum vitamin B-12 concentrations, and serum folate concentrations did not significantly affect the results. CONCLUSIONS: The plasma amino acid profiles of elderly patients with aMCI or AD show abnormalities in aromatic and basic amino acids that potentially affect neurotransmitter biosynthesis. PMID- 15277175 TI - Dietary glycemic load and risk of age-related cataract. AB - BACKGROUND: Metabolism of many of the most commonly consumed carbohydrates in the United States results in a high plasma glucose response, which can be quantified by the glycemic load. Although hyperglycemia is a risk factor for cataract, there is no information on the potential effect of a high dietary glycemic load on the incidence of age-related cataract. OBJECTIVE: Our objective was to prospectively examine the association between dietary glycemic load and incident age-related cataract. DESIGN: We studied 2 cohorts-71 919 women and 39 926 men-aged > or =45 y who had no previous diagnosis of cataract, diabetes mellitus, or cancer and who were followed for 14 and 12 y, respectively, for the occurrence of cataract extraction. We calculated dietary glycemic load from data reported on multiple validated food-frequency questionnaires and used pooled logistic regression models to estimate the association with incident cataract extraction. We performed analyses separately for each cohort and then calculated pooled estimates across cohorts. RESULTS: During 980 683 person-years of follow-up, we confirmed 4865 incident age-related cataract extractions. After adjustment for age, cigarette smoking, body mass index, total caloric intake, dietary intake of lutein and zeaxanthin, and alcohol consumption, there was no significant relation of dietary glycemic load to risk of cataract extraction (P for trend = 0.10). The pooled relative risk between the highest and lowest quintiles of dietary glycemic load was 0.95 (95% CI: 0.81, 1.11; P for heterogeneity by cohort = 0.1). CONCLUSION: These prospective epidemiologic data do not support the hypothesis that a high dietary glycemic load, primarily a result of consumption of refined carbohydrates, increases the risk of cataract extraction. PMID- 15277176 TI - Hormonal and lifestyle determinants of appendicular skeletal muscle mass in men: the MINOS study. AB - BACKGROUND: Aging-related sarcopenia is characterized by a loss of muscle mass and strength and increased fatigability. However, studies of its determinants in elderly men are scarce. OBJECTIVE: We investigated risk factors for sarcopenia in a large cohort of men. DESIGN: We analyzed 845 men aged 45-85 y who belonged to the MINOS cohort. Lifestyle factors (physical activity, tobacco smoking, alcohol intake, caffeine intake) were evaluated by using a standardized questionnaire. Appendicular skeletal muscle mass (ASM) was estimated by using dual-energy X-ray absorptiometry. The relative appendicular skeletal muscle mass index (RASM) was calculated as ASM/body height(2.3). Apparent free testosterone concentration (AFTC) and free testosterone index (FTI) were calculated on the basis of concentrations of total testosterone and sex hormone-binding globulin. RESULTS: RASM decreased with age (r = -0.29, P < 0.0001). Current smokers had lower RASM than did subjects who never smoked (-3.2%; P < 0.003). RASM increased with the intensity of physical activity at work (P for trend < 0.001). Men who participated in regular exercise during leisure time had 2.2% higher RASM than did those who did not (P < 0.03). Men whose values for AFTC, FTI, or 25 hydroxycholecalciferol [25(OH)D] were >2 SDs below the mean for young men had significantly lower RASM than did men with higher values. Men with sarcopenia, defined as the lowest quartile of RASM in the studied cohort (<6.32 kg/m(2.3)), were significantly older than men with normal RASM, weighed significantly less, smoked more, and spent significantly less time on leisure-time activities. Sarcopenic men also had lower values for testosterone, AFTC, FTI, and 25(OH)D. CONCLUSION: In elderly men, low physical activity, tobacco smoking, thinness, low testosterone (AFTC and FTI), and decreased 25(OH)D concentrations are risk factors for sarcopenia. PMID- 15277177 TI - Food patterns measured by factor analysis and anthropometric changes in adults. AB - BACKGROUND: Sixty-five percent of US adults are overweight, and 31% of these adults are obese. Obesity results from weight gains over time; however, dietary determinants of weight gain remain controversial. OBJECTIVE: Our objective was to examine whether food patterns derived from exploratory factor analysis are related to anthropometric changes. We hypothesized that we would derive a healthy food pattern and that it would predict smaller changes in body mass index (BMI; in kg/m(2)) and waist circumference (in cm) than would other food patterns in models adjusted for baseline anthropometric measures. DESIGN: The subjects were 459 healthy men and women participating in the Baltimore Longitudinal Study of Aging. Diet was assessed by using 7-d dietary records, from which 40 food groups were formed and entered into a factor analysis. RESULTS: Six food patterns were derived. Factor 1 (reduced-fat dairy products, fruit, and fiber) was most strongly associated with fiber (r = 0.39) and loaded heavily on reduced-fat dairy products, cereal, and fruit and loaded moderately on fruit juice, nonwhite bread, nuts and seeds, whole grains, and beans and legumes. In a multivariate-adjusted model in which the highest and lowest quintiles were compared, factor 1 was inversely associated with annual change in BMI (beta = -0.51; 95% CI: -0.82, 0.20; P < 0.05; P for trend < 0.01) in women and inversely associated with annual change in waist circumference (beta = -1.06 cm; 95% CI: -1.88, -0.24 cm; P < 0.05; P for trend = 0.04) in both sexes. CONCLUSION: Our results suggest that a pattern rich in reduced-fat dairy products and high-fiber foods may lead to smaller gains in BMI in women and smaller gains in waist circumference in both women and men. PMID- 15277178 TI - Obesity during childhood and adolescence augments bone mass and bone dimensions. AB - BACKGROUND: Studies of the effect of childhood obesity on bone accrual during growth have yielded conflicting results, largely related to the failure to adequately characterize the confounding effects of growth, maturation, and body composition. OBJECTIVE: The objective of this study was to determine the effect of childhood obesity on skeletal mass and dimensions relative to height, body composition, and maturation in males and females. DESIGN: In 132 nonobese (body mass index < 85th percentile) and 103 obese (body mass index > or = 95th percentile) subjects aged 4-20 y, whole-body and vertebral bone mineral content (BMC) was determined by using dual-energy X-ray absorptiometry, and bone area, areal bone mineral density (BMD), and fat and lean masses were measured. Vertebral volumetric BMD was estimated as BMC/area(1.5). RESULTS: Obesity was associated with greater height-for-age, advanced maturation for age, and greater lean mass for height (all P < 0.001). Sex-specific multivariate regressions with adjustment for maturation showed that obesity was associated with greater vertebral areal BMD for height, greater volumetric BMD, and greater vertebral BMC for bone area (all P < 0.05). After adjustment for maturation and lean mass, obesity was associated with significantly greater whole-body bone area and BMC for age and for height (all P < 0.001). CONCLUSIONS: In contrast with the results of prior studies, obesity during childhood and adolescence was associated with increased vertebral bone density and increased whole-body bone dimensions and mass. These differences persisted after adjustment for obesity-related increases in height, maturation, and lean mass. Future studies are needed to determine the effect of these differences on fracture risk. PMID- 15277179 TI - Proportion of the US population whose intakes of added sugars exceed the suggested maximum in the dietary reference intakes. PMID- 15277181 TI - Maternal nutrition and fetal growth: bias introduced because of an inappropriate statistical modeling strategy may explain null findings. PMID- 15277183 TI - Primary role of sweeteners in the body mass indexes of women from developing countries: implications for risk of chronic disease. PMID- 15277184 TI - Western soy intake is too low to produce health effects. PMID- 15277186 TI - From bedside to bench to bedside: role of renin-angiotensin-aldosterone system in remodeling of resistance arteries in hypertension. PMID- 15277188 TI - The powerful microbubble: from bench to bedside, from intravascular indicator to therapeutic delivery system, and beyond. AB - This review discusses the development, current applications, and therapeutic potential of ultrasound contrast agents. Microbubbles containing gases act as true, intravascular indicators, permitting a noninvasive, quantitative analysis of the spatial and temporal heterogeneity of blood flow and volumes within the microvasculature. These shelled microbubbles are near-perfect reflectors of acoustic ultrasound energy and, when injected intravenously into the bloodstream, reflect ultrasound waves within the capillaries without disrupting the local environment. Accordingly, microbubble ultrasound contrast agents are clinically useful in enhancing ultrasound images and improving the accuracy of diagnoses. More recently, ultrasound contrast agents have been used to directly visualize the vasa vasorum and neovascularization of atherosclerotic carotid artery plaques, thus suggesting a new paradigm for diagnosis and treatment of atherosclerosis. Future applications of these microscopic agents include the deliver of site-specific therapy to targeted organs in the body. Medical therapies may use these microbubbles as carriers for newer therapeutic options. PMID- 15277189 TI - Cellular approaches to tissue repair in cardiovascular disease: the more we know, the more there is to learn. PMID- 15277190 TI - Stable benefit of embryonic stem cell therapy in myocardial infarction. AB - Conventional therapies for myocardial infarction attenuate disease progression without contributing significantly to repair. Because of the capacity for de novo cardiogenesis, embryonic stem cells are considered a potential source for myocardial regeneration, yet limited information is available on their ultimate therapeutic value. We treated infarcted rat hearts with CGR8 embryonic stem cells preexamined for cardiogenicity, serially probed left ventricular function, and determined final pathological outcome. Stem cell delivery generated new cardiomyocytes of embryonic stem cell origin that integrated with host myocardium within infarct regions. This resulted in a functional benefit within 3 wk that remained sustained over 12 wk of continuous follow-up and included a vigorous inotropic response to beta-adrenergic challenge. Integration of stem cell-derived cardiomyocytes was associated with normalized ventricular architecture, little scar, and a decrease in signs of myocardial necrosis. In contrast, sham-treated infarcted hearts exhibited ventricular cavity dilation and aneurysm formation, poor ventricular function, and a lack of response to beta-adrenergic stimulation. No evidence of graft rejection, ectopy, sudden cardiac death, or tumor formation was observed after therapy. These findings indicate that embryonic stem cells, through differentiation within the host myocardium, can contribute to a stable beneficial outcome on contractile function and ventricular remodeling in the infarcted heart. PMID- 15277191 TI - Tissue-engineered microvessels on three-dimensional biodegradable scaffolds using human endothelial progenitor cells. AB - Tissue engineering may offer patients new options when replacement or repair of an organ is needed. However, most tissues will require a microvascular network to supply oxygen and nutrients. One strategy for creating a microvascular network would be promotion of vasculogenesis in situ by seeding vascular progenitor cells within the biopolymeric construct. To pursue this strategy, we isolated CD34(+)/CD133(+) endothelial progenitor cells (EPC) from human umbilical cord blood and expanded the cells ex vivo as EPC-derived endothelial cells (EC). The EPC lost expression of the stem cell marker CD133 but continued to express the endothelial markers KDR/VEGF-R2, VE-cadherin, CD31, von Willebrand factor, and E selectin. The cells were also shown to mediate calcium-dependent adhesion of HL 60 cells, a human promyelocytic leukemia cell line, providing evidence for a proinflammatory endothelial phenotype. The EPC-derived EC maintained this endothelial phenotype when expanded in roller bottles and subsequently seeded on polyglycolic acid-poly-l-lactic acid (PGA-PLLA) scaffolds, but microvessel formation was not observed. In contrast, EPC-derived EC seeded with human smooth muscle cells formed capillary-like structures throughout the scaffold (76.5 +/- 35 microvessels/mm(2)). These results indicate that 1) EPC-derived EC can be expanded in vitro and seeded on biodegradable scaffolds with preservation of endothelial phenotype and 2) EPC-derived EC seeded with human smooth muscle cells form microvessels on porous PGA-PLLA scaffolds. These properties indicate that EPC may be well suited for creating microvascular networks within tissue engineered constructs. PMID- 15277192 TI - Autologous vascular smooth muscle cell-based myocardial gene therapy to induce coronary collateral growth. AB - For therapeutic angiogenesis to achieve clinical relevance, it must be effective, with minimal side effects to other end organ systems. We developed a cardiac specific gene delivery mechanism by transfecting autologous vascular smooth muscle cells (VSMC) with VEGF and administering these cells via intracoronary injection. We evaluated the efficacy of this protocol by its ability to stimulate angiogenesis in the presence of a subthreshold stimulus for collateralization. A modified canine repetitive coronary occlusion model was utilized in these experiments with left anterior descending coronary artery occlusions for 2 min every 2 h four times per day for 21 days. An intramyocardial catheter in the perfusion territory of the left anterior descending coronary artery measured proteins in the myocardial interstitial fluid. VSMC from jugular vein explants were isolated, amplified in culture for 3 wk, and transfected with a plasmid expressing VEGF-165 and/or enhanced green fluorescent protein. Cells were injected before commencement of occlusions. VEGF levels in myocardial interstitial fluid were significantly higher in VEGF-transfected animals than in sham (repetitive occlusions without cell transplantation) and control (repetitive occlusions with enhanced green fluorescent protein-transfected cells) animals at the onset of occlusions (P < 0.05). In the VEGF group, collateral flow was increased at day 7 and remained higher than in sham and control groups thereafter. We found that intracoronary administration of VEGF-transfected autologous VSMC effectively promotes collateral development. This approach may provide a way to confine delivery of a gene to a specified organ, thus minimizing complications related to gene transfection in nontargeted organ systems. PMID- 15277193 TI - Use of blood outgrowth endothelial cells as virus-producing vectors for gene delivery to tumors. AB - Cell-based delivery of therapeutic viruses has potential advantages over systemic viral administration, including attenuated neutralization and improved viral targeting. One of the exciting new areas of investigation is the potential ability of endothelial-lineage cells to deliver genes to the areas of neovascularization. In the present study, we compared two types of endothelial lineage cells [outgrowth endothelial cells (OECs) and culture-modified mononuclear cells (CMMCs), also known as "endothelial progenitor cells"] for their ability to be infected with adenovirus and to home to the areas of neovascularization. Both cell types were isolated from peripheral blood of healthy human donors and expanded in culture. We demonstrate that OECs are more infectable and home better to tumors expressing VEGF on systemic administration. Furthermore, we used an adenoviral/retroviral chimeric system to convert OECs to retrovirus-producing cells. When injected systemically into tumor-bearing mice, OECs retain their ability to produce retrovirus and infect surrounding tumor cells. Our data demonstrate that OECs could be efficient carriers for viral delivery to areas of tumor neovascularization. PMID- 15277194 TI - Autologous stem cell transplantation for myocardial repair. AB - Current therapies for heart failure due to transmural left ventricular (LV) infarction are limited. We have developed a novel patch method for delivering autologous bone marrow stem cells to sites of myocardial infarction for the purpose of improving LV function and preventing LV aneurysm formation. The patch consisted of a fibrin matrix seeded with autologous porcine mesenchymal stem cells labeled with lacZ. We applied this patch to a swine model of postinfarction LV remodeling. Myocardial infarction was produced by using a 60-min occlusion of the left anterior descending coronary artery distal to the first diagonal branch followed by reperfusion. Results were compared between eight pigs with stem cell patch transplantation, six pigs with the patch but no stem cells (P), and six pigs with left anterior descending coronary artery ligation alone (L). Magnetic resonance imaging data collected 19 +/- 1 days after the myocardial infarction indicated a significant increase of LV systolic wall thickening fraction in the infarct zone of transplanted hearts compared with P or L hearts. Blue X-gal staining was observed in the infarcted area of transplanted hearts. PCR amplification of specimens from the X-gal-positive area revealed the Ad5 RSV-lacZ vector fragment DNA sequence. Light microscopy demonstrated that transplanted cells had differentiated into cells with myocyte-like characteristics and a robust increase of neovascularization as evidenced by von Willebrand factor positive angioblasts and capillaries in transplanted hearts. Thus this patch based autologous stem cell procedure may serve as a therapeutic modality for myocardial repair. PMID- 15277195 TI - c-kit-immunopositive vascular progenitor cells populate human coronary in-stent restenosis but not primary atherosclerotic lesions. AB - Progress in the treatment of human in-stent restenosis (ISR) is hampered by an imprecise understanding of the nature of the cells that occlude vascular stents. Recent studies suggest that circulating vascular progenitor cells may mediate vascular repair and lesion formation. Moreover, functional endothelial progenitor cells appear to play a protective role in attenuating vascular lesion formation. Hence, we sought to answer two important questions: 1). Are primitive cells found in ISR lesions? 2). Is the abundance of cultured angiogenic cells (CACs) in patients with ISR different from that in patients with non-ISR lesions or normal controls? Human coronary atherectomy tissue from 13 ISR, 6 postangioplasty restenosis (RS), and 14 primary (PR) atherosclerotic lesions, as well as 15 postmortem coronary artery cross sections from young individuals without atherosclerosis, were studied. All 13 ISR and 4 of 6 RS tissue specimens contained cells that immunolabeled for the primitive cell marker c-kit and smooth muscle alpha-actin, whereas the intima and media of PR lesions and normal arteries were devoid of c-kit-immunopositive cells. The abundance of peripheral blood mononuclear cell-derived CACs was assessed in 10 patients with ISR, 6 patients with angiographically verified patent stents, and 6 individuals with no clinical evidence of coronary artery disease. CACs were less abundant in ISR patients than in non-ISR controls (13.9 +/- 3.1 vs. 22.3 +/- 6.7 cells/high-power field, P < 0.05), and both of these groups had fewer CACs than non-coronary artery disease patients (37.6 +/- 3.8, P < 0.05). These findings suggest a unique pathogenesis for ISR and RS lesions that involves c-kit-immunopositive smooth muscle cells. Moreover, the paucity of CACs in patients with ISR may contribute to the pathogenesis of ISR, perhaps because of attenuated reendothelialization. PMID- 15277196 TI - Myocardial neovascularization by bone marrow angioblasts results in cardiomyocyte regeneration. AB - The primary cardiac response to ischemic insult is cardiomyocyte hypertrophy, which initiates a genetic program culminating in apoptotic myocyte loss, progressive collagen replacement, and heart failure, a process termed cardiac remodeling. Although a few cardiomyocytes at the peri-infarct region can proliferate and regenerate after injury, no approaches are known to effectively induce endogenous cardiomyocytes to enter the cell cycle. We recently isolated, in human adult bone marrow, endothelial progenitor cells, or angioblasts, that migrate to ischemic myocardium, where they induce neovascularization and prevent myocardial remodeling. Here we show that increasing the number of angioblasts trafficking to the infarct zone results in dose-dependent neovascularization with development of progressively larger-sized capillaries. This results in sustained improvement in cardiac function by mechanisms involving protection against apoptosis and, strikingly, induction of proliferation/regeneration of endogenous cardiomyocytes. Our results suggest that agents that increase myocardial homing of bone marrow angioblasts could effectively induce endogenous cardiomyocytes to enter the cell cycle and improve functional cardiac recovery. PMID- 15277197 TI - Hypotonic activation of volume-sensitive outwardly rectifying chloride channels in cultured PASMCs is modulated by SGK. AB - The serum- and glucocorticoid-inducible kinase (SGK) is a serine/threonine protein kinase (PK) transcriptionally regulated by corticoids, serum, and cell volume. SGK regulates cell volume of various cells by effects on Na(+) and K(+) transport through membrane channels. We hypothesized a role for SGK in the activation of volume-sensitive osmolyte and anion channels (VSOACs) in cultured canine pulmonary artery smooth muscle cells (PASMCs). Intracellular dialysis through the patch electrode of recombinant active SGK, but not kinase-dead Delta60-SGK-K127M, heat-inactivated SGK, or active Akt1, partially activated VSOACs under isotonic conditions. Dialysis of active SGK before cell exposure to hypotonic medium significantly accelerated the activation kinetics and increased the maximal density of VSOAC current. Exposure of PASMCs to hypotonic medium (230 mosM) activated phosphatidylinositol 3-kinases (PI3Ks) and their downstream targets Akt/PKB and SGK but not PKC-epsilon. Inhibition of PI3Ks with wortmannin reduced the activation rate and maximal amplitude of VSOACs. Immunoprecipitated ClC-3 channels were phosphorylated by PKC-epsilon but not by SGK in vitro, suggesting that SGK may activate VSOACs indirectly. These data indicate that the PI3K-SGK cascade is activated on hypotonic swelling of PASMCs and, in turn, affects downstream signaling molecules linked to activation of VSOACs. PMID- 15277198 TI - Hyperoxia causes oxygen free radical-mediated membrane injury and alters myocardial function and hemodynamics in the newborn. AB - Newborn children can be exposed to high oxygen levels (hyperoxia) for hours to days during their medical and/or surgical management, and they also can have poor myocardial function and hemodynamics. Whether hyperoxia alone can compromise myocardial function and hemodynamics in the newborn and whether this is associated with oxygen free radical release that overwhelms naturally occurring antioxidant enzymes leading to myocardial membrane injury was the focus of this study. Yorkshire piglets were anesthetized with pentobarbital sodium (65 mg/kg), intubated, and ventilated to normoxia. Once normal blood gases were confirmed, animals were randomly allocated to either 5 h of normoxia [arterial Po(2) (Pa(O(2))) = 83 +/- 5 mmHg, n = 4] or hyperoxia (Pa(O(2)) = 422 +/- 33 mmHg, n = 6), and myocardial functional and hemodynamic assessments were made hourly. Left ventricular (LV) biopsies were taken for measurements of antioxidant enzyme activities [superoxide dismutase (SOD), glutathione peroxidase (GPx), and catalase (CAT)] and malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE) as an indicator of oxygen free radical-mediated membrane injury. Hyperoxic piglets suffered significant reductions in contractility (P < 0.05), systolic blood pressure (P < 0.03), and mean arterial blood pressure (P < 0.05). Significant increases were seen in heart rate (P < 0.05), whereas a significant 11% (P < 0.05) and 61% (P < 0.001) reduction was seen in LV SOD and GPx activities, respectively, after 5 h of hyperoxia. Finally, MDA and 4-HNE levels were significantly elevated by 45% and 38% (P < 0.001 and P = 0.02), respectively, in piglets exposed to hyperoxia. Thus, in the newborn, hyperoxia triggers oxygen free radical-mediated membrane injury together with an inability of the newborn heart to upregulate its antioxidant enzyme defenses while impairing myocardial function and hemodynamics. PMID- 15277199 TI - Effect of estrogen on cerebrovascular prostaglandins is amplified in mice with dysfunctional NOS. AB - Chronic estrogen treatment increases endothelial vasodilator function in cerebral arteries. Endothelial nitric oxide (NO) synthase (eNOS) is a primary target of the hormone, but other endothelial factors may be modulated as well. In light of possible interactions between NO and prostaglandins, we tested the hypothesis that estrogen treatment increases prostanoid-mediated dilation using NOS deficient female mouse models, i.e., mice treated with a NOS inhibitor [N(G) nitro-l-arginine methyl ester (l-NAME)] for 21 days or transgenic mice with the eNOS gene disrupted (eNOS(-/-)). All mice were ovariectomized; some in each group were treated chronically with estrogen. Cerebral blood vessels then were isolated for biochemical and functional analyses. In vessels from control mice, estrogen increased protein levels of eNOS but had no significant effect on cyclooxygenase (COX)-1 protein, prostacyclin production, or constriction of pressurized, middle cerebral arteries to indomethacin, a COX inhibitor. In l-NAME-treated mice, however, cerebrovascular COX-1 levels, prostacyclin production, and constriction to indomethacin, as well as eNOS protein, were all greater in estrogen-treated animals. In vessels from eNOS(-/-) mice, estrogen treatment also increased levels of COX-1 protein and constriction to indomethacin, but no effect on prostacyclin production was detected. Thus cerebral blood vessels of control mice did not exhibit effects of estrogen on the prostacyclin pathway. However, when NO production was dysfunctional, the impact of estrogen on a COX-sensitive vasodilator was revealed. Estrogen has multiple endothelial targets; estrogen effects may be modified by interactions among these factors. PMID- 15277200 TI - Caveolin-3 and SAP97 form a scaffolding protein complex that regulates the voltage-gated potassium channel Kv1.5. AB - The targeting of ion channels to particular membrane microdomains and their organization in macromolecular complexes allow excitable cells to respond efficiently to extracellular signals. In this study, we describe the formation of a complex that contains two scaffolding proteins: caveolin-3 (Cav-3) and a membrane-associated guanylate kinase (MAGUK), SAP97. Complex formation involves the association of Cav-3 with a segment of SAP97 localized between its PDZ2 and PDZ3 domains. In heterologous expression systems, this scaffolding complex can recruit Kv1.5 to form a tripartite complex in which each of the three components interacts with the other two. These interactions regulate the expression of currents encoded by a glycosylation-deficient mutant of Kv1.5. We conclude that the association of Cav-3 with SAP97 may constitute the nucleation site for the assembly of macromolecular complexes containing potassium channels. PMID- 15277201 TI - Reactive oxygen species stimulate central and peripheral sympathetic nervous system activity. AB - Recent studies have implicated reactive oxygen species (ROS) in the pathogenesis of hypertension and activation of the sympathetic nervous system (SNS). Because nitric oxide (NO) exerts a tonic inhibition of central SNS activity, increased production of ROS could enhance inactivation of NO and result in activation of the SNS. To test the hypothesis that ROS may modulate SNS activity, we infused Tempol (4-hydroxy-2,2,6,6-tetramethyl piperidinoxyl), a superoxide dismutase mimetic, or vehicle either intravenously (250 microg x kg(-1) x min(-1)) or in the lateral ventricle (50 microg x kg body wt(-1) x min(-1)), and we determined the effects on blood pressure (BP), norepinephrine (NE) secretion from the posterior hypothalamus (PH) measured by the microdialysis technique, renal sympathetic nerve activity (RSNA) measured by direct microneurography, the abundance of neuronal NO synthase (nNOS)-mRNA in the PH, paraventricular nuclei (PVN), and locus coeruleus (LC) measured by RT-PCR, and the secretion of nitrate/nitrite (NO(x)) in the dialysate collected from the PH of Sprague-Dawley rats. Tempol reduced BP whether infused intravenously or intracerebroventricularly. Tempol reduced NE secretion from the PH and RSNA when infused intracerebroventricularly but raised NE secretion from the PH and RSNA when infused intravenously. The effects of intravenous Tempol on SNS activity were blunted or abolished by sinoaortic denervation. Tempol increased the abundance of nNOS in the PH, PVN, and LC when infused intracerebroventricularly, but it decreased the abundance of nNOS when infused intravenously. When given intracerebroventricularly, Tempol also reduced the concentration of NO(x) in the dialysate collected from the PH. Pretreatment with N(omega)-nitro-l-arginine methyl ester did not abolish the effects of intracerebral Tempol on BP, heart rate, NE secretion from the PH, and RSNA suggesting that the effects of Tempol on SNS activity may be in part dependent and in part independent of NO. In all, these studies support the notion that ROS may raise BP via activation of the SNS. This activation may be mediated in part by downregulation of nNOS and NO production, in part by mechanisms independent of NO. The discrepancy in results between intracerebroventricular and intravenous infusion of Tempol can be best explained by direct inhibitory actions on SNS activity when given intracerebral. By contrast, Tempol may exert direct vasodilation of the peripheral circulation and reflex activation of the SNS when given intravenously. PMID- 15277202 TI - Neural regulation of the proinflammatory cytokine response to acute myocardial infarction. AB - Within minutes of acute myocardial infarction (MI), proinflammatory cytokines increase in the brain, heart, and plasma. We hypothesized that cardiac afferent nerves stimulated by myocardial injury signal the brain to increase central cytokines. Urethane-anesthetized male Sprague-Dawley rats underwent ligation of the left anterior descending coronary artery (LAD) or sham LAD ligation after bilateral cervical vagotomy, sham vagotomy, or application of a 10% phenol solution to the epicardial surface of the myocardium at risk. MI caused a significant increase in tumor necrosis factor (TNF)-alpha and interleukin (IL) 1beta in the plasma and heart, which was blunted by vagotomy. MI also caused a significant increase in hypothalamic TNF-alpha and IL-1beta, which was not affected by vagotomy. In contrast, epicardial phenol blocked MI-induced increases in hypothalamic TNF-alpha and IL-1beta without affecting increases in the plasma and heart. These findings demonstrate that the appearance of proinflammatory cytokines in the brain after MI is independent of blood-borne cytokines and suggest that cardiac sympathetic afferent nerves activated by myocardial ischemia signal the brain to increase cytokine production. In addition, an intact vagus nerve is required for the full expression of proinflammatory cytokines in the injured myocardium and in the circulation. We conclude that the sympathetic and parasympathetic innervation of the heart both contribute to the acute proinflammatory response to MI. PMID- 15277203 TI - Gene transfer of extracellular superoxide dismutase improves relaxation of aorta after treatment with endotoxin. AB - Lipopolysaccharide (LPS) impairs vascular function, in part by generation of reactive oxygen species. One goal of this study was to determine whether gene transfer of extracellular SOD (ECSOD) improves vascular responsiveness in LPS treated rats. A second goal was to determine whether effects of ECSOD are dependent on the heparin-binding domain of the enzyme, which facilitates binding of ECSOD to the outside of cells. Adenoviruses containing ECSOD (AdECSOD), ECSOD with deletion of its heparin-binding domain (AdECSOD-HBD), or a control virus (AdLacZ) were injected intravenously into rats. Three days later, vehicle or LPS (10 mg/kg ip) was injected. After 24 h, vascular reactivity was examined in aortic rings in vitro. Maximum relaxation to acetylcholine was 95 +/- 1% (means +/- SE) after AdlacZ plus vehicle and 77 +/- 3% after AdlacZ plus LPS (P < 0.05). Responses to calcium ionophore A-23187 and submaximal concentrations of nitroprusside also were impaired by LPS. Gene transfer of ECSOD, but not AdECSOD HBD, improved (P < 0.05) relaxation to acetylcholine and A-23187 after LPS. Maximum relaxation to acetylcholine was 88 +/- 3% after LPS plus AdECSOD. Superoxide was increased in aorta after LPS, and the levels were reduced after AdECSOD but not AdECSOD-HBD. LPS-induced adhesion of leukocytes to aortic endothelium was reduced by AdECSOD but not by AdECSOD-HBD. We conclude that after gene transfer in vivo, binding of ECSOD to arteries effectively decreases the numbers of adherent leukocytes and levels of superoxide and improves impaired endothelium-dependent relaxation produced by LPS. PMID- 15277204 TI - Is there a correlation between ventricular fibrillation cycle length and electrophysiological and anatomic properties of the canine left ventricle? AB - We hypothesized that myocardial infarction-related alterations in ventricular fibrillation (VF) cycle length (VFCL) would correlate with changes in local cardiac electrophysiological and anatomic properties. An electrophysiological study was performed in normal, subacute, and chronic infarction mongrel dogs. VF was induced by programmed electrical stimulation and mean and minimum early and late VFCL was determined and correlated with local electrophysiological and anatomic properties. Effective refractory period (ERP), activation recovery time (ART), ERP/ART ratio, threshold, and ERP and ART dispersion were determined at 112 sites on the anterior left ventricle. Wave front progression was analyzed over a 2-s period. The extent of local tissue necrosis and of myocardial fiber disarray was also evaluated. The early mean VFCL was significantly longer in the subacute infarction (149 +/- 35 ms) and chronic infarction dogs (129 +/- 18 ms) compared with control dogs (102 +/- 15 ms; P < 0.0001 for both comparisons) as was the early minimum VFCL with similar trends seen during late VF. Complete epicardial reentrant circuits were significantly more common in normal dogs (4.3 +/- 2.4, 22.4% of cycles) than in subacute (0.75 +/- 0.96, 5.3% of cycles, P < 0.05 vs. normal) and chronic infarction dogs (1.3 +/- 1.3, 7.5% of cycles, P < 0.05 vs. normal). There was a poor correlation between the mean and minimum early and late VFCL and local electrophysiological and anatomic properties (R(2) < 0.2 for all comparisons) with a much better correlation between average mean and minimum VFCL (over the entire plaque) and global ERP and ART dispersion during early and late VF. In conclusion, VFCL in normal and infarcted myocardium shows a poor correlation with local ventricular electrophysiological and anatomic properties measured in sinus rhythm. However, there was a much better correlation between the average VFCL with global dispersion of repolarization. The lack of correlation between local VFCL and refractoriness and the infrequent occurrence of epicardial reentry suggests that intramural reentry may be the primary mechanism of VF in this model. PMID- 15277205 TI - Theoretical simulation of K(+)-based mechanisms for regulation of capillary perfusion in skeletal muscle. AB - Muscle fibers release K(+) into the interstitial space upon recruitment. Increased local interstitial K(+) concentration ([K(+)]) can cause dilation of terminal arterioles, leading to perfusion of downstream capillaries. The possibility that capillary perfusion can be regulated by vascular responses to [K(+)] was examined using a theoretical model. The model takes into account the spatial relationship between functional units of muscle fiber recruitment and capillary perfusion. Diffusion of K(+) in the interstitial space was simulated. Two hypothetical mechanisms for vascular sensing of interstitial [K(+)] were considered: direct sensing by arterioles and sensing by capillaries with stimulation of feeding arterioles via conducted responses. Control by arteriolar sensing led to poor tissue oxygenation at high levels of muscle activation. With control by capillary sensing, increases in perfusion matched increases in oxygen demand. The time course of perfusion after sudden muscle activation was considered. Predicted capillary perfusion increased rapidly within the first 5 s of muscle fiber activation. The reuptake of K(+) by muscle fibers had a minor effect on the increase of interstitial [K(+)]. Uptake by perfused capillaries was primarily responsible for limiting the increase in [K(+)] in the interstitial space at the onset of fiber activation. Vascular responses to increasing interstitial [K(+)] may contribute to the rapid increase in blood flow that is observed to occur after the onset of muscle contraction. PMID- 15277206 TI - Voluntary physical activity alterations in endothelial nitric oxide synthase knockout mice. AB - One of the main factors that control vasoreactivity and angiogenesis is nitric oxide produced by endothelial nitric oxide synthase (eNOS). We recently showed that knocking out eNOS induces an important reduction of mitochondrial oxidative capacity in slow-twitch skeletal muscle. Here we investigated eNOS's role in physical activity and contribution to adaptation of muscle energy metabolism to exercise conditions. Physical capacity of mice null for the eNOS isoform (eNOS-/ ) was estimated for 8 wk with a voluntary wheel-running protocol. In parallel, we studied energy metabolism enzyme profiles and their response to voluntary exercise in cardiac and slow-twitch soleus (Sol) and fast-twitch gastrocnemius (Gast) skeletal muscles. Weekly averaged running distance was two times lower for eNOS-/- (4.09 +/- 0.42 km/day) than for wild-type (WT; 7.74 +/- 0.42 km/day; P < 0.01) mice. Average maximal speed of running was also lower in eNOS-/- (17.2 +/- 1.4 m/min) than WT (21.2 +/- 0.9 m/min; P < 0.01) mice. Voluntary exercise influenced adaptation to exercise specifically in Sol muscle. Physical activity significantly increased Sol weight by 22% (P < 0.05) in WT but not eNOS-/- mice. WT Sol muscle did not change its metabolic profile in response to exercise, in contrast to eNOS-/- muscle, in which physical activity decreased cytochrome-c oxidase (COX; -36%; P < 0.05), citrate synthase (-37%; P < 0.06), and creatine kinase (-24%, P < 0.01) activities. Voluntary exercise did not change energy enzyme profile in heart (except for 39% increase in COX activity in WT) or Gast muscle. These results suggest that eNOS is necessary for maintaining a suitable physical capacity and that when eNOS is downregulated, even moderate exercise could worsen energy metabolism specifically in oxidative skeletal muscle. PMID- 15277207 TI - Effect of NADPH oxidase inhibition on cardiopulmonary bypass-induced lung injury. AB - Cardiopulmonary bypass (CPB) causes acute lung injury. Reactive oxygen species (ROS) from NADPH oxidase may contribute to this injury. To determine the role of NADPH oxidase, we pretreated pigs with structurally dissimilar NADPH oxidase inhibitors. Low-dose apocynin (4-hydroxy-3-methoxy-acetophenone; 200 mg/kg, n = 6), high-dose apocynin (400 mg/kg, n = 6), or diphenyleneiodonium (DPI; 8 mg/kg) was compared with diluent (n = 8). An additional group was treated with indomethacin (10 mg/kg, n = 3). CPB was performed for 2 h with deflated lungs, complete pulmonary artery occlusion, and bronchial artery ligation to maximize lung injury. Parameters of pulmonary function were evaluated for 25 min following CPB. Blood chemiluminescence indicated neutrophil ROS production. Electron paramagnetic resonance determined the effect of apocynin and DPI on in vitro pulmonary endothelial ROS production following hypoxia-reoxygenation. Both apocynin and DPI attenuated blood chemiluminescence and post-CPB hypoxemia. At 25 min post-CPB with Fi(O(2)) = 1, arterial Po(2) (Pa(o(2))) averaged 52 +/- 5, 162 +/- 54, 335 +/- 88, and 329 +/- 119 mmHg in control, low-dose apocynin, high-dose apocynin, and DPI-treated groups, respectively (P < 0.01). Indomethacin had no effect. Pa(O(2)) correlated with blood chemiluminescence measured after drug administration before CPB (R = -0.60, P < 0.005). Neither apocynin nor DPI prevented the increased tracheal pressure, plasma cytokine concentrations (tumor necrosis factor-alpha and IL-6), extravascular lung water, and pulmonary vascular protein permeability observed in control pigs. NADPH oxidase inhibition, but not xanthine oxidase inhibition, significantly blocked endothelial ROS generation following hypoxia-reoxygenation (P < 0.05). NADPH oxidase-derived ROS contribute to the severe hypoxemia but not to the increased cytokine generation and pulmonary vascular protein permeability, which occur following CPB. PMID- 15277208 TI - Loss of PKC-delta alters cardiac metabolism. AB - PKC-delta is believed to play an essential role in cardiomyocyte growth. In the present study, we investigated the effect of PKC-delta on cardiac metabolism using PKC-delta knockout mice generated in our laboratories. Proteomic analysis of heart protein extracts revealed profound changes in enzymes related to energy metabolism: certain isoforms of glycolytic enzymes, e.g., lactate dehydrogenase and pyruvate kinase, were absent or decreased, whereas several enzymes involved in lipid metabolism, e.g., phosphorylated isoforms of acyl-CoA dehydrogenases, showed a marked increase in PKC-delta(-/-) hearts. Moreover, PKC-delta deficiency was associated with changes in antioxidants, namely, 1-Cys peroxiredoxin and selenium-binding protein 1, and posttranslational modifications of chaperones involved in cytoskeleton regulation, such as heat shock protein (HSP)20, HSP27, and the zeta-subunit of the cytosolic chaperone containing the T-complex polypeptide 1. High-resolution NMR analysis of cardiac metabolites confirmed a significant decrease in the ratio of glycolytic end products (alanine + lactate) to end products of lipid metabolism (acetate) in PKC-delta(-/-) hearts. Taken together, our data demonstrate that loss of PKC-delta causes a shift from glucose to lipid metabolism in murine hearts, and we provide a detailed description of the enzymatic changes on a proteomic level. The consequences of these metabolic alterations on sensitivity to myocardial ischemia are further explored in the accompanyingpaper (20). PMID- 15277210 TI - Measures of venous function: technical issues and novelty of findings. PMID- 15277209 TI - Ischemic preconditioning exaggerates cardiac damage in PKC-delta null mice. AB - Ischemic preconditioning confers cardiac protection during subsequent ischemia reperfusion, in which protein kinase C (PKC) is believed to play an essential role, but controversial data exist concerning the PKC-delta isoform. In an accompanying study (26), we described metabolic changes in PKC-delta knockout mice. We now wanted to explore their effect on early preconditioning. Both PKC delta(-/-) and PKC-delta(+/+) mice underwent three cycles of 5-min left descending artery occlusion/5-min reperfusion, followed by 30-min occlusion and 2 h reperfusion. Unexpectedly, preconditioning exaggerated ischemia-reperfusion injury in PKC-delta(-/-) mice. Whereas ischemic preconditioning increased superoxide anion production in PKC-delta(+/+) hearts, no increase in reactive oxygen species was observed in PKC-delta(-/-) hearts. Proteomic analysis of preconditioned PKC-delta(+/+) hearts revealed profound changes in enzymes related to energy metabolism, e.g., NADH dehydrogenase and ATP synthase, with partial fragmentation of these mitochondrial enzymes and of the E(2) component of the pyruvate dehydrogenase complex. Interestingly, fragmentation of mitochondrial enzymes was not observed in PKC-delta(-/-) hearts. High-resolution NMR analysis of cardiac metabolites demonstrated a similar rise of phosphocreatine in PKC delta(+/+) and PKC-delta(-/-) hearts, but the preconditioning-induced increase in phosphocholine, alanine, carnitine, and glycine was restricted to PKC-delta(+/+) hearts, whereas lactate concentrations were higher in PKC-delta(-/-) hearts. Taken together, our results suggest that reactive oxygen species generated during ischemic preconditioning might alter mitochondrial metabolism by oxidizing key mitochondrial enzymes and that metabolic adaptation to preconditioning is impaired in PKC-delta(-/-) hearts. PMID- 15277211 TI - Caspase-mediated degeneration in Alzheimer's disease. PMID- 15277212 TI - Long-term accumulation of amyloid-beta, beta-secretase, presenilin-1, and caspase 3 in damaged axons following brain trauma. AB - Plaques composed of amyloid beta (Abeta) have been found within days following brain trauma in humans, similar to the hallmark plaque pathology of Alzheimer's disease (AD). Here, we evaluated the potential source of this Abeta and long-term mechanisms that could lead to its production. Inertial brain injury was induced in pigs via head rotational acceleration of 110 degrees over 20 ms in the coronal plane. Animals were euthanized at 3 hours, 3 days, 7 days, and 6 months post injury. Immunohistochemistry and Western blot analyses of the brains were performed using antibodies specific for amyloid precursor protein (APP), Abeta peptides, beta-site APP-cleaving enzyme (BACE), presenilin-1 (PS-1), caspase-3, and caspase-mediated cleavage of APP (CCA). Substantial co-accumulation for all of these factors was found in swollen axons at all time points up to 6 months following injury. Western blot analysis of injured brains confirmed a substantial increase in the protein levels of these factors, particularly in the white matter. These data suggest that impaired axonal transport due to trauma induces long-term pathological co-accumulation of APP with BACE, PS-1, and activated caspase. The abnormal concentration of these factors may lead to APP proteolysis and Abeta formation within the axonal membrane compartment. PMID- 15277213 TI - Degranulation of paneth cells via toll-like receptor 9. AB - The release of antimicrobial peptides and growth factors by Paneth cells is thought to play an important role in protecting the small intestine, but the mechanisms involved have remained obscure. Immunohistochemistry and immunofluorescence showed that Paneth cells express Toll-like receptor 9 (TLR9) in the granules. Injection of mice with oligonucleotides containing CpG sequence (CpG-ODNs) led to a down-modulation of TLR9 and a striking decrease in the number of large secretory granules, consistent with degranulation. Moreover CpG-ODN treatment increased resistance to oral challenge with virulent Salmonella typhimurium. Moreover, our findings demonstrate a sentinel role for Paneth cells through TLR9. PMID- 15277214 TI - Regulation of fibrillin-1 by biglycan and decorin is important for tissue preservation in the kidney during pressure-induced injury. AB - There is growing evidence that the two small leucine-rich proteoglycans biglycan and decorin regulate the assembly of connective tissues and alter cell behavior during development and pathological processes. In this study, we have used an experimental animal model of unilateral ureteral ligation and mice deficient in either biglycan or decorin. We discovered that pressure-induced injury to the wild-type kidneys led to overexpression of decorin, biglycan, fibrillin-1, and fibrillin-2. In contrast, in biglycan-deficient kidneys the overexpression of fibrillin-1 was markedly attenuated and this was associated with cystic dilatation of Bowman's capsule and proximal tubules. Notably, we found that in ligated kidneys from decorin-null mice, fibrillin-1 expression was initially enhanced to the same extent as in wild-type animals. However, long-term obstruction resulted in down-regulation of fibrillin-1 and concurrent cystic dilatation of Bowman's capsule in 33% of kidneys at 5 months after obstruction. In all of the genotypes, no differences in fibrillin-2 expression were observed. These in vivo data correlated with a significant induction of fibrillin-1 expression in renal fibroblasts and mesangial cells by recombinant biglycan and decorin. Our results indicate a novel role for decorin and biglycan during pressure-induced renal injury by stimulating fibrillin-1 expression. PMID- 15277215 TI - Differential gene expression in ovarian carcinoma: identification of potential biomarkers. AB - Ovarian cancer remains the fifth leading cause of cancer death for women in the United States. In this study, the gene expression of 20 ovarian carcinomas, 17 ovarian carcinomas metastatic to the omentum, and 50 normal ovaries was determined by Gene Logic Inc. using Affymetrix GeneChip HU_95 arrays containing approximately 12,000 known genes. Differences in gene expression were quantified as fold changes in gene expression in ovarian carcinomas compared to normal ovaries and ovarian carcinoma metastases. Genes up-regulated in ovarian carcinoma tissue samples compared to more than 300 other normal and diseased tissue samples were identified. Seven genes were selected for further screening by immunohistochemistry to determine the presence and localization of the proteins. These seven genes were: the beta8 integrin subunit, bone morphogenetic protein-7, claudin-4, collagen type IX alpha2, cellular retinoic acid binding protein-1, forkhead box J1, and S100 calcium-binding protein A1. Statistical analyses showed that the beta8 integrin subunit, claudin-4, and S100A1 provided the best distinction between ovarian carcinoma and normal ovary tissues, and may serve as the best candidate tumor markers among the seven genes studied. These results suggest that further exploration into other up-regulated genes may identify novel diagnostic, therapeutic, and/or prognostic biomarkers in ovarian carcinoma. PMID- 15277216 TI - Increased angiogenic response in aortic explants of collagen XVIII/endostatin null mice. AB - Endostatin, a proteolytic fragment of basement membrane-associated collagen XVIII, has been shown to be a potent angiogenesis inhibitor both in vivo and in vitro when given at high concentrations. The precise molecular mechanisms by which it functions and whether or not it plays a role in physiological regulation of angiogenesis are not clear. In mice with targeted null alleles of Col18a1, there appears to be no major abnormality in vascular patterns or capillary density in most organs. Furthermore, the growth of experimental tumors is not increased. However, a detailed analysis of induced angiogenesis in these mice has not been performed. Therefore, we compared the angiogenic responses induced by in vitro culture of aortic explants from collagen XVIII/endostatin-null mice (ko) to wild-type (wt) littermates. We found a twofold increase in microvessel outgrowth in explants from ko mice, relative to wt explants. This increased angiogenesis was reduced to the wt level by the addition of low levels (0.1 microg/ml) of recombinant mouse or human endostatin during the culture period. To address cellular/molecular mechanisms underlying this difference in angiogenic response between ko and wt mice, we isolated endothelial cells from both strains and compared their biological behavior. Proliferation assays showed no difference between the two types of endothelial cells. In contrast, adhesion assays showed a striking difference in their ability to adhere to fibronectin suggesting that collagen XVIII/endostatin may regulate interactions between endothelial cells and underlying basement membrane-associated components, including fibronectin, such that in the absence of collagen XVIII/endostatin, endothelial cells are more adhesive to fibronectin. In the aortic explant assay, characterized by dynamic processes of microvessel elongation and regression, this may result in stabilization of newly formed vessels, reduced regression, and a net increase in microvessel outgrowth in explants from ko mice compared to the wt littermates. PMID- 15277217 TI - Antigen transport and cytoskeletal characteristics of a distinct enterocyte population in inflammatory bowel diseases. AB - Intestinal antigen uptake is enhanced in inflammatory bowel disease. We analyzed transcellular transport routes of antigens in different compartments of normal enterocytes and atypical intestinal epithelial cells called "rapid antigen uptake into the cytosol enterocytes" (RACE cells). These cells constitute a recently described population of enterocyte-derived cells, which are increased in inflammatory bowel disease. Mucosa of freshly resected specimens were incubated with the antigens ovalbumin or horseradish peroxidase. Ultrastructural labeling patterns of differentiation-dependent proteins, the brush-border enzyme sucrase isomaltase and the cytoskeleton proteins villin and actin, were determined in enterocytes. Apoptosis was investigated biochemically and ultrastructurally by cleavage of caspase-3. Both antigens were transported to late endosomes and to trans-Golgi vesicles of enterocytes in inflammatory bowel disease and control specimens. Quantitative evaluation revealed a significantly increased transepithelial antigen transport in both compartments of RACE relative to normal enterocytes. Labeling densities for sucrase-isomaltase, villin, and actin were decreased in RACE relative to normal enterocytes. Caspase-3 was not increased in RACE cells relative to controls. RACE cells are characterized by increased antigen transport to late endosomes and the trans-Golgi network, a disassembled cytoskeleton and lower concentrations of proteins that are markers of cell differentiation. PMID- 15277218 TI - Targeted deletion of CC chemokine receptor 2 attenuates left ventricular remodeling after experimental myocardial infarction. AB - A key component of cardiac remodeling after acute myocardial infarction (MI) is the inflammatory response, which modulates cardiac tissue repair. The purpose of this study was to investigate the relationship between the monocytic inflammatory response and left ventricular remodeling after MI using mice deficient in CC chemokine receptor 2 (CCR2), the primary receptor for the critical regulator of CC chemokine ligand 2. Immunohistochemical analysis revealed rapid infiltration of macrophages into infarcted tissue within 7 days in wild-type (WT) mice. However, this process was greatly impaired in CCR2-deficient (CCR2(-/-)) mice. Echocardiography demonstrated beneficial effects of CCR2 deficiency on left ventricular remodeling at 7 and 28 days after MI. In situ zymography showed augmented gelatinolytic activity in WT mice within 7 days after MI, whereas gelatinolytic activity was barely detectable in CCR2(-/-) mice. Moreover, the distribution of gelatinolytic activity in serial sections was very similar to the distribution of macrophages rather than neutrophils. Expression of matrix metalloproteinases and tumor necrosis factor-alpha mRNAs was up-regulated in infarcted regions from WT mice compared to CCR2(-/-) mice at 3 days after MI. Direct inhibition of CCR2 functional pathway might contribute to the attenuation of left ventricular remodeling after MI. PMID- 15277219 TI - Increased blood clotting, microvascular density, and inflammation in eotaxin secreting tumors implanted into mice. AB - An important theme that is emerging in cancer research is the interaction between tumor cells and the host stroma. Because many types of human cancer are infiltrated by eosinophils that are believed to mediate an anti-tumor cytotoxic effect, we developed and studied a transfected B16 murine melanoma cell line that secretes high levels (510 pg/ml/100,000 cells/day) of eotaxin, a chemokine that recruits and activates primarily eosinophils. Here we report that there was increased inflammation (eosinophils, mast cells, mononuclear cells), blood clotting, and microvascular density within the tumors produced by subcutaneous implants of eotaxin-secreting tumor cells in 10 C57BL/6 compared to tumors produced by wild-type tumor cells. The extensive blood clotting in the eotaxin transfected tumors was associated with significantly decreased blood flow to the tumors as measured by magnetic resonance imaging [(mean maximum signal enhancement of eotaxin-secreting tumors, 147 +/- 57 (n = 7) compared to 202 +/- 36 signal enhancement units (n = 8) for the wild-type melanoma cells; P = 0.04 by two-tailed, unpaired t-test]. Surprisingly, there was no significant difference between the growth rates or mean masses of the eotaxin-secreting tumors (750 +/- 280 mg, n = 10) and the wild-type tumors (780 +/- 290, n = 10) after 20 days of growth in vivo, despite the significantly slower growth rate in vitro of the eotaxin-secreting tumor cells. We conclude that eotaxin and the resultant tumor infiltrating inflammatory cells are not likely to mediate a significant anti tumor effect in vivo. Instead, elevated eotaxin is associated with increased inflammation, microvascular density, and blood clotting. Thus, eotaxin and eosinophils may play a more complex role in modulating the growth of tumors than the simple, anti-tumor cytotoxic effect that has been previously proposed. PMID- 15277221 TI - Interleukin-6/soluble interleukin-6 receptor signaling attenuates proliferation and invasion, and induces morphological changes of a newly established pleomorphic malignant fibrous histiocytoma cell line. AB - Pleomorphic malignant fibrous histiocytoma (MFH) is occasionally associated with inflammatory paraneoplastic syndrome (PNS). Recently, we reported that interleukin (IL)-6, one of the candidate cytokines, which induces such systemic inflammatory reaction, may be a tumor-associated factor involved in the pathogenesis and its clinical manifestations of MFH. In the local microenvironment, tumor-induced inflammatory reaction may play a role favoring tumor progression. To clarify the biological relevance of IL-6 in MFH, we established a human MFH cell line, named MIPS-2, derived from a resected specimen of a patient presenting with PNS. In this patient, the serum IL-6 level ran parallel to the disease course: elevated serum IL-6 concentration normalized immediately after radical surgery, and re-elevation occurred on tumor recurrence. MIPS-2 presented pleomorphic appearance, severe nuclear abnormalities with prominent nucleoli, and tumorigenesis in nude mice. MIPS-2 expressed IL-6, IL-6 receptor (IL-6R), and glycoprotein 130 (gp130) but lacked the soluble form of IL 6R (sIL-6R), as determined by flow cytometry and reverse transcriptase-polymerase chain reaction analyses. Stimulation of MIPS-2 with IL-6 combined with exogenous sIL-6R induced phosphorylation of both signal transducer and activator of transcription 3 (STAT3) and mitogen-activated protein kinase (MAPK), decreased cell proliferation, attenuated invasion, and induced morphological changes. Collectively, these data suggested that the IL-6/sIL-6R signaling pathway plays a pivotal role for proliferation, invasion, and morphology of MFH via STAT3 and MAPK pathway as autocrine and/or paracrine manner, and proposed the therapeutic potential for the use of both anti-growth factor and proinflammatory cytokine targeting strategies to combat devastating MFH. PMID- 15277220 TI - Insulin-like growth factor-I plays a pathogenetic role in diabetic retinopathy. AB - Diabetic retinopathy is a leading cause of blindness in the Western world. Aberrant intercellular adhesion molecule-1 expression and leukocyte adhesion have been implicated in its pathogenesis, raising the possibility of an underlying chronic inflammatory mechanism. In the current study, the role of insulin-like growth factor (IGF)-I in these processes was investigated. We found that systemic inhibition of IGF-I signaling with a receptor-neutralizing antibody, or with inhibitors of PI-3 kinase (PI-3K), c-Jun kinase (JNK), or Akt, suppressed retinal Akt, JNK, HIF-1alpha, nuclear factor (NF)-kappaB, and AP-1 activity, vascular endothelial growth factor (VEGF) expression, as well as intercellular adhesion molecule-1 levels, leukostasis, and blood-retinal barrier breakdown, in a relevant animal model. Intravitreous administration of IGF-I increased retinal Akt, JNK, HIF-1alpha, NF-kappaB, and AP-1 activity, and VEGF levels. IGF-I stimulated VEGF promoter activity in vitro, mainly via HIF-1alpha, and secondarily via NF-kappaB and AP-1. In conclusion, IGF-I participates in the pathophysiology of diabetic retinopathy by inducing retinal VEGF expression via PI-3K/Akt, HIF-1alpha, NF-kappaB, and secondarily, JNK/AP-1 activation. Taken together, these in vitro and in vivo signaling studies thus identify potential targets for pharmacological intervention to preserve vision in patients with diabetes. PMID- 15277222 TI - Cytogenetic alterations affecting BCL6 are predominantly found in follicular lymphomas grade 3B with a diffuse large B-cell component. AB - Recently, classical banding cytogenetic studies suggested that follicular lymphomas (FLs) grade 3 with preserved maturation to centrocytes (FL3A) are closely related to FL grades 1 and 2 and frequently harbor the t(14;18), whereas FL grade 3B, consisting of centroblasts exclusively, do frequently show 3q27 alterations. To clarify the prevalence of BCL6 and BCL2 rearrangements in FL and diffuse large B-cell lymphomas (DLBLs), we performed a large scale bicolor interphase cytogenetic (fluorescence in situ hybridization) study on 188 well characterized B-NHLs classified according to the World Health Organization Classification of Tumors of the Lymphoid Tissues. BCL6 rearrangements were detected in a significantly higher number of FL3B with a DLBL component (12 of 22, 55%) compared with purely diffuse nodal DLBLs (19 of 77, 25%) and DLBLs with a well-documented primary extranodal origin (2 of 27, 7%) (P < 0.001). Five FL3B without a DLBL component were negative for both t(14;18) and 3q27 aberrations. FL grades 1/2 and FL3A were t(14;18)-positive in 88% and 64% of cases, respectively, but 3q27 alterations were identified in only four FL3A. These data exemplify different genetic pathways in the genesis of FLs with a high content of centroblasts and suggest that 3q27 rearrangements are predominantly associated with FL grade 3B harboring a DLBL component. PMID- 15277224 TI - Effective hepatocyte transplantation using rat hepatocytes with low asialoglycoprotein receptor expression. AB - Development of a reliable method of isolating highly proliferative potential hepatocytes provides information crucial to progress in the field of hepatocyte transplantation. The aim of this study was to develop reliable hepatocyte transplantation using highly proliferative, eg, progenitor-like hepatocytes, based on asialoglycoprotein receptor (ASGPR) expression levels for hepatocyte transplantation. We have previously reported that mouse hepatocytes with low ASGPR expression levels have highly proliferative potential and can be used as progenitor-like hepatocytes. We therefore fractionated F344 male rat hepatocytes expressing low and high levels of ASGPR and determined the liver repopulation capacity of hepatocytes according to low and high ASGPR expression in the liver. Next, 2 x 10(5) cells of each type were transplanted into female liver regenerative model dipeptidyl peptidase-deficient rats, and we estimated the rate of liver repopulation by the transplanted hepatocytes in the host liver, as determined by recognition of the Sry gene on the Y-chromosome. At 60 days after hepatocyte transplantation, the transplanted hepatocytes occupied approximately 76% of the total hepatocyte mass in the case of the transplantation of hepatocytes with low ASGPR expression, but accounted for approximately 12% and 17% of the mass in the case of the transplantation of hepatocytes with high ASGPR expression and unfractionated hepatocytes, respectively. In conclusion, these findings suggest that hepatocytes with low ASGPR expression can result in normal liver function and a high repopulation capacity in vivo. These results provide insight into development of a strategy for effective liver repopulation using transplanted hepatocytes. PMID- 15277223 TI - Number of mast cells in the peritoneal cavity of mice: influence of microphthalmia transcription factor through transcription of newly found mast cell adhesion molecule, spermatogenic immunoglobulin superfamily. AB - The mi (microphthalmia) locus of mice encodes a transcription factor, MITF. B6 tg/tg mice that do not express any MITF have white coats and small eyes. Moreover, the number of mast cells decreased to one-third that of normal control (+/+) mice in the skin of B6-tg/tg mice. No mast cells were detectable in the stomach, mesentery, and peritoneal cavity of B6-tg/tg mice. Cultured mast cells derived from B6-tg/tg mice do not express a mast cell adhesion molecule, spermatogenic immunoglobulin superfamily (SgIGSF). To obtain in vivo evidence for the correlation of nonexpression of SgIGSF with decrease in mast cell number, we used another MITF mutant, B6-mi(vit)/mi(vit) mice that have a mild phenotype, ie, black coat with white patches and eyes of normal size. B6-mi(vit)/mi(vit) mice had a normal number of mast cells in the skin, stomach, and mesentery, but the number of peritoneal mast cells decreased to one-sixth that of +/+ mice. Cultured mast cells and peritoneal mast cells of B6-mi(vit)/mi(vit) mice showed a reduced but apparently detectable level of SgIGSF expression, demonstrating the parallelism between mast cell number and expression level of SgIGSF. The number of peritoneal mast cells appeared to be influenced by MITF through transcription of SgIGSF. PMID- 15277226 TI - Active caspase-6 and caspase-6-cleaved tau in neuropil threads, neuritic plaques, and neurofibrillary tangles of Alzheimer's disease. AB - Previously, we have shown that caspase-6 but not caspase-3 is activated by serum deprivation and induces a protracted cell death in primary cultures of human neurons (LeBlanc AC, Liu H, Goodyer C, Bergeron C, Hammond J: Caspase-6 role in apoptosis of human neurons, amyloidogenesis and Alzheimer's disease. J Biol Chem 1999, 274:23426-23436 and Zhang Y, Goodyer C, LeBlanc A: Selective and protracted apoptosis in human primary neurons microinjected with active caspase-3, -6, -7, and -8. J Neurosci 2000, 20:8384-8389). Here, we show with neoepitope antibodies that the p20 subunit of active caspase-6 increases twofold to threefold in the affected temporal and frontal cortex but not in the unaffected cerebellum of Alzheimer's disease brains and is present in neurofibrillary tangles, neuropil threads, and the neuritic plaques. Furthermore, a neoepitope antibody to caspase 6-cleaved Tau strongly detects intracellular tangles, extracellular tangles, pretangles, neuropil threads, and neuritic plaques. Immunoreactivity with both antibodies in pretangles indicates that the caspase-6 is active early in the pathogenesis of Alzheimer's disease. In contrast to the nuclear and cytosolic localization of active caspase-6 in apoptotic neurons of fetal and adult ischemic brains, the active caspase-6 in Alzheimer's disease brains is sequestered into the tangles or neurites. The localization of active caspase-6 may strongly jeopardize the structural integrity of the neuronal cytoskeletal system leading to inescapable neuronal dysfunction and eventual cell death in Alzheimer's disease neurons. Our results suggest that active caspase-6 is strongly implicated in human neuronal degeneration and apoptosis. PMID- 15277225 TI - The oncogenic activity of RET point mutants for follicular thyroid cells may account for the occurrence of papillary thyroid carcinoma in patients affected by familial medullary thyroid carcinoma. AB - Activating germ-line point mutations in the RET receptor are responsible for multiple endocrine neoplasia type 2-associated medullary thyroid carcinoma (MTC), whereas somatic RET rearrangements are prevalent in papillary thyroid carcinomas (PTCs). Some rare kindreds, carrying point mutations in RET, are affected by both cancer types, suggesting that, under specific circumstances, point mutations in RET can drive the generation of PTC. Here we describe a family whose siblings, affected by both PTC and MTC, carried a germ-line point mutation in the RET extracellular domain, converting cysteine 634 into serine. We tested on thyroid follicular cells the transforming activity of RET(C634S), RET(K603Q), another mutant identified in a kindred with both PTC and MTC, RET(C634R) a commonly isolated allele in MEN2A, RET(M918T) responsible for MEN2B and also identified in kindreds with both PTC and MTC, and RET/PTC1 the rearranged oncogene that characterizes bona fide PTC in patients without MTC. We show that the various RET point mutants, but not wild-type RET, scored constitutive kinase activity and exerted mitogenic effects for thyroid PC Cl 3 cells, albeit at significantly lower levels compared to RET/PTC1. The low mitogenic activity of RET point mutants paralleled their reduced kinase activity compared to RET/PTC. Furthermore, RET point mutants maintained a protein domain, the intracellular juxtamembrane domain, that exerted negative effects on the mitogenic activity. In conclusion, RET point mutants can behave as dominant oncogenes for thyroid follicular cells. Their transforming activity, however, is rather modest, providing a possible explanation for the rare association of MTC with PTC. PMID- 15277227 TI - Phenotypic switch from paracrine to autocrine role of hepatocyte growth factor in an androgen-independent human prostatic carcinoma cell line, CWR22R. AB - Support mechanisms involved in growth of androgen-independent prostate cancer are primarily unknown. Hepatocyte growth factor (HGF)/Met has been suggested to be one of them based primarily on immunohistochemical studies. We conducted a series of experiments to assess the role of the HGF/Met system in an androgen-dependent human prostate carcinoma, CWR22 and its androgen-independent derivative, CWR22R. We found that action of HGF changed from paracrine to autocrine in progression to androgen-independent state. CWR22 tumors did not express HGF but expressed Met, whereas prostate stromal cells expressed HGF at a high level. Growth of CWR22 was stimulated either by addition of HGF to the culture or by the presence of prostate stromal cells. On the other hand, CWR22R cells expressed both HGF and Met. Knockdown of Met expression by RNA interference method suppressed the growth of CWR22R cells. Our data suggest that HGF is intimately involved in growth of human prostate cancer and that progression from the androgen-dependent to the androgen-independent state is associated with an adaptive switch in support mechanism from paracrine to autocrine. Our data offer one mechanism to account for androgen-independent human cancer growth. PMID- 15277228 TI - Expression of the type-1 repeats of thrombospondin-1 inhibits tumor growth through activation of transforming growth factor-beta. AB - In the present study, the type-1 repeats of thrombospondin-1 (TSP-1) were transfected into A431 cells. Expression of all three type-1 repeats (3TSR) and expression of just the second type-1 repeat containing the transforming growth factor (TGF)-beta activating sequence KRFK (TSR2 + KRFK) significantly inhibited in vivo tumor angiogenesis and growth in nude mice. These tumors expressed increased levels of both active and total TGF-beta. A431 cells expressing the second type-1 repeat without the KRFK sequence (TSR2 - KRFK) produced tumors that were slightly larger than the 3TSR and TSR2 + KRFK tumors. These tumors expressed elevated levels of active TGF-beta but levels of total TGF-beta were not different from control tumors. Injection of the peptide, LSKL, which blocks TSP-1 activation of TGF-beta, reversed the growth inhibition observed with cells expressing TSR2 + KRFK to a level comparable to controls. Various residues in the WSHWSPW region and the VTCG sequence of both TSR2+/- KRFK were mutated. Although mutation of the VTCG sequence had no significant effect on tumor growth, mutation of the WSHWSPW sequence reduced inhibition of tumor growth. These findings suggest that the inhibition of tumor angiogenesis and growth by endogenous TSP-1 involves regulation of both active and total TGF-beta and the sequences KRFK and WSHWSPW in the second type-1 repeat. PMID- 15277229 TI - Expression of histone deacetylase 8, a class I histone deacetylase, is restricted to cells showing smooth muscle differentiation in normal human tissues. AB - Histone deacetylases (HDACs) were originally identified as nuclear enzymes involved in gene transcription regulation. Until recently, it was thought that their activity was restricted within the nucleus, with histones as unique substrates. The demonstration that specific HDACs deacetylate nonhistone proteins, such as p53 and alpha-tubulin, broadened the field of activity of these enzymes. HDAC8, a class I HDAC, is considered to be ubiquitously expressed, as suggested by results of Northern blots performed on tissue RNA extracts, and transfection experiments using various cell lines have indicated that this enzyme may display a prominent nuclear localization. Using immunohistochemistry, we unexpectedly found that, in normal human tissues, HDAC8 is exclusively expressed by cells showing smooth muscle differentiation, including visceral and vascular smooth muscle cells, myoepithelial cells, and myofibroblasts, and is mainly detected in their cytosol. These findings were confirmed in vitro by nucleo cytoplasmic fractionation and immunoblot experiments performed on human primary smooth muscle cells, and by the cytosolic detection of epitope-tagged HDAC8 overexpressed in fibroblasts. Immunocytochemistry strongly suggested a cytoskeleton-like distribution of the enzyme. Further double-immunofluorescence staining experiments coupled with confocal microscopy analysis showed that epitope-tagged HDAC8 overexpressed in murine fibroblasts formed cytoplasmic stress fiber-like structures that co-localized with the smooth muscle cytoskeleton protein smooth muscle alpha-actin. Our works represent the first demonstration of the restricted expression of a class I HDAC to a specific cell type and indicate that HDAC8, besides being a novel marker of smooth muscle differentiation, may play a role in the biology of these contractile cells. PMID- 15277230 TI - Molecular classification of parathyroid neoplasia by gene expression profiling. AB - The current classification of sporadic parathyroid neoplasia, specifically the distinction of adenoma from multiple gland neoplasia (double adenoma and nonfamilial primary hyperplasia) is problematic and results in a relatively high rate of clinical error. Oligonucleotide microarrays (Affymetrix U133A) were used to evaluate parathyroid samples from 61 patients; 35 adenomas, 10 nonfamilial multiple gland neoplasia, 3 familial primary hyperplasia, 8 renal-induced hyperplasia, and 5 from patients without parathyroid disease (normals). A multiclass comparison using supervised clustering identified distinct gene signatures for each class of parathyroid samples. We developed a predictor model that correctly identified 34 of 35 cases of adenoma, 9 of 10 cases of nonfamilial multiple gland neoplasia, and identified a minimum set of 11 genes for the distinction of adenoma versus multiple gland neoplasia. All methods of unsupervised clustering showed two related but different types of parathyroid adenomas that we have arbitrarily designated as type 1 and type 2 adenomas. Multiple gland parathyroid neoplasia, which represents either synchronous or asynchronous autonomous growth in two, three, or all four parathyroid glands, is a distinct molecular entity and does not represent the molecular pathogenesis of adenoma occurring in multiple glands. PMID- 15277231 TI - The 14-3-3 protein epsilon isoform expressed in reactive astrocytes in demyelinating lesions of multiple sclerosis binds to vimentin and glial fibrillary acidic protein in cultured human astrocytes. AB - The 14-3-3 protein family consists of acidic 30-kd proteins expressed at high levels in neurons of the central nervous system. Seven isoforms form a dimeric complex that acts as a molecular chaperone that interacts with key signaling components. Recent studies indicated that the 14-3-3 protein identified in the cerebrospinal fluid of various neurological diseases including multiple sclerosis (MS) is a marker for extensive brain destruction. However, it remains unknown whether the 14-3-3 protein plays an active role in the pathological process of MS. To investigate the differential expression of seven 14-3-3 isoforms in MS lesions, brain tissues of four progressive cases were immunolabeled with a panel of isoform-specific antibodies. Reactive astrocytes in chronic demyelinating lesions intensely expressed beta, epsilon, zeta, eta, and sigma isoforms, among which the epsilon isoform is a highly specific marker for reactive astrocytes. Furthermore, protein overlay, mass spectrometry, immunoprecipitation, and double immunolabeling analysis showed that the 14-3-3 protein interacts with both vimentin and glial fibrillary acidic protein in cultured human astrocytes. These results suggest that the 14-3-3 protein plays an organizing role in the intermediate filament network in reactive astrocytes at the site of demyelinating lesions in MS. PMID- 15277232 TI - Cathepsin K is the principal protease in giant cell tumor of bone. AB - Giant cell tumor (GCT) of bone is a neoplasm of bone characterized by a localized osteolytic lesion. The nature of GCT is an enigma and the cell type(s) and protease(s) responsible for the extensive localized clinicoradiological osteolysis remain unresolved. We evaluated protease expression and cellular distribution of the proteolytic machinery responsible for the osteolysis. mRNA profiles showed that cathepsin K, cathepsin L, and matrix metalloproteinase (MMP) 9 were the preferentially expressed collagenases. Moderate expression was found for MMP-13, MMP-14, and cathepsin S. Specific protease activity assays revealed high cathepsin K activity but showed that MMP-9 was primarily present (98%) as inactive proenzyme. Activities of MMP-13 and MMP-14 were low. Immunohistochemistry revealed a clear spatial distribution: cathepsin K, its associated proton pump V-H(+)-ATPase, and MMP-9 were exclusively expressed in osteoclast-like giant cells, whereas cathepsin L expression was confined to mononuclear cells. To explore a possible role of cathepsin L in osteolysis, GCT derived, cathepsin L-expressing, mononuclear cells were cultured on dentine disks. No evidence of osteolysis by these cells was found. These results implicate cathepsin K as the principal protease in GCT and suggest that osteoclast-like giant cells are responsible for the osteolysis. Inhibition of cathepsin K or its associated proton-pump may provide new therapeutic opportunities for GCT. PMID- 15277233 TI - Vascular gene expression in nonneoplastic and malignant brain. AB - Malignant gliomas are uniformly lethal tumors whose morbidity is mediated in large part by the angiogenic response of the brain to the invading tumor. This profound angiogenic response leads to aggressive tumor invasion and destruction of surrounding brain tissue as well as blood-brain barrier breakdown and life threatening cerebral edema. To investigate the molecular mechanisms governing the proliferation of abnormal microvasculature in malignant brain tumor patients, we have undertaken a cell-specific transcriptome analysis from surgically harvested nonneoplastic and tumor-associated endothelial cells. SAGE-derived endothelial cell gene expression patterns from glioma and nonneoplastic brain tissue reveal distinct gene expression patterns and consistent up-regulation of certain glioma endothelial marker genes across patient samples. We define the G-protein-coupled receptor RDC1 as a tumor endothelial marker whose expression is distinctly induced in tumor endothelial cells of both brain and peripheral vasculature. Further, we demonstrate that the glioma-induced gene, PV1, shows expression both restricted to endothelial cells and coincident with endothelial cell tube formation. As PV1 provides a framework for endothelial cell caveolar diaphragms, this protein may serve to enhance glioma-induced disruption of the blood-brain barrier and transendothelial exchange. Additional characterization of this extensive brain endothelial cell gene expression database will provide unique molecular insights into vascular gene expression. PMID- 15277235 TI - Lack of integrin alpha1beta1 leads to severe glomerulosclerosis after glomerular injury. AB - Severity of fibrosis after injury is determined by the nature of the injury and host genetic susceptibility. Metabolism of collagen, the major component of fibrotic lesions, is, in part, regulated by integrins. Using a model of glomerular injury by adriamycin, which induces reactive oxygen species (ROS) production, we demonstrated that integrin alpha1-null mice develop more severe glomerulosclerosis than wild-type mice. Moreover, primary alpha1-null mesangial cells produce more ROS both at baseline and after adriamycin treatment. Increased ROS synthesis leads to decreased cell proliferation and increased glomerular collagen IV accumulation that is reversed by antioxidants both in vivo and in vitro. Thus, we have identified integrin alpha1beta1 as a modulator of glomerulosclerosis. In addition, we showed a novel pathway where integrin alpha1beta1 modulates ROS production, which in turn controls collagen turnover and ultimately fibrosis. Because integrin alpha1beta1 is expressed in many cell types this may represent a generalized mechanism of controlling matrix accumulation, which has implications for numerous diseases characterized by fibrosis. PMID- 15277234 TI - Loss of LFA-1, but not Mac-1, protects MRL/MpJ-Fas(lpr) mice from autoimmune disease. AB - Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by immune complex-mediated tissue injury. Many different adhesion molecules are thought to participate in the development of SLE; however, few studies have directly examined the contributions of these proteins. Here we demonstrate that LFA-1 plays an essential role in the development of lupus in MRL/MpJ-Fas(lpr) mice. Mice deficient in LFA-1, but not Mac-1, showed significantly increased survival, decreased anti-DNA autoantibody formation, and reduced glomerulonephritis. The phenotype of the LFA-1-deficient mice was similar to that observed in beta(2) integrin-deficient (CD18-null) MRL/MpJ-Fas(lpr) mice, suggesting a lack of redundancy among the beta(2) integrin family members and other adhesion molecules. These studies identify LFA-1 as a key contributor in the pathogenesis of autoimmune disease in this model, and further suggest that therapeutic strategies targeting this adhesion molecule may be beneficial for the treatment of SLE. PMID- 15277236 TI - Indigenous pulmonary Propionibacterium acnes primes the host in the development of sarcoid-like pulmonary granulomatosis in mice. AB - Although many cases of sarcoidosis are self-limiting with spontaneous remission, uncontrolled pulmonary granulomatosis with fibrosis produces intolerable long term respiratory symptoms in a minority of patients. Individuals with chronic pulmonary sarcoidosis require an alternative therapy to corticosteroidal treatment because of its insufficient effectiveness. Although many researchers have considered infection as the triggering factor for this disease, the mechanisms by which the candidate causative organisms induce this disorder remain unclear. We report here that extrapulmonary sensitization to Propionibacterium acnes, which is one of the candidates to date, induced pulmonary Th-1 granulomas mainly in the subpleural and peribronchovascular regions often observed in sarcoidosis. These granulomas appear to be caused by indigenous P. acnes pre existing in the lower respiratory tract of the normal lung, which is believed to be germ-free, and by an influx of P. acnes-sensitized CD4(+) T cells from the circulation. Importantly, the eradication of indigenous P. acnes with antibiotics alleviated the granulomatous lung disease. This is the first report to present clear evidence of the contribution of an indigenous pulmonary bacterium to the formation of granulomatous lesions in the lung. We propose that treatment targeting indigenous P. acnes in the lung may be a possible remedy for pulmonary sarcoidosis. PMID- 15277238 TI - Functional defects in the fanconi anemia pathway in pancreatic cancer cells. AB - Biallelic BRCA2-mutations can cause Fanconi anemia and are found in approximately 7% of pancreatic cancers. Recently, several sequence changes in FANCC and FANCG were reported in pancreatic cancer. Functional defects in the Fanconi pathway can result in a marked hypersensitivity to interstrand crosslinking agents, such as mitomycin C. The functional implications of mutations in the Fanconi pathway in cancer have not been fully studied yet; these studies are needed to pave the way for clinical trials of treatment with crosslinking agents of Fanconi-defective cancers. The competence of the proximal Fanconi pathway was screened in 21 pancreatic cancer cell lines by an assay of Fancd2 monoubiquitination using a Fancd2 immunoblot. The pancreatic cancer cell lines Hs766T and PL11 were defective in Fancd2 monoubiquitination. In PL11, this defect led to the identification of a large homozygous deletion in FANCC, the first cancer cell line found to be FANCC-null. The Fanconi-defective cell lines Hs766T, PL11, and CAPAN1 were hypersensitive to the crosslinking agent mitomycin C and some to cis platin, as measured by cell survival assays and G(2)/M cell-cycle arrest. These results support the practical exploration of crosslinking agents for non-Fanconi anemia patients that have tumors defective in the Fanconi pathway. PMID- 15277237 TI - B Lymphocyte signaling established by the CD19/CD22 loop regulates autoimmunity in the tight-skin mouse. AB - Systemic sclerosis (SSc) is characterized by fibrosis and autoimmmunity. Peripheral blood B cells from SSc patients specifically overexpress CD19, a critical cell-surface signal transduction molecule in B cells. CD19 deficiency in B cells also attenuates skin fibrosis in the tight-skin (TSK/+) mouse, a genetic model for SSc. Herein we analyzed two transgenic mouse lines that overexpress CD19. Remarkably, 20% increase of CD19 expression in mice spontaneously induced SSc-specific anti-DNA topoisomerase I (topo I) antibody (Ab) production, which was further augmented by 200% overexpression. In TSK/+ mice overexpressing CD19, skin thickness did not increase, although anti-topo I Ab levels were significantly augmented, indicating that abnormal CD19 signaling influences autoimmunity in TSK/+ mice and also that anti-topo I Ab does not have a pathogenic role. The molecular mechanisms for abnormal CD19 signaling were further assessed. B-cell antigen receptor crosslinking induced exaggerated calcium responses and augmented activation of extracellular signal-regulated kinase in TSK/+ B cells. CD22 function was specifically impaired in TSK/+ B cells. Consistently, CD19, a major target of CD22-negative regulation, was hyperphosphorylated in TSK/+ B cells. These findings indicate that reduced inhibitory signal provided by CD22 results in abnormal activation of signaling pathways including CD19 in TSK/+ mice and also suggest that this disrupted B cell signaling contribute to specific autoantibody production. PMID- 15277239 TI - Thy-1 expression regulates the ability of rat lung fibroblasts to activate transforming growth factor-beta in response to fibrogenic stimuli. AB - Distinct subpopulations of fibroblasts contribute to lung fibrosis, although the mechanisms underlying fibrogenesis in these subpopulations are not clear. Differential expression of the glycophosphatidylinositol-linked protein Thy-1 affects proliferation and myofibroblast differentiation. Lung fibroblast populations selected on the basis of Thy-1 expression by cell sorting were examined for responses to fibrogenic stimuli. Thy-1 (-) and Thy-1 (+) fibroblast populations were treated with platelet-derived growth factor-BB, interleukin 1beta, interleukin-4, or bleomycin and assessed for activation of transforming growth factor (TGF)-beta, Smad3 phosphorylation, and alpha-smooth muscle actin and fibronectin expression. Thy-1 (-) fibroblasts responded to these stimuli with increased TGF-beta activity, Smad3 phosphorylation, and expression of alpha smooth muscle actin and fibronectin, whereas Thy-1 (+) fibroblasts resisted stimulation. The unresponsiveness of Thy-1 (+) cells is not because of defective TGF-beta signaling because both subsets respond to exogenous active TGF-beta. Rather, Thy-1 (-) fibroblasts activate latent TGF-beta in response to fibrogenic stimuli, whereas Thy-1 (+) cells fail to do so. Defective activation is common to multiple mechanisms of TGF-beta activation, including thrombospondin 1, matrix metalloproteinase, or plasmin. Thy-1 (-) lung fibroblasts transfected with Thy-1 also become resistant to fibrogenic stimulation, indicating that Thy-1 is a critical biological response modifier that protects against fibrotic progression by controlling TGF-beta activation. These studies provide a molecular basis for understanding the differential roles of fibroblast subpopulations in fibrotic lung disease through control of latent TGF-beta activation. PMID- 15277240 TI - Role of interleukin-1 in prion disease-associated astrocyte activation. AB - Prion-induced chronic neurodegeneration has a substantial inflammatory component, and the activation of glia cells may play an important role in disease development and progression. However, the functional contribution of cytokines to the development of the gliosis in vivo was never systematically studied. We report here that the expression of interleukin-1beta (IL-1beta), IL-1beta converting enzyme, and IL-1 receptor type 1 (IL-1RI) is up-regulated in a murine scrapie model. The scrapie-induced gliosis in IL-1RI(-/-) mice was characterized by an attenuated activation of astrocytes in the asymptomatic stage of the disease and a reduced expression of CXCR3 ligands. Furthermore, the accumulation of the misfolded isoform of the prion protein PrP(Sc) was significantly delayed in the IL-1RI(-/-) mice. These observations indicate that IL-1 is a driver of the scrapie-associated astrocytosis and possibly the accompanying amyloid deposition. In addition, scrapie-infected IL-1RI-deficient (IL-1RI(-/-)) mice showed a delayed disease onset and significantly prolonged survival times suggesting that an anti-inflammatory therapeutical approach to suppress astrocyte activation and/or glial IL-1 expression may help to delay disease onset in established prion infections of the central nervous system. PMID- 15277244 TI - Interspecific variation of plant traits associated with resistance to herbivory among four species of Ficus (moraceae). AB - BACKGROUND AND AIMS: To understand the defensive characteristics of interspecies varieties and their responses to herbivory damage, four species of Ficus plants (Ficus altissima, F. auriculata, F. racemosa and F. hispida) were studied. They were similar in life form, but differed in successional stages. Of these, Ficus altissima is a late successional species, F. hispida is a typical pioneer and F. auriculata and F. racemosa are intermediate successional species. We addressed the following questions: (1) What is the difference in plant traits among the four species and are these traits associated with differences in herbivory damage levels? (2) What is the difference in the damage-induced changes among the four species? METHODS: Herbivory damage was measured in the field on randomly planted seedlings of the four species of the same age. Defences to herbivory were also tested by feeding leaves of the four species to larvae of Asota caricae in the laboratory. A total of 14 characters such as water content, thickness, toughness, pubescence density on both sides, leaf expansion time, lifetime and the contents of total carbon (C), nitrogen (N), phosphorous (P), potassium (K), magnesium (Mg) and calcium (Ca) were measured. Leaf calcium oxalate crystal (COC) density, total Ca and N content, leaf toughness and height were measured to investigate induced responses to artificial herbivory among the four species. KEY RESULTS: and conclusions Herbivory damage in the four studied species varied greatly. The pioneer species, F. hispida, suffered the most severe herbivory damage, while the late successional species, F. altissima, showed the least damage. A combination of several characteristics such as high in content of N, Ca and P and low in leaf toughness, lifetime and C : N ratio were associated with increased herbivore damage. The late successional species, F. altissima, might also incorporate induced defence strategies by means of an increase in leaf COC and toughness. PMID- 15277242 TI - Modulation of notch signaling elicits signature tumors and inhibits hras1-induced oncogenesis in the mouse mammary epithelium. AB - Deregulation of Notch signaling, which normally affects a broad spectrum of cell fates, has been implicated in various neoplastic conditions. Here we describe a transgenic mouse model, which demonstrates that expression of a constitutively active form of the Notch1 receptor in the mammary epithelium induces the rapid development of pregnancy/lactation-dependent neoplasms that consistently exhibit a characteristic histopathological pattern. These signature tumors retain the ability to respond to apoptotic stimuli and regress on initiation of mammary gland involution, but eventually appear to progress in subsequent pregnancies to nonregressing malignant adenocarcinomas. Additionally, we present evidence indicating that cyclin D1 is an in vivo target of Notch signals in the mammary glands and demonstrate that we can effectively inhibit Hras1-driven, cyclin D1 dependent mammary oncogenesis by transgenic expression of the Notch antagonist Deltex. PMID- 15277243 TI - Effect of drought stress on lipid metabolism in the leaves of Arabidopsis thaliana (ecotype Columbia). AB - BACKGROUND AND AIMS: Cell membranes are major targets of environmental stresses. Lipids are important membrane components, and changes in their composition may help to maintain membrane integrity and preserve cell compartmentation under water stress conditions. The aim of this work was to investigate the effects of water stress on membrane lipid composition and other aspects of lipid metabolism in the leaves of the model plant, Arabidopsis thaliana. METHODS: Arabidopsis thaliana (ecotype Columbia) plants were submitted to progressive drought stress by withholding irrigation. Studies were carried out in plants with hydration levels ranging from slight to very severe water deficit. Enzymatic activities hydrolysing MGDG, DGDG and PC were measured. Expression of several genes essential to lipid metabolism, such as genes coding for enzymes involved in lipid biosynthesis (MGDG synthase, DGDG synthase) and degradation (phospholipases D, lipoxygenase, patatin-like lipolytic-acylhydrolase), was studied. KEY RESULTS: In response to drought, total leaf lipid contents decreased progressively. However, for leaf relative water content as low as 47.5 %, total fatty acids still represented 61 % of control contents. Lipid content of extremely dehydrated leaves rapidly increased after rehydration. The time-course of the decrease in leaf lipid contents correlated well with the increase in lipolytic activities of leaf extracts and with the expression of genes involved in lipid degradation. Despite a decrease in total lipid content, lipid class distribution remained relatively stable until the stress became very severe. CONCLUSIONS: Arabidopsis leaf membranes appeared to be very resistant to water deficit, as shown by their capacity to maintain their polar lipid contents and the stability of their lipid composition under severe water loss conditions. Moreover, arabidopsis displayed several characteristics indicative of a so far unknown adaptation capacity to drought-stress at the cellular level, such as an increase in the DGDG : MGDG ratio and fatty acid unsaturation. PMID- 15277241 TI - Mice deficient in glutathione transferase zeta/maleylacetoacetate isomerase exhibit a range of pathological changes and elevated expression of alpha, mu, and pi class glutathione transferases. AB - Glutathione transferase zeta (GSTZ1-1) is the major enzyme that catalyzes the metabolism of alpha-halo acids such as dichloroacetic acid, a carcinogenic contaminant of chlorinated water. GSTZ1-1 is identical with maleylacetoacetate isomerase, which catalyzes the penultimate step in the catabolic pathways for phenylalanine and tyrosine. In this study we have deleted the Gstz1 gene in BALB/c mice and characterized their phenotype. Gstz1(-/-) mice do not have demonstrable activity with maleylacetone and alpha-halo acid substrates, and other GSTs do not compensate for the loss of this enzyme. When fed a standard diet, the GSTZ1-1-deficient mice showed enlarged liver and kidneys as well as splenic atrophy. Light and electron microscopic examination revealed multifocal hepatitis and ultrastructural changes in the kidney. The addition of 3% (w/v) phenylalanine to the drinking water was lethal for young mice (<28 days old) and caused liver necrosis, macrovesicular steatosis, splenic atrophy, and a significant loss of circulating leukocytes in older surviving mice. GSTZ1-1 deficient mice showed constitutive induction of alpha, mu, and pi class GSTs as well as NAD(P)H:quinone oxidoreductase 1. The overall response is consistent with the chronic accumulation of a toxic metabolite(s). We detected the accumulation of succinylacetone in the serum of deficient mice but cannot exclude the possibility that maleylacetoacetate and maleylacetone may also accumulate. PMID- 15277245 TI - Relationships among vernalization, shoot apex development and frost tolerance in wheat. AB - BACKGROUND AND AIMS: Frost tolerance of wheat depends primarily upon a strong vernalization requirement, delaying the transition to the reproductive phase. The aim of the present study was to learn how saturation of the vernalization requirement and apical development stage are related to frost tolerance in wheat. METHODS: 'Mironovskaya 808', a winter variety with a long vernalization requirement, and 'Leguan', a spring variety without a vernalization requirement, were acclimated at 2 degrees C at different stages of development. Plant development (morphological stage of the shoot apex), vernalization requirement (days to heading) and frost tolerance (survival of the plants exposed to freezing conditions) were evaluated. KEY RESULTS: 'Mironovskaya 808' increased its frost tolerance more rapidly; it reached a higher level of tolerance and after a longer duration of acclimation at 2 degrees C than was found in 'Leguan'. The frost tolerance of 'Mironovskaya 808' decreased and its ability to re-acclimate a high tolerance was lost after saturation of its vernalization requirement, but before its shoot apex had reached the double-ridge stage. The frost tolerance of 'Leguan' decreased after the plants had reached the floret initiation stage. CONCLUSIONS: The results support the hypothesis that genes for vernalization requirement act as a master switch regulating the duration of low temperature induced frost tolerance. In winter wheat, due to a longer vegetative phase, frost tolerance is maintained for a longer time and at a higher level than in spring wheat. After the saturation of vernalization requirement, winter wheat (as in spring wheat) established only a low level of frost tolerance. PMID- 15277246 TI - Genetic structure and outcrossing rates in Flourensia cernua (Asteraceae) growing at different densities in the South-western Chihuahuan Desert. AB - BACKGROUNDS AND AIMS: Flourensia cernua is a partially self-incompatible, wind pollinated shrub that grows in two scrub types of contrasting densities. It was anticipated that differences in plant density would affect the amount of genotype availability, and thus higher outcrossing rates and less genetic differentiation would be found at high-density sites. METHODS: At five high-density sites and at five low-density sites, 11 allozyme loci were analysed in adults. Outcrossing rates were estimated using five allozyme loci sampled from eight families from each scrub type. KEY RESULTS: High levels of genetic variation were found at all sites (ranging from P = 82-100 %, He = 0.33-0.45, and Ho = 0.4-0.59). Heterozygotes were found in excess (FIS = -0.15 +/- 0.06 s.d.), suggesting that natural selection favours heterozygosity, and there was little differentiation between sites (FST = 0.08 +/- 0.02 s.d.). Life history attributes, such as long lived habit and wide geographic distribution, as well as the presence of a self incompatibility system may explain these results. Outcrossing rates did not differ from 1.0 in both scrub types, and there was no genetic differentiation between scrub types (FST = -0.01 +/- 0.004 s.d.). CONCLUSIONS: The high rate of outcrossing favoured by partial incompatibility may generate unrestricted gene flow between scrub types and thus may explain the lack of differentiation between them. High heterozygosity could be expected in long-lived plants of arid zones as they confront a variable and stressing environment. PMID- 15277249 TI - Intra-inflorescence variation in floral traits and reproductive success of the hermaphrodite Silene acutifolia. AB - BACKGROUND AND AIMS: Intraspecific variation in floral components and reproductive success is often located at the intra-individual level. The arrangement of flowers within inflorescences may explain a great deal of this variation. The variation in number of ovules, fruit set, number of seeds per fruit, seed set, seed weight and seed germination is investigated at different positions within the inflorescence of Silene acutifolia. METHODS: Data were obtained in natural populations, and germination experiments were conducted in a germination chamber. KEY RESULTS: The number of ovules, fruit set, number of seeds, seed set and seed weight, showed a significant decline from early (primary) position to later (tertiary) position. The patterns of intra inflorescence variation were consistent in different populations and years of study. Seed germination showed an opposite pattern, seeds from primary position showed the lowest germination percentages and seeds from tertiary position the highest, although the effect of position on germination was only marginally significant. There was significant among-population variation in number of ovules per flower. Fruit set also varied significantly among populations, with lower fruit set in the smaller and more isolated population. No significant among population differences were detected in number of seeds per fruit and seed set. Seeds from the smallest and more isolated population (Arnado) were the lightest. Seed germination showed strong differences between populations, seeds from Arnado started to germinate later, and showed the lowest final germination percentages. CONCLUSIONS: Architectural effects or resource competition are the most commonly proposed hypothesis to explain these patterns. Data suggest that there is less pollen available to pollinate tertiary flowers, and that there is not enough outcross pollen in Arnado. The germination percentages suggest that there is variation in the source of pollen within inflorescences, with high probability of receiving outcross pollen in flowers from primary position, and higher probability of geitonogamous crosses in tertiary flowers. PMID- 15277247 TI - Soluble inorganic tissue phosphorus and calcicole-calcifuge behaviour of plants. AB - BACKGROUND AND AIMS: Natural and semi-natural, non-fertilized calcareous soils are consistently low in soluble and easily exchangeable phosphate. An over utilization, or possibly an immobilization, of inorganic P in the tissues of calcifuge plants may take place, if such plants are forced to grow on a calcareous soil, though this has not been experimentally demonstrated. The objectives of this study are, therefore, to elucidate if calcifuge plants, when forced to develop on a calcareous soil, not only have lower total P (Ptot) concentrations in their leaves than calcicole plants grown on such soil, but also a lower proportion of Ptot as water-soluble, inorganic phosphate. Such differences may be of importance in understanding the calcicole-calcifuge behaviour of plants. MATERIALS AND METHODS: Plants of five calcicole and five calcifuge herbs and three calcicole and three calcifuge grasses were cultivated in a glasshouse on a moderately acid Cambisol and a calcareous Rendzic Leptosol using seeds of wild populations from southern Sweden. The calcifuges were: Corynephorus canescens, Deschampsia flexuosa, Holcus mollis, Digitalis purpurea, Lychnis viscaria, Rumex acetosella, Scleranthus annuus and Silene rupestris. The calcicoles were: Melica ciliata, Phleum phleoides, Sesleria caerulea, Arabis hirsuta, Sanguisorba minor, Scabiosa columbaria, Silene uniflora ssp. petraea and Veronica spicata. KEY RESULTS: At harvest, calcifuges had much lower leaf tissue concentrations of Ptot and Pi than calcicoles when grown on the calcareous soil, and biomass production of the calcifuges was poor on this soil. Moreover, the calcifuge herbs had, on average, a lower proportion of their Ptot as Pi than had the calcicole herbs. The calcifuge herbs were also unable to avoid excessive uptake of Ca from the calcareous soil. The calcifuge grasses maintained a similar proportion of Ptot as Pi as the calcicole grasses, but their growth was still poor on the calcareous soil. CONCLUSIONS: On calcareous soil, very little Pi in the tissues of calcifuge herbs is, at any time, available for use in various physiological functions. This is of importance to their photosynthesis, growth, competition and final survival on such soils. PMID- 15277248 TI - Variation in floral sex allocation in Polygonatum odoratum (Liliaceae). AB - BACKGROUND AND AIMS: It is well known that resource allocation to male and female functions can be highly variable in hermaphroditic plants. The purpose of this study was to investigate variations in sexual investment at different levels (flower, plant and population) in Polygonatum odoratum, a plant with sequentially opening flowers. METHODS: Pollen and ovule production in base, middle and top flowers of P. odoratum flowering shoots from two natural populations were quantified. Plant measurements of phenotypic and functional gender were calculated in both populations. Total leaf number was used to investigate the relationship between gender assessments and plant size. KEY RESULTS: Pollen and ovule production varied depending on flower position, although the precise pattern differed between both studied populations; only investment in female floral function decreased markedly from base to top flowers in both populations. The frequency distribution of phenotypic gender and their relationship with plant size differed between populations. Phenotypic and functional gender were correlated in both populations. CONCLUSIONS: Sexual investment in P. odoratum has shown a marked variability within plants, among plants, and between populations, which confirms the importance of analysing sex expression in plants of this type. Differences in relative investment in male and female components (phenotypic gender) are reflected in the functional gender and it would be expected that the evolution of sexual specialization in Polygonatum odoratum would be promoted. PMID- 15277250 TI - Day-night changes of energy-rich compounds in crassulacean acid metabolism (CAM) species utilizing hexose and starch. AB - BACKGROUND AND AIMS: Plants with crassulacean acid metabolism (CAM) can be divided into two groups according to the major carbohydrates used for malic acid synthesis, either polysaccharide (starch) or monosaccharide (hexose). This is related to the mechanism and affects energy metabolism in the two groups. In Kalanchoe pinnata and K. daigremontiana, which utilize starch, ATP-dependent phosphofructokinase (tonoplast inorganic pyrophosphatase) activity is greater than inorganic pyrophosphate-dependent phosphofructokinase (tonoplast adenosine triphosphatase) activity, but the reverse is the case in pineapple (Ananas comosus) utilizing hexose. To test the hypothesis that the energy metabolism of the two groups differs, day-night changes in the contents of ATP, ADP, AMP, inorganic phosphate (Pi), phosphoenolpyruvate (PEP) and inorganic pyrophosphate (PPi) in K. pinnata and K. daigremontiana leaves and in pineapple chlorenchyma were analysed. METHODS: The contents of energy-rich compounds were measured spectrophotometrically in extracts of tissue sampled in the light and dark, using potted plants, kept for 15 d before the experiments in a growth chamber. KEY RESULTS: In the three species, ATP content and adenylate energy charge (AEC) increased in the dark and decreased in the light, in contrast to ADP and AMP. Changes in ATP and AEC were greater in Kalanchoe leaves than in pineapple chlorenchyma. PPi content in the three species increased in the dark, but on illumination it decreased rapidly and substantially, remaining little changed through the rest of the light period. Pi content of Kalanchoe leaves did not change between dark and light, whereas Pi in pineapple chlorenchyma increased in the dark and decreased in the light, and the changes were far greater than in Kalanchoe leaves. Light-dark changes in PEP content in the three species were similar. CONCLUSIONS: These results corroborate our hypothesis that day-night changes in the contents of energy-rich compounds differ between CAM species and are related to the carbohydrate used for malic acid synthesis. PMID- 15277252 TI - Endometrial cancer: a frequent orphan disease. PMID- 15277251 TI - Contribution of root cap mucilage and presence of an intact root cap in maize (Zea mays) to the reduction of soil mechanical impedance. AB - BACKGROUND AND AIMS: The impedance to root growth imposed by soil can be decreased by both mucilage secretion and the sloughing of border cells from the root cap. The aim of this study is to quantify the contribution of these two factors for maize root growth in compact soil. METHODS: These effects were evaluated by assessing growth after removing both mucilage (treatment I -- intact) and the root cap (treatment D -- decapped) from the root tip, and then by adding back 2 micro L of mucilage to both intact (treatment IM -- intact plus mucilage) and decapped (treatment DM -- decapped plus mucilage) roots. Roots were grown in either loose (0.9 Mg m(-3)) or compact (1.5 Mg m(-3)) loamy sand soils. Also examined were the effects of decapping on root penetration resistance at three soil bulk densities (1.3, 1.4 and 1.5 Mg m(-3)). KEY RESULTS: In treatment I, mucilage was visible 12 h after transplanting to the compact soil. The decapping and mucilage treatments affected neither the root elongation nor the root widening rates when the plants were grown in loose soil for 12 h. Root growth pressures of seminal axes in D, DM, I and IM treatments were 0.328, 0.288, 0.272 and 0.222 MPa, respectively, when the roots were grown in compact soil (1.5 Mg m(-3) density; 1.59 MPa penetrometer resistance). CONCLUSIONS: The contributions of mucilage and presence of the intact root cap without mucilage to the lubricating effect of root cap (percentage decrease in root penetration resistance caused by decapping) were 43 % and 58 %, respectively. The lubricating effect of the root cap was about 30 % and unaffected by the degree of soil compaction (for penetrometer resistances of 0.52, 1.20 and 1.59 MPa). PMID- 15277253 TI - Thalidomide in cancer medicine. AB - Thalidomide, an oral agent with antiangiogenic and immunomodulatory properties, is being investigated extensively in the management of advanced cancer. Multiple studies with large numbers of patients have confirmed that this drug has significant activity in multiple myeloma. Some patients with myelofibrosis or myeodysplatic syndromes may reduce their need for transfusions after thalidomide treatment. The activity of thalidomide in solid tumors is less prominent. Studies in Kaposi's sarcoma, malignant melanoma, renal cell carcinoma and prostate cancer appear more promising especially when thalidomide is combined with biological agents or with chemotherapy. Limited activity was demonstrated in patients with glioma, while thalidomide appears to be inactive in patients with head and neck cancer, breast or ovarian cancer. PMID- 15277254 TI - Primary chemotherapy for breast cancer: the evidence and the future. AB - The first generation of randomised trials assessing the role of primary chemotherapy in breast cancer has failed to demonstrate the expected survival benefit. However, it has established the role of this treatment in 'downstaging' tumours of patients with locally advanced disease and, consequently, in improving breast conservation rates. Also, a number of surrogates of outcome have been identified, which will hopefully lead to earlier results in breast cancer clinical trials. Encouraging results have also been reported in trials investigating a number of novel approaches. PMID- 15277255 TI - Phase III randomized trial of doxorubicin + cisplatin versus doxorubicin + 24-h paclitaxel + filgrastim in endometrial carcinoma: a Gynecologic Oncology Group study. AB - BACKGROUND: This study was performed to determine whether 24-h paclitaxel plus doxorubicin and filgrastim was superior to cisplatin plus doxorubicin in patients with endometrial cancer with respect to response, progression-free survival (PFS) and overall survival (OS). PATIENTS AND METHODS: Eligible chemotherapy-naive patients were randomly assigned to doxorubicin 60 mg/m2 intravenously (i.v.) followed by cisplatin 50 mg/m2 i.v. (arm 1, n=157) or doxorubicin 50 mg/m2 i.v. followed 4 h later by paclitaxel 150 mg/m2 i.v. over 24 h plus filgrastim 5 microg/kg on days 3-12 (arm 2, n=160). Starting doses were reduced for prior pelvic radiotherapy and age > 65 years. Both regimens were to be repeated every 3 weeks for a maximum of seven cycles. RESULTS: There was no significant difference in response rate (40% versus 43%), PFS (median 7.2 versus 6 months) or OS (median 12.6 versus 13.6 months) for arm 1 and arm 2, respectively. Toxicities were primarily hematological, with 54% (arm 1) and 50% (arm 2) of patients experiencing grade 4 granulocytopenia. Gastrointestinal toxicities were similar in both arms. CONCLUSIONS: Doxorubicin and 24-h paclitaxel plus filgrastim was not superior to doxorubicin and cisplatin in terms of response, PFS or survival in advanced endometrial cancer. PMID- 15277256 TI - Patterns of failure, prognostic factors and survival in locoregionally advanced head and neck cancer treated with concomitant chemoradiotherapy: a 9-year, 337 patient, multi-institutional experience. AB - BACKGROUND: Locoregionally advanced, stage IV head and neck cancer has traditionally carried a poor prognosis. We sought to assess changes in patterns of failure, prognostic factors for recurrence, and overall outcome, using two different strategies of chemoradiotherapy conducted in prospective, multi institutional phase II trials. PATIENTS AND METHODS: Three hundred and thirty seven stage IV patients were treated from 1989 to 1998. We compared locoregional and distant recurrence rates, overall survival and progression-free survival from two different treatment strategies: intensive induction chemotherapy followed by split-course chemoradiotherapy (type 1, n=127), or intensified, split-course, hyperfractionated multiagent chemoradiotherapy alone (type 2, n=210). Univariate and multivariate analyses of 12 chosen covariates were assessed separately for the two study types. RESULTS: The pattern of failure varied greatly between study types 1 and 2 (5-year locoregional failure of 31% and 17% for study types 1 and 2, respectively, P=0.01; 5-year distant failure rate of 13% and 22% for study types 1 and 2, P=0.03). Combined 5-year overall survival was 47% [95% confidence interval (CI) 41% to 53%) and progression-free survival was 60% (95% CI 55% to 66%). Both treatment strategies yielded similar survival rates. Poor overall survival and distant recurrence were best predicted by advanced nodal stage. Locoregional recurrence was extremely rare for patients with T0-T3 tumor stage, regardless of lymph-node stage. CONCLUSIONS: This analysis suggests that pattern of failure in primary head and neck cancer may be dependent upon treatment strategy. Randomized clinical trials of induction chemotherapy are warranted as a means to determine if a decrease in distant metastases can lead to an increase in survival rates in the setting of effective chemoradiotherapy for locoregional control. Additionally, this analysis provides impetus for randomized clinical trials of organ preservation chemoradiotherapy in sites outside the larynx and hypopharynx. PMID- 15277257 TI - Phase II study of the farnesyl transferase inhibitor R115777 in patients with sensitive relapse small-cell lung cancer. AB - BACKGROUND: R115777 (tipifarnib, Zarnestra) is a farnesyl transferase inhibitor that blocks the farnesylation of proteins involved in signal transduction pathways critical for cell proliferation and survival. This multicenter phase II study was conducted to determine the efficacy, tolerability and pharmacokinetics of R115777 in patients with relapsed small-cell lung cancer (SCLC). PATIENTS AND METHODS: Patients who had a partial or complete response to their initial chemotherapy regimen, followed by at least 3 months off treatment before relapse (sensitive relapse) were eligible. R115777 was administered in 3-week cycles at a dose of 400 mg orally twice daily for 14 consecutive days followed by 7 days off treatment. RESULTS: Twenty-two patients were enrolled. The median progression free survival was 1.4 months and median overall survival was 6.8 months. Non hematological toxicities were predominantly grade 1-2 and included nausea (64%) and fatigue (60%). Grade 3-4 granulocytopenia and thrombocytopenia occurred in 27% and 23% of patients, respectively. Febrile neutropenia was not observed. Pharmacokinetic studies demonstrated peak plasma concentrations of R115777 2.6 4.5 h after oral dosing and no significant drug accumulation. The trial was terminated because no objective responses were observed in 20 patients evaluable for response. CONCLUSIONS: R115777 showed no significant antitumor activity as a single agent in sensitive-relapse SCLC. PMID- 15277258 TI - Single nucleotide polymorphisms and outcome in docetaxel-cisplatin-treated advanced non-small-cell lung cancer. AB - BACKGROUND: Platinum-based doublets are the standard chemotherapy for advanced non-small-cell lung cancer (NSCLC). Excision-repair cross-complementing 1 (ERCC1), xeroderma pigmentosum group D (XPD) and ribonucleotide reductase subunit M1 (RRM1) are essential to the repair of cisplatin DNA adducts. Multidrug resistance 1 (MDR1) has been related to antimicrotubule resistance. We assessed whether single nucleotide polymorphisms (SNPs) in ERCC1, XPD, RRM1 and MDR1, and ERCC1 mRNA expression, predicted survival in docetaxel-cisplatin-treated stage IV NSCLC patients. PATIENTS AND METHODS: Using the TaqMan 5' nuclease assay, we examined ERCC1 118, XPD 751 and 312, RRM1 -37C/A, and MDR1 C3435T SNPs in peripheral blood lymphocytes (PBLs) obtained from 62 docetaxel-cisplatin-treated advanced NSCLC patients. ERCC1 expression was measured in RNA isolated from PBLs using real-time reverse transcriptase PCR. RESULTS: Overall median survival was 10.26 months. Median survival was 9.67 months for 34 patients with ERCC1 118 C/T, 9.74 months for 17 patients with T/T, and not reached for 11 patients with C/C (P=0.04). Similar significant differences in time to progression were observed according to ERCC1 118 genotype (P=0.03). No other significant differences were observed. CONCLUSIONS: Patients homozygous for the ERCC1 118 C allele demonstrated a significantly better survival. ERCC1 SNP assessment could be an important component of tailored chemotherapy trials. PMID- 15277259 TI - Combination of irinotecan (CPT-11) plus oxaliplatin (L-OHP) as first-line treatment in locally advanced or metastatic gastric cancer: a multicentre phase II trial. AB - BACKGROUND: The purpose of this study was to evaluate the efficacy and tolerance of irinotecan (CPT-11) in combination with oxaliplatin (L-OHP) as first-line treatment in patients with locally advanced or metastatic gastric cancer (AGC). PATIENTS AND METHODS: Thirty-two patients with AGC who had not received previous therapy for metastatic disease were enrolled. The median age was 62.5 years and the World Health Organization performance status was 0-1 in 29 patients; 13 (40.6%) patients had previous surgery and three (9.4%) had adjuvant chemotherapy. L-OHP (85 mg/m2 as a 2-h i.v. infusion) followed by CPT-11 (200 mg/m2 as a 30-min i.v. infusion) was given on day 1, in cycles of 21 days. RESULTS: All patients were evaluable for toxicity and 31 were evaluable for response. Complete response was achieved in one (3.1%) patient and a partial response was achieved in 15 (46.9%) [overall response rate = 50% (95% confidence interval 38.7-72.4%)]. Eight (25%) patients had stable disease, and eight (25%) had progressive disease. The median duration of response was 5 months and the median time to disease progression was 5.5 months. After a median follow-up period of 16 months, the median survival time was 8.5 months. Grade 3-4 neutropenia occurred in six (18.6%) patients, febrile neutropenia in two (6.2%) and grade 3 anaemia in one (3.1%). Grade 3 diarrhoea was observed in two (6.2%) patients, grade 1 neurotoxicity in five (15.6%) and grade 3 asthenia in two (6.2%). There was no treatment-related death. CONCLUSIONS: The combination of CPT-11/L-OHP is an active regimen as front-line treatment in AGC with a favourable toxicity profile and deserves further evaluation in randomised studies. PMID- 15277260 TI - Oxaliplatin reintroduction in patients previously treated with leucovorin, fluorouracil and oxaliplatin for metastatic colorectal cancer. AB - BACKGROUND: FOLFOX, a bimonthly combination of leucovorin, 5-fluorouracil and oxaliplatin, is active in metastatic colorectal cancer, but sometimes causes cumulative sensory neurotoxicity. This retrospective study investigated FOLFOX reintroduction after a break in treatment or following disease progression on another regimen. PATIENTS AND METHODS: FOLFOX was reintroduced in 29 patients. During their previous FOLFOX therapy, 24 had achieved a response, four were stable and one had progression. Median progression-free survival (PFS) was 33 weeks. Grade 3 neuropathy developed in nine and grade 2 neuropathy in eight patients. RESULTS: Following FOLFOX reintroduction, six patients (21%) showed a response, 15 (52%) were stable and eight (28%) had progression. Median PFS was 18 weeks. Grade 3 neuropathy developed in four patients and grade 2 neuropathy in 11. Two patients with previous grade 3 neuropathy had no recurrence of neuropathy after eight and 18 cycles, respectively. Among 13 patients who received no treatment between periods of FOLFOX therapy, four (31%) had a response and eight (62%) had stable disease. CONCLUSION: Reintroduction of oxaliplatin was feasible and achieved a response or stabilization in 73% of patients. These results support the concept of intensified, repeated short courses of FOLFOX, a strategy currently being evaluated prospectively in the OPTIMOX study. PMID- 15277261 TI - Anthracycline-based chemotherapy as primary treatment for intravascular lymphoma. AB - BACKGROUND: Optimal therapeutic management of intravascular lymphoma (IVL) lacks precise guidelines. PATIENTS AND METHODS: The clinico-pathological features of 38 HIV-negative patients with IVL were reviewed to define efficacy of chemotherapy in these malignancies. Clinical characteristics of 22 patients treated with chemotherapy and of 16 untreated patients were compared in order to understand better the impact and causes of potential patient selection. RESULTS: Median age was 70 years (range 34-90), with a male/female ratio of 0.9; 23 (61%) patients had Eastern Cooperative Oncology Group performance status (ECOG-PS) > 1; 21 (55%) had systemic symptoms. Cutaneous lesions and anemia were significantly more common among patients treated with chemotherapy; central nervous system (CNS) and renal involvement were significantly more common among untreated patients. Chemotherapy was associated with a response rate of 59% and a 3-year overall survival of 33 +/- 11%. Five of six patients with CNS involvement received chemotherapy: four of them died early; only one patient, treated with adriamycin, cyclophosphamide, vincristine, methotrexate, bleomycin and prednisolone (MACOP-B) followed by high-dose chemotherapy and autologous stem cell transplantation (ASCT), was alive at 19 months. High-dose chemotherapy supported by ASCT was indicated at diagnosis in another patient (43 years of age, stage I), who was alive at 71 months, and at relapse after cyclophosphamide, doxorubicin, vincristine and prednisone (CHOP) in two patients who died early after transplantation. PS < or = 1, disease limited to the skin, stage I, and use of chemotherapy were independently associated with better outcome. CONCLUSIONS: Anthracycline-based chemotherapy is the standard treatment for IVL. However, survival is disappointing, with a relevant impact of diagnostic delay and lethal complications. More intensive combinations, containing drugs with higher CNS bioavailability, are needed in cases with brain involvement, and the role of high dose chemotherapy supported by ASCT should be further investigated in younger patients with unfavorable features. PMID- 15277263 TI - Gemtuzumab ozogamicin (MylotargTM) is infrequently associated with sinusoidal obstructive syndrome/veno-occlusive disease. AB - BACKGROUND: Gemtuzumab ozogamicin (GO) is approved for the treatment of older adults with acute myeloid leukemia in first relapse. Several reports have suggested an association between GO administration and hepatic veno-occlusive disease (VOD), which has recently been termed sinusoidal obstructive syndrome (SOS). However, the majority of these studies were done in patients who had undergone high-dose therapy with stem cell transplantation or when GO was administered in combination with other cytotoxic chemotherapy. PATIENTS AND METHODS: We performed a retrospective review of all patients treated at our institution with single-agent GO, either as initial therapy or in the relapsed and refractory setting. All patients were planned to receive GO 9 mg/m2 in two doses, 14 days apart. We reviewed liver function tests before and after administration and analyzed hepatic injuries in the context of patients' other comorbid conditions. Patients were classified as experiencing liver toxicity if their liver function(s) abnormality lasted for > 7 days, as documented by repeated serum studies. RESULTS: Forty-seven patients were analyzed. Response rate (27.2%) and median duration of response (6 months) were comparable to other reports. All patients were assessable for liver toxicity, of which 23 (48%) had elevation of at least one of their liver function tests (alanine aminotransferase, aspartate aminotransferase, total bilirubin or alkaline phosphatase). Elevations in liver function test(s) were noted at a median of 14 days (range 7-175 days). Eight patients had other comorbid conditions that could explain their liver abnormality, making the incidence of direct GO-induced liver injury 31%. However, only one patient had radiographic and clinical evidence suggesting SOSVOD. CONCLUSIONS: When administered using the recommended dose and schedule, GO has little association with VODSOS if given as a single agent. In this retrospective review, the incidence of GO-related SOSVOD is as low as 2%. PMID- 15277262 TI - Outcome and prognostic factors in advanced Hodgkin's disease treated with high dose chemotherapy and autologous stem cell transplantation: a study of 341 patients. AB - BACKGROUND: The reported probability of survival of patients with Hodgkin's disease (HD) following high-dose chemotherapy with autologous stem cell transplantation (HDC/ASCT) is 35-65% at 5 years. The Polish Lymphoma Research Group investigated retrospectively prognostic factors for overall survival (OS) and event-free survival (EFS), and the risk of secondary malignancies in a large series of patients who underwent HDC/ASCT. PATIENTS AND METHODS: The data of 341 consecutive patients treated in 10 centers from 1990 to 2002 were collected and analyzed. RESULTS: The actuarial 5-year OS and EFS were 64% [95% confidence interval (CI) 57% to 71%] and 45% (95% CI 39% to 51%), respectively. In the multivariate model, unfavorable prognostic factors for EFS were less than partial response at the time of ASCT [relative risk (RR), 2.92 (95% CI 1.68-5.08); P<0.001] and three or more previous chemotherapy lines (RR, 2.16; 95% CI 1.42 3.30; P<0.001). These two factors were also associated with unfavorable OS (RR, 3.32; 95% CI 1.90-5.79; P<0.001 and RR, 2.34, 95% CI 1.51-3.64; P<0.001). Five year cumulative risk of secondary malignancy was 8.4% (95% CI 2% to 13%) and the only identified risk factor was splenectomy (P=0.02). CONCLUSIONS: HDC/ASCT should be considered early in the course of disease for patients with a response after standard therapy. PMID- 15277264 TI - Physical performance, depression, immune status and fatigue in patients with hematological malignancies after treatment. AB - BACKGROUND: Fatigue is a frequent and severe problem after treatment of patients with hematological malignancies. This symptom has been associated with anemia, reduced physical performance, mood, endocrine disorders and impaired nutritional status. Recently, it has been suggested that fatigue can be related to a persistent activation of the immune system with increased production of proinflammatory cytokines. However, there is no conclusive evidence regarding the role of the immune system in the origin of fatigue in cancer patients. PATIENTS AND METHODS: We evaluated the correlation of fatigue with thyroid function, markers of immune activity [interleukin (IL)-1alpha, IL-1 soluble receptor, IL-6, C-reactive protein and neopterin], liver and kidney function, mood and physical ability in 71 patients with hematological malignancies. All patients had been free of relapse and not received treatment (chemotherapy, radiotherapy or immune modulators) for at least 3 months. RESULTS: Fatigue was related to depression (r=0.84; P<0.0001) and reduced performance status (r=-0.61; P<0.0001). However, there was no correlation between fatigue and thyroid, liver and kidney function, anemia, albumin concentration or markers of immune activity (all r-values <0.20; P>0.05). CONCLUSIONS: We conclude that fatigue in relapse-free patients with hematological malignancies is associated with depressive mood and reduced physical performance, but not with impairment of thyroid function, anemia or persistent activation of the immune system. PMID- 15277265 TI - Recognition of distress and psychiatric morbidity in cancer patients: a multi method approach. AB - BACKGROUND: This study aimed to determine the prevalence of psychiatric morbidity and distress among 189 consecutively recruited cancer patients upon admission to surgical oncology wards, and to investigate the recognition of distressed patients by medical staff. PATIENTS AND METHODS: Assessment consisted of a diagnostic psychiatric interview (SCID, DSM-IV), patient-reported distress using a standardised questionnaire (Hospital Anxiety and Depression Scale), and physicians' and nurses' estimates of patients' distress. Twenty-eight per cent of patients were assigned a psychiatric diagnosis, with adjustment disorder predominating. RESULTS: Surgeons accurately recognised marked distress in 77% of patients with a psychiatric disorder and nurses did so in 75%. Because of low specificity, the positive predictive value was only 39% in surgeons and 40% in nurses. However, recognition of distress translated into referral to the psychosocial liaison service for only a minor proportion of distressed patients. CONCLUSIONS: Since a remarkable proportion of distressed patients remained unrecognised by the medical staff, only systematic screening of patients upon admission allows timely support to those who are most in need. PMID- 15277266 TI - A prospective study on lung toxicity in patients treated with gemcitabine and carboplatin: clinical, radiological and functional assessment. AB - BACKGROUND: Small series and retrospective studies have suggested that treatment with gemcitabine may be associated with pulmonary toxicity. However, a prospective evaluation of cancer patients treated with gemcitabine-based chemotherapy without neoplastic involvement of the thorax and without administration of radiotherapy has not been performed. PATIENTS AND METHODS: To investigate this issue, 41 consecutive patients receiving gemcitabine and carboplatin underwent prospective evaluation of lung function, which included pulmonary symptoms, pulmonary function tests, arterial blood gases and radiographic studies. Assessment was performed before and after completion of chemotherapy in all patients. Patients with a substantial decline in diffusion capacity for carbon monoxide (DLCO), defined as a drop of > or = 20%, were reassessed 2 months later. RESULTS: After chemotherapy, there were no significant changes in forced vital capacity (FVC), forced expiratory volume in 1 s (FEV1), FEV1/FVC ratio, alveolar volume or total lung capacity. In contrast, there was a significant decline in DLCO (73 +/- 22 versus 67 +/- 24% predicted; P = 0.017) and in carbon monoxide transfer coefficient (KCO) (89 +/- 24 versus 80 +/- 24% predicted; P = 0.004). Arterial blood gases did not change following treatment. Ten of the 41 patients (24%) exhibited a substantial decline in DLCO, which, however, recovered within 2 months (DLCO at baseline, immediately after therapy and at 2 months after completion of treatment, 84 +/- 14, 58 +/- 16 and 77 +/- 17% predicted, respectively; P < 0.001; baseline DLCO versus DLCO at 2 months, P > 0.05). Four of the 41 patients (10%) experienced dyspnea, which was self limiting, with the exception of one patient who developed interstitial lung fibrosis. Among the various risk factors examined, older age, female gender and lower baseline DLCO were associated with more profound changes in DLCO post treatment. CONCLUSIONS: This prospective analysis showed that the combination of gemcitabine and carboplatin induces a significant, but reversible, decrease in diffusion capacity, which is mostly asymptomatic. Thus, this regimen is safe as regards clinically significant lung toxicity. PMID- 15277267 TI - Hypertension as a risk factor for glioma? Evidence from a population-based study of comorbidity in glioma patients. AB - BACKGROUND: Little is known about the aetiology of glioma. Research is often hampered by the low incidence and high mortality of the disease. Concomitant diseases in glioma patients may indicate possible aetiological pathways. We therefore studied comorbidity in glioma patients. PATIENTS AND METHODS: We performed a case-control study using population-based data from the Eindhoven Cancer Registry. We compared prevalences of concomitant diseases in 510 glioma patients with two reference cancer populations from the same registry. RESULTS: Compared with all other cancer patients, a significantly higher prevalence of hypertension was found in glioma patients for age categories 60-74 years [odds ratio (OR) 1.37; 95% confidence interval (CI) 1.02-1.84] and 75+ years (OR 2.37; 95% CI 1.34-4.21). The association was most pronounced in elderly men and in astrocytic glioma, with a maximum in age category 75+ years (OR 5.86; 95% CI 2.20 15.7). The prevalence of cerebrovascular disease was higher in glioma patients >45 years old (OR 1.67; 95% CI 1.12-2.47), whereas the prevalence of other cancers was lower (OR 0.64; 95% CI 0.48-0.87). No consistent associations were detected for several other concomitant diseases. CONCLUSIONS: Our data suggest an association between hypertension and glioma, although questions remain about causality and the possible mechanisms. We hypothesise that this association is mediated through potentially neurocarcinogenic effects of antihypertensive medication. PMID- 15277268 TI - Circulating angiogenic factor levels correlate with extent of disease and risk of recurrence in patients with soft tissue sarcoma. AB - BACKGROUND: Tumor angiogenesis, or new blood vessel formation, is regulated by a balance between pro-angiogenic factors such as vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF), and anti-angiogenic factors such as endostatin. PATIENTS AND METHODS: To investigate this angiogenic balance in soft tissue sarcomas (STS), blood samples were collected from 76 STS patients and 15 healthy controls, and analyzed for VEGF, bFGF and endostatin using quantitative enzyme-linked immunosorbent assays (ELISA). RESULTS: Forty-one patients (54%) had primary tumors, 20 (26%) had local recurrences and 15 (20%) had metastatic disease with or without local disease. Levels of all three angiogenic factors were highly variable in STS patients. Mean levels of VEGF and bFGF were 12 and 14 times higher, respectively, in patients compared with controls (P<0.0001). VEGF levels correlated with size of tumor, with the highest levels found in tumors >10 cm in size. Patients with metastases had endostatin levels 45% lower than patients without metastases (P=0.047). In 54 patients who underwent resection of primary disease or local recurrence, low pre-operative bFGF level was associated with a higher risk of subsequent recurrence (P=0.044). CONCLUSIONS: STS secrete widely variable levels of angiogenic factors, and levels of specific factors may correlate with extent of disease, predict risk of recurrence and possibly guide the use of anti-angiogenic agents. PMID- 15277269 TI - The difference between study recommendations, stated policy, and actual practice in a clinical trial. AB - BACKGROUND: We determined whether physicians involved in a clinical trial adhere to the study recommendations or the stated policy of their treatment centre with respect to the administration of boost radiation after breast conserving surgery. PATIENTS AND METHODS: Boost radiation treatment policy was determined by survey at 25 oncology centres involved in a randomised trial of breast or breast plus nodal radiation in Canada. Actual practice was compared with stated policy and study recommendations. RESULTS: Among 248 subjects, 201 (81%) were treated according to stated policy [kappa=0.40, 95% confidence intervals (CI) 0.27-0.52; P<0.0001], indicating only a fair to moderate agreement between stated and actual practice, while 232 (94%) were treated according to study recommendations (kappa=0.59, 95% CI 0.40-0.77; P<0.0001), indicating moderate to near substantial agreement between study recommendations and actual practice (P=0.88 for z-test of difference). In a multivariate analysis, subjects who had invasive disease at a resection margin were more likely to get a boost than those with margins clear of invasive tumour by 2 mm [odds ratio (OR) 49, 95% CI 7.6-322; P<0.0001]. CONCLUSIONS: Physicians appear compliant with study recommendations for a non randomised manoeuvre in a clinical trial, possibly at the expense of compliance with stated local policy. Clinical trial protocols should incorporate standard practice. PMID- 15277270 TI - A phase I trial of a Bcl-2 antisense (G3139) and weekly docetaxel in patients with advanced breast cancer and other solid tumors. AB - PURPOSE: Expression of the Bcl-2 protein confers resistance to various apoptotic signals. G3139 [oblimersen sodium (Genasense)] is a phosphorothioate antisense oligodeoxynucleotide that targets Bcl-2 mRNA, downregulates Bcl-2 protein translation, and enhances the antitumor effects of subtherapeutic doses of docetaxel (Taxotere). PATIENTS AND METHODS: We performed a phase I trial to determine the maximum tolerated dose (MTD) and safety profile of combined therapy with G3139 and weekly docetaxel in patients with advanced Bcl-2-positive solid tumors. Cohorts of three to six patients were enrolled to escalating doses of G3139 and a fixed dose of weekly docetaxel using either of two schedules. In part I, G3139 was administered by continuous infusion for 21 days (D1-22), and docetaxel (35 mg/m2) was given weekly on days 8, 15 and 22. In part II, G3139 was given by continuous infusion for 5 days before the first weekly dose of docetaxel, and for 48 h before the second and third weekly docetaxel doses. For both schedules, cycles were repeated every 4 weeks. RESULTS: Twenty-two patients were enrolled. Thirteen patients were treated on the part I schedule with doses of G3139 escalated from 1 to 4 mg/kg/day. Nine patients were on the part II schedule of shorter G3139 infusion at G3139 doses of 5-9 mg/kg/day. Hematologic toxicities were mild, except for one case of persistent grade 3 thrombocytopenia in part I. The most common adverse events were cumulative fatigue and transaminase elevation, which prevented further dose escalation beyond 4 mg/kg/day for 21 days with the part I schedule. In part II of the study, using the abbreviated G3139 schedule, even the highest daily doses were tolerated without dose-limiting toxicity or the need for dose modification. Objective tumor response was observed in two patients with breast cancer, including one whose cancer previously progressed on trastuzumab plus paclitaxel. Four patients had stable disease. Pharmacokinetic results for G3139 were similar to those of other trials. CONCLUSIONS: G3139 in combination with standard-dose weekly docetaxel was well tolerated. The shortened and intermittent G3139 infusion had less cumulative toxicities and still allowed similar total G3139 delivery as the longer infusion. Further studies should examine the molecular effect of the regimen, as well as clinical activities in malignancies for which taxanes are indicated. PMID- 15277272 TI - Oxaliplatin in non-seminomatous germ-cell tumors. PMID- 15277271 TI - A phase I clinical and pharmacokinetic study of the camptothecin glycoconjugate, BAY 38-3441, as a daily infusion in patients with advanced solid tumors. AB - BACKGROUND: The aim of this study was to define the maximum tolerated dose (MTD), dose-limiting toxicity (DLT) and pharmacokinetics of the camptothecin glycoconjugate BAY 38-3441, administered as an infusion for 30 min on two separate schedules every 3 weeks. PATIENTS AND METHODS: A total of 81 patients with advanced solid tumors were treated with BAY 38-3441 either at doses of 20, 40, 67, 100, 140, 210, 315, 470 and 600 mg/m2/day for 1 day every 3 weeks (single dose schedule), or at doses of 126, 189, 246, 320 and 416 mg/m2/day once daily for three consecutive days every 3 weeks (3-day schedule). Plasma sampling was performed to characterize the pharmacokinetics of BAY 38-3441 and camptothecin with these schedules. RESULTS: DLTs included renal toxicity, granulocytopenia and thrombocytopenia on the single-day schedule at doses > or = 470 mg/m2/day, and diarrhea and thrombocytopenia on the 3-day schedule at doses > or = 320 mg/m2/day. Other non-DLTs were gastrointestinal, dermatological and hematological. Pharmacokinetics of BAY 38-3441 and camptothecin appear to be dose dependent, but not linear. CONCLUSIONS: Renal toxicity was dose-limiting for BAY 38-3441 using 30-min infusions on the single-dose schedule. Dose escalation to 470 mg/m2/day is feasible using a 2-h infusion. However, because of the superior safety profile, we recommend the 3-day schedule for BAY 38-3441 at a dose of 320 mg/m2/day as 30-min infusions for further phase II studies. PMID- 15277273 TI - Neomycin as secondary prophylaxis for irinotecan-induced diarrhea. PMID- 15277274 TI - Unusual extramedullary relapse of CML. PMID- 15277276 TI - A new editorial team for Archives of Internal Medicine: the choices we make. PMID- 15277275 TI - Rituximab-CHOP or 2-weekly CHOP + G-CSF in aggressive lymphoma of the elderly? PMID- 15277277 TI - Gastroesophageal reflux disease, Barrett esophagus, and esophageal adenocarcinoma. AB - The incidence of esophageal adenocarcinoma has been rising rapidly over the past few decades. The major risk factors predisposing to the development of adenocarcinoma are long-standing gastroesophageal reflux disease and Barrett esophagus, but other factors may be involved as cancer can occur in their absence. In patients with Barrett esophagus, the extent and degree of dysplasia influence the risk of esophageal adenocarcinoma. As neither medical nor surgical therapies have been proven to prevent adenocarcinoma, endoscopic screening of patients with chronic reflux and endoscopic surveillance of patients diagnosed with Barrett esophagus are usually performed in an effort to detect adenocarcinomas at earlier stages. The evidence supporting strategies in the management of patients with gastroesophageal reflux and Barrett esophagus is presented. PMID- 15277278 TI - A practical and evidence-based approach to cardiovascular disease risk reduction. AB - Implementation of the numerous lifestyle and medical management options for secondary prevention of cardiovascular disease remains a daunting goal for primary care physicians and cardiologists alike. Despite the existence of expert consensus guidelines on cardiovascular prevention by the American College of Cardiology and the American Heart Association, therapies known to improve patient care and decrease morbidity and mortality remain underutilized. This review attempts to simplify cardiovascular risk reduction by summarizing key clinical trials in an "ABC" format. We believe that if health care providers and patients use such a format, important lifestyle and pharmacologic options will more likely be addressed. PMID- 15277279 TI - Resuscitating advance directives. AB - Advance directives have not fulfilled their promise of facilitating decisions about end-of-life care for incompetent patients. Many legal requirements and restrictions concerning advance directives are counterproductive. Requirements for witnessing or notarizing advance directives make it difficult for patients to complete a written directive during a physician visit. State laws that establish a hierarchy of family surrogates for incompetent patients who have not appointed a proxy are inflexible and may not apply to common clinical situations. Advance directives would be more useful if they emphasized discussing end-of-life care with physicians rather than completing a legal document. State laws should be revised to encourage patients to discuss advance directives with physicians and to complete them during an office visit. Such patient-physician discussions about end-of-life care can lead to more informed patient decisions. Procedures for written advance directives should be simplified. Patients should be able to designate health care proxies through oral statements to physicians. These reforms will encourage discussions between patients and physicians about advance directives and may lead to more informed decisions near the end of life. PMID- 15277281 TI - Social support and its relationship to morbidity and mortality after acute myocardial infarction: systematic overview. AB - Among the commonly understood socioeconomic determinants of health, social change, disorganization, and poverty have been associated with an increased risk of morbidity and mortality. One of the postulated mechanisms through which these determinants have been linked to health and illness is their relationship to social support. The health determinant, social isolation or lack of a social support network (SSN), and its effects on premature mortality after acute myocardial infarction mandate further scrutiny by the cardiovascular community for several reasons. First, as a predictor of 1-year mortality, low SSN is equivalent to many of the classic risk factors, such as elevated cholesterol level, tobacco use, and hypertension. Second, treatment of acute myocardial infarction is costly. Because low social support is associated with an increased 1-year mortality, neglecting the role of the SSN may diminish the possible gains accrued during acute-phase treatment. Therefore, lack of an SSN should be considered a risk factor for subsequent morbidity and mortality after a myocardial infarction. Finally, cardiac rehabilitation programs and other extant prevention strategies can be better used to reduce mortality after myocardial infarction. This article systematically reviews recent evidence related to SSNs to provide an update on the role of social support in cardiovascular disease related outcomes. PMID- 15277280 TI - Timing in the communication of pain among nursing home residents, nursing staff, and clinicians. AB - BACKGROUND: The management of nursing home (NH) residents' pain requires adequate nursing assessment and clinician knowledge of pain therapies. However, the timely communication of pain from residents to nurses and from nurses to clinicians is equally necessary. Using a 4-step model (nursing assessment of pain, notification of clinicians regarding pain assessment, clinicians' assessment of pain and intervention), and nursing reassessment following an intervention, we describe the timing with which each of these steps occur. METHODS: In a telephone survey of directors of nursing from 63 of the 68 nursing homes in New Haven County, Connecticut, we determined (1) how often nurses assess pain in residents, (2) when nurses notify clinicians about residents' pain, (3) how often clinicians assess pain, and (4) when nurses reassess pain after a clinician's intervention. RESULTS: Whereas in 76% of NHs nurses assessed pain in residents without pain at least "quarterly," only in 46% of NHs was pain assessed in residents with pain at least "every shift." In 42% of NHs nurses notified clinicians at least when the regimen was "ineffective." Only 55% of directors of nursing reported that clinicians assessed pain at least every 30 to 60 days. Finally, in 73% of NHs nursing reassessment occurred at least 1 hour after intervention. CONCLUSIONS: There is considerable variability in how frequently nurses and clinicians assess pain, when clinicians are notified about pain, and how frequently nurses reassess pain. Studies are needed to determine optimal timing in the communication process of pain to allow better pain management outcomes and quality of care for NH residents. PMID- 15277282 TI - Lifetime nonnarcotic analgesic use and decline in renal function in women. AB - BACKGROUND: Analgesics are commonly used and may impair kidney function. However, limited prospective information is available on the long-term effects of aspirin, other nonsteroidal anti-inflammatory drugs (NSAIDs), and acetaminophen on renal function. METHODS: A total of 1697 women participating in the Nurses' Health Study provided information on a mailed questionnaire in 1999 about lifetime use of acetaminophen, aspirin, and NSAIDs and provided blood samples in 1989 and 2000. The main outcome was change in estimated glomerular filtration rate (GFR) in 11 years. Multivariate logistic regression was used to determine the odds of developing the outcome according to lifetime analgesic intake. RESULTS: The mean +/- SD estimated GFR decreased from 88 +/- 17 to 79 +/- 17 mL/min per 1.73 m(2). There were no substantial differences in the unadjusted or estimated GFR levels among the categories of lifetime intake for the 3 analgesic groups at baseline or after 11 years. Acetaminophen use was associated with an increased risk of a GFR decline of at least 30 mL/min per 1.73 m(2) (P trend =.01) and a GFR decline of 30% or greater (P trend<.001), but aspirin and NSAID use were not. Compared with women consuming less than 100 g of acetaminophen, multivariate-adjusted odds ratio (95% confidence intervals) for a decline in GFR of at least 30 mL/min per 1.73 m(2) for women consuming more than 3000 g was 2.04 (1.28-3.24). CONCLUSIONS: Higher lifetime use of aspirin and NSAIDs is not associated with renal function decline, but high acetaminophen use may increase the risk of loss of renal function. The absolute risk of renal function decline due to even high lifetime analgesic intake seems to be modest. PMID- 15277283 TI - National trends in osteoporosis visits and osteoporosis treatment, 1988-2003. AB - BACKGROUND: Research is limited on physicians' prescribing practices for osteoporosis treatment. We investigated patterns of pharmacotherapy from 1988 to 2003 and the impact of new medications on identification and treatment of patients with osteoporosis. METHODS: We tracked trends from 1988 through 2003 in the frequency of osteoporosis visits and patterns of pharmacotherapy associated with these visits using nationally representative data on prescribing patterns by office-based US physicians from the IMS HEALTH National Disease and Therapeutic Index. RESULTS: The number of physician visits for osteoporosis increased 4-fold between 1994 (1.3 million visits) and 2003 (6.3 million visits), whereas it had remained stable in prior years. This increase coincided with the availability of oral daily bisphosphonates and the selective estrogen receptor modulator raloxifene. The annualized percentage of osteoporosis visits where medications were prescribed increased from 82% in 1988 to 97% by 2003. Prior to 1994, the leading choices for osteoporosis therapy were calcium and estrogens, with lesser roles played by calcitonins and bisphosphonates. Between 1994 and 2003, the percentage of visits where bisphosphonates and raloxifene were prescribed increased from 14% to 73% and from 0% to 12%, respectively, while prescriptions for other medications declined. CONCLUSIONS: New medications for osteoporosis offering improved efficacy and convenient dosing were associated with increased frequency of patient visits and treatment. This finding suggests that new drug therapy contributed to increased disease recognition and treatment. PMID- 15277284 TI - Difficult end-of-life treatment decisions: do other factors trump advance directives? AB - BACKGROUND: Advance directives are widely promoted as a means to plan for patients' decisional incapacity, yet there is little evidence of their effectiveness. We devised a study to assess physicians' compliance with hypothetical advance directives and further examine their clinical reasoning. METHODS: The study consisted of an analysis of a mailed written survey containing 6 hypothetical cases of seriously ill patients. Each case contained an explicit advance directive with potential conflict between the directive and (1) prognosis, (2) wishes of family or friends, or (3) quality of life. Data were collected on the clinical treatment decisions made by physicians and the reasons for those decisions. Study participants were all internal medicine faculty and resident physicians from a single academic institution. RESULTS: A total of 47% analyzable surveys (117/250) were returned. Decisions by faculty and residents were not consistent with the advance directive in 65% of cases. This inconsistency was similar for faculty and residents (68% and 61%, respectively; P>.05). When physicians made decisions inconsistent with the advance directive, they were more likely to list reasons other than the directive for their decisions (89%; P<.001). CONCLUSIONS: Internists frequently made treatment decisions that were not consistent with an explicit advance directive. In difficult clinical situations, internists appear to consider other factors such as prognosis, perceived quality of life, and the wishes of family or friends as more determinative than the directive. Future work needs to explore the generalizability of these findings and examine how strictly patients desire their advance directives to be followed. PMID- 15277285 TI - The protective effect of habitual tea consumption on hypertension. AB - BACKGROUND: Tea has long been believed to possess hypotensive effects in popular Chinese medicine. However, conflicting results have been shown among human trials and animal studies on the relation between tea consumption and blood pressure. Epidemiological evidence about the long-term effect of tea on hypertensive risk is also inconsistent. METHODS: We examined the effect of tea drinking, measured in detail for the past decades, on the risk of newly diagnosed hypertension in 1507 subjects (711 men and 796 women), 20 years or older, who did not have a hypertensive history during 1996 in Taiwan. RESULTS: Six hundred subjects (39.8%) were habitual tea drinkers, defined by tea consumption of 120 mL/d or more for at least 1 year. Compared with nonhabitual tea drinkers, the risk of developing hypertension decreased by 46% for those who drank 120 to 599 mL/d and was further reduced by 65% for those who drank 600 mL/d or more after carefully adjusting for age, sex, socioeconomic status, family history of hypertension, body mass index, waist-hip ratio, lifestyle factors (total physical activity, high sodium intake, cigarette smoking, alcohol consumption, and coffee drinking), and dietary factors (vegetable, fruit, unrefined grain, fish, milk, visible-fat food, and deep fried food intake). However, tea consumption for more than 1 year was not associated with a further reduction of hypertension risk. CONCLUSION: Habitual moderate strength green or oolong tea consumption, 120 mL/d or more for 1 year, significantly reduces the risk of developing hypertension in the Chinese population. PMID- 15277286 TI - Twenty-one-year trends in the use of inferior vena cava filters. AB - BACKGROUND: Improved inferior vena cava (IVC) filters have led to liberalization of the indications for insertion. Increased use, however, has been followed with a potential for unwarranted insertion. There are only sparse data on trends in the use of IVC filters in patients with pulmonary embolism (PE), patients with deep venous thrombosis (DVT) alone, and patients at high risk. We analyzed the National Hospital Discharge Survey (NHDS) database for such trends. METHODS: We used data from the NHDS, which is based on a national probability sample of discharges from short-stay nonfederal hospitals in 50 states and the District of Columbia. The numbers of sampled patients with DVT, PE, and IVC filters were determined from the International Classification of Diseases, Ninth Revision, Clinical Modification codes at discharge. RESULTS: The number of patients who had IVC filters increased from 2000 in 1979 to 49 000 in 1999. In 1999, 45% of IVC filter insertions were in patients with DVT alone, 36% were in patients with PE, and 19% were in patients who presumably were at high risk but did not have DVT or PE listed as a discharge code. The use of IVC filters was more frequent in northeastern states than in western states (P =.01). CONCLUSIONS: The use of IVC filters increased markedly during the last 2 decades in patients with PE, patients with DVT alone, and patients at risk who had neither PE nor DVT. Randomized controlled trials may lead to improved risk stratification and limit the number of unnecessary filter insertions. PMID- 15277287 TI - Uric acid level as a risk factor for cardiovascular and all-cause mortality in middle-aged men: a prospective cohort study. AB - BACKGROUND: Despite abundant epidemiologic evidence, the role of elevated serum uric acid level as a cardiovascular risk factor is controversial. We assessed the predictive value of serum uric acid levels for cardiovascular and overall mortality. METHODS: A population-based prospective cohort study was performed of 1423 middle-aged Finnish men initially without cardiovascular disease, cancer, or diabetes. The main outcome measure was death from cardiovascular disease and any cause. RESULTS: The mean follow-up was 11.9 years. There were 157 deaths during follow-up, of which 55 were cardiovascular. In age-adjusted analyses, serum uric acid levels in the upper third were associated with a greater than 2.5-fold higher risk of death from cardiovascular disease than levels in the lower third. Taking into account cardiovascular risk factors and variables commonly associated with gout increased the relative risk to 3.73. Further adjustment for factors related to the metabolic syndrome strengthened the risk to 4.77. Excluding the 53 men using diuretics did not alter the results. In age-adjusted analyses, men with serum uric acid levels in the upper third were 1.7-fold more likely to die of any cause than men with levels in the lower third. Adjustment for further risk factors strengthened the association somewhat. CONCLUSIONS: Serum uric acid levels are a strong predictor of cardiovascular disease mortality in healthy middle-aged men, independent of variables commonly associated with gout or the metabolic syndrome. Serum uric acid measurement is an easily available and inexpensive risk marker, but whether its relationship to cardiovascular events is circumstantial or causal remains to be answered. PMID- 15277288 TI - Randomized trials of vitamin E in the treatment and prevention of cardiovascular disease. AB - BACKGROUND: Observational epidemiological studies consistently show that individuals who choose to take high amounts of vitamin E through diet or supplements experience cardiovascular benefits, for which basic research provides plausible mechanisms. However, because the size of the postulated benefit is small to moderate, the confounding inherent in observational studies is as great as the effect size. Before the availability of randomized evidence, about 1 in 4 adults was taking vitamin E supplements in the United States. METHODS: We conducted a computerized search of the English-language literature from 1990 to the present and found 7 large-scale randomized trials of the effectiveness vitamin E in the treatment and prevention of cardiovascular disease. Data were available on myocardial infarction, stroke, or cardiovascular death. RESULTS: Six of the 7 trials showed no significant effect of vitamin E on cardiovascular disease. In an overview, vitamin E had neither a statistically significant nor a clinically important effect on any important cardiovascular event (odds ratio [OR], 0.98; 95% confidence interval [CI], 0.94-1.03) or its components: nonfatal myocardial infarction (OR, 1.00; 95% CI, 0.92-1.09), nonfatal stroke (OR, 1.03; 95% CI, 0.93-1.14), or cardiovascular death (OR, 1.00; 95% CI, 0.94-1.05). CONCLUSIONS: The ORs and CIs provide strong support for a lack of statistically significant or clinically important effects of vitamin E on cardiovascular disease. The use of agents of proven lack of benefit, especially those easily available over the counter, may contribute to underuse of agents of proven benefit and failure to adopt healthy lifestyles. PMID- 15277289 TI - The effect of excessive anticoagulation on mortality and morbidity in hospitalized patients with anticoagulant-related major hemorrhage. AB - BACKGROUND: We aimed to determine the effect of excessive anticoagulation on morbidity and mortality in hospitalized patients with major anticoagulant associated hemorrhage. METHODS: We prospectively identified 101 consecutive inpatients admitted to Brigham and Women's Hospital with major bleeding occurring during administration of warfarin sodium, unfractionated heparin (UFH), or low molecular-weight heparin (LMWH). RESULTS: Fifty patients had excessive and 51 had nonexcessive anticoagulation. The overall mortality at 60 days was 26% (13/50) in the excessive group compared with 10% (5/51) in the nonexcessive group (P =.03). Excessive warfarin therapy was associated with an increased 60-day mortality (P =.049), in contrast to excessive anticoagulation with UFH or LMWH alone (P =.27) or UFH or LMWH as a "bridge" to warfarin therapy (P =.10). Multivariate regression identified excessive anticoagulation as an independent predictor of 60 day mortality (adjusted hazard ratio [HR], 4.17; 95% confidence interval [CI], 1.39-12.49; P =.01), along with intracranial hemorrhage (adjusted HR, 6.16; 95% CI, 1.75-21.67; P =.005) and active cancer (adjusted HR, 3.79; 95% CI, 1.13 12.70; P =.03). Excessive anticoagulation was also a significant predictor of the combined nonfatal end point of stroke, myocardial infarction, hypotension, critical anemia, and surgical or angiographic intervention at 30 days (HR, 2.17; 95% CI, 1.25-3.78; P =.006). CONCLUSION: In a cohort of patients with anticoagulation-associated hemorrhage, excessive anticoagulation contributed independently to increased morbidity and mortality. PMID- 15277291 TI - Benzodiazepine use and hip fractures in the elderly: who is at greatest risk? AB - BACKGROUND: It remains unclear whether benzodiazepine use increases hip fracture incidence. We studied this relationship in a large cohort, controlling for multiple potential confounders. METHODS: We analyzed 42 months of New Jersey Medicaid health care claims data for all enrollees. Each eligible person-day was assigned to categories of benzodiazepine exposure and categories of other predictors, based on prior and current medication dispensing and diagnosis information. Hip fractures were identified based on hospital claims with primary discharge diagnosis International Classification of Diseases, Ninth Revision (ICD 9) codes 820.xx. RESULTS: Cohort members (n = 125 203) contributed 194 071 person years and had 2312 eligible hip fractures. After adjustment for age, sex, race, Medicaid nursing home residence, exposure to other psychoactive medications, including antiparkinsonian medications, diagnoses of epilepsy and dementia, and hospitalization in the previous 6 months, the incidence rate of hip fracture was significantly higher compared with no benzodiazepine use for exposure to any benzodiazepine (incidence rate ratio [IRR], 1.24; 95% confidence interval [CI], 1.06-1.44), to a short half-life, high-potency benzodiazepine (IRR, 1.27; 95% CI, 1.01-1.59), during the first 2 weeks after starting a benzodiazepine (IRR, 2.05; 95% CI, 1.28-3.28), during the second 2 weeks after starting a benzodiazepine (IRR, 1.88; 95% CI, 1.15-3.07), and for continued use (IRR, 1.18; 95% CI, 1.03 1.35). CONCLUSIONS: The incidence of hip fracture appears to be associated with benzodiazepine use. Contrary to several previous studies, short half-life benzodiazepines are not safer than long half-life benzodiazepines. Hip fracture risk is highest during the first 2 weeks after starting a benzodiazepine and declines thereafter. PMID- 15277290 TI - Laboratory tests to determine the cause of hypokalemia and paralysis. AB - BACKGROUND: Hypokalemia and paralysis may be due to a short-term shift of potassium into cells in hypokalemic periodic paralysis (HPP) or due to a large deficit of potassium in non-HPP. Failure to make a distinction between HPP and non-HPP may lead to improper management. Therefore, we evaluated the diagnostic value of spot urine tests in patients with hypokalemia and paralysis during 3 years. METHODS: Before therapy, the urine potassium concentration, potassium creatinine ratio, and transtubular potassium concentration gradient were determined in a second voided urine sample. RESULTS: Forty-three patients with hypokalemia and paralysis were identified: 30 had HPP and 13 had non-HPP. There was no significant difference in the plasma potassium or bicarbonate concentrations and in the pH of arterial blood between the 2 groups. All but 2 patients in the non-HPP group had urine potassium concentration values less than 20 mmol/L. Although the potassium concentration was significantly lower in the HPP group, there was some overlap. In contrast, the transtubular potassium concentration gradient and potassium-creatinine ratio differentiated patients with HPP vs non-HPP. Although only a mean +/- SD of 63 +/- 36 mmol of potassium chloride was administered in the patients with HPP, rebound hyperkalemia (>5 mmol/L) occurred in 19 (63%) of these 30 patients. CONCLUSIONS: Calculating the transtubular potassium concentration gradient and potassium-creatinine ratio provided a simple and reliable test to distinguish HPP from non-HPP. Minimal potassium chloride supplementation should be given to avoid rebound hyperkalemia in patients with HPP. PMID- 15277292 TI - The quality of care plans for patients with do-not-resuscitate orders. AB - BACKGROUND: Care plans for patients with do-not-resuscitate (DNR) orders often fail to define limits other than cardiopulmonary resuscitation and fail to address other patient care needs. We studied the explicitness and comprehensiveness of care plans for patients with DNR orders and what factors were associated with this aspect of the quality of their care. METHODS: A cross sectional study was conducted at Georgetown University Medical Center (GUMC), Washington, DC, and St Vincent Catholic Medical Centers (SVCMC), St Vincent's Hospital-Manhattan, New York, NY. Participants included 189 consecutive medical inpatients with DNR orders. RESULTS: A previously validated medical chart review technique termed concurrent care concerns (CCCs) measured whether 11 possible patient care needs had been addressed within 2 days of the DNR order. Reasons for the DNR order were documented in only 55% of cases, and a consent conversation was documented in only 69%. The mean number of total CCCs per DNR order was 1.55 (1.84 at GUMC and 1.29 at SVCMC; (P =.007). In a multivariate logistic regression analysis of low (or=2) CCCs, patients with malignancy (P =.002), higher APACHE III (Acute Physiology and Chronic Health Evaluation III) scores (P =.007), and a documented consent conversation (P =.009) and those at Georgetown (P =.005) were more likely to have high attention to CCCs. Patients with dementia were the least likely to have high attention to CCCs. CONCLUSIONS: Documented consent conversations and care plans for patients with DNR orders are less than ideal. Care plans differ in quality by diagnosis, institution, and whether or not a consent conversation is documented. These observations might help to guide interventions that aim to improve the care of patients with DNR orders. PMID- 15277293 TI - Religion, spirituality, and acute care hospitalization and long-term care use by older patients. AB - BACKGROUND: The impact of religion and spirituality on acute care hospitalization (ACH) and long-term care (LTC) in older patients before, during, and after ACH is not well known. METHODS: Patients 50 years or older consecutively admitted to the general medical service at Duke University Medical Center were interviewed shortly after admission (N = 811). Measures of religiosity were organized religious activity (ORA), nonorganizational religious activity (NORA), religiosity through religious radio and/or television (RTV), intrinsic religiosity, and self-rated religiousness. Measures of spirituality included self rated spirituality and daily spiritual experiences (DSE). Primary outcome was number of ACH days during an average 21-month observation period. Secondary outcomes were times hospitalized and number of days spent in a nursing home or rehabilitation setting (collectively, long-term care: LTC). Race and sex interactions were examined. RESULTS: In the cross-sectional analysis, ORA was the only religious variable related to fewer ACH days and fewer hospitalizations, an effect that is fully explained by physical health status and that disappeared when examined prospectively. The number of LTC days was inversely related to NORA, RTV, and DSE, effects that were partially explained by social support but not by severity of medical illness. Interactions with race and sex were notable but reached statistical significance only among African Americans and women. In those groups, religious and/or spiritual characteristics also predicted future LTC use independent of physical health and baseline LTC status. CONCLUSIONS: Relationships with ACH were weak, were confined to ORA only, and disappeared in prospective analyses. However, robust and persistent effects were documented for religiousness and/or spirituality in the use of LTC among African Americans and women. PMID- 15277294 TI - Dispersal of HIV positive asylum seekers: national survey of UK healthcare providers. PMID- 15277295 TI - Hypnosis for pain relief in labour and childbirth: a systematic review. AB - BACKGROUND: In view of widespread claims of efficacy, we examined the evidence regarding the effects of hypnosis for pain relief during childbirth. METHODS: Medline, Embase, Pubmed, and the Cochrane library 2004.1 were searched for clinical trials where hypnosis during pregnancy and childbirth was compared with a non-hypnosis intervention, no treatment or placebo. Reference lists from retrieved papers and hypnotherapy texts were also examined. There were no language restrictions. Our primary outcome measures were labour analgesia requirements (no analgesia, opiate, or epidural use), and pain scores in labour. Suitable comparative studies were included for further assessment according to predefined criteria. Meta-analyses were performed of the included randomized controlled trials (RCTs), assessed as being of "good" or "adequate" quality by a predefined score. RESULTS: Five RCTs and 14 non-randomized comparisons (NRCs) studying 8395 women were identified where hypnosis was used for labour analgesia. Four RCTs including 224 patients examined the primary outcomes of interest. One RCT rated poor on quality assessment. Meta-analyses of the three remaining RCTs showed that, compared with controls, fewer parturients having hypnosis required analgesia, relative risk=0.51 (95% confidence interval 0.28, 0.95). Of the two included NRCs, one showed that women using hypnosis rated their labour pain less severe than controls (P<0.01). The other showed that hypnosis reduced opioid (meperidine) requirements (P<0.001), and increased the incidence of not requiring pharmacological analgesia in labour (P<0.001). CONCLUSION: The risk/benefit profile of hypnosis demonstrates a need for well-designed trials to confirm the effects of hypnosis in childbirth. PMID- 15277297 TI - High-frequency oscillatory ventilation and an interventional lung assist device to treat hypoxaemia and hypercapnia. AB - A male patient accidentally aspirated paraffin oil when performing as a fire eater. Severe acute respiratory distress syndrome (Pa(o(2))/Fi(o(2)) ratio 10.7 kPa) developed within 24 h. Conventional pressure-controlled ventilation (PCV) with high airway pressures and low tidal volumes failed to improve oxygenation. Hypercapnia (Pa(co(2)) 12 kPa) with severe acidosis (pH<7.20) ensued. Treatment with high-frequency oscillatory ventilation (HFOV) and a higher adjusted airway pressure (35 cm H(2)O) improved the Pa(o(2))/Fi(o(2)) ratio within 1 h from 10.7 to 22.9 kPa, but the hypercapnia and acidosis continued. Stepwise reduction of the mean airway pressure (26 cm H(2)O), and oscillating frequencies (3.5 Hz), as well as increasing the oscillating amplitudes (95 cm H(2)O) resulted in an unchanged Pa(co(2)), but oxygenation worsened. The new pumpless extracorporeal interventional lung assist device (ILA, NovaLung, Hechingen, Germany) was therefore used for carbon dioxide elimination to enable a less aggressive ventilation strategy. Pa(co(2)) normalized after initiation of ILA. HFOV with a mean airway pressure of 32 cm H(2)O was maintained, but with a higher oscillatory frequency (9 Hz) and very low oscillatory amplitude (25 cm H(2)O). After 6 days, the patient was transferred to a conventional ventilator, and ILA was discontinued after 13 days without complications. PMID- 15277296 TI - Perioperative systemic haemostatic agents. AB - Skilful surgery combined with blood-saving methods and careful management of blood coagulation will all help reduce unnecessary blood loss and transfusion requirements. Excessive surgical bleeding causes hypovolaemia, haemodynamic instability, anaemia and reduced oxygen delivery to tissues, with a subsequent increase in postoperative morbidity and mortality. The role of anaesthetists in managing surgical blood loss has increased greatly in the last decade. Position of the patient during surgery and the provision of a hypotensive anaesthetic regimen were once considered the most important contributions of the anaesthetist to decreasing blood loss. Now, several pharmacological haemostatic agents are being used by anaesthetists as blood-saving agents. After a brief discussion of the physiology of haemostasis, this article will review the evidence for the role of such agents in reducing perioperative blood loss and transfusion requirements. PMID- 15277298 TI - Estimation of errors in determining intrathoracic blood volume using thermal dilution in pigs with acute lung injury and haemorrhage. AB - BACKGROUND: Global end diastolic volume (GEDV) has a constant and predictable relationship to intrathoracic blood volume (ITBV). The present study assesses the difference between ITBV derived from GEDV and ITBV measured directly in pigs with acute lung injury (ALI) and mild haemorrhage. METHODS: We caused ALI in 12 anaesthetized pigs by i.v. injection of oleic acid and removed 10% of estimated blood volume. EVLW, GEDV, ITBV (COLD; Pulsion Medical Systems), Pa(o(2))/Fi(o(2)), lung compliance and haemodynamic variables were measured at baseline (time 0) and at 30 and 120 min. All animals were volume-resuscitated, followed by measurements at 180 min. A linear equation estimated from the 44 pairs of ITBV and GEDV values in 11 animals was applied iteratively to the four GEDV measurements in the 12th animal, enabling 48 comparisons between measured (ITBVm) and derived ITBV (ITBVd) to be made. RESULTS: Increase in extravascular lung water index (EVLWi) was associated with significant pulmonary hypertension, worsening of oxygenation and compliance (repeated measures ANOVA; P<0.05). There was good within-subject correlation and agreement between ITBV(m) and ITBV(d) (r=0.72, mean bias 0.8 ml; sd 32 ml). Mean error in deriving ITBV from GEDV was 4.5%. (sd 4.2%; range 0.05-19%). There were no significant differences in errors in the presence of small (up to 10%) deficits in blood volume (F=1.0; P=0.41). CONCLUSIONS: ITBV estimated by thermodilution alone is comparable to measurements made by the thermo-dye dilution technique in the presence of pulmonary hypertension and mild deficits in total blood volume. PMID- 15277299 TI - Dynamic aspects of acute tolerance to allopregnanolone evaluated using anaesthesia threshold in male rats. AB - BACKGROUND: It is unclear if allopregnanolone (AlloP) anaesthesia can induce tolerance. Acute tolerance is defined as altered sensitivity to a drug during a single continuous exposure. METHODS: Induction of acute tolerance to AlloP was studied in male rats using a threshold technique of deep anaesthesia. AlloP was infused at a dose rate of 4.0 mg kg(-1) min(-1). The infusion was stopped when a burst suppression of 1 s or more (the "silent second", SS) occurred in the EEG. To maintain anaesthesia, the infusion was restarted when no SS had been seen in the EEG for 1 min. This interrupted targeted infusion towards an EEG end-point (SS) was continued until 30, 60 or 90 min of anaesthesia had been reached. At these times the rats were killed and AlloP concentrations in serum, muscle, fat and different brain regions were determined by radioimmunoassay. RESULTS: Maintenance dose rate (MDR) was calculated using 20-min intervals. During anaesthesia the MDR increased (P<0.001) from 0.67 (sem 0.03) mg kg(-1) min(-1) (in the interval 10-30 min) to 0.98 (0.04) mg kg(-1) min(-1) (in the interval 65 85 min). After 60 min a slight increase in MDR was observed. After 90 min of anaesthesia the AlloP concentrations in the hippocampus and brainstem had increased by more than 50% compared with control values of 25.2 (1.13) and 52.7 (5.81) nmol g(-1) respectively, and after 60 min to around 40%. At 30 min no increase was seen in any brain region analysed. CONCLUSIONS: Measurements in vivo and in vitro record acute tolerance to AlloP occurring with a delay. PMID- 15277300 TI - Effects of thoracic epidural anaesthesia on microvascular gastric mucosal oxygenation in physiological and compromised circulatory conditions in dogs. AB - BACKGROUND: The effects of thoracic epidural anaesthesia (TEA) on gastric mucosal microvascular haemoglobin oxygenation (microHbO(2)) are unclear. At the splanchnic level, reduction of sympathetic tone may promote vasodilation and increase microHbO(2). However, these splanchnic effects are counteracted by systemic effects of TEA (e.g., decreased cardiac output (CO) and mean arterial pressure (MAP)), thus making the net effect on microHbO(2) difficult to predict. In this respect, effects of TEA on microHbO(2) may differ between physiological and compromised circulatory conditions, and additionally may depend on adequate fluid resuscitation. Furthermore, TEA may alter the relationship between regional microHbO(2) and systemic oxygen-transport (DO(2)). METHODS: Chronically instrumented dogs (flow probes for CO measurement) were anaesthetized, their lungs ventilated and randomly received TEA with lidocaine (n=6) or epidural saline (controls, n=6). Animals were studied under physiological and compromised circulatory conditions (PEEP 10 cm H(2)O), both with and without fluid resuscitation. We measured gastric mucosal microHbO(2) by reflectance spectrophotometry, systemic DO(2), and systemic haemodynamics (CO, MAP). RESULTS: Under physiological conditions, TEA preserved microHbO(2) (47 (3)% and 49 (5)%, mean (sem)) despite significantly decreasing DO(2) (11.3 (0.8) to 10.0 (0.7) ml kg(-1) min(-1)) and MAP (66 (2) to 59 (3) mm Hg). However, during compromised circulatory conditions, TEA aggravated the reduction in microHbO(2) (to 32 (1)%), DO(2) (to 6.7 (0.8) ml kg(-1) min(-1)) and MAP (to 52 (4) mm Hg), compared with controls. During TEA, fluid resuscitation completely restored these variables. TEA preserved the correlation between microHbO(2) and DO(2), compared with controls. CONCLUSIONS: TEA maintains microHbO(2) under physiological conditions, but aggravates the reduction of microHbO(2) induced by cardiocirculatory depression, thereby preserving the relationship between gastric mucosal and systemic oxygenation. PMID- 15277301 TI - Management of severe postpartum haemorrhage by uterine artery embolization. AB - We report a case of postpartum haemorrhage which was successfully treated by embolization of the uterine artery. This technique is not well known and is thought to be underused in this condition. We wish to alert medical personnel to its role in this life-threatening situation. PMID- 15277302 TI - Ultrasound guidance in regional anaesthesia. AB - The technology and clinical understanding of anatomical sonography has evolved greatly over the past decade. In the Department of Anaesthesia and Intensive Care Medicine at the Medical University of Vienna, ultrasonography has become a routine technique for regional anaesthetic nerve block. Recent studies have shown that direct visualization of the distribution of local anaesthetics with high frequency probes can improve the quality and avoid the complications of upper/lower extremity nerve blocks and neuroaxial techniques. Ultrasound guidance enables the anaesthetist to secure an accurate needle position and to monitor the distribution of the local anaesthetic in real time. The advantages over conventional guidance techniques, such as nerve stimulation and loss-of resistance procedures, are significant. This review introduces the reader to the theory and practice of ultrasound-guided anaesthetic techniques in adults and children. Considering their enormous potential, these techniques should have a role in the future training of anaesthetists. PMID- 15277303 TI - Effects of propofol and sevoflurane on the excitability of rat spinal motoneurones and nociceptive reflexes in vitro. AB - BACKGROUND: Spinal actions of halogenated ethers are widely recognized, whereas spinal actions of intravenous anaesthetics like propofol are less clear. The aim of this study was to compare the spinal effects of propofol and sevoflurane. METHODS: We used an isolated spinal cord in vitro preparation from rat pups and superfused the anaesthetics at known concentrations. Responses of motoneurones to single and repetitive C-fibre intensity stimulation (trains of 20 stimuli at 1 Hz) of a lumbar dorsal root were recorded from the corresponding ventral root via a suction electrode. RESULTS: Stimulation trains produced a wind-up of action potentials in motoneurones. Both propofol and sevoflurane produced a significant concentration-dependent depression of the evoked wind-up, although at clinically relevant concentrations sevoflurane exhibited a larger intrinsic efficacy. Applied at anaesthetic concentrations, sevoflurane 250 micro M abolished action potentials whereas propofol 1 micro M only produced a reduction close to 50%. At these concentrations, sevoflurane produced a large depressant effect on the monosynaptic reflex whereas propofol was ineffective. CONCLUSIONS: Sevoflurane produces large inhibitory effects on nociceptive and non-nociceptive reflexes which are likely to contribute to immobility during surgery. Compared with sevoflurane, propofol appears to have much weaker effects on spinal reflexes such as those recorded in an isolated preparation. PMID- 15277305 TI - Facial processing deficits and social dysfunction: how are they related? PMID- 15277307 TI - Evaluating the role of the cerebellum in temporal processing: beware of the null hypothesis. PMID- 15277309 TI - PET studies and physiopathology of motor fluctuations in Parkinson's disease. PMID- 15277311 TI - Methyllycaconitine, alpha-bungarotoxin and (+)-tubocurarine block fast ATP-gated currents in rat dorsal root ganglion cells. AB - The effects of nicotinic acetylcholine receptor antagonists were studied on currents evoked by application of ATP to rat isolated dorsal root ganglion cells, and human embryonic kidney 293 cells expressing rat P2X(3) and P2X(2/3) receptors. The rapidly desensitising (within 100 ms) current in dorsal root ganglion cells was inhibited by methyllycaconitine, alpha-bungarotoxin and (+) tubocurarine (concentrations giving half-maximal inhibition were approximately 40, 60 and 800 nm, respectively), but not by hexamethonium (100 microm) or mecamylamine (100 microm). The sustained (>250 ms) current in dorsal root ganglion cells was inhibited by (+)-tubocurarine (80% by 10 microm), but not by methyllycaconitine (200 nm), alpha-bungarotoxin (200 nm), mecamylamine (100 microm) or hexamethonium (100 microm). Rapidly desensitising currents evoked by alpha,betamethylene-ATP in human embryonic kidney cells expressing P2X(3) receptors were inhibited by methyllycaconitine and alpha-bungarotoxin, at concentrations similar to those effective in dorsal root ganglion cells. The results indicate that some nicotinic acetylcholine receptor antagonists are potent blockers of P2X receptors on neurons, particularly the homo-oligomeric P2X(3) receptor. This finding suggests that these drugs should be used with care to discriminate between P2X and neuronal acetylcholine receptor types. PMID- 15277312 TI - Ranolazine: ion-channel-blocking actions and in vivo electrophysiological effects. AB - Ranolazine is a novel anti-ischemic drug that prolongs the QT interval. To evaluate the potential mechanisms and consequences, we studied: (i) Ranolazine's effects on HERG and IsK currents in Xenopus oocytes with two-electrode voltage clamp; (ii) effects of ranolazine, compared to d-sotalol, on effective refractory period (ERP), QT interval and ventricular rhythm in a dog model of acquired long QT syndrome; and (iii) effects on selected native currents in canine atrial myocytes with whole-cell patch-clamp technique. Ranolazine inhibited HERG and IsK currents with different potencies. HERG was inhibited with an IC(50) of 106 micromol l(-1), whereas the IC(50) for IsK was 1.7 mmol l(-1). d-Sotalol caused reverse use-dependent ERP and QT interval prolongation, whereas ranolazine produced modest, nonsignificant increases that plateaued at submaximal doses. Neither drug affected QRS duration. d-Sotalol had clear proarrhythmic effects, with all d-sotalol-treated dogs developing torsades de pointes (TdP) ventricular tachyarrhythmias, of which they ultimately died. In contrast, ranolazine did not generate TdP. Effects on I(Kr) and I(Ks) were similar to those on HERG and IsK. Ranolazine blocked I(Ca) with an IC(50) of approximately 300 micromol l(-1). I(Na) was unaffected. We conclude that ranolazine inhibits I(Kr) by blocking HERG currents, inhibits I(Ca) at slightly larger concentrations, and has modest and self-limited effects on the QT interval. Unlike d-sotalol, ranolazine does not cause TdP in a dog model. The greater safety of ranolazine may be due to its ability to inhibit I(Ca) at concentrations only slightly larger than those that inhibit I(Kr), thus producing offsetting effects on repolarization. PMID- 15277313 TI - Cannabinoids and intestinal motility: welcome to CB2 receptors. AB - Delta(9)-tetrahydrocannabinol (the active ingredient of marijuana), as well as endogenous and synthetic cannabinoids, exert many biological functions by activating two types of cannabinoid receptors, CB(1) receptors (expressed by central and peripheral neurons) and CB(2) receptors (that occur mainly in immune cells). Convincing evidence has accumulated in recent years that cannabinoids inhibit gastric and intestinal motility through activation of enteric CB(1) receptors. However, a report in this issue of British Journal of Pharmacology has highlighted the possibility that CB(2) receptors in the rat intestine could contribute to reducing the increase of intestinal motility induced by an endotoxic inflammation. By minimizing the adverse psychotropic effects associated with brain cannabinoid receptors, the CB(2) receptor represents a new molecular target for the treatment of motility disorders associated with intestinal inflammation. PMID- 15277314 TI - Intestinal Na+-K+-ATPase activity and molecular events downstream of interferon gamma receptor stimulation. AB - This study evaluated the effects of human interferon-gamma (IFN-gamma) on Na(+) K(+)-ATPase activity and the intracellular signaling pathways involved in human intestinal epithelial Caco-2 cells. Na(+)-K(+)-ATPase activity was determined as the difference between total and ouabain-sensitive ATPase. p38 MAP kinase activity was analyzed by Western blotting using the p38 MAP kinase assay kit. Total and phosphorylated STAT1 protein levels were detected using the PhosphoPlus Stat1. IFN-gamma decreased Na(+)-K(+)-ATPase activity in a time- and concentration-dependent manner. The IFN-gamma-induced decrease in Na(+)-K(+) ATPase activity was accompanied by no changes in the abundance of alpha(1) subunit Na(+)-K(+)-ATPase. Downregulation of protein kinase C (PKC) with phorbol 12,13-dibutyrate (PDBu) prevented the inhibitory effect of IFN-gamma on Na(+) K(+)-ATPase activity. Inhibition of Raf-1, mitogen-activated protein kinase kinase (MAPKK/MEK), p38 MAPK and STAT1 with, respectively, GW 5074, PD 98059, SB 203580 and epigallocatechin gallate prevented inhibition of Na(+)-K(+)-ATPase activity by IFN-gamma. Treatment with IFN-gamma markedly increased the expression of total and phospho-STAT1, this being accompanied by activation of p38 MAPK. Activation of phospho-STAT1 by IFN-gamma was almost abolished by epigallocatechin gallate and markedly reduced by SB 203580, but insensitive to downregulation of PKC. The increase in short circuit current (I(sc)) by 1.0 and 2.5 micrograms ml( 1) amphotericin B was markedly attenuated in IFN-gamma-treated cells. However, the inhibitory effect of PDBu on the amphotericin B-induced increase in I(sc) was of similar magnitude in vehicle- and IFN-gamma-treated cells. It is concluded that IFN-gamma markedly attenuates Na(+)-K(+)-ATPase activity. The transduction mechanisms set into motion by IFN-gamma involve the activation of PKC downstream STAT1 phosphorylation and Raf-1, MEK, ERK2 and p38 MAPK pathways, in a complex sequence of events. PMID- 15277315 TI - Anandamide acts as a vasodilator of dural blood vessels in vivo by activating TRPV1 receptors. AB - Migraine pathophysiology is believed to involve the release of neuropeptides via the activation of trigeminal afferents that innervate the cranial vasculature. Anandamide, the endogenous ligand to the cannabinoid receptor, is able to inhibit neurogenic dural vasodilatation, calcitonin gene-related peptide (CGRP)-induced and nitric oxide-induced dural vessel dilation in the intravital microscopy model. In an in vitro setting anandamide is also able to activate the vanilloid type 1 (TRPV1) receptor and cause vasodilation, via the release of CGRP. In this study we used intravital microscopy to study whether anandamide behaves as a TRPV1 receptor agonist in the trigeminovascular system. We examined if anandamide induced dural vasodilation involves CGRP release that can be reversed by the CGRP receptor antagonist, CGRP(8-37), and whether like capsaicin the anandamide effect could be reversed by the TRPV1 receptor antagonist, capsazepine. Anandamide 1 (19+/-9%, n=12), 3 (29+/-5%, n=37), 5 (74+/-7%, n=13) and 10 mg kg(-1) (89+/-18%, n=6) was able to cause a dose-dependent increase in dural vessel diameter. Capsazepine (3 mg kg(-1), t(5)=6.2, P<0.05) and CGRP(8-37) (300 micrograms kg( 1), t(6)=11.1, P<0.05) attenuated the anandamide-induced dural vessel dilation when compared to control (Student's paired t-test). AM251 (3 mg kg(-1)), a cannabinoid type 1 (CB(1)) receptor antagonist, was unable to reverse this anandamide-induced dilation. The study demonstrates that anandamide acts as a TRPV1 receptor agonist in the trigeminovascular system, activating TRPV1 receptors that promote CGRP release and cause vasodilation independent of any action at the CB(1) receptor. Anandamide has been shown previously to inhibit trigeminovascular neurons and prevent vasodilation, through an action at CB(1) receptors. PMID- 15277316 TI - The cannabinomimetic arachidonyl-2-chloroethylamide (ACEA) acts on capsaicin sensitive TRPV1 receptors but not cannabinoid receptors in rat joints. AB - The vasoactive effects of the synthetic cannabinoid (CB) arachidonyl-2 chloroethylamide (ACEA) was tested in the knee joints of urethane-anaesthetised rats. Experiments were also performed to determine whether these vasomotor responses could be blocked by the selective CB(1) receptor antagonists AM251 (N (piperidin-1-yl)-5-(4-iodophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3 carboxamide) (10(-9) mol) and AM281 (1-(2,4-dichlorophenyl)-5-(4-iodophenyl)-4 methyl-N-4-morpholinyl-1H-pyrazole-3-carboxamide) (10(-8) mol), as well as the selective CB(2) receptor antagonist AM630 (6-iodo-2-methyl-1-[2 4(morpholinyl)ethyl]-[1H-indol-3-yl](4-methoxyphenyl)methanone) (10(-8) mol). Peripheral application of ACEA (10(-14)-10(-9) mol) onto the exposed surface of the knee joint capsule caused a dose-dependent increase in synovial blood flow. The dilator action of the CB occurred within 1 min after drug administration and rapidly returned to control levels shortly thereafter. The maximal vasodilator effect of ACEA corresponded to a 30% increase in articular perfusion compared to control levels. The hyperaemic action of ACEA was not significantly altered by coadministration of AM251, AM281 or AM630 (P>0.05; two-way ANOVA). The transient receptor potential channel vanilloid receptor 1 (TRPV(1)) antagonist capsazepine (10(-6) mol) significantly reduced the vasodilator effect of ACEA on joint blood vessels (P=0.002). Furthermore, destruction of unmyelinated and thinly myelinated joint sensory nerves by capsaicin (8-methyl-N-vanillyl-6-nonenamide) treatment also attenuated ACEA responses (P<0.0005). These data clearly demonstrate a vasodilator effect of the cannabinomimetic ACEA on knee joint perfusion. Rather than a classic CB receptor pathway, ACEA exerts its vasomotor influence by acting via TRPV(1) receptors located on the terminal branches of capsaicin-sensitive afferent nerves innervating the joint. PMID- 15277317 TI - Effect of aldosterone antagonism on myocardial dysfunction in hypertensive patients with diastolic heart failure. AB - BACKGROUND: Specific treatments targeting the pathophysiology of hypertensive heart disease are lacking. As aldosterone has been implicated in the genesis of myocardial fibrosis, hypertrophy, and dysfunction, we sought to determine the effects of aldosterone antagonism on myocardial function in hypertensive patients with suspected diastolic heart failure by using sensitive quantitative echocardiographic techniques in a randomized, double-blinded, placebo-controlled study. METHODS AND RESULTS: Thirty medically treated ambulatory hypertensive patients (19 women, age 62+/-6 years) with exertional dyspnea, ejection fraction >50%, and diastolic dysfunction (E/A <1, E deceleration time >250 m/sec) and without ischemia were randomized to spironolactone 25 mg/d or placebo for 6 months. Patients were overweight (31+/-5 kg/m2) with reduced treadmill exercise capacity (6.7+/-2.1 METS). Long-axis strain rate (SR), peak systolic strain, and cyclic variation of integrated backscatter (CVIB) were averaged from 6 walls in 3 standard apical views. Mean 24-hour ambulatory blood pressure at baseline (133+/ 17/80+/-7 mm Hg) did not change in either group. Values for SR, peak systolic strain, and CVIB were similar between groups at baseline and remained unchanged with placebo. Spironolactone therapy was associated with increases in SR (baseline: -1.57+/-0.46 s(-1) versus 6-months: -1.91+/-0.36 s(-1), P<0.01), peak systolic strain (-20.3+/-5.0% versus -26.9+/-4.3%, P<0.001), and CVIB (7.4+/-1.7 dB versus 8.6+/-1.7 dB, P=0.08). Each parameter was significantly greater in the spironolactone group compared with placebo at 6 months (P=0.05, P=0.02, and P=0.02, respectively), and the increases remained significant after adjusting for baseline differences. The increase in strain was independent of changes in blood pressure with intervention. The spironolactone group also exhibited reduction in posterior wall thickness (P=0.04) and a trend to reduced left atrial area (P=0.09). CONCLUSIONS: Aldosterone antagonism improves myocardial function in hypertensive heart disease. PMID- 15277320 TI - Carotid plaque echolucency increases the risk of stroke in carotid stenting: the Imaging in Carotid Angioplasty and Risk of Stroke (ICAROS) study. AB - BACKGROUND: Carotid artery stenting (CAS) has recently emerged as a potential alternative to carotid endarterectomy. Cerebral embolization is the most devastating complication of CAS, and the echogenicity of carotid plaque has been indicated as one of the risk factors involved. This is the first study to analyze the role of a computer-assisted highly reproducible index of echogenicity, namely the gray-scale median (GSM), on the risk of stroke during CAS. METHODS AND RESULTS: The Imaging in Carotid Angioplasty and Risk of Stroke (ICAROS) registry included 418 cases of CAS collected from 11 international centers. An echographic evaluation of carotid plaque with GSM measurement was made preprocedurally. The onset of neurological deficits during the procedure and the postprocedural period was recorded. The overall rate of neurological complications was 3.6%: minor strokes, 2.2%, and major stroke, 1.4%. There were 11 of 155 strokes (7.1%) in patients with GSM < or =25 and 4 of 263 (1.5%) in patients with GSM >25 (P=0.005). Patients with severe stenosis (> or =85%) had a higher rate of stroke (P=0.03). The effectiveness of brain protection devices was confirmed in those with GSM >25 (P=0.01) but not in those with GSM < or =25. Multivariate analysis revealed that GSM (OR, 7.11; P=0.002) and rate of stenosis (OR, 5.76; P=0.010) are independent predictors of stroke. CONCLUSIONS: Carotid plaque echolucency, as measured by GSM < or =25, increases the risk of stroke in CAS. The inclusion of echolucency measured as GSM in the planning of any endovascular procedure of carotid lesions allows stratification of patients at different risks of complications in CAS. PMID- 15277321 TI - Occupational, commuting, and leisure-time physical activity in relation to total and cardiovascular mortality among Finnish subjects with type 2 diabetes. AB - BACKGROUND: Some previous studies have assessed the association between leisure time physical activity and mortality among patients with diabetes, but the potential effect of occupational and commuting physical activity remains uncertain. METHODS AND RESULTS: We prospectively followed up 3316 Finnish participants 25 to 74 years of age with type 2 diabetes. The association of different types of physical activity with mortality was examined with Cox proportional-hazard models. During a mean follow-up of 18.4 years, 1410 deaths were recorded, 903 of which were due to cardiovascular disease (CVD). The multivariate-adjusted (age, sex, study year, body mass index, systolic blood pressure, cholesterol, smoking, and the 2 other types of physical activity) hazard ratios associated with light, moderate, and active work were 1.00, 0.86, and 0.60 (P(trend)<0.001) for total mortality and 1.00, 0.91, and 0.60 (P(trend)<0.001) for CVD mortality, respectively. The multivariate-adjusted hazard ratios associated with low, moderate, and high leisure-time physical activity were 1.00, 0.82, and 0.71 (P(trend)<0.001) for total mortality and 1.00, 0.83, and 0.67 (P(trend)=0.005) for CVD mortality, respectively. Active commuting had significant inverse associations with total and CVD mortality, but these relations were no longer significant after additional adjustment for occupational and leisure-time physical activity. CONCLUSIONS: Moderate or high levels of physical activity reduce total and CVD mortality among patients with type 2 diabetes. Not only leisure-time physical activity but also occupational and commuting physical activities are important components of a healthy lifestyle among patients with diabetes. PMID- 15277322 TI - Randomized trial of atorvastatin for reduction of myocardial damage during coronary intervention: results from the ARMYDA (Atorvastatin for Reduction of MYocardial Damage during Angioplasty) study. AB - BACKGROUND: Small myocardial infarctions after percutaneous coronary intervention have been associated with higher risk of cardiac events during follow-up. Observational studies have suggested that statins may lower the risk of procedural myocardial injury. The aim of our study was to confirm this hypothesis in a randomized study. METHODS AND RESULTS: One hundred fifty-three patients with chronic stable angina without previous statin treatment were enrolled in the study. Patients scheduled for elective coronary intervention were randomized to atorvastatin (40 mg/d, n=76) or placebo (n=77) 7 days before the procedure. Creatine kinase-MB, troponin I, and myoglobin levels were measured at baseline and at 8 and 24 hours after the procedure. Detection of markers of myocardial injury above the upper normal limit was significantly lower in the statin group versus the placebo group: 12% versus 35% for creatine kinase-MB (P=0.001), 20% versus 48% for troponin I (P=0.0004), and 22% versus 51% for myoglobin (P=0.0005). Myocardial infarction by creatine kinase-MB determination was detected after coronary intervention in 5% of patients in the statin group and in 18% of those in the placebo group (P=0.025). Postprocedural peak levels of creatine kinase-MB (2.9+/-3 versus 7.5+/-18 ng/mL, P=0.007), troponin I (0.09+/ 0.2 versus 0.47+/-1.3 ng/mL, P=0.0008), and myoglobin (58+/-36 versus 81+/-49 ng/mL, P=0.0002) were also significantly lower in the statin than in the placebo group. CONCLUSIONS: Pretreatment with atorvastatin 40 mg/d for 7 days significantly reduces procedural myocardial injury in elective coronary intervention. These results may influence practice patterns with regard to adjuvant pharmacological therapy before percutaneous revascularization. PMID- 15277324 TI - Modern pacemaker and implantable cardioverter/defibrillator systems can be magnetic resonance imaging safe: in vitro and in vivo assessment of safety and function at 1.5 T. AB - BACKGROUND: MRI has unparalleled soft-tissue imaging capabilities. The presence of devices such as pacemakers and implantable cardioverter/defibrillators (ICDs), however, is historically considered a contraindication to MRI. These devices are now smaller, with less magnetic material and improved electromagnetic interference protection. Our aim was to determine whether these modern systems can be used in an MR environment. METHODS AND RESULTS: We tested in vitro and in vivo lead heating, device function, force acting on the device, and image distortion at 1.5 T. Clinical MR protocols and in vivo measurements yielded temperature changes <0.5 degrees C. Older (manufactured before 2000) ICDs were damaged by the MR scans. Newer ICD systems and most pacemakers, however, were not. The maximal force acting on newer devices was <100 g. Modern (manufactured after 2000) ICD systems were implanted in dogs (n=18), and after 4 weeks, 3- to 4 hour MR scans were performed (n=15). No device dysfunction occurred. The images were of high quality with distortion dependent on the scan sequence and plane. Pacing threshold and intracardiac electrogram amplitude were unchanged over the 8 weeks, except in 1 animal that, after MRI, had a transient (<12 hours) capture failure. Pathological data of the scanned animals revealed very limited necrosis or fibrosis at the tip of the lead area, which was not different from controls (n=3) not subjected to MRI. CONCLUSIONS: These data suggest that certain modern pacemaker and ICD systems may indeed be MRI safe. This may have major clinical implications for current imaging practices. PMID- 15277325 TI - Morphology of atrial myocardial extensions into human caval veins: a postmortem study in patients with and without atrial fibrillation. AB - BACKGROUND: Atrial fibrillation (AF) may be triggered from arrhythmogenic foci originating from atrial muscular sleeves that extend into the caval veins (CVs). The aim of this anatomic study was to evaluate both the extent and arrangement of atrial myocardial fibers in CVs in subjects with and without a history of AF. METHODS AND RESULTS: Twenty-five human autopsied hearts (15 men; mean age, 65.5+/ 12 years; range, 39 to 80 years) were studied. Seven subjects had a previous history of AF. The presence and morphology of atrial myocardial extensions were studied microscopically in both CVs. Such extensions were found in 38 of 50 CVs (76%). Their average length in the superior vena cava reached 13.7+/-13.9 mm (maximum, up to 47 mm) and in the inferior vena cava, 14.6+/-16.7 mm (maximum, up to 61 mm). The thickness of atrial myocardium extending into the CVs was 1.2+/ 1.0 mm (maximum, 4 mm) for the superior vena cava and 1.2+/-0.9 mm for the inferior vena cava (maximum, 3 mm). The majority of myocardial extensions revealed discontinuous and circular patterns. Degenerative changes were found in approximately half of the subjects. There was no significant difference between patients with and without a history of AF. CONCLUSIONS: Atrial myocardial extensions into both CVs are present in the majority of human beings, both with and without a history of AF. The extensions are localized on the outer side of venous adventitia. Arrangement, length, and thickness of myocardial sleeves onto the CVs vary individually, and many of them contain degenerative changes. PMID- 15277326 TI - C-reactive protein induces apoptosis in human coronary vascular smooth muscle cells. AB - BACKGROUND: Accumulating evidence suggests that C-reactive protein (CRP), in addition to being a predictor of coronary events, may have direct actions on the vessel wall in the evolution of atherosclerosis. Although accumulation of vascular smooth muscle cells (VSMCs) in the intima is a key event in the development of arterial lesions, apoptosis of VSMCs also plays an important role in progression of atherosclerotic lesions and contributes to increased plaque vulnerability. METHODS AND RESULTS: In the present study we demonstrate that CRP induces caspase-mediated apoptosis of human coronary VSMCs. DNA microarray analysis was used to identify CRP-regulated genes. The growth arrest- and DNA damage-inducible gene 153 (GADD153) mRNA expression was prominently upregulated by CRP. As confirmed by Northern blot analysis, CRP induced a time- and dose dependent increase of GADD153 mRNA expression. GADD153, a gene involved in growth arrest and apoptosis in vascular and nonvascular cells, is regulated at both transcriptional and posttranscriptional levels. CRP regulation of GADD153 mRNA expression in VSMCs occurs primarily at the posttranscriptional level by mRNA stabilization. Small interfering RNA (siRNA) specifically targeted to GADD153 reduced CRP-induced apoptosis. GADD153 also specifically colocalized to apoptotic VSMCs in human coronary lesions, further supporting a functional role for GADD153 in CRP-induced cell death. CONCLUSIONS: These results demonstrate that GADD153 is a CRP-regulated gene in human VSMCs and plays a causal role in CRP-induced apoptosis. Pharmacological targeting of CRP expression or action may provide a novel therapy for atherosclerosis. PMID- 15277327 TI - Increase in serum amyloid a evoked by dietary cholesterol is associated with increased atherosclerosis in mice. AB - BACKGROUND: Elevated serum amyloid A (SAA) levels are associated with increased cardiovascular risk. SAA levels can be increased by dietary fat and cholesterol. Moreover, SAA can cause lipoproteins to bind extracellular vascular proteoglycans, a process that is critical in atherogenesis. Therefore, we hypothesized that diet-induced increases in SAA would increase atherosclerosis independent of their effect on plasma cholesterol levels. METHODS AND RESULTS: Female LDL-receptor-null (LDLR-/-) mice were fed high-saturated fat diets (21%, wt/wt), with or without added cholesterol (0.15%, wt/wt), for 10 weeks. Compared with chow-fed controls, the high-fat diets increased plasma SAA levels. Addition of cholesterol further increased SAA levels 2-fold (P<0.05) without further increasing plasma cholesterol levels. Addition of dietary cholesterol also increased atherosclerosis (P<0.05). Four lines of evidence suggest that SAA actually might cause atherosclerosis: (1) SAA levels when mice were euthanized correlated with the extent of atherosclerosis (r=0.49; P<0.02); (2) SAA levels after 5 weeks of diet correlated with the extent of atherosclerosis at 10 weeks (r=0.66; P<0.01); (3) binding of HDL from these animals to proteoglycans in vitro was related to the HDL-SAA content (r=0.65; P<0.01); and (4) immunoreactive SAA was present in lesion areas enriched with both proteoglycans and apolipoprotein A I, the major HDL apolipoprotein. CONCLUSIONS: Addition of cholesterol to a high fat diet increased plasma SAA levels and atherosclerosis independent of an adverse effect on plasma lipoproteins, consistent with the hypothesis that SAA may promote atherosclerosis directly by mediating retention of SAA-enriched HDL to vascular proteoglycans. PMID- 15277328 TI - Distal protection with a filter device during coronary stenting in patients with stable and unstable angina. AB - BACKGROUND: Filter protection after percutaneous coronary intervention (PCI) is now available to prevent distal embolization. The aims of this study were (1) to evaluate the microembolization phenomenon during procedures of stent implantation in native coronary arteries of patients with stable and unstable angina, (2) to assess the amount and characteristics of the debris captured by the Angioguard, and (3) to investigate the relation between clinical and angiographic variables and pathological data. METHODS AND RESULTS: Elective coronary stenting with the use of a protective filter was attempted in 39 consecutive coronary artery lesions with >60% stenosis (mean, 67.6+/-8.79%). Debris was present in 75.6% of the filters. Particle size ranged from 47.16 to 2503.48 microm (mean, 518.83+/ 319.61 microm) in the major axis. Particles >300 microm were found in 24 of 28 filters with debris (85.7%), and particles >1000 microm were present in 10 of 28 filters (35.7%). Patients with unstable angina had greater particles (mean maximum longitudinal diameter, 1098.33+/-714.3 microm) than those with stable angina (412.91+/-453 microm; P<0.001). The presence of unstable angina (OR, 65; CI, 1.2 to 3420; P=0.03) and age >67 years (OR, 42; CI, 1 to 1698; P=0.04) were found to be the only independent predictors of embolic particle size. CONCLUSIONS: By limiting embolization, protective devices may prevent a number of potentially unfavorable events, thereby improving outcome. Our data support the use of these devices, especially in lesions with higher embolic potential, such as those occurring in older patients and in those with unstable angina. PMID- 15277329 TI - Inhibition of diet-induced atherosclerosis and endothelial dysfunction in apolipoprotein E/angiotensin II type 1A receptor double-knockout mice. AB - BACKGROUND: Angiotensin II type 1 (AT1) receptor activation is potentially involved in the multifactorial pathogenesis of atherosclerosis. METHODS AND RESULTS: Apolipoprotein E-deficient (ApoE-/-) mice were crossed with AT1A receptor-deficient (AT1-/-) mice to obtain homozygous double-knockout animals (ApoE-/--AT1-/- mice). Wild-type (C57BL/6J), ApoE-/-, AT1-/-, and ApoE-/--AT1-/- mice were fed a high-cholesterol diet for 7 weeks. In contrast to wild-type and AT1-/- mice, this treatment led to severe atherosclerotic lesion formation in the aortic sinus and the aorta (oil red O staining) and to an impaired endothelium dependent vasodilation (organ chamber experiments with isolated aortic segments) in ApoE-/- mice. In the age-matched ApoE-/--AT1-/- littermates, development of diet-induced endothelial dysfunction and atherosclerotic lesion formation was profoundly inhibited. Concomitantly, aortic release of superoxide radicals was increased 2-fold in ApoE-/- mice compared with wild-type animals, whereas aortic superoxide production was normalized in ApoE-/--AT1-/- mice (L-012 chemiluminescence). There were no significant differences in plasma cholesterol levels between ApoE-/- and ApoE-/--AT1-/- animals. Systolic blood pressure was significantly lower in ApoE-/--AT1-/- animals than in ApoE-/- mice (tail-cuff measurements). Oral treatment of ApoE-/- mice with either hydralazine or irbesartan reduced systolic blood pressure to the same level; however, only AT1 receptor antagonist treatment reduced atherosclerotic lesion formation and improved endothelial function. CONCLUSIONS: Genetic disruption of the AT1A receptor leads to inhibition of vascular oxidative stress, endothelial dysfunction, and atherosclerotic lesion formation in ApoE-/- mice irrespective of blood pressure and plasma cholesterol levels. These results indicate a fundamental role of AT1 receptor activation in atherogenesis. PMID- 15277330 TI - Role for matrix metalloproteinase-2 in oxidized low-density lipoprotein-induced activation of the sphingomyelin/ceramide pathway and smooth muscle cell proliferation. AB - BACKGROUND: Oxidized LDLs (oxLDLs) and matrix metalloproteinases (MMPs) are present in atherosclerotic lesions. OxLDLs activate various signaling pathways potentially involved in atherogenesis. OxLDLs induce smooth muscle cell (SMC) proliferation mediated by the activation of the sphingomyelin/ceramide pathway and tyrosine kinase receptors. MMPs are also able to induce SMC migration and proliferation in addition to extracellular matrix degradation. The present study was designed to investigate whether MMPs play a role in the mitogenic effect of oxLDLs. METHODS AND RESULTS: OxLDLs induce the release of activated MMP-2 in SMC culture medium. MMP-2 was identified by its 65-kDa gelatinase activity on zymography and by using specific blocking antibodies and MMP-2-/- cells. MMP inhibitors (batimastat and Ro28-2653) and the blocking antibodies anti-MMP-2 and anti-membrane type 1-MMP inhibited the oxLDL-induced sphingomyelin/ceramide pathway activation and subsequent activation of ERK1/2 and DNA synthesis but did not inhibit the oxLDL-induced epidermal growth factor receptor and platelet derived growth factor receptor activation. Exogenously added activated MMP-2 or membrane type 1-MMP-1 triggered the activation of both sphingomyelin/ceramide and ERK1/2 pathways and DNA synthesis. Conversely, suppression of MMP-2 expression in MMP-2-/- cells or in SMCs treated by small-interference RNA also blocked both sphingomyelin/ceramide signaling and DNA synthesis. CONCLUSIONS: Together, these data demonstrate that MMP-2 plays a pivotal role in oxLDL-induced activation of the sphingomyelin/ceramide signaling pathway and subsequent SMC proliferation. These pathways may constitute a potential therapeutic target for modulating the oxLDL-induced proliferation of SMCs in atherosclerosis or restenosis. PMID- 15277331 TI - Different effects of antihypertensive therapies based on losartan or atenolol on ultrasound and biochemical markers of myocardial fibrosis: results of a randomized trial. AB - BACKGROUND: In hypertensive left ventricular hypertrophy (LVH), myocardial texture is altered by a disproportionate increase in fibrosis, but there is insufficient clinical evidence whether antihypertensive therapy or individual agents can induce regression of myocardial fibrosis. METHODS AND RESULTS: We compared the effects of an angiotensin II receptor antagonist with a beta-blocker on myocardial collagen volume (assessed by echoreflectivity and serum collagen markers) in 219 hypertensive patients with echocardiographically documented LVH. Patients were allocated randomly to receive losartan 50 to 100 mg/d (n=111) or atenolol 50 to 100 mg/d (n=99) with or without hydrochlorothiazide 12.5 to 25 mg/d for 36 weeks. Echoreflectivity analysis was conducted on ultrasound tracings of the midapex septum with specifically designed and validated software. A color histogram of reflecting echoes was obtained, and its spread (broadband [BB], previously shown to correlate directly with collagen volume fraction on endomyocardial biopsies) was used as the primary outcome measure. Mean color scale and serum markers of collagen synthesis (PIP, PIIIP) or degradation (CITP) were secondary outcome variables. Echoreflectivity analysis proved feasible in 106 patients (losartan 52, atenolol 54). Losartan reduced BB over 36 weeks (from 114.5 to 104.3 color levels, P<0.02), whereas atenolol treatment was associated with an increase in BB (from 109.0 to 113.6 color levels, P=NS), the difference between treatments being -12.8 color levels (95% CI -23.6 to -2.0, P=0.02). Secondary end points (mean color scale and collagen markers) also changed in the direction of decreased collagen in patients receiving losartan, but differences between groups were not statistically significant. CONCLUSIONS: In hypertensive patients with LVH, losartan decreases myocardial collagen content, whereas atenolol does not. The difference between the 2 treatments is statistically significant. PMID- 15277332 TI - Delayed ischemic preconditioning activates nuclear-encoded electron-transfer chain gene expression in parallel with enhanced postanoxic mitochondrial respiratory recovery. AB - BACKGROUND: Delayed ischemic preconditioning promotes cardioprotection via genomic reprogramming. We hypothesize that molecular regulation of mitochondrial energetics is integral to this cardioprotective program. METHODS AND RESULTS: Preconditioning was induced by use of 3 episodes of 3-minute coronary artery occlusion separated by 5 minutes of reperfusion. Twenty-four hours later, infarct size was reduced by 58% after preconditioning compared with sham-operated controls (P<0.001). Cardiac mitochondria were isolated from sham and preconditioned rat hearts. Mitochondrial respiration and ATP production were similar between the groups; however, preconditioned mitochondria exhibit modest hyperpolarization of the inner mitochondrial membrane potential (> or =22% versus control, P<0.001). After 35-minute anoxia and reoxygenation, preconditioned mitochondria demonstrated a 191+/-12% improvement in ADP-sensitive respiration (P=0.002) with preservation of electron-transfer-chain (ETC) activity versus controls. This augmented mitochondrial recovery was eradicated when preconditioning was abolished by the antioxidant 2-mercaptopropionyl glycine (2 MPG). These biochemical modulations appear to be regulated at the genomic level in that the expression of genes encoding rate-controlling complexes in the ETC was significantly upregulated in preconditioned myocardium, with a concordant induction of steady-state protein levels of cytochrome oxidase, cytochrome c, and adenine nucleotide translocase-1. 2-MPG abolished preconditioning induction of these transcripts. Moreover, transcripts of nuclear regulatory peptides known to orchestrate mitochondrial biogenesis, nuclear respiratory factor-1 and peroxisome proliferator-activated receptor gamma coactivator 1alpha, were significantly induced in preconditioned myocardium. CONCLUSIONS: Delayed preconditioned mitochondria display increased tolerance against anoxia-reoxygenation in association with modifications in mitochondrial bioenergetics, with concordant genomic induction of a mitochondrial energetic gene regulatory program. This program appears to be mediated by reactive oxygen species signaling. PMID- 15277333 TI - Cardiopulmonary exercise testing: how do we differentiate the cause of dyspnea? PMID- 15277334 TI - Images in cardiovascular medicine. Sleep (vagal)-induced atrial fibrillation. PMID- 15277335 TI - Vagal nerve stimulation in chronic heart failure: an antiinflammatory intervention? PMID- 15277336 TI - Metabolic syndrome: a definition in progress. PMID- 15277337 TI - Myocardial injury and cardiac troponin I release after off-pump versus on-pump coronary surgery. PMID- 15277338 TI - Homocysteine is an independent risk factor for myocardial infarction, in particular fatal myocardial infarction in middle-aged women. PMID- 15277339 TI - Serum homocysteine and endothelial dysfunction in circulatory disorders in women. PMID- 15277340 TI - Cardiovascular protections in severely impaired hemostasis. PMID- 15277341 TI - RNA reference materials for gene expression studies. Difficult first steps. PMID- 15277342 TI - RNA reference materials for gene expression studies. RNA metrology: forecast calls for partial clearing. PMID- 15277343 TI - Homocysteine and folate in pregnancy. PMID- 15277344 TI - SNPs for sale. Cheap! PMID- 15277345 TI - Biopsy or biomarkers: is there a gold standard for diagnosis of liver fibrosis? PMID- 15277347 TI - Detection of placental transcription factor mRNA in maternal plasma. PMID- 15277346 TI - Development of an ELISA for measurement of BCAR1 protein in human breast cancer tissue. AB - BACKGROUND: High concentrations of breast cancer anti-estrogen resistance 1 (BCAR1) protein measured by Western blotting in primary breast tumor cytosols are associated with early disease progression and failure of tamoxifen therapy. The aim of the present study was to develop an ELISA to measure BCAR1 quantitatively in extracts of human breast cancer tissue. METHODS: A recombinant fragment of BCAR1 (the human homolog of murine p130Cas) was produced in bacterial M15 cells, purified, and injected into chickens and rabbits. The generated antibodies were affinity-purified and used for the construction of an ELISA. After validation, the results obtained with the ELISA were compared with Western blot findings on primary breast tumors. RESULTS: The detection limit the BCAR1 ELISA was 0.0031 microg/L, and the within-run imprecision (CV) was <20% at concentrations down to 0.004 microg/L. The within-run imprecision (CV) was 1.0-7.2%, and the between-run CV was 3.6-5.4%. There was no cross-reactivity with family member HEF1. The assay exhibited parallelism of results between serial dilutions and a mean recovery (range) of 96 (79-118)%. CONCLUSIONS: The ELISA measures BCAR1 in human breast cancer cytosols with high sensitivity and specificity. The assay can be used to confirm and to quantitatively extend previous semiquantitative Western blot data on the prognostic and predictive value of BCAR1 in human breast cancer; it can also be applied for other diseases. PMID- 15277348 TI - Candidate reference method for the quantification of circulating 25 hydroxyvitamin D3 by liquid chromatography-tandem mass spectrometry. PMID- 15277349 TI - Convenient single-nucleotide polymorphism typing from whole blood by probe extension and bioluminescence detection. PMID- 15277350 TI - Effect of blood contamination on delta 450 bilirubin measurement: an in vitro comparison of two corrective methods. PMID- 15277351 TI - Interassay and interobserver variability in the detection of anti-neutrophil cytoplasmic antibodies in patients with ulcerative colitis. PMID- 15277352 TI - Survivin mRNA copy number in bladder washings predicts tumor recurrence in patients with superficial urothelial cell carcinomas. PMID- 15277353 TI - Biological variation of plasma F2-isoprostane-III and arachidonic acid in healthy individuals. PMID- 15277354 TI - Specimen dilution for C2 monitoring with the Abbott TDxFLx cyclosporine monoclonal whole blood assay. PMID- 15277355 TI - Direct detection of exon deletions/duplications in female carriers of and male patients with Duchenne/Becker muscular dystrophy. PMID- 15277356 TI - Protein profiling in urine for the diagnosis of bladder cancer. PMID- 15277357 TI - Electronic microarray technique for detection of nine base substitutions including single-nucleotide polymorphisms in the human OGG1 gene. PMID- 15277358 TI - Automated turbidimetric benzalkonium chloride method for measurement of protein in urine and cerebrospinal fluid. PMID- 15277359 TI - Circulating soluble CD14 monocyte receptor is associated with increased alanine aminotransferase. PMID- 15277360 TI - Determination of guanidinoacetate and creatine in urine and plasma by liquid chromatography-tandem mass spectrometry. PMID- 15277361 TI - Ki-67 protein concentrations in urothelial bladder carcinomas are related to Ki 67-specific RNA concentrations in urine. PMID- 15277362 TI - PCR-based haplotype determination to distinguish CYP2B6*1/*7 and *5/*6. PMID- 15277363 TI - Interference by gelatin-based plasma substitutes in capillary zone electrophoresis. PMID- 15277364 TI - Clarification in the point/counterpoint discussion related to surface-enhanced laser desorption/ionization time-of-flight mass spectrometric identification of patients with adenocarcinomas of the prostate. PMID- 15277365 TI - Serum 99th percentile reference cutoffs for seven cardiac troponin assays. PMID- 15277366 TI - Rapid detection of UGT1A1 gene polymorphisms by newly developed Invader assay. PMID- 15277367 TI - Methylation of mitochondrial DNA is not a useful marker for cancer detection. PMID- 15277368 TI - Increased selenium concentrations in seronorm trace elements serum (level 2). PMID- 15277369 TI - Urinary methylmalonic acid test may have greater value than the total homocysteine assay for screening elderly individuals for cobalamin deficiency. PMID- 15277370 TI - Does the glucose-dependent insulin secretion mechanism itself cause oxidative stress in pancreatic beta-cells? AB - Glucose-dependent insulin secretion (GDIS), reactive oxygen species (ROS) production, and oxidative stress in pancreatic beta-cells may be tightly linked processes. Here we suggest that the same pathways used in the activation of GDIS (increased glycolytic flux, ATP-to-ADP ratio, and intracellular Ca2+ concentration) can dramatically enhance ROS production and manifestations of oxidative stress and, possibly, apoptosis. The increase in ROS production and oxidative stress produced by GDIS activation itself suggests a dual role for metabolic insulin secretagogues, as an initial sharp increase in insulin secretion rate can be accompanied by progressive beta-cell injury. We propose that therapeutic strategies targeting enhancement of GDIS should be carefully considered in light of possible loss of beta-cell function and mass. PMID- 15277371 TI - PYY3-36 reinforces insulin action on glucose disposal in mice fed a high-fat diet. AB - Peptide YY(3-36) (PYY(3-36)) is released by the gut in response to nutrient ingestion. It modulates the activities of orexigenic neuropeptide Y (NPY) neurons and anorexigenic proopiomelanocortin (POMC) neurons in the hypothalamus to inhibit food intake. Because both NPY and POMC have also been shown to impact insulin action, we wondered whether PYY(3-36) could improve insulin sensitivity. To address this question, we examined the acute effect of intravenous PYY(3-36) on glucose and free fatty acid (FFA) flux during a hyperinsulinemic-euglycemic clamp in mice maintained on a high-fat diet for 2 weeks before the experiment. We also evaluated the effects of PYY(3-36) infusion on glucose uptake in muscle and adipose tissue in this experimental context. Under basal conditions, none of the metabolic parameters were affected by PYY(3-36). Under hyperinsulinemic conditions, glucose disposal was significantly increased in PYY(3-36)-infused compared with vehicle-infused mice (103.8 +/- 10.9 vs. 76.1 +/- 11.4 micromol.min(-1).kg(-1), respectively; P = 0.001). Accordingly, glucose uptake in muscle and adipose tissue was greater in PYY(3-36)-treated animals, although the difference with controls did not reach statistical significance in adipose tissue (muscle: 2.1 +/- 0.5 vs. 1.5 +/- 0.5 micromol/g tissue, P = 0.049; adipose tissue: 0.8 +/- 0.4 vs. 0.4 +/- 0.3 micromol/g tissue, P = 0.08). In contrast, PYY(3-36) did not impact insulin action on endogenous glucose production or FFA metabolism. These data indicate that PYY(3-36) reinforces insulin action on glucose disposal in mice fed a high-fat diet, through a mechanism that is independent of food intake and body weight. In contrast, it leaves glucose production and lipid flux largely unaffected in this experimental context. PMID- 15277372 TI - Potential role for AMP-activated protein kinase in hypoglycemia sensing in the ventromedial hypothalamus. AB - The mechanisms by which specialized glucose-sensing neurons within the hypothalamus are able to detect a falling blood glucose remain largely unknown but may be linked to some gauge of neuronal energy status. We sought to test the hypothesis that AMP-activated protein kinase (AMPK), an intracellular kinase purported to act as a fuel sensor, plays a role in hypoglycemia sensing in the ventromedial hypothalamus (VMH) of the Sprague-Dawley rat by chemically activating AMPK in vivo through bilateral microinjection, before performing hyperinsulinemic-hypoglycemic or hyperinsulinemic-euglycemic clamp studies. In a subgroup of rats, H3-glucose was infused to determine glucose kinetics. The additional chemical activation by AICAR of AMPK in the VMH during hypoglycemia markedly reduced the amount of exogenous glucose required to maintain plasma glucose during hypoglycemia, an effect that was almost completely accounted for by a three- to fourfold increase in hepatic glucose production in comparison to controls. In contrast, no differences were seen between groups in hypoglycemia induced rises in the principal counterregulatory hormones. In conclusion, activation of AMPK within the VMH may play an important role in hypoglycemia sensing. The combination of hypoglycemia- and AICAR-induced AMPK activity appears to result in a marked stimulus to hepatic glucose counterregulation. PMID- 15277373 TI - The regulation of glucose-excited neurons in the hypothalamic arcuate nucleus by glucose and feeding-relevant peptides. AB - Glucosensing neurons in the hypothalamic arcuate nucleus (ARC) were studied using electrophysiological and immunocytochemical techniques in neonatal male Sprague Dawley rats. We identified glucose-excited and -inhibited neurons, which increase and decrease, respectively, their action potential frequency (APF) as extracellular glucose levels increase throughout the physiological range. Glucose inhibited neurons were found predominantly in the medial ARC, whereas glucose excited neurons were found in the lateral ARC. ARC glucose-excited neurons in brain slices dose-dependently increased their APF and decreased their ATP sensitive K+ channel (KATP channel) currents as extracellular glucose levels increased from 0.1 to 10 mmol/l. However, glucose sensitivity was greatest as extracellular glucose decreased to <2.5 mmol/l. The glucokinase inhibitor alloxan increases KATP single-channel currents in glucose-excited neurons in a manner similar to low glucose. Leptin did not alter the activity of ARC glucose-excited neurons. Although insulin did not affect ARC glucose-excited neurons in the presence of 2.5 mmol/l (steady-state) glucose, they were stimulated by insulin in the presence of 0.1 mmol/l glucose. Neuropeptide Y (NPY) inhibited and alpha melanocyte-stimulating hormone stimulated ARC glucose-excited neurons. ARC glucose-excited neurons did not show pro-opiomelanocortin immunoreactivity. These data suggest that ARC glucose-excited neurons may serve an integrative role in the regulation of energy balance. PMID- 15277374 TI - Effects of different hypocaloric diets on protein secretion from adipose tissue of obese women. AB - Little is known about common factors (e.g., macronutrients and energy supply) regulating the protein secretory function of adipose tissue. We therefore compared the effects of randomly assigned 10-week hypoenergetic (-600 kcal/day) diets with moderate-fat/moderate-carbohydrate or low-fat/high-carbohydrate content on circulating levels and production of proteins (using radioimmunoassays and enzyme-linked immunosorbent assays) from subcutaneous adipose tissue in 40 obese but otherwise healthy women. Similar results were obtained by the two diets. Body weight decreased by approximately 7.5%. The secretion rate of leptin decreased by approximately 40%, as did that of tumor necrosis factor-alpha (TNF alpha), and interleukin (IL)-6 and -8 decreased by 25-30%, whereas the secretion of plasminogen activator inhibitor 1 (PAI-1) and adiponectin did not show any changes. Regarding mRNA expression (by real-time PCR), only that of leptin and IL 6 decreased significantly. Circulating levels of leptin and PAI-1 decreased by 30 and 40%, respectively, but there were only minor changes in circulating TNF alpha, IL-6, or adiponectin. In conclusion, moderate caloric restriction but not macronutrient composition influences the production and secretion of adipose tissue-derived proteins during weight reduction, leptin being the most sensitive and adiponectin and PAI-1 the least sensitive. PMID- 15277375 TI - Islet allograft survival induced by costimulation blockade in NOD mice is controlled by allelic variants of Idd3. AB - NOD mice develop type 1 autoimmune diabetes and exhibit genetically dominant resistance to transplantation tolerance induction. These two phenotypes are genetically separable. Costimulation blockade fails to prolong skin allograft survival in (NOD x C57BL/6)F1 mice and in NOD-related strains made diabetes resistant by congenic introduction of protective major histocompatibility complex (MHC) or non-MHC Idd region genes. Here, we tested the hypothesis that the genetic basis for the resistance of NOD mice to skin allograft tolerance also applies to islet allografts. Surprisingly, costimulation blockade induced permanent islet allograft survival in (NOD x C57BL/6)F1 mice but not in NOD mice. After costimulation blockade, islet allograft survival was prolonged in diabetes resistant NOD.B6 Idd3 mice and shortened in diabetes-free C57BL/6 mice congenic for the NOD Idd3 variant. Islet allograft tolerance could not be induced in diabetes-resistant NOD.B10 Idd5 and NOD.B10 Idd9 mice. The data demonstrate that 1) NOD mice resist islet allograft tolerance induction; 2) unlike skin allografts, resistance to islet allograft tolerance is a genetically recessive trait; 3) an Idd3 region gene(s) is an important determinant of islet allograft tolerance induction; and 4) there may be overlap in the mechanism by which the Idd3 resistance locus improves self-tolerance and the induction of allotolerance. PMID- 15277376 TI - Increased serum levels of MRP-8/14 in type 1 diabetes induce an increased expression of CD11b and an enhanced adhesion of circulating monocytes to fibronectin. AB - The recruitment of monocytes from the bloodstream is crucial in the accumulation of macrophages and dendritic cells in type 1 diabetic pancreases. Adhesion via integrins to endothelium and extracellular matrix proteins, such as fibronectin (FN), and the production of myeloid-related protein (MRP)-8, -14, and -8/14 by recently transmigrated monocytes are thought to be instrumental in such recruitment. We determined the FN-adhesive capacity and integrin expression of monocytes of type 1 and type 2 diabetic patients and related them to the subjects' serum levels of MRP-8, -14 and -8/14. Monocytes of type 1 diabetic patients displayed an increased adhesion to fibronectin in comparison with type 2 patients and healthy control subjects but had a normal expression of the FN binding integrins CD29, CD49a, CD49d, and CD49e (although CD11b and CD18 expression was increased). MRP-8/14, which was increased in the sera of type 1 diabetic patients, induced healthy donor monocytes to adhere to FN and upregulate CD11b expression in a dosage-dependent manner. The observed MRP-induced increased adhesion of monocytes to FN and upregulation of CD11b most likely contributed to a facilitated accumulation of monocytes and monocyte-derived cells at the site of inflammation, in this case the pancreatic islets. PMID- 15277377 TI - GAD65-specific CD4+ T-cells with high antigen avidity are prevalent in peripheral blood of patients with type 1 diabetes. AB - Negative selection of self-reactive T-cells during thymic development, along with activation-induced cell death in peripheral lymphocytes, is designed to limit the expansion and persistence of autoreactive T-cells. Autoreactive T-cells are nevertheless present, both in patients with type 1 diabetes and in at-risk subjects. By using MHC class II tetramers to probe the T-cell receptor (TcR) specificity and avidity of GAD65 reactive T-cell clones isolated from patients with type 1 diabetes, we identified high-avidity CD4+ T-cells in peripheral blood, coexisting with low-avidity cells directed to the same GAD65 epitope specificity. A variety of cytokine patterns was observed, even among T-cells with high MHC-peptide avidity, and the clones utilize a biased set of TcR genes that favor two combinations, Valpha12-beta5.1 and Valpha17-Vbeta4. Presence of these high-avidity TcRs indicates a failure to delete autoreactive T-cells that likely arise from oligoclonal expansion in response to autoantigen exposure during the progression of type 1 diabetes. PMID- 15277378 TI - Flt3-ligand treatment prevents diabetes in NOD mice. AB - The mechanism by which mixed chimerism reverses autoimmunity in type 1 diabetes has not been defined. NOD mice have a well-characterized defect in the production of myeloid progenitors that is believed to contribute significantly to the autoimmune process. We therefore investigated whether chimerism induces a correction of this defect. Mixed chimerism restored production of myeloid progenitors in NOD mice to normal levels. Notably, NOD bone marrow cells as well as donor bone marrow cells produced the mature myeloid progeny, and the level of donor chimerism was not correlated with the degree of restoration of the defect. Moreover, NOD bone marrow cells cultured with Flt3-ligand developed a heat-stable antigen-positive/Ly6C+ population comprised primarily of mature myeloid dendritic cells, suggesting that the underlying abnormality is not cell intrinsic but rather due to a block in development of mature myeloid progeny, including myeloid dendritic cells. Strikingly, treatment of NOD mice with Flt3-ligand significantly decreased insulitis and progression to diabetes and was associated with a significant increase in myeloid dendritic cells and in vivo induction of CD4+/CD25+ cells in the pancreatic lymph node. Therefore, Flt3-ligand treatment and/or the establishment of mixed chimerism in prediabetic candidates may provide a benign and novel approach to treat diabetes. PMID- 15277379 TI - Hyperresponsiveness, resistance to B-cell receptor-dependent activation-induced cell death, and accumulation of hyperactivated B-cells in islets is associated with the onset of insulitis but not type 1 diabetes. AB - B-cells proliferate after B-cell receptor (BCR) stimulation and are deleted by activation-induced cell death (AICD) during negative selection. We report that B cells from type 1 diabetes-susceptible NOD and type 1 diabetes-resistant but insulitis-prone congenic NOD.B6Idd4B and NOR mice, relative to B-cells from nonautoimmune disease-prone C57BL/6 and BALB/c mice, display a hyperproliferative response to BCR stimulation and lower activation threshold in the absence or presence of interleukin 4 (IL-4). This hyperproliferation is associated with an increased proportion of NOD and NOR B-cells that enter into the S phase of the cell cycle and undergo cell division. The relative resistance to BCR-induced AICD of B-cells from NOD, NOR, and NOD.B6Idd4B mice, all of which develop insulitis, correlates with the presence of a higher percentage of hyperactivated B-cells in the spleen and islets of these mice than in nonautoimmune disease-prone C57BL/6 and BALB/c mice. The NOD islet-infiltrated activated B-cells are more responsive to further stimulation by IL-4 than activated spleen B-cells. Our results suggest that resistance to AICD and accumulation of hyperactivated B-cells in islets is associated with the onset of an inflammatory insulitis, but not type 1 diabetes. PMID- 15277380 TI - Major histocompatibility complex class I shedding and programmed cell death stimulated through the proinflammatory P2X7 receptor: a candidate susceptibility gene for NOD diabetes. AB - It has been hypothesized that type 1 diabetes is initiated by neonatal physiological pancreatic beta-cell death, indicating that the early stages of this autoimmune response may reflect a dysregulated response to immune "danger" signals. One potential danger signal is ATP, high concentrations of which stimulate the purinergic receptor P2X7 on hematopoietic cells. We compared the sensitivity of lymphocytes from model type 1 diabetic (NOD) and control (C57BL/10) mice to activation of this pathway. Stimulation of the P2X7 receptor of NOD mice resulted in more pronounced shedding of the lymphocyte homing receptor CD62L and in increased programmed cell death. Levels of major histocompatibility complex class I molecules, which have previously been reported to be poorly expressed on NOD lymphocytes, were initially normal, but the molecules were shed preferentially from NOD cells after P2X7 receptor stimulation. Thus, although NOD lymphocytes have been considered resistant to programmed cell death, they are highly sensitive to that stimulated through the P2X7 receptor. Because NOD mice express a low activation threshold allele of the P2X7 receptor and the P2X7 gene maps to a locus associated with disease, P2X7 is a good candidate susceptibility gene for NOD diabetes. PMID- 15277381 TI - Autocrine regulation of single pancreatic beta-cell survival. AB - Function and survival of cells depend in part on the presence of growth factors. We explored the autocrine regulation of insulin and nerve growth factor (NGF) on single adult rat pancreatic beta-cell survival and hormone secretion. When NGF or insulin signaling were blocked in culture media, cell survival decreased compared with control cells, with apoptosis being the main mechanism of cell death. To further explore the role of glucose in beta-cell survival, we cultured the cells for 16 h in 2.6 mmol/l glucose and observed that nearly 17% of the cells developed apoptosis; this effect was partially prevented by NGF and almost completely inhibited by insulin treatment. A high K+ concentration had the same effect, suggesting that insulin and NGF secretion by the cells was responsible for the survival effects and not glucose per se. Blocking NGF signaling with an NGF antibody or with K252a reduced insulin biosynthesis and secretion in the cells that survived the treatment. Moreover, the functional beta-cell subpopulation with a higher insulin secretion rate is more susceptible to K252a. These results further indicate that NGF and insulin play important autoregulatory roles in pancreatic beta-cell survival and function and strongly suggest the need to explore new focuses in diabetes treatment. PMID- 15277382 TI - Inhibition of activin signaling induces pancreatic epithelial cell expansion and diminishes terminal differentiation of pancreatic beta-cells. AB - Activins regulate the growth and differentiation of a variety of cells. During pancreatic islet development, activins are required for the specialization of pancreatic precursors from the gut endoderm during midgestation. In this study, we probed the role of activin signaling during pancreatic islet cell development and regeneration. Indeed, we found that both activins and activin receptors are upregulated in duct epithelial cells during islet differentiation. Interestingly, the expression of endogenous cellular inhibitors of activin signaling, follistatin and Cripto, were also found to be augmented. Inhibition of activins significantly enhanced survival and expansion of pancreatic epithelial cells but decreased the numbers of differentiated beta-cells. Our results suggest that the homeostasis of growth and terminal differentiation requires a precise context dependent regulation of activin signaling. Follistatin participates in this process by promoting expansion of precursor cells during pancreas growth. PMID- 15277383 TI - Extracellular matrix protects pancreatic beta-cells against apoptosis: role of short- and long-term signaling pathways. AB - We have shown previously that culture of beta-cells on matrix derived from 804G cells and rich in laminin-5 improves their function. The purpose of this study was to investigate whether this matrix protects beta-cells against apoptosis and to elucidate signaling pathways involved. Matrix protected sorted rat beta-cells against apoptosis under standard conditions (11.2 mmol/l glucose, 10% serum), after serum deprivation (1% serum), and in response to interleukin-1beta (IL 1beta; 2 ng/ml), compared with control (poly-L-lysine [pLL]). Caspase-8 activity was reduced in cells cultured on matrix, whereas focal adhesion kinase (FAK), protein kinase B (PKB, or Akt), and extracellular signal-regulated kinase (ERK) phosphorylation was augmented. Treatment (4 h) with an anti-beta1 integrin antibody, with the ERK pathway inhibitor PD98059, and/or with the phosphatidylinositol 3-kinase inhibitor LY294002 augmented cell death on 804G matrix but not on pLL. In long-term assays (48 h), PD98059 but not LY294002 drastically augmented cell death on 804G matrix but did so to a lesser extent on pLL. The protein inhibitor of nuclear factor-kappaB (IkappaBalpha) was overexpressed in cells cultured 18 h on matrix with partial blockade by PD98059. In summary, this study provides evidence for activation of signaling pathways and gene expression by extracellular matrix leading to improved beta-cell survival. PMID- 15277384 TI - Insulin dose-response curves for stimulation of splanchnic glucose uptake and suppression of endogenous glucose production differ in nondiabetic humans and are abnormal in people with type 2 diabetes. AB - To determine whether the insulin dose-response curves for suppression of endogenous glucose production (EGP) and stimulation of splanchnic glucose uptake (SGU) differ in nondiabetic humans and are abnormal in type 2 diabetes, 14 nondiabetic and 12 diabetic subjects were studied. Glucose was clamped at approximately 9.5 mmol/l and endogenous hormone secretion inhibited by somatostatin, while glucagon and growth hormone were replaced by an exogenous infusion. Insulin was progressively increased from approximately 150 to approximately 350 and approximately 700 pmol/l by means of an exogenous insulin infusion, while EGP, SGU, and leg glucose uptake (LGU) were measured using the splanchnic and leg catheterization methods, combined with a [3-3H]glucose infusion. In nondiabetic subjects, an increase in insulin from approximately 150 to approximately 350 pmol/l resulted in maximal suppression of EGP, whereas SGU continued to increase (P < 0.001) when insulin was increased to approximately 700 pmol/l. In contrast, EGP progressively decreased (P < 0.001) and SGU progressively increased (P < 0.001) in the diabetic subjects as insulin increased from approximately 150 to approximately 700 pmol/l. Although EGP was higher (P < 0.01) in the diabetic than nondiabetic subjects only at the lowest insulin concentration, SGU was lower (P < 0.01) in the diabetic subjects at all insulin concentrations tested. On the other hand, in contrast to LGU and overall glucose disposal, the increment in SGU in response to both increments in insulin did not differ in the diabetic and nondiabetic subjects, implying a right shifted but parallel dose-response curve. These data indicate that the dose-response curves for suppression of glucose production and stimulation of glucose uptake differ in nondiabetic subjects and are abnormal in people with type 2 diabetes. Taken together, these data also suggest that agents that enhance SGU in diabetic patients (e.g. glucokinase activators) are likely to improve glucose tolerance. PMID- 15277385 TI - Splanchnic cortisol production occurs in humans: evidence for conversion of cortisone to cortisol via the 11-beta hydroxysteroid dehydrogenase (11beta-hsd) type 1 pathway. AB - Glucocorticoids are potent regulators of protein, fat, and carbohydrate metabolism. To determine if cortisol production occurs within the splanchnic bed in humans, 11 nondiabetic subjects were studied using the hepatic/leg catheterization method along with an infusion of [9,11,12,12-2H4] cortisol (D4 cortisol) as proposed by Andrews et al. In the fasting state, there was net release (P < 0.05) of cortisol from the splanchnic bed (6.1 +/- 2.6 microg/min) and net uptake (P < 0.05) by the leg (1.7 +/- 0.7 microg/min). This, along with cortisol production by other tissues (e.g., the adrenals), resulted in a total body cortisol appearance rate of 18.1 +/- 1.9 microg/min. Fractional splanchnic D4-cortisol extraction averaged 12.9 +/- 1.3% (P < 0.001), splanchnic cortisol uptake 14.8 +/- 2.0 microg/min (P < 0.001), and splanchnic cortisol production 22.2 +/- 3.3 microg/min (P < 0.001). On the other hand, fractional leg D4 cortisol extraction averaged 5.6 +/- 1.8% (P < 0.02), leg cortisol uptake 2.3 +/- 0.7 microg/min (P < 0.01), and leg cortisol production 0.4 +/- 0.4 microg/min, which did not differ from zero. Because D4-cortisol loses a deuterium during conversion to [9,12,12-2H3] cortisone (D3-cortisone), which in turn generates [9,12,12(2)H3] cortisol (D3-cortisol) via 11-beta hydroxysteroid dehydrogenase (11beta-HSD) type 1, D3-cortisol production can be used as an index of 11beta-HSD type 1 activity. Net splanchnic D3-cortisol release (3.9 +/- 0.4 microg/min) and splanchnic D3-cortisol production (7.1 +/- 0.7 microg/min) occurred (P < 0.01) in all subjects. In contrast, there was minimal leg D3-cortisol production (0.04 +/- 0.01 microg/min), resulting in a strong correlation between splanchnic D3 cortisol production and total-body 3D-cortisol production in both the fasting state (r = 0.84; P < 0.02) and during an infusion of insulin (r = 0.97; P < 0.01). Thus, splanchnic production of cortisol occurs in nondiabetic humans at rates approximating that which occurs in the remainder of the body. These data support the possibility that alterations in splanchnic cortisol production contribute to visceral fat accumulation and the hepatic insulin resistance of obesity or type 2 diabetes. PMID- 15277386 TI - Interactions between endothelin and nitric oxide in the regulation of vascular tone in obesity and diabetes. AB - Endothelial dysfunction reflects an imbalance of vasodilators and vasoconstrictors. Endogenous endothelin activity seems to be increased in human obesity and type 2 diabetes, and cellular studies suggest that this factor may itself reduce bioavailable nitric oxide (NO). We studied 20 lean, 20 obese, and 14 type 2 diabetic individuals under three protocols, measuring leg vascular responses to intra-arterial infusions of NG-monomethyl-l-arginine (l-NMMA; an inhibitor of NO synthase) alone or in combination with BQ123 (an antagonist of type A endothelin receptors) or phentolamine (used as a control vasodilator). NO synthase inhibition alone (study 1) produced an approximately 40% increase in leg vascular resistance (LVR) in all three participant groups, which was not statistically different across groups (increase in LVR: lean, 135 +/- 28; obese, 140 +/- 32; type 2 diabetic, 184 +/- 51 units; NS). By design, BQ123 at the infused rate of 3 micromol/min produced equivalent approximately 35% reductions in LVR across groups. The subsequent addition of l-NMMA produced a greater increase in LVR among obese participants than lean or type 2 diabetic participants (study 2: lean, 182 +/- 48; obese, 311 +/- 66; type 2 diabetic, 186 +/- 40; P = 0.07). Compared with study 1, the effect of l-NMMA was magnified by BQ123 in obese participants but not in lean or type 2 diabetic participants (P = 0.005, study 1 vs. 2; P = 0.03 for group effect). Phentolamine (75 mg/min) produced vasodilation in obese participants comparable to that seen with BQ123 but failed to augment the L-NMMA response. Endothelin antagonism unmasks or augments NO synthesis capacity in obese but not type 2 diabetic participants. This suggests that impaired NO bioavailability as a result of endogenous endothelin may contribute to endothelial dysfunction in obesity, in addition to direct vasoconstrictor effects of endothelin. In contrast, endothelin antagonism alone is insufficient to restore impaired NO bioavailability in diabetes. PMID- 15277387 TI - Partial gene deletion of endothelial nitric oxide synthase predisposes to exaggerated high-fat diet-induced insulin resistance and arterial hypertension. AB - Nitric oxide (NO) plays a major role in the regulation of cardiovascular and metabolic homeostasis, as evidenced by insulin resistance and arterial hypertension in endothelial NO synthase (eNOS) null mice. Extrapolation of these findings to humans is difficult, however, because eNOS gene deficiency has not been reported. eNOS gene polymorphism and impaired NO synthesis, however, have been reported in several cardiovascular disease states and could predispose to insulin resistance. High-fat diet induces insulin resistance and arterial hypertension in normal mice. To test whether partial eNOS deficiency facilitates the development of insulin resistance and arterial hypertension during metabolic stress, we examined effects of an 8-week high-fat diet on insulin sensitivity (euglycemic clamp) and arterial pressure in eNOS(+/-) mice. When fed a normal diet, these mice had normal insulin sensitivity and were normotensive. When fed a high-fat diet, however, eNOS(+/-) mice developed exaggerated arterial hypertension and had fasting hyperinsulinemia and a 35% lower insulin-stimulated glucose utilization than control mice. The partial deletion of the eNOS gene does not alter insulin sensitivity or blood pressure in mice. When challenged with nutritional stress, however, partial eNOS deficiency facilitates the development of insulin resistance and arterial hypertension, providing further evidence for the importance of this gene in linking metabolic and cardiovascular disease. PMID- 15277388 TI - Vasomotor responses to hypoxia in type 2 diabetes. AB - Type 2 diabetes is associated with vascular dysfunction, accelerated atherosclerotic morbidity, and mortality. Abnormal vasomotor responses to chemoreflex activation may contribute to the acceleration of atherosclerotic diabetes complications, but these responses have not previously been investigated. We measured forearm mean blood flow (MBF) and mean vascular conductance (MVC) responses to isocapnic hypoxia in seven healthy and eight type 2 diabetic subjects during local intra-arterial saline infusion and alpha adrenergic blockade (phentolamine). The effects of hypoxia on saline and phentolamine responses significantly differed between groups; relative to normoxia, the %DeltaMVC with hypoxia during saline was -3.3 +/- 11.2% in control and 24.8 +/- 13.3% in diabetic subjects, whereas phentolamine increased hypoxic %DeltaMVC to similar levels (39.4 +/- 9.7% in control subjects and 48.0 +/- 11.8% in diabetic subjects, P < 0.05, two-way ANOVA). Absolute normoxic MBF responses during saline infusion were 91.9 +/- 21.1 and 77.9 +/- 15.3 in control and diabetic subjects, respectively, and phentolamine increased normoxic MBF to similar levels (165.2 +/- 40.1 ml/min in control subjects and 175.9 +/- 32.0 ml/min in diabetic subjects; both P < 0.05). These data indicate that diabetic and control subjects exhibit similar responses to hypoxia in the presence of alpha-adrenergic blockade despite evidence of exaggerated alpha-mediated vasoconstriction at rest. PMID- 15277389 TI - Proinflammatory cytokines, markers of cardiovascular risks, oxidative stress, and lipid peroxidation in patients with hyperglycemic crises. AB - Acute and chronic hyperglycemia are proinflammatory states, but the status of proinflammatory cytokines and markers of oxidative stress and cardiovascular risks is not known in hyperglycemic crises of diabetic ketoacidosis (DKA) and nonketotic hyperglycemia (NKH). We studied 20 lean and 28 obese patients with DKA, 10 patients with NKH, and 12 lean and 12 obese nondiabetic control subjects. We measured 1) proinflammatory cytokines (tumor necrosis factor-alpha, interleukin [IL]-6, IL1-beta, and IL-8), 2) markers of cardiovascular risk (C reactive protein [CRP], homocysteine, and plasminogen activator inhibitor-1 [PAI 1]), 3) products of reactive oxygen species (ROS; thiobarbituric acid [TBA] reacting material, and dichlorofluorescein [DCF]), and 4) cortisol, growth hormone (GH), and free fatty acids (FFAs) on admission (before insulin therapy) and after insulin therapy and resolution of hyperglycemia and/or ketoacidosis. Results were compared with lean and obese control subjects. Circulating levels of cytokines, TBA, DCF, PAI-1, FFAs, cortisol, and GH on admission were significantly increased two- to fourfold in patients with hyperglycemic crises compared with control subjects, and they returned to normal levels after insulin treatment and resolution of hyperglycemic crises. Changes in CRP and homocysteine in response to insulin therapy did not reach control levels after resolution of hyperglycemia. We conclude that DKA and NKH are associated with elevation of proinflammatory cytokines, ROS, and cardiovascular risk factors in the absence of obvious infection or cardiovascular pathology. Return of these values to normal levels with insulin therapy demonstrates a robust anti-inflammatory effect of insulin. PMID- 15277390 TI - Intra-abdominal fat is a major determinant of the National Cholesterol Education Program Adult Treatment Panel III criteria for the metabolic syndrome. AB - The underlying pathophysiology of the metabolic syndrome is the subject of debate, with both insulin resistance and obesity considered as important factors. We evaluated the differential effects of insulin resistance and central body fat distribution in determining the metabolic syndrome as defined by the National Cholesterol Education Program (NCEP) Adult Treatment Panel III. In addition, we determined which NCEP criteria were associated with insulin resistance and central adiposity. The subjects, 218 healthy men (n = 89) and women (n = 129) with a broad range of age (26-75 years) and BMI (18.4-46.8 kg/m2), underwent quantification of the insulin sensitivity index (Si) and intra-abdominal fat (IAF) and subcutaneous fat (SCF) areas. The metabolic syndrome was present in 34 (15.6%) of subjects who had a lower Si [median: 3.13 vs. 6.09 x 10(-5) min( 1)/(pmol/l)] and higher IAF (166.3 vs. 79.1 cm2) and SCF (285.1 vs. 179.8 cm2) areas compared with subjects without the syndrome (P < 0.001). Multivariate models including Si, IAF, and SCF demonstrated that each parameter was associated with the syndrome. However, IAF was independently associated with all five of the metabolic syndrome criteria. In multivariable models containing the criteria as covariates, waist circumference and triglyceride levels were independently associated with Si and IAF and SCF areas (P < 0.001). Although insulin resistance and central body fat are both associated with the metabolic syndrome, IAF is independently associated with all of the criteria, suggesting that it may have a pathophysiological role. Of the NCEP criteria, waist circumference and triglycerides may best identify insulin resistance and visceral adiposity in individuals with a fasting plasma glucose <6.4 mmol/l. PMID- 15277391 TI - Impaired glucose tolerance, but not impaired fasting glucose, is associated with increased levels of coronary heart disease risk factors: results from the Baltimore Longitudinal Study on Aging. AB - Impaired fasting glucose (IFG) and impaired glucose tolerance (IGT) identify individuals at high risk for progression to diabetes. Whether IFG and IGT have comparable coronary heart disease (CHD) risk factor profiles, independent of their progression to diabetes, is unclear. We determined CHD risk factor levels in 937 nondiabetic individuals at baseline and biannually over a mean follow-up period of 9.5 years. Subjects had no known CHD at baseline and had > or =2 (mean 4.2) oral glucose tolerance tests during follow-up. We classified glucose tolerance categories using American Diabetes Association diagnostic criteria or modified criteria that redefined IFG as 100-126 mg/dl, creating a similar baseline prevalence of IFG and IGT. Subjects who developed diabetes during follow up were excluded from our analysis. Baseline CHD risk factors were similar in subjects with normal glucose tolerance (NGT) and IFG, but significantly more atherogenic in those with IGT or IFG + IGT. These findings were unchanged when the modified criteria were used, suggesting that IGT is phenotypically different from IFG and is associated with increased levels of CHD risk factors. Subjects with isolated IFG had similar levels of CHD risk factors as NGT subjects, even when IFG was redefined with a lower threshold. Although CHD risk factors were increased in the IGT group, the incidence of CHD events was not significantly different among groups, perhaps owing to the limited number of events. The differences in CHD risk factors among prediabetic groups may have clinical implications for screening strategies and CHD risk stratification of individuals with IFG and IGT. PMID- 15277392 TI - Diminished loss of proteoglycans and lack of albuminuria in protein kinase C alpha-deficient diabetic mice. AB - Activation of protein kinase C (PKC) isoforms has been implicated in the pathogenesis of diabetic nephropathy. We showed earlier that PKC-alpha is activated in the kidneys of hyperglycemic animals. We now used PKC-alpha(-/-) mice to test the hypothesis that this PKC isoform mediates streptozotocin-induced diabetic nephropathy. We observed that renal and glomerular hypertrophy was similar in diabetic wild-type and PKC-alpha(-/-) mice. However, the development of albuminuria was almost absent in the diabetic PKC-alpha(-/-) mice. The hyperglycemia-induced downregulation of the negatively charged basement membrane heparan sulfate proteoglycan perlecan was completely prevented in the PKC-alpha( /-) mice, compared with controls. We then asked whether transforming growth factor-beta1 (TGF-beta1) and/or vascular endothelial growth factor (VEGF) is implicated in the PKC-alpha-mediated changes in the basement membrane. The hyperglycemia-induced expression of VEGF165 and its receptor VEGF receptor II (flk-1) was ameliorated in PKC-alpha(-/-) mice, whereas expression of TGF-beta1 was not affected by the lack of PKC-alpha. Our findings indicate that two important features of diabetic nephropathy-glomerular hypertrophy and albuminuria are differentially regulated. The glucose-induced albuminuria seems to be mediated by PKC-alpha via downregulation of proteoglycans in the basement membrane and regulation of VEGF expression. Therefore, PKC-alpha is a possible therapeutic target for the prevention of diabetic albuminuria. PMID- 15277393 TI - Oxidative stress affects synaptosomal gamma-aminobutyric acid and glutamate transport in diabetic rats: the role of insulin. AB - Evidence suggests that oxidative stress is involved in the pathophysiology of diabetic complications and that insulin has a neuroprotective role in oxidative stress conditions. In this study, we evaluated the in vitro effect of insulin in the susceptibility to oxidative stress and in the transport of the amino acid neurotransmitters gamma-aminobutyric acid (GABA) and glutamate in a synaptosomal fraction isolated from male type 2 diabetic Goto-Kakizaki (GK) rat brain cortex. The ascorbate/Fe(2+)-induced increase in thiobarbituric acid reactive substances (TBARSs) was similar in Wistar and GK rats and was not reverted by insulin (1 micromol/l), suggesting that other mechanisms, rather than a direct effect in membrane lipid peroxidation, may mediate insulin neuroprotection. Diabetes did not affect GABA and glutamate transport, despite the significant decrease in membrane potential and ATP/ADP ratio, and insulin increased the uptake of both GABA and glutamate in GK rats. Upon oxidation, there was a decrease in the uptake of both neurotransmitters and an increase in extrasynaptosomal glutamate levels and in ATP/ADP ratio in GK rats. Insulin treatment reverted the ascorbate/Fe(2+) induced decrease in GABA accumulation, with a decrease in extrasynaptosomal GABA. These results suggest that insulin modulates synaptosomal GABA and/or glutamate transport, thus having a neuroprotective role under oxidizing and/or diabetic conditions. PMID- 15277394 TI - Surface expression of collagen receptor Fc receptor-gamma/glycoprotein VI is enhanced on platelets in type 2 diabetes and mediates release of CD40 ligand and activation of endothelial cells. AB - Diabetes is associated with an enhanced collagen-mediated platelet activation that contributes significantly to thromboischemic complications. In this study, the platelet collagen receptor glycoprotein VI (GPVI) was studied in 385 patients with type 2 diabetes. Surface expression of the platelet Fc receptor that forms a functional complex with GPVI was significantly increased in patients with diabetes compared with those without diabetes (P = 0.02). Fc receptor expression correlated with GPVI expression and was found to be independently associated with diabetes (r = 0.529, P < 0.001). Stimulation of GPVI through a specific anti-GPVI monoclonal antibody significantly enhanced surface expression of CD40L (P = 0.006). Because CD40L is a potent platelet-derived cytokine that is involved in thrombosis and atherosclerosis, we evaluated the effect of GPVI-mediated release of CD40L on activation of endothelial cells. Coincubation of GPVI-stimulated platelets resulted in substantial enhanced endothelial surface expression of CD62P, alphavbeta3, and intercellular adhesion molecule 1 (P < 0.05) and secretion of monocyte chemoattractant protein 1 of cultured human umbilical vein endothelial cells (P < 0.01). These results suggest that the function of collagen receptor GPVI is altered in type 2 diabetes and may play an important role in atherothrombotic complications. Inhibition of GPVI may be a promising pharmacological target in the treatment of high-risk diabetic patients. PMID- 15277395 TI - Polymorphic variations in the neurogenic differentiation-1, neurogenin-3, and hepatocyte nuclear factor-1alpha genes contribute to glucose intolerance in a South Indian population. AB - The neurogenic differentiation-1 (NEUROD1), neurogenin-3 (NEUROG3), and hepatic nuclear factor-1alpha (TCF1) genes are interacting transcription factors implicated in controlling islet cell development and insulin secretion. Polymorphisms of these genes (Ala45Thr [NEUROD1], Ser199Phe [NEUROG3], and Ala98Val [TCF1]) have been postulated to influence the development of type 2 diabetes. We have investigated the role and interaction between these variants using PCR/restriction fragment-length polymorphism assays in 454 subjects recruited as part of a population survey in South India. Additionally, 97 South Indian parent-offspring trios were studied. Polymorphisms of all three genes were associated with either fasting blood glucose (FBG) and/or 2-h blood glucose (BG) in either the total dataset or when restricted to a normoglycemic population. A monotonically increasing effect, dependent on the total number of risk-associated alleles carried, was observed across the whole population (P < 0.0001 for FBG and 2-h BG), raising FBG by a mean of 2.9 mmol/l and 2-h BG by a mean of 4.3 mmol/l. Similarly, an ascending number of the same risk alleles per subject increased the likelihood of type 2 diabetes (P = 0.002). In conclusion, we observed a combined effect of variations in NEUROD1, NEUROG3, and TCF1 in contributing to overall glucose intolerance in a South Indian population. PMID- 15277396 TI - Variation at the insulin gene VNTR (variable number tandem repeat) polymorphism and early growth: studies in a large Finnish birth cohort. AB - Variation at the insulin gene (INS-)VNTR (variable number of tandem repeats) minisatellite polymorphism has been reported to be associated with both early growth and adult metabolic phenotypes. However, the samples studied have been small and the relationship between INS-VNTR variation and parameters of early growth inconsistent, with four previous studies producing conflicting results. We have studied the relationship between INS-VNTR class (measured by genotyping the nearby -23HphI variant with which it is in tight linkage disequilibrium) and early growth in 5,646 members of the Northern Finnish Birth Cohort of 1966. Comparing class III homozygotes with other genotypes using multivariate linear regression analysis, we found no significant associations with any early growth measure (birth weight, birth length, ponderal index, and head circumference at 1 year), even after stratifying subjects by growth trajectory during infancy and/or birth order. For example, among infants with limited postnatal growth realignment (n = 2,470), class III/III infants were no heavier at birth (difference [+/-SE] in the means [fully adjusted], 58 +/- 51 g; P = 0.26) than class I/- infants. No significant associations were detected following reanalysis with an additive model (for example, for birth weight, beta = 20 g [95% CI -3 to 44], P = 0.09). Studies of this large population-based cohort have failed to generate convincing evidence that INS-VNTR variation influences early growth. PMID- 15277397 TI - Adiponectin receptor 1 gene (ADIPOR1) as a candidate for type 2 diabetes and insulin resistance. AB - Considerable data support adiponectin as an important adipose-derived insulin sensitizer that enhances fatty acid oxidation and alters hepatic gluconeogenesis. Adiponectin acts by way of two receptors, ADIPOR1 and ADIPOR2. ADIPOR1 is widely expressed in tissues, including muscle, liver, and pancreas, and binds the globular form of adiponectin with high affinity. To test the hypothesis that sequence variations in or near the ADIPOR1 gene contribute to the risk of developing type 2 diabetes and the metabolic syndrome, we screened the eight exons (including the untranslated exon 1) of the ADIPOR1 gene with flanking intronic sequences and the 5' and 3' flanking sequences. We identified 22 single nucleotide polymorphisms (SNPs) in Caucasian and African-American subjects, of which a single nonsynonymous SNP (N44K) in exon 2 was present only in African American subjects. We typed 14 sequence variants that had minor allele frequencies >5%. No SNP was associated with type 2 diabetes in Caucasians or African Americans, and no SNP was a determinant of insulin sensitivity or insulin secretion among nondiabetic members of high-risk Caucasian families. However, the two alleles of a SNP in the 3' untranslated region were expressed unequally, and ADIPOR1 mRNA levels were significantly lower among transformed lymphocytes from diabetic African-American individuals than among control cell lines. This altered gene expression might suggest a role for ADIPOR1 in the metabolic syndrome. PMID- 15277398 TI - Quantitative trait loci near the insulin-degrading enzyme (IDE) gene contribute to variation in plasma insulin levels. AB - Insulin-degrading enzyme (IDE) plays a principal role in the proteolysis of several peptides in addition to insulin and is encoded by IDE, which resides in a region of chromosome 10q that is linked to type 2 diabetes. Two recent studies presented genetic association data on IDE and type 2 diabetes (one positive and the other negative), but neither explored the fundamental question of whether polymorphism in IDE has a measurable influence on insulin levels in human populations. To address this possibility, 14 single nucleotide polymorphisms (SNPs) from a linkage disequilibrium block encompassing IDE have been genotyped in a sample of 321 impaired glucose tolerant and 403 nondiabetic control subjects. Analyses based on haplotypic genotypes (diplotypes), constructed with SNPs that differentiate common extant haplotypes extending across IDE, provided compelling evidence of association with fasting insulin levels (P = 0.0009), 2-h insulin levels (P = 0.0027), homeostasis model assessment of insulin resistance (P = 0.0001), and BMI (P = 0.0067), with effects exclusively evident in men. The strongest evidence for an effect of a single marker was obtained for rs2251101 (located near the 3' untranslated region of IDE) on 2-h insulin levels (P = 0.000023). Diplotype analyses, however, suggest the presence of multiple interacting trait-modifying sequences in the region. Results indicate that polymorphism in/near IDE contributes to a large proportion of variance in plasma insulin levels and correlated traits, but questions of sex specificity and allelic heterogeneity will need to be taken into consideration as the molecular basis of the observed phenotypic effects unfolds. PMID- 15277399 TI - Prospective isolation of multipotent pancreatic progenitors using flow-cytometric cell sorting. AB - During pancreatic development, neogenesis, and regeneration, stem cells might act as a central player to generate endocrine, acinar, and duct cells. Although these cells are well known as pancreatic stem cells (PSCs), indisputable proof of their existence has not been reported. Identification of phenotypic markers for PSCs leads to their prospective isolation and precise characterization to clear whether stem cells exist in the pancreas. By combining flow cytometry and clonal analysis, we show here that a possible pancreatic stem or progenitor cell candidate that resides in the developing and adult mouse pancreas expresses the receptor for the hepatocyte growth factor (HGF) c-Met, but does not express hematopoietic and vascular endothelial antigens such as CD45, TER119, c-Kit, and Flk-1. These cells formed clonal colonies in vitro and differentiated into multiple pancreatic lineage cells from single cells. Some of them could largely expand with self-renewing cell divisions in culture, and, following cell transplantation, they differentiated into pancreatic endocrine and acinar cells in vivo. Furthermore, they produced cells expressing multiple markers of nonpancreatic organs including liver, stomach, and intestine in vitro. Our data strongly suggest that c-Met/HGF signaling plays an important role in stem/progenitor cell function in both developing and adult pancreas. By using this antigen, PSCs could be isolated prospectively, enabling a detailed investigation of stem cell markers and application toward regenerative therapies for diabetes. PMID- 15277400 TI - SREBF-1 gene polymorphisms are associated with obesity and type 2 diabetes in French obese and diabetic cohorts. AB - Sterol regulatory element-binding protein (SREBP)-1 transcription factors play a central role in energy homeostasis by promoting glycolysis, lipogenesis, and adipogenesis. The sterol regulatory element-binding protein gene (SREBF)-1 is a good candidate gene for obesity and obesity-related metabolic traits such as type 2 diabetes and dyslipidemia. The SREBF-1 molecular screening of 40 unrelated obese patients by PCR/single-strand conformation polymorphism revealed 19 single nucleotide polymorphisms (SNPs). Six SNPs were genotyped for an association study in large French obese and nonobese cohorts. Case-control studies using two independent nonobese cohorts indicated that SNP17 (54G/C, exon 18c) is associated with morbid obesity (odds ratio 1.5, P = 0.006 and P = 0.02, respectively). SNP3 (-150G/A, exon 1a), SNP5 (-36delG, exon 1a), and SNP17 are found in high linkage disequilibrium (D' > 0.8). The haplotype including wild-type alleles of these SNPs (C/G/G/T/C/G, HAP2) is identified as a risk factor for morbid obesity (P = 0.003). In the obese group, SNP3, SNP5, and SNP17 are associated with male specific hypertriglyceridemia (P = 0.07, P = 0.01, and P = 0.05, respectively). SNP17 is also associated with type 2 diabetes (P = 0.03) and increased prevalence of nephropathy (P = 0.028) in a diabetic cohort. Our results indicate a role of the SREBF-1 gene in genetic predisposition of metabolic diseases such as obesity, type 2 diabetes, and dyslipidemia. PMID- 15277401 TI - DPB1 alleles are associated with type 1 diabetes susceptibility in multiple ethnic groups. AB - Genetic associations between type 1 diabetes and alleles at the HLA class II locus DPB1 have been previously reported. Observed associations could be due to variation in the DPB1 locus itself or to linkage disequilibrium (LD) between DPB1 alleles and other susceptibility loci. One measure of whether the association of an allele with a disease reflects a true effect of the locus or is simply due to LD is the observation of that association in multiple ethnic groups. Previous type 1 diabetes associations have been reported for DPB1*0301 and DPB1*0202 (predisposing) and for DPB1*0402 (protective). In this study, results are reported from testing these associations in three different sample sets: 1) Puerto Rican case and control subjects, 2) Mexican-American simplex families, and 3) high-risk (DR3/DR4) individuals with and without an affected relative. DPB1*0301 was associated in all three groups, even after accounting for LD with DRB1-DQB1. DPB1*0202 and DPB1*0402 were positively and negatively associated, respectively, in two of the three populations. These results suggest that the observed DPB1 associations, especially that of the DPB1*0301 allele, with type 1 diabetes are likely to be true associations. This supports the concept that multiple genes in the HLA region can affect type 1 diabetes susceptibility. PMID- 15277402 TI - Severe persistent hyperinsulinemic hypoglycemia due to a de novo glucokinase mutation. AB - Glucokinase (GK) is a glycolytic key enzyme that functions as a glucose sensor in the pancreatic beta-cell, where it governs glucose-stimulated insulin secretion (GSIS). Heterozygous inactivating mutations in the glucokinase gene (GCK) cause a mild form of diabetes (maturity-onset diabetes of the young [MODY]2), and activating mutations have been associated with a mild form of familial hyperinsulinemic hypoglycemia. We describe the first case of severe persistent hyperinsulinemic hypoglycemia due to a "de novo" mutation in GCK (Y214C). A baby girl presented with hypoglycemic seizures since the first postnatal day as well as with inappropriate hyperinsulinemia. Severe hypoglycemia persisted even after treatment with diazoxide and subtotal pancreatectomy, leading to irreversible brain damage. Pancreatic histology revealed abnormally large and hyperfunctional islets. The mutation is located in the putative allosteric activator domain of the protein. Functional studies of purified recombinant glutathionyl S transferase fusion protein of GK-Y214C showed a sixfold increase in its affinity for glucose, a lowered cooperativity, and increased kcat. The relative activity index of GK-Y214C was 130, and the threshold for GSIS predicted by mathematical modeling was 0.8 mmol/l, compared with 5 mmol/l in the wild-type enzyme. In conclusion, we have identified a de novo GCK activating mutation that causes hyperinsulinemic hypoglycemia of exceptional severity. These findings demonstrate that the range of the clinical phenotype caused by GCK mutations varies from complete insulin deficiency to extreme hyperinsulinemia. PMID- 15277403 TI - Effects of rosiglitazone and metformin on liver fat content, hepatic insulin resistance, insulin clearance, and gene expression in adipose tissue in patients with type 2 diabetes. AB - Both rosiglitazone and metformin increase hepatic insulin sensitivity, but their mechanism of action has not been compared in humans. The objective of this study was to compare the effects of rosiglitazone and metformin treatment on liver fat content, hepatic insulin sensitivity, insulin clearance, and gene expression in adipose tissue and serum adiponectin concentrations in type 2 diabetes. A total of 20 drug-naive patients with type 2 diabetes (age 48 +/- 3 years, fasting plasma glucose 152 +/- 9 mg/dl, BMI 30.6 +/- 0.8 kg/m2) were treated in a double blind randomized fashion with either 8 mg rosiglitazone or 2 g metformin for 16 weeks. Both drugs similarly decreased HbA1c, insulin, and free fatty acid concentrations. Body weight decreased in the metformin (84 +/- 4 vs. 82 +/- 4 kg, P < 0.05) but not the rosiglitazone group. Liver fat (proton spectroscopy) was decreased with rosiglitazone by 51% (15 +/- 3 vs. 7 +/- 1%, 0 vs. 16 weeks, P = 0.003) but not by metformin (13 +/- 3 to 14 +/- 3%, NS). Rosiglitazone (16 +/- 2 vs. 20 +/- 1 ml.kg(-1).min(-1), P = 0.02) but not metformin increased insulin clearance by 20%. Hepatic insulin sensitivity in the basal state increased similarly in both groups. Insulin-stimulated glucose uptake increased significantly with rosiglitazone but not with metformin. Serum adiponectin concentrations increased by 123% with rosiglitazone but remained unchanged during metformin treatment. The decrease of serum adiponectin concentrations correlated with the decrease in liver fat (r = -0.74, P < 0.001). Rosiglitazone but not metformin significantly increased expression of peroxisome proliferator-activated receptor-gamma, adiponectin, and lipoprotein lipase in adipose tissue. In conclusion, rosiglitazone but not metformin decreases liver fat and increases insulin clearance. The decrease in liver fat by rosiglitazone is associated with an increase in serum adiponectin concentrations. Both agents increase hepatic insulin sensitivity, but only rosiglitazone increases peripheral glucose uptake. PMID- 15277404 TI - CD16+ human monocyte-derived dendritic cells matured with different and unrelated stimuli promote similar allogeneic Th2 responses: regulation by pro- and anti inflammatory cytokines. AB - We previously demonstrated that tumor necrosis factor (TNF)-alpha-matured CD16- and CD16+ human monocyte-derived dendritic cells (16-mDC and 16+mDC) differentially stimulate naive CD4+ lymphocytes by inducing Th1- and Th2-like responses, respectively. Here, we further characterized the role of different DC maturation factors on Th polarization. Immature 16+mDC and 16-mDC (iDC) obtained by culture of purified monocytes with GM-CSF and IL-4 were maturated with (i) Toll-like receptor (TLR) ligands [lipopolysaccharide (LPS)], (ii) lymphocyte derived (soluble CD40 ligand, IFN-gamma) and (iii) endogenous inflammatory stimuli [TNF-alpha, prostaglandin (PG)E2]. After activation with these stimuli, DC secrete IL-12 only in presence of LPS, and 16+mDC produced lower amounts of IL 12 and IL-10 than 16-mDC. Allogeneic CD4+CD45RO- lymphocytes co-cultured with 16+mDC secreted higher levels of IL-4 and IL-10 than those co-cultured with 16 mDC, regardless of the maturation stimuli. Results were similar when DC were activated with TLR-2 or TLR-3 ligands. The higher induction of IL-4 by 16+mDC was primarily dependent on IL-12, IL-4 and IL-10. IFN-gamma production by CD4+ T cells was similar with all the conditions except with LPS-16+mDC, which induced reduced amounts of this cytokine. Those differences were totally eliminated by neutralization of IL-12, IL-4 or IL-10. Finally, 16-mDC could reverse the Th2 phenotype of already committed lymphocytes toward a Th1 pattern in short-term cultures, whereas 16+mDC had less ability to skew this phenotype. These results indicate that 16+mDC elicit superior Th2 responses independently of the maturation factors that they received, and suggest that they could represent an important population of regulatory DC. PMID- 15277407 TI - Disruption of ultradian and circadian rhythms of blood pressure in nondipper hypertensive patients. AB - Ultradian rhythms in blood pressure (BP) are known to exist, but their modification in hypertension is largely unknown. The present study was undertaken to assess the integrity of ultradian and 24-hour BP rhythms in dipper (n=100) and nondipper (n=20) hypertensive patients compared with 44 dipper normotensive individuals. Fourier analysis was used to fit ultradian (12, 8, and 6 hour) and 24-hour rhythms in BP and heart rate (HR). Mesor, amplitude, and acrophase were calculated for individual and overall rhythm curves. All subjects showed significant ultradian or 24-hour BP and HR rhythms. Systolic and diastolic BP mesor was higher in hypertensive patients compared with normotensive patients. The percentage of variability in ambulatory BP that could be explained by fitting ultradian and 24-hour rhythms was reduced in nondippers compared with normotensives or dippers. Amplitude of ultradian and 24-hour rhythms in BP increased in dippers and decreased in nondippers. Ultradian and 24-hour rhythms in HR did not differ among the 3 groups examined. Results indicate that in nondippers, blunted ultradian and 24-hour rhythm amplitude in BP was accompanied by a loss of rhythm integrity. PMID- 15277406 TI - Renal and hormonal effects of water deprivation in late-term pregnant rats. AB - Water-retaining hormones are stimulated during pregnancy allowing normal volume expansion. Because pregnant rats actively retain water, we postulate that water deprivation (WD) would cause a greater reduction in plasma volume in pregnant than in nonpregnant rats. To test this hypothesis, Sprague-Dawley pregnant and nonpregnant rats were water-deprived for 48 hours. At day 19 of pregnancy, or in the corresponding day in nonpregnant rats, they were randomly assigned to either a WD or a control (C) pair-fed group (n=10 to 12 per group). WD significantly reduced body weight, food intake, and creatinine clearance, and increased urinary osmolality in nonpregnant and pregnant rats. WD reduced plasma volume in a similar proportion in nonpregnant and pregnant rats (nonpregnant rats C=13.1+/ 0.4, WD=11.0+/-0.2; pregnant rats C=19.4+/-0.7, WD=16.8+/-0.5 mL, P<0.001). Both groups of pregnant rats had a similar reduction in blood pressure. Plasma renin activity (nonpregnant rats C=6.1+/-1.1, WD=20.5+/-2.0; pregnant rats C=49+/-9.7, WD=94+/-12 ng angiotensin I/mL per hour, P<0.001) and plasma aldosterone levels were increased by pregnancy and further increased by WD. WD significantly reduced urinary kallikrein. WD caused a significant reduction in fetal but not placental weights. Present data indicate that 48-hour WD reduced renal kallikrein and further stimulated water-retaining hormones. We speculate that these are compensatory changes contributing to the maintenance of pregnancy in response to WD. PMID- 15277408 TI - Circulating 1,5-anhydroglucitol levels in adult patients with diabetes reflect longitudinal changes of glycemia: a U.S. trial of the GlycoMark assay. AB - OBJECTIVE: 1,5-Anhydroglucitol (1,5AG) is a major circulating polyol arising primarily from ingestion and excreted competitively with glucose. Japanese studies have demonstrated reduced concentrations of 1,5AG in serum in hyperglycemic patients in comparison with euglycemic subjects and a gradual normalization of 1,5AG values for patients responding to antihyperglycemic therapies. In this first U.S. study, we assessed the ability of 1,5AG measurements to monitor glycemic control in a cohort of 77 patients with diabetes (22 with type 1 diabetes, 55 with type 2 diabetes) who presented with suboptimal glycemic control at baseline (defined as HbA(1c) >or=7%). RESEARCH DESIGN AND METHODS: Each patient received therapies consisting of combinations of diabetes education, nutritional counseling, and addition or dose adjustment of various insulins or oral antihyperglycemic medications. Therapy was targeted to reduce mean HbA(1c) by >or=1.0% over the monitoring period. 1,5AG, HbA(1c), fructosamine, and random glucose measurements were performed at baseline and at 2, 4, and 8 weeks after the initiation of therapy. RESULTS: 1,5AG, fructosamine, and glucose values progressed significantly toward euglycemia by week 2 of monitoring (Wilcoxon's signed-rank test, P < 0.05), with median changes of 93, 7, and -13% for 1,5AG, fructosamine, and glucose, respectively. In contrast, HbA(1c) values did not respond significantly to therapy until week 4. On an individual patient basis, 89.6% of patients displayed longitudinal changes of 1,5AG from baseline to week 8 in concordance with HbA(1c). 1,5AG was also highly correlated with HbA(1c) and fructosamine (Spearman rho = -0.6459 and -0.6751, respectively; both P < 0.0001). CONCLUSIONS: We conclude that 1,5AG responds sensitively and rapidly to changes in glycemia and monitors glycemic control in accordance with established markers. PMID- 15277409 TI - Improved plasma glucose control, whole-body glucose utilization, and lipid profile on a low-glycemic index diet in type 2 diabetic men: a randomized controlled trial. AB - OBJECTIVE: To determine whether a chronic low-glycemic index (LGI) diet, compared with a high-glycemic index (HGI) diet, has beneficial effects on plasma glucose control, lipid metabolism, total fat mass, and insulin resistance in type 2 diabetic patients. RESEARCH DESIGN AND METHODS: Twelve type 2 diabetic men were randomly allocated to two periods of 4 weeks of an LGI or HGI carbohydrate diet separated by a 4-week washout interval, in a crossover design. RESULTS: The LGI diet induced lower postprandial plasma glucose and insulin profiles and areas under the curve than after the HGI diet. At the end of the two dietary periods, the 7-day dietary records demonstrated equal daily total energy and macronutrient intake. Body weight and total fat mass were comparable. Four-week LGI versus HGI diet induced improvement of fasting plasma glucose (P < 0.01, Delta changes during LGI vs. HGI), HbA(1c) (P < 0.01), and whole-body glucose utilization measured by the euglycemic-hyperinsulinemic clamp (P < 0.05). LGI diet induced a decrease in fasting plasma total and LDL cholesterol (Delta changes LGI vs. HGI, P < 0.01), free fatty acids (P < 0.01), apolipoprotein B, and plasminogen activator inhibitor 1 activity. CONCLUSIONS: Only 4 weeks of an LGI diet was able to improve glycemic control, glucose utilization, some lipid profiles, and the capacity for fibrinolysis in type 2 diabetes. Even if changes in glycemic control were modest during the 4-week period, the use of an LGI diet in a longer-term manner might play an important role in the treatment and prevention of diabetes and related disorders. PMID- 15277410 TI - Treatment of diabetic ketoacidosis with subcutaneous insulin aspart. AB - OBJECTIVE: In this prospective, randomized, open trial, we compared the efficacy and safety of aspart insulin given subcutaneously at different time intervals to a standard low-dose intravenous (IV) infusion protocol of regular insulin in patients with uncomplicated diabetic ketoacidosis (DKA). RESEARCH DESIGN AND METHODS: A total of 45 consecutive patients admitted with DKA were randomly assigned to receive subcutaneous (SC) aspart insulin every hour (SC-1h, n = 15) or every 2 h (SC-2h, n = 15) or to receive IV infusion of regular insulin (n = 15). Response to medical therapy was evaluated by assessing the duration of treatment until resolution of hyperglycemia and ketoacidosis. Additional end points included total length of hospitalization, amount of insulin administration until resolution of hyperglycemia and ketoacidosis, and number of hypoglycemic events. RESULTS: Admission biochemical parameters in patients treated with SC-1h (glucose: 44 +/- 21 mmol/l [means +/- SD], bicarbonate: 7.1 +/- 3 mmol/l, pH: 7.14 +/- 0.09) were similar to those treated with SC-2h (glucose: 42 +/- 21 mmol/l, bicarbonate: 7.6 +/- 4 mmol/l, pH: 7.15 +/- 0.12) and IV regular insulin (glucose: 40 +/- 13 mmol/l, bicarbonate 7.1 +/- 4 mmol/l, pH: 7.11 +/- 0.17). There were no statistical differences in the mean duration of treatment until correction of hyperglycemia (6.9 +/- 4, 6.1 +/- 4, and 7.1 +/- 5 h) or until resolution of ketoacidosis (10 +/- 3, 10.7 +/- 3, and 11 +/- 3 h) among patients treated with SC-1h and SC-2h or with IV insulin, respectively (NS). There was no mortality and no differences in the length of hospital stay, total amount of insulin administration until resolution of hyperglycemia or ketoacidosis, or the number of hypoglycemic events among treatment groups. CONCLUSIONS: Our results indicate that the use of subcutaneous insulin aspart every 1 or 2 h represents a safe and effective alternative to the use of intravenous regular insulin in the management of patients with uncomplicated DKA. PMID- 15277411 TI - The incidence of congestive heart failure in type 2 diabetes: an update. AB - OBJECTIVE: The aims of this study were to update previous estimates of the congestive heart failure (CHF) incidence rate in patients with type 2 diabetes, compare it with an age- and sex-matched nondiabetic group, and describe risk factors for developing CHF in diabetic patients over 6 years of follow-up. RESEARCH DESIGN AND METHODS: We performed a retrospective cohort study of 8,231 patients with type 2 diabetes and 8,845 nondiabetic patients of similar age and sex who did not have CHF as of 1 January 1997, following them for up to 72 months to estimate the CHF incidence rate. In the diabetic cohort, we constructed a Cox regression model to identify risk factors for CHF development. RESULTS: Patients with diabetes were much more likely to develop CHF than patients without diabetes (incidence rate 30.9 vs. 12.4 cases per 1,000 person-years, rate ratio 2.5, 95% CI 2.3-2.7). The difference in CHF development rates between persons with and without diabetes was much greater in younger age-groups. In addition to age and ischemic heart disease, poorer glycemic control (hazard ratio 1.32 per percentage point of HbA(1c)) and greater BMI (1.12 per 2.5 units of BMI) were important predictors of CHF development. CONCLUSIONS: The CHF incidence rate in type 2 diabetes may be much greater than previously believed. Our multivariate results emphasize the importance of controlling modifiable risk factors for CHF, namely hyperglycemia, elevated blood pressure, and obesity. Younger patients may benefit most from risk factor modification. PMID- 15277412 TI - Incidence of lower-extremity amputation in American Indians: the Strong Heart Study. AB - OBJECTIVE: To define incidence and predictors of nontraumatic lower-extremity amputation (LEA) in a diverse cohort of American Indians with diabetes. RESEARCH DESIGN AND METHODS: The Strong Heart Study is a study of cardiovascular disease and its risk factors in 13 American-Indian communities. Data on the presence/absence of amputations were collected at each of three serial examinations (1989-1992, 1993-1995, and 1997-1999) by direct examination of the lower extremity. The logistic regression model was used to quantify the relationship between risk of LEA and potential risk factors, including diabetes duration, HbA(1c), peripheral arterial disease, and renal function. RESULTS: Of the 1,974 individuals with diabetes and without prevalent LEA at baseline, 87 (4.4%) experienced an LEA during 8 years of follow-up, and a total of 157 anatomical sites were amputated among these individuals. Amputation of toes was most common, followed by below-the-knee and above-the-knee amputations. Age adjusted odds of LEA were higher among individuals with unfavorable combinations of risk factors, such as albuminuria and elevated HbA(1c). Multivariable modeling indicated that male sex, renal dysfunction, high ankle-brachial index, longer duration of diabetes, less than a high school education, increasing systolic blood pressure, and HbA(1c) predicted LEA risk. CONCLUSIONS: The 8-year cumulative incidence of LEA in American Indians with diabetes is 4.4%, with marked differences in risk by sex, educational attainment, renal function, and glycemic control. PMID- 15277413 TI - Are we underestimating diabetes-related lower-extremity amputation rates? Results and benefits of the first prospective study. AB - OBJECTIVE: The objective of this study was to accurately determine the incidence of lower-extremity amputation using prospective data collection and to compare the results with those obtained by retrospective methods. RESEARCH DESIGN AND METHODS: The study was carried out over a 3-year period in a large district general hospital covering a clearly defined and relatively static population. All diabetic inpatients with foot problems were identified and followed-up until discharge or death. The demographic and admission details, medical history, investigations, procedures, and history and etiology of the foot lesion were collected twice weekly by a specialist nurse and podiatrist from all relevant wards. Thus, all subjects who underwent amputation could be identified. For comparison, retrospective data were collected from the hospital coding activities database, operating theater log books, anesthetic database, and limb-fitting records. RESULTS: The total population of the region in 2000 was 337,859, of which 9,183 were known to have diabetes. The total number of amputations during the 3-year survey period was 79, of which 45 were major and 34 minor. In our local population, the mean incidence during the survey period (1997-2000) equates to 7.8/100,000 general population and 2.85/1,000 diabetic population for all amputations, 4.5/100,000 general population and 1.62/1,000 diabetic population for major amputations, and 3.3/100,000 general population and 1.23/1,000 diabetic population for minor amputations. The prospective survey detected all lower extremity amputations identified by the various retrospective methods; however, for the reverse, this was not the case. All of the retrospective methods, including the most commonly used (ICD-9 and OPCS-4 coding), failed to detect all of the cases revealed by the prospective survey (error rate ranging from 4.2 to 90.6%), and between 4.5 and 17.4% of amputations were misclassified. CONCLUSIONS: This study demonstrates the advantages of prospective data collection as a means of determining the incidence of lower-extremity amputations and highlights the limitations of retrospective data collection methods, which underestimate the incidence. In particular, the operating theater records, which have been the gold standard for many surveys, were found to be unreliable. Moreover, we have shown a 47% reduction in the major amputations during the survey period. Thus, we recommend that a prospective audit be incorporated into the activities of the specialist foot care team as a means of assessing and improving clinical care. PMID- 15277414 TI - Cost-effectiveness of early irbesartan treatment versus control (standard antihypertensive medications excluding ACE inhibitors, other angiotensin-2 receptor antagonists, and dihydropyridine calcium channel blockers) or late irbesartan treatment in patients with type 2 diabetes, hypertension, and renal disease. AB - OBJECTIVE: The aim of this study was to determine the most cost-effective time point for initiation of irbesartan treatment in hypertensive patients with type 2 diabetes and renal disease. RESEARCH DESIGN AND METHODS: This study was a Markov model-simulated progression from microalbuminuria to overt nephropathy, doubling of serum creatinine, end-stage renal disease, and death in hypertensive patients with type 2 diabetes. Two irbesartan strategies were created: early irbesartan 300 mg daily (initiated with microalbuminuria) and late irbesartan (initiated with overt nephropathy). These strategies were compared with control, which consisted of antihypertensive therapy with standard medications (excluding ACE inhibitors, other angiotensin-2 receptor antagonists, and dihydropyridine calcium channel blockers) with comparable blood pressure control, initiated at microalbuminuria. Transition probabilities were taken from the Irbesartan in Reduction of Microalbuminuria-2 study, Irbesartan in Diabetic Nephropathy Trial, and other published sources. Costs and life expectancy, discounted at 3% yearly, were projected over 25 years for 1,000 simulated patients using a third-party payer perspective in a U.S. setting. RESULTS: Compared with control, early and late irbesartan treatment in 1,000 patients were projected to save (mean +/- SD) 11.9 +/- 3.3 million dollars and 3.3 +/- 2.7 million dollars, respectively. Early use of irbesartan added 1,550 +/- 270 undiscounted life-years (discounted 960 +/- 180), whereas late irbesartan added 71 +/- 40 life-years (discounted 48 +/- 27) in 1,000 patients. Early irbesartan treatment was superior under a wide-range of plausible assumptions. CONCLUSIONS: Early irbesartan treatment was projected to improve life expectancy and reduce costs in hypertensive patients with type 2 diabetes and microalbuminuria. Later use of irbesartan in overt nephropathy is also superior to standard care, but irbesartan should be started earlier and continued long term. PMID- 15277415 TI - Excess hospitalizations, hospital days, and inpatient costs among people with diabetes in Andalusia, Spain. AB - OBJECTIVE: The goal of this study was to estimate the excess hospitalizations, hospital days, and inpatient costs attributable to diabetes in Andalusia, Spain (37 hospitals, 7,236,459 inhabitants), during 1999 compared with those without diabetes. RESEARCH DESIGN AND METHODS: This study was an analysis of all hospital discharges. Those with an ICD-9-CM code of 250 as either the main or secondary diagnosis were considered to have been admissions of individuals with diabetes. An estimate of costs was applied to each inpatient admission by assigning a cost weight based on the diagnostic-related group (DRG) related to each admission. RESULTS: A total of 538,580 admissions generated 4,310,654 hospital bed-days and total costs of 940,026,949 euro. People with diabetes accounted for 9.7% of all hospital discharges, 13.8% of total stays, and 14.1% of the total cost. Of the total cost for individuals with diabetes (132,509,217 euro), 58.3% were excess costs, of which 47% was attributable to cardiovascular complications and 43% to admissions for comorbid diseases. Individuals 45-75 years of age accounted for 75% of the excess costs. The rate of admissions during the study year was 145 per 1,000 inhabitants for individuals with diabetes compared with 70 admissions per 1,000 inhabitants for individuals without diabetes. CONCLUSIONS: The costs arising from hospitalization of individuals with diabetes are disproportionate in relation to their prevalence. For those aged >or=45 years, cardiovascular complications were clearly the most important factor determining increased costs from diabetes. PMID- 15277416 TI - Additive effects of glucagon-like peptide 1 and pioglitazone in patients with type 2 diabetes. AB - OBJECTIVE: To evaluate the effect of combination therapy with pioglitazone and glucagon-like peptide (GLP)-1 in patients with type 2 diabetes. RESEARCH DESIGN AND METHODS: Eight patients with type 2 diabetes (BMI 32.7 +/- 1.3 kg/m(2) and fasting plasma glucose 13.5 +/- 1.2 mmol/l) underwent four different treatment regimens in random order: saline therapy, monotherapy with continuous subcutaneous infusion of GLP-1 (4.8 pmol x kg(-1) x min(-1)), monotherapy with pioglitazone (30-mg tablet of Actos), and combination therapy with GLP-1 and pioglitazone. The observation period was 48 h. End points were plasma levels of glucose, insulin, glucagon, free fatty acids (FFAs), and sensation of appetite. RESULTS: Fasting plasma glucose decreased from 13.5 +/- 1.2 mmol/l (saline) to 11.7 +/- 1.2 (GLP-1) and 11.5 +/- 1.2 (pioglitazone) and further decreased to 9.9 +/- 1.0 (combination) (P < 0.001). Eight-hour mean plasma glucose levels were reduced from 13.7 +/- 1.1 mmol/l (saline) to 10.6 +/- 1.0 (GLP-1) and 12.0 +/- 1.2 (pioglitazone) and were further reduced to 9.5 +/- 0.8 (combination) (P < 0.0001). Insulin levels increased during monotherapy with GLP-1 compared with monotherapy with pioglitazone (P < 0.01). Glucagon levels were reduced in GLP-1 and combination therapy compared with saline and monotherapy with pioglitazone (P < 0.01). FFAs during breakfast (area under the curve, 0-3 h) were reduced in combination therapy compared with saline (P = 0.03). Sensation of appetite was reduced during monotherapy with GLP-1 and combination therapy (P < 0.05). CONCLUSIONS: GLP-1 and pioglitazone show an additive glucose-lowering effect. A combination of the two agents may, therefore, be a valuable therapeutic approach for the treatment of type 2 diabetes. PMID- 15277417 TI - The effect of liraglutide, a long-acting glucagon-like peptide 1 derivative, on glycemic control, body composition, and 24-h energy expenditure in patients with type 2 diabetes. AB - OBJECTIVE: Glucagon-like peptide (GLP)-1 is a gut hormone that exerts incretin effects and suppresses food intake in humans, but its therapeutic use is limited due to its short half-life. This was a randomized, double-blind, parallel-group, placebo-controlled trial investigating the effect of the long-acting GLP-1 derivative liraglutide (NN2211) on glycemic control, body weight, body composition, and 24-h energy expenditure in obese subjects with type 2 diabetes. RESEARCH DESIGN AND METHODS: Thirty-three patients (mean +/- SD) aged 60.0 +/- 9.5 years, with HbA(1c) 7.5 +/- 1.2% and BMI 36.6 +/- 4.1 kg/m(2), were randomized to treatment with a single daily subcutaneous dose of 0.6 mg liraglutide (n = 21) or placebo (n = 12) for 8 weeks. In addition to weight and glycemic parameters, body composition was assessed by dual-energy X-ray absorptiometry (DEXA) scanning and 24-h energy expenditure in a respiratory chamber. RESULTS: After 8 weeks, liraglutide reduced fasting serum glucose (liraglutide, -1.90 mmol/l, and placebo, 0.27 mmol/l; P = 0.002) and HbA(1c) (liraglutide, -0.33%, and placebo, 0.47%; P = 0.028) compared with placebo. No change in body weight was detected (liraglutide, -0.7 kg, and placebo, -0.9 kg; P = 0.756). There was a nonsignificant trend toward a decrease in total fat mass (liraglutide, -0.98%, and placebo, -0.12%; P = 0.088) and toward an increase in lean body mass (liraglutide, 1.02%, and placebo, 0.23%; P = 0.118) in the liraglutide group compared with the placebo group. Twenty-four-hour energy expenditure was unaffected by the treatment (liraglutide, -12.6 kJ/h, and placebo, -13.7 kJ/h; P = 0.799). CONCLUSIONS: Eight weeks of 0.6-mg liraglutide treatment significantly improved glycemic control without increasing weight in subjects with type 2 diabetes compared with those on placebo. No influence on 24 h energy expenditure was detected. PMID- 15277418 TI - Evaluating the accuracy of continuous glucose-monitoring sensors: continuous glucose-error grid analysis illustrated by TheraSense Freestyle Navigator data. AB - OBJECTIVE: The objective of this study was to introduce continuous glucose-error grid analysis (CG-EGA) as a method of evaluating the accuracy of continuous glucose-monitoring sensors in terms of both accurate blood glucose (BG) values and accurate direction and rate of BG fluctuations and to illustrate the application of CG-EGA with data from the TheraSense Freestyle Navigator. RESEARCH DESIGN AND METHODS: We approach the design of CG-EGA from the understanding that continuous glucose sensors (CGSs) allow the observation of BG fluctuations as a process in time. We account for specifics of process characterization (location, speed, and direction) and for biological limitations of the observed processes (time lags associated with interstitial sensors). CG-EGA includes two interacting components: 1) point-error grid analysis (P-EGA) evaluates the sensor's accuracy in terms of correct presentation of BG values and 2) rate-error grid analysis (R EGA) assesses the sensor's ability to capture the direction and rate of BG fluctuations. RESULTS: CG-EGA revealed that the accuracy of the Navigator, measured as a percentage of accurate readings plus benign errors, was significantly different at hypoglycemia (73.5%), euglycemia (99%), and hyperglycemia (95.4%). Failure to detect hypoglycemia was the most common error. The point accuracy of the Navigator was relatively stable over a wide range of BG rates of change, and its rate accuracy decreased significantly at high BG levels. CONCLUSIONS: Traditional self-monitoring of BG device evaluation methods fail to capture the important temporal characteristics of the continuous glucose monitoring process. CG-EGA addresses this problem, thus providing a comprehensive assessment of sensor accuracy that appears to be a useful adjunct to other CGS performance measures. PMID- 15277419 TI - Elevated plasma levels of Nt-proBNP in patients with type 2 diabetes without overt cardiovascular disease. AB - OBJECTIVE: The NH(2)-terminal portion of the precursor of brain natriuretic peptide (Nt-proBNP) has been reported to be elevated in left ventricular dysfunction. This peptide is a split product from the proBNP molecule, and its level in the circulation is not, as the mature BNP peptide, dependent on the peripheral number of BNP receptors. We aimed to test the hypothesis that asymptomatic left ventricular dysfunction (ALVD), as estimated by Nt-proBNP, would be more prevalent in patients with type 2 diabetes without overt cardiovascular disease in comparison with matched control subjects. RESEARCH DESIGN AND METHODS: The study population consisted of 253 patients with type 2 diabetes and 230 matched control subjects aged 40-70 years without any overt heart disease from primary care centers in Western Finland and Southern Sweden. Nt-proBNP was measured in plasma by competitive enzyme immunosorbent assay. RESULTS: Patients with type 2 diabetes were shown to have higher Nt-proBNP values (360.9 pmol/l [262.6-467.9]) than control subjects (302.7 pmol/l [215.4-419.2]) (P < 0.001). Nt-proBNP levels were independently related to diabetes after adjustment for age, sex, systolic and diastolic blood pressure, BMI, heart rate, drug treatment, serum creatinine, and cystatin C. CONCLUSIONS: Our data suggest that the secretion of Nt-proBNP is increased in type 2 diabetic patients with no overt heart disease, suggesting that type 2 diabetes is associated with a higher prevalence of ALVD than hitherto thought. Nt-proBNP may thus serve as a screening instrument to select patients with type 2 diabetes who could benefit from an echocardiographical examination. PMID- 15277420 TI - Distinct diagnostic criteria of fulminant type 1 diabetes based on serum C peptide response and HbA1c levels at onset. AB - OBJECTIVE: Diagnostic criteria in fulminant type 1 diabetes, a novel subtype of type 1 diabetes, remain unclear. RESEARCH DESIGN AND METHODS: We analyzed basal and longitudinal changes of serum C-peptide levels during a 75-g oral glucose tolerance test (OGTT) in 125 consecutively recruited patients with type 1 diabetes including fulminant type 1 diabetes (n = 25) and acute-onset type 1 diabetes (n = 100). Discriminating criteria of fulminant type 1 diabetes were examined using receiver-operating characteristic curve analysis and multiple logistic regression analysis. RESULTS: The integrated values of serum C-peptide response during OGTT (SigmaC-peptide) in fulminant type 1 diabetes at onset, 1 year, and 2 years after onset were markedly lower than those in acute-onset type 1 diabetes. None of the patients with fulminant type 1 diabetes had improvement of C-peptide response to OGTT. Fasting C-peptide values at onset in fulminant type 1 diabetes were significantly lower than those in acute-onset type 1 diabetes. We established diagnostic criteria of serum C-peptide and HbA(1c) levels at onset that discriminate fulminant type 1 diabetes from acute-onset type 1 diabetes with high sensitivity and specificity: a criterion in which the levels of both the fasting C-peptide is 5 years' duration. MDL showed a linear decrease with increasing duration of diabetes. Distal leg IENF showed significant negative correlations with warm (P < 0.02) and cold (P < 0.05) thermal threshold, heat pain (P < 0.05), pressure sense (P < 0.05), and neurological disability score total sensory (P < 0.03) and total neuropathy (P < 0.03) values. CONCLUSIONS: IENF was not significantly altered in these patients at <5 years' duration of diabetes, but fell significantly after 5 years of diabetes. MDL exhibited a linear loss with time, suggesting a different mechanism of change. MDL and IENF together may prove a useful end point in therapeutic trials for neuropathy. PMID- 15277427 TI - Variability in activity may precede diabetic foot ulceration. AB - OBJECTIVE: To evaluate the role of activity in the development of neuropathic foot ulceration in individuals with diabetes. RESEARCH DESIGN AND METHODS: We evaluated the first 100 consecutive individuals with diabetes (95.0% male, aged 68.5 +/- 10.0 years with concomitant neuropathy, deformity, and/or a history of lower-extremity ulceration/partial foot amputation) enrolled in an ongoing prospective longitudinal activity study. Subjects used a high-capacity continuous computerized activity monitor. Data were collected continuously over a minimum of 25 weeks (or until ulceration) with daily activity units expressed as means +/- SD. RESULTS: Eight subjects ulcerated during the evaluation period of 37.1 +/- 12.3 weeks. The average daily activity was significantly lower in individuals who ulcerated compared with individuals who did not ulcerate (809.0 +/- 612.2 vs. 1,394.5 +/- 868.5, P = 0.03). Furthermore, there was a large difference in variability between groups. The coefficient of variation was significantly greater in the ulceration group compared with the no ulceration group (96.4 +/- 50.3 vs. 44.7 +/- 15.4%, P = 0.0001). In the 2 weeks preceding the ulcerative event, the coefficient of variation increased even further (115.4 +/- 43.0%, P = 0.02), but there was no significant difference in average daily activity during that period (P = 0.5). CONCLUSIONS: The results of this study suggest that individuals with diabetes who develop ulceration may actually have a lower overall activity than their counterparts with no ulceration, but the quality of that activity may be more variable. Perhaps modulating the "peaks and valleys" of activity in this population through some form of feedback might prove to reduce risk for ulceration in this very-high-risk population. PMID- 15277428 TI - Male sex increases the risk of autoimmunity but not type 1 diabetes. AB - OBJECTIVE: The goal of this study was to explore the role of sex on the prevalence of autoantibodies, protective genetic subtypes, beta-cell function, and the incidence of type 1 diabetes in a population of first- and second-degree relatives of patients with type 1 diabetes (probands). We examined both the effect of the sex of the individual screened as well as the effect of the sex of the individual's proband on diabetes risk variables tested. RESEARCH DESIGN AND METHODS: The Diabetes Prevention Trial-Type 1 has screened 93,188 relatives of type 1 diabetic patients from February 1994 to January 2002. After observing that more men than women were islet cell autoantibody (ICA) positive for the group as a whole, we further explored the role of sex by detailed analysis of variables in this population. RESULTS: Our data suggest only an influence of sex on the type 1 diabetes disease process. After adjustment for race, age, and relationship to proband, male sex was associated with the appearance of autoimmunity, i.e., the presence of ICA and having two or more antibodies. There was no effect of sex on the presence of other autoantibodies, insulin secretion, results of oral glucose tolerance test, or development of diabetes. CONCLUSIONS: Our finding that male sex conveys an independent increased risk for development of ICA and multiple antibodies, while at the same time finding no difference with respect to the development of diabetes, suggests that male relatives with the known risk factor of ICA are less likely than comparable female relatives to progress to overt disease, that the pathogenesis of type 1 diabetes among men is slower compared with women, or that women develop diabetes manifesting different antibody responses. PMID- 15277429 TI - Non-HDL cholesterol and apolipoprotein B predict cardiovascular disease events among men with type 2 diabetes. AB - OBJECTIVE: To evaluate the role of non-HDL cholesterol and apolipoprotein (apo)B, markers of all potentially atherogenic lipoproteins, as predictors of cardiovascular disease (CVD) in comparison with LDL cholesterol in patients with type 2 diabetes. RESEARCH DESIGN AND METHODS: We prospectively followed 746 diabetic men in the Health Professionals' Follow-up Study who were aged 46-81 years and free of CVD or cancer at the time of blood draw in 1993-1994. During 6 years of follow-up, we ascertained 103 incident CVD cases. RESULTS: We used Cox proportional hazard modeling to estimate the relative risk (RR) of CVD. After adjustment for age, BMI, and other lifestyle risk factors, the multivariate RR of CVD (the highest versus the lowest quartile) was 2.34 (95% CI 1.26-4.32) for non HDL cholesterol, 2.31 (1.23-4.35) for apoB, and 1.74 (0.99-3.06) for LDL cholesterol. Comparisons of nested models indicate that non-HDL cholesterol, but not apoB, adds significantly to the prediction of CVD risk beyond LDL cholesterol. The area under the receiver operating characteristic curve was 0.685, 0.691, 0.695, and 0.722 for the CVD risk-prediction model with LDL cholesterol, apoB, non-HDL cholesterol, and total cholesterol-to-HDL cholesterol ratio (or the non-HDL-to-HDL cholesterol ratio), respectively. CONCLUSIONS: Non HDL cholesterol and apoB are more potent predictors of CVD incidence among diabetic men than LDL cholesterol. Statistically, the ratio of total to HDL cholesterol is the best predictor of CVD in this cohort of diabetic men. PMID- 15277430 TI - Impact of degree of obesity on surrogate estimates of insulin resistance. AB - OBJECTIVE: To evaluate the role of adiposity in the relationship between specific and surrogate estimates of insulin-mediated glucose uptake (IMGU) in a large nondiabetic population. RESEARCH DESIGN AND METHODS: Healthy volunteers were classified by BMI into normal weight (<25.0 kg/m(2), n = 208), overweight (25.0 29.9 kg/m(2), n = 168), and obese (>or=30.0 kg/m(2), n = 109) groups. We then assessed how differences in BMI affect the correlation between steady-state plasma glucose (SSPG) concentration at the end of a 180-min infusion of octreotide, glucose, and insulin (a specific measure of IMGU) and five surrogate estimates: fasting plasma glucose, fasting plasma insulin, homeostasis model assessment of insulin resistance (HOMA-IR), quantitative insulin sensitivity check index (QUICKI), and area under the curve for insulin in response to oral glucose (I-AUC). RESULTS: Correlation coefficients (r values) between SSPG and surrogate measures of IMGU were all significant (P < 0.05), but the magnitude varied between BMI groups: normal weight: fasting plasma glucose 0.20, fasting plasma insulin 0.33, HOMA-IR 0.36, QUICKI -0.33, and I-AUC 0.69; overweight: fasting plasma glucose 0.19, fasting plasma insulin 0.55, HOMA-IR 0.55, QUICKI 0.54, and I-AUC 0.72; and obese: fasting plasma glucose 0.40, fasting plasma insulin 0.56, HOMA-IR 0.60, QUICKI -0.61, and I-AUC 0.69. CONCLUSIONS: The relationship between direct and surrogate estimates of IMGU varies with BMI, with the weakest correlations seen in the normal-weight group and the strongest in the obese group. In general, I-AUC is the most useful surrogate estimate of IMGU in all weight groups. Fasting plasma insulin, HOMA-IR, and QUICKI provide comparable information about IMGU. Surrogate estimates of IMGU based on fasting insulin and glucose account for no more than 13% of the variability in insulin action in the normal-weight group, 30% in the overweight group, and 37% in the obese group. PMID- 15277431 TI - Phenotype-genotype correlations in a series of wolfram syndrome families. AB - OBJECTIVE: Wolfram syndrome is an extremely rare autosomal-recessive disorder that predisposes the development of type 1 diabetes in association with progressive optic atrophy. The genetic basis of this disease has been shown to be due to mutations in the WFS1 gene. The WFS1 gene encodes a novel transmembrane protein called wolframin, which recent evidence suggests may serve as a novel endoplasmic reticulum calcium channel in pancreatic beta-cells and neurons. Genotype-phenotype correlations in this syndrome are becoming apparent and may help in explaining some of the variable characteristics observed in this disease. RESEARCH DESIGN AND METHODS: In this report, we have studied 13 patients with Wolfram syndrome from nine families to further define the relationship between mutation site and type with specific disease characteristics. RESULTS: A severe phenotype was seen in patients with mutations in exon 4 and with a large deletion encompassing most of exon 8. In total, nine novel mutations were identified as well as three new silent polymorphisms. CONCLUSIONS: Similar to all other mutation reports, most causative changes identified in the WFS1 gene occurred in exon 8, and only one was identified outside this region in exon 4. PMID- 15277432 TI - Adiponectin and leptin concentrations may aid in discriminating disease forms in children and adolescents with type 1 and type 2 diabetes. AB - OBJECTIVE: The incidence of pediatric type 2 diabetes has recently seen an alarming increase. To improve our understanding of pediatric type 2 diabetes and identify markers that discriminate these subjects from those with type 1 diabetes, we performed a multivariant analysis associating serum adiponectin and leptin levels with anthropometrical parameters and disease state. RESEARCH DESIGN AND METHODS: Samples from children and adolescents with type 1 diabetes (n = 41) and type 2 diabetes (n = 17) and from nondiabetic individuals of similar age from the general population (n = 43) were investigated. An analysis included the parameters of matching for BMI and Tanner stage. Receiver-operator characteristic (ROC) curves were established to assess these analytes' association with disease. RESULTS: Contrary to studies of adult type 1 diabetes, adiponectin levels in our pediatric type 1 diabetic subjects (10.2 microg/ml [95% CI 8.6-11.7]) did not differ from those of healthy control subjects (10.6 microg/ml [9.2-12.0]; P = NS). Children with type 2 diabetes (5.5 microg/ml [4.8-6.2]) had significantly lower adiponectin levels than both of those groups. Conversely, type 2 diabetic subjects showed marked elevations in serum leptin concentrations (24.3 ng/ml [17.1-31.5]) compared with healthy control subjects (2.7 ng/ml [1.3-4.1]; P < 0.001) and type 1 diabetic subjects (5.1 ng/ml [3.5-6.7]; P < 0.001). Importantly, each of the properties ascribed to pediatric type 2 diabetes was present when the comparison was restricted to healthy children or type 1 diabetic patients whose BMI was >85th percentile or who had Tanner stage 4 and 5. The evaluation of adiponectin-to-leptin ratios revealed a striking difference between children with type 1 diabetes (6.3 [3.8-8.8]) and type 2 diabetes (0.3 [0.2-0.5]; P < 0.001). CONCLUSIONS: In pediatric diabetes, where diagnosis of disease is often difficult, these studies suggest that the adiponectin-to-leptin ratio may provide additional help in the discrimination between type 1 and type 2 diabetes. PMID- 15277433 TI - The +276 G/T single nucleotide polymorphism of the adiponectin gene is associated with coronary artery disease in type 2 diabetic patients. AB - OBJECTIVE: Two single nucleotide polymorphisms (SNPs) at the adiponectin locus (+45T>G and +276G>T) have been associated with low circulating adiponectin levels, insulin resistance, and type 2 diabetes. We investigated whether these genetic markers are determinants of coronary artery disease (CAD) in type 2 diabetic patients. RESEARCH DESIGN AND METHODS: A total of 376 consecutive type 2 diabetic patients were studied: 142 case subjects with coronary stenosis >50% or previous myocardial infarction and 234 control subjects with no symptoms, no electrocardiogram (ECG) signs of myocardial ischemia, and a normal ECG stress test (n = 189) and/or (n = 45) with coronary stenosis T polymorphism is a determinant of CAD risk in type 2 diabetic patients. This marker may assist in the identification of diabetic individuals at especially high risk of CAD, so that preventive programs can be targeted at these subjects. PMID- 15277434 TI - Aldose reductase gene polymorphisms and peripheral nerve function in patients with type 2 diabetes. AB - OBJECTIVE: We screened the human aldose reductase (ALR) gene for DNA sequence variants in type 2 diabetic and nondiabetic subjects and investigated whether the previously reported and novel polymorphisms were associated with neurophysiologic deterioration and clinical peripheral neuropathy. RESEARCH DESIGN AND METHODS: The study population included 85 Finnish type 2 diabetic and 126 nondiabetic subjects. The genetic analyses were performed using the PCR, single-strand conformation polymorphism, restriction fragment-length polymorphism, and automated laser fluorescence scanning analyses. A detailed neurologic examination and neurophysiologic analyses were performed at the time of diagnosis and at the 10-year examination. RESULTS: The genetic screening identified four polymorphisms: C-106T, C-11G, A11370G, and C19739A. The C and Z-2 alleles of the C-106T polymorphism and the previously reported (CA)(n) repeat marker were more frequent in type 2 diabetic subjects than in nondiabetic subjects. At baseline, the diabetic subjects with the T allele of the C-106T polymorphism had lower sensory response amplitude values in the peroneal (P = 0.025), sural (P = 0.007), and radial (P = 0.057) nerves and, during follow-up, a greater decrease in the conduction velocity of the motor peroneal nerve than those with the C-106C genotype. No associations were found between the polymorphisms examined and clinical polyneuropathy. CONCLUSIONS: The C-106T polymorphism of the ALR gene may contribute to an early development of neurophysiologic deterioration in type 2 diabetic patients. PMID- 15277435 TI - Prevalence and factor analysis of metabolic syndrome in an urban Korean population. AB - OBJECTIVE: The aim of this study was to explore the prevalence and pattern of the metabolic syndrome and its association with hyperinsulinemia in an urban Korean population of 269 men and 505 women. RESEARCH DESIGN AND METHODS: The National Cholesterol Education Program Adult Treatment Panel (ATP) III guidelines were used to calculate the sex-specific prevalence of the metabolic syndrome. After excluding individuals taking medication for hypertension, diabetes, or dyslipidemia, we used factor analysis to examine the pattern of the metabolic syndrome in 206 men and 449 women. RESULTS: The prevalence of metabolic syndrome was 16.0% in men and 10.7% in women aged 30-80 years. However, ATP III criteria for central obesity are not optimal for an Asian-Pacific population; when waist circumference is reduced from 102 to 90 cm in men and 88 to 80 cm in women, the prevalence of the metabolic syndrome increased to 29.0 and 16.8%, respectively. Sex-specific factor analysis showed four factors in men (obesity, glucose intolerance, hypertension, and dyslipidemia) and three in women (obesity hypertension, glucose intolerance, and obesity-dyslipidemia). Insulin resistance estimated from fasting insulin levels clustered with three of the four factors in men and two of the three factors in women. By ATP III or Asian-Pacific waist circumference criteria, the prevalence of the metabolic syndrome increased with increasing tertiles of insulin resistance, which was estimated by a homeostasis model assessment. CONCLUSIONS: The metabolic syndrome is common in an urban Korean population when using Asian-Pacific waist criteria. The prevalence of the metabolic syndrome increased with increasing tertiles of insulin resistance. PMID- 15277436 TI - Inflammation, insulin resistance, and adiposity: a study of first-degree relatives of type 2 diabetic subjects. AB - OBJECTIVE: Inflammatory markers such as C-reactive protein (CRP) are associated with insulin resistance, adiposity, and type 2 diabetes. Whether inflammation causes insulin resistance or is an epiphenomenon of obesity remains unresolved. We aimed to determine whether first-degree relatives of type 2 diabetic subjects differ in insulin sensitivity from control subjects without a family history of diabetes, whether first-degree relatives of type 2 diabetic subjects and control subjects differ in CRP, adiponectin, and complement levels, and whether CRP is related to insulin sensitivity independently of adiposity. RESEARCH DESIGN AND METHODS: We studied 19 young normoglycemic nonobese first-degree relatives of type 2 diabetic subjects and 22 control subjects who were similar for age, sex, and BMI. Insulin sensitivity (glucose infusion rate [GIR]) was measured by the euglycemic-hyperinsulinemic clamp. Dual-energy X-ray absorptiometry determined total and abdominal adiposity. Magnetic resonance imaging measured abdominal adipose tissue volumes. RESULTS: First-degree relatives of type 2 diabetic subjects had a 20% lower GIR than the control group (51.8 +/- 3.9 vs. 64.9 +/- 4.6 micromol x min(-1) x kg fat-free mass(-1), P = 0.04). However, first-degree relatives of subjects with type 2 diabetes and those without a family history of diabetes had normal and comparable levels of CRP, adiponectin, and complement proteins. When the cohort was examined as a whole, CRP was inversely related to GIR (r = -0.33, P = 0.04) and adiponectin (r = -0.34, P = 0.03) and positively related to adiposity (P < 0.04). However, CRP was not related to GIR independently of fat mass. In contrast to C3 (r = 0.41, P = 0.009) and factor B (r = 0.43, P = 0.005), CRP was unrelated to factor D. CONCLUSIONS: The insulin resistant state is not associated with changes in inflammatory markers or complement proteins in subjects at high risk of type 2 diabetes. Our study confirms a strong relationship between CRP and fat mass. Increasing adiposity and insulin resistance may interact to raise CRP levels. PMID- 15277437 TI - Sagittal abdominal diameter is a strong anthropometric marker of insulin resistance and hyperproinsulinemia in obese men. AB - OBJECTIVE: It is clinically important to find noninvasive markers of insulin resistance and hyperproinsulinemia because they both predict cardiovascular and diabetes risk. Sagittal abdominal diameter (SAD) or "supine abdominal height" is a simple anthropometric measure previously shown to predict mortality in men, but its association with insulin resistance and hyperproinsulinemia is unknown. RESEARCH DESIGN AND METHODS: In a common high-risk group of 59 moderately obese men (aged 35-65 years, BMI 32.6 +/- 2.3 kg/m(2)), we determined anthropometry (SAD, BMI, waist girth, and waist-to-hip ratio [WHR]); insulin sensitivity (euglycemic-hyperinsulinemic clamp); and plasma concentrations of intact proinsulin, specific insulin, C-peptide, glucose, and serum IGF binding protein-1 (IGFBP-1). To compare SAD with other anthropometric measures, univariate and multiple regression analyses were used to determine correlations between anthropometric and metabolic variables. RESULTS: SAD showed stronger correlations to all measured metabolic variables, including insulin sensitivity, than BMI, waist girth, and WHR. SAD explained the largest degree of variation in insulin sensitivity (R(2) = 0.38, P < 0.0001) compared with other anthropometric measures. In multiple regression analyses, including all anthropometric measures, SAD was the only independent anthropometric predictor of insulin resistance (P < 0.001) and hyperproinsulinemia (P < 0.001). CONCLUSIONS: In obese men, SAD seems to be a better correlate of insulin resistance and hyperproinsulinemia (i.e., cardiovascular risk) than other anthropometric measures. In overweight and obese individuals, SAD could represent a simple, cheap, and noninvasive tool that could identify the most insulin resistant in both the clinic and clinical trials evaluating insulin sensitizers. These results need confirmation in larger studies that also include women and lean subjects. PMID- 15277438 TI - Caffeine impairs glucose metabolism in type 2 diabetes. PMID- 15277439 TI - Matrix metalloproteinase-9 and tissue inhibitor of metalloproteinase-1 and -2 in type 2 diabetes: effect of 1 year's cardiovascular risk reduction therapy. PMID- 15277440 TI - Pancreatic elastase-1 in stools, a marker of exocrine pancreas function, correlates with both residual beta-cell secretion and metabolic control in type 1 diabetic subjects. PMID- 15277441 TI - Impairment of coronary circulation by acute hyperhomocysteinemia in type 2 diabetic patients. PMID- 15277442 TI - Approach to the pathogenesis and treatment of nonalcoholic steatohepatitis. AB - Nonalcoholic steatohepatitis (NASH) represents an advanced stage of fatty liver disease developed in the absence of alcohol abuse. Its increasing prevalence in western countries, the diagnostic difficulties by noninvasive tests, and the possibility of progression to advanced fibrosis and even cirrhosis make NASH a challenge for hepatologists. NASH is frequently associated with type 2 diabetes and the metabolic syndrome, and several genetic and acquired factors are involved in its pathogenesis. Insulin resistance plays a central role in the development of a steatotic liver, which becomes vulnerable to additional injuries. Several cyclic mechanisms leading to self-enhancement of insulin resistance and hepatic accumulation of fat have been recently identified. Excess intracellular fatty acids, oxidant stress, tumor necrosis factor-alpha, and mitochondrial dysfunction are causes of hepatocellular injury, thereby leading to disease progression and to the establishment of NASH. Intestinal bacterial overgrowth also plays a role, by increasing production of endogenous ethanol and proinflammatory cytokines. Therapeutic strategies aimed at modulating insulin resistance, normalizing lipoprotein metabolism, and downregulating inflammatory mediators with probiotics have promising potential. PMID- 15277443 TI - Weight management through lifestyle modification for the prevention and management of type 2 diabetes: rationale and strategies: a statement of the American Diabetes Association, the North American Association for the Study of Obesity, and the American Society for Clinical Nutrition. PMID- 15277444 TI - Inpatient diabetes control: rationale. PMID- 15277445 TI - Diastolic function and type 1 diabetes. PMID- 15277446 TI - Poor correlation of pulse-wave velocity and intima-media thickness in diabetic subjects. PMID- 15277447 TI - The human insulin analog aspart can induce insulin allergy. PMID- 15277448 TI - Hypoadiponectinemia is closely associated with nonalcoholic hepatic steatosis in obese subjects. PMID- 15277449 TI - Consensus development conference on antipsychotic drugs and obesity and diabetes: response to consensus statement. PMID- 15277450 TI - Consensus development conference on antipsychotic drugs and obesity and diabetes: response to consensus statement. PMID- 15277451 TI - Consensus development conference on antipsychotic drugs and obesity and diabetes: response to consensus statement. PMID- 15277452 TI - Consensus development conference on antipsychotic drugs and obesity and diabetes: response to consensus statement. PMID- 15277454 TI - Randomized trial evaluating a predominately fetal growth-based strategy to guide management of gestational diabetes in caucasian women: response to Schaefer-Graf et al. PMID- 15277456 TI - A standardized triglyceride and carbohydrate challenge: response to Mohanlal and Holman. PMID- 15277458 TI - Role of simvastatin as an immunomodulator in type 2 diabetes: response to Lopes Virella et al. PMID- 15277459 TI - Improper insulin compliance may lead to hepatomegaly and elevated hepatic enzymes in type 1 diabetic patients: response to Yu and Howard. PMID- 15277460 TI - Peripheral arterial disease in people with diabetes: response to consensus statement. PMID- 15277462 TI - Thiazolidinedione use, fluid retention, and congestive heart failure: a consensus statement from the American Heart Association and American Diabetes Association: response to Nesto. PMID- 15277464 TI - Seeing the unseen: Microarray-based gene expression profiling in vision. PMID- 15277465 TI - Structural and elemental evidence for edema in the retina, retinal pigment epithelium, and choroid during recovery from experimentally induced myopia. AB - PURPOSE: The purpose of this study was to monitor temporal changes in the retina, retinal pigment epithelium (RPE), and choroid of chick eyes using biometric, ultrastructural, and elemental microanalysis techniques as a means of visualizing more detailed signs of the physiological processes underlying choroidal expansion and refractive normalization during recovery from form deprivation. METHODS: Axial dimensions and refractions were measured on form-deprived and fellow eyes of 117 experimental chickens reared with monocular translucent occlusion from days 1 to 15 and given different lengths of visual experience (T = 0-144 hours) before death. Tissue was analyzed ultrastructurally by electron microscopy and relative sodium (Na) and chloride (Cl) ion abundances, by using x-ray microanalysis to determine changes in the presence of these indicators of tissue hydration. RESULTS: Refractive error decreased from more than 20 D of myopia almost linearly over the first 144 hours after occlusion. Concurrent changes in thickness in the retina, RPE, and choroid were seen as a series of thickness increases and edema, which returned to normal thickness, first in the retina, and did not reach maximum until 3 days after occluder removal in the choroid. In freeze-dried tissue, Na and Cl ion concentrations were greatest in the RPE photoreceptor outer segments and extravascular choroid at T = 0, decreasing toward fellow eye levels by T = 48 in the RPE and choroid. Na and Cl ion abundances in the frozen lymph of choroidal lymphatics were nearly at control levels (T = 0) and increased later as the vessels became more distended after the extravascular edema became significant. CONCLUSIONS: The results suggest that occluder removal induces edema across the retina and choroid and that this fluid may be the vector eliciting choroidal expansion during recovery from form deprivation possibly driven by the hyperosmolarity in the choroid, RPE, and photoreceptor outer segments that accompanies deprivation. PMID- 15277466 TI - Bves is expressed in the epithelial components of the retina, lens, and cornea. AB - PURPOSE: To demonstrate the expression pattern and subcellular localization of Bves/Pop1a protein, a newly identified cell adhesion molecule, during eye development and corneal regeneration. METHODS: Staged embryonic and adult eyes were assayed using fluorescence immunohistochemistry to detect the Bves protein. A human corneal epithelial (HCE) cell line was used as a model to examine Bves localization during corneal growth and regeneration, with and without antisense morpholino treatment. RESULTS: The data detail the expression and localization of Bves protein before, during, and after differentiation of the eye. In these analyses, Bves was localized to an apical-lateral position in the initial epithelial primordia of the eye. Later, Bves became localized to specific cell types and subcellular domains in the retina, lens, and cornea, indicating changes in Bves expression in the differentiated eye. Finally, an in vitro model of corneal wound healing showed that Bves staining was missing at the epithelial surface during cellular migration across the wound, but it reappeared at points of cell contact during the reinitiation of epithelial continuity. When epithelial sheets were treated with Bves antisense morpholinos to inhibit Bves function, disruption of epithelial integrity was observed. After injury, similar treatment resulted in an acceleration of cell movement at the wound surface but regeneration of an intact epithelium was ultimately impeded. CONCLUSIONS: Taken together, these studies suggest that Bves is expressed in epithelial elements of the developing eye and may have a role in corneal epithelial growth and regeneration. PMID- 15277467 TI - BRAF mutations in conjunctival melanoma. AB - PURPOSE: An activating mutation in exon 15 of the BRAF gene has been found in a high proportion of cutaneous melanomas and cutaneous nevi but not in uveal melanoma. Conjunctival melanoma shows greater clinical similarity to cutaneous melanoma than does uveal melanoma. The purpose of this study was to determine whether the T1799A BRAF mutation found in cutaneous melanoma is also present in conjunctival melanoma. METHODS: DNA was extracted from paraffin sections obtained from glutaraldehyde or formalin-fixed, paraffin-embedded conjunctival melanomas. Forty-two specimens were identified from 25 patients. Seminested PCR was used to amplify exon 15 of the BRAF gene, and the resultant PCR product was purified and directly sequenced. Sequences from conjunctival melanomas were compared with the wild-type sequence of the BRAF gene. The presence or absence of the BRAF mutation was compared with the clinicopathological features. RESULTS: The T1799A (V600E) mutation was detected by sequencing in melanomas from 5 of 22 patients as well as in the positive control, a cutaneous melanoma cell line. In this small series, no statistically significant associations between the presence of the BRAF mutation and clinicopathological characteristics were detected, although tumors with this mutation tended to have a larger diameter and greater depth of invasion and to contain epithelioid cells. CONCLUSIONS: Others have demonstrated a BRAF T1799A activating mutation in cutaneous but not uveal melanoma. In this study, this BRAF mutation was demonstrated in some conjunctival melanoma tissue samples, suggesting that some conjunctival melanomas may share biological features in common with cutaneous melanoma. PMID- 15277468 TI - Downregulation of ATP synthase subunit-6, cytochrome c oxidase-III, and NADH dehydrogenase-3 by bright cyclic light in the rat retina. AB - PURPOSE: Retinas of albino rats born and raised in bright cyclic light (300-800 lux) are less susceptible to light-induced apoptosis than retinas of animals born and raised in dim cyclic light (5 lux). In this study, the objective was to study the mechanisms of neuroprotection in the bright cyclic light-reared retina by identification of differentially expressed genes with differential display (DD) PCR. METHODS: Albino rats were born and raised in 5- or 400-lux cyclic light (12 hours on/off). At 6 to 8 weeks of age, animals were either killed to harvest retinas or exposed to 1700 lux illumination for 12 or 24 hours. The neuroprotection of 400-lux cyclic light rearing was evaluated by DNA fragmentation and quantitative histology. The differentially expressed candidate genes were identified by DD-PCR. Northern blot analysis was used to quantitate differential expression of selected genes. Differential expression of protein was determined by Western blot and enzyme activity analysis. Cellular localization of transcripts was determined by in situ hybridization. RESULTS: DNA fragmentation and quantitative histology results indicated that 400-lux cyclic light rearing protected the retina from light-induced apoptosis compared with 5-lux cyclic light rearing. DD-PCR analysis showed that a 283-bp expressed sequence tag (EST) was downregulated in retinas of rats raised from birth in 400-lux cyclic light. A BLAST search identified the EST as the 3'-terminal sequence of mitochondria encoded NADH dehydrogenase subunit 3 (ND-3). Northern blot analysis showed that the EST hybridized to two mRNA transcripts, the larger of which was confirmed to encompass the adenosine triphosphate (ATP) synthase subunit 6 (ATPase-6), cytochrome c oxidase subunit III (CO-III), ND-3, and tRNA-Gly. Northern blot analysis demonstrated that CO-III and ATPase-6 were downregulated 1.8- and 2.3 fold by 400-lux cyclic light compared with 5-lux cyclic light, respectively; however, there was no change in cytochrome c oxidase subunit I and II (CO-I and II) or in 12S ribosomal RNA (12S rRNA), a mitochondrial housekeeping gene. Western blot analysis using anti-CO-III antibody showed more CO-III protein in retinal mitochondria from dim-light-raised rats. The enzyme activity of CO was two times higher in retinal homogenates from dim-light-raised rats than those from bright-light-raised rats. In situ hybridization using a (35)S-labeled CO-III probe showed that the CO-III transcript was present and downregulated in most of the retinal layers of bright-light-reared animals. CONCLUSIONS: Rearing in cyclic light at 400-lux downregulates the expression of ATPase-6, CO-III, and ND-3 compared with rearing in 5-lux cyclic light. The authors hypothesize that these changes are adaptive responses to light stress that provide neuroprotection to retinal cells by elevating the level of stress-related factors and reducing the level of oxidized cytochrome c, the form that activates the apoptotic cascade of cell death. PMID- 15277469 TI - Effects of rolipram, a selective inhibitor of type 4 phosphodiesterase, on lipopolysaccharide-induced uveitis in rats. AB - PURPOSE: To investigate effects of rolipram, an inhibitor of type 4 phosphodiesterase, on lipopolysaccharide (LPS)-induced uveitis in Wistar rats. METHODS: A total of 100 microg LPS was injected into the rat footpad. Rolipram (Wako Pure Chemical, Osaka, Japan) was injected intraperitoneally 30 minutes before administration of LPS. Levels of intracameral protein, cells, E-selectin, tumor necrosis factor (TNF)-alpha, interleukin (IL)-6, and nitrite were determined. E-selectin, TNF-alpha, IL-6, and inducible nitric oxide synthase (iNOS) mRNAs and immunohistochemical reactivity of nuclear factor (NF)-kappa B and TNF-alpha were also examined in the iris-ciliary body. RESULTS: After LPS injection, intracameral protein and cells increased from 18 to 30 hours later. Rolipram, however, inhibited elevation of protein and cells. After LPS injection, mRNA levels of E-selectin, TNF-alpha, and IL-6 in the iris-ciliary body increased 3 hours later, and iNOS mRNA increased 6 hours later. Elevation of mRNA levels for E-selectin, TNF-alpha, and IL-6 was inhibited by rolipram. After LPS injection, intracameral TNF-alpha and IL-6 levels increased 4 to 6 hours later, and nitrite levels increased 14 to 20 hours later. Elevation of TNF-alpha and IL 6 levels was decreased by rolipram. Rolipram did not affect iNOS mRNA and nitrite levels. Immunoreactivity of NF-kappa B was strong 1 hour after LPS injection, and was decreased by rolipram. Immunoreactivity of TNF-alpha was strong 4 hours after LPS injection and was decreased by rolipram. CONCLUSIONS: NF-kappa B translocation and expression of E-selectin, TNF-alpha, and IL-6 are involved in the pathogenesis of LPS-induced uveitis and are inhibited by rolipram. The inhibitory effect of rolipram in uveitis may be independent of iNOS synthesis. PMID- 15277470 TI - Gene expression profiling of purified rat retinal ganglion cells. AB - PURPOSE: The phenotype of specialized cells arises, in part, from their characteristic gene expression patterns. Retinal ganglion cells (RGCs) are of wide interest in neuroscience and die in glaucoma and other optic neuropathies. In this study the genes expressed by RGCs were profiled by expressed sequence tag (EST) analysis. METHODS: ESTs were generated from a cDNA library constructed from RGCs isolated by immunopanning. The RGC genes were compared with published microarray expression profiles from 13 different neural regions. Immunohistochemistry was performed by standard methods. RESULTS: Clustering of 4791 RGC ESTs identified 2360 unique gene clusters. Of these, 60% represented known genes, 27% uncharacterized genes/ESTs, and 13% novel sequence. Unexpectedly, one of the largest RGC clusters, RESP18, corresponded to a neuroendocrine-specific gene preferentially expressed in the hypothalamus. RESP18 immunoreactivity within the retina was found mainly in the RGC layer. DDAH1, a gene involved in nitric oxide metabolism, was localized to RGC and amacrine layers. Comparison of gene expression patterns across neuronal regions revealed a prominent subset of RGC genes that were overexpressed in dorsal root and trigeminal ganglia. To narrow the search for candidate disease-related genes, RGC genes were mapped to known disease loci for optic neuropathies. CONCLUSIONS: This work is one of the first efforts to profile gene expression in a purified population of retinal neurons, the RGCs. The profiling, in addition to revealing both known and novel genes underlying the RGC phenotype, also uncovered common patterns of gene expression between RGCs and other sensory ganglia. PMID- 15277471 TI - Modulating expression of peripherin/rds in transgenic mice: critical levels and the effect of overexpression. AB - PURPOSE: Mutations in the photoreceptor-specific protein peripherin/rds are associated with multiple retinal diseases. To date, attempts to achieve complete structural and functional rescue in animal models of peripherin/rds-induced retinal degeneration have not been successful. Gene therapy-directed approaches have been hindered by the haploinsufficiency phenotype, which dictates well regulated expression of peripherin/rds protein levels. METHODS: Using a transgenic mouse line expressing wild-type peripherin/rds (NMP), the authors evaluated the critical in vivo level of peripherin/rds needed to maintain photoreceptor structure and ERG function and assessed the consequences of peripherin/rds overexpression in both rods and cones by Western blot and immunoprecipitation analyses, immunohistochemistry, electron microscopy, and electroretinography. The NMP transgene included a C-terminal modification (P341Q) to facilitate detection of the transgenic protein in the presence of wild-type peripherin/rds, using the monoclonal antibody 3B6. RESULTS: Peripherin/rds protein levels in NMP homozygotes were approximately 60% of wild-type levels. Western blot and immunoprecipitation analyses confirmed normal biochemical properties of the NMP protein when compared with wild-type peripherin/rds. Immunohistochemistry demonstrated appropriate localization of transgenic peripherin/rds protein to the disc rim region of photoreceptor outer segments. Total peripherin/rds levels in the retina were modulated by crossing NMP transgenic mice into different rds genetic backgrounds. A positive correlation was observed between peripherin/rds expression levels and the structural and functional integrity of photoreceptor outer segments. Overexpression of peripherin/rds caused no detectable adverse effects on rod or cone structure and function. CONCLUSIONS: These findings may have significant implications regarding therapeutic intervention in peripherin/rds-associated retinal diseases. PMID- 15277473 TI - The wing 2 region of the FOXC1 forkhead domain is necessary for normal DNA binding and transactivation functions. AB - PURPOSE: To determine the biochemical defects that underlie Axenfeld-Rieger malformations, to determine a functional role for wing 2 in FOXC1, and to understand how mutations in this region disrupt FOXC1 function. METHODS: Sequencing DNA from patients with Axenfeld-Rieger malformation resulted in the identification of two novel missense mutations (G165R and R169P) in wing 2 of FOXC1. Site-directed mutagenesis was used to introduce these mutations, as well as previously reported mutation (M161K), into the FOXC1 cDNA. These FOXC1 mutants were evaluated to determine their ability to localize to the nucleus, bind DNA and activate gene expression. RESULTS: Two novel missense mutations were identified in unrelated patients, in wing 2 of the FOXC1 forkhead domain. Because there had been no previous biochemical analysis, the mutation M161K was also investigated. All three mutant proteins localized correctly to the nucleus. The G165R mutation maintained wild-type levels of DNA binding; however, both the M161K and R169P mutations displayed reduced DNA binding ability. Biochemical analysis showed that all three mutations disrupt FOXC1's transactivation ability. CONCLUSIONS: Biochemical analysis of mutations G165R and R169P and of a previously reported mutation, M161K, demonstrate the functional significance of wing 2. M161K and R169P disrupt DNA binding of FOXC1, consistent with the hypothesis that wing 2 is necessary for DNA binding. The results also suggest that wing 2 plays a role in gene activation. These results provide the first insights into how mutations in wing 2 disrupt FOXC1 function. PMID- 15277472 TI - Kruppel-like factor 15, a zinc-finger transcriptional regulator, represses the rhodopsin and interphotoreceptor retinoid-binding protein promoters. AB - PURPOSE: To identify novel transcriptional regulators of rhodopsin expression as a model for understanding photoreceptor-specific gene regulation. METHODS: A bovine retinal cDNA library was screened in a yeast one-hybrid assay, with a 29 bp bovine rhodopsin promoter fragment as bait. Expression studies used RT-PCR and beta-galactosidase (LacZ) histochemistry of retinas from transgenic mice heterozygous for a targeted LacZ replacement of KLF15. Promoter transactivation assays measured luciferase expression in HEK293 cells transiently transfected with bovine rhodopsin or IRBP promoter-reporter constructs and expression constructs containing cDNAs for full or truncated KLF15, Crx (cone rod homeobox), and/or Nrl (neural retina leucine zipper). Data were analyzed with general linear models. RESULTS: The zinc-finger transcription factor KLF15 was identified as a rhodopsin-promoter-binding protein in a yeast one-hybrid screen. Expression was detected by RT-PCR in multiple tissues, including the retina, where KLF15-LacZ was observed in the inner nuclear layer, ganglion cell layer, and pigmented epithelial cells, but not in photoreceptors. KLF15 repressed transactivation of rhodopsin and IRBP promoters alone and in combination with the transcriptional activators Crx and/or Nrl. Repressor activity required both a 198-amino-acid element in the N-terminal domain and the C-terminal zinc finger DNA-binding domains. CONCLUSIONS: The zinc finger containing transcription factor KLF15 is a transcriptional repressor of the rhodopsin and IRBP promoters in vitro and, in the retina, is a possible participant in repression of photoreceptor-specific gene expression in nonphotoreceptor cells. PMID- 15277474 TI - The need for routine eye examinations. AB - PURPOSE: To determine the incidence and causes of vision loss to assist in the objective determination of the preferred frequency of routine screening for those with normal vision. METHODS: A prospective, population-based study of people aged 40 or more years. Standardized examination protocols were used that included presenting and best corrected visual acuity, visual field testing, and comprehensive eye examination with dilation. RESULTS: There were 2529 people with a full data set, including 1590 with a normal baseline examination. The 5-year incidence of vision loss (<6/12 presenting acuity in the worse eye) was 2.39%. Overall, 24 (63%) of 38 of those with vision loss had noticed a change in their vision, and 18 (75%) of these 24 had attended an eye examination. This left only 14 (0.88%) people who had had normal baseline examination results and had asymptomatic vision loss develop over this 5-year period. CONCLUSIONS: Frequent routine eye examinations of those with normal examination results will have a low yield and may not be cost effective. Health promotion messages should target those who notice a change in vision and those at higher risk such as those with diabetes or a family history of eye disease. PMID- 15277476 TI - The impact of hydrogel lens settling on the thickness of the tears and contact lens. AB - PURPOSE: To investigate the effect of contact lens insertion on the thickness on the prelens tear film (PLTF), the contact lens, and the postlens tear film (PoLTF). METHODS: Twelve contact lens wearers (mean age, 32.7 years; four males) inserted etafilcon A hydrogel lenses (power: -2.00 D, base curve: 8.3 mm) in both eyes immediately before testing. Previously described interference techniques, based on oscillations in reflectance spectra, were used to measure the thickness of the PLTF, contact lens, and PoLTF. The thickness of the layers is derived from the frequency of the oscillations. Spectra were captured 1 minute after lens insertion and every minute thereafter for 30 minutes. Least squares regression fits were used to determine the relation between thickness of each layer and time. RESULTS: The combined data from all subjects for PLTF thickness were fit with an exponential decay plus a constant thickness; the initial thickness was 4.5 micrometers, the time constant was 7.1 minutes (P < 0.001), and final thickness was 2.5 micrometers. The apparent thickness of the contact lens declined linearly at an average rate of 0.051 micrometers/minute (P < 0.001). The PoLTF thickness remained constant at 2.5 micrometers (P = 0.46). CONCLUSIONS: For most subjects, the PLTF thinned significantly over the course of the first 30 minutes of lens wear. The apparent thinning of the contact lens may be caused by a real thinning of the lens, but also may have a contribution from improved centration over the 30-minute period. The PoLTF remained relatively stable during this period. PMID- 15277475 TI - Delivery of gentamicin to the rabbit eye by drug-loaded hydrogel iontophoresis. AB - PURPOSE: To assess the corneal iontophoretic delivery of gentamicin by drug loaded hydrogel probe, and to determine the resultant ocular disposition and elimination of the drug from the cornea and anterior chamber. METHODS: Corneal iontophoresis of gentamicin sulfate was studied in healthy white rabbits by using drug-loaded disposable hydroxyethyl methacrylate (HEMA) hydrogel disk probes and a portable mini-ion device designed in the authors' laboratory. The iontophoretic treatment was performed with a current intensity of 1 mA for 60 seconds only. Three control groups were used: mock iontophoresis (no current) for 60 seconds, topical eye drops of fortified gentamicin (1.4%) every 5 minutes for 1 hour, and subconjunctival injection of 0.25 mL of 40 mg/mL gentamicin solution. The animals in the iontophoretic experimental groups were killed at predetermined time points. The gentamicin concentrations in the cornea and aqueous humor were assayed with a fluorescence polarization immunoassay. Analysis of the gentamicin eye pharmacokinetics was performed with a modeling approach. RESULTS: Peak gentamicin concentrations in the cornea (363.1 +/- 127.3 microg/g) and in the aqueous humor (29.4 +/- 17.4 microg/mL) were reached at 0 and 2 hours after the iontophoretic treatment, respectively. The peak gentamicin concentrations after a single iontophoresis treatment were 12 to 15 times higher than those obtained after gentamicin injection or after topical eye drop instillation, and much higher than in mock iontophoresis. The concentration versus time profile of gentamicin in the cornea and the anterior chamber after iontophoresis was appropriately described by applying a two-compartment pharmacokinetic model. CONCLUSIONS: A short iontophoretic treatment using gentamicin-loaded hydrogels has potential clinical value in increasing drug penetration to the anterior segments of the eye and maintaining therapeutic drug levels in the cornea for more than 8 hours. PMID- 15277477 TI - Monitoring of rabbit cornea response to dehydration stress by optical coherence tomography. AB - PURPOSE: To evaluate the application of optical coherence tomography (OCT) for continuous noninvasive monitoring and quantification of the dynamics of corneal response after exposure of the cornea to dehydrating stress. METHODS: The changes in central corneal thickness (CCT) and scattering properties of the cornea were monitored with OCT in rabbit cornea in vivo after topical application of a glycerin-based hypertonic agent (HA) or prolonged surface evaporation of the cornea. The observed changes in backscatter were correlated with the changes in corneal hydration. RESULTS: An inverse relationship was found between the logarithm of the intensity of backscatter within the cornea and the degree of corneal hydration at which the intensity of the backscatter changed up to 20 times between the peak of the de- and rehydration phases. An analytical relationship is derived between the magnitude of the backscatter from the stroma and the extent of corneal hydration. Furthermore, depending on the concentration of the drug, a peak overshoot in corneal thickness in the range of 40% to 90% was detected during the rehydration process after topical application of the HA. At a 100% concentration of HA, the average dehydration rate was 74 microm/min, whereas the average rehydration rate was 12.4 microm/min. The same parameters for surface evaporation were 2.7 and 1.5 microm/min, respectively. CONCLUSIONS: OCT may offer a unique capability to quantitatively monitor the dynamics of corneal response and to assess corneal function based on noninvasive detection of the changes in the optical properties and morphology of the cornea after topical application of dehydrating agents. PMID- 15277478 TI - Effect of environmental conditions on tear dynamics in soft contact lens wearers. AB - PURPOSE: Dry eye symptoms are often associated with soft contact lens (SCL) wear and may be affected by environmental conditions. This study was conducted to determine the effects of temperature and humidity on the tear film in SCL wearers. METHODS: All 11 enrolled subjects were males (mean age, 23.5 +/- 5.2 [SD] years), and all wore SCL daily. They were exposed in different sessions to four different conditions in an environmental chamber with the air temperature (AT) and relative humidity (RH) set at 5 degrees C/10% (AT/RH), 15 degrees C/20%, 25 degrees C/40%, or 35 degrees C/50%. Two different types of hydrogel SCL (SCL-a and SCL-b; water content 72.0% and 37.5%, respectively) were used. The meniscus tear volume was determined on a video meniscometer by measuring the tear meniscus radius (TMR) with and without SCL. The tear interference patterns on the contact lens (TIPCL) were classified into five grades (the higher the grade, the thinner the film). Using a video interferometer, the non-invasive tear film breakup time (NIBUT) was recorded with and without SCLs; ocular dryness was also scored with and without SCLs. RESULTS: Under the environmental conditions examined, there were no significant differences in the TMR without or with SCL, regardless of their type. As AT and RH decreased, there was a significant increase in the TIPCL grade (CL-a: P = 0.042; CL-b: P = 0.002), a significant decrease in NIBUT (CL-a: P = 0.004; CL-b: P = 0.001), and a significant increase in the dryness score (without SCL P = 0.023; with CL-a P = 0.009; with CL-b P = 0.003). The dryness scores were higher with CL-a than CL-b (P = 0.011 at 15 degrees C/20%). Under identical experimental conditions, we observed no significant change in NIBUT in the absence of an SCL. CONCLUSIONS: AT and RH apparently had no effect on the tear volume in the presence of SCLs. As AT and RH decreased, the tear film on the SCL became thinner, NIBUT became shorter, and dryness increased. Dryness was more pronounced in eyes with SCL of the higher water content. PMID- 15277480 TI - Sonic hedgehog expression and role in healing corneal epithelium. AB - PURPOSE: To examine the expression pattern and roles of Sonic hedgehog (Shh) in healing corneal epithelium. METHODS: Immunofluorescent staining and Western blot analysis were used to detect Shh, patched 1 (Ptc 1) receptors, and Gli transcription factors in corneal epithelium of Wistar rats (n = 44) at various intervals after an epithelial defect. Effects of exogenous Shh on cell proliferation and cyclin D1 expression were determined in healing corneal epithelium of organ-cultured mouse eyes. RESULTS: Uninjured rat corneal epithelium was not labeled by anti-Shh antibody, but weakly positive for Ptc 1. Basal cells of limbal and conjunctival epithelia were labeled by antibodies against Shh and Ptc 1. Shh protein was transiently upregulated in limbal epithelium in 2 hours and was also transiently expressed in the migrating corneal epithelium with its peak at 12 hours postdebridement. Such upregulation of Shh expression was associated with a transient nuclear translocation of Gli-3 without lifting the suppression of cell proliferation in migrating epithelium postdebridement in vivo. However, an addition of Shh protein to culture medium resulted in nuclear accumulation of cyclin D1 and marked acceleration of epithelial cell proliferation in migrating corneal epithelium of an organ cultured mouse eye. CONCLUSIONS: Corneal epithelial debridement causes a transient upregulation of Shh expression and activation of Shh/Gli-3 signaling cascade in healing corneal and limbal epithelia. Although exogenous Shh promotes epithelial cell proliferation in corneal organ culture, its expression in migrating epithelium in vivo does not counteract the suppression of cell proliferation at the early healing phase of epithelium debridement. PMID- 15277479 TI - Role of EGFR transactivation in preventing apoptosis in Pseudomonas aeruginosa infected human corneal epithelial cells. AB - PURPOSE: To determine the role of epidermal growth factor (EGF) receptor (EGFR) mediated signaling pathways in preventing infection-induced apoptosis in human corneal epithelial cells (HCECs). METHODS: Epithelial monolayers of a telomerase immortalized HCEC line, HUCL, and primary culture of HCECs were infected with Pseudomonas aeruginosa in the presence of the EGFR inhibitor tyrphostin AG1478, the extracellular signal-regulated kinase (ERK) inhibitor U0126, the phosphoinositide 3-kinase (PI3K) inhibitor LY294002, the heparin-binding EGF-like growth factor (HB-EGF) antagonist CRM197, the HB-EGF neutralizing antibody, or the matrix metalloproteinase inhibitor GM6001. The activation of EGFR was analyzed by immunoprecipitation using EGFR antibodies, followed by Western blot analysis with phosphotyrosine antibody. Phosphorylation of ERK and Akt, a major substrate of PI3K, and generation of cleaved caspase-3 and poly (ADP-ribose) polymerase (PARP) were determined by Western blot analysis. Apoptotic cells were characterized by positive staining of active caspase-3, loss of mitochondrial cytochrome c, and condensation of chromosomes. Apoptosis was also confirmed by measuring caspase-3 activity and assessing the generation of cleaved caspase-3 and PARP. RESULTS: P. aeruginosa infection of HUCL cells resulted in EGFR activation and EGFR-dependent ERK1/2 and PI3K phosphorylation. Inhibition of EGFR, ERK1/2, and PI3K activities with kinase-specific inhibitors (AG1478, U0126, and LY294002, respectively) resulted in an increase in the number of apoptotic cells, in elevated cellular caspase-3 activity, and/or in increased cleaved PARP in P. aeruginosa-infected HUCL cells or primary culture of HCECs. Blocking HB-EGF ectodomain shedding by inhibition of matrix metalloproteinase-mediated proteolysis, downregulation of HB-EGF, or neutralization of its activity retarded infection-induced EGFR transactivation and, as a consequence, increased infection induced HUCL apoptosis. CONCLUSIONS: Bacterial infection of HCECs induces EGFR transactivation through HB-EGF ectodomain shedding. EGFR and its downstream ERK and PI3K signaling pathways play a role in preventing epithelial apoptosis in the early stage of bacterial infection. PMID- 15277481 TI - In vitro antiangiogenic activity in ex vivo expanded human limbocorneal epithelial cells cultivated on human amniotic membrane. AB - PURPOSE: To compare the in vitro antiangiogenic activities of ex vivo expanded human limbocorneal epithelial (HLE) cells cultivated on preserved human amniotic membrane (AM) and to identify factors responsible for the activities. METHODS: The antiangiogenic effects were compared of culture media conditioned by AM, HLE cells, or HLE cells cultivated on intact AM (HLE/IAM), on denuded AM (HLE/DAM), or on DAM cocultured with 3T3 fibroblasts (HLE/DAM/3T3). A monolayer culture of human umbilical vein endothelial cells (ECs) was used in a proliferation and migration assay. ECs suspended in type I collagen gel were used to assess capillary tube formation. Quantitative analyses of tissue inhibitor of metalloproteinase (TIMP)-1, thrombospondin (TSP)-1, pigment epithelium-derived factor (PEDF), and endostatin (proteolytic fragment of collagen XVIII) were performed by ELISA. Immunoconfocal microscopy was performed to localize the site of endostatin expression in HLE cells and AM. RESULTS: HLE cell- but not AM conditioned medium (CM) inhibited the proliferation and migration of ECs, and coculture of HLE cells, but not of AM, with ECs inhibited capillary tube formation. Although some data from HLE cells alone are not significantly different from the control, increased inhibitory activity was expressed by HLE/IAM and HLE/DAM and was most significantly expressed by HLE/DAM/3T3. Quantitation of TIMP-1, TSP-1, PEDF, and endostatin revealed that only the level of endostatin showed an increased expression by HLE cells cultivated on AM. Neutralizing antibody to endostatin substantially abrogated the inhibitory effect on EC proliferation and migration, but was less effective on EC differentiation. Endostatin signal was more prominent in the basement membrane zone of HLE cells cultivated on denuded AM than in those cultivated on intact AM. CONCLUSIONS: The antiangiogenic effect of HLE cells was enhanced when they were cultivated on AM and cocultured with 3T3 fibroblasts, and endostatin-related antiangiogenic factor may play a major role. This highlights the significance of cell-matrix and cell cell interaction in the regulation of antiangiogenic factor secretion by HLE cells. PMID- 15277482 TI - Using unsupervised learning with variational bayesian mixture of factor analysis to identify patterns of glaucomatous visual field defects. AB - PURPOSE: To determine whether an unsupervised machine learning classifier can identify patterns of visual field loss in standard visual fields consistent with typical patterns learned by decades of human experience. METHODS: Standard perimetry thresholds for 52 locations plus age from one eye of each of 156 patients with glaucomatous optic neuropathy (GON) and 189 eyes of healthy subjects were clustered with an unsupervised machine classifier, variational Bayesian mixture of factor analysis (vbMFA). RESULTS: The vbMFA formed five distinct clusters. Cluster 5 held 186 of 189 fields from normal eyes plus 46 from eyes with GON. These fields were then judged within normal limits by several traditional methods. Each of the other four clusters could be described by the pattern of loss found within it. Cluster 1 (71 GON + 3 normal optic discs) included early, localized defects. A purely diffuse component was rare. Cluster 2 (26 GON) exhibited primarily deep superior hemifield defects, and cluster 3 (10 GON) held deep inferior hemifield defects only or in combination with lesser superior field defects. Cluster 4 (6 GON) showed deep defects in both hemifields. In other words, visual fields within a given cluster had similar patterns of loss that differed from the predominant pattern found in other clusters. The classifier separated the data based solely on the patterns of loss within the fields, without being guided by the diagnosis, placing 98.4% of the healthy eyes within the same cluster and spreading 70.5% of the eyes with GON across the other four clusters, in good agreement with a glaucoma expert and pattern standard deviation. CONCLUSIONS: Without training-based diagnosis (unsupervised learning), the vbMFA identified four important patterns of field loss in eyes with GON in a manner consistent with years of clinical experience. PMID- 15277483 TI - Thickness and birefringence of healthy retinal nerve fiber layer tissue measured with polarization-sensitive optical coherence tomography. AB - PURPOSE: Thinning of the retinal nerve fiber layer and changes in retinal nerve fiber layer (RNFL) birefringence may both precede clinically detectable glaucomatous vision loss. Early detection of RNFL changes may enable treatment to prevent permanent loss of vision. Polarization-sensitive optical coherence tomography (PS-OCT) can provide objective information on RNFL thickness and birefringence. METHODS: PS-OCT scans around the optic nerve head (ONH) of two healthy young volunteers were made using 10 concentric circles of increasing radius. Both the mean RNFL thickness and mean retinal nerve fiber birefringence for each of 48 sectors on a circle were determined with data analysis. RESULTS: Both the RNFL thickness and birefringence varied as a function of sector around the ONH. The RNFL became thinner with increasing distance from the ONH. In contrast, the birefringence did not vary significantly as a function of radius. CONCLUSIONS: Birefringence of healthy RNFL is constant as a function of scan radius but varies as a function of position around the ONH, with higher thickness values occurring superior and inferior to the ONH. Measured double-pass phase retardation per unit depth around the ONH ranged between 0.10 and 0.35 deg/microm, equivalent to birefringences of 1.2 x 10(-4) and 4.1 x 10(-4) respectively, measured at a wavelength of 840 nm. Consequently, when a spatially constant birefringence around the ONH is assumed, the conversion of scanning laser polarimetry (SLP) phase-retardation measurements to RNFL thickness may yield incorrect values. The data do not invalidate the clinical value of a phase retardation measurement, but affect the conversion of phase retardation to RNFL thickness. PMID- 15277484 TI - Predictive factors of the optic nerve head for development or progression of glaucomatous visual field loss. AB - PURPOSE: To evaluate which morphologic features of the optic disc are predictive factors for the development or progression of visual field loss in chronic open angle glaucoma. METHODS: The prospective observational clinical study included 763 eyes of 416 white subjects with ocular hypertension and chronic open-angle glaucoma. During the follow-up time (mean, 67.4 months; median, 65.1; range, 6.2 104.5), all patients underwent repeated qualitative and morphometric evaluation of color stereo optic disc photographs and white-on-white visual field examination. Progression of glaucomatous visual field damage was defined by point wise regression analysis for each of the 59 locations in the visual field. Outcome measures were qualitative and quantitative morphologic optic nerve head parameters. RESULTS: Development or progression of glaucomatous visual field defects was detected in 106 (13.9%) eyes. At baseline of the study, neuroretinal rim area was significantly (P < 0.002) smaller, the beta zone of parapapillary atrophy (P < 0.003, nasal sector) was significantly larger, and age was significantly higher (P < 0.003) in the progressive study group than in the nonprogressive study group. Both study groups did not vary significantly in size of the optic disc and the alpha zone of parapapillary atrophy. Cox proportional hazard regression analysis revealed that the progression of glaucomatous visual field loss depended significantly on the area of the neuroretinal rim (P < 0.001) and age (P < 0.001), but was independent of diameter of the retinal arterioles and veins. CONCLUSIONS: Morphologic predictive factors for development or progression of glaucomatous visual field defects in whites are small neuroretinal rim area and large beta zone of parapapillary atrophy. Age is an additional nonmorphologic parameter. Progression of glaucomatous optic nerve head changes is independent of the size of the optic disc and alpha-zone of parapapillary atrophy and retinal vessel diameter. PMID- 15277485 TI - Alkylphosphocholines: a new therapeutic option in glaucoma filtration surgery. AB - PURPOSE: To investigate the effect of alkylphosphocholines (APCs) on human Tenon fibroblast (HTF) proliferation, migration, and cell-mediated collagen gel contraction. METHODS: HTFs were isolated from tissue samples of three patients obtained during surgery and cultured in DMEM and 10% fetal calf serum (FCS). HTFs (passage 3-6) were treated with one APC in different concentrations spanning the 50% inhibitory concentration (IC(50)), as determined previously. Inhibition of cell proliferation was assessed by the tetrazolium dye reduction assay. Migration was determined in chemoattractant chambers with fibronectin-coated polycarbonated membranes. For inhibition of contraction, three-dimensional collagen gels were seeded with HTFs, and the gel size was measured. Cell viability was determined by the trypan blue exclusion assay. For analysis of the mechanism of action, protein kinase C (PKC) activity was measured. RESULTS: The IC(50) varied between 7.0 and 10.5 microM for all APCs tested. At this concentration, all four APCs inhibited HTF migration and cell-mediated collagen gel contraction in the presence of serum. The inhibitory effects on HTF proliferation, migration, and contraction were observed at nontoxic concentrations. PKC activity was reduced to 50% of control level at the IC(50) of all APCs applied. CONCLUSIONS: APCs are effective inhibitors of HTF proliferation, migration, and cell-mediated contraction of collagen gels at nontoxic concentrations. Their mechanism of action seems to involve the inhibition of the PKC pathway. PMID- 15277486 TI - Efficacy and safety of memantine treatment for reduction of changes associated with experimental glaucoma in monkey, I: Functional measures. AB - PURPOSE: To determine, using electrophysiological measures of visual system function, whether oral daily dosing of memantine is both safe and effective to reduce the injury associated with experimental glaucoma in primates. METHODS: Argon laser treatment of the anterior chamber angle was used to induce chronic ocular hypertension (COHT) in the right eye of 18 macaque monkeys. Nine animals were orally dosed daily with 4 mg/kg memantine while the other nine animals received an oral dose of vehicle only. Using both conventional and multifocal methods, recordings of the electroretinogram (ERG) were made at approximately 3, 5, and 16 months after elevation of the intraocular pressure (IOP). Recordings of the visually-evoked cortical potential (VECP) were also made at the 16-month time point. RESULTS: Chronic ocular hypertension was associated with a reduction in the amplitude of components of the multifocal ERG response and visually-evoked cortical potential. Memantine-treated animals suffered less amplitude reduction for these measures than did vehicle-treated animals, though this treatment effect on the ERG measures was observed only at the early time points (3 and 5 months post IOP elevation). Memantine treatment was not associated with an effect on either the kinetics or amplitude of ERG or VECP response measures obtained from the normotensive eyes. CONCLUSIONS: Systemic treatment with memantine, a compound which does not lower intraocular pressure, was both safe and effective for reduction of functional loss associated with experimental glaucoma. PMID- 15277487 TI - Efficacy and safety of memantine treatment for reduction of changes associated with experimental glaucoma in monkey, II: Structural measures. AB - PURPOSE: To determine, using anatomic measurements, whether daily oral dosing with memantine is both safe and effective to reduce the injury associated with experimental glaucoma in primates. METHODS: Argon laser treatment of the anterior chamber angle was used to induce chronic ocular hypertension (COHT) in the right eyes of 18 macaque monkeys. Nine animals were daily orally dosed with 4 mg/kg memantine while the other nine animals received vehicle only. Measurements of intraocular pressure (IOP) from both eyes of all animals were made at regular intervals. Appearance of the optic nerve head, retinal vessels, and surrounding retina was documented with stereo fundus photographs obtained at multiple time points throughout the study. Measurements of optic nerve head topography were obtained from confocal laser scans made from animals with the highest IOPs at approximately 3, 5, and 10 months after elevation of IOP. At approximately 16 months after IOP elevation, animals were killed and histologic counts of cells in the retinal ganglion cell (RGC) layer were made. RESULTS: Histologic measurements showed that, for animals with moderate elevation of IOP, memantine treatment was associated with an enhanced survival of RGCs in the inferior retina. Measurements of optic nerve head topography showed less IOP-induced change in memantine treated animals. This effect was seen in measurements of both the cup and the neuroretinal rim. A comparison of these same histologic and morphologic measurements in normotensive eyes from the two treatment groups showed that memantine treatment was not associated with any significant effects on these eyes. CONCLUSIONS: Histologic measurements of RGC survival as well as tomographic measurements of nerve head topography show that systemic treatment with memantine, a compound which does not lower intraocular pressure, is both safe and effective to reduce changes associated with experimental glaucoma. PMID- 15277488 TI - Analysis of porcine optineurin and myocilin expression in trabecular meshwork cells and astrocytes from optic nerve head. AB - PURPOSE: To determine the cDNA sequences and analyze the expression of porcine optineurin and myocilin in trabecular meshwork cells (TMCs) and astrocytes from the optic nerve head under normal and experimental conditions. METHODS: Both porcine optineurin and myocilin were cloned to determine the cDNA sequences. Porcine TMCs and astrocytes were isolated and treated with dexamethasone (500 nM) for 2 weeks, incubated under hypoxic conditions (7% O(2)) for 72 hours, or exposed to 33 mm Hg hydrostatic pressure for 72 hours. A 10% mechanical stretch for 24 hours was also performed on TMCs. The expression level of the optineurin and myocilin transcripts was analyzed by real-time quantitative PCR. RESULTS: The sequences of porcine optineurin and myocilin cDNA were determined, and the expression of both genes was confirmed in both TMCs and astrocytes. Amino acid sequences of porcine optineurin and myocilin were homologous to those of humans by 84% and 82%, respectively, and shared protein motifs and modification sites. The expression of myocilin mRNA by TMCs and astrocytes was increased by 8.0- and 5.5-fold, respectively, after exposure to dexamethasone. In contrast, the expression of optineurin was suppressed to 68% in TMCs and 48% in astrocytes after exposure to dexamethasone. A significant reduction of myocilin expression was observed after 72 hours of incubation under hypoxic conditions in both types of cells, whereas optineurin was not affected. Hydrostatic pressure for 72 hours and mechanical stretching for 24 hours had minimal affects on gene expression of both optineurin and myocilin. CONCLUSIONS: The high homology of porcine optineurin and myocilin to the comparable human genes indicates that pigs can be used to study changes in gene expression in hypertensive eyes. The alterations in expression of myocilin but not of optineurin under stress suggest that different mechanisms in the phenotype of glaucoma associated with the two genes are involved in development of glaucoma. PMID- 15277489 TI - Lamina cribrosa thickness and spatial relationships between intraocular space and cerebrospinal fluid space in highly myopic eyes. AB - PURPOSE: To evaluate the spatial relationships of the intraocular space, the cerebrospinal fluid space, and the lamina cribrosa in highly myopic eyes. METHODS: The study included 36 human globes with an axial length of more than 26.5 mm that showed marked glaucomatous optic nerve damage (n = 29; highly myopic glaucomatous group) or in which the optic nerve was affected by neither glaucoma nor any other disease (n = 7; highly myopic normal group). Two non-highly myopic control groups included 53 globes enucleated because of malignant choroidal melanoma (n = 42; non-highly myopic normal group) or because of painful absolute secondary angle-closure glaucoma (n = 11; non-highly myopic glaucomatous group). Anterior-posterior histologic sections through the pupil and the optic disc were morphometrically evaluated. RESULTS: In both highly myopic groups compared with both non-highly myopic groups and in the highly myopic glaucomatous group compared with the highly myopic normal group, the lamina cribrosa was significantly (P < 0.001) thinner. Correspondingly, the distance between the intraocular space and the cerebrospinal fluid space was significantly (P < 0.05) shorter in the highly myopic normal group than in the non-highly myopic normal group and in the highly myopic glaucomatous group than in the highly myopic normal group. CONCLUSIONS: In highly myopic eyes, the lamina cribrosa is significantly thinner than in non-highly myopic eyes, which decreases the distance between the intraocular space and the cerebrospinal fluid space and steepens the translaminar pressure gradient at a given intraocular pressure, which may explain the increased susceptibility to glaucoma in highly myopic eyes. As in non-highly myopic eyes, thinning of the lamina cribrosa gets more pronounced in highly myopic eyes if glaucoma is also present. PMID- 15277490 TI - Inhibition of hemangiogenesis and lymphangiogenesis after normal-risk corneal transplantation by neutralizing VEGF promotes graft survival. AB - PURPOSE: To evaluate the occurrence and time course of hem- and lymphangiogenesis after normal-risk corneal transplantation in the mouse model and to test whether pharmacologic strategies inhibiting both processes improve long-term graft survival. METHODS: Normal-risk allogeneic (C57BL/6 to BALB/c) and syngeneic (BALB/c to BALB/c) corneal transplantations were performed and occurrence and time course of hem- and lymphangiogenesis after keratoplasty was observed, by using double immunofluorescence of corneal flatmounts (with CD31 as a panendothelial and LYVE-1 as a lymphatic vascular endothelium-specific marker). A molecular trap designed to eliminate VEGF-A (VEGF Trap(R1R2); 12.5 mg/kg) was tested for its ability to inhibit both processes after keratoplasty and to promote long-term graft survival (intraperitoneal injections on the day of surgery and 3, 7, and 14 days later). RESULTS: No blood or lymph vessels were detectable immediately after normal-risk transplantation in either donor or host cornea, but hem- and lymphangiogenesis were clearly visible at day 3 after transplantation. Both vessel types reached donor tissue at 1 week after allografting and similarly after syngeneic grafting. Early postoperative trapping of VEGF-A significantly reduced both hem- and lymphangiogenesis and significantly improved long-term graft survival (78% vs. 40%; P < 0.05). CONCLUSIONS: There is concurrent, VEGF-A-dependent hem- and lymphangiogenesis after normal-risk keratoplasty within the preoperatively avascular recipient bed. Inhibition of hem and lymphangiogenesis (afferent and efferent arm of an immune response) after normal-risk corneal transplantation improves long-term graft survival, establishing early postoperative hem- and lymphangiogenesis as novel risk factors for graft rejection even in low-risk eyes. PMID- 15277491 TI - Role of tumor necrosis factor receptor expression in anterior chamber-associated immune deviation (ACAID) and corneal allograft survival. AB - PURPOSE: To determine the role of tumor necrosis factor receptors (TNFRs) in corneal allograft rejection. METHODS: Corneal epithelial and endothelial cells were examined by flow cytometry for the expression of TNFRI and TNFRII and their susceptibility to TNF-alpha-induced apoptosis. Corneal allografts from normal and TNFRI and TNFRII knockout (KO) C57BL/6 mice were transplanted to BALB/c hosts, and the fate of the allografts was monitored. C57BL/6 spleen cells were injected into the anterior chamber (AC) of BALB/c mice to induce anterior chamber associated immune deviation (ACAID) and promote corneal allograft survival. The presence of ACAID suppressor cells in corneal allograft recipients was tested using a local adoptive transfer (LAT) assay. RESULTS: Murine corneal epithelial and endothelial cells expressed TNFRI and TNFRII and were susceptible to TNF alpha-induced apoptosis, yet corneal allografts from either TNFRI or TNFRII donors did not enjoy a lower incidence of rejection or a prolongation in survival time compared to corneal allografts from normal C57BL/6 donors. Moreover, all 31 of the TNFRII KO corneal grafts were rejected by naive BALB/c hosts. Rejection of TNFRII KO corneal grafts occurred even though suppressor cells developed in the hosts and inhibited the expression of delayed-type hypersensitivity to donor alloantigens. CONCLUSIONS: Expression of TNFRII on corneal cells conveys a degree of protection against immune rejection of corneal allografts by a mechanism that is independent of ACAID. Moreover, induction of ACAID before the application of TNFRII KO corneal allografts fails to improve survival and does not replace the TNFRII-dependent protective mechanism. PMID- 15277492 TI - Analysis of the optical quality of intraocular lenses. AB - PURPOSE: To evaluate the optical quality of different intraocular lenses (IOLs). METHODS: An optical test bench and suitable software were used to assist in analysis of the optical Fourier transform (OFT) of a test image and to determine the quality of the lens in terms of spatial frequency response. The OFT was automatically converted, by means of an optical-electronic calibration procedure, into a modulation transfer function (MTF) for each lens. The passband value calculated by computer analysis of the MTF is an objective index of the lens quality. Three randomly acquired samples of 24 different models of foldable IOLs were compared. Statistical analysis was performed with two-way and one-way ANOVA for repeated measurements and with the Ryan-Einot-Gabriel-Welsch multiple F test. RESULTS: The method was demonstrated to be precise and accurate. A large range of passband values was found. Statistically significant differences between the mean passband values for different lenses were found. The lowest passband value (125.60 line pairs [lp]/mm) was measured for the IOL (Lenstec LH3000; Lenstec, Inc., St. Petersburg, FL) and the highest (191.48 lp/mm) for the Acrysof SA30AL (Alcon, Fort Worth, TX). CONCLUSIONS: Different IOLs can transmit different spectra of spatial frequencies. The best frequency response was provided by acrylic IOLs, particularly those with an asymmetrically biconvex profile. This could be due to a reduction of optical degradation provided by this type of profile. A lens with a higher frequency response should determine a better quality of vision once implanted and the frequency response should therefore be considered when choosing the intraocular lens model. PMID- 15277493 TI - Qualitative effect of zonular tension on freshly extracted intact human crystalline lenses: implications for the mechanism of accommodation. AB - PURPOSE: To determine the topographic effects of zonular tension on the anterior surface of the human crystalline lens. METHODS: Real-time topography of the anterior surface of seven fully relaxed, freshly extracted intact, clear, human crystalline lenses aged 3, 17, 45, 54, 54, 56, and 56 years was qualitatively obtained before, during, and after the application of zonular traction. Zonular traction was applied manually either by grasping a group of zonules 180 degrees apart with tying forceps (three lenses), or with micrometers by clamping four portions of the ciliary body that were 90 degrees apart (four lenses). RESULTS: Zonular tension began with the lenses in the fully relaxed, baseline state. As zonular tension was increased across one meridian of all seven lenses, the center of the anterior surface steepened while the periphery of the anterior surface flattened across that meridian of traction. When the tension was reduced across that meridian of traction, the center of the lens flattened while the periphery steepened in that meridian. Four-point zonular traction applied 90 degrees apart produced symmetrical central steepening (four lenses). Reduction of zonular tension across both orthogonal meridians caused symmetrical central flattening. CONCLUSIONS: These observations reveal that when zonular tension is applied to the fully relaxed lens, the center steepens and its periphery flattens in the meridian (or meridians) in which zonular tension is applied. The reverse of this process demonstrates that as tension is reduced, the center of the lens flattens while the periphery steepens either in the meridian of relaxation or symmetrically when zonular tension is released from two orthogonal meridians. These results are opposite to what would have been predicted on the basis of Helmholtz's theory of accommodation. PMID- 15277494 TI - Hepatocyte growth factor induces proliferation of lens epithelial cells through activation of ERK1/2 and JNK/SAPK. AB - PURPOSE: Posterior capsule opacification (PCO) is caused by proliferation and migration of lens epithelial cells (LECs) remaining after cataract surgery. In this study, the effect of HGF in LECs and the signaling pathways that contribute to HGF-induced proliferation were investigated. METHODS: Capsular bags prepared from porcine eyes were maintained in serum-free DMEM. The human lens epithelial B3 cells (HLE B3) and rat lens epithelial explants were cultured in MEM supplemented with 20% FCS and medium 199 with 0.1% BSA, respectively. Cell proliferation was determined by MTT assay, proliferating cell nuclear antigen (PCNA) expression, or flow cytometry. An antisense oligonucleotide was used to inhibit cyclin D1 expression. Activation of the mitogen activated protein kinase (MAPK) and phosphatidylinositol 3-kinase (PI3K) pathways was detected by immunoblot analysis. RESULTS: The proliferation of LECs in a capsular bag culture was significantly inhibited by treatment with the neutralizing antibody for HGF receptor. Stimulation of HLE B3 with hepatocyte growth factor (HGF) activated the MAPKs, ERK, and JNK/SAPK, but not p38. Activation of both ERK and JNK/SAPK was necessary for the HGF-stimulated induction of cyclin D1, which in turn was necessary for the HGF-induced proliferation of LECs. PI3K also participated in the regulation of cyclin D1 expression upstream of ERK and JNK/SAPK. CONCLUSIONS: The data indicate that HGF is a potent growth factor for LECs and may contribute to the development of PCO and suggest that the signaling pathways involved in HGF stimulated proliferation may constitute potential therapeutic targets in the treatment of PCO. PMID- 15277495 TI - Proteomic and sequence analysis of chicken lens crystallins reveals alternate splicing and translational forms of beta B2 and beta A2 crystallins. AB - PURPOSE: To characterize the adult chicken lens proteome using mass spectrometry and two-dimensional gel electrophoresis (2-DE). METHODS: Lens proteins from 10 week old chickens were separated by gel filtration and reversed-phase chromatography, and whole protein masses were measured with electrospray mass spectrometry. Water-soluble lens proteins were separated by 2-DE and identified by tandem mass spectrometry of in-gel digests. RESULTS: Whole protein masses were consistent with all major chicken lens crystallin sequences, except for beta B2 and beta B3. Subsequent cDNA sequencing revealed errors in published sequences translating into 2- and 7-amino-acid differences, respectively, for beta B2 and beta B3, which were in better agreement with the measured masses. Previously uncharacterized forms of beta A2 and beta B2 were observed. The novel form of beta A2 had four fewer amino acids, was more abundant, and resulted from translation at a second start codon. The novel form of beta B2 contained 14 additional amino acids in the interdomain linker and resulted from alternate splicing within intron 4 of the transcript. All examined crystallins, except beta A3, for which data could not be obtained, were N-terminally acetylated, and all beta-crystallins lacked an initial methionine, except for the smaller beta A2 form. In-gel digests identified 29 proteins on the 2-DE map and indicated that truncation occurs within N-terminal extensions of beta-crystallins during lens maturation. CONCLUSIONS: The complementary techniques 2-DE, mass spectrometry, and DNA sequencing were used to provide the most complete description of the adult chicken lens proteome to date and identified alternate forms of beta A2 and beta B2. PMID- 15277496 TI - A homozygous splice mutation in the HSF4 gene is associated with an autosomal recessive congenital cataract. AB - PURPOSE: To map the locus and identify the gene causing autosomal recessive congenital cataracts in a large consanguineous Tunisian family. METHODS: DNA was extracted from blood samples from a large Tunisian family with an autosomal recessive, congenital, total white cataract. A genome-wide scan was performed with microsatellite markers. All exons and the splice sites of the HSF4 gene were sequenced in all members of the Tunisian family and in control individuals. RT PCR was used to detect different transcripts of the HSF4 gene in the human lens. The transcripts were cloned in a TA cloning vector and sequenced. RESULTS: Two point linkage analyses showed linkage to markers on 16q22 with a maximum lod score of 17.78 at theta = 0.01 with D16S3043. Haplotype analysis refined the critical region to a 1.8-cM (4.8-Mb) interval, flanked by D16S3031 and D16S3095. This region contains HSF4, some mutations of which cause the autosomal dominant Marner cataract. Sequencing of HSF4 showed a homozygous mutation in the 5' splice site of intron 12 (c.1327+4A-->G), which causes the skipping of exon 12. A more detailed study of the transcripts resulting from alternative splicing of the HSF4 gene in the lens is also reported, showing the major transcript HSF4b. CONCLUSIONS: This is the first report describing association of an autosomal recessive cataract with the HSF4 locus on 16q21-q22.1 and the first description of HSF4 splice variants in the lens showing that HSF4b is the major transcript. PMID- 15277497 TI - Controlled drug release from an ocular implant: an evaluation using dynamic three dimensional magnetic resonance imaging. AB - PURPOSE: The ability of an episcleral implant at the equator of the eye to deliver drugs to the posterior segment was evaluated, using a sustained-release implant containing gadolinium-DTPA (Gd-DTPA). The movement of this drug surrogate was assessed with magnetic resonance imaging (MRI) in the rabbit eye. The results were compared with a similar implant placed in the vitreous cavity through a scleral incision at the equator. METHODS: Polymer-based implants releasing Gd DTPA were manufactured and placed in the subconjunctival space on the episclera or in the vitreous cavity in live rabbit eyes (in vivo) and in freshly enucleated eyes (ex vivo). Release rates of implants in vitro were also determined. Dynamic three-dimensional MRI was performed using a 4.7-Tesla MRI system for 8 hours. MR images were developed and analyzed on computer. RESULTS: Episcleral implants in vivo delivered a mean total of 2.7 microg Gd-DTPA into the vitreous, representing only 0.12% of the total amount of compound released from the implant in vitro. No Gd-DTPA was detected in the posterior segment of the eye. Ex vivo, the Gd-DTPA concentration in the vitreous was 30 times higher. In vivo eyes with intravitreal implants placed at the equator delivered Gd-DTPA throughout the vitreous cavity and posterior segment. Compartmental analysis of the ocular drug distribution from an episcleral implant showed that the elimination rate constant of Gd-DTPA from the subconjunctival space into the episcleral veins and conjunctival lymphatics was 3-log units higher than the transport rate constant for Gd-DTPA movement into the vitreous. CONCLUSIONS: In vivo, episcleral implants at the equator of the eye did not deliver a significant amount of Gd-DTPA into the vitreous, and no compound was identified in the posterior segment. A 30-fold increase in vitreous Gd-DTPA concentration occurred in the enucleated eyes, suggesting that there are significant barriers to the movement of drugs from the episcleral space into the vitreous in vivo. Dynamic three-dimensional MRI using Gd-DTPA, and possibly other contrast agents, may be useful in understanding the spatial relationships of ocular drug distribution and clearance mechanisms in the eye. PMID- 15277498 TI - H-7 effect on outflow facility after trabecular obstruction following long-term echothiophate treatment in monkeys. AB - PURPOSE: To determine whether H-7 can enhance outflow facility after trabecular meshwork obstruction by extracellular material that accumulates after long-term treatment of monkeys with the cholinesterase inhibitor echothiophate iodide (ECHO). METHODS: Cynomolgus monkeys were treated topically with 150 microg ECHO in one (n = 4 eyes) or both (n = 8 eyes) eyes for up to 48 weeks. Accommodation response to topical pilocarpine was monitored periodically. Outflow facility response to H-7 was measured by two-level constant pressure perfusion on three or four different occasions after intraocular pressure was elevated for 12 to 18 weeks. RESULTS: Long-term treatment with ECHO decreased the accommodative response to pilocarpine and increased intraocular pressure, as has been reported. Baseline outflow facility was decreased by 46% +/- 7% (n = 12, P < 0.001). H-7 partially restored baseline outflow facility measured during subsequent perfusions while ECHO treatment was continued. Concurrent H-7 enhanced outflow facility by 73% +/- 18% (n = 12, P < 0.005) beyond the same-day baseline in ECHO treated eyes. Cessation of ECHO treatment further restored baseline outflow facility, and the outflow facility response to H-7. CONCLUSIONS: H-7 can enhance OF in the presence of trabecular obstruction produced by long-term ECHO treatment. This suggests that H-7 may be useful in treating glaucoma, even in the presence of accumulated plaque material that has been described previously. PMID- 15277499 TI - Temporal changes in gene expression after injury in the rat retina. AB - PURPOSE: The goal of this study was to define the temporal changes in gene expression after retinal injury and to relate these changes to the inflammatory and reactive response. A specific emphasis was placed on the tetraspanin family of proteins and their relationship with markers of reactive gliosis. METHODS: Retinal tears were induced in adult rats by scraping the retina with a needle. After different survival times (4 hours, and 1, 3, 7, and 30 days), the retinas were removed, and mRNA was isolated, prepared, and hybridized to the Affymatrix RG-U34A microarray (Santa Clara, CA). Microarray results were confirmed by using RT-PCR and correlation to protein levels was determined. RESULTS: Of the 8750 genes analyzed, approximately 393 (4.5%) were differentially expressed. Clustering analysis revealed three major profiles: (1) The early response was characterized by the upregulation of transcription factors; (2) the delayed response included a high percentage of genes related to cell cycle and cell death; and (3) the late, sustained profile clustered a significant number of genes involved in retinal gliosis. The late, sustained cluster also contained the upregulated crystallin genes. The tetraspanins Cd9, Cd81, and Cd82 were also associated with the late, sustained response. CONCLUSIONS: The use of microarray technology enables definition of complex genetic changes underlying distinct phases of the cellular response to retinal injury. The early response clusters genes associate with the transcriptional regulation of the wound-healing process and cell death. Most of the genes in the late, sustained response appear to be associated with reactive gliosis. PMID- 15277500 TI - Fundus autofluorescence in patients with leber congenital amaurosis. AB - PURPOSE: Fundus autofluorescence (FAF), as an index of lipofuscin accumulation in the retinal pigment epithelium (RPE), provides indirect information on the level of metabolic activity of the RPE and thus the integrity of the RPE/photoreceptor complex. To investigate whether the photoreceptor/RPE complex is still viable in patients with Leber congenital amaurosis (LCA), FAF imaging was performed. METHODS: Three patients with LCA (patients A, B, and C; ages, 24, 15, and 37 years, respectively) were enrolled and one patient with RP with preserved visual acuity (age, 28 years) was included as a control. The diagnosis was based on history, visual function, and Ganzfeld electroretinography (International Society for Clinical Electrophysiology of Vision [ISCEV] standard). FAF was recorded with a confocal scanning laser ophthalmoscope (cSLO; Heidelberg Retina Angiograph; Heidelberg Engineering, Heidelberg, Germany). RESULTS: All patients with LCA had vision reduced to perception of light and had undetectable ERGs. FAF was normal in patient A. In patient B, there was a parafoveal ring of mildly increased FAF. The midperiphery showed mildly decreased FAF. Patient C showed a parafoveal ring of moderately increased FAF. FAF was moderately decreased along the arcades and the midperiphery. The patient with RP showed a parafoveal ring of moderately increased FAF and severely decreased FAF eccentric to the macula including the periphery. CONCLUSIONS: The FAF findings in these patients with LCA suggest that there is continuous metabolic demand from the photoreceptors and that the RPE/photoreceptor complex is, at least in part, anatomically intact, but the photoreceptors have lost function. These findings may have implications for future treatment. It is notable that more than 20 years of severe visual impairment associated with LCA can be associated with normal FAF, indicating that photoreceptor function may be rescuable. PMID- 15277501 TI - Neuroprotection of photoreceptors by minocycline in light-induced retinal degeneration. AB - PURPOSE: Microglial cells have been found to play pivotal roles in various neuronal degenerative diseases such as Parkinson's and Alzheimer's diseases. Minocycline, a microglial inhibitor, has recently been shown to be neuroprotective in various models of cerebral ischemia and degenerative diseases of the brain. This study was conducted to evaluate the neuroprotective effect of minocycline and the role of microglia in light-induced retinal degeneration. METHODS: BALB/cJ mice were exposed to intense green light for 3 hours and observed during 1, 3, or 7 days of dark recovery. The animals received intraperitoneal injections of minocycline or vehicle 1 day before exposure to light for 2, 4, or 8 days, depending on the periods of survival. Morphologic, morphometric, immunohistochemical, and electrophysiological studies were performed to evaluate the efficacy of minocycline in the amelioration of light induced retinal degeneration and the possible involvement of microglial cells. RESULTS: Minocycline treatment provided marked amelioration in the loss of photoreceptors in light-induced retinal degeneration, as evidenced by morphologic, morphometric, and electrophysiologic criteria. Morphologically, the minocycline-treated group showed markedly better preservation of the outer retina after exposure to light. Morphometrically, at 7 days after exposure to light, in the minocycline-treated animals, 89.1% of the normal-appearing photoreceptor nuclei remained, but in the retinas of the vehicle-control group only 38.0% of these nuclei remained. This difference was statistically significant (P < 0.001). At 7 days after exposure to light electroretinography (ERG) showed that minocycline significantly preserved the amplitudes of dark-adapted a- and b-wave and light-adapted b-wave, which were all significantly reduced after exposure to light. Concomitant with this protective effect, at 3 days after exposure to light, the CD11b(+) microglial cells in the outer nuclear layer (ONL) and subretinal space in the minocycline-treated group were significantly decreased (by 63.5%) when compared with those in the light-exposed, vehicle-treated control group (P < 0.01). CONCLUSIONS: Minocycline is neuroprotective against light induced loss of photoreceptors, possibly through the inhibition of retinal microglial activation. PMID- 15277502 TI - Expression of apoptosis markers in the retinas of human subjects with diabetes. AB - PURPOSE: To investigate the expression of the apoptotic mediators in the retinas from human subjects with diabetes mellitus. METHODS: Ten donor eyes from five subjects with diabetes mellitus, and eight eyes from four nondiabetic subjects without known ocular disease serving as control subjects were examined. Immunohistochemical techniques were used with antibodies directed against glial fibrillary acidic protein (GFAP), caspase-3, Fas, Fas ligand (FasL), Bax, Bcl-2, survivin, p53, extracellular signal-regulated kinases (ERK1/2), and p38. RESULTS: In retinas from all subjects without diabetes, weak Bcl-2 immunoreactivity was confined to GFAP-positive glial cells in the nerve fiber layer. Weak immunoreactivity for ERK1/2 was noted in a few nuclei in the inner nuclear layer and in a few Muller cell processes. Cytoplasmic immunostaining for survivin was noted in the retinal pigment epithelial cells. There was no immunoreactivity for the other antibodies tested. All diabetic retinas showed cytoplasmic immunoreactivity for caspase-3, Fas, and Bax in ganglion cells. FasL immunoreactivity was detected in GFAP-positive cells. Upregulation of Bcl-2 immunoreactivity was noted in GFAP-positive cells in nerve fiber and ganglion cell layers, and Bcl-2 induction was noted in Muller cell processes. Strong immunoreactivity for ERK1/2 was observed in many nuclei in the inner nuclear layer in GFAP-positive cells in the nerve fiber and ganglion cell layers and numerous Muller cell processes. Survivin immunoreactivity was not altered in the diabetic retinas. There was no immunoreactivity for p53 and p38. CONCLUSIONS: Ganglion cells in diabetic retinas express several proapoptosis molecules, suggesting that these cells are the most vulnerable population. Glial cells in diabetic retinas are activated and express several antiapoptosis molecules in addition to the cytotoxic effector molecule FasL, suggesting a possible role of glial cells in induction of apoptosis in ganglion cells. PMID- 15277503 TI - Severe retinal degeneration associated with disruption of semaphorin 4A. AB - PURPOSE: Semaphorin 4A (Sema4A) is a member of the transmembrane class 4 family of semaphorins. It has recently been shown to participate in cell-cell communication in the immune system. High levels of sema4A are also present in brain and eye, but its function in the central nervous system has not been studied. To investigate the function of Sema4A, we generated mice deficient in this transmembrane signaling molecule. METHODS: An embryonic stem (ES) cell clone with a retroviral gene-trap insertion in the sema4A gene was used to generate mice lacking this transmembrane semaphorin. Fundus photography, fluorescein angiography, and electroretinography were used to evaluate retinal anatomy and physiology in mice lacking Sema4A. Electron microscopy and immunohistochemistry with cell-type-specific markers were used to characterize retinal development. In situ hybridization with sema4A-specific riboprobes was used to localize expression of this gene in the developing and adult eye. RESULTS: Fundus photography performed at 14 weeks of age revealed severe retinal degeneration, attenuated retinal vessels, and depigmentation in mice lacking Sema4A. At this age, the outer nuclear layer was reduced to a single row of photoreceptor cells, and the outer plexiform layer was thin and disorganized. Disruption of Sema4A also compromised the physiological function of both rod and cone photoreceptors. Developmental studies in Sema4A-deficient mice revealed abnormal morphology of photoreceptor outer segments during the time at which they establish contacts with apical microvilli of the retinal pigment epithelium (RPE). Sema4A is expressed in the inner retina and RPE during the time at which photoreceptor outer segments elongate. CONCLUSIONS: These findings identify a previously unknown function of Sema4A in the developing visual system and provide a useful model for understanding cell-cell interactions that occur between photoreceptors and the RPE. PMID- 15277504 TI - Safety testing of indocyanine green and trypan blue using retinal pigment epithelium and glial cell cultures. AB - PURPOSE: Indocyanine green (ICG) and trypan blue have been advocated as vital stains for use during macular surgery. The safety of these agents was tested using a cell culture model. METHODS: Human retinal pigment epithelium (RPE) and Muller cell lines were exposed to ICG over a range of concentrations up to 0.5%, and trypan blue up to 0.2%. Cells were exposed to each dye for 5, 15, or 30 minutes, rinsed, and incubated 24 hours. Cell viability was measured using a mitochondrial dehydrogenase-assay and fluorescent live-dead probe. Experiments were repeated using 0.5% and 1% ICG and 0.06% and 0.12% trypan blue, with follow up at 0, 1, 5, and 15 days. ICG experiments were repeated in the presence of illumination from a xenon light-source channeled through a surgical endolight, and using reduced osmolarity solutions of 0.1%, 0.5%, and 1% (185 vs. 275 mOsM). RESULTS: There was no clear relationship between cell viability and the concentration of the agent or duration of follow-up, except in RPE cells exposed to 1% ICG. These showed a linear (R(2) 0.9952) decline in viability with time, with a significant reduction by day 15 (P = 0.016). RPE cells exposed to ICG and illumination were not significantly different from the negative control, but when illumination was combined with low osmolarity, viability was reduced (P = 0.0016). ICG and illumination reduced Muller cell viability (P < 0.0001 for both 185 and 275 mOsM). Muller cells incubated with 185 mOsM 1% ICG showed a significant reduction in viability (P < 0.0001) not seen with the 185 mOsM 0.5% or 0.1% solutions or in the low-osmolarity RPE groups. CONCLUSIONS: The combination of exposure to 0.5% ICG and the newer endoillumination light-sources can damage cultured Muller cells. Although the preparations of ICG most commonly used clinically did not produce significant damage, relatively small changes in ICG osmolarity and concentration did. This suggests that safety margins are not large. Trypan blue is safe in a cell culture model. PMID- 15277505 TI - Basement membrane-dependent modification of phenotype and gene expression in human retinal pigment epithelial ARPE-19 cells. AB - PURPOSE: To use porcine lens capsule (PLC) as basement membrane for ARPE-19 cells and to characterize its effects on cell differentiation and gene expression. METHODS: Postconfluent cultures of ARPE-19 cells were established on either porous polyester filters or PLC membranes and characterized by electron microscopy, immunocytochemistry, and transepithelial electrical resistance measurements. Metabolic activity was assessed by measuring phagocytosis of rod outer segments. mRNA populations of ARPE-19 cells grown on polyester and PLC membranes were compared by suppressive subtractive hybridization. Differentially regulated messages were subsequently identified by DNA sequencing and their altered expression confirmed by Northern or virtual Northern blot analysis. RESULTS: Culture of ARPE-19 cells on PLC membrane induced the formation of apical microvilli and the ability to phagocytose rod outer segments. These culture conditions also led to enhanced junctional distribution of ZO-1 and occludin, the formation of polarized membrane domains, and a significant increase in transepithelial resistance. Gene expression was significantly altered by growth on PLC membranes and 29 differentially expressed transcripts were identified. CONCLUSIONS: Culture of ARPE-19 cells on PLC membranes resulted in a more differentiated phenotype and in expression of a specific set of transcripts encoding protein products that may affect epithelial differentiation, polarity and survival. PMID- 15277506 TI - Characterization of smooth muscle cell and pericyte differentiation in the rat retina in vivo. AB - PURPOSE: To identify and apply a range of suitable mural cell markers and undertake an in vivo characterization of pericyte and smooth muscle cell (SMC) differentiation in the developing rat retina. METHODS: Pericyte and SMC differentiation was characterized by immunohistochemistry with antibodies to NG2, desmin, alpha-smooth muscle actin (SMA), calponin, and caldesmon. RESULTS: Immature mural precursor cells (MPCs) were scattered throughout the primitive capillary plexus in the rat retina at embryonic day (E)20. The postnatal differentiation of pericytes and arteriolar and venous SMCs followed with distinct intermediate phenotypes. SMC differentiation coincided with selection of major vessels from the primordial capillary bed. Maturation of radial arteriolar SMCs was indicated by the expression of calponin and caldesmon, proteins that play a role in the regulation of SMC contraction. The mere presence of immature mural cells did not confer vessel stability; rather vessel stability in the developing rat retina coincided with caldesmon and calponin expression in arteriolar SMCs. CONCLUSIONS: This normative data and the identification of suitable in vivo markers of pericytes and SMCs will allow meaningful interpretation of the changes in these cell types. When examining the role of mural cells in developmental and pathologic vascularization, the results show that there is a need to use multiple-marker immunohistochemistry because of significant mural cell heterogeneity. The observation that the expression of caldesmon and calponin in arteriolar SMCs coincides with resistance to hyperoxia in the developing rat retina, lead us to suggest that maturation of SMCs and their consequent ability to regulate blood flow may play a key role in vessel stabilization. PMID- 15277507 TI - Expression of photoreceptor-specific nuclear receptor NR2E3 in rod photoreceptors of fetal human retina. AB - PURPOSE: To study the physiological function of NR2E3 and possible molecular mechanisms underlying enhanced short-wavelength cone syndrome (ESCS) pathogenesis in developing human retina, and to compare its expression to that of Neural Retina Leucine zipper (NRL), a transcription factor essential for rod differentiation. METHODS: Expression of NR2E3, a photoreceptor-specific orphan nuclear receptor, was examined in human retinas between fetal weeks (Fwk) 9 to 22 by reverse transcription-polymerase chain reaction (RT-PCR) and in situ hybridization. Both NR2E3 and NRL expression patterns were followed by immunocytochemistry. The human retina develops in a central to peripheral pattern, in which a protein may take weeks to be expressed throughout the entire retina. This allowed a detailed temporal analysis of NR2E3 and NRL expression. RESULTS: NR2E3 expression was detected shortly after the appearance of NRL in putative immature rods on the foveal edge at Fwk 11.7. Expression of both markers was maintained in rod opsin expressing fetal photoreceptors. NR2E3 expression was not detected in either long/medium- or short-wavelength cones. Its absence from cones was also supported by the position of labeled nuclei deep in the outer nuclear layer, and by the absence of NR2E3 from the fovea. CONCLUSIONS: A role for NR2E3 in the rod developmental pathway is suggested. The closely related expression patterns of NRL and NR2E3 supported an interactive function, where both transcription factors determine the rod fate and suppress immature rods from adopting the S-cone fate. PMID- 15277508 TI - Cultured embryonic retina systems as a model for the study of underlying mechanisms of Toxoplasma gondii infection. AB - PURPOSE: Toxoplasma gondii, the most common cause of retinochoroiditis in humans, is an obligate intracellular protozoan parasite that depends on te host cell's microenvironment to proliferate. Because congenital infection is associated with a higher risk of ocular involvement than a postnatally acquired infection, this study was conducted to investigate the ability of Toxoplasma gondii to infect retinal tissue during development, when cellular environmental changes normally occur. METHODS: Retinas from 5- to 9-day-old chick embryos were used. Stationary cultures were prepared in 24-well cell culture dishes and maintained at 37 degrees C in DMEM plus 5% fetal bovine serum for 2 to 6 days. Then the wells were infected with 4 x 10(5) tachyzoites. Retina explants and aggregate cell cultures were maintained in DMEM under rotation at 37 degrees C. T. gondii proliferation was measured using [(3)H]-thymidine incorporation after 72 hours. Ornithine and arginine decarboxylase (ODC and ADC) activities were determined by measuring CO(2) production from [1-(14)C]-ornithine and [1-(14)C]-arginine, respectively. RESULTS: The proliferation of tachyzoites was high in dense, stationary cultures expressing elevated ODC and ADC activity. The addition of ODC or ADC inhibitors reduced T. gondii proliferation by approximately 20% to 40%. As for cultured retina cells, retina explants also allowed T. gondii proliferation whenever ODC activity was high. CONCLUSIONS: The data suggest a direct correlation between retinal polyamine biosynthesis and the proliferation of T. gondii, in agreement with the observation that individuals infected congenitally have a greater risk of development of toxoplasmic retinochoroiditis. PMID- 15277509 TI - RPE cells internalize low-density lipoprotein (LDL) and oxidized LDL (oxLDL) in large quantities in vitro and in vivo. AB - PURPOSE: To determine whether plasma low-density lipoprotein (LDL) could be internalized by the RPE and which receptors may be involved. A secondary objective was to determine whether ARPE19 cells could be used as a model to investigate cholesterol processing in the RPE. METHODS: Commercially available human LDL was labeled with rhodamine or AlexaFluor 568. Immunofluorescence was performed using commercially available antibodies to LDL-R, CD36, and LOX-1. Cells and tissues were imaged using epifluorescence and confocal fluorescence microscopy. Immunoblot analysis and RT-PCR were performed using published techniques. RESULTS: Intravenously injected rhodamine-labeled LDL (rhoLDL) was detected in the rat RPE by fluorescence confocal microscopy 24 hours after injection. The rhoLDL was present in some areas and absent in others. Cultured ARPE19 cells were also found to internalize LDL and oxidized LDL (oxLDL) readily. Using AlexaFluor 568-labeled LDL we determined that the average cultured RPE cell could internalize approximately 12 to 16 pg of LDL and oxLDL in 24 hours. Immunoblots readily detected the presence of CD36 and LDL-R in the cultured RPE cells but not LOX-1, whereas RT-PCR detected mRNA for all three receptors. Dual labeling experiments using AlexaFluor 568-labeled LDL and AlexaFluor 488 for the immunolocalization of the receptors showed colocalization of LDL-R with the internalized LDL and CD36 with oxLDL particles. CONCLUSIONS: Plasma LDL readily enters the RPE through the choriocapillaris but is not found homogeneously throughout the retina. This may suggest some form of regulation to the permeability of the fenestrated choroidal endothelial cell junctions. ARPE19 cells are a good model for studying the internalization mechanisms of LDL and oxLDL in vitro. LDL may be used as a vector to carry hydrophobic molecules into the RPE. PMID- 15277510 TI - Cytotoxicity of oxidized low-density lipoprotein in cultured RPE cells is dependent on the formation of 7-ketocholesterol. AB - PURPOSE: To determine which components present in oxidized LDL are responsible for the cytotoxicity associated with its internalization by culture ARPE19 cells. METHODS: ARPE19 cells were grown in 24-well and 96-well plates. Cell viability was measured by MTT and/or adenosine triphosphate (ATP) content. LDL was oxidized with Cu(+2) and oxysterol content analyzed by a novel HPLC method. RESULTS: OxLDL showed increased cytotoxicity with prolonged oxidation. Analysis of the oxLDL showed a predominance of the 7-oxygenated products, 7 alpha-hydroxycholesterol (7 alpha HCh), 7 beta-hydroxycholesterol (7 beta HCh), and 7-ketocholesterol (7kCh). Addition of these oxysterols to the ARPE19 cell in free form indicated that 7kCh is the most cytotoxic of the oxysterols but at physiologically unrealistic concentrations. Partitioning of individual oxysterols into nonoxidized LDL at concentrations similar to those found in the oxLDL also indicated that 7kCh is the most cytotoxic of the oxysterols. Transition metals are tightly bound by LDL and play an important role in the oxidation of LDL, but do not seem to enhance its cytotoxicity directly. CONCLUSIONS: Prolonged oxidation of LDL increases the levels of 7kCh due to further oxidation of 7 alpha HCh and 7 beta HCh. The formation of 7KCh seems to be responsible for most of the cytotoxicity associated with oxLDL internalization in ARPE19 cells. PMID- 15277511 TI - IGF-1-induced VEGF and IGFBP-3 secretion correlates with increased HIF-1 alpha expression and activity in retinal pigment epithelial cell line D407. AB - PURPOSE: To examine insulin-like growth factor (IGF)-1 stimulation of expression of hypoxia inducible factor (HIF)-1 alpha and secretion of vascular endothelial growth factor (VEGF) and IGF binding protein (IGFBP)-3 in human retinal pigment epithelial (RPE) cell line D407. METHODS: D407 cells cultured in dishes or Transwell inserts were treated with cobalt chloride or varying doses of IGF-1. Whole cell lysates were assayed by immunoblot for HIF-1 alpha expression, whereas conditioned medium was TCA precipitated and assayed by immunoblot for VEGF and ligand blot for IGFBP-3. Cells grown on coverslips were similarly treated and probed with antibodies to HIF-1 alpha, VEGF, and IGFBP-3, and visualized by epifluorescence microscopy. Cells grown on Transwell inserts were probed with antibodies to the Na(+)/K(+)-ATPase alpha-1 subunit and either the alpha or beta subunits of the IGF-1 receptor and visualized in Z-section using confocal microscopy. RESULTS: Immunoblot analysis of whole cell lysates from IGF-1-treated D407 cells revealed the upregulation of HIF-1 alpha protein. Epifluorescence microscopy demonstrated a positive correlation between HIF-1 alpha expression and nuclear localization, VEGF and IGFBP-3 synthesis and export, and IGF-1 action. Western and ligand blot analyses of RPE cell-conditioned medium indicated that IGF-1 induced increases in VEGF and IGFBP-3 secretion. Cells grown on Transwell inserts exhibited constitutive apical secretion of VEGF and IGFBP-3, which increased on apical or basolateral treatment with IGF-1. Confocal analysis of Transwell-cultured D407 cells confirmed the apical localization of the Na(+)/K(+) ATPase alpha-1 subunit, characteristic of polarized RPE, with IGF-1 receptor alpha and beta subunits exhibiting a nonpolarized distribution. CONCLUSIONS: IGF 1 stimulates increased HIF-1 alpha expression as well as VEGF and IGFBP-3 secretion in D407 cells. Similar to their in vivo counterparts, D407 cells maintain reversed epithelial polarity. Apical secretion of VEGF and IGFBP-3 increases in response to either apical or basolateral IGF-1 stimulation consistent with the nonpolarized distribution of IGF-1 receptors. PMID- 15277512 TI - Insulin-like growth factor binding proteins modulate Muller cell responses to insulin-like growth factors. AB - PURPOSE: Muller cells are consistently identified in diabetic fibrocontractive ocular tissues and, in response to insulin-like growth factor I, generate tractional forces of the type that cause retinal detachment. Recent studies suggest that diabetes-associated increases in vitreous insulin-like growth factor activity cannot be attributed to simple increases in concentration alone, suggesting that more complex biochemical changes in vitreous growth factor control mechanisms are involved. The goal of this study was to evaluate the contributions of vitreous insulin-like growth factor-binding proteins (IGFBPs) toward control of growth factor activity. METHODS: Native and recombinant IGFBPs effects were evaluated on IGF-I- and -II-stimulated Muller cells in tissue culture assays that involved cell incubation on three-dimensional collagen gels and that monitored progressive matrix condensation. IGFBP degradation by Muller cell-secreted proteases was assessed in Western ligand blots, and direct stimulatory effects were evaluated by incubating cells with IGFBPs alone. RESULTS: IGFBP direct stimulatory effects on Muller cells were significant, but relatively modest, and IGFBP modulation through Muller cell-secreted proteases was undetectable. In contrast, IGFBP inhibitory effects on IGF-I and -II were highly variable and, in some cases, profound. IGFBP-3 effectively inhibited IGF-I and -II stimulation with detectable effects at concentrations equimolar to the growth factor. IGFBP-1, -2, -4, and -5 were of intermediate effectiveness as inhibitors, 3- to 11-fold less active than IGFBP-3. IGFBP-6 had virtually no inhibitory effects on IGF-I, but was moderately effective against IGF-II. CONCLUSIONS: IGFBP effects on IGF-I- and -II-stimulated Muller cells are primarily inhibitory with only modest direct stimulatory effects of limited physiologic relevance. IGFBP-2 and -3, the major binding proteins identified in vitreous, most likely function as the vitreous growth factor sink and control ligand activity through sequestration. PMID- 15277513 TI - Association of ocular dominance and anisometropic myopia. AB - PURPOSE: To determine the association between ocular dominance and degree of myopia in patients with anisometropia. METHODS: Fifty-five subjects with anisometropic myopia were recruited. None of them had amblyopia. Refractive error and axial length were measured in each subject. Ocular dominance was determined using the hole-in-the-card test and convergence near-point test. RESULTS: There was a threshold level of anisometropia (1.75 D) beyond which the dominant eye was always more myopic than the nondominant eye. Of the 33 subjects with anisometropia of < or =1.75 D, the dominant eye was more myopic in 17 (51.5%) subjects. Dominant eyes, determined by the hole-in-the-card test, had a significantly greater myopic spherical equivalent (-5.27 +/- 2.45 D) than nondominant eyes (-3.94 +/- 3.10 D; P < 0.001). Dominant eyes also had a longer axial length than nondominant eyes (25.15 +/- 0.96 mm vs. 24.69 +/- 1.17 mm, respectively; P < 0.001). The difference was more evident in those subjects with higher anisometropia (>1.75 D), but was not significant in those with lower anisometropia (< or =1.75 D). Similar results were obtained using the convergence near-point test. CONCLUSIONS: The present study shows that the dominant eye has a greater degree of myopia than the nondominant eye in subjects with anisometropic myopia. Taking ocular dominance into account in the design of randomized clinical trails to assess the efficacy of myopia interventions may provide useful information. PMID- 15277514 TI - Red-green chromatic mechanisms in normal aging and glaucomatous observers. AB - PURPOSE: This study was designed to determine whether normal aging and glaucoma are associated with red-green (R/G) chromatic processing abnormalities, a function that is primarily performed by the parvocellular visual pathway. METHODS: Chromatic processing mechanisms were examined in 98 glaucomatous observers (between the ages of 49 and 93 years; mean age, 70.8 +/- 9.4 [SD]) and 67 normal observers (between the ages of 49 and 88; mean age, 70.6 +/- 10.6 years) with the use of the minimum-motion and motion-nulling paradigms. Phakic glaucomatous (n = 60; mean age, 68.7 +/- 8.9 years) and normal (n = 32; mean age, 69.8 +/- 10.6 years) and pseudophakic glaucomatous (n = 38; mean age, 74 +/- 9.4 years) and normal (n = 35; mean age, 71.4 +/- 10.6 years) subjects were tested to evaluate the effects of lenticular aging on color perception. RESULTS: Phakic observers (normal or glaucomatous) displayed significantly different minimum motion values than did both their younger counterparts and all the pseudophakic subjects. These results suggest that normal aging with the presence of a natural lens is accompanied by a significant decrease in green-light sensitivity, an effect that is not exacerbated by glaucoma and is primarily related to optical factors. The data also revealed no differences in color motion perception between groups, indicating that the higher cortical mechanisms of the parvocellular pathway implicated in the analysis of information about the middle and long wavelengths of the visible spectrum are not selectively affected by the disease process and normal aging. CONCLUSIONS: Normal aging and glaucoma do not produce significant R/G chromatic processing deficits at retinal and postretinal levels when optical factors are excluded. The authors propose the hypothesis that glaucoma-related effects on motion perception and blue-on-yellow perimetry should be viewed as evidence of loss of ganglion cells that necessitates integration of information over larger retinal areas and more receptor cells than in the R/G chromatic system. Ganglion cells with large receptive fields involve more neural connections and are less numerous than those that respond to R/G information. The functional consequence of this could be that the loss of a single ganglion cell with a larger receptive field would have a greater impact on visual function than the loss of a ganglion cell with a smaller receptive field, such as the ones that process R/G information. The authors believe that glaucoma-induced functional loss is best viewed as related to receptive field structure and function rather than to anatomic cell-type damage. PMID- 15277515 TI - The Duncker illusion: intersubject variability, brief exposure, and the role of eye movements in its generation. AB - PURPOSE: The Duncker illusion, also known as induced motion, is the illusory component of an object's motion that results from background movement. The origins of this robust phenomenon are still subject to debate. The goal of this study was to examine the role eye movements play in its generation. METHODS: The Duncker illusion was generated by rear-projecting an LED target and a random-dot background onto a semitranslucent screen. Each moved under independent control of mirror galvanometers. The background was either stationary or moved vertically, whereas the target moved horizontally with various degrees of a vertical component. Using a two-alternative, forced-choice cancellation paradigm involving multiple interleaved staircases, seven human subjects quantified the illusion under a variety of conditions. These conditions included varying the exposure time from 50 to 200 ms. RESULTS: The strength of the illusion did not decrease, even when exposures too brief to generate eye movement were used. Four of the subjects showed no difference between long and short exposures (P > 0.12). In two the illusion was stronger for short exposures (P < 0.05), and in one the illusion was not quantifiable. There was little intrasubject variability across trial types, yet there was a large intersubject variability in the strength of the illusion. CONCLUSIONS: The illusion is robust in that it is perceived under a wide variety of conditions by all observers. By varying the parameters, it was possible to demonstrate that eye movements do not play a role in generating the illusion and that this robust illusion is most likely caused by the visual system attributing relative motion between target and background to target motion and ignoring any true background movement. The mechanism by which this occurs varies in degree between individuals. PMID- 15277516 TI - Intranuclear trafficking: organization and assembly of regulatory machinery for combinatorial biological control. AB - The molecular logistics of nuclear regulatory processes necessitate temporal and spatial regulation of protein-protein and protein-DNA interactions in response to physiological cues. Biochemical, in situ, and in vivo genetic evidence demonstrates the requirement for intranuclear localization of regulatory complexes that functionally couple cellular responses to signals that mediate combinatorial control of gene expression. We have summarized evidence that subnuclear targeting of transcription factors mechanistically links gene expression with architectural organization and assembly of nuclear regulatory machinery for biological control. The compromised intranuclear targeting of regulatory proteins under pathological conditions provides options for the diagnosis and treatment of disease. PMID- 15277517 TI - Antizyme targets cyclin D1 for degradation. A novel mechanism for cell growth repression. AB - Overproduction of the ornithine decarboxylase (ODC) regulatory protein ODC antizyme has been shown to correlate with cell growth inhibition in a variety of different cell types. Although the exact mechanism of this growth inhibition is not known, it has been attributed to the effect of antizyme on polyamine metabolism. Antizyme binds directly to ODC, targeting ODC for ubiquitin independent degradation by the 26 S proteasome. We now show that antizyme induction also leads to degradation of the cell cycle regulatory protein cyclin D1. We demonstrate that antizyme is capable of specific, noncovalent association with cyclin D1 and that this interaction accelerates cyclin D1 degradation in vitro in the presence of only antizyme, cyclin D1, purified 26 S proteasomes, and ATP. In vivo, antizyme up-regulation induced either by the polyamine spermine or by antizyme overexpression causes reduction of intracellular cyclin D1 levels. The antizyme-mediated pathway for cyclin D1 degradation is independent of the previously characterized phosphorylation- and ubiquitination-dependent pathway, because antizyme up-regulation induces the degradation of a cyclin D1 mutant (T286A) that abrogates its ubiquitination. We propose that antizyme-mediated degradation of cyclin D1 by the proteasome may provide an explanation for the repression of cell growth following antizyme up-regulation. PMID- 15277518 TI - Inhibition of SNAP-25 phosphorylation at Ser187 is involved in chronic morphine induced down-regulation of SNARE complex formation. AB - Opiate abuse has been shown to cause adaptive changes in presynaptic release and protein phosphorylation-mediated synaptic plasticity, but the underlying mechanisms remain unclear. Neuronal SNARE proteins serve as important regulatory molecules underlying neural plasticity in view of their major role in the process of neurotransmitter release. In the present study, the expression of SNAP-25, a t SNARE protein essential for vesicle release, was found to be dramatically regulated in hippocampus after chronic morphine treatment, which was visualized with two-dimensional gel electrophoresis. The spots of SNAP-25 in the gel were shifted along the dimension of isoelectric point, indicating a likely change of the post-transcriptional modification. Immunoblotting analysis with specific antibody to Ser187, a protein kinase C (PKC) phosphorylation site of SNAP-25, revealed that the specific phosphorylation was correspondingly decreased, which was correlated with morphine-induced inhibition of PKC activity. Moreover, the level of ternary complex of SNARE proteins in either synaptosomes or PC12 cells was significantly reduced after chronic morphine treatment. This suggests a causal relationship between the inhibition of PKC-dependent SNAP-25 phosphorylation and the down-regulation of SNARE complex formation after chronic morphine treatment. Further analysis of SNARE complex formed by transfection of the wild-type or Ser187 mutants of SNAP-25 showed that only wild-type-formed complex was inhibited by morphine treatment. Thus, these results indicate that chronic morphine treatment inhibits phosphorylation of SNAP-25 at Ser187 and leads to a down-regulation of SNARE complex formation, which presents a potential molecular mechanism for the alteration of exocytotic process and neural plasticity during opiate abuse. PMID- 15277519 TI - Coordination of peroxisomal beta-oxidation and fatty acid elongation in HepG2 cells. AB - A major product of mitochondrial and peroxisomal beta-oxidation is acetyl-CoA, which is essential for multiple cellular processes. The relative role of peroxisomal beta-oxidation of long chain fatty acids and the fate of its oxidation products are poorly understood and are the subjects of our research. In this report we describe a study of beta-oxidation of palmitate and stearate using HepG2 cells cultured in the presence of multiple concentrations of [U (13)C(18)]stearate or [U-(13)C(16)] palmitate. Using mass isotopomer analysis we determined the enrichments of acetyl-CoA used in de novo lipogenesis (cytosolic pool), in the tricarboxylic acid cycle (glutamate pool), and in chain elongation of stearate (peroxisomal pool). Cells treated with 0.1 mm [U-(13)C(18)]stearate had markedly disparate acetyl-CoA enrichments (1.1% cytosolic, 1.1% glutamate, 10.7% peroxisomal) with increased absolute levels of C20:0, C22:0, and C24:0. However, cells treated with 0.1 mm [U-(13)C(16)]palmitate had a lower peroxisomal enrichment (1.8% cytosolic, 1.6% glutamate, and 1.1% peroxisomal). At higher fatty acid concentrations, acetyl-CoA enrichments in these compartments were proportionally increased. Chain shortening and elongation was determined using spectral analysis. Chain shortening of stearate in peroxisomes generates acetyl CoA, which is subsequently used in the chain elongation of a second stearate molecule to form very long chain fatty acids. Chain elongation of palmitate to stearate appeared to occur in a different compartment. Our results suggest that 1) chain elongation activity is a useful and novel probe for peroxisomal beta oxidation and 2) chain shortening contributes a substantial fraction of the acetyl-CoA used for fatty acid elongation in HepG2 cells. PMID- 15277520 TI - The propeptide domain of lysyl oxidase induces phenotypic reversion of ras transformed cells. AB - Lysyl oxidase is an extracellular enzyme critical for the normal biosynthesis of collagens and elastin. In addition, lysyl oxidase reverts ras-mediated transformation, and lysyl oxidase expression is down-regulated in human cancers. Since suramin inhibits growth factor signaling pathways and induces lysyl oxidase in ras-transformed NIH3T3 cells (RS485 cells), we sought to investigate the effects of suramin on the phenotype of transformed cells and the role of lysyl oxidase in mediating these effects. Suramin treatment resulted in a more normal phenotype as judged by growth rate, cell cycle parameters, and morphology. beta aminopropionitrile, the selective inhibitor of lysyl oxidase enzyme activity, was remarkably unable to block suramin-induced reversion. By contrast, ectopic antisense lysyl oxidase demonstrated that lysyl oxidase gene expression mediated phenotypic reversion. Since lysyl oxidase is synthesized as a 50 kDa precursor and processed to a 30 kDa active enzyme and 18 kDa propeptide, the effects of these two products on the transformed phenotype of RS485 cells were then directly assessed in the absence of suramin. Here we report, for the first time, that the lysyl oxidase propeptide, and not the lysyl oxidase enzyme, inhibits ras dependent transformation as determined by effects on cell proliferation assays, growth in soft agar, and Akt-dependent induction of NF-kappaB activity. Thus, the lysyl oxidase propeptide, which is released during extracellular proteolytic processing of pro-lysyl oxidase, functions to inhibit ras-dependent cell transformation. PMID- 15277521 TI - SUMO modification of septin-interacting proteins in Candida albicans. AB - The initiation of bud and hyphal growth in the opportunistic fungal pathogen Candida albicans both involve polarized morphogenesis. However, there are many differences including the function of the septin proteins, a family of proteins involved in membrane organization in a wide range of organisms. Septins form a characteristic ring on the inner surface of the plasma membrane at the bud neck, whereas the septins are diffusely localized across emerging hyphal tips. In addition, septin rings are maintained at sites of septum formation in hyphae rather than being disassembled immediately after cytokinesis. The possibility that C. albicans septins are regulated by the small ubiquitin-like protein SUMO was examined in this study because the Saccharomyces cerevisiae septins were shown previously to be modified by SUMO (Smt3p). However, SUMO conjugation to septins was not detected during budding or hyphal morphogenesis in C. albicans. These results are supported by the lack of conserved SUMO consensus motifs between septins from the two organisms even after adjusting the predicted Cdc3p and Cdc12p septin sequences to account for mRNA splicing in C. albicans. Interestingly, a homolog of the Smt3p SUMO was identified in the C. albicans genome, and an epitope tagged version of Smt3p was conjugated to a variety of proteins. Immunofluorescence analysis showed prominent Smt3p SUMO localization at bud necks and sites of septum formation in hyphae similar to the septins. However, Smt3p was primarily detected on the mother cell side of the septin ring. A subset of these Smt3p-modified proteins co-immunoprecipitated with the septin Cdc11p. These results indicate that septin-associated proteins and not the septins themselves are the key target of SUMO modification at the bud neck in C. albicans. PMID- 15277522 TI - Interaction of telomestatin with the telomeric single-strand overhang. AB - The extremities of chromosomes end in a G-rich single-stranded overhang that has been implicated in the onset of the replicative senescence. The repeated sequence forming a G-overhang is able to adopt a peculiar four-stranded DNA structure in vitro called a G-quadruplex, which is a poor substrate for telomerase. Small molecule ligands that selectively stabilize the telomeric G-quadruplex induce telomere shortening and a delayed growth arrest. Here we show that the G quadruplex ligand telomestatin has a dramatic effect on the conformation of intracellular G-overhangs. Competition experiments indicate that telomestatin strongly binds in vitro and in vivo to the telomeric overhang and impairs its single-stranded conformation. Long-term treatment of cells with telomestatin greatly reduces the G-overhang size, as evidenced by specific hybridization or telomeric oligonucleotide ligation assay experiments, with a concomitant delayed loss of cell viability. In vivo protection experiments using dimethyl sulfate also indicate that telomestatin treatment alters the dimethyl sulfate effect on G overhangs, a result compatible with the formation of a local quadruplex structure at telomeric overhang. Altogether these experiments strongly support the hypothesis that the telomeric G-overhang is an intracellular target for the action of telomestatin. PMID- 15277523 TI - In vivo and in vitro reconstitution of Atg8 conjugation essential for autophagy. AB - In an analogous manner to protein ubiquitination, The C terminus of Atg8p, a yeast protein essential for autophagy, conjugates to a head group of phosphatidylethanolamine via an amide bond. Though physiological role of this reaction is assigned to membrane organization during autophagy, its molecular details are still unknown. Here, we show that Escherichia coli cells coexpressed Atg8p, Atg7p (E1), and Atg3p (E2) allowed to form conjugate of Atg8p with endogenous PE. Further, we established an in vitro Atg8p-PE reconstitution system using purified Atg8pG116, Atg7p, Atg3p, and PE-containing liposomes, demonstrating that the Atg7p and the Atg3p are minimal catalysts for Atg8p-PE conjugate reaction. Efficiency of this lipidation reaction depends on the state of the substrate, PE (phospholipid bilayer and its lipid composition). It is also suggested that the lipidation induces a conformational change in the N-terminal region of Atg8p. In vitro system developed here will provide a powerful system for further understanding the precise role of lipidation and interaction of two ubiquitin-like systems essential for autophagy. PMID- 15277524 TI - Autophosphorylation suppresses whereas kinase inhibition augments the translocation of protein kinase Calpha in response to diacylglycerol. AB - We have seen that protein kinase Calpha (PKCalpha) is transiently translocated to the plasma membrane by carbachol stimulation of neuroblastoma cells. This is induced by the Ca2+ increase, and PKCalpha does not respond to diacylglycerol (DAG). The unresponsiveness is dependent on structures in the catalytic domain of PKCalpha. This study was designed to investigate if and how the kinase activity and autophosphorylation are involved in regulating the translocation. PKCalpha enhanced green fluorescent protein translocation was studied in living neuroblastoma cells by confocal microscopy. Carbachol stimulation induced a transient translocation of PKCalpha to the plasma membrane and a sustained translocation of kinase-dead PKCalpha. In cells treated with the PKC inhibitor GF109203X, wild-type PKCalpha also showed a sustained translocation. The same effects were seen with PKCbetaI, PKCbetaII, and PKCdelta. Only kinase-dead and not wild-type PKCalpha translocated in response to 1,2-dioctanoylglycerol. To examine whether autophosphorylation regulates relocation to the cytosol, the autophosphorylation sites in PKCalpha were mutated to glutamate, to mimic phosphorylation, or alanine, to mimic the non-phosphorylated protein. After stimulation with carbachol, glutamate mutants behaved like wild-type PKCalpha, whereas alanine mutants behaved like kinase-dead PKCalpha. When the alanine mutants were treated with 1,2-dioctanoylglycerol, all cells showed a sustained translocation of the protein. However, neither carbachol nor GF109203X had any major effects on the level of autophosphorylation, and GF109203X potentiated the translocation of the glutamate mutants. We, therefore, hypothesize that 1) autophosphorylation of PKCalpha limits its sensitivity to DAG and 2) that kinase inhibitors augment the DAG sensitivity of PKCalpha, perhaps by destabilizing the closed conformation. PMID- 15277525 TI - The transcription factor CCAAT/enhancer-binding protein alpha is required for the intracellular retention of GLUT4. AB - Insulin modulates glucose uptake into adipocytes by regulating the trafficking of the GLUT4 glucose transporter. GLUT4 is mostly excluded from the surface of unstimulated cells because it is much more slowly exocytosed than it is endocytosed. GLUT4 traffics through an adipocyte-specific, specialized endosomal recycling pathway that only partially overlaps with compartments of the general endosomal recycling pathway. Insulin stimulates GLUT4 exocytosis and partially inhibits its endocytosis, resulting in GLUT4 redistribution to the cell surface. Insulin does not stimulate glucose uptake into adipocytes lacking the CCAAT/enhancer-binding protein alpha (C/EBPalpha) transcription factor. Here we show that these adipocytes do not properly traffic GLUT4. In these adipocytes, GLUT4 was rapidly exocytosed in basal conditions, resulting in an accumulation of GLUT4 on the plasma membrane. Although the kinetics of GLUT4 trafficking were altered, GLUT4 was still targeted to specialized intracellular compartments in adipocytes lacking C/EBPalpha, demonstrating an uncoupling of the targeting of GLUT4 to a specialized, adipocyte-specific insulin-regulated pathway from the regulation of the movement of GLUT4 through this pathway. Re-expression of C/EBPalpha in adipocytes lacking C/EBPalpha restored normal GLUT4 trafficking. We propose that C/EBPalpha controls the expression of the proteins that determine the basal, slow exocytosis of GLUT4, but not the proteins required to make the adipocyte-specific compartments through which GLUT4 traffics. Furthermore, these data support a model in which insulin stimulates GLUT4 exocytosis by releasing an inhibitor of GLUT4 movement to the cell surface, and it is this clamp on basal exocytosis that is missing in adipocytes lacking C/EBPalpha. PMID- 15277526 TI - Microphthalmia transcription factor induces both retinal pigmented epithelium and neural crest melanocytes from neuroretina cells. AB - Mitf encodes a basic helix-loop-helix transcription factor that plays an essential role in the differentiation of the retinal pigmented epithelium (RPE) and neural crest-derived melanocytes. As cells containing melanogenic enzymes (TRP2) are found in Mitf mouse mutants, it is not clear whether Mitf is a downstream factor or a master regulator of melanocyte differentiation. To further study the role of Mitf in committing cells to the melanocyte lineage, we express Mitf in the cultured quail neuroretina cells. This leads to the induction of two types of pigmented cells: neural crest-derived melanocytes, according to their dendritic morphology, physiology, and gene expression pattern are observed together with pigmented epithelial RPE-like cells. The expression of Mitf is lower in pigmented epithelial RPE-like cells than in neural crest-derived melanocytes. Accordingly, overexpression of Mitf in cultured quail RPE causes cells to develop into neural crest-like pigmented cells. Thus, Mitf is sufficient for the proper differentiation of crest-like pigmented cells from retinal cells and its expression level may determine the type of pigment cell induced. PMID- 15277527 TI - The essential ATP-binding cassette protein RLI1 functions in translation by promoting preinitiation complex assembly. AB - RLI1 is an essential yeast protein closely related in sequence to two soluble members of the ATP-binding cassette family of proteins that interact with ribosomes and function in translation elongation (YEF3) or translational control (GCN20). We show that affinity-tagged RLI1 co-purifies with eukaryotic translation initiation factor 3 (eIF3), eIF5, and eIF2, but not with other translation initiation factors or with translation elongation or termination factors. RLI1 is associated with 40 S ribosomal subunits in vivo, but it can interact with eIF3 and -5 independently of ribosomes. Depletion of RLI1 in vivo leads to cessation of growth, a lower polysome content, and decreased average polysome size. There was also a marked reduction in 40 S-bound eIF2 and eIF1, consistent with an important role for RLI1 in assembly of 43 S preinitiation complexes in vivo. Mutations of conserved residues in RLI1 expected to function in ATP hydrolysis were lethal. A mutation in the second ATP-binding cassette domain of RLI1 had a dominant negative phenotype, decreasing the rate of translation initiation in vivo, and the mutant protein inhibited translation of a luciferase mRNA reporter in wild-type cell extracts. These findings are consistent with a direct role for the ATP-binding cassettes of RLI1 in translation initiation. RLI1-depleted cells exhibit a deficit in free 60 S ribosomal subunits, and RLI1-green fluorescent protein was found in both the nucleus and cytoplasm of living cells. Thus, RLI1 may have dual functions in translation initiation and ribosome biogenesis. PMID- 15277528 TI - Rho and Rho-kinase mediate thrombin-induced phosphatidylinositol 4-phosphate 5 kinase trafficking in platelets. AB - Phosphatidylinositol 4-phosphate 5-kinase (PIP5K) catalyzes the rate-limiting step in the production of phosphatidylinositol 4,5-bisphosphate (PIP(2)), a signaling phospholipid that contributes to actin dynamics. We have shown in transfected tissue culture cells that PIP5K translocates from the cytosol to the plasma membrane following agonist-induced stimulation of Rho family GTPases. Nonetheless, it is unclear whether Rho GTPases induce PIP5K relocalization in platelets. We used PIP5K isoform-specific immunoblotting and lipid kinase assays to examine the intracellular localization of PIP5K in resting and activated platelets. Using differential centrifugation to separate the membrane skeleton, actin filaments and associated proteins, and cytoplasmic fractions, we found that PIP5K isoforms were translocated from cytosol to actin-rich fractions following stimulation of the thrombin receptor. PIP5K translocation was detectable within 30 s of stimulation and was complete by 2-5 min. This agonist-induced relocalization and activation of PIP5K was inhibited by 8-(4 parachlorophenylthio)-cAMP, a cAMP analogue that inhibits Rho and Rac. In contrast, 8-(4-parachlorophenylthio)-cGMP, a cGMP analogue that inhibits Rac but not Rho, did not affect PIP5K translocation and activation. This suggests that Rho GTPase may be an essential regulator of PIP5K in platelets. Consistent with this hypothesis, we found that C3 exotoxin (a Rho-specific inhibitor) and HA1077 (an inhibitor of the Rho effector, Rho-kinase) also eliminated PIP5K activation and trafficking into the membrane cytoskeleton. Thus, these data indicate that Rho GTPase and its effector Rho-kinase have an intimate relationship with the trafficking and activation of platelet PIP5K. Moreover, these data suggest that relocalization of platelet PIP5K following agonist stimulation may play an important role in regulating the assembly of the platelet cytoskeleton. PMID- 15277529 TI - Microglia-derived pronerve growth factor promotes photoreceptor cell death via p75 neurotrophin receptor. AB - Reports implicating microglia-derived nerve growth factor (NGF) during programmed cell death in the developing chick retina led us to investigate its possible role in degenerative retinal disease. Freshly isolated activated retinal microglia expressed high molecular weight forms of neurotrophins including that of nerve growth factor (NGF), brain-derived neurotrophic factor, neurotrophin-3, and neurotrophin-4. Conditioned media from cultured retinal microglia (MGCM) consistently yielded a approximately 32-kDa NGF-reactive band when supplemented with bovine serum albumin (BSA) or protease inhibitors (PI); and promoted cell death that was suppressed by NGF immunodepletion in a mouse photoreceptor cell line (661w). The approximately 32 kDa protein was partially purified (MGCM/p32) and was highly immunoreactive with a polyclonal anti-pro-NGF antibody. Both MGCM/p32 and recombinant pro-NGF protein promoted cell death in 661w cultures. Increased levels of pro-NGF mRNA and protein were observed in the RCS rat model of retinal dystrophy. MGCM-mediated cell death was reversed by p75NTR antiserum in p75NTR(+)/trkA(-) 661w cells. Our study shows that a approximately 32 kDa pro NGF protein released by activated retinal microglia promoted degeneration of cultured photoreceptor cells. Moreover, our study suggests that defective post translational processing of NGF might be involved in photoreceptor cell loss in retinal dystrophy. PMID- 15277530 TI - The forkhead transcription factor FoxC2 inhibits white adipocyte differentiation. AB - In this study, we show that expression of FoxC2 blocks the capacity of 3T3-L1 preadipocytes to undergo adipogenesis in the presence of dexamethasone, isobutylmethylxanthine, and insulin. This block is characterized by an extensive decrease in the expression of proteins associated with the function of the mature fat cell, most notably C/EBPalpha, adiponectin, perilipin, and the adipose specific fatty acid-binding protein, FABP4/aP2. Since the expression of these proteins lies downstream of PPARgamma, we overexpressed PPARgamma in Swiss mouse fibroblasts to promote adipocyte differentiation. We show that FoxC2 blocks the ability of PPARgamma to induce adipogenic gene expression in response to exposure of the cells to dexamethasone, isobutylmethylxanthine, insulin, and a PPARgamma ligand. Interestingly, the expression of aP2 escapes the inhibitory action of FoxC2 under conditions that promote maximum PPARgamma activity. In contrast, FoxC2 inhibits the expression of C/EBPalpha, perilipin, and adiponectin even in the presence of potent PPARgamma ligands. Finally, we show that FoxC2 does not affect the ability of PPARgamma to bind to or transactivate from a PPARgamma response element. These data suggest that FoxC2 blocks adipogenesis by inhibiting the capacity of PPARgamma to promote the expression of a subset of adipogenic genes. PMID- 15277531 TI - Jamip1 (marlin-1) defines a family of proteins interacting with janus kinases and microtubules. AB - Jamip1 (Jak and microtubule interacting protein), an alias of Marlin-1, was identified for its ability to bind to the FERM (band 4.1 ezrin/radixin/moesin) homology domain of Tyk2, a member of the Janus kinase (Jak) family of non receptor tyrosine kinases that are central elements of cytokine signaling cascades. Jamip1 belongs to a family of three genes conserved in vertebrates and is predominantly expressed in neural tissues and lymphoid organs. Jamip proteins lack known domains and are extremely rich in predicted coiled coils that mediate dimerization. In our initial characterization of Jamip1 (73 kDa), we found that it comprises an N-terminal region that targets the protein to microtubule polymers and, when overexpressed in fibroblasts, profoundly perturbs the microtubule network, inducing the formation of tight and stable bundles. Jamip1 was shown to associate with two Jak family members, Tyk2 and Jak1, in Jurkat T cells via its C-terminal region. The restricted expression of Jamip1 and its ability to associate to and modify microtubule polymers suggest a specialized function of these proteins in dynamic processes, e.g. cell polarization, segregation of signaling complexes, and vesicle traffic, some of which may involve Jak tyrosine kinases. PMID- 15277532 TI - Sef interacts with TAK1 and mediates JNK activation and apoptosis. AB - Sef was recently identified as a negative regulator of fibroblast growth factor (FGF) signaling in a genetic screen of zebrafish and subsequently in mouse and humans. By inhibiting FGFR1 tyrosine phosphorylation and/or Ras downstream events, Sef inhibits FGF-mediated ERK activation and cell proliferation as well as PC12 cell differentiation. Here we show that Sef and a deletion mutant of Sef lacking the extracellular domain (SefIC) physically interact with TAK1 (transforming growth factor-beta-associated kinase) and activate JNK through a TAK1-MKK4-JNK pathway. Sef and SefIC overexpression also resulted in apoptotic cell death, while dominant negative forms of MKK4 and TAK1 blocked Sef-mediated JNK activation and attendant 293T cell apoptosis. These investigations reveal a novel activating function of Sef that is distinct from its inhibitory effect on FGF receptor signaling and ERK activation. PMID- 15277533 TI - Cytosolic RNase inhibitor only affects RNases with intrinsic cytotoxicity. AB - Cytosolic RNase inhibitor binds to and neutralizes most members of the pancreatic type RNase superfamily. However, there are a few exceptions, e.g. amphibian onconase and bovine seminal RNase, and these are endowed with cytotoxic activity. Also, RNase variants created by mutagenesis to partially evade the RNase inhibitor acquire cytotoxic activity. These findings have led to the proposal that the cytosolic inhibitor acts as a sentry to protect mammalian cells from foreign RNases. We silenced the expression of the gene encoding the cytosolic inhibitor in HeLa cells and found that the cells become more sensitive to foreign cytotoxic RNases. However foreign, non-cytotoxic RNases remain non-cytotoxic. These results indicate that the cytosolic inhibitor neutralizes those foreign RNases that are intrinsically cytotoxic and have access to the cytosol. However, its normal physiological role may not be to guard against foreign RNases in general. PMID- 15277534 TI - Protective effect of phosphatidylinositol 4,5-bisphosphate against cortical filamentous actin loss and insulin resistance induced by sustained exposure of 3T3-L1 adipocytes to insulin. AB - Muscle and fat cells develop insulin resistance when cultured under hyperinsulinemic conditions for sustained periods. Recent data indicate that early insulin signaling defects do not fully account for the loss of insulin action. Given that cortical filamentous actin (F-actin) represents an essential aspect of insulin regulated glucose transport, we tested to see whether cortical F-actin structure was compromised during chronic insulin treatment. The acute effect of insulin on GLUT4 translocation and glucose uptake was diminished in 3T3 L1 adipocytes exposed to a physiological level of insulin (5 nm) for 12 h. This insulin-induced loss of insulin responsiveness was apparent under both low (5.5 mm) and high (25 mm) glucose concentrations. Microscopic and biochemical analyses revealed that the hyperinsulinemic state caused a marked loss of cortical F actin. Since recent data link phosphatidylinositol 4,5-bisphosphate (PIP(2)) to actin cytoskeletal mechanics, we tested to see whether the insulin-resistant condition affected PIP(2) and found a noticeable loss of this lipid from the plasma membrane. Using a PIP(2) delivery system, we replenished plasma membrane PIP(2) in cells following the sustained insulin treatment and observed a restoration in cortical F-actin and insulin responsiveness. These data reveal a novel molecular aspect of insulin-induced insulin resistance involving defects in PIP(2)/actin regulation. PMID- 15277535 TI - Comparison of the efficacy, toxicity, and pharmacokinetics of a uracil/tegafur (UFT) plus oral leucovorin (LV) regimen between Japanese and American patients with advanced colorectal cancer: joint United States and Japan study of UFT/LV. AB - PURPOSE: To compare the efficacy, toxicities, and pharmacokinetics of an oral regimen consisting of uracil/tegafur (UFT) and leucovorin (LV) between Japanese patients and patients in the United States with previously untreated metastatic colorectal cancer. PATIENTS AND METHODS: Forty-four Japanese patients and 45 patients in the United States were enrolled in concurrent nonrandomized phase II trials. UFT 300 mg/m2/d and leucovorin 75 mg/d were administered orally for 28 days followed by a 7-day rest period. The total daily dose of each drug was divided into three equal doses. Treatment was repeated every 5 weeks until disease progression. Blood samples for the pharmacokinetic study were obtained after the initial dose on day 1 of the first course. RESULTS: The response rate for the Japanese patients and the patients in the United States was 36.4% (95% CI, 22.4% to 52.2%) and 34.1% (95% CI, 20.5% to 49.9%), respectively. The only major toxicity was diarrhea, and other toxicities were mild in both populations. The incidence of grade 3 or higher diarrhea in the Japanese and Americans was 9% and 22%, respectively. Although the area under the curve and maximum concentration of fluorouracil were found to be slightly higher in the Japanese patients than the patients in the United States, and area under the curve adjusted body surface area appeared to be comparable between the two groups. CONCLUSION: The efficacy and pharmacokinetic parameters of UFT and LV are comparable in Japanese and American patients; however, a difference in toxicity profile, specifically diarrhea, was noted. This oral regimen of UFT and LV is considered to have similar activity against metastatic colorectal cancer and to have acceptable toxicity in patients in both countries. PMID- 15277536 TI - Retinoids in lung cancer: friend, foe, or fellow traveler? PMID- 15277537 TI - Pattern of care at the end of life: does age make a difference in what happens to women with breast cancer? AB - PURPOSE: In the last 40 years, palliative care has become the standard of care at the end of life. However, there are limited data about the degree of access to such care at the population level. METHODS: Using administrative databases, a care-oriented profile score was created to describe the care received during the last 6 months of life for 2,291 women who were dying of breast cancer in the province of Quebec, Canada, during the years 1992 to 1998. The care received was described through indicators of care that would reflect a palliative care philosophy. An ordinal score was developed for comparisons among age groups of women using a proportional odds ordinal regression model. RESULTS: We found that only 6.9% of women died at home, while 69.6% of them died in acute care beds. While most women (75%) had few indicators indicating provision of palliative care during the last 6 months of life, younger women (< 50 years) were even less likely (odds ratio, 0.70; 95% CI, 0.54 to 0.90) to receive such care compared with middle aged women (50 to 59 years; serving as the reference group), while older women (> 70 years) were more likely (odds ratio, 1.85; 95% CI, 1.49 to 2.29). CONCLUSION: Our study indicates that a sizeable proportion of women terminally ill from breast cancer do not have access to palliative care-an issue that health care policy makers may wish to explore further. PMID- 15277538 TI - Bridging the divide: integrating cancer-directed therapy and palliative care. PMID- 15277539 TI - Cancer drug development: for populations or for individuals? PMID- 15277540 TI - Promoter methylation of retinoic acid receptor beta 2 and the development of second primary lung cancers in non-small-cell lung cancer. AB - PURPOSE: To investigate whether the promoter hypermethylation of retinoic acid receptor beta 2 (RARbeta2) is associated with the development of second primary lung cancers (SPLCs) differentially according to smoking status in primary non small-cell lung cancer (NSCLC). PATIENTS AND METHODS: We retrospectively analyzed the relationship between RARbeta2 methylation and the SPLC development in a total of 342 NSCLCs. The methylation status of RARbeta2 was determined by using methylation-specific polymerase chain reaction. The difference in the time to SPLC development was analyzed by using the log-rank test and the Cox proportional hazards model. The median follow-up was 4.1 years. RESULTS: SPLCs developed in 19 (5.6%) of the 342 NSCLCs, and overall incidence rate of SPLC development was 1.54 per 100 patient-years. SPLCs did not occur in 39 patients who had not smoked. After controlling for possible confounding factors, the hazard of failure for former smokers with RARbeta2 hypermethylation was about 2.87 (95% CI, 0.92 to 13.64; P =.08) times higher compared to those without RARbeta2 methylation. However, for current smokers, hypermethylation of the RARbeta2 was found to have a protective effect against the SPLC development (hazard ratio = 0.23; 95% CI, 0.11 to 0.87; P =.03). CONCLUSION: Hypermethylation of RARbeta2 promoter had a differential effect on the development of SPLCs in NSCLC, and this was dependent on smoking status. Our study suggests that a combination of retinoids and/or a demethylating agent may be effective in the prevention of SPLCs in never-smokers and former smokers with NSCLC. PMID- 15277541 TI - Muscle satellite cells adopt divergent fates: a mechanism for self-renewal? AB - Growth, repair, and regeneration of adult skeletal muscle depends on the persistence of satellite cells: muscle stem cells resident beneath the basal lamina that surrounds each myofiber. However, how the satellite cell compartment is maintained is unclear. Here, we use cultured myofibers to model muscle regeneration and show that satellite cells adopt divergent fates. Quiescent satellite cells are synchronously activated to coexpress the transcription factors Pax7 and MyoD. Most then proliferate, down-regulate Pax7, and differentiate. In contrast, other proliferating cells maintain Pax7 but lose MyoD and withdraw from immediate differentiation. These cells are typically located in clusters, together with Pax7-ve progeny destined for differentiation. Some of the Pax7+ve/MyoD-ve cells then leave the cell cycle, thus regaining the quiescent satellite cell phenotype. Significantly, noncycling cells contained within a cluster can be stimulated to proliferate again. These observations suggest that satellite cells either differentiate or switch from terminal myogenesis to maintain the satellite cell pool. PMID- 15277542 TI - Monitoring disulfide bond formation in the eukaryotic cytosol. AB - Glutathione is the most abundant low molecular weight thiol in the eukaryotic cytosol. The compartment-specific ratio and absolute concentrations of reduced and oxidized glutathione (GSH and GSSG, respectively) are, however, not easily determined. Here, we present a glutathione-specific green fluorescent protein based redox probe termed redox sensitive YFP (rxYFP). Using yeast with genetically manipulated GSSG levels, we find that rxYFP equilibrates with the cytosolic glutathione redox buffer. Furthermore, in vivo and in vitro data show the equilibration to be catalyzed by glutaredoxins and that conditions of high intracellular GSSG confer to these a new role as dithiol oxidases. For the first time a genetically encoded probe is used to determine the redox potential specifically of cytosolic glutathione. We find it to be -289 mV, indicating that the glutathione redox status is highly reducing and corresponds to a cytosolic GSSG level in the low micromolar range. Even under these conditions a significant fraction of rxYFP is oxidized. PMID- 15277545 TI - To freeze or not to freeze: adaptations for overwintering by hatchlings of the North American painted turtle. AB - Many physiologists believe that hatchling painted turtles (Chrysemys picta) provide a remarkable, and possibly unique, example of 'natural freeze-tolerance' in an amniotic vertebrate. However, the concept of natural freeze-tolerance in neonatal painted turtles is based on results from laboratory studies that were not placed in an appropriate ecological context, so the concept is suspect. Indeed, the weight of current evidence indicates that hatchlings overwintering in the field typically withstand exposure to ice and cold by avoiding freezing altogether and that they do so without benefit of an antifreeze to depress the equilibrium freezing point for bodily fluids. As autumn turns to winter, turtles remove active nucleating agents from bodily fluids (including bladder and gut), and their integument becomes a highly efficient barrier to the penetration of ice into body compartments from frozen soil. In the absence of a nucleating agent or a crystal of ice to 'catalyze' the transformation of water from liquid to solid, the bodily fluids remain in a supercooled, liquid state. The supercooled animals nonetheless face physiological challenges, most notably an increased reliance on anaerobic metabolism as the circulatory system first is inhibited and then caused to shut down by declining temperature. Alterations in acid/base status resulting from the accumulation of lactic acid may limit survival by supercooled turtles, and sublethal accumulations of lactate may affect behavior of turtles after the ground thaws in the spring. The interactions among temperature, circulatory function, metabolism (both aerobic and anaerobic), acid/base balance and behavior are fertile areas for future research on hatchlings of this model species. PMID- 15277543 TI - Zinc and the Msc2 zinc transporter protein are required for endoplasmic reticulum function. AB - In this report, we show that zinc is required for endoplasmic reticulum function in Saccharomyces cerevisiae. Zinc deficiency in this yeast induces the unfolded protein response (UPR), a system normally activated by unfolded ER proteins. Msc2, a member of the cation diffusion facilitator (CDF) family of metal ion transporters, was previously implicated in zinc homeostasis. Our results indicate that Msc2 is one route of zinc entry into the ER. Msc2 localizes to the ER when expressed at normal levels. UPR induction in low zinc is exacerbated in an msc2 mutant. Genetic and biochemical evidence indicates that this UPR induction is due to genuine ER dysfunction. Notably, we found that ER-associated protein degradation is defective in zinc-limited msc2 mutants. We also show that the vacuolar CDF proteins Zrc1 and Cot1 are other pathways of ER zinc acquisition. Finally, zinc deficiency up-regulates the mammalian ER stress response indicating a conserved requirement for zinc in ER function among eukaryotes. PMID- 15277544 TI - Activation of Cdc42 by trans interactions of the cell adhesion molecules nectins through c-Src and Cdc42-GEF FRG. AB - Nectins, Ca2+ -independent immunoglobulin-like cell-cell adhesion molecules, initiate cell-cell adhesion by their trans interactions and recruit cadherins to cooperatively form adherens junctions (AJs). In addition, the trans interactions of nectins induce the activation of Cdc42 and Rac small G proteins, which increases the velocity of the formation of AJs. We examined here how nectins induce the activation of Cdc42 in MDCK epithelial cells and L fibroblasts. Nectins recruited and activated c-Src at the nectin-based cell-cell adhesion sites. FRG, a GDP/GTP exchange factor specific for Cdc42, was then recruited there, tyrosine phosphorylated by c-Src, and activated, causing an increase in the GTP-bound active form of Cdc42. Inhibition of the nectin-induced activation of c-Src suppressed the nectin-induced activation of FRG and Cdc42. Inhibition of the nectin-induced activation of FRG or depletion of FRG by RNA interference suppressed the nectin-induced activation of Cdc42. These results indicate that nectins induce the activation of Cdc42 through c-Src and FRG locally at the nectin-based cell-cell adhesion sites. PMID- 15277546 TI - The electric fish Brachyhypopomus pinnicaudatus produces jamming avoidance responses to signals that are harmonically related to its own discharges. AB - Jamming avoidance responses (JARs) are exhibited by pairs of pulse type electric fish that discharge with similar frequencies whenever their individual pulses are about to coincide: responses consist of the transient shortenings in inter discharge intervals in the fish with the higher frequency. This study describes and models novel forms of JARs observed in sexually mature male or female Brachyhypopomus pinnicaudatus. One novel JAR was observed in male-female pairs in their natural habitat. It happened when the baseline frequencies were not similar but, rather, when one was almost twice that of the other; moreover, the transient interval shortenings occurred not in the fish with the higher frequency but in the slower one. Transient interval shortenings similar to those in all natural JARs were observed in individual fish in tanks and submitted to periodic electrical pulse trains. They happened not only when pulse frequencies were slightly lower than the unperturbed frequency emitted by the fish but also when slightly lower than the frequency's sub- or higher harmonics (e.g. one half or twice). The proposed model satisfactorily reproduces all experimental observations. In it, forthcoming inter-pulse intervals reflect the differences between the cophases of pulses that arrive within the 'sensitive windows' belonging to either consecutive (i.e. one and the next) or alternating (e.g. every other, every three) intervals. Paired pulse fish embody interacting oscillators, and, in particular, JARs embody either quasiperiodic phase walk throughs and intermittencies or periodic and locked forms. Hence, their study would profit by the powerful theories and approaches advanced by nonlinear dynamics. PMID- 15277547 TI - Ontogenesis of the attachment ability in the bug Coreus marginatus (Heteroptera, Insecta). AB - Each tarsus of Coreus marginatus L. (Coreidae) bears a pair of smooth flexible pulvilli adapted for attachment to relatively smooth surfaces, such as their host plant Rumex crispus L. (Polygonaceae). This account quantifies insect attachment abilities on smooth surfaces at various stages of ontogenesis. Friction (shear) force (FF) of adults and juvenile insects was measured by the use of a computer controlled centrifugal force tester equipped with a fibre optical sensor. Pad area, body size and body mass were determined individually for each experimental insect. Light microscopy revealed no difference in pulvilli area between different leg pairs. Pulvilli area demonstrated a stronger increase with increasing linear dimensions, as predicted by scaling laws. Since friction coefficient (relationship between FF and body weight) (FC) was always higher than 1, it was concluded that adhesion has strongly contributed to the measured friction. The frictional properties of pulvilli do not change during ontogenesis. Thus, only the growth of pulvilli and, therefore, the increased contact area, contribute to the increasing attachment ability in insects at later larval stages. Due to different scaling of the body mass and area of attachment organs, smaller insects attach relatively more strongly. Both FF and FC were higher in experiments in which higher angular acceleration (AC) was applied. Lateral tenacity determined individually for experimental insects and pooled for all animals and accelerations is 0.097 N m(-2). These data led us to suggest that viscosity of the pad secretion and/or visco-elastic properties of the foam-like material of pulvilli play an important role in the attachment ability of insects. PMID- 15277548 TI - Thermal stability and muscle efficiency in hovering orchid bees (Apidae: Euglossini). AB - To test whether variation in muscle efficiency contributes to thermal stability during flight in the orchid bee, Euglossa imperialis, we measured CO2 production, heat loss and flight kinematics at different air temperatures (Ta). We also examined the relationship between wingbeat frequency (WBF) and Ta in five additional species of orchid bees. Mean thoracic temperature (Tth) for Eg. imperialis hovering in a screened insectary and in the field was 39.3+/-0.77 degrees C (mean +/- 95% C.I.), and the slope of Tth on Ta was 0.57. Head and abdominal temperature excess ratios declined with Ta, indicating that Eg. imperialis were not increasing heat dissipation from the thorax at high Ta. Elevation of Tth above Ta was correlated with WBF, but Tth alone was not. Estimates of heat production from both respirometry and heat loss experiments decreased 33% as Ta rose from 24 to 34 degrees C. Mean muscle efficiency over this temperature range was 18% assuming perfect elastic energy storage and 22% assuming zero elastic energy storage. Both efficiency estimates increased significantly as Ta rose from 24 to 34 degrees C. In all six species examined, WBF declined significantly with Ta. These data indicate that hovering orchid bees regulate heat production through changes in wingbeat kinematics and consequent changes in energy conversion by the flight motor. Temperature-dependent variation in elastic energy storage or muscle contraction efficiency or both may contribute to the observed trends. PMID- 15277549 TI - Cytotoxicity of diatom-derived oxylipins in organisms belonging to different phyla. AB - The cytotoxicity of several saturated and unsaturated marine diatom-derived aldehydes and an oxo-acid have been screened in vitro and in vivo against different organisms, such as bacteria, algae, fungi, echinoderms, molluscs and crustaceans. Conjugated unsaturated aldehydes like 2E,4E-decadienal, 2E,4E octadienal, 5E,7E-9-oxo-nonadienoic acid and 2E-decenal were active against bacteria and fungi and showed weak algicidal activity. By contrast, the saturated aldehyde decanal and the non-conjugated aldehyde 4Z-decenal had either low or no significant biological activity. In assays with oyster haemocytes, 2E,4E decadienal exhibited a dose-dependent inhibition of cytoskeleton organisation, rate of phagocytosis and oxidative burst and a dose-dependent promotion of apoptosis. A maternal diatom diet that was rich in unsaturated aldehydes induced arrest of cell division and apoptotic cell degradation in copepod embryos and larvae, respectively. This wide spectrum of physiological pathologies reflects the potent cell toxicity of diatom-derived oxylipins, in relation to their non specific chemical reactivity towards nucleophilic biomolecules. The cytotoxic activity is conserved across six phyla, from bacteria to crustaceans. Deregulation of cell homeostasis is supposed to induce the elimination of damaged cells through apoptosis. However, efficient protection mechanisms possibly exist in unicellular organisms. Experiments with a genetically modified yeast species exhibiting elevated membrane and/or cell wall permeability suggest that this protection can be related to the inability of the oxylipin compounds to enter the cell. PMID- 15277550 TI - Structure and properties of the glandular surface in the digestive zone of the pitcher in the carnivorous plant Nepenthes ventrata and its role in insect trapping and retention. AB - Carnivorous plants of the genus Nepenthes grow in nutrient-poor habitats and have evolved specialised trapping organs, known as pitchers. These are composed of different surface zones serving the functions of attraction, capture and digestion of insects, which represent a main source of nitrogen. To investigate the role of the glandular digestive zone in the trapping mechanism of the pitcher, structural, mechanical and physico-chemical studies were applied to N. ventrata and combined with insect behavioural experiments. It was found that the glandular surface is microscopically rough since it is regularly structured with multicellular glands situated in epidermal depressions. The presence of downward directed 'hoods' over the upper part of glands and sloped depressions in the proximal direction of the pitcher causes a marked anisotropy of the surface. The glandular zone surface is composed of relatively stiff material (Young's modulus, 637.19+/-213.44 kPa). It is not homogeneous, in terms of adhesive properties, and contains numerous areas without adhesion as well as adhesive areas differing greatly in tenacity values (range, 1.39-28.24 kPa). The surface is readily wettable with water (contact angle, 31.9-36.0 degrees C) and has a high surface free energy (56.84-61.93 mN m(-1)) with a relatively high polar component (33.09 52.70 mN m(-1)). To examine the effect of the glandular secretion on attachment systems of insects having hairy and smooth adhesive pads, forces generated on different surfaces by Calliphora vicina flies and Pyrrhocoris apterus bugs, respectively, were measured. Flies attached equally well to both fresh and air dried glandular surfaces whereas bugs generated a significantly lower force on the fresh glandular surface compared with the air-dried one. It is assumed that the contribution of the glandular surface to insect retention, due to its effect on insect attachment, differs depending on insect weight and the type of insect attachment system. Surface anisotropy does not facilitate effective claw interlocking so that insects possessing only claws are probably not able to cling to the glandular surface. However, stiffness of the pitcher wall material in the digestive zone can provide claw clinging via punching of the pitcher wall by claws. Small insects lacking pads may use adhesive areas on the plant surface to attach themselves, but such solitary points with very strong adhesion possibly impede their overall locomotion and chance of escape. Pad-bearing insects are presumably able to attach to smooth parts of the glandular surface located between glands. High free surface energy of the plant substrate may promote adhesion. Gland secretion may decrease attachment ability in insects with smooth adhesive pads but not influence attachment of insects with hairy attachment systems. PMID- 15277551 TI - Magnetic sense in the Japanese eel, Anguilla japonica, as determined by conditioning and electrocardiography. AB - Magnetosensitivity of the Japanese eel, Anguilla japonica, was examined by conditioning and electrocardiography. Marine eels, river eels and farmed eels were conditioned to an imposed magnetic field ranging from 12,663 nT to 192,473 nT parallel to the fish body, which was placed along the earth's west-east axis. Electrocardiograms were recorded with electrodes placed close to the fish body inside a PVC pipe shelter. After 10-40 conditioning runs, all the eels exhibited a significant conditioned response (i.e. slowing of the heart beat) to a 192,473 nT magnetic field and even to a 12,663 nT magnetic field, respectively equivalent to 5.92x and 0.38x the horizontal geomagnetic field (32,524 nT) at our laboratory. The west-east vector of the imposed magnetic field (12,663 nT) combined with that of the geomagnetic field and produced a horizontal resultant magnetic field of 21 degrees easterly. Therefore, Japanese eel are magnetosensitive whether they are at sea, in the river or in the farm. Results of the present study were compared with those of past studies that showed no magnetic sense in the American eel, Anguilla rostrata, and the European eel, Anguilla anguilla. PMID- 15277552 TI - The importance of the lateral line in nocturnal predation of piscivorous catfish. AB - In a previous study we showed that nocturnal piscivorous catfish track the wake left by a swimming prey fish to locate it, following past locations to detect the present location of the prey. In a wake there are hydrodynamic as well as chemical signatures that both contain information on location and suitability of the prey. In order to determine how these two wake stimuli are utilised in prey tracking, we conducted experiments in catfish in which either the lateral line or the external gustation was ablated. We found that a functional lateral line is indispensable for following the wake of swimming prey. The frequency of attack and capture was greatly diminished and the attacks that did occur were considerably delayed when the lateral line was ablated. In contrast, catfish with ablated external taste still followed the wakes of their prey prior to attacking, albeit their attacks were delayed. The external taste sense, which was reported earlier to be necessary for finding stationary (dead) food, seems to play a minor role in the localisation of moving prey. Our finding suggests that an important function of the lateral line is to mediate wake-tracking in predatory fish. PMID- 15277553 TI - Does condition of Atlantic cod (Gadus morhua) have a greater impact upon swimming performance at Ucrit or sprint speeds? AB - To compare the sensitivity of sprint and critical (Ucrit) swimming speeds to the condition of Atlantic cod (Gadus morhua) and to identify the best anatomic, behavioural and biochemical correlates of these types of swimming, we established two groups of cod that were fed or starved for 12 weeks. We evaluated sprint swimming and Ucrit performance as well as the speed at which repeated burst-coast movements began in the Ucrit test before measuring the metabolic capacities of red and white muscle sampled caudally, centrally and rostrally and the anatomic characteristics of the cod. White muscle lactate was measured directly after the Ucrit test. As expected, the twofold difference in Fulton's condition factor (0.5+/-0.04 for starved and 1.0+/-0.1 for fed cod) was accompanied by large differences in the anatomic and biochemical parameters measured. Despite the relative sparing of muscle aerobic capacity during starvation and despite the greater use of oxidative fibres during Ucrit compared with sprint swimming, these types of swimming differed by much the same extent between starved and fed cod. In the Ucrit tests, white muscle lactate levels and lactate accumulation per burst-coast movement were considerably higher in fed than starved cod, suggesting more intensive use of fast muscle fibres in cod in good condition. Multiple regression analysis indicated strong correlations between Ucrit, the speed at which regular burst-coasting began and the activity of pyruvate dehydrogenase (PDH) in red muscle (both caudal and central positions). PDH activity may limit the rate of oxidative ATP production by red muscle. The activity of cytochrome c oxidase in rostral white muscle was the strongest correlate of sprint swimming, suggesting that aerobic preparation of white muscle facilitates rapid contraction. The correlation between Ucrit and sprint swimming was weak, perhaps due to inter-individual differences in sensitivity during sprint tests. PMID- 15277554 TI - Characterization and expression of plasma membrane Ca2+ ATPase (PMCA3) in the crayfish Procambarus clarkii antennal gland during molting. AB - The discontinuous pattern of crustacean cuticular mineralization (the molting cycle) has emerged as a model system to study the spatial and temporal regulation of genes that code for Ca2+-transporting proteins including pumps, channels and exchangers. The plasma membrane Ca2+-ATPase (PMCA) is potentially of significant interest due to its role in the active transport of Ca2+ across the basolateral membrane, which is required for routine maintenance of intracellular Ca2+ as well as unidirectional Ca2+ influx. Prior research has suggested that PMCA expression is upregulated during periods of elevated Ca2+ influx associated with postmolt cuticular mineralization. This paper describes the cloning, sequencing and functional characterization of a novel PMCA3 gene from the antennal gland (kidney) of the crayfish Procambarus clarkii. The complete sequence, the first obtained from a non-genetic invertebrate species, was obtained through reverse transcription-polymerase chain reaction (RTPCR) and rapid amplification of cDNA ends (RACE) techniques. Crayfish PMCA3 consists of 4148 bp with a 3546 bp open reading frame coding for 1182 amino acid residues with a molecular mass of 130 kDa. It exhibits 77.5-80.9% identity at the mRNA level and 85.3-86.9% identity at the protein level with PMCA3 from human, mouse and rat. Membrane topography was typical of published mammalian PMCAs. Northern blot analysis of total RNA from crayfish gill, antennal gland, cardiac muscle and axial abdominal muscle revealed that a 7.5 kb species was ubiquitous. The level of PMCA3 mRNA expression in all tissues (transporting epithelia and muscle) increased significantly in pre/postmolt stages compared with relatively low abundance in intermolt. Western analysis confirmed corresponding changes in PMCA protein expression (130 kDa). PMID- 15277555 TI - The effects of intense wing molt on diving in alcids and potential influences on the evolution of molt patterns. AB - Large and medium-sized alcids have a very intense wing molt wherein many flight feathers are shed in rapid succession and wing surface area is reduced by as much as 40%. Although these birds are rendered flightless during wing molt, they must still use their wings to propel themselves underwater. A molt-induced loss of wing area could simply reduce wing propulsion such that more muscular work would be required to maintain a given speed. Alternatively, molt could reduce drag on the wings, making a bird more penguin-like and actually enhancing diving ability. I addressed this issue by filming captive common guillemots Uria aalge and tufted puffins Fratercula cirrhata using an array of video cameras to plot the birds' movements in three dimensions. From these coordinate data I calculated swimming velocities, angles of descent and absolute depths. These values allowed me to estimate the forces due to drag and buoyancy that must be counteracted by flapping, which in turn yielded estimates of the amount of work generated during each flap as well as the average power and cost of transport. Within-bird comparisons of diving performance when wings were intact and during several stages of wing molt indicated that molt is associated with more frequent flapping, reduced displacement during the flap cycle, and possibly reduced work per flap. These negative effects on diving may explain why primary and secondary molts were offset in the birds I studied such that the period during which all of the flight feathers are effectively missing is minimized. PMID- 15277556 TI - Postprandial increases in nitrogenous excretion and urea synthesis in the giant mudskipper Periophthalmodon schlosseri. AB - The objective of this study was to determine the effects of feeding on the excretory nitrogen (N) metabolism of the giant mudskipper, Periophthalmodon schlosseri, with special emphasis on the role of urea synthesis in ammonia detoxification. The ammonia and urea excretion rates of P. schlosseri increased 1.70- and 1.92-fold, respectively, within the first 3 h after feeding on guppies. Simultaneously, there were significant decreases in ammonia levels in the plasma and the brain, and in urea contents in the muscle and liver, of P. schlosseri at 3 h post-feeding. Thus, it can be concluded that P. schlosseri was capable of unloading ammonia originally present in some of its tissues in anticipation of ammonia released from the catabolism of excess amino acids after feeding. Subsequently, there were significant increases in urea content in the muscle, liver and plasma (1.39-, 2.17- and 1.62-fold, respectively) at 6 h post-feeding, and the rate of urea synthesis apparently increased 5.8-fold between 3 h and 6 h. Increased urea synthesis might have occurred in the liver of P. schlosseri because the greatest increase in urea content was observed therein. The excess urea accumulated in the body at 6 h was completely excreted between 6 and 12 h, and the percentage of waste-N excreted as urea-N increased significantly to 26% during this period, but never exceeded 50%, the criterion for ureotely, meaning that P. schlosseri remained ammonotelic after feeding. By 24 h, 62.7% of the N ingested by P. schlosseri was excreted, out of which 22.6% was excreted as urea N. This is the first report on the involvement of increased urea synthesis and excretion in defense against ammonia toxicity in the giant mudskipper, and our results suggest that an ample supply of energy resources, e.g. after feeding, is a prerequisite for the induction of urea synthesis. Together, increases in nitrogenous excretion and urea synthesis after feeding effectively prevented a postprandial surge of ammonia in the plasma of P. schlosseri as reported previously for other fish species. Consequently, contrary to previous reports, there were significant decreases in the ammonia content of the brain of P. schlosseri throughout the 24 h period post-feeding, accompanied by a significant decrease in brain glutamine content between 12 h and 24 h. PMID- 15277558 TI - Experimental studies of the material properties of the forewing of cicada (Homoptera, Cicadidae). AB - Detailed investigations on the structural and mechanical properties of the forewing of the cicada were carried out. Measurement of the structures of the wings showed that the thickness of the membrane of each cell and the diameter of each vein were non-uniform in both the longitudinal and transverse directions, and their means were approximately 12.2 and 133.3 microm, respectively. However, the aspect ratios of the wings and the bodies were quite uniform and were approximately equal to 2.98 and 2.13, respectively. Based on the measured thickness, mass and area of the membranes of the cells, the mean density and the mean area density of the wing were approximately 2.3 g cm(-3) and 2.8x10(-3) g cm(-2), respectively. In addition, the diameters of the veins of the wings, including the diameters of the holes in the vein of the leading edge, were examined. The mechanical properties of the wing were investigated separately by nanoindentation and tensile testing. The results indicated that the mean Young's modulus, hardness and yield stress of the membranes of the wings were approximately 3.7 GPa, 0.2 GPa and 29 MPa, respectively, and the mean Young's modulus and strength of the veins along the direction of the venation of wings were approximately 1.9 GPa and 52 MPa, respectively. Finally, the relevant results were briefly analyzed and discussed, providing a guideline to the biomimetic design of the aerofoil materials of micro air vehicles. PMID- 15277557 TI - Expression of hypothalamic arginine vasotocin gene in response to water deprivation and sex steroid administration in female Japanese quail. AB - Arginine vasotocin (AVT) is a neurohypophyseal hormone involved in reproductive function and control of osmoregulation in birds. In view of the dual function of AVT, the present experiment was designed to observe the effect of water deprivation (WD) and sex steroid [estradiol benzoate (EB) and testosterone propionate (TP)] treatment independently, as well as simultaneously, on the profile/activity of the hypothalamic AVT system. WD resulted in a significant increase in plasma osmolality, sodium ion concentration and AVT concentration, but administration of sex steroids had no significant influence on these parameters. By contrast, the amount of hypothalamic AVT transcript (northern analysis) and the size of immunoreactive vasotocin (ir-AVT) neurons and hybridization signals (in the form of silver grains), representing AVT mRNA in corresponding neurons of paraventricular nuclei (PVN), increased significantly in all the treated groups compared with controls. Our findings indicate that although sex steroid administration has no effect on plasma osmolality and AVT concentration, unlike water deprivation, it may stimulate the profile/activity of AVT neurons of PVN, supporting the possibility of sex steroid receptors on these neurons. It is concluded that in quail, osmotic stress not only upregulates the expression of the AVT gene in existing neurons but also recruits many more neurons to increase the rate of AVT synthesis and secretion, while sex steroids appear to have a stimulatory effect only on the existing number of neurons and only at the level of transcription/translation and hence may influence/modulate hypothalamic AVT gene expression in response to osmotic stress. This study also suggests an interrelationship between reproduction and AVT system/function in birds. PMID- 15277559 TI - The biomechanical and neural control of hydrostatic limb movements in Manduca sexta. AB - Caterpillars are ecologically successful soft-bodied climbers. They are able to grip tightly to foliage using cuticular hooks at the tips of specialized abdominal limbs called prolegs. The neural control of proleg retraction has been examined in some detail but little is known about how prolegs extend and adduct. This is of particular interest because there are no extensor muscles or any obvious mechanisms for directing hydraulic flow into the proleg. In restrained tobacco hornworms (Manduca sexta), adduction can be evoked by stimulating mechanosensory hairs on the medial surface of the proleg. 3-D kinematics show that extension and adduction occur simultaneously through an unfolding of membrane between the pseudo segments. Hemolymph pressure pulses are not necessary to extend the proleg; instead, the pressure at the base of the proleg decreases before adduction and increases before retraction. It is proposed that these pressure changes are caused by muscles that stiffen and relax the body wall during cycles of retraction and adduction. Electromyographic recordings show that relaxation of the principal planta retractor muscle is essential for normal adduction. Extracellular nerve and muscle recordings in reduced preparations show that medial hair stimulation of one proleg can strongly and bilaterally excite motoneurons controlling the ventral internal lateral muscles of all the proleg bearing segments. Ablation, nerve section and electromyographic experiments show that this muscle is not essential for adduction in restrained larvae but that it is coactive with the retractors and may be responsible for stiffening the body wall during proleg movements. PMID- 15277560 TI - Functional roles of the transverse and longitudinal flagella in the swimming motility of Prorocentrum minimum (Dinophyceae). AB - Equations describing the motion of the dinoflagellate Prorocentrum minimum, which has both a longitudinal and a transverse flagellum, were formulated and examined using numerical calculations based on hydrodynamic resistive force theory. The calculations revealed that each flagellum has its own function in cell locomotion. The transverse flagellum works as a propelling device that provides the main driving force or thrust to move the cell along the longitudinal axis of its helical swimming path. The longitudinal flagellum works as a rudder, giving a lateral force to the cell in a direction perpendicular to the longitudinal axis of the helix. Combining these functions results a helical swimming motion similar to the observed motion. Flagellar hairs present on the transverse flagellum are necessary to make the calculated cell motion agree with the observed cell motion. PMID- 15277561 TI - Prey snapping and visual distance estimation in Texas horned lizards, Phrynosoma cornutum. AB - Captive Texas horned lizards were high-speed videotaped while feeding on ants in order to study the role of vision in facilitating tongue-protrusion capture of prey. Analysis of tongue movements revealed that prey snapping in these lizards is not a typical fixed-action pattern. By contrast, it is variable in performance and duration. Lizards adjusted head and tongue direction during the strike, within a few milliseconds, in response to movements of the prey. The duration of a typical tongue strike was 100-150 ms. The strike duration was prolonged after ophthalmic lenses were placed in front of one or both eyes. These lenses were used to investigate whether horned lizards use accommodation to judge prey distance. Focal changes of negatively powered ophthalmic lenses (employed monocularly) induced a clear underestimation of prey distance by the lizards, confirming the hypothesized expectation that accommodation is used for depth perception. The effect of the lenses was different in the two animals tested with monocular restriction. This, together with the lack of difference in responses by the lizards when untreated and when both eyes were lens covered (binocular treatment of equal power, -9 D), illustrates that horned lizards also use other visual parameters for depth perception. PMID- 15277562 TI - When vortices stick: an aerodynamic transition in tiny insect flight. AB - We have used computational fluid dynamics to study changes in lift generation and vortex dynamics for Reynolds numbers (Re) between 8 and 128. The immersed boundary method was used to model a two-dimensional wing through one stroke cycle. We calculated lift and drag coefficients as a function of time and related changes in lift to the shedding or attachment of the leading and trailing edge vortices. We find that the fluid dynamics around the wing fall into two distinct patterns. For Re> or =64, leading and trailing edge vortices are alternately shed behind the wing, forming the von Karman vortex street. For Re< or =32, the leading and trailing edge vortices remain attached to the wing during each 'half stroke'. In three-dimensional studies, large lift forces are produced by 'vortical asymmetry' when the leading edge vortex remains attached to the wing for the duration of each half stroke and the trailing edge vortex is shed. Our two-dimensional study suggests that this asymmetry is lost for Re below some critical value (between 32 and 64), resulting in lower lift forces. We suggest that this transition in fluid dynamics is significant for lift generation in tiny insects. PMID- 15277563 TI - Peripheral innervation patterns and central distribution of fin chromatophore motoneurons in the cuttlefish Sepia officinalis. AB - Body patterning behavior in unshelled cephalopod molluscs such as squid, octopuses, and cuttlefish is the ability of these animals to create complex patterns on their skin. This behavior is generated primarily by chromatophores, pigment-containing organs that are directly innervated by central motoneurons. The present study focuses on innervation patterns and location of chromatophore motoneurons in the European cuttlefish Sepia officinalis, specifically those motoneurons that control chromatophores of the fin. The fin is known to be innervated by the large, branching fin nerve. This study further characterizes the innervation of fin chromatophores by the fin nerve, generates a reference system for the location of fin nerve branches across individuals, and localizes the neurons whose axons innervate fin chromatophores through the fin nerve. Data from extracellular stimulation of fin nerve branches in intact animals demonstrate topographic innervation of fin chromatophores, while retrograde labeling data reveal the posterior subesophageal mass of the brain as the primary location of fin chromatophore motoneurons. PMID- 15277564 TI - Effect of water depth and water velocity upon the surfacing frequency of the bimodally respiring freshwater turtle, Rheodytes leukops. AB - This study examines the effect of increasing water depth and water velocity upon the surfacing behaviour of the bimodally respiring turtle, Rheodytes leukops. Surfacing frequency was recorded for R. leukops at varying water depths (50, 100, 150 cm) and water velocities (5, 15, 30 cm s(-1)) during independent trials to provide an indirect cost-benefit analysis of aquatic versus pulmonary respiration. With increasing water velocity, R. leukops decreased its surfacing frequency twentyfold, thus suggesting a heightened reliance upon aquatic gas exchange. An elevated reliance upon aquatic respiration, which presumably translates into a decreased air-breathing frequency, may be metabolically more efficient for R. leukops compared to the expenditure (i.e. time and energy) associated with air-breathing within fast-flowing riffle zones. Additionally, R. leukops at higher water velocities preferentially selected low-velocity microhabitats, presumably to avoid the metabolic expenditure associated with high water flow. Alternatively, increasing water depth had no effect upon the surfacing frequency of R. leukops, suggesting little to no change in the respiratory partitioning of the species across treatment settings. Routinely long dives (>90 min) recorded for R. leukops indicate a high reliance upon aquatic O2 uptake regardless of water depth. Moreover, metabolic and temporal costs attributed to pulmonary gas exchange within a pool-like environment were likely minimal for R. leukops, irrespective of water depth. PMID- 15277565 TI - How the house sparrow Passer domesticus absorbs glucose. AB - According to the hypothesis that most glucose absorption occurs passively across intestinal tight junctions (paracellular absorption), one would predict fairly similar rates of in vivo absorption of L-glucose, the stereoisomer of D-glucose that is absorbed only passively and is not catabolized, and of 3-O-methyl-D glucose (3OMD-glucose), the D-glucose analogue that is actively and passively transported and not catabolized. In house sparrows Passer domesticus, we applied a pharmacokinetic method to measure simultaneous in vivo absorption of [14C]L glucose and [3H]3OMD-glucose in a situation in which intestinal glucose transporters were relatively saturated (gavage solution contained 200 mmol l(-1) 3OMD-glucose). Fractional absorptions (F) were not significantly different between [3H]3OMD- and [14C]L-glucose (0.80 vs 0.79), and the apparent rates of absorption did not differ significantly. When we performed the same experiment on other sparrows in a situation in which intestinal glucose transporters were relatively unsaturated (200 mmol l(-1) mannitol replaced 3OMD-glucose in the gavage solution), the apparent rate of absorption was significantly reduced for [14C]l-glucose by 39% and for [3H]3OMD-glucose by 26%. A simulation model showed that a reduction is not predicted if most of the [3H]3OMD-glucose is actively absorbed, because the absorption rate of the tracer should increase when competitive inhibitor (unlabeled 3OMD-glucose) is removed. The similar extent and rates of absorption of [3H]3OMD- and [14C]L-glucose, and the acceleration of their rates of absorption in the presence of luminal 3OMD-glucose, are most consistent with Pappenheimer's hypothesis that the majority of dietary D-glucose is absorbed passively. PMID- 15277566 TI - Antisecretory factor expression is regulated by inflammatory mediators and influences the severity of experimental autoimmune encephalomyelitis. AB - Antisecretory factor (ASF) was originally identified as a potent inhibitor of intestinal fluid secretion induced by a number of enterotoxins. In addition to its involvement in intestinal fluid secretion, ASF modulates the proliferation of memory/effector T cells and is expressed by cells of the immune system. This report describes the role of ASF in modulating immune responses and assesses the regulation of ASF during an in vivo immunological reaction. ASF expression was redistributed during adoptively transferred experimental autoimmune encephalomyelitis (EAE), and in response to other inflammatory stimuli. Administration of the anti-ASF antibody TLD-1A8A increased the clinical severity and duration of the disease. Consistent with these findings, addition of TLD-1A8A to T cell proliferation assays resulted in up-regulation of the proinflammatory cytokines IL-18 and IL-6 and in down-regulation of IL-10. Furthermore, we identified cytokines that regulated the expression of ASF at both the mRNA and protein level. ASF, therefore, appears to play a previously unappreciated and potentially important role in the regulation of immune responses. PMID- 15277567 TI - MC-3 receptor and the inflammatory mechanisms activated in acute myocardial infarct. AB - Investigation of the mechanisms activated by endogenous inhibitory pathways can lead to identification of novel targets for cardiovascular inflammatory pathologies. Here we exploited the potential protective role that melanocortin receptor type 3 (MC3-R) activation might play in a myocardial ischemia reperfusion injury model. In resting conditions, mouse and rat heart extracts expressed MC3-R mRNA and protein, without changes following ischemia-reperfusion. At the cellular level heart macrophages, but not fibroblasts or cardiomyocytes, expressed this receptor, as demonstrated by immunogold labeling. In vivo, administration of the melanocortin agonist MTII (10 microg per mouse equivalent to 9.3 nmol) 30 min prior to ischemia (25 min) attenuated mouse heart 2 h reperfusion injury by approximately 40%, an effect prevented by the mixed MC3/4-R antagonist SHU9119 but not by the selective MC4-R antagonist HS204. Similar results were obtained when the compound was given at the beginning of the reperfusion period. Importantly, delayed myocardial damage as measured 24 h post reperfusion was equally protected by administration of 10 microg MTII. The focus on MC3-R was also substantiated by analysis of the recessive yellow (e/e) mouse, bearing a mutated (inactive) MC1-R, in which MTII was fully protective. Myocardial protection was associated with reduced markers of systemic and local inflammation, including cytokine contents (interleukin-1 and KC) and myeloperoxidase activity. In conclusion, this study has highlighted a previously unrecognized protective role for MC3-R activation on acute and delayed heart reperfusion injury. These data may open new avenues for therapeutic intervention against heart and possibly other organ ischemia-reperfusion injury. PMID- 15277568 TI - Polymorphonuclear leukocytes from patients with severe sepsis have lost the ability to degrade fibrin via u-PA. AB - Fibrin persistence in the vasculature is an important complication of sepsis that can often lead to mortality. We have previously established that polymorphonuclear leukocytes (PMN) from healthy individuals have the capacity to degrade fibrin via urokinase-type plasminogen activator (u-PA). We have also demonstrated an increase in u-PA antigen in the plasma of patients suffering from septic shock. In this study, we investigate the hypothesis that PMN from patients with sepsis have lost their fibrinolytic ability and that this might contribute to the persistence of fibrin deposits. We show here that PMN from these patients do not express any u-PA activity, despite retaining some u-PA antigen. Additionally, thrombi prepared from the whole blood of the patients exhibit reduced endogenous lysis compared with those from healthy individuals. These data indicate that loss of fibrinolytic activity from PMN may be a contributing factor in fibrin persistence in the microvasculature in sepsis. PMID- 15277569 TI - Plasminogen activator inhibitor-2 (PAI-2) in eosinophilic leukocytes. AB - Plasminogen activator inhibitor-2 (PAI-2) as a potential eosinophil protein was inferred from our gene microarray study of mouse eosinophilopoiesis. Here, we detect 47 kDa intracellular and approximately 60 kDa secretory forms of PAI-2 in purified human eosinophil extracts. PAI-2 is present at variable concentrations in eosinophil lysates, ranging from 30 to 444 ng/10(6) cells, with a mean of 182 ng/10(6) cells from 10 normal donors, which is the highest per-cell concentration among all leukocyte subtypes evaluated. Enzymatic assay confirmed that eosinophil derived PAI-2 is biologically active and inhibits activation of its preferred substrate, urokinase. Immunohistochemical and immunogold staining demonstrated PAI-2 localization in eosinophil-specific granules. Immunoreactive PAI-2 was detected in extracellular deposits in and around the eosinophil-enriched granuloma tissue encapsulating the parasitic egg in livers of wild-type mice infected with the helminthic parasite Schistosoma mansoni. Among the possibilities, we consider a role for eosinophil-derived PAI-2 in inflammation and remodeling associated with parasitic infection as well as allergic airways disease, respiratory virus infection, and host responses to tumors and metastasis in vivo. PMID- 15277570 TI - Insulin-like growth factor-I stimulates IL-10 production in human T cells. AB - There is vast body of evidence that the insulin-like growth factor (IGF)-I exerts immunomodulatory effects in vitro and in vivo. In vitro studies indicate that stimulatory effects of IGF-I may be exerted through augmentation of inflammatory cytokine production. To further explore the immunomodulatory effects of IGF-I through regulation of cytokine production, we tested the in vitro effects of IGF I on the secretion of inflammatory T helper cell type 1 (Th1) and Th2 cytokines by human peripheral blood mononuclear cells (PBMC). To this end, PBMC were stimulated with the T cell mitogen phytohemagglutinin (PHA), and cytokines in the culture media were assessed after 18, 42, 66, and 80 h of culture. We found that IGF-I stimulated the secretion of the Th2 cytokine interleukin (IL)-10 by 40-70% in PHA-stimulated PBMC. In addition, we observed a small stimulatory effect (15%) on the secretion of another Th2 cytokine IL-4. The secretion of IL-2, IL-5, IL-6, interferon-gamma, and the inflammatory cytokines IL-1beta, IL-8, and tumor necrosis factor alpha was not or was hardly affected. IL-10 secretion was also stimulated in purified T cells, and we established that IGF-I also stimulated IL 10 mRNA expression by 100-150%. The monocyte-activating bacterial cell-wall product lipopolysaccharide induced IL-10 production in PBMC, but this was not affected by IGF-I. As IL-10 predominantly exerts anti-inflammatory actions and suppresses Th1-dependent immune responses, our results indicate that IGF-I may exert inhibitory actions on inflammatory and Th1-mediated cellular immune responses through stimulation of IL-10 production in T cells. PMID- 15277571 TI - Towards understanding the molecular mechanism of sperm chemotaxis. PMID- 15277572 TI - CFTR channel pharmacology: novel pore blockers identified by high-throughput screening. PMID- 15277573 TI - Revisiting the role of H+ in chemotactic signaling of sperm. AB - Chemotaxis of sperm is an important step toward fertilization. During chemotaxis, sperm change their swimming behavior in a gradient of the chemoattractant that is released by the eggs, and finally sperm accumulate near the eggs. A well established model to study chemotaxis is the sea urchin Arbacia punctulata. Resact, the chemoattractant of Arbacia, is a peptide that binds to a receptor guanylyl cyclase. The signaling pathway underlying chemotaxis is still poorly understood. Stimulation of sperm with resact induces a variety of cellular events, including a rise in intracellular pH (pHi) and an influx of Ca2+; the Ca2+ entry is essential for the chemotactic behavior. Previous studies proposed that the influx of Ca2+ is initiated by the rise in pHi. According to this proposal, a cGMP-induced hyperpolarization activates a voltage-dependent Na+/H+ exchanger that expels H+ from the cell. Because some aspects of the proposed signaling pathway are inconsistent with recent results (Kaupp, U.B., J. Solzin, J.E. Brown, A. Helbig, V. Hagen, M. Beyermann, E. Hildebrand, and I. Weyand. 2003. Nat. Cell Biol. 5:109-117), we reexamined the role of protons in chemotaxis of sperm using kinetic measurements of the changes in pHi and intracellular Ca2+ concentration. We show that for physiological concentrations of resact (<25 pM), the influx of Ca2+ precedes the rise in pHi. Moreover, buffering of pHi completely abolishes the resact-induced pHi signal, but leaves the Ca2+ signal and the chemotactic motor response unaffected. We conclude that an elevation of pHi is required neither to open Ca(2+)-permeable channels nor to control the chemotactic behavior. Intracellular release of cGMP from a caged compound does not cause an increase in pHi, indicating that the rise in pHi is induced by cellular events unrelated to cGMP itself, but probably triggered by the consumption and subsequent replenishment of GTP. These results show that the resact-induced rise in pHi is not an obligatory step in sperm chemotactic signaling. A rise in pHi is also not required for peptide-induced Ca2+ entry into sperm of the sea urchin Strongylocentrotus purpuratus. Speract, a peptide of S. purpuratus may act as a chemoattractant as well or may serve functions other than chemotaxis. PMID- 15277574 TI - Discovery of glycine hydrazide pore-occluding CFTR inhibitors: mechanism, structure-activity analysis, and in vivo efficacy. AB - The cystic fibrosis transmembrane conductance regulator (CFTR) protein is a cAMP regulated epithelial Cl- channel that, when defective, causes cystic fibrosis. Screening of a collection of 100,000 diverse small molecules revealed four novel chemical classes of CFTR inhibitors with Ki < 10 microM, one of which (glycine hydrazides) had many active structural analogues. Analysis of a series of synthesized glycine hydrazide analogues revealed maximal inhibitory potency for N (2-naphthalenyl) and 3,5-dibromo-2,4-dihydroxyphenyl substituents. The compound N (2-naphthalenyl)-[(3,5-dibromo-2,4-dihydroxyphenyl)methylene]glycine hydrazide (GlyH-101) reversibly inhibited CFTR Cl- conductance in <1 min. Whole-cell current measurements revealed voltage-dependent CFTR block by GlyH-101 with strong inward rectification, producing an increase in apparent inhibitory constant Ki from 1.4 microM at +60 mV to 5.6 microM at -60 mV. Apparent potency was reduced by lowering extracellular Cl- concentration. Patch-clamp experiments indicated fast channel closures within bursts of channel openings, reducing mean channel open time from 264 to 13 ms (-60 mV holding potential, 5 microM GlyH 101). GlyH-101 inhibitory potency was independent of pH from 6.5-8.0, where it exists predominantly as a monovalent anion with solubility approximately 1 mM in water. Topical GlyH-101 (10 microM) in mice rapidly and reversibly inhibited forskolin-induced hyperpolarization in nasal potential differences. In a closed loop model of cholera, intraluminal GlyH-101 (2.5 microg) reduced by approximately 80% cholera toxin-induced intestinal fluid secretion. Compared with the thiazolidinone CFTR inhibitor CFTR(inh)-172, GlyH-101 has substantially greater water solubility and rapidity of action, and a novel inhibition mechanism involving occlusion near the external pore entrance. Glycine hydrazides may be useful as probes of CFTR pore structure, in creating animal models of CF, and as antidiarrheals in enterotoxic-mediated secretory diarrheas. PMID- 15277575 TI - Toll-like receptor 4 (TLR4) of retinal pigment epithelial cells participates in transmembrane signaling in response to photoreceptor outer segments. AB - Retinal pigment epithelial (RPE) cells mediate the recognition and clearance of effete photoreceptor outer segments (POS), a process central to the maintenance of normal vision. Given the emerging importance of Toll-like receptors (TLRs) in transmembrane signaling in response to invading pathogens as well as endogenous substances, we hypothesized that TLRs are associated with RPE cell management of POS. TLR4 clusters on human RPE cells in response to human, but not bovine, POS. However, TLR4 clustering could be inhibited by saturating concentrations of an inhibitory anti-TLR4 mAb. Furthermore, human POS binding to human RPE cells elicited transmembrane metabolic and calcium signals within RPE cells, which could be blocked by saturating doses of an inhibitory anti-TLR4 mAb. However, the heterologous combination of bovine POS and human RPE did not trigger these signals. The pattern recognition receptor CD36 collected at the POS-RPE cell interface for both homologous and heterologous samples, but human TLR4 only collected at the human POS-human RPE cell interface. Kinetic experiments of human POS binding to human RPE cells revealed that CD36 arrives at the POS-RPE interface followed by TLR4 accumulation within 2 min. Metabolic and calcium signals immediately follow. Similarly, the production of reactive oxygen metabolites (ROMs) was observed for the homologous human system, but not the heterologous bovine POS-human RPE cell system. As (a) the bovine POS/human RPE combination did not elicit TLR4 accumulation, RPE signaling, or ROM release, (b) TLR4 arrives at the POS-RPE cell interface just before signaling, (c) TLR4 blockade with an inhibitory anti-TLR4 mAb inhibited TLR4 clustering, signaling, and ROM release in the human POS-human RPE system, and (d) TLR4 demonstrates similar clustering and signaling responses to POS in confluent RPE monolayers, we suggest that TLR4 of RPE cells participates in transmembrane signaling events that contribute to the management of human POS. PMID- 15277576 TI - Gating transitions in bacterial ion channels measured at 3 microns resolution. AB - Ion channels of high conductance (>200 pS) are widespread among prokaryotes and eukaryotes. Two examples, the Escherichia coli mechanosensitive ion channels Ec MscS and Ec-MscL, pass currents of 125-300 pA. To resolve temporal details of conductance transitions, a patch-clamp setup was optimized for low-noise recordings at a time resolution of 3 microns (10-20 times faster than usual). Analyses of the high-resolution recordings confirm that Ec-MscL visits many subconductance states and show that most of the intersubstate transitions occur more slowly than the effective resolution of 3 micros. There is a clear trend toward longer transition times for the larger transitions. In Ec-MscS recordings, the majority of the observed full conductance transitions are also composite. We detected a short-lived (approximately 20 microns) Ec-MscS substate at 2/3 of full conductance; transitions between 2/3 and full conductance did not show fine structure and had a time course limited by the achieved resolution. Opening and closing transitions in MscS are symmetrical and are not preceded or followed by smaller, rapid currents ("anticipations" or "regrets"). Compared with other, lower-conductance channels, these measurements may detect unusually early states in the transitions from fully closed to fully open. Increased temporal resolution at the single-molecule level reveals that some elementary steps of structural transitions are composite and follow several alternative pathways, while others still escape resolution. High-bandwidth, low-noise single-channel measurements may provide details about state transitions in other high-conductance channels; and similar procedures may also be applied to channel- and nanopore-based single molecule DNA measurements. PMID- 15277577 TI - Can Shaker potassium channels be locked in the deactivated state? AB - For structural studies it would be useful to constrain the voltage sensor of a voltage-gated channel in its deactivated state. Here we consider one Shaker potassium channel mutant and speculate about others that might allow the channel to remain deactivated at zero membrane potential. Ionic and gating currents of F370C Shaker, expressed in Xenopus oocytes, were recorded in patches with internal application of the methanethiosulfonate reagent MTSET. It appears that the voltage dependence of voltage sensor movement is strongly shifted by reaction with internal MTSET, such that the voltage sensors appear to remain deactivated even at positive potentials. A disadvantage of this construct is that the rate of modification of voltage sensors by MTSET is quite low, approximately 0.17 mM( 1).s(-1) at -80 mV, and is expected to be much lower at depolarized potentials. PMID- 15277578 TI - Regulation of K+ flow by a ring of negative charges in the outer pore of BKCa channels. Part I: Aspartate 292 modulates K+ conduction by external surface charge effect. AB - The pore region of the majority of K+ channels contains the highly conserved GYGD sequence, known as the K+ channel signature sequence, where the GYG is critical for K+ selectivity (Heginbotham, L., T. Abramson, and R. MacKinnon. 1992. Science. 258:1152-1155). Exchanging the aspartate residue with asparagine in this sequence abolishes ionic conductance of the Shaker K+ channel (D447N) (Hurst, R.S., L. Toro, and E. Stefani. 1996. FEBS Lett. 388:59-65). In contrast, we found that the corresponding mutation (D292N) in the pore forming alpha subunit (hSlo) of the voltage- and Ca(2+)-activated K+ channel (BKCa, MaxiK) did not prevent conduction but reduced single channel conductance. We have investigated the role of outer pore negative charges in ion conduction (this paper) and channel gating (Haug, T., R. Olcese, T. Ligia, and E. Stefani. 2004. J. Gen Physiol. 124:185 197). In symmetrical 120 mM [K+], the D292N mutation reduced the outward single channel conductance by approximately 40% and nearly abolished inward K+ flow (outward rectification). This rectification was partially relieved by increasing the external K+ concentration to 700 mM. Small inward currents were resolved by introducing an additional mutation (R207Q) that greatly increases the open probability of the channel. A four-state multi-ion pore model that incorporates the effects of surface charge was used to simulate the essential properties of channel conduction. The conduction properties of the mutant channel (D292N) could be predicted by a simple approximately 8.5-fold reduction of the surface charge density without altering any other parameter. These results indicate that the aspartate residue in the BKCa pore plays a key role in conduction and suggest that the pore structure is not affected by the mutation. We speculate that the negative charge strongly accumulates K+ in the outer vestibule close to the selectivity filter, thus increasing the rate of ion entry into the pore. PMID- 15277580 TI - Amplifying cancer vaccine responses by modifying pathogenic gene programs in tumor cells. AB - Immunosuppressive factors, such as vascular endothelial growth factor, transforming growth factor-beta, prostaglandin E2, interleukin (IL)-10, and IL-6, are made frequently by cancer cells. These factors, along with others, can inhibit the development and function of tumor-reactive effector T cells and the clinical results of cancer vaccines. Production of these factors by tumor cells is associated with disease progression and may represent an active immune surveillance escape mechanism. However, a number of factors appear to be made directly in response to signaling molecules, such as RAS, AKT, and signal transducer and activator of transcription 3, which are activated as a result of genetic events that occur during oncogenesis. Methods to overcome the negative effects of immunosuppressive factors, which are "hard wired" into gene programs of cancer cells, might then improve the results of cancer vaccines. For example, specific blocking antibodies, which recognize such factors, or kinase inhibitors, which block the signaling pathways that lead to their production, could potentially be used as vaccine adjuvants. The effects of immunosuppressive factors may also be "turned off" by cytokines with tumor suppressor properties. The enhanced clinical and immunological effects of melanoma vaccines observed after the administration of high doses of interferon-alpha2b provide a "proof of principle" in human patients, that agents which counter the gene programs of cancer cells, causing them to intrinsically resist tumor-reactive T cells, may improve significantly the efficacy of cancer vaccines. PMID- 15277579 TI - Regulation of K+ flow by a ring of negative charges in the outer pore of BKCa channels. Part II: Neutralization of aspartate 292 reduces long channel openings and gating current slow component. AB - Neutralization of the aspartate near the selectivity filter in the GYGD pore sequence (D292N) of the voltage- and Ca(2+)-activated K+ channel (MaxiK, BKCa) does not prevent conduction like the corresponding mutation in Shaker channel, but profoundly affects major biophysical properties of the channel (Haug, T., D. Sigg, S. Ciani, L. Toro, E. Stefani, and R. Olcese. 2004. J. Gen. Physiol. 124:173-184). Upon depolarizations, the D292N mutant elicited mostly gating current, followed by small or no ionic current, at voltages where the wild-type hSlo channel displayed robust ionic current. In fact, while the voltage dependence of the gating current was not significantly affected by the mutation, the overall activation curve was shifted by approximately 20 mV toward more depolarized potentials. Several lines of evidence suggest that the mutation prevents population of certain open states that in the wild type lead to high open probability. The activation curves of WT and D292N can both be fitted to the sum of two Boltzmann distributions with identical slope factors and half activation potentials, just by changing their relative amplitudes. The steeper and more negative component of the activation curve was drastically reduced by the D292N mutation (from 0.65 to 0.30), suggesting that the population of open states that occurs early in the activation pathway is reduced. Furthermore, the slow component of the gating current, which has been suggested to reflect transitions from closed to open states, was greatly reduced in D292N channels. The D292N mutation also affected the limiting open probability: at 0 mV, the limiting open probability dropped from approximately 0.5 for the wild-type channel to 0.06 in D292N (in 1 mM [Ca2+]i). In addition to these effects on gating charge and open probability, as already described in Part I, the D292N mutation introduces a approximately 40% reduction of outward single channel conductance, as well as a strong outward rectification. PMID- 15277581 TI - ABT-963 [2-(3,4-difluoro-phenyl)-4-(3-hydroxy-3-methyl-butoxy)-5-(4 methanesulfonyl-phenyl)-2H-pyridazin-3-one], a highly potent and selective disubstituted pyridazinone cyclooxgenase-2 inhibitor. AB - Nonsteriodal anti-inflammatory drugs (NSAIDs) are efficacious for the treatment of pain associated with inflammatory disease. Clinical experience with marketed selective cyclooxygenase-2 (COX-2) inhibitors (celecoxib, rofecoxib, and valdecoxib) has confirmed the utility of these agents in the treatment of inflammatory pain with an improved gastrointestinal safety profile relative to NSAID comparators. These COX-2 inhibitors belong to the same structural class. Each contains a core heterocyclic ring with two appropriately substituted phenyl rings appended to adjacent atoms. Here, we report the identification of vicinally disubstituted pyridazinones as potent and selective COX-2 inhibitors. The lead compound in the series, ABT-963 [2-(3,4-difluoro-phenyl)-4-(3-hydroxy-3-methyl butoxy)-5-(4-methanesulfonyl-phenyl)-2H-pyridazin-3-one], has excellent selectivity (ratio of 276, COX-2/COX-1) in human whole blood, improved aqueous solubility compared with celecoxib and rofecoxib, high oral anti-inflammatory potency in vivo, and gastric safety in the animal studies. After oral administration, ABT-963 reduced prostaglandin E2 production in the rat carrageenan air pouch model (ED50 of 0.4 mg/kg) and reduced the edema in the carrageenan induced paw edema model with an ED30 of 1.9 mg/kg. ABT-963 dose dependently reduced nociception in the carrageenan hyperalgesia model (ED50 of 3.1 mg/kg). After 14 days of dosing in the adjuvant arthritis model, ABT-963 had an ED(50) of 1.0 mg/kg in reducing the swelling of the hind paws. Magnetic resonance imaging examination of the diseased paws in the adjuvant model showed that ABT-963 significantly reduced bone loss and soft tissue destruction. ABT-963 is a highly selective COX-2 inhibitor that may have utility in the treatment of the pain and inflammation associated with arthritis. PMID- 15277582 TI - A novel nonpeptide antagonist of the kinin B1 receptor: effects at the rabbit receptor. AB - The kinin B1 receptor (B1R) has attracted interest as a potential therapeutic target because this inducible G protein-coupled receptor is involved in sustained inflammation and inflammatory pain production. Compound 11 (2-[(2R)-1-[(3,4 dichlorophenyl) sulfonyl]-3-oxo-1,2,3,4-tetrahydroquinoxalin-2-yl]-N-[2-[4-(4,5 dihydro-1H-imidazol-2-yl)phenyl]ethyl]acetamide) is a high-affinity nonpeptide antagonist for the human B1R, but it is potent at the rabbit B1R as well: its Ki value for the inhibition of [3H]Lys-des-Arg9-BK (bradykinin) binding to a novel myc-labeled rabbit B1R expressed in COS-1 is 22 pM. In contractility tests (organ bath pharmacology), we found that compound 11 is an apparently surmountable antagonist of des-Arg9-BK- or Lys-des-Arg9-BK-induced contraction of the rabbit isolated aorta (pA2 values of 10.6+/-0.14 and 10.4+/-0.12, respectively). It did not influence contractions induced by angiotensin II in the rabbit aorta or by BK or histamine in the jugular vein, but it suppressed the prostaglandin-mediated relaxant effect of des-Arg9-BK on the rabbit isolated mesenteric artery. Compound 11 (1 nM) inhibited both the phosphorylation of the extracellular signal regulated kinase1/2 mitogen-activated protein kinases induced by Lys-des-Arg9-BK in serum-starved rabbit aortic smooth muscle cells and the agonist-induced translocation of the fusion protein B1R-yellow fluorescent protein expressed in human embryonic kidney (HEK) 293 cells. Compound 11 does not importantly modify the expression of myc-B1R over 24 h in HEK 293 cells (no detectable action as "pharmacological chaperone"). The present results support that compound 11 is a potent and highly selective antagonist suitable for further investigations of the role of the kinin B1R in models of inflammation, pain, and sepsis based on the rabbit. PMID- 15277584 TI - Bridging disciplines: an introduction to the special issue on public health and pediatric psychology. PMID- 15277583 TI - Inhibition of tumor cell proliferation by sigma ligands is associated with K+ Channel inhibition and p27kip1 accumulation. AB - Previous studies have shown that sigma receptors are overexpressed in tumor cells. However, the role of sigma receptors remains enigmatic. Recently, we and others have demonstrated that sigma-1 receptor modulates K+ channels in pituitary. In the present report, patch-clamp and Western blot assays were used in small cell lung cancer (SCLC, NCI-H209, and NCI-H146) and leukemic (Jurkat) cell lines to investigate the effects of sigma ligands on voltage-gated K+ channels and cell proliferation. The sigma ligands (+)-pentazocine, igmesine, and 1,3-di(2-tolyl)guanidine (DTG) all reversibly inhibited voltage-activated K+ currents in both cell lines. The potency of sigma ligand-induced inhibition (10 microM) was igmesine = (+)-pentazocine > DTG, pointing to the involvement of sigma-1 receptors. Addition of the K+ channel blockers tetraethylammonium (TEA) and 4-aminopyridin or one of cited sigma ligands in the culture media reversibly inhibited Jurkat cell growth. Interestingly, K+ channel blockers and sigma ligands caused an accumulation of the cyclin-dependent kinase inhibitor p27kip1 and a decrease in cyclin A expression in Jurkat and SCLC cells, whereas no effect could be detected on p21cip1. Moreover, sigma ligands and TEA had no effect on caspase 3 activity. Accordingly, incubation of cells with sigma ligands did not provoke DNA laddering. These data demonstrate that sigma ligands and voltage dependent channel blockers inhibit cell growth through a cell cycle arrest in the G1 phase but not via an apoptotic mechanism. Altogether, these results indicate that the sigma-1 receptor-induced inhibition of the cell cycle is, at least in part, the consequence of the inhibition of K+ channels. PMID- 15277585 TI - Understanding toddlers' in-home injuries: I. Context, correlates, and determinants. AB - Multimethod strategies (i.e., questionnaires, parents' observations, injury-event recording diaries, telephone and home interviews) were used to study in-home injuries experienced by toddlers over a 3-month period. Cuts, scrapes, and puncture wounds were the most common injuries. The majority of injuries affected children's limbs, and injuries most often occurred in the morning. Boys were injured most often in rooms designated for play, and a majority of their injuries followed from misbehavior. Girls were most often injured in nonplay areas of the home, with the majority of injuries occurring during play activities. Boys experienced more frequent and severe injuries than girls, although girls reacted more than boys to their injuries. Child factors relevant to injury included: risk taking, sensation seeking, and ease of behavior management. Temperament factors did not relate to child injury. Parent factors relevant to child injury included parents' beliefs about control over their child's health, protectiveness, and beliefs about child supervision. Regression analyses revealed that both child (i.e., risk taking) and parent (i.e., protectiveness) factors were significant determinants of child injury. PMID- 15277586 TI - Understanding toddlers' in-home injuries: II. Examining parental strategies, and their efficacy, for managing child injury risk. AB - Multimethod strategies (i.e., questionnaires, injury-event recording diaries, and telephone and home interviews) were used to study in-home injuries experienced by toddlers over a 3-month period and to identify anticipatory prevention strategies implemented by parents, on a room-by-room basis, that effectively reduced child injury risk. Three types of prevention strategies were used by parents: environmental (e.g., hazard removal, safety devices to prevent access), parental (e.g., increased supervision, parent modification of their own behavior to decrease injury risk for their child), and child based (e.g., teaching rules or prohibitions to promote safety), with parents often using a combination of these. Use of these strategies, and their efficacy to reduce injury risk, varied on a room-by-room basis. Nonetheless, two general conclusions are supported: (1) An emphasis on child-based strategies never decreases, and often elevates, risk of injury to toddlers; and (2) parental and environmental strategies, either singularly or in combination, serve protective functions that significantly reduce children's risk of in-home injury. Although it is commonplace for parents of children between 2 and 3 years of age to transition from environmental and supervision strategies to the use of teaching and rule-based ones to manage injury risk, doing so too early clearly elevates children's risk of injury in the home. PMID- 15277587 TI - Perceived risk, risk taking, estimation of ability and injury among adolescent sport participants. AB - OBJECTIVE: To determine the predictive validity of perceived risk, risk taking, estimation of ability, overefficacy, and previous injuries on actual injury among adolescents in sport; and to examine sex differences on these factors. METHODS: A cohort of 260 (148 male, 112 female) soccer players aged 11 to 14 years participated in a 3-month prospective injury study. Preseason written measures included self-reported perceived risk, previous injuries, risk taking and estimation of ability. RESULTS: Low levels of perceived risk and estimation of ability were associated with a significant increase in risk of injury, with odds ratios (ORs) ranging from 3.77-7.92. Positive relationships between injury and both estimation of ability and overestimation of ability were supported. Estimation of ability was also positively related to risk taking. In this study, however, risk taking was not directly related to injury, nor were previous injuries. Girls reported higher levels of perceived risk and lower levels of risk taking than boys. However, boys and girls reported similar estimation of ability and overestimation of ability and subsequently incurred similar numbers of injuries. CONCLUSIONS: Perceived risk and estimation of ability represent significant psychological risk factors for injury in adolescent sports. Sex differences in perceived risk, risk taking, and previous injuries should be considered when developing environmental and behavioral injury-prevention programs. PMID- 15277588 TI - Brief report: the adaptation of Project Northland for urban youth. AB - OBJECTIVE: Summarizes the research and intervention design of a new trial to evaluate an adaptation of Project Northland, a multicomponent, community-wide alcohol prevention program for culturally diverse youth living in a large city. The original Project Northland was successful in reducing alcohol use among a sample of mostly White, rural adolescents. METHODS: We highlight the steps taken to adapt the intervention strategies for culturally diverse inner-city youth, families, and neighborhoods. The research design is a randomized controlled trial to evaluate the effectiveness of the adapted Project Northland for reducing the early onset and prevalence of alcohol use among young urban adolescents. CONCLUSION: The information gained from this trial, including the process of adaptation of prevention strategies, will be beneficial for alcohol-use prevention efforts within diverse urban communities across the country. PMID- 15277589 TI - Diffusion of an integrated health education program in an urban school system: planet health. AB - OBJECTIVE: Assessed the feasibility, acceptability, and sustainability of Planet Health, an interdisciplinary, integrated health education curriculum implemented in six public middle schools. METHODS: Workshops on Planet Health implementation were attended by 129 teachers (language arts, math, science, and social studies) over three school years (1999-2000, 2000-2001, and 2001-2002). Questionnaires were administered post-implementation and in the fall and spring of each year. Outcomes were dose, acceptability, feasibility, and intent to continue use. RESULTS: The average number of lessons taught per teacher per year was 1.7 to 3.1, compared to a goal of 2 to 3. Each year, teachers reported high acceptability and perceived feasibility of the intervention, and the majority indicated they intended to continue using the curriculum. CONCLUSIONS: Planet Health was feasible and acceptable in a participatory research model involving a public school-university partnership, and it was also sustainable independent of the research effort. PMID- 15277591 TI - Is resurgent Na+ current an alpha-subunit-specific property? Maybe not. Focus on "Sodium currents in subthalamic nucleus neurons from Nav1.6-null mice". PMID- 15277592 TI - Respiratory-like rhythmic activity can be produced by an excitatory network of non-pacemaker neuron models. AB - It is still unclear whether the respiratory-like rhythm observed in slice preparations containing the pre-Botzinger complex is of pacemaker or network origin. The rhythm persists in the absence of inhibition, but blocking pacemaker activity did not always result in rhythm abolition. We developed a computational model of the slice to show that respiratory-like rhythm can emerge as a network property without pacemakers or synaptic inhibition. The key currents of our model cell are the low- and high-threshold calcium currents and the calcium-dependent potassium current. Depolarization of a single unit by current steps or by raising the external potassium concentration can induce periodic bursting activity. Gaussian stimulation increased the excitability of the model without evoking oscillatory activity, as indicated by autocorrelation analysis. In response to hyperpolarizing pulses, the model produces prolonged relative refractory periods. At the network level, an increase of external potassium concentration triggers rhythmic activity that can be attributed to cellular periodic bursting, network properties, or both, depending on different parameters. Gaussian stimulation also induces rhythmic activity that depends solely on network properties. In all cases, the calcium-dependent potassium current has a central role in burst termination and interburst duration. However, when periodic inhibition is considered, the activation of this current is responsible for the characteristic amplification ramp of the emerged rhythm. Our results may explain controversial results from studies blocking pacemakers in vitro and show a shift in the role of the calcium-dependent potassium current in the presence of network inhibition. PMID- 15277593 TI - Light-evoked oscillatory discharges in retinal ganglion cells are generated by rhythmic synaptic inputs. AB - In the visual system, optimal light stimulation sometimes generates gamma-range (ca. 20 approximately 80 Hz) synchronous oscillatory spike discharges. This phenomenon is assumed to be related to perceptual integration. Applying a planar multi-electrode array to the isolated frog retina, Ishikane et al. demonstrated that dimming detectors, off-sustained type ganglion cells, generate synchronous oscillatory spike discharges in response to diffuse dimming illumination. In the present study, applying the whole cell current-clamp technique to the isolated frog retina, we examined how light-evoked oscillatory spike discharges were generated in dimming detectors. Light-evoked oscillatory ( approximately 30 Hz) spike discharges were triggered by rhythmic ( approximately 30 Hz) fluctuations superimposed on a depolarizing plateau potential. When a suprathreshold steady depolarizing current was injected into a dimming detector, only a few spikes were evoked at the stimulus onset. However, repetitive spikes were triggered by a gamma-range sinusoidal current superimposed on the steady depolarizing current. Thus the light-evoked rhythmic fluctuations are likely to be generated presynaptically. The light-evoked rhythmic fluctuations were suppressed not by intracellular application of N-(2,6-dimethyl phenylcarbamoylmethyl)triethylammonium bromide (QX-314), a Na(+) channel blocker, to the whole cell clamped dimming detector but by bath-application of tetrodotoxin to the retina. The light-evoked rhythmic fluctuations were suppressed by a GABA(A) receptor antagonist but potentiated by a GABA(C) receptor antagonist, whereas these fluctuations were little affected by a glycine receptor antagonist. Because amacrine cells are spiking neurons and because GABA is one of the main transmitters released from amacrine cells, amacrine cells may participate in generating rhythmically fluctuated synaptic input to dimming detectors. PMID- 15277594 TI - Phasic stimuli evoke precisely timed spikes in intermittently discharging mitral cells. AB - Mitral cells, the principal cells of the olfactory bulb, respond to sensory stimulation with precisely timed patterns of action potentials. By contrast, the same neurons generate intermittent spike clusters with variable timing in response to simple step depolarizations. We made whole cell recordings from mitral cells in rat olfactory bulb slices to examine the mechanisms by which normal sensory stimuli could generate precisely timed spike clusters. We found that individual mitral cells fired clusters of action potentials at 20-40 Hz, interspersed with periods of subthreshold membrane potential oscillations in response to depolarizing current steps. TTX (1 microM) blocked a sustained depolarizing current and fast subthreshold oscillations in mitral cells. Phasic stimuli that mimic trains of slow excitatory postsynaptic potentials (EPSPs) that occur during sniffing evoked precisely timed spike clusters in repeated trials. The amplitude of the first simulated EPSP in a train gated the generation of spikes on subsequent EPSPs. 4-aminopyridine (4-AP)-sensitive K(+) channels are critical to the generation of spike clusters and reproducible spike timing in response to phasic stimuli. Based on these results, we propose that spike clustering is a process that depends on the interaction between a 4-AP-sensitive K(+) current and a subthreshold TTX-sensitive Na(+) current; interactions between these currents may allow mitral cells to respond selectively to stimuli in the theta frequency range. These intrinsic properties of mitral cells may be important for precisely timing spikes evoked by phasic stimuli that occur in response to odor presentation in vivo. PMID- 15277595 TI - Influence of age on adaptability of human mastication. AB - The objective of this work was to study the influence of age on the ability of subjects to adapt mastication to changes in the hardness of foods. The study was carried out on 67 volunteers aged from 25 to 75 yr (29 males, 38 females) who had complete healthy dentitions. Surface electromyograms of the left and right masseter and temporalis muscles were recorded simultaneously with jaw movements using an electromagnetic transducer. Each volunteer was asked to chew and swallow four visco-elastic model foods of different hardness, each presented three times in random order. The number of masticatory cycles, their frequency, and the sum of all electromyographic (EMG) activity in all four muscles were calculated for each masticatory sequence. Multiple linear regression analyses were used to assess the effects of hardness, age, and gender. Hardness was associated to an increase in the mean number of cycles and mean summed EMG activity per sequence. It also increased mean vertical amplitude. Mean vertical amplitude and mean summed EMG activity per sequence were higher in males. These adaptations were present at all ages. Age was associated with an increase of 0.3 cycles per sequence per year of life and with a progressive increase in mean summed EMG activity per sequence. Cycle and opening duration early in the sequence also fell with age. We concluded that although the number of cycles needed to chew a standard piece of food increases progressively with age, the capacity to adapt to changes in the hardness of food is maintained. PMID- 15277596 TI - Precision of spike trains in primate retinal ganglion cells. AB - Recent studies have revealed striking precision in the spike trains of retinal ganglion cells in several species and suggested that this precision could be an important aspect of visual signaling. However, the precision of spike trains has not yet been described in primate retina. The spike time and count variability of parasol (magnocellular-projecting) retinal ganglion cells was examined in isolated macaque monkey retinas stimulated with repeated presentations of high contrast, spatially uniform intensity modulation. At the onset of clearly delineated periods of firing, retinal ganglion cells fired spikes time-locked to the stimulus with a variability across trials as low as 1 ms. Spike count variance across trials was much lower than the mean and sometimes approached the minimum variance possible with discrete counts, inconsistent with Poisson statistics expected from independently generated spikes. Spike time and count variability decreased systematically with stimulus strength. These findings were consistent with a model in which firing probability was determined by a stimulus driven free firing rate modulated by a recovery function representing the action potential absolute and relative refractory period. PMID- 15277597 TI - Context contingent signal processing in the cerebellar flocculus and ventral paraflocculus during gaze saccades. AB - The vestibuloocular reflex (VOR) functions to stabilize gaze when the head moves. The flocculus region (FLR) of the cerebellar cortex, which includes the flocculus and ventral paraflocculus, plays an essential role in modifying signal processing in VOR pathways so that images of interest remain stable on the retina. In squirrel monkeys, the firing rate of most FLR Pk cells is modulated during VOR eye movements evoked by passive movement of the head. In this study, the responses of 48 FLR Purkinje cells, the firing rates of which were strongly modulated during VOR evoked by passive whole body rotation or passive head-on trunk rotation, were compared to the responses generated during compensatory VOR eye movements evoked by the active head movements of eye-head saccades. Most (42/48) of the Purkinje cells were insensitive to eye-head saccade-related VOR eye movements. A few (6/48) generated bursts of spikes during saccade-related VOR but only during on-direction eye movements. Considered as a population FLR Pk cells were <5% as responsive to the saccade-related VOR as they were to the VOR evoked by passive head movements. The observations suggest that the FLR has little influence on signal processing in VOR pathways during eye-head saccade related VOR eye movements. We conclude that the image-stabilizing signals generated by the FLR are highly dependent on the behavioral context and are called on primarily when external forces unrelated to self-generated eye and head movements are the cause of image instability. PMID- 15277598 TI - Activation of a calcium-activated cation current during epileptiform discharges and its possible role in sustaining seizure-like events in neocortical slices. AB - Epileptic seizures are composed of recurrent bursts of intense firing separated by periods of electrical quiescence. The mechanisms responsible for sustaining seizures and generating recurrent bursts are yet unclear. Using whole cell voltage recordings combined with intracellular calcium fluorescence imaging from bicuculline (BCC)-treated neocortical brain slices, I showed isolated paroxysmal depolarization shift (PDS) discharges were followed by a sustained afterdepolarization waveform (SADW) with an average peak amplitude of 3.3 +/- 0.9 mV and average half-width of 6.2 +/- 0.6 s. The SADW was mediated by the calcium activated nonspecific cation current (I(can)) as it had a reversal potential of 33.1 +/- 6.8 mV, was unaffected by changing the intracellular chloride concentrations, was markedly diminished by buffering [Ca(2+)](i) with intracellular bis-(o-aminophenoxy)-N,N,N',N'-tetraacetic acid (BAPTA), and was reversibly abolished by the I(can) blocker flufenamic acid (FFA). The Ca(2+) influx responsible for activation of I(can) was mediated by both N-methyl-d aspartate-receptor channels, voltage-gated calcium channels and, to a lesser extent, internal calcium stores. In addition to isolated PDS discharges, BCC treated brain slices also produced seizure-like events, which were accompanied by a prolonged depolarizing waveform underlying individual ictal bursts. The similarities between the initial part of this waveform and the SADW and the fact it was markedly reduced by buffering [Ca(2+)](i) with BAPTA strongly suggested it was mediated, at least in part, by I(can). Addition of FFA reversibly eliminated recurrent bursting, and transformed seizure-like events into isolated PDS responses. These results indicated I(can) was activated during epileptiform discharges and probably participated in sustaining seizure-like events. PMID- 15277599 TI - Generalized integrate-and-fire models of neuronal activity approximate spike trains of a detailed model to a high degree of accuracy. AB - We demonstrate that single-variable integrate-and-fire models can quantitatively capture the dynamics of a physiologically detailed model for fast-spiking cortical neurons. Through a systematic set of approximations, we reduce the conductance-based model to 2 variants of integrate-and-fire models. In the first variant (nonlinear integrate-and-fire model), parameters depend on the instantaneous membrane potential, whereas in the second variant, they depend on the time elapsed since the last spike [Spike Response Model (SRM)]. The direct reduction links features of the simple models to biophysical features of the full conductance-based model. To quantitatively test the predictive power of the SRM and of the nonlinear integrate-and-fire model, we compare spike trains in the simple models to those in the full conductance-based model when the models are subjected to identical randomly fluctuating input. For random current input, the simple models reproduce 70-80 percent of the spikes in the full model (with temporal precision of +/-2 ms) over a wide range of firing frequencies. For random conductance injection, up to 73 percent of spikes are coincident. We also present a technique for numerically optimizing parameters in the SRM and the nonlinear integrate-and-fire model based on spike trains in the full conductance based model. This technique can be used to tune simple models to reproduce spike trains of real neurons. PMID- 15277600 TI - Presynaptic activity and Ca2+ entry are required for the maintenance of NMDA receptor-independent LTP at visual cortical excitatory synapses. AB - We have shown that some neural activity is required for the maintenance of long term potentiation (LTP) at visual cortical inhibitory synapses. We tested whether this was also the case in N-methyl-d-aspartate (NMDA) receptor-independent LTP of excitatory connections in layer 2/3 cells of developing rat visual cortex. This LTP occurred after 2-Hz stimulation was applied for 15 min and always persisted for several hours while test stimulation was continued at 0.1 Hz. When test stimulation was stopped for 1 h after LTP induction, only one-third of the LTP instances disappeared, but most did disappear under a pharmacological suppression of spontaneous firing, indicating that LTP maintenance requires either evoked or spontaneous activities. LTP was totally abolished by a temporary blockade of action potentials with lidocaine or the removal of extracellular Ca(2+) after LTP induction, but it persisted under a voltage clamp of postsynaptic cells or after a temporary blockade of postsynaptic activity with the glutamate receptor antagonist kynurenate, suggesting that LTP maintenance requires presynaptic, but not postsynaptic, firing and Ca(2+) entry. More than one-half of the LTP instances were abolished after a pharmacological blockade of P-type Ca(2+) channels, whereas it persisted after either L-type or Ni(2+)-sensitive Ca(2+) channel blockades. These results show that the maintenance of NMDA receptor independent excitatory LTP requires presynaptic firing and Ca(2+) channel activation as inhibitory LTP, although the necessary level of firing and Ca(2+) entry seems lower for the former than the latter and the Ca(2+) channel types involved are only partly the same. PMID- 15277601 TI - Decoding continuous and discrete motor behaviors using motor and premotor cortical ensembles. AB - Decoding motor behavior from neuronal signals has important implications for the development of a brain-machine interface (BMI) but also provides insights into the nature of different movement representations within cortical ensembles. Motor control can be hierarchically characterized as the selection and planning of discrete movement classes and/or postures followed by the execution of continuous limb trajectories. Based on simultaneous recordings in primary motor (MI) and dorsal premotor (PMd) cortices in behaving monkeys, we demonstrate that an MI ensemble can reconstruct hand or joint trajectory more accurately than an equally sized PMd ensemble. In contrast, PMd can more precisely predict the future occurrence of one of several discrete targets to be reached. This double dissociation suggests that a general-purpose BMI could take advantage of multiple cortical areas to control a wider variety of motor actions. These results also support the hierarchical view that MI ensembles are involved in lower-level movement execution, whereas PMd populations represent the early intention to move to visually presented targets. PMID- 15277602 TI - Perceptual illusion of "paradoxical heat" engages the insular cortex. AB - Paradoxical heat (PH), the illusion of skin heat, accompanies many neurological disorders. Using the technique of percept-related functional MRI, we found a region of the right insular cortex specifically activated when subjects perceive a heat sensation in their right hand even though their skin temperature is cool or at neutral. This region was suppressed during mild skin cooling. We propose that this differential response is a manifestation of the role of the insula in signaling temperature perceptions regardless of the actual temperature of the skin. These findings suggest that a region within the insula has a complex role in heat perception, perhaps contributing to a specific, rather than general, thermosensory perception. These data provide insight to our basic understanding of normal and pathological thermosensory perceptions. PMID- 15277603 TI - Statement of retraction. Estradiol rapidly activates Akt via the ErbB2 signaling pathway. PMID- 15277606 TI - Donepezil therapy for neuropsychiatric symptoms in AD: methods make the message. PMID- 15277607 TI - Menstrual migraine: timing is everything. PMID- 15277608 TI - Gliomatosis cerebri: better definition, better treatment. PMID- 15277609 TI - West Nile virus and "poliomyelitis". AB - West Nile virus (WNV) has recently been associated with a syndrome of acute flaccid paralysis. Most cases of WNV-associated weakness have clinical, histopathologic, and electrophysiologic characteristics indistinguishable from those of poliomyelitis caused by infection with poliovirus. There is debate about the nomenclature of this manifestation of WNV infection. An historical perspective of the term "poliomyelitis" suggests that the term "WNV poliomyelitis" seems appropriate, but members of the neurologic and infectious disease communities should engage in discussion regarding the terminology of this syndrome. PMID- 15277610 TI - Modulation of the renin-angiotensin-aldosterone system for the secondary prevention of stroke. AB - Recurrent stroke is a major public health concern and new treatment strategies are needed. While modulation of the renin angiotensin aldosterone system (RAAS) has proven effective in reducing recurrent cardiac events, its role in preventing recurrent cerebrovascular events remains unclear. RAAS is both a circulating and tissue based hormonal system that regulates homeostasis and tissue responses to injury in both the CNS and the periphery, via the activity of angiotensin II (Ang II). Vascular and hematologic effects induced by Ang II including endothelial dysfunction, vascular structural changes, inflammation, hemostasis, and fibrinolysis are increasingly linked to the occurrence of cerebrovascular events. Animal models have shown that RAAS modulation may be protective in cerebrovascular disease. The HOPE and LIFE trials support the role of blood pressure independent mechanisms of RAAS modulation for improving outcomes in a broad range of patients with cardiovascular disease but do not specifically address recurrent stroke prevention. PROGRESS, a trial of secondary stroke prevention, demonstrates that blood pressure reduction with a combination strategy including the routine use of ACE inhibitors prevents recurrent stroke. Current evidence suggests that the RAAS plays an important role in the development and progression of cerebrovascular disease. Modulation of the RAAS holds promise for the secondary prevention of stroke, however, ongoing clinical trials will better define the exact role of ACE inhibitor and angiotensin II Type 1 receptor blocker therapy in stroke survivors. PMID- 15277611 TI - The efficacy of donepezil in the treatment of neuropsychiatric symptoms in Alzheimer disease. AB - OBJECTIVE: To determine the efficacy of donepezil in the treatment of neuropsychiatric symptoms in patients with Alzheimer disease (AD) in a randomized withdrawal study. METHOD: Patients with mild to moderate AD with marked neuropsychiatric symptoms at baseline (Neuropsychiatric Inventory [NPI] > 11 points) were treated openly with donepezil 5 mg daily for 6 weeks followed by 10 mg daily for a further 6 weeks. Patients were then randomized (60:40) to either placebo or 10 mg donepezil daily. All patients were assessed at 6 weeks and provided there was no marked cognitive deterioration their blinded treatment was continued for a further 6 weeks. NPI and carer distress were assessed at 6 weekly intervals throughout the study. RESULTS: A total of 134 patients participated. Following randomization patients who continued on donepezil 10 mg for 12 weeks had improvements in NPI compared with the placebo group (mean change -2.9 vs 3.3 points; ITT-LOCF p = 0.02) and in NPI-Distress scores (median change -2.0 vs 1.0 points; ITT-LOCF p = 0.01). During the open-label phase the total NPI and NPI Distress scores were lower after 12 weeks treatment with open label donepezil compared with baseline (total NPI 22 points vs13 points; ITT-LOCF p < 0.0001; NPI Distress 13.5 vs 7.9 points; ITT-LOCF p < 0.0001). In the open-label phase all domains of the NPI (with the exception of elation) were improved (all p < 0.05 after Bonferroni correction). CONCLUSIONS: Donepezil has significant efficacy in the treatment of neuropsychiatric symptoms in patients with mild to moderate AD. PMID- 15277613 TI - Fasting insulin and incident dementia in an elderly population of Japanese American men. AB - OBJECTIVE: To evaluate the association of fasting insulin level to incident dementia in a cohort of elderly men. METHODS: Data are from the Honolulu-Asia Aging Study, a community-based study of Japanese-American men, aged 71 to 91 years in 1991. Serum insulin was measured in 1991 and participants were grouped based on their insulin levels. Dementia was ascertained in 1991, 1994, and 1996 according to international guidelines. The 2,568 men dementia-free in 1991 were reexamined in 1994 and 1996; 244 new cases of dementia were diagnosed. Survival analysis with age as the time scale was used to estimate the risk (hazard ratio [HR] and 95% CI) for incident dementia associated with levels of insulin. RESULTS: The risk of dementia was increased at the two extremes of the insulin distribution (lower and upper 15th percentiles). Compared to the rest of the cohort subjects in the lowest 15th percentile and highest 15th percentile had an increased risk for dementia (HR = 1.54, CI 1.11 to 2.11 and HR = 1.54, CI 1.05 to 2.26). In men with insulin levels <22.2 mIU/L the risk for dementia decreased with increased levels of insulin (HR = 0.76, CI 0.72 to 0.79 for each increase of one logarithmic unit -2.72 mIU/L of insulin). In men with insulin levels >/=22.2 mIU/L the risk for dementia increased with increasing levels of insulin (HR = 1.64, CI = 1.07 to 2.52 for each 2.72 mIU/L). CONCLUSIONS: Both low and high levels of insulin are associated with an increased risk of developing dementia. PMID- 15277612 TI - Memory impairment, but not cerebrovascular disease, predicts progression of MCI to dementia. AB - BACKGROUND: Mild cognitive impairment (MCI) is widely viewed as the transition phase between normal aging and Alzheimer disease (AD). Given that MCI can also result from cerebrovascular disease (CVD), the authors used clinical, MRI, and cognitive measures of AD and CVD to test the hypothesis that CVD increases the likelihood of progression from MCI to dementia within 3 years. OBJECTIVE: To examine the impact of CVD on progression of MCI to dementia. METHODS: Fifty-two consecutive patients with MCI (71% men) including many with symptomatic CVD were longitudinally evaluated for 3.1 +/- 1.3 years. MCI was defined as a Clinical Dementia Rating Scale (CDR) score of 0.5. Dementia was defined as progression to a CDR score of > or =1.0. RESULTS: Forty-four percent of the MCI patients had MRI infarcts, 50% of which were symptomatic. Thirty-three percent of patients progressed to dementia, and 37.8% of these had MRI infarcts. Clinically probable or possible AD was diagnosed in approximately 82% of converters. Of the clinical and MRI measures, only hippocampal volume was associated with increased risk to progression (hazard ratio [HR] = 0.31 [95% CI 0.1 to 0.92], p = 0.03). When neuropsychological measures were included in the analysis, memory (HR = 0.90 [95% CI 0.84 to 0.96], p = 0.002) and executive function (HR = 0.96 [95% CI 0.92 to 1.0], p = 0.045) were associated with increased risk of dementia progression, whereas APOE genotype, cerebrovascular risk factors, clinical stroke, presence or absence of lacunes, and extent of white matter hyperintensities did not predict progression. CONCLUSION: Within a heterogenous group of MCI patients, including many with clinically significant CVD, baseline memory and executive performance significantly predicted likelihood to develop dementia. PMID- 15277614 TI - Hereditary dementia with intracerebral hemorrhages and cerebral amyloid angiopathy. AB - BACKGROUND: Deposition of the beta-amyloid peptide (Abeta) in neuritic plaques is a hallmark of Alzheimer disease (AD). Mutations in genes encoding amyloid precursor protein (APP) and presenilin 1 and 2 (PSEN1, PSEN2) are associated with increased accumulation of Abeta in neuritic plaques or in the walls of cerebral vessels. Intracerebral hemorrhage occasionally affects patients with AD. METHODS: A Finnish family with dementia in four generations and with frequent co occurrence of dementia and intracerebral hemorrhage was identified. Clinical features of 14 family members with a cognitive decline were evaluated. All exons in genes encoding APP, PSEN1, PSEN2, cystatin C, transthyretin, gelsolin, and ITM2B were sequenced, and an association study of APP was conducted by identification of single-nucleotide polymorphisms. RESULTS: Neuropathologic examination revealed Alzheimer-type changes with Abeta in neuritic plaques and vessel walls, but the cognitive profile of the patients differed from that in AD, as the visuoconstructive functions and verbal fluency were well preserved even in the moderate stage of the disease. In addition to cognitive decline, five patients had had lobar intracerebral hemorrhages and one was diagnosed with hemosiderin deposits in MRI, suggesting previous cerebral microbleeds. No causative mutations were identified in candidate genes associated with amyloid diseases, but linkage to APP region could not be entirely excluded. CONCLUSIONS: The family presents an autosomal dominant form of beta-amyloidogenic disease that resembles the Italian, Flemish, and Iowa types of AD. No amyloidogenic mutations were identified, but the role of the APP region could not be entirely excluded. PMID- 15277615 TI - Insulin-degrading enzyme and Alzheimer disease: a genetic association study in the Han Chinese. AB - BACKGROUND: The gene for insulin-degrading enzyme (IDE) represents a strong positional and biologic candidate for late-onset Alzheimer disease (LOAD) susceptibility. IDE is located on chromosome 10q23.3 close to a region of linkage for LOAD. In addition, many studies have identified a possible role of IDE in the degradation of amyloid beta-protein and the intracellular amyloid precursor protein (APP) domain released by gamma-secretase processing. OBJECTIVE: To examine the association of IDE with AD in the Han Chinese. METHODS: Four IDE polymorphisms (three in 5'-untranslated region and one in intron 21) were analyzed, using a population of 210 patients with LOAD and 200 control subjects well matched for age, sex, and ethnic background. RESULTS: Among the four polymorphisms studied, only the C allele of single-nucleotide polymorphism (SNP) IDE2 showed association with AD (p = 0.005). Stratification of the data by APOE epsilon4 status indicated that the association between IDE2 and AD was confined to APOE epsilon4 carriers only. No association was found between all variants studied and AD within APOE epsilon4-negative subjects. The global haplotype frequencies showed significant differences between AD patients and control subjects. Furthermore, overrepresentation of GCTG haplotype in the AD group was found. It may be a risk haplotype for AD. CONCLUSIONS: These results suggest a possible synergic interaction between IDE and APOE epsilon4 in the risk to develop late-onset sporadic AD. IDE might modify the effect of the APOE epsilon4 risk factor in the Han Chinese population. PMID- 15277616 TI - White matter lesions impair frontal lobe function regardless of their location. AB - OBJECTIVE: To analyze the effect of white matter lesions in different brain regions on regional cortical glucose metabolism, regional cortical atrophy, and cognitive function in a sample with a broad range of cerebrovascular disease and cognitive function. METHODS: Subjects (n = 78) were recruited for a study of subcortical ischemic vascular disease (SIVD) and Alzheimer disease (AD) contributions to dementia. A new method was developed to define volumes of interest from high-resolution three-dimensional T1-weighted MR images. Volumetric measures of MRI segmented white matter signal hyperintensities (WMH) in five different brain regions were related to regional PET glucose metabolism (rCMRglc) in cerebral cortex, MRI measures of regional cortical atrophy, and neuropsychological assessment of executive and memory function. RESULTS: WMH was significantly higher in the prefrontal region compared to the other brain regions. In all subjects, higher frontal and parietal WMH were associated with reduced frontal rCMRglc, whereas occipitotemporal WMH was only marginally associated with frontal rCMRglc. These associations were stronger and more widely distributed in nondemented subjects where reduced frontal rCMRglc was correlated with WMH for all regions measured. In contrast, there was no relationship between WMH in any brain region and rCMRglc in either parietal or occipitotemporal regions. WMHs in all brain regions were associated with low executive scores in nondemented subjects. CONCLUSIONS: The frontal lobes are most severely affected by SIVD. WMHs are more abundant in the frontal region. Regardless of where in the brain these WMHs are located, they are associated with frontal hypometabolism and executive dysfunction. PMID- 15277618 TI - A randomized trial of frovatriptan for the intermittent prevention of menstrual migraine. AB - BACKGROUND: Menstrually associated migraine (MAM) is often prolonged and difficult to manage with conventional therapies. Frovatriptan is a new selective 5HT(1B/1D) receptor agonist indicated for short-term management of migraine. It has a long half-life and good tolerability. These characteristics suggest that frovatriptan may be useful for the intermittent prevention of MAM. METHODS: The study was a randomized, double-blind, placebo-controlled, three-way crossover design. Patients treated each of three perimenstrual periods (PMPs) with placebo, frovatriptan 2.5 mg QD, and frovatriptan 2.5 mg BID. The 6-day treatment started 2 days before the anticipated start of MAM headache. The primary efficacy endpoint was incidence of MAM headache during the 6-day PMP. RESULTS: The population comprised 546 women (mean age, 37.6 years). Use of frovatriptan reduced the occurrence of MAM headache. The incidence of MAM headache during the 6-day PMP was 67% for placebo, 52% for frovatriptan 2.5 mg QD, and 41% for frovatriptan 2.5 mg BID. Both frovatriptan regimens were superior to placebo (p < 0.0001), and the BID regimen was superior to the QD regimen (p < 0.001). Both frovatriptan regimens also reduced MAM severity (p < 0.0001), duration (p < 0.0001), and the use of rescue medication (p < 0.01 QD; p < 0.0001 BID) in a dose dependent manner. The incidence and type of adverse events for both regimens were similar to placebo and consistent with those reported for short-term migraine management. CONCLUSION: Frovatriptan given prophylactically for 6 days was effective in reducing the incidence of menstrually associated migraine. More than half of patients who used frovatriptan 2.5 mg BID had no menstrually associated migraine headache during the 6-day perimenstrual period. The findings are consistent with the long duration of action and good tolerability of frovatriptan observed in short-term migraine management. PMID- 15277617 TI - Total homocysteine and cognition in a tri-ethnic cohort: the Northern Manhattan Study. AB - OBJECTIVE: Several studies implicate elevated homocysteine as a risk factor for dementia and cognitive decline, but most studies have involved subjects older than 55 years from homogeneous populations. The authors examined homocysteine and cognition in a tri-ethnic community sample 40 years and older. METHOD: The Northern Manhattan Study includes 3,298 stroke-free subjects. Of these 2,871 had baseline fasting total homocysteine (tHcy) levels and Mini-Mental State Examination (MMSE) scores available. The authors used multiple linear regression to examine the cross-sectional association between baseline tHcy levels and mean MMSE scores adjusting for sociodemographic and vascular risk factors. RESULTS: Homocysteine levels were related to age, renal function, and B12 deficiency. Those with B12 deficiency had tHcy levels five points higher (9.4 vs 14.4 nmol/L). Mean MMSE scores differed by age, sex, and race-ethnic group. Those with hypertension, diabetes, cardiac disease, and B12 deficiency had lower MMSE scores. In multivariate analyses, elevated tHcy was associated with lower mean MMSE scores for those older than 65 but not for those 40 to 64. Adjusting for B12 deficiency and sociodemographic factors the mean MMSE was 2.2 points lower for each unit increase in the log tHcy level (95% CI -3.6, -0.9). Adding vascular risk factors to the model did not attenuate this effect (mean MMSE -2.2 points; 95% CI -3.5, -0.9). CONCLUSIONS: Elevated homocysteine was independently associated with decreased cognition in subjects older than 65 in this tri-ethnic cohort, adjusting for sociodemographic and vascular risk factors. PMID- 15277619 TI - Initial chemotherapy in gliomatosis cerebri. AB - BACKGROUND: Because of the diffuse nature of gliomatosis cerebri (GC), surgery is not suitable, and large field radiotherapy carries the risk of severe toxicity. In this setting, initial chemotherapy warrants further investigation. METHODS: The authors treated 63 consecutive patients with GC with initial chemotherapy consisting of either PCV (procarbazine, 60 mg/m2 on days 8 to 21; CCNU, 110 mg/m2 on day 1; and vincristine, 1.4 mg/m2 on days 8 and 29) or temozolomide (TMZ; 150 to 200 mg/m2 for 5 days every 4 weeks). There were 40 men and 23 women, with a median age of 48 years (range, 17 to 74 years) and a median Karnofsky performance status of 90 (range, 50 to 100). GC was initially present at diagnosis in 49 patients (primary GC), whereas 14 patients with a circumscribed glioma at onset developed secondary GC after a median follow-up period of 5.11 years. GC was classified based on the predominant tumor cells as astrocytic, oligodendroglial, or mixed GC. RESULTS: Seventeen patients received 1 to 6 cycles (median, 5) of PCV, and 46 received 2 to 24 courses (median, 13) of TMZ. Grade 3 to 4 hematologic toxicity was seen in 4 of 17 (23.5%) patients treated with PCV and in 4 of 46 (8.6%) of those treated with TMZ. Clinical objective responses were observed in 21 of 63 (33%) patients, and radiologic responses were seen in 16 of 62 (26%), with no significant difference between the two regimens. For all patients combined, the median progression-free survival (PFS) and overall survival (OS) were 16 months and 29 months, respectively. Regardless of the chemotherapeutic regimen, oligodendroglial GC had a better prognosis than astrocytic and oligoastrocytic GC in terms of PFS (p < 0.02) and OS (p < 0.0001). CONCLUSION: Initial chemotherapy is useful for some patients with gliomatosis cerebri. Temozolomide is well tolerated and appears to be a valuable alternative to procarbazine-CCNU-vincristine, especially for those with slow-growing, low grade GC. PMID- 15277620 TI - Season of birth and risk of brain tumors in adults. AB - BACKGROUND: Recent studies demonstrated an excess of winter births in children with brain tumors and in adults with various neurologic or psychiatric diseases relative to the general population. OBJECTIVE: To investigate a possible association between month of birth and risk of brain tumors in adults using data from a large, hospital-based case-control study. METHODS: Cases were patients with incident glioma (n = 489) or meningioma (n = 197) diagnosed at hospitals in Boston, MA, Phoenix, AZ, and Pittsburgh, PA. Controls (n = 799) were patients hospitalized for a variety of nonmalignant conditions and frequency matched to cases by hospital, age, sex, race/ethnicity, and distance of residence from hospital. Odds ratios (ORs) were calculated using multivariate unconditional logistic regression allowing for cyclic variation in risk with month of birth. RESULTS: A relationship between month of birth and risk of adult glioma and meningioma was found, best described by a 12-month periodic function with peaks in February and January and troughs in August and July. The association between month of birth and risk of glioma differed significantly by handedness, with left handed and ambidextrous subjects born during late fall through early spring being at particularly high risk of adult glioma as compared with those born at other times of the year. CONCLUSION: These findings suggest the importance of seasonally varying exposures during the pre- or postnatal period in the development of brain tumors in adults. PMID- 15277621 TI - Antibody-positive paraneoplastic neurologic syndromes: value of CT and PET for tumor diagnosis. AB - OBJECTIVE: To assess use of whole-body 18F fluoro-2-deoxy-glucose (FDG)-PET and CT for diagnosing tumor in patients with antibody-positive paraneoplastic neurologic syndromes (PNS). METHODS: In order to directly compare CT and FDG-PET imaging in patients with various antineuronal antibodies, the authors performed in parallel CT scanning and FDG-PET in a series of 13 consecutive patients (9 women, 4 men, aged 59 +/- 14 years) with positive antineuronal antibodies (anti Hu: 8, anti-Yo: 4, anti-Tr: 1) in whom the authors were searching for a tumor or tumor recurrence. RESULTS: A new tumor or tumor recurrence was found in 10/13 patients (5 small cell lung cancer, 2 ovarian cancer, and 1 each neuroblastoma, Hodgkin's lymphoma, and lymph node metastasis of adenocarcinoma). All tumors except one with good clinical evidence for small cell lung cancer were confirmed histologically. For detection of tumor or tumor recurrence, CT was positive in 3/10 patients (sensitivity of 30%), and FDG-PET in 9/10 (sensitivity of 90%, difference between methods p < 0.01), but the combination of both methods showed a sensitivity of 100%. CONCLUSIONS: FDG-PET imaging is useful in tumor screening of patients with antineuronal antibodies, but should be complemented by CT scanning to increase sensitivity and accuracy of tumor diagnosis. PMID- 15277622 TI - Functional reorganization of spatial transformations after a parietal lesion. AB - BACKGROUND: Mental spatial transformations are ubiquitous and necessary for everyday spatial cognition, such as packing luggage into a car or repairing a broken vase. The posterior parietal cortex is known to be involved in performing such transformations. OBJECTIVE: To measure reorganization after lesioning of posterior parietal cortex areas subserving spatial transformation. METHOD: Brain activity in a patient who underwent a resection of right parietal cortex to manage intractable epilepsy was measured using fMRI while he performed a set of spatial transformation tasks. These data were compared with data from a group of healthy control subjects. RESULTS: During spatial transformations, activity in the regions overlapping the resection was reduced in the patient compared with control subjects, but activity in the contralateral cortex was greater than that of control subjects. CONCLUSIONS: After a lesion the left hemisphere can adopt components of spatial reasoning normally subserved by the right hemisphere. This converges with evidence that components of language processing normally subserved by the left hemisphere can be taken over by the right hemisphere, suggesting that plasticity of function in the adult human cortex is a general characteristic. PMID- 15277623 TI - Psychiatric comorbidities in patients with Parkinson disease and psychosis. AB - OBJECTIVE: To determine the prevalence and impact of comorbid psychiatric disturbances in Parkinson disease (PD) patients with psychosis. METHODS: Subject data were derived from a research database of 116 PD patients participating in standardized motor, cognitive, psychiatric, and caregiver assessments. RESULTS: There were 25 patients (22%) with psychosis manifest as hallucinations (n = 9), delusions (n = 1), or hallucinations and delusions (n = 15) and 25 patients (22%) who had no current or past psychiatric comorbidities (PDN). In the psychotic group, 44% had psychosis only (PSY), and 56% had psychosis plus at least one other comorbid psychiatric disturbance (PSY+), including depressive disorders (71%), anxiety disorders (21%), apathetic syndromes (14%), and delirium (14%). There were no differences in age, sex, education, or age onset or duration of PD among the PSY, PSY+, and PDN groups. Both psychotic groups had greater motor, functional, and frontal cognitive deficits and increased caregiver burden scores relative to PDN. PSY+ showed greater global and selective cognitive deficits compared to PDN. Psychosis was a primary predictor of caregiver burden, whereas depressive symptoms indirectly enhanced motor impairments. CONCLUSIONS: Nonpsychotic psychiatric disturbances, especially affective disturbances, are common comorbidities in PD patients with psychosis and warrant clinical attention to reduce morbidity and caregiver distress. PMID- 15277624 TI - Pergolide use in Parkinson disease is associated with cardiac valve regurgitation. AB - OBJECTIVE: To determine if pergolide injures heart valves, by comparing echocardiographic findings in pergolide-treated patients with those of a historical control group. METHODS: Letters were sent to all patients in the authors' practice believed to be taking pergolide, and those responders who wished to continue it were urged to undergo echocardiography. Echocardiograms were obtained on 46 patients, and scores for valvular regurgitation were compared with those from an age-matched control group derived from the Framingham Study. The composite valve regurgitation score was modeled as a linear function of total milligrams lifetime use of pergolide, controlling for age. RESULTS: Eighty-nine percent of pergolide-treated patients had some degree of valvular insufficiency. For each of the three valves for which there are control data, we found an approximately 2- to 3-fold increased risk of abnormal valves in the pergolide patients (odds ratio [OR] approximately 3) and an estimated 14-fold increased risk of concerning tricuspid regurgitation (OR = 18.4). The composite valve score (the sum of valve scores for each of the four valves) was a function of lifetime pergolide use. CONCLUSION: Pergolide may injure cardiac valves, resulting most commonly in tricuspid regurgitation. PMID- 15277625 TI - No evidence for heritability of Parkinson disease in Swedish twins. AB - BACKGROUND: Although several genes are implicated in Parkinson disease (PD), they explain only a small fraction of cases. The etiology of most cases is yet unknown. OBJECTIVE: To evaluate heritability of PD in same-sexed and opposite sexed twin pairs in the Swedish Twin Registry (STR). METHODS: All twins in the STR born in 1950 or earlier and alive in 1998 (n = 50,150) were included. The authors screened 33,780 twins in 14,082 pairs for PD by telephone interviews and linked the STR to the Swedish Inpatient Discharge Register. Two hundred forty seven twins with self-reported PD or a PD diagnosis in the Inpatient Discharge Register (called "possible PD") and 517 twins who reported parkinsonian symptoms or use of antiparkinsonian medication ("suspected parkinsonism or movement disorder") were identified. RESULTS: For possible PD, there were only two concordant pairs, both female dizygotic. Similarly, concordances were low in all zygosity groups when the definition of affected was expanded to include twins with suspected parkinsonism or movement disorder in addition to possible PD. Sex differences in the relative importance of genetic and environmental effects were indicated with a marginally larger familial component in women. The best-fitting structural equation model included only environmental components of variance. CONCLUSIONS: These results suggest that environmental factors are most important in the etiology of PD. Compared with other complex diseases, the importance of genetic effects in PD is notably low. The preponderance of discordant twin pairs provides an ideal material for studying environmental risk factors and potential genotype-by-environment interaction. PMID- 15277626 TI - Cooling for Acute Ischemic Brain Damage (COOL AID): a feasibility trial of endovascular cooling. AB - OBJECTIVE: To report results of a randomized pilot clinical feasibility trial of endovascular cooling in patients with ischemic stroke. METHODS: Forty patients with ischemic stroke presenting within 12 hours of symptom onset were enrolled in the study. An endovascular cooling device was inserted into the inferior vena cava of those randomized to hypothermia. A core body temperature of 33 degrees C was targeted for 24 hours. All patients underwent clinical assessment and MRI initially, at days 3 to 5 and days 30 to 37. RESULTS: Eighteen patients were randomized to hypothermia and 22 to receive standard medical management. Thirteen patients reached target temperature in a mean of 77 +/- 44 minutes. Most tolerated hypothermia well. Clinical outcomes were similar in both groups. Mean diffusion-weighted imaging (DWI) lesion growth in the hypothermia group (n = 12) was 90.0 +/- 83.5% compared with 108.4 +/- 142.4% in the control group (n = 11) (NS). Mean DWI lesion growth in patients who cooled well (n = 8) was 72.9 +/- 95.2% (NS). CONCLUSIONS: Induced moderate hypothermia is feasible using an endovascular cooling device in most patients with acute ischemic stroke. Further studies are needed to determine if hypothermia improves outcome. PMID- 15277627 TI - Use of antihypertensive agents in the management of patients with acute ischemic stroke. AB - BACKGROUND: To protect the ischemic penumbra, guidelines have recommended against treating all but the severest elevations in blood pressure during acute ischemic stroke. OBJECTIVE: To determine how often antihypertensive agents were used in routine clinical practice and whether this use was consistent with guideline recommendations. METHODS: The records of patients discharged with ischemic stroke in 2000 at Baystate Medical Center in Springfield, MA, were reviewed. Adherence was evaluated by examining the use of antihypertensive agents in the context of daily blood pressure recordings during the first 4 days of hospitalization. Therapy was considered appropriate in the setting of severe hypertension (systolic blood pressure of >220 mm Hg or mean arterial blood pressure of >130 mm Hg) and potentially harmful in the setting of relative (systolic blood pressure of <120 mm Hg or mean arterial blood pressure of <85 mm Hg) or absolute (systolic blood pressure of <90 mm Hg or mean arterial blood pressure of <60 mm Hg) hypotension. RESULTS: One hundred (65%) of the 154 ischemic stroke patients were treated with antihypertensive agents. Forty-two percent of those who had received therapy prior to admission had their regimen intensified, and 36% of previously untreated patients had therapy initiated. Sixteen (11%) patients had hypertension severe enough to warrant treatment upon arrival, and 34 (22%) had at least one episode of severe hypertension during the first 4 hospital days. Sixty-five (65%) patients developed relative hypotension on a day when antihypertensive agents were administered, and five (5%) developed absolute hypotension. CONCLUSIONS: Most patients with acute ischemic stroke are treated with antihypertensive agents despite the absence of severe hypertension. Although low blood pressure is common among treated patients, frank hypotension is unusual. PMID- 15277628 TI - EKG abnormalities in children and adolescents with symptomatic temporal lobe epilepsy. AB - BACKGROUND: Changes in cardiac rate and rhythm are often found in adult patients with temporal lobe epilepsy (TLE) and could be involved in the pathogenesis of sudden unexplained death (SUDEP). However, little is known about heart rate (HR) variability in pediatric patients with TLE. OBJECTIVES: To investigate ictal and peri-ictal HR abnormalities in children and adolescents with medically refractory symptomatic TLE and to determine the influence of focus localization and laterality. METHODS: Patients younger than 18 years, with drug-resistant unilateral symptomatic TLE and presenting with at least one habitual complex partial seizure (CPS) during presurgical noninvasive video-EEG monitoring, were enrolled. Synchronous single-channel EKG recordings were analyzed during the preictal, ictal, and postictal stages. RESULTS: Twenty patients fulfilled the inclusion criteria. Seventy-two temporal lobe seizures (TLSs) were analyzed. Ictal tachycardia was found in 71 TLSs (98%), whereas ictal bradycardia was not observed. During preictal stages, tachycardia occurred in 20 seizures and mild bradycardia in 3. In 44 seizures (62%), tachycardia was still present >60 seconds after EEG seizure termination. Cluster analysis revealed significant differences in HR evolution depending on location and side of seizure onset: Early and high HR increase was primarily associated with right mesial TLSs. CONCLUSIONS: Cardiovascular dysregulation is common during temporal lobe CPSs in children. These results confirm a right hemispheric lateralization of sympathetic cardiac control. PMID- 15277629 TI - Effect of localization of missense mutations in SCN1A on epilepsy phenotype severity. AB - BACKGROUND AND METHODS: Many missense mutations in the voltage-gated sodium channel subunit gene SCN1A were identified in patients with generalized epilepsy with febrile seizures plus (GEFS+) and severe myoclonic epilepsy of infancy (SMEI), although GEFS+ is distinct from SMEI in terms of clinical symptoms, severity, prognosis, and responses to antiepileptic drugs. The authors analyzed the localization of missense mutations in SCN1A identified in patients with GEFS+ and SMEI to clarify the phenotype-genotype relationships. RESULTS: Mutations in SMEI occurred more frequently in the "pore" regions of SCN1A than did those in GEFS+. These SMEI mutations in the "pore" regions were more strongly associated than mutations in other regions with the presence of ataxia and tendency to early onset of disease. The possibility of participation of ion selectivity dysfunction of the channel in the pathogenesis of SMEI was suggested by a mutation in the pore region (R946C) identified in a SMEI patient. CONCLUSIONS: There was a significant phenotype-genotype relationship in generalized epilepsy with febrile seizures plus and severe myoclonic epilepsy of infancy with SCN1A missense mutations. More severe sodium channel dysfunctions including abnormal ion selectivity that are caused by mutations in the pore regions may be involved in the pathogenesis of SMEI. PMID- 15277630 TI - Differences in cognitive impairment of relapsing remitting, secondary, and primary progressive MS. AB - OBJECTIVE: To investigate the cognitive skills of patients with relapsing remitting multiple sclerosis (RRMS), secondary progressive MS (SPMS), and primary progressive MS (PPMS) relative to healthy control subjects and to assess whether there is heterogeneity in the type of cognitive disabilities demonstrated by patients with different MS phenotypes. METHODS: RRMS patients (n = 108), SPMS patients (n = 71), PPMS patients (n = 55), and healthy control subjects (n = 67) underwent neuropsychological assessment with the Brief Repeatable Battery of Neuropsychological Tests. RESULTS: Relative to controls, cognitive performance of RRMS patients was deficient when tasks required higher-order working memory (WM) processes (Word List Generation, 10/36 Spatial Recall Test, Symbol Digit Modalities Test). PPMS and SPMS patients performed poorer than control subjects on all tasks. SPMS patients performed more poorly than PPMS patients when tasks required higher-order WM processes, except when speed of information processing played a relatively important role (Symbol Digit Modalities Test, Paced Auditory Serial Addition Test). Whereas RRMS patients generally performed better than the progressive subtypes, they showed relatively poor verbal fluency. CONCLUSION: MS patients with different disease courses have different cognitive profiles. PMID- 15277631 TI - Americo Negrette (1924 to 2003): diagnosing Huntington disease in Venezuela. AB - OBJECTIVE: To elucidate the role of Dr. Americo Negrette in diagnosing and reclassifying the dancing mania in Maracaibo, Venezuela as Huntington disease (HD). METHODS: All of the medical and nonmedical books and articles by Negrette were collected and reviewed. Personal interviews with Negrette were performed to confirm the details of his life and original work. His childhood, medical education, and contribution to HD, as well as contributions to medicine, art, and poetry, were covered in interview sessions. Excerpts from his autobiography, Ciudad de Fuego, were translated into English for publication. A complete bibliography of 70 books and research articles authored by Negrette was assembled. Negrette himself reviewed the compilation of this manuscript just days before his sudden death on September 14, 2003. RESULTS: Americo Negrette, a Venezuelan physician, biochemist, artist, and poet, observed a dancing epidemic in 1952. He acquired, through interviews with locals, the knowledge that there were two other small towns along Lake Maracaibo devastated by a syndrome called el mal (the bad). Negrette changed the diagnosis of these patients from a dancing mania to what he believed was Huntington chorea, later termed HD. He presented his findings at the Venezuelan Sixth Congress of Medical Science in 1955. He was met with reluctance in the local scientific community and a passive ear from government authorities. His description, written in Spanish, was not widely distributed beyond Venezuela, and its importance would have to wait until one of his students shared his observations (more than a decade later) with the HD research community and the rest of the world. CONCLUSION: The "dancing mania" of Maracaibo, because of the work of Americo Negrette, has been reclassified as Huntington disease. PMID- 15277632 TI - Willisian collateralization. PMID- 15277633 TI - Paroxysmal dysarthria and ataxia after midbrain infarction. AB - The authors describe a patient who showed paroxysmal dysarthria and right-limb ataxia after midbrain infarction. SPECT imaging showed marked hypoperfusion in the left parietal lobe while the patient was having frequent paroxysmal attacks. After treatment with phenytoin, the symptoms and hypoperfusion in SPECT imaging improved. The authors conclude that dysfunction of the cerebellothalamocortical pathway after midbrain infarction may cause paroxysmal dysarthria and ataxia. PMID- 15277634 TI - Elicited repetitive daily blindness: a new familial disorder related to migraine and epilepsy. AB - The authors report a family affected by multiple daily episodes of transient visual loss, elicited repetitive daily blindness (ERDB); the onset was early in life, and the disease followed a benign course. ERDB is associated with childhood epilepsy and familial hemiplegic migraine, apparently segregating as a monogenic, autosomal dominant condition with variable expression. Genetic linkage to CACNA1A was excluded. PMID- 15277635 TI - Prevalence of migraine on each day of the natural menstrual cycle. AB - Diary data from 155 women were analyzed using within-woman analysis. Compared with all other times of the cycle, migraine was 1.7 times more likely to occur during the 2 days before menstruation and 2.1 times more likely to be severe and 2.5 times more likely to occur during the first 3 days of menstruation and 3.4 times more likely to be severe. This confirms that migraine at menstruation is different from nonmenstrual attacks, even within individuals. PMID- 15277636 TI - Chemotherapy as initial treatment in gliomatosis cerebri: results with temozolomide. AB - The optimal therapy for gliomatosis cerebri is unclear, and the rate of response to chemotherapy is not known. Eleven radiotherapy-naive patients received a median number of 10 treatment cycles of temozolomide. An objective response was documented in 45%, and the median time to tumor progression was 13 months with a progression-free survival of 55% at 12 months. These results indicate that radiotherapy to extensive brain regions can be deferred until progressive disease is observed. PMID- 15277637 TI - Reanalysis of genotype distributions published in "Neurology" between 1999 and 2002. AB - The authors tested 123 genotypes described in 54 papers published in the journal Neurology between 1999 and 2002 to ascertain whether these genotype distributions deviated from Hardy-Weinberg equilibrium (HWE). Unreported deviations from HWE in 19 genotype distributions described in 11 of the papers were discovered. The authors also report additional information that could have been extracted after calculating HWE and conclude that HWE values should be mandatory in population genetic studies published in Neurology. PMID- 15277638 TI - DD genotype of ACE gene is a risk factor for intracerebral hemorrhage. AB - Genetic factors may play a role in susceptibility to stroke. The angiotensin converting enzyme (ACE) gene is a candidate gene for two phenotypically different types of stroke affecting small perforating arteries: spontaneous intracerebral hemorrhage (SIH) and ischemic stroke due to small vessel disease (SVD). The authors report evidence that ACE gene DD homozygosity of the I/D polymorphism in intron 16 is an independent risk factor for SIH, and not for SVD stroke, in a Polish population. PMID- 15277639 TI - Fragile X premutation alleles in SCA, ET, and parkinsonism in an Asian cohort. AB - Among 367 subjects, the authors analyzed 167 patients with essential tremor, sporadic progressive cerebellar ataxia, multiple-system atrophy, and atypical parkinsonism and 200 healthy control subjects for FMR1 premutation alleles. None of the subjects carried alleles within the premutation range. These findings suggest that in the absence of other supportive clinical or imaging features, the cost-effectiveness of routine fragile X tremor/ataxia syndrome screening in this Asian cohort with movement disorders was low. PMID- 15277640 TI - Prion protein codon 129 polymorphism and risk of Alzheimer disease. AB - The authors investigated the PRNP Met129Val polymorphism in 1,393 subjects including 482 patients with Alzheimer disease (AD) and two independent control groups. In patients, PRNP Met homozygosity conferred increasing risk with decreasing age at onset (onset: 61 to 70 years, n = 151, p = 0.02, odds ratio [OR] = 1.72, 95% CI = 1.2 to 2.53; onset: < or =60 years, n = 138, p = 0.013, OR = 1.92, 95% CI = 1.31 to 2.87), whereas no association was obtained in patients with onset at older than 70 years. The results suggest involvement of the prion protein in the pathogenesis of early-onset AD. PMID- 15277641 TI - Radiosurgery for cerebral arteriovenous malformations in hereditary hemorrhagic telangiectasia. AB - The authors evaluated the efficacy of radiosurgery (RS) for cerebral arteriovenous malformations in hereditary hemorrhagic telangiectasia (HHT AVMs). Two patients with seven HHT AVMs were treated by linear accelerator-RS. Complete obliteration was achieved 18 to 24 months post-treatment without side effects. Because HHT AVMs are small and multiple, RS is superior to microsurgery because it is noninvasive and all AVMs can be treated in one session regardless of their location. PMID- 15277642 TI - Ictal head version in generalized epilepsy. AB - Focal physical signs may occur with generalized seizures. The authors retrospectively reviewed video-EEG studies of 20 patients with apparent primary generalized epilepsy to assess the frequency of head version during generalized tonic-clonic seizures. Five patients (25%) demonstrated forced head version, including two with turning to the right and left on consecutive seizures. Forced head turning can occur in generalized epilepsy and may not necessarily indicate focal seizure origin. PMID- 15277643 TI - Lamotrigine-induced seizure aggravation and negative myoclonus in idiopathic rolandic epilepsy. AB - The authors describe a paradoxical reaction to lamotrigine (LTG) treatment in a patient with idiopathic rolandic epilepsy characterized by seizure deterioration, the appearance of new seizure type, and transient cognitive impairment. This phenomenon was present at a low dose after a slow titration and promptly reverted on LTG discontinuation. This rare event may have similarities with carbamazepine induced seizure worsening caused by the Na++ channel inhibitory effect of the two antiepileptic drugs. PMID- 15277644 TI - L-dopa responsiveness in dementia with Lewy bodies, Parkinson disease with and without dementia. AB - The authors analyzed whether nondemented (PD) and demented Parkinson patients (PDD) and patients with dementia with Lewy bodies (DLB) respond similarly in the levodopa test (LDT). Percentage of motor improvement was similar in the three groups; the proportion of patients with 10% and more improvement was greater in PD than in PDD and DLB. Positive LDT was predictive for favorable response in chronic levodopa treatment, but also some nonresponsive demented patients profited from chronic levodopa therapy. PMID- 15277645 TI - Neonatal lower motor neuron syndrome associated with maternal neuropathy with anti-GM1 IgG. AB - The authors report a newborn with motor neuropathy associated with anti-GM1 antibodies from an affected mother. This finding suggests that the disorder was due to transplacental transfer of pathogenic antibodies. PMID- 15277646 TI - Neuro-Behcet disease with predominant involvement of the brainstem. AB - We report a patient with relapsing-remitting meningoencephalitis secondary to neuro-Behcet disease which resulted in recurrent brainstem encephalitis. MRI revealed increased signal intensity on the proton density and T2-weighted images and gadolinium enhancement during relapses. Autopsy revealed acute on chronic meningoencephalomyelitis involving the entire CNS, which was most marked in the brainstem. PMID- 15277647 TI - Deterioration of the auditory brainstem response in children with type 3 Gaucher disease. AB - Enzyme replacement therapy (ERT) has improved the quality of life in patients with Gaucher disease (GD). An emerging concern is whether ERT can also halt the neurologic progression in type 3 GD. The authors examined the auditory brainstem response (ABR) in eight children with type 3 GD undergoing high-dose ERT and found a consistent deterioration in ABR response. They conclude that ERT does not halt brainstem degeneration and that alternative therapies must be sought. PMID- 15277648 TI - Schistosoma mansoni myelopathy: clinical and pathologic findings. AB - Thirteen patients with Schistosoma mansoni myelopathy are reported. Neurologic syndromes included acute areflexic flaccid paraplegia (three), thoracic myelopathy with hyperreflexia and Babinski sign (six), and a cauda equina syndrome (four). Inflammatory granulomas and a schistosome worm in a leptomeningeal vein of the spinal cord were observed in the one patient coming to necropsy. PMID- 15277649 TI - Reversible hyperintensity lesion on diffusion-weighted MRI in hypoglycemic coma. AB - In a woman aged 73 years who recovered from hypoglycemic coma without neurologic deficit after glucose infusion, admission diffusion-weighted MRI showed the presence of hyperintensive lesions. The lesions regressed after glucose infusion. PMID- 15277650 TI - Wernicke encephalopathy. PMID- 15277651 TI - Balint syndrome due to Creutzfeldt-Jakob disease. PMID- 15277652 TI - Basal ganglia hyperperfusion in a patient with systemic lupus erythematosus related parkinsonism. PMID- 15277653 TI - The epidemiology of the Lambert-Eaton myasthenic syndrome in the Netherlands. PMID- 15277654 TI - Improvement of SSPE by intrathecal infusion of alpha-IFN. PMID- 15277655 TI - Bilateral facial nerve palsy as first indication of relapsing hairy cell leukemia after 36 years. PMID- 15277656 TI - Pneumocephalus after air travel. PMID- 15277657 TI - Polymyositis: an overdiagnosed entity. PMID- 15277658 TI - Unicorns, dragons, polymyositis, and other mythical beasts. PMID- 15277659 TI - MRI of idiopathic lumbosacral plexopathy. PMID- 15277660 TI - Asymmetric freezing of gait in hemiparkinsonism-hemiatrophy. PMID- 15277661 TI - Nanometer-localized multiple single-molecule fluorescence microscopy. AB - Fitting the image of a single molecule to the point spread function of an optical system greatly improves the precision with which single molecules can be located. Centroid localization with nanometer precision has been achieved when a sufficient number of photons are collected. However, if multiple single molecules reside within a diffraction-limited spot, this localization approach does not work. This paper demonstrates nanometer-localized multiple single-molecule (NALMS) fluorescence microscopy by using both centroid localization and photobleaching of the single fluorophores. Short duplex DNA strands are used as nanoscale "rulers" to validate the NALMS microscopy approach. Nanometer accuracy is demonstrated for two to five single molecules within a diffraction-limited area. NALMS microscopy will greatly facilitate single-molecule study of biological systems because it covers the gap between fluorescence resonance energy transfer-based (<10 nm) and diffraction-limited microscopy (>100 nm) measurements of the distance between two fluorophores. Application of NALMS microscopy to DNA mapping with <10-nm (i.e., 30-base) resolution is demonstrated. PMID- 15277662 TI - Studies of stereocontrolled allylation reactions for the total synthesis of phorboxazole A. AB - A highly convergent total synthesis of the potent anticancer agent (+) phorboxazole A (1) is accomplished. Four components (3-6) are assembled with considerations for control of absolute and relative stereochemistry. Iterative asymmetric allylation methodology addresses key stereochemical features in the preparation of the 2,6-cis- and 2,6-trans-tetrahydropyranyl rings of the C3-C19 component (3). The stereocontrolled asymmetric allylation process is also used for development of the C28-C41 fragment (4). Novel Barbier coupling reactions of alpha-iodomethyl oxazoles and related thiazoles are described with samarium iodide. The convergent assembly of components 4 and 5 features formation of the fully substituted C22-C26 pyran by intramolecular capture of an allyl cation intermediate with high facial selectivity, and further efforts lead to E-C19/C20 olefination. The synthesis culminates with use of a modified Julia olefination for attachment of the C42-C46 segment and subsequent late-stage macrocyclization by installation of the (Z)-C2/C3 alpha,beta-unsaturated lactone. PMID- 15277663 TI - Bioreactor-based bone tissue engineering: the influence of dynamic flow on osteoblast phenotypic expression and matrix mineralization. AB - An important issue in tissue engineering concerns the possibility of limited tissue ingrowth in tissue-engineered constructs because of insufficient nutrient transport. We report a dynamic flow culture system using high-aspect-ratio vessel rotating bioreactors and 3D scaffolds for culturing rat calvarial osteoblast cells. 3D scaffolds were designed by mixing lighter-than-water (density, <1g/ml) and heavier-than-water (density, >1g/ml) microspheres of 85:15 poly(lactide-co glycolide). We quantified the rate of 3D flow through the scaffolds by using a particle-tracking system, and the results suggest that motion trajectories and, therefore, the flow velocity around and through scaffolds in rotating bioreactors can be manipulated by varying the ratio of heavier-than-water to lighter-than water microspheres. When rat primary calvarial cells were cultured on the scaffolds in bioreactors for 7 days, the 3D dynamic flow environment affected bone cell distribution and enhanced cell phenotypic expression and mineralized matrix synthesis within tissue-engineered constructs compared with static conditions. These studies provide a foundation for exploring the effects of dynamic flow on osteoblast function and provide important insight into the design and optimization of 3D scaffolds suitable in bioreactors for in vitro tissue engineering of bone. PMID- 15277664 TI - Cardioprotective effects of thioredoxin in myocardial ischemia and reperfusion: role of S-nitrosation [corrected]. AB - Apoptosis contributes to myocardial ischemia/reperfusion (MI/R) injury, and both thioredoxin (Trx) and nitric oxide have been shown to exert antiapoptotic effects in vitro. Recent evidence suggests that this particular action of Trx requires S nitrosation at Cys-69. The present study sought to investigate whether or not exogenously applied Trx reduces MI/R injury in vivo and to which extent this effect depends on S-nitrosation. Adult mice were subjected to 30 min of MI and treated with either vehicle or human Trx (hTrx, 2 mg/kg, i.p.) 10 min before reperfusion. Native hTrx was incorporated into myocardial tissue as shown by immunostaining, and reduced MI/R injury as evidenced by decreased terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling (TUNEL) staining, DNA fragmentation, caspase-3 activity, and infarct size. When hTrx was partially S nitrosated by preincubation with S-nitrosoglutathione, its cardioprotective effect was markedly enhanced. Treatment with hTrx significantly reduced p38 mitogen-activated protein kinase (MAPK) activity, and this effect was also potentiated by S-nitrosation. To further address the role of S-nitrosation for the overall antiapoptotic effect to Trx, the action of Escherichia coli Trx (eTrx) was investigated in the same model. Whereas eTrx inhibited MI/R-induced apoptosis to a degree similar to hTrx, S-nitrosation of this protein, which lacks Cys-69, failed to further enhance its antiapoptotic action. Collectively, our results demonstrate that systemically applied Trx is taken up by the myocardium to exert potent cardioprotective effects in vivo, offering interesting therapeutic avenues. In the case of hTrx, these effects are further potentiated by S-nitrosation, but this posttranslational modification is not essential for protection. PMID- 15277665 TI - Reproductive ground plan may mediate colony-level selection effects on individual foraging behavior in honey bees. AB - The colony-level phenotype of an insect society emerges from interactions between large numbers of individuals that may differ considerably in their morphology, physiology, and behavior. The proximate and ultimate mechanisms that allow this complex integrated system to form are not fully known, and understanding the evolution of social life strategies is a major topic in systems biology. In solitary insects, behavior, sensory tuning, and reproductive physiology are linked. These associations are controlled in part by pleiotropic networks that organize the sequential expression of phases in the reproductive cycle. Here we explore whether similar associations give rise to different behavioral phenotypes in a eusocial worker caste. We document that the pleiotropic genetic network that controls foraging behavior in functionally sterile honey bee workers (Apis mellifera) has a reproductive component. Associations between behavior, physiology, and sensory tuning in workers with different foraging strategies indicate that the underlying genetic architectures were designed to control a reproductive cycle. Genetic circuits that make up the regulatory "ground plan" of a reproductive strategy may provide powerful building blocks for social life. We suggest that exploitation of this ground plan plays a fundamental role in the evolution of social insect societies. PMID- 15277666 TI - Rpe65 Leu450Met variant is associated with reduced levels of the retinal pigment epithelium lipofuscin fluorophores A2E and iso-A2E. AB - There is a growing body of evidence that the nondegradable fluorophores that accumulate as the lipofuscin of retinal pigment epithelium (RPE) are involved in mechanisms leading to the degeneration of RPE in macular degeneration. Most of the constituents of RPE lipofuscin are inadvertent products of the retinoid visual cycle, the enzymatic pathway by which the 11-cis-retinal chromophore of rhodopsin is generated. Indeed, a major constituent of RPE lipofuscin, the pyridinium bisretinoid A2E, is a diretinal conjugate that forms in photoreceptor cells and is deposited in RPE cells as a consequence of the phagocytosis of the outer segment membrane by RPE cells. Given the adverse effects of A2E, there is considerable interest in combating its deposition so as to protect against vision loss. These efforts, however, necessitate an understanding of factors that modulate its formation. Here we show that an amino acid variant in murine Rpe65, a visual-cycle protein required for the regeneration of 11-cis-retinal, is associated with reduced A2E accumulation. PMID- 15277667 TI - Tissue remodeling of rat pulmonary arteries in recovery from hypoxic hypertension. AB - The reversibility of tissue remodeling is of general interest to medicine. Pulmonary arterial tissue remodeling during hypertension induced by hypoxic breathing is well known, but little has been said about the recovery of the arterial wall when the blood pressure is lowered again. We hypothesize that tissue recovery is a function of the oxygen concentration, blood pressure, location on the vascular tree, and time. We measured the changes of blood pressure, vessel lumen, vessel wall thicknesses, and opening angle of each segment of the blood vessel at its zero-stress state after step changes of the oxygen concentration in the breathing gas. The zero-stress state of each vessel is emphasized because it is important to the analysis of stress and strain and in morphometry. Experimental results are presented as histories of tissue parameters after step changes of the oxygen level. Tissue characteristics are examined under the hypothesis that they are linearly related to changes in the local blood pressure. Under this linearity hypothesis, each aspect of the tissue change can be expressed as a convolution integral of the blood pressure history with a kernel called the indicial response function. It is shown the indicial response function for rising blood pressure is different from that for falling blood pressure. This difference represents a major nonlinearity of the tissue remodeling process of the blood vessels. PMID- 15277668 TI - Worm genomes hold the smoking guns of intron gain. PMID- 15277669 TI - Integrin beta3 regions controlling binding of murine mAb 7E3: implications for the mechanism of integrin alphaIIbbeta3 activation. AB - Abciximab, a derivative of the murine mAb 7E3, protects against ischemic complications of percutaneous coronary interventions by inhibiting ligand binding to the alphaIIbbeta3 receptor. In this study we identified regions on integrin beta3 that control 7E3 binding. Murine/human amino acid substitutions were created in two regions of the betaA domain that previous studies found to influence 7E3 binding: the C177-C184 loop and K125-N133. The T182N substitution and a K125Q mutation reduced 7E3 binding to human beta3 in complex with alphaIIb. The introduction of both the human C177-C184 region and human W129 into murine beta3 was necessary and sufficient to permit 7E3 binding to the human alphaIIb/murine beta3 complex. Although we cannot exclude allosteric effects, we propose that 7E3 binds between C177-C184 and W129, which are within 15 A of each other in the crystal structure and close to the beta3 metal ion-dependent adhesion site. We previously demonstrated that 7E3 binds more rapidly to activated than unactivated platelets. Because it has been proposed that alphaIIbbeta3 changes from a bent to an extended conformation upon activation, we hypothesized that 7E3 binds less well to the bent than the extended conformation. In support of this hypothesis we found that 7E3 bound less well to an alphaIIbbeta3 construct locked in a bent conformation, and unlocking the conformation restored 7E3 binding. Thus, our data are consistent with alphaIIbbeta3 existing in variably bent conformations in equilibrium with each other on unactivated platelets, and activation resulting in alphaIIbbeta3 adopting a more extended conformation. PMID- 15277670 TI - The SapB morphogen is a lantibiotic-like peptide derived from the product of the developmental gene ramS in Streptomyces coelicolor. AB - SapB is a morphogenetic peptide that is important for aerial mycelium formation by the filamentous bacterium Streptomyces coelicolor. Production of SapB commences during aerial mycelium formation and depends on most of the genes known to be required for the morphogenesis of aerial hyphae. Furthermore, the application of purified SapB to mutants blocked in morphogenesis restores their capacity to form aerial hyphae. Here, we present evidence that SapB is a lantibiotic-like peptide that is derived by posttranslational modification from the product of a gene (ramS) in the four-gene ram operon, which is under the control of the regulatory gene ramR. We show that the product of another gene in the operon (ramC) contains a region that is similar to enzymes involved in the biosynthesis of lantibiotics, suggesting that it might be involved in the posttranslational processing of RamS. We conclude that SapB is derived from RamS through proteolytic cleavage and the introduction of four dehydroalanine residues and two lanthionine bridges. We provide an example of a morphogenetic role for an antibiotic-like molecule. PMID- 15277671 TI - Mapping glycoside hydrolase substrate subsites by isothermal titration calorimetry. AB - Relating thermodynamic parameters to structural and biochemical data allows a better understanding of substrate binding and its contribution to catalysis. The analysis of the binding of carbohydrates to proteins or enzymes is a special challenge because of the multiple interactions and forces involved. Isothermal titration calorimetry (ITC) provides a direct measure of binding enthalpy (DeltaHa) and allows the determination of the binding constant (free energy), entropy, and stoichiometry. In this study, we used ITC to elucidate the binding thermodynamics of xylosaccharides for two xylanases of family 10 isolated from Geobacillus stearothermophilus T-6. The change in the heat capacity of binding (DeltaCp = DeltaH/DeltaT) for xylosaccharides differing in one sugar unit was determined by using ITC measurements at different temperatures. Because hydrophobic stacking interactions are associated with negative DeltaCp, the data allow us to predict the substrate binding preference in the binding subsites based on the crystal structure of the enzyme. The proposed positional binding preference was consistent with mutants lacking aromatic binding residues at different subsites and was also supported by tryptophan fluorescence analysis. PMID- 15277672 TI - Infecting the brain to stop addiction? PMID- 15277673 TI - In vivo positron-emission tomography imaging of progression and transformation in a mouse model of mammary neoplasia. AB - Imaging mouse models of human cancer promises more effective analysis of tumor progression and reduction of the number of animals needed for statistical power in preclinical therapeutic intervention trials. This study utilizes positron emission tomography imaging of 2-[18F]-fluoro-deoxy-D-glucose to monitor longitudinal development of mammary intraepithelial neoplasia outgrowths in immunocompetent FVB/NJ mice. The mammary intraepithelial neoplasia outgrowth tissues mimic the progression of breast cancer from premalignant ductal carcinoma in situ to invasive carcinoma. Progression of disease is clearly evident in the positron emission tomography images, and tracer uptake correlates with histological evaluation. Furthermore, quantitative markers of disease extracted from the images can be used to track proliferation and progression in vivo over multiple time points. PMID- 15277674 TI - RNA dynamics in live Escherichia coli cells. AB - We describe a method for tracking RNA molecules in Escherichia coli that is sensitive to single copies of mRNA, and, using the method, we find that individual molecules can be followed for many hours in living cells. We observe distinct characteristic dynamics of RNA molecules, all consistent with the known life history of RNA in prokaryotes: localized motion consistent with the Brownian motion of an RNA polymer tethered to its template DNA, free diffusion, and a few examples of polymer chain dynamics that appear to be a combination of chain fluctuation and chain elongation attributable to RNA transcription. We also quantify some of the dynamics, such as width of the displacement distribution, diffusion coefficient, chain elongation rate, and distribution of molecule numbers, and compare them with known biophysical parameters of the E. coli system. PMID- 15277675 TI - Mechanical switching and coupling between two dissociation pathways in a P selectin adhesion bond. AB - Many biomolecular bonds exhibit a mechanical strength that increases in proportion to the logarithm of the rate of force application. Consistent with exponential decrease in bond lifetime under rising force, this kinetically limited failure reflects dissociation along a single thermodynamic pathway impeded by a sharp free energy barrier. Using a sensitive force probe to test the leukocyte adhesion bond P-selectin glycoprotein ligand 1 (PSGL-1)-P-selectin, we observed a linear increase of bond strength with each 10-fold increase in the rate of force application from 300 to 30,000 pN/sec, implying a single pathway for failure. However, the strength and lifetime of PSGL-1-P-selectin bonds dropped anomalously when loaded below 300 pN/sec, demonstrating unexpectedly faster dissociation and a possible second pathway for failure. Remarkably, if first loaded by a "jump" in force to 20-30 pN, the bonds became strong when subjected to a force ramp as slow as 30 pN/sec and exhibited the same single pathway kinetics under all force rates. Applied in this way, a new "jump/ramp" mode of force spectroscopy was used to show that the PSGL-1-P-selectin bond behaves as a mechanochemical switch where force history selects between two dissociation pathways with markedly different properties. Furthermore, replacing PSGL-1 by variants of its 19-aa N terminus and by the crucial tetrasaccharide sialyl LewisX produces dramatic changes in the failure kinetics, suggesting a structural basis for the two pathways. The two-pathway switch seems to provide a mechanism for the "catch bond" response observed recently with PSGL-1-P-selectin bonds subjected to small-constant forces. PMID- 15277676 TI - The educative impact of health care treatment on malarial prevention behavior for the poor in Guinea, West Africa. AB - We analyze the malarial health behavior of rural populations by using data from the 1999 Demographic and Health Survey for Guinea, West Africa. We find that prior formal health care treatment is associated with heightened malaria prevention behaviors for the poorest uneducated populations in this rural cohort. Individuals from this subgroup that report no history of malarial infection and exclude themselves from health care treatment further appear to be misdiagnosing the disease at a substantial level. We conjecture that the use of formal health care options provides informational exposure to the clinical aspects of malarial pathogenesis. For individuals steeped in the most severe poverty, this exposure appears to have a particularly robust educative effect. The health behavioral dynamics we observe here have putative extensions for regional health policy as well with other infectious diseases, such as HIV/AIDS. PMID- 15277677 TI - Carbohydrate mimicry between human ganglioside GM1 and Campylobacter jejuni lipooligosaccharide causes Guillain-Barre syndrome. AB - Molecular mimicry between microbial and self-components is postulated as the mechanism that accounts for the antigen and tissue specificity of immune responses in postinfectious autoimmune diseases. Little direct evidence exists, and research in this area has focused principally on T cell-mediated, antipeptide responses, rather than on humoral responses to carbohydrate structures. Guillain Barre syndrome, the most frequent cause of acute neuromuscular paralysis, occurs 1-2 wk after various infections, in particular, Campylobacter jejuni enteritis. Carbohydrate mimicry [Galbeta1-3GalNAcbeta1-4(NeuAcalpha2-3)Galbeta1-] between the bacterial lipooligosaccharide and human GM1 ganglioside is seen as having relevance to the pathogenesis of Guillain-Barre syndrome, and conclusive evidence is reported here. On sensitization with C. jejuni lipooligosaccharide, rabbits developed anti-GM1 IgG antibody and flaccid limb weakness. Paralyzed rabbits had pathological changes in their peripheral nerves identical with those present in Guillain-Barre syndrome. Immunization of mice with the lipooligosaccharide generated a mAb that reacted with GM1 and bound to human peripheral nerves. The mAb and anti-GM1 IgG from patients with Guillain-Barre syndrome did not induce paralysis but blocked muscle action potentials in a muscle-spinal cord coculture, indicating that anti-GM1 antibody can cause muscle weakness. These findings show that carbohydrate mimicry is an important cause of autoimmune neuropathy. PMID- 15277678 TI - Venomous protease of aphid soldier for colony defense. AB - In social aphids, morphological, behavioral, and physiological differences between soldiers and normal insects are attributed to differences in gene expression between them, because they are clonal offspring parthenogenetically produced by the same mothers. By using cDNA subtraction, we identified a soldier specific cysteine protease of the family cathepsin B in a social aphid, Tuberaphis styraci, with a second-instar soldier caste. The cathepsin B gene was specifically expressed in soldiers and first-instar nymphs destined to be soldiers. The cathepsin B protein was preferentially produced in soldiers and showed a protease activity typical of cathepsin B. The cathepsin B mRNA and protein were localized in the midgut of soldiers. For colony defense, soldiers attack enemies with their stylet, which causes paralysis and death of the victims. Notably, after soldiers attacked moth larvae, the cathepsin B protein was detected from the paralyzed larvae. Injection of purified recombinant cathepsin B protein certainly killed the recipient moth larvae. From these results, we concluded that the cathepsin B protein is a major component of the aphid venom produced by soldiers of T. styraci. Soldier-specific expression of the cathepsin B gene was found in other social aphids of the genus Tuberaphis. The soldier-specific cathepsin B gene showed an accelerated molecular evolution probably caused by the action of positive selection, which had been also known from venomous proteins of other animals. PMID- 15277679 TI - Venezuelan equine encephalitis emergence: enhanced vector infection from a single amino acid substitution in the envelope glycoprotein. AB - In 1993 and 1996, subtype IE Venezuelan equine encephalitis (VEE) virus caused epizootics in the Mexican states of Chiapas and Oaxaca. Previously, only subtype IAB and IC VEE virus strains had been associated with major outbreaks of equine and human disease. The IAB and IC epizootics are believed to emerge via adaptation of enzootic (sylvatic, equine-avirulent) strains for high titer equine viremia that results in efficient infection of mosquito vectors. However, experimental equine infections with subtype IE equine isolates from the Mexican outbreaks demonstrated neuro-virulence but little viremia, inconsistent with typical VEE emergence mechanisms. Therefore, we hypothesized that changes in the mosquito vector host range might have contributed to the Mexican emergence. To test this hypothesis, we evaluated the susceptibility of the most abundant mosquito in the deforested Pacific coastal locations of the VEE outbreaks and a proven epizootic vector, Ochlerotatus taeniorhynchus. The Mexican epizootic equine isolates exhibited significantly greater infectivity compared with closely related enzootic strains, supporting the hypothesis that adaptation to an efficient epizootic vector contributed to disease emergence. Reverse genetic studies implicated a Ser --> Asn substitution in the E2 envelope glycoprotein as the major determinant of the increased vector infectivity phenotype. Our findings underscore the capacity of RNA viruses to alter their vector host range through minor genetic changes, resulting in the potential for disease emergence. PMID- 15277680 TI - Reinitiation involving upstream ORFs regulates ATF4 mRNA translation in mammalian cells. AB - During cellular stresses, phosphorylation of eukaryotic initiation factor-2 (eIF2) elicits gene expression designed to ameliorate the underlying cellular disturbance. Central to this stress response is the transcriptional regulator activating transcription factor, ATF4. Here we describe the mechanism regulating ATF4 expression involving the differential contribution of two upstream ORFs (uORFs) in the 5' leader of the mouse ATF4 mRNA. The 5' proximal uORF1 is a positive-acting element that facilitates ribosome scanning and reinitiation at downstream coding regions in the ATF4 mRNA. When eIF2-GTP is abundant in nonstressed cells, ribosomes scanning downstream of uORF1 reinitiate at the next coding region, uORF2, an inhibitory element that blocks ATF4 expression. During stress conditions, phosphorylation of eIF2 and the accompanying reduction in the levels of eIF2-GTP increase the time required for the scanning ribosomes to become competent to reinitiate translation. This delayed reinitiation allows for ribosomes to scan through the inhibitory uORF2 and instead reinitiate at the ATF4 coding region. Increased expression of ATF4 would contribute to the expression of genes involved in remediation of cellular stress damage. These results suggest that the mechanism of translation reinitiation involving uORFs is conserved from yeast to mammals. PMID- 15277681 TI - The ion channel of F-ATP synthase is the target of toxic organotin compounds. AB - ATP is the universal energy currency of living cells, and the majority of it is synthesized by the F1F0 ATP synthase. Inhibitors of this enzyme are therefore potentially detrimental for all life forms. Tributyltin chloride (TBT-Cl) inhibits ATP hydrolysis by the Na(+)-translocating ATP synthase of Ilyobacter tartaricus or the H(+)-translocating counterpart of Escherichia coli with apparent Ki of 200 nM. To target the site of this inhibition, we synthesized a tritium-labeled derivative of TBT-Cl in which one of the butyl groups was replaced by a photoactivatable aryldiazirine residue. Upon illumination, subunit a of the ATP synthase becomes specifically modified, and this labeling is suppressed in the presence of the original inhibitor. In case of the Na+ ATP synthase, labeling is also suppressed in the presence of Na+ ions, suggesting an interference in Na+ or TBT-Cl binding to subunit a. This interference is corroborated by the protection of ATP hydrolysis from TBT-Cl inhibition by 105 mM Na+. TBT-Cl strongly inhibits Na+ exchange by the reconstituted I. tartaricus ATP synthase. Taken together these results indicate that the subunit a ion channel is the target site for ATPase inhibition by toxic organotin compounds. An inhibitor interacting specifically with this site has not been reported previously. PMID- 15277682 TI - Subfertility and defective folliculogenesis in female mice lacking androgen receptor. AB - The roles of the androgen receptor (AR) in female fertility and ovarian function remain largely unknown. Here we report on the generation of female mice lacking AR (AR(-/-)) and the resulting influences on the reproductive system. Female AR( /-) mice appear normal but show longer estrous cycles and reduced fertility. The ovaries from sexually mature AR(-/-) females exhibited a marked reduction in the number of corpora lutea. After superovulation treatment, the AR(-/-) ovaries produced fewer oocytes and also showed fewer corpora lutea. During the periovulatory period, an intensive granulosa apoptosis event occurs in the AR(-/ ) preovulatory follicles, concurrent with the down-regulation of p21 and progesterone receptor expression. Furthermore, the defective conformation of the cumulus cell-oocyte complex from the AR(-/-) females implies a lower fertilization capability of the AR(-/-) oocytes. In addition to insufficient progesterone production, the diminished endometrial growth in uteri in response to exogenous gonadotropins indicates that AR(-/-) females exhibit a luteal phase defect. Taken together, these data provide in vivo evidence showing that AR plays an important role in female reproduction. PMID- 15277683 TI - G protein-coupled receptors: in silico drug discovery in 3D. AB - The application of structure-based in silico methods to drug discovery is still considered a major challenge, especially when the x-ray structure of the target protein is unknown. Such is the case with human G protein-coupled receptors (GPCRs), one of the most important families of drug targets, where in the absence of x-ray structures, one has to rely on in silico 3D models. We report repeated success in using ab initio in silico GPCR models, generated by the predict method, for blind in silico screening when applied to a set of five different GPCR drug targets. More than 100,000 compounds were typically screened in silico for each target, leading to a selection of <100 "virtual hit" compounds to be tested in the lab. In vitro binding assays of the selected compounds confirm high hit rates, of 12-21% (full dose-response curves, Ki < 5 microM). In most cases, the best hit was a novel compound (New Chemical Entity) in the 1- to 100-nM range, with very promising pharmacological properties, as measured by a variety of in vitro and in vivo assays. These assays validated the quality of the hits as lead compounds for drug discovery. The results demonstrate the usefulness and robustness of ab initio in silico 3D models and of in silico screening for GPCR drug discovery. PMID- 15277684 TI - A naphthyridine carboxamide provides evidence for discordant resistance between mechanistically identical inhibitors of HIV-1 integrase. AB - The increasing incidence of resistance to current HIV-1 therapy underscores the need to develop antiretroviral agents with new mechanisms of action. Integrase, one of three viral enzymes essential for HIV-1 replication, presents an important yet unexploited opportunity for drug development. We describe here the identification and characterization of L-870,810, a small-molecule inhibitor of HIV-1 integrase with potent antiviral activity in cell culture and good pharmacokinetic properties. L-870,810 is an inhibitor with an 8-hydroxy-(1,6) naphthyridine-7-carboxamide pharmacophore. The compound inhibits HIV-1 integrase mediated strand transfer, and its antiviral activity in vitro is a direct consequence of this ascribed effect on integration. L-870,810 is mechanistically identical to previously described inhibitors from the diketo acid series; however, viruses selected for resistance to L-870,810 contain mutations (integrase residues 72, 121, and 125) that uniquely confer resistance to the naphthyridine. Conversely, mutations associated with resistance to the diketo acid do not engender naphthyridine resistance. Importantly, the mutations associated with resistance to each of these inhibitors map to distinct regions within the integrase active site. Therefore, we propose a model of the two inhibitors that is consistent with this observation and suggests specific interactions with discrete binding sites for each ligand. These studies provide a structural basis and rationale for developing integrase inhibitors with the potential for unique and nonoverlapping resistance profiles. PMID- 15277685 TI - Discriminatory aptamer reveals serum response element transcription regulated by cytohesin-2. AB - Cytohesins are a family of highly homologous guanine nucleotide exchange factors (GEFs) that act on ADP-ribosylation factors (ARFs). The small ARF-GEFs are involved in integrin signaling, actin cytoskeleton remodeling, and vesicle transport. Here, we selected and applied a specific inhibitor for ARF nucleotide binding site opener (ARNO)/cytohesin-2, an RNA aptamer that clearly discriminates between cytohesin-1 and cytohesin-2. This reagent bound to an N-terminal segment of cytohesin-2 and did not inhibit ARF-GEF function in vitro. When transfected into HeLa cells, it persisted for at least 6 h without requiring stabilization. Its effect in vivo was to down-regulate gene expression mediated through the serum-response element and knockdown mitogen-activated protein kinase activation, indicating that cytohesin-2 acts by means of mitogen-activated protein kinase signaling. We conclude that the N-terminal coiled-coil and parts of the Sec7 domain of cytohesin-2 are required for serum-mediated transcriptional activation in nonimmune cells, whereas cytohesin-1 is not. Our results indicate that intramer technology can be used not only for assigning novel biological functions to proteins or protein domains but also to prove nonredundancy of highly homologous proteins. PMID- 15277686 TI - Transcriptional repression of target genes by LEUNIG and SEUSS, two interacting regulatory proteins for Arabidopsis flower development. AB - Transcription repression plays important roles in preventing crucial regulatory proteins from being expressed in inappropriate temporal or spatial domains. LEUNIG (LUG) and SEUSS (SEU) normally act to prevent ectopic expression of the floral homeotic gene AGAMOUS in flowers. LUG encodes a protein with sequence similarities to the yeast Tup1 corepressor. SEU encodes a plant-specific regulatory protein with sequence similarity in a conserved dimerization domain to the LIM-domain binding 1/Chip proteins in mouse and Drosophila. Despite the molecular isolation of LUG and SEU, the biochemical function of these two proteins remains uncharacterized, and the mechanism of AGAMOUS repression remains unknown. Here, we report that LUG and SEU interact directly in vitro and in vivo. Furthermore, LUG exhibits a strong repressor activity on several heterologous promoters in yeast and plant cells. SEU, in contrast, does not exhibit any direct repressor activity, but can repress reporter gene expression only in the presence of LUG, indicating a possible role of SEU as an adaptor protein for LUG. Our results demonstrate that LUG encodes a functional homologue of Tup1 and that SEU may function similarly to Ssn6, an adaptor protein of Tup1. We have defined the LUG/LUH, Flo8, single-strand DNA-binding protein domain of LUG as both necessary and sufficient for the interaction with SEU and two domains of LUG as important for its repressor function. Our work provides functional insights into plant transcriptional corepressors and reveals both conservation and distinctions between plant corepressors and those of yeast and animals. PMID- 15277687 TI - Pheromone anosmia in a scarab beetle induced by in vivo inhibition of a pheromone degrading enzyme. AB - Previous biochemical evidence suggests that a cytochrome P450 specific to male antennae of the pale-brown chafer, Phyllopertha diversa, has evolved as a pheromone-degrading enzyme. By using a bioinformatics approach, we have now cloned three P450 cDNAs: CYP4AW1, CYP4AW2, and CYP6AT1. RT-PCR indicated that CYP4AW2 is expressed in all tissues examined, that CYP6AT1 is antennae-rich, and that CYP4AW1 is antennae-specific. Both tissue specificity and electrophysiological studies strongly support that CYP4AW1 in P. diversa is a pheromone-degrading enzyme involved in pheromone inactivation. Highly sensitive, pheromone-specific olfactory receptor neurons in male antennae were completely desensitized by direct application of metyrapone into the sensillar lymph. When tested in the same or different individuals, the metyrapone treatment had no effect on olfactory receptor neurons tuned to the plant volatile (Z)-3-hexenyl acetate, which might be inactivated by an esterase. Metyrapone treatment did not affect pheromone reception in the Japanese beetle, Popillia japonica, in the scarab beetle, Anomala octiescostata, or in the Oriental beetle, Exomala orientalis. Metyrapone-induced anosmia was restricted to the pheromone detectors in P. diversa, which became insensitive to physiological concentrations of pheromones for a few minutes. As opposed to previous trials, the specificity of the inhibitor and pheromone system led to unambiguous evidence for the role of pheromone-degrading enzymes in the fast inactivation of pheromones. PMID- 15277688 TI - Specific and differential inhibition of very-long-chain fatty acid elongases from Arabidopsis thaliana by different herbicides. AB - In higher plants, very-long-chain fatty acids (VLCFAs) are the main constituents of hydrophobic polymers that prevent dessication at the leaf surface and provide stability to pollen grains. Of the 21 genes encoding VLCFA elongases (VLCFAEs) from Arabidopsis thaliana, 17 were expressed heterologously in Saccharomyces cerevisiae. Six VLCFAEs, including three known elongases (FAE1, KCS1, and KCS2) and three previously uncharacterized gene products (encoded by At5g43760, At1g04220, and At1g25450) were found to be enzymatically active with endogenous yeast fatty acid substrates and to some extent with externally supplied unsaturated substrates. The spectrum of VLCFAs accumulated in expressing yeast strains was determined by gas chromatography/mass spectrometry. Marked specificity was found among elongases tested with respect to their elongation products, which encompassed saturated and monounsaturated fatty acids 20-30 carbon atoms in length. The active VLCFAEs revealed highly distinct patterns of differential sensitivity to oxyacetamides, chloroacetanilides, and other compounds tested, whereas yeast endogenous VLCFA production, which involves its unrelated elongase (ELO) in sphingolipid synthesis, was unaffected. Several compounds inhibited more than one VLCFAE, and some inhibited all six active enzymes. These findings pinpoint VLCFAEs as the target of the widely used K(3) class herbicides, which have been in commercial use for 50 years, provide important clues as to why spontaneous resistance to this class is rare, and point to complex patterns of substrate specificity and product spectrum among members of the Arabidopsis VLCFAE family. PMID- 15277689 TI - Toward the synthesis of the carbacylic ansa antibiotic kendomycin. AB - The convergent synthesis of a benzofuran analog of the carbacyclic ansa compound kendomycin has been achieved by assembling three major fragments by means of epoxide opening and directed ortho lithiation. The crucial tetrahydropyran ring was formed by a highly stereoselective cationic cyclization. Analysis of all synthesized tetrahydropyran-arene compounds reveals a hindered sp(2)-sp(3) rotation, which results in rotational isomers or atropisomers affecting macrocyclization reactions. The latter could only be achieved by means of Horner Wadsworth-Emmons olefination. PMID- 15277690 TI - Human placenta feeder layers support undifferentiated growth of primate embryonic stem cells. AB - Various undifferentiated embryonic stem (ES) cells can grow on mouse embryonic fibroblast (MEF) feeders. However, the risk of zoonosis from animal feeders to human ES cells generally excludes the clinical use of these human ES cells. We have found that human placenta is a useful source of feeder cells for the undifferentiated growth of primate ES cells. As on MEF feeders, primate ES cells cultured on human amniotic epithelial (HAE) feeder cells and human chorionic plate (HCP) cells had undifferentiated growth. The cultured primate ES cells expressed Oct-4, alkaline phosphatase, and SSEA-4. The primate ES cells on HAE feeder cells produced typical immature teratomas in vivo after injection into severe combined immunodeficient mice. Human placenta is quite novel and important because it would provide a relatively available source of feeders for the growth of human ES cells for therapeutic purposes that are also free of ethical complications. PMID- 15277691 TI - Breaking the species barrier: derivation of germline-competent embryonic stem cells from Mus spretus x C57BL/6 hybrids. AB - Embryonic stem (ES) cells, which can differentiate into almost all types of cells, have been derived from the house mouse Mus musculus, rat, rabbit, humans, and other species. Transmission of the genotype to the offspring of chimeras has been achieved only with M. musculus ES cells, limiting targeted mutagenesis using ES cells to this species. Mus spretus, which exhibits many genetic polymorphisms with M. musculus, displays dominant resistance to cancer and inflammation, making derived inbred strains very useful in positional cloning and interspecies mapping. We show here for the first time the derivation of ES cells from hybrid blastocysts, obtained by the mating of two different species, namely Mus musculus and Mus spretus, and their use for the generation of chimeric mice that transmit the Mus spretus genotype and phenotype to the offspring. These hybrid ES cells allow the genetic manipulation of Mus spretus, as an alternative to Mus musculus. PMID- 15277692 TI - Human embryonic stem cells possess immune-privileged properties. AB - Human embryonic stem cells (hESCs) are envisioned to be a major source for cell based therapies. Efforts to overcome rejection of hESCs include nuclear transfer and collection of hESC banks representing the broadest diversity of major histocompatability complex (MHC) polymorphorisms. Surprisingly, immune responses to hESCs have yet to be experimentally evaluated. Here, injection of hESCs into immune-competent mice was unable to induce an immune response. Undifferentiated and differentiated hESCs failed to stimulate proliferation of alloreactive primary human T cells and inhibited third-party allogeneic dendritic cell mediated T-cell proliferation via cellular mechanisms independent of secreted factors. Upon secondary rechallenge, T cells cocultured with hESCs were still responsive to allogeneic stimulators but failed to proliferate upon re-exposure to hESCs. Our study demonstrates that hESCs possess unique immune-privileged characteristics and provides an unprecedented opportunity to further investigate the mechanisms of immune response to transplantation of hESCs that may avoid immune-mediated rejection. PMID- 15277693 TI - BMP4: its role in development of the hematopoietic system and potential as a hematopoietic growth factor. AB - Blood formation occurs throughout the life of an individual in a process driven by hematopoietic stem cells (HSCs). The ability of bone marrow (BM) and cord blood (CB) HSC to undergo self-renewal and develop into multiple blood lineages has made these cells an important clinical resource. Transplantation with BM- and CB-derived HSCs is now used extensively for treatment of hematological disorders, malignancies, and immunodeficiencies. An understanding of the embryonic origin of HSC and the factors regulating their generation and expansion in vivo will provide important information for the manipulation of these cells ex vivo. This is critical for the further development of CB transplantation, the potential of which is limited by small numbers of HSC in the donor population. Although the origins of HSCs have become clearer and progress has been made in identifying genes that are critical for the formation and maintenance of HSCs, less is known about the signals that commit specific populations of mesodermal precursors to hematopoietic cell fate. Critical signals acting on these precursor cells are likely to be derived from visceral endoderm in yolk sac and from underlying stroma in the aorta-gonad-mesonephros region. Here we summarize briefly the origin of yolk sac and embryonic HSCs before detailing evidence that bone morphogenic protein-4 (BMP4) has a crucial role in Xenopus and mammalian HSC development. We discuss evidence that BMP4 acts as a hematopoietic growth factor and review its potential to modulate HSC in ex vivo expansion cultures from cord blood. PMID- 15277694 TI - Dendritic cell development in long-term spleen stromal cultures. AB - The cellular microenvironments in which dendritic cells (DCs) develop are not known. DCs are commonly expanded from CD34+ bone marrow precursors or blood monocytes using a cocktail of growth factors including GM-CSF. However, cytokine supported cultures are not suitable for studying the intermediate stages of DC development, since progenitors are quickly driven to become mature DCs that undergo limited proliferation and survive for only a short period of time. This lab has developed a long-term culture (LTC) system from spleen which readily generates a high yield of DCs. Hematopoietic cells develop under more normal physiological conditions than in cultures supplemented with cytokines. A spleen stromal cell monolayer supports stem cell maintenance, renewal, and the specific differentiation of only DCs and no other hematopoietic cells. Cultures maintain continuous production of a small population of small-sized progenitors and a large population of fully developed DCs. Cell-cell interaction between stromal cells and progenitor cells is critical for DC differentiation. The progenitors maintained in LTC appear to be quite distinct from bone marrow-derived DC progenitors that respond to GM-CSF. The majority of cells produced in LTC are large-sized cells with a phenotype reflecting myeloid-like DC precursors or immature DCs. These cells are highly endocytotic and weakly immunostimulatory for T cells. This model system predicts in situ production of DCs in spleen from endogenous progenitors, as well as a central role for spleen in DC hematopoiesis. PMID- 15277695 TI - Plasticity of bone marrow-derived stem cells. AB - Stem cell plasticity refers to the ability of adult stem cells to acquire mature phenotypes that are different from their tissue of origin. Adult bone marrow cells (BMCs) include two populations of bone marrow stem cells (BMCs): hematopoietic stem cells (HSCs), which give rise to all mature lineages of blood, and mesenchymal stem cells (MSCs), which can differentiate into bone, cartilage, and fat. In this article, we review the literature that lends credibility to the theory that highly plastic BMCs have a role in maintenance and repair of nonhematopoietic tissue. We discuss the possible mechanisms by which this may occur. Also reviewed is the possibility that adult BMCs can change their gene expression profile after fusion with a mature cell, which has brought into question whether this stem cell plasticity is real. PMID- 15277696 TI - Cellular immunotherapy with dendritic cells in cancer: current status. AB - Dendritic cells (DCs) are specialized antigen-presenting cells whose immunogenicity leads to the induction of antigen-specific immune responses. DCs can easily be generated ex vivo from peripheral blood monocytes or bone marrow/circulating hematopoietic stem cells cultured in the presence of cytokine cocktails. DCs have been used in numerous clinical trials to induce antitumor immune responses in cancer patients. The studies carried out to date have demonstrated that DCs pulsed with tumor antigens can be safely administered, and this approach produces antigen-specific immune responses. Clinical responses have been observed in a minority of patients. It is likely that either heavy medical pretreatment or the presence of large tumor burdens (or both) is among the causes that impair the benefits of vaccination. Hence, the use of DCs should be considered in earlier stages of disease such as the adjuvant setting. Prospective applications of DCs extend to their use in allogeneic adoptive immunotherapy to specifically target the graft versus tumor reaction. DCs continue to hold promise for cellular immunotherapy, and further investigation is required to determine the clinical settings in which patients will most benefit from the use of this cellular immune adjuvant. PMID- 15277697 TI - Changes in gene expression at the precursor --> stem cell transition in leech. AB - The glossiphoniid leech, Theromyzon trizonare, displays particularly large and accessible embryonic precursor/stem cells during its early embryonic cleavages. We dissected populations of both cell types from staged embryos and examined gene expression profiles by differential display polymerase chain reaction methodology. Among the approximately 10,000 displayed cDNA fragments, 56 (approximately 0.5%) were differentially expressed at the precursor --> stem cell transition; 29 were turned off (degraded, precursor-specific); and 27 were turned on (transcribed, stem cell-specific). Several putative differentially expressed cDNAs from each category were confirmed by Northern blot analysis on staged embryos. DNA sequencing revealed that 19 of the cDNAs were related to a spectrum of genes including the CCR4 antiproliferation gene, Rad family members, and several transcriptional regulators, while the remainder encoded hypothetical (10) or novel (27) sequences. Collectively, these results identify dynamic changes in gene expression during stem cell formation in leech and provide a platform for examining the molecular aspects of stem cell genesis in a simple invertebrate organism. PMID- 15277698 TI - Maintenance of pluripotency in human embryonic stem cells is STAT3 independent. AB - The preservation of "stemness" in mouse embryonic stem (mES) cells is maintained through a signal transduction pathway that requires the gp130 receptor, the interleukin-6 (IL-6) family of cytokines, and the Janus Kinase-signal transducer and activator (JAK/STAT) pathway. The factors and signaling pathways that regulate "stemness" in human embryonic stem (hES) cells remain to be elucidated. Here we report that STAT3 activation is not sufficient to block hES cell differentiation when the cells are grown on mouse feeder cells or when they are treated with conditioned media from feeder cells. Human ES cells differentiate in the presence of members of the IL-6 family of cytokines including leukemia inhibitory factor (LIF) and IL-6 or in the presence of the designer cytokine hyper-IL-6, which is a complex of soluble interleukin-6 receptor (IL-6R) and IL-6 with greatly enhanced bioactivity. Human ES cells express LIF, IL-6, and gp130 receptors, as well as the downstream signaling molecules. Stimulation of human and mouse ES cells with gp130 cytokines resulted in a robust phosphorylation of downstream ERK1, ERK2, and Akt kinases, as well as the STAT3 transcription factor. Loss of the pluripotency markers Nanog, Oct-4, and TRA-1-60 was observed in hES cells during gp130-dependent signaling, indicating that signaling through this pathway is insufficient to prevent the onset of differentiation. These data underscore a fundamental difference in requirements of murine versus hES cells. Furthermore, the data demonstrate the existence of an as-yet-unidentified factor in the conditioned media of mouse feeder layer cells that acts to maintain hES cell renewal in a STAT3-independent manner. PMID- 15277699 TI - Efficient transfection of embryonic and adult stem cells. AB - The ability of embryonic stem cells and adult stem cells to differentiate into specific cell types holds immense potential for therapeutic use in cell and gene therapy. Realization of this potential depends on efficient and optimized protocols for genetic manipulation of stem cells. In the study reported here, we demonstrate the use of nucleofection as a method to introduce plasmid DNA into embryonic and adult stem cells with significantly greater efficiency than electroporation or lipid-based transfection methods have. Using enhanced green fluorescent protein (eGFP) as a reporter gene, mouse embryonic stem cells were transfected both transiently and stably at a rate nearly 10-fold higher than conventional methods. The transfected cells retained their stem cell properties, including continued expression of the stem cell markers SSEA1, Oct4, and Rex1; formation of embryoid bodies; differentiation into cardiomyocytes in the presence of appropriate inducers; and, when injected into developing blastocysts, contribution to chimeras. Higher levels of transfection were also obtained with human embryonic carcinoma and human embryonic stem cells. Particularly hard-to transfect adult stem cells, including bone marrow and multipotent adult progenitor cells, were also transfected efficiently by the method of nucleofection. Based on our results, we conclude that nucleofection is superior to currently available methods for introducing plasmid DNA into a variety of embryonic and adult stem cells. The high levels of transfection achieved by nucleofection will enable its use as a rapid screening tool to evaluate the effect of ectopically expressed transcription factors on tissue-specific differentiation of stem cells. PMID- 15277700 TI - Hyaluronic acid facilitates the recovery of hematopoiesis following 5 fluorouracil administration. AB - The fate of hematopoietic stem cells (HSCs) is determined by microenvironmental niches, but the molecular structure of these local networks is not yet completely characterized. Our recent observation that glycosaminoglycan hyaluronic acid (HA), a major component of the bone marrow extracellular matrix, is required for in vitro hematopoiesis led us to suggest a role for HA in structuring the hematopoietic niche. Accordingly, HA deprivation induced by various treatments might lead to an imbalance of normal HSC homeostasis. Since 5-fluorouracil (5-FU) administration sharply decreases the amount of cell surface-associated HA in bone marrow, we examined whether the administration of exogenous HA enhances suppressed hematopoiesis in 5-FU-treated mice. HA administered to mice following 5-FU infusion facilitated the recovery of leukocytes and thrombocytes in the peripheral blood. Intravenously infused HA was found in the bone marrow, where it bound endothelial cells and resident macrophages and increased expression of the hematopoiesis-supportive cytokines interleukin-1 and interleukin-6. In agreement with these observations, enhanced hematopoietic activity was detected in the bone marrow, as measured by elevated counts of long-term culture-initiating cells (LTC ICs), committed progenitors, and the total number of mature bone marrow cells. Overall, our results suggest that HA is required for regulation of the hematopoiesis-supportive function of bone marrow accessory cells and, therefore, participates in hematopoietic niche assembly. PMID- 15277701 TI - Analysis of primitive CD34- and CD34+ hematopoietic cells from adults: gain and loss of CD34 antigen by undifferentiated cells are closely linked to proliferative status in culture. AB - There is limited understanding of CD34- hematopoietic cells and the linkage between CD34 antigen expression and cell proliferation. In this study, early CD34 CD38- LIN- (CD34-) cells were purified from mobilized adult peripheral blood and carefully analyzed in vitro for growth and modulation of CD34. Mobilized CD34+CD38- LIN- (CD34+) cells were used for comparison. Expression of CD34, CD38, and LIN antigens was determined, and proliferative responses were assessed with PKH tracking dye, expression of Ki67 antigen, and uptake of pyronin Y. Suspension cultures of adult CD34- cells generated CD34+ cells and progenitors for >8 weeks. Stromal cultures demonstrated the presence of long-term culture-initiating cells within the CD34- fraction. While CD34- cells were slower to initiate growth than the CD34+ cells were, no significant difference in hematopoietic cell output was found. Upon cultivation of CD34- cells, CD34 antigen appeared within 48 hours but was restricted to those cells that had initiated growth. Surprisingly, CD34+ precursors lost CD34 expression in culture if they remained in G0 for more than 2 days. Those cells later regained expression of CD34 antigen upon initiation of growth. Comparison of cells that did or did not rapidly modulate CD34 antigen revealed no differences in long-term growth potential. In conclusion, in vitro expression of CD34 by CD34- and CD34+ populations is tightly linked to cellular proliferation. In this culture system, expression of CD34 antigen by LIN- cells constitutes an early hallmark of growth. Measurement of CD34 expression by LIN- cells in expansion culture underestimates the total content of hematopoietic cells. PMID- 15277702 TI - Clonal analysis of individual marrow-repopulating cells after experimental peripheral blood progenitor cell transplantation. AB - Methods to analyze the clonality of an adverse event in preclinical or clinical retroviral stem cell gene therapy protocols are needed. We analyzed the progeny of retrovirally transduced human peripheral blood progenitor cells (PBPCs) after transplantation and engraftment in immune-deficient mice. The integration site of the provirus serves as a unique tag of the individual transduced PBPC. A plasmid library of junctions between proviral and human genomic DNA was generated. We were able to detect individual transduced cell clones that amounted to 0.14% 0.0001% of chimeric bone marrow cells. This is the first report in which the contribution of individual marrow-repopulating cells to human hematopoiesis is directly quantified. PMID- 15277703 TI - Expression and function of homing-essential molecules and enhanced in vivo homing ability of human peripheral blood-derived hematopoietic progenitor cells after stimulation with stem cell factor. AB - Hematopoietic stem cell (HSC) homing from blood to bone marrow is a multistep process involving rolling, extravasation, migration, and finally adhesion in the correct microenvironment. With view to the hematopoietic recovery after clinical stem cell transplantation, we investigated the effect of stem cell factor (SCF) on the expression and the adhesive function of the alpha4beta1 and alpha5beta1 integrins very-late antigen (VLA)-4 and VLA-5 on peripheral blood-derived hematopoietic progenitor cells. After SCF stimulation, the expression of VLA-4 and VLA-5 on CD34+/c-kit+ cells obtained from healthy donors increased from 54% to 90% and from 3% to 82%, respectively. For patient-derived cells, the increase was 67% to 90% and 12% to 46%. The proportion of mononuclear cells adhering to the fibronectin fragment CH296 increased by stimulation with SCF from 14% to 23%. Accordingly, functional studies showed an approximate 30% increase of adherent long-term culture-initiating cell. The improvement of the homing abilities of SCF stimulated HSC was confirmed by transplantation into sublethally irradiated nonobese diabetic-scid/scid mice. Six weeks after the transplantation, in eight of eight animals receiving human HSC with the addition of SCF, a profound multilineage hematopoietic engraftment was detected, whereas in the control group receiving only HSC, none of eight animals engrafted. Our data provide the first in vivo evidence that stimulation with cytokines improves the homing ability of transplanted human hematopoietic progenitor cells. PMID- 15277704 TI - The hematopoietic stem cell in myelodysplasia. AB - The mechanisms underlying hematopoietic stem cell or progenitor cell abnormalities in myelodysplastic syndromes (MDSs) remain poorly characterized. Current evidence exists for multiple intrinsic and extrinsic influences upon the stem cell in these disorders. These influences are outlined in this review and include: stem cell characteristics in MDSs, as compared with those in acute myelogenous leukemia; the role of increased apoptosis; the role of signaling pathway abnormalities; the influences of immune modulation; and the effect of stromal cells and stromal cell cytokine production. Despite numerous studies that have examined these factors, how they converge to produce a situation in which accelerated proliferation and accelerated death occur simultaneously remains largely an unexplored area. It is anticipated that future studies that focus on well-characterized and purified progenitor populations in these disorders will elucidate the process by which ineffective hematopoiesis results from the influences of stem cell abnormalities versus abnormalities in the stem cell's microenvironmental and immunologic milieu. PMID- 15277705 TI - Differentiation prevents assessment of neural stem cell pluripotency after blastocyst injection. AB - Earlier studies reported that neural stem (NS) cells injected into blastocysts appeared to be pluripotent, differentiating into cells of all three germ layers. In this study, we followed in vitro green fluorescent protein (GFP)-labeled NS and embryonic stem (ES) cells injected into blastocysts. Forty-eight hours after injection, significantly fewer blastocysts contained GFP-NS cells than GFP-ES cells. By 96 hours, very few GFP-NS cells remained in blastocysts compared with ES cells. Moreover, 48 hours after injection, GFP-NS cells in blastocysts extended long cellular processes, ceased expressing the NS cell marker nestin, and instead expressed the astrocytic marker glial fibrillary acidic protein. GFP ES cells in blastocysts remained morphologically undifferentiated, continuing to express the pluripotent marker stage-specific embryonic antigen-1. Selecting cells from the NS cell population that preferentially formed neurospheres for injection into blastocysts resulted in identical results. Consistent with this in vitro behavior, none of almost 80 mice resulting from NS cell-injected blastocysts replaced into recipient mothers were chimeric. These results strongly support the idea that NS cells cannot participate in chimera formation because of their rapid differentiation into glia-like cells. Thus, these results raise doubts concerning the pluripotency properties of NS cells. PMID- 15277706 TI - Erythropoietin overcomes imatinib-induced apoptosis and induces erythroid differentiation in TF-1/bcr-abl cells. AB - Targeting BCR-ABL tyrosine kinase by treatment with the selective inhibitor imatinib (formerly STI571, Gleevec) has proved to be highly efficient for inhibiting leukemic growth in vitro. In addition, in clinical trials, imatinib has produced high response rates in patients with chronic myeloid leukemia (CML) in chronic phase and blastic crisis. However, episodes of severe cytopenia were also frequently observed, leading to discontinuation of therapy in some cases. Therefore, it is important to examine whether administration of cytokines overcomes the adverse effects of imatinib in in vitro systems. In this study, we examine the effects of granulocyte-macrophage colony-stimulating factor (GM-CSF) and erythropoietin (EPO) on TF-1/bcr-abl (which was generated by transduction of a bcr-abl fusion gene into the TF-1 cell line) as a model system for CML with blastic crisis. Imatinib induced apoptosis in TF-1/bcr-abl cells but not in the parental TF-1 cells. However, GM-CSF, a survival factor of the parental TF-1 cells, protected TF-1/bcr-abl cells from imatinib-induced apoptosis in a dose dependent manner. Concomitantly, constitutive phosphorylation of Stat5 and FKHRL1 was significantly inhibited by imatinib, and the inhibition was canceled by the addition of GM-CSF, accompanied by upregulation of Bcl-xL and downregulation of p27/Kip1. In addition, although untreated TF-1/bcr-abl cells had lost responsiveness to both GM-CSF and EPO and showed autonomous growth, GM-CSF enhanced phosphorylation of Stat5 and FKHRL1 in these cells. Importantly, imatinib-treated TF-1/bcr-abl cells differentiated into hemoglobin-positive cells in the presence of EPO, as in the case for the parental TF-1 cells. Taken together, imatinib-treated CML cells may differentiate into mature cells in the presence of differentiation-inducing cytokines such as EPO. PMID- 15277707 TI - Skeletal myogenic differentiation of mesenchymal stem cells isolated from human umbilical cord blood. AB - Human umbilical cord blood (UCB) has been regarded as an alternative source for cell transplantation and cell therapy because of its hematopoietic and nonhematopoietic (mesenchymal) potential. Although there has been debate about whether mesenchymal stem cells (MSCs) are invariably present in UCB, several reports showed that MSC-like cells could be consistently derived from human UCB and, moreover, could differentiate into various cells of a mesodermal origin. However, it remains unclear whether these UCB-derived MSCs are also capable of differentiating into skeletal muscle cells. In this study, we isolated MSCs from human UCB and induced them to differentiate into skeletal muscle cells. During cell culture expansion, UCB-derived mononuclear cells gave rise to adherent layers of fibroblast-like cells expressing MSC-related antigens such as SH2, SH3, alpha-smooth muscle actin, CD13, CD29, and CD49e. More important, when these UCB derived MSCs were incubated in promyogenic conditions for up to 6 weeks, they expressed myogenic markers in accordance with myogenic differentiation pattern. Both flow cytometric and reverse transcriptase-polymerase reaction analyses showed that two early myogenic markers, MyoD and myogenin, were expressed after 3 days of incubation but not after 2 weeks. At week 6, more than half of UCB derived MSCs expressed myosin heavy chain, a late myogenic marker. Our results demonstrate that UCB-derived MSCs possess a potential of skeletal myogenic differentiation and also imply that these cells could be a suitable source for skeletal muscle repair and a useful tool of muscle-related tissue engineering. PMID- 15277708 TI - Critical parameters for the isolation of mesenchymal stem cells from umbilical cord blood. AB - Evidence has emerged that mesenchymal stem cells (MSCs) represent a promising population for supporting new clinical concepts in cellular therapy. However, attempts to isolate MSCs from umbilical cord blood (UCB) of full-term deliveries have previously either failed or been characterized by a low yield. We investigated whether cells with MSC characteristics and multi-lineage differentiation potential can be cultivated from UCB of healthy newborns and whether yields might be maximized by optimal culture conditions or by defining UCB quality criteria. Using optimized isolation and culture conditions, in up to 63% of 59 low-volume UCB units, cells showing a characteristic mesenchymal morphology and immune phenotype (MSC-like cells) were isolated. These were similar to control MSCs from adult bone marrow (BM). The frequency of MSC-like cells ranged from 0 to 2.3 clones per 1 x 10(8) mononuclear cells (MNCs). The cell clones proliferated extensively with at least 20 population doublings within eight passages. In addition, osteogenic and chondrogenic differentiation demonstrated a multi-lineage capacity comparable with BM MSCs. However, in contrast to MSCs, MSC-like cells showed a reduced sensitivity to undergo adipogenic differentiation. Crucial points to isolate MSC-like cells from UCB were a time from collection to isolation of less than 15 hours, a net volume of more than 33 ml, and an MNC count of more than 1 x 10(8) MNCs. Because MSC-like cells can be isolated at high efficacy from full-term UCB donations, we regard UCB as an additional stem cell source for experimental and potentially clinical purposes. PMID- 15277709 TI - Modeling for Lesch-Nyhan disease by gene targeting in human embryonic stem cells. AB - Human embryonic stem (ES) cells are pluripotent cells derived from blastocyst stage embryos. It has been suggested that these cells should play a major role in transplantation medicine and be able to advance our knowledge in human embryology. We propose that these cells should also play a vital role in the creation of models of human disorders. This aspect would be most valuable where animal models failed to faithfully recapitulate the human phenotype. Lesch-Nyhan disease is caused by a mutation in the HPRT1 gene that triggers an overproduction of uric acid, causing gout-like symptoms and urinary stones, in addition to neurological disorders. Due to biochemical differences between humans and rodents, a mouse lacking the HPRT expression will fail to accumulate uric acid. In this research we demonstrate a model for Lesch-Nyhan disease by mutating the HPRT1 gene in human ES cells using homologous recombination. We have verified the mutation in the HPRT1 allele at the DNA and RNA levels. By using selection media, we show that HPRT1 activity is abolished in the mutant cells, and the HPRT1-cells show a higher rate of uric acid accumulation than the wild-type cells. Therefore, these cells recapitulate to some extent the characteristics of Lesch-Nyhan syndrome and can help researchers further investigate this genetic disease and analyze drugs that will prevent the onset of its symptoms. We therefore suggest that human diseases may be modeled using human ES cells. PMID- 15277710 TI - The molecular perspective: nicotine and nitrosamines. PMID- 15277711 TI - Dr. Koscak Maruyama, Professor Emeritus of Chiba University. PMID- 15277712 TI - Hormonal control of meiosis initiation in the testis from Japanese newt, Cynops pyrrhogaster. AB - Meiosis is an event that occurs prerequisitely and specifically in gametogenesis. However, the mechanisms of conversion from mitosis to meiosis are poorly understood. I will review the results so far obtained by us using newt testis as a model system, and discuss about the extrinsic mechanism(s) controlling the conversion from mitosis to meiosis. In the newt spermatogonia enter meiosis in the 8th generation after 7 mitotic divisions. We developed organ and reaggregate culture systems with a chemically defined medium in which porcine follicle stimulating hormone (pFSH) promotes spermatogonial proliferation and differentiation into primary spermatocytes. Human recombinant stem cell factor (RhSCF) in vitro stimulates the spermatogonial proliferation and progression to the 7th generation, but not the differentiation into primary spermatocytes; instead they die of apoptosis. The reason why rhSCF does not stimulate meiosis entrance seems to be due to the low level expression of c-kit protein at the 7th generation of spermatogonia. Ovine PRL induces apoptosis in the 7th generation of spermatogonia in vivo and in vitro. Incubation of newts at low temperature causes spermatogonial apoptosis by the elevation of plasma PRL titer. In the absence of FSH in organ culture spermatogonia can progress until the 7th generation, but the 8th generation never appear due to the apoptosis. Altogether there seems to be a regulatory checkpoint for entrance into meiosis in the 7th generation. Spermatogonia could circumvent the checkpoint by the influence of some factor(s) produced by Sertoli cells upon activation by FSH. Trial to isolate factor(s) responsible for the meiosis-initiation is now underway. PMID- 15277713 TI - Copulation in the cricket is performed by chain reaction. AB - The male and female genitalia are finely designed to match each other for copulation in the cricket Gryllus bimaculatus. Copulatory acts of the male, stereotyped and time-fixed, are elicited by stimulation of mechanoreceptors on particular regions of the abdomen, cerci and genitalia. Sequential execution of each motor act proceeds as a chain reaction in which one act stimulates some receptors which in turn elicits another act and so on, while the female remains immobile on the male's back. Each key stimulus for a motor act appears as a result of the male's own act, except for copulatory papilla protrusion by the female. The final sequence of spermatophore extrusion and transfer are irreversible fixed motor actions which are triggered when the female copulatory papilla stimulates the epiphallic hairs. They proceed without continual central drive from the brain, and apparently without sensory feedback. In addition, they are well coordinated with movement and posture in the entire body. Some neural mechanisms of controlling mating behavior and switching the reproductive cycle are discussed. PMID- 15277714 TI - Not Just an empty cavity: the inter-rhabdomeral space in the Jamaican cavefly Neoditomyia farri (Diptera, Mycetophilidae). AB - Contrary to most other Diptera, the inter-rhabdomeral spaces of the retina of the Jamaican cavefly Neoditomya farri are filled neither by extracellular matrix nor dense cytoplasmic material. Instead, a foamy organization of loose vacuoles, measuring approximately 0.7 microm in diameter, appears to keep the rhabdomeres of retinula cells 7 and 8 in place. The vacuoles are bounded by membranes and traces of actin, determined immunocytochemically, are present. The origin of the vacuoles is unclear, but evidence in support of a retinula cell rather than cone cell origin is advanced. PMID- 15277715 TI - A non-social and isolate rearing condition induces an irreversible shift toward continued fights in the male fighting fish (Betta splendens). AB - Effects of rearing conditions were examined in the development of agonistic behaviors in the male fighting fish. In group-I (highly social), fish were communally reared. In group-II (highly social and isolate), fish were individually housed and exposed to the group-I fish through transparent walls until the sexual maturity (from 6 to 12 weeks post-hatch). In group-III (social and isolate), individually housed fish were similarly exposed to other fish within the group. In group-IV (non-social and isolate), individually housed fish were further visually isolated. Agonisitc behaviors were compared among males of the groups-II, -III, and -IV in their fights against the group-I male. The group IV males showed significantly higher rate of wins than the groups-II and -III males, without differences in the incidence of agonistic behaviors (butt-or-bite, chase, and gill-cover erect) before the termination of the mutual fights. Increased incidence of agonistic behaviors was found after the termination (particularly in the unilateral chase), suggesting that the group-IV males continued to fight even after the opponent male displayed a submission. The aggression was also enhanced in the group-II, when they were thereafter reared in a social isolation after the sexual maturation; a critical period was thus not found. The enhanced aggression was not reversed in the group-IV, when they were thereafter exposed to social stimuli; shift to the continued fights was irreversible. Possible fitness gain of the enhanced aggression was discussed in terms of the adjustability to altered biological resources. PMID- 15277716 TI - Photosensitivity of the central nervous system of Limulus polyphemus. AB - The photosensitivity of the central nervous system (CNS) of the horseshoe crab, Limulus polyphemus, was investigated by analyzing changes in motor nerve activity in the segmental nerves of prosomal and opisthosomal ganglia. Spontaneous efferent impulses were recorded in the dark from all the investigated segmental nerves. Impulse trains from the 7th dorsal nerve in the prosomal CNS were inhibited in response to illumination of the whole CNS. Impulse trains from each of the 9-13th dorsal nerves in the isolated opisthosomal CNS were inhibited, and the impulse train from each the 14-16th dorsal nerve was elicited or inhibited upon illuminating the whole CNS. Spontaneous rhythmic bursts at 20-80 s intervals were recorded in the dark from the ventral nerves of the isolated opisthosomal CNS. In the presence of light, the rhythmicity of spontaneous bursts disappeared and other species of impulse trains were elicited. In single ganglion preparations, isolated from the rest of the CNS by surgically severing the connectives, similar photoresponses were recorded before and after isolation. These results demonstrate that the CNS of Limulus is a photosensitive organ. PMID- 15277717 TI - Intraovarian cavity leucocytes of viviparous fish, Neoditrema ransonneti (Perciformes, Embiotocidae). AB - To obtain basic information on the properties of the intraovarian cavity leucocytes (IOCLs) of the viviparous teleost, Neoditrema ransonneti, morphological characteristics and numerical changes of IOCLs during the reproductive cycle were investigated. In the ovaries of newborn females, leucocytes exuded into the lumen were observed first in November, prior to insemination of semen. These cells were primarily macrophages, neutrophils and lymphocytes. Among them, macrophages were invariably the largest population throughout the reproductive cycle. They began to phagocytize spermatozoa in December, when spermatozoa were first detected in the ovary. The number of IOCLs gradually increased from November in the newborn female. However, this increase is not ascribed to the effect of copulation or the presence of semen, because the number of leucocytes also increased in non-mating fish. While developing embryos were discharged into the ovarian lumen at the latest in January, a number of spermatozoa and spermatozoa-phagocytizing macrophages were seen until March. Even after the extinction of sperm cells, numerous IOCLs remained in the lumen and coexisted with fetuses until their parturition. These results suggest that IOCLs play roles in successful pregnancy, besides elimination of remaining spermatozoa. PMID- 15277718 TI - Embryonic stages from cleavage to gastrula in the loach Misgurnus anguillicaudatus. AB - Early developmental staging from the zygote stage to the gastrula is a basic step for studying embryonic development and biotechnology. We described the early embryonic development of the loach, Misgurnus anguillicaudatus, based on morphological features and gene expression. Synchronous cleavage was repeated for 9 cycles about every 27 min at 20 degrees C after the first cleavage. After the 10th synchronous cleavage, asynchronous cleavage was observed 5.5 h post fertilization (hpf), indicating the mid-blastula transition. The yolk syncytial layer (YSL) was formed at this time. Expressions of goosecoid and no tail were detected by whole-mount in situ hybridization from 6 hpf. This time corresponded to the late-blastula period. Thereafter, epiboly started and a blastoderm covered over the yolk cell at 8 hpf. At 10 hpf, the germ ring and the embryonic shield were formed, indicating the stage of early gastrula. Afterward, the epiboly advanced at the rate of 10% of the yolk cell each hour. The blastoderm covered the yolk cell completely at 15 hpf. The embryonic development of the loach resembled that of the zebrafish in terms of morphological change and gene expression. Therefore, it is possible that knowledge of the developmental stages of the zebrafish might be applicable to the loach. PMID- 15277719 TI - Localization of ghrelin-producing cells in the stomach of the rainbow trout (Oncorhynchus mykiss). AB - Ghrelin, an endogenous ligand for the growth hormone secretagogue receptor (GHS R), was isolated from the rat stomach and determined to be n-octanoylated 28 amino-acid peptide. In this study, we studied the distribution of ghrelin producing cells (ghrelin cells) in the gastrointestinal tract of male and female rainbow trout (Oncorhynchus mykiss) by immunohistochemistry using N-terminal region-recognizing antibody and also by in situ hybridization using a trout ghrelin-specific cRNA probe. Ghrelin cells were found in the mucosal layer of the stomach but not in the myenteric plexus, and no ghrelin cells were observed in other regions of the gastrointestinal tract. Ghrelin cells could be classified into two types: closed- and opened-type cells. The density of ghrelin cells increased gradually in the direction from the cardiac to pyloric portions of the stomach in both sexes. The number of ghrelin cells per unit area seemed to be higher in females than in males. In conclusion, trout ghrelin cells exist in the stomach and are classified into two types of cells, closed- and opened-type cells. PMID- 15277720 TI - Human type II GnRH receptor mediates effects of GnRH on cell proliferation. AB - GnRH (gonadotropin-releasing hormone) is well-known as the central regulator of the reproductive system through its stimulation of gonadotropin release from the pituitary. Progress in studies on GnRH demonstrated that GnRH has both inhibitory and stimulatory effects on cell proliferation depending on the cell type, and the mechanisms of these effects have been intensively studied. However, even human GnRH receptors which mediate GnRH stimulation have not been completely identified. In the present study, we showed that the inhibitory and stimulatory effects of GnRH on colony-formation using four cell lines and have demonstrated that the inhibitory and stimulatory effects of GnRH exhibit distinctly different patterns of ligand sensitivity. This result strongly suggests that the two opposite effects of GnRH occur via different types of GnRH receptors, however expressional analyses of human GnRH receptors did not exhibit the significantly different pattern between negatively and positively responding cell lines. Then, in order to identify the GnRH receptors involved in the two opposite effects, effects of GnRH were analysed under the conditions that human GnRH receptors were knocked down by the technique of RNA interference. Consequently, it was found that human type II GnRH receptor mediates GnRH stimulation and its splice variant determines the direction of the response to GnRH. These results are the first clear evidence for the functionality of human type II GnRH receptor. Therefore our novel findings are quite noticeable and will greatly contribute to the studies on the mechanisms of the effects of GnRH on cell proliferation in the future. PMID- 15277721 TI - Distribution of albatross remains in the Far East regions during the Holocene, based on zooarchaeological remains. AB - Many albatross remains have been found in the Japanese Islands and the surrounding areas, such as Sakhalin and South Korea. These remains are interesting for two reasons: numerous sites from which albatross remains have been found are located in coastal regions of the Far East where no albatrosses have been distributed recently, and there are some sites in which albatross remains represent a large portion of avian remains, although albatrosses are not easily preyed upon by human beings. We collected data on albatross remains from archaeological sites in the Far East regions during the Holocene and arranged the remains geographically, temporally and in terms of quantity. Based on these results, we showed that coastal areas along the Seas of Okhotsk and Japan have rarely been used by albatrosses in Modern times, though formerly there were many albatrosses. We proposed two explanations for the shrinkage of their distributional range: excessive hunting in the breeding areas, and distributional changes of prey for albatrosses. PMID- 15277722 TI - Close relationship between Asterina and Solasteridae (Asteroidea) supported by both nuclear and mitochondrial gene molecular phylogenies. AB - Phylogenetic relationships among asteroids remain to be extremely controversial in spite of many morphological and molecular studies have been applied to this issue. In the present study, especially focusing on resolving the relationship of Asterina and Solasteridae, we reconstructed the molecular phylogenetic tree of asteroids using nuclear 18S rDNA. A close relationship between Asterina and Solasteridae, which has been suggested from analyses of mitochondrial 12S rDNA and 16S rDNA, is supported here by the nuclear 18S rDNA dataset. The support is even stronger when the sequences of mitochondrial rDNAs and nuclear 18S rDNA are combined as a total dataset. The independent support from both nuclear 18S rDNA and mitochondrial rDNAs strongly argues for a close relationship between the Asterina and Solasteridae. PMID- 15277723 TI - Mitochondrial gene introgression between spined loaches via hybridogenesis. AB - This report deals with an unusual mode of mitochondrial gene introgression between Cobitis hankugensis (C. sinensis) and C. longicorpus which is mediated by a unisexual hybridogenetic system of diploid-triploid C. hankugensis-longicorpus complex. Mitochondrial DNA sequences of 3329-3330bp encompassing from upstream ND6 to 12S rDNA indicated that mitochondrial genomes from the diploid hybrids, triploid hybrids, and their parental species are almost identical. Because triploid hybrids produce haploid ova with C. hankugensis chromosome set, normal diploid C. hankugensis regenerates upon insemination with C. hankugensis sperm. If the hybrid carries C. longicorpus mitochondrial genome, the regenerated C. hankugensis is a nucleo-cytoplasmic hybrid, thus accomplishing the unusual mode of mitochondrial gene introgression. PMID- 15277724 TI - Pharmacological role of HB-EGF shedding by angiotensin II in cardiomyocytes. AB - Angiotensin II (AGII) is known to induce cardiac hypertrophy. Prolonged hypertrophy leads to reduced cardiac performance. The cardiac protective effect of AGII antagonist implies the potential role of AGII in the transition from cardiac hypertrophy to heart failure. Here we report a new mechanism of HB-EGF mediated cardiac hypertrophy induced by AGII. HB-EGF is a EGF family growth factor synthesized as the membrane anchored form and released by protease cleavage to activate its receptor. In cultured cardiomyocytes, AGII induced the transactivation of EGF receptor, which was blocked by metalloproteinase inhibitor KBR-7785 and HB-EGF-neutralizing antibody #19. Both KBR-7785 and #19 attenuated the cardiac hypertrophy by AGII in vitro and in vivo. Thus we conclude that AGII activates metalloproteinase and sheds HB-EGF. Released HB-EGF bound to EGF receptor, leading to the cardiac hypertrophy. Recently, similar transactivation of EGF receptor by a GPCR agonist has been reported in various organs, indicating that EGF receptor transactivation by HB-EGF might play the general role of pharmacological reaction by AGII. PMID- 15277725 TI - Chymase-derived angiotensin II and arrhythmias after myocardial infarction. AB - Mast cell-derived chymase seems to be important in the regulation of local angiotensin (A) II formation in cardiovascular tissues. In human heart, chymase accounts for 80% of A II formation. Therefore, the chymase-dependent A II pathway may play an important role in the pathogenesis of A II-related cardiovascular diseases. For example, cardiac chymase was activated earlier than ACE and this activation lasted longer than that of ACE after myocardial infarction (MI) in hamsters. Treatment with a specific chymase inhibitor treatment, but not an ACE inhibitor, improved post-MI survival as well as cardiac function and the extent of the beneficial effects was similar to that observed for an AT1-receptor antagonist treatment in this model. The survival benefit after MI seems to be related to an antiarrhythmic effect of the chymase inhibitor because chymase inhibition reduces the incidence of ventricular arrhythmias during periods of heart ischemia in a dog MI model. Thus, an antiarrhythmic effect of the chymase inhibitor may contribute to a reduction in mortality rate during the acute phase after MI. PMID- 15277726 TI - Additive improvement of left ventricular remodeling by aldosterone receptor blockade with eplerenone and angiotensin II type 1 receptor antagonist in rats with myocardial infarction. AB - We investigated the effects of the aldosterone blocker eplerenone alone and in combination with angiotensin II type 1 receptor antagonist on ventricular remodeling in rats with left ventricular (LV) dysfunction after extensive myocardial infarction (MI). Adding an aldosterone antagonist to an ACE inhibitor reduces mortality and morbidity in heart failure. Starting 1 day after MI, rats were treated with placebo, eplerenone (100 mg/kg/day), the angiotensin type 1 receptor antagonist candesartan (1 mg/kg/day), or a combination of both for nine weeks. Both monotherapies attenuated the rise in LV end-diastolic dimension (LVDd) and LV end-diastolic volume (LVEDV) compared with placebo, whereas combined treatment further attenuated LVDd and LVEDV and significantly improved LV function. Increased collagen type I and III gene expressions in the noninfarcted LV myocardium from MI placebo rats was attenuated by candesartan, but almost completely prevented by eplerenone and eplerenone/candesartan. The addition of eplerenone to candesartan prevented the increases in LV gene expression of ANP and BNP more effectively than either monotherapy. The aldosterone blocker eplerenone improved LV remodeling in rats with LV dysfunction after extensive MI. Combination therapy with an candesartan substantially potentiates this effect by a complementary prevention of LV fibrosis, cardiac hypertrophy, and molecular alterations. PMID- 15277727 TI - Mineralocorticoid receptor antagonist spironolactone improves left ventricular remodeling in patients with congestive heart failure and acute myocardial infarction. AB - Randomized aldactone evolution study (RALES) and eplerenone post-AMI heart failure efficacy and survival study (EPHESUS) had shown that aldosterone blockade (AB) with standard therapy resulted in a reduction in mortality in patients with congestive heart failure (CHF) and acute myocardial infarction (AMI). However, the mechanism for the beneficial effect of AB remains unknown. To evaluate the effect of spironolactone (Spi) on left ventricular (LV) remodeling, 34 CHF patients with DCM were randomly divided into the Spi (+) or Spi (-) groups. Four months of treatment with Spi improved the LV volume and mass. To evaluate the effect of Spi on post-infarct LV remodeling, 134 patients with first anterior AMI were randomly divided into the Spi (+) or Spi (-) groups after revascularization. LV ejection fraction was significantly improved after 1 month in the Spi (+) group compared with that in the Spi (-) group. LV end-diastolic volume index was significantly suppressed in the Spi (+) group compared with that in the Spi (-) group. Transcardiac extraction of aldosterone through the heart was significantly suppressed in the Spi (+) group and was significant lower in the Spi (+) group compared with the Spi (-) group. These findings indicate that AB combined with standard therapy can prevent LV remodeling in patients with CHF and AMI, suggesting that the failing heart is the target organ of the aldosterone. PMID- 15277728 TI - Aldosterone and renal injury. AB - Our recent efforts have been focused on the mechanisms responsible for the progression of aldosterone-induced renal injury. We have demonstrated in rats that chronic treatment with aldosterone (0.75 micro g/H, SC) and 1% NaCl (in drinking solution) results in severe proteinuria and glomerular injury, characterized by cell proliferation and mesangial matrix expansion. Increased renal cortical NAD(P)H oxidase expression, reactive oxygen species (ROS) generation, and mitogen-activated protein kinase (MAPK) activation were also observed. Treatment with a selective mineralocorticoid receptor antagonist, eplerenone(0.125% in chow), or an antioxidant, tempol (3 mM in drinking solution), prevented elevations of ROS levels and MAPK activity, as well as ameliorating glomerular injury, indicating that aldosterone-induced glomerular injury is associated with redox-sensitive MAPK activation. In vitro studies showed that mineralocorticoid receptors are highly expressed in rats mesangial cells, particularly in the cytoplasm. Aldosterone (100 nM) application activates MAPK and causes cellular proliferation and deformation. These data suggest that aldosterone contributes to the progression of glomerular injury through its direct actions. PMID- 15277729 TI - Application of in vivo patch-clamp technique to pharmacological analysis of synaptic transmission in the CNS. AB - Slice preparations as well as acutely dissociated and cultured neurons from various regions in the CNS have been widely used to analyze pharmacological properties of synaptic responses and receptors expressed at the pre- and post synaptic sites. However, the essential properties are not obviously different from neuron to neuron. These characteristics of neurons in the CNS make it difficult to elucidate their functional significances. It is, therefore, preferable that the pharmacological analysis should be made from identified neurons by stimulation of identified inputs. The in vivo patch-clamp recording technique allows us to clarify the synaptic responses evoked by the various known natural stimuli applied to the skin or other parts and makes it possible to interpret with more certainty the behavioral changes by synaptic plasticity observed at the single cell level. Although the in vivo technique has obvious advantages in analysis of physiological responses, this method is, however, confined so far to neurons located at the near surface of the CNS for pharmacological analysis, because of the diffusion problem of the chemical to deeper neurons. Thus, combinatorial studies with dissociated or cultured neurons or with slice preparations are clearly required for further understanding of pharmacological properties of neurons in the CNS. PMID- 15277730 TI - Long-term prognosis of patients with paroxysmal atrial fibrillation depends on their response to antiarrhythmic therapy. AB - BACKGROUND: The rhythm control treatment strategy for persistent atrial fibrillation (AF) has been shown not to improve quality of life or prognosis any more than rate control. It is unclear whether the prognosis of the patients with paroxysmal AF (PAF) is influenced by the response to antiarrhythmic drug therapy (AAT). METHODS AND RESULTS: The relationship between the response to AAT and long term prognosis was evaluated in 290 patients with PAF (mean age, 69 years). During a mean follow-up period of 51 months, 114 patients (39%) had no recurrence of AF (Group 1), 113 (39%) had repeated AF recurrence (Group 2), and the remaining 63 (22%) had permanent AF despite AAT (Group 3). The survival rate without any cardiovascular deaths at 60 months was 99% in Group 1, 95% in Group 2 and 94% in Group 3 (p=NS among 3 groups). Survival rate without symptomatic ischemic stroke was 99% in Group 1, 88% in Group 2 and 76% in Group 3 (p<0.05 Group 1 vs Groups 2 and 3). The annual rate of stroke in the patients with warfarin treatment was similar among the 3 groups, whereas that in the patients without warfarin was higher in Groups 2 and 3 than in Group 1. CONCLUSIONS: Long term prognosis of patients with PAF varies with the response to AAT: When sinus rhythm is maintained, the prognosis is good even without anticoagulation therapy. PMID- 15277731 TI - Risk factors for patients developing a fulminant course with acute myocarditis. AB - BACKGROUND: A fulminant course can be difficult to predict at the onset of acute myocarditis, so the aim of the present study was to identify the predictive clinical symptoms/signs or laboratory findings. METHODS AND RESULTS: Thirty-nine patients with acute lymphocytic myocarditis, excluding 8 who manifested shock at admission, were studied. The fulminant group was defined as 12 patients who developed shock after admission, requiring intraaortic balloon pumping or percutaneous cardiopulmonary support, and the non-fulminant group comprised the 27 patients without shock. Various parameters at admission were compared between the 2 groups, together with multiple logistic regression analysis, excluding 6 patients with partially missing values. In the fulminant group, C-reactive protein (7.0 +/- 7.0 vs 2.3 +/- 2.2 mg/dl, p<0.01) and creatine kinase (1,147 +/- 876 vs 594 +/- 568 IU/L, p<0.05) concentrations were higher, intraventricular conduction disturbances were more frequent (9/12 vs 7/27 patients, p<0.01) and the left ventricular ejection fraction was lower (40.7 +/- 13.9 vs 50.1 +/- 10.6%, p<0.05) than in the non-fulminant group. In the multiple logistic regression analysis model with the presence/absence of a fulminant course considered as the independent variable, and C-reactive protein, creatine kinase, intraventricular conduction disturbances, and left ventricular ejection fraction as dependent variables, a high-risk group (expected proportion of fulminant course > or = 0.5) and a low-risk group (<0.5) could be differentiated. A fulminant course occurred in 9/13 (69%) patients in the high-risk group, but in only 2/20 (10%) patients in the low risk group (p<0.001). CONCLUSIONS: The risk of a fulminant course of acute myocarditis was high in patients with elevated C reactive protein, and creatine kinase concentrations, decreased left ventricular ejection fraction, and intraventricular conduction disturbances at the time of admission. PMID- 15277732 TI - Genetic analysis of Brugada syndrome in Western Japan: two novel mutations. AB - BACKGROUND: Brugada syndrome is a form of idiopathic ventricular fibrillation characterized by right bundle-branch block pattern and ST elevation in the right precordial leads of the ECG. The SCN5A gene encodes the alpha-subunit of the human heart sodium channel, which plays a critical role in cardiac excitability, and mutations of SCN5A could underlie Brugada syndrome. METHODS AND RESULTS: To detect mutations of SCN5A, DNA samples from 12 Japanese patients with Brugada syndrome were analyzed using direct sequencing. Two patients had novel mutations, G292S and S835L, but no other mutations of SCN5A were detected in the remaining patients. The first mutation, G292S, was identified adjacent to the pore-lining region between the DIS5 and DIS6 transmembrane segments of SCN5A, and the second mutation, S835L, was in the intracellular loop connecting the DIIS4 to DIIS5. Both mutations were not detected in 100 unrelated control subjects. CONCLUSION: Two novel SCN5A mutations have been found in Japanese patients with Brugada syndrome. PMID- 15277733 TI - Ongoing myocardial damage in chronic heart failure is related to activated tumor necrosis factor and Fas/Fas ligand system. AB - BACKGROUND: Elevated concentrations of cardiac troponin T and heart-type fatty acid-binding protein (H-FABP) identify patients with chronic heart failure (CHF) and ongoing myocardial damage (OMD) who are at increased risk for future cardiac events. Cardiomyocyte necrosis and/or apoptosis via activated tumor necrosis factor (TNF) and the Fas/Fas ligand (FasL) system may be related to the development of OMD. METHODS AND RESULTS: The serum concentrations of H-FABP, a sensitive marker of membrane damage of cardiomyocytes, soluble Fas (sFas) and TNF alpha were measured in 38 patients with CHF. The concentrations of H-FABP, TNF alpha and s-Fas in patients with New York Heart Association (NYHA) III + IV were all significantly higher than in those patients in NYHA II (H-FABP; III + IV 9.3+/-5.9 vs II 5.1+/-1.8 ng/ml, p=0.003, TNF-alpha; III + IV 10.5+/-3.8 vs II 8.0+/-2.7 pg/ml, p=0.02, sFas; III + IV 3.36+/-1.37 vs II 2.58 +/-0.84 ng/ml, p=0.03). Increased concentrations of H-FABP significantly correlated with the concentrations of TNF- alpha (r=0.57, p=0.0001) and sFas (r=0.69, p<0.0001), independent of renal function. CONCLUSION: OMD detected by H-FABP, a marker of membrane damage, is related to activated TNF and the Fas/FasL system, which suggests a pathophysiological role of cardiomyocyte necrosis and/or apoptosis in patients with worsening heart failure. PMID- 15277734 TI - New holter monitoring analysis system: a calculation of the lead vectors. AB - BACKGROUND: A new system of synthesizing a 12-lead electrocardiogram (Syn-ECG) with practically identical waveforms to the standard 12-lead ECG (Stn-ECG) from 3 channel ECGs recorded by Holter monitoring has been developed. METHODS AND RESULTS: The study group comprised 16 healthy individuals and 13 patients with abnormal ECGs. The bipolar eV1, eV5 and eVF leads were recorded using digital Holter monitoring and nine Syn-ECGs, corresponding to each lead of the Stn-ECG, were synthesized. The 9 ECGs consisted of a theoretical Syn-ECG and 8 Syn-ECGs positioned around the theoretical Syn-ECG at 3 cm intervals on the Frank's image surface. Of the 9 ECGs, the Syn-ECG with the maximum product of the cross correlation coefficient of the QRS wave and that of the T wave, was automatically selected as the optimal Syn-ECG. The amplitude data from the QRS wave, R wave, T wave, and ST level, and also the amplitude ratio of the R wave, T wave to the QRS wave, were significantly well correlated between the Syn-ECG and Stn-ECG. CONCLUSIONS: A practically identical ECG morphology, comparable with a Stn-ECG, was successfully created using this system. PMID- 15277735 TI - Heart rate variability and ambulatory blood pressure monitoring in young patients with hypertrophic cardiomyopathy. AB - BACKGROUND: Sudden cardiac death commonly occurs in young patients with hypertrophic cardiomyopathy (HCM); however, their heart rate variability (HRV) and blood pressure (BP) response to daily life activities is not well known. METHODS AND RESULTS: HRV and ambulatory BP monitoring were performed in 20 patients (age range: 7-21 years) and 57 age-matched healthy volunteers (age range: 10-22 years). Time domain variables and spectral data were obtained at hourly intervals throughout the day. To determine the BP response to daily life activities, the ratios of the mean BP and pulse pressure in the morning, afternoon, and night to those during sleeping were calculated. The association between the BP level and HRV was also evaluated. The HCM patients showed significantly increased sympathovagal imbalance and decreased parasympathetic activity in the early morning, around noon, and in the early evening. This abnormality was independent of cardiac symptoms. Symptomatic patients showed a significantly lower systolic BP response in the morning, and a higher incidence of dissociation between sympathetic activity and BP response than asymptomatic patients. CONCLUSION: An abnormal BP response in the presence of impaired HRV appears to be predictive for cardiac events in young patients with HCM. PMID- 15277736 TI - Effects of a distal protection device during primary stenting in patients with acute anterior myocardial infarction. AB - BACKGROUND: The angiographic no-reflow phenomenon is an adverse prognostic factor in patients with acute myocardial infarction (AMI). The aim of the present study was to evaluate the effects of an occlusive balloon type distal protection device (PercuSurge GuardWire: GW) during primary stenting in patients with anterior AMI. METHODS AND RESULTS: The GW group included 42 patients treated by primary stenting with GW protection and the control group included 30 patients treated by primary stenting after thrombectomy without distal protection. Left ventricular (LV) function was measured and compared by left ventriculography obtained soon after percutaneous coronary intervention (PCI) and 3 weeks after onset. The corrected TIMI frame count values were lower in the GW group than in the control group (27.5+/-2.3 vs 35.1 +/-2.5, p=0.030). The number of patients with myocardial blush grade 3 after PCI was higher in the GW group than in the control group (45.7 vs 20.0%, p=0.029). Peak concentration of creatine kinase myocardial fraction was lower in the GW group than in the control group (326.6+/-41.5 vs 454.9+/-46.2 mg/dl, p=0.043). GW patients showed greater improvement at 3 weeks after PCI in terms of LV ejection fraction (+4.6+/-1.2 vs -1.1+/-1.5, p=0.004), LV end-systolic volume index (+0.5+/-2.4 vs +9.0+/-2.7, p=0.023), and regional wall motion abnormalities (-2.03+/-0.14 vs -2.51+/-0.14, p=0.018). CONCLUSION: Primary stenting with GW protection can restore epicardial coronary flow and myocardial perfusion, and also preserve LV function in anterior AMI. PMID- 15277737 TI - New method of measuring coronary diameter by electron-beam computed tomographic angiography using adjusted thresholds determined by calibration with aortic opacity. AB - BACKGROUND: In a previous study the adjusted thresholds at which the diameters of coronary arteries determined by enhanced electron-beam computed tomography (CT) scans are equal to the corresponding quantitative coronary angiography measurements were analyzed, and their correlation with maximum CT values for the vessel short axes was determined. A rapid accurate method for such measurements was sought by substituting maximum CT values for the descending aorta in the corresponding axial images for those for the short axes. METHODS AND RESULTS: In 8 patients, 179 sites were measured. Means (+/- SD) of adjusted thresholds and the maximum CT values for vessel short axes and the descending aorta in the corresponding axial images for all vessels were 108 +/-66, 227+/-80, and 363+/-75 Hounsfield Unit (HU), respectively. Adjusted thresholds correlated with the maximum CT values for the corresponding vessel short axes and the descending aorta in the corresponding axial images, with R2=0.55, 0.33, p<0.01, respectively. An abbreviated formula for use of maximum CT values for the descending aorta in the corresponding axial images was y=0.5x-75 (HU) (y= adjusted threshold, x= maximum CT value for the descending aorta in the corresponding axial image). CONCLUSIONS: The abbreviated formula provided a rapid, accurate method for measurements independent of arterial enhancement. PMID- 15277738 TI - Early aerobic training increases end-tidal CO2 pressure during exercise in patients after acute myocardial infarction. AB - BACKGROUND: End-tidal CO2 partial pressure (PETCO2) has been suggested as a noninvasive index reflecting cardiac output under constant ventilation. The aim of this study was to examine whether PETCO2 does reflect cardiac output, even during exercise, in patients with acute myocardial infarction (AMI) undergoing exercise training early after onset. Method and Results Patients aged 47-73 years were randomly assigned to either a training group (n=18) or a control group (n=18) 1 week after the onset of AMI. Those in the training group performed exercise training under supervision at the anaerobic threshold level for 2 weeks, while patients in the control group followed a conventional walking regimen. In the training group, but not in the control group, PETCO2 at the respiratory compensation point increased significantly from 39.1+/-3.5 to 41.1+/-3.7 mmHg (p<0.01). Similarly, the cardiac index at peak exercise increased only in the training group (from 6.04+/-0.98 to 7.31+/-0.97 L/min per m2, p<0.01). These 2 measurements correlated well both before and after the study period. Peak oxygen uptake and anaerobic threshold were increased only in the training group. Conclusions Aerobic exercise training early after the onset of AMI significantly increased PETCO2 during exercise, which may reflect an improvement in cardiac output during exercise in response to physical training via a decreased ventilation-perfusion mismatch. PMID- 15277739 TI - Different effects of pravastatin and cerivastatin on the media of the carotid arteries as assessed by integrated backscatter ultrasound. AB - BACKGROUND: Currently, there are various types of statins used in the treatment of hyperlipidemia and coronary artery disease. The purpose of this study was to compare the effects of a lipophilic statin (cerivastatin) with those of a hydrophilic statin (pravastatin) on the carotid arterial media using integrated backscatter (IB) ultrasound. Cerivastatin (C) has a strong anti-proliferative effect (APE) on smooth muscle cells (SMCs), whereas pravastatin (P) has a weak effect. METHODS AND RESULTS: The IB values in the media of 72 segments of carotid arteries were measured in 36 patients with hyperlipidemia before and after statin therapy or diet for 6 months (C, n=13: P, n=12: diet, n=11). In addition, IB values of 34 segments of carotid arteries were measured in 34 patients without coronary risk factors. Intima - media thickness (IMT) and arterial stiffness (stiffness beta) were measured by conventional echo at the same time. IB values did not significantly change in the P group (12.8+/-3.5 vs 12.7+/-2.7 dB), but decreased in the C group (12.1 +/-2.9 vs 10.0+/-2.7 dB, p<0.01). Also, stiffness beta did not significantly change in the P group (8.3+/-3.1 vs 7.6+/-2.5), but decreased in the C group (10.1+/-4.3 vs 7.9+/-3.3, p<0.05). IB values correlated with age (r=0.70, p<0.01) and stiffness beta (r=0.67, p<0.01) in the 34 patients without coronary risk factors. CONCLUSIONS: Statin therapy with cerivastatin, but not pravastatin, decreased the IB values of the carotid media and arterial stiffness. The difference between these 2 statins may be related to their effective dose range. PMID- 15277740 TI - Reconstruction of bilateral branch pulmonary artery stenosis caused by Takayasu's aortitis. AB - A 63 year-old female presented with dyspnea on exertion. Her chest X-ray showed cardiomegaly, and right ventricular overload and tricuspid regurgitation were detected. Her pulmonary ventilation and blood flow scintigraphy findings were suspicious of pulmonary vascular disease; the diagnosis was pulmonary hypertension and bilateral branch pulmonary artery stenosis. After the inflammation settled, the stenotic bilateral branch pulmonary artery was reconstructed with a prosthetic vessel and the pulmonary pressure normalized immediately. A resected specimen revealed that the stenotic changes were from Takayasu's disease. The patient's postoperative course was uneventful, and pulmonary ventilation and blood scintigraphy returned to an almost normal range. At follow-up 5 years and 6 months after the operation, there was no evidence of pulmonary artery disease (eg, stenosis and/or ischemia) or of any change in the central vessels of the retina, the so-called Takayasu's retinopathy. PMID- 15277741 TI - Giant organized thrombus in the left sinus of valsalva causing intermittent left coronary obstruction: an unusual case of acute myocardial infarction. AB - A 48-year-old Japanese man was admitted to hospital for acute myocardial infarction associated with a giant organized thrombus occupying the left sinus of Valsalva. Cardiac catheterization revealed no organic stenosis in either coronary artery, but left ventriculography and aortography showed a filling defect above the left coronary cusp. Transesophageal echocardiography was immediately performed and showed a round mass filling the left sinus of Valsalva. A solid, round mass approximately 2.5 cm in diameter was removed during emergency surgery and determined to be a thrombus on the basis of microscopic findings. This is the second report of a giant organized thrombus occupying the entire left sinus of Valsalva, obstructing the ostium of the left coronary artery intermittently, and leading to acute myocardial infarction. PMID- 15277742 TI - Injury to the tricuspid valve and membranous atrioventricular septum caused by huge calcified right ventricular myxoma: report of a case. AB - A 64-year-old female, admitted because of severe dyspnea on exertion and facial edema, showed echocardiographic findings of a large tumor in the right ventricle (RV). Echocardiography revealed a cardiac mass extending from the RV across the tricuspid valve into the right atrium, synchronized with the cardiac cycle, and severe tricuspid regurgitation was apparent. The mass was removed under cardiopulmonary bypass. It measured 7 x 5 x 5 cm with diffuse superficial calcification and arose from the posterior wall of the RV, just under the tricuspid valve ring, with a short pedicle. During the same procedure, after the successful excision of the tumor, small atrial and ventricular septal defects were found that had been caused by the tumor and these were closed directly. The tricuspid valve was repaired with valvuloplasty, chordoplasty and annuloplasty. The microscopic findings were of typical myxoma; however, a right ventricular myxoma protruding into the right atrium is exceedingly rare. PMID- 15277743 TI - Failure of follow-up gallium single-photon emission computed tomography and fluorine-18-fluorodeoxyglucose positron emission tomography to predict the deterioration of a patient with cardiac sarcoidosis. AB - Although gallium-67-citrate (67Ga) scanning and single-photon emission computed tomography (SPECT) are useful in the assessment of disease activity in cardiac sarcoidosis, a patient with cardiac sarcoidosis in whom SPECT imaging with 67Ga failed to predict the deterioration in the clinical course is presented. A 53 year-old woman diagnosed with cardiac sarcidosis had 67Ga scanning and 67Ga SPECT, both of which showed abnormal high uptake. After treatment with corticosteroid, there was an apparent improvement in the 67Ga SPECT findings, and the dose of the corticosteroid was reduced. Subsequently, the disease activity of the cardiac sarcoidosis was thought to be well controlled, because abnormal uptake was not found on repeat 67Ga SPECT. However, 4 years after initial diagnosis, thinning at the basal ventricular septal wall and complete atrioventricular block were noted. Despite repeating the evaluation with 67Ga SPECT and additional fluorine-18-fluorodeoxyglucose positron emission tomography (18FDG PET) after discovering this progression, neither of these examinations showed any abnormality. Unfortunately, in this patient, the disease activity of cardiac sarcoidosis was underestimated by the diagnostic imaging modalities. PMID- 15277744 TI - Purification and characterization of a 24 kDa protein from tartary buckwheat seeds. AB - A 24 kDa protein was isolated from tartary buckwheat seeds by using chromatography of Superdex 75 gel filtration and Resource Q ion-exchange column. SDS-PAGE and Sephacryl S-200 gel filtration were used to provide information about the molecular mass of the protein purified from tartary buckwheat. The protein was composed of 215 amino acid residues and showed strong IgE binding activity in an ELISA test to the sera colleted from two patients allergic to buckwheat. These results suggested that the purified 24 kDa protein from tartary buckwheat seeds was an important functional protein and was relatively specific for buckwheat-allergic patients. It should be a very useful tool in the diagnosis of buckwheat allergy in the future. PMID- 15277745 TI - Cloning and characterization of the catabolite control protein A gene from Bacillus stearothermophilus No. 236. AB - The gene encoding the catabolite control protein A (CcpA) of Bacillus stearothermophilus No. 236, a strong xylanolytic bacterium, was cloned, sequenced, and expressed in Escherichia coli. The nucleotide sequence of the ccpA gene corresponded to an open reading frame of 1,005 bp that encodes a polypeptide of 334 amino acid residues with a calculated molecular mass of 36,902 kDa. The CcpA protein belonging to the LacI/GalR family of transcriptional regulators was produced by a recombinant E. coli strain expressing the B. stearothermophilus No. 236 ccpA gene and purified to apparent homogeneity. The transcription start site was mapped at a position 63 nucleotides upstream of the translation initiation codon, and a presumed promoter sequence was also identified. The deduced amino acid sequence of the ccpA gene product contained the helix-turn-helix motif found in many DNA-binding proteins, and showed the highest identity (62%) with CcpA from B. subtilis. The B. stearothermophilus No. 236 ccpA gene was demonstrated to be able to complement a B. subtilis ccpA mutant that exhibited two distinct mutant phenotypes: a growth defect and a release of carbon catabolite repression (CCR). These results indicate that the ccpA gene product of B. stearothermophilus No. 236 is functionally active also in B. subtilis. Electrophoretic mobility shift assay with the purified CcpA revealed that the CcpA of B. stearothermophilus No. 236 bound specifically to the xynA creB (catabolite responsive element B) sequence. Taken together, these results strongly suggest that the CcpA protein participates in CCR of B. stearothermophilus No. 236 xynA gene. PMID- 15277746 TI - Mass spectrometric assignment of Smith degradation glycopeptides derived from ribonuclease B. AB - We established a method to determine the glycosyl linkage structure by a combination of Smith degradation and liquid chromatography-electrospray ionization-quadrupole-time of flight-mass spectrometry (LC-ESI-Q-TOF-MS) and tandem MS (MS/MS). To assign the sugar linkage of N-glycoprotein, we employed a typical ribonuclease B containing oligosaccharides (Man5-9GlcNAc2). Tryptic digestion of ribonuclease B provided a mixture of high-mannose glycopeptides consisting of the four amino acids, Asn34-Leu-Thr-Lys37 (NLTK, T6). The mixture of glycopeptides was separated by high-performance liquid chromatography (HPLC) in a reversed phase column and was characterized by ESI-Q-TOF-MS and MS/MS. Comparison of the data with and without Smith degradation allowed us to make reasonable assignments to support such linkage patterns as (1-->2), (1-->3), (1- >6) and their multiples. These assignments were limited to six mannoses or lower due to the unstable nature of the higher derivatives. This method should be applicable to determine the linkage pattern of an unknown glycoprotein in about a 6-microgram amount. PMID- 15277747 TI - Cloning and characterization of the genes encoding enzymes for the protocatechuate meta-degradation pathway of Pseudomonas ochraceae NGJ1. AB - The 2-pyrone-4,6-dicarboxylate lactonase gene (proL), the protocatechuate 4,5 dioxygenase alpha and beta subunits genes (proOa and proOb), and the 4-carboxy-2 hydroxymuconate-6-semialdehyde dehydrogenase gene (proD) were cloned from the chromosomal DNA of Pseudomonas ochraceae NGJ1. These genes were in the order proLOaObD on the DNA, and a possible transcription terminator sequence followed. The proL and proD genes were over-expressed in Escherichia coli, and their gene products were purified for identification, while the expression of proOaOb was at a lower level. The protocatechuate meta-degradation operon was reconstituted with the recombinant plasmids and expressed successfully in E. coli. PMID- 15277748 TI - Superior molasses assimilation, stress tolerance, and trehalose accumulation of baker's yeast isolated from dried sweet potatoes (hoshi-imo). AB - Yeast strains were isolated from dried sweet potatoes (hoshi-imo), a traditional preserved food in Japan. Dough fermentation ability, freeze tolerance, and growth rates in molasses, which are important characteristics of commercial baker's yeast, were compared between these yeast strains and a commercial yeast derivative that had typical characteristics of commercial strains. Classification tests including pulse-field gel electrophoresis and fermentation/assimilation ability of sugars showed that almost the stains isolated belonged to Saccharomyces cerevisiae. One strain, ONY1, accumulated intracellular trehalose at a higher level than commercial strain T128. Correlated with intracellular trehalose contents, the fermentation ability of high-sugar dough containing ONY1 was higher. ONY1 also showed higher freeze tolerance in both low-sugar and high sugar doughs. The growth rate of ONY1 was significantly higher under batch and fed-batch cultivation conditions using either molasses or synthetic medium than that of strain T128. These results suggest that ONY1 has potential commercial use as baker's yeast for frozen dough and high-sugar dough. PMID- 15277749 TI - Characterization of the L-lysine biosynthetic pathway in the obligate methylotroph Methylophilus methylotrophus. AB - The L-lysine biosynthetic pathway of the gram-negative obligate methylotroph Methylophilus methylotrophus AS1 was examined through characterization of the enzymes aspartokinase (AK), aspartsemialdehyde dehydrogenase, dihydrodipicolinate synthase (DDPS), dihydrodipicolinate reductase, and diaminopimelate decarboxylase. The AK was inhibited by L-threonine and by a combination of L threonine and L-lysine, but not by L-lysine alone, and the activity of DDPS was moderately reduced by L-lysine. In an L-lysine producing mutant (G49), isolated as an S-(2-aminoethyl)-L-cysteine (lysine analog) resistant strain, both AK and DDPS were partially resistant to feedback inhibition. The ask and dapA genes encoding AK and DDPS respectively were isolated from the parental strain, AS1, and its G49 derivative. Comparison of the sequences revealed a point mutation in each of these genes in G49. The mutation in the ask gene altered aspartic acid in a key region involved in the allosteric regulation common to AKs, while a novel mutation in the dapA gene altered tyrosine-106, which was assumed to be involved in the binding of L-lysine to DDPS. PMID- 15277750 TI - Preparation and biological activity of molecular probes to identify and analyze jasmonic acid-binding proteins. AB - Several types of jasomonic acid (JA) derivatives, including JA--amino acid conjugates, a JA--biotin conjugate, a JA--dexamethasone heterodimer, and a JA fluoresceine conjugate, were prepared as candidates for molecular probes to identify JA--binding proteins. These JA derivatives, excepting the JA- fluoresceine conjugate, exhibited significant biological activities in a rice seedling assay, a rice phytoalexin-inducing assay, and/or a soybean phenylalanine ammonia-lyase-inducing assay. These JA derivatives could therefore be useful probes for identifying JA--binding proteins. The activity spectra of the prepared compounds were different from each other, suggesting that different types of JA receptors were involved in the perception of JA derivatives in the respective bioassays. PMID- 15277751 TI - Divergent structures of carbazole degradative car operons isolated from gram negative bacteria. AB - Southern hybridization analysis of the genomes from the newly-isolated 10 carbazole (CAR)-utilizing bacteria revealed that 8 of the isolates carried gene clusters homologous to the CAR-catabolic car operon of Pseudomonas resinovorans strain CA10. Sequencing analysis showed that two car operons and the neighboring regions of Pseudomonas sp. strain K23 are nearly identical to that of strain CA10. In contrast to strains CA10 and K23, carEF genes did not exist downstream of the car gene cluster of Janthinobacterium sp. strain J3. In the car gene clusters, strains CA10, K23 and J3 have Rieske-type ferredoxin as a component of carbazole dioxygenase, although Sphingomonas sp. strain KA1 possesses a putidaredoxin-type ferredoxin. We confirmed that this putidaredoxin-type ferredoxin CarAc can function as an electron mediator to CarAa of strain KA1. In the upstream regions of the carJ3 and carKA1 gene clusters, ORFs whose deduced amino acid sequences showed homology to GntR-family transcriptional regulators were identified. PMID- 15277752 TI - Cloning and overexpression of the Exiguobacterium sp. F42 gene encoding a new short chain dehydrogenase, which catalyzes the stereoselective reduction of ethyl 3-oxo-3-(2-thienyl)propanoate to ethyl (S)-3-hydroxy-3-(2-thienyl)propanoate. AB - Exiguobacterium sp. F42 was screened as a producer of an enzyme catalyzing the NADPH-dependent stereoselective reduction of ethyl 3-oxo-3-(2-thienyl)propanoate (KEES) to ethyl (S)-3-hydroxy-3-(2-thienyl)propanoate ((S)-HEES). (S)-HEES is a key intermediate for the synthesis of (S)-duloxetine, a potent inhibitor of the serotonin and norepinephrine uptake carriers. The responsible enzyme (KEES reductase) was partially purified, and the gene encoding KEES reductase was cloned and sequenced via an inverse PCR approach. Sequence analysis of the gene for KEES reductase revealed that the enzyme was a member of the short chain dehydrogenase/reductase family. The probable NADPH-interacting site and 3 catalytic residues (Ser-Tyr-Lys) were fully conserved. The gene was highly expressed in Escherichia coli, and the gene product was purified to homogeneity from the recombinant E. coli by simpler procedures than from the original host. The molecular mass of the purified enzyme was 27,500 as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and 55,000 as determined by gel filtration chromatography. Our results show that this enzyme can be used for the practical production of (S)-HEES. PMID- 15277753 TI - Genetic analysis of chs1+ and chs2+ encoding chitin synthases from Schizosaccharomyces pombe. AB - To explore the function of chitin in Schizosaccharomyces pombe, we have cloned chs1+ and chs2+, encoding putative chitin synthases, based on sequences in the Sanger Centre database. The synthetic lethal phenotype of the S. cerevisiae chs1 chs2 chs3 mutant was complemented by expression of S. pombe chs1+ or chs1+, indicating that both chs1+ and chs2+ in fact encode chitin synthase. The homothallic Deltachs1 strain formed abnormal asci that contained 1, 2, or 3 spores, while the Deltachs2 strain had no noticeable phenotype. The chs1 chs2 double disruptant looked similar phenotypically to the Deltachs1 strain. The Chs2 GFP fusion protein predominantly localized at the septum after the septum was formed during vegetative growth. The level of chs2+ mRNA increased just before the septum was formed. Levels of Chs2-13Myc synthesis also changed during the cell cycle. Thus, chs1+ is required for proper spore formation, and chs2+ is perhaps involved in septum formation. PMID- 15277754 TI - Absorption of acylated anthocyanins in rats and humans after ingesting an extract of Ipomoea batatas purple sweet potato tuber. AB - We evaluated the absorbability of anthocyanins in humans and rats administered with a beverage prepared from an extract of the tuber of purple sweet potato (Ipomoea batatas Cultivar Ayamurasaki), or with an anthocyanin concentrate. Two major anthocyanin components, cyanidin 3-O-(2-O-(6-O-(E)-caffeoyl-beta-D glucopyranosyl)-beta-D-glucopyranoside)-5-O-beta-D-glucopyranoside) and peonidin 3-O-(2-O-(6-O-(E)-caffeoyl-beta-D-glucopyranosyl)-beta-D-glucopyranoside)-5-O beta-D-glucopyranoside), were detected in the plasma and urine of both rats and humans by HPLC or liquid chromatography/mass spectrometry (LC/MS). The plasma concentration of anthocyanins in humans reached a maximum 90 minutes after ingestion, and the recovery of anthocyanins in the urine was estimated as 0.01 0.03%. These results indicate that acylated anthocyanins could be selectively absorbed after ingesting food. PMID- 15277755 TI - Molecular cloning of a genomic DNA encoding yam class IV chitinase. AB - Genomic DNA for a class IV chitinase was cloned from yam (Dioscorea opposita Thunb) leaves and sequenced. The deduced amino acid sequence shows 50 to 59% identity to class IV chitinases from other plants. The yam chitinase, however, has an additional sequence of 8 amino acids (a C-terminal extension) following the cysteine that was reported as the last amino acid for other class IV chitinases; this extension is perhaps involved in subcellular localization. A homology model based on the structure of a class II chitinase from barley was used as an aid to interpreting the available data. The analysis suggests that the class IV enzyme recognizes an even shorter segment of the substrate than class I or II enzymes. This observation might help to explain why class IV enzymes are better suited to attack against pathogen cell walls. PMID- 15277756 TI - Intracellular localization of a class IV chitinase from yam. AB - Genomic DNA encoding a class IV chitinase was cloned from yam (Dioscorea opposita Thunb) leaves in previous research (Biosci. Biotechnol. Biochem., 68, 1508-1517 (2004)). But this chitinase had an additional sequence composed of eight amino acids (a C-terminal extension) at the C-terminal, compared with class IV chitianses from other plants. In order to clarify the role of this C-terminal extension in cellular localization, plants and suspension-cultured cells of Nicotiana tabacum were transformed with either the cloned yam class IV chitinase gene carrying the C-terminal extension or its truncated gene by the Agrobacterium mediated method, and then their localization was investigated. The results suggest that the C-terminal extension of yam class IV chitinase plays a role as a targeting signal for plant vacuoles. This is the first report presenting the existence of vacuolar type class IV chitinase. PMID- 15277757 TI - Metabolism of benzoquinone by yeast cells and oxidative characteristics of corresponding hydroquinone: application to highly sensitive measurement of yeast cell density by using benzoquinone and a chemiluminescent probe. AB - The metabolic efficiency of seven derivatives of 1,4-benzoquinone (BQ) by yeast cells and the oxidative characteristics of the corresponding hydroquinones (HQs) were studied by electrochemical, spectrophotometric and chemiluminescent methods. The spectrophotometric method was based on the reduction of a tetrazolium salt to formazan dye during the autoxidation of HQs generated by yeast cells under alkaline conditions. The amounts of HQs detected directly by the electrochemical method did not agree with those calculated from the formazan dye obtained by the spectrophotometric method. A tetrazolium salt was reduced to a formazan dye by both the superoxide anion radical (O2-*) generated during the autoxidation of 2,3,5,6-tetramethyl-1,4-HQ and by HQ itself. Little formazan dye was formed, and hydrogen peroxide (H2O2) was then finally produced during the autoxidation of 1,4 HQ or 2-methyl-1,4-HQ. Formazan dye and H2O2 were generated at a certain ratio during the autoxidation of derivatives of dimethyl-1,4-HQ or 2,3,5-trimethyl-1,4 HQ. The analytical method based on chemiluminescence with lucigenin and 2,3,5,6 tetramethyl-1,4-BQ was applied to highly sensitive measurement of the yeast cell density. A linear relationship between the chemiluminescence intensity and viable cell density was obtained in the range of 1.2 x 10(3) - 4.8 x 10(4) cells/ml. The detection limit was 4.8 x 10(2) cells/ml. PMID- 15277758 TI - Gene cloning and characterization of a Bacillus vietnamensis metalloprotease. AB - A Bacillus vietnamensis metalloprotease (BVMP) with high affinity toward collagen was isolated and purified from the culture supernatant of Bacillus vietnamensis 11-4 occurring in Vietnamese fish sauces. The BVMP gene was cloned and its nucleotide and coded amino acid sequences determined. BVMP consists of 547 amino acid residues, with the zinc-binding sites conserved in common metalloproteases. It shares 57% amino acid identity with thermolysin originating from Bacillus thermoproteolyticus. The three-dimensional structure of BVMP was deduced by computer-aided modeling with the use of the known three-dimensional thermolysin structure as a template. Like thermolysin, BVMP cleaved the oxidized insulin B chain at the peptide bonds involving the N-terminal sides of hydrophobic and aromatic amino acids. BVMP also showed high hydrolytic activity toward gelatin, collagen, casein, and elastin, especially toward the skeletal proteins at increased NaCl concentration. The high activity was found to be due to enhanced affinity to the substrates. Kinetical data on BVMP indicated that the Km values for the hydrolysis of Cbz-GPGGPA as a collagen model decreased as the concentration of added NaCl increased. Some contribution of this enzyme during the aging of fish sauces at high salt concentrations can thus be expected. PMID- 15277759 TI - apg15-1, a UGA mutant allele in the Saccharomyces cerevisiae APG16 gene, and its suppression by a cytoplasmic factor. AB - Autophagy is a complex cellular process by which starving cells utilize cytoplasmic macromolecules as nutritional resources. In Saccharomyces cerevisiae, more than 15 genes are involved in this process and most of them have been cloned and characterized by now. But there remains a complementation group represented by a single mutation, apg15-1, unclear as to its molecular nature. We obtained DNA fragments that functionally complemented apg15-1 and found that the responsible ORF, YMR159C, was already assigned as APG16. It was further found that apg15-1 was a UGA allele in which the 243rd base of the 450 bp coding region of APG16 was converted from C to T, and that the previously observed complementation between apg15-1 and apg16D was attributable to the action of a cytoplasmic omnipotent suppressor. This suppressor was readily cured by guanidine HCl and also by overexpression or disruption of HSP104, indicating its close similarity to the PSI prion-like factor. Since apg15-1 is a mutation highly sensitive to termination suppression, it can be used as a tool to detect weak termination suppressors. PMID- 15277760 TI - Glucosyldiacylglycerol enhances reciprocal activation of prourokinase and plasminogen. AB - Reciprocal activation of prourokinase (pro-u-PA) and plasminogen is an important mechanism in the initiation and propagation of local fibrinolytic activity. We found that glucosyldiacylglycerol (GDG) enhanced the reciprocal activation by 1.5 to 2-fold at 0.7-16 microM, accompanying increased conversions of both zymogens to active two-chain forms. The reciprocal activation system consists of (i) plasminogen activation by pro-u-PA to form plasmin, (ii) pro-u-PA activation by the resulting plasmin to form two-chain u-PA (tcu-PA), and (iii) plasminogen activation by the resulting tcu-PA. Whereas GDG minimally affected steps (ii) and (iii) in isolated systems, it markedly enhanced step (i) in the absence of the conversion of pro-u-PA to tcu-PA. GDG significantly increased the intrinsic fluorescence of pro-u-PA (6.7%), but not that of tcu-PA or plasminogen. The large change in intrinsic fluorescence suggests that GDG selectively affects pro-u-PA to alter its conformation, and this mechanism may account for enhancement of its intrinsic plasminogen activator activity. PMID- 15277761 TI - Cloning of the Ruminococcus albus cel5D and cel9A genes encoding dockerin module containing endoglucanases and expression of cel5D in Escherichia coli. AB - An EcoRI chromosomal DNA fragment of Ruminococcus albus F-40 that conferred endoglucanase activity on Escherichia coli was cloned. An open reading frame (ORF1) and another incomplete reading frame (ORF2) were found in the EcoRI fragment. The ORF2 was completed using inverse PCR genome walking technique. ORF1 and ORF2, which confront each other, encoded cellulases belonging to families 5 and 9 of the glycoside hydrolases and were designated cel5D and cel9A respectively. The cel5D gene encodes 753 amino acids with a deduced molecular weight of 83,409. Cel5D consists of a signal peptide of 24 amino acids, a family 5 catalytic module, a dockerin module, and two family-4 carbohydrate-binding modules (CBMs). The cel9A gene encodes 936 amino acids with a deduced molecular weight of 104,174, consisting of a signal peptide, a family-9 catalytic module, a family-3 CBM, and a dockerin module. The catalytic module polypeptide (rCel5DCat) derived from Cel5D was constructed, expressed, and purified from a recombinant E. coli. The truncated enzyme hydrolyzed cellohexaose, cellopentaose, and cellotetraose to yield mainly cellotriose and cellobiose with glucose as a minor product, but the enzyme was less active toward cellotriose and not active toward cellobiose, suggesting that this enzyme is a typical endoglucanase. rCel5DCat had a Km of 3.9 mg/ml and a Vmax of 37.2 micromol/min/mg for carboxymethycellulose. PMID- 15277762 TI - Expression of a single-chain antibody against indole-3-acetic acid in Escherichia coli. AB - A hybridoma cell line that produces a monoclonal antibody specific for indole-3 acetic acid (IAA) was prepared. The DNA fragments coding the variable regions of the light and the heavy chains of the antibody were prepared by PCR using the cDNA of the antibody as a template. A chimera DNA for a single chain variable fragment (scFv) was constructed, and expressed in Escherichia coli. The scFv antibody expressed in E. coli as well as the original monoclonal antibody showed a specific binding to IAA. PMID- 15277763 TI - Effects of protein transport inhibitors on the distribution and secretion of the fusion protein RntA-EGFP in Aspergillus oryzae. AB - The distribution of the secreted protein ribonuclease T1 (RntA) fused with the enhanced green fluorescent protein (EGFP), RntA-EGFP, was visualized in hyphae of Aspergillus oryzae in the presence of a protein transport inhibitor, brefeldin A, cytochalasin A, or nocodazole. During treatment with the protein transport inhibitors, the distribution of RntA-EGFP changed and distinct patterns of fluorescence accumulation were observed. The addition of brefeldin A caused RntA EGFP fluorescence to appear in reticular networks, and the disruption of the polymerization of actin filaments by cytochalasin A caused an increase in RntA EGFP fluorescence intensity in the hyphae without accumulation in a specific cellular component. In contrast, RntA-EGFP fluorescence was distributed in different parts of a hypha during treatment with nocodazole, a compound that depolymerizes microtubules. In addition, quantitative analysis was performed using the RntA-EGFP visualization system to analyze the relative amount of RntA EGFP secreted into the culture medium during treatment with the protein transport inhibitors. PMID- 15277764 TI - Identification of (+)-phyllocladene, (--)-sandaracopimaradiene, and (+)-kaurene as new fungal metabolites from fusicoccin-producing Phomopsis amygdali F6. AB - A chemical analysis of the diterpene hydrocarbons produced by fusicoccin producing fungus Phomopsis amygdali F6 identified five phyllocladene-related tri- and tetracyclic diterpene hydrocarbons. The presence of (+)-phyllocladene, (--) sandaracopimaradiene, (+)-isopimara-8,15-diene, and (+)-pimara-8(14),15-diene in the fungus was demonstrated by GC-MS, 1H-NMR, and [alpha]D measurements. (+) Kaurene was also identified by GC-MS and chiral capillary GC. The possible biosynthetic relationship of these metabolites is discussed. PMID- 15277765 TI - Running inhibits osteoporosis induced by protein-deficient (PD) food intake. AB - Running at 0.7 km/h for 10 min every day inhibited development of osteoporosis caused by protein deficient (PD) food intake. Urine alkaline phosphatase (ALP), a marker of bone formation osteoporosis, was not elevated in rats fed PD, when the osteoporosis was inhibited by running. Estrogen supplementation increased bone breaking energy (BBE), but did not increase bone mineral density (BMD), and did not decrease urinary ALP levels. PMID- 15277766 TI - Purification and characterization of O-Acetylserine sulfhydrylase of Corynebacterium glutamicum. AB - We highly purified O-acetylserine sulfhydrylase from the glutamate-producing bacterium Corynebacterium glutamicum. The molecular mass of the purified enzyme was 34,500 as determined by SDS-polyacrylamide gel electrophoresis, and 70,800 as determined by gel filtration chromatography. It had an apparent Km of 7.0 mM for O-acetylserine and a Vmax of 435 micromol min-1 (mg x protein)-1. This is the first report of the cysteine biosynthetic enzyme of C. glutamicum in purified form. PMID- 15277767 TI - New 2(3-->20)abeotaxane and 3,11-cyclotaxane from needles of Taxus cuspidata. AB - Two new taxoid metabolites, 2alpha,7beta,10beta-triacetoxy-5alpha,13alpha dihydroxy-2(3-->20)abeotaxa-4(20),11-dien-9-one (1) and 2alpha-acetoxy-5alpha cinnamoyloxy-9alpha,10beta-dihydroxy-3,11-cyclotax-4(20)-en-13-one (2), were isolated from the methanol extract of needles of the Japanese yew, Taxus cuspidata. PMID- 15277768 TI - 8-Hydroxydaidzein, an aldose reductase inhibitor from okara fermented with Aspergillus sp. HK-388. AB - The aldose reductase (AR) inhibitor, 8-hydroxydaidzein, was isolated and identified from a methanolic extract of okara (soybean pulp) fermented with the fungal strain, Aspergillus sp. HK-388. 8-Hydroxydaidzein showed non-competitive inhibition of human recombinant AR with respect to DL-glyceraldehyde, its Ki value being evaluated as 7.0 microM. PMID- 15277770 TI - Growth and carotenoid production of Thraustochytrium sp. CHN-1 cultured under superbright red and blue light-emitting diodes. AB - Isomers of astaxanthin produced by Thraustochytrium sp. CHN-1 are identified as (3S,3S')-trans-astaxanthin, (3R,3R')-trans-astaxanthin and (3S,3S')-cis astaxanthin by chirality column HPLC, and 1H and 13C NMR. We studied the effects of light generated by superbright blue, red and near-red LEDs on the growth and carotenoid production of Thraustochytrium sp. CHN-1. Thraustochytrium sp. CHN-1 responded to blue LEDs light: It produced carotenoid pigments (astaxanthin) PMID- 15277769 TI - Mitogenic properties of pokeweed lectin-D isoforms on human peripheral blood lymphocytes: non-mitogen PL-D1 and mitogen PL-D2. AB - We investigated native structures and mitogenic properties of pokeweed lectin-D isoforms (PL-D1 and -D2) on human peripheral blood lymphocytes along with other isolectins (PL-A to -C). Both native PL-D isoforms appeared to behave as monomers. PL-D2 proliferated the lymphocytes like PL-C, whereas PL-D1 had no mitogenicity. PL-D1 acquired mitogenic activity after trimming of the C-terminal dipeptide. PMID- 15277771 TI - Enhanced calcium absorption in the small intestine by a phytate-removed deamidated soybean globulin preparation. AB - Soybean globulins were deamidated after removing phytate using ion-exchange resins, and then hydrolyzed by digestive enzymes. The phytate-removed deamidated soybean globulins (PrDS) retained high calcium-binding ability even after the hydrolysis by digestive enzymes. PrDS and its hydrolysates enhanced calcium absorption from the small intestine when injected into the small intestine together with a calcium solution. PMID- 15277772 TI - Antimicrobial activities of diterpene dialdehydes, constituents from myoga (Zingiber mioga Roscoe), and their quantitative analysis. AB - The antimicrobial activities of the three diterpene dialdehydes, miogadial, galanal A and galanal B, isolated from flower buds of the myoga (Zingiber mioga Roscoe) plant were investigated with some strains of bacteria, yeasts and molds. Among the three compounds, miogadial exhibited relatively greater antimicrobial activity than the others against Gram-positive bacteria and yeasts. Galanals A and B also behaved as antimicrobial agents against Gram-positive bacteria and yeasts. The content of miogadial in the flower buds was much higher than that in the leaves, whereas galanals A and B were contained at high levels in the leaves and rhizomes. PMID- 15277773 TI - Classification of brassinosteroid-regulated genes based on expression profiles in bri1 and in response to a protein kinase inhibitor, staurosporin. AB - To gain insight into the brassinosteroid (BR) signaling pathway, the expression of BR-regulated genes was analysed in the BR-signaling mutant br-insensitive 1 (bri1), and in the presence of a protein kinase inhibitor, staurosporin. BR regulated genes were classified based on the results. This classification will perhaps prove useful in BR-signalling studies using BR-regulated genes as molecular markers. PMID- 15277774 TI - Fusicocca-3(16),10(14)-diene, and beta- and delta-araneosenes, new fusicoccin biosynthesis-related diterpene hydrocarbons from Phomopsis amygdali. AB - Further isolation and examination of fusicoccane hydrocarbons biosynthetically related to fusicoccin from Phomopsis amygdali allowed us to identify new fungal diterpene hydrocarbons of fusicoccadiene and araneosene. These were assigned as (+)-fusicocca-3(16),10(14)-diene, and (+)-beta- and (+)-delta-araneosenes. These findings led to the experimental clarification of the structures of the biosynthetic hydrocarbon intermediates presumed earlier. PMID- 15277775 TI - Production of lupin acid phosphatase in transgenic rice for use as a phytate hydrolyzing enzyme in animal feed. AB - The acid phosphatase gene from lupin was expressed in transgenic rice plants under the control of the maize ubiquitin promoter or rice chlorophyll a/b binding protein (Cab) promoter. Transgenic rice leaves exhibited up to an 18-fold increase in phytate-hydrolyzing activity. Based on the phytate-hydrolyzing activity at pH 5.5, more than 85% this activity was retained after heat-treatment at 80 degrees C for 15 min, and the heterologous enzyme in leaf sections and leaf extracts was relatively stable during storage. A distinct increase in released phosphate was observed when the heterologous enzyme was mixed with the feed extract. These results suggest that the heterologous enzyme in rice plants may maintain its desired characteristics as a phytate-hydrolyzing enzyme when added to animal feed. PMID- 15277776 TI - Dihydrocoronatine, promising candidate for a chemical probe to study coronatine-, jasmonoid- and octadecanoid-binding protein. AB - Coronatine (1), its synthetic analogs (6-13) and jasmonic acid induced various volatiles in rice leaves. In the range of 0.01-0.1 mM, dihydrocoronatine (7) exhibited 4-687 times higher activity for linalool emission than that of 1. The radioactive derivative of 7, [4,5-3H]-7, was employed to identify the putative coronatine-binding protein in rice leaves. 7 would be a promising candidate for a chemical probe to study cornatine-binding protein related to the jasmonoid and octadecanoid signaling pathway in higher plants. A detailed study of coronatine binding protein in rice leaves and cell culture with [4,5-3H]-7 is now in progress. PMID- 15277777 TI - Interaction between a negative regulator (Msa2/Nrd1) and a positive regulator (Cpc2) of sexual differentiation in Schizosaccharomyces pombe. AB - The sexual differentiation of Schizosaccharomyces pombe is controlled by many cellular components which have not been fully characterized. We isolated a gene called msa2 as a multi-copy suppressor of a sporulation abnormal mutant (sam1). Msa2p is identical with Nrd1p which has been characterized as a factor that blocks the onset of sexual differentiation. The yeast two-hybrid system was used to identify Cpc2p, a fission yeast homolog of the RACK1 protein, that interacted with Msa2p/Nrd1p. We confirmed that Msa2p/Nrd1p interacted with Cpc2p in S. pombe cells. An epistatic analysis of msa2/nrd1 and cpc2 suggests that Msa2p/Nrd1p was an upstream regulator for Cpc2p. A localization analysis of Cpc2p and Msa2p/Nrd1p indicates that both proteins were predominantly localized in the cytoplasm. The interaction of negative regulator Msa2p/Nrd1p with positive regulator Cpc2p suggests a new regulatory circuit in the sexual differentiation of S. pombe. PMID- 15277778 TI - A risk-adjusted approach to comparing the return on investment in health care programs. AB - The league table approach to rank ordering health care programs according to the incremental cost-effectiveness ratio is a common method to guide policy makers in setting priorities for resource allocation. In the presence of uncertainty, however, ranking programs is complicated by the degree of variability associated with each program. Confidence intervals for cost-effectiveness ratios may be overlapping. Moreover, confidence intervals may include negative ratios and the interpretation of negative cost-effectiveness ratios is ambiguous. We suggest to rank mutually exclusive health care programs according to their rate of return which is defined as the net monetary benefit over the costs of the program. However, how does a program with a higher expected return but higher uncertainty compare to a program with a lower expected return but lower risk? In the present paper we propose a risk-adjusted measure to compare the return on investment in health care programs. Financing a health care program is treated as an investment in a risky asset. The risky asset is combined with a risk-free asset in order to construct a combined portfolio. The weights attributed to the risk-free and risky assets are chosen in such a manner that all programs under consideration exhibit the same degree of uncertainty. We can then compare the performance of the individual programs by constructing a risk-adjusted league table of expected returns. PMID- 15277779 TI - Changes in hospital quality after conversion in ownership status. AB - This paper examines the effects of conversions between For-Profit and Not-For Profit forms on quality of medical care in California hospitals. The sample includes elderly patients treated in California's private hospitals from 1990 to 1998 for Acute Myocardial Infarction and Congestive Heart Failure. The results suggest that converted hospitals have experienced quality changes before conversion and that ignoring these changes may bias the estimates of conversion effects. Both conversions are found to have some adverse consequences: Hospitals that converted to FP form show an increase in AMI mortality rates, while those converted to NFP status indicate an increase in CHF mortality outcomes. PMID- 15277780 TI - New evidence on hospital profitability by payer group and the effects of payer generosity. AB - This study provides (a) new estimates of U.S. hospital profitability by payer group, controlling for hospital characteristics, and (b) evidence about the intensity of care for particular diseases associated with the generosity of the patient's payer and other payers at the same hospital. The conceptual framework is a variant of the well-known model of a local monopolist selling in a segmented market. Effects of two kinds of regulation are considered. The data are taken from hospital accounting reports in four states in FY2000, and detailed discharge summaries from the Healthcare Cost and Utilization Project of the Agency for Healthcare Research and Quality. The profitability of inpatient care for privately insured patients was found to be about 4% less than for Medicare, but 14% higher than for Medicaid and only 9% higher than for self-pay patients. We found significant direct associations but not external effects of payer generosity on the intensity of care. PMID- 15277781 TI - Multiple prior years of health expenditures and Medicare health plan choice. AB - This study examines the effect of multiple prior years of health expenditures on the probability of enrollment in a Medicare HMO. Beneficiaries may require more than one year of prior expenditure data to form a reliable estimate of future expenditures if health expenditures have a significant transitory component. We used a logit model to estimate the influence of 1991-1993 Part A expenditures and demographic data on the choice of health plans in 1994. The results indicate that beneficiaries use multiple years of expenditures in their choice of health plan and the effect of prior spending declined with time. PMID- 15277782 TI - Multi-bed vascular disease: past, present, and future. PMID- 15277783 TI - Modulating thrombotic potential in catheter-based percutaneous coronary and peripheral vascular interventions. AB - Thrombosis is an obligatory consequence of all percutaneous vascular interventions. Balloon angioplasty, intravascular stents and other devices routinely used to facilitate dilatation of critical vascular stenoses result in fracture of the intima and exposure of the thrombogenic subendothelium with initiation and perpetuation of platelet activation and aggregation. This not uncommonly results in thrombus formation that may lead to abrupt vessel closure, distal ischemia and tissue infarction, and target organ dysfunction. Fortunately, advances in our understanding of the mechanisms that underlie vascular thrombosis have led to advances in the use of adjunctive pharmacological agents that modulate this pathophysiological response and have led to important reductions in the incidence and severity of thrombotic complications of percutaneous transluminal interventions. PMID- 15277784 TI - The efficacy and safety of perioperative antiplatelet therapy. AB - Widespread adoption of the antiplatelet agents into everyday clinical practice has revolutionized contemporary care of the cardiovascular patient. Major adverse cardiovascular events including death, myocardial infarction, stroke, and recurrent angina have all been shown to be significantly decreased when these agents are employed in the treatment of coronary atherosclerosis, acute coronary syndromes, myocardial infarction, and in the setting of percutaneous coronary intervention. As a growing number of patients on antiplatelet therapy are undergoing various surgical procedures, the potential risks and benefits these drugs pose perioperatively will become increasingly important. Available data indicate that, when used appropriately, these drugs can be used safely prior to surgery. Efficacy in improving surgical outcomes and in preventing adverse cardiovascular events postoperatively has also been demonstrated. The purpose of this review is to examine the perioperative safety and efficacy of the most widely used antiplatelet agents: aspirin; the thienopyridine clopidogrel; and the glycoprotein (GP) IIb/IIIa inhibitors abciximab, eptifibatide, and tirofiban. This information, coupled with emerging platelet monitoring techniques, may help provide additional assistance to the clinician to manage therapy and guide appropriate timing of both cardiac and noncardiac surgery. PMID- 15277785 TI - Facilitating optimal care of acute coronary, cerebrovascular and peripheral vascular syndromes in the emergency department: the role of oral antiplatelet therapy. AB - The benefits of aspirin use in the emergent care of MI and stroke have been well established. Recent studies have further demonstrated the importance of antiplatelet therapy in the acute setting, primarily with the use of intravenous glycoprotein IIb/IIIa receptor inhibitors. Aspirin and the thienopyridines (ticlopidine and clopidogrel) are oral antiplatelet agents that interfere with platelet activation in complementary, but separate pathways. Combination therapy of clopidogrel and aspirin has demonstrated benefit for the management of acute coronary syndromes, ischemic cerebrovascular disease and peripheral vascular disease in several large trials. This article reviews the pathophysiology of platelet activation, landmark trials on oral antiplatelet agents, and the current recommendation for the use of oral antiplatelet agents in the emergency department. PMID- 15277786 TI - Evolving concepts in the triad of atherosclerosis, inflammation and thrombosis. AB - Recent developments into antherothrombosis, the leading cause of morbidity and mortality in Western Society, may help to change our treatment strategy to a more casual approach. The composition of the atherosclerotic plaque, rather than the percent stenosis, appears to be a critical predictor for both risk of plaque rupture and subsequent thrombogenicity. A large lipid core, rich in tissue factor (TF) and inflammatory cells including macrophages, and a thin fibrous cap with compromise of its structural integrity by matrix degrading enzymes, such as metalloproteinases (MMPs), render a lesion susceptible to rupture and subsequent acute thrombosis. Thrombosis may lead to a complete occlusion or, in the case of mural thrombus or intraplaque hemorrhage, to plaque progression. Disruption of a vulnerable or unstable plaque (type IV and Va lesions of the AHA classification) with a subsequent change in plaque geometry and thrombosis may result in an acute coronary syndrome. The high-risk plaque tend to be relatively small, but soft or vulnerable to "passive" disruption because of high lipid content. Inflammatory processes are important components of all stages of atherosclerotic development, including plaque initiation and disruption. As such the early steps in atherosclerotic lesion formation are the over expression of endothelial adhesive protein (i.e. selectins, VCAM and ICAM), chemotactic factors (MCP-1), growth factors (M-CSF), and cytokines (IL-2) that will facilitate the recruitment, internalization and survival of blood-borne inflammatory cells into the vascular wall. Macrophages, following what appears to be a defense mission by protecting the vessel wall from excess lipid accumulation, may eventually undergo apoptosis with release of MMPs and TF. Specific cell recruitment in the vessel wall and build-up of the extracellular matrix are coordinated by a wide variety of stimulators and inhibitors. Active interaction of immune competent cells within the atherosclerotic lesions appears to play a pivotal role in the control of atherosclerotic plaque evolution and, therefore, deserves particular attention from the research community with the ultimate goal of improving preventive and therapeutic medical approaches. Inflammation, thrombosis and atherosclerosis are interdependent and define a triad within the complex pathogenic process of atherothrombosis. PMID- 15277787 TI - Utilization of guidelines and computer-based technology to achieve optimal care in atherothrombotic vascular disease. AB - Recently there have been numerous advances in the treatment of UA/NSTEMI based on large randomized trials. However, there are also data from several registries that have shown underutilization of these therapies. National guidelines, such as the ACC/AHA Guidelines are designed to provide clinicians recommendations for optimal care. There are some initial data from the TIMI 3 and GUARANTEE registries that publication of such guidelines does appear to help improve compliance with evidence-based optimal care, however, it appears that critical pathways and specific tools such as standardized orders or checklists are needed to more fully implement the recommendations. One newer type of tool are Palm Pilot programs such as the 3 presented here, where it is hoped that their use at the bedside with provide clinicians rapid access to evidence-based medicine ina format that can be useful in a busy practice setting. PMID- 15277788 TI - Multi-bed vascular disease and atherothrombosis: scope of the problem. AB - While atherosclerosis has traditionally been divided into three types of disease, coronary artery or coronary heart disease (CHD), cerebrovascular disease, and peripheral vascular or peripheral arterial disease (PAD), it is now clear that atherosclerosis is a systemic disease caused by the same pathologic processes regardless of the vascular bed involved. The burden of disease is enormous both in the US and around the world with 61,800,000 Americans affected with one or more types of CVD, responsible for 958,775 deaths annually at a cost of approximately US 329.2 billion dollars annually. Despite trends of decreasing cardiovascular mortality, the global burden of cardiovascular disease is expected to rise, with CHD and stroke becoming the first and fourth most common causes of mortality and morbidity globally. Atherosclerosis is a multibed process with a substantial portion of patients afflicted with disease in more than one bed, although often assymptomatic. Now that there are multiple therapies available to modify and treat atherosclerosis and atherosclerotic risk factors, identification and treatment of these patients are important since their leading cause of death is from co-existing cardiovascular disease. PMID- 15277791 TI - Use of fast-setting hydroxyapatite cement for secondary craniofacial contouring. AB - Patients who have previously had surgical correction of major craniofacial deformities will often have residual contour deformities they wish to have improved at a later date. The development of hydroxyapatite cement has simplified these procedures. The setting time is reduced to 5 to 8 minutes by mixing the cement with a phosphate-based solution, increasing the tensile strength, and maintaining the same biocompatibility and osseoconductivity. This study includes 48 patients who presented with a variety of residual contour irregularities secondary to a craniofacial congenital anomaly or a posttraumatic defect. All but one of the patients with congenital craniofacial conditions had their initial surgical correction performed by the senior author (Magee) and had regular follow up visits. Variable amounts of hydroxyapatite cement were used according to the size of the defect to be corrected. Five patients had a postoperative complication: two infections, one seroma, one persistent swelling, and one drain retention. Patients were followed from 6 months to 3 years (mean, 1 year 5 months). Good results were achieved in 38 patients, acceptable results with minor asymmetries were seen in seven patients, and three other patients required a second intervention to obtain a better contour. Cranioplasty with fast-setting hydroxyapatite cement is a simple and reliable procedure, with a low complication rate. Attention to simple technical and operative principles can provide excellent results. PMID- 15277789 TI - Patient-specific antiplatelet therapy. AB - Platelets are key players in thrombosis, and have thus become the main targets in the acute treatment of, as well as in the primary and secondary prevention against, thrombotic cardiovascular diseases. Three main classes of anti-platelet agents are currently available for clinical use: aspirin, the thienopyridines, and the intravenous GPIIb/IIIa antagonists. While these therapies are beneficial in the mean patient population, they may produce adverse effects in selected patient subgroups. In this article, we review the three main classes of antiplatelet drugs, discussing them in terms of the patient characteristics that are likely to influence their overall efficacy and safety. PMID- 15277792 TI - Solvent-dehydrated calvarial allografts in craniofacial surgery. AB - Craniofacial surgery almost always requires the use of bone grafting. Although autografts are the standard procedure for bone grafting, it is sometimes not possible to harvest bone, and autografts have particular risks. The use of allograft bone provides a reasonable alternative to meet the need for graft material. Solvent dehydration is a multistage procedure in which human cadaveric bone is processed by osmotic exchange baths and gamma sterilization. This processing avoids the risk of infection transmission, decreases antigenicity, and does not weaken the mechanical properties of the bone. Solvent-dehydrated, gamma irradiated human calvarial bone allografts were used for reconstruction of craniofacial deformities in 24 patients between 1988 and 2002. Resorption of the allografts and results of the surgical intervention were evaluated with plain radiographs and three-dimensional computed tomography 12 months after surgery, in 21 patients. Serologic tests for human immunodeficiency virus-1 antibody, hepatitis B surface antigen, and hepatitis C antigen were also performed. Biopsy specimens were taken from the allografts. Average follow-up in this group was 30 months (range, 8 to 60 months), and results of serologic tests were negative in all patients. Seventy-one percent of the patients (15 of 21) showed no resorption, with partial and complete allograft fusion. One patient had nearly total graft loss and the remaining five patients had 10 to 25 percent graft resorption. Rigid fixation of the allograft, contact with the dura and periosteum, and prevention of dead spaces around the allograft are the most important factors in achieving a satisfactory result. In solvent-dehydrated bone allografts, sterilization and antigenic tissue cleaning are achieved after several steps with a minimal dose of radiation. The result is a nonantigenic, sterile mechanical scaffold that can tolerate external forces. Although autografts are the standard in craniofacial surgery, solvent-dehydrated calvarial bone allografts produced successful results in selected cases. PMID- 15277793 TI - Surgical anatomy of the levator veli palatini: a previously undescribed tendinous insertion of the anterolateral fibers. AB - The purpose of this study was to describe the previously unreported tendinous insertion of the anterolateral fibers of the levator veli palatini (levator) and discuss possible implications for levator function and cleft palate repair. The velopharyngeal anatomy in normal adult cadavers was studied, with histologic confirmation of anatomical findings. These findings were compared with a more limited study of levator anatomy in cleft palates at the time of intraoperative muscle dissection. Just before entering the velum, the levator divides into two parts. The smaller bundle of muscle fibers (anterolateral part) runs anteriorly, close to the lateral pharyngeal wall, and inserts into the palatine aponeurosis through a number of fine tendons. The main part of the muscle runs medially into the velum, where it fans out and forms the levator sling with the contralateral levator. The possible function of the anterolateral part of the levator is discussed. Inadequate release of the tendinous insertions at the time of palate repair may tether the levator anteriorly and compromise muscle retropositioning or may result in splitting of the levator, so that only part of the levator is retropositioned. PMID- 15277794 TI - Nasal reconstruction with a forehead flap in children younger than 10 years of age. AB - Nasal reconstruction has been analyzed extensively in adults but not in children. The purpose of this article is to review the authors' experience with the forehead flap for nasal reconstruction in 10 children under the age of 10 during a 10-year period. Outcomes were assessed by an objective grading system for cosmetic surgical results. Subjective criteria were also applied by an assistant surgeon and by the patients' relatives. Appropriate results were obtained by the following principles: (1) A modified approach that considers three subunits consisting of the dorsum, tip, and ala was used; (2) a forehead flap is the best option for an entire subunit or a full-thickness defect repair; (3) the forehead flap design should be paramedian, oblique, and opposite to the major defect to avoid the hairline and allow better caudal advancement; (4) ear or costal cartilages are good options for structural support (the septum is a nasal growth center that should not be touched); (5) infundibular undermining of vestibular mucosa, turnover flaps, and skin grafts are good options for internal lining; (6) reconstruction is a three-stage procedure (an intermediate operation is added to thin the flap and perform secondary revisions for lining and support); (7) reconstruction should be completed before the child is school aged, to achieve good aesthetic results immediately and avoid psychosocial repercussions; and (8) the reconstructed nose, with skin, lining, and support, will grow with the child (no final surgery should be planned at the age of 18, other than revisions of late complications). PMID- 15277796 TI - Chewing and swallowing after surgical treatment for oral cancer: functional evaluation in 196 selected cases. AB - One hundred ninety-six patients treated for oral cancer between 1992 and 1999 self-scored their speech, chewing, and swallowing using a new self-questionnaire (Functional Intraoral Glasgow Scale) developed at Canniesburn Hospital, Glasgow, to assess the functional efficiency of patients treated for intraoral cancer. The patients were distributed into 12 homogeneous groups, according to the site and size of surgical resection, carefully mapped out on standard diagrams of the oral cavity. The functional outcome for chewing and swallowing was correlated to the site and size of resected tissue, to the reconstruction modality, and to radiotherapy and compared with the speech quality. The general trend is very similar for both chewing and swallowing; the smaller the resections, the better the functional outcome. Chewing was mostly affected by resections of the floor of the mouth, whereas swallowing was mostly affected by demolition of the base of the tongue and of the retromolar trigone. Speech showed a better postoperative recovery than chewing and swallowing. The reconstruction modality did not influence the eventual outcome for either function. Radiotherapy in combination with surgery is a negative functional prognostic factor. A correlation between site and size of excision and functional outcome is presented using color multiple-view diagrams for immediate appreciation to identify positive and negative prognostic factors. PMID- 15277797 TI - Extended vertical trapezius myocutaneous flap in head and neck reconstruction as a salvage procedure. AB - In surgical treatment of head and neck cancer, when local tumor recurrence or failure of the previous reconstruction method occurs, reoperation for reconstruction of complicated soft-tissue defects can become a challenge for the plastic surgeon. This article describes the authors' experience with the extended vertical trapezius myocutaneous flap for head and neck complicated soft-tissue defects in nine patients ranging in age from 17 to 72 years. The causes of the defects were squamous cell carcinoma of the external ear (n = 2), lip (n = 2), larynx (n = 1), and oral cavity floor (n = 1); congenital hemifacial atrophy temporomandibular joint ankylosis (n = 1); synovial sarcoma at the mandibular ramus (n = 1); and malignant fibrous histiocytoma at the posterior cranial fossa (n = 1). Eight of the nine patients had previously been operated on using other flap procedures, including free flaps and/or distant pedicled flaps (pectoralis major and deltopectoral flaps). One patient had been operated on using a graft procedure. After failure of the previous flap procedures in four patients and tumor recurrence in five patients, the extended vertical trapezius myocutaneous pedicled flap was used as a salvage procedure. The mean flap size was 7 x 34 cm. The flap was based solely on the transverse cervical artery. Superior muscle fibers of the trapezius were preserved and the caudal end of the flap was extended from 10 to 13 cm beyond the caudal end of the trapezius muscle. Three weeks postoperatively, the pedicle was separated. No flap failure occurred. The donor sites were closed primarily. There were no disabilities with regard to shoulder motion. Tumor recurrence was observed in two patients. In conclusion, for complicated soft-tissue defects of the head and neck, the extended vertical trapezius flap can be preferred as a salvage procedure because it is a simple, reliable, large flap that is located far enough from the damaged area. PMID- 15277798 TI - Rhinophyma: dispelling the myths. AB - Rhinophyma is a relatively common condition in the west of Scotland. The Canniesburn Plastic Surgery Unit receives 12 to 13 new patients per year for surgical treatment. The reported incidence of simultaneous carcinoma in the setting of rhinophyma is on the order of 15 to 30 percent. There are conflicting reports about the association between alcohol and rhinophyma in the literature, and these are supported with little or no statistical evidence. Retrospective epidemiologic data on 45 cases of rhinophyma are presented. An audit of case notes was performed to examine histology and also alcohol consumption in these cases. The authors found no coincidental malignancies at the time of surgery, which is contrary to many previous publications. The alcohol consumption of the rhinophyma cases was compared with that of a control group that consisted of 48 men presenting for blepharoplasty. The series did not demonstrate a positive association between alcohol and rhinophyma when compared with a similar cohort of patients presenting for blepharoplasty surgery (p > 0.20) or with statistics available from the Scottish Health Survey. PMID- 15277799 TI - Cadaveric study of the arterial anatomy of the upper lip. AB - Arterial distribution of the upper lip was investigated in this study. The location, course, length, and diameter of the superior labial artery and its alar and septal branches were determined on 14 preserved cadaver heads. Another cadaver head was used to show the arterial tree by the colored silicone injection technique. The superior labial artery was the main artery of the upper lip and always originated from the facial artery. The superior labial artery was 45.4 mm in length, with a range from 29 to 85 mm. The mean distance of the origin of the superior labial artery from the labial commissura was 12.1 mm. The superior labial artery was 1.3 mm in external diameter at its origin. The mean distance of origin of the superior labial artery from the lower border of the mandible was 46.4 mm. The alar division of the superior labial artery was mostly found as a single branch (82 percent). Its mean length was 14.8 mm and the mean diameter at the origin was 0.5 mm. The distance between the origins of the superior labial artery and the septal branch was 33.3 mm. The septal branch was single in most of the cases (90 percent). The mean length of the septal branch was 18.0 mm and the diameter at its origin was 0.9 mm. After all dissections, it was concluded that the arterial distribution of the upper lip was not constant. The superior labial artery can occur in different locations unilaterally and bilaterally, with the branches showing variability. PMID- 15277800 TI - Satisfaction with breast reconstruction in women with bilateral prophylactic mastectomy: a descriptive study. AB - Prophylactic bilateral mastectomy is an option for women who are at an increased risk of developing breast cancer. Prophylactic mastectomy is often performed with immediate reconstruction (i.e., at the same time and under the same anesthetic as the mastectomy). Satisfaction with reconstruction has been described previously for women with mastectomy for breast cancer. However, the authors know of no previous research that has reported on satisfaction with reconstruction in patients who have electively sought mastectomy for the prevention of breast cancer. Women in the province of Ontario who had undergone prophylactic bilateral mastectomy plus breast reconstruction between 1991 and 2000 were asked to rate their level of satisfaction with the cosmetic results of their mastectomy and reconstruction and their overall satisfaction with their decision to have prophylactic mastectomy. Women were also asked whether they experienced complications associated with their surgery and what types of complications they experienced. Thirty-seven women completed questionnaires for this study, and all of them had immediate breast reconstruction after prophylactic mastectomy. The majority of women (70.3 percent) reported being satisfied or extremely satisfied with the cosmetic results of their breast reconstruction. Women with self reported postsurgical complications (16.2 percent) were significantly less satisfied with reconstruction than those who did not report complications (p = 0.009). Personal subjective risk of breast cancer before prophylactic mastectomy was negatively correlated with satisfaction with reconstruction (r = -0.38, p = 0.024) and with subjective risk estimation after prophylactic surgery (r = -0.54, p = 0.001). Women who did not worry about developing breast cancer after prophylactic mastectomy had significantly higher levels of satisfaction with breast reconstruction than those who continued to worry (p < 0.001). Women who reported an improved body image after reconstruction were significantly more likely to report higher levels of satisfaction than those who reported a diminished body image (p = 0.007). The majority of women were satisfied with the cosmetic results of breast reconstruction after prophylactic mastectomy. Women who overestimated their breast cancer risk had lower satisfaction levels. Correcting overestimation of breast cancer risk in women who have prophylactic mastectomy may improve satisfaction with reconstruction following prophylactic mastectomy. PMID- 15277801 TI - The oblique rectus abdominis musculocutaneous flap: revisited clinical applications. AB - The authors present their experience with a previously described but infrequently used variation of the rectus abdominis myocutaneous flap. Skin paddles angled obliquely from the line of the rectus abdominis and toward the rib cage were successfully carried on periumbilical perforators from the inferior epigastric system. Skin paddle dimensions ranged from 6.5 to 12 cm in width and from 10 to 27 cm in length in 14 consecutive patients. In six of the 14 patients, the flap was used intraabdominally to obliterate radiated pelvic defects and to close radiated vaginal defects. Five flaps were placed externally to repair radiated wounds of the perineum, thigh, and trunk, and the remaining three cases were performed as free tissue transfers. One cadaver injection study was performed to redemonstrate the preferential flow of fluid in a superior-oblique direction from periumbilical perforators. Termed the oblique rectus abdominis musculocutaneous ("ORAM") flap, this flap variation has significant advantages in terms of ease of dissection and versatility over its flap cousins the vertical rectus abdominis musculocutaneous flap and the transverse rectus abdominis musculocutaneous flap. PMID- 15277802 TI - Technical variations of the bipedicled TRAM flap in unilateral breast reconstruction: effects of conventional versus microsurgical techniques of pedicle transfer on complications rates. AB - In cases of unilateral breast reconstruction with a transverse rectus abdominis musculocutaneous (TRAM) flap, poorly perfused tissue, which is normally excised to avoid subsequent fat necrosis, must sometimes be used to achieve adequate breast size and projection. In such cases, incorporation of a second vascular pedicle into the flap design improves perfusion. The authors retrospectively examined their experience with bipedicled TRAM flap-based unilateral breast reconstruction to determine whether the use of microsurgical rather than conventional (nonmicrosurgical) techniques for flap transfer resulted in lower incidences of flap-site fat necrosis and donor-site hernia/bulge. The authors retrospectively reviewed the medical records of all patients who underwent unilateral breast reconstruction with a bipedicled TRAM or deep inferior epigastric perforator flap between January of 1991 and March of 2001. Group 1 consisted of patients who had undergone flap transfer using a conventional technique for both pedicles; group 2, patients who had flap transfer using a conventional technique for one pedicle and a microsurgical technique for the other; and group 3, patients who had flap transfer using a microsurgical technique for both pedicles. Of the 863 patients identified, 72 (8.3 percent) had undergone reconstruction using a bipedicled flap. There were 43 patients in group 1, 24 patients in group 2, and five patients in group 3. Only one case of total flap loss had occurred (group 1). Partial flap loss occurred in two patients in group 1 (5 percent) and three patients in group 2 (13 percent). Fat necrosis occurred more frequently in groups 1 (23 percent) and 2 (29 percent) than in group 3 (0 percent) (p = 0.5, Fisher's exact test). Similarly, bulge or hernia was more common in groups 1 (12 percent) and 2 (4 percent) than in group 3 (0 percent) (p = 0.6, Fisher's exact test). In this study, patients who received a bipedicled TRAM flap using microsurgical techniques alone (group 3) appeared to have better flap perfusion and less frequent hernia/bulge than did patients who underwent flap transfer using conventional (group 1) or combined techniques (group 2). However, these differences were not statistically significant, and this trend must be verified in a larger study. PMID- 15277804 TI - Distally based dorsal forearm fasciosubcutaneous flap. AB - Use of a local flap is often required for the reconstruction of a skin defect on the dorsum of the hand. For this purpose, a distally based dorsal forearm fasciosubcutaneous flap based on the perforators of the posterior interosseous artery was developed. From 1997 until 2002, this flap was used to reconstruct skin defects on the dorsum of the hand in nine patients at Chonnam National University Medical School. The sizes of these flaps ranged from 10 to 14 cm in length and from 5 to 7 cm in width. The flaps survived in all patients. Marginal loss over the distal edge of the flap was noted in one patient. Three flaps that developed minimal skin-graft loss were treated successfully with a subsequent split-thickness skin graft. The long-term follow-up showed good flap durability and elasticity. The distally based dorsal forearm fasciosubcutaneous flap is a convenient and reliable alternative for reconstructing skin defects of the dorsum of the hand involving vital structure exposure. It obviates the need for more complicated and time-consuming procedures. PMID- 15277806 TI - Upper extremity limb salvage with microvascular reconstruction in patients with advanced sarcoma. AB - Limb salvage is a viable alternative to amputation in many cases of advanced sarcoma. The authors examined their experience with microvascular reconstruction of upper extremity defects after sarcoma resection, focusing on oncologic and functional outcomes. A retrospective analysis yielded 17 patients who underwent 18 free flap procedures and met the inclusion criteria. Most patients (71 percent, n = 12) had recurrent sarcoma at presentation to the authors' institution. Malignant fibrous histiocytoma was the most common pathologic subtype (n = 6). High-grade tumors were present in 94 percent of patients (n = 16). The free flap survival rate was 100 percent. The rectus abdominis flap was the most common free flap used (39 percent; n = 7). Local recurrence occurred in nine flaps (50 percent), and five patients ultimately required amputations. Six patients (35 percent) had distant recurrence. The mean Enneking score for limb function was 73 percent of the maximum (21.9 of 30). The 5-year disease-specific survival rate was 61.3 percent. In select patients with advanced upper extremity sarcoma undergoing limb salvage, microvascular flap reconstruction can provide reliable, safe coverage with reasonable preservation of function. PMID- 15277808 TI - Free dorsoulnar perforator flap transfers for the reconstruction of severely injured digits. AB - The aim of this study was to investigate the feasibility of transferring the free dorsoulnar perforator flap nourished by the cutaneous perforator branched dorsoulnar artery to reconstruct severely injured fingers under upper arm anesthesia. Between April of 2001 and April of 2002, 13 free dorsoulnar perforator flaps were used in 13 patients. There were 11 men and two women ranging in age from 18 to 64 years, with an average age of 38 years. The affected fingers were one thumb, four index fingers, five middle fingers, two ring fingers, and one little finger. All cases were performed under upper arm anesthesia combined with intravenous local anesthesia. The operative time ranged from 103 to 140 minutes, with an average time of 120 minutes. The flap size ranged from 1 x 3 to 3 x 4 cm, and was transferred from the same forearm of the injured finger. All donor sites were closed primarily without a skin graft. The aim of reconstruction for fingers was to repair a traumatic defect (five cases), partial necrosis following replantation (two cases), and soft-tissue defects resulting from resection of a scar (three cases) and to revascularize ischemic fingers (three cases). All flaps survived completely. After repair of the flow through circulation of the common digital artery and ischemic finger, a postoperative angiogram showed the vascular patency and hypervascularity of the reconstructed fingers, and the patients' complaints were reduced. The free dorsoulnar perforator flap under regional anesthesia is first reported; it may become one valuable option as a very small flap for the treatment of repairing intercalated or segmental defects as a flow-through flap for soft-tissue defects and ischemic fingers. PMID- 15277809 TI - Surgical treatment of osteoarthritis of the carpometacarpal joint of the thumb: a systematic review. AB - In most cases of basal joint osteoarthritis, surgery becomes an option at stages II, III, and IV, as classified by Eaton. Controversy exists regarding which technique achieves the best outcome. This systematic review was undertaken to address the question of which technique, if any, offers the best outcome to patients with osteoarthritis of the first carpometacarpal joint greater than stage II. A thorough search of the electronic databases Cochrane, Cinahl, Healthstar, and MEDLINE/PubMed was undertaken to identify reviews and articles on primary comparative studies of the different surgical options. The methodological quality of the retrieved articles was assessed on the basis of specific criteria. Inclusion criteria were applied to 44 of 254 possibly relevant articles. Eight reviews and 18 comparative studies met the criteria and were reviewed. Each of the techniques, arthrodesis, trapeziectomy with or without biological/synthetic interposition, osteotomy, and joint replacement, was associated with unique benefits and risks. There was great variability in outcome measurements. The majority of retrieved review articles claim that ligamentous reconstruction and tendon interposition may represent the best option; however, validity assessment of these studies revealed methodological flaws. Furthermore, results from the articles on comparative studies indicate that ligamentous reconstruction and tendon interposition may provide no additional benefit when compared with arthrodesis and trapeziectomy alone or with tendon interposition. There is no consensus as to which clinical outcomes are most important in thumb basal joint surgery and how these should be measured. This renders the appraisal and comparison of such studies a challenging task. Until large randomized controlled trials that compare techniques in similar populations with respect to staging and prognostic factors are undertaken and the clinical outcomes are clearly defined, surgeons will continue to claim superiority of one technique over another without supporting evidence. PMID- 15277810 TI - A New Surgical Technique for Polysyndactyly of the Toes without Skin Graft. AB - Reconstruction for polysyndactyly of the toes aims at cosmetic improvement. A previous method that uses a skin graft has inherent disadvantages of mismatched pigmentation between the graft and the surrounding skin and scar formation at the donor site. The authors' new improved surgical technique for the treatment of polysyndactyly of the toes does not require a skin graft and therefore avoids these problems. The authors designed a subcutaneous flap from the distal portion of a rectangular flap of skin from the dorsal side of the interdigital webbing and moved the former flap to the sidewall of the base of a toe. Both flaps are the same size; therefore, an interdigital space had to be of sufficient size to accommodate both of them. To ensure an adequate blood supply to the flap, careful handling of the subcutaneous flap is essential for success. This procedure can apply to polysyndactyly of the fourth, fifth, and sixth toes when the fourth and fifth toes adhere over the distal side of the distal interphalangeal joint and when the skin on the dorsal side of the fifth toe, regarded as the excessive one, is at lease twice the size of the dorsal rectangular flap. Ten patients with polysyndactyly of the toe were treated with this method. Aesthetically good results were obtained. PMID- 15277811 TI - Long-term follow-up of total penile reconstruction with sensate osteocutaneous free fibula flap in 18 biological male patients. AB - Surgical reconstruction of the penis is challenging because of the many cosmetic and functional (e.g., sexual intercourse and voiding) requirements that must be addressed. Since the free sensate osteocutaneous fibula flap was first described for total penile reconstruction in 1993 it has been widely accepted, with its advantages and minimal shortcomings. In this article, the authors present the longest follow-up of biologically male patients with free fibular phalloplasties. Since 1994, 18 biologically male patients with total penile losses for various reasons were treated with free sensate osteocutaneous fibula flaps. All patients were included in the study. The ages of the patients ranged between 20 and 26 years (mean, 22.2 years). The average follow-up period was 5.4 years (range, 1 to 9 years). Patient satisfaction was evaluated by a questionnaire regarding both quality of orgasm and daily activities. Conventional radiographic imaging, magnetic resonance imaging, and bone mineral densitometry were performed to evaluate the fate of the bony component of the flap. Also, sensibility was evaluated by bulbocavernous reflex and penile somatosensory evoked potentials testing in nine patients. Six patients married, and five of them had six children. Most patients and their partners reported pleasurable sexual intercourse and orgasm. Conventional radiographs of the fibular bone in neophalluses showed robust, calcified bone structure without any evidence of bone resorption or fracture. The magnetic resonance images showed the cortical substance and spongiosum of the bone marrow, which are characteristic signs of bone viability. After intravenous injection of gadolinium, the neophallus bone showed uptake of contrast medium. Viability of neophallus bone was shown even at 9-year follow-up (the longest follow-up in the literature). Dual energy x-ray absorptiometry measurements of the penile bone grafts showed that fibular components in the penis had bone mineral density values that were close to but lower than those of intact fibula in the same subjects. These results were considered as evidence of viability of bone grafts. Neural integrity was found between the nerves of the neophallus and the residual penile bodies by both bulbocavernous reflex and penile somatosensory evoked potentials tests. In conclusion, free sensate fibula flap phalloplasty provides the cosmetic and functional requirements that an ideal penis should have. All results put an end to the discussion that the fibular component of the neophallus could resorb. Constitution of neural integrity is important in terms of pleasurable sexual intercourse. The authors believe the free sensate osteocutaneous fibula flap should be considered as the standard in penile reconstruction. PMID- 15277813 TI - Emergency room coverage: an evolving crisis. AB - Historically, a newly graduated plastic surgeon in the United States could build a practice from his or her emergency room coverage. The historical cliche was for the surgeon to be affable, able, and available, and from that basis one's practice would grow. Emergency room exposure was an avenue for starting a practice, developing recognition, and, after that, building a referral pattern. Recently, the cross-shifting influence of management care, rising malpractice insurance costs, and risk ratio are changing this cliche to a crisis. An evaluation of a 2 1/2-year exposure to emergency room coverage has revealed a completely different profile. A total of 300 patient visits resulting in 69 surgical operations were evaluated for insurance and remuneration history. The findings indicated a significant remuneration dilemma for emergency room coverage. Interestingly, a remuneration problem exists in a market different from what one would expect. In this study, a sample from a suburban hospital, rather than an inner-city university hospital, is the greater problem. PMID- 15277814 TI - Smoke in the operating theater: an unregarded source of danger. AB - Monopolar electrocautery devices are being used in operating theaters worldwide and have become a "sine qua non" in modern surgery. Despite being widespread, the use of electrocautery is not harmless, because by burning the tissue with rather low temperatures as compared with usual combustion, toxic gases evolve and particles are dispersed and are inhaled by the staff in the operating theater. Samples of this smoke, which evolves particularly densely during reduction mammaplasty, were analyzed using a carbon dioxide laser photoacoustic spectrometer. Eleven gas components could be identified and quantified. In particular, the established concentration of 2-fur-ancarboxaldehyde (furfural) measured at 2 cm from the point of origin was outstandingly high, being 12 times higher than the occupational exposure limit. More than half of the identified gases do not even have any occupational exposure limit specifications. Because of the expected dilution at the height of the operating distance (the surgeon's nose), the present measured concentrations do not allow any conclusion on a direct health danger to the operating team. Because of laser spectroscopy, the present work reveals not only the involved gases but also their respective concentrations near the point of origin. These data are prerequisite for further studies, which are mandatory, verifying the effective concentrations of the inhaled gases. PMID- 15277815 TI - Reduction of adhesions with composite AlloDerm/polypropylene mesh implants for abdominal wall reconstruction. AB - Ventral hernia repair often includes the use of structural prosthetic materials, such as polypropylene mesh, that can induce dense abdominal adhesions to peritoneal structures. AlloDerm (LifeCell Corp., Branchburg, N.J.), a commercially available decellularized human dermal analogue with its native basement membrane components intact, is gradually revascularized and replaced with autologous tissue after implantation. The authors hypothesized that AlloDerm integrated with polypropylene mesh would reduce adhesions and provide a biodegradable scaffold to generate an autologous vascularized tissue layer separating the abdominal viscera from the mesh. Ventral hernia defects (3 x 1 cm) in 19 guinea pigs were repaired using an inlay technique with polypropylene mesh alone (n = 6) or with composite implants constructed by integrating polypropylene mesh and AlloDerm with its basement membrane surface oriented toward (polypropylene/AlloIn, n = 7) or away from (polypropylene/ AlloOut, n = 6) the peritoneal cavity. At 4 weeks, the authors determined the amount of mesh implant surface area covered by adhesions, the strength of the adhesions [graded from 0 (none) to 3], and the incidence of bowel adhesions. Histologic analyses were performed on full-thickness tissue sections from the repair sites. The mean surface areas affected by adhesions and mean adhesion strength were significantly lower in the polypropylene/AlloIn (area, 12.4 percent; mean grade, 1.0) and polypropylene/AlloOut (area, 9.5 percent; mean grade, 0.5) groups than in the polypropylene group (area, 79.5 percent; mean grade, 2.9); there were no such differences between the polypropylene/AlloIn and polypropylene/AlloOut groups. The bowel was adherent to 67 percent of polypropylene repairs and 0 percent of the composite mesh repairs. The AlloDerm was remodeled to form a vascularized tissue layer beneath the mesh in composite repairs, unlike the significantly thinner, dense scar layer that formed in the polypropylene repairs. Immunohistochemical labeling for factor VIII showed neovascularization throughout the AlloDerm. The AlloDerm thus functioned as a biodegradable tissue scaffold, guiding the formation of a thick, well-vascularized tissue layer separating the polypropylene mesh from intraperitoneal structures. This significantly reduced both the amount of surface area covered by adhesions and adhesion strength. Basement membrane orientation had no effect. Composite mesh implants composed of structural prosthetic materials integrated with AlloDerm may have useful clinical applications for abdominal wall reconstruction by reducing adhesions and providing a vascularized tissue layer to separate and protect the peritoneal structures from polypropylene mesh fibers. PMID- 15277816 TI - Ultrasound-assisted liposuction in the treatment of lipodystrophy secondary to cushingoid-type disease of unknown cause. PMID- 15277817 TI - Chemotherapy-induced neuropathy: treatment by decompression of peripheral nerves. PMID- 15277818 TI - The "silicone suture" for tissue expansion without an expander: a new device for repair of soft-tissue defects after burns. PMID- 15277820 TI - Long-term experience with endoscopic surgical treatment for congenital muscular torticollis in infants and children: a review of 85 cases. PMID- 15277821 TI - The Tinel sign: a historical perspective. AB - The Tinel sign is one of the most well-known and widely used clinical diagnostic tools in medicine. Aside from Jules Tinel, after whom the sign is named, several authors have described the famous "tingling" sign seen in regenerating injured nerves. In fact, Tinel was not the first to present the sign to the scientific community. The clinical value and utility of the Tinel sign have remained in question since its introduction; many may misinterpret the sign as a prelude to complete functional recovery of injured nerves, when in fact it only signals the progress of nerve regeneration. Today the Tinel sign is widely associated with the diagnosis of carpal tunnel syndrome and in the evaluation of regenerating peripherally injured nerves. Knowledge of the history and misconceptions surrounding the sign provides clinicians today with a greater appreciation of current debates on the use of the Tinel sign. PMID- 15277822 TI - Shelling and plastic surgery. PMID- 15277823 TI - Hemodynamic physiology and thermoregulation in liposuction. AB - Little is known about the physiology of large-volume liposuction. Patients are exposed to prolonged procedures, general anesthesia, fluid shifts, and infusion of high doses of epinephrine and lidocaine. Consequently, the authors examined the thermoregulatory and cardiovascular responses to liposuction by assessing multiple physiologic factors. The aims of their study were to serially determine hemodynamic parameters perioperatively, to quantify perioperative and postoperative plasma epinephrine levels, and to chronologically document fluctuations in core body temperature. Five female volunteers with American Society of Anesthesiologists' physical status I and II underwent moderate- to large-volume liposuction. Heart rate, blood pressure, mean pulmonary arterial pressure, cardiac index, and central venous pressure were monitored. Serum epinephrine levels and core body temperature were assessed perioperatively. The hemodynamic responses to liposuction were characterized by an increase in cardiac index (57 percent), heart rate (47 percent), and mean pulmonary arterial pressure (44 percent) (p < 0.05). Central venous pressure was not significantly altered. Maximum epinephrine levels were observed 5 to 6 hours after induction. Significant correlations between cardiac index and epinephrine concentrations were shown intraoperatively (r = 0.75). All patients developed intraoperative low body temperatures (mean 35.5 degrees C). An overall enhanced cardiac function was observed in patients subsequent to large-volume liposuction. The etiology of the altered cardiac parameters was multifactorial but may have been attributable in part to the administration of epinephrine, which counters the effects of general anesthesia and operative hypothermia. Additional explanations for raised cardiac output may be hemodilution or emergence from general anesthesia. Elevated mean pulmonary arterial pressure may be a result of subclinical fat embolism demonstrated in previous porcine studies, although fat was not observed in urine. The unchanged central venous pressure levels indicate that young healthy patients with compliant right ventricles can accommodate the fluid loads of large-volume liposuction. Overall hemodynamic parameters remained within safe limits. Within these surgical parameters, patients should be clinically screened for cardiovascular and blood pressure disorders before liposuction is undertaken, and preventative measures should be taken to limit intraoperative hypothermia. PMID- 15277825 TI - Pharmacokinetics and safety of lidocaine and monoethylglycinexylidide in liposuction: a microdialysis study. AB - High doses of lidocaine are administered to patients undergoing liposuction. Monoethylglycinexylidide, the active metabolite of lidocaine, is 80 to 90 percent as potent as lidocaine, and its relative toxicity is approximately that of lidocaine. Monoethylglycinexylidide has not previously been measured in studies on lidocaine in liposuction. The aims of this study were to characterize systemic exposure to lidocaine and monoethylglycinexylidide and to measure lidocaine and monoethylglycinexylidide levels within the tissues. Five female volunteers between the ages of 29 and 40 years underwent liposuction. Lidocaine (1577 to 2143 mg, corresponding to 19.9 to 27.6 mg/kg) was infiltrated during the procedure. Levels of lidocaine and monoethylglycinexylidide in blood and lipoaspirate were assessed perioperatively. Tissue lidocaine and monoethylglycinexylidide levels were measured postoperatively using a microdialysis technique in vivo. The peak (maximal) concentration of lidocaine plus monoethylglycinexylidide was 2.2 to 2.7 microg/ml. Time to peak lidocaine plus monoethylglycinexylidide was 8 to 28 hours after infiltration began. Absorbed lidocaine was estimated to be 911 to 1596 mg; therefore, 45 to 93 percent (mean, 64 percent) of the infiltrated dose was ultimately absorbed. Lipoaspirate analysis showed that 9.1 to 10.8 percent (mean, 9.7 percent) of the infiltrated dose was removed during the procedure. Tissue lidocaine levels below 5 microg/ml were demonstrated from 4 to 8 hours postoperatively. The peak lidocaine plus monoethylglycinexylidide concentration was within safe limits in this group of subjects. Time to peak lidocaine plus monoethylglycinexylidide signifies a delayed peak and therefore a longer period of potential lidocaine toxicity than was originally thought. Microdialysis results demonstrated that tissue lidocaine levels may be subtherapeutic within 4 to 8 hours of the procedure. Investigation into factors controlling the resorption of lidocaine during liposuction is warranted in an effort to improve the duration of effect. Furthermore, considering the active metabolite monoethylglycinexylidide, longitudinal studies are necessary to determine whether improving the side effect profile of lidocaine by reducing the dose administered during liposuction may be possible without decreasing the perioperative analgesic effect. PMID- 15277827 TI - Nose surgery: how to prevent a middle vault collapse--a review of 50 patients 3 to 21 years after surgery. AB - There is a conception, likely a misconception, that when performing a nasal osteotomy with a concomitant dorsal hump removal, the upper lateral cartilages are detached or damaged and, over the long-term, respiratory difficulties result because of a middle vault collapse or interference with the internal nasal valve. A follow-up of 50 patients between 3 and 21 years postoperatively provides evidence that this can be prevented. The vast majority (82 percent) reported they were breathing very well for an average of 6.5 years postoperatively. Of the authors' own 38 primary rhinoplasty patients, only two patients (5 percent) reported respiratory difficulties. The authors are unable to substantiate that either the osteotomy or the dorsal hump removal was responsible. Of the 12 patients who had their primary rhinoplasty performed elsewhere, six (50 percent) reported respiratory difficulties before the secondary rhinoplasty at this clinic. Furthermore, an appreciable improvement in breathing was reported by 66.7 percent of these patients after the secondary rhinoplasty. The authors conclude that their gentle proper surgical technique, combined with a good understanding of nasal physiology (with respect to the septum, inferior turbinates, and external and internal valves), allows them to perform a concomitant dorsal hump removal and osteotomy without interfering with nasal physiology. PMID- 15277828 TI - Balanced angular profile analysis. AB - To evaluate current preferences and ethnic differences of female soft-tissue profiles, 71 profile photographs of famous female models were collected from Internet Web pages and divided into four groups (Korean, 22; Japanese, 15; Chinese, 16; and Western, 18). Eleven soft-tissue landmarks were recorded on each photograph and 16 angular measurements were made by using V-ceph (CyberMed, Inc., Seoul, Korea). Data from each group are presented to show the means, ranges, p and F values, standard deviations, and standard errors of each measurement. In addition, individual measurements for each group were compared with those of the other groups by one-way analysis of variance using a p value corrected for multivariable testing. Between-group mean value differences were calculated using a Tukey's studentized range test (HSD), at a significance level of p = 0.05. Most of the variables were similar in the groups. Significant between-group differences (p < 0.05) were found for angle of alar curvature point, profile convexity, interlabial contour, and nasolabial contour. In addition, we divided all data into two groups (Western and Asian). The t test (with significance level set to p = 0.05) was performed to compare the two. Significant between-group differences (p < 0.05) were found for angle of alar curvature, angle of labiale inferius, profile convexity, and lower lip projection angle, but no significant racial differences were found in terms of several profile angles. These findings suggest that point of ala curvature point, subnasale, and the labiale inferius of Asian models may differ from those of Western models. These peculiar angular patterns of Asian models led the authors to create a new characteristic angular concept, termed the "ethnic pyramid," which is composed of soft-tissue profile points of alar curvature point, subnasale, pronasale, and labiale inferius. This ethnic pyramid describes the characteristic patterns of the ethnic differences. The results of this study suggest that the soft-tissue profiles of famous female models have some common features but also show differences among ethnic groups and races. This simple method of profile analysis may provide aesthetic surgeons with a simple formula and reference data for creation and application of an attractive face. On the basis of their balanced angular profile analysis data, the authors suggest that appropriate and harmonious aesthetic operations reflecting these differences should be considered. PMID- 15277829 TI - A cadaveric analysis of the ideal costal cartilage graft for Asian rhinoplasty. AB - Augmentation rhinoplasty of the Asian nose may be effectively accomplished with alloplastic materials. However, certain circumstances mandate the use of autologous grafts (e.g., dorsal augmentation that exceeds 8 mm and patient intolerance of alloplastic implants). Septal and auricular cartilages are inadequate for dorsal augmentation of the Asian nose. The use of costal cartilage for autologous augmentation in select Asian patients has proven to be a reliable method in more than 500 operative cases during a 10-year period. This study was designed to evaluate the ideal costal cartilage graft for augmentation rhinoplasty. Forty-two preserved cadavers were studied for the relationship of the individual rib cartilages to the surrounding tissue and for the length and caliber of each costal cartilage. The seventh rib was found to be the ideal rib graft by virtue of its safe location and overall size for grafting. The seventh rib is situated over the abdominal cavity, so the risk of pneumothorax is insignificant. The internal thoracic artery and vein descend in close apposition behind the first to sixth ribs but begin a course medial to the ribs inferior to this point, and therefore vascular injury during seventh-rib harvesting is unknown. The seventh rib also provides the greatest overall available length (90.7 mm, right; 89.6 mm, left) and thickness (17.6 mm, right; 17.5 mm, left). Despite the more conspicuous location of the incision required to harvest the seventh rib, the limited 3-cm incision that is used has healed favorably in almost all cases. The other major drawback for seventh-rib harvesting is the dissection required through the overlying rectus abdominis muscle, but little technical difficulty or postoperative morbidity is added with muscle dissection. The seventh rib is advocated as the ideal choice for augmentation rhinoplasty and potentially other recipient sites. PMID- 15277830 TI - Treatment of facial fat atrophy related to treatment with protease inhibitors by autologous fat injection in patients with human immunodeficiency virus infection. AB - A parallel effect of the use of protease inhibitors in human immunodeficiency virus-positive patients is the appearance of facial fat atrophy. To correct this, the authors propose lipoinjection of autologous fat into the areas of facial atrophy by a technique based on the atraumatic procurement of fat and posterior treatment with decantation, centrifugation, and cleaning of other material obtained by aspiration. The method of fat injection is also important and is performed by means of interlaced tunnels and the introduction of a small volume of fat into each tunnel as a graft. In 30 percent of the cases, reinjection of fat was required during the first postoperative months. The results obtained after the experience of 2 years were very satisfactory. PMID- 15277832 TI - SMAS graft of the nasolabial area during deep plane rhytidectomy. AB - The extensive list of treatments for the nasolabial area illustrates that the problem is not solved yet. Although a better understanding of the anatomy and physiology of the nasolabial area has been developed, the ideal treatment has not been found. It seems that a combined approach based on summation of partially effective modalities is the best option. The authors present their experience with combined deep plane rhytidectomy, malar fat lift, and superficial musculoaponeurotic system graft in the treatment of the aging nasolabial area. Results in 70 consecutive patients are presented with this simple, nonscarring, inexpensive technique. PMID- 15277834 TI - Submammary flap for correction of severe sequelae from augmentation mammaplasty. AB - A submammary flap was used in 20 patients with severe cicatricial retractions and loss of the inferior pole of the breast caused by inadequately treated mammary implant infections. This axial flap can be used with a medial pedicle, based on the perforating branches of the epigastric artery or the distal part of the internal mammary artery, or laterally based, nourished by the intercostal perforators. After 6 months, reimplantation was performed in 15 patients. The authors' follow-up ranged between 8 months and 6 years. This transverse adipocutaneous flap procedure is very simple to perform, the donor site is sutured primarily without additional undermining, and the resultant scar lies hidden within the submammary fold. It provides tissue with similar skin texture and color match. Its versatility allows it to be used as a full-thickness tissue replacement or partially or totally de-epithelialized for soft-tissue reconstruction. It can also be used as an island flap. For all of these reasons, in emotionally distressed patients with low compliance with surgical treatments and additional scars, this flap is the authors' flap of choice for reconstruction. PMID- 15277836 TI - Sensibility of the abdomen after abdominoplasty. AB - Abdominal skin hypesthesia may occur after abdominoplasty. The purpose of this study was to find out (1) which sensibility modalities are decreased and (2) which areas of the abdominal wall are affected, so that patients can be warned preoperatively about this condition. Forty patients were divided in two groups of 20 patients each. In the control group, patients had no previous abdominal incisions. The sensibility evaluation of patients from the experimental group was made from 12 to 60 months after abdominoplasty, with an average of 31.5 months. These patients were divided into two groups of 10 patients each, a short-term follow-up group (12 to 30 months postoperatively) and a long-term follow-up group (31 to 60 months postoperatively). The abdominal skin was divided into 12 areas; nine were above the abdominoplasty incision and three were below it. Sensibility to superficial touch, superficial pain, and hot and cold modalities was recorded as positive in all areas by a variable number of patients of the experimental group. However, in area 8 (hypogastric area), a statistically significant number of patients had decreased sensibility in all sensibility modalities (Fisher's test and t test). Patients in the experimental group also showed decreased sensibility to hot and cold temperature in area 11 (pubic area). Sensibility to pressure decreased significantly in all areas of the abdomen when compared with the control group (t test). When patients of the short-term follow-up group were compared with those of the long-term follow-up group, there was no statistically significant difference for all modalities of sensibility in the areas studied, except for area 5. In this area it was found that long-term follow-up patients recovered sensibility to cold and hot temperatures. These findings help plastic surgeons to orient their patients about possible risk of exposure to injuries in the areas with decreased sensibility after abdominoplasty. Most importantly, as these patients have decreased sensibility to pressure and hot temperature in a more extensive area of the abdomen, they are exposed to a higher risk of burn injury. PMID- 15277838 TI - More caveats for plastic surgeons. PMID- 15277839 TI - "If you were beginning your career...". PMID- 15277840 TI - Bioactive glass for bone replacement in craniomaxillofacial reconstruction. PMID- 15277843 TI - Gracilis flap: a variation of the main vascular pedicle. PMID- 15277844 TI - Medicine and plastic surgery: science or art? PMID- 15277846 TI - The influence of fibroblasts on the differentiation of cultured epithelial cells in vitro. PMID- 15277847 TI - Presenting postoperative images in a better light. PMID- 15277848 TI - Nipple reconstruction using autologous graft. PMID- 15277850 TI - Cosmetic breast implants and suicide. PMID- 15277851 TI - Tumescent technique in capsulotomies: a useful adjunct. PMID- 15277852 TI - Necessity of inspection of superficial palmar arch by blunt dissection from exit portal in endoscopic carpal tunnel release. PMID- 15277853 TI - The benefit of transverse carpal ligament reconstruction following open carpal tunnel release. PMID- 15277855 TI - Smoking and elective surgery: a survey of United kingdom plastic surgery consultants. PMID- 15277856 TI - A more reliable fixation method for the high-density porous polyethylene implant. PMID- 15277858 TI - Maxillofacial reconstruction with prefabricated osseous free flaps: a 3-year experience with 24 patients. PMID- 15277860 TI - Use of nasal prosthesis for anesthesia in head and neck surgery. PMID- 15277861 TI - Various materials may aid in teaching surgical procedures. PMID- 15277862 TI - An application of a rice bag stitch. PMID- 15277863 TI - A broken forceps for anterior scoring: a cheap and simple device for anterior scoring. PMID- 15277864 TI - Using a scalpel as a screwdriver. PMID- 15277865 TI - Constructing your own suction drainage system. PMID- 15277866 TI - Donor-site recurrence of rectal adenocarcinoma after reverse latissimus dorsi flap. PMID- 15277867 TI - Occult primary melanoma of the urethra: a long-term survival case. PMID- 15277868 TI - A simplified technique for umbilical reconstruction. PMID- 15277869 TI - An inexpensive exhaust system for electrocautery. PMID- 15277870 TI - Surgical glove finger pieces for hair separation. PMID- 15277871 TI - The use of a simple syringe as a stent for McIndoe vaginal construction. PMID- 15277872 TI - "Pocket measure": an exclusive tool for measuring and recording pressure ulcer pockets. PMID- 15277873 TI - Use of acutely burned skin of a digit as a flap donor site. PMID- 15277874 TI - Memorable medical mentors: VI. Thomas S. Cullen (1868-1953). PMID- 15277895 TI - Patient choice cesarean: an evidence-based review. AB - Primary elective cesarean performed on a patient's request now comprises 4% to 18% of all cesareans and 14% to 22% of elective cesareans in reported series. Patients most commonly choose cesarean because of tocophobia, or fear of childbirth. Almost two thirds of obstetricians surveyed are willing to perform cesarean on request, citing decreased risk of pelvic floor or fetal injury, maintenance of sexual functioning, and physician and patient convenience. Contrasting these beliefs are the limited available data on short- and long-term maternal and perinatal morbidity and mortality that generally favor vaginal delivery. Moreover, comprehensive economic impact assessments of cesarean on request are lacking, and professional organizations do not agree on the ethics of offering patient choice cesarean. TARGET AUDIENCE: Obstetricians & Gynecologists, Family Physicians. LEARNING OBJECTIVES: After completion of this article, the reader should be able to list the reasons that women and obstetricians choose elective cesarean delivery, to outline the ethical aspects of cesarean delivery, and to describe the material and fetal morbidity and mortality associated with cesarean delivery compared to vaginal delivery. PMID- 15277896 TI - Pathophysiology of fetal growth restriction: implications for diagnosis and surveillance. AB - Normal fetal growth depends on the genetically predetermined growth potential and is modulated by fetal, placental, maternal, and external factors. Fetuses with intrauterine growth restriction (IUGR) are at high risk for poor short- and long term outcome. Although there are many underlying etiologies, IUGR resulting from placental insufficiency is most relevant clinically because outcome could be altered by appropriate diagnosis and timely delivery. A diagnostic approach that aims to separate IUGR resulting from placental disease from constitutionally small fetuses and those with other underlying etiologies (e.g., aneuploidy, viral infection, nonaneuploid syndromes) needs to integrate multiple imaging modalities. In placental-based IUGR, cardiovascular and behavioral responses are interrelated with the disease severity. Ultrasound assessment of fetal anatomy, amniotic fluid volume, and growth is complementary to the Doppler investigation of fetoplacental blood flow dynamics. A diagnostic approach to IUGR combining these modalities is presented in this review. TARGET AUDIENCE: Obstetricians & Gynecologists, Family Physicians. LEARNING OBJECTIVES: After completion of this article, the reader should be able to describe the development of the placental interface, to outline the mechanisms of placental insufficiency, and to list the manifestations of placental insufficiency and the tests that can be used to diagnose fetal growth restriction. PMID- 15277897 TI - Contemporary management of type 1 diabetes mellitus in pregnancy. AB - Type 1 diabetes in pregnancy can result in significant short- and long-term morbidity to both mother and offspring if management is suboptimal. This morbidity imposes a considerable financial and health burden on the individual and society at large. There is currently a significant body of knowledge to offer guidance on optimal obstetric management of the woman with type 1 diabetes. Utilization of appropriate management guidelines preconception and during pregnancy is an effective strategy to limit complications of type 1 diabetes and should therefore become the standard of care. TARGET AUDIENCE: Obstetricians & Gynecologists, Family Physicians. LEARNING OBJECTIVES: After completion of this article, the reader should be able to describe the features of a type I diabetic patient, to outline the goals of preconception care in this population of patients, to list the potential adverse effects of diabetes in pregnancy, and to summarize a potential strategy for the management of insulin administration in pregnancy. PMID- 15277904 TI - Are local anesthetics needed for local anesthesia? PMID- 15277905 TI - Anesthetic neurotoxicity: the collision between laboratory neuroscience and clinical medicine. PMID- 15277906 TI - Anesthesia-induced developmental neuroapoptosis. Does it happen in humans? PMID- 15277907 TI - Sleeping with uncertainty: anesthetics and desiccated absorbent. PMID- 15277908 TI - Anesthetic requirement is increased in redheads. AB - BACKGROUND: Age and body temperature alter inhalational anesthetic requirement; however, no human genotype is associated with inhalational anesthetic requirement. There is an anecdotal impression that anesthetic requirement is increased in redheads. Furthermore, red hair results from distinct mutations of the melanocortin-1 receptor. Therefore, the authors tested the hypothesis that the requirement for the volatile anesthetic desflurane is greater in natural redheaded than in dark-haired women. METHODS: The authors studied healthy women with bright red (n = 10) or dark (n = 10) hair. Blood was sampled for subsequent analyses of melanocortin-1 receptor alleles. Anesthesia was induced with sevoflurane and maintained with desflurane randomly set at an end-tidal concentration between 5.5 and 7.5%. After an equilibration period, a noxious electrical stimulation (100 Hz, 70 mA) was transmitted through bilateral intradermal needles. If the volunteer moved in response to stimulation, desflurane was increased by 0.5%; otherwise, it was decreased by 0.5%. This was continued until volunteers "crossed over" from movement to nonmovement (or vice versa) four times. Individual logistic regression curves were used to determine desflurane requirement (P50). Desflurane requirements in the two groups were compared using Mann-Whitney nonparametric two-sample test; P < 0.05 was considered statistically significant. RESULTS: The desflurane requirement in redheads (6.2 vol% [95% CI, 5.9-6.5]) was significantly greater than in dark haired women (5.2 vol% [4.9-5.5]; P = 0.0004). Nine of 10 redheads were either homozygous or compound heterozygotes for mutations on the melanocortin-1 receptor gene. CONCLUSIONS: Red hair seems to be a distinct phenotype linked to anesthetic requirement in humans that can also be traced to a specific genotype. PMID- 15277909 TI - Effect of clonidine on cardiovascular morbidity and mortality after noncardiac surgery. AB - BACKGROUND: Perioperative myocardial ischemia occurs in 20-40% of patients at risk for cardiac morbidity and is associated with a ninefold increase in risk of cardiac morbidity. METHODS: In a prospective, double-blinded, clinical trial, we studied 190 patients with or at risk for coronary artery disease in two study groups with a 2:1 ratio (clonidine, n = 125 vs. placebo, n = 65) to test the hypothesis that prophylactic clonidine reduces the incidence of perioperative myocardial ischemia and postoperative death in patients undergoing noncardiac surgery. Clonidine (0.2 mg orally as well as a patch) or placebo (tablet and patch) was administered the night before surgery, and clonidine (0.2 mg orally) or placebo (tablet) was administered on the morning of surgery. The patch or placebo remained on the patient for 4 days and was then removed. RESULTS: The incidence of perioperative myocardial ischemia was significantly reduced with clonidine (intraoperative and postoperative, 18 of 125, 14% vs. placebo, 20 of 65, 31%; P = 0.01). Prophylactic clonidine administration had minimal hemodynamic effects. Clonidine reduced the incidence of postoperative mortality for up to 2 yr (clonidine, 19 of 125 [15%] vs. placebo, 19 of 65 [29%]; relative risk = 0.43 [confidence interval, 0.21-0.89]; P = 0.035). CONCLUSIONS: Perioperative administration of clonidine for 4 days to patients at risk for coronary artery disease significantly reduces the incidence of perioperative myocardial ischemia and postoperative death. PMID- 15277910 TI - Evaluation of the Alaris Auditory Evoked Potential Index as an indicator of anesthetic depth in preschool children during induction of anesthesia with sevoflurane and remifentanil. AB - BACKGROUND: Autoregressive modeling with exogenous input of middle latency auditory evoked potentials (A-Line autoregressive index [AAI]) has been proposed for monitoring depth of anesthesia in adults. The aim of this study was to evaluate the performance of the AAI during induction of anesthesia with sevoflurane and remifentanil in pediatric patients. METHODS: Twenty preschool children were anesthetized with sevoflurane and remifentanil. AAI, heart rate, and mean arterial pressure were compared for their ability to distinguish between different hypnotic states before inhalation induction and during sevoflurane anesthesia with and without remifentanil infusion. The prediction probability was calculated for discrimination between the predefined case milestones Awake, Spontaneous Eye Closure, and insertion of a laryngeal mask airway during general anesthesia (Laryngeal Mask Insertion). RESULTS: The AAI (mean +/- SD) in Awake children was 79 +/- 10, declining to 59 +/- 22 at Spontaneous Eye Closure and 34 +/- 13 when anesthetized. AAI values significantly overlapped between anesthetic states. For the AAI, the prediction probabilities regarding the ability to discriminate the hypnotic state at the case milestones Awake versus Spontaneous Eye Closure and Awake versus Laryngeal Mask Insertion were 0.77 and 0.99, respectively. In terms of prediction probability values, heart rate and mean arterial pressure were not indicative for anesthetic states. Remifentanil did not influence the AAI. CONCLUSION: During induction of pediatric patients with sevoflurane, the AAI is of higher value in predicting anesthetic states than hemodynamic variables and reliably differentiates between the awake and anesthetized states. However, individual AAI values demonstrate significant variability and overlap between different clinical conditions. PMID- 15277911 TI - Cardioprotective properties of sevoflurane in patients undergoing coronary surgery with cardiopulmonary bypass are related to the modalities of its administration. AB - BACKGROUND: Experimental studies have related the cardioprotective effects of sevoflurane both to preconditioning properties and to beneficial effects during reperfusion. In clinical studies, the cardioprotective effects of volatile agents seem more important when administered throughout the procedure than when used only in the preconditioning period. The authors hypothesized that the cardioprotective effects of sevoflurane observed in patients undergoing coronary surgery with cardiopulmonary bypass are related to timing and duration of its administration. METHODS: Elective coronary surgery patients were randomly assigned to four different anesthetic protocols (n = 50 each). In a first group, patients received a propofol based intravenous regimen (propofol group). In a second group, propofol was replaced by sevoflurane from sternotomy until the start of cardiopulmonary bypass (SEVO pre group). In a third group, propofol was replaced by sevoflurane after completion of the coronary anastomoses (SEVO post group). In a fourth group, propofol was administered until sternotomy and then replaced by sevoflurane for the remaining of the operation (SEVO all group). Postoperative concentrations of cardiac troponin I were followed during 48 h. Cardiac function was assessed perioperatively and during 24 h postoperatively. RESULTS: Postoperative troponin I concentrations in the SEVO all group were lower than in the propofol group. Stroke volume decreased transiently after cardiopulmonary bypass in the propofol group but remained unchanged throughout in the SEVO all group. In the SEVO pre and SEVO post groups, stroke volume also decreased after cardiopulmonary bypass but returned earlier to baseline values than in the propofol group. Duration of stay in the intensive care unit was lower in the SEVO all group than in the propofol group. CONCLUSION: In patients undergoing coronary artery surgery with cardiopulmonary bypass, the cardioprotective effects of sevoflurane were clinically most apparent when it was administered throughout the operation. PMID- 15277912 TI - Effects of Bispectral Index monitoring on ambulatory anesthesia: a meta-analysis of randomized controlled trials and a cost analysis. AB - BACKGROUND: Ambulatory surgery is growing in popularity worldwide. For example, 50-70% of surgical procedures in North America are performed on an ambulatory basis. Use of Bispectral Index (BIS) monitoring for titration of general anesthesia may allow use of less anesthetics, reduction in side effects, and faster patient recovery. METHODS: MEDLINE and other databases were searched for randomized controlled trials examining the use of BIS monitoring versus standard practice in ambulatory surgery patients. Outcomes were extracted from these articles, and a meta-analysis was performed. RESULTS: One thousand three hundred eighty subjects from 11 trials were included in the meta-analysis. The use of BIS monitoring significantly reduced anesthetic consumption by 19%, reduced the incidence of nausea/vomiting (32% vs. 38%; odds ratio, 0.77), and reduced time in the recovery room by 4 min. However, these benefits did not result in significant reduction in time until patient discharge from the ambulatory surgery unit. Cost analysis using pooled costs to reflect North America, Europe, and Asia indicated that use of BIS monitoring increased the cost per patient by 5.55 US dollars because of the cost of BIS electrodes. CONCLUSIONS: The use of BIS monitoring modestly reduced anesthetic consumption, risk of nausea and vomiting, and recovery room time. These benefits did not reduce time spent in the ambulatory surgery unit, and cost of the BIS electrode exceeded any cost savings. PMID- 15277913 TI - A randomized controlled trial comparing the ProSeal Laryngeal Mask Airway with the Laryngeal Tube Suction in mechanically ventilated patients. AB - BACKGROUND: The ProSeal Laryngeal Mask Airway (PLMA) (Laryngeal Mask Company, Henley-on-Thames, United Kingdom) is a new laryngeal mask with a modified cuff designed to improve its seal and a drain tube for gastric tube placement. Similarly, the Laryngeal Tube Suction (LTS) (VBM Medizintechnik Gmbh, Sulz a.N, Germany) is a new laryngeal tube that also has an additional channel for gastric tube placement. This study compared the placement and functions of these two devices. METHODS: One hundred fifty patients undergoing general anesthesia for elective surgery were randomly allocated to the PLMA (n = 75) or LTS (n = 75). Oxygenation and ventilation, ease of insertion, fiberoptic view, oropharyngeal leak pressure, ventilatory data, ease of gastric tube insertion, and postoperative airway morbidity were determined. RESULTS: After successful insertion of the devices in 96% of patients with the PLMA and in 94.4% with the LTS it was possible to maintain oxygenation, ventilation, and respiratory mechanics during the entire duration of surgery. Successful first and second attempt insertion rates were 57 patients (76%) and 15 patients (20%), respectively, for the PLMA and 60 patients (80%) and 11 patients (14.6%), respectively, for the LTS. Airway placement was unsuccessful with the PLMA in three patients and with the LTS in four patients. Time to achieve an effective airway was 36 +/- 24 s with the PLMA versus 34 +/- 25 s with the LTS. Gastric tube insertion was possible in 97.3% of patients with the PLMA and in 96% with the LTS. CONCLUSIONS: With respect to both physiologic and clinical function, the PLMA and LTS are similar and either device can be used to establish a safe and effective airway in mechanically ventilated anesthetized adult patients. PMID- 15277914 TI - Signal verification of middle latency auditory evoked potentials by automated detection of the brainstem response. AB - BACKGROUND: The midlatency components of auditory evoked potentials (AEPs) are gradually suppressed with increasing concentrations of anesthetics. Thus, they have been proposed as a monitor of anesthetic depth. However, undetected malfunction or disconnection of headphones and undetected hearing loss also result in suppressed midlatency AEPs that in turn may be misinterpreted as signs of deep anesthesia. As the brainstem component of the AEP is minimally influenced by anesthetics, its presence or absence can be used to verify that the recorded signal is a true AEP rather than an artifact. In this study, an online-capable procedure for detection of the brainstem component of the AEP was developed. METHODS: One hundred and ninety perioperatively recorded AEPs (binaural stimuli, 500 sweeps) were selected from a database with electroencephalographic and concomitant AEP stimulus information. Identical electroencephalogram regions were used to produce nonstimulus synchronized averaged signals (500 sweeps, "non AEP"). The 190 AEPs and 190 "non-AEPs" were used to develop a detector of the brainstem component of AEPs. AEPs and "non-AEPs" were wavelet transformed (discrete wavelet decomposition, biorthogonal 2.2 mother-wavelet), and the coefficient with the best separation of the two classes of signals was selected. Receiver operating characteristic curve analysis was performed to determine the optimum threshold value for this coefficient. RESULTS: The third coefficient of the third level was selected. In AEP signals, retransform of this coefficient produces a peak that resembles peak V of the brainstem response. The developed detector of the brainstem component of AEP had a sensitivity of 97.90% and a specificity of 99.48%. CONCLUSIONS: This detector of the AEP brainstem component can be used to verify that the signal reflects the response to an auditory stimulus. An alternative approach, used in the Danmeter AEP monitor, is based on the signal-to-noise ratio of the midlatency components of the AEP. Because the midlatency components of AEP are suppressed by anesthesia, a false alarm "low AEP/no AEP" is generated during deep anesthesia. This, in turn, may suggest disconnection of headphones or technical problems whenever anesthesia is deep. This disadvantage has been overcome by our detector, which is based on the identification of the brainstem component of AEP. PMID- 15277915 TI - Hemofiltration but not steroids results in earlier tracheal extubation following cardiopulmonary bypass: a prospective, randomized double-blind trial. AB - BACKGROUND: Activation of the inflammatory cascade is thought to account for some of the respiratory dysfunction and prolonged mechanical ventilation associated with cardiopulmonary bypass. The objective of this investigation was to identify whether perioperative steroids or hemofiltration during cardiopulmonary bypass, by their attenuation of inflammation, would reduce duration of mechanical ventilation after cardiac surgery. METHODS: After Institutional Review Board approval and informed consent, 192 patients scheduled to undergo elective primary coronary artery bypass grafting or valvular replacement or repair were randomized in a double-blind prospective study into three groups. One group (Control) received saline at induction and at 6-h intervals for four doses. Another group (Hemofil) received saline and hemofiltration to obtain 27 ml/kg of hemofiltrate. The final group (Steroid) received 1 g methylprednisolone before anesthesia induction and then 4 mg of dexamethasone at 6-h intervals for four doses. All patients underwent normothermic cardiopulmonary bypass and received propofol for postoperative sedation. Separate two-sample comparisons were performed to compare each experimental group versus the control group using the Wilcoxon rank sum test for continuous variables and Fisher exact test for categorical variables. In all cases, two-tailed P values 70 kg) with a sex-based formula (size 4 for women and size 5 for men) for selecting the appropriate size of ProSeal laryngeal mask airway. METHODS: Two hundred thirty seven healthy, anesthetized, paralyzed adult patients (American Society of Anesthesiologists physical status I or II; age, 18-80 yr) were randomly allocated for weight- or sex-based size selection. An experienced user inserted the ProSeal laryngeal mask airway with the digital technique. The following were compared: ease of insertion, oropharyngeal leak pressure, ease of ventilation, gas exchange, location of gas leak, anatomic position, mucosal injury, and postoperative pharyngolaryngeal problems. Intraoperative and postoperative data collection were unblinded and blinded, respectively. RESULTS: Ease of insertion, anatomic position, gas exchange, mucosal injury, and postoperative pharyngolaryngeal problems were similar between groups. For the sex-based group, larger ProSeal laryngeal mask airways were selected more frequently (P < 0.0001), oropharyngeal leak pressure (P = 0.02) was higher, leak volume (P = 0.004) and leak fraction (P = 0.007) were lower, and oropharyngeal leaks (P = 0.03) were detected less frequently. CONCLUSION: Size selection for the ProSeal laryngeal mask airway is equally effective using the manufacturer's weight-based formula or the sex-based formula in healthy, anesthetized, paralyzed adult patients, but leakage of small volumes of air from the mouth occurs less frequently with the sex-based formula. PMID- 15277917 TI - Effects of anesthetic agents on focal adhesion kinase (pp125FAK) tyrosine phosphorylation in rat hippocampal slices. AB - BACKGROUND: Tyrosine protein kinase proteins exert a prominent control on signaling pathways and may couple rapid events, such as action potential and neurotransmitter release, to long-lasting changes in synaptic strength and survival. Whether anesthetics modulate tyrosine kinase activity remains unknown. The aim of the current study was therefore to examine the effects of intravenous and volatile anesthetics on the phosphorylation of focal adhesion kinase (ppFAK), a functionally important nonreceptor tyrosine kinase, in the rat hippocampus. METHODS: Phosphorylation of ppFAK was examined in hippocampal slices by immunoblotting with both antiphosphotyrosine and specific anti-ppFAK antibodies. Experiments were performed in the absence (control) or presence of various concentrations of pharmacologic or anesthetic agents or both. RESULTS: Clinically relevant concentrations of thiopental, propofol, etomidate, isoflurane, sevoflurane, and desflurane induced a concentration-related increase in tyrosine phosphorylation. In contrast, ketamine (up to 100 microm) and the nonimmobilizer F6 (1,2-dichlorohexafluorocyclobutane, 25 microm) did not significantly affect ppFAK phosphorylation. The anesthetic-induced increase in ppFAK phosphorylation was blocked by GF 109203X, RO 318220, and chelerythrin (100 microm), three structurally distinct inhibitors of protein kinase C and U 73122 (50 microm), an inhibitor of phospholipase C. The propofol- and isoflurane-induced increase in ppFAK phosphorylation was reversible and showed nonadditivity of effects with phorbol 12-myristate 13-acetate (an activator of protein kinase C, 0.1 microm). In contrast, ketamine (up to 100 microm), MK801 (10 microm, an N-methyl-d aspartate receptor antagonist), bicuculline (10 microm, a gamma-aminobutyric acid type A receptor antagonist), and dantrolene (30 microm, an inhibitor of the ryanodine receptor) were ineffective in blocking anesthetic-induced activation of tyrosine phosphorylation. CONCLUSION: Except for ketamine, anesthetic agents markedly increase tyrosine phosphorylation of ppFAK in the rat hippocampus, most likely via the phospholipase C-protein kinase C pathway, whereas the nonimmobilizer F6 does not. These results suggest that ppFAK represents a target for anesthetic action in the brain. PMID- 15277918 TI - Direct cardiac effects of coronary site-directed thiopental and its enantiomers: a comparison to propofol in conscious sheep. AB - BACKGROUND: Previous evidence from laboratory animal studies indicates that R thiopental has a greater margin of safety than either the more potent S thiopental or the clinically used rac-thiopental. Although thiopental can cause cardiovascular depression from direct myocardial effects as well as indirect central nervous system and peripheral effects, no studies have yet determined whether its myocardial effects are enantioselective. A lesser direct effect would provide further evidence supporting R-thiopental as a preferred single enantiomer replacement for rac-thiopental. METHODS: The direct myocardial effects of the thiopental enantiomers were compared to those of rac-thiopental and propofol, using a crossover design with small incremental doses infused over 3 min, on separate days, into the left coronary arteries of conscious sheep. Hemodynamic and electrocardiographic measurements were acquired, and serial blood samples were collected during the studies for drug analyses. RESULTS: All three forms of thiopental and propofol produced significant hemodynamic effects consisting of dose-related and rapid-onset decreases in left ventricular dP/dtmax and stroke volume, and increases in left coronary blood flow and heart rate. Cardiac output, mean arterial blood pressure, and central venous pressure remained unaltered. The effects did not differ significantly among rac-thiopental, enantiopure R- or S thiopental, or propofol. Arterial blood drug concentrations were consistently less than those associated with systemic effects. CONCLUSIONS: Although previous evidence indicates that R-thiopental could make a suitable single-enantiomer replacement for rac-thiopental, the current study did not find a significant difference in direct cardiac effects among the thiopental enantiomers, racemate, or propofol. PMID- 15277919 TI - Mitochondrial complex I function affects halothane sensitivity in Caenorhabditis elegans. AB - BACKGROUND: : The gene gas-1 encodes a subunit of complex I of the mitochondrial electron transport chain in Caenorhabditis elegans. A mutation in gas-1 profoundly increases sensitivity of C. elegans to volatile anesthetics. It is unclear which aspects of mitochondrial function account for the hypersensitivity of the mutant. METHODS: : Oxidative phosphorylation was determined by measuring mitochondrial oxygen consumption using electron donors specific for either complex I or complex II. Adenosine triphosphate concentrations were determined by measuring luciferase activity. Oxidative damage to mitochondrial proteins was identified using specific antibodies. RESULTS: : Halothane inhibited oxidative phosphorylation in isolated wild-type mitochondria within a concentration range that immobilizes intact worms. At equal halothane concentrations, complex I activity but not complex II activity was lower in mitochondria from mutant (gas 1) animals than from wild-type (N2) animals. The halothane concentrations needed to immobilize 50% of N2 or gas-1 animals, respectively, did not reduce oxidative phosphorylation to identical rates in the two strains. In air, adenosine triphosphate concentrations were similar for N2 and gas-1 but were decreased in the presence of halothane only in gas-1 animals. Oxygen tension changed the sensitivity of both strains to halothane. When nematodes were raised in room air, oxidative damage to mitochondrial proteins was increased in the mutant animal compared with the wild type. CONCLUSIONS: : Rates of oxidative phosphorylation and changes in adenosine triphosphate concentrations by themselves do not control anesthetic-induced immobility of wild-type C. elegans. However, they may contribute to the increased sensitivity to volatile anesthetics of the gas-1 mutant. Oxidative damage to proteins may be an important contributor to sensitivity to volatile anesthetics in C. elegans. PMID- 15277920 TI - Effects of volatile anesthetics on store-operated Ca(2+) influx in airway smooth muscle. AB - BACKGROUND: In airway smooth muscle (ASM), volatile anesthetics deplete sarcoplasmic reticulum (SR) Ca(2+) stores by increasing Ca(2+) "leak." Accordingly, SR replenishment becomes dependent on Ca(2+) influx. Depletion of SR Ca(2+) stores triggers Ca(2+) influx via specific plasma membrane channels, store operated Ca(2+) channels (SOCC). We hypothesized that anesthetics inhibit SOCC triggered by increased SR Ca(2+) "leak," preventing SR replenishment and enhancing ASM relaxation. METHODS: In porcine ASM cells, SR Ca was depleted by cyclopiazonic acid or caffeine in 0 extracellular Ca(2+), nifedipine and KCl (preventing Ca(2+) influx through L-type and SOCC channels). Extracellular Ca(2+) was rapidly introduced to selectively activate SOCC. After SOCC activation, SR was replenished and the protocol repeated in the presence of 1 or 2 minimum alveolar concentration halothane, isoflurane, or sevoflurane. In other cells, characteristics of SOCC and interactions between acetylcholine (Ach) and volatile anesthetics were examined. RESULTS: Cyclopiazonic acid produced slow SR leak, whereas the caffeine response was transient in ASM cells. Reintroduction of extracellular Ca(2+) rapidly increased [Ca(2+)]i. This influx was insensitive to nifedipine, SKF-96365, and KBR-7943, inhibited by Ni and blockade of inositol 1,4,5-triphosphate-induced SR Ca(2+) release, and enhanced by ACh. Preexposure to 1 or 2 minimum alveolar concentration halothane completely inhibited Ca(2+) influx when extracellular Ca(2+) was reintroduced, whereas isoflurane and sevoflurane produced less inhibition. Only halothane and isoflurane inhibited ACh induced augmentation of Ca(2+) influx. CONCLUSION: Volatile anesthetics inhibit a Ni/La-sensitive store-operated Ca(2+) influx mechanism in porcine ASM cells, which likely helps maintain anesthetic-induced bronchodilation. PMID- 15277921 TI - Protein kinase C-epsilon primes the cardiac sarcolemmal adenosine triphosphate sensitive potassium channel to modulation by isoflurane. AB - BACKGROUND: Cardioprotection by volatile anesthetic-induced preconditioning is known to involve intracellular signaling pathways. Recent studies have shown that protein kinase C (PKC) plays an important role in anesthetic-induced preconditioning. In this study, the effects of the activation of specific isozymes of PKC, specifically PKC-epsilon and -delta, on the modulation of the sarcolemmal adenosine triphosphate-sensitive potassium (sarcKATP) channel by isoflurane were investigated. METHODS: The sarcKATP current was measured in ventricular myocytes isolated from guinea pig hearts using the whole cell configuration of the patch clamp technique. Peptides that induced the translocation of specific PKC isozymes were used to activate PKC-epsilon and PKC delta. RESULTS: Under whole cell conditions, isoflurane alone was unable to elicit the opening of the sarcKATP channel. Pretreatment with the specific PKC epsilon activator, PP106, primed the sarcKATP channel to open in the presence of isoflurane. The resulting sarcKATP current densities in the presence of 0.88 mm isoflurane were 6.5 +/- 6.0 pA/pF (n = 7) and 40.4 +/- 18.2 pA/pF (n = 7) after pretreatment with 100 and 200 nm PP106, respectively. The PKC-epsilon antagonist PP93 abolished this effect. A scrambled peptide of the PKC-epsilon activator PP105 did not prime the sarcKATP channel. The PKC-delta activator PP114 was significantly less effective in priming the sarcKATP channel. 5-Hydroxydecanoate significantly attenuated the effect of the PKC-epdsilon activator on the sarcKATP channel. In addition, immunohistochemical analysis showed that the PKC-epsilon isoform translocated to both the mitochondria and sarcolemma after anesthetic induced preconditioning, whereas the PKC-delta isoform translocated to the mitochondria. CONCLUSION: The PKC-epsilon isozyme primed the sarcKATP channel to open in the presence of isoflurane. The PKC-delta isozyme was significantly less effective in modulating the isoflurane effect on this channel. PMID- 15277922 TI - Molecular mechanisms of the inhibitory effects of bupivacaine, levobupivacaine, and ropivacaine on sarcolemmal adenosine triphosphate-sensitive potassium channels in the cardiovascular system. AB - BACKGROUND: Sarcolemmal adenosine triphosphate-sensitive potassium (KATP) channels in the cardiovascular system may be involved in bupivacaine-induced cardiovascular toxicity. The authors investigated the effects of local anesthetics on the activity of reconstituted KATP channels encoded by inwardly rectifying potassium channel (Kir6.0) and sulfonylurea receptor (SUR) subunits. METHODS: The authors used an inside-out patch clamp configuration to investigate the effects of bupivacaine, levobupivacaine, and ropivacaine on the activity of reconstituted KATP channels expressed in COS-7 cells and containing wild-type, mutant, or chimeric SURs. RESULTS: Bupivacaine inhibited the activities of cardiac KATP channels (IC50 = 52 microm) stereoselectively (levobupivacaine, IC50 = 168 microm; ropivacaine, IC50 = 249 microm). Local anesthetics also inhibited the activities of channels formed by the truncated isoform of Kir6.2 (Kir6.2 delta C36) stereoselectively. Mutations in the cytosolic end of the second transmembrane domain of Kir6.2 markedly decreased both the local anesthetics' affinity and stereoselectivity. The local anesthetics blocked cardiac KATP channels with approximately eightfold higher potency than vascular KATP channels; the potency depended on the SUR subtype. The 42 amino acid residues at the C terminal tail of SUR2A, but not SUR1 or SUR2B, enhanced the inhibitory effect of bupivacaine on the Kir6.0 subunit. CONCLUSIONS: Inhibitory effects of local anesthetics on KATP channels in the cardiovascular system are (1) stereoselective: bupivacaine was more potent than levobupivacaine and ropivacaine; and (2) tissue specific: local anesthetics blocked cardiac KATP channels more potently than vascular KATP channels, via the intracellular pore mouth of the Kir6.0 subunit and the 42 amino acids at the C-terminal tail of the SUR2A subunit, respectively. PMID- 15277923 TI - Hemorrhagic shock in swine: nitric oxide and potassium sensitive adenosine triphosphate channel activation. AB - BACKGROUND: To determine the role of nitric oxide and adenosine triphosphate sensitive potassium (KATP) vascular channels in vascular decompensation during controlled hemorrhagic shock in swine. METHODS: Thirty instrumented, anesthetized adolescent Yorkshire swine were subjected to controlled isobaric hemorrhage to a mean arterial pressure of 40 mmHg for 2 h (n = 6) or 4 h (n = 10) or 50 mmHg for 4 h (n = 8). An additional six animals were used as anesthetized instrumented time controls. During controlled hemorrhage, plasma and tissue samples were obtained every 30 to 60 min. Before euthanasia, tissue (carotid artery, lung, liver, and aorta) was obtained for analysis of nitrate concentrations and nitric oxide synthase activity. Isolated carotid artery ring reactivity to norepinephrine was also determined with and without glibenclamide. RESULTS: Animals hemorrhaged to 40 mmHg decompensated earlier than animals hemorrhaged to 50 mmHg. Plasma nitrate concentrations and nitric oxide synthase activity rose consistently throughout hemorrhage in both groups. However, they were substantially higher in the mean arterial pressure 40 group. Constitutive nitric oxide synthase activity was the major contributor to total nitric oxide synthase activity throughout the protocol with only the animals maintained at 40 mmHg for 4 h showing evidence of inducible nitric oxide synthase activity. Profound KATP channel activation and hyporeactivity of isolated vessel rings to norepinephrine was not observed until 4 h after the initiation of hemorrhagic shock. Only those animals with inducible nitric oxide synthase activity showed a decreased response to norepinephrine, and this hyporeactivity was reversed with the KATP channel inhibitor, glibenclamide. CONCLUSIONS: The data indicate that profound KATP activation associated with increased nitric oxide concentrations and inducible nitric oxide synthase induction is a key factor in vascular smooth muscle hyporeactivity characteristic of the late decompensatory phase of hemorrhagic shock in swine. PMID- 15277924 TI - Temperature-independent Inhibition of L-type calcium currents by halothane and sevoflurane in human atrial cardiomyocytes. AB - BACKGROUND: Cardiac L-type calcium currents (ICa,L) are affected by volatile anesthetics, possibly contributing to their side effects. Actions of anesthetics on ion channels are usually studied in vitro at room temperature. However, the solubility of anesthetic gases as well as ICa,L are markedly sensitive to the study temperature. Therefore, temperature-dependent effects of halothane and sevoflurane on cardiac ICa,L were analyzed. METHODS: ICa,L were studied at 21 degrees C and 36 degrees C with the patch clamp technique in isolated human atrial cardiomyocytes. Concentrations of anesthetics brought into solution by gassing at both temperatures were determined with gas chromatography. RESULTS: The aqueous concentrations of halothane and sevoflurane were linearly related to their concentration in the gas phase (1 to 3 minimum alveolar concentration [MAC]). At 21 degrees C, the slope of this relation was 0.52 and 0.12 mm/vol % for halothane and sevoflurane, respectively, and decreased at 36 degrees C to 0.29 and 0.09 mm/vol %, respectively. ICa,L displayed significantly higher current amplitudes at 36 degrees C than at 21 degrees C and significantly accelerated time-dependent inactivation. Halothane (1-2 MAC) and sevoflurane (1-3 MAC) evoked stronger inhibitions of ICa,L at 21 degrees C than at 36 degrees C. In spite of different temperature-dependent current amplitudes, the fractional (percent) inhibition of ICa,L showed the same linear relationship to the concentrations of halothane and sevoflurane in the bath medium at both temperatures, as revealed from present and previous experiments. CONCLUSIONS: Inhibition of ICa,L by halothane and sevoflurane is determined by the aqueous concentration of the anesthetics, independently of the temperature. Increased solubility may explain the stronger effects of the anesthetics at lower temperatures. PMID- 15277925 TI - Relative amnesic potency of five inhalational anesthetics follows the Meyer Overton rule. AB - BACKGROUND: Doses of volatile anesthetics around 0.3 minimum alveolar concentration (MAC) inhibit learning. However, threshold amnesic doses and relative potencies between agents are not well established. The authors determined amnesic potency in rats for four common volatiles and nitrous oxide. METHODS: After institutional review board approval, adult Sprague-Dawley rats received inhibitory avoidance training during exposure to either air or various subanesthetic doses of desflurane, sevoflurane, isoflurane, halothane, or nitrous oxide (4-21 rats/dose). Animals were trained to remain in a starting "safe" compartment for 100 consecutive seconds by administering a foot shock (0.3 mA) each time they entered an adjacent "shock" compartment. Memory was assessed at 24 h. Anesthetic effects on pain thresholds were separately determined. RESULTS: Learning: Only relatively higher doses of sevoflurane, halothane, and desflurane increased the number of shocks required for task acquisition. Memory: Significantly decreased retention performance (P < 0.05) was found at relatively low inspired concentrations of 0.2% isoflurane, 0.3% sevoflurane and halothane, 0.44% desflurane, and 20% nitrous oxide. Amnesic potency was nitrous oxide >/= desflurane > sevoflurane >/= isoflurane >> halothane, (rank-ordered ED50 values as %MAC). Amnesic potency correlated with oil:gas partition coefficients (r = 0.956, P < 0.007). Halothane, only at 0.08%, enhanced retention (P < 0.01). All agents were analgesic at higher doses. CONCLUSIONS: Amnesic potency differs between agents; nitrous oxide is most potent and halothane is least potent relative to MAC. The amnesic threshold ranges from 0.06 to 0.3 MAC. The correlation between potency and oil:gas partition coefficients suggests a fundamental role for hydrophobicity in mediating amnesia, similar to its association with MAC. Some agents (e.g., halothane) may enhance aversive memory retention at doses typically encountered during emergence. PMID- 15277926 TI - Retigabine stimulates human KCNQ2/Q3 channels in the presence of bupivacaine. AB - BACKGROUND: Inhibition of KCNQ2/Q3 channels may cause convulsion in humans. The interaction of bupivacaine with these channels is unknown. The anticonvulsant retigabine activates KCNQ2/Q3 channels and may reverse inhibitory actions of bupivacaine. Potassium channel stimulation may thus constitute a novel approach to treat local anesthetic-induced seizures. The aim of this study was to characterize bupivacaine effects on KCNQ2/Q3 channels and to investigate whether retigabine reverses the effects of the local anesthetic. METHODS: KCNQ2/Q3 channels were transiently expressed in Chinese hamster ovary cells. The effects of bupivacaine and retigabine were studied with the patch-clamp technique. RESULTS: Bupivacaine inhibited KCNQ2/Q3 channels in a concentration-dependent and reversible manner. The concentration-response curve was described by a Hill equation (IC50 = 173 +/- 7 microm, Hill coefficient = 1.4 +/- 0.1, mean +/- SEM, n = 37). The inhibitory effect did not differ between bupivacaine and levobupivacaine (42 +/- 4%, n = 7, versus 42 +/- 5%, n = 10; P > 0.05). Ropivacaine was four times less potent than bupivacaine. The inhibition of KCNQ2/Q3 channels by bupivacaine resulted in a significant and reversible depolarization of the membrane potential. Retigabine (300 nm-10 microm) reversed the inhibitory action of bupivacaine on KCNQ2/Q3 channels as well as the depolarization of the membrane potential. CONCLUSIONS: The anticonvulsant retigabine at nanomolar concentrations reverses the inhibitory effect of micromolar concentrations of bupivacaine. Our results allow the hypothesis that activation of KCNQ2/Q3 channels by retigabine may offer a novel therapeutic approach for the treatment of bupivacaine-induced seizures. PMID- 15277927 TI - Epidural neostigmine combined with sufentanil provides balanced and selective analgesia in early labor. AB - BACKGROUND: This study evaluated the efficacy of an epidural single dose of neostigmine combined with sufentanil to provide selective and balanced analgesia at the beginning of labor. METHODS: After informed consent, 125 healthy parturients were randomly allocated to receive, after a test dose, a single injection of either epidural sufentanil 20 micrograms (minimal analgesic dose) or 10 micrograms or a combination of sufentanil 10 micrograms with neostigmine 250, 500, or 750 micrograms in a total volume of 12 ml. Pain scores were recorded at regular intervals to determine onset and duration of analgesia. Maternal and fetal vital parameters as well as side effects were closely monitored. RESULTS: Parturients did not differ concerning demographic data. Epidural neostigmine 500 micrograms with sufentanil 10 micrograms produced effective analgesia (visual analog scale <30 mm within 10 min in 72% parturients and within 15 min in 85% parturients; average duration of 119 min, confidence interval 96-142 min) that was as effective as epidural sufentanil 20 micrograms. Epidural combination with neostigmine 250 micrograms was ineffective, whereas 750 micrograms did not produce higher effect than 500 micrograms. No motor block was recorded. Maternal and fetal vital parameters remained stable during labor. CONCLUSIONS: Epidural combination of neostigmine 500 micrograms (e.g., 6-7 micrograms/kg) with sufentanil 10 micrograms provides similar duration of analgesia as epidural sufentanil 20 micrograms and allows effective and selective analgesia devoid of side effects in the first stage of labor. PMID- 15277928 TI - Lumbar plexus in children. A sonographic study and its relevance to pediatric regional anesthesia. AB - BACKGROUND: Pediatric regional anesthesia has gained increasing interest over the past decades. The current study was conducted to investigate the lumbar paravertebral region and the lumbar plexus at L3-L4 and L4-L5 by means of sonography to obtain fundamentals for the performance of ultrasound-guided posterior lumbar plexus blocks. METHODS: Thirty-two children (12 boys, 20 girls) with American Society of Anesthesiologists physical status I or II were enrolled in the current study. The lumbar paravertebral region was visualized at L3-L4 and L4-L5 on two corresponding posterior sonograms (longitudinal, transverse). The lumbar plexus had to be delineated, and skin-plexus distances were measured. In a series of five pediatric patients undergoing inguinal herniotomy, ultrasound guided posterior lumbar plexus blocks at L4-L5 were performed. RESULTS: The children were stratified into three age groups (group 1: > 3 yr and 5 yr and 8 yr and 5.7 points on the Action Research Arm test) was 27% in group B (four patients) and 8% in group A (one patient). The differences in functional gain and success rate were not statistically significant, neither were the differences between the two groups on the secondary outcome measures. CONCLUSION: The difference between the two stimulation strategies was not statistically significant. PMID- 15277961 TI - Effects of respiratory muscle weakness on daily living function, quality of life, activity levels, and exercise capacity in mild to moderate Parkinson's disease. AB - OBJECTIVE: To investigate respiratory muscle strength in mild to moderate Parkinson's disease in comparison with prediction equations and age-matched controls. To identify correlations between respiratory muscle strength and daily living function, activity levels, quality of life, and exercise capacity. DESIGN: A total of 66 participants with Parkinson's disease and 32 age-matched, healthy controls participated. Respiratory mouth pressures (representing respiratory muscle strength) were compared with predicted values. A comparison was also made between participants with Parkinson's disease and age-matched controls. Respiratory mouth pressures were correlated with results of the Barthel index (daily living function), a modified Baecke activity questionnaire (activity levels), the Parkinson's disease questionnaire (quality of life in Parkinson's disease), peak heart rate, peak oxygen consumption, blood lactate thresholds, and the number of stages completed during an incremental cycle ergometer test (representing exercise capacity). RESULTS: Respiratory mouth pressures were significantly lower (P < 0.05) in the Parkinson's disease group, but this did not influence the measures of daily living function, activity levels, and quality of life. Respiratory mouth pressures correlated with lactate thresholds (r = 0.308, P < 0.01) and the number of completed stages of the cycle ergometer test (r = 0.490, P < 0.01). CONCLUSIONS: In mild to moderate Parkinson's disease, there is a significant weakness of the respiratory muscles. This can affect the individual during exercise but has no apparent effect on activities that do not require a large effort of the respiratory muscles. PMID- 15277962 TI - Comparison of peak cough flows measured by pneumotachograph and a portable peak flow meter. AB - OBJECTIVE: To compare peak cough flows (PCF) obtained with a pneumotachograph (PCFp) with those measured using a portable peak flow meter (PCFm) in a population of healthy subjects and patients with neuromuscular disease. DESIGN: A total of 30 healthy subjects and 32 medically stable patients with neuromuscular diseases were studied. Using an oronasal mask connected, in a randomized order, to a pneumotachograph and to a portable peak flow meter, PCFp and PCFm were measured as every subject performed maximal cough efforts from total lung capacity. RESULTS: PCFp measurements were 377.70 +/-179.28 liters/min and PCFm measurements were 377.50 +/- 172.98 liters/min (not significant). The two measurements were correlated (r = 0.95, P < 0.001). The random error obtained was 9.37%. The mean of the differences between the values obtained using both instruments +/- limits of agreement (+2 SD) was 0.20 +/- 109.78 liters/min (not significant). When the agreement was tested over three ranges of flows (< 270 liters/min, 270-480 liters/min, and >480 liters/min), no statistically significant differences were obtained for the population as a whole in any range; however, in the lower flow range, 14 patients had PCFm significantly greater than PCFp (213.93 +/- 55.92 vs. 180.86 +/- 57.22, P = <0.01). CONCLUSION: PCF measurements made with a portable peak flow meter such as the asmaPLAN+ (Vitalograph, Ennis, Ireland) are reproducible and reasonably accurate when flows are >270 liters/min. However, some caution must be exercised in clinical practice at lower PCF because of the tendency of this portable device to overestimate the lower flows. PMID- 15277963 TI - Effort-limited treadmill walk test: reliability and validity in subjects with postpolio syndrome. AB - OBJECTIVE: To determine the reliability and construct validity of an effort limited treadmill walk test to measure functional ability in subjects with postpolio syndrome in an outpatient postpolio clinic. DESIGN: Functioning and distance walked on a treadmill to a Borg "hard" effort level were measured three times, a week apart, by two blinded raters in 15 subjects with postpolio syndrome, aged 37-67 yrs, with new weakness, fatigue, and pain but with no other cause of symptomatology or condition-limiting walking. One rater tested them twice. Fatigue activity level, mobility, and health-related quality of life (Medical Outcome Study Short Form Health Survey [SF-36]) defined functioning. Generalizability correlation coefficients determined intrarater, test-retest and interrater reliability. The correlations relating the distance walked and functioning determined construct validity. RESULTS: Reliability for generalizability correlation coefficients were: intrarater, 0.91; test-retest, 0.85; and interrater, 0.58. Interrater reliability improved to 0.91 with adherence to a standardized protocol. Validity was established with correlations between the distance walked and SF-36 physical component score (0.66), physical role (0.60), bodily pain (0.60), and vitality (0.55). CONCLUSIONS: The treadmill walk test provides a reproducible and valid measure of ability in persons with postpolio syndrome with a single rater, but a standardized protocol is essential for reliability. PMID- 15277964 TI - Effect of rehabilitation on hip and knee proprioception in older adults after hip fracture: a pilot study. AB - OBJECTIVE: Impaired proprioception may predispose patients with hip fracture to increased risk of future disability. The purpose of the study was to determine the effect of rehabilitation on proprioceptive changes in both the hip and knee joints of patients after hip fracture. DESIGN: Data were collected on 30 patients with hip fracture (mean age, 79.6 +/- 6.7 yrs) who attended physical and occupational therapy sessions five times per week during a rehabilitation hospital stay of 24.8 +/- 8.1 days. Proprioception was assessed with an electrogoniometer within 48 hrs of admission to and discharge from the rehabilitation unit. The passive-to-active reproduction of joint angle technique determined absolute angular error in non-weight-bearing positions at 15, 30, and 60 degrees of hip flexion and knee extension in both injured and noninjured sides. RESULTS: Absolute angular error decreased significantly (P < 0.05) from admission (5.3 +/- 2.6 degrees, 4.1+/- 3.1 degrees) to discharge (3.0 +/- 2.3 degrees, 2.8 +/- 3.1 degrees) in hip flexion and knee extension, respectively, on the injured side. Absolute angular error was significantly less (P < 0.05) at 15 degrees compared with 30 and 60 degrees of hip flexion at admission and discharge on the injured side. CONCLUSIONS: Hip and knee joint proprioception significantly improved in the injured side after the rehabilitation program. This may be an important outcome regarding future disability in this population. PMID- 15277965 TI - Functional recovery and length of stay after hip fracture in patients taking corticosteroids. AB - OBJECTIVE: To evaluate the functional recovery and the length of stay after hip fracture in patients receiving corticosteroids. DESIGN: A total of 796 inpatients with hip fracture consecutively admitted to our rehabilitation hospital were included in this retrospective study. A total of 36 of 796 were currently treated with either oral (n = 23) or inhaled (n = 13) corticosteroids. RESULTS: No significant differences were shown between corticosteroid users and controls for Barthel index score at admission or discharge, change in Barthel index score resulting from rehabilitation, and length of stay. Multiple regression, including 11 confounding variables, showed that several factors, but not the treatment with corticosteroids, were significantly associated with the Barthel index score or the length of stay. The results were similar when the two subgroups of patients receiving corticosteroids were evaluated separately. In the subgroup of the patients receiving oral corticosteroids, no meaningful correlations were observed between the daily dose (milligrams of prednisone equivalent) and the Barthel index score, the change in the Barthel index score attributable to rehabilitation, or the length of stay. CONCLUSIONS: After hip fracture, neither the functional recovery nor the length of stay were significantly affected by the current treatment with corticosteroids. PMID- 15277966 TI - Effect of wheelchair stroke pattern on mechanical efficiency. AB - OBJECTIVE: To investigate the effect of different wheelchair stroke patterns on efficiency and propulsion technique (force application and timing). DESIGN: Inexperienced, able-bodied subjects were randomly divided into two velocity groups (1.11 m/sec [n = 13] and 1.39 m/sec [n = 11]). An external (medium) load was set at 0.23 N/kg. Subjects performed four 4-min exercise blocks on a wheelchair ergometer. The first block was performed with a freely chosen movement pattern of the hand. Thereafter, the pumping, semicircular, or single-looping over propulsion pattern were performed in a counterbalanced order. Gross mechanical efficiency and propulsion technique variables were measured with Oxycon Alpha and an instrumented wheelchair ergometer. RESULTS: A significant difference was found for mechanical efficiency, with pumping showing the highest efficiency and semicircular the lowest efficiency, regardless of velocity. Timing variables and negative power deflections before and after the push phase showed significant differences between the stroke patterns. CONCLUSIONS: Pumping is the energetically most efficient stroke pattern in contrast to the semicircular pattern in this subject group. Propulsion technique could not explain the difference in efficiency. PMID- 15277967 TI - Muscle strength in patients with unicompartmental arthroplasty. AB - OBJECTIVE: The aim of the current study was to evaluate the isokinetic strength of the knee in patients with unicondylar prostheses, to compare these results with healthy control subjects of the same age, and to correlate these results with clinical scores. DESIGN: Seventeen patients were examined an average of 21.5 mos after surgery. Clinical examination was done using the Hospital for Special Surgery, Knee Society, and patellar scores and a visual analog scale for pain. Quality of life was assessed by the Short Form 36 Health Questionnaire. Isokinetic evaluation of knee extensor and flexor muscles was done using a Cybex 6000 dynamometer at angular velocities of 60 and 180 degrees/sec. Eleven healthy subjects of comparable age served as a control group. RESULTS: Clinical results differed significantly in all categories. Quality of life differed only in the items of physical functioning, role limitation because of physical problems, and bodily pain. Isokinetic strength in patients showed a loss of torque of approximately 30% in extension and flexion at 60 and 180 degrees/sec compared with the control group. The flexion and extension ratio and the angles of maximum torque did not differ between the groups. CONCLUSION: In comparison with healthy control subjects, persons with an implanted unilateral sledge prosthesis have strength deficits in extension and flexion. A deficit of the extensor muscles, as it has been described previously for patients with a total knee arthroplasty, could not be found. PMID- 15277969 TI - Dynamic hip flexion contractures. PMID- 15277970 TI - Phantom breast pain as a source of functional loss. AB - Although physicians are aware of phantom limb pain, which can occur in up to 85% of patients who undergo amputation, and its potential effect on functional status, the presence of phantom pain after amputation of other body parts such as the breast and its effect on function may be less appreciated. We report the case of a 63-yr-old woman with multiple sclerosis who underwent a modified radical mastectomy for left intraductal breast carcinoma. After her mastectomy, she required a brief course of inpatient rehabilitation and was discharged from rehabilitation independent, with feeding, dressing, hygiene, and transfers. Two months after her mastectomy, she had difficulty with these tasks because of phantom breast pain. Accurate diagnosis of her pain and successful treatment resulted in a return to premorbid functional status. PMID- 15277972 TI - The young cervical spinal stenotic. PMID- 15277974 TI - [Jubile of Professor Philippe Vague, Marseille, France, May 2004]. PMID- 15277976 TI - Positive octreotide scintigraphy and determination of lanreotide activity in Paget's disease of bone associated with phosphate diabetes: a case report. AB - A patient with Paget's disease developed phosphate diabetes (phosphate: 1.6 mg/dl (2.5-4.4 mg/dl), with 29 ml/min phosphate clearance (Nl<15ml/min) and a 65% phosphate reabsorption rate (Nl>85%). As previously demonstrated in tumor-induced osteomalacia, we hypothesized that osteoblasts might manifest somatostatin receptor activity. The patient underwent an octreotide scan which demonstrated increased uptake localized in affected bone. Under lanreotide treatment (40 mg i.m.), the patient's bone pain improved with a concomitant decrease in phosphate alkaline level. Phosphate clearance and tubular readsorption rate of phosphate did not change significantly. We reviewed previously reported cases of associated Paget's bone disease and phosphate diabetes. PMID- 15277975 TI - [Sexual dysfunction among 78 Moroccan male hemodialysis patients: clinical and endocrine study]. AB - PURPOSE: To investigate the correlation between biochemical and endocrine variables with sexual disorders in 78 male patients on chronic hemodialysis at the Ibn Rochd University Hospital (Casablanca-Morocco). METHODS: Seventy-eight male hemodialysis patients with chronic renal failure were evaluated with regard to their sexual function. All patients answered a personal questionnaire on their sexual activity. Hormone (FSH, LH, prolactin, testosterone, and parathyroid hormone) and zinc and ferritin assays were also performed. RESULTS: The men reported erection (44.9%), libido (44.9%), ejaculation (26.8%), and orgasm (21.8%) disorders. Gynecomastia was observed in 17.9% of the patients. There was no correlation with weight nor the nature of the causal nephropathy nor with duration of dialysis. Levels of gonadotropins (FSH, LH), prolactin, and parathyroid hormone were elevated. Testosterone levels were low. Ferritinemia was elevated but there was no significant variation in zincemia. There was a negative curvilinear relationship between serum testosterone and sexual disorders, and between gynecomastia and ferritinemia. LH and prolactin levels were positively correlated with gynecomastia. CONCLUSION: Abnormal hormonal and iron overload could be important factors involved in the complex pathogenesis of sexual dysfunction in chronic renal failure patients undergoning hemodialysis. PMID- 15277977 TI - [Intrathyroid metastasis: 11 cases]. AB - INTRODUCTION: Non-thyroid cancers rarely metastasize into the thyroid gland. The aim of this retrospective study was to report a series of thyroid metastases and to emphasize their unusual occurrence and their poor prognosis. METHODS: Between January 1987 and June 1999 eleven patients underwent thyroidectomy for isolated, metastatic diseases of non thyroidal origin (mean age 61 yrs, 54.5% female). The primary tumors were: pulmonary squamous cell carcinoma (n=5), renal cell carcinomas (n=2), esophageal squamous cell carcinoma (n=1), leiomyosarcoma (n=1), oropharynx squamous cell carcinoma (n=1), and breast carcinoma (n=1). Analyzing these cases, there is a marked preponderance of lung cancers, renal cancer coming second in order of frequency. Clinical manifestations are: thyroid nodule without hormonal disturbance; others signs are dysphonia and/or dysphagia. RESULTS: Ten patients underwent preoperative fine-needle aspiration, nine of ten were suggestive of metastatic disease. The mean time from resection of the primary tumor to thyroid metastases was 25 months (range 1-96 months). Total thyroidectomy (n=9) or lobectomy (n=2) was performed without morbidity or mortality. No patients have had recurrent disease in the neck. Median survival after treatment was 10 months (range 1-29 months). Course of death were mainly disseminated metastases. CONCLUSION: For isolated metastatic cancer to the thyroid, surgical resection should be performed in order to avoid potential morbidity of tumor recurrence in the neck, even if the prognosis remains poor, for the majority of the cases. PMID- 15277978 TI - Imaging features of intrasellar tuberculoma: two cases. AB - Hypophyseal tuberculoma is extremely rare. It may be confused with other more common sellar tumors such as adenomas. Characteristic, but not specific, radiological features are in the majority of cases: intense enhancement on contrast CT and thickening of the pituitary stalk better visible on MRI. We describe imaging findings in two patients with pituitary tuberculosis. In these cases an accurate non-invasive diagnosis was found to be important as antituberculous chemotherapy is curative. PMID- 15277979 TI - [Etiologic and therapeutic aspects of acquired central diabetes insipidus]. AB - Acquired central diabetes insipidus (CDI) is a rare disease due to anatomic lesions of the hypothalamo-pituitary system. We discuss the etiologic and therapeutic aspects of CDI. Through 5 cases and a review of the literature. We report: Two cases of Langerhans histiocytosis, a 21 year old man and a 37 year old women. The CDI was the only endocrine manifestation in the man, but it was associated with panhypopituitarism and infiltration of the thyroid gland by histiocytosis cells in the women. One case of suprasellar germinoma, a 18 year old adolescent had hypocorticism, hypothyroidism, and hypogonadism associated with CDI and hyperprolactinemia. One case of pituitary cystic lesion with extension to the suprasellar area, a 36 year old women presenting with hypocorticism, growth hormone deficiency, and hyperprolactinemia. One case of neuro-Behcet's disease in a 47 year old man. He presented with strokes and CDI. When the CDI is isolated Langerhans Histiocytosis should be considered first. The exploration of the hypothalamo-pituitary region using MRI showed different anatomic lesions in patients with CDI. It is very difficult to determine definitive diagnosis before surgery in the cases of cystic lesions. PMID- 15277980 TI - [Intrathyroidal parathyroid adenoma. Study of a case]. AB - Intrathyroidal parathyroid adenoma is an infrequent lesion which can be explained by abnormalities during embryonic migration of the parathyroid glands. That abnormality can be a cause of failed cervical exploration for primary hyperparathyroidism. From a case report and literature data, we propose an exploration processes to search those ectopic parathyroid adenoma. PMID- 15277981 TI - [Severe primary hyperparathyroidism and vitamin D deficiency]. AB - Primary hyperparathyrodism is a common disease, often asymptomatic. A young post partum woman was hospitalized for functional impotence of the upper right limb and poor health status. Laboratory tests revealed severe primary hyperparathyroidism (osteitis fibrosa cystica and nephrolithiasis) associated with vitamin D deficiency. Technetium 99m and thallium parathyroid scintigraphy showed increased uptake under the left thyroid lobe. After surgical resection of a parathyroid adenoma, serum calcium fell markedly and parathyroid levels declined but remained above normal. Calcium and 25(OH)-vitamin D supplementation led to normal calcium and vitamin D levels in three Months, with marked improvement in the bone lesions visualized on the six-Month x-rays. Based on this observation, we describe the many radiological aspects of bone involvement in long-standing hyperparathyroidism aggravated by vitamin D deficiency. Early diagnosis of hyperparathyroidism is crucial. PMID- 15277982 TI - [Ectopic thyroid: two cases]. AB - Ectopic thyroid is a rare condition (1/4000 to 1/8000 among patients with hypothyroidism). The underlying etiological pathogenic mechanisms remain unknown. Diagnosis is established on the basis of imaging findings. We report two cases of hypothyroidism in adult females who had ectopic sublingual thyroid glands. The first patient was a 20-Year-old woman who had been treated for hypothyroidism since the age of 13 Years before the diagnosis of ectopic thyroid 7 Years later. In both patients, the thyroid gland was palpable. In the first patient the physical examination revealed an ectopic sublingual gland. Scintigraphy confirmed the diagnosis in both patients. The CT-scan and MRI were positive in the second patient. Hormonal substitution therapy using L-thyroixine was given. PMID- 15277983 TI - Involvement of interleukin-18 in the inflammatory response against oropharyngeal candidiasis. AB - BACKGROUND: Oral candidiasis is a collective name for a group of disorders caused by the dimorphic fungus Candida albicans (C. albicans). Host defenses against C. albicans essentially fall into two categories: specific immune mechanisms and local oral mucosal epithelial cell defenses. The rationale of this study was to investigate the involvement of IL-18 in the inflammatory response against oral candidiasis. MATERIAL/METHODS: We first used human oral mucosa tissue and saliva to assess the production of Il-18. Second, we engineered human oral mucosa using only normal human oral epithelial cells and fibroblasts. Tissues were infected with C. albicans at different time points. RESULTS: Tissue and saliva analyses demonstrated that constitutively produced and secreted IL-18 was up-regulated following Candida-infection. With our engineered model, we showed that C. albicans significantly increased the secretion of active IL-18 by infected epithelial cells. Interestingly, a significant secretion of IFNg functionally supported the up-regulation of active IL-18 in C. albicans-infected tissues. We also showed that rhIL-18 increased the expression and production of endogenous IL 18 and ICE in C. albicans-infected tissues, which was paralleled by a significant increase in IFNg secretion. CONCLUSIONS: These data suggest that (i) oral epithelial cells are involved in local host defenses against C. albicans infections, via IFNg induced-IL-18, and (ii) that IL-18 and IFNg secretions may be related to epithelial cells. Given that our experimental model closely mimics the natural interface between the oral mucosa and C. albicans, it appears that IL 18 meets the requirements of being a cytokine that epithelial cells use to control C. albicans infections. PMID- 15277984 TI - Somatic mutations in peroxisome proliferator-activated receptor-gamma are rare events in human cancer cells. AB - BACKGROUND: Peroxisome proliferator-activated receptor-gamma (PPAR-gamma) is a nuclear hormone receptor activated after binding a lipophilic ligand, such as naturally occurring 15dPGJ2. There is striking evidence that PPAR-gamma activation leads not only to an increase in insulin sensitivity, but also to tumor cell apoptosis and cell cycle arrest. A growing number of diabetes mellitus II patients currently benefit from treatment with synthetic PPAR-gamma agonists, the thiazolidinediones (TZDs), which sensitize peripheral cells towards insulin. Furthermore, some TZDs are undergoing clinical investigations for the treatment of malignant diseases. Therefore, detailed information on the frequency of genetic alterations of PPAR-gamma in malignant tumor cells is necessary. MATERIAL/METHODS: PPAR-gamma DNA of 33 histologically different tumor cells was isolated, purified, and all coding regions were separately amplified by PCR. The coding exons were then analyzed by single-stranded conformational polymorphism (SSCP) and bidirectional DNA sequencing. RESULTS: In five breast cancer brain metastasis samples from patients and 28 cancer cell lines derived from lymphoma, glioblastoma, and breast carcinoma we found only one coding region shift in exon 5b of the glioblastoma U373 DNA. This silent mutation does not lead to a change in amino acid alignment. No further polymorphisms, including those which have already been described, could be detected in any other sample. CONCLUSIONS: We conclude that somatic mutations in the PPAR gene are exceedingly rare events in malignant tumor cells. This makes PPAR-gamma more unlikely to act as a tumor suppressor gene, making it a stable and suitable target for TZD biological cancer therapy. PMID- 15277985 TI - Growth hormone enhances gastric ulcer healing in rats. AB - BACKGROUND: Gastric ulcer healing requires the reconstitution of epithelial structures and underlying connective tissue through cellular proliferation, migration, and differentiation. The systemic application of growth hormone (GH) has shown anabolic effects in postoperative and burn therapy by increasing protein synthesis and attenuating protein catabolism. There is also evidence that GH stimulates cell proliferation and differentiation. In this study we evaluated the impact of GH on gastric ulcer healing. MATERIAL/METHODS: Gastric ulcers were induced with a cryoprobe in male Wistar rats (285+/-11 g). The first group of rodents (n=10) received a daily subcutaneous dose of 2.5 mg/kg growth hormone for seven days. The second group (n=10) received only vehicle. After 7 days, ulcer size was determined photoplanimetrically. Cell proliferation and new vessel growth in the ulcer margin were evaluated by quantitative immunohistochemical staining for proliferating cell nuclear antigen (PCNA) and smooth muscle actin (SMA), respectively. RESULTS: The systemic application of GH caused a significant increase in body weight (332+/-7 g vs. 289+/-13 g; p=0.007). Ulcer size was also reduced significantly compared with controls (5.6+/-0.8 mm2 vs. 9.9+/-1 mm2; p=0.005). Immunohistochemical analysis revealed a significant increase in cell proliferation (79.7+/-0.9% pos. cells vs. 64.7+/-1.9% pos. cells; p=0.0001) as shown by PCNA expression, and a significant increase in new vessel growth as demonstrated by SMA expression (1762+/-124 cells/mm2 vs. 1067+/-77 cells/mm2; p=0.0001). CONCLUSIONS: Growth hormone accelerates gastric ulcer healing by stimulating cell proliferation and angiogenesis. PMID- 15277986 TI - NK1.1 cells are required to control T cell hyperactivity during Trypanosoma cruzi infection. AB - BACKGROUND: This study evaluated the regulatory function of NK1.1+ cells during Trypanosoma cruzi infection. MATERIAL/METHODS: Both thymectomized (Tx C57Bl/6) and euthymic C57Bl/6 mice (C57Bl/6) were infected intraperitoneally with the Tulahuen strain. NK1.1+ cells were depleted in vivo by anti-NK1.1 mAb. Spleen cells were analyzed by flow cytometry for the expression of CD44 and CD69 on T cells. Supernatants from splenocytes were used to measure nitrite concentration (quantified by Griess reagent). Interleukin 2 and IFN-gamma levels were determined by ELISA. The protocols used herein were approved by the Institutional Committee for Ethics. Student's t or Kruskal-Wallis tests were applied, as indicated. RESULTS: The number of T cells expressing CD69 increased progressively during T. cruzi infection in NK1.1 cell-depleted C57Bl/6 mice. In spite of an increased early T cell activation during infection, the percentage of CD4+ CD44high T cells did not augment in NK1.1 cell-depleted C57Bl/6 mice compared with untreated C57Bl/6 controls. Serum levels of IFN-gamma in anti-NK1.1-treated mice were higher than in non-depleted animals. Con-A-stimulated spleen cell supernatants from NK1.1 cell-depleted animals contained increased levels of IL-2 and nitric oxide (NO) during early infection. CONCLUSIONS: After the first week of infection, NO overproduction and high levels of IFN-gamma in anti-NK1.1-tre ated C57Bl/6 mice appeared to be related to susceptibility and hyperactivation of peripheral T cells. Finally, this study suggests a novel regulatory function of NK1.1+ cells during T. cruzi infection. Without NK1.1 cells, T lymphocytes are hyperactivated but do not differentiate to effector/memory T cells in infected C57Bl/6 mice. PMID- 15277987 TI - Does the crural diaphragm share in the contractile activity of the costal diaphragm? The concept of an "autonomous esophageal crus" and its role in esophageal competence. AB - BACKGROUND: We investigated the hypothesis that the esophageal crus (EC, part of the crus surrounding the esophagus) is an "individual muscle" and does not share in the contractile activity of the costal diaphragm (CD). MATERIAL/METHODS: The electric activities of the EC and the costal diaphragm (CD) were recorded in 21 subjects (12 men, 9 women, aged 41.6+/-10.4 years) scheduled for laparotomy. One needle electrode was introduced into the EC, one into each vertebral crus (VC), and one in the CD. Recording was performed before and after diaphragmatic paralysis by curarization. While the diaphragm was paralyzed, the responses of the EC and the 2 VCs to CD stimulation were registered. RESULTS: The CD had significantly higher resting motor unit action potentials (MUAPs) than the EC and the 2 VCs (p<0.05), and the EC higher MUAPs than the 2 VCs (p<0.05). During diaphragmatic contraction on inspiration or by stimulation (5 square pulses, 1 ms apart, threshold 32.2+/-6.3 mA), the MUAPs of the CD increased significantly (p<0.01), while those of the EC or VCs exhibited no significant change (p<0.05). CONCLUSIONS: The current findings suggest that the EC is an autonomous muscle which does not share in the contractile activity of the CD. The EC seems to be essentially involved in the competent mechanism of the lower esophagus and has no role in respiratory function. PMID- 15277988 TI - Catalytic properties of IgMs with amylolytic activity isolated from patients with multiple sclerosis. AB - BACKGROUND: Recently, amylolytic activity was detected in IgMs isolated from the sera of the patients with multiple sclerosis. MATERIAL/METHODS: All purified samples of IgM were electrophoretically homogenous and did not contain any co purified a-amylase and a-glucosidase activities, in accordance with a set of criteria developed for abzymes. The amylolytic activity of abzymes was studied in the hydrolysis of p-nitrophenyl a-D-maltooligosaccharides with different degrees of polymerization from 1 to 8 by TLC and reverse-phase HPLC techniques. RESULTS: All IgM samples isolated from 54 patients with clinically definite multiple sclerosis demonstrated hydrolytic activity towards the above artificial substrates. The Michaelis constant values (Km) in the hydrolysis of p-nitrophenyl a-D-maltoheptaoside were in the range of 10 p-nitrophenyl or p-nitrophenyl a-D glucosides, thus indicating the presence of an a-D-glucosidase activity. For a number of the investigated samples, specific amylolytic activity increased depending on the length of substrates (from p-nitrophenyl maltopentaoside to p nitrophenyl maltohexaoside); for other IgMs, the opposite dependence was observed. All IgMs studied did not exhibit any other glycoside hydrolase activities toward p-nitrophenyl glycoside substrates. CONCLUSIONS: Abzyme fractions from different donors demonstrated catalytic heterogeneity in Michaelis Menten parameters and different modes of action in the hydrolysis of p nitrophenyl maltooligosaccharides. Enzymatic properties of the IgMs tested varied from human a-amylases. All investigated abzyme samples did not show transglycosylating ability. PMID- 15277989 TI - Evaluation of homeopathy in broiler chickens exposed to live viral vaccines and administered Calendula officinalis extract. AB - BACKGROUND: In this study it was determined that a Calendula officinalis water extract can reduce the immune response to three different viruses in broiler chickens, associated with improvement in body weights. MATERIAL/METHODS: The experiment was conducted on broiler chickens divided into two groups of 105 birds each. The first group received a Calendula officinalis water extract orally, while the second group received drinking water only. All birds in the two groups were similarly exposed to three different live vaccine viruses. Quantitative assessment of humoral immunity to each of the 3 viruses and records of bursal and thymus weight indices were taken. Performance, as observed in weight records at 21 and 41 days of age, feed conversion, and% mortality up to market age, was also evaluated. RESULTS: There was a reduction in immune response to IB virus at 42 days of age, to ND virus at 29 and 42 days of age, and to IBD virus at 14, 29, and 42 days of age in the Calendula officinals-treated birds in comparison with controls. This immune reduction in Calendula officinalis-treated birds was associated with insignificant reduction in the bursal weight index at 42 days of age and an improvement in mean weights at 21 and 41 days of age; the feed conversion and mortality rates were similar in the two groups (P>0.05). CONCLUSIONS: Calendula officinalis had an immunomodulation effect against three different live viruses in broiler chickens. PMID- 15277990 TI - Liquid 2-P-HEMA for endovascular tumor therapy: in vivo feasibility study. AB - BACKGROUND: To evaluate a new liquid polymer developed for selective arterial tumor embolization. MATERIAL/METHODS: In six rabbits we embolized the renal artery with pure 2-P-HEMA. Three rabbits were killed immediately after embolization, one after 12 h, and two animals underwent follow-up angiography after two weeks. In a second part of the study we embolized the left renal artery in three mini-swine with pure 2-P-HEMA. The kidneys were examined post mortem by soft-tissue radiography and kidney-tissue section microscopy. RESULTS: Complete occlusion of small distal arteries, most of the arterioles together with the precapillary vascular bed, was achieved in all animals. The speed of injection influenced the grade of distal occlusion; repeated injections through the same microcatheter were possible. The relatively fast injection speed of 0.3 ml/min led to an optimal distribution of the liquid embolic. There was no reperfusion in the two follow-up angiographies two weeks after embolization. Histopathologically, there was no angionecrosis and subintimal bleeding. No marked inflammatory reaction in the vessel wall or perivascular tissue was observed in the embolized arteries. CONCLUSIONS: This study suggests that liquid 2-P-HEMA may be feasible for the treatment of tumors. PMID- 15277991 TI - Cytotoxic and genotoxic effects of ss-(triphenylpho-s-phonio)ethyl carboxylate and of N,N'-bis(dihexylphos-phinoylmethyl)-1,4-diaminocyclohexane. AB - BACKGROUND: Several organophosphorous compounds (OPs) are now being tested therapeutically. Cholinesterase inhibition, which in large doses makes these agents effective pesticides, may also be useful in other doses for treating dementia. Metrifonate, for example, has been used to treat schistosomiasis and is undergoing trials for the treatment of primary degenerative dementia. MATERIAL/METHODS: Here we report the characterization of newly synthesized OPs from the group of phosphobetaines [beta-(triphenylphosphonio)ethyl carboxylate, PB] and of alpha-aminophosphoryl compounds [N,N'-bis(dihexylphosphinoylmethyl) 1,4-diaminocyclohexane, AP] according to their toxic and genotoxic properties determined in prokaryotic and eukaryotic test systems. RESULTS: The absence of toxicity towards Gram-negative bacteria and of genotoxicity in Ames mutagenicity assay and in SOS-chromotest did not exclude the cytotoxic effect of PB towards NIH3T3 mouse fibroblasts, which supports the notion of an extremely diverse interspecies response to OPs. In contrast, AP demonstrated toxic properties detected by antibacterial effect as well as by the inhibition of the proliferation and respiration of fibroblasts. The enzymatic transformation of the compound is necessary to reveal the genotoxic properties of AP. The role of mammalian microsomal enzymes and of bacterial C-P lyase in the formation of AP genotoxic metabolites is under discussion. CONCLUSIONS: Neither toxicity nor genotoxicity of PB was found in bacterial tests. Cytotoxic and mutagenic effects of AP were detected. The data contribute to the investigation of the biological activity of novel organophosphates which could be useful for the future development of OP-based therapeutics. PMID- 15277992 TI - Lack of relationship between metallothionein (MT) expression and proliferation exponents in cells of primary ductal breast cancer of G2 grade of differentiation. AB - BACKGROUND: The metallothioneins (MTs) are a group of proteins which, due to their unique structure, fulfil numerous functions in the cell. They participate in growth, differentiation, and reparative processes, protect cells against free radicals, and are responsible for heavy metal homeostasis. Their involvement has been reported in the multidrug resistance to cytostatic drugs. Numerous reports document MT presence in cells of various tumors, including breast cancer. Augmented expression of MTs has been reported in less differentiated tumors. MT expression used to be linked to higher proliferative activity of tumor cells, shorter survival of the patients, and tamoxifen-resistance. The present study aimed at examining the relation between MT expression and the manifestation of proliferation exponents (Ki67, nucleolar organizers--AgNORs) in cells of ductal breast cancer of G2 grade of malignancy. MATERIAL/METHODS: Reactions were performed to detect MTs (clone E9), Ki67 (clone MIB-1) (immunocytochemistry), and AgNORs (silver impregnation) in paraffin sections of breast cancers in G2 grade originating from 60 females. Results of the reactions were subjected to statistical analysis using Statistica 98 PL software. RESULTS: Statistical analysis (Spearman's rank correlation) demonstrated no relationships between the studied markers (p>0.05). CONCLUSIONS: There is no correlation between metallothionein expression and proliferation and between Ki67 and AgNORs in ductal breast cancers of G2 grade of differentiation. PMID- 15277993 TI - Effect of long-term hemodialysis on plasma lecithin: cholesterol acyltransferase activity and the amounts and compositions of HDL2 and HDL3 in hemodialysis treated patients with chronic renal failure: a 9-year longitudinal study. AB - BACKGROUND: Atherosclerosis was correlated with hemodialysis duration (HD) in chronic renal failure (CRF) patients. Dyslipidemias were identified as atherogenic risk factors. MATERIAL/METHODS: To investigate variations in HDL2 and HDL3 composition and lecithin: cholesterol acyltransferase (LCAT) activity as functions of HD, 20 CRF patients were selected for maintenance hemodialysis during the 9 years from April 1, 1991 to March 31, 2000. Blood samples were drawn four times: 1991 (T0, study begin), 1994 (T1), 1997 (T2), and 2000 (T3). T0 results were taken as references. RESULTS: Triacylglycerol concentrations were 1.12-fold higher at T1 (P<0.01), 1.31-fold at T2, and 1.63-fold at T3 (P<0.001). Increases of 14% and 33% of total cholesterol were noted at T2 (P<0.05) and T3 (P<0.001). Hypertriglyceridemia correlated with HD (r=0.70, P<0.05). LCAT activity decreased by 27%, 39%, and 51% at times T1, T2, and T3, respectively, this activity being negligible in 30% of patients at T2 and 40% at T3. An inverse relationship was noted between LCAT activity and HD (r=-0.80, P<0.001). Increases in HDL2-unesterified cholesterol (UC) and HDL3-UC were obtained at T2 and T3 (P<0.05), and high HDL2-triacylglycerols (TG) and HDL3-TG were noted at T1, T2, and T3 (P<0.001). HDL3-phospholipids (PL) values were diminished by 9% at T1 (P<0.05), 17% at T2, and 19% at T3 (P<0.001). CONCLUSIONS: Long-term hemodialysis aggravates lipid anomalies following CRF. Alterations in HDL composition contribute to the reduced efficacy of reverse cholesterol transport and patients are submitted to a greater risk for atherosclerosis. PMID- 15277994 TI - Depression and physical health among family caregivers of geriatric patients with cancer--a longitudinal view. AB - BACKGROUND: We investigated, in a sample of 491 cancer patient/caregiver dyads, the impact of caregiving on caregivers' mental and physical health over a period of one year (four assessments). Our model postulates that patient and caregiver characteristics impact caregiver experiences, and all of these in turn affect the mental and physical health of the caregiver. MATERIAL/METHODS: Random-effects regression methods were employed to investigate how patient and caregiver characteristics affect caregiver experiences, and how these same patient and caregiver characteristics affect caregiver physical and psychological health, and are mediated by caregiver experiences. RESULTS: Caregivers' personal perceptions of the caregiving experience (impact on schedule, social functioning, abandonment) played a central role as determinants of caregiver outcomes (depression and physical health). The contextual elements that came to the foreground as either direct or indirect determinants of caregiver outcomes were the patient characteristics symptoms, depression, treatment, comorbidity and cancer site, and the caregiver characteristics education, esteem derived from caregiving, and living arrangement. CONCLUSIONS: Our findings suggest that physicians, oncologists and other health care providers involved in the care of cancer patients should be cognizant of the demands put on caregivers. Periodic assessments and dialogue with the caregiver about their experiences, needs and concerns in combination with a review of the patient's illness trajectory may be necessary to insure that caregivers are able to provide quality care to their patients and not succumb to the burdens of caregiving. PMID- 15277995 TI - Increased plasma homocysteine concentrations in healthy people with hostile behavior: the ATTICA study. AB - BACKGROUND: One of the psychological factors showing significant association with the development of coronary heart disease is hostility. However, the pathway by which hostility may affect coronary risk is not fully understood. Thus we evaluated the association between hostility and inflammation (thrombotic marker) in a population-based sample of males and females with no clinical evidence of cardiovascular disease. MATERIAL/METHODS: The ATTICA study is a health and nutrition survey carried out in the province of Attica, Greece, during 2001-2002. 410 participants (200 men 39+/-12 years old, and 210 women 35+/-10 years old) completed the Hostility and Direction of Hostility questionnaire (range 0-55) and had blood taken for the assessment of high sensitivity C--reactive protein, fibrinogen, white blood cell counts, and plasma homocysteine concentrations. RESULTS: 111 (27%) of the participants were classified in the upper quartile of the hostility scale (>21 score) and 119 (29%) were classified in the lower quartile (<11 score). Multivariate linear regression analysis revealed that hostility score associated positively only with homocysteine levels (standardized Beta=0.124, adj. R2=6%, p=0.015), after controlling for age, gender, educational status, body mass index, physical activity levels, and dietary habits of the participants. In particular, a 10-unit increase in the hostility scale was associated with a 2.9 micromol/l rise in homocysteine levels. CONCLUSIONS: Our findings suggest a positive relationship between homocysteine and hostility; however, whether hostility influences inflammation or the thrombotic process remains to be evaluated by future prospective studies. PMID- 15277996 TI - Neutrophil elastase and interleukin-8 as inflammatory mediators in mechanically ventilated children. AB - BACKGROUND: Neutrophil elastase is a proteolytic enzyme which can have a destructive effect on respiratory tract structures. Interleukin-8 (IL-8), a proinflammatory cytokine, is an important chemoattractant for neutrophils. The aim of our study was to assess inflammatory states by determining elastase in complex with alpha1-proteinase inhibitor (E-alpha1PI) and IL-8 in children requiring mechanical ventilation. MATERIAL/METHODS: Plasma and respiratory tract lavage fluid (RTLF) levels of E-a1PI and IL-8 were measured (ELISA) in 31 children with (group I) and 22 without (group II) respiratory tract infection. Plasma results were compared with a group of healthy controls. Results are given as medians and ranges. Additionally, the percentage content of neutrophils in RTLF was determined. RESULTS: Significantly higher (p<0.00004) plasma levels of E alpha1PI were found in group I than in group II. In group II, there were significantly higher (p<0.002) RTLF levels of E-alpha1PI and IL-8 than in group I. In both groups, the percentage content of neutrophils in RTLF exceeded 60%. A negative correlation was found between the plasma and RTLF levels of E-alpha1PI (r=-0.69; p<0.0004), between E-alpha1PI and percentage neutrophil content (r= 0.6; p<0.006), and between IL-8 and percentage neutrophil content (r=-0.5; p<0.04) in RTLF in group II. CONCLUSIONS: The determination of E-alpha1PI and IL 8 may be useful in the assessment of the inflammatory state in children requiring mechanical ventilation. PMID- 15277997 TI - Screening for CDG type Ia in Joubert syndrome. AB - BACKGROUND: The features of Joubert syndrome include hypotonia, ataxia, characteristic neuro-imaging findings, episodic hypoventilation, psychomotor retardation, and abnormal eye movements. Common symptoms in congenital disorders of glycosylation (CDG) type Ia are muscle hypotonia, cerebellar hypoplasia, ataxia, mental retardation, ophthalmologic involvement, failure to thrive, abnormal fat distribution, and hepatopathy. It has been postulated that some Joubert syndrome patients might have an underlying disorder of protein glycosylation. MATERIAL/METHODS: Screening for disorders of glycosylation was performed in five children diagnosed with Joubert syndrome. Data were retrospectively collected from clinical charts, the patients were reexamined by clinical geneticists, and available neuro-imaging data were also reanalyzed. Diagnoses were established based on results of serum transferrin isoelectric focusing, phosphomannomutase enzyme activity measurements, and DNA mutation analysis. RESULTS: We confirmed the diagnoses of CDG type Ia in two of the five children originally diagnosed with Joubert syndrome. The symptoms of the two syndromes were clearly distinguishable. CONCLUSIONS: Syndromic patients with congenital vermis malformations should be screened for congenital disorders of glycosylation. PMID- 15277998 TI - Angiotensin-converting enzyme gene polymorphism and risk of myocardial infarction in Colombia. AB - BACKGROUND: The ACE gene insertion/deletion polymorphism has been studied as a risk factor for acute myocardial infarction (AMI) in different populations with conflicting results. MATERIAL/METHODS: We conducted a case-control study in first AMI cases matched by age (+/-5 years) and sex to controls with non-cardiac diseases to ascertain whether Colombian carriers of the DD genotype were at higher risk of AMI than carriers of the ID and II genotypes. Zygosity for the deletion-insertion (D-I) of the ACE gene polymorphism was determined by polymerase chain reaction. Logistic regression with adjustment for matching factors was used to estimate the independent effect of the DD polymorphism after controlling for traditional cardiovascular risk factors. RESULTS: Participants (n=202) had a mean age of 62 years and 32% were women. The distribution of the polymorphism was significantly different (p=0.001) in cases (II 8.9%; ID 51.5%; DD 39.6%) and controls (II 8.9%; ID 64.4%; DD 26.7%). After adjustment for other risk factors, the risk of AMI in subjects with genotype DD was 1.98 times higher than the risk in the combined group of genotypes II and ID (95% CI: 1.01, 3.87; p=0.04). There was a significant DD by age interaction (p=0.002). In subjects p=0.86), while in subjects <60 years the risk increased 5.16 times (95% CI: 1.68, 15.90; p=0.004). CONCLUSIONS: These results support an increased risk of AMI in Colombian subjects <60 years with the ACE DD genotype. PMID- 15277999 TI - Comparison of simple indices of insulin sensitivity using the euglycemic hyperinsulinemic clamp technique. AB - BACKGROUND: Insulin resistance is an important factor in the pathogenesis of type 2 diabetes mellitus and other diseases, collectively known as "metabolic syndrome". The gold standard in measuring insulin resistance is glucose clamp, but this method is difficult to apply in large studies. Therefore, indirect indices of insulin sensitivity are proposed. The aim of the present study was to compare these simple indices with data from clamp studies. MATERIAL/METHODS: We examined 51 obese subjects, 23 with impaired glucose tolerance (IGT), 28 with normal (obese-NGT), and 37 healthy lean controls. Insulin sensitivity was determined with the euglycemic hyperinsulinemic clamp technique. We estimated indices of insulin sensitivity: fasting plasma insulin (INS), logarithm INS (log [INS]), homeostasis model assessment (HOMA), logarithm HOMA (log [HOMA]) and quantitative insulin sensitivity check index (QUICKI). RESULTS: With clamp technique, we demonstrated a decrease in insulin sensitivity in both obese groups vs controls, and also in IGT compared to NGT subjects. The differences were significant when we used other indices of insulin sensitivity, except those for INS, log [INS] and HOMA between the two obese groups. Indirect indices correlated with insulin sensitivity derived from clamp in the whole population and in the subgroups of control and NGT-obese subjects. In the IGT group, only the correlations with log [INS], log [HOMA] and QUICKI were significant. CONCLUSIONS: Simple indices may give valuable information about insulin sensitivity in large studies. Indices based on log-transformed plasma glucose and insulin levels are recommended in subjects with IGT. PMID- 15278000 TI - Bone marrow transplantation in the course of hematological malignancies--follow up study in blood serum by 31P MRS. AB - BACKGROUND: Bone marrow transplantation (BMT) is one of the most efficient methods for the treatment of hematological neoplasms. Its successful application requires previous complete remission (CR) and exclusion of residual disease. MATERIAL/METHODS: This 31P MRS study reports data collected during several years of observation of nineteen patients. Duration of observation varied from 1 to 8.5 years from the time of diagnosis. Data collected after BMT covered periods ranging from 1.5 months to 6.5 years. RESULTS: 31P MRS spectra of normal sera showed four peaks, that at the lowest field due to Pi and the other three to phospholipids (PL), i.e. phosphatidylethanolamine + sphingomyelin (PE + SM), lysophosphatidylcholine (LPC), and phosphatidylcholine (PC). Long-term follow-up studies showed good correlation between the 31P MR spectral features of sera and disease response to therapy. In particular, at the time of diagnosis spectra showed significant decreases in the levels of all detected phospholipids (PC, LPC, PE and SM). These decreases, dependent upon disease severity, changed during chemotherapy according to the individual response of the patient. During chemotherapy and after BMT, the spectral profile changed in responding patients to resemble that of normal serum with the typical, higher peak intensities. CONCLUSIONS: Longitudinal analysis of in vitro 31P spectra of sera of patients with hematological malignancies represents a reliable method to evaluate the susceptibility of the neoplasm to a cytostatic treatment. The same approach seems to be useful in evaluating the effectiveness and outcome of bone marrow transplantation over time. PMID- 15278001 TI - A case of tuberculous granuloma at the supra-sternal notch that was difficult to differentiate from a thyroid tumor. AB - BACKGROUND: Despite a decline after World War II, the rate of tuberculosis remains higher in Japan than in other countries. We report a case of tuberculous granuloma at the supra-sternal notch that was difficult to differentiate from a thyroid tumor. CASE REPORT: The patient was a 75-year-old Japanese woman who was referred to our hospital for further investigations and treatment of an anterior neck tumor, that was diagnosed as a suspected of thyroid malignancy by another institute. The thyroid function and biological data were normal except for an elevated erythrocyte sedimentation rate. Imaging studies showed a mass at the supra-sternal notch, and the border between the tumor and the thyroid gland was indistinct. The tuberculosis bacillus group was identified by fine needle aspiration cytology. The patient was treated surgically for tuberculous granuloma, and histopathological findings revealed that the lymph node tuberuculosis had invaded the thyroid gland. We started anti-tuberculous therapy after the operation. The post-operative course was uneventful with good wound healing. CONCLUSIONS: When a markedly elevated erythrocyte sedimentation rate and c-reactive protein value are associated with an anterior neck mass, tuberculosis should be considered in the differential diagnosis of thyroid swelling. Fine needle aspiration cytology is a rapid, simple and effective diagnostic method for extra-pulmonary tuberculous lesions involving the neck. When there is abscess formation or features of compression, or if the mass cannot be differentiated from a thyroid tumor, combined therapy involving anti-tuberculous agents and surgery must be considered. PMID- 15278002 TI - Herpes simplex encephalitis: a case report. AB - BACKGROUND: Despite advances in antiviral therapy over the past 2 decades, herpes simplex encephalitis (HSE) remains a serious illness with significant risk of morbidity and death. HSE is the most common cause of sporadic viral encephalitis, with a predilection for the temporal lobes and a range of clinical presentations, from aseptic meningitis and fever to a severe rapidly progressive form involving altered consciousness. Clinical features of HSE include fever with mental status changes (depressed level of consciousness, confusion, disorientation, personality changes) sometimes with seizures (focal or generalized), dysphagia, or other focal neurological signs. Symptoms vary in intensity early in the disease, but tend to progress rapidly. CT and MRI can play an important role in determining the diagnosis and extent of the disease. CASE REPORT: This case report refers to a 17-year-old girl, previously diagnosed with herpes encephalitis, and presents the outcome of rehabilitation on the patient's mental state during a 7-year follow-up period. CONCLUSIONS: The prognosis for patients with HSE has been dramatically improved by the availability of specific antiviral therapy; sequelae in surviving patients may include severe neurological deficits, seizures, and/or neuropsychological dysfunctions that greatly impair quality of life. To improve the prognosis for patients with HSE, acyclovir treatment should be initiated as soon as HSE is suspected. After discharge, rehabilitation should be provided, in the effort to improve the patient's self-reliance in everyday life. PMID- 15278003 TI - Primary testicular carcinoid. AB - BACKGROUND: Testicular tumors in general can be classified as seminoma, embryonal carcinoma, teratocarcinoma, choriocarcinoma, and mixed tumors. Nevertheless, other histological types may also be observed. In rare cases, primary or secondary neuroendocrine tumors derived from chromaffine cells can be observed in the testicle. CASE REPORT: A 51-year-old patient was hospitalized on account of a two-month-old painless tumescence of the right testicle. Radical orchiectomy revealed a solid, inhomogeneous intratesticular tumor 3 cm in diameter. Pathohistological findings and immunohistochemical staining with different neuroendocrine markers revealed a testicular carcinoid classified as pT1. The excretion of 5-hydroxyindole-acetic acid (5-HIAA) was within the normal range. No further carcinoid tumor site was found. CONCLUSIONS: In case of a testicular carcinoid, exclusion of a primary carcinoid tumor site in other organs is mandatory. PMID- 15278004 TI - Other malignant neoplasms in patients with gastrointestinal stromal tumors (GIST). PMID- 15278005 TI - 14C-urea breath test as a 'gold standard' for detection of Helicobacter pylori infection. PMID- 15278006 TI - Music and medical research. PMID- 15278007 TI - Levosimendan in patients with cardiogenic shock undergoing surgical revascularization: a case series. AB - BACKGROUND: Levosimendan is a new calcium sensitizer, which enhances myocardial contractility while simultaneously having vasodilatory and cardioprotective effects. These properties could be advantageous in patients with myocardial ischemia requiring inotropic support. MATERIAL/METHODS: 10 patients with acute myocardial ischemia, cardiogenic shock, and/or cardiopulmonary resuscitation undergoing emergency surgical revascularization were treated with levosimendan (bolus 6 microg.kg(-1), continuous infusion 0.2 microg.kg(-1).min(-1)) in addition to catecholamines. RESULTS: All patients treated with levosimendan and catecholamines were weaned successfully from CPB on the 1st attempt. All patients needed additional norepinephrine because of vasodilation. In 4 patients, levosimendan was stopped in the early postoperative period. 2 patients died, 8 patients survived without any multiorgan failure. Only 2 patients needed an additional intra-aortic balloon pump. In the surviving patients, postoperative ventilation lasted for 8-72 hours. CONCLUSIONS: Levosimendan may have exerted positive inotropic and cardioprotective effects in these high risk patients with acute myocardial ischemia. However, these preliminary results supporting the use of levosimendan as an inoprotective drug need to be confirmed by a large randomized prospective trial. PMID- 15278008 TI - Topical honey application vs. acyclovir for the treatment of recurrent herpes simplex lesions. AB - BACKGROUND: The objective of this research was to investigate the effect of the topical application of honey on recurrent attacks of herpes lesions, labial and genital, as compared to acyclovir cream. MATERIAL/METHODS: Sixteen adult patients with a history of recurrent attacks of herpetic lesions, 8 labial and 8 genital, were treated by topical application of honey for one attack and acyclovir cream for another attack. RESULTS: For labial herpes, the mean duration of attacks and pain, occurrence of crusting, and mean healing time with honey treatment were 35%, 39%, 28% and 43% better, respectively, than with acyclovir treatment. For genital herpes, the mean duration of attacks and pain, occurrence of crusting, and mean healing time with honey treatment were 53%, 50%, 49% and 59% better, respectively, than with acyclovir. Two cases of labial herpes and one case of genital herpes remitted completely with the use of honey. The lesions crusted in 3 patients with labial herpes and in 4 patients with genital herpes. With acyclovir treatment, none of the attacks remitted, and all the lesions, labial and genital, developed crust. No side effects were observed with repeated applications of honey, whereas 3 patients developed local itching with acyclovir. CONCLUSIONS: Topical honey application is safe and effective in the management of the signs and symptoms of recurrent lesions from labial and genital herpes. PMID- 15278009 TI - Molecular mechanisms and clinical applications of ginseng root for cardiovascular disease. AB - Ginseng root is used extensively in traditional Chinese medicine for its alleged tonic effect and possible curative and restorative properties. There are increasing evidences in the literature on the potential role of ginseng in treating cardiovascular diseases. We herein examine the history of ginseng usage and review the current literature on a myriad pharmacological function of ginseng on the cardiovascular system. From the published studies involving cell cultures and animal models, ginseng is shown to have potential benefits on the cardiovascular system through diverse mechanisms, including antioxidant, modifying vasomotor function, reducing platelet adhesion, influencing ion channels, altering autonomic neurotransmitters release, improving lipid profiles, and involving in glucose metabolism and glycemic control. In addition, the relevant clinical trials regarding the effects of ginseng on the cardiovascular disease are summarized, particularly in managing hypertension and improving cardiovascular function. Finally, the controversies in the literature and the possible adverse interactions between ginseng and other drugs are discussed. This review underscores the potential benefit effects of ginseng on cardiovascular diseases, highlights the gaps in our current research, and emphasizes the necessity for more rigorous systemic investigation. PMID- 15278010 TI - Can a Mediterranean diet moderate the development and clinical progression of coronary heart disease? A systematic review. AB - BACKGROUND: It has been suggested that adherence to a Mediterranean diet reduces all causes of mortality, especially death rates due to coronary heart disease. In this review we summarize the findings of observational studies that evaluated the effect of the Mediterranean dietary pattern in the primary and secondary prevention of coronary heart disease. MATERIAL/METHODS: We retrieved published results from prospective and case-control studies which evaluated the association between adherence to a Mediterranean diet and the occurrence of coronary heart disease outcomes. RESULTS: The benefits from the Mediterranean diet were significant in all studies. The reduction in the risk of coronary heart disease varied from 8% to 45%, depending on the increment used by the investigators in the presentation of their results. CONCLUSIONS: The systematically reviewed studies reveal a cardio-protective effect of the Mediterranean diet and point to this dietary pattern as highly appropriate for public health objectives. PMID- 15278011 TI - Advancing the treatment of anxiety disorders: new findings and novel uses for atypical antipsychotics. PMID- 15278012 TI - Studying non-human primates: a gateway to understanding anxiety disorders. AB - Non-human primates, such as the rhesus monkey, provide excellent models of human fear and anxiety because of similarities in behavioral responses and brain function. Studies of rhesus monkeys demonstrate that animals with an anxious temperament exhibit inappropriately exaggerated responses to fearful situations, extreme asymmetrical electrical activity in the right prefrontal cortex, and dysregulation of the corticotropin-releasing factor system. Similar findings have been observed in anxious or behaviorally inhibited children who are at greater risk of developing anxiety disorders later in life. Characterization of distinct behavioral and neurobiological features in anxious rhesus monkeys may one day form the basis of tools to identify children who are at risk to develop anxiety disorders and other stress-related problems later in life. Psychopharmacology Bulletin. 2004;38(Suppl 1):8-13. PMID- 15278013 TI - Early-Life Adversity, CRF Dysregulation, and Vulnerability to Mood and Anxiety Disorders. AB - A large and growing literature suggests that traumatic experiences early in life increase the risk of mood and anxiety disorders in genetically predisposed persons. Findings from laboratory animal studies as well as studies of women with histories of early-life trauma demonstrate that long-lived alterations in the corticotropin-releasing factor (CRF) system and stress responses underlie this vulnerability. Women with histories of abuse plus current depression exhibit the greatest abnormalities in the hypothalamic-pituitary response to stress and may represent a unique cohort of patients. Studies in laboratory animals also suggest that persistent changes in the CRF system may be reversed by antidepressants or surrogate parenting, which underscores the urgent need for primary and secondary prevention studies in children who are living in adverse or dangerous environments. Psychopharmacology Bulletin. 2004;38(Suppl 1): 14-20. PMID- 15278014 TI - Understanding psychiatric disorders through functional brain imaging. AB - Functional brain imaging technologies, such as positron emission tomography (PET) and single-photon emission tomography (SPECT), are widely used to measure hemodynamic and metabolic neural events associated with psychiatric illness or an experimental challenge. Psychopharmacologic treatment produces adaptive changes in neural circuitry that correspond with changes in behavior. This concept has been tested in studies that were designed to visualize changes in neural receptors and transporters subsequent to pharmacotherapy. The use of functional neuroimaging in distributed neural processing studies offers the hope of identifying individuals at risk of developing psychiatric disorders in addition to treatment responders and non-responders. This technology also has the potential to identify new treatments and novel uses for existing treatments. Psychopharmacology Bulletin. 2004;38(Suppl 1): 21-24. PMID- 15278015 TI - Late-life anxiety disorders. AB - The prevalence, natural course, risk profile, and treatment of anxiety disorders in the elderly are remarkably understudied. Anxiety disorders are less prevalent in the elderly than in younger adults, but rates of subsyndromal anxiety disorders in elderly persons are nearly as high as in their younger cohorts. The most common late-life anxiety disorders are mixed anxiety-depression and generalized anxiety disorder. Though the benzodiazepines are widely used in this population and are considered relatively safe given appropriate dosing and safety monitoring, important liabilities remain with the use of these agents. Antidepressants also are widely used in elderly patients, but there are no randomized controlled anxiety disorder treatment trials in this population. Gabapentin and low-dose atypical antipsychotics are beginning to be used and studies of the atypical antipsychotics are ongoing. Until studies are completed, treatment of late-life anxiety will continue to be guided by extrapolating data from the general adult population. Psychopharmacology Bulletin. 2004;38(Suppl 1): 25-30. PMID- 15278016 TI - Unmet needs in the treatment of anxiety disorders. AB - There are a number of unmet needs in the treatment of anxiety disorders including the need for more effective, rapidly acting, and better tolerated medications; early identification of nonresponse; effective treatments for refractory disorders; prevention of relapse; and promotion of resilience and long-lasting response. Rates of response to contemporary antidepressants and other anxiolytics are often less robust than might be hoped, and remission rates, which have until recently been infrequently measured, are even lower. A small number of mostly uncontrolled studies suggest a role for augmentation of initial therapy with a second modality in patients who do not fully remit to treatment. There also is a small but growing literature which suggests the use of novel anticonvulsants and atypical antipsychotics in the treatment of anxiety disorders should be further studied. However, a definitive place for these newer therapeutic strategies in the anxiety disorder treatment armamentarium awaits evidence from large, controlled studies. Psychopharmacology Bulletin. 2004;38(Suppl 1): 31-37. PMID- 15278017 TI - Drug development for anxiety disorders: new roles for atypical antipsychotics. AB - Anxiety disorders are prevalent and frequently comorbid with depression. Rates of response and remission for anxiety disorders are low despite marked improvements in treatment in the past several decades. Antidepressants and anxiolytics remain the most frequently prescribed agents for anxiety disorders, but the numbers of prescriptions for novel forms of therapy, such as anticonvulsants and atypical antipsychotics are increasing. For the atypical antipsychotics, agonist activity at the 5-HT1A receptor has been hypothesized to translate into anxiolytic effects. A small, but growing, literature suggests that atypical antipsychotics are useful as augmentation therapy for treatment of refractory anxiety disorders. The next generation antipsychotic, aripiprazole, has a unique mechanism of action (ie, combined D2 and 5-HT1A partial agonist and 5-HT2A antagonist) and improves depressive and depressive/anxiety symptoms in patients with schizophrenia. Further studies examining the effect of aripiprazole and other atypical antipsychotic drugs on depressive and anxiety symptoms in patients with refractory anxiety disorders are warranted. Psychopharmacology Bulletin. 2004;38(Suppl 1): 38-45. PMID- 15278018 TI - The challenge of severe acute respiratory syndrome (SARS) in dentistry. AB - Severe Acute Respiratory Syndrome (SARS) is caused by a newly identified coronavirus, called SARS-associated coronavirus (SARS-CoV) that appears to be transmitted primarily through droplets of saliva. This is the reason why the most important international organizations recommend that the dentists adopt a unique preventive approach to the problem: SARS patients should not be treated in the dental office. This is possible only if a suspected case of SARS is correctly and promptly identified. But a correct identification is made difficult by several factors, such as the incubation period, a possibly asymptomatic onset of the illness, the still low specificity and sensitivity of laboratory and instrumental tests. A case or suspected case of SARS may thus unwillingly be treated at the dental office. It is therefore necessary to adopt protective measures for the dental personnel and to implement and enforce infection control measures in order to eliminate the risk of viral contamination. Nonetheless, these procedures do not ensure a complete elimination of SARS-CoV contamination risk since a major risk is represented by blood-borne infection, which is originated by the mouth of patients, and the contamination of dental units water lines (DUWLs) is most difficult to control. Blood-borne contamination may be achieved only by adopting a high level, between-patients disinfection protocol of the DUWLs based on the use of chemical agents with biocidal activity against spores, viruses, bacteria and fungi (Autosteril method). In conclusion a fully effective control of the cross-infection risk will be obtained only by adopting a correct, integrated use of different infection control procedures. PMID- 15278019 TI - A protocol for the treatment of mandibular condylar fractures. AB - The authors suggest a protocol for the treatment of mandibular condylar fractures classified by fracture level: the condylar head, the condylar neck and subcondylar fractures. The protocol provides a guide to individualizing intervention according to the type of fracture, the patient's age, the degree of restriction of mandible movement and the possible presence of fractured condyle displacement, which is often associated with functional disorders. The age of the patient is a key factor in the choice of treatment. The therapeutic goal in adult patients differs from that in growing patients, since in children the condyle is a major growth center for the mandible. Management may be surgical or nonsurgical; surgical intervention may be conservative with or without immobilization with closed or open reduction and fixation. PMID- 15278020 TI - Maxillary post-traumatic outcome correction. Literature review and personal experience. Part III: loss of maxillary substance (free flaps-distraction osteogenesis). AB - Revascularised free flaps retain dual vascularisation, both periosteal and medullary, undoubtedly present optimal survival and minimal re-absorption in view of the prevalence of osteogenetic rather than osteoclastic phenomena. A revascularised free bone flap involves the transfer of a certain amount of bone tissue, whether or not associated with a muscular, skin and/or facial component, with the features of an axial flap, dissecting the vascular stalk of the donor site and re-anastomosing both the arterial and the venous components on to recipient vessels in the site of the primary defect. The vessels in question measure only about 2-4 mm, so that micro-surgery techniques must be applied. For bone defects less than 6 cm, with upkeep of the mandibular or maxillary cortical bone and preservation of the soft tissues, with residual bone of at least 8 mm in height and 4 mm in thickness, alveolar distraction may represent a valid alternative to bone grafts, at the same time as ensuring an increase of the alveolar bone and intraoral soft tissues involved in the distraction process. PMID- 15278021 TI - Diagnosis of temporomandibular disorders according to RDC/TMD axis I findings, a multicenter Italian study. AB - AIM: The aim of the present study was to evaluate the prevalence of the different Research Diagnostic Criteria for Temporomandibular Disorders (RDC/TMD) Axis I types of temporomandibular disorders (TMD) in a population of subjects seeking for TMD treatment at 2 University Departments in Italy and to compare it with data from similar studies in the literature, in order to assess the usefulness of the RDC/TMD classification system as a tool for epidemiological data gathering and multicenter and cross-cultural comparison. METHODS: A total of 210 consecutive patients seeking for TMD treatment at the Section of Prosthetic Dentistry, Department of Neuroscience, University of Pisa, Italy, and 109 consecutive patients seeking treatment at the Section of Prosthetic Dentistry and Temporo-mandibular Disorders, University of Pavia, Italy, were assessed using RDC/TMD. RESULTS: Only 181 of the patients referring to the University of Pisa and 104 of those referring to the University of Pavia satisfied the criteria for inclusion. Findings from the 2 study populations were very similar. Mean age of the patients was 40 years, with a female:male ratio of 3.5:1 (222 females, 77.9%; 63 males, 22.1%). The prevalence of RDC/TMD Axis I diagnoses was 50.2% (143/285) for Group I disorders (muscle disorders), 38.6% (110/285) for Group II disorders (disc displacement), and 50.2% (143/285) for Group III disorders (arthralgia, osteoarthritis, osteoarthrosis). CONCLUSION: Results from the present investigation have confirmed the usefulness of the RDC/TMD classification system for research purposes and for data gathering in cross-cultural and multicenter comparisons. PMID- 15278022 TI - Is clinical assessment valid for the diagnosis of temporomandibular joint disk displacement? AB - AIM: The aim of this work was to evaluate the predictive value of clinical examination for magnetic resonance imaging (MRI) findings of temporomandibular joint (TMJ) disk position abnormalities. METHODS: Participants in this study were 51 consecutive patients with signs and symptoms of temporomandibular disorders (TMD). All 102 temporomandibular joints (TMJ) were evaluated to detect disk position abnormalities by means of a standardized clinical assessement according to Research Diagnostic Criteria for Temporomandibular Disorders (RDC/TMD) and MRI performed by a blinded radiologist at the Section of Prosthetic Dentistry, University of Pisa, Italy. The accuracy of clinical assessment was evaluated with respect to MRI. RESULTS: Clinical assessment showed a good predictive value (PV) for the diagnosis of normal disk position (86.2%) and an acceptable PV for the diagnosis of disk displacement with reduction (70.3%), while it seems less accurate in predicting MRI diagnosis of disk displacement without reduction. The overall agreement between clinical RDC/TMD examination and MRI for the evaluation of disk position was 77.3%. CONCLUSION: Clinical RDC/TMD examination proved to be accurate in detecting normal disk position and disk displacement with reduction but not reliable in predicting MRI diagnosis of disk displacement without reduction in the temporomandibular joint. PMID- 15278023 TI - Pediatric cranio-facial surgery. First-year's experience with a gun applying bioabsorbable tacks. AB - AIM: In 2001 a prototype of a gun to apply bioabsorbable tacks in cranio-facial surgery has been developed. METHODS: From May 2001 to May 2002 this device has been used in the University Hospital of Innsbruck (Austria) for different cranioplasty procedures, in 34 children, showing its reliability for cranio facial bone fixation. The children were affected by isolated craniosynostosis or by syndromical synostosis (Apert, Crouzon) and in a case of benign tumor of the parietal skull vault. The range of age, at the time of surgery, was between 3 months and 204 months of age. Bone segments were fixed using self-reinforced polylactide plates and tacks. RESULTS: Firm fixation was obtained intra operatively and the operative time was reduced about 25-30 minutes as compared to use of plates and screws. This device has just one limitation in its own spring force: sometimes the bone thinner than 1 mm has been broken applying the tacks. CONCLUSION: After the first-year's experience it is possible to confirm that this device reduces, in selected cases, operative time, blood loss, risk of infection and, as a result, the costs. PMID- 15278024 TI - Abnormalities of canines eruption. AB - The aim of this article is to present the personal experience on the treatment of 2 groups of patients presenting the same problem: the recovery of teeth 13 and 23 in the dental arch. The 1(st) group consists of adult patients aged approximately 20-28 years with a retained position of the permanent canines and permanence of the respective deciduous teeth in the dental arch. In the 2(nd) group, the loss of the deciduous teeth had not been followed by the physiological eruption of the permanent teeth. The authors' aim is to demonstrate that similar orthodontic treatments can represent valid solutions for the treatment of pathologies with different etiopathogenesis. PMID- 15278025 TI - Granular cell tumor of the tongue (Abrikossoff's tumor). A case report and review of the literature. AB - Granular cell tumor (GCT), also known as Abrikossoff's tumor, is a relatively uncommon neoplasm presenting as a benign, single, well-circumscribed nodule, usually arising on the tongue. Histologically, GCT shows numerous strands of large polyhedral granular cells, separated by collagen bundles, with no evidence of encapsulation. Recurrences may occur following inadequate excision. A 32-year old woman, presenting a non-painful nodule embedded in the chorion of her tongue, underwent a first excisional biopsy which revealed a GCT and the margins of the bioptic sample appeared free. Immunohistochemistry for protein S-100 revealed the tumor cells invading the margins of the sample. A second intervention was made to excise the possible remaining neoplastic tissue. This case, owing to its uncommon clinical appearance, seemed particularly interesting. Immunohistochemical assay for S-100, whenever a CGR is suspected, is always mandatory to discover tumor cells infiltrating the margins of the sample. Furthermore, immunohistochemical assay can make the diagnosis more precise excluding other more serious tumors which could arise in the oral cavity. In any event, the patient should be reassured of the benign nature of the tumor and a careful follow-up is necessary in order to diagnose relapses. PMID- 15278026 TI - An experimental swine model of small bowel cross transplantation. AB - AIM: In this study we evaluated the possibility of performing a cross small bowel transplantation (CrSBTx) in which, at the same time, 2 pigs were both donors and later recipients of intestinal grafts. The hemodynamic and metabolic impact of this original transplantation model on the animals was determined. METHODS: Ten large White adult female pigs underwent a 2 stage procedure. The principal intraoperative hemodynamic and metabolic parameters were measured at different times during the operation. In the 3 days that followed the operation, renal function, liver and pancreatic damage were investigated. RESULTS: Our surgical model permits us to keep excellent hemodynamic and metabolic stability with low mortality. CONCLUSION: The need of half of animals with respect to conventional models represents an ethical and economic advantage of CrSBTx and we propose it for intestinal transplant studies in large animals. PMID- 15278027 TI - [The role of superextended lymphadenectomy (D4) in gastric cancer]. AB - AIM: The outcome of surgery in gastric cancer differs in Japan and Western countries and the extension of lymphadenectomy may play a crucial role in survival. In Japan the choice of performing extended (D2) and superextended (D4) lymphadenectomies is based on retrospective studies, and a prospective randomized study comparing D2 and D4 is still in course. In Western countries the randomized trials comparing D1 and D2 could not provide definite indications, D2 is not yet performed as a routine procedure and D4 is accepted only by few surgeons. We report our experience and discuss indications and results. METHODS: Since January 2000 through December 2002 we performed 27 superextended lymphadenectomies for the radical treatment of advanced gastric cancer. Early gastric cancers and patients over 80 years of age received conventional D2 gastrectomies. Selection of patients for D4 was made after laparotomy, when intraoperative peritoneal lavage cytology could rule out the presence of malignant cells, while D2 was done in case of peritoneal micrometastases. RESULTS: Every patients had 39.5 nodes removed on average (range 17-94), and micrometastases in tier 16 were found in 7 cases (26%). Early post-operative surgical morbidity was 18% (5 patients) and mortality was 3.7% (1 patient). As much as 30% of patients complained of diarrhea as a late complication. The follow up could demonstrate a 3 year overall actuarial survival of 76%. Actuarial survival was 100% for N- and 70% for N+. A remarkable data was that 4 out of 5 patients who died from recurrence in the follow-up, were N4+. Actuarial survival at 3 years for N4+ patients was 34%, and the difference in survival between N4+ and other N+ was statistically significant (p<0.05). CONCLUSIONS: Superextended lymphadenectomy in gastric cancer is feasible with postoperative morbidity and mortality rates not exceeding the rates of other lymphadenectomies. Actuarial survival at 3 years with D4 was better than in previous personal experience with D2, although the patients who underwent D4 were selected by intraperitoneal lavage cytology, while D2 patients had not been selected. The prognosis for N4- patients was better than for N4+ with micrometastases in tier 16. The presence of N4 micrometastases worsens the prognosis, but it is still uncertain whether D4 does improve survival: it is undoubtedly a new means of more accurate staging in gastric cancer surgery. The newer TNM classification regards the number of nodes removed as an indicator of radicality. Every surgeon should consider that superextended lymphadenectomies could comply with R0 radicality, and perform it within the ranges of low morbidity and mortality, until randomized trials with definitive results are available. PMID- 15278028 TI - [Gastroesophageal reflux and laryngopharyngeal disorders: pH-metric essay of the acid exposure of the proximal and distal esophagus]. AB - AIM: The purpose of this study is to verify the role of gastroesophageal reflux as responsible for otolaryngological symptoms; it is often difficult to identify, both because of its subtle and aspecific clinical-endoscopic picture and because the traditional distal esophageal pHmetry may show readings which are still within the norm at this level. The esophageal pHmetry carried out at proximal level gives greater diagnostic accuracy. METHODS: We use a catheter with 3 antimony electrodes, 2 of them placed at the esophageal level, respectively at 5 and 20 cm above the inferior esophageal sphincter, and 1 in the stomach. We studied 3 groups of patients: 41 asymptomatic no-refluxer patients (group I, control), 59 refluxer patients with only typical esophageal symptoms (group II) and 68 patients with laryngo-pharyngeal symptoms suffering from acid reflux (group III). RESULTS: At both the proximal and distal esophageal levels, the reflux, with reference to control, turned out to be significantly higher in groups II and III, both as regards the number of episodes (p<0.01), the time of acid exposition (p<0.01), the length of the longer reflux (p<0.01), whereas no differences were found between groups II and III. The proximal esophageal acid refluxes in both groups II and III occurred mainly in the upright position: alone (57.2% and 62.5% respectively) or combined with supine position (23.8% and 34.0% respectively). CONCLUSION: Our experience suggests that the esophago-gastric pHmetry with 3 electrodes represents an effective procedure for the correlation between otolaryngological symptoms and gastro-esophageal reflux. PMID- 15278029 TI - [Ultrasound-guided endoscopic drainage, without radiological examination, in patients with neoplastic biliary obstruction. Preliminary results]. AB - AIM: Endoscopic stent insertion has become the preferred method for palliation of malignant biliary obstruction. Currently, endoscopic stent placement involves the use of contrast media and radiological equipment to achieve direct opacification of the biliary duct systems, and to determine the location and the extension of biliary obstruction. This report proposes a new combination of ultrasonography and biliary endoscopy, with endoscopic stent placement entirely performed under US-guidance. METHODS: US-guided stent placement was carried out in 8 patients. A guide-wire and a guiding-catheter were endoscopically introduced and identified, by US, the common bile duct across the stricture. Hydromer-coated polyurethane angled stents (10F) were finally inserted over the guide-wire/guiding-catheter by a pusher tube system. RESULTS: Successful stent insertion was achieved in all patients. There were no complications. Successful drainage, with substantial reduction in bilirubin level, was achieved in all patients (14.2+/-9.5 vs 4.2+/ 2.9 mg/dl at 1 week). CONCLUSION: Endoscopic stent placement performed under US guidance, is safe and effective. Further studies in a larger series, including more proximal strictures are suggested. PMID- 15278030 TI - [Dysplastic cysts of the liver: our experience]. AB - AIM: Systematic surveys with advanced non-invasive imaging techniques have revealed that hepatic cysts are quite common in the general population. Therefore, we retrospectively examined our case series and compared it with the literature. METHODS: Between January 1990 and December 2000, 228 patients with non-parasitic liver cysts were referred to the outpatients section of the Department of Surgery of the University of Cagliari and 23 were submitted to treatment: 14 patients (60.8%) for solitary cyst and 9 (39.2%) for multiple simple cysts of the liver. One patient (4.5%) had right upper quadrant pain. Eleven (47.8%) patients were asymptomatic: 7 (63.7%) required treatment for other pathologies, 3 (27.3%) for a progressive enlargement of the cyst and 1 (9%) for a suspected hydatid disease. Mean diameter of the treated cysts measured by preoperative CT or US was 8.8 cm (range 7-14). Percutaneous aspiration-injection reaspiration (PAIR) was performed in 5 patients (21.7%), US-guided in 2 cases (40%) and CT-guided in 3 (60%). Twenty patients (86.9%) underwent cysts unroofing, 18 (78.2%) with open surgical fenestration and the latest 2 cases with a laparoscopic approach. Two patients had PAIR as second treatment for recurrence: CT-guided in one and US-guided in the other case. RESULTS: Four (25%) out of 16 patients treated exclusively for cystic liver disease, had fever in 3 cases and nausea and vomiting in 1 case; 8 patients (50%) had an intraperitoneal drainage for a mean of 6-7 days (range 4-11) and of 116 cc of serum-hematic liquid. CONCLUSIONS: In our opinion the choice of an adequate treatment must be based on an accurate evaluation of the clinical aspects of the patients and on the characteristics of cystic lesions such as number, size and location. These data let us to choose a surgical treatment rather than a strict US follow-up and to get the best outcome in terms of absence of recurrence, and less biological and economic costs. PMID- 15278031 TI - [Omentoplasty is effective in lowering the complications of ano-rectal resections]. AB - AIM: The aim of this prospective, randomized study was to investigate the influence of omentoplasty on complications following colorectal resection, Hartmann's intervention and abdominoperineal amputation. METHODS: One hundred and seventy-one patients undergoing elective or emergency surgery for malignancy, benign tumor, diverticular disease and others were randomly assigned to omentoplasty (OP group) or not (NO group). The primary goal was to evaluate the rate of clinical and radiological anastomotic leakage. The secondary goal was to assess the morbidity (mainly septic complications) following Hartmann's and Miles' procedures. RESULTS: In colorectal anastomosis, 18 patients (14.3%) had anastomotic leakage, 4 (6.4%) in the OP group and 14 (21.9%) in the NO group. Differences between the 2 groups were also found in terms of repeat operations (3.2% vs 14.1%) and deaths (3.2% vs 7.8%). Other factors associated with anastomotic leakage were the distal site of anastomosis (<5 cm from anal verge) and the emergency. In Hartmann's and Miles' procedures, septic complications were reduced in the OP group. CONCLUSION: Omental wrap, with its mechanical and biological properties, seems to be effective in lowering the rate and the severity of complications after colorectal and anal surgery. PMID- 15278032 TI - [Tension-free laparoscopic versus open inguinal hernia repair]. AB - AIM: During the last decade laparoscopic techniques have been applied to the treatment of inguinal hernia to combine tension-free technique, esthetic, and functional benefits of mini-invasive surgery. Anyway controversy persists regarding the most effective inguinal hernia repair. The aim of this study is to compare the open technique and the laparoscopic approach concerning: complications, recurrences, recovery time and return to usual activity. METHODS: A randomized prospective analysis of 121 consecutive inguinal hernia repairs was performed over a 12-month period. Male well-informed patients with primary monolateral inguinal hernia (ASA I-II) were divided into 2 groups and consecutively treated; group A was treated with laparoscopic transabdominal preperitoneal approach (TAPP) (median age 47+/-7 years, 57 patients), group B with open mesh herniorrhaphy (45+/-6 years, 64 patients). RESULTS: Complication rate was 5.26% for group A (none needed conversion) and 4.68% for group B. All complications were considered minor. No recurrences were observed over a 12-month follow-up in both groups. Post-operative hospital stay and return to activity show statistically significant differences. Median post-hospital stay was 1.7 days for group A while it was longer (2.9 days) for group B. Significant difference was observed in the duration of convalescence too (group A 9.3+/-7.2 days; group B 12.1+/-7. 1 days). CONCLUSION: On the basis of our experience, even if a longer follow-up is needed, the validity of laparoscopic approach to inguinal hernia is confirmed. General anesthesia and higher costs are reasonable compromises for a shorter period of discomfort in patients with a low ASA index and busy job/sport activity. PMID- 15278033 TI - [500 reconstructive flaps in oncological surgery of the head and neck: critical review of 10 years experience]. AB - AIM: Oncologic surgery of the head and neck, according to the principle of oncological radicality, involves large demolition often including skin, soft tissues and bone structures. The aim of this study is to provide a high-level perspective of results achieved in terms of functionality and softness with the different reconstructive techniques during the last ten years. METHODS: The test group was composed of 467 patients, hospitalised in the "Regina Elena" National Cancer Institute in Rome and treated for head and neck cancer; 86% of the treated patients suffered from stage III or IV of the disease. For the reconstructive phase, 506 flaps were used, 45.5% were myocutaneous flaps, 37.1% cutaneous and fasciocutaneous flaps and 17.4% free flaps. Ischemic complications occurred in 5.2% of the myocutaneous flaps group and in 10.3% of the free flaps group. RESULTS: "Minor complications" affected 3.9% of the cases in the free flaps group and 20.8% of cases in the myocutaneous flaps group. CONCLUSION: The analysis of the results shows that generally free flaps are more reliable than myocutaneous flaps in terms of minor complications, however they tend to prolong the patient's hospitalisation and increase the overall treatment cost. Furthermore in morpho functional terms, free flaps ensure results, in defined anatomical areas (such as the cervical-esophageal region, jaw, tongue) that today cannot be compared to the "conventional" procedures. As for myocutaneous and cutaneous/ fasciocutaneous flaps, according to personal opinion they are still the first choice for reconstruction having minimal or no functional implications. PMID- 15278034 TI - [Surgical treatment of pilonidal disease. Results with the Bascom's technique]. AB - AIM: Pilonidal sinus is a considerable source of problems in young patients both in terms of discomfort and in time off to work. Many procedures have been proposed for its treatment but most of them present substantial persistence/recurrence rates. Surgical procedures avoiding a wound in the midline are most likely to succeed. Bascom's technique is the simplest and successful method. The aim of this study is to retrospectively evaluate the results of the Bascom's procedure performed by the authors as to healing time and recurrence rate. All patients with chronic pilonidal disease, treated with Bascom's technique were re-viewed. Complications, healing time and long-term follow-up were considered. RESULTS: A total of 74 patients (52 males, and 22 females), were admitted to the study. The mean age was 26 years; 69 had a small sinus with 1-2 tracks. Three patients (4%) had postoperative bleeding or wound infection. Mean healing time was 39 days but all patients were able to return to work within 1 week from the operation. The mean period of follow-up was 45 months. Six patients developed recurrence (9,2%). Only 3 of them, (because symptomatic) required a second operation. CONCLUSION: Bascom's technique is simple and suitable for one day surgery with local anesthesia. It also gives favorable results as to return to work and rate of recurrence. Therefore, it is suggested as the procedure of choice in the initial treatment of symptomatic pilonidal disease. PMID- 15278035 TI - [Surgical complications for gastric and small bowel metastases due to primary lung carcinoma]. AB - Lung carcinoma is the first cause of death for tumors; in Italy there are from 35,000 to 40,000 new cases a year, with a global survival to 5 years from the diagnosis in 13% of cases. The gastric and small bowel metastases are rare, respectively 0.4% and 1.1% and the cases reported in the literature are rare. There is often a poor symptomatology and diagnosis is usually based on post mortem examination. Yet, sometimes, the first and only demonstration of the secondary illness is a surgical complication, whether it be a haemorrhage, a perforation or an occlusion. The authors report their experience, from April 1999 to March 2003, of 3 cases of small bowel metastases, presented in 1 case with a perforation and in 2 cases with an occlusion and of 1 case of gastric metastasis, which presented with a haemorrhage. These patients were treated by emergency surgery, with 1 case only of postoperative mortality. The 1st patient died 6 months after surgery, and the 2nd patient 6 days after, the 3rd patient 4 months after the 1st operation and the 4th patient is still in follow-up after 6 months. The authors present a review of the literature and some considerations of diagnosis and surgical treatment. PMID- 15278036 TI - [Acute abdomen due to rupture of mesenteric cysts. Observations on a clinical case and review of the literature]. AB - Personal experience based on a clinical case of a young woman with acute abdominal pain referable to acute appendicitis is presented. The surgical procedure was performed through a Mc Burney incision and revealed the rupture of mesenteric cysts; removal of the cysts was carried out without intestinal resection. Post-operative course was uneventful and 2 years follow-up showed no recurrence. Mesenteric cysts are an uncommon pathology, mainly in adult ages. After an analysis of the incidence and etiology, the pathological features and types of clinical presentation are discussed. Diagnosis in asymptomatic cases is usually made in search of other diseases. Complications are rare: rupture, infection and intestinal obstruction. In such cases, the clinical presentation is usually attributable to the main causes of acute abdomen, unless ultrasonography or CT scan are performed. When mesenteric cyst is diagnosed, a laparoscopic approach should be performed, even if in emergency traditional surgery is justified. Total excision of the cyst is necessary to avoid recurrence and obtain a correct pathologic evaluation. PMID- 15278037 TI - Is video-assisted thoracic surgery always safe? PMID- 15278038 TI - Qualification of the remnant gland after subtotal thyroidectomy. PMID- 15278039 TI - Training guideline for use of propofol in gastrointestinal endoscopy. PMID- 15278040 TI - A prospective, randomized trial comparing mechanical methods of hemostasis plus epinephrine injection to epinephrine injection alone for bleeding peptic ulcer. AB - BACKGROUND: The hemostatic efficacy of mechanical methods of hemostasis, together with epinephrine injection, was compared with that of epinephrine injection alone in bleeding peptic ulcer. METHODS: Ninety patients with a peptic ulcer with active bleeding or a non-bleeding visible vessel were randomly assigned to undergo a mechanical method of hemostasis (23 hemoclip application, 22 band ligation) plus epinephrine injection, or epinephrine injection alone. RESULTS: The two groups were similar with respect to all background variables. Initial hemostasis was achieved in 44/45 (97.8%) patients in both groups. The mean number of hemoclips and elastic bands applied were 2.8: 95% CI[2.5, 3.1] and 1.1: 95% CI[1.0, 1.2], respectively, and the mean volume of epinephrine injected was 19.9 mL: 95% CI[19.3 mL, 20.5 mL]. The rate of recurrent bleeding in the combination group (2/44, 4.5%) was significantly lower in comparison with the injection group (9/44, 20.5%, p < 0.05). The mean number of therapeutic endoscopic sessions needed to achieve permanent hemostasis in the combination group (1.04: 95% CI[1.01, 1.07]) was significantly lower vs. the injection group (1.22: 95% CI[1.15, 1.30]). CONCLUSIONS: The combination of an endoscopic mechanical method of hemostasis plus epinephrine injection is more effective than epinephrine injection alone for the treatment of bleeding peptic ulcer. PMID- 15278041 TI - Endoscopic sphincterotomy vs. endoscopic papillary balloon dilation for choledocholithiasis in patients with liver cirrhosis and coagulopathy. AB - BACKGROUND: To determine whether endoscopic papillary balloon dilation decreases the risk of hemorrhage without increasing the risk of acute pancreatitis, the results of endoscopic papillary balloon dilation were compared with those of endoscopic biliary sphincterotomy in patients with cirrhosis and coagulopathy. METHODS: Twenty-one patients with liver cirrhosis with coagulopathy had endoscopic papillary balloon dilation for choledocholithiasis from January 2001 to September 2003. Twenty patients with cirrhosis and coagulopathy who underwent endoscopic biliary sphincterotomy from January 1998 to December 2000, served as a historical control group. RESULTS: The rate of endoscopic biliary sphincterotomy related hemorrhage was 30% (6/20), whereas the rate for endoscopic papillary balloon dilation related hemorrhage was 0% (p=0.009). With regard to rates of hemorrhage in relation to Child-Pugh class, most (n=5) of the bleeding complications occurred in patients with Child-Pugh class C cirrhosis; bleeding occurred in only one patient with Child-Pugh B cirrhosis. There was no significant difference between the endoscopic biliary sphincterotomy and the endoscopic papillary balloon dilation groups for procedure-related pancreatitis (10% vs. 4.7%, respectively; p>0.05). CONCLUSIONS: Endoscopic papillary balloon dilation may significantly reduce the risk of bleeding compared with endoscopic biliary sphincterotomy in patients with advanced cirrhosis and coagulopathy. In these patients, the substitution of endoscopic papillary balloon dilation for endoscopic biliary sphincterotomy is recommended for treatment of choledocholithiasis. PMID- 15278042 TI - Endoscopic papillary balloon dilation causes transient pancreatobiliary and duodenobiliary reflux. AB - BACKGROUND: Endoscopic papillary balloon dilation reduces sphincter function at least transiently or partially, which may allow pancreatobiliary and duodenobiliary reflux to occur. This study prospectively evaluated pancreatobiliary and duodenobiliary reflux after endoscopic papillary balloon dilation. METHODS: In 12 patients with choledocholithiasis, ductal bile was sampled for amylase concentration and bacterial culture during ERCP, before and at 7 days to 5 years after endoscopic papillary balloon dilation. To provide comparative and control data, ductal bile was sampled in 12 patients with gallbladder cholesterol polyps and 6 with anomalous pancreaticobiliary junction who did not undergo endoscopic papillary balloon dilation. RESULTS: Amylase concentrations in ductal bile from patients with choledocholithiasis before endoscopic papillary balloon dilation were marginally significantly higher (before Bonferroni correction) compared with concentrations in bile from patients with gallbladder polyps. The concentration of amylase in bile was significantly increased at 7 days after endoscopic papillary balloon dilation compared with that before endoscopic papillary balloon dilation; the level was comparable with that of patients with an anomalous pancreaticobiliary junction. Subsequently, the amylase concentration gradually decreased and was approximately equal to the pre endoscopic papillary balloon dilation level at 1 year. Bacteriocholia was frequent (67%-92%) for up to 3 months after endoscopic papillary balloon dilation but was rare thereafter. CONCLUSIONS: Endoscopic papillary balloon dilation causes transient pancreatobiliary and duodenobiliary reflux. However, reflux is no longer present at 1 year after endoscopic papillary balloon dilation. PMID- 15278043 TI - Hemorrhoidal elastic band ligation with flexible videoendoscopes: a prospective, randomized comparison with the conventional technique that uses rigid proctoscopes. AB - BACKGROUND: Elastic band ligation by means of a rigid proctoscope is the treatment of choice for patients with symptoms caused by internal hemorrhoids of grade 2 to 3. In contrast to the flexible videoendoscope, the rigid proctoscope has limited maneuverability, has a narrower field of view, and does not allow adequate documentation. Therefore, a randomized trial was conducted to compare the safety and the efficacy of conventional elastic band ligation with videoendoscopic elastic band ligation. METHODS: A total of 100 consecutive patients (mean age 47 [12] years) with chronically bleeding grade 2 or 3 internal hemorrhoids were randomized to elastic band ligation or videoendoscopic elastic band ligation. For videoendoscopic elastic band ligation, a reusable multiband ligator was attached to the end of a therapeutic upper videoendoscope. From one to 3 bands were placed per session in both groups. Re-treatment was performed every 2 to 3 weeks in both groups until cessation of bleeding and eradication of the hemorrhoids (at least grade 2) were achieved. Thereafter, the patients were followed to assess complications and efficacy. Recurrent bleeding was considered a treatment failure. RESULTS: To achieve the desired therapeutic aims, a significantly lower number of treatment sessions was required in the videoendoscopic elastic band ligation group (1.8 [0.8] vs. 2.4 [0.9]; p < 0.01) and the total number of bands applied was significantly less (2.8 [1.1] vs. 3.7 [1.4]; p < 0.01). Pain was noted after ligation by 25% of patients in the elastic band ligation group compared with 27% of those who had videoendoscopic elastic band ligation. However, analgesic medications were required in only 7% after elastic band ligation and 9% after videoendoscopic elastic band ligation (NS). Post-ligation bleeding that had to be treated endoscopically occurred in 3.5% of the patients of the elastic band ligation group and 3.2% of those in the videoendoscopic elastic band ligation group (NS). Blood transfusion was not required. At a median follow-up of 12 months, there was no recurrence of bleeding in 40 patients (80%) in the conventional elastic band ligation group vs. 43 (86%) in the videoendoscopic elastic band ligation (NS). CONCLUSIONS: The long-term efficacy and safety of conventional elastic band ligation and videoendoscopic elastic band ligation are highly comparable. However, when videoendoscopic elastic band ligation is performed, significantly fewer treatment sessions are required. PMID- 15278044 TI - Training with a computer-based simulator achieves basic manual skills required for upper endoscopy: a randomized controlled trial. AB - BACKGROUND: Changes in medical practice have constrained the time available for education and the availability of patients for training. Computer-based simulators have been devised that can be used to achieve manual skills without patient contact. This study prospectively compared, in a clinical setting, the efficacy of a computer-based simulator for training in upper endoscopy. METHODS: Twenty-two fellows with no experience in endoscopy were randomly assigned to two groups: one group underwent 10 hours of preclinical training with a computer based simulator, and the other did not. Each trainee performed upper endoscopy in 19 or 20 patients. Performance parameters evaluated included the following: esophageal intubation, procedure duration and completeness, and request for assistance. The performance of the trainees also was evaluated by the endoscopy instructor. RESULTS: A total of 420 upper endoscopies were performed; the computer pretrained group performed 212 and the non-pretrained group, 208. The pretrained group performed more complete procedures (87.8% vs. 70.0%; p < 0.0001), required less assistance (41.3% vs. 97.9%; p < 0.0001), and the instructor assessed performance as "positive" more often for this group (86.8% vs. 56.7%; p < 0.0001). The length of procedures was comparable for the two groups. CONCLUSIONS: The computer-based simulator is effective in providing novice trainees with the skills needed for identification of anatomical landmarks and basic endoscopic maneuvers, and in reducing the need for assistance by instructors. PMID- 15278045 TI - Detection of colorectal polyps by multislice CT colonography with ultra-low-dose technique: comparison with high-resolution videocolonoscopy. AB - BACKGROUND: This prospective study compared multislice CT colonography with ultra low-dose technique to high-resolution videocolonoscopy as the standard for detection of colorectal cancer and polyps. METHODS: After standard bowel preparation, 115 patients underwent multislice CT colonography with an ultra-low dose multislice CT colonography protocol immediately before videocolonoscopy. After noise reduction by using a mathematical algorithm, ultra-low-dose multislice CT colonographic images were analyzed in blinded fashion, and the results were compared with the results of high-resolution videocolonoscopy. RESULTS: A total of 150 lesions were detected by high-resolution videocolonoscopy in 115 patients. For ultra-low-dose multislice CT colonography, sensitivities for detection of polyps less than 5 mm in size, 5 to 10 mm, and greater than 10 mm in diameter were 76%, 91%, and 100%, respectively. Although the sensitivity for detection of flat lesions was only 50%, the sensitivity and the specificity for detection of polyps 5 mm or greater in size were 94% and 84%, respectively. For adenomatous lesions greater than 5 mm in size, sensitivity was 94% and specificity was 92%. The overall specificity was 79%. The calculated effective radiation dose ranged between 0.75 and 1.25 mSv. CONCLUSIONS: Compared with high resolution videocolonoscopy, ultra-low-dose multislice CT colonography has excellent sensitivity and specificity for detection of colorectal lesions 5 mm or greater in size, and the radiation exposure is relatively low. However, before this technique can be generally recommended for colorectal screening, further improvement in the detection of flat and extremely small lesions must be achieved. PMID- 15278046 TI - Risk factors for acute biliary pancreatitis. AB - BACKGROUND: Few studies have analyzed both stone-related and pancreatobiliary anatomical factors that predispose to acute biliary pancreatitis. Both of these factor types were studied by multivariate analysis. METHODS: A total of 143 patients with (n=43) or without (n=100) recent acute biliary pancreatitis who underwent cholecystectomy for gallbladder stones after ERCP were prospectively studied. The interval between the onset of pancreatitis and ERCP ranged from 12 to 35 days (mean, 20 days). Univariate and multivariate analyses for 15 potential risk factors for acute biliary pancreatitis, including operative and ERCP findings, were performed. RESULTS: Univariate analysis identified 5 significant predictive factors for pancreatitis: a diameter of the smallest gallbladder stone of 5 mm or less, a cystic duct diameter of 5 mm or more, 20 or more gallbladder stones, a diameter of the largest gallbladder stone of 5 mm or less, and irregular gallstone surface. Of these 5 factors, the first 3 remained significant in the multivariate analysis. CONCLUSIONS: Both stone-related factors (small and multiple stones) and an anatomical factor (enlarged cystic duct) may contribute to the development of biliary pancreatitis. These features should be carefully considered during management of patients with gallstones. PMID- 15278047 TI - Healing rate of EMR-induced ulcer in relation to the duration of treatment with omeprazole. AB - BACKGROUND: Although EMR-induced ulcers heal faster and recur less often than noniatrogenic gastric ulcers, there is no consensus regarding the duration of therapy for these ulcers. This study prospectively evaluated healing of EMR induced ulcers according to the duration of omeprazole therapy. METHODS: A total of 69 patients were randomly assigned, after EMR, to treatment with omeprazole (20 mg per day) for 7 days (1-week group) or with omeprazole (20 mg per day) for 28 days (4-week group). Four weeks after EMR, ulcer size and stage were compared with those of the initial EMR-induced ulcer. Each patient kept a daily diary of drugs consumed and ulcer-related symptoms during the 4-week period after EMR. RESULTS: Thirty-four patients were randomized to the 4-week group, and 26 were randomized to the 1-week group. No significant differences were observed between the two groups at 4 weeks after EMR in terms of ulcer reduction ratio (p=0.29) or stage (p=0.11). In addition, no difference was observed between the two groups with respect to ulcer-related symptoms or use of additional gastric-coating medication (p=0.48). CONCLUSIONS: For EMR-induced ulcer, treatment with omeprazole for 1 week is equivalent to treatment for 4 weeks. Short-term therapy with omeprazole can be considered for EMR-induced ulcer. PMID- 15278048 TI - Endoscopic band ligation without electrosurgery: a new technique for excision of small upper-GI leiomyoma. AB - BACKGROUND: Leiomyoma is a relatively common submucosal tumor in the upper-GI tract. The efficacy of a new method for resection of these tumors, endoscopic band ligation, was evaluated. METHODS: The study included 59 patients with 64 small upper-GI leiomyomas arising in the muscularis propria as determined by endoscopy, EUS, and EUS-guided FNA. The distribution of the 64 leiomyomas was the following: esophageal, 50; gastric, 12; duodenal, 2. A standard endoscope with a transparent cap attached to the tip was used. The cap was placed over the lesion, maximum sustained suction was applied, and an elastic band was released around the base. Beginning 2 weeks after banding, the lesions were observed endoscopically once per week until healing was complete. RESULTS: The 50 esophageal leiomyomas sloughed completely. The mean time required for complete healing after band ligation was 3.6 weeks. Nine of the 12 gastric leiomyomas sloughed completely; the resulting ulcer defect was healed at a mean of 4.5 weeks. The other 3 lesions did not slough because they were not completely ligated. The two duodenal lesions sloughed completely after banding, and the mean time until healing of the defect was 4.5 weeks. No perforation occurred. Follow up ranged from 16 to 31 months, during which time no recurrence was observed. CONCLUSIONS: Endoscopic band ligation is an effective and safe treatment for small upper-GI leiomyoma. PMID- 15278049 TI - Reliability of the "immersion technique" during routine upper endoscopy for detection of abnormalities of duodenal villi in patients with dyspepsia. AB - BACKGROUND: Upper endoscopy is not routinely performed to directly detect abnormalities of the duodenal villi. The reliability of the immersion technique for assessment of duodenal villi was evaluated in a series of patients with dyspepsia. METHODS: A total of 396 patients who were to undergo standard EGD for dyspeptic symptoms were enrolled. Patients with suspected malabsorption were excluded. By performing a "modified immersion technique," duodenal villi were scored as the following: definitely present, partially present, or definitely absent. Three duodenal biopsy specimens were obtained from each patient, and villi also were scored histologically as the following: normal, partial villous pattern, or total villous atrophy. RESULTS: Sensitivity, specificity, and positive and negative predictive values of the modified immersion technique for detection of total villous atrophy were 100%, 99.7%, 85.7%, and 100%, respectively. Sensitivity, specificity, and positive and negative predictive values of modified immersion technique for detection of partial villous patterns were 75%, 99.5%, 60%, and 99.7%, respectively. Sensitivity, specificity, and positive and negative predictive values for modified immersion technique detection of any villous abnormality (partial or total villous atrophy) were 90.9%, 99.5%, 83.3%, and 99.7%, respectively. CONCLUSIONS: During standard EGD, duodenal evaluation by modified immersion technique can reliably detect abnormalities of duodenal villi. This simple diagnostic technique may be performed routinely during endoscopic exploration of duodenum. PMID- 15278050 TI - Endoscopic balloon dilation for benign gastric outlet obstruction with or without Helicobacter pylori infection. AB - BACKGROUND: Endoscopic balloon dilation has been used to treat patients with gastric outlet obstruction caused by peptic stricture. This study assessed the role of endoscopic balloon dilation in patients with gastric outlet obstruction with or without Helicobacter pylori infection. METHODS: Consecutive patients seen between January 1996 and September 2001 with benign gastric outlet obstruction (defined as stenosis preventing the passage of a 9-mm diameter endoscope, vomiting, succussion splash, and recent weight loss) were prospectively studied. Exclusion criteria were the following: refusal to undergo dilation, and gastric outlet obstruction because of malignancy. At endoscopy, antral biopsy specimens were obtained for histopathologic evaluation and for a rapid urease test for Helicobacter pylori infection. Patients then underwent dilation with through-the scope balloons. After balloon dilation, patients with Helicobacter pylori infection were treated to eradicate the infection. RESULTS: Fifty-one patients (33 men, 18 women; median age 65 years; IQR 44-79 years) were studied; 33 consented to endoscopic balloon dilation. Symptom resolution occurred in 25 patients (14 Helicobacter pylori positive, 11 Helicobacter pylori negative). During a median follow-up of 24 months (IQR 16-40 months), 3 of 14 patients in the Helicobacter pylori positive group and 6 of 11 in the Helicobacter pylori negative group developed further ulcer complications (p=0.039). CONCLUSIONS: After endoscopic dilation for gastric outlet obstruction, eradication of Helicobacter pylori infection is associated with fewer ulcer complications. PMID- 15278051 TI - EMR of large sessile colorectal polyps. AB - BACKGROUND: EMR optimizes histopathologic assessment of resected lesions. This study evaluated the outcome of EMR of large sessile colorectal polyps in terms of complications and recurrence. METHODS: An uncontrolled prospective study was conducted of a cohort of 136 patients with sessile colorectal polyps referred for EMR. After submucosal injection, EMR was performed piecemeal by either snare polypectomy alone or with cap aspiration. RESULTS: In 136 patients, a total of 139 sessile polyps were resected, 86 of which were in the right colon. Median polyps diameter was 20 mm in the right colon and 30 mm in the other colonic segments. Intraprocedure bleeding occurred after 15 polypectomies (10.8%) and was controlled endoscopically in all cases; there was no delayed bleeding. Post polypectomy syndrome occurred in 5 patients (3.7%). There was no perforation. Invasive carcinoma was found in 17 sessile colorectal polyps, and surgery was performed in 10 of 17 cases. Follow-up colonoscopy in 93 patients without invasive carcinoma (96 polyps), over a median of 12.3 months, disclosed local recurrence of 21 adenomatous polyps (21.9%). Colonoscopic follow-up in 5 of the 7 patients, who had sessile colorectal polyps with invasive carcinoma and did not undergo surgery, disclosed no local recurrence. CONCLUSIONS: EMR, including EMR with cap aspiration, is effective and safe for removal of sessile colorectal polyps throughout the colon. PMID- 15278052 TI - Predictive factors for lymph node metastasis of differentiated submucosally invasive gastric cancer. AB - BACKGROUND: For early gastric cancer, submucosal invasion may be unrecognized until histopathologic examination of the specimen obtained by EMR. Gastrectomy with lymphadenectomy is the standard treatment for such submucosal cancers. However, approximately 80% of submucosal cancers do not have lymph node metastasis. Unnecessary surgery could be avoided if a subgroup of patients with submucosal cancer with negligible risk of lymph node metastasis can be defined. This study was conducted to define such a subgroup. METHODS: Data from 104 patients surgically treated for differentiated submucosal cancers were retrospectively collected. A multivariate analysis of clinicopathologic factors was performed to identify predictive factors for lymph node metastasis. RESULTS: Three independent risk factors, namely, female gender (p=0.0174), deep invasion (> or =500 microm) into the submucosal layer (p=0.001), and presence of lymphatic involvement (p < 0.0001) were associated with lymph node metastasis. Lymph node metastasis was not observed in any patient who had limited submucosal invasion and absence of lymphatic involvement. The rate of lymph node metastasis was calculated to be 80% in patients who had both deep submucosal invasion and lymphatic involvement. CONCLUSIONS: If endoscopic resection specimens exhibit no deep penetration (<500 microm) into the submucosal layer and lymphatic involvement is absent, EMR may be sufficient treatment for submucosal well differentiated early gastric cancers. A long-term follow-up study of patients with such lesions treated by EMR alone is required. PMID- 15278053 TI - Reliability, cost-effectiveness, and safety of reuse of ancillary devices for ERCP. AB - BACKGROUND: The choice between reusable and single-use devices for ERCP depends on various medical and economic criteria. This study evaluated the reliability and the safety (risk of cross-contamination) of reusable devices. A cost analysis of the use of reusable devices also was conducted. METHODS: All patients referred for ERCP that required use of a sphincterotome or a retrieval basket were eligible for inclusion in a clinical study of 4 different devices (3 types of sphincterotome, 1 type of retrieval basket). All devices were steam sterilized. Before each use, each device was subjected to bacteriologic and virologic tests (hepatitis C virus, hepatitis B virus markers). Devices were examined before and after each procedure. The numbers of safe and efficient procedures that could be performed with each device were assessed. Three strategies were compared in a cost analysis: internal reprocessing (strategy 1), external reprocessing (strategy 2), and single-use (strategy 3). Inputs used were the results of the clinical study, hospital data for 1 year of endoscopic activity, and market prices. RESULTS: A total of 342 patients underwent the following procedures: sphincterotomy (248 patients), stent insertion (59 patients), use of basket without sphincterotomy (14 patients), and diagnostic ERCP/unsuccessful cannulation (21 patients). At the time of ERCP, 36 patients had viral or bacterial infection. Fifty instruments were used (20 single-lumen sphincterotomes, 10 double lumen sphincterotomes, 20 retrieval baskets). Overall, the median number of efficient uses per device was 10. The median number of efficient uses by each type of device was the following: single-lumen sphincterotome, 12; double-lumen sphincterotome, 8; and, retrieval baskets, 10. All virologic and bacteriologic tests for all instruments were negative. The cost optimization analysis found that strategy 1 is cost effective (euro37,283/y) compared with strategy 2 (euro40,101/y) and especially with Strategy 3 (euro115,210/y). CONCLUSIONS: Reuse of the sphincterotomes and baskets evaluated in this study during ERCP is safe in terms of infectious hazards. Because they endure numerous uses, reusable instruments are cost effective, especially when compared with single-use accessories. PMID- 15278054 TI - Classifications of esophagitis: who needs them? PMID- 15278055 TI - Colonic abscess. PMID- 15278056 TI - Pedunculated colonic lipoma. PMID- 15278057 TI - Terminal ileal lipoma. PMID- 15278058 TI - Ileocolic intussusception. PMID- 15278059 TI - Meckel's diverticulum mimicking small bowel tumor. PMID- 15278060 TI - Bleeding Meckel's diverticulum. PMID- 15278061 TI - Metastatic renal cell cancer. PMID- 15278062 TI - Perforated duodenal ulcer. PMID- 15278063 TI - EUS-guided FNA. PMID- 15278064 TI - Fluorescence in situ hybridization of cytologic specimens from Barrett's esophagus: a pilot feasibility study. AB - BACKGROUND: Endoscopic brush cytology is a promising surveillance technique for Barrett's esophagus. However, there is a need for ancillary biomarkers to increase the sensitivity of cytology and to allow identification of patients at increased risk for disease progression. The aims of this study were to evaluate the feasibility of fluorescence in situ hybridization of endoscopic brush cytology specimens and to determine if there are specific chromosomal changes in cytologic specimens from patients with cancer that are not present in patients without dysplasia. METHODS: Archival cytology slides from 16 patients with Barrett's esophagus were studied: 8 negative for dysplasia and 8 positive for adenocarcinoma. Fluorescence in situ hybridization was used to detect two alterations: HER-2 gene (17q11.2-q12) and 20q13.2 region amplification. OBSERVATIONS: For 7 of 8 adenocarcinoma cases, there was amplification/aneusomy of at least one of the two analyzed regions by fluorescence in situ hybridization. None of the samples negative for dysplasia were abnormal for either of the two genomic regions studied. CONCLUSIONS: Fluorescence in situ hybridization is feasible by using routine Barrett's esophagus cytologic specimens. Differences in genomic makeup can be detected in cells from patients negative for dysplasia and in those with adenocarcinoma. PMID- 15278065 TI - Initial evaluation of a duodenoscope modified to allow guidewire fixation during ERCP. AB - BACKGROUND: Catheter/guidewire exchanges during ERCP require the coordinated efforts of an endoscopist and endoscopy assistant. A prototype duodenoscope was developed to improve the control of catheter/guidewire exchange by enabling fixation of guidewires at the elevator lever. METHODS: An initial prototype duodenoscope and a subsequent modification of this instrument were used to perform ERCP in 7 and 10 patients, respectively. The following were recorded: total procedure time, fluoroscopy time, catheter/guidewire exchange time, guidewire repositioning, loss of guidewire access, success or failure of fixation, and endoscopist satisfaction. OBSERVATIONS: The initial and the modified prototype duodenoscopes were used in a variety of catheter/guidewire exchanges (n=46). Guidewire fixation was achieved in 75% of catheter/guidewire exchanges with the initial prototype and in 93% with the modified prototype and was reflected in shorter exchange times. Access to the desired duct was not lost during any exchange, and the need for repositioning was eliminated. CONCLUSIONS: A new prototype duodenoscope with an elevator lever that enables guidewire fixation will improve the ease and efficiency of catheter/guidewire exchange during ERCP. Modifications made to the original prototype improved reliability of guidewire fixation. PMID- 15278066 TI - Safety and efficacy of ERCP in pregnancy. AB - BACKGROUND: Choledocholithiasis during pregnancy increases the risk of morbidity and mortality for both fetus and mother because of cholangitis and pancreatitis. ERCP has been advocated as safe and effective in pregnant women, but fetal radiation exposure is not routinely monitored. The aim of this study was to record fetal exposure to ionizing radiation during ERCP and to assess outcome. METHODS: Seventeen ERCPs were performed in pregnant women between January 1995 and August 2003. Techniques to minimize fluoroscopy were used, and fluoroscopy times were recorded. Thermoluminescent dosimeters affixed to the skin of the mother were used to estimate fetal radiation exposure. OBSERVATIONS: Mean gestational age was 18.6 (8.9) weeks (range 5-33 weeks). Mean fluoroscopy time was 14 (13) seconds (range 1-48 seconds). Estimated fetal radiation exposure was 40 (46) mrad (range 1-180 mrad). There was a correlation between fluoroscopy time and radiation exposure, but there was a wide range of radiation exposure for individual fluoroscopy times. Complications included post-sphincterotomy bleeding in one patient (controlled by hemoclip placement) and post-ERCP pancreatitis in one patient that necessitated 3 days of hospitalization. Two women developed third-trimester preeclampsia, and labor was induced in both. Thirteen of the 15 patients who delivered were contacted and they confirmed that their child was in good health. CONCLUSIONS: ERCP with modified techniques is safe during pregnancy. Dosimetry should be routinely recorded. PMID- 15278067 TI - EUS-guided FNA of lung masses adjacent to or abutting the esophagus after unrevealing CT-guided biopsy or bronchoscopy. AB - BACKGROUND: The accuracy, the safety, and the cost-effectiveness of EUS-guided FNA for screening patients with lung cancer for mediastinal metastasis are well established, but the utility of EUS-guided FNA in evaluating lung mass per se has not been investigated. This study retrospectively evaluated experience with EUS guided FNA of lung mass lesions after unsuccessful attempts by CT-guided or bronchoscopic tissue sampling to establish a tissue diagnosis. METHODS: A database was searched for all patients who had EUS-guided FNA of lung mass lesions over a 3-year period. The diagnostic yield and safety of EUS-guided FNA were evaluated. OBSERVATIONS: Eighteen patients (11 men, 7 women) underwent EUS guided FNA of lung mass lesions adjacent to or abutting the esophagus. The indication for EUS-guided FNA was evaluation of the mediastinum of patients with lung mass of unclear etiology. EUS-guided FNA yielded tissue for diagnosis in 100% of patients: 15 non-small-cell lung cancer, one small-cell lung cancer, two metastatic lung disease. Ten patients had unresectable disease; in 8, the mass was confined to the lung parenchyma. The mean number of needle passes required to establish a diagnosis was two (range 1-6). No complication was encountered (mean follow-up 141 days; range 72-396 days). Five patients underwent curative surgery, and 13 had palliative chemoradiation. CONCLUSIONS: In this study, EUS-guided FNA of lung mass was safe, and it established a diagnosis in all patients with accessible lesions. Given these preliminary data, a prospective evaluation of this new indication for EUS-guided FNA is justified. PMID- 15278068 TI - Monolobar Caroli's disease complicated by cholangiocarcinoma in a 70-year-old man, previously asymptomatic. PMID- 15278069 TI - Primary squamous cell carcinoma of the bile duct diagnosed by transpapillary cholangioscopy: case report and review. PMID- 15278070 TI - Chronic ischemic proctitis: case report and review. PMID- 15278071 TI - Cap-assisted endoclip placement for recurrent ulcer hemorrhage after repeatedly unsuccessful endoscopic treatment and angiographic embolization: case report. PMID- 15278072 TI - Pemphigus vulgaris with exclusive involvement of the esophagus: case report and review. PMID- 15278073 TI - Peutz-Jeghers-type hamartomatous polyp in a patient without Peutz-Jeghers syndrome. PMID- 15278074 TI - Treatment of refractory ulcerative proctitis with argon plasma coagulation: case report. PMID- 15278075 TI - Endoscopic diagnosis of a congenital ileal duplication cyst. PMID- 15278076 TI - Endoscopically visualized suction catheter food disimpaction from a metallic esophageal stent. PMID- 15278079 TI - Fasciotomy worsens the amount of myonecrosis in a porcine model of crotaline envenomation. AB - STUDY OBJECTIVE: We evaluate the efficacy of fasciotomy or crotaline snake antivenom in reducing myonecrosis. METHODS: We used a randomized, blinded, controlled acute animal preparation. Twenty anesthetized swine were injected intramuscularly in the anterior tibiales muscle of both hind limbs with 6 mg/kg of Crotalus atrox venom (total of 12 mg/kg of venom per animal). Immediately after venom injection, the right hind limb underwent fasciotomy. Muscle biopsies were obtained from the fasciotomized hind limb at 0, 4, and 8 hours and from the other hind limb at the conclusion of the study (8 hours). In addition, animals received either 8 vials of reconstituted Crotalidae polyvalent immune Fab (ovine) (CroFab; FabAV) or an equal volume of normal saline solution intravenously 1 hour after venom injection. A pathologist blinded to the study determined the percentage of myonecrotic cells in each biopsy. Statistical analysis was performed using repeated measures analysis of variance for compartment pressure. Rank-order methods were used for comparison of myonecrosis between groups. RESULTS: Biopsies from hind limbs undergoing fasciotomy revealed a progressive increase in the amount of myonecrosis over time (myonecrosis median at 0, 4, or 8 hours [or death]: 0%, 14%, or 14.5%, respectively; P<.001). Comparison of the amount of myonecrosis of biopsies at death or 8 hours revealed that limbs that underwent fasciotomy had significantly more myonecrosis than those that did not (myonecrosis median: 14.5% versus 2.5%, P=.048). No difference was detected in the amount of myonecrosis when FabAV was compared with normal saline solution on final biopsies from either fasciotomy or nonfasciotomy hind limb (myonecrosis median: 10.0% versus 10.0%, P=.64). CONCLUSION: Fasciotomy significantly worsens the amount of myonecrosis in a porcine model of intramuscular crotaline venom injection. No change in the amount of myonecrosis was detected with the use of FabAV treatment at the dosages used in this animal model. PMID- 15278080 TI - Can steel heal a compartment syndrome caused by rattlesnake venom? PMID- 15278081 TI - Comparison of consciousness level assessment in the poisoned patient using the alert/verbal/painful/unresponsive scale and the Glasgow Coma Scale. AB - STUDY OBJECTIVE: We determine how the alert/verbal/painful/unresponsive (AVPU) responsiveness scale (alert, responsive to verbal stimulation, responsive to painful stimulation, and unresponsive) corresponds to the Glasgow Coma Scale (GCS) when assessing consciousness level in the poisoned patient. METHODS: Consciousness level was assessed using the AVPU responsiveness scale and the GCS in all patients admitted to the hospital during a 6-month period with deliberate or accidental poisoning. An AVPU responsiveness scale algorithm and details of the individual components of the GCS were provided. Data were recorded prospectively on admission to the toxicology ward by nursing staff in the majority of cases and from case records for the small number of patients admitted directly to the ICU. Nursing staff also recorded any difficulty assessing consciousness level using either scoring system. RESULTS: Of the 1,384 patients studied, 1,138 patients were alert, 114 patients responded to a verbal stimulus, 87 patients responded to a painful stimulus, and 15 patients were unresponsive. The median GCS scores with interquartile ranges (IQR) for each AVPU responsiveness category were 15 (IQR 15), 13 (IQR 12 to 14), 8 (IQR 7 to 9), and 3 (IQR 3), respectively. There was a degree of overlap between the range of GCS scores for each category. Nursing staff recorded more difficulty using the GCS than the AVPU responsiveness scale. Alcohol-intoxicated patients proved to be the most difficult to assess. All patients who were unresponsive required intubation. No patient with a GCS score greater than 6 was intubated. CONCLUSION: Each AVPU category can be shown to correspond to a range of GCS scores. The AVPU responsiveness scale appears to provide a rapid simple method of assessing consciousness level in most poisoned patients, but difficulty was still observed in assessing alcohol-intoxicated patients. PMID- 15278082 TI - The Meixner test in the detection of alpha-amanitin and false-positive reactions caused by psilocin and 5-substituted tryptamines. AB - STUDY OBJECTIVE: The Meixner test has been suggested to identify the presence of alpha-amanitin, one of the toxic compounds in Amanita mushrooms. We attempted to determine the detection limit of the Meixner test for alpha-amanitin and to determine the percentage of positive sample interpretation compared with other mushroom indole compounds. METHODS: This was a 2-part in vitro experiment. In part 1, we applied the Meixner test to a series of dilutions of alpha-amanitin (0 microg, 0.8 microg, 1.0 microg, 2.0 microg, and 4.0 microg) on telephone book paper, which were then presented to 5 blinded emergency physicians. We sought to determine the lowest amount of alpha-amanitin that was universally recognized as positive by the physicians (the detection limit). In the second part, 5 emergency physicians were presented simultaneously with 10 Meixner processed samples, including the mushroom indole compounds alpha-amanitin (2 microg and 10 microg), psilocin (20 microL and 60 microL of mushroom extract), 5-hydroxytryptamine (100 microg and 200 microg), and 5-methoxy-N,N-dimethyltryptamine (100 microg and 200 microg), as well as 2 negative controls (20 microL of water and methanol). We determined how likely these other indoles are to be mistaken for a positive alpha amanitin Meixner reaction result by determining the percentage of positive sample interpretation for each compound and comparing them with the rate for alpha amanitin. Fisher's exact test was used to determine any significant difference (P<.05) between the samples. RESULTS: The minimum amount of alpha-amanitin that was identified with 100% agreement by testers was 2 microg. For the second part, there was 100% agreement that psilocin gives a positive Meixner test (100% false positive) and a 35% recognition rate of the 5-substituted tryptamine compounds as a positive Meixner test. There was no statistical difference between the interpretation of alpha-amanitin and psilocin, suggesting the test is unable to differentiate between them. CONCLUSION: Although the Meixner test has a good detection limit for toxic amounts of alpha-amanitin, a positive Meixner reaction does not adequately distinguish between alpha-amanitin and other mushroom indoles. PMID- 15278083 TI - A novel pain management strategy for combat casualty care. AB - STUDY OBJECTIVE: Pain control in trauma patients should be an integral part of the continuum of care, beginning at the scene with out-of-hospital trauma management, sustained through the evacuation process, and optimized during hospitalization. This study evaluates the effectiveness of a novel application of a pain control medication, currently indicated for the management of chronic and breakthrough cancer pain, in the reduction of acute pain for wounded Special Operations soldiers in an austere combat environment. METHODS: Doses (1,600 microg) of oral transmucosal fentanyl citrate were administered by medical personnel during missions executed in support of Operation Iraqi Freedom from March 3, 2003, to May 3, 2003. Hemodynamically stable casualties presenting with isolated, uncomplicated orthopedic injuries or extremity wounds who would not have otherwise required an intravenous catheter were eligible for treatment and evaluation. Pretreatment, 15-minute posttreatment, and 5-hour posttreatment pain intensities were quantified by the verbal 0-to-10 numeric rating scale. RESULTS: A total of 22 patients, aged 21 to 37 years, met the study criterion. The mean difference in verbal pain scores (5.77; 95% confidence interval [CI] 5.18 to 6.37) was found to be statistically significant between the mean pain rating at 0 minutes and the rating at 15 minutes. However, the mean difference (0.39; 95% CI 0.18 to 0.96) was not statistically significant between 15 minutes and 5 hours, indicating the sustained action of the intervention without the need for redosing. One patient experienced an episode of hypoventilation that resolved readily with administration of naloxone. Other documented adverse effects were minor and included pruritus (22.7%), nausea (13.6%), emesis (9.1%), and lightheadedness (9.1%). CONCLUSION: Oral transmucosal fentanyl citrate can provide an alternative means of delivering effective, rapid-onset, and noninvasive pain management in an out-of-hospital, combat, or austere environment. PMID- 15278084 TI - Emergency medicine research on the front lines. PMID- 15278085 TI - Nebulized lidocaine decreases the discomfort of nasogastric tube insertion: a randomized, double-blind trial. AB - STUDY OBJECTIVE: Nasogastric tube insertion is a common emergency department (ED) procedure that is associated with considerable patient discomfort. The safety and efficacy of nebulized lidocaine for upper airway anesthesia have previously been demonstrated. We determine whether nebulized lidocaine administered before nasogastric tube insertion significantly reduces patient discomfort. METHODS: A double-blind, placebo-controlled, randomized clinical trial of adult patients was conducted in the EDs of 2 university hospitals. Twenty-nine participants were administered nebulized lidocaine (4 mL 10%), and 21 participants received nebulized normal saline solution. Patient discomfort was measured using a 100-mm visual analog scale. The difficulty of nasogastric tube insertion was evaluated using a 5-point Likert scale. RESULTS: There was a clinical and statistical significant difference in patient discomfort associated with the passage of the nasogastric tube between nebulized lidocaine and placebo groups (mean visual analog scale score 37.7 versus 59.3 mm, respectively; difference between group means 21.6 mm; 95% confidence interval [CI] 5.3 to 38.0 mm). There was not a detectable difference in difficulty with the passage of the nasogastric tube between the 2 groups (median 2 versus 2; median difference 0; 95% CI -1 to 1). Epistaxis occurred more frequently in the lidocaine group (17% versus 0%; difference 17%; 95% CI 3.5% to 31%). CONCLUSION: Nebulized lidocaine decreases the discomfort of nasogastric tube insertion and should be considered before passing a nasogastric tube. An increased frequency of epistaxis, however, may be associated with its use. PMID- 15278086 TI - Nasogastric tubes: hard to swallow. PMID- 15278087 TI - Lateral automobile impacts and the risk of traumatic brain injury. AB - STUDY OBJECTIVES: We determine the relative risk and severity of traumatic brain injury among occupants of lateral impacts compared with occupants of nonlateral impacts. METHODS: This was a secondary analysis of the National Highway Traffic Safety Administration's National Automotive Sampling System, Crashworthiness Data Systems for 2000. Analysis was restricted to occupants of vehicles in which at least 1 person experienced an injury with Abbreviated Injury Scale score greater than 2. Traumatic brain injury was defined as an injury to the head or skull with an Abbreviated Injury Scale score greater than 2. Outcomes were analyzed using the chi2 test and multivariate logistic regression, with adjustment of variance to account for weighted probability sampling. RESULTS: Of the 1,115 occupants available for analysis, impact direction was lateral for 230 (18.42%) occupants and nonlateral for 885 (81.58%) occupants. One hundred eighty-seven (16.07%) occupants experienced a traumatic brain injury, 14.63% after lateral and 16.39% after nonlateral impact. The unadjusted relative risk of traumatic brain injury after lateral impact was 0.89 (95% confidence interval [CI] 0.51 to 1.56). After adjusting for several important crash-related variables, the relative risk of traumatic brain injury was 2.60 (95% CI 1.1 to 6.0). Traumatic brain injuries were more severe after lateral impact according to Abbreviated Injury Scale and Glasgow Coma Scale scores. The proportion of fatal or critical crash-related traumatic brain injuries attributable to lateral impact was 23.5%. CONCLUSION: Lateral impact is an important independent risk factor for the development of traumatic brain injury after a serious motor vehicle crash. Traumatic brain injuries incurred after lateral impact are more severe than those resulting from nonlateral impact. Vehicle modifications that increase head protection could reduce crash-related severe traumatic brain injuries by up to 61% and prevent up to 2,230 fatal or critical traumatic brain injuries each year in the United States. PMID- 15278088 TI - Risk analysis in road traffic injury research. PMID- 15278089 TI - Unprecedented nationwide paid media campaign on impaired driving yields strong results: "You Drink and Drive. You Lose." campaign scores with target market. PMID- 15278090 TI - Commentary: "You Drink and Drive. You Lose." Reaching the target audience is not enough. PMID- 15278091 TI - Ultrasonography of the internal jugular vein in patients with dyspnea without jugular venous distention on physical examination. AB - STUDY OBJECTIVE: Accurate physical examination of patients with dyspnea is important. Jugular venous distention, however, can be difficult to assess in patients. The purpose of this case series is to serve as a pilot study of how ultrasonographic examination of the internal jugular vein compares with other measures of dyspnea. METHODS: This was a case series of 8 patients presenting with dyspnea without jugular venous distention on physical examination. Each patient underwent ultrasonographic examination of the internal jugular vein and inferior vena cava by an emergency physician sonographer blinded to all other clinical information after initial evaluation by another emergency physician for dyspnea. Results of ultrasonographic examination of the internal jugular vein and inferior vena cava were subsequently compared with initial emergency physician physical examination findings, initial chest radiography interpreted by radiologists, initial B-type natriuretic peptide levels, and final hospital discharge diagnosis. RESULTS: Ultrasonographic examination of the internal jugular vein compared more favorably with B-type natriuretic peptide levels and chest radiographic findings than ultrasonographic examination of the inferior vena cava in these patients with dyspnea but not jugular venous distention on physical examination. It was able to identify every patient diagnosed with cardiogenic pulmonary edema on hospital discharge. CONCLUSION: Ultrasonographic examination of the internal jugular vein appears to be helpful in patients who present with dyspnea but do not have evidence of jugular venous distention on physical examination. PMID- 15278092 TI - Presenting at journal club: A guide. PMID- 15278093 TI - Is doing "everything" enough? PMID- 15278094 TI - Management of recurrent disease after radical prostatectomy. AB - Prostate cancer has undergone a stage migration since the advent of widespread PSA testing, yet still a significant number of men develop PSA recurrence following radical prostatectomy. This causes anxiety to the patient and the urologist. This review examines the clinical significance of biochemical relapse and the role of imaging modalities and anastomotic biopsies. The importance of the radical prostatectomy pathological features and the PSA kinetics in determining the site of recurrence and the best treatment modality is emphasised. The optimal timing and dose of salvage radiotherapy and the role of hormonal therapy is discussed. PMID- 15278095 TI - Timing and choice of androgen ablation. AB - Hormone therapy remains central to the management of advanced prostate cancer. Evidence has accrued to suggest benefits from earlier treatment. Trials which demonstrate significant improvements in disease-specific survival may be underpowered to show overall survival benefits, due to the effect of mortality from unrelated causes. Toxicity, particularly the risk of osteoporosis, causes increasing concern, particularly where many years of use may be contemplated as in early relapse after curative treatment. Strategies to reduce toxicity include use different drugs, notably antiandrogens and with renewed interest in oestrogens in low dose or administered parenterally, and deferred and intermittent treatment. Further improvements in drug development, reducing toxicity, and identification of the cause of hormone refractory disease and its prevention would revolutionise the use of hormone therapy. Currently, deferred treatment, used selectively and with careful follow-up, remains an option for selected patients. PMID- 15278096 TI - Postradical prostatectomy TRUS-guided anastomotic biopsy. Where do we stand today? AB - The issue of performing tissue sampling from the vesicourethral anastomotic area postradical prostatectomy (transrectal ultrasound-guided biopsy) after radical surgical treatment of local disease has failed, still remains controversial. We review a selection of articles that evaluate this procedure as well as newer diagnostic modalities and we discuss how this technique may have a position in our treatment dilemmas in cases with biochemical failure of undetermined origin. PMID- 15278097 TI - Glutamate as a therapeutic target in psychiatric disorders. AB - Glutamate is the primary excitatory neurotransmitter in the mammalian brain. Glutamatergic neurotransmission may be modulated at multiple levels, only a minority of which are currently being exploited for pharmaceutical development. Ionotropic receptors for glutamate are divided into N-methyl-D-aspartate receptor (NMDAR) and AMPA receptor subtypes. NMDAR have been implicated in the pathophysiology of schizophrenia. The glycine modulatory site of the NMDAR is currently a favored therapeutic target, with several modulatory agents currently undergoing clinical development. Of these, the full agonists glycine and D-serine have both shown to induce significant, large effect size reductions in persistent negative and cognitive symptoms when added to traditional or newer atypical antipsychotics in double-blind, placebo-controlled clinical studies. Glycine (GLYT1) and small neutral amino-acid (SNAT) transporters, which regulate glycine levels, represent additional targets for drug development, and may represent a site of action of clozapine. Brain transporters for D-serine have recently been described. Metabotropic glutamate receptors are positively (Group I) or negatively (Groups II and III) coupled to glutamatergic neurotransmission. Metabotropic modulators are currently under preclinical development for neuropsychiatric conditions, including schizophrenia, depression and anxiety disorders. Other conditions for which glutamate modulators may prove effective include stroke, epilepsy, Alzheimer disease and PTSD. PMID- 15278098 TI - Inhibition of system A-mediated glycine transport in cortical synaptosomes by therapeutic concentrations of clozapine: implications for mechanisms of action. AB - Clozapine is an atypical antipsychotic with particular efficacy in schizophrenia, possibly related to potentiation of brain N-methyl-D-aspartate receptor (NMDAR) mediated neurotransmission. NMDARs are regulated in vivo by glycine, which is regulated in turn by glycine transporters. The present study investigates transport processes regulating glycine uptake into rat brain synaptosomes, along with effects of clozapine on synaptosomal glycine transport. Amino-acid uptake of amino acids was assessed in rat brain P2 synaptosomal preparations using a radiotransport assay. Synaptosomal glycine transport was inhibited by a series of amino acids and by the selective System A antagonist MeAIB (2-methyl aminoisobutyric acid). Clozapine inhibited transport of both glycine and MeAIB, but not other amino acids, at concentrations associated with preferential clinical response (0.5-1 microg/ml). By contrast, other antipsychotics studied were ineffective. The novel glycine transport inhibitor N[3-(4'-fluorophenyl)-3 (4'-phenylphenoxy)propyl]sarcosine (NFPS) produced biphasic inhibition of [(3)H]glycine transport, with IC(50) values of approximately 25 nM and 25 microM, respectively. NFPS inhibition of [(3)H]MeAIB was monophasic with a single IC(50) value of 31 microM. Clozapine significantly inhibited [(3)H]glycine binding even in the presence of 100 nM NFPS. In conclusion, this study suggests first that System A transporters, or a subset thereof, may play a critical role in regulation of synaptic glycine levels and by extension of NMDA receptor regulation, and second that System A antagonism may contribute to the differential clinical efficacy of clozapine compared with other typical or atypical antipsychotics. PMID- 15278099 TI - C957T polymorphism of the dopamine D2 receptor (DRD2) gene affects striatal DRD2 availability in vivo. PMID- 15278100 TI - Chronic central administration of ghrelin reverses the effects of leptin. AB - OBJECTIVE: To determine whether chronic central administration of ghrelin can block the effects of leptin on food intake, adiposity, and plasma concentrations of metabolic parameters and hormones. DESIGN: Intracerebroventricular (ICV) infusions of leptin (5 microg/day) for 7 days, with or without ghrelin (1.2 microg/day), in rats. Rats administered leptin plus ghrelin were divided into ad lib-fed and food-restricted groups. MEASUREMENT: Body weight and food intake were monitored daily. Following killing on day 8, epididymal fat weight and fasting plasma concentrations of glucose, insulin, leptin, ghrelin, IGF-1, and adiponectin were determined. RESULTS: ICV infusion of leptin decreased food intake by 39% and fat weight by 41%. Leptin decreased plasma concentrations of glucose, insulin, and leptin and increased plasma ghrelin levels. Central coadministration of ghrelin blocked the effects of leptin. Most of the effects of ghrelin were diminished by food restriction but ghrelin effect on adiposity and plasma insulin concentrations remained in food-restricted rats. CONCLUSION: Chronic central administration of ghrelin reversed the effects of leptin, primarily by altering food intake, but ghrelin may have regulatory effects on adiposity and plasma insulin levels independent of feeding effect. PMID- 15278101 TI - The influence of the stigma of obesity on overweight individuals. AB - OBJECTIVE: To investigate the internalization of anti-fat bias among overweight individuals across a variety of attitudes and stereotypes. DESIGN: Two studies were conducted using the Implicit Association Test (IAT), a performance-based measure of bias, to examine beliefs among overweight individuals about 'fat people' vs 'thin people'. Study two also contained explicit measures of attitudes about obese people. SUBJECTS: Study 1 participants were 68 overweight patients at a treatment research clinic (60 women, 8 men; mean Body Mass Index (BMI) of 37.1+/-3.9 kg/m(2)). Study 2 involved 48 overweight participants (33 women, 15 men) with a BMI of 34.5+/-4.0 kg/m(2). RESULTS: Participants exhibited significant anti-fat bias on the IAT across several attributes and stereotypes. They also endorsed the explicit belief that fat people are lazier than thin people. CONCLUSION: Unlike other minority group members, overweight individuals do not appear to hold more favorable attitudes toward ingroup members. This ingroup devaluation has implications for changing the stigma of obesity and for understanding the psychosocial and even medical impact of obesity on those affected. PMID- 15278102 TI - Night eating and weight change in middle-aged men and women. AB - OBJECTIVE: To examine the association between the habit of eating at night, and the 5-y preceding and 6-y subsequent weight changes in a middle-aged population, with particular focus on the obese. DESIGN: Prospective study with initial examination of the cohort in 1982-83, re-examination in 1987-88 and a third examination in 1992-93. SUBJECTS: The Danish MONICA cohort includes an age- and sex-stratified random sample of the population from the Western part of the Copenhagen County. Out of 2,987 subjects participating in 1987-88, a total of 1,050 women and 1,061 men had been examined in 1982-83, and 1993-94 too. Subjects working night shifts were excluded. MEASUREMENTS: Night eating in 1987-88, 5-y preceding and 6-y subsequent weight change. RESULTS: In total, 9.0% women and 7.4% men reported 'getting up at night to eat'. Obese women with night eating experienced an average 6-y weight gain of 5.2 kg (P=0.004), whereas only 0.9 kg average weight gain was seen among obese women who did not get up at night to eat. No significant associations were found among all women, or between night eating and the 5-y preceding weight change for women. Night eating and weight change were not associated among men. CONCLUSION: Night eating was not associated with later weight gain, except among already obese women, suggesting that getting up at night to eat may be a contributor to further weight gain among the obese. PMID- 15278103 TI - Hepatic steatosis in obese Chinese children. AB - OBJECTIVES: THE AIMS OF OUR STUDY WERE: (1) to determine the prevalence of asymptomatic hepatic steatosis and presumed nonalcoholic steatohepatitis, in our local population of obese Chinese children referred for medical assessment; and (2) to assess the correlation between severity of ultrasonographic hepatic steatosis and degree of obesity, insulin resistance and serum biochemical abnormalities. DESIGN: Cross-sectional study. METHODS: In total, 84 obese children, 25 girls and 59 boys with median age and body mass index (BMI) of 12.0 years (interquartile range (IR): 9.5-14.0) and 30.3 kg/m(2) (IR: 27.1-33.4), respectively, referred for medical assessment were studied. All subjects underwent physical examination, anthropometric and dual energy X-ray absorptiometry (DEXA) scan measurements and real-time ultrasonographic (US) examination of the liver. Fasting blood samples were collected for the measurement of liver function, hepatitis status, levels of serum glucose and insulin and lipid profile. Degree of fatty infiltration of the liver was graded according to ultrasonic appearance of liver echotexture, liver-diaphragm differentiation in echo amplitude, hepatic echo penetration and clarity of hepatic blood vessels. RESULTS: All recruited subjects had no history of alcohol abuse and tests for Hepatitis B or C virus were negative. Thorough examination showed all of them to be in general good health without signs of chronic liver disease. Hepatic steatosis identified by defined ultrasonic appearances was diagnosed in 65 subjects (77%); 17 girls and 48 boys. The severity of fatty liver was positively related to anthropometric measurements including BMI, waist and hip circumference, subscapular skinfold thickness; insulin resistance markers [QUICKI and homeostasis model assessment (HOMA)], and hypertriglyceridaemia. Multvariate ordinal regression analysis showed that BMI and raised alanine aminotransferase (ALT) were positively associated with fatty liver. Combination of hepatic steatosis with raised ALT (presumptive NASH) was found in 19 subjects (24%). This group of patients had significantly higher waist hip ratio and conicity index compared to those with isolated hepatic steatosis. Boys with presumed NASH were also found to have significantly higher insulin resistance. CONCLUSION: Nonalcoholic fatty liver disease (NAFLD) was common among our cohort of obese children referred for medical assessment. The prevalence of simple steatosis and presumed NASH was 77 and 24%, respectively. The severity of US steatosis was positively correlated with BMI, raised ALT, insulin resistance and hypertryglyceridaemia. Ultrasonography being noninvasive and readily available could be used for the monitoring of the progression of hepatic steatosis. Further longitudinal studies are required to determine the natural disease progression and the role of insulin resistance and other factors in the pathophysiology of NAFLD. PMID- 15278104 TI - Relation of BMI to fat and fat-free mass among children and adolescents. AB - OBJECTIVE: Although the body mass index (BMI, kg/m2) is widely used as a surrogate measure of adiposity, it is a measure of excess weight, rather than excess body fat, relative to height. We examined the relation of BMI to levels of fat mass and fat-free mass among healthy 5- to 18-y-olds. METHODS AND PROCEDURES: Dual-energy X-ray absorptiometry was used to measure fat and fat-free mass among 1196 subjects. These measures were standardized for height by calculating the fat mass index (FMI, fat mass/ht2) and the fat-free mass index (FFMI, fat-free mass/ht2). RESULTS: The variability in FFMI was about 50% of that in FMI, and the accuracy of BMI as a measure of adiposity varied greatly according to the degree of fatness. Among children with a BMI-for-age > or =85th P, BMI levels were strongly associated with FMI (r=0.85-0.96 across sex-age categories). In contrast, among children with a BMI-for-age <50th P, levels of BMI were more strongly associated with FFMI (r=0.56-0.83) than with FMI (r=0.22-0.65). The relation of BMI to fat mass was markedly nonlinear, and substantial differences in fat mass were seen only at BMI levels > or =85th P. DISCUSSION: BMI levels among children should be interpreted with caution. Although a high BMI-for-age is a good indicator of excess fat mass, BMI differences among thinner children can be largely due to fat-free mass. PMID- 15278105 TI - Is serum leptin related to physical function and is it modifiable through weight loss and exercise in older adults with knee osteoarthritis? AB - OBJECTIVE: To determine the effect of weight loss and exercise interventions on serum leptin and to investigate the relationship of physical function and osteoarthritis (OA) severity with serum leptin in older overweight and obese adults with knee OA. In addition, the study examined if serum leptin predicts weight loss. DESIGN: Longitudinal, controlled clinical trial of weight loss and exercise interventions. SUBJECTS: Community dwelling, older, overweight and obese adults (n=316; >60 years of age; body mass index >/=28.0 kg m(-2)) with symptomatic knee OA and self-reported difficulty in performing selected physical activities were recruited. INTERVENTIONS: Participants were randomized into one of four groups for the 18-month study duration: Healthy Lifestyle Controls, Dietary Weight Loss (Diet), Exercise Training (Exercise), and a combination of Dietary Weight Loss and Exercise Training (Diet+Exercise). The weight loss goal for the two Diet groups was 5% from baseline at 18 months. Participants in the Exercise groups were trained for 3 days week(-1), 60 min day(-1). MEASUREMENTS: Body weight, body mass index, serum leptin, physical function, and OA severity were measured at baseline, 6 months, and 18 months. RESULTS: Diet and Diet+Exercise groups lost 5.3 and 6.1% of their weight, respectively, at 18 months with the Exercise group losing 2.9%. There was a significant main effect of weight loss on serum leptin with a decrease in serum leptin averaged across the 6- and 18-month time points for the Diet and Diet+Exercise groups compared to the other two groups (beta=0.245; P<0.01). No main effect for exercise training was observed. Serum leptin was related to self-reported physical function. In all participants, a mixed model analysis demonstrated that lower levels of baseline serum leptin predict larger weight loss (beta=-2.779; P=0.048). CONCLUSION: Decreases in serum leptin may be one mechanism by which weight loss improves physical function and symptoms in OA patients. PMID- 15278106 TI - Differences between recumbent length and stature measurement in groups of 2- and 3-y-old children and its relevance for the use of European body mass index references. AB - OBJECTIVE: To compare length and stature measurements of young children and to examine the relevance of any difference for comparison with body mass index (BMI) references designed for use from birth to adulthood. SUBJECTS: A total of 426 2-y old and 525 3-y-old children included in the Dortmund Nutritional and Anthropometrical Longitudinally Designed (DONALD) Study. DATA ANALYSIS: Length and stature were measured to the nearest millimetre using a stadiometre. Agreement between both measurements at age 2 and 3 y, respectively, was determined by mean differences and by comparison with the German BMI reference. RESULTS: The average length of 2-y-old girls and boys was 88.3 (3.1) and 89.9 (3.2) cm, mean differences (stature minus length) were -0.47 (0.65) and -0.45 (0.64) cm. The corresponding BMI values were 16.18 (1.3) and 16.46 (1.2) kg/m2, with mean differences of +0.17 (0.24) and +0.16 (0.23). According to stature, 9.4% of the girls and 10.8% of the boys were overweight (>90th percentile), while length classified 7.1 and 9.4% as overweight. Similar mean differences between length and stature were observed at age 3 y: -0.53 (0.62) and -0.47 (0.65) cm in height and +0.17 (0.20) and +0.14 (0.20) kg/m2 in the BMI of girls and boys, respectively. According to stature, 7.6 and 7.3% were overweight as opposed to 5.4 and 4.8% using length. The observed differences increased with higher BMI levels. CONCLUSION: Changing measurements from length to stature results in an upward shift of BMI, not reflected in current European BMI references. This small but systematic error may result in misinterpretation of individual BMI levels or trend observations. PMID- 15278107 TI - Perceptions of healthy and desirable body size in urban Senegalese women. AB - OBJECTIVE: To examine the cultural ideals for body size held by urban Senegalese women; to determine the body size that women associate with health; and to estimate the change in prevalence of female obesity in an urban neighbourhood of Dakar. DESIGN: Cross-sectional, population-based study in the subject's home, using a structured interviewer-administered questionnaire, conducted in the same Dakar neighbourhood as that of a previous survey conducted in 1996. SUBJECTS: A total of 301 randomly selected women, aged 20-50 y, living in a specific Dakar neighbourhood, Senegal. MEASUREMENTS: A total of 32 items concerning body satisfaction, social status, health and individual attributes to associate with one of six photographic silhouettes; body mass index (BMI), waist circumference, waist-to-hip ratio by anthropometry; and measures of economic status. RESULTS: In all, 26.6% of women were overweight (BMI 25-29.9 kg/m2) and 18.6% were obese (BMI > or =30 kg/m2) compared with 22.4 and 8.0% respectively in 1996. Overweight was the most socially desirable body size, although obesity itself was seen as undesirable, associated with greediness and the development of diabetes and heart disease. Lay definitions of overweight and normal weight differed substantially from health definitions, as one-third of the sample saw the 'overweight' category as normal. Over a third of women with BMI > or =25 kg/m2 wanted to gain more weight. CONCLUSION: There has been a sharp rise in the prevalence of obesity in Senegalese women living in a Dakar neighbourhood over the last 7 y. In general, overweight body sizes (but not obese) were seen in a positive light. The finding that the term 'overweight' made little sense to these Senegalese women could have important implications for developing public health policies. PMID- 15278108 TI - Physical activity is inversely related to waist circumference in 12-y-old French adolescents. AB - BACKGROUND AND OBJECTIVE: Waist circumference (W) has been shown to be a good predictor of cardiovascular risk. The aim of this study was to investigate whether physical activity (PA) is related to W in adolescents as previously shown in adults. DESIGN AND SUBJECTS: Cross-sectional population-based survey of 2,714 12-y-old adolescents from the eastern part of France. MEASUREMENTS: Body mass index (BMI) and W were measured. Structured PA, active commuting to and from school and sedentary activities (SED), for example television viewing, computer/video games and reading and different potential confounders (dietary habits, parental overweight, family annual income tax and educational level) were assessed by a questionnaire. RESULTS: The adolescents had a mean BMI of 19.0+/ 3.4 kg/m2, and 20.2% of them were overweight, with no gender difference. Boys had a greater W than girls (67.6+/-9.1 vs 65.7+/-8.9 cm, P<0.0001). In all, 42% of the girls and 25% of the boys did not practice any structured PA outside school and less than 40% of the adolescents commuted actively to school more than 20 min/day. About one-third of the adolescents devoted more than 2 h/day to SED. In univariate analyses, BMI was negatively associated with structured PA but significantly only for girls (P<0.01) and positively associated with SED for both genders (P<0.0001 for girls, P<0.01 for boys). W was negatively associated with structured PA and positively associated with SED both in girls (P<0.0001 and P=0.03, respectively) and boys (P<0.01 and P=0.08, respectively). Multiple general linear models show that SED is associated with BMI, independently of structured PA, in both genders. On the other hand, structured PA was inversely associated with W, independently of SED. The inverse relation between structured PA and W persisted after additional adjustment on BMI, with a greater effect of PA for the adolescents with higher BMI. CONCLUSIONS: In 12-y-old adolescents, structured PA is inversely associated with W, an indicator of total adiposity but also more specifically of abdominal fat. This suggests that PA may have a beneficial effect on youth metabolic and cardiovascular risks, in particular in the presence of overweight.. PMID- 15278109 TI - Anionic NaCl-type frameworks of [Mn(II)(HCOO)(3)(-)], templated by alkylammonium, exhibit weak ferromagnetism. AB - We present the synthesis, characterization by IR, TGA, single crystal X-ray structure and magnetic properties of a novel series of NaCl-type frameworks of [AmineH(+)][Mn(HCOO)(3)(-)], templated by alkylammonium. The anionic NaCl framework of [Mn(HCOO)(3)(-)] is counter-balanced by the alkylammonium cations located in the cavities of the framework to which they are hydrogen-bonded. The divalent manganese ions have octahedral geometry and are bridged by the formate in an anti-anti mode of coordination. All the compounds exhibit long-range antiferromagnetism below 9 K with a slight non-collinear arrangement of the moments. The canting, likely due to second-order spin-orbit coupling, is via a Dzyaloshinski-Moriya antisymmetric exchange mechanism. A spin-flop is observed in each case at fairly low fields. An orthorhombic to monoclinic transformation was observed for the protonated cyclotrimethyleneamine that is accompanied by localization of the cations into two positions below 240 K from the rapid dynamic flipping of the ring observed at room temperature. PMID- 15278110 TI - Crystal-to-crystal transformations of a microporous metal-organic laminated framework triggered by guest exchange, dehydration and readsorption. AB - Single crystals of a neutral, microporous, laminated metal-organic framework (MOF)[Fe(pydc)(4,4'-bipy)]xH(2)O (1xH(2)O)(H(2)pydc = 2,5-dicarboxypyridine, 4,4' bipy = 4,4'-bipyridine) were generated by hydrothermal synthesis, and its crystal structure was determined. 1xH(2)O retains the framework robustness to ca. 370 degrees C and is insoluble in common organic solvents. By soaking in MeOH and EtOH solutions, 1xH(2)O was transformed directly from the parent single crystals into single crystals of 1xMeOH or 1xEtOH, respectively. Meanwhile, 1xH(2)O shrank to the guest-free framework (h) or (v), respectively, under appropriate heating (up to 160 degrees C in N(2)) or vacuum treatment (10 mmHg) at room temperature. Compared to that of 1xH(2)O, the unit-cell volume of 1xEtOH slightly increases by 2.9%, whereas those of (h) or (v) are reduced by 8.2 and 6.6%, respectively. The anhydrous (v) was found to be highly chemically reactive, taking up ethanol vapor to furnish the solvated crystal structure of an 'expanded' framework 1xEtOH. In a mixture of ethanol-DMF or ethanol-benzene, a selective exchange process was observed, with only ethanol molecules exchanged into the structure due to the limited free size of the channels in the framework of. All the transformed crystals have also been characterized by X-ray single-crystal diffraction to understand the crystal-to-crystal transformation, which have different free volumes (6.5-20.4%). PMID- 15278111 TI - The NaNO(3)/KNO(3) system: the position of the solidus and sub-solidus. AB - The existing evidence for the nature of the phase diagram for the binary system sodium nitrate-potassium nitrate is reviewed and in particular whether the system is of the continuous solid solution type, as has often been stated in the last 80 years, or whether this system is of the eutectic type as was earlier believed and has again been asserted recently. Additional evidence from Raman spectroscopy and Raman mapping on the 50 : 50 mol%(minimum melting point) composition is now presented, supporting the eutectic classification. Abrupt changes in wavenumber, or in the wavenumber-temperature gradient of five Raman bands indicate a solid state transition at about 115 degrees C and are attributed to a phase transition in KNO(3)-rich areas. On a fast cooled sample, Raman bands attributed to sodium nitrate-rich and potassium nitrate-rich areas were found to persist up to and slightly beyond the melting point, and although their wavenumber-positions converged, the apparent single band could still be resolved into the two bands which could be attributed to the Na-rich and K-rich areas. On cooling, the reverse change took place quickly. Measurements with the initially slow cooled sample, where these areas were bigger, showed that the spectral bands reverted to the room-temperature wavenumber values, after holding at 22 degrees C for only 60 90 min. PMID- 15278112 TI - Zinc complexes of T-shaped trans-1,2,3-propenetricarboxylic acid with 1-D ribbon like chain, 2-D rhombus-grid-like and herringbone-like layers, and non interpenetrating 3-D open framework. AB - Five zinc complexes of T-shaped trans-1,2,3-propenetricarboxylic acid (H(3)L) with 1-D ribbon-like chain, 2-D rhombus-grid-like and herringbone-like layers, and non-interpenetrating 3-D open framework have been obtained based on different building blocks. X-ray single crystal analyses show that the T-shaped information of H(3)L was nicely expressed in the construction of herringbone-like or ladder like structure. All compounds were found to display blue fluorescent emissions. PMID- 15278113 TI - A high-throughput approach to lanthanide complexes and their rapid screening in the ring opening polymerisation of caprolactone. AB - Libraries of lanthanide complexes supported by nitrogen and oxygen containing ligands have been synthesised using a high-throughput approach. The complexes were employed in the ring-opening polymerisation of epsilon-caprolactone, in some cases giving polycaprolactone of controlled molecular weight and narrow polydispersity. The libraries, based on twenty-one ligands and eight lanthanide reagents, were developed in order to determine the best combination of lanthanide metal and ligand. They were prepared via transamination reactions of [Ln[N(SiMe(3))(2)](3)] complexes with tetradentate dianionic ligands containing oxygen and nitrogen donors. 1H NMR spectroscopy was used to screen polymerisation activity. The steric demand of the ligand has a significant effect on the polymerisation process, as do the type of nitrogen donor and the size of the central Ln(3+) ion. Ligands containing aryl rings with bulky substituents such as tert-pentyl groups afforded species capable of performing controlled polymerisation of caprolactone, whereas less bulky groups such as methyl were not effective. Yttrium and mid-sized lanthanides such as samarium showed increased activity compared with the larger lanthanides, lanthanum and praseodymium, and the smaller lanthanides like ytterbium. X-ray crystal structures of a sterically demanding chelating amine-bis((2-hydroxyaryl)methyl) ligand and a chloride bridged dinuclear gadolinium complex are reported. The centrosymmetric molecule contains gadolinium in distorted capped trigonal prismatic environments bonded to two amine, two phenolate, one THF and two chloride donors. PMID- 15278114 TI - Hydrosilylation of dienes by yttrium hydrido complexes containing a linked amido cyclopentadienyl ligand. AB - The dimeric hydrido complex [Y(L)(THF)(mu-H)](2)() containing the CH(2)SiMe(2) linked amido-cyclopentadienyl ligand L = C(5)Me(4)CH(2)SiMe(2)NCMe(3)(2-) catalyzed the hydrosilylation of 1,5-hexadiene, 1,7-octadiene and vinylcyclohexene by PhSiH(3). As demonstrated for 1,7-octadiene, the product distribution of the hydrosilylation strongly depends on the molar ratio of the reagents. In the absence of PhSiH(3), the stoichiometric reaction of with 1,5 hexadiene gave the isolable crystalline cyclopentylmethyl complex [Y(L)[CH(2)CH(CH(2))(4)](THF)]. Internal olefins such as trans-stilbene and alkynes such as tert-butylacetylene were not hydrosilylated by. trans-Stilbene was inserted into the yttrium-hydride bond of to give the 1,2-diphenylethyl complex [Y(L)[CH(CH(2)Ph)Ph](THF)]. tert-Butylacetylene reacted with to give the dimeric acetylide [Y(L)(C[triple bond]CCMe(3))](2). In an attempt to detect the monomeric hydrido species as a DME adduct [Y(L)H(DME)], complex was reacted with DME to form the sparingly soluble, dimeric 2-methoxyethoxy complex [Y(L)(mu OCH(2)CH(2)OMe-kappaO)](2) under C-O splitting. PMID- 15278115 TI - Chiral biarylamido/anisole complexes of yttrium in enantioselective aminoalkene hydaroamination/cyclisation. AB - A group of chiral, dibasic, biaryl-bridged amido proligands containing peripheral methoxyphenyl (anisole) ligation are developed for the synthesis of new amide complexes of yttrium and lanthanum. A potentially tetradentate bis(amidoanisole) system gives, on reaction with [Y[N(SiMe(2)H)(2)](3)(THF)] a crystallographically characterised bis complex [Y(H)] presumably as a result of low steric demand, since a more bulky version gives the target [Y[N(SiMe(2)H)(2)](THF)]. The molecular structure of the latter reveals a similar cis-alpha structure to our recently reported Schiff-base analogue. Variable-temperature NMR studies are consistent with low rigidity in the molecular structure. A potentially tridentate, amidoanisolyl/amido proligand gives complexes [M[N(SiMe(2)H)(2)](THF)(n)](M = Y, n= 1; M = La, n= 2). Chiral non-racemic versions of the above complexes were tested in the hydroamination/cyclisation of 2,2'-dimethylaminopentane to the corresponding pyrrolidine. Activities were relatively low compared to recently reported examples, and ee values were in the range 20-40% despite the well-expressed chirality of the catalysts. PMID- 15278116 TI - Arylaminopyridinato complexes of zirconium. AB - A range of 2-arylaminopyridines (HL) are synthesised readily from bromopyridines and amines using palladium-catalysed amination. Protonolysis reactions of these proligands with ZrX(4)(X = NMe(2), CH(2)Ph, CH(2)Bu(t)) yield zirconium complexes of the type [ML(n)X(4-n)], several of which have been characterised by X-ray crystallography. Control of metal/ligand stoichiometry and structure is pursued by investigation of the effects on substitution patterns of the pyridine and aryl rings. Some distinct patterns emerged; (i) the 6-methyl position on the pyridine appears to be particularly important with regards to control of stoichiometry, although there are co-ligand effects; (ii) structures of the metal alkyl derivatives [Zr(n)(CH(2)R)(4-n)] are dominated by aromatic pi-pi stacking, even when bulky arene substituents are employed at. This leads to the complexes adopting a C(2v)-symmetric core; (iii) the amides [Zr(2)(NMe(2))(2)] have structures for which aromatic pi-pi stacking is unfeasible, and correspondingly C(2)-symmetric or similar structures are adopted. All the structural data presented is consistent with a trans influence order at zirconium Me(2)N > RCH(2) > py. PMID- 15278117 TI - Crystal structures of imidazolium bis(trifluoromethanesulfonyl)imide 'ionic liquid' salts: the first organic salt with a cis-TFSI anion conformation. AB - Crystal structures of two examples of an important class of ionic liquids, 1,3 dimethylimidazolium and 1,2,3-triethylimidazolium bis(trifluoromethanesulfonyl)imide have been characterized by single crystal X ray diffraction. The anion in the 1,3-dimethylimidazolium example (mp 22 degrees C), adopts an unusual cis-geometry constrained by bifurcated cation-anion C-H. . .O hydrogen-bonds from the imidazolium cation to the anion resulting in the formation of fluorous layers within the solid-state structure. In contrast, in the 1,2,3-triethylimidazolium salt (mp 57 degrees C), the ions are discretely packed with only weak C-H. . .O contacts between the ions close to the van der Waals separation distances, and with the anion adopting the twisted conformation observed for all other examples from the limited set of organic bis(trifluoromethanesulfonyl)imide crystal structures. The structures are discussed in terms of the favorable physical properties that bis(trifluoromethanesulfonyl)imide anions impart in ionic liquids. PMID- 15278118 TI - Reactions of Li- and Yb-coordinated N,N'-bis(trimethylsilyl)-beta-diketiminates: one- and two-electron reductions, deprotonation, and C-N bond cleavage. AB - The synthesis and characterisation of novel Li and Yb complexes is reported, in which the monoanionic beta-diketiminato ligand has been (i) reduced (SET or 2 [times] SET), (ii) deprotonated, or (iii) C-N bond-cleaved. Reduction of the lithium beta-diketiminate Li(L(R,R'))[L(R,R')= N(SiMe(3))C(R)CHC(R')N(SiMe(3))] with Li metal gave the dilithium derivative [Li(tmen)(mu-L(R,R'))Li(OEt(2))](R = R'= Ph; or, R = Ph, R[prime or minute]= Bu(t)). When excess of Li was used the dimeric trilithium [small beta]-diketiminate [Li(3)(L(R,R[prime or minute]))(tmen)](2)(, R = R'= C(6)H(4)Bu(t)-4 = Ar) was obtained. Similar reduction of [Yb(L(R,R'))(2)Cl] gave [Yb[(mu-L(R,R'))Li(thf)](2)](, R = R[prime or minute]= Ph; or, R = R'= C(6)H(4)Ph-4 = Dph). Use of the Yb-naphthalene complex instead of Li in the reaction with [Yb(L(Ph,Ph))(2)] led to the polynuclear Yb clusters [Yb(3)(L(Ph,Ph))(3)(thf)], [Yb(3)(L(Ph,Ph))(2)(dme)(2)], or [Yb(5)(L(Ph,Ph))(L(1))(L(2))(L(3))(thf)(4)] [L(1)= N(SiMe(3))C(Ph)CHC(Ph)N(SiMe(2)CH(2)), L(2)= NC(Ph)CHC(Ph)H, L(3)= N(SiMe(2)CH(2))] depending on the reaction conditions and stoichiometry. The structures of the crystalline complexes 4, 6x21/2(hexane), 5(C(6)D(6)), and have been determined by X-ray crystallography (and have been published). PMID- 15278119 TI - [upper bond 1 start]PdCl[PPh(2)CH(2)CH(2)C[upper bond 1 end]HCH(2)CH(2)PPh(2)] as a precursor to homo- and hetero-metallic species directed by ESMS (electrospray ionisation mass spectrometry). AB - The cyclometallated Pd compound [upper bond 1 start]PdCl[PPh(2)CH(2)CH(2)C[upper bond 1 end]HCH(2)CH(2)PPh(2)], obtained from PdCl(2)(Ph(2)P(CH(2))(5)PPh(2)) in refluxing DMF (N,N-dimethylformamide), was characterized by NMR and X-ray single crystal diffraction analysis. The cyclometallation of MCl(2)(Ph(2)P(CH(2))(5)PPh(2))(M = Pd, Pt) to give and its Pt(ii) analogue was probed using electrospray ionisation mass spectrometry (ESMS). The reactivity of towards neutral ligands such as phosphines and pyridines as well as basic metal complexes such as pyridine-thiolate compounds of Au(i), Hg(ii) and Pt(ii) in solution was also investigated. The results showed that the chloride trans to the Pd-C bond is susceptible to ligand replacement. A number of entry metalloligands was examined in an attempt to establish a route to cyclometallated aggregates and clusters. PMID- 15278120 TI - Redox and complexation chemistry of the Cr(VI)/Cr(V)-D-galacturonic acid system. AB - The oxidation of d-galacturonic acid by Cr(VI) yields the aldaric acid and Cr(III) as final products when a 30-times or higher excess of the uronic acid over Cr(VI) is used. The redox reaction involves the formation of intermediate Cr(IV) and Cr(V) species, with Cr(VI) and the two intermediate species reacting with galacturonic acid at comparable rates. The rate of disappearance of Cr(VI), Cr(IV) and Cr(V) depends on pH and [substrate], and the slow reaction step of the Cr(VI) to Cr(III) conversion depends on the reaction conditions. The EPR spectra show that five-coordinate oxo-Cr(V) bischelates are formed at pH < or = 5 with the uronic acid bound to Cr(V) through the carboxylate and the alpha-OH group of the furanose form or the ring oxygen of the pyranose form. Six-coordinated oxo Cr(V) monochelates are observed as minor species in addition to the major five coordinated oxo-Cr(V) bischelates only for galacturonic acid : Cr(VI) < or =10 : 1, in 0.25-0.50 M HClO(4). At pH 7.5 the EPR spectra show the formation of a Cr(V) complex where the vic-diol groups of Galur participate in the bonding to Cr(V). At pH 3-5 the Galur-Cr(V) species grow and decay over short periods in a similar way to that observed for [Cr(O)(alpha-hydroxy acid)(2)](-). The lack of chelation at any vic-diolate group of Galur when pH < or = 5 differentiates its ability to stabilise Cr(V) from that of neutral saccharides that form very stable oxo-Cr(V)(diolato)(2) species at pH > 1. PMID- 15278121 TI - Extended metal atom chains (EMACs) of five chromium or cobalt atoms: symmetrical or unsymmetrical? AB - Structural studies of pentachromium and pentacobalt extended metal atom chain (EMAC) systems are presented in which the metal chains are helically wrapped by either the pentadentate dianion tripyridyldiamide (tpda) or its diethyl substituted analogue bis(4-ethyl-2-pyridylamido)pyridine (etpda). The compound Cr(5)(tpda)(4)(NCS)(2), has alternating long and short Cr-Cr distances, contrary to recent reports that describe it as symmetrical with essentially equally spaced chromium atoms. The linear Cr(5)(10+) chain is composed of two Cr(2)(4+) quadruply bonded units and an isolated high spin Cr(II) unit. The new compounds Cr(5)(etpda)(4)Cl(2), [Cr(5)(etpda)(4)FCl]PF(6), and Co(5)(etpda)(4)(NCS)(2)() employ the etpda ligand as "insulation" around a central pentametal-atom "wire." Compound, like all other oxidized pentachromium compounds, has very disparate alternating short-long-short-long Cr-Cr distances, 2.032(3) A, 2.560(6) A, 1.873(5) A, and 2.509(4) A. Compound shows a nearly uniform spacing of the Co atoms, although the outer Co-Co distances (2.279[4] A) are slightly longer than the inner ones (2.239[4] A). PMID- 15278122 TI - Dinegative tetrahedral oxoanion complexation; structural and solution phase observations. AB - A structural and solution phase study of complexation of dinegative oxoanionic guests by protonated azacryptand hosts is presented. The interactions involve a complex balance of competing factors: anion free energy of hydration, host basicity and solvation, and complementarity of steric matching between host and guest. The formation constants of dinegatively charged anion cryptates are much larger than the analogous values for the mononegative analogues perchlorate or perrhenate. PMID- 15278123 TI - Coordinative flexibility of 1,2-bis[1,4,7-triazacyclonon-1-yl]propan-2-ol in mononuclear and binuclear Ni(II) complexes. AB - Reaction of 1,2-bis[1,4,7-triazacyclonon-1-yl]propan-2-ol hexabromide (T(2)PrOH.6HBr) with Ni(ClO(4))(2)[middle dot]6H(2)O and adjustment of the pH to 7 resulted in the crystallization of pink and blue products from the one reaction mixture. The analytical data and X-ray structure determinations establish compositions corresponding to [Ni(T(2)PrOH)]Br(ClO(4))xH(2)O (pink crystals) and [Ni(2)(T(2)PrO)(OH(2))(3)Br]Br(ClO(4))x2H(2)O (blue crystals). A repeat synthesis of the latter yielded the diperchlorate monohydrate [Ni(2)(T(2)PrO)(OH(2))(3)Br](ClO(4))(2)xH(2)O. In the mononuclear complex, the 2 propanol group connecting the two 1,4,7-trizacyclononane (tacn) rings is protonated, the six nitrogen donors from the T(2)PrOH ligand coordinating to a single Ni(II) centre in a distorted octahedral geometry. In the binuclear complexes and, three coordination sites on each distorted octahedral Ni(II) centre are occupied fac by three nitrogen donors from the one tacn ring, the two metal centres being linked by an endogenous alkoxo bridge. A notable common feature of the two identical cations is that for one Ni(II) centre the remaining two sites are occupied by two water ligands, while in the other a bromo ligand replaces one ligated water. Similar binuclear systems have been recently defined [Zn(2)(T(2)PrO)X(H(2)O)(2)](ClO(4))(2)(X = Cl, Br), two complexes that exhibit coordination asymmetry with one pseudo-octahedral and one pseudo-square pyramidal Zn(ii) centre. The weak antiferromagnetic coupling in and is discussed and compared to di-phenoxo-bridged Ni(II) examples. PMID- 15278124 TI - Biphenyl derived Schiff-base vanadium(V) complexes with pendant OH-groups- structure, characterization and hydrogen peroxide mediated sulfide oxygenation. AB - A series of mononuclear oxovanadium(v) complexes of tridentate Schiff bases HL(1) HL(4) and H(2)L(5)-H(2)L(8) derived from 6-phenylsalicylaldehyde and 6-(2 hydroxyphenyl)salicylaldehyde and four different amines was synthesized. The systematically selected ligands contain aliphatic or aromatic nitrogen, or alkoxy and phenoxy-oxygen as third donor atom. The complexes were characterised by spectroscopic methods in solution and the solid state. Single-crystal X-ray analyses were performed with VO(2)L(1)(), VO(2)L(3)x1/2EtOH (), VO(2)L(4)(), VO(OiPr)L(7)xiPrOH, VO(OiPr)L(8) and H(2)L(8). For all compounds the vanadium(v) cores contain distorted tetragonal pyramidal geometry around the dioxo- and oxovanadium site at which the N(2)O- and NO(2)-donor ligands bind equatorially. Complexes and display intramolecular hydrogen bonding of the pendant hydroxyphenyl group to a coordinated oxygen trans to a nitrogen atom and therefore serve as suitable models for the native site of vanadium dependent haloperoxidases. Variable-temperature (1)H NMR spectra revealed significant hydrogen bond interaction in acetonitrile solution. In situ prepared catalysts are active for hydrogen peroxide mediated oxygenation of ethyl phenyl sulfide and showed complete conversion of the substrate to ethyl phenyl sulfoxide, together with small amounts of the corresponding sulfone, as detected by GC/MS after 10 min. The complex of H(2)L(7) turned out to be most efficient while HL(1) and HL(2) were completely inactive. Catalysis is supported by the pendant OH group in the complex of HL(3), the catalyst is twice as active as the complex of HL(4). PMID- 15278125 TI - Reaction behaviour of dinuclear copper(I) complexes with m-xylyl-based ligands towards dioxygen. AB - Intramolecular ligand hydroxylation was observed during the reactions of dioxygen with the dicopper(I) complexes of the ligands L(1)(L(1)=alpha,alpha'-bis[(2 pyridylethyl)amino]-m-xylene) and L(3)(L(3)=alpha, alpha'-bis[N-(2-pyridylethyl) N-(2-pyridylmethyl)amino]-m-xylene). The dinuclear copper(I) complex [Cu(2)L(3)](ClO(4))(2) and the dicopper(II) complex [Cu(2)(L(1) O)(OH)(ClO(4))]ClO(4) were characterized by single-crystal X-ray structure analysis. Furthermore, phenolate-bridged complexes were synthesized with the ligand L(2)-OH (structurally characterized [Cu(2)(L(2)-O)Cl(3)] with L(2)=alpha, alpha'-bis[N-methyl-N-(2-pyridylethyl)amino]-m-xylene; synthesized from the reaction between [Cu(2)(L(2)-O)(OH)](ClO(4))(2) and Cl(-)) and Me-L(3)-OH: [Cu(2)(Me-L(3)-O)(mu-X)](ClO(4))(2)xnH(2)O (Me-L(3)-OH = 2,6-bis[N-(2 pyridylethyl)-N-(2-pyridylmethyl)amino]-4-methylphenol and X = C(3)H(3)N(2)( )(prz), MeCO(2)(-) and N(3)(-)). The magnetochemical characteristics of compounds were determined by temperature-dependent magnetic studies, revealing their antiferromagnetic behaviour [-2J(in cm(-1)) values: -92, -86 and -88; -374]. PMID- 15278126 TI - A study on the mimics of Cu-Zn superoxide dismutase with high activity and stability: two copper(II) complexes of 1,4,7-triazacyclononane with benzimidazole groups. AB - Two copper(II) complexes [CuL(1)Cl]ClO(4) and [CuL(2)MeCN](ClO(4))(2)xH(2)O were synthesized (L(1)= 1-(benzimidazole-2-ylmethyl)-1,4,7-triazacyclononane, L(2)= 1,4-bis(benzimidazole-2-ylmethyl)-1,4,7-triazacyclonone). The benzimidazole groups were N-substituents of tacn, and the complexes are more stable than their parents. They are able to catalyse the dismutation of superoxide anion in aqueous solutions at physiological pH and in bovine serum albumin solution (0.5 mg ml( 1)). X-ray structure analysis and EPR and electronic spectra show that the structure of complex is more similar to the Cu(II) centre of Cu(2)Zn(2)SOD than that. Comparing with other Cu(II) complexes, the complex possesses both high SOD activity and highly thermodynamic stability. PMID- 15278127 TI - Reactions of [Ru(H(2)O)(6)](2+) with water-soluble tertiary phosphines. AB - In aqueous solutions under mild conditions, [Ru(H(2)O)(6)](2+) was reacted with various water-soluble tertiary phosphines. As determined by multinuclear NMR spectroscopy, reactions with the sulfonated arylphosphines L =mtppms, ptppms and mtppts yielded only the mono- and bisphosphine complexes, [Ru(H(2)O)(5)L](2+), cis-[Ru(H(2)O)(4)L(2)](2+), and trans-[Ru(H(2)O)(4)L(2)](2+) even in a high ligand excess. With the small aliphatic phosphine L = 1,3,5-triaza-7 phosphatricyclo-[3.3.1.1(3,7)]decane (pta) at [L]:[Ru]= 12:1, the tris- and tetrakisphosphino species, [Ru(H(2)O)(3)(pta)(3)](2+), [Ru(H(2)O)(2)(pta)(4)](2+), [Ru(H(2)O)(OH)(pta)(4)](+), and [Ru(OH)(2)(pta)(4)] were also detected, albeit in minor quantities. These results have significance for the in situ preparation of Ru(II)-tertiary phosphine catalysts. The structures of the complexes trans-[Ru(H(2)O)(4)(ptaMe)(2)](tos)(4)x2H(2)O, trans [Ru(H(2)O)(4)(ptaH)(2)](tos)(4)[middle dot]2H(2)O, and trans-mer [RuI(2)(H(2)O)(ptaMe)(3)]I(3)x2H(2)O, containing protonated or methylated pta ligands (ptaH and ptaMe, respectively) were determined by single crystal X-ray diffraction. PMID- 15278128 TI - Electronic and photophysical properties of a novel phenol bound dinuclear ruthenium complex: evidence for a luminescent mixed valence state. AB - A novel dinuclear ruthenium(II) complex bridged by dianionic bridge 3-(2-phenol) 5-(pyridin-2-yl)-1,2,4-triazole in which the ruthenium metal atoms are bound through N,N coordination to the pyridine and triazole and O,N coordination to the triazole and phenolate is described. The electrochemical, spectroscopic and photophysical behaviour of the dimer is compared with its associated N,N- and O,N coordinated mononuclear complexes. The mixed valence complex was prepared electrochemically and a weak inter-valence charge transfer transition is observed which from Hush theory provides an electronic coupling matrix element of 666 cm( 1), suggesting the complex is weakly coupled and valence trapped. In its native state the dinuclear compound is essentially non-emissive but upon the oxidation of the O,N moiety luminescence from the complex is reversibly switched on at 0.3 V and reversibly switched off by application of 1.3 or 0 V. To our knowledge this is the first report of a luminescent mixed valence ruthenium complex. PMID- 15278129 TI - Appraisal of the redox behaviour of the antimetastatic ruthenium(III) complex [ImH][RuCl(4)(DMSO)(Im)], NAMI-A. AB - The imidazolium trans-tetrachloro(dimethylsulfoxide)imidazoleruthenate(III) complex [ImH][Ru(III)Cl(4)(DMSO)(Im)], NAMI-A, has shown an interesting antimetastatic activity. Since Ru(III) complexes are coordinatively more inert than the corresponding Ru(II) derivatives, an "activation by reduction" mechanism has been proposed to explain the biological activity of NAMI-A, thus acting as a pro-drug. We report here an electrochemical study on NAMI-A in aqueous solutions which emphasizes the structural and chemical consequences accompanying the easy Ru(III)/Ru(II) electron transfer (e.g., axial imidazole/water exchange in acidic solution in the short timescale of cyclic voltammetry followed by equatorial chloride/water exchange in the longer timescale of macroelectrolysis). PMID- 15278130 TI - Electrochemical oxidation of methanol using dppm-bridged Ru/Pd, Ru/Pt and Ru/Au catalysts. AB - The electrochemical oxidation of methanol was carried out using a series of dppm bridged Ru/Pd, Ru/Pt and Ru/Au heterobimetallic complexes as catalysts. The major oxidation products were formaldehyde dimethyl acetal (dimethoxymethane, DMM) and methyl formate (MF). The Ru/Pd and Ru/Pt bimetallic catalysts generally afforded lower product ratios of DMM/MF and higher current efficiencies than the Ru/Au catalysts. The Ru/Au bimetallics exhibited product ratios and current efficiencies similar to those obtained from the Ru mononuclear compound CpRu(PPh(3))(2)Cl. Increasing the methanol concentration afforded higher current efficiencies, while the addition of water to the samples shifted the product distribution toward the more highly oxidized product, MF. PMID- 15278131 TI - Electrochemistry of a labile average-valence dicopper system. AB - Electrochemical studies on an average-valent dicopper cryptate demonstrate the existence of a solvent-assisted disproportionation in acetonitrile, which is absent in water. The existence of this process was confirmed via absorption spectroscopy, which allowed the evaluation of thermodynamic parameters. The disproportionation equilibrium is enthalpically controlled, and the results suggest an upper limit for the bond energy of the one-electron Cu-Cu bond of 200 kJ mol(-1). PMID- 15278132 TI - Mono- and dinuclear complexes of sulfones with the tetrachlorides of group 4. AB - The reactions of dialkyl sulfones [R(2)SO(2): R = Me, Et, Ph, R(2)=-(CH(2))(4)-] with the metal tetrachlorides of Group 4 [MCl(4): M = Ti, Zr, Hf] give different products mainly depending on the sulfone/M molar ratio. Compounds of formula [M(2)Cl(8)(R(2)SO(2))(2)][M = Ti, R(2)=-(CH(2))(4)-; M = Zr, R = Et, R = Ph] and [MCl(4)(R(2)SO(2))(2)](sulfone/M = 2)[M = Ti, R = Me; M = Zr, R = Me, R = Ph, R(2)=-(CH(2))(4)-; M = Hf, R = Me, R(2)=-(CH(2))(4)-] have been obtained. By X ray diffraction methods the dinuclear titanium and zirconium adducts, [Ti(2)Cl(8)(mu-sulfolane-O,O')(2)] and [Zr(2)Cl(8)(mu-Ph(2)SO(2)-O,O')(2)] have been established to contain bridging sulfone and hexacoordinated metal centres, while the mononuclear zirconium complex [ZrCl(4)(Me(2)SO(2))(2)] has cis monodentate sulfones in a slightly distorted octahedral geometry. The reaction between TiCl(4) and sulfolane (tetrahydrothiophene 1,1-dioxide) in SOCl(2) affords the 1:1 adduct independent of the sulfone/Ti molar ratio. Ligand-exchange and inter-conversion between mononuclear and dinuclear species have been observed by NMR, while the spectral features of the SO(2) moiety have been assigned by IR- and Raman spectroscopies. PMID- 15278133 TI - Organotrichlorogermane synthesis by the reaction of elemental germanium, tetrachlorogermane and organic chloride via dichlorogermylene intermediate. AB - Organotrichlorogermanes were synthesized by the reaction of elemental germanium, tetrachlorogermane and organic chlorides, methyl, propyl, isopropyl and allyl chlorides. Dichlorogermylene formed by the reaction of elemental germanium with tetrachlorogermane was the reaction intermediate, which was inserted into the carbon-chlorine bond of the organic chloride to give organotrichlorogermane. When isopropyl or allyl chloride was used as an organic chloride, organotrichlorogermane was formed also in the absence of tetrachlorogermane. These chlorides were converted to hydrogen chloride, which subsequently reacted with elemental germanium to give the dichlorogermylene intermediate. The reaction of elemental germanium, tetrachlorogermane and organic chlorides provides a simple and easy method for synthesizing organotrichlorogermanes, and all the raw materials are easily available. PMID- 15278134 TI - Synthesis, characterisation and optical spectroscopy of platinum(II) di-ynes and poly-ynes incorporating condensed aromatic spacers in the backbone. AB - A series of protected and terminal dialkynes with extended pi-conjugation through a condensed aromatic linker unit in the backbone, 1,4 bis(trimethylsilylethynyl)naphthalene, 1,4-bis(ethynyl)naphthalene, 9,10 bis(trimethylsilylethynyl)anthracene, 9,10-bis(ethynyl)anthracene, have been synthesized and characterized spectroscopically. The solid-state structures of and have been confirmed by single crystal X-ray diffraction studies. Reaction of two equivalents of the complex trans-[Ph(Et(3)P)(2)PtCl] with an equivalent of the terminal dialkynes 1,4-bis(ethynyl)benzene and, in (i)Pr(2)NH-CH(2)Cl(2), in the presence of CuI, at room temperature, afforded the platinum(II) di-ynes trans [Ph(Et(3)P)(2)Pt-C[triple bond, length as m-dash]C-R-C[triple bond, length as m dash]C-Pt(PEt(3))(2)Ph](R = benzene-1,4-diyl; naphthalene-1,4-diyl and anthracene 9,10-diyl ) while reactions between equimolar quantities of trans [((n)Bu(3)P)(2)PtCl(2)] and under similar conditions readily afforded the platinum(II) poly-ynes trans-[-((n)Bu(3)P)(2)Pt-C[triple bond]C-R-C[triple bond]C ](n)(R = naphthalene-1,4-diyl and anthracene-9,10-diyl ). The Pt(II) diynes and poly-ynes have been characterized by analytical and spectroscopic methods, and the single crystal X-ray structures of and have been determined. These structures confirm the trans-square planar geometry at the platinum centres and the linear nature of the molecules. The di-ynes and poly-ynes are soluble in organic solvents and readily cast into thin films. Optical spectroscopic measurements reveal that the electron-rich naphthalene and anthracene spacers create strong donor-acceptor interactions between the Pt(II) centres and conjugated ligands along the rigid backbone of the organometallic polymers. Thermogravimetry shows that the di-ynes possess a somewhat higher thermal stability than the corresponding poly-ynes. Both the Pt(II) di-ynes and the poly-ynes exhibit increasing thermal stability along the series of spacers from phenylene through naphthalene to anthracene. PMID- 15278135 TI - Dendrimers as guests in molecular recognition phenomena. AB - Dendrimers are highly branched macromolecules which may engage in host-guest interactions, acting as either hosts or guests; this review is specifically concerned with the binding behavior of dendrimers containing single or multiple guest residues interacting with individual, freely diffusing hosts. PMID- 15278136 TI - Entrapment of enzymes and nanoparticles using biomimetically synthesized silica. AB - Entrapment of enzymes and nanoparticles using biosilicification reactions. PMID- 15278137 TI - Single-walled carbon nanotubes filled with M OH (M = K, Cs) and then washed and refilled with clusters and molecules. AB - Heating single-walled carbon nanotubes (SWNTs) with molten hydroxides MOH (M = K, Cs) gave MOH@SWNT in good yield; high resolution transmission electron microscopy (HRTEM) indicated that CsOH in CsOH@SWNT often adopts twisted 1D crystal structures inside SWNTs; treating MOH@SWNT with water at room temperature removes the soluble hydroxide filling and the resulting SWNTs may then be filled using aqueous solutions of uranyl acetate or uranyl nitrate at rt giving SWNTs filled with UO(2) clusters and uranyl acetate molecules. PMID- 15278138 TI - First observation of capping/uncapping by a ligand of a Zn porphyrin adsorbed on Ag(100). AB - A significant first step towards creation of catalytically active porphyrin functionalised metal surfaces has been achieved. PMID- 15278139 TI - Constellational diastereomers in encapsulation complexes. AB - Three chiral guests in a cylindrical capsule led to six diastereomeric complexes. PMID- 15278140 TI - [Re6Te8(CN)6][(Ir(CO)(PPh3)2)6](OTf)2 : a new Re6 cluster-supported iridium(I) compound. AB - A new hexanuclear rhenium cluster encapsulated by six iridium complexes, [Re6Te8(CN)6][(Ir(CO)(PPh3)2)6](OTf)2 (3), which is effective in catalyzing the hydrogenation of p-CH3C6H4C[triple bond]CH to p-CH3C6H4CH=CH2 has been prepared. PMID- 15278141 TI - Two dimensional PNA/DNA arrays: estimating the helicity of unusual nucleic acid polymers. AB - We extend the generality of nucleic acid-based structural nanotechnology by incorporating non-natural nucleic acids into a DNA double crossover (DX) molecule; visualizing two-dimensional arrays of these DX molecules by Atomic Force Microscopy (AFM) enables us to measure the helical repeat of any heteroduplex sequence capable of forming the outer arms of a DX. PMID- 15278142 TI - New fascaplysin-based CDK4-specific inhibitors: design, synthesis and biological activity. AB - The first biologically active non-planar analogues of the toxic anti-cancer agent, fascaplysin, have been produced; we present the design, synthesis and biological activity of three tryptamine derivatives. PMID- 15278143 TI - Dimerization of a guanidiniocarbonyl pyrrole cation in DMSO that can be controlled by the counteranion. AB - In the presence of chloride anions cation 1 dimerizes in DMSO with a surprisingly high association constant of > 10(3) M(-1) whereas the addition of picrate disrupts these dimers by formation of even more stable discrete pi-stacked ion pairs. PMID- 15278144 TI - Cruciform pi-systems: effect of aggregation on emission. AB - The solid state properties of cruciform pentamers are examined in thin film preparation, in the single crystalline state and in nanoparticle formulations; emission behavior was found to vary substantially with the solid state morphology. This type of behaviour is an excellent way to manipulate the emissive properties of conjugated [small pi]-systems. PMID- 15278145 TI - Vibronic coupling in the ground and excited states of the naphthalene cation. AB - The hole-vibrational coupling in naphthalene is studied using high-resolution gas phase photoelectron spectroscopy and density functional theory calculations (DFT), and a remarkable increase of the coupling with low-frequency vibrations is observed in the excited states. PMID- 15278146 TI - Base-discriminating fluorescent (BDF) nucleoside: distinction of thymine by fluorescence quenching. AB - A novel fluorescence BDF probe containing pyrene-labeled 7-deaza-2[prime or minute]-deoxyadenosine has been developed for the detection of thymine base on a target DNA. PMID- 15278147 TI - A convergent, versatile route to two synthetic conjugate anti-toxin malaria vaccines. AB - The synthesis of two glycosylphosphatidyl inositol (GPI) glycans that constitute the malaria toxin and promising anti-toxin vaccine constructs using a scalable route is described. PMID- 15278148 TI - A Suzuki-Miyaura coupling mediated deprotection as key to the synthesis of a fully lipidated malarial GPI disaccharide. AB - Ligandless palladium-catalyzed Suzuki-Miyaura coupling converted an inert p bromobenzyl ether to a DDQ-labile p-(3,4-dimethoxyphenyl) benzyl ether in the presence of azide functionality and this strategy serves as a key step for the convergent synthesis of a fully lipidated malaria GPI disaccharide. PMID- 15278149 TI - Mesoporous silica-supported zirconocene catalysts for highly isotactic polypropylene. AB - A new bis(indenyl)zirconocene bearing a pendant Si-Cl anchor has been grafted onto MCM-41, SBA-15, MCM-48 and a disordered mesoporous silica for the polymerisation of ethylene and propylene, producing polymers with very high molecular weights, low polydispersities and, in the case of polypropylene, higher levels of isotacticity than obtainable with analogous homogeneous systems. PMID- 15278150 TI - Unprecedented eight-palladium(I) crown-cycle with metal-metal unsupported bonds. AB - Tri(N-pyrrolyl)phosphine reacted with the sigma/pi complex [Pd(mu-Cl)(COD-MeO)]2 to give the octa-cycle [Pd(mu-Cl)[P(pyrl)3]]8 containing four Pd(I)-Pd(I) unbridged bonds. PMID- 15278151 TI - H-D exchange reaction on benzene ring of polystyrene in hydrothermal deuterium oxide with platinum(IV) oxide catalyst. AB - Benzene rings of polystyrene samples are labelled with deuterium oxide and catalytic amount of platinum(iv) oxide under hydrothermal conditions. PMID- 15278152 TI - Mixed metal bis(mu-oxo) complexes with [CuM(mu-O)2]n+(M = Ni(III) or Pd(II)) cores. AB - Two highly reactive heterodinuclear bis(mu-oxo) complexes were prepared by combining mononuclear peroxo species with reduced metal precursors at -80 degrees C and were identified by UV-vis, EPR/NMR, and resonance Raman spectroscopy, with corroboration in the case of the CuPd system from density functional calculations. PMID- 15278153 TI - Novel synthesis of FAU-type zeolite membrane with high performance. AB - FAU-type zeolite membranes were synthesized by the vapor phase transformation (VPT) methods with or without prior seeding on the substrate, and it was revealed that the CO(2)/N(2) selectivity of the seeded membrane is greater than that of the unseeded membrane. PMID- 15278154 TI - Highly selective formation of linear esters from terminal and internal alkenes catalysed by palladium complexes of bis-(di-tert-butylphosphinomethyl)benzene. AB - The methoxycarbonylation of terminal or internal alkenes catalysed by palladium complexes of bis-(di-tert-butylphosphinomethyl)benzene under mild conditions leads to linear esters in 99% selectivity via a hydride mechanism. PMID- 15278155 TI - Facile synthesis of membrane-embedded peptides utilizing lipid bilayer-assisted chemical ligation. AB - Lipid bilayer-assisted chemical ligation between thiolester and N-terminal cysteine peptides has been developed with successful application to the synthesis of membrane protein segments possessing both two transmembrane and one extracellular regions. PMID- 15278156 TI - Lipase catalysed Michael addition of secondary amines to acrylonitrile. AB - A new enzymatic process is described. Different preparations of lipase B from Candida antarctica are able to catalyse Michael-type addition of secondary amines to acrylonitrile. This new reaction widens the applicability of these biocatalysts in organic synthesis. PMID- 15278157 TI - Gold(I) or gold(III) as active species in AuCl(3)-catalyzed cyclization/cycloaddition reactions? A DFT study. AB - In a B3LYP mechanistic study, AuCl and AuCl(3) catalysts feature similar overall barriers for a reaction sequence of 2-ethynyl benzaldehyde and ethyne to 1 naphthyl carbaldehyde, comprising cyclization, [3+2] cycloaddition, and two rearrangements. PMID- 15278158 TI - Methane activation by silica-supported Zr(IV) hydrides: the dihydride [(triple bond)SiO)2ZrH2] is much faster than the monohydride [(triple bond)SiO)3ZrH]. AB - The silica-supported Zr(iv) dihydride [(triple bond)SiO)2ZrH2] reacts quickly and completely with methane to yield [(triple bond)SiO)2ZrMe2] through the intermediate [(triple bond)SiO)2ZrHMe], while its monohydride analogue [(triple bond)SiO)3ZrH] yields the monomethylated product [(triple bond)SiO)3ZrMe] slowly and incompletely. PMID- 15278159 TI - Fe-Ga multiple bonding? Synthesis, spectroscopic and structural characterization of a transition metal complex containing a cationic two-coordinate gallium centre. AB - This communication reports the synthesis and characterization of the cationic iron complex [((eta5-C5Me5)Fe(CO)2))2Ga]+[BAr(f)4]- [Ar(f) = C6H3(CF3)2-3,5] containing a symmetrically bridging two-coordinate gallium atom and a delocalised Fe-Ga-Fe pi system incorporating partial Fe-Ga multiple bond character. PMID- 15278160 TI - Microwave-assisted sidewall functionalization of single-wall carbon nanotubes by Diels-Alder cycloaddition. AB - The first Diels-Alder cycloaddition of o-quinodimethane to SWNT has been performed under microwave irradiation. PMID- 15278161 TI - Design of a doubly-hydrophilic block copolypeptide that directs the formation of calcium carbonate microspheres. AB - The crystallization of calcium carbonate into microspheres has been accomplished using the rationally-designed, doubly-hydrophilic block copolypeptide poly(Nepsilon-2[2-(2-methoxyethoxy)ethoxy]acetyl-L-lysine)(100)-b-poly(L aspartate sodium salt)30 as a structure-directing agent. PMID- 15278162 TI - Novel organocycloborates via Grignard reagents. AB - An unprecedented cyclisation reaction of alpha,omega-borylbromoalkanes is seen upon treatment with magnesium turnings, forming organocycloborates in high yield. The novel boratacyclopentanes and -hexanes have been characterised by X-ray crystallography. PMID- 15278163 TI - Mixed ligand system of cysteine and thioglycolic acid assisting in the synthesis of highly luminescent water-soluble CdTe nanorods. AB - Highly luminescent water-soluble CdTe nanorods were prepared with the assistance of the mixed ligand system of cysteine and thioglycolic acid; the aspect ratio and photoluminescence of the CdTe nanorods could be controlled by the refluxing time. PMID- 15278164 TI - A general one-pot process leading to highly functionalised ordered mesoporous silica films. AB - Various organic moieties are homogeneously introduced in high quantities into mesostructured porous silica films through a general co-condensation process, which influences the self-assembly mechanism, depending on the physico-chemical properties of each function. PMID- 15278166 TI - Catalytic activity of dodecacarbonyltetracobalt in aqueous media: a "greening" of the Pauson-Khand reaction. AB - The unprecedented reactivity of Co(4)(CO)(12) with enynes under aqueous conditions, representing the development of a mild and simple aqueous-phase cobalt-catalyzed PK reaction protocol, is described herein. PMID- 15278165 TI - Binding of CO to structural models of the bimetallic subunit at the A-cluster of acetyl coenzyme A synthase/CO dehydrogenase. AB - Trinuclear Ni-Cu-Ni and Ni-Ni-Ni complexes derived from an Ni(ii)-dicarboxamido dithiolato metallosynthon exhibit redox behavior and CO binding properties similar to those of the A-cluster in acetyl coenzyme A synthase/CO dehydrogenase (ACS/CODH). PMID- 15278167 TI - Protonation of a lanthanum phosphide-alkyl occurs at the P-La not the C-La bond: isolation of a cationic lanthanum alkyl complex. AB - Protonation of the heteroleptic, cyclometalated lanthanum phosphide complex [((Me3Si)2CH)(C6H4-2-CH2NMe2)P]La(THF)[P(C6H4-2-CH2NMe2)(CH(SiMe3)(SiMe2CH2))] with [Et3NH][BPh4] yields the cationic alkyllanthanum complex [(THF)4La[P(C6H4-2 CH2NMe2)(CH(SiMe3)(SiMe2CH2))]][BPh4]. PMID- 15278168 TI - Magnetic behavior of tetrakis[4-(N-tert-butyl-N-oxylamino)pyridine]bis(isocyanato N)cobalt(II) in frozen solution. AB - A complex of cobalt(II)(OCN)(2) coordinated with four pyridines having a stable tert-butyl aminoxyl exhibited in frozen MTHF a slow magnetic relaxation for the reorientation of magnetization with activation barrier, Delta/k(B) = 50 K, and a hysteresis loop having a fast relaxation at 0 Oe below 2.5 K. PMID- 15278169 TI - Hydrogen storage on fullerenes: hydrogenation of C59N. using C60H36 as the source of hydrogen. AB - C(60)H(36) has been used as the source of hydrogen for the in situ hydrogenation of (C(59)N)(2), leading to C(59)NH(5) as the main reaction product identified by negative-ion mass spectrometry and providing evidence of the usage of C(60) as a storage device for hydrogen. PMID- 15278170 TI - Superelectrophilic activation of polyfunctional organic compounds using zeolites and other solid acids. AB - Zeolites and other available solid acids have been successfully applied to initiate reactions, which were earlier recognised to involve superelectrophilic intermediates and thus required excess of superacids to be carried out. PMID- 15278171 TI - Excess electron transfer in G-quadruplex. AB - The excess electron transfer in a G-quadruplex is successfully probed by using the reaction of hydrated electrons with quadruplex complex of pentamers and the 8 bromoguanine moieties as the detection system. PMID- 15278172 TI - Tris(thioacetals) from benzene hexathiol: towards covalent self-assembly. AB - Mixtures of isomers are available from the reaction of benzene hexathiol with three equivalents of p-tolualdehyde and kinetic traps avoided under the reported catalytic conditions, establishing tris(thioacetals) as potential building blocks for covalently self-assembled complex structures. PMID- 15278173 TI - Template synthesis of functionalized polystyrene in ordered silicate channels. AB - A mesostructured nanocomposite was fabricated by using a novel electroactive, polymerizable surfactant as a template in a sol-gel process, and the first example of well-resolved polystyrene with redoxactive functional group synthesized in silicate matrices was provided. PMID- 15278174 TI - Determination of cysteine concentration by fluorescence increase: reaction of cysteine with a fluorogenic aldehyde. AB - A fluorogenic method for the determination of cysteine concentration has been developed. PMID- 15278175 TI - Preparation and structural characterisation of methoxybis(trimethylsilyl)silyl potassium and its condensation product. AB - Depending on the conditions the reaction of tris(trimethylsilyl)methoxysilane (1) with potassium tert-butoxide either in benzene and in the presence of 18-crown-6 or in THF gives either the crown ether adduct of potassium methoxybis(trimethylsilyl)silane (2), or 2 methoxytetrakis(trimethylsilyl)disilanyl potassium (3). PMID- 15278176 TI - Glow discharge growth of SnO2 nano-needles from SnH4. AB - Single crystalline SnO(2) nano-needles with length up to 6-7 microm and diameter less than 300 nm are synthesized on an Au-coating porous silicon substrate from SnH(4) source via a glow discharge process. PMID- 15278177 TI - Supported ionic liquids: ordered mesoporous silicas containing covalently linked ionic species. AB - Ordered mesoporous silicas with hexagonal or lamellar architectures incorporating covalently bound ionic species were synthesized via a template directed hydrolysis-polycondensation of tetraethoxysilane (TEOS) with triethoxysilylated imidazole [(EtO)(3)Si(CH(2))(3)-Im] or alkylimidazolium halides [(EtO)(3)Si(CH(2))(3)-Im(+)-R Hal(-)]. PMID- 15278178 TI - Novel coordinating motifs for lanthanide(III) ions based on 5-(2 pyridyl)tetrazole and 5-(2-pyridyl-1-oxide)tetrazole. Potential new contrast agents. AB - Water-soluble and neutral Ln(III) and Zn (II) complexes of pyridine- and (pyridine-1-oxide)tetrazole have been synthesized and the Gd derivatives have great potential as high-relaxivity low-osmolarity MRI contrast agents. PMID- 15278179 TI - Intramolecular C-H insertions adjacent to sulfur for the diastereoselective synthesis of thienofuranones. AB - A new approach to the diastereoselective synthesis of thienofuranones is described in which an intramolecular 1,5-carbenoid C-H insertion adjacent to sulfur features as a key step. PMID- 15278180 TI - Tuning iridium(III) phenylpyridine complexes in the "almost blue" region. AB - We report on the synthesis and photophysical properties of blue emitting iridium(iii) complexes. The use of a negatively charged ligand, such as a triazolyl pyridine, allows a facile preparation, maintaining the high energy emission (blue region) of heteroleptic complexes. We discuss the role played by electron withdrawing substituents of a different nature and also how the substitution position of the same group influences the spectroscopical behaviour. PMID- 15278181 TI - Identification of peptides that promote the rapid precipitation of germania nanoparticle networks via use of a peptide display library. AB - Peptides that promote the rapid, room-temperature precipitation of amorphous germania nanoparticle networks from solution have been identified via use of a combinatorial peptide display library. PMID- 15278182 TI - Current role of local ablative treatments for hepatocellular carcinoma. AB - Due to modern diagnostic imaging and the sensitive alpha-fetoprotein test, small hepatocellular carcinoma can now be detected at an early stage. Studies have shown that surgical resection of the tumors is a valuable treatment. Local treatment under ultrasound guidance was initially considered as an alternative when patients' liver reserves were not good enough for surgical resection; however, this technique has been improved and the results indicate that its survival rate can compete with that of surgical resection. In follow-up studies of patients with small hepatocellular carcinoma, a 5-year survival of 60% has been achieved after percutaneous ethanol injection therapy. Percutaneous microwave coagulation therapy and percutaneous radiofrequency ablation therapy have been shown to have some advantages over percutaneous ethanol injection therapy, although the follow-up durations of these studies were not long enough. Percutaneous ethanol injection therapy, percutaneous microwave coagulation therapy and percutaneous radiofrequency ablation therapy have become the 3 most widely used techniques for the treatment of hepatocellular carcinomas that are less than 5 cm in diameter and have a tumor number less than 3. In general, a tumor size of 3 to 5 cm is a good candidate for radiofrequency ablation and a tumor size of 2 to 3 cm is suitable for radiofrequency ablation or microwave coagulation. If the tumor size is around 2 cm or less, microwave coagulation or ethanol injection is often chosen due to the relatively low cost and similar efficacy. Ethanol injection also has the advantage of needing only a fine needle for injection. Informed selection of the appropriate technique, or combining a technique with transcatheter hepatic arterial embolization according to the tumor size and number, might provide the most effective treatment and achieve better results for hepatocellular carcinoma, even if the liver reserve is not good. However, large-scale, randomized, controlled trials are required before a definitive conclusion can be reached. PMID- 15278183 TI - Antituberculosis drug resistance among retreatment tuberculosis patients in a referral center in Taipei. AB - BACKGROUND AND PURPOSE: To determine the prevalence of antituberculosis drug resistance among retreatment tuberculosis patients in a referral center in Taipei. METHODS: We reviewed the register of susceptibility testing of the mycobacteriology laboratory of the Chronic Disease Control Bureau to identify patients with positive culture for Mycobacterium tuberculosis in the year 2000 2001. Medical charts were reviewed to determine patients' tuberculosis treatment histories. Patients who had multidrug-resistant (MDR) tuberculosis, defined as documentation of isolates resistant to at least isoniazid and rifampin, were identified. Retreatment tuberculosis patients without prior evidence of MDR tuberculosis were classified into 3 categories, i.e., relapse, treatment after default and treatment after failure, and the frequency and patterns of antituberculosis drug resistance were determined. RESULTS: A total of 317 patients who had received antituberculosis treatment for more than 1 month were identified. Among them, 183 were retreatment cases without prior evidence of MDR tuberculosis, including 93 with relapse, 57 with treatment after default, and 33 with treatment after failure. Among the 183 patients, the prevalence of resistance to any drug was 42.6%; 14.2% were resistant to 1 drug, 13.7% to 2 drugs, 7.1% to 3 drugs, 7.7% to 4 drugs or more, and 24.6% had MDR tuberculosis. The prevalence of any drug resistance among patients with relapse, treatment after default and treatment after failure was 33.3%, 42.1%, and 69.7%, respectively, while the prevalence of MDR tuberculosis in these groups was 12.9%, 19.3% and 66.7%, respectively. CONCLUSIONS: If susceptibility results are unavailable, the World Health Organization-recommended retreatment regimen may be used in retreatment tuberculosis patients. However, the high proportion of MDR tuberculosis among patients with treatment after failure poses a challenge to the efficacy of the retreatment regimen. PMID- 15278184 TI - Clinical features and outcome analysis of infective endocarditis in elderly patients. AB - BACKGROUND AND PURPOSE: Diagnosis and management of infective endocarditis (IE) in elderly patients remains a difficult problem. This study evaluated the clinical and microbiologic characteristics and outcome of IE in elderly patients. METHODS: From 1996 to 2002, clinical and microbiologic data from 67 patients with IE aged > or = 65 years at National Taiwan University Hospital were reviewed and analyzed. RESULTS: The median age of the 67 patients was 74 years (range, 66 to 95 years). Staphylococci and streptococci were the 2 leading etiologies of IE. Forty seven of the cases were defined as community-acquired. IE had not been diagnosed at admission in nearly two-thirds of cases. Of the 20 episodes of nosocomial IE, the median interval to development of IE after hospitalization was 30 days. Staphylococci constituted 90% of causative pathogens in nosocomial IE. The in-hospital mortality rate was 29.9%. Univariate analysis revealed that staphylococci as the causative pathogen and nosocomial IE were predictors of fatal outcome (p <0.05). When significant variables related to a fatal outcome on univariate analysis, i.e., staphylococcal IE, nosocomial IE, acute renal failure and cardiac complications, were entered in the multivariate analysis, acute renal failure and development of congestive heart failure or new conduction disturbance were both significantly associated with in-hospital death. CONCLUSIONS: Diagnosis of IE in the elderly was often delayed and a substantial proportion of patients died during hospitalization. Staphylococcal IE and nosocomial IE were significant predictors of in-hospital death. PMID- 15278185 TI - Feasibility and short-term results of total arterial coronary artery bypass in Taiwanese. AB - BACKGROUND AND PURPOSE: Long-term results of coronary artery bypass are limited by progressive atherosclerosis in venous conduits. Arterial conduits are believed to have a better patency rate. A radial artery in composite graft with the left internal thoracic artery makes total arterial revascularization possible in almost all patients. We sought to evaluate the feasibility and short-term results of this grafting strategy. METHODS: A total of 381 consecutive patients received elective coronary artery bypass grafting between 1 May 2000 and 31 August 2002. Patients with associated procedures were also included. There were 333 patients without venous conduits. Patients with left ventricular dysfunction were not excluded from total arterial revascularization. Follow-up time was 13.7 +/- 7.6 months. RESULTS: Total arterial revascularization was achieved in 88% of the patients. Arterial harvest was easy and simple and complete revascularization could be achieved. The 30-day overall mortality rate was 3.29%. In the patients with left ventricle ejection fraction greater than 0.4, the mortality rate was 2.32%. There were 6 late deaths: heart failure in 2, sepsis in 2, respiratory failure in 1, and pneumonia in 1. CONCLUSIONS: It is feasible to use total arterial grafts for coronary artery bypass graft in patients with coronary artery disease. Myocardial revascularization can be complete and the early mortality rate is acceptable. PMID- 15278186 TI - Determinants of in-hospital mortality after surgery for acute type A aortic dissection. AB - BACKGROUND AND PURPOSE: Acute type A aortic dissection presents a formidable challenge for the cardiac surgeon, although remarkable improvements have been achieved in diagnosis, surgical techniques and perioperative management. The aim of this study was to identify the most important variables associated with in hospital mortality in patients undergoing surgery for this condition. METHODS: Between July 1998 and June 2002, 80 patients underwent surgery for acute type A aortic dissection. Univariate and multivariate analyses were performed to identify the variables independently correlated with in-hospital mortality. RESULTS: The overall in-hospital mortality rate was 20% (16/80 patients). Univariate analysis revealed 24 preoperative and operative variables, including type of surgery, cardiopulmonary bypass (CPB) time, aortic cross-clamp time, diabetes mellitus, and postoperative (postoperative 24 hours) bleeding > or = 1500 mL, as factors associated with in-hospital death. Stepwise logistic regression analysis showed the factors independently associated with in-hospital death were CPB time, diabetes mellitus, and postoperative bleeding > or = 1500 mL (p <0.05). CONCLUSIONS: Multiple factors affect in-hospital mortality after surgery for acute type A aortic dissection. This study suggests that CPB time, diabetes mellitus and postoperative bleeding > or = 1500 mL are the main determinants of in-hospital death. PMID- 15278187 TI - Outcome predictors of fulminant hepatic failure in children. AB - BACKGROUND AND PURPOSE: Mortality of fulminant hepatic failure (FHF) is high in children. It is crucial to identify patients with poor prognosis for timely referral to liver transplantation. This study aimed to assess the clinical and laboratory characteristics of children with FHF and evaluate their correlation with outcome. METHODS: Retrospective review was conducted of a total of 31 pediatric patients aged 1 month to 14 years with a diagnosis of FHF from January 1984 to December 2001. Twenty seven clinical and laboratory parameters were analyzed using univariate and multivariate analysis. RESULTS: The etiology of FHF was HBV infection in 10 patients, Wilson's disease in 3, enterovirus infection in 1, herpes simplex virus infection in 1, herpes simplex virus 6 infection with hemophagocytic syndrome in 1, leptospirosis infection in 1, and cryptogenic in 14. Overall mortality was 71% (22/31). The patients who eventually died tended to be older in age, were more likely to have fever during the disease course, had higher peak bilirubin, indirect bilirubin, and ammonia levels, and had more advanced stages of encephalopathy. Multivariate analysis revealed that peak ammonia level > or = 200 microM/L was the only independent factor significantly associated with mortality. The mortality rate in patients with ammonia levels > or = 200 microM/L was 93% compared to 43% in patients with ammonia levels < 200 microM/L (p = 0.013). CONCLUSION: Childhood FHF had a high mortality rate. Ammonia level > or = 200 microM/L was associated with a higher mortality rate. Liver transplantation is recommended for these patients. PMID- 15278188 TI - Predictive factors for QTc prolongation in schizophrenic patients taking antipsychotics. AB - BACKGROUND AND PURPOSE: The rate-corrected electrocardiographic QT (QTc) interval may significantly increase in schizophrenic patients taking antipsychotics. QTc prolongation is a risk factor for development of arrhythmia. The objective of this study was to assess the predictors of QTc prolongation in schizophrenic patients taking antipsychotic medication. METHODS: Electrocardiograms were obtained from 138 controls and 412 schizophrenic inpatients taking antipsychotics. QTc prolongation was defined as a mean value of 2 standard deviations above the controls. Predictors were analyzed with a logistic regression model. RESULTS: Based on data obtained from controls, QTc prolongation was defined as a QTc greater than 421 ms. Logistic regression analysis showed that significant predictors for QTc prolongation were: female gender (odds ratio, 3.355 [95% CI, 1.767-6.371]); increased age (1.040 [1.011-1.069]); and increased doses of some antipsychotics, including clozapine (1.006 [1.003-1.008]), chlorpromazine (1.003 [1.002-1.005]), thioridazine (1.007 [1.003-1.011]), and sulpiride (1.001 [1.001-1.002]). CONCLUSIONS: Predictors of the QTc prolongation in schizophrenic patients taking antipsychotic medications were female gender, old age, and treatment with clozapine, chlorpromazine, thiroridazine, or sulpiride. PMID- 15278189 TI - Bone mineral density in women receiving thyroxine suppressive therapy for differentiated thyroid carcinoma. AB - BACKGROUND AND PURPOSE: Most patients with well-differentiated thyroid carcinoma have an excellent prognosis and are likely to live long enough to be subjected to osteoporosis. The purpose of this study was to investigate the consequences of treatment with a supraphysiological dose of levothyroxine (l-T4) on bone mineral density (BMD) in Taiwanese women with differentiated thyroid cancer. METHODS: A total of 69 (44 premenopausal, 25 postmenopausal) Taiwanese women with differentiated thyroid cancer were included in this retrospective study. These patients were free of disease recurrence after initial near-total thyroidectomy and I-131 radioablation, and had undergone regular l-T4 suppressive therapy for more than 3 years (mean, 7.3 +/- 3.0 years; range, 3 to 15 years). The degree of thyroid-stimulating hormone (TSH) suppression was determined based on the mean TSH score for each patient which was determined by analysis of all available follow-up TSH data, where 1 = undetectable TSH (< 0.2 mIU/mL); 2 = subnormal TSH (0.2 to 0.39 mIU/mL); 3 = normal TSH (0.4 to 4.0 mIU/mL); and 4 = elevated TSH (> 4.0 mIU/mL). The patients were divided into a full TSH suppression group with a mean TSH score in the range 1.0 to 1.99, and a partial TSH suppression group with a mean TSH score in the range 2.0 to 2.99. BMD was measured by dual-energy X-ray absorptiometry at the lumbar spine, femoral neck, Ward's triangle and total hip. Comparisons between subgroups of patients and controls were performed by unpaired t test. Correlation between BMD and other clinical variables was assessed by Pearson's correlation analysis. RESULTS: Postmenopausal patients (aged 57.7 +/- 6.9 years) had significantly higher serum calcium levels and decreased BMD at all sites of the spine and hip as compared with premenopausal patients (aged 38.6 +/- 6.7 years) with similar BMI and duration of TSH suppression. Comparison of BMD between postmenopausal patients and BMI- and age-matched controls revealed that the patient group had decreased BMD at all sites of measurement, although this difference was not significant. This phenomenon was not observed in the premenopausal patients. Furthermore, when BMD was compared between patients categorized as having full and partial suppression of TSH, only patients with full suppression in the postmenopausal group showed a tendency to lower BMD. There was a strong correlation of BMD with age, BMI and serum calcium level. However, no correlation was found between BMD and degree of TSH suppression or duration of l-T4 suppression therapy. CONCLUSION: Women with differentiated thyroid cancer who had long-term (mean, 7.3 +/- 3.0 years) l-T4 therapy and suppressed TSH levels had no evidence of lower BMD. However, patients with full suppression in the postmenopausal group showed a tendency towards lower BMD. Therefore, careful monitoring of BMD in postmenopausal women during suppression therapy is mandatory. PMID- 15278190 TI - Proton magnetic resonance spectroscopy-guided biopsy for cerebral glial tumors. AB - BACKGROUND AND PURPOSE: Although application of proton magnetic resonance (MR) spectroscopy in the diagnosis of brain tumors has been reported, the role of this technique as guidance for targeting biopsy of brain tumors is not well established. The usefulness and limitations of predicting tumor proliferative activity and pathological grading of brain gliomas based on samples obtained from proton MR spectroscopy-guided stereotactic biopsy also remains unclear. The purpose of this study was to evaluate the usefulness of single-voxel MR spectroscopy-guided stereotactic biopsy for cerebral gliomas and to correlate the findings of MR spectroscopy with proliferative activity (measured by Ki-67 labeling index) of tumors and pathological diagnosis. METHODS: Localized proton spectra were obtained before stereotactic/surgical biopsy in 7 patients with glioma (8 lesions). Metabolic values in the spectra were measured semiquantitatively and correlated with the Ki-67 labeling index and pathological grade of each surgical specimen. RESULTS: MR spectroscopy-guided biopsy was effective in obtaining a representative specimen for accurate pathological diagnosis in all patients, including 1 patient with multifocal glioma and 2 with diffusely infiltrated gliomas (gliomatosis cerebri). Those lesions with higher choline complex/creatine ratio (Cho/Cr) and lower N-acetyl-L-aspartate/creatine values in MR spectroscopy were higher grade tumors. Higher Ki-67 labeling index (indicating higher proliferative activity of tumor) with higher Cho/Cr ratios in MR spectroscopy were significantly correlated with tumor grade. CONCLUSIONS: MR spectroscopy-guided biopsy was effective in obtaining a representative specimen for accurate pathological diagnosis, and the Cho/Cr ratio of MR spectra and Ki-67 labeling index were reliable predictors of glioma grade. PMID- 15278191 TI - Lymphadenoma lacking sebaceous differentiation in the parotid gland. AB - Sebaceous lymphadenoma is a rare salivary gland neoplasm that is composed of solid or cystic sebaceous elements in a dense lymphoid background. An even rarer tumor resembling sebaceous lymphadenoma but without the sebaceous element has also been recognized. This unusual tumor has been named "lymphadenoma lacking sebaceous differentiation" (LLSD) or simply "lymphadenoma". Because of the rarity of this entity, detailed clinicopathological manifestations have not been properly characterized. We report a case of LLSD occurring in the left parotid gland of a 49-year-old man. Histologically, the tumor was well circumscribed and composed of solid squamous islands within dense lymphoid stroma. No sebaceous differentiation could be identified in the epithelial nest. The similarity of the histological picture of the tumor to metastatic carcinoma may lead to misdiagnosis. It is therefore important for the pathologist to be aware of the distinguishing features of this unusual entity. PMID- 15278192 TI - Mesenteric adenitis caused by Salmonella enterica serovar Enteritidis. AB - Mesenteric adenitis is a self-limited condition characterized by fever, localized right lower quadrant abdominal pain, and frequent leukocytosis, making it difficult to differentiate from appendicitis. We report a case of mesenteric adenitis in an 8-year-old boy who presented at the emergency department with right lower quadrant abdominal pain, diarrhea, and fever up to 40 degrees C. Acute appendicitis was initially suspected, but further abdominal ultrasound and contrast enhanced computed tomography studies showed a normal appendix with marked mesenteric adenopathy. Symptomatic treatment was given and pain and fever subsided 2 days later. Follow-up sonography showed resolution of adenopathy, confirming the diagnosis of mesenteric adenitis. The admission stool cultures grew Salmonella enterica serovar Enteritidis (S. Enteritidis). Unlike previous reports in western countries where Yersinia species prevails and was thought to be self-limited, S. Enteritidis carries potential risk for serious systemic complications, such as meningitis or septic arthritis. The isolation of this unusual microbiological species thus has both therapeutic and epidemiological implications for mesenteric adenitis in Taiwan. PMID- 15278193 TI - Angiomyofibroblastoma of the vulva. AB - Angiomyofibroblastoma is a rare, recently described, distinctive benign mesenchymal tumor that occurs mainly in the vulvar region of premenopausal women. We report a case of angiomyofibroblastoma of the vulva. A 45-year-old woman reported a small, painless nodule on the left vulva which rapidly enlarged during the 6 months before she visited the hospital. Local examination showed a pedunculated, mobile mass on the superior aspect of the left labium majus (13 x 12 x 10 cm) with elastic texture. Left vulvectomy was performed and the excised specimen revealed a well-defined and multilobulated tumor with edematous changes on gross examination. Microscopically, the tumor had alternate hyper- and hypocellular areas characterized by benign-looking round- to spindle-shaped cells with eosinophilic cytoplasm. The tumor cells were admixed with scattered thin walled vessels in an edematous stroma. Immunohistochemical analysis was diffusely positive for desmin, estrogen receptor and progesterone receptor; reactive in small focus for actin; and negative for S-100 and CD34. Angiomyofibroblastoma, a mesenchymal tumor, was diagnosed. The postoperative course was smooth with no recurrence during the following 8 months. Simple excision for angiomyofibroblastoma is recommended because local recurrence or metastasis has not been reported. PMID- 15278194 TI - [Pulmonary hypertension. Pathophysiology and current concepts of medication therapy]. AB - Chronic pulmonary hypertension (PHT) is characterized by permanently increased pulmonary artery pressure. Diagnostic criteria are a mean pulmonary arterial pressure above 25 mmHg at rest and above 30 mmHg during exercise. Pulmonary arterial hypertension is characterized by progressive obliteration of the pulmonary vascular bed, which results in progressive right heart failure and death. Pathologic processes behind the complex vascular changes associated with PHT include vasoconstrictor/vasodilator imbalance, thrombosis, misguided angiogenesis and inflammation. The function of the pulmonary endothelium is altered with decreased production of vasodilators such as prostacyclin and nitric oxide and an increased production of endothelins, finally resulting in pulmonary vascular remodelling. A new diagnostic classification of pulmonary hypertension (PHT) was proposed at the World Health Organization (WHO) Pulmonary Hypertension Meetings held in Evian in 1998 and in Venice in 2003. This classification reflects recent advances in the understanding of pulmonary hypertensive diseases. Depending on the underlying disease and the localization of the vascular lesion, five different subgroups of PHT are formed. An exact diagnostic classification is necessary for application of the current treatment options for the different forms of PHT. Target of therapy is besides avoiding local thrombosis by anticoagulation and treatment of vasoconstriction, the prevention of vascular remodelling. For patients with advanced pulmonary arterial hypertension (PAH; NYHA stages III and IV) treatment with prostanoids (inhalative, oral, subcutaneous or intravenous), with endothelin-receptor antagonists or with a phosphodiesterase inhibitor can be indicated. Whether initial or adjunct combined therapy provides additional clinical benefits to patients with severe pulmonary arterial hypertension needs further investigation, but first results are promising. PMID- 15278195 TI - [Detection and identification of the pathogenic cause of a brain abscess by molecular genetic methods]. AB - Brain abscesses are life-threatening and detection and identification of the causative pathogens are crucial for substantiating the diagnosis and for selecting the optimal antibiotic regimen. In approximately 20% of the patients microbiological cultures of abscess material remain sterile. The polymerase chain reaction (PCR) provides a methodological alternative, but data about the use of broad spectrum PCR assays to detect the causative pathogens in brain abscesses are rare. We report on the case of a 65-years-old patient with a brain abscess caused by Fusobacterium spp., which was only diagnosed by broad spectrum PCR. To our knowledge this is the second report about a brain abscess, where Fusobacterium spp. was identified only by broad spectrum PCR and subsequent DNA sequencing. PMID- 15278196 TI - [Preclinical management of accidental methadone intoxication of a 4-year-old girl. Antagonist or intubation?]. AB - We report on the preclinical management of a 4-year-old child who was found in a comatose condition with respiratory failure after accidental ingestion of methadone. Emergency airway management was carried out with endotracheal intubation instead of administering the antagonist naloxone. The child could be extubated 12 h later and was released from hospital after 3 days with no neurological symptoms. The authors attempt to formulate an algorithm for the preclinical management of opioid intoxication with reference to the literature and own experience. Endotracheal intubation seems to be superior to the use of the antagonist naloxone, especially in a critical situation. This is the only way to ensure a rapid oxygenation with adequate airway protection and with the simultaneous avoidance of the side-effects of naloxone. A restrictive and critical administration of the opioid antagonist naloxone is recommended when there is suspicion of opioid ingestion but no signs of intoxication. PMID- 15278198 TI - Selection for aneuploid potato hybrids combining a low wild genome content and resistance traits from Solanum commersonii. AB - A breeding scheme based on the production of progenies with odd ploidy was followed to introduce useful genes from the wild Solanum commersonii (cmm) into S. tuberosum (tbr) genome. Hybrids from 5 x x 4 x crosses were characterized for traits of interest, and selection was assisted by amplified fragment length polymorphism (AFLP) analysis. As expected, most of the hybrids were aneuploids, with a trend towards a low degree of aneuploidy. Despite the fact that aneuploidy has often been associated with a reduction in male and female fertility, most of the hybrids were fertile following crosses with tbr, making it possible to produce viable offspring. A screening for resistance traits deriving from cmm was also carried out. With respect to freezing resistance, the killing temperatures of cold-acclimated genotypes were distributed between the wild and cultivated parental values, with some hybrids displaying an acclimation capacity higher than 3 degrees C. A wide variability was also found for tuber soft rot resistance, and hybrids with high levels of resistance were identified. Selection of hybrids was based on a two-stage scheme that consisted of conventional phenotypic selection followed by an estimation of the wild genome content still present in order to identify hybrids combining noteworthy traits with a low wild genome content. Previously selected cmm-specific AFLPs were used to monitor the degree of wild genome content still present in each hybrid. The percentage of cmm-specific markers ranged from 59% to 91%, with an average value of 75%. AFLP analysis was employed to assist in the selection of valuable hybrids for further breeding efforts. PMID- 15278199 TI - Substitutes for genome differentiation in tuber-bearing Solanum: interspecific pollen-pistil incompatibility, nuclear-cytoplasmic male sterility, and endosperm. AB - The cultivated potato, Solanum tuberosum L. (2n=4x=48), has a very large number of related wild and cultivated tuber-bearing species widely distributed in the Americas. These species, grouped in 16 taxonomic series, range from the diploid to the hexaploid level. Polyploid species are either disomic or polysomic, and sexual polyploidization via genetically controlled 2 n gametes has played a major role in their evolution. Species are separated in nature by geographical and ecological barriers. However, there are several examples of sympatric species that share the same niches but do not readily cross (i.e., the diploids S. commersonii and S. chacoense in certain areas of Argentina). External barriers alone are, therefore, not sufficient to explain species integrity. In addition, there is no strong evidence indicating that genome differentiation is important in the group. In this review we present evidence supporting the assertion that interspecific pollen-pistil incompatibility, nuclear-cytoplasmic male sterility, and the endosperm are major forces that strengthen the external hybridization barriers allowing, at the same time and under specific circumstances, a certain amount of gene exchange without jeopardizing the integrity of the species. PMID- 15278200 TI - [Endourethral brachytherapy for the prevention of recurrent strictures following internal urethrotomy]. AB - METHODS AND MATERIALS: Between November 2000 and December 2002 endoscopic incision or transurethral scar resection was followed by endourethral brachytherapy (BT) which was performed in patients with recurrent bulbar strictures ( n=9), bladder neck stenosis after transurethral prostatectomy (TUR P) ( n=3), anastomotic stricture after radical prostatectomy ( n=2) or penile urethral stricture ( n=1). High dose rate (HDR) iridium-192 BT started on the day of the endoscopic incision or resection and continued for the following 3 days. The BT fractionation scheme was 4x3 Gy in the first three patients (until first relapse) and 4x4 Gy in all following patients. The dose was calculated at 3 mm tissue depth using 3-dimensional CT-planning. As of February 2004, the median follow-up of all patients reached 22 months. RESULTS: Seven of 15 patients (46%) are recurrence free. In two patients (13.3%), recurrent strictures developed 12 month later, outside of the region of initial treatment. In six patients (40%) the treatment was considered to be unsuccessful as recurrent strictures were found between 2 and 12 months after the initial or second course of treatment. CONCLUSION: Endourethral brachytherapy after endoscopic incision or resection is a promising treatment for the prevention of recurrent strictures of the urethra, bladder neck or vesicourethral anastomosis. The initial results have been very good, but with longer follow-up recurrence occurred in the irradiated area in 40% of patients. Prospective randomized studies in patients with a strictly defined type of recurrent stricture, or even after the first internal urethrotomy, should be done in the future. PMID- 15278202 TI - [Botulinum toxin in urology. An inventory]. AB - Botulinum toxin (BTX) is highly potent in neurogenic and non-neurogenic voiding disorders. Experience with it in neurourology began 15 years ago in the treatment of neurogenic detrusor-sphincter dyssynergia. Indications were expanded not only to neurogenic detrusor hyperactivity but also to non-neurogenic detrusor hyperactivity, other forms of dysfunctional voiding, and some types of pelvic pain syndrome. Sphincter injections can be recommended for patients with symptomatic post-voiding residual urine due to insufficient detrusor contractility, and detrusor injections can be recommended for patients with neurogenic detrusor hyperactivity in which anticholinergic drugs are not sufficient. Because of the lack of evidence-based studies, botulinum toxin is not approved for urologic use, although there is a desperate need for it. PMID- 15278203 TI - [The significance of comorbidity and age in radical prostatectomy]. AB - The coexistence of another diagnosis beside the index disease under study is defined as comorbidity. Comorbidity increases with advancing age. In candidates for radical prostatectomy, two aspects of comorbidity are of interest: its association with perioperative complications and long-term survival. The Charlson score is the most extensively studied comorbidity classification for the prediction of long-term outcome. Several studies have identified this score as an independent prognostic factor in the prostate cancer setting. In addition to the Charlson score, data collected during the preoperative cardiopulmonary risk assessment may deliver information identifying patients with an increased long term mortality risk. The meaningfulness of comorbidity in predicting overall mortality seems to be comparable to that of the Gleason score, the most important tumor-related predictor of survival in prostate cancer. The identification of prognostically relevant single conditions and the development of a "radical prostatectomy-specific" comorbidity classification might improve the stratification of candidates for radical prostatectomy in the future. PMID- 15278204 TI - [Sexuality in the aged]. AB - The demographic development in the western industrial societies makes the quality of life of older people a very important issue. Leading a satisfying sexual life is part of this. The picture of older people as asexual humans can no longer be maintained. Recent studies show that the fundamental psychosociological need for acceptance, tenderness, warmness and security does not disappear with age. Sexual activity in the age primarily depends on the existence of a partnership. Sexual dysfunctions in aging result from physical, psychological, and partner-related changes, as well as lifestyles. This has to be considered in diagnosis and treatment. Principally, there is no difference between the treatment of older and younger sexual partners PMID- 15278205 TI - [Therapeutic options in the treatment of cartilage defects. Techniques and indications]. AB - BACKGROUND: Cartilage is composed of chondrocytes embedded within an extracellular matrix of collagens, proteoglycans, and noncollagenous proteins. Together, these structures maintain the unique mechanical properties and manifest its striking inability to heal even the most minor injury. METHODS: This review presents the principles of cartilage structure and the biological background of cartilage repair and gives information about the surgical techniques for treating cartilage defects. RESULTS: The response of cartilage to injuries differs from that of other tissues because of its avascularity, the immobility of chondrocytes and the limited ability of mature chondrocytes to proliferate and alter their synthetic patterns. Surgical therapeutic efforts in treating cartilage defects have focused on bringing new cells and tissues capable of chondrogenesis into the lesions and facilitating the access to the vascular system. CONCLUSION: The right indication and the treatment of joint instability and axis deformation are essential for the successful use of cartilage repair procedures. PMID- 15278206 TI - [MRI in the follow-up of matrix-supported autologous chondrocyte transplantation (MACI) and microfracture]. AB - AIM: Matrix-guided autologous chondrocyte implantation (MACI) was compared with microfracture (MFX) to demonstrate the reconstitution of cartilage over a two year period using the morphological capabilities of MRI. PATIENTS AND METHODS: 27 patients (9 females and 18 males, mean age 33 years) underwent MACI on the knee joint. The defects originated from trauma (15 cases), osteochondritis dissecans (8 cases) and chronic repetitive trauma (4 cases) and were localized at the condyles (24 cases) or patella (3 cases). All patients were examined postoperatively after 1, 3, 6, 12 and 24 months with a 1,5 T unit (Gyroscan, Philips) using proton- and T2w spinecho and T1w fatsuppressed 3D gradientecho sequences. We measured the signal intensities of the implant and neighbouring cartilage to calculate the contrast-to-noise ratio (CNR), and the thickness of cartilage and implant layers to define the defect filling rate. Finally, partial and complete remission was defined on MRI and compared with clinical data and morphology on MRI. Additionally, 7 patients were treated with MFX and, subsequently examined on MRI with the same protocol. RESULTS: After MACI, MRI showed a partial but no complete equilibration of signal intensities of implant and adjacent cartilage over the 1 and 2 year follow-up periods which was shown by reduction of CNR from 21 to 10 on 3D-GE and from 26 to 9 on T2w SE sequences. Continuous growth of the implants resulted in an increased filling of the defects starting at 40% after 0.5 year to 85% after 1 or 2 years. Complete remission was found on MRI in 17/27 cases, and remission rate was influenced by etiology of cartilage defect but not by age and gender of patients or size and location of defects. The Lysholm-Gillquist score improved from 49.7 to 97.3. After MFX equilibration of signal intensities and growth of the regenerating fibrous cartilage was less pronounced and complete remission was found in only 2/7 cases. In addition, the clinical score improved from 45.5 to 74.2. CONCLUSION: Direct imaging of cartilage with MRI and assessment of clinical scores allowed improved documentation of the outcome after MACI and MFX. MRI showed that MACI is superior to MFX concerning rate of complete remissions and filling of the defect with regenerating tissue. Clinical examinations showed better scores for MACI than for MFX. PMID- 15278207 TI - [Osteochondritis dissecans of the knee joint]. AB - BACKGROUND: Osteochondritis dissecans (OD) is an affection of the subchondral bone involving progressive detachment of an osteochondral fragment. METHODS: This article describes the epidemiology, etiology, clinical appearance, possibilities for radiological diagnostics, and classification of OD and presents conservative and surgical therapeutic techniques. RESULTS: Treatment of OD depends on the size, location, stability of the fragment, and skeletal maturity. The majority of young patients can be treated conservatively. Surgical interventions include antegrade or retrograde drilling with optional refixation and osteochondral transplantation as well as autologous chondrocyte transplantation. CONCLUSION: The results of surgical intervention are quite promising; nevertheless, further prospective comparative studies are necessary to evaluate effectivity. PMID- 15278213 TI - Symmetry is in the eye of the beeholder: innate preference for bilateral symmetry in flower-naive bumblebees. AB - Bilateral symmetry has been considered as an indicator of phenotypic and genotypic quality supporting innate preferences for highly symmetric partners. Insect pollinators preferentially visit flowers of a particular symmetry type, thus leading to the suggestion that they have innate preferences for symmetrical flowers or flower models. Here we show that flower-naive bumblebees (Bombus terrestris), with no experience of symmetric or asymmetric patterns and whose visual experience was accurately controlled, have innate preferences for bilateral symmetry. The presence of color cues did not influence the bees' original preference. Our results thus show that bilateral symmetry is innately preferred in the context of food search, a fact that supports the selection of symmetry in flower displays. Furthermore, such innate preferences indicate that the nervous system of naive animals may be primed to respond to relevant sensory cues in the environment. PMID- 15278214 TI - Analysis of the Henze precipitate from the blood cells of the ascidian Phallusia mammillata. AB - The Henze precipitate, a peculiar blue-green microparticulate obtained by lysis of the blood cells of the ascidian Phallusia mammillata (Protochordata), was investigated with atomic force microscopy (AFM), scanning electron microscopy (SEM) and X-ray microanalysis. The precipitate was collected from the Henze solution, an unstable red-brown product obtained by treating blood with distilled water, whose degradation yields a characteristic blue-green product. The microparticulates measured 50-100 micro m in diameter and appeared irregular in shape. SEM examination showed smooth, roughly round boundaries. The microparticulate surface examined with AFM appeared as an irregular matrix formed by 70-320-nm-wide mammillate composites, including and embedding small (500-800 nm wide) crystal-like multilayered formations. X- ray analysis showed that the elements present in these same precipitates were mainly C, Si, Al and O. The microparticulate composition appeared close to those of natural waxes or lacquers, embedding amorphous silicates and/or other Si-Al components. The unusual occurrence of Si in ascidian blood and its role are discussed. PMID- 15278215 TI - Brain mechanisms that control sleep and waking. AB - This review paper presents a brief historical survey of the technological and early research that laid the groundwork for recent advances in sleep-waking research. A major advance in this field occurred shortly after the end of World War II with the discovery of the ascending reticular activating system (ARAS) as the neural source in the brain stem of the waking state. Subsequent research showed that the brain stem activating system produced cortical arousal via two pathways: a dorsal route through the thalamus and a ventral route through the hypothalamus and basal forebrain. The nuclei, pathways, and neurotransmitters that comprise the multiple components of these arousal systems are described. Sleep is now recognized as being composed of two very different states: rapid eye movements (REMs) sleep and non-REM sleep. The major findings on the neural mechanisms that control these two sleep states are presented. This review ends with a discussion of two current views on the function of sleep: to maintain the integrity of the immune system and to enhance memory consolidation. PMID- 15278217 TI - Spectrographic analysis of the ultrasonic vocalisations of adult male and female BALB/c mice. AB - In this study, a spectrographic analysis was designed to improve the description of the shape, the modulations, the rate, length and frequencies of BALB/c mouse calls in different behavioural situations. Male and female calls emitted during investigation of cages with clean bedding, soiled with male or female bedding, and during same-sex encounters, were recorded and described. BALB/c male mice uttered different types of vocalisations both when investigating counterpart odour cues and when interacting with same-sex counterparts. BALB/c female mice vocalised solely during same-sex counterpart encounters and it appeared that calls were uttered mainly by the resident females. Male and female mice present a complex array of calls, which seem to be linked to particular behavioural situations. Further studies using this technology may help to improve our understanding of the role of vocal communication in natural rodent populations. PMID- 15278216 TI - Comparative genomics: methods and applications. AB - Interpreting the functional content of a given genomic sequence is one of the central challenges of biology today. Perhaps the most promising approach to this problem is based on the comparative method of classic biology in the modern guise of sequence comparison. For instance, protein-coding regions tend to be conserved between species. Hence, a simple method for distinguishing a functional exon from the chance absence of stop codons is to investigate its homologue from closely related species. Predicting regulatory elements is even more difficult than exon prediction, but again, comparisons pinpointing conserved sequence motifs upstream of translation start sites are helping to unravel gene regulatory networks. In addition to interspecific studies, intraspecific sequence comparison yields insights into the evolutionary forces that have acted on a species in the past. Of particular interest here is the identification of selection events such as selective sweeps. Both intra- and interspecific sequence comparisons are based on a variety of computational methods, including alignment, phylogenetic reconstruction, and coalescent theory. This article surveys the biology and the central computational ideas applied in recent comparative genomics projects. We argue that the most fruitful method of understanding the functional content of genomes is to study them in the context of related genomic sequences. In particular, such a study may reveal selection, a fundamental pointer to biological relevance. PMID- 15278218 TI - Circadian consequences of social organization in the ant species Camponotus compressus. AB - The locomotor activity rhythm of different castes of the ant species Camponotus compressus was monitored individually under laboratory light/dark (LD) cycles, and under continuous darkness (DD). The colony of this ant species comprises two sexual castes, the queens and the males, and three worker castes, namely the major, media, and minor workers. The virgin males and virgin queens display rhythmic activity patterns, but the mated queens were arrhythmic while laying eggs, with the rhythmicity resuming soon after egg-laying. Under the LD regime, major workers showed nocturnal patterns, while about 75% of the media workers displayed nocturnal patterns and about 25% showed diurnal patterns. Under the DD regime, most major workers exhibited circadian rhythm of activity with a single steady state, whereas media workers displayed two types of activity patterns, with activity patterns changing after 6-9 days in DD (turn-arounds). The pre-turn around tau of the ants that showed nocturnal activity patterns during LD entrainment was <24 h after release into DD, which then became >24 h, after 6-9 days. On the other hand, the pre-turn-around tau of those ants that exhibited diurnal patterns during LD entrainment was first >24 h after release into DD, and then became <24 h, after 6-9 days. The activity of the minor workers neither entrained to LD cycles nor showed any sign of free-run in DD. It appears that the circadian clocks of the ant species C. compressus are flexible, and may perhaps depend upon the tasks assigned to them in the colony. PMID- 15278220 TI - Exceptionally high levels of multiple mating in an army ant. AB - Most species of social insects have singly mated queens, although there are notable exceptions. Competing hypotheses have been proposed to explain the evolution of high levels of multiple mating, but this issue is far from resolved. Here we use microsatellites to investigate mating frequency in the army ant Eciton burchellii and show that queens mate with an exceptionally large number of males, eclipsing all but one other social insect species for which data are available. In addition we present evidence that suggests that mating is serial, continuing throughout the lifetime of the queen. This is the first demonstration of serial mating among social hymenoptera. We propose that high paternity within colonies is most likely to have evolved to increase genetic diversity and to counter high pathogen and parasite loads. PMID- 15278221 TI - Does the queen win it all? Queen-worker conflict over male production in the bumblebee, Bombus terrestris. AB - Social insects provide a useful model for studying the evolutionary balance between cooperation and conflict linked to genetic structure. We investigated the outcome of this conflict in the bumblebee, Bombus terrestris, whose annual colony life cycle is characterized by overt competition over male production. We established artificial colonies composed of a queen and unrelated workers by daily exchange of callow workers between colony pairs of distinct genetic make up. Using microsatellite analysis, this procedure allowed an exact calculation of the proportion of worker-derived males. The development and social behavior of these artificial colonies were similar to those of normal colonies. Despite a high worker reproduction attempt (63.8% of workers had developed ovaries and 38.4% were egg-layers), we found that on average 95% of the males produced during the competition phase (CPh) were queen-derived. However, in four colonies, queen death resulted in a considerable amount of worker-derived male production. The different putative ultimate causes of this efficient control by the queen are discussed, and we suggest a possible scenario of an evolutionary arms race that may occur between these two female castes. PMID- 15278222 TI - The bacterium Xenorhabdus nematophila inhibits phospholipases A2 from insect, prokaryote, and vertebrate sources. AB - The bacterium, Xenorhabdus nematophila, is a virulent insect pathogen. Part of its pathogenicity is due to impairing cellular immunity by blocking biosynthesis of eicosanoids, the major recognized signal transduction system in insect cellular immunity. X. nematophila inhibits the first step in eicosanoid biosynthesis, phospholipase A(2) (PLA(2)). Here we report that the bacterium inhibits PLA(2) from two insect immune tissues, hemocytes and fat body, as well as PLA(2)s selected to represent a wide range of organisms, including prokaryotes, insects, reptiles, and mammals. Our finding on a bacterial inhibitor of PLA(2) activity contributes new insight into the chemical ecology of microbe host interactions, which usually involve actions rather than inhibitors of PLA(2)s. PMID- 15278223 TI - Gamergates in the Australian ant subfamily Myrmeciinae. AB - Ant workers can mate and reproduce in a few hundreds of species belonging to the phylogenetically basal poneromorph subfamilies ( sensu Bolton 2003). We report the first occurrence of gamergates (i.e. mated reproductive workers) in a myrmeciomorph subfamily. In a colony of Myrmecia pyriformis that was collected without a queen, workers continued to be produced over a period of 3 years in the laboratory. Behavioural observations and ovarian dissections indicated that three workers were mated and produced the diploid offspring. The Myrmeciinae are thus another taxon in which the selective benefits of sexual reproduction by workers can be investigated. PMID- 15278224 TI - Overestimates of black carbon in soils and sediments. AB - Several recent reports suggest that black carbon (BC), which broadly encompasses charcoal, soot, and other forms of pyrogenic carbon, may constitute a significant proportion of the refractory carbon in soil and sedimentary organic matter. BC is a sink for biospheric and atmospheric carbon dioxide, and is intimately tied to the biogeochemical cycling of both carbon and oxygen through its role in organic matter cycling. Additionally, BC may represent a large fraction of the "missing carbon sink" in global carbon accounting. Here, we demonstrate that documented measurements of BC may be the result of methodological artifacts, which inadvertently overestimate the amount of BC. We found that a widely used thermal oxidative method can create a residue that falls under the operational definition of BC in samples that are relatively BC-free. Moreover, during this procedure, labile organic matter constituents are condensed into pyrogenic carbon, implying that the labile components are present in lesser quantities. These methodological deficiencies are promoting overestimates in the amount of refractory carbon in soil and sedimentary organic matter and may endorse inaccuracies in the rates of carbon fluxes, the mean residence times of terrestrial carbon, and organic matter burial rates in oceanic environments. PMID- 15278225 TI - [Influence of circadian rhythms on cardiovascular function]. AB - The clinical importance of circadian biological rhythms has been strengthened by a number of studies showing a circadian distribution of cardiovascular events like myocardial infarction, stroke, complex arrhythmia, or sudden cardiac death. Incidence of diseases showed a maximum during the early morning hours after awakening from sleep. In addition, a number of pathophysiological mechanisms has been identified to coincide with this peak including blood pressure and heart rate surges, decreased endothelial dilatory capacity of peripheral and coronary arteries, enhanced sympathetic activity, decreased cardiac electrical stability, and increased platelet aggregation. This time window of high risk for the incidence of cardiovascular events has been identified as a target for new treatment and prevention strategies including new release forms of antihypertensive and coronary-dilatory drugs. The use of melatonin as an antihypertensive drug has been successfully explored and opens new opportunities for the management of cardiovascular dysfunction and disease from a circadian perspective. PMID- 15278226 TI - [Frey's syndrome of the external auditory canal]. AB - Frey's syndrome of the external auditory canal is extremely rare. A 55-year old woman presented with a 6 month history of unilateral gustatory otorrhea. She never complained of hearing impairment, tinnitus, vertigo or otalgia. No trauma or surgical signs were evident near the ear or parotid gland. Examination of the ear showed an intact tympanic membrane without disease. A diagnosis of gustatory sweating syndrome was suggested by the observation of sweat production after chewing and by Minor's starch-iodine test. Symptoms were relieved after tympanic neurectomy. The pathogenesis, differential diagnosis and treatment options are discussed. PMID- 15278227 TI - [Chronic nose blockade and rhinorrhea]. PMID- 15278228 TI - [Temporary, undefined, unilateral eyeball movement disorder]. PMID- 15278231 TI - [News in medical jurisprudence from October to April 2004]. PMID- 15278232 TI - [Auditory steady-state response. On the threshold of clinical usage?]. PMID- 15278233 TI - [Olfactory dysfunction. Epidemiology, pathophsiological classification, diagnosis and therapy]. AB - The present manuscript is the result of a collaborative effort within the framework of the Working Group of Olfactology and Gustology of the German Society for ENT, Head and Neck Surgery. It provides a comprehensive overview about the current views on the epidemiology, terminology, diagnostics, and therapy of olfactory dysfunction, and aims to offer a framework for the standardized procedures for the diagnosis and therapy of olfactory disorders. PMID- 15278234 TI - [Sentinel lymph node mapping in gastric and esophageal carcinomas]. AB - During the last 5 years, the concept of sentinel lymph nodes has been investigated in a variety of solid tumors. Despite the multidirectional and complex lymphatic drainage of the stomach, early gastric cancer has been shown to be a suitable model for sentinel lymph node mapping. In contrast, sentinel lymph node mapping of esophageal cancer is compromised by the anatomic location of the esophagus and its lymphatic drainage in the closed space of the mediastinum. The technique and clinical application of sentinel lymph node mapping thus differ between esophageal and gastric cancer. Reliable detection of sentinel lymph nodes in the mediastinum requires radioisotope labelling, while blue dye and radioisotope labelling are both feasible for gastric cancer. In patients with early gastric cancer, laparoscopic resection with sentinel node negative status is already under investigation in clinical trials. In esophageal cancer, sentinel node mapping is still considered an experimental technique. Preliminary data, however, indicate that it may be reliable and feasible in patients with early adenocarcinoma of the distal esophagus. PMID- 15278235 TI - [Humeral shaft fractures]. AB - Since Lorenz Bohler postulated in his 1964 summary with the title "Against the operative treatment of fresh humeral shaft fractures" that the operative treatment is the exception in the therapy of humeral fractures times have changed. In the last years a conservative treatment of a humeral fracture is the exception and only used after straight indications. The operative therapy nowadays is the gold standard because of the development of new intramedullar and rotation stable implants in addition to the classical osteosynthesis with the plate. But even the external fixator for primary stabilisation in polytrauma patients or as rescue procedure after complications should be in repertory of every orthopedic surgeon. Attention should be put on the avoidance of primary and the correct treatment of secondary nerval lesions, esp. of the radial nerve. Here we are tending to the operative revision of the nerve in indistinct cases. In the treatment of the seldom humeral shaft fracture of the child conservative treatment is to prefer; in complications a resolute shift to a final operative stabilisation of the fracture is necessary. PMID- 15278236 TI - [Sentinel lymph node in melanoma]. AB - The procedure of sentinel node biopsy (SNB) has emerged as an important advance especially with respect to staging of malignant melanoma. Elective (prophylactic) lymph node dissection that had been practiced in primary melanoma with a suspected increased risk of (clinically occult) lymphatic metastasis has been replaced by SNB. Patients with proven metastatic involvement of the sentinel node (12-25%) can be specifically selected for regional lymph node dissection. Metastatic involvement of the sentinel node (SN) is a significant independent prognostic factor. The value of detecting metastasis by highly sensitive diagnostic tools such as RT-PCR is just as uncertain as is the therapeutic benefit of operative or conservative therapies in sentinel node-positive patients with respect to improving prognosis and is currently under study. PMID- 15278240 TI - Oxidation and ring cleavage of dibenzofuran by the filamentous fungus Paecilomyces lilacinus. AB - The ability of the imperfect soil fungus Paecilomyces lilacinus to transform the environmental pollutant dibenzofuran was investigated. Transformation of dibenzofuran and related derivatives lead to 14 products, which were identified by UV spectroscopy, mass spectrometry, and proton nuclear magnetic resonance spectroscopy. Biotransformation was initiated by two separate hydroxylation steps, leading to the accumulation of 4-monohydroxylated and 4 dihydroxylateddibenzofurans. Hydroxylation at both aromatic rings produced 2,7 dihydroxydibenzofuran, 3,7-dihydroxydibenzofuran, and 2,8-dihydroxydibenzofuran. Further oxidation yields ring cleavage of dibenzofuran, which has not been described before for filamentous fungi. The ring fission products were identified as benzo[ b]furo[3,2-d]-2-pyrone-6-carboxylic acid and [2-(1-carboxy-methylidene) benzofuran-3-ylidene]-hydroxy-acetic acid and its derivatives hydroxylated at carbon 7 and 8 at the non-cleaved ring. Other metabolites were riboside conjugates of 2-hydroxydibenzofuran and 3-hydroxydibenzofuran. The results showed that P. lilacinus transforms the hydrophobic compound dibenzofuran by phase I/phase II reactions to produce hydroxylated products and excretable sugar conjugates. PMID- 15278241 TI - Pseudomonas aeruginosa dihydroorotases: a tale of three pyrCs. AB - Pseudomonas aeruginosa PAO1 was shown to contain three pyrC sequences. Two of these genes, designated pyrC (PA3527) and pyrC2 (PA5541), encode polypeptides with dihydroorotase (DHOase) activity, while the third, pyrC' (PA0401), encodes a DHOase-like polypeptide that lacks DHOase activity, but is necessary for the structure and function of ATCase. Both pyrC and pyrC2 were cloned and complemented an Escherichia coli pyrC mutant. In addition, pyrC and pyrC2 were individually inactivated in P. aeruginosa by homologous exchange with a mutated allele of each. The resulting mutant strains were prototrophic. A pyrC, pyrC2 double mutant was also constructed, and this strain had an absolute requirement for pyrimidines. The transcriptional activity of pyrC and pyrC2 was measured using lacZ promoter fusions. While pyrC was found to be constitutively expressed, pyrC2 was expressed only in the pyrC mutant background. An in vitro transcriptional/translational system was used to estimate the size of the pyrC2 gene product. The expressed polypeptide was approximately 47 kDa, which is in keeping with the theoretical molecular mass of 48 kDa, making it the largest prokaryotic DHOase polypeptide identified to date. To our knowledge, this is the first report of a true DHOase mutant in P. aeruginosa and also the first confirmation that pyrC2 encodes a polypeptide with DHOase activity. PMID- 15278242 TI - Genetic diversity in Rhizopus oryzae strains as revealed by the sequence of lactate dehydrogenase genes. AB - Twenty-seven strains of Rhizopus oryzae accumulating predominantly lactic acid were shown to possess two ldh genes, ldhA and ldhB, encoding NAD-dependent lactate dehydrogenases. Variation in nucleotide sequence was identified for each gene from different strains, and similar phylogenetic trees were obtained based on the nucleotide sequences of both genes. The other 21 strains of R. oryzae accumulating predominantly fumaric and malic acids contained a single ORF of ldhB. Compared to the strains accumulating predominantly lactic acid, a lower degree of sequence divergence was found in ldhB, resulting in a separate cluster in the phylogenetic tree. The high similarity (>90%) spanning the ORF and adjacent regions demonstrates that ldhA and ldhB are derived from the same ancestor gene. The strains accumulating predominantly fumaric and malic acids lack functional ldhA, which plays a role in lactic acid synthesis and may form a lineage separated from the strains accumulating predominantly lactic acid in the genus Rhizopus. PMID- 15278243 TI - Ribosome modulation factor protects Escherichia coli during heat stress, but this may not be dependent on ribosome dimerisation. AB - The role of ribosome modulation factor (RMF) in protecting heat-stressed Escherichia coli cells was identified by the observation that cultures of a mutant strain lacking functional RMF (HMY15) were highly heat sensitive in stationary phase compared to those of the parent strain (W3110). No difference in heat sensitivity was observed between these strains in exponential phase, during which RMF is not synthesised. Studies by differential scanning calorimetry demonstrated that the ribosomes of stationary-phase cultures of the mutant strain had lower thermal stability than those of the parent strain in stationary phase, or exponential-phase ribosomes. More rapid breakdown of ribosomes in the mutant strain during heating was confirmed by rRNA analysis and sucrose density gradient centrifugation. Analyses of ribosome composition showed that the 100S dimers dissociated more rapidly during heating than 70S particles. While ribosome dimerisation is a consequence of the conformational changes caused by RMF binding, it may not therefore be essential for RMF-mediated ribosome stabilisation. PMID- 15278247 TI - Special report on the official positions of the International Society for Clinical Densitometry. AB - The International Society for Clinical Densitometry (ISCD) periodically holds Position Development Conferences (PDCs) for the purpose of establishing standards and guidelines for indications, acquisition, and interpretation of bone density tests. Topics are selected for consideration by the ISCD Scientific Advisory Committee, reviewed by scientific working groups, and presented to an international panel of experts. Topic categories addressed to date include indications for bone density testing, selection of reference databases for T scores and Z-scores, clinical applications for central and peripheral bone densitometry, serial bone density testing, instrument precision assessment, phantom scanning and calibration testing, requirements for a bone density report, nomenclature, and diagnosis of osteoporosis in postmenopausal women, premenopausal women, men, and children. Following an open session for public comment and discussion, the panel convenes for consideration of each topic and makes recommendations for positions to the ISCD Board of Directors. Recommendations that are accepted become the Official Positions of the ISCD. This Special Report summarizes the methodology of the ISCD PDCs and presents selected Official Positions of general interest. PMID- 15278250 TI - Anterior enterocele: a report of three cases. AB - Anterior enterocele is an uncommon finding in patients with pelvic organ prolapse. We reviewed 490 consecutive operations for pelvic organ prolapse . Three anterior enteroceles were identified in a series of 193 enterocele repairs (1.6%). The presentation and treatment of each of these patients is reviewed. PMID- 15278249 TI - Sexual function after tension-free vaginal tape (TVT) for stress incontinence: results of a mailed questionnaire. AB - In a retrospective study we evaluated sexual function after tension-free vaginal tape (TVT) placement for urinary stress incontinence based on responses to a mailed questionnaire at least 3 months after the operation, to a maximum of 1 year. From 1999 to 2002, a sexual function questionnaire was mailed to 128 women (and their partners) who had undergone a TVT procedure for genuine urinary stress incontinence, without pelvic organ prolapse or detrusor instability. The questionnaire was returned by 96 women (75%), 69 (72%) of whom reported being sexually active. Mean frequency of intercourse did not change. Overall, 26% described improved intercourse compared to before the operation. Only one patient described worsening of intercourse after the TVT operation because of an increase in her incontinence. Overall, in this study the technique of tension-free vaginal tape as such seems to have no negative impact on sexual function. However, because of its successful outcome on incontinence, it has a positive overall effect on sexual function. The possible causes of postoperative partner discomfort require further investigation. PMID- 15278251 TI - Is the leak point pressure alone an accurate indicator of intrinsic sphincteric deficiency? AB - The aim of this study was to determine the characteristics of women who meet the criteria for intrinsic sphincteric deficiency (ISD) on maximum urethral closure pressure (MUCP) but not on leak point pressure (LPP) measurement. We performed a cross-sectional chart review of every patient who underwent multichannel, microtransducer urodynamic testing in our center between 1994 and 1996 (n=423). From this population we culled a sub-population of women who fit into one of the following two groups: women with no evidence of ISD on MUCP or LPP and women with evidence of ISD on MUCP only. Logistic regression was used to identify independent predictors of group membership. Increasing age (>60.5 years) and a positive supine empty stress test were the only independent predictors of membership in the group of women with ISD on MUCP only. Knowledge of these risk factors may help clinicians in choosing appropriate pre-operative testing. PMID- 15278252 TI - Urinary retention caused by adenocarcinoma arising in recurrent urethral diverticulum. AB - We present a case of adenocarcinoma arising in a urethral diverticulum. A 71-year old woman presented with urinary retention and a mass in the anterior vaginal wall. A biopsy was performed and revealed adenocarcinoma. Anterior pelvic exenteration with continent urinary diversion was performed followed by adjuvant radiation therapy as the tumor was large and poorly differentiated. She developed stomal stenosis and underwent stomal revision. The patient has done well since, with no evidence of recurrence at 1-year follow-up at the time of this report. PMID- 15278253 TI - The tensile strength of Cooper's ligament suturing: comparison of abdominal and transvaginal techniques. AB - This study was designed to compare the strength and position of sutures anchored into Cooper's ligament utilizing a minimally invasive transvaginal suturing technique, versus the 'open' abdominal approach. In 12 fresh cadavers, Cooper's ligament was accessed via abdominal and vaginal incisions. After randomization, polytetrafluoroethylene (00) sutures were spaced along one ligament with the transvaginal device (n=36). Contralaterally, sutures were placed abdominally (n=36). Progressive load was applied until suture breakage or dislodgement, and tensile strength was measured using a digital tensiometer. Peak tension averaged 14.5 psi for abdominal and 12.96 psi for vaginal (p=0.28). Suture breakage rather than ligament 'pullout' was more likely for abdominal (95 vs. 56%, p=0.0001). Vaginal and abdominal sutures demonstrated nearly identical mean distances from mid-symphysis (4.62 vs. 4.24 cm, p=0.56). Peak tension was not correlated with suture location (r2=0.17, p=0.28). We conclude that transvaginal suturing, using the minimally invasive device, achieved similar tensile strength and position to the open technique. Transvaginal sutures were associated with greater likelihood of ligament 'pullout' before suture breakage under maximal load; however, the clinical implications of this finding are uncertain. PMID- 15278254 TI - Utility of urine reagent strip in screening women with incontinence for urinary tract infection. AB - The aim of this study was to determine the utility of urine reagent strips in screening women with urinary incontinence for urinary tract infections. Subjects were all women presenting with urinary incontinence. A urine specimen was screened for urinary tract infection with a urine reagent strip and urine culture. Using the urine culture result as the gold standard, the sensitivity, specificity, and positive and negative predictive values of the urine reagent strips were calculated. A total of 265 pairs of reagent strips and urine cultures were evaluated. Thirty-one cultures grew out of a single organism; nine of these had a positive urine reagent strip, yielding a sensitivity of 29%. The specificity of the urine reagent strip was 99%, the positive predictive value was 82% and the negative predictive value was 92%. For women presenting with urinary incontinence, the sensitivity of a urine reagent test for diagnosing urinary tract infection was low. PMID- 15278255 TI - Pelvic floor muscle strength and thickness in continent and incontinent nulliparous pregnant women. AB - The aim of the study was to measure pelvic floor muscle function in continent and incontinent nulliparous pregnant women. The study group consisted of 103 nulliparous pregnant women at 20 weeks of pregnancy. Women reporting urinary incontinence once per week or more during the previous month were classified as incontinent. Function was measured by vaginal squeeze pressure (muscle strength) and increment in thickness of the superficial pelvic floor muscles (urogenital diaphragm) assessed by perineal ultrasound. Seventy-one women were classified as continent and 32 women as incontinent. Continent women had statistically significantly higher maximal vaginal squeeze pressure and increment in muscle thickness when compared with incontinent women. There was a strong correlation between measurements of vaginal squeeze pressure and perineal ultrasound measurements of increment in muscle thickness. This study demonstrates statistically significant differences in pelvic floor muscle function measured by strength and thickness in continent compared with incontinent nulliparous pregnant women. PMID- 15278257 TI - Paravaginal defects: prevalence and accuracy of preoperative detection. AB - The objective of this study was to determine the prevalence of paravaginal defects and to report the correlation between diagnosing a paravaginal defect preoperatively and observing the presence of one intraoperatively. This was a prospective study in which 77 patients with at least stage 2 prolapse of the anterior vaginal wall who desired surgical correction of their prolapse were assessed pre- and intraoperatively for the detection of a paravaginal defect. In order to differentiate a midline or central defect from a paravaginal defect, an index finger or ring forceps was placed vaginally toward each ischial spine separately. If the prolapse became reduced, the patient was clinically diagnosed with a paravaginal defect on that side. The intraoperative visualization or palpation of the pubocervical fascia detached from the arcus tendineus fasciae pelvis was used as the gold standard in diagnosing a paravaginal defect. The overall prevalence of a paravaginal defect in patients with at least stage 2 prolapse of the anterior vaginal wall was 37.7%. The sensitivities for detecting a left, right and bilateral paravaginal defect were 47.6, 40.0 and 23.5%, respectively, while the specificities for each side were 71.4, 67.3, and 80.0%, respectively. The overall prevalence of a paravaginal defect in patients with anterior vaginal wall prolapse is low. The standard clinical evaluation used to preoperatively detect a paravaginal defect in our hands is a poor predictor for the actual presence of a paravaginal defect. PMID- 15278258 TI - Voiding dysfunction in young, nulliparous women: symptoms and urodynamic findings. AB - The objective was to determine urodynamic findings in young, premenopausal, nulliparous women with bothersome lower urinary tract symptoms and assess whether or not symptoms are predictive of specific urodynamic abnormalities. The records of 57 women were reviewed. Those with neurological disease or a primary complaint of stress incontinence were excluded. All completed the American Urological Association Symptom Index (AUASI) and underwent videourodynamics. Symptoms were compared in patients with and without bladder dysfunction and/or voiding phase dysfunction. Bladder dysfunction was diagnosed in 86% of patients with urge incontinence vs. 17% of those without (p<0.0001). Patients with voiding phase dysfunction had higher total and voiding AUASI scores. Occult neurological disease was later diagnosed in 4 women (24%) with urge incontinence and bladder dysfunction. Urge incontinence and voiding symptoms are frequently associated with urodynamically demonstrable abnormalities. Urge incontinence and bladder dysfunction may be a sign of occult neurological disease in this population. The presenting symptoms are useful in determining the utility of urodynamics in this population. PMID- 15278263 TI - Apoptosis in right-ventricle biopsy is not predictive of graft survival. AB - Myocardial dysfunction is common in grafted hearts from brain-dead donors, but the mechanisms involved remain unclear, although apoptosis has been suggested to play an important role. In this study, we investigated the presence of apoptotic myocardial cells in donor hearts as compared to control hearts to determine whether pre-existing apoptosis can predict donor heart dysfunction. Apoptosis was studied by in situ DNA fragmentation assay and by Western Blotting for caspase-3, the pivotal executive caspase of the apoptotic pathway. We show that brain-death induced myocardial apoptosis was not predictive of myocardial dysfunction in transplanted hearts. PMID- 15278264 TI - Successful treatment of accelerated vascular rejection in a highly immunised renal transplant recipient with immunoadsorption and 15-deoxyspergualin. PMID- 15278265 TI - Consumption of blood products during mechanical circulatory support in children: comparison between ECMO and a pulsatile ventricular assist device. AB - OBJECTIVE: Different mechanical circulatory support (MCS) systems are used in children with intractable heart failure. However, the need for anticoagulation leads to hemorrhage with subsequent use of blood products. We compared the coagulation disorders and the need for blood products in children treated either with extracorporeal membrane oxygenation (ECMO) or with the Berlin Heart pulsatile pneumatic ventricular assist device. PATIENTS: We retrospectively reviewed the first 8-day course of 64 children who were on MCS for more than 2 days between 1990 and 2002. Thirty children (median age 7.4 years, weight 25.5 kg) received Berlin Heart support and 34 children (median age 1.8 years, weight 9.2 kg) ECMO. Anticoagulation was accomplished by continuous infusion of heparin. Red blood cell count, platelet count, aPTT, AT III, fibrinogen, and ACT were measured regularly. Depending on blood loss and the coagulation disorder, red blood cells, fresh frozen plasma, platelets, and AT III were substituted. RESULTS: There were no preoperative differences in hematological parameters between the two groups. In the Berlin Heart group platelet transfusion was 4.3 ml x kg x day vs 24.6 ml x kg x day in the ECMO group. Red blood cell substitution was 17.2 vs 60.3 ml.kg.day. Fresh frozen plasma substitution was 8.5 ml x kg x day vs 46.9 ml x kg x day (P<0.001). Even in the congenital heart defect subgroups, when MCS was implanted without recent cardiotomy, the differences were significant. Nevertheless, the mean daily values for hemoglobin, platelets, and fibrinogen were lower in the ECMO group. There was lower overall mortality in the Berlin Heart group. CONCLUSIONS: Compared to ECMO, use of the Berlin Heart in children results in less blood loss and lower consumption of red blood cells, platelets, and fresh frozen plasma. PMID- 15278266 TI - The longer patients are in hospital before Intensive Care admission the higher their mortality. AB - OBJECTIVE: To explore the relationship between hospital mortality and time spent by patients on hospital wards before admission to the intensive care unit (ICU). DESIGN: Observational study of prospectively collected data. SETTING: Participating intensive care units within the North East Thames Regional Database. PATIENTS AND PARTICIPANTS: Patients, 7,190, admitted to ICU from the hospital wards of 24 hospitals. INTERVENTIONS: None. MEASUREMENTS AND RESULTS: Of ICU admissions from the wards, 40.1% were in hospital for more than 3 days and 11.7% for more than 15 days. ICU patients who died in hospital were in-patients longer (p=0.001) before admission (median 3 days; interquartile range 1-9) than those discharged alive (median 2 days; interquartile range 1-5). Hospital mortality increased significantly (p<0.0001) in relation to time on hospital wards before ICU: 47.1% (standardised mortality ratio 1.09) for patients in hospital 0-3 days before ICU admission up to 67.2% (standardised mortality ratio 1.39) for patients on the wards for more than 15 days before ICU. Length of stay before ICU admission was an independent predictor of hospital mortality (odds ratio per day 1.019; 95% confidence interval 1.014-1.024). There were significant differences (p<0.001) in patient age, APACHE II score and predicted mortality in relation to time on wards before ICU admission. CONCLUSIONS: Mortality was high among patients admitted from the wards to ICU; many were inpatients for days or weeks before admission. The longer these patients were in hospital before ICU admission, the higher their mortality. Patients with delayed admission differed in some respects compared to those admitted earlier. PMID- 15278268 TI - Procoagulant and fibrinolytic activity in ventilator-associated pneumonia: impact of inadequate antimicrobial therapy. AB - OBJECTIVE: To determine the homeostatic balance of patients with ventilator associated pneumonia (VAP) with respect to the adequacy of antimicrobial therapy. DESIGN AND SETTING: Descriptive observational study in a 12-bed medical intensive care unit in a university-affiliated hospital. PATIENTS: Twenty-nine patients with VAP documented by quantitative culture of bronchoalveolar secretions and a control group of eight mechanically ventilated patients. METHODS: Serial bronchoalveolar lavage fluid (BALF) samples were assayed for prothrombin activation fragment (F1+2), thrombin-antithrombin (TAT) complex, fibrinolytic activity, urokinase-type plasminogen activator (u-PA), and plasminogen activator inhibitor type 1 (PAI-1) on days 1, 4, and 7 after VAP onset. RESULTS: Pathogens isolated from patients with inadequate empirical antimicrobial coverage included methicillin-resistant Staphylococcus aureus (n=2), Pseudomonas aeruginosa (n=4), and Acinetobacter baumannii (n=1). Compared to those who received adequate antibiotic therapy, TAT, F1+2, and PAI-1 levels increased while u-PA levels remained unchanged. Despite antibiotic adjustment on day 4, TAT levels remained elevated in those who lacked adequate antimicrobial coverage and were significantly correlated with PaO(2)/FIO(2). The procoagulant activity was accompanied by a local depression of fibrinolytic capacity that was attributed mainly to increased BALF PAI-1 levels. Nonsurvivors showed significantly higher levels of TAT and PAI-1 than survivors. No significant correlation between the bacterial burden and the homeostatic derangements was documented. CONCLUSIONS: The lung inflammatory response seems to promulgate a local procoagulant activity associated with hypoxemia in those with inadequate antibiotic therapy. The homeostatic derangement seems to be independent of the lung bacterial burden. PMID- 15278267 TI - Coagulation disorders of cardiopulmonary bypass: a review. AB - BACKGROUND: Postoperative bleeding is one of the most common complications of cardiac surgery. DISCUSSION: Extensive surgical trauma, prolonged blood contact with the artificial surface of the cardiopulmonary bypass (CPB) circuit, high doses of heparin, and hypothermia are all possible triggers of a coagulopathy leading to excessive bleeding. Platelet activation and dysfunction also occur and are caused mainly by heparin, hypothermia, and inadequate protamine administration. Heparin and protamine administration based on heparin concentrations as opposed to fixed doses may reduce coagulopathy and postoperative blood loss. CONCLUSIONS: A better comprehension of the multifactorial mechanisms of activation of coagulation, inflammation, and fibrinolytic pathways during CPB may enable a more effective use of the technical and pharmaceutical options which are currently available. PMID- 15278269 TI - Comment on "Role of vasopressin in the management of septic shock" by Mutlu and Factor. PMID- 15278271 TI - Comment on "Surviving sepsis campaign guidelines for the management of severe sepsis and septic shock" by Dellinger et al. PMID- 15278272 TI - Pulse oximetry. PMID- 15278273 TI - Benefits of minocycline and rifampin-impregnated central venous catheters. A prospective, randomized, double-blind, controlled, multicenter trial. AB - OBJECTIVE: To determine the efficacy of minocycline and rifampin-impregnated catheters compared to non-impregnated catheters in critically ill patients. DESIGN: Prospective, randomized, double-blind, controlled, multicenter trial. SETTING: Intensive care units of seven acute-care teaching hospitals in Spain. PATIENTS. Intensive care unit patients requiring triple-lumen central venous catheter for more than 3 days. INTERVENTIONS: At catheter insertion, 228 patients were randomized to minocycline and rifampin-impregnated catheters and 237 to non impregnated catheters. Skin, catheter tip, subcutaneous segment, hub cultures, peripheral blood and infusate cultures were performed at catheter withdrawal. The rate of colonization, catheter-related bloodstream infection (CRBSI) and catheter related clinical infectious complications (purulence at the insertion site or CRBSI) were assessed. MEASUREMENTS AND MAIN RESULTS: In the intention-to-treat analysis (primary analysis), the episodes per 1000 catheter days of clinical infectious complications decreased from 8.6 to 5.7 (RR =0.67, 95% CI 0.31-1.44), CRBSI from 5.9 to 3.1 (RR =0.53, 95% CI 0.2-1.44) and tip colonization from 24 to 10.4 (RR =0.43, 95% CI 0.26-0.73). Antimicrobial-impregnated catheters were associated with a significant decrease of coagulase-negative staphylococci colonization (RR =0.24, 95% CI 0.13-0.45) and a significant increase of Candida spp. colonization (RR =5.84, 95% CI 1.31-26.1). CONCLUSIONS: The use of antimicrobial-impregnated catheters was associated with a significantly lower rate of coagulase-negative staphylococci colonization and a significant increase in Candida spp. colonization, although a decrease in CRBSI, increase in 30-day survival or reduced length of stay was not observed. PMID- 15278274 TI - [Botulinum toxin A in the treatment of infantile cerebral palsy. Taking into account multilevel, integrated treatment]. AB - Botulinum toxin A represents a significant development in the management of children and adolescents with spastic cerebral palsy. Prerequisites for an adequate result are a correct indication, an exact injection technique and an intensive post-treatment programme. Spastic muscle overactivity and the constant tendency of the involved muscles to shorten with growth cannot be treated by only one method. Therefore a multilevel approach and an integrated treatment schedule including plaster of Paris, orthoses and physiotherapy are currently the best ways to modify the disease process. The inclusion of objective clinical documentation techniques combined with 3-D instrumented gait analysis allows the determination of the indications more exactly and for monitoring the post treatment results. If started early and correctly, this integrated management approach has the potential to modify the natural history of the disorder, and to reduce the frequency of later surgery. PMID- 15278275 TI - [Titanium deposits on the ceramic heads of dislocated total hip replacements]. AB - INTRODUCTION: In cases of revision total hip replacement (THR) having acetabular titanium components with dislocations in the past, metallic deposits are frequently found on the ceramic heads. The aim of this study was to determine whether the metallic deposits increase surface roughness in definite areas. MATERIALS AND METHODS: Ten ceramic heads of revised THR which showed metallic deposits were investigated. The patients suffered from one to six dislocations. The prosthesis lifetime ranged from 2 weeks to 12 years. Surface structure was investigated in a scanning electron microscope (LEO 1525), and the metallic deposits characterised by means of energy dispersive x-ray analysis (EDX). RESULTS: In the area of the metallic deposits, titanium was detected by EDX. Edges with a significant increase in surface roughness were observed, partially accompanied by damage to the surface structure and loosening of Al(2)O(3) particles. DISCUSSION: Titanium deposits increase surface roughness in definite areas, which probably causes wear to the ceramic-ceramic or ceramic-polyethylene articulations due to different roughness values and surface properties. Further investigations are necessary in order to determine the importance of these findings with respect to wear and loosening of total hip arthroplasty. PMID- 15278276 TI - [Accusations of malpractice in chirotherapeutic treatment]. AB - BACKGROUND: Little is known about the incidence of adverse effects after chiropractic manipulation. Over representation of severe and under representation of less severe complications has to be assumed. MATERIAL AND METHODS: A total of 57 expert opinions from the malpractice advisory board of the North Rhine General Medical Council (Nordrheinische Arztekammer), as well as judgments from German courts since 1949, were analyzed. RESULTS AND CONCLUSIONS: A total of 16 of 57 cases of chiropractic manipulation (since 1975) were attested as malpractice by expert opinion, seven of which had significant negative consequences. In nine cases, the judgments of German courts refer to manual therapy. Of these, five deal with informed consent. Observance of the "Bingen Declaration" would have avoided all cases of malpractice found by the advisory board over 29 years. PMID- 15278277 TI - [Heparin induced thrombocytopenia. A frequently unrecognised complication after major orthopedic surgery]. AB - The application of unfractioned (UFH) and low molecular weight heparins (LMH) has reduced the incidence of thromboembolic events. However, the frequency of heparin induced thrombocytopenia (HIT II) in orthopedic patients, particularly susceptible for both thromboembolic complications and HIT II with potentially life threatening complications, is about 0.5% for LMW and 3% UFH. Induced by an immune response, the excessive activation of platelets and endothelial cells causes massive thrombin generation and, as a result, thrombotic vessel occlusion. The rates of mortality and amputation in HIT II are estimated to be 30% and 20%, respectively. The clinical course is highly dependent on early therapeutic intervention, consisting of compatible and adequately dosed anticoagulation drugs. Vitamin K antagonists as well as platelet substitution may lead to disastrous sequelae. We summarize the current state of the pathophysiology, diagnosis and therapy of HIT II and illustrate therapeutic mistakes in a case report. PMID- 15278278 TI - Screening of depression in patients with diabetes mellitus. PMID- 15278279 TI - Creating a pandemic of prediabetes: the proposed new diagnostic criteria for impaired fasting glycaemia. AB - AIMS/HYPOTHESIS: In November 2003 the American Diabetes Association expert committee on the diagnosis and classification of diabetes mellitus suggested a revision of the diagnostic criteria for IFG, lowering the diagnostic threshold from 6.1 to 5.6 mmol/l. The aim of the present study was to evaluate the consequences of this change with respect to: (i) the prevalence of IFG in five different countries; (ii) the concordance between IFG and IGT (classification of individuals); and (iii) the cardiovascular risk profile of these groups. Finally we discuss the likelihood that intervention for cardiovascular risk and prevention strategies developed for individuals with IGT are applicable to subjects with IFG. METHODS: The first part of the study is based on the population-based Danish Inter99 study, where 6265 individuals, aged 30 to 60 years and without previously diagnosed diabetes, underwent an oral glucose tolerance test. The second part is based on the DETECT-2 project, in which studies from China, India, France and USA were used to analyse the impact of the proposed revision of the diagnostic criteria in different ethnic groups. RESULTS: The proposed change in diagnostic criteria would increase the prevalence of IFG in Denmark from 11.8 to 37.6%. The proposed IFG category would identify 60.0% of all subjects with IGT compared to 29.2% with the old criteria, but among individuals with the new IFG category only 18.5% would also have IGT. Individuals with isolated IFG had lower insulin levels and a lower cardiovascular risk profile with the proposed criteria compared with the current WHO criteria. Data from the DETECT-2 study confirmed the marked increase in the prevalence of IFG, and the estimated number of individuals in the age range 40 to 64 years with IFG in urban India, urban China and the USA would increase by 78%, 135% and 193% respectively. CONCLUSIONS/INTERPRETATION: The proposed revised diagnostic criteria will lead to a dramatic increase in the prevalence of IFG, but the concordance rate between IFG and IGT remains low. This new IFG group will have a more favourable cardiovascular risk profile than the current IFG group as defined by the WHO. This seriously questions whether the existing intervention strategies are applicable to the new category of individuals with IFG. PMID- 15278280 TI - Davis BJ, Forbes JM, Thomas MC et al. (2004) Superior renoprotective effects of combination therapy with ACE and AGE inhibition in the diabetic spontaneously hypertensive rat. Diabetologia 47:89-97. PMID- 15278281 TI - The changing phenotype of the human species (affluent variety). PMID- 15278282 TI - Open-nucleus breeding strategies compared with population-wide positive assortative mating: I. Equal distribution of testing efforts. AB - Positive assortative mating (PAM) can enhance the additive genetic variance in a breeding population(BP). This increases the potential for gains in the production population (PP, selected subset of the BP) for recurrent selection programs in forest trees. The assortment of mates can be either: (1) by individual tree rank across the whole BP (PAM), or (2) trees of similar rank can be merged into larger hierarchical groups and then mated randomly within group ("open"-nucleus breeding,NB). The objective of this study was to compare PAM and NB in quantitative terms. The NB simulation model assumed two tiers (nucleus, main) with unrestricted migration between the tiers. Clonal tests were used to predict breeding values and test resources per mate were kept constant for all mates. Both gain and diversity were combined into a single selection criterion, "group merit selection." Alternatives were compared over five breeding cycles by considering genetic gain and diversity in a selected PP established in a seed orchard. The assortment of mates in both alternatives enhanced additive variance and increased the additive effect in the BP, leading to additional gain in the PP. Gains generated under PAM always exceeded gains under NB. Thus, the main message from this study is that PAM in both the short- and long-term results in more gain at any target level of diversity in the PP (the breeder's target) than is achieved by the NB alternative. The optimum size of the nucleus varies with the desired level of seed orchard diversity. At lower target diversity, smaller nucleus sizes are favorable, while larger sizes result in more gain when seed orchard diversity is considered more important. PMID- 15278283 TI - Mapping of a locus for adult plant resistance to downy mildew in broccoli (Brassica oleracea convar. italica). AB - The identification of the gene Pp523, conferring downy mildew resistance to adult plants of broccoli (Brassica oleracea convar. italica), led to the construction of a genetic map that included this resistance locus, 301 amplified fragment length polymorphisms, 55 random amplified polymorphic DNAs, 46 inter-simple sequence repeats, three simple sequence repeats, four other PCR markers and a flower colour locus, all gathered into nine major linkage groups. Nineteen additional molecular markers were clustered into one group of four markers, one group of three markers and six pairs of markers. The map spans over 731.9 cM, corresponding to 89.5% of the 818 cM estimated to be the total genome length. A significant number of the mapped markers, 19.3%, showed distorted segregation. The average distance between mapped adjacent markers is 1.64 cM, which places this map among the densest published to date for this species. Using bulked segregant analysis, we identified a group of molecular markers flanking and closely linked in coupling to the resistance gene and included these in the map. Two markers linked in coupling, OPK17_980 and AT.CTA_133/134, are located at 3.1 cM and 3.6 cM, respectively, at each side from the resistance gene. These markers can be used for marker-assisted selection in breeding programs aiming at the introgression of this gene in susceptible B. oleracea genotypes. The fine mapping of the genomic region surrounding the Pp523 resistance gene is currently being carried out, a basic condition for its isolation via positional cloning. PMID- 15278284 TI - Identification of flanking SSR markers for a major rice gall midge resistance gene Gm1 and their validation. AB - Host-plant resistance is the preferred strategy for management of Asian rice gall midge (Orseolia oryzae), a serious pest in many rice-growing countries. The deployment of molecular markers linked to gall midge resistance genes in breeding programmes can accelerate the development of resistant cultivars. In the present study, we have tagged and mapped a dominant gall midge resistance gene, Gm1, from the Oryza sativa cv. W1263 on chromosome 9, using SSR markers. A progeny-tested F2 mapping population derived from the cross W1263/TN1 was used for analysis. To map the gene locus, initially a subset of the F2 mapping population consisting of 20 homozygous resistant and susceptible lines each was screened with 63 parental polymorphic SSR markers. The SSR markers RM316, RM444 and RM219, located on chromosome 9, are linked to Gm1 at genetic distances of 8.0, 4.9 and 5.9 cM, respectively, and flank the gene locus. Further, gene/marker order was also determined. The utility of the co-segregating SSR markers was tested in a backcross population derived from the cross Swarna/W1263//Swarna, and allelic profiles of these markers were analysed in a set of donor rice genotypes possessing Gm1 and in a few gall midge-susceptible, elite rice varieties. PMID- 15278288 TI - Expression studies of GUP1 and GUP2, genes involved in glycerol active transport in Saccharomyces cerevisiae, using semi-quantitative RT-PCR. AB - Glycerol active uptake in Saccharomyces cerevisiae, characterised physiologically as a proton symport, was previously described as repressed by glucose, induced by growth on non-fermentable carbon sources and unresponsive to growth under salt stress. GUP1 and GUP2 were identified and characterised as genes involved in glycerol active uptake. Using semi-quantitative RT-PCR, GUP1 and GUP2 transcription was measured. Unlike active transport activity determined previously, this was shown to be constitutive and not affected by either glucose repression or growth under salt stress. Furthermore, transcription of GUP1 and GUP2 was not affected in the gpd1gpd2 mutant strain grown under salt stress in the presence of small amounts of glycerol, in which case a very high Vmax of glycerol uptake was reported. Intracellular compounds were determined. Glycerol, acetate and trehalose were found to be the major compounds accumulated. Surprisingly, the gpd1gpd2 mutant was found to produce significant amounts of glycerol. Yet, the results provide no evidence for a correlation between the amount of each compound and the glycerol transport activity in any of the strains. PMID- 15278289 TI - Cloning, functional expression and characterization of three Phanerochaete chrysosporium endo-1,4-beta-xylanases. AB - Three Phanerochaete chrysosporium endo-1,4-beta-xylanase genes were cloned and expressed in Aspergillus niger. Two of these genes, xynA and xynC, encode family 10 glycoside hydrolases, while the third, xynB, codes for a family 11 glycoside hydrolase. All three xylanases possess a type I carbohydrate-binding domain connected to the catalytic domain by a linker region. The three xylanases were purified to homogeneity by weak anion or Avicell column chromatography and subsequently characterized. The XynA, XynB and XynC enzymes have molecular masses of 52, 30 and 50 kDa, respectively. Optimal activity was obtained at pH 4.5 and 70 degrees C with the family 10 xylanases and pH 4.5 and 60 degrees C with the family 11 xylanase. The measured Km when using birchwood xylan as the substrate was 3.71 +/- 0.69 mg/ml for XynA and XynC and was 9.96 +/- 1.45 mg/ml for XynB. Substrate specificity studies and the products released during the degradation of birchwood xylan suggest differences in catalytic properties between the two family 10 xylanases and the family 11 xylanase. PMID- 15278290 TI - [Bioptic diagnosis of chronic hepatitis. Results of an evidence-based consensus conference of the German Society of Pathology, of the German Society for Digestive and Metabolic Diseases and of Compensated Hepatitis (HepNet)]. PMID- 15278291 TI - Stability of a model of human granulopoiesis using continuous maturation. AB - A class of mathematical models involving a convection-reaction partial differential equation (PDE) is introduced with reference to recovering human granulopoiesis after high dose chemotherapy with stem cell support. The stability properties of the model are addressed by means of numerical investigations and analysis. A simplified model with proliferation rate and mobilization rate independent of maturity shows that the model is stable as the maturation rate grows without bounds, but may go through stable and non-stable regimens as the maturation rate varies. It is also shown that the system is stable when parameters are chosen to approximate a real physiological situation. System characteristics do not change profoundly by introduction of a maturity-dependent proliferation and mobilization rate, as is necessary to make the model operate more in accordance with hematological observations. However, by changing the system mitotic responsiveness with respect to changes in cytokine level, the system is still stable but may show persistent oscillations much resembling clinical observations of cyclic neutropenia. Furthermore, in these cases, changes in the model feedback signal caused by, for instance, an impaired effective cytokine elimination by cell receptors may enforce these oscillations markedly. PMID- 15278292 TI - A new interpretation of the Keller-Segel model based on multiphase modelling. AB - In this paper an alternative derivation and interpretation are presented of the classical Keller-Segel model of cell migration due to random motion and chemotaxis. A multiphase modelling approach is used to describe how a population of cells moves through a fluid containing a diffusible chemical to which the cells are attracted. The cells and fluid are viewed as distinct components of a two-phase mixture. The principles of mass and momentum balance are applied to each phase, and appropriate constitutive laws imposed to close the resulting equations. A key assumption here is that the stress in the cell phase is influenced by the concentration of the diffusible chemical. By restricting attention to one-dimensional cartesian geometry we show how the model reduces to a pair of nonlinear coupled partial differential equations for the cell density and the chemical concentration. These equations may be written in the form of the Patlak-Keller-Segel model, naturally including density-dependent nonlinearities in the cell motility coefficients. There is a direct relationship between the random motility and chemotaxis coefficients, both depending in an inter-related manner on the chemical concentration. We suggest that this may explain why many chemicals appear to stimulate both chemotactic and chemokinetic responses in cell populations. After specialising our model to describe slime mold we then show how the functional form of the chemical potential that drives cell locomotion influences the ability of the system to generate spatial patterns. The paper concludes with a summary of the key results and a discussion of avenues for future research. PMID- 15278296 TI - Monoclonal gammopathy of undetermined significance (MGUS) in patients with solid tumors: effects of chemotherapy on the monoclonal protein. AB - The aim of this study was to assess the effect of systemic chemotherapy on the monoclonal protein levels of patients with solid tumors who also have a monoclonal gammopathy of undetermined significance (MGUS). All patients with solid tumors who were referred to our department for consideration of systemic chemotherapy were evaluated with serum protein electrophoresis (SPEP) for the presence of MGUS. When MGUS was confirmed with immunofixation, serial SPEP was performed during and after completion of chemotherapy. Over a 6-year period, 21 patients with solid tumors and MGUS were prospectively identified and assessed. At least 50% reduction of serum monoclonal protein was noted in 4 of 11 patients treated with paclitaxel or docetaxel with a platinum analogue and in 5 of 7 patients who received an irinotecan-containing regimen. Our data indicate that in MGUS patients treated with irinotecan-containing chemotherapy regimens, a high incidence of reduction in their monoclonal protein levels is observed. Since topotecan, another topoisomerase I inhibitor, has some activity in multiple myeloma, further evaluation of irinotecan may be warranted. Evaluation of larger numbers of MGUS patients treated with chemotherapy for their underlying malignancy may help identify "in vivo" potentially active agents and regimens for patients with overt myeloma. PMID- 15278297 TI - Immunofluorescent analysis with the anti-PML monoclonal antibody PG-M3 for rapid and accurate genetic diagnosis of acute promyelocytic leukemia. AB - Genetic diagnosis is currently considered the most reliable method to accurately identify patients with acute promyelocytic leukemia (APL) requiring tailored therapy including all- trans retinoic acid (ATRA). We investigated the clinical effectiveness of immunofluorescence techniques with the anti-PML monoclonal antibody PG-M3 for rapid and accurate diagnosis of APL. PML immunofluorescence staining was analyzed in 164 patients with acute myeloblastic leukemia (AML), including APL (110 patients) and non-APL subtypes (54 patients). All 54 patients with an AML phenotype, in whom tests for t(15;17) or its fusion gene PML/ RARalpha were negative, showed a speckled (macrogranular) nuclear pattern. Of the 110 genetically diagnosed APL patients, 108 showed a microgranular pattern that confirmed PG-M3 positivity. The remaining two patients were not evaluable for PG M3 reactivity because of scarcity of cells. No patient with APL showed a normal pattern. The high sensitivity and specificity of immunolabeling using PG-M3 monoclonal antibody show that it is a highly efficient and reliable tool to identify PML/ RARalpha-positive patients with APL and that it should be standardized as a first-line diagnostic procedure. In addition, it is technically simple, fast, and cheap, only requiring small tissue samples and non sophisticated equipment. PMID- 15278298 TI - Anagrelide therapy in pregnancy: report of a case of essential thrombocythemia. PMID- 15278299 TI - Congenital dyserythropoietic anemias: epidemiology, clinical significance, and progress in understanding their pathogenesis. AB - The congenital dyserythropoietic anemias (CDAs) comprise a group of rare hereditary disorders of erythropoiesis, characterized by ineffective erythropoiesis as the predominant mechanism of anemia and by distinct morphological abnormalities of the majority of erythroblasts in the bone marrow. The classification in three types as proposed in 1968 is still valid, but there is genetic heterogeneity within each type, and there are additional variants of unknown genetic basis. CDA II is the most frequent, and the nonfamilial type of CDA III the rarest group. The genes of CDA II and CDA III were mapped to chromosome 20 and 15, respectively, and the gene of CDA I on 15q was recently cloned. Therapeutic decision making requires definition of the type, an estimate of individual severity, and presence of or risk for complications. Therapeutic measures include interferon-alpha for CDA I, splenectomy for CDA II, and iron depletion for all individuals at risk for secondary hemochromatosis. PMID- 15278300 TI - Comments about "Vascularised pedicled iliac crest graft for selected total hip acetabular reconstructions: a cadaver study". PMID- 15278306 TI - Analysis of gene expression profiles of hepatocellular carcinomas with regard to 18F-fluorodeoxyglucose uptake pattern on positron emission tomography. AB - PURPOSE: 18F-fluorodeoxyglucose (FDG) uptake on positron emission tomography (PET) scan has been found to reflect tumour aggressiveness and prognosis in various types of cancer. In this study, the gene expression profiles of hepatocellular carcinomas (HCCs) were evaluated to determine whether HCCs with high 18F-FDG uptake have more aggressive biological potential than those with low uptake. METHODS: Surgical specimens were obtained from ten patients with HCC (six males and four females, age range 38-68 years). The tumour samples were divided into two groups based on the 18F-FDG PET scan findings: high 18F-FDG uptake (n=4) and low 18F-FDG uptake (n=6). RESULTS: The pathological tumour grade was closely correlated with the 18F-FDG uptake pattern: HCCs with high 18F-FDG uptake were pathologically Edmondson-Steiner grade III, while those with low uptake were either grade II or grade II with a focal area of grade III. The total RNA was extracted from the frozen tissues of all HCCs (n=10) and adjacent non-cancerous tissue (n=7). The gene expression profiles were evaluated using an oligoDNA microarray. The HCCs with high 18F-FDG uptake showed increased expression of 11 genes--including vascular cell adhesion molecule-1, vinexin beta and core 1 UDP galactose:N-acetylgalactosamine-alpha-R-beta 1,3-galactosyltransferase and the natural killer cell inhibitory receptor--compared to those with low uptake (p<0.005). Nine genes, including regulator of mitotic spindle assembly 1, grb2 related adaptor protein and beta-1,3-n-acetylglucosaminyltransferase, were repressed. CONCLUSION: Gene expression is closely related to cell survival, cell to-cell adhesion or cell spreading; therefore, HCCs with high 18F-FDG uptake appear to have more aggressive biological properties than those with low uptake. PMID- 15278308 TI - Pre-operative localisation of hyperfunctional parathyroid tissue with 11C methionine PET. AB - PURPOSE: Previous studies have shown high sensitivity of positron emission tomography (PET) with 11C-methionine in the pre-operative localisation of parathyroid adenoma and hyperplasia. Nonetheless, in secondary and tertiary hyperparathyroidism (HPT) and in patients with recurrent disease, pre-operative localisation of adenomatous (PTA) or hyperplastic tissue is still a problem with all available methods. The aim of this study was to define the optimal imaging protocol and to compare the diagnostic value of 11C-methionine PET and 99mTc methoxyisobutylisonitrile (MIBI) single-photon emission computed tomography (SPECT): in particular, we wished to define the benefit of 11C-methionine in those patients with inconclusive or negative conventional imaging. METHODS: Thirty highly pre-selected patients with HPT were enrolled. Sixteen patients had primary HPT, 12 patients had secondary HPT, and two patients had recurrences of parathyroid carcinomas. All patients had ultrasound of the neck, dual-phase scintigraphy with 99mTc-MIBI and PET with 11C-methionine. SUV(parathyroid)/SUV(cervical soft tissue) (target-to-background) and SUV(parathyroid tissue)/SUV(thyroid tissue) (target-to-non-target) ratios were calculated. After surgery, histology of specimens was obtained in all patients but one. RESULTS: In 12 patients with secondary or tertiary HPT, 36 hyperplastic parathyroid glands were histologically verified. Twenty-five of 36 lesions (69%) were detected with 11C-methionine PET and 17 (47%) with 99mTc-MIBI scintigraphy. PET studies were positive in 17/18 (94%) cases in which HPT was related to adenomas or carcinomas. 99mTc-MIBI scintigraphy/SPECT yielded pathological lesions in 9/18 cases (50%). All eight atypical localisations of parathyroid glands were detected with PET but only six of the eight were detected with 99mTc MIBI scintigraphy/SPECT. In 10/11 patients with recurrent HPT and non-diagnostic scintigraphy/SPECT, hyperfunctional parathyroid tissue was identified with 11C methionine PET. The highest SUV(parathyroid)/SUV(cervical soft tissue) ratio was found 10 min, and the highest SUV(parathyroid tissue)/SUV(thyroid tissue) ratio 40 min post injection. In three patients clear delineation of hyperfunctional tissue was only achieved after 40 min post injection. CONCLUSION: 11C-methionine PET is a clinically useful method in highly pre-selected patients with recurrent primary HPT as well as in secondary and tertiary HPT if ultrasound and 99mTc-MIBI SPECT are inconclusive or negative. PET imaging of atypical PTA localisations is more accurate than conventional scintigraphy. In order to achieve optimal contrast of parathyroid glands versus thyroid tissue and adjacent soft tissue, imaging at both 10 min and 40 min is recommended. PMID- 15278309 TI - Characterization in Pseudomonas putida Cg1 of nahR and its role in bacterial survival in soil. AB - Sequencing, RFLP analyses and experiments utilizing a lacZ transcriptional reporter fused to the promoter regions of nahR and nahG in Pseudomonas putida Cg1 confirmed that regulation of naphthalene degradation in both P. putida Cg1 and the type strain, P. putida NCIB 9816-4, is consistent with that of NAH7 from P. putida G7. Two nahR knockout strains (RK1 and Cg1-NAHR from P. putida NCIB 9816-4 and Cg1, respectively) showed a growth defect in the presence of naphthalene as sole carbon and energy source. We hypothesized that nahR influences ecological fitness of bacteria in naphthalene-contaminated soil and tested this hypothesis using both parent and nahR-knockout strains introduced to soil microcosms with and without added naphthalene. After 21 days, loss of cell viability was pronounced in the presence of added naphthalene crystals for nahR mutants of both test bacteria, relative to the wild types. Diminished viable counts were attributed to toxicity. Thus, our data indicated that NahR in P. putida Cg1 is virtually identical to its homologues in other pseudomonads and that nahR is required for resistance to naphthalene toxicity, hence the persistence of bacterial cells in soil with high concentrations of naphthalene. PMID- 15278311 TI - Inhibiting mild steel corrosion from sulfate-reducing and iron-oxidizing bacteria using gramicidin-S-producing biofilms. AB - A gramicidin-S-producing Bacillus brevis 18-3 biofilm was shown to reduce corrosion rates of mild steel by inhibiting both the sulfate-reducing bacterium Desulfosporosinus orientis and the iron-oxidizing bacterium Leptothrix discophora SP-6. When L. discophora SP-6 was introduced along with D. orientis to a non antimicrobial-producing biofilm control, Paenibacillus polymyxa ATCC 10401, a corrosive synergy was created and mild steel coupons underwent more severe corrosion than when only D. orientis was present, showing a 2.3-fold increase via electrochemical impedance spectroscopy (EIS) and a 1.8-fold difference via mass loss measurements. However, when a gramicidin-S-producing, protective B. brevis 18-3 biofilm was established on mild steel, the metal coupons were protected against the simultaneous attack of D. orientis and L. discophora SP-6. EIS data showed that the protective B. brevis 18-3 biofilm decreased the corrosion rate about 20-fold compared with the non-gramicidin-producing P. polymyxa ATCC 10401 biofilm control. The mass loss for the protected mild steel coupons was also significantly lower than that for the unprotected ones (4-fold decrease). Scanning electron microscope images corroborated the corrosion inhibition by the gramicidin-S-producing B. brevis biofilm on mild steel by showing that the metal surface remained untarnished, i.e., the polishing grooves were still visible after exposure to the simultaneous attack of the sulfate-reducing bacterium and the iron-oxidizing bacterium. PMID- 15278313 TI - Deficiency in the glycerol channel Fps1p confers increased freeze tolerance to yeast cells: application of the fps1delta mutant to frozen dough technology. AB - Intracellular glycerol content affects the freeze-thaw stress tolerance of Saccharomyces cerevisiae. We have recently reported that intracellular-glycerol enriched cells cultured in glycerol medium acquire tolerance to freeze stress and retain high leavening ability even in dough after frozen storage [Izawa et al. (2004) Appl Microbiol Biotechnol http://dx.doi.org/10.1007/s00253-004-1624-4]. A deletion mutant of the FPS1 gene, which encodes a glycerol channel, accumulates glycerol inside the cell without an exogenous supply of glycerol into the medium. We found that the fps1delta cells acquired tolerance to freeze stress and retained high leavening ability in dough after frozen storage for 7 days. These results suggest that the fps1delta mutant is a useful strain for developing better frozen-dough with a commercial advantage. PMID- 15278317 TI - Microbial characterization of toluene-degrading denitrifying consortia obtained from terrestrial and marine ecosystems. AB - The degradation characteristics of toluene coupled to nitrate reduction were investigated in enrichment culture and the microbial communities of toluene degrading denitrifying consortia were characterized by denaturing gradient gel electrophoresis (DGGE) technique. Anaerobic nitrate-reducing bacteria were enriched from oil-contaminated soil samples collected from terrestrial (rice field) and marine (tidal flat) ecosystems. Enriched consortia degraded toluene in the presence of nitrate as a terminal electron acceptor. The degradation rate of toluene was affected by the initial substrate concentration and co-existence of other hydrocarbons. The types of toluene-degrading denitrifying consortia depended on the type of ecosystem. The clone RS-7 obtained from the enriched consortium of the rice field was most closely related to a toluene-degrading and denitrifying bacterium, Azoarcus denitrificians (A. tolulyticus sp. nov.). The clone TS-11 detected in the tidal flat enriched consortium was affiliated to Thauera sp. strain S2 (T. aminoaromatica sp. nov.) that was able to degrade toluene under denitrifying conditions. This indicates that environmental factors greatly influence microbial communities obtained from terrestrial (rice field) and marine (tidal flat) ecosystems. PMID- 15278319 TI - Skeletal tuberculosis in children. AB - The objective of this review is to present the imaging findings of skeletal tuberculosis in children. The incidence of tuberculosis is increasing and skeletal tuberculosis accounts for 10-20% of all extra-pulmonary cases. The most common manifestations of skeletal tuberculosis in children are spondylitis, arthritis and osteomyelitis. Tuberculous spondylitis involves the intervertebral disc only late in the disease. Subligamentous spread of the infection may lead to multiple levels of vertebral body involvement that may either be continuous or skipped. Extension of the disease into the paravertebral or extra-dural space may occur. Tuberculous arthritis usually occurs as a result of metaphyseal spread to the joint. Tuberculous osteomyelitis may appear as cystic, well-defined lesions, infiltrative lesions or spina ventosa. The latter is a term used to describe a form of tuberculous osteomyelitis where underlying bone destruction, overlying periosteal reaction and fusiform expansion of the bone results in cyst-like cavities with diaphyseal expansion. Radiographs are still the mainstay of evaluation of patients with bony lesions. Ultrasonography can detect soft-tissue extension of the bony lesions and guide drainage or biopsy procedures. CT accurately demonstrates bony sclerosis and destruction, especially in areas difficult to assess on radiographs such as the posterior elements of the vertebral body. MRI is the modality of choice in evaluating early marrow involvement and soft-tissue extension of the lesion. PMID- 15278321 TI - Fetal MRI: obstetrical and neurological perspectives. AB - Despite major advances in the understanding and in the genetics of several diseases of the developing brain, early prediction of the neurological prognosis of brain abnormality discovered in utero or of white matter damage discovered in a preterm neonate remains particularly difficult. Advances in prenatal diagnosis and the increased rate of survival of extremely preterm infants who are at higher risk of developing white matter damage underline the critical and urgent need for reliable predictive techniques. New imaging techniques such as diffusion-weighted imaging, magnetic resonance spectroscopy or functional MRI applied to the fetus represent promising tools in this perspective. PMID- 15278320 TI - Pediatric applications of abdominal vascular Doppler imaging: Part I. AB - Ultrasound is a remarkably powerful and versatile modality for pediatric imaging, without requiring exposure to radiation or sedatives. By providing information on blood flow, Doppler sonography can reveal details about normal physiology and disease processes not discernable from gray-scale anatomic images alone. However, in routine practice in many institutions Doppler remains underutilized, in part due to uncertainty of the meaning of changes in Doppler waveforms. In part I of this review, the basics of hemodynamics and how changes in blood flow affect the Doppler waveform are reviewed. Clinical applications in the investigation of hepatic disease are then discussed. PMID- 15278322 TI - Complications of pediatric paranasal sinusitis. AB - Acute paranasal sinus infection in children is often diagnosed clinically without the need for radiographic confirmation. Most cases have a favorable outcome following appropriate antibiotic therapy. A small percentage of cases where symptoms and signs are persistent or severe will require emergent imaging to rule out complications related to local spread of disease intraorbitally or intracranially. A strong index of suspicion is required in such cases, and cross sectional imaging evaluation with CT and MRI should include axial and coronal images of the paranasal sinuses and, where appropriate, the orbits and brain (with attention to the cavernous sinus). There is no role for plain radiography in the evaluation of the complications of acute sinusitis in the pediatric patient. PMID- 15278323 TI - Sialoblastoma: MRI findings. AB - Sialoblastoma is a rare, aggressive and potentially malignant salivary gland tumour diagnosed in the neonatal period. A total of 28 cases have been reported in the literature, but reports of the imaging findings are limited. We describe a neonate with a right parotid sialoblastoma. MRI showed a large facial mass, which was mostly hypointense to the brain on T1-weighted images and mildly hyperintense on T2-weighted images. There were foci of haemorrhage and necrosis. Heterogeneous and weak contrast enhancement was detected on contrast-enhanced images. The tumour invaded the maxilla and adjacent muscles. PMID- 15278324 TI - Long-term (6-year) follow-up of untreated multiple aggressive vertebral haemangiomas in an adolescent. PMID- 15278325 TI - Does the transition zone reliably delineate aganglionic bowel in Hirschsprung's disease? AB - BACKGROUND: Knowledge of the extent of aganglionic bowel is important for preoperative planning of trans-anal surgery in patients with Hirschsprung's disease (HD). OBJECTIVE: To evaluate the accuracy of the transition zone, as identified by contrast enema study, for identifying the extent of aganglionic bowel. MATERIALS AND METHODS: A total of 32 patients with preoperative contrast enema studies and pathologic identification of aganglionic extent were reviewed. Two pediatric radiologists independently reviewed the contrast enema studies. The radiographic transition zone was compared to the pathological extent of aganglionic bowel. RESULTS: Radiologist agreement of the site of transition zone on contrast enema was 90.6%. The concordance between the radiographic transition zone and pathologic extent of aganglionic bowel was 62.5%. The subgroup of patients with long-segment HD revealed a concordance of only 25%. CONCLUSION: Contrast enema delineation of the transition zone in HD needs to be regarded with caution. This is especially true in long-segment disease, where knowledge of the extent of aganglionic bowel is most crucial to surgical planning. PMID- 15278326 TI - Protective effect of aminoguanidine against nephrotoxicity induced by amikacin in rats. AB - Aminoglycoside antibiotics have long been used in antibacterial therapy. Despite their beneficial effects, aminoglycosides have considerable nephrotoxic and ototoxic side effects. It has been reported that reactive oxygen radical species (ROS) play role in the pathophysiology of aminoglycosides-induced nephrotoxicity. Aminoguanidine (AG) is an effective antioxidant and free radical scavenger which has long been known to protect against nephrotoxicity. We investigated the effects of AG on amikacin (AK)-induced changes of renal malondialdehyde (MDA), glutathione (GSH), blood urea nitrogen (BUN), serum creatinine (Cr) and albumin (Alb) which are used to monitor the development of renal tubular damage. Morphological changes in the kidney were also examined using light microscopy. A total of 21 rats were equally divided into three groups which were: (1) injected with saline, (2) injected with AK, and (3) injected with AK + AG, respectively. AK administration to control rats increased renal MDA and decreased GSH levels. AG administration before AK injection caused significant decreases in MDA and increases in GSH levels in kidneys compared to rats treated with AK alone. The serum BUN level increased slightly, Cr and serum Alb did not change as a result of any treatment. AG tended to decrease the level of serum BUN and did not cause any change in Alb or Cr levels. Morphological changes, including glomerular, tubular epithelial alterations and interstitial edema, were clearly observed in AK-treated rats. In addition, AG reversed the morphological damage to the kidney induced by AK. The results show that AG has a protective effect on nephrotoxicity induced by AK and may therefore improve the therapeutic index of AK. PMID- 15278327 TI - Safety profile of celecoxib as used in general practice in England: results of a prescription-event monitoring study. AB - AIMS: A post-marketing surveillance study using the technique of Prescription Event Monitoring was undertaken to monitor the safety of celecoxib, a cyclo oxygenase (COX)-2 inhibitor, as prescribed in primary care in England. METHODS: Patients were identified from dispensed British National Health Service prescription data supplied in confidence by the Prescription Pricing Authority for celecoxib between May and December 2000. Simple questionnaires were sent to the prescribing general practitioner at least 6 months after the date of the first dispensed prescription for each individual patient. Event incidence densities (IDs) [the number of 1st reports per 1000 patient-months of exposure (pme)] were calculated. ID differences for events reported in month 1 (ID1) and months 2-6 (ID2) were examined for temporal changes in event rate. Information on suspected adverse drug reactions (ADRs), reasons for stopping treatment, outcome of pregnancies and cause of death were also requested. Data were gathered on potential gastrointestinal (GI) risk factors [recent use of other non-steroidal anti-inflammatory drugs (NSAIDs), past history of upper GI disorders and concomitant gastro-irritant agents or anti-ulcer drugs]. Crude IDs per 1000 pme and ID ratios were calculated according to potential risk factors, and age (> or = 65 years, < or = 64 years). RESULTS: The cohort comprised of 17,458 patients [median age 62 years (IQR 51,73); 68.3% female]. The most common specified indication was osteoarthritis (28.1%, n = 4905). Not effective was the event with the highest ID1 (139.9 per 1000 pme). The clinical events with the highest ID1 were dyspepsia (25.4 per 1000 pme) followed by abdominal pain (10.6). These were also given frequently as reasons for stopping (551 and 174 of 9126 reports). Of 436 events in 325 patients (1.9% of total cohort) that were reported as ADRs, the most frequent were events within the alimentary system (186 reports). Uncommon events reported during treatment (not necessarily as ADRs) included allergy (0.10%, n = 17), anaphylaxis (0.01%, n = 2), angioneurotic oedema (0.02%, n = 3) and bronchospasm (0.05%, n = 9). There were 103 reports of events associated with thromboembolism and 111 reports of serious GI events [90 GI bleeds (upper and lower); 21 peptic ulcers] received during treatment or within 1 month of stopping. A past history of dyspeptic/other upper GI conditions and use of concomitant gastro-protective drugs were each associated with a significantly increased risk of dyspepsia and abdominal pain. CONCLUSIONS: Frequently reported adverse events were those GI events commonly associated with treatment with other NSAIDS. Stratification by identified risk factors suggested that channelling of high-risk patients is likely. Serious upper and lower GI events, and thromboembolic events did occur during this study, although the incidence was low (< 1%). Doctors should continue to prescribe NSAIDs, including COX-2-specific inhibitors, with caution. PMID- 15278329 TI - Modification of the human motor cortex by associative stimulation. AB - Manipulation of afferent input is capable of inducing reorganisation of the motor cortex. For example, following 1 h of paired electrical stimulation to the motor point of two hand muscles ("associative stimulation") the excitability of the corticospinal projection to the stimulated muscles is increased. Here we investigated the mechanisms responsible for such change using transcranial magnetic stimulation (TMS). Cortical excitability changes were investigated by measuring motor evoked potentials (MEPs), intracortical inhibition (ICI), intracortical facilitation (ICF), and short-interval intracortical facilitation (SICF). Following 1 h of associative stimulation MEP amplitudes in the stimulated muscles significantly increased. Additionally, there was a significant increase in ICF and of SICF at interstimulus intervals in the range of 2.3-3.3 ms. There was no significant change in ICI. These findings confirm previous observations that a 1-h period of associative stimulation can increase the excitability of the cortical projection to stimulated muscles. Additionally, these results suggest that the observed modifications of excitability are due to changes in intracortical excitatory circuits. PMID- 15278330 TI - Can the motor system resolve a premovement bias in grip aperture? Online analysis of grasping the Muller-Lyer illusion. AB - The goal of the present investigation was to determine the time-course by which the motor system might resolve the context-dependent effects of a visual illusion [i.e., the Muller-Lyer (ML) figure]. Specifically, we asked participants to scale their grip aperture (GA) to the perceived size of an object embedded within a ML figure in advance of closed-loop (CL) and open-loop (OL) grasping movements. As a result, premovement GA was biased in a direction consistent with the perceptual effects of the illusion. We reasoned that such a manipulation might provide a novel opportunity to determine whether the motor system is able to resolve a biased GA immediately following the onset of a response [i.e., in accord with the perception/action model (PAM); Milner and Goodale 1995, The visual brain in action, Oxford University Press], or gradually as the action unfolds [i.e., in accord with the planning/control model (PCM); Glover and Dixon 2002, Percept Psychophys 64:266-278]. It was found that biasing GA in advance of movement resulted in a reliable effect of the ML figure throughout CL and OL trials (i.e., up to 80% of grasping time). Although the present findings appear contrary to the theoretical tenets of the PAM and the PCM, it is proposed that biasing GA in advance of movement leads to offline visual processing and a feedforward mode of grasping control, thus accounting for the illusion-induced effect throughout the grasping response. PMID- 15278331 TI - Optical microarray biosensors. PMID- 15278332 TI - Simultaneous determination of the Cd and Zn total body burden of individual, nearly microscopic, nanoliter-volume aquatic organisms ( Hyalella azteca) by rhenium-cup in-torch vaporization (ITV) sample introduction and axially viewed ICP-AES. AB - The Cd and Zn total body burden of individual, up to 7-day-old aquatic organisms (Hyalella azteca benthic amphipod) with an average volume of approximately 100 nL was determined simultaneously by using rhenium-cup (Re-cup) in-torch vaporization (ITV) sample introduction and an axially viewed inductively coupled plasma atomic emission spectrometry (ICP-AES) system. The direct elemental analysis capabilities of this system (i.e., no sample digestion) reduced sample preparation time, eliminated contamination concerns from the digestion reagent and, owing to its detection limits (e.g., in the low pg range for Cd and Zn), vit enabled simultaneous determinations of Cd and Zn in individual, neonate and young juvenile specimens barely visible to the unaided eye (e.g., nearly microscopic). As for calibration, liquid standards and the standard additions method were tested. Both methods gave comparable results, thus indicating that in this case liquid standards can be employed for calibration, and in the process making use of the standard additions method unnecessary. Overall, the ITV-ICP-AES approach by-passed the time-consuming acid digestions, eliminated the potential for contamination from the digestion reagents, improved considerably the speed of acquisition of analytical information and enabled simultaneous determinations of two elements using individual biological specimens. PMID- 15278333 TI - Chromatographic molecular recognition for catechol-related compounds using thiacalix[4]arene as an effective selector. AB - Thiacalix[4]arene (5,11,17,23-tetra- tert-butyl-25,26,27,28-tetrahydroxy 2,8,14,20-tetrathiacalix[4]arene) is an amphiphilic molecule comprising four p tert-butylphenol-like groups ortho-linked by single sulfur atoms. This molecule has a high electron density area owing to the close proximity of the hydroxyl groups and sulfur atoms. We studied the applicability of this interesting compound as a selector for high-performance liquid chromatography (HPLC) thereby presumably exploiting this feature. Firstly, uniformly sized polymer particles were prepared by using a multi-step swelling and polymerization method with ethylene glycol dimethacrylate (EDMA) as a cross-linker. Methacrylic acid (MAA) was introduced onto the surface of the resulting polymer particles through a new modification method. Thiacalix[4]arene was chemically bonded through the MAA group by using 1,4-dibromobutane as a spacer to reduce steric hindrance around the MAA and the polymer particle itself. The performance of the prepared polymer based thiacalix[4]arene-modified stationary phase was evaluated with HPLC. Specific chromatographic retention behavior was observed for catechol relative to positional isomers of xylene, cresol, and benzene-diol. Catecholamine and catechol showed specific chromatographic retention behavior. PMID- 15278335 TI - Ultratrace voltammetric determination of DNA-bound platinum in patients after administration of oxaliplatin. AB - Oxaliplatin, a novel diaminocyclohexane-platinum complex, is used for the treatment of metastatic colorectal cancer. The amount of DNA-adduct formation of this drug in white blood cells of patients is determined after isolation of the DNA by density gradient centrifugation and a four-step solid phase extraction procedure. DNA is quantified by UV spectrometry, and platinum is determined after mineralization of the DNA sample by adsorptive stripping voltammetry (formazone method). It is possible to determine Pt-nucleotide ratios in clinical samples down to five Pt atoms in 10(8) nucleotides, and the dynamic range of the method covers three orders of magnitude. An absolute amount of 25 microg of DNA is sufficient for such measurements. With the method described, the time-dependent formation of oxaliplatin DNA adducts can be monitored in clinical studies, which may help us to understand inter-individual differences in the responses of patients to oxaliplatin-based therapy. PMID- 15278336 TI - Total protein determinations by particle beam/hollow cathode optical emission spectroscopy (PB/HC-OES) system III: investigation of carrier salts for enhanced particle transport. AB - Particle beam hollow cathode optical emission spectroscopy (PB/HC-OES) is evaluated as a generic tool for total protein determinations by monitoring the carbon atomic emission (C (I) 193.0 nm) resultant from dissociated analyte species. Previous studies demonstrated the capability of the PB/HC-OES system for total protein determinations with limits of detection for bovine serum albumin (BSA) samples being at the single-nanogram level for 200 microl injections. Non linear behavior across the concentration range in the calibration curve was observed due to the poor transport of small particles (owing to low analyte concentrations) through the PB interface. The potential use of non-volatile salts as carrier agents is investigated in the determination of protein samples by PB/HC-OES. A range of chloride salts (different cations), potassium salts (different anions), and an organic modifier (ammonium acetate) is investigated here for possible use as carriers upon addition as sample injection matrices for protein samples. The analyte response curves of BSA samples with KCl added as the sample injection matrix show higher sensitivity, better linearity (R2) and subsequently lower detection limits in comparison to those obtained with water, HCl, KNO3 or ammonium acetate as carrier matrices. PMID- 15278337 TI - Trace analysis of rapamycin in human blood by micellar electrokinetic chromatography. AB - A capillary electrophoretic method with UV detection at 278 nm has been developed for analysis of the immunosuppressant rapamycin (sirolimus) in human blood at low microgram per liter levels. Separation has been achieved in an acidic carrier electrolyte containing sodium dodecylsulfate and 30% (v/v) acetonitrile. For sample clean-up and preconcentration, an off-line solid-phase extraction step using a silica-based reversed-phase material and an on-capillary focussing technique were employed. The latter allows the injection of increased sample volumes without excessive band broadening. Although this new method is less sensitive than existing liquid chromatographic procedures combined with mass spectrometry, it is fully suited to routine analysis of rapamycin in blood from patients treated with this drug. Last but not least the low costs make it an attractive alternative to established methods. PMID- 15278339 TI - Application of SEC-ICP-MS for comparative analyses of metal-containing species in cancerous and healthy human thyroid samples. AB - Size exclusion chromatography (SEC) was coupled on-line to inductively coupled plasma mass spectrometry (ICP-MS) for speciation study of trace metals in cancerous thyroid tissues in comparison to healthy thyroids aimed to estimation of changes in metalloprotein speciation in pathological tissue. The study showed a presence of species binding Cu, Zn, Cd and Pb in healthy thyroid tissue with a good reproducibility of chromatographic results, whereas the same species could not be detected in cancerous tissues. Thus, remarkable differences with respect to metal-binding species were revealed between healthy and pathological thyroid samples, pointing out a completely different distribution of trace metals in cancerous tissues. The metal-binding species could not be identified in the frame of this work because of a lack of appropriate standards. Nevertheless, the results obtained confirm the suitability of SEC-ICP-MS for monitoring of changes in trace metal distribution in cancerous tissue and will help to better understand the role of metal-containing species in thyroid pathology. PMID- 15278340 TI - Application of luminescent nanocrystals as labels for biological molecules. AB - Luminescent semiconductor nanocrystals, so called "quantum dots" (QD), have attracted increasing interest for bioanalytical labeling applications in recent years. This review describes the major optical and (bio)chemical features of this class of label, compared with organic dyes. Different conjugation methods are also discussed and the most important recent applications are presented. An overview over the current state-of-the-art is given, as also is an outlook on possibilities and limitations. PMID- 15278341 TI - Raman microscopic investigations of BaTiO(3) precursors with core-shell structure. AB - Due to their outstanding dielectric and ferroelectric properties, barium titanate (BaTiO(3))-based ceramics have found many applications in electronic devices. To optimise the final quality of such ceramics, a detailed knowledge of the complex processes involved in the formation of BaTiO(3) is required. The phase formation process in ordered structures of the BaCO(3)/TiO(2) system was analysed by X-ray diffraction and by Raman spectral imaging (RSI) as a function of the annealing temperature. RSI was used for the first time as a locally resolving method for phase analysis, and proved to be a useful tool in examining the formation process of BaTiO(3) starting from spherical, core-shell structured precursors of the type TiO(2) core/BaCO(3) shell. The Raman spectra of different BaO-TiO(2) phases appearing as intermediate phases during the formation of BaTiO(3) were recorded for separately-prepared pure substances. Using these spectra as fingerprints, and choosing phase filters by setting wave number windows, "phase landscape pictures" of the samples at different temperatures during the genesis of BaTiO(3) could be created with a lateral resolution of up to 200 nm. These pictures confirm shell like formation of the different barium titanate phases according to the diffusion of barium and oxygen ions from the Ba-rich shell into the TiO(2) core. At an intermediate state of the phase formation process, the phase sequence Ba(2)TiO(4), BaTiO(3), BaTi(2)O(5), BaTi(4)O(9) and BaTi(5)O(11) to TiO(2) was detected from the outer to the inner parts of the core-shell structures. PMID- 15278342 TI - High-throughput chemiluminometric determination of prostate-specific membrane antigen mRNA in peripheral blood by RT-PCR using a synthetic RNA internal standard. AB - A quantitative reverse transcriptase polymerase chain reaction (RT-PCR) method, employing internal standard (IS) RNA and a simplified chemiluminometric hybridization assay, is described for the determination of prostate-specific membrane antigen (PSMA) mRNA. The recombinant RNA IS has the same binding sites and size as the amplified PSMA mRNA. Biotinylated PCR products (263 bp) from PSMA mRNA and RNA IS are captured in microtiter wells coated with streptavidin, and hybridized with alkaline phosphatase-conjugated probes. The bound alkaline phosphatase (AP) is measured by using a chemiluminogenic substrate. The ratio of the luminescence values obtained for PSMA mRNA and the RNA IS is a linear function of the initial amount of PSMA mRNA present in the sample before RT-PCR. The linear range extends from 500 to 5,000,000 PSMA mRNA copies and the overall reproducibility of the assay, including RT-PCR and hybridization, ranges from 7.4 to 16.6%. Samples containing total RNA from PSMA-expressing LNCaP cells give luminescence ratios linearly related to the number of cells in the range 0.5 5,000 cells. PMID- 15278344 TI - Simultaneous determination of zinc, copper, lead, and cadmium in fuel ethanol by anodic stripping voltammetry using a glassy carbon-mercury-film electrode. AB - A new, versatile, and simple method for quantitative analysis of zinc, copper, lead, and cadmium in fuel ethanol by anodic stripping voltammetry is described. These metals can be quantified by direct dissolution of fuel ethanol in water and subsequent voltammetric measurement after the accumulation step. A maximum limit of 20% ( v/ v) ethanol in water solution was obtained for voltammetric measurements without loss of sensitivity for metal species. Chemical and operational optimum conditions were analyzed in this study; the values obtained were pH 2.9, a 4.7-microm thickness mercury film, a 1,000-rpm rotation frequency of the working electrode, and a 600-s pre-concentration time. Voltammetric measurements were obtained using linear scan (LSV), differential pulse (DPV), and square wave (SWV) modes and detection limits were in the range 10(-9)-10(-8) mol L(-1) for these metal species. The proposed method was compared with a traditional analytical technique, flame atomic absorption spectrometry (FAAS), for quantification of these metal species in commercial fuel ethanol samples. PMID- 15278345 TI - Dose and schedule determinants of cocaine choice under concurrent variable interval schedules in rhesus monkeys. AB - RATIONALE: Drug abuse can be characterized as a condition in which the choice to self-administer a drug is excessive, even exclusive of the choice of other reinforcers. Under concurrent interval schedules of reinforcement, subjects typically distribute behavior to match reinforcement allocation. However, research has shown that when behavior is maintained by different doses of cocaine under concurrent variable-interval (conc VI) schedules, exclusive choice of the higher dose is the rule. OBJECTIVE: The present study was designed to examine the generality of this finding to other behavioral conditions. METHODS: Rhesus monkeys ( n=5) lever pressed under a conc VI 60-s VI 60-s or a conc VI 600-s VI 600-s schedule of cocaine (i.v.) presentation. Doses differing by 4-fold (0.025 versus 0.1, 0.05 versus 0.2 mg/kg per injection) were available for lever pressing. RESULTS: Monkeys responded more on the lever associated with the higher dose when saline or a lower dose was the alternative. The distribution of responses was well predicted by relative drug intake, but with consistent undermatching. Exclusive high-dose responding was seen in about half of the individual session intervals under the shorter schedule, rarely under the longer schedule, and was not seen over the session. CONCLUSION: Under conc VI schedules, behavior was apportioned between two different doses in a manner that favored the higher dose but undermatched relative intake. Exclusive high-dose choice may occur when cocaine is frequently available but is not an invariable outcome of the choice between a low and a high dose of cocaine. PMID- 15278346 TI - Benzodiazepine receptor and serotonin 2A receptor modulate the aversive-like effects of nitric oxide in the dorsolateral periaqueductal gray of rats. AB - RATIONALE: Escape reactions induced by electrical stimulation of the dorsolateral periaqueductal gray (dlPAG) are inhibited by local administration of benzodiazepine (BZ) or serotonin (5-HT) receptor agonists. Nitric oxide (NO) is a gas messenger that may mediate aversive behaviors. NO donors injected into the dlPAG induce escape reactions. OBJECTIVES: To test the hypothesis that the escape reactions induced by a NO donor in the dlPAG would be attenuated by pre-treatment with BZ-receptor or 5-HT-receptor agonists. METHODS: Male Wistar rats with cannulae aimed at the dlPAG received microinjections of vehicle (0.2 microl), the BZ midazolam maleate (80 nmol), the 5-HT(1A)-receptor agonist 8-OH-DPAT (8 nmol or 16 nmol) or the 5-HT(2A/2C)-receptor agonist DOI (16 nmol) 10 min before the administration of the NO donor SIN-1 (150 nmol). Behavioral observation took place immediately after the last injection in an open arena over a 10-min period. RESULTS: SIN-1 induced escape reactions characterized by running and jumps. Pre treatment with DOI, but not 8-OH-DPAT, partially inhibited the effects of SIN-1. Pre-treatment with midazolam maleate, however, completely prevented the effects of the NO donor. CONCLUSION: The results suggest that the aversive-like effects of NO donor in the dlPAG may be modulated by the BZ and 5-HT(2A/2C) receptors. PMID- 15278352 TI - Cardiac output and oxygen release during very high-intensity exercise performed until exhaustion. AB - Our objectives were firstly, to study the patterns of the cardiac output (Q(.)) and the arteriovenous oxygen difference [(a-nu(-))O(2)] responses to oxygen uptake (V(.)O(2)) during constant workload exercise (CWE) performed above the respiratory compensation point (RCP), and secondly, to establish the relationships between their kinetics and the time to exhaustion. Nine subjects performed two tests: a maximal incremental exercise test (IET) to determine the maximal V(.)O(2) (V(.)O(2)peak), and a CWE test to exhaustion, performed at p Delta50 (intermediate power between RCP and V(.)O(2)peak). During CWE, V(.)O(2) was measured breath-by-breath, Q(.) was measured beat-by-beat with an impedance device, and blood lactate (LA) was sampled each minute. To calculate ( a-nu( )O(2), the values of V(.)O(2) and Q(.) were synchronised over 10 s intervals. A fitting method was used to describe the V(.)O(2), Q(.) and ( a-nu(-))O(2) kinetics. The ( a-nu(-)O(2) difference followed a rapid monoexponential function, whereas both V(.)O(2) and Q(.) were best fitted by a single exponential plus linear increase: the time constant (tau) V(.)O(2) [57 (20 s)] was similar to tau ( a-nu(-)O(2), whereas tau for Q(.) was significantly higher [89 (34) s, P <0.05] (values expressed as the mean and standard error). LA started to increase after 2 min CWE then increased rapidly, reaching a similar maximal value as that seen during the IET. During CWE, the rapid component of V(.)O(2) uptake was determined by a rapid and maximal ( a-nu(-)O(2) extraction coupled with a two-fold longer Q(.) increase. It is likely that lactic acidosis markedly increased oxygen availability, which when associated with the slow linear increase of Q(.), may account for the V(.)O(2) slow component. Time to exhaustion was larger in individuals with shorter time delay for ( a-nu(-)O(2) and a greater tau for Q(.). PMID- 15278353 TI - Decreased lung capillary blood volume post-exercise is compensated by increased membrane diffusing capacity. AB - The diffusing capacity of the lung for carbon monoxide (DLCO) decreases to below the pre-exercise value in the hours following a bout of intense exercise. Two mechanisms have been proposed: (1) development of pulmonary oedema and (2) redistribution of central blood volume to peripheral muscles causing a reduction in pulmonary capillary blood volume ( V(c)). In the present study DLCO, V(c) and the membrane diffusing capacity ( D(m)) were measured in nine healthy females using a rebreathing method, in contrast to the single breath technique employed in previous studies. DLCO, V(c) and D(m) were measured before and at 1, 2, 3, 16 and 24 h following maximal treadmill exercise. Compared with pre-exercise values, DLCO was depressed by up to 8.9 (3.0)% ( P<0.05) for the first 3 h following exercise, but had returned to pre-exercise values by 16 h post-exercise. V(c) fell by 21.2 (4.1)% ( P<0.05) at 3 h post-exercise, but at the same time D(m) increased by 14.7 (9.1)%. It was concluded that: (1) the increase in D(m) made it unlikely that the fall in DLCO was due to interstitial oedema and injury to the blood gas barrier; (2) on the other hand, the reduction in DLCO following exercise was consistent with a redistribution of blood away from the lungs; and (3) the trend for D(m) and V(c) to reciprocate one another indicates a situation in which a fall in V(c) nevertheless promotes gas transfer at the respiratory membrane. It is suggested that this effect is brought about by the reorientation of red blood cells within the pulmonary capillaries following exercise. PMID- 15278355 TI - Human adductor muscles atrophy after short duration of unweighting. AB - The purpose of this study was to investigate the effect of 20 days of bed rest (BR) on the physiological cross-sectional area (PCSA) of the adductor (AD) muscle group. Five healthy men and five healthy women were kept on a horizontal bed for 20 days. To calculate the PCSA of the AD muscle group, transaxial magnetic resonance imaging of the right thigh was acquired four times for each of the subjects: (1) pre BR, (2) on the tenth day of BR, (3) post BR, and (4) 1 month after BR. Overall, the PCSA measurements of the AD muscle group and its individual muscles were significantly decreased on the tenth day of BR and at post BR; however, they had recovered to the baseline within 1 month of the reambulation period. The amount and pattern of relative change in the PCSA measurements of the AD muscle group were similar to those of the knee extensor (KE) and knee flexor (KF) muscle groups studied previously. These results suggested that the AD muscle group showed an atrophic response that was similar to the KE and KF muscle groups after unweighting; therefore, we should pay much more attention to the AD muscle group when considering countermeasures for future human spaceflight projects. PMID- 15278354 TI - A wide range of baroreflex stimulation does not alter forearm blood flow. AB - The contribution to the regulation of forearm blood flow (FBF) by different baroreceptor populations has previously only been studied over a limited range of stimuli. Therefore, FBF and R-R interval were recorded during neck suctions and neck pressures ranging from -60 to +40 mmHg. The change in R-R interval (DeltaR R) during neck suction was significantly increased at each stage when compared to the control ( P<0.05). DeltaR-R did not show any significant change during any of the neck pressure stages ( P>0.05). Suction or pressure applied to the neck did not elicit any significant changes in FBF when compared to the control ( P>0.05). These data show that widening the range of applied stimuli to carotid sinus baroreceptors does not induce a change in FBF. However, the small transient changes reported previously cannot be discounted. PMID- 15278357 TI - Validity of heart rate as an indicator of aerobic demand during soccer activities in amateur soccer players. AB - In order to validate the use of heart rate (HR) in describing and monitoring physiological demands during soccer activities, the HR versus oxygen uptake ( V(.)O(2)) relationship determined on the field during soccer-specific exercises was compared to that found in the laboratory during treadmill exercise. Seven male amateur soccer players [mean (SE), age 25.3 (1.2) years, body mass 72.9 (2.1) kg, stature 1.76 (0.03) m] performed three trials on the field (two laps of a purpose-made circuit including a variety of soccer activities) at different intensities (moderate, high and very high, according to their rate of perceived exertion) and an incremental test on a treadmill in the laboratory. HR increased linearly with V(.)O(2) during both field and laboratory tests according to exercise intensity ( P<0.01). The mean correlation coefficients of the HR- V(.)O(2) relationships obtained in the laboratory and on the field were 0.984 (0.012) and 0.991 (0.005) ( P<0.001), respectively. The mean value of the HR- V(.)O(2) regression equation slope and intercept obtained in laboratory [0.030 (0.002) and 79.6 (4.6), respectively] were not significantly different compared to those found on the field [0.032 (0.003) and 76.7 (9.7)]. The present study seems to confirm that HR measured during soccer exercises effectively reflects the metabolic expenditure of this activity. Thus, with the aid of laboratory reference tests, the physiological demands of soccer activities can be correctly estimated from HR measured on the field in amateur soccer players. PMID- 15278359 TI - Expression of the cholinergic gene locus in the rat placenta. AB - High amounts of acetylcholine (ACh) and its synthesising enzyme choline acetyltransferase (ChAT) have been detected in the placenta. Since the placenta is not innervated by extrinsic or intrinsic cholinergic neurons, placental ACh and ChAT originate from non-neuronal sources. In neurons, cytoplasmic ACh is imported into synaptic vesicles by the vesicular acetylcholine transporter (VAChT), and released through vesicular exocytosis. In view of the coordinate expression of VAChT and ChAT from the "cholinergic gene locus" in neurons, we asked whether VAChT is coexpressed with ChAT in rat placenta, and investigated this issue by means of RT-PCR, in situ hybridisation, western blot and immunohistochemistry. Messenger RNA and protein of the common type of ChAT (cChAT), its splice variant peripheral ChAT (pChAT), and VAChT were detected in rat placenta with RT-PCR and western blot. ChAT in situ hybridisation signal and immunoreactivity for cChAT and pChAT were observed in nearly all placental cell types, while VAChT mRNA and immunolabelling were detected in the trophoblast, mesenchymal cells and the visceral yolk sac epithelial cells. While ChAT is nearly ubiquitously expressed in rat placenta, VAChT immunoreactivity is localised cell type specifically, implying that both vesicular and non-vesicular ACh release machineries prevail in placental cell types. PMID- 15278363 TI - Inhibition of melanoma metastases by fenofibrate. AB - The effect of fenofibrate, a ligand of peroxisome proliferator-activated receptor (PPAR) alpha, on the growth and metastatic potential of Bomirski hamster melanoma s.c. tumors, pigmented line (BHM Ma) was investigated in vivo. RT-PCR and Western blot analyses revealed the presence of mRNA and protein of PPAR alpha in BHM Ma cells. The animals treated orally with fenofibrate developed significantly fewer metastatic foci in the lungs, as compared to the control group; however, primary tumor growth remained unaltered. This observation is interesting in respect of the potential use of fenofibrate in melanoma chemoprevention. PMID- 15278364 TI - Molecular interactions of B-CAM (basal-cell adhesion molecule) and laminin in epithelial skin cancer. AB - Molecular events underlying the progression of malignant tumors through the surrounding tissue are largely mediated by membrane-bound adhesion molecules. Basal-cell adhesion molecule (B-CAM), a 90-kDa laminin receptor of the immunoglobulin superfamily, is induced in some epithelial malignancies. Its function in these tumors, however, still remains obscure. We demonstrated that expression of B-CAM is very weak, if detectable at all, in normal epidermis but is strongly induced in both basal cell carcinomas and squamous cell carcinomas of the skin, and most pronounced at the basal surface of the tumor nests. Interestingly, the only known B-CAM ligand, laminin, was markedly upregulated within corresponding microanatomical sites surrounding the tumor nests, suggesting that both molecules may interact there. Consistent with this hypothesis, we were able to directly demonstrate binding of a B-CAM/Fc chimeric molecule to the peritumoral stroma in situ. Finally, in proof-of-principle experiments, human B-CAM was overexpressed both in murine and in human fibroblasts. The haptotactic migration of these novel B-CAM+ cell populations on a laminin matrix was significantly increased (P = 0.02) as compared to mock transfected cells when integrin-mediated adhesion was blocked by chelation of divalent cations. Thus, our findings provide the first direct experimental evidence that interactions of B-CAM and laminin may be involved in progression of epithelial skin tumors. PMID- 15278365 TI - Cyclic AMP differentially regulates cell proliferation of normal human keratinocytes through ERK activation depending on the expression pattern of B Raf. AB - Intracellular cyclic AMP (cAMP) increased by extracellular stimuli induces various biological effects, such as cell proliferation, differentiation, and migration. Previous reports regarding the effect of cAMP on keratinocyte proliferation are contradictory and indicate that the effect apparently depends on cellular density. Recent studies have revealed that cAMP signaling regulates cell proliferation by modulating mitogen-activated protein kinase (MAPK) activity. The precise mechanism by which cAMP affects keratinocyte proliferation and/or the crosstalk between the cAMP and MAPK signaling pathways, however, remain to be determined. Using normal human keratinocytes (NHK), we investigated the effect of cAMP on keratinocyte proliferation and its molecular mechanism in terms of cellular density. In confluent NHK, cyclic AMP decreased extracellular regulated kinase (ERK) phosphorylation and cell proliferation in a Ras independent and Rap1-dependent manner. The decreased cell proliferation by cAMP was blocked by the MEK-1 inhibitor, PD98059. In contrast, in subconfluent NHK, cAMP increased ERK phosphorylation and cell proliferation. Western blot analysis revealed that NHK expressed B-Raf and Rap-1. Although both 95 kDa and 62 kDa B Raf isoforms were expressed in subconfluent NHK, only 62 kDa B-Raf was detected in confluent NHK. Transfection of 95 kDa B-Raf into confluent NHK resulted in a cAMP-dependent increase in ERK phosphorylation and cell proliferation. These findings indicate that differential expression of B-Raf isoforms is critical for cAMP-dependent regulation of NHK proliferation that depends on phosphorylation of ERK. PMID- 15278366 TI - Activation of NFkappaB signal pathways in keloid fibroblasts. AB - Keloids are characterized as an "over-exuberant" healing response resulting in a disproportionate extracellular matrix (ECM) accumulation and tissue fibrosis. In view of the integral role of inflammation and cytokines in the healing response, it is logical to assume that they may play a part in orchestrating the pathology of this "abnormal" healing process. Tumor necrosis factor-alpha (TNF-alpha) is a potent proinflammatory cytokine involved in activation of signaling events and transcriptional programs, such as NFkappaB. This study attempts to determine the difference in NFkappaB and its related genes expression and DNA binding activity between keloid and normal skin fibroblasts. Three keloid and normal skin tissues (NSk) and their derived fibroblasts were used to determine NFkappaB signaling pathway expression using specific cDNA microarrays, Western blot analysis and immunohistochemistry. Electrophoretic mobility gel shift assay (EMSA) was used to assess NFkappaB-binding activity, all assays were performed in the presence and absence of TNF-alpha. TNF-alpha up-regulated 15% of NFkappaB signal pathway related genes in keloid fibroblast compared to normal skin. At the protein level, keloid fibroblasts and tissues showed higher basal levels of TNF- receptor associated factors-TRAF1, TRAF2-TNF-alpha, inhibitor of apoptosis (c-IAP-1), and NFkappaB, compared with NSk. Keloid fibroblasts showed a constitutive increase in NFkappaB-binding activity in comparison to NSk both with and without TNF-alpha treatment. NFkappaB and its targeted genes, especially the antiapoptotic genes, could play a role in keloid pathogenesis; targeting NFkappaB could help in developing therapeutic interventions for the treatment of keloid scarring. PMID- 15278367 TI - Catecholamine effects on human melanoma cells evoked by alpha1-adrenoceptors. AB - The biological effects of catecholamines in mammalian pigment cells are poorly understood. Our previous results showed the presence of alpha(1)-adrenoceptors in SK-Mel 23 human melanoma cells. The aims of this work were to (1) characterize catecholamine effects on proliferation, tyrosinase activity and expression, (2) identify the alpha(1)-adrenoceptor subtypes, and (3) verify whether chronic norepinephrine (NE) treatment modified the types and/or pharmacological characteristics of adrenoceptors present in SK-Mel 23 human melanoma cells. Cells treated with the alpha(1)-adrenergic agonist, phenylephrine (PHE, 10(-5) or 10( 4) M), for 24-72 h, exhibited decreased cell proliferation and enhanced tyrosinase activity, but unaltered tyrosinase expression as compared with the control. The proliferation and tyrosinase activity responses were inhibited by the alpha(1)-adrenergic antagonist prazosin, suggesting they were evoked by alpha(1)-adrenoceptors. The presence of actinomycin D, a transcription inhibitor, did not diminish PHE-induced effects. RT-PCR assays, followed by cloning and sequencing, demonstrated the presence of alpha(1A)- and alpha(1B)-adrenoceptor subtypes. NE-treated cells (24 or 72 h) were used in competition assays, and showed no significant change in the competition curves of alpha(1)-adrenoceptors as compared with control curves. Other adrenoceptor subtypes were not identified in these cells, and NE pretreatment did not induce their expression. In conclusion, the activation of SK-Mel 23 human melanoma alpha(1)-adrenoceptors elicit biological effects, such as proliferation decrease and tyrosinase activity increase. Desensitization or expression of other adrenoceptor subtypes after chronic NE treatment were not observed. PMID- 15278368 TI - Complete resolution of psoriasis in a patient treated with stealth liposomal doxorubicin and carboplatin for ovarian cancer. PMID- 15278369 TI - Intramedullary k-wire fixation of metacarpal fractures. AB - INTRODUCTION: The majority of metacarpal fractures can be treated conservatively. Nevertheless, surgical treatment is justified in certain cases. Palmar dislocation of >30 degrees and shortening of >5 mm will significantly affect extension and flexion of the hand. Consequently, surgical treatment is indicated. The aim of our study was to evaluate the clinical results of intramedullary Kirschner-wire fixation of metacarpal fractures. MATERIAL AND METHODS: In a retrospective study we analyzed the clinical results of 35 patients with metacarpal fractures that had been treated by closed reduction and elastic fixation with at least two intramedullary k-wires. RESULTS: Most of the patients were young, with good bone quality and low anesthetic risk, and they had suffered the fractures as a result of a direct trauma. Predominantly uncomplicated, the fractures were metaphyseal, subcapital and of the fifth metacarpal bone (750.3-B1 fractures). Surgical treatment was indicated for a palmar axis dislocation of >20 degrees or if a rotatory deficiency was present. Metacarpal joint function and correction of rotatory displacement could be assessed on median after a period of 1.1 year. In 34 patients flexion and extension was normal on both sides. In one patient we found an extension deficiency of 15 degrees and a rotatory deficiency of 10 degrees . In 34 out 35 patients with metacarpal fractures, minimally invasive intramedullary k-wire osteosynthesis resulted in complete restoration. CONCLUSIONS: Intramedullary k-wire fixation is a minimally invasive method for stabilizing metacarpal fractures. The excellent long-term clinical results are due to the fact that the gliding tissue around the fracture will not be affected at all by the surgical procedure. PMID- 15278370 TI - Effects of age and dietary restriction on oxidative DNA damage, antioxidant protection and DNA repair in rats. AB - BACKGROUND: Experimentally imposed dietary restriction is known to extend the lifespan of rodents, perhaps by slowing the accumulation of oxidative damage that is thought to be one of the causes of aging. AIM OF THE STUDY: We examined the effects of restricted total food intake, and protein and calorie restriction, on DNA oxidation and related biomarkers in rats. METHODS: From 1 to 17 months, rats in group 1 received normal diet ad libitum. Group 2 received 70% of the quantity consumed by the first group. Group 3 had the same quantity as group 2, but with a reduction in protein (from 18% to 10% of the diet by weight), and group 4 were further restricted with a 30% decrease in calories. Lymphocytes were isolated from blood samples taken every two months. DNA breaks, oxidised pyrimidines, resistance to H2O2-induced damage, and strand break repair were measured with the comet assay. Organs were isolated from rats killed at 17 months, with 1 month-old rats for comparison; DNA oxidation and antioxidant enzyme activities were measured. RESULTS: DNA breaks in lymphocytes increased from 1 to 3 months but thereafter declined with age, except in ad libitum fed rats. Oxidised pyrimidines did not change significantly. Resistance to H2O2-induced damage was least at 3 months, and increased with age. Repair of DNA strand breaks was efficient at all ages. Diet had little effect on these endpoints. Diet had no influence on 8-oxo 7.8-dihydroguanine levels in DNA from liver, testis and brain of 17 month old rats. Combining data from all four groups, the levels in brain and liver were significantly higher at 17 months compared with 1 month. Antioxidant enzyme activities tended to increase between 1 and 17 months; effects of diet were not so consistent. CONCLUSIONS: While DNA damage shows a modest increase with age in some organs, antioxidant status and DNA strand break repair do not decline with age. Restricted diets (including protein and calorie restriction) have no effect on any of these markers of genetic stability. PMID- 15278371 TI - No effect on adenoma formation in Min mice after moderate amount of flaxseed. AB - BACKGROUND & AIM: The mammalian lignan enterolactone (ENL) produced from plant lignans, e. g. secoisolariciresinol diglycoside (SDG), may protect against various cancers in humans. The present work aims to evaluate the effect of flaxseed on tumour formation in multiple intestinal neoplasia (Min) mice, a model for colon tumorigenesis. DESIGN: Male and female Min mice were fed either with a non-fibre control diet or the same diet supplemented with 0.5 % (w/w) defatted flaxseed meal. Conversion of SDG to the mammalian lignans enterodiol (END) and ENL in the gut, and plasma ENL, were measured by HPLC with coulometric electrode array detector (CEAD) and timeresolved fluoroimmunoassay, respectively. Wild-type mice were also fed with the experimental diets in order to see whether lignan metabolism is different in Min and wild-type mice. RESULTS: The total number of adenomas or their size in the small intestine was not different in the flaxseed and control groups. The flaxseed group had a tendency for a decreased number of colon adenomas in both genders. Gender and genotype based differences were found in the intestinal ENL levels. When compared to Min females, Min males in the flaxseed group had several fold higher ENL levels in the small intestine (Min males 125 +/- 124.5 nmol/g vs. females 22.8 +/- 16.0 nmol/g, P = 0.048) and caecum (47.6 +/- 31.6 nmol/g vs. females 14.5 +/- 6.6 nmol/g, P = 0.001). Presence of adenomas in the gut influences the intestinal lignan metabolism. Min mice had less intestinal END and ENL as compared with the wild-type mice (P < 0.05). Mean plasma ENL increased 7-fold during the flaxseed feeding (7 nmol/L in control vs. 50 nmol/L in flaxseed group) but no differences between gender and genotype were found. The plasma ENL level did not correlate with adenoma number in the small intestine and colon. CONCLUSION: The number of intestinal adenomas in the Min mouse model is not related to ENL level in plasma nor is it associated with the levels of intestinal lignans. A gender difference in ENL lignan metabolism was found in the gut but not in the plasma. PMID- 15278372 TI - The antioxidative effect of the bacteria Dienococcus radiophilus against LDL lipid peroxidation. AB - BACKGROUND: Lipid peroxidation is an important process in the development of atherosclerosis. Thus agents with antioxidant properties may play an important role in the inhibition of atherosclerosis. OBJECTIVES: In this study we aimed to show that the lipid extract of the bacteria Deinococcus radiophilus (leDR) has antioxidant properties against LDL oxidation. RESULTS: This antioxidant effect was shown in both transition metal ion and free radical generating systems. We also showed that leDR can protect LDL from UV light-induced oxidative damage. The antioxidative capacity of leDR is partly due to copper ion chelation. CONCLUSION: We conclude that some specific bacteria constituent has the ability to inhibit LDL oxidation and, thus, to attenuate atherogenesis. PMID- 15278374 TI - Thoracic neuroblastoma: outcome of incomplete resection. AB - The prognosis for thoracic neuroblastoma has been documented as good, but the reasons have not been elucidated. We reviewed our experience of patients with thoracic neuroblastoma who were treated over the past decade. Among 102 patients treated for neuroblastoma at our hospital between December 1987 and June 1997, 20 patients had thoracic neuroblastoma (stage 1: nine, stage 2: five, stage 3: three, stage 4: three). Tumor characteristics and survival rate were compared between thoracic and nonthoracic neuroblastoma. The surgical margin was positive in 13 of the 20 patients with thoracic neuroblastoma. However, local recurrence was observed in only one patient who later underwent complete resection. All patients survived 4-14 years of follow-up. Among those over 1 year old, thoracic neuroblastoma was detected at an earlier stage than in their nonthoracic counterparts (stages 1 and 2 vs. 3 and 4: 6/3 vs. 1/17, p=0.003), and the 5-year survival rate was better than in their nonthoracic counterparts (100% vs. 44.5%, p=0.015). The incidence of ganglioneuroblastoma was significantly higher in the thoracic group at the age of >1 year ( p=0.003). In six of nine patients from the thoracic group who were >1 year old, small areas of ganglioneuroma were identified in the tumor tissue. There was a stronger tendency for the maturation of neuroblastoma into ganglioneuroma in the thoracic group. Complete resection is not required for thoracic neuroblastoma regardless of the patient's age. PMID- 15278375 TI - Hindgut duplication--case report and literature review. AB - Colonic duplication is a rare congenital abnormality that can present diagnostic difficulties. A case of duplication of the entire large bowel and terminal ileum is presented, and a framework for diagnostic work-up and treatment options in these patients is discussed. PMID- 15278376 TI - Perforation of the piriform recessus by a swallowed glass splinter presenting as pneumomediastinum in a child. AB - The swallowing of sharp glass splinters is rare due to the difficulty of swallowing such objects, and perforation of the piriform recessus and mediastinitis are unusual complications. Perforation of the piriform recessus due to a swallowed sharp glass splinter requires prompt treatment. This report describes a child who was operated on immediately after the diagnosis of perforation of the piriform recessus and was managed successfully. Light-guided pharyngoscopy was very useful for detecting the region of perforation. The literature on such injuries is reviewed here, and the problems associated with treating children with perforation of the piriform recessus and mediastinitis caused by swallowed glass splinters are discussed. PMID- 15278377 TI - The results of antegrade continence enema using a retubularized sigmoidostomy. AB - Left colonic antegrade continence enema (ACE) has been reported only as an alternative to right colonic ACE-the Malone appendicostomy and Monti retubularized ileostomy. This paper evaluated the advantages of left colonic ACE using a retubularized sigmoidostomy (RS) as an appropriate method for maintaining fecal continence and as a first-line surgical treatment for patients with fecal incontinence or intractable constipation. Ten patients underwent surgery between March 2002 and June 2003: seven with meningomyelocele, one with cloacal anomaly, one with anorectal malformation, and one with lipoma of the spine. An RS tube was fashioned and then implanted using a segment of the sigmoid colon and exteriorized through the umbilicus. An enema was done 10 days after surgery using only normal saline. The outcomes were assessed after adjusting to the appropriate enema regimen for the 10 cases. The mean duration of the enema was 23.0+/-8.4 min, with 250 ml (range 80-800) as the median volume of fluid used. The enema interval ranged from 1-3 days. No patient showed any abdominal discomfort or soiling episodes, with the exception of one who experienced daytime fecal staining, but this occurred less than once per month. The self-cosmesis for the umbilical stoma was satisfactory. The RS procedure provided excellent continence control, with a shortening of the enema duration, a lower fluid volume, and good cosmesis, and without any ACE-related abdominal pain. This procedure can be used as a first-choice surgical treatment for intractable constipation and fecal incontinence. PMID- 15278380 TI - Separation surgery for total vertical craniopagus twins. AB - OBJECTIVE: A pair of conjoined twins aged 11 months underwent investigations, followed by surgical separation in Singapore General Hospital in April 2001. They were joined at the skull vertex and facing in opposite directions. METHODS: Radiological investigations including cerebral angiography, magnetic resonance imaging and computerized tomographic scans were performed, leading to the diagnosis of total vertical craniopagus. There were two separate brains, with separate arterial circulations, but with a common superior sagittal sinus. Tissue expanders were inserted in the subgaleal space for 6 months of scalp expansion prior to surgery. Pre-operative planning involved the use of virtual reality equipment and life-sized polymer models of the conjoined skulls and brains. Surgical separation of the twins was achieved after approximately 100 h of operating time, using intraoperative image guidance, microsurgical techniques and intraoperative neurophysiologic monitoring. Reconstruction of the dura, calvarium and scalp was performed with artificial dura, absorbable plates and split skin grafts. Postoperative complications included focal cortical infarction, meningitis, and hydrocephalus. CONCLUSION AND OUTCOME: Despite these complications, the twins recovered satisfactorily and were discharged to their home country within 6 months. The 3-month outcome was minor disability in one twin and severe developmental delays in the other. Separation surgery is possible for complex cranially-conjoined twins but requires detailed planning and extensive surgical management. PMID- 15278381 TI - Advanced "tactile" medical imaging for separation surgeries of conjoined twins. AB - BACKGROUND: Understanding the anatomy of conjoined twins is difficult because of the rarity of this congenital malformation and the scarcity of reported cases in medical literature. NEW TECHNOLOGY: Advances in radiologic imaging, computer modeling, and sophisticated manufacturing techniques enable medical imaging datasets to be translated into accurate, solid, life-size models. These models, which can be designed to include various combinations of anatomical features revealed by established imaging modalities, are important for pre-surgical assessments and planning, as well as for reference during the actual operative procedure. In addition, they provide a valuable basis for communication between the groups of specialists who are involved in these cases. CASE STUDIES: This article will describe the advances in technology behind this process and illustrate its value in two cases of craniopagus twins. PMID- 15278382 TI - The embryology of conjoined twins. AB - INTRODUCTION: Attention is drawn to the spontaneous incidence of twinning, both dizygotic and monozygotic in different mammalian species. Conjoined twinning, however, only arises when the twinning event occurs at about the primitive streak stage of development, at about 13-14 days after fertilisation in the human, and is exclusively associated with the monoamniotic monochorionic type of placentation. It is believed that the highest incidence of conjoined twinning is encountered in the human. While monozygotic twinning may be induced experimentally following exposure to a variety of agents, the mechanism of induction of spontaneous twinning in the human remains unknown. All agents that are capable of acting as a twinning stimulus are teratogenic, and probably act by interfering with the spindle apparatus. DISCUSSION: The incidence of the various types of conjoined twinning is discussed. Information from the largest study to date indicates that the spontaneous incidence is about 10.25 per million births. The most common varieties encountered were thoraco-omphalopagus (28%), thoracopagus (18.5%), omphalopagus (10%), parasitic twins (10%) and craniopagus (6%). Of these, about 40% were stillborn, and 60% liveborn, although only about 25% of those that survived to birth lived long enough to be candidates for surgery. Conjoined twinning occurs by the incomplete splitting of the embryonic axis and, with the exception of parasitic conjoined twins, all are symmetrical and "the same parts are always united to the same parts". Fusion of monozygotic twins is no longer believed to be the basis of conjoined twinning. Accounts are provided of the anatomical features of each of the commonly encountered varieties. PMID- 15278383 TI - Separation of craniopagus twins in the era of modern neuroimaging, interventional neuroradiology, and frameless stereotaxy. AB - INTRODUCTION: Craniopagus twinning is a rare embryological event occurring in 1 in 2.5 million births. CASE REPORT: We present our recent experience with the separation of total vertical craniopagus twins in the modern era of neuroimaging and interventional neuroradiology. Three-dimensional CT images revealed the twins' heads were axially rotated 30 degrees. MRI showed deficient dura between the brains of the twins, and some sharing of parietal brain tissue. Cerebral angiography showed a dominant arterial circulation of one twin with unilateral middle cerebral artery (MCA) branches feeding the other twin. The twins shared a common superior sagittal sinus in its middle segment where a circular sinus was formed. Prior to surgery, endovascular separation of the twins' arterial and venous circulations was achieved in part using tantalum coils for the MCA feeders in one twin, and balloon occlusion of the anterior superior sagittal sinus of the other. Using the ISG wand intraoperatively, surgery proceeded stepwise and included the circumferential removal of bone and opening of the dura, separation of the twins' brains along leptomeningeal planes, and identification of the major draining veins from the superficial cerebral cortex. DISCUSSION: The separation of craniopagus twins demands a multidisciplinary team approach. Utilizing preoperative neuroendovascular techniques to occlude shared vascular anastomotic channels, complex total vertical craniopagus twins can now be successfully separated in a one-stage procedure. PMID- 15278384 TI - Ischiopagus and pygopagus conjoined twins: neurosurgical considerations. AB - BACKGROUND: Neurosurgeons are familiar with the challenges presented by craniopagus twins, but other types of conjoined twins may also have neurosurgical implications. We report our experience in the management of ischiopagus and pygopagus conjoined twins. METHODS: This is a retrospective review of the management of conjoined twins at Red Cross Children's Hospital in Cape Town, South Africa. RESULTS: Twenty-three pairs of symmetrical conjoined twins were managed over a 40-year period (1964-2003), of which 16 (70%) were separated. Of these cases, 6 are the focus of this study, namely 4 pairs of ischiopagus twins and 2 pairs of pygopagus twins seen between 1993 and 2003. In 2 cases, there was direct involvement of the nervous system at the site of union, with 1 pair of ischiopagi manifesting end-to-end union of their spinal cords, while a pair of pygopagi had back-to-back fusion of the conus. Another pair of ischiopagi had a fused dural sac without joined neural elements, but one of these children developed syringomyelia 2 years after separation. Neuroimaging was invaluable in detecting these abnormalities. The one pair of ischiopagi who died before separation were HIV positive and had severe brain atrophy and cystic encephalmalacia at autopsy. Nine of the 12 children (75%) had bony abnormalities of the spine remote from the area of conjunction. The most common finding was the presence of hemivertebrae, usually in the thoracic spine. Six children manifested scoliosis, which has already progressed in the oldest two. Technical aspects such as timing and sequence of separation, the division of neural tissues and reconstruction are discussed, as are the long-term complications of their spinal abnormalities. CONCLUSIONS: Ischiopagus and pygopagus conjoined twins manifest an interesting array of spinal abnormalities, which present challenges, not only at the time of separation, but also in their long-term management. PMID- 15278385 TI - Craniopagus twins: embryology, classification, surgical anatomy, and separation. AB - INTRODUCTION: With recent advances in brain imaging and neurosurgical techniques, there has been a renewed interest in the surgical separation of craniopagus twins. Successful separation in recent cases, along with widespread publicity, has attracted craniopagus twins from all over the world to be referred to pediatric neurosurgical centers for evaluation and consideration for surgical separation. SEPARATION OF BLOOD SUPPLY: It has become apparent, however, that the most critical decisions in surgical planning are related to separation of the blood supply to the conjoined brains. In fact, in craniopagus twins that survive pregnancy or the first few days of life, there is usually little shared brain tissue. The shared blood supply is far and away the more critical issue. It is very difficult to successfully separate craniopagus twins in one surgical procedure. Staged separation, with gradual re-routing of the shared blood supply, has been a successful alternative. CASE STUDIES AND DISCUSSION: We discuss here our experience with three sets of craniopagus twins and our approach to staged separation. PMID- 15278386 TI - Complications of bioresorbable fixation systems in pediatric neurosurgery. AB - OBJECTIVE: Bioresorbable devices are an attractive alternative to metal instrumentation for internal fixation of bone, and have been used extensively in orthopedic and craniofacial surgery. In neurosurgery, the reported literature is predominantly confined to pediatric craniofacial procedures, with encouraging results and minimal complications. We have used bioreabsorbable plates and screws in cranial and spinal pediatric neurosurgery procedures. We report four complications related to their usage. MATERIALS AND METHODS: Bioabsorbable instrumentation was used in pediatric patients for fixation of bone after cranial or spinal procedures. RESULTS: Four patients developed complications related to the instrumentation: 2 following cranial surgery for epilepsy, 1 after correction of a growing skull fracture, and 1 after laminotomy for an intramedullary tumor. Two patients had fibrous encapsulation with granuloma formation and 2 patients had osteolysis following the fixation. CONCLUSION: Bioabsorbable fixation devices for the stabilization of bone following craniotomy and laminotomy in pediatric patients may be associated with complications, including granuloma formation and osteolysis. PMID- 15278387 TI - Coronary artery calcification detected by a mobile helical computed tomography unit and future cardiovascular death: 4-year follow-up of 6120 asymptomatic Japanese. AB - In the present study, we performed a prospective follow-up study in a population which underwent chest computed tomography (CT) screening. A total of 6120 participants underwent a chest CT medical examination for lung cancer and tuberculosis in Nagano Prefecture, Japan, between 1996 and 1997. Computed tomography scanning was performed from the apex of the lung to the diaphragm at a tube voltage of 120 kV and a tube current of 50 mA. We measured the CT density of the coronary arteries in 5-7 slices where coronary arteries were detected. The CT density threshold for determining coronary artery calcification (CAC) was above +110 HU. In 2000, we investigated the number of deaths due to cardiac and noncardiac disease among the participants. Of the 6120 participants, 14 died of cardiac disease (9, myocardial infarction; 4, heart failure; and 1, angina pectoris) and 64 died of other diseases. Coronary artery calcification was detected in 10 of the patients who died of cardiac disease, and in 31 of those who died of other diseases. The prevalence of CAC was higher in the former than in the latter (71.4% vs 48.4%, P = 0.084). The relative risk of CAC for cardiac death was 2.66 (95% confidence interval: 0.76, 9.37). The findings of this study suggested that CAC detected in a mass chest CT screening by a mobile helical CT unit was predictive of future cardiovascular death. PMID- 15278388 TI - Prothrombin G20210A gene mutation with LightCycler polymerase chain reaction in venous thrombosis and healthy population in the southeast of Turkey. AB - Venous thrombosis (VT) is a common disease, with an annual incidence in the general population of approximately 1 per 1000. The prevalence of genetic risk factors for thrombosis varies greatly in different parts of the world. Prothrombin G20210A (PT G20210A) gene mutation has been recently identified as a common risk factor in venous thrombosis. Sixty-one patients with VT, differing in age and sex, and 340 healthy subjects were consecutively enrolled into our study to determine the prevalence of PT G20210A in VT and in the healthy population of the southeast of Turkey. The mutation was identified with fluorescence resonance energy transfer (FRET) with the LightCycler polymerase chain reaction. The PT G20210A mutation was found to be 6.5% (4/61) in the VT group and 1.2% (4/340) in the healthy group ( P = 0.021). Three patients with VT had a heterozygous PT G20210A mutation, and the other patient with VT had both Factor V Leiden and PT G20210A mutations. We showed that this method may be used safely for detection of the PT G20210A gene mutation, and the prevalence of PT G20210A mutation is significantly higher in patients with VT than in the healthy population in the southeast of Turkey. PMID- 15278389 TI - In vivo effect of losartan on platelet aggregation in patients with hypertension. AB - The angiotensin II receptor, losartan, has been found to inhibit platelet aggregability to some extent in in vitro experiments. There have been conflicting results about the in vivo effects of losartan. We sought to clarify the in vivo effect of losartan on platelet aggregation. Forty patients with grade I essential hypertension were treated with losartan for 3 weeks. Platelet aggregation tests with adenosine diphosphate (ADP) and ristocetin were analyzed and compared before and at the end of the study. Losartan effectively decreased systolic (SBP) and diastolic (DBP) blood pressure. Mean SBP before and after treatment was 159.6 +/- 12.8 and 149.2 +/- 17.3 mmHg, respectively. Mean DBP decreased from 93.7 +/- 8.2 to 87.7 +/- 10.3 mmHg after treatment. The results of the platelet aggregation tests with ADP and ristocetin were not significantly different when both rate and amplitude of maximal aggregation were included. Peak platelet aggregation with ADP regarding the lowest light transmission in the aggregometer was 59.8% +/- 24.3% before and 58.3% +/- 18.1% after the treatment. The same variables with ristocetin were 66.8% +/- 21.6% and 60.8% +/- 23.3%, respectively. In vivo effects of losartan on platelet aggregation with ADP and ristocetin were insignificant. PMID- 15278390 TI - Impact of mitral regurgitation on long-term survival in patients with ischemic cardiomyopathy: efficacy of combined mitral valve repair and revascularization. AB - Ischemic cardiomyopathy complicated by severe mitral regurgitation (MR) has a poor prognosis. In such cases, whether mitral valve repair for MR improves the prognosis of survival remains unclear. In this study, 50 patients diagnosed with ischemic cardiomyopathy at our hospital between August 1991 and August 1996 were studied to examine the long-term prognosis and factors determining the prognosis. Among 17 patients with the complication of severe MR, 11 underwent mitral valve repair (repair group) and 6 did not (nonrepair group). Among the 33 patients without MR, 15 underwent revascularization (revascularization group) and 18 received medical treatment alone (medical group). Patients with MR showed significantly poorer baseline activities of daily living (ADL) [New York Heart Association (NYHA) class III or above: MR(+) vs MR(-) = 14 vs 8; P = 0.0001] and survival rate [MR(+) vs MR(-); log rank = 3.8, P = 0.05]. In contrast, patients in whom mitral valve repair was actively performed to resolve MR had favorable outcomes for both ADL (NYHA class improved from 3.9 +/- 0.3 to 2.7 +/- 1.0; P = 0.0004) and survival rate (MV repair vs nonrepair: long rank = 10.1, P = 0.0015). In addition, among patients without MR, the revascularization group showed more favorable results in terms of ADL (NYHA class improved from 3.5 +/- 0.7 to 2.5 +/ 0.8; P = 0.0059) and survival rate (revascularization vs medical: log rank = 3.7, P = 0.05), irrespective of improvement of left ventricular function. When the factors determining the prognosis for ischemic cardiomyopathy were examined by multivariate analysis, whether or not revascularization was conducted, the presence or absence of mitral regurgitation, and if present, whether or not mitral valve repair was performed were identified as independent factors determining the prognosis (revascularization: hazard ratio = 0.121, P = 0.012; absence of MR: hazard ratio = 0.104, P = 0.050; mitral valve repair: hazard ratio = 0.018, P = 0.005). These results showed that revascularization should be conducted as actively as possible in patients with ischemic cardiomyopathy; in addition, for those patients with mitral regurgitation, mitral valve repair should be conducted actively to relieve it. PMID- 15278391 TI - Mid-term results of mitral valve repair for complicated active bacterial endocarditis in high-risk patients. AB - Mitral valve repair in endocarditis achieves a competent valve and prevents septic embolization and acute left ventricular failure, in which operative mortality could be increased. Early and mid-term results were examined to establish whether emergency mitral valve repair offers an advantage in complicated active endocarditis. Ten patients with complicated active native valve endocarditis underwent mitral valve repair. The mean age was 45.8 +/- 18.5 years; two patients were female (20%). All patients had severe mitral regurgitation, which combined in one patient with mitral valve stenosis. New York Heart Association (NYHA) functional class was IV in all patients. The macroscopically infected tissue with vegetation in all patients was excised. Multiple techniques were required to achieve valve competence. There was one (10%) hospital death in a patient with persistent congestive heart failure, and a reoperation in another (10%) after 2 years. Mean follow-up was 32.1 +/- 12.7 months (range 1-45 months) and was complete. There were no late deaths, recurrent endocarditis, or thromboembolic events. Seven patients (77.7%) were in NYHA functional class I, and two (22.2%) were in class II. Mitral valve repair in complicated active bacterial endocarditis limited to leaflet tissues has a low operative mortality and valve-related morbidity, with promising mid-term survival in high-risk patients. PMID- 15278392 TI - Left ventricular remodeling and aortic distensibility in elite power athletes. AB - The aim of this study was to determine left ventricular (LV) morphology and aortic function in power athletes and to compare them with normal subjects. Thirty-two elite male wrestlers and 15 age-matched healthy male controls were included. All subjects underwent echocardiographic examination. Measurements included LV cavity dimension at systole and diastole, wall thickness, diastolic parameters, and aortic diameter, 3 cm above aortic valve, at systole and diastole. Left ventricular mass and mass index were found to be higher in the athletes than in control subjects. The aortic distensibility index was found to be reduced in the athletes compared with controls (2.53 +/- 0.91 vs 3.94 +/- 1.77 cm(2) dyne(-1) 10(-6), P = 0.003), while the aortic stiffness index was significantly higher in the athletes than in controls (9.12 +/- 3.23 vs 6.65 +/- 2.35, P = 0.02). However, LV end-systolic wall stress was lower in the athletes than in controls. Furthermore, transmitral early ( E) and late ( A) peak velocity, peak velocity of the myocardial systolic wave ( S(m)), and early ( E(m)) and atrial ( A(m)) diastolic waves at the inferior wall were higher in the athletes than in controls. Reduced aortic distensibility in elite power athletes may be one of the cardiovascular adaptation factors which affect LV hypertrophy. PMID- 15278393 TI - Effect of age on carotid arterial intima-media thickness in childhood. AB - To investigate relationships between carotid arterial intima-media thickness (IMT) and age in childhood, we performed high-resolution carotid arterial ultrasonography in 60 healthy children (27 boys, 33 girls; age range, 5-14 years) determined by screening to have no dyslipidemia or hypertension. No plaque formation was found, and irregularity of IMT (root mean square roughness of IMT) did not correlate with age. Mean IMT increased in a linear manner with age [IMT in millimeters = (0.009 x age in years) + 0.35] ( r = 0.39, P = 0.002). This correlation remained significant after adjustment for gender, parental smoking, systolic and diastolic blood pressure, body mass index, and serum concentrations of low-density lipoprotein cholesterol, high-density lipoprotein cholesterol, and triglycerides. None of these known cardiovascular disease risk factors in adults had a significant relationship with age-adjusted IMT in children. While circumferential wall stress and diastolic blood pressure were not correlated with age, mean IMT and lumen diameter showed significant positive relationships with circulating blood volume, which was calculated as the function of height and weight. These data suggested that age-dependent physiologic thickening of arterial walls begins in childhood. PMID- 15278394 TI - Double left anterior descending coronary artery arising from the left and right coronary arteries: a rare congenital coronary artery anomaly. AB - Double left anterior descending coronary artery arising from the left and right coronary arteries is a very rare congenital coronary artery anomaly. In this report, we describe a patient with double left anterior descending coronary artery originating from the left and right coronary arteries. To the best of our knowledge, dual connection of the left anterior descending coronary artery to the left and right coronary arteries has been described in only five patients. PMID- 15278395 TI - A case of primary cardiac lymphoma located in the pericardial effusion. AB - Primary cardiac lymphoma is a rare disorder with a poor prognosis. We present here a case of 77-year-old woman who was diagnosed as having cardiac lymphoma antemortem according to a cytologic examination of the pericardial effusion. Determination of the levels of serum-soluble interleukin-2 receptor and serum deoxythymidine kinase was useful for the diagnosis. Echocardiography, computed tomography, magnetic resonance imaging, and gallium scan revealed neither lymphadenopathy nor tumor in the heart, so she was diagnosed as having malignant lymphoma that probably originated from the pericardium. Systemic chemotherapy with CHOP (cyclophosphamide, farmorubicin, oncovin, and prednisolone) resulted in a complete resolution of the pericardial effusion. She has been in remission 48 months after discontinuation of the chemotherapy. PMID- 15278396 TI - Living-related lobar transplantation and simultaneous atrial septal defect closure in a young patient with irreversible pulmonary hypertension: a case report. AB - A 6-year-old boy was diagnosed as having atrial septal defect with oversystemic pulmonary hypertension, and gradually developed hypoxia and heart failure. At the age of 11, living-related bilateral lobar lung transplantation from his parents was indicated, because of his critical condition. The estimated forced vital capacity calculated by the donors' lower lobes was 104% of his age. The operation was carried out with simultaneous closure of the atrial septal defect under full cardiopulmonary bypass. The postoperative course was complicated with pulmonary edema and phrenic nerve palsy, which were eventually resolved. Six months after surgery he was free from heart failure and rehabilitating at home without oxygen. The final diagnosis was primary pulmonary hypertension with atrial septal defect. Living-related bilateral lobar lung transplantation with simultaneous intracardiac repair may be an optional strategy for children with Eisenmenger syndrome or primary pulmonary hypertension with intracardiac defect when cadaveric transplantation is not possible. PMID- 15278397 TI - Rupture of a thoracic aortic aneurysm: a rare adverse reaction following systemic tissue plasminogen activator infusion. AB - We present a patient with rupture of a thoracic aortic aneurysm occurring after systemic infusion of tissue plasminogen activator for the treatment of acute ischemic stroke, which was successfully treated with the placement of an endovascular stent-graft. PMID- 15278398 TI - Respiration of individual honeybee larvae in relation to age and ambient temperature. AB - The CO(2) production of individual larvae of Apis mellifera carnica, which were incubated within their cells at a natural air humidity of 60-80%, was determined by an open-flow gas analyzer in relation to larval age and ambient temperature. In larvae incubated at 34 degrees C the amount of CO(2) produced appeared to fall only moderately from 3.89 +/- 1.57 microl mg(-1) h(-1) in 0.5-day-old larvae to 2.98 +/- 0.57 microl mg(-1) h(-1) in 3.5-day-old larvae. The decline was steeper up to an age of 5.5 days (0.95 +/- 1.15 microl mg(-1) h(-1)). Our measurements show that the respiration and energy turnover of larvae younger than about 80 h is considerably lower (up to 35%) than expected from extrapolations of data determined in older larvae. The temperature dependency of CO(2) production was determined in 3.5-day-old larvae, which were incubated at temperatures varying from 18 to 38 degrees C in steps of 4 degrees C. The larvae generated 0.48+/-0.03 microl mg(-1) h(-1) CO(2) at 18 degrees C, and 3.97 +/- 0.50 microl mg(-1) h(-1) CO(2) at 38 degrees C. The temperature-dependent respiration rate was fitted to a logistic curve. We found that the inflection point of this curve (32.5 degrees C) is below the normal brood nest temperature (33-36 degrees C). The average Q(10) was 3.13, which is higher than in freshly emerged resting honeybees but similar to adult bees. This strong temperature dependency enables the bees to speed up brood development by achieving high temperatures. On the other hand, the results suggest that the strong temperature dependency forces the bees to maintain thermal homeostasis of the brood nest to avoid delayed brood development during periods of low temperature. PMID- 15278399 TI - The role of call frequency and the auditory papillae in phonotactic behavior in male Dart-poison frogs Epipedobates femoralis (Dendrobatidae). AB - Territorial males of the pan-Amazonian Dart-poison frog, Epipedobates femoralis, are known to present stereotypic phonotactic responses to the playback of conspecific and synthetic calls. Fixed site attachment and a long calling period within an environment of little temperature change render this terrestrial and diurnal pan-Amazonian frog a rewarding species for field bioacoustics. In experiments at the field station Aratai, French Guiana, we tested whether the prominent frequency modulation of the advertisement-call notes is critical for eliciting phonotactic responses. Substitution of the natural upward sweep by either a pure tone within the species frequency range or a reverse sweep did not alter the males' phonotactic behavior. Playbacks with artificial advertisement calls embedded in high levels of either low-pass or high-pass masking noise designed to saturate nerve fibers from either the amphibian papilla or basilar papilla showed that male phonotactic behavior in this species is subserved by activation of the basilar papilla of the inner ear. PMID- 15278400 TI - Saccadic head and thorax movements in freely walking blowflies. AB - Visual information processing is adapted to the statistics of natural visual stimuli, and these statistics depend to a large extent on the movements of an animal itself. To investigate such movements in freely walking blowflies, we measured the orientation and position of their head and thorax, with high spatial and temporal accuracy. Experiments were performed on Calliphora vicina, Lucilia cuprina and L. caesar. We found that thorax and head orientation of walking flies is typically different from the direction of walking, with differences of 45 degrees common. During walking, the head and the thorax turn abruptly, with a frequency of 5-10 Hz and angular velocities in the order of 1,000 degrees /s. These saccades are stereotyped: head and thorax start simultaneously, with the head turning faster, and finishing its turn before the thorax. The changes in position during walking are saccade-like as well, occurring synchronously, but on average slightly after the orientation saccades. Between orientation saccades the angular velocities are low and the head is held more stable than the thorax. We argue that the strategy of turning by saccades improves the performance of the visual system of blowflies. PMID- 15278401 TI - Rapid firing rates from mechanosensory neurons in copepod antennules. AB - Small detection distances coupled with rapid movements require copepods to respond to stimuli with behavioral latencies on the order of milliseconds. Receiving adequate sensory information in such a short time necessitates extremely rapid firing rates of the efferent receptors. Here we show that copepod mechanoreceptors can fire at frequencies up to 5 kHz in response to fluid mechanical stimuli. Neural activity at these frequencies enables these animals to code for a range of fluid velocities thus providing important information regarding the nature of different fluid disturbances. PMID- 15278407 TI - Rectal endometriosis: MRI study with rectal coil. PMID- 15278409 TI - Percutaneous thrombin injection for the treatment of a post-pancreatitis pseudoaneurysm of the gastroduodenal artery. PMID- 15278412 TI - Influence of detector collimation on SNR in four different MDCT scanners using a reconstructed slice thickness of 5 mm. AB - The purpose of this paper is to compare the influence of detector collimation on the signal-to-noise ratio (SNR) for a 5.0 mm reconstructed slice thickness for four multi-detector row CT (MDCT) units. SNRs were measured on Catphan test phantom images from four MDCT units: a GE LightSpeed QX/I, a Marconi MX 8000, a Toshiba Aquilion and a Siemens Volume Zoom. Five-millimetre-thick reconstructed slices were obtained from acquisitions performed using detector collimations of 2.0-2.5 mm and 5.0 mm, 120 kV, a 360 degrees tube rotation time of 0.5 s, a wide range of mA and pitch values in the range of 0.75-0.85 and 1.25-1.5. For each set of acquisition parameters, a Wiener spectrum was also calculated. Statistical differences in SNR for the different acquisition parameters were evaluated using a Student's t-test (P<0.05). The influence of detector collimation on the SNR for a 5.0-mm reconstructed slice thickness is different for different MDCT scanners. At pitch values lower than unity, the use of a small detector collimation to produce 5.0-mm thick slices is beneficial for one unit and detrimental for another. At pitch values higher than unity, using a small detector collimation is beneficial for two units. One manufacturer uses different reconstruction filters when switching from a 2.5- to a 5.0-mm detector collimation. For a comparable reconstructed slice thickness, using a smaller detector collimation does not always reduce image noise. Thus, the impact of the detector collimation on image noise should be determined by standard deviation calculations, and also by assessing the power spectra of the noise. PMID- 15278415 TI - Dobutamine stress MRI. Part I. Safety and feasibility of dobutamine cardiovascular magnetic resonance in patients suspected of myocardial ischemia. AB - The aim of the study was to evaluate safety and feasibility of dobutamine cardiovascular magnetic resonance (CMR) in patients with proven or suspected coronary artery disease. Dobutamine CMR was evaluated retrospectively in 400 consecutive patients with suspicion of myocardial ischemia. Dobutamine was infused using an incremental protocol up to 40 microg/kg body weight per minute. All anti-anginal medication was stopped 4 days before the CMR study and infusion time of dobutamine was 6 min per stage. Hemodynamic data, CMR findings and side effects were reported. Patients with contraindications to CMR (metallic implants and claustrophobia) were excluded from analysis. Dobutamine CMR was successfully performed in 355 (89%) patients. Forty-five (11%) patients could not be investigated adequately because of non-cardiac side effects in 29 (7%) and cardiac side effects in 16 (4%) patients. Hypotension (1.5%) and arrhythmias (1%) were the most frequent cardiac side effects. One patient developed a severe complication (ventricular fibrillation) at the end of the study. There were no myocardial infarctions or fatal complications of the stress test. The most frequent non-cardiac side effects were nausea, vomiting and claustrophobia. Age >70 years, prior myocardial infarction and rest wall motion abnormalities showed no significant differences with side effects (P>0.05). Dobutamine CMR is safe and feasible in patients with suspicion of myocardial ischemia. PMID- 15278416 TI - Dobutamine stress MRI. Part II. Risk stratification with dobutamine cardiovascular magnetic resonance in patients suspected of myocardial ischemia. AB - The aim of this study was to determine the prognostic value of dobutamine cardiovascular magnetic resonance (CMR) in patients suspected of myocardial ischemia. Clinical data and dobutamine-CMR results were analyzed in 299 consecutive patients. Follow-up data were analyzed in categories of risk levels defined by the history of coronary artery disease and presence of rest wall motion abnormalities (RWMA). Major adverse cardiac events (MACE) as evaluated end points included cardiac death, nonfatal myocardial infarction and clinically indicated coronary revascularization. Follow-up was completed in 214 (99%) patients with a negative dobutamine-CMR study (no signs of inducible myocardial ischemia) with an average of 24 months. The patients with a negative dobutamine CMR study and RWMA showed a significantly higher annual MACE rate (18%) than the patients without RWMA (0.56%) ( P<0.001). Patients without RWMA showed an annual MACE rate of 2% when they had a history of coronary artery disease and <0.1% without a previous coronary event ( P<0.001). Dobutamine-CMR showed a positive and negative predictive value of 95 and 93%, respectively. The cardiovascular occurrence-free survival rate was 96.2%. In patients suspected of myocardial ischemia, dobutamine-CMR is able to assess risk levels for coronary events with high accuracy. PMID- 15278417 TI - Evaluation of haemoglobin (erythrogen): for improved somatic embryogenesis and plant regeneration in cotton (Gossypium hirsutum L. cv. SVPR 2). AB - Somatic embryogenesis in cotton (Gossypium hirsutum L.) is accelerated when the plant regeneration medium is supplemented with haemoglobin (erythrogen). In cotton SVPR 2 lines, a higher frequency of embryoid formation was observed when the medium contained 400 mg/l haemoglobin. Fresh weight of the callus, rate of embryoid induction, number of embryoids formed and the percentage of plant regeneration from somatic embryos were increased. Among the two different cultivars tested, MCU 11 showed no response to the presence of haemoglobin when compared to SVPR 2, and embryogenic callus formation was completely absent in the former. Medium containing MS salts, 100 mg/l myo-inositol , 0.3 mg/l thiamine HCL, 0.3 mg/l Picloram (PIC), 0.1 mg/l kinetin and 400 mg/l haemoglobin effected a better response with respect to embryogenic callus induction. After 8 weeks of culture, a high frequency of embryoid induction was observed on medium containing MS basal salts, 100 mg/l myo-inositol, 0.3 mg/l PIC , 0.1 mg/l isopentenyl adenine, 1.0 g/l NH4NO3 and 400 mg/l haemoglobin. Plant regeneration was observed in 75.8% of the mature somatic embryos, and whole plant regeneration was achieved within 6-7 months of culture. The regenerated plantlets were fertile and similar to in vivo-grown, seed-derived plants except that they were phenotypically smaller. A positive influence of haemoglobin was observed at concentrations up to 400 mg/l at all stages of somatic embryogenesis. The increase in the levels of antioxidant enzyme activities, for example superoxide dismutase and peroxidase, indicated the presence of excess oxygen uptake and the stressed condition of the plant tissues that arose from haemoglobin supplementation. This increased oxygen uptake and haemoglobin-mediated stress appeared to accelerate somatic embryogenesis in cotton. PMID- 15278418 TI - Identification, cloning and sequence analysis of a dwarf genome-specific RAPD marker in rubber tree [Hevea brasiliensis (Muell.) Arg]. AB - High-yielding dwarf clones of Hevea brasiliensis are tolerant to wind damage and therefore useful for high-density planting. The identification of molecular markers for the dwarf character is very important for isolating true-to-type high yielding dwarf hybrid lines in the early stage of plant breeding programs. We have identified a dwarf genome-specific random amplified polymorphic DNA (RAPD) marker in rubber tree. A total of 115 random oligonucleotide 10-mer primers were used to amplify genomic DNA by PCR, of which 19 primers produced clear and detectable bands. The primer OPB-12 generated a 1.4-kb DNA marker from both natural and controlled F(1) hybrid progenies (dwarf stature) derived from a cross between a dwarf parent and a normal cultivated clone as well as from the dwarf parent; it was absent in other parent (RRII 118). To validate this DNA marker, we analyzed 22 F(1) hybrids (13 with a dwarf stature and nine with a normal stature); the dwarf genome-specific 1.4-kb RAPD marker was present in all dwarf stature hybrids and absent in all normal-stature hybrids. This DNA marker was cloned and characterized. DNA marker locus specificity was further confirmed by Southern blot hybridization. Our results indicate that Southern blot hybridization of RAPD using probes made from cloned DNA fragments allows a more accurate analysis of the RAPD pattern based on the presence/absence of specific DNA markers than dye-stained gels or Southern blot analysis of RAPD blots using probes made from purified PCR products. Detection of RAPD markers in the hybrid progenies indicates that RAPD is a powerful tool for identifying inherited genome segments following different hybridization methods in perennial tree crops. PMID- 15278420 TI - Expression of a salt-induced protein (SALT) in suspension-cultured cells and leaves of rice following exposure to fungal elicitor and phytohormones. AB - Phytohormones are essential signal compounds in the regulation of stress-related and defense-related genes. However, there is no clear evidence for any effect of these signal molecules and biotic elicitors on the regulation of the SALT gene in suspension-cultured rice cells. We characterized the expression of a SALT gene following treatment with fungal elicitor, phytohormones, cycloheximide, and inhibitors of protein kinase/phosphatases. SALT expression was up-regulated following treatment with a fungal elicitor, jasmonic acid (JA), abscisic acid (ABA), and NaCl. However, salicylic acid (SA) alone or in combination with one of the other elicitors not only strongly inhibited SALT gene expression but also exhibited an antagonistic effect in suspension cells and leaves. Cycloheximide inhibited SALT accumulation in suspension cells and in leaves, but the inhibitors of protein kinase/phosphatase did not. Immunolocalization revealed that SALT protein was present in xylem parenchyma cells of vascular bundles in the major and minor leaf veins. PMID- 15278421 TI - The rice metallothionein gene promoter does not direct foreign gene expression in seed endosperm. AB - We generated transgenic tobacco and rice plants harboring a chimeric gene consisting of the 5'-upstream sequence of the rice metallothionein gene (ricMT) fused to the beta-glucuronidase (GUS) gene. The activity and tissue-specific expression of the ricMT promoter were demonstrated in these transgenic plants. In the transgenic rice plants, despite substantial levels of GUS activity in the shoot and root, almost no GUS signal was detected in the endosperm. Thus, the ricMT promoter could be useful in avoiding accumulation of undesired proteins in the seed endosperm. PMID- 15278422 TI - Paroxysmal nocturnal hemoglobinuria in a girl with hemolysis and "hematuria". AB - Paroxysmal nocturnal hemoglobinuria (PNH) is a rare disorder of unknown frequency. In its classic form, PNH is characterized by hemolysis accompanied by nocturnal hemoglobinuria. The clinical course is unpredictable and may vary from severe hemolysis and recurrent venous thrombosis to latent periods with milder symptoms. We report a 15-year-old girl with hemolytic episodes, abdominal pain, and passage of dark urine. Hemoglobinuria was demonstrated by a "blood"-positive dipstick test in the absence of red blood cells in the urinary sediment. The diagnosis of PNH was confirmed by flow cytometry. PMID- 15278423 TI - Geographic variation in patterns of nestedness among local stream fish assemblages in Virginia. AB - Nestedness of faunal assemblages is a multi-scale phenomenon, potentially influenced by a variety of factors. Prior small-scale studies have found freshwater fish species assemblages to be nested along stream courses as a result of either selective colonization or extinction. However, within-stream gradients in temperature and other factors are correlated with the distributions of many fish species and may also contribute to nestedness. At a regional level, strongly nested patterns would require a consistent set of structuring mechanisms across streams, and correlation among species' tolerances of the environmental factors that influence distribution. Thus, nestedness should be negatively associated with the spatial extent of the region analyzed and positively associated with elevational gradients (a correlate of temperature and other environmental factors). We examined these relationships for the freshwater fishes of Virginia. Regions were defined within a spatial hierarchy and included whole river drainages, portions of drainages within physiographic provinces, and smaller subdrainages. In most cases, nestedness was significantly stronger in regions of smaller spatial extent and in regions characterized by greater topographic relief. Analysis of hydrologic variability and patterns of faunal turnover provided no evidence that inter-annual colonization/extinction dynamics contributed to elevational differences in nestedness. These results suggest that, at regional scales, nestedness is influenced by interactions between biotic and abiotic factors, and that the strongest nestedness is likely to occur where a small number of organizational processes predominate, i.show $132#e., over small spatial extents and regions exhibiting strong environmental gradients. PMID- 15278424 TI - Herbivore population suppression by an intermediate predator, Phytoseiulus macropilis, is insensitive to the presence of an intraguild predator: an advantage of small body size? AB - Recent work in terrestrial communities has highlighted a new question: what makes a predator act as a consumer of herbivores versus acting as a consumer of other predators? Here we test three predictions from a model (Rosenheim and Corbett in Ecology 84:2538-2548) that links predator foraging behavior with predator ecology: (1) widely foraging predators have the potential to suppress populations of sedentary herbivores; (2) sit and wait predators are unlikely to suppress populations of sedentary herbivores; and (3) sit and wait predators may act as top predators, suppressing populations of widely foraging intermediate predators and thereby releasing sedentary herbivore populations from control. Manipulative field experiments conducted with the arthropod community found on papaya, Carica papaya, provided support for the first two predictions: (1) the widely foraging predatory mite Phytoseiulus macropilis strongly suppressed populations of a sedentary herbivore, the spider mite Tetranychus cinnabarinus, whereas (2) the tangle-web spider Nesticodes rufipes, a classic sit and wait predator, failed to suppress Tetranychus population growth rates. However, our experiments provided no support for the third hypothesis; the sit and wait predator Nesticodes did not disrupt the suppression of Tetranychus populations by Phytoseiulus. This contrasts with an earlier study that demonstrated that Nesticodes can disrupt control of Tetranychus generated by another widely foraging predator, Stethorus siphonulus. Behavioral observations suggested a simple explanation for the differing sensitivity of Phytoseiulus and Stethorus to Nesticodes predation. Phytoseiulus is a much smaller predator than Stethorus, has a lower rate of prey consumption, and thus has a much smaller requirement to forage across the leaf surface for prey, thereby reducing its probability of encountering Nesticodes webs. Small body size may be a general means by which widely foraging intermediate predators can ameliorate their risk of predation by sit and wait top predators. This effect may partially or fully offset the general expectation from size-structured trophic interactions that smaller predators are subject to more intense intraguild predation. PMID- 15278425 TI - Bush selection along foraging pathways by sympatric impala and greater kudu. AB - In order to identify the selection mechanism of two sympatric African browsers, we analysed encounter rates and selection of bushes along foraging pathways. We monitored the tracks, left overnight, by kudu and impala on an experimental plot of natural Acacia nilotica and Dichrostachys cinerea in the highveld of Zimbabwe, and recorded the number of bushes attacked in each category. Both ungulates were selective for the bush categories, but kudu were consistently more selective than impala, and showed a higher preference for the larger A. nilotica and D. cinerea bushes, which had a significantly greater number of bites which were not reachable by impala. For both kudu and impala, the probability of attacking larger bushes increased significantly with the proportion of large bushes encountered along the foraging pathways, whereas the consumption of smaller bushes was apparently unpredictable. For the most abundant food item (medium D. cinerea), the probability of attack by impala along a pathway decreased with increasing proportions of larger bushes in the experimental area, but was also dependent on impala group size and season. In addition, we found that encounter rates with larger bushes were significantly higher for kudu than for impala. Experimentally reducing the availability of the larger bushes had little effect on both impala and kudu during the following rainy season. However, during the following cool dry season, kudu showed an increased selectivity with a strong preference for the remaining large bushes (large A. nilotica), followed by a sharp decrease in selectivity in the hot dry season when they also fed from significant numbers of medium trees. Impala had little reaction to the experimental changes in the availability of bush categories in either season. We suggest that both kudu and impala selected bushes on the basis of the potential number of bites they can provide, and this resulted in different search strategies. Kudu focussed on the larger bushes which have a larger number of twigs which are out of reach of impala and kudu also probably directed their path preferentially towards the few larger bushes to maximize encounter rates with this favoured bush category. These differences in bush selection process lead to a low overlap in resource use between the two browsers in this type of savanna. PMID- 15278426 TI - Zebra mussels affect benthic predator foraging success and habitat choice on soft sediments. AB - The introduction of zebra mussels ( Dreissena spp.) to North America has resulted in dramatic changes to the complexity of benthic habitats. Changes in habitat complexity may have profound effects on predator-prey interactions in aquatic communities. Increased habitat complexity may affect prey and predator dynamics by reducing encounter rates and foraging success. Zebra mussels form thick contiguous colonies on both hard and soft substrates. While the colonization of substrata by zebra mussels has generally resulted in an increase in both the abundance and diversity of benthic invertebrate communities, it is not well known how these changes affect the foraging efficiencies of predators that prey on benthic invertebrates. We examined the effect of zebra mussels on the foraging success of four benthic predators with diverse prey-detection modalities that commonly forage in soft substrates: slimy sculpin ( Cottus cognatus), brown bullhead ( Ameirus nebulosus), log perch ( Percina caprodes), and crayfish ( Orconectes propinquus). We conducted laboratory experiments to assess the impact of zebra mussels on the foraging success of predators using a variety of prey species. We also examined habitat use by each predator over different time periods. Zebra mussel colonization of soft sediments significantly reduced the foraging efficiencies of all predators. However, the effect was dependent upon prey type. All four predators spent more time in zebra mussel habitat than in either gravel or bare sand. The overall effect of zebra mussels on benthic feeding fishes is likely to involve a trade-off between the advantages of increased density of some prey types balanced against the reduction in foraging success resulting from potential refugia offered in the complex habitat created by zebra mussels. PMID- 15278427 TI - Field transplants reveal summer constraints on a butterfly range expansion. AB - The geographic ranges of most species are expected to shift to higher elevations and latitudes in response to global warming. But species react to specific environmental changes in individualistic ways, and we are far from a detailed understanding of range-shifts. Summer temperature often limits the ranges of insects and plants, so many range-shifts are expected to track summer warming. I explore this potential range-limiting factor in a case study of a northwardly expanding American butterfly, Atalopedes campestris (Lepidoptera, Hesperiidae). This species has recently colonized the Pacific Northwest, USA, where the mean annual temperature has risen 0.8-1.8 degrees C over the past 100 years. Using field transplant experiments across the current range edge, I measured development time, survivorship, fecundity and predation rates along a naturally occurring thermal gradient of 3 degrees C. Development time was significantly slower outside the current range in eastern Washington (WA), as expected because of cooler temperatures there. Slower development would reduce the number of generations possible per year outside the current range, dramatically lowering the probability that a population could survive there. Differences in survivorship, fecundity and predation rate across the range edge were not significant. The interaction between summer and winter temperature appears to be crucial in defining the current range limit. The estimated difference in temperature required to affect the number of generations is greater than the extent of summer warming observed over the past century, however, and thus historically winter temperature alone probably limited the range in southeastern WA. Nonetheless, extraordinarily warm summers may have improved colonization success, increasing the probability of a range expansion. These results suggest that extreme climatic events may influence rates of response to long-term climate change. They also demonstrate that range-limiting factors can change over time, and that the asymmetry in seasonal warming trends will have biological consequences. PMID- 15278428 TI - A comparison of morphological and chemical fruit traits between two sites with different frugivore assemblages. AB - Large-scale comparisons might reveal matching between fruit traits and frugivore assemblages that might be cryptic on a local scale. Therefore, we compared morphological (colour, size, husk thickness) and chemical fruit traits (protein, nitrogen, sugar, lipid, tannin and fibre content) between Malagasy and South African tree communities with different frugivore communities. In Madagascar, where lemurs are important seed dispersers, we found more tree species with fruit colour classified as "primate fruits". In contrast, in South Africa we found more tree species with fruits classified as "bird coloured". This correlated with the greater importance of frugivorous birds in South Africa vs. Madagascar. Additionally, we found higher sugar concentrations in fruits from the South African tree community and higher fibre content in fruits from the Malagasy tree community. However, fibre content could be related to differences in abiotic conditions between the two study sites. This suggests that fruit colour more than other morphological and chemical fruit traits, reflects food selection by the different frugivore assemblages of those two sites. PMID- 15278429 TI - El Nino droughts and their effects on tree species composition and diversity in tropical rain forests. AB - In this study I investigated the effects of the extreme, 1997/98 El Nino related drought on tree mortality and understorey light conditions of logged and unlogged tropical rain forest in the Indonesian province of East Kalimantan (Borneo). My objectives were to test (1) whether drought had a significant effect on tree mortality and understorey light conditions, (2) whether this effect was greater in logged than in undisturbed forest, (3) if the expected change in tree mortality and light conditions had an effect on Macaranga pioneer seedling and sapling densities, and (4) which (a)biotic factors influenced tree mortality during the drought. The 1997/1998 drought led to an additional tree mortality of 11.2, 18.1, and 22.7% in undisturbed, old logged and recently logged forest, respectively. Mortality was highest in logged forests, due to extremely high mortality of pioneer Macaranga trees (65.4%). Canopy openness was significantly higher during the drought than during the non-drought year (6.0, 8.6 and 10.4 vs 3.7, 3.8 and 3.7 in undisturbed, old logged and recently logged forest, respectively) and was positively correlated with the number of dead standing trees. The increase in light in the understorey was accompanied by a 30 to 300 fold increase in pioneer Macaranga seedling densities. Factors affecting tree mortality during drought were (1) tree species successional status, (2) tree size, and (3) tree location with respect to soil moisture. Tree density and basal area per surface unit had no influence on tree mortality during drought. The results of this study show that extreme droughts, such as those associated with El Nino events, can affect the tree species composition and diversity of tropical forests in two ways: (1) by disproportionate mortality of certain tree species groups and tree size classes, and (2) by changing the light environment in the forest understorey, thereby affecting the recruitment and growth conditions of small and immature trees. PMID- 15278430 TI - Aphid effects on rhizosphere microorganisms and microfauna depend more on barley growth phase than on soil fertilization. AB - This paper gives the first reports on aphid effects on rhizosphere organisms as influenced by soil nutrient status and plant development. Barley plants grown in pots fertilized with N but without P (N), with N and P (NP), or not fertilized (0) were sampled in the early growth phase (day 25), 1 week before and 1 week after spike emergence. Aphids were added 16 days before sampling was carried out. In a separate experiment belowground respiration was measured on N and NP fertilized plant-soil systems with aphid treatments comparable to the first experiment. Aphids reduced numbers of rhizosphere bacteria and fungal feeding nematodes 1 week before spike emergence. Before spike emergence, aphids reduced belowground respiration in NP treatments. These findings strongly indicate that aphids reduced allocation of photoassimilates to roots and deposition of root exudates in the growth phase of the plant. Contrary to this, 1 week after spike emergence numbers of bacteria, fungal feeding nematodes and Protozoa were higher in rhizospheres of plants subjected to aphids probably because aphids enhanced root mortality and root decomposition. Protozoa and bacterial feeding nematodes were stimulated at different experimental conditions with nematodes being the dominant bacterial grazers at N fertilization and Protozoa in the NP treatment before spike emergence. PMID- 15278431 TI - Effects of behavioral and morphological plasticity on risk of predation in a Neotropical tadpole. AB - Predator-induced phenotypic plasticity is widespread among aquatic animals, however the relative contributions of behavioral and morphological shifts to reducing risk of predation remain uncertain. We tested the phenotypic plasticity of a Neotropical tadpole ( Rana palmipes) in response to chemical cues from predatory Belostoma water bugs, and how phenotype affects risk of predation. Behavior, morphology, and pigmentation all were plastic, resulting in a predator induced phenotype with lower activity, deeper tail fin and muscle, and darker pigmentation. Tadpoles in the predator cue treatment also grew more rapidly, possibly as a result of the nutrient subsidy from feeding the caged predator. For comparison to phenotypes induced in the experiment, we quantified the phenotype of tadpoles from a natural pool. Wild-caught tadpoles did not match either experimentally induced phenotype; their morphology was more similar to that produced in the control treatment, but their low swimming activity was similar to that induced by predator cues. Exposure of tadpoles from both experimental treatments and the natural pool to a free-ranging predator confirmed that predator-induced phenotypic plasticity reduces risk of predation. Risk of predation was comparable among wild-caught and predator-induced tadpoles, indicating that behavioral shifts can substantially alleviate risk in tadpoles that lack the typical suite of predator-induced morphological traits. The morphology observed in wild-caught tadpoles is associated with rapid growth and high competition in other tadpole species, suggesting that tadpoles may profitably combine a morphology suited to competition for food with behaviors that minimize risk of predation. PMID- 15278432 TI - Root foraging for patchy resources in eight herbaceous plant species. AB - The root foraging strategy of a plant species can be characterized by measuring foraging scale, precision, and rate. Trade-offs among these traits have been predicted to contribute to coexistence of competitors. We tested for trade-offs among root foraging scale (total root mass and length of structural roots), precision (ln-ratio of root lengths in resource-rich and resource-poor patches), and rate (days required for roots to reach a resource-rich patch, or growth rate of roots within a resource-rich patch) in eight co-occurring species. We found that root foraging scale and precision were positively correlated, as were foraging scale and the rate of reaching patches. High relative growth rate of a species did not contribute to greater scale, precision, or rate of root foraging. Introduced species had greater foraging scale, precision, and rate than native species. The positive correlations between foraging scale and foraging precision and rate may give larger species a disproportionate advantage in competition for patchy soil resources, leading to size asymmetric competition below ground. PMID- 15278434 TI - Immunolocation of proteoglycans and bone-related noncollagenous glycoproteins in developing acellular cementum of rat molars. AB - To elucidate the roles of proteoglycans of (PGs), bone sialoprotein (BSP), and osteopontin (OPN) in cementogenesis, their distribution was investigated in developing and established acellular cementum of rat molars by an immunoperoxidase method. To characterize PGs, antibodies against five species of glycosaminoglycans (GAGS), chondroitin-4-sulfate (C4S), chondroitin-6-sulfate (C6S), unsulfated chondroitin (C0S), dermatan sulfate (DS), and keratan sulfate (KS) were used. Routine histological staining was also applied. With onset of dentin mineralization, the initial cementum appeared on the dentin surface as a hematoxylin-stained fibril-poor layer. Subsequently, primitive principal fibers attached to the initial cementum. As the acellular cementum containing extrinsic fibers covered the initial cementum, the intal cementum formed the cemento dentinal junction. Following immunohistochemistry at the earliest time of cementogenesis, the initial cementum was intensely immunoreactive for C4S, C6S, C0S, BSP, and OPN. After the initial cementum was embedded, neither the cemento dentinal junction nor the cementum was immunoreactive for any GAG species. However, the cementum was immunoreactive for any GAG species. However, the cementum and cemento-dentinal were consistently immunoreactive for BSP. Although the cemento-dentinal junction was consistently immunoreactive for OPN, the remaining cementum showed no significant immunoreactivity. Thus, initial acellular cementogenesis requires a dense accumulation of PGs, BSP, and OPN, which may be associated with the mineralization process independently of collagen fibrils and initial principal fiber attachment. PMID- 15278435 TI - Association between evolutionary history of angiotensinogen haplotypes and plasma levels. AB - Over the last decade, considerable effort has been invested in studying the associations between angiotensinogen (AGT) variants, AGT plasma levels and high blood pressure. Evidence accumulated to date consistently supports the relationship between the AGT locus and the protein level, while an influence on blood pressure has been difficult to establish; in both instances the predisposing molecular variants are not fully defined. An evolutionary approach, taking into account the phylogenetic relationship between all the polymorphisms at this locus, may improve our understanding of the genetic nature of these quantitative phenotypes. Accordingly we sequenced a 6.8 kb region of the AGT gene in 57 Nigerian individuals (29 with high AGT plasma levels and 28 with low AGT plasma levels). Haplotypes were grouped into seven major haplogroups and their phylogenetic relationship was established. The association between haplogroups and AGT plasma levels was investigated. A significant linear correlation was detected between haplogroup genetic distance and AGT levels, suggesting a nonrandom accumulation of risk-associated mutations during the evolutionary history of the AGT gene. PMID- 15278436 TI - Variations in the C3, C3a receptor, and C5 genes affect susceptibility to bronchial asthma. AB - Bronchial asthma (BA) is a common chronic inflammatory disease characterized by hyperresponsive airways, excess mucus production, eosinophil activation, and the production of IgE. The complement system plays an immunoregulatory role at the interface of innate and acquired immunities. Recent studies have provided evidence that C3, C3a receptor, and C5 are linked to airway hyperresponsiveness. To determine whether genetic variations in the genes of the complement system affect susceptibility to BA, we screened single nucleotide polymorphisms (SNPs) in C3, C5, the C3a receptor gene (C3AR1), and the C5a receptor gene (C5R1) and performed association studies in the Japanese population. The results of this SNP case-control study suggested an association between 4896C/T in the C3 gene and atopic childhood BA (P = 0.0078) as well as adult BA (P = 0.010). When patient data were stratified according to elevated total IgE levels, 4896C/T was more closely associated with adult BA (P = 0.0016). A patient-only association study suggested that severity of childhood BA was associated with 1526G/A of the C3AR1 gene (P = 0.0057). We identified a high-risk haplotype of the C3 gene for childhood (P = 0.0021) and adult BA (P = 0.0058) and a low-risk haplotype for adult BA (P = 0.00011). We also identified a haplotype of the C5 gene that was protective against childhood BA (P = 1.4 x 10(-6)) and adult BA (P = 0.00063). These results suggest that the C3 and C5 pathways of the complement system play important roles in the pathogenesis of BA and that polymorphisms of these genes affect susceptibility to BA. PMID- 15278437 TI - Scd1ab-Xyk: a new asebia allele characterized by a CCC trinucleotide insertion in exon 5 of the stearoyl-CoA desaturase 1 gene in mouse. AB - We describe here a spontaneous, autosomal recessive mutant mouse suffering from skin and hair defects, which arose in the outbred Kunming strain. By haplotype analysis and direct sequencing of PCR products, we show that this mutation is a new allele of the asebia locus with a naturally occurring mutation in the Scd1 gene (a CCC insertion at nucleotide position 835 in exon 5), which codes for stearoyl-CoA desaturase 1. This mutation introduces an extra proline residue at position 279 in the Scd1 protein. The mutant mice, originally designated km/km but now assigned the name Scd1ab-Xyk (hereafter abbreviated as abXyk/abXyk), have a similar gross and histological phenotype to that reported for previously characterized allelic asebia mutations (Scd1ab, Scd1abJ, Scd1ab2J, and Scd1tm1Ntam). Histological analysis showed they were also characterized by hypoplasic sebaceous glands and abnormal hair follicles. In a cross between Kunming- abXyk/abXyk and ABJ/Le-abJ/abJ mice, all the progeny showed the same phenotype, indicating that the two mutations were non-complementing and therefore allelic. Comparisons with the other four allelic mutants indicate that the Scd1ab Xyk mutation causes the mildest change in Scd1 function. This new mouse mutant is a good model not only for the study of scarring alopecias in humans, which are characterized by hypoplasic sebaceous glands, but also for studying the structure and function of the Scd1 protein. PMID- 15278438 TI - Nematode parasites and scrapie: experiments in sheep and mice. AB - To demonstrate the possible role of nematode parasites in the modification of host susceptibility to scrapie, experiments were conducted using sheep naturally exposed to scrapie, chosen by their genotype at the PrP gene, and infected with Teladorsagia circumcincta. Two 4-year duration experiments demonstrated that the nematode infection shortened the development of scrapie with a significant regression between the level of infection and age at first scrapie symptoms (P < 0.006). Investigations by ELISA tests in different species of nematode parasites of the digestive tract collected from scrapie infected ewes did not reveal the presence of PrPSc. In scrapie-infected C57BL mice, infected or not with Heligmosoides polygyrus at various times, parasitized animals showed a slight but significantly longer survival period. Assays on transmission by the larvae hatching from eggs collected from scrapie-infected mice were unsuccessful. We concluded that nematodes modify host susceptibility to scrapie, but their role in the horizontal transmission of the disease was not demonstrated. PMID- 15278439 TI - The life cycle of Anisakis simplex in the Norwegian Deep (northern North Sea). AB - Copepoda (Calanus finmarchicus n = 1,722, Paraeuchaeta norvegica n = 1,955), Hyperiidae (n = 3,019), Euphausiacea (Meganyctiphanes norvegica n = 4,780), and the fishes Maurolicus muelleri (n = 500) and Pollachius virens (n = 33) were collected in the Norwegian Deep (northern North Sea) during summer 2001 to examine the importance of pelagic invertebrates and vertebrates as hosts of Anisakis simplex and their roles in the transfer of this nematode to its final hosts (Cetaceans). Third stage larvae (L3) of A. simplex were found in P. norvegica, M. muelleri and P. virens. The prevalence of A. simplex in dissected P. norvegica was 0.26%, with an intensity of 1. Prevalences in M. muelleri and P. virens were 49.6% and 100.0%, with mean intensities of 1.1-2.6 (total fish length >or=6.0-7.2) and 193.6, respectively. All specimens of C. finmarchicus and M. norvegica examined were free of anisakid nematode species and no other parasites were detected. P. norvegica, which harboured the third stage larvae, is the obligatory first intermediate host of A. simplex in the investigated area. Though there was no apparent development of larvae in M. muelleri, this fish can be considered as the obligatory second intermediate host of A. simplex in the Norwegian Deep. However, it is unlikely that the larva from P. norvegica can be successfully transmitted into the cetacean or pinniped final hosts, where they reach the adult stage. An additional growth phase and a second intermediate host is the next phase in the life cycle. Larger predators such as P. virens serve as paratenic hosts, accumulating the already infective stage from M. muelleri. The oceanic life cycle of A. simplex in the Norwegian Deep is very different in terms of hosts and proposed life cycle patterns of A. simplex from other regions, involving only a few intermediate hosts. In contrast to earlier suggestions, euphausiids have no importance at all for the successful transmission of A. simplex in the Norwegian Deep. This demonstrates that this nematode is able to select definite host species depending on the locality, apparently having a very low level of host specificity. This could explain the wide range of different hosts that have been recorded for this species, and can be seen as the reason for the success of this parasite in reaching its marine mammal final hosts in an oceanic environment. PMID- 15278440 TI - The efficacy of inhibitors involved in spermidine metabolism in Plasmodium falciparum, Anopheles stephensi and Trypanosoma evansi. AB - In the present study, we have tested the effect of different polyamine inhibitors of the spermidine metabolizing enzymes deoxyhypusine synthase and homospermidine synthase in different chloroquine resistant Plasmodium falciparum strains, in the mosquito Anopheles stephensi (Diptera: Culicidae) and in a Trypanosoma evansi clone I from strain STIB 806 K China. Recent experiments have shown that agmatine is a growth inhibitor of the malaria parasite P. falciparum (Kaiser et al. 2001) in vitro. A comparison of agmatine efficacy with the new antimalarials artemisinin, triclosan and conventional chloroquine showed similar or even better results on the basis of growth inhibition and the reduction of developmental forms. However, no effect of triclosan or agmatine was observed at the ribonucleic acid level. In a second set of experiments, we tested the effect of 1,7-diaminoheptane and agmatine on oocyst formation in A. stephensi after infection with Plasmodium yoelii. Agmatine had an antisporozoite effect since 1,000 microM led to a 59.5% inhibition of oocysts. A much weaker inhibitor of oocyst formation was 1,7-diaminoheptane. The most effective in in vitro inhibition of T. evansi was dicyclohexylamine, an inhibitor of spermidine biosynthesis with an IC(50 ) value of 47.44 microM and the deoxyhypusine inhibitor 1,7-diaminoheptane with an IC(50) value of 47.80 microM. However, both drugs were ineffective in in vivo experiments in a Trypanosoma mouse model. Two different spermidine analogues, 1,8-diaminooctane and 1,3-diaminopropane with IC(50) values of 171 microM and 181.37 microM, respectively, were moderate inhibitors in vitro and ineffective in vivo. PMID- 15278441 TI - The phylogeny of diphyllobothriid tapeworms (Cestoda: Pseudophyllidea) based on ITS-2 rDNA sequences. AB - Phylogenetic analysis of sequences of the ITS-2 rRNA genes of 20 samples of pseudophyllidean cestodes of the family Diphyllobothriidae (Ligula, Digramma, Diphyllobothrium, and Schistocephalus) from different hosts and geographical regions revealed that: (1) the inclusion of ligulids (previously family Ligulidae) to the Diphyllobothriidae is correct; (2) Schistocephalus appears as the most basal taxon of the Diphyllobothriidae, well separated from Ligula and Digramma, thus supporting the validity of Schistocephalinae Dubinina, 1962; (3) Digramma belonged with samples of Ligula, thus suggesting its invalidity as a genus; and (4) isolates of Ligula, presumably belonging to Ligula intestinalis, are paraphyletic, indicating that this species may represent a complex of separate taxa. Our results indicate the necessity for a taxonomic revision of the family Diphyllobothriidae. PMID- 15278442 TI - Duffy antigen is important for the lethal effect of the lethal strain of Plasmodium yoelii 17XL. AB - We studied the potential role of the Duffy antigen and glycophorin A as receptors for rodent malaria parasite invasion of erythrocytes. Parasitemia increased exponentially after infection with Plasmodium berghei NK65, P. chabaudi, and P. vinckei in Duffy antigen knockout, glycophorin A knockout, and wild-type mice, indicating that the Duffy antigen and glycophorin A are not essential for these malaria parasites. However, parasitemia of the Duffy antigen knockout mice infected with P. yoelii 17XL remained constant from day 5 to 14 after infection, and then decreased, resulting in autotherapy. The treatment of P. yoelii 17XL infected Duffy antigen knockout mice with anti-CD4 antibody increased the parasitemia 15 days after infection and the mice eventually died, indicating that CD-4-positive cells play an important role in the clearance of P. yoelii 17XL at the late stage of the infection. PMID- 15278443 TI - The efficacy of enrofloxacin, alone or combined with metronidazole, in the therapy of canine leishmaniasis. AB - We evaluated the efficacy of enrofloxacin, alone or combined with metronidazole, against Leishmania infantum. The in vitro activity of this fluoroquinolone was assessed using two different methods: a direct test aimed at assessing the drug activity on the parasite, and an indirect test aimed at evaluating the drug effect on macrophage killing, lymphomonocyte activation and nitric oxide production. An in vivo test was also performed on 36 dogs with leishmaniasis, subdivided into three groups, one treated with enrofloxacin, another with enrofloxacin plus metronidazole, and a control group with meglumine antimoniate. The direct test did not show any action of enrofloxacin on the parasite, while the indirect testing showed an enhancement of macrophage killing and an increase in nitric oxide production. These findings show that enrofloxacin does not exert a direct anti-leishmanial activity in vitro. However, on the basis of the positive immunostimulation results shown in vitro and the clinical improvement, particularly of the cutaneous lesions, obtained in several dogs in the in vivo trial, the use of enrofloxacin in association with a specific anti-leishmanial drug can be proposed in the therapeutic protocol of canine leishmaniasis. PMID- 15278444 TI - Soboliphyme occidentalis sp. nov. (Nematoda, Soboliphymidae) from the Iberian mole Talpa occidentalis (Insectivora, Talpidae) in Spain. AB - A new species of Soboliphyme from the endemic Iberian mole (Talpa occidentalis) is described. Soboliphyme occidentalis sp. nov. can be readily distinguished from all of its congeners primarily by the position of the vulva, which clearly shows a posterior oesophageal location, and the number of male caudal papillae. S. occidentalis sp. nov. is the only species that has four pairs of caudal papillae. S. abei, S. caucasica and S. jamesoni can be distinguished from S. occidentalis sp. nov. by not having a notched sucker, the anterior position of the vulva and two polar plugs in the eggs. S. jamesoni has an armate oral sucker and longer spicule; S. caucasica a longer spicule and shorter eggs, and S. abei has shorter eggs, which separate these species from S. occidentalis sp. nov. In the rest of the species with a notched oral sucker, S. baturini and S. hirudiniformis are differentiated from S. occidentalis sp. nov. by the anterior position of the vulva, two polar plugs in the egg and the spicule length in S. baturini and S. hirudiniformis and the size of eggs in S. baturini and S. hirudiniformis. S. ataahai, S. soricis and S. urotrichi have the vulva at the oesophago-intestinal junction, 9-10 male caudal papillae (S. ataahai and S. urotrichi), absence of male caudal papillae (S. soricis), armate oral sucker and long spicule in S. ataahai and one row of six circumoral spines in S. urotrichi. A key to the species of Soboliphyme is presented. PMID- 15278445 TI - A preliminary investigation on the infectivity of Trichinella larvae in traditional preparations of walrus meat. AB - This study evaluated the infectivity of Trichinella nativa in freshly frozen walrus meat and traditionally aged walrus meat (igunaq) associated with two human outbreaks of trichinellosis in the Canadian Arctic. Trichinella larvae recovered from walrus meat stored at -20 degrees C for up to 20 months remained infective for guinea pigs inoculated with 135 or 716 larval doses. However, none of the 4-5 and 10-month-old igunaq preparations contained infective T. nativa larvae as measured by bioassays using mice and guinea pigs at inoculation doses ranging from 6 to 500 larvae. This indicates that the degradation process that occurred in the field can be sufficient to either kill Trichinella larvae or render them non-infective for mice and guinea pigs. Further research is needed to evaluate the food safety risk of traditional walrus igunaq aged under different field conditions and storage times. PMID- 15278446 TI - The effects of the ectoparasite Tracheliastes polycolpus (Copepoda: Lernaeopodidae) on the fins of rostrum dace (Leuciscus leuciscus burdigalensis). AB - Rostrum dace (Leuciscus leuciscus burdigalensis) from the River Viaur were found to be infested with the ectoparasite Tracheliastes polycolpus (Copepoda: Lernaeopodidae). Samples from five study sites along the river revealed different patterns of parasite infestation. Heavily infested fish were found at the upper study sites whereas much lower infestation levels were observed at the lower study sites. The copepods showed an aggregated dispersion pattern on host fins. The results showed significantly preferred microhabitats, with adult females being more abundant on the anal, pelvic and along the external part of the pectoral fins. The anal and pelvic fins were damaged by the parasite with a loss of their surface area. These fin alterations may reduce the fish's swimming ability and therefore affect the rostrum dace population. Our findings highlight the need to study the effects of parasites on stream fish populations. PMID- 15278448 TI - Anti-hepatitis B surface antigen IgG1 subclass is predominant in individuals who have recovered from hepatitis B virus infection, chronic carriers, and vaccinees. AB - Patterns of each IgG-specific subclass for hepatitis B virus (HBV) core antigen (anti-HBc) are remarkably different among individuals with different infection status, i.e., completely recovered or chronic carrier. Each of the IgG-specific subclasses of HBV surface antigen (anti-HBs) was tested for ELISA sensitivity using four commercially available hepatitis B surface antigen (HBsAg) kits and one self-prepared plate. The specificity in 18 serum samples obtained from chronic HBV carriers, recovered individuals, vaccinees and non-infected individuals was investigated. Differences in absorbance values were obtained by comparing results from these different plates. Data on the absorbance values of anti-HBs IgG subclasses obtained indicated that one to four subjects had a false negative or false-positive result using the four commercial plates. Only the self prepared plate demonstrated 100% specificity and sensitivity for anti-HBs subclasses. Moreover, the results indicate that anti-HBs subclass IgG1 was predominant in cured patients, chronic carriers and vaccinees. The samples from both chronic carriers and vaccinees exhibited a significantly higher concentration of total IgG and IgG1 than samples in recovered individuals (P<0.05). PMID- 15278449 TI - Metachronous multicentric giant-cell tumor of the bone in the lower limb. Case report and Ki-67 immunohistochemistry study. AB - Multicentric giant-cell tumors of the bone (GCTs) are rare. Little is known about the mechanisms by which these tumors spread and how 1% of GCT turn out to be multicentric. We report the case of a 19-year-old woman with metachronous multiple and recurrent GCTs that were unusual in their pattern of progression along the right lower limb over a 23-year period. Histology showed no evidence of malignant transformation. The treatment was repeated curettage and packing with cement. This did not permit a wide surgical margin, but avoided amputation and preserved full limb function. We tested the proliferation index marker Ki-67 in the tumor specimens. Ki-67 expression was limited to the mononuclear cell component of the tumors. The proliferation index was similar in each new tumor and higher in recurrences for each location. In this case, proliferation was initially low in the new tumor location, despite the time difference and independent from the initial clone evolution. Proliferation index increased in recurrent GCTs after marginal margin resection. PMID- 15278450 TI - Generalized crystal-storing histiocytosis as a presentation of multiple myeloma: a case with a possible pro-aggregation defect in the immunoglobulin heavy chain. AB - Crystal-storing histiocytosis (CSH) with massive accumulation of particulate immunoglobulins is a rare phenomenon accompanying B-cell dyscrasias. In the reported case (M51), the disease presented as systemic CSH and later was proved to be a frank multiple myeloma. The aggregates of crystal-laden histiocytes were demonstrated in the bone marrow, lungs, kidney, and liver. Additionally, the crystalline immunoglobulin particles were identified in renal stromal cells and in hepatocytes. The patient developed lung adenocarcinoma and died 12 months after the presentation, shortly after the lobectomy. In this paper, we report the results of morphological (including electron microscopy), immunohistochemical, and biochemical analysis. The tendency for aggregation of the IgG kappa monoclonal protein was due to the abnormal physicochemical properties of its heavy chain. Massive accumulation of crystal-storing histiocytes surpassed the myeloma tumor burden and markedly contributed to the severity of the disease. PMID- 15278451 TI - Neurogenesis in the chilopod Lithobius forficatus suggests more similarities to chelicerates than to insects. AB - In a recent comparative study on neurogenesis in the diplopod Glomeris marginata we have shown that the millipede and the spider share several features that cannot be found in homologous form in insects and crustaceans. The most distinctive difference is that groups of neural precursors are singled out from the neuroectoderm of the spider and the diplopod, rather than individual cells (i.e. neuroblasts) as in insects or crustacean. This observation constitutes the first morphological indication for a close myriapod/chelicerate relationship that has otherwise only been suggested by molecular phylogenetic analysis. To see whether the pattern of neurogenesis described for the diplopod is representative for myriapods, we analysed neurogenesis in the basal chilopod Lithobius forficatus. We show here that groups of cells invaginate from the chilopod neuroectoderm at strikingly similar positions as the invaginating cell groups of the diplopod and the spider. Furthermore, the expression patterns of the proneural and neurogenic genes reveal more similarities to the chelicerate and the diplopod than to insects. Thus, chelicerates and myriapods share the developmental mechanism for neurogenesis, either because they are true sister groups, or because this reflects the ancestral state of neurogenesis in arthropods. PMID- 15278452 TI - Identification of regulatory modules mediating specific expression of the roughest gene in Drosophila melanogaster. AB - Roughest (Rst) is a cell adhesion molecule of the immunoglobulin superfamily with pleiotropic functions during the development of Drosophila melanogaster. It has been shown to be involved in cell sorting before apoptosis in the developing compound eye, in fusion processes of embryonic muscle development and in axonal pathfinding. In accordance with its multiple functions, the rst gene shows a dynamic expression pattern throughout the development of Drosophila. In order to understand the transcriptional regulation of rst expression we have identified rst cis regulatory sequences in an enhancer detection screen. By dissection of the identified rst cis regulatory sequences we identified several distinct rst regulatory modules. Among others these include elements for expression in interommatidial cells of the pupal eye disc at a time when apoptotic decisions are made in these cells and elements for expression in the embryonic mesoderm. The expression of rst in the embryonic mesoderm is regulated by at least two separate modules. PMID- 15278453 TI - A single amino acid substitution in soybean VSPalpha increases its acid phosphatase activity nearly 20-fold. AB - Soybean [Glycine max (L.) Merr.] contains two proteins called vegetative storage proteins (VSPs) that function as temporary storage reserves, but are also closely related to plant acid phosphatases of the haloacid dehalogenase (HAD) superfamily. This study examined the biochemical basis for the relatively low catalytic activity previously reported for these VSPs. The specific activity of purified recombinant VSPalpha on GMP was about 40-fold lower than for a related soybean root nodule acid phosphatase (APase), which had a specific activity of 845 U mg(-1) protein. Conversion of Ser106 to Asp increased VSPalpha activity about 20-fold. This Asp residue is present in nodule APase and is a highly conserved nucleophile in the HAD superfamily. Related VSPs from cultivated soybean and from three wild perennial soybeans, as well as a pod storage protein (PSP) from Phaseolus vulgaris L. all lack the catalytic Asp, suggesting they too are catalytically inefficient. Phylogenetic analysis showed the VSPs and PSP are more closely related to each other than to 21 other VSP-like proteins from several plant species, all of which have the nucleophilic Asp. This study suggests that loss of catalytic activity may be a requirement for the VSPs and PSP to function as storage proteins in legumes. PMID- 15278454 TI - Cloning, characterization and expression of OsGLN2, a rice endo-1,3-beta glucanase gene regulated developmentally in flowers and hormonally in germinating seeds. AB - We report here the isolation and characterization of a new endo-1,3-beta glucanase (1,3-beta-GLU) cDNA, OsGLN2, that is expressed both in flowers and in germinating seeds of rice (Oryza sativa L.). The isolated OsGLN2 gene encoded a protein which displayed 72%, 93% and 92% identity at the amino acid level with those encoded by barley GII, rice Gns4 and glu1 1,3-beta-GLU genes, respectively. A GST-OsGLN2 recombinant protein expressed in Escherichia coli preferentially hydrolyzed Laminaria digitata 1,3;1,6-beta-glucan and liberated only oligosaccharides, suggesting that the enzyme can be classified as a 1,3-beta-GLU. Northern analysis with a 3'-UTR gene-specific probe revealed that OsGLN2 is expressed exclusively in the paleae and lemmas during flowering, and no expression of OsGLN2 was detected in other tissues such as leaf blades, leaf sheaths, stems, nodes and roots in mature rice plants. The OsGLN2 gene is also expressed in germinating seeds, where its expression is predominant in endosperms rather than embryos. In de-embryonated rice half-seeds, addition of gibberellin A3 (GA) greatly enhanced expression of the OsGLN2 gene, while the GA-induced gene expression was suppressed strongly by abscisic acid (ABA). This is the first report, to our knowledge, that OsGLN2 encodes a 1,3-beta-GLU and is expressed specifically in paleae and lemmas during flowering and in germinating seeds, where its expression is enhanced by GA and suppressed by ABA. PMID- 15278455 TI - Localization and function of SulP, a nuclear-encoded chloroplast sulfate permease in Chlamydomonas reinhardtii. AB - Recent work [H.-C. Chen et al. (2003) Planta 218:98-106] reported on the genomic, proteomic, phylogenetic and evolutionary aspects of a putative nuclear gene ( SulP) encoding a chloroplast sulfate permease in the model green alga Chlamydomonas reinhardtii. In this article, evidence is provided for the envelope localization of the SulP protein and its function in the uptake and assimilation of sulfate by the chloroplast. Localization of the SulP protein in the chloroplast envelope was concluded upon isolation of C. reinhardtii chloroplasts, followed by fractionation into envelope and thylakoid membranes and Western blotting of these fractions with specific polyclonal antibodies raised against the recombinant SulP protein. The function of the SulP protein was probed in antisense transformants of C. reinhardtii having lower expression levels of the SulP gene. Results showed that cellular sulfate uptake capacity was lowered as a consequence of attenuated SulP gene expression in the cell, directly affecting rates of de novo protein biosynthesis in the chloroplast. The antisense transformants exhibited phenotypes of sulfate-deprived cells, displaying slow rates of light-saturated oxygen evolution, low levels of Rubisco in the chloroplast and low steady-state levels of the photosystem-II D1 reaction-center protein. The role of the chloroplast sulfate transport in the uptake and assimilation of sulfate in C. reinhardtii is discussed along with its impact on the repair of photosystem-II from a frequently occurring photo-oxidative damage and potential use for the elucidation of the H(2)-evolution-related metabolism in this green alga. PMID- 15278456 TI - The type-1 and type-2 ribosome-inactivating proteins from Iris confer transgenic tobacco plants local but not systemic protection against viruses. AB - The antiviral activity of the type-2 ribosome-inactivating protein (RIP) IRAb from Iris was analyzed by expressing IRAb in tobacco (Nicotiana tabacum L. cv. Samsun NN) plants and challenging the transgenic plants with tobacco mosaic virus (TMV). Although constitutive expression of IRAb resulted in an aberrant phenotype, the plants were fertile. Transgenic tobacco lines expressing IRAb showed a dose-dependent enhanced resistance against TMV infection but the level of protection was markedly lower than in plants expressing IRIP, the type-1 RIP from Iris that closely resembles the A-chain of IRAb. To verify whether IRIP or IRAb can also confer systemic protection against viruses, transgenic RIP expressing scions were grafted onto control rootstocks and leaves of the rootstocks challenged with tobacco etch virus (TEV). In spite of the strong local antiviral effect of IRIP and IRAb the RIPs could not provide systemic protection against TEV. Hence our results demonstrate that expression of the type-1 and type 2 RIPs from Iris confers tobacco plants local protection against two unrelated viruses. The antiviral activity of both RIPs was not accompanied by an induction of pathogenesis-related proteins. It is suggested that the observed antiviral activity of both Iris RIPs relies on their RNA N-glycohydrolase activity towards TMV RNA and plant rRNA. PMID- 15278457 TI - The challenges of moving chemicals within and out of cells: insights into the transport of plant natural products. PMID- 15278458 TI - Differential expression patterns of two cellulose synthase genes are associated with primary and secondary cell wall development in aspen trees. AB - The quality and quantity of cellulose deposited in the primary and secondary cell walls of plants vary in accordance with their biological function. However, the molecular basis of such cellulose heterogeneity has so far remained unclear. Since enrichment of better-quality cellulose, in terms of increased degree of polymerization and crystallinity, is one of the goals of forest biotechnology, our main objective is to decipher the roles of distinct cellulose synthase (CesA) genes in tree development, with special reference to wood production. Here, we report two full-length CesA cDNAs, PtrCesA3 and PtrCesA4, from an economically important tree aspen (Populus tremuloides). PtrCesA3 is orthologous to the Arabidopsis AtCesA4 gene involved in secondary wall formation, whereas PtrCesA4 is orthologous to the Arabidopsis AtCesA1 gene involved in primary cell wall formation. To define the specific cell types expressing these CesA genes, we explored the natural distribution patterns of PtrCesA3 and PtrCesA4 transcripts in a variety of aspen organs, such as leaves, petiole, stem, and roots, using in situ hybridization with hypervariable region-specific antisense riboprobes. Such a side-by-side comparison suggested that PtrCesA3 is exclusively expressed in secondary-wall-forming cells of xylem and phloem fibers, whereas PtrCesA4 is predominantly expressed in primary-wall-forming expanding cells in all aspen organs examined. These findings suggest a functionally distinct role for each of these two types of PtrCesAs during primary and secondary wall biogenesis in aspen trees, and that such functional distinction appears to be conserved between annual herbaceous plants and perennial trees. PMID- 15278460 TI - Properties of 3D rotations and their relation to eye movement control. AB - Rotations of the eye are generated by the torques that the eye muscles apply to the eye. The relationship between eye orientation and the direction of the torques generated by the extraocular muscles is therefore central to any understanding of the control of three-dimensional eye movements of any type. We review the geometrical properties that dictate the relationship between muscle pulling direction and 3D eye orientation. We then show how this relation can be used to test the validity of oculomotor control hypotheses. We test the common modeling assumption that the extraocular muscle pairs can be treated as single bidirectional muscles. Finally, we investigate the consequences of assuming fixed muscle pulley locations when modeling the control of eye movements. PMID- 15278463 TI - Causal transfer function analysis to describe closed loop interactions between cardiovascular and cardiorespiratory variability signals. AB - Although the concept of transfer function is intrinsically related to an input output relationship, the traditional and widely used estimation method merges both feedback and feedforward interactions between the two analyzed signals. This limitation may endanger the reliability of transfer function analysis in biological systems characterized by closed loop interactions. In this study, a method for estimating the transfer function between closed loop interacting signals was proposed and validated in the field of cardiovascular and cardiorespiratory variability. The two analyzed signals x and y were described by a bivariate autoregressive model, and the causal transfer function from x to y was estimated after imposing causality by setting to zero the model coefficients representative of the reverse effects from y to x. The method was tested in simulations reproducing linear open and closed loop interactions, showing a better adherence of the causal transfer function to the theoretical curves with respect to the traditional approach in presence of non-negligible reverse effects. It was then applied in ten healthy young subjects to characterize the transfer functions from respiration to heart period (RR interval) and to systolic arterial pressure (SAP), and from SAP to RR interval. In the first two cases, the causal and non-causal transfer function estimates were comparable, indicating that respiration, acting as exogenous signal, sets an open loop relationship upon SAP and RR interval. On the contrary, causal and traditional transfer functions from SAP to RR were significantly different, suggesting the presence of a considerable influence on the opposite causal direction. Thus, the proposed causal approach seems to be appropriate for the estimation of parameters, like the gain and the phase lag from SAP to RR interval, which have a large clinical and physiological relevance. PMID- 15278464 TI - Metabolic profile of patients after elective open heart surgery. AB - To evaluate the surgical stress of open heart surgery with moderate hypothermic cardiopulmonary bypass (CPB), oxygen consumption (VO2), carbon dioxide production (VCO2), resting energy expenditure (REE), respiratory quotient (RQ), 24 hour urinary urea nitrogen excretion (UUN), and glucose, fat and protein utilization were determined in 20 patients before and after open heart surgery. Proteins (albumin, prealbumin and transferin) and body weight were measured preoperatively and on 6th postoperative day (POD). Preoperative predicted EE as determined by the Harris-Benedict equation was correlated with measured REE. No significant alteration in VO2, VCO2, REE, 24 hour UUN and protein utilization was observed on the first 6 PODs. RQ decreased significantly on the 1st, 3rd and 4th POD. This was attributed to greater fat utilization due to reduced calorie intake during the early postoperative period. Transport proteins reduced slightly but insignificantly. There was a significant reduction in body weight at the end of the study period due probably to loss of body water. We conclude that patients in the early postoperative period after uneventful open heart surgery are neither hypermetabolic nor hypercatabolic when compared with their stable state before operation. PMID- 15278465 TI - Hypertensive crisis during the resection of an adrenal tumor in primary aldosteronism. AB - Primary aldosteronism is one of the few causes of hypertension which is subject to total surgical treatment, but a hypertensive crisis can occur during the resection of the adrenal tumor. We undertook this study to evaluate the relationship between hormonal factors and a hypertensive crisis during surgery. Sixteen patients with primary aldosteronism who were scheduled for the resection of an adrenal tumor were participated in this investigation. Hormonal factors and hemodynamic variables were evaluated before induction of anesthesia, after induction of anesthesia, 30 minutes after the incision, during tumor manipulation, during resection, and immediately after surgery. During tumor manipulation, a hypertensive crisis occurred in six patients (hypertensive group; HG) but not in any others (non-hypertensive group; NHG). There were no differences in hormonal levels, except epinephrine, during tumor manipulation between HG and NHG. Hemodynamic evaluations revealed an increase of systemic vascular resistance during the hypertensive crisis. We conclude that the hypertensive crisis during the manipulation of an adrenal tumor is caused by the rapid release of epinephrine from the manipulated adrenal gland. PMID- 15278466 TI - Hemodynamic and oxygen delivery-consumption changes during partial liver resection. AB - The effects of partial liver resection on hemodynamics and the oxygen delivery consumption relationship were evaluated in ten patients with hepatocellular carcinoma. The cardiac index and oxygen delivery were increased significantly (P < 0.05) at 30 minutes after incision, 30 min after liver resection and in the recovery room. Oxygen delivery decreased significantly (P < 0.05) during liver resection. Oxygen consumption remained low throughout the procedure. We did not discover any flow-dependent change in oxygen consumption. Although our patients persisted a hyperdynamic state throughout surgery, their arterial ketone body ratio remained low. Therefore, it may be necessary to maintain a hyperdynamic state during partial liver resection in order to increase hepatic blood flow. PMID- 15278467 TI - Dose-response relation and time course of action of pipecuronium in patients anesthetized with nitrous oxide and sevoflurane. AB - The dose-response relation of pipecuronium, the time course of its neuromuscular blocking effects, and the reversibility of the residual block by neostigmine have been investigated in patients under sevoflurane/N2O Anesthesia using a neuromuscular transmission analyzer (Accelograph, Biometer, Denmark). After an initial dose of pipecuronium (0.04 mg.kg(-1), i.v.), the maximum block rate, onset time, the time from administration until 25% recovery and 50% recovery of control twitch height of the first response to train-of-four nerve stimulation and the interval time of administration of maintenance dose (0.005 mg.kg(-1), i.v.) were 93.7 +/- 7.68%, 5.0 +/- 1.84, 55.4 +/- 23.92, 73.0 +/- 29.44 and 38.7 +/- 15.50 minutes, respectively. The average intubation score (excellent; 0, good; 1 fair; 2, poor; 3) was 0.63 +/- 0.56 at the level of 95.88 +/- 5.06% block. Neostigmine (1.5 mg) promptly reversed the residual neuromuscular blockade induced by pipecuronium (reversal time: 10.1 +/- 2.98 minutes). No side effects attributable to pipecuronium was seen in this study. In conclusion, pipecuronium is a very useful nondepolarizing neuromuscular blocking agent especially for moderately long surgical procedure over 4-5 hours. PMID- 15278468 TI - Endocrine and hemodynamic changes during liver surgery in patients with compensated liver cirrhosis. AB - The purpose of this study was to determine hormonal levels in compensated liver cirrhotic patients under general anesthesia before and after liver surgery. We measured plasma norepinephrine, epinephrine, arginine vasopressin, and aldosterone levels and renin activity in non-cirrhotic and compensated cirrhotic patients undergoing liver resection after induction of anesthesia but before skin incision and after the end of operation but before discontinuation of nitrous oxide. We simultaneously measured hemodynamic variables. Plasma levels of norepinephrine (P < 0.001), epinephrine (P < 0.001), arginine vasopressin (P < 0.05), renin (P < 0.05) and aldosterone (P < 0.001) significantly increased after completion of surgery compared with those before incision in both groups. There was a significant positive correlation between plasma renin and aldosterone (r = 0.56, P < 0.01) levels in non-cirrhotics, but no correlation was observed in cirrhotics; and there was a significant positive correlation between plasma norepinephrine and arginine vasopressin (r = 0.45, P < 0.05) levels in non cirrhotics, but no correlation in cirrhotics. Cardiac index and arterial pressure increased after the end of operation (P < 0.05). This increase after the operation was the same between cirrhotic and non-cirrhotic groups. There were no changes in heart rate, mean pulmonary arterial pressure, and pulmonary capillary wedge pressure after the end of operation. We conclude that hemodynamic and endocrinological changes were similar between compensated cirrhotic patients and non-cirrhotic patients during liver surgery. Endocrine changes might partly explain the hemodynamic changes during surgery. PMID- 15278469 TI - The intrathecal spread of hyperbaric dibucaine in adolescents. AB - We investigated the spread of spinal anesthesia with hyperbaric dibucaine in 20 adolescents aged 9-18 yr and 20 adults aged 23-53 yr. No significant difference was found between the two groups with regard to height, whereas a statistical significant was found between the two groups with regard to weight. Spinal anesthesia was conducted with Neo-Percamin S injected at the L3-L4 interspace through a 25-gauge spinal needle. Injected volumes of the anesthetic solution were calculated from the patients' height at 0.01 ml.cm(-1). In adolescents, 1.6 +/- 0.1 ml (mean +/- SD) of the anesthetic solution produced 19.4 +/- 1.5 spinal segments blocked. In adults, 1.6 +/- 0.1 ml of the solution produced 13.4 +/- 1.6 spinal segments blocked. A high spinal anesthesia above T5 was achieved in 17 (85%) patients in adolescents, whereas such a high level of spinal anesthesia was not experienced in adults. These results suggest that the hyperbaric dibucaine solution for spinal anesthesia in adolescents may have a tendency to produce an unexpectedly extensive spread of anesthesia. PMID- 15278471 TI - Effects of magnesium on isolated canine coronary arterial tension. AB - The effects of magnesium on the tension of isolated canine coronary arterial strips were studied. In the solution containing K+ of 20 mEq.l(-1), Ca2+ of 4 mEq.l(-1), and Na+ of 127 mEq.l(-1), the tension was 811 +/- 111 mg with Mg2+ of 1 mEq.l(-1), 494 +/- 135 mg with Mg2+ of 10 mEq.l(-1), 272 +/- 126 mg with Mg2+ of 20 mEq.l(-1), -52 +/- 63 mg with Mg2+ of 30 mEq.l(-1), -69 +/- 80 mg with Mg2+ of 40 mEq.l(-1). In the solution containing K+ of 20 mEq.l(-1), Na+ of 12 mEq.l( 1) and Ca2+ of 0 mEq.l(-1), the tension was 102 +/- 22 mg with Mg2+ of 1 mEq.l( 1), 3 +/- 35 mg with Mg2+ of 10 mEq.l(-1), -49 +/- 33 mg with Mg2+ of 20 mEq.l( 1), -59 +/- 49 mg with Mg2+ of 30 mEq.l(-1), -65 +/- 54 mg with Mg2+ of 40 mEq.l( 1). The data demonstrated that Mg2+ above 30 mEq.l(-1) inhibited the increase in tension caused by Ca2+ and Mg2+ above 20 mEq.l(-1) inhibited the increase in tension caused by low Na+ concentration. PMID- 15278470 TI - Intra-operative blood pressure control by prostaglandin E1 in patients with hypertension and ischemic heart disease--a multi-center study. AB - The purpose of this multi-center study was to evaluate the efficacy and safety of prostaglandin E1 (PGE1) administration in achieving deliberate hypotension and in treating intraoperative hypertension for patients with a history of hypertension and ischemic heart disease. PGE1 (0.08 microg.kg(-1).min(-1)) decreased systolic blood pressure from 125 +/- 29 to 106 +/- 22 mmHg (mean +/- SD) in the deliberate hypotension group (n = 158) and from 155 +/- 34 to 125 +/- 32 mmHg in the antihypertension group (n = 55). The heart rate significantly increased from 80 +/- 15 to 85 +/- 18 beats.min(-1) in the deliberate hypotension group, but was not significantly altered in the antihypertension group. The time required to obtain the desired level of blood pressure was approximately 20 min in the deliberate hypotension group. When the infusion was stopped, blood pressure returned approximately to the preinfusion level within about 20 min. No rebound hypertension was observed. PGE1 significantly increased the urine flow in patients who had a low urine flow before PGE1 infusion. Thirteen out of 213 patients (5.6%) had side effects such as excessive hypotension (1%), phlebitis (3%), and unexpected tachycardia (1%), which were alleviated gradually after discontinuation of PGE1 infusion. No dysarrhythmia and further ST segment changes in the electrocardiograms were observed. These findings suggest that PGE1 can be safely used to control arterial blood pressure during surgery in patients having preoperative hypertension and ischemic heart disease. PMID- 15278472 TI - Evaluation of histamine-releasing property of propofol in whole blood in vitro. AB - We examined the property of emulsion form of propofol (ICI 35 868) to release histamine in whole blood in vitro. Heparinized whole blood from 10 healthy volunteers were incubated with medium and propofol at the final concentration of 0, 1, 10 and 100 microg.ml(-1). The concentration of histamine in supernatant fluid after incubation was measured by radioimmunoassay. Histamine release was expressed as the percentage of the concentration of histamine released into supernatant fluid relative to the total cellular histamine content, which was yielded by destroying cell components in the whole blood. Histamine release in the presence of propofol at the concentrations of 1, 10 and 100 microg.ml(-1) were almost the same as histamine release in the absence of propofol. We conclude that emulsion form of propofol has no property to release histamine in whole blood in vitro. PMID- 15278473 TI - Inhibition of lipid peroxidation by some dihydropyridine derivatives. AB - The efficacy of dihydropyridine derivatives in inhibiting lipid peroxidation was studied using modified Buege and Aust's method. The method first involves keeping a decapitated rat head at 37 degrees C for 30 min in order to induce global ischemia. Then, the cortex is removed and homogenized, and the homogenate is subsequently exposed to air for 30 min for reoxygenation. Finally, the amount of thiobarbituric acid-reactive substances (TBAR) is measured. With this method, nisoldipine, nimodipine, nitrendipine, nifedipine and nicardipine were all shown to have an antioxidant activity that correlated with their lipophilicity, which was determined by their octanol/water partition coefficients. PMID- 15278474 TI - The effect of vagal afferent on total vascular compliance in rats. AB - This study was designed to investigate the effect of vagal afferent stimulation on total vascular compliance (TVC). Rats were anesthetized with sodium pentobarbital and artificially ventilated, TVC was determined together with stressed and unstressed blood volumes by measuring mean circulatory filling pressure (Pmcf) at three different levels of circulating blood volume. Measurements was repeated with the intact vagus, after vagotomy and during stimulation of vagal afferents. Vagotomy caused no change in TVC, Pmcf, and stressed and unstressed blood volumes. On the other hand, electrical stimulation of the vagal afferents for 30 sec increased TVC from 3.03 +/- 0.51 to 3.39 +/- 0.44 ml.mmHg(-1).kg(-1) (P < 0.05) and decreased Pmcf from 7.83 +/- 1.40 to 7.22 +/- 1.21 mmHg (P < 0.05). Neither stressed nor unstressed blood volume was changed by vagal stimulation. These results indicate that excitation of vagal afferent causes venodilation and increases TVC without changing stressed and unstressed blood volumes. PMID- 15278475 TI - Plasma histamine levels during induction of anesthesia with propofol in dogs. AB - We examined a property of emulsion formation of propofol (ICI 35868) to release histamine into circulating plasma in dogs. Plasma histamine was measured with radioimmunoassay before (baseline), and 1, 5 and 10 min after the administration of 15 mg.kg(-1) propofol. There were no significant differences between the plasma histamine levels at 1, 5 and 10 min after the administration of propofol and the baseline level. We conclude that the emulsion formation of propofol of 15 mg.kg(-1) does not release histamine during induction of anesthesia in dogs. PMID- 15278476 TI - Fentanyl antagonizes diazepam on carotid sinus baroreflex control of circulation in rabbits. AB - To investigate the effects of a combination of fentanyl and diazepam on carotid sinus baroreflex in conscious rabbits, we examined the responses of mean systemic arterial pressure (MAP), heart rate (HR) and total peripheral resistance (TPR) to bilateral carotid occlusion (BCO). Seven rabbits were given 0.5 mg.kg(-1) of diazepam i.v. followed by 10 mg.kg(-1) of fentanyl i.v. at 5 min intervals (group 1), and the drugs were given in the reverse order to 5 other rabbits (group 2). BCO was repeated in conscious state (control) and after each drug injection. MAP responses did not differ from control response in either group when both drugs were given. In group 1, however, diazepam decreased HR response to 71.4% of control, and increased TPR response by 36%. Fentanyl administration reversed diazepam-induced changes in BCO responses to the control level. In group 2, fentanyl decreased TPR response to 61.6% of control and increased HR response by 41.5%. Administration of diazepam following fentanyl restored HR and TPR responses to control levels. Carotid sinus baroreflex gain was 3.1 +/- 0.4 (mean +/- SEM) in control and 3.1 +/- 0.4 after administration of both drugs in 12 rabbits. The results suggest that a sedative dose of either fentanyl or diazepam antagonizes the other drug's action on the carotid sinus baroreflex. The combination of fentanyl and diazepam has little influence on carotid sinus baroreflex control of the circulation in rabbits. PMID- 15278477 TI - Effects of halothane on carotid occlusion in rabbits. AB - Effects of halothane on the carotid sinus baroreflex control of circulation were studied in chronically instrumented rabbits. The carotid sinus baroreflex was evaluated by the hemodynamic responses to bilateral carotid occlusion (BCO). Either 0.5 or 1.0 MAC of halothane inhalation did not alter mean arterial pressure (MAP) or total peripheral resistance (TPR), but significantly increased heart rate (HR). Carotid occlusion produced a significant increase in MAP and HR, and both responses were attenuated dose-dependently by halothane. Halothane depressed the reflex gain of arterial pressure from 3.5 +/- 0.3 at conscious state to 1.3 +/- 0.2 at 1.0 MAC halothane. Response of cardiac output (CO) to BCO was attenuated significantly only at 1.0 MAC compared with those responses at conscious state and at 0.5 MAC. Response of TPR was attenuated at both 0.5 and 1.0 MAC halothane as compared with at conscious state but no significant difference existed between the two concentrations of halothane. These data suggested that halothane could attenuate the carotid occlusion responses to various degrees in the involved effector components. 0.5 MAC halothane attenuated MAP response to BCO predominantly by attenuating reflex peripheral vasoconstriction. The reduced CO response was mainly responsible for further attenuation of MAP response at 1.0 MAC halothane. PMID- 15278478 TI - Hepatic rupture after cardiac arrest--a case report. PMID- 15278479 TI - Air ventilation during pulmonary artery banding operation. PMID- 15278480 TI - Tracheal tear during esophageal blunt resection. PMID- 15278481 TI - Neuromuscular effects of sevoflurane in a patient with Myasthenia Gravis. PMID- 15278482 TI - General anesthesia for cesarean section in a parturient with quintuplet pregnancy. PMID- 15278483 TI - Superimposed high frequency jet ventilation for the treatment of cardiogenic pulmonary edema in a case of viral myocarditis. PMID- 15278484 TI - Anaphylactoid reaction associated with blood transfusion during anesthesia--a case of characteristic hemodynamic changes. PMID- 15278485 TI - Coronary artery spasm occurring twice the patient received general anesthesia. PMID- 15278486 TI - The effect of sevoflurane on rat liver mitochondrial respiration. AB - The effect of sevoflurane on rat liver mitochondrial respiration has been investigated with a Clark type oxygen electrode at 25 degrees C, pH 7.4. The higher susceptibility of NADH-linked substrate (glutamate) oxidation (with respect to succinate oxidation) to the damages by sevoflurane has been confirmed. PMID- 15278487 TI - Blood sampling through a peripheral venous infusion line. AB - The usefulness of a blood sampling technique for the measurement of blood glucose concentration through a peripheral venous infusion line was investigated with patients under general anesthesia. An infusion line was connected through a T connector to a venous catheter indwelled in a forearm vein, and lactated Ringer's solution was infused at a flow rate of 100 ml/hour. At an arbitrary time, 0.5 ml of blood sample was obtained through the T-connector after discarding 1.5 ml of blood, while a vein in another arm was punctured to obtain 0.5 ml of blood. As a result, the glucose concentration in the samples from the venous infusion line was strongly correlated with that obtained by direct puncture, and the regression line passed through the origin. This suggests that blood sample from a peripheral venous line can be used in place of samples obtained by direct puncture. PMID- 15278488 TI - Whole body oxygen consumption after hypothermic cardiopulmonary bypass. AB - Whole body oxygen consumption and the substrate for energy production during the post-bypass period have not been clarified. We hypothesized that the substrate composition for energy production during post-bypass period might be different from that during pre-bypass period because of surgical diabetic state induced by hypothermic cardiopulmonary bypass (CPB). We measured whole body oxygen consumption, carbon dioxide production and respiratory quotient by the gas exchange method using the Datex Deltatrac before and after hypothermic cardiopulmonary bypass. We also measured oxygen consumption by Fick's principle. Whole body oxygen consumption (P < 0.001) and carbon dioxide production (P < 0.05) increased significantly above pre-CPB values after the termination of CPB. Respiratory quotient (P < 0.01) decreased significantly below pre-CPB values after the termination of CPB. We conclude that oxygen consumption increased significantly above pre-bypass values after the termination of hypothermic cardiopulmonary bypass at least under the fentanyl, diazepam, chlorpromazine anesthesia with continuous infusion of nitroglycerin and nicardipine. The changes in respiratory quotient suggest a relatively higher ratio of lipid metabolism for energy production during post-bypass period. PMID- 15278489 TI - Sustained effects of plasma norepinephrine levels on femoral-radial pressure gradient after cardiopulmonary bypass. AB - In order to determine the influence of the sympathetic nervous system upon the femoral-radial artery pressure gradient after cardiopulmonary bypass (CPB), we examined plasma norepinephrine levels in 34 adult male patients undergoing coronary artery bypass grafting. Cardiovascular parameters, including systolic arterial pressure, mean arterial pressure, cardiac index (CI), systemic vascular resistance index (SVRI), pulmonary artery pressure (PAP), hemoglobin (Hb) and peak dP/dt of radial and femoral artery pressures were measured after sternotomy, and immediately after the discontinuation of CPB and 90 min after CPB. Plasma norepinephrine levels were measured after sternotomy, after aortic declamping and 90 min after CPB. The patients were divided into two groups. Group A consisted of 17 patients whose femoral minus radial systolic pressure difference was 15 mmHg or more at 90 min after CPB, while Group B consisted of 17 patients with the difference less than 15 mmHg. Group A patients had significantly longer time values in the duration of both CPB (Group A 175 +/- 10 min; Group B 115 +/- 12 min, P < 0.001) and aortic cross clamping (Group A 116 +/- 7 min, Group B 71 +/- 9 min, P < 0.001). Although there was no significant difference in Hb or PAP of 90 min after CPB in Groups A and B, the following values, listed in the order of A to B, were obtained; CI, 2.79 +/- 0.10 versus 3.46 +/- 0.16 l.min(-1).m(-2) (P < 0.01); mean radial artery pressure (MRP), 58.7 +/- 2.4 versus 65.1 +/- 1.8 mmHg (P < 0.05); peak dP/dt of radial artery pressure, 568 +/- 64 versus 1026 +/- 61 mmHg.sec(-1) (P < 0.001); and plasma norepinephrine concentration, 1.81 +/- 0.25 versus 0.98 +/- 0.10 ng.ml(-1) (P < 0.01), which were statistically significant. The higher femoral-radial artery pressure gradient after CPB was observed in patients with both a longer CPB time and a higher plasma norepinephrine concentration. These results suggest that a marked constriction of peripheral arteries might have produced a damped transmission of the pressure pulse to the radial artery. PMID- 15278490 TI - Postoperative recovery of arterial oxygen saturation determined by pulse oximetry in pediatric patients. AB - Small children are physiologically subject to arterial oxygen desaturation. However, few reports have referred to the risk factors related to postanesthetic hypoxemia and the duration of hypoxemia. The purpose of this study was to clarify these two aspects. Eighty-five ASA physical status I infants and children were included in the study. They were scheduled for minor surgery. Fifty-six underwent oral endotracheal intubation, and 29 patients breathed from a mask. Anesthesia was maintained with Enflurane or Halothane and nitrous oxide. Arterial oxygen saturation was measured with a pulse oximeter. The measurements were started shortly after patients' arrival in the recovery room, and conducted every 5 min at least for 1 hour. Ten patients had SpO2 values of less than 95%. In all except one, SpO2 decreased within 10 min after arrival in the recovery room. Age, height, and weight of these 10 children were significantly different from the remaining 75, but there were no significant differences in anesthetic duration and postanesthetic awakefulness between the group with postanesthetic hypoxemia and the one without. The importance of monitoring the clinical condition of pediatric patients after general anesthesia is universally acknowledged. Monitoring with the pulse oximeter has proven very useful and shows that, unless oxygen saturation is monitored, all children should receive supplemental oxygen. PMID- 15278491 TI - Evaluation of the penetration depth of transdermally applied 3% GA MHPh 2Na-10% lidocaine gel in man. AB - In order to estimate the penetration depth of transdermal 3% GA MHPh 2Na-10% lidocaine gel mixture, the following physiological functions of the skin were examined before and after a 60 min occulusive application of the gel in 16 adult volunteers. Thermal sweat expulsins ceased completely on the gel-treated ventral surface of one forearm in all the firs 5 subjects, though it continued on the untreated contrast area of the other forearm. Sympathetic skin response (SSR) was also no longer induced on the gel-treated middle finger in 1 of another 3 subjects and was severely depressed in the other 2 subjects, while the SSR on the untreated index finger appeared constantly. Vasomotion of the skin circulation on another 3 subjects, remained unaffected on both the gel-treated and the untreated fingers. Extraction of a leg-hair in the treated area did not induce pain sensation in all the last 5 subjects. In addition to the transcellular main roots, some of the transcutaneously applied gel seems to penetrate deeply into the skin through the appendageal roots such as the eccrine sweat glands and the pilosebaceous glands. PMID- 15278492 TI - Rationale for preoperative screening of anti-HCV antibody. AB - We investigated the incidence of the anti-HCV antibody and associated factors in 1,031 surgical patients who had received blood transfusion during or after operation from October 1988 to April 1991, at Kyushu University Hospital. One hundred fifteen patients (11.2%) were anti-HCV positive. Sixty of the 219 patients (27.%) with a history of transfusion were positive, as were 55 of 812 (6.8%) without it. Patients aged under 40 showed a 0.6% positive rate (1 of 175) as did 8.5% (54 of 637) of those 40 and over in the no transfusion history group. Among the 637 patients without transfusion histories and aged over 40, patients with preoperative maximum ALT value over 36 IU. l(-1) had significantly higher positivity (16.0%, 29/181) than those with ALT values less than 35 IU. l(-1) (5.5%, 25/456, P < 0.01). The incidence of anti-HCV antibody in preoperative surgical patients in our hospital is ten times higher than that of donors. Anti HCV are associated with transfusion, age, and liver dysfunction. Operating room personnel are at high risk because of contact with many HCV carrier patients. PMID- 15278493 TI - A new infrared tympanic thermometer in surgery and anesthesia. AB - We have evaluated a new infrared tympanic thermometer, IT-10, as an intraoperative temperature monitor in patients with or without open abdominal surgery. It determines temperature by measuring infrared radiation given off by a warm object. Temperatures measured with this device were closely correlated with those measured with rectal and bladder thermometries. We conclude that this new tympanic thermometer is safe, convenient, accurate, and easily usable in the clinical situation. PMID- 15278494 TI - The effects of an intravenous nicardipine injection on baroreflex control of heart rate in man. AB - The effects of nicardipine injection on baroreflex control of heart rate were investigated by both pressor and depressor tests in 17 adult patients. Baroreflex sensitivity was attenuated after nicardipine injection by the pressor test using phenylephrine, whereas it was not changed by the depressor test using nitroglycerine. No resetting of the baroreflex occurred after nicardipine injection. By the pressor test, the plasma norepinephrine level was decreased, indicating that parasympathetic activity increased, and by the depressor test, the plasma norepinephrine concentration was increased, indicating that sympathetic activity increased. These results suggest that it is safe to use nicardipine clinically even when reduction in blood pressure for hypovolemia or unclamping the main artery is expected, and it is disadvantageous to administer the drug when an increase in blood pressure due to cross-clamping of the main artery is forecasted. PMID- 15278495 TI - Modification of hepatic protein kinase C with phorbol myristate acetate and staurosporine alters hemodynamics in the perfused rat liver. AB - Activation of protein kinase C (PKC) has been implicated in the pathogenesis of endotoxicosis and severe sepsis. Since hepatic blood flow and metabolism have been known to be altered in endotoxicosis and sepsis, we studied the hemodynamic effect of PKC modulation with phorbol 12-myristate 13-acetate (PMA) and staurosporine (St) on the perfused rat liver. The liver was isolated from overnight-fasted male Sprague-Dawley rats and placed in a recirculating perfusion apparatus. The liver was perfused with Krebs-Ringer-bicarbonate solution at a constant pressure of 12 cmH2O. Flow to the liver was continuously monitored with an electric magnetic flowmeter. PMA at an initial concentration of 2 x 10(-8) M significantly decreased hepatic flow. Staurosporine (St), a potent PKC inhibitor at 4 x 10(-7) M produced a small increase in hepatic flow. Pretreatment with St significantly attenuated the flow reduction by PMA. St significantly suppressed the flow reductions by 4 x 10(-6) M of prostaglandin E2 and D2. These results suggest that the PKC inside the liver may play an important role in the regulation of hepatic blood flow during endotoxicosis and sepsis. PMID- 15278496 TI - Prostaglandin E1 (PGE1) attenuates vasoconstriction induced by PGE2, PGD2 and phorbol myristate acetate in the perfused rat liver. AB - It has been shown that prostaglandins (PGs) produced by Kupffer and endothelial cells play an important role in mediating physiological responses to various immunological stimuli. We studied the effect of prostaglandin E1 (PGE1) on the hemodynamic and metabolic changes induced by prostaglandin E2 (PGE2), D2 (PGD2) and phorbol 12-myristate 13-acetate (PMA), a potent inducer of PGs in the isolated rat liver perfused with Krebs-Ringer-bicarbonate (KRB) solution at a constant pressure of 12 cmH2O. The liver was taken from overnight-fasted male Sprague-Dawley rats weighing 260 to 310 g. Both PGE2 and PGD2 significantly decreased hepatic flow when their initial concentration was elevated to micromolar range. Although 1 x 10(-6) M of PGE1 did not have a major effect on hepatic flow, it significantly attenuated the declines of hepatic flow produced by 4 x 10(-6) M of PGE2 and PGD2. However, none of PGs tested influenced glucose and lactate concentrations in the medium. Continuous infusion of PGE1 into the medium at a rate of 5 microg.min(-1) significantly diminished the decreases in hepatic flow and oxygen consumption induced by 2 x 10(-8) M of PMA. These results suggest that administration of PGE1 may preserve hepatic blood flow by modifying the intrahepatic regulatory mechanism involving the activation of Kupffer and endothelial cells. PMID- 15278497 TI - Effects of surfactant on lung injury induced by hyperoxia and mechanical ventilation in rabbits. AB - We evaluated the effects of exogenous surfactant on lung injury caused by 100% oxygen and mechanical ventilation in rabbits. Surfactant-treated rabbits (n = 9) were ventilated with 100% oxygen for 36 hours and bovine surfactant was given via the trachea 12 hours after the start of mechanical ventilation. Saline-treated (n = 9) rabbits were treated identically, except that they received saline without surfactant. There were no significant changes in hemodynamics, lung mechanics, or arterial oxygen tension during artificial ventilation. Albumin concentration in the bronchoalveolar lavage fluid (BALF) of saline-treated rabbits was slightly higher than those in surfactant-treated rabbits and significantly higher than in non-treated rabbits. C3a concentration in BALF was significantly higher in saline treated rabbits than in surfactant-treated and non-treated rabbits. In addition, the wet-to-dry lung weight ratio was significantly lower in surfactant-treated rabbits than in saline-treated rabbits (5.06 +/- 0.10 vs. 5.67 +/- 0.14, P < 0.05). Light microscopy revealed hyaline membrane formation in saline-treated rabbits, but fewer changes were observed in surfactant-treated rabbits. Electron microscopy revealed extensive endothelial cell destruction in saline-treated rabbits, while such changes except endothelial cell swelling were not observed in surfactant-treated rabbits. We conclude that exogenous surfactant attenuated lung injury caused by oxygen exposure and ventilation. PMID- 15278498 TI - Strain-differences of sensitivity to volatile anesthetics and their genetic character in mice. AB - Using loss of the righting reflex, we determined the ED50 values for enflurane, isoflurane, sevoflurane and halothane in white-haired ddN mice and black-haired C57BL mice. The ED50s (Mean +/- SEM) in ddN and C57BL mice for enflurane were 1.65 +/- 0.01 and 1.19 +/- 0.01% atm, for isoflurane 1.02 +/- 0.01 and 0.74 +/- 0.01% atm, for sevoflurane 2.29 +/- 0.03 and 1.95 +/- 0.03% atm, and for halothane 0.97 +/- 0.01 and 0.97 +/- 0.01% atm, respectively. The results indicate that the ddN strain is more resistant to enflurane, isoflurane and sevoflurane than the C57BL strain. The sensitivities to enflurane and isoflurane is F1 progeny of reciprocal crosses between ddN and C57BL mice revealed that in the ddN strain enflurane resistance is an incompletely dominant or polygenic character, isoflurane resistance in ddN strain is an autosomal recessive character and both are controlled by genes on the sex (X) chromosome. Enflurane and isoflurane resistances are controlled by at least 2 genes, one on the X chromosome, and each resistance is controlled by a different genetic mode PMID- 15278499 TI - Emergency laparotomy in uncontrolled thyrotoxic patient with preoperative fulminant hepatic failure. PMID- 15278500 TI - Changes in free and total catecholamine concentrations in plasma in patients undergoing coronary artery bypass grafting under high-dose fentanyl anesthesia. AB - We measured free and total catecholamine in ten patients undergoing coronary artery-bypass grafting under high-dose fentanyl (93.9 +/- 2.2 microg.kg(-1), mean +/- SE) anesthesia. Arterial blood samples were obtained: 1) before induction of anesthesia (control), 2) 1 min after intubation, 3) 1 min after skin incision, 4) 1 min after median sternotomy and, 5) just before termination of cardiopulmonary bypass (CPB). The concentrations of free and total catecholamine were measured by HPLC using fully automated analyzer, 8030-TOHSO. Free and total catecholamine concentrations did not change significantly before CPB. At the termination of CPB, however, the levels in free dopamine, norepinephrine and epinephrine all increased several fold as compared with control. Similarly, total norepinephrine and epinephrine also increased at the end of PCB, while total dopamine did not change. Present results indicated that 1) the measurement of free CAs is more significant than the measurement of total CAs for the assessment of sympathoadrenal responses to surgical stimuli, and that 2) high-doses of fentanyl produce hemodynamic stability by suppressing sympathoadrenal responses elicited by the usual surgical procedures. However, stress triggered by CPB could not be suppressed totally by fentanyl even with high dose. PMID- 15278501 TI - Severe aryepiglottic edema following extubation in a patient with rheumatoid arthritis. PMID- 15278502 TI - Effect of nicardipine on pulmonary hypertension after repair of congenital heart defects in early postoperative period. AB - We examined the effect of nicardipine and hyperventilatin on pulmonary arterial pressure (PAP) in four patients with pulmonary hypertension in congenital heart defect especially in the early postoperative period. There was a significant positive correlation between the values of arterial carbon dioxide tension (PaCO2) and the ratio of mean PAP to mean systemic arterial pressure (Pp/Ps) in two patients whose Heath Edwards classification was Grade II; one of them also had 20% reduction of mean PAP by nicardipine without changing mean arterial pressure. In the remaining two patients showing no correlation between the values of PaCO2 and Pp/Ps, nicardipine did not reduce PAP. Although nicardipine reduced PAP without changing systemic arterial pressure in only one out of four patients, these results suggest that nicardipine may be a drug for control of PAP during weaning phase from the ventilator especially in patients whose PAP decreases under hyperventilation. PMID- 15278503 TI - Nystagmus following intrathecal morphine administration. PMID- 15278504 TI - Perioperative management of a patient with congenital antithrombin III deficiency. PMID- 15278505 TI - Perioperative management of two patients with respiratory problems undergoing Abdominal surgery with high spinal anesthesia. PMID- 15278506 TI - Pharyngeal perforation following endotracheal intubation in a patient with rheumatoid arthritis. PMID- 15278507 TI - Cardiac and respiratory arrest following aortocaval fistula. PMID- 15278508 TI - An analysis of flow volume curves during artificial ventilation. AB - Expiratory flow-volume curves during artificial ventilation (FV-av) were analyzed in 48 patients undergoing general anesthesia. They were divided into 4 groups according to preoperative respiratory disorders; obstructive type (group 1), restrictive type (group 2), small airway disease (group 3) and normal control (group 4). Expiratory flow rates and volumes during artificial ventilation were plotted on an X-Y recorder to calculate V50/V25, mean time constant ratio (MTCR), obstructive index (OI) and slope ratio (SR). FV-av values were compared among groups. FV-av values in groups 2 were significantly higher than those in group 4. The values in group 1 and those in group 3 were not significantly different from those in group 4. FV-av values may reflect restrictive respiratory dysfunctions but they are not sensitive enough to detect obstructive lung disease. PMID- 15278510 TI - Lack of the mechanoreceptor influences on ventilatory control during halothane anesthesia in humans. AB - Mechanical influences independent of chemoreceptor function on ventilatory control were studied in halothane-anesthetized, artificially ventilated patients using the technique reported by Altose et al. (Respir Physiol 66: 171-180, 1986). Contribution of mechanical factor was indirectly assessed by comparing the values of arterial carbon dioxide tension at which the subjects started breathing efforts during CO(2) loading induced by the following two methods. 1) Partial rebreathing of expired gas and 2) Mechanical hypoventilation (successive decrease in inflation volume). These two maneuvers resulted in a similar rate of increase in end-expiratory carbon dioxide tension. However, contrary to the observation made by Altose et al. in awake volunteers, we found comparable values of ventilatory recruitment threshold for Pa(CO)(2). Thus, we speculate that halothane anesthesia and/or loss of consciousness impair transmission of afferent information from the lung and/or chest wall musculature. Such effects may be responsible for the depression of load compensatory mechanism during anesthesia. PMID- 15278511 TI - Effects of intravenously administered lidocaine on pulmonary vagal afferents and phrenic nerve activity in cats. AB - The ability of lidocaine to suppress activity of single vagal afferent fiber and that of phrenic nerve was studied in 20 cats anesthetized with pentobarbital. Slowly adapting stretch receptors (SAR, n = 16) and rapidly adapting stretch receptors (RAR, n = 7) were identified by their discharge pattern to pulmonary inflation. Intravenous lidocaine (1 mg.kg(-1) or 2 mg.kg(-1)) produced a suppression of SAR activity but not of RAR activity. Suppression of phrenic nerve activity lasted much longer than that of SAR. These findings indicate that iv lidocaine acts more dominantly on CNS than on peripherals. We conclude that iv lidocaine prevents cough and hemodynamic changes caused by airway manipulation mainly through its action on CNS and not on peripherals (peripheral nerves or their receptor). PMID- 15278513 TI - Cardio-respiratory changes with increased intra-bladder pressure in prone position during anesthesia. AB - Twenty non-obese patients, 13-29 years of age, operated on for scoliosis were examined for cardio-respiratory changes that occur during positioning on bolsters, and the effect on the cardio-respiratory system of raised intra abdominal pressure was evaluated. Hemodynamic and respiratory responses were measured when the position was changed from supine to prone and back to supine during anesthesia. We measured the intra-bladder pressure using a transurethral catheter (IBP) as an index of the intra-abdominal pressure. When the position was changed from supine to prone, the cardiac index (CI) decreased by 10-30% and the systemic vascular resistance index increased by 8-14%. IBP rose significantly ( P < 0.001), but it remained below 5 mmHg. These changes continued to be mild until the patient was returned to the supine position. Pa(O)(2), A-aD(O)(2)and Qs/Qt remained unchanged. CI decreased significantly ( P < 0.001) when IBP was increased to 10 mmHg by abdominal compression, but was not affected when IBP was increased to only 5 mmHg. It was concluded that mild abdominal compression in the prone position during anesthesia has little effect on the cardio-respiratory system in lean young subjects. PMID- 15278512 TI - Hemodynamic effects of nicardipine-induced hypotension during enflurane/nitrous oxide anesthesia in man. AB - Hemodynamic effects of nicardipine-induced hypotension during enflurane/nitrous oxide were evaluated in 10 surgical patients. An infusion of nicardipine was titrated to maintain mean arterial pressure at 60 to 70 mmHg under enflurane 1.5 to 2.0 vol% and nitrous oxide 60 vol%. Mean arterial pressure was well controlled with the nicardipine infusion, whereas cardiac index increased with decreased systemic vascular resistance. Heart rate increased concomitantly with decreased blood pressure, which indicated that enflurane 1.5 to 2.0 vol% did not suppress baroreceptor reflex during nicardipine administration. However, rate-pressure product was not increased by the nicardipine. Right and left ventricular systolic work indices were not increased by the nicardipine. Right ventricular ejection fraction was not also changed by the nicardipine. Although serum norepinephrine level increased during the nicardipine infusion, the values remained within physiological ranges. Our results suggest that nicardipine-induced hypotension may be safely performed during enflurane/nitrous oxide anesthesia because neither ventricular work nor myocardial oxygen demand was increased by nicardipine. PMID- 15278514 TI - Free radical formation during splanchnic artery occlusion shock. AB - Free radical (FR) formation in the rat intestinal lumen was measured using the spin-trapping technique and electron paramagnetic resonance spectroscopy. Intestinal ischemia was produced by occluding the celiac and the superior mesenteric arteries for 30 min followed by reperfusion. The lumen was filled with a solution of PBN (N-tert-butyl-alpha-phenyl-nitrone) and the intestine was squeezed to enhance the interaction between the PBN solution and the intestinal mucosal cells. Free radicals were produced upon reperfusion, with peaks at 5 and 90 min. Post-ischemic treatment with superoxide dismutase (20 mg.kg(-1)) inhibited the increase of FR production during the second peak by 36%. In a single study in a group of leucocytopenic rats (WBC < 1500/mm(3)), the increase of FR production during the second peak was decreased by 80%. However, these treatments did not inhibit the FR production during the first peak in either group. In contrast, pretreatment with allopurinol (40 and 100 mg.kg(-1) injection at 24 and 3 hours before ischemia, respectively) inhibited the FR production during the first peak by 76%, but did not inhibit during the second peak. The changes in lipid peroxidation in the intestinal mucosa, specific gravity of the intestine and in the hematocrit were correlated to the FR production in the second peak. These results suggest that a major cause of tissue injury after reperfusion in the ischemic intestine may largely be produced by neutrophils. PMID- 15278515 TI - Serum fluoride concentration after sevoflurane anesthesia in ethanol treated rats: special reference to cytochrome P-450 in the liver. AB - The relationship between serum concentration of inorganic fluoride (F(-)) and cytochrome P-450 content after sevoflurane anesthesia was investigated in ethanol treated rats. Twenty male Wistar rats were randomly divided into 2 isocaloric diet groups of 10 rats each: one group receiving a standard diet and the other an ethanol diet. After 28 days on the diets the animals were administered 2.5% sevoflurane for 2 hr with 30% oxygen and 70% nitrous oxide. Cytochrome P-450 and cytochrome b(5) were induced by the ethanol diet. In the ethanol diet group serum concentration of F(-) was significantly higher than that of the standard diet group after sevoflurane anesthesia. These results suggest that cytochrome P-450 and b(5), which were induced by ethanol, enhanced sevoflurane defluorination. PMID- 15278516 TI - Effects of high pressure on polarity change of the water-liposome interface induced by volatile anesthetics. AB - The effects of high pressure on the interactions between volatile anesthetics and three kinds of phosphatidyl choline liposomes were investigated by fluorometry, using a thiacarbocyanine dye (3,3"-dioctadecyl-2,2"-thiacarbocyanine) which is sensitive to the environmental viscosity and dielectric constant. Seven general anesthetics, halothane, enflurane, isoflurane, methoxyflurane, sevoflurane, diethylether and chloroform were used. We have previously reported that these anesthetics decreased the phase transition temperature and increased the effective dielectric constant of the water-liposome interface using dipalmitoyl phosphatidylcholine. In this study, it was confirmed that the effects of anesthetics on the effective dielectric constant were not altered by the use of a gel or liquidcrystal membrane, and were reversed by the application of high pressure (<800 bars). The increase of the effective dielectric constant was attributed to the perturbation of hydrogen bonds at the liposomal interface. High pressure is considered to promote hydration. Our results obtained under high pressure supported previous observations made at ambient pressure, which suggested that the perturbation of hydrogen bonds at the water-liposome interface correlates with the mechanism of anesthesia. PMID- 15278517 TI - Action of opioid agonist-antagonist drugs on the pupil and nociceptive responses in mice. AB - Opioid derivatives with mixed agonist-antagonist activities are becoming increasingly more popular in analgesia. We tested the mydriatic and analgesic activity of morphine in mice in comparison with similar effects of three agonist antagonist agents: buprenorphine, butorphanol and nalbuphine. We also examined the antagonistic action of these three drugs by evaluating the analgesia and mydriasis in animals pretreated with morphine. The analgesic effect was assayed using the hot plate method while the pupillary responses were measured with a binocular operating microscope. Morphine produced dose-dependent mydriasis and analgesia in mice. The morphine-type agent buprenorphine and two nalorphine-type agonist-antagonists, butorphanol and nalbuphine, caused agonistic mydriatic and analgesic effects, usually less effective then morphine. Buprenorphine proved to have higher agonist activity than butorphanol and nalbuphine. The difference between butorphanol and nalbuphine was not statistically significant. A correlation between the mydriatic and the analgesic activity, known to exist among opioid derivatives with agonist activity only, was also demonstrated in the three investigated agonist-antagonist agents. Morphine-induced mydriasis and analgesia were reversed by all three agonist-antagonist drugs, but buprenorphine is a significantly weak antagonist in comparison with butorphanol and nalbuphine. An antagonistic property (antimydriatic and antianalgesic effects after pretreatment with morphine) of both nalorphine-type investigated drugs was not statistically significant, except for the antianalgesic effect of nalbuphine in doses 1 and 3 mg.kg(-1), which was higher in comparison with butorphanol. PMID- 15278518 TI - Effects of prostacyclin analogue, OP-2507, on function and metabolism in the ischemic working rat heart. AB - We examined the effects of a new stable prostacyclin analogue, OP-2507, on myocardial function and metabolism in the ischemic working rat heart preparation. The hearts were perfused with Krebs-Henseleit bicarbonate (KHB) buffer, and whole heart ischemia was induced by one-way aortic valve for 15 min follows by reperfusion for 30 min. In the treated hearts, OP-2507, 20 ng.ml(-1), was administered to KHB buffer from the beginning to the end of experiment. During ischemia, coronary flow in the OP-2507 group increased significantly more than that in the control group. The mechanical performance of both groups was impaired after ischemia. However, the recovery of coronary flow, cardiac output, peak systolic pressure and LV dP/dT(max) was significantly higher in the treated group than in the control group. The incidence of ventricular fibrillation during reperfusion was 100% and 25% in the control and the OP-2507 groups, respectively. Myocardial ATP content was significantly higher in the treated hearts than that in the control hearts. These results indicate that this stable prostacyclin analogue is beneficial in myocardial ischemia, even without its well known action of preventing platelet aggregation. PMID- 15278519 TI - Effects of tilting in the sagittal plane on the cephalad spread of anesthesia. AB - The effect of tilting in the sagittal plane on the spread of anesthesia was studied in 30 healthy male patients. Two ml of 0.3% hyperbaric dibucaine was used for intrathecal injection in the lateral position. After 3 min of resting on their side, 15 patients were placed in the horizontal supine position. Other 15 patients were turned to the contralateral side at 7-8 degrees in the sagittal plane. Cephalad spread of sensory analgesia by the pin-prick method, degree of motor blockade by Bromage score, mean arterial pressure, and heart rate were assessed. Mean spread of sensory analgesia in the non-dependent side on the dural puncture was significantly higher in the sagittal-tilt group (T7.4 +/- 2.6 at 15 min) compared the horizontal group (T9.5 +/- 1.4 at 15 min). There was no significant difference in the mean cephalad spread of the analgesic level in the dependent side between the two groups. Unilateral motor anesthesia of the dependent side seemed to be canceled by the sagittal tilting maneuver. A 7 to 8 degree tilt in the sagittal plane is recommended to facilitate the cephalad spread of analgesia and to avoid unilateral anesthesia. PMID- 15278520 TI - Systemic administration of procaine suppresses the somato-sympathetic reflex discharges in anesthetized cats. AB - Somato-sympathetic reflex discharges (SSRDs) which were induced from the lumbar sympathetic truck elicited by train pulse stimulation of femoral nerves in anesthetized cats were examined during and after intravenous infusion of 1 mg.kg( 1).min(-1) of procaine. The infusion demonstrated that amplitudes were suppressed with prolonged latencies and/or their tendencies in A- and C-reflex potentials of SSRDs. Heart rate and blood pressure decreased slightly, and the decreases correlated with time courses of suppressed SSRDs. These results suggest that the systemic procaine infusion may be a reliable method to alleviate somato sympathetic reflexes under such conditions as extreme stimulation of somatic afferent nerves observed in surgical procedures. PMID- 15278521 TI - Effects of surgical site and inspired gas warming devices on body temperature during lower abdominal and thoracic surgery. AB - To evaluate the effects of surgical site and inspired gas warming and humidifying devices on body temperature, we studied rectal, tympanic membrane, and esophageal temperature changes in 48 patients. The patients were divided into 4 groups (n = 12), according to surgical site, lower abdominal surgery and thoracic surgery, and according to the warming device used, heat and moisture exchanger (ThermoVent 600) and heated humidifier (Cascade 1). The heated humidifier was controlled to warm inspired gases to about 35 degrees C. All body temperatures fell significantly during surgery. There was no difference in the tympanic membrane and esophageal temperature declines between the two surgical sites, but the decline in rectal temperature was larger in the lower abdominal surgery than in the thoracic surgery. At the end of surgery, all temperatures returned to the value before surgery, and the rectal and tympanic membrane temperatures even exceeded them. There was no difference between the effects of the ThermoVent 600 and Cascade 1. These results suggest that rectal temperature is influenced by the ambient temperature during lower abdominal surgery and that warming and humidifying devices for inspired gases do not prevent, but can restore the decline in body temperature during lower abdominal and thoracic surgery. The heated humidifier showed no advantage over the heat and moisture exchanger in our study. PMID- 15278522 TI - Blood pressure, heart rate and catecholamine response during fiberoptic nasotracheal intubation under general anesthesia. AB - Arterial blood pressure (ABP) and heart rate were recorded at one-minute intervals during several stages of intubation in the fiberscope group and the laryngoscope group, to determine if fiberoptic nasotracheal intubation would result in fewer hemodynamic and catecholamine responses than when intubation was performed with a Macintosh laryngoscope. Blood samples were also taken to measure plasma catecholamine concentration immediately after intubation with the fiberscope. The mean ABP in the laryngoscope group was slightly greater than that of the fiberscope group for 4 min after intubation. Heart rates at 2 min and 4 min after intubation in the laryngoscope group were significantly greater than those for the fiberscope group. Even immediately after intubation, the mean plasma levels of epinephrine and norepinephrine were unchanged in the fiberscope group. Arterial oxygen saturation (Sp(O)(2)) was maintained within normal range during both of intubation procedures, although the time required for intubation was longer than in the laryngoscope group. Other cardiovascular complications were more common in the laryngoscope group than in the fiberscope group. These results suggest that fiberoptic intubation results in less severe stress than does laryngoscopic intubation. Fiberoptic intubation should therefore be used not only in patients with difficult airway, hypertension, ischemic heart disease, or cerebrovascular atherosclerosis, but also it is recommended for all patients for whom nasotracheal intubation is indicated. PMID- 15278523 TI - The effects of feeding on the development of metabolic acidosis in the rat: comparison between perfused liver in situ and whole animal. AB - Rat liver perfused in situ was charged with two concentrations of halothane. Lactate increased in a time- and dose-dependent manner in the liver of the fed rat, whereas its increase in the starved rat was much milder (1.7 --> 9.2 mmol. l(-1) vs 0.6 --> 1.4 mmol. l(-1) after a 3 hr charge with 6.0% halothane). Base excess decreased also more markedly in the fed rat. Glucose increased 2.3 times the control value in the fed rat, whereas it did not change significantly in the starved rat. Changes produced by enflurane were very similar to those produced by halothane. It was inferred that in the presence of halothane and enflurane, hepatic glycogen was transformed into glucose and then to lactate by the inhibition of NADH dehydrogenase. In the liver of the starved rat, glucose, hence lactate, did not increase because of the depletion of glycogen. When halothane (1.9%) was given to the whole animal, changes in lactate, base excess and glucose in the arterial blood were very mild. Marked disparities in these parameters between the two experimental models were inferred to be due to: 1) possible insinuation of anaerobic metabolism in the perfusion experiments, 2) a well-kept balance between the suppression of cellular metabolic activity and inhibition of energy production by halothane in the whole animal, and 3) involvement of neural and humoral factors in the intact whole animal. PMID- 15278524 TI - Tracheostomy for a patient of sleep apnea syndrome in an emergency situation. PMID- 15278525 TI - Anesthesia and perioperative management for Kasabach-Merritt syndrome--report of a case. PMID- 15278526 TI - Greater than normal variability of Ca-induced Ca release in muscle fibers of a patient with a positive family history of malignant hyperthermia. PMID- 15278527 TI - Anesthesia in Shy-Drager syndrome. PMID- 15278528 TI - Recurrent coronary artery spasm during a non-cardiac surgical procedure. PMID- 15278529 TI - Spinal morphine injections for treatment of post-spinal headache. PMID- 15278530 TI - Is there a connection between postsurgical infections after cardiac surgery and the sort of anesthesia used? PMID- 15278531 TI - Is it safe to use trans-epidural spinal cord stimulating needle? PMID- 15278533 TI - Recombinant human-type SOD attenuates circulatory disorders after reperfusion of splanchnic organs in rats. AB - Oxygen free radicals (OFRs) have been reported to play pivotal roles in the pathogenesis of cell damage induced by ischemia and reperfusion. The efficacy of recombinant human superoxide dismutase (rh-SOD) in the treatment of circulatory disorders after reperfusion of the splanchnic area was investigated in rats. All rats died within 3 hours after release of 60-min superior mesenteric artery occlusion (SMAO) when no treatment was given. Animals which received rh-SOD, 2 mg.100 g(-1)BW, at reperfusion followed by a continuous infusion of rh-SOD 0.67 mg.100 g(-1)BW.hr(-1), exhibited prolonged survival times compared with no treatment rats (231 +/- 35 min and 149 +/- 43 min, respectively). Mean blood pressure in rats treated with rh-SOD was higher than in controls after reperfusion, and was concomitant with improvement in splanchnic perfusion. The results suggest excessive activity of OFRs in reperfused organs and a possible scavenging effect of rh-SOD as a means of eliminating them. PMID- 15278534 TI - Hypoglycemia enhances bupivacaine-induced cardiotoxicity in the rat. AB - The effect of blood glucose concentration on bupivacaine-induced cardiotoxicity was investigated in normoglycemic and hypoglycemic adult rats and compared to that of equipotent doses of lidocaine. The anesthetic agents were injected intraperitoneally into tracheostomized animals anesthetized with ketamine. ECG and direct blood pressure measurements were recorded continuously. Femoral arterial blood was used for determinations of glucose level, potassium concentration and base deficit values. Blood was drawn from the heart at the time of death for local anesthetic levels. In hypoglycemic animals, bupivacaine rapidly produced serious dysrhythmias leading to asystole. In normoglycemic rats, only ST-segment changes followed bupivacaine injection and death ensued from hypoxemia secondary to respiratory failure. With lidocaine, both hypoglycemic and normoglycemic rats died of hypoxemia following respiratory paralysis without antecedent dysrhythmias. Thus, hypoglycemia enhanced the cardiac effects of bupivacaine but not those of lidocaine. PMID- 15278535 TI - Cardiovascular responses to fiberoptic intubation: a comparison of orotracheal and nasotracheal intubation. AB - We compared the cardiovascular responses between nasal and oral intubation with a fiberoptic bronchoscope under the combination of neuroleptic analgesia (NLA) and topical anesthesia. The 16 patients studied were divided into 2 groups: the nasal intubation group (N group: 8 patients) and the oral intubation group (O group: 8 patients). There were significant changes in systolic, diastolic and mean arterial pressures in the N group and in the pressure rate quotient in the O group. Diastolic arterial pressure and heart rate were significantly higher in the N group than in the O group before induction of general anesthesia. The rate pressure product (RPP) was significantly higher in the N group than in the O group at some points during the procedure. The individual RPP in both groups was relatively stable except for one patient in the N group, who had a marked increase in RPP during the procedure. We conclude that, under the combination of NLA and topical anesthesia, the cardiovascular responses to oral fiberoptic intubation are less severe than those to the nasal approach. The oral approach is recommended, especially in patients with coronary artery disease, taking into consideration of the cardiovascular responses to fiberoptic intubation. PMID- 15278536 TI - The effects of urinastatin on the plasma levels of granulocyte elastase during open heart surgery under simple deep hypothermia. AB - Changes in granulocyte elastase (GLE) and Beta-gluculonidase (Beta-gl) were observed during open heart surgeries which were performed under deep hypothermia with surface cooling. In addition, the effect of urinary trypsin inhibitor, urinastatin, on the activities of these enzymes was studied. The patients were divided into three groups, namely group U-I with intravenous injection of 6000 u.kg(-1) of urinastatin before cooling, group U-II administered with an additional 6000 u.kg(-1) after warming to 30 degrees C, and an untreated group (Group C). The plasma level of GLE increased significantly in the three groups compared with the level before cooling respectively. In the group U-II, the GLE level after the warming was lower than that in the control group. The serum level of Beta-gl increased significantly in the three groups at the end of rewarming (36 degrees C). The release of GLE from lysosomes in granulocytes was inhibited in the group U-II. The insufficient inhibition of GLE release in the group U-I is probably due to relatively short half-life of urinastatin. Therefore double administration of 6000 u.kg(-1), before and after the cooling, may be required to achieve the therapeutic effect. Consequently, urinastatin appears to be useful in open heart surgery under deep hypothermia with surface cooling. PMID- 15278537 TI - Interactions between ORG9426 and other non-depolarizing neuromuscular blocking agents in rats in vivo. AB - In this study, combined neuromuscular blocking effects of ORG9426 with other non depolarizing neuromuscular blocking agents were investigated. About 20% steady state neuromuscular block was established by a continuous infusion of one of 6 neuromuscular blocking agents (ORG9426, vecuronium, pancuronium, pipecuronium, d tubocurarine and metocurine). Then 1/7 of the ED50 of ORG9426 or one of other neuromuscular blocking agents was administered in a single injection, and the increase in the neuromuscular block was observed. The combined neuromuscular blocking effect of ORG9426 and d-tubocurarine or ORG9426 and metocurine was significantly ( P < 0.05) greater than that of each corresponding control (the combination of same neuromuscular blocking agent). The effect of d-tubocurarine was also potentiated by vecuronium, pancuronium and pipecuronium. These potentiations were not observed between ORG9426 and pancuronium, pipecuronium or vecuronium. Possible mechanisms of these synergistic interactions were discussed. PMID- 15278538 TI - Cardiac and hepatic metabolism in spontaneously hypertensive rats following acute blood loss. AB - Seven spontaneously hypertensive rats (SHRs) and eight Wistar-St rats were used to assess the influence of hemorrhage on myocardial and hepatic energy metabolism. They received 2% halothane and pancuronium, 0.3 mg.kg(-1), during preparation. After discontinuation of halothane, blood (2 ml.100 g body weight( 1)) was gradually withdrawn over a 5 min period from a femoral artery. Thirty min after induction of hemorrhage, the heart and liver were removed and myocardial and hepatic metabolites (ATP, lactate, pyruvate and glycogen) were measured by the enzymatic methods. Acidosis and decreased hematocrit were noted in the both groups after hemorrhage. Mean arterial pressure (MAP) in SHR was significantly higher than that in Wistar rat before hemorrhage. However, there were no significant differences in MAP and heart rate between the two groups after hemorrhage. Although there were no significant differences in cardiac metabolites, a significant decrease of hepatic ATP and an increase of hepatic lactate/pyruvate ratio were found in SHR when compared with Wistar rat. These results suggest that human hypertensive disease may run a high risk in connection with acute hemorrhage. PMID- 15278539 TI - Epidural administration and analgesic spread: comparison of injection with catheters and needles. AB - This study was designed to investigate differences in epidural analgesic spread between catheter and needle injections in 48 patients with comparable physical characteristics. The spread of analgesia in the catheter injection group with a 0.24 ml.sec(-1) injection rate (n = 16) was 16.8 +/- 1.5 spinal segments and that in the needle injection group at the same injection rate (n = 16) was 12.5 +/- 1.8 spinal segments ( P < 0.01). Needle injection at the faster rate of 1.2 ml.sec(-1) (n = 16) produced a significantly greater spread of analgesia than with the 0.24 ml.sec(-1) rate through the needle (16.2 +/- 1.6 vs 12.5 +/- 1.8 spinal segments, P < 0.01). Thirteen of 16 patients receiving the fast needle injection complained of back compression or discomfort during the injection. The injection through an epidural catheter and the fast (1.2 ml.sec(-1)) injection through a needle produced extensive and equivalent epidural analgesic spread. However, because of patients discomfort with fast injection through the needle, the authors conclude that when using continuous epidural anesthesia, the initial injection of local anesthetic should be administered through the epidural catheter not the needle. PMID- 15278540 TI - Prostaglandin E1 reduces blood loss during and after resection of lumbar herniated disc. AB - Controlled hypotension was employed during resection of lumbar herniated disc on 10 patients. Prostaglandin E(1) (PG) was used as a hypotensive agent. The systolic blood pressure was lowered less than 100 mmHg in the hypotensive group. The average blood loss during surgery was 95 +/- 41 ml for the hypotensive group compared with 154 +/- 81 ml for the normotensive group ( P < 0.05). The blood loss after surgery was also significantly less in the hypotensive group than in the normotensive group ( P < 0.05). We conclude that PG is an effective hypotensive agent on blood loss during and after surgery. PMID- 15278541 TI - Analysis of oxygen transport to the brain when two or more parameters are affected simultaneously. AB - We composed a model, combining oxygen transport system from blood to tissue with the oxygen consumption system at the tissue. The aim of this study is to apply it to the brain tissue under conditions when two or more oxygen transport parameters are affected simultaneously. The following values were assumed. Critical tissue P(O)(2) (Pcrit(O)(2)) 2 mmHg; oxygen consumption above this level 3 ml.min( 1).100 g(-1); diffusion coefficient from blood vessel to tissue (Dvt) 0.2 ml.min( 1).mmHg(-1).100 g(-1); cerebral bloow flow (CBF) 50 ml.min(-1).100 g(-1); hemoglobin 15 g.100 ml(-1). The Hill equation was used for oxygen dissociation curve with n of 2.7 and P(50) of 27.0 mmHg. The changes of oxygen consumption of the brain (V(O)(2)) were analyzed when 2 or more of 5 parameters, Pa(O)(2), CBF, Dvt, P(50) and hemoglobin decreased simultaneously from their respective normal values. As the number of parameters affected increased, the level at which oxygen consumption begins to be affected became higher. With all five parameters combined, a reduction down to 78 per cent of normal resulted in tissue hypoxia. We conclude that the oxygen consumption of the brain is fairly resistant when only one parameter is affected, but it becomes increasingly vulnerable when several parameters are affected simultaneously. A clinically important finding is that the brain is particularly vulnerable to a combination of hypocapnia and a decreased level of 2,3DPG. PMID- 15278542 TI - Does propofol enhance GABA-mediated inhibition? AB - The effects of propofol on synaptic transmission were characterized and compared with pentobarbital in the rat hippocampal slice preparation. Hippocampal CA1 population spike after stimulation of Schaffer collaterals indicated that the postsynaptic response was primarily mediated by non-N-methyl-D-aspartate class glutamate receptors since it was abolished by the presence of 6,7 dinitroquinoxaline-2,3-dione (DNQX). Propofol and pentobarbital depressed CA1 population spike amplitude in a dose dependent fashion. Dose-response curves for population spike amplitudes were determined for propofol and pentobarbital and the concentrations producing a half-maximum response (ED(50)) were 110 microM and 160 microM for propofol and pentobarbital, respectively. By contrast, when GABA A mediated inhibition was blocked by addition of 100 microM pictotoxin, propofol, in concentrations up to 400 microM had no significant effect on population spike amplitudes. These results suggest that propofol attenuates synaptic transmission in the central nervous system in part by enhancing GABA A-mediated inhibition and not by depressing glutamate-mediated excitation, as occurs with pentobarbital. PMID- 15278543 TI - Adsorption of sevoflurane by soda-limes. AB - Dry soda-lime adsorbs significant quantities of halothane, thus influencing on the speed of the induction of anaesthesia with the agent and also on the recovery from anesthesia. Sevoflurane is a new inhaled anesthetic. Although the chemical degradation of sevoflurane with soda-lime has been studied, no information is available about its adsorption by soda-lime. This issue can not be neglected clinically. Two different soda-limes were placed in saturated vapour of sevoflurane for 17 hours to weight adsorbed sevoflurane. Then soda-limes adsorbing sevoflurane was sealed in a test tube after air-drying for 1) 0 min, 2) 10 min, 3) 30 min and 4) 17 hours. The vapour phase of sevoflurane in the test tube at various temperatures were determined using gas chromatography. Sevoflurane vapour concentrations in the test tubes increased in a temperature dependent manner. Those in the conventional soda-lime were higher than those in the new soda-lime under any experimental conditions. Sevoflurane was released from soda-limes even after air-drying for 17 hours. These results show that much amount of sevoflurane is adsorbed by soda-limes and is released easily in the air. Thus there is a possibility for our patients to inhale unexpected inhaled anesthetics, if we use our anesthetic machine repeatedly. PMID- 15278544 TI - Efficacy of thoracic sympathetic ganglion block and prediction of complications: clinical evaluation of the anterior paratracheal and posterior paravertebral approaches in 234 patients. AB - In the 10 years from 1980 to 1989, a total of 234 patients underwent 557 thoracic sympathetic ganglion blocks. The block was performed by the anterior paratracheal approach in 129 cases and by the posterior paravertebral approach in 428 cases. The procedures for using these two approaches are presented here. The efficacy of thoracic sympathetic ganglion blockade was evaluated as follows; marked efficacy was defined by the complete control of sweating in the palms, moderate efficacy was defined by a decrease in palmar sweating which persisted for at least one week, and minor efficacy was defined by a decrease in sweating followed by recurrence of hyperhidrosis within one week with maintenance of palmar warmth. in addition, the results were retrospectively reviewed in relation to the age and sex of the patients, the technique used, the laterality of the block, the disease treated, the doses of local anesthetic and neurolytic agents, and the number of blocks. The posterior approach was significantly more successful than the anterior approach, and the treatment of both T2 and T3 by the posterior approach was significantly more effective than the treatment of either nerve alone by the same approach ( P < 0.01). The efficacy rate was significantly lower for hyperhidrosis than for the other diseases ( P < 0.01). Complete cessation of hyperhidrosis was significantly less common in the over-60 age group ( P < 0.01). Regarding the dose of neurolytic, the complete cessation of hyperhidrosis was achieved significantly more frequently with doses of 2.5 ml or higher than with lower doses ( P < 0.01) when both T2 and T3 wee treated by the posterior approach. A dose-dependent response if hyperhidrosis was noted at dose levels higher than 2.5 ml. Thoracic sympathetic ganglion blockade was only occasionally associated with complications, and no serious complications were observed. Before injecting the neurolytic agent, a mixture of contrast medium and local anesthetic was injected to determine the three-dimensional distribution of the contrast and to assess the scope of the analgesia produced by the local anesthetic. Significant complications could thus be avoided. PMID- 15278545 TI - Neuromuscular pharmacology update. PMID- 15278546 TI - Vecuronium and danger of vagal induced cardiac arrest during laparotomy: a case report and literature review. PMID- 15278547 TI - Anesthetic management of a child with Maroteau-Lamy syndrome. PMID- 15278548 TI - Hypothermia associated with intrathecal morphine. PMID- 15278549 TI - Paraplegia associated with hyperthermia during repair of coarctation of the aorta. PMID- 15278550 TI - Coronary artery spasm immediately following extubation of the trachea. PMID- 15278551 TI - Bilateral pneumothoraces after transtracheal high frequency jet ventilation during emergency tracheostomy. PMID- 15278552 TI - Hemomediastinum induced by heparinization after stellate ganglion blockada. PMID- 15278553 TI - Evaluation of gastric tube with esophageal thermister (Thermosump). AB - The accuracy and the feasibility of esophageal temperature measured by a new gastric tube. Thermosump, which is incorporated with a esophageal thermister, was evaluated in anesthetized dogs (n = 6) and men (n = 59). In dogs, esophageal temperature measured by Thermosump was correlated well with the temperatures measured by the conventional esophageal thermister, and also by the pulmonary artery catheter (r = 0.98, 0.98, respectively). In anesthetized men, correlation between esophageal temperature by Thermosump and rectal, or bladder temperature was good during surgery of extremities (r = 0.81, 0.80, respectively). But during abdominal surgery, correlation between esophageal and bladder temperature was relatively poor (r = 0.50). Insertion of the tube, and suction of gastric fluid through the tube were easy without any complication. This newly developed gastric tube with a esophageal thermister was safe, and useful for measuring esophageal temperature. PMID- 15278554 TI - Distribution of hypobaric tetracaine within cerebrospinal fluid in a spinal canal model. AB - We studied the distribution of hypobaric tetracaine within cerebrospinal fluid (CSF) using a spinal canal model to examine the spread of a hypobaric anesthetic solution during spinal anesthesia. In our study, 0.2% tetracaine colored with methylene blue was observed to migrate upwards rapidly and spread horizontally in the upper portion of the model placed horizontally and filled with CSF which was collected from several neurosurgical patients. The boundary between the hypobaric solution and CSF could be clearly identified. These results suggest that the hypobaric tetracaine will distribute in the upper portion of the spinal canal during spinal anesthesia. It can be used to produce unilateral spinal blockade in the lateral decubitus position despite a small difference in specific gravity between the hypobaric anesthetic and CSF. In addition, the fact that the hypobaric solution showed a rapid horizontal spread suggests that correct positioning both during and following administration of the anesthetic is important to control the level of anesthesia. PMID- 15278555 TI - A device for tracheal tube during CO2 laser irradiation in laryngomicrosurgery. AB - We devised that the segment of commercially available defensor II tube coming in contact with the vocal cord was concaved. We used this new tube during CO(2) laser irradiation in laryngomicrosurgery. We came to the conclusion that it was much more superior to the conventional tube in safety and resistance of the material to Co(2) laser irradiation and in increase of the operation field. PMID- 15278556 TI - Preventative effect of PGE1 for postoperative liver damage. AB - The efficacy of a low dose of PGE(1)-use on the postoperative liver damage was evaluated. PGE(1) was infused in with the mean rate of 0.026 microg.kg(-1).min( 1) during surgical procedure to 93 patients under GO-enflurane anesthesia (the PG). Serum GOT, GPT and total bilirubin (TBIL) values measured before, at the end of (End) and 3 days (3d) after the operation were compared to those obtained from 43 patients without PGE(1) administration (the control). This dose of PGE(1) did not change blood pressure and heart rate, but slightly decreased Pa(O)(2). In patients with preoperative normal values of GOT, GPT and TBIL, increases in GOT, GPT and TBIL observed at End in the PG were significantly lower than those in the control (31.9 vs 72.2 IU, 25.9 vs 61.9 IU, 0.68 vs 0.83 mg.dl(-1), respectively). GOT, GPT and TBIL at 3d significantly increased in both groups, and these levels were identical between the two groups. In patients with preoperative abnormal values, only GOT at End increased in both groups, while no significant difference between the PG and the control group was noted. GOT at 3d and GPT at End and 3d did not significantly changed in either group. These results suggest that the low dose of PGE(1) administered during an operation prevents the development of postoperative liver damage, but does not treat the damaged hepatic cells. PMID- 15278557 TI - Duration of action of supplemental doses of vecuronium is related to the duration after the initial dose. AB - Vecuronium was administered in an initial dose of 0.1 approximately 0.3 mg.kg(-1) and in supplemental doses of 0.03 mg.kg(-1) or 0.05 mg.kg(-1) in 74 patients (ASA class 1 or 2) scheduled for abdominal surgery. The duration of the neuromuscular blockade provided by vecuronium after both the initial and supplemental doses was determined using the evoked integrated electromyographic device. A statistically significant positive correlation (correlation coefficient: 0.83 approximately 0.91) was found between the duration of action of the initial dose and that of the first to fourth supplemental doses. The regression lines of each of the first four supplemental doses to the initial dose were very similar to each other. These results suggest that, since the duration of action of supplemental doses of vecuronium was prolonged in patients showing a long duration of action of the initial dose, it would be wise to avoid blind adherence to a predetermined schedule for supplemental administration. Rather, anesthesiologists should take into account the patient's response to the initial dose and then decide the most appropriate timing for supplemental doses. Moreover, since vecuronium shows little cumulative effect even after 4 supplemental administrations in clinical range doses, it can be concluded that vecuronium can be safely used in a wide dose range. PMID- 15278559 TI - Calcium entry blocker nicardipine inhibits sodium and inorganic phosphate reabsorption independent of renal circulation in dogs. AB - The effects of nicardipine on renal function were studied in anesthetized dogs. The changes in the tubular sodium (Na) and inorganic phosphate (PO(4)) reabsorption caused by the drug infusion into the renal artery without altered systemic and real circulation were especially evaluated. In dogs receiving a smaller dose of nicardipine (5 ng.kg(-1).min(-1)) into the left renal artery the blood pressure and renal circulation did not change, but urine volume and urinary Na and PO(4) excretion increased significantly. In dogs receiving a larger dose of nicardipine (50 ng.kg(-1).min(-1)) into the renal artery, renal plasma flow, urine volume and urinary Na and PO(4) excretion increased significantly, but creatinine clearance did not. The fractional distal Na reabsorption did not change with nicardipine infusion in either group. PO(4) reabsorption is considered to occur mainly in the proximal renal tubule, so its appearance in urine in increased quantities without the changes of systemic and renal circulation suggests proximal activity of the drug. PMID- 15278558 TI - Changes in erythrocyte membrane fluidity by endotoxin in rats. AB - The effect of endotoxin on fluidity and lipid composition of the erythrocyte membrane was studied in rats following the intraperitoneal administration of endotoxin (30 mg.kg(-1) body weight). Erythrocyte membrane fluidity measured with 16-stearic acid spin label (16-SAL) was significantly decreased in the endotoxin treated rats as compared with control. A decrease of lysophosphatidylcholine in the membrane lipid was evident in the endotoxin-treated rats. The cholesterol to phospholipid molar ratios and other phospholipid fractions did not differ significantly in the two groups. The levels of plasma Beta-glucuronidase activity and lipoperoxide were significantly increased in the endotoxin-treated rats when compared to controls. There were significant correlations between the parameter of 16-SAL in erythrocytes and plasma Beta-glucuronidase activities or lipoperoxide from both endotoxin-treated and control groups, P < 0.005 or P < 0.02 respectively. In conclusion endotoxin decreased rat erythrocyte membrane fluidity in vivo. Since membrane fluidity is closely related to the vital functions of the membranes, the change described could be related to the abnormality of cell membrane functions in endotoxin shock state. PMID- 15278560 TI - Changes in plasma catecholamine levels following injection of prostaglandin F2alpha into the basal cistern in rabbits. AB - We measured plasma epinephrine and norepinephrine concentrations in a rabbit model simulating subarachnoid hemorrhage (SAH), following the injection of prostaglandin F(2alpha) (PGF(2alpha)) into the basal cistern. In this model, plasma epinephrine values increased significantly (to 4.2-fold those before injection), substantially more than norepinephrine (which increased 1.3-fold) at 5 minutes (min) after PGF(2alpha) injection. Dissection of autonomic outflow from the cervical spinal cord or ligation of the suprarenal veins reduced the changes in plasma catecholamine concentrations associated with PGF(2alpha) injection. These results suggest that the sympathetic discharge seen after PGF(2alpha) injection into the basal cistern in rabbits occurred through the sympatho-adrenal pathways. PMID- 15278561 TI - Effects of immobilization stress and of a benzodiazepine derivative on rat central dopamine system. AB - The effects of a novel benzodiazepine derivative, Ro 16-6028 on rat brain dopamine system were examined under stress and non-stress conditions. Thirty minutes restraint stress increased dopamine synthesis in two dopamine neuron regions, prefrontal cortex and nucleus accumbence. Ro 16-6028 inhibited potently dopamine synthesis in prefrontal cortex, nucleus accumbens and striatum in dose dependent manner under non-stress condition. Furthermore, Ro 16-6028 reverses the stress-induced augmentation of the synthesis in prefrontal cortex. These findings indicate that Ro 16-6028 has an anxiolytic profile and that central dopamine system plays an important role in stress reaction. PMID- 15278562 TI - Effects of isoflurane and halothane on the calcium ion-tension curve in rat myocardium. AB - In order to clarify the interaction of volatile anesthetics and extracellular calcium ion on the myocardial contraction, effects of both isoflurane (1.0%) and halothane (0.5%) on the extracellular calcium ion concentration ([Ca(2+)](O)) tension curve were studied. Increasing [Ca(2+)](O) enhanced the myocardial contraction response, and the maximal response was obtained at [Ca(2+)](O) of 3.0 mM. Halothane depressed the maximal value of the tension development in response to increasing [Ca(2+)](O), while isoflurane did not ( P < 0.01). The probit response of the developed tension to the changes in [Ca(2+)](O) indicated that isoflurane increased the median effective concentration (EC(50)) of [Ca(2+)](O) significantly from 0.484 +/- 0.051 (mean +/- SEM) to 0.870 +/- 0.056 mM ( P = 0.001), but halothane did not ( P = 0.018). Therefore, 1.0% isoflurane was concluded to move the [Ca(2+)](O)-tension curve to the right, while a downwards shift occurred with 0.5% halothane. PMID- 15278563 TI - Potentiation of the cardiovascular effects of nicardipine by enflurane anesthesia in canine blood-perfused heart preparations. AB - The cardiovascular interaction between nicardipine (N) and enflurae (E) was examined with blood perfused isolated paplillary muscle preparations (PMP) and sinoatrial node preparations (SNP). Blood flow to these preparations was supplied by either conscious or 1.7% E-anesthetized-donor dogs. N was administered continuously at a rate of 2.0 microg.kg(-1).min(-1) into the donor dogs for 60 min. Measurements were as follows: mean arterial blood pressure (MAP), heart rate (DHR), PQ interval (PQ) in the electrocardiogram, developed tension (DT) in PMP, sinoatrial rate (SAR) in SNP and blood flow (BF) to PMP and SNP. There were no significant differences in PQ, SAR and BF between two groups. However, N further decreased MAP, DHR and DT that were already decreased by E significantly. The authors conclude that the cardiovascular interaction between N and E was generally additive but that concerning negative inotropism was synergistic. PMID- 15278564 TI - Retrospective study of post-anesthetic mild liver disorder associated with inhalation anesthetics, halothane and enflurane. AB - The incidence of post-anesthetic mild liver disorder (PAMLD) was compared between 928 patients administered halothane and 1766 patients administered enflurane. They were selected from 19 504 surgical patients administered general anesthesia at Kyushu University Hospital over the past 6 years and 4 months. They had had normal liver function before operation and had no history of blood transfusion. Alanine aminotransferase (ALT) levels exceeding 70 IU. l(-1) within 180 days after operation were found in 226 patients in the halothane group (24.4%), and in 250 patients in the enflurane group (14.2%) ( P < 0.01). Both maximum ALT levels and duration of ALT elevation were higher and longer in the halothane group ( P < 0.01). These results suggest that, not only in the development of fulminant hepatitis but also in PAMLD, enflurane is less hepatotoxic than halothane. PMID- 15278565 TI - What anesthesiologist should know about neuromuscular monitoring today? PMID- 15278566 TI - A cardiovascular collapse during cemented total hip replacement in a diabetic patient. PMID- 15278567 TI - A case of thyroid crisis occurring during surgery. PMID- 15278568 TI - Anesthetic management of combined caesarean section and pheochromocytoma removal. PMID- 15278569 TI - Anesthetic management for a patient with amyloidosis. PMID- 15278570 TI - Abdominal total hysterectomy in a patient with isolated angiitis of the central nervous system. PMID- 15278571 TI - Corticosteroid effect on down regulation in beta adrenergic receptors. PMID- 15278572 TI - Fiberoptic evaluation of peripheral airways of two patients with acute respiratory failure during mechanical ventilation. PMID- 15278573 TI - Hyperpotassemia during major vascular surgery: a possible indicator of visceral infarction. PMID- 15278574 TI - Oxygen uptake and carbon dioxide elimination during epinephrine-induced arrhythmias in humans. PMID- 15278575 TI - Programming a predictive formula for angina and other risk factors in patients with cardiac diseases undergoing noncardiac operations. AB - We programmed a formula which predicts the incidence of either myocardial infarction or cardiac death during the postoperative period. The original formula was proposed by Shah et al, based on their own data and analysis. The program is simple and is written in a language called Quick Basic. The use of this program is also simple. Such a program has improved the use of this analysis substantially. The program has been posted on to a few Computer network services as a free software. PMID- 15278576 TI - The neuromuscular effects of sevoflurane and isoflurane alone and in combination with vecuronium or atracurium in the rat. AB - Sevoflurane was compared to isoflurane anesthesia alone and in combination with atracurium or vecuronium in 84 rats using the sciatic nerve-anterior tibialis muscle preparation. Both bolus injection and infusion rate techniques were used to evaluate these drug interactions. The ED(50) (dose which produced a 50% depression of twitch tension) of atracurium was 311 +/- 31 and 360 +/- 32 microg.kg(-1) during 1.25 MAC sevoflurane and isoflurane anesthesia respectively. The ED(50) of vecuronium was 190 +/- 27 and 149 +/- 14 microg.kg(-1) during 1.25 MAC sevoflurane and isoflurane anesthesia respectively. The mean infusion rates of atracurium and vecuronium required to maintain a 50% depression of twitch tension were 5.04 +/- 0.7 and 2.02 +/- 0.3 mg.kg(-1).hr(-1). These infusion rates were 5.04 +/- 0.7 and 2.02 +/- 0.3 mg.kg(-1).hr(-1) during 1.25 MAC sevoflurane and 3.73 +/- 0.3 and 1.81 +/- 0.4 mg.kg(-1).hr(-1) during 1.25 MAC isoflurane anesthesia respectively. With both atracurium and vecuronium, the infusion rate required to maintain a 50% depression twitch of tension was inversely related to the concentrations of isoflurane and sevoflurane. The authors conclude that sevoflurane is similar in potency to that of isoflurane in augmenting a vecuronium or atracurium induced neuromuscular blockade in a dose-dependent manner. PMID- 15278577 TI - Changes in end-tidal CO2 level following tourniquet deflation during orthopedic surgery. AB - We studied the changes in end-tidal CO(2) (ET(CO)(2)) and systemic responses after tourniquet deflation in spontaneously breathing and ventilation-controlled patients during orthopedic surgery of both the upper and/or the lower extremities. In most patients, increases in ET(CO)(2), heart rate, and Pa(CO)(2), as well as decreases in blood pressure and pH were observed. In every spontaneously breathing patient, the respiratory rate began to increase before the ET(CO)(2) reached a maximum. Arterial blood gas analysis suggested that the increase in ET(CO)(2) closely reflected the increase in Pa(CO)(2). Our study yielded new information on the ET(CO)(2) changes as follows: 1) the time for ET(CO)(2) level to reach a peak (peak time) was almost constant despite the considerable differences in the increases in ET(CO)(2) both in spontaneous breathing and ventilation-controlled groups and the peak time in the former group was shorter than that in the latter group; and 2) it was suggested that the increase in ET(CO)(2) in the spontaneously breathing patients was smaller than that in ventilation-controlled patients when both patients were subjected to the same conditions on tourniquet time and tourniqueted area. Our data showed that the increase in ET(CO)(2) (or Pa(CO)(2)) can be large and prolonged in some situations. Thus, we recommend continuous ET(CO)(2) monitoring and the proper hyperventilation at tourniquet deflation in order to minimize any adverse effects of acidosis. PMID- 15278578 TI - Premedication with metoclopramide decreases the frequency of methohexital induced hiccup. AB - Metoclopramide is one of many drugs that have been recommended for the treatment of intractable hiccup. Methohexital may produce hiccup during induction of general anesthesia. 211 women received methohexital for induction and maintenance of general anesthesia for short gynaecological procedures. All the patients were premedicated with fentanyl, diazepam and atropine. 109 patients were randomly selected to receive metoclopramide before induction of anesthesia; the remaining 102 patients served as a control group, and were anesthetized without metoclopramide premedication. The frequency of hiccup was compared between the two groups. 7 patients had hiccup in the metoclopramide premedicated group, as compared to 17 patients in the control group. This difference was statistically significant. We conclude that metoclopramide reduces the frequency of methohexital induced hiccup, and recommend that metoclopramide be routinely used for the premedication of methohexital injection. PMID- 15278579 TI - Anesthetic management with eptazocine hydrobromide in patients receiving long term antipsychotic medication. AB - We evaluated the usefulness of eptazocine hydrobromide as an adjuvant in patients receiving antipsychotics for long periods. Patients anesthetized with enflurane alone (enflurane group, n = 11), were compared with those anesthetized with enflurane and eptazocine hydrobromide 1 mg.kg(-1) (eptazocine group, n = 10). The mean daily dose of the antipsychotics, converted into the amount of chlorpromazine, was 345 mg in the eptazocine group and 366 mg in the enflurane group. The duration of antipsychotic medication was 14 years in the eptazocine group and 17 years in the enflurane group. The maintenance concentration of enflurane was 0.37% in the eptazocine group and 0.67% in the enflurane group, being significantly lower in the former group. The interval between the termination of operation and removal of the endotracheal tube was slightly shorter in the eptazocine group. The discriminant function of circulatory stability obtained from the measurement of systolic blood pressure and heart rate during anesthesia in the eptazocine group was 481, being significantly lower than 539 in the enflurane group. Both absolute and relative instabilities of systolic blood pressure and heart rate were slightly smaller in the eptazocine group. No side effects associated with eptazocine hydrobromide administration were observed. These results suggest the safety and usefulness of this analgesic in the anesthetic management of patients receiving long-term antipsychotic medication. PMID- 15278580 TI - Pharmacokinetics and pharmacodynamics of vecuronium bromide. AB - The pharmacokinetics and pharmacodynamics of vecuronium bromide were studied in patients under general anesthesia of enflurane and nitrous oxide in oxygen. Eighteen patients were randomly divided into two groups which received either 0.05 mg.kg(-1) (low dose group) or 0.20 mg.kg(-1) (high dose group) of vecuronium intravenously. The plasma concentration of vecuronium was determined by high performance liquid chromatography. The neuromuscular blocking effect was assessed by measuring the twitch tension of the adductor pollicis muscle elicited by supramaximal electrical stimulation. Pharmacokinetic analysis was carried out using a two compartment model. The relationship between the T4/T1 and T1/ control T1 ratios differed during onset and spontaneous offset of the blockade; the T4/T1 ratios were significantly higher during onset than during offset, although there were large variations of fade in the train-of-four response in each patient during offset. These results suggest that it is difficult to estimate the T1/control T1 ratio by the T4/T1 ratio during offset. Pharmacokinetic analysis revealed that the high dose group had a shorter elimination half-life than did the low dose group. A shorter elimination half-life at a high dose may be to some extent due to hepatic clearance. The pharmacokinetic parameters bore no fixed relationship to the pharmacodynamics in each patient. PMID- 15278581 TI - Interactions between volatile anesthetics and dipalmitoyl phosphatidylcholine liposomes as studied by fluorometry with a thiacarbocyanine dye. AB - The effects of volatile anesthetics on the properties of dipalmitoyl phosphatidylcholine liposome were investigated by fluorescence spectroscopy with a thiacarbocyanine dye (3,3"-dioctadecyl-2,2"-thiacarbocyanine) which is sensitive to the viscosity and the dielectric constant of the environment. Seven volatile anesthetics, halothane, enflurane, isoflurane, methoxyflurane, sevoflurane, diethylether and chloroform were used. All anesthetics decreased the phase transition temperature of the liposome and increased the effective dielectric constant of the water-liposome interface. The increase of the effective dielectric constant was attributed to the release of the hydrated water molecules from the membrane surface. The increment of the effective dielectric constant depended on the thermodynamic activity of anesthetics in the solution, and was not affected seriously by the kind of anesthetics. On the other hand, the degree of the depression of the phase transition temperature depended on the molar concentrations of anesthetics. Considering from the Ferguson's report, which is dealt with the relationship between the physiological effect and the thermodynamic activity, the effect of anesthetics on the effective dielectric constant of the membrane surface is more correlated to the anesthetic action than the effect on the phase transition temperature. PMID- 15278582 TI - Effects of prostaglandin E1 on left ventricular performance in dogs; comparisons with trinitroglycerin and adenosine triphosphate. AB - To examine the cardiovascular response to prostaglandin E1 infusion, we observed hemodynamic changes including left ventricular diameter (an ultrasonic crystal pair) during PGE(1)-induced hypotension in anesthetized open-chest dogs. Left ventricular contractility was assessed primarily by measuring the slope of the left ventricular endsystolic pressure-diameter relation (ESPDR) determined by combining end-systolic points from a vena caval occlusion. The cardiovascular effects of induced hypotension by infusions of trinitroglycerin and adenosine triphosphate were also examined at the equivalent magnitude of hypotension. Approximately 25% reduction of systemic blood pressure was produced by the three agents. PGE(1) significantly increased cardiac output from 1200 +/- 132 to 1439 +/- 162 ml.min(-1) (mean +/- SE, P < 0.05), stroke volume from 9.1 +/- 1.1 to 10.0 +/- 1.0 ml (P < 0.05), and %-diameter shortening from 10.4 +/- 0.8 to 14.4 +/- 0.8% ( P < 0.01), but the slope of ESPDR was unchanged. Similar changes were also observed during adenosine triphosphate-induced hypotension. PGE(1) significantly decreased end-diastolic diameter in a similar manner to trinitroglycerin. Thus PGE(1) appears to have little influence on left ventricular contractility aside from its effects on afterload and preload, indicating that it is a useful agent for producing controlled hypotension during anesthesia. PMID- 15278583 TI - Analysis of oxygen transport and oxygen utilization combined. AB - We propose a model which combines oxygen transport system from blood to tissue with oxygen utilization system at the tissue. The model consists of 3 equations; the relationship between tissue P(O)(2) (Pts(O)(2)) and O(2) utilization (Vrc(O)(2)), diffusion from vessel to tissue, and Fick equation. This model has two advantages. First, it is self-consistent. Varying Vrc(O)(2) varies the oxygen transport. Second, it enables to analyze the effects of various factors of oxygen transport/utilization on other factors. We applied this model to the brain tissue. Following values were assumed. Critical tissue P(O)(2) (Pcrit(O)(2)) 2 mmHg; oxygen utilization above this level 3 ml.min(-1).100 g(-1); diffusion coefficient from blood vessel to tissue (D) 0.2 ml.min(-1).mmHg(-1).100 g(-1); cerebral blood flow (CBF) 50 ml.min(-1).100 g(-1); hemoglobin 15 g.100 ml(-1). Hill equation was used for oxygen dissociation curve with n of 2.7 and P50 of 27.0 mmHg. From these, the following values were obtained; Pv(O)(2), Pts(O)(2) and Vrc(O)(2). The changes were analyzed for the 5 input values, Pa(O)(2), CBF, D, P50 and Hb, changing from zero to their respective normal values. A reduction of a single parameter down to 50% of normal barely affected oxygen utilization. A further reduction resulted in significant oxygen utilization. Under conditions studied, a decrease in P50 reduced oxygen utilization faster than that in any other parameters. PMID- 15278584 TI - The influence of obesity and underweightness on respiratory function of geriatric patients undergoing surgery. AB - We investigated the influence of obesity and underweightness on the respiratory function of 228 patients over 65 ys. old undergoing elective surgery. The parameters we studied were preoperative Pa(O)(2) (Pa(O)(2)-pre), Pa(O)(2) under general anesthesia (Pa(O)(2)-op) and preoperative spirometric values including data from flow-volume curves and closing volumes. Triceps skinfold thickness (TSF), body mass index (BMI), Broca's index (BI) and Onodera's prognostic nutritional index (PNI) were measured or calculated. Respiratory parameters were compared between 3 groups; overweight (BMI >23), normal weight (BMI 20-22), underweight (BMI <19). Single and multiple correlations were analyzed between 3 nutritional parameters (BMI, TSF, PNI) and respiratory values. As a result, Pa(O)(2)-pre and Pa(O)(2)-op in overweight group were lower than those in the other groups. None of other parameters showed significant differences between the 3 groups. In multiple regression analysis, BMI correlated with Pa(O)(2)-pre (r = 0.24), Pa(O)(2)-op (r = -0.43), %VC (r = 0.18), peak flow rate (PFR, r = 0.17) and V(50)/HT (r = 0.18). TSF correlated with Pa(O)(2)-pre (r = -0.22), %MVV (r = 0.28) and RV/TLC (r = 0.28). PNI correlated with PFR (r = 0.23). We concluded that overweightness has greater influence on respiratory function of elderly patients than underweightness and that arterial blood gas analysis is essential in preoperative assessment of obese geriatric patients. PMID- 15278585 TI - Hemodynamic responses during induction of anesthesia with halothane-nitrous oxide in children with or without atropine premedication. AB - Forty three children ranged from 1 yr. to 6 yr. were randomly assigned to non atropinized group (n = 20; A(-)) and atropinized group (0.015 mg.kg(-1) i.m., n = 23; A(+)). Control hemodynamics were measured under 0.5% halothane and 67% nitrous oxide and 33% oxygen for three minutes, and then halothane was increased to 2.5% and maintained for 15 min. In the A(-) group, stroke volume (SV) decreased to 64%, heart rate (HR) increased from 100/min to 111/min, and blood pressure (BP) decreased from 65 mmHg to 62 mmHg. Skin blood flow (SBF) concomitantly measured by a laser doppler flowmeter decreased to 48% and total peripheral resistance (TPR) increased to 128%. In the A(+) group, HR increased from 117/min to 132/min ( P < 0.05, vs. A(-) group), BP decreased from 67 mmHg to 66 mmHg. SV decreased to 71% ( P < 0.05, vs. A(-) group). Changes in SBF and TPR were 68% and 128% respectively. End-expired halothane concentration in the A(+) group increased slower than in the A(-) group but not significantly. The results indicate increased sympathetic tone would work as a compensating mechanism for decreased SV and CO. Atropine premedication attenuated cardiovascular depression by maintaining HR and possibly by delaying induction speed of anesthesia. In conclusion, halotane-nitrous oxide anesthesia decreased SV without a marked decrease in heart rate and blood pressure in children. This decrease in SV and BP was attenuated by atropine premedication. PMID- 15278586 TI - Comparison of visceral pain incidence during cesarean section performed under spinal or epidural anesthesia. AB - The incidence of visceral pain during cesarean section performed under regional anesthesia was studied in 80 unpremedicated patients. They were divided in two similar groups concerning age, weight and height. Group 1 consisted of 40 patients submitted to cesarean section under spinal anesthesia, while in group 2 (40 patients) epidural anesthesia was used. Surgery was totally painless for all patients of group 1 patients, whereas in group 2 intraoperative analgesia was complete for 11, good in 18 and fair in 10 patients. One patient of group 2 required general anesthesia due to excrutiating pain during exteriorization of uterus despite a seemly adequate lebel of cutaneous analgesia of T(6). The authors conclude that spinal anesthesia favorably compares with epidural anesthesia for cesarean section, because the incidence of visceral pain with the former was nill and because both techniques are equally safe for mothers and neonates. PMID- 15278587 TI - Improved PCO2 monitoring during high frequency jet ventilation. AB - During high frequency jet ventilation (HFJV), it has been shown that Pa(CO)(2) can be predicted by capnography when the frequency was temporarily reduced to obtain a steady expiratory CO(2) tension (P et(CO)(2)). The influence of the sampling site of expiratory gas in the airway and the driving pressure of the ventilator on the difference between Pa(CO)(2) and P et(CO)(2) was investigated in ten adult patients who underwent general anesthesia. During HFJV (frequency: 100 bpm, inspiratory duty cycle; 30%), its frequency was temporarily reduced to 10 bpm in twelve different condition; i.e., 3 different driving pressures (Pd; 20, 30, and 40 psi) at 4 different sampling sites (d; 0, 2, 5, and 10 cm in the endotracheal tube). Both P et(CO)(2) and Pa(CO)(2) were measured simultaneously, and their difference was evaluated by using Student's t-test. The difference between Pa(CO)(2) and Pmax(CO)(2) (the maximum P(CO)(2) value in the airway during exhalation) was minimal, when Pd was greater than 30 psi, and d was greater than 5 cm. The present study suggests that better prediction of Pa(CO)(2) can be done by P et(CO)(2) during HFJV, when d and Pd were set as large as possible to obtain stable expiratory P(CO)(2) curve. PMID- 15278588 TI - Changes in coagulation and fibrinolytic activity associated with tracheal intubation. AB - This study was carried out to clarify the effect of tracheal intubation on the coagulation and fibrinolytic system. It was performed on 20 patients (ASA class 1 2) undergoing elective surgery. Before and after tracheal intubation, hemodynamics, ACTH, cortisol, catecholamines, and several coagulation and fibrinolytic factors were measured. Tracheal intubation was accompanied by significant increases in the blood pressure, heart rate, and norepinephrine level. No changes were observed in fibrinopetide A, fibrinopeptide B(Beta15-42), tissue plasminogen activator antigen, plasminogen, fibrinogen, and Alpha(2) plasmin inhibitor. Patients exposed to long intubation time (>20 seconds) were found to have a significantly higher level of fibrinopeptide A than patients with short tracheal intubation time ( isoflurane > enflurane > methoxyflurane > halothane. This partly supports our hypothesis as far as halogenated ethers are concerned. Halothane rarely produced opisthotonus. In the sevoflurane, isoflurane, and methoxyflurane groups, incidence was lower in middle-aged than in young or elderly mice, while incidence increased with age in the enflurane group. PMID- 15278624 TI - Hypoxic ventilatory response in cats lightly anesthetized with ketamine: effects of halothane and sevoflurane in low concentrations. AB - The effect of low concentration sevoflurane and halothane on the ventilatory response to isocapnic hypoxia was studied in sixteen cats. The cats were divided into two groups, sevoflurane group and halothane group, of eight subjects each. As parameters of the hypoxic ventilatory response, A value [the slope of the hyperbolic curve, V(E) = V(0) + A/(Pa(O)(2)-32)] and ratio of V(50) (the minute volume obtained from the hyperbolic equation when Pa(O)(2) = 50 mmHg) to V(0) were studied. These two parameters were examined at three states, sedative state with ketamine as the control, ketamine plus 0.1 MAC inhalation anesthetic, and ketamine plus 0.5 MAC inhalation anesthetic. In the sevoflurane group, the A values were 4789 +/- 1518, 2187 +/- 1214, 1730 +/- 880 (mean +/- SE. ml.min( 1).mmHg) at the control state, 0.1 MAC and 0.5 MAC, respectively. In the halothane group, the A values were 6411 +/- 2368, 2529 +/- 842 and 2372 +/- 545, respectively. The ratios of V(50) to V(0) were 1.32 +/- 0.09, 1.22 +/- 0.09, 1.25 +/- 0.08 in the sevoflurane group, 1.47 +/- 0.18, 1.32 +/- 0.11, 1.54 +/- 0.18 in the halothane group, respectively. The A value at 0.1 MAC of the halothane group was less than the control value significantly. This proved that even low concentration halothane depressed the hypoxic ventilatory responses. The depression of hypoxic ventilatory response could cause postanesthetic hypoventilation. On the other hand, we could not find significant depression on the hypoxic ventilatory response in the sevoflurane group, but we should notice that variances of the hypoxic ventilatory response were large. PMID- 15278625 TI - Prevention of succinylcholine-induced myalgia with lidocaine pretreatment. AB - We studied the effects of 3 mg.kg(-1) lidocaine iv on the succinylcholine (SCh) induced myalgia in 94 unpremedicated ambulant patients undergoing dilatation and curettage of the uterus. The post-SCh myalgia was confirmed through interview by telephone. The data were correlated with the degree of fasciculation and changes in the serum electrolytes and creatine kinase (CK) levels following SCh administration. Pretreatment with lidocaine, 3 mg.kg(-1) iv, significantly reduced the incidence of myalgia from 40.4% of control group to 12.8% lidocaine treated group, but not the CK levels. The severity of myalgia was not related to the intensity of fasciculation assessed by visual observation. The pretreatment with lidocaine had no untoward effect on the circulation, although the peak arterial and peak venous lidocaine levels achieved were 29.6 +/- 23 micro g.ml( 1) and 10.1 +/- 3.3 micro g.ml(-1) respectively. These finding indicated that the pretreatment with lidocaine, 3 mg.kg(-1) iv, was effective in prevention of SCh induced myalgia. PMID- 15278626 TI - A selective thromboxane synthetase inhibitor, OKY-046, fails to improve blood rheology in endotoxin-shocked rabbits. AB - Effects of a selective thromboxane synthetase inhibitor, (E)-3-[4-(1 imidazolylmethyl)phenyl]-2-propenoic acid hydrochloride monohydrate (OKY-046), were studied hemorheologically in endotoxin shocked-rabbits. The animals were intravenously administrated with 0.1 mg of endotoxin 3 times at intervals of 3 days. At 7 days after the last endotoxin injection, endotoxin (0.2 mg.kg(-1)) was intravenously administrated to induce a shock. OKY-046 (30 mg.kg(-1)) was administrated after hypotension was developed by the endotoxin treatment and, then, it was continuously injected at 0.03 mg.kg(-1).min(-1). Blood pressure remained unchanged and hypotensive was maintained during the treatment with OKY 046. Blood was sampled from the femoral artery 15 (before the administration of OKY-046), 45, and 120 minutes after the final administration of endotoxin. Pa(O)(2) increased, and Pa(CO)(2), arterial pH, and base excess (BE) decreased during the endotoxin shock. The decrease of pH and BE was prevented by the administration of OKY-046. In the endotoxin-shocked animals, hematocrit, whole blood viscosity, erythrocyte deformability, plasma fluidity, and the ratio of hematocrit to whole blood viscosity showed no significant differences between the OKY-046 treated animals and non-treated ones. These data show that a selective thromboxane synthetase inhibitor (OKY-046) does not improve the blood rheology during endotoxin shock, although it seems to prevent the acidosis in some extent. PMID- 15278627 TI - Difference in the effect of pancuronium and vecuronium on baroreflex control of heart rate in humans. AB - The effects of pancuronium and vecuronium, each in doses of 0.05 and 0.08 mg.kg( 1), on the baroreflex control of the heart rate were studied in 40 adult patients of either sex (21 men and 19 women) during stable nitrous oxide-oxygen-fentanyl anesthesia. The blood pressure was elevated by intravenous infusion of phenylephrine (4 micro g.kg(-1).min(-1)) for the pressor test, and lowered by a bolus injection of nitroglycerin (0.3-0.5 mg) for the depressor test. Baroreflex sensitivity was judged from the slope of the regression of the systolic blood pressure on the succeeding R-R intervals on the ECG. There was no significant difference between the baseline blood pressure at which both tests were carried out. Nitrous oxide-oxygen-fentanyl anesthesia alone suppressed the baroreflex sensitivity to a level which was at the lower limit of the physiological and non anesthetized state. The 0.08 mg.kg(-1) dose of pancuronium significantly suppressed the reflex sensitivity in both the pressor and depressor tests. However, the 0.05 mg.kg(-1) dose of pancuronium and both doses of vecuronium did not cause any significant change in the test results. PMID- 15278628 TI - Enflurane suppresses phrenic nerve-diaphragm transmission in vivo. AB - We examined the effects of enflurane on the diaphragmatic function in 15 pentobarbital-anesthetized, mechanically ventilated dogs. They were divided into three groups of five animals each, according to the administered concentration of enflurane. The diaphragmatic function was assessed from transdiaphragmatic pressure (Pdi) and integrated diaphragmatic electromyography (Edi) developed at functional residual capacity against an occluded airway during bilateral supramaximal phrenic nerve stimulation at 0.5, 10, 20, 50 and 100 Hz under quasiisometric condition. After a control measurement, enflurane was administered at a constant end-expired concentration (0, 0.5 and 1 MAC) and the measurement was repeated after 1 hour of exposure. The Pdi amplitude generated by single twitch (0.5 Hz) and during 10, 20 and 50 Hz stimulation was unchanged between the groups. No change in Pdi during 100 Hz stimulation was noted during 0 and 0.5 MAC exposure, while it was reduced by 1 MAC of enflurane. When the values of Pdi were expressed as % of maximum Pdi (%Pdi,max) that developed during control measurement and analyzed in terms of %Pdi,max-stimulus frequency relationship, a significant decrease in %Pdi,max was noted for 100 Hz stimulation in 0.5 and 1 MAC groups compared to the control. Similarly, Edi during 100 Hz stimulation obtained in 0.5 and 1 MAC groups was markedly depressed compared to the control. Edi during 50 Hz stimulation was also decreased at 1 MAC. Relative changes in Edi following enflurane administration were greater than the corresponding changes of Pdi. These results demonstrate that enflurane impairs diaphragmatic function through its inhibitory effects on neuromuscular transmission. PMID- 15278629 TI - Circulatory and metabolic changes in the brain during induced hypotension- comparison among trimetaphan, glycerin trinitrate and prostaglandin E1. AB - Induced hypotension was carried out using trimetaphan (TMP), glycerin trinitrate (GTN) and prostaglandin E(1) (PGE(1)) in 45 patients received elective abdominal surgery under anesthesia with enflurane in N(2)O/O(2) in order to evaluate and compare the effects of these three agents on cerebral circulation and metabolism. Upon reduction of mean arterial blood pressure to 60-65 mmHg, cerebral blood flow decreased in the TMP and GTN groups but increased in the PGE(1) group. The changes were quite proportional to those in cardiac index in the three groups. Cerebral oxygen consumption decreased only in the TMP group. Changes in cerebrospinal fluid pressure were not in parallel with those in cerebral blood flow. The former decreased slightly in the TMP group but increased in the GTN and PGE(1) groups. These results offered a great caution for induction of artificial hypotension using these agents. PMID- 15278630 TI - Placental transfer and effects of famotidine on neonates. AB - The effect of famotidine on neonates was studied in 34 obstetric patients who underwent elective cesarean section. In the famotidine group, 20 mg of famotidine was intramuscularly injected at 60 min before induction of anesthesia, and 0.5 mg of atropine was injected at 30 min before induction. In the control group, only atropine was given. Ratio of famotidine concentration in the umbilical venous blood to that in the maternal venous blood was determined as 0.64 +/- 0.13 (mean +/- SD). No significant differences were noted in the Apgar scores, neonatal gastric acidity, and results of liver function tests between the two groups. No side effect, such as the development of gastrointestinal infections, was observed. PMID- 15278631 TI - Effects of abdominal surgery on somatosensory evoked potentials during nitrous oxide-enflurane anesthesia. AB - The effect of abdominal surgery on median nerve somatosensory evoked potentials (SEPs) was studied in 8 enflurane and nitrous oxide anesthesia (GOE) patients. We further compared the effect of epidural anesthesia. The first recording was done immediately prior to induction. Anesthesia was then induced with 5 mg.kg(-1) i.v. of thiopental and maintained with 1.0% enflurane, 66% N(2)O and 33% O(2). Before skin incision for abdominal surgery, the second recording was performed under GOE anesthesia and the third recording during surgery. Then 2% lidocaine was injected into the epidural space through a preinserted catheter, and after 15 min the fourth recording was obtained. The latencies of peaks N1, P2 and N2 and the amplitudes of N1-P2 and P2-N2 were measured. The latencies of N1, P2 and N2 increased and the amplitudes of N1-P2 and P2-N2 deceased significantly after the induction of anesthesia compared with the control values. During abdominal surgery the latencies of N1 and P2 decreased and the amplitudes of N1-P2 and P2 N2 increased. After epidural anesthesia, however, the latencies of N1 and P2 increased and the amplitudes of N1-P2 and P2-N2 decreased significantly and returned almost to the values recorded under preoperative GOE anesthesia. These phenomena indicated that the excitations produced by surgical stimulation in nerve ending might have been transmitted to the central nervous system via spinal nerves and blocked by epidural anesthesia. PMID- 15278632 TI - Effect of inhalation anesthetics on swimming activity of artemia salina. AB - The swimming movement of artemia salina in the artificial sea water was measured by using the video camera system in the absence and presence of anesthetics, i.e. enflurane, halothane, and isoflurane. The movement of artemia looked random at a glance but the obtained distribution curve for the swimming speed was skewed toward the high speed side somewhat resembling a Maxwellian distribution curve seen in the statistics of ideal gases. When anesthetic were added, the distribution curve became sharpened and shifted to the low speed side, which is similar to a behavior of ideal gases when they are cooled down. The mean swimming speed was decreased eventually leading to an irreversible death with increasing the anesthetic dose. The activity was analyzed by using the hydrodynamic equation. The ED(50), which is a dose that causes a 50% reduction in the activity, of all anesthetics used in this study was quite similar to the MAC values for human. It was also suggested that an interaction between anesthetics and artemia was highly cooperative since the larger Hill coefficients were obtained for all three anesthetics used. PMID- 15278633 TI - General anesthesia for a case of abdominal musculature deficiency, prune belly, syndrome. PMID- 15278634 TI - Changes in hemodynamics of anaphylactic reaction induced by transfused blood during operation. PMID- 15278635 TI - Severe hypoxia following spinal anesthesia: possible association with pulmonary embolism. PMID- 15278636 TI - Urgent surgical reperfusion for intraoperative myocardial infarction following coronary artery bypass grafting. PMID- 15278637 TI - Hyperventilation induced coronary artery spasm during anesthesia for neurosurgery. PMID- 15278638 TI - Two cases of asymptomatic epiglottic cyst confirmed by lateral cervical roentgenogram. PMID- 15278639 TI - Impending vagus nerve paralysis accelerated to full manifestation following cervical intrathecal neurolysis--case report. PMID- 15278640 TI - Bronchospasm-induced massive lung collapse during thoracotomy. PMID- 15278641 TI - Epidural spinal cord stimulation for treatment of outpatients with intractable pain-report of three cases. PMID- 15278642 TI - Combined effects of inversed ratio ventilation (IRV) with positive end-expiratory pressure ventilation (PEEP) on cardiorespiratory function in acute respiratory failure. AB - Combined effects of inversed ratio ventilation (IRV) with positive end-expiratory pressure (PEEP) on cardiorespiratory function were examined in 24 patients with acute respiratory failure. Patients were divided into two groups: the IRV group (n = 12) who showed no significant increase in Pa(O)(2) with a 6 cmH(2)O of PEEP and PEEP group (n = 12) who were ventilated mechanically with PEEP only at maximum level of 10 cmH(2)O. In IRV group step-wise prolongation of the I : E ratio from 1 : 1.9 to 2.6 : 1 or 4 : 1 was applied as a Pa(O)(2) was improved and in PEEP group also level of PEEP was increased from 0, 5 to 10 cmH(2)O after one hour period irrespective of Pa(O)(2). Inversed ratio ventilation and PEEP increased significantly Pa(O)(2)/F i(O)(2), the increase being observed 6 hrs (I : E = 2 : 1) and 2 hrs (10 cmH(2)O) after starting IRV or PEEP. Further improvement of oxygenation was not observed in IRV even if I : E ratio was prolonged up to 2.6 : 1 or 4 : 1. These results suggested that combinations of IRV with PEEP were effective and an I : E ratio of 2 : 1 may be optimal, and IRV is advantageous compared to PEEP, but will take more long time to improve oxygenation than PEEP. PMID- 15278643 TI - The effects of calcium antagonists on EEG, evoked potentials and neurologic recovery after complete global brain ischemia for 15 minutes in dogs. AB - The effects of three calcium antagonists on the recovery from neurologic damages after complete global brain ischemia were examined by evaluating the change of a electroencephalogram (EEG), evoked potentials (EP) and a neurologic recovery score (NRS) in dogs. Fifteen minutes global brain ischemia was achieved by occluding the ascending aorta and the caval veins. Nicardipine (NC), flunarizine (FL) and diltiazem (DL) were administered with continuous infusions for three days after the ischemia. The EEG-EP scores (0 : no response - 6 : normal) 3 hr after the ischemia were 1.4 +/- 0.4 (mean +/- SE) in the control, 2.2 +/- 0.3 in the NC, 2.2 +/- 0.4 in the FL and 2.1 +/- 0.2 in the DL. There were no significant differences between the 4 groups. The survival rates on the 7th day after the ischemia were 67% (6/9) in the control, 78% (7/9) in the NC, 56% (5/9) in the FL and 89% (8/9) in the DL. No significant differences were presented between the 4 groups. The NRSs (0 : death - 100 : normal) on the 7th day were 40.3 +/- 7.3 in the control, 59.0 +/- 8.5 in the NC, 63.2 +/- 9.7 in the FL and 55.7 +/- 3.3 in the DL. Each treated group showed a tendency to have a higher NRS than that in the untreated control group. The NRS in all dogs treated by the Ca(++) antagonists on the 7th day was 58.7 +/- 4.1, which was significantly higher than that in the control group ( P < 0.05). We conclude that the continuous administration of calcium antagonists for three days after the global brain ischemia would be beneficial for the neurologic recovery. PMID- 15278644 TI - Prevention of postanesthetic shivering with intravenous administration of aspirin. AB - There have been many reports of postanesthetic shivering (PAS); however, the causes have not been defined clearly, and the reported methods of inhibiting PAS are not always available clinically. In the present study, we assessed the effect of the intravenous administration of aspirin on the prevention of PAS in 62 patients undergoing oral or maxillofacial surgery, anesthetized with enflurane nitrous oxide. Thirty of the patients were randomly selected, and received intravenous aspirin DL-lysine 900 mg (equivalent to aspirin 495 mg) before the end of surgery. No significant differences were noted in the rectal temperatures between the group given aspirin and the control group. Shivering was observed in 17 of the 32 patients of control group. In contrast, shivering was observed in 5 of the 30 patients who received aspirin. This was a statistically significant difference ( P < 0.01). These data indicate that intravenous administration of aspirin significantly inhibited PAS. The finding suggests that PAS is related to prostaglandin synthesis or to the formation of derivatives of arachidonic acid, since aspirin inhibits both the synthesis of prostaglandins and the formation of derivatives of arachidonic acid. PMID- 15278645 TI - Evaluation of alkalinized lidocaine solution in brachial plexus blockade. AB - The effect of alkalinization of lidocaine solution in brachial plexus blockade was evaluated in a double blind study. Commercial 1.5% lidocaine with epinephrine 1 : 200,000 (pH 5.72) was compared with an alkalinized solution of lidocaine (pH 7.12). 10 mg.kg(-1) of each solution was administrated by the axillary perivascular technique in 34 adult patients scheduled for elective surgery. The onset and spread of sensory blockade and the intensity of motor blockade were determined. An alkalinized lidocaine solution produced more complete sensory blockade in all of four main nerves of the upper extremity as compared with the control lidocaine solution. The onset of sensory blockade in the musculocutaneous, radial, ulnar and median nerves was shortened 58%, 40%, 30% and 28%, respectively, by employing the alkalinized lidocaine solution. Also the analgesic onset in the radial and musculocutaneous nerves was significantly faster than the other two nerves ( P < 0.05 and P < 0.01). Furthermore, the intensity of motor blockade was greatly potentiated when alkalinized lidocaine solution was employed. There was no significant increase in plasma concentration of lidocaine in patients who were given alkalinized solution. PMID- 15278646 TI - Contrast radiography and effects of thoracic sympathetic ganglion block- anatomical analysis. AB - Thoracic sympathetic ganglion block is less effective than lumbar sympathetic ganglion block due to differences in the anatomical structure of these regions. Contrast radiographic findings and an analysis of the effects of lumbar sympathetic ganglion block have been reported, but there are few reports concerning thoracic sympathetic ganglion block. The relationship between contrast radiography findings and the effects of thoracic sympathetic ganglion block were studied in 131 block procedures which mainly had hyperhidrosis. PMID- 15278647 TI - Accidental extubations during respiratory management in a children's hospital. AB - An investigation was conducted on the frequency of accidental extubations at Shizuoka Children's Hospital during the past 12 years. The study was performed on 150 randomly selected patients who received respiratory support for more than 24 hr. Fifteen accidental extubations occurred in 9 patients. Most of them (87%) occurred in the neonatal intensive care unit (NICU), and the rate was 1 per 54 days of intubation. The time at which these accidents happened varied, although they were more common during the day-time. The reasons of accidental extubation could not be specified in two-thirds of the cases. It became clear that more immature babies were more likely to suffer accidental extubation, perhaps reflecting the fact that most of the immature babies in the NICU were intubated orally, and that a larger proportion of them required a longer period of respiratory support. Therefore, early weaning from respiratory support is recommended if it is possible. In conclusion, increased surveillance and more secure methods of taping of endotracheal tubes are crucial for preventing life threatening accidental extubations during respiratory support. PMID- 15278648 TI - A close relationship between post-tetanic twitch and train-of-four responses during neuromuscular blockade by vecuronium. AB - The relationship between post-tetanic twitch (PTT) and train-of-four (TOF) responses after intravenous administration of vecuronium were studied using EMG in 20 patients under nitrous oxide and enflurane anesthesia. After the initial dose (0.2 mg.kg(-1)) of vecuronium, the detectable first twitch of PTT (PTT(1)) always preceded that of TOF (TOF(1)) with the mean time interval of 10.7 +/- 2.6 min. The post-tetanic count (PTC) which coincided with the first appearance of TOF(1) was 9.4 +/- 2.6. After the appearance of TOF(1), the magnitude of TOF(1) was almost identical to that of PTC(10) until full recovery from neuromuscular blockade was observed, whether the supplemental doses of vecuronium (0.03-0.04 mg.kg(-1) i.v.) were administered or not. The magnitude of TOF(2) was slightly lower than that of PTC(20). These results suggest that there is a close relationship between these two types of response, and by evaluating not only PTC but also the magnitude of each PTT, the recovery of TOF responses can be predicted and its extent be estimated fairly accurately. PMID- 15278649 TI - Respiration by tracheal insufflation of oxygen (TRIO) at high flow rates in apneic dogs. AB - Tracheal insufflation of oxygen (TRIO) is a technique in which oxygen is introduced into the trachea at a constant flow rate via a catheter advanced to the level of the carina. We studied the effects of flow rates (0.5, 1.0, 1.5 and 2.0 l.kg(-1).min(-1)) on arterial blood gases during TRIO in 6 apneic dogs. The constant flow was administered through the tip of a catheter (I.D. 2.0 mm) advanced to a site of 1 cm above the carina. After 30 min of TRIO, the mean Pa(CO)(2) at the flow rates of 0.5, 1.0, 1.5 and 2.0 l.kg(-1).min(-1) were 88 +/- 20, 76 +/- 20, 64 +/- 23 and 52 +/- 18 mmHg, respectively. CO(2) elimination increased as the flow rates increased from 0.5 to 2.0 l.kg(-1).min(-1). Based on the above study, we examined the effects of TRIO at a flow rate of 3 l.kg( 1).min(-1) in another 5 apneic dogs. TRIO, at a flow rate of 3 l.kg(-1).min(-1), was able to maintain normocarbia over 4 hr. The mean Pa(O)(2) and Pa(CO)(2) at 4.0 hr were 465 +/- 77 and 41 +/- 4 mmHg. Although the mechanism of pulmonary gas exchange during TRIO is unclear, our study is the first to document that normocarbia can be maintained by high-flow TRIO in apneic dots. PMID- 15278650 TI - The use of the laryngeal mask airway in pediatric anesthesia. AB - Laryngeal mask airway (LMA) insertion was tried in 120 pediatric cases, from 2 months to 12 years of age. Initial indications for LMA were the same as for a face mask, except for two additional conditions; anticipation of difficulty with intubation and difficulty in management by a face mask. Size 2 LMA was used in the vast majority of cases. The insertion was successful on the first trial in 108 cases. More than one trial was necessary in 9 cases but only 3 cases required more than 3 trials. Insertion could not be completed in 3 cases. The relationship between the depth of LMA at the front teeth and age could be roughly described by "depth = 10 cm + 0.3 x Age". LMA was found to provide a better and more secure airway than the face mask without direct tracheal intervention. Heart rate did not increase with LMA insertion. It is easy to use and can be used in place of the face mask, but complications such as stomach air inflation due to too vigorous manual ventilation, slight pharyngeal injury, and airway obstruction due to kinking of LMA can occur. These complications can be avoided and must be kept in mind during it's use. LMA itself can be used to obtain a patent airway where an endotracheal airway is difficult to obtain. LMA-aided tracheal intubation can be extremely useful in obtaining endotracheal airways. Non-blind techniques can be used with LMA to increase safety. LMA is a very useful addition to pediatric anesthesia practice. PMID- 15278651 TI - Circulatory stability and plasma lidocaine levels during continuous and intermittent thoracic epidural analgesia. AB - Circulatory stability and plasma levels of lidocaine were investigated in 20 patients who received thoracic epidural analgesia with plain lidocaine during elective abdominal surgery under general anesthesia. In one group, bolus injection of 8 ml of 2% lidocaine was followed by volumetric continuous pump driven infusion (CPI) of 8 ml of 1.5% lidocaine per hour. In the other group, the same initial bolus injection was followed by repetitive intermittent bolus infusions (RII) of 6 ml of 1.5% lidocaine at a 45 min-interval. Circulatory stability was evaluated by a discriminant function. The results showed that epidural analgesia produced smaller circulatory fluctuations with CPI than with RII. Venous plasma lidocaine levels were consistently higher with CPI than with RII. Plasma levels increased stepwise with RII and kept constant with CPI. Differences in plasma levels were significant from 20 min after the initial injection to 135 min. We therefore conclude that epidural analgesia with CPI is superior to that with RII. However, it must be remembered that higher plasma levels may occur with CPI than with RII. PMID- 15278652 TI - Effects of calcium and temperature on tension in isolated canine coronary artery. AB - The effects of calcium and temperature on the tension of isolated canine coronary arterial strips were studied. In 20 mEq. l(-1) K solution, the tension was significantly increased from 0 mg with 0 mEq. l(-1) Ca to 33 +/- 18 mg with 0.2 mEq. l(-1) Ca at 37 degrees C, from -40 +/- 18 mg with 0 mEq. l(-1) Ca to -17 +/- 11 mg with 0.2 mEq. l(-1) Ca at 30 degrees C, from -77 +/- 19 mg with 0 mEq. l( 1) Ca to -52 +/- 17 mEq. l(-1) with 1 mEq. l(-1) Ca at 25 degrees C, from -88 +/- 13 mg with 0 mEq. l(-1) Ca to -41 +/- 18 mg with 2 mEq. l(-1) Ca at 20 degrees C, from -125 +/- 16 mg with 0 mEq. l(-1) Ca to -116 +/- 13 mg with 2 mEq. l(-1) Ca at 15 degrees C. Ca higher than 0.2 mEq. l(-1) produced a dose-dependent increase in tension between 37 degrees C and 15 degrees C. In spite of the presence of 4 mEq. l(-1) Ca, the development of tension was strongly supressed by lowering the temperature below 20 degrees C, and completely inhibited at 10 degrees C. The rate of a decrease in tension caused by cooling was about 5.5 mg. degrees C(-1). This study demonstrated that Ca(2+) produced a dose-dependent increase in tension in high-K solution, which was suppressed as the temperature was lowered. PMID- 15278653 TI - Lung mechanics with tension pneumothorax during mechanical ventilation. PMID- 15278654 TI - Three cases of reflex sympathetic dystrophy in the lower extremity treated with lumbar sympathetic ganglion block. PMID- 15278655 TI - A devised method for the fiberoptic nasotracheal intubation under general anesthesia. PMID- 15278656 TI - Anesthetic management of a patient with Hallermann-Streiff syndrome. PMID- 15278657 TI - Cesarean section of a pregnant woman associated with infection of rubella. PMID- 15278658 TI - Pulmonary edema occurring immediately after surgery. PMID- 15278659 TI - Age-dependent alterations in the response of isolated rat aortas to thiobarbiturates. AB - Responses to thiamylal and thiopental were compared in helical strips of rat thoracic aortas of different ages (5-6, 10-12 and 20-22 weeks old), precontracted partially with phenylephrine or prostaglandin F(2alpha) (PGF(2alpha)). Thiamylal and thiopental, in concentrations of 3 x 10(-5) to 10(-4) M, produced a dose dependent relaxation in aortas at 5-6 weeks of age, no significant change of tension in those at 10-12 weeks of age, and a marked constriction in those at 20 22 weeks of age. These thiobarbiturates, in a high concentration of 10(-3) M, produced a profound relaxation in aortas at any age studied. It is concluded that the responses to thiobarbiturates of thoracic aorta precontracted with phenylephrine or PGF(2alpha) alter with age. PMID- 15278660 TI - Plasma vasopressin response to contrast medium during cardiac catheterization in infants and children. AB - Fifteen infants and children (M = 7, F = 8), aged from 0 to 13 years, who underwent cardiac catheterization and cardioangiography under ketamine-diazepam anesthesia were the subjects of this study. The effect of a contrast medium, isolamate sodium (66.8%) on the plasma somolality and vasopressin concentration was studied. The plasma osmolality was significantly elevated after contrast medium administration (289 +/- 3 vs. 303 +/- 8 mosmol.kg(-1)) as well as plasma vasopressin (from 2.1 +/- 0.9 vs. 4.7 +/- 2.0 micro-unit.ml(-1)). It is concluded that the administration of contrast medium for cardioangiography causes elevation of plasma osmolality, which leads to the elevation of plasma vasopressin concentration. PMID- 15278662 TI - Large dose of flunitrazepam attenuates baroreflex control of heart rate in man. AB - The effects of intravenous tranquilizers such as flunitrazepan, diazepam and droperidol on baroreflex control of heart rate were investigated using both pressor and depressor tests. The dose-dependency was also studied by the administration of the drugs at the usual or twofold dosage. The usual dosage caused no significant change in neuronal controlling mechanism of circulation. However, flunitrazepam at double the usual dosage produced a decrease in baroreflex sensitivity at depressor tests. The results suggest that flunitrazepam when given repeatedly might accelerate hemodynamic derangements induced by an acute decrease in blood pressure. PMID- 15278661 TI - The pre- and postjunctional components of the neuromuscular effect of antibiotics. AB - The relative contributions of the pre- and postsynaptic components of the myoneural blocking effect of different antibiotics were studied using: (a) a radio-active method that measures selectively the Ca(2+)-dependent, stimulation evoked, quantally released, (3)H-acetylcholine ((3)H-ACh) from the mouse in vitro phrenic nerve-hemidiaphragm preparation without cholinesterase inhibition; (b) measurement of the force of contraction of the indirectly or directly stimulated muscle. The antibiotics studied (neomycin, polymyxin B and lincomycin), reduced the release of (3)H-ACh evoked by stimulation (18 trains of 40 shocks at 50 Hz) in a concentration dependent manner. While the inhibitory effect of neomycin was inversely related to [Ca(2+)](o), that of lincomycin was moderately and that of polymyxin B was not affected by increasing [Ca(2+)](o) from 0.75 to 5.0 mM. Similarly, the d-tubocurarine (d-Tc)-induced inhibition of the release of (3)H ACh was independent of [Ca(2+)](o). The K-channel blocking agent, 4-aminopyridine (4-AP), enhanced the release of ACh in a concentration dependent manner and prevented the neuromuscular effect of neomycin. However, the neuromuscular effect of polymyxin B and of lincomycin was not affected by 4-AP. Atropine, enhanced the release of (3)H-ACh. Antibiotics, however, were still able to reduce the release of ACh when the negative muscarinic feedback mechanism of ACh release was eliminated by atropine. Our findings indicate that the antibiotics studied possess both pre- and postsynaptic effects. Presynaptically they reduce the evoked release of ACh; postsynaptically they inhibit muscle contractility. The rank order of presynaptic action is neomycin >polymyxin B >lincomycin. PMID- 15278663 TI - Diaphragmatic fatigue and its recovery are influenced by cardiac output. AB - The effects of cardiac output (Qt) on diaphragmatic blood flow (Qdi) and diaphragmatic fatigue and its recovery were studied in dogs. Qdi was estimated by measurement of the left inferior phrenic arterial blood flow (QdiL). There was a significant positive correlation between % Qt and % QdiL. Transdiaphragmatic pressure (Pdi) was significantly smaller in the lowered Qt group than the control at the end of fatigue, and the speed of recovery from the fatigue was also significantly slower in the lowered Qt group. It is concluded that Qt was considered to play an important role in the pathogenesis of diaphragmatic fatigue and its recovery in the mechanically ventilated dogs. PMID- 15278664 TI - Changes in the plasma histamine concentration after the administration of vecuronium bromide. AB - Clinical symptoms of anaphylactoid reaction to muscle relaxants vary from localized flush to cardiovascular collapse. Vecuronium bromide is reported to have very little histamine releasing property. However, there are some reports of anaphylaxis or anaphylactoid reaction to vecuronium. We studied plasma histamine concentration after the intravenous injection of vecuronium to confirm the histamine release. Twenty patients were randomly allocated to one of two groups, each group comprising of 10 patients: one group was to receive vecuronium 0.1 mg.kg(-1) and the other 0.2 mg.kg(-1) using the priming principle. Blood samples were taken prior to and 1, 3, 5, 8 and 13 min after the administration of vecuronium. The plasma histamine concentration was measured by radioimmunoassay with monoclonal antibody. There were no significant changes in plasma histamine concentration over 13 min after the administration of vecuronium compared with the baseline value. There were also no significant differences between these two groups. We concluded that vecuronium up to 0.2 mg.kg(-1) did not change the plasma histamine concentration in the patients having no previous history of allergy or atopic tendencies. PMID- 15278665 TI - Comparative study on the cardio-respiratory change during prostaglandin E1 induced hypotension in the patients in the supine and prone position. AB - Prostaglandin E(1)-induced hypotension (25% reduction from the preadministration level in mean arterial pressure) was applied to thirteen patients. Eight patients among them were operated in the supine position (group I) and other five in the prone position (group II). The maintenance dose of PGE(1) was considerably lower in group II than in group I (0.067 micro g.kg(-1).min(-1) vs. 0.119 micro g.kg( 1).min(-1)). In group I, there was a significant increase in CI, with a significant decrease in SVRI and PVRI during PGE(1)-induced hypotension. Such a high dose of PGE(1) (0.119 micro g.kg(-1).min(-1)) was considered to have a direct dilating action on the systemic resistance bed as well as on the pulmonary vasculature. It was considered that the suppression of hypoxic pulmonary vasoconstriction could be a mechanism to increase venous admixture during PGE(1) induced hypotension. In group II, there was no significant increase in CI, and no significant decrease in SVRI and PVRI. PGE(1)-induced hypotension can be safely applied to the anesthetized patients, but we should be careful to apply it to the patients in the prone position, because lower dose of PGE(1) can induce severe hypotension, which is not accompanied by the increase in CI as occurres in the patients in the supine position. PMID- 15278666 TI - Fluctuating CPAP (F-CPAP) versus conventional CPAP (C-CPAP) in dogs with blood aspiration. AB - Fluctuating CPAP(F-CPAP) is a combination of spontaneous ventilation and fluctuating PEEP, in which end-expiratory pressure (EEP) is periodically changed within a certain range. In a dog model with localized lung injury induced by the aspiration of non-heparinized blood (2 ml.kg(-1) body weight), we carried out a comparative study of the effects of F-CPAP in which the EEP was cyclically changed from 4 to 12 cmH(2)O with periods of 10 min and those of conventional CPAP with a fixed EEP of 8 cmH(2)O (C-CPAP), on hemodynamics and pulmonary oxygenation. The blood aspiration produced significant increases in the intrapulmonary shunt (Qsp/Qt), the alveolar-arterial difference of partial pressure of oxygen (A-aD o(2)), and the respiratory rate (RR). Although both F CPAP and C-CPAP reduced Qsp/Qt and A-aD o(2) and RR, 7 dogs treated with F-CPAP showed a significantly greater recovery of Qsp/Qt and A-aD o(2) than 7 dogs treated with C-CPAP. There were no significant differences in hemodynamic variables between the two groups. These results suggest that F-CPAP is more useful in the treatment of some kinds of hypoxic respiratory failure due to uneven distribution of lung injury. PMID- 15278667 TI - Intraoperative changes in blood coagulation and the effectiveness of ulinastatin during liver resection. AB - The blood coagulation status of 16 patients undergoing liver resection was monitored by thrombelastograph (TEG). Coagulation test by TEG was performed at three different times: before and one hour after induction of anesthesia and after liver resection. The four variables such as r (reaction time), k (coagulation velocity), ma (maximum amplitude) and me (maximum elasticity) were measured. In 8 patients, Ulinastatin was not administered during the operation and FFP was transfused after the second measurement of TEG (group I). The other 8 patients were administered totally 300,000 units of Ulinastatin after induction until the second measurement of TEG, thereafter FFP was transfused (group II). The TEG showed poor preoperative coagulation state in both groups. In group I, TEG variables showed coagulopathy was exacerbated significantly during liver resection. In group II TEG variables showed no significant changes during operation. Between the two groups there were statistical differences in the TEG variables during the operation. The TEG was useful for monitoring coagulation function during liver resection. It was impossible to improve TEG data by only replacement of FFP. Ulinastatin was useful in normalizing the coagulation function and in preventing the changes in TEG measurements during liver resection. PMID- 15278668 TI - Combined negative inotropic effects of calcium entry blockers and isoflurane on canine isolated heart muscles. AB - The combined negative inotropic effects of isoflurane and calcium entry blockers (verapamil, diltiazem, nifedipine, nicardipine) were studied utilizing isolated heart preparations of ventricular muscles from dogs. All of these calcium entry blockers exerted dose-dependent decreases in maximal velocity of shortening (Vmax), maximal developed isometric force (Fm), and the maximal first derivative of Fm (maximal dF/dt). Dose-dependent decreases of these variables of muscle mechanics were augmented in isoflurane-depressed myocardium. At equimolar concentrations, direct myocardial depression was demonstrated in the following order of severity: nifedipine > diltiazem = verapamil > nicardipine. Percent depressions of Vmax, Fm and maximal dF/dt were significantly greater in muscles when calcium entry blockers were combined with 1MAC isoflurane than in muscles of calcium entry blockers alone. These data suggest that the negative inotropic effects of verapamil, diltiazem, nifedipine, and nicardipine were potentiated by isoflurane. PMID- 15278669 TI - Effects of sodium and temperature on tension in isolated canine coronary artery. AB - The effects of sodium and temperature on tension of isolated canine coronary arterial strips were studied. In 20 mEq. l(-1) K solution, the strength of tension was inversely related to the Na concentration. At 37 degrees C, the tension was significantly increased at 70 mEq. l(-1) Na and below. The tension was gradually suppressed by lowering of the temperature from 37 degrees C to 10 degrees C. At 10 degrees C, tension did not developed significantly at Na concentrations between 127 mEq. l(-1) and 12 mEq. l(-1). It was concluded that the decrease in Na concentrations increased the tension of the canine coronary artery and the lowering of temperature supressed the tension inducted by the decrease in Na concentrations. PMID- 15278670 TI - The effects of sevoflurane on lidocaine-induced convulsions. AB - The influence of sevoflurane on lidocaine-induced convulsions was studied in cats. The convulsive threshold (mean +/- SD) was 41.4 +/- 6.5 mg. l(-1) with lidocaine infusion (6 mg.kg(-1).min(-1)), increasing significantly to 66.6 +/- 10.9 mg. l(-1) when the end-tidal concentration of sevoflurane was 0.8%. However, the threshold (61.6 +/- 8.7 mg. l(-1)) during 1.6% sevoflurane was not significant from that during 0.8% sevoflurane, indicating a celling effect. There was no significant difference in the convulsive threshold between sevoflurane and enflurane. The rise in blood pressure became less marked when higher concentrations of sevoflurane or enflurane were administered and the blood pressure at convulsions decreased significantly in 1.6% sevoflurane, and in 0.8% and 1.6% enflurane. However, there was no significant difference in the lidocaine concentrations measured when the systolic blood pressure became 70 mmHg. Apamin, a selective blocker of calcium-dependent potassium channels, was administered intracerebroventricularly in rats anesthetized with 0.8% sevoflurane to investigate the mechanism of the anticonvulsive effects. Apamin (10 ng) had a tendency to decrease the convulsive threshold (21.6 +/- 2.2 to 19.9 +/- 2.5 mg. l(-1)) but this was not statistically significant. It is suggested that sevoflurane reduces the convulsive effect of lidocaine toxicity but carries some risk due to circulatory depression. PMID- 15278671 TI - Internal jugular vein cannulation: time required under ultrasonographic guidance with a valsalva maneuver. AB - Time required for the cannulation of right internal jugular vein (IJV) under ultrasonographic guidance with a Valsalva maneuver was investigated in surgical patients. The degree of distension of the IJV and the changes in blood pressure (BP) during the maneuver were also examined. The required time for the cannulation was 11 +/- 5 sec (mean +/- SD, n = 21), and there was no failure in the cannulation. The mean BP decreased significantly during the maneuver, while the vessels enlarged approximately twofold, which facilitated the cannulation. The decrease in BP and the enlargement of the vessels during a Valsalva maneuver were confirmed in the left side, which indicated the feasibility of left IJV cannulation under ultrasonographic guidance. PMID- 15278672 TI - The effects of a thromboxane A2 receptor antagonist on neurologic recovery after 15 min complete global brain ischemia in dogs. AB - The effects of a thromboxane A(2) receptor antagonist (anti-TXA(2); ONO-3708) on the neurologic recovery after 15 min complete cerebral ischemia were investigated in dogs. Complete cerebral ischemia was achieved by occlusion of the trunk of the aorta, the superior and the inferior caval vein. Seventeen dogs were divided into 2 groups; 1) control group (no drug, n = 9), 2) Anti-TXA(2) group (anti-TXA(2) 200 mcg.kg(-1) in iv bolus soon after recirculation followed by continuous infusion at a rate of 10 mcg.kg(-1).min(-1) for 3 days, n = 8). EEG, auditory brainstem response (ABR), middle latency response (MLR), and somatosensory evoked potential (SEP) recordings were obtained before ischemia, 120 min after re circulation and on the 7th day after ischemia. The neurologic recovery score (NRS) were determined on the 3rd and the 7th day. Impaired EEG score was significantly higher in the anti-TXA(2) group on 7th day after ischemia ( P < 0.05). Anti-TXA(2) increased the reappearance rates of the ABR-Vth ( P < 0.05) and the SEP-N(3) waves ( P < 0.01) at 120 min after ischemia. The survival rate tended to be higher in the anti-TXA(2) group. However, NRS did not significantly differ in the groups. PMID- 15278673 TI - Peripheral vascular permeability following a thermal injury to the airway. AB - Effects of thermal injury to the airway on the vascular permeability in the region of head and neck, were studied in the canine models. The thermal airway injury was produced by an inhalation of a gas burner's flame through the metallic tracheostomy cannula. The changes in vascular permeability were evaluated by calculating the reflection coefficient, which was obtained by the protein washdown technique into lymph. The reflection coefficient after the flame inhalation did not show any increases, while it increased significantly after a histamine infusion into the carotic artery. We concluded, that the vascular permeability in the unburned area does not increase at least in the first 3 hr after a thermal injury to the airway. PMID- 15278674 TI - False hyperchloremia in bromism. AB - Plasma chloride concentration measured by an ion-specific electrode can be interfered by other ions. The authors experienced a case of phantom limb pain with a marked hyperchloremia (251 mEq. l(-1)) which was measured by the ion specific electrode method. The patient was diagnosed as bromide intoxication due to chronic ingestion of analgesic tablets which contain bromvalerylurea. A toxic level of plasma bromide concentration supported the diagnosis. Elevated plasma chloride and bromide concentrations were normalized in three weeks after discontinuation of the analgesic intake. Laboratory study revealed that fluoride ion did not affect chloride concentration measured by an ion-specific electrode. Bromide and iodide ions, however, interfered with the electrode and produced a large overestimation of chloride concentration. Hyperchloremia should be interpreted carefully when chloride was measured by an ion-specific electrode method. PMID- 15278675 TI - An example of facilities for intraoperative radiotherapy. PMID- 15278676 TI - A safe approach to percutaneous cannulation of the internal jugular vein in children. PMID- 15278677 TI - An attack of bronchial asthma during spinal anesthesia. PMID- 15278678 TI - Intrathoracic migration of an epidural catheter. PMID- 15278679 TI - The first attempt of CO2 removal with an artificial heart-lung machine--revisited a quarter of century later. PMID- 15278680 TI - Unmet needs in information flow between breast cancer patients, their spouses, and physicians. AB - This study focused on the needs and sources of disease information of breast cancer patients and their spouses during early disease in two settings: at the department of oncology (AD) and on a rehabilitation course (RC). The aim was to characterize those patients and spouses who are not content with average information. Eighty percent of AD and 31% of RC patients were content with the available information (p < 0.001) and 75% of AD spouses and 43% of RC spouses reported similarly (p = 0.008). Higher education, younger age, and shorter time (<1 year) since diagnosis indicated a greater need for information among patients, whereas among spouses, only education level was associated with it. More information was needed on prognosis, cancer as a disease, its influence on daily life, and treatment effects. In both groups, the same proportion of patients reported to have felt involved in decision making sufficiently (60%), inadequately (27%), and 19% versus 16% did not want to be actively participating in decision making. The patients were mostly satisfied with participation in decision making, but they expressed unsatisfactory needs on information during early years of breast cancer. Similarly, their spouses were not content with available information. PMID- 15278682 TI - Chemotherapy-induced nausea and vomiting in routine practice: a European perspective. AB - GOALS OF WORK: The aim of this study was to evaluate the occurrence of chemotherapy-induced nausea and vomiting (CINV) and its effect on patients' ability to carry out daily life activities following moderately to highly emetogenic, first-cycle chemotherapy in routine practice in cancer centers of four different European countries. PATIENTS AND METHODS: This was a prospective, cross-sectional, nonrandomized, self-assessment study in 249 patients enrolled from cancer centers in Spain, Austria, Germany, and Switzerland. The study population consisted of 78% women, with a mean age of 54. Breast, lung, and ovarian cancers made up 75% of all cancers in the study. Patients received a mean of 2.0 chemotherapy agents and 2.5 antiemetic drugs. MAIN RESULTS: A total of 450 emetic episodes experienced by 243 patients was recorded over 5 days following chemotherapy, with an average of 1.8 episodes per patient (range: 0-28). A higher percentage of patients (38%) suffered from delayed compared to acute emesis (13%). Between 42% and 52% of all patients suffered from nausea (visual analogue scale > or = 5 mm) on any one day, peaking at day 3. Using the Functional Living Index for Emesis (FLIE) questionnaire, 75% of patients with nausea and 50% with vomiting reported a negative impact of these conditions on performance of daily living. CONCLUSIONS: CINV remains a significant problem in routine practice, particularly in the delayed phase posttreatment. Overall, CINV had a negative impact on patients' daily life. PMID- 15278681 TI - Investigating the utility of serum cytokine measurements in a multi-institutional cancer anorexia/weight loss trial. AB - BACKGROUND: Interleukin-1 beta (IL-1beta), tumor necrosis factor alpha (TNFalpha), and interleukin-6 (IL-6) have been implicated in the cancer anorexia/weight loss syndrome. However, previous smaller studies have yielded conflicting results as to whether circulating, serum concentrations of these cytokines are in fact elevated. As the translational component of a large multi institutional trial, this study assessed the clinical value of serum concentrations of these cytokines in patients with this syndrome. METHODS: Patients with incurable cancer with anorexia and/or weight loss were eligible. All underwent weekly weight measurements and appetite assessment for the first month and then monthly assessments thereafter. Serum was obtained at baseline and at 1 month, and all three cytokines were measured with the Immunolite assay. RESULTS: A total of 118 patients participated. At baseline, 99%, 54%, and 47% of patients' samples had undetectable IL-1beta, TNFalpha, and IL-6, respectively. Similar results were obtained at 1 month. No correlations were observed between serum cytokine concentrations and changes in weight or appetite. Baseline serum IL-6 predicted a diminished survival but only after adjustment for age and cancer site. CONCLUSION: Serum concentrations of IL-1beta, TNFalpha, and IL-6, as measured in this study, provide data of limited clinical value for patients with the cancer anorexia/weight loss syndrome. PMID- 15278684 TI - Human projected area factors for detailed direct and diffuse solar radiation analysis. AB - Projected area factors for individual segments of the standing and sedentary human body were modelled for both direct and diffuse solar radiation using detailed 3D geometry and radiation models. The local projected area factors with respect to direct short-wave radiation are a function of the solar azimuth angle (alpha) between 0 degrees < alpha<360 degrees and the solar altitude (beta) angles between -90 degrees < beta<+90 degrees . In case of diffuse solar radiation from the isotropic sky the local human projected area factors were modelled as a function of the ground albedo (rho) ranging between 0< rho<1. The model was validated against available experimental data and showed good general agreement with projected area factors measured for both the human body as a whole and for local quantities. Scientists can use the equations to predict the inhomogeneous irradiation and absorption of direct and diffuse solar radiation and UV-radiation at surfaces of the human body. In conjunction with detailed multi-node models of human thermoregulation the equations can be used to predict the physiological implications of solar radiation and outdoor weather conditions on humans. PMID- 15278685 TI - A review of indicators of climate change for use in Ireland. AB - Impact indicators are systems/organisms, the vitality of which alters in response to changes in environmental condition. The indicators assessed in this review fall within the impact category of the driver-pressure-state-impact-response (DPSIR) framework. Instrumental records have shown unequivocal changes in climatic conditions over the past 30 years at a global level but impact indicators allow these changes to be monitored at a finer resolution. Our main aim was to review sets of indicators of climate change currently used in various countries and to make recommendations for their use in the Irish environment. We review a preliminary set of climate change impact indicators in five sectors: agriculture; plant and animal distribution patterns; phenology; palaeoecology and human health. Currently, the most effective impact indicators of climate change have proved to be phenological observations of tree developmental stages. The strongest factor limiting the use of indicators is the lack of long-term data sets from which a climatic signal can be extracted. PMID- 15278686 TI - Crop growth and development effects on surface albedo for maize and cowpea fields in Ghana, West Africa. AB - The albedo (alpha) of vegetated land surfaces is a key regulatory factor in atmospheric circulation and plays an important role in mechanistic accounting of many ecological processes. This paper examines the influence of the phenological stages of maize (Zea mays) and cowpea (Vigna unguiculata) fields on observed albedo at a tropical site in Ghana. The crops were studied for the first and second planting dates in the year 2002. Crop management was similar for both seasons and measurements were taken from 10 mx10-m plots within crop fields. Four phenological stages were distinguished: (1) emergence, (2) vegetative, (3) flowering, and (4) maturity. alpha measured from two reference surfaces, short grass and bare soil, were used to study the change over the growing seasons. Surface alpha was measured and simulated at sun angles of 15, 30, 45, 60, and 75 degrees . Leaf area index (LAI) and crop height (CH) were also monitored. Generally, alpha increases from emergence to maturity for both planting dates in the maize field but slightly decreases after flowering in the cowpea field. For maize, the correlation coefficient ( R) between alpha and LAI equals 0.970, and the R between alpha and CH equals 0.969. Similarly, for cowpea these Rs are 0.988 and 0.943, respectively. A modified albedo model adequately predicted the observed alphas with an overall R>0.860. The relative difference in surface alpha with respect to the alpha values measured from the two reference surfaces is discussed. Data presented are expected to be a valuable input in agricultural water management, crop production models, eco-hydrological models and in the study of climate effects of agricultural production, and for the parameterization of land-surface schemes in regional weather and climate models. PMID- 15278688 TI - [Migrainous vertigo]. AB - Migrainous vertigo is one of the commonest cause of episodic vertigo and is increasingly recognized among neurootologists and migraine specialists. The clinical presentation is heterogeneous with spontaneous and positional vertigo lasting seconds to days and inconsistent temporal relationship to headache and other migrainous features. Findings during the acute episode suggest that several pathophysiological mechanisms can be involved. There are no evidence-based guidelines for therapy available. Presently, migrainous vertigo is not included in the classification of the International Headache Society (IHS). We propose that the diagnosis of definite migrainous vertigo can be made in a patient with migraine according to the IHS with at least two attacks of vestibular vertigo accompanied by migrainous symptoms when other causes are excluded. PMID- 15278689 TI - [Pain assessment in the geriatric patient. Part I: pain diagnostics]. AB - The geriatric pain patient is a frail patient at risk of losing the ability to take care of him/herself and, therefore, at risk of losing personal independence. There is a growing probability of comorbidity and of cognitive impairment with increasing age. One of the main objectives of geriatrics is the preservation and promotion of the capacity to perform everyday tasks (activity) and to participate in social life (participation). Comorbidity impacts on disability to a higher degree than pain does. Cognitive restrictions contribute to a decreased validity of the assessment instruments. For these reasons, geriatric assessment has to be multidimensional and has to take into account the patient's psychological and social situation in addition to the pain and physical status. In geriatrics, a set of standardized instruments is used to accomplish this task. It may not be practical to handle the whole set in the practice of the pain specialist; nevertheless, some simple measures for the assessment of functional as well as cognitive status should be incorporated into the diagnostic procedure for the elderly pain patient. Written questionnaires should not be used in cases of sensory or cognitive impairment of the patient. The adequate tool for these patients is a structured pain interview. PMID- 15278690 TI - Treatment strategies in patients with chronic renal disease: ACE inhibitors, angiotensin receptor antagonists, or both? AB - We discuss the evidence supporting the use of angiotensin-converting enzyme inhibitors (ACEI), angiotensin II type 1 receptor blockers (ARB), or the combination of both in children with chronic renal disease. Several large-scale, prospective, randomized studies with clinical end points have been performed in adult patients, but studies in children are relatively scarce. In adult patients with chronic renal diseases, ACEI clearly delay the progression of chronic non diabetic renal diseases, and nephropathy in patients with type 1 diabetes. The benefits of ACEI are most apparent in glomerular diseases with marked proteinuria but extend also to kidney diseases with lower proteinuria. This notion is also supported by several smaller or retrospective trials in children. Therefore, ACEI should be given to children with chronic renal diseases, particularly if high blood pressure and/or proteinuria are present. In adults, large-scale trials have documented that ARB exert similar effects as ACEI but tend to exert fewer undesired side effects. Data on ARB in children with chronic renal disease are still very scarce, but these substances offer an alternative for patients who cannot tolerate ACEI due to unwarranted side effects. Combination therapy with ARB plus ACEI may be more effective than either drug class alone. However, we will need the results of further long-term prospective clinical studies, as well as a better understanding of the role of the AT(2) receptor, before combination therapy can be widely recommended. A trial of ARB plus ACEI is justified in selected patients if blood pressure and/or proteinuria cannot adequately be lowered by ACEI or ARB alone. PMID- 15278691 TI - Experimental impact of the history of packing on the mean pressure in silos. AB - The role played by the history of packing in the behaviour of a granular material confined in a column is studied experimentally. The mean pressure applied by the granular assembly to the base is measured as a function of the height of material poured in the column. We obtain reproducible mean vertical normal stress measurements without using a particular procedure to mobilise the wall friction. We focus on the influence of the filling method on the mean vertical normal stress. Filling the column via the edges induces a higher apparent mass of grain than filling it via the centre. Particular attention is devoted to the measurement of the effect of the force sensor stiffness. We show that the lower the base stiffness, the lower the mean pressure on the base. We also vary the packing fraction. We obtain an increasing relation between the apparent mass and the mean packing fraction, and show that this relation depends on the filling method. PMID- 15278693 TI - Spreading dynamics of 2D dipolar Langmuir monolayer phases. AB - We study the spreading of a liquid 2D dipolar droplet in a Langmuir monolayer. Interfacial tensions (line tensions) and microscopic contact angles depend on the scale on which they are probed and obey a scaling law. Assuming rapid equilibration of the microscopic contact angle and ideal slippage of the 2D solid/liquid and solid/gas boundary, the driving force of spreading is merely expressed by the shape-dependent long-range interaction integrals. We obtain good agreement between experiment and numerical simulations using this theory. PMID- 15278694 TI - Instabilities in thin-film binary mixtures. AB - Binary mixtures of ultra-thin films are subject to both height and concentration fluctuations, either of which, under the appropriate conditions, can become unstable. We investigate a simple general thermodynamic model which shows that, due to the constraint imposed upon coupling between these fluctuations, a thin film binary mixture will be less stable than if either height or concentration fluctuations are considered in isolation. We investigate the height dependence of the stability condition, discuss equilibrium conditions when higher-order contributions to the free energy are negligible, and predict the scaling behaviour. PMID- 15278692 TI - DNA renaturation at the water-phenol interface. AB - We study the renaturation of complementary single-stranded DNAs in a water-phenol two-phase system, with or without shaking. In very dilute solutions, each single stranded DNA is strongly adsorbed at the interface at high salt concentrations. The adsorption of the single-stranded DNA is specific to phenol and relies on stacking and hydrogen bonding. We establish the interfacial nature of DNA renaturation at high salt, either with vigorous shaking (in which case the reaction is known as the Phenol Emulsion Reassociation Technique or PERT) or without. In the absence of shaking, the renaturation involves a surface diffusion of the single-stranded DNA chains. A comparison of PERT with other known renaturation reactions shows that PERT is the most efficient one and reveals similarities between PERT and the renaturation performed by single-stranded nucleic acid binding proteins. The most efficient renaturation reactions (either with PERT or in the presence of condensing agents) occur in heterogeneous systems, in contrast with standard thermal renaturation, which takes place in the bulk of a homogeneous phase. This work highlights the importance of aromaticity in molecular biology. Our results lead to a better understanding of the partitioning of nucleic acids, and should help to design improved extraction procedures for damaged nucleic acids. We present arguments in favor of interfacial scenarios involving phenol in prebiotic chemistry. PMID- 15278695 TI - The hydrodynamic hypothesis versus the bulk flow hypothesis. PMID- 15278697 TI - AAA--abdominal aortic altruism. PMID- 15278698 TI - Transient edema of the spinal cord as a result of spontaneous acute epidural hematoma in the thoracic spine. AB - We present an unusual case of spontaneous epidural hemorrhage in the thoracic spine resulting in rapid onset of transient and extensive edema in the spinal cord. The patient presented with acute onset of midscapular back pain, bilateral lower extremity weakness, and bladder dysfunction. Repeat MRI 20 days after decompression of the hematoma showed residual hematoma and complete resolution of the spinal cord edema. The implications and differential diagnosis of spinal cord edema in this clinical setting are discussed. PMID- 15278699 TI - Controversies in cervical spine imaging in trauma patients. AB - No area of emergency radiology has generated as much discussion in recent years as the subject of cervical spine imaging for trauma patients. This review will be in three parts. The first will examine the indications for cervical imaging and will focus on those factors that make patients at high risk or low risk for cervical injury. The second part will discuss the merits of radiography and computed tomography as the main screening diagnostic examination. In addition to the roles of each modality in the evaluation process, such factors as efficacy of diagnosis, time (duration) of study, and cost will be discussed. Finally, the third part will explore the methods currently employed to "clear" the cervical spine in comatose patients. PMID- 15278700 TI - Early intrathoracic migration of Kirschner wires used for percutaneous osteosynthesis of a two-part humeral neck fracture: a case report. AB - We present an unusual case of early migration of three Kirschner wires used for percutaneous osteosynthesis of a two-part humeral neck fracture, causing hemothorax. An 85-year-old woman was admitted to the emergency room after casual accident. She was found to have suffered a two-part fracture of the surgical neck of the right humerus. The humeral fracture was treated by closed reduction and percutaneous osteosynthesis with three threaded Kirschner wires, which were bent subcutaneously. Ten days after the accident the patient presented with dyspnea and laterocervical pain. Plain X-rays and complementary CT demonstrated intrathoracic migration of the three Kirschner wires with hemothorax. Two of the wires were seen under the right clavicle and adjacent to the C7 vertebra. The third wire reached the lateral chest wall. Immediate surgery was performed, with withdrawal of the wires and placement of a drainage tube. The patient had an uneventful recovery after surgery. The humeral fracture resulted in a nonunion, which was well tolerated by the patient and was left untreated. The use of Kirschner wires for osteosynthesis of proximal humeral fractures may cause significant thoracic morbidity, even if various prophylactic measures, including the use of threaded wires, subcutaneous bending, and close radiographic follow up, are adopted. The use of Kirschner wires should anyway be restricted to carefully selected cases, in order to avoid major complications. PMID- 15278701 TI - Frequently missed fractures in children (value of comparative views). AB - Fractures in infants and children are different to those seen in adults. Many are very subtle and difficult to detect with certainty. Furthermore, variations in bone contour and epiphyseal plate configuration are endless and at first may suggest pathology. Comparative views are therefore invaluable and are emphasized throughout this communication, which deals with: (1) plastic bending fractures, (2) hairline fractures, (3) impaction fractures, (4) subtle epiphyseal metaphyseal Salter-Harris fractures, and (5) subtle angle buckle fractures. Helpful points to assist one in detecting these fractures are presented. PMID- 15278702 TI - Acute myocardial infarction demonstrated by multidetector CT scanning. AB - A 55-year-old man was brought to the emergency room complaining of left-sided chest pain. His electrocardiogram was indeterminate, and a multidetector computed tomogram (MDCT) was performed to exclude aortic dissection. The patient's aorta was normal, but an area of hypoperfusion was evident in the lateral ventricular myocardial wall. The ability to diagnose myocardial ischemia and infarcts on nongated MDCT is of particular clinical interest. As more imaging technology is devoted to imaging the heart, the greater expectations of radiologists' ability to diagnose cardiac disease in the emergency room will become. PMID- 15278703 TI - Intrathecal injection of epidural blood patch: a case report and review of the literature. AB - Epidural blood patch (EBP) is a commonly performed procedure for the treatment of persistent severe post- dural-puncture headache (PDPH). It has a high success rate with a low incidence of complications. We report the case of a 27-year-old woman who developed progressive back pain and radicular symptoms after an EBP was performed for PDPH. An emergency MRI showed a subarachnoid hematoma. Gradual recovery occurred without the need for intervention. To our knowledge, this is the only case demonstrating the MRI findings of a rare complication of a common procedure. Radiologists may benefit from familiarity with epidural blood patching, including the technique, risks, benefits, and potential complications PMID- 15278704 TI - Acute lymphoblastic leukemia of the skin and subcutaneous tissues; the first manifestation of disease in a 6-month-old infant: a case report with literature review. AB - Leukemic infiltrate involving the skin and subcutaneous tissue was the first manifestation of disease in a 6-month-old female infant. Knowledge of age-related distribution patterns of the red (cellular) and yellow (fatty) marrow is crucial for the interpretation of magnetic resonance imaging (MRI) studies. Diffusely decreased signal intensity throughout the bone marrow on the T1-weighted images specifically involving the epiphyseal ossification centers in infants 6 months after their appearance should be suggestive of a marrow infiltrative/replacement process. Correlation with the peripheral blood smear and bone marrow aspirate are necessary for the diagnosis of leukemia. PMID- 15278705 TI - Primary epiploic appendagitis: CT diagnosis. AB - The purpose of this study was to analyze the CT signs of primary epiploic appendagitis. A retrospective search of the CT database over 12 months for this diagnosis revealed 11 cases. The clinical findings were recorded. Softcopy CT images were reviewed by two experienced abdominal radiologists (KS, DM) for location of lesion, size, shape, presence of central hyperdense focus, degree of bowel wall thickening, mass effect, and ancillary signs. Abdominal pain was the primary symptom in all patients. Preliminary diagnoses were appendicitis (n=2), diverticulitis (n=5), pancreatitis (n=1), ovarian lesion (n=1), or unknown (n=2). Abdominal examination and white blood cell count were uninformative. CT examination revealed a solitary (n=11), ovoid (n=9) fatty lesion with some soft tissue stranding adjacent to the left colon (n=6), transverse colon (n=3), or right colon (n=2). Central hyperdensity (n=5), mild bowel wall thickening (n=2), and parietal peritoneal thickening (n=4) were also seen. In 4 patients the lesions were not visible on follow-up CT examination performed 23-184 days later. Primary epiploic appendagitis can clinically mimic other, more serious inflammatory conditions. Knowledge of its findings on CT would help the radiologist make the diagnosis and allow a more conservative approach to patient care. PMID- 15278706 TI - The field of emergency radiology--recognition at last? PMID- 15278707 TI - Controversies in emergency radiology. CT versus ultrasound in the evaluation of blunt abdominal trauma. AB - There has been controversy regarding ultrasonography (US) versus CT in blunt abdominal trauma (BAT). Each modality has its strengths and weaknesses. US is fast and allows resuscitative efforts to proceed while the patient is being scanned. However, the sensitivity of US is inferior to that of CT, and there is user variability. CT is better at determining the extent, type, and grade of injury, resulting in a more tailored therapeutic plan and safe conservative management of many patients. However, CT involves ionizing radiation, cannot be performed portably, and requires only visual monitoring while scanning. Given each modality's strengths and weaknesses we conclude that CT is the preferred examination when the BAT patient is stable or moderately stable, enough to be taken to CT. If a BAT patient is unstable, US is beneficial in screening for certain injuries or large hemoperitoneum prior to an exploratory laparotomy. PMID- 15278708 TI - The use of sonography versus computed tomography in the triage of blunt abdominal trauma: the European perspective. AB - The management of the trauma-emergency patient has become an important political and economic issue and one of the major challenges of the industrialized countries. In Europe ultrasonography is always part of the basic work-up, following physical examination, whereas computed tomography (CT) remains a second line investigation. Injury prevalence, radiation dose exposure, practicability, and costs are relevant considerations in our emergency departments, where we have a growing number of patients seeking medical attention. The radiologist's task is to decide which imaging modality is most appropriate after the clinical context has been taken into consideration. The clinical value of CT is unquestioned; what is questionable is only its systematic use. With the growing demand for trauma care, screening ultrasonography can lower the number of inappropriate CT examinations. PMID- 15278709 TI - Emergency percutaneous biliary drainage in patients post liver transplantation. AB - The aim of this study was to evaluate the efficacy of emergent biliary drainage in post-liver-transplant patients. We reviewed the results of 17 biliary drainages in 15 post-liver-transplant patients. After biliary drainage there was a 72.8% decrease in direct bilirubin and 52.5% decrease in gamma-glutamyl transpeptidase (GGTP). Anastomotic stricture was the most common cause of biliary obstruction. Emergent biliary decompression in post-liver-transplant patients is a life-saving procedure with a high success rate and immediate positive results. PMID- 15278710 TI - Cholecystocolonic fistula: serial CT imaging features. AB - We report the CT imaging findings of an unusual case of cholecystocolonic fistula, which had presented in the emergency department with melena. It is rare for the fistulous communication to occur between gallbladder and the colon. We describe the serial imaging findings, which were diagnostic of this condition. PMID- 15278711 TI - Penetrating injuries of the neck and the increasing role of CTA. AB - Vascular injuries of the neck are most frequently the result of penetrating trauma. Diagnostic evaluation of hemodynamically stable patients who have suffered penetrating neck wounds is challenging and remains controversial. In order to reduce morbidity and mortality, prompt diagnosis and subsequent treatment of these injuries is critical. Traditionally, these patients undergo direct contrast angiography. However, this technique has limitations including its invasive nature and potential complications. The use of routine screening angiography has also been questioned because of the low rate of positive examinations. More recently, helical and multislice CT angiography (CTA) has emerged as a fast, minimally invasive accurate study to evaluate penetrating neck injuries. CTA is not operator-dependent and the results can be reproduced easily by using established technical parameters. It is readily available in most centers and allows the simultaneous evaluation of the extravascular soft tissues and bones. PMID- 15278712 TI - Patient encounter time intervals in the evaluation of emergency department patients requiring abdominopelvic CT: oral contrast versus no contrast. AB - The aim of the study was to assess various time intervals during patient encounters involving unenhanced (NECT) versus oral-contrast-enhanced (CECT) abdominopelvic (A/P) CT performed in the emergency department (ED) on adult patients presenting with acute abdominal pain. Computerized patient order entry and administrative data as well as scans themselves were retrospectively evaluated at a high-volume (107,000 visits per annum) regional medical center urban ED for a period of 30 consecutive days. All adult patients who had CT of abdomen and pelvis for abdominal pain during the 30 days of the study period were included. Data collected included demographic information, time of registration, time of first encounter in the ED, time of CT order, clinical indication for scan, time of scan, time of disposition (i.e., discharge or admit), and final disposition. Patients were excluded if they were less than 16 years old, pregnant, or met criteria for major trauma and evaluation in the trauma suite. Patients were also excluded from analysis if they received more than one scan on the same day (3 patients). Of 183 patients, 102 underwent NECT and 81 CECT. Some of the patients who underwent NECT had urinary colic. Among patients who did not have urinary colic there is a statistically significant difference in the median time intervals between: (1) patient arrival in the ED and evaluation by a physician (NECT 57 min, CECT 84 min, P<0.001); (2) patient exam by the physician and the time the A/P CT was ordered (NECT 35 min, CECT 63 min, P<0.01); (3) receipt of the CT order and the time of the scan (NECT 104 min, CECT 172 min, P<0.001); and (4) time of arrival in ED and disposition (NECT 358 min, CECT 599 min, P<0.001). There are significant time interval differences between CECT and NECT during patient encounters involving adults presenting with abdominal pain to the ED. The differences are greater than the amount of time allotted for opacification of small bowel (90 min). Baseline data such as these may prove useful in assessing the efficacy of scan techniques and improving resource utilization. PMID- 15278713 TI - Evaluation of the use of the X-ray department with regard to plain chest radiography on acute general medical admissions in the context of recently introduced UK guidelines. AB - All requests for radiographs in the United Kingdom should comply with Royal College of Radiologists (RCR) guidelines, which aim to encourage more appropriate use of diagnostic radiology and so reduce the number of clinically unhelpful radiographic examinations. The aim of this study was to evaluate the use of the X ray department during the normal working day and out of hours with regard to plain chest radiography on acute general medical admissions. A prospective study was performed using a questionnaire filled in by the requesting clinician, radiographer, and ward staff. Of 265 forms were used in a 1-month period, 221 were completed satisfactorily, and the data from these were used in the study. The median time taken for the chest X-ray to be performed was 80 min; once performed it took a median of 9 min to be returned to the ward. The requesting clinician took a median of 25 min to review the chest X-ray. However, the most striking statistic is that only 22% of the chest X-ray taken were reviewed by the junior doctors; the other 78% were looked at for the first time on the "post take" ward rounds. The questionnaire form allowed requests to be graded as urgent, "soon," and nonurgent. There was very little difference in how long it took X-rays in these categories to be performed. This study shows that requests for the chest X-rays on acute medical admissions are more or less routine. Even though many are requested and performed as urgent examinations, 78% of all chest X-rays on these patients are not reviewed until the following morning. While many of these examinations may be deemed necessary, it is unlikely that all will comply with the RCR guidelines. PMID- 15278715 TI - Urine protein as a rapid screen for renal function in the ED: can it replace serum creatinine in selected patients? AB - Many radiology departments are unwilling to perform studies that require contrast administration to adult emergency department patients over the age of 35 without having a documented serum creatinine concentration of less than 2.0 mg/dl within a week of the study. Significant diagnostic delays may ensue waiting for this serum test. The present study was performed to determine whether a negative urine protein test, obtained by dip testing, will serve as a marker for a serum creatinine concentration below 2.0 mg/dl in emergency department patients who give no history of a disease which potentially could cause renal insufficiency. Emergency patients aged 35 years or more presenting to a university hospital who did not volunteer a history of hypertension, diabetes mellitus, multiple myeloma, or systemic lupus erythematosus in triage were enrolled. Only patients with a negative urine protein test whose serum creatinine was tested for other reasons were included. Of the 310 patients who had no protein in their urine and no history of disease which potentially could cause renal insufficiency, none had a serum creatinine concentration greater than 2.0 mg/dl (mean=0.82 mg/dl, SD 0.28). Ages ranged from 35 to 96 years (mean=59.7 years, SD=17.5). All patients would have qualified for a contrast load for contrast computed tomography studies or for intravenous pyelogram. In patients who do not have a known history of hypertension, multiple myeloma, systemic lupus erythematosus, diabetes mellitus, or specific renal disease, urine dip testing for protein in the emergency department may be a rapid and safe screen for imaging studies requiring contrast without having to await serum creatinine testing. PMID- 15278716 TI - Validity of plain films in intussusception. AB - The aim of this study was to re-evaluate the specificity of plain film findings in intussusception. The plain film findings in 80 cases of proven intussusception were reviewed. Findings documented were: (1) presence or absence of small bowel obstruction, (2) paucity of right lower quadrant gas, (3) presence of an intracolonic mass, (4) presence of a rim or target sign, and (5) presence of the classic triad of intestinal obstruction, intracolonic mass, and paucity of right lower quadrant gas. Intestinal obstruction was present in 54% of patients. In 19 patients (24%) the abdominal films were completely normal. Paucity of right lower quadrant gas was seen in 10% of patients, while specific findings of a mass or a target (rim) sign were seen in 29% of patients. The classic triad of an intracolonic mass, obstruction, and paucity of gas in the right lower quadrant occurred in only 1 patient (1%). Plain films of the abdomen were diagnostic of intussusception in only 29% of cases. A completely normal gas pattern was seen in one-quarter of our patients. This being the case, most patients with suspected intussusception will require further imaging, either by ultrasound or contrast enema. In our institution we favor the ultrasound study. PMID- 15278717 TI - Alternative diagnoses to stone disease on unenhanced CT to investigate acute flank pain. AB - Acute flank pain is a common problem in emergency medicine. The most frequent cause is urolithiasis, but many other entities can cause the same clinical presentation. In many institutions unenhanced computed tomography (CT) of the abdomen is used in this setting. One of the major advantages of unenhanced CT is its ability to detect other pathologies causing flank pain. In this pictorial review, we present the CT findings of pathologies other than stone disease in patients with acute flank pain. PMID- 15278718 TI - Traumatic diaphragmatic rupture: can oral contrast increase CT detectability? AB - Traumatic diaphragmatic rupture is a frequently missed diagnosis. We present a patient with traumatic diaphragmatic hernia. Diagnosis was suggested by a emergent computed tomography (CT) examination without oral contrast. Diaphragmatic rupture and herniation of stomach were confirmed by repeating CT examination after the administration of oral contrast and using multiplanar reconstruction. PMID- 15278719 TI - Large wooden foreign body in the hand: recognition of occult fragments with ultrasound. AB - A 37-year-old man with pain and swelling of the thenar was referred to the emergency department. On ultrasound a 3 x 1 x 0.2 cm large wooden foreign body was depicted in the thenar region. In addition, ultrasonography (US) was able to show multiple smaller fragments adjacent to the larger foreign body. To give a better overview of the position relative to tendons and muscles, CT with soft tissue window settings was performed. CT gave a good anatomic overview but was not able to show the smaller fragments. A total of six additional fragments were depicted at US. Performing US is mandatory in patients with penetrating injuries by foreign bodies because it is very sensitive. Using US in an emergency setting can avoid retained fragments and depict other soft tissue complications. PMID- 15278720 TI - An 810 nm diode laser in the treatment of small (< or = 1.0 mm) leg veins: a preliminary assessment. AB - A consistently effective treatment for small leg veins (< or = 1.0 mm) is still being sought. The efficacy of an 810 nm diode laser in vein removal was assessed in a preliminary study. Fifteen females, skin types I to III, vein diameters 0.5 1 mm, aged from 25 to 42 years, participated in the study. An 810 nm diode laser (90 W, 20 ms/pulse, 10 Hz rep rate, 4.0 mm hand piece) was applied along the target veins. Biopsies were taken from two patients before and after the first treatment session. No compression was applied post-treatment. Four weeks later, a second treatment was given. Results were assessed subjectively from the patients' satisfaction index (SI) and objectively from clinical photography done by an independent clinician, who also judged the venous morphology before and 4 weeks after the second session. All patients completed the trial. Pain was moderate to severe at the time of treatment and erythema which was mild, which was seen in all 15 patients; oedema occurred in 12 patients and blistering in only one. No scarring was noticed. The overall satisfaction indices at the 4- and 8-week assessments were 20.7% and 55.1%, respectively. No patient got worse. The objective evaluations at the 4- and 8-week assessments showed increasing improvement in all aspects examined. Pain at the time of treatment was a problem for all patients, so epidermal cooling should be added. Despite this, the 810 laser diode was an interesting and promising device for treatment of small leg veins, warranting further study in larger patient cohorts with a longer-term follow up. PMID- 15278721 TI - Hair removal with intense pulsed light. AB - The use of light and laser for hair removal has evolved during the past few years. Laser systems such as the ruby laser (694 nm), alexandrite laser (755 nm), diode laser (810 nm) and neodymium:yttrium-aluminium-garnet (Nd:YAG) laser (1,064 nm) are commonly used in hair removal. However, permanent hair removal has been difficult to achieve using lasers owing to the long growth/rest cycle of normal human hair follicles. There is still an increasing demand for safer and more efficient hair removal techniques. The latest and most effective choice in the treatment of hair removal is non-coherent intense pulsed light (IPL), which is both efficient and safe for hair removal. A group of 210 patients with skin type III-V were treated for superfluous hair in different areas of the body (face, extremities, axillae, bikini line and back) for three to five sessions at 6-week intervals using IPL. There was a significant hair reduction of about 80% with no side effects and minimal complications. Follow-up was done 6 months after the last session. In conclusion, IPL is very effective and safe for hair removal. PMID- 15278722 TI - Impact of holmium:YAG and neodymium:YAG lasers on the efficacy of DNA delivery in transitional cell carcinoma. AB - New approaches in the treatment of transitional cell carcinoma (TCC) are using gene therapy to influence the disease at the genetic level. Technical advances in genomics, the availability of tissue-specific gene promoters and other developments have made this approach more realistic. Transporting the gene into the target cell is still the major problem. Several transfection techniques have been introduced. Transfection of naked DNA is one of the simplest to perform but transfection rates have been very poor. We investigated the influence of laser energy on transfection efficacy in urothelial cancer cells in vitro with two types of medical lasers. A suspension of human transitional cancer cells (UM-UC3; 3.5 million cells/ml) was mixed with 200 microg of plasmid DNA (pEGFP-N1). Two types of laser energy, neodymium:YAG (Nd:YAG) and holmium:YAG (Ho:YAG), were applied to the cell suspension in different energy settings. Twenty four hours after treatment, transfection rates were measured with FACS analysis. Energy setting parameters that determine the efficacy of laser were investigated. The significance of different transfection rates was estimated with the student's t test. We demonstrated that the Nd:YAG laser was not suitable for achieving significant transfection of the reporter gene to the cells. In contrast, the Ho:YAG laser produced satisfactory transfection rates. There was an increase in transfection with increasing frequency of laser pulses, from 16% with 2 Hz up to 40% with 10 Hz (p < 0.0005). Pulse frequency was therefore stabilised at 10 Hz. Pulse energy (mJ) showed the same dependency: a transfection rate of 18.3% was achieved with 1,000 mJ and 53.8% with 2,000 mJ (p > 0.0005). Additionally, we investigated the impact of total pulse number (imp) with different pulse energies. At 1,000 mJ, a transfection rate of 18.3% was estimated with 200 imp and 48.56% with 750 imp, (p < 0.0005). At 2,000 mJ, a transfection rate of 53.8% was achieved with 200 imp and 58.26% with 500 imp. The optimal laser setting observed in this experiment was 10 Hz, 2,000 mJ and 500 imp. This study indicates that the efficacy of naked DNA delivery into TCC in vitro is improvable by application of Ho:YAG laser energy. The Nd:YAG laser did not increase transfection rates in our model. Our results with the Ho:YAG laser are encouraging for further studies to optimise DNA delivery. As TCC tissue is relatively easy to access, this method could become an effective and minimally invasive procedure in urothelial cancer treatment. PMID- 15278723 TI - The 980-nm diode laser for brain surgery: histopathology and recovery period. AB - The 980-nm diode laser has been under investigation for neurosurgery because of a local peak in the absorption spectra of water around this wavelength. This work was carried out to examine the extent of thermal changes and the recovery process of laser-induced brain lesions. In order to study the quality of the lesions, a conventional monopolar electrocoagulation technique was applied comparatively. An in vivo stereotaxic neurosurgical procedure was performed on Wistar rats. Bilateral brain lesions of the same size (2-3 mm diameter) were created with a diode laser and via electrocoagulation. Subjects were sacrificed 0, 2 and 7 days after surgery in order to observe the healing process of the necrotic tissue. The surgical after-effects of both types of lesions were identified through immunohistochemical staining with CD68 macrophage marker and haematoxylin eosin (H&E). CD68 was found to be more efficient than H&E in determining the thermally altered areas. Histological examinations showed that the 980-nm diode laser system has a remarkable ablating ability with minimal thermal damage of nearby tissue. PMID- 15278724 TI - External temperature during KTP-Nd:YAG laser irradiation in root canals: an in vitro study. AB - To avoid the damage of periodontal tissues during laser irradiation of the root canal, the conditions of lasering must be carefully controlled. The aim of this study was to determine the safety parameters of the irradiation conditions during the use of a KTP-Nd:YAG laser in root canals. Root canals of 60 freshly extracted teeth were prepared (step-back technique) and filled with a photosensitiser (Acid Red 52). Different irradiation conditions [output power (P), pulse width (PW), pulse repetition rate (PRR, Hz)] were used. The laser beam was delivered by means of the KTP-Nd:YAG fibre tip with a beam spot-size diameter of 200 microm. The temperature increases were measured on the external apical third of the root surface using a thermocouple. The results showed that the temperature rise was always below 7 degrees C at the following laser settings: (1) single irradiation, P < or = 4 W, PW < or = 2.55 ms and PRR < or = 20 Hz; (2) repetitive irradiation series, five pulses of 1 s each, four resting times of 1 s each, P < or = 2.25 W, PW < or = 2.55 ms and PRR < or = 20 Hz. The use of resting times was necessary so as to avoid thermal cumulative effects. In these in vitro irradiation conditions, the use of the KTP-Nd:YAG laser in endodontics may be considered harmless for periodontal tissues. PMID- 15278725 TI - Nitrogen laser irradiation (337 nm) causes temporary inactivation of clinical isolates of Mycobacterium tuberculosis. AB - We have investigated the effect of nitrogen laser irradiation (337 nm) on viability of clinical isolates of Mycobacterium tuberculosis. Bacteria were exposed to a nitrogen laser (average power 2.0 mW) in vitro at power density of 70 +/- 0.7 W/m2 for 0-30 min, and the cell viability was determined by luciferase reporter phage (LRP) assay. Immediately after laser exposure, all the clinical isolates investigated showed a dose-dependent decrease in cell viability. However, when the laser-exposed isolates were incubated in broth medium for 3 days, most of these showed significant recovery from laser-induced damage. Addition of 5.0 microg/ml acriflavine (a DNA repair inhibitor) in the incubation medium had no significant effect on recovery. This suggests that DNA damage may not be involved in the cell inactivation. Electron paramagnetic resonance (EPR) studies using 5-doxyl strearic acid (5-DS) as a probe suggest alterations in lipid regions of the cell wall. Implications of these results for understanding therapeutic effect of nitrogen laser on drug-resistant tuberculosis are discussed. PMID- 15278726 TI - Effects of an Er:YAG laser on mitochondrial activity of human osteosarcoma derived osteoblasts in vitro. AB - The aim of the present study was to investigate the collateral damage of an Er:YAG laser on bone cells in vitro using a special application tip designed for treatment of periimplantitis. Before laser irradiation, SaOs-2 osteoblasts (2 x 10(4) cells) were inoculated into 96-well tissue culture plates and incubated for 48 h under standardised conditions. A total of 120 cell cultures were irradiated with an Er:YAG laser using a cone-shaped quartz glass fibre tip at energy settings of 40, 60, 80 and 100 mJ at 10 Hz (energy densities of 5.08, 7.62, 10.16 and 12.7 J cm(-2)) for 10 s. Each energy setting was used at a distance of 1, 2 and 3 mm between the application tip and the bottom of the culture plate. Following irradiation, mitochondrial activity of the cells was measured using a luminescent cell viability assay. After laser irradiation, mitochondrial activity of SaOs-2 osteoblasts was significantly reduced when compared with nonirradiated cells (P < 0.001), dependent on the energy setting used and the distance between the application tip and the bottom of the culture plate. Mitochondrial activity increased significantly with decreasing energy settings and increasing distances (P < 0.001). Within the limits of the present study, it was concluded that an Er:YAG laser, used with a cone-shaped glass fibre tip designed for treatment of periimplantitis, has detrimental effects on mitochondrial activity of SaOs-2 osteoblasts in vitro at energy settings of 40, 60, 80 and 100 mJ (10 Hz). PMID- 15278727 TI - Risk of Borrelia burgdorferi infection in western Switzerland following a tick bite. AB - The aim of this study was to define the risk of developing Lyme borreliosis after a tick bite. A survey was conducted from 1993 to 1995 in the western part of Switzerland in a group of patients who presented for treatment of a recent tick bite. Only patients with negative serological tests (enzyme-linked fluorescent assay screening test, and IgG and IgM immunoblots) at the first consultation and for whom a second blood sample was available 2 months later were included in the study. Of the 376 patients included, 266 had no clinical manifestation (group 1) and 110 had a small local cutaneous reaction (<2 cm) (group 2). The tick was available for 160 patients. Seroconversion was observed in 4.5% of 376 patients, 3.4% in group 1 and 7.2% in group 2. Typical erythema migrans, confirmed by seroconversion, was observed in three of 376 (0.8%) patients, while five of 376 (1.3%) patients developed a skin lesion without seroconversion. No other clinical manifestation of Lyme borreliosis was observed among these 376 patients. Borrelia detection in ticks did not correlate significantly with the risk of Lyme borreliosis. In conclusion, the risk of developing Lyme borreliosis in western Switzerland after a tick bite is low, and therefore, prophylactic antibiotics are not required. PMID- 15278728 TI - Meningococcal urethritis in a heterosexual man. PMID- 15278729 TI - First isolation of Rickettsia conorii from humans in the Trakya (European) region of Turkey. AB - In the Trakya region of Turkey, located in the European part of the country, presumptive cases of Mediterranean spotted fever have been diagnosed and treated every summer since the beginning of the 1990s. The aim of this prospective study was to isolate and identify the rickettsial strains from blood samples of 11 patients and from skin biopsies of 10 of these 11 patients with the diagnosis of spotted fever in the Trakya region of Turkey in 2003. Immunofluorescence assay was performed with acute-phase and convalescent-phase serum samples of 11 patients. All patients had significant antibody titers against spotted fever group rickettsiae. Rickettsia conorii was isolated from the skin biopsies of three of ten patients and was also demonstrated by polymerase chain reaction in skin biopsies of nine of ten patients. In southeastern Europe, the Balkan Peninsula (including the Trakya region of Turkey) is an area where arthropods are endemic and where new arthropod-borne infections can be detected. PMID- 15278730 TI - Cavitating pneumonia after treatment with infliximab and prednisone. AB - Tumor necrosis factor (TNF)-alpha antagonists constitute a novel class of immunomodulating drugs that are used for the treatment of an increasing number of inflammatory disorders. These agents are associated with an increased risk of tuberculosis, but the risk of other infections is less clear. Reported here is the case of a patient who developed cavitary pneumonia after treatment with infliximab (monoclonal TNF-alpha antibodies) and corticosteroids for rheumatoid arthritis. Cryptococcus neoformans was the only pathogen isolated from bronchoalveolar lavage fluid. The patient responded well to fluconazole. The risk of infection after treatment with TNF-alpha antagonists and the possible causative microorganisms are discussed. PMID- 15278732 TI - Prevalence of mutations in dihydrofolate reductase and dihydropteroate synthetase genes of Plasmodium falciparum: analysis of imported cases of malaria in Italy. PMID- 15278733 TI - The role of humans in the cognitive development of apes revisited. PMID- 15278743 TI - Mixed connective tissue disease with interstitial pneumonia in HTLV-1 carrier: case report and review of the literature. AB - We report on a carrier of human T-lymphotropic virus type 1 (HTLV-1) who developed mixed connective tissue disease (MCTD). This patient suddenly manifested clinical symptoms and interstitial pneumonia ascribable to MCTD following long-term infection with HTLV-1. After initiation of oral prednisolone all manifestations quickly improved in parallel with a decrease in inflammatory reactions. In this patient HTLV-1 infection might have played an important role in the pathogenesis of MCTD. Since HTLV-1 can cause adult T-cell leukemia and HTLV-1-associated myelopathy, and also collagen diseases including MCTD, careful observation is necessary even in a carrier, particularly when autoantibodies are detectable in serum. PMID- 15278746 TI - Lupus arthropathy: historical evolution from deforming arthritis to rhupus. AB - Systemic lupus erythematosus is an autoimmune and inflammatory disease with multiple clinical manifestations, including arthropathy. The clinical presentation of articular involvement is variable, ranging from arthralgia without erosions or deformity to an erosive arthropathy and severe functional disability. A subset of patients with this articular involvement have Jaccoud's arthropathy, and others have an arthropathy with clinical findings similar to rheumatoid arthritis that has been called "rhupus." In this paper we review the historical evolution of concepts of lupus arthropathy, from deforming arthritis to rhupus, and conclude that rhupus is not a combination of rheumatoid arthritis and lupus. Instead, rhupus arthropathy should be regarded as a variant of the arthropathy of systemic lupus erythematosus. PMID- 15278747 TI - Cellulitis caused by Capnocytophaga cynodegmi associated with etanercept treatment in a patient with rheumatoid arthritis. PMID- 15278749 TI - Effects of alendronate on metacarpal and lumbar bone mineral density, bone resorption, and chronic back pain in postmenopausal women with osteoporosis. AB - The purpose of this study was to investigate the effect of alendronate on metacarpal and lumbar bone mineral density (BMD), bone resorption, and chronic back pain in postmenopausal women with osteoporosis. Eighty postmenopausal women with osteoporosis, 59-88 years of age, were divided into two groups of 40 each according to the site of BMD measurement: the metacarpus (M) and the lumbar spine (L). All of them were treated with alendronate (5 mg/day) for 12 months. Metacarpal or lumbar BMD was measured by computed X-ray densitometry or dual energy X-ray absorptiometry in the M or the L group, respectively, at baseline and every 6 months. Urinary cross-linked N-terminal telopeptides of type I collagen (NTX) were measured by enzyme-linked immunosorbent assay, and chronic back pain was evaluated by face scale score at baseline and every 6 months in both groups. There were no significant differences in baseline characteristics, including age, body mass index, years since menopause, urinary NTX level, face scale score, or number of prevalent vertebral fractures per patient between the two groups. Urinary NTX level was reduced and chronic back pain was improved similarly in both groups. Whereas metacarpal BMD did not significantly change in the M group (0.20% increase), lumbar BMD increased by 8.15% in the L group. These results suggest that although alendronate increases BMD of the lumbar spine, which is rich in cancellous bone, and improves chronic back pain, with suppression of bone resorption in postmenopausal women with osteoporosis, it may fail to increase cortical BMD of the metacarpus, a distal site of the upper extremity. PMID- 15278750 TI - Extracorporeal shock wave therapy for chronic calcific tendinitis of the shoulder. PMID- 15278751 TI - Autoimmune manifestations in common variable immunodeficiency. AB - Common variable immunodeficiency (CVID) is a disorder characterized by decreased serum immunoglobulin concentrations and increased incidence of recurrent infections. Interestingly 20-25% of patients with CVID develop clinical features suggestive of an autoimmune disease. Although this association is well established, the immunodeficiency background of CVID patients manifesting autoimmune disorders is often overlooked. This study describes three CVID patients displaying a variety of autoimmune manifestations. The pathophysiologic mechanisms of autoimmunity in CVID are also reviewed. PMID- 15278752 TI - Pattern of neuropsychiatric manifestations and outcome in juvenile systemic lupus erythematosus. AB - The aim of this study was to study the neuropsychiatric (NP) manifestations, diagnostic evaluation, treatment and outcome in juvenile systemic lupus erythematosus (SLE). We reviewed the charts of all children with SLE and evidence of NP manifestations as defined by the presence of at least one of the following: headache, cerebrovascular accident (CVA), chorea, seizure, papilledema, and psychiatric or spinal cord manifestations. Out of 90 children with SLE, 20 (16 female) had NP manifestations. The mean age at onset was 8.8 years. The mean period between onset of SLE and NP manifestations was 10.2 months. NP manifestations were the presenting feature in 3 patients. Eleven patients had headache, 10 had psychiatric manifestations, 10 had seizure and 6 had CVA. Coma was seen in 5 patients, chorea in 4, transverse myelitis in 2 and papilledema in 2. Anticardolipin antibodies were high in 12 patients. Five patients had an abnormal CSF study. Nine patients had EEG abnormalities and 13 showed MRI abnormalities. All patients received oral prednisone and 17 were treated with IVMP and immunosuppressive therapy (cyclophosphamide or azathioprine); 85% of our patients recovered completely, but 15% had persistent NP sequelae; 10% died from severe infection. In conclusion, NP involvement in juvenile SLE is common. However, early diagnosis and early treatment with adjunctive intravenous pulse cyclophosphamide may improve the outcome. PMID- 15278753 TI - Oral enzyme combination versus diclofenac in the treatment of osteoarthritis of the knee--a double-blind prospective randomized study. AB - The aim of this study was to compare the efficacy and safety of an oral enzyme rutosid combination (ERC) containing rutosid and the enzymes bromelain and trypsin, with that of diclofenac in patients with osteoarthritis (OA) of the knee. A total of 103 patients presenting with painful episodes of OA of the knee were treated for 6 weeks in two study centers in a randomized, double-blind, parallel group trial. Altogether, 52 patients were treated in the ERC group and 51 patients were treated in the diclofenac group. Primary efficacy criteria were Lequesne's Algofunctional Index (LFI) and a 'complaint index', including pain at rest, pain on motion and restricted function. The efficacy criteria were analyzed by applying the Wilcoxon-Mann-Whitney test that provides the Mann-Whitney estimator (MW) as a measure of relevance. Non-inferiority was considered to be proven if the lower bound of the 97.5% one-sided confidence interval (CI-LB) was higher than MW = 0.36 (benchmark of not yet relevant inferiority). Both treatments resulted in clear improvements. Within the 6-week observation period, the mean value of the LFI decreased from 13.0 to 9.4 in the ERC group and from 12.5 to 9.4 in the diclofenac group. Non-inferiority of ERC was demonstrated by both primary criteria, LFI (MW = 0.5305; CI-LB = 0.4171) and complaint index (MW = 0.5434; CI-LB = 0.4296). Considerable improvements were also seen in secondary efficacy criteria, with a slight tendency towards superiority of ERC. The global judgment of efficacy by physician resulted in at least good ratings for 51.4% of the ERC patients, and for 37.2% of the diclofenac patients. In the majority of patients tolerability was judged in both drug groups as very good or good. The current study indicates that ERC can be considered as an effective and safe alternative to NSAIDs such as diclofenac in the treatment of painful episodes of OA of the knee. Placebo-controlled studies are now needed to confirm these results. PMID- 15278754 TI - The significance of IgA class of antineutrophil cytoplasmic antibodies (ANCA) in childhood Henoch-Schonlein purpura. AB - Antineutrophil cytoplasmic antibodies (ANCA) have been identified in a wide variety of vasculitic disorders, but it is controversial whether ANCA are present in the sera of patients with HSP. This prospective study was designed to assess the place of ANCA, particularly their IgA subclass, in HSP. Thirty-five patients (18 boys, 17 girls) aged 9.4 +/- 4 (3-16) years with a clinical diagnosis of HSP based on American College of Rheumatology (ACR) criteria were enrolled. Thirteen patients (6 boys, 7 girls) aged 8.3 +/- 5.5 (2-21) years with other vasculitides, consisting of classic polyarteritis nodosa (PAN) (n = 2); cutaneous polyarteritis nodosa (n = 1); acute infantile hemorrhagic edema (n = 2); acute urticarial vasculitis (n = 2); hypocomplementemic vasculitis (n = 1); and unclassified vasculitis (n = 5) served as disease controls and 10 healthy children served as normal controls. Twenty-five HSP patients and 7 disease controls were re evaluated in the resolution phase that was described as 4-6 weeks after all symptoms subsided and all medications were stopped. Blood samples for ANCA and IgA rheumatoid factor (RF) were studied by indirect immunofluorescence (IIF) and ELISA, respectively. IgG ANCA was significantly lower in percentage in HSP patients (2.8%) than in disease controls (40%) (p = 0.002). In contrast, IgA ANCA in cytoplasmic pattern was detected in a significantly higher percentage of HSP patients (82.3%) in the acute phase compared to those in the disease controls (38%) (p = 0.004). In the resolution phase, IgA ANCA was negative in 88% of the patients (p = 0.001 for acute vs resolution phases). Neither IgG nor IgA ANCA were seen in normal controls. No relationship was found between disease severity of HSP and IgA ANCA. Positive IgA rheumatoid factor was present in only two patients with HSP. In conclusion, our results suggest that IgA ANCA may be useful to confirm the diagnosis of HSP in children. PMID- 15278755 TI - Successful treatment of rheumatoid arthritis is associated with a reduction in serum sE-selectin and thrombomodulin level. AB - The aim of this study was to investigate the changes in serum levels of endothelial cell injury markers, soluble (s) E-selectin and thrombomodulin (TM), in patients with rheumatoid arthritis (RA) before and after antirheumatic drug treatment and to assess the relationship between these changes and clinical responses to the drug treatment. Eleven patients with RA having active arthritis and 12 healthy volunteers were enrolled in the study. They were monitored by clinical and laboratory parameters while receiving a combination of methotrexate, hydroxychloroquine and sulphasalazine. Pre- and post-treatment clinical and laboratory parameters, including sE-selectin and sTM levels, were measured. The ages of the patients were comparable with those of the control groups. Significant improvements were detected in erythrocyte sedimentation rate, C reactive protein, hemoglobin, morning stiffness, patients' global assessment, physicians' global assessment, number of tender joints and number of swollen joints improved at the end of the therapy (for each parameter p < 0.05). Significant improvements were detected in clinical and laboratory parameters. In the patient group there were significant decreases in the levels of sTM and sE selectin after treatment (p < 0.05). The patient group had significantly higher sTM and sE-selectin levels than the control group at the beginning of the study (p < 0.01), but the difference returned to normal after the treatment (p > 0.05). The sE-selectin and sTM levels significantly correlated with each other, and also with clinical and laboratory findings. Combination treatment successfully treated RA patients. sE-selectin and sTM levels probably reflect disease activity and can be helpful in monitoring disease status and response to therapy. PMID- 15278756 TI - Alexithymia and anger in patients with fibromyalgia. AB - Our objective was to delineate the relevance of the personality construct alexithymia and anger-in in patients with fibromyalgia syndrome. Fifty subjects with fibromyalgia syndrome were compared to 20 subjects with rheumatoid arthritis and 42 healthy controls on the measures of anxiety, depression, anger, alexithymia, pain intensity and disability. There was a significant difference on the measures of anxiety and anger between FMS and RA groups, and also between FMS patients and healthy controls. There was a significant difference between FMS patients and healthy controls on the measures of depression, difficulty in identifying feelings subscale of TAS (TAS-dif), and total alexithymia scores. When the severity of pain was controlled for, there was a significant difference on the measures of anger and alexithymia between the FMS and the RA groups. Fibromyalgia patients were more alexithymic than rheumatoid arthritis patients even when the level of depression was controlled for. Anger towards oneself, which is anger-in, was higher in patients with fibromyalgia patients than in the rheumatoid arthritis sample. A stepwise regression model showed that the anger out scores and the anxiety scores predicted the level of pain severity, and this explained 32% of the variance in the fibromyalgia syndrome group. Although anger in is consistently higher in fibromyalgia patients, it is the behavioral expression of anger, together with anxiety, that predicts the severity of the pain. The difficulty of identifying feelings, rather than other dimensions of alexithymia, seems to be associated with fibromyalgia. PMID- 15278757 TI - Clinical and radiographic features mimicking pulmonary embolism as the first manifestation of Takayasu's arteritis. AB - The current report describes a female patient with Takayasu's arteritis who was hospitalized because of respiratory complaints and a ventilation-perfusion (V/P) scan demonstrating severe perfusion defects with normal ventilation. Her symptoms and V/P scan results were initially interpreted as massive pulmonary embolism, but a spiral computed tomography (CT) of the chest and a repeated lung scan indicated inflammatory involvement of the pulmonary artery accounting for the positive lung scan study. Review of her medical records from 17 years ago revealed that similar symptoms and a positive V/P scan were the initial features of her disease. PMID- 15278758 TI - Is local subacromial corticosteroid injection beneficial in subacromial impingement syndrome? AB - Subacromial corticosteroid injection is one of the most frequently used management tools in subacromial impingement syndrome (SIS) despite controversial reports on the efficacy. Our purpose, in this single blinded, randomised and controlled study was to clarify whether the corticosteroid injection provides additional benefit when used with other conservative treatment modalities in 48 patients with stage 2 SIS. The patients were randomly divided into three groups according to the two therapeutic injections applied with a 10-day interval: group 1: 10 cc of 1% lignocaine + 40 mg of methylprednisolone for the first and second injections, group 2: 10 cc of 1% lignocaine + 40 mg of methylprednisolone for the first injection and only 10 cc of 1% lignocaine for the second injection, group 3: only 10 cc of 1% lignocaine for the first and second injections. All the patients were prescribed 500 mg of naproxen sodium to use two times daily, instructed to rest and perform Codman's pendulum exercises during the first 15 days. Shoulder pain during rest, activity, and causing disturbance of sleep was evaluated using a visual analogue scale and shoulder function was investigated by total Constant score and its subsectional parameters which are pain, daily living activities, active range of motion and strength before the therapy and 1 and 3 months after the therapy onset. Significant improvements from the baseline values in all pain and function parameters were observed at the first and second evaluation in all groups. Group 1 patients had more favourably improved values in pain causing sleep disturbance and daily living activity parameters than group 2 and 3 patients only in the 1st month after therapy onset. We found that subacromial corticosteroid injections in the acute or subacute phase of SIS provided additional short-term benefit without any complication when used together with nonsteroidal anti-inflammatory drugs (NSAIDs) and exercise. PMID- 15278759 TI - Knee osteoarthritis: interpretation variability of radiological signs. AB - The aim of this study was (1) to determine the variability in detecting radiological signs of knee osteoarthritis (OA) between an orthopaedics specialist, a fourth-year resident in the speciality and a recently qualified doctor and (2) to determine which of the existing criteria show the greatest variability when used by the three participants to detect the degree of evolution of the pathology. This observational study included radiographs of 95 patients with knee pain. Osteophytes, narrowing of joint space (excluding inter-osteophyte bridges) subchondral sclerosis, subchondral cysts, collapse of the central joint cortical bone and lateral deformity, according to the criteria of Kellgren and Lawrence, modified by Kallman et al. were evaluated. Anteroposterior radiographs were used. Knees that had undergone previous surgery were excluded. Cohen's kappa index was used to calculate the degree of agreement between observers. The concordance analysis showed a low level of agreement among the three observers of the radiological variables with a maximum of 50% in some parameters. The authors discuss the possible causes of this low level of agreement. The low degree of agreement of 50% among the three observers is in line with previous reports and suggests that better training of observers is necessary and that the use of any classification is problematic. PMID- 15278761 TI - CTLA4 exon 1 polymorphism, rheumatoid arthritis and autoimmune endocrinopathy. PMID- 15278762 TI - Molecular genetic analyses of beta-thalassemia in South India reveals rare mutations in the beta-globin gene. AB - beta-Thalassemia is the most prevalent single-gene disorder. Since no viable forms of treatment are available, the best course is prevention through prenatal diagnosis. In the present study, the prevalence of beta-thalassemia was extensively investigated in the South Indian population, especially from the state of Andhra Pradesh. Screening for causal mutations was carried out on genomic DNA isolated from patient blood samples by using the routine reverse dot blot (RDB) and amplification refractory mutation system-polymerase chain reaction (ARMS-PCR) techniques. DNA sequencing was performed wherever necessary. Among the nine mutations identified, four, including IVS-1-5 (G-C) (IVS1+5G>T), codon 41/42 (-TTCT) (c.124_127delTTCT), codon 15 (G-A) (c.47G>A), and HbS (sickle mutation) (c.20A>T) mutations, accounted for about 98% of the total positive cases. Two mutations viz. codon 8/9 (+G) (c.27_28insG) and HbE (codon 26 G-A) (c.79G>A) exhibited a very low frequency of occurrence, whereas the IVS-1-1 (G-T) (IVS1+1G>T) and the 619 bp deletion (c.366_494del) mutations were absent. We also identified certain rare mutations during the diagnostic evaluation. Gene sequencing confirmed the codon 30 (G-C) (c.92G>C) mutation and the rare codon 5 ( CT) (c.17_18delCT) and IVS-II-837 (T-G) (IVSII-14T>G) mutations. This is the first report of the IVS II 837 mutation in the Indian population. We also report a novel diagnostic application during RDB-based screening for the detection of the (c.92G>C) mutations. Such a comprehensive mutation screening is essential for prenatal diagnosis of beta-thalassemia and control of this highly prevalent monogenic disorder in the Indian population. PMID- 15278763 TI - Phylogeographic analysis of mitochondrial DNA haplogroup F2 in China reveals T12338C in the initiation codon of the ND5 gene not to be pathogenic. AB - In this report, we studied on a homoplasmic T12338C change in mitochondrial DNA (mtDNA), which substituted methionine in the translational initiation codon of the NADH dehydrogenase subunit 5 gene ( ND5) with threonine. This nucleotide change was originally identified in two mtDNAs belonging to haplogroup F2 by our previous complete sequencing of 48 mtDNAs. Since then, a total of 76 F2 mtDNAs have been identified by the variations occurring in the hypervariable segments and coding regions among more than 3,000 individuals across China. As the T12338C change was detected in 32 samples representing various sub-clades of the F2 haplogroup while not in 14 non-F2 controls, we believe that the T12338C change is specific to the F2 haplogroup. As F2 and its sub-clades were widely distributed in normal individuals of various Chinese populations, we conclude that T12338C is not pathogenic. In addition, based on the average distribution frequency, haplotype diversity and nucleotide diversity of haplogroup F2 in the populations across China, the T12338C nucleotide substitution seems to have been occurred in north China about 42,000 years ago. Our results provided a good paradigm for distinguishing a polymorphic change from a pathogenic mutation based on mtDNA phylogeny. PMID- 15278764 TI - Identification of variants in cyclin D1 ( CCND1) and B-Cell CLL/lymphoma 2 ( BCL2). AB - CCND1 is an important cell-cycle regulatory protein associated with cell proliferation, poor prognosis and recurrence in cancer, while BCL2 is an important anti-apoptotic protein that plays a vital role in the regulation of the life span by controlling the rate of apoptosis. Recent studies have shown that CCND1 and BCL2 may be responsible for the body mass and the regulation of various metabolic processes. In an effort to discover additional polymorphism(s), we scrutinized the genetic polymorphisms in the CCND1 and BCL2. By direct DNA sequencing in 24 individuals, we identified 22 sequence variants within the 16 kb of whole CCND1 gene: one in exon 4, 17 in introns and four in the 3' UTR region. We also found eight sequence variants within 7.5 kb exon-intron boundaries of BCL2 gene: one in promoter, three in exon 1, and four in the 3' UTR region. Haplotypes, their frequencies and linkage disequilibrium coefficients (| D'| and r(2)), among polymorphisms were estimated. Among identified variants, seven and six variants of CCND1 and BCL2 were genotyped in a larger series of subjects ( n=320). Statistical analyses of CCND1 and BCL2 polymorphisms with two metabolic phenotypes revealed no significant association. The information concerning genetic polymorphisms of CCND1 and BCL2 might provide valuable information for future genetic studies of diseases. PMID- 15278765 TI - Comparison between various strategies for the disease-gene mapping using linkage disequilibrium analyses: studies on adenine phosphoribosyltransferase deficiency used as an example. AB - Recently, linkage disequilibrium analyses have been used to detect disease causing loci based on the common disease-common variant hypothesis. To see what methods can effectively identify the genes, we have to apply them to the practical data obtained from the human population. We extensively performed linkage disequilibrium and haplotype analyses on adenine phosphoribosyltransferase ( APRT) genes in both control and deficient subjects. To examine the power to detect disease-causing loci, we analyzed SNPs, STRPs, and VNTR within and around the APRT gene. When only SNPs were used, P values did not necessarily show significant difference, even at loci close to the mutation site for APRT*J that is exclusively observed among Japanese. However, the examination of the same samples with haplotypes based on the haplotype block data gave sufficient significance. In the case of STRP and VNTR, some single-marker loci showed significant difference. Our study suggested that the use of haplotype analysis based on the haplotype-block structure is more powerful than single marker locus analysis for the detection of disease-related loci. PMID- 15278766 TI - Large scrotal hernia: a complicated case of mesh migration, ascites, and bowel strangulation. AB - A 30-year-old male with 1 1/2-year history of an asymptomatic, large, reducible right indirect scrotal hernia presented to the emergency department complaining of a 2-week history of increasing abdominal distension and daily emesis. He had recently undergone an emergent exploratory laparotomy in which his asymptomatic hernia was repaired with a mesh plug from an intra-abdominal approach. The mesh plug subsequently migrated into the patient's scrotum resulting in a strangulating bowel obstruction. This paper discusses a serious complication that may result from inappropriate use and placement of a mesh plug and our approach to correct the situation utilizing a bioabsorbable mesh prosthesis. PMID- 15278768 TI - Engineering the thermostability of Trichoderma reesei endo-1,4-beta-xylanase II by combination of disulphide bridges. AB - Disulphide bridges were introduced in different combinations into the N-terminal region and the single alpha-helix of mesophilic Trichoderma reesei xylanase II (TRX II). We used earlier disulphide-bridge data and designed new disulphide bridges for the combination mutants. The most stable mutant contained two disulphide bridges (between positions 2 and 28 and between positions 110 and 154, respectively) and the mutations N11D, N38E, and Q162H. With a half-life of approximately 56 h at 65 degrees C, the thermostability of this sevenfold mutant was approximately 5,000 times higher than that of TRX II, and the half-life was 25 min even at 75 degrees C. The thermostability of this mutant was approximately 30 times higher than that of the corresponding mutant missing the bridge between positions 2 and 28. The extensive stabilization at two protein regions did not alter the kinetic properties of the sevenfold mutant from that of the wild-type TRX II. The combination of disulphide bridges enhanced significantly the pH dependent stability in a wide pH range. PMID- 15278770 TI - Watching for falling apples. PMID- 15278771 TI - Significance of preoperative position of the femoral head in failed closed reduction in developmental dislocation of the hip: surgical results. AB - Long-term follow-up results of open reduction for developmental dislocation of 83 hips via the extensive anterolateral approach were retrospectively analyzed. Open reduction was performed in infancy, and the follow-up period ranged from 12 to 24 years. This procedure is a complete circumferential dissection of the joint capsule and produces sufficient concentric reduction of the femoral head in the acetabulum immediately after the surgery. A lateral arthrographic classification of interposed limbus and the preoperative position of the unreduced femoral head is introduced, and is related to operative findings and surgical results, including Severin's classification. The results at the final follow-up were: Severin's group I in 35 hips, group II in 19 hips, group III in 10 hips, and group IV in four hips. According to the classification of the preoperative position of the femoral head, there were 31 hips of the intracapsular type and 37 hips of the extracapsular type. Thirty-four of the 37 hips of the extracapsular type were classified in Severin's group I or II (92%). Twenty of the 31 hips of the intracapsular type were classified in Severin's group I or II (65%). A very significantly greater number of hips with good radiological outcome were in the extracapsular type than in the intracapsular type. PMID- 15278772 TI - Arthroscopic surgery for traumatic triangular fibrocartilage complex injury. AB - Clinical characteristics, diagnostic imaging, arthroscopic findings, and surgical results of arthroscopic treatment are reported in 62 hands with traumatic triangular fibrocartilage complex (TFCC) injury treated between 1995 and 2002. This retrospective study also describes and compares the results of a novel arthroscopic suture technique for repairing tears with Palmer's class 1B and 1D tears. According to Palmer's classification, there were 10 class 1A, 27 1B, 8 1C, and 17 1D injuries. The arthroscopic suture and debridement treatments were done under brachial plexus block as a 1-day in-hospital procedure. Class 1D tears required transradial fixation to anchor the TFCC. Surgical results of suture groups according to Minami's criteria were excellent in 16 hands, good in 15 hands, fair in 1 hand, and poor in 1 hand. The results of the debridement group were excellent in 16 hands, good in 10 hands, fair in 2 hands, and poor in 1 hand. The results of the current study indicate that the novel suture technique were easy to perform, provided strong, tight repair for class 1D injuries, and was applicable for class 1B injuries in this small case series. PMID- 15278773 TI - Physical and mental health in young adults operated on for idiopathic scoliosis. AB - In this study, physical and mental health were investigated in 30 young adults who were operated on for idiopathic scoliosis, 2-3 years after surgery, and the results compared to an age- and sex-matched control group of 40 individuals. We used the short form of the 36 health survey (SF-36 version 1.2), which is a 36 item questionnaire measuring health functioning on eight scales: physical functioning, role limitations due to physical functioning, bodily pain, general health perceptions, vitality, social functioning, emotional role limitation, and general mental health. The patient scores indicated lower than the controls in physical health but higher in mental health. Otherwise, there were no significant differences between the two groups. Overall, this study showed that young adults operated on for idiopathic scoliosis were satisfied, and that their mental health was even better than the normal group, but their physical health was somewhat poorer. Thus, the surgical procedure was well tolerated and had not traumatized the patient. PMID- 15278774 TI - Effects of potassium and anion channel blockers on the cellular response of human osteoarthritic chondrocytes. AB - The aim of this article is to determine to what extent the proliferation, CD44 expression, and apoptosis behavior of cells can be influenced by the modulation of ion channel activity on the cell membrane of human osteoarthritic chondrocytes. The potassium channel blocker 4-aminopyridine (4-AP) and the chloride and anion channel blocker 4-acetamido-4'-isothiocyano-2,2'-disulfonic acid stilbene (SITS) were used as ion channel modulators. Assessment of the proliferation was done by incorporation of (3)H-thymidine. The detection of apoptotic cells and expression of the hyaluronic acid binding CD44-receptor were determined by flow cytometry. The results showed that 4-AP and SITS lead to a temporary increase in (3)H-thymidine incorporation, followed by a suppression of proliferation after a 12-day incubation. 4-AP causes considerable cytotoxic effects. SITS leads to necrotic cell damage. CD44 expression is increased up to 43% after incubation with 4-AP for 24 or 48 h, whereas prolonged incubation under SITS influence leads to a clear inhibition of CD44 expression. In conclusion, proliferation, CD44 expression, and apoptosis behavior of human chondrocytes can be influenced by modulation of ion channel activity. These results serve as a basis for further investigations to extend the therapeutic possibilities in the treatment of arthritis. PMID- 15278775 TI - Double-band reconstruction of the ACL using a synthetic implant: a cadaveric study of knee laxity. AB - In this study, the anterior laxity and internal rotation of five cadaveric knee joints were compared when the anterior cruciate ligament (ACL) was intact, after its reconstruction with the anteromedial band (AMB) only, then after its reconstruction with the double band, with the posterolateral band (PLB) tensioned first at 20 degrees and then at 90 degrees, and finally with the ACL resected. The tests were performed using a mechanical apparatus that allowed the joint 6 degrees of freedom and also the application of external loads and torques on the tibia. The loads used were 50, 90, and 130 N for the anterior laxity test, and a torque of 2, 3, and 4 Nm in the internal rotation test. In all cases, laxity with double-band reconstruction was closer to the natural value than when it was constructed with the AMB only. In some cases, double-band reconstruction imposed a higher constraint on the joint than did the natural ACL. Measurement of the residual tension on the PLB after its final anchoring was also performed during passive flexion. This test revealed a high tension on this band with the knee in hyperextension, followed by a decrease in value through to 45 degrees and a slight increase at 90 degrees, thus following a similar trend to that of the natural PLB. PMID- 15278776 TI - Factors for the presence of anteromedial rotatory instability of the knee. AB - Anteromedial rotatory instability (AMRI) of the knee joint was investigated with an instrument newly designed to simulate the manual AMRI test and to quantify its magnitude. Thirty healthy subjects, 20 patients with anterior cruciate ligament (ACL) injury, and 10 with both ACL and medial collateral ligament (MCL) injuries were examined. Using the instrument, 100 N of anterior force was applied to the proximal part of the tibia with the foot in neutral rotation, 30 degrees of internal rotation, and 30 degrees of external rotation, and the magnitude of anterior displacement was recorded. The measurement was carried out at 20 degrees and 90 degrees of flexion. A significant increase in anterior laxity was observed in all three rotation positions in the injured patients. However, the magnitude of laxity in external rotation was less than that in neutral rotation in the ACL injured patients, whereas it was the greatest in external rotation in ACL + MCL injured patients. Thus, we conclude that an injury involving both the ACL and MCL causes AMRI. PMID- 15278777 TI - Electrophysiological study on primary afferent properties of a chronic constriction nerve injury model in spinal rats. AB - A chronic constriction nerve injury (CCI) model of the rat sciatic nerve is known to exhibit neuropathic pain behavior. The authors conducted electrophysiological analysis for the primary afferent properties of this model in a decerebrate spinal preparation. In the CCI model, electrical transcutaneous stimulation for A delta and C-fibers showed a low current threshold to elicit the flexion withdrawal reflex. The antidromic ectopic firing activity recorded from the sural nerve showed abnormal firing patterns, which were not seen in normal rats, as follows: (1) an increase of spontaneous firing frequency, (2) development of an on-off pattern that consisted of cyclic burst spikes, and (3) increased firing number under the hypoxic condition. The amplitude of the A-delta component in the antidromic sensory nerve-evoked potential was lower than that in normal rats. The current study clarified the electrophysiological parameters reflecting pathological hypersensivity and excitability of primary afferents in the CCI model, which could not found by behavioral analysis. These results may be useful in future studies evaluating possible treatments of neuropathic pain. PMID- 15278778 TI - Ankle tuberculosis: a report of four cases in a Japanese hospital. AB - We report four patients suffering from ankle tuberculosis, which is an uncommon disease in Japan. All patients complained of swelling and pain in the affected ankle. Ankle tuberculosis is a disorder that can be easily misdiagnosed. The mean delay in diagnosis was 5.3 months in our series. All patients had already been diagnosed as having ankle tuberculosis by the time they visited our hospital in this series. The patients had been definitively diagnosed to have ankle tuberculosis by means of a needle biopsy in two cases, an open biopsy in one case, and on the basis of the clinical features in one case. All patients received antitubercular therapy and non-weight-bearing protective treatment. Antitubercular therapy was performed for an average of 14.3 months, and non weight-bearing treatment was maintained for a mean of 4.5 months. The main treatments for such cases include open biopsy for an uncertain diagnosis, debridement for intractable synovitis, and arthrodesis for severely destroyed ankles with pain. All patients received some form of surgical intervention either from previous doctors or from us in this series. We conclude that most patients who suffer from ankle tuberculosis seem to need surgery in addition to adequate chemotherapy and non-weight-bearing protective treatment. PMID- 15278779 TI - MR imaging findings of an unusual case of myositis ossificans presenting as a progressive mass with features of fluid-fluid level. AB - We present a case of myositis ossificans involving the upper arm in which features of fluid-fluid level became apparent on magnetic resonance imaging (MRI). Serial MR images obtained over a 6-month period exhibited progressive appearances in the absence of dense mineralization in the early and mid phases, which simulate neoplastic conditions. Twenty-four weeks following the biopsy, MR images revealed that the mass had decreased in size with the disappearance of most of the features of fluid-fluid level. To the best of our knowledge, no example of myositis ossificans accompanied by features of fluid-fluid level, which was closely monitored by MRI, exists in the literature. These features of image examination should be evaluated cautiously to avoid unnecessary surgical intervention, especially in instances where lesions exhibit expansive appearance. This case provided beneficial information regarding the sequence of changes in terms of MR appearance of myositis ossificans. PMID- 15278780 TI - Acute calcific tendinitis of the gluteus medius: a case report with serial magnetic resonance imaging findings. AB - A case of calcific tendinitis of the gluteus medius is presented. Calcification was evident in the soft tissue adjacent to the greater trochanter on plain radiographs. On the initial magnetic resonance images (MRI), inflammatory edematous change was detected not only in the gluteus medius but also in the bone marrow of the greater trochanter, corresponding to the painful area. Three months later, calcification disappeared on plain radiographs and the femur showed normal signal intensity on MRI. Initial MRI excluded other diseases' including infection and bone tumor, and serial MRI confirmed that the change in extraosseous and intraosseous findings were in accordance with self-limiting clinical symptoms. PMID- 15278781 TI - Restoring the continuity of the tibialis posterior tendon in the treatment of symptomatic accessory navicular with flat feet. PMID- 15278782 TI - Distinct osteogenic mechanisms of bones of distinct origins. AB - Mammalian bones have three distinct origins (paraxial mesoderm, lateral plate mesoderm, and neural crest) and undergo two different modes of formation (intra membranous and endochondral). Bones derived from the paraxial mesoderm and lateral plate mesoderm mainly form through the endochondral process. During this process, hypertrophic chondrocytes play a vital role in inducing both osteogenesis and angiogenesis. One of the essential osteogenic factors secreted from hypertrophic chondrocytes is Indian hedgehog (Ihh). In contrast, bones derived from the neural crest mainly form through the intramembranous pro-cess and do not require Ihh. Thus, depending on their origin, bones have distinct signaling properties, which need to be considered in the research and application of bone biology. PMID- 15278783 TI - Metastatic bone disease: pathogenesis and new strategies for treatment. AB - Bone is the third leading site of metastatic disease, after the lung and liver. Pain, pathological fractures, neurological deficits, and forced immobilization significantly decrease the quality of life of patients with bone metastasis. The development of metastasis, from the migration of malignant cells from the primary tumor to their proliferation at a distant site, involves a series of sequential steps: angiogenesis, matrix degradation, cell motility, cell attachment, and cellular proliferation. A better understanding of the pathogenesis of metastasis may be expected to lead to the development of new treatment modalities for bone metastasis. Currently, antiangiogenic agents, matrix metalloproteinase (MMP) inhibitors, and hyperthermia are some of the newer therapeutic modalities that seem to hold promise for the treatment of metastatic bone disease. PMID- 15278784 TI - Solvent-based deuterium isotope effects on the redox thermodynamics of cytochrome c. AB - The reduction thermodynamics of cytochrome c (cytc), determined electrochemically, are found to be sensitive to solvent H/D isotope effects. Reduction of cytochrome c is enthalpically more favored in D(2)O with respect to H(2)O, but is disfavored on entropic grounds. This is consistent with a reduction induced strengthening of the H-bonding network within the hydration sphere of the protein. No significant changes in E degrees ' occur, since the above variations are compensative. As a main result, this work shows that the oxidation-state dependent differences in protein solvation, including electrostatics and solvent reorganization effects, play an important role in determining the individual enthalpy and entropy changes of the reduction process. It is conceivable that this is a common thermodynamic feature of all electron transport metalloproteins. The isotope effects turn out to be sensitive to buffer anions which specifically bind to cytc. Evidence is gained that the solvation thermodynamics of both redox forms of cytc are sensibly affected by strongly hydrated anions. PMID- 15278788 TI - Inhalation of calcium channel blocking agents protects against methacholine induced bronchoconstriction. AB - The calcium channel blockers, diltiazem and verapamil, and the beta agonist orciprenaline sulfate all demonstrated significant protection against methacholine-induced bronchoconstriction in 11 stable asthmatics (5 males and 6 females). Ten and 20 mg of inhaled diltiazem, 5 mg of verapamil or 30 mg of orciprenaline administered 15 min before stepwise increasing doses of methacholine hydrochloride produced significant reduction in respiratory resistance (Rrs), minimum dose of methacholine hydrochloride required for Rrs increase (Dmin) and bronchial reactivity measured with an Astograph. The mechanism of action of the calcium channel blockers is presumably at the level of the smooth muscle cells themselves. The combination of positive influence and lack of any adverse effect on blood pressure or heart rate with any of the agents tested indicates that their clinical application for alleviation of acute asthma can be recommended. PMID- 15278785 TI - SufA/IscA: reactivity studies of a class of scaffold proteins involved in [Fe-S] cluster assembly. AB - IscA/SufA proteins belong to complex protein machineries which are involved in iron-sulfur cluster biosynthesis. They are defined as scaffold proteins from which preassembled clusters are transferred to target apoproteins. The experiments described here demonstrate that the transfer reaction proceeds in two observable steps: a first fast one leading to a protein-protein complex between the cluster donor (SufA/IscA) and the acceptor (biotin synthase), and a slow one consisting of cluster transfer leading to the apoform of the scaffold protein and the holoform of the target protein. Mutation of cysteines in the acceptor protein specifically inhibits the second step of the reaction, showing that these cysteines are involved in the cluster transfer mechanism but not in complex formation. No cluster transfer from IscA to IscU, another scaffold of the isc operon, could be observed, whereas IscU was shown to be an efficient cluster source for cluster assembly in IscA. Implications of these results are discussed. PMID- 15278789 TI - Neuromuscular effects of pipecuronium during sevoflurane anesthesia compared with isoflurane and enflurane anesthesia. AB - We evaluated the neuromuscular effects of pipecuronium during anesthesia with equipotent concentrations of either sevoflurane, isoflurane or enflurane. Twenty seven patients scheduled for minor elective otolaryngeal or plastic surgery were studied and randomly assigned to 3 groups, one group per anesthetic agent. Anesthesia was induced with thiamylal 5 mg.kg(-1) and the trachea was intubated with succinylcholine 1 mg.kg(-1), then anesthesia was maintained with 60% nitrous oxide in oxygen and sevolfurane, isoflurane or enflurane, depending on the group. Neuromuscular blocking effects were monitored by recording the electromyographic activity of the adductor pollicis muscle from supramaximal stimulation of the ulnar nerve at 10-s intervals. Pipecuronium 40 microg.kg(-1) was administered when electromyographic activity had reached a stable state, 30 min after succinylcholine administration. The maximum effect (% block of control) and clinical duration (time to 25% recovery) of pipecuronium were 99.1 +/- 1.4% and 63.7 +/- 14.7 min (mean +/- S.D.) for sevoflurane, 99.0 +/- 2.0% and 60.9 +/- 20.5 min for isoflurane, and 98.0 +/- 2.5% and 62.8 +/- 28.7 min for enflurane, respectively. There were no significant differences in these values between the anesthetics. Cardiovascular stimulant effects were not observed in any of the groups. We conclude that the effect of pipecuronium under seveflurane anesthesia is similar to that under isoflurane and enflurane anesthesia. PMID- 15278790 TI - The effects of butorphanol on baroreflex control of heart rate in man. AB - The effects of butorphanol injection on baroreflex control of heart rate were investigated using both pressor and depressor tests in eighteen adult patients. Baroreflex sensitivity was attenuated after butorphanol injection in the pressor test using phenylephrine, whereas it was unchanged in the depressor test using nitroglycerine. No resetting of the baroreflex occurred after butorphanol injection. After the administration of butorphanol, plasma epinephrine and norepinephrine levels increased. These results suggest that it is safe to use butorphanol clinically even when a reduction in blood pressure due to hypovolemia or unclamping of the major artery is expected and that it is disadvantageous to administer the drug when an increase in blood pressure due to cross-clamping of the major artery is predicted. PMID- 15278791 TI - Natural frequency/damping coefficient relationship of the catheter-manometer system required for high-fidelity measurement of the pulmonary arterial pressure. AB - Using a digital simulation method, we analyzed the relationship between natural frequency (fn) and damping coefficient (Zeta) of the catheter-manometer system required for high-fidelity measurement of the pulmonary arterial pressure. The pulmonary artery pressure waveform was obtained with a catheter-tip transducer and it was fed into a dynamic simulator programmed on a computer. The original waveform and the output of the simulator were compared and judged visually for the fidelity. From this analysis, the combination of fn and Zeta was obtained and was plotted on a fn-Zeta diagram. It showed as an area, which was convex on the left side and open on the right side. The left-convex endpoint was located at a damping coefficient of about 0.7. At a lower heart rate, this area was extended to the lower frequency side, while, at a higher heart rate, this area was limited to the higher frequency side. The fn-Zeta diagram was also constructed theoretically by calculating the relations between natural frequencies and damping coefficients of a second order system with the amplitude and phase error tolerance set at +/-5% respectively. PMID- 15278792 TI - Simultaneous laser-Doppler flowmetry of canine spinal cord and cerebral blood flow: responses to PaCO2 and blood pressure changes. AB - We observed the relative changes of both spinal cord blood flow (local SCBF) and local cerebral blood flow (local CBF) using independent laser-Doppler flowmeters (LDF) in 12 dogs under N2O(50%)-O2-enflurane(1.0%) anesthesia. The dorsal surface of the lumbar spinal cord and the parietal surface of the brain were partially exposed. Two fine LDF probes were placed between the exposed surfaces and the dura maters at each site. Both local SCBF and local CBF decreased simultaneously with hyperventilation and increased with hypoventilation within several seconds. The local SCBF responses to PaCO2 changes were similar in direction and degree as those of the local CBF. Autoregulation of local SCBF to arterial blood pressure (ABP) changes was abolished, though that of the local CBF was still recognized in a blunted fashion within a mean ABP range of 50 to 150 mmHg. PMID- 15278793 TI - Effects on enflurane on gill withdrawal behaviors and the ability of gill motor neurones to elicit gill contractions in aplysia. AB - We used the Aplysia gill withdrawal reflex model system in order to study how enflurane effected both gill withdrawal adaptive behaviors and the activity of single identified neurones which are involved with the mediation of the gill withdrawal response. We found that a continuous superfusion of enflurane (0.5 and 1.0%) solution over the abdominal ganglion (the CNS) resulted in an increase in the spontaneous gill respiratory movements; an increase in the spontaneous discharges in identified central motor neurones; and a depolarizing shift in the resting membrane potential of these neurones. Enflurane also significantly effected the ability of the gill motor neurones to elicit a gill contraction when the motor neurone was depolarized to produce action potentials by passing depolarizing current into the neurone. Although in most cases the ability of the motor neurone to elicit a gill withdrawal contraction was decreased, that in one third of the cases was increased. Enflurane may exert its actions by effecting the activity of CNS control neurones which exert both facilitatory and suppressive control over the peripheral nervous system in the gill as well as by having direct effects on the motor neurones. PMID- 15278794 TI - Effect of halothane on intercellular adhesion molecule-1 (ICAM-1) in melanoma cells. AB - There have only been a few reports relating to the effect of inhalational anesthetics on the tumor cell morphology in cancer patients undergoing surgery. We hypothesized that some anesthetic agents might influence the spread of unresectable cancer cells and might additionally worsen the condition of the patient due to depressed host immune surveillance. We therefore evaluated the influence of halothane on tumor cell adhesion, which is closely linked to tumor cell metastasis. Human melanoma cells from SK-MEL-37 cell-line were exposed to 4% halothane for 3, 6, 12 or 24 hours, respectively. Furthermore, after 24 hours halothane exposure, they were incubated in a 5% CO2 atmosphere for 12 or 24 hours. The cells were then analyzed using a fluorescence flowcytometer and intercellular adhesion molecule-1 (ICAM-1) expression in SK-MEL-37 cells was quantified as the intensity of fluorescence of ICAM-1 expressed in 10,000 cells. ICAM-1 expression in cells exposed to halothane for 3, 6, 12 or 24 hours was lower than that of non-exposed cells and returned to control level after further incubation in 5% CO2 atmosphere for either 12 or 24 hours. We conclude that halothane might affect the progression of tumor cell metastasis in vitro. PMID- 15278795 TI - Deuterium chemical shift imaging for the estimation of cerebral perfusion in rabbit infarction model. AB - In order to develop a new technique for the measurement of local cerebral blood flow (CBF), the deuterium chemical shift imaging (2H-CSI) technique, an application of in vivo nuclear magnetic resonance (NMR), was used for the estimation of cerebral perfusion in rabbit infarction model. The 2H chemical shift images of rabbit brain were obtained every 30 seconds before and after intravenous injection of deuterated saline. The changes in 2H NMR signal intensity documented that the cerebral perfusion in the damaged area due to infarction decreased obviously compared to that in the intact area. These findings indicate that the 2H-CSI technique can be applied to the measurement of local CBF. The readily availability and limited toxicity of deuterated water may make possible to use this method in clinical cases. PMID- 15278796 TI - Effects of halothane and isoflurane anesthesia on sympathetic adrenal nerve responses to carbon dioxide challenge in rats. AB - We studied the influence of two volatile anesthetics, halothane and isoflurane, on the circulatory and sympathetic nerve responses to carbon dioxide (9% CO2) in rats. Systolic blood pressure was depressed throughout the CO2 challenge and after an initial reduction, a gradual increase was observed in heart rate. Sympathetic adrenal nerve action potentials (SANA) significantly increased in contrast to negative responses in the circulatory functions. SANA responses against time were trapezoid in shape. There were no significant differences in SANA responses between 1% (1 MAC) and 1.5% (1.5 MAC) halothane groups, nor between 1.4% (1 MAC) and 2% (1.5 MAC) isoflurane groups. Halothane and isoflurane, therefore, did not produce dose-dependent effects on sympathetic response to hypercapnia within these concentrations. The maximum changes in SANA from the baseline values were 110% and 40% for the halothane and isoflurane groups, respectively. The sympathetic reflex response to hyperacapnia was retained at 1.5 MAC for both anesthetics, though isoflurane depressed these responses more markedly than halothane. Our results suggest that halothane is a more preferable anesthetic than isoflurane when viewed from the standpoint of preservation of sympathetic nerve response in such undesirable situations as severe hypercapnia occurring during anesthesia. PMID- 15278797 TI - Naloxone and flumazenil fail to antagonize the isoflurane-induced suppression of dorsal horn neurons in cats. AB - Effects of naloxone and flumazenil on isoflurane activities were examined on dorsal horn neurons in cats. Isoflurane suppressed bradykinin-induced nociceptive responses in transected feline spinal cords. The bradykinin-induced neuronal firing rates were significantly suppressed by 60.0%, 35.3% and 32.2% at 10, 20 and 30 min after isoflurane administration, respectively. The 32.3% suppression on bradykinin-induced neuronal responses at 30 min after isoflurane administration was not reversed 5 min after administration of naloxone (36.4% suppression). The suppressive effects of isoflurane were not reversed by naloxone (0.2 mg.kg(-1), i.v.). Similarly, the benzodiazepine antagonist, flumazenil (0.2 mg.kg(-1), i.v.), did not affect the suppressive effects of isoflurane. Failure of naloxone and flumazenil to reverse the suppressive effects of isoflurane suggests that isoflurane interacts with neither opioid nor benzodiazepine receptors in producing its suppressive action on nociceptive responses in dorsal horn neurons of the feline spinal cord. PMID- 15278798 TI - Compartment syndrome after prolonged lithotomy position in patient receiving combined epidural and general anesthesia. PMID- 15278799 TI - Changes in plasma concentrations of free amino acids during and after the anhepatic period. PMID- 15278800 TI - Massive extrapleural hematoma after attempted internal jugular vein cannulation. PMID- 15278801 TI - Total spinal anesthesia for resection of pheochromocytoma. PMID- 15278802 TI - Pulmonary edema induced by laryngospasm. PMID- 15278803 TI - Anesthetic management of an infant with Cockayne's syndrome. PMID- 15278804 TI - Changes in laryngeal mask airway cuff pressure under general anesthesia with and without nitrous oxide. PMID- 15278805 TI - Coronary artery spasm after intraperitoneal administration of cisplatin and etoposide during anesthesia. PMID- 15278806 TI - Deliberate hypotension induced by epidural anesthesia. PMID- 15278807 TI - Emergency operation in a patient with asymptomatic pheochromocytoma. PMID- 15278808 TI - A case of absent brainstem responses with electroencephalographic activity. PMID- 15278809 TI - The N-methyl-D-aspartate (NMDA) receptor antagonist dizocilpine (MK-801) suppresses enflurane-induced opisthotonus in mice. AB - We determined whether enflurane-induced opisthotonus in ddN mice is mediated by N methyl-D-aspartate (NMDA) receptor using NMDA receptor antagonists dizocilpine (MK-801) and ketamine. Animals were given intraperitoneal injections of 0.2 ml saline (control), 2.5 or 5.0 mg.kg(-1) dizocilpine in saline, or 20 or 40 mg.kg( 1) ketamine is saline 20 min prior to exposure to 2.0% enflurane. Incidence of opisthotonus measured during exposure to enflurane for 20 min was 49% (n = 51) in saline (control) group, 6.7 (P < 0.01 vs control, n = 30) and 15.0% (P < 0.01, n = 40) in 2.5 and 5.0 mg.kg(-1) dizocilpine group, respectively, and 43.9 (NS, n = 41) and 40.0% (NS, n = 40) in 20 and 40 mg.kg(-1) ketamine group, respectively. These results strongly suggest that enflurane-induced opisthotonus is mediated by NMDA receptor. Ketamine failed to suppress significantly due to possibly small dosages. Further, dizocilpine itself produced severe seizures during preenflurane period (30.0 and 40.0% in 2.5 and 5.0 mg.kg(-1), respectively), which may be a novel finding. PMID- 15278810 TI - A case of pharyngeal perforation following endotracheal intubation at anaesthesia for total knee replacement in a 63 year old with rheumatoid arthritis. PMID- 15278811 TI - Epidural block for treatment of intestinal obstruction--clinical and experimental studies. AB - Seventy patients with intestinal obstruction were managed with usual conservative treatments and epidural anesthesia to block splanchnic and somatic nervous systems, for nine years from 1981 to 1990. Improvement of clinical symptoms and general conditions was accomplished in 48 patients (68.6%). In these 48 patients, 41 patients (58.6%) had complete remission of intestinal obstruction, showing flatus in 8.3 hours on an average, but seven (10.0%) had incomplete remission. For these seven, after improvements of their clinical symptoms, elective radical operations were performed within three weeks. In 22 (31.4%) patients whose symptoms were not improved at all with the epidural block, emergency exploratory celiotomies were performed, 15.4 hours on an average after the initial epidural block. Indications for surgical intervention of intestinal obstruction were decided by the absent movement of gas in the bowel in a series of plain X-rays. The effectiveness of the epidural block on the motility of the obstructed intestinal loop was experimentally confirmed in monkeys. We suggested that the epidural block, accompanied with usual conservative treatments, be recommended as the initial treatment for intestinal obstruction. PMID- 15278813 TI - Differences in the assessment of postoperative pain when evaluated by patients and doctors. AB - This study was undertaken to compare the assessment of pain intensity by 59 patients and by their doctors according to a visual analogue scale (VAS) at rest and when coughing at 5 and 20 hr after major abdominal surgery. The rating given by the patients, who received epidural analgesia to relieve postoperative pain, was significantly above, and moreover, significantly correlated with that given by the doctors at any time or under any condition of the assessment. However, the correlation between the ratings given by patients and doctors at rest at 5 hr after surgery was low (r = 0.39, rs = 0.38) and significantly different from that when coughing at 20 hr after the operation (r = 0.79, rs = 0.80). Our findings indicate that the assessment of postoperative pain may be associated with some unreliability, especially during early periods, when using the subjective or objective-rated VAS at rest separately, and thus requires the combined use or the concomitant use of the VAS when coughing. Substitutional use of the objective rated VAS for the subjective-rated VAS is not advised. PMID- 15278812 TI - Metabolic changes associated with malnutrition in the patients with multiple organ failure. AB - To clarify the metabolic changes associated with malnutrition in the patients with multiple organ failure (MOF), we measured energy expenditure, nitrogen excretion, nonprotein respiratory quotient (NPRQ), caloric intake, and cumulative caloric balance (CCB) in 20 MOF patients (12 survivors and 8 non-survivors). The non-survivors exhibited significantly greater cumulative caloric deficit than the survivors. Metabolic activity tended to decline to normal in the survivors as organ failures were overcome. In the non-survivors, on the contrary, regardless of large caloric deficit hypermetabolism persisted and characteristically followed by the sudden decrease in metabolic activity at the time immediately prior to death. Compared to the survivors, the non-survivors generally exhibited poorer response in metabolic activity and greater NPRQ change to the altered amount of caloric intake. It seemed that protein sparing effect by increased caloric intake was preserved in both the survivors and the non-survivors only with CCB above -5 times basal energy expenditure. These results suggest that persistent hypermetabolism and poor metabolic response to nutritional support are partly responsible for existing organ failures and poor outcome in MOF patients. PMID- 15278814 TI - Does a lidocaine patch reduce the pain at venous cannulation in adults? AB - In this study we evaluated whether a lidocaine patch reduces the pain relating to a venous cannulation in adults. The patch is consisted of the base containing 50% lidocaine on a thin polyester membrane. Its surface area is 15 cm2. Twenty-six adult patients scheduled for elective surgery (11 males and 15 females) were randomly divided into two groups according to application periods: Group A for 15 min and Group B for 30 min. Either the dorsal part of the hand or the radial side of the wrist was chosen and covered with the patch. Pain assessment was made by patients using a 0-100 point visual analog scale (VAS). In 7 patients of Group A, plasma lidocaine levels were measured 15 min after application by homogeneous enzyme immunoassay. The levels were further measured 30 and 60 min after application in 3 of those patients. The mean VAS score was 28.4 +/- 13.1 (mean +/ SD) for Group A and 51.8 +/- 15.9 for Group B, and the difference was statistically significant (P < 0.05). Plasma lidocaine levels were always below 0.2 microg.ml(-1). The results indicate that the skin was partially anesthetized by the lidocaine patch. A lidocaine patch may be useful and safely applicable for venous cannulation in adult patients. PMID- 15278815 TI - Does preoperative oral clonidine inhibit salivary secretion during general anesthesia? AB - Clonidine is known to inhibit salivary secretions and cause dryness of the mouth. We evaluated the effect of preoperative oral clonidine on salivary secretions before and during general anesthesia. Twenty-eight adult patients, equally divided into four groups, received the following premedication 2 hr prior to induction of anesthesia. Group 1 patients received oral ranitidine 5 mg.kg(-1) alone. Groups 2 and 3 patients received oral clonidine 1 microg.kg(-1) and 3 microg.kg(-1) respectively with oral ranitidine 5 mg.kg(-1). Group 4 patients received no premedication and served as control. The volume of salivary secretions was determined by calculating the change in weight of four cotton wool cylinders placed in the oral space 10 min before and 30, 60 and 120 min after induction of anesthesia. Salivary volume was significantly less in the clonidine treatment groups before induction of anesthesia. After induction of anesthesia, there were no significant differences in salivary secretions among the four groups. No severe hypotension or bradycardia was seen in any patient of four groups. Preoperative oral ranitidine 5 mg.kg(-1) had no effect on salivary secretion. In conclusion, clonidine did not decrease salivary secretions further over the already decreased level during general anesthesia. PMID- 15278816 TI - Changes in venous capacitance during prostaglandin E1-induced hypotension; comparisons with trinitroglycerin. AB - The purpose of this study was to examine the effects of prostaglandin E1 (PGE1) on venous capacitance during controlled hypotension. Trinitroglycerin (TNG) was used as a control agent. In rats anesthetized with ketamine, mean arterial pressure was lowered to 70 mmHg and subsequently 50 mmHg by intravenous infusion of PGE1 or TNG. Venous capacitance was assessed before and during induced hypotension by measuring the mean circulatory filling pressure (MCFP). MCFP was measured after briefly arresting the circulation by inflating an indwelling balloon in the right atrium. MCFP was significantly decreased by PGE1 from 7.9 +/ 0.3 to 6.9 +/- 0.3 mmHg at mean arterial pressure of 70 mmHg and to 6.9 +/- 0.2 mmHg at mean arterial pressure of 50 mmHg. The decrease in MCFP by PGE1 at mean arterial pressure of 70 mmHg was not significantly different from TNG. However, the decrease in MCFP by PGE1 at mean arterial pressure of 50 mmHg was significantly less than that by TNG. The results suggest that the venous capacitance may be increased by PGE1 to a similar degree with TNG at doses to produce a comparable level of moderate hypotension, but the increase in venous capacitance may be less in PGE1 than TNG at doses to produce deep hypotension. PMID- 15278817 TI - Effects of continuous negative extrathoracic pressure ventilation on left ventricular dimensions and hemodynamics in dogs. AB - It has been reported that continuous negative extrathoracic pressure ventilation (CNETPV) depresses cardiac output less than continuous positive pressure ventilation (CPPV) does, and this difference may be related to the different effects of two ventilatory modes on preload. We performed simultaneous measurements of hemodynamics and left ventricular short axis dimensions by transesophageal echocardiography (TEE) to evaluate left ventricular preload and function during CNETPV and CPPV in normal dogs. Hemodynamic measurements and simultaneous TEE recording were performed at 5 successive periods; 1) the first control period of intermittent positive pressure ventilation (IPPV1), 2) CNETPV with negative end-expiratory pressure (NEEP) of -10 cmH2O (CNET10), 3) CNETPV with NEEP of -15 cmH2O (CNET15), 4) the second control period of IPPV (IPPV2), and 5) CPPV with PEEP of 15 cmH2O (CPPV15). Left ventricular end-systolic and end diastolic dimension (LVESD and LVEDD), ejection fraction (EF) and fractional shortening (FS) were measured from TEE recordings. Both CNET10 and CNET15 induced no significant changes in hemodynamics and left ventricular dimensions, compared with those during IPPV1. However, CPPV15 reduced cardiac output and stroke volume (SV) and increased heart rate significantly, compared with IPPV2. CPPV15 significantly decreased LVEDD compared with IPPV2. Neither EF nor FS showed any significant change throughout the experiment. These results indicate that CNETPV preserved cardiac output because it maintained the preload and the left ventricular function. PMID- 15278818 TI - Changes in circulating blood volume following isoflurane or sevoflurane anesthesia. AB - Changes of circulating blood volume (CB volume) measured by the dual indicator dilution method were observed in 33 chronically instrumented mongrel dogs following either alpha-chloralose-urethane (C group), additive isoflurane (I group) or sevoflurane anesthesia (S group). These anesthetic groups were each divided into two subgroups with regard to respiratory care, namely Cp, Ip and Sp for those with intermittent positive pressure ventilation (six animals per subgroups), and Cs, Is and Ss for those with spontaneous breathing (five animals per subgroups). The CB volume under positive pressure ventilation remained unchanged in the Ip and Sp groups at both 0.5 and 1.0 MAC, and in the Cp group. The CB volume remained essentially unchanged in the Cs and Is groups at both 0.5 or 1.0 MAC, but the plasma volume tended to increase slightly in the Is group at 1.0 MAC. In the Ss group under spontaneous breathing, however, the CB volume increased from 84.4 +/- 7.0 to 91.4 +/- 7.7 at 0.5 MAC, and to 91.4 +/- 10.2 ml.kg(-1) at 1.0 MAC (0.01 < P < 0.05). These increases were caused by an increase in the plasma volume. The above data suggests that a concomitant increase in the venous pressure associated with an increase in the intrathoracic pressure produced by positive pressure ventilation would attenuate changes in the CB volume during sevoflurane anesthesia. PMID- 15278819 TI - 31P and 23Na nuclear magnetic resonance study on forebrain ischemia in rats with shift reagent Dy(TTHA). AB - 31P and 23Na nuclear magnetic resonance (NMR) spectroscopy was employed to study the dynamic changes in intracellular high-energy phosphates and sodium during 15 min of forebrain ischemia and recirculation in in vivo rat brain. In the presence of the shift reagent Dysprosium triethylenetetramine-N,N,N",N",N"',N"'-hexaacetic and [Dy(TTHA)], the sodium peak separated into two peaks, unshifted and shifted. During 15 min of ischemia, the unshifted sodium peak decreased and the shifted sodium peak increased. With recirculation, the unshifted and the shifted sodium peaks returned to the preischemia level within 10 min, but the shifted one increased during 30-60 min. Intracellular high-energy phosphates and intracellular pH (pHi) decreased during 15 min of ischemia and returned to the preischemia levels within 20 min of recirculation. We conclude that the decrease in unshifted sodium peak during ischemia is due to the decrease in subarachnoid sodium and the cellular influx of interstitial sodium would be minimum. The increase in shifted sodium peak during ischemia is considered to be due to the dilatation of cerebral blood vessels and the increase in interstitial sodium which was transported from subarachnoid space. PMID- 15278820 TI - Elevation of the extracellular glutamate concentration in the hippocampus after total cerebral ischemia related to the deterioration of the recovery in EEG and evoked potentials in dogs. AB - The concentrations of extracellular glutamate (Glu), aspartate (Asp) and glycine (Gly) were measured by microdialysis method in the cortex and hippocampus before, during and after 15 min of total cerebral ischemia in dogs. The correlations between the concentrations of amino acids and the changes in EEG and evoked potentials (EP) after ischemia were evaluated. Total cerebral ischemia was achieved by occluding the ascending aorta and the caval veins. The concentrations of Glu in the hippocampus significantly increased from 1.73 +/- 0.59 (mean +/- SEM) nmol.ml(-1) at pre-ischemia to 5.46 +/- 1.34 (P < 0.05) during ischemia and 14.37 +/- 3.70 (P < 0.01) 0-15 min after ischemia, and returned to the pre ischemic level 30 min after ischemia. The concentration of hippocampal Glu 0-15 min after ischemia had significant negative correlations with the EEG-EP scores (0 = serious deterioration of electrical function and 6 = normal electrical function) 30 min, 3 hr and 5 hr after ischemia (r = -0.69, P < 0.05 : r = -0.67, P < 0.05 : r = -0.70, P < 0.05, respectively). The increase of the extracellular Glu concentration in the hippocampus immediately after ischemia may aggravate the neurological outcome after total cerebral ischemia. PMID- 15278821 TI - Isosorbide dinitrate attenuated coronary artery spasm during general anesthesia for non-cardiac surgery. PMID- 15278822 TI - A case with intractable pain suffering from pancoast syndrome. PMID- 15278823 TI - Cardiopulmonary bypass and plasma taurine. PMID- 15278824 TI - Burning leg pain during spinal anesthesia in a diabetic patient. PMID- 15278825 TI - Usefulness of laryngeal mask airway in tracheostomy. PMID- 15278826 TI - Prostaglandin E1 increased cardiac contractility in cardiac arrest during open heart surgery. PMID- 15278827 TI - An increase in plasma concentrations of granulocyte elastase during and after bench surgery of the liver. PMID- 15278828 TI - Anesthetic management for liver transplantation from living donor to adult recipient--a case report. PMID- 15278829 TI - Recurrence of asthma after removal of adrenaline secreting pheochromocytoma. PMID- 15278830 TI - Respiratory rate change during balloon valvuloplasty. PMID- 15278832 TI - Two cases of bronchospasm during operation in a prone position. PMID- 15278831 TI - Two cases of postintubation subglottic granuloma with dyspnea. PMID- 15278833 TI - Sleeping sites of Rhinopithecus bieti at Mt. Fuhe, Yunnan. AB - Data on sleeping site selection were collected for a group of black-and-white snub-nosed monkeys (Rhinopithecus bieti; around 80) at Mt. Fuhe, Yunnan, China (99 degrees 20'E, 26 degrees 25'N, about 3,000 m asl) from November 2000 to January 2002. At the site mainly three vegetation types were present in an elevation-ascending order: deciduous broad leaf forest, mixed coniferous and broad leaf forest, and dark coniferous forest. In addition, bamboo forest presented in areas burned in 1958. Sleeping sites (n =10) were located in the coniferous forest, where trees were the tallest, bottommost branches were the highest, the diameter of crowns was the second largest, and the gradient of the ground was the steepest. Monkeys usually kept quiet during entering and staying at a sleeping site. The site choice and the quietness may be tactics to avoid potential predators. In the coniferous forest, however, monkeys did not sleep in the valley bottom where trees were the largest, but frequently slept in the middle of the slope towards the east/southeast, in the shadow of ridges in three other directions, to avoid strong wind and to access sunshine; in winter-spring, they ranged in a more southern and lower area than in summer-autumn. These may be behavioral strategies to minimize energy stress in the cold habitat. Monkeys often slept in the same sleeping site on consecutive nights, which reflected a reduced pressure of predation probably due to either the effectiveness of anti predation through sleeping site selection, or the population decline of predators with increasing human activities in the habitat. The group's behavioral responses to interactive and sometimes conflicting traits of the habitat are site-specific and conform to expectations for a temperate zone primate. PMID- 15278837 TI - A rheological network model for the continuum anisotropic and viscoelastic behavior of soft tissue. AB - The mechanical behavior of soft tissue demonstrates a number of complex features including nonlinearity, anisotropy, viscoelasticity, and growth. Characteristic features of the time-dependent and anisotropic behavior are related to the properties of various components of the tissue such as fibrous collagen and elastin networks, large proteins and sugars attached to these networks, and interstitial fluid. Attempts to model the elastic behavior of these tissues based on assumptions about the behavior of the underlying constituents have been reasonably successful, but the essential addition of viscoelasticity to these models has been met with varying success. Here, a new rheological network model is proposed using, as its basis, an orthotropic hyperelastic constitutive model for fibrous tissue and a viscoelastic reptation model for soft materials. The resulting model has been incorporated into numerical and computational models, and is shown to capture the mechanical behavior of soft tissue in various modes of deformation including uniaxial and biaxial tension and simple shear. PMID- 15278838 TI - Adaptive site selection rules and variation in group size of barn swallows: individual decisions predict population patterns. AB - Variation in group size is ubiquitous among socially breeding organisms. An alternative to the traditional examination of average reproductive success in groups of different sizes is to examine individual decision making by determining the cues used for site selection. Once factors used for decision making are known, one can determine whether group-level patterns, such as group size variation, are emergent properties of individual-level decision rules. The advantage of this alternative approach is that it can explain the distribution of group sizes rather than just the occurrence of optimal group sizes. Using barn swallows, I tested, but did not support, the hypothesis that individuals settle at sites based on the previous success of conspecifics (i.e., performance-based conspecific attraction). Instead, I demonstrate that an adaptive site selection decision rule--to breed where it is possible to reuse previously constructed nests--predicts 83% of the variation in the number of breeding pairs at a site. Furthermore, experimental nest removals demonstrated that settlement decisions are also strongly influenced by site familiarity. I discuss the interaction of the cue-based site selection rule with the occurrence of site fidelity and how, more generally, a consideration of individual-level decision rules can improve our understanding of variation in many social behaviors. PMID- 15278839 TI - The kin composition of social groups: trading group size for degree of altruism. AB - Why some social systems form groups composed of kin, while others do not, has gone largely untreated in the literature. Using an individual-based simulation model, we explore the demographic consequences of making kinship a criterion in group formation. We find that systems where social groups consist of one generation breeding associations may face a serious trade-off between degree of altruism and group size that is largely mediated by their kin composition. On the one hand, restricting groups to close kin allows the evolution of highly altruistic behaviors but may limit group size to suboptimal levels, the more severely so the smaller the intrinsic fecundity of the species and the stricter the kin admission rule. Group size requirements, on the other hand, can be met by admitting nonkin into groups, but not without limiting the degree of altruism that can evolve. As a solution to this conundrum, we show that if helping roles within groups are assigned through a lottery rather than being genetically determined, maximum degrees of altruism can evolve in groups of nonrelatives of any size. Such a "lottery" mechanism may explain reproductive and helping patterns in organisms as varied as the cellular slime molds, pleometrotic ants, and Galapagos hawks. PMID- 15278840 TI - Reconstructing the origin of Helianthus deserticola: survival and selection on the desert floor. AB - The diploid hybrid species Helianthus deserticola inhabits the desert floor, an extreme environment relative to its parental species Helianthus annuus and Helianthus petiolaris. Adaptation to the desert floor may have occurred via selection acting on transgressive, or extreme, traits in early hybrids between the parental species. We explored this possibility through a field experiment in the hybrid species' native habitat using H. deserticola, H. annuus, H. petiolaris, and two populations of early-generation (BC(2)) hybrids between the parental species, which served as proxies for the ancestral genotype of the ancient hybrid species. Character expression was evaluated for each genotypic class. Helianthus deserticola was negatively transgressive for stem diameter, leaf area, and flowering date, and the latter two traits are likely to be advantageous in a desert environment. The BC(2) hybrids contained a range of variation that overlapped these transgressive trait means, and an analysis of phenotypic selection revealed that some of the selective pressures on leaf size and flowering date, but not stem diameter, would move the BC(2) population toward the H. deserticola phenotype. Thus, H. deserticola may have originated from habitat-mediated directional selection acting on hybrids between H. annuus and H. petiolaris in a desert environment. PMID- 15278841 TI - Stoichiometrically explicit competition between grazers: species replacement, coexistence, and priority effects along resource supply gradients. AB - Assuming key trade-offs among interactors, several models (resource ratio, keystone predation, intraguild predation) predict changes in species composition over resource supply gradients. Ecological stoichiometry could also predict compositional shifts of grazers over gradients of nutrient and light supply through a mechanism involving (mis)matches between elemental body composition of grazers and plants. This hypothesis is explored here using a suite of two-grazer, one-plant models that incorporate three key components: plant production depends on light and nutrients, nutrient content of plants can vary, and homeostatic grazers can be carbon or nutrient limited. The results from this suite closely resemble the classical resource ratio model describing plant competition for two resources. Here, the models predict shifts of grazer composition along resource supply gradients if species trade off competitive abilities for plant carbon and nutrients. Given this trade-off, superior nutrient competitors should dominate low nutrient environments, and superior carbon competitors should dominate high nutrient environments. At intermediate nutrient supply, species can coexist at a stable equilibrium, or alternative stable states emerge, depending on how grazers impact their resources. These results depend on food web architecture, however. For instance, predators can alter or reduce possibilities for stoichiometry mediated coexistence of grazers. PMID- 15278842 TI - The evolution of female preferences for multiple indicators of quality. AB - In a variety of species, females exhibit preferences for multiple male ornaments. Several hypotheses have been proposed to explain this phenomenon. Which, if any, of these hypotheses is the most plausible in general remains largely unresolved based on the available empirical data. Yet theoretical studies conclude that the evolution of preferences for multiple signals of male quality is unlikely, especially when the use of an additional cue in mate choice strongly increases the overall cost of choice. This would imply that most male courtship characters do not reflect the male's genetic quality but instead evolved through Fisherian sexual selection. However, the existing models focus on ornaments that signal overall genetic quality and do not address the possibility that different ornaments provide information about different aspects of quality. Therefore, we develop a model in which the ornaments act as signals for distinct quality components. When the ornaments provide overlapping information about these quality components, we retrieve the results of earlier models. However, when the ornaments provide independent information, preferences for multiple ornaments may evolve, even when exhibiting multiple preferences is costly. We discuss our results in relation to the multiple-message and redundant-signal hypotheses for ornament diversity and identify parallels between Fisherian and good-genes mechanisms for the evolution of multiple ornaments. PMID- 15278843 TI - Temporal variation can facilitate niche evolution in harsh sink environments. AB - We examine the impact of temporal variation on adaptive evolution in "sink" environments, where a species encounters conditions outside its niche. Sink populations persist because of recurrent immigration from sources. Prior studies have highlighted the importance of demographic constraints on adaptive evolution in sinks and revealed that adaptation is less likely in harsher sinks. We examine two complementary models of population and evolutionary dynamics in sinks: a continuous-state quantitative-genetics model and an individual-based model. In the former, genetic variance is fixed; in the latter, genetic variance varies because of mutation, drift, and sampling. In both models, a population in a constant harsh sink environment can exist in alternative states: local maladaptation (phenotype comparable to immigrants from the source) or adaptation (phenotype near the local optimum). Temporal variation permits transitions between these states. We show that moderate amounts of temporal variation can facilitate adaptive evolution in sinks, permitting niche evolution, particularly for slow or autocorrelated variation. Such patterns of temporal variation may particularly pertain to sinks caused by biotic interactions (e.g., predation). Our results are relevant to the evolutionary dynamics of species' ranges, the fate of exotic invasive species, and the evolutionary emergence of infectious diseases into novel hosts. PMID- 15278844 TI - The influence of ecology and genetics on behavioral variation in salamander populations across the Eastern Continental Divide. AB - Understanding the unique contributions of ecology and history to the distribution of species within communities requires an integrative approach. The Eastern Continental Divide in southwestern Virginia separates river drainages that differ in species composition: the more aquatic, predatory Desmognathus quadramaculatus is present only in the New River drainage (which drains to the Gulf of Mexico), while Desmognathus monticola is present in both the New River drainage and the James River drainage (which drains to the Atlantic Ocean). We investigated natural distributions, behavioral variation in experimental mesocosms, population genetic, and phylogenetic implications of community structure. The presence of D. quadramaculatus increased the terrestriality of D. monticola in natural and experimental situations but to different degrees in allopatric and sympatric populations. Our ecological data suggest that the degree of terrestriality in D. monticola is a result of a balance between the optimal aquatic habitat and risks of predation. Our genetic analyses suggest that D. monticola has experienced a recent range expansion and has only a recent history of association with D. quadramaculatus in Virginia. This is surprising given the strong behavioral variation that exists in populations experiencing unique community compositions over a scale of meters. This study demonstrates the need to combine both ecology and genetics toward an understanding of the factors affecting species distributions, behavioral variation between populations, and patterns of genetic variation across a landscape. PMID- 15278845 TI - The robustness of Hamilton's rule with inbreeding and dominance: kin selection and fixation probabilities under partial sib mating. AB - Assessing the validity of Hamilton's rule when there is both inbreeding and dominance remains difficult. In this article, we provide a general method based on the direct fitness formalism to address this question. We then apply it to the question of the evolution of altruism among diploid full sibs and among haplodiploid sisters under inbreeding resulting from partial sib mating. In both cases, we find that the allele coding for altruism always increases in frequency if a condition of the form rb>c holds, where r depends on the rate of sib mating alpha but not on the frequency of the allele, its phenotypic effects, or the dominance of these effects. In both examples, we derive expressions for the probability of fixation of an allele coding for altruism; comparing these expressions with simulation results allows us to test various approximations often made in kin selection models (weak selection, large population size, large fecundity). Increasing alpha increases the probability of fixation of recessive altruistic alleles (h<1/2), while it can increase or decrease the probability of fixation of dominant altruistic alleles (h>1/2). PMID- 15278846 TI - Linking optimal foraging behavior to bird community structure in an urban-desert landscape: field experiments with artificial food patches. AB - Urban bird communities exhibit high population densities and low species diversity, yet mechanisms behind these patterns remain largely untested. We present results from experimental studies of behavioral mechanisms underlying these patterns and provide a test of foraging theory applied to urban bird communities. We measured foraging decisions at artificial food patches to assess how urban habitats differ from wildlands in predation risk, missed-opportunity cost, competition, and metabolic cost. By manipulating seed trays, we compared leftover seed (giving-up density) in urban and desert habitats in Arizona. Deserts exhibited higher predation risk than urban habitats. Only desert birds quit patches earlier when increasing the missed-opportunity cost. House finches and house sparrows coexist by trading off travel cost against foraging efficiency. In exclusion experiments, urban doves were more efficient foragers than passerines. Providing water decreased digestive costs only in the desert. At the population level, reduced predation and higher resource abundance drive the increased densities in cities. At the community level, the decline in diversity may involve exclusion of native species by highly efficient urban specialists. Competitive interactions play significant roles in structuring urban bird communities. Our results indicate the importance and potential of mechanistic approaches for future urban bird community studies. PMID- 15278847 TI - Convergence, divergence, and homogenization in the ecological structure of emydid turtle communities: the effects of phylogeny and dispersal. AB - Studies that have explored the origins of patterns of community structure from a phylogenetic perspective have generally found either convergence (similarity) in community structure between regions through adaptive evolution or lack of convergence (dissimilarity) due to phylogenetic conservatism in the divergent ecological characteristics of lineages inhabiting different regions. We used a phylogenetic approach to document a third pattern in the structure of emydid turtle communities. Emydid communities in southeastern North America tend to have a higher proportion of aquatic species than those in the northeast. This pattern reflects phylogenetic conservatism in the ecology and biogeography of two basal emydid clades, limiting convergence in community structure between these regions. However, differences in community structure between northeastern and southeastern North America have also been homogenized considerably by the dispersal of species with phylogenetically conserved ecological characteristics between regions. This pattern of ecologically conservative dispersal may be important in many continental and oceanic systems. PMID- 15278848 TI - Large species shifts triggered by small forces. AB - Changes in species composition of communities seem to proceed gradually at first sight, but remarkably rapid shifts are known to occur. Although disrupting disturbances seem an obvious explanation for such shifts, evidence for large disturbances is not always apparent. Here we show that complex communities tend to move through occasional catastrophic shifts in response to gradual environmental change or evolution. This tendency is caused by multiple attractors that may exist in such systems. We show that alternative attractors arise robustly in randomly generated multispecies models, especially if competition is symmetrical and if interspecific competition is allowed to exceed intraspecific competition. Inclusion of predators as a second trophic level did not alter the results greatly, although it reduced the probability of alternative attractors somewhat. These results suggest that alternative attractors may commonly arise from interactions between large numbers of species. Consequently, the response of complex communities to environmental change is expected to be characterized by hysteresis and sudden shifts. Some unexplained regime shifts observed in ecosystems could be related to alternative attractors arising from complex species interactions. Additionally, our results support the idea that ancient mass extinctions may partly be due to an intrinsic loss of stability of species configurations. PMID- 15278849 TI - Measles metapopulation dynamics: a gravity model for epidemiological coupling and dynamics. AB - Infectious diseases provide a particularly clear illustration of the spatiotemporal underpinnings of consumer-resource dynamics. The paradigm is provided by extremely contagious, acute, immunizing childhood infections. Partially synchronized, unstable oscillations are punctuated by local extinctions. This, in turn, can result in spatial differentiation in the timing of epidemics and, depending on the nature of spatial contagion, may result in traveling waves. Measles epidemics are one of a few systems documented well enough to reveal all of these properties and how they are affected by spatiotemporal variations in population structure and demography. On the basis of a gravity coupling model and a time series susceptible-infected-recovered (TSIR) model for local dynamics, we propose a metapopulation model for regional measles dynamics. The model can capture all the major spatiotemporal properties in prevaccination epidemics of measles in England and Wales. PMID- 15278850 TI - Distraction sneakers decrease the expected level of aggression within groups: a game-theoretic model. AB - Hawk-dove games have been extensively used to predict the conditions under which group-living animals should defend their resources against potential usurpers. Typically, game-theoretic models on aggression consider that resource defense may entail energetic and injury costs. However, intruders may also take advantage of owners who are busy fighting to sneak access to unguarded resources, imposing thereby an additional cost on the use of the escalated hawk strategy. In this article we modify the two-strategy hawk-dove game into a three-strategy hawk-dove sneaker game that incorporates a distraction-sneaking tactic, allowing us to explore its consequences on the expected level of aggression within groups. Our model predicts a lower proportion of hawks and hence lower frequencies of aggressive interactions within groups than do previous two-strategy hawk-dove games. The extent to which distraction sneakers decrease the frequency of aggression within groups, however, depends on whether they search only for opportunities to join resources uncovered by other group members or for both unchallenged resources and opportunities to usurp. PMID- 15278851 TI - Markov chain analysis of succession in a rocky subtidal community. AB - We present a Markov chain model of succession in a rocky subtidal community based on a long-term (1986-1994) study of subtidal invertebrates (14 species) at Ammen Rock Pinnacle in the Gulf of Maine. The model describes successional processes (disturbance, colonization, species persistence, and replacement), the equilibrium (stationary) community, and the rate of convergence. We described successional dynamics by species turnover rates, recurrence times, and the entropy of the transition matrix. We used perturbation analysis to quantify the response of diversity to successional rates and species removals. The equilibrium community was dominated by an encrusting sponge (Hymedesmia) and a bryozoan (Crisia eburnea). The equilibrium structure explained 98% of the variance in observed species frequencies. Dominant species have low probabilities of disturbance and high rates of colonization and persistence. On average, species turn over every 3.4 years. Recurrence times varied among species (7-268 years); rare species had the longest recurrence times. The community converged to equilibrium quickly (9.5 years), as measured by Dobrushin's coefficient of ergodicity. The largest changes in evenness would result from removal of the dominant sponge Hymedesmia. Subdominant species appear to increase evenness by slowing the dominance of Hymedesmia. Comparison of the subtidal community with intertidal and coral reef communities revealed that disturbance rates are an order of magnitude higher in coral reef than in rocky intertidal and subtidal communities. Colonization rates and turnover times, however, are lowest and longest in coral reefs, highest and shortest in intertidal communities, and intermediate in subtidal communities. PMID- 15278852 TI - Closing the gap between academic surgery and community practice. PMID- 15278853 TI - Facial computed tomography use in trauma patients who require a head computed tomogram. AB - PURPOSE: Head-injured patients admitted to a trauma center may or may not have associated facial fractures. Most head-injured patients undergo head computed tomography (CT) scan early in their evaluation. The question of adding a facial CT at the time of the head CT can be unclear. The aims of our study are 1) to analyze how the facial CT is used in conjunction with the head CT in facial fracture trauma patients, 2) to recognize unique identifiers that would aid the surgeon's decision-making process to order a facial CT in continuity with a head CT, and 3) to examine what is characteristic of head trauma patients who receive a facial CT separately, at some point after the head CT. Materials and methods Data were retrospectively reviewed for a 5-year period at a level I trauma center in which all patients who present with craniomaxillofacial trauma are managed by the oral and maxillofacial surgery (OMFS) service. Included patients must have obtained a head CT during initial resuscitation and be diagnosed with a facial fracture during the same hospital stay. These patients were divided into 3 groups: those who had a 1) head CT only, 2) head CT and facial CT at the same time, and 3) head CT with the facial CT performed at a later time. RESULTS: A total of 9,871 patients were admitted to the trauma service during a 5-year period and 4,926 patients (49.9%) had head CT performed. Of this group, 12% had facial fractures, and the most common associated injury in this group was facial lacerations and concussions. The nasal fracture followed by the orbital fracture was the most common fracture type. Eighty-four percent of the time, the facial CT was used to help diagnose facial fractures in this patient population. The 3 different groups showed unique trends. CONCLUSION: Six points were identified in our study that can augment the physical examination in patients who require head CT. The following points can help prompt the clinician to order a combination head and facial CT: 1) 12% of trauma patients who require a head CT will have a facial fracture, whereas half of these patients will have multiple facial fractures. 2) Orbital fractures are commonly missed in this group and often require a secondary scan such as coronal views for accurate diagnosis. 3) Facial lacerations correlate with ordering a combination head and facial CT. 4) The most common facial fracture identified among patients receiving a trauma head CT is the nasal fracture. 5) The use of the facial CT in more severely injured patients tended to be delayed and was related to increased hospital and intensive care unit days. 6) Only 16% of facial fracture patients who had received an initial trauma head CT did not require further facial CT scanning. PMID- 15278854 TI - Resorbable poly-L-lactide plates and screws for the treatment of mandibular condylar process fractures: a clinical and radiologic follow-up study. AB - PURPOSE: The purpose of this study was to determine whether a resorbable poly-l lactide (PLLA) miniplate system could be used to treat mandibular condylar process fracture. PATIENTS AND METHODS: Fourteen patients (12 males, 2 females, aged 23.1 +/- 5.7 years) who had mandibular condylar process fractures treated with PLLA implants were recalled for follow-up clinical and radiologic examinations at 3 years. RESULTS: Mouth opening recovered to more than 35 mm and occlusion was stable in all patients. There was no facial asymmetry 3 months postoperatively. Two patients had mild chronic postoperative tenderness at the implantation site; however, there was no wound infection. All fractured mandibular condyles showed anatomic good reduction and long-term stability with the use of resorbable miniplates and screws. Bone healing was satisfactory in all patients, and there was no evidence of abnormal resorption of the condylar process. The screw holes remained evident after 3 years. Screw holes in 2 patients showed enlargement on radiographic examination. CONCLUSION: The PLLA miniplate system provides reliable stability when used for the fixation of mandibular condylar process fractures. PMID- 15278855 TI - Psychologic functioning and needs of indigent patients with facial injury: a prospective controlled study. AB - PURPOSE: This study sought to examine 1) temporal changes in psychologic functioning over 12 months and 2) baseline differences in mental health and social service needs between orofacial injury patients and sociodemographically comparable controls undergoing elective oral surgery. MATERIALS AND METHODS: Prospective, case-control study of patients treated at a public hospital in Los Angeles, CA. A total of 336 subjects with mandible fractures and 119 subjects undergoing elective removal of their third molars participated in structured, repeated follow-up assessments (10 days, 6 months, and 12 months after their surgical procedures). Multiple imputation was used to manage incomplete data, and propensity score analysis was used to correct for covariate imbalances between the injury and the control cohort. A series of ANOVAs, chi(2) analyses, and odds ratios was conducted. RESULTS: The injury patients continue to experience significant psychologic distress for up to 12 months following the traumatic event. Orofacial injury patients also tend to report more lifetime and current mental health and social service needs than the sociodemographically similar elective-surgery cohort. CONCLUSION: The management of facial injuries in disadvantaged individuals should integrate case management that addresses psychosocial sequelae and service needs of patients. PMID- 15278856 TI - Clinical investigation of amelanotic malignant melanoma in the oral region. AB - PURPOSE: Amelanotic oral malignant melanoma (AOMM) is a rare tumor that is difficult to diagnose. We studied the clinical and pathologic features of nine cases of this tumor to define diagnostic criteria and estimate prognoses for 2 different types of AOMM. PATIENTS AND METHODS: Nine patients with 2 different types of primary AOMM were examined between 1970 and 2002. The histopathology of surgical specimens was studied, uncertain diagnoses were supported by immunohistochemical reactions, and electron microscopy and prognoses were reviewed retrospectively. RESULTS: AOMM without radial growth phase may be particularly difficult to diagnose correctly without immunohistochemical assistance. Tumors consisted of a mixture of polygonal and spindle cells in different ratios in tumors with and without radial growth phase. The life span ranged from 3 months to 6 years 3 months, and all 9 patients died of the tumor. In 7 of the 9 cases, distant metastases were found. CONCLUSIONS: AOMM without radial growth phase may be misdiagnosed as epulis or squamous cell carcinoma. Questionable lesions, particularly maxillary and palatal lesions, must be biopsied for histopathologic and possibly immunohistochemical examinations followed by rapid treatment. The prognosis of AOMM was poor. PMID- 15278857 TI - The correlation between mandible fractures and loss of consciousness. AB - PURPOSE: The purpose of the study was to evaluate the relationship between mandible fractures and loss of consciousness (LOC) in patients sustaining maxillofacial trauma in a level I trauma center. PATIENTS AND METHODS: The study was a retrospective chart review performed at Lincoln Medical and Mental Health Center, a level I trauma center. A total of 225 hospital charts of patients who sustained mandible fractures during the period from 1997 to 2001 were reviewed. The site of fracture, number of fractures per mandible, degree of displacement, and mechanism of injury were recorded. Percentages of each category were compared among patients with positive and negative LOC. Patients with any concomitant craniofacial fractures or injuries were excluded from the study. RESULTS: The incidence of LOC in patients sustaining mandible fractures was 17.6%. Young men were most likely to sustain mandibular fractures. The male-to-female ratio was 5.6:1. More than 70% of mandible fractures were due to interpersonal violence. Among various causes of interpersonal violence, use of a fist was most common. Overall, the number of fractures per patient, the incidence of singular versus multiple mandibular fractures, and the degree of displacement were less in the group with positive LOC. Body and angle regions were subjected to higher numbers of fractures, with slight variation among the 2 groups. The most common locations for mandibular fractures were the body (26% in negative LOC group; 42% in positive LOC group) and the angle (31% in negative LOC group; 26% in positive LOC group). CONCLUSION: Nearly 1 in 5 mandible fractures was associated with positive LOC. The patient group with a positive LOC, on average, sustained fewer fractures. It is possible that when the mandible sustained fewer fractures, the dissipation of energy was reduced and more force was transmitted to the cranial vault, thereby resulting in a higher incidence of LOC. PMID- 15278858 TI - Intraoral maxillary quadrangular Le Fort II osteotomy: a long-term follow-up study. AB - PURPOSE: This study evaluated the clinical outcome and skeletal stability of the intraoral maxillary quadrangular Le Fort II osteotomy (IQLO) with wire or rigid internal fixation following horizontal maxillary advancement. PATIENTS AND METHODS: All 21 patients who had undergone the IQLO were analyzed for operation time, blood loss, length of hospitalization, intraoperative and postoperative complications, and radiographic abnormalities. Lateral cephalometric radiographs were taken preoperatively (T1), postoperatively (T2) and late postoperatively (T3) to analyze skeletal movement. Two maxillary landmarks (posterior nasal spine [PNS] and A point) and 2 dental landmarks (the distobuccal cusp tip of the maxillary left second molar [2M] and the maxillary incisal tip [CI]) were used to determine horizontal and vertical changes for each time period. Student t test was used to evaluate early postoperative changes and late postoperative stability. In addition, 21 patients completed a questionnaire at the most recent follow-up visit regarding personal intentions, perceived outcome, and overall satisfaction. RESULTS: Twenty-one patients (9 females, 12 males) with an average age of 20.3 years diagnosed with horizontal maxillary-zygomatic deficiency underwent IQLO by 1 surgeon with an average follow-up of 6.3 years. Nine patients received mini-plate osseous segment fixation and 12 patients received wire osseous segment fixation. The mean time from surgery to the first postoperative radiograph (T2) was 4.4 weeks (range 1.0 to 6.7 weeks) and the mean time from surgery to the late postoperative radiograph (T3) was 6.2 years (range, 7.9 to 176.3 months). Statistical analysis of cephalometric landmarks revealed the following significant late postsurgical movements (T3-T2) for wire fixation: PNS moved 1.0 mm inferiorly (SD, 1.2), and 2M moved 1.5 mm inferiorly (SD, 2.2). The remaining cephalometric landmarks for rigid and wire fixation showed no statistically significant late postsurgical movement. Clinical outcome analysis revealed few complications, low surgical and postsurgical morbidity, and excellent patient satisfaction. CONCLUSION: The IQLO is a predictable procedure that exhibits long-term skeletal stability. Long-term retrospective review revealed low postsurgical morbidity and high patient satisfaction. PMID- 15278859 TI - The application of magnetic resonance imaging-guided fine-needle aspiration cytology in the diagnosis of deep lesions in the head and neck. AB - PURPOSE: Image-guided fine-needle aspiration cytology (FNAC) may be useful as an alternative diagnostic approach to lesions in the head and neck. This study reports on the use of magnetic resonance imaging (MRI)-guided FNAC for diagnostic evaluation of deep lesions in this region. MATERIALS AND METHODS: This was a prospective study of 12 patients with deep lesions in the head and neck who underwent MRI-guided FNAC at the Shanghai 9th People's Hospital. A 0.2-T open magnet was used for MRI and localization of the 20-gauge MRI-compatible needle. All of the aspirated samples were stained with hematoxylin and eosin and examined by a cytopathologist. RESULTS: The needle in all 12 cases was displayed on MRI in the central portion of the lesion under the guidance of MRI; 12 of 12 patients (100%) had diagnostic aspirations and none needed open biopsy for more specific histologic interpretation. Six of these 12 patients with tumors (4 malignant, 2 benign) underwent operative treatment with positive postoperative pathologic results. One patient had a diagnosis of inflammation. The diagnostic accuracy was 91.67% (11 of 12), the sensitivity was 85.71% (6 of 7), and the specificity was 100% (5 of 5). There were no false-positive results and 1 false-negative result, for a false-negative rate of 14.29% (1 of 7). All aspiration procedures were well tolerated and without complications. CONCLUSIONS: MRI-guided FNAC is a cost effective tool for establishing tissue diagnosis as a primary investigative modality. It is helpful and accurate in the diagnosis of deep lesions in the head and neck and in follow-up of patients, thereby avoiding further surgical intervention. PMID- 15278860 TI - Early repair of avulsive facial wounds secondary to trauma using interpolation flaps. AB - PURPOSE: This article describes the use of various local interpolation flaps for the reconstruction of facial defects resulting from trauma. PATIENTS AND METHODS: Fifteen interpolation flaps were used to restore missing tissue of 14 patients who sustained trauma. Two of the procedures were performed as a single stage, whereas 13 flaps required a separate surgery to "take down" the pedicle. All flap procedures were performed within 72 hours from the time of trauma. Four types of interpolation flaps were used. RESULTS: All flaps healed without evidence of infection, dehiscence, or necrosis. Two patients required secondary treatment 6 months after the flap procedure. CONCLUSION: Various local interpolation flaps provide a reliable and aesthetic treatment option for early repair of soft tissue defects secondary to trauma. PMID- 15278861 TI - Management of descending necrotizing mediastinitis. AB - PURPOSE: One of the most dreaded and the most lethal form of mediastinitis is descending necrotizing mediastinitis (DNM). PATIENTS AND METHODS: Between January 1990 and June 2001, 6 patients (mean age, 54.5 years; age range, 19 to 72 years) with DNM were treated in the Department of Thoracic Surgery of General Hospital of Attica "K.A.T." The primary etiology was odontogenic abscess in 3 patients and peritonsillar abscess in the other 3. Diagnosis was confirmed by computed tomography of the neck and chest. All patients underwent surgical drainage of the involved cervical region and mediastinum by monolateral cervicotomy and left thoracotomy. RESULTS: The delay between the occurrence of thoracic symptoms and mediastinal drainage varied from 1 to 4 days. The thoracic approach and the side of the thoracotomy depended on the involved mediastinal compartments and side of pleural effusion. The duration of mediastinal drainage varied from 8 to 22 days (mean, 12.5 days). One patient died of multiorgan failure related to postoperative septic shock. CONCLUSION: Delayed diagnosis and inadequate drainage are the main causes of the high mortality rate of DNM. Routine use of the computed tomography scan is highly recommended in patients with a deep cervical infection for early detection of mediastinitis at a time when the chest roentgenogram is still normal. If one realistically hopes to avoid the high mortality rate, aggressive surgical drainage and debridement of the neck and drainage of the mediastinum via a posterolateral thoracotomy by a multidisciplinary team of surgeons are required. PMID- 15278862 TI - Occlusal caries experience in patients with asymptomatic third molars. AB - PURPOSE: Our goals were to determine the prevalence of caries experience, carious lesions, or restorations on the occlusal surface, in asymptomatic third molars erupted to the occlusal plane, and to examine the association between caries experience in other molars and third molars within the same mouth and quadrant. PATIENTS AND METHODS: Clinical data assessing oral health were collected from healthy patients (ASA I, II). The presence or absence of caries experience on the occlusal surface of third molars and on any surface of the first and second molars was recorded during clinical and radiographic examinations. The occurrence of caries experience for younger and older subjects was compared using the general association Cochran-Mantel-Haenszel statistic and the association of occurrence in the maxilla and mandible by the McNemar test. The association between caries experience in a third molar and caries experience in first and second molars also was assessed. RESULTS: Overall, 28% of the 303 patients with at least 1 third molar at the occlusal plane were affected by third molar caries. Patients 25 years or older had more caries experience in a third molar than those younger than 25 years, 39% versus 11% (P <.0001). Mandibular third molars were affected more often than maxillary third molars, 24% versus 18% (P <.0001). Nearly all patients, 76 of 80 (95%), with third molar caries experience also had caries experience in first/second molars, but only 80 of 223 (36%) of patients with first/second molar caries experience had a history of third molar caries. CONCLUSIONS: The prevalence of caries in third molars erupted to the occlusal plane in these young patients was high, but not unique to third molars, particularly in those 25 years of age and older. Although these results provide a baseline description of the association between caries experience in first/second molars and associated third molars, data are needed from longitudinal studies to determine the value of first/second molar caries experience in predicting the risk of caries in third molars. PMID- 15278863 TI - Ex vivo produced human conjunctiva and oral mucosa equivalents grown in a serum free culture system. AB - PURPOSE: We sought to develop full-thickness ex vivo produced human conjunctiva and oral mucosa equivalents using a serum-free culture system without a feeder layer and to compare conjunctiva and oral mucosa equivalents to assess their suitability as graft materials for eyelid reconstruction. MATERIALS AND METHODS: Human conjunctival and oral mucosal keratinocytes were cultured, expanded, and seeded onto AlloDerm (LifeCell Corp, Branchburg, NJ), a cadaveric, acellular dermis, to produce ex vivo produced full-thickness mucosa equivalents. Histology of equivalents and their expression of immunoreactive Ki-67, a proliferation marker, and GLUT1, a membrane antigen seen in barrier tissues, were examined at 4, 11, and 18 days after seeding onto AlloDerm. RESULTS: Progressive epithelial stratification was observed on day 4, 11, and 18 conjunctiva and oral mucosa equivalents. Ki-67 immunoreactivity progressively increased with cultured time in both types of equivalent, indicating the continued presence of actively proliferating cells. GLUT1 immunoreactivity, concentrated in the basal keratinocytes of stratified epithelia of both types of equivalents, mimicked native tissue and indicated a high glycolytic state of the basal cells. CONCLUSIONS: Conjunctival and oral mucosal equivalents are similar to native tissue and demonstrate high proliferative and glycolytic states. Due to the similarity to conjunctiva, oral mucosal equivalents may be useful for eyelid reconstruction. Their advantages for surgical reconstruction include 1) ease of obtaining autogenous oral epithelium for expansion in vitro without the possibility of contaminating cellular- or serum-borne biologic agents, 2) growth of intact, confluent epithelia on rigid, transplantable human allogeneic dermis that may be surgically transplanted, and 3) reduced donor site morbidity and surgical time. PMID- 15278864 TI - A role for cyclooxygenase II inhibitors in modulating temporomandibular joint inflammation from a meal pattern analysis perspective. AB - PURPOSE: Developing a valid noninvasive animal model to study temporomandibular joint (TMJ) inflammation/pain has proved difficult. However, its has been recently demonstrated that meal pattern analysis, and in particular meal duration, can be used as a biologic marker for TMJ inflammation/pain induced by bilateral injections of complete Freund's adjuvant (CFA). The present study was undertaken to confirm previous findings and extend them by using rofecoxib (VIOXX; Merck and Co, West Point, PA), a selective cyclooxygenase-2 inhibitor (COX-2-I). MATERIALS AND METHODS: Forty-eight male rats were assigned to 1 of 4 groups: group 1, no CFA and no COX-2-I treatment; group 2, no CFA and treatment with the COX-2-I; group 3, bilateral TMJ CFA injection and no COX-2-I treatment; and group 4, CFA injection and treatment with the COX-2-I. Food intake was recorded by computer 24 hours before and for 48 hours after CFA injection. TMJ swelling, chromodacryorrhea, and meal patterns were quantified. RESULTS: CFA increased swelling (P <.05), chromodaccryorrhea (P <.05), meal duration at 24 and 48 hours, and TMJ retrodiscal tissue interleukin-1beta (P < 0.01) in group 3, but treatment with the COX-2-I attenuated these effects in group 4, (CFA + COX-2-I). CONCLUSIONS: These data confirm that meal pattern analysis, and in particular meal duration, is a noninvasive measure of TMJ inflammation/pain. However, this experiment has extended this model as a marker of drug treatment efficacy, specifically the efficacy of COX-2-I in treatment of orofacial inflammation/pain. PMID- 15278865 TI - Large lytic lesion of the ascending ramus, the condyle, and the infratemporal region. PMID- 15278866 TI - Bispectral analysis: an objective method of assessing anesthetic depth. PMID- 15278867 TI - Bispectral index monitor for conscious sedation: the case against its use. PMID- 15278868 TI - Chronic recurrent parotitis: a closer look at its origin, diagnosis, and management. PMID- 15278869 TI - Axillary nodal metastasis from oral and maxillofacial cancers: a report of 3 cases. PMID- 15278870 TI - Brown tumor of the mandible as first manifestation of atypical parathyroid adenoma. PMID- 15278871 TI - The silent sinus syndrome: report of 2 cases. PMID- 15278872 TI - Posttransplant lymphoproliferative disorder resembling a chronic orocutaneous infection in an immunosuppressed patient. PMID- 15278873 TI - Maxillary distraction osteogenesis in a patient with pycnodysostosis: a case report. PMID- 15278874 TI - Transoral osteosynthesis at the mandibular ramus and subcondyle using angular screwing instrument and biodegradable miniplate system. PMID- 15278875 TI - Early cellular alterations in bone after radiation therapy and its relation to osteoradionecrosis. PMID- 15278876 TI - Platelet-rich plasma: evidence to support its use. PMID- 15278895 TI - 2003 annual report: ESRD clinical performance measures project. PMID- 15278897 TI - Differential expression and activation of Stat3 during mouse embryo implantation and decidualization. AB - Signal transducer and activator of transcription (STATs) can be activated by many cytokines and growth factors. Stat3, a member of STAT family, is essential for embryonic development. Stat3 is specifically activated during mouse embryo implantation. This study was to investigate the expression, activation, and regulation of Stat3 in mouse uterus during early pregnancy, pseudopregnancy, delayed implantation, artificial decidualization, and hormonal treatments using in situ hybridization and immunohistochemistry. There was a strong level of Stat3 phosphorylation in the luminal epithelium only at the midnight of day 4 pregnancy, which coincides with attachment reaction between the blastocyst and luminal epithelium. However, there was no detectable Stat3 phosphorylation at the corresponding period during pseudopregnancy. On day 5 of pregnancy, Stat3 phosphorylation was strongly observed in the luminal epithelium and the stroma surrounding the implanting blastocyst at implantation sites, but not at the inter implantation sites. Stat3 phosphorylation was also not detected on day 5 of pseudopregnancy. Stat3 phosphorylation was at a high level in the decidual cells on days 6-8 of pregnancy. Under artificial decidualization, Stat3 was also phosphorylated in the decidual cells. In the ovariectomized mice, there was no Stat3 expression and activation in the uterus. Progesterone had no obvious effects. However, Stat3 mRNA expression and phosphorylation were significantly stimulated by estrogen treatment. Our data suggest that Stat3 phosphorylation may be important for mouse embryo implantation and decidualization, and may also be regulated by maternal estrogen. PMID- 15278898 TI - Timing of Plk1 and MPF activation during porcine oocyte maturation. AB - A Polo-like kinase 1 (Plk1) appears involved in an autocatalytic loop between CDC25C phosphatase and M phase promoting factor (MPF) in Xenopus oocytes and leads to activation of MPF that is required for germinal vesicle breakdown (GVBD). Although similar evidence for such a role of Plk1 in MPF activation during maturation of mammalian oocytes is absent, changes in Plk1 enzyme activity correlate with MPF activation, Plk1 co-localizes with MPF, and microinjection of antibodies neutralizing Plk1 delays GVBD. In this study, we exploited the prolonged time required for maturation of porcine oocytes to define precisely the timing of Plk1 and MPF activation during maturation. GVBD typically occurs between 24 and 26 hr of culture in vitro and meiotic maturation is completed after 40-44-hr culture. We find that Plk1 is activated before MPF, which is consistent with its role in activating MPF in mammalian oocytes. PMID- 15278899 TI - Vitellogenin mRNA expression in Cherax quadricarinatus during secondary vitellogenic at first maturation females. AB - PCR products of 1.1 and 0.9 kb were generated using Cherax quadricarinatus genomic DNA in the first case, and hepatopancreas and ovary cDNAs in the second case. These PCR products were cloned and analyzed for nucleotide sequences. The 1.1 kb fragment was used as a probe for Northern hybridization, revealing a transcript of approximately 8 kb in both tissues. Results from both Northern blot and RT-PCR analyses showed that the mRNA enconding the 3' end of the vitellogenin cDNA was present simultaneously in both hepatopancreas and ovary tissues in secondary vitellogenic at first maturation females, but was not detected in male hepatopancreas. The deduced amino acid sequences of Vitellogenin (Vg) cDNAs from ovary and hepatopancreas confirmed the existence at least two different Vg genes, and two different sites of synthesis. PMID- 15278900 TI - Effect of paternal tamoxifen on the expression of insulin-like growth factor 2 and insulin-like growth factor type 1 receptor in the post-implantation rat embryos. AB - Nuclear transplantation studies demonstrated the importance of paternal contribution to embryogenesis. Paternal treatment with agents like cyclophosphamide and 5-azacytidine has been shown to cause an increase in pre implantation loss (PIL) and post-implantation loss (POL). Studies from our laboratory have shown that paternal tamoxifen treatment increases PIL and POL. It was observed that the PIL occurred at day 2 of gestation (embryo at 2-4 cell stage) and the POL occurred around day 9 of gestation (mid-gestation). The insulin-like growth factor (IGF) system represents one of the major growth controlling system expressed in the embryo. Several studies suggest that in rodents, insulin-like growth factor 2 (Igf2) signaling through the insulin-like growth factor type 1 receptor (Igf1r) modulates embryo growth at around days 9-11 of gestation (mid-gestation). The present study was undertaken to evaluate the expression of Igf2 and Igf1r transcript by RT-PCR in the post-implantation embryos obtained after paternal tamoxifen treatment. It was observed that both the genes were down regulated in resorbed embryos (POL). Since Igf2 is an imprinted gene and the imprint mark is established during spermatogenesis, the present study suggests that paternal tamoxifen treatment may have affected imprinting of the gene during spermatogenesis thereby decreasing its expression and leading to increase in POL. This is to our knowledge the first study correlating the increase in post-implantation embryo loss obtained after paternal drug treatment with the decrease in the expression of Igf2 in these embryos. PMID- 15278901 TI - Rapid and improved method for windowing eggs accessing the stage X chicken embryo. AB - The chick stage X blastoderm is routinely accessed through a small window in a freshly laid egg. However, windowing severely compromises embryo survival with hatch rates as low as a few percent. We previously reported a simple modification to the standard method that reduced introduction of air into the sealed egg and improved the hatchability to 32%. Here, we describe an even simpler and more rapid method for sealing a windowed egg using hot glue or paraffin in which the hatch rate increased to an average of 63% of the unwindowed control eggs. The primary reason for low hatchability can be attributed to air trapped within the egg during windowing and/or leakage during incubation, as shown by increased lethality by artificially introducing air into windowed and sealed eggs. Although the hatch rate was considerably improved, air can still enter the egg during incubation and is likely to account for less than 100% hatchability of the sealed eggs. The success of this new windowing method will facilitate high throughput for the production of transgenic birds and find use in developmental biology, toxicity testing, and avian disease research. PMID- 15278902 TI - Sexing of in vitro produced ovine embryos by duplex PCR. AB - The aim of this article was to develop a fast and easy duplex polymerase chain reaction (PCR) method, for sex determination of ovine in vitro produced embryos prior to implantation. We tested the approach with 107 samples of autosomal cells (oviductal sheep cells and male lamb fibroblasts), divided into three groups for each sex according to the number of cells employed (30, 5, 2, respectively). We then used the test on 21 embryos at blastocyst stage. On the same day the embryos were transferred in pairs into 11 recipient synchronized ewes. The PCR utilized two different sets of primers: the first pair recognized a bovine Y-chromosome specific sequence (SRY), that showed 100% homology with the corresponding sequence of the ovine Y-chromosome and is amplified in males only. The second pair recognized the bovine 1.715 satellite DNA (SAT) which was amplified in all ovine samples but, when submitted to the GenBank database did not show homology with any of the reported ovine sequences. However, after sequencing, ovine amplification product showed 98% homology with the bovine specific satellite sequence. The autosomal samples were amplified with 85.0% efficiency and 91.2% accuracy, while amplification was successful with all 21 embryos (100% efficiency). Eight lambs were born and the sex as determined by PCR corresponded to the anatomical sex in seven (87.5% accuracy). These results confirm that this method can be applied in ovine breeding programs to manipulate sex ratio of offspring. PMID- 15278903 TI - Identification of differentially regulated genes in bovine blastocysts using an annealing control primer system. AB - The identification of embryo-specific genes would provide insights into early embryonic development. However, the current methods employed to identify the genes that are expressed at a specific developmental stage are labor intensive and suffer from high rates of false positives. Here we employed a new and accurate reverse transcription-polymerase chain reaction (RT-PCR) technology that involves annealing control primers (ACPs) to identify the genes that are specifically or prominently expressed in bovine early blastocysts and hatched blastocysts produced in vitro. Using these techniques, a total of nine expressed sequence tags (ESTs) of genes that were differentially expressed in hatched blastocysts, as compared to blastocyst embryos, were cloned and sequenced. The cloned genes or ESTs (C1-C9) all exhibited significant sequence similarity with known bovine genes (99-100%; FTL, RPS12, LAPTM4a, and RPL12) or ESTs (80-94%; AIBP, CULLIN-1, HDLP, COX5a, and RECS1) of other species. As revealed by real time RT-PCR, these genes were regulated upstream in the hatched blastocyst stage during early implantation. These results suggest that this new, PCR-based differential display RT-PCR technique is a very useful tool for the identification of stage-specific genes of preimplantation embryos. PMID- 15278904 TI - Effect of chemically defined culture medium supplemented with beta mercaptoethanol and amino acids on implantation and development of different stage in vivo- or in vitro-derived mouse embryos. AB - In vitro culture (IVC) systems are required for many biotechnological and assisted reproductive technologies and the researchers have been modifying in vitro embryo culture conditions to reach the comparable efficiencies provided in vivo. In the present study, the effects of beta-mercaptoethanol (Beta-ME) and amino acids (AA) on the development of mouse embryos obtained in vivo or in vitro at different stages were investigated. Chemically defined potassium simplex optimized medium (KSOM) was used as basic culture medium and six experimental groups were established and by supplementation of Beta-ME and AA into KSOM media. The quality of blastocysts was evaluated by counting the cells and determining the ratio of inner cell mass (ICM) to trophoectoderm (TE) cells. In addition, embryo transfer (ET) was performed to investigate the rate of implantation and live fetuses. The results obtained in the present study demonstrated that the combined treatment of Beta-ME and AA to 1-cell stage embryos not only enhanced in vitro development to the blastocyst stage but also improved both the number of blastocysts cells and live fetuses. PMID- 15278905 TI - Beta 2-agonist treatment enhances uterine oxytocin receptor mRNA expression in pregnant rats. AB - The objective of this study was to disclose an interaction between Beta(2) adrenergic (Beta(2)-ARs) and oxytocin (OT) receptors (OTRs) in the late-pregnant rat uterus. We investigated the level of uterine OTR mRNA expression after the administration of Beta(2)-AR agonists fenoterol and hexoprenaline to rats from day 18 to 22 of pregnancy, and also tested the effect of fenoterol on uterine explants. Hexoprenaline induced a maximum 24% increase of OTR mRNA. Fenoterol in vivo elicited a maximum 125% increase of OTR mRNA, in vitro produced a maximum fourfold increase in OTR mRNA. In fenoterol-treated rats the maximal contractility increasing effect of OT on isolated uterine rings was significantly higher than in intact term pregnant rats, but the EC50 values were not statistically different. It was concluded that the enhanced expression of OTR mRNA induced by Beta(2)-agonists in the late-pregnant rat uterus may be a possible drawback to effective therapy of preterm uterine contractions with Beta(2)-agonists. PMID- 15278906 TI - Fate of the first polar bodies in mouse oocytes. AB - Both nuclear transfer and intracytoplasmic sperm injection (ICSI) practice necessitates studies on the spatial relationship between the MII spindle and the first polar bodies (FPB). Although recent observations have shown that the FPB position does not predict accurately the location of the meiotic spindle in metaphase II oocytes of monkey, hamster, and human, detailed studies on FPB deviation and its affecting factors are lacking. Since polar bodies can be used for genetic testing and oocyte quality grading, their life span under different conditions should be studied. The timing of formation and degeneration and the position relative to the MII spindle of the FPB and the factors affecting FPB deviation and degeneration during in vivo and in vitro aging of both in vivo and in vitro matured mouse oocytes were investigated in this study. Mice of the Kun ming breed were used, and the intact and degenerated FPB were identified through microscopic morphology in combination with propidium iodide (PI) exclusion test and the chromosomes visualized by Hoechst staining. Results are summarized as follows: (i) oocytes started FPB extrusion at 8 hr after the onset of in vivo or in vitro maturation, but the number of FPB reached maximum much later in vitro (14 hr of culture) than in vivo (10 hr post hCG). (ii) Some FPB began to degenerate before ovulation and around 70% became degenerated within 6 hr after maximal nuclear maturation both in vivo and in vitro; they disappeared faster during in vivo than in vitro aging but turned from intact to degenerated at a similar tempo. (iii) Some FPB began to deviate from the MII spindle 10 hr after hCG injection or in vitro culture and the distance between FPB and the spindle increased with time during both in vivo and in vitro aging. (iv) FPB deviated more slowly in the in vitro matured oocytes than in in vivo matured. (v) Denudation performed after FPB extrusion markedly enhanced its deviation. (vi) The perivitelline space (PVS) increased with time during maturation and aging in vivo and in vitro and the values of PVS and the percentages of FPB adjacent to the spindle were significantly negatively correlated. (vii) Cytochalasin B and colchicine had no effect on FPB deviation. (viii) None of the more than 3,500 FPBs observed was found to be dividing or have divided into two cells at any time points before or after ovulation or in vitro maturation. Our results were consistent with the possibility that the displacement of the FPB was a time- and PVS-dependent process, indicating that PVS would increase with time and its formation and enlargement would facilitate the lateral displacement of the degenerating FPB. PMID- 15278907 TI - Phosphatidylinositol 3-kinase and Akt participate in the FSH-induced meiotic maturation of mouse oocytes. AB - Phosphatidylinositol 3-kinase (PI3K) is known to play critical roles in signal transduction processes related to a variety of cellular activities. In the present study, we investigated the role of PI3K during meiotic maturation in mouse oocytes using a specific inhibitor, LY294002. In follicle-stimulating hormone (FSH)-induced reversal of hypoxanthine-mediated meiotic arrest of cumulus oocyte complexes (COCs), LY294002 suppressed germinal vesicle breakdown (GVBD), first polar body (PB1) emission, and cumulus expansion. To examine the effect of LY294002, denuded oocytes (DOs) were cultured in medium containing follicular fluid meiosis-activating sterol (FF-MAS) since absence of gonadotropin receptors in oocytes has been reported and FSH did not stimulate meiotic maturation of DOs in the presence of hypoxanthine. In FF-MAS-induced maturation of DOs, LY294002 suppressed PB1emission, but not GVBD. In spontaneous gonadotropin-independent oocyte maturation, LY294002 had no effect on COCs and DOs. Akt/protein kinase B, a serine-threonine kinase, is a key downstream effector of the PI3K pathway. Therefore, we also examined the distribution of Akt during FSH-induced meiotic maturation. The distribution of Ser(473) phosphorylated Akt was similar to the localization of microtubules, while Thr(308) phosphorylated Akt was present in the pericentriolar materials (PCM) in metaphase I (MI) and II (MII) oocytes. LY294002 decreased the amount of Thr(308) phosphorylated Akt to very low to undetectable levels in MI and MII oocytes. Ser(473) phosphorylated Akt showed aberrant distribution and very low to undetectable levels of expression in LY294002-treated MI and MII oocytes, respectively. These results suggest that PI3K and Akt participate in mouse meiotic maturation. PMID- 15278908 TI - Proteasomal activity in mammalian spermatozoa. AB - The proteasome is a multicatalytic cellular complex, which possess three different enzymatic activities, trypsin-like, chymotrypsin-like, and peptidylglutamyl peptidase. Its function is to remove abnormal or aged proteins. Recently, it has been suggested the participation of the sperm proteasome during mammalian fertilization. In this study, we present evidence that indicates that sperm extracts from several mammalian species, including hamster, mice, rats, bovine, rabbits, and humans all possess proteasome activity. We characterized the three specific activities of the proteasome using specific synthetic substrates and specific proteasome inhibitors. The results indicates that the highest specific activity detected was in mouse sperm toward the trypsin substrates and it was 1,114% of the activity of human sperm toward the chymotrypsin substrate Suc-Leu-Leu-Val-Tyr-AMC (SLLVY-AMC, which was considered as 100%). In all cases, the lowest activity was toward substrates for the peptidylglutamyl peptidase hydrolyzing activity, and it was lowest for rabbit sperm (1.7% of the activity of human sperm toward the chymotrypsin substrate SLLVY-AMC). In addition, specific proteasome inhibitors were able to block all proteasome activities almost 100%, with the exception of clasto-Lactacystin beta-lactone upon rat sperm. All sperm extracts tested evidenced bands of about 29-32 kDa by Western blots using a monoclonal antibody against proteasome subunits alpha 1, 2, 3, 5, 6, and 7. In conclusion, sperm from several mammals possess enzymatic activities that correspond to the proteasome. The proteasome from the different species hold similar but distinctive enzymatic characteristics. PMID- 15278909 TI - The regulation of competence to replicate in meiosis by Cdc6 is conserved during evolution. AB - DNA replication licensing is an important step in the cell cycle at which cells become competent for DNA replication. When the cell cycle is arrested for long periods of time, this competence is lost. This is the case for somatic cells arrested in G0 or vertebrate oocytes arrested in G2. CDC6 is a factor involved in replication initiation competence which is necessary for the recruitment of the MCM helicase complex to DNA replication origins. In Xenopus, we have previously shown that CDC6 is the only missing replication factor in the oocyte whose translation during meiotic maturation is necessary and sufficient to confer DNA replication competence to the egg before fertilization (Lemaitre et al., 2002: Mol Biol Cell 13:435-444; Whitmire et al., 2002: Nature 419:722-725). Here, we report that this oogenesis control has been acquired by metazoans during evolution and conserved up to mammals. We also show that, contrary to eukaryotic metazoans, in S. pombe cdc18 (the S. pombe CDC6 homologue), CDC6 protein synthesis is down regulated during meiosis. As such, the lack of cdc18 prevents DNA replication from occurring in spores, whereas the presence of cdc6 makes eggs competent for DNA replication. PMID- 15278910 TI - Retinoid-dependent mRNA expression and poly-(A) contents in bovine oocytes meiotically arrested and/or matured in vitro. AB - The presence of retinoic acid (RA) during in vitro maturation (IVM) improves bovine oocyte quality and developmental potential. In this work, we investigated the underlying molecular mechanisms. Cumulus-oocyte complexes were meiotically arrested by roscovitine and/or matured in defined medium containing RA, 1% ethanol (vehicle), or no additives. Cumulus-free oocytes were analyzed for poly (A) mRNA contents and relative mRNA expression of genes involved in cell cycle regulation (cyclin B1 and H1) and antioxidative defence (Mn-superoxide dismutase and glucose-6-phosphate dehydrogenase). Poly-(A) mRNA increased after meiotic inhibition and decreased with IVM completion, both in meiotically arrested and permissively matured oocytes, i.e., matured without previous meiotic arrest. RA dramatically increased poly-(A) mRNA in meiotically arrested oocytes, but more than half of the poly-(A) mRNA disappeared during maturation. Irrespective of oocyte origin, transcripts were detected for all the genes analyzed. IVM, with or without previous meiotic inhibition, increased expression of cyclin B1 and glucose-6-phosphate dehydrogenase, and decreased cyclin H1 and Mn-superoxide dismutase. Except for a decreasing of Mn-superoxide dismutase in meiotically arrested and matured oocytes, RA did not affect mRNA expression. Ethanol led to an abnormal poly-(A) mRNA profile and expression of all the genes analyzed. RA does not modify expression of cyclin B1 and HI genes in the bovine oocyte, and probably does not generate oxidative stress. In addition, RA enhanced mRNA amount as measured by poly-(A) mRNA contents. PMID- 15278911 TI - Vertebrate yolk proteins: a review. PMID- 15278912 TI - Relationships between cagA, vacA, and iceA genotypes of Helicobacter pylori and DNA damage in the gastric mucosa. AB - Helicobacter pylori (H. pylori) is believed to predispose carriers to gastric cancer by inducing chronic inflammation. The inflammatory processes may result in the generation of reactive oxygen and nitrogen species that damage DNA. In this study, we investigated the relationships between DNA damage in the gastric mucosa and cagA, vacA, and iceA genotypes of H. pylori. The study was conducted with biopsies from the gastric antrum and corpus of 98 H. pylori-infected and 26 uninfected control patients. H. pylori genotypes were determined by PCR and DNA damage was measured in gastric mucosal cells by the Comet assay (single cell gel electrophoresis). All patients were nonsmokers, not abusing alcohol, and not using prescription or recreational drugs. Levels of DNA damage were significantly higher (P < 0.0001) in the H. pylori-infected patients than in uninfected patients. In comparison with the level of DNA damage in the uninfected controls, the extent of DNA damage in both the antrum (OR = 8.45; 95% CI = 2.33-37.72) and the corpus (OR = 6.55; 95% CI = 2.52-17.72) was related to infection by cagA+/vacAs1m1 and iceA1 strains. The results indicate that the genotype of H. pylori is related to the amount of DNA damage in the gastric mucosa. These genotypes could serve as biomarkers for the risk of extensive DNA damage and possibly gastric cancer. PMID- 15278913 TI - Effect of artificial mixtures of environmental polycyclic aromatic hydrocarbons present in coal tar, urban dust, and diesel exhaust particulates on MCF-7 cells in culture. AB - Human exposure to polycyclic aromatic hydrocarbons (PAHs) occurs through complex mixtures. The National Institute of Standards and Technology has established standard reference materials (SRMs) for selected PAH mixtures that are composed of carcinogenic, noncarcinogenic, and weakly carcinogenic compounds, such as those derived from coal tar (SRM 1597), atmospheric particulate matter (SRM 1649), and diesel particulate matter (SRM 1650). To study the effects of PAHs with different carcinogenic potential in complex mixtures, and to investigate the metabolic activation of noncarcinogenic and weakly carcinogenic PAHs to DNA binding derivatives, artificial mixtures (1597H, 1649H, and 1650H) were prepared in the laboratory. These artificial mixtures contained the same relative ratios of noncarcinogenic and weakly carcinogenic PAHs present in SRM 1597, SRM 1649, and SRM 1650. The human mammary carcinoma-derived cell line MCF-7 was treated with these artificial mixtures and analyzed for PAH-DNA adduct formation and the induction of cytochrome P450 (CYP) enzymes. We found that the artificial mixtures formed lower but detectable levels of DNA adducts 24 and 48 hr after treatment than benzo[a]pyrene. Induction of CYP enzyme activity was measured by the ethoxyresorufin-O-deethylase assay, and the expression of CYP1A1 and CYP1B1 was confirmed by immunoblots. Both noncarcinogenic and weakly carcinogenic PAHs present in the artificial mixtures have the ability to induce CYP1A1 and CYP1B1 in MCF-7 cells and contribute to DNA binding. Therefore, it is necessary to take into account the noncarcinogenic and weakly carcinogenic PAHs present in environmental mixtures in assessing the potential risk associated with human exposure. PMID- 15278914 TI - Dose- and time-dependent responses for micronucleus induction by X-rays and fast neutrons in gill cells of medaka (Oryzias latipes). AB - Medaka fish (Oryzias latipes) were exposed to various doses of X-rays or fast neutrons, and the frequency of micronucleated cells (MNCs) was measured in gills sampled at 12- or 24-hr intervals from 12 to 96 hr after exposure. The resulting time course of MNC frequency was biphasic, with a clear peak 24 hr after exposure, irrespective of the kind of radiation applied and the dose used. The half-life of MNCs induced in the gill tissues by the two exposures fluctuated around 28 hr, with no significant dose-dependent trend for either X-ray- or neutron-exposed fish. As assayed 24 hr after exposure, the MNC frequency increased linearly over the control level with increasing doses of both X-rays and fast neutrons. The relative biological effectiveness (RBE) of fast neutrons to X-rays for MNC induction was estimated to be 4.3 +/- 0.6. This value is close to the RBE value of 5.1 +/- 0.3 reported for fast neutron induction of somatic crossing-over mutations in Drosophila melanogaster that arise from recombination repair of DNA double-strand breaks. These results and other data support our conclusion that the medaka gill cell micronucleus assay is a reliable short-term test for detecting potential inducers of DNA double-strand breaks. PMID- 15278915 TI - Chemopreventive effects of vanadium toward 1,2-dimethylhydrazine-induced genotoxicity and preneoplastic lesions in rat colon. AB - In the present study, we have evaluated the antitumor effects of vanadium by monitoring DNA damage and chromosomal aberrations (CAs) during the early preneoplastic stage of 1,2-dimethylhydrazine (1,2-DMH)-induced colon cancer in male rats. Treatment with 20 mg/kg 1,2-DMH for 6 weeks resulted in the formation of aberrant crypt foci (ACF), a putative preneoplastic lesion associated with colon cancer development, while cotreatment with ammonium monovanadate (0.5 ppm in the drinking water) reduced ACF formation by 50% (P < 0.001). The 6-week treatment with 1,2-DMH also resulted in significantly higher levels of DNA damage in rat colon as measured by the Comet assay (higher mean values for length-to width ratios (L:W) of DNA mass (P < 0.01) and mean frequencies of cells with comets (P < 0.001)). The vanadium cotreatment reduced DNA damage in colon cells by 32% (P < 0.02 and P < 0.001 for L:W and tailed cells, respectively). 1,2-DMH treatment also produced a 10-fold increase in the frequency of CAs in rat colon (P < 0.001), while cotreatment with vanadium resulted in a reduction in CAs after 2, 4, and 6 weeks of 1,2-DMH exposure (P < 0.01). Analysis of antioxidant defense enzyme activity in colonic mucosa indicated that glutathione reductase and catalase activities were increased in 1,2-DMH-treated rats; cotreatment with vanadium reduced these activities when compared to the carcinogen control (P < 0.001 and P < 0.02). These results demonstrate that the early protective effect of vanadium in chemically induced rat colon carcinogenesis may be mediated by a reduction of carcinogen-induced DNA damage. PMID- 15278916 TI - Spontaneous mutant frequency and mutation spectrum for gene A of phiX174 grown in E. coli. AB - The use of transgenic targets for measuring mutant frequencies in mammalian tissue requires an estimate of the mutant frequency that results from recovery of the transgene in bacterial recovery systems. In this study, we have determined the spontaneous mutant frequency, estimated the mutation rate, and ascertained the mutation spectrum for gene A of phiX174 grown in E. coli strain CQ2 from 156 small independent cultures. The mutant frequency of 12 of the 156 cultures was 17 +/- 1.0 x 10(-6) and the estimated mutation rate per gene replication was 7.4 +/- 2.3 x 10(-6). The mutant frequency and spectrum from E. coli were not significantly different from that of solvent-treated embryonic mouse cells in culture, 19 +/- 0.5 x 10(-6) (Valentine CR et al. [2002]: Environ Mol Mutagen 39:55-68), indicating that those spontaneous mutants were primarily derived from E. coli. The E. coli spectrum was heavily weighted toward two major target sites (hot spots), 4225A-->G (56%) and 4218G-->A or C (20%). Four new target sites and one new mutational event were recovered by the gene A forward assay. A mutant spectrum from an expanded phage stock was also determined to assess the effects of propagating the virus. This mutant frequency was higher (6 x 10(-4)), contained more double mutants (15% compared to 0.6%), and had a significantly different spectrum from the spectrum for independent cultures (fewer A:T-->G:C and G:C-->C:G changes and more G:C-->A:T; P < 0.002). The E. coli mutation spectrum will be useful for determining the origin of gene A mutation in tissues of phiX174 transgenic mice. PMID- 15278917 TI - In vivo mutation in gene A of splenic lymphocytes from phiX174 transgenic mice. AB - Single-burst analysis was applied to a forward assay for gene A mutation in splenic lymphocytes of phiX174 transgenic mice for the purpose of optimizing analytical parameters for identifying in vivo mutations. The effect of varying the cutoff value for an in vivo burst on induced mutant frequency, fold increase, and the significance of the difference between control and N-ethyl-N-nitrosourea (ENU)-treated mice was calculated by two different methods. The plating density was reduced to an average of less than 10 background mutant plaques per aliquot in order to separate in vitro bursts. The spectrum of mutations contributing < 60 plaques per aliquot from control animals was not significantly different from the control spectra from E. coli or transgenic phiX174 cells in culture. The mutant spectra from ENU-treated animals was highly different between mutant bursts of > 80 plaques per aliquot compared to mutations contributing < 60 plaques per aliquot (P < 0.000001), the former fitting the spectrum expected for ENU-induced mutations. The latter spectrum was also different from control animals and E. coli (P < 0.000001), suggesting the difference was caused by ex vivo mutation. With the mutations found in this study, the total number of reported target sites for gene A is now 33. The results support the interpretation that, in contrast to results for the lacI transgene, 100% of mutants isolated in gene A from control animals and cells were fixed in E. coli. We attribute the difference between the two genes to hot-spot sites for mutation in gene A and to a testable hypothesis that the mosaic plaque assay for the lacI transgene underestimates the frequency of ex vivo mutants. PMID- 15278918 TI - DNA damage produced in HaCaT cells by combined fluoranthene exposure and ultraviolet A irradiation. AB - Fluoranthene is a polycyclic aromatic hydrocarbon (PAH) and a principal constituent of PAH-contaminated aquatic systems. In the present study, fluorescein diacetate uptake and the Comet assay were used to assess the cytotoxicity and genotoxicity of fluoranthene in HaCaT (human adult low calcium high temperature) cells in the presence or absence of ultraviolet A (UVA) irradiation. Exposure of cells to 0.1, 0.25, 0.75, 2, and 5 microM fluoranthene alone for 30 min or to 6.1 +/- 0.07 J/cm2 UVA alone did not cause cytotoxicity or cellular DNA damage. However, concomitant exposure to both caused a nonlinear dose-response in cytotoxicity to HaCat cells. The same exposure conditions also resulted in a dose-responsive DNA damage in HaCaT cells. Because DNA damage mainly was detected at relatively high levels of cytotoxicity, we cannot rule out the possibility that it occurred as a consequence of cellular toxicity mechanisms. PMID- 15278920 TI - Evaluation of DNA intercalation potential of pharmaceuticals and other chemicals by cell-based and three-dimensional computational approaches. AB - To what extent noncovalent chemical-DNA interactions, in particular weak nonbonded DNA intercalation, contribute to genotoxic responses in mammalian cells has not been fully elucidated. Moreover, with the exception of predominantly flat, multiple-fused-ring structures, our ability to predict intercalation ability of novel compounds is nearly completely lacking. Computational programs such as DEREK and MCASE recognize primarily those molecules that can form irreversible covalent adducts with DNA since their learning sets, for the most part, have not been populated by compounds for which a relationship between noncovalent interaction and genotoxicity exists. We describe here a novel three dimensional (3D) computational DNA-docking model for prediction of DNA intercalative activity of molecules with both classical and nonclassical intercalating structures. The 3D docking results show a remarkable concordance with results obtained from testing these molecules directly in the Chinese hamster V79 cell-based bleomycin amplification system suggesting that either or both of these approaches may have utility in defining noncovalent chemical-DNA interactions. The ability to predict and/or demonstrate cellular DNA intercalation of novel molecules may well provide fresh insights into the nature and mechanistic basis of structurally unexpected genotoxicity observed during safety testing. PMID- 15278919 TI - Comparison of two particulate hexavalent chromium compounds: Barium chromate is more genotoxic than lead chromate in human lung cells. AB - Particulate hexavalent chromium [Cr(VI)] compounds are well-established human lung carcinogens. However, their carcinogenic mechanisms are poorly understood as most investigators have used soluble Cr(VI) compounds. Recent work from our laboratory has found that barium chromate (BC) is also cytotoxic and clastogenic. To understand how BC relates to existing data on other particulate Cr(VI) compounds, we compared its cytotoxicity and clastogenicity with lead chromate (LC), which has been used as a prototypical particulate Cr(VI) compound, in WTHBF 6 cells, a near-normal human lung cell line. We found that BC is a more potent cytotoxicant, inducing 67%, 12%, 3%, and 0% relative survival at concentrations of 0.1, 0.5, 1, and 5 microg/cm2, respectively, while LC induced 90%, 71%, 43%, and 15% survival at these same concentrations. We found that BC was also more clastogenic, damaging 22% and 49% of metaphase cells at 0.1 and 0.5 microg/cm2, and causing complete cell cycle arrest at 1 and 5 microg/cm2. By contrast, 0.1, 0.5, and 1.0 microg/cm2 LC damaged 10%, 27%, and 37% of metaphase cells, respectively, and complete cell cycle arrest was not observed until a concentration of 5 microg/cm2 was reached. We found that BC and LC both partially dissolved in complete medium in the presence of cells, producing similar extracellular concentrations. Both compounds were also comparable with respect to particle uptake and the amount of intracellular Cr ions. Considering previous reports showing that lead ions were inactive and that sodium chromate and LC have similar clastogenic potencies, these data suggest that BC genotoxicity may not be solely mediated by Cr ions, but also involve some clastogenic activity of barium ions. PMID- 15278921 TI - Folate supplementation of cyclophosphamide-treated mothers diminishes micronucleated erythrocytes in peripheral blood of newborn rats. PMID- 15278922 TI - Closure of skin incisions in rabbits by laser soldering: I: Wound healing pattern. AB - BACKGROUND AND OBJECTIVES: Temperature-controlled tissue laser soldering is an innovative sutureless technique awaiting only solid experimental data to become the gold-standard surgical procedure for incision closure. The goals of the current study were: (1) to define the optimal laser soldering conditions, (2) to explore the immediate skin reparative healing events after sealing the wound, and (3) to determine the long-term trajectory of skin wound healing. STUDY DESIGN/MATERIALS AND METHODS: Skin incisions were generated over rabbit dorsa and were closed using different wound-closure interventions, in three groups: (a) closure, using a temperature-controlled infrared fiberoptic CO2 laser system, employing 47% bovine serum albumin as a solder; (b) wound closure by cyanoacrylate glues; and (c) wound closure by sutures. The reparative outcomes were evaluated macroscopically and microscopically, employing semi-quantitative grading indices. RESULTS: Laser soldering of incisions at T = 65 degrees C emerged as the optimal method achieving immediate wound sealing. This in turn induced accelerated reparative events characterized by a reduced inflammatory reaction, followed by minimal scarring and leading to a fine quality healing. CONCLUSIONS: Temperature-controlled laser soldering offers an accelerated wound reparative process with numerous advantages over the conventional methods. Further investigations may reveal additional benefits in the spectrum of advantages that this innovative surgical technology has to offer. This can introduce new scientific insight that will pave the way for clinical use. PMID- 15278923 TI - Closure of skin incisions in rabbits by laser soldering II: Tensile strength. AB - BACKGROUND AND OBJECTIVES: The basic characteristic property of wound closure is the immediate and long-term tensile strength (LTS). The objective of the current study was to compare tissue laser soldering to other available methods (i.e., cyanoacrylate glues and sutures) in the performance and outcome of wound closure and reparative healing process, with an emphasis on the immediate and LTS. STUDY DESIGN/MATERIALS AND METHODS: The animals were divided into three groups according to the type and details of the closure procedure. Group A: laser treatments at different temperatures were compared to sutured incisions, emphasizing the LTS after 10 days. Group B: laser soldering at 65 +/- 5 degrees C was compared to chemical glues (i.e., Histoacryl and Dermabond), emphasizing the immediate tensile strength (ITS). Group C: LTS of laser soldered incisions was compared to that of sutured incisions at various time intervals emphasizing LTS (3, 7, 14, 28 days). RESULTS: Group A: LTS at 60 degrees C exhibited the highest values (0.48 MPa). Group B: no ITS difference was detected between laser soldering and chemical glues. Group C: soldered incisions at 65 degrees C exhibited higher LTS (1.81 MPa) than that of sutured incisions (1.08 MPa) (P < 0.043). CONCLUSIONS: Temperature-controlled laser soldering at 65 degrees C provided sufficient ITS and higher bonding LTS values compared with sutures, resulting in better wound healing characteristics. The laser soldering system presented here should be tested on larger animal models before adopting it for clinical usage. PMID- 15278924 TI - Theoretical and experimental investigation of the thermal effects within body cavities during transendoscopical CO2 laser-based surgery. AB - BACKGROUND AND OBJECTIVES: The recent development of flexible hollow waveguides for MID-IR lasers may be utilized transendoscopically to ablate selectively neoplastic, superficial tissues within body cavities. Study goals are to investigate theoretically and experimentally heat distribution and thermal response of cavity lining, during CO2 laser minimally invasive surgery (MIS), and to thermally optimize the procedure under practical conditions. STUDY DESIGN/MATERIALS AND METHODS: Mathematical model was developed to predict temperature distribution along cavity lining. Experimental setup was built, including all the necessary components for a fully feedback-controlled MIS, i.e., laser generator, gas insufflating system, surgical suction, and infrared imaging feedback mechanism, all controlled by central PC-based program. Thermal images of cavity lining were recorded and analyzed throughout varying conditions. RESULTS: Thermal gradients along the cavity lining, during and after the laser irradiation, were obtained mathematically and experimentally. Diverse modes of heat dispersions were observed, as well as the relative contributions of user controlled parameters to the maximal heat of cavity lining. The software controlled setup has demonstrated the capacity to instantly manage varying conditions, by which it automatically protects cavity lining from getting overheated. CONCLUSIONS: Analytical predictions and experimental measurements were highly correlated. The software-controlled system may serve a powerful tool to control thermal side effects during MIS within body cavities. PMID- 15278925 TI - On appropriate pathology for photothermal surgery. AB - BACKGROUND AND OBJECTIVES: The standard hematoxylin and eosin (H&E) stain can yield dramatic underestimates of tissue death when pathology is performed immediately after radiative surgery. Our objectives were to identify an accurate, consistent histology, and to understand under what conditions the problem is manifested. STUDY DESIGN/MATERIALS AND METHODS: Photothermal surgery with highly concentrated sunlight was performed ex vivo on chicken livers, and in vivo on healthy live rats that were sacrificed at periods of 0, 24, 48, and 72 hours. Pathology with both H&E and an alternative NADH-diaphorase (NADH) stain were prepared. RESULTS: H&E pathology performed shortly after surgery errs by close to an order of magnitude in appraising lesion volume, but the shortfall is mitigated when histological examination is postponed for 24 hours. The biochemical signals missed by the H&E technique at short times can be captured by the NADH stain which reveals the cessation of enzymatic activity, albeit at greater expense and complexity. CONCLUSIONS: The significance lies in avoiding false positives in surgery, providing reliable estimates of photothermal surgical efficacy, and highlighting the relative merits of alternative histological practices with photothermal surgery. PMID- 15278926 TI - The vascular response induced by transmyocardial laser revascularization is determined by the size of the channel scar: Results of CO2, holmium and excimer lasers. AB - BACKGROUND AND OBJECTIVES: To investigate the angiogenic effect of CO2, Ho:YSGG, and XeCl excimer TMLR in a rat model with morphologic characteristics of chronic myocardial ischemia. STUDY DESIGN/MATERIALS AND METHODS: Two channels (200-320 microm) were created per rat heart. After 14 days, vessel numbers and densities in and around laser scars were assessed. RESULTS: Capillary densities in the laser scars were equal between the three lasers ( approximately 130 vessels/mm2) but much lower than in control areas ( approximately 2,100 vessels/mm2). Vessel densities excluding capillaries were significantly higher in Ho:YSGG and CO2 scars compared to excimer scars, while only Ho:YSGG scars contained significantly more large vessels (diameter > or = 20 microm) than control areas. Only rarely, extension of vascular growth into adjacent myocardium was observed in any of the three groups. CONCLUSIONS: These results indicate that the angiogenic response following TMLR is limited to the channel scar and related to the scar size rather than the specific laser type. PMID- 15278927 TI - Thermal therapy of canine cerebral tumors using a 980 nm diode laser with MR temperature-sensitive imaging feedback. AB - BACKGROUND AND PURPOSE: The laser-induced thermal therapy (LITT) of cerebral tumors has conventionally been performed using Nd:YAG lasers and is associated with a risk of high focal temperatures potentially followed by cavitation that could result in boiling and/or explosive char. We have developed small diffusing laser fiber tips to better distribute the energy deposition and a computer controlled feedback system to monitor therapy and prevent excess temperature buildup. In this study, we evaluated the feasibility of using magnetic resonance temperature imaging (MRTI)-based feedback system for the thermal treatment of experimental intracerebral tumors using 980 nm laser irradiation delivered through these diffusing tips. STUDY DESIGN/MATERIALS AND METHODS: Transmissible venereal tumors (TVTs) were grown via inoculation in the right cerebral hemisphere of seven canines. The laser fiber tips were inserted into a total of 10 independent TVT-suspected regions in the seven animals. Margins for the target area in each animal were prescribed on the basis of pretreatment MR images. MRTI based feedback software was used to measure and regulate both temperature and the delivered thermal dose to achieve the desired thermal ablation and prevent excess heating. The effects of treatment were verified by results of histologic analyses. RESULTS: Treatments resulted in contiguous areas of thermal necrosis in tumors and adjacent brain margin. The feedback software successfully cut off the laser power once the desired treatment volume was achieved, and prevented focal temperatures from exceeding predefined thresholds. Follow-up MRI studies showed 1.4- to 2.9-fold LITT-induced lesion expansion within 1-6 days after treatment. CONCLUSIONS: Targeted thermal coagulation of small intracerebral tumors is feasible using MRTI-based feedback and diffused 980 nm diode laser light. PMID- 15278928 TI - Scanning electron microscopic evaluation of resin-dentin interface after Er:YAG laser preparation. AB - BACKGROUND AND OBJECTIVES: The aim of this study was to observe the morphological characteristics of dentin-resin interfaces when a composite resin was bonded to Er:YAG laser irradiated dentin. STUDY DESIGN/MATERIALS AND METHODS: Human dentin surfaces were divided into three equal areas. One third was prepared using a carbide bur and etched with phosphoric acid, the second third was conditioned using an Er:YAG laser, the third one was irradiated and etched. A hybrid composite resin was bonded on dentin surfaces using a single-component adhesive system. Longitudinal sections were exposed to 5 N HCl then to 1% NaOCl. The thickness of the hybrid layer and the dimensions of the resin tags were measured using scanning electron microscopy (SEM) pictures and an image analysis software. RESULTS: When acid etching was performed, a hybrid layer as well as the characteristic funnel-shaped resin tags were observed. When Er:YAG laser was used alone, no hybrid layer could be detected. The resin tags appeared thinner and exhibited a cylindrical shape. CONCLUSIONS: The acid pre-treatment of the irradiated surface allowed both the seal of the dentinal surface and the increase of the diameter of the resin tags. PMID- 15278929 TI - CO2 laser resurfacing and photocarcinogenesis: an experimental study. AB - BACKGROUND AND OBJECTIVES: To examine whether carbon dioxide (CO2) laser treatment has a carcinogenic potential or may influence ultraviolet (UV)-induced carcinogenesis. STUDY DESIGN/MATERIALS AND METHODS: Hairless mice (n = 211) were treated with a Sharplan CO2 laser with FeatherTouch scanner. Simulated solar irradiations were administrated either pre-operatively or pre- and post operatively. Weekly clinical assessments of skin tumors were performed blinded during the entire observation period of 12 months. RESULTS: No tumors appeared (a) in mice just treated with CO2 laser, (b) in mice exposed to UV irradiation only before CO2 laser treatment or (c) in untreated control mice. Tumors developed in CO2 laser treated mice that were exposed to UV-irradiation both pre- and post-operatively and in UV-irradiated control mice. The time to first, second, and third tumors ranged from 18 to 20 weeks and no significant differences were demonstrated. CONCLUSIONS: CO2 laser treatment does not have a carcinogenic potential in itself, nor does CO2 laser treatment influence UV induced carcinogenesis. PMID- 15278930 TI - Treatment of leg veins with the long pulse dye laser using variable pulse durations and energy fluences. AB - BACKGROUND AND OBJECTIVES: Attempts at using the pulsed dye laser (PDL) operating at 585 nm wavelength and 0.45 milliseconds pulse duration to treat leg veins have been notably unsuccessful. The aim of this study is to evaluate the efficacy and complications of the 595 nm long PDL in the treatment of leg veins. STUDY DESIGN/PATIENTS AND METHODS: Fourteen patients with 38 lesions were treated using a long PDL equipped with a cryogen spray cooling (CSC) device. Variable pulse durations ranging from 1.5 to 20 milliseconds and energy fluences from 10 to 20 J/cm2 were utilized depending on the size of treated vessels. All patients received two laser treatments at an interval of 2 months. RESULTS: Of patients with veins type 1A and type 1, 6 (100%) and 3 (13%) had complete clearing, respectively. Of patients with veins type 1 and type 2, 18 (78.3%) and 2 (22.2%) lesions showed excellent response, respectively. Hyperpigmentation was the only observed complication lasting several months in 57.9% of treated sites. CONCLUSIONS: The long PDL operating at 595 nm is a safe and effective treatment for leg veins, especially in type 1A and type1 vessels. PMID- 15278931 TI - Long pulse Nd:YAG laser for treatment of leg veins in 40 patients with assessments at 6 and 12 months. AB - BACKGROUND AND OBJECTIVES: This study assessed subjectively and objectively the efficacy of a long-pulsed Nd:YAG laser system in clearing dermal leg veins, successful treatment of which remains problematic. STUDY DESIGN/PATIENTS AND METHODS: Forty female patients (24-58 years old, skin types II-IV) with leg veins were treated with synchronized micropulses from a long-pulsed 1,064 nm Nd:YAG laser, 6 mm diameter spot size, 130 and 140 J/cm2. One to three treatments were given at 6-week intervals, with post-treatment assessments at 6 and 12 months. Patients assessed improvement subjectively with a satisfaction index (SI). Objective assessment was based on the clinical photography, and in addition on computer-generated data from a Canny operator-based edge-detection program. RESULTS: The overall patient satisfaction rates and objective assessments at the 6 and 12 month assessments were 42.5 and 57.5%, and 75 and 82.5%, respectively. CONCLUSIONS: The long-pulsed Nd:YAG laser offered efficient treatment of leg veins. Side effects were minimal and transient. The edge-detection program may help patients appreciate better the actual results of the treatment. PMID- 15278932 TI - Differences in production of melanin radicals by 694 nm ruby laser and UVA radiation. AB - BACKGROUND AND OBJECTIVES: The 694 nm ruby laser is used clinically for hair removal and the mechanism is predominantly photo thermal via melanin targeting. We investigated 694 nm laser-irradiation of human hair, and laser-irradiation of synthetic dopa melanin to establish whether photolysis and oxygen radical production is also contributory, and which may have side effects. STUDY DESIGN/MATERIALS AND METHODS: Ultraviolet-A (UVA) irradiation of melanin was used as a positive control for radical production. Laser- and UVA-irradiated hair samples, and synthetic dopa melanin in media of different viscosity, were analyzed using electron spin resonance spectroscopy, and compared. The spin trap 5,5-dimethyl- 1-pyrroline N-oxide (DMPO) was used to probe laser-irradiated dopa melanin for superoxide radical production. RESULTS: Comparable to UVA, laser irradiation of hair increased the signal-intensity of the intrinsic melanin radical. UVA-induced radicals decay rapidly; however, laser-induced radicals decayed slowly and did not fully revert to original levels after 24 hours. Laser induced radicals were increasingly stable with viscosity of the medium. Superoxide radicals were detected using DMPO in UVA- but not laser-irradiated synthetic dopa-melanin at pH 4.5. CONCLUSIONS: Laser-irradiation of melanin does not result in oxygen radical formation; however, a paramagnetic species, long lived in rigid media, is detected which is worth further investigation. PMID- 15278933 TI - Comparison of the effects of laser, ultrasound, and combined laser + ultrasound treatments in experimental tendon healing. AB - BACKGROUND AND OBJECTIVE: Therapeutic ultrasound (US) and laser (L) treatments accelerate and facilitate wound healing, and also have beneficial effects on tendon healing. This randomized control study was designed to evaluate the effects of low-intensity US and low-level laser therapy (LLLT) on tendon healing in rats. STUDY DESIGN/MATERIALS AND METHODS: Eighty-four healthy male Swiss Albino rats were divided into three groups consisting of 28 rats, the left Achilles tendons were used as treatment and the right Achilles tendons as controls. The right and left Achilles tendons of rats were traumatized longitudinally. The treatment was started on postinjury day one. We applied the treatment protocols including low-intensity US treatment in Group I (US Group), Sham US in Group II (SUS Group), LLLT in Group III (L Group), Sham L in Group IV (SL Group), US and LLLT in Group V (US + L Group), and Sham US and Sham L in Group VI (SUS + SL Group). The US treatment was applied with a power of 0.5 W/cm2, a frequency of 1 MHz, continuously, 5 minutes daily. A low-level Ga-As laser was applied with a 904 nm wavelength, 6 mW average power, 1 J/ cm2 dosage, 16 Hz frequency, for 1 minute duration, continuously. In the control groups, the similar procedures as in the corresponding treatment groups were applied with no current (Sham method). The treatment duration was planned for 9 days (sessions) in all groups, except the rats used for biochemical evaluation on the 4th day of treatment, which were treated for 4 days. We measured the levels of the tissue hydroxyproline for biochemical evaluation on the 4th, 10th, and 21st days following the beginning of treatment and the tendon breaking strength on the 21st day following the beginning of treatment for biomechanical evaluation. Seven rats in each group were killed on the 4th, 10th, and 21st days for biochemical evaluation and on the 21st day for biomechanical evaluation. RESULTS: The hydroxyproline levels were found to be significantly increased in the treatment groups on the 10th and 21st days compared to their control groups (P < 0.05). In comparison of the treatment groups on the 4th, 10th, and 21st days of the treatment, the levels of tissue hydroxyproline were found to be more increased in combined US+L Group compared with US Group and L Group, but the difference was not significant (P > 0.05). In comparison of the tendon breaking strengths, it was found as significantly increased in the treatment groups compared with their control groups (P < 0.05), although there was no significant difference between the treatment groups. CONCLUSIONS: Although US, L, and combined US + L treatments increased tendon healing biochemically and biomechanically more than the control groups, no statistically significant difference was found between them. Also we did not find significantly more cumulative positive effects of combined treatment. As a result, both of these physical modalities can be used successfully in the treatment of tendon healing. PMID- 15278935 TI - Visualizing cell death in the developing heart. PMID- 15278936 TI - Interactive web-based programs to teach functional anatomy: the pterygopalatine fossa. AB - Certain areas of the body contain structures that are difficult to envision in their proper spatial orientations and whose functions are complex and difficult to grasp. This is especially true in the head, where many structures are relatively small and inaccessible. To address this problem, we are designing Web based programs that consist of high-resolution interactive bitmap illustrations, prepared using Adobe Photoshop, and vector-based animations, prepared via Macromedia Flash. Flash action script language is used for the animations. We have used this approach to prepare a program on the pterygopalatine fossa, an important neurovascular junction in the deep face that is especially difficult to approach by dissection and to depict in static images in an atlas. The program can be viewed online at http://cds.osr.columbia.edu/anatomy/ppfossa/. A table of contents simplifies navigation through the program and a menu enables the user to identify each of the vascular and neuronal components and either to insert or to remove each from its position in the fossa. The functional anatomy of the nerves in the fossa is animated. For example, users can activate and subsequently follow action potentials as they course along axons to their targets. This high degree of interactivity helps promote learning. PMID- 15278937 TI - Making tracks: the forensic analysis of footprints and footwear impressions. AB - Analysis of footwear characteristics, impressions, and track ways can provide important evidence in a crime scene investigation. In this article, we present examples of how students can be involved in hands-on laboratory-based activities as a means of introducing the forensic sciences. The teaching methodology employs active learning strategies that allow students to discover scientific principles for themselves, develop techniques of critical thinking and problem solving, and gain appreciation for how knowledge arises. By including forensic sciences in the science curriculum, students develop an appreciation for the interrelatedness of all the sciences. From this series of activities, i.e., examining analyses of footprint and footwear impressions, students working as teams will gather information, analyze data, and draw conclusions. Moreover, students will be able to assess the significance of the quality and variability in the data collection process as well as learn the value of controls and experimental design through comparison of results with other groups. PMID- 15278938 TI - Anxiety and dissection of the human cadaver: an unsolvable relationship? AB - Anxiety is an emotional reaction frequently shown by students when a human cadaver is being dissected. Nonetheless, few studies analyze the nature of the anxiety response in this situation and the ones that do exist are mainly limited to English-speaking countries. Our research has three aims: to study the characteristic anxiety reaction to dissection practices, to determine the weight exerted by internal and environmental variables on this anxiety reaction, and to design practices aimed at reducing the state of anxiety experienced by pupils in their human anatomy practices. The studies were carried out in the dissection room of the Department of Human Anatomy and Embryology II at the Faculty of Medicine of the Complutense University, Madrid, during the 3 academic years 2000 2003. The anxiety response to the first dissection of a human cadaver is mainly determined by a situation considered to be threatening, with novelty as its main characteristic. The students' anxiety response is first determined by the situation itself and reactions depend on individual differences. Repeated or gradual exposure (detailed verbal information on the situation, visits to dissecting rooms when no cadaver is present, videos showing pictures of human dissections, etc.) before carrying out the first dissection reduce the students' anxiety response. PMID- 15278939 TI - Microsmatic primates: reconsidering how and when size matters. AB - The terms "microsmatic" and "macrosmatic" refer to species with lesser or greater levels, respectively, of olfactory function. Historically, primates are considered microsmats (olfactory sense reduced) with a concomitant increased emphasis on vision. The olfactory bulbs (forebrain centers that receive peripheral olfactory input) are proportionately smaller in primates compared to most other mammals. Similarly, the regions of the nasal cavity that are covered with olfactory epithelium (containing receptor cells) have proportionately less surface area in primates than other mammals. Thus, the generalization that primates are microsmatic is most frequently stated in terms of the proportional rather than absolute size of olfactory structures. Yet the importance of scaling to body size is unclear in regard to the chemical senses such as the olfactory or vomeronasal systems-do chemosensory structures such as olfactory bulbs and olfactory epithelium exhibit the same neural relationship to body mass that is seen for neural tissues that supply innervation to musculature or the skin? Previous studies examining neuronal density, volume, and/or surface area of the olfactory epithelium illustrate that different conclusions may be supported based on the parameter used. Plots of olfactory bulb volume versus body mass that generated for large-scale taxonomic studies or growth studies benefit from body mass (or total brain volume) with a comparative perspective. However, our examination of proportional versus absolute measurements implies that in comparisons within taxa, body size adjustments needlessly distort the data. As a final consideration, another embryonic derivative of the nasal placode, the vomeronasal organ, may warrant consideration regarding a definition of microsomia versus macrosomia. PMID- 15278940 TI - On the reliability of recent tests of the Out of Africa hypothesis for modern human origins. AB - In this paper we critique two recent studies that have been claimed to disprove the Out of Africa hypothesis for modern human origins (Hawks et al., 2000; Wolpoff et al., 2001). We show that the test prediction employed by Hawks et al. (2000) and Wolpff et al. (2001) is not relevant to many versions of the Out of Africa hypothesis, and that the key specimens they used are problematic in terms of morphological representativeness. We also show that there are significant problems with the character state datasets employed in the studies. Lastly, we highlight evidence that the main method used in the studies (pairwise difference analysis) is not reliable when applied to the type of data employed by Hawks et al. (2000) and Wolpoff et al. (2001). In view of the foregoing, we contend that Hawks et al.'s (2000) and Wolpoff et al.'s (2001) claim to have disproved the Out of Africa hypothesis cannot be sustained. PMID- 15278941 TI - Subectodermal microfibrillar bundles are organized into a distinct parallel array in the developing chick limb bud. AB - In this study, a unique fiber system in the subectodermal mesenchyme of the chick limb bud was visualized immunohistochemically with the use of a novel monoclonal antibody termed "FB1." This antibody stained a subset of extracellular fibers in the embryonic mesenchyme. Among the fibers visualized, those running perpendicularly to the limb bud ectoderm became progressively prominent in their thickness and length, and organized into a parallel array in the subectodermal region. This fiber system was distinct from that of major collagens, fibronectin, or tenascin. A molecule immunoprecipitated with FB1 comigrated with JB3 antigen, or chicken fibrillin-2. The fibers visualized immunohistochemically by FB1 and JB3 were indistinguishable from each other, and ultrastructurally appeared to be bundles composed of tubular-like microfibrils that originated directly from the ectodermal basal lamina. They lacked the amorphous deposits that are characteristic of elastin. A similar array of subectodermal fibers was also found in the developing axilla and some truncal regions, again well before the development of a definitive dermis. These findings suggest that a parallel array of subectodermal FB1-positive fibers constitutes a precocious fiber system in the presumptive dermis prior to the substantial formation of collagenous fibers. These fibers could be developmentally linked to oxytalan fibers, which are known to be present in the papillary dermis in mature cutaneous tissue. PMID- 15278942 TI - Age-related expression patterns of Bag-1 and Bcl-2 in growth plate and articular chondrocytes. AB - Aging cartilage displays increased chondrocyte apoptosis and decreased responsiveness of chondrocytes to growth factors. The molecular mechanisms responsible for these changes have not been identified. Bag-1 is a Bcl-2-binding protein that promotes cell survival, interacts with a diverse group of cellular proteins, and may integrate multiple pathways involved in controlling cell survival, growth, and phenotype. Bcl-2 is important for maintaining chondrocyte phenotype and delaying terminal differentiation and apoptosis of chondrocytes. Comparatively little is known about the role of Bag-1 in cartilage. Here we show that both growth plate and articular chondrocytes in the mouse express the Bag-1 protein. In the growth plate, Bag-1 expression is prominent in the late proliferative and prehypertrophic chondrocytes, displaying a pattern similar to what has been reported for Bcl-2. Further, the expression of both Bcl-2 and Bag-1 declines with age in the articular cartilage. Growth assays demonstrate that knocking down Bag-1 expression causes a decrease in growth rate. These results suggest that Bag-1 is involved in the regulation of chondrocyte phenotype and cartilage aging. PMID- 15278943 TI - Expression of the Ellis-van Creveld (Evc) gene in the rat tibial growth plate. AB - Ellis-van Creveld (EvC) syndrome is an autosomal recessive chondrodysplasia characterized by short limbs, postaxial polydactyly, natal teeth, and dysplastic nails. The Ellis-van Creveld (EVC) gene, which is mutated in patients with EvC syndrome, has been identified by positional cloning. However, the physiological roles of the EVC gene have not been elucidated. Histopathological analyses of EvC syndrome have shown disturbed chondrocytic phenotypes during cartilage development. We therefore postulated that the EVC gene is a critical factor for chondrocytes during endochondral ossification. The present study focuses on the relationship between the Evc gene and chondrocytes, and examines Evc gene expression in the rat tibial growth plate at the mRNA and protein levels. Evc mRNA in tibial epiphyseal cartilage was expressed at postnatal day (P) 1, P28, and P56 by RT-PCR. Immunohistochemical analyses localized the Evc protein mainly in prehypertrophic and hypertrophic chondrocytes of the epiphyseal growth plate in the tibia during the embryonic and postnatal periods. Evc mRNA was also detected in prehypertrophic and hypertrophic chondrocytes by in situ hybridization. These results indicate that the Evc gene functions mainly in the prehypertrophic and hypertrophic chondrocytes of the epiphyseal growth plate. The data presented here are important for future studies of the underlying mechanism of chondrodysplasia in EvC syndrome. PMID- 15278944 TI - Remodeling capacity of calcified fibrocartilage of the hip. AB - It is well known that the incidence of hip fractures increases exponentially with age and that hip fractures can be a major cause of morbidity and morality among elderly humans; this has prompted substantial research on hip fractures. The majority of research on hip fractures has focused on morphological changes of the proximal femur with age. Recently calcified fibrocartilage in the proximal femur has been shown to increase in fractional area with age and can ultimately make up to 60% of the fractional area of the cortex. However, the capacity of the tissue to remodel and heal is currently unknown. The purpose of the present study was to determine the remodeling capacity of calcified fibrocartilage on the proximal femur compared to the underlying cortical bone. The remodeling capacity of calcified fibrocartilage and cortical bone was assessed in adult female sheep by means of tetracycline labeling. The number of double and single labels within each tissue type was quantified and analyzed with a paired t-test. The data showed very few labels in the calcified fibrocartilage compared to the cortical bone. This indicated that calcified fibrocartilage lacked a capacity to remodel in a manner similar to bone. The results of this investigation demonstrate that while the underlying cortical bone was actively remodeling, the calcified fibrocartilage appeared to have no remodeling capacity similar to that of cortical bone. PMID- 15278945 TI - Changing intracellular compartmentalization of beta-galactosidase in the ROSA26 reporter mouse during embryonic development: a light- and electron-microscopic study. AB - The beta-geo (LacZ) reporter gene encodes for beta-galactosidase (beta-gal) in all cells of the ROSA26 mouse during embryonic development. As such, beta-gal activity constitutes an excellent marker for in situ labeling of expressing cells. However, the intracellular distribution of beta-gal differs between cells, and changes during embryonic development. Therefore, we studied LacZ-encoded beta gal using light and electron microscopy in the heart, lung, liver, and small intestine on days 13 and 16 of gestation, and the kidney on day 16 of gestation in ROSA26 mice. The Bluo-gal method was carried out under standardized conditions, including fixation, washing, and incubation procedures. Intracellular beta-gal staining is encountered in a combination of membranous compartments, including the nuclear envelope, the endoplasmic reticulum, and the plasma membrane. Its exact localization depends on the cell type and is regulated during development. Therefore, one must take the compartmental transition of intracellular beta-gal staining into consideration when interpreting results obtained from experiments using ROSA26 mice. PMID- 15278947 TI - Spatiotemporal distribution of apoptosis during normal cloacal development in mice. AB - To understand normal cloacal developmental processes, serial sagittal sections of mouse embryos were made every 6 hrs from embryonic day 11.5 (E11.5) to E13.5. During cloacal development to form the urogenital sinus and anorectal canal, fusion of the urorectal septum with the cloacal membrane was not observed, and the ventral and dorsal parts of the cloaca were continuously connected by the canal until disappearance of the cloacal membrane to open the vestibule formed by the urogenital sinus and anorectal canal to the outside at E13.5. Ventral shifting of the dorsal part of the cloaca was observed until E12.5. The dorsal part was transformed in accordance with ventral shifting. In addition, apoptosis was seen to occur around the dorsal part. However, from E12.25, apoptotic cells are observed in a linear arrangement in the urorectal septum just ventral to the peritoneal cavity. Interestingly, extension of this line reaches the area of the cloacal membrane disintegrated by apoptosis. The present findings suggest that in the early stages (until E12.0), distribution of apoptosis in mesenchyme around the dorsal part of the cloaca might be strongly related to the transformation and ventral shifting of this part. Conversely, the apoptosis pattern in urorectal septum mesenchyme in later stages (from E12.0) might be involved in transformation of the urorectal septum and disintegration of the cloacal membrane. PMID- 15278946 TI - Adult human mylohyoid muscle fibers express slow-tonic, alpha-cardiac, and developmental myosin heavy-chain isoforms. AB - Some adult cranial muscles have been reported to contain unusual myosin heavy chain (MHC) isoforms (i.e., slow-tonic, alpha-cardiac, embryonic, and neonatal), which exhibit distinct contractile properties. In this study, adult human mylohyoid (MH) muscles obtained from autopsies were investigated to detect the unusual MHC isoforms. For comparison, the biceps brachii and masseter muscles of the same subjects were also examined. Serial cross-sections from the muscles studied were incubated with a panel of isoform-specific anti-MHC monoclonal antibodies that distinguish major and unusual MHC isoforms. On average, the slow type I and fast type II MHC-containing fibers in the MH muscle accounted for 54% and 46% of the fibers, respectively. In contrast to limb and trunk muscles, the adult human MH muscle was characterized by a large proportion of hybrid fibers (85%) and a small percentage of pure fibers (15%; P < 0.01). Of the fast fiber types, the proportion of the type IIa MHC-containing fibers (92%) was much greater than that of the type IIx MHC-containing fibers (8%; P < 0.01). Our data demonstrated that the adult human MH fibers expressed the unusual MHC isoforms that were also identified in the masseter, but not in the biceps brachii. These isoforms were demonstrated by immunocytochemistry and confirmed by electrophoretic immunoblotting. Fiber-to-fiber comparisons showed that the unusual MHC isoforms were coexpressed with the major MHC isoforms (i.e., MHCI, IIa, and IIx), thus forming various major/unusual (or m/u) MHC hybrid fiber types. Interestingly, the unusual MHC isoforms were expressed in a fiber type specific manner. The slow-tonic and alpha-cardiac MHC isoforms were coexpressed predominantly with slow type I MHC isoform, whereas the developmental MHC isoforms (i.e., embryonic and neonatal) coexisted primarily with fast type IIa MHC isoform. There were no MH fibers that expressed exclusively unusual MHC isoforms. Approximately 81% of the slow type I MHC-containing fibers expressed slow-tonic and alpha-cardiac MHC isoforms, whereas 80% of the fast type IIa MHC containing fibers expressed neonatal MHC isoform. The m/u hybrid fibers (82% of the total fiber population) were found to constitute the predominant fiber types in the adult human MH muscle. At least seven m/u MHC hybrid fiber types were identified in the adult human MH muscle. The most common m/u hybrid fiber types were found to be the MHCI/slow-tonic/alpha-cardiac and MHCIIa/neonatal, which accounted for 39% and 33% of the total fiber population, respectively. The multiplicity of MHC isoforms in the adult MH fibers is believed to be related to embryonic origin, innervation pattern, and unique functional requirements. PMID- 15278948 TI - Localization of estrogen and androgen receptors in male reproductive tissues of mice and rats. AB - Using immunohistochemical methods, we studied the cell-type- and species-specific expressions of estrogen receptor (ER) isoforms (ER alpha and ER beta) and androgen receptors (ARs) in the male reproductive tract and accessory sex glands of mature mice and rats. ER alpha and ER beta showed cell-type- and species specific distributions, respectively. In contrast, AR was localized in the epithelial and stroma cells of all tissues examined in this study, in both species. In mice, the epithelial cells of the ductuli efferentes showed a strong ER alpha-immunoreaction, and those of the caput epididymis, coagulating glands, and prostate also exhibited a positive reaction. Stroma cells, except in the ductuli efferentes, showed a positive ER alpha-immunostaining. In rats, ER alpha was detected in very few cell types: the epithelial cells of the ductuli efferentes showed a strong reaction, and the stroma cells of the ampullary and urethral glands exhibited a weak reaction. ER beta was localized in the epithelial cells of the prostate in mice, while the reaction was faint or negative in both the epithelial and stroma cells of other tissues. In rats, the ER beta-immunoreaction was strongest in the epithelial cells of the ventral prostate. The epithelial cells of the corpus and cauda epididymis, ductus deferens, and urethral glands, and the stroma cells of the urethral glands were also positively ER beta-immunostained. Almost the same AR distribution pattern was observed in both species. In particular, strong AR-immunostaining was present in the epithelial cells of the caput and corpus epididymis, seminal vesicle, and ventral prostate. These results indicate that species and tissues differences should be taken into careful consideration in assessing the physiological and pharmacological effects of sex steroids (particularly estrogens) on the reproductive tissues of male rodents. PMID- 15278949 TI - Cementum-like tissue deposition on the resorbed enamel surface of human deciduous teeth prior to shedding. AB - Prior to the shedding of human deciduous teeth, odontoclastic resorption takes place at the pulpal surface of the coronal dentin, and this resorption occasionally extends coronally from the dentinoenamel junction into the enamel. After the end of resorption, however, the resorbed enamel surface is repaired by the deposition of a cementum-like tissue. Using this phenomenon as an observation model, in this study we examined the sequence of cellular and extracellular/matrix events involved in the enamel resorption repair by light and electron microscopy. As the odontoclast terminated its resorption activity, it detached from the resorbed enamel surface; thereafter, numerous mononuclear cells were observed along the resorbed enamel surface. Most of these mononuclear cells made close contact with the resorbed enamel surface, and coated pits or patches were observed on their plasma membrane facing this surface. Furthermore, they frequently contained thin needle- or plate-like enamel crystals in their cytoplasmic vacuoles as well as secondary lysozomes. Following the disappearance of these monononuclear cells, the resorbed enamel surface now displayed a thin coat of organic matrix. Ultrastructurally, this organic layer was composed of a reticular and/or granular organic matrix, but contained no collagen fibrils. Energy-dispersive X-ray microanalysis of this thin organic layer in undecalcified sections revealed small spectral peaks of Ca and P. Cementum-like tissue initially formed along this thin organic layer, increased in width, and appeared to undergo mineralization as time progressed. The results of our observations demonstrate that regardless of type of matrix of dental hard tissues, tooth repair may be coupled to tooth resorption, and suggest that mononuclear cells and an organic thin layer found on the previously resorbed enamel surface may play an important role in the repair process initiated after resorption of the enamel. PMID- 15278950 TI - Alveolar bone Sharpey fibers of the rat incisor in normal and altered functional conditions examined by scanning electron microscopy. AB - The morphology and the area density of Sharpey fibers in the socket of the rat incisor under normo-, hyper-, and hypofunctional conditions were evaluated using scanning electron microscopy. Sharpey fibers appeared either as dome-shaped projections, when highly mineralized, or as depressions when less mineralized. Near the alveolar crest, most of the fibers were fully mineralized and arranged in compact longitudinal rows. Toward the basal end of the socket, the rows became interrupted, forming islets of gradually smaller size and number. The density of the Sharpey fibers was higher (P < 0.01) in the mesial and distal faces than in the lingual face in most of the socket length. In normofunctional conditions, in all faces the density decreased 70 to 90 times from the crestal toward the basal region of the socket (P < 0.01). The degree of mineralization of the Sharpey fibers also decreased steadily in the same direction, indicating that, for support, the periodontal ligament matures from basal to incisal and is fully developed only in the crestal region. In hyper- and hypofunctional conditions, the same distribution was observed. The area density of the Sharpey fibers in the hyperfunctional condition showed a slight increase at the basal levels of the socket mesial and distal faces (P < 0.01 or P < 0.05). In hypofunctional incisors, the density decreased significantly (P < 0.01) at the mesial and distal faces in all regions of the socket, and at the lingual face, the decrease (P < 0.05) was restricted to the incisal regions. This may be one of the factors for the weakening of the periodontal ligament in hypofunctional incisors. PMID- 15278951 TI - Distribution of calcitonin gene-related peptide at the neuromuscular junction of mdx mice. AB - In normal skeletal muscle, the protein dystrophin is associated with plasma membrane glycoproteins and may be involved in the stabilization of the sarcolemma. Mutant mdx mice are markedly deficient in dystrophin and show muscle fiber necrosis followed by regeneration. Changes in the distribution of acetylcholine receptors (AChRs) have been reported at the neuromuscular junction of mdx mice possibly as a result of alterations in the release or response to neural trophic factors. One such factor is calcitonin gene-related peptide (CGRP), which has been implicated in AChR synthesis and function. In this study, we used rhodamine-alpha-bungarotoxin and anti-CGRP IgG FITC to study AChR and CGRP distribution at the neuromuscular junction of mdx mice. Using laser scanning fluorescence confocal microscopy, it was possible to see that CGRP-like immunoreactivity had a presynaptic distribution, covering the AChRs. Thirty-four percent of dystrophic junctions were found to be labeled with CGRP compared to 80% of control endplates. Since CGRP-positive and -negative fibers showed similar changes in AChR distribution, it is suggested that CGRP is probably not directly involved in the altered pattern of AChR seen in dystrophin-deficient muscle fibers of mdx mice. PMID- 15278952 TI - A caution regarding U-500 insulin by continuous subcutaneous infusion. PMID- 15278953 TI - Free cortisol and critically ill patients. PMID- 15278954 TI - Free cortisol and critically ill patients. PMID- 15278955 TI - Free cortisol and critically ill patients. PMID- 15278956 TI - Ductal carcinoma in situ of the breast. PMID- 15278957 TI - Ductal carcinoma in situ of the breast. PMID- 15278958 TI - Ductal carcinoma in situ of the breast. PMID- 15278959 TI - Beethoven's deafness. 1958. PMID- 15278960 TI - Autoimmune sensorineural hearing loss. 1979. PMID- 15278961 TI - Editorial: How to write a scientific paper. 1989. PMID- 15278962 TI - Healthcare workers sentenced to death in Libya. PMID- 15278963 TI - Call for volunteers for AIDS vaccine trials. PMID- 15278964 TI - Self-serving and other-serving: matters of trust and intent. PMID- 15278965 TI - Nursing shortage or nursing famine: looking beyond numbers? PMID- 15278966 TI - Conceptual models of nursing: international in scope and substance? The case of the Roy adaptation model. PMID- 15278967 TI - Bibliography. Current world literature. Renal pathophysiology. PMID- 15278968 TI - Findings of scientific misconduct. PMID- 15278969 TI - Reactions vary to Pope's comments on feeding. PMID- 15278970 TI - The whole truth: drug companies must tell us all they know about the medicines we take. PMID- 15278972 TI - Abstracts of the Specialist Registrars in Obstetrics and Gynaecology meeting, Belfast, June 2003. PMID- 15278971 TI - Gene privacy for Icelanders. PMID- 15278973 TI - Publish or be damned. PMID- 15278974 TI - Abstracts from the 2nd Mediterranean Emergency Medicine Congress. Sitges, Spain, September 13-17, 2003. PMID- 15278975 TI - Nuchal translucency screening and informed consent. PMID- 15278976 TI - Electroconvulsive treatment, neurology, and psychiatry. PMID- 15278978 TI - A critique of medical coercive psychiatry, and an invitation to dialogue. AB - The medical model is the dominant ideology of psychiatry. It is used to justify involuntary institutionalization, forced drugging, and electroshock. By medical model is meant the medicalization of human thoughts, feelings, and behaviors. The medical model is an ideology because it serves the interests of and justifies the public demand for a covert form of social control. Psychiatry, the government, and the pharmaceutical industry have formed a mutually compatible view of human problems and their solutions, a "gleichschalten"--a marching in step. In their common view, certain human problems are psychiatric illnesses which, like other medical illnesses, are biological but, unlike other medical illnesses, can be treated with confinement and coercion. This ideology is accepted and adopted by the public without critical examination or self-reflection. Critical examination of the medical model has been systematically repressed by academic and organized psychiatry and their supporters. Recently, however, an increasing number of people are questioning the medical model. They are doubtful about the validity and social utility of explaining an expanding category of human problems caused by errant brain chemistry for which persons bear no responsibility and for which the treatment is pharmaceutical drugs. They are rising in protest against psychiatric repression, oppression and abuse. This essay presents 10 points of criticism of medical coercive psychiatry. It was first presented as the accusation at the Foucault Tribunal in Berlin in 1998. It has been reframed in this essay as an invitation to medical coercive psychiatry to dialogue with its critics. PMID- 15278977 TI - Pharmaceutical industry agenda setting in mental health policies. AB - The development of political agenda-setting through the use of sophisticated public relations techniques is threatening to undermine the delicate balance of representative democracy. This has important ramifications for policies aimed at providing mental health services and the implementation of mental health laws. The principal agenda setters in this area are pharmaceutical companies with commercial reasons to promote public policies that expand the sales of their products. They have manufactured highly effective advocacy coalitions that incorporate front groups in order to set the policy agenda for mental health. However, policies tailored to their commercial purpose are not necessarily beneficial either for patients or the society at large. PMID- 15278979 TI - The ethics of informed parental consent to psychiatric drugging of children. AB - That so many children are taking prescribed psychiatric drugs means that millions of parents have agreed to this recommendation for their children. Most pertinent to this fact is the question whether these parents have been given the opportunity to make their decisions in accordance with the legal standard of informed consent which requires a physician to disclose sufficient information for a patient to make an "informed" decision about a proposed treatment. At the least, such legitimate consent entails an opportunity to evaluate knowledgeably the options available and the risks attendant upon each, without coercion. This article examines the ethics of informed consent in psychiatry, and extant practices in the field, especially in regard to the psychiatric drugging of school age children in the United States. The authors argue that informed consent is systematically violated in this domain, and present a look at what authentic informed consent for parents to decide about psychiatric drugs for their children would entail. PMID- 15278980 TI - [Third Congress of Cardiologists and Angiologists of Bosnia-Herzegovina with International Participation, Tuzla, Bosnia-Herzegovina 27-30 May 2004. Proceedings and abstracts]. PMID- 15278981 TI - The need for "ethical" human sciences and services. PMID- 15278982 TI - Empirical, ethical, and political perspectives on the use of methylphenidate. AB - The case for methylphenidate (MPH) (Ritalin) rests on claims that Attention Deficit Hyperactivity Disorder (ADHD) is a disorder, that ADHD is biologically rooted, that MPH is effective against ADHD, that nontreatment is developmentally hazardous, and that MPH is a relatively innocuous rather than dangerous treatment. It will be found that the case for MPH rests on the hope that ADHD is a biological disorder and on MPH's short-term suppression of disruptive behavior. The explosion of MPH use on this flimsy evidential base can be explained within a political economy analysis but is difficult to justify ethically. It suggests a disturbing reliance upon drugs to solve problems stemming from a combination of inadequate support for families in modern societies and educational systems pushed to do more with less. Reliance upon a psychoactive medication to make children "ready to learn" is not only unsupported by the evidence, it teaches children incorrect lessons about how to meet the stresses and challenges of life. PMID- 15278983 TI - A boy who stops taking stimulants for "ADHD": commentaries on a Pediatrics case study. AB - This article presents eight commentaries on a case study published in the journal Pediatrics in 2001. The case study, introduced in Pediatrics to highlight the issue of adolescents' compliance with drug treatment in a "high-prevalence neurobehavioral condition," briefly describes an adolescent boy who announces that he no longer needs the methylphenidate he was prescribed for ADHD and had been taking for the last five years. We invited experienced professionals from the disciplines of psychiatry, psychology, counseling, education, and occupational therapy to comment on the case and make recommendations to the adolescent boy in the case study, his family, and professionals. Their commentaries highlight issues rarely discussed in the mainstream literature, including: the extent to which ADHD is erroneously portrayed and vigorously managed as a disease; the lack of validity of the ADHD construct in adolescence; the widespread use of stimulants as performance-enhancing drugs; the need to respect an adolescent's gut instinct and developing decisions; the importance of family dynamics in ADHD-like situations; the need to ease stimulant withdrawal effects; and the human rights of children prescribed psychotropic drugs. PMID- 15278984 TI - Research into the origins and characteristics of unicorns: mental illness as the unicorn. AB - Basic research, particularly into the psychological and neurological underpinnings of schizophrenia and other "mental illnesses," is flawed because of its adherence to the ideology that unwanted, hard-to-understand behavior constitutes true medical illness. It is argued here that psychiatric diagnostic terms represent moral judgments rather than medical entities. By reducing experimental subjects to a moral label, and assuming that neurological differences associated with unwanted behavior are brain diseases, researchers fail to take into account the conscious experience, organization of self and self image, patterns of motivation, history and social contexts of their patients. The failure to consider the psychology of their subjects renders the results of these studies ambiguous and irrelevant for any uses except bolstering the biomedical model of psychiatry. PMID- 15278985 TI - Better never than late: peer review and the preservation of prejudice. AB - This article documents some difficulties authors face who challenge faulty research claims published in mainstream literature. Editors of "reputable journals" may react with stonewalling tactics that tend to enshrine these faulty results. A case in point is the mental test literature, which has long been beset with racist myths. In 1985, Arthur Jensen added a new myth, his "Spearman Hypothesis," which asserts that a positive correlation between White/Black means differences in scores on mental tests and the loadings of the first principal component confirms the existence of a general intelligence factor ("g"). It can be shown by mathematical and geometric deduction, by computer stimulation, and by reference to "real data," including Jensen's own, that the assertion is unwarranted, and that the relationship Jensen observed is an artifact that has nothing to do with ethnicity or "g." Nevertheless, it proved impossible for more than 12 years to record this challenge to Jensen's claims in any of the leading journals in psychology and statistics. Typically, their editors invoked arguments having nothing to do with the fundamental question of whether Jensen's claims are true or false. It is concluded that, in view of the transparent racist implications of these claims, such editorial policies--regardless of their motivations--contribute to the preservation of ethnic stereotypes and prejudice. PMID- 15278986 TI - Active and passive consent: a comparison of actual research with children. AB - Passive consent, which is ethically questionable, requires parents to sign and return a form if they refuse to allow their child to participate in research. Active consent requires parents to sign and return a form if they consent for their child to participate. To compare passive and active consent research projects, we evaluated 15 published examples (since 1995) of passive consent and the adjacent experimental article (active consent). Passive consent projects involved significantly higher response rates, more subjects, greater likelihood of being conducted in school rather than in clinical settings, but about the same age of participants as active consent projects. We recommend that: (a) Institutional Review Boards scrutinize all passive consent projects and consider whether the consent procedure is ethical for the research sample; (b) editors and reviewers examine all manuscripts for the consent procedure used; and (c) ethicists and researchers debate the appropriateness/ethics of passive consent. PMID- 15278987 TI - The origins of coercion in Assertive Community Treatment: a review of early publications from the special treatment unit of Mendota State Hospital. AB - This article argues that Assertive Community Treatment (ACT) is fundamentally and historically based on the uncritical, but societally well-accepted view, that medically justified coercion (punishment or unwanted treatment) is therapeutic. It documents this claim by reviewing the early professional history and the resultant publications of the inventors of ACT (originally known as Training in Community Living), consisting of psychiatrists, social workers, and psychologists who trained and worked during the 1960s through the 1980s, at Mendota State Hospital (eventually renamed Mendota Mental Health Institute) in Wisconsin. PMID- 15278988 TI - Electroshock: a crime against the spirit. AB - Since its introduction in 1938, electroshock (electroconvulsive treatment, ECT) has been to its proponents a blessing and to its critics a curse. The author, himself an insulin coma-electroshock survivor, sides with the critics arguing that ECT is inherently harmful and dehumanizing. To support his views, he cites findings and comments from the professional literature in four areas: brain damage, memory loss, death, and brainwashing. The author also presents seven reasons for the continuing use of ECT, including profitability, value as a reinforcer of the biological model of mental illness, the absence of informed consent, the procedure's function as a "treatment of next resort," government and media support, and the public's failure to hold psychiatrists accountable for their conduct. The author concludes the article with his poem "Aftermath." PMID- 15278989 TI - Screening for depression: preventive medicine or telemarketing? AB - In this editorial, the authors critically examine a recent recommendation by the United States Preventive Services Task Force that all adults visiting primary care practitioners be routinely "screened" for depression. Drug industry funding for depression screening campaigns, popular but unproven assertions that depression is a "biochemical disorder," and the banality of antidepressant prescribing are seen to motivate the proposal. The full report cited by the Task Force omits information on the lack of effectiveness of antidepressants compared to placebo and recognizes that the potential harms of screening have not been studied. PMID- 15278990 TI - Images in emergency medicine. High-altitude pulmonary edema (HAPE). PMID- 15278991 TI - Synchronized cardioversion of unstable supraventricular tachycardia resulting in ventricular fibrillation. PMID- 15278992 TI - SARS decision rule: who's a suspect? PMID- 15278993 TI - Out-of-hospital pediatric airway management. PMID- 15278994 TI - Diagnostic pitfall: nontraumatic spinal epidural hematoma mimicking a brainstem stroke. PMID- 15278995 TI - Propofol and pediatric sedation. PMID- 15278996 TI - Emergency department phlebotomist: a failed experiment. PMID- 15278997 TI - A strange bullet. PMID- 15278998 TI - International HIV vaccine trial under way in Africa and the USA. PMID- 15278999 TI - First candidate enrolled in new HIV vaccine trial. PMID- 15279000 TI - Oxygen 2002. Proceedings of the 10th Symposium on Underwater and Hyperbaric Physiology. La Jolla, California, USA, July 1-2, 2002. Symposium in honor of Dr. Christian J. Lambertsen. PMID- 15279001 TI - Prison's second death row. PMID- 15279002 TI - Thailand glimpses success. PMID- 15279004 TI - [Proceedings of the 1st National Meeting of Nephrology Residents. Spain]. PMID- 15279003 TI - Is involuntary outpatient commitment a remedy for community mental health service failure? AB - Involuntary outpatient commitment (IOC) statutes exist in response to disorganized community mental health service delivery and perceived treatment non compliance. These statutes attempt to force psychiatric patients to comply with outpatient mental health services. Mental health service consumers, providers, and advocates have increasingly questioned the necessity and legality of IOC. Credible research indicates that IOC does not substantially benefit consumers and may increase mental health deterioration. IOC has proven difficult to implement, enforce, and successfully measure. Rather than resorting to expanding coercive measures, mental health systems and policymakers must ensure provision of voluntary and accessible mental health services. Furthermore, IOC cannot be legally or ethically justified even if hypothetical research supporting its alleged effectiveness exists. This article summarizes influential and contradictory IOC research, explores legal issues, and proposes that providing voluntary consumer-driven services would reduce IOC usage and prevent criminalizing individuals experiencing serious emotional distress. PMID- 15279005 TI - The dilemma of early intervention: some problems in mental health screening and labeling. AB - Motivated in part by a desire to identify and manage individuals considered to be at risk for serious mental disorders, early intervention programs are becoming increasingly accepted by the medical community, increasingly supported by the pharmaceutical industry, and increasingly unchallenged by the public. This article examines some of the conceptual and practical difficulties associated with early, or preventive, mental health care. Examples are drawn from contemporary research as well as historical commentaries in the fields of industrial psychology, sociology, and education. The tone of the discussion is cautionary, but the goal is to encourage mental health professionals and consumers to consider the potential harmfulness of early intervention strategies before these programs are uncritically copied, expanded, or renewed. PMID- 15279006 TI - Genetics and antisocial behavior. AB - This commentary article reviews a recent meta-analysis of genetic influences on antisocial behavior by Rhee and Waldman (2002). The authors combined the results of 51 twin and adoption studies and concluded that antisocial behavior has an important genetic component. However, twin and adoption studies contain several methodological flaws and are subject to the confounding influence of environmental factors. Therefore, Rhee and Waldman's conclusions in favor of genetic influences are not supported by the evidence. Two additional topics are Rhee and Waldman's incorrect description of the heritability concept and their failure to discuss several German criminal twin studies published during the Nazi era. PMID- 15279007 TI - Biomedical cooptation of the psychological care and support continuum for severely distressed persons. AB - In this issue of Ethical Human Sciences and Services, Lehrman describes his vision, focusing on continuity of care, of a mental health care system for psychosis. His main argument is that the same psychiatrist should treat an individual patient whatever the settings (hospital, community) that the patient shifts through, and that the psychiatrist is the key treating professional and as such should manage and direct care over the continuum. While continuity of psychiatric care is desirable and indeed noncontroversial and feasible, psychiatric services are not central to the care continuum, and its psychiatric control will lead to even more biomedical orientation in patient care. This would undermine another key aspect of Lehrman's vision: a mental health system focused upon the whole person in interaction with the environment. The history of psychiatry shows that, as a medical specialty, psychiatry will inevitably focus upon the brain rather than the person. Psychiatric care could serve as adjunct to appropriate supports, services, and treatment, but if it determines them, they will merely become a downplayed accessory of biomedical (i.e., psychotropic drug) treatment. PMID- 15279008 TI - Psychiatry and politics: some preliminary considerations. AB - This article explores the usually invisible relationship of psychiatry and politics. It is argued that all human relationships can be understood in terms of the type of political interactions existing between citizens, and that these types include the anarchistic, authoritarian, and democratic. It is theorized that each type of politics is both the result and expression of specific forms of interpersonal and intrapersonal relationships. The author presents the opinion that psychiatry functions according to the authoritarian mode of politics and explores relational consequences of this praxis. The author concludes with a description of his personal, democratic alternative to current psychiatric politics. PMID- 15279009 TI - Call me antipsychiatry activist--not "consumer". AB - The author, a long-time human rights activist, explains why he calls himself an anti-psychiatry activist rather than a "mental health consumer." He believes that the latter term is nonsensical in the current mental health system, characterized by lack of consumer choice and an explosion of involuntary interventions. He is a member of People Against Coercive Treatment and the Ontario Coalition Against Poverty. PMID- 15279010 TI - The impact of the Food and Drug Administration Modernization Act on the recruitment of children for research. AB - This article argues that contrary to the claims made by research stakeholders in industry, academia and government, the shift in public policy since the enactment of the Food and Drug Administration Modernization Act (FDAMA) of 1997 and its financial incentives to industry to test drugs on children, has had a deleterious impact on children's dignity, health and welfare. Those lucrative incentives offered an opportunity to accelerate the pace of FDA approval for pediatric drug marketing. FDAMA resulted in a radical shift in federal policy to accommodate an expansion of pediatric trials. Children who are precluded from exercising a human adult's right to informed consent to research are increasingly sought as test subjects even when the trials offer no potential benefit for them. Prior to FDAMA children were protected under federal regulations that prohibited their recruitment for experiments that were not in their best interest. This article discusses eight cases and controversies demonstrating that children have been subjected to experiments that exposed them to pain, discomfort, and serious risks of harm. Babies have died testing a lethal heartburn drug; children have been subjected to "forced dose titration" in antidepressant drug trials that resulted in several suicide attempts. Toddlers are currently being subjected to methylphenidate dose tolerance tests without evidence of any pathological condition. Healthy teenagers are being exposed to antipsychotic drugs known to induce severe pathological side effects in speculative "schizophrenia prevention" experiments. PMID- 15279012 TI - Proceedings of the 8th International Congress of Therapeutic Drug Monitoring and Clinical Toxicology. September 7-11, 2003. PMID- 15279011 TI - Organizational ethics in managed behavioral health care: perspectives from executives and leaders. AB - Managed behavioral health care (MBHC) is frequently criticized on ethical grounds for the way it undermines classical ideals of professionalism in mental health and addiction treatment. There is an implied assumption that practitioners who are executives and leaders in MBHC companies have moved away from clinical ethics to the adoption of business and financial models. This qualitative study explores perceptions of organizational ethical issues from the point of view of leaders working in MBHC settings and how their perspectives contribute to our current schemas for analyzing the ethical complexities of MBHC. Twenty-seven participants from across the United States were interviewed using an interview guide that relied on open-ended questions and probes. Inquiry findings present four major themes and describe participant material in a way that enhances sensitivity and understanding to organizational ethics in MBHC and behavioral health services and research. PMID- 15279013 TI - Categorical diagnosis and a poetics of obligation: an ethical commentary on psychiatric diagnosis and treatment. AB - Contemporary psychiatric practice reflects the hegemony of the DSM model of human emotional distress. Algorythms are used to decontextualize complaints of distress and classify them into distinct categories of "disorders" based on the presence or absence of specific "symptoms." This is usually followed by the prescription of psychotropic drugs. Such a reductionistic perspective and its mechanical approach to diagnosis and treatment have far reaching epistemological and moral consequences. In this article I address the moral issue from the postmodern ethic articulated by philosopher and theologian John Caputo, which he calls a "poetics of obligation." This position argues for the restoration of respect for the singularity, uniqueness and complexity of each doctor-patient encounter. I further argue for a disempowering of medical model psychiatry and a retrieval of an eclectic holism in psychiatric discourse. PMID- 15279014 TI - [200th anniversary of the birth of Karl Rokitansky]. PMID- 15279015 TI - Abstracts of the International Society for Pharmacoeconomics and Outcomes Research (ISPOR) 9th Annual International Meeting. May 16-19, 2004, Arlington, Virginia, USA. PMID- 15279016 TI - Results of the Heart Protection Study. PMID- 15279017 TI - A subanalysis of the Captopril Prevention Project. PMID- 15279018 TI - Abstracts of papers presented at the XXXVI Nordic Meeting of Gastroenterology, the XXVII Nordic Meeting of Digestive Endoscopy, the XV Nordic Meeting of Gastrointestinal Motility, the Annual Endoscopy/Gastroenterology Nurses's/Assistants' Meeting. Oslo, Norway, 2-5 June 2004. PMID- 15279019 TI - Insulin pump therapy versus multiple daily injections. PMID- 15279020 TI - The quality of fat in the diet and its impact on insulin sensitivity. PMID- 15279021 TI - Nonsteroidal anti-inflammatory drugs for the prevention of Alzheimer's disease: a systematic review. AB - OBJECTIVE: Alzheimer's disease, the most prevalent dementia, is a prominent source of chronic illness in the elderly. Laboratory evidence suggests that nonsteroidal anti-inflammatory drugs (NSAIDs) might prevent the onset of Alzheimer's disease. Since the early 1990s numerous observational epidemiological studies have also investigated this possibility. The purpose of this meta analysis is to summarize and evaluate available evidence regarding exposure to nonaspirin NSAIDs and risk of Alzheimer's disease using meta-analyses of published studies. METHODS: A systematic search was conducted using Medline, Biological Abstracts, and the Cochrane Library for publications 1960 onwards. All cross-sectional, retrospective, or prospective observational studies of Alzheimer's disease in relation to NSAID exposure were included in the analysis. At least 2 of 4 independent reviewers characterized each study by source of data and design, including method of classifying exposure and outcome, and evaluated the studies for eligibility. Discrepancies were resolved by consensus of all 4 reviewers. RESULTS: Of 38 publications, 11 met the qualitative criteria for inclusion in the meta-analysis. For the 3 case-control and 4 cross-sectional studies, the combined risk estimate for development of Alzheimer's disease was 0.51 (95% Cl=0.40-0.66) for NSAID exposure. In the prospective studies, the estimate was 0.74 (95% Cl=0.62-0.89) for 4 studies reporting lifetime NSAID exposure and it was 0.42 (95% Cl=0.26-0.66) for the 3 studies reporting a duration of use of 2 or more years. CONCLUSIONS: Based on analysis of prospective and nonprospective studies, NSAID exposure was associated with decreased risk of Alzheimer's disease. An issue that requires further exploration in future trials or observational studies is the temporal relationship between NSAID exposure and protection against Alzheimer's disease. PMID- 15279022 TI - Abstracts of the 2004 ASGE (American Society for Gastrointestinal Endoscopy) Meeting, May 18-21. Orlando, Florida, USA. PMID- 15279023 TI - Linking toxicology to epidemiology: biomarkers and new technologies. Abstracts of the ICT-X Satellite Meeting on Molecular Biology. 8-10 July 2004, Haikko, Finland. PMID- 15279024 TI - Abstracts of the 26th ISCVS (International Society for Cardiovascular Surgery) World Congress. PMID- 15279025 TI - Abstracts of the XXXIX Annual Scientific Meeting of the Society for the Spontaneously Hypertensive Rat (SHR). June 27-28, 2003, Tokyo, Japan. PMID- 15279026 TI - Orthopaedic proceedings, 2003, 2004. PMID- 15279027 TI - Cardiostim 2004. Abstracts of the 14th World Congress in Cardiac Electrophysiology and Cardiac Techniques. Nice Acropolis, France, June 16-19, 2004. PMID- 15279028 TI - Coral bleaching: a potential biomarker of environmental stress. AB - Coral bleaching refers to the loss of symbiotic algae by host corals, or to the loss of pigmentation by the algae themselves, causing corals to appear white or "bleached." Some corals may regain algae or pigmentation and survive, but when bleaching is severe the host coral dies. Coral bleaching events have increased dramatically in the last two decades, and coral reefs throughout the world have been extensively degraded as a result. This article reviews coral bleaching for investigators working in the field of toxicology and environmental health, a group of scientists not normally exposed to this issue. Several environmental stressors have been correlated with bleaching, including fluctuations in sea surface temperatures and salinity, increased sedimentation, increased solar radiation, and contaminants such as oil and herbicides. Molecular mechanisms of bleaching are only beginning to be investigated and are thus far poorly understood. Toxicologists have the potential to make significant contributions toward understanding anthropogenic aspects of coral bleaching and elucidating molecular mechanisms of this important environmental problem. PMID- 15279029 TI - Amorphous silica: a review of health effects from inhalation exposure with particular reference to cancer. AB - Silicas and silicates are some of the most abundant compounds found naturally in the earth's crust. Excessive exposure to crystalline silicas can cause serious lung disease such as silicosis and has been associated with lung cancer in some studies, but the potential health effects of amorphous silicas (silicon dioxide without crystalline structure) have not been well studied. Results from animal studies of amorphous silicas, unlike those seen with crystalline silicas, have suggested limited and largely reversible cytotoxic and possibly fibrogenic effects associated with some forms, but data on cancer outcomes are scanty and for the most part negative. Epidemiologic investigations to date for any potential cancer risk are not informative because the effects of crystalline and amorphous silicas have not been separated. Any future epidemiologic study should attempt to clarify the health effects of amorphous silicas from those of crystalline silicas, particularly with regard to any potential for carcinogenicity. PMID- 15279030 TI - Reassessment of the carcinogenicity of polychlorinated biphenyls (PCBs). AB - The current policy for regulating polychlorinated biphenyls (PCBs) is based on one chronic bioassay that examined the carcinogenicity of a 60% chlorinated PCB (Norback & Weltman, 1985). All studies originally considered by the U.S. Environmental Protection Agency (EPA) for use in calculating a cancer slope factor (CSF) for PCBs were reevaluated and new CSFs calculated based on the results of a pathology reassessment (Moore et al., 1994). When studies of 60% chlorine PCBs from 3 different laboratories were compared, there was no scientific basis for selecting only 1 data set for deriving CSF estimates. Using a geometric mean to calculate a CSF based on all studies of PCBs with 60% chlorine replaces the current value of 7.7 (mg/kg/d)(-1) with a value of 1.9 (mg/kg/d)(-1). CSFs for PCBs containing less than 60% chlorine (54% and 42%) were less than 1.0 (mg/kg/d)(-1). Using a toxic equivalency factor (TEF) approach similar to that of 2,3,7,8-tetrachlorodibenzo-p-dioxin shows no correlation between toxic equivalent dose and CSFs, indicating that use of TEFs is not predictive of cancer potency for PCBs. Based on these findings, PCB cancer risk assessment policy would more closely reflect scientific data if (1) separate risk assessments were developed for each major PCB formulation and (2) all appropriate data were used when calculating cancer potency for PCB mixtures of 60% chlorine. PMID- 15279031 TI - Biological monitoring of exposure to chlorpyrifos-methyl by assay of urinary alkylphosphates and 3,5,6-trichloro-2-pyridinol. AB - The results of biological monitoring by assay of urinary 3,5,6-trichloro-2 pyridinol and alkylphosphates (DMP, DMTP) in groups of 9 and 2 workers exposed to chlorpyrifos-methyl during vine spraying and manual leaf thinning 5-11 d after spraying, respectively, are reported. The results are compared with those of a control group of 46 subjects not occupationally exposed to organophosphate insecticides. Significantly higher urinary excretion of metabolites (Mann-Whitney U-test) was found in both groups than in controls. Levels of 3,5,6-trichloro-2 pyridinol (mean +/- SD) were 15.9 + 10.6 nmol/g creatinine (n = 33) for controls, 92.4 + 162.5 nmol/g creatinine (n = 20) for manual workers, and 675.5 + 1110.8 nmol/g creatinine (n = 48) for workers spraying and mixing the insecticide. Levels of DMP (mean +/- SD) were 63.8 + 100.1 nmol/g creatinine (n = 42), 123.0 + 79.0 nmol/g creatinine (n = 20), and 577.2 + 1003.2 nmol/g creatinine (n = 61), respectively, for the same 3 groups. Levels of DMTP (mean +/- SD) were 153.4 + 164.4 nmol/g creatinine (n = 43), 489.3 + 288.3 nmol/g creatinine (n = 20), and 297.6 + 215.4 nmol/g creatinine (n = 61), respectively, for the same 3 groups. Good correlations were found between urinary excretion of 3,5,6-trichloro-2 pyridinol and DMP (r = .776 for manual workers; r = .775 for workers mixing and spraying the insecticide) or DMTP (r = .558 and r = .746, respectively for the same 2 groups). The peak of excretion of the three metabolites was found in urine samples collected the night after the spraying or leaf thinning operations. PMID- 15279032 TI - Characterization of methemoglobinemia induced by 3,5-xylidine in rats. AB - The objective of this study was to characterize the dose effect and kinetics of methemoglobinemia in rats following oral or intravenous administration of 3,5 xylidine (XYL). The first set of experiments involved the intravenous administration of 0.06, 0.12, 0.24, 0.48, or 0.60 mmol XYL/kg to groups of 3 rats each and the serial sampling of blood from the tail vein of individual animals for the determination of methemoglobin levels. An additional series of experiments involved the oral administration of 0.24, 0.48, 0.72, 0.96, 1.2, 1.8, 2.4, or 4.8 mmol XYL/kg and the serial sampling of blood for the determination of methemoglobin levels. The results showed a dose-dependent induction of methemoglobinemia by XYL in the rat, for both routes of administration. The maximal percent methemoglobin observed in the treated animals was 28.90 +/- 0.34% and 32.67 +/- 2.14% for the intravenous (0.6 mmol/kg) and oral (4.8 mmol/kg) routes, respectively. The dose levels of 0.06 mmol/kg (iv) and 0.96 mmol/kg (po) were the no-observable-adverse-effect levels with respect to XYL-induced methemoglobinemia in the rat. The dose-effect information on XYL-induced methemoglobinemia obtained in this study may be useful for the characterization of noncarcinogenic risks of acute human exposure to this chemical. PMID- 15279033 TI - Lymphocyte proliferative response and tissue distribution of methylmercury sulfide and chloride in exposed rats. AB - The immunotoxic effects and tissue distribution of different forms of methylmercury compounds were studied in rats. Methylmercury sulfide or methylmercury chloride was fed to rats at concentrations of 5 or 500 microg/L in drinking water for 8 wk. T-cell lymphocyte proliferative response to phytohemagglutinin (PHA) and determination of tissue distribution of mercury by gas chromatography using electron capture were assayed. Four different forms of mercury compounds were employed: MeHgS-, (MeHg)2S, (MeHg)3S+, and MeHgCl. Results indicated that exposure to methylmercury significantly enhanced lymphocyte responsiveness in most of the exposed groups at the low concentration of 5 microg/L, with the highest proliferative response (fourfold increase) in the MeHgCl group. At 500 microg/L, a significant decrease in the lymphocyte proliferative response was observed in the (MeHg)3S+ and MeHgCl groups; conversely, the MeHgS(-)- and (MeHg)2S-exposed animals had a modest increase of the lymphocyte proliferative response. The largest concentrations of all four mercury forms were detected in the kidney and spleen. The levels of mercury found in kidney, spleen, liver, brain, and testis were lower in the MeHgCl group than in those exposed to (MeHg)2S and (MeHg)3S+. These data indicate that the organ distribution of mercury and immune alteration may vary according to the chemical structure of the compound. This observation may have important implications in humans potentially exposed to low levels of methylmercury present in the environment, since the immune system plays an important regulatory role in the host-defense mechanisms. PMID- 15279034 TI - [Comparative botanical and phytochemical investigations on the Gentiana species native to Hungary]. AB - The aim of this work was to compare the three Gentiana species native to Hungary (Gentiana asclepiadea L., Gentiana pneumonanthe L. and Gentiana cruciata L.--all protected) based on their botanical and phytochemical properties. Furthermore we planned the exploration of the biodiversity of these species and the examination of the correlations of the variable elements observed to establish the importance of these factors. Morphological, histological and phytochemical investigations were made. A new analytical method was developed through the systematic optimization of sample preparation and chromatographic (HPLC-DAD) separation. The results of the morphological, histological and phytochemical investigations were analysed by multivariate exploratory techniques. Based on our results, the investigated species can be distinguished as they are strongly heterogenous regarding certain morphological, histological and phytochemical characteristics. We made the first study on the biodiversity of these Gentiana species in their whole habitat in Hungary. Based on the results of the investigations mentioned above, distinct interspecific variability was found, in addition some results showed significant intraspecific variability as well. Numerous correlations were revealed between the variable characteristics, for example significant relationship was found between the average gentiopicroside content of the underground parts of Gentiana cruciata on the biotopes and the slope of the biotope. The knowledge of this variability and these correlations can be a benefit during the introduction and the improvement of medicinal plants with high contents of agents. PMID- 15279035 TI - [Applications of metal ions and their complexes in medicine I]. AB - The "inorganic medical chemistry" is a rapidly developing field with enormous potential for applications, which offers new possibilities to the pharmaceutical industry. For example, the titanocene dichloride is already in clinical use, and antimetastatic activity of a range of Ru(III) complexes is also well established. There are ways to minimize the toxicity of Gd(III) complexes and therefore they can be safely injected as MRI contrast agents. The so called "ligand design" allows paramagnetic ions to be targeted to specific organs. Such designed ligands also enable the targeting of radiodiagnostic (99mTc) and radiotherapeutic (186Re) isotopes. There is a significant progress in understanding the coordination chemistry and biochemistry of metal ion(s) containing complexes such as Au antiarthritic and Bi antiulcer drugs. Further, currently developing areas include Mn (SOD mimics), V (insulin mimics), Ru (NO scavengers), Ln-based photosensitizers, metal-targeted organic agents and the Fe overload. The expanding knowledge of the role of metals in biochemistry is expected to provide scope for the design of new drugs in many other areas too, for example neuropharmaceutical and antiaffective agents. Progress in coordination chemistry is strongly dependent on understanding not only the thermodynamics of reactions, but also the kinetics of metal complexes under biologically relevant conditions. PMID- 15279036 TI - [Pathogenesis and treatment of Wilson's disease]. AB - Authors review the pathogenesis, symptoms and diagnosis of Wilson's disease. Wilson's disease or hepatolenticular degeneration is an autosomal recessive disorder. It is caused by defective hepatic excretion of copper. The disease is fatal without treatment. The prevention of severe permanent damage depends upon early recognition and diagnosis followed by appropriate lifelong anticopper treatment. The purpose of the therapy of Wilson's disease is to eliminate the copper by chelators (D-penicillamine, triethylene tetramine, ammonium tetrathiomolibdate) and to inhibit the absorption and accumulation of copper by zinc salts (zinc sulphate, zinc acetate, zinc gluconate). PMID- 15279037 TI - [Thermostability testing of iron(II) sulphate for formulation of drug product]. AB - The aim of the experimental study was to determine which iron(II) sulphate form, the monohydrate or the heptahydrate, is more suitable for formulation of a retard dosage form with melt technology, and to control the suitable compound from a thermostability aspect according to the ICH (International Conference on Harmonization). Iron(II) sulphate is susceptible to oxidation and a rise in temperature increases the rate of this redox change. The presence of Fe3+ is definitely harmful in the case of enteral administration. First, the activation energy of the oxidation process was quantified for both iron(II) sulphate forms. It was found that the monohydrate is oxidized four times more slowly than the monohydrate form. It follows from this that the monohydrate well resists short lived high temperature. The thermostability tests on the monohydrate showed that samples, stored at high temperature and high relative humidity for a long time, oxidized only slowly. On the basis of all these findings, the monohydrate form may be suggested for melt technology. PMID- 15279038 TI - [Investigation of the mechanism of cell membrane-active cyclic lipodepsipeptides compounds]. AB - The cyclic lipodepsipeptides produced by Pseudomonas syringae pv. syringae possess fungicide properties. They inhibit much of the cell functions, presumably on the basis of their pore-forming activity. The syringomycin E, studied earlier by our research group, formed pores on human red blood cells (RBC), and caused also partially hemolysis. To reach selective toxicity, knowledge of the relationship between the structure and function of the CLPs is required. To fulfill this requirement, the pore forming properties of two other CLPs, the syringopeptin22A (SP22A) and the syringotoxin (ST), were investigated on RBCs. On the basis of our transport kinetic measurements on RBC, the SP22A and the ST formed pores built up by few monomers. ST pores inactivated at 37 degrees C, and at 20 degrees C, however, they remained stable at 8 degrees C, similarly to the SRE. Inactivation of the SRE and ST pores was faster at 37 degrees C than at 20 degrees C. SP22A pores did not inactivate even at 37 degrees C. On the basis of our results, we concluded that fluidity has a crucial role in the process and the dimension of the hydrophobic part and the number of the positive charges of the polar headgroup influence the stability of the pores. To study the interactions of CLPs with membranes on molecular level we applied EPR spectroscopy and spectral simulation methods for liposomes of different composition. CLPs decreased the fluidity at all depths of the membrane: the motional and rotational freedom of the lipid molecules decreased, while their ordering increased. These observations in conjunction with the concentration dependence of the CLP's action suggested that CLPs form pores involving the lipids, too. Investigation of the temperature dependence of the CLPs' action showed that to get their complete effect a given temperature range is required. It corroborated that the fluidity has a crucial role in the interactions between CLPs and membrane lipids. From the results, obtained with DPPC-DOPC or DPPC-cholesterol liposomes we concluded that the membrane-inhomogeneity decreases the effectiveness of the CLPs. Dynamic light scattering measurements were used to determine the size-distribution of the liposomes. According to our experiences CLPs provoke fusion and/or aggregation, which can have a role in the cell-killing properties of the CLPs. PMID- 15279039 TI - [Interview with Professor Tivadar Tulassay, the member of the Hungarian Academy of Sciences, the Rector of Semmelweis University]. PMID- 15279040 TI - The Schizophrenia Patient Outcomes Research Team (PORT): updated treatment recommendations 2003. AB - Since publication of the original Schizophrenia Patient Outcomes Research Team (PORT) treatment recommendations in 1998, considerable scientific advances have occurred in our knowledge about how to help persons with schizophrenia. Today an even stronger body of research supports the scientific basis of treatment. This evidence, taken in its entirety, points to the value of treatment approaches combining medications with psychosocial treatments, including psychological interventions, family interventions, supported employment, assertive community treatment, and skills training. The most significant advances lie in the increased options for pharmacotherapy, with the introduction of second generation antipsychotic medications, and greater confidence and specificity in the application of psychosocial interventions. Currently available treatment technologies, when appropriately applied and accessible, should provide most patients with significant relief from psychotic symptoms and improved opportunities to lead more fulfilling lives in the community. Nonetheless, major challenges remain, including the need for (1) better knowledge about the underlying etiologies of the neurocognitive impairments and deficit symptoms that account for much of the disability still associated with schizophrenia; (2) treatments that more directly address functional impairments and that promote recovery; and (3) approaches that facilitate access to scientifically based treatments for patients, the vast majority of whom currently do not have such access. PMID- 15279041 TI - Differential effectiveness of clozapine for patients nonresponsive to or intolerant of first generation antipsychotic medications. AB - This report examines whether the gains associated with changing to clozapine are greater for people who have been intolerant of first generation antipsychotic medications versus those who have been treatment-nonresponsive to previous agents. We examined data from an open-label, randomized trial that compared clozapine to usual care with first generation agents (n = 227). While most patients (n = 173, 76%) entered that study because they were nonresponsive to at least two first generation antipsychotic medications (treatment nonresponsive [TNR]), 24 percent (n = 54) were eligible because they experienced intolerable side effects (treatment intolerant [TI]). Significantly more TI patients discontinued their clozapine trial during the 2-year study compared to TNR patients, and TI patients taking clozapine were more likely to develop agranulocytosis or severe leukopenia. However, TI patients who remained on clozapine showed significant reductions in problematic behaviors and greater movement toward independent living situations than TNR patients. Clinicians should give serious consideration to offering clozapine and other second generation antipsychotic medications to patients who have demonstrated intolerance to first generation antipsychotic medications. PMID- 15279042 TI - Weight gain with clozapine compared to first generation antipsychotic medications. AB - Few studies have examined gender differences in the propensity to gain weight on clozapine. Weight gain increases risk for many medical illnesses and is of particular concern for people with schizophrenia who are more overweight than the general population. Long-stay patients in Connecticut state hospitals were randomly assigned to switch to open-label treatment with clozapine (n = 138) or to continue receiving first generation (conventional) antipsychotic medications (n = 89). Using survival and random regression models, we examined percentage of body weight gained during 2 years for patients assigned to clozapine versus those who continued taking first generation antipsychotic medications. We also examined the impact of gender on weight gain. Patients who switched to clozapine gained a greater percentage of weight (13 pounds, 7%) than did patients remaining on first generation medications (5 pounds, 4%) at the end of 2 years. Normal-weight patients on clozapine were more likely to become obese (body mass index [BMI] > or = 30). Patients gained weight whether they switched to clozapine or remained on first generation antipsychotic medications, but weight gain was significantly greater (1 BMI unit) in the clozapine-treated group, particularly among women. PMID- 15279043 TI - Neurological soft signs and their relationship to cognitive and clinical efficacy of atypical antipsychotics in schizophrenia. AB - Neurological soft signs (NSS) are nonspecific indicators of brain dysfunction that are found to be in excess and correlated with cognitive dysfunction and psychopathology in patients with schizophrenia. The aim of the study was to examine whether the severity of NSS determines the efficacy of atypical antipsychotics in schizophrenia. Forty-three patients with schizophrenia were assessed on psychopathology and cognitive domains of executive functioning, memory, attention, and psychomotor speed at baseline and 6 months after they had been switched from typical to atypical antipsychotics. NSS were examined at baseline. The high-NSS group showed more severe psychopathology and greater impaired cognitive function than the low-NSS group at baseline. Following treatment, there were improvements in cognitive functioning and psychopathology with the low-NSS group, which showed significant improvements on measures of verbal fluency, memory, and psychomotor speed and negative symptoms. The high-NSS group also showed improvements on most of these measures, but the improvement was less than that seen in the low-NSS group. The presence of high NSS in schizophrenia patients impedes the improvement in cognitive function with atypical antipsychotics treatment. PMID- 15279044 TI - Poor antipsychotic adherence among patients with schizophrenia: medication and patient factors. AB - Many patients with schizophrenia are poorly adherent with antipsychotic medications. The newer, atypical antipsychotics may be more acceptable to patients and result in increased adherence. We used national Department of Veterans Affairs (VA) pharmacy data to examine whether patients receiving atypical agents are more adherent with their medication and explored patient factors associated with adherence. Patients who received a diagnosis of schizophrenia or schizoaffective disorder between October 1, 1998, and September 30, 1999, were identified in the VA National Psychosis Registry. We calculated medication possession ratios (MPRs) for patients filling prescriptions for one (n = 49,003) or two (n = 14,211) antipsychotics during the year. We examined cross sectional relationships among adherence, type of antipsychotic, and patient characteristics and explored adherence among patients switching antipsychotics during the year. Among patients receiving one antipsychotic, 40 percent had MPRs < 0.8, indicating poor adherence. African-Americans and younger patients were more likely to be poorly adherent. Cross-sectionally, patients on atypical agents were more likely to be poorly adherent (41.5%) than patients on conventional agents (37.8%). However, among a small group of patients switching from a conventional to an atypical agent (n = 1,661) during the year, the percentage who were poorly adherent decreased from 46 percent to 40 percent. We describe the continuum of antipsychotic adherence among a large sample of patients with schizophrenia and confirm that poor adherence is common. African-Americans and younger patients are particularly at risk. Unfortunately, atypical antipsychotics may not be associated with substantial improvements in adherence. More intensive interventions are likely needed. PMID- 15279045 TI - Premorbid functioning in schizophrenia: relation to baseline symptoms, treatment response, and medication side effects. AB - Impaired premorbid functioning prior to the onset of acute psychosis has frequently been noted in schizophrenia. This study examined retrospectively the premorbid status of patients in their first episode of psychosis in order to determine relationships with baseline symptoms, treatment response, and medication side effects. One hundred eleven schizophrenic and schizoaffective patients participating in a large prospective study of first episode schizophrenia were evaluated with the Premorbid Adjustment Scale (PAS). Premorbid functioning in males became progressively worse over time. Deficit state patients exhibited worse premorbid functioning. A third of patients exhibited sustained poor premorbid functioning. At various developmental stages, lower "sociability and withdrawal" scores correlated with increased time to treatment response, more severe negative symptoms, increased drug-induced parkinsonism, and deterioration of premorbid functioning. Various mean PAS scores predicted susceptibility to tardive dyskinesia. Our findings suggest that prior to acute psychosis onset there are certain behavioral precursors reflected in premorbid functioning that may predict subsequent illness manifestations. Measures of premorbid functioning indicate that disease pathogenesis is manifest, albeit more subtly, prior to presentation of first psychotic symptoms. PMID- 15279046 TI - The global costs of schizophrenia. AB - Schizophrenia is a chronic disease associated with a significant and long-lasting health, social, and financial burden, not only for patients but also for families, other caregivers, and the wider society. Many national and local studies have sought to estimate the societal burden of the illness--or some components of it--in monetary terms. Findings vary. We systematically reviewed the literature to locate all existing international estimates to date. Sixty-two relevant studies were found and summarized. Within- and between-country differences were analyzed descriptively. Despite the wide diversity of data sets and methods applied, all cost-of-illness estimates highlight the heavy societal burden of schizophrenia. Such information helps us to understand the health, health care, economic, and policy importance of schizophrenia, and to better interpret and explain the large within- and across-country differences that exist. PMID- 15279047 TI - Direct costs of schizophrenia and related disorders in Italian community mental health services: a multicenter, prospective 1-year followup study. AB - The behavior that accompanies schizophrenia and related disorders interferes with professional and social activities. As a result, schizophrenia is one of the most costly psychiatric illnesses. Direct medical costs associated with schizophrenia were estimated from the Italian National Health Service perspective. This was a multicenter observational 1-year study conducted in 14 Italian community mental health centers (CMHCs). Eligible patients were those with a diagnosis of schizophrenia or schizoaffective or schizophreniform disorder who had been followed by the CMHCs for at least 2 years at study entry. Exactly 643 patients were enrolled in the study. The mean direct cost per year was 6,964 (27,025 for schizophrenia and 6,587 for patients with related psychotic disorders) (1998 exchange rate U.S.$1 = 1.121). The present study provides further estimates of the cost of schizophrenia treatment in Italian mental health services and highlights the variability in the single cost components across clinically defined subgroups of patients. PMID- 15279048 TI - Offspring of parents with schizophrenia: mental disorders during childhood and adolescence. AB - Although offspring of parents with schizophrenia are at risk for schizophrenic illness as adults, little is known about their pattern of symptoms as children and adolescents. Lifetime Axis I and II DSM-III-R diagnoses were made for 116 young people (ages 12-22). Forty-one subjects had a parent with schizophrenia, 39 had a parent with a nonschizophrenic mental disorder, and 36 had parents with no mental illness. Schizophrenia spectrum disorders occurred at higher rates among young people with a parent with schizophrenia (17.1%) than in other young people (5.3%), even after controlling for mental disorder in the nonproband parent. Schizophrenia and schizotypal personality disorder occurred exclusively in offspring of parents with schizophrenia. Offspring of parents with schizophrenia were also at increased risk for avoidant personality disorder but not paranoid personality disorder. Although lifetime anxiety disorders were common in all young people regardless of parent diagnosis, current anxiety disorders were more prevalent for the adolescent offspring of parents with schizophrenia. These data strongly suggest familial vulnerability to schizophrenia spectrum disorder that is observable before adulthood, particularly for males. PMID- 15279049 TI - Multiple dimensions of schizotypy in first degree biological relatives of schizophrenia patients. AB - Considerable research has been devoted to identifying individuals predisposed to schizophrenia, with much of the effort devoted to identifying the personality characteristics of the biological relatives of schizophrenia patients. Although resource-consuming interviews have yielded promising results, investigators have long sought self-report measures that index genetic risk for schizophrenia. The Schizotypal Personality Questionnaire (SPQ) is a self-report measure that assesses the nine features of DSM-defined schizotypy. The SPQ, modified to include validity scales, was administered to 135 nonpsychotic first degree relatives of schizophrenia patients and 112 healthy controls. Principal components analysis (PCA) yielded three factors that correlated highly with previously reported factors (social-interpersonal, cognitive-perceptual, and disorganization). Social-interpersonal deficits were found to best differentiate relatives from controls. Contrary to the hypothesis that schizophrenia relatives are more defensive in responding to schizotypy questionnaires, relatives were significantly less defensive than controls. The results demonstrate that a multidimensional paper-and-pencil measure can characterize schizotypal features in schizophrenia relatives, which will be useful for the further delineation of the heritable schizophrenia spectrum phenotype. PMID- 15279050 TI - Investigating graphesthesia task performance in the biological relatives of schizophrenia patients. AB - This study compared the performance of 39 biological relatives of persons with schizophrenia to that of 30 normal adult controls on graphesthesia processing, a complex somatosensory processing task. The relatives performed significantly worse on the graphesthesia task compared to the healthy controls. The relatives and control subjects, however, did not differ on two neurocognitive control tasks. These data are interpreted within the context of a somatosensory deficit linked to schizophrenia liability. PMID- 15279051 TI - Factorial composition of self-rated schizotypal traits among young males undergoing military training. AB - The aim of this study within the Athens Study of Psychosis Proneness and Incidence of Schizophrenia (ASPIS) was the examination of the latent structure of schizotypal dimensions among a large population of young male conscripts in the Greek Air Force during their first week of military training. Confirmatory factor analysis (CFA) was conducted on 1,355 reliable responders to the self-rated Schizotypal Personality Questionnaire (SPQ), which covers all nine aspects of DSM III-R schizotypal personality disorder (SPD). A four-factor model (cognitive/perceptual, paranoid, negative, and disorganization schizotypal dimensions) provided a better fit to the data than did other competing models (one-, two-, three-, four, and five-factor models). This result is in agreement with recent findings supporting the notion of a multidimensional construct of the schizotypy and related schizophrenia phenotype. PMID- 15279052 TI - Earlier age of first diagnosis in schizophrenia is related to impaired motor control. AB - We examined the control of motor behavior in relation to age of first diagnosis (AFD; an approximation of age of onset) in schizophrenia. We hypothesized that earlier AFD reflects increased vulnerability to the disorder, vulnerability that may be indexed by elevated levels of motor abnormality. AFD, symptom and demographic features, motor performance on a line drawing task, and the presence and severity of dyskinesia and extrapyramidal side effects were evaluated in 65 chronic schizophrenia subjects. More severely impaired motor control was significantly related to an earlier age of diagnosis. Potential confounds, including age, gender, education, length of illness, current medication dosage, symptom status, and motor side effects, did not appear to influence this relationship, although greater chronicity appeared to be independently related to more severely impaired motor control. In summary, the data are consistent with the hypothesis that an earlier AFD is associated with more pronounced motor impairment. PMID- 15279053 TI - Discriminating value of total minor physical anomaly score on the Waldrop physical anomaly scale between schizophrenia patients and normal control subjects. AB - Minor physical anomalies (MPAs) are slight structural aberrations that are believed to be associated with abnormal neurodevelopment. Studies of schizophrenia patients show that these patients score higher in MPAs than normal controls. The present study attempted to assess the potential value of MPAs as a classifying test in the status schizophrenia patient versus normal control. Seventy-six schizophrenia patients and 82 normal controls were assessed for MPAs using the Waldrop Physical Anomaly Scale, and specificity, sensitivity, and predictive value of the total MPA score were determined. A significantly higher percentage of schizophrenia patients than normal controls had high numbers of MPAs. Total MPA scores higher than 4 showed the most balanced set of sensitivity (76.3%), specificity (72.0%), and positive (71.6%) and negative (76.6%) predictive values for schizophrenia and were the cutoff scores that optimally discriminate schizophrenia patients from normal controls. Schizophrenia patients showed a higher percentage of subjects with prominent MPA scores. The results are consistent with the hypothesis that MPAs might reflect extragenetic stressful events and present total MPA score as a reliable index in distinguishing between schizophrenia patients and normal controls. PMID- 15279054 TI - What is the functional significance of hippocampal pathology in schizophrenia? AB - The hippocampal formation (HF) is one of the brain structures most consistently altered in schizophrenia, yet the contribution of HF pathology to severe mental illness is poorly understood. We present evidence that our current ignorance is attributable to the fact that the anterior HF is heavily involved in schizophrenia but has been inadequately examined by schizophrenia investigators. We propose that the anterior HF in humans, and its counterpart in rodents (ventral HF), constrain diverse responses to psychological stimuli and that disruption of this function contributes to schizophrenia. While current data suggest that hallmark symptoms of schizophrenia most likely result from the role of the anterior HF in the integrated neurocircuit that includes the prefrontal cortex, ventral striatum, and ventral tegmental area, better characterized and phylogenetically preserved neurocircuits may be similarly affected by anterior HF pathology and account for associated findings of the disorder. We propose that focusing on the impact of ventral HF pathology on these simpler circuits and functions in rodents may provide insight into the pathophysiology of severe mental illness in humans. We review several associated findings in schizophrenia to assess the likelihood that each could be a product of this putative anterior HF dysfunction and could therefore be productively studied in rodents by probing ventral HF function. PMID- 15279055 TI - Multiple structural brain measures obtained by three-dimensional magnetic resonance imaging to distinguish between schizophrenia patients and normal subjects. AB - This study was designed to investigate the extent to which schizophrenia patients can be differentiated from normal subjects by structural brain measures. High resolution magnetic resonance imaging scans were performed on 57 schizophrenia patients (30 males, 27 females) and 47 normal controls (25 males, 22 females). Significant enlargements of the left and right body of the lateral ventricle, the left and right sylvian fissure, and the third ventricle were observed in the male patients. Significant enlargements of the left inferior horn, and the left and right sylvian fissure, and a significant volume reduction of the right temporal lobe were observed in the female patients. Discriminant function analysis using brain anatomical measures as variables allowed correct classification of 80.0 percent of the male patients, 80.0 percent of the male controls, 77.8 percent of the female patients, and 86.4 percent of the female controls. These findings support the view that schizophrenia patients have structural deviations in multiple brain areas and that a combination of structural brain measures can distinguish between patients and controls. PMID- 15279056 TI - Orbitofrontal cortical dysfunction in akinetic catatonia: a functional magnetic resonance imaging study during negative emotional stimulation. AB - Catatonia is a psychomotor syndrome characterized by concurrent emotional, behavioral, and motor anomalies. Pathophysiological mechanisms of psychomotor disturbances may be related to abnormal emotional-motor processing in prefrontal cortical networks. We therefore investigated prefrontal cortical activation and connectivity patterns during emotional-motor stimulation using functional magnetic resonance imaging (FMRI). We investigated 10 akinetic catatonic patients in a postacute state and compared them with 10 noncatatonic postacute psychiatric controls (age-, sex-, diagnosis-, and medication-matched) and 10 healthy controls. Positive and negative pictures from the International Affective Picture System were used for emotional stimulation. FMRI measurements covered the whole frontal lobe, activation signals in various frontal cortical regions were obtained, and functional connectivity between the different prefrontal cortical regions was investigated using structural equation modeling. Catatonic patients showed alterations in the orbitofrontal cortical activation pattern and in functional connectivity to the premotor cortex in negative and positive emotions compared to psychiatric and healthy controls. Catatonic behavioral and affective symptoms correlated significantly with orbitofrontal activity, whereas catatonic motor symptoms were rather related to medial prefrontal activity. It is concluded that catatonic symptoms may be closely related to dysfunction in the orbitofrontal cortex and consequent alteration in the prefrontal cortical network during emotional processing. Because we investigated postacute patients, orbitofrontal cortical alterations may be interpreted as a trait marker predisposing for development of catatonic syndrome in schizophrenic or affective psychosis. PMID- 15279057 TI - Left prefrontal repetitive transcranial magnetic stimulation in schizophrenia. AB - In a double-blind, controlled study, we examined the therapeutic effects of high frequency left prefrontal repetitive transcranial magnetic stimulation (rTMS) on schizophrenia symptoms. A total of 22 chronic hospitalized schizophrenia patients were randomly assigned to 2 weeks (10 sessions) of real or sham rTMS. rTMS was given with the following parameters: 20 trains of 5-second 10-Hz stimulation at 100 percent motor threshold, 30 seconds apart. Effects on positive and negative symptoms, self-reported symptoms, rough neuropsychological functioning, and hormones were assessed. Although there was a significant improvement in both groups in most of the symptom measures, no real differences were found between the groups. A decrease of more than 20 percent in the total PANSS score was found in 7 control subjects but only 1 subject from the real rTMS group. There was no change in hormone levels or neuropsychological functioning, measured by the MMSE, in either group. Left prefrontal rTMS (with the used parameters) seems to produce a significant nonspecific effect of the treatment procedure but no therapeutic effect in the most chronic and severely ill schizophrenia patients. PMID- 15279058 TI - Cerebrovascular response to cognitive tasks in patients with schizophrenia measured by near-infrared spectroscopy. AB - We assessed the cerebral hemoglobin oxygenation response in the left frontal area in 62 schizophrenia patients and 31 healthy subjects during a verbal fluency test (VF) and letter number span test (LN) measured by near-infrared spectroscopy (NIRS). Oxygenated hemoglobin (oxyHb) increased during VF and LN in both groups. Schizophrenia patients showed lower VF and LN performance and a smaller increase in oxyHb during VF than controls. A reduced oxyHb response during VF in schizophrenia patients was also observed even when their VF performance was matched with controls' performance. On the other hand, increase in oxyHb during LN in schizophrenia patients was comparable with that in controls. In addition, patients medicated with atypical antipsychotics showed a larger increase in oxyHb during VF and LN than those medicated with typical antipsychotics. The present study confirmed functional hypofrontality in schizophrenia patients reported by other modalities such as position emission tomography, single-photon emission tomography, and functional magnetic resonance imaging and suggested that the hypofrontality may be task dependent. PMID- 15279059 TI - At issue: siblings of patients with schizophrenia: sibling bond, coping patterns, and fear of possible schizophrenia heredity. AB - Siblings of schizophrenia patients are from the patient's perspective important support providers, but most studies on family burden have focused on the parental role. This study aims to develop a detailed analysis of the psychological aspects of having a sibling with schizophrenia. We did a qualitative study with audiotaped semistructured interviews of 16 siblings. The reliability of the inductive categorization of data was high. A unifying theme appeared to be an emotional sibling bond characterized by feelings of love, sorrow, anger, envy, guilt, and shame. The major categories linked to coping with the situation were avoidance, isolation, normalization, caregiving, and grieving. A third major theme consisted of a fear of possible schizophrenia heredity. The siblings described concerns about the impact of a family history of psychiatric illness, a fear of becoming mentally ill, and reflections about "bad genes." Our findings support earlier findings of coping patterns but complement them by providing a model that includes awareness of genetic vulnerability as an important part of siblings' subjective burden. PMID- 15279060 TI - Stages of change in smokers with schizophrenia or schizoaffective disorder and in the general population. AB - This study compared smoking behavior and motivation to quit smoking, assessed with a "stages of change" questionnaire, in outpatients with schizophrenia or schizoaffective disorder and in a representative sample of the general population. We conducted a mail survey in a representative sample of the general population of Geneva, Switzerland, in 1996 (n = 742); and a survey of 151 patients with schizophrenia (84%) or schizoaffective disorder (16%) who attended a Geneva ambulatory psychiatric clinic in 2000. There were more smokers (70% vs. 28%, p < 0.001) in patients with schizophrenia or schizoaffective disorder than in the general population, and fewer ex-smokers (15% vs. 52%, p < 0.001). Patients with schizophrenia or schizoaffective disorder smoked more than smokers in the general population (22 vs. 16 cigarettes per day, p < 0.001). Among current smokers, the distribution of stages of change was similar in patients with schizophrenia or schizoaffective disorder (precontemplation 79%, contemplation 18%, preparation 3%) and in the general population sample (74%, 22%, and 4%, p = 0.6). In both samples, similar proportions of smokers had made an attempt to quit in the previous year (27% vs. 22%, p = 0.3). These results suggest that a substantial minority of smokers with schizophrenia or schizoaffective disorder are motivated to quit smoking, try to quit, and succeed in quitting. PMID- 15279061 TI - First person account: how insight poetry helped me to overcome my illness. AB - The article that follows is part of the Schizophrenia Bulletin's ongoing First Person Account series. We hope that mental health professionals--the Bulletin's primary audience--will take this opportunity to learn about the issues and difficulties confronted by consumers of mental health care. In addition, we hope that these accounts will give patients and families a better sense of not being alone in confronting the problems that can be anticipated by persons with serious emotional difficulties. We welcome other contributions from patients, ex patients, or family members. Our major editorial requirement is that such contributions be clearly written and organized, and that a novel or unique aspect of schizophrenia be described, with special emphasis on points that will be important for professionals. Clinicians who see articulate patients with experiences they believe should be shared might encourage these patients to submit their articles to Schizophrenia Bulletin, First Person Accounts, EEI Communications, 66 Canal Center Plaza, Suite 200, Alexandria, VA 22314.--The Editors. PMID- 15279062 TI - Pharmacogenetics of antipsychoatics. AB - Although a number of antipsychotics have been introduced for the treatment of schizophrenia, inter-individual differences of in antipsychotic response and the number of refractory schizophrenic patients have become two of the most challenging problems in clinical psychiatry. Thus, the pharmacogenetics of antipsychotics have been aimed at providing genetic components of this inter individual variability in antipsychotic response in order to establish an individually-based pharmacotherapy for schizophrenia and to elucidate the mechanism of antipsychotic response so as to solve the refractoriness of schizophrenia. Pharmacogenetics, which is defined as the science of pharmacological response and its modification by hereditary influence can be divided into two categories: the genetic background of pharmacokinetics, i.e. the absorption, distribution, tissue localization, biotransformation and excretion of drugs, and pharmacodynamics, i.e. the biochemical and physiological consequences of a drug and its mechanism of action. Pharmacokinetics of antipsychotics has been focused mainly on the association between genetic polymorphisms in CYP genes, including CYP2D6, and the metabolism of these drugs. Polymorphism in CYP2D6 enables a division of individuals within a given population into at least two groups, i.e. poor metabolizers (PMs), extensive metabolizers (EMs), and ultrarapid metabolizers (UMs) of certain drugs. PMs have higher plasma concentrations of and more adverse effects from antipsychotics. UMs could be one of the important factors that induce treatment-refractoriness to antipsychotics. Genetic polymorphisms in serotonin and dopamine receptors that have a high affinity for antipsychotics have so far been extensively investigated in the pharmacodynamics of this type of drug. Not just one gene but multiple genes play a role in complex phenotypes, including the clinical response to medication. Thus, a multiple candidate genes approach has recently been adopted in the pharmacogenetics of antipsychotics. The new field of pharmacogenomics using DNA microarray analysis, which focuses on the genetic determinants of drug response at the level of the entire human genome, is important for development and prescription of safer and more effective individually-tailored antipsychotics. PMID- 15279063 TI - Prolonged cytostatic tumor dormancy induced by serial exchange of chemotherapy in colorectal carcinoma. AB - BACKGROUND: To improve quality of life cytostatic effect of serially changed chemotherapy was investigated. MATERIALS AND METHODS: Nonrandomized controlled trial in 17 patients with diagnosis of metastasis or recurrence following primary colorectal carcinoma was conducted from 1996 through 2001. Patients underwent low dose CDDP+5-FU monitoring continual CEA level. Whenever uninterrupted increase for minimally 3 times of CEA level was observed, the next chemotherapy was chosen from the following chemotherapy: l-Leucovorin+5-FU, low-dose CPT-11. RESULTS: Six were died of carcinoma. Median survival time from primary surgery and those from the day of diagnosis of metastasis were 48.6 and 23.3 months, respectively. Most of the patients experienced decrease in CEA level after continuous increase. No severe side effects were observed in them except one who died of hyperosmolar diabetic syndrome. CONCLUSIONS: Although the present trial should await further follow-up to confirm the clinical relevance of its modality, longer survival attained by the serially exchanged chemotherapy would implicate future chemotherapeutic strategy. PMID- 15279064 TI - Videofluorographic observations on swallowing in patients with dysphagia due to neurodegenerative diseases. AB - We examined three intervention methods for their efficacy in preventing aspiration in 25 patients with Parkinson's disease (PD) and 23 patients with degenerative cerebellar ataxia (CA). On videofluoroscopic examination. 13 patients with PD (52%) and 7 patients with CA (30.4%) showed aspiration. In all PD patients and 5 patients with CA, no aspiration was observed after changing the food form. With the chin down posture and supraglottic swallow techniques, no aspiration was observed in only 1 PD patient. Among 7 patients with CA, the chin down posture and supraglottic swallow techniques resulted in the disappearance of aspiration in 4 patients. This indicates that changing the food form (ex. jelly) was effective in preventing aspiration in both PD and CA patients with a history of aspiration. In addition, the chin down posture and supraglottic swallow techniques were effective in preventing aspiration in CA patients with good sitting-position balance and cervical control. PMID- 15279065 TI - A quantitative study of nerve fiber density in the submandibular gland of rats. AB - The route and three dimensional distribution of nerve terminals in the submandibular gland were investigated in rats using immunohistochemistry for the protein gene product (PGP) 9.5, as a marker of neuronal elements. Thick fiber bundles were found along the wall of the excretory duct. Many fine fibers from these thick bundles were distributed each lobule of the submandibular gland. A large number of single fibers terminated in the area around the striated, intercalated ducts and the acini. The densities of PGP 9.5 immunoreactive terminals were measured by a computer aided analysis system in the three areas: the striated duct, the intercalated duct, and the acini, whose densities (microm/microm2) were 0.23, 0.39 and 0.05 respectively. The relatively high density of nerve terminals in the intercalated duct suggests that the duct system probably plays an unexpectedly important role in the functional aspects. PMID- 15279066 TI - Incorporation, remodeling and re-expression of exogenous gangliosides in human cancer cell lines in vitro and in vivo. AB - Human neuroblastomas and gliomas express high levels of GD2 ganglioside. Mechanisms for the re-expression of GD2 after the incorporation of an exogenous precursor structure were analyzed using a human heterophilic monoclonal antibody (mAb) together with mouse anti-GD3 and mouse anti-GD2 mAbs. First, mouse anti-GD2 mAb 220-51 was generated and its reactivity was confirmed to be almost identical with that of the well-known mAb 3F8 antibody. As reported previously for GD3 variants, new ganglioside antigens reactive with human mAb 32-27 were analyzed by culturing an astrocytoma cell line AS in the presence of NeuGc-GM3. Analysis of the extracted gangliosides from AS thus cultured revealed a new component detected with mAb 32-27, migrating similarly to GD2. Incorporated NeuGc-GM3 seemed to be converted to NeuAc-NeuGc-type GD3, and then to NeuAc-NeuGc-type GD2 with alpha2,8-sialyltransferase and beta1,4-GalNAc transferase, respectively. In addition, AS was inoculated into nude mice, and glycolipids were extracted from generated tumors. Analysis of the ganglioside components using mAbs indicated that NeuAc-NeuGc-type GD2 was generated in the xenogeneic tumors by incorporating NeuGc-GM3 from mouse blood. These results indicated the presence of a pathway for utilization of exogenous gangliosides for remodeling and re-expression in vivo. PMID- 15279067 TI - Genotype announcement in a genetic polymorphism study for health checkup examinees at Nagoya University Hospital. AB - On June 9, 2003, we started free genetic tests of eight polymorphisms for health checkup examinees who attended a basic course at Nagoya University Hospital. They were informed of their genotypes within four weeks after blood donation for research purposes. The genotypes were those of alcohol dehydrogenase 2 (ADH2) Arg47His, aldehyde dehydrogenase 2 (ALDH2) Glu487Lys, NAD(P)H: quinone oxidoreductase (NQO1) C609T, glutathione S transferase M1 (GSTM1), glutathione S transferase T1 (GSTT1), interleukin-1B (IL-1B) C-31T, and tumor necrosis factor A (TNF-A) T-1031C, angiotensin-converting enzyme (ACE) Ins/Del. In the first three months, 227 (89.4%) out of 254 examinees participated in the free tests, having been informed of the research aims, after which they consented to our use of research data. To date, there have been no complaints from the participants, indicating that the announcement of polymorphism genotypes may be accepted differently from that of hereditary disease genotypes. PMID- 15279068 TI - Associations between disease risk and eight polymorphisms adopted for genotype announcements at Nagoya University Hospital. AB - Genetic polymorphisms have the potential to predict disease susceptibility. This may be especially useful among individuals with a high-risk lifestyle, so that the genotyping could be adopted for disease prevention through modifications toward a lower-risk lifestyle. We started a program of free genotype announcements in a polymorphism study among health checkup examinees at the Nagoya University Hospital on June 9, 2003. Since such announcements remain controversial for fear of unexpected harmful effects and counseling system, the accumulated evidence on the association between disease risk and genotypes announcements in our study was reviewed in this article. The genotypes used were those of alcohol dehydrogenase 2 (ADH2) Arg47His, aldelhyde dehydlrogenase 2 (ALDH2) Glu487Lys, NAD(P)H: quinone oxidoreductase (NQO1) C609T, glutathlione S transferase M1 (GSTM1), glutathione S-transferase T1 (GSTT1), interleukin-1B (IL 1B) C-31T, and tumor necrosis factor A (TNF-A) T-1031C, angiotensin converting enzyme (ACE) Ins/Del. Since showed a potential for widespread use in health checkups, the information on the above polymorphisms seems worth documenting. Although there have been no complaints from the participants to date, careful treatments are requested. PMID- 15279069 TI - Baseline and CRH-stimulated ACTH and cortisol levels after administration of the peroxisome proliferator-activated receptor-gamma ligand, rosiglitazone, in Cushing's disease. AB - The ability of acute rosiglitazone administration in influencing ACTH/cortisol secretion in basal conditions and after CRH stimulation was studied in patients with Cushing's disease. Ten patients (8 women and 2 men, aged 18-65 yr) with Cushing's disease were enrolled in the study: 6 of them had previously undergone unsuccessful surgery and 4 were untreated. Plasma ACTH and serum cortisol levels were evaluated at serial time points for 3 h during saline infusion and after the administration of rosiglitazone (8 mg, po) and for 1 h after the injection of CRH (1 microg/kg iv) given alone or 30 min following rosiglitazone administration. The 4 tests were performed in all subjects in randomized order on different days. No significant difference was observed between the pattern of hormone secretion during saline alone and after rosiglitazone, as evaluated by two-way analysis of variance (ANOVA). The integrated areas under the curves (AUCs) were also not significantly different (ACTH: 5683 +/- 1038 vs 6111 +/- 1007 pg/ml/180 min; cortisol: 2333 +/- 267 vs 2902 +/- 486 microg/dl/180 min). In addition, there was no difference for ACTH and cortisol responses to CRH given either alone or after rosiglitazone, when evaluated as peak, increment or AUC; the pattern of the responses analyzed by two-way ANOVA was also similar. IN CONCLUSION: 1) the administration of a single dose of rosiglitazone did not decrease ACTH/cortisol levels or blunt their response after CRH injection; 2) the activation of PPAR gamma receptors by rosiglitazone seems unable to affect ACTH and cortisol secretion, at least in acute conditions, in patients with ACTH-secreting pituitary adenomas. PMID- 15279070 TI - Meal intake similarly reduces circulating concentrations of octanoyl and total ghrelin in humans. AB - ABSTRACT. Several data show that meal intake and nutritional status regulate circulating ghrelin concentrations in humans. Ghrelin mainly circulates in two different forms: octanoyl and des-octanoyl ghrelin. Most circulating ghrelin is des-octanoyl ghrelin which is considered inactive because it lacks endocrine activity. However, recent evidence suggests that des-octanoyl ghrelin exerts biological activity such as stimulation of adipogenesis, cardiovascular effects and control of cell growth. In healthy humans, although the total ghrelin concentration is known to peak before meals and to be reduced by food intake, no data are available about the octanoyl ghrelin response in the absorptive state. Therefore, after an overnight fast, we compared the effects of a mixed meal ingestion (meal study) or of additional 240 min fasting (control study) on plasma concentrations of octanoyl and total ghrelin in 6 healthy subjects (body mass index: 23 +/- 0.7). At baseline, octanoyl-ghrelin accounted for 3.15 +/- 0.2% of total circulating ghrelin without differences between the two sessions. A similar ratio was maintained in the absorptive state with no differences between the studies and basal values. Compared with control, meal intake significantly suppressed (nadir at 90 min) octanoyl and total ghrelin by 38 +/- 3 and 40 +/- 3% of basal values, respectively. In the meal study, multivariate analysis of variance showed that serum insulin best predicted plasma octanoyl-ghrelin concentrations accounting for 97% of its variation (r2 = -0.97,p = 0.0016). IN CONCLUSION: in healthy humans, octanoyl-ghrelin represents about 3-4% of total circula-ting ghrelin and this ratio is closely maintained in post-absorptive and absorptive states. PMID- 15279071 TI - Elite volunteer athletes of different sport disciplines may have elevated baseline GH levels divorced from unaltered levels of both IGF-I and GH-dependent bone and collagen markers: a study on-the-field. AB - Seventy-seven Italian eliteathletes(42 M, 35 F, mean age +/- SE: 24.4-0.7 yr, age range: 17-47 yr) of different sport disciplines (sprinters, triathletes, middle distance runners, road-walkers, cyclists, rowing athletes, skiers, roller hockey players, swimmers) were sampled on-the-field (before a training session) for the determination of basal GH, IGF-I, C-terminal cross-linked telopeptide of type I collagen (ICTP) and amino-terminal propeptide of type III procollagen (PIIINP) levels, two GH-dependent peripheral markers of bone and collagen turnover, respectively. Basal GH concentrations were significantly higher (p<0.001) in female (5.8 +/- 1.0 ng/ml) vs male athletes (1.8 +/- 0.5 ng/ml), with a large spread of values in either gender. Mean GH levels of athletes were significantly higher than those recorded in age-matched sedentary controls (females: 2.5 +/- 0.5 ng/ml, p<0.001; males: 0.5 +/- 0.2 ng/ml, p<0.05). Among female athletes, 7/35 had basal GH values higher than the upper limit of control values (>9.5 ng/ml), while among males 7/42 had values higher than the upper limit of male sedentary controls (>3.6 ng/ml). No significant differences in basal GH concentrations were found between females taking oral contraceptives (OC) and those who did not receive this treatment (5.0 +/- 2.1 vs 6.0 +/- 1.2 ng/ml). IGF I levels (236.4 +/- 7.8 ng/ml) were in the normal range for age in all athletes (except for 1 athlete with slightly increased levels), no significant correlation being found between GH and IGF-I levels (R2=0.0393). Mean ICTP (4.6 +/- 0.2 ng/ml) and PIIINP (4.4-0.1 ng/ml) concentrations of elite athletes were not significantly different from those recorded in age and matched healthy sedentary subjects; 4 athletes showed increased PIIINP levels and 2 had increased ICTP levels. ICTP and PIIINP levels were positively correlated with chronological age (p<0.001), a positive correlation being also found between the two markers (p<0.001). On the contrary, no significant correlation was found between basal GH/IGF-I levels and ICTP/PIIINP levels. In conclusion, the present study demonstrates that: 1) elite athletes (particularly females), which have frequently increased basal GH on-the-field, have actually normal IGF-I levels; 2) ICTP and PIIINP levels of athletes are similar to those recorded in healthy sedentary, being significantly higher in younger subjects of both groups; 3) the presence of increased basal GH levels, being associated with normal IGF-I, ICTP and PIIINP levels, is probably the result of a transient GH peak in this study group. Further additional studies are requested to verify the possible use of these peripheral GH-dependent markers for detecting exogenous chronic administration of recombinant GH in athletes. PMID- 15279072 TI - Effect of raloxifene and its interaction with human PTH on bone formation. AB - We studied the of effects raloxifene alone or in combination with human PTH (hPTH) 1-34 in mineralizing cultures of SaOS-2 cells. Raloxifene (10(-8)-10(-6) M) increased bone nodule formation in cultures of SaOS-2 cells when added intermittently from day 8 to day 17. A single 24-h treatment with 10(-8) M hPTH (1-34) at day 8 reduced the nodule area by 75.6% at day 17, and raloxifene added concomitantly with hPTH (1-34) reduced this inhibitory effect in a dose-dependent manner. Raloxifene also reduced the hPTH (1-34)-induced inhibition of alkaline phosphatase (ALP) activity. The 10-fold stimulation of c-fos mRNA expression by hPTH (1-34) was not influenced by raloxifene co-treatment. The protein kinase A (PKA) inhibitor 6-22 amide (1.7 nM) and the protein kinase C (PKC) inhibitor bisindolylmaleimide 1 (10 nM) did not influence the separate effects of PTH and raloxifene on mineralized bone nodule formation. This is the first report on the interaction of PTH and raloxifene in an osteoblast culture system. PMID- 15279073 TI - Body composition, fat distribution and metabolic characteristics in lean and obese women with polycystic ovary syndrome. AB - The polycystic ovary syndrome (PCOS), characterized by chronic anovulation and hyperandrogenism, has many features of metabolic syndrome and can be considered a metabolic disease. Approximately 50% of patients with PCOS are overweight or obese with abdominal fat accumulation. Some metabolic alterations and abdominal fat distribution have also been reported in lean women with PCOS. The aim of this study was to evaluate the effect, if any, of obesity on metabolic features, body composition and fat distribution in patients with PCOS. Body composition and abdominal fat distribution (evaluated by DEXA), waist circumference, blood pressure, lipid profile, glucose tolerance and homeostasis model assessment index were determined in 23 lean [mean age 23 +/- 5 yr, mean body mass index (BMI) 22 +/- 2 kg/m2] and 27 overweight-obese (mean age 21 +/- 5 yr, mean BMI 32 +/- 5 kg/m2) patients with PCOS and in 20 age- and weight-matched eumenorrhoic women. Patients exhibited slight but non-significant differences in metabolic parameters, waist circumference, blood pressure and total and abdominal fat content compared with weight-matched controls. None of the lean subjects suffered from metabolic syndrome according to the National Cholesterol Education Program- Adult Treatment Panel III (NCEP-ATPIII) criteria as opposed to 10 overweight obese patients and three overweight-obese control subjects (37% and 33.3% of each subgroup, respectively). Our data do not show significant metabolic alterations in lean PCOS women. Results indicate that obesity seems to underpin the metabolic alterations exhibited by the overweight-obese patients. However, since women with PCOS are at increased cardiovascular risk, further studies are needed to evaluate metabolic alterations and body composition in these patients. PMID- 15279074 TI - Mutations in the SLC26A4 (pendrin) gene in patients with sensorineural deafness and enlarged vestibular aqueduct. AB - Pendred syndrome and the enlarged vestibular aqueduct (EVA) are considered phenotypic variations of the same entity due to mutations in the SLC26A4 (pendrin) gene. Pendred syndrome consists in sensorineural deafness, goiter and impaired thyroid hormone synthesis while in EVA thyroid function seems to be preserved. The aim of this study was to evaluate thyroid function and morphology and to look for mutations in the SLC26A4 gene in patients presented with EVA. Among 57 consecutive patients with sensorineural deafness 15 with EVA, as assessed by magnetic resonance imaging (MRI), were identified and studied. A complete evaluation of thyroid function including thyroid echography and perchlorate discharge test was carried out in all patients with EVA; all exons of the SLC26A4 gene were amplified from peripheral leukocytes and directly sequenced, using specific intronic primers. Out of 15 patients with EVA, goiter was present in 8 (53%), hypothyroidism in 7 (47%), increased serum thyroglobulin levels in 8 (53%) and a positive perchlorate discharge test in 10 (67%). Nine alleles of the SLC26A4 gene were mutated: 2 novel mutations (L465W and G497R) and 4 already known mutations (T410M, R409H, T505N and IVS1001+1G>A) were found. Four subjects were compound heterozygous and 1 heterozygous (G497R/wt). All patients harbouring mutations in the SLC26A4 gene had goiter and a positive perchlorate discharge test: 3 were slightly hypothyroid and 2 euthyroid. The remaining 10 patients had no mutations in the SLC26A4 gene: 4 of them were hypothyroid, 2 with goiter and positive perchlorate discharge test, 2 without goiter and with negative perchlorate discharge test. Two patients without mutations were euthyroid with positive perchlorate discharge test. Patients with mutations in the SLC26A4 gene had larger thyroid volume (p<0.002), higher serum thyroglobulin (Tg) levels (p<0.002) and greater radioiodine discharge after perchlorate (p=0.09) than patients without mutations. The results of the present study lend support to the concept that all patients with mutated SLC26A4 gene have abnormalities of thyroid function tests. PMID- 15279075 TI - The adrenal sensitivity to ACTH stimulation is preserved in anorexia nervosa. AB - Hyperactivity of hypothalamus-pituitary-adrenal (HPA) axis in anorexia nervosa (AN) has been demonstrated and is likely to reflect a central nervous system (CNS)-mediated effect of starvation. Alterations in the adrenal response to ACTH in AN have also been reported by some authors. In order to define the adrenal sensitivity to ACTH in this condition, we studied cortisol (F), aldosterone (A) and DHEA responses to the sequential administration of low and supramaximal ACTH 1-24 doses (0.06 microg/m2 ACTH 1-24 iv at 0 min and 250 microg ACTH 1-24 iv at +60 min, respectively) in 10 young women with AN [ANW, age 21.2 +/- 0.9 yr, body mass index (BMI) 15.7 +/- 0.6 kg/m2]. The results in this group were compared with those recorded in 10 healthy normal women (HW, 23.4 +/- 1.1 yr, 21.9 +/- 0.9 kg/m2). In ANW urinary F levels were similar to those in HW. Basal serum F, A and DHEA levels in ANW were not significantly different from those in HW. In HW the lowest ACTH dose induced a significant (p<0.05) increase of F, A and DHEA. The maximal ACTH dose induced F, A and DHEA increases greater (p<0.05) than those induced by the lowest ACTH dose. In ANW both ACTH doses induced significant (p<0.05) F and DHEA increases which were not significantly different from those in HW, though a trend toward a lower cortisol response after ACTH 0.06 microg/m2 in ANW was present. Like in HW, in ANW the maximal ACTH dose induced F and DHEA increases greater (p<0.01) than those induced by the lowest dose. Unlike HW, in ANW A levels did not increase after the lowest ACTH dose while they increased after the maximal one overlapping the response in HW. In conclusion, the cortisol and DHEA responses to a very low and a supra-maximal ACTH dose in patients with AN were similar to those in healthy subjects, indicating that the sensitivity to ACTH of the fasciculata and reticularis adrenal zones is preserved in this condition. On the other hand, a reduced sensitivity to ACTH of the glomerularis adrenal zone in patients with AN is suggested by the lack of aldosterone response to the lowest corticotropin dose. PMID- 15279076 TI - Postmenopausal serum androstenedione levels are associated with the calcitonin receptor gene polymorphism T1377c. A pilot study. AB - We tested the hypothesis that homeostasis of sex-steroids is related to the calcitonin receptor (CALCR) genotype. To determine the CALCR genotype PCR amplification followed by digestion with Alul restriction enzyme were carried out according to Nakamura et al. Indeed, a single nucleotide difference at position 1377 of cDNA generates two alleles (CC genotype or TT genotype). Serum estradiol, testosterone and their precursors androstendione (AD) and DHEA levels were estimated in a cohort of 113 postmenopausal women. While serum DHEA levels did not differ between the individual allele combinations, AD levels as well as AD/DHEA ratio were higher in carriers of TC and CC genotypes than those with TT genotype (p<0.05 and p<0.02, respectively, ANCOVA). We postulate that the 3beta hydroxysteroid dehydrogenase activity is associated with C allele at least in C19 steroids. The data correspond with the functionality of CALCR. PMID- 15279077 TI - Comparison of c-Fos immunoreactivity in pancreatic beta cells and cells with neural crest, endoderm and mesoderm origin in rats. AB - AIMS: Pancreatic beta cells are known to share many similarities with neuronal cells, but their origin remains controversial. It has been hypothesized that pancreatic beta cells are derived from neural crest cells. The aim of this study was to investigate the similarities between pancreatic beta cells and cells with neural crest, endoderm and mesoderm origin with respect to c-Fos immunoreactivity (c-Fos-ir), which has a role in important cellular processes including cellular proliferation, growth, differentiation and apoptosis. METHODS: c-Fos-ir was analyzed by immunohistochemical methods in formaline-fixed, paraffin-embedded sections of rat pancreatic beta cells (BCs), in adrenal medullary chromaffin cells (CCs) that are derived from neural crest, in exocrine pancreatic acinar cells (ACs) that are derived from endoderm, and in adrenal cortex zona reticularis cells (RCs) that are derived from mesoderm. RESULTS: The statistical comparisons revealed no significant differences between BCs and CCs with respect to c-Fos-ir (p>0.05). However, a highly significant difference (p<0.001) with respect to c-Fos-ir both between ACs and RCs, and between these two cell types and each of the two other cell types was noted. CONCLUSIONS: As opposed to findings in cells without neural crest origin, the observed similarity between BCs and CCs with respect to c-Fos-ir, provides additional evidence for the similarity of these cells with cells derived from neural crest. PMID- 15279078 TI - Evaluation of two replacement regimens in primary adrenal insufficiency patients. effect on clinical symptoms, health-related quality of life and biochemical parameters. AB - OBJECTIVE: To evaluate clinical symptoms, health-related quality of life (HRQL) and biochemical parameters in patients with primary adrenal insufficiency under treatment with two different hydrocortisone regimens (20 mg-0 mg-10 mg/day and 10 mg-5 mg-5 mg/day), each maintained for 3 months and compare results obtained with those in healthy controls. DESIGN, PATIENTS AND METHODS: Twelve patients with primary adrenal insufficiency were studied. Clinical symptoms and HRQL with the Nottingham Health Profile (NHP) were evaluated and Na, K and serum cortisol determined at 09:00 h, 12:30 h and 17:30 h and urinary free cortisol (UFC) throughout the day. Control group comprised 19 healthy subjects. RESULTS: No differences in specific adrenal insufficiency symptoms were detected between the two regimens. HRQL was worse in energy dimension assessed by the NHP compared to the general population, regardless of 20 mg-0 mg-10 mg/day or 10 mg-5 mg-5 mg/day treatment (p=0.03 and p=0.013). The total NHP score was only adversely affected when patients were on the 10 mg-5 mg-5 mg/day hydrocortisone replacement regimen (p=0.008). Serum cortisol concentrations were higher than controls at 09:00 h, and lower at 17:30 h with both regimens, whereas serum cortisol at 12:30 h and UFC were within the 5th-95th percentile normal range only with the 10 mg-5 mg-5 mg/day regimen. CONCLUSIONS: Patients with primary adrenal insufficiency had worse HRQL in the NHP energy dimension compared with the general population, regardless of the hydrocortisone regimen although total score for HRQL was worse only with the 10 mg-5 mg-5 mg/day regimen. Patients on the thrice-daily hydrocortisone regimen showed a more physiological cortisol profile, leading us to recommend initially treating patients with this dose and increasing it in the case of impaired HRQL. PMID- 15279079 TI - Relationship between insulin resistance assessed by HOMA-IR and exercise test variables in asymptomatic middle-aged patients with type 2 diabetes. AB - We investigated the relationship between index of insulin resistance (IR) and exercise test variables in middle-aged asymptomatic patients with Type 2 diabetes. METHODS: 90 patients (48 men, 42 women; age: 49 +/- 6 yr) were included in the study. We used homeostasis model assessment for IR (HOMA-IR) index as index of IR. All patients were subjected to treadmill exercise test. Four subjects were tested positive (4.4%). Study patients were separated into three groups: group I (no.=26) HOMA-IR index <2.24; group II (no.=26) index 2.24-3.59; group III (no.=38) index >3.59. RESULTS: group I had less frequency of cardiovascular risk factors than group II and III (p=0.001). Systolic blood pressure baseline as well as peak exercise values, were higher in group III than in group I and II (p=0.048 vs p=0.01, respectively). Higher total exercise time and peak workload were found in group I than group II and III (p=0.04). The recovery of heart rate (delta HR(pr)) was similar among the study groups. We found significant negative correlations between HOMA-IR and total exercise time and peak workload. In addition we found significant negative correlations between age vs chronotrophic index (CI), delta HR(pr), and peak workload. There were also similar negative correlations between duration of diabetes vs CI and delta HR(pr). CONCLUSIONS: IR is associated with a variety of cardiovascular risk factors. Some exercise test variables point out changes of autonomic tone during exercise in elevated IR group. Negative correlation between HOMA-IR and peak exercise capacity (METs) may well confirm increased mortality in hyperinsulinemia. PMID- 15279080 TI - Spontaneous regression of hypercalcemia in a patient with primary hyperparathyroidism and prolactinoma. AB - We describe a unique case of spontaneous resolution of hyperparathyroidism in a lady with combined parathyroid adenoma and prolactinoma, raising the possibility of underlying multiple endocrine neoplasia (MEN) 1 syndrome. We also discuss the mechanism and natural history of such spontaneous remission. PMID- 15279082 TI - Metastatic renal cell carcinoma to the pituitary presenting with hyperprolactinemia. AB - Metastases to the pituitary gland from systemic cancers is a rare phenomenon and usually occurs in patients with disseminated disease. The neurohypophysis is the most commonly involved site, and diabetes insipidus is the most common presentation in these patients. Breast and lung cancer are the most common cancers metastasizing to the pituitary. Involvement of the pituitary by renal cell carcinoma (RCC) is exceedingly rare. Mild-to-moderate degree of hyperprolactinemia is a rare presentation of pituitary metastases. We report the case of a woman with metastatic RCC to the pituitary presenting an unusually high degree of hyperprolactinemia. PMID- 15279081 TI - Normal pregnancy in a woman with nesidioblastosis treated with somatostatin analog octreotide. AB - OBJECTIVE: We report the case of a 36-yr-old woman with nesidioblastosis treated throughout pregnancy with high doses of octreotide. We studied the course of blood glucose, foetal growth and development. METHODS: Blood samples were obtained every month throughout pregnancy and taken at birth from the umbilical cord. Sonography was performed repeatedly to monitor foetal growth. RESULTS: The daily dose of octreotide was adapted to blood glucose levels: a dose of 1000 microg was infused during the first part of pregnancy, then it was decreased step by step during the last trimester of gestation. An elective cesarean section was performed at 32 weeks of gestation. High octreotide concentrations were obtained during the first part of gestation (range 2888-5021 pg/ml). During the third trimester of pregnancy blood glucose increased despite high insulin levels attesting physiological insulin-resistance. Plasma levels of placental GH and IGF 1 levels were similar to those observed in a normal pregnancy. Despite the presence of octreotide in the umbilical cord, TSH, free T4, PRL and pituitary GH concentrations were normal at birth. The female newborn (weight 3520 g, length 52 cm) had no malformation, and presented with normal postnatal development. CONCLUSION: Our study demonstrates that: 1) octreotide treatment can be effective in controlling endogenous hyperinsulinism during pregnancy; 2) octreotide does not affect physiological changes during pregnancy such as insulin-resistance or placental GH level; 3) exposure of the foetus to octreotide throughout pregnancy does not induce any malformation and does not affect foetal development. PMID- 15279083 TI - Fatal toxic epidermal necrolysis in autoimmune polyglandular syndrome type I. AB - Autoimmune polyglandular syndrome Type I (APS I) is a disorder defined by the presence of at least two of the following diseases: Addison's disease, hypoparathyroidism, and chronic mucocutaneous candidiasis. We present the case of a 45-yr-old woman, affected by APS I, in chronic treatment with betamethasone. She was referred to a Division of General Medicine for jaundice, ascites and peripheral edema attributed to worsening of pre-existing autoimmune chronic hepatitis. During hospitalization, the following drugs were given: Amoxicillin/Clavulanic acid and Levofloxacin for bronchopneumonia, Furosemide and Canreonate for renal impairment, Pantoprazole for gastric protection, and Itraconazole for oral candidiasis. After about a month, she developed widespread, sheet-like, epidermal detachment, with painful lesions of the conjunctiva, lips and mouth. Toxic epidermal necrolysis (TEN) was diagnosed, and the patient was transferred to a Burn Center, where she died 10 days after the first onset of cutaneous rash. Autoptic and histopathological findings (epidermal necrosis and detachment, lymphomonocytic infiltration of the dermis) confirmed the clinical diagnosis. TEN is a usually drug-induced cutaneous inflammatory disorder characterized by extensive epidermal detachment and frequent mucosal involvement. It has also been associated with immuno-mediated disorders (HIV infection, graft vs-host disease, systemic lupus erythematosus, mixed essential cryoglobulinemia), in keeping with immuno-mediated pathogenesis. We present, to our knowledge, the first report of TEN in a patient with APS I, and suggest that some pathogenetic mechanisms of APS I are shared with TEN. We stress how such a disease can occur in an autoimmune syndrome, even during corticosteroid treatment. PMID- 15279084 TI - Autoimmune diabetes: more than just one flavor? PMID- 15279085 TI - Reduction of cardiovascular morbidity and mortality in type 2 diabetes. A rational approach to hypoglycemic therapy. AB - Type 2 diabetes mellitus is the single most important risk factor for the development of coronary artery disease. Unfortunately, the traditional therapeutic strategies for the treatment of hyperglycemia have proven to be ineffective in preventing cardiovascular complications. In recent years the number of available hypoglycemic agents has increased and considerable progress has been made regarding the comprehension of the pathophysiology of diabetes and its vascular complications. In the present article we firstly present benefits and risks of intensive vs standard hypoglycemic intervention, and the pros and cons of therapy targeted to postprandial hyperglycemia. Secondly, we discuss the cardiovascular effects of sulfonylurea agents and insulin, focusing on the role of intensive insulin treatment in the context of acute coronary syndromes. Thirdly, we review the epidemiological, clinical and experimental evidence linking insulin resistance and cardiovascular disease. Finally, we present the rationale and the role of metformin and thiazolidinedionetherapy in the prevention of cardiovascular complications. We conclude that the optimal use of the full spectrum of hypoglycemic agents has the potential to play a key role in the prevention of diabetes-related macrovascular complications. PMID- 15279087 TI - Ten years of democracy--what is the impact on oral health? PMID- 15279088 TI - Facial profile changes with various orthodontic premolar extraction sequences during growth. AB - The Visual Treatment Objective (VTO) is an extremely valuable tool in the process of working out the treatment plan for the individual. This is especially true for the inexperienced operator. Often it becomes clear that extractions are indicated, but there remains uncertainty as to which teeth should be chosen for extractions to give the best results. As it is necessary to express the expected change in profile due to growth of the nose and also of the chin, VTO construction is helpful in determining the extraction decision. Clinical experience and research projects have indicated that the use of mean values has definite limitations when constructing a VTO for an individual. The primary objective of this investigation was to provide formulae by means of which anteroposterior changes in the facial profile of growing individuals for whom four premolar extraction treatment is proposed can be predicted with a greater measure of certainty. The records of 248 growing Caucasian orthodontic patients (120 males and 128 females) were selected and divided into Group 44 (four first premolar extractions), Group 45 (upper first premolar and lower second premolar extractions) and Group 55 (four second premolar extractions). Formulae are presented for the prediction of the amount of incisor retraction, anteroposterior nose growth and for chin growth. The formulae were derived by regression of incisor retraction, anteroposterior nose growth and chin growth on various predictors, and could be used in the construction of VTO's for selected cases. PMID- 15279089 TI - Analysis of attendance rates at Soweto dental clinics 1995-2002. AB - BACKGROUND: Patient attendance rates at Soweto dental clinics increased during the year after the implementation of free primary oral health care in 1995. OBJECTIVE: This study was performed to examine if the attendance rates continued to increase between April 1995 and March 2002. MATERIALS AND METHODS: Monthly clinic records were used to record casual (pain and sepsis treatment) and booked patient attendance (restorative, prosthetic and orthodontic treatment) and number of dental operators in the nine primary health care clinics and one hospital clinic in Soweto. Data were analysed with SAS and Prism software. RESULTS: Total patient attendances in the primary health care clinics significantly increased from 6,161 in 1995 to 10,519 in 2002 (P<0.05) due to an increase in casual patients Booked patients decreased and patients treated per operator increased. In the hospital clinic the casual patient attendances decreased but booked patients significantly increased (P<0.005). CONCLUSIONS: Patient attendance rates increased between 1995 and 2002 with an increase in dental operator workload. PMID- 15279086 TI - Magnetic resonance imaging of the hypothalamus-pituitary unit in childrensuspected of hypopituitarism: who, how and when toinvestigate. AB - The magnetic resonance (MR) identification of pituitary hyperintensity in the posterior part of the sella has been the most striking recent finding contributing to the diagnosis of "idiopathic" and permanent GH deficiency (GHD). Moreover, advancements in DNA technology have shed new light on the study of the genetic causes of hypopituitarism. Abnormalities in two genes, the GH-N encoding the GH and the GHRH receptor (GHRH-R), have been identified, while mutations in five other gene-encoding transcription factors such as Pit-1, Prop-1, Hesx-1, Lhx 3 and Lhx-4 involved in anterior pituitary development, have also been described. MR imaging shows marked differences in pituitary morphology indicating different GHD etiologies and different prognoses. Ectopic posterior pituitary is a specific marker of permanent GHD. These patients do not have Pit-1, Prop-1, or Lhx-3 mutations and should be carefully monitored for evolving pituitary hormone defects, though they do not require GH re-evaluation in adulthood; selected cases may have Hesx-1 or Lhx-4 mutations. MR evidence of normal or small anterior pituitary gland, enlarged empty sella, pituitary hyperplasia and/or intrasellar or suprasellar mass when associated with combined pituitary hormone deficiency call for molecular analysis of Pit-1, Prop-1, Hesx-1, or Lhx-3. Limitation of neck rotation and Chiari-I malformation may suggest Lhx-3 or Lhx-4 mutations (exceedingly rare). In "idiopathic" isolated GHD, evidence of normal anterior or small anterior pituitary size with normal location of posterior pituitary and normal connection between the hypothalamus and pituitary gland is suggestive of "transitory" or false positive GHD; patients with such characteristics should be re-evaluated well before reaching adult height. In selected cases, anterior pituitary height that is 2 SD below age-adjusted normal pituitary height could be suggestive of GHRH-R gene defect; it is worth pointing out that normal pituitary MR together with severe GHD has been observed, though rarely, in subjects with a genetic origin of GHD. PMID- 15279090 TI - A simple appliance for the management of obstructive sleep apnoea--the MEDUNSA Anti-Snoring Device (MASD). AB - Obstructive sleep apnoea (OSA) is a breathing abnormality that occurs during sleep. It is characterised by snoring and episodes of sleep disturbance that can occur from 10 to 500 times per night causing excessive daytime exhaustion. A large tongue and a retrognathic mandible have been listed as aetiological factors. Orthodontic appliances that have been used to manage the condition are usually complicated, uncomfortable, expensive and delicate. The aim of this clinical procedure is to design a simple, inexpensive, durable and user-friendly appliance that can be used in the management of OSA. The procedure involves taking upper and lower impressions of a patient with OSA and articulating the study casts in a registered protrusive bite. Gum guards of the dental arches are then constructed and fused together in the protrusive position with a 5 to 10 mm open bite. The forward and downward advancement of the mandible caused by the appliance drags the tongue and other structures along, and thus opens the posterior pharyngeal airway during sleep. For ease of reference this appliance has been named the MEDUNSA Anti-Snoring Device (MASD) and it differs from similar appliances in that it is entirely toothborne and does not impinge on the surrounding soft tissues. The MASD was used in a patient with OSA at the MEDUNSA Oral Health Centre. Pre-treatment and post treatment airway dimensions from the lateral cephalograms confirm an increase of 50 per cent in airway size at the level of the epiglottis with a corresponding reduction in clinical symptoms. The MASD is an extremely valuable tool to dentists, orthodontists, as well as any other health professionals managing sleep-breathing abnormalities. While the MASD has been successful in this preliminary case, clinical trials are under way to further evaluate the MASD. PMID- 15279091 TI - Essential oils and interdental hygiene. PMID- 15279092 TI - A review of foetal alcohol syndrome and the role of the oral health care worker. AB - Foetal Alcohol Syndrome (FAS) consists of multi-system abnormalities and is caused by the excessive intake of alcohol during pregnancy. The teratogenic effect of alcohol on humans has now been established beyond reasonable doubt and FAS is one of the most important human teratogenic conditions known today. Many investigators have catalogued, quantified and refined these hallmarks of FAS over the years and have established that the most consistent consequences of heavy maternal drinking during pregnancy are prenatal and postnatal growth deficiency, and brain and craniofacial abnormalities. Many practitioners are unable to recognise the often subtle features associated with FAS and fail to diagnose it. The purpose of this article is to review the epidemiology, pathophysiology, clinical presentation and discuss the relevance of FAS to clinical dental practice. PMID- 15279093 TI - An update on the pathophysiology of sepsis. AB - Mortality among critically ill patients has been attributed to the development of sepsis. About 28% of patients with sepsis still die, despite numerous interventions. Trials on sepsis investigated mostly anti-inflammatory strategies, based on the prevailing theory that sepsis represents an uncontrolled inflammatory response. None of these showed convincing benefit in humans, despite promising results in animal studies. The reason for this is becoming clear: sepsis represents a biphasic response to infection, and the initial pro inflammatory response that we have targeted thus far is invariably followed by a prolonged period of immune suppression. In addition, a patient may oscillate between a pro- and anti-inflammatory state repeatedly. A single magic bullet therapy is thus unlikely to work. The mediators of this process are the cytokines, and a lot of research is focussed on modulating these to achieve a better outcome. In addition, the central role of the coagulation cascade in mediating inflammation and sepsis is becoming clear, and therapies addressing this mechanism are promising. PMID- 15279094 TI - General practitioner's radiology case 21. PMID- 15279095 TI - Metal content in river suspended particulate matter: data on Po River. AB - The present study summarizes the last ten years of literature on heavy metal distribution in Suspended Particulate Matter (SPM) and dissolved phase in the Po River (Italy). The work compares different methodologies employed to collect, concentrate and fractionate the samples. The importance of metal speciation as a function of particle size is underlined and two approaches to metal speciation in the colloidal fraction of Po River SPM are presented: Sedimentation Field-Flow Fractionation (SdFFF) and pH-dependent extractions. Finally, emphasis is placed on the need for comparison with a reference "natural background level" of the metal load in Po River particulate matter in order to determine the real human contribution to river pollution. The high values of some trace transition elements, such as Cr and Ni, have been compared with clay sediments around Ferrara and with bricks in historic buildings. The highly comparable natural concentration of these metals in Po fine sedimentary rocks and in historic brick buildings of Ferrara (XII-XVI centuries) can provide information on natural geochemical anomalies. PMID- 15279096 TI - Spectrophotometric determination of aluminium and nickel. AB - A simple spectrophotometric method, based on the complexes with xylenol orange (XO) and EDTA, is presented for the rapid determination of aluminium and nickel, respectively, in synthetic samples of hydrotalcite. The method only requires the solubilization in sulphuric acid of the inorganic material before the ligand addition. Under optimum conditions, the complexes Al-XO and Ni-EDTA showed maximum absorption at 554 nm and 380 nm, respectively. The method obeyed Beer's law in the concentration range 0.14-1.8 microg mL(-1) of aluminium, and 30-2730 microg mL(-1) of nickel. Molar absorptivities were 2.45 x 10(4) and 14.85 L mol( 1) cm(-1) while Sandell's sensitivities were 1.1 x 10(-3) and 3.9 microg cm(-2) for aluminium and nickel, respectively. The standard addition technique was used and the recoveries obtained revealed that the proposed procedure shows good accuracy. PMID- 15279097 TI - Chemometric studies in the Lagoon of Venice, Italy. Annual evolution of sulphur species and relationship to biogeochemical cycles in lagoon water. AB - During the period March 1997-March 1998 dimethyl sulphide (DMS), dimethylsulphoniopropionate (DMSP) and carbon disulphide (CS2) were determined weekly in the water of the Lagoon of Venice, Italy (at three stations located in the Giudecca Canal, the San Secondo Canal and the Rio di San Nicolo). At the same time, the following hydrological and biological variables were also measured: tide height, temperature, transmittance, fluorescence, pH, salinity, chlorinity, sulphate, ammonia, nitrite, nitrate, phosphate, silicate, chlorophyll a, phaeopigments, phytoplankton (abundance and biomass). Principal component analysis (PCA), applied as a dimension reduction tool, made it possible to summarize multivariate information in a small number of components, which highlighted the relationships between the temporal evolutions of the sulphur compounds with hydrological and biological variables in the seasonal biogeochemical cycle of the lagoon. In particular the overall temporal cycle, which begins with the development of biological activity in late winter and spring, followed by the predominance of degradation processes during the late summer and the remineralization of nutrients in autumn, is clearly described in the plane of the first two principal components, together with the interrelationships between all the relevant variables. PMID- 15279098 TI - Chitosan membrane: tool for chromium (III) recovery from aqueous solutions. AB - In this study four kinds of Chitosan membranes were prepared. The permeability of the membranes was then investigated by permeation of K+ in these membranes. The performances of the prepared membranes for recovery of chromium (III) from aqueous solution were evaluated. The effects of operation conditions on permeation characteristics were determined, and permeation mechanism was discussed. The stability of the membranes was also studied. PMID- 15279099 TI - Improving primary treatment of urban wastewater with lime-induced coagulation. AB - The enhancement of primary treatment efficiency through the coagulation process may yield several advantages, including lower aeration energy in the subsequent biological unit and higher recovery of biogas from sludge digestion. In this work sewage coagulation with lime was studied at pilot plant level, using degritted sewage from the city of Rome. The work aimed at optimising the operating conditions (coagulant dosage or treatment pH, and mixing conditions in the coagulation and flocculation tanks), in order to maximise the efficiency of suspended Chemical Oxygen Demand (COD) removal and to minimise sludge production. Lime dosage optimisation resulted in an optimal treatment pH of 9. Lime addition up to pH 9 may increase COD removal rate in the primary treatment from typical 30 35% of plain sedimentation up to 55-70%. Within the velocity gradients experimented in this work (314-795 s(-1) for the coagulation tank and 13-46 s(-1) for the flocculation tank), mixing conditions did not significantly affect the lime-enhanced process, which seems to be controlled by slow lime dissolution. Sludge produced in the lime-enhanced process settled and compacted easily, inducing an average 36% decrease in sludge volume with respect to plain settling. However excess sludge was produced, which was not accounted for by the amount of suspended solids removed. This is probably due to incomplete dissolution of lime, which may be partially incorporated in the sludge. PMID- 15279100 TI - Comparison of three sequential extraction procedures (original and modified 3 steps BCR procedure) applied to sediments of different origin. AB - The 3 steps sequential extraction procedure proposed by the Standards Measurements and Testing program (SM&T--formerly BCR) of the European Union has been applied for the speciation of metals in sediments. Results obtained by the application of the BCR standardized procedure were compared to those of two four step sequential extraction procedures, which are different from the BCR procedure only for the introduction of an additional step with NaOCl, as 2nd and 3rd step respectively. Five different metals have been taken into consideration: Cd, Cu, Ni, Pb and Zn. The analytical performances of the laboratory have been evaluated using three certified reference materials: the BCR 601 lake sediment for the BCR sequential extraction procedure, PACS-1 and MESS-1 for total metal concentration. Results showed that the efficiency of NaOCl treatment is higher or at least equal to that of H2O2 treatment and that its selectivity is quite satisfying. Moreover the NaOCl treatment doesn't significantly influence the extraction of the easily reducible fraction. PMID- 15279101 TI - Photocatalytic mineralisation of aniline derivatives in aquatic systems using semiconductor oxides. AB - Photocatalytic degradation of aqueous solution of aniline derivatives such as ortho-nitroaniline (ONA), meta-nitroaniline (MNA), para-nitroaniline (PNA), 4 bromoaniline (4-BrA) and 2-chloroaniline (2-ClA) were carried out over ZnO or TiO2 (anatase and rutile) in a photocatalytic reactor. The observed results revealed that the order of photocatalytic activity for degradation of selected compound was ZnO > TiO2 (rutile) > TiO2 (anatase) with the ratio of the rate constants to the surface area of 3.2 x 10(-3), 1.9 x 10(-3) and 1.0 x 10(-3) respectively. The effect of some physical and chemical parameters such as amount of photocatalyst, pH, time of irradiation and solvent were studied. Degradation kinetic was according to Longmuir behaviour. Spectrophotometric methods and TOC analysis supported that aniline derivatives almost completely mineralized. PMID- 15279102 TI - Chromium speciation by surfactant assisted solid-phase extraction and flame atomic absorption spectrometric detection. AB - A new procedure has been developed for chromium speciation in aqueous solution by the use of micellar, ion-association, solid-phase extraction techniques (SPE) followed by flame atomic absorption spectrometry. The method was based on the use of C-18 bonded phase silica SPE disks for retention of ion-associated Cr(VI) with cetyl trimethyl ammonium bromide (CTAB), elution of the retained species and subsequent detection by flame atomic absorption spectrometry (FAAS). Cr(III) was oxidized by potassium persulfate to Cr(VI), then the total chromium was retained on the disk and determined by FAAS. The amount of Cr(III) was calculated by the difference between the total and Cr(VI) values. The calculated limit of detections (LOD) (based on 3sigma) are 15 microg L(-1) and 20 microg L(-1) for Cr(VI) and Cr(III) respectively. No considerable interferences have been observed from other investigated anions and cations and the method has been successfully applied to water samples taken from the Karoon River in Khuzestan province. PMID- 15279103 TI - Differential pulse polarographic determination of lead in standard alloys and biological samples after separation and preconcentration with PAN. AB - A highly selective, sensitive, rapid and economical differential pulse polarographic method has been developed for the determination of trace amount of lead in various samples after adsorption of its 1-(2-pyridylazo)-2-naphthol (PAN) on naphthalene in the pH range of 8.4 - 11.5. After filtration, the solid mass is shaken with 9.0 ml of 1 M hydrochloric acid and lead is determined by differential pulse polarography (DPP). Lead can alternatively be quantitatively adsorbed on [1-(2-pyridylazo)-2-naphthol]-naphthalene adsorbent packed in a column and determined similarly. The detection limit is 0.1 microg/ml (signal to noise ratio = 2) and the linearity is maintained in the concentration range 0.3 - 300 microg/ml with a correlation coefficient of 0.9996 and relative standard deviation of +/- 1.1%. Characterization of the electroactive process included an examination of the degree of reversibility. Various parameters such as the effect of pH, volume of aqueous phase and interference of a number of metal ions on the determination of lead has been studied in detail to optimize the conditions for determination in alloys and biological samples. PMID- 15279104 TI - Octadecyl silica membrane disks modified with a new Schiff's base for the preconcentration of lead and copper before their determination in water samples. AB - A simple and fast method for the extraction and determination of ppt level of Pb2+ and Cu2+ ions using octadecyl-bonded silica membrane disks modified by a new tetradentates Schiffs base [Bis(2,4-dimethoxy benzaldehyde) ethylen diimine](TDSB) and inductively coupled plasma atomic emission spectrometry is described. Extraction efficiency, and influence of flow rate, pH, type and the least amount of acid for stripping of Cu2+ and Pb2+ from the modified disks and break through volume were evaluated. The maximum capacity of the membrane disks modified by 5 mg of TDSB used was found to be 347 +/- 7 and 470 +/- 6 microg of copper and lead, respectively. The concentration factor is 500 (for 2500 mL water sample and flow rate of 20 mL min(-1)) and detection limit of the proposed method is 12.5 and 150.5 pg/ml for Cu2+ and Pb2+, respectively. The method was applied to the determination of Cu2+ and Pb2+ ions from various water, wastewater, black tea, and hot pepper samples. PMID- 15279105 TI - A nitric oxide biosensor based on horseradish peroxidase/kieselguhr co-modified pyrolytic graphite electrode. AB - A reagentless nitric oxide (NO) biosensor was prepared using a pyrolytic graphite (PG) electrode modified with a composite film containing horseradish peroxidase (HRP) and kieselguhr. Noticeably, the electron-transfer reactivity of HRP was significantly enhanced when incorporated in the inorganic kieselguhr material. Consequently, we observed the direct electrochemical response of HRP in this composite film, which would be otherwise electrochemically silent (in the absence of kieselguhr). Importantly, this modified electrode demonstrated nice catalytic activity, as well as high stability, towards the reduction of NO. The peak current related to NO was linearly proportional to its concentration from 2.0 x 10(-7) to 2.0 x 10(-5) mol/L, and the relative standard deviation was 4 % for five successive determinations at a NO concentration of 1.0 x 10(-5) mol/L. The critical level in concentration was estimated to be (4.0 +/- 0.3) x 10(-8) mol/L. PMID- 15279106 TI - Effect of CO2 concentration in air on electrolytic conductivity of aqueous solutions of KCl. PMID- 15279107 TI - Much ado about nothing. Between last year's Medicare bill and the November elections, healthcare legislation may be high on drama, low on substance. AB - That sound you hear from Washington? It's the hum of talk about healthcare, and that's about all you can expect for the rest of the year-and no action. While Senate Majority Leader Bill Frist, left, has vowed to continue fighting for malpractice liability reform, "It's a tough lift this year," says AHIP's Karen Ignagni. PMID- 15279108 TI - Increased friction. Tax on ambulatory surgery centers aimed at boosting funds for N.J. hospitals. PMID- 15279109 TI - Stay out of Florida. State bans single-specialty facilities for cardiac, orthopedic, cancer treatment. PMID- 15279110 TI - Scoring big. Two major initiatives in works to gauge hospital value. PMID- 15279111 TI - Blues unite. BCBS of Arkansas, Florida to consolidate products. PMID- 15279112 TI - Rehab backlash. As new regs take effect, lobbying mounts for update. PMID- 15279113 TI - The unusual suspects. Whistle-blowers target contractor, Bear Stearns. PMID- 15279114 TI - Governance gap. Alaska hospitals try to improve CEO-board relations. PMID- 15279115 TI - Tough enough? Rules about drug firms' gifts get new look from docs. PMID- 15279116 TI - Mission unaccomplished. Hospitals need to act now to show that serving patients is their only business. PMID- 15279117 TI - On pricing, think wholesale. PMID- 15279118 TI - One step at a time. Preparation, orderly implementation are keys to achieving clinical, financial goals for information technology. PMID- 15279119 TI - A new degree of expertise. Organizations, universities helping doctors graduate to IT proficiency. PMID- 15279120 TI - Mixing policy and politics. As Kerry's domestic policy director, Sarah Bianchi blends campaign experience, healthcare expertise. PMID- 15279121 TI - California Substance Abuse Research Consortium, May 2003: update on recent substance abuse trends. AB - California Substance Abuse Research Consortium (SARC) meetings have become a mainstay in supporting alcohol and other drug (AOD) information exchange on research to policy and other initiatives within the state. One integral component of SARC is discussion of regional data on emerging and changing substance abuse trends. This article provides highlights from the substance abuse epidemiology portion of the May 2003 meeting, as presented for the San Francisco Bay Area, Fresno County, Los Angeles County, and San Diego County. PMID- 15279122 TI - Pilot-testing a statewide outcome monitoring system: overview of the California Treatment Outcome Project (CALTOP). AB - Timely information provided by an effective outcome monitoring system (OMS) is key to making improvements in treatment program effectiveness, service provision, and client outcomes. The California Treatment Outcome Project (CalTOP) developed and pilot-tested an automated outcome monitoring system for California's alcohol and other drug (AOD) system of care. CalTOP was designed to track client movement through treatment programs, measure standardized assessment of client service needs, record service utilization, assess treatment outcomes and client satisfaction, and determine the extent to which treatment produces cost-offsets in other health and social service systems. Information collected by CalTOP revealed that client problem severity at admission was high, services needed were diverse, and treatment services were generally not well matched to the level of problem severity or needs of clients. Also, client retention and length of stay in treatment were generally insufficient to maximize the potential benefits associated with treatment. This article presents the type of information on client demographics and treatment retention that was provided by CalTOP and outlines recommendations for implementing an AOD outcome monitoring system statewide. PMID- 15279123 TI - Diffusion of substance abuse treatment: will buprenorphine be a success? AB - The availability of buprenorphine promises to return the treatment of heroin addiction to mainstream medicine in the United States for the first time in nearly a century. This new treatment also provides an opportunity to reflect on the introduction of methadone and LAAM and their subsequent successes and failures in clinical implementation. The three articles that follow in this issue highlight the potential pharmacological, clinical, and logistic advantages of buprenorphine and provide clinicians with guidelines for its use in an integrated treatment setting. The clinical success of buprenorphine, however, depends in large part on factors beyond its pharmacological advantages. Clinician attitude and the extent to which they embrace buprenorphine will play an important role in determining the future success of this medication. PMID- 15279124 TI - Buprenorphine in the treatment of opiate dependence: its pharmacology and social context of use in the U.S. AB - Buprenorphine's physiological effects are produced when it attaches to specific opiate receptors that are designated mu, kappa, or delta. Buprenorphine, a partial agonist at the mu receptor and an antagonist at the kappa receptor, produces typical morphine-like effects at low doses. At higher doses, it produces opiate effects that are less than those of full opiate agonists. Knowledge of the physiological effects of opiate receptors and the way they interact with opiate agonists, partial opiate agonists, and opiate antagonists is fundamental to understanding the safety and efficacy of buprenorphine in treatment of pain and opiate addiction. Knowledge of the historical and social context of opiate agonist treatment of opiate dependence is fundamental to understanding how nonpharmacological factors may limit the clinical adoption and utility of a safe and effective medication in treatment of opiate dependence. This article reviews the pharmacology of sublingual buprenorphine and the historical context of opiate agonist therapy; delineates classes of opiate receptors and their interaction with opiate agonists, partial agonists, and antagonists; and describes the commercially available pharmaceutical formulations of buprenorphine. It focuses on sublingual buprenorphine tablets, Subutex and Suboxone, the FDA-approved formulations of buprenorphine for treatment of opiate dependence. Sublingual buprenorphine, and the combination of sublingual buprenorphine/naloxone, have unique pharmacological properties that make them a logical first-line intervention in the treatment of opioid dependence. PMID- 15279125 TI - Evolving use of buprenorphine in the treatment of addiction. AB - Two formulations of buprenorphine were approved in the United States for treatment of opioid dependence in 2002. This newly available treatment offers a safe and effective alternative to addicted individuals who are not currently in treatment. This article focuses on the steps that physicians and patients may take, as of this writing, if they should wish to participate in this new treatment. The content is clinically oriented and intelligible to an audience that is not medically trained. In the article, buprenorphine is placed in context of the standard opioid pharmacotherapy of methadone maintenance, and the expansion of opioid pharmacotherapy into the office setting is described. PMID- 15279126 TI - Counseling buprenorphine patients: information and treatment approaches for counselors. AB - The Drug Addiction Treatment Act of 2000 allows qualifying physicians to prescribe buprenorphine out of office-based practices to treat opioid-dependent patients, with the requirement that they have the ability to refer them to ancillary counseling services. It is likely that these patients will be seen by a wide range of counselors with varying experience in treating addictions. In order to enable the spectrum of counselors working with buprenorphine patients to receive information regarding opioids, buprenorphine, opioid dependency, and relevant counseling approaches, an online course has been developed. The information and rationale provided in this course is summarized in this article, including: background information regarding opioids and buprenorphine; general issues in counseling patients with substance abuse disorders; and approaches to counseling buprenorphine patients. PMID- 15279127 TI - California drug courts: a methodology for determining costs and avoided costs. AB - A significant body of outcome evaluation research on drug courts exists; however, few studies have investigated the cost implications of these collaborative justice models. This study focuses on creating a sound research design that can be utilized for a statewide and national cost-assessment of drug courts by conducting an in-depth case study of three adult drug courts in California. A transactional costs analysis (TCA) approach was utilized, allowing the researcher to calculate costs based on every individual's transactions within the drug court and the traditional criminal justice system. This model allows for the identification of each agency's resource contribution to the system and their avoided costs due to system outcomes. Cost results in all three sites indicate that participation in drug court, regardless of graduation status; saves taxpayers significant money over time. Expenditure and savings varied considerably among the agencies involved. Some agencies, such as the Department of Corrections, contribute little to the drug court system but experience substantial costs avoidance due to a reduction in recidivism among drug court participants. In order to validate study results and test the research design, the TCA methodology will be applied in six additional courts in the second phase of the project. PMID- 15279128 TI - Alcohol and violence: connections, evidence and possibilities for prevention. AB - This article reviews a number of theoretical and substantive arguments and models concerning the link between alcohol and violent crime which have appeared in the research literature in the past decade. These arguments and models form a firm foundation for the expectation that alcohol plays a causal role in violent crime, and that interventions designed to reduce or eliminate this link between alcohol and violence have the potential to become effective violence prevention policies. Four studies on the relationship between alcohol and violence are summarized, including one in which a natural alcohol policy experiment is evaluated. Taken together, these studies provide substantial empirical evidence that alcohol policy can be an effective crime prevention tool. PMID- 15279129 TI - Evaluation of the substance abuse and crime prevention act: treatment clients and program types during the first year of implementation. AB - The Substance Abuse and Crime Prevention Act (SACPA) represents a major shift in criminal justice policy. Eligible offenders can now be sentenced to drug treatment instead of either supervision without treatment or incarceration. UCLA is conducting an independent evaluation of SACPA that will continue over a 5 1/2 year period ending June 30, 2006. Analysis of data collected during the first year of implementation provides information on the flow of offenders through the SACPA "pipeline" from the initial decision to participate through treatment entry. Also available are characteristics of SACPA clients entering treatment and information on programs treating SACPA clients. Results show that most SACPA eligible offenders chose to participate in SACPA and that almost two-thirds of those who chose SACPA went on to enter treatment. Also, compared to other treatment clients in California, SACPA treatment clients included fewer women, were older, were more likely to use methamphetamine, and had been using drugs for a longer period of time. Finally, most SACPA clients were referred to outpatient drug-free treatment, regardless of primary drug problem, and very few heroin users in SACPA were referred to methadone. Future reports will cover the possible cost-saving associated with SACPA, outcomes for SACPA clients, and overall lessons learned. PMID- 15279130 TI - When you assume...the reality of implementing a legally mandated substance abuse treatment program. AB - In 2000, the voters of California approved the Substance Abuse and Crime Prevention Act (SACPA), mandating that substance-abusing offenders who met the eligibility criteria receive treatment and not jail time. In preparation for implementation, the Santa Clara County Department of Alcohol and Drug Services Research Institute worked to identify the number and type of clients who would be eligible to receive treatment. The purpose of this article is to analyze the differences between a profile of SACPA-eligible clients that was based on pre SACPA implementation clients and assumptions that were made concerning eligibility for the program and the reality of post-SACPA implementation clients. This article examines causes and consequences of the discrepancies between policy and implementation in the area of substance abuse treatment. PMID- 15279131 TI - Standardizing evaluation of on-line continuing medical education: physician knowledge, attitudes, and reflection on practice. AB - INTRODUCTION: Physicians increasingly earn continuing medical education (CME) credits through on-line courses, but there have been few rigorous evaluations to determine their effects. The present study explores the feasibility of implementing standardized evaluation templates and tests them to evaluate 30 on line CME courses. METHODS: A time series design was used to compare the knowledge, attitudes, and reported changes in practice of physician participants who completed any of 30 on-line CME courses that were hosted on an academic CME Web site and a CME Web portal during the period from August 1, 2002, through March 31, 2003. Data were collected at baseline, at course completion, and 4 weeks later Paired t tests were used to compare the means of responses across time. RESULTS: U.S. physicians completed 720 post-tests. Quality of content was the characteristic of most importance to participants; too little interaction was the largest source of dissatisfaction. Overall mean knowledge scores increased from 58.1% to 75.6% at post-test and then decreased to 68.2% at 4 weeks following the course. Effect sizes of increased knowledge immediately following the course were larger for case-based than for text-based courses. Nearly all physicians reported making changes in practice following course completion, although reported changes differed from expected changes. CONCLUSIONS: Increases in physician knowledge and knowledge retention were demonstrated following participation in on-line CME courses. The implementation of standardized evaluation tests proved to be feasible and allowed longitudinal evaluation analyses across CME providers and content areas. PMID- 15279132 TI - Severe acute respiratory syndrome and the delivery of continuing medical education: case study from Toronto. AB - INTRODUCTION: Severe acute respiratory syndrome (SARS) struck Toronto in the spring of 2003, causing many deaths, serious morbidity, forced quarantine of thousands of individuals, and the closure of all provincial hospitals for several weeks. Given the direction by public health authorities to cancel or postpone all continuing medical education (CME) courses, including those sponsored by the University of Toronto Faculty of Medicine, SARS has had a profound effect on the delivery of CME in Toronto and beyond. METHOD: Case study design using existing documents and self-report. RESULTS: The immediate, specific response of the University of Toronto CME program to SARS is described for the period from March 2003 to September 2003. DISCUSSION: During major outbreaks of infectious disease, continuing education providers should maintain regular contact with public health authorities and learners, enact a rational process for postponing or canceling courses, and implement a disaster plan flexible enough to ensure the deliver, of education using technological advances. PMID- 15279133 TI - Assessing the health of future physicians: an opportunity for preventive education. AB - INTRODUCTION: Research shows that physicians who model prevention are more likely to encourage preventive behaviors in their patients. Therefore, understanding the health of medical students ought to provide insight into the development of health promotion programs that influence the way these future physicians practice medicine. A university-based General Clinical Research Center (GCRC) provides a venue well suited to the health assessment and education of medical students. This research explores the utility of a GCRC in a program measuring the prevalence of clinical risk factors and related health behaviors in first-year medical students. METHODS: A 6-year cross-sectional study of first-year medical students measured clinical and behavioral variables associated with metabolic syndrome. Statistical testing was used to determine the prevalence of risk factors and the influence of gender in these variables. RESULTS: This group of medical students displayed better health indicators than did the general young adult population; however a small proportion of medical students exhibited early risk factors for chronic disease. There were significant gender differences in mean values for clinical risk factors, with males displaying higher cardiovascular risk overall. Males and females demonstrated significant differences in dietary intake and exercise programs. DISCUSSION: A GCRC can be used to provide a health assessment of medical students. Moreover, some students may benefit from health promotion programs incorporated into medical school curricula. This study provides a foundation for further research on the health of future physicians and the development of health promotion programs for this population. It also begins to explore the use of a GCRC as a teaching resource for medical students. PMID- 15279134 TI - Technology-enabled knowledge translation: frameworks to promote research and practice. AB - Knowledge translation articulates how new scientific insights can be implemented efficiently into clinical practice to reap maximal health benefits. Modern information and communication technologies can be effective tools to help in the collection, processing, and targeted distribution of information from which clinicians, researchers, administrators, policy makers in health, and the public can benefit. Effective implementation of knowledge translation through the use of information and communication technologies, or technology-enabled knowledge translation (TEKT), would benefit both the individual health professional and the health system. Successful TEKT in health requires cultivation and acceptance in the following key domains: Perceiving types of knowledge and ways in which clinicians acquire and apply knowledge in practice. Understanding the conceptual and contextual frameworks of information and communication technologies applied to health systems, particularly the push, pull, and exchange communication models. Comprehending essential issues in implementation of information and communication technologies and strategies to take advantage of emerging opportunities and overcome existing barriers. Establishing a common and widely acceptable evaluation framework in order that researchers can compare various methodologies in their rightful contexts in TEKT research and adoption. Achieving harmony and common understanding in these areas will go a long way in fostering a fertile and innovative environment to encourage research and advance understanding in this exciting domain of TEKT. PMID- 15279135 TI - Setting quality standards for web-based continuing medical education. AB - Continuing medical education (CME) on the Internet has grown steadily over the past several years. However, the quality of Web-based CME has received limited attention in the medical literature, and there have been few attempts to articulate quality standards. This article describes five sets of standards published in the distance education literature and explores whether the standards might be used to inform and enhance approaches to designing and delivering Web based CME programs. Standards synthesize practical knowledge, best practices, and research findings. They vary in their perspectives on quality, fall short of being comprehensive, and convey many elements that apply to Web CME. We conclude that published standards in the distance education literature can provide valuable guidance to Web CME providers, and there is a clear need for additional research into questions about what works in Web-based education and why. PMID- 15279136 TI - Build-a-case: a brand new continuing medical education technique that is peculiarly familiar. AB - An observation at a problem-based learning, case-building meeting prompted the realization that building cases might itself be an effective educational intervention. We developed a process for a new continuing medical education technique that is peculiarly familiar that we call "build-a-case." Build-a-case has now been used for teaching and learning in many clinical situations and with several kinds of health professionals. Subjective evaluations of the approach are consistently positive, and people feel that they learn from it because it prompts the discussion of practice as it is in their clinics and communities. In what follows, we describe the build-a-case process and our experiences with it and suggest several theoretical constructs that might be useful in promoting thoughtful research on what may become a useful new tool for continuing education. PMID- 15279137 TI - The contribution of hospital library services to continuing medical education. AB - Much of the literature relating to continuing medical education programs laments the lack of effectiveness of traditional lecture-based format, the most often used method of presentation in hospitals. A gap exists between the content taught in lectures and the application of that knowledge in actual patient care. The services of the medical librarian, already employed in most hospitals, can help ameliorate this problem. Further, libraries help to support quality improvement efforts. These three functions (library services, continuing medical education, and quality improvement) are interdependent. Each lends strength to the other, and, ideally, all are coordinated within the hospital structure. PMID- 15279138 TI - [Assessment of two DNA extraction methods to amplify the pneumolysin gene (PLY) from blood culture samples of Streptococcus pneumoniae]. AB - BACKGROUND: Streptococcus pneumoniae is a common etiologic agent of invasive respiratory infections among children under 5 years of age and older adults. Isolation rates of S. pneumoniae by traditional culture techniques are low. AIM: To study the sensitivity and specificity of two different DNA extraction methods to amplify the ply gene, applied to three different types of blood culture broths, experimentally inoculated with S. pneumoniae. MATERIAL AND METHODS: DNA was extracted from the cultures using an organic method or a technique that consists in dilution, washing with NaOH and concentration of the sample. This was followed by PCR amplification of a 355 pb fragment of the pneumolysin gene (ply). RESULTS: The organic DNA extraction method inhibited the PCR reaction at all concentrations studied (0.6 to 10(6) colony forming units/mL). Using the NaOH extraction, ply gene amplification was positive in all three blood culture broths, but only at concentrations of 10(3) colony forming units/mL, or higher. Using the same DNA extraction method, PCR was negative when the broths were inoculated with seven other related bacterial species, which results in a 100% specificity. CONCLUSIONS: Detection of S. pneumoniae by amplification of ply gene from blood cultures using the protocol of NaOH for DNA extraction is specific and provides results in a short lapse. However, the diagnostic sensitivity is not optimal, which limits its clinical use. PMID- 15279139 TI - [Hereditary nonpolyposis colorectal cancer: surgical treatment and pedigree analysis]. AB - BACKGROUND: Hereditary nonpolyposis colorectal cancer (HNPCC) accounts for 3 to 5% of all colorectal cancer (CC). It is an autosomal dominant syndrome with 80% of penetrance for this disease. AIM: To analyze the pedigree and surgical treatment of HNPCC. PATIENTS AND METHODS: We retrospectively analyzed our database of CC selecting patients with HNPCC according to clinical criteria (Amsterdam II). We characterized our patient's pedigrees with telephonic interviews. RESULTS: From 1111 patients operated on with CC we identified 13 (1.17%) with HNPCC. The mean age at diagnosis was 41.6 years (range: 23-75). Sixty two percent presented in International Union Against Cancer (UICC) stages I or II and none in stage IV. Seventy one percent of tumors were proximal to splenic flexure. In 5 patients the diagnosis of HNPCC was made postoperatively, after diagnosis of CC in their relatives. In all but one of the 8 patients with preoperative diagnosis of HNPCC, we performed a total colectomy. From the remaining 6 patients with partial colectomy, 2 developed metachronic CC. Two patients died of cancer. From 101 persons in the 4 families, 25 have developed neoplasia: 18 CC, 3 endometrial cancer and 4 other tumors. Twenty eight relatives were eligible for colonoscopic screening, but only 21% of them have been screened appropriately. CONCLUSIONS: Preoperative diagnosis should change the surgical treatment of HNPCC, preventing metachronic disease. Primary colonoscopic screening allowed us to diagnose CC in early stages, nonetheless most of eligible relatives have not followed recommended frequency for colonoscopy. PMID- 15279140 TI - [Serotypes and antimicrobial susceptibility of Streptococcus agalactiae]. AB - BACKGROUND: Streptococcus agalactiae or group B streptococcus, GBS, is the leading cause of neonatal and maternal infections and an opportunistic pathogen in adults with underlying disease. In the last decade, a dramatic increase in the resistance of this microorganism to erythromycin and clindamycin has been observed. AIM: To determine the serotype distribution and antimicrobial susceptibility of isolates of S. agalactiae collected from infections and colonization and to assess the genetic mechanisms of macrolide and clindamycin resistance. MATERIAL AND METHODS: A total of 100 GBS isolates were collected between 1998 and 2002, in Santiago, Chile. They were isolated from the amniotic fluid from patients with premature rupture of membranes (7 isolates), blood from neonatal sepsis (10 isolates), neonate colonizations (2 strains), skin and soft tissue infections (7 isolates), urinary tract infections (5 isolates), genital infections (3 isolates), articular fluid (one isolate), and 65 strains were recovered from vaginal colonization55. RESULTS: Serotypes Ia, II and III were the predominant serotypes identified in our study, accounting for 90 (90%) of the strains. Five isolates belonged to serotypes Ib (5%) and two (2%) to serotype V respectively; no strains belonging to serotype IV were found. All strains were susceptible to penicillin G, ampicillin and cefotaxime, and four isolates (4%) were resistant to both erythromycin (MIC >64 microg/ml) and clindamycin (MIC >64 microg/ml). The strains had a constitutive macrolide-lincosamide-streptogramin (cMLSB) resistance phenotype and the erm(A) gene was present in the four isolates. CONCLUSIONS: Serotypes Ia, II and III were the predominant serotypes in this study. All strains were susceptible to penicillin G, ampicillin and cefotaxime, and four (4%) strains were resistant to both erythromycin and clindamycin. The cMLSB resistance phenotype, and the erm(A) gene was detected in resistant strains. PMID- 15279141 TI - [Immediate results of palliative surgery for different forms of univentricular heart using the Norwood procedure]. AB - BACKGROUND: Norwood procedure is used as the first stage in the palliative treatment of the hypoplastic heart syndrome and can be used, with some technical modifications, in other forms of univentricular heart with aortic stenosis or hypoplasia. These patients have a high mortality (50%), derived from the procedure itself and from their abnormal physiological status. AIM: To report our experience with the Norwood procedure. PATIENTS AND METHODS: Retrospective analysis of all patients subjected to the Norwood procedure between February, 2000 and June 2003. RESULTS: Thirteen patients (9 females, age range 5-60 days and median weight of 3.3 kg) were operated. Eight had hypoplastic heart syndrome and five had a single ventricle with aortic arch hypoplasia. The diagnosis was done in utero in eight patients. All technical variations, according to the disposition and anatomy of the great vessels, are described. Cardiac arrest with profound hypothermia was used in all and regional cerebral perfusion was used in nine. Three patients died in the perioperative period and three died in the follow up (two, four and 10 months after the procedure). Gleen and Fontan procedures were completed in five and one patients, respectively. CONCLUSIONS: Our results with the Norwood procedure are similar to other series. There is an important mortality in the immediate operative period and prior to the Glenn procedure. PMID- 15279142 TI - [Clinical and epidemiological profile of intestinal intussusception among infants of Metropolitan Santiago]. AB - BACKGROUND: Intussusception (IS) is a potentially severe disease that affects an undetermined number of Chilean infants. The withdrawal of a rotavirus vaccine in 1999 due to its association with IS, highlighted the need for updated information on IS worldwide including Chile, before introduction of new vaccines. AIM: To estimate the incidence and to describe the epidemiology and clinical presentation of IS in the Metropolitan Area of Chile. MATERIAL AND METHODS: IS cases occurring between 1996 and 2001 in the seven public pediatric hospitals and in six private clinics (during 2000 and 2001) were identified. Incidence rates were calculated using updated population estimates. A systematic review of the medical charts of IS cases occurring in the public hospitals for 2000-2001 was performed. RESULTS: IS incidence rates for the Public Sector ranged from 32 to 39 per 100,000 children < 2 years of age. These figures did not vary significantly among the different Health Care Services, nor after inclusion of the private clinics. IS was more common in males (66%) and infants younger than 12 months (83%), with 67% of cases occurring between 3 and 8 months of age. The most common presenting symptoms were abdominal pain (90%), vomiting (86%), and rectal bleeding (75%). Ileocolic IS predominated (83%) and surgical correction was the preferred treatment (81%). No death occurred in this series. CONCLUSIONS: IS incidence rates were intermediate compared to other series, stable over time, and similar between the public and private sector. Clinical characteristics were similar to those previously reported with a disproportionately high use of surgical correction over enema, currently considered the preferred treatment option. PMID- 15279143 TI - [Biomedical impact of traveling for Chilean elderly]. AB - BACKGROUND: In Chile there is a program named "Vacations for Elderly during Low Season". AIM: To characterize participants of this program and to measure the impact of traveling in their health and wellbeing. MATERIAL AND METHODS: Two anonymous and voluntary questionnaires were applied to 4200 participants, before and after a ten days vacation package. RESULTS: Before traveling, questionnaires were answered by 802 subjects, and after traveling by 4057 (69% women, 22% older than 75 years old, 15.8% living alone). The presence and maintaining of good health were most appreciated at this age and 59% classified their health as good or excellent. Twenty five percent referred sensory problems (seeing or hearing), 12% reported urinary incontinence and 21% presented falls in the last three months; depression screening (GDS-5) was positive in 16%. Chronic disease prevalence was similar to the general Chilean elderly population. After traveling they reported significant improvements in the items sociability, wellbeing, mood, appetite, insomnia and ostheoarthritic pain. CONCLUSIONS: Elderly who traveled were mainly women, who thought that maintaining good health is the most precious value. After traveling they improved significantly different aspects of wellbeing. Promotion of this kind of recreation programs is an important tool for integration and enhancement of quality of life in elderly subjects in our country. PMID- 15279144 TI - [Recent changes in prostate cancer mortality in Chile. Trends analysis from 1955 to 2001]. AB - BACKGROUND: By the year 2000, prostate cancer became the second leading cause of cancer death in Chilean men of all ages and is the leading cause of cancer deaths in men of eighty years of age or older. AIM: To analyze the trends in mortality rates from prostate cancer in Chile in a fifty years series, estimating the rate of increase of such rates and their changes in time. MATERIAL AND METHODS: A trend analysis for age standardized mortality rates was performed, using join point regression analysis, which allows estimation of the annual percent change of rates and to find significant changes in such trend. RESULTS: Age standardized mortality rates in Chile reached their peak value in 1996, becoming apparently stable from then on. Crude rates have had a steady increase during the whole period. The trends analysis identified three different periods in the growth of the age standardized rates: a first one of slot increase in rates between 1955 and 1981 (0.9% annual increase), a second one of more aggressive growth starting in 1981 (2.6% annual increase), and a third period starting in 1996, in which rates slowly decline at an annual rate of 1%. CONCLUSIONS: The tendency of prostate cancer seen in Chile resembles that of industrialized countries, with an increase in its age standardized death rates that suffers a downturn by the end of the past decade. Besides early detection techniques, a substantial part of the reduction in mortality from prostate cancer could be explained by therapeutic improvements. PMID- 15279145 TI - [Bacteremic pneumococcal pneumonia in 45 immunocompromised hospitalized adults]. AB - BACKGROUND: The presence of bacteremia during a pneumococcal pneumonia is a sign of bad prognosis. AIM: To report a clinical experience with bacteremic pneumococcal pneumonia. PATIENTS AND METHODS: We reviewed the clinical and laboratory data from 45 adults (36 male, aged 17 to 97 years) with community acquired pneumonia (CAP) and Streptococcus pneumoniae bacteremia, hospitalized between January 1997 and August 2002 at the Puerto Montt Hospital (Southern Chile). RESULTS: Eighty four percent of patients bad underlying aggravating conditions, mainly alcoholism (40%), chronic obstructive lung disease (17.8%) and renal failure (17.8%). Seven percent were homeless. Fever, cough, dyspnea and sputum were the most common presenting symptoms. Five patients had pleural involvement. Four strains (8.9%) of S. pneumoniae had diminished susceptibility to penicillin. Nine patients died (case-fatality rate of 20%), but mortality was attributed to pneumonia in only three of them. Main factors associated with a higher mortality were renal failure, absence of cough, an arterial pH < 7.3 on admission, ICU hospitalization, shock, mechanical ventilation and an APACHE score > 16. CONCLUSIONS: The high death rate of these patients could be explained mainly by underlying conditions. ICU management and higher cost preventive measures could reduce this rate. PMID- 15279146 TI - [Prediction of the pressure ulcer development in elderly women using the Braden scale]. AB - BACKGROUND: Pressure ulcers are a common complication among elderly patients confined to bed for long periods. The Braden scale is a commonly used risk assessment tool. AIM: To evaluate the use of Braden scale. PATIENTS AND METHODS: Seventy women aged 61 to 96 years, admitted to the Internal Medicine Service of Barros Luco-Trudeau Hospital, were studied. Their risk was evaluated using the Braden scale. The presence of pressure ulcer was diagnosed according to the National Pressure Ulcer Advisory Panel on admission, two weeks later and at discharge. RESULTS: On admission, mean Braden scale score was 16.6+/-2.8 and 34 women had a score of 16 or less, that is considered of risk. Twenty five women (20 with a score of 16 or less) developed pressure ulcers, mostly superficial. The odds ratio of a score of 16 or less for the development of ulcers was 4.2 (95% CI 1.8-11.7, p <0.001). The sensitivity and specificity of such score were 80 and 69% respectively. CONCLUSIONS: The Braden scale predicts the risk of developing pressure ulcers with a good sensitivity and specificity in female elderly patients. PMID- 15279147 TI - [Albumin dialysis MARS (Molecular Adsorbent Recirculating System) as a bridge for liver transplantation in acute liver failure. Report of three cases]. AB - The most successful therapy for acute liver failure is liver transplantation. However, due to the low number of donors, organ support therapies need to be used as a bridge to liver transplantation. Molecular Adsorbents Recirculating System (MARS) is a dialysis treatment that uses a recirculating dialysate containing albumin. This allows the removal of both hydrosoluble and albumin-related substances. This system improves hepatic encephalopathy, renal dysfunction and some clinical parameters in acute liver failure, but there is no clear decrease in mortality. We report three women aged 23, 21 and 61 years, that were subjected to liver transplantation, in whom this therapy was successfully used. PMID- 15279148 TI - [Paroxysmal tachycardia in a patient with a permanent form of junctional reciprocating tachycardia. A case report]. AB - Junctional reciprocating tachycardia is an atrioventricular reentrant tachycardia whose anterograde conduction occurs via the His Purkinje and the retrograde conduction via an accessory pathway with slow conduction. The most common form is incessant tachycardia but a paroxysmal form also exists. We report a 35 years old female with recurrent paroxysmal tachycardia, that underwent electrophysiological evaluation. A left posterolateral accessory pathway was documented. Reciprocating paroxysmal tachycardia was induced by electrical stimulation and a successful pathway ablation was performed. PMID- 15279149 TI - [Recessive dystrophic epidermolysis bullosa. Report of one case]. AB - We report a female newborn with a dystrophic epidermolysis bullosa. The diagnosis was made by electron microscopy of a bullous skin lesion. The importance of reaching a specific diagnosis is underscored. Close relatives can therefore be informed and educated about prognosis, etiology and the possibility of having new affected offspring. An accurate diagnosis can be reached through electronic microscopy or modern immunohistochemical techniques. Further complementary information given by conventional histology is required. A complete study is recommended to minimize errors in the intepretation of morphology. PMID- 15279150 TI - [Integrons and resistance gene cassettes: structure and role against antimicrobials]. AB - Bacteria have developed sophisticated and successful genetic mechanisms to evade the action of antimicrobials. Bacterial multiresistance has caused serious problems in the treatment of nosocomial infections. Integrons and gene cassettes are considered the main genetic elements in the evolution of plasmids and transposons that actively participate in the mobilization of genes, codifying different bacterial resistance mechanisms. This article reviews the historical and structural aspects of integrons and resistance gene cassettes and the presence of these structures in gram negative bacteria isolated from Chilean hospitals in the last ten years. PMID- 15279151 TI - [Pancreatic neuroendocrine tumors]. AB - All pancreatic neuroendocrine tumors derive from Langerhans islet cells and have a low incidence. Half of them are functioning tumors that produce diverse hormones and occasionally cause serious clinical endocrine syndromes. They may be malignant, but they have a better survival, if compared to pancreatic ductal adenocarcinoma. Insulinoma, gastrinoma, glucagonoma, VIPoma (VIP=vasoactive intestinal peptide), somatostatinoma and ACTHoma are functioning tumors and they may also be part of Multiple Endocrine Neoplasia type I (MEN 1) syndrome and of von Hippel-Lindau disease. Diagnosis of non-functioning tumors is usually late, when they reach a big size and have even developed nodal and hepatic metastases. Nowadays, there are effective medical treatments for the medical problems secondary to excessive hormone production. For example, the hypergastrinemia typical of the Zollinger-Ellison syndrome in gastrinoma, can be adequately managed. Surgical resection is the most advisable therapy for pancreatic endocrine tumors, especially when they are small, when long time survival is better. Pre and intra operative imagenology is a great aid to locate these tumors. There are several surgical alternatives, according to the tumor size and location within the pancreas. Furthermore, palliative therapy can be used in disseminated disease. Treatment success is the result of a multidisciplinary medical team work of endocrinologists, surgeons, gastroenterologists, pathologists and geneticists. PMID- 15279152 TI - [Comfort and satisfaction of physicians with their medical practice in a reforming health system]. AB - The subjective wellbeing of physicians and the degree of dissatisfaction with their practice is nowadays an important issue in English speaking journals. There is an international perception of a growing and deepening crisis in health systems that is affecting the professional and personal life of physicians. A multidisciplinary group, from two main Universities in Chile, has been formed to investigate this situation locally. The results of this investigation will be published in successive issues of Revista Medica de Chile. PMID- 15279153 TI - [Changes in the environment of medical practice]. AB - Changes in medical practice have been very deep in the last 50 years. These changes have happened together with other external variations in the economy, demography, culture and information, generating a sense of uneasiness and unhappiness among the medical profession. This problem is visible when interviewing clinicians in a wide range of medical settings, independently of their contractual arrangement. The medical work model in Chile has changed from a situation in which there was a sole employer, the National Health Service, one single training school, the University of Chile, and a small minority of powerful and influential intellectual leaders. At the beginning of the XXI century, there are several modes of medical work, distant and unconnected among them. On the other side of the problem many factors add complexity, such as the demographic and epidemiological transition, technological advance, the dual rich-poor medical care, the emergence of malpractice procedures and the new, better informed and more demanding medical consumer. On top of this, different health care systems reform initiatives appear, providing more uncertainty to a nearly unbearable situation. This bibliographic review shows us that similar feelings are experienced in advanced countries and in developing societies such as Chile. PMID- 15279154 TI - [A proposal for training in hematology, in Chile]. PMID- 15279155 TI - [Hormones of the cardiovascular system]. PMID- 15279156 TI - [Change of the antioxidant status and free radicals production in hemolymph of Galleria mellonella larvae in microsporidiosis]. PMID- 15279157 TI - [Activity of antioxidant enzymes in coelomocytes of Eupentacta fraudatrix]. PMID- 15279158 TI - [Heterogeneity of olfactory transduction mechanisms in the frog Rana temporaria]. PMID- 15279159 TI - [Regulatory effect of magnesium ions on prolactin and somatotropin interaction with receptors of granulosa cells in Bos taurus]. PMID- 15279160 TI - [Affect of peptide and non-peptide opioids on the defensive reaction of the cockroach Periplaneta americana in the "hot chamber"]. PMID- 15279161 TI - [On some peculiarities of the phonotaxis selective mechanisms in the cricket Gryllus bimaculatus]. PMID- 15279162 TI - [Temperature affect on bringing up a first hatching by females of the ant Lasius niger (Hymenoptera, formicidae): the latitude variation of a reaction norm]. PMID- 15279163 TI - [Dynamics of thyroid hormones during the early stages in the development of the sturgeon Acipenser guldenstadti]. PMID- 15279164 TI - [Autogenic somatomotor activation of the heart in growing rats]. PMID- 15279165 TI - [Hemodynamics of the microcirculation in the cerebral cortex of rats with nitrite methemoglobinemia]. PMID- 15279166 TI - [Development of the rhesus macaque (Macaca mulatta) during early ontogenesis artificially fed and brought up by man]. PMID- 15279167 TI - [Evolutionary advantage of vasopressin substitution for vasotocin in mineral metabolism of the mammals]. PMID- 15279168 TI - [Calcium binding proteins of the turtle pretectum (immunochemical study). ]. PMID- 15279169 TI - [Current problems of microorganisms physiology]. PMID- 15279170 TI - Electrochemical reduction of carbon dioxide on pyrite as a pathway for abiogenic formation of organic molecules. AB - A wide spectrum of electrode potentials of minerals that compose sulfide ores enables the latter, when in contact with hydrothermal solutions, to form galvanic pairs with cathode potentials sufficient for electrochemical reduction of CO2. The experiments performed demonstrated the increase of cathode current on the rotating pyrite disc electrode in a range of potentials more negative than -800 mV in presence of CO2. In high-pressure experiments performed in a specially designed electrochemical cell equipped with a pyrite cathode and placed into autoclave, accumulation of formate was demonstrated after 24 hr passing of CO2 (50 atm, room temperature) through electrolyte solution. The formation of this product started on increasing the cathode potential to -800 mV (with respect to saturated silver chloride electrode). The yield grew exponentially upon cathode potential increase up to -1200 mV. The maximum current efficiency (0.12%) was registered at cathode potentials of about -1000 mV. No formate production was registered under normal atmospheric pressure and in the absence of imposed cathode potential. Neither in experiments, nor in control was formaldehyde found. It is proposed that the electrochemical reduction of CO2 takes part in the formation of organic molecules in hydrothermal solutions accompanying sulfide ore deposits and in 'black smokers' on the ocean floor. PMID- 15279171 TI - Prebiotic adenine revisited: eutectics and photochemistry. AB - Recent studies support an earlier suggestion that, if adenine was formed prebiotically on the primitive earth, eutectic freezing of hydrogen cyanide solutions is likely to have been important. Here we revisit the suggestion that the synthesis of adenine may have involved the photochemical conversion of the tetramer of hydrogen cyanide in eutectic solution to 4-amino-5-cyano-imidazole. This would make possible a reaction sequence that does not require the presence of free ammonia. It is further suggested that the reaction of cyanoacetylene with cyanate in eutectic solution to give cytosine might have proceeded in parallel with adenine synthesis. PMID- 15279172 TI - Stepwise evolution of nonliving to living chemical systems. AB - Steps by which a nonliving chemical system could have transformed into a living system are described and discussed, assuming general features of Wachtershauser's chemo-autotrophic surface theory of the origin of life. Environmental species such as CO2 and H2S are proposed to have reacted to form a quasi-steady state metal-bound intermediate (CH3-M) that slowly decayed into waste (CH4). Unpredictable dispersive reactions expanded the system to include surface-bound forms of the citric acid cycle intermediates (oxaloacetate-->citrate). Further reaction yielded an autocatalytic system in which raw materials are converted into the system at exponential rates. Combinatorial dispersive reactions that improved the performance of this system were automatically selected and incorporated into it. Systems evolved critical features of living systems (proteins, membranes, proteins, nucleic acids, etc.) using two related mechanisms called grafting and waste-conversion. Such living systems were transformed from less recognizable types (characterized by autocatalytic spreading, decentralization, poorly defined boundaries, etc.) into more recognizable ones (encapsulated by membranes, controlled by single-molecule genomes, etc.) that self-replicated by a cell division cycle and could evolve by the standard gene based Darwinian mechanism. The resulting systems are viewed as having an autocatalytic network composed of three linked autocatalytic subreactions. PMID- 15279173 TI - Symmetry breaking by spontaneous crystallization--is it the most plausible source of terrestrial handedness we have long been looking for?--A reappraisal. AB - In the light of recent and controversial findings on spontaneous resolution of racemates and their implications in the origin of homochirality on earth, we present here a detailed review of this important topic. Although spontaneous resolution cannot at this moment be reliably predicted, there has also been considerable progress in crystal structure prediction and, not only thermodynamic factors, but also kinetic ones, play important roles in the efficiency of packing and crystallization. In addition, self-association and supramolecular control phenomena may be identified in cases where spontaneous resolution of enantiomers is actually occurring. While this contribution summarizes our current understanding of this intriguing phenomena, it is hoped that future work on crystalline conglomerates (or homochiral crystals) of prebiotic importance will be of further help to understand the general problem of terrestrial chirogenesis. PMID- 15279174 TI - On the classes of aminoacyl-tRNA synthetases, amino acids and the genetic code. AB - The division of the aminoacyl-tRNA synthetases in two classes is compared with a division of the amino acids in two classes, obtained from the AAIndex databank by a principal component analysis. The division of the enzymes in Classes I and II follows to a great extent a division in the chemical and biological properties of their cognate amino acids. Furthermore, the phylogenetic trees of Classes I and II enzymes are highly correlated with dendrograms obtained for their cognate amino acids by using the indices in the AAIndex database. We argue that the evolution of aminoacyl-tRNA synthetases was determined by the characteristics of their corresponding amino acids. We interpret these results considering models for the origin and evolution of the genetic code in which an initial version, containing fewer amino acids, was modified by the incorporation of new amino acids following duplication and divergence of previous synthetases and tRNA molecules. PMID- 15279175 TI - Microbial life and temperature: a semi empirical approach. AB - Many groups have examined the effect of temperature on the survival of microorganisms, resulting in the development of several models. Some of these models are based on the Arrhenius equation and the others are based on multidimensional response surface equations. We argue that the former are inadequate and the latter lack biological meaning. We show that an equation (the GLE equation) deduced from the Theory of Rate Processes is more accurate than the Arrhenius equation. The excellent standard deviation values of the apparent free energy of activation obtained with the GLE equation for microbial growth, embryogenic and other processes show that this equation is more suitable than the Arrhenius equation. The GLE equation shows how temperature affects survival. Thus, organisms survive longer at low temperatures than at normal temperatures. The recent discovery of microorganisms in Siberian permafrost samples that are several million years old, in deep oil fields, mines and other extreme habitats appears to be consistent with the GLE equation. Another example, the enhanced resistance of spores at extreme temperatures can be easily explained by their high apparent free energy of activation We also examined the implications of the GLE equation on food sterilization practices and on exobiology. PMID- 15279176 TI - Neoadjuvant chemotherapy of breast cancer. AB - The use of neoadjuvant chemotherapy in the treatment of breast cancer has been evolving during the past two decades. Fisher's group accomplished the scientific rationale in mouse models in the 1970s. The Milan group was the first to use neoadjuvant therapy in locally advanced breast cancer and also determined that adjuvant sequential doxorubicin with cyclophosphamide/methotrexate/fluorouracil (CMF) offered improved survival when compared to alternating CMF with doxorubicin. Other groups (M. D. Anderson and NSABP) have evaluated similar doxorubicin-based neoadjuvant therapies in locally advanced breast cancer (LABC) and in early disease. These studies have shown an increase in breast-conserving therapy (BCT) and an increase in pathologic complete response (pCR) when neoadjuvant therapy was used. Due to anthracycline resistance, taxanes were added to the doxorubicin-based neoadjuvant chemotherapy regimen with an increase in BCT and pCR than when an anthracycline regimen was used alone. Overall survival has yet to be determined when comparing an anthracycline-based regimen to the same regimen with the addition of a sequential taxane. Therefore, a combined treatment of an anthracycline/cyclophosphamide/taxane regimen is the recommended neoadjuvant chemotherapy for patients with breast cancer. PMID- 15279177 TI - The operative note as billing documentation: a preliminary report. AB - Certified professional coders from a multispecialty academic surgical practice used operative notes to identify 10 of the most common deficiencies for reimbursement of services. These 10 deficiencies were then used as evaluation criteria to audit the operative notes used as billing documentation. Twenty-four per cent of operative notes contained no deficiencies, whereas the remaining 76 per cent contained one or more audit criteria deficiencies. The three most common deficiencies identified included an incomplete description of all surgical procedures performed (56%), an inadequate description of the indications for procedures (49%), and only 45 per cent of the operative notes were dictated within 24 hours of the procedure. Thirty-nine per cent were dictated by faculty surgeons, whereas 61 per cent were dictated by surgical residents. Twenty-nine per cent of the operative notes that were dictated by faculty surgeons contained no deficiencies as compared with 20 per cent of the operative notes that were dictated by surgical residents. For a multispecialty academic surgical practice, the operative note is the document of justification for 75 per cent of revenue generated. We conclude that 1) the operative note represents the most important document for justification of reimbursement for surgical services, 2) surgeons should reassess the operative note as a billing document and provide the information necessary to expedite reimbursement, 3) surgical residents should be instructed in the details of an operative report as a billing document, and 4) most of the information needed in the operative note for billing purposes is simple and straightforward data that is important not only for reimbursement but also from a medico-legal and medical records standpoint. PMID- 15279178 TI - Surgeon-performed ultrasound in the management of thyroid malignancy. AB - Surgeon-controlled real-time ultrasound (US) is a new adjunct in the management of patients with thyroid malignancy. The introduction of US as a routine evaluation tool has increased the recognition of nonpalpable thyroid cancers and cervical lymph node metastases. We report our experience and the change in management of patients with thyroid cancer due to the use of US. We reviewed the records of all patients undergoing neck operations for thyroid cancer since 2002. US was performed by a surgeon preoperatively in all patients and intraoperatively when non-palpable cervical lymph nodes were present. Suspicious nonpalpable thyroid nodules underwent US-guided fine-needle aspiration (FNA) for cytology. Seventy-two patients underwent operations for thyroid cancer. US influenced the management in 57 per cent (41/72) of patients. US was useful in 1) identification and guidance for the FNA of nonpalpable cancers in 28 per cent (20/72), 2) identification of nonpalpable nodules in the contralateral lobe in 38 per cent (27/72), 3) preoperative diagnosis of nonpalpable metastatic lymph nodes in 24 per cent (17/72), and intraoperative guidance for their excision. Surgeon performed US changed and enhanced the pre- and intraoperative management in more than half the patients with thyroid cancer. PMID- 15279179 TI - Twenty-one cases of aortoenteric fistula: lessons for the general surgeon. AB - We retrospectively reviewed our experience from 1984 to 2001 with 21 cases of aortoenteric fistula (AEF) in 19 patients. The majority of cases were in men (13 of 19, 68%). One AEF was spontaneous, the other 20 developed after prior vascular reconstruction (95%). The majority of AEF were duodenal (48%) followed by small bowel (38%), colon (10%), and esophageal AEF (5%). The proximal anastomosis of the prior vascular repair was the site of AEF origin in 62 per cent of cases, the distal anastomosis accounted for 19 per cent, and the body of the graft for 14 per cent. The intestinal repair was chosen on a case-by-case basis by the general surgeon and consisted of a simple primary repair in 48 per cent, resection with primary anastomosis in 38 per cent, and patching with pleura or omentum in individual cases. Colostomies were created in the two cases with colonic AEF. The duodenum was excluded in one of 10 duodenal AEF. Six patients (32%) died in the 90 days following surgery. The biggest risk of postoperative death was presentation with sepsis (P = 0.069); interestingly, women were more likely to present with sepsis (P = 0.019) and experienced a disproportionate rate of postoperative death (male 23%, female 50%, P = 0.24). The method used to repair the bowel was linked to a higher rate of postoperative death, and patients that required bowel resection died more frequently (66%) than those who had a simple repair (10%, P = 0.07). Overall mortality with AEF remains high despite routine SICU care. The biggest risk for death is preoperative sepsis. Women presented with sepsis more frequently than men. The method of bowel repair appears to be related to overall survival and along with sepsis is, perhaps, a surrogate for the degree of erosion present at the site of the AEF. Simple bowel repairs were sufficient when technically possible. Duodenal exclusion is not an obligatory adjunct to duodenal repairs. PMID- 15279180 TI - The utility of a rapid parathyroid assay for uniglandular, multiglandular, and recurrent parathyroid disease. AB - The rapid parathyroid hormone assay (rPTH) is an effective tool in minimally invasive resections of parathyroid adenomas. However, there are relatively few reports examining its utility in the full spectrum of parathyroid disease. The purpose of this study was to examine the utility of the rapid parathyroid hormone assay in uniglandular, multiglandular, and recurrent hyperparathyroidism. A retrospective analysis of all patients undergoing parathyroid resection from June 2001 to March 2003 was undertaken. All patients underwent preoperative localization studies. Rapid parathyroid hormone (PTH) levels were drawn immediately prior to and 10 minutes following parathyroid resection. A decline of > or = 50 per cent rPTH qualified as a successful/complete resection. Additional intraoperative samples were drawn as needed for patients with multiglandular disease. Of 46 treated patients who were examined (average age, 54 years), 30 had single, 12 patients had multigland disease, and 4 had recurrent/persistent hyperparathyroidism. Thirty-seven patients had primary hyperparathyroidism (32 with single and 3 with double adenomas; 2 with hyperplasia), 4 patients had secondary hyperparathyroidism, and 5 tertiary hyperparathyroidism. All procedures were considered successful, as every patient exhibited a postresection decrement in rPTH exceeding 50 per cent (average decrement, 91%). Although 2 patients exhibited a postoperative PTH increase exceeding 50 per cent of the preoperative value, all remained asymptomatic and eucalcemic (median follow-up, 21.5 months). The rPTH assay is an effective tool in determining success of parathyroidectomy in patients with primary uni- and multiglandular as well as recurrent hyperparathyroidism. It can be used to achieve minimally invasive neck dissections or guide surgical decision-making in more complex cases. PMID- 15279181 TI - Adenocarcinoid of the appendix: is right hemicolectomy necessary? A meta-analysis of retrospective chart reviews. AB - The purpose of this study was to determine the efficacy of simple appendectomy versus right hemicolectomy in the treatment of localized adenocarcinoid of the appendix. A literature review of retrospective chart reviews from 1966 to March 1, 2003, was performed. Outcomes of retrospective chart reviews were assessed on the basis of treatment modality. Meta-analysis of studies by determining odds ratios for appendectomy versus extended resection using the Hunter-Schmidt meta analytic method was performed. One hundred patients from 13 studies met inclusion criteria. Seven per cent failure rate with appendectomy alone and 10 per cent with extended resection were observed [OR 1.9 (0.6-5.8); association chi2 1.15, 1 df, P = 0.28]. Our data supports the use of appendectomy alone in localized cases of adenocarcinoid of the appendix provided there is no cecal involvement and the tumor's histology is low grade. PMID- 15279182 TI - Treatment of superior mesenteric and portal vein thrombosis with direct thrombolytic infusion via an operatively placed mesenteric catheter. AB - Acute superior mesenteric vein (SMV) and portal vein (PV) thrombosis can be a complication of hypercoagulable, inflammatory, or infectious states. It can also occur as a complication of medical or surgical intervention. Management of mesenteric and portal vein thrombosis includes both operative and nonoperative approaches. Operative interventions include thrombectomy with thrombolysis; this is often employed for patients who present with signs of peritoneal irritation. Nonoperative approaches can be either noninvasive or invasive. Treatment with anticoagulation has been shown to be efficacious, though its rate of recanalization is not as high as with intravascular infusion of thrombolytics. Intravenous catheterization and thrombolytic infusion has the advantage of direct pharmacologic thrombolysis of clot, with decreased infusion required and the possibility to carry out dilation or thrombectomy concurrently. We report the use of recombinant tissue-plasminogen activator (rt-PA) infusion via an operatively placed multi side-hole catheter/5-Fr introducer sheath into the right portal and superior mesenteric vein clot, inserted through a small jejunal vein, in a patient who presented with acute gangrenous appendicitis and thrombosis of the main portal trunk and superior mesenteric vein. A temporary abdominal closure was maintained until 36 hours after the start of infusion of the rt-PA. At this time venous system had normal flow, with complete recanalization of the right portal and superior mesenteric veins. PMID- 15279183 TI - Pseudoangiomatous stromal hyperplasia of the breast in two adolescent females. AB - The majority of breast lesions that present in adolescent girls are benign, with most being fibroadenomas. Rarely, a large and rapidly growing breast mass may be found to be the more recently described entity named pseudoangiomatous stromal hyperplasia (PASH). The pathogenesis of this lesion is attributed to hyperplasia of stromal myofibroblasts in response to hormonal stimuli. To date, PASH has rarely been described in adolescence. We describe the presentation of PASH in two adolescent patients. The first is a 12-year-old girl who found a mass in her left breast 3 months prior to presentation. An excision of an 11.5 x 10 x 3.5 cm lesion weighing 347 g was performed via breast-conserving incisions. The second patient is a 16-year-old girl who also had a rapidly enlarging left breast mass removed in a similar fashion. This mass measured 12 x 11 x 6 cm and weighed 460 g. Both tumors were noted to have a smooth capsule. Histologic appearance consisted of the typical features of PASH; fibrous stroma containing numerous anastomosing slit-like spaces; some compressed and others with discernible lumina. Both patients had complete excisions and have since not experienced recurrence. Each has had excellent cosmetic results with symmetrical breast development since their resection. Pseudoangiomatous stromal hyperplasia is a rare tumor that arises in the breast. These tumors may grow quickly and often are mistaken for fibroadenomas, phylloides tumor, or angiosarcoma. They must be resected with careful attention to resection around the capsule of the tumor with breast conservation as a goal. Long-term follow-up is necessary, as some have been reported to recur. PMID- 15279184 TI - Contralateral portal vein embolization for hepatectomy in the setting of hepatic steatosis. AB - Portal vein embolization is evolving as an important adjunctive tool in hepatic surgery. In select patients, preoperative hypertrophy of the future remnant liver via contralateral portal vein embolization decreases postoperative liver dysfunction. Hepatic steatosis is the most common liver parenchymal disorder in Western populations. Moderate and severe degrees of hepatic steatosis convey an increased risk of postoperative liver dysfunction following major hepatic resections, but no studies exist examining the role of preoperative portal vein embolization in patients with hepatic steatosis. In this manuscript, we review the indications for portal vein embolization currently supported by the literature and present a patient with moderate to severe steatosis who successfully underwent portal vein embolization and a subsequent major liver resection. PMID- 15279185 TI - Primary duodenal low-grade mucosa-associated lymphoid tissue lymphoma presenting with outlet obstruction. AB - Low-grade lymphoma arising in mucosa-associated lymphoid tissue (MALT) of the duodenum represents a very rare neoplasm. We report an unusual presentation of primary duodenal MALT lymphoma in a 78-year-old man. The patient initially presented with a suspected pulmonary embolus and was anticoagulated, which precipitated a major gastrointestinal hemorrhage. A large atypical ulcer with narrowing of the duodenum beyond the bulb was seen on endoscopy. Biopsies revealed atypical lymphoid cells. Abdominal CT scan revealed a mass in either the duodenum or head of the pancreas. An endoscopic retrograde cholangiopancreatography (ERCP) was performed, which revealed a normal pancreatic duct with a large calculus in the common bile duct, which was extracted after sphincterotomy. Elective surgery was planned for suspected lymphoma of the duodenum. The patient developed severe nausea, vomiting, and fullness after meals. The patient underwent pancreaticoduodectomy for a neoplastic mass causing duodenal obstruction. Pathological examination of the resected specimen revealed a low-grade B-cell lymphoma (MALToma) arising in the duodenum and invading the pancreas. Flow cytometry confirmed the phenotype typical of MALT lymphoma. Celiac, peripancreatic, pelvic, and cervical nodes were also involved with tumor. Bone marrow was also positive for metastasis. The patient was postoperatively treated with chemotherapy for stage IV disease. PMID- 15279186 TI - In situ femoral to popliteal bypass graft using superficial femoral vein to popliteal vein. AB - During the evolution of vascular surgery as a specialty, many conduits have been used to revascularize the lower extremities. Superficial veins and prosthetic materials make up the majority of materials used to bypass diseased segments of native artery. The deep veins of the thigh have also been reported as alternatives for arterial bypass. However, the use of the in situ superficial femoral and popliteal vein bypass has not been reported to our knowledge in current literature. We report a 79-year-old white female with lower extremity rest pain who underwent an in situ femoral popliteal bypass graft for limb salvage. PMID- 15279187 TI - Unusual cases of jaundice secondary to non-neoplastic bile duct obstruction. AB - Obstructive jaundice secondary to common bile duct stricture is most commonly attributed to malignancy. Here we present three unusual cases that mimicked carcinoma in presentation but were histologically diagnosed as benign inflammatory processes during operative care. The first case was attributed to obstruction-induced chronic pancreatitis secondary to Crohn's disease of the head of the pancreas, the second was due to sarcoidosis within periportal and extrahepatic biliary lymph nodes and distal common bile duct, and the third case was due to tuberculosis of biliary lymph nodes. All were successfully managed surgically, but potentially these patients may have been effectively treated pharmacologically, without the need for invasive surgical intervention, if an earlier diagnosis were available to the clinicians. A retrospective and comparative review of the data of each case demonstrated subtle clues such as multiple enlarged biliary lymph node involvement and only moderately elevated bilirubin levels that pointed toward possible nonmalignant processes. PMID- 15279188 TI - Subareolar injection of radioactive colloid for sentinel lymph node identification in breast cancer patients. AB - Sentinel lymph node biopsy (SLNB) is becoming the standard for staging the axilla in breast cancer patients in many institutions. The best method of injection is still questionable. The purpose of this study was to compare the results of SLNB using the peritumoral or the subareolar injection site. Between December 1997 and March 2000, we performed 100 SLNBs. Technecium-labeled colloidal human serum albumin was injected peritumorally (Group A, 31 patients; Group B, 31 patients) or subareolarly (Group C, 38 patients). Patent blue dye was given periareolarly (Group A) or peritumorally (Groups B and C). Preoperative lymphoscintigraphy was performed in all patients. SLNB was successful in 94 patients (94%). The identification rate improved from 80 per cent (first 25 patients) to 99 per cent (last 75). The subareolar injection of the colloid did not adversely influence the results of SLNB compared with the peritumoral injection (identification rate, 100% vs 97%; false negative rate, 6% vs 14%). The subareolar injection of colloid is a simple and at least as accurate technique as the peritumoral one. This technique can also improve the identification rate of SLNB for breast cancer patients. PMID- 15279189 TI - Changes in abdominal aortic aneurysm size after endovascular repair with Zenith, AneuRx, and custom-made stent-grafts. AB - The aim of this study was to determine the maximal aneurysm diameter (MAD), the total aneurysm volume (TAV), the intra-aneurysm vascular channel (IAVC), and total thrombus volume (TTV) and compare changes in those parameters during a 12 month time period. In addition, these parameters for three different endovascular grafts were compared. A retrospective review of 42 patients who had undergone endovascular aneurysm repair (EVAR) between July 1999 and March 2001, and without evidence of an endoleak or migration, was performed. The minimum follow-up in this group was 12 months. The three grafts deployed were Dacron-stainless steel bifurcated grafts with suprarenal fixation [Zenith; Cook, Inc. (n = 14)], Dacron stainless steel aorto uni-iliac grafts with suprarenal fixation [custom-made (n = 10)], and externally supported Dacron nitinol bifurcated grafts [AneuRx; Medtronic, Inc. (n = 18)]. Volumetric measurements were obtained from CT images performed preoperatively, at 1 month and 12 months thereafter, using a 3-D Magicview 1000 workstation (Siemens, Inc.). Regardless of the type of endograft, a significant change in MAD and TAV (P = 0.008), IAVC (P = 0.031), and TTV (P = 0.001) was observed over the 12-month postoperative period. Both maximum diameter and total aneurysm volume appear to reflect accurately successful aneurysm exclusion. We conclude that both two-dimensional, maximal aneurysm diameter and three-dimensional, total aneurysm volume accurately reflect changes in morphology after endovascular aneurysm repair. PMID- 15279190 TI - Pinch-off syndrome: case report and collective review of the literature. AB - Pinch-off syndrome (POS) occurs when a long-term central venous catheter is compressed between the clavicle and the first rib. The compression can cause transient obstruction of the catheter and may result in a tear or even complete transsection and embolization of the catheter. POS may be preceded by a finding of "pinch-off sign" on chest X-ray (CXR) films in which the catheter is indented as it passes beneath the clavicle. We performed a collective review of the 109 cases of POS in the medical literature and report 3 new cases. On average, POS occurs 5.3 months after the insertion of the catheter but has ranged from immediately after insertion to 60 months later. If the subclavian vein is used for access, then an upright CXR should be obtained after the procedure and periodically thereafter to rule-out POS. Treatment of POS is removal of the catheter. If the tip of the catheter has embolized, it can usually be retrieved percutaneously with a transvenous snare. POS can be prevented by using the internal jugular vein for access rather than the subclavian vein. PMID- 15279191 TI - Lymph node metastasis at the splenic hilum in proximal gastric cancer. AB - We performed splenectomy on patients with macroscopic advanced gastric cancer located at the proximal part of the stomach to achieve complete D2 lymphadenectomy. The aim of this study was to clarify the survival benefit of splenectomy in the treatment of gastric cancer. The clinical records of 225 patients who underwent total gastrectomy with splenectomy for gastric cancers involving the proximal part of the stomach were analyzed retrospectively. Nodal involvement at the splenic hilum (no. 10) was detected in 47 cases (20.9%). All of these cases were macroscopically diagnosed as positive for serosal invasion or regional lymph node metastasis at the time of surgery. In considering the lymphatic pathway from the primary tumor to no. 10 lymph nodes, metastasis at lymph nodes along the lesser curvature (no. 3), the short gastric vessels, or the gastroepiploic vessels (no. 4) may be good indicators of no. 10 lymph node metastasis. The overall survival of 47 patients with positive no. 10 lymph nodes was extremely poor. However, when curative surgery was performed, the survival of no. 10 positive patients was not different from that of no. 10 negative patients. Thus, for patients with advanced gastric cancer located in the proximal part of the stomach, D2 lymphadenectomy with splenectomy is recommended when patients show macroscopic evidence of serosal invaded tumor with regional lymph node metastasis. PMID- 15279192 TI - Primary malignant melanoma of the right colon. AB - The small and large intestines are the most common sites for metastases from cutaneous malignant melanoma. However, primary melanomas in these sites are exceedingly rare. There are several case reports of patients with primary melanoma of the small bowel, but finding of a solitary primary melanoma in the colon is exceedingly rare. We describe a patient that was operated on for bowel obstruction due to colonic intussusception resulting from a right colonic tumor. Histopathological examination confirmed a diagnosis of malignant melanoma. A thorough postoperative investigation did not reveal a primary lesion in any other site. Two years after surgery, there was no evidence for recurrent disease. The treatment and prognosis of metastatic and primary melanoma of the gastrointestinal tract is discussed as well as the embryonic base for development of primary malignant melanoma of the intestine. Primary malignant melanoma of the intestine is an extremely rare lesion that may arise in the large bowel as well. It must be differentiated from other intestinal tumors and mandates a thorough investigation to rule out the possibility of being a metastasis from another more common primary site. PMID- 15279193 TI - The open peritoneal cavity: etiology correlates with the likelihood of fascial closure. AB - The use of laparostomy in damage control surgery and uncontrolled intra-abdominal infection has been well described. We examined 71 patients who required laparostomy to see if trends in management and outcome could be identified based on the underlying disease state. The underlying etiology included gastrointestinal sepsis (n = 25), pancreatitis (n = 21), or trauma (n = 25). Pancreatitis patients required more operations per patient (P < 0.05). The likelihood and type of closure (fascial, mesh, or none) was related to the underlying etiology: trauma patients were more likely to have fascial closure (P < 0.02), patients with GI sepsis were more likely to require mesh closure, and pancreatitis patients were more likely to have no formal closure (P < 0.02). Only 29 per cent of patients achieved definitive fascial closure. Mortality in trauma patients was 20 per cent, 36 per cent for GI sepsis, and 43 per cent in patients with pancreatitis. Complications of laparostomy included enterocutaneous fistula (16.9%) and abscess formation (7%). Though the use of laparostomy has become more prevalent, it is still associated with significant hospital stay, morbidity, and mortality. In our study, the number of operations and likelihood of abdominal closure appears to correlate with the etiology of the underlying disease requiring use of laparostomy. PMID- 15279194 TI - [Recent trends of diabetic neuropathy]. PMID- 15279195 TI - [Central post-stroke pain in Wallenberg syndrome]. AB - We analyzed clinical characteristics of central post-stroke pain (CPSP), constant and burning pain, in 32 patients with Wallenberg syndrome due to infarction. CPSP developed in 44% (14/32) of the patients; 8 in acute stage (within 4 days after the stroke) and 8 in chronic stage(10-120 days after the stroke). Apart from one exceptional case, CPSP was present in the hypalgesic side of face and extremities. In 9 cases of typical type (Currier's distribution of sensory disturbance), CPSP occurred in the ipsilateral face to the lesion during acute stage. Among them, 2 cases developed severe lancinating pain in chronic stage. In 5 cases of ventral type, it occurred in the contralateral extremities during chronic stage. Spino- and trigemino-thalamic tract are injured but medullary reticular formation is intact in Wallenberg syndrome. It is, therefore, considered that CPSP in Wallenberg syndrome is caused by denervation sensitivity of "paleo-reticulothalamic" tract within the reticular formation. PMID- 15279196 TI - [Examination of NIRS for evaluation of frontal lobe function in diffuse axonal injury (DAI) patients]. AB - The purpose of this study is to compare diffuse axonal injury (DAI) patients with healthy controls by using near infrared spectroscopy (NIRS). The Wisconsin Card Sorting Test Keio Version (KWCST), a standard task paradigm to detect human frontal lobe dysfunction was set as a method. The result of the examination showed that compared with DAI patients, wider increase of total hemoglobin was admitted in the frontal part of the brain of healthy people during the KWCST. This suggests that NIRS would serve as an objective indicator to evaluate the frontal lobe function in DAI patients. PMID- 15279197 TI - [Immunosuppressive-associated encephalopathy in bone marrow transplant recipients]. AB - We studied clinical features of immunosuppressive (cyclosporine, tacrolimus) associated encephalopathy in bone marrow transplant patients. 378 cases of allogeneic bone marrow transplant recipients over fifteen years old of chronic and acute leukemia (CML, ANLL, ALL) (n = 311), myelodysplastic syndrome (MDS) (n = 42) and severe aplastic anemia (SAA) (n = 25) were investigated. Immunosuppressive associated encephalopathy occurred in 12 cases. The rate of incidence was significantly higher in SAA and MDS (7 cases) than in leukemia. The cases which showed typical radiological abnormality in MRI were limited in SAA and hypoplastic MDS. 10 cases died, which revealed worse than an overall survival rate of recipients without immunosupressive-associated encephalopathy. 5 of 7 cases in SAA and MDS had taken cyclosporine as treatment of the disease before bone marrow transplantation and that might influence the incidence of encephalopathy. PMID- 15279198 TI - [Visual search training for a case of homonymous field defect with multiple visual dysfunctions]. AB - A 72-year-old right handed man developed right homonymous hemianopia without macular sparing, left homonymous lower quadranopia with macular sparing, cerebral amblyopia, cerebral achromatopsia, impaired form vision, and mild right hemispatial neglect, after multiple cerebral infarctions, involving bilateral occipital cortices. His intelligence and memory were deteriorated moderately. He failed to notice objects located in the affected visual field, because of his severely impaired visual search. When ordinary lighting was used, he showed severe right-sided omissions on the line cancellation test. However, omissions were less marked under the brighter lighting. By using a modified method of Kerkhoff and Vianen (1994), he was trained to make saccadic eye movements toward affected regions to find a target and to search and point at targets arranged randomly. As the sensitivity for contrast of isoluminante red and green stimuli was preserved well at high spatial frequencies despite the decreaced contrast sensitivity for brightness, we used green targets as the training stimuli. After the training, search field and pointing range that could be covered by the patient increased in size for both green and white targets, and daily activities improved. Moreover, after the training, he no longer showed discrepancy in line cancellation performances between ordinary and brighter lighting conditions. In the follow up period, the search field and the performance on the line cancellation test were maintained, while the performance of pointing targets array declined. The family members complained of mild re-deterioration of daily activities. Then, the training for searching and pointing re-introduced at home. After the training, his pointing performance and daily activities, evaluated by questionnaires to his family members, improved again. In conclusion, it was suggested that disordered visual search after a homonymous field defect can be treated effectively, even if multiple visual dysfunctions were associated. PMID- 15279199 TI - [A case of acute distigmine bromide intoxication in the therapeutic dosage for treatment of underactive neurogenic bladder]. AB - Distigmine bromide (Ubretid) is a long-acting anti-cholinesterase, widely used for the treatment of underactive neurogenic bladder and myasthenia gravis. Our study concerns a 73-year-old man treated with a potentially life-threatening cholinergic state due to distigmine bromide. He had been administered distigmine bromide orally for over two years at a daily dosage of 10 mg as a treatment for underactive neurogenic bladder. He suddenly developed diarrhea and consciousness disturbance during treatment of his urinary tract infection. Bradycardia and miosis were noted. Blood examination revealed extremely low levels of the plasma cholinesterase activity. The condition was diagnosed as distigmine bromide intoxication. All cholinergic symptoms disappeared in several days after the administration of distigmine bromide was terminated. Cholinergic crisis due to overdosage with anticholinesterases is well known, and the myasthenic patients are usually supervised in the early stages of dosage regulation to guard against the possibility of cholinergic crisis. However the use of oral distigmine bromide, even in therapeutic doses for urinary retention, could result in cholinergic crisis. We therefore conclude that extreme caution must be used in administering distigmine bromide. PMID- 15279200 TI - [High-dose intravenous immunoglobulin in the treatment of sensory ataxic neuropathy with Sjogren's syndrome: a case report]. AB - We report herein a case of sensory ataxic neuropathy with Sjogren's syndrome (SS SAN) who became dramatically improved in response to high-dose intravenous immunoglobulin treatment (IVIg). An 81-year-old man began to feel numbness in his hands and feet in August 2002. Because he became unsteady and could not do skillfull movement, he was admitted to our hospital in May 2003. On neurological examination, all tendon reflexes were absent. His vibratory and position senses were severely impaired to knees and elbows. Touch, temperature, and pinprick sensations were mildly disturbed in a glove-stocking distribution. Coordination was clumsy in all limbs because of sensory loss. He had gait ataxia with Romberg sign. Nerve conduction study revealed that sensory nerve action potentials were absent. He was diagnosed as having SS-SAN because Schirmer test, Saxon test and both SS-A and SS-B antibodies were positive. Thereafter, Mg, 400 mg/kg daily for 5 days, was administered. His sensory impairment began to improve 2 days after Mg. Subsequently, he could walk steadily without ataxia. It is considered that IVIg may be an effective treatment for SS-SAN. PMID- 15279201 TI - [Rapid improvement of callosal edema by thiamine administration in Marchiafava Bignami disease: a case report]. AB - Marchiafava-Bignami disease (MBD) associated with chronic alcoholism is a fatal disorder characterized by demyelination of the corpus callosum. A 62-year-old Japanese man, a heavy drinker for his last over 10 years, was admitted to our hospital because of acute onset of speech disturbance. The first MR images showed abnormal signal intensity of the corpus callosum, which was a typical finding of MBD, but no signal abnormality on diffusion-weighted images. At three days after large doses of thiamine administration, MR studies revealed the disappearance of callosal high signal intensity. His symptom gradually improved, the pathogenesis and therapy of MBD were discussed. PMID- 15279202 TI - [A case of amyloid angiopathy mimicking multiple metastatic tumors]. PMID- 15279203 TI - [Transient arterial hyperintensity on fast fluid-attenuated inversion recovery images in hyperacute cardiogenic cerebral embolism]. PMID- 15279204 TI - [Transient lesion in the isolated cerebral cortex in a case with status epilepticus]. PMID- 15279205 TI - [An autopsy case of 54-year-old man with progressive dementia]. PMID- 15279206 TI - [The hot summer of 2003: heat stroke in Maastricht]. AB - Two male patients, 46 and 62 years of age, were brought to the emergency department on a hot summer's day. Both wore excessive clothing. The first patient had a temperature of 43 degrees C and was comatose. Heteroanamnesis indicated that he was suffering from schizophrenia. Although the prognosis seemed to be poor, his condition improved after treatment in intensive care, consisting of cooling and supportive treatment, but the patient had considerable permanent neurological impairment. The second patient had a temperature of 40.3 degrees C, was confused and had an atactic gait. He was cooled immediately and recovered swiftly without complications. Heat stroke is a life-threatening illness, which is defined as a body temperature above 40 degrees C and central nervous-system dysfunction. Heat stroke may be attended by many serious complications, including multi-organ failure and residual brain damage. Prompt recognition and rapid treatment, consisting of adequate cooling, are required. PMID- 15279207 TI - [The practice guideline 'Hormonal contraception' (second revision) from the Dutch College of General Practitioners; a response from the perspective of general practice]. AB - The second revision of the Dutch College of General Practitioners' practice guideline on hormonal contraception discusses the various methods of hormonal birth control and can help the general practitioner to inform the patient when choosing the most desirable method. A thorough anamnesis is necessary with regard to the increased risks for breast cancer, uterine cancer and cardiovascular diseases in women using hormonal contraception, especially in those who smoke. The guideline also discusses non-hormonal methods briefly. PMID- 15279208 TI - [The practice guideline 'Hormonal contraception' (second revision) from the Dutch College of General Practitioners; a response from the perspective of obstetrics & gynaecology]. AB - The Dutch College of General Practitioners' (NHG) guideline on hormonal contraception does not follow the WHO criteria for the use of oral contraceptives in contrast to the guideline of the Dutch Society of Obstetrics and Gynaecology. Contrary to the WHO criteria, the NHG guideline considers a blood-pressure measurement before starting with an oral contraceptive to be unnecessary. It also considers no form of migraine to be a contraindication for oral contraceptives. The NHG guideline further disclaims the (slightly) increased risk of developing breast cancer in women using oral contraceptives. It advises initiation of oral contraceptive use two weeks postpartum in non-breastfeeding women and six weeks postpartum in breastfeeding women, instead of the three weeks and six months, respectively, indicated in the WHO guideline. Lastly, the NHG guideline is too optimistic as to the reliability of oral-contraceptive use, as no distinction is made between efficacy and effectiveness. Such discrepancies between two Dutch guidelines can be detrimental to women's health care. The WHO criteria for contraceptive use may be a valuable tool to overcome differences of opinion so as to achieve a badly needed full consensus. PMID- 15279209 TI - [Bispectral analysis of the electroencephalogram: a new method for recording the level of consciousness during anaesthesia]. AB - Until recently, no measure was available that provided objective and reproducible information on the level of consciousness in patients under general anaesthesia. Several decades of research to find a reliable measure for determining the depth of anaesthesia has now led to the clinical introduction of the bispectral index scale (BIS), a parameter derived from the electroencephalogram. Implementation of the BIS-monitor in anaesthetic practice leads to a reduced use of hypnotic agents, a more rapid recovery phase and possibly a reduced incidence of awareness. PMID- 15279210 TI - [Bronchoscopy: new diagnostic and therapeutic technology, new indications]. AB - Over the last few years developments have been concentrated on the diagnostic and therapeutic properties of bronchoscopy. Autofluorescence and fluorescence bronchoscopy significantly enhance the detection rate of premalignant and early neoplastic endobronchial lesions. Unfortunately, this technique is hampered by a low specificity. Endobronchial ultrasound examination is the first tool that has enabled the bronchoscopist to get an impression of the tracheal wall beyond its epithelial surface including the mediastinal and the hilar structures. Currently, the complexity of the procedure prevents its widespread application. Intervention bronchoscopy involves mechanical and laser removal of processes that may impede the central airways. Electrocauterization and laser dissection are regularly used to remove intrabronchial tumour depositions. Airway stenting is indicated if there are stenoses caused by compression from abnormalities located externally to the airways. PMID- 15279211 TI - [Summary of the practice guideline 'Hormonal contraception' (second revision) from the Dutch College of General Practitioners]. AB - When choosing a method of contraception, a woman must consider the pros and cons of various methods together with her family physician. In this process, the doctor provides information on the advantages and disadvantages, while the woman decides. A sub-50 pill of the second-generation preparation is the oral contraceptive of choice. If the woman chooses a newly developed method of contraception, she must be carefully informed about the uncertainties with regard to reliability and safety. Oral contraceptives are absolutely contra-indicated in the following cases: a history of myocardial infarction, stroke (CVA), venous thromboembolism, a known coagulation-factor deficiency, breast or endometrial carcinoma or severe liver-function disorders. Non-hormonal methods of contraception are preferred in such a case. If there are two or more risk factors for cardiovascular disease, the doctor and the patient must consider the pros and cons of hormonal contraception. In this connection, stopping smoking is more effective than not using an oral contraceptive. A prescription for an oral contraceptive can be given without a physical examination, not even a measurement of the blood pressure; follow-up is only necessary in the case of side effects or questions. Progestagen-only contraceptives are absolutely contraindicated in the following cases: current venous thromboembolism, vaginal bleeding of unknown origin, progestagen-dependent tumours such as breast cancer, and severe liver function disorders. PMID- 15279212 TI - [Diagnostic image (195). A man with blue finger tips]. AB - A 42-year-old man had ischaemia of three fingers of his left hand caused by thromboangiitis obliterans (Buerger's disease). PMID- 15279213 TI - [Successful breastfeeding after reduction mammaplasty]. AB - OBJECTIVE: To determine if it is possible to successfully breastfeed after reduction mammaplasty with intact continuity between papilla and glandular tissue. DESIGN: Retrospective by use of a written questionnaire. METHOD: Between 1986 and 2000, 994 women under the age of 35 underwent a reduction mammaplasty at the Medical Centre in Leeuwarden, The Netherlands. They were all sent a questionnaire of which 585 were completed correctly and returned. RESULTS: Of the 585 women, 215 had had one or more children and 90 had started breastfeeding, 57 of whom were successful (63%). In non-operated women this figure is 69%. The advice to try to breastfeed was associated with a higher percentage of success than the advice not to try--78% and 42% respectively. CONCLUSION: It appears to be justified to encourage women who have had a breast reduction to attempt to breastfeed after the birth of a child. PMID- 15279214 TI - [Pregnancy termination in the second trimester with vaginally administered dinoprostone followed by intravenous sulprostone, for the indication 'foetal congenital defects'; results of a retrospective study]. AB - OBJECTIVE: To obtain insight into treatment duration and complications of the currently accepted method for pregnancy termination in the second trimester using a combination of prostaglandine-E2 medications. DESIGN: Retrospective study of medical records. METHOD: Data were collected for all second trimester pregnancy terminations performed on foetal indication in the Erasmus Medical Center in Rotterdam, The Netherlands, in the years 1998-2001. The treatment consisted of vaginal administration of dinoprostone, followed by intravenous administration of sulprostone. RESULTS: A total of 134 pregnant women were involved in the analysis. The median age was 33 years and the median pregnancy duration 19 weeks. The median duration of hospital stay was 3 days (range: 2-11 days). The median duration of treatment for the total study population was 19 hours (4-172). For nulliparae this was 23 hours, and for multiparae 17 hours (p < 0.05). Of the total study population, 91 women (68%) delivered within 24 hours. The percentage of multiparae that delivered within 24 hours was higher than the percentage of nulliparae that delivered in this time (76% compared to 53%: p < 0.05). Operative removal of a--partially--retained placenta was conducted in 51 women (38%). This percentage was higher among women with treatment duration longer than 24 hours than among women who delivered within 24 hours (51% versus 32%; p < 0.05). The treatment duration and the complication percentage were less favourable than those described in a number of publications relating to the combination mifepristone and misoprostol. PMID- 15279215 TI - [A child with traumatic hemobilia]. AB - A 4-year-old boy was hit by a car travelling at 40 km/h and was admitted 3.5 h later to the department of paediatric surgery. Because he was haemodynamically unstable and needed blood transfusion, the patient underwent an emergency operation. The liver was ruptured in the right lobe. A large haematoma was found in the serosa of the duodenum, along with a Meckel's diverticle, which was left in place. The liver rupture was covered and sealed. One month after the accident the patient was re-admitted, because of abdominal pain and gastrointestinal bleeding. The cause was thought to be the Meckel's diverticle, which was removed later. Two months after the trauma the patient was re-admitted with abdominal pain, again with haematemesis and melaena. The diagnosis of hemobilia was obtained with MRI and angiography, which revealed a ruptured pseudoaneurysm of the ramus dexter of the proper hepatic artery. The patient was successfully treated with embolization. The diagnostic delay was two months, which illustrates the importance of considering the possibility of the diagnosis hemobilia in case of gastrointestinal haemorrhage combined with biliary symptoms. PMID- 15279216 TI - [Uterine hyperstimulation following cervix ripening with dinoprostone in a vaginal insert system]. AB - Three women, aged 28, 29 and 31 years, primigravidae, with an unripe cervix and an indication for induction of labour, were administered dinoprostone in a controlled vaginal insert system (VIS). A few hours after the insertion of the VIS strong, prolonged contractions occurred with bradycardia in the foetus, resulting in an emergency caesarean section. The children and the mothers recovered well. A potential adverse drug reaction of prostaglandins is uterine hyperstimulation. The sustained-release intravaginal dinoprostone was expected to be safer than the intravaginal or intracervical application of a prostaglandin gel. But data from the literature are conflicting. The risk of uterine hyperstimulation by prostaglandins including the sustained-release dinoprostone system necessitates a re-evaluation of the indications for induction of labour and the procedures of cervical priming. Up-to-date guidelines are an essential tool for the safe use of prostaglandins in daily obstetric practice. PMID- 15279217 TI - [The hidden treasure-chests of the regional cancer registries]. PMID- 15279218 TI - ['The rumbling of shaking brains'; the treatment of traumatic skull and brain injury in the Netherlands in the 17th century: 7 case reports from Observationes medicae by Nicolaes Tulp]. PMID- 15279219 TI - [Cutaneous nocardiosis as an opportunistic infection]. PMID- 15279220 TI - [Important prognostic significance of a sentinel-node biopsy in patients with malignant melanoma]. PMID- 15279221 TI - [Diagnostic image (184). Two women with painful bands after axillary lymph node removal]. PMID- 15279222 TI - [Amitraz intoxications in the horse: cases and backgrounds]. AB - Since the only registrated anti-ectoparasiticum for horses (foxim) is no longer available in The Netherlands, some cases of amitraz intoxication of the horse have occurred. In this article the literature concerning amitraz intoxications and experiments will be discussed and some cases will be described. PMID- 15279223 TI - [Is porcine reproductive and respiratory syndrome (PRRS) of Aujeszky from the 21st century? Round table conference about PRRS prevention and control]. PMID- 15279224 TI - [Homeopathic information]. PMID- 15279225 TI - [Predecessors: veterinarians from earlier times (54). Pierre-Victor Galtier (1846 1908)]. PMID- 15279226 TI - [Reaction to 'Alienation from the society']. PMID- 15279228 TI - Significant perfection. PMID- 15279227 TI - [Biomechanical aspects of the claw-floor interaction in dairy cattle: implications for locomotion and claw disorders]. AB - The prevalence of claw disorders is still high among cows housed on concrete floors. Concrete floors affect the locomotion of cattle, their natural behavior. Although many factors affect the development of claw disorders and locomotor problems, biomechanical aspects have hardly been analysed. In this thesis, mechanical (over)loading of the claw and its significance for claw disorders and lameness are discussed. The mechanical characteristics a floor needs to provide in order to enable unrestrained locomotory behavior. This biomechanical approach, which is a relatively new approach in cattle locomotion, has provided new insights. Despite preventive trimming, the weakest parts of the claw capsule are loaded relatively the most. Concrete floors provide too little friction to enable unrestricted cattle locomotion. PMID- 15279229 TI - New patient communication. PMID- 15279230 TI - Implementing a tobacco control plan in the dental office. PMID- 15279231 TI - Proximal margin modifications for all-ceramic veneers. AB - Conventional ceramic veneers circumvent contact areas and extend palatally in the gingival third of the tooth only. Veneers are no longer considered reversible therapy. With the objective of longevity in mind, proximal extensions beyond contact areas maximize the functional and aesthetic potential of veneers and maintain conservative tooth preparations. The clinical and laboratory advantages of proximal veneer extensions for anterior teeth significantly outweigh the disadvantage of increased tooth structure removal. This article presents a modified preparation technique for the proximal extension of ceramic veneer preparations in the anterior region. PMID- 15279232 TI - Ensuring success with newer materials and techniques. PMID- 15279233 TI - Aesthetic rehabilitation of severely discolored anterior dentition: restorative considerations using all-ceramic veneers. PMID- 15279234 TI - Lasers in modern caries management. PMID- 15279236 TI - Immediate Occlusal Loading (IOL) of dental implants: predictable results through DIEM guidelines. AB - For years, dental implants have been loaded immediately upon implant placement with varying degrees of success. As clinicians' understanding of the biological and mechanical factors involved in immediate occlusal loading (IOL) has evolved, the success of these procedures has increased--particularly as a treatment option for the restoration of the edentulous mandible or the mandible that will be rendered edentulous during treatment. Due to increasing interest in this treatment alternative, the authors have provided a clear definition of the terminology associated with IOL and have demonstrated the DIEM Guidelines used to increase the success and predictability of such treatment. This presentation also introduces new implant components that simplify the clinical application of the immediate loading concept, enhancing its benefits and acceptance among dental patients and practices alike. PMID- 15279237 TI - Advancements in dentistry: digital radiography and its application. PMID- 15279238 TI - Finishing and polishing techniques: direct composite resin restorations. AB - Contemporary clinicians are responsible for delivering realistic restorations that closely resemble the existing natural tooth structures. In order to achieve such aesthetic results, the use of exemplary materials is required. Recent advancements in composite resin systems have improved the practitioner's ability to deliver optimal results using chairside techniques. The incorporation of a precise finishing and polishing protocol further allows the clinician to seal the restoration and maintain natural surface luster and contour. This article presents the necessary finishing and polishing protocol to ensure long-term aesthetic results. PMID- 15279239 TI - Aesthetic tooth replacement and restoration of the anterior maxilla with pressed ceramic restorations. PMID- 15279240 TI - Occlusal parameters for ceramic restorations: biological and functional considerations. PMID- 15279241 TI - Minimally invasive aesthetic treatment for discolored and fractured teeth in adolescents: a case report. AB - This article describes a minimally invasive treatment of a young patient who presented with generalized enamel white spots and hypocalcification in combination with a severely fractured central incisor. Enamel microabrasion technique was used to remove discolored and/or pitted enamel. A direct composite restoration was then placed according to the anatomical layering technique. These procedures are described step-by-step herein. The advantages of microabrasion and composite restoration over conventional prosthetic procedures are also discussed. PMID- 15279242 TI - Contemporary implant concepts in aesthetic dentistry--part 3: adjacent immediate implants in the aesthetic zone. AB - This series of articles has discussed contemporary concepts in the treatment planning of dental implants in edentulous ridges and immediate single-tooth implants in the aesthetic zone. This presentation broadens the discussion to include the wider application of immediate implants, highlighting the surgical and restorative considerations affecting their immediate placement and provisionalization. It highlights the application of immediate implants in both single- and multiple-unit applications and provides detailed demonstrations of the involved implant and restorative components required by such procedures. PMID- 15279243 TI - Shared genetic basis of resistance to Bt toxin Cry1ac in independent strains of pink bollworm. AB - Classical and molecular genetic analyses show that two independently derived resistant strains of pink bollworm, Pectinophora gossypiella (Saunders), share a genetic locus at which three mutant alleles confer resistance to Bacillus thuringiensis (Bt) toxin Cry1Ac. One laboratory-selected resistant strain (AZP-R) was derived from individuals collected in 1997 from 10 Arizona cotton fields, whereas the other (APHIS-98R) was derived from a long-term susceptible laboratory strain. Both strains were previously reported to show traits of "mode 1" resistance, the most common type of lepidopteran resistance to Cry1A toxins. Inheritance of resistance to a diagnostic concentration of Cry1Ac (10 microg per gram of diet) was recessive in both strains. In interstrain complementation tests for allelism, F1 progeny from crosses between the two strains were resistant to the diagnostic concentration of Cry1Ac. These results indicate that a major resistance locus is shared by the two strains. Analysis of DNA from the pink bollworm cadherin gene (BtR) using allele-specific polymerase chain reaction (PCR) tests showed that the previously identified resistance alleles (r1, r2, and r3) occurred in both strains, but their frequencies differed between strains. In conjunction with previous findings, the results reported here suggest that PCR based detection of the three known cadherin resistance alleles might be useful for monitoring resistance to Cry1Ac-producing Bt cotton in field populations of pink bollworm. PMID- 15279244 TI - Influence of brood, vent screening, and time of year on honey bee (Hymenoptera: Apidae) pollination and fruit quality of greenhouse tomatoes. AB - Greenhouse tomatoes, Lycopersicon esculentum Miller (Solanaceae), are autogamous, but facilitated pollination results in increased fruit size and set. Previous research examining honey bee pollination in greenhouse tomato crops established that fruit quality resulting from honey bee visitation is often comparable to bumble bees (Bombus spp.) and significantly better than in flowers that receive no facilitated pollination. However, management alternatives have not been studied to improve tomato fruit quality when honey bees are the only pollination option available for the high-value greenhouse industry. We investigated whether the quantity of brood (eggs, larvae, and pupae) in a honey bee colony in the winter and screening on greenhouse vents in the summer would encourage honey bee foraging on tomato flowers. We also established the influence of time of year on the potential for honey bees to be effective pollinating agents. We constructed small honey bee colonies full of naive forager bees with either two frames of brood ("brood colonies") or two empty frames ("no-brood") and compared total fruit set and the number of tomato seeds resulting from fruit potentially visited by honey bees in each of these treatments to bagged flowers that received no facilitated pollination. There was no significant difference in the quality of fruit resulting from honey bees from "brood" and "no-brood" colonies. However, these fruits produced significantly more seeds than bagged flowers restricted from facilitated pollination. Honey bees from brood and no-brood colonies also resulted in 98% fruit set compared with 80% fruit set in bagged flowers that received no facilitated pollination. During the summer, the number of seeds per fruit did not differ significantly between unbagged flowers potentially visited by honey bees in screened greenhouses and unscreened greenhouses and bagged flowers that received no facilitated pollination. However, time of year did have a significant influence on the quality of fruit produced by honey bees compared with flowers that received no facilitated pollination, because no difference in seed number was observed between the treatments after mid-April. The results from this study demonstrate that the management of brood levels and vent screening cannot be used to improve the quality of fruit resulting from honey bee pollination and that honey bees can be a feasible greenhouse pollination alternative only during the winter. PMID- 15279245 TI - Nectar robbery by bees Xylocopa virginica and Apis mellifera contributes to the pollination of rabbiteye blueberry. AB - Honey bees, Apis mellifera L., probe for nectar from robbery slits previously made by male carpenter bees, Xylocopa virginica (L.), at the flowers of rabbiteye blueberry, Vaccinium ashei Reade. This relationship between primary nectar robbers (carpenter bees) and secondary nectar thieves (honey bees) is poorly understood but seemingly unfavorable for V. ashei pollination. We designed two studies to measure the impact of nectar robbers on V. ashei pollination. First, counting the amount of pollen on stigmas (stigmatic pollen loading) showed that nectar robbers delivered fewer blueberry tetrads per stigma after single floral visits than did our benchmark pollinator, the southeastern blueberry bee, Habropoda laboriosa (F.), a recognized effective pollinator of blueberries. Increasing numbers of floral visits by carpenter bee and honey bee robbers yielded larger stigmatic loads. As few as three robbery visits were equivalent to one legitimate visit by a pollen-collecting H. laboriosa female. More than three robbery visits per flower slightly depressed stigmatic pollen loads. In our second study, a survey of 10 commercial blueberry farms demonstrated that corolla slitting by carpenter bees (i.e., robbery) has no appreciable affect on overall V. ashei fruit set. Our observations demonstrate male carpenter bees are benign or even potentially beneficial floral visitors of V ashei. Their robbery of blueberry flowers in the southeast may attract more honey bee pollinators to the crop. PMID- 15279246 TI - Altered physiology in worker honey bees (Hymenoptera: Apidae) infested with the mite Varroa destructor (Acari: Varroidae): a factor in colony loss during overwintering? AB - The ectoparasitic mite Varroa destructor (Anderson & Trueman) is the most destructive pest of the honey bee, Apis mellifera L., in Europe and the United States. In temperate zones, the main losses of colonies from the mites occur during colony overwintering. To obtain a deeper knowledge of this phenomenon, we studied the mites' impact on the vitellogenin titer, the total protein stores in the hemolymph, the hemocyte characteristics, and the ecdysteroid titer of adult honey bees. These physiological characteristics are indicators of long-time survival and endocrine function, and we show that they change if bees have been infested by mites during the pupal stage. Compared with noninfested workers, adult bees infested as pupae do not fully develop physiological features typical of long-lived wintering bees. Management procedures designed to kill V. destructor in late autumn may thus fail to prevent losses of colonies because many of the adult bees are no longer able to survive until spring. Beekeepers in temperate climates should therefore combine late autumn management strategies with treatment protocols that keep the mite population at low levels before and during the period when the winter bees emerge. PMID- 15279247 TI - Brood pheromone regulates foraging activity of honey bees (Hymenoptera: Apidae). AB - Brood pheromone modulated the foraging behavior of commercial honey bee, Apis mellifera L., colonies pollinating a 10-ha market garden of cucumber, Cucurbita pepo L., and zucchini, Cucumis saticus L., in Texas in late autumn. Six colonies were randomly selected to receive 2000 larval equivalents of brood pheromone and six received a blank control. The ratio of pollen to nonpollen foragers entering colonies was significantly greater in pheromone-treated colonies 1 h after treatment. Pheromone-treated foragers returned with pollen load weights that were significantly heavier than controls. Pollen returned by pheromone-treated foragers was 43% more likely to originate from the target crop. Number of pollen grains washed from the bodies of nonpollen foragers from pheromone-treated colonies was significantly greater than controls and the pollen was 54% more likely to originate from the target crop. Increasing the foraging stimulus environment with brood pheromone increased colony-level foraging and individual forager efforts. Brood pheromone is a promising technology for increasing the pollination activity and efficiency of commercial honey bee colonies. PMID- 15279248 TI - Bacterial probiotics induce an immune response in the honey bee (Hymenoptera: Apidae). AB - To explore immune system activation in the honey bee, Apis mellifera L., larvae of four ages were exposed through feeding to spores of a natural pathogen, Paenibacillus larvae larvae, to cells of a diverse set of related nonpathogenic bacteria, and to bacterial coat components. These larvae were then assayed for RNA levels of genes encoding two antibacterial peptides, abaecin and defensin. Larvae exposed to either P. l. larvae or a mix of nonpathogenic bacteria showed high RNA levels for the abaecin gene relative to controls. First instars responded significantly to the presence of the nonpathogenic mix within 12 h after exposure, a time when they remain highly susceptible to bacterial invasion. This response was sustained for two successive instars, eventually becoming 21 fold higher in larvae exposed to probiotic spores versus control larvae. The mixture of nonpathogenic bacteria is therefore presented as a potential surrogate for assaying the immune responses of different honey bee lineages. It also is proposed that nonpathogenic bacteria can be used as a probiotic to enhance honey bee immunity, helping bee larvae, and other life stages, survive attacks from pathogens in the field. PMID- 15279249 TI - Evaluation of methods for extracting Xylella fastidiosa DNA from the glassy winged sharpshooter. AB - The recent spread of the plant pathogenic bacterium Xylclla fastidiosa Wells et al. by an invasive vector species, Homalodisca coagulata Say, in southern California has resulted in new epidemics of Pierce's disease of grapevine. Our goal is to develop an efficient method to detect low titers of X. fastidiosa in H. coagulata that is amenable to large sample sizes for epidemiological studies. Detection of the plant pathogenic bacterium X. fastidiosa in its insect vector is complicated by low titers of bacteria, difficulty in releasing it from the insect mouthparts and foregut, and the presence of substances in the insect that inhibit polymerase chain reaction (PCr). To select the optimal protocol for DNA extraction to be used with PCR, we compared three standard methods and 11 commercially available kits for relative efficiency of X. fastidiosa DNA extraction in the presence of insect tissue. All of the protocols tested were proficient at extracting DNA from pure bacterial culture (1 x 10(5) cells), and all but one protocol successfully extracted sufficient bacterial DNA in the presence of insect tissue. Three DNA extraction techniques, immunomagnetic separation, the DNeasy Tissue kit (Qiagen, Hercules, CA), and Genomic DNA Purification kit (Fermentus, Hanover, MD), were compared more closely using a dilution series of X. fastidiosa (5000-0 cells) with and without insect tissue present. The DNeasy Tissue kit was the best kit tested, allowing detection of 5 x 10(3) X. fastidiosa cells with an insect head background. PMID- 15279250 TI - Viability of Claviceps africana spores ingested by adult corn earworm moths, Helicoverpa zea (Boddie) (Lepidoptera: Noctuidae). AB - A study was conducted in College Station, TX, to determine the viability of Claviceps africana spores in the digestive tract of adult corn earworm moths, Helicoverpa zea (Boddie). Both sexes were exposed to ergot-infected sorghum panicles for 30 min, and spores were recovered from excreta of the moths at 24-, 48-, and 72-h intervals after feeding. Recovered spores were quantified, and viability was determined by the germination rate of macroconidia. Nearly a 100 fold greater concentration of spores was recovered from female excreta at the three time intervals compared with male excreta. Concentration of spores in female and male excreta was greatest at 24 h, with a significant reduction at the later time intervals. Spore germination rates for both sexes were greater at 24 h, with survival being significantly reduced at the 72-h interval. Spores in female excreta survived longer than those from male excreta. Spore survival over time was significantly reduced in male excreta. Spore concentration and survival were greater from female excreta, which is key, because egg-laying activities on sorghum panicles intensify during flowering, and this source of ergot spores could contribute to the spread of the disease. This study demonstrates that corn earworm moths can internally carry viable ergot spores for several days and can act as primary dispersal agents for the fungus. This is important because contaminated moths migrating from areas in Mexico and southern Texas where ergot is endemic could transmit and spread the disease to other sorghum growing regions of the United States. PMID- 15279251 TI - Effect of environmental conditions and leafhopper gender on Maize chlorotic dwarf virus transmission by Graminella nigrifrons (Homoptera: Cicadellidae). AB - To determine the most economical and efficient means to maintain cultures of Maize chlorotic dwarf virus (MCDV) and to screen for host plant resistance to MCDV, we evaluated the effects of temperature, light intensity, daylength, atmospheric pressure, and leafhopper gender on the frequency of transmission of MCDV by Grarminella nigrifrons Forbes (Homoptera: Cicadellidae). Female leafhoppers transmitted at higher frequencies than males under most conditions. In temperature studies, transmission rates for both male and female leafhoppers progressively increased as temperatures rose from 20 to 30 degrees C. At high light intensities, both males and females transmitted at greater frequencies than they did at low. Similarly, longer day lengths were correlated with higher transmission rates for both sexes. No significant differences in transmission rates were observed in response to differences in atmospheric pressure. The results also showed that transmission rates under most conditions are high enough to overcome potential ambiguities caused by inoculated susceptible plants that do not become infected (disease escapes) when screening for resistance. PMID- 15279252 TI - Two new bacterial pathogens of Colorado potato beetle Coleoptera: Chrysomelidae). AB - Other than Bacillus thuringiensis Berliner, few bacteria are lethal to the Colorado potato beetle (Leptinotarsa decemlineata [Say]), a major pest of potatoes and eggplant. Expanded use of biologicals for the control of Colorado potato beetle will improve resistance management, reduce pesticide use, and produce novel compounds for potential use in transgenic plants. Using freeze dried, rehydrated artificial diet in pellet form to screen bacteria lethal to other insects, we determined that strains of Photorhabdus luminescens killed Colorado potato beetle larvae. The LC50 for second instar larvae of strain HM5-1 was 6.4 +/- 1.87 x 10(7) cells per diet pellet. In an attempt to find additional naturally occurring P. luminescens strains toxic to Colorado potato beetle larvae, we recovered, from soil, bacteria that produced a purple pigment. This bacterial strain, identified as Chromobacterium sp. by 16S ribosomal DNA sequencing, was also toxic to Colorado potato beetle larvae within 3 d. The LC50 for second instar larvae for these bacteria was 2.0 +/- 0.79 x 10(8) cells per diet pellet, while the LC50 was approximately 1 log lower for third instar larvae. P. luminescens appeared to kill by means of a protein toxin that may be similar to the described lepidopteran protein toxins. Based on the heat and acid stability, the toxin or toxins that Chromobacterium sp. produces, while not fully characterized, do not appear to be typical proteins. In both bacteria, the toxins are made after exponential growth ceases. PMID- 15279253 TI - Quality assessment of selected commercially available whitefly and aphid biological control agents in the United States. AB - This study assessed the quality of three commercially available natural enemies used for pest management in greenhouses: the whitefly parasitoid Encarsia formosa Gahan (Hymenoptera: Aphelinidae), the aphid parasitoid Aphidius colemani Viereck (Hymenoptera: Braconidae), and the aphid predatory midge Aphidoletes aphidimlyza (Rondani) (Diptera: Cecidomyiidae). Shipment packaging was consistent for all natural enemies. However, there was high variability in delivery punctuality, product cost, and product information provided by each of the six selected companies. Product quantity, percentage of emergence upon arrival, percentage of total emergence, percentage of females, and percentage of flying insects were assessed using International Organization for Biological Control (IOBC) recommended procedures. The parameters with greatest variability between companies were percentage of emergence upon arrival (0.9-10.5%) and percentage of flying insects (35.4-85.0%) for E. formnosa; product quantity (623.3-833.8 aphid mummies), percentage of emergence upon arrival (6.1-41.2%) and percentage of females (51.1-54.8%) for A. colemani; and percentage of emergence upon arrival (0.0-7.7%) and percentage of females (54.6-76.2%) for A. aphlidimyza. Results are discussed in terms of the value to consumers and compared with IOBC standards. PMID- 15279254 TI - Impact of peach extrafloral nectar on key biological characteristics of Trichogramma minutum (Hymenoptera: Trichogrammatidae). AB - This study investigated the longevity, fecundity, and host feeding of a wild and a commercially reared strain of Trichogramma minutum Riley (Hymenoptera: Trichogrammatidae) when presented with peach extrafloral nectar. Both the wild and commercial T. minutum strains lived longer and parasitized more Grapholita molesta (Busck) eggs when provided peach extrafloral nectar and water than when provided only water. Nectar-fed wild and commercially reared females lived 11.6 and 9.9 d and produced 105.2 and 61.0 offspring, respectively. When provided only water, wild females lived 3.3 d and produced 52.8 offspring, whereas commercially reared females lived 2.0 d and produced 24.4 offspring. Nectar feeding significantly increased the number of G. molesta eggs destroyed by host feeding by both wild and commercial strains. PMID- 15279255 TI - Effects of greenhouse pesticides on the soil-dwelling predatory mite Stratiolaelaps scimitus (Acari: Mesostigmata: Laelapidae) under laboratory conditions. AB - Knowledge of the effects of pesticides on biological control agents is required for the successful implementation of integrated pest management (IPM) programs in greenhouse production systems. Laboratory assays were conducted to assess the effects of an acaricide (dicofol), two insecticides (chlorpyrifos and pyriproxyfen), and two fungicides (fosetyl-Al and mefenoxam) on Stratiolaelaps scimitus (Womersley), a soil-dwelling predatory mite widely marketed in North America under the name Hypoaspis miles (Berlese) as a biological control agent of dark-winged fungus gnats (Bradysia spp.). Eggs, larvae, protonymphs, deutonymphs, and adult male and female mites were first assayed using dicofol, an acaricide used in the experiments as a positive control, applied to filter paper in an enclosed arena. Protonymphs were assayed for lethal and sublethal effects against the remaining pesticides at maximum label-recommended rates applied to filter paper, by using dicofol as a positive control and water as a negative control. The larva and protonymph were the life stages most susceptible to dicofol, with estimated 24-h LC50 values of 9 and 26 mg m(-2), respectively. Chlorpyrifos was highly toxic to the protonymphs of S. scimitus, causing >95% mortality after 24-h exposure and 100% mortality after 48 h. In contrast, the insect growth regulator (IGR) pyriproxyfen was much less toxic to protonymphs of S. scimitus; pyriproxyfen caused no significant mortality, compared with <5% mortality in the water control. Mortality caused by the fungicides was also relatively low; 72-h exposure to fosetyl-Al and mefenoxam resulted in 17.4 and 27.5% mortality, respectively. The IGR and fungicides increased the duration of the protonymphal stage by 1.2-1.8-fold, but they had no effect on the duration of subsequent life stages, nor on the duration of preoviposition, oviposition, and postoviposition periods of adult females. Total numbers and viability of eggs laid by mites exposed to the IGR and fungicides did not differ from the negative control, although the average rate of egg production during the oviposition of mites exposed to fosetyl-Al was increased. Pyriproxyfen, fosetyl-Al, and mefenoxam are likely to be compatible with S. scimitus under field conditions, because these pesticides caused little mortality of protonymphs, and they did not negatively affect the development and reproduction of S. scimitus under extreme laboratory conditions. In contrast, the use of chlorpyrifos in conjunction with S. scimitus is not recommended unless more comprehensive testing under semifield or field conditions demonstrates compatibility. PMID- 15279256 TI - Comparison of susceptibility of pest Euschistus servus and predator Podisus maculiventris (Heteroptera: Pentatomidae) to selected insecticides. AB - Susceptibility of the brown stink bug, Euschistus serous (Say), and the spined soldier bug, Podisus maculiventris (Say), to acetamiprid, cyfluthrin, dicrotophos, indoxacarb, oxamyl, and thiamethoxam, was compared in residual and oral toxicity tests. Generally, susceptibility of P. maculiventris to insecticides was significantly greater than or not significantly different from that of E. servus. Cyfluthrin and oxamyl were more toxic to the predator than to E. servus in residual and feeding tests, respectively. Dicrotophos is the only compound that exhibited both good residual and oral activity against E. servus, but even this toxicant was more toxic to the predator than to the pest in oral toxicity tests. Feeding on indoxacarb-treated food caused high mortality for both nymphs and adults of P. maculiventris. In contrast, E. servus was unaffected by feeding on food treated with this compound. Insecticide selectivity to P. maculiventris was detected only with acetamiprid for adults in residual toxicity tests and for nymphs in oral toxicity tests. Because insecticide selectivity to P. maculiventris was limited, it is extremely important to conserve P. maculiventris in cotton fields by applying these insecticides for control of brown stink bugs only when the pest reaches economic threshold. PMID- 15279257 TI - Life history and cost analysis for continuous rearing of Perillus bioculatus (Heteroptera: Pentatomidae) on a zoophytogenous artificial diet. AB - The impact of a zoophytogenous, insect-free artificial diet on the developmental rate, life history parameters, and fertility was examined over 11 consecutive generations for domesticated Perillus biocullatus (F.) (Heteroptera: Pentatomidae). This study showed that when fed an insect-free artificial diet during both the nymphal and adult stages, developmental times were prolonged, and the net reproductive rates (R0) and the intrinsic rates of increase (r(m)) were significantly lower than when fed larval Trichoplusia ni at both nymphal and adult stages. Moreover, the cost to rear P. bioculatus on the artificial diet approached 1.1 times the cost of rearing P. bioculatus on natural prey. These results demonstrate the effectiveness and potential cost-savings of the zoophytogenous artificial diet for rearing a beneficial pentatomid. PMID- 15279258 TI - Feeding and egg distribution studies of Heringia calcarata (Diptera: Syrphidae), a specialized predator of woolly apple aphid (Homoptera: Eriosomatidae) in Virginia apple orchards. AB - Predation by the aphidophagous syrphid fly Heringia calcarata (Loew) on woolly apple aphid, Eriosoma lanigerum (Hausmann), was studied in the laboratory and in Virginia apple orchards. Feeding studies compared the prey suitability of three temporally sympatric aphid pests of apple: spirea aphid, Aphis spiraecola Patch; rosy apple aphid, Dysaphis plantaginea (Passerini); and woolly apple aphid. Significantly more H. calcarata larvae survived and completed development on a pure diet of woolly apple aphid than on rosy apple aphid, and none survived on spirea aphid. Final larval weights were significantly greater, and the larval developmental period was significantly shorter on woolly apple aphid than on rosy apple aphid, but neither the duration of pupal development nor adult weight differed between diets. H. calcarata larvae consumed an average of 105 woolly apple aphids during their development. Naive, neonate larvae given access to all possible pair combinations of woolly apple aphid, rosy apple aphid, and spirea aphid consumed significantly more woolly apple aphids in all pairings that included woolly apple aphid. When given a choice of rosy apple aphid and spirea aphid, significantly more rosy apple aphids were consumed. Weekly counts of syrphid eggs found in woolly apple aphid, rosy apple aphid, and spirea aphid colonies collected from apple trees showed that two generalist hover fly predators, Eupeodes americanus (Wiedemann) and Syrphus rectus Osten Sacken, were present in colonies of all three aphid species and that E. americanus was the most abundant syrphid predator in A. spiraecola and D. plantaginea colonies. H. calcarata eggs were found only in woolly apple aphid colonies and were more abundant there than E. americanus and S. rectus. These data suggest that H. calcarata is a specialized predator of woolly apple aphid in the apple ecosystem in Virginia. PMID- 15279259 TI - Effects of low temperature on egg and larval stages of the lesser appleworm (Lepidoptera: Tortricidae). AB - Lesser appleworm, Grapholita prunivora (Walsh), eggs were subjected to cold storage conditions at 2.0 degrees C +/- 0.2 degrees C for 0-90 d. The most tolerant embryonic stage was the blackhead stage (96-120-h-old eggs) with an LT90 of 25 d. The four instars of lesser appleworm were subjected to cold storage conditions at 2.0 degrees C +/- 0.2 degrees C for 0-280 d. The fourth instar was the most tolerant to cold storage, with an LT90 of 71.5 d. Exposure to low temperatures such as those commonly used for fruit storage shows promise as an alternative to fumigation for lesser appleworm eggs and larvae on apples and pears after harvest. PMID- 15279260 TI - Ionizing irradiation quarantine treatment against oriental fruit moth (Lepidoptera: Tortricidae) in ambient and hypoxic atmospheres. AB - Oriental fruit moth, Grapholita molesta (Busck), is a pest of many rosaceous temperate fruits, including pomes, Malus spp., and stone fruits, Prunus spp., in much of the world. However, some areas are free of the pest, and shipments of fruit hosts from infested to noninfested areas may be regulated. Current quarantine treatments for oriental fruit moth include methyl bromide fumigation and cold storage for several weeks. Methyl bromide use is being restricted because it is a stratospheric ozone-depleting substance, and alternatives are sought. Cold is not tolerated by many hosts of oriental fruit moth. The objective of this research was to develop irradiation quarantine treatments against the pest under ambient and hypoxic storage conditions because some hosts of oriental fruit moth are stored in hypoxic atmospheres, and hypoxia is known to lessen the effects of irradiation. In ambient atmospheres, no adults emerged from 58,779 fifth instars (the most radiotolerant stage present in fruit) irradiated with a target dose of 200 Gy (195-232 Gy measured). In atmospheres flushed with nitrogen, 5.3% of adults emerged from 44,050 fifth instars irradiated with a target dose of 200 Gy (194-230 Gy measured), but they died at a faster rate than control adults and without laying eggs. A dose of 232 Gy (the maximum recorded when 200 Gy was targeted) is recommended to disinfest any fruit of oriental fruit moth under ambient and hypoxic atmospheres. PMID- 15279261 TI - Female European corn borer (Lepidoptera: Crambidae) ovarian developmental stages: their association with oviposition and use in a classification system. AB - Reproductive development of female European corn borer, Ostrinia nubilalis (Hubner), was investigated and a classification system proposed. Females collected in a blacklight trap during 1982 and 1983 were dissected and their reproductive system examined. Female reproductive systems were divided into six stages based on ovum development within the ovarioles, ovum depletion, ovariole appearance, and fat body color and shape. The female reproductive systems were also staged on the basis of spermatophore appearance. The time necessary to classify a female is also reported. Based on the classification system, the relationship between female age and stage of ovarian development was quantified under three temperature regimes. Females were found to experience a 3- to 5-d preoviposition period before initiation of egg deposition under optimal temperature conditions. This delay between adult emergence and initiation of egg laying corresponded with more advanced ovarian developmental stages collected in blacklight traps and indicates that actively ovipositing females are primarily being collected in blacklight traps. PMID- 15279262 TI - Oil-soluble dyes incorporated in meridic diet of Diatraea grandiosella (Lepidoptera: Crambidae) as markers for adult dispersal studies. AB - Mark-release-recapture experiments to study insect dispersal require the release of marked insects that can be easily identified among feral conspecifics. Oil soluble dyes have been used successfully to mark various insect species. Two oil soluble dyes, Sudan Red 7B (C.I. 26050) and Sudan Blue 670 (C.I. 61554), were added to diet of the southwestern corn borer, Diatraea grandiosella Dyar, and evaluated against an untreated control diet. Survival, diet consumption, larval and pupal weight, development time, fecundity, longevity, and dry weight of the adults were measured. Adults reared on the three diets were also tested for mating success. Some minor effects were observed for southwestern corn borers reared on the marked diets. Eggs, larvae, pupae, and adults were all reliably marked and readily identifiable. Adults retained color for their entire life span. Adults from each diet mated successfully with adults from the other diets. F1 progeny from the different mating combinations survived to the second instar but tended to lose the marker after 3-4 d on untreated diet. Both Sudan Red 7B and Sudan Blue 670 can be used to mark southwestern corn borer adults and thus should be useful for mark-release-recapture dispersal studies. The dyes will also be useful for short-term studies with marked larvae and oviposition behavior. PMID- 15279263 TI - Aromatherapy in the Mediterranean fruit fly (Diptera: Tephritidae): sterile males exposed to ginger root oil in prerelease storage boxes display increased mating competitiveness in field-cage trials. AB - Previous research showed that exposure to ginger root, Zingiber officinale Roscoe, oil increased the mating success of mass-reared, sterile males of the Mediterranean fruit fly, Ceratitis capitata (Wiedemann). This work, however, involved the exposure of small groups of males (n = 25) in small containers (volume 400 ml). Several sterile male release programs use plastic adult rearing containers (so-called PARC boxes; hereafter termed storage boxes; 0.48 by 0.60 by 0.33 m) to hold mature pupae and newly emerged adults before release (approximately = 36,000 flies per box). The objective of the current study was to determine whether the application of ginger root oil to individual storage boxes increases the mating competitiveness of sterile C. capitata males. Irradiated pupae were placed in storage boxes 2 d before adult emergence, and in the initial experiment (adult exposure) ginger root oil was applied 5 d later (i.e., 3 d after peak adult emergence) for 24 h at doses of 0.0625, 0.25, 0.5, 1.0, and 2.0 ml. In a second experiment (pupal-adult exposure), ginger root oil was applied to storage boxes immediately after pupal placement and left for 6 d (i.e., 4 d after peak adult emergence) at doses of 0.25 and 1.0 ml. Using field cages, we conducted mating trials in which ginger root oil-exposed (treated) or nonexposed (control) sterile males competed against wild-like males for copulations with wild-like females. After adult exposure, treated males had significantly higher mating success than control males for all doses of ginger root oil, except 2.0 ml. After pupal-adult exposure, treated males had a significantly higher mating success than control males for the 1.0-ml but not the 0.25-ml dose of ginger root oil. The results suggest that ginger root oil can be used in conjunction with prerelease, storage boxes to increase the effectiveness of sterile insect release programs. PMID- 15279264 TI - Demography of soybean aphid (Homoptera: Aphididae) at summer temperatures. AB - Soybean aphid, Aphis glycines Matsumura, is now widely established in soybean, Glycine max L., production areas of the northern United States and southern Canada and is becoming an important economic pest. Temperature effect on soybean aphid fecundity and survivorship is not well understood. We determined the optimal temperature for soybean aphid growth and reproduction on soybean under controlled conditions. We constructed life tables for soybean aphid at 20, 25, 30, and 35 degrees C with a photoperiod of 16:8 (L:D) h. Population growth rates were greatest at 25 degrees C. As temperature increased, net fecundity, gross fecundity, generation time, and life expectancy decreased. The prereproductive period did not differ between 20 and 30 degrees C; however, at 30 degrees C aphids required more degree-days (base 8.6 degrees C) to develop. Nymphs exposed to 35 degrees C did not complete development, and all individuals died within 11 d. Reproductive periods were significantly different at all temperatures, with aphids reproducing longer and producing more progeny at 20 and 25 degrees C than at 30 or 35 degrees C. Using a modification of the nonlinear Logan model, we estimated upper and optimal developmental thresholds to be 34.9 and 27.8 degrees C, respectively. At 25 degrees C, aphid populations doubled in 1.5 d; at 20 and 30 degrees C, populations doubled in 1.9 d. PMID- 15279265 TI - Active ingredients in cade oil that synergize attractiveness of alpha-ionol to male Bactrocera latifrons (Diptera: Tephritidae). AB - Cade oil, a commercially available essential oil produced by destructive distillation of juniper, Juniperus oxycedrus L., twigs, is known to synergize the attractancy of alpha-ionol to male Bactrocera latifrons (Hendel). Through chemical fractionation and outdoor olfactometer-based bioassays, seven compounds in cade oil were identified that potentially could provide some level of synergism. Tests with sterile laboratory flies showed that four of the seven compounds (eugenol, isoeugenol, 2-methoxy-4-ethylphenol, and 2-methoxy-4 propylphenol), together with a closely related compound not found in cade oil, 2 methoxy-4-methylphenol, are capable of synergizing the attractiveness of alpha ionol to male B. latifrons under field conditions. The similarity in structures of these five synergistic compounds shows that there is a response to a core 2 methoxyphenol structure, with fly response little affected by some variation in the composition of the side chain on the number 4 carbon. Because identified synergists were structurally similar, only one compound, eugenol, was selected for further field studies. In an 8-wk weathering test, using released sterile flies, traps baited with alpha-ionol + eugenol had catches comparable with catches at traps baited with alpha-ionol + cade oil, with catches generally increased with a higher eugenol loading. For both eugenol and cade oil, catches tended to be better when these synergists were deployed on separate wicks from the alpha-ionol. Eugenol and alpha-ionol, however, were unable to provide attraction comparable with that of cade oil and alpha-ionol in tests with wild fly populations. PMID- 15279266 TI - Role of egg density on establishment and plant-to-plant movement by western corn rootworm larvae (Coleoptera: Chrysomelidae). AB - The effect of egg density on establishment and dispersal of larvae of the western corn rootworm, Diabrotica virgifera virgifera LeConte, was evaluated in a 3-yr field study. Implications of these data for resistance management plans for Bt crops are discussed. Viable egg levels of 100, 200, 400, 800, and 1600 eggs per infested plant were evaluated in 2000, 2001, and 2002. A 3200 viable egg level was also tested in 2001 and 2002. All eggs were infested on one plant per subplot in a field that was planted to soybean, Glycine max (L.), in the previous year. For each subplot, the infested plant, three plants down the row, the closest plant in the adjacent row of the plot, and a control plant at least 1.5 m from any infested plant (six plants total) were sampled. In 2000, there were five sample dates between egg hatch and pupation, and in 2001 and 2002, there were six sample dates. On each sample date, four replications of each egg density were sampled for both larval recovery and plant damage. Initial establishment on a corn plant seemed to not be density-dependent because a similar percentage of larvae was recovered from all infestation rates. Plant damage and, secondarily, subsequent postestablishment larval movement were density-dependent. Very little damage and postestablishment movement occurred at lower infestation levels, but significant damage and movement occurred at higher infestation rates. Movement generally occurred at a similar time as significant plant damage and not at initial establishment, so timing of movement seemed to be motivated by available food resources rather than crowding. At the highest infestation level in 2001, significant movement three plants down the row and across the 0.76 m row was detected, perhaps impacting refuge strategies for transgenic corn. PMID- 15279267 TI - Distribution and population development of Nasonovia ribisnigri (Homoptera: Aphididae) in iceberg lettuce. AB - A field study was conducted to determine the distribution and development of aphid Nasonovia ribisnigri (Mosley) (Homoptera: Aphididae) populations in iceberg lettuce, Lactuca sativa L. 'Salinas'. Lettuce plants were transplanted and caged individually in the field and inoculated with apterous N. ribisnigri at 0, 1, 2, 3, and 4 wk after transplanting in spring and fall 2002. Plants were harvested 15 50 d after inoculations; numbers of alates and apterous N. ribisnigri were counted or estimated on each leaf for each plant. Inoculations during all 5 wk of plant development resulted in successful colonization of lettuce heads. Results indicated that head formation did not reduce the risk of colonization by N. ribisnigri to iceberg lettuce; plants were susceptible to colonization by N. ribisnigri throughout their development. For later inoculations, N. ribisnigri populations were relatively smaller, and aphids were found mostly within the heads. For earlier inoculations, N. ribisnigri populations were larger, and within-plant distributions shifted toward frame leaves. The shift of population distributions toward frame leaves correlated significantly with increases in N. ribisnigri population density. For most inoculations, more aphids were present on wrapper leaves than on other leaves. The proportion of alates did not vary significantly with population density. Population development of N ribisnigri also correlated significantly with heat unit accumulation. Yellow sticky cards were used to monitor alates in each cage. Catches of N. ribisnigri alates on yellow sticky cards were significantly correlated with total numbers of alates as well as with total population sizes on individual lettuce plants. PMID- 15279268 TI - Developing an ecotoxicological testing standard for predatory mites in Australia: acute and sublethal effects of fungicides on Euseius victoriensis and Galendromus occidentalis (Acarina: Phytoseiidae). AB - Laboratory bioassays for testing the effect of agrochemicals on Euseius victotiensis (Womersley) and Galendromus occidentalis (Nesbitt) on detached leaves of Glycine max (L.) (soybean) and Phaseolus vulgaris L. (French bean) were developed. The tests allowed standardized comparisons between mite species and leaf substrates, under "worst-case scenario" exposure, comparable with commercial pesticide application. Young juveniles, along with their initial food and the entire water supply, were sprayed to the point of runoff by using a Potter spray tower. The highest registered field rate concentration used on French bean was adjusted to deliver the same pesticide dose per higher runoff point spray volume on soybean. Cumulative mortality was assessed at 48 h, 4 d, and 7 d after spray application. Fecundity was assessed for 7 d from the onset of egg lay. Boscalid (Filan 500 WG), dithianon (Delan 700 WG), and kresoxim-methyl (Stroby 500 WG) caused no significant 7-d mortality or fecundity reduction to G. occidentalis or E. victoriensis compared with controls, and are classified as harmless to both species. Mancozeb (Mancozeb 750 WG) was highly toxic to both species, resulting in severe mortality and fecundity reduction and is considered incompatible with integrated pest management programs that use these species. Metiram (Polyram 700 WG) was highly toxic to E. victoriensis but only moderately toxic to G. occidentalis. Analyses of mortality proportions, including, and excluding unaccounted escapees, produced the same results. Test standardization on leaf substrates provides an alternative approach to standardization via residue on glass used by International Organisation for Biological and Integrated Control or Noxious Animals and Plants/West Palaearctic Regional Section regulatory testing in the European Union. PMID- 15279269 TI - Effect of buprofezin on survival of immature stages of Harmonia axyridis, Stethorus punctum picipes (Coleoptera: Coccinellidae), Orius tristicolor (Hemiptera: Anthocoridae), and Geocoris spp. (Hemiptera: Geocoridae). AB - The effect of buprofezin, a chitin synthesis inhibitor, on development and survival of immature stages of Harmonia axyridis (Pallas), Stethortus punctum picipes Casey, Orius tristicolor (White), Geocoris pallens Stal, and Geocoris punctipes (Say), was examined in a series of laboratory bioassays. Very few H. axyridis larvae (3.1%) treated with buprofezin reached adulthood, although 65% of treated pupae emerged successfully. Buprofezin caused no mortality to eggs of S. punctum picipes but 71.1% of treated early instar larvae failed to complete development. Eighty percent of treated late instars and 92.3% of pupae produced viable adults. Early instar nymphs of O. tristicolor were unaffected by buprofezin, whereas 47.7 and 85% of G. punctipes and G. pallens nymphs, respectively, failed to complete development. Treated eggs of G. pallens hatched successfully. The use of buprofezin in integrated pest management in Washington state wine grapes is discussed. PMID- 15279270 TI - Effects of azadirachtin/Neemazal on different stages and adult life table parameters of Trichogramma cacoeciae (Hymenoptera: Trichogrammatidae). AB - The effects of azadirachtin/Neemazal on adults, emergence, and life table parameters of Trichogramma cacoeciae Marchal were studied. The adults were exposed to fresh residues of the insecticide applied on glass plates. Based on the dose-response study, the LC50 value was 1330 ppm or 13.3 microg (AI)/ml. The effect of Neemazal on three developmental stages of the parasitoid was tested by dipping parasitized Sitotroga cerealella (Olivier) and Cydia pomonella (L.) eggs at the field-recommended concentration 3, 6, and 9 d after parasitization corresponding to larval, prepupal, and pupal stages. The emergence of adult parasitoids was adversely affected in both hosts, but the adverse effect was more in S. cerealella eggs compared with C. pomonella. The adult emergence was reduced by 73.3 and 33.76% in Sitotroga and Cydia eggs compared with controls, respectively. Longevity and progeny production of the emergent adults did not differ significantly from control. Neemazal affected stable population parameters (r(m), T, and DT) significantly. The intrinsic rate of increase for the control and Neemazal-exposed populations was 0.340 and 0.335 female offspring per female per day, respectively. Because some of postemergence life table parameters of adults were significantly reduced by the insecticide treatment, emergence rate of the parasitoid from treated eggs is not an adequate measure of ecological selectivity, and field studies are needed to determine the actual impact of neem on T. cacoeciae. PMID- 15279271 TI - Toxicity of spinosad in protein bait to three economically important tephritid fruit fly species (Diptera: Tephritidae) and their parasitoids (Hymenoptera: Braconidae). AB - The feeding toxicity of the natural insecticide spinosad in Provesta protein bait was evaluated for three economically important fruit fly species, the Mediterranean fruit fly, Ceratitis capitata (Wiedemann); the melon fly, Bactrocera cucurbitae Coquillett; and the oriental fruit fly, Bactrocera dorsalis Hendel. Both females and males were evaluated. Spinosad was remarkably similar in toxicity to all three fruit fly species. Male C. capitata (24 h LC50 values and 95% fiducial limits = 2.8 [2.60-3.0] mg/liter spinosad) were significantly, although only slightly more susceptible to spinosadthan females (4.2 [3.8-4.6] mg/liter). Male (5.5 [4.7-6.6] mg/liter) andfemale (4.3 [3.7-4.9] mg/liter) B. cucurbitae were equally susceptible to spinosad. Female (3.3 [3.1-3.6] mg/liter) and male (3.1 [2.9-3.3] mg/liter) B. dorsalis also were equally susceptible to spinosad. Provesta bait containing spinosad also was evaluated against two parasitoids of tephritid fruit flies, Fopius arisanus (Sonan) and Pysttalia fletcheri (Silvestri). These parasitoids did not feed on the bait, so a contact toxicity test was conducted. Significant amounts of mortality were found only after exposure of parasitoids to spinosad-coated glass vials with concentrations > or =500 mg/liter spinosad. Parasitoids were less susceptible than fruit flies to such a degree that use of spinosad in bait spray should be compatible with these parasitoid species. Because the fruit flies tested in this study were so susceptible to spinosad, this product seems to be promising as a bait spray additive and a replacement for malathion for control of these species. PMID- 15279272 TI - Behavioral and developmental effects of neem extracts on Clavigralla scutellaris (Hemiptera: Heteroptera: Coreidae) and its egg parasitoid, Gryon fulviventre (Hymenoptera: Scelionidae). AB - Extracts of neem, Azadirachta indica A. Juss, negatively affected feeding and development of Clavigralla scutellaris (Westwood), a coreid pest of pigeonpea, Cajanus cajan (L.) Millspaugh. Labial dabbing, pod wall penetration, and seed damage by fifth instars were significantly reduced on beans, Phaseolus vulgaris (L.), that had been dipped in aqueous, methanolic, or hexane extracts of neem seed kernel. When fourth instars were dipped directly into aqueous extract, developmental abnormalities of the wings occurred at all levels tested and fecundity dropped to zero at concentrations above 0.3125%. The LC50 value was 3.14% (220 ppm azadirachtin) at 8 d. The scelionid wasp Gryon fulviventre (Crawford) is an important natural enemy of Clavigralla spp.; egg mortality from this parasitoid ranged from 37 to 85% during the fall cropping season. Feeding by newly emerged wasps was dramatically reduced when honey was mixed with aqueous neem suspension, but 6-d survivorship of adults did not differ significantly from that of the control. Wasp oviposition behavior was altered slightly when coreid eggs were treated with neem: the period of antennation was significantly extended, but time for drilling, oviposition, and marking was unaffected. Neem dipped eggs were accepted for oviposition and progeny emerged successfully from these treated eggs. Exposure of already parasitized eggs to neem did not interfere with progeny emergence, longevity, or sex ratio. Thus, neem extract and egg parasitoids seem to be compatible and promising control strategies for C. scutellaris. Our results suggest that use of neem against pod-sucking bugs will not interfere with natural control provided by G. fulviventre. PMID- 15279273 TI - Injury to preflowering and flowering cotton by brown stink bug and southern green stink bug. AB - The impact of brown stink bug, Euschistus servus (Say), and southern green stink bug, Nezara viridula (L.), injury was evaluated on preflowering and flowering cotton, Gossypium hirsutum L., plants in no-choice tests. Vegetative stage cotton seedlings and reproductive structures, including flower buds (square) and bolls, were infested with adults and/or nymphs of both species. There were no significant differences in height, height to node ratio, square retention, and flower initiation for cotton seedlings or plants with a match-head square between southern green stink bug adult- or brown stink bug adult-infested and noninfested treatments. Abscission for individual large squares (precandle) and multiple squares (medium and small square on the same sympodial branch) was not significantly different among infested and noninfested treatments for the following species and developmental stages: brown stink bug adults, southern green stink bug adults, and third and fourth to fifth instar southern green stink bug nymphs. In boll infestation studies, the relationship between boll maturity, expressed as heat units beyond anthesis, and boll growth, abscission, hard locked carpels, seedcotton yield, and seed germination was measured. Brown stink bug induced abscission in bolls that had accumulated > 0-350 heat units beyond anthesis. Boll growth and seedcotton yield was significantly lower for bolls infested with brown stink bug through 266.5 and 550 heat units beyond anthesis, respectively. The proportion of hard locked carpels per boll was significantly greater for the infested treatment in a cohort of bolls that accumulated from 51 to 400 heat units beyond anthesis. Seed germination in bolls infested with brown stink bug was significantly lower in bolls aged 101-600 heat units beyond anthesis. PMID- 15279274 TI - Potential for transport of boll weevils (Coleoptera: Curculionidae) to the cotton gin within cotton modules. AB - There is concern that cotton gins located in boll weevil, Anthonomus grandis grandis Boheman, eradication zones serving customers in adjacent infested zones may serve as a site for boll weevil reintroductions if weevils are transported alive inside cotton modules. We surveyed fields in three distinct areas of Texas and found that weevils can be present in large numbers in cotton fields that have been defoliated and desiccated in preparation for harvest, both as free adults and as immatures inside unopened bolls. Harvested cotton taken from module builders indicated that approximately = 100-3700 adult boll weevils were packed inside modules constructed at the sampled fields. Marked weevils were forced through a laboratory field cleaner (bur extractor) commonly mounted on stripper harvesters, and 14% were recovered alive in the seed cotton fraction and lived at least to 24 h. Survival of weevils placed inside modules declined over time up to 7 d, but the magnitude of the decline varied with experimental conditions. In one experiment, 91% of the weevils survived to 7 d, whereas under harsher environmental conditions, only 11% survived that long. Together, our results indicate that when cotton is harvested in an infested area, boll weevils likely will be packed alive into cotton modules, and many will still be alive by the time the module is fed into the gin, at least up to 7 d after the module's construction. PMID- 15279275 TI - A new method for loading Bursaphelenchus xylophilus (Nematoda: Aphelenchoididae) on adult Monochamus alternatus (Coleoptera: Cerambycidae). AB - A new method was developed for loading the pinewood nematode, Bursaphelenchus xylophilus (Steiner et Buhrer) Nickle, on the beetle Monochamus alternatus Hope. Postdiapause beetle larvae were sterilized with 70 and 99.9% aqueous ethanol and placed singly in flasks where B. xylophilus reproduced on the fungus Ophiostoma minus (Hedgcock) H. et P. Sydow that had been grown on autoclaved barley grain and Pinus densifiora Sieb. et Zucc. wood chips. The fungus produced a large nematode population that developed to a high proportion of third-stage dispersal juveniles that molted to the fourth-stage dispersal juveniles. The survival rate was 80%, and the mean nematode load was 10,096. It took a mean of 5 wk to obtain the nematode-infested beetles after the initiation of nematode rearing. PMID- 15279276 TI - Sex pheromone of Argyrotaenia pomililiana (Lepidoptera: Tortricidae), a leafroller pest of apples in Argentina. AB - Sex pheromone gland extracts of Argyrotaenia pomililiana Trematerra & Brown females contained seven 14-chain compounds, the Z and E isomers of 11 tetradecenyl acetate, 11-tetradecen-1-ol, and 11-tetradecenal, respectively, together with tetradecyl acetate. In field trapping tests, a 100:5 blend of Z11 14:Ac and Z11-14:Al was shown to be suitable for detection and monitoring of A. pomililiana. This species-specific lure will facilitate the use of mating disruption against codling moth, Cydia pomonella (L.), in Argentine fruit orchards. PMID- 15279277 TI - Effects of fungicide residues on the survival, fecundity, and predation of the mites Tetranychus urticae (Acari: Tetranychidae) and Galendromus occidentalis (Acari: Phytoseiidae). AB - Representative fungicides from three or four families used for management of powdery mildew and other diseases in tree fruits were evaluated for their effects on a common spider mite and predator mite species, respectively. A modified Munger cell technique was effective in measuring the response of phytophagous and predaceous mites to fungicide residues on detached leaves in the laboratory. Demethylation-inhibiting (DMI) (imidazole [triflumazole] and triazole [myclobutanil]) and strobilurin (trifloxystrobin) fungicides were not toxic to female Tetranychus urticae Koch and Galendromus occidentalis (Nesbitt), and no sublethal effects were found on fecundity and predation rate after 3-5-d exposure to residues. Benomyl, a benzimidazole fungicide, increased adult mortality and reduced fecundity for both mite species; however, it did not alter the predation rate of G. occidentalis females on T. urticae eggs and larvae. Female G. occidentalis that survived the lethal effects of benomyl and the comparison acaricide pyridaben were unimpaired in predation. Our results for benomyl substantiate those of earlier studies and provide evidence for nontoxic effects of DMI and strobilurin fungicides on mites. We propose that DMI and strobilurin fungicides are a good fit for integrated mite management programs due to conservation of phytoseiid predatory mites. PMID- 15279278 TI - Utilization of blueberry by the lappet moth, Streblote panda Hubner (Lepidoptera: Lasiocampidae): survival, development, and larval performance. AB - The lappet moth, Streblote panda Hubner [1820] (Lasiocampidae), is a common species found in blueberry, Vaccinium spp. (Ericaceae) fields of Western Andalusia. The biology of this species as well as the extent to which its larvae can use and survive on blueberry is unknown. In this study, the suitability to larvae of several blueberry cultivars was studied. Larvae were grown under controlled laboratory conditions on excised foliage of six blueberry cultivars. Survival, development, and food use were determined for first and fifth instars. According to our results, blueberry has become an alternative host plant for S. panda in southwestern Andalusia. Low growth rates and efficiencies of use of food were observed. Lower gross efficiency of growth was found for larvae fed blueberry 'Sharpblue', despite a higher apparent digestibility of this cultivar. Larvae reared on this cultivar had the highest mortality, increased developmental time, and used a greater part of metabolism for maintenance. Herbivore pressure may be increased with the widespread planting of the most suitable cultivars 'Misty' and 'O'Neal', whereas 'Sharpblue' and'Climax' seem to be the least suitable host plants. These data provide useful information for planning and managing blueberry orchards in the presence of S. panda populations. PMID- 15279279 TI - Application of molecular techniques to distinguish Liriomyza trifolii from L. sativae (Diptera: Agromyzidae) on tomato cultivation in Japan. AB - A molecular method is applied for differentiating the morphologically related leafminers Liriomyza trifolli and L. sativae on tomato cultivation. The method requires multiplex polymerase chain reaction (PCR) amplification of a region of mitochondrial cytochrome oxidase DNA using multiprimer sets. The method divides the mitochondrial fragment of L. trifolli into two fragments and L. sativae into three fragments. It is faster, less costly, and easier than random amplified polymorphic DNA-PCR, PCR-restriction fragment-length polymorphism, and DNA sequencing and more sensitive than the enzyme electrophoresis method, which are other ways to differentiate these two species. We applied the method to samples from populations of another place, sex, and stage and obtained the same results. PMID- 15279280 TI - Effect of sucrose octanoate on survival of nymphal and adult Diaphorina citri (Homoptera: Psyllidae). AB - Asian citrus psylla, Diaphorina citri Kuwayama (Homoptera: Psyllidae) was detected for the first time in the United States near Delray Beach, FL, on 2 June 1998 and is continuing to spread and multiply throughout southern Florida. This psyllid is the vector of Liberobacter asiaticum, a phloem-limited bacterium that causes citrus greening disease. This pathogen has not been found in the Western Hemisphere to date. Furthermore, high infestation levels of D. citri can impact citrus plant health, fruit quality, or yield. Replicated laboratory and spray booth bioassays were conducted to determine the insecticidal activity of a synthetic analog of natural sugar esters found in leaf trichomes of wild tobacco, Nicotiatna gossei Domin, to nymphal and adult D. citri. Field trials were initiated in Fort Pierce, FL, in 2000 to determine activity of the sugar ester formulation (sucrose octanoate) on D. citri and other citrus pests, including immature Asian citrus leafminer, Phyllocnistis citrella Stainton and mites. Sucrose octanoate rates tested ranged from 400 to 8000 ppm (0.1-2% formulated product). Our data suggest that both nymphal and adult D. citri as well as the mite complex tested would be equally controlled to levels of >90% at the higher concentrations of sucrose octanoate and that good coverage is key to efficacy. PMID- 15279281 TI - Seasonal susceptibility of 'Bartlett' pears to codling moth (Lepidoptera: Tortricidae) infestation and notes on diapause induction. AB - Seasonal susceptibility of 'Bartlett' pear, Pyrus communis L., to codling moth, Cydia pomonella (L.), infestation, successful completion of larval development after infestation, and the induction of C. pomonella diapause was studied from 1992 through 1995. The seasonal variation in C. pomonella infestation and larval survival were effected by changes in fruit maturity. In late May through mid June, pears were hard and were not as successfully infested by C. pomonella and produced less larvae compared with fruit later in the season. In late June to mid July, pears became more suitable for infestation and a greater percentage of the larvae completed their development. In late July through mid-August, pears were susceptible to infestation, but the larvae were less likely to successfully complete development than in the late June to mid-July period due to pear tissue breakdown. From mid-August through September, pears are unsuitable for infestation, and few larvae were produced. When fruit were infested with neonate larvae in late May and mature larvae emerged from the fruit in July, a low percentage of the larvae entered diapause. However, when fruit were infested with neonate larvae in early July and mature larvae emerged from the fruit in early August, the majority of the larvae entered diapause. When fruit were infested with neonate larvae in late July through September and mature larvae emerged from the fruit after mid-August, nearly all C. pomonella larvae had entered diapause. PMID- 15279282 TI - Potential resistance of crape myrtle cultivars to flea beetle (Coleoptera: Chrysomelidae) and Japanese beetle (Coleoptera: Scarabaeidae) damage. AB - Field and laboratory studies were conducted to identify potential resistance among crape myrtles, Lagerstroemia spp., to Japanese beetle, Popillia japonica Newman and to flea beetles, Altica spp. Damage ratings revealed variation among cultivars in susceptibility to beetle feeding. Cultivars with Lagerstroemia fauriei Koehne in their parentage exhibited the least amount of damage in choice and no-choice experiments, with few exceptions. The data indicate that both beetle species cause more feeding damage on certain cultivars of Lagerstroemia indica L., such as 'Country Red', 'Twilight', and 'Carolina Beauty' than interspecific cultivars with L. fauriei in their parentage, such as 'Natchez', 'Tonto', and 'Muskogee'. When comparing the effect of parentage on all of the major pests of crape myrtle, L. faurei confers resistance to all pests except crape myrtle aphid. No correlation was found between leaf toughness, leaf color, and leaf nutrients in estimating flea beetle cultivar preference. With this information, growers can more effectively target scouting measures to the most susceptible cultivars. and breeders can select plants that will require the fewest chemical inputs. PMID- 15279283 TI - Effects of ground cover treatments and insecticide use on population density and damage caused by Lygus lineolaris (Heteroptera: Miridae) in apple orchards. AB - We conducted a 2-yr study in commercial apple orchards in Nova Scotia to assess the effects of ground cover treatments and insecticides on population density and fruit injury caused by tarnished plant bug, Lygus lineolaris (Palisot de Beauvois). The design was a split-plot with insecticides applied to whole orchard blocks and ground cover treatments applied to plots nested within orchard blocks. Ground cover treatments were 1) standard herbicide use, 2) enhanced weed control in tree rows, and 3) treatment two plus use of a selective herbicide in laneways. Treatments had few significant effects on vegetation in the tree row, but in laneways, known dicot hosts of L. lineolaris were suppressed and nonhost grasses promoted with treatment 3. Ground cover treatments did not affect cumulative captures of adult tarnished plant bugs on white sticky traps located in the plots but did affect captures in sweep nets. Split-plot ANOVA indicated no significant effect of insecticides on injury in either year, but ground cover treatments were significant in 2001. The lowest ranking rates of injury in both years were in orchards treated before bloom with a pyrethroid insecticide, either cyhalothrin lambda or cypermethrin. The highest ranking rate of injury occurred in an orchard where insecticide was not applied until after bloom despite a high prebloom capture of L. lineolaris adults on orchard perimeter sticky traps. Fruit injury values for the ground cover treatment 3 were 63.3% (n.s.) and 50.0% (P < 0.05), respectively, of those in the standard treatment in 2000 and 2001. PMID- 15279284 TI - Binomial sequential sampling plans for late instars of European corn borer (Lepidoptera: Crambidae), corn earworm (Lepidoptera: Noctuidae), and damaged kernels in sweet corn ears. AB - Late-season infestations of European corn borer, Ostrinia nubilalis (Hubner), and corn earworm, Helicoverpa zea (Boddie), were sampled to develop binomial sequential sampling plans for larval infestations and damaged kernels in sweet corn, Zea mays L., ears, near harvest. Fields were sampled to obtain a range of larval densities likely to be encountered over a range of infestation levels and field conditions. Binomial sampling plans were developed for O. nubilalis larvae, H. zea larvae, O. nubilalis, and H. zea larvae combined, and for damaged sweet corn kernels. Observed densities ranged from 0.01 to 4.40 larvae per ear for O. nubilalis, 0.005-1.62 larvae per ear for H. zea, and 0.004-36.12 damaged kernels per ear. Results of resampling analyses, based on the proportion of ears infested with one or more larvae, or damaged kernels, indicated an average sample size of 34-37 ears was necessary to classify whether larval infestations, or the incidence of damaged kernels, exceeded 5%. Two operating characteristic curves are presented for each of the four sampling plans. Initial results, with upper bounds of 0.10, and alpha (type I) and beta (type II) error rates at 0.10 and 0.05, respectively, resulted in a 90% probability of making the correct management decision at infestation levels >10%. To improve performance of the sampling plans, we modified the binomial plans by reducing the upper bound to 0.075, while maintaining the same error rates. This plan resulted in a higher probability (>95%) of making the correct management decision to reject a sweet corn load when infestation levels are >10%. PMID- 15279285 TI - Effects of moisture on the toxicity of inorganic and organic insecticidal dust formulations to German cockroaches (Blattodea: Blattellidae). AB - Toxicity of boric acid (40-99% [AI]), silica gel, eugenol, and deltamethrin dust formulations to adult male German cockroaches, Blattella germanica (L.), was evaluated at five different relative humidities ranging from 0 to 100% and in the presence of 0 to 1 ml of water. Victor boric acid dust was generally the most toxic boric acid formulation at all relative humidities, despite having the lowest percentage (40%) of boric acid; however, this was the only formulation to have sucrose and other edible ingredients. There was no consistent effect of relative humidity on dust toxicity; LT50 values of Roach Prufe (98% boric acid), Victor, and Drione (silica gel and synergized pyrethrins) increased significantly linearly with relative humidity, whereas other formulations were unaffected. The LT50 values of all boric acid-based dust formulations declined exponentially when wetted with increasing volumes of water. Water did not affect the toxicity of deltamethrin and eugenol dusts, but it caused a linear decline in toxicity of silica gel (Dri-Die). The toxicity of a formulation containing silica gel and synergized pyrethrins (Drione) increased exponentially with increasing amounts of water. Moisture in the form of relative humidity does not strongly affect the toxicity of most insecticidal dust formulations. Presence of water, however, increases the toxicity of boric acid dusts and Drione. Toxicity of the hydrophobic deltamethrin (DeltaDust) and eugenol (EcoPCO D) dusts were unaffected by water. PMID- 15279286 TI - Delayed toxicity as a critical factor in the efficacy of aqueous baits for controlling Argentine ants (Hymenoptera: Formicidae). AB - Boric acid, imidacloprid, and thiamethoxam in sucrose aqueous baits had different delayed toxicities to worker Argentine ants, Linepithema humile (Mayr). The concentrations required to produce an LT50 (time required to produce 50% mortality) within 1-4 d were 3.63-0.55% boric acid, 9.2 x 10(-3) to 7.1 x 10(-4)% imidacloprid, and 3 x 10(-4) to 2 x 10(-5)% thiamethoxam. The three toxicants were not repellent. Other laboratory trials showed that 1% boric acid, 5 x 10(-4) to 5 x 10(-3)% imidacloprid, and 1 x 10(-5) to 1 x 10(-3)% thiamethoxam had delayed toxic effects, whereas 0.5% boric acid and < 5 x 10(-3)% imidacloprid did not. Baits that provided an LT50 between days 1 and 4 were considered to have delayed toxic effects. The utility of aqueous sucrose baits and toxicants soluble in such systems and the negative impact of fast-acting toxicants on trail following, recruitment, trophallaxis, and control of Argentine ants are discussed. PMID- 15279287 TI - Effect of a lignin-degrading fungus on feeding preferences of Formosan subterranean termite (Isoptera: Rhinotermitidae) for different commercial lumber. AB - The feeding preferences of the Formosan subterranean termite, Coptotermes formosanus Shiraki, for commercial lumber Alaska yellow cedar, Chamaecyparis nootkatensis (D. Don) Spach; yellow birch, Betula alleghaniensis Britton; northern red oak, Quercus rubra L.; redwood, Sequoia sempervirers (D. Don) Endl; and spruce (Picea spp.) were examined to determine whether the presence of the lignin-degrading basidiomycete Marasmiellus troyanus (Murrill) Singer could alter the relative preference of termites for these wood species. In paired choice tests with fungus-inoculated sawdust versus control sawdust, termites showed a strong preference for the fungus-inoculated sawdust for all wood species tested, except for Alaska yellow cedar. In a multiple-choice test using sawdust without fungus, termites showed a very strong preference for red oak sawdust over the other three species. In a paired choice test using fungus-inoculated sawdust, termites showed a preference for redwood over red oak sawdust. In a feeding test using autoclaved wood blocks without fungal decay, there was no difference in termite consumption of birch, red oak, or redwood. The relative preference of termites for redwood increased when blocks were decayed by M. troyanus for 3 and 8 wk. These results indicate that chemical modifications due to fungal decay affected the feeding preference of termites for different commercial lumber. PMID- 15279288 TI - Frequency of kdr gene in house fly field populations: correlation of pyrethroid resistance and kdr frequency. AB - The frequency of the L1014 F kdr mutation was determined in 14 field populations of house flies, Musca domestica L., with resistance factors at LD50 for pyrethrin/piperonyl butoxide and bioresmethrin/piperonyl butoxide from 4 to 29 and 2 to 98, respectively. A polymerase chain reaction test for identifying kdr homo- or heterozygote house flies was used to determine the frequency of kdr. The L1014 F allele was found in all populations tested. The frequency of kdr in the field populations was high and varied from 0.46 to 0.99. Eleven of the populations were in Hardy-Weinberg equilibrium, whereas two strains had higher number of heterozygotes than expected, indicating a possible heterozygote advantage. The frequency of kdr was strongly correlated with the reduced mortality observed in the bioassays with pyrethrum and bioresmethrin synergized by piperonyl butoxide. This indicates that kdr is a major mechanism for pyrethroid resistance in these field populations. Five field populations had resistance factors >25 and >10 for bioresmethrin/piperonyl butoxide and pyrethrin/piperonyl butoxide, respectively. The frequencies of kdr in these five populations varied from 0.89 to 0.99. The frequencies of kdr in the field populations showing no or a low level of resistance had frequencies of kdr from 0.46 to 0.75, which indicates that the L1014 F kdr allele is a fully recessive genetic trait in house flies. We have shown that the molecular diagnostic PASA method to determine the resistance phenotypes and the frequency of kdr is a powerful tool, which could be used to get information to make recommendations about pest and resistance management. PMID- 15279289 TI - Susceptibility of spinosad in Musca domestica (Diptera: Muscidae) field populations. AB - The toxicity of spinosad was determined in one susceptible and five insecticide resistant laboratory strains of house fly, Musca domestica L. Spinosad was relatively slow-acting, but highly toxic to house flies. In a feeding bioassay, spinosad LC50 at 72 h was 0.51 microg of spinosad per gram of sugar, making it 6.3- and 3.5-fold more toxic to house flies compared with azamethiphos and methomyl, respectively. In topical application bioassay, the LD50 at 48 h of spinosad in susceptible house flies was 40 ng per 20 mg of house fly, making spinosad less toxic than the pyrethroid bioresmethrin synergized by piperonyl butoxide and the organophosphate dimethoate. The insecticide-resistant laboratory strains had resistance factors to spinosad at LC50 in feeding bioassay from 1.5 to 5.5 and at LD50 in topical application bioassay from 2.5 to 4.7, indicating that in house fly cross-resistance to the major insecticide classes will not initially be of major concern for the use of spinosad for house fly control. The toxicity of spinosad was also evaluated against 31 field populations of house flies collected from livestock farms across Denmark. The field populations were 2.2- to 7.5-fold resistant to spinosad at 72 h in feeding bioassay, but based on steep slopes in the bioassay and the limited variation of spinosad toxicity against the various field populations, we consider the field populations to be spinosad-susceptible. We propose a diagnostic dose of 12 microg of spinosad per gram of sugar in feeding bioassay with impregnated sugar for determination of resistant house flies, which is 10x the LC95 of the susceptible strain WHO and approximately = 2x the LD95 of the field populations. Spinosad showed no substantial cross-resistance to the pyrethroid bioresmethrin synergized by piperonyl butoxide, the anticholinesterases dimethoate, azamethiphos, methomyl, and spinosad in house fly field populations. PMID- 15279290 TI - Cross-resistance of Cry1Ab-selected Ostrinia nubilalis (Lepidoptera: Crambidae) to Bacillus thuringiensis delta-endotoxins. AB - Corn plants expressing the toxin from Bacillus thuringiensis (Berliner) have proven to be effective in controlling lepidopteran pests such as the European corn borer, Ostrinia nubilalis (Hubner) (Lepidoptera: Crambidae). Several Bt toxins are being tested and incorporated into crop genomes, although tests for cross-resistance among different toxins have been limited by a lack of resistant colonies. Four different colonies of O. nubilalis selected with full-length Cry1Ab incorporated into artificial diet developed significant levels of resistance (2.0- to 10-fold) within 10 generations. Additionally, selection with Cry1Ab resulted in decreased susceptibility to a number of other toxins to which the selected colonies were not previously exposed. Significantly, levels of resistance were highest to Cry1Ac with resistance ratios up to 51.0-fold. Low levels (less than five-fold) of cross-resistance were detected with Cry1F. In contrast, Cry9C susceptibility was unaffected by selection with Cry1Ab. These results indicate that the availability of multiple toxins could improve resistance management strategies, provided cross-resistance among toxins is not a factor. PMID- 15279291 TI - Genetics of resistance to transgenic Bacillus thuringiensis poplars in Chrysomela tremulae (Coleoptera: Chrysomelidae). AB - The area under genetically engineered plants producing Bacillus thuringiensis (Bt) toxins is steadily increasing. This increase has magnified the risk of alleles conferring resistance to these toxins being selected in natural populations of target insect pests. The speed at which this selection is likely to occur depends on the genetic characteristics of Bt resistance. We selected a strain of the beetle Chrysomela tremulae Fabricius on a transgenic Bt poplar clone Populus tremula L. x Populus tremuloides Michx producing high levels of B. thuringiensis Cry3Aa toxin. This strain was derived from an isofemale line that generated some F2 offspring that actively fed on this Bt poplar clone. The resistance ratio of the strain was >6400. Susceptibility had decreased to such an extent that the mortality of beetles of the strain fed Bt poplar leaves was similar to that of beetles fed nontransgenic poplar leaves. Genetic crosses between susceptible, resistant, and F1 hybrids showed that resistance to the Cry3Aa toxin was almost completely recessive (D(LC) = 0.07) and conferred by a single autosomal gene. The concentration of Cry3Aa produced in the transgenic Bt poplar used in this study was 6.34 times higher than the LC99 of the F1 hybrids, accounting for the complete recessivity (D(ML) = 0) of survival on Bt poplar leaves. Overall, the genetic characteristics of the resistance of C. tremulae to the Cry3Aa toxin are consistent with the assumptions underlying the high-dose refuge strategy, which aims to decrease the selection of Bt resistance alleles in natural target pest populations. PMID- 15279292 TI - Field evaluation of emmer wheat-derived synthetic hexaploid wheat for resistance to Russian wheat aphid (Homoptera: Aphididae). AB - Broadening the genetic base for resistance to Russian wheat aphid, Diuraphis noxia (Mordvilko) (Homoptera: Aphididae), in bread wheat, Triticum aestivum L., is desirable. To date, identified Russian wheat aphid resistance genes are either located to the D chromosomes or to rye translocation of wheat, and resistance derived from the A or B genomes of tetraploid Triticum spp. would therefore be highly beneficial. Fifty-eight synthetic hexaploid wheat, derived from interspecific crosses of Triticum dicoccum Schrank. and Aegilops tauschii (Coss.) Schmal. and their parents were evaluated for resistance to Russian wheat aphid under field conditions. Plots infested with aphids were compared with plots protected with insecticides. The T. dicoccum parents were highly resistant to Russian wheat aphids, whereas the Ae. tauschii parents were susceptible. Resistance levels observed in the synthetic hexaploids were slightly below the levels of their T. dicoccum parents when a visual damage scale was used. but no major resistance suppression was observed among the synthetics. Russian wheat aphid infestation on average reduced plant height and kernel weight at harvest in the synthetic hexaploids and the T. dicoccum parents by 3-4%, whereas the susceptible control 'Seri M82' suffered losses of above 20%. Because resistance in the synthetic hexaploid wheat is derived from their T. dicoccum parent, resistance gene(s) must be located on the A and/or B genomes. They must therefore be different from previously identified Russian wheat aphid resistance genes, which have all been located on the D genome of wheat or on translocated segments. PMID- 15279293 TI - Resistance of Glycine species and various cultivated legumes to the soybean aphid (Homoptera: Aphididae). AB - The soybean aphid, Aphis glycines Matsumura, is a new pest of soybean, Glycine max (L.) Merr., in North America. It has become widespread on soybean in North America since it was first identified in the Midwest in 2000. Species of Rhamnus L. (buckthorn) are the primary hosts of A. glycines, and soybean is known as a secondary host. There is limited information about the secondary host range of A. glycines. Aphid colonization on various legume hosts was compared in choice experiments. Aphid colonization occurred on species in the genus Glycine Wild. No colonization occurred on Lablab purpureus (L.) Sweet, Lens culinaris Medik, Phaseolus vulgaris L., Pisum sativum L., or species of Vicia L. and Vigna Savi. Colonization was limited or aphids were transient on species of Medicago L., Phaseolus L., and Trifolium L. There were significant differences in aphid colonization among Medicago truncatula accessions with numbers ranging from 7 to 97 aphids per plant. Six Glycine soja Sieb. & Zucc. accessions were as resistant as G. max accessions to A. glycines; these may represent novel sources of A. glycines resistance not found in G. max. Antibiosis was found to play a large role in the expression of resistance in three of the G. soja accessions. Results of this study indicated that G. max and G. soja were the best secondary hosts of A. glycines; however, its secondary host range may include other leguminous species. Therefore, A. glycines did not seem to have a highly restricted monophagous secondary host range. PMID- 15279294 TI - Tomato psyllid behavioral responses to tomato plant lines and interactions of plant lines with insecticides. AB - Adult tomato psyllid, Bactericerca (Paratrioza) cockerelli (Sulc) (Homoptera: Psyllidae), behavioral responses were evaluated for five tomato plant lines and for the interactions of insecticides with four commercial cultivars. Plant lines tested included the commercial 'Shady Lady', 'Yellow Pear', '7718 VFN', 'QualiT 21', and the plant introduction line PI 134417. Insecticides included a kaolin particle film, pymetrozine, pyriproxyfen, spinosad, and imidacloprid. Psyllids spent significantly more time feeding on 'Yellow Pear' than all other plant lines except '7718 VFN'. In comparisons among plant lines, psyllids exposed to the wild accession PI 1.34417 showed a 98% reduction in feeding, a significant increase in jumping behavior, and a significant tendency to abandon the leaves, thereby demonstrating repellency, not just an antixenosis response. Interactions between plant lines and insecticides influenced behavioral responses. All insecticides tested significantly reduced feeding durations on all cultivars except the preferred 'Yellow Pear'. However, nonfeeding activities such as walking, probing, resting, and jumping varied substantially with chemical and cultivar combination. The behavior assay results offered insight into host resistance mechanisms, provided a useful technique for measuring effects of interaction of plant lines with insecticides, and generated information for selecting insecticides for specific cultivars used in integrated pest management program for the tomato psyllid. PMID- 15279295 TI - Characterization of oxidative enzyme changes in buffalograsses challenged by Blissus occiduus. AB - This research investigated the role of oxidative enzymes in the defense response of buffalograss, Buchloe dactyloides (Nuttall) Engelmann, to Blissus occiduus Barber. Changes in catalase and peroxidase activity were observed in both resistant and susceptible buffalograsses in response to chinch bug feeding. Susceptible plants were shown to have a lower level of catalase activity compared with their respective control plants. By contrast, catalase activities of resistant plants were similar between infested and control buffalograsses throughout the study. Resistant plants had higher levels of peroxidase activity compared with their control plants, whereas peroxidase activities for control and infested susceptible plants remained at similar levels or were slightly lower for infested plants. These findings suggest that chinch bug feeding leads to a loss in catalase activity in susceptible buffalograsses. In contrast, resistant buffalograsses may be able to tolerate chinch bug feeding by increasing their peroxidase activity. Polyphenol oxidase activities were similar between control and infested plants for the buffalograsses evaluated. Among the enzymes examined, no differences in isozyme profiles for peroxidase and polyphenol oxidase were detected between control and infested 378, NE91-118, Cody, and Tatanka plants. Gels stained for catalase identified differences in the isozyme profiles of infested and uninfested 378 plants; however, infested and control NE91-118, Tatanka, and Cody plants has similar isozyme profiles. No differences in protein profiles were observed between chinch buginfested 378, NE91-118, Cody, and Tatanka plants and their respective uninfested controls. PMID- 15279296 TI - Influence of Bemisia argentifolii (Homoptera: Aleyrodidae) infestation and squash silverleaf disorder on zucchini seedling growth. AB - We investigated the effect of different levels of infestation by whiteflies, Bemisia argentifolii Bellows & Perring, on the growth and pigment concentrations of seedlings of zucchini, Cucurbita pepo L., that differed in their tolerance to squash silverleaf disorder. Genetically similar sister lines that were either tolerant (ZUC76-SLR) or susceptible (ZUC61) to silverleaf disorder exhibited reduced plant height, internode length, plant dry weight, and petiole length in response to whitefly feeding. Similar plant growth responses to whitefly feeding were observed despite that the foliage of ZUC61 silvered severely, whereas the foliage of ZUC76-SLR showed no silvering in a greenhouse experiment conducted in the spring and showed only minimal silvering in a similar greenhouse experiment conducted in the fall. In plants of both sister lines infested with 50 pairs of whiteflies and their progeny, petioles, but not the leaf blades, of uninfested leaves had reduced chlorophyll content. In another experiment, two different genetic sources of tolerance to silverleaf disorder (ZUC33-SLR/PMR and ZUC76-SLR) and a commercial silverleaf-susceptible zucchini hybrid ('Zucchini Elite') responded similarly to whitefly feeding, except the tolerant genotypes did not exhibit leaf silvering. All genotypes, silverleaf tolerant or not, had reduced dry weight, plant height, and internode length that became more pronounced as whitefly infestation increased. All genotypes had reduced levels of chlorophylls and carotenoids in uninfested young leaf blades and petioles from infested plants. Petioles, however, were more affected by feeding than leaf blades, showing a 66% reduction in chlorophylls a+b and carotenoids at the lowest infestation level (30 pairs of whitefly and their progeny), whereas pigments in leaf blades declined more slowly in response to whitefly feeding density, averaging 14-15% less at the highest infestation level (90 pairs of whitefly and their progeny). We conclude that tolerance to silverleaf disorder does not prevent stunting in zucchini seedlings nor does it protect against the systemic loss of photosynthetic and protoprotectant pigments induced by feeding of B. argentifolii whiteflies. PMID- 15279297 TI - Effect of three resistant soybean genotypes on the fecundity, mortality, and maturation of soybean aphid (Homoptera: Aphididae). AB - The fecundity, longevity, mortality, and maturation of the soybean aphid, Aphis glycines Matsumura (Homoptera: Aphididae), were characterized using three resistant soybean, Glycine max (L.) Merrill, genotypes ('Dowling', 'Jackson', and PI200538 'Sugao Zarai') and two susceptible genotypes ('Pana' and 'Loda'). Antibiosis in the resistant genotypes was demonstrated by a significant decrease in fecundity and longevity and increased mortality of A. glycines. Aphid fecundity, measured as number of offspring produced in the first 10 d by each viviparous aptera, was higher on Pana than on the resistant genotypes. Aphid longevity, the mean number of days a 1-d-old adult lived, was 7 d longer on Pana than on Dowling and Jackson. The mortality of both viviparous apterae and nymphs on resistant genotypes was significantly higher than on susceptible genotypes. A greater number of first instars survived to maturation stage (date of first reproduction) on susceptible plants than on resistant plants. None of the first instars placed on Dowling and PI200538 leaves survived to maturation. Observations of aphid behavior on leaves indicated that aphids departed from the leaves of resistant plants 8-24 h after being placed on them, whereas they remained indefinitely on leaves of susceptible cultivars and developed colonies. Reduced feeding due to ingestion of potentially toxic compounds in soybean may explain the possible mechanism of resistance to the soybean aphid. PMID- 15279298 TI - Identification of Russian wheat aphid (Homoptera: Aphididae) populations virulent to the Dn4 resistance gene. AB - The Russian wheat aphid, Diuraphis noxia (Mordvilko), is a serious worldwide pest of wheat and barley. Russian wheat aphid populations from Hungary, Russia, and Syria have previously been identified as virulent to D. noxia (Dn) 4, the gene in all Russian wheat aphid-resistant cultivars produced in Colorado. However, the virulence of Russian wheat aphid populations from central Europe, North Africa, and South America to existing Dn genes has not been assessed. Experiments with plants containing several different Dn genes demonstrated that populations from Chile, the Czech Republic, and Ethiopia are also virulent to Dn4. The Czech population was also virulent to plants containing the Dnx gene in wheat plant introduction PI220127. The Ethiopian population was also virulent to plants containing the Dny gene in the Russian wheat aphid-resistant 'Stanton' produced in Kansas. The Chilean and Ethiopian populations were unaffected by the antibiosis resistance in Dn4 plants. There were significantly more nymphs of the Chilean population on plants of Dn4 than on Dn6 plants at both 18 and 23 d postinfestation, and the Ethiopian population attained a significantly greater weight on Dn4 plants than on plants containing Dn5 or Dn6. These newly characterized virulent Russian wheat aphid populations pose a distinct threat to existing or proposed wheat cultivars possessing Dn4. PMID- 15279299 TI - Evaluation of sequential presence-absence sampling plans for the diamondback moth (Plutellidae: Lepidoptera) in cruciferous crops in Australia. AB - Two sets of sequential presence-absence sampling plans for decision-making in the management of diamondback moth, Plutella xylostella (L.), were developed and evaluated. One set of sampling plans targeted the classification of proportions of infested plants, and the other set of sampling plans targeted the classification of larval density. The action thresholds investigated were 0.15, 0.25, 0.35, and 0.45 proportion of plants infested with larvae, and 0.2, 0.4, 0.6, and 0.8 larvae per plant. They are representative of the action thresholds currently practiced by Australian crucifer growers. For each sampling plan, the population range within which a minimal correct decision rate of 95% can be expected at a maximal average sample size of 50 plants (OC95ASN50) was specified. The closeness of an OC95ASN50 range to the target action threshold is a measure of the expected performance of the sampling plan. A closer distance reflects better performance. The OC95ASN50 ranges of the proportion-classification sampling plans were within 33-53% of the target action thresholds. The width of these OC95ASN50 ranges represents 73-87% of the entire population range (0-1). For the classification of larval density, an empirical proportion-density model was first established using data from different states and different cruciferous crops. The OC95ASN50 ranges of the density-classification sampling plans were within 57-75% of the target action threshold. Simulated sampling of 20 independent data sets showed that for most data sets the correct classification rate was at least 98% and the matching average sample size was <50 plants. PMID- 15279301 TI - Accounting for cluster sampling in constructing enumerative sequential sampling plans. AB - Green's sequential sampling plan is widely used in applied entomology. Green's equation can be used to construct sampling stop charts, and a crop can then be surveyed using a simple random sampling (SRS) approach. In practice, however, crops are rarely surveyed according to SRS. Rather, some type of hierarchical design is usually used, such as cluster sampling, where sampling units form distinct groups. This article explains how to make adjustments to sampling plans that intend to use cluster sampling, a commonly used hierarchical design, rather than SRS. The methodologies are illustrated using diamondback moth, Plutella xylostella (L.), a pest of Brassica crops, as an example. PMID- 15279300 TI - Efficacy of 1,4-diaminobutane (putrescine) in a food-based synthetic attractant for capture of Mediterranean and Mexican fruit flies (Diptera: Tephritidae). AB - Field trials were conducted in Guatemala to evaluate the importance of 1,4 diaminobutane (putrescine) in traps baited with ammonium acetate, trimethylamine, and putrescine. For the Mediterranean fruit fly, Ceratitis capitata (Wiedemann), there were no differences in percentage of females captured in coffee and citrus or in percentage of males captured in citrus in traps with ammonium acetate and trimethylamine lures (females in coffee, 26.4 +/- 6.27%; females in citrus, 35.7 +/- 5.35%; males in citrus, 37.7 +/- 7.48%) versus ammonium acetate, trimethylamine, and putrescine lures (females in coffee, 36.6 +/- 9.64%; females in citrus, 41.1 +/- 5.18%; males in citrus, 37.1 +/- 6.09%). Percentage of males captured in coffee was reduced significantly when putrescine was not used with the ammonium acetate and trimethylamine (39.9 +/- 4.34 versus 31.6 +/- 5.29%). Lower percentages were captured in traps baited with ammonium acetate and putrescine, and the lowest percentages were captured in traps baited with putrescine and trimethylamine. When population level as indicated by capture in traps baited with ammonium acetate, trimethylamine, and putrescine was considered, a higher percentage of C. capitata males were captured in traps baited with all three components when one or more flies per trap per day were captured in coffee, and a higher percentage of females were captured when less than one fly per trap per day was captured in citrus. Percentage of the Mexican fruit fly, Anastrepha ludens (Loew), captured was significantly higher in traps baited with ammonium acetate and putrescine and significantly lower in traps baited putrescine and trimethylamine than in all other treatments. Results indicate that putrescine may be deleted when monitoring established populations of C. capitata but should be used in traps used to monitor A. ludens or to detect new infestations of C. capitata. PMID- 15279302 TI - Influence of adding borax and modifying pH on effectiveness of food attractants for melon fly (Diptera: Tephritidae). AB - The melon fly, Bactrocera cucurbitae (Coquillett) (Diptera: Tephritidae), is the most damaging pest of cucurbits in Reunion Island. The influence of adding borax and modifying pH on the effectiveness of different food attractants for both sexes of the melon fly is analyzed by a release-recapture method in field cages. Adding borax to protein hydrolysates Nulure and Buminal strongly reduced their attractiveness for B. cucurbitae. Acidification of 5% Buminal solution (from pH 6 to pH 3) doubled its attractiveness for melon fly. Conversely, Torula yeast at pH 10.5 was significantly more attractive than the standard Torula yeast at pH 9 (28% of captured flies compared with 17%). However, a further pH increase of the yeast solution does not improve its attractiveness. The results are discussed in relation to other studies on pH modification of various baits for Tephritidae. PMID- 15279304 TI - Movement and distribution of adult rusty grain beetle, Cryptolestes ferrugineus (Coleoptera: Laemophloeidae), in stored wheat in response to different temperature gradients and insect densities. AB - The movement and distribution of adult Cryptolestes ferrugineus (Stephens) (Coleoptera: Laemophloeidae) in grain provide important information for detection of insect pests and for simulations of their distribution in grain bins. Adult movement and distribution were determined in 100 by 100 by 1000-mm wheat (14.5 +/ 0.2% moisture content) columns at four insect densities, three temperature gradients, and dynamic (changing) temperature conditions. Insect density was a minor factor influencing insect movement and distribution in grain columns with temperature gradients. Dispersal resulted in a uniform distribution at a higher insect density (higher than two adults per kilogram of wheat), and aggregation occurred at a low insect density. Adults wandered in the first 6 h after introduction, and there were fewer adults wandering in the vertical direction than in the horizontal direction. Adults moved faster in the vertical direction than in the horizontal direction, and the maximum speed of the movement was 6 m/d in the horizontal direction, and >10.8 m/d in the vertical direction through wheat. Adults could detect temperature gradients in <1 h and preferred warmer temperatures when they had a choice. Insect distribution in horizontal wheat columns at any temperature gradient was unstable for 24 h. Twenty-four hours after introduction, adults gradually overcame their positive geotactic behavior if the upper temperature was more biologically suitable or was not <27.5 degrees C. Adults responded faster to higher temperature gradients than to lower temperature gradients. There was a similar pattern of adult distribution in 144 h. PMID- 15279303 TI - Biological activity of volatile di-n-propyl disulfide from seeds of neem, Azadirachta indica (Meliaceae), to two species of stored grain pests, Sitophilus oryzae (L.) and Tribolium castaneum (Herbst). AB - Head space volatiles, including 73% di-n-propyl disulfide, were collected from freshly crushed neem seeds. This compound along with previously reported diallyl disulfide (di-2-propenyl disulfide) were toxic when applied topically or as a fumigant to Tribolium castaneum adults and 8-, 12-, and 16-d-old larvae, and Sitophilus oryzae adults. Di-n-propyl disulfide significantly decreased the growth rate and dietary utilization with moderate inhibition of food consumption in both insects. The total coefficient of deterrence for this compound ranged between 68.5 and 178.6, which suggests that it has medium to very good deterrent activity vis-a-vis the treatment concentration and instar. Di-n-propyl disulfide and diallyl disulfide presented a similar effect on efficiency of conversion of ingested food, which is reduced 3-fold; this implies that both compounds are physiological toxicants. Present studies clearly demonstrate that di-n-propyl disulfide could be a potent toxicant, fumigant, and feeding deterrent for stored grain pests, if a suitable formulation and application procedure are developed. PMID- 15279305 TI - Chronological age-grading of three species of stored-product beetles by using near-infrared spectroscopy. AB - The accuracy of near-infrared spectroscopy (NIRS) for predicting the chronological age of adults of the rice weevil, Sitophilus oryzae (L.); the lesser grain borer, Rhyzopertha dominica (F.); and the red flour beetle, Tribolium castaneum (Herbst), three pests of stored grain, was examined. NIRS predicted age correlated well with actual age of these three species. Age predictions in S. oryzae by using the NIRS method are not dependent upon adult sex or temperatures to which adult weevils are exposed. Results indicated that water content decreased with increasing age in rice weevil adults, and excluding wavelengths at which water absorbs NIR radiation reduced the accuracy of correct classification. Additionally, removing cuticular lipids from insects resulted in a significant decrease in classification accuracy of weevils, indicating that these compounds may be partly responsible for the ability of NIRS to differentiate young from old beetles. NIRS is a nondestructive technique that can be used to age-grade large numbers of adult stored-product beetles, information that could help to increase the accuracy of population models for these pest species. PMID- 15279306 TI - Microwave radar detection of stored-product insects. AB - A microwave radar system that senses motion was tested for capability to detect hidden insects of different sizes and activity levels in stored products. In initial studies, movements of individual adults or groups of Lasioderma serricorne (F.), Oryzaephilus surinamensis (L.), Attagenus unicolor (Brahm), and Tribolium castaneum (Herbst) were easily detected over distances up to 30 cm in air. Boxes of corn meal mix and flour mix were artificially infested with 5-100 insects to estimate the reliability of detection. The likelihood that a box was infested was rated by the radar system on a quantitative scale. The ratings were significantly correlated with the numbers of infesting insects. The radar system has potential applications in management programs where rapid, nondestructive targeting of incipient insect infestations would be of benefit to the producers and consumers of packaged foods. PMID- 15279307 TI - Survival of stored-product insect natural enemies in spinosad-treated wheat. AB - The survival of stored product insect natural enemies in wheat treated with spinosad was investigated in laboratory and pilot scale experiments. The predator Xylocoris flavipes (Reuter), the warehouse pirate bug, and the parasitoids Habrobracon hebetor (Say), Theocolax elegans (Westwood), and Anisopteromalus calandrae (Howard) were exposed to wheat treated with aliquots of water or spinosad at 0.05-1 mg ([AI])/kg. X. flavipes was the only species that survived (92% survival) in spinosad-treated wheat at 1 mg/kg. X. flavipes suppressed populations of immature Tribolium castaneum (Herbst), the red flour beetle, by nearly 90% compared with a water-treated control, but 100% suppression of immatures was achieved in wheat receiving spinosad or spinosad + X. flavipes treatments. A 3-mo pilot scale experiment to evaluate T. castaneum suppression in drums holding 163.3 kg of wheat showed that the pest populations increased throughout the study in the control treatment, but peaked after 1 mo in the X. flavipes-treated drums. By comparison, better T. castaneum population suppression was achieved in spinosad or spinosad + X. flavipes treatments. Although X. flavipes can survive and reproduce in spinosad-treated wheat, under our test conditions spinosad alone provided adequate suppression of T. castaneum populations in stored wheat. PMID- 15279308 TI - Postlarval fitness of transgenic strains of Cochliomyia hominivorax (Diptera: Calliphoridae). AB - Eight transgenic strains of Cochliomyia hominivorax (Coquerel) (Diptera: Calliphoridae) were compared with the wild-type parental laboratory strain (P95) in colony. Measurements of average weight of pupae, percentage of adults emerging from pupae, ratio of males to total emerged adults, and mating competitiveness were analyzed. The parental strain colony was subcultured and exposed to handling procedures equivalent to transgenic strains for valid comparison of overall colony fitness. None of the transgenic colonies exhibited significantly lower fitness characteristics than the control parental colony. One transgenic colony had a higher ratio of adults emerging from pupae, and five colonies had higher average pupal weight; because fitness cost would only be indicated by lower values, the statistical variations were not significant. Males of one transgenic strain were shown to mate with equal frequency compared with males of the parental strain. Hence, the presence of the transgene used to produce the strains tested did not incur a fitness cost to the colonies of laboratory-reared C. hominivorax. PMID- 15279309 TI - Patterns of variation within and between Greek populations of Ceratitis capitata suggest extensive gene flow and latitudinal clines. AB - Four natural Greek populations of Mediterranean fruit fly, Ceratitis capitata (Wiedemann), was studied for genetic variability at 25 enzyme loci. The comparison of polymorphism within and between populations shows two loci with high between-population heterozygosity (HT) and very high fixation index (F(ST)) values, suggesting the presence of balancing selection. The gradual decline of common allele frequency of the polymorphic loci tested indicated that latitudinal clines are present in Greece. Indirect estimates of gene flow based both on Wright's method (Nm*) and on the Slatkin's method (Nm*), which depends on the frequencies of rare alleles found in only one population, revealed a substantial amount of gene flow (Nm = 3.493 and Nm* = 3.197). These estimates of gene flow may well explain why the "introduced" Greek populations of C. capitata, in spite of their low genetic variability, display the same polymorphic loci. Gene flow in combination with natural selection and genetic drift may have played an important role to genetic differentiation in this species in Greece. PMID- 15279310 TI - Psychological sequelae of remote exposure to the September 11th terrorist attacks in Canadians with and without panic. AB - The purpose of the present investigation was to examine the psychological impact of remote exposure to the events and aftermath of the terrorist attacks in the USA on September 11th, 2001, and to assess what differences, if any, exist between individuals classified with probable panic disorder and those without. Telephone interviews were conducted with 122 residents of the capital city of the Canadian prairie province of Saskatchewan in spring 2002 in order to gather information regarding current mood, fears and avoidance behaviours as well as current post-traumatic stress disorder symptoms specific to September 11th. Consistent with previous findings and despite the remote nature of exposure, results indicated that the psychological well-being and behaviour of participants with probable panic disorder was more adversely affected by the events and aftermath of September 11th than those without panic disorder. These results suggest that remote viewing of traumatic events can have a significant and lingering impact on psychological well-being and behaviour and that these effects are more pronounced in those with panic disorder. Implications of the findings and future research directions are discussed. PMID- 15279311 TI - Effects of September 11 on patients with obsessive compulsive disorder. AB - Effects of the September 11, 2001 terrorist attacks in the USA were investigated in 25 patients with obsessive compulsive disorder and 27 normal controls 4-6 months after the attacks. Participants completed a 15-item questionnaire to retrospectively assess changes in mood, cognition, behavior and somatic complaints since September 11, 2001. Overall, both patients with obsessive compulsive disorder and normal controls reported minor changes in mood, behavior and somatic complaints. However, normal controls reported severe to extreme initial impact, slightly more cognitive symptoms (uncertainty about the future, intrusive recollections and greater desire to be with loved ones) and a slightly greater degree of overall impact on emotion and behavior at 1, 2 and 3 months after September 11 than did patients with obsessive compulsive disorder. Results support previous research that has found a relatively minor lasting impact of September 11 on both clinical and normal populations. Differences in cognition and coping mechanisms between normal controls and patients with obsessive compulsive disorder are proposed. PMID- 15279312 TI - Threat appraisals, distress and the development of positive life changes after September 11th in a Canadian sample. AB - Several surveys have reported the negative psychological impact on the general public of the terrorist attacks in the USA of September 11th, 2001. Yet the attacks also led many people to make positive changes in their relationships, values and priorities. A survey of 80 adults in Ottawa, Canada demonstrated that greater perceived threat and greater initial distress reactions significantly predicted the extent to which people reported positive changes in their lives (e.g. closer to family, refocused priorities). Initial distress and greater perceived threat also correlated positively with whether people provided help after the disaster. Follow-up data on 40 of these participants 11 months later revealed significant stability over time for the extent of positive life changes reported, and demonstrated that degree of initial distress and perceived threat continued to correlate positively with life change reports at this later point in time. The data are consistent with the argument that the perception of growth may develop out of one's personal experience of emotional pain. PMID- 15279313 TI - Norwegian version of the automatic thoughts questionnaire: a reliability and validity study. AB - This study investigated the reliability and validity of the Norwegian version of the Automatic Thoughts Questionnaire. Three samples were used: (a) 344 male military recruits; (b) 142 psychiatric outpatients; and (c) 41 healthy controls. The Automatic Thoughts Questionnaire was found to have high internal consistency and satisfactory temporal stability. It correlated positively with the Beck Depression Inventory and was found to discriminate significantly between clinically depressed and non-depressed psychiatric patients and healthy controls. In general the results are satisfactory, suggesting that the Norwegian version of the Automatic Thoughts Questionnaire has adequate reliability and validity properties. The Automatic Thoughts Questionnaire was shown to be a useful measure of frequency of automatic negative thoughts in both clinical and non-clinical populations. PMID- 15279314 TI - Psychometric properties of the Norwegian version of the Dysfunctional Attitude Scale (Form A). AB - The present study reports the reliability and validity of the Norwegian version of the Dysfunctional Attitude Scale in non-clinical and clinical populations. The participants were 344 young male military recruits, 41 healthy controls and 142 psychiatric outpatients. All the participants completed the Dysfunctional Attitude Scale, the Beck Depression Inventory and the Automatic Thoughts Questionnaire. The analysis of the Dysfunctional Attitude Scale revealed a Cronbach's alpha of 0.85, indicating satisfactory reliability. Evidence for the construct validity was obtained by the correlation between the Dysfunctional Attitude Scale and the Beck Depression Inventory (r = 0.47) and the Dysfunctional Attitude Scale and the Automatic Thoughts Questionnaire (r = 0.47). Finally, the Dysfunctional Attitude Scale significantly discriminated between clinically depressed, non-depressed psychiatric patients and healthy controls. The results showed that the Norwegian version of the Dysfunctional Attitude Scale possess satisfactory psychometric properties suggesting that this instrument is appropriate for use as a cognitive measure in a Norwegian cultural context. PMID- 15279315 TI - Deficits in perceived social support associated with generalized social phobia. AB - Social phobia is a common anxiety disorder associated with significant impairment in social and occupational functioning. To date, few studies have examined the relationship between social phobia and perceived social support, a construct with important relationships to physical and mental health. The present study examined data from 2 widely used measures of perceived social support administered to 132 individuals with DSM-IV generalized social phobia. These data were compared with those obtained from a healthy control group and from several clinical and non clinical samples reported in the literature. Persons with generalized social phobia scored significantly lower on both measures of social support compared with all other groups. It is suggested that deficits in perceived social support associated with generalized social phobia may play a role in the development of co-morbid problems and should be explicitly targeted by treatments for social phobia. Low correlations between perceived social support and social anxiety measures suggest that perceived support should be specifically evaluated in this population. PMID- 15279316 TI - Is traumatic amnesia nothing but psychiatric folklore? AB - Some psychotherapists believe that certain experiences are so overwhelmingly traumatic that some victims become incapable of remembering their worst trauma except under special circumstances (e.g. therapy) many years later. Unfortunately, clinicians who endorse this concept of traumatic amnesia often misinterpret the very studies they adduce in support of it. More specifically, they misinterpret other, unrelated memory phenomena as evidence for traumatic amnesia, such as ordinary forgetfulness, psychogenic amnesia, organic amnesia, incomplete encoding of traumatic experiences, non-disclosure of remembered trauma, and simply not thinking about something for a long time. The purpose of this article is to dispel confusions rampant in this literature. PMID- 15279317 TI - Amnesia, folklore and folks: recovered memories in clinical practice. PMID- 15279318 TI - IVF and ICSI reimbursed in Belgium. PMID- 15279319 TI - Vitality of oligozoospermic semen samples is improved by both swim-up and density gradient centrifugation before cryopreservation. AB - PURPOSE: To ascertain whether washing sperm from oligozoospermic and normozoospermic samples before cryopreservation improves post-thaw vitality. METHODS: Normozoospermic (n = 18) and oligozoospermic (n = 16) samples were divided into three aliquots. The first aliquot remained untreated and the second and third aliquots were subjected to the swim-up and discontinuous density gradient sperm washing techniques respectively. Vitality staining was performed, samples mixed with cryopreservation media and frozen. Spermatozoa were thawed, stained, and vitality quantified and expressed as the percentage of live spermatozoa present. RESULTS: Post-thaw vitality in untreated aliquots from normozoospermic samples (24.9% +/- 2.3; mean +/- SEM) was significantly higher (unpaired t-tests; P < 0.01) than untreated oligozoospermic samples (11.9% +/- 2.3). Post-thaw vitality was significantly higher after swim-up in normozoospermic samples (35.6% +/- 2.1; P < 0.001; one-way ANOVA) and oligozoospermic samples (27.7% +/- 1.7; P < 0.01). Density gradient centrifugation significantly improved post-thaw vitality in oligozoospermic (22.4% +/- 1.0; P < 0.01) but not normozoospermic (30.8% +/- 1.8) samples. CONCLUSIONS: Sperm vitality in cryopreserved oligozoospermic samples was improved by both the swim-up and density gradient centrifugation washing techniques prior to freezing. PMID- 15279320 TI - Impact of semen characteristics on the success of intrauterine insemination. AB - PURPOSE: To evaluate the influence of sperm characteristics on the outcome of infertility treatment using intrauterine insemination (IUI). METHODS: Retrospective study of 431 infertility couples who underwent 1007 IUI treatment cycles from June 1999 to October 2002. Sperm parameters before and after preparation for IUI were evaluated and correlated with pregnancy outcome. RESULTS: Clinical pregnancy occurred in 12% of cycles and 28% of patients. Initial sperm motility and processed forward progression were independently associated with pregnancy after IUI. The mean number of cycles per patient was 4.3. Although pregnancy rate per cycle did not differ from cycle to cycle, the cumulative pregnancy rate approached plateau after five cycles. CONCLUSIONS: Sperm motility is an independent factor influencing IUI-related pregnancy. A forward progression score of 3 to 4 in a processed specimen is necessary for IUI success. The number of IUI attempts per patient should be individualized depending upon the needs of patients. PMID- 15279321 TI - Low-level gonosomal mosaicism in women undergoing ICSI cycles. AB - PURPOSE: Females with low-level gonosomal mosaicism (LLGM) scheduled for ICSI were retrospectively analyzed and compared to control groups. The prevalence of this mosaicism as well as its impact for an ICSI treatment were evaluated. METHODS: Routine cytogenetic analysis was done in 891 females scheduled for ICSI treatment. For comparison 294 females with recurrent abortions and 104 women with clinical or cytogenetical affected children, who also had routine chromosome analysis, were acquired. RESULTS: In the ICSI group the incidence of LLGM was significantly lower compared to the group of females with recurrent abortions (3.9% vs. 8.5%, p < 0.01). The incidence was similar between the groups of ICSI females and females with cytogenetical or clinically affected children (5.8%, p = 0.43). Neither anamnestic factors nor outcome parameters were different between females with and without LLGM undergoing ICSI. CONCLUSION: There is no higher incidence of LLGM in females undergoing ICSI. In those who show LLGM, fertility capacity is not reduced. PMID- 15279322 TI - Reproductive performance of couples discordant for hepatitis B and C following IVF treatment. AB - PURPOSE: To examine the reproductive performance of hepatitis B (HBV) and C (HCV) discordant couples following IVF-ET. METHODS: A retrospective cohort study of 25 IVF-ET cycles in HBV and HCV discordant couples was performed. Thirteen patients in the study cohort were discordant for HBV (10 males and 3 females), and 12 (9 males and 3 females) for HCV. Twenty-seven consecutive age matched patients comprised the control group. All patients underwent controlled ovarian hyperstimulation using the long downregulation protocol followed by IVF or ICSI. RESULTS: Patients in the three groups (HBV, HCV, and controls) had similar ages, and day 3 FSH concentrations. Despite comparable response to COH, and similar fertilization, and cleavage rates in the three groups, couples discordant for HBV or HCV had significantly poorer implantation and pregnancy rates (7.7%, 0% respectively) compared with controls (41%). CONCLUSIONS: Despite comparable response to COH, HBV and HCV positive discordant couples, have significantly lower implantation and pregnancy rates compared with age-matched controls. PMID- 15279323 TI - Feasibility of human telomerase reverse transcriptase mRNA expression in individual blastomeres as an indicator of early embryo development. AB - PURPOSE: The study was undertaken to test whether human telomerase reverse transcriptase (hTERT) transcripts in an individual blastomere could be used as an indicator for embryo development. METHODS: Group A consisted of day 3 normal cleaving embryos at 4- to 8-cell stage, which were surplus and not allocated for uterine transfer. Group B consisted of arrested or fragmented embryos at the same stage, which were considered to be compromised. After blastomere dissociation, RNA purification, reverse transcription, and hTERT PCR amplification, the amplified product was separated by 3% gel electrophoresis. RESULTS: Eighty-six (90.5%) of the 95 intact blastomeres in group A and 78 (70.9%) of the 110 blastomeres in group B demonstrated hTERT mRNA expression. The difference was statistically significant (P < 0.05, chi-square). Eight (38.1%) of the 21 embryos in group A and 22 (62.9%) of the 35 embryos in group B had at least one blastomere that did not express hTERT mRNA under this procedure. The difference was not significant (P > 0.05, chi-square). CONCLUSIONS: General hTERT mRNA transcripts can be detected in most of the individual blastomeres but cannot be used as an indicator for early embryo development. Further investigations are necessary to elucidate its clinical application. PMID- 15279324 TI - The effect of a two-hour, room temperature incubation of human spermatozoa in TEST-yolk buffer on the rate of fertilization in vitro. AB - PURPOSE: To reassess the use of TEST-yolk buffer (TYB) in an in vitro fertilization (IVF) program by comparing fertilization rates achieved in a glucose-free cleavage medium by the standard IVF preparation of sperm versus a 2 h, room temperature incubation of sperm in TYB. METHODS: Oocytes collected for IVF were randomly split into two groups and inseminated with either TYB-treated sperm or IVF-prepared sperm. SETTING: Stanford Reproductive Endocrinology and Infertility Center. PATIENTS: Fifty couples undergoing IVF with at least 10 mature oocytes. MAIN OUTCOME MEASURES: Fertilization rates in vitro. RESULTS: Fertilization rates were significantly higher (p = 0.015) with TYB treatment. The average 2PN fertilization rate was 49.6% (188/379) for the IVF group and 57.4% (221/385) in the IVF with TYB group. CONCLUSIONS: A 2-h, room temperature incubation of sperm in TYB produces significantly higher 2PN fertilization rates as compared to standard IVF preparation of sperm in a current generation cleavage medium. PMID- 15279326 TI - Bilateral ovarian pregnancy after in vitro fertilization and embryo transfer in a patient with tubal factor infertility. AB - Primary ovarian pregnancy is very rare event after natural pregnancy or assisted reproductive technology (ART) procedures. Although there are a few reports about unilateral ovarian pregnancy after in vitro fertilization and embryo transfer (IVF-ET), there has been no report about bilateral ovarian pregnancy. Moreover, it is difficult to diagnose an ovarian pregnancy following in vitro fertilization and embryo transfer because of enlarged ovary, fluid collection in pelvic cavity, and its low incidence. We present a case of a patient who underwent IVF-ET due to tubal factor infertility, but the patient developed bilateral ovarian pregnancy and was performed both ovarian wedge resection through laparotomy. PMID- 15279325 TI - Tissue perfusion essential for spermatogenesis and outcome of testicular sperm extraction (TESE) for assisted reproduction. AB - PURPOSE: In order to determine if there are areas of major and minor perfusion in a single testicle and if the quality of sperm is correlated with quantity of perfusion we collected testicle tissue for TESE in accordance to the local testicle tissue perfusion. METHODS: A patient undergoing TESE underwent testicular perfusion mapping using contrast enhanced ultrasound. The exposed tissue was scanned with a Laser Doppler scanner and perfusion rates were determined measuring tissue perfusion units (TPUs). Tissue was biopsied and sperm were selected and prepared for assisted reproduction. RESULTS: The total amount of isolated sperm correlated highly with the intensity of tissue perfusion showing high number of sperm in areas with high TPUs. CONCLUSIONS: This is the first demonstration that sperm quality and quantity is depending on tissue perfusion within the testicle. To further improve infertility treatment we propose that random biopsies could be replaced by perfusion-dependent collection of testicular tissue. PMID- 15279327 TI - Private self-consciousness factors: relationships with need for cognition, locus of control, and obsessive thinking in Iran and the United States. AB - The authors measured Internal State Awareness (ISA) and Self-Reflectiveness (SR) factors from the Private Self-Consciousness Scale in Iranian (N = 325) and U.S. (N = 401) university students. In both societies, positive correlations with Need for Cognition and Internal Control and negative correlations with external control and obsessive thinking confirmed ISA as an adaptive form of self consciousness. In partial correlations in which the authors controlled for ISA, SR was associated cross-culturally with greater Obsessive Thinking. This outcome conformed with the authors' expectations that SR would have negative mental health implications, but other data revealed complexities in the SR association with adjustment. Differences between samples failed to yield any simple support for F. Fukuyama's (1992) suggestion that Iranians might be more "alienated" (pp. 236-237) in their psychological functioning. The present study most importantly offered cross-cultural evidence in favor of the claim that better measures of an introspective self-awareness need to be developed. PMID- 15279328 TI - Intergroup perception as a compromise between in-group bias and fair-mindedness. AB - Previously, perceived competence of and attraction toward targets categorized by race showed in-group bias and no bias, respectively. Consequently, previous investigators regarded intergroup perception as a compromise between the norms of in-group bias and fair-mindedness. An alternative hypothesis for such findings is that attraction is not as relevant a dimension for intergroup discrimination as is competence. To test contrasting predictions of these hypotheses, the present authors asked participants from the majority and minority groups in Singapore (ns = 320) to evaluate either competence of or attraction toward one of the five targets. Consistent with the hypothesis that intergroup perception is a compromise, both dimensions yielded a uniform but weak in-group bias. The participants' equating of the in-group with one out-group further illustrated fair-mindedness. The authors discussed implications of the findings. PMID- 15279329 TI - Work-family conflict, work- and family-role salience, and women's well-being. AB - The author considered both the direct effect and the moderator effect of role salience in the stress-strain relationship. In contrast to previous studies that have examined the effects of salience on well-being within specific social roles, the present study focused on the work-family interface. From a sample of 147 employed English women with children, the present results of the regression analyses showed that both effects are possible, depending on the outcome measures used. The author observed a direct effect of role salience in the prediction of job satisfaction; work salience was positively related to job satisfaction, over and above the main-effect terms of work-interfering-with-family (WIF) conflict and family-interfering-with-work (FIW) conflict. In contrast, the author found a moderator effect of role salience and conflict for symptoms of psychological distress. However, contrary to predictions, the author found that work salience exacerbated the negative impact of WIF conflict, rather than FIW conflict, on well-being. The author discussed these results in relation to the literature on work-family conflict, role salience, and the issue of stress-strain specificity. PMID- 15279330 TI - World assumptions and combat-related posttraumatic stress disorder. AB - The authors examined the association between (a) personal world assumptions and (b) combat stress reactions (CSRs), posttraumatic stress disorder (PTSD), and PTSD's course among three groups of Israeli veterans: 109 veterans who suffered from CSR on the battlefield, 98 decorated veterans, and 189 control participants. Participants completed standardized questionnaires that measured PTSD and world assumption. Both CSR and chronic PTSD were associated with lower levels of self worth and beliefs about the benevolence of people. In addition, the authors found a linear association between self-worth perceptions and levels of mental status. The authors examined the results of the study considering the extraordinary characteristics and meaning of war. PMID- 15279331 TI - Highly dominating, highly authoritarian personalities. AB - The author considered the small part of the population whose members score highly on both the Social Dominance Orientation scale and the Right-Wing Authoritarianism scale. Studies of these High SDO-High RWAs, culled from samples of nearly 4000 Canadian university students and over 2600 of their parents and reported in the present article, reveal that these dominating authoritarians are among the most prejudiced persons in society. Furthermore, they seem to combine the worst elements of each kind of personality, being power-hungry, unsupportive of equality, manipulative, and amoral, as social dominators are in general, while also being religiously ethnocentric and dogmatic, as right-wing authoritarians tend to be. The author suggested that, although they are small in number, such persons can have considerable impact on society because they are well-positioned to become the leaders of prejudiced right-wing political movements. PMID- 15279332 TI - Severity of measles: a study at the Queen Sirikit National Institute of Child Health. AB - INTRODUCTION: Thousands of measles cases are reported annually in Thailand even though measles vaccine has been introduced in the expanded program of immunization for every 9-month-old infant for nearly 20 years. Severe cases are admitted to the hospital, usually with complications, some cases lead to death. OBJECTIVES: To study the clinical presentations of severe cases of measles and its complications and find the correlations of severity of pneumonia with age, nutritional status and history of vaccination. MATERIAL AND METHOD: The hospital charts of measles patients admitted to the Queen Sirikit National Institute of Child Health (QSNICH) during 1998-2002 were retrospectively reviewed. Demographic data, history including history of measles vaccination, physical examinations, laboratory investigations, treatment and hospital course which were relevant were recorded. Paired t-test and Pearson's correlation were used for data analysis. RESULTS: There were 156 cases of measles admitted to the QSNICH. There were 95 boys and 61 girls and the male to female ratio was 1.56:1. The age range was 2 months to 14.8 years, median = 1.5 years, mode 8 months. Fifty-nine percent of the cases were under 2 years of age; 40% under one year and 23.9% were under 9 months. About 44% of the cases had one dose of previous measles vaccination, no history of measles vaccination in 91.4% of cases whose age was under 1 year in contrast to 80% of cases over 5 years that had a history of measles vaccination. Sixty-six percent of the cases had normal nutritional status while 12.4%, 4.8% and 2.1% had mild, moderate and severe protein calorie malnutrition. Fourteen cases (9%) had underlying diseases. At least 3 of the classical signs and symptoms of measles (rash, cough and coryza) were found in 92.3% of the cases. The mean duration of fever at the time of admission was 5.3 days. The common complications in admitted measles cases were pneumonia (62.2%) and diarrhea (38.1%). The likely causes of pneumonitis were measles viruses (52.6%) and bacteria (47.4%). There was one dead case with severe pneumonia, with ARDS and respiratory failure. Young infants had a higher incidence of diarrhea with dehydration (p = 0.000) but severity of pneumonia was not different from older children (p = 0.512). The severity of pneumonia was not correlated with the age (r = 0.087), nutritional status (r = 0122) or the history of receiving measles vaccine (r = 0.116). CONCLUSION: Measles is one of the important diseases of in patients admitted to the QSNICH, because of the severity of the diseases due to pneumonia and diarrhea. One severe case died because of severe pneumonia that lead to ARDS and respiratory failure. Young infants had a higher incidence of diarrhea and dehydration, while there was no correlation between severe pneumonia with age, nutritional status and history of vaccination. PMID- 15279333 TI - Is calculated LDL-C by using the new modified Friedewald equation better than the standard Friedewald equation? AB - The patients who have CHD or CHD risk equivalents should have LDL-C level less than 100 mg/dL because of the great reduction of risk for major coronary events. Direct measurement of LDL-C is the most accurate but is expensive. But with the practical use of the Friedewald equation for calculating LDL-C, the authors noticed that the accuracy declined with triglyceride level being higher than 300 mg/dL. The authors determined the correlation of direct measurement of LDL-C with calculation LDL-C from the Friedewald equation and postulated the new modified Friedewald equation for calculating LDL-C by using 1/6 triglyceride to minus. From a total of 1079 fasting serum samples analysis, and determining the correlation of LDL-C from the direct measurement (dm LDL) while calculating LDL-C from equations of the standard Friedewald (sf LDL), and the new modified Friedewald (mf LDL), by using 1/6 triglyceride to minus instead of 1/5 triglyceride, if triglyceride was over 200 mg/dL. The authors found an excellent correlation within 0 +/- 10% difference of dm LDL and sfLDL if triglyceride was less than 200 mg/dL, but sf LDL is less accurate when the triglyceride level is high, and mf LDL has better correlation with dm LDL within 0 +/- 10% that sf LDL vs mf LDL, 72.3% vs 91.6% (p = 0.0001), and 58.3% vs 83.3%, (p = 0.01) when the triglyceride level is 200-299, and 300-399 mg/dL respectively. It is shown that sf LDL has more underestimation than mf LDL when compared with dm LDL (more than 10 mg/dL) as 26.9% vs 2.5% (p < 0.0001) and 41.6% vs 5.6% (p = 0.0003 ) with triglyceride of 200-299, and 300-399 mg/dL respectively, although mf LDL showed overestimation of more than 10 mg/dL difference with dm LDL as sf LDL vs mf LDL of 0.8% vs 5.8% (p = 0.03), and 0.0% vs 11.1% (p=0.03) if the triglyceride is in the range of 200-299 and 300-399 mg/dL respectively, even with a triglyceride level of 400-499 mg/dL, mfLDL still has good correlation with dm LDL up to 75.0%. The authors conclude that the standard Friedewald equation is excellent for LDL calculation if triglyceride is less than 200 mg/dL, but the accuracy is declined when triglyceride is over 200 mg/dL, the authors offer a new modified Friedewald equation to calculate LDL-C if triglyceride is in the range of 200-499 mg/dL which has a better correlation with direct measured LDL-C. However this new modified Friedewald equation needs to be testified again especially with dyslipidemic patient sera. PMID- 15279334 TI - Changes in hematologic markers in patients with mitral stenosis after successful percutaneous balloon mitral valvuloplasty. AB - Systemic embolism is a major complication of mitral stenosis which is usually related to a presence of left atrial thrombus. Percutaneous balloon mitral valvuloplasty (PBMV) was previously reported to reduce the incidence of this complication. However, the mechanisms of this beneficial procedure was under investigated. The aim of this study was to investigate the changes in coagulation activity, platelet activity and endocardial function in 29 patients with mitral stenosis after successful PBMV. All subjects had good left ventricular systolic function and 48.3% had atrial fibrillation. There was a significant reduction in thrombin-antithrombin complex (TAT) after a successful procedure and the level of thrombomodulin was also significantly higher one month after successful procedure. However, the level of platelet factor 4 (PF4) and beta-thromboglobulin (beta-TG) were increased after this procedure but not achieved the statistical significance. In conclusion, successful PBMV can reduce the prethrombotic state in patients with mitral stenosis. In addition, it may improve endocardial function of the left atrium in those without atrial fibrillation. PMID- 15279335 TI - Role of bronchial washing in the diagnosis of endoscopically visible lung cancer. AB - OBJECTIVE: To evaluate the usefulness of bronchial washings in addition to endobronchial biopsies and/or bronchial brushings for the pathological diagnosis of endoscopically visible lung cancer. MATERIAL AND METHOD: A retrospective study of patients diagnosed as lung cancer by bronchoscopy between January 1995 and December 1998. Patients were included in the study if they had 1) endoscopically visible tumors (exophytic mass or irregular mucosa) and 2) bronchial washings (BWs) performed together with either endobronchial biopsies (EBBs) or bronchial brushings (BRs). Patients were classified into 3 groups according to the result of the histocytology as follows: A) positive in both BWs and EBBs/BRs, B) positive in only EBBs/BRs and C) positive in only BWs. A number of patients in each group were analyzed to see the benefit of BWs as an add-on diagnostic tool. The authors also evaluated the benefits of BWs in the subgroup of patients who had necrotic and bleeding tumor Statistical analysis of the data was performed by using the likelihood-ratio chi-square test. RESULTS: Two hundred and twenty-two patients were included in the present study. The number of patients in group A, B and C was 108, 108, and 6, respectively. Therefore, BW was the only diagnostic procedure in 6 patients (2.7%). Those 6 patients all had incurable non-small cell lung cancer The likelihood of a positive BWs in an exophytic mass was no different from irregular mucosa. The likelihood of a positive BWs in a tumor with necrosis was higher than in a tumor without necrosis. In contrast, tumors with active bleeding had a lower likelihood of positive BWs when compared with those without bleeding. The likelihood ratio showed no statistical significance in any of the groups. CONCLUSION: The addition of BWs to either EBBs or BRs is beneficial, but it may not be cost-effective. This procedure may be useful in patients with an endoscopically visible necrotic tumor In contrast, the bronchoscopic finding of a bleeding tumor may be a negative predictor. This procedure may be a suitable approach when performed only in selected cases, such as necrotic tumor or negative initial EBBs/BRs. PMID- 15279336 TI - Prevalence of lipodystrophy in Thai-HIV infected patients. AB - To determine the prevalence and clinical characteristics of lipodystrophy in HIV infected Thai patients, a cross-sectional study was performed on 278 HIV-infected patients at Bamrasnaradura Infectious Disease Institute. Laboratory data related to lipid and glucose metabolism were obtained from both patients who self reported fat maldistribution or diagnosed by a physician. The history of antiretroviral treatment and HIV infection were recorded. Prevalence of lipodystrophy found in the present study was 17%. Lipodystrophy was reported mostly on the face, buttock, legs, arms, and abdomen respectively. Two-thirds of these patients had mixed syndromes of fat accumulation and fat wasting and the others had only fat wasting. Ninety-three percent of lipodystrophic patients had at least 1 abnormality in either lipid or glucose metabolism. Eighty-eight percent had dyslipidemia, 21% had impaired glucose tolerance, 30% had insulin resistance and 27% had diabetes mellitus. Lipodystrophic patients have a high rate of lipid and glucose metabolism abnormalities which are the major risk factors for cardiovascular events. PMID- 15279337 TI - Primary congenital hypothyroidism: clinical characteristics and etiological study. AB - Forty-eight children with primary congenital hypothyroidism, who attended Chiang Mai University Hospital, during 1977-2000, were reviewed. The female to male ratio was 2:1. The age at diagnosis ranged from 1 month to 12 years 4 months, with 27% of the cases diagnosed within the first three months of life, 37.5% within the first year, and 62.5% after one year of age. Constipation, delayed development and growth, feeding problems, prolonged neonatal jaundice and goiter were more common. Prolonged neonatal jaundice was found in every case diagnosed within the first three months. The other common signs were dry or mottled skin, abdominal distension, macroglossia, short stature, puffy face and umbilical hernia. Kocher-Debre-Semelaigne syndrome comprised 18.7% of cases with a 2:1 female to male ratio, and it was found in various forms of hypothyroidism. Thyroid scintigrams were done in 47 patients. Thyroid dysgenesis was the most common etiology (80.9%), which consisted of 40.4% athyreosis, 4.3% hypoplasia, and 36.2% thyroid ectopy. Thyroid dyshormonogenesis accounted for 18.9%, in which only 4 of 9 presented with goiter. Two-thirds of these patients showed a positive result to the perchlorate discharge test, indicating an organification defect. A 11 patients had elevated serum TSH level greater than 50 mU/L. The serum T4 level below 2 microg/dL was observed in 17 of 19 patients with athyreosis, 11 of 1 7 with thyroid ectopy, and 6 of 9 with thyroid dyshormonogenesis. These findings including retarded bone age were unable to differentiate among different groups of hypothyroidism. PMID- 15279338 TI - Meniscal lesions in the anterior cruciate insufficient knee: the accuracy of clinical evaluation. AB - OBJECTIVE: The purpose of this study was to find out the accuracy of certain symptoms and examination findings that are used to diagnose meniscal injury associated with a torn anterior cruciate ligament. STUDY DESIGN: Cross-sectional study. MATERIAL AND METHOD: The authors studied one hundred consecutive patients with anterior cruciate ligament insufficiency who were scheduled for surgery. During preoperative admission, one of the authors (KT) examined the patients and recorded the demographic data, duration of symptoms, and the clinical findings including Ballottement sign, joint line tenderness, Childress' sign, Merke's sign, Steinmann I sign, McMurray test, and Apley test. All patients underwent arthroscopically assisted anterior cruciate reconstruction by the senior author (PC). Specific meniscal procedures were performed according to the surgeon's preference at the time of surgery. Predictive results of preoperative examination tests for meniscal tears were compared with the findings at surgery and analyzed using arthroscopic findings as the gold standard. RESULTS: There were one hundred patients included in the present study. Out of 100 patients, 75% had meniscal tears and 6% had both meniscal and cartilage lesions. The most sensitive test was Childress' sign (68%), which also had the highest accuracy (66%). The most specific tests were Steinmann I sign and Apley test (100%). CONCLUSION: Childress' sign was more accurate than other tests for detecting meniscal lesions in anterior cruciate insufficient knees. Steinmann I sign and Apley test had the highest specificity. PMID- 15279339 TI - The role of SMAS flap in preventing Frey's syndrome following standard superficial parotidectomy. AB - Between 1992 and 2002, 46 patients who underwent standard superficial parotidectomy with a superficial muscoloaponeurotic system (SMAS) preservation technique were included in a retrospective study. Twenty-six patients were evaluated by questionnaire for subjective symptoms of gustatory sweating and flushing as well as satisfaction with the aesthetic appearance of their cheek. Six of twenty-six patients (23.1%) complained of symptoms of Frey's syndrome. Seven of twenty-six patients (26.9%) demonstrated a positive Minor's starch iodine test. By this technique the incidence of Frey's syndrome is substantially reduced from 48% by subjective review and 72% by objective measurement reported in the previous study by the same group of surgeons without using the SMAS preservation technique. This study supports the role of the SMAS flap in preventing Frey's syndrome following standard superficial parotidectomy. PMID- 15279340 TI - Maternal satisfaction to epidural and spinal anesthesia for cesarean section. AB - BACKGROUND AND OBJECTIVE: The differences between epidural (EA) and spinal (SA) anesthesia that can affect maternal satisfaction are the procedures, quality of anesthesia and postoperative events. Dominantly, postoperative events such as postdural puncture headache, pruritus and nausea or vomiting after spinal anesthesia are claimed to be its disadvantages. However, maternal satisfactory perception to theses two techniques has not been revealed. The authors' purpose was to compare maternal satisfaction regarding the techniques and their outcomes between EA and SA by the developed valid and reliable tool. MATERIAL AND METHOD: Patients were randomly classified into two groups: epidural (Group E, n=56) and spinal (Group S, n = 58). Epidural and spinal anesthesia were administered with bupivacaine, 20 mL 0.5% with 1:200,000 epinephrine combined with two doses of 5 mg morphine and hyperbaric bupivacaine 2.2-2.4 mL 0.5% combined with 0.2 mg morphine respectively. Guidelines for treatment of intraoperative and postoperative events, which might be the confounding factors, were set up. Maternal satisfaction was evaluated by the 11-item, qualified, self-administered questionnaire comprised of 4 common factors. The score of 0-10 Visual analog scale was used to access the degree of satisfaction. Trained personnel performed data collections in the post-anesthesia care unit and ward. The means of the factor and total satisfaction scores were compared between the two groups by Mann Whitney U test. A p-value < 0.05 considered significant. RESULTS: There was no statistical difference in the factor scores between the two groups. The total satisfactory score was 89.48 +/- 9.31 and 90.03 +/- 11.26 in Group E and S respectively. No statistical difference of the total satisfaction score was detected. CONCLUSION: There was no difference in maternal satisfaction regarding to the techniques and the outcomes between EA and SA. PMID- 15279341 TI - Endometrial abnormalities in postmenopausal breast cancer patients. AB - OBJECTIVE: To determine the prevalence of endometrial thickening and pelvic pathologies in postmenopausal breast cancer patients. MATERIAL AND METHOD: A total of 66 postmenopausal breast cancer patients receiving treatment at Srinagarind Hospital from 1 July 1999 to 31 August 2000 were included in the study. Patients who had been treated with hormones such as tamoxifen or progestins were not eligible for this study. Thorough history taking and physical examination as well as transvaginal ultrasonography were conducted in all study patients. Fractional curettage was carried out and the specimens obtained were sent for pathological examination in all patients whose endometrial thickness was found to be greater than 5 mm on the ultrasound scan. RESULTS: Among the 66 patients included in this study, the mean age was 54.97 years. The majority of patients (75.76%) had stage II disease. The mean +/- SD of endometrial thickness found in this study was 3.55 +/- 1.72 mm. The prevalence of thickened endometrium (defined as ET > 5 mm from TVS) was 10.60% (7 from 66 cases). Other pelvic pathologies detected by ultrasonography were myoma uteri (4.55%) and ovarian mass (1.52%). Among the seven patients whose endometrial thickness was found to be greater than 5 mm, results of curettage specimens revealed inadequate tissue obtained (42.85%), atrophic endometrium (28.57%), active endometrial gland (14.29%), and scanty stromal cell (14.29%). CONCLUSION: The prevalence of thickened endometrium in postmenopausal breast cancer patients found in this study was interestingly high. The pathological results of such cases, however, turned out to be negative for neoplastic changes in all cases. Further study, thus, is needed before precise recommendation could be made regarding the value of TVS screening in breast cancer patients not taking tamoxifen. PMID- 15279342 TI - Randomized, double-blind clinical trial of a lactose-free and a lactose containing formula in dietary management of acute childhood diarrhea. AB - BACKGROUND: Refeeding of artificially fed infants with lactose-containing formula after oral rehydration therapy in the treatment of acute diarrhea was concluded to be indifferent to non-lactose formula by a meta-analysis. In Thai as well as Asian infants and children with low lactase level from genetically determinant and with rotavirus infection, lactose malabsorption is most likely to occur and cause delayed recovery. The aim of this study was to compare the effect of a lactose-free and a lactose-containing formula in dietary management of acute childhood diarrhea. PATIENTS AND METHOD: A randomized, double-blind clinical trial of 80 male children, formula-fed, aged 3 to 24 months, admitted with acute watery diarrhea and mild or moderate dehydration, was carried out. All children received oral rehydration therapy for the first 4 hours. After appropriate rehydration, they were fed either a lactose-free formula (Dumex Lactose-Free Formula; treatment group, n = 40) or a lactose-containing formula (Dumex Infant Formula; control group, n = 40) in adjunction with oral rehydration solution. In addition, the infants were fed rice gruel as tolerated. Comparisons of duration of diarrhea, weight gain, vomiting, biochemical changes, stool frequency and weight and unscheduled intravenous fluid were made. RESULTS: Three children (2 treatment, and 1 control) dropped out from the study. The total number of unscheduled intravenous infusions were 6 of 80 children (7.5%), including 2 (5.0%) in the treatment group and 4 (10.0%) in the control group. Three children in the control group did not resolve from diarrhea within 7 days of treatment. Rotavirus was identified in approximately 50% of the children in each group. Using survival analysis, the median duration of diarrhea was significantly shortened by 20.5 hours in the treatment group compared to the control group (77.0 hours in the treatment group vs 97.5 hours in the control group; P = 0.002). Significantly decrease in stool frequency and increase in percent weight gain were seen in the treatment group at 24 hours. Moderate acidosis cleared up to near normal at 24 hours in the treatment group but acidosis persisted in the control group. In the rotavirus diarrhea subgroup, moderate acidosis turned to be mild in treatment group, but acidosis was unchanged with increased plasma chloride level in the control at 24 hours thus suggesting that the children in the control group might have lactose malabsorption and osmotic diarrhea. Duration of rotavirus diarrhea was shortened 23.6 hours in treatment group compared to the control (P = 0.0034). CONCLUSIONS: In this study, lactose-free formula was shown to be effective in the dietary management of acute childhood diarrhea. Duration of diarrhea was shortened, weight gain was better, and stool frequency was less when compared to lactose-containing formula. Moderate acidosis cleared up spontaneously at 24 hours. Unscheduled IV could be decreased by 50%. Children receiving lactose-free formula tolerated it well. Data of subgroup analysis of rotavirus diarrhea revealed lactose-free formula scored higher than the control group for all parameters studied. PMID- 15279343 TI - Treatment of pain after spinal surgery in the recovery room by single dose lornoxicam: a randomized, double blind, placebo-controlled trial. AB - BACKGROUND: Lornoxicam has been used in microsurgical lumbar discectomy. However, there is no data about controlling pain after open discectomy or laminectomy. OBJECTIVE: To compare the efficacy of a single dose of 16 mg of lornoxicam for the treatment of pain after disectomy or laminectomy with placebo in the PACU. STUDY DESIGN: Randomized, double blind, placebo-controlled trial. MATERIAL AND METHOD: Fifty-six patients who underwent discectomy or laminectomy were randomly allocated to receive 16 mg lornoxicam (Group L), or placebo (Group P) at the beginning of wound closure. Pain scores at rest (using a verbal numeric rating scale: VNRS 0-10), time to first analgesia requirement, morphine consumption during the first 2 hr after surgery and adverse effects were all recorded. The outcomes were assessed on admission to the PACU (T0), then at 1 (T1) and 2 (T2) hr after surgery. RESULTS: Baseline data were comparable between the two groups. The proportion of patients with VNRS > 5 at T0 in both groups were not significantly different (44.4% in group P vs 50.0% in group L, CI of difference: 32.4%, 21.3%, p = 0.68). The mean VNRS scores, at T0 and T1 were > 5 and at T2 was < 5 in both groups. There was no difference between the two groups. The morphine consumption in both groups was not different (9.0 mg vs 9.3 mg) as well as the time to first analgesia requirement (35 min vs 40 min). Patients in the two groups had no significant difference in the symptoms or degree of nausea/vomiting. The number of patients with excessive sedation and the proportion of patients needing oxygen during transportation to the ward were not different. CONCLUSION: Lornoxicam 16 mg given intravenously before wound closure provides inadequate pain relief immediately after disectomy or laminectomy in the PACU. However, adequate pain relief was demonstrated at 2 hr after surgery, which was similar to the placebo. PMID- 15279344 TI - Propofol-based fast-track for ambulatory surgery. AB - The purpose of this study was to provide data of propofol-based total intravenous anesthesia (TIVA) for ambulatory surgery in developing a fast-track technique. One hundred and forty-two patients scheduled for elective surgery were studied: mean (SD) age 42.21 (16.23) years, male to female 72:70, mean (SD) body weight 60.75 (11.67) kg and American Society of Anesthesiologists (ASA) physical status I/II/III 66/38/38. Mean (SD) thiopental induction 225 (55.69) mg was maintained with mean (SD) propofol 199.64 (86.26) mg for mean (SD) anesthetic time 29.02 (11.21) minutes. Various narcotics were used: fentanyl 73.48 +/- 24.38 microg for 123 cases, morphine 3.27 +/- 1.10 mg for 10 cases, remifentanil 492 +/- 105.26 microg for 7 cases and pethidine 23.33 +/- 2.88 mg for 2 cases. Midazolam was given 2.70 +/- 1.05 mg. Patients were positioned in supine, lithotomy or lateral decubitus. One-fourth were PS III with a diagnosis of chronic renal failure and renal transplants coming for incision and drainage of perianal abscess. The mean (SD) wake-up time was 36.02 (17.69) seconds. Only one case (chronic renal failure) had severe hypotension after induction. Anesthetic agents and ideas of fast-track anesthesia were discussed. PMID- 15279345 TI - Perianal blockage with 0.5% bupivacaine for postoperative pain relief in hemorrhoidectomy. AB - Hemorrhoidectomy can be done in many positions under many anesthetic techniques as an ambulatory surgery. Post-procedural pain is frequently severe enough to delay home discharge. A combination between preincisional local anesthetics and general anesthesia looks attractive in terms of preemptive analgesia and starting time of surgery. The study aimed to compare anesthetic time, pain-free period and pain relief in patients with and without 0.5% plain bupivacaine infiltration after mask inhalation, total intravenous anesthesia or endotracheal tube general anesthesia. MATERIAL AND METHOD: 142 patients were randomized into control (C) and study (S) groups with n = 70 and 72 respectively. Patient characteristics in both groups were: age 40.45 +/- 13.03 VS 37.48 +/- 13.63 years old, BW 59.77 +/- 11.19 VS 58.80 +/- 9.76 kg, male:female 31/39 VS 43/29, PS 1/2/3/E = 48/19/1/2 VS 53/15/3/1 for C and S respectively. All underwent surgery in lithotomy under ET/TIVA/mask: 53/13/4 VS 22/27/23 and anesthetic time was 49.02 +/- 18.04 VS 33.33 +/- 10.31 min (p < 0.05). RESULTS: Pain-free periods in C and S were 204.44 +/- 878.07 and 540 +/- 298.03 min with median times of 57.5 (n = 67) VS 560 (n = 58) min. Pain severity in S was mainly none or mild degree while in C it was moderate or severe, apparently when analysed in subgroups of ET and TIVA. Analgesic requirements were statistically more in group C. CONCLUSION: Better postoperative pain relief could be accomplished by preincisional 0.5% plain bupivacaine infiltration after general anesthesia. The technique helped relax anal muscles for surgical ease and avoided patient discomfort in case of a prolonged procedure. Preemptive analgesia and key pain management were discussed. PMID- 15279346 TI - Microsurgical free flap and replantation without antithrombotic agents. AB - Microsurgical reconstructions for free flap transfer, digits and limb replantation have been highly successful applications in the past decades. Antithrombotic prophylactic agents, such as low-molecular-weight dextran, aspirin and heparin have been routinely used for the prevention of microvascular thrombosis. Even though these agents are efficacious in microsurgery, some systemic morbidity is still reported. Forty cases of microsurgical reconstruction over the last five years are reported. They include 22 cases of free flap transfer and 18 cases of replantation (19 fingers, 2 toes and one hand). The surgery was performed by the same group of plastic surgeons and no antithrombotic agent was given intraoperatively or during the post operative period Results show one partial flap loss, two replantation losses due to severe crush injuries of the digits and one toe replantation failure in a two year old. The failure of toe replantation was due to surgical technique and poor post operative immobilization. The result shows that successful microsurgical reconstruction depends on many factors. One of the most important factors is microsurgical technique. Use of antithrombotic agents alone does not appear to play a significant role in the patency of microvascular structures. PMID- 15279347 TI - The combined oral contraceptive pill versus bromocriptine to suppress lactation in puerperium: a randomized double blind study. AB - A randomised double blind comparative study of 230 HIV infected mothers who had a normal delivery at 37-42 weeks' gestation were divided into two groups; 116 combined pill users and 114 bromocriptine users to suppress lactation. There were 33 cases (28.5%) of combined pills users and 29 cases (25.4%) of bromocriptine users who had breast engorgement without statistical difference. All of them had mild breast engorgement without any treatment except one case (0.9%) in the bromocriptine group had severe breast engorgement with puerperal fever and needed an analgesic drug. There were no side effects of the drugs. This study showed that combined pills were beneficial to suppress lactation in HIV infected mothers to prevent postnatal mother-to-child transmission because of low risk and low cost. PMID- 15279348 TI - Association between serum homocysteine, folate and B12 concentration with coronary artery disease in Thai patients. AB - BACKGROUND: Hyperhomocysteinemia, associated with low folate and low B12 levels, is known to be an independent risk factor for atherosclerosis. Only a few available data has been demonstrated in Thai patients. OBJECTIVE: To evaluate serum fasting homocysteine, folate and B12 levels whether to see they are associated with coronary artery disease (CAD). METHOD AND RESULT: Three hundred and one consecutive suspected CAD patients who underwent coronary angiography at the Police General Hospital were studied. The mean age of the patients, 195 males and 106 females, was 63.0 +/- 10.0 year (range 39-85). A total of 218 patients were angiographically demonstrated as having CAD. The mean serum homocysteine level of CAD patients had a non significant higher level than those of 83 non CAD patients: 11.4 +/- 6.2 vs 10.2 +/- 4.2 umol/L, p = 0.06. Means of folate and B12 level in the CAD patients and non CAD patients were 6.6 +/- 4.6 vs 7.0 +/- 4.3 nmol/L, p = 0.49 and 650.9 +/- 415.4 vs 613.3 +/- 443.2 pmol/L, p = 0.56 respectively. No significant correlations were found between homocysteine with folate and B12 levels. Logistic regression analysis showed a significant association between homocysteine and CAD with OR = 1.08 (95%CI, 1.01-1.16), p = 0.03 after being adjusted for age, sex, DM, HT history of hyperlipidemia, smoking, BMI, folate and B12 levels. No significant association between homocysteine level with the number of coronary vessel stenosis, age, BMI, DM, HT smoking and history of hyperlipidemia was observed in the present study. CONCLUSION: Hyperhomocyteinemia, but not folate and B12 levels, may be an independent risk factor for coronary artery disease in Thai patients. PMID- 15279349 TI - Vision screening in schoolchildren: two years results. AB - OBJECTIVE: To assess the prevalence of visual impairment and ocular abnormalities among schoolchildren in Chiang Mai. DESIGN: A community-based survey. SUBJECTS AND METHOD: The vision screening project was conducted from June 2000 to March 2002. Students in grade I in the Chiang Mai municipal area were examined for visual acuity (VA), color vision, ocular alignment, anterior segment and fundus. Subjective refraction was done in students with subnormal vision (VA 20/30 or less). Referral to the hospital for further evaluation and treatment was made for students with strabismus, amblyopia and other ocular abnormalities. RESULTS: A total of 3,431 and 3,467 students were enrolled in 2000 and 2001, respectively. The prevalence of normal vision (VA 20/20), VA 20/30 or better in at least one eye and 20/40 or less in at least one eye were similar in both years (87%, 5.7%, 7.3% and 85%, 6.4%, 8.7%, respectively). There was no statistically significant difference in visual acuity among boys and girls in either year (p = 0.6 and p = 0.2). Prevalence of abnormal color vision was 4.2%. Other causes of visual impairment in both years included strabismus (1.5% and 6.2%), amblyopia (1.1% and 1.4%) and some congenital abnormalities. Most cases of amblyopia were due to uncorrected refractive errors. CONCLUSION: The authors found that over 10% of school-aged children had subnormal vision. The important causes of visual deterioration came from refractive errors, strabismus and amblyopia. The authors concluded that vision screening is a cost-effective way of reducing visual morbidity from preventable visual impairment, which is a tragedy that cannot be ignored. PMID- 15279350 TI - Prevalence and determinants of overweight and obesity in Thai adults: results of the Second National Health Examination Survey. AB - To describe the prevalence of overweight and obesity and examine their relationship with socio-demographic factors in Thai adults. Using data from a cross-sectional survey, the National Health Examination Survey II (NHESII), the authors examined the prevalence of overweight (BMI > or = 25 kg/m2) and obesity (BMI > or = 30 kg/m2) in 3,220 Thai adults aged 20-59 yr. Univariate analyses and Logistic regression models were used to examine the association of overweight and obesity with socio-demographic and behavioral risk factors. The overall age adjusted prevalence of overweight and obesity were 28.3% and 6.8% respectively, with a higher prevalence for women than for men (overweight: 33.9% vs 19.2% and obesity: 8.8% vs 3.5%). The prevalence of overweight and obesity was greater among older compared to younger people and among residents of urban (34.8% and 9.9%) compared to rural areas (26.4% and 5.9%). The prevalence of overweight and obesity varied by region in line with the level of economic development--Bangkok, Central, North, South and North-East. By using logistic regression analysis, overweight was associated with a number of characteristics as follows: age (per ten years increase) with adjusted Odds Ratio (OR) of 1.3; women 1.4; married 2.2; being a current smoker 0.4, and living in Bangkok and the central region 1.6 (compare to North-East). There was no clear difference in prevalence of overweight and obesity among education levels and type of occupation after controlling for other covariates. In conclusion, women of middle age, married, and living in Bangkok and the Central region, are at greater risk of overweight and obesity. Without effective lifestyle modification programs to curb these physiologic risk factors at population level, it is likely that related disease burden will ensue. Public health surveillance and intervention to modify the risk factors of excessive weight should be implemented. PMID- 15279351 TI - Acute suppurative parotitis. AB - The parotid is the salivary gland most affected by inflammatory process. The authors reviewed the records of 36 patients with acute suppurative parotitis admitted between July 1996 and June 2002. All patients had unilateral swelling of the cheek that extended to the angle of the jaw; 23 had fever (64%) and 6 diabetes (17%). Even though Staphylococcus aureus is the usual pathogen in acute suppurative parotitis, in the present study, alpha-hemolytic streptococci predominated. Adequate hydration and administration of parenteral antimicrobials are essential managements of suppurative parotitis. In the event a well formed abscess, surgical drainage was necessary. The choice of antibiotics depends on the causal microbes. PMID- 15279352 TI - Microbial air quality in mass transport buses and work-related illness among bus drivers of Bangkok Mass Transit Authority. AB - The air quality in mass transport buses, especially air-conditioned buses may affect bus drivers who work full time. Bus numbers 16, 63, 67 and 166 of the Seventh Bus Zone of Bangkok Mass Transit Authority were randomly selected to investigate for microbial air quality. Nine air-conditioned buses and 2-4 open air buses for each number of the bus (36 air-conditioned buses and 12 open-air buses) were included. Five points of in-bus air samples in each studied bus were collected by using the Millipore A ir Tester Totally, 180 and 60 air samples collected from air-conditioned buses and open-air buses were cultured for bacterial and fungal counts. The bus drivers who drove the studied buses were interviewed towards histories of work-related illness while working. The results revealed that the mean +/- SD of bacterial counts in the studied open-air buses ranged from 358.50 +/- 146.66 CFU/m3 to 506 +/- 137.62 CFU/m3; bus number 16 had the highest level. As well as the mean +/- SD of fungal counts which ranged from 93.33 +/- 44.83 CFU/m3 to 302 +/- 294.65 CFU/m3; bus number 166 had the highest level. Whereas, the mean +/- SD of bacterial counts in the studied air conditioned buses ranged from 115.24 +/- 136.01 CFU/m3 to 244.69 +/- 234.85 CFU/m3; bus numbers 16 and 67 had the highest level. As well as the mean +/- SD of fungal counts which rangedfrom 18.84 +/- 39.42 CFU/m3 to 96.13 +/- 234.76 CFU/m3; bus number 166 had the highest level. When 180 and 60 studied air samples were analyzed in detail, it was found that 33.33% of the air samples from open air buses and 6.11% of air samples from air-conditioned buses had a high level of bacterial counts (> 500 CFU/m3) while 6.67% of air samples from open-air buses and 2.78% of air samples from air-conditioned buses had a high level of fungal counts (> 500 CFU/m3). Data from the history of work-related illnesses among the studied bus drivers showed that 91.67% of open-air bus drivers and 57.28% of air conditioned bus drivers had symptoms of work-related illnesses, p = 0.0185. PMID- 15279354 TI - Mycobacterial skin infections: comparison between histopathologic features and detection of acid fast bacilli in pathologic section. AB - BACKGROUND: Detection of acid fast bacilli (AFB) in chronic granulomatous inflammation is an important clue for mycobacterial infection. DESIGN: A retrospective review of 104 pathologic sections (from 1994 to 2001) of suspected cases of mycobacterial (tuberculous and nontuberculous) skin infections to study histopathologic features and the correlation with the presence of AFB in the section was performed. RESULTS: All cases showed granulomatous inflammations that can be categorized into 4 types: mixed cell, suppurative, tuberculoid and palisading granuloma. AFB was found in 32 sections (30.77%). Ninety five specimens from 104 specimens were simultaneously cultured. AFB positive cases yielded higher positive cultural results, 17 from 29 cases (58.62%) compared to the AFB negative group, 23 from 66 cases, (34.85%). Mixed cell granuloma was the most common histologic feature, but suppurative granuloma was the most common histological feature (56.25%) in which AFB could be found, which was statistically significantly different from other types of granuloma. Tuberculoid granuloma was more common in the AFB negative group (20.83%) compared to the AFB positive group (9.37%) but the difference was not statistically significant. In cases that AFB could not be found, the inflammation tended to be located in the upper half of the dermis. CONCLUSION: AFB can be more frequently detected in suppurative granuloma that might be located in any portion of the dermis. This finding was not species specific. PMID- 15279353 TI - Comparative study of renal function between standard and modified anatrophic nephrolithotomy by radionuclide renal scans. AB - OBJECTIVE: To compare the changes in renal function after surgery between standard and modified anatrophic nephrolithotomy using the technetium 99m-DTPA renal scan in patients with complex staghorn calculi. MATERIAL AND METHOD: From July 2001 to March 2002, standard anatrophic nephrolithotomy (sANL) was performed in 7 patients with complex staghorn calculi and modified anatrophic nephrolithotomy (mANL) was performed in another group of 8 patients with the same condition. Preoperative and postoperative renal function were assessed by technetium 99m-DTPA renal scan. RESULTS: Mean patient age was 41 years in the sANL group and 45 years in the mANL group. Male to female ratio was 4:3 in the sANL group and 5:3 in the mANL group. Median operative time was 205 minutes in the sANL group compare with 180 minutes in the mANL group (P = 0.03). Median estimated blood loss was 300 ml. in the sANL group and 275 ml. in the mANL group (P = 0.17). Median percent reduction of GFR on the operated kidney was 9.13 ( 30.03 to -3.15) in the sANL group and 27.25 (-41.81 to -1.55) in the mANL group (P = 0.13). Residual small stone was seen in one patient of the sANL group and ESWL was performed for stone fragmentation. There were no serious short-term complications. CONCLUSION: The average operative time of sANL was longer than mANL however, sANL preserved more renal function than mANL. This study suggested that sANL should be the procedure of choice in patients who have compromised renal function. PMID- 15279355 TI - Bronchiolitis obliterans syndrome after heart-lung transplantation: a case report and review of literature. AB - Bronchiolitis obliterans syndrome (BOS) is regarded as a manifestation of chronic rejection after lung transplantation and remains the major cause of late morbidity and mortality after lung and heart-lung transplantation. The authors, herein, reported the first documented case of a patient who receiving heart-lung transplantation at our institute and developed BOS as a late complication. The patient presented 5 years after received heart-lung transplantation with progressive shortness of breath due to obstructive lung disease. He was diagnosed with BOS by typical clinical presentation, pulmonary function test and radiographic findings and there were no other identified etiologies of airway obstruction. The authors also reviewed the recent update on the diagnosis and management of BO after lung transplantation. PMID- 15279356 TI - Monosomy 7 in patients with aplastic anemia and paroxysmal nocturnal hemoglobinuria with evolution into acute myeloid leukemia. AB - We report two cases of Thai patients with aplastic anemia/paroxysmal nocturnal hemoglobinuria (AA/PNH) who subsequently developed acute myeloid leukemia (AML) at their terminal phase. Monosomy 7 was demonstrated upon karyotypic analysis of bone marrow in both cases at the time leukemia developed The first patient was a 25-year-old man diagnosed with AA at age 14, recovered from AA at age 15, developed PNH at age 21 and turned into AML at age 25. The second patient was a 27-year-old man diagnosed with PNH at age 22, developed severe AA at age 25 and turned into AML at age 27. This latter patient received anti-lymphocyte globulin when he developed severe AA but did not respond well whereas the first patient fully recovered from AA with anabolic hormone treatment. Time to diagnosis of AML in the patient who received immunosuppressive therapy was strikingly shorter than that who received conventional androgen therapy (2 years vs 11 years after AA, respectively). The presence of monosomy 7 in leukemic cells of both patients emphasizes its central role in the development of AML from AA/PNH. However, other factors such as choice of AA/PNH therapy and patients response may modulate the time to emergence of monosomy 7-carrying AML clone and frank leukemia. Further studies into the biologic and genetic mechanisms involved in the development of leukemic clone arising from AA/PNH should be explored. PMID- 15279357 TI - Dysphagia after total laryngectomy resulting from hypocalcemia: case report. AB - Although hypocalcemia is a common postoperative complication of patients who have undergone a total laryngectomy with total thyroidectomy for treatment of laryngeal cancer or cancer of adjacent organs and hypocalcemia can produce the symptom of dysphagia, there has never been a report that hypocalcemia is the cause of dysphagia in these patients. The authors reported two cases who had hypocalcemia after total laryngectomy with total thyroidectomy and presented with sudden and severe dysphagia. However, the symptom of dysphagia was dramatically improved after calcium replacement therapy. PMID- 15279358 TI - Hydatid disease of the liver: the first indigenous case in Thailand and review of the literature. AB - The authors describe the first indigenous case of hepatic hydatid disease in Thailand. A 58-year-old female presented with progressive right upper quadrant abdominal discomfort over a 6-month period. Ultrasonography and computed tomography showed a solitary cystic lesion 11 x 12 x 13 centimeter in size at the left lobe of the liver. She had never been abroad and had no livestock exposure. The first operation was complicated by spillage of the parasite which required a combination of albendazole and praziquantel and a second operation for intracystic instillation of hypertonic saline solution. Unfortunately, uncontrolled generalized seizures developed due to severe hypernatremia. She never regained consciousness and expired 3 weeks after admission. In addition, the authors also reviewed previous reports of hydatid disease in Thailand. To date, only 9 cases have been reported since 1932. PMID- 15279359 TI - Use predictive modeling to improve profitability under cap. PMID- 15279360 TI - Study reveals 'best practices' in plan administration. PMID- 15279361 TI - More california rates show some IPAs still fully capitated. PMID- 15279362 TI - Do capitated physicians have more incentive to prescribe beta-blocker therapy? PMID- 15279363 TI - Does higher Medicare spending reflect better care? PMID- 15279364 TI - [IgA nephropathy in pediatrics]. AB - IgA nephropathy is a primitive cronic idiopatic glomerulonephritis, characterized by diffuse depositis of IgA in the glomeruler mesangium. Familial cases are also descripted. IgA nephropaty is more frequent in males and in white rase. In Italy it's the most frequently recognized glomerulonephritis in renal biopsia (20%), especially in patients with dismorfic micro or macroematuria and nephrotic proteinuria. Clinical presentation is often in association with respiratory tract or gastrointestinal disorders. The most relevant pathogenetic hypothesis suggest an IgA abnormal glycosilation, with mesangial IgA aggregation, increased mesangial reactivity and release of inflammatory mediators and fibrotic agents. Treatment is considered in rapidly progressing forms. At the present, there is no treatment of proven value in all patients, althoug interesting results have been published with prednison, ACE-inhibitors or fish-oil in decresing renal deterioration rate. Natural history varies in different series. Renal survival at 10 years is 85% in Italy, 94% in France, 97% in the USA. Poor prognostic factor are heavy proteinuria and hypertension. However a wide inter-individual variability is observed. PMID- 15279365 TI - [Respiratory diseases associated to surfactant proteins B and C deficiency]. AB - Pulmonary surfactant is a mixture of lipids and proteins necessary to reduce alveolar surface tension and prevent end expiratory atelectasis. The hydrophobic surfactant proteins B and C are essential for lung function and pulmonary homeostasis after birth. Mutations in the genes encoding surfactant proteins B and C are associated with acute respiratory failure and interstitial lung disease. Diagnosis is possible through DNA analysis from blood leucocytes and immunostaining from lung tissue. PMID- 15279366 TI - [Head trauma: clinical and diagnostic factors in pediatric emergency]. AB - The Authors describe the clinical spectrum of head trauma. The importance of history (the way the trauma occurred) and of the intrinsic dynamics of the lesions are emphasized, as is their role for the outcome. They delineate the major intervention the pediatrician should perform in emergency, and the diagnostic and therapeutical approach. In particular, recommendations are made about the best neuroradiological test which should be done. PMID- 15279367 TI - [The management of the child with head injury: initial assessment]. AB - In western countries, head injury is very important in public health; infact it is the main cause of mortality after infancy and a significant cause of long term disability. The patients with severe head injury needs an intensive care unit management, but the initial approach is a critical phase, too. In fact, it is demonstrated that trauma mortality and morbility are significantly reduced by a quick and well conducted approach. This article focuses the initial management of the injured child. PMID- 15279368 TI - [The management of the child with head injury: our experience]. AB - OBJECTIVES: To analyze the group of children admitted to our Pediatric Intensive Care Unit (PICU) for neurotrauma and describe the management algorithms adopted by us for pediatric head injury. METHODS: All the children affected by head injury and admitted to PICU since november 1992 to november 2000 have been examined. Injury severity has been classified using the Glasgow Coma Score (GCS), while the long term neurological outcome with the Glasgow Outcome Score (GOS). We have described the clinical presentation, the kind and dynamics of injury and the clinical outcome one month after discharge. RESULTS: 210 children with head injury have been identified. Among them: 38 were affected by mild head injury, 50 by a moderate one and 122 by a severe one. The most frequent cause of injury has been represented by car accidents and motorbike or bicycle falls. The overall outcome has resulted good (GOS 4-5) in all children affected by mild or moderate head injury; on the other side, patients with severe injury have presented the following scores: GOS 1 (dead children) 14.7%, GOS 2 (persistent vegetative state) 1.6%, GOS 3 (severe disability) 22.2%, GOS 4 (mild disability) and GOS 5 (no disability) 61.5%. CONCLUSION: A correct management of children with head injury helps clinicians to improve outcome and to reduce mortality. Therapeutics algorithms suggested by us could be useful for the management of this kind of patients, not only when they are affected by a severe head injury but, also, when they suffer from a mild one, that is the most common event in the emergency room departments. PMID- 15279369 TI - [Visual motor and visual defects in spina bifida]. AB - Among the numerous problems that spina bifida (SB) patients are faced with, impairments to the visual apparatus are often considered late and are not covered extensively in the literature. At the Pediatric Department of the University of Padua, an assessment of the visual function of 59 SB patients between 5 months and 26 years of age (29 male and 30 female) has been carried out by means of an ophthalmologic protocol. As far as the alteration of the exstrinsic ocular mobility is concerned, 44% (26/59) of our patients revealed a manifest squint and only 3% (two patients) suffered from a latent squint. The most frequent type is a convergent squint (80%). The assessment of visual acuity made through Optotype was good in 82% of the cases (unlike what is commonly reported in the literature) and mild in 18%. None of the patients manifested hypovision. Refraction defects were present in 59% (34/59) of the patients. Regular ophthalmologic evaluations from birth or from diagnosis allow ophthalmologic treatments that are tailored to children suffering from SB and also enable them to reach and maintain a good visual standard and to observe the subtle symptoms of endocranial hypertension sooner. An early discovery and correct treatment of visual problems improves cognitive and motive performance as well as the autonomy of SB patients. PMID- 15279370 TI - [Kluver Bucy syndrome and central diabetes insipidus: two uncommon complications of herpes simplex encephalitis]. AB - Herpes Simplex Encephalitis (HSE) is an uncommon but severe disease with high mortality and morbidity. The major clinical manifestations are deteriorating consciousness with confusion, drowsiness or coma, altered behaviour, convulsions and a variety of neurological signs (hemiplegia, aphasia, ataxia, etc.). An uncommon complication of HSE is Kluver Bucy syndrome (KBS), characterized by hyperorality, bulimia and changes in emotional behaviour. Neuroimaging studies frequently show an involvement of the temporal lobes and limbic areas. Another uncommon complication of HSE is central diabetes insipidus as a result of herpes simplex infection of the hypothalamus. We report two pediatric cases of HSE complicated with Kluver Bucy syndrome and central diabetes insipidus. PMID- 15279371 TI - [Kearns-Sayre syndrome associated with growth hormone deficiency]. AB - The Authors report a 10 years old boy with Kearns-Sayre syndrome and growth hormone (GH) deficiency. The patient was treated with human recombinant GH for 6 months but the growth velocity/year did not change. The Authors report a brief review of the literature on the ethiopatogenesis of GH deficiency observed in this patient is given. PMID- 15279372 TI - [Ascariasis as a cause of acute abdomen: a case report]. AB - Ascaris Lumbricoides Infestation (ALI) is one of the most common helmintic disease of the gastrointestinal tract, and may cause severe surgical complications, especially in children. ALI is frequent in tropical and subtropical countries. We present our experience with a case of a 5-years old pakistan girl treated in Italy for acute abdomen in which ALI was detected during surgical exploration. PMID- 15279373 TI - The development of peace. PMID- 15279374 TI - [Can the scientist play his role in the maintaining of peace?]. PMID- 15279375 TI - Tolerance as a way to accept and enjoy the diversity. PMID- 15279376 TI - "Blessed are those who work for peace". PMID- 15279377 TI - The immeasurable value of the "breath of life". PMID- 15279378 TI - Personal experience in Bosnia. PMID- 15279379 TI - [Introduction to the "Queen Sofia Institute" X international symposium on nephrologic research]. PMID- 15279380 TI - [Statins, inflammation and atherosclerotic damage]. PMID- 15279381 TI - [Nitric oxide and extracell matrix deterioration. A role in atherosclerosis?]. PMID- 15279382 TI - [Oxidative stress in hypertension: from the SNPs to the clinical phenotype]. PMID- 15279383 TI - [Does the lipidic metabolism of myocardium modulate cardiac hypertrophy?]. PMID- 15279384 TI - Cardiovascular disease in chronic renal failure. Risk factors and prevention. PMID- 15279385 TI - [Vascular calcification mechanisms in uremia]. PMID- 15279386 TI - [Left ventricular hypertrophy after renal transplant: prevention and treatment]. PMID- 15279387 TI - [Analyses of genetic risk factors associated to chronic nephropathy of renal transplant: genetic polymorphisms of cytokynes, adhesion molecules, platelet coagulation system and cardiovascular risk markers]. PMID- 15279388 TI - [Protocol biopsies: the importance of the renal transplant vasculopathy]. PMID- 15279389 TI - [Anti HLA post-transplant antibodies. A new method of monitorization]. PMID- 15279391 TI - Overseas transcription: is it safe? PMID- 15279390 TI - [Immunologic mechanisms inducing tolerance to allografts]. PMID- 15279392 TI - Clinical guidelines sword or shield? PMID- 15279393 TI - I cut incoming calls 40%. PMID- 15279394 TI - Our unspoken promise. PMID- 15279395 TI - 5-6 months out: office design and supplies. PMID- 15279396 TI - What to buy for a new practice. PMID- 15279397 TI - In this border-town dilemma. PMID- 15279398 TI - Your staff needs CME, too. PMID- 15279399 TI - Medical errors: lessons from aviation. PMID- 15279400 TI - Helping patients. Protecting yourself. PMID- 15279401 TI - Your liability for a locum. PMID- 15279402 TI - Pharmacological treatment of dementia. PMID- 15279403 TI - [The biology of myelodysplastic syndromes]. AB - In the past 10 years a large amount of data has been published with respect to myelodysplasia. However, this newly emerged data, to date, has not been integrated into an all encompassing cohesive concept. The more knowledge we have about this illness to more precise diagnostic classifications, prognostications and eventually curative treatment modalities are possible. The ineffective haemopoiesis that is a hallmark of the disease may be explained by the apoptosis, hence antiapoptotic therapy, it appears, in some cases results in improvement. It is known that the malfunction of the mitochondria is responsible for accumulating the iron in sideroblastic anemia. The same mitochondrial malfunction may also be a factor in the development and progression of the non-sideroblastic forms of myelodysplasia as well. Promising research is on the way in examining the interaction between stroma cells of the bone marrow and the stem cells. Several publications study the hypothesis that the stroma also plays a significant role in the development of myelodysplasia clone. This role may be a causative one or perhaps a more passive/permissive one. This paper attempts to provide an overview of the newly emerged data, published in the most recent years, concerning myelodysplasia. PMID- 15279404 TI - [Examination of hepatitis c virus antibody seropositive blood donors and their relatives]. AB - INTRODUCTION: 239 anti-HCV seropositive blood donors (132 male, 107 female, age: 19-61, mean: 40.59 y.) and 174 family members of them (74 male, 100 female, age: 4-65, mean: 23.67 y.) were studied for chronic hepatitis C virus infection and chronic liver disease. Detailed virus serology, ultrasonography, and 6 months follow-up and--in patients with HCV RNA--liver biopsy were made. RESULTS: HCV RNA was determined in 165 patients. 70% of them were HCV RNA positive. The ALT level was normal in 95 cases (57%), and lower, than twice of the normal was in 34 cases (20%) among them. Liver biopsy was made in 79 patients; chronic C hepatitis was proven in 75 cases (steatosis in 3 cases, alcoholic liver disease in 1 case was observed). Inflammatory activity was minimal (HAI < 3) in 17, mild (HAI: 3-6) in 41, moderate (HAI: 7-9) in 7, and severe (HAI > 9) in 10 cases. There was no correlation between the serum ALT levels and the severity of the histological activity of chronic C hepatitis. Authors stress the importance of the fact, that 2 patients had normal ALT and 5 patients ALT levels were lower, than the twice of the normal of the 17 patients with significant inflammatory activity (HAI < 6). Chronic C hepatitis need for antiviral therapy was occurred in 45% of patients who known themselves previously healthy. CONCLUSIONS: The necessity of the systematic examination of anti-HCV seropositive patients and of the importance of the liver biopsy in patients with HCV RNA positivity is stressed. 3 anti-HCV seropositive cases of 174 family members of the blood donors were observed, but none of them was HCV RNA positive. It seems to be, family members of the HCV infected patients have no increased risk for HCV infection. PMID- 15279405 TI - [Microcirculatory changes in patients with chronic venous and lymphatic insufficiency and heavy leg symptoms before and after therapy with procyanidol oligomers (laser-Doppler study)]. AB - INTRODUCTION: To study the efficacy of venotonica and medicaments having anti edema effect is very actual. AIMS: Patients suffering from venous-lymphatic insufficiency were treated with procyanidol oligomers in an open, prospective study. The effective substance of this medication protects the fibrous connective tissue protein, thus improving the function of the venous and lymphatic capillaries. METHODS: For three months, the daily dose was 2 x 150 mg 30 patients were included into the study. The efficacy of the treatment was evaluated by statistical methods analysing the changes of the clinical symptoms and the satisfaction of the patients. For the first time, laser-Doppler method was used to determine the pathological condition of the dermal microcirculation before and after 3 months of treatment. The laser-Doppler tests used as indicators of the functional changing of the microcirculations were as follows: venoarterial response, reactive hyperemic response, and thermal stimulation response. The authors established the standard values and compared the measurement scores to those. According to the resulting scores, the status before and after the treatment can be compared by statistic analysis. Subjective complaints of the patients such as pain, heaviness of the leg, nightly muscle cramps were estimated by a visual analog scale. RESULTS: The treatment (procyanidol oligomers) significantly reduced lymphedema (p < 0.001) in the "heavy leg" syndrome. Symptoms as pain, heaviness of the leg, muscle cramps, improved significantly (p < 0.001) after treatment. The microcirculatory status was pathologic before treatment. After treatment, there was no significant improvement in the laser Doppler results. CONCLUSIONS: The study revealed that in venous-lymphatic insufficiency, the arterial-capillary system is also damaged. The authors bring to attention that the disorder of the dermal arteries playing a role in the pathogenesis of venous-lymphatic insufficiency first detected in this study needs to be further investigated. PMID- 15279406 TI - [DOOR (deafness, onychodystrophy, osteodystrophy, mental retardation) syndrome]. AB - Authors highlight the difficulties of syndrome identification through reporting the first case of DOOR syndrome in Hungary (the 28th case worldwide). The awareness and appropriate weighing of the importance of vestigial nails (onychodystrophy) was crucial for the correct diagnosis. Based on the normal level of 2-oxoglutarate excretion, the patient can be categorized as type 2. This is associated with better survival, which does not mean a substantial difference in quality of life. Although, prenatal diagnosis is not possible at present, knowledge of the enzyme defect and detection of the reduced activity of the 2 oxoglutarate dehydrogenase E1 component may provide an opportunity. If parents opt to have another child, a 25% risk is to be taken into account. PMID- 15279407 TI - [Treatment of community acquired pneumonia]. PMID- 15279408 TI - ["Horns" of man]. PMID- 15279409 TI - [About the nephrosis-syndrome]. PMID- 15279410 TI - [Generations' succession]. PMID- 15279411 TI - [Timolol topical ophthalmic combination]. AB - The primary standard therapy in patients with open-angle glaucoma and ocular hypertension is carried out by means of monotherapy with synthetic prostaglandin analogues. Most of the glaucoma patients need more than one medication for adequate intraocular pressure control. Timolol represents the basic component of these combinations. Timolol topical ophthalmic combinations with dorzolamide (Cosopt), pilocarpine, latanoprost (Xalacom), travoprost and unoprostone are thoroughly described. Most antiglaucoma medications achieve on one side directly baroprotection by decreasing intraocular pressure and on the other side they produce indirectly vasoprotection, that is secondary to intraocular pressure lowering. The Cosopt, due to its Dorzolamide component, makes an exception since it produces directly both baroprotection by aqueous humor flow suppression and vasoprotection by increasing the blood flow within the retina, choroid, optic nerve head and retrobulbar area. Given these considerations as well as the fact that the ocular hypotensive effect of both the Cosopt and the latanoprost are equally potent, a question seems reasonable according to accepted standard i.e. Cosopt or latanoprost as primary glaucoma therapy? PMID- 15279412 TI - [Peripheral alexias]. AB - The brain lesions could lead to impairments of the comprehension and production of written language. This acquired inability is named alexia. It is a significant problem for neurologists and ophthalmologists. Our study presents a classification of the alexias, whose pathology was describe first by Dejerine (1891; 1892). There are two varieties of alexias: central alexias and peripheral alexias (especially agnozic alexia and attentional alexia). In agnozic alexia, the patient cannot read, but can write, understand and speak. It results from a type of cerebral disconnection in which the angular gyrus of the dominant hemisphere is disconnected from its bilateral visual input. The most commonly reported pathology is occlusion of the dominant (left) posterior cerebral artery, which leads to infarction of both the left occipital lobe (causing partial or complete right homonymous hemianpsia) and the splenium of the corpus callosum. PMID- 15279413 TI - [Nonconventional ophthalmological diagnosis--case report]. AB - The paper brings up a case of ophthalmological simulation in a ten-year old girl. The sequence of investigations and functional explorations performed is reviewed, in chronological order, as well as the unconventional method of restoring the visual function. PMID- 15279414 TI - [Systemic thrombosis in central retinal artery occlusion--case report]. AB - Central retinal artery occlusion is a dramatic event, a major emergency; presentation is with an acute, painless and profound loss of vision. The patient usually comes too late to the ophthalmologist, the treatment becoming inefficient. Therefore we present a case of CRA occlusion managed by systemic thrombolysis. PMID- 15279415 TI - [Persistent hyperplastic primary vitreous associated with retinal folds]. AB - The paper presents the case of a 18 years old male suffering from persistent hyperplastic primary vitreous (PHPV) associated with congenital retinal folds. The clinical features and the pathogenic correlations of the two affections are discussed. Comparing to the PHPV, consequence of a embryogenesis flow appeared in the development of the primary hyaloid-vitreous complex, the congenital retinal folds are considered to be the expression of secondary changes, generated by the background of a varied vitreo-retinal pathology. PMID- 15279416 TI - [LASIK after radial keratotomy--case report]. AB - LASIK after radial keratotomy is difficult and intraoperative risks are high. In this article I present the case of a patient who underwent such an operation. PMID- 15279417 TI - [Structural and immunohistochemical changes of conjunctiva induced by topical glaucoma medication]. AB - PURPOSE: The topical medication represents the first line therapy for the primary open angle glaucoma. The study is aimed at assessing the structural and immunohistochemical changes of conjunctiva induced by topical glaucoma medication. METHODS: For this purpose, we carried out a 40 weeks, prospective, experimental, epidemiological-operational and randomized study enrolling 18 patients (36 eyes) with recently primary open angle glaucoma. The eyes were divided into treatment (non-selective beta blockers or selective, prostaglandin analogues, topical carbonic anhydrase inhibitors) in four groups. The assessment was performed by comparison with control group (4 patients) who was instilled with natural tears (with different preservative). Both the cytology and the conjunctival biopsy specimens were investigated by histological exams and immunohistochemistry using different monoclonal antibodies. The study was performed by collaboration with The National Institute of Research and Development in Pathology and Biomedical Sciences-"V. Babes"-Bucharest. RESULTS: The morphometric analysis of histological sections of conjunctiva showed the following changes: squamous metaplasia (significant increases in the thickness and number of epithelial cell layers), inflammation (increase the number of lymphocytes, macrophages and fibroblasts) and subconjunctival fibrosis. According to the type of medication, we observed the significant increase of subepithelial collagen density and degenerative changes of fibrocytes, the reduction of extracellular matrix and also the up-regulation of antibodies against matrix metalloproteinase and allergic changes. According to structural changes, the immunohistochemistry confirmed the tendency of chronic inflammation. CONCLUSIONS: This study revealed important structural and immunohistochemical changes of conjunctiva after topical glaucoma medication. The category and the intensity of these changes are dependent on the sort of therapy and the topical treatment period. The findings showed that benzalkonium chloride (the most common preservative of antiglaucoma drugs) is a major factor for conjunctival metaplasia. PMID- 15279418 TI - [The adaptometry study in patients with diabetic retinopathy]. AB - PURPOSE: The aim of this paper is to study if the cones and rods adaption is influenced by the degree of the diabetic retinopathy or by the retinal photocoagulation treatment. MATERIAL AND METHOD: I analysed three groups of diabetic patients: the first group without diabetic retinopathy, the second with different stages of diabetic retinopathy without laser treatment and the last one treated by panphotocoagulation. RESULTS: The exploration of the global light and dark adaptation of the retina using Hartinger's adaptometer does not allow a qualitative and selective appreciation of this function. However it suggests quantitative alterations of the terminal limit in patients with diabetic retinopathy treated or not by panphotocoagulation. CONCLUSIONS: The diabetic retinopathy produces minor symptoms of low dark adaptation even in the advanced phase. The diabetic retinopathy induces minimal changes of the graph of the adaptation limits even in the patients without treatment. PMID- 15279419 TI - [Phacoemulsification--personal experience on my first 507 cases]. AB - PURPOSE: The evaluation of the intra and post operative early complications of the phacoemulsification with posterior chamber artificial lens implant on my first 507 cases. METHODS: A retrospective study on 507 eyes operated by cataract through phacoemulsification technique at the Emergency Military Hospital from Cluj-Napoca during the 1st of May-30th of June 2002. The surgeries were performed by the same surgeon through a corneal or a scleral incision using rigid and foldable artificial lenses. The evaluation period lasted 2 months, the types and frequency of the intraoperative and early postoperative complications being analyzed. RESULTS: The intraoperative complications were represented by the posterior capsule's efraction in 17 cases (3.37%); zonular rupture in 3 cases (0.59%); the loss of the nucleus into vitreous 4 cases (0.79%); the luxation of the crystalline lens in the anterior chamber in 5 cases (0.99%); the hyphaema and iris trauma in 14 cases (2.76%); incomplete cortical clean-up in 9 cases (1.78%); the failed capsulorhexis in 7 cases (1.38%) and the conversion to ECCE in 8 cases (1.58%). The early postoperative complications met into the study were: the corneal edema in 34 cases (6.71%); raised IOP in 18 cases (3.55%); the retained lens material in 9 cases (1.78%); the late inflammatory reactions in 6 cases (1.18%); hypopyon in 3 cases (0.59%); endophthalmitis in 3 cases (0.59%); retinal detachment in 1 case (0.2%) and the anterior capsule fibrosis in 4 cases (0.79%). CONCLUSIONS: The phacoemulsification is a high security technique because of the small incision and the sharp accuracy. The complications correlate themselves with the surgeon experience and with some particularities of the cataract (the brunescent cataract at the old patient, the medium dilatation of the pupil, etc.). PMID- 15279420 TI - [The visual evoked potentials in diabetic retinopathy]. AB - PURPOSE: The recording of Visual Evoked Potential alterations at the patients with diabetic retinopathy. MATERIAL AND METHODS: It was performed the Visual Evoked Potential recordings at 24 patients with diabetic retinopathy in different stages of evolution, with or without complications. The type of Visual Evoked Potential recording was pattern reversal with vertical bars. We followed the diagram alterations in correlation with the evolution stages of diabetic retinopathy and the visual parameter alterations. RESULTS: In all cases we recorded alteration of the Visual Evoked Potential. In nonproliferative diabetic retinopathy was noticed the delay of P100 wave with inconstant presence of the N75 and N135 waves. In proliferative diabetic retinopathy and its complications the alterations of the tract were important. CONCLUSIONS: The gradual alteration of the Visual Evoked Potential tract at the patients with diabetic retinopathy represents a prognosis of the disease. PMID- 15279421 TI - [Lens extraction in malignant myopia]. AB - This issue presents the results of cristalin extraction in myopic eyes, analysing especially the postoperatory visual acuity and refraction. Some particularities of lens extraction in myopic eyes are pointed and the place of lens extraction as a method of refractive surgery of myopic eyes is analysed. PMID- 15279422 TI - [The visual evoked potentials in open angle glaucoma]. AB - PURPOSE: The study purpose to analyse the variations of Visual Evoked Potentials on a group of patients with Open Angle Glaucoma. MATERIAL AND METHOD: It was performed the Visual Evoked Potential recordings on a number of patients with Open Angle Glaucoma in different stages of evolutions. Some of the patients were monitored by the Visual Evoked Potentials recordings in a dynamic way, before and after the law pressure treatment. The type of Visual Evoked Potentials recording was pattern reversal with vertical bars. RESULTS: In incipient stage of Open Angle Glaucoma the alteration of the tract was minimal with inconstant presence of the N75 and N135 waves and just a few variation of the delay of P100. In advanced stages of Open Angle Glaucoma the alterations of the tract were important: the delay of P100 wave. In absolute stage the tract was null. CONCLUSIONS: The alterations of Visual Evoked Potentials tract was influenced by the different stages of evolution of Open Angle Glaucoma. The gradual alteration of the Visual Evoked Potential tract represents a prognosis of the disease. PMID- 15279423 TI - [Ocular parasitosis--particular aspects]. AB - The goal of this study cases are presented in this study. The first case was emphasized in a patient with an epibulbar tumor and the second case was diagnosed with Toxocara Canis. In both cases the ophthalmoscopic images and the clinical evolution were unusual on the other hand etiological diagnosis in both cases was difficult. In the first case, the surgical procedure was effective, but in the second case the specific antiparasitic treatment could not stop the evolution of corioretinal's lesions; it required laser therapy. PMID- 15279424 TI - [Lacrimal secretion in hormonal imbalance]. AB - The aim of this study is the alteration of lacrimal secretion on a group of female patients with deregulations of the hormonal balance, by the influence of age factor. We have to mention that our female patients have no ocular pathology. MATERIAL AND METHOD: The study was conducted on a group of patients aged between 20-70 years old, which has been kept in observation in the Endocrinology Clinic and Obstetrics-Gynecology Clinics of Emergency Hospital, during March-August 2003. Their lacrimal secretion was monitored by volumetric tests (Schirmer). RESULTS: We studied the alteration of the lacrimal secretion on female patients with deregulations of the hormonal balance, by the influence of age factor. CONCLUSIONS: It was recorded the alteration of lacrimal secretion on the female patients with aforementioned dysfunction, the age factor being influential. PMID- 15279425 TI - [Incidence of the uveal pseudoexfoliation syndrome in patients with diabetes mellitus]. AB - PURPOSE: To study the incidence of the uveal pseudoexfoliation syndrome in diabetic patients with or without diabetic retinopathy. MATERIAL AND METHOD: There were studied a group of diabetic patients with different ages, with or without diabetic retinopathy; the control group was represented by the non diabetic patients with similar ages. RESULTS: It has been recorded a high incidence of the uveal pseudoexfoliation syndrome in diabetic patients compare with non-diabetic patients at the same age. CONCLUSIONS: It seems that uveal pseudoexfoliation syndrome is determined by a collagen metabolic alteration which is also present in diabetes mellitus, among other metabolic alterations; this will be the explication for high incidence in diabetic patients. PMID- 15279426 TI - [Axial length and branch retinal vein occlusion]. AB - PURPOSE: To determine whether there is a connection between the axial length and the occurrence of the branch retinal vein occlusion (BRVO). METHOD: 18 patients that had been diagnosed with BRVO have been selected retrospectively. The diagnosis has been made according to the clinical criteria (fundus aspect). A control group of 18 patients has ben constituted. The patients in the control group matched the age distribution and the prevalence of high arterial blood pressure and diabetes mellitus of the patients in the BRVO group. The measurements of the axial lengths have been performed by the same examiner, for the affected and fellow eye, in the BRVO group and for the right eye, in the control group. The data were interpreted by the help of the descriptive statistics. The Student t test has been used and was considered significant, if the p value was < 0.05. RESULTS: The mean axial length of the eyes with BRVO was 22.42 mm (range, 21.5 mm-24.1 mm) and of the fellow eyes, 22.44 mm (range, 21.3 mm-24.2 mm). The difference is not statistically significant: p = 0.776. The mean axial length of the eyes in the control group was 23.42 mm (range, 21.5 mm-24.4 mm), the difference being statistically significant when compared with the BRVO eyes: p = 0.00064. DISCUSSIONS: The possible connections between a shorter axial length and the incidence of BRVO are analyzed, taking into account the known factors that are involved in the pathogenesis of this disease. CONCLUSION: The shorter axial length is a risk factor for BRVO. PMID- 15279427 TI - [Treatment modalities in Duane syndrome]. AB - Stilling-Turk-Duane syndrome is a restrictive syndrome caused by an abnormal innervation of the lateral rectus muscle, which is the result of nerve VI agenesia. One of the most important feature is represented by "co-contraction phenomenon" which means the simultaneous contraction of lateral and medial muscles in adduction. The treatment is individualised and most patients may benefit of surgical procedures. The goals of surgical treatment is: improved head posture, elimination of vertical deviation, reduced of enophthalmos and normal ocular alignment. This issue represent the surgical management and the postoperative results in 3 cases of Stilling-Turk-Duane syndrome treated in Eye Clinic, Iasi. PMID- 15279428 TI - Vitrectomy for severe penetrating trauma involving the posterior segment. PMID- 15279429 TI - [Lagophthalmy of VII nerve palsy--therapeutical approaches]. AB - Facial nerve palsy is impairing orbicular muscle function leading to lagophthalmos. This disease impairs patients' quality of life by exposure keratopathy which is accompanied by ocular pain and deteriorate visual acuity at a variable degree. The paper's aim is to show up the place of lagophthalmos in anterior ocular segment pathology and to evidence the results obtained by us with a new therapeutical method applied in several cases. All cases with lagophthalmos have ocular complications like exposure kerato-conjunctivitis. The treatment was both conservative and surgical which consisted in majority of cases in blepharoraphy with its inconveniencies, aesthetical and functional. We have applied a new surgical method--gold weight superior lid implantation--in 4 cases and the results are promising. PMID- 15279430 TI - [Our experience in using laser excimer]. AB - In this paper we present our experience concerning 658 excimer laser operations: statistical data about the number of operations, preoperative investigations, postoperative care and postoperative complications. PMID- 15279431 TI - Effect of articulatory suppression on task-switching performance: implications for models of working memory. AB - In a series of experiments, we examine some effects of articulatory suppression in task switching. The results from Experiments 1a and 2a showed that switch costs in the articulatory suppression condition were larger than those in the control and tapping conditions when the switching cues were not provided. On the other hand, articulatory suppression did not have any effect on switch costs in Experiments 1b and 2b, where the switching cues were provided. In Experiment 3, using a computer-assisted experimentation, this pattern of data was replicated in a two-factor design with articulatory suppression and switching cues factors. The results indicate that a specific component in working memory, the phonological loop, might contribute to the performance in task switching, at least in situations where the external task cues were not available. The data reported here suggest that the phonological loop plays an important role in one of the executive control processes, and challenge the traditional idea that the slave systems are simply governed by the central executive in the working memory. PMID- 15279432 TI - Simulating episodic memory deficits in semantic dementia with the TraceLink model. AB - Although semantic dementia is primarily characterised by deficits in semantic memory, episodic memory is also impaired. Patients show poor recall of old autobiographical and semantic memories, with better retrieval of recent experiences; they can form new memories, and normal performance on pictorial recognition memory has been demonstrated. As these abnormalities in episodic memory are virtually a mirror image of those seen in the amnesic syndromes, semantic dementia poses a challenge to extant models of remote memory and amnesia. Here, we show that one such model, TraceLink, can reproduce some of the principal findings on episodic memory in semantic dementia. A loss of nodes and connections within the trace system, which can be identified with the temporal neocortical memory storage sites implicated in semantic dementia, simulates without further assumptions the findings reported above. PMID- 15279433 TI - Memory conjunction errors for autobiographical events: more than just familiarity. AB - Two diary studies investigated the prevalence of memory conjunction errors for real-life events. In both studies, participants completed detailed diary pages over the course of several weeks. Participants in both diary studies committed memory conjunction errors on a later recognition memory test. In the second diary study participants also made remember/know judgements. For a large proportion of their memory conjunction errors participants indicated that they "remembered" the event occurring in that context. These diary studies demonstrate that memory conjunction errors do occur for autobiographical memories, and these errors can seem compelling. PMID- 15279434 TI - Effects of working memory load on long-term word priming. AB - Three experiments assessed the role of verbal and visuo-spatial working memory in supporting long-term repetition priming for written words. In Experiment 1, two priming tasks (word stem completion and category-exemplar production) were included with three levels of load on working memory: (1) without memory load, (2) memory load that involved storing a string of six digits, and (3) memory load that involved storing a graphic shape. Experiments 2 and 3 compared the effects of a verbal (Experiment 2) or a visual (Experiment 3) working memory load at encoding on both an implicit (word stem completion) and an explicit test (cued recall). The results show no effect of memory load in any of the implicit memory tests, suggesting that priming does not rely on working memory resources. By contrast, loading working memory at encoding causes a significant disruptive effect on the explicit memory test for words when the load is verbal but not visual. PMID- 15279435 TI - Opposing effects of phonological similarity on item and order memory of words and nonwords in the serial recall task. AB - This study shows that the classical phonological similarity effect (PSE) in immediate serial recall is critically affected by the lexicality of list items, the type of phonological similarity involved, and the scoring procedure. PSE was present in the serial recall score when phonologically distinct words were compared to words that share the middle vowel and end consonant (rhyming lists). PSE was absent in the serial recall score when phonologically distinct words were compared to words that share the initial and final consonants (consonant frame lists). There was a reversal of PSE in serial recall of nonwords when comparing distinct lists to both types of similar lists. Recall accuracy on the other hand was higher for distinct lists regardless of lexicality. Item errors dominated in relation to order errors in the case of nonwords, whereas order errors dominated in relation to item errors in the case of words. Furthermore, order errors were more common for phonologically similar lists, whereas item errors were more common for phonologically distinct lists. This may be the result of intra-list and inter-list interference, respectively. The dominance of the former error type may cause a classical PSE, whereas the dominance of the latter error type may cause a reversal of PSE. Finally, an item identification task yielded no evidence of an association between intra-list interference and discriminability of items in a list. PMID- 15279436 TI - Complexity factors in visuo-spatial working memory. AB - Three experiments are presented that use a technique of selective interference- irrelevant pictures--to develop our understanding of visuo-spatial working memory. Visual noise fields are used as the irrelevant pictures. Using two related measures of simple visual complexity, the experiments demonstrate that the greater the complexity the greater the degree of interference, even within a paradigm where subjects are instructed to look at but otherwise ignore the irrelevant pictures. Both the number of dots and the density of the dots comprising the visual noise fields affect the degree of interference in a concurrent memory task. In addition, increasing the size of the field increases the amount of interference. It is argued that the results give insight into the properties of visual working memory and contribute to its theoretical development. For example, it is argued that the store is directly accessible by externally presented interference and that particular aspects of the noise displays cause interference with visual memory. PMID- 15279437 TI - Reminiscence, forgetting, and hypermnesia using face-name learning: isolating the effects using recall and recognition memory measures. AB - A face-name learning paradigm was used to study phenomena involved in reminiscence, forgetting, and hypermnesia. Individuals introduced themselves on videotape while participants tried to learn their names. The presence of cues during testing increased overall performance but decreased hypermnesia in Experiment 1. Significant recognition memory effects were found for reminiscence and hypermnesia in Experiments 2 and 3. Experiment 3 also showed no interference from activities between testing sessions, but did show facilitating effects from exposure to photographs of target faces and to exposure of target names. The results were interpreted as showing support for reminiscence effects being primarily caused by imagery redintegration and effects consistent with stimulus sampling theories. PMID- 15279438 TI - Scented memories of literature. AB - This study examined the effects on recall of story details of congruity or incongruity between the hedonic valence of literary texts and odours inhaled while reading them. During the reading session, 24 undergraduates (12 males and 12 females) read two passages involving positive subject matter and two with negative subject matter while sniffing pleasant or unpleasant odours in a within subject fully counterbalanced design. Subjects rated their experience of each text on eleven 7-point scales. During the test session 48 hours later, subjects read a two-word title associated with each of the passages and inhaled the odour that was paired with it in the reading session. They also rated their experience on six of the scales that had been used during the reading session. Results showed that hedonic congruence between the passage and the odour fostered enhanced recall during the test session. The combination of positive subject matter and positive odour was reflected in more accurate recall of character details, while pairing negative subject matter and negative odour resulted in more accurate recall of setting details. Regression analysis showed that overall recall accuracy was increased by identifying with the characters in the stories and for passages that were found pleasing and personally meaningful. Consistent with the literature on implicit learning involving odours, recall accuracy varied inversely with perceived odour intensity. Implicit learning involving odours and literary passages is therefore fostered by unity in the reading experience. PMID- 15279439 TI - Effects of environmental context manipulated by the combination of place and task on free recall. AB - Three experiments, using a 2 (study context) x 2 (test context) between-subjects design, were conducted to examine the effects of environmental context manipulated by the combination of two contextual elements, place and task, on free recall. Undergraduates individually studied nouns and received a free-recall test, with a 10-minute filled retention interval. The contexts were manipulated by the combination of task and place in Experiment 1, by place alone in Experiment 2, and by task alone in Experiment 3. For the manipulation of place and task, two perceptually distinctive places and two distinctive tasks (a calculation task and a fine-motor task) were used. Tasks were imposed before and after studying target items and before a free-recall test. Significant environmental-context effects were yielded in Experiment 1, but not in the other experiments. The implications of the results are discussed. PMID- 15279440 TI - [Minimally invasive treatment of common bile duct calculi]. AB - Miniinvasive treatment of lithiasis of the common bile duct hasn't reached yet a standard end point. There are multiple techniques used for it. In this study we evaluate its indications, possibilities and limits. MATERIAL AND METHOD: In a series of 14,024 biliary patients operated over 9 years 719 patients underwent open choledocolithotomy. In 173 (1.2%) of patients we used miniinvasive procedures as follows: 71 cases underwent sequential treatment, 91 laparoscopic treatment and in 11 cases the remnant calculi were extracted 1-6 days postoperative. RESULTS: Sequential treatment was the preferred treatment when the lithiasis of the common bile duct was detected preoperative. Transcistic extraction was more often performed for the lithiasis diagnosed intraoperative. The conversion to open surgery was performed in 13 cases, remnant calculi were early diagnosed in 11 patients and late diagnosed in 14 cases (the calculi were extracted by endoscopic sphyncterotomy). All patients were healed. DISCUSSIONS: The miniinvasive procedures have to be practiced as frequently as possible because of rapid healing and early recovery of the patients, despite some disadvantage of them. The choledocotomy and choledocoduodenostomy are exceptional techniques to be used in specific cases. PMID- 15279441 TI - Differentiated approach to surgical treatment of patients with perforated duodenal ulcer. AB - From 1995-2001, 264 patients with perforated duodenal ulcer were treated by the Emergency Surgical Service of Azerbaijan State Medical University in Baku. In a time of evolving standards and scientific understanding of acid peptic disease, a treatment formula was applied with excellent results. Treatment was tailored to the stage of peritonitis by time from perforation. This time was objectively evaluated by analysis of the peritoneal exudate. The value of laparoscopy was also assessed. Simple closure was performed in 94 patients (open procedure in 46, laparoscopic suture of perforated ulcer in 48 patients). Resection was performed in 170 patients (partial gastrectomy Billroth I in 118 patients, partial gastrectomy Billroth II in 18 patients and antrectomy vagotomy in 34). Application of the algorithm reduced mortality after simple closure to 6.4% and after resection to 1.2%. PMID- 15279442 TI - [Surgical treatment of hepatocellular carcinoma--indications, limitations, realities]. AB - The present study analyzes the importance of the factors in improving the resectability, obtaining morbidity and mortality rates in accordance to the actual exigencies. In The III-rd Surgical Clinic, "Sf.Spiridon" Hospital, Iasi, during 1998-2003, 24 cases of CHC (19 men, 5 women with a median age of 58.5 years), usually developed on a cirrhotic liver, benefited by surgical approach. The tumoral mass (median size 7.8 cm) was situated in the left liver (15 cases- 62.5%), right liver--13 cases and for 1 case with multiple localization (the segments VI-VII and III). For 12 cases (50%) various extensions of liver resections have been undergone: left lobectomies II-III--4 cases, left hepatectomy--1 case, segmentectomies VI--3 cases, segmentectomy III + bisegmentectomy VI-III--1 case, atypical hepatectomy--3 cases. Only 2 cases benefited by right portal vein ligation prior to resection. In 12 cases intraoperative exploration and US examination (4 cases) contraindicated the resection. One patient deceased on the entire lot (4.16%); post-resection mortality--8.33%. In conclusion, the early diagnostic of CHC developed on cirrhotic liver, the patients selection, the use of laparoscopy and intraoperative US, the available devices (CUSA dissector), selective ligation of portal branch prior to resection represents imperative elements in improving the resectability in safe conditions for the patient. PMID- 15279443 TI - [Pancreatic pseudocyst--diagnosis, evolution, surgical treatment]. AB - BACKGROUND: The pancreatic pseudocyst (PP) represents one of the most common complications of acute and chronic pancreatitis, patients with this severe pathology often undergoing surgical treatment. PATIENTS AND METHOD: This paper evaluates 133 cases that underwent surgical procedures for PP in the 3rd Surgical Clinic Cluj-Napoca during January 1993-April 2003, from a diagnostic and therapeutic point of view, emphasizing the modern imagistic methods for diagnosis and the modern surgical approach to PP. RESULTS: In 105 cases (79%) an internal drainage procedure was performed (including three cases that underwent laparoscopic drainage procedures and one case undergoing a laparoscopy assisted drainage procedure, all four cases with favorable postoperative outcome), external drainage procedures were performed in 25 cases (19%) and in three cases (2%) a pancreatic resection was performed. External drainage was performed only when internal drainage was not possible. Postoperative complications occurred only in patients with externally drained PP. DISCUSSION: The results of this study confirm that the internal drainage of PP, especially using the limb of jejunum Roux-en-Y technique (67 cases, 50%), represents the best approach to PP surgery, laparoscopic procedures being a valid option in performing an internal drainage. PMID- 15279444 TI - [Suppurative complications of acute pancreatitis]. AB - BACKGROUND/AIMS: This retrospective study investigated the management of the suppurative complication of acute pancreatitis in our clinic. MATERIALS AND METHODS: A retrospective study was made in the Surgery no. 1 Clinic of Timisoara during the period of 6 years (1996-2001) and on the subject of the acute pancreatitis with suppurative complications. RESULTS: As a result the study found a number of 16 cases of suppurative complications out of a total number of 224 acute pancreatitis (7.14%). The acute pancreatitis with suppurative complications was most frequent of biliary etiology (56.2%). All suppurations have occurred on serious necrotizing hemorrhagic acute pancreatitis background. The major elements that contributed to determining the diagnosis were: systemic toxicity, the permanent sepsis, the association of multiple organ failure, but the element that provided certainty in the diagnosis was the computed tomography (CT scan). The surgical mortality was 18.7%. CONCLUSIONS: The acute pancreatitis that evolves unfavorable under medical treatment must be operated. Necrotic tissues and purulent collections must be evacuated and drained efficiently. Open drainage through laparotomy lowered postoperative mortality. The early surgical intervention had a favorable effect on the prognostic. PMID- 15279445 TI - [Hereditary pancreatitis]. AB - We present the cases of three brothers (a woman and two men) with recurrent attacks of necrotic acute pancreatitis that were treated in our clinic. Two of them have diabetes mellitus controlled through insulin treatment. All patients have presented the first episode of acute pancreatitis around age of 35. We have observed a high level of serum triglyceride at admission, without evidence of lipid disorder. In addition we couldn't identify other causes of these episodes of acute pancreatitis (biliary stones, alcohol, trauma, drugs, lipid disorders). During the last year two members of this family presented recurrent attacks of abdominal pain without any biochemical or imaging signs of acute pancreatitis. PMID- 15279446 TI - [Late intestinal perforations after alloplastic repair of postoperative eventrations]. AB - Late intestinal perforation is the most rare and the most serious complication following the repair of abdominal parietal defects with alloplastic material. The authors have gathered in 7 years 6 cases of perforations which occurred between 1 and 10 years after using nonabsorbable synthetic mesh prostheses made of polyester (4 cases) and polypropylene (2 cases) as parietal substitution materials. The perforations were discovered either by the appearance of complete enteric fistulas after exploration of circumscribed parietal suppurations (3 cases) or during surgery of suppurations without external fistulas. All cases involved perforations of the small intestine and in only one case there was also a perforation of the transverse colon. In 4 cases, where eventration relapse co existed, the polyester prostheses were found partially or totally detached from the abdominal wall. The treatment of enteric perforation consisted of 5 segmental enterectomies (one of those a double one) followed by anastomoses and one enteroraphy. The colic perforation was excised and sutured. The mesh was totally removed in 5 cases and partially removed in 1 case (around the perforation). The postoperative evolution was simple in only 3 cases. In the other 3 cases anastomotic fistulas occurred and one of them (with reduced leakage) was treated by nonoperative management. It did not close spontaneously even after 12 months. The other 2 cases required multiple reinterventions (enterectomies, ileostomies, ileocoloanastomosis). One of these patients has been lost away after 200 days of hospitalization and multiple reinterventions. In all cases, the closure of the abdominal wall was simplificad by cutaneous suture only whereas consecutive relapsed eventrations remain to be treated by subsequent reinterventions. PMID- 15279447 TI - [Hepatic actinomycosis--pseudotumoral form]. AB - Hepatic actinomycosis is a pretty rare anatomo-clinical entity that often induces a wrong pre-operative diagnosis. By presenting a case of hepatic actinomycosis, apparently primitive and diagnosed pre-operatively as a neoplastic lesion, the authors discuss the diagnosis and treatment of this disease, mentioning that in the solid, tumoral forms, the hepatic resection is necessary. The authors consider that the post-operative addition of antibiotherapy (mega doses of penicillin) is useful for the eradication of inflammatory residual hepatic parenchyma as well as for the cicatrisation of the parietal fistulae. PMID- 15279448 TI - [Gall bladder carcinoma]. AB - The study comprises a lot of 989 cholecystectomies for gall bladder lithiasis performed in the First Surgery Clinic of the University Emergency Hospital of Bucharest throughout three years and a half (2000-June 2003). Half out of them- 493 cases (50%)--were laparoscopic interventions. All of the organ material was submitted to pathological examination. In 4 cases (0.4%), all females, the pathologist surprisingly unveiled the diagnosis of gall bladder carcinoma in the 3rd stage of evolution for all of them. None of the observations presented beforehand or per operative hints suggestive for a malignant affliction. Three patients associated multiple pathology such as acute pancreatitis (obs No2), common bile duct lithiasis (obs No3) and cholecysto-duodenal fistula (obs No4). The postoperative evolution of gall bladder neoplasia of the 4 cases studied was extremely fast, the patients soon being readmitted in our clinic for jaundice, subhepatic tumoral mass and liver metastasis, on average after 31 days after cholecystectomy. That was why the reinterventions were confined to limit at exploratory laparotomies only. PMID- 15279449 TI - [Clinical course of an ileal carcinoid with mesenteric metastasis]. AB - This paper aim is to present the case of a male patient, age 63 admitted in hospital for non-specific gastroenterologic symptoms easily attributed to colecyst calculosis; the anamnesis and careful clinical examination have avoided a useless cholecystectomy and permitted to establish indication for laparotomy; during that was found an ileal tumor with massive metastasis in the base of mesentery. The surgical attitude was dictated by histological diagnostic difficulties (prostate adenocarcinoma metastasis), so it has given up the idea of tumor excision (unjustified intraoperative risk for a metastasis), performing a bypass of the ileal tumor by an enteroenterostomy, then the patient was sent to the Oncological Department. Postoperative evaluation of the patient finally permitted to establish the neuroendocrine origin of the tumor; after oncological treatment (chemotherapy), about 10 months after operation the patient didn't present any sign of tumor (tumor markers and CT scan within normal). The paper emphasizes the problem of practicing an aggressive surgical treatment for carcinoid intestinal tumors with mesenteric metastasis but also brings attention to nonsymptomatic forms of cholecyst calculosis that may hide associated malignant tumors undetected by laparoscopic cholecystectomy. PMID- 15279450 TI - [Carcinoid tumors, leiomyosarcoma and invasive adenocarcinoma metastasis of the Treitz angle]. AB - Our paper studies the quite rare occurrence of the digestive superior occlusive pathology, namely some causes which might induce partial or even total obstruction of the digestive tract, at the distal anatomical limits of the duodenum, also known as the Treitz angle. The first two described cases were carcinoid tumors, obstructive and ulcerated in the lumen of the same angle. The last two cases were an obstructive leiomyosarcoma and an invasive mesenteric metastasis from a right colon cancer, which cause a total external compression of the Treitz angle, clinically manifested as a complete food intolerance, as a first symptom. Concerning the clinical evolution, these are totally different lesions, malign, metastasis and neuroendocrine tumors, which occurred at the same level, had a totally different clinical evolution and surgical approach, only three of them developing at good postoperative course. The imaging, clinical and pathological diagnosis problems, the different specific surgical solutions, the postoperative care and finally the rarity of this level of obstruction of the small bowel, are the aim of this paper. PMID- 15279451 TI - [Frey operation--valuable alternative in the surgical treatment of chronic pancreatitis]. AB - The "golden standard" of the surgical treatment of chronic pancreatitis with an inflammatory mass in the head of the pancreas seems to be the duodenum preserving resection of the head of the pancreas as described by Beger. However, in some cases, the inflammatory process may induce an encasement of the retropancreatic intestinal vessels making the dissection of the portal vein very difficult. The local resection of the head of the pancreas combined with longitudinal pancreaticojejunostomy (Frey operation) was developed in order to provide a simple and less time consuming procedure, that avoids the dissection of the portal vein and is especially indicated in cases with severe inflammatory and edematous alterations of the head of the pancreas at this level and with dilated pancreatic duct. Two patients with chronic pancreatitis with severe pain, addiction to analgesics and weight loss underwent a Frey procedure. In both patients an inflammatory mass in the head of the pancreas and dilated pancreatic duct were demonstrated. The freeing of the head of the pancreas from the portal vein was not possible because of the intense inflammatory process. The local resection of the pancreatic head and the longitudinal pancreatico-jejunostomy was successfully performed. There were no postoperative mortality or morbidity and the short and long term results (pain relief and nutritional status) are excellent. PMID- 15279452 TI - Percentage of coronary anomalies in a population of patients undergoing coronary angiography: a retrospective study. AB - Coronary artery anomalies, although less frequent than congenital anomalies of the heart chambers and valve morphology, should be considered in a wide range of ages, in both sexes and as a possible etiology in myocardial ischemia, infarction, and sudden death, as well as in the planning of heart surgery for coronary revascularization, correction of congenital heart malformations or valve replacement. Between January 1996 and June 2002 we reviewed our catheterization database and carried out a retrospective study of the 3660 angiographies performed in our cardiology department. The patients were referred for positive ischemic test, acute coronary syndrome and/or valvular heart disease. From the 3660 angiographies we identified 25 patients (0.68%) with coronary artery anomalies and report the prevalence and types of these anomalies in the population studied. We also assessed the presence of coronary artery disease. PMID- 15279453 TI - PAMI risk score for mortality prediction in acute myocardial indarction treated with primary angioplasty. AB - OBJECTIVE: Based on the PAMI 1 and 2, AIR PAMI, and STENT PAMI trials, a risk score to predict mortality in patients undergoing primary angioplasty was recently proposed--the PAMI risk score. It includes only 6 parameters. As one of the first tools available to predict mortality in this group of patients, it results from controlled trials, with restricted inclusion criteria. It was our objective to evaluate how the PAMI risk score applies to "real world" patients. METHODS: 149 patients (mean age 58.2 +/- 13.6 years, 113 male) undergoing primary angioplasty were included. The PAMI risk score was applied and the patients were divided in 3 groups: 0 to 2 points (group A), 3 to 6 points (group B) and > or =7 points (group C). RESULTS: Sixty-eight patients (46%) were included in group A, 41 (27%) in group B and 40 (27%) in group C. There were no significant differences in pain-to-balloon times between the 3 groups. Immediate mortality (0%, 2.4% and 15%: p = 0.001), in-hospital mortality (2.9%, 7.3% and 37.5%; p < 0.001), 30-day mortality (2.9%, 7.3% and 37.5%; p < 0.001) and 6-month mortality (4.4%, 14.6% and 45%; p < 0.001) were significantly different between the 3 groups. CONCLUSIONS: The PAMI risk score is a simple prognostic tool, with parameters that can be easily acquired, enabling reliable prediction of immediate, in-hospital, 30-day and 6-month mortality in patients with acute myocardial infarction treated with primary angioplasty. PMID- 15279454 TI - Acute coronary syndromes in smokers: clinical and angiographic characteristics. AB - Smoking is a major and reversible risk factor for coronary artery disease. The present work aims to define the risk factors, angiographic and clinical characteristics and evolution of acute coronary syndromes in smokers. METHODS: We studied 521 consecutive patients with acute coronary syndrome admitted to the intensive care unit who underwent catheterization. We assessed the population in terms of risk factors, pathology (unstable angina or acute myocardial infarction), coronary morphology, left ventricular function, the need for intervention, evolution and complications over a one-year period. The characteristics of smokers were then compared with those of non-smokers. RESULTS: Of the 521 patients with acute coronary syndrome (391 men), 182 (35 %) were smokers. The smokers were younger than the non-smokers (56.3+/-9.5 versus 66.4 +/ 7.8; p < 0.001), were more frequently male (91 versus 66%; p < 0.001), and presented more risk factors (43% with 3 or more risk factors versus 17% in non smokers; p < 0.001), more obesity (11 versus 5%; p < 0.01), and less diabetes (19 versus 37%; p < 0.001). Smokers presented greater prevalence of acute myocardial infarction (57 versus 40%; p < 0.001) and less unstable angina. Coronary morphology was not significantly different in smokers compared to non- smokers and left ventricular function after the aculte coronary syndrome was similar in both groups. Smokers less frequently underwent surgery during hospitalization (22% versus 35%; p < 0.01) but needed angioplasty as often as non-smokers (48% versus 16%; NS). Smokers presented more frequent complications (angina, heart failure, re-infarction or CABG) than non-smokers (26% versus 17%; p < 0.01), during the first year of follow-up. One-year mortality was similar in both groups. The results were not significantly different when adjusted for gender. CONCLUSIONS: On average, acute coronary syndrome occurred 10 years earlier in smokers than in non-smokers. The former generally presented more risk factors, lower prevalence of diabetes and higher of obesity, more myocardial infarctions and less unstable angina. After the acute coronary syndrome, at one year, smokers presented more complications than non-smokers but had similar mortality. PMID- 15279455 TI - Transcatheter device occlusion of atrial septal defects and patent foramen ovale under intracardiac echocardiographic guidance. AB - BACKGROUND: Transcatheter device occlusion of atrial septal defects is becoming an effective alternative to surgery. In order to avoid the discomfort of transesophageal echocardiography and the need for general anesthesia we started using intracardiac ultrasound (ICUS) to guide device implantation in adult patients. OBJECTIVE: To assess the efficacy and safety of transcatheter closure of atrial septal defect (ASD) and patent foramen ovale (PFO) assisted by ICUS. METHODS: Between April 2001 and November 2002, 27 patients with ASD and four patients with PFO were selected for transcatheter device occlusion. Patient selection was based on previous transesophageal echocardiogram and clinical data. All patients were treated without anesthesia or sedation. The percutaneous occlusion technique was assisted by ICUS and fluoroscopy. Morphological aspects of the defect and measurement of its largest diameter were used to select device diameter. RESULTS: Thirty-one patients were treated (27 with ASD, four with PFO); 21 (68%) were female, mean age was 40.1+/-13.5 years, and mean ASD diameter was 20.1+/-6.4 mm. We implanted ASD Amplatzer occluding devices in all ASD patients, the size being selected by adding 4 to 6 mm to the maximum ASD diameter. The patients with PFO had an Amplatzer PFO occluder. Mean total procedure time was 50.1+/-18.9 minutes and mean fluoroscopy time was 11.7+/-5.9 minutes. All devices were successfully deployed. We had neither complications nor mortality. Follow-up evaluation was performed by transthoracic echocardiography at 24 hours, one, six and 12 months after occlusion. During a mean follow-up period of 8.5+/-7.1 months, all patients were free of symptoms and on transthoracic echocardiographic evaluation at 24 hours all but three patients had complete occlusion (three ASD patients had insignificant residual shunt), with 100% occlusion rate at six months. CONCLUSIONS: Transcatheter occlusion of atrial septal defects can be effectively and safely performed using ICUS to guide device implantation in selected adult patients. PMID- 15279456 TI - Viridans streptococcus endocarditis associated with spondylodiscitis. AB - The authors report a case of a 78-year-old male, admitted to the Hospital with fever, lumbar pain and a systolic murmur. Viridans streptococcus endocarditis associated with spondylodiscitis was diagnosed. Images and results of the exams are presented. This case is compared with similar studies in the literature. PMID- 15279457 TI - Right atrial rupture. AB - Cardiac rupture is a rare complication of primary cardiac tumors. We present a very uncommon case of cardiac angiosarcoma presenting as a right atrial rupture. A 58-year-old patient presented progressive dyspnea, weight loss, anorexia and asthenia. A chest X-ray revealed global cardiac enlargement, mainly from the right atrium. Transthoracic echocardiography gave rise to the suspicion of right atrial pseudoaneurysm, which was confirmed by transesophageal echocardiography. The ruptured atrial wall was clearly seen using contrast echo. In addition, magnetic resonance imaging provided an excellent tool in the anatomic evaluation of this uncommon complication. The diagnosis of angiosarcoma was hystologically confirmed. PMID- 15279458 TI - Randomization methods in clinical trials. AB - In recent times there has been an enormous growth in the number of randomized controlled trials (RCTs). Their importance for clinical practice is inarguable, so it is essential for the practitioner to acquire a basic knowledge of their methodology, in order to accurately evaluate this type of study. Randomization is a central concept, since it constitutes the basis for production of results that are valid, important and applicable. In this paper we set out to show the importance of randomization by explaining its basic concepts, its practical use, the different types available, and the best ways to put them into practice. PMID- 15279459 TI - Feeding difficulties with a cardiovascular etiology. PMID- 15279460 TI - Introduction to research in cardiology. Evidence-based medicine--stratification of the evidence. PMID- 15279461 TI - [The weight of epidemiologic studies]. PMID- 15279462 TI - [Reflections on the profile of scientific research in Portugal]. PMID- 15279463 TI - [CILAB--a PC-based laboratory speech processor for implementation and evaluation of new stimulation strategies for cochlear implants]. AB - CILab is a computer-based versatile laboratory system for the implementation and evaluation of innovative stimulation strategies for cochlear implants. In contrast to existing laboratory systems the entire signal processing from the input signal to the creation of the data word for the implant is effected with the aid of a personal computer (PC). This permits rapid implementation of new stimulation strategies or psycho-acoustic tests. Real-time audio processing is also possible by using the CILab as a cochlear implant speech processor. The laboratory system has been employed with success for the evaluation of new strategies and numerous psycho-acoustic tests. PMID- 15279464 TI - [Comparative analyses of the reliability of automatic external defibrillators]. AB - Automatic external defibrillators are gaining increasing acceptance. Last year 6000 devices were installed in Germany. Since the average user has only limited medical knowledge, high demands have to be made on the automatic ECG diagnosis (fibrillation detection). Within the framework of this study a fully automatic test system that permits an objective comparison of the performance of the various devices available on the market was constructed. Older devices in particular do not always meet the requirements defined by international standards with regard to sensitivity and specificity. In addition, company philosophy appears to differ in terms of the preferential emphasis on sensitivity or specificity. Purchasers of such devices need take these findings into consideration. PMID- 15279465 TI - [The suitability of digital colposcopy for telematic applications]. AB - Since carcinoma of the cervix is one of the most common cancers in women, screening of the cervix has acquired considerable importance. Colposcopy is a simple diagnostic method of detecting suspicious changes at an early stage. Shortcomings of this method are its low specificity and high inter- and intra observer variability. A clinical pilot study was therefore carried out to investigate the advantages of a digital colposcopic system comprising a binocular colposcope coupled to a CCD camera and a computer. The aim of the study was to evaluate the reliability of diagnostic findings of the cervix obtained with digital colposcopy in comparison with standard binocular colposcopy, and to assess its suitability for telematic applications (teleconsultation, telediagnostics, treaching). A total of 315 patients were examined and statistically analysed. The patients were first submitted to a conventional colposcopic examination and a diagnosis was established. During the colposcopic examination camera images were stored on a computer, on the basis of which a second physician experienced in colposcopy reviewed the initial diagnosis. The primary and secondary findings of each patient were classified into 4 categories and compared following the Rome classification system. Agreement between the primary and secondary diagnosis was established in 69% of the cases (kappa = 0.60 +/- 0.03). No bias was observed in terms of under- or overrating. The percentage of non-assessable colposcopic examinations was 9.2%. Digital colposcopy is therefore suitable for reproducing diagnostic findings on the computer, given adequate digital image quality and a suitable classification model. The method has clear advantages with regard to follow-up, internal quality control of the diagnosis, and the training and further education of physicians and students. In the future, telecolposcopy may open up new opportunities in gynaecology. PMID- 15279466 TI - [Properties and degradation of a new bioresorbable bone glue]. AB - In trauma surgery gluing is an attractive method of bonding fractured bone, which is rapid and does not require the use of screws and plates. The purpose of this study was to analyze in vitro the properties of a new bioresorbable bone glue, and in vivo its structure and degradation. The newly developed bone glue is based on alkylenbis(oligolactoyl)methacrylates and employs a two-component initiator system. Starting components for synthesis are ethylene glycol, lactic acid and methacrylic acid. In vitro the solidified glue is degraded via hydrolysis of ester bonds. Degradation products are ethylene glycol, lactic acid and oligomeres of methacrylic acid. After the first week polymer pellets (MMA, HEMALA, ELAMA) showed a weight loss of 12%. From week 2-20 a linear weight loss of 1.5% per week, that is 40% after 20 weeks, was observed. The in vivo investigations of the ultrastructure of the glue revealed a transparent and homogeneous mass with large electron-tight vacuoles. Differences in structure and degradation were not observed. Degradation of glue by hydrolysis and phagocytosis, with good biocompatibility was demonstrated. PMID- 15279467 TI - [Experimental investigation on the reproducibility of ensemble-averaged electromyographic gait analysis data in the area of experimental and clinical orthopaedics]. AB - With suitable application and signal processing methods, surface electromyography is a comparatively simple instrument for investigating the temporal pattern of the muscular activity of a walking subject. The influence of changes both in the external experimental conditions (e.g. orthopedic shoe design) and in the human locomotor system (due to disease or therapy) on the individual muscular gait characteristics can be documented in this way. The usefulness of this kind of investigation is basically limited by the reproducibility of the gait analytical findings of the subject, who is examined at different times with unchanged bodily state and under identical experimental conditions unchanged. In our experiments we observed that the reproducibility of electromyographic activity curves obtained by ensemble averaging over a sufficiently high number of full strides differs for different muscles and in different subjects. Within the same experimental session it is very high and considerably better than in experiments done on different days. In examinations done on different days the basic characteristics of the activity curves are reproduced better than the absolute height of the amplitudes. In view of these findings the differences observed in the gait analysis of patients in the course of operative or conservative therapy have to be interpreted very carefully as to their true origin. PMID- 15279468 TI - [A gearing mechanism with 4 degrees of freedom for robotic applications in medicine]. AB - Applications in robot-aided surgery are currently based on modifications of manipulators used in industrial manufacturing processes. In this paper we describe novel rotatory kinematics for a manipulator, specially developed for deployment in robot-aided surgery. The construction of the gearing mechanism used for the positioning and orientation of a linkage point is described. Forward and inverse kinematics were calculated, and a constructive solution proposed. The gearing mechanism is based on two disk systems, each of which consists of two opposing rotatable discs. The construction was designed in such a way that the linkage point can be positioned freely anywhere within the mechanism's range of motion. The kinematics thus permits an x-y-positioning via rotating movements only. The spatial arrangement of two of such disc systems permits movements in four degrees of freedom (DOF). The construction is compact, but can be further miniaturized, is flexible and manufacturing costs are low. On the basis of this mechanical concept a new, small automated manipulator for surgical application will be developed. PMID- 15279469 TI - [Primary stability of cement free hip cup prostheses--morphometric comparative analysis of two cup types]. AB - An uncemented implanted acetabular cup must ensure primary stability and postoperative osteointegration. For both conditions the immediate postoperative contact between bone and the structured surface of the cup is of decisive importance. Two different types of acetabular cups manufactured by ESKA (standard model and Kapuziner model) were implanted in the innominate bones of elderly deceased persons. After plastination, thin serial sections were obtained. On the basis of X-rays of these sections, contact between bone and the reticulate surface of the cups was quantified. The quality of the contact was analysed densitometrically. PMID- 15279470 TI - [Management of vesical diverticula]. AB - Vesical diverticula refer to hernias of the vesical mucous membrane in the detrusor. The diverticulum wall is therefore constituted by the chorion urothelium. The muscular dehiscence that is at the origin of the diverticulum may be either congenital or degenerative. Two important complications of the diverticulum-that are sometimes interwoven-may occur: a draining defect (responsible for infections, lithiasis, and functional signs of the lower urinary tract), and the development of an urothelial tumour in the diverticulum cavity. For such complicated diverticula, surgery is indicated, by endoscopic or retropubic approach. Results may be excellent, provided the surgical intervention focuses at the same time on the management of the associated sub-vesical obstacle in case of acquired diverticulum. PMID- 15279471 TI - [Uneasy nephrectomies]. AB - Kidney exeresis, which is usually a simple surgical procedure, may sometimes be uneasy due to multiple difficulties related to the size of the lesion or anatomic presentation. In this chapter, such difficulties and their related complications are considered from a pragmatic angle, in order to identify some practical solutions. PMID- 15279472 TI - What does it all really mean? PMID- 15279473 TI - Pulp reaction to vital bleaching. AB - This study evaluated the histological changes in dental pulp after nightguard vital bleaching with 10% carbamide peroxide gel. Fifteen patients between 12 and 26 years of age with caries-free first premolars scheduled for orthodontic extraction were treated with 10% Opalescence (Ultradent Products, Inc). Tooth #5 had four days of bleaching, tooth #12 was treated for two weeks, tooth #21 was bleached for two weeks followed by two weeks without treatment and tooth #28, serving as the control, was without treatment. All teeth were extracted at the same time. Immediately after extraction, 4 mm of the most apical portion of the root was sectioned off and each specimen was placed in a vial containing 10% neutral buffered formalin. The samples were prepared for histological evaluation at the Scandinavian Institute of Dental Materials (NIOM) and microscopically examined independently at both NIOM and Indiana University School of Dentistry (IUSD). Pulp reactions were semi-quantitatively graded as none, slight, moderate and severe. Slight pulpal changes were detected in 16 of the 45 bleached teeth. Neither moderate nor severe reactions were observed. The findings indicate that the slight histological changes sometimes observed after bleaching tend to resolve within two weeks post-treatment. Statistical differences existed only between the untreated control and the four-day (p=0.0109) and two-week (p=0.0045) treatment groups. The findings from this study demonstrated that nightguard vital bleaching procedures using 10% carbamide peroxide might cause initial mild, localized pulp reactions. However, the minor histological changes observed did not affect the overall health of the pulp tissue and were reversible within two weeks post-treatment. Therefore, two weeks of treatment with 10% carbamide peroxide used for nightguard vital bleaching is considered safe for dental pulp. PMID- 15279474 TI - In vivo antibacterial effects of dentin primer incorporating MDPB. AB - This study examined the hypothesis that experimental primer containing the antibacterial monomer 12-methacryloyloxydodecylpyridinium bromide (MDPB), which was previously reported to show bactericidal effects in vitro, inhibits bacteria in cavities under in vivo conditions. The number of bacteria resulting from applying primer solution to cavities in dog teeth infected with Streptococcus mutans was determined. The infected cavities were also restored using primer and the pulp response was histopathologically examined after 7, 30 and 75 days. No bacteria were recovered after applying the experimental primer, although the bactericidal effects of the proprietary primer were insignificant. Restoration with the experimental primer resulted in little or no pulpal inflammation for all periods; whereas, mild to moderate inflammatory response was observed when using proprietary primer. These results indicate that the experimental primer containing MDPB could exhibit in vivo antibacterial effects, suggesting its possible clinical benefit. PMID- 15279475 TI - A randomized, controlled trial evaluating the three-year clinical effectiveness of two etch & rinse adhesives in cervical lesions. AB - A three-year randomized, controlled prospective study evaluated the clinical performance of two three-step etch & rinse adhesives (OptiBond FL, Kerr: O-FL; PermaQuick, Ultradent: PMQ) in Class V cervical erosion-abrasion lesions. The latter adhesive was also tested with two restorative composites with contrasting stiffness in order to evaluate the effect composite stiffness might have on the clinical longevity of cervical restorations. A total of 150 lesions were randomly restored in pairs of the three adhesive/composite combinations (PMQ combined with Amelogen Hybrid: PMQ/A-Hy, Ultradent; PMQ combined with Amelogen Microfill: PMQ/A Mi, Ultradent; O-FL combined with Prodigy: O-FLJPro, Kerr) per patient and evaluated at baseline, after six months, one year, two years and three years of clinical service. After three years, the retention rate was 100% for O-FL/Pro and 98% for both PMQ/A-Hy and PMQ/A-Mi, thereby, satisfying the "full acceptance" guidelines specified by the American Dental Association. A pairwise comparison showed no significant difference in adhesive performance between restorations made using the microfilled and hybrid composite for any evaluation criteria (p>0.05). PMID- 15279476 TI - Analysis of longitudinal marginal deterioration of ceramic inlays. AB - This study quantitatively and morphologically analyzed and clarified the longitudinal marginal changes of ceramic inlays and determined the mechanism for those changes. Epoxy replicas of 15 Class II ceramic inlays in permanent premolars prepared at baseline, 6, 12, 24, 48, 72 and 96 months after placement were selected. A CCD optical laser scanner was employed to measure quantitative changes in the occlusal surfaces of restored teeth. Longitudinal cross-sections of marginal areas of a ceramic inlay were computed, and two profiles of the same location obtained at different periods were superimposed using software. The area enclosed by the two profiles obtained at different periods was defined as the quantitative marginal change, and both the area and maximum depth in the area enclosed were calculated with picture analysis software. The marginal deterioration pattern was analyzed by drawing a longitudinal curve of quantitative change for each restoration. Morphological observation was carried out by scanning electron microscopy at magnifications from 20x to 75x. Quantitative measurement and morphological observation identified a sequential three-stage pattern of marginal deterioration; initial rapid progress of wear of resin composite cement in the first stage, followed by a second stage without any remarkable visible change, then rapid progression of microfractures of ceramics and/or enamel in the third stage. Boundaries between the first and second stage were found in the six and 21-month period, and those between the second and third stage at 72 months. It was concluded that longitudinal marginal deterioration of fired ceramic inlays progressed in a sequential three-stage pattern. PMID- 15279477 TI - Repair or replacement of amalgam restorations: decisions at a USA and a UK dental school. AB - Whereas replacement of failed restorations is the major treatment for adults in dental practice, repair is an important alternative with the potential to save tooth structure and increase the longevity of restorations at a lower cost. This in vitro study recorded the choices of treatment for the same set of teeth with defective Class II amalgam restorations by students and faculty at two dental schools (University of Manchester, UK and University of Florida, USA). Treatment options (monitor, refurbish, repair and replace) and reason(s) for the choice of treatment for 24 marked amalgam restorations were selected. Overall, participants more frequently chose replacement of restorations; whereas, repair was the least favored option. The reasons cited the most to replace restorations were secondary caries including unsightly appearance, partially lost restoration and tooth fracture; for repair, the major reasons included loss of part of the restoration and marginal ditching; and for refurbishment, the major reasons included poor anatomic form and marginal ditching. There was a significant difference between the students and faculties at the two sites in their choice of treatment (p<0.0001; Chi-square test). The treatment decision to "monitor" the restorations was more frequent for the Manchester site than the Florida site. Conversely, the combined treatment decisions to "refurbish, repair and replace" were more frequently chosen in Florida than in Manchester. PMID- 15279478 TI - The effect of home bleaching agents on the surface roughness of tooth-colored restoratives with time. AB - This study evaluated the effects of home bleaching agents on the surface roughness of composite restoratives. Two home bleaching gels (10% and 15% carbamide peroxide, Opalescence) and five different tooth-colored restorative materials from the same manufacturer (3M-ESPE) were selected. They included microfill (Filtek A110 [FO]), flowable (Filtek Flow [FF]), polyacid-acid modified (F2000 [FT]) and minifill (Z100 [ZO]; Filtek Z250 [ZT]) composites. Thirty-six specimens of each material were fabricated, randomly divided into three groups (n=12) and treated as follows: Group 1-Stored in distilled water, Group 2 Bleached with 10% carbamide peroxide (CP) eight hours/day; Group 3-Bleached with 15% CP eight hours/day. All treatment was conducted at 37 degrees C and fresh gel applied and rinsed off daily for eight weeks. For the bleached groups, the specimens were stored in distilled water at 37 degrees C during the hiatus periods. All the specimens were subjected to roughness testing (Ra) at weeks 0, 1, 2, 4, 6 and 8 using a profilometer. The results were analyzed using general linear model with Scheffe's post-hoc tests at significance level 0.05. The results showed that the effect of bleaching on surface roughness was material and time dependent. ZT was not affected by bleaching treatment, while FT was significantly roughened after one week of bleaching with 15% CP compared to the control group. FO, FF and ZO were not significantly roughened until eight weeks of bleaching. Repolishing or replacement of tooth-colored restorations may be required after bleaching procedures. PMID- 15279479 TI - Bending resistance of prefabricated titanium posts following molten cast core attachment. AB - Posts and cores are used to restore endodontically treated teeth that have substantial loss of the coronal tooth structure. This in vitro study was designed to determine the mechanical properties of prefabricated titanium posts following attachment of their metal cores by molten casting (caston). Prefabricated tapered titanium posts (ER post-restoring system, Komet, Lemgo, Germany) in three diameter sizes (ISO 50, 90, 110) (n=9) were cast over with the metal cores of three different alloys (Au-Ag-Pt, Au-Pt-Pd, Co-Cr-Mo). Also, posts of each size were precision fit into the central core channels of the different cast metal cores to serve as control specimens. The 0.2% yield strengths (R0.2) of all specimens were tested on a universal testing machine. Statistical analyses of the results were carried out with an analysis of variance (ANOVA, one-way, two-way) and Bonferroni-Dunn's multiple comparisons post-hoc analysis for test groups (alpha=0.05). There was a significant decrease in yield strength (p<0.05) as a result of casting the various metals over the different post sizes, considered to be due to the detrimental thickening and porosity formation of the titanium surface oxide layer. Twenty-one percent, 51% and 33% reduction in yield strength, respectively, was obtained for the ISO 50, ISO 90 and ISO 110 cast-on groups relative to controls (p<0.05). Statistically significant differences in various core alloys were found only for the Au-Ag-Pt alloy compared to the Co-Cr-Mo alloy (post size ISO 50) and the Au-Pt-Pd alloy compared to the Co-Cr-Mo alloy (post size ISO 110) (p>0.05). Prefabricated titanium posts with metal cores cast over them showed inferior mechanical properties compared to precision-fit posts. These results indicate greater strength of the titanium posts when their cast cores were attached mechanically rather than by the molten casting method. PMID- 15279480 TI - Influence of curing lights and modes on cross-link density of dental composites. AB - This study investigated the influence of curing lights and modes on the cross link density of dental composites. Four LED/halogen curing lights (LED-Elipar Freelight [FL], 3M-ESPE and GC e-light [EL], GC; high intensity halogen-Elipar Trilight [TL], 3M-ESPE; very high intensity halogen-Astralis 10 [AS], Ivoclar Vivadent) were selected for this study. Pulse (EL1), continuous (FL1, EL2, TL1), turbo (EL3, AS) and soft-start (FL2, EL4, TL2) curing modes of the various lights were examined. A conventional, continuous cure halogen light (Max [MX], Dentsply Caulk) was used for comparison. Six composite (Z100, 3M-ESPE) specimens were made for each light-curing mode combination. After polymerization, the specimens were stored in air at 37 degrees C for 24 hours and subjected to hardness testing using a digital microhardness tester (load=500 g; dwell time=15 seconds). The specimens were then placed in 75% ethanol-water solution at 37 degrees C for 24 hours and post-conditioning hardness was determined. Mean hardness (HK)/change in hardness (deltaHK) was computed and the data subjected to analysis using one-way ANOVA/Scheffe's test and Independent Samples t-test (p<0.05). Softening upon storage in ethanol (deltaHK) was used as a relative indication of cross-link density. Specimens polymerized with AS, TL2 and all modes of both LED lights were significantly more susceptible to softening in ethanol than specimens cured with MX. No significant difference in cross-link density was observed among the various modes of EL and FL. For TL, curing with continuous mode resulted in specimens with significantly higher cross-link density than curing with the soft start mode. PMID- 15279481 TI - Effects of multiple adhesive coatings on dentin bonding. AB - Simple changes to bonding techniques can improve resin-dentin bond strengths. This study evaluated the effect of multiple consecutive coatings of adhesive resin on dentin by measuring both microtensile bond strength and nano-leakage following exposure to ammoniacal silver nitrate. Resin-dentin bonded specimens were prepared using two total-etch adhesives (OptiBond Solo Plus/Kerr or Single Bond/3M ESPE). During bonding, resin application and air evaporation were done 1, 2, 3, 4, 6 or 8 times on acid-etched, moist dentin surfaces. Mean microtensile bond strengths were evaluated by two-way ANOVA and Fisher's PLSD test (p<0.05; n=16 for each group). Additionally, nanoleakage of silver nitrate was evaluated by transmission electron microscopy (TEM). The results indicated that bond strengths increased with each coating up to four coats. Nanoleakage decreased with each coat, becoming very small after four or more coats. This adhesive application method can be easily applied to clinical practice, thereby improving the quality of resin-dentin bonds. PMID- 15279482 TI - Composite bond strength to enamel with self-etching primers. AB - This study compared the shear bond strength (SBS) to enamel of five self-etching primer/adhesive systems and one total-etch, one-bottle adhesive system. Sixty freshly extracted bovine incisors were mounted, polished to 600-grit and randomly assigned to six groups (n=10): Adper Prompt Self-Etch (AD), OptiBond Solo Plus Self-Etch (OP), AdheSE (AS), Tyrian (TY) and Clearfil SE Bond (SE) as self etching systems; and Single Bond (SB) as a total-etch system (control). The respective hybrid composite was applied in a #5 gelatin capsule and light-cured. After 500 thermal cycles (5 degrees C-55 degrees C), the specimens were loaded in shear using an Instron at 5 mm/minute. Mean bond strengths were analyzed with one way ANOVA, followed by a Duncan's post-hoc test (p 0.05). SBS (mean +/- SD) were: AD=13.0(+/- 2.5); OP=5.6(+/-2.3);AS=12.6(+/-3.7); TY=7.6(+/-2.6); SE=17.6(+/-4.5) and SB=17.9(+/-4.4). ANOVA showed a significant difference at p<0.0001. Duncan's post-hoc test ranked this difference in three homogeneous subsets. Only SE showed similar enamel SBS compared to the total-etch system tested (SB). AD and AS were ranked in the intermediary Duncan's subset, while TY and OP resulted in the lowest SBS. SBS to enamel with self-etching primers may depend on its specific composition. PMID- 15279483 TI - Influence of the use of Er:YAG laser for cavity preparation and surface treatment in microleakage of resin-modified glass ionomer restorations. AB - This study quantitatively assessed the amount of microleakage on Class V cavities prepared by Er:YAG laser and high-speed handpiece, varying the surface treatment and restoring with a resin-modified glass ionomer cement. Fifty cavities were prepared using either an Er:YAG laser device or a carbide bur at high speed. The surface treatment was performed as follows: Er:YAG laser irradiation (G1); 40% polyacrylic acid (G2); laser + acid (G3); finishing with low speed + laser + acid (G4); conventional bur preparation + acid (G5-control). The samples were restored with Fuji II LC, thermocycled, isolated and immersed in a 50% AgNO3 solution. The restorations were serially sectioned and the extent of dye penetration was measured in milimeters using specific computer software. Data were analyzed by two-way ANOVA and Tukey test. The lowest degree of microleakage was observed for G5, which was statistically similar (p>0.05) to G4 but different (p<0.05) from all the other experiental groups. Lesser microleakage was observed at the occlusal margins than at the cervical margins (p<0.05). It may be concluded that the use of Er:YAG laser for cavity preparation and surface treatment negatively affected the marginal sealing of resin-modified glass ionomer restorations. PMID- 15279484 TI - The influence of salivary contamination on shear bond strength of dentin adhesive systems. AB - This study evaluated the influence of salivary contamination during dentin bonding procedures on shear bond strength and investigated the effect of contaminant-removing treatments on the recovery of bond strength for two dentin bonding agents. One hundred and ten human molars were embedded in cylindrical molds with self-curing acrylic resin. The occlusal dentin surface was exposed by wet grinding with #800 silicon carbide abrasive paper. The teeth were divided into five groups for One-step (OS) (BISCO, Inc) and six groups for Clearfil SE Bond (SE) (Kuraray Co, Ltd, Osaka, Japan). For One-step, the grinding surface was treated with 32% phosphoric acid; BAC (BISCO Inc) and divided into five groups: OS control group (uncontaminated), OS I (salivary contamination, blot dried), OS II (salivary contamination, completely dried), OS III (salivary contamination, wash and blot dried) and OS IV (salivary contamination, re-etching for 10 seconds, wash and blot dried). For SE bond, the following surface treatments were done: SE control group (primer applied to the fresh dentin surface), SE I (after salivary contamination, primer applied), SE II (primer, salivary contamination, dried), SE III (primer, salivary contamination, wash and dried), SE IV (after procedure of SE II, re-application of primer) and SE V (after procedure of SE III, re-application of primer). Each bonding agent was applied and light cured for 10 seconds. Clearfil AP-X (Kuraray Co, Ltd) composite was packed into the Ultradent mount jig mold and light cured for 40 seconds. The bonded specimens were stored for 24 hours in a 37 degrees C waterbath. The shear bond strengths were measured using an Instron testing machine (Model 4202, Instron Corp). The data for each group were subjected to one-way ANOVA followed by the Newman-Keuls test to make comparisons among the groups. The results were as follows: In the One-step groups, the OS II group showed statistically significant lower shear bond strength than the OS control, I, III and IV (p<0.05). In the Clearfil SE Bond groups, the SE II and SE III groups had decreased shear bond strength compared with the control and SE I, SE IV and SE V groups (p<0.05). In conclusion, when using One-step total etch adhesive and when the etched surface is contaminated by saliva, blotting the surface and applying the primer can recover the bond strength. Complete drying of the salivary contaminated surface should be avoided. In the Clearfil SE Bond groups, the re-priming treatment (SE IV and SE V) resulted in the recovery of shear bond strength in the specimens contaminated after priming. PMID- 15279485 TI - In vitro microleakage of four tracers with multiple applications to the same tooth. AB - Microleakage testing continues to be undertaken using a variety of techniques and methodologies. This study compared four microleakage tracers to determine if a difference exists in their ability to demonstrate microleakage on a single dental amalgam restorative material by testing in two phases. Class V amalgam restorations were placed on the facial surfaces of 105 extracted human premolars with all margins in enamel. The teeth were stored at 37 degrees C in water for two weeks except during thermocycling for 2500 cycles between 8 degrees C and 48 degrees C. The teeth were prepared for microleakage testing by sealing the external surfaces with nail polish and tinfoil, leaving the restoration and surrounding 1 mm exposed. In the first phase, four groups of 15 teeth were randomly assigned to 0.5% basic fuchsin dye, 2.0% fluorescent dye, 1.5% reactive orange 14 and 45Ca. In the second phase, another three groups of 15 teeth were immersed in 45Ca, then immersed in one of the remaining three tracers. Ridit analysis and Newman-Keuls multiple comparisons were used to compare the groups at a significance level of 0.05. The results indicate that there are differences in observed microleakage between tracers and there is no statistical influence on dye tracers by initial immersion in 45Ca. PMID- 15279486 TI - Elution of leachable components from composites after LED and halogen light irradiation. AB - This study investigated the influence of curing lights and modes on the elution of leachable components from dental composites. Four LED/halogen curing lights (LED-Elipar Freelight [FL], 3M-ESPE and GC e-light [EL], GC; high intensity halogen-Elipar Trilight [TL], 3M-ESPE; very high intensity halogen-Astralis 10 [AS], Ivoclar Vivadent) were selected for this study. Pulse (EL1), continuous (FL1, EL2, TL1), turbo (EL3, AS) and soft-start (FL2, EL4, TL2) curing modes of the various lights were examined. A conventional continuous cure halogen light (Max [MX], Dentsply-Caulk) was used for comparison. Three composite (Z100, 3M ESPE) specimens (6.5 mm in diameter and 1-mm thick) were made for each curing light-mode combination. After polymerization, the specimens were stored in air at 37 degrees C for 24 hours and incubated in acetonitrile at 37 degrees C for 24 hours. BisGMA and TEGDMA extracts were isolated by high performance liquid chromatography (HPLC). Data were subjected to analysis using one-way ANOVA/Scheffe's post-hoc test and Independent Samples t-test at significance level 0.05. The total monomer (BisGMA and TEGDMA) eluted ranged from 8.75 to 27.97 ppm for FL1 and AS, respectively. Significantly more unreacted monomers were leached from composites cured with all modes of EL and AS when compared to MX. No significant difference in the total monomer eluted was observed between the two modes of FL/TL and MX Although composites cured with EL2 released significantly less monomer than EL1, 3 and 4, no significant difference in the total monomer eluted was observed between the continuous and soft-start modes of FL and TL. The elution of leachable components from composites appears to be curing light specific rather than light source (LED or halogen) and curing mode specific. PMID- 15279487 TI - The influence of dentin adhesives on the demineralization of irradiated and non irradiated human root dentin. AB - This study determined the caries-protective effects of two different dentin bonding systems (Syntac, Scotchbond) on sound and irradiated root surfaces in vitro. The root surfaces of 30 freshly extracted caries-free human molars were used. The teeth were bisected in the mesio-distal direction and all lingual halves of the teeth were irradiated. The irradiation dose of 60 Gy was fractionally applied over six weeks (2 Gy/day, 5d/wk). All halves were then coated with acid-resistant nail varnish, exposing two rectangular windows 6 mm2 each on the dentinal root surface. One window served as an untreated control, while the other was treated with one of the above mentioned dentin adhesive systems. The specimens were randomly distributed among the four experimental groups as follow: Group A: Syntac, non-irradiated; Group AR: Syntac, irradiated; Group B: Scotchbond, non-irradiated; Group BR: Scotchbond, irradiated. Subsequently, all specimens were demineralized for 14 days with acidified gel (HEC, pH 4.8, 37 degrees C). From each window, two dentinal slabs were cut. The slabs were ground to a thickness of 80 microm and submerged in water. The depth of the lesions was determined using a polarized light microscope. The non irradiated control specimens showed lesions with an average depth of 63 microm (+/-10,2 microm). In the case of the irradiated control specimen, the lesion depth was not significantly different. In all experimental groups, the lesion depth was significantly reduced compared to the control groups. Statistical analysis revealed no significant differences between the irradiated and non irradiated specimens. It can be concluded that demineralization of the root surface can be hampered by application of the dentin adhesive systems tested. In this study, no differences between irradiated and sound root surfaces could be detected. PMID- 15279488 TI - Effects of cavity configuration on composite restoration. AB - This study evaluated the effects of various cavity configurations on the bond strength, microleakage, flexural strength and elastic modulus of a hybrid (Clearfil AP-X) and a microhybrid (Esthet-X) composite restorative. After the specimens were made with C-factors of less than 1, 2.4 and 3.4, flexural strength and elastic modulus were evaluated in three-point bending using a mechanical testing machine. Fragments of the fractured specimens were selected randomly and the fracture surfaces were examined in SEM. To evaluate the microtensile bond strength and microleakage of composite restorations in bovine cavities, C-factors (ratio of bonded to non-bonded cavity surface) were controlled as 1.0, 2.3, 3.0 and 3.7. All specimens were stored in distilled water at 37 degrees C for 24 hours and tested in a universal testing machine (EZ Test, Shimadzu, Japan). For the microleakage test, teeth with restorations were stained with silver nitrate and examined by two examiners under a stereomicroscope at 40x magnification. The hybrid composite showed higher mechanical properties than the microhybrid composite. The flexural strength and elastic modulus of both composites decreased when polymerized under greater constraint, that is, with increasing C-factor. Mean microtensile bond strength to dentin was also decreased with increasing C factor for both types of composites. Microleakage scores for the hybrid composite restorations were generally higher than the microhybrid composite. PMID- 15279489 TI - Polytetrafluoroethylene (PTFE) tape as a matrix in operative dentistry. AB - A method for using PTFE tape as a matrix to prevent the etching and/or bonding of adjacent tooth structure was presented. This technique is simple, quick and inexpensive. PFTE tape in the form of plumber's tape is readily available at any hardware store. Despite manufacturers' varying thicknesses of PTFE tape, it can be stretched, yielding a matrix thinner than other matrix systems. The tape will also adhere and conform to adjacent tooth structure, offering the operator an unhindered access to perform sculpting of the restorative material. PMID- 15279490 TI - Establishing proximal contacts with pre-polymerized composite inserts. AB - A technique of restoring Class II posterior restorations with resin-based composite and a pre-polymerized composite insert was presented. This technique is intended to aid the practitioner in obtaining tight, broad, proximal contacts without having to purchase special instruments or materials. This method is not time-consuming, nor technically difficult. Another benefit of this technique is that it reduces the total amount of volumetric shrinkage of restorative material inside the cavity preparation, as the composite insert is pre-polymerized before placement. PMID- 15279491 TI - Salvaging a compromised tooth using a combination amalgam core/composite window technique. PMID- 15279492 TI - Expression levels of genes likely involved in glucose-sensing in the obese Zucker rat brain. AB - It has been suggested that certain cells in the brain, like pancreatic beta cells, use glucose transporter-2 (GLUT-2), glucokinase and glucagon-like peptide 1 receptor (GLP-1R) to sense glucose in the service of multiple aspects of energy balance. The obese Zucker rat displays numerous disturbances in energy homeostasis and may provide a model of dysfunctional expression of genes related to nutrient control systems. Using real-time RT-PCR we measured gene expression for three of the pancreatic glucose-sensing markers and neuropeptide Y (NPY) in the medial, lateral hypothalamus and hindbrain of lean and obese Zucker rats of both genders. Additionally, we measured circulating levels of glucose, leptin, insulin, corticosterone and glucagon. The results indicate that GLUT-2 mRNA expression is decreased, whereas glucokinase is increased in the hindbrain of obese rats. NPY mRNA level is significantly higher, whereas GLP-1R is significantly lower in the medial hypothalamus in obese individuals. Gender related differences were found in the hindbrain and medial hypothalamus for GLUT 2 and in the lateral hypothalamus for GLP-1R and they may be related to the fact that the female Zucker rats do not develop diabetes as readily as males. Furthermore, the hindbrain may be an important site for glucose-sensing where major phenotypic changes occur for glucose-sensing genes expression. PMID- 15279493 TI - Blueberry supplemented diet: effects on object recognition memory and nuclear factor-kappa B levels in aged rats. AB - It has been reported that an antioxidant-rich, blueberry-supplemented rat diet may retard brain aging in the rat. The present study determined whether such supplementation could prevent impaired object recognition memory and elevated levels of the oxidative stress-responsive protein, nuclear factor-kappa B (NF kappaB) in aged Fischer-344 rats. Twelve aged rats had been fed a 2% blueberry supplemented diet for 4 months prior to testing. Eleven aged rats and twelve young rats had been fed a control diet. The rats were tested for object recognition memory on the visual paired comparison task. With a 1-h delay between training and testing, aged control diet rats performed no better than chance. Young rats and aged blueberry diet rats performed similarly and significantly better than the aged control diet group. Levels of NF-kappaB in five brain regions of the above subjects were determined by western blotting assays. In four regions, aged control diet rats had significantly higher average NF-kappaB levels than young animals on the control diet. In four regions, aged blueberry diet rats had significantly lower levels of NF-kappaB than aged control diet rats. Normalized NF-kappaB levels (averaged across regions and in several individual regions) correlated negatively and significantly with the object memory scores. PMID- 15279494 TI - The effects of an essential fatty acid compound and a cholecystokinin-8 antagonist on iron deficiency induced anorexia and learning deficits. AB - Iron deficiency (ID) is among the most common nutritional diseases, causing deleterious effects that include decreases in cognitive function and weight loss. The ID also induces a reduction in the number and affinity of dopaminergic D2 receptors. The new finding that ID induces an increase in the pancreas cells, leads to the hypothesis that cholecystokinin-8 (CCK-8) is involved in the ID effects. The level of CCK-8 was higher among ID rats, compared with normal rats. The ID rats in our study were anorectic and performed poorly in learning tests (Morris water maze and passive avoidance learning). Essential fatty acids (EFA) mediate dopamine activity and have been found to rehabilitate learning deficits. Treatment with a fatty acid compound blocked both the learning deficits and the anorexia, while a CCK-8 antagonist was successful only against the anorectic effects. PMID- 15279495 TI - Effects of fish oil on the central nervous system: a new potential antidepressant? AB - In the last 100 years major depression has increased worldwide. In this study we provided coconut fat (CF, rich in saturated fatty acids) or fish oil (FO, rich in n-3 polyunsaturated fatty acids) to female rats throughout pregnancy and lactation and then to their offspring post-weaning and examined lipid brain profile and the possible effect of FO as antidepressant agent in the offspring in adulthood (F1). Rats were submitted to forced swimming test, elevated plus maze, Morris water maze and open field. Peroxidation rate in the cerebral cortex and hippocampus were measured. Docosahexaenoic acid (DHA) concentration in dam's milk, eicosapentaenoic acid (EPA) and DHA concentration in hippocampus and cerebral cortex from F1 rats FO supplemented increased significantly when compared to control (C) and CF rats. Arachidonic acid/EPA ratio in the cerebral cortex and hippocampus decreased in rats submitted to forced swimming test. Peroxidation rate were not different between the groups. Immobility time in the forced swimming test in FO group was reduced (p < 0.01) when compared to C and CF rats. We conclude that lifelong intake of FO was able to induce an antidepressant effect with EPA and DHA concentration increased in the cerebral cortex and hippocampus. PMID- 15279496 TI - Conductor compounds of phenylpentane in Mycoleptodonoides aitchisonii mycelium enhance the release of dopamine from rat brain striatum slices. AB - Monoterpene compound is a major component of essential oils in various aromatic species. Previous reports about the monoterpene compound linalool and its effect on the brain neurotransmitters glutamic acid, GABA and acetylcholine, but not catecholamines, have been reported. In this study, we investigated the effect of linalool or conductor compounds of phenylpentane, including 1-phenyl-3-pentanol and 1-phenyl-3-pentanone, on dopamine release using rat striatal slices. The edible mushroom Mycoleptodonoides aitchisonii belongs to the Climacodontaceae family, and its cultivate medium or mycelium contains derivatives of the fragrant conductor compound, phenylpentane. Compared to basal levels, 2.5 microg linalool increased dopamine from striatal slices 3-fold. A 4-fold increase in dopamine release resulted from 2.5 microg 1-phenyl-3-pentanol administration, while a half dose of this compound induced a 2.5-fold increase. A greater than 2-fold increase resulted with 2.5 microg 1-phenyl-3-pentanone. These data indicate that striatum has sensitivity for these fragrant compounds and different releasing effects result with differ structures. These actions may affect other neurotransmitters and influence brain function. PMID- 15279497 TI - Co-existence of anemia, vitamin A deficiency and growth retardation among children 24-84 months old in Maracaibo, Venezuela. AB - Iron deficiency anemia has been associated with alterations in child development and psychomotor function, being myelination and dopaminergic functioning especially vulnerable. Iron deficiency, at different ages, has different reversible and irreversible effects on CNS. Anemia has also been related to vitamin A deficiency (VAD) and growth retardation. The aim of the present paper was to determine the coexistence of micronutrient deficiency, iron and vitamin A, and macronutrient deficiency (growth retardation). The sample consisted of 202 Venezuelan children, aged 24-84 month old, (104 girls, 98 boys); Anemia, VAD and growth retardation was evaluated by means of blood hemoglobin concentration analysis, HPLC serum retinol (values <20 microg/dl reveal VAD) and height/age and weight/age Z scores (< or = - 2 SD express stunting and underweight). Prevalence of anemia was 38.11%; VAD, 21.78%; stunting, 14.36% and underweight, 9.40%. Anemia and VAD clustered in 7.92%; anemia + stunting or + underweight coexisted in 5.94% and 2.97%, respectively. Stunting and underweight clustered with VAD in 2.97% and 1.48%. The three-way combination with anemia was only seen with stunting in 0.99% of the sample. The prevalence of micronutrient deficiencies remain as significant public health problems which should be simultaneously treated as virtually independent, giving priority to infant, toddler and preschool age groups. PMID- 15279498 TI - The effects of nicotine and sucrose on spatial memory and attention. AB - Both nicotine and sucrose can enhance performance on cognitive tasks. However, little is known about whether nicotine and sucrose could act jointly to augment mental performance. To investigate if there is an interaction between nicotine and sucrose on cognitive behavior, performance on a continuous performance task (CPT) and a spatial memory task was examined in 14 healthy smokers after they had drunk 8 oz of either a sucrose- or aspartame-containing beverage, and then chewed a piece of gum containing either 2 mg nicotine or no nicotine. To assess changes in mood as a function of nicotine and sucrose intake, the profile of mood states (POMS) test was administered three times during each test session. Participants made significantly more correct responses and significantly fewer incorrect responses on the CPT when they received nicotine than when they received the placebo gum. Closer analysis of the data revealed that there was an interaction between sucrose consumption and nicotine intake. Nicotine increased hits and decreased misses when participants were given the sucrose-containing beverage, but not when they were given the aspartame-containing beverage. Neither nicotine nor sucrose affected spatial memory or mood across experimental sessions. However, when data were analyzed for just the first session, participants who drank the sucrose-containing beverage performed significantly better on the spatial memory task than those who drank the aspartame-containing beverage. No gender differences in the effects of nicotine or sucrose on cognitive performance were detected. The results provide support that both nicotine and sucrose have positive effects on cognitive behavior, and that under some conditions the two variables have additive effects on performance. PMID- 15279499 TI - Sex differences in injecting practices and hepatitis C: a systematic review of the literature. PMID- 15279500 TI - Fifteen best practice recommendations for bar-code medication administration in the Veterans Health Administration. AB - BACKGROUND: Since 2000, the Veterans Health Administration (VHA) has pioneered the development and deployment of a bar-code medication administration (BCMA) system. Based on VHA experience, 15 "best practices" for BCMA implementation, integration, and maintenance are recommended. METHODS: Data were collected on potential barriers to the effectiveness of BCMA to improve patient safety by direct observation of medication administration, simulated BCMA use in a laboratory setting, a survey of nursing informatics specialists regarding policies and procedures, and 30 unstructured interviews with diverse stakeholders. RECOMMENDATIONS: Fifteen practices were proposed, categorized by implementation and continuous improvement, training, troubleshooting, contingency planning, equipment maintenance, medication administration, and maintenance of paper patient wristbands. For example, Recommendation 15 ("Periodic replacement of wristbands") advises weekly bar-coded wristband replacement in long term care settings to improve the scanning reliability. DISCUSSION: Lessons learned about best practices to address challenges may offer insight to others considering implementation of bar-code technology. PMID- 15279501 TI - Using intranet-based order sets to standardize clinical care and prepare for computerized physician order entry. AB - BACKGROUND: The high cost of computerized physician order entry (CPOE) and physician resistance to standardized care have delayed implementation. An intranet-based order set system can provide some of CPOE's benefits and offer opportunities to acculturate physicians toward standardized care. INTRANET CLINICIAN ORDER FORMS (COF): The COF system at the Medical University of South Carolina (MUSC) allows caregivers to enter and print orders through the intranet at points of care and to access decision support resources. RESULTS: Work on COF began in March 2000 with transfer of 25 MUSC paper-based order set forms to an intranet site. Physician groups developed additional order sets, which number more than 200. Web traffic increased progressively during a 24-month period, peaking at more than 6,400 hits per month to COF. Decision support tools improved compliance with Centers for Medicare & Medicaid Services core indicators. DISCUSSION: Clinicians demonstrated a willingness to develop and use order sets and decision support tools posted on the COF site. COF provides a low-cost method for preparing caregivers and institutions to adopt CPOE and standardization of care. The educational resources, relevant links to external resources, and communication alerts will all link to CPOE, thereby providing a head start in CPOE implementation. PMID- 15279502 TI - Extending the nurse practitioner concurrent intervention model to community acquired pneumonia and chronic obstructive pulmonary disease. AB - BACKGROUND: A Nurse Practitioner (NP) Concurrent Intervention Model shown effective for controlling telemetry usage was extended to patients with community acquired pneumonia (CAP) and patients with chronic obstructive pulmonary disease (COPD). METHODS: In spring 2000, investigators at Hackensack University Medical Center and the University of Medicine and Dentistry of New Jersey-New Jersey Medical School began an intervention to increase compliance with the Centers for Medicare & Medicaid Services (CMS) performance measures for CAP. Cost-reduction efforts were introduced by using previously described criteria for switching from intravenous to oral medication and for hospital discharge. RESULTS: Use of the NP intervention model for patients admitted with CAP and for COPD patients resulted in significant reductions in length of stay and cost savings. DISCUSSION: Concurrent intervention by a nurse practitioner can help achieve excellent compliance with performance measures for CAP and be applied to other chronic respiratory diseases such as COPD. PMID- 15279503 TI - Physician-perceived barriers to adopting a critical pathway for unity-acquired pneumonia. AB - BACKGROUND: A proven efficacious and evidence-based critical pathway for community-acquired pneumonia (CAP) was implemented in six hospitals across a health service region (Edmonton, Canada). After one year (November 2000-November 2001), the pathway had reduced average length of stay by 1 day (from 10.8 to 9.8 days, p < .001). However, great variation was observed in physician adherence to the pathway. METHODS: Physician-perceived barriers to adoption of the CAP pathway were identified through in-depth interviews. Data saturation was reached after 10 physicians, representing a convenience sample of those willing to participate, were interviewed. RESULTS: Self-reported adherence to the CAP pathway was 75% (range 50%-100%). Qualitative analysis of the interview data indicated that comments could be grouped into five themes: (1) limited applicability, (2) lack of flexibility to accommodate atypical clinical presentations, (3) perception of insufficient evidence to support recommendations, (4) local organizational barriers, and (5) need for local adaptation. For example, one physician remarked that his community hospital had insufficient staff to support collection of lab samples for all patients. DISCUSSION: Interventions to increase pathway adoption and further improve quality of CAP care should address the identified barriers. For example, local audit and feedback of outcomes data to persuade physicians of the benefits of CAP pathways will need to be instituted. PMID- 15279504 TI - The North Carolina experience with the diabetes health disparities collaboratives. AB - BACKGROUND: The Bureau of Primary Health Care (BPHC) adopted a collaborative approach that used the Chronic Care Model and quality improvement methods. The North Carolina Diabetes Prevention and Control Branch has partnered with the 12 participating community health centers since early 2000. METHODS: Team leaders of the first four centers that participated in the collaboratives were interviewed. Information obtained included previous diabetes efforts, benefits of the collaborative, success factors, and barriers to sustainability. CASE STUDY: In one of two case studies, a nonprofit community health center made Chronic Care Model-based changes to the organization of health care, clinical information systems, and delivery system design. RESULTS: Centers tracked used the electronic registry to establish a baseline, trend key process and outcome measures, and raise the standard of care. Success factors included senior leadership support, physician champions, multidisciplinary teams, and priority of collaborative activities. Barriers included staff turnover and low priority in strategic planning. Glycohemoglobin (A1C) values from aggregated reports demonstrated improvement. DISCUSSION: Useful strategies for future collaboratives may include providing provider-specific data, imparting vision to new team members, ensuring that leadership provides collaborative structure and resources, and pairing veteran and new participating sites. PMID- 15279505 TI - Righting wrong site surgery. AB - BACKGROUND: As defined by the Joint Commission on Accreditation of Healthcare Organizations (JCAHO), wrong site surgery includes wrong side or siteof the body, wrong procedure, and wrong-patient surgeries. Although many health care organizations are implementing guidelines and procedures to decrease the occurrence of wrong site surgery, numerous barriers to their effectiveness have been identified. HUMAN FACTORS ENGINEERING (HFE) ANALYSIS: A human factors system analysis can be used to better understand how elements of a work system combine andinteract to contribute to breakdowns in the system. A case study of wrong site surgery in an outpatient setting illustrates how the different work systtem elements can contribute to the occurrence of a wrong site surgery. In analyzing the care process, it is particularly important to identify the transitions of care, which can be sources of patient safety problems when deficits in communication and information transfer occur (for example, miscommunication, information not transmitted on time, wrong information transmitted, misunderstanding of the information transmitted). RECOMMENDATIONS: After a wrong site surgery, conduct a root cause analysis that uses the work system model and includes a surgery care process analysis similar to the one described in the case study; collaborate with human factors engineers to learn how to apply the work system model; apply the work system model to process analysis; and optimize work systems. PMID- 15279506 TI - Risk management lessons from one hospital's IT shutdown. PMID- 15279507 TI - Photocatalytic coatings for environmental applications. AB - A series of nano- and micronparticle-grade anatase and rutile titanium dioxide pigments have been prepared with various densities of surface treatments, particle size and surface area. Their photocatalytic activites have been determined in a series of paint films by FTIR, chalking, color, gloss change and weight loss after artifical weathering. The pigments have also been examined by rapid assessment methodologies using photodielectric microwave spectroscopy, 2 propanol oxidation and hydroxyl analysis. The microwave response under light and dark cycles provides an extended timescale probe of charge-carrier dynamics in the pigments. Pigment particle size, surface area and properties clearly play an important role in dispersion and any polymer-pigment interactions. Photooxidation studies on several types of paint films show a clear demarcation between nanoparticle- and pigmentary-grade titanium dioxide, with the former being more active because of their greater degree of catalytic surface activity. The photosensitivity of titanium dioxide is considered to arise from localized sites on the crystal surface (i.e. acidic OH), and occupation of these sites by surface treatments inhibits photoreduction of the pigment by ultraviolet radiation; hence, the destructive oxidation of the binder is inhibited. Coatings containing 2-5% by weight alumina or alumina and silica are satisfactory for general-purpose paints. If greater resistance to weathering is desired, the pigments are coated more heavily to about 7-10% weight. The coating can consist of a combination of several materials, e.g. alumina, silica, zirconia, aluminum phosphates of other metals. For example, the presence of hydrous alumina particles lowers van der Waals forces between pigments particles by several orders of magnitude, decreasing particle-particle attractions. Hydrous aluminum oxide phases appear to improve dispersibility more effectively than most of the other hydroxides and oxides. Coated nanoparticles are shown to exhibit effective light stabilization in various water- and oil-based paint media in comparison with conventional organic stabilizers. Hindered piperidine stabilizers are shown to provide no additional benefits in this regard, often exhibiting strong antagonism. The use of photocatalytic titania nanoparticles in the development of self-cleaning paints and microbiological surfaces is also demonstrated in this study. In the former case, surface erosion is shown to be controlled by varying the ratio of admixture of durable pigmentary-grade rutile (heavily coated) and a catalytic grade anatase nanoparticle. For environmental applications in the development of coatings for destroying atmospheric pollutants such as nitrogen oxide gases (NO(X)), stable substrates are developed with photocatalytic nanoparticle-grade anatase. In this study, porosity of the coatings through calcium carbonate doping is shown to be crucial in the control of the effective destruction of atmospheric NO(X) gases. For the development of microbiological substrates for the destruction of harmful bacteria, effective nanoparticle anatase titania is shown to be important, with hydrated high surface area particles giving the greatest activity. PMID- 15279508 TI - Temporal specificity of extinction in autoshaping. AB - Three experiments investigated the effects of varying the conditioned stimulus (CS) duration between training and extinction. Ring doves (Streptopelia risoria) were autoshaped on a fixed CS-unconditioned stimulus (US) interval and extinguished with CS presentations that were longer, shorter, or the same as the training duration. During a subsequent test session, the training CS duration was reintroduced. Results suggest that the cessation of responding during an extinction session is controlled by generalization of excitation between the training and extinction CSs and by the number of nonreinforced CS presentations. Transfer of extinction to the training CS is controlled by the similarity between the extinction and training CSs. Extinction learning is temporally specific. PMID- 15279509 TI - Persistence of preference for a flavor presented in simultaneous compound with sucrose. AB - Rats exposed to a simultaneous compound of a flavor and sucrose subsequently exhibited a preference for the flavor over water. This preference persisted across repeated testing even though the flavor was presented in the absence of sucrose. The preference did, however, extinguish if the rats were hungry when trained or tested, or if they had been reexposed to sucrose between training and test. Though failing to extinguish the preference, presentation of the flavor outside the compound protected it from the effects of sucrose devaluation, indicating that these presentations extinguished the within-compound association between the flavor and sucrose. The authors conclude that the hedonic reaction elicited by sucrose imbues the flavor with the same hedonic properties, and these properties maintain the preference independently of the flavor-sucrose association. PMID- 15279510 TI - Temporal coding in conditioned inhibition: analysis of associative structure of inhibition. AB - Two experiments with rats as subjects were conducted to investigate the associative structure of temporal control of conditioned inhibition through posttraining manipulation of the training excitor-unconditioned stimulus (US) temporal relationship. Experiment 1 found that following simultaneous Pavlovian inhibition training (i.e., A --> US/XA-no US) in which a conditioned stimulus (CS A) was established as a delay excitor, maximal inhibition was observed on a summation test when CS X was compounded with a delay transfer CS. Furthermore, posttraining shifts in the A-US temporal relationship from delay to trace resulted in maximal inhibition of a trace transfer CS. Experiment 2 found complementary results to Experiment 1 with an A-US posttraining shift from serial to simultaneous. These results suggest that temporal control of inhibition is mediated by the training excitor-US temporal relationship. PMID- 15279511 TI - Sequential responding and planning in chimpanzees (Pan troglodytes) and rhesus macaques (Macaca mulatta). AB - Chimpanzees (Pan troglodytes) and rhesus macaques (Macaca mulatta) selected either Arabic numerals or colored squares on a computer monitor in a learned sequence. On shift trials, the locations of 2 stimuli were interchanged at some point. More errors were made when this interchange occurred for the next 2 stimuli to be selected than when the interchange was for stimuli later in the sequence. On mask trials, all remaining stimuli were occluded after the 1st selection. Performance exceeded chance levels for only 1 selection after these masks were applied. There was no difference in performance for either stimulus type (numerals or colors). The data indicated that the animals planned only the next selection during these computerized tasks as opposed to planning the entire response sequence. PMID- 15279512 TI - Temporal learning in random control procedures. AB - Experiments 1 and 2 delivered conditioned stimuli (CSs) at random times and unconditioned stimuli (USs) at either fixed (Experiment 1) or random (Experiment 2) intervals. In Experiment 3, CS duration was manipulated, and US deliveries occurred at random during the background. In all 3 experiments, the mean rate of responding (head entries into the food cup) in the background was determined by the mean US-US interval, and the mean rate during the CS was a linear combination of responding controlled by the mean US-US and mean CS onset-US intervals; the pattern of responding in time was determined by the interval distribution form (fixed or random). An event-based timing account, Packet theory, provided an explanation of the results. PMID- 15279513 TI - A role for CS-US contingency in Pavlovian conditioning. AB - Two experiments evaluated the role of conditioned stimulus-unconditioned stimulus (CS-US) contingency in appetitive Pavlovian conditioning in rats. In both experiments, some groups received a positively contingent CS signaling an increased likelihood of the US relative to the absence of the CS. These groups were compared with control treatments in which the likelihood of the US was the same in the presence and absence of the CS. A trial marker served as a trial context. Experiment 1 found contingency sensitivity. There was a reciprocal relationship between responding to the CS and the trial marker. Experiment 2 showed that this result was not stimulus or response specific. These results are consistent with associative explanations and the idea that rats are sensitive to CS-US contingency. PMID- 15279514 TI - A partial reinforcement extinction effect despite equal rates of reinforcement during Pavlovian conditioning. AB - In 4 experiments rats received appetitive Pavlovian conditioning followed by extinction. Food accompanied every trial with the conditioned stimulus (CS) for the continuously reinforced groups and only half of the trials for the partially reinforced groups. In contrast to previous experiments that have compared the effects of partial and continuous reinforcement, the rate at which food was delivered during the CS was the same for both groups. The strength of the conditioned response during extinction weakened more rapidly in the continuously than in the partially reinforced groups. The results demonstrate that the partial reinforcement extinction effect is a consequence of the nonreinforced trials with the CS, rather than the rate at which the unconditioned stimulus is delivered during the CS. PMID- 15279515 TI - The impact of sense of coherence and negative affectivity on the work stressor- strain relationship. AB - This study investigated different ways in which work stressors, sense of coherence (SOC), and negative affectivity (NA) might influence strain. Three models covering direct, moderating, and mediating effects between the variables were tested for hyperresponsitivity, causality, perception, selection, and stressor-creation mechanisms. The sample consisted of 205 hospital employees (nurses, physicians, and medical technicians). Work stressors, SOC, NA, and strain were measured and analyzed in structural equation models and with analyses of variance. Besides direct effects on strain, significant perception, selection, and stressor-creation mechanisms of SOC could be identified. After controlling for SOC and NA, work stressors remain substantial predictors of strain. PMID- 15279516 TI - Aggression and violence against home care workers. AB - This article describes the development of the Violence and Aggression in Health Care Questionnaire (VAQ) and the application of the measure in the field of home care. In a 1st sample of 361 German home care workers, the scales of the VAQ reached internal consistencies from .72 to .93. A confirmatory factor analysis gave evidence to the hypothesized factor structure. Significant correlation with indicators of psychophysical strain and health resulted in a 2nd sample of 180 home care workers. In multiple regression analysis based on a 3rd sample of 180 home care workers, verbal aggression by patients was a significant predictor of negative psychological outcomes. The relationship is completely mediated by negative emotional reactions after aggressive incidents. PMID- 15279517 TI - Burnout patterns in rehabilitation: short-term changes in job conditions, personal resources, and health. AB - This longitudinal study reports the patterning of the burnout symptoms and the changes in employees' job conditions, personal resources, and psychological health 4 months after a rehabilitation intervention. The data were gathered by means of questionnaires before and after a rehabilitation period. Four patterns were identified: not burned out (n = 55), exhausted and cynical (n = 36), burned out (n = 26), and low professional efficacy (n = 18). These patterns differed in terms of job resources, personal resources, and depression. There were both positive and negative changes detected in participants' psychological health and job resources at the follow-up. The study shows the importance of identifying different burnout patterns in order to focus rehabilitation activities more effectively. PMID- 15279518 TI - Intimate partner abuse perpetrated by employees. AB - This exploratory study examined partner abuse perpetration in the context of employment. Qualitative and quantitative data from 29 men convicted of partner abuse were gathered through focus groups and a brief survey. Men attributed absences, reduced productivity, and errors to their perpetration of abuse and described harassing their victims using employers' phones, vehicles, e-mails and by enlisting coworkers. Findings also suggest that zero-tolerance policies may be ineffective and that employers may benefit from partner abuse training. For example, participants reported that their employers offered them alcohol-abuse services instead of batterer intervention, despite the fact that substance abuse was not a factor in all of the cases. These findings need to be tested through a larger scale, empirical investigation. PMID- 15279519 TI - Repeated downsizing contact: the effects of similar and dissimilar layoff experiences on work and well-being outcomes. AB - In this longitudinal study, the authors compared 1,244 white- and blue-collar workers who reported 0, 1, or 2 contacts with layoffs; all were employees of a large manufacturing company that had engaged in several mass waves of downsizing. Consistent with a stress-vulnerability model, workers with a greater number of exposures to both direct and indirect downsizing reported significantly lower levels of job security and higher levels of role ambiguity, intent to quit, depression, and health problems. Findings did not support the idea that workers became more resilient as they encountered more layoff events. The authors found only partial evidence that the similarity or dissimilarity of the type of repeated downsizing exposure played a role in how workers reported changes in these outcome variables. PMID- 15279520 TI - Comprehensive health promotion interventions at the workplace: experiences with health circles in Germany. AB - Health circles, the central element of a comprehensive health promotion approach that has been developed in Germany in recent years, emphasize organizational and psychosocial factors while actively involving employees in the process. Through an extensive review the authors identified 11 studies, presenting the results of 81 health circles. The scientific quality of the data is limited: only 3 studies used (nonrandomized) control groups, whereas the remaining studies are based on retrospective before-and-after comparison. Nonetheless, the available data suggest that health circles are an effective tool for the improvement of physical and psychosocial working conditions and have a favorable effect on workers' health, well-being, and sickness absence. More rigorous studies are needed to confirm these results. PMID- 15279521 TI - Both smoking reduction with nicotine replacement therapy and motivational advice increase future cessation among smokers unmotivated to quit. AB - Smokers not currently interested in quitting (N = 616) were randomized to receive telephone-based (a) reduction counseling plus nicotine replacement therapy (NRT) plus brief advice to quit, (b) motivational advice plus brief advice, or (c) no treatment. More smokers in the reduction (43%) and motivational (51%) conditions made a 24-hr quit attempt over 6 months than smokers in the no-treatment condition (16%; p < or = .01), but the 2 active conditions did not differ (p > or = .05). Similarly, 18%, 23%, and 4% of each condition were abstinent (7-day point prevalence) at 6 months (p < or = .01). Results indicate smoking reduction using NRT does not undermine cessation but rather increases the likelihood of quitting to a degree similar to motivational advice. PMID- 15279522 TI - Self-perceptions of competence in children with ADHD and comparison children. AB - The self-perceptions of children with attention-deficit/hyperactivity disorder (ADHD; n = 487) were compared with those of children in a local normative comparison group (n = 287), relative to teacher- and parent-rated perceptions of their competence. Children were participants in the ongoing follow-up portion of the Multimodal Treatment Study of Children with ADHD. Children with ADHD were much more likely than comparison children to overestimate their competence relative to adult report, regardless of who was used as the criterion rater (teacher, mother, or father). Examination by comorbidity subgroups revealed that children with ADHD inflated their self-perceptions the most in domains of greatest deficit. Gender effects also are reported. PMID- 15279523 TI - Cognitive mediation of treatment change in social phobia. AB - Ninety individuals with social phobia (social anxiety disorder) participated in a randomized controlled trial and completed cognitive-behavioral group therapy, exposure group therapy without explicit cognitive interventions, or a wait-list control condition. Both treatments were superior to the wait-list group in reducing social anxiety but did not differ from one another at posttest. Changes in estimated social cost mediated treatment changes in both treatment conditions from pre- to posttest. However, only participants who received cognitive behavioral therapy showed continued improvement from posttest to 6-month follow up, which was associated with a reduction of estimated social cost from pretest to posttest. These results suggest that cognitive intervention leads to better maintenance of treatment gains, which is mediated through changes in estimated social cost. PMID- 15279524 TI - Developmental trajectories of cigarette smoking and their correlates from early adolescence to young adulthood. AB - Smoking initiation typically occurs in adolescence and increases over time into emerging adulthood. Thus adolescence and emerging adulthood compose a critical time period for prevention and intervention efforts. To inform these efforts, this study used latent growth mixture modeling to identify 6 smoking trajectories from ages 13 to 23 among 5,914 individuals: nonsmokers (28%), stable highs (6%), early increasers (10%), late increasers (10%), decreasers (6%), and triers (40%). By age 23, the trajectories merged into 2 distinct groups of low- and high frequency and their standing on age 23 outcomes reflected this grouping. Consideration of these results can help researchers identify at-risk individuals before their smoking becomes too problematic, providing an opportunity for intervention and possible prevention of nicotine dependence. PMID- 15279525 TI - Therapeutic alliance, negative mood regulation, and treatment outcome in child abuse-related posttraumatic stress disorder. AB - This study examined the related contributions of the therapeutic alliance and negative mood regulation to the outcome of a 2-phase treatment for childhood abuse-related posttraumatic stress disorder (PTSD). Phase 1 focused on stabilization and preparatory skills building, whereas Phase 2 was comprised primarily of imaginal exposure to traumatic memories. Hierarchical regression analyses indicated the strength of the therapeutic alliance established early in treatment reliably predicted improvement in PTSD symptoms at posttreatment. Furthermore, this relationship was mediated by participants' improved capacity to regulate negative mood states in the context of Phase 2 exposure therapy. In the treatment of childhood abuse-related PTSD, the therapeutic alliance and the mediating influence of emotion regulation capacity appear to have significant roles in successful outcome. PMID- 15279526 TI - Positive tertiary appraisals and posttraumatic stress disorder in U.S. male veterans of the war in Vietnam: the roles of positive affirmation, positive reformulation, and defensive denial. AB - A 70.9% majority of the U.S. male veterans in a nationwide sample appraised the impact of their service in Vietnam on their present lives as mainly positive. A substantial minority, 41.7%, judged the effects to be highly salient. With controls on level of exposure to war-zone stressors measured with data from military records, the valence and salience of these appraisals are investigated in relation to posttraumatic stress disorder (PTSD) and other indicators of wartime and postwar functioning. The results are consistent with the hypothesis that mainly positive tertiary appraisals are affirmations of successful wartime and postwar adaptation rather than defensive denials related to maladaptive outcomes. The possibility that mainly positive tertiary appraisals also contribute to successful postwar adaptation is discussed. PMID- 15279527 TI - Targeting misperceptions of descriptive drinking norms: efficacy of a computer delivered personalized normative feedback intervention. AB - The authors evaluated the efficacy of a computer-delivered personalized normative feedback intervention in reducing alcohol consumption among heavy-drinking college students. Participants included 252 students who were randomly assigned to an intervention or control group following a baseline assessment. Immediately after completing measures of reasons for drinking, perceived norms, and drinking behavior, participants in the intervention condition were provided with computerized information detailing their own drinking behavior, their perceptions of typical student drinking, and actual typical student drinking. Results indicated that normative feedback was effective in changing perceived norms and alcohol consumption at 3- and 6-month follow-up assessments. In addition, the intervention was somewhat more effective at 3-month follow-up among participants who drank more for social reasons. PMID- 15279528 TI - Disentangling the effects of safety-behavior utilization and safety-behavior availability during exposure-based treatment: a placebo-controlled trial. AB - The primary aim of the current study was to further investigate the deleterious effects of safety-seeking behaviors on fear reduction by disentangling the effects of perceived availability of threat-relevant safety behaviors during treatment versus their actual use. Participants (N=72) displaying marked claustrophobic fear were randomly assigned to 1 of 5 conditions: (a) exposure only (EO), (b) exposure with phobic safety-behavior availability (SBA), (c) exposure with safety-behavior utilization (SBU), (d) credible placebo treatment (PL), or (e) wait list (WL). High end-state functioning rates at posttreatment were as follows: EO = 94%, SBA = 45%, SBU = 44%, PL = 25%, and WL = 0%. Findings suggest that it is the perception of the availability of safety aids as opposed to their actual use that exerts a disruptive effect on fear reduction. Clinical implications are discussed. PMID- 15279529 TI - Brief treatments for cannabis dependence: findings from a randomized multisite trial. AB - This study evaluated the efficacy of 2 brief interventions for cannabis-dependent adults. A multisite randomized controlled trial compared cannabis use outcomes across 3 study conditions: (a) 2 sessions of motivational enhancement therapy (MET); (b) 9 sessions of multicomponent therapy that included MET, cognitive behavioral therapy, and case management; and (c) a delayed treatment control (DTC) condition. Participants were 450 adult marijuana smokers with a Diagnostic and Statistical Manual of Mental Disorders (4th ed.; American Psychiatric Association, 1994) diagnosis of cannabis dependence. Assessments were conducted at baseline, and at 4, 9, and 15 months postrandomization. The 9-session treatment reduced marijuana smoking and associated consequences significantly more than the 2-session treatment, which also reduced marijuana use relative to the DTC condition. Most differences between treatments were maintained over the follow-up period. Discussion focuses on the relative efficacy of these brief treatments and the clinical significance of the observed changes in marijuana use. PMID- 15279530 TI - Preventing disruptive behavior in elementary schoolchildren: impact of a universal classroom-based intervention. AB - A population-based, randomized universal classroom intervention trial for the prevention of disruptive behavior (i.e., attention-deficit/hyperactivity problems, oppositional defiant problems, and conduct problems) is described. Impact on developmental trajectories in young elementary schoolchildren was studied. Three trajectories were identified in children with high, intermediate, or low levels of problems on all 3 disruptive behaviors at baseline. The intervention had a positive impact on the development of all disruptive behavior problems in children with intermediate levels of these problems at baseline. Effect sizes of mean difference at outcome were medium or small. In children with the highest levels of disruptive behavior at baseline, a positive impact of the intervention was found for conduct problems. PMID- 15279531 TI - The Hartford study of supported employment for persons with severe mental illness. AB - The authors compared 3 approaches to vocational rehabilitation for severe mental illness (SMI): the individual placement and support (IPS) model of supported employment, a psychosocial rehabilitation (PSR) program, and standard services. Two hundred four unemployed clients (46% African American, 30% Latino) with SMI were randomly assigned to IPS, PSR, or standard services and followed for 2 years. Clients in IPS had significantly better employment outcomes than clients in PSR and standard services, including more competitive work (73.9% vs. 18.2% vs. 27.5%, respectively) and any paid work (73.9% vs. 34.8% vs. 53.6%, respectively). There were few differences in nonvocational outcomes between programs. IPS is a more effective model than PSR or standard brokered vocational services for improving employment outcomes in clients with SMI. PMID- 15279532 TI - Stages of change or changes of stage? Predicting transitions in transtheoretical model stages in relation to healthy food choice. AB - Relatively little research has examined factors that account for transitions between transtheoretical model (TTM) stages of change. The present study (N = 787) used sociodemographic, TTM, and theory of planned behavior (TPB) variables, as well as theory-driven interventions to predict changes in stage. Longitudinal analyses revealed that sociodemographic, TPB, and 1 of the interventions predicted transitions between most stages of change. In fact, only progression from the preparation stage was not predictable. However, given that this change of stage marks the transition between cognition and actual behavior, the identification of variables that bridge this gap is crucial for the development of interventions to promote stage transitions. PMID- 15279533 TI - Parent-child interaction therapy with physically abusive parents: efficacy for reducing future abuse reports. AB - A randomized trial was conducted to test the efficacy and sufficiency of parent child interaction therapy (PCIT) in preventing re-reports of physical abuse among abusive parents. Physically abusive parents (N=110) were randomly assigned to one of three intervention conditions: (a) PCIT, (b) PCIT plus individualized enhanced services, or (c) a standard community-based parenting group. Participants had multiple past child welfare reports, severe parent-to-child violence, low household income, and significant levels of depression, substance abuse, and antisocial behavior. At a median follow-up of 850 days, 19% of parents assigned to PCIT had a re-report for physical abuse compared with 49% of parents assigned to the standard community group. Additional enhanced services did not improve the efficacy of PCIT. The relative superiority of PCIT was mediated by greater reduction in negative parent-child interactions, consistent with the PCIT change model. PMID- 15279534 TI - Intergenerational transmission of depression: test of an interpersonal stress model in a community sample. AB - An interpersonal stress model of depression transmission was tested in a community sample of nearly 800 depressed and never-depressed women and their 15 year-old children. It was hypothesized that maternal depression (and depression in the maternal grandmother) contributed to chronic interpersonal stress in the mothers, affecting quality of parenting and youths' social competence. In turn, poor social functioning and interpersonal life events caused at least in part by the youths were predicted to be the proximal predictors of current depressive symptoms and diagnoses. Structural equation modeling confirmed the predicted associations among variables and the link between youth chronic and episodic interpersonal stress and depression. Additionally, the association between maternal and child depression was entirely mediated by the predicted family and interpersonal stress effects. PMID- 15279535 TI - Relative efficacy of criminological, clinical, and personality measures of future risk of offending in mentally disordered offenders: a comparative study of HCR 20, PCL:SV, and OGRS. AB - The authors compared the ability of 3 commonly used measures of risk of future offending in a sample of 315 mentally disordered offenders discharged from a medium-secure unit in the United Kingdom. The authors explored whether the same criminogenic factors that predict recidivism in the general population also predict recidivism in mentally disordered offenders. The actuarial measure, using mainly criminological variables, provided the best prediction of recidivism compared with measures based on personality or clinical information, which provided no incremental validity over the actuarial measure. The authors suggest that for maximum efficacy clinical risk should be rated at a time of active symptoms rather than at discharge when symptoms are minimal. PMID- 15279536 TI - Do assault-related variables predict response to cognitive behavioral treatment for PTSD? AB - This study examined the hypothesis that variables such as history of prior trauma, assault severity, and type of assault, previously found to be associated with natural recovery, would also predict treatment outcome. Trauma-related variables were examined as predictors of posttreatment posttraumatic stress disorder (PTSD) severity in a sample of 73 female assault victims with chronic PTSD who completed treatment in a comparative outcome study (E. B. Foa et al., 1999). Results indicated that after controlling for initial severity of PTSD symptoms, the experience of trauma in childhood and sustaining physical injury during the adult assault were predictive of greater PTSD severity following treatment. PMID- 15279537 TI - Brief family intervention effects on adolescent substance initiation: school level growth curve analyses 6 years following baseline. AB - This study examines the effects of 2 brief family-focused interventions on the trajectories of substance initiation over a period of 6 years following a baseline assessment. The 2 interventions, designed for general-population families of adolescents, were the 7-session Iowa Strengthening Families Program (ISFP) (Molgaard & Spoth, 2001) and the 5-session Preparing for the Drug Free Years Program (PDFY) (Catalano, Kosterman, Haggerty, Hawkins, & Spoth, 1999). Thirty-three rural public schools were randomly assigned to the ISFP, the PDFY, or a minimal-contact control condition. The authors evaluated the curvilinear growth observed in school-level measures of initiation using a logistic growth curve analysis. Alcohol and tobacco composite use indices--as well as lifetime use of alcohol, cigarettes, and marijuana--and lifetime drunkenness, were examined. Significant intervention-control differences were observed, indicating favorable delays in initiation in the intervention groups. PMID- 15279538 TI - Sequential treatment for child abuse-related posttraumatic stress disorder: methodological comment on Cloitre, Koenen, Cohen, and Han (2002). AB - M. Cloitre, K. Koenen, L. R. Cohen, and H. Han (2002; see record 2002-18226-001) presented results of a randomized trial that clearly demonstrate the safety and efficacy of a treatment program delivering skills training in affective and interpersonal regulation (STAIR) prior to conducting imaginal exposure (IE) to trauma memories for adults with posttraumatic stress disorder (PTSD) related to childhood abuse. In this comment the authors review the results presented by Cloitre et al and specifically compare the impact of the STAIR and IE phases of the treatment on affect regulation and psychopathology measures. Evidence for adverse events associated with exposure therapy is reviewed. The authors emphasize that the present study should not be interpreted as evidence that pretreatment with STAIR is additively helpful or necessary prior to IE for PTSD associated with child abuse and that a between-groups comparison is necessary before such conclusions can be drawn. PMID- 15279540 TI - G protein-coupled receptor fusion proteins in drug discovery. AB - A wide range of peptides and polypeptides can be appended to either the N- or C terminus of G protein-coupled receptors without disrupting substantially ligand binding and signal transduction. Following fusion of fluorescent proteins, reporter gene constructs or G protein alpha subunits to the C-terminal tail of a receptor high content and G protein activation assays can be employed to identify agonist ligands. Further modification of the receptor fusions to introduce enhanced levels of constitutive activity and to physically destabilise the protein allows antagonist/inverse agonists screens to be developed in parallel. Equivalent C-terminal addition of pairs of complementary, non-functional, polypeptide fragments allows the application of enzyme complementation techniques. Introduction of N-terminal tags to receptors has also allowed the introduction of novel assay techniques based on a pH-sensitive cyanine dye. These have the capacity to overcome certain limitations of GPCR-fluorescent protein fusions. PMID- 15279541 TI - Allosteric modulation of G protein-coupled receptors. AB - G protein-coupled receptors (GPCRs) constitute the largest receptor superfamily in the human genome and represent the most common targets of drug action. Classic agonist and antagonist ligands that act at GPCRs tend to bind to the receptor's orthosteric site, that is, the site recognized by the endogenous agonist for that receptor. However, it is now evident that GPCRs possess additional, extracellular, allosteric binding sites that can be recognized by a variety of small molecule modulator ligands. Allosteric modulators offer many advantages over classic orthosteric ligands as therapeutic agents, including the potential for greater GPCR-subtype selectivity and safety. However, the manifestations of allosterism at GPCRs are many and varied and, in the past, traditional screening methods have generally failed to detect many allosteric modulators. More recently, there have been a number of major advances in high throughput screening, including the advent of cell-based functional assays, which have led to the discovery of more allosteric modulator ligands than previously appreciated. In addition, a number of powerful analytical techniques have also been developed exclusively for detecting and quantifying allosteric effects, based on an increased awareness of various mechanisms underlying allosteric modulator actions at GPCRs. Together, these advances promise to change the current paucity of GPCR allosteric modulators in the clinical setting and yield novel therapeutic entities for the treatment of numerous disorders. PMID- 15279542 TI - Bivalent ligands for G protein-coupled receptors. AB - Bivalent ligands have been developed for a variety of G protein-coupled receptor targets, including opioid, dopamine, serotonin and muscarinic receptors. The most successful application of the bivalent ligand approach has been in the development of selective opioid antagonists, such as norbinaltorphimine. Several important principles have emerged from the study of norbinaltorphimine and related compounds, including the utility of bivalent ligands for targeting particular receptor classes and serving as a scaffold for specific interactions with unique amino acid residues that render receptor subtype selectivity. In recent years, several novel bivalent compounds were synthesized and characterized for activity at muscarinic receptors. The compounds display an interesting profile of high binding affinity, strong agonist potency and receptor subtype selectivity. Bivalent ligands represent an important starting point for the development of selective muscarinic agonists with potential utility in treating a variety of neurological disorders, including Alzheimer's disease and schizophrenia. The bivalent ligand approach may be generally applicable to other G protein-coupled receptors. PMID- 15279543 TI - A(1) adenosine receptor agonists: medicinal chemistry and therapeutic potential. AB - Adenosine receptors are widely distributed in the body and modulate numerous physiological processes. Four receptor subtypes (termed A(1), A(2A), A(2B) and A(3)) have been identified based on their pharmacological profile and cloning. Activation of the A(1) adenosine receptors produces a number of effects including a reduction in heart rate and atrial contractility, the attenuation of the stimulatory actions of catecholamines on the heart as well as a reduction of lipolysis in adipose tissue. As a result, A(1)AR agonists have been targeted as anti-arrhythmic and cardioprotective agents. This review discusses the synthesis, structure-activity relationships and therapeutic potential of A(1)AR agonists. PMID- 15279544 TI - Small-molecule antagonists of CCR5 and CXCR4: a promising new class of anti-HIV-1 drugs. AB - Combination therapy with reverse transcriptase and protease inhibitors greatly reduces morbidity and mortality in HIV-1-infected individuals. However, current anti-retroviral treatment cannot eradicate the virus from infected individuals and is often limited by the emergence of drug-resistant HIV-1 strains and long term toxicity. These problems emphasize the need to develop new anti-HIV-1 drugs targeting different steps in the viral replication cycle. HIV-1 entry into host cells represents a complex sequence of events involving several viral and cellular proteins that are potential drug targets. In particular, HIV-1 entry requires a sequential interaction of the viral envelope glycoprotein gp120 with CD4 and a co-receptor on the host cell plasma membrane. The CC-chemokine receptor 5 (CCR5) and the CXC-chemokine receptor 4 (CXCR4) are the primary HIV-1 co receptors in vivo, and are attractive targets for the development of new anti-HIV 1 drugs. CCR5 and CXCR4 belong to the protein superfamily of G protein-coupled receptors (GPCRs). Many orally bioavailable small-molecules interact with specific GPCRs and many existing drugs are orally bioavailable small-molecule agonists or antagonists of GPCRs. Several small-molecule antagonists of CCR5 and CXCR4 that block chemokine binding and HIV-1 entry have been identified in recent years and are now in pre-clinical or clinical development as drug candidates. This review discusses structural and functional aspects of these compounds and summarizes recent insights into how small-molecule antagonists interact with CCR5 and CXCR4, focusing on drug development programs that are well documented in the scientific literature. PMID- 15279545 TI - Treatment of chronic hepatitis B: from research to clinical practice via the consensus conferences. AB - The aim of antiviral therapy of chronic hepatitis B is to control Hepatitis B Virus (HBV) replication and to cure liver disease avoiding the progression of chronic hepatitis to cirrhosis and the end stage complications of cirrhosis. HBeAg/anti-HBe seroconversion is the hallmark of response in hepatitis B "e" antigen (HBeAg) positive patients. In the patients with antibody against HBeAg (anti-HBe positive) the combination of HBV DNA and anti-HBc IgM tests provides adequate diagnostic accuracy. Patients with biochemical and/or histological disease activity are eligible to therapy. The drug choice is based on age, disease severity, risk of complications, side effects and compliance, particularly in anti-HBe positive patients where prolonged treatment is needed. Interferon (5-6 MU daily or 9-10 MU thrice weekly for 4-6 months) is the first line therapy for HBeAg positive patients and (5-6 MU thrice weekly for 12-24 months) for anti-HBe positive patients. When IFN is contraindicated or ineffective, Lamivudine (100 mg) or Adefovir Dipivoxil (10 mg) are given as long as 4-6 months after HBeAg/anti-HBe seroconversion or for long-term treatments in HBeAg positive non-responders and anti-HBe positive patients. Patients with more advanced forms of cirrhosis and portal hypertension are to be treated within liver transplantation programs. Fifteen to 30% of treated patients achieve sustained response and more than 60% of them experience long-term disease remission during therapy. In perspectives, currently available molecular and immunologic tools and modelling of viral dynamics will help to address the therapy issue with more complex, efficacious and individually tailored treatment schedules. PMID- 15279546 TI - Treatment of acute hepatitis C. AB - Progression from acute to chronic HCV infection occurs in 50% to 84% of cases. In light of the risk of developing chronic disease and the response rate to treatment once the disease is established, it is important to consider early treatment of acute HCV infection before it progresses to the chronic state. Several studies evaluated the efficacy of either alpha or beta IFN monotherapy in patients with acute hepatitis C, but nearly all trials are small and present great variability regarding timing, schedule, response definition and patient characteristics. To overcome these limits, IFN efficacy has been assessed by meta analyses demonstrating that antiviral therapy during the acute phase of HCV significantly reduces evolution to chronic hepatitis. Accordingly, treatment of persons with acute hepatitis C is warranted. However, several issues remain to be addressed, such as the optimal regimen and timing. Recent data would indicate that induction with daily IFN is needed to optimize response and pegylated IFN monotherapy could be the best option. Combination therapy with ribavirin does not seem to increase the response rate but could be proposed as a second choice to patients non responding to IFN monotherapy. Delaying treatment by 2-3 months might allow the identification of cases who would spontaneously resolve without compromising efficacy. However, additional data are required to improve the selection of those patients at great risk of progressing to chronic disease, and also to establish the optimal treatment in terms of risk/benefit and cost effectiveness ratio. PMID- 15279547 TI - The role of ribavirin in the combination therapy of hepatitis C virus infection. AB - Ribavirin is a very broad-spectrum anti-viral agent used clinically to treat infections by Lassa fever virus, respiratory syncytial virus (RSV) and, in combination with Interferon-alpha (IFN-alpha), hepatitis C virus (HCV). Although it was originally synthesized over 30 years ago, the precise mechanisms of its therapeutic activities are still not fully understood. Ribavirin was shown to possess both direct and indirect action mechanisms against several DNA and RNA viruses. These include direct inhibition of viral RNA-dependent RNA polymerases, inhibition of the host inosine monophosphate dehydrogenase, modulation of the host immune response and inhibition of viral capping enzymes. More recently, ribavirin was demonstrated to be able to act as an RNA virus mutagen, increasing mutations in the RNA virus genome and reducing their infectivity. Still the real challenge is to identify which of its biological properties is responsible for the observed clinical efficacy on specific infections. Under this aspect, renewed interest results from its synergistic enhancement of interferon-alpha (IFN-alpha) therapy, which could open the way to develop more powerful anti-HCV compounds. This work purpose is to provide a broad overview of all the recognized ribavirin action mechanisms against HCV, which can possibly also explain its synergistic behavior with IFN-alpha. An overview on the corresponding HCV treatment clinical observations is also provided in the second part of this work. PMID- 15279548 TI - The immune responsiveness in hepatitis C virus infected patients: effects of interferon-alfa/ribavirin combined treatment on the lymphocyte response with special reference to B cells. AB - Previous data demonstrated that an elevated percentage of hepatitis C virus (HCV) infected patients are endotoxemic. Endotoxemic patients are poor responders to the interferon (IFN)- alpha/ribavirin (RIB) treatment and exhibit lower serum levels of IFN-gamma and interleukin (IL)-10 than the responder counterpart. Here we provide evidence that in endotoxemic HCV+ patients absolute numbers of CD19(+) cells (B cells) are higher than those observed in the non-endotoxemic counterpart at the end of the combined treatment. Moreover, anti lactoferrin (LF) antibodies are more elevated in non-responder HCV+ patients than in the responders. In turn, these autoantibodies may affect the antiviral activity of LF, on the one hand, and, on the other hand abrogate the LF binding to lipopolysaccharides (LPS). Such an interaction hampers the binding of LPS to LPS binding protein, thus inhibiting LPS fixation to CD14(+) cells and, ultimately, leading to a decreased release of proinflammatory cytokines. PMID- 15279549 TI - Artificial neural networks for the prediction of response to interferon plus ribavirin treatment in patients with chronic hepatitis C. AB - Combined therapy using Interferon alfa (IFN) and Ribavirin (RIB) represents the standard treatment in patients with chronic hepatitis C. However, the percentage of responders to this regimen is still low, while its cost and side effects are elevated. Therefore, the possibility to predict patient's response to the above treatment is of paramount importance. The progress in the field of informatics and its large use for decision making has led to the development of novel techniques related to the so-called Artificial Intelligence, even including artificial neural networks (ANNs). In chronic viral hepatitis data are lacking. By means of an artificial neural network (ANN), 300 patients treated with IFN plus RIB were retrospectively analyzed with the aim to predict the response to the treatment. One hundred patients resulted responders and 200 non-responders at the end of treatment and during the follow up. For evaluating the prediction of treatment response, six ANNs with 16 neurons of input, an hidden layer with 7 neurons and an output layer with one neuron were utilized. The ANN model generated a positive predictive value (i.e. posterior probability of treatment response) ranging from 57% to 75% while the negative one (i.e. posterior probability of no response to treatment) was comprised between 52% and 71%. The highest level of diagnostic accuracy was 70%. In conclusion, ANNs appear to be a promising tool in the prediction of treatment response in patients with chronic hepatitis C. However, additional prospective studies are necessary to ultimately validate this predictive method. PMID- 15279550 TI - Therapy of chronic hepatitis C virus infection in HIV co-infected people. AB - The introduction of highly active anti-retroviral therapy (HAART) for Human Immunodeficiency Virus (HIV) infection has significantly improved the life expectancy of HIV positive patients. Hepatitis C virus (HCV) co-infection is common in HIV infected patients and is now a significant cause of morbidity and mortality. Optimal management and treatment of HCV in HIV infected patients is therefore essential. Interferon-alpha (IFN-alpha) and ribavirin is the mainstay of treatment for HCV infection in HIV infected people. The sustained virological response rate (SVR) with combination therapy is lower than that commonly observed in HCV mono-infected patients. This is, at least in part, due to the very high treatment drop out rates. Ribavirin in combination with HAART is associated with particular side effects such as mitochondrial toxicity. Therefore, vigilant monitoring of patients during therapy, in specialist centers is essential. Pegylated interferon (PEG-IFN) plus ribavirin is particularly promising as it is easier to administer and will probably become the treatment of choice for co infected patients. A SVR is associated with genotype 2 and 3, in addition to a high CD4+ cell count and a low HCV load prior to therapy. The progression of HCV related liver disease in HIV positive patients is faster than in subjects with HCV infection alone. As a result, there is an increasing incidence of cirrhosis and end-stage liver disease in co-infected patients. Liver transplantation is being evaluated in many centers. To date the experiences are very limited but encouraging in term of survival rate. PMID- 15279551 TI - The impact of antiviral treatments on the course of chronic hepatitis C: an evidence-based approach. AB - Hepatitis C virus chronic infection is currently the most common cause of end stage liver disease. The benefit of antiviral therapy on liver histology and its impact on the long-term course of the disease has been extensively studied. However, the results are still equivocal and the overall assessment of treatment effect remains difficult to evaluate. Although the conclusions of the last National Institute of Health Consensus Development Conferences on Hepatitis C have recently been published, several important issues still remain unanswered. We review the available data by an evidence-based approach and conclude that: 1) peginterferon alfa is more effective than conventional interferon in improving liver histology; 2) monotherapy with PEG-interferon induces a marked reduction in staging in virological sustained responders and to a lesser degree in relapsers, but provides no benefit to nonresponders after 24-48 weeks of treatment; 3) maintenance therapy aiming to improve histology in virological nonresponders should be considered experimental and of unproven benefit; 4) although the reduction in the number of events in sustained responders suggests a long-term benefit of IFN therapy, available evidence is still insufficient to confirm that IFN prolongs life in HCV infected patients. Data of the long-term benefit of subjects treated with IFN plus ribavirin are still not available; 5) pooling of published data suggests a slight preventive effect of IFN on HCC development in patients with HCV-related cirrhosis. The magnitude of this effect is low and the observed benefit might be due to spurious associations. The preventive effect is more evident among sustained responders to interferon. PMID- 15279552 TI - Structural bioinformatics and its impact to biomedical science. AB - During the last two decades, the number of sequence-known proteins has increased rapidly. In contrast, the corresponding increment for structure-known proteins is much slower. The unbalanced situation has critically limited our ability to understand the molecular mechanism of proteins and conduct structure-based drug design by timely using the updated information of newly found sequences. Therefore, it is highly desired to develop an automated method for fast deriving the 3D (3-dimensional) structure of a protein from its sequence. Under such a circumstance, the structural bioinformatics was emerging naturally as the time required. In this review, three main strategies developed in structural bioinformatics, i.e., pure energetic approach, heuristic approach, and homology modeling approach, as well as their underlying principles, are briefly introduced. Meanwhile, a series of demonstrations are presented to show how the structural bioinformatics has been applied to timely derive the 3D structures of some functionally important proteins, helping to understand their action mechanisms and stimulating the course of drug discovery. Also, the limitation of these approaches and the future challenges of structural bioinformatics are briefly addressed. PMID- 15279553 TI - Prediction of protein function in the absence of significant sequence similarity. AB - Tremendous progress in DNA sequencing has yielded the genomes of a host of important organisms. The utilisation of these resources requires understanding of the function of each gene. Standard methods of functional assignment involve sequence alignment to a gene of known function; however such methods often fail to find any significant matches. Here we discuss a number of recent alternative methods that may be of use when sequence alignment fails. Function can be defined in a number of ways including E.C. number and MIPS and KEGG functional classes. Phylogenetic profiles show the pattern of presence or absence of a protein between genomes. Protein-protein interactions can be identified by searching for interacting pairs of proteins that are fused to a single protein chain in another organism. The gene neighbour method uses the observation that if the genes that encode two proteins are close on a chromosome, the proteins tend to be functionally related. More general methods use sequence properties such as amino acid composition, mean hydrophobicity, predicted secondary structure and post translational modification sites. Data mining methods devise rules in the form of IF... THEN statements that make predictions of function using sequence based attributes, predicted secondary structure and sequence similarity. Finally, structural features can be used, after modelling the structure of a protein from its sequence or solving its structure. Protein fold class can be strongly indicative of function, while other structural features, such as secondary structure content, cleft size and 3D structural motifs are also useful. PMID- 15279554 TI - High density DNA microarrays: algorithms and biomedical applications. AB - DNA microarrays are devices capable of detecting the identity and abundance of numerous DNA or RNA segments in samples. They are used for analyzing gene expressions, identifying genetic markers and detecting mutations on a genomic scale. The fundamental chemical mechanism of DNA microarrays is the hybridization between probes and targets due to the hydrogen bonds of nucleotide base pairing. Since the cross hybridization is inevitable, and probes or targets may form undesirable secondary or tertiary structures, the microarray data contain noise and depend on experimental conditions. It is crucial to apply proper statistical algorithms to obtain useful signals from noisy data. After we obtained the signals of a large amount of probes, we need to derive the biomedical information such as the existence of a transcript in a cell, the difference of expression levels of a gene in multiple samples, and the type of a genetic marker. Furthermore, after the expression levels of thousands of genes or the genotypes of thousands of single nucleotide polymorphisms are determined, it is usually important to find a small number of genes or markers that are related to a disease, individual reactions to drugs, or other phenotypes. All these applications need careful data analyses and reliable algorithms. PMID- 15279555 TI - P-selectin cell adhesion molecule in inflammation, thrombosis, cancer growth and metastasis. AB - P-selectin (CD62P) is a member of the selectin family of cell adhesion molecules. It is expressed on stimulated endothelial cells and activated platelets and mediates leukocyte rolling on stimulated endothelial cells and heterotypic aggregation of activated platelets onto leukocytes. It also mediates heterotypic aggregation of activated platelets to cancer cells and adhesion of cancer cells to stimulated endothelial cells. Using P-selectin knockout mice, the importance of P-selectin-mediated cell adhesive interactions in the pathogeneses of inflammation, thrombosis, growth and metastasis of cancers has been clearly demonstrated. Here we will summarize the current knowledge and highlight the important progress in the biomedical research of P-selectin biology, providing a suitable target for therapeutic interventions developed through both experimental and bioinformatic approaches. PMID- 15279556 TI - The design and development of deferiprone (L1) and other iron chelators for clinical use: targeting methods and application prospects. AB - Iron is essential for all human cells as well as neoplastic cells and invading microbes. Natural and synthetic iron chelators could affect biological processes involving iron and other metal ions in health and disease states. Iron overload is the most common metal toxicity condition worldwide. There are currently two iron chelating drugs, which are mostly used for the treatment of thalassaemia and other conditions of transfusional iron overload. Deferoxamine was until recently the only approved iron chelating drug, which is effective but very expensive and administered parenterally resulting in low compliance. Deferiprone (L1 or 1,2 dimethyl-3-hydroxypyrid-4-one) is the world's first and only orally active iron chelating drug, which is effective and inexpensive to synthesise thus increasing the prospects of making it available to most thalassaemia patients in third world countries who are not currently receiving any form of chelation therapy. Deferiprone has equivalent iron removal efficacy and comparable toxicity to deferoxamine. There are at least four other known iron chelators, which are currently being developed. Even if successful, these are not expected to become available for clinical use in the next five years and to be as inexpensive as deferiprone. The variation in the chemical, biological, pharmacological, toxicological and other properties of the chelating drugs and experimental chelators provide evidence of the difference in the mode of action of chelators and the need to identify and select molecular structures and substituents based on structure/activity correlations for specific pharmacological activity. Such information may increase the prospects of designing new chelating drugs, which could be targeted and act on different tissues, organs, proteins and iron pools that play important role not only in the treatment of iron overload but also in other diseases of iron and other metal imbalace and toxicity including free radical damage. Chelating drugs could also be designed, which could modify the enzymatic activity of iron and other metal containing enzymes, some of which play a key role in many diseases such as cancer, inflammation and atherosclerosis. Other applications of iron chelating drugs could involve the detoxification of toxic metals with similar metabolic pathways to iron such as Al, Cu, Ga, In, U and Pu. PMID- 15279557 TI - Analysis of protein glycation products by matrix-assisted laser desorption ionization time-of-flight mass spectrometry. AB - The term protein glycation summarizes non-enzymatic reactions between amino groups of proteins and sugars or sugar degradation products, leading to early glycation products (intact sugar attached) and advanced glycation end-products (AGEs). Protein glycation is involved in the progression of several diseases, such as diabetes, uremia, and atherosclerosis. However, qualitative and quantitative analysis of in vitro or in vivo glycated proteins is still a challenging task. The introduction of matrix-assisted laser desorption ionization time-of-flight technique (MALDI-TOF) changed mass spectrometry (MS) into a valuable tool for biomedical analysis, because the soft ionization procedure allows the measurement of proteins up to 100 kDa. In the last few years, MALDI TOF-MS was applied to the investigation of glycation processes: the analyses of plasma proteins from diabetic or uremic patients allowed a precise determination of the average number of sugar residues attached to serum albumin or immunoglobulins of each patient. Thus, a more individualized diagnosis of each patient was achieved by MALDI-TOF-MS than by other diagnostic tools. In a similar way, the glycation rate of hemoglobin, isolated from diabetic blood and of beta-2 microglobulin isolated from amyloid plaques from uremic patients was determined. The application of MALDI-TOF-MS for in vitro studies revealed important new insights into glycation mechanisms. Whereas the measurement of the intact proteins allows the determination of the average glycation rate, peptide mapping prior to MALDI-TOF-MS can reveal the exact structures of the glycation products and the glycation site. Furthermore, when the unmodified peptide is used as internal standard, MALDI-TOF-MS can also be used for reliable, site specific relative quantification of defined glycation products. PMID- 15279558 TI - Lactate in solid malignant tumors: potential basis of a metabolic classification in clinical oncology. AB - A number of studies have demonstrated that malignant transformation is associated with an increase in glycolytic flux and in anaerobic and aerobic cellular lactate excretion. Using quantitative bioluminescence imaging in various primary carcinomas in patients (uterine cervix, head and neck, colorectal region) at first diagnosis of the disease, we showed that lactate concentrations in tumors in vivo could be relatively low or extremely high (up to 40 micromol/g) in different individual tumors or within the same lesion. In all tumor entities investigated, high molar concentrations of lactate were correlated with a high incidence of distant metastasis already in an early stage of the disease. Low lactate tumors (< median of approx. 8 micromol/g) were associated with both a longer overall and disease free survival compared to high lactate lesions (lactate > approx. 8 micromol/g). Lactate dehydrogenase was found to be upregulated in most of these tumors compared to surrounding normal tissue. Numerous recent reports support these data by demonstrating various biological activities of lactate that can enhance the malignant behavior of cancer cells. These mechanisms include the activation of hyaluronan synthesis by tumor associated fibroblasts, upregulation of VEGF and of HIF-1 alpha, and direct enhancement of cellular motility which generates favorable conditions for metastatic spread. Thus, lactate accumulation not only mirrors but also actively enhances the degree of tumor malignancy. We propose that determination of lactate in primary tumors may serve as a basis for a novel metabolic classification which can lead to an improvement of prognosis and therapy in clinical oncology. PMID- 15279559 TI - Function and regulation of tumor necrosis factor receptor type 2. AB - TNF is a major proinflammatory cytokine, which exerts its effects through two different receptors known as TNF-R1 and TNF-R2. Whereas TNF-R1 induced signaling pathways have been very well characterized during the past years, TNF-R2 has been much less well studied. Nevertheless, the function of TNF-R2 should not be underestimated, the more because this receptor shows a much more restricted but inducible expression. Several inflammatory diseases and cancers show upregulated levels of soluble TNF-R2 or are associated with TNF-R2 polymorphisms, implicating an important role for TNF-R2 as a therapeutic target. Here we will review the mechanisms that regulate TNF-R2 expression, as well as discuss TNF-R2 induced signal transduction and cross-talk with TNF-R1. PMID- 15279560 TI - Advances in the discovery of novel antibacterial agents during the year 2002. AB - The development of bacterial resistance is a significant problem in the treatment of infection, and the importance of research directed toward the discovery of novel agents to treat infections cannot be underestimated. In the past, discovery programs have focused on modification of natural products or existing classes of marketed antibacterial agents. A significant period of time lapsed between the introduction of the nalidixic acid-based quinolones and the next novel antibacterial agent (Zyvox). However, the advent of the "genomics era" has provided a wealth of new targets that afford the opportunity to discover novel antibacterial agents. This review reports on the state of antibacterial research directed toward the development of novel antibacterial agents with novel mechanisms of action for the calendar year, 2002. While variations on existing drug classes continue to appear, we have chosen to limit our discussion to novel classes of antibacterial agents which have not yet been marketed. PMID- 15279561 TI - Monoamine oxidases: certainties and uncertainties. AB - A great deal has been learned about the behaviour of monoamine oxidase in the 75 years since it was first discovered, but there is still a great deal left to understand. This review concentrates on the dynamic aspects of our knowledge of the interactions of MAO with substrates and inhibitors and how it may collaborate with other enzymes, with particular emphasis on aspects that remain to be clarified. PMID- 15279562 TI - Structure and mechanism of monoamine oxidase. AB - Monoamine oxidases A and B (MAO A and MAO B) are mitochondrial outer membrane bound flavoproteins that catalyze the oxidative deamination of neurotransmitters and biogenic amines. A number of mechanism-based inhibitors (MAOI's) have been developed for clinical use as antidepressants and as neuroprotective drugs. To facilitate the development of more effective and specific inhibitors, a detailed understanding of the structures and catalytic mechanisms of these enzymes is required. The recent development of high level expression systems for producing recombinant human liver MAO A and MAO B in Pichia pastoris has facilitated the determination of the three dimensional crystal structures of MAO B (up to 1.7 angstroms resolution) in complex with different reversible (isatin, 1,4-diphenyl 2-butene) and irreversible inhibitors (pargyline, N-(2-aminoethyl)-p chlorobenzamide, and trans-2-phenylcyclopropylamine). The binding of substrates or inhibitors to MAO B involves an initial negotiation of a protein loop occurring near the surface of the membrane and two hydrophobic cavities; an "entrance" cavity and an "active site" cavity. These two cavities can either be separate or in a fused state depending on the conformation of the Ile199 side chain, which appears to function as a gate. The amine function of the bound substrate approaches the re face of the bent and "puckered" covalent FAD through an "aromatic cage" formed by two tyrosine residues that are perpendicular to the plane of the flavin ring. No amino acid residues that could function as acids or bases are found near the catalytic site. The existing structural data on MAO B support previous QSAR results and are also supportive of a proposed polar nucleophilic mechanism for MAO A and B catalysis rather than the alternatively proposed single electron transfer mechanism. PMID- 15279563 TI - Regulation of MAO-A and MAO-B gene expression. AB - MAO A and B genes are made of 15 exons with identical exon-intron organization. They are located on X-chromosome organized in opposite direction, tail to tail with 24 kb apart. Both promoters are GC-rich and regulated by transcription factor Sp1. However, they have distinctly different features. MAO B gene, but not MAO A gene, has TATA box. MAO B promoter contains two clusters of overlapping Sp1 sites, the CACCC repressor element. Transcription factors Sp1 and Sp4 can activate MAO B promoter activity through the proximal cluster of Sp1 sites and its activation can be repressed by the over-expression of Sp3 and a related family member, BTEB2. Decreased methylation and transcription repressor Sp3 upregulate human MAO B, but not MAO A, gene expression during Caco-2 differentiation. MAO B, but not MAO A gene, could be activated by PMA (phorbol 12 myristate 13-acetate) by protein kinase C, MAPkinase signal transduction pathway involves cJun and Egr-1. The differences in MAO A and B gene regulation may explain the different tissue-specific expression and functions of these two important isoenzymes. PMID- 15279564 TI - Platelet MAO and personality--function and dysfunction. AB - Research on the association between platelet monoamine oxidase (MAO) activity and personality traits, such as sensation seeking and impulsiveness, is reviewed with an emphasis on early history and current situation. The effects of MAO-inhibiting compounds in cigarette smoke for the interpretation of this association are discussed and recent results confirming a true association between platelet MAO activity and personality and vulnerability, for e.g. type 2 alcoholism are presented. From a clinical point of view, the link between platelet MAO activity, which is highly genetically regulated and is stable in the individual, and personality traits, has had its greatest impact on the understanding of the nature of constitutional factors making individuals vulnerable, for e.g. substance abuse and other forms of sociopathic behaviour. The molecular mechanisms underlying the association between platelet MAO and behaviour are discussed and evidence that common transcriptional factors, e.g. within the AP-2 family, regulating both the expression of platelet MAO and components of the central monoaminergic systems, such as synthesising enzymes, receptors and transporters, are presented. A hypothesis is put forward, that such common transcription factors may not directly regulate platelet MAO expression, but rather mitochondrial density, or outer mitochondrial membrane surface. PMID- 15279565 TI - Pharmacological aspects of (-)-deprenyl. AB - Deprenyl, the selective irreversible inhibitor of monoamine oxidase-B (MAO-B), has been synthesised as a potential antidepressant, however, due to its dopamine potentiating capacity, became a registered drug in the treatment of Parkinson's disease. Deprenyl possesses a wide range of pharmacological activities; some of them are not related to its MAO-B inhibitory potency. Beside its dopamine potentiating effect, it renders protection against a number of dopaminergic, cholinergic and noradrenergic neurotoxins with a complex mechanism of action. By inducing antioxidant enzymes and decreasing the formation of reactive oxygen species, deprenyl is able to combat an oxidative challenge implicated as a common causative factor in neurodegenerative diseases. In a dose substantially lower than required for MAO-B inhibition (10(-9)-10(-13) M), deprenyl interferes with early apoptotic signalling events induced by various kinds of insults in cell cultures of neuroectodermal origin, thus protecting cells from apoptotic death. Deprenyl requires metabolic conversion to a hitherto unidentified metabolite to exert its antiapoptotic effect, which serves to protect the integrity of the mitochondrion by inducing transcriptional and translational changes. Pharmacokinetic and metabolism studies have revealed that deprenyl undergoes intensive first pass metabolism, and its major metabolites also possess pharmacological activities. The ratio of the parent compound and its metabolites reaching the systemic circulation and the brain are highly dependent on the routes of administration. Therefore, in the treatment of neurodegenerative diseases, reconsideration of the dosing schedule, by lowering the dose of deprenyl and choosing the most appropriate route of administration, would diminish undesired adverse effects, with unaltered neuroprotective potency. PMID- 15279566 TI - Clinical applications of MAO-inhibitors. AB - Monoamine oxidase inhibitors (MAO-I) have been useful in the treatment of both psychiatric and neurological disorders over centuries. Here we focus on the development of this drug treatment. Focus is given on the use of irreversible MAO I's as well as on reversible ones. Benefit and side effects are reported for Parkinson's disease, Alzheimer's dementia, depression syndrome and panic disorders. The preclinical and clinical effects of selegiline with regard to neuroprotection are highlightened and the conclusion is drawn that there is good evidence for a clinical neuroprotective capacity based on the assumption that the 50 percent recovery of MAO-B is obtained already after a 10 days withdrawal of selegiline. There is also a focus on selegilines metabolism to amphetamine and metamphetamine. In order to avoid any such effects of metabolic compounds on the cardiovascular system Zydis Selegiline, a melt-tablet avoid of major metabolism to amphetamine and metamphetamine is described in detail. Developments in MAO-I research are discussed in detail as there are moclobemide, lacabemide, rasagiline. Interactions of MAO-I' with tricyclics and serotonin selective reuptake inhibitors (SSRI's) are described as there is mentioning of interactions of MAO-I's with other compounds in general. Tables and figures report on clinical studies and on pharmacological properties of MAO-I's. PMID- 15279567 TI - Tachykinins and tachykinin receptors: structure and activity relationships. AB - In addition to the classical neurotransmitters, acetylcholine and noradrenaline, a wide number of peptides with neurotransmitter activity have been identified in the past few years. Among them, the tachykinins substance P (SP), neurokinin A (NKA) and neurokinin B (NKB) appear to act as mediators of nonadrenergic, noncholinergic (NANC) excitatory neurotransmission. Tachykinins interact with specific membrane proteins, belonging to the family of G protein-coupling cell membrane receptors. Until now, three tachykinin receptors termed NK1 (NK1R), NK2 (NK2R) and NK3 (NK3R) have been cloned in different species. A large amount of reports suggests that these peptides are involved in nociception and neuroimmunomodulation, and in the development of different diseases such as bronchial asthma, inflammatory bowel syndrome and psychiatric disorders. Tachykinin receptor antagonists are therefore promising, therapeutically relevant agents. However, and in spite of extensive research, the obtention of selective antagonists of tachykinin receptors have revealed very difficult. An understanding of how ligands interact with their receptors is essential to permit a rational design of compounds acting selectively at the tachykinin receptor level. The major aim of the present article is to review the structure-activity data that exist for tachykinins and their receptors, with the purpose of getting insight into basic structural requirements that determine ligand/receptor interaction. PMID- 15279568 TI - A molecular understanding of mast cell activation and the promise of anti allergic therapeutics. AB - Mast cells are central to allergic disease. Their immediate (exocytosis of granule-stored allergic-mediators) and delayed (de novo synthesis of inflammatory mediators) response to an allergen underlies the symptoms seen in acute; and chronic allergic disease. Thus, intervention in the allergen-mediated activation of mast cells is a long sought after goal in the treatment and management of allergic disease. The recent gain in deciphering the molecular mechanisms underlying immunoglobulin E (IgE)-mediated mast cell activation has provided optimism for the development of new therapeutic strategies. Among the most promising is the use of humanized anti-IgE antibodies that inhibit binding of IgE to its high affinity receptor (FcepsilonRI) on the mast cell. Other strategies target molecules proximal to FcepsilonRI, whose activities are central in mast cell activation. One such molecule, Syk kinase, has been targeted by various approaches including a small molecule inhibitor that specifically abrogates mast cell degranulation. More recently, various molecules that function to promote protein-protein interactions (adapters) were demonstrated as essential to mast cell degranulation and cytokine production. It remains to be seen if these molecules hold therapeutic promise for disease intervention. Additional studies identifying molecules required for mast cell granule fusion and content exocytosis also bodes well for discovery of new therapeutic targets. While our understanding of IgE-mediated mast cell activation is still at its inception, the modest success in identifying molecules essential to this process affords some confidence for better treatment of allergic disease. PMID- 15279569 TI - Multidrug resistance and anticonvulsants: new studies with some enaminones. AB - The multidrug resistance (MDR), often conferred by the active extrusion of drugs from the cell, is a phenomenon often seen in cancer cells that may become resistant to a wide spectrum of drugs with varying chemical structures or cellular targets. This event has recently been reported for anticonvulsants. Studies in our laboratories on this occurrence with some enaminones have shown that the enaminones display high efflux ratios and are recognized by P glycoprotein (P-gp) and/or the multidrug resistance protein (MRP), which have been reported as the main efflux transporters responsible for the development of MDR. Recent studies have uncovered interesting structural analogues that can modulate the functional activity of P-gp, suggesting a possible increase in the bioavailabillity of P-gp substrate drugs when administered concurrently. PMID- 15279570 TI - Strategies for the stereocontrolled formation of oxygen analogues of penicillins and cephalosporins. AB - The synthesis of oxacephalotin and oxacephamandol, which are more active than natural, sulfur-containing congeners, and the isolation of clavulanic acid, a potent inhibitor of beta-lactamase enzymes, directed attention of many academic and industrial laboratories the synthesis of oxygen analogues of penicillins and cephalosporins. The present review focuses attention on the problem of stereocontrol in the formation of a desired configuration of the bridgehead carbon atom in the title compounds. Five feasible synthetic methods leading to the basic skeletons of clavams and 5-oxacephams are discussed. Three of them involve the nucleophilic substitution at C-4 of the azetidin- 2-ones performed as inter- or intramolecular process and the remaining two involve cycloaddition reactions between ketenes and iminoethers, or between vinyl ethers and isocyanates. Owing to the general application, stereospecificity and high asymmetric induction, the last method seems to be most advantageous. The weak point of the nucleophilic substitution methodology is that a nucleophile approaches the 3-substituted azetidin-2-one ring preferentially anti to the existing substituent and in the case where there is no substituent at C-3, that the stereoselectivity of formation of the new chirality center at C-4 is low. All discussed methods are illustrated by the examples taken from the literature. PMID- 15279571 TI - Asymmetric synthesis of beta-lactams through the Staudinger reaction and their use as building blocks of natural and nonnatural products. AB - In the last two decades, the better understanding of the mechanistic aspects of the beta-lactams' biological activity and their inhibition, and the chemical exploitation of beta-lactams as synthetic intermediates in organic chemistry, have experienced a continuous and somewhat complementary advance. A prerequisite for such a development has been the accessibility of enantiopure beta-lactams. The latter are now routinely prepared most often through the ketene-imine cycloaddition reaction, also termed the Staudinger reaction. This review accounts for the recent progress made in the asymmetric synthesis of beta-lactams (with special emphasis in the Staudinger reaction approach), as well as in their use as synthetic intermediates en route to natural products, including alpha- and beta amino acids and peptides derived therefrom. PMID- 15279572 TI - Beta-lactam cholesterol absorption inhibitors. AB - beta-Lactams have recently been identified as potent, highly efficacious cholesterol absorption inhibitors (CAIs). The discovery, SAR, and asymmetric synthesis of this class of hypolipidemic agents are described. Metabolism studies of the first clinical candidate, Sch 48461, led to the identification of a more potent second generation clinical candidate, Sch 58235 (ezetimibe) incorporating key structural elements of the active metabolites. A summary of preclinical and early clinical studies of ezetimibe as monotherapy and in combination with statins is also presented. Efforts to identify a pharmacophore model has led to the development of conformationally constrained analogs and analogs with conformational biases based on intramolecular hydrogen bonding possibilities. Finally, mechanism of action studies have led to the development of many biochemical tools for the investigation and identification of novel proteins involved in cholesterol uptake. PMID- 15279573 TI - Azetidin-2-ones, synthon for biologically important compounds. AB - Azetidin-2-one, a four-membered cyclic lactam (beta-lactam) skeleton has been recognised as a useful building block for the synthesis of a large number of organic molecules by exploiting the strain energy associated with it, in addition to its use in the synthesis of a variety of beta-lactam antibiotics. Efforts have been made in exploring such new aspects of beta-lactam chemistry using enantiomerically pure beta-lactams as versatile intermediates for the synthesis of aromatic beta-amino acids and their derivatives, peptides, polyamines, polyamino alcohols, amino sugars and polyamino ethers. The development of methodologies based on beta-lactam nucleus is now referred as 'the beta-lactam synthon methods'. The selective bond cleavage of the strained ring coupled with further interesting transformation render this fascinating molecule as a powerful building block. This provides an access to diverse structural type of synthetic target molecules lacking beta-lactam ring structure. This review provides an account of synthesis of organic compounds having biological significance at the same time lacking beta-lactam ring, by using beta-lactam as synthon. PMID- 15279574 TI - Beta-lactams as versatile synthetic intermediates for the preparation of heterocycles of biological interest. AB - Since the advent of penicillin, the beta-lactam antibiotics have been the subject of much discussion and investigation, within both the scientific and public sectors. The primary biological targets of the beta-lactam antibacterial drugs are the penicillin binding proteins, a group of transpeptidases anchored within the bacterial cellular membrane, which mediate the final step of cell wall biosynthesis. The extensive use of common beta-lactam antibiotics such as penicillins and cephalosporins in medicine has resulted in an increasing number of resistant strains of bacteria through mutation and beta-lactamase gene transfer. Thus, a handful of nonconventional fused polycyclic beta-lactams have been described in the literature in order to overcome the defence mechanisms of the bacteria. In fact, tricyclic beta-lactam antibiotics, generally referred to as trinems, are a new class of synthetic antibacterial agents featuring good resistance to beta-lactamases and dehydropeptidases. In addition, recent discoveries have shown other biological properties of these compounds apart from their antibacterial action. In this sense, beta-lactams can serve as inhibitors of serine proteases, such as human leukocyte elastase (HLE) or thrombin, acyl-CoA cholesterol acyltransferase inhibitors and inhibitors of human cytomegalovirus. Additional impetus for research efforts on beta-lactam chemistry has been provided by the introduction of the beta-lactam synthon method, a term coined by Ojima 20 years ago, according to which 2-azetidinones can be employed as useful intermediates in organic synthesis. The usefulness of beta-lactams in the stereocontrolled synthesis of heterocycles of biological significance is based on the impressive variety of transformations, which can be derived from this system, due inter alia to a high chirality content that can be transferred into a variety of products. The cyclic 2-azetidinone skeleton has been extensively used as a template on which to build the heterocyclic structure fused to the four-membered ring, using the chirality and functionalisation of the beta-lactam nucleus as a stereocontrolling element. Alternatively, the direct one-pot generation of fused nitrogen heterocyclic systems from the nitrogen framework of 2-azetidinone derivatives has been achieved by selective bond breakage and rearrangement. It is our aim in this Review to highlight the state of the art in this endeavour, consisting either of the stereocontrolled synthesis of fused polycyclic beta lactams of antibacterial interest, or stereoselective synthesis of different sized heterocycles of biological significance. Representative examples of the latter include indolizidines, pyrrolizidines, pirrolidines, pyrroles, taxoids and macrolide natural products. PMID- 15279575 TI - The discovery and development of modified penicillin- and cephalosporin-derived beta-lactamase inhibitors. AB - While beta-lactam antibiotics remain among the most commonly prescribed pharmaceutical products, their effectiveness is currently threatened by the development of bacterial resistance. One key resistance mechanism is the ability to destroy the antibiotic through utilization of one or more types of beta lactamase. An effective countermeasure is to employ a combination product, consisting of both a beta-lactam antibiotic and a beta-lactamase inhibitor. Unfortunately, currently available inhibitors narrowly target only class A beta lactamases. This review will detail our research, directed toward the development of a useful broad-spectrum beta-lactamase inhibitor. In the process, we have discovered new inhibitors capable of simultaneously inactivating class A, C, and D beta-lactamase, produced conjugate siderophore/beta-lactamase inhibitors, and explored the SAR's of tunable, cephalosporin-derived beta-lactamase inactivators. Useful synthetic methodology will be described, which simplifies the large scale production of many known inhibitors and which allows the rapid preparation of libraries of prospective inhibitors. PMID- 15279576 TI - Chemical diversity of bioactive marine natural products: an illustrative case study. AB - The marine environment contains a number of plants, animals and micro organisms, which, due to the unique adaptations to their habitat, elaborate a wide diversity of natural products with specific bioactivities. These products provide a rich source of chemical diversity that can be used to design and develop new potentially useful therapeutic agents. The huge variety of the structures present in marine organisms has been illustrated through the case study of the sponge Plakortis simplex, whose chemical analysis, started in our laboratories about ten years ago, revealed an incredible variety and abundance of secondary metabolites. The obtained results have been presented with the intention of drawing some conclusions of general relevance. Particularly, the problem of the limited availability of natural compounds for both structural and preliminary pharmacological studies has been discussed, this issue becoming a serious obstacle when the pharmacological research reaches a more advanced stage. Furthermore, the origin of the chemodiversity in Plakortis simplex and, in general, in marine invertebrates has been discussed; in this respect, the possible cooperative role of symbiotic micro-organisms in the biosynthesis of the varied metabolic content typical of these organisms has been considered. PMID- 15279577 TI - Advanced preclinical and clinical trials of natural products and related compounds from marine sources. AB - The marine environment has proven to be a very rich source of extremely potent compounds that have demonstrated significant activities in anti-tumor, anti inflammatory, analgesia, immuno-modulation, allergy and anti-viral assays. Although the case can and has been made that the nucleosides such as Ara-A and Ara-C are derived from knowledge gained from investigations of bioactive marine nucleosides, no drug directly from marine sources (whether isolated or by total synthesis) has yet made it to the commercial sector in any human disease. However, as shown in this review, there are now significant numbers of very interesting molecules that have come from marine sources, or have been synthesized as a result of knowledge gained from a prototypical compound, that are either in or approaching Phase III clinical trials in cancer, analgesia and allergy, with a very substantial number of other, quite different potential agents following in their wake, in these and in other diseases. PMID- 15279578 TI - Drugs from the sea: conopeptides as potential therapeutics. AB - Marine cone snails from the genus Conus are estimated to consist of up to 700 species. These predatory molluscs have devised an efficient venom apparatus that allows them to successfully capture polychaete worms, other molluscs or in some cases fish as their primary food sources. The toxic venom used by the cone shells contains up to 50 different peptides that selectively inhibit the function of ion channels involved in the transmission of nerve signals in animals. Each of the 700 Conus species contains a unique set of peptides in their venom. Across the genus Conus, the conotoxins represent an extensive array of ion channel blockers each showing a high degree of selectivity for particular types of channels. We have undertaken a study of the conotoxins from Australian species of Conus that have the capacity to inhibit specifically the nicotinic acetylcholine receptors in higher animals. These conotoxins have been identified by mass spectroscopy and their peptide sequences in some cases deduced by the application of modern molecular biology to the RNA extracted from venom ducts. The molecular biological approach has proven more powerful than earlier protein/peptide based technique tor the detection of novel conotoxins [1,2]. Novel conotoxins detected in this way have been further screened for their abilities to modify the responses of tissues to pain stimuli as a first step in describing their potential as lead compounds for novel drugs. This review describes the progress made by several research groups to characterise the properties of conopeptides and to use them as drug leads for the development of novel therapeutics for the treatment of a range of neurological conditions. PMID- 15279580 TI - New trends in the design of drugs against Alzheimer's disease. AB - First described by Alois Alzheimer in 1907, Alzheimer's disease (AD) is the most common dementia type, affecting approximately 20 million people worldwide. As the population is getting older, AD is a growing health problem. AD is currently treated by symptomatic drugs, the acetylcholinesterase inhibitors, based on the cholinergic hypothesis (1976). During the past decade, advances in neurobiology have conducted to the identification of new targets. Although some of these innovative approaches tend to delay onset of AD, others are still symptomatic. In this review, we present an overview of the several strategies and new classes of compounds against AD. PMID- 15279581 TI - Endothelin and subarachnoid hemorrhage-induced cerebral vasospasm: pathogenesis and treatment. AB - Endothelin (ET)-mediated vasoconstriction has been implicated in the pathophysiology of various disorders, e.g. hypertension, chronic heart failure, acute renal failure, pulmonary hypertension, and subarachnoid hemorrhage (SAH) induced cerebral vasospasm. The potential involvement of ETs in cerebral vasospasm following SAH has triggered considerable interest in designing therapeutic strategies to inhibit biological effects of ET. Major approaches include: (a) reducing the levels of circulating ET- 1 by the the specific anti- ET- 1 antibodies, (b) antagonizing the ET receptors, and (c) suppressing the biosynthesis of ET-1. To date, numerous antagonists of ET(A) and/or ET(B) receptors have been discovered, and some are under clinical evaluation. Inhibitors of endothelin-converting enzymes (ECEs), which catalyze the biosynthesis of ET-1, have also been synthesized. Two types of ECE-1 inhibitors have been evaluated in various animal disease models: dual ECE-1/neutral endopeptidase 24.11 (NEP) inhibitors and selective ECE-1 inhibitors. In this article, the effects of ET receptor antagonists and ECE-1 inhibitors on the prevention and reversal of SAH-induced cerebral vasospasm in preclinical animal models are reviewed. PMID- 15279582 TI - Engineered killer mimotopes: new synthetic peptides for antimicrobial therapy. AB - This review deals with a novel approach to produce synthetic antibiotic peptides (killer mimotopes), similar to those described for the conversion of epitopes into peptide mimotopes, allowing their use as surrogate vaccines. Synthetic peptides pertaining to the complementary determining regions (CDRs) of a recombinant antiidiotypic antibody (PaKTscFv), which mimic the wide spectrum of microbicidal activity of a killer toxin produced by the yeast Pichia anomala (PaKT), have proven to act as structural or functional mimotopes of PaKT. This activity appeared to be mediated by interaction with specific cell wall killer toxin receptors (KTRs), mainly constituted by beta glucans. Killer mimotopes have shown in vitro an impressive microbicidal activity against Candida albicans. They were adopted as a model of PaKT- and PaKTscFv-susceptible microorganisms. Optimization through alanine scanning led to the generation of an engineered decapeptide (KP) of a CDR-L1 pertaining antibody fragment with an enhanced in vitro microbicidal activity. It had a potent therapeutic effect against experimental vaginal and systemic candidiasis in normal and immunodeficient mice caused by flucanozole susceptible and resistant yeast isolates. KP exerted a microbicidal activity in vitro against multidrug-resistant eukaryotic and prokaryotic pathogenic microorganisms, which was neutralized by interaction with laminarin (beta 1,3-glucan). To our knowledge, KP represents the prototype of an engineered peptide fragment derived from a microbicidal recombinant antiidiotypic antibody. It is capable of exerting antimicrobial activity in vitro and a therapeutic effect in vivo presumably acting through interaction with the beta glucan KTR component in the cell walls of pathogenic microorganisms. PMID- 15279583 TI - Chemical mediators of gallbladder dysmotility. AB - In order to accomplish its contribution to the digestive process, the gallbladder must contract appropriately during its emptying phases and it must be able to relax adequately for filling to occur. A variety of neuro-hormonal inputs to gallbladder smooth muscle coordinate the gallbladder emptying process with other events occurring in the bowel. Gallbladder dysmotility can disrupt the normal flow of bile to the small bowel, resulting in digestive dysfunction. In addition to this, alterations in gallbladder motility may play a role in pathological conditions, such as cholesterol gallstone formation and cholecystitis. It is still not entirely clear whether impaired gallbladder emptying is a cause or consequence of cholesterol gallstones, but recent experimental evidences demonstrate that cholesterol can directly affect the plasma membrane of gallbladder smooth muscle cells to cause impaired contraction. In addition, gallbladder emptying is impaired in acute gallbladder inflammation, probably as the result of the deleterious neural and muscular actions of inflammatory mediators such as reactive oxygen species, prostaglandins and histamine. It should also be noted that opiate treatments in critically ill patients can reduce gallbladder motility by inhibiting neurotransmitter release, and may contribute to the onset of acalculous cholecystitis, which is associated with significant morbidity in these patients. PMID- 15279579 TI - Mechanism targeted discovery of antitumor marine natural products. AB - Antitumor drug discovery programs aim to identify chemical entities for use in the treatment of cancer. Many strategies have been used to achieve this objective. Natural products have always played a major role in anticancer medicine and the unique metabolites produced by marine organisms have increasingly become major players in antitumor drug discovery. Rapid advances have occurred in the understanding of tumor biology and molecular medicine. New insights into mechanisms responsible for neoplastic disease are significantly changing the general philosophical approach towards cancer treatment. Recently identified molecular targets have created exciting new means for disrupting tumor specific cell signaling, cell division, energy metabolism, gene expression, drug resistance and blood supply. Such tumor-specific treatments could someday decrease our reliance on traditional cytotoxicity-based chemotherapy and provide new less toxic treatment options with significantly fewer side effects. Novel molecular targets and state-of-the-art, molecular mechanism-based screening methods have revitalized antitumor research and these changes are becoming an ever-increasing component of modern antitumor marine natural products research. This review describes marine natural products identified using tumor-specific mechanism-based assays for regulators of angiogenesis, apoptosis, cell cycle, macromolecule synthesis, mitochondrial respiration, mitosis, multidrug efflux and signal transduction. Special emphasis is placed on natural products directly discovered using molecular mechanism-based screening. PMID- 15279584 TI - Adherence to antiretroviral therapies: state of the science. AB - HIV-related morbidity and mortality have been dramatically improved in populations treated with combination antiretroviral therapy. Although it is widely recognized that adherence to the antiretroviral medication regimens is vital to treatment success, rates of adherence to the regimens are often poor. There is a large body of research exploring the problem of adherence to antiretroviral medications. The literature is, to date, dominated by reports identifying factors that are predictive or associated with antiretroviral adherence. Adherence is increasingly understood as a dynamic behavior influenced by a matrix of interrelated factors that change over time. Preliminary reports suggest varying degrees of success with strategies designed to improve adherence. Multifaceted strategies appear to be the most promising; however, there are few controlled studies substantiating the effectiveness of these approaches and the mechanisms by which the interventions promote adherence are not well understood. More well powered, rigorously evaluated antiretroviral adherence intervention trials are urgently needed. Further, problems in the field exist because of limitations in the available adherence measures. This paper provides a comprehensive review and analysis of the state-of-the-science of this body of work. Despite substantial attention to antiretroviral adherence in recent years, there remain significant gaps in our understanding. PMID- 15279585 TI - Pathogenesis of HIV-associated non-Hodgkin lymphoma. AB - In the current era of highly active antiretroviral therapy (HAART), the prevalence of HIV-associated non-Hodgkin lymphoma (H-NHL) is not as high as in the beginning of the acquired immunodeficiency syndrome (AIDS) epidemic, but still remains above that of non-HIV-infected individuals. Therefore, the epidemiology suggests that the pathogenesis of H-NHL may be multifactoral, involving the interaction of the immune system with HIV or other pathogens. Although HIV is a retrovirus, it is not characterized with the typical oncogenic potential associated with insertional mutagenesis. HIV integration occurs as a necessary step in the life cycle allowing for replication and transcription of the viral genome to take place. While HIV is thought to integrate randomly, more recent studies have suggested that integration occasionally occurs in a less random fashion. While the majority of H-NHL may be secondary to immune dysfunction, this non-random integration may be a factor leading to pathogenesis of a small subset of H-NHL since the pathways involved in malignancies usually require an accumulation of abnormalities leading to proliferation and transformation. A common element among lymphomas in this setting of immune dysfunction is the inability to control a pathogen that acts as an ongoing immune activator. Thus, an increased risk of H-NHL emerges from the combination of immunodeficiency, increased immune activation and possible HIV insertional mutagenesis. The focus of this review will be on the role viral pathogenesis plays in oncogenic transformation. PMID- 15279586 TI - Recent advances in understanding the molecular mechanisms of HIV-1 entry and fusion: revisiting current targets and considering new options for therapeutic intervention. AB - Recent advances in our understanding of the cellular and molecular mechanisms of HIV-1 entry provide the basis for novel therapeutic strategies that prevent viral penetration of the target cell-membrane, while reducing detrimental virus and treatment effects on cells and prolonging virion exposure to immune defenses. A number of potential sites for therapeutic intervention become accessible during the narrow window between virus attachment and the subsequent fusion of viral envelope with the cell membrane. Initial approaches considered for prevention of HIV-1 entry included the use of natural ligands, small-molecule inhibitors and/or monoclonal antibodies, which could interfere with gp120-CD4 and/or gp120 CXCR4/CCR5 interaction. Others avenues pursued were the use of agents that interfere with the conformational changes of gp120 or gp41 leading to subsequent fusion of viral and cellular membranes. More recently, strategies have emerged that involve inhibition of thiol/disulfide oxidoreductases (factors which facilitate Env transition from an inactive to a fusion-competent conformation) to block redox exchange, thereby impeding the entry process. This review provides a summary of the cellular and viral factors mediating the HIV-1 entry process, with an emphasis on novel therapeutics targeting Env-receptor/coreceptor interaction, Env conformational change and the membrane fusion process. PMID- 15279587 TI - Immune restoration disease: a consequence of dysregulated immune responses after HAART. AB - Immune Restoration Diseases (IRD) are a collection of atypical 'opportunistic infections' and inflammatory diseases seen in human immunodeficiency virus (HIV) patients after HIV viraemia is suppressed by highly active antiretroviral therapy (HAART). IRD probably reflect dysregulated immune responses against pre-existing infections by opportunistic pathogens, with different immunopathological mechanisms for different pathogens. For example, mycobacterial IRD are associated with delayed type hypersensitivity (DTH) responses to mycobacterial antigens, whereas patients who experience cytomegalovirus (CMV) IRD have elevated plasma levels of soluble CD30, a marker of a T2 cytokine environment expressed by activated CD8 T-cells. As IRD are often compartmentalised to organs, monitoring serological markers such as pathogen-specific IgG antibody, may be informative, as demonstrated for CMV and hepatitis C virus (HCV)-associated IRD. Genetic studies have provided evidence of distinct immunopathological mechanisms and inherited susceptibility to IRD associated with mycobacterial and herpesviridae infections. The expansion of HAART in the developing world where many HIV patients have low CD4+ T-cell counts and high rates of concomitant infections will place a large number of patients at-risk of developing IRD. It is therefore important to understand the immunopathology so that prevention, diagnosis and treatment can be improved. PMID- 15279588 TI - Glycosylation of the ENV spike of primate immunodeficiency viruses and antibody neutralization. AB - Neutralizing antibody titers have been correlated with protection following vaccination against many viral pathogens. The logical target of protective antibody responses elicited by potential HIV vaccines should be the viral Env spike on the surface of the virion. However, the potency and titers of neutralizing antibodies that arise during HIV infection are generally discouragingly low and the antibodies that do arise recognize mainly autologous virus. This is thought to be a result of a combination of immunodominance of hypervariable regions of the Env protein that can easily escape neutralization, antibody reactivity to gp160 "decoy" protein in cell surface debris or monomeric gp120, conformational constraints within the Env trimer that create unfavorable antibody binding conditions and extensive glycosylation of the exposed regions of Env within the trimer. This review will describe current knowledge regarding glycosylation as a mechanism of neutralization resistance and discuss experimental approaches used to overcome this resistance. Part of the strategy toward development of an optimally immunogenic Env spike will likely require modification of Env glycosylation. PMID- 15279589 TI - Vpu: a multifunctional protein that enhances the pathogenesis of human immunodeficiency virus type 1. AB - The Vpu protein is the smallest of the proteins encoded by human immunodeficiency virus type 1 (HIV-1). This transmembrane protein interacts with the CD4 molecule in the rough endoplasmic reticulum (RER), resulting in its degradation via the proteasome pathway. Vpu also has been shown to enhance virion release from infected cells. While much has been learned about the function of Vpu in cell culture systems, its exact role in HIV-1 pathogenesis is still unknown. This has been primarily due to the lack of a suitable primate model system since vpu is found only in HIV-1 and simian immunodeficiency viruses isolated from chimpanzees (SIVcpz), and three species of old world monkeys within the genus Cercopithecus. Several laboratories have developed pathogenic molecular clones of simian-human immunodeficiency virus (SHIV) in which the tat, rev, vpu and env genes of HIV-1 are expressed in the genetic background of SIV. The availability of such clones has allowed investigators to assess the role of Vpu in pathogenesis using a relevant animal model. This review will focus on the current understanding of the structure-function relationships of Vpu protein and recent advances using the SHIV model to assess the role of Vpu in HIV-1 pathogenesis. PMID- 15279590 TI - HIV-1 superinfection: evidence and impact. AB - Superinfection is defined as infection by a second virus during an immunologic steady state, following infection by a primary virus. It is now well established that superinfection with HIV-1 occurs in humans. Detection of an increasing number of circulating recombinant forms, which result from infection of a cell by two or more clades, suggests that superinfection occurs more frequently than previously thought. The second virus (from a different clade or the same clade as the primary virus) can superinfect some time after the first and this is associated with rapid viral rebound and immune decline. Primary infection with a specific clade appears not to provide cross-protection against superinfection with a different clade or the same clade. Estimating the overall impact of HIV-1 superinfection on pathogenesis and attempts to create a broadly protective prophylactic HIV-1 vaccine is complicated by our inability to quantify the true incidence and prevalence. PMID- 15279591 TI - Cytopenias in HIV infection: mechanisms and alleviation of hematopoietic inhibition. AB - Hematopoietic abnormalities including anemia, cytopenias, and alterations of the stem cell plasticity in the bone marrow microenvironment commonly occur in HIV infected patients. These observations suggest that HIV-1 infection may affect processes important during early stages of hematopoiesis or stem cell differentiation. Hematopoietic abnormalities may be caused by altered stem cell differentiation possibly due to abnormal lineage specific expression of certain cellular genes such as cytokines relevant to hematopoiesis. These cytokines could affect regulatory signals important in hematopoiesis. However, in HIV infected individuals, it is not only the virus but also the highly active antiretroviral therapy (HAART) that both contribute to persistent hematopoietic suppression and ensuing cytopenias. Even if a lowering of HIV replication by HAART were to occur in infected individuals, prolonged HAART by itself and/or appearance of drug resistant mutants can contribute to hematopoietic suppression and resulting cytopenias. However, confounding factors such as opportunistic infections, immune mediated effects, or the consequences of prolonged physiological stress, which could contribute to decreased hematopoiesis in patients or other individuals, make the causative role of HIV in vivo, uncertain. The severe combined immunodeficient mouse transplanted with human fetal thymus and liver tissues (SCID-hu) is a small animal model which mimics HIV infection in humans, and is useful to determine the mechanisms of HIV-1 induced hematopoietic inhibition and development of drug therapies for interventions of stem cell differentiation. Further, SCID mouse serves as a useful small animal recipient of human progenitor cells and also allows us to study the differentiation of these cells in vivo. Results from our studies are expected to provide relief for HIV infected individuals from hematopoietic inhibition and ensuing cytopenias. PMID- 15279592 TI - The challenge of antiretroviral-drug-resistant HIV: is there any possible clinical advantage? AB - Resistance to antiretroviral drugs is associated with reduced treatment options and, therefore, increased risk of disease progression or death. Despite the main goal of antiretroviral therapy should be achievement of complete suppression of HIV replication, the accumulation of resistance mutations in patients with multiple treatment failure makes this objective often difficult, or even impossible to obtain. Thus, clinicians should be aware about the complex relationship between drug pressure and viral replication capacity and about some potential advantages related to antiretroviral drug resistance. The two main biological mechanisms that can be at the origin of these clinical benefits are: reduction of viral fitness and viral hypersusceptibility. The term "fitness" indicates the ability of HIV to maintain high rate of replication capacity in presence of antiretroviral drugs. Consequently, replication capacity, high in presence of wild type virus, tends to decrease when HIV must adapt its enzymes to work in presence of drugs. A reduction of viral fitness is observed in patients harbouring mutations conferring resistance to all the three classes of antiretroviral drugs currently in use. Particularly, the effects on reduction of replication capacity related to M184V mutation in reverse transcriptase are analyzed. Viral isolates with reduced susceptibility or resistance to some antiretroviral drugs may exhibit significant increased susceptibility to other drugs acting on the same enzyme. This phenomenon is known as hypersusceptibility and can be demonstrated in vitro by phenotypic assays. Phenotypic hypersusceptibility has been demonstrated for all three drug classes. Particularly, NNRTI hypersusceptibility, associated with NRTI mutations, is analyzed and discussed. PMID- 15279593 TI - First and second generations of COX-2 selective inhibitors. AB - The identification and characterization of the inducible form of cyclooxygenases (COX-2) stimulated the investigations to develop efficient, non-steroidal anti inflammatory drugs (NSAIDs) with reduced side effects (essentially gastro intestinal toxicity) compared to classical NSAIDs. This review focuses on the chemical and pharmacological properties (pre-clinical data) of marketed COX-2 inhibitors. PMID- 15279594 TI - Structural approach for COX-2 inhibition. AB - The design of selective COX-2 inhibitors is a new approach to obtain potent, anti inflammatory drugs but with less side effects. Several families of such inhibitors were reported in literature. In this review, the drug design processes used to understand their binding mode and the origin of selectivity of these compounds are described. PMID- 15279595 TI - The second generation of COX-2 inhibitors: clinical pharmacological point of view. AB - Valdecoxib, parecoxib, etoricoxib and lumiracoxib represent the second generation of selective COX-2 inhibitors. In comparison to the first generation, they show an at least equivalent efficacy in the treatment of pain and inflammation. However, the postulated gain of safety is yet difficult to determine and seems to be, if any, small. PMID- 15279596 TI - COX-2 selective inhibitors, carbonic anhydrase inhibition and anticancer properties of sulfonamides belonging to this class of pharmacological agents. AB - The sulfonamides constitute an important class of drugs, with several types of pharmacological agents possessing antibacterial, anti-carbonic anhydrase, diuretic, hypoglycaemic, antithyroid, protease inhibitory and anticancer activity among others. A recently developed class of pharmacological agents incorporating primary sulfamoyl moieties in their molecule is constituted by the COX-2 selective inhibitors, with at least two clinically used drugs, celecoxib and valdecoxib. Another drug of this class, rofecoxib, does not contain sulfonamide moieties, but the isosteric and isoelectronic methylsulfone group. It was recently shown that the sulfonamide COX-2 selective inhibitors (but not the methylsulfone ones) also act as nanomolar inhibitors of several isozymes of the metallo-enzyme carbonic anhydrase (CA), some of which are strongly involved in tumourigenesis. In consequence, the potent anticancer effects of the sulfonamide COX-2 selective inhibitors and the much weaker such effects of rofecoxib, reported ultimately by many researchers, may be explained by the contribution of CA inhibition to such processes in addition to COX-2 inhibition. PMID- 15279597 TI - Recent development in the field of dual COX / 5-LOX inhibitors. AB - Cyclooxygenases and lipoxygenase are key enzymes in the arachidonic acid metabolism. Dual inhibitors are drugs able to block both the COX and the 5-LOX metabolic pathways. Compared to COX or LOX pathways single inhibitors, dual inhibitors present at least two major advantages. First, dual inhibitors, by acting on the two major arachidonic acid metabolic pathways, possess a wide range of anti-inflammatory activities. Secondly, dual inhibitors appear to be almost exempt from gastric toxicity, which is the most troublesome side effect of non selective COX inhibitors. PMID- 15279598 TI - Advance in understanding the biosynthesis of prostacyclin and thromboxane A2 in the endoplasmic reticulum membrane via the cyclooxygenase pathway. AB - Recent advances in topological and structural characterization of the prostacyclin (PGI(2)) and thromboxane A(2) (TXA(2)) synthases have led to the understanding of the biosynthesis of PGI(2) and TXA(2) at a structural level. This mini-review focuses on the molecular mechanism of the isomerization of the prostaglandin H(2) to PGI(2)and TXA(2) by their synthases in the endoplasmic reticulum (ER) membrane coordinated with cyclooxygenase-1 or -2. This review summarizes the evidences in which the biosynthesis of PGI(2)and TXA(2) are influenced/modulated by the membrane anchor residues of the synthases and the ER membrane itself, and provides the structural basis for engineering the synthases for the next generation of gene therapy and drug designs targeting the specific synthases. PMID- 15279599 TI - New developments on thromboxane modulators. AB - Thromboxane A(2) (TXA(2)) is a labile product formed from arachidonic acid by cyclooxygenase. The pathogenesis of numerous cardiovascular, pulmonary, and thromboembolic diseases can be related to this metabolite. Therefore, TXA(2) modulators have been developed for 20 years. This review will highlight the evolution in the field of TXA(2) modulators. PMID- 15279600 TI - Modulation of the arachidonic cascade with omega3 fatty acids or analogues: potential therapeutic benefits. AB - Increasing interest in the role of omega3 fatty acids has arisen in these latest years since evidence of their implication in the cardioprotective fish based diet of the Inuit has been demonstrated. Furthermore, several in vitro, in vivo and epidemiological studies support the benefit of this fatty acids intake in various pathological states such as in the cardiovascular, cancer, inflammation, psychiatric, paediatric, pulmonary, dermatological and ophthalmologic fields. This review will focus on metabolism and pharmacological implication of omega3 fatty acids intake as well as its interest in the prevention or treatment of the above-mentioned pathologies. PMID- 15279601 TI - "Dipeptoids": from the chemical structure of the endogenous peptide to the design of peptidomimetics. AB - The present review details the rational multi-step process followed for the discovery of a family of non-peptide CCK receptor ligands ("dipeptoids"), starting from the structure of the endogenous peptide, CCK(8). Emphasis will be made on the N- and C-terminal modifications, on the singular effects of the stereochemical changes and the incorporation of conformational constraints into the structure of "dipeptoids", and on the modifications directed to improve the pharmacological profile of these compounds to afford valuable clinical candidates. PMID- 15279602 TI - Predicting synthetic accessibility: application in drug discovery and development. AB - The estimation and use of synthetic accessibility in the drug discovery process is discussed. A distinction is drawn between synthetic feasibility and accessibility and the practical uses of an accessibility score are examined. Various techniques used in the estimation of accessibility are described and their utility and potential accuracy compared. PMID- 15279603 TI - The development of inhibitors of heparanase, a key enzyme involved in tumour metastasis, angiogenesis and inflammation. AB - Heparanase is an endo-beta-glucuronidase that degrades the glycosaminoglycan heparan sulfate, a major component of the extracellular matrix and basement membranes, and has been implicated in such processes as inflammation, angiogenesis and metastasis. The identification of inhibitors of heparanase is an attractive approach towards developing new therapeutics for metastatic tumours and chronic inflammatory diseases. This review focuses on heparanase inhibitors that have been isolated or synthesised to date. More recent developments in the understanding of heparanase structure and function that may ultimately aid in the future design of inhibitors with improved potency and specificity, are also discussed. PMID- 15279604 TI - LPS signal transduction: the picture is becoming more complex. AB - Lipopolysaccharide (LPS) is the principal initiator of septic shock and it is to a large extent responsible for post-operative mortality. The use of antibiotics is still the most successful therapy against infection that may lead to sepsis and septic shock. With the advent of antibiotic resistant strains like MRSA the usefulness of conventional antibiotics is declining and new treatment strategies for LPS-mediated septic shock are called for. In this review we discuss the molecular mechanisms that are involved in the recognition of LPS and in the initiation of an immune response. Furthermore, we also review the recent insights in the signal transduction including receptor clustering and signalosome activation. Further insight into LPS-dependent signal transduction will assist the development of novel rational therapy. PMID- 15279605 TI - Endotoxins: relationships between structure, function, and activity. AB - Molecules of endotoxin (lipopolysaccharides, LPS), forming a unique molecular class with peculiar physico-chemical properties, impart a very important role in the formation and function of the outer membrane (OM). The latter is strictly asymmetric with the LPS monolayer forming the outer leaflet and the phospholipid (PL) monolayer forming the inner leaflet. Thus, the OM builds a functional lipid environment for the OM proteins (Omp's, porins) and the LPS layer is the first locus of interaction of the bacterial cells with components of the host's immune system,. Therefore its physical state and biochemical parameters (such as the fluidity of the lipid A acyl chains and the backbone charge density) essentially influence the defense of bacteria against the attack of the human immune systems such as the complement and antimicrobial peptides/proteins. LPS, released from the bacterial cell, is responsible for a variety of biological effects which can be ascribed to the unique structural features of LPS- the three-dimensional supramolecular structure and the intramolecular conformation - which are essential determinants of the bioactivity of endotoxins. Here, the physico chemical parameters which are important on the one side for the function of the OM and on the other side for the activity of isolated LPS are reviewed. PMID- 15279606 TI - Inhibition of endotoxin response by synthetic TLR4 antagonists. AB - Endotoxin, from the outer membrane of Gram-negative bacteria, has been implicated as the etiological agent of a variety of pathologies ranging from relatively mild (fever) to lethal (septic shock, organ failure, and death). While endotoxin (also known as lipopolysaccharide or LPS) is a complex heterogeneous molecule, the toxic portion of LPS (the lipid A portion) is relatively similar across a wide variety of pathogenic strains of bacteria, making this molecule an attractive target for the development of an LPS antagonist. Research over the past fifteen years focused on the design of various lipid A analogs including monosaccharide, acyclic and disaccharide compounds has lead to the development of E5564, an advanced, unique and highly potent LPS antagonist. E5564 is a stable, pure LPS antagonist that is selective against endotoxin-mediated activation of immune cells in vitro and in animal models. In Phase I clinical trials, we have developed an ex vivo endotoxin antagonism assay that has provided results on pharmacodynamic activity of E5564 in addition to the more typical safety and pharmacokinetic evaluations. Results from these assays have been reinforced by analysis of in vivo antagonistic activity using a human endotoxemia model. Results from all of these studies indicate that E5564 is an effective in vivo antagonist of endotoxin, and may prove to be of benefit in a variety of endotoxin mediated diseases. This review discusses the evolution of synthetic LPS antagonists with emphasis on the SAR and development of E5564 and its precursors. PMID- 15279607 TI - Endotoxin neutralizing peptides. AB - Neutralization and sequestration of bacterial lipopolysaccharide which plays a key role in gram-negative sepsis is required to block the progression of sepsis at early stages in addition to destroying bacteria. Many of the host defense peptides which have antimicrobial activity are also able to bind to and neutralize LPS, however, these two activities do not necessarily correlate. Due to its toxicity application of polymyxin B as the prototype of LPS neutralizing peptide is limited to topical applications and extracorporeal removal of endotoxin. Development of novel endotoxin neutralizing peptides without the toxicity of polymyxin B have been based on the natural host defense peptides, fragments of LPS binding proteins and engineered peptides. Neutralization of LPS can be achieved through several different peptide fold motifs, which are reviewed in this article. Endogenous host defense peptides, fragments of endotoxin-binding proteins and synthetic anti-endotoxin peptides fold into alpha-helical, beta hairpin, extended and compact conformations without regular secondary structure. In animal models many of the peptides have demonstrated good in vitro and in vivo endotoxin neutralizing activity but up to now none of the peptides has been approved for clinical application with an anti-endotoxin indication. Recent developments include preparation of novel types of endotoxin neutralizing compounds such as peptides modified by lipophilic moieties and non-peptidic molecules, particularly lipopolyamines and on the other hand additional medical applications such as extracorporeal endotoxin removal, targeting to inflammation sites or endotoxoid based vaccines. PMID- 15279608 TI - The search for molecular determinants of LPS inhibition by proteins and peptides. AB - Lipo-poly-saccharide (LPS) induced Gram-negative sepsis and septic shock remain lethal in up to 60 % of cases, and LPS antagonists that neutralize its endotoxic action are the subject of intensive research. The molecular motifs of specific binding of LPS by antiendotoxin proteins and peptides may lead to an understanding of LPS action at the atomic level and provide clues for the development of new immunomodulatory compounds for use as therapy in the treatment of Gram-negative bacterial sepsis. The interaction of LPS with its cognate binding proteins has been structurally elucidated in the single case of the X-ray crystallographic structure of LPS in complex with the integral outer membrane protein FhuA from E. coli K-12 (Ferguson et al., Science 1999, 282, 2215). This structure and other known structures of LPS binding proteins have been used to propose a common binding motif of LPS to proteins. Another independent source of structural information are solution structures of peptides in complex with LPS that can be determined using the transferred NOE effect. The molecular mechanisms of biological activity of bacterial endotoxins can additionally be probed by theoretical means. The growing structural knowledge is opening pathways to the design of peptides or peptidomimetics with improved antiendotoxin properties. PMID- 15279609 TI - A review of depsipeptide and other histone deacetylase inhibitors in clinical trials. AB - The histone deacetylase inhibitors (HDIs) are a new class of antineoplastic agents currently being evaluated in clinical trials. While these agents have been studied extensively in the laboratory, only recently has their mechanism of action begun to be elucidated. Several structural classes of compounds have been shown to exert histone deacetylase inhibition, including sodium n-butyrate, suberoylanilide hydroxamic acid, LAQ824, CI-994, MS-275, and depsipeptide. The HDIs have been shown to induce differentiation, to decrease cell proliferation, and to induce cell death. HDIs are thought to exert their anti-neoplastic effects by altering the expression of genes that play a role in the control of cell growth, and transformation. The HDIs have specific and well-defined effects on cancer cells. Preliminary results from clinical trials suggest that these agents are very promising. While there were sporadic case reports of activity using the early generation HDIs, dramatic responses have recently been observed in patients with T-cell lymphomas treated with depsipeptide, one of the newer agents. With the well-defined molecular effects on cancer cells, surrogate markers can be analyzed for evidence of activity and efficacy using either tumor samples or normal tissue. Presented in this review are details from clinical trials with both earlier and newer generations of HDIs. Toxicities specific to this class of agents are outlined and possibilities for rational combination therapies are discussed. PMID- 15279610 TI - Membrane disrupting lytic peptides for cancer treatments. AB - Membrane disrupting lytic peptides are abundant in nature and serve insects, invertebrates, vertebrates and humans as defense molecules. Initially, these peptides attracted attention as antimicrobial agents; later, the sensitivity of tumor cells to lytic peptides was discovered. In the last decade intensive research has been conducted to determine how lytic peptides lyse bacteria and tumor cells. A number of synthetic peptides have been designed to optimize their antibiotic and anti-tumor properties and improve their therapeutic capabilities. The sequences of alpha-helical cationic membrane disrupting peptides has been discussed, their proposed mechanisms of action reviewed, and their roles in cell selectivity and tumor cell destruction considered. Parameters important for the selection and design of lytic peptides for cancer treatments include increased activities against tumor cells, low cytolytic activities to normal mammalian cells and erythrocytes. The conjugation of lytic peptides with hormone ligands and the production of pro-peptides provide methods for targeting of cancer cells. The therapeutic possibilities in cancer treatment by targeted lytic peptides are broad and offer improvement to currently used chemotherapeutical drugs. Lytic peptides interact with the tumor cell membrane within minutes, and their activity is independent of multi-drug resistance. Lytic peptide-chorionic gonadotropin (CG) conjugates destroy primary tumors, prevent metastases and kill dormant and metastatic tumor cells. These conjugates do not destroy vital organs; they are not antigenic, and are more toxic to tumor cells than to non-malignant cells. PMID- 15279611 TI - Small molecule toxins targeting tumor receptors. AB - Targeting toxic therapeutics to tumors through receptors over expressed on the surface of cancer cells can reduce systemic toxicity and increase the effectiveness of the targeted compounds. Small molecule targeted therapeutics have a number of advantages over toxic immunoconjugates including better tumor penetration, lack of neutralizing host immune response and superior flexibility in selection of drug components with optimal specificity, potency and stability in circulation. Three major components of the targeted drug, the toxic warhead, tumor-specific ligand and the linker can influence the properties of each other and thus have to be optimized for each system. All receptor-targeted drugs are delivered inside the cells through endocytosis and undergo processing liberating the toxins in endosomes and lysosomes. Common delivery route defines a number of general requirements for each drug component. The review addresses currently known possible receptor targets and their ligands along with toxins that have been used and that have a potential to be successfully applied in tumor targeting. Linkers that are stable in circulation, but efficiently cleaved in lysosomes constitute an essential component of receptor-targeted drugs and are evaluated in greater detail. PMID- 15279612 TI - Cancer-specific ligands identified from screening of peptide-display libraries. AB - Although monoclonal antibodies have demonstrated clinical potentials as tumor targeting agents, poor tumor penetration of the antibodies due to the size of molecules and liver/bone marrow toxicity by non-specific uptake of the antibodies are the two major limitations of antibody therapy. Peptidic targeting agents may ease the problems associated with antibody cancer therapy. Combinatorial libraries displayed on microorganisms have successfully been utilized to discover cell surface-binding peptides, which can be tumor-targeting agents. Among many molecular display techniques, phage display has been the most popular approach. Peptides can be used as targeting molecules of receptor-targeted toxins and gene therapy, imaging and/or therapeutic agents, and nano-medical technologies. Recent results from preclinical studies with various peptides support their targeting potential and suggest that the role of peptides as targeting molecules in drug development should be further exploited. PMID- 15279613 TI - Role of progastrins and gastrins and their receptors in GI and pancreatic cancers: targets for treatment. AB - Accumulating evidence in literature suggests that amidated and non-amidated gastrins (gastrin precursors) may play an important role in the proliferation and carcinogenesis of gastrointestinal and pancreatic cancers, especially in the presence of DNA damaging agents and/or infectious agents. Amidated gastrins appear to have a protective role, while progastrins exert growth promoting effects in cancers. Several receptor subtypes and signal transduction pathways mediate the biological effects of the gastrin peptides. Progastrin and gastrins also exert anti-apoptotic effects, which may additionally contribute to the growth and co-carcinogenic effects of these peptides on GI mucosal cells in vivo. Amidated gastrins additionally play an important role in the migration of GI epithelial cells, and in glandular morphogenesis, while progastrins may play an important role in invasion and metastasis. Therefore, targeting progastrins, gastrins, and their cognate receptors may provide a therapeutic tool for treating GI and pancreatic cancers. Targeting CCK2-receptors has, so far, not provided optimal beneficial effects. However, targeting gastrins via a vaccine approach has provided some encouraging results for treating GI and pancreatic cancers. It is expected that targeting precursor gastrins (progastrins and gly-gastrins), exclusively rather than amidated gastrins, may be more effective for treating GI cancers. Since GI cancers at advanced stages are largely responsive to autocrine and intracrine progastrins, down-regulation of intracellular progastrins will likely be more effective at this stage. PMID- 15279614 TI - The urokinase receptor as a potential target in cancer therapy. AB - The glycolipid-anchored receptor for urokinase-type plasminogen activator (uPAR) is essential for cell-surface associated plasminogen activation and is overexpressed at the invasive tumor-stromal microenvironment in many human cancers. In line with this, uPAR and uPA levels in both resected tumor tissue and plasma are of independent prognostic significance for patient survival in several types of human cancer. As the expression of both uPAR and its cognate protease ligand thus appears to be correlated with tumor malignancy, the uPA-uPAR interaction represents an attractive target for the development of either antagonists with possible anti-invasive effects or cytotoxic agents with anti tumor effects. In this review we discuss recent achievements in the development of protein and peptide based drug candidates targeting uPAR. The majority of these compounds has been optimized for human uPAR and exhibits a pronounced species-specificity showing little or no reactivity with murine uPAR. This evidently complicates their application in preclinical intervention studies, since an intimate interplay between the tumor and its associated stroma is a distinct feature of the invasive phenotype of many human cancers. The virtues and drawbacks of various mouse tumor models as surrogates for human cancer are also discussed in relation to uPAR targeting. PMID- 15279615 TI - An expanding appreciation of the role chemokine receptors play in cancer progression. AB - The contribution of small molecular weight chemoattractant cytokines (chemokines) and their receptors in the trafficking of tumor, immune and vascular cells pertaining to the development and progression of cancer has begun to be investigated. The current literature indicates that interactions between the immune network, angiogenic and cell survival cascades are important for the trafficking and progression of human cancer and that chemokines and chemokine receptors play a central role in these complex inter-related pathways. Several therapeutic approaches have been reviewed and suggest that the most promising arise from the development of combinations of chemokine receptor antagonists. PMID- 15279616 TI - The MEP pathway: a new target for the development of herbicides, antibiotics and antimalarial drugs. AB - Isoprenoids, a diverse group of compounds derived from the five-carbon building units isopentenyl diphosphate (IPP) and its isomer dimethylallyl diphosphate (DMAPP), are essential for survival in all organisms. Animals synthesize their isoprenoids from mevalonic acid (MVA), whereas most pathogenic bacteria and the malaria parasites utilize a completely different pathway for IPP and DMAPP synthesis, the methylerythritol phosphate (MEP) pathway. Plants use both pathways for the synthesis of isoprenoid precursors. The recent elucidation of the MEP pathway has opened the possibility to develop new strategies against microbial pathogens. Novel immunotherapeutic agents can be developed based on the MEP pathway intermediates known to activate the proliferation of human V-delta-9 V gamma-2 T-cells after infection by many pathogenic bacteria and protozoa. Moreover, the design of specific inhibitors of MEP pathway enzymes (which are highly conserved but show no homology to mammalian proteins) should result in herbicides and drugs with broad-spectrum antimicrobial activity without mechanism based toxicity to humans. A good example is the cure of bacterial infections and malaria with fosmidomycin, a highly stable inhibitor of the MEP pathway. The use of plants as test systems has led to the identification of additional inhibitors such as ketoclomazone. Biochemical, genetic and crystallographic approaches with the MEP pathway enzymes are now starting to characterize the inhibition kinetics and identify which residues play a structural or catalytic role. Current efforts should eventually contribute to an effective drug designed to fight against microbial pathogens that show resistance to currently available agents. PMID- 15279617 TI - The therapeutic role of taurine in ischaemia-reperfusion injury. AB - As a non-toxic endogenous antioxidant, the semi-essential amino acid taurine is a potential attenuator of oxidative damage such as that produced by ischaemia reperfusion injury. Ischaemia-reperfusion injury is a well established if paradoxical phenomenon whereby ischaemic tissue, doomed to necrosis if it is not reperfused, is actually further damaged by oxidative attack when perfusion is restored. This paper is a review of the literature concerning therapeutic strategies in ischaemia-reperfusion injury, including non-pharmacological and pharmacological interventions. There is consistent experimental evidence of an important role of taurine in ischaemia-reperfusion injury, with a clinical role emerging in human trials of taurine administered prior to coronary artery bypass grafting and heart valve surgery. PMID- 15279618 TI - Mitochondrial DNA and aging. AB - Among the numerous theories that explain the process of aging, the mitochondrial theory of aging has received the most attention. This theory states that electrons leaking from the ETC (electron transfer chain) reduce molecular oxygen to form O2*- (superoxide anion radicals). O2*-, through both enzymic and non enzymic reactions, can cause the generation of other ROS (reactive oxygen species). The ensuing state of oxidative stress results in damage to ETC components and mtDNA (mitochondrial DNA), thus increasing further the production of ROS. Ultimately, this 'vicious cycle' leads to a physiological decline in function, or aging. This review focuses on recent developments in aging research related to the role played by mtDNA. Both supportive and contradictory evidence is discussed. PMID- 15279619 TI - B-group vitamin supplementation mitigates oxidative damage after acute ischaemic stroke. AB - Evidence shows that there is a rapid increase in the production of markers of oxidative damage immediately following acute stroke and that endogenous antioxidant defences are rapidly depleted, thus permitting further tissue damage. Several studies point to an antioxidant effect of B-group vitamins and a pro oxidant effect of elevated plasma tHcy (total homocysteine). In the present study, we assessed whether supplementary B-group vitamins during this critical period will enhance antioxidant capacity and mitigate oxidative damage. Forty eight patients with acute ischaemic stroke within 12 h of symptom onset were assigned to receive daily oral supplements of B-group vitamins comprising 5 mg of folate, 5 mg of vitamin B2, 50 mg of vitamin B6 and 0.4 mg of vitamin B12 (n=24) or no supplements (n=24) for 14 days. The treatment group and controls were matched for stroke subtype and age. Blood samples were obtained before intervention and also at 7 and 14 days post-recruitment for measurement of the following biomarkers: red cell folate (whole blood folate corrected with haematocrit), erythrocyte glutathione reductase activity coefficient (EGRAC; measure of vitamin B2 status), plasma pyridoxal phosphate (vitamin B6 status), plasma vitamin B12, plasma alpha-tocopherol, plasma ascorbic acid, plasma TAOC (total antioxidant capacity), plasma MDA (malondialdehyde), plasma tHcy and CRP (C-reactive protein). Supplementation for 14 days with B-group vitamins significantly increased the plasma concentrations of pyridoxal phosphate and red blood cell folate and improved a measure of B2 status compared with the control group (P<0.05). Plasma tHcy decreased in both groups albeit less in the control group, but differences in cumulative changes were not significant. There was, however, a decrease in plasma MDA concentration in the treatment group, in contrast with the increase seen in the control group and these differences were significant (P=0.05). CRP concentration, a marker of tissue inflammation, was significantly lower in the treatment group compared with controls (P<0.05). In conclusion, B-group vitamin supplementation immediately post-infarct may have antioxidant and anti-inflammatory effects in stroke disease independent of a homocysteine-lowering effect. PMID- 15279620 TI - Crystal structures of the antitermination factor NusB from Thermotoga maritima and implications for RNA binding. AB - NusB is a prokaryotic transcription factor involved in antitermination processes, during which it interacts with the boxA portion of the mRNA nut site. Previous studies have shown that NusB exhibits an all-helical fold, and that the protein from Escherichia coli forms monomers, while Mycobacterium tuberculosis NusB is a dimer. The functional significance of NusB dimerization is unknown. We have determined five crystal structures of NusB from Thermotoga maritima. In three crystal forms the protein appeared monomeric, whereas the two other crystal forms contained assemblies, which resembled the M. tuberculosis dimers. In solution, T. maritima NusB could be cross-linked as dimers, but it migrated as a monomer in gel-filtration analyses, suggesting a monomer/dimer equilibrium with a preference for the monomer. Binding to boxA-like RNA sequences could be detected by gel shift analyses and UV-induced cross-linking. An N-terminal arginine-rich sequence is a probable RNA binding site of the protein, exhibiting aromatic residues as potential stacking partners for the RNA bases. Anions located in various structures support the assignment of this RNA binding site. The proposed RNA binding region is hidden in the subunit interface of dimeric NusB proteins, such as NusB from M. tuberculosis, suggesting that such dimers have to undergo a considerable conformational change or dissociate for engagement with RNA. Therefore, in certain organisms, dimerization may be employed to package NusB in an inactive form until recruitment into antitermination complexes. PMID- 15279621 TI - Higher IL-10 levels are associated with less effective clearance of Plasmodium falciparum parasites. AB - The implications of high levels of the immune regulatory cytokine IL-10 in Plasmodium falciparum malaria are unclear. IL-10 may down-regulate pro inflammatory responses and also exacerbate disease by inhibiting anti-parasitic immune functions. To study possible inhibiting effects on parasite clearance, IL 10 plasma levels were determined in 104 Tanzanian children, 1 to 4 years old, with acute uncomplicated P. falciparum malaria, and analysed for association with parasite densities during 3 days of anti-malarial treatment. Higher baseline IL 10 plasma levels were associated with statistically significantly higher parasite densities after 24, 48 and 72 h of treatment. These associations could not be explained by differences in initial parasitaemia, temperature, age, sex or type of treatment. Induction of high IL-10 production might be a direct or indirect mechanism whereby the parasite evades the immune response. PMID- 15279622 TI - Monoclonal antibodies against a 62 kDa proteinase of Trichomonas vaginalis decrease parasite cytoadherence to epithelial cells and confer protection in mice. AB - Trichomonas vaginalis infects the epithelium of the genital tract. The mechanism by which it invades the tissue leading to the disease is not thoroughly understood. However, results of several studies seem to agree that parasite adhesion to epithelium cells is the initial step leading to infection in women. T. vaginalis is associated with high levels of proteolytic activity. The role of some of these proteinases in the development of infection has been demonstrated. The current study establishes the role of a 62 kDa excretion-secretion proteinase in parasite cytoadherence. Monoclonal antibodies (MAbs) against this enzyme were tested for their ability to inhibit this process. Three stable hybrid producers of IgG(1)class MAbs (4D8, 1A8, 3C11) against the 62 kDa proteinase were obtained. Two of them (4D8 and 1A8) showed parasite recognition by immunofluorescence. Parasite cytoadherence to a monolayer of HeLa cells was inhibited by the 4D8, 1A8 and 3C11 antibodies. MAb 4D8 administered 24 h before a challenge with T. vaginalis by the intraperitoneal route was able to protect the majority of mice. Nitric oxide levels in the serum of animals inoculated with MAb 4D8 and challenged with the parasite were significantly different from those recorded in mice treated with an unrelated MAb. These studies show that an appropriate antibody against 62 kDa proteinase can help the host resist a challenge by the intraperitoneal route with T. vaginalis. PMID- 15279623 TI - Th1 and Th2 immunological profile induced by cysteine proteinase in murine leishmaniasis. AB - This study evaluated the immune response to three synthetic peptides (pI, VMVEQVICFD; pII, VGGGLCFE; pIII, PYFLGSIMNTCHYT) from the COOH-terminal region of Leishmania amazonensis cysteine proteinases, in BALB/c- and CBA-infected mice with this parasite. Only BALB/c mice, previously inoculated with pI, showed a distinct exacerbation of the lesion. Blastogenesis assays were performed with lymph node cells from the group of mice infected with L. amazonensis, but not inoculated with the peptides, and we detected lymphoproliferative responses in BALB/c and CBA mice with a 5.0x and 2.5x stimulation index, respectively. Cell phenotypes were evaluated and in both mouse strains CD8(+)cells proliferated more extensively than CD4(+)cells. INF-gamma and nitric oxide were detected only in supernatants obtained from CBA mouse lymph node cell cultures, whereas IL-4 was detected in supernatant cultures from both strains of mice. Our results indicate a preferential stimulation of CD8(+)T-cell subsets by the pI cysteine proteinase peptide and the induction of both Th1 and Th2 phenotypes during L. amazonensis infections in both BALB/c and CBA mice. We suggest that the pI peptide from the COOH-terminal region of the cysteine proteinase induces immune responses different from those elicited by the whole molecule. PMID- 15279624 TI - Inhibition of neutrophil recruitment by ES of Nippostrongylus brasiliensis. AB - It has been reported that excretory-secretory (ES) material from the parasitic nematode Nippostrongylus brasiliensis has potential modulatory effects on the host's immune system. We observed that intratracheal instillation of ES from the L3 stage of the parasite reduced neutrophil numbers in LPS-induced inflammation as assessed by bronchoalveolar lavage. PMID- 15279625 TI - Trichomonas vaginalis-induced apoptosis in RAW264.7 cells is regulated through Bcl-xL, but not Bcl-2. AB - The purpose of this study was to determine whether anti-apoptotic proteins of the Bcl-2 family such as Bcl-2 and Bcl-x(L), proteins that confer resistance to apoptotic death from some stimuli, block apoptotic cell death in RAW264.7 cells upon treatment with Trichomonas vaginalis. In this study, the expression level of Bcl-2 was unchanged throughout the course of apoptotic cell death, and overexpressed Bcl-2 did not prevent release of cytochrome c, the significant change of the membrane potential, activation of caspases, and PARP cleavage in T. vaginalis-treated RAW264.7 cells. On the other hand, Bcl-x(L)expression was decreased after T. vaginalis treatment accompanied with Bax activation. Furthermore, we showed that release of mitochondrial cytochrome c, cleavage of caspase-9 and PARP during apoptosis in T. vaginalis-treated RAW264.7 cells were considerably diminished by transfection with overexpressed Bcl-x(L), and overexpressed Bcl-x(L)could inhibit T. vaginalis-induced apoptosis in RAW264.7 cells. In addition, interestingly, pre-treatment with caspase inhibitors, Boc-D FMK and Z-DEVD-FMK, significantly abolished T. vaginalis-induced down-regulation of Bcl-x(L), suggesting that caspase-3 may play a pivotal role in the process of apoptosis as well as the down-regulation of Bcl-x(L)by T. vaginalis. Therefore, these results suggest that T. vaginalis-induced apoptosis in RAW264.7 cells can occur via a Bcl-x(L)-dependent apoptotic mechanism. PMID- 15279626 TI - Infection of BALB/c mice with Angiostrongylus costaricensis decreases pulmonary inflammatory response to ovalbumin. AB - SUMMARY The prevalence of asthma in developing countries is lower than in developed countries. Viral, bacterial and parasitic infections may be associated with this discrepancy. The relationship between parasitic infection and asthma prevalence is not clear. Previous controversial data have demonstrated that parasitic infection may either predispose or protect against the development of asthma. The aim of this study is to determine whether infection with Angiostrongylus costaricensis (A. costaricensis) decreases inflammatory lung response to ovalbumin (OVA) in mice. Seven BALB/c mice were infected with A. costaricensis by orogastric gavage (10 larvae/mouse) on day (D) 0. The mice were immunized against OVA by intraperitoneal injection on D 5 and D 12 and received an intranasal OVA challenge (40 micro L) on D 15 and D 17. On D 19 bronchoalveolar lavage (BAL) was performed. Six BALB/c mice (control group) were immunized with OVA using the same protocol, but were not infected with A. costaricensis. Interleukin (IL)-1beta and IL-6 levels were measured in the BAL fluid by using commercial ELISA assays. Total cell counts and differential cell counts were performed in the BAL fluid samples. The group infected with A. costaricensis had lower total cell count in the BAL fluid when compared with the control group (0.11 x 10(6)cells/mL and 0.3 x 10(6)cells/mL, respectively; P = 0.013). BAL fluid IL-1beta levels in the infected group were significantly lower than in the control group (P = 0.008). IL-6 levels in BAL fluid were not different between the groups studied. We conclude that Angiostrongylus costaricensis infection in mice decreases pulmonary inflammatory response to OVA. PMID- 15279628 TI - Metallothionein expression in human neoplasia. AB - The metallothionein family is a class of low-molecular-weight, cysteine-rich proteins with high affinity for metal ions. Four major isoforms (metallothionein 1, -2, -3, and -4) have been identified in mammals, involved in many pathophysiological processes, including metal ion homeostasis and detoxification, protection against oxidative damage, cell proliferation and apoptosis, drug and radiotherapy resistance and several aspects of the carcinogenic process. In the present review we examine the expression of metallothionein in different human tumours and its correlation with histopathological variables, tumour cell proliferation or apoptosis, resistance to radiation or chemotherapy, patient survival and prognosis. A variable profile of metallothionein and its isoforms' expression has been observed in different cancer types. Although metallothionein expression has been implicated in carcinogenic evolution, its use as a marker of tumour differentiation, cell proliferation and prognosis predictor remains unclear. Detailed studies focused on the expression of metallothionein isoforms and isotypes in different tumour types could elucidate the role of this group of proteins in the carcinogenic process, delineating its possible clinical significance for the management of patients. PMID- 15279629 TI - The spectrum of pathological changes in severe acute respiratory syndrome (SARS). AB - AIMS: To analyse the lung pathology of severe acute respiratory syndrome (SARS) and correlate the findings with the time sequence of the disease. METHODS AND RESULTS: Ten patients with a clinical diagnosis of SARS, and virological confirmation of SARS coronavirus infection were identified. Histology in most cases showed diffuse alveolar damage, from early to late phases, and the changes corresponded to the time sequence. Other variable features include multinucleated giant cells, pneumocytes with cytomegaly and variable amounts of inflammatory cells and foamy macrophages. One case showed superimposed bronchopneumonia. No viral inclusions were found. Coronavirus particles were identified in pneumocytes by electron microscopy. CONCLUSIONS: The predominant pathological process of SARS is diffuse alveolar damage and, in patients who die from the disease, there is evidence of organization and fibrosis. There are apparently no histological features specific for this disease, and the aetiological diagnosis depends on virological and ultrastructural studies. PMID- 15279630 TI - Expression of TTF-1 and cytokeratins in primary and secondary epithelial lung tumours: correlation with histological type and grade. AB - AIMS: To assess cytokeratin (CK) and thyroid transcription factor (TTF)-1 expression in primary epithelial lung tumours by comparison with non-pulmonary carcinomas and to correlate it with their histological type and grade. METHODS AND RESULTS: Immunohistochemistry using antibodies against CKs 5/6, 7, 19, 20 and TTF-1 was applied to 165 primary and 37 secondary epithelial lung tumours. CK5/6 is a sensitive and specific marker of lung squamous carcinomas being positive in 100% of cases. CK7 is a common marker of primary lung adenocarcinomas (100% of cases) but with a lower specificity since it is also observed in other primary lung carcinomas (70% of large-cell neuroendocrine carcinomas, 40% of large-cell carcinomas, 23% of squamous carcinomas) but also in 27% of non-pulmonary adenocarcinomas. Addition of an anti-CK20 may be useful to prove or disprove the pulmonary origin of an adenocarcinoma when there is a history of colon cancer. CK19 is ubiquitous but a predominant or exclusive 'dot-like' pattern is very suggestive of high-grade neuroendocrine carcinoma. TTF-1 is a very sensitive and specific marker to document the pulmonary origin of an adenocarcinoma if a thyroid origin is excluded. Its expression in neuroendocrine lung tumours depends on the tumour grade. CONCLUSIONS: Immunohistochemical expression of CKs and TTF-1 may be correlated with histological type and grade of lung primary epithelial tumours and may allow them to be distinguished from non-pulmonary carcinomas. PMID- 15279631 TI - p57(KIP2) immunohistochemical staining of gestational trophoblastic tumours does not identify the type of the causative pregnancy. AB - AIM: To determine whether immunohistochemical staining for p57(KIP2), the product of the maternally expressed gene CDKN1C, can be used to differentiate between gestational trophoblastic tumours arising from a complete hydatidiform mole and those originating from non-molar pregnancies. METHODS: The immunohistochemical expression of p57(KIP2) was investigated in 23 cases of choriocarcinoma and 17 placental site trophoblastic tumours. Fourteen of the tumours examined were shown by DNA analysis to have arisen from complete hydatidiform moles and 26 from non molar pregnancies. RESULTS: Five of 11 (45%) post-complete hydatidiform mole choriocarcinomas and two of three (67%) post-complete hydatidiform mole placental site trophoblastic tumours were found to be p57(KIP2)+ and showed similar immunostaining characteristics to tumours that developed from non-molar pregnancies. Although there was a statistically significant reduction in the proportion of cases showing positive p57(KIP2) staining in post-complete hydatidiform mole tumours compared with those originating in non-molar pregnancies [proportion difference 0.35 [95% confidence interval (CI) 0.05, 0.61], P = 0.02], immunostaining did not provide diagnostically useful information to differentiate between these tumours in clinical practice. There was no significant difference between the extent of staining in choriocarcinoma versus placental site trophoblastic tumours [proportion difference 0.17 (95% CI - 12, 42), P = 0.19]. The majority of both types of gestational trophoblastic tumour were positive for the presence of the p57(KIP2) protein irrespective of their genetic origin. CONCLUSION: Immunostaining for p57(KIP2) fails to discriminate between gestational trophoblastic tumours that have arisen from complete hydatidiform moles and those that have originated from other types of pregnancy. PMID- 15279632 TI - Increased cell size and Akt activation in HER-2/neu-overexpressing invasive ductal carcinoma of the breast. AB - AIMS: To determine whether cell size is related to HER-2/neu status and/or to Akt activation in breast carcinomas. HER-2/neu overexpression is observed in 20-30% of invasive breast carcinomas with poor pronostic features, but little is known about the cell phenotype associated with HER-2/neu activation. Akt has been found to be involved in the HER-2/neu signal transduction pathway and Akt activation has been associated with increased cell size in various models. METHODS AND RESULTS: A case-control study of invasive ductal carcinoma of the breast was carried out, including 21 cases displaying HER-2/neu overexpression and 20 HER 2/neu negative controls. Cytoplasmic and nuclear sizes were measured on digitized histological pictures using cell image analysis software. Akt expression analysis was performed by immunohistochemistry on formalin-fixed histological sections using an anti-phosphorylated-Akt (Ser473) antibody. RESULTS: HER-2/neu overexpressing carcinomas had a mean nuclear size of 75 +/- 22.2 micro m(2) and a mean cytoplasmic size of 187 +/- 52.3 micro m(2). Both values were higher than the nuclear and cytoplasmic size of HER-2/neu-negative cases (nucleus = 58 +/- 24.5 micro m(2), cytoplasm = 133 +/- 56.6 micro m(2); P = 0.02 and P =0.003, respectively). Up to 75% of the tumours with a cell size over 140 micro m(2) were HER-2/neu-positive. Immunohistochemical Akt expression was observed in 19/40 (47.5%) cases. The immunoreactivity was localized in the cytoplasm in eight cases, on the cell membrane in four cases and at both sites in seven cases. One case was not interpretable. Comparison between HER-2/neu and Akt status showed that Akt was detectable at the cell membrane in 43% (9/21) of HER-2/neu-positive and in 10% (2/19) of HER-2/neu-negative cases (P = 0.02). CONCLUSIONS: HER-2/neu overexpression was consistently associated with increased cell size in invasive ductal carcinoma of the breast. This increase may be related to concomitant Akt activation. The assessment of activated pathways in HER-2/neu-overexpressing breast carcinomas may provide useful information for optimized individual HER 2/neu-targeted therapy. PMID- 15279633 TI - Surgical excision is warranted following a core biopsy diagnosis of mucocoele like lesion of the breast. AB - AIMS: Mucocoele-like lesions (MLLs) of the breast are unusual lesions in which mucin-filled ducts or cysts are accompanied by extrusion of mucin into surrounding stroma. A possible diagnosis of MLL may be suggested by the finding of mucin-filled ducts or cysts and/or stromal mucin in a core biopsy sample. Whether such findings should prompt immediate open diagnostic biopsy to exclude malignancy is currently uncertain, although this represents current practice in our institution. In this study we have reviewed 11 cases of possible MLL on core biopsy correlating both pathological and radiological findings in order to determine the risks of associated malignancy and whether excision is the most appropriate management option. METHODS AND RESULTS: Eleven cases of possible MLL presenting via the Breast Screening and Assessment Unit in Leeds since April 1999 were identified by review of pathological records. Histological slides, mammograms and ultrasound images were reviewed. Ten of the 11 had undergone open surgical biopsy for diagnosis. Three of the 10 (30%) proved to derive from malignant lesions. Two were ductal carcinoma in situ (DCIS) and one was an invasive mucinous carcinoma. All three cases had an associated atypical epithelial proliferation which, in a surgical excision, would be classified as atypical ductal hyperplasia (ADH) at least, as well as mucin in the core biopsy sample. The majority of possible MLLs presented radiologically as coarse calcification, but two of four (50%) which had a radiological mass subsequently proved malignant. Seven cases were without atypia on the core and all subsequently proved benign. Three of these, however, were associated with ADH on the excision biopsy. CONCLUSION: Surgical excision is warranted following a core biopsy suggestion of possible MLL when mucin-filled ducts or cysts and stromal mucin have been seen. The risk of malignancy is high when the core biopsy also contains an atypical epithelial proliferation (100% in our series) and also when there is an associated radiological mass lesion. In cases without atypia on the core a significant proportion of cases (43%) are associated with ADH on excision. PMID- 15279634 TI - The frequency of intratubular embryonal carcinoma: implications for the pathogenesis of germ cell tumours. AB - AIMS: To define the frequency and distribution of intratubular embryonal carcinoma (IEC) in an attempt to shed light on the pathogenesis of non seminomatous germ cell tumours (NSGCTs). Intratubular germ cell neoplasia of unclassified type (IGCNU) is common in NSGCT; however, IEC is rarely described. METHODS AND RESULTS: Sixty-two germ cell tumours were reviewed. Immunochemistry for CD30, placental alkaline phosphatase (PLAP) and c-kit was performed. The distribution, immunohistochemistry and morphology of the intratubular neoplasia were noted. All cases showed widespread IGCNU with PLAP and c-kit staining. CD30 showed strong focal intratubular positivity in 20/31 NSGCTs, 1/29 seminomas and 1/4 mixed seminomas/NSGCTs. In 17 of these cases, the CD30+ tubules were not easily identified as IEC on routine stains. These tubules were scanty in number and c-kit was negative, though some showed patchy PLAP staining. The cells within these tubules differed morphologically from IGCNU. CONCLUSIONS: IEC defined by CD30 positivity is not always easily identified on haematoxylin and eosin staining. We suggest that IEC is a common intermediate step between IGCNU and NSGCTs. The patchy and focal distribution of IEC suggests it may evolve quickly to invasive disease. PMID- 15279635 TI - Malignant transformation within benign adnexal skin tumours. AB - AIMS: To report five malignant trichogenic tumours arising in longstanding, previously benign adnexal neoplasms through malignant transformation. Malignant trichogenic adnexal tumours are extremely rare neoplasms. METHODS AND RESULTS: The patients were between 55 years and 79 years of age. Three of the tumours were located on the arms, two on the face. Three of our patients had a history of chronic lymphocytic leukaemia, one patient had a history of colonic adenocarcinoma. The duration of the tumour nodules was reported as between 20 and 40 years before sudden changes occurred. These changes included rapid growth, pain, itching, ulceration and bleeding. Histologically, all tumours were well circumscribed and encapsulated. There was a residual benign tumour component and morphological signs such as bone formation, dystrophic calcification and sclerosis suggesting long duration of the lesions. All patients except for one, who refused further clinical investigation due to her advanced age of 79 years, had an underlying systemic malignancy. CONCLUSIONS: The growth stimulus in these benign adnexal neoplasms resulting in malignant transformation may be attributed to the acquisition of additional genetic events or to immunosuppression due to an underlying neoplastic disease. Therefore, patients with systemic diseases or malignancy should be carefully examined and followed for sudden changes in pre existing benign cutaneous tumours. PMID- 15279636 TI - Nasopharyngeal intraepithelial lesion: latent Epstein-Barr virus infection with malignant potential. AB - AIMS: To study the morphology and immunohistochemical expression of nasopharyngeal intraepithelial lesions and to understand their place in nasopharyngeal carcinogenesis. METHODS AND RESULTS: Nine cases of nasopharyngeal intraepithelial lesion (NPIL) were diagnosed during nasopharyngeal biopsy screening for nasopharyngeal carcinoma (NPC). Two cases were associated with early invasion. All cases demonstrated specific histological features and consistent positivity on in-situ hybridization for Epstein-Barr virus (EBV) encoded RNA. Pure NPIL lesions showed low-grade morphology while lesions associated with early invasion were high grade. Immunohistochemical studies showed increased expression of bcl-2 and essentially negative findings for BZLF1 and LMP1. High-grade lesions had relatively stronger expression of bcl-2 and p53. CONCLUSIONS: NPIL harbours latent EBV infection and has malignant potential. Multiple steps are involved in its occurrence and progression. Low-grade and high grade lesions should be managed differently. PMID- 15279637 TI - Survivin expression is a negative prognostic marker in laryngeal squamous cell carcinoma and is associated with p53 accumulation. AB - AIMS: To investigate survivin expression in laryngeal squamous cell carcinoma (LSCC), its prognostic significance and relation to p53 status. Survivin is an inhibitor of apoptosis protein that is overexpressed in cancer. It has been implicated in both prevention of apoptosis and cell cycle regulation. It has been suggested that wild-type p53 represses survivin expression. METHODS AND RESULTS: Expression of survivin and p53 was analysed in 68 archival biopsy specimens of LSCC by immunohistochemistry. Survivin was detected in 67 of 68 LSCC cases; the proportion of survivin-positive cells varied from 8.2% to 100%. It was localized in the nucleus and/or cytoplasm of tumour cells. Of LSCC cases, 31.8% were p53+. The number of survivin-positive cells was significantly higher in the p53+ group. A high level of survivin expression and a supraglottic site of the tumour were two independent adverse prognostic factors in LSCC. CONCLUSIONS: Survivin is expressed in a varying proportion of cells in virtually all cases of LSCC. A high level of its expression predicts poor survival. Loss of wild-type p53 is a possible mechanism of survivin up-regulation in LSCC. PMID- 15279638 TI - Pleomorphic adenoma and carcinoma ex pleomorphic adenoma: immunohistochemical demonstration of the association between tenascin expression and malignancy. AB - AIMS: To investigate tenascin expression in salivary gland tumours. Tenascin is a matricellular protein that has been studied in several tumour types. Its expression has been correlated with tumour morphogenesis as well as with local invasiveness and tumour metastatic behaviour. METHODS AND RESULTS: The distribution pattern of tenascin in a series of 63 pleomorphic adenomas (PA) and 20 carcinomas ex- pleomorphic adenoma (Ca ex PA) was studied immunohistochemically. Ten normal adult salivary glands were used as controls. Tenascin surrounded the excretory ducts of normal adult salivary gland tissue. It was absent in the basement membrane compartment of both benign and malignant mixed tumours. In the interstitial compartment of the extracellular matrix, the fibro-hyaline type expressed tenascin in a statistically significantly (P < 0.001) lower number of PA cases (25%) in comparison with both malignant and benign areas of Ca ex PA (75% and 90%, respectively). In the Ca ex PA group, a statistically significantly difference (P < 0.001) was found in the frequency of tenascin deposits around aggregates of neoplastic cells between metastasizing (73%) and non-metastasizing neoplasms (0%). CONCLUSIONS: These findings strongly support the hypothesis that tenascin deposition is involved in the mechanisms of malignant transformation of pleomorphic adenomas into carcinomas as well as being associated with clinical disease progression. PMID- 15279639 TI - Dome magnifiers. PMID- 15279640 TI - Histiocyte-rich dermatoses in two patients with acute lymphocytic leukaemia treated with cytarabine. PMID- 15279641 TI - Recurrent polymorphous haemangioendothelioma. PMID- 15279642 TI - Correlation between Alcian blue-periodic acid-Schiff stain and immunohistochemical expression of mucin 2 in Barrett's oesophagus. PMID- 15279643 TI - Spirochaetes within the cysts of pneumatosis coli. PMID- 15279644 TI - E-cadherin and catenin expression in normal and neoplastic endocervical glandular epithelium. PMID- 15279645 TI - 'Ectopic' ectopic hamartomatous thymoma. PMID- 15279647 TI - Bone remodeling during prenatal morphogenesis of the human mental foramen. AB - From a morphogenetic point of view, the mental foramen of the mandible is a highly suitable model to study the interactions of different tissues such as nerves, vessels, mesenchymal cells, cartilage, and bone. In previous work, we provided a three-dimensional description of the mental foramen at different developmental stages, and now we complement those studies with a three dimensional visualization of different bone remodeling activities around the mental foramen. Histological serial sections of human embryos and fetuses, ranging in size from 25 to 117 mm crown-rump-length (CRL), were used to characterize the bone remodeling activity (apposition, inactivity, and resorption). We quantified and reconstructed this activity in three dimensions, and included information on the spatial relationship of the nerves, vessels, and dental primordia. In general, the mandible showed strong apposition at its outer surfaces. The brim of the mental foramen, however, displayed changing remodeling activity at different stages. In the depth of the bony gutter, which provides space for the nerve and the blood vessels, we found bone resorption beneath the inferior alveolar vein. Bone was also resorbed in proximity to the dental primordia. In future studies, we will relate gene expression data to these morphological findings in order to identify molecular mechanisms that regulate this complex system. PMID- 15279648 TI - Prediction of success and failure of behavior modification as treatment for dental anxiety. AB - Behavior modification techniques are effective in the treatment of extreme dental anxiety, but their success is by no means absolute. In the present article, the Corah Dental Anxiety Scale (DAS), the self-report symptom inventory SCL-90R and a questionnaire accessing subjects' daydreaming styles (the Short Imaginal Process Inventory) were used to develop possible predictive measures for success and failure of behavior modification as a treatment for dental fear. The patients' level of distractibility and mind wandering, initial dental anxiety and somatization significantly predicted the success of therapy. The odds ratio indicated that the risk of therapy failure increased about 11 times with an increase of one scale of the Poor Attention Control Scale, about three times with an increase of one level of the mean DAS score, and 0.17 times with an increase of one level of somatization. The predictive value of the chosen scales was 80%. Thus, the use of these scales as part of an initial admittance process for patients who suffer from dental anxiety can enhance our ability to better recognize patients who are prone to fail behavior therapy as treatment for their problem, and enable their referral for other possible modes of treatment. PMID- 15279649 TI - The 3020insC mutation of the NOD2/CARD15 gene in patients with periodontal disease. AB - The 3020insC mutation of the NOD2/CARD15 gene leads to impaired activation of nuclear factor-kappa B (NF-kappaB) in vitro. As the destruction of periodontal tissue is mediated via activation of NF-kappaB, with subsequent transcription of proinflammatory cytokines, the c-insertion mutation of the NOD2/CARD15 gene might contribute to the proposed genetic background of periodontitis. The present study analysed the frequency of this mutation in 80 patients with chronic periodontal disease and 122 healthy controls. The 3020insC mutation was identified by employing the polymerase chain reaction followed by restriction fragment length polymorphism analysis. The prevalence of the 3020insC mutation of the NOD2/CARD15 protein in patients with periodontitis was 1.9% (three of 160) and that for the control group was 2.0% (five of 244) (P = 0.942). Hence, unlike in Crohn's disease, the 3020insC mutation of the NOD2/CARD15 gene does not seem to influence the pathophysiology of periodontitis. PMID- 15279650 TI - Supramolecular pellicle precursors. AB - Saliva contacting with solid surfaces in the oral cavity forms a coat termed the pellicle. However, its formation is not fully understood. Although indications for the existence of supramolecular pellicle precursors have been reported, the possible relationship between them and pellicle formation is unclear. This study investigates the ability of supramolecular precursors to form the pellicle via interaction with a solid surface. Fixed and unfixed salivary globes were spread onto a microscopic grid and examined by transmission electron microscopy. Biochemical pretreatment of saliva revealed that neither disulphide links nor transglutaminase-mediated crosslinking are responsible for maintaining the salivary globes, i.e. supramolecular pellicle precursors. However, the detergent, sodium dodecyl sulphate, caused dissociation of the salivary globes, indicating their micellar nature. Saliva contacting a formvar film for 10 s did not form a complete surface coating, but single supramolecular pellicle precursors were observed attached to the surface. After extension of the contact time to 60 s, a surface layer was formed by clustering and fusion of the supramolecular pellicle precursors. The supramolecular pellicle precursors are unstable and attain a thermodynamically more favourable state by adhesion to a solid surface. As a result, a layer of fused precursors covering the solid surface is formed -- the salivary pellicle. PMID- 15279651 TI - In vitro antibacterial effect of calcium hydroxide combined with chlorhexidine or iodine potassium iodide on Enterococcus faecalis. AB - Several studies have shown a higher success rate of root canal therapy when the canal is free from bacteria at the time of obturation. Treatment strategies that are designed to eliminate this microflora should include agents that can effectively disinfect the root canal. Enterococcus faecalis is often associated with persistent endodontic infections. While in vivo studies have indicated calcium hydroxide to be the most effective all-purpose intracanal medicament, iodine potassium iodide (IKI) and chlorhexidine (CHX) may be able to kill calcium hydroxide-resistant bacteria. Supplementing the antibacterial activity of calcium hydroxide with IPI or CHX preparations was studied in bovine dentine blocks. While calcium hydroxide was unable to kill E. faecalis in the dentine, calcium hydroxide combined with IKI or CHX effectively disinfected the dentine. The addition of CHX or IKI did not affect the alkalinity of the calcium hydroxide suspensions. It may be assumed that combinations also have the potential to be used as long-term medication. Cytotoxicity tests using the neutral red method indicated that the combinations were no more toxic than their pure components. PMID- 15279652 TI - Effects of an inducible nitric oxide synthase inhibitor on experimentally induced rat pulpitis. AB - Nitric oxide (NO) is a biological effector molecule involved in a large variety of reactions and, as synthesized by inducible nitric oxide synthase (iNOS), has many important roles in inflammatory conditions. This study aimed to evaluate the anti-inflammatory effects of an iNOS-specific inhibitor, N-(3 (aminomethyl)benzyl)acetamidine (1400W), on experimentally induced rat pulpitis in the upper incisors of 6-wk-old male Wistar rats. 1400W (1 mg kg(-1)), the non specific NOS inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME, 50 mg kg(-1)), or sterile saline (control) were administered before the application of lipopolysaccharide (LPS). Rats were killed 3, 6, 9, 12, 24, 48, and 72 h after LPS application, and immunocompetent cells were detected immunohistochemically. The numbers of granulocytes infiltrating into the pulp were significantly depressed in the 1400W group compared with the saline and L-NAME groups. The kinetics of the macrophages and Ia(+) cells in the 1400W group were similar to those in the L-NAME group, while the maximum numbers in both groups were significantly reduced compared with those in the saline group. These results suggest that NO may be responsible for the infiltration of immunocompetent cells in the progress of pulpitis, and that 1400W is a promising candidate for controlling pulpal inflammatory reactions. PMID- 15279653 TI - Mechanism of Fas-mediated cell death and its enhancement by TNF-alpha in human salivary gland adenocarcinoma cell line HSG. AB - Fas-mediated cell death in a human salivary gland adenocarcinoma cell line (HSG) was induced by treatment of the cells with agonistic anti-Fas antibody (CH-11), and this cell death was enhanced by pretreatment with tumor necrosis factor alpha (TNF-alpha). The mode of cell death was apoptosis, because it was accompanied by caspase activation and the cleavage of poly(ADP-ribose) polymerase. The TNF-alpha treatment of the cells increased the expression of Fas, which was accompanied by the activation of nuclear factor kappaB (NFkappaB). These results suggest that the enhancement of the apoptosis caused by TNF-alpha resulted from increased sensitivity of the HSG cells to CH-11-mediated apoptosis due to induction of Fas protein by TNF-alpha via the activation of NFkappaB. In order to elucidate the apoptosis signaling pathway, we examined the effect of various caspase inhibitors on the apoptosis induced by CH-11. Fas-mediated apoptosis of HSG cells was slightly inhibited by the caspase-9 inhibitor although it was mainly inhibited by that for caspase-8. Based on this finding, we consider CH-11-induced apoptosis in HSG cells to be mainly mediated by the type I death signaling pathway that is caused by a caspase cascade initiated by the activation of caspase-8 at the death inducing signaling complex (DISC). PMID- 15279654 TI - Undermining of enamel as a mechanism of abfraction lesion formation: a finite element study. AB - Many workers have suggested that abfraction lesion formation is caused by the physical overloading of enamel. However, an alternative mechanism, involving undermining of the cervical enamel along the amelodentinal junction (ADJ), may be a more realistic explanation. The aim of this study was to examine what effect undermining of the buccal cervical enamel would have on the stress distribution in upper teeth. Two-dimensional plain strain finite element meshes of an upper incisor, canine and first premolar and the supporting periodontal ligament and alveolar bone were developed. Each tooth was loaded with an oblique 100 N load, and the nodal maximum principal stresses (MPS) along a buccal horizontal sampling plane 1.1 mm above the amelo-cemental junction was measured. A discontinuity between the cervical enamel and dentine elements was then introduced (0.1 mm wide) using gap elements. The vertical extent of this defect varied from 0.1 to 0.5 mm. The value of the MPS varied from 1.8 to 209 Mpa, and the lowest values were found for the intact teeth (range 0.6-30.3 MPa). The discontinuity caused a dramatic increase in the numerical values of the MPS, and in many instances these exceeded the known failure stress for enamel. PMID- 15279655 TI - The adhesion between fiber posts and root canal walls: comparison between microtensile and push-out bond strength measurements. AB - Aim of the study was to compare the trimming and non-trimming variants of the microtensile technique with the 'micro' push-out test in the ability to measure accurately the bond strength of fiber posts luted inside root canals. In 15 endodontically treated teeth (Group A), fiber posts were cemented with Excite DSC in combination with Variolink II (Ivoclar-Vivadent). In 15 roots RelyX Unicem (3M ESPE) was used for fiber post luting (Group B). Within each group, the bond strength of cemented fiber posts was assessed with the trimming and non-trimming microtensile technique, as well as with the push-out test. The great number of premature failures (16.9% in Group A, 27.5% in Group B) and the finding of high standard deviation values make questionable the reliability of the trimming microtensile technique. With the non-trimming microtensile technique, only five sticks were obtained from a total of six roots. The remaining specimens failed prematurely during the cutting phase. With the push-out test no premature failure occurred, the variability of the data distribution was acceptable, and regional differences in bond strength among root levels could be assessed. Relatively low values of bond strength were, in general, recorded for luted fiber posts. In conclusion, when measuring the bond strength of luted fiber posts, the push-out test appears to be more dependable than the microtensile technique. PMID- 15279656 TI - Evidence of chemisorption of maleic acid to enamel and hydroxyapatite. AB - Maleic acid has been used as an etchant or non-rinse conditioner in adhesive dentistry. However, the inherent mechanisms of the interaction of maleic acid with hydroxyapatite/enamel have never been fully elucidated. The purpose of this study was to provide evidence for the chemisorption of maleic acid onto hydroxyapatite/enamel, and to identify the reaction products obtained following the interaction of maleic acid with hydroxyapatite. Hydroxyapatite particles were dissolved in a 15% (w/v) aqueous solution of maleic acid (pH = 0.98). Half of the solution was dried to obtain a desiccated mixture. This mixture, hydroxyapatite, maleic acid and self-prepared calcium maleate were analysed by X-ray diffraction (XRD). Acetone was added to the other half of the solution to obtain a precipitate. This precipitate, hydroxyapatite, maleic acid, unetched enamel and maleic acid-etched enamel were analysed by X-ray photoelectron spectroscopy (XPS). The precipitate was also analysed by (1)H NMR. A new binding energy, indicating carboxylate groups, was detected by XPS on the precipitate and maleic acid-etched enamel surface. XRD data indicated the formation of calcium maleate and calcium hydrogen phosphate after the reaction. NMR data revealed that one carboxylic group of maleic acid reacted with hydroxyapatite. Hence, maleic acid can chemisorb to hydroxyapatite and enamel via ionic interactions. PMID- 15279657 TI - In vivo degradation of resin-dentin bonds produced by a self-etch vs. a total etch adhesive system. AB - The purpose of this study was to evaluate the long-term durability of in vivo bond strengths and the morphological changes of interfaces between dentin and two adhesive systems. Class V preparations were prepared on the facial surfaces of 14 intact teeth of two monkeys and restored with a combination of Unifil Bond/Z250 or Single Bond/Z250. One year later, 10 additional teeth were restored with the same materials and the monkeys were killed after 24 h. All of the restored teeth were subjected to microtensile bond strength ( micro TBS) testing. The debonded surfaces of the dentin sides were morphologically observed using Fe-scanning electron microscopy (SEM), as were the polished cross-sections of resin-dentin interfaces. For both Unifil Bond and Single Bond, the micro TBS at 24 h was significantly higher than that at 1 yr. Fe-SEM observations of polished cross sectioned and fractured surfaces showed that porosity within the hybrid layers produced by Single Bond increased over time. However, the interface produced by Unifil Bond revealed no noticeable changes in morphology between 24-h and 1-yr specimens. It is concluded that even though the bond strengths of both adhesive systems declined over time, the bonding interface using self-etching primers was relatively stable over time compared to the wet bonding system. PMID- 15279658 TI - The effect of a self-etching primer on the continuous demineralization of dentin. AB - Self-etching primers (SEP), used for adhesion of resin restorations, contain an acidic monomer that is not rinsed off after application; therefore, residual acid could further demineralize dentin, jeopardizing adhesion. This study evaluated whether dentin demineralization continues after a 20-s application of a SEP and also after polymerization of the adhesive. Surface recession was measured, using atomic force microscopy (AFM), between the masked surface and dentin etched with SEP (Clearfil SE Bond) or phosphoric acid (pH 1.94) immediately, 5 min, 2 h or 12 h after application. AFM-based nanoindentations were performed to determine mechanical property profiles across resin-bonded interfaces of two sequence groups: SG 1, where the adhesive was applied and polymerized immediately after application of the primer and specimens were immersed in Hank's solution after 5 min, 2 h and 12 h; or SG 2, where specimens were placed in an incubator before the application of the adhesive, for the same time periods. Significant surface recession occurred over time for all groups, except for SEP before desiccation. Nanoindentations yielded decreased hardness and elastic modulus below the hybrid layer after application of the initial primer, even after polymerization of the adhesive. The results reveal a demineralized dentin zone below the hybrid layer formed by the SEP, not fully protected by the adhesive, which could jeopardize bond strength. PMID- 15279659 TI - Effects of a fluoride etchant on resin bonding to titanium-aluminum-niobium alloy. AB - This investigation was carried out in order to evaluate ammonium hydrogen fluoride (AHF) and cupric chloride (CC) as components of a metal etchant. The surface of cast titanium-aluminum-niobium (Ti-6Al-7Nb) was air-abraded with alumina, etched for 10 s, and rinsed with water. A phosphate or a thiophosphate primer was applied to the bonding area, and an acrylic rod was bonded to the specimen with a tri-n-butylborane-initiated self-curing luting agent. Shear bond strengths were determined after thermocycling (4 degrees C and 60 degrees C) for 10,000 cycles. The average bond strength was significantly influenced by thermocycling, AHF, and primer, but was not influenced by CC. The maximum average bond strengths were obtained when the etchant consisted of 5mass% AHF, with and without 0.3mass% CC. Microphotographs showed that numerous micropits were created on the etched surface, suggesting increased micromechanical retention. In conclusion, chemical etching with 5mass% AHF significantly improved the durability of resin bonding to Ti-6Al-7Nb. PMID- 15279660 TI - Management of haemophilia and its complications in developing countries. AB - Eighty per cent of people with haemophilia live in developing countries, where technical expertise and health care facilities may be less than optimal. Haemophilia is a relatively rare disease and high-cost, technology-intensive therapy is not a high priority for the governments of developing countries. The rapid spread of transfusion-related viral infections in many developing countries presents further problems for haemophiliacs. However, it is possible to manage haemophiliacs patients with limited resources. Strategies for conserving factor concentrates, include education of doctors and patients, prenatal diagnosis, increasing the use of anti-fibrinolytic agents, physiotherapy, the use of fibrin glue, and simple orthotics and prosthetic measures. These approaches are helpful in the majority of these patients. Meanwhile, with the help of the World Federation of Haemophilia (WFH), all developing countries are gradually improving management skills for this relatively rare but disabling disease. The present review broadly describes the management of various aspects of severe haemophilia in developing countries. PMID- 15279661 TI - Comparison of haemoglobinometry by WHO Haemoglobin Colour Scale and copper sulphate against haemiglobincyanide reference method. AB - Although estimation of haemoglobin is essential for diagnosing anaemia and assessing its severity, many health centres in developing countries do not have the facilities for haemoglobinometry. The WHO Haemoglobin Colour Scale (HCS) method is a simple and inexpensive clinical device that was recently developed in order to diagnose anaemia in such centres. In Indonesia, the copper sulphate specific gravity method is used for blood donor screening and also in primary health clinics in the rural and remote areas. In this study, the HCS method is compared with the copper sulphate method and with an earlier paper scale, the Tallquist method, against the standard haemiglobincyanide spectrophotometric method. The HCS method showed an acceptable level of precision and accuracy for use as a reliable screening tool to diagnose anaemia in patients and also for blood donor screening. PMID- 15279662 TI - Absence of macrocytic anaemia in Alzheimer's disease. AB - There is an association between Alzheimer's disease (AD) and low serum levels of vitamin B12 and folic acid. Patients also have elevated serum levels of homocysteine and disease progression might therefore be associated with the development of a macrocytic anaemia. We investigated the relationship between disease duration, homocysteine and haematological indices in patients with clinically diagnosed AD and healthy elderly controls. Haemoglobin and platelet counts fell only slightly with increasing dementia duration, but there were no other changes in haematological indices. In particular, macrocytosis and red cell distribution width were unrelated to disease duration and no patients were anaemic. Our results support previous observations that the neurological and haematological features of B12 and folate deficiency are often unrelated in these patients. PMID- 15279663 TI - Plasma basic fibroblast growth factor and bone marrow fibrosis in clonal myeloproliferative disorders. AB - Basic fibroblast growth factor (bFGF) is an important growth factor involved in clonal hematopoietic expansion, neoangiogenesis, and bone marrow fibrosis, all of which are important pathobiologic features of clonal chronic myeloproliferative disorders (CMPD) and myelodysplastic syndromes (MDS). The aim of this study was to assess circulating bFGF concentrations in patients with CMPD and MDS with respect to the presence of bone marrow fibrosis in histopathologic examination. The study group comprised 18 patients with CMPD (six female, 12 male; median age 50 years), seven patients with MDS (one female, six male; median age 66 years) and 10 healthy adults as controls (four female, six male; median age 29 years). CMPD group included six chronic myelogenous leukemia (CML), seven essential thrombocythemia (ET), three polycythemia vera (PV), two agnogenic myeloid metaplasia (AMM). All seven MDS patients were the FAB subtype of refractory anemia (RA). Bone marrow biopsy sections stained with hematoxylin and eosin (H & E) and for reticulin were examined for the presence of fibrosis. The median plasma bFGF level was 18.2 pg/ml (interquartile range, IQR: 15.2-26.7) in patients with CMPD, 18.0 pg/ml (IQR: 15.8-26.4) in patients with MDS, 13.6 pg/ml (IQR: 9.9-20.0) in the control group. The bFGF levels were significantly higher in patients with CMPD in comparison with the healthy control group (P = 0.031). Circulating bFGF tended to be significantly lower in relation to the development of marrow fibrosis (P = 0.028). The complicated interactions of bFGF and fibrosis in the context of CMPD may be either 'cause' or 'effect'. The bFGF might represent an important link between angiogenesis, fibrosis, and clonal neoplastic hematopoiesis during the development of CMPD. PMID- 15279664 TI - Systemic haemostasis after intermittent pneumatic compression. Clues for the investigation of DVT prophylaxis and travellers thrombosis. AB - Intermittent pneumatic compression (IPC) is known to provide effective prophylaxis against post-surgical deep-vein thrombosis (DVT), and other procedures based on reducing venous stasis have been promoted recently to minimize the risk of thromboembolism after long-haul travel ('travellers thrombosis'). This study sought to measure the effects of IPC on systemic haemostasis, which are currently disputed. IPC was applied for 120 min on 21 male, non-smoking volunteers ranging in age from 19 to 47 years. IPC promoted a significant increase in global fibrinolytic potential. Levels of urokinase plasminogen activator activity (uPA) measured using an amidolytic assay were raised after IPC. However, enzyme-linked immunosorbent assays (ELISA) of uPA antigen, and the activities of tissue plasminogen activator (tPA) and plasminogen activator inhibitor 1 (PAI-1) were not statistically different from those in control experiments. IPC led to highly significant falls in factor VIIa, associated with increased levels of tissue factor pathway inhibitor (TFPI). IPC enhances fibrinolysis and suppresses procoagulant activation. Measurements of specific fibrinolytic components do not reflect overall fibrinolytic activity and are highly dependent on the method of assay. The results provide important clues for detailed studies of the effects of haemodynamics on systemic haemostasis. PMID- 15279665 TI - Evaluation of refrigerated platelet concentrates supplemented with low doses of second messenger effectors. AB - With the goal of producing haemostatically effective platelet concentrates (PCs) with a longer shelf-life, we aimed to identify a simple combination of platelet inhibitors, with a low pharmacological load, which could avoid the unacceptable loss of platelets stored under refrigerated conditions. PCs stored with different combinations of second messenger effectors were analysed at days 5, 10 and 15 of storage and compared with those supplemented with ThromboSol--a combination of six platelet inhibitors that protects cells from cold damage. The following parameters were analysed: platelet counts, biochemical parameters (glucose, pH, bicarbonate, lactate), cell lysis (lactic dehydrogenase, LDH), membrane glycoproteins (GPs), platelet aggregation, fibrinogen binding and hypotonic shock response. We characterized the combination of amiloride and sodium nitroprusside (at 1/2 the dose included in ThromboSol). This was found to be similar to ThromboSol and superior to nontreated units in the prevention of cold-induced platelet aggregation at day 15 of storage (maintenance of 78% and 80% of initial platelet counts, respectively), preservation of GPIbalpha (11% and 12% better maintenance of mean fluorescence intensity compared with control units, respectively), and reduced cell lysis (13% and 11% decrease in supernatant LDH, respectively). The reduced pharmacological load with the identified solution compared with ThromboSol is an argument in favour of the potential use of these agents when designing strategies to improve PC storage. PMID- 15279666 TI - Pseudobasophilia as an erroneous white blood cell differential count with a discrepancy between automated cell counters: report of two cases. AB - We report two cases that showed erroneous white blood cell differential counts by automated cell counters. Each case showed an interesting discrepancy of differential count between cell counters, and marked pseudobasophilia was observed by one of the two counters. The first patient was a 44-year-old female who suffered from multiple myeloma for more than one and a half years. Increased myeloma cells (43%) in peripheral blood were counted as basophils by the ADVIA 120, and as monocytes by SE-9000, respectively. The second patient was a 72-year old female diagnosed as having chronic myelomonocytic leukemia. Dysgranulopoietic neutrophils (50%) and monocytes (31%) were increased in the peripheral blood. Dysgranulopoietic neutrophils were counted as basophils by STKS. In contrast, about half of the increased monocytes were counted as neutrophils by the ADVIA 120. These interesting findings highlight the importance of microscopic examination of the blood film in routine laboratory practice, and automated cell counters, especially for the hematologic patients, cannot completely substitute for it. These results also imply that at least some subpopulations with different membrane or cytoplasmic properties may exist even in the similarly classified cells. PMID- 15279667 TI - Evans' syndrome following autologous peripheral blood stem cell transplantation for non-Hodgkin's lymphoma. AB - A 34-year-old woman with non-Hodgkin's lymphoma received high-dose chemotherapy followed by autologous peripheral blood stem cell transplantation (PBSCT). She developed Evans' syndrome, the association of immune thrombocytopenia and autoimmune hemolytic anemia, 49 days after transplantation. Multiple autoimmune disorders may occur concurrently after autologous PBSCT. PMID- 15279668 TI - Fatal peripheral neuropathy following FLA chemotherapy. AB - We discuss a case with significant progressive peripheral neurological deterioration following administration of both fludarabine and cytarabine as part of the FLA (fludarabine and cytarabine) regime. Of particular interest is that toxicity only occurred during the second course of FLA and sixth course of Ara-C containing chemotherapy. At this point, a new antifungal agent had been commenced, suggesting a possible drug interaction enhancing the risk of known neurological toxicity with this regime. PMID- 15279669 TI - Coexistence of congenital red cell pyruvate kinase and band 3 deficiency. AB - The authors report the case of a 9-year-old Caucasian girl, born in northern Portugal, with chronic nonspherocytic haemolytic anaemia and without family history of anaemia. The aethiological study of this anaemia revealed pyruvate kinase deficiency (PKD), because of two previously described mutations (426Arg- >Trp and 510Arg-->Gln). Since the blood smear revealed features not fully compatible with PKD diagnosis, additional tests were performed for the propositus and her parents, namely red blood cell membrane protein analysis. A decrease in proteins band 3 (15%) and 4.2 (18%) was found in the propositus. Her father presented only a decrease in band 3 (11%). Coexistence of PKD and erythrocyte membrane proteins deficiency in the same patient is very uncommon. Our findings suggest that a careful blood smear observation may lead to the identification of a combined deficiency in erythrocyte membrane proteins and enzymopathies. PMID- 15279670 TI - Increase of immature reticulocyte fraction in myelodysplastic syndromes. PMID- 15279671 TI - Overexpression of p16INK4A and p14ARF in haematological malignancies. PMID- 15279672 TI - Automated haemoglobinopathy screening. PMID- 15279674 TI - "Is there nothing more practical than a good theory?": Why innovations and advances in health behavior change will arise if interventions are used to test and refine theory. AB - Theoretical and practical innovations are needed if we are to advance efforts to persuade and enable people to make healthy changes in their behavior. In this paper, I propose that progress in our understanding of and ability to promote health behavior change depends upon greater interdependence in the research activities undertaken by basic and applied behavioral scientists. In particular, both theorists and interventionists need to treat a theory as a dynamic entity whose form and value rests upon it being rigorously applied, tested and refined in both the laboratory and the field. To this end, greater advantage needs to be taken of the opportunities that interventions afford for theory-testing and, moreover, the data generated by these activities need to stimulate and inform efforts to revise, refine, or reject theoretical principles. PMID- 15279675 TI - Estimating age conditional probability of developing disease from surveillance data. AB - Fay, Pfeiffer, Cronin, Le, and Feuer (Statistics in Medicine 2003; 22; 1837-1848) developed a formula to calculate the age-conditional probability of developing a disease for the first time (ACPDvD) for a hypothetical cohort. The novelty of the formula of Fay et al (2003) is that one need not know the rates of first incidence of disease per person-years alive and disease-free, but may input the rates of first incidence per person-years alive only. Similarly the formula uses rates of death from disease and death from other causes per person-years alive. The rates per person-years alive are much easier to estimate than per person years alive and disease-free. Fay et al (2003) used simple piecewise constant models for all three rate functions which have constant rates within each age group. In this paper, we detail a method for estimating rate functions which does not have jumps at the beginning of age groupings, and need not be constant within age groupings. We call this method the mid-age group joinpoint (MAJ) model for the rates. The drawback of the MAJ model is that numerical integration must be used to estimate the resulting ACPDvD. To increase computational speed, we offer a piecewise approximation to the MAJ model, which we call the piecewise mid-age group joinpoint (PMAJ) model. The PMAJ model for the rates input into the formula for ACPDvD described in Fay et al (2003) is the current method used in the freely available DevCan software made available by the National Cancer Institute. PMID- 15279676 TI - Similar group mean scores, but large individual variations, in patient-relevant outcomes over 2 years in meniscectomized subjects with and without radiographic knee osteoarthritis. AB - BACKGROUND: Epidemiological studies have, so far, identified factors associated with increased risk for incident or progressive OA, such as age, sex, heredity, obesity, and joint injury. There is, however, a paucity of long-term data that provide information on the nature of disease progression on either group or individual levels. Such information is needed for identification of study cohorts and planning of clinical trials. The aim of the study was, thus, to assess the variation in pain and function on group and individual level over 2 years in previously meniscectomized individuals with and without radiographic knee osteoarthritis (OA). METHODS: 143 individuals (16% women, mean age at first assessment 50 years [range 27-83]) were assessed twice; approximately 14 and 16 years after isolated meniscectomy, with a median interval of 2.3 years (range 2.3 3.0). Radiographic OA (as assessed at the time of second evaluation) was present in the operated knee in 40%, and an additional 19% had a single osteophyte grade 1 in one or both of the tibiofemoral compartments. Subjects completed the self administered and disease-specific Knee injury and Osteoarthritis Outcome Score (KOOS). RESULTS: There were no significant changes in the group mean KOOS subscale scores over the 2-year period. However, a great variability over time was seen within individual subjects. Out of 143 subjects, 16% improved and 12% deteriorated in the subscale Pain, and 13% improved and 14% deteriorated in the subscale ADL > or = 10 points (the suggested threshold for minimal perceptible clinical change). Similar results were seen for remaining subscales. CONCLUSION: Group mean scores for this study cohort enriched in incipient and early-stage knee OA were similar over 2 years, but pain, function and quality of life changed considerably in individuals. These results may be valid also for other at risk groups with knee OA, and motivate further careful examination of the natural history of OA, as well as properties of the OA outcome instruments used. Longitudinal outcome data in OA studies need to be analyzed both on an individual and a group level. PMID- 15279677 TI - Use and comparison of different internal ribosomal entry sites (IRES) in tricistronic retroviral vectors. AB - BACKGROUND: Polycistronic retroviral vectors that contain several therapeutic genes linked via internal ribosome entry sites (IRES), provide new and effective tools for the co-expression of exogenous cDNAs in clinical gene therapy protocols. For example, tricistronic retroviral vectors could be used to genetically modify antigen presenting cells, enabling them to express different co-stimulatory molecules known to enhance tumor cell immunogenicity. RESULTS: We have constructed and compared different retroviral vectors containing two co stimulatory molecules (CD70, CD80) and selectable marker genes linked to different IRES sequences (IRES from EMCV, c-myc, FGF-2 and HTLV-1). The tricistronic recombinant amphotropic viruses containing the IRES from EMCV, FGF-2 or HTLV-1 were equally efficient in inducing the expression of an exogenous gene in the transduced murine or human cells, without displaying any cell type specificity. The simultaneous presence of several IRESes on the same mRNA, however, can induce the differential expression of the various cistrons. Here we show that the IRESes of HTLV-1 and EMCV interfere with the translation induced by other IRESes in mouse melanoma cells. The IRES from FGF-2 did however induce the expression of exogenous cDNA in human melanoma cells without any positive or negative regulation from the other IRESs present within the vectors. Tumor cells that were genetically modified with the tricistronic retroviral vectors, were able to induce an in vivo anti-tumor immune response in murine models. CONCLUSION: Translation of the exogenous gene is directed by the IRES and its high level of expression not only depends on the type of cell that is transduced but also on the presence of other genetic elements within the vector. PMID- 15279678 TI - Investigating the utility of combining phi29 whole genome amplification and highly multiplexed single nucleotide polymorphism BeadArray genotyping. AB - BACKGROUND: Sustainable DNA resources and reliable high-throughput genotyping methods are required for large-scale, long-term genetic association studies. In the genetic dissection of common disease it is now recognised that thousands of samples and hundreds of thousands of markers, mostly single nucleotide polymorphisms (SNPs), will have to be analysed. In order to achieve these aims, both an ability to boost quantities of archived DNA and to genotype at low costs are highly desirable. We have investigated phi29 polymerase Multiple Displacement Amplification (MDA)-generated DNA product (MDA product), in combination with highly multiplexed BeadArray genotyping technology. As part of a large-scale BeadArray genotyping experiment we made a direct comparison of genotyping data generated from MDA product with that from genomic DNA (gDNA) templates. RESULTS: Eighty-six MDA product and the corresponding 86 gDNA samples were genotyped at 345 SNPs and a concordance rate of 98.8% was achieved. The BeadArray sample exclusion rate, blind to sample type, was 10.5% for MDA product compared to 5.8% for gDNA. CONCLUSIONS: We conclude that the BeadArray technology successfully produces high quality genotyping data from MDA product. The combination of these technologies improves the feasibility and efficiency of mapping common disease susceptibility genes despite limited stocks of gDNA samples. PMID- 15279679 TI - Injuries at the Canadian National Tae Kwon Do Championships: a prospective study. AB - BACKGROUND: The purpose of this prospective study was to assess the injury rates in male and female adult Canadian Taekwondo athletes relative to total number of injuries, type and body part injured. METHODS: Subjects (219 males, 99 females) participated in the 1997 Canadian National Taekwondo Championships in Toronto, Canada. Injuries were recorded on an injury form to documents any injury seen and treatment provided by the health care team. These data were later used for this study. The injury form describes the athlete and nature, site, severity and mechanism of the injury. RESULTS: The overall rate of injuries was 62.9/1,000 athlete-exposures (A-E). The males (79.9/1,000 A-E) sustained significantly more injuries than the females (25.3/1,000 A-E). The lower extremities were the most commonly injured body region in the men (32.0 /1,000 A-E), followed by the head and neck (18.3/1,000 A-E). Injuries to the spine (neck, upper back, low back and coccyx) were the third most often injured body region in males (13.8/1,000 A-E). All injuries to the women were sustained to the lower extremities. The most common type of injury in women was the contusion (15.2/1,000 A-E). However, men's most common type of injury was the sprain (22.8/1,000 A-E) followed by joint dysfunction (13.7/1,000 A-E). Concussions were only reported in males (6.9/1,000 A-E). Compared to international counterparts, the Canadian men and women recorded lower total injury rates. However, the males incurred more cerebral concussions than their American colleagues (4.7/1,000 A-E). CONCLUSIONS: Similar to what was found in previous studies, the current investigation seems to suggest that areas of particular concern for preventive measures involve the head and neck as well as the lower extremities. This is the first paper to identify spinal joint dysfunction. PMID- 15279680 TI - 8-Cl-Adenosine enhances 1,25-dihydroxyvitamin D3-induced growth inhibition without affecting 1,25-dihydroxyvitamin D3-stimulated differentiation of primary mouse epidermal keratinocytes. AB - BACKGROUND: Epidermal keratinocytes continuously proliferate and differentiate to form the mechanical and water permeability barrier that makes terrestrial life possible. In certain skin diseases, these processes become dysregulated, resulting in abnormal barrier formation. In particular, skin diseases such as psoriasis, actinic keratosis and basal and squamous cell carcinomas are characterized by hyperproliferation and aberrant or absent differentiation of epidermal keratinocytes. We previously demonstrated that 8-Cl-adenosine (8-Cl Ado) can induce keratinocyte growth arrest without inducing differentiation. RESULTS: To determine if this agent might be useful in treating hyperproliferative skin disorders, we investigated whether 8-Cl-Ado could enhance the ability of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3], a known keratinocyte differentiating agent and a clinical treatment for psoriasis, to inhibit keratinocyte growth. We found that low concentrations of 8-Cl-Ado and 1,25(OH)2D3 appeared to act additively to reduce proliferation of primary mouse epidermal keratinocytes. However, another agent (transforming growth factor-beta) that triggers growth arrest without inducing differentiation also coincidentally inhibits differentiation elicited by other agents; inhibition of differentiation is suboptimal for treating skin disorders, as differentiation is often already reduced. Thus, we determined whether 8-Cl-Ado also decreased keratinocyte differentiation induced by 1,25(OH)2D3, as measured using the early and late differentiation markers, keratin 1 protein levels and transglutaminase activity, respectively. 8-Cl-Ado did not affect 1,25(OH)2D3-stimulated keratin 1 protein expression or transglutaminase activity. CONCLUSIONS: Our results suggest that 8 Cl-Ado might be useful in combination with differentiating agents for the treatment of hyperproliferative disorders of the skin. PMID- 15279681 TI - The "Goldilocks effect" in cystic fibrosis: identification of a lung phenotype in the cftr knockout and heterozygous mouse. AB - BACKGROUND: Cystic Fibrosis is a pleiotropic disease in humans with primary morbidity and mortality associated with a lung disease phenotype. However, knockout in the mouse of cftr, the gene whose mutant alleles are responsible for cystic fibrosis, has previously failed to produce a readily, quantifiable lung phenotype. RESULTS: Using measurements of pulmonary mechanics, a definitive lung phenotype was demonstrated in the cftr-/- mouse. Lungs showed decreased compliance and increased airway resistance in young animals as compared to cftr+/+ littermates. These changes were noted in animals less than 60 days old, prior to any long term inflammatory effects that might occur, and are consistent with structural differences in the cftr-/- lungs. Surprisingly, the cftr+/- animals exhibited a lung phenotype distinct from either the homozygous normal or knockout genotypes. The heterozygous mice showed increased lung compliance and decreased airway resistance when compared to either homozygous phenotype, suggesting a heterozygous advantage that might explain the high frequency of this mutation in certain populations. CONCLUSIONS: In the mouse the gene dosage of cftr results in distinct differences in pulmonary mechanics of the adult. Distinct phenotypes were demonstrated in each genotype, cftr-/-, cftr +/-, and cftr+/+. These results are consistent with a developmental role for CFTR in the lung. PMID- 15279683 TI - Stem cells and prostate cancer. PMID- 15279682 TI - A computer simulation analysis of the accuracy of partial genome sequencing and restriction fragment analysis in estimating genetic relationships: an application to papillomavirus DNA sequences. AB - BACKGROUND: Determination of genetic relatedness among microorganisms provides information necessary for making inferences regarding phylogeny. However, there is little information available on how well the genetic relationships inferred from different genotyping methods agree with true genetic relationships. In this report, two genotyping methods - restriction fragment analysis (RFA) and partial genome DNA sequencing - were each compared to complete DNA sequencing as the definitive standard for classification. RESULTS: Using the Genbank database, 16 different types or subtypes of papillomavirus were selected as study samples, because numerous complete genome sequences were available. RFA was achieved by computer-simulated digestion. The genetic similarity of samples, based on RFA, was determined from the proportion of fragments that matched in size. DNA sequences of four specific genes (E1, E6, E7, and L1), representing partial genome sequencing, were also selected for comparison to complete genome sequencing. Laboratory error was not taken into account. Evaluation of the correlation between genetic similarity matrices (Mantel's r) and comparisons of the structure of the derived dendrograms (partition metric) indicated that partial genome sequencing (for single genes) had higher agreement with complete genome sequencing, achieving a maximum Mantel's r = 0.97 and a minimum partition metric = 10. RFA had lower agreement, with a maximum Mantel's r = 0.60 and a minimum partition metric = 18. CONCLUSIONS: This simulation indicated that for smaller genomes, such as papillomavirus, partial genome sequencing is superior to restriction fragment analysis in representing genetic relatedness among isolates. The generalizability of these results to larger genomes, as well as the impact of laboratory error, remains to be demonstrated. PMID- 15279685 TI - Docetaxel improves survival in metastatic androgen-independent prostate cancer. PMID- 15279686 TI - Screening for prostate cancer. AB - Prostate cancer is a highly prevalent disease in the Western world. In the United States alone, prostate cancer affects approximately 230,000 men and causes the death of 30,000 American men annually. Several theoretical health care measures may be implemented to decrease the morbidity and mortality of any disease. These measures include prevention, screening, improved curative treatment, and the transformation of an acute lethal disease to a chronic, tolerable one. This summary focuses on the screening aspects of prostate cancer. PMID- 15279687 TI - Prostatic intraepithelial neoplasia: an update. AB - High-grade prostatic intraepithelial neoplasia (HGPIN) is commonly encountered on prostate needle biopsies and, based on epidemiologic, molecular, and animal models, has proven to be the most significant risk factor for prostate cancer and likely represents the premalignant phase of prostatic adenocarcinoma. This lesion is characterized by cellular proliferations within pre-existing ducts and glands, with nuclear and nucleolar enlargement similar to prostate cancer. However, unlike cancer, HGPIN retains a basal cell layer identifiable by immunohistochemistry with the basal cell-specific antibody cytokeratin 34bE12. The incidence of HGPIN identified in needle biopsies is as high as 25%, increases with age, and coexists with prostate cancer in approximately 85% of cases. There appears to be no causal relationship between HGPIN and serum prostate-specific antigen (total, percent free, or density) or radiographic characteristics on transrectal ultrasound. In a large series, the identification of HGPIN on initial needle biopsy is associated with about a 35% risk of prostate cancer on subsequent biopsies. Thus, the finding of HGPIN on prostate needle biopsy necessitates a second biopsy in a patient eligible for curative treatment. As a precursor lesion, HGPIN is currently a target for chemopreventive strategies, including antiandrogens and nutritional supplementation. PMID- 15279688 TI - Ethnic variation in localized prostate cancer: a pilot study of preferences, optimism, and quality of life among black and white veterans. AB - Ethnic variations that may influence the preferences and outcomes associated with prostate cancer treatment are not well delineated. Our objective was to evaluate prospectively preferences, optimism, involvement in care, and quality of life (QOL) in black and white veterans newly diagnosed with localized prostate cancer. A total of 95 men who identified themselves as black/African-American or white who had newly diagnosed, localized prostate cancer completed a "time trade-off" task to assess utilities for current health and mild, moderate, and severe functional impairment; importance rankings for attributes associated with prostate cancer (eg, urinary function); and baseline and follow-up measures of optimism, involvement in care, and QOL. Interviews were scheduled before treatment, and at 3 and 12 months after treatment. At baseline, both blacks and whites ranked pain, bowel, and bladder function as their most important concerns. Optimism, involvement in care, and QOL were similar. Utilities for mild impairment were lower for blacks than whites, but were similar for moderate and severe problems. Decline in QOL at 3 and 12 months compared to baseline occurred for both groups. However, even with adjustment for marital status, education level, and treatment, blacks had less increase in nausea and vomiting and more increase in difficulty with sexual interest and weight gain compared with whites. Black and white veterans entered localized prostate cancer treatment with similar priorities, optimism, and involvement in care. Quality-of-life declines were common to both groups during the first year after diagnosis, but ethnic variation occurred with respect to nausea and vomiting, sexual interest, and weight gain. PMID- 15279689 TI - Assessing a prostate cancer brachytherapy technique using early patient-reported symptoms: a potential early indicator for technology assessment? AB - Brachytherapy for early prostate cancer can cause long-term urinary, bowel, and sexual dysfunction. Modifying technique may mitigate complications, but definitive outcome assessment requires long-term follow-up. Although radiation dose plausibly mediates all treatment-related toxicity, short-term symptoms may indicate long-term outcomes. We sought an early indication of whether a modified brachytherapy technique successfully decreased toxicity in the anticipated direction by assessing changes in symptoms and symptom distress 3 months after treatment. In a prospective study of clinically localized prostate cancer using a validated, patient-reported questionnaire, we assessed 85 men, whose primary treatment was brachytherapy alone, prior to treatment and 3 months after the procedure. Twenty-two men received standard ultrasound-guided brachytherapy (SB), and 63 men received magnetic resonance imaging-guided brachytherapy (MB), a technique intended to decrease urinary toxicity by reducing urethral irradiation. Patient age and other sociodemographic variables were similar in the 2 groups. The MB group experienced a greater increase in urinary obstruction/irritation symptoms (P = 0.02) and sexual function distress, but not sexual dysfunction (P = 0.22), whereas the SB group reported a smaller increase in bowel symptoms (P = 0.04) and bowel distress (P = 0.02). We found reduced short-term urinary obstruction/irritation and increased bowel problems after MB consistent with the hypothesized effects of the modified technique, although no obvious mechanism explains the decreased sexual function distress in MB patients. Whether these short-term changes predict long-term outcome differences will require much longer follow-up. However, these results suggest that measuring early symptoms may indicate whether an altered brachytherapy treatment technique has intended favorable consequences, potentially accelerating technology assessment. PMID- 15279690 TI - Analysis of prostate-specific antigen rebound interval in patients with prostate cancer receiving hormonal therapy and external-beam radiation therapy. AB - The goal of this investigation is to characterize the clinical significance of the rebound interval (RI) after neoadjuvant short-course hormonal therapy (HT) and external-beam radiation therapy (RT), during which the prostate-specific antigen (PSA) may rise because of hormone withdrawal prior to full RT efficacy. The charts of 257 consecutive patients with localized prostate cancer who received short-course neoadjuvant HT and RT were reviewed. A piecewise-linear log PSA versus time curve was generated for each patient and averaged over the population to facilitate identification of the RI start and end dates. Existing definitions of biochemical failure--American Society for Therapeutic Radiology and Oncology (ASTRO), Vancouver and Houston--were applied, as were these same definitions modified to exclude failures during the RI. Sensitivity and specificity were analyzed, using no evidence (by digital rectal examination or radiology) of disease failure as the gold standard. The 5-year biochemical survival with different failure definitions were ASTRO versus ASTRO-modified: 81.6% versus 86.7%; Houston versus Houston-modified: 71.4% versus 76.7%; and Vancouver versus Vancouver-modified: 83.5% versus 85.6%. The sensitivity and specificity comparisons were ASTRO versus ASTRO-modified 58.3% versus 33.3%; 91.4% versus 94.3%, Vancouver versus Vancouver-modified: 50% versus 50%; 92.7% versus 95.5%, Houston versus Houston-modified: 100% versus 66.7%; 90.6% versus 92.2%. The RI after HT and RT is likely not merely an artifact of hormone withdrawal but is correlated with ultimate clinical outcome. Excluding RI failures can marginally improve specificity but may possibly have an unacceptable risk of lowering sensitivity. Further work is needed to design and validate definitions of failure, which account for the RI. PMID- 15279691 TI - Plasma levels of heat shock protein 70 in patients with prostate cancer: a potential biomarker for prostate cancer. AB - Heat shock protein 70 (Hsp70) is a stress-inducible protein that is also known for its inhibitory effects on apoptosis. Increased Hsp70 expression is reported in a variety of tumor tissues. Heat shock protein 70 is detectable in plasma and could potentially be used as a biomarker for diagnosis or disease stratification. The relationship between plasma levels of Hsp70 and prostate cancer status has not been well studied. Our study was designed to test this relationship. One hundred twenty-five patients with localized/untreated or hormone-refractory prostate cancer were identified. Forty-five healthy male blood donors between 50 and 73 years of age served as controls. EDTA plasma was subjected to quantitative sandwich immunoassays for both Hsp70 and prostate-specific antigen (PSA). Wilcoxon rank-sum tests were used to examine differences by category. Maximally selected c2 statistics were used to identify cutoff points to best distinguish between categories. Plasma Hsp70 levels in the patients with localized untreated disease (n = 68; median, 0.8 ng/mL; interquartile range, 0.5-2.0) were significantly higher than those in the control group (n = 45; median, 0.5 ng/mL; interquartile range, 0.3-0.8; P = 0.0037). Although the primary cutoff point (1.15 ng/mL) significantly distinguished the localized untreated patients from the control group, plasma Hsp70 levels did not prove more effective than PSA as a predictor for diagnosis or stratification of patients with prostate cancer in the context of group comparisons. Nonetheless, several patients in the localized untreated group showed higher plasma levels of Hsp70 than the primary cutoff point even though their PSA levels were within normal range (< 4 ng/mL). Heat shock protein 70 is a marker of prostate cancer, although its clinical utility is uncertain. It is possible that when used in conjunction with PSA it might prove useful in identifying patients with early-stage prostate cancer who might otherwise be missed by PSA screening alone. PMID- 15279692 TI - Thalidomide and analogues: current proposed mechanisms and therapeutic usage. AB - Microvessel density is a prognostic factor for many cancers, including prostate. For this reason, several studies and therapeutic approaches that target the tumor microvasculature have been attempted. Thalidomide has long been recognized as an antiangiogenic molecule. Recently, this drug has regained favor as an anticancer agent and is in clinical trial for multiple myeloma and prostate cancer, among others. This article will briefly review the proposed mechanisms of action for thalidomide, discuss why these activities are of therapeutic value in diseases currently undergoing clinical trials, and summarize the current status of clinical trials for prostate cancer. The focus will be predominantly on the relationship of thalidomide to angiogenesis, as well as on the future and potential value of thalidomide-inspired structural derivatives. PMID- 15279694 TI - Structural similarities in the cellular receptors used by adenovirus and reovirus. AB - Adenovirus and reovirus are nonenveloped viruses that engage cell-surface receptors using filamentous attachment proteins with head-and-tail morphology. The coxsackievirus and adenovirus receptor (CAR) and reovirus receptor junctional adhesion molecule 1 (JAM1) are immunoglobulin superfamily members that form homodimers stabilized by ionic and hydrophobic contacts between their N-terminal immunoglobulin-like domains. Both proteins are expressed at regions of cell-cell contact and contain sequences in their cytoplasmic tails that anchor the proteins to the actin cytoskeleton. Like CAR and JAM1, the attachment proteins of adenovirus and reovirus, fiber and sigma1, respectively, also share key structural features. Both fiber and sigma1 have defined regions of flexibility within the tail, which is constructed in part using an unusual triple beta-spiral motif. The head domains of both proteins are formed by an 8-stranded beta-barrel with identical beta-strand connectivity. Strikingly, both adenovirus fiber and reovirus 1 engage their receptors by interacting with sequences that also mediate formation of receptor homodimers. Therefore, while adenovirus and reovirus belong to different virus families and have few overall properties in common, the observed similarities between the receptors and attachment proteins of these viruses suggest a conserved mechanism of attachment and an evolutionary relationship. PMID- 15279695 TI - Structural constraints on viral escape from HIV- and SIV-specific cytotoxic T lymphocytes. AB - Cytotoxic T lymphocytes (CTL) play a central role in controlling lentiviral infections in both humans and monkeys. While they contain the spread of human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV), CTL are not capable of fully eradicating virus following infection. Ongoing viral replication can therefore lead to the accumulation of viral mutations within CTL epitopes that can undermine cellular immune control of virus. Here we review the importance of CTL in controlling HIV/SIV infection and how immunologic pressure exerted by effector T cells selects for viral variants that escape CTL recognition. We review two examples of viral escape from CTL at highly conserved epitopes that illustrate the extraordinary capacity of lentiviruses to adapt to their immunologic environment despite structural constraints on the ability of the virus to accommodate mutations. PMID- 15279696 TI - T cells in coxsackievirus-induced myocarditis. AB - Myocarditis is a complex disease in which distinct immunopathogenic mechanisms cause tissue injury. In some but not all cases, autoimmunity is a major pathogenic factor. Cross-reactivity between viral and myosin epitopes underlies both cellular and humoral autoimmunity in myocarditis. Thus, the genetics of the host as well as the virus determine disease pathogenicity. Innate immunity, as represented by gammadelta+ T cells, is important in determining disease susceptibility. The innate effectors rapidly localize in the infected myocardium and through release of IFNgamma (Vgamma4+ cells; BALB/c) or IL-4 (Vgamma1+ cells; C57Bl/6), modulate the developing adaptive immune response to either a Th1 or Th2 response, respectively. The Vgamma4+ cells in BALB/c mice recognize CD1d, a major histocompatibility complex class I-like antigen. The ligand for Vgamma1+ cells is unknown. Only infected myocytes up-regulate CD1d. Signaling through both infection (double stranded RNA) and TNFalpha is required for CD1d up-regulation. PMID- 15279697 TI - Pathogenesis of respiratory syncytial virus infection. AB - Respiratory syncytial virus (RSV) is recognized as the most important cause of serious lower respiratory tract illness in infants and young children worldwide causing repeat infections throughout life with serious complications occurring in the elderly and immune compromised patient. The level of disease pathogenesis associated with RSV infection is balanced between virus elimination and the nature of the immune response to infection. The innate and adaptive immune responses to RSV infection are not fully elucidated; however, significant progress has been made in understanding the virus-host relationship and mechanisms associated with disease pathogenesis. This review summarizes important aspects of these findings, and provides current perspective on processes that may contribute to RSV disease pathogenesis. PMID- 15279698 TI - Modifying adenoviral vectors for use as gene-based cancer vaccines. AB - The past decade has produced significant advances in our understanding of antigen presenting cells, tumor antigens, and other components of the immune response to cancer. Gene-based vaccination is emerging as one of the more promising approaches for loading dendritic cells (DC) with tumor-associated antigens. In this respect, it is proposed that adenoviral (AdV) vectors can deliver high antigen concentrations, promote effective processing and MHC expression, and stimulate potent cell-mediated immunity. While AdV vectors have performed well in pre-clinical vaccine models, their application to patient care has limitations. The in vivo administration of AdV vectors is associated with both innate and adaptive host responses that result in tissue inflammation and injury, viral neutralization, and premature clearance of AdV-transduced cells. A variety of strategies have been developed to address these limitations. The ideal vaccine would avoid vector-related immune responses, have relative specificity for transducing DC, and induce high levels of transgene expression. This review describes the range of host responses to AdV vaccines, identifies strategies to reduce viral recognition and enhance transgene antigen expression, and suggests future approaches to vector development and administration. There is every reason to believe that safer and more effective forms of AdV-based vaccines can be developed and applied to patient therapy. PMID- 15279699 TI - T cell responses to influenza virus infection: effector and memory cells. AB - New approaches to visualizing antigen-specific primary responses to influenza and the development of memory subsets in distinct sites suggest that both CD4 and CD8 T cells play complex roles in primary viral clearance and have the potential to contribute to protection from secondary infection. PMID- 15279700 TI - Host evasion by emerging paramyxoviruses: Hendra virus and Nipah virus v proteins inhibit interferon signaling. AB - Interferon (IFN) can activate Signal Transducer and Activator of Transcription (STAT) proteins to establish a cellular antiviral response and inhibit virus replication. Many viruses have evolved strategies to inhibit this antiviral mechanism, but paramyxoviruses are unique in their abilities to directly target the IFN-responsive STAT proteins. Hendra virus and Nipah virus (Henipaviruses) are recently emerged paramyxoviruses that are the causative agents of fatal disease outbreaks in Australia and peninsular Malaysia. Similar to other paramyxoviruses, Henipaviruses inhibit IFN signal transduction through a virus encoded protein called V. Recent studies have shown that Henipavirus V proteins target STAT proteins by inducing the formation of cytoplasmically localized high molecular weight STAT-containing complexes. This sequestration of STAT1 and STAT2 prevents STAT activation and blocks antiviral IFN signaling. As the V proteins are important factors for host evasion, they represent logical targets for therapeutics directed against Henipavirus epidemics. PMID- 15279701 TI - Innate immune responses in respiratory syncytial virus infections. AB - Respiratory syncytial virus (RSV) is the most important viral respiratory pathogen of early life. Studies of the immune response in general (and the innate response in particular) to this agent are of interest for a number of reasons. First, severe forms of illness may be a result of enhanced immunologic responsiveness to viral constituents at the time of infection. Secondly, the immune response to RSV may consist principally of innate immune responses at the time of maximum severity of illness. Third, RSV infection in infancy may be linked via immune mechanisms to the development of childhood wheezing. Finally there are no meaningfully effective forms of therapy for RSV infection, and elucidation of the immune response may suggest new therapeutic approaches. This review will summarize our current knowledge of innate immune responses to RSV infection. Specifically we will review early interactions of the virus with surfactant proteins and Toll-like receptors, chemokine release from infected cells, cytokine release from activated inflammatory cells, activation of neuroimmune pathways, generation of dendritic cells, the release of soluble mediators of airway obstruction, and genetic polymorphisms associated with RSV related illness. PMID- 15279702 TI - Regulation of cellular gene expression in endothelial cells by sin nombre and prospect hill viruses. AB - Mechanisms of hantavirus-induced vascular leakage remain unknown. This study was initiated to determine whether hantavirus-induced changes in endothelial cell gene expression may provide insight into disease mechanisms. Additionally, by using pathogenic Sin Nombre virus (SNV) and non-pathogenic Prospect Hill virus (PHV), we wanted to identify cellular responses that are likely to differentiate pathogenic from nonpathogenic hantaviruses. Using the Affymetrix DNA Array, we found that PHV and SNV did not significantly differ in the number of activated genes (18 versus 14 genes) in infected endothelial cells at 4 h PI. However, a smaller group of genes (36) were up-regulated by PHV compared to SNV (175) at 12 h PI. Only two genes were down-regulated in SNV-infected cells. Expression of the functionally diverse group of genes was altered at an early stage of infection (4 and 12 h PI). The genes affected include putative anti-viral factors, transcription factors, growth factors, chemokines, receptors, structural proteins, metabolism, and kinases. Although many genes were activated in cells infected with SNV and PHV, overall cellular transcriptional responses were more altered by pathogenic SNV compared to non-pathogenic PHV. PMID- 15279703 TI - Modulation of dengue virus infection of dendritic cells by Aedes aegypti saliva. AB - Dengue virus (DV) is a flavivirus carried by the Aedes aegypti mosquito that causes a spectrum of illnesses in the tropics, including dengue fever, dengue hemorrhagic fever, and dengue shock syndrome. Dendritic cells (DCs) are professional antigen presenting cells recently shown to be permissive for DV, and implicated as the primary targets of initial DV infection. DV is transmitted to human host by infected mosquitoes during a blood meal, but it is currently unknown whether transmission is modified by vector saliva that is also deposited in the host's skin during feeding. Previous studies evaluated only the outcome of DV infection of DCs, and did not address the influence of mosquito saliva. To more fully characterize natural transmission of DV, we evaluated the effects of Ae. aegypti saliva on DV infection of human myeloid DCs. We found that saliva inhibited DV infection in DCs. Moreover, pre-sensitization of DCs with saliva, prior to DV infection, enhanced inhibition. In addition, enhanced production of IL-12p70 and TNF-alpha were detected in DV-infected DC cultures exposed to mosquito saliva. The proportion of dead cells was also significantly reduced in these cultures. These data contribute to the overall understanding of the natural pathogenesis of DV infection and suggest that there is a protective role for mosquito saliva that limits viral uptake by DCs. PMID- 15279704 TI - Vertical transmission of a murine retrovirus, ts1. AB - Mechanism of maternal retroviral transmission remains an unsolved problem. The current investigation is a part of our ongoing research on vertical transmission of MoMuLV-TB ts1 in BALB/c mice. A total of 270 adult mice and 165 fetuses were used. Forty-four experimental mice were injected with 0.1 mL of 4.0 x 10(6) ffu/mL of ts1 virus at 72 h after birth; 24 controls were injected with DMEM. Almost half of the females went through two rounds of pregnancies. In the first round, 135 experimental and 57 control pups were produced. Forty-three experimental and 20 control pups were followed until they developed clinical symptoms. The second round of pregnancy produced a total of 46 mid-gestational and 119 full-term fetuses. PCR, and light and electron microscopy were performed to evaluate viral transmission. Overall, 99% vertical transmission occurred in pups of infected mothers. Twelve percent of mid-gestational and 39% full-term fetuses were PCR positive. We have established that, if mothers are infected with ts1 virus at 72 h after birth, then nearly 100% vertical transmission occurs, via in utero, intrapartum, or breast milk. Thirty-nine percent transmission occurred in utero alone. This is an excellent model to study the transplacental and post gestational transmission of retroviruses, such as ts1. PMID- 15279705 TI - Immune correlates of virological response in HIV-positive patients after highly active antiretroviral therapy (HAART). AB - Correlates of immune reconstitution after highly active antiretroviral therapy (HAART) are not completely understood, in particular as far as viro-immunological discordant responses are concerned. HIV-positive patients on stable HAART for > or = 1 year were recruited. Viro-immunological responses were categorized according to positive or negative area under the curve (AUC) variations for HIV plasma viral load (pVL) and CD4+ T-cell counts measured at least every 4 months. The following parameters were evaluated: lymphocyte spontaneous apoptosis (LSA), intracellular Bcl-2 expression in both CD4-CD45RA+ and CD4-CD45R0+, IL-7 and IL 15 plasma concentrations, and lymphocyte TRECs levels. Sixty-one patients were enrolled. A significant inverse correlation was found between CD4+ T-cell count and pVL AUC (r = 0.45; p = 0.0003). Patients with pVL response had higher levels of Bcl-2 in CD4-CD45R0+ (mean 65,409 MESF vs. 54,018 MESF; p = 0.089) and higher IL-15 (mean 1.34 pg/mL vs. 1.05 pg/mL; p = 0.069, respectively). Higher LSA and lower TRECs levels were found in viro-immunological non-responder patients with respect to those who had viro-immunological response (mean 24.84% vs. 14.89%; p = 0.01, and mean 17,796 copies/10(6) cells vs. 29,251 copies/10(6) cells; p = 0.68, respectively). Virological suppression may allow Bcl-2 and IL-15 hyperexpression during incomplete immune-reconstitution phase, while more complete immune reconstitution appeared to be marked by both high TRECs and low LSA levels, possibly indicating both central and peripheral CD4+ T-cell repopulations at this stage. PMID- 15279706 TI - mRNA expression of proinflammatory cytokines in mouse CNS correlates with replication rate of semliki forest virus but not with the strain of viral proteins. AB - Tissue expression in viral infection of immunological effector molecules may depend on virus structure or replication or both. We analyzed cytokine mRNA expression in the central nervous system (CNS) of Balb/c mice during viral infection with Semliki Forest virus (SFV) clones, which varied either in structure or virulence or both. Highly neurovirulent SFV4 effectively induced IFN gamma, TNF-alpha, IL-6 and TGF-beta, but its avirulent derivative V4-opal with nsP3 arginine-476 to opal mutation, elicited only weak induction of these cytokines. Structurally different, avirulent rA774, obtained by cloning from avirulent SFV A7(74) strain, did not induce synthesis of proinflammatory Th1 or Th2 cytokines in murine CNS, but increased synthesis of TGF-beta transcripts. In contrast, structurally identical but moderately virulent rA774-arg virus with sense codon at opal position in nsP3, markedly stimulated synthesis of IFN-gamma, TNF-alpha, and IL-10 transcripts, without, however, reaching the levels elicited by lethal SFV4. The rA774-arg clone was more potent in attracting peripheral immune cells into the CNS than the completely avirulent strains. In conclusion, induction of proinflammatory cytokine mRNA in the CNS by SFV infection seemed to correlate with the rate of viral replication and was not significantly influenced by the virus envelope or nonstructural protein primary structure. The results also have relevance for development of CNS gene therapy vectors as SFV4 and A774 display differences in CNS infection characteristics. PMID- 15279707 TI - T cell immunity to measles viral proteins in infants and adults after measles immunization. AB - Vaccination of infants against measles remains of global importance, and proposed new vaccine strategies include the use of measles proteins or synthetic peptides as immunogens. We studied cell-mediated immunity to whole measles antigen and measles proteins in immune adults and infants after measles vaccine. Further, we measured CD8+ T cell responses to peptide pools corresponding to the nucelocapsid (N) measles protein in adults given measles vaccine. Cell-mediated immune responses to three of four measles proteins were equivalent to those against whole measles antigen in immune adults. Responses to the fusion (F) protein were lower in infants compared to whole measles antigen (p < or = 0.03). Infant responses to both whole measles antigen and the F protein were lower compared with these responses in adults (p < or = 0.001). CD8+ T cell responses to N peptide pools varied, and differed between immune HLA-A2-positive individuals compared with naive and HLA-A2-negative subjects after measles vaccination. The measles-specific T cell adaptive response of infants is limited compared to adults, including responses to the F protein. PMID- 15279708 TI - Herpes simplex virus 1 induced LOX-1 expression in an endothelial cell line, ECV 304. AB - Infections, such as by Chlamydophilia pneumoniae, cytomegalovirus, herpes simplex virus, and Helicobacter pylori, have been shown to be involved in atherogenesis. Herpes simplex virus I (HSV-1) could infect vascular endothelial cells, and it has been shown that, when endothelial cells were activated with oxidized LDL (oxLDL), a number of cellular events are occurred, leading to endothelial cell dysfunction. Since LOX-1 is a major receptor for oxLDL on endothelial cells and its expression was increased in atherosclerosis, we investigated whether HSV1 infection can lead to the increase expression of LOX-1 in endothelial cells. LOX 1 mRNA expression determined by RT-PCR and LOX-1 promoter activity measured by luciferase assay were increased in endothelial cells following HSV-1 infection. This suggests that one of the mechanisms by which HSV-1 is involved in atherogenesis maybe the enhanced uptake of oxLDL via the increased expression of LOX-1 in endothelial cells. PMID- 15279709 TI - Detection of Puumala hantavirus antibody with ELISA using a recombinant truncated nucleocapsid protein expressed in Escherichia coli. AB - A truncated recombinant nucleocapsid protein (rNp118) consisting of the first 118 amino-terminal amino acids (AA) of the Puumala hantavirus (PUUV) nucleocapsid protein expressed in Escherichia coli, was evaluated for its antigenicity and reliability as serodiagnostic antigen in an enzyme-linked immunosorbent assay (ELISA) for detection of PUUV antibodies in human sera. The PUUV nucleocapsid protein has been shown to contain several B-cell epitopes, mapped within the first 118 amino-terminal AA. This finding makes the rNp118 an interesting recombinant protein to use as serodiagnostic antigen. The sensitivity of this new PUUV-rNp118 ELISA, was compared with those of a commercially available PUUV ELISA assay and an home-made ELISA based on a recombinant whole nucleocapsid protein of PUUV. Eighty-six human serum samples clinically suspected for PUUV-induced nephropathia epidemica, and previously screened with the reference assays, were tested. The sensitivity of the new assay was compared with that of the reference assays and an excellent correlation between the assays was found. Sera found to be negative by other methods were also negative in our assay. The ELISA based on rNp118 represents an alternative and valid test for detection of antibodies to PUUV in human sera. PMID- 15279710 TI - Brain cancer mortality in the United States, 1986 to 1995: a geographic analysis. AB - The Atlas of Cancer Mortality in the United States, 1950-94 (Devesa et al.) published in 1999 by the National Institutes of Health suggests that there are elevated rates of brain and other nervous system cancer in the northwestern, north central, and southeastern parts of the country. Being descriptive in nature, the atlas does not evaluate whether observed patterns are simply due to random variation or if they are reflective of true geographical differences in disease risk or treatment practices. To formally test for geographical clustering of disease, we analyzed U.S. brain cancer mortality data from 1986 to 1995 with Tango's Excess Events test, the Cuzick-Edwards k-Nearest-Neighbors test, and the spatial scan statistic. All tests revealed statistically significant geographical clustering for both adult men and women. The spatial scan statistic indicated that the most likely cluster of high mortality was in parts of Arkansas, Mississippi, and Oklahoma (relative risk [RR] = 1.22, P < 0.0001) for women and in parts of Tennessee and Kentucky (RR = 1.15, P < 0.0001) for men. Several secondary clusters were detected, but there were no statistically significant clusters of a very localized nature and a high RR. For childhood brain cancer, there were no statistically significant geographical clusters. It is reassuring that no local brain cancer mortality "hot spots" with very high RRs were found. While the causes of the large geographical clusters with modest RRs are unclear, the geographical pattern of brain cancer mortality provides valuable information that can help in formulating etiological hypotheses and in targeting high-risk populations for further epidemiological and health services research. PMID- 15279711 TI - Induction of membrane-type-1 matrix metalloproteinase by epidermal growth factor mediated signaling in gliomas. AB - Increased expression of membrane-type matrix metalloproteinases (MT-MMPs) has previously been reported to correlate with increasing grade of malignancy in gliomas, a relationship shared with alterations in epidermal growth factor receptor (EGFR) signaling. To investigate the possibility of a causative role for EGFR signaling in increasing MT-MMP expression and subsequent peritumoral proteolysis, we characterized glioma cell lines for expression of MT1-MMP, MT2 MMP, MT3-MMP, and MT5-MMP by Western blotting and by quantitative real-time polymerase chain reaction analysis, and for MMP-2 activity following epidermal growth factor (EGF) stimulation. EGF stimulation of glioma cell lines resulted in a 2- to 4-fold increase in MT1-MMP mRNA levels. Although there were slight differences in MT2-, MT3-, and MT5-MMP mRNA expression following EGF stimulation, none of these demonstrated an increase similar to that of MT1-MMP expression. Treatment of high-grade glioma cell lines U251MG and IPSB-18 with EGF for 24 h resulted in a several-fold increase in MT1-MMP protein (2.5- and 5.1-fold, respectively) and in cyclin D1 (2.9-fold), as compared to untreated controls. No significant increase was detected in other MT-MMPs at the protein level. Although there was no detectable increase in proMMP-2 protein, there was an increase in MMP-2 activity. Furthermore, the MT1-MMP induction by EGF was prevented by pretreatment with the EGFR-specific tyrphostin inhibitor AG1478. Similarly, treatment with the phosphatidylinositol 3-kinase inhibitor LY294002 prevented the induction of MT1-MMP protein by EGF stimulation. These compounds additionally inhibited EGF-stimulated invasion in Matrigel Transwell assays. Our results indicate that one mechanism of EGFR-mediated invasiveness in gliomas may involve the induction of MT1-MMP. PMID- 15279712 TI - Intercellular heterogeneity of expression of the MGMT DNA repair gene in pediatric medulloblastoma. AB - DNA methylation and epigenetic inactivation of the O6-methylguanine methyltransferase (MGMT) gene induces MGMT deficiency, reducing the tumor cell's DNA repair capacity and increasing its susceptibility to alkylating chemotherapeutic agents. Consequently, adult patients whose tumors are deficient in MGMT have better outcomes with alkylator chemotherapy, and MGMT methylation has been proposed as a screening marker of deficient tumors. In order to test the feasibility of this approach for medulloblastoma, a common brain tumor in children, we determined the methylation status, mRNA expression pattern, and protein expression of MGMT in a panel of clinical specimens. Methylation-specific polymerase chain reaction analysis revealed methylation of MGMT in 28 of 37 tumor samples. Quantitative real-time reverse transcriptase-polymerase chain reaction showed a range of expression of MGMT mRNA varying more than 20-fold. However, there was no correlation found between MGMT methylation and mRNA expression. Immunohistochemistry demonstrated that all tumors were immunoreactive for MGMT in the nucleus of the medulloblastoma cells in a heterogeneous pattern. The intercell variability of MGMT complement explained the discordance between methylation and expression. Therefore, MGMT methylation as determined by methylation-specific polymerase chain reaction cannot be used as a marker for MGMT deficiency in medulloblastoma. Further, these findings support the use of pharmacological MGMT depletion as a rational approach for intensification of alkylator chemotherapy in the treatment of medulloblastoma. PMID- 15279713 TI - Poliovirus receptor CD155-targeted oncolysis of glioma. AB - Cell adhesion molecules of the immunoglobulin superfamily are aberrantly expressed in malignant glioma. Amongst these, the human poliovirus receptor CD155 provides a molecular target for therapeutic intervention with oncolytic poliovirus recombinants. Poliovirus has been genetically modified through insertion of regulatory sequences derived from human rhinovirus type 2 to selectively replicate within and destroy cancerous cells. Efficacious oncolysis mediated by poliovirus derivatives depends on the presence of CD155 in targeted tumors. To prepare oncolytic polioviruses for clinical application, we have developed a series of assays in high-grade malignant glioma (HGL) to characterize CD155 expression levels and susceptibility to oncolytic poliovirus recombinants. Analysis of 6 HGL cases indicates that CD155 is expressed in these tumors and in primary cell lines derived from these tumors. Upregulation of the molecular target CD155 rendered explant cultures of all studied tumors highly susceptible to a prototype oncolytic poliovirus recombinant. Our observations support the clinical application of such agents against HGL. PMID- 15279714 TI - Antitumor effects of specific telomerase inhibitor GRN163 in human glioblastoma xenografts. AB - Telomerase is a ribonucleoprotein complex that elongates telomeric DNA and appears to play an important role in cellular immortalization of cancers. Because telomerase is expressed in the vast majority of malignant gliomas but not in normal brain tissues, it is a logical target for gliomaspecific therapy. The telomerase inhibitor GRN163, a 13-mer oligonucleotide N3'-->P5' thio phosphoramidate (Geron Corporation, Menlo Park, Calif.), is complementary to the template region of the human telomerase RNA subunit hTR. When athymic mice bearing U-251 MG human brain tumor xenografts in their flanks were treated intratumorally with GRN163, a significant growth delay in tumor size was observed (P < 0.01 in all groups) as compared to the tumor size in mice receiving a mismatched oligonucleotide or the carrier alone. We also investigated biodistribution of the drug in vivo in an intracerebral rat brain-tumor model. Fluorescein-labeled GRN163 was loaded into an osmotic minipump and infused directly into U-251 MG brain tumors over 7 days. Examination of the brains revealed that GRN163 was present in tumor cells at all time points studied. When GRN163 was infused into intracerebral U-251 MG tumors shortly after their implantation, it prevented their establishment and growth. Lastly, when rats with larger intracerebral tumors were treated with the inhibitor, GRN163 increased animal survival times. Our results demonstrate that the antitelomerase agent GRN163 inhibits growth of glioblastoma in vivo, exhibits favorable intracerebral tumor uptake properties, and prevents the growth of intracerebral tumors. These findings support further development of this compound as a potential anticancer agent. PMID- 15279715 TI - Prognostic factors for survival of patients with glioblastoma: recursive partitioning analysis. AB - Survival for patients with glioblastoma multiforme is short, and current treatments provide limited benefit. Therefore, there is interest in conducting phase 2 trials of experimental treatments in newly diagnosed patients. However, this requires historical data with which to compare the experimental therapies. Knowledge of prognostic markers would also allow stratification into risk groups for phase 3 randomized trials. In this retrospective study of 832 glioblastoma multiforme patients enrolled into prospective clinical trials at the time of initial diagnosis, we evaluated several potential prognostic markers for survival to establish risk groups. Analyses were done using both Cox proportional hazards modeling and recursive partitioning analyses. Initially, patients from 8 clinical trials, 6 of which included adjuvant chemotherapy, were included. Subsequent analyses excluded trials with interstitial brachytherapy, and finally included only nonbrachytherapy trials with planned adjuvant chemotherapy. The initial analysis defined 4 risk groups. The 2 lower risk groups included patients under the age of 40, the lowest risk group being young patients with tumor in the frontal lobe only. An intermediate-risk group included patients with Karnofsky performance status (KPS) >70, subtotal or total resection, and age between 40 and 65. The highest risk group included all patients over 65 and patients between 40 and 65 with either KPS<80 or biopsy only. Subgroup analyses indicated that inclusion of adjuvant chemotherapy provides an increase in survival, although that improvement tends to be minimal for patients over age 65, for patients over age 40 with KPS less than 80, and for those treated with brachytherapy. PMID- 15279716 TI - Results of a phase 1 study utilizing monocyte-derived dendritic cells pulsed with tumor RNA in children and young adults with brain cancer. AB - We conducted a phase 1 study of 9 pediatric patients with recurrent brain tumors using monocyte-derived dendritic cells pulsed with tumor RNA to produce antitumor vaccine (DCRNA) preparations. The objectives of this study included (1) establishing safety and feasibility and (2) measuring changes in general, antigen specific, and tumor-specific immune responses after DCRNA. Dendritic cells were derived from freshly isolated monocytes after 7 days of culture with IL-4 and granulocyte-macrophage colony-stimulating factor, pulsed with autologous tumor RNA, and then cryopreserved. Patients received at least 3 vaccines, each consisting of an intravenous and an intradermal administration at biweekly intervals. The study showed that this method for producing and administering DCRNA from a single leukapheresis product was both feasible and safe in this pediatric brain tumor population. Immune function at the time of enrollment into the study was impaired in all patients tested. While humoral responses to recall antigens (diphtheria and tetanus) were intact in all patients, cellular responses to mitogen and recall antigens were below normal. Following DCRNA vaccine, 2 of 7 patients showed stable clinical disease and 1 of 7 showed a partial response. Two of 7 patients who were tested showed a tumor-specific immune response to DCRNA. This study showed that DCRNA vaccines are both safe and feasible in children with tumors of the central nervous system with a single leukapheresis. PMID- 15279717 TI - Phase 1 study of 28-day, low-dose temozolomide and BCNU in the treatment of malignant gliomas after radiation therapy. AB - We conducted a study to determine the dose-limiting toxicity of an extended dosing schedule of temozolomide (TMZ) when used with a fixed dose of BCNU, or 1,3 bis(2-chloroethyl)-1-nitrosourea (carmustine), taking advantage of TMZ's ability to deplete O6-alkylguanine-DNA-alkyltransferase and the synergistic activity of these two agents. Patients with malignant gliomas who had undergone radiation therapy were eligible. Patients were treated with TMZ for 28 days, followed by a 28-day rest (1 cycle). The TMZ was started at 50 mg/m2 and increased in 10-mg/m2 increments; a fixed dose of BCNU (150 mg/m2) was given within 72 h of starting TMZ. A standard phase 1 dose-escalation scheme was used with 3 patients per cohort. Fourteen glioblastoma patients and 10 anaplastic astrocytoma patients were treated. The dose-limiting toxicity was myelosuppression at 90 mg/m2 of TMZ. The total number of cycles given was 73 (median number was 2). Six patients (25%) required a dose reduction in BCNU, and six were removed from study for hematologic toxicity after cycle 1; three patients overlapped. The median time to progression and overall survival were, respectively, 82 and 132 weeks for anaplastic astrocytomas and 14 and 69 weeks for glioblastomas. We conclude that the combination of BCNU and the extended dosing schedule of TMZ is feasible and that the maximal tolerated dose of a 28-day course of TMZ is 80 mg/m2 when combined with a fixed dose of BCNU at 150 mg/m2. This is the recommended dose for phase 2, but myelosuppression after cycle 1 suggests that long-term treatment may be difficult. PMID- 15279718 TI - 13-cis-retinoic acid in the treatment of recurrent glioblastoma multiforme. AB - Basic science and clinical investigations have demonstrated that 13-cis-retinoic acid (cRA) has activity against malignant gliomas. To assess its effectiveness in the setting of recurrent glioblastoma multiforme (GBM), we performed a retrospective analysis of the medical records and neuroimaging results of patients with recurrent GBM who were treated with cRA. The toxicity profile of cRA, response, and effect on progression-free survival from initiation of treatment were end points of our analysis. Eighty-two of 85 patients with a median age of 51 years received at least 1 full cycle of cRA. At the initiation of cRA treatment, the median Karnofsky performance score was 80. All patients had failed conventional radiotherapy. Seven patients were chemonaive, whereas 75 patients had received some form of chemotherapy. Radiographic partial responses, minor responses, and stable disease were seen in 4%, 8%, and 34% of patients, respectively. Two patients were not assessable. Progression-free survival and overall survival after initiation of cRA were 10.0 and 24.6 weeks, respectively. Six-month progression-free survival was 19% for the entire group. Grade 3 or 4 toxicity developed in 14 patients (16%), one of whom developed pancreatitis and died. The results of this study demonstrate only modest efficacy for cRA therapy in this cohort of heavily pretreated patients with recurrent GBM. This data supports the use of cRA in such patients, but its further evaluation in larger, prospective, controlled studies with or without other noncytotoxic and cytotoxic agents may be warranted. PMID- 15279719 TI - Transmission of glioblastoma multiforme following bilateral lung transplantation from an affected donor: case study and review of the literature. AB - Donor-acquired solid organ malignancy is a rare complication of organ transplantation. We report a case of a patient who received bilateral lung transplants for pulmonary fibrosis from a donor with known glioblastoma multiforme (GBM). The lungs, heart, liver, and kidneys were harvested after a lethal intracranial bleed and accepted for transplantation by four centers. An enlarged hilar lymph node sampled at the time of transplant was found to contain GBM. Four months later, the patient developed diffuse interstitial pulmonary infiltrates with mediastinal lymphadenopathy. Lung biopsy confirmed metastatic GBM. The patient died 2 weeks after the diagnosis was established. The patient receiving the donor liver also developed GBM. We present a case study, review of the literature, and suggested interventions to minimize the risk of transmission. PMID- 15279723 TI - Latanoprost eye drops increase concentration of glycosaminoglycans in posterior rabbit sclera. AB - PURPOSE: Glycosaminoglycans are important components of ocular tissues such as the sclera. The pressure reducing effect of a new antiglaucoma drug, latanoprost, is based on an increase in the uveo-scleral outflow by way of modulation of the intracellular matrix of the ciliary body. The purpose of the study was to test the effect of latanoprost on the content of glycosaminoglycans in rabbit cornea and sclera. METHODS: Twelve rabbits were studied. Six rabbits were treated for 12 weeks with latanoprost eye drops and 6 with hydroxypropylmethylcellulose, dextran 70 eye drops for control. Samples were taken from cornea and anterior, lateral, and posterior sclera. Glycosaminoglycans were determined quantitatively by spectrophotometry (uronic acids). RESULTS: A significant increase in the concentration of uronic acids was found in all three localisations of sclera from latanoprost-treated animals. The increase was 26%, 24%, and 20% in anterior, lateral, and posterior sclera, respectively. CONCLUSIONS: Long-term treatment with latanoprost induces biochemical changes in sclera. The results indicate that topically applied latanoprost reaches the posterior parts of the rabbit eye. PMID- 15279724 TI - Structural requirements of flavonoids for increment of ocular blood flow in the rabbit and retinal function recovery in rat eyes. AB - PURPOSE: We have recently reported that the effect of a flavonoid on ocular blood flow depends upon the number of hydroxy (OH) groups in its backbone structure. To elucidate the structural features on the number and type of functional groups present in the flavonoid molecule plus the number of OH groups, flavonoids with four to five OH groups, with or without methoxy groups, were studied on their effects to affect the ocular blood flow and the retinal function recovery. METHODS: A colored microsphere technique was used to determine the ocular blood flow in albino rabbit eyes and electroretinography was used to measure the retinal function recovery. RESULTS: Flavonols with four free OH groups produced no effects on the ocular blood flow (fisetin, kaempferol), whereas flavanone and flavones with four free OH groups and without the C2-C3 double bond produced the rapid increment on ocular blood flow (dihydrofisetin and luteolin, respectively). Similarly, flavonols with five free OH groups produced no effects on the ocular blood flow (morin, quercetin). Yet, flavanone with five free OH groups and without the C2-C3 double bond produced the rapid increment on ocular blood flow (dihydroquercetin). Flavanols with five free OH groups and without the C2-C3 double bond and the carbonyl group produced no effects on the ocular blood flow (catechin). Flavonols with four free OH groups and a methoxy group on the 7 position produced no effects on the ocular blood flow (Rhamnetin). Flavonols with four free OH groups and a methoxy group at the 5 (5-methylquercetin) or 3' position (isorhamnetin) produced positive effects on the ocular blood flow also. Flavonol with five methoxy groups but no OH group produced positive effects on the ocular blood flow (pentamethylquercetin). Flavonols with an excessive number of OH groups, having both a catechol-like structure in the C ring and a catechol at the B ring, produced no effect on the ocular blood flow (rhamnetin, quercetin). Parallel results were obtained on retinal function recovery after ischemic insult. CONCLUSION: The presence of OH groups at certain positions and the double bond at C2-C3 in the flavonoid molecules, which produces lipophilic action, can affect the increment on ocular blood flow and retinal function recovery. O-methylation can increase ocular blood flow and retinal function recovery as well. PMID- 15279725 TI - Inhibition of endothelin-1 and KCL-induced increase of [CA2+]i by antiglaucoma drugs in cultured A7r5 vascular smooth-muscle cells. AB - Over contraction of vascular smooth muscle may result in ischemia to ocular neuronal cells and deteriorate the glaucoma. The purpose of this study was to investigate the inhibitory effects of various commercial antiglaucoma drugs including brimonidine, dipivefrin, betaxolol, timolol, levobunolol, carteolol, brinzolamide, dorzolamide, unoprostone, latanoprost, pilocarpine, and preservative benzalkonium chloride on endothelin-1(ET-1) and KCl-induced increase of intracellular free Ca2+ ([Ca2+]i) in cultured rat A7r5 vascular smooth muscle cells. These drugs were diluted from original concentrations to 1/100, 1/1000, and 1/10000. [Ca2+]i mobility was analyzed by spectrofluorometry after loading with fura-2-AM. Betaxolol, timolol, levobunolol, and carteolol were found to inhibit KCl-induced release of [Ca2+]i in a dose-dependent manner. High concentrations of betaxolol, timolol, levobunolol, carteolol, and unoprostone also inhibited ET-1-induced increase of [Ca2+]i in A7r5 cells. However, ET-1- and KCl-induced increase of [Ca2+]i was not diminished by other drugs including brimonidine, dipivefrin, brinzolamide, dorzolamide, latanoprost, pilocarpine, and benzalkonium chloride. These results indicate that high concentrations of unoprostone and beta-adrenergic blocking agents including betaxolol, timolol, levobunolol, and carteolol may inhibit ET-1-induced increase of [Ca2+]i. The mechanism may be mediated by inhibition of extracellular calcium influx via blocking of L-type voltage-dependent Ca2+ channel in A7r5 cells. PMID- 15279726 TI - Aqueous penetration of gatifloxacin and levofloxacin into the rabbit aqueous humor following topical dosing. AB - The aqueous penetration of the commercial preparations of the fluoroquinolone antibiotics ofloxacin, ciprofloxacin, levofloxacin, and gatifloxacin were compared following topical dosing in a rabbit model. Levofloxacin achieved the highest aqueous concentrations, with a mean aqueous level of 4.8014 microcg/mL (p = 0.002, p = 0.00002, p = 0.015.) Ofloxacin (2.5136 microcg/mL) and gatifloxacin (2.4817 microcg/mL) achieved statistically equal aqueous concentrations (p = 0.479). Ciprofloxacin reached the lowest levels in the aqueous humor (0.9616 microcg/mL, p = 0.00002, 0.00004, 0.008). Gatifloxacin alone achieved concentrations in excess of the MIC90s of gram-positive pathogens of concern. PMID- 15279728 TI - Effects of carteolol hydrochloride on the in vitro production of LPS-induced proinflammatory cytokines by murine macrophage. AB - We investigated whether carteolol hydrochloride, which has intrinsic sympathomimetic activity (ISA), inhibits the production of proinflammatory cytokines using mouse macrophages (MPs) and peripheral-blood mononuclear cells (PBMCs). MPs and PBMCs were collected from BALB/C strain mice, treated simultaneously with lipopolysaccharide (LPS) and test agents (carteolol hydrochloride, timolol maleate, betaxolol hydrochloride, levobunolol hydrochloride, or nipradilol) in medium, and incubated in a CO2 incubator. TNF alpha and IL-6 in medium were measured by ELISA. Carteolol hydrochloride significantly inhibited the production of TNF-alpha and IL-6 by MPs at 10(-5) M and higher (p < 0.01) or PBMCs at 10(-6) M and higher (p < 0.01) compared to the controls, while the other test agents had no inhibitory effect. Carteolol hydrochloride inhibited the production of proinflammatory cytokines by inflammatory cells, raising the possibility that this intraocular hypotensive drug may be expected to have anti-inflammatory effects in patients with increased intraocular tension and postoperative inflammation. PMID- 15279727 TI - Preventive versus treatment effect of AG3340, a potent matrix metalloproteinase inhibitor in a rat model of choroidal neovascularization. AB - PURPOSE: AG3340 (prinomastat) is a nonpeptidic, small-molecular-weight, synthetic matrix metalloproteinase inhibitor (MMPI) with selective inhibitory action of MMP 2, MMP-9, MMP-3, and MT-MMP1. We evaluated AG3340 injected intravitreally to treat choroidal neovascularization in a laser induced rat CNV model. METHODS: In the pretreatment group, the drug was injected the same day after induction of choroidal neovascularization by diode laser. In the treatment group, the drug was injected 2 weeks after induction of choroidal neovascularization (CNV). Fluorescein and indocyanine green angiography were performed to evaluate CNV. ERG recordings and histology were performed to assess toxicity and the CNV lesions. RESULTS: When used at the time of CNV induction, 62.8% of lesions in control versus 22.8% of the laser lesions in treated eyes developed CNV (p < 0.0001). The invading fibrovascular complex was thicker in the control eyes than that in the treated eyes. No signs of toxicity were detected. When used to treat established CNV, the percentage of leakage in treated and control eyes were 54.1% and 58.9% respectively (p > 0.05). Prinomastat was effective when given at the time of induction of CNV in the rat model. Administration of prinomastat 2 weeks after laser induction did not show efficacy. CONCLUSION: Prinomastat was active in the earliest stages of experimental CNV. It might be best used in combination with photodynamic therapy to inhibit recurrence of CNV from temporarily closed new vessels. PMID- 15279729 TI - Laser treatment of cultured retinal pigment epithelial cells-evaluation of the cellular damage in vitro. AB - PURPOSE: Evaluation of the effects of laser photocoagulation on cultured primary retinal pigment epithelial cells. METHODS: Cells were treated by a diode laser (678 nm) with 800 and 1600 mW for 0.186 second. Cell toxicity was tested by the WST-1 assay, and the uptakes of glutamate and gamma-aminobutyric acid (GABA) were measured. RESULTS: Laser photocoagulation (1600 mW) caused cell damage and the mitochondrial enzyme activity evaluated by a WST-1 test significantly decreased by 20%-30%. Laser treatment caused a dose-dependent decrease in glutamate uptake but increased GABA uptake. CONCLUSIONS: Laser treatment and the laser-induced increase in temperature influence transport processes in retinal pigment epithelial cells and may cause cell damage in the posterior part of the retina. PMID- 15279730 TI - The disposition and bioavailability of 35S-GSH from 35S-GSSG in BSS PLUS in rabbit ocular tissues. AB - The purpose of this study was to evaluate the biodistribution and uptake of 35S GSH into intraocular tissues following the administration of BSS PLUS containing 35S-GSSG by either an anterior chamber or intravitreal injection. This study evaluated the disposition and uptake of the 35S-radiolabel, the intracellular concentrations of 35S-GSH from extracellular 35S-GSSG, and the percentage of 35S GSH to the total cellular GSH pool. Glutathione was analyzed by high-performance liquid chromatography (HPLC) using fluorescence detection after derivitizing the thiols in situ with monobromobimane. The effluent from the GSH peak was then collected for measurement of 35S-GSH. After an anterior chamber injection of 35S BSS PLUS, 35S-radioactivity rapidly disappeared from the aqueous humor between 0.5 and 2 hours; corneal 35S-radioactivity remained constant over time. 35S-GSH was detected in the iris and ciliary body. However, in the cornea, 35S-GSH became the predominant radioactive thiol in the stroma, endothelium, and epithelium; the corneal stroma appeared to be a possible GSH reservoir for the adjacent corneal layers. After an intravitreal injection, 35S-radioactivity slowly decreased in the vitreous humor but was readily taken up by the tissues of the posterior segment, especially the retina and choroid, which showed the greatest concentrations of 35S-GSH of all tissues studied. The data from this study demonstrate that 35S-GSSG in BSS PLUS is metabolized and taken up by ocular cells and that 35S-GSH becomes incorporated into the intracellular GSH pool of ocular tissues. PMID- 15279731 TI - Ocular pharmacokinetics of fluocinolone acetonide after Retisert intravitreal implantation in rabbits over a 1-year period. AB - PURPOSE: The present study was designed to examine the pharmacokinetics of a fluocinolone acetonide (FA) intravitreal implant in pigmented rabbits. METHODS: Pigmented rabbits were randomly assigned to receive either a 0.5 mg or 2.0 mg FA intravitreal implant (Retisert). Four animals were sacrificed per time point (2 hours; 2 weeks; and 3, 6, 9, and 12 months after implantation) for FA intraocular levels determination. RESULTS: In the vitreous, concentration of FA was relatively constant from the first time point, 2 hours, through 1 year, and dose related, approximately seven- to eight-fold greater in the 2-mg implant. Concentrations of FA were generally higher in the vitreous (11-18 and 75-146 ng/g) and retina (42-87 and 224-489 ng/g) than in the aqueous humor (0.21-1.1 and 2.6-13.0 ng/g) for the 0.5- and 2-mg implants, respectively. Urine and plasma values were below the lower limit of quantitation (200 pg/mL) for all observations, indicating no evidence of systemic absorption. CONCLUSIONS: In this rabbit study, the Retisert provides relatively constant levels of FA in the posterior pole, which is consistent with previous reports of its clinical utility. PMID- 15279732 TI - An integrated approach to MS management. PMID- 15279733 TI - Multiple sclerosis in children. AB - Multiple sclerosis can develop during childhood, even among children under 10 years of age, and the initial diagnosis can be difficult. A first demyelinating event in children may be an episode of monophasic acute disseminated encephalomyelitis or a first episode of a macrophage activation syndrome, angiitis affecting the central nervous system or MS. The risk of developing MS is lower if: the child is younger than 10 years old; onset is associated with severely altered consciousness; presentation is polysymptomatic; or there are large and poorly limited lesions of the white matter. MS in children probably has a slightly better outcome than MS in adults. Initial treatment mainly relies on methylprednisolone, and there is little information on the results of beta interferon treatment in children with MS. PMID- 15279734 TI - Certification of multiple sclerosis nurses: an international perspective. AB - The Multiple Sclerosis Nursing International Certification Board was formed as a special initiative of the International Organization of Multiple Sclerosis Nurses (IOMSN). The aim was to develop a certification examination for MS nursing practice. This certification should achieve the vision of the IOMSN and unite MS nurses worldwide through standard practices. Such practices are based on common knowledge and skills, and tasks that encourage the best outcome for the nurse patient collaborative relationship. Certification allows recognition of an individual nurse's skill level, establishes a standard for all nurses treating patients with MS, improves patient care, and also benefits the neurological community. The Multiple Sclerosis Certified Nurse is revolutionary, as certification is an international effort to enhance and standardize MS care and develop MS nurse professionalism across borders. PMID- 15279735 TI - The effectiveness and practicality of immunosuppressive drugs. AB - Immunosuppression in MS is fundamentally linked to inflammation. Given that the irreversible deficits in MS are the consequence of axonal loss (the impact of other factors, such as neuronal loss, is less well understood), the critical therapeutic issue in MS is the relationship between inflammation and axonal injury. Two hypotheses for this relationship have been suggested: that inflammation and axonal injury are independent pathologies; or that they are dependent. In this latter case, an argument can be made for using an immunosuppressant. The optimal ways of administering immunosuppressive therapy may be early in the disease course, or as part of induction or combination therapy. One immunosuppressant (mitoxantrone) is licensed for the treatment of MS and some treatment efficacy and long-term surveillance data are available. More studies on the efficacy and use of immunosuppressants in MS are needed, however. PMID- 15279736 TI - Neuroprotection in multiple sclerosis report from the MS Forum 16th Modern Management Workshop, March 2004, Cernobbio, Italy. AB - Multiple sclerosis and other neurological disorders, such as stroke and Parkinson's disease, have a neurodegenerative component. This workshop reviewed the neurodegenerative processes involved in MS and other neurological conditions, with the aim of identifying neuroprotective strategies that could slow or halt MS disease progression. PMID- 15279737 TI - Transvaginal sonography--technique and limitations. PMID- 15279738 TI - Transvaginal versus transabdominal sonography in the evaluation of pelvic pathology. AB - OBJECTIVE: To find the accuracy of sonographic information provided by transvaginal sonography (TVS) in pelvic pathology as compared to transabdominal sonography (TAS). DESIGN: A comparative study. PLACE AND DURATION OF STUDY: The study was carried out in Military Hospital (MH) and Combined Military Hospital (CMH) Rawalpindi from January 2002 to June 2002. MATERIALS AND METHODS: Hundred patients were included in the study from a total of 212 referred for pelvic sonography. Two radiologists independently performed transabdominal and transvaginal sonography of these patients. An independent observer compared the findings. TVS was graded as superior, equal or inferior to TAS depending on the score assigned by them. RESULTS: TVS was considered superior in 63%, equal in 27% and inferior in 10% of the cases as compared to transabdominal sonography. It was graded inferior to TAS in cases with large pelvic masses and superior in majority of cases of ovarian follicle monitoring, polycystic ovaries, endometrial carcinoma and suspected ectopic pregnancy. Cases in which both techniques were considered equal included patients with no abnormal finding, some pelvic masses and advanced pelvic inflammatory disease. CONCLUSION: Transvagival sonography is superior to transabdominal sonography in most cases of pelvic pathology. However, TAS should still be the initial sonographic technique for routine evaluation of the female pelvis followed by TVS if indicated. In cases of ovarian follicle monitoring, suspected polycystic ovaries, endometrial pathology and suspected ectopic pregnancy, TVS may be used as the initial sonographic technique and can even replace TAS. PMID- 15279739 TI - Surgical management of epithelial parotid tumours. AB - OBJECTIVE: To describe the clinicopathological presentation and treatment options in epithelial parotid tumours with emphasis on surgery. DESIGN: Descriptive study. PLACE AND DURATION OF STUDY: ENT Departments of Pakistan Institute of Medical Sciences, Islamabad and Holy Family Hospital, Rawalpindi and Surgical Units at Rawalpindi General Hospital and DHQ Teaching Hospital, Rawalpindi, during a ten year period 1993-2003. SUBJECTS AND METHODS: Epithelial parotid tumours diagnosed and operated by an ENT surgeon and a general surgeon in 10 years during their posting in different teaching hospitals were included in the study. Clinical presentation, preoperative investigations, operative procedure, histopathology report, postoperative complications and further management were recorded. The data was collected and reviewed from the records of all the patients maintained by the authors. RESULTS: Fifty-two patients presented with parotid tumour. Average age was 38 years. Commonest presentation was a painless lump over the parotid region (85%), pain (15%), facial palsy, and enlarged neck nodes. Majority of tumours were benign, only two were recurrent. Parotid pleomorphic Adenoma (PPA) was the commonest benign tumour, others being Warthin's tumour and monomorphic adenoma. Adenoid cystic carcinoma was the commonest malignant tumour 29% followed by mucoepidermoid carcinoma. Others were carcinoma in PPA, squamous cell carcinoma, malignant mixed tumour, malignant lymphoepithelioma and undifferentiated carcinoma. Superficial parotidectomy (SP) was the commonest operation performed in 69%. Other procedures were total conservative parotidectomy in 11%, total radical surgery in 9% and enucleation in only one patient earliest in the series. Neck node dissection was done in 2 patients. Except for one child, rest of the 13 patients received postoperative radiotherapy and one patient of lymphoepithelioma received chemotherapy in addition. Commonest postoperative complication was temporary facial weakness in 35% (18/52). Permanent facial palsy occurred in 08 patients. Of these 07 had a malignant process and only one patient had excision biopsy. CONCLUSION: Benign and malignant epithelial parotid tumours can be diagnosed by their clinical presentation supplemented with FNAC. Superficial parotidectomy (SP) was the operation of choice. Facial nerve can be saved in total conservative parotidectomy for benign tumour in deep lobe and early malignant tumour. Radical parotidectomy followed by radiotherapy and in selected cases neck node dissection are the recommended procedures for advanced malignant parotid tumours. PMID- 15279740 TI - Glucose 6 phosphate dehydrogenase deficiency in adults. AB - OBJECTIVE: To determine the frequency of glucose-6-phosphate dehydrogenase (G6PD) deficiency in adults presented with anemia. DESIGN: Case-review. PLACE AND DURATION OF STUDY: District Headquarter Hospital Batkhela, Malakand Agency, from March 1999 to August 2000. SUBJECTS AND METHODS: Eighteen months admission data was reviewed for G6PD deficiency as a cause of anemia. Anemia was defined by world health organization (WHO) criteria as haemoglobin less than 11.3 gm%. G6PD activity was measured by Sigma dye decolorisation method. All patients were screened for complications of hemolysis and its possible cause. Patients with more than 13 years of age were included in the study. RESULTS: Out of 3600 patients admitted, 1440 were found anaemic and 49 as G6PD deficient. So the frequency of G6PD deficiency in anaemic patients was 3.4% and the overall frequency is 1.36%. G6PD deficiency among males and females was three and six percent respectively. Antimalarials and antibiotics containing sulphonamide group were the most common precipitating factors for hemolysis. Anemia and jaundice were the most common presentations while malaria was the most common associated disease. Acute renal failure was the most severe complication occurring in five patients with two deaths. CONCLUSION: G6PD deficiency is a fairly common cause of anemia with medicine as common precipitating factor for hemolysis. Such complications can be avoided with early recognition of the disease and avoiding indiscriminate use of medicine. PMID- 15279741 TI - Hematological evaluation of splenomegaly. AB - OBJECTIVE: To find out the relative frequency of clinical conditions associated with splenomegaly that require hematological evaluation in our set up. DESIGN: Cross sectional study. PLACE AND DURATION OF STUDY: Combined Military Hospital, Quetta, Balochistan, from July 2000 to July 2003. SUBJECTS AND METHODS: Patients of either gender and all age groups with palpable spleen were included. Patients with splenomegaly due to liver disease, malarial parasites on thick or thin blood film, positive Widal test, or positive blood cultures were excluded from study. Patients were initially evaluated with clinical history, microscopic examination of blood smear, and blood counts. Depending upon provisional diagnosis bone marrow examination or investigations for hemolytic anemia were performed. RESULTS: One hundred patients were received. Seventy-eight patients were adults and 22 patients were of pediatric age group. In the adults, hematological malignancies were seen in 37%, malarial parasites in bone marrow in 20.5%, megaloblastic anemia in 13%, bacterial infections in 9%, hemolytic anemia in 9%, tropical splenomegaly in 5%, and positive bone marrow culture for salmonella in 6.5%. In children, hematological evaluation revealed hematological malignancies in 18%, beta thalassaemia in 55%, other hemolytic anemias in 13.5%, congenital sideroblastic anemia in 4.5%, and storage disorder in 9%. CONCLUSION: Hematological workup is informative in most of the cases. Bone marrow examination is the key investigation, hematological malignancies constituted 37% of the adult and 18% of pediatric age group patients. Hemolytic anemia constituted 68% of pediatric age group. PMID- 15279742 TI - Brain abscess--diagnosis and management. AB - OBJECTIVE: To evaluate the clinical presentation, diagnosis, sources of infection, surgical management outcome and microorganisms involved in the brain abscess in our locality. DESIGN: Descriptive study. PLACE AND DURATION OF STUDY: The department of Neurosurgery, Chandka Medical College Hospital, Larkana from July 1998 to June 2003. SUBJECTS AND METHODS: All patients who were confirmed cases of brain abscess were entered into the study. Data collected on proforma, contained categories of age, gender, clinical presentation, diagnostic laboratory findings, computed tomography scan reports, associated anomalies, surgical management, culture reports antibiotic therapy, microbiologic features and treatment out come. RESULTS: Out of 82 patients, 58 were males and 24 females. Mean age was 18 years (range 05 months to 55 years). Headache with papilloedema was the commonest presentation (82%). Neurological deficit was present in 46%. A source of infection was present in 89%. Otogenic source was the commonest (63%). CT scan was diagnostic in all (100%) cases. Solitary abscess was found in 79% of the cases while in 21% of the cases multiple abscess were found. Temporal lobe was the commonest site involved (55%). Cultures were found positive for microorganism in 82% of the cases. Bacteriodes (38%) and Streptococci (25%) were the commonest isolates. Burr hole aspiration was done in only 38% of the cases while excision of the capsule along with aspiration was carried out in 62% of the cases. Over all morality was 22% in this series; causes of death were septicemia, ventriculitis and pneumonia. CONCLUSION: Diagnosis with CT scan, appropriate antibiotic therapy and complete removal of abscess along with excision of capsule could reduce the mortality and neurological deficits from brain abscess. PMID- 15279743 TI - Renal and post-renal causes of acute renal failure in children. AB - OBJECTIVE: To identify the causes of acute renal failure (ARF) in pediatric population along with the identification of the age and gender most affected by the failure. DESIGN: Descriptive study. PLACE AND DURATION OF STUDY: This study was carried out at the Pediatric Medical Ward of National Institute of Child Health (NICH), Karachi over a period of two years from 1996 to 1998. SUBJECTS AND METHODS: The study included children under the age of 12 years who presented with signs and symptoms suggestive of ARF (oliguria/anuria, vomiting, acidotic breathing etc.) along with raised blood urea nitrogen (BUN) serum creatinine and metabolic acidosis as shown by arterial blood gases (ABGs). Patients were divided into two groups on the basis of age; group A consisting of 0-2 years and group B from >2 years. Patients presenting with transient pre-renal azotaemia were excluded from the study. After providing initial emergency cover, detailed history, physical examination and investigations were carried out according to a proforma specially designed to ascertain the cause of ARF. Patients were managed for ARF as per standard recommendations and investigations completed or repeated as and when required. RESULTS: A total of 119 patients with ARF were admitted in the ward over a period of two years constituting 1.36% of the total admissions and 16.39% of the admissions due to renal pathology. Mean age of presentation was 4.5 years with 56.7% of the patients under the age of 5 years. Male predominance was noted in all ages with an overall male to female ratio of 2.3:1. Most common cause leading to ARF in younger age group was found to be hemolytic uremic syndrome [25(54.34%)] followed by septicemia [7(15.21%)]. In older patients renal calculus disease was the most common [22(30.13%)] underlying pathology followed by pre-existing, undiagnosed chronic renal failure [16(21.91%)]. More than 90% of cases were related, directly or indirectly, to infections. CONCLUSION: ARF is fairly common in children especially under the age of 5 years showing a male predominance. More than 90% of the cases can be prevented by improving primary health care and by early and prompt treatment of infections. PMID- 15279744 TI - Degloving injuries of the lower limb. AB - OBJECTIVE: To identify the pattern of degloving injuries of the lower limb to help in designing a management protocol for such injuries. DESIGN: Descriptive study. PLACE AND DURATION OF STUDY: Department of Plastic and Reconstructive Surgery, Hayatabad Medical Complex, Peshawar over a period of 3 years from September 1998 to August 2001. SUBJECTS AND METHODS: Fifty patients with degloving injuries of the lower limb were included, type, cause, extent and location of degloving injury was identified in all of them. RESULTS: Majority of the patients were males (74%) with a median age of 14 years. The most common cause of trauma was roadside accidents (96%). Left lower limb involvement was more common (72%). Sixty-six per cent of the cases sustained trauma to the leg. The type of degloving injury observed more frequently was open degloving in 94% of the patients. Fifty one percent patients had proximally attached degloved skin. CONCLUSION: The pattern of degloving injuries of the lower limb emerging from this study will help clinicians to identify the pattern of these injuries and develop a management protocol. PMID- 15279745 TI - Smear-negative pulmonary tuberculosis and lymphocyte subsets. AB - OBJECTIVE: To correlate and quantitate lymphocyte subsets with clinically diagnosed smear-negative pulmonary tuberculosis and severity of disease. DESIGN: Case-control study. PLACE AND DURATION OF STUDY: Military Hospital and Armed Forces Institute of Pathology Rawalpindi. 1999-2000. SUBJECTS AND METHODS: Freshly diagnosed, well-characterized smear-negative patients (n=15) of pulmonary tuberculosis were selected. Non-induced three-consecutive negative smears of sputum with simultaneous culture for AFB for 6-8 weeks, positive Mauntoux test (Z10 mm), blood complete picture with ESR and chest x-rays were done. Selected panel of monoclonal antibodies against specific CD markers were used. Statistical analysis done by student t-test or Mann-Whitney rank-sum test with the help of Sigma State software. RESULTS: Hemoglobin and total lymphocyte counts were significantly reduced whereas total leukocyte counts with absolute neutrophil counts were increased. Fraction of CD4+ and CD8+ T lymphocytes with HLA-DR expression was reduced while no significant change in rest of the TB and NK lymphocytes. CONCLUSION: Low hemoglobin level, high neutrophil count and low total lymphocytes suggest possible direct relationship with extent of disease. The number of activated CD4+, CD8+, ab and gd TCR T cells have tendency to increase during Mycobacterium infection. This seems to have a potential of being a good, non-invasive prognostic indicator for patients with pulmonary tuberculosis. PMID- 15279746 TI - Klippel Trenaunay syndrome. AB - Klippel-Trenaunay syndrome (KTS) is a congenital vascular disorder of unknown cause, characterized by port wine stain (capillary malformations), venous malformation and limb hypertrophy. We present a case of this rare syndrome in a young girl. PMID- 15279747 TI - Epiploic appendagitis. AB - This case report describes epiploic appendagitis in an elderly lady who presented with acute abdomen. The condition was diagnosed on CT scan and the patient treated conservatively. PMID- 15279748 TI - Two displaced intrauterine contraceptive devices (copper-T). AB - An unusual case of two displaced intrauterine contraceptive devices (copper-T) is described in a 34 years old lady. One copper-T was found in the urinary bladder with stone formation around it and the other embedded in the posterior wall of the urinary bladder. This was managed operatively and the patient made a smooth recovery. PMID- 15279749 TI - Cardiac metastases of osteosarcoma. AB - Osteosarcoma is a malignancy whose various sites of metastasis greatly modify its ultimate prognosis. We report a case of simultaneous pulmonary and cardiac metastases in a 41-year-old male patient with osteosarcoma of the tibia, presenting after more then one year of completion of adjuvant therapy with progressive dyspnea and cyanosis. Diagnosis was made on computerized tomogram and echocardiogram. The metastatic mass entirely occupying the right ventricle and the pulmonary artery proved fatal. PMID- 15279750 TI - Salmonella osteomyelitis. AB - A case report of a young girl with salmonella osteomyelitis of left radius is presented having no constitutional symptoms. X-ray, changes are described. Curettage tissue sent for culture and histology. Histology showed non-specific osteomyelitis while salmonella was grown. She was treated accordingly and got cured with three year follow-up. PMID- 15279751 TI - Severe rhabdomyolysis due to drug overdose. AB - A case of a young girl is described who presented in a state of intoxication and later developed acute renal failure secondary to severe rhabdomyolysis. With vigorous fluid therapy she regained normal renal function. Biochemical testing finally confirmed over the counter drug overdose as the cause for this presentation. PMID- 15279752 TI - Irritable bowel syndrome in patients evaluated for organic bowel pathology. PMID- 15279753 TI - Acute respiratory infections in Pakistan: have we made any progress? AB - Acute respiratory infections (ARI) are the leading cause of death in young children in Pakistan, responsible for 20-30% of all child deaths under age 5 years. This paper summarizes the research and technical development efforts over the last 15 years which have contributed to improving the effectiveness of the case management strategy to reduce mortality from pneumonia in children in Pakistan. Community intervention is viable, effective and practical. Rising antimicrobial resistance among commonly used and low-cost oral agents is of significant concern. Appropriate monitoring and evaluation of the impact of the ARI control programme is lacking. Lack of funding for programmatic activities, lack of coordination with other child survival programs, inadequate training for community health workers and general practitioners in the private sector, lack of public awareness about seeking timely and appropriate care, and insufficient planning and support for ARI programmatic activities at provincial and district levels are major hindrances in decreasing the burden of ARI in the country. The recent introduction of the community-based Lady Health Worker (LHW) Programme and WHO and UNICEF-sponsored integrated management of childhood illness initiative present ideal opportunities for re-emphasizing early case detection and appropriate case management of ARI. Ultimately, focusing on preventive strategies such as improving nutrition, reducing indoor pollution, improving mass vaccination, as well as introduction of new vaccines effective against important respiratory pathogens will likely have the most impact on reducing severe ARI and deaths from severe disease. PMID- 15279754 TI - Health systems research and management: building capacity of district health managers. PMID- 15279755 TI - Comparative study of Mantoux test versus BCG inoculation as a diagnostic tool for pulmonary tuberculosis in children up to 5 years of age. PMID- 15279756 TI - Role of fixateur interne in thoracolumbar junction injuries. PMID- 15279757 TI - Surgical Treatment of Refractory Epilepsy. AB - The extension of cortical resection to treat children with intractable epilepsy is one of the most exciting advances in pediatric neurology in the past two decades. Many children with epilepsy who previously had little hope have been given a new chance at life. Many patients who were not considered for surgery are now recognized to be excellent surgical candidates. Most notably, children with generalized seizures such as infantile spasms or gelastic seizures caused by hypothalamic hamartomas now have the opportunity to benefit from surgery. In adults, there is one goal for epilepsy surgery: freedom from seizures. This is an important goal but it is not the only one in children. It may not even be the most important goal. When operating on young children with epilepsy, we seek to alter the long-term developmental and behavioral outcome. Although there are many significant recent advances in pediatric epilepsy surgery, three are particularly important. These are 1) hemispherotomy for children who require hemispheric resection; 2) resection of hypothalamic hamartomas through an innovative transcallosal approach; and 3) resection of multiple tubers in children with tuberous sclerosis complex. PMID- 15279759 TI - New Treatments in Pediatric Brain Tumors. AB - The management of childhood brain tumors is likely to change dramatically during the next few years. Current treatment has improved outcome in some types of brain tumors, but for most patients, survival rates have not changed during the past two decades. Advances in surgery, radiation therapy, and chemotherapy are likely to marginally increase survival, and possibly improve the quality of life for long-term survivors. As the molecular factors underlying childhood brain tumors are better elucidated, molecular-targeted therapy will become a major modality of treatment with the promise of not only increasing the likelihood of survival, but also decreasing treatment-related sequelae. PMID- 15279758 TI - Stroke in Children with Sickle Cell Disease. AB - Children with sickle disease are at high risk for ischemic stroke and transient ischemic attacks, usually secondary to intracranial arteriopathy involving the terminal internal carotid and proximal middle cerebral and anterior cerebral arteries, which may be diagnosed using transcranial Doppler ultrasound or magnetic resonance angiography (MRA). Other central nervous system (CNS) complications include seizures and coma, which may be secondary to ischemic stroke, sinovenous thrombosis, reversible posterior leukoencephalopathy, or acute demyelination. The immediate priority after an acute CNS event is to improve cerebral oxygenation with oxygen supplementation to maintain peripheral saturation measured using pulse oximetry between 96% and 99%, and a simple transfusion of packed cells within an hour of presentation if the patient's hemoglobin is less than 10 g/dL. The patient then should have erythrocytapheresis or manual exchange to reduce the hemoglobin S percentage to below 30%. Computed tomography to exclude hemorrhage is mandatory and MR T2-weighted imaging with MRA, fat-saturated imaging of the neck (dissection), MR venography (sinovenous thrombosis), and diffusion-weighted imaging usually distinguishes between arterial ischemic stroke and the differential diagnoses. Comatose patients with widespread focal or global cerebral edema may have good functional outcome after surgical decompression. Anticoagulation may be indicated for dissection or sinovenous thrombosis and steroids for demyelination. Blood pressure should be reduced slowly if raised in patients with reversible posterior leukoencephalopathy. Seizures should be treated aggressively and electroencephalogram monitoring should be done to exclude subclinical seizures if the patient is unconscious. Hemorrhagic stroke may require craniectomy and drainage and/or management of vasospasm. Interventional neuroradiology with coils is an alternative to surgical clipping for aneurysms. For secondary prevention, regular blood transfusion to hemoglobin S of less than 30% reduces the risk of recurrent stroke from approximately 67% to approximately 10%. Hydroxyurea and phlebotomy may be used in patients who are alloimmunized. Moyamoya syndrome is a risk factor for recurrence despite prophylactic blood transfusion. Revascularization may prevent additional stroke. Bone marrow transplantation may be offered to patients with human leukocyte antigen-compatible siblings. Blood transfusion prevents stroke in patients with velocities greater than 200 cm per second on TCD; a phase III trial studying the prevention of the progression of silent infarction is being done. Emerging primary prophylaxis regimens being tested include citrulline and arginine, aspirin, and overnight oxygen supplementation. Physicians caring for children with sickle cell disease also should ensure adequate nutrition, including five servings of fruit and vegetables a day. The role of vitamin supplementation is controversial, particularly when patients must take daily penicillin prophylaxis. PMID- 15279760 TI - Early Diagnosis of Autism Spectrum Disorders. AB - Autistic spectrum disorders (ASD) are an often-disabling continuum of disorders affecting two to four in 1000 children. These disorders have a core set of defining features including impaired verbal and nonverbal communication, impaired social interaction, and restricted or repetitive patterns of behavior. The cause of autism is unclear. The disorder can be defined only by related behaviors. Although there has been considerable improvement in standardized screening techniques for ASD in the past 10 years, screening and diagnostic practices in medicine and education lag far behind clinical research. Various studies have found the average age of diagnosis to be between 3 and 6 years, with significant differences as a function of ethnicity and socio-economic status. Preliminary research suggests that in some populations, missed diagnosis and misdiagnosis of ASD are common. This may be caused partly by inadequate screening practices. It also may reflect that presentation of symptoms varies from patient to patient. Lack of resources for appropriate referral, diagnosis, and treatment may play an important role. This article discusses recent progress in ASD screening, what is known of current screening and diagnostic practices, and future directions for research and practice improvement. The best practice model for the screening and early diagnosis of autism spectrum disorders and other developmental disabilities should include routine developmental surveillance as part of well-child pediatric care. General developmental screening should be followed by autism-specific screening for those children who fail the initial developmental screen, or whose parents report suspect behaviors. PMID- 15279761 TI - Poststroke Neuropsychiatric Illness: An Integrated Approach to Diagnosis and Management. AB - Patients who have had stroke are at significant risk for various neuropsychiatric illnesses. The most common and important of these are poststroke depression and poststroke dementia (attributable to vascular dementia, Alzheimer's dementia, or a combination of mechanisms). Poststroke neuropathology may lead some patients to experience concurrent and "overlapping" mood and cognitive symptoms. Less frequently, poststroke anxiety disorders, psychosis, isolated pathologic expressions of emotions, and apathy or fatigue may be encountered. The authors review the current literature on poststroke neuropsychiatry and offer an integrated approach to pathophysiologic concepts and clinical surveillance, screening, diagnosis, and evidence-based pharmacologic and nonpharmacologic intervention for these clinical problems on the clinical boundary between neurology and psychiatry. PMID- 15279762 TI - Substance Use Disorders and Neurologic Illness. AB - Because of the high rates of substance use disorders among the general and clinical populations, and the abuse potential of many medications commonly used in the treatment of neurologic illnesses, the treating neurologist must deal with drug misuse and abuse in practice. The most important tool neurologists must have in their arsenal is the ability to assess for and recognize substance use disorders in their patients. Any treatment plan developed for such patients must include ongoing management of substance abuse issues. After a substance use disorder is diagnosed, the neurologist must make proper referrals to adjunctive support interventions (Alcoholics Anonymous or Narcotics Anonymous) and chemical dependency specialists, and work closely and in collaboration with these components of the patient's overall treatment. The treating neurologist should be aware of the myriad neurologic sequelae of drug use, because most drugs of abuse, including alcohol, can have neurologic manifestations resulting from acute intoxication, acute withdrawal, or chronic use. Drug use (past and present) should be included in the differential diagnosis for any patient with an atypical constellation of symptoms or with isolated neurologic deficits. If the treatment of a neurologic condition requires the use of a potentially addictive substance, particularly when the patient has a history of substance use disorders, then the clinician must minimize the risk of addiction by giving the least addictive substances and developing a plan to manage the use of the drug throughout the treatment period. PMID- 15279764 TI - The cellular response to general and programmed DNA double strand breaks. AB - DNA double strand breaks (DSBs) are among the most dangerous lesions that can occur in the genome of eukaryotic cells. Proper repair of chromosomal DSBs is critical for maintaining cellular viability and genomic integrity and, in multi cellular organisms, for suppression of tumorigenesis. Thus, eukaryotic cells have evolved specialized and redundant molecular mechanisms to sense, respond to, and repair DSBs. In this chapter, we provide an overview of the progress that has been made over the last decade in elucidating the identity and function of components that participate in the cellular response to chromosomal DSBs. Then, we discuss, in more depth, the response to DSBs that occur in the context of the V(D)J recombination and IgH class switch recombination reactions that occur in cells of the lymphocyte lineage. PMID- 15279765 TI - DSB repair: the yeast paradigm. AB - Genome stability is of primary importance for the survival and proper functioning of all organisms. Double-strand breaks (DSBs) arise spontaneously during growth, or can be created by external insults. In response to even a single DSB, organisms must trigger a series of events to promote repair of the DNA damage in order to survive and restore chromosomal integrity. In doing so, cells must regulate a fine balance between potentially competing DSB repair pathways. These are generally classified as either homologous recombination (HR) or non homologous end joining (NHEJ). The yeast Saccharomyces cerevisiae is an ideal model organism for studying these repair processes. Indeed, much of what we know today on the mechanisms of repair in eukaryotes come from studies carried out in budding yeast. Many of the proteins involved in the various repair pathways have been isolated and the details of their mode of action are currently being unraveled at the molecular level. In this review, we focus on exciting new work eminating from yeast research that provides fresh insights into the DSB repair process. This recent work supplements and complements the wealth of classical genetic research that has been performed in yeast systems over the years. Given the conservation of the repair mechanisms and genes throughout evolution, these studies have profound implications for other eukaryotic organisms. PMID- 15279766 TI - The mechanism of vertebrate nonhomologous DNA end joining and its role in V(D)J recombination. AB - The vertebrate immune system generates double-strand DNA (dsDNA) breaks to generate the antigen receptor repertoire of lymphocytes. After those double strand breaks have been created, the DNA joinings required to complete the process are carried out by the nonhomologous DNA end joining pathway, or NHEJ. The NHEJ pathway is present not only in lymphocytes, but in all eukaryotic cells ranging from yeast to humans. The NHEJ pathway is needed to repair these physiologic breaks, as well as challenging pathologic breaks that arise from ionizing radiation and oxidative damage to DNA. PMID- 15279767 TI - Homologous recombination-mediated double-strand break repair. AB - Exchange of DNA strands between homologous DNA molecules via recombination ensures accurate genome duplication and preservation of genome integrity. Biochemical studies have provided insights into the molecular mechanisms by which homologous recombination proteins perform these essential tasks. More recent cell biological experiments are addressing the behavior of homologous recombination proteins in cells. The challenge ahead is to uncover the relationship between the individual biochemical activities of homologous recombination proteins and their coordinated action in the context of the living cell. PMID- 15279768 TI - DNA recombination, chromosomal stability and carcinogenesis: insights into the role of BRCA2. AB - Germline mutations affecting a single allele of BRCA2 increase susceptibility to breast and ovarian cancer, whilst germline inheritance of certain bi-allelic mutations causes a Fanconi anaemia-like syndrome. Here, we review current knowledge of the BRCA2 protein, focussing on recent studies that provide mechanistic insight into its biological function in regulating DNA recombination reactions mediated by the RAD51 recombinase. We argue that the chromosomal instability and cancer predisposition provoked by BRCA2 inactivation are a consequence of the failure to re-start stalled DNA replication, and to repair DNA double-strand breaks, through error-free pathways that depend on homologous pairing between DNA strands. PMID- 15279769 TI - The Mre11 complex and the metabolism of chromosome breaks: the importance of communicating and holding things together. AB - The conserved Mre11 complex (Mre11, Rad50, and Nbs1) plays a role in each aspect of chromosome break metabolism. The complex acts as a break sensor and functions in the activation and propagation of signaling pathways that govern cell cycle checkpoint functions in response to DNA damage. In addition, the Mre11 complex influences recombinational DNA repair through promoting recombination between sister chromatids. The Mre11 complex is required for mammalian cell viability but hypomorphic mutants of Mre11 and Nbs1 have been identified in human genetic instability disorders. These hypomorphic mutations, as well as those identified in yeast, have provided a benchmark for establishing mouse models of Mre11 complex deficiency. In addition to consideration of Mre11 complex functions in human cells and yeast, this review will discuss the characterization of mouse models and insight gleaned from those models regarding the metabolism of chromosome breaks. The current picture of break metabolism supports a central role for the Mre11 complex at the interface of chromosome stability and the regulation of cell growth. Further genetic analysis of the Mre11 complex will be an invaluable tool for dissecting its function on an organismal level and determining its role in the prevention of malignancy. PMID- 15279770 TI - NBS1 and its functional role in the DNA damage response. AB - Nijmegen breakage syndrome is a recessive genetic disorder, characterized by elevated sensitivity to ionizing radiation, chromosome instability and high frequency of malignancies. Since cellular features partly overlap with those of ataxia-telangiectasia (A-T), NBS was long considered an A-T clinical variant. NBS1, the product of the gene underlying the disease, contains three functional regions: the forkhead-associated (FHA) domain and BRCA1 C-terminus (BRCT) domain at the N-terminus, several SQ motifs (consensus phosphorylation sites by ATM and ATR kinases) at a central region and MRE11-binding region at the C-terminus. NBS1 forms a multimeric complex with hMRE11/hRAD50 nuclease at the C-terminus and recruits or retains them at the vicinity of sites of DNA damage by direct binding to histone H2AX, which is phosphorylated by ATM in response to DNA damage. The combination of the FHA/BRCT domains has a crucial role for the binding of NBS1 to H2AX. Thereafter, the NBS1 complex proceeds to rejoin double-strand breaks predominantly by homologous recombination repair in vertebrates, while it also might be involved in suppression of inter-chromosomal recombination even for V(D)J recombination. These processes collaborate with cell cycle checkpoints to facilitate DNA repair, while defects of these checkpoints in NBS cells are partial in nature. A possible explanation for these moderate defects are the redundancy of multiple checkpoint regulations in vertebrates, or the modulator role of NBS1, in which NBS1 amplifies ATM activation by accumulation of the MRN complex at damaged sites. This molecular link of NBS1 to ATM may explain the phenotypic similarity of NBS to A-T. PMID- 15279771 TI - Structure and function of the double-strand break repair machinery. AB - The discovery of the recA gene toward the middle of the 20th century sparked work in perhaps one of the most biochemically and biophysically intriguing systems of DNA repair-homologous recombination. The inner workings of this system, in particular those of the more complex eukaryotes, have been and in many ways remain mysterious. Yet at the turn of this century, a wealth of structural and genetic results has unveiled a detailed picture of the roles, relationships, and mechanics of interacting homologous recombination proteins. Here we focus on the predominant questions addressed by these exciting 21st century structural results from detection of broken DNA ends to coordination of pathway progression. The emerging structural view of double-strand break repair, therefore, reveals the molecular basis both for functions specific to DNA recombination and for general features characterizing DNA repair processes. PMID- 15279772 TI - DNA single-strand breaks and neurodegeneration. AB - The association of human genetic disorders with defects in the DNA damage response is well established. Most of the major DNA repair pathways are represented by diseases in which that pathway is absent or impaired, including those responsible for repairing DNA double-strand breaks. Conspicuous by their absence, however, have been human disorders associated with defects in the repair or response to DNA single-strand breaks (SSBs). However, three papers have recently associated hereditary spinocerebellar ataxia with mutations in genes connected with SSBR. The emerging links between SSBR and neurodegeneration are discussed. PMID- 15279773 TI - PI 3-kinase related kinases: 'big' players in stress-induced signaling pathways. AB - The phosphoinositide 3-kinase related kinases (PIKKs) comprise a family of high molecular mass signaling proteins that play central roles in the control of cell growth, gene expression, and genome surveillance and repair in eukaryotic cells. Mammalian cells express six PIKK family members, five of which-ATM, ATR, mTOR, DNA-PK, and hSMG-1-function as protein serine-threosine kinases. This overview provides some general insights into the pharmacology, biochemistry, and function of this nonconventional group of protein kinases. PMID- 15279774 TI - DNA damage-induced activation of ATM and ATM-dependent signaling pathways. AB - Ataxia-telangiectasia mutated (ATM) plays a key role in regulating the cellular response to ionizing radiation. Activation of ATM results in phosphorylation of many downstream targets that modulate numerous damage response pathways, most notably cell cycle checkpoints. In this review, we describe recent developments in our understanding of the mechanism of activation of ATM and its downstream signaling pathways, and explore whether DNA double-strand breaks are the sole activators of ATM and ATM-dependent signaling pathways. PMID- 15279775 TI - Regulation of DNA replication by ATR: signaling in response to DNA intermediates. AB - The nuclear protein kinase ATR controls S-phase progression in response to DNA damage and replication fork stalling, including damage caused by ultraviolet irradiation, hyperoxia, and replication inhibitors like aphidicolin and hydroxyurea. ATR activation and substrate specificity require the presence of adapter and mediator molecules, ultimately resulting in the downstream inhibition of the S-phase kinases that function to initiate DNA replication at origins of replication. The data reviewed strongly support the hypothesis that ATR is activated in response to persistent RPA-bound single-stranded DNA, a common intermediate of unstressed and damaged DNA replication and metabolism. PMID- 15279776 TI - Role of DNA-PK in the cellular response to DNA double-strand breaks. AB - The DNA-dependent protein kinase (DNA-PK) plays a critical role in DNA double strand break (DSB) repair and in V(D)J recombination. DNA-PK also plays a very important role in triggering apoptosis in response to severe DNA damage or critically shortened telomeres. Paradoxically, components of the DNA-PK complex are present at the mammalian telomere where they function in capping chromosome ends to prevent them from being mistaken for double-strand breaks. In addition, DNA-PK appears to be involved in mounting an innate immune response to bacterial DNA and to viral infection. As DNA-PK localizes very rapidly to DNA breaks and phosphorylates itself and other damage-responsive proteins, it appears that DNA PK serves as both a sensor and a transducer of DNA-damage signals. The many roles of DNA-PK in the mammalian cell are discussed in this review with particular emphasis on recent advances in our understanding of the phosphorylation events that take place during the activation of DNA-PK at DNA breaks. PMID- 15279777 TI - The ATM-related kinase, hSMG-1, bridges genome and RNA surveillance pathways. AB - Recent studies have identified, hSMG-1 as the newest member of the phosphoinositide 3-kinase(PI3-kinase)-related kinase (PIKK) family. The protein kinase activity of hSMG-1 resembles that of the related PIKK, ATM, both in terms of substrate specificity and its sensitivity to inhibition by the fungal metabolite wortmannin. hSMG-1 is the ortholog of a Caenorhabditis elegans protein, CeSMG-1, which has been genetically linked to a critical mRNA surveillance pathway termed nonsense-mediated decay (NMD). The function of NMD is to mark for rapid degradation mRNAs that bear a premature termination codon. Compelling evidence now indicates that hSMG-1 is also a central player in the NMD pathway in human cells. In addition, hSMG-1, like ATM, appears to be involved in the recognition and/or repair of damaged DNA in these cells. In this review, we introduce a model in which hSMG-1 teams with ATM and ATR to insure the overall quality of the transcriptome in human cells. PMID- 15279778 TI - The multifaceted role of mTOR in cellular stress responses. AB - The mammalian target of rapamycin (mTOR) is a large multidomain protein whose function is inhibited by the immunosuppressant drug rapamycin. mTOR (or its homologues in lower eukaryotes) plays roles in cell growth and the cell cycle, control of the cytoskeleton and nutrient transport, protein and RNA stability and transcription and translation. In mammalian cells, the best understood effectors of mTOR are proteins involved in controlling the translational machinery. Signalling through mTOR is stimulated by amino acids and by hormones and mitogens. On the other hand, mTOR signaling is impaired in response to a range of stressful stimuli. These include DNA damage, nutrient withdrawal and depletion of cellular energy, as well as hypoxia. In response, e.g. to DNA damage, impairment of mTOR signaling appears to precede the commitment of cells to apoptosis. The mechanisms by which these stressful conditions still remain largely unclear. However, these responses make physiological sense, as impairment of mTOR signalling under these conditions will tend to inhibit anabolic processes and cell growth and division. PMID- 15279779 TI - The DNA double-strand break response pathway: becoming more BRCAish than ever. AB - Breast carcinoma is the leading cause of cancer incidence, and second in cancer mortality to lung cancer, in women of the Western hemisphere. Germ line mutations in the breast cancer susceptibility gene, BRCA1, is responsible for half of all cases of hereditary breast cancer, which constitutes about 5-10% of all cases of breast cancer. Current hypothesis has ascribed a role for Brca1 in maintaining genomic stability, through its involvement in cellular response pathway to the DNA double-strand breaks (DSB). DNA DSB, which are the most deleterious form of DNA damage, are repaired through a series of coordinated steps embedded in a signal transduction pathway that ultimately ensure the elimination of potentially harmful mutations to the genome. This pathway can be crudely divided into a primary and secondary phase. The primary response phase is initiated by sensor proteins that activate transducer protein kinases Atm and Atr, which target downstream effector proteins, such as Chk1 and Chk2, to elicit the secondary response phase. Brca1 has been intimately linked with various aspects of this signaling pathway. However, the precise role of Brca1 in this process remains unclear. In this review, we will provide a simple model in an attempt to clarify the role of Brca1 during cellular response to DNA DSB. PMID- 15279780 TI - 53BP1, an activator of ATM in response to DNA damage. AB - p53 Binding protein 1 (53BP1) belongs to a family of evolutionarily conserved DNA damage checkpoint proteins with C-terminal BRCT domains and is most likely the human ortholog of the budding yeast Rad9 protein, the first cell cycle checkpoint protein to be described. 53BP1 localizes rapidly to sites of DNA double strand breaks (DSBs) and its initial recruitment to these sites has not been shown to be dependent on any other protein. Initially, 53BP1 was thought to be a mediator of DNA DSB signaling, but now it has been shown to function upstream of ataxia telangiectasia mutated (ATM), in one of at least two parallel pathways leading to ATM activation in response to DNA damage. Currently, only a single tudor and two BRCT domains are recognized in 53BP1; however, their precise functional role is not understood. Elucidating the function of 53BP1 will be critical to understanding how cells recognize DNA DSBs and how ATM is activated. PMID- 15279781 TI - MDC1/NFBD1: a key regulator of the DNA damage response in higher eukaryotes. AB - The protein MDC1/NFBD1 contains a forkhead-associated (FHA) domain and two BRCA1 carboxyl-terminal (BRCT) domains. It interacts with several proteins involved in DNA damage repair and checkpoint signalling, and is phosphorylated in response to DNA damage and during mitosis. Upon treatment of cultured human cells with DNA damaging agents, MDC1/NFBD1 translocates to sites of DNA lesions, where it collaborates with other proteins and with phosphorylated histone H2AX to mediate the accumulation of checkpoint and repair factors into nuclear foci. Down regulation of MDC1/NFBD1 expression levels by small interfering RNA (siRNA) renders cells hyper-sensitive to DNA damaging agents and leads to defects in cell cycle checkpoint activation and apoptosis. Thus, MDC1/NFBD1 appears to be a key regulator of the DNA damage response in mammalian cells. PMID- 15279782 TI - H2AX: the histone guardian of the genome. AB - At close hand to one's genomic material are the histones that make up the nucleosome. Standing guard, one variant stays hidden doubling as one of the core histones. But, thanks to its prime positioning, a variation in the tail of H2AX enables rapid modification of the histone code in response to DNA damage. A role for H2AX phosphorylation has been demonstrated in DNA repair, cell cycle checkpoints, regulated gene recombination events, and tumor suppression. In this review, we summarize what we have learned about this marker of DNA breaks, and highlight some of the questions that remain to be elucidated about the physiological role of H2AX. We also suggest a model in which chromatin restructuring mediated by H2AX phosphorylation serves to concentrate DNA repair/signaling factors and/or tether DNA ends together, which could explain the pleotropic phenotypes observed in its absence. PMID- 15279783 TI - Interplay between chromatin and cell cycle checkpoints in the context of ATR/ATM dependent checkpoints. AB - Maintenance of both genome stability and its structural organization into chromatin are essential to avoid aberrant gene expression that could lead to neoplasia. Genome integrity being threatened by various sources of genotoxic stresses, cells have evolved regulatory mechanisms, termed cell cycle checkpoints. In general, these surveillance pathways are thought to act mainly to coordinate proficient DNA repair with cell cycle progression. To date, this cellular response to genotoxic stress has been viewed mainly as a DNA-based signal transduction pathway. Recent studies, in both yeast and human, however, highlight possible connections between chromatin structure and cell cycle checkpoints, in particular those involving kinases of the ATM and ATR family, known as key response factors activated early in the checkpoint pathway. In this review, based on this example, we will discuss hypotheses for chromatin-based events as potential initiators of a checkpoint response or conversely, for chromatin-associated factors as targets of checkpoint proteins, promoting changes in chromatin structure, in order to make a lesion more accessible and contribute to a more efficient repair response. PMID- 15279784 TI - Telomeres and the DNA damage response: why the fox is guarding the henhouse. AB - DNA double strand breaks (DSBs) are repaired by an extensive network of proteins that recognize damaged DNA and catalyze its repair. By virtue of their similarity, the normal ends of linear chromosomes and internal DNA DSBs are both potential substrates for DSB repair enzymes. Thus, telomeres, specialized nucleo protein complexes that cap chromosomal ends, serve a critical function to differentiate themselves from internal DNA strand breaks, and as a result prevent genomic instability that can result from their inappropriate involvement in repair reactions. Telomeres that become critically short due to failure of telomere maintenance mechanisms, or which become dysfunctional by loss of telomere binding proteins, elicit extensive checkpoint responses that in normal cells blocks proliferation. In this situation, the DNA DSB repair machinery plays a major role in responding to these "damaged" telomeres - creating chromosome fusions or capturing telomeres from other chromosomes in an effort to rid the cell of the perceived damage. However, a surprising aspect of telomere maintenance is that many of the same proteins that facilitate this repair of damaged telomeres are also necessary for their proper integrity. Here, we review recent work defining the roles for DSB repair machinery in telomere maintenance and in response to telomere dysfunction. PMID- 15279785 TI - The switch from survival responses to apoptosis after chromosomal breaks. AB - Eukaryotic cells have evolved highly sophisticated cellular responses to DNA double strand breaks (DSBs) that increase the likelihood of survival. However, cells can also respond to DSBs by undergoing programmed cell death. The mechanisms underlying the cellular decision on whether to repair and survive or to die are not well understood but may be related to the efficiency of repair or the extent of the damage. Presumably, a few easily reparable DSBs will not result in cell death in most cell types. However, abundant complex DSBs will present a severe challenge to the repair machineries with repeated attempts at repair likely to result in genome instability. For multicellular eukaryotes at least, struggling to complete repair is folly, whereas removal of severely damaged cells is a more sensible strategy. Here we discuss recent evidence linking DSBs to a highly regulated form of cell death termed, apoptosis. In particular, we focus on the roles of the tumour suppressor, p53 and a recently discovered role for an isotype of the linker histone H1. We present a hypothesis that the elevated levels of ssDNA produced during ongoing attempts at DSB repair may be involved in the switch from repair to apoptosis. PMID- 15279786 TI - Mammalian cell cycle checkpoints: signalling pathways and their organization in space and time. AB - The major mission of the cell division cycle is a faithful and complete duplication of the genome followed by an equal partitioning of chromosomes to subsequent cell generations. In this review, we discuss the advances in our understanding of how mammalian cells control the fidelity of these fundamental processes when exposed to diverse genotoxic insults. We focus on the most recent insights into the molecular pathways that link the sites of DNA lesions with the cell cycle machinery in specific phases of the cell cycle. We also highlight the potential of a new technology allowing direct visualization of molecular interactions and redistribution of checkpoint proteins in live cell nuclei, and document the emerging significance of live-cell imaging for elucidation of the spatio-temporal organization of the DNA damage response network. PMID- 15279787 TI - Dial 9-1-1 for DNA damage: the Rad9-Hus1-Rad1 (9-1-1) clamp complex. AB - Genotoxic stress activates checkpoint signaling pathways that block cell cycle progression, trigger apoptosis, and regulate DNA repair. Studies in yeast and humans have shown that Rad9, Hus1, Rad1, and Rad17 play key roles in checkpoint activation. Three of these proteins-Rad9, Hus1, and Rad1-interact in a heterotrimeric complex (dubbed the 9-1-1 complex), which resembles a PCNA-like sliding clamp, whereas Rad17 is part of a clamp-loading complex that is related to the PCNA clamp loader, replication factor-C (RFC). In response to genotoxic damage, the 9-1-1 complex is loaded around DNA by the Rad17-containing clamp loader. The DNA-bound 9-1-1 complex then facilitates ATR-mediated phosphorylation and activation of Chk1, a protein kinase that regulates S-phase progression, G2/M arrest, and replication fork stabilization. In addition to its role in checkpoint activation, accumulating evidence suggests that the 9-1-1 complex also participates in DNA repair. Taken together, these findings suggest that the 9-1-1 clamp is a multifunctional complex that is loaded onto DNA at sites of damage, where it coordinates checkpoint activation and DNA repair. PMID- 15279788 TI - Replication protein A phosphorylation and the cellular response to DNA damage. AB - Defects in cellular DNA metabolism have a direct role in many human disease processes. Impaired responses to DNA damage and basal DNA repair have been implicated as causal factors in diseases with DNA instability like cancer, Fragile X and Huntington's. Replication protein A (RPA) is essential for multiple processes in DNA metabolism including DNA replication, recombination and DNA repair pathways (including nucleotide excision, base excision and double-strand break repair). RPA is a single-stranded DNA-binding protein composed of subunits of 70-, 32- and 14-kDa. RPA binds ssDNA with high affinity and interacts specifically with multiple proteins. Cellular DNA damage causes the N-terminus of the 32-kDa subunit of human RPA to become hyper-phosphorylated. Current data indicates that hyper-phosphorylation causes a change in RPA conformation that down-regulates activity in DNA replication but does not affect DNA repair processes. This suggests that the role of RPA phosphorylation in the cellular response to DNA damage is to help regulate DNA metabolism and promote DNA repair. PMID- 15279789 TI - Chk1 in the DNA damage response: conserved roles from yeasts to mammals. AB - Chk1 is an evolutionarily conserved protein kinase that functions to ensure genomic integrity upon genotoxic stress. Studies to date have revealed striking similarities among Chk1 pathways of different organisms. In this review we discuss what is known about Chk1 activation and what downstream factors are regulated by Chk1 to counter replication blocks and DNA damage induced by UV, IR, and other genotoxic agents. Where applicable, we also compare the role of Chk1 with that of the Chk2 protein kinase in the checkpoint responses. PMID- 15279790 TI - Claspin, a regulator of Chk1 in DNA replication stress pathway. AB - Regulation of the vertebrate checkpoint kinase Chk1 involves several protein complexes including the recently identified protein Claspin. Claspin associates with Chk1 upon replication stress and DNA damage and is required for Chk1 activation in both Xenopus and human systems. More importantly, Claspin is involved in regulation of cell cycle checkpoints. Here, we discuss the emerging roles of Claspin in the Chk1 pathway and its functions in checkpoint control. PMID- 15279792 TI - The p53 response to DNA damage. AB - The p53 tumour suppressor protein is a highly potent transcription factor which, under normal circumstances, is maintained at low levels through the action of MDM2, an E3 ubiquitin ligase which directs p53 ubiquitylation and degradation. Expression of the mdm2 gene is stimulated by p53 and this reciprocal relationship forms the basis of a negative feedback loop. Both genotoxic and non-genotoxic stresses that induce p53 focus principally on interruption of the p53-MDM2 loop with the consequence that p53 becomes stabilised, leading to changes in the expression of p53-responsive genes. The biological outcome of inducing this pathway can be either growth arrest or apoptosis: factors affecting the functioning of the loop, the biochemical activity of p53 itself and the cellular environment govern the choice between these outcomes in a cell type- and stress specific manner. PMID- 15279791 TI - The Chk2 protein kinase. AB - Checkpoint kinase 2 (Chk2) is a multifunctional enzyme whose functions are central to the induction of cell cycle arrest and apoptosis by DNA damage. Insight into Chk2 has derived from multiple approaches. Biochemical studies have addressed Chk2 structure, domain organization and regulation by phosphorylation. Extensive work has been done to identify factors that recognize and respond to DNA damage in order to activate Chk2. In turn a number of substrates and targets of Chk2 have been identified that play roles in the checkpoint response. The roles and regulation of Chk2 have been elucidated by studies in model genetic systems extending from worms and flies to mice and humans. The relationship of Chk2 to human cancer studies is developing rapidly with increasing evidence that Chk2 plays a role in tumor suppression. PMID- 15279793 TI - Constitutively active DNA damage checkpoint pathways as the driving force for the high frequency of p53 mutations in human cancer. AB - If the major function of the p53 protein is to function as a DNA damage checkpoint protein, then it is reasonable to hypothesize that its inactivation in human cancer must be related to its DNA damage checkpoint function. This hypothesis further implies that in tumor cells one or more of the DNA damage checkpoint pathways has been activated. Otherwise, p53 would not be active and there would be no selective pressure for TP53 mutations. I make the argument that tumorigenic transformation is intrinsically associated with formation of DNA DSBs in every cell cycle leading to activation of DNA damage checkpoint pathways. In turn, activation of the DNA DSB checkpoint provides the selective pressure for the high frequency of p53 inactivation in human cancer. PMID- 15279794 TI - The interplay of Fanconi anemia proteins in the DNA damage response. AB - Fanconi anemia (FA) is a rare autosomal recessive disease characterized by chromosome instability and cancer predisposition. At least 11 complementation groups for FA have been identified, and eight FA genes have been cloned. Interestingly, the eight known FA proteins cooperate in a common pathway leading to the interaction of monoubiquitinated FANCD2 and BRCA2 in damaged chromatin. Disruption of this pathway results in the clinical and cellular abnormalities common to all FA subtypes. This review will examine the interaction of the cloned FA proteins with each other and with other DNA damage response proteins (i.e., ATM, ATR, and NBS1). Also, somatic (acquired) disruption of the FA pathway in human tumors appears to account for their chromosome instability and crosslinker hypersensitivity. PMID- 15279795 TI - The emerging role of E2F-1 in the DNA damage response and checkpoint control. AB - Genotoxic stress triggers a myriad of cellular responses including cell cycle arrest, stimulation of DNA repair and apoptosis. A central role for the E2F-1 transcription factor in the DNA damage response pathway is gaining support. E2F-1 is phosphorylated by DNA damage responsive protein kinases, which leads to E2F-1 accumulation and the induction of apoptosis. In addition, emerging information suggests that E2F-1 may play a role in the detection and subsequent repair of damaged DNA. PMID- 15279796 TI - The XRCC genes: expanding roles in DNA double-strand break repair. AB - Functional analysis of the XRCC genes continues to make an important contribution to the understanding of mammalian DNA double-strand break repair processes and mechanisms of genetic instability leading to cancer. New data implicate XRCC genes in long-standing questions, such as how homologous recombination (HR) intermediates are resolved and how DNA replication slows in the presence of damage (intra-S checkpoint). Examining the functions of XRCC genes involved in non-homologous end joining (NHEJ), paradoxical roles in repair fidelity and telomere maintenance have been found. Thus, XRCC5-7 (DNA-PK)-dependent NHEJ commonly occurs with fidelity, perhaps by aligning ends accurately in the absence of sequence microhomologies, but NHEJ-deficient mice show reduced frequencies of mutation. NHEJ activity seems to be involved in both mitigating and mediating telomere fusions; however, defective NHEJ can lead to telomere elongation, while loss of HR activity leads to telomere shortening. The correct functioning of XRCC genes involved in both HR and NHEJ is important for genetic stability, but loss of each pathway leads to different consequences, with defects in HR additionally leading to mitotic disruption and aneuploidy. Confirmation that these responses are likely to contribute to cancer induction and/or progression, is given by studies of humans and mice with XRCC gene disruptions: those affecting NHEJ show increased lymphoid tumours, while those affecting HR lead to breast cancer and perhaps to gynaecological tumours. PMID- 15279797 TI - Mismatch repair and DNA damage signalling. AB - Postreplicative mismatch repair (MMR) increases the fidelity of DNA replication by up to three orders of magnitude, through correcting DNA polymerase errors that escaped proofreading. MMR also controls homologous recombination (HR) by aborting strand exchange between divergent DNA sequences. In recent years, MMR has also been implicated in the response of mammalian cells to DNA damaging agents. Thus, MMR-deficient cells were shown to be around 100-fold more resistant to killing by methylating agents of the S(N)1type than cells with functional MMR. In the case of cisplatin, the sensitivity difference was lower, typically two- to three-fold, but was observed in all matched MMR-proficient and -deficient cell pairs. More controversial is the role of MMR in cellular response to other DNA damaging agents, such as ionizing radiation (IR), topoisomerase poisons, antimetabolites, UV radiation and DNA intercalators. The MMR-dependent DNA damage signalling pathways activated by the above agents are also ill-defined. To date, signalling cascades involving the Ataxia telangiectasia mutated (ATM), ATM- and Rad3-related (ATR), as well as the stress-activated kinases JNK/SAPK and p38alpha have been linked with methylating agent and 6-thioguanine (TG) treatments, while cisplatin damage was reported to activate the c-Abl and JNK/SAPK kinases in MMR-dependent manner. MMR defects are found in several different cancer types, both familiar and sporadic, and it is possible that the involvement of the MMR system in DNA damage signalling play an important role in transformation. The scope of this article is to provide a brief overview of the recent literature on this subject and to raise questions that could be addressed in future studies. PMID- 15279798 TI - PARP-1, PARP-2 and ATM in the DNA damage response: functional synergy in mouse development. AB - Poly(ADP-ribosyl)ation is an immediate DNA damage-dependent posttranslational modification of histones and nuclear proteins that contributes to the survival of injured proliferating cells. Poly(ADP-ribose) polymerases (PARPs) now constitute a superfamily of 18 proteins, encoded by different genes and displaying a common conserved catalytic domain. PARP-1 (113kDa), the founding member, and PARP-2 (62kDa) are both involved in DNA-break sensing and signaling when single strand break repair (SSBR) or base excision repair (BER) pathways are engaged. The generation by homologous recombination of deficient mouse models have confirmed the caretaker function of PARP-1 and PARP-2 in mammalian cells under genotoxic stress. This review summarizes our present knowledge on their physiological role in the cellular response to DNA damage and on the genetic interactions between PARP-1, PARP-2, Atm that play an essential role during early embryogenesis. PMID- 15279799 TI - DNA damage responses to oxidative stress. AB - The DNA damage response is a hierarchical process. DNA damage is detected by sensor proteins such as the MRN complex that transmit the information to transducer proteins such as ATM and ATR, which control the damage response through the phosphorylation of effector proteins. The extent of the DNA damage determines cell fate: cell cycle arrest and DNA repair or the activation of apoptotic pathways. In aerobic cells, reactive oxygen species (ROS) are generated as a by-product of normal mitochondrial activity. If not properly controlled, ROS can cause severe damage to cellular macromolecules, especially the DNA. We describe here some of the cellular responses to alterations in the cellular redox state during hypoxia or oxidative stress. Oxidative damage in DNA is repaired primarily via the base excision repair (BER) pathway which appears to be the simplest of the three excision repair pathways. To allow time for DNA repair, the cells activate their cell cycle checkpoints, leading to cell cycle arrest and preventing the replication of damage and defective DNA. PMID- 15279800 TI - The role of ATM and ATR in the cellular response to hypoxia and re-oxygenation. AB - ATM and ATR are stress-response kinases which respond to a variety of insults including ionizing radiation, replication arrest, ultraviolet radiation and hypoxia/re-oxygenation. Hypoxia occupies a unique niche in the study of both ATR- and ATM-mediated checkpoint pathways. Hypoxia is a physiologically significant stress that occurs in virtually all solid tumors and differs from most other stresses in that it does not induce DNA damage. Previous studies have indicated that hypoxia provides a unique way to induce ATR in response to inhibition of DNA replication. During tumor expansion hypoxia is inevitably followed by periods of re-oxygenation which in vitro has been shown to induce significant levels of DNA damage and an ATM response. Therefore both ATR and ATM have a role to play in hypoxia/re-oxygenation. PMID- 15279801 TI - Network responses to DNA damaging agents. AB - Global transcriptional profiling and large scale phenotypic studies have shown that eukaryotic cells mount a robust and complex response to damage. Further, systems biology approaches have employed powerful analytical methods to integrate global data sets with regulatory sequences, protein-DNA and protein-protein interactions, which have led to the identification of large networked responses to damage. A number of groups have shown that damage responsive networks embrace groups of highly connected genes and proteins and have illustrated that multiple interconnected cellular pathways respond to damage and moreover, dictate viability post damage. This review highlights some of the global studies that examine cellular responses to damage, and proposes that we should be moving from pathways to networks, in order to gain better insight into cellular defense mechanisms. PMID- 15279802 TI - Chromosome damage and progression into and through mitosis in vertebrates. AB - How cells behave as they divide in the presence of chromosome (DNA) damage is only just beginning to be explored. It appears to depend on the cell type and organism, the stage of development, how extensive the damage is and when it occurs. The existing data support the conclusion that vertebrate somatic cells lack a conventional DNA damage checkpoint during mitosis, and that when damaged DNA does prolong mitosis it is mediated by the spindle assembly checkpoint. As a rule, in the presence of DNA damage cells ultimately undergo an aberrant mitosis and enter the ensuing G1. They then either die, via apoptosis or mitotic catastrophe, or survive with an altered genome. To avoid these outcomes, cells with DNA damage are normally prevented from entering mitosis by a number of G2 checkpoint control pathways. PMID- 15279803 TI - The DNA double-strand break response in the nervous system. AB - DNA double-strand breaks (DSBs) require a coordinated molecular response to ensure cellular or organism survival. Many factors required for the DSB response, including those involved in non-homologous end joining (NHEJ) and homologous recombination repair (HRR) are essential during nervous system development. Additionally, human syndromes resulting from defective responses to DNA damage often feature overt neuropathology such as neurodegeneration. Thus, appropriate responses to DSBs are critical for the normal development and maintenance of the nervous system. PMID- 15279804 TI - The role of the DNA double-strand break response network in meiosis. AB - Organisms with sexual reproduction have two homologous copies of each chromosome. Meiosis is characterized by two successive cell divisions that result in four haploid sperms or eggs, each carrying a single copy of homologous chromosome. This process requires a coordinated reorganization of chromatin and a complex network of meiotic-specific signaling cascades. At the beginning of meiosis, each chromosome must recognize its homolog, then the two become intimately aligned along their entire lengths which allows the exchange of DNA strands between homologous sequences to generate genetic diversity. DNA double-strand breaks (DSBs) initiate meiotic recombination in a variety of organisms. Numerous studies have identified both the genomic loci of the initiating DSBs and the proteins involved in their formation. This review will summarize the activation and signaling networks required for the DSB response in meiosis. PMID- 15279805 TI - Interactions of viruses with the cellular DNA repair machinery. AB - Mammalian cells are equipped with complex machinery to monitor and repair damaged DNA. In addition to responding to breaks in cellular DNA, recent studies have revealed that the DNA repair machinery also recognizes viral genetic material. We review some examples that highlight the different strategies that viruses have developed to interact with the host DNA repair apparatus. While adenovirus (Ad) inactivates the host machinery to prevent signaling and concatemerization of the viral genome, other viruses may utilize DNA repair to their own advantage. Viral interactions with the repair machinery can also have detrimental consequences for the host cells and their ability to maintain the integrity of the host genome. Exploring the interactions between viruses and the host DNA repair machinery has revealed novel host responses to virus infections and has provided new tools to study the DNA damage response. PMID- 15279806 TI - Reverse genetic studies of the DNA damage response in the chicken B lymphocyte line DT40. AB - In the 'post-genome' era, reverse genetics is one of the most informative and powerful means to investigate protein function. The chicken B lymphocyte line DT40 is widely used for reverse genetics because the cells have a number of advantages, including efficient gene targeting as well as a remarkably stable phenotype. Furthermore, the absence of functional p53 in DT40 cells enables identification of DNA damage using chromosome analysis by suppressing damage induced apoptosis during interphase. This review summarizes the contribution of DT40 cells to reverse genetic studies of DNA damage response pathways in higher eukaryotic cells. PMID- 15279807 TI - Ataxia-telangiectasia, an evolving phenotype. AB - Ataxia-telangiectasia (A-T) is a progressive neurodegenerative disorder, with onset in early childhood and a frequency of approximately 1 in 40,000 births in the United States. A-T is seen among all races and is most prominent among ethnic groups with a high frequency of consanguinity. The syndrome includes: progressive cerebellar ataxia, dysarthric speech, oculomotor apraxia, choreoathetosis and, later, oculocutaneous telangiectasia. Immunodeficiency with sinopulmonary infections, cancer susceptibility (usually lymphoid), and sensitivity to ionizing radiation are also characteristic. Laboratory findings include: (1) elevated alphafetoprotein (AFP), (2) cerebellar atrophy on magnetic resonance imaging, (3) reciprocal translocations between chromosomes 7 and 14 in lymphocytes, (4) absence or dysfunction of the ATM protein, (5) radiosensitivity, as demonstrated by colony survival assay (CSA), and (6) mutations in the ATM gene. The latter are usually truncating or splicing mutations; approximately 10% are missense mutations. Mutations are found across the entire gene. Almost all recurring mutations are found on unique haplotypes that represent founder effects and ancestral relationships between patients. In addition to radiosensitivity and sensitivity to radiomimetic chemicals, the phenotype of A-T cells includes defective damage-induced activation of the cell cycle checkpoints at G1, S and G2/M. With the aid of molecular testing, A-T can now be distinguished from other autosomal recessive cerebellar ataxias (ARCAs) such as Friedreich ataxia, Mre11 deficiency (AT-like disease), and the oculomotor apraxias 1 (aprataxin deficiency) and 2 (senataxin deficiency). Other "A-T variants" include: (1) Nijmegen breakage syndrome (NBS) or nibrin/Nbs1 deficiency, with microcephaly and mental retardation but without ataxia, apraxia, or telangiectasia, and 2) A T(Fresno), a phenotype that combines features of both NBS and A-T, with mutations in the ATM gene. The term "A-T variant" has a diminishing usefulness. PMID- 15279808 TI - Functional consequences of sequence alterations in the ATM gene. AB - The product of the gene (ATM) mutated in the human genetic disorder ataxia telangiectasia (A-T) is a high molecular weight, protein ( approximately 350kDa) containing a C-terminal protein kinase domain and a number of other putative domains not yet functionally defined. The majority of ATM gene mutations in A-T patients are truncating, resulting in prematurely terminated products that are highly unstable. Missense mutations within the kinase domain and elsewhere in the molecule alter the stability of the protein and lead to loss of protein kinase activity. Only rarely are patients observed with two missense mutations and this gives rise to a milder disease phenotype. Evidence for a dominant interfering effect on normal ATM kinase activity has been reported in cell lines transfected with missense mutant ATM and in cell lines from some A-T heterozygotes. The dominant negative effect of mutant ATM is manifested by an enhancement of cellular radiosensitivity and may be responsible for the cancer predisposition observed in carriers of ATM missense mutations. In this review, we explore the domain structure of the ATM molecule, sites of interaction with other proteins and the consequences of specific amino acid changes on function. PMID- 15279809 TI - Nijmegen breakage syndrome: clinical manifestation of defective response to DNA double-strand breaks. AB - Nijmegen breakage syndrome is a rare autosomal recessive genetic disease belonging to a group of disorders often called chromosome instability syndromes. In addition to a characteristic facial appearance and microcephaly, patients suffering from Nijmegen breakage syndrome have a range of symptoms including radiosensitivity, immunodeficiency, increased cancer risk and growth retardation. The underlying gene, NBS1, is located on human chromosome 8q21 and codes for a protein product termed nibrin, Nbs1 or p95. Over 90% of patients are homozygous for a founder mutation: a deletion of five base pairs which leads to a framehift and protein truncation. The protein nibrin/Nbs1 is suspected to be involved in the cellular response to DNA damage caused by ionising irradiation, thus accounting for the radiosensitivity of Nijmegen breakage syndrome. We review here some of the more recent findings on the NBS1 gene and discuss how they impinge on the clinical manifestation of the disease. PMID- 15279810 TI - Ataxia-telangiectasia-like disorder (ATLD)-its clinical presentation and molecular basis. AB - Comparison of the clinical and cellular phenotypes of different genomic instability syndromes provides new insights into functional links in the complex network of the DNA damage response. A prominent example of this principle is provided by examination of three such disorders: ataxia-telangiectasia (A-T) caused by lack or inactivation of the ATM protein kinase, which mobilises the cellular response to double strand breaks in the DNA; ataxia-telangiectasia-like disease (ATLD), a result of deficiency of the human Mre11 protein; and the Nijmegen breakage syndrome (NBS), which represents defective Nbs1 protein. Mre11 and Nbs1 are members of the Mre11/Rad50/Nbs1 (MRN) protein complex. MRN and its individual components are involved in different responses to cellular damage induced by ionising radiation and radiomimetic chemicals, including complexing with chromatin and with other damage response proteins, formation of radiation induced foci, and the induction of different cell cycle checkpoints. The phosphorylation of Nbs1 by ATM would indicate that ATM acts upstream of the MRN complex. Consistent with this were the suggestions that ATM could be activated in the absence of fully functional Nbs1 protein. In contrast, the regulation of some ATM target proteins, e.g. Smc1 requires the MRN complex as well as ATM. Nbs1 may, therefore, be both a substrate for ATM and a mediator of ATM function. Recent studies that indicate a requirement of the MRN complex for proper ATM activation suggest that the relationship between ATM and the MRN complex in the DNA damage response is yet to be fully determined. Despite the fact that both Mre11 and Nbs1 are part of the same MRN complex, deficiency in either protein in humans does not lead to the same clinical picture. This suggests that components of the complex may also act separately. PMID- 15279811 TI - An overview of three new disorders associated with genetic instability: LIG4 syndrome, RS-SCID and ATR-Seckel syndrome. AB - Around 15-20 hereditary disorders associated with impaired DNA damage response mechanisms have been previously described. The range of clinical features associated with these disorders attests to the significant role that these pathways play during development. Recently, three new such disorders have been reported extending the importance of the damage response pathways to human health. LIG4 syndrome is conferred by hypomorphic mutations in DNA ligase IV, an essential component of DNA non-homologous end-joining (NHEJ), and is associated with pancytopaenia, developmental and growth delay and dysmorphic facial features. Radiosensitive severe combined immunodeficiency (RS-SCID) is caused by mutations in Artemis, a protein that plays a subsidiary role in non-homologous end-joining although it is not an essential component. RS-SCID is characterised by severe combined immunodeficiency but patients have no overt developmental abnormalities. ATR-Seckel syndrome is caused by mutations in ataxia telangiectasia and Rad3 related protein (ATR), a component of a DNA damage signalling pathway. ATR-Seckel syndrome patients have dramatic microcephaly and marked growth and developmental delay. The clinical features of these patients are considered in the light of the function of the defective protein. PMID- 15279812 TI - Genetic biomarkers of therapeutic radiation sensitivity. AB - The occurrence of acute or late normal tissue reactions after therapeutic radiotherapy and cellular responses in in vitro radiosensitivity assays do not correlate well suggesting that to date no one test system is suitable for predicting the risk or severity of such reactions. New insights into the underlying molecular mechanisms of this sensitivity are coming from studies that assess associations between common polymorphisms in DNA damage detection and repair genes and the development of adverse reactions to radiotherapy. The presence of such variants may alter protein function and an individual's capacity to repair damaged DNA modifying the response of the normal tissue. Polymorphisms in the XRCC1, ATM, hHR21 and TGFbeta1 genes have been shown to be associated with an increased risk of developing an adverse normal tissue reaction to radiotherapy, whilst one variant in the ATM gene has been reported to be radioprotective. Functional studies, taking into account either the haplotypes or the combined genotypes when multiple polymorphisms in a gene are present, will be necessary to establish the mechanistic basis of these associations. In the future association studies can only benefit from the analysis of multiple genes in large, well-characterized cohorts in particular to identify genetic factors that might specifically influence the temporal occurrence of these adverse reactions. PMID- 15279813 TI - Relevance and irrelevance of DNA damage response to radiotherapy. AB - Ionizing radiation (IR) has been used to treat human malignancies since the early part of the 20th century. To date, most of the advances in radiotherapy have focused on optimization of treatment delivery schedules and technologic improvements in the physical targeting of dose. By comparison, many of the discoveries regarding the molecular basis of DNA damage and repair have not yet been translated to clinical practice. This article offers some perspectives regarding modulators of radiation effects and the challenges faced as we approach newer molecular targets. Our goal is to frame the issues that contribute to the apparent disconnect between laboratory discoveries and improvements in clinically relevant therapeutics. PMID- 15279814 TI - Why is it important to diagnose chorionicity and how do we do it? AB - Because the monochorionic (MC) placenta is designed for a singleton fetus, and might not provide adequate physiological support for twins, obstetric problems are more frequent in MC than dichorionic (DC) twins. Problems arise because asymmetric cord insertions cause growth discordance as a result of unequal sharing of placental tissue. Approximately 95% of MC twin placentas contain interfetal vascular connections of some kind, sometimes in several combinations. Such connections can cause twin-twin transfusion syndrome and twin reversed arterial perfusion. The survivor can also suffer damage if the co-twin dies spontaneously or from inappropriate methods of selective termination. These complications are progressive and often advanced by 18 weeks gestation. Monoamniotic twins carry greater risks than diamniotic twins, especially entangled cords. MC twins are often discordant for congenital anomalies. Diagnosis of MC twinning is optimal in the first trimester. Optimal management of these MC twin disorders is not yet established; long-term follow-up studies are unsatisfactory. In clinical practice, chorionicity is not always determined in the first trimester. PMID- 15279815 TI - What prenatal diagnosis should be offered in multiple pregnancy? AB - The issues surrounding prenatal diagnosis in multiple pregnancy are complex. Accurate determination of chorionicity is vital and an inability to determine this should trigger consideration for referral to a specialist. The choice of screening method for detection of chromosomal abnormality is limited, and existing data demonstrates the advantages of nuchal translucency screening. The possibility of obtaining discordant results and options for management should be discussed in advance. Invasive tests are technically more difficult and associated with a higher risk of procedure-related pregnancy loss than less invasive methods. Repeat invasive testing is required more often in multiple pregnancies than in singleton pregnancies. Selective termination is technically feasible in both mono- and dichorionic pregnancies, although the risks are higher with the former. It is likely to be more acceptable than high-order multifetal reduction performed in the absence of fetal abnormality. PMID- 15279816 TI - How can we diagnose and manage twin-twin transfusion syndrome? AB - Severe mid-trimester twin-twin transfusion syndrome (TTS) complicates about 15% of monochorionic twin pregnancies. If left untreated, the mortality is 80-100%. The pathophysiological prerequisite for the onset of TTS is unequal blood flow via arteriovenous placental anastomoses from the so-called donor to the recipient twin. This can result in hypovolemia, hypotension and oligo- or anuria in the donor, and hypervolemia, hypertension, polyuria and finally heart failure in the recipient. Leading sonographic signs of TTS include severe oligo- or anhydramnios and a small or absent bladder filling in the donor in contrast to polyhydramnios with increased bladder filling in the recipient. Patients might present with clinical symptoms due to massive polyhydramnios. In severe mid-trimester TTS, fetoscopic laser occlusion of the anastomosing vessels on the placental surface under local anaesthesia plus subsequent amniodrainage is the most promising therapeutic option at present. In acute TTS after 26 weeks of gestation, amniodrainage is the therapy of choice. All patients suspected of this high-risk condition should be referred to a specialized fetal medicine centre. PMID- 15279817 TI - Obstetric complications of twin pregnancies. AB - Advances in assisted reproductive technology and increases in the proportion of maternities in older women have both contributed to the steep increase in the incidence of twin pregnancies since the 1980s. Maternal and perinatal complications are higher in twins than in singleton pregnancies. A significant proportion of perinatal mortality and morbidity among twins is due to the high incidence of preterm delivery and the added complication of twin-to-twin transfusion syndrome (TTTS) in monochorionic twins. Monochorionic twins also have a much higher rate of perinatal mortality than dichorionic twins, the greatest risk being before fetal viability (<24 weeks gestation). Early diagnosis of twins and their chorionicity, close fetal surveillance, particularly of monochorionic twins, and prompt therapeutic intervention in TTS are necessary to reduce perinatal mortality. Intrapartum management in the hospital setting with anaesthetic and neonatal facilities, as well as critical assessment of mode of delivery, have led to better outcomes. Ultrasonography is a valuable tool in the management of twin pregnancy. This chapter briefly summarises these topics, with a particular focus on recent literature. PMID- 15279818 TI - Management of other complications specific to monochorionic twin pregnancies. AB - Monochorionic (MC) twins have a 3-10-fold higher perinatal mortality and morbidity than dichorionic twins. This is largely attributable to their common vascular architecture and the high rate of discordant fetal growth, growth restriction and congenital abnormalities. In the event of a single intrauterine death (IUD), intertwin agonal transfusion results in up to a 38% risk of death and a 46% risk of neurological injury to the co-twin. This chapter addresses the management of complications unique to MC twins. The primary aim of management is to prevent single IUD or, if inevitable, prevent agonal transfusion occurring by vascular occlusive selective feticide. Older fetoscopic techniques have been replaced by the simpler ultrasound-guided techniques of interstitial laser and bipolar cord occlusion. Their application in twin reversed-arterial perfusion sequence has been associated with a 50% reduction of perinatal mortality in the pump twin. Moreover, prophylactic interstitial laser therapy in early pregnancy might obviate the technical and clinical difficulties in the presence of fetal decompensation in later pregnancy. Recent strategies to reduce the high perinatal mortality due to cord entanglement in antenatally diagnosed monoamniotic twins including medical amnioreduction and elective caesarean delivery at 32 weeks, are also discussed. PMID- 15279819 TI - Do reduced multiples do better? AB - Dramatic successes in infertility care have allowed millions of previously fertile women to have their own children. However, an epidemic of multiple pregnancies has resulted, with catastrophic increases in morbidity and mortality, and in the economic costs to society. Multifetal pregnancy reduction (MFPR) has been used to decrease fetal number in the late first trimester and has dramatically improved outcomes. Recent data suggest that pregnancies starting with three or four, and in some cases five fetuses, which are reduced to twins, do as well as starting with twins. Patients with triplets do better reduced to twins. Reduction to a singleton is becoming more common, particularly for women over 40. Combining MFPR with chorionic villus sampling in patients over 30 years of age has enabled couples to maximize the health of the resultant children. PMID- 15279820 TI - Is it normal for multiples to be smaller than singletons? AB - This section discusses fetal growth in multiple pregnancy from various perspectives. Whereas the entire 'fetal mass' of a multiple pregnancy can exceed the 90th birth weight percentile of a singleton of the same gestational age, the individual fetuses might exhibit growth patterns compatible with adaptation to the limited uterine environment. Adaptation can take the form of relative growth restriction (i.e. discordance), whereby not all fetuses show a decelerating growth pattern. When adaptation fails, fetal growth is genuinely restricted. However, even in cases with significant discordance, about 40% of the smaller twins weigh above the 10th birth weight percentile. The key in these circumstances might be found in the ability to recognize the transition from the physiologic adaptation into the pathologic process of growth restriction. PMID- 15279821 TI - Delivery of the term twin. AB - The ever-increasing incidence of twin pregnancies world wide, together with the increasing trend to caesarean delivery, has resulted in intense scrutiny of the most appropriate method of twin delivery. The term twin has an increased risk of twin mortality compared to term singletons and this might be a result of the increase risk of labour and delivery compared to that of singletons. There are three ways to address this from the literature. The first is to compare outcome for the second twin versus the first twin, and to compare these outcomes in those twins delivered vaginally compared to those delivered by lower section caesarean section (LSCS). The second is to compare outcomes for twins delivered vaginally and for those delivered by caesarean section (CS). These data show higher rates of adverse perinatal outcome for the twin at or near term if delivery is vaginal versus CS. The third method is to compare outcomes for twins delivered by planned vaginal birth (VB; actual VB plus emergency CS) versus planned CS. This chapter will review this data thus outline an ongoing randomized controlled trial--the Twin Birth Study. PMID- 15279822 TI - How and why are triplets disadvantaged compared to twins? AB - The current epidemic of triplets, a result of the widespread use of assisted reproduction, started less than two decades ago. Its full impact has been appreciated only recently. Triplets are disadvantaged from every perinatal perspective compared to twins--preterm birth, low birth weight, morbidity and mortality--because the human uterus probably is better equipped to carry twins than triplets. Although modern neonatal care has improved survival rates of preterm as well as low-birth-weight triplets, other complications remain and are of great clinical importance. The alternative to carrying triplets--multifetal pregnancy reduction--is associated with improved outcomes, as expected from comparing twin to higher-order multiples. However, the improved outcomes of triplets in recent years might call for second thoughts about the frequent recommendation of multifetal pregnancy reduction of triplets to twins. PMID- 15279823 TI - Psychological adjustment to twins after infertility. AB - The birth of twins and other multiples is physically and emotionally stressful. The increase in the use of the assisted reproductive technologies has lead to an exponential increase in the rates of twins and triplets in the US. Whereas the medical complications of twins and other multiples has been well studied, the psychological and social implications of these events has not. Very little empirical research has been conducted to assess the differential impact of twins, as compared to singletons, on maternal adjustment, postpartum depression and marital functioning. In addition, assessment of infant health, disposition and behavior and its relation to maternal adjustment is lacking. The birth of twins after a period of infertility complicates the clinical picture and the impact of infertility on subsequent parental adjustment is only beginning to be understood. Although research suggests that infertile couples often desire multiples, the experience of parenting multiples after infertility has not been studied. Research on fertile couples indicate that: (i) approximately 10% of women develop postpartum depression and; (ii) marital adjustment declines after the birth of the first child. Because of the unique demands of parenting multiples, it is hypothesized that mothers of twins who have a history of infertility would be at increased risk for depression and marital decline. Descriptive studies of these families support this view, although additional studies are needed to determine the degree and extent of the problem. Additionally, variables such as, prepregnancy adjustment, equitable division of child-care tasks and perceived social support should be studied to determine if they buffer against the expected effects. PMID- 15279824 TI - Economic and social implications of multiple birth. AB - This paper provides the background statistics and trends on multiple births. It highlights a number of factors that influence parents with multiples, including social situation, psychological adjustment and economic circumstances. It is often assumed that these areas of concern arise only after the babies have been born, but clinicians should be aware that many are seen in the antenatal period as well. Apart from the socioeconomic cost to individual families, multiple births also carry a cost to society. A number of self-help groups and voluntary organisations can assist parents of multiples. This is especially important if parents find that family and friends cannot or will not come forward. PMID- 15279825 TI - Cytogenetic monitoring of personnel working in angiocardiography laboratories in Iran hospitals. AB - Angiocardiography is an X-ray examination of the blood vessels or chambers of the heart for evaluation of the number and severity of blockages in arteries that supply blood to the heart. Cardiologists and staff members applying these procedures are exposed to high levels of scattered radiation. In the present study we analyzed and followed-up on the cytogenetic effects of X-ray angiography in personnel of laboratories for treatment of cardiovascular disease. According to film dosimeter analysis, personnel received 0.25-15 mSv during the previous year (average of 3 mSv/y), which indicated an exposure below the limit established by the International Commission on Radiological Protection (ICRP). Samples of peripheral blood were collected from cardiologists, nurses and technicians and from a matched control group. The incidence of unstable chromosomal aberrations (dicentrics, acentric fragments, ring chromosomes) and cytokinesis-blocked micronuclei were analyzed. Results show a high frequency of acentric fragments in cardiologists, nurses and technicians compared to controls (P < 0.001). When the exposed groups were compared, a higher percentage of acentric fragments was observed in the nurses and technicians compared to cardiologists (P = 0.004). Six individuals presented with dicentric chromosomes and their equivalent whole-body doses ranged from 0.05 to 0.10 Gy. No correlation was observed between chromosome aberrations and annual effective dose or age of the exposed groups. Although the mean frequency of chromosome aberrations in the male workers was slightly higher than in the females, no significant difference was observed between male and female workers in each group (P = 0.86). The mean number of micronuclei per 1000 binucleated cells was significantly higher in the exposed groups compared with the matched control group (P < 0.001). In a follow up study, chromosome aberrations in lymphocytes of 23 exposed personnel were analyzed five times during 3 years. Results show that the mean number of acentric fragments decreased gradually during 36 months in those workers who followed the radiation protection guides. The results of this study emphasize the importance of individual biomonitoring, limiting exposure and radiation safety programs. PMID- 15279826 TI - Facilitated detection of chromosome break and repair at low levels of ionizing radiation by addition of wortmannin to G1-type PCC fusion incubation. AB - We have measured rejoining kinetics of chromosome breaks using a modified cell fusion-based premature chromosome condensation (PCC) technique in confluent cultures of normal human fibroblasts irradiated at low doses of X-rays. In order to enhance the sensitivity of the fusion-based PCC assay, we added a DNA double strand break (DSB) repair inhibitor wortmannin during the incubation period for PCC/fusion process resulting in a significantly higher yield of G1-type chromosome breaks. The initial number of chromosome breaks (without repair) gave a linear dose response with about 10 breaks per cell per Gy which is about two times higher than the value with the conventional G1-type PCC method. The chromosome rejoining kinetics at 0.5 and 2.0 Gy X-rays reveal a bi-phasic curve with both a fast and a slow component. The fast component (0-30 min) is nearly identical for both doses, but the slow component for 2 Gy kinetics is much slower than that for 0.5 Gy, indicating that the process occurring during this period may be crucial for the ultimate fate of irradiated cells. The chromosome rejoining kinetics obtained here is similar to that of other methods of detecting DNA DSB repair such as the gammaH2AX assay. Our chromosome repair assay is useful for evaluating the accuracy of other assays measuring DNA DSB repair at doses equal or less than 0.5 Gy of ionizing radiation. PMID- 15279827 TI - Identification of cytochrome P-450s involved in the formation of APNH from norharman with aniline. AB - Mutagenic 9-(4'-aminophenyl)-9H-pyrido[3,4-b]indole (aminophenylnorharman, APNH), formed from norharman and aniline in the presence of S9 mix, is thought to be accountable for the co-mutagenic action of norharman. Our previous studies suggest that cytochrome P-450s (CYPs) are involved in the generation of APNH. In order to identify the responsible CYP species in the present study, norharman (8 mg) and aniline (4 mg) were incubated with individual recombinant human CYPs (2 nmol) at 37 degrees C for 20 min. Formation of APNH was observed with CYP1A1, CYP1A2, CYP1B1, CYP2B6, CYP2D6, CYP2E1 and CYP3A4, but not with CYP2A6, CYP2C9 and CYP2C19. The amounts of APNH from norharman and aniline were 33 ng for CYP1A1, 15 ng for CYP3A4, 7 ng for CYP2D6, 6 ng for CYP1A2 and 5 ng for CYP2B6. APNH formation in the presence of CYP1B1 and CYP2E1 was very low at around one fiftieth of that with CYP3A4. When CYP selective chemical inhibitors, such as furafylline for CYP1A2 and ketoconazole for CYP3A4, were added to the reaction mixture of norharman, aniline and human microsomes, formation of APNH was decreased to 14 and 16% of the control level, respectively. Moreover, human lung microsomes also showed the activity of APNH formation from norharman and aniline, albeit at only one hundredth of that with liver microsomes. In general, content in human liver microsomes is rather high for CYP3A4 and CYP1A2 but relatively low for CYP2D6 and CYP2B6, at about 30, 10, 1.5% and less than 1% of the total CYP, respectively. Although CYP1A1 showed the highest APNH formation activity, its expression in human liver is reported to be below the level of detection. Based on these observations, it is suggested that the practical major contributors to the formation of APNH from norharman and aniline are CYP3A4 and CYP1A2, the responsible reactions mainly occurring in the liver. PMID- 15279828 TI - Clastogenic effect for cigarette smoking but not areca quid chewing as measured by micronuclei in exfoliated buccal mucosal cells. AB - The objective of this study was to use the micronuclei from exfoliated buccal mucosal cells to investigate the clastogenic effects of areca quid chewing and cigarette smoking, as well as the interaction between the two. The study population was selected from residents of seven villages recruited for a community-cohort study. A total of 141 subjects were recruited based on the regular consumption of cigarettes and betel quid. Salient personal characteristics were collected from interview using a specially designed questionnaire. Micronuclei were scored on Feulgen/fast green-stained smear preparations of exfoliated cells obtained by scraping the surface of the buccal mucosa. There was no significant interaction between the chewing of betel nut and cigarette smoking. Heavy smoking was positively associated with MN frequency, with areca quid chewing negatively associated. A significant positive trend was demonstrated for the relationship between MN frequency and either daily cigarette consumption or cumulative smoking pack-years. By contrast, negative trends were demonstrated for the analogous relationships with areca quid chewing. These results indicate that heavy smoking, but not areca quid chewing, increases MN formation. These findings suggest that the carcinogenesis of the oral cancers induced by areca quid chewing in Taiwan may be through a pathway other than genotoxicity. PMID- 15279829 TI - Mutagenicity in Salmonella of halonitromethanes: a recently recognized class of disinfection by-products in drinking water. AB - Halonitromethanes (HNMs) are a recently identified class of disinfection by products (DBPs) in drinking water. They include chloronitromethane (CHN), dichloronitromethane (DCNM), trichloronitromethane (TCNM), bromonitromethane (BNM), dibromonitromethane (DBNM), tribromonitromethane (TBNM), bromochloronitromethane (BCNM),dibromochloronitromethane (DBCNM), and bromodichloronitromethane (BDCNM). Previous studies of TCNM, DCNM, CNM, and TBNM found that all four were mutagenic in bacteria, and a recent study showed that all nine induced DNA damage in CHO cells. Here, all nine HNMs were evaluated in the Salmonella plate-incorporation assay +/- S9 using strains TA98, TA100, TA104, TPT100, and the glutathione transferase theta (GSTT1-1)-expressing strain RSJ100. All were mutagenic, most with and without S9. In the absence of S9, six were mutagenic in TA98, six in TA100, and three in TA104; in the presence of S9, these numbers were five, seven, and three, respectively. Thus, the HNMs-induced base substitutions primarily at GC sites as well as frameshifts. Although five HNMs were activated to mutagens in RSJ100 -S9, they produced < or =2-fold increases in revertants and potencies <506 rev/micromol. The rank order of the HNMs by mutagenic potency in TA100 +S9 was (BCNM DBNM) > (TBNM CNM > BNM DCNM BDCNM) > (TCNM = DBCNM). The mean rev/micromol for the three groupings, respectively, were 1423, 498, and 0, which classifies the HNMs as weak mutagens in Salmonella. Reaction of the dihalo and monohalo HNMs with GSH, possibly GSTT1-1, is a possible mechanism for formation of ultimate mutagenic products. Because the HNMs are mutagenic in Salmonella (present study) and potent clastogens in mammalian cells [Environ. Sci. Technol. 38 (2004) 62], their presence in drinking water warrants further research on their potential health effects. PMID- 15279830 TI - Chromosome aberrations among cigarette smokers in Colombia. AB - Worldwide, the annual morbimortality caused by cigarette smoking is a major public health concern. In Colombia, up to 33% of the adult population has smoked at some point in life, raising important national issues on the disease burden from tobacco. The aim of this study was to establish whether cigarette smoking increases the frequency of chromosome aberrations (CA) in peripheral blood lymphocytes of smokers (n = 52) compared with non-smokers (n = 52) in Popayan, Colombia. After signing a consent form, volunteers provided a blood sample (20 ml) to establish cell cultures at 52 h. For CA analysis, 100 complete metaphase cells from each subject were evaluated. The CA frequency was significantly higher in smokers (8.38 +/- 0.61) than in non-smokers (3.13 +/- 0.29), showing the highest number of CA (14.83 +/- 1.01) among heavy smokers (>20 pack-years). Interestingly, light smokers (< or =10 pack-years) also showed a significant increase in CA when compared to non-smokers (6.62 +/- 0.53 versus 3.13 +/- 0.29, P < 0.01, respectively). In addition, a significant positive correlation was found between the frequency of CA and the intensity of smoking in pack-years (R2 = 0.60). Our study indicates that the genotoxic effects in lymphocytes from smokers are most likely caused by cigarette smoke constituents, providing scientific evidence to encourage national campaigns to prevent tobacco consumption. PMID- 15279831 TI - Chromosome and DNA damage analysis in individuals occupationally exposed to pesticides with relation to genetic polymorphism for CYP 1A1 gene in Ecuador. AB - DNA damage was measured by using the alkaline comet assay and the chromosomal aberration (CA) test using peripheral blood samples from 45 pesticide sprayers from Cayambe, Ecuador. From a total of approximately 200 nuclei scored for each donor with the comet assay, a highly significant increase in DNA migration was observed when compared with a similar unexposed control population. Additionally, in the CA test, the exposed individuals were found to be significantly different when compared to the control population. Polymorphisms for the CYP 1A1 (Msp I and Ile/Val) in exposed individuals were analyzed by PCR-RFLP and allele-specific PCR techniques. No association was found between the polymorphisms and higher levels of DNA damage as assessed by the comet assay. PMID- 15279832 TI - Genotoxicity of goniothalamin in CHO cell line. AB - Goniothalamin (GTN) is a styrylpyrrone derivative from Goniothalamus umbrosus and other Annonaceae species. It has been shown to have anti-cancer and apoptosis inducing properties against various human tumour and animal cell lines. The compound has also been shown to be active in vivo against DMBA-induced rat mammary tumours and was reported as an anti-fertility agent in rats. The aim of our study was to assess the genotoxicity of GTN in CHO cells using the UKEMS guidelines. A metabolic activation fraction (S9) was prepared according to standard methods. The methylthiazoletetrazolium (MTT) screening assay was then carried out to determine the cytotoxicity index (IC50) of GTN. The average IC50 value was 12.45 (+/- 3.63)microM. The mitotic index (MI) assay was then performed to determine the clastogenicity indices (MI(C25), MI(C50) and MI(C100)) of GTN. The chromosome aberration (CA) induction assay using air-dried metaphase spread was then performed to investigate the clastogenic effects of goniothalamin. Benzo[a]pyrene (BaP) and ethylmethanesulphonate (EMS) were used as positive controls in the presence and absence of S9 metabolic activation, respectively. The anti-genotoxicity effect of GTN was also assessed using a combination of GTN and EMS, and GTN and BaP. Dose-responses of CA frequencies were determined for both, the genotoxicity and anti-genotoxicity effects. GTN on its own and when combined with positive controls, was found to induce and enhance CA, respectively. Chromatid and whole chromosome breaks/gaps, as well as interchanges, endoreduplications and ring chromosomes were the main types of aberration induced by GTN. The overall clastogenic effect of GTN was statistically significant. In conclusion, GTN is potentially a genotoxic or clastogenic substance without any anti-genotoxic properties. PMID- 15279833 TI - Non-smoking coke oven workers show an occupational PAH exposure-related increase in urinary mutagens. AB - We examined the urinary mutagenicity in the YG1024 Salmonella typhimurium strain in the presence of S9 mix, of 31 male non-smoking coke oven workers and an equal number of controls matched for gender and dietary habits. Occupational PAH exposure to the workers was assessed by means of the individual urinary post shift excretion of 1-pyrenol (mean +/- S.D.: 5.41 +/- 6.06 micromole/mol creatinine). Eleven urinary extracts of workers (35.5%) were clearly mutagenic (with at least a doubling of the number of spontaneous revertants), against only two samples in the control group (6.5%) (chi2-test; chi2 = 7.883; P < 0.01). Moreover, the mean mutagenic activity level corrected for dilution/concentration of the urine was about three times higher in coke oven workers than in matched controls (mean +/- S.D. (range) 495 +/- 407 (89.7-1603) versus 186 +/- 113 (14.2 524) net revertants/mmol creatinine; Mann-Whitney U-test, z = 3.86, P < 0.001). Simple linear regression analysis showed that the coke workers' urinary mutagenic activity is associated with the PAH occupation-related urinary excretion of 1 pyrenol (r = 0.41, P = 0.0215). This study definitely demonstrates an occupation related exposure of coke oven workers' bladder epithelium to mutagenic PAH metabolites. This factor, mainly in the case of high exposure studied here, may account for a higher bladder cancer risk in coke oven workers. PMID- 15279834 TI - Genetic damage in exfoliated cells from oral mucosa of individuals exposed to X rays during panoramic dental radiographies. AB - The genotoxic effects of X-ray emitted during dental panoramic radiography were evaluated in exfoliated cells from oral epithelium through a differentiated protocol of the micronucleus test. Thirty-one healthy individuals agreed to participate in this study and were submitted to this procedure for diagnosis purpose after being requested by the dentist. All of them answered a questionnaire before the examination. Cells were obtained from both sides of the cheek by gentle scrapping with a cervical brush, immediately before the exposure and after 10 days. Cytological preparations were stained according to Feulgen Rossenbeck reaction and analyzed under light and laser scanning confocal microscopies. Micronuclei, nuclear projections (buds and broken eggs) and degenerative nuclear alterations (condensed chromatin, karyolysis and karyorrhexis) were scored. The frequencies of micronuclei, karyolysis and pycnosis were similar before and after exposure (P > 0.90), whereas the condensation of the chromatin and the karyorrhexis increased significantly after exposure (P < 0.0001). In contrast, both bud and broken egg frequencies were significantly higher before the examination (P < 0.005), suggesting that these structures are associated to the normal epithelium differentiation. The results suggest that the X-ray exposure during panoramic dental radiography induces a cytotoxic effect by increasing apoptosis. We also believe that the score of other nuclear alterations in addition to the micronucleus improves the sensitivity of genotoxic effects detection. PMID- 15279835 TI - Inflammatory and genotoxic effects of diesel particles in vitro and in vivo. AB - Recent studies have identified an indirect genotoxicity pathway involving inflammation as one of the mechanisms underlying the carcinogenic effects of air pollution/diesel exhaust particles (DEP). We investigated the short-term effects of DEP on markers of inflammation and genotoxicity in vitro and in vivo. DEP induced an increase in the mRNA level of pro-inflammatory cytokines and a higher level of DNA strand breaks in the human lung epithelial cell line A549 in vitro. For the in vivo study, mice were exposed by inhalation to 20 or 80 mg/m3 DEP either as a single 90-min exposure or as four repeated 90-min exposures (5 or 20 mg/m3) and the effects in broncho-alveolar lavage (BAL) cells and/or lung tissue were characterized. Inhalation of DEP induced a dose-dependent inflammatory response with infiltration of macrophages and neutrophils and elevated gene expression of IL-6 in the lungs of mice. The inflammatory response was accompanied by DNA strand breaks in BAL cells and oxidative DNA damage and increased levels of bulky DNA adducts in lung tissue, the latter indicative of direct genotoxicity. The effect of a large single dose of DEP was more pronounced and sustained on IL-6 expression and oxidative DNA damage in the lung tissue than the effect of the same dose administered over four days, whereas the reverse pattern was seen in BAL cells. Our results suggest that the effects of DEP depend on the rate of delivery of the particle dose. The mutation frequency (MF), after DEP exposure, was determined using the transgenic Muta Mouse and a similar exposure regimen. No increase was observed in MF in lung tissue 28-days after exposure. In conclusion, short-term exposure to DEP resulted in DNA strand breaks in BAL cells, oxidative DNA damage and DNA adducts in lungs; and suggested that DNA damage in part is a consequence of inflammatory processes. The response was not associated with increased MF, indicating that the host defence mechanisms were sufficient to counteract the adverse effects of inflammation. Thus, there may be thresholds for the inflammation-associated genotoxic effects of DEP inhalation. PMID- 15279836 TI - Inter-rater agreement on chromosome 5 breakage in FISH-based mutagen sensitivity assays (MSAs). AB - In chromosome breakage assays, validated, universal criteria for selection of cells and classification of chromosome aberrations may enhance their utility for cancer susceptibility screening. To standardize a fluorescence in situ hybridization (FISH) modification of the mutagen sensitivity assay (MSA), scoring criteria were evaluated by web-based validation. Two hundred digital FISH images were assigned random identification numbers. With this set of images, criteria for inclusion of cells and measurement of the frequency of abnormal cells were evaluated by eight observers, all of whom had five or more years of experience. Observers included doctoral and MS/BS level cytogeneticists, and were drawn from a randomized pool of 54 volunteers. Questions addressed were: (1) how uniformly were criteria applied to analysis of a standard digital FISH image set and (2) did concordance vary with educational level? These data suggest inter-rater agreement within a factor of 2 for average breakage frequency, but revealed greater variability in cell selection. These results aid in estimating the components of assay variance due to definitions, technical parameters and biological variables. PMID- 15279837 TI - Phototoxicity and DNA damage induced by the cosmetic ingredient chemical azulene in human Jurkat T-cells. AB - Previous study showed that the cosmetic ingredient chemical azulene and its derivative gauiazulene exhibited photomutagenicity four- to five-fold higher than spontaneous mutation in Salmonella typhimurium TA102. In this study, phototoxicity including photogenotoxicity of azulene in human Jurkat T-cells is reported. When the cell suspensions are irradiated by light (UVA plus visible light) in the presence of azulene, an azulene dose-dependent cellular DNA damage is observed. At the highest azulene concentration of 50 microM, the average DNA fragmentation is 33 +/- 10%, determined by single cell gel electrophoresis (Comet assay). Cell viability assay using fluorescein diacetate indicates that the cells could endure the damage and remain viable. Further study revealed that the combination of light and azulene can cause single-strand cleavage on pure PhiX174 plasmid DNA in solution. Studies using scavengers reveal that singlet oxygen and free radicals are involved in causing DNA cleavage. This suggests that the photomutagenicity of azulene in S. typhimurium TA102 could be due to DNA fragmentation caused by the concurrent exposure to azulene and light. PMID- 15279838 TI - Establishment of ten strains of genetically engineered Salmonella typhimurium TA1538 each co-expressing a form of human cytochrome P450 with NADPH-cytochrome P450 reductase sensitive to various promutagens. AB - We newly developed 10 Salmonela typhimurium TA1538 strains each co-expressing a form of human cytochrome P450s (P450 or CYP) together with NADPH-cytochrome P450 reductase (CPR) for highly sensitive detection of mutagenic activation of mycotoxins, polycyclic aromatic hydrocarbons, heterocyclic amines, and aromatic amines at low substrate concentrations. Each form of P450 (CYP1A1, CYP1A2, CYP2A6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, CYP3A4 or CYP3A5) expressed in the TA1538 cells efficiently catalyzed the oxidation of a representative substrate. Aflatoxin B1 was mutagenically activated effectively by CYP1A1, CYP1A2, and CYP3A4 and weakly by CYP2A6 and CYP2C8 expressed in S. typhimurium TA1538. CYP1A1 and CYP1A2 were responsible for the mutagenic activation of 2 amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and 2-acetylaminofluorene. Benzo[a]pyrene was also activated efficiently by CYP1A1 and weakly by CYP1A2, CYP2C9, CYP2C19, and CYP3A4 expressed in TA1538. These results suggest that the newly developed S. typhimurium TA1538 strains are applicable for detecting the activation of promutagens of which mutagenic activation is not or weakly detectable with N-nitrosamine-sensitive YG7108 strains expressing human P450s. PMID- 15279839 TI - Hitting the chemotherapy jackpot: strategy, productivity and chemistry. PMID- 15279840 TI - Targeting tumors by necrosis. PMID- 15279841 TI - Parasite movement: crucial anchor revealed. PMID- 15279842 TI - New ray of hope for cancer patients. PMID- 15279843 TI - Bob Ruffolo discusses the rhythm of drug development. AB - Since 2002, Robert Ruffolo has been President, Research & Development, Wyeth Pharmaceuticals, and Senior Vice President of Wyeth (http://www.wyeth.com). Following his postdoctoral fellowship at the National Heart, Lung and Blood Institute, he has worked in the pharmaceutical industry, initially spending 6 years at Lilly and then 16 years at SmithKline Beecham. He joined Wyeth in 2000 as Executive Vice President, Pharmaceutical R&D. Also an Adjunct Professor in the College of Pharmacy at Ohio State University, Bob has published nearly 500 full length papers and has received many awards more commonly presented to academics, such as the John Jacob Abel award from the American Society of Pharmacology and Experimental Therapeutics (ASPET) in 1998. PMID- 15279844 TI - A quiet revolution in lead optimisation services? PMID- 15279845 TI - The impact of cytochrome P450 allosterism on pharmacokinetics and drug-drug interactions. PMID- 15279846 TI - Fighting superbugs with superdrugs. PMID- 15279847 TI - From magic bullets to designed multiple ligands. AB - Increasingly, it is being recognised that a balanced modulation of several targets can provide a superior therapeutic effect and side effect profile compared to the action of a selective ligand. Rational approaches in which structural features from selective ligands are combined have produced designed multiple ligands that span a wide variety of targets and target classes. A key challenge in the design of multiple ligands is attaining a balanced activity at each target of interest while simultaneously achieving a wider selectivity and a suitable pharmacokinetic profile. An analysis of literature examples reveals trends and insights that might help medicinal chemists discover the next generation of these types of compounds. PMID- 15279848 TI - Whatever happened to cassette-dosing pharmacokinetics? AB - Cassette dosing is a procedure that is used for rapidly assessing the pharmacokinetics of a series of discovery drug candidates by dosing a mixture of compounds rather than a single compound. Cassette dosing has advantages and disadvantages associated with its use, which leads to controversy about how and if it should be used. To assess the current practices of the pharmaceutical industry regarding cassette dosing, a survey of several pharmaceutical companies was conducted. Analysis of the survey revealed that opinion on this subject is divided within the pharmaceutical industry. In addition, it was determined that approximately only a half of those companies that perform in vivo pharmacokinetic screening use cassette dosing for this purpose. PMID- 15279849 TI - Utility of homology models in the drug discovery process. AB - Advances in bioinformatics and protein modeling algorithms, in addition to the enormous increase in experimental protein structure information, have aided in the generation of databases that comprise homology models of a significant portion of known genomic protein sequences. Currently, 3D structure information can be generated for up to 56% of all known proteins. However, there is considerable controversy concerning the real value of homology models for drug design. This review provides an overview of the latest developments in this area and includes selected examples of successful applications of the homology modeling technique to pharmaceutically relevant questions. In addition, the strengths and limitations of the application of homology models during all phases of the drug discovery process are discussed. PMID- 15279850 TI - Ultrasound and transdermal drug delivery. AB - Transdermal drug delivery offers an attractive alternative to the conventional drug delivery methods of oral administration and injection. However, the stratum corneum acts as a barrier that limits the penetration of substances through the skin. Application of ultrasound to the skin increases its permeability (sonophoresis) and enables the delivery of various substances into and through the skin. This review presents the main findings in the field of sonophoresis, namely transdermal drug delivery and transdermal monitoring. Particular attention is paid to proposed enhancement mechanisms and future trends in the field of cutaneous vaccination and gene delivery. PMID- 15279855 TI - Traditional indigenous healing: Part I. AB - Traditional indigenous healing is widely used today, as it has been since time immemorial. This article describes the following areas in regards to traditional healing: (a) an explanation of indigenous peoples, (b) a definition of traditional indigenous healing, (c) a portrayal of traditional healers, (d) health within indigenous culture, (e) traditional healing techniques, (f) utilization of traditional healing, (g) how to find a traditional healer, and (h) comparing traditional healing principles with mainstream ways. It is important to have knowledge about this method of holistic healing so health care providers and nurses can integrate it into the health care for individuals and/or families that choose traditional indigenous healing. PMID- 15279856 TI - 'Take small steps to go a long way' consumer involvement in research into complementary and alternative therapies. AB - This investigation set out to learn about consumer involvement in complementary medicine research from those who have experience of practice in this area. A literature search was combined with written and oral responses from key people and organisations in the UK. Letter or e-mail contact was made with 59 key people and organisations and 43 people responded. Eighteen respondents were interviewed. The overall level of consumer involvement was low but participants provided examples of experiences of consumer involvement in commissioning, designing, carrying out, and disseminating research. Clear roles and tasks and a consumer friendly research environment, enabled consumers to contribute, gain confidence, and gradually widen their areas of involvement. There appears to be no single 'right way' for researchers and consumers to work together, but with experience and mutual respect researchers became increasingly enthusiastic about the value of the consumer perspective. As one consumer said: 'You have to take small steps to go a long way'. PMID- 15279857 TI - Complementary therapies to reduce physiological stress in pregnancy. AB - Pregnancy is a period of enormous physio-pathological and psychosocial adaptation in a woman's life. Although it is usually a time of joy and anticipation, many women experience some degree of anxiety, concern and fear regarding their own health and that of their babies, as well as the approaching labour. Worry about social, financial, occupational and relationship issues can often add to their stress levels which increases the possibility of pregnancy complications. However while mild to moderate stress facilitates successful adjustment to these demands, a significant increase in the levels of stress hormones may compromise the health of both mother and fetus. Complementary therapies are increasingly popular with expectant mothers and are gradually being integrated into conventional maternity care, primarily by midwives. However, there is debate about whether these therapies simply provide a form of relaxation for pregnant women with psychological stress or whether they could-or should-be used more constructively to deal with physio-pathological stress. This paper considers the physiological effects of certain complementary therapies in reducing the impact of stress in pregnancy. PMID- 15279858 TI - Development and evaluation of an inpatient [correction of impatient] holistic nursing care services department. AB - This paper describes the development and evaluation of a holistic nursing department at a 261-bed conventional, community hospital. Through the holistic nursing department, a nurse visits hospitalized inpatients. The visit might include complementary and alternative modalities (CAM) therapies, such as relaxation techniques, therapeutic touch, aromatherapy, and therapeutic suggestion. Evaluation of visits occurred through a retrospective chart review and patient satisfaction surveys. Main outcome measures were patient satisfaction, physiological changes, and pre- and post-distress scores. Discomfort and distress was decreased and patient satisfaction high when CAM therapies were used in conjunction with traditional inpatient medical and nursing care. PMID- 15279859 TI - The geography of complementary medicine: perspectives and prospects. AB - While complementary and alternative medicine (CAM) research has benefited from a range of social scientific perspectives, geographical contributions have been only slowly forthcoming. In this context, this paper illuminates the possibilities for CAM researchers to develop dedicated geographical perspectives. Some fundamental changes in the empirical and theoretical foci of medical/health geography are outlined, from a concern with mapping services and diseases in macro-space to investigating the dynamic between health and place. Highlighted are some important relational dynamics among CAM providers, patients/consumers and places of treatment and some general issues that could benefit from a geographical analysis. The concurrent research agenda is located across the sub disciplinary strata of human geography. PMID- 15279860 TI - Psychological treatment of three urban fire fighters with post-traumatic stress disorder using eye movement desensitisation reprocessing (EMDR) therapy. AB - Fire fighters are at increased risk of developing mental health problems due to the nature of their work, which can sometimes be extremely traumatic. Arranging for immediate access to mental health specialists can often take a protracted time to arrange, leading to the individual remaining disabled and off work. The South Wales fire and rescue service have responded to this challenge and formed a partnership with their local NHS traumatic stress service. This has enabled fire fighters to receive early psychological assessment and treatment from a nurse therapist trained in cognitive behaviour therapy or referred to a consultant liaison psychiatrist. This paper will describe 3 cases which all suffered with PTSD and were treated via the partnership with a controversial therapy EMDR. PMID- 15279861 TI - Paediatric nurses' attitudes to massage and aromatherapy massage. AB - Complementary therapies have continued to increase in popularity in healthcare and it is widely accepted that they can be incorporated into the nursing role. However, this acceptance does not necessarily mean that the introduction of therapies into the nursing arena has been without confusion and without professional and legal implications. Consequently, this small-scale, qualitative study aimed to explore the perceptions and lived experiences of paediatric nurses of two therapies, namely massage and aromatherapy massage. There is a dearth of literature exploring nurses' perceptions to the incorporation of these therapies, especially in the arena of paediatric nursing where massage and aromatherapy massage are common practice. Semi-structured interviews were undertaken with qualified nurses and revealed the themes of 'benefit', 'family centred care', 'nursing care' and 'being held back'. It was found that at some stage during their professional career each nurse had performed massage and/or aromatherapy massage. All nurses were able to recall certain benefits of the therapies for the children that they had observed and many discussed the importance of involving the family as a way of including them in to the care of their child. However, for the nurses in this study, it was evident that the incorporation of complementary therapies into the nursing role was determined by the context in which they practised. Due to the dominance of the medical model, nurses faced pressures and conflicts in the realities of their nursing work, which meant they were often unable to carry out these therapies. PMID- 15279862 TI - ASMS board of directors. PMID- 15279863 TI - Directory of corporate members. PMID- 15279864 TI - Directory of members. PMID- 15279865 TI - Medical risks in epilepsy: a review with focus on physical injuries, mortality, traffic accidents and their prevention. AB - The present review aims at highlighting selective aspects of the medical risks in epilepsy and their prevention. Emphasis is put on accidents and physical injuries, including risk factors and effectiveness of prevention; mortality, its causes, risk factors and prevention of seizure-related deaths, as well as traffic accidents, their risk factors and the effectiveness of prevention. Accidents and injuries are slightly more frequent among people with epilepsy than in the general population. This increased risk is probably most prevalent in patients with symptomatic epilepsy and frequent seizures, most often in combination with associated handicaps. The majority of accidents are trivial and occur at home. The most frequent injuries among patients with epilepsy are contusions, wounds, fractures, abrasions and brain concussions. The standardised mortality ratio (SMR; the ratio of observed number of deaths in a population with epilepsy to that expected, based on age and sex-specific mortality rates in a reference population) in population-based studies of epilepsy is 2-3 compared to the general population. This increased mortality is largely related to the etiology of the epilepsy and is probably not influenced by the treatment of the epilepsy. On the other hand, most fatalities in patients with chronic, therapy resistant epilepsy seem to be seizure-related and often sudden unexpected deaths (SUDEP). The frequency of such seizure-related deaths is most likely to be reduced by intensified treatment aiming at early seizure control, although appropriate studies for definitive evidence are still lacking. Apparently, there is an increased rate of traffic accidents in drivers with epilepsy, even if population based prospective data are lacking. Many of these accidents are seizure-related. Probably, the extent to which physicians report their patients with uncontrolled epilepsy to the authorities is too low, but this has not yet been explored. Moreover, the preventive measures in legislation may be ignored by many people with epilepsy. PMID- 15279866 TI - MPEP, an antagonist of metabotropic glutamate receptors, exhibits anticonvulsant action in immature rats without a serious impairment of motor performance. AB - An antagonist of type I metabotropic glutamate receptors MPEP was found to exhibit anticonvulsant action in adult rodents. Present experiments were focused on action of this drug against pentetrazol-induced motor seizures in immature rats 12-, 18- and 25-days old. Dose of pentetrazol (100 mg/kg s.c.) was chosen to elicit minimal clonic seizures and (after a longer latency) generalized tonic clonic seizures. Pretreatment with MPEP (doses from 10 to 80 mg/kg i.p.) resulted in a dose-dependent suppression of the tonic phase of generalized tonic-clonic seizures in all age groups studied. Efficacy of MPEP was higher and the effect lasted longer in 12- than in 25-day-old rats. In addition, minimal clonic seizures were suppressed in 18-day-old rats. Motor abilities of immature animals were not compromised by MPEP in doses of 20 and/or 40 mg/kg i.p., only righting reflex was a little slowed down in 12- and 18-day-old rats. In contrast to antagonists of ionotropic glutamate receptors anticonvulsant doses of MPEP do not induce unwanted side effects in motor performance of developing rats. PMID- 15279867 TI - Brain-derived neurotrophic factor (BDNF) Val66Met polymorphisms in febrile seizures. AB - Various studies have shown that brain-derived neurotrophic factor (BDNF) increased neuronal excitability. We tested that BDNF might be involved in the etiology of febrile seizures (FSs). A total of 186 Taiwanese children were divided into two groups: (1) FSs (n = 104); (2) normal control subjects (n = 83). A single base pair polymorphism SNP6265 (Val66Met) at position 196 was analyzed. Our findings suggest that the BDNF polymorphisms were not candidate genetic markers. PMID- 15279868 TI - Modeling remission and relapse in pediatric epilepsy: application of a Markov process. AB - Seizure outcome is frequently described in terms of patients ever attaining remission or being in terminal remission. Outcomes are more complicated and, over many years, repeated remission and relapses may occur. These are difficult to quantify with standard survival techniques used in analysis of remission and relapse. The Markov process, which allows one to track a patient's state (remission or not) over time, provides a suitable approach for studying repeated remission and relapse. In a prospective community-based study of children followed from the point of the initial diagnosis of epilepsy, we examined the probability of repeated remission and relapse over up to three different remission episodes (minimum 1 year each) per patient. The role of epilepsy syndrome was the main determinant of remission-relapse patterns considered in the analysis. Two different Markov models were used, one involving three states and the other seven states. Of 613 children initially recruited into the study, 602 were followed at least 1 year and thus eligible for the analysis. Almost 90% of the cohort experienced a remission; however, almost half then relapsed. Second remissions occurred in 81% of those who relapsed of whom 38% relapsed again. A third remission occurred in 82% of those after a second relapse of whom 58% relapsed yet again. After the first 2 years, approximately 70% of the cohort was in remission, 20% was no longer in remission having relapsed, and 10% had never been in remission. Significant differences were seen by underlying epilepsy syndrome. Children with one of the epileptic encephalopathies were least likely of all syndrome groups ever to remit. Those with symptomatic partial epilepsies were less likely to remit than children with any of the other syndromes, idiopathic partial or generalized, cryptogenic partial, and unclassified. Differences between these last groups became apparent when considering their subsequent remission and relapse histories. These differences were best seen in the seven-state model. For example, idiopathic partial epilepsies were most likely to enter remission and never relapse. By contrast, idiopathic generalized and cryptogenic partial epilepsies were more likely to remit and relapse repeatedly. The Markov approach provides an alternative to standard survival techniques for understanding remission and relapse outcomes in epilepsy. Its advantage is that it allows one to track the individuals' outcome over time even as the condition fluctuates. The technique would also be applicable in virtually any remitting-relapsing disorder. PMID- 15279869 TI - Inhibition of aconitase in astrocytes increases the sensitivity to chemical convulsants. AB - Although there is evidence that astrocytes support neuronal function, the contribution of astrocytes to seizure onset and termination is not known. To determine whether there are changes in seizure susceptibility or neuronal damage when the ability of astrocytes to generate ATP is reduced, 0.5 nmol of fluorocitrate (FC) was injected into the right ventricle. Injection of FC alone did not produce electrographic or behavioral seizures and did not stress or injure neurons or astrocytes, as measured with silver stain and immunohistochemistry for HSP32 or HSP72. However, in animals pretreated with FC, administration of kainic acid, at a dose that does not initiate seizures in control animals (7 mg/kg), caused wet dog shakes and neuronal damage in the hilus. Wet dog shakes did not cause any neuronal damage in control animals. If the dose of FC was increased to 0.75 nmol, then subsequent administration of the same dose of kainic acid (7 mg/kg) caused stage 3-5 seizures. Injection of FC also reduced the dose of pilocarpine needed to produce seizures. Given simultaneously with FC, isocitrate, which bypasses the biochemical inhibition of aconitase, blocked the effects of FC in both kainic acid and pilocarpine treated animals. The data demonstrate that inhibition of aconitase in astrocytes lowers the doses of both kainic acid and pilocarpine that will cause behavioral seizures and may increase neuronal vulnerability to seizures. PMID- 15279870 TI - 18FDG-PET in epilepsies of infantile onset with pharmacoresistant generalised tonic-clonic seizures. AB - AIMS: To investigate the pathophysiology of pharmacoresistant epilepsies with cryptogenic generalised tonic-clonic seizures (GTCS) from infancy. METHODS: 18F Deoxy-Glucose-Positron Emission Tomography 18FDG-PET) with statistical parametric mapping (SPM). Inclusion criteria were: pharmacoresistant chronic epilepsy with GTCS commencing in infancy, no focal seizures except alternating hemiconvulsions and no focal epileptic discharges in the EEG during the first year of the disease, no focal changes upon routine neuroradiological investigations, no indication of brain damage according to history and clinical examination. RESULTS: 15 boys and 15 girls with a mean age of 6.4 years (range l-14 years) were included. All still suffered from seizures despite past treatment with a mean of five drugs. Nearly all were mentally retarded, 19 to a severe and 10 to a minor degree. Fifteen were ataxic and 11 hypotonic. The EEG in 23 showed irregular generalised spike-wave discharges. PET SPM analysis revealed bioccipital hypometabolism related to sedation. Pathological monofocal hypometabolic areas were found in three, multifocal hypometabolic areas in 22 and diffuse bilateral hypometabolism in three patients. Frontal hypometabolism correlated to the degree of mental retardation, hypotonia, and ataxia. Temporomesial hypometabolism correlated to the occurrence of obtunded states and prominent delta rhythms in the EEG. Central and parietal changes were associated with the occurrence of myoclonic seizures and spike-wave discharges. CONCLUSIONS: 18FDG-PET in many of these children with cryptogenic generalised epilepsies showed multifocal hypometabolic areas of unknown aetiology. Primary cortical microdysgenesis and secondary changes due to the severe and long-standing epilepsy must be considered. Only a minority of patients showed restricted focal hypometabolism as a possible indication for surgical treatment. PMID- 15279871 TI - Vinpocetine inhibits the epileptic cortical activity and auditory alterations induced by pentylenetetrazole in the guinea pig in vivo. AB - Here we investigate the effect of the neuroprotective drug, vinpocetine on the epileptic cortical activity, on the alterations of the later waves of brainstem auditory evoked potentials (BAEPs) and on the hearing decline induced by the convulsing agent, pentylenetetrazole (PTZ). Vinpocetine at doses from 2 to 10 mg/kg inhibits the tonic-clonic convulsions induced by PTZ (100 mg/kg). Vinpocetine injected at a dose of 2 mg/kg 4 h before PTZ completely prevents the characteristic electroencephalogram (EEG) changes induced by PTZ for the ictal and post-ictal periods. Vinpocetine also abolished the PTZ-induced changes in the amplitude and latency of the later waves of the BAEPs in response to pure tone burst monoaural stimuli (frequency 8 or 4 kHz intensity 100 dB), and the PTZ induced increase in the BAEP threshold. These results show the antiepileptic potential of vinpocetine and indicate the capability of vinpocetine to prevent the changes in the BAEP waves associated with the hearing loss observed during generalized epilepsy. PMID- 15279872 TI - NTP center for the evaluation of risks to human reproduction reports on phthalates: addressing the data gaps. PMID- 15279874 TI - Effect of the anti-androgenic endocrine disruptor vinclozolin on embryonic testis cord formation and postnatal testis development and function. AB - Vinclozolin is a systemic dicarboximide fungicide that is used on fruits, vegetables, ornamental plants, and turf grass. Vinclozolin and its metabolites are known to be endocrine disruptors and act as androgen receptor antagonists. The hypothesis tested in the current study is that transient embryonic exposure to an anti-androgenic endocrine disruptor at the time of testis determination alters testis development and subsequently influences adult spermatogenic capacity and male reproduction. The effects of vinclozolin on embryonic testicular cord formation in vitro were examined, as well as the effects of transient in utero vinclozolin exposure on postnatal testis development and function. Embryonic day 13 (E13, sperm-positive vaginal smear day = E0) gonads were cultured in the absence or presence of vinclozolin (50-500microM). Vinclozolin treated gonads had significantly fewer cords (P < 0.05) and the histology of the cords that formed were abnormal as compared to vehicle-treated organs. Pregnant rats were exposed to vinclozolin (100 mg/kg/day) between embryonic days 8 and 14 (E8-E14) of development. Testis morphology and function were analyzed from postnatal day (P) 0, pubertal P20, and adult P60. No significant effect of vinclozolin on testis histology or germ cell viability was observed in P0 testis. The pubertal P20 testis from vinclozolin exposed animals had significantly higher numbers of apoptotic germ cells (P < 0.01), but testis weight was not affected. The adult P60 sperm motility was significantly lower in vinclozolin exposed males (P < 0.01). In addition, apoptotic germ cell number in testis of vinclozolin exposed animals was higher in adult P60 animals. Observations demonstrate that vinclozolin can effect embryonic testicular cord formation in vitro and that transient in utero exposure to vinclozolin increases apoptotic germ cell numbers in the testis of pubertal and adult animals. This correlated to reduced sperm motility in the adult. In conclusion, transient exposure to vinclozolin during the time of testis differentiation (i.e. cord formation) alters testis development and function. Observations indicate that transient exposure to an anti-androgenic endocrine disruptor during embryonic development causes delayed effects later in adult life on spermatogenic capacity. PMID- 15279875 TI - High-resolution image cytometry of rat sperm nuclear shape, size and chromatin status. Experimental validation with the reproductive toxicant vinclozolin. AB - Recent studies have shown that the complex inter-related processes of sperm chromatin organization and nuclear morphogenesis, both of which are important fertility determinants, may be disrupted by chemicals. A high-resolution image cytometry method has been developed, using the fluorescent dye bisbenzimide, for the measurement of 20 features of the sperm nucleus related to size, form and chromatin status in the rat. For the complete set of features measured and from a total of 150 spermatozoa assessed per sample, the overall coefficient of reproducibility was 5%. Then, an experimental validation of the method was carried out in rats chronically exposed to the antiandrogenic reproductive toxicant vinclozolin and control animals. Univariate statistics revealed significant vinclozolin-induced changes for 19 out of 20 morphometric and chromatin features. Stepwise linear discriminant analysis classified correctly 84.3% of the sperm nuclei with only four features selected. The accuracy and reproducibility of the cytometry assessment of the sperm nuclei together with the results of the experimental validation suggest this method may be a new powerful tool for use in reproductive toxicology. PMID- 15279876 TI - P-glycoprotein expression and distribution in the rat placenta during pregnancy. AB - P-glycoprotein (P-gp) is a drug efflux transporter that limits the entry of various potentially toxic drugs and xenobiotics into the fetus and is thus considered a placental protective mechanism. In this study, P-gp expression was investigated in the rat chorioallantoic placenta over the course of pregnancy. Three methods have been employed: real-time RT-PCR, western blotting and immunohistochemistry. The expression of mdr1a and mdr1b genes was demonstrated as early as on the 11th gestation day (gd) and increased with advancing gestation. Western blotting analysis revealed the presence of P-gp in the rat placenta starting from gd 13 onwards. P-gp was localized in the developing labyrinth zone of the placenta on gd 13; from gd 15 up to the term P-gp was seen as a dot like continuous line in the syncytiotrophoblast layers. Our data confirm the presence of P-gp in the rat chorioallantoic placenta starting soon after its development, which may signify the involvement of P-gp in transplacental pharmacokinetics during the whole period of placental maturing. PMID- 15279877 TI - Histomorphometric alteration and cell-type specific modulation of arylhydrocarbon receptor and estrogen receptor expression by 2,3,7,8-tetrachlorodibenzo-p-dioxin and 17beta-estradiol in mouse experimental model of endometriosis. AB - Our purpose was to examine the effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), either singly (10 microg/kg) or with 17beta-estradiol (E2, 100 microg/kg), on the growth of endometriosis in a mouse endometriosis model by employing histo-morphometrical analysis as well as expression of arylhydrocarbon receptor (AhR) and estrogen receptor (ER). Epithelial height, stromal thickness, and proliferative activity of the endometriotic lesions were significantly increased by E2 in ovariectomized mice, whereas co-administered TCDD significantly reduced these effects. TCDD alone did not affect the proliferative activity but rather reduced the epithelial height and stromal thickness. ER expression in the luminal epithelium was decreased by E2 compared with ovariectomy alone, while TCDD significantly increased it. On the other hand, stromal ER expression was significantly increased by ovariectomy and decreased by E2, though TCDD did not further enhance this expression. These results indicate that a short-term exposure to TCDD failed to increase the growth of endometriotic lesion and the direct effect of TCDD probably depends on a cell-specific interaction with ovarian steroids mediated by their own receptors. These initial findings in intact tissue of mouse endometriosis may suggest critical roles of steroid hormones in the pathogenesis of endometriosis in relation to endocrine disruptors. PMID- 15279878 TI - Effects of maternal xenoestrogen exposure on development of the reproductive tract and mammary gland in female CD-1 mouse offspring. AB - The objective of this study was to examine the effects of maternal exposure to xenoestrogen, at levels comparable to or greater than human exposure, on development of the reproductive tract and mammary glands in female CD-1 mouse offspring. Effects of genistein (GEN), resveratrol (RES), zearalenone (ZEA), bisphenol A (BPA) and diethylstilbestrol (DES) were examined. Beginning on gestational day 15, pregnant CD-1 mice were administered four daily subcutaneous injections with 0.5 or 10 mg/kg/day of GEN, RES, ZEA or BPA, 0.5 or 10 microg/kg/day of DES dissolved in dimethylsulfoxide (DMSO), or DMSO vehicle (n = 6). Vaginal opening was monitored, 6 animals per group were autopsied at 4, 8, 12 and 16 weeks of age and estrous cyclicity was monitored from 9 to 11 weeks of age. Maternal exposure to xenoestrogen accelerated puberty onset (vaginal opening) and increased the length of the estrous cycle; mice treated with GEN, RES, BPA or DES spent more time in diestrus, and ZEA-treated mice spent more time in estrus. Lack of corpora lutea and vaginal cornification were observed at 4 weeks of age in the high-dose GEN (33%) and RES (17%) groups, and in the high- and low-dose BPA groups (33 and 50%, respectively) and DES groups (83 and 100%, respectively). Lack of corpora lutea and vaginal cornification was observed in the high-dose ZEA group at 4, 8, 12 and 16 weeks of age (83, 100, 83 and 33%, respectively). Mammary gland differentiation was accelerated in ZEA- and BPA treated mice with corpora lutea at 4 weeks of age. ZEA-treated mice without corpora lutea showed mammary growth arrest at 8, 12 and 16 weeks of age; their mammary glands consisted only of a dilatated duct filled with secreted fluid. Mammary gland growth was similar with xenoestrogens other than ZEA or BPA to that of the controls at all time points. High-dose GEN and RES and high- and low-dose BPA and DES exerted transient effects on the reproductive tract and mammary glands, whereas ZEA exerted prolonged effects. PMID- 15279879 TI - Is embryo-cryopreservation really neutral? A new long-term effect of embryo freezing in mice: protection of adults from induced cancer according to strain and sex. AB - BACKGROUND: Beneficial or harmful effects of embryo freezing have been described in man and animals, raising the question of the neutrality of this technique. OBJECTIVE: We examined, in mice, the possibility that embryo freezing influences the probability of the emergence, in adults, of an induced urinary bladder cancer. METHODS: The experiment was conducted in mice derived from embryos of two different genotypes. Females receiving embryos were parsed into two groups according to whether these embryos were cryopreserved or not. The derived adults received the chemical carcinogen N-butyl-N-4hydroxybutylnitrosamine (BBN), in the drinking water. Time to death since the onset of treatment was measured for each animal until the 300th day. RESULTS: In females from one of the two strains tested, embryo freezing led to a favorable long-term effect on the probability of resistance to induced cancer. CONCLUSION: This beneficial effect, taken together with other effects reported in the literature be they beneficial or harmful, suggests that embryo freezing in mice may not be neutral. PMID- 15279881 TI - Influence of colloid, preservation medium and trimetazidine on renal medulla injury. AB - In organ transplantation, preservation injury is an important factor which could influence short-term and long-term graft outcome. The renal medulla is particularly sensitive to oxidant stress and ischemia-reperfusion injury (IRI). Using an autotransplant pig kidney model, we investigated renal function and medullary damage determined between day 1 and week 2 after 24- or 48-h cold storage in different preservation solutions: University of Wisconsin solution (UW), Hopital Edouard Herriot solution (a high Na+ version of UW), ECPEG (high Na+ preservation solution with PEG) and ICPEG (a high K+ version of ECPEG) with or without trimetazidine (TMZ). TMZ improved renal preservation and increased renal function when added in each preservation solution (particularly HEH and ECPEG). Medullary damage led to the early appearance of trimethylamine-N-oxide (TMAO) followed by 1H-NMR in urine and plasma. TMZ and ECPEG is the most efficient association to reduce medullary damage. This study clarifies the role of colloid and polarity solution and the role of mitochondrial protection by TMZ. PMID- 15279880 TI - Reproductive evaluation of aqueous crude extract of Achillea millefolium L. (Asteraceae) in Wistar rats. AB - The present study was conducted to evaluate the toxicity of the exposure to the aqueous extract from leaves (AE) of Achillea millefolium L. on reproductive endpoints in Wistar rats. Adult male rats were treated daily with yarrow extract (0.3, 0.6 and 1.2 g/kg/day) during 90 days by oral gavage. Endpoints including reproductive organ weights, sperm and spermatid numbers as well as sperm morphology were evaluated. No clinical signs of toxicity were detected over the treatment period, and body weight gain was similar in all groups. A significant increase in the percentage of abnormal sperm in the group treated with the highest dose of yarrow extract was detected with no other important changes in the other reproductive endpoints studied in the male rats. Furthermore, a possible estrogenic/antiestrogenic activity of the yarrow extract screened after a 3-day treatment of immature female rats which did not show any uterotrophic effects. PMID- 15279882 TI - Pioglitazone induces plasma platelet activating factor-acetylhydrolase and inhibits platelet activating factor-mediated cytoskeletal reorganization in macrophage. AB - Platelet activating factor (PAF) is a key molecule for inflammation. To examine a role of peroxisome proliferator-activated receptor gamma (PPARgamma) in inflammatory reactions of atherosclerosis, we investigated the effects of 15 deoxy-(Delta12,14)-Prostaglandin J2 (15d-PGJ2) and pioglitazone, PPARgamma ligands, on plasma PAF-acetylhydrolase (PAF-AH) expression in THP-1 macrophages. PAF-AH mRNA and protein were up-regulated by the PPARgamma ligands. Prostaglandin F2alpha (PGF2alpha), a PARgamma inhibitor, abrogated the up-regulation of PAF-AH mRNA by pioglitazone, suggesting that PPARgamma activation is involved in the induction of PAF-AH by pioglitazone. As PAF promotes the cell motility with cytoskeletal reorganization, we investigated the effect of pioglitazone on PAF mediated morphological changes in THP-1 macrophages. In the absence of pioglitazone, PAF promoted the elongation of actin cytoskeleton, which was inhibited by pretreatment with pioglitazone. In contrast, pioglitazone was not able to inhibit the morphological changes induced by C-PAF, a non-hydrolyzable PAF agonist. Thus, it is suggested that PAF-induced morphological changes could be inhibited by pioglitazone through PAF-AH, which rapidly hydrolyzed PAF. These data propose that PPARgamma/PAF-AH pathway is a clinical target for the prevention against atherosclerosis. PMID- 15279883 TI - Characterization of lectin aggregates in the hemolymph of freshwater prawn Macrobrachium rosenbergii. AB - In invertebrates, lectins play relevant roles in innate immunity; however, their regulatory mechanisms have not been identified yet. In this work, we purified, by gel filtration and affinity chromatography, lectin aggregates circulating in the hemolymph of the freshwater prawn Macrobrachium rosenbergii and compared their physicochemical properties with a previously described lectin (MrL). High molecular weight MrL aggregates (MrL-I) lack hemagglutinating activity and showed bands of 62.1, 67.1 and 81.4 kDa, whereas MrL-III, which corresponds to MrL, showed hemagglutinating activity and is constituted by a single 9.6-kDa band as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS PAGE) analysis. MrL-I and MrL-III showed similar amino acid composition but different carbohydrates concentration. Edman degradation indicated NH2-terminal sequence of five amino acids for the 9.6-kDa MrL-III (DVPLL/A) and eleven for the main 81.4-kDa band identified in MrL-I (DVPLL/AXKQQQD); analysis by MALDI-TOF indicated a different tryptic pattern for MrL-I and MrL-III. MrL-I was recognized by monoclonal antibodies against MrL-III. Circular dichroism indicated that the secondary structure in both proteins is similar and contains 23% of beta-sheet and 24% of alpha-helix. Our results suggest that differential posttranslational processes that favor aggregation are involved in regulating the activity of the lectin. PMID- 15279884 TI - Peptides binding to a Gb3 mimic selected from a phage library. AB - Peptides binding to a Gb3 mimic were selected from 12-mer peptide library. The self-assembled monolayer (SAM) of a Gb3 mimic was formed on the gold surface, and biopanning was carried out with the phage display peptide library. After three rounds of biopanning, four individual sequences were obtained from 10 phage clones, and the selected peptides having the specific 7-mer sequence (FHENWPS) showed affinities to the Gb3 mimic as strong as to RCA120. Molecular dynamics calculations suggested that the peptides bound to the Gb3 mimic by hydrophobic interaction and hydrogen bonding formation, and the cooperative interactions played an important role in the recognition. The Stx-1 binding was inhibited by the peptides. PMID- 15279885 TI - The interaction of trypsin with trehalose: an investigation of protein preservation mechanisms. AB - Preservation of the native protein structure and biological activity in dry protein/excipient mixtures has been previously attributed to either the glass forming properties of the additives or to their ability to hydrogen bond to the protein. There is evidence that both processes are important but it has not yet been elucidated which is the limiting factor that determines the efficiency of a given molecule as a protectant. In this work, gravimetric measurements together with enzyme activity assays have been employed to investigate the protection of proteins by sugars, through direct interaction via hydrogen bonding and as the result of glass formation. As a model protein, trypsin has been employed and the modes of action of two similar disaccharides, sucrose and trehalose, which offer different levels of protection, evaluated and compared. Data obtained on freeze dried formulations indicate that protein and sugars interact through hydrogen bonding to protein hydration sites. The extent of interaction is found to change dramatically at elevated temperatures; sucrose showing a significantly decreased, and trehalose a considerably increased level of interaction. Protein preservation is shown to be directly related to the number of hydrogen bonds formed. Possible reasons why trehalose interacts more extensively with the protein than sucrose are discussed in terms of differences in the anhydrous structures and molecular mobilities of the sugar molecules. PMID- 15279886 TI - A metabolic model describing the H2O2 elimination by mammalian cells including H2O2 permeation through cytoplasmic and peroxisomal membranes: comparison with experimental data. AB - We have constructed a metabolic model describing the H2O2 elimination by mammalian cells. It comprises three compartments (medium, cytosol, and peroxisome) separated by cytoplasmic and peroxisomal membranes, and H2O2 moves across the membranes with different permeation rate constants. Catalase localizes to peroxisomes, while glutathione peroxidase (GPx) and GSH recycling system (glutathione reductase (GR) and the oxidative pentose phosphate pathway (PPP)) localize to cytosol. The rates of individual enzyme reactions were computed using the experimentally determined activities and rate equations known for mammalian enzymes. Using the model, the concentration dependence of H2O2 elimination rate was obtained by numerical simulation and was compared with experimental data obtained previously with cultured mammalian cells (fibroblasts, human umbilical vein endothelial cells (HUVEC), and PC12 cells). The model was shown to be able to reproduce the data well by assuming appropriate values for the permeability rate constants. The H2O2 permeability coefficients thus estimated for cytoplasmic and peroxisomal membranes were in the same order of magnitude, except that the value for cytoplasmic membrane of PC12 cell was significantly smaller. The results suggest that the membrane permeability is one of the rate-limiting factors in the H2O2 elimination by mammalian cells. Using the model and estimated parameter values, we have examined the rate-limiting enzyme of the metabolic system, as well as the intracellular H2O2 concentration under steady-state and non-steady-state conditions. PMID- 15279887 TI - Interleukin-1 inhibits voltage-dependent P/Q-type Ca2+ channel associated with the inhibition of the rise of intracellular free Ca2+ concentration and catecholamine release in adrenal chromaffin cells. AB - Effects of interleukin (IL) on intracellular free Ca2+ concentration ([Ca2+]i) rise and catecholamine (CA) release were examined in isolated, cultured bovine adrenal chromaffin cells. IL-1alpha and IL-1beta inhibited the rise of [Ca2+]i and CA release induced by acetylcholine (ACh) and excess KCl both in normal and in Ca2+-sucrose medium. Pretreatment by IL-1 receptor antagonist (IL-1RA) blocked the inhibitory actions of IL-1alpha. IL-1alpha reduced CA release induced by veratridine in normal medium but not in the presence of diltiazem. Analysis using specific blockers for voltage-operated Ca2+ channels (VOCC) revealed that IL 1alpha and IL-1beta specifically inhibited the P/Q-type Ca2+ channel to reduce [Ca2+]i rise induced by excess KCl. IL-1 did not affect [Ca2+]i rise induced either by bradykinin or caffeine in Ca2+-deprived medium or via activation of store-operated Ca2+ channel (SOC). The inhibitory effects of IL-1alpha were blocked by pretreatments with herbimycin A, U0126 and PD 98054, but not with SB202190, SP 600125 or pertussis toxin (PTX). These results demonstrated that IL 1 inhibits stimulation-evoked [Ca2+]i rise and CA release in chromaffin cells by blocking voltage-operated P/O-type Ca2+ channels. The inhibitory action of IL-1 may be mediated through the tyrosine kinase and MEK/ERK pathways. PMID- 15279888 TI - Interaction of mushroom tyrosinase with aromatic amines, o-diamines and o aminophenols. AB - 3-Amino-L-tyrosine was found to be a substrate of mushroom tyrosinase, contrary to what had previously been reported in the literature. A series of amino derivatives of benzoic acid were tested as substrates and inhibitors of the enzyme. 3-Amino-4-hydroxybenzoic acid, 4-amino-3-hydroxybenzoic acid and 3,4 diaminobenzoic acid were oxidized by this enzyme, as previously reported for Neurospora crassa tyrosinase, but 4-aminobenzoic acid and 3-aminobenzoic acid were not. Interestingly, 3-amino-4-hydroxybenzoic acid was oxidized five times faster than 4-amino-3-hydroxybenzoic acid, confirming the importance of proton transfer from the hydroxyl group at C-4 position. All compounds inhibited the monophenolase activity but their effect on the diphenolase activity was small or negligible. 3-Amino-4-hydroxybenzoic acid was a stronger inhibitor than 4-amino-3 hydroxybenzoic acid, indicating their different binding affinity to the oxy form of the enzyme. Both, however, were weaker inhibitors than 3-amino-L-tyrosine, 4 methoxy-o-phenylenediamine and 3,4-diaminobenzoic acid, which was the strongest inhibitor from among the compounds tested. These results show that the relative positioning of the amino group and the hydroxy group in o-aminophenols with respect to the side chain is important both for binding to the dicopper center and for catalysis. PMID- 15279889 TI - Heparan sulfate and control of endothelial cell proliferation: increased synthesis during the S phase of the cell cycle and inhibition of thymidine incorporation induced by ortho-nitrophenyl-beta-D-xylose. AB - The effect of xylosides on the synthesis of [35S]-sulfated glycosaminoglycans by endothelial cells in culture was investigated. Ortho-nitrophenyl-beta-D-xylose (10(-3)M) produces a dramatic enhancement on the synthesis of heparan sulfate and chondroitin sulfate secreted to the medium (20- and 100-fold, respectively). Para nitrophenylxyloside, at the same concentration, produces an enhancement of only 37- and 3-fold of chondroitin sulfate and heparan sulfate, respectively. These differences of action seem to be related with the higher lipophilic character of ortho-nitrophenyl-xyloside. A lower enhancement of the synthesis of the two glycosaminoglycans is also observed with 2-naphtol beta-D-xylose and cis/trans decahydro-2-naphtol beta-D-xylose. Besides stimulating the synthesis, O nitrophenyl-beta-D-xylose as PMA [J. Cell. Biochem. 70 (1998) 563] also inhibits [3H]-thymidine incorporation by quiescent endothelial cells stimulated for growth by fetal calf serum (FCS). The combination of xylosides with PMA produced some cumulative effect. PMA stimulates the synthesis of heparan sulfate mainly at G1 phase whereas the highest enhancement of synthesis produced by the xylosides is in the S phase of the endothelial cell cycle. PMID- 15279890 TI - Evaluation of active recombinant catalytic domain of human ErbB-2 tyrosine kinase, and suppression of activity by a naturally derived inhibitor, ZH-4B. AB - Human cancers frequently express high levels of ErbB-2 tyrosine kinase, which is associated with aggressive tumor behavior and poor prognosis. ErbB-2 is thus a promising target for cancer therapy. Here we express the catalytic domain of ErbB 2 as a soluble active kinase, and investigate the correlations between its activity and kinase concentration, ATP concentration, substrate concentration and divalent cation type. A simple and effective screening model is established to identify and evaluate potential inhibitors of ErbB-2 kinase. ZH-4B, a naturally derived small molecule compound that potently inhibits ErbB-2 kinase activity with an IC50 value of 2.45+/-0.56 microM, is identified. In SK-OV-3 human ovarian cancer cells and SK-BR-3 human breast carcinoma cells, ZH-4B blocks epidermal growth factor (EGF)-induced phosphorylation of ErbB-2 in a dose-dependent manner. Our data collectively indicate that ZH-4B is a potential novel anti-cancer agent that deserves further investigation. PMID- 15279891 TI - Expression of erbB receptors mRNA in thyroid tissues. AB - ErbB family members, such as epidermal growth factor receptor 1 (erbB1), erbB2, erbB3 and erbB4, are widely distributed in organ tissues, and these receptors are suspected tumorigenesis factors. We measured erbB mRNA in thyroid tissues of benign and malignant thyroid tumors or Graves' disease using Genescan. ErbB2 is associated with aggressive cancers and is used as a biological marker for the disease; Northern blot and reverse transcription-polymerase chain reaction (RT PCR) analyses have shown it to be increased in Graves' disease. Additional studies indicated a similar result in papillary carcinoma cells; mRNAs of erbB2 and erbB3 were increased but erbB4 mRNA was decreased, suggesting distorted erbB expression may be associated with tumorigenesis. However, only erbB2 overexpression is associated with Graves' disease. These data further implicate transmembrane-type receptors in tumorigenesis in the thyroid. PMID- 15279892 TI - Regulation of transcription and activity of Rhizobium etli glutaminase A. AB - The present study determines the regulatory mechanisms that operate on Rhizobium etli glutaminase A. glsA gene expression levels were evaluated under several metabolic conditions by fusions of the glsA gene promoter and the transcriptional reporter cassette uidA2-aad. glsA expression was directly correlated to the glutaminase A activity found under the tested growth conditions, reaching its maximum level in the presence of glutamine and during exponential growth phase. Glutamine induces glsA expression. The influence of allosteric metabolites on glutaminase A activity was also determined. The purified enzyme was inhibited by 2-oxoglutarate and pyruvate, whereas oxaloacetate and glyoxylate modulate it positively. Glutaminase A is not inhibited by glutamate and is activated by ammonium. Glutaminase A participates in an ATP-consuming cycle where glutamine is continually degraded and resynthesized by glutamine synthetase (GS). GS and glutaminase A activities appear simultaneously during bacterial growth under different metabolic conditions and their control mechanisms are not reciprocal. Slight overproduction in glutaminase A expression causes a reduction in growth yield and a dramatic decrease in bacterial growth. We propose a model for regulation of glutaminase A, and discuss its contribution to glutamine cycle regulation. PMID- 15279893 TI - Dietary fat mediates hyperglycemia and the glucogenic response to increased protein consumption in an insect, Manduca sexta L. AB - Many insects display non-homeostatic regulation over blood sugar level. The concentration of trehalose varies dramatically depending on physiological and nutritional state. In the absence of dietary carbohydrate, blood trehalose in larvae of the lepidopteran insect Manduca sexta is maintained by gluconeogenesis and is dependent on dietary protein consumption. In the present study, the effect of dietary fat on the glucogenic response of insects to increased dietary protein was examined by NMR analysis of (2-13C)pyruvate metabolism. Last instar larvae were maintained on a carbohydrate-free chemically defined artificial diet having variable levels of casein with and without corn oil. Gluconeogenic flux, the ratio of the rate of gluconeogenesis to the rate of glycolysis, was estimated from the 13C distribution in trehalose arising by gluconeogenesis and the 13C enrichment of alanine due to pyruvate cycling. Insects grew well on carbohydrate free diets and growth increased with increasing dietary protein level. At all dietary protein levels, larvae grew better on diets with fat. Without dietary fat, larvae were glucogenic but displayed low blood trehalose concentrations, <30 mM, regardless of protein consumption. When fat was included in the diet, however, gluconeogenic flux and blood trehalose level increased sharply in response to increased dietary protein level, with trehalose concentrations >50 mM at higher levels of protein consumption. When offered a choice of a high carbohydrate and a high protein diet, larvae maintained on diets with fat displayed a food preference related to blood sugar level. Those with low blood sugar fed on carbohydrate, while those with high blood sugar preferred protein. Trehalose synthesized from (2-13C)pyruvate exhibited asymmetry in the 13C distribution in individual glucose molecules, indicating a disequilibrium at the triose phosphate isomerase-catalyzed step of the gluconeogenic pathway. In trehalose from larvae on diets with fat, the asymmetric 13C distribution was higher than in trehalose from insects on diets lacking fat. This may partially result from isotopic disequilibrium when unenriched glycerol is metabolized to dihydroxyacetone phosphate following fat hydrolysis. The asymmetry in 13C distribution, however, also occurred in insects on diets without fat and decreased with increased gluconeogenic flux suggesting that true disequilibrium between the triose phosphates is the principal reason for the asymmetry. PMID- 15279895 TI - Inhibition of metalloproteinase activity by fruit extracts. AB - While the metalloproteinase enzymes are essential for development and remodeling of tissues, aberrant over expression of these enzymes contributes to several pathologic conditions. In particular, metalloproteinase over expression in cancer plays a significant role in metastasis by providing a mechanism for invasion and spread. The data presented here indicate that water extracts of raspberries, blackberries and muscadine grapes inhibit the activities of metalloproteinases 2 and 9. This inhibition could contribute to the suppression of carcinogenesis by diets high in fruit content. PMID- 15279894 TI - DNA hypomethylation induced by non-genotoxic carcinogens in mouse and rat colon. AB - The ability of non-genotoxic colon carcinogens to induce DNA hypomethylation was evaluated. Administering 0, 0.2 and 0.4 mg/kg of 5-aza-2'-deoxycytidine to female mice for 5 days resulted in a dose-related decrease in 5-methylcytosine in colon DNA. Rutin (3.0 mg/kg) and five bile acids (4.0 mg/kg) were administered in the diet to male F344 rats for 14 days. Rutin and four bile acids that promote colon cancer, deoxycholic acid, chenodeoxycholic acid, cholic acid and lithocholic acid caused DNA hypomethylation, while ursodeoxycholic acid that prevents colon cancer did not. Bromodichloromethane (BDCM) was administered to male F344 rats and B6C3F1 mice by gavage at 0, 50 and 100 mg/kg or in their drinking water at 0, 350 and 700 mg/l for up to 28 days. In rats, BDCM decreased DNA methylation, being more effective when administered by gavage, correlating to its greater carcinogenic potency by this route. In mice, BDCM did not decrease DNA methylation, corresponding to its lack of carcinogenic activity in the colon of this species. In summary, the ability of non-genotoxic colon carcinogens to cause DNA hypomethylation correlated with their carcinogenic activity in the colon. PMID- 15279896 TI - p-Nonylphenol pretreatment during the late neonatal period has no effect on 3,2' dimethyl-4-aminobiphenyl-induced prostate carcinogenesis in male F344 rats. AB - The modifying effects of late neonatal administration of p-nonylphenol (NP), a suspected xenoestrogen, on 3,2'-dimethyl-4-aminobiphenyl (DMAB)-induced prostatic carcinogenesis were investigated in male F344 rats. Three-week-old rats received 25, 250 or 2000 ppm of NP in the diet for 3 weeks prior to DMAB treatment and were sacrificed at 67 weeks of age for histopathological assessment of lesions and Ki-67 immunohistochemical analysis of cell cycle kinetics. Dietary administration of NP during the sexually immature period had no effects on maturation of male sex organs. Incidence, multiplicity and areas of neoplastic lesions in the prostate and seminal vesicles, and Ki-67 labeling indices in normal-looking epithelium were not significantly different among the experimental groups. These results indicate that late neonatal treatment with NP has no modulating effects on DMAB-induced rat prostatic carcinogenesis. PMID- 15279897 TI - Magnetite nanoparticle-loaded anti-HER2 immunoliposomes for combination of antibody therapy with hyperthermia. AB - Anti-HER2 antibody can induce antitumor responses, and can be used in delivering drugs to HER2-overexpressing cancer. Previously, we produced hyperthermia using magnetite nanoparticles that generate heat in an alternating magnetic field. In the present study, we constructed anti-HER2 immunoliposomes containing magnetite nanoparticles, which act as tumor-targeting vehicles, combining anti-HER2 antibody therapy with hyperthermia. The magnetite nanoparticle-loaded anti-HER2 immunoliposomes exerted HER2-mediated antiproliferative effects on SKBr3 breast cancer cells in vitro. Moreover, 60% of magnetite nanoparticles were incorporated into SKBr3, and the cells were then heated at 42.5 degrees C under an alternating magnetic field, resulting in strong cytotoxic effects. These results suggest that this novel therapeutic tool is applicable to treatment of HER2-overexpressing cancer. PMID- 15279898 TI - Theanine, a specific glutamate derivative in green tea, reduces the adverse reactions of doxorubicin by changing the glutathione level. AB - We previously showed that theanine, a specific glutamate derivative in green tea, decreased doxorubicin (DOX)-induced adverse reactions such as the induction of the lipid peroxide level and the reduction of glutathione peroxidase activity in normal tissues. In order to clarify how theanine attenuates the adverse reactions of DOX, we have focused on the effects of theanine on glutamate and glutathione (GSH) levels in normal tissues. The administration of theanine to mice increased the glutamate concentration in the liver and heart, and not in tumors. In vitro examinations indicated that theanine was metabolized to glutamate mainly in the liver. Moreover, theanine inhibited GSH reduction induced by DOX in the liver and heart. Therefore, these results suggested that theanine attenuated the DOX induced adverse reactions involved in oxidative damage, due to increase in glutamate and the recovery of GSH levels in normal tissues. PMID- 15279899 TI - C-terminal domain of human CAP18 antimicrobial peptide induces apoptosis in oral squamous cell carcinoma SAS-H1 cells. AB - Mammalian myeloid and epithelial cells express many antimicrobiotic peptides that contribute to innate host defense against invading microbes. In the present study, a 27-mer peptide of the C-terminal domain (hCAP18(109-135)) and analogs of the antimicrobial peptide human cathelicidin LL-37/human cationic antimicrobial protein 18 (hCAP18) were examined for tumoricidal activity. In vitro results showed that hCAP18(109-135) induced apoptosis in human oral squamous cell carcinoma (OSCC), SAS-H1 cells. The hCAP18(109-135) induced mitochondrial depolarization and apoptosis in SAS-H1 cells, but not in healthy human gingival fibroblasts (HGF) and human keratinocyte line HaCaT cells. Caspases were not activated during hCAP18(109-135)-induced apoptosis in SAS-H1 cells. The results indicate that hCAP18(109-135) may induce caspase-independent apoptosis in OSCC but not in normal cells. hCAP18(109-135) can therefore be a useful anti-tumor therapeutic agent in the treatment of OSCC. PMID- 15279900 TI - Decreased expression of the seven ARP2/3 complex genes in human gastric cancers. AB - The Arp2/3 complex and filamins play important roles in organization of actin cytoskeleton, and thus in cellular morphology and locomotion. We recently identified decreased expression of a gene for one of seven subunits of the Arp2/3 complex, the p41-Arc gene, and silencing of a filamin gene, the FLNc gene, in human gastric cancers. In this study, gene expressions of the seven subunits of the Arp2/3 complex, including p41-Arc, and their methylation statuses were analyzed in human gastric cancers. Quantitative real-time RT-PCR analysis of 32 primary gastric cancer samples and eight gastric cancer cell lines revealed that expressions of all the seven genes were significantly decreased. All the 32 primary cancer samples showed decreased expression of at least one subunit, and 25 samples showed decreased expressions of four or more of the seven subunits. Methylation-specific PCR analysis showed that none of the CpG islands in the 5' regions of the six genes other than p41-Arc were methylated in primary gastric cancers or cell lines. The consistent decrease of the Arp2/3 complex genes and its important role in actin organization suggested that the decrease could be involved in cancer phenotypes, such as dysplastic morphology. PMID- 15279901 TI - A methylated oligonucleotide induced methylation of GSTP1 promoter and suppressed its expression in A549 lung adenocarcinoma cells. AB - Glutathione S-transferase P1 (GSTP1) belongs to xenobiotic enzymes, and is supposed to contribute to chemoresistance. Though it was reported that GSTP1 gene is suppressed by cytosine-guanine (CpG) island methylation of its promoter, this promoter is not strongly methylated and GSTP1 protein is highly expressed in lung cancer. We intended to induce methylation of GSTP1 CpG island by using a methylated sense oligonucleotide complementary to this region. When we transduced the methylated oligonucleotides to A549 lung adenocarcinoma cells, methylation of the GSTP1 promoter and reduction of GSTP1 expression was induced, cell viability was reduced; however, chemoresistance against cisplatin has not clearly changed. PMID- 15279902 TI - Hydrogen peroxide negatively modulates Wnt signaling through downregulation of beta-catenin. AB - The Wnt signal transduction pathway plays an important role in organogenesis and carcinogenesis. In an effort to better understand the action of oxidative stress induced cellular signaling, we investigate the effect of exogenous H2O2 on the Wnt signal pathway. H2O2 decreases the amount of nuclear beta-catenin and Tcf/Lef dependent transcription. Overexpression of Dvl-1 abrogated H2O2-induced downregulation of beta-catenin. Pretreatment with LiCl or Wnt-3a conditioned medium completely inhibited H2O2-induced release of mitochondrial cytochrome c and DNA fragmentation. These results suggest that H2O2 negatively modulates the Wnt signal pathway through downregulation of beta-catenin. PMID- 15279903 TI - Galectin-3 expression in non-small cell lung carcinoma. AB - Galectins are a family of animal beta-galactoside-binding lectins involved in malignant transformation and progression. The present study evaluated the immunohistochemical expression of galectin-3 in a consecutive series of 81 radically resected non-small cell lung carcinomas (NSCLCs). The main pattern of galectin-3 expression was cytoplasmic (median percentage of cells with cytoplasmic positivity: 80.0%). The median percentage of tumor cells with nuclear and cytoplasmic co-expression of galectin-3 was 3.5%. No cases with exclusive nuclear immunostaining were observed. Functional interaction between galectin-3 and the thyroid transcription factor-1 (TTF-1) was previously demonstrated by cotransfection experiments. In the present study, concomitant expression of nuclear galectin-3 and TTF-1 was independently associated with a worse clinical outcome (HR 2.0; P = 0.01). PMID- 15279904 TI - Microsatellite instability in esophageal adenocarcinoma. AB - The frequency of microsatellite instability (MSI), a result of defective mismatch repair during DNA replication, has been reported inconsistently in primary esophageal adenocarcinoma (EADC). Using a panel of 15 markers, the primary aim of this study was to analyze the frequency of MSI in a well-characterized series of 27 primary EADCs, defined according to strict clinicopathologic criteria. Polymerase chain reaction was used to amplify the following microsatellite repeat loci: D2S123, D10S197, D2S119, D11S904, D2S147, D3S1764, D7S1830, D7S1805, D2S434, D9S299, BAT25, BAT26, D5S346, D17S250, and TGF-beta-RII. Tumors were classified as microsatellite-stable (MSS) when no alterations were seen in tumor DNA compared to matched normal tissues, low-level MSI (MSI-L) when 1-5 of 15 markers were altered, and high-level MSI (MSI-H) when more than five markers were altered. Using these stringent criteria, 9/27 (33%) tumors were MSS, 18/27 (67%) tumors were MSI-L, and no tumor was MSI-H. Immunohistochemistry demonstrated cell nuclear expression of DNA mismatch repair proteins (both hMLH1 and hMSH2) in 78% (21/27) of tumors. No associations were seen between MSI and immunohistochemical expression of hMLH1, hMSH2, alterations in p53 or MBD4, tumor grade, pathologic stage, or patient survival. In conclusion, the finding of low levels of MSI in most tumors suggests an inherent baseline genomic instability, and potentially increased susceptibility to mutations during the progression of esophageal adenocarcinoma. PMID- 15279905 TI - Reduced expression of liver-intestine cadherin is associated with progression and lymph node metastasis of human colorectal carcinoma. AB - Liver-intestine cadherin (LI-cadherin) is a recently identified member of the cadherin superfamily. We examined LI-cadherin expression in four human colorectal carcinoma cell lines and 45 human primary colorectal carcinomas using a monoclonal antibody against LI-cadherin. We also investigated the correlation between LI-cadherin expression and clinicopathologic parameters. Among the cell lines, LI-cadherin expression was detected in a differentiated phenotype, but not in dedifferentiated phenotypes. Among the 45 tumor samples, LI-cadherin expression was preserved in 28 (62%) and reduced in 17 (38%). Reduced LI-cadherin expression was significantly associated with a high tumor grade (P = 0.015), lymphatic invasion (P = 0.033), lymph node metastasis (P = 0.015), and an advanced pTNM stage (P = 0.033). These results suggest that analysis of LI cadherin expression may help to indicate the biological aggressiveness of this malignancy. PMID- 15279906 TI - Isolation and characterization of the 5'-flanking region of the growth hormone secretagogue receptor gene from black seabream Acanthopagrus schlegeli. AB - Ghrelin, the recently discovered endogenous ligand for growth hormone secretagogue receptor (GHSR), is widely expressed and involved in regulating diverse physiological functions in addition to stimulation of growth hormone (GH) secretion. Previous studies have demonstrated the functional significance of the ghrelin/GHSR system, yet the transcriptional regulation of the ghrelin and GHSR genes are poorly understood. We have recently cloned the GHSR cDNA from the pituitary of black seabream Acanthopagrus schlegeli. In the present study, we have isolated a 2.1 kb 5'-flanking region of the GHSR gene from the same species and have investigated, for the first time, the transcriptional regulation of GHSR from a non-human species. The 5'-flanking region of the seabream GHSR gene was found to contain a number of unique putative transcription factor-binding sites different from the human counterpart. Functional characterization of the 5' flanking region in several cell lines indicates that the region between -1423 and +19 contains sufficient elements for promoter function. Moreover, progressive 3' deletion analysis suggests the presence of negative regulatory element(s) and essential cis-acting element(s) at -514/+19 and -928/-515, respectively. Furthermore, we have shown that the promoter activity is significantly enhanced by a GHSR agonist in a cell line stably expressing the seabream GHSR, and this stimulatory effect could be completely blocked by a GHSR antagonist. These results suggest that homologous up-regulation plays an important role in the transcriptional control of the teleostean GHSR gene. This is in big contrast to the human situation in which a homologous down-regulation of the GHSR gene transcription by its own ligand has been previously demonstrated. PMID- 15279907 TI - Functional significance of the BBXXB motif reversed present in the cytoplasmic domains of the human follicle-stimulating hormone receptor. AB - The minimal structural motif, BBXXB (where B represents a basic amino acid residue and X a non-basic residue), located in particular regions of the intracellular domains of cell surface membrane receptors is involved in the G protein-activating activity of a number of G protein-coupled receptors. The human FSH receptor (hFSHR) exhibits a reversed BBXXB motif (BXXBB) in the juxtamembrane region of the third intracellular loop (IL3) and the carboxyl terminus (Ctail) of the receptor; however the importance of this sequence on receptor function remains unclear. In the present study, we analyzed the effects of mutations in this structural motif on hFSHR expression, receptor-mediated effector activation and agonist-provoked receptor internalization. Human embryonic kidney 293 cells were transiently transfected with plasmids containing the cDNA of the wild-type (Wt) hFSHR or several hFSHR mutants in which basic amino acids of the minimal structural motif at the IL3 and Ctail were replaced with alanine (i.e. AXXAA, AXXBB, BXXAB and BXXBA mutants). Alanine substitution of the three basic residues present in the IL3-BXXBB (IL3-AXXAA mutant) yielded a < or =60 kDa possibly under glycosylated form of the FSHR, whereas the same substitutions in the Ctail resulted in the immature >62 kDa form of the receptor; both AXXAA hFSHR mutants completely failed to bind agonist and activate effector. Individual substitutions resulted in different cAMP responses to agonist stimulation: the IL3-AXXBB and IL3-BXXBA mutant hFSHRs failed to evoke Gs protein activation, whereas agonist stimulated cAMP production was completely normal when the IL3-BXXAB mutant was expressed. All three IL3 mutants bound [125I]-labelled FSH in a similar fashion to the Wt hFSHR. Ligand-binding, cell surface membrane receptor expression and agonist-provoked effector activation were significantly affected by the individual substitutions at the Ctail-BXXBB motif: the Ctail-AXXBB variant exhibited reduced (approximately 50%) maximal cAMP response and ability to bind ligand, whereas both ligand binding and effector activation was severely reduced or abolished by expression of the Ctail-BXXBA and -BXXAB hFSHR mutants; the expression levels of the 80 kDa form of the receptor correlated with the magnitude of ligand-provoked cAMP production and binding capability of the mutant receptors. Upon stimulation by agonist, all mutants with detectable ligand binding activity internalized following the pattern exhibited by the Wt hFSHR species. These results indicate that the BXXBB motif at the IL3 of the hFSHR is essential for coupling the activated receptor to the Gs protein, whereas the same motif in the Ctail is apparently more important for membrane expression. PMID- 15279908 TI - Estrogen and phytoestrogen regulate the mRNA expression of adrenomedullin and adrenomedullin receptor components in the rat uterus. AB - The serum estrogen surge in the uterus triggers precisely-timed physiological and biochemical responses required establishing and maintaining pregnancy. Previous reports have shown that consumption of phytoestrogen-containing plants may disrupt the precise control of pregnancy. To evaluate the effects of phytoestrogens in the uterus, we screened for estradiol (E2)-inducible genes in immature rat uteri. We identified the gene for receptor-activity-modifying protein 2 (Ramp2), known to be a component of the adrenomedullin (ADM) receptor, as responsive to both E2 and the phytoestrogen coumestrol (Cou). We further examined the expression of ADM and ADM signaling components Ramp2, Ramp3, and CRLR in the immature rat uterus and found that both E2 and Cou regulated these genes expression. In addition, treatment with ADM increased uterine weight and edema similar to that observed after Cou treatment. Our findings indicated that the phytoestrogen caused the abnormal induction of vasoactive factors in the uterus. PMID- 15279909 TI - Activation of second messenger-dependent protein kinases induces muscarinic acetylcholine receptor desensitization in rat thyroid epithelial cells. AB - Internalization and phosphorylation of G protein-coupled receptors (GPCR) are considered two important regulatory events of receptor signal transduction. In Fischer rat thyroid (FRT) epithelial cells, we have shown that muscarinic acetylcholine receptor (mAChR) stimulation induces intracellular Ca2+ mobilization via Ca2+ store release, capacitative Ca2+ entry and voltage dependent Ca2+ channels activation. In the present study, the role of mAChR internalization and phosphorylation on receptor signalling pathway was examined by means of intracellular Ca2+ measurement in these cells. Exposure of FRT cells to carbachol (Cch), a mAChR agonist, resulted in a desensitization of receptor mediated intracellular Ca2+ mobilization and induced the internalization of constitutively expressed mAChR in this cell type. Treatment of FRT cells with hypertonic sucrose, which markedly reduced agonist-receptor complex internalization, or phenylarsine oxide (PAO) diminished the Cch-induced intracellular Ca2+ response. Moreover, pretreatment of cells with phorbol-12 myristate-13-acetate (PMA), an activator of protein kinase C (PKC), completely abolished Cch-evoked Ca2+ mobilization, whereas it was significantly increased by the preincubation of cells with GF109203X, a selective inhibitor of PKC. We also found a marked decrease on Cch-stimulated Ca2+ mobilization in pretreated FRT cells with forskolin, an activator of protein kinase A (PKA), but the preincubation of cells with genistein, an inhibitor of protein tyrosine kinases, had no effect on Ca2+ mobilization induced by Cch. These findings seem to indicate that mAChR in FRT cells exhibit a desensitization, which may be mediated, at least in part, through activation of second messenger-dependent protein kinases and that receptor internalization could be necessary for signalling. PMID- 15279910 TI - Expression of 17beta- and 3beta-hydroxysteroid dehydrogenases and steroidogenic acute regulatory protein in non-luteinizing bovine granulosa cells in vitro. AB - Granulosa cells of small follicles differentiate in vitro in serum-free medium, resulting in increased estradiol secretion and abundance of mRNA encoding cytochrome P450aromatase (P450arom). We tested the hypothesis that differentiation in vitro also involves increased expression of 3beta- and 17beta hydroxysteroid dehydrogenases (HSD) in the absence of steroidogenic acute regulatory protein (StAR) expression, as has been observed in vivo. Granulosa cells from small (<6 mm diameter) follicles were cultured for up to 6 days, and mRNA levels quantified by Northern hybridization or RT-PCR. Estradiol and progesterone concentrations in medium increased with time in culture, as did mRNA encoding P450arom, 3beta- and 17beta-HSD but not P450scc. Both P450arom and 17beta-HSD were significantly correlated with estradiol accumulation in culture medium. Progesterone secretion was correlated with 3beta-HSD but not P450scc mRNA levels. StAR mRNA was detectable by RT-PCR, did not change with duration of culture and was not correlated with progesterone secretion. FSH significantly stimulated P450arom and 17beta-HSD mRNA levels. Cell origin (from the antral or the basal layer of the membrana granulosa) did not affect steroidogenesis. We conclude that under the present cell culture system granulosa cells do not luteinize, and show expression of key steroidogenic enzymes in patterns similar to those occurring in differentiating follicles in vivo. Further, the data suggest that 17beta-HSD may be as important as P450arom in regulating estradiol secretion, and that 3beta-HSD is more important than P450scc as a regulator of progesterone secretion in non-luteinizing granulosa cells. PMID- 15279911 TI - Differential expression of adenylyl cyclase subtypes in human cardiovascular system. AB - In human myocardium, beta1-adrenoceptor stimulation achieves maximal inotropic response but less than 50% of maximal adenylyl cyclase activation, whereas the reverse is true of the beta2-adrenoceptor. Four types of adenylyl cyclase, type IV-VII, have been described in mammalian heart, but their expression and relative distribution in human heart and blood vessels is not known. We found that type IV, V, VI and VII adenylyl cyclases were all expressed in cardiomyocytes. Whereas types IV and VII RNA were more abundant in extra-cardiac than cardiac tissues, both absolute and relative expression of type VI was greatest in heart, and lower in tissues lacking a beta1-adrenoceptor. Type V expression was virtually confined to atrium. In situ mRNA hybridisation showed that the beta1-adrenoceptor co localised with type VI adenylyl cyclase but not other subtypes in juxtoglomerular cells of human kidney. The tissue specific expression of these adenylyl cyclase subtypes may favour its coupling to corresponding receptors expressed in the given tissue type. PMID- 15279912 TI - Cloning, characterisation, and expression of three oestrogen receptors (ERalpha, ERbeta1 and ERbeta2) in the European sea bass, Dicentrarchus labrax. AB - Three oestrogen receptor [ER] subtypes have been described in teleost fish, namely ERalpha, and two ERbeta subtypes, called ERbeta1 and ERbeta2 (or ERbeta and ERgamma in Atlantic croaker). Their expression during embryonic development and gonadal growth has evoked interest in their potential role in sexual differentiation and gonadal development in fish. We cloned three oestrogen receptors from adult liver (sb-ERalpha cDNA) and ovary (partial sb-ERbeta1 and sb ERbeta2 cDNAs) of the European sea bass, and according to their phylogenetic relatedness to other ERs in teleosts, named them sea bass [sb-] ERalpha, ERbeta1 and ERbeta2. Deduced amino acid numbers for sb-ERalpha, sb-ERbeta1 and sb-ERbeta2 were 639, 517 and 608, respectively, representing in the case of sb-ERbeta1 and sb-ERbeta2 about 90% of the open reading frame. Highest amino acid identities were found for sb-ERalpha with eelpout ERalpha (88.7%), for sb-ERbeta1 with Atlantic croaker ERgamma (85.8%), and for sb-ERbeta2 with Atlantic croaker ERbeta (90.1%). Southern analysis confirmed that all three sea bass oestrogen receptors (sb-ERs) are the products of three distinct genes. In adult sea bass, ERalpha was predominantly expressed in liver and pituitary, while sb-ERbeta1 and sb-ERbeta2 were more ubiquitously expressed, with highest expression levels in pituitary. In a mixed-sex population of juvenile sea bass, sb-ERalpha expression was significantly elevated in gonads at 200 days posthatch (dph), while for sb ERbeta1 and sb-ERbeta2 highest expression levels were observed in gonads at 250 dph. For sb-ERbeta2, expression was also significantly higher in the brain at 250 dph. The cloning of these three ER subtypes in the European sea bass together with the results obtained on expression levels in adult and juvenile animals has given us the foundation to investigate their possible role in sexual differentiation and development in this species in future studies. PMID- 15279913 TI - Comparative analysis and characterization of mutated thyroid peroxidases with disturbance expressed on the cell surface. AB - Five mutated thyroid peroxidases (TPO) with varying degrees of disturbance in cell surface expression, probably owing to misfolding, were comparatively analyzed. CHO-K1 cells transfected with these mutated mRNAs expressed TPO protein in 65.6-82.1% of cells in antibody staining, and the TPOs were located in intracellular structures like the nuclear envelope and ER as well as cytoplasmically like wild-type TPO. When cell surface expression was examined, three mutated TPOs, G533C-, D574/L575del-, and G771R-TPOs, were expressed to varying degrees. In contrast, R175Q- and R665W-TPOs were thought not to be expressed on the cell surface, although a vague increment in R175Q-TPO was observed with increasing amounts of mRNA. In the kinetic study, three mutated TPOs having insufficient expression on the cell surface showed delays in decrease at 4 and 8 h after chase, although between 8 and 24 h after chase they decreased rapidly, as did the two other mutated TPOs. In immunoprecipitation by anti-TPO antibody, G533C-, D574/L575del-, and G771R-TPOs exhibited increasing interaction with calnexin. The combined evidence suggested that some of the mutated TPOs with disturbance in cell surface expression, probably owing to misfolding, exhibited the delay in kinetics of newly synthesized protein as a result of increasing interaction with calnexin and that such TPOs could be expressed to some extent on the cell surface. PMID- 15279914 TI - Nitric oxide influences the maturation of cumulus cell-enclosed mouse oocytes cultured in spontaneous maturation medium and hypoxanthine-supplemented medium through different signaling pathways. AB - Nitric oxide (NO) has been recently shown to act with a dual action in mouse oocyte meiotic maturation depending on its concentration, but the mechanism(s) through which it influences oocyte maturation has not been fully clarified to date. The purpose of this study was to test the hypothesis that different signaling mechanisms exist for NO-stimulated and NO-inhibited in vitro maturation of meiosis in cumulus cell-enclosed oocytes (CEOs) from PMSG-primed immature female mice. CEOs were cultured in both spontaneous maturation model and hypoxanthine (HX) arrested model to investigate the mechanism(s). Sodium nitroprusside (SNP, an NO donor) at a concentration of 1mM delayed significantly germinal vesical breakdown (GVBD) during the first 5 h of incubation period and further inhibited the formation of first polar body (PB1) at the end of 24 h of incubation. While SNP, at a concentration of 10 microM, stimulated significantly the meiotic maturation of oocytes by overcoming the inhibition of HX. Methinine blue (MB, 10 microM) or 1-H-[1,2,4] oxadiazolo-[4,3-a]quinoxalin-1-one (ODQ, 10 microM)), two soluble guanylate cyclase (sGC) inhibitors, could reverse SNP inhibited spontaneous oocyte maturation, but had no effect on SNP-stimulated meiotic maturation in the presence of HX. 8-Br-cGMP (1mM), a cell-permeating cGMP analogue, demonstrated a significant inhibitory effect on both spontaneous meiotic maturation and HX-arrested meiotic maturation. The delay effect of SNP on GVBD occurrence was similar to that of forskolin (6 microM, an adenylate cyclase stimulator) and rolipram (250 microM, a phosphodiesterase 4 inhibitor), two cAMP elevating reagents. Both forskolin and rolipram reversed significantly the SNP stimulated meiotic maturation, but did not reverse the SNP-inhibited spontaneous meiotic maturation. Cilostamide (1 microM), the selective inhibitor of phosphodiestrase 3 (PDE3), could mimic the inhibitory effect of HX on the spontaneous meiotic maturation in CEOs and this inhibitory effect could also be reversed by SNP (10 microM). Moreover, sphingosine (3 microM), a protein kinase C (PKC) inhibitor, blocked the SNP-inhibited spontaneous meiotic maturation, but did not block the SNP-stimulated meiotic maturation. Clearly, these results suggest that pathway differences are present between SNP-inhibited spontaneous meiotic maturation and SNP-stimulated meiotic maturation of mouse oocytes. PMID- 15279915 TI - Interactive video behavioral intervention to reduce adolescent females' STD risk: a randomized controlled trial. AB - A longitudinal randomized design was used to evaluate the impact of a theoretically based, stand-alone interactive video intervention on 300 urban adolescent girls' (a) knowledge about sexually transmitted diseases (STDs), (b) self-reported sexual risk behavior, and (c) STD acquisition. It was compared to two controls, representing high-quality informational interventions. One used the same content in book form; the other used commercially available brochures. Following randomization, the interventions were administered at baseline, with booster sessions at 1, 3, and 6 months. Self-reports revealed that those assigned to the interactive video were significantly more likely to be abstinent in the first 3 months following initial exposure to the intervention, and experienced fewer condom failures in the following 3 months, compared to controls. Six months after enrollment, participants in the video condition were significantly less likely to report having been diagnosed with an STD. A non-significant trend in data from a clinical PCR assay of Chlamydia trachomatis was consistent with that finding. PMID- 15279916 TI - Unmet need for contraception in Kuwait: issues for health care providers. AB - Based on a nationally representive household survey of Kuwaiti women held in 1999 (n = 1502) unmet need for contraception was analyzed in Kuwait, an oil-rich Muslim country. It was found that 9.7% currently married women had an unmet need for contraception. Of those, 6.1% wanted to stop child bearing, while 3.6% wanted to space their children. A bivariate comparison of the women with unmet need and current contraceptors showed that the unmet need group comprised of relatively older women with a significantly higher level of parity and ones where husband or wife disapproved of contraception. Also, larger percentages of the unmet need group belonged to relatively lower socio-economic status and were Bedouins. Among the reasons for current non-use, two-third believed that they had a low risk of pregnancy due to infrequent sexual activity or sub-fecundity, and 22% were not using a method because of health concerns. A significantly larger percentage of the unmet need group disapproved of contraception, and believed that Islam forbids family planning, compared to current users (30% and 15%, respectively). The logistic regression analysis showed that the wife's perception of the husband's disapproval of contraceptive use had the strongest negative association with unmet need. We conclude that the contraceptive needs of about 90% of all non pregnant currently married women who wanted to delay or limit children were being met adequately despite the absence of a formal family planning program, while about 10% women had an unmet need. Issues for health care providers are discussed and family planning counseling is recommended for higher risk older women with unmet need. PMID- 15279917 TI - Transactional sex among women in Soweto, South Africa: prevalence, risk factors and association with HIV infection. AB - Sex workers have long been considered a high-risk group for HIV infection, but to date little quantitative research has explored the association between HIV risk and exchange of sex for material gain by women in the general population. The objective of this study was to estimate the prevalence of such transactional sex among women attending antenatal clinics in Soweto, South Africa, to identify demographic and social variables associated with reporting transactional sex, and to determine the association between transactional sex and HIV serostatus. We conducted a cross-sectional study of women seeking antenatal care in four Soweto health centres who accepted routine antenatal HIV testing. Private face-to-face interviews covered socio-demographics, sexual history and experience of gender based violence. 21.1% of participants reported having ever had sex with a non primary male partner in exchange for material goods or money. Women who reported past experience of violence by male intimate partners, problematic substance use, urban residence, ever earning money, or living in substandard housing were more likely to report transactional sex, while women who reported delayed first coitus, were married, or had a post-secondary education were less likely to report transactional sex. Transactional sex was associated with HIV seropositivity after controlling for lifetime number of male sex partners and length of time a woman had been sexually active (OR = 1.54, 95% CI: 1.07, 2.21). Women who reported non-primary partners without transactional sex did not have increased odds of being HIV seropositive (OR = 1.04, 95% CI: 0.75, 1.43). We conclude that transactional sex may place women at increased risk for HIV, and is associated with gender-based violence, substance use and socio-economic disadvantage. Research, policy and programmatic initiatives should consider the role of transactional sex in women's HIV risk, with attention to the intersecting roles of violence, poverty, and substance use in shaping women's sexual behaviour. PMID- 15279918 TI - What constitutes success in preventive health care? A case study in assessing the benefits of hip protectors. AB - The economic success of preventive health programs is typically assessed by the net health-related utility gain or loss to society relative to the cost. Issues relating to the positive or negative utility associated with participating in a preventive health program are often ignored. However, it is likely that calls for informed consumer choice and respect for patient autonomy will provide an impetus to examine utility associated with the process and outcomes of preventive health programs. In this paper, we outline the nature of the ex ante and ex post perspective in evaluating benefits and the presence of process utility and the utility of gambling in individual's utility function for preventive health care. The implications of including process attributes and psychological states when assessing benefits to society are discussed in relation to an empirical study on the value of external hip protectors for the prevention of hip fractures. We demonstrate that wearing hip protectors and the psychological outcomes of being a participant in the program can have a significant impact on individual's assessment of the benefits. Furthermore, point of reference plays a crucial role in their valuation. Individuals who did not consent to participate in a trial of hip protectors valued all states significantly lower than those who did participate in the trial. We argue that the utility associated with adherence to the intervention is an important issue for preventive health policy. From the viewpoint of applied welfare economics, evaluation of preventive health programs should allow for both process and outcome utility when assessing benefits. In this context, success might be viewed as maximising the opportunity for individuals to make an informed choice. PMID- 15279919 TI - 'In the eye of the beholder': perceptions of local impact in English Health Action Zones. AB - Contemporary efforts to promote population health improvement and to reduce inequalities in the UK are characterised by their complexity as they engage with a multiplicity of agencies and sectors. Additionally, the emphasis on promoting evidence-based practice has challenged evaluators tasked with collecting and interpreting evidence of impact in complex local health economies. National policy makers, local implementers and other stakeholders will have varying perspectives on impact and the Labour Government's centralising tendencies have acted to 'crowd out' local voices from the policy process. Drawing on the national evaluation of Health Action Zones (HAZ) this article 'gives voice' to local stakeholders and their perceptions of impact. Informed by a Theories of Change perspective, we explore HAZ interventions to articulate the nature of impact and its limits. We analyse the claims made by local HAZs with reference to the evidence base and examine their significance in the context of overall HAZ objectives. We conclude that local implementer perspectives are no less sophisticated than those at the policy centre of central government, but that they are informed by three important factors: the local context, a need to be pragmatic and the limited potency of evidence in the public policy system. PMID- 15279920 TI - Insights into the 'healthy immigrant effect': health status and health service use of immigrants to Canada. AB - This paper combines multiple cross-sections of data drawn from the National Population Health Survey and Canadian Community Health Survey to confirm the existence of the 'healthy immigrant effect', specifically that immigrants are in relatively better health on arrival in Canada compared to native-born Canadians, and that immigrant health converges with years in Canada to native-born levels. The paper finds robust evidence that the healthy immigrant effect is present for the incidence of chronic conditions for both men and women, and results in relatively slow convergence to native-born levels. There is only weak evidence in terms of self-assessed health status. The inclusion of controls for region of origin and year of arrival does not account for the observed effects, although region of origin is an important determinants of immigrant health. The paper then considers some alternative explanations for the observed differences, and support is found for the idea that the healthy immigrant effect reflects convergence in physical health rather than convergence in screening and detection of existing health problems. PMID- 15279921 TI - Socioeconomic, cultural, and personal influences on health outcomes in low income Mexican-origin individuals in Texas. AB - Several studies have suggested that the health of Mexican-Americans is better than expected given their low socioeconomic status. The healthy migrant hypothesis and the acculturation hypothesis, stating that the foreign-born and the less acculturated enjoy better health, have been proposed as possible complementary explanations. However, it is not clear which are the socioeconomic, cultural, and personal characteristics that favor good health and that differentiate foreign-born from US-born and unacculturated from acculturated Mexicans. In this paper, we compare, by nativity and acculturation level, the socioeconomic, cultural, and personal characteristics in a sample of low income mostly female Mexican-origin individuals living in Texas and investigate their contribution to differences in self-reported physical health, mental health, and self-rated health (SRH) status. Using a multistage probability sample, we completed 1745 interviews with Mexican-origin individuals. The survey instrument included the SF-12, demographic and socioeconomic information, and questions on social support, religiosity, fear of victimization, trust, perceived racism, and perceived opportunity. Nativity and use of the Spanish language were combined into a nativity/acculturation variable. We estimated multivariate regressions and ordered logit regressions to investigate the association of health outcomes to nativity/acculturation and socioeconomic, cultural, and personal characteristics. Overall, the distribution of strengths (more social support, trust, perceived personal opportunities and less perceived victimization) reflected a nativity based income gradient and an education gradient reflecting language use. Health outcomes varied by nativity/acculturation after controlling for socioeconomic, cultural, and personal characteristics. Physical health differed by nativity, supporting the healthy migrant hypothesis, while nativity-based differences in mental health were explained by socioeconomic and personal characteristics. SRH varied by language use, suggesting a culturally conditioned response. The socioeconomic, cultural, and personal factors affected health outcomes differently. These findings suggest a complicated interaction between nativity, acculturation, and economic factors in determining social and personal strengths and their influences on health. PMID- 15279922 TI - Utilization of well-baby care visits provided by Taiwan's National Health Insurance Program. AB - In April of 1995, Taiwan's National Health Insurance Program started providing each eligible child a total of six well-baby care visits. The first four are for infancy, the fifth is for the second and the third years of life, and the sixth is for the fourth year. These services are in addition to neonatal screening and a series of primary immunizations that have been publicly financed and utilized conventionally for years. The purposes of this study were to investigate the utilization level of these well-baby care visits, and explore relevant factors. The results reveal that 36% of eligible children did not use any of the first four visits, 58% did not utilize the fifth, and 82% did not use the sixth in the late 1990s. It appears that the take-up of these services is much less than satisfactory. Maternal awareness of and attitudes toward the services appeared to be the most important factors influencing utilization. These two factors not only were most influential, but also significantly contributed to disparities in utilization among different regions and types of residential districts. As a result, they should be the focus of interventions for advancing well-baby care. While these two factors are at the individual level, they are not independent from the health care system because the health care system has impacts on individual factors. Since physicians can serve as a good vehicle for teaching parents about relevant information and correct attitudes, and most physicians in Taiwan complained about the payment scheme, offering stronger incentives for physicians to promote such services might be helpful for achieving a high utilization level of well-baby care. PMID- 15279923 TI - Committed to health for all? How the G7/G8 rate. AB - The G7/G8 group of nations dominate the world political and economic order. This article reports selected results from an investigation of the health implications of commitments made at the 1999, 2000 and 2001 Summits of the G7/G8, with special reference to the developing world. We emphasize commitments that relate to the socioeconomic determinants of health (primarily to reducing poverty and economic insecurity) and to the ability of national governments to make necessary basic investments in health systems, education and nutrition. We conclude that without a stronger commitment to redistributive policy measures on the part of the G7/G8, historic commitments on the part of the international community to providing health for all are likely not to be fulfilled. PMID- 15279924 TI - Research report: do general practitioners tell their patients about side effects to common treatments? AB - The principle of respect for patients' autonomy, or right to self-determination, has gained increasing importance in health care legislation during the last decade. To respect this principle the patients' informed consent to a proposed treatment is required. In relation to ordinary treatments in general practice, where several reasonable alternatives may be available and where non-treatment may be an acceptable alternative, this requirement is at least as strong as in other parts of the health care system. In this context, information about side effects may be crucial for the patient's decision to accept a proposed treatment or not. The aims of this study were to investigate the extent to which general practitioners in Denmark inform their patients about possible side effects without being asked when a common treatment is proposed. We also wished to examine the relation between physicians' estimation of the severity and frequency of these side effects, and their willingness to inform patients spontaneously as well as their preferred reasons for choosing to inform or not inform the patients. A questionnaire was sent to a random sample of 450 Danish general practitioners. The respondents differed considerably with regard to their willingness to inform patients about side effects but they were significantly more likely to give the information spontaneously if they considered the side effects frequent than when side effects were considered rare. In contrast, estimations of severity did not seem to be of any importance. The majority of the respondents informed their patients primarily to enable them to react appropriately to the side effects in question or to make sure that the patient would comply with the treatment. These findings indicate that the information given to patients about side effects by Danish general practitioners is not in accordance with the principle of respect for the patients' autonomy and not in accordance with the requirements of Danish legislation. PMID- 15279925 TI - Truth telling and truthfulness in the care for patients with advanced dementia: an ethnographic study in Dutch nursing homes. AB - The purpose of this study was to investigate and analyze the moral tension that exists in the care for demented nursing home patients, between the principle of respect for autonomy and the value that is attached to respect for the subjective world of the patient. To this end an ethnographical field study was carried out by two researchers in two Dutch nursing homes. Among the central topics that evolved were the different moral problems that nurses experience concerning truth telling and acting truthfully in relation to demented patients. In situations unrelated to the dementia and its diagnosis, the right to be informed is in principle respected, even if the information is sometimes painful. More specific questions of demented patients about their situation are a regular cause of embarrassment for their carers, who rely on various treatment strategies to deal with such questions. These strategies are often successful. However, when they fail, the nurses are faced with a problem they cannot solve, namely the loss of a common shared world and the resulting unmentionable truth about the diagnosis of dementia, as objective basis and legitimization for their approach to the demented patient. We conclude that in the training and professional support given to nurses, more attention should be paid to (awareness of) the moral problems that arise from this loss of a common shared world, so that they can react to the subjective world of demented patients without feeling that they are deceiving them. PMID- 15279926 TI - Relief, risk and renewal: mixed therapy regimens in an Australian suburb. AB - In recent decades, health care has been transformed by the proliferation of non biomedical therapies that are mostly unsupported by government-authorized systems of health care, and are referred to in the health social science literature by terms such as "alternative" or "complementary" medicine, or the acronym "CAM". From the perspective of users, who are often pluralistic and pragmatic in their orientation to healing modalities, "mixed therapy regimens" is a less dichotomized and more appropriate conceptualization of the process of seeking health care from diverse sources of expertise. This paper reports on an anthropological study of residents of an Australian suburb, where it was found that 24% of interviewees in a community-based sample reported they or members of their household had used non-biomedical therapies. It was found that users of mixed therapy regimens regard the felt effectiveness of therapies to be more important than legitimacy deriving from a professionalised scientific knowledge system. "Natural" therapies, variously defined (and including vitamins), are valued as a counterweight to forms of disability and malaise that are often associated with risks of modern life. Pharmaceuticals (especially prescription products), by contrast, were not infrequently subjected to radical critique and various forms of practical resistance. The contrast between pharmaceuticals and "natural" therapies in many residents' accounts points to the moral criteria that are an important dimension along which the effectiveness of care is evaluated. "Natural" remedies and mixed therapy regimens can also be interpreted as antidotes for the experience of living in a "risk society", part of a project of resistance to the hazards of modernity carried out at the site of the body. PMID- 15279927 TI - Street children in Pakistan: a situational analysis of social conditions and nutritional status. AB - This paper examines the social conditions and nutritional status of street children in Pakistan. Nutritional status is evaluated by an assessment of height and weight relative to age. A heterogeneous sample of 108 street children in the twin cities of Rawalpindi and Islamabad was studied. One hundred and one of them were children "on the street" having regular family contact; seven were "of the street", without any family contact. Most street children came from large families which had recently moved to the city in search of economic opportunities. Their parents had low education levels and were either unemployed or employed in unskilled occupations. Poverty clearly was an important factor. The majority of the children moved to the street to augment family income. Most of the children were males (81%) and the average age at beginning life on the street was under 10 years. The majority was working 8-12 h daily with an average income of Rs. 40-60 per day (USD 1 = Rs. 60). Important issues were parental exploitation, police harassment, abuse, and the impact of other street peers in their lives. The distribution of height-for-age relative to the National Center for Health Statistics (NCHS) reference standard showed that 20% were stunted and 12% had wasting. Wasting was equal between sexes, while fewer girls than boys were stunted. The study concludes that the issue of street children in Rawalpindi and Islamabad is mainly one of "children on the street," while "children of the street" are an exceptional phenomenon. It was noted that, with some exceptions, street children in other world regions share similar risk factors and backgrounds. Findings from this study will facilitate both the identification of high-risk families, i.e. those whose children are likely to take to the street, and timely preventive and rehabilitative measures. PMID- 15279928 TI - Job stress, social support, and prevalence of insomnia in a population of Japanese daytime workers. AB - To clarify the relationship between perceived job stress, social support and prevalence of insomnia in Japanese daytime workers, 1161 male white-collar employees of an electric equipment manufacturing company (age, 23-60 years, mean age of 37.0) were surveyed by means of a mailed questionnaire. Perceived job stress was evaluated with the Japanese version of the generic NIOSH job stress questionnaire. Insomnia was diagnosed if workers had at least 1 of 3 types of symptoms on an almost nightly basis. The symptoms were (1) taking more than 30 min to fall asleep (Difficulty Initiating Sleep, DIS), (2) difficulty in maintaining sleep (DMS), or (3) early morning awakening (EMA). The overall prevalence of insomnia was 23.6% and the prevalences of the three subtypes were 11.3% for DIS, 14.2% for DMS, and 1.9% for EMA. Workers with high intragroup conflict (OR 1.6), high job dissatisfaction (OR 1.5), and high symptoms of depression (OR 2.0) (CES-D > 16) had a significantly increased risk for insomnia after adjusting for multiple confounding factors. Low employment opportunities, physical environment and low coworker support also were weakly associated with risk for insomnia among workers. Furthermore, high depressive symptoms significantly increased the risk of DIS (OR 2.4). Therefore in white-collar male daytime workers, psychological job stress factors such as interpersonal conflicts with fellow employees, job satisfaction, and social support were independently associated with a modestly increased risk of insomnia that included three different subtypes that were considered to be defining for the disorder. PMID- 15279929 TI - The effect of hospital bed reduction on the use of beds: a comparative study of 10 European countries. AB - In Europe, the reduction of acute care hospital beds has been one of the measures implemented to restrict hospital expenditure. The aim of this study is to gain insight into the effect bed reductions have on the use of the remaining beds within different healthcare systems. We concentrated on two healthcare system elements: hospital financing system (per diem and global budget systems) and physician remuneration system (fee-for-service and salary systems). We also controlled for technological development and demand for healthcare. We used data from the OECD health data files of 10 North-Western European countries on hospital bed supply and use. The hospital bed indicators used were occupancy rate, average length of stay and admission rate. The data were analysed with multilevel analysis. We found some indication that the different financial incentives of hospital financing systems do indeed influence hospital bed use in the case of reductions in acute care hospital bed supply in different ways. However, we found significant effects only for the hospital bed use indicators "occupancy rate" and "admission rate". For physician financing systems, no significant effects were found. PMID- 15279930 TI - Arsenic contamination awareness among the rural residents in Bangladesh. AB - Arsenic poisoning of tubewell water, which constitutes the primary source of drinking water, has become the greatest health threat to the people of rural Bangladesh. Somewhere between 35 to 57 million people in the country are now suspected of being affected by drinking water contaminated with arsenic. While the Bangladesh government, non-government organizations (NGOs), and bilateral and multilateral assistance agencies are involved in combating this dreadful problem, all of their efforts to date have proceeded without having grassroots information about arsenic poisoning. The objectives of this study are to investigate the level of knowledge rural residents have regarding arsenic poisoning and to identify the correlates of that knowledge. Questionnaire surveys administered among residents of four rural areas in Bangladesh provided the major data source for this study. Twenty villages were selected from moderate and low arsenic risk regions and a total of 356 respondents, 177 from medium risk regions and 179 from low risk regions, were interviewed. Analysis of the survey data reveals that arsenic awareness is currently not widespread in the study villages, particularly in the low arsenic risk region. There are also gaps in arsenic knowledge regarding the diseases caused by arsenic poisoning and mitigating measures available to prevent contamination. This study identified arsenic risk region, level of education, gender, and age as important determinants of arsenic knowledge. The findings of this study will aid in making existing health education programs more effective and in reducing the risk of developing arsenic related illnesses. PMID- 15279931 TI - Does traditional birth attendant training improve referral of women with obstetric complications: a review of the evidence. AB - This narrative and meta-analytic review of the effectiveness of traditional birth attendant (TBA) training to improve access to skilled birth attendance for obstetric emergencies produced mixed results. Among 16 studies that fit the inclusion criteria, there is a medium, positive, non-significant association between training and TBA knowledge of risk factors and conditions requiring referral; and small, positive, significant associations between TBA referral behavior and maternal service use. These results cannot be causally attributed to TBA training because of the overall quality of studies; moreover, in several studies TBA training was a component of integrated intervention packages. The effort and expense of more rigorous research focusing on TBA training to improve access to emergency obstetric care are difficult to justify. The referral process is complex; the real effects of TBA training on TBA and maternal behavior are likely to be small; and while the proportion of TBA-attended births worldwide varies, it is, on average, quite low. The behavioral determinants and logistical barriers to care seeking for emergency obstetric care are generally well known. We suggest a more promising research agenda would reposition the questions surrounding referral into a broader ecological perspective. PMID- 15279932 TI - Denial of impending death: a discourse analysis of the palliative care literature. AB - Terminally ill patients and their families are often referred to as being "in denial" of impending death. This study uses the qualitative method of discourse analysis to investigate the usage of the term "denial" in the contemporary hospice and palliative care literature. A Medline search (1970-2001) was performed combining the text words "deny" and "denial" with the subject headings "terminal care", "palliative care" and "hospice care," and restricted to English articles discussing death denial in adults. The 30 articles were analysed using a constant comparison technique and emerging themes regarding the meaning and usage of the words "deny" and "denial" identified. This paper focusses on the theme of denial as an individual psychological process. Three dominant subthemes were distinguished: denial as an unconscious "defence mechanism", denial as "healthy" and denial as temporary. The analysis focusses on the intertextuality of these themes with each other and with previous texts on the denial of death. Elements of the psychoanalytic definition of denial as an unconscious defence mechanism are retained in the literature but are interwoven with new themes on patient choice. The result is an overall discourse that is conflictual and at times self contradictory but overall consistent with the biomedical model of illness. I suggest that the representation of death denial elaborated in these articles may be related to a larger discourse on dying in contemporary Western society, which both invites patients to participate in the planning of their death and labels those who do not comply. PMID- 15279933 TI - First use of two-dimensional polyacrylamide gel electrophoresis to determine phylogenetic relationships. AB - Methods for microbial classification are not always capable of distinguishing between isolates at the species level. We have previously characterised four Ferroplasma isolates that were >98.9% similar at the 16S rDNA level, the isolates showed marked phenotypic differences, and one isolate was borderline on the 70% species boundary from DNA-DNA similarity data. In this study we have used statistical comparisons of two-dimensional polyacylamide gel electrophoresis gels for classification of closely related isolates. From the protein profile similarities an un-rooted tree was constructed that was congruent with a tree derived from DNA-DNA similarities. PMID- 15279934 TI - Development of a microtitre-based spectrophotometric method to monitor Babesia divergens growth in vitro. AB - Babesia divergens multiplication cycle involves erythrocyte invasion, intracellular division, and erythrocyte lysis with the simultaneous liberation of hemoglobin. We have decided to set up a spectrophotometric protocol based on hemoglobin concentration in the culture supernatants to monitor B. divergens in vitro growth. After the selection of 405 nm as the most appropriate endpoint hemoglobin wavelength in our conditions (hemoglobin concentration in the supernatant), cultures were standardized [1 x 10(9) red blood cell (RBC)/ml, 1 2.5 x 10(5) infected red blood cell (iRBC)/ml] to allow their monitoring over 3 days. The protocol was then compared to the most commonly used growth measurement methods: parasitemia counting and [(3)H]hypoxanthine incorporation. An excellent correlation was demonstrated between A(405) of the culture supernatant and parasitemia of the iRBC, whatever the RBC concentration used in the medium. This correlation was also evidenced between A(405) and [(3)H]hypoxanthine incorporation for [(3)H]hypoxanthine concentrations lower than 4 microCi/ml. Our assays also highlighted the inhibitory effect of [(3)H]hypoxanthine on B. divergens growth even when used at low concentrations (0.8 microCi/ml) and for a short incorporation duration (24 h). This effect was confirmed by both A(405) and parasitemia counting. In conclusion, A(405) measurement of B. divergens culture supernatant represents a simple, rapid, safe, and reliable way to measure the in vitro growth of this parasite. Generation times of three different B. divergens strains were then determined by the protocol described here and varied between 8 h 36 min and 13 h 8 min. PMID- 15279935 TI - Binding interactions of Escherichia coli with globotetraosylceramide (globoside) using a surface plasmon resonance biosensor. AB - Surface plasmon resonance with an alkane L1 chip was used to investigate the binding of uropathogenic Escherichia coli, carrying adhesion receptors, to globotetraosylceramide (globoside; GbO4). The immobilization of globoside was reproducible and resulted in a stable globoside layer on the L1 chip. The data indicated that the globoside-immobilized L1 chip could be used for studying interactions with live or chemically fixed E. coli. The results indicated that the dissociation time was significantly reduced in glutaraldehyde-fixed E. coli as compared to living cells. Overall, the report demonstrates the significance of the L1 chip in terms of sensitivity, specificity, handling, and speed when studying globoside/E. coli interactions. This model may assist in screening for compounds that can inhibit the binding of uropathogenic E. coli to glycolipid ligands on target cells. PMID- 15279936 TI - Improvement of recoveries for the determination of protozoa Cryptosporidium and Giardia in water using method 1623. AB - The U.S. Environmental Protection Agency has developed method 1623 for simultaneous detection of Cryptosporidium oocysts and Giardia cysts in water. Method 1623 includes four major steps: filtration, immunomagnetic separation (IMS), fluorescent antibody (FA) staining and microscopic examination. It was noted that the recovery levels following IMS-FA and FA staining were high, averaging more than 92.0% and 89.0% for C. parvum oocysts and G. lamblia cysts, respectively. In contrast, when the filtration step was incorporated, the recovery level of C. parvum oocysts declined significantly to 18.1% in seeded tap water, while a relatively high recovery level of 77.2% for G. lamblia cysts could still be achieved. Further study indicated that the recovery level of C. parvum oocysts could be enhanced significantly when an appropriate amount of silica particles was added to a water sample. The recovery level of C. parvum oocysts was affected by particle size and concentration. The optimal silica particle size was determined to be within the range of 5-40 microm, and the corresponding optimal silica concentration was 1.42 g for 10-l tap water. When both G. lamblia cysts and C. parvum oocysts were spiked into the tap water sample containing the optimum amount of silica particles, the average recovery levels of oocysts and cysts were 82.7% and 75.4%, respectively. The results obtained clearly suggested that addition of an appropriate amount of silica particles could improve the recovery level of C. parvum oocysts significantly and yet there was no noticeable deleterious effect on the recovery level of G. lamblia cysts. Further study indicated that the rotation time in the IMS procedure using the Dynal GC-Combo IMS kit (which was recommended in method 1623) was important for G. lamblia cyst detection. In contrast, the recovery level of C. parvum oocysts was not affected by the rotation time. Furthermore, it was found that the recovery levels of C. parvum oocysts using methods 1622 and 1623 were quite close although different IMS kits were used in the two methods. PMID- 15279937 TI - Development of a robust microtiter plate-based assay method for assessment of bioactivity. AB - A microtiter plate-based assay was developed for the quantitative monitoring of bioactive compound production in Streptomyces hygroscopicus fermentation samples. The method reported demonstrates the successful application of the theories of disk diffusion based methods of bioactivity assessment, to a microtiter assay for high throughput analysis. The assay method facilitates the generation of the dose response curve of test organisms (Escherichia coli, Bacillus subtilis and Saccharomyces cerevisiae) to a bioactive compound. Using this dose-response curve, the method facilitates definition of three distinct Minimum Inhibitory Concentration (MIC) values for use in the characterisation of the bioactive attributes of a sample. The assay uses established standard procedures to facilitate adaptation of the assay for use with a wider range of test microorganisms. Errors due to the assumption of a linear relationship between turbidity and biomass concentration are also reduced, due to incorporation of a step to convert turbidity to biomass concentration, for use in the calculation of bioactivity. PMID- 15279938 TI - Identification and quantification of arsC genes in environmental samples by using real-time PCR. AB - The arsC gene is responsible for the first step in arsenate biotransformation encoding the enzyme arsenate reductase. The quantitative real-time PCR method was developed to quantify the abundance of the arsC genes in environmental samples contaminated with arsenic. Two sets of primers that showed high specificity for the target arsC gene were designed based on consensus sequences from 13 bacterial species. The arsC gene was used as an external standard instead of total DNA in the calibration curve for real-time PCR, which was linear over six orders of magnitude and the detection limit was estimated to be about three copies of the gene. Soil samples from arsenic contaminated sites were screened for arsC genes by using PCR and showed the presence of this gene. The copy numbers of the gene ranging from 0.88 x 10(4) to 1.56 x 10(5) per ng total DNA were found in eight arsenic contaminated samples. Soil samples from a bioreactor containing pulp mill biomass and high concentration of arsenate showed a tenfold higher count of arsC gene copies than soil samples collected underground from an arsenic-rich gold mine. PMID- 15279939 TI - A simple screening protocol for the identification of quorum signal antagonists. AB - Quorum sensing (QS) is a mechanism by which diverse microorganisms can control specific processes in response to population density. A relatively well-known form of QS among Proteobacteria involves production and subsequent response to acylated homoserine lactones (AHLs). Quorum sensing inhibition (QSI), targeting AHL-dependent signaling, has been reported as a strategy for the control of biofilm formation used by several marine organisms. We developed a simple soft agar overlay protocol, based on pigmentation inhibition, to rapidly screen for the presence of potential QSI by bacteria and plants. For bacterial screens, test organisms are first streaked onto their appropriate media and incubated overnight. For plant screens, the plant material (leaf, stem, flower, etc.) is placed onto LB agar. The bacterial growth or plant samples are then covered with an overlay of LB soft agar containing an inoculum of either Pseudomonas aureofaciens 30-84 or Chromobacterium violaceum ATCC 12472 (indicator cultures) and then incubated overnight. These indicator bacteria regulate pigment production by N-hexanoyl-HSL (C6-HSL) QS and are readily inhibited by AHL analogues and other antagonists. QSI is indicated by the lack of pigment production of the indicator culture in the vicinity of the test sample. Growth inhibition of the indicator culture indicates possible antibiotic production. Two different biosensor organisms based on derivatives of Agrobacterium tumefaciens and C. violaceum, capable of detecting a range of AHLs were used to determine whether QSI is due to the production of interfering AHLs competing with the C6 HSL regulation of C. violaceum and P. aureofaciens pigment production. This simple protocol will facilitate the screening of multiple organisms for the production of potential antifouling compounds. PMID- 15279940 TI - Chemotactic responses of the fish-parasitic scuticociliate Philasterides dicentrarchi to blood and blood components of the turbot Scophthalmus maximus, evaluated using a new microplate multiassay. AB - This study describes a new capillary-type microplate multiassay for characterization of protozoal chemotactic responses, allowing up to 32 assays to be run simultaneously. We used the new multiassay to evaluate the chemoattractant activity of turbot blood components and turbot cells for the facultative parasite Philasterides dicentrarchi, which is responsible for significant losses in turbot farming. Preliminary tests indicated that the assay requires 3-4 h for detection of chemoattractant activity, that it can be performed effectively using the ciliate axenic culture medium, and that it distinguishes clearly between different concentrations of chemoattractant. Application of the assay indicated that whole blood and serum from normal turbot, and especially infected turbot, have strong chemoattractant activity for P. dicentrarchi trophozoites, whereas neither turbot blood cells nor other turbot cells nor bacteria were significant chemoattractants. These results raise the possibility that turbot serum components are involved in host detection and host invasion by P. dicentrarchi, in line with previous findings indicating that turbot with skin lesions show increased susceptibility to P. dicentrarchi infection. PMID- 15279941 TI - An optical microsensor to measure fluorescent light intensity in biofilms. AB - We have developed an optical microsensor to quantify fluorescent light intensity distribution in biofilms. The optical system consisted of a beam splitter, light couplers, filters and a spectrophotometer able to accept the fiberoptic cable to measure fluorescent light intensity. The emitted light, fluorescence from the biofilm, was collected at the tip of the optical microsensor and was transferred to a spectrophotometer via a fiberoptic cable. The total fluorescent light intensity was evaluated from the emission spectrum by numerical integration. The newly developed fiberoptic microsensor was tested using a Staphylococcus aureus strain producing yellow fluorescent protein (YFP) grown as biofilm. We used a 405 nm violet laser diode for excitation, and measured the emission intensity between 480 nm and 540 nm. The optical microsensor that quantifies fluorescent light intensity is a promising tool in biofilm research which often requires detection and quantification of fluorescent light intensity distribution generated by various fluorescent proteins. PMID- 15279942 TI - Quantitative and qualitative comparison of density-based purification methods for detection of Cryptosporidium oocysts in turbid environmental matrices. AB - Purification methods for Cryptosporidium oocysts are usually selected on the basis of recovery yield, but the amount of particulate debris in environmental matrices could limit efficiency of oocyst detection by microscopic examination or PCR detection. Previous studies have shown that the standard immunomagnetic separation (IMS) procedure would not be the most suitable method for oocyst purification from turbid matrices. We compared the capacity of Percoll-sucrose flotation and six other density-based purification methods to achieve selective separation of Cryptosporidium oocysts from particulate debris. Rate of oocyst recovery and particulate loading in the purified suspensions were chosen as comparison criteria for the different purification methods. In most earlier studies, the chemical treatments employed to obtain a purified oocyst suspension modify the surface properties of oocysts in spiked samples. Assuming this produces unrealistic conditions affecting the evaluation of purification methods, we performed the present study with native oocysts. Flotation and gradient procedures were tested with and without formaldehyde ethyl acetate (FEA) separation. FEA separation was found to be unsuitable. Filtration and Percoll gradient did not allow selective oocyst separation from debris. Among the purification methods suitable for routine microscopic examination, Percoll sucrose flotation provided the best recovery rates. For automated enumeration systems or PCR detection, potassium bromide and especially Nycodenz gradients appeared to be the most suitable purification methods. Potassium bromide and Nycodenz gradients provided the best balance between oocyst recovery and particulate load. PMID- 15279943 TI - A combined dielectrophoresis, traveling wave dielectrophoresis and electrorotation microchip for the manipulation and characterization of human malignant cells. AB - The study of the dielectric properties of micrometer- or nanometer-scale particles is of particular interest in present-day applications of biomedical engineering. Electrokinetics utilises electrically energised microelectrode structures within microfluidic chambers to noninvasively probe the physiological structure of live cancer cells. A system is described that combines the three complementary techniques of dielectrophoresis (DEP), travelling wave dielectrophoresis (TWD) and electrorotation (ROT) for the first time on a single, integrated chip (3 x 6 mm). The chip employs planar microelectrode arrays fabricated on a silicon substrate to facilitate the synthesis of the various nonuniform electric fields required for the controlled manipulation, measurement and characterization of mammalian cells. A study of the dielectric properties of human malignant cells (Daudi and NCI-H929) was performed to demonstrate the potential and the versatility of the system in providing a fully programmable microsystem. PMID- 15279944 TI - Application of oligonucleotide array technology for the rapid detection of pathogenic bacteria of foodborne infections. AB - A rapid and accurate method for detection for common pathogenic bacteria in foodborne infections was established by using oligonucleotide array technology. Nylon membrane was used as the array support. A mutation region of the 23S rRNA gene was selected as the discrimination target from 14 species (genera) of bacteria causing foodborne infections and two unrelated bacterial species. A pair of universal primers was designed for PCR amplification of the 23S rRNA gene. Twenty-one species (genera)-specific oligonucleotide detection probes were synthesized and spotted onto the nylon membranes. The 23S rRNA gene amplification products of 14 species of pathogenic bacteria were hybridized to the oligonucleotide array. Hybridization results were analyzed with digoxigenin linked enzyme reaction. Results indicated that nine species of pathogenic bacteria (Escherichia coli, Campylobacter jejuni, Shigella dysenteriae, Vibrio cholerae, Vibrio parahaemolyticus, Proteus vulgaris, Bacillus cereus, Listeria monocytogenes and Clostridium botulinum) showed high sensitivity and specificity for the oligonucleotide array. Two other species (Salmonella enterica and Yersinia enterocolitica) gave weak cross-reaction with E. coli, but the reaction did not affect their detection. After redesigning the probes, positive hybridization results were obtained with Staphylococcus aureus, but not with Clostridium perfringens and Streptococcus pyogenes. The oligonucleotide array can also be applied to samples collected in clinical settings of foodborne infections. The superiority of oligonucleotide array over other tests lies on its rapidity, accuracy and efficiency in the diagnosis, treatment and control of foodborne infections. PMID- 15279945 TI - Development of multi-color FISH method for analysis of seven Bifidobacterium species in human feces. AB - We have developed a multi-color fluorescence in situ hybridization (FISH) method which detects, by a single reaction, all seven species of Bifidobacterium (B. adolescentis, B. angulatum, B. bifidum, B. breve, B. catenulatum, B. dentium, and B. longum), the dominant bacteria in human feces. First, eight new types of oligonucleotide probe were designed, complementary with the 16S rRNA sequence specific to genus Bifidobacterium and each bifidobacterial species described above. Using whole cell hybridization, the fluorescent intensity was measured against the bacterial species targeted by each probe, to show that each probe is specific to the targeted bacteria and that the relative fluorescent intensity (RFI) as an indicator of probe accessibility is high at 61-117%. Then, bacterial species-specific probes were labeled with fluorochromes (FITC, TAMRA and Cy5) in seven different ways, singly or in combination. Using these probes, seven species of Bifidobacterium were differentially stained in mixed samples of cultured bacteria and feces from adult volunteers, proving the efficacy of this technique. PMID- 15279946 TI - Biogenic amine receptors in parasitic nematodes: what can be learned from Caenorhabditis elegans? AB - The biogenic amines, serotonin, octopamine, tyramine and dopamine regulate many essential processes in parasitic nematodes, such as pharyngeal pumping, muscle contraction, and egg-laying, as well as more complex behaviors, such as mechanosensation and foraging, making biogenic amine receptors excellent targets for drug discovery. This review is designed to summarize our knowledge of nematode biogenic amine signaling and preliminarily identify some of the key receptors involved in the regulation of biogenic amine-dependent behaviors through an analysis of the free-living nematode, Caenorhabditis elegans. PMID- 15279947 TI - Localization of organellar proteins in Plasmodium falciparum using a novel set of transfection vectors and a new immunofluorescence fixation method. AB - The apicoplast and mitochondrion of the malaria parasite Plasmodium falciparum are important intracellular organelles and targets of several anti-malarial drugs. In recent years, our group and others have begun to piece together the metabolic pathways of these organelles, with a view to understanding their functions and identifying further anti-malarial targets. This has involved localization of putative organellar proteins using fluorescent reporter proteins such as green fluorescent protein (GFP). A major limitation to such an approach is the difficulties associated with using existing plasmids to genetically modify P. falciparum. In this paper, we present a novel series of P. falciparum transfection vectors based around the Gateway recombinatorial cloning system. Our system makes it considerably easier to construct fluorescent reporter fusion proteins, as well as allowing the use of two selectable markers. Using this approach, we localize proteins involved in isoprenoid biosynthesis and the posttranslational processing of apicoplast-encoded proteins to the apicoplast, and a protein putatively involved in the citric acid cycle to the mitochondrion. To confirm the localization of these proteins, we have developed a new immunofluorescence assay (IFA) protocol using antibodies specific to the apicoplast and mitochondrion. In comparison with published IFA methods, we find that ours maintains considerably better structural preservation, while still allowing sufficient antibody binding as well as preserving reporter protein fluorescence. In summary, we present two important new tools that have enabled us to characterize some of the functions of the apicoplast and mitochondrion, and which will be of use to the wider malaria research community in elucidating the localization of other P. falciparum proteins. PMID- 15279948 TI - A Plasmodium berghei reference line that constitutively expresses GFP at a high level throughout the complete life cycle. AB - Green fluorescent protein (GFP) is a well-established reporter protein for the examination of biological processes. This report describes a recombinant Plasmodium berghei, PbGFPCON, that constitutively expresses GFP in a growth responsive manner in its cytoplasm from a transgene that is integrated into the genome and controlled by the strong promoter from a P. berghei elongation factor 1alpha gene. All life cycle forms of PbGFPCON except for male gametes can be easily visualized by fluorescent microscopy. PbGFPCON showed similar growth characteristics to wild type P. berghei parasites throughout the whole life cycle and can therefore be used as a reference line for future investigations of parasite-host cell interactions. The principle of automated fluorescence-based counting and sorting of live parasites from host cell backgrounds and different parasite forms from complex mixtures such as asynchronous blood stages is established. PbGFPCON allows the visualization and investigation of live parasite stages that are difficult and labor-intensive to observe, such as the liver and mosquito stages. PbGFPCON can be employed to establish the phenotype of independent mutant parasites. With the recent development of a second, independent selectable marker in P. berghei, PbGFPCON is a useful tool to investigate the effect of further genetic modifications on host-parasite interactions. PMID- 15279949 TI - Quantification of Plasmodium falciparum gametocytes in differential stages of development by quantitative nucleic acid sequence-based amplification. AB - Two quantitative nucleic acid sequence-based amplification assays (QT-NASBA) based on Pfs16 and Pfs25, have been developed to quantify sexual stage commitment and mature gametocytes of Plasmodium falciparum. Pfs16 mRNA is expressed in all sexual forms including sexually committed ring stages while expression of Pfs25 mRNA is restricted to late stage gametocytes. Both assays showed a sensitivity of one sexual stage parasite/microl of blood. Blood samples from experimentally infected non-immune human volunteers were tested for Plasmodium falciparum by standard microscopy, a previously developed asexual 18S rRNA QT-NASBA, Pfs16 and Pfs25 mRNA QT-NASBA. Pfs16 QT-NASBA was positive in 9 out of 10 volunteers within 48 h after first detection of 18S rRNA, mostly before or at the day of positive microscopy. In contrast, the Pfs25 mRNA QT-NASBA was negative during the 28 days of follow-up, but consistently positive in gametocyte samples from naturally infected Kenyan patients. These data suggest that sexual stage commitment can occur early in the blood-stage infection without successful maturation into infectious gametocytes. In conclusion, Pfs16 and Pfs25 QT-NASBA assays in combination with a previously developed asexual stage QT-NASBA allow for the separate quantification of all developmental stages present in the circulation. The application of sexual stage QT-NASBA assays may contribute to a better understanding of the biology and epidemiology of malaria transmission. PMID- 15279950 TI - Molecular characterization of bifunctional hydroxymethyldihydropterin pyrophosphokinase-dihydropteroate synthase from Plasmodium falciparum. AB - A 2118-base pair gene encoding the bifunctional hydroxymethyldihydropterin pyrophosphokinase-dihydropteroate syntheses of Plasmodium falciparum (pfPPPK DHPS) was expressed under the control of the T5 promoter in a DHPS-deficient Escherichia coli strain. The enzyme was purified to near homogeneity using nickel affinity chromatography followed by gel filtration and migrates as an intense band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis with apparent mass of approximately 83 kDa. Gel filtration suggested that the native pfPPPK DHPS might exist as a tetramer of identical subunits. The enzyme was found to be Mg2+ - and ATP-dependent and had optimal temperature ranging from 37 to 45 degrees C with peak activity at pH 10. Sodium chloride and potassium chloride at 0.2 and 0.4 M, respectively, activated the activity of the enzyme but higher salt concentrations were inhibitory. Guanidine-HCl and urea inhibited the enzyme activity by 50% at 0.25 and 0.9 M, respectively. Kinetic properties of the recombinant pfPPPK-DHPS were investigated. Sulfathiazole and dapsone were potent inhibitors of pfPPPK-DHPS, whilst sulfadoxine, sulfanilamide, sulfacetamide and p aminosalicylic acid were less inhibitory. Our construct provides an abundant source of recombinant pfPPPK-DHPS for crystallization and drug screening. PMID- 15279951 TI - Functional interdependence of the DBLbeta domain and c2 region for binding of the Plasmodium falciparum variant antigen to ICAM-1. AB - Cytoadherence of Plasmodium falciparum-infected erythrocytes is associated with severe malaria and is primarily mediated through binding of the variant surface antigen P. falciparum erythrocyte membrane protein 1 (PfEMP1) to specific host ligands. Infected erythrocyte binding to Intercellular Adhesion Molecule 1 (ICAM 1) has been implicated as having a role in cerebral malaria, a major cause of death from P. falciparum infection. We have examined ICAM-1-binding PfEMP1 proteins in the cytoadhesive P. falciparum strain IT4/25/5 in order to extend our understanding of binding. For A4tres, the ICAM-1 binding region was previously shown to reside within contiguous DBL2beta and c2 domains. We determined the gene sequence encoding IT-ICAM var, and showed that ICAM-1 binding in this protein also maps to DBL2betac2 domains that have 48% amino acid identity to A4tres. By truncation and chimera analysis, most of the DBL2beta and the first half of the c2 region were required for A4tres binding to ICAM-1, suggesting this tandem should be considered a structural-functional combination for ICAM-1 binding. Of interest, a chimera formed between two different ICAM-1 binding domains did not bind ICAM-1, suggesting a functional interdependence between DBL2beta and c2 from the same protein. As gene recombination and gene conversion are important mechanisms for generating diversity in the PfEMP1 protein family, this finding implies an extra level of constraint on the functional evolution of binding traits. Knowledge about the PfEMP1::ICAM-1 interaction may allow the development of interventions to prevent binding and disease. PMID- 15279952 TI - Translation initiation factor eIF-5A from Plasmodium falciparum. AB - Eukaryotic translation initiation factor (eIF-5A) is a highly conserved and essential protein that contains the unique amino acid hypusine. The first step in the post-translational biosynthesis of hypusine, the transfer of an aminobutyl moiety from the polyamine substrate spermidine to the -amino group of a specific lysine residue in the eIF-5A precursor, is catalyzed by the enzyme deoxyhypusine synthase. A cDNA encoding a protein homologous to eIF-5A was isolated by plaque hybridization from a cDNA library of Plasmodium falciparum. The cloned cDNA contains an open reading frame encoding a protein of 161 amino acids, which shares a high sequence identity with other eukaryotic eIF-5A sequences. A phylogenetic tree constructed with eIF-5A from P. falciparum and 16 other eIF-5A sequences of eukaryotic and archaeal origin reveals that plasmodial eIF-5A together with other apicomplexan eIF-5A show a higher degree of homology to plant proteins than to animal and fungal sequences. The plasmodial eIF-5A gene was expressed as a six-histidine tagged fusion protein in Escherichia coli. Radioactive incorporation studies with [1,8-3H] spermidine indicated that this protein can serve as a substrate for human deoxyhypusine synthase. Results of quantitative real-time PCR studies with synchronized erythrocytic stages of P. falciparum revealed no significant induction or downregulation but only some variation in the expression level of plasmodial eIF-5A in ring, trophozoite and schizont stage. PMID- 15279953 TI - Transgene expression in Schistosoma mansoni: introduction of RNA into schistosomula by electroporation. AB - Despite their significance in human and veterinary medicine, and the ability to maintain the parasites in the mouse, relatively little functional detail is available regarding the biology of schistosomes. This deficit is due largely to the lack of well-developed molecular tools for manipulating gene expression in these parasites. Here, we describe an electroporation protocol that provides a routine approach for efficiently introducing nucleic acids into schistosomes. Using luciferase-encoding RNA for electroporation, and luciferase activity as a read-out, we established 400 microg/ml of RNA, and a 20 ms pulse at 125 V using a square wave electroporation generator to be optimal for electroporating schistosomes. Under these conditions schistosomula from 1 hr to 18 hr old could be successfully electroporated, the majority of parasites within a population expressed the introduced RNA, and acute mortality was negligible. Electroporation, as described here, makes possible experimental studies using transiently expressed constitutively active and/or dominant negative mutant proteins, etc. In addition, the finding that electroporation can be used to introduce RNA into schistosomula raises the possibility of using this approach to introduce either DNA constructs or dsRNA sequences, both of which might be expected to have longer-term, ideally inheritable, effects. PMID- 15279955 TI - MSP domain proteins show enhanced expression in male germ line cells. AB - Nematode sperm utilize a crawling motility based on polymerization- depolymerization of a nematode-specific cytoskeletal molecule-major sperm protein (MSP). While several proteins that interact with and regulate MSP filament formation have been identified using in vitro approaches, it is likely that additional molecules participate in vivo in the regulation of MSP cytoskeletal dynamics. By comparing EST data generated from an Ascaris suum testis germinal zone cDNA library with EST data from other tissue- and stage-specific A. suum cDNA libraries and with expression profile data from Caenorhabditis elegans, 42 genes were selected with exclusive or enhanced expression in male germ line cells. In addition to 11 protein kinases and seven protein phosphatases, 10 genes encoding proteins with protein-protein interaction domains were identified. These potential cytoskeletal modifiers included five novel MSP-domain proteins (As MDPs). All five As-mdps were highly expressed in male germ line cells, but only As-mdp-2, 3 and 5 were transcribed exclusively in the testis. The prediction that As-MDP-1, 2 and 3 were cytosolic components and that As-MDP-4 and 5 were associated with the sperm cell membrane proteins was supported by the results of immunoblotting experiments. The 23 members of the MDP family of proteins from C. elegans were predicted to be transcribed in the testis. The findings provide additional candidates to the growing list of molecules that regulate MSP cytoskeletal dynamics. PMID- 15279956 TI - Fluorescent protein tagging in Toxoplasma gondii: identification of a novel inner membrane complex component conserved among Apicomplexa. AB - Toxoplasma gondii is an obligate intracellular parasite, and its sub-cellular organization shows clear adaptations to this life-style. In addition to organelles shared among all eukaryotes, the organism possesses a number of specialized compartments with important roles in host cell invasion and intra cellular survival. These unique aspects of the parasite's biology are also reflected in its genome. The ongoing genome sequencing efforts for T. gondii and related apicomplexans predict a high proportion of genes unique to the phylum, which lack homologs in other model organisms. Knowing the sub-cellular localization of these gene products will be an important first step towards their functional characterization. We used a library approach wherein parasite genomic DNA was fused to the yellow fluorescent protein (YFP) gene. Parasites transformed with this library were screened by flow cytometry and fluorescence microscopy. Clones tagged in a wide variety of sub-cellular compartments (nucleus, mitochondria, ER, dense granules (secreted), spliceosome, plasma membrane, apicoplast, inner membrane complex) were isolated and confirmed using compartment specific markers. Clones with tags in parasite-specific localizations were subjected to insert rescue and phenotypic verification using an in vitro recombination system. Among the genes identified is a novel inner membrane complex gene (IMC3) conserved among Apicomplexa. PMID- 15279957 TI - A GFP-based motif-trap reveals a novel mechanism of targeting for the Toxoplasma ROP4 protein. AB - The protozoan parasite Toxoplasma gondii is a highly specialized eukaryote that contains a remarkable number of intracellular compartments, some unique to Apicomplexans and others typical of eukaryotes in general. We have established a green fluorescent protein (GFP)-based motif-trap to identify proteins targeted to different intracellular locations and subsequently the signals responsible for this sorting. The motif-trap involves the transfection and integration of a linearized GFP construct which lacks a promoter and an initiator methionine codon. FACS is used to isolate parasites in which GFP fuses in-frame into a coding region followed by screening by fluorescence microscopy for those containing GFP targeted to specific intracellular compartments. GFP trapping was successful using vectors designed for integration into regions encoding exons and vectors that were engineered with a splice acceptor site for integration into regions encoding introns. This strategy differs from most protein traps in that the resulting fusions are expressed from the endogenous promoter and starting methionine. Thus, problems from inappropriate expression levels or the creation of fortuitous targeting signals seen in library-based traps are diminished. Using this approach, we have trapped GFP localized to a number of intracellular compartments including the nucleus, nucleolus, endoplasmic reticulum, cytosol, parasite surface and rhoptries of Toxoplasma. Further analysis of a parasite clone containing GFP targeted to the rhoptries shows GFP fused to the gene encoding the rhoptry protein ROP4 and has elucidated an additional mechanism for targeting of this protein. PMID- 15279958 TI - Identification of a mitochondrial superoxide dismutase with an unusual targeting sequence in Plasmodium falciparum. AB - The intraerythrocytic stages of Plasmodium falciparum are exposed to oxidative stress and require functional anti-oxidant systems to survive. In addition to the parasite's known iron-dependent superoxide dismutase PfSOD1, a second SOD gene (PfSOD2) interrupted by 8 introns was identified on chromosome 6. Molecular modelling shows that the structure of PfSOD2 is similar to other iron-dependent SODs and phylogenetic analysis suggests PfSOD1 and PfSOD2 are the result of an ancestral gene duplication. The deduced amino acid sequence of PfSOD2 is similar to PfSOD1 but has a long N-terminal extension. Immunofluorescence studies show that PfSOD1 is cytosolic, whereas the N-terminal extension of PfSOD2 targets a green fluorescent protein fusion into the parasite's mitochondrion. Both SOD genes are transcribed during the erythrocytic cycle with PfSOD1 mRNA levels up to 35-fold higher than those of PfSOD2. Northern blots demonstrated that the mRNA levels of both SOD genes are up-regulated upon exposure to oxidative stress. PMID- 15279959 TI - Predominant clonal evolution leads to a close parity between gene expression profiles and subspecific phylogeny in Trypanosoma cruzi. AB - We investigated the relationships between overall phylogenetic diversity in Trypanosoma cruzi evidenced by multilocus markers (MLEE and RAPD) on the one hand, and gene expression patterns, revealed by mRNA analysis on the other hand. Nineteen laboratory-cloned stocks representative of this parasite's overall phylogenetic diversity and ecogeographical range were analyzed using random amplified differentially expressed sequences (RADES). The bat trypanosome T. cruzi marinkellei was taken as outgroup. The profiles obtained showed that RADES polymorphism cannot be considered as a simple subsample of general RAPD polymorphism. Indeed, many RADES bands were not present in general RAPD profiles, and vice versa. Phylogenies obtained from RADES on the one hand, and MLEE/RAPD on the other hand, were very similar. This suggests that in spite of the recent observation of hybrid genotypes and mosaic genes in T. cruzi, clonal evolution in this parasite has been preponderant enough on an evolutionary scale to carve the polymorphism on all types of DNA sequences, including expressed genes, although these genes are assumed to undergo natural selection pressure contrary to noncoding sequences and neutral polymorphisms. PMID- 15279960 TI - Inhibitory and neutral antibodies to Plasmodium falciparum MSP119 form ring structures with their antigen. AB - Blood-stage malaria vaccine candidates include surface proteins of the merozoite. Antibodies to these proteins may either block essential steps during invasion or render the merozoite susceptible to phagocytosis or complement-mediated degradation. Structural information on merozoite surface proteins complexed to antibodies provides crucial information for knowledge-based vaccine design. The major merozoite surface protein MSP1 is an abundant surface molecule in Plasmodium falciparum. Only a subset of antibodies against MSP119 inhibits invasion (inhibitory antibodies), whereas other antibodies binding to MSP119 have no effect on invasion (neutral antibodies). Here we report on the complex of MSP119 with both inhibitory monoclonal antibody 12.10 and neutral monoclonal antibody 2F10. The complexes were established using both whole IgG's and Fab fragments, and analysed by dynamic light scattering, electron microscopy and analytical ultra centrifugation. Specific ring structures were formed in the ternary complex with the two antibodies, providing direct evidence of non overlapping epitopes on MSP119. Mutational studies also indicated that the epitopes of the inhibitory and neutral antibodies are spatially remote. PMID- 15279961 TI - Ancylostoma ceylanicum anticoagulant peptide-1: role of the predicted reactive site amino acid in mediating inhibition of coagulation factors Xa and VIIa. AB - Hookworm infection is a leading cause of gastrointestinal blood loss and iron deficiency anemia in developing countries. Ancylostoma hookworms secrete potent anticoagulants, which have been shown to target coagulation factors Xa and the factor VIIa/Tissue Factor complex, respectively. The goal of these experiments was to determine the mechanism of action of three recombinant hookworm anticoagulants using in vitro assays. Three hookworm coagulation inhibitors were expressed and purified, along with site directed mutants targeting each of the predicted P1 inhibitory reactive site amino acid residues. Using chromogenic assays, it has been confirmed that Ancylostoma caninum Anticoagulant Peptide 5 (AcAP5) inhibits coagulation factor Xa (fXa) by a canonical, substrate-like mechanism. In contrast, Ancylostoma ceylanicum Anticoagulant Peptide-1 (AceAP1) binds to and inhibits fXa by both active site and non-active site mediated interactions. Data from in vitro studies also demonstrates that AceAP1 inhibits the factor VIIa/Tissue complex (fVIIa/TF) and displays a distinct pattern of fXa binding. Together, these data suggest that the human hookworm A. ceylanicum has evolved a single anticoagulant that targets multiple components of the mammalian coagulation response, effectively mimicking the concerted action of the two related inhibitors from A. caninum. Despite the amino acid sequence similarity, AceAP1 appears to interact with coagulation proteases fXa and fVIIa by a novel mechanism, perhaps explaining its spectrum of inhibitory activity. PMID- 15279954 TI - A survey of Leishmania braziliensis genome by shotgun sequencing. AB - We have carried out a survey of the genome of Leishmania (Viannia) braziliensis by shotgun sequencing. Approximately 15% of the haploid genome of the parasite (5.15 Mb of genomic sequence) was obtained. A large number of known and putative genes, predicted to be involved in several cellular processes, were identified. Some genomic features were investigated, such as the general G + C content, which was found to be lower than L. major (57% versus 63%). BlastN searches revealed that 60.2% of the clusterized GSS sequences displayed similarity to L. major genomic sequences, while a BlastX search showed that 45.3% of the thus obtained predicted protein sequences showed similarity to annotated proteins of L. major. Further comparison of the degree of conservation between L. major and L. braziliensis revealed that coding regions are much more conserved than non-coding ones. The shotgun sequence analysis of Leishmania braziliensis appears to be an efficient and suitable strategy contributing to the search for vaccines and novel drug targets. The sequence data described in this paper have been submitted to the dbGSS database under the following accession numbers (BX530413 to BX530454; BX530456 to BX530718; BX538354 to BX539305; BX539350 to BX540325; BX541002 to BX544869; BX544893 to BX545685; BX897701 to BX897710; BX905184 to BX907797; BX907798 to BX908381; BX908403 to BX908718). All data including sequences are also available at (www.ebi.ac.uk/embl/). PMID- 15279962 TI - Transcriptome of axenic liver stages of Plasmodium yoelii. AB - Plasmodium liver stages or early exo-eythrocytic forms (EEFs) contain antigens that are essential for achieving sterile, protective immunity against malaria. Yet, attempts at identifying these antigens have been hampered by the challenge of obtaining large numbers of purified EEFs, uncontaminated with hepatocyte material. Using a recently described system for producing axenically cultured EEFs from Plasmodium yoelii, we have constructed a cDNA library and generated 1453 expressed sequence tags (ESTs) resulting in 652 unique transcripts. Analysis of the library provides insight into processes required for the initiation and development of Plasmodium liver stages, such as protein degradation, cell cycle progression and nutrient transport. Analysis of the gene expression profile of liver stages, as revealed by this library, suggests that liver stages represent a shift from "sporozoite-like" to "blood-stage-like". This is the first study of the transcriptional repertoire of Plasmodium liver stages. PMID- 15279963 TI - Cloning and characterization of an immunoreactive gene encoding a calcium-binding protein from Naegleria fowleri. PMID- 15279965 TI - Intron encoded sequences necessary for trans splicing in transiently transfected Brugia malayi. PMID- 15279964 TI - Trichomonas vaginalis ribosomal DNA: analysis of the intergenic region and mapping of the transcription start point. PMID- 15279966 TI - Genetic complementation of Leishmania deficient in PSA (GP46) restores their resistance to lysis by complement. PMID- 15279967 TI - Diterpenes from the brown algae Dictyota dichotoma and Dictyota linearis. AB - Nineteen secondary metabolites of the brown alga Dictyota dichotoma (Huds.) Lam. and fifteen metabolites of the brown alga D. linearis (Ag.) Grev. were isolated and their chemical structures were elucidated on the basis of their NMR and mass spectral data. The diterpenes isopachydictyolal (1) from D. dichotoma and 4alpha acetyldictyodial (2) from D. linearis are new natural products. The antiviral activity of metabolites isolated in adequate amounts was evaluated in laboratory assays against Herpes simplex virus I (HSV I) and Poliomyelitis Virus I, using Vero cells as hosts. PMID- 15279968 TI - Limonoids from Turraea floribunda (Meliaceae). AB - Six novel limonoids and limonoid derivatives, turraflorins D-I along with the known turraflorins A and B have been isolated from seed of the South African Turraea floribunda (Meliaceae). PMID- 15279969 TI - Constituents of two Flourensia species. AB - The MeOH extract of aerial parts of Flourensia riparia Grisebach (Asteraceae) afforded a sesquiterpene lactone, 4beta-hydroxy-4,10alpha-dimethyl 7alphaH,8alphaH-eudesman-11-ene-8,12-olide, together with septuplinolide, its isomer at positions C-5 and C-10. In addition, known flavonoids, p hydroxyacetophenone derivatives, carabrone and isoalantolactone were identified. Three known flavonoids and a benzofuran were isolated from Flourensia campestris Wedd. PMID- 15279970 TI - Sesquiterpenes from the east African sandalwood Osyris tenuifolia. AB - The essential oil of the east African sandalwood Osyris tenuifolia was investigated by chromatographic and spectroscopic methods. Beside several already known sesquiterpenes four new compounds could be isolated by preparative gas chromatography and their structures investigated by mass spectroscopy and NMR techniques. Two of the new compounds--tenuifolene (17) and ar-tenuifolene (15)- show a new sesquiterpene backbone. 2,(7Z,10Z)-Bisabolatrien-13-ol (23) and the cyclic ether lanceoloxide (21) belong to the bisabolanes. PMID- 15279971 TI - Amritosides A, B, C and D: clerodane furano diterpene glucosides from Tinospora cordifolia. AB - Four new clerodane furano diterpene glucosides (amritosides A, B, C and D) were isolated as their acetates from Tinospora cordifolia stems. The structures of these compounds were established on the basis of spectroscopic studies. PMID- 15279972 TI - Terpenes from Otostegia integrifolia. AB - The essential oil and chloroform extract of air-dried leaves of Otostegia integrifolia Benth. were investigated for the first time using analytical and preparative gas chromatography (GC), GC-mass spectrometry (MS) and NMR techniques. A total of 40 constituents including monoterpenes, sesquiterpenes, diterpenes and their derivatives were identified. A prenylbisabolane type diterpene, 1-methyl-4-(5,9-dimethyl-1-methylene-deca-4,8-dienyl)cyclohexene was identified as a major component. The chloroform extract of the leaves yielded two labdane type diterpenoids, 15,16-epoxy-3alpha,9alpha-dihydroxy-labda-13(16),14 diene and 9(13),15(16)-diepoxy-3alpha-hydroxy-16-dihydrolabda-14-ene, a saturated hydrocarbon, pentatriacontane, and stigmasterol. The structures of the isolated compounds were established by spectroscopic methods. PMID- 15279973 TI - Trihydroxylated linear diterpenes from the brown alga Bifurcaria bifurcata. AB - Two novel polar diterpenes were isolated from the brown alga Bifurcaria bifurcata collected off the Atlantic coast of Morocco, and their structures established by spectral methods. Both compounds are trihydroxylated acyclic diterpenes derived from 12-hydroxygeranylgeraniol. They were tested in vitro for their cytotoxicity and proved to be active against the NSCLC-N6 cell line. Their absolute configuration at the C-12 position has been determined with a modified Mosher's method [J. Am. Chem. Soc. 113 (1991) 4092] and that of the 12 hydroxygeranylgeraniol (bifurcadiol) has been revised. PMID- 15279974 TI - Kaurane and abietane diterpenoids from Tripterygium doianum (Celastraceae). AB - Extraction of Tripterygium doianum (Celastraceae) afforded five new diterpenoids and 11 known diterpenoids belonging to the ent-kaurane and abietane families. Their structures were established based on spectroscopic studies. The isolated compounds showed moderate cytotoxicity against human tumor cell assays. PMID- 15279975 TI - Chemical constituents of leaves and stem bark of Plumeria obtusa. AB - The continued studies on the constituents of the fresh leaves and stem bark of Plumeria obtusa Linn. have led to the isolation and characterization of four new triterpenoids, dammara-12,20(22)Z-dien-3-one (1), dammara-12,20(22)Z-dien-3beta ol (2), olean-12-en-3beta,27-diol (3), and 27-hydroxyolean-12-en-3-one (4) and 12 known compounds, which included eight triterpenoids; dammara-3beta,20(S),25-triol (5), urs-12-en-3beta-hydroxy-27-Z-feruloyloxy-28-oic acid (6), 3beta-hydroxyolean 12-en-28-oic acid (7), 3beta,27-dihydroxylupan-29-ene (8), 3beta-hydroxylupan-29 en-28-oic acid (9), 3beta-hydroxyursan-12-en-28-oic acid (11), 3beta-hydroxy-27-p coumaroyloxy-olea-12-en-28-oic acid (12) and urs-12-en-3-one (15); an iridoid 1alpha-plumieride (10); a cardenolide 3alpha,14beta-dihydroxy-17beta-card-20(22) enolide (13); a fatty acid ester methyl n-octadecanoate (14) and a steroid 3beta hydroxy-delta5-stigmastane (16). The new constituents were characterized through spectroscopic studies including 1D (1H and 31C NMR) and 2D (COSY-45, NOESY, J resolved, HMQC and HMBC) NMR and chemical transformations. This is the first report on the isolation of dammarane triterpenoids from P. obtusa. Compounds 5 and 6 are hitherto unreported from P. obtusa. The known compounds were identified by comparison of their spectral data with those reported in the literature. PMID- 15279976 TI - Azorellane diterpenes from Azorella cryptantha. AB - Azorella cryptantha yielded the diterpenes, azorellolide and the dihydroderivative, dihydroazorellolide, together with the known yaretol and 1alpha,10beta,4beta,5alpha-diepoxy-7alpha-germacran-6beta-ol. Both possess a carbon skeleton type that may originate from rearrangement of the mulinane skeleton. PMID- 15279977 TI - Cytotoxic phenylethanol glycosides from Psidium guaijava seeds. AB - Phytochemical investigations of the acetone extract of Psidium guaijava seeds has led to the isolation of five known flavonoid glycosides, two phenolic glycosides and two new phenylethanoid glycosides which have been identified as 1-O-3,4 dimethoxy-phenylethyl-4-O-3,4-dimethoxy cinnamoyl-6-O-cinnamoyl-beta-D glucopyranose and 1-O-3,4-dimethoxyphenylethyl-4-O-3,4-dimethoxy cinnamoyl-beta-D glucopyranose, on the basis of chemical, physical and spectroscopic methods of analysis. PMID- 15279978 TI - Three acylated flavone glycosides from Sideritis ozturkii Aytac & Aksoy. AB - From the aerial parts of Sideritis ozturkii, three new flavonoids, chrysoeriol 7 O-[2'''-O-caffeoyl-O-acetyl-beta-D-glucopyranosyl-(1-->2)-beta-D glucopyranoside], chrysoeriol 7-O[2'''-O-caffeoyl-beta-D-glucopyranosyl-(1 -->2) beta-D-glucopyranoside] and chrysoeriol 7-O[2'''-O-p-coumaroyl-6'''-beta-O-acetyl D-glucopyranosyl-(1-->2)-beta-D-glucopyranoside] named as ozturkosides A, B and C, respectively, were isolated, along with three known phenylethanoid glycosides, verbascoside, leucoseptoside A, martynoside and five known diterpenoids, 7 epicandicandiol, linearol, sidol, sideroxol, epoxyisolinearol. The structures were elucidated mainly by spectroscopic methods. PMID- 15279979 TI - Lignans from the roots of Echinops giganteus. AB - Two new lignans, (+)-4-(3-methylbutanoyl)-2,6-di(3,4-dimethoxy)phenyl-3,7 dioxabicyclo[3.3.0]octane and (+)-4-hydroxy-2,6-di(3,4-dimethoxy)phenyl-3,7 dioxabicyclo[3.3.0]octane, together with the known lupeol and sitosteryl beta-D glucopyranoside, have been isolated from the roots of Echinops giganteus var. lelyi C. D. Adams (Compositae). The structures were elucidated on the basis of spectral studies and comparison with published data. PMID- 15279980 TI - Secondary mould metabolites of Cladosporium tenuissimum, a hyperparasite of rust fungi. AB - Investigation of the extracts of a culture of Cladosporium tenuissimum, a known hyperparasite of several rust fungi, gave rise to the isolation of cladosporols B E (2-5). Their structure and stereochemistry were elucidated on the basis of 1H and 13C NMR evidence and CD measures. Cladosporols 1-5 were active in inhibiting the urediniospore germination of the bean rust agent Uromyces appendiculatus. PMID- 15279981 TI - (-)-Amarbellisine, a lycorine-type alkaloid from Amaryllis belladonna L. growing in Egypt. AB - A new lycorine-type alkaloid, named (-)-amarbellisine, was isolated from the bulbs of Egyptian Amaryllis belladonna L. together with the well known alkaloids (-)-lycorine, (-)-pancracine, (+)-vittatine, (+)-11-hydroxyvittatine, and (+) hippeastrine. The new alkaloid, containing the pyrrolo[de]phenanthridine ring system, was essentially characterised by spectroscopic and optical methods, and proved to be the 2-methoxy-3a,4,5,7,11b,11c-hexahydro-1H-[1,3]dioxolo[4,5 j]pyrrolo[3,2,1-de]phenanthridinol. By using HPTLC technique we also carried out a comparative study of the relative and total alkaloidal content at two different stages of plant growth. Finally, the antimicrobial activity of the isolated alkaloids was assayed. PMID- 15279982 TI - Venalstonine and dioxokopsan derivatives from Kopsia fruticosa. AB - Two new venalstonine derivatives, viz., venacarpines A and B, and one new dioxokopsan derivative, kopsorinine, in addition to the kopsifolines A-F, and 11 other known alkaloids, were isolated from a Malayan Kopsia species. The structures of the new alkaloids were determined using NMR and MS analysis. PMID- 15279983 TI - Alkaloids from Delphinium pentagynum. AB - Aerial parts of a collection of Delphinium pentagynum Lam. from Niebla, Southern Spain, furnished one diterpene alkaloid, 2-dehydrodeacetylheterophylloidine, two norditerpene alkaloids, 14-demethyl-14-isobutyrylanhweidelphinine and 14-demethyl 14-acetylanhweidelphinine, the known alkaloids 14-deacetylnudicauline, methyllycaconitine, 14-deacetyl-14-isobutyrylnudicauline, 14-acetylbrowniine, browniine, delcosine, lycoctonine, 18-methoxygadesine, neoline, karakoline and the aporphine alkaloid magnoflorine. Structures of the alkaloids were established by MS, 1D and 2-D NMR techniques. PMID- 15279984 TI - Iridoid glucosides of Paederota lutea and the relationships between Paederota and Veronica. AB - In a chemical investigation of the water soluble compounds in Paederota lutea eight known iridoids were isolated together with a new one with a 8,9-double bond, namely paederotoside (10-O-benzoyl-6'-O-alpha-arabino(1-->6)-beta glucopyranosyl arborescosidic acid) and the 6-hydroxy-flavone glucoside 4'-O methylscutellarein 7-O-beta-glucopyranoside. The known iridoid glucosides were 8 epiloganic acid, gardoside, aucubin, catalpol and the 6-O-esters of catalpol: veronicoside, catalposide, amphicoside and verproside. The compounds isolated show that Paederota has a glycoside content almost identical to that of a general Veronica species, and this is in good agreement with the results from recent investigations of the phylogeny of Veronica and its closest relatives, where Paederota is placed as a sister-group next to Veronica. In an analysis of the iridoid glucosides from some of these relatives, it is shown that Veronica, Paederota, Picrorhiza and Veronicastrum are all characterized by containing 6-O esters of catalpol. Some less closely related taxa namely: Lagotis, Wulfenia, Plantago, Aragoa and Globularia instead contain 10-O-esters of catalpol or aucubin. PMID- 15279985 TI - Acylated iridoid glucosides from Veronica anagallis-aquatica. AB - Three new (1-3) and four known iridoid glucosides (4-7) as well as a known phenylethanoid glycoside (8) were isolated from the aerial parts of Veronica anagallis-aquatica and their structures were determined as 6'-O-benzoyl-8 epiloganic acid named aquaticoside A (1), 6'-O-p-hydroxybenzoyl-8-epiloganic acid named aquaticoside B (2), 6'-O-benzoyl-gardoside named aquaticoside C (3), veronicoside (4), catalposide (5), verproside (6), verminoside (7) and martynoside (8) on the basis of 1D and 2D NMR spectral analysis. PMID- 15279986 TI - Acetophenone C-glucosides and stilbene O-glucosides in Upuna borneensis. AB - Three acetophenone C-glycosides; 2,4,6-trihydroxyacetophenone 3-C-beta-(2'-O-p hydroxybenzoyl)-glucopyranoside, 2,4,6-trihydroxyacetophenone 3-C-beta-(2'-O-E coumaroyl)-glucopyranoside, 2,4,6-trihydroxyacetophenone 3-C-beta-(2'-O-E cinnamoyl)-glucopyranoside, and two resveratrol O-glycosides; piceid 2'-O-p hydroxybenzoate and, piceid 2'-O-E-ferulate, together with three known compounds were isolated from the acetone soluble part of stem of Upuna borneensis (Dipterocarpaceae). The structures of isolates were determined by spectral analysis including extensive 2D-NMR spectral analyses. PMID- 15279987 TI - Amides from Streptomyces hygroscopicus and their antimicrobial activity [corrected]. AB - Three amides, N-salicyloyl-2-aminopropan-1,3-diol (1) and 1-acetyl-N-salicyloyl-2 aminopropan-3-ol (2) including a natural product, N-salicyloyl-2-aminopropan-1-ol (3) were isolated from an ethyl acetate extract of the culture filtrate of Streptomyces hygroscopicus [corrected] The structures of these compounds were unambiguously established by interpretation of their spectral data including, a series of 1D and 2D-NMR and MS analyses. Compounds 1-3 showed significant antibacterial activity against a wide range of Gram positive and Gram negative bacteria. PMID- 15279988 TI - Coumaroyl triterpene lactone, phenolic and naphthalene glycoside from stem bark of Diospyros angustifolia. AB - From the ethanolic extract of stem bark of D. angustifolia three new compounds, a coumaroyl triterpene lactone, diospyrosooleanolide (1), a phenolic glycoside, diospyrososide (2) and a naphthalene glycoside, diospyrosonaphthoside (3) were isolated along with five known compounds (4-8). The structures of these compounds were established on the basis of spectroscopic and chemical evidences. PMID- 15279989 TI - A Dhb-microcystin from the filamentous cyanobacterium Planktothrix rubescens. AB - A Dhb-microcystin variant was isolated from the filamentous cyanobacterium Planktothrix rubescens. Its structure was elucidated as (E)-Dhb-microcystin-HilR ([D-Asp3, (E)-Dhb7]microcystin-HilR) on the basis of spectral data and amino acid analysis after acid hydrolysis. PMID- 15279990 TI - Volatile constituents of Capillipedium parviflorum. AB - The essential oil of aerial parts of Capillipedium parviflorum (family Poaceae) was obtained by hydrodistillation in 0.4% yield on dry weight basis. The oil was analysed by capillary GC and GC-MS techniques. Two new compounds from plant source 4-nonanol 51.7% and 4-undecanone 23.5% predominated in the essential oil and were separated by column chromatography, identified and confirmed by 1H and 13C NMR. Thirty one compounds were identified from the essential oil accounting for 96% of total identifications. PMID- 15279991 TI - Phenylpropanoid derivatives from edible canna, Canna edulis. AB - Two phenylpropanoid sucrose esters were isolated from dry rhizomes of Canna edulis Ker Gawl., along with a known phenylpropanoid sucrose ester and four known phenylpropanoids. On the basis of analysis of spectroscopic data and chemical evidence, these two phenylpropanoid sucrose esters were shown to be 3-O-p coumaroyl-6-O-feruloyl-beta-D-fructofuranosyl 6-O-acetyl-alpha-D-glucopyranoside and 3,6-di-O-p-coumaroyl-beta-D-fructofuranosyl 6-O-acetyl-alpha-D glucopyranoside. PMID- 15279992 TI - Insect antifeedant compounds from Nothofagus dombeyi and N. pumilio. AB - A bioassay-guided purification of the extracts of Nothofagus dombeyi and N. pumilio leaves yielded several triterpenes and flavonoids including 2-O acetylmaslinic acid, 3-O-acetyl 20,24,25-trihydroxydammarane, and 3,20,24,25 tetrahydroxydammarane as new natural products. All the isolated compounds were assessed for antifeeding activity against the 5th instar larvae of Ctenopsteustis obliquana. 12-Hydroxyoleanolic lactone and pectolinarigenin from N. dombeyi and dihydrooroxylin A from N. pumilio, showed significant antifeeding activity. PMID- 15279993 TI - Seasonal cardenolide production and Dop5betar gene expression in natural populations of Digitalis obscura. AB - Productivity variations and seasonal fluctuations of cardenolides have been studied in 10 natural populations of Digitalis obscura distributed in three bioclimatic belts. Main cardenolides in D. obscura plants are those of the series A and such predominance (ca. 80-85%) over the series B metabolites is independent of the population studied or the degree of maturity of the leaves. Primary glycosides represent ca. 50-60% of total cardenolides; this percentage did not vary among populations or with the leaf age but increased in summer and decreased in winter. A correlation analysis between plant biomass and cardenolide content showed a positive relationship of these parameters, which, according to the bioclimatic distribution of the populations, suggests that certain environmental conditions may cause marked decreases in plant biomass together with a reduction in productivity. Cardenolide contents changed in the timecourse of the four seasons as a multiple response to distinct plant and/or environmental factors. The lowest production was recorded in May, followed by a fast cardenolide accumulation in summer, a decreasing phase in autumn, and a stationary phase in winter. We also analysed the seasonal expression of the gene encoding the progesterone 5beta-reductase, enzyme producing the required 5beta-configured intermediaries of cardenolides. A fragment of the isolated partial genomic sequence was used as a probe for Northern analysis to study the seasonal gene expression in selected populations. The expression pattern showed increasing levels from February to July and a further reduction in autumn, although harmful climatic conditions seems to induce overexpression of this gene. PMID- 15279994 TI - The class III peroxidase multigenic family in rice and its evolution in land plants. AB - Plant peroxidases (class III peroxidases, E.C. 1.11.1.7) are secreted glycoproteins known to be involved in the mechanism of cell elongation, in cell wall construction and differentiation, and in the defense against pathogens. They usually form large multigenic families in angiosperms. The recent completion of rice (Oryza sativa japonica c.v. Nipponbare) genome sequencing allowed drawing up the full inventory of the genes encoding class III peroxidases in this plant. We found 138 peroxidase genes distributed among the 12 rice chromosomes. In contrast to several other gene families studied so far, peroxidase genes are twice as numerous in rice as in Arabidopsis. This large number of genes results from various duplication events that were tentatively traced back using a phylogenetic tree based on the alignment of conserved amino acid sequences. We also searched for peroxidase encoding genes in the major phyla of plant kingdom. In addition to gymnosperms and angiosperms, sequences were found in liverworts, mosses and ferns, but not in unicellular green algae. Two rice and one Arabidopsis peroxidase genes appeared to be rather close to the only known sequence from the liverwort Marchantia polymorpha. The possible relationship of these peroxidases with the putative ancestor of peroxidase genes is discussed, as well as the connection between the development of the class III peroxidase multigenic family and the emergence of the first land plants. PMID- 15279995 TI - The sesquiterpene hydrocarbons of maize (Zea mays) form five groups with distinct developmental and organ-specific distributions. AB - The sesquiterpene hydrocarbon chemistry of maize (Zea mays) inbred line B73 was analyzed by both direct solvent extraction and headspace sampling. In seedlings, 15 olefinic compounds were identified, and 21 olefins were detected in mature plants after anthesis. Both solvent extracts and collections of headspace terpenes were found to contain the same compounds in the same relative proportions suggesting that there is no selective barrier to release from plants. Approximately 25% of the stored pool was found to be released from young seedlings per hour. The individual sesquiterpenes varied extensively in their abundance among different organs and developmental stages. Compounds could be divided into five different groups such that the members of each group always occur together in the same constant ratios to one another. Each group has a distinct distribution pattern. Group A includes the two dominant compounds, (E) beta-farnesene and alpha-bergamotene, and appears only after herbivore damage in seedlings, but is constitutively present in the leaves and husks after anthesis. The major compounds of group B, including alpha-copaene, germacrene D and delta cadinene, were present throughout the seedling but found only in husks of mature plants. The group C compounds, beta-bisabolene and an unknown sesquiterpene olefin, are restricted to the roots. The presence of group D and E compounds was confined to the leaves and husk of mature plants. The complex sesquiterpene mixture of group D is identical to the products formed by the previously identified terpene synthase TPS4, suggesting that each of the four other sesquiterpene hydrocarbon mixtures may also represent the products of a single terpene synthase. PMID- 15279996 TI - Changes in glycosidase activities during galactoglucomannan oligosaccharide inhibition of auxin induced growth. AB - The inhibition of 2,4-D-induced elongation growth by galactoglucomannan oligosaccharides (GGMOs) in pea stem segments (Pisum sativum L. cv. Tyrkys) after 18 h of incubation results in changes of extracellular, intracellular and cell wall glycosidase activities (beta-D-glucosidase, beta-D-mannosidase, beta-D galactosidase, beta-D-xylosidase, alpha-D-galactosidase, and alpha-L arabinosidase). GGMOs lowered the glycosidase activities in the extracellular fraction, while in the cell wall fractions their activities were markedly increased. The intracellular enzyme alpha-d-galactosidase increased while the beta-d-galactosidase decreased in activity in response to the GGMO treatment. Extracellular enzymes showed low values of activities in comparison with intracellular and cell wall glycosidases. It is evident that GGMOs can alter auxin induced elongation and glycosidase activities in different compartments of the cell, however, the mode and site of their action remains unclear. PMID- 15279997 TI - Salt-induced lipid changes in Catharanthus roseus cultured cell suspensions. AB - Salt treatment strongly affected cell growth by decreasing dry weight. Exposure of Catharanthus roseus cell suspensions to increasing salinity significantly enhanced total lipid (TL) content. The observed increase is mainly due to high level of phospholipids (PL). Hundred mM NaCl treatment increased phospholipid species phosphatidylcholine (PC) and phosphatidylethanolamine (PE), whereas it reduced glycolipid ones monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG) but not sulfoquinovosyldiacylglycerol (SQDG). Moreover, fatty acid composition was clearly modified when cells were cultured in the presence of 100 mM NaCl, whereas only few changes occurred at 50 mM. Salt treatment decreased palmitic acid (16:0) level and increased that of linolenic acid (18:2). Such effect was observed in phospholipid species PC and PE and in glycolipid DGDG. Double bond index (DBI) was enhanced more than 2-fold in fatty acids of either glycolipids or phospholipids from cells submitted to 100 mM NaCl. Free sterol content was also significantly enhanced, especially at 100 mM NaCl, whereas free sterols/phospholipids (St/PL) ratio was slightly decreased. All these salt-induced changes in membrane lipids suggest an increase in membrane fluidity of C. roseus cells. PMID- 15279998 TI - Isolation and characterization of wound-inducible carboxypeptidase inhibitor from tomato leaves. AB - In a previous report [Mol. Gen. Genet. 228 (1991) 281], carboxypeptidase inhibitor protein (CPI) mRNA was found to accumulate in leaves of wounded tomato plants, but CPI protein could not be detected. In contrast, we found that CPI protein does accumulate in tomato leaves in response to wounding, and also in response to treatment with either systemin, methyl jasmonate (MeJ), oligogalacturonic acid, or chitosan. Identification of CPI protein was confirmed by its inhibition of metallo-carboxypeptidase A (CPAase), which was used as an assay during purification of the inhibitor from leaves of MeJ-treated tomato plants. Amino acid sequence analysis and mass spectroscopic analyses of the pure protein confirmed its identity as CPI. The pure protein inhibited CPAase in a 1:1 stoichimetric interaction. Time course analyses of the induction of CPI mRNA in tomato leaves in response to wounding indicated that the gene is a member of the group of "late genes" that code for defensive proteins synthesized in leaves in response to herbivore attack. PMID- 15279999 TI - Flavonoids from shoots and roots of Trifolium repens (white clover) grown in presence or absence of the arbuscular mycorrhizal fungus Glomus intraradices. AB - White clover (Trifolium repens) plants were grown in the presence or absence of the arbuscular mycorrhizal fungus Glomus intraradices. Flavones, 4',5,6,7,8 pentahydroxy-3-methoxyflavone and 5,6,7,8-tetrahydroxy-3-methoxyflavone, as well as two flavones 3,7-dihydroxy-4'-methoxyflavone and 5,6,7,8-tetrahydroxy-4' methoxyflavone never previously reported in plants, were isolated. The known 3,5,6,7,8-pentahydroxy-4'-methoxyflavone, 2',3',4',5',6'-pentahydroxy-chalcone, 6 hydroxykaempferol, 4',5,6,7,8-pentahydroxyflavone and 3,4'-dimethoxykaempferol were also obtained. Analysis of extracts obtained from roots and shoots revealed that the compositions of the flavonoid mixtures varied with growing conditions. Quercetin, acacetin and rhamnetin accumulated in roots of inoculated plants, whereas they were not detected in non-inoculated plants. PMID- 15280000 TI - Mulinane-type diterpenoids from Azorella compacta display antiplasmodial activity. AB - Two mulinane-type diterpenoids were isolated from Azorella compacta; namely 20 hydroxymulin-11,13-dienyl acetate and 13,14-dihydroxymulin-11-en-20-oic acid. The structures were elucidated by analysis of their spectroscopic data. These compounds, as well as three previously isolated diterpenes, were evaluated as potential in vivo growth inhibitors of Plasmodium berghei NK 65 on infected mice at an intraperitoneal dose of 10 mg/kg/day. Sixty percent and forty-two percent growth inhibition were obtained with 17-acetoxymulin-11,13-dien-20-oic acid and 13, 14-dihydroxymulin-11-en-20-oic acid, respectively. PMID- 15280001 TI - Fresh organically grown ginger (Zingiber officinale): composition and effects on LPS-induced PGE2 production. AB - Gas chromatography in conjunction with mass spectrometry, a technique previously employed to analyze non-volatile pungent components of ginger extracts modified to trimethylsilyl derivatives, was applied successfully for the first time to analyze unmodified partially purified fractions from the dichloromethane extracts of organically grown samples of fresh Chinese white and Japanese yellow varieties of ginger, Zingiber officinale Roscoe (Zingiberaceae). This analysis resulted in the detection of 20 hitherto unknown natural products and 31 compounds previously reported as ginger constituents. These include paradols, dihydroparadols, gingerols, acetyl derivatives of gingerols, shogaols, 3-dihydroshogaols, gingerdiols, mono- and diacetyl derivatives of gingerdiols, 1 dehydrogingerdiones, diarylheptanoids, and methyl ether derivatives of some of these compounds. The thermal degradation of gingerols to gingerone, shogaols, and related compounds was demonstrated. The major constituent in the two varieties was [6]-gingerol, a chemical marker for Z. officinale. Mass spectral fragmentation patterns for all the compounds are described and interpreted. Anti inflammatory activities of silica gel chromatography fractions were tested using an in vitro PGE2 assay. Most of the fractions containing gingerols and/or gingerol derivatives showed excellent inhibition of LPS-induced PGE2 production. PMID- 15280002 TI - Minor pyrano-isoflavones from Eriosema kraussianum: activity-, structure-, and chemical reaction studies. AB - The isolation and identification of two minor pyrano-isoflavones from Eriosema kraussianum is described. New studies on the original pyrano-isoflavones shows that: (i) kraussianone 2 (a major compound in the plant) can be cyclised under acid conditions, (ii) kraussianones 3 and 5 cause contraction (not relaxation as anticipated) of corpus cavernosum tissue and (iii) the structures proposed previously for 4 and 5 are confirmed by the data obtained from an X-ray study of 5. PMID- 15280003 TI - Insect growth inhibition by tocotrienols and hydroquinones from Roldana barba johannis. AB - The methanol extract from the aerial parts of Roldana barba-johannis (Asteraceae) afforded sargachromenol, sargahydroquinoic acid, and sargaquinoic acid. These natural products and their corresponding acetylated and methylated derivatives showed insecticidal and insect growth regulatory activities against the Fall Armyworm [Spodoptera frugiperda J.E. Smith, (Lepidoptera: Noctuidae)], an important insect pest of corn. The most active compounds were sargachromenol and its acetylated derivative; sargahydroquinoic acid and its acetylated derivative; and a mixture of sargachromenol, sargahydroquinoic acid, and sargaquinoic acid (6:3:1) and the acetylated form of this mixture. All these compounds and mixtures had significant inhibitory effects between 5.0 and 20.0 ppm in diets. Most compounds were insecticidal to larvae, with lethal doses between 20 and 35 ppm. In addition, these substances also demonstrated scavenging properties toward 2,2 diphenyl-1-picrylhydrazyl radical in TLC autographic and spectrophotometric assays. These compounds appear to have selective effects on the pre-emergence metabolism of the insect. The results from these compounds were fully comparable in activity to those known natural insect growth inhibitors such as gedunin and methanol extracts of Cedrela salvadorensis and Yucca periculosa. These substances may be useful as natural insecticidal agents. PMID- 15280004 TI - Antioxidant phenolic and quinonemethide triterpenes from Cheiloclinium cognatum. AB - The triterpenes, 22beta-hydroxypristimerin and cognatine, were isolated together with the known compounds pristimerin, maytenin, 20alpha-hydroxymaytenin, 22beta hydroxymaytenin, netzahualcoyol, netzahualcoyondiol and netzahualcoyone from root bark of Cheiloclinium cognatum. The structures of the isolated compounds were elucidated by interpretation of their spectral data, including gHMQC and gHMBC experiments. The isolates were investigated for their radical scavenging abilities through a spectrophotometric assay involving reduction of 2,2-diphenyl picryl hydrazyl (DPPH). PMID- 15280005 TI - Water-soluble polysaccharides from Salvia officinalis L. possessing immunomodulatory activity. AB - A water-soluble polysaccharide complex (A) composed of galactose (17.9%), 3-O methyl-galactose (3.0%), glucose (15.5%), mannose (8.3%), arabinose (30.4%), xylose (7.6%), fucose (2.6%), rhamnose (6.7%), and uronic acids (8.0%) has been isolated from the aerial parts of sage (Salvia officinalis L.) by cold water extraction. It showed a broad molecular-mass distribution pattern (Mw approximately 2000-93,000) with a predominance of polymers with Mw< 10,000. Ion exchange chromatography of A afforded six polymeric fractions (A1-A6) in which arabinogalactans associated with galacturonan and/or rhamnogalacturonan backbones prevail. Sage polysaccharides were examined for their immunomodulatory activity in the comitogenic thymocyte test which is interpreted as being an in vitro correlate of adjuvant activity. The acidic polysaccharide fractions A2, A3 and A4 exhibited the highest mitogenic and comitogenic activities of all fractions tested, and relatively high SI(comit)/SI(mit) ratios approximately 3 indicate potential adjuvant properties of these polysaccharides. PMID- 15280006 TI - Metabolomic analysis of Strychnos nux-vomica, Strychnos icaja and Strychnos ignatii extracts by 1H nuclear magnetic resonance spectrometry and multivariate analysis techniques. AB - 1H Nuclear magnetic resonance spectrometry and multivariate analysis techniques were applied for the metabolic profiling of three Strychnos species: Strychnos nux-vomica (seeds, stem bark, root bark), Strychnos ignatii (seeds), and Strychnos icaja (leaves, stem bark, root bark, collar bark). The principal component analysis (PCA) of the 1H NMR spectra showed a clear discrimination between all samples, using the three first components. The key compounds responsible for the discrimination were brucine, loganin, fatty acids, and Strychnos icaja alkaloids such as icajine and sungucine. The method was then applied to the classification of several "false angostura" samples. These samples were, as expected, identified as S. nux-vomica by PCA, but could not be clearly discriminated as root bark or stem bark samples after further statistical analysis. PMID- 15280007 TI - Constituents from the bark of Tabebuia impetiginosa. AB - The bark of Tabebuia impetiginosa afforded nineteen glycosides, consisting of four iridoid glycosides, two lignan glycosides, two isocoumarin glycosides, three phenylethanoid glycosides and eight phenolic glycosides. Their structures were determined using both spectroscopic and chemical methods. Iridoid glycosides, phenylethanoid glycosides and lignan glycosides had ajugol, osmanthuside H and secoisolariciresinol 4-O-beta-D-glucopyranoside as their structural elements, respectively, whereas the aglycone moieties of the isocoumarin glycosides were considered to be (-)-6-hydroxymellein. Phenolic glycosides had 4-methoxyphenol, 2,4-dimethoxyphenol, 3,4-dimethoxyphenol, 3,4,5-trimethoxyphenol and vanillyl 4 hydroxybenzoate as each aglycone moiety. Additionally, the sugar chains of these isocoumarin glycosides and phenolic glycosides were concluded to be beta-D apiofuranosyl-(1-->6)-beta-D-glucopyranoside as well as those of osmanthuside H and above phenylethanoid glycosides. PMID- 15280008 TI - Epidermal growth factor, neurotrophins and the metastatic cascade in prostate cancer. AB - Although cancer of the prostate (CaP) is the most commonly occurring cancer in males, there are major limitations in its diagnosis and long-term cure. Consequently, understanding the molecular mechanisms involved in the progression of CaP is of particular importance for production of pharmacological and biological agents to manage the disease. The development of the normal prostate is regulated by stromal-epithelial interactions via endocrine and paracrine factors, such as androgens and growth factors, which act as precise homeostatic regulators of cellular proliferation. Importantly, after a period of hormonal therapy, CaP shifts from an androgen-dependent to an androgen-independent state with a concomitant switch from paracrine to autocrine growth factor stimulation and subsequent upregulation of growth factor expression. Thus, growth factors and their receptors have a pivotal role in CaP. This is emphasized by current evidence obtained from clinical specimens as well as several in vitro and in vivo models strongly suggesting that epidermal growth factor and the neurotrophins (nerve growth factor, brain derived neurotrophin factor, neurotrophin-3 and neurotrophin-4/5) together with their tyrosine kinase receptors could play a very significant role in CaP progression. PMID- 15280009 TI - Fine-tuning the hydrophobicity of a mitochondria-targeted antioxidant. AB - The mitochondria-targeted antioxidant MitoQ comprises a ubiquinol moiety covalently attached through an aliphatic carbon chain to the lipophilic triphenylphosphonium cation. This cation drives the membrane potential-dependent accumulation of MitoQ into mitochondria, enabling the ubiquinol antioxidant to prevent mitochondrial oxidative damage far more effectively than untargeted antioxidants. We sought to fine-tune the hydrophobicity of MitoQ so as to control the extent of its membrane binding and penetration into the phospholipid bilayer, and thereby regulate its partitioning between the membrane and aqueous phases within mitochondria and cells. To do this, MitoQ variants with 3, 5, 10 and 15 carbon aliphatic chains were synthesised. These molecules had a wide range of hydrophobicities with octan-1-ol/phosphate buffered saline partition coefficients from 2.8 to 20000. All MitoQ variants were accumulated into mitochondria driven by the membrane potential, but their binding to phospholipid bilayers varied from negligible for MitoQ3 to essentially total for MitoQ15. Despite the span of hydrophobicites, all MitoQ variants were effective antioxidants. Therefore, it is possible to fine-tune the degree of membrane association of MitoQ and other mitochondria targeted compounds, without losing antioxidant efficacy. This indicates how the uptake and distribution of mitochondria-targeted compounds within mitochondria and cells can be controlled, thereby facilitating investigations of mitochondrial oxidative damage. PMID- 15280010 TI - His507 of acylaminoacyl peptidase stabilizes the active site conformation, not the catalytic intermediate. AB - Acylaminoacyl peptidase is a member of the prolyl oligopeptidase family. Amino acid sequence alignment suggests that the stabilization of the tetrahedral intermediate should be mediated by His507 rather than by a tyrosine residue found in the other family members of this serine peptidase group. The pH dependence of k(cat)/K(m) did not reveal any effect of His507. Substitution of an alanine for His507 gave the same bell-shaped pH rate profile with the same pK(a) values (7.0 and 8.7). However, the value of the rate constant was 85 times lower with the modified enzyme, which indicated that His507 is an important residue that is probably involved in the formation of the 3-dimensional structure. PMID- 15280011 TI - Identification of O2-induced peptides in an obligatory anaerobe, Clostridium acetobutylicum. AB - Clostridium acetobutylicum DSM792 (=ATCC824), a solvent producing obligate anaerobe, grew well after a shift in growth conditions from anoxic to microoxic at the mid exponential phase. In two-dimensional gel electrophoresis, a spot migrating at 45 kDa and three spots at 23 kDa accumulated after 30 min of flushing with 5% O(2)/95% N(2). Based on peptide mass fingerprints, the 45 kDa polypeptide was determined to be NP_347663 (A-type flavoprotein homologue) and the 23 kDa polypeptides were determined to be NP_350180 or NP_350181 (novel type rubrerythrin homologue). Northern blot analysis indicated that the expressions of these peptide transcripts were upregulated within 10 min after flushing with 5% O(2)/95% N(2). PMID- 15280012 TI - Two independent light signals cooperate in the activation of the plastid psbD blue light-responsive promoter in Arabidopsis. AB - The psbD blue light-responsive promoter (BLRP), whose activation has been considered to require strong blue light, is recognized only by SIG5 among six sigma factors of plastid RNA polymerase in Arabidopsis. We found SIG5 transcript accumulation was rapidly induced after a 30-min induction time by blue light (470 nm) with an intensity threshold of 5 micromol m(-2)s(-1) through cryptochromes. Besides this weak blue light, the psbD BLRP activation required the stronger light such as 50 micromol m(-2)s(-1) irrespective of blue or red light (660 nm). Thus, the two independent light signalings, the cryptochrome-mediated signaling to induce SIG5 transcription and the stronger light-dependent signaling, cooperate to activate the psbD BLRP. PMID- 15280013 TI - Tomatidine and lycotetraose, hydrolysis products of alpha-tomatine by Fusarium oxysporum tomatinase, suppress induced defense responses in tomato cells. AB - Many fungal pathogens of tomato produce extracellular enzymes, collectively known as tomatinases, that detoxify the preformed antifungal steroidal glycoalkaloid alpha-tomatine. Tomatinase from the vascular wilt pathogen of tomato Fusarium oxysporum f. sp. lycopersici cleaves alpha-tomatine into the aglycon tomatidine (Td) and the tetrasaccharide lycotetraose (Lt). Although modes of action of alpha tomatine have been extensively studied, those of Td and Lt are poorly understood. Here, we show that both Td and Lt inhibit the oxidative burst and hypersensitive cell death in suspension-cultured tomato cells. A tomatinase-negative F. oxysporum strain inherently non-pathogenic on tomato was able to infect tomato cuttings when either Td or Lt was present. These results suggest that tomatinase from F. oxysporum is required not only for detoxification of alpha-tomatine but also for suppression of induced defense responses of host. PMID- 15280014 TI - Presence of different phospholipase C isoforms in the nucleus and their activation during compensatory liver growth. AB - Phospholipase C (PLC) was purified from the membrane-depleted rat liver nuclei. About 60% of the total PLC-activity corresponded to beta1b isoform, 30% to PLC gamma1 and less than 10% to PLC-delta1. PLC-beta1b and -gamma1 were found in the nuclear matrix, while PLC-delta1 was detected in the chromatin. Two peaks of an increase in the total PLC-activity were detected occurring at 6 and 20 h after partial hepatectomy. An early increase in PLC-beta1b activity in the nuclear matrix was associated with serine phosphorylation of the enzyme, while the later increase paralleled the increase in the amount of protein. The increase in the PLC-gamma1 activity measured at 6 and 20 h after partial hepatectomy was associated with tyrosine phosphorylation of the enzyme. The activity of PLC delta1 and the amount of the protein found in the chromatin was increased only at 20 h after partial hepatectomy. PMID- 15280015 TI - Pim-1 kinase promotes inactivation of the pro-apoptotic Bad protein by phosphorylating it on the Ser112 gatekeeper site. AB - Constitutive expression of the Pim-1 kinase prolongs survival of cytokine deprived FDCP1 cells, partly via maintenance of Bcl-2 expression. Here, we show that Pim-1 colocalizes and physically interacts with the pro-apoptotic Bad protein and phosphorylates it in vitro on serine 112, which is a gatekeeper site for its inactivation. Furthermore, wild-type Pim-1, but not a kinase-deficient mutant, enhances phosphorylation of this site in FDCP1 cells and protects cells from the pro-apoptotic effects of Bad. Our results suggest that phosphorylation of Bad by Pim-1 is one of several mechanisms via which the Pim-1 kinase can enhance Bcl-2 activity and promote cell survival. PMID- 15280016 TI - Inhibitory action of novel aromatic diamine compound on lipopolysaccharide induced nuclear translocation of NF-kappaB without affecting IkappaB degradation. AB - 4-Methyl-N1-(3-phenyl-propyl)-benzene-1,2-diamine (JSH-23) is a novel chemically synthetic compound. The aromatic diamine JSH-23 compound exhibited inhibitory effect with an IC(50) value of 7.1 microM on nuclear factor (NF)-kappaB transcriptional activity in lipopolysaccharide (LPS)-stimulated macrophages RAW 264.7, and interfered LPS-induced nuclear translocation of NF-kappaB without affecting IkappaB degradation. This mechanism of action is very rare for controlling NF-kappaB activation. Furthermore, the compound inhibited not only LPS-induced expressions of tumor necrosis factor-alpha, interleukin (IL)-1beta, IL-6 and inducible nitric oxide synthase and cyclooxygenase-2 but also LPS induced apoptosis of the RAW 264.7 cells. PMID- 15280017 TI - The intracellular dissipation of cytosolic calcium following glucose re-addition to carbohydrate depleted Saccharomyces cerevisiae. AB - Glucose re-addition to carbohydrate starved yeast cells leads to a transient elevation of eytosolic calcium (TECC). Concomitantly, a cytosolic proton extrusion occurs through the activation of the vacuolar H(+)-ATPase and the plasma membrane H(+)-ATPases. This study addressed the dissipation of the TECC through intracellular compartmentalization and the possible affects of the H(+) ATPases on this process. Both the vacuole and the Golgi-ER apparatus were found to play important roles in distributing calcium to internal stores. Additionally, the inhibition of cytosolic proton extrusion augmented cytosolic calcium responses. A model where pH dependent cytosolic calcium buffering plays an important role in the dissipation of the TECC in Saccharomyces cerevisiae is proposed. PMID- 15280018 TI - NO-degradation by alfalfa class 1 hemoglobin (Mhb1): a possible link to PR-1a gene expression in Mhb1-overproducing tobacco plants. AB - Tobacco plants overproducing alfalfa class 1 hemoglobin (HOT plants) have been shown to have reduced necrotic symptom development. Here, we show that this altered pathogenic response is linked to a significant increase in the nitric oxide (NO)-affected pathogenesis-related (PR-1a) transcript accumulation in the transgenic plants. Homogenates of HOT transgenic seedlings were also found to have higher NO-scavenging activity than non-transformed ones. The NO-scavenging properties of recombinant alfalfa class1 hemoglobin have been examined. Recombinant Mhb1 (rMhb1) was produced in bacteria and purified using polyethylene glycol (10-25%) fractionation, chromatography on DEAE-Sephacel, and Phenyl Superose columns. After the final purification step, the obtained preparations were near homogeneous and had a molecular weight of 44 kDa determined by size exclusion chromatography and 23 kDa by SDS-PAGE, indicating that rMhb1 is a dimer. The protein participated in NO-degradation activity with NAD(P)H as a cofactor. After ion-exchange columns, addition of FAD was necessary for exhibiting maximal NO-degradation activity. The NAD(P)H-dependent NO-scavenging activity of rMhb1, which is similar to that of barley hemoglobin, supports a conclusion that both monocot and dicot class 1 hemoglobins can affect cellular NO levels by scavenging NO formed during hypoxia, pathogen attack and other stresses. PMID- 15280019 TI - Functional characterization of recombinant batroxobin, a snake venom thrombin like enzyme, expressed from Pichia pastoris. AB - A thrombin-like enzyme of Bothrops atrox moojeni venom, batroxobin, specifically cleaves fibrinogen alpha chain, resulting in the formation of non-crosslinked fibrin clots. The cDNA encoding batroxobin was cloned, expressed in Pichia pastoris and the molecular function of purified recombinant protein was also characterized. The recombinant batroxobin had an apparent molecular weight of 33 kDa by SDS-PAGE analysis and biochemical activities similar to those of native batroxobin. The purified recombinant protein strongly converted fibrinogen into fibrin clot in vitro, and shortened bleeding time and whole blood coagulation time in vivo. However, it did not make any considerable alterations on other blood coagulation factors. Several lines of experimental evidence in this study suggest that the recombinant batroxobin is a potent pro-coagulant agent. PMID- 15280020 TI - hOLF44, a secreted glycoprotein with distinct expression pattern, belongs to an uncharacterized olfactomedin-like subfamily newly identified by phylogenetic analysis. AB - Secreted proteins are indispensable for the development and differentiation of multicellular organisms. Cloning and characterization of novel or hypothetical genes encoding these proteins are therefore inviting great incentives. Using bioinformatics tools and experimental approaches, we isolated and characterized a human secreted glycoprotein, hOLF44, which contains a highly conserved olfactomedin-like (OLF) domain in the C-terminal. However, phylogenetic analysis revealed that hOLF44 is not clustered into any of the OLF subfamilies containing characterized members, and obviously falls into a newly identified uncharacterized OLF subfamily. Western blot analysis showed that hOLF44 protein is robustly secreted from the transfected COS-7 cells. Expression levels of hOLF44 mRNA are abundant in placenta, moderate in liver and heart, whereas fairly weak in other tissues examined. Immunohistochemical study on human term placenta demonstrated that hOLF44 is mainly localized extracellularly surrounding the syncytiotrophoblastic cells and very rarely expressed in the maternal decidua layer. These results suggest that hOLF44 may have matrix-related function involved in human placental and embryonic development, or play a similar role in other physiological processes. The further functional characterization of hOLF44 may provide insights into a better understanding of the newly identified OLF subfamily. PMID- 15280021 TI - Gliadin stimulates human monocytes to production of IL-8 and TNF-alpha through a mechanism involving NF-kappaB. AB - Wheat gliadin is the triggering agent in coeliac disease. In this study, we documented that proteolytic fragments of gliadin, in contrast to other food antigens, induced interleukin (IL)-8 and tumour necrosis factor-alpha (TNF-alpha) production and significantly increased interferon (IFN)-gamma-induced cytokine secretion in human monocytic line THP-1 cells. Stimulation with gliadin resulted in elevated phosphorylation of the IkappaBalpha molecule and increased NF kappaB/DNA binding activity that was inhibited by sulfasalazine, l-1-tosylamido-2 phenylethyl chloromethyl ketone and pyrrolidine dithiocarbamate (PDTC). The activation pathway was shown to be independent of the CD14 molecule. Less mature U-937 monocytes responded to gliadin stimulation by low IL-8 secretion, TNF-alpha production was not detectable. We propose that gliadin-induced activation of monocytes/macrophages can participate in mechanisms leading to the impairment of intestinal mucosa in coeliac patients. PMID- 15280022 TI - Characterisation of the Rab binding properties of Rab coupling protein (RCP) by site-directed mutagenesis. AB - Rab coupling protein (RCP) is a member of the Rab11-family of interacting proteins (Rab11-FIPs). Family members are characterised by their ability to interact with Rab11. This property is mediated by a conserved Rab binding domain (RBD) located at their carboxy-termini. Several Rab11-FIPs can also interact with other small GTPases. RCP interacts with Rab4 in addition to Rab11. To dissect out the individual properties of the Rab4 and Rab11 interactions with RCP, conserved amino acids within the RBD of RCP were mutated by site-directed mutagenesis. The effect of these mutations on Rab4 and Rab11 binding, and the intracellular localisation of RCP, was examined. Our results indicate that Rab11, rather than Rab4, mediates the intracellular localisation of RCP, and that the class I Rab11 FIPs compete for binding to Rab11. PMID- 15280023 TI - Gene selection and classification from microarray data using kernel machine. AB - The discrimination of cancer patients (including subtypes) based on gene expression data is a critical problem with clinical ramifications. Central to solving this problem is the issue of how to extract the most relevant genes from the several thousand genes on a typical microarray. Here, we propose a methodology that can effectively select an informative subset of genes and classify the subtypes (or patients) of disease using the selected genes. We employ a kernel machine, kernel Fisher discriminant analysis (KFDA), for discrimination and use the derivatives of the kernel function to perform gene selection. Using a modified form of KFDA in the minimum squared error (MSE) sense and the gradients of the kernel functions, we construct an effective gene selection criterion. We assess the performance of the proposed methodology by applying it to three gene expression datasets: leukemia dataset, breast cancer dataset and colon cancer dataset. Using a few informative genes, the proposed method accurately and reliably classified cancer subtypes (or patients). Also, through a comparison study, we verify the reliability of the gene selection and discrimination results. PMID- 15280024 TI - Biochemical engineering of the N-acyl side chain of sialic acid leads to increased calcium influx from intracellular compartments and promotes differentiation of HL60 cells. AB - Sialylation of glycoconjugates is essential for mammalian cells. Sialic acid is synthesized in the cytosol from N-acetylmannosamine by several consecutive steps. Using N-propanoylmannosamine, a novel precursor of sialic acid, we are able to incorporate unnatural sialic acids with a prolonged N-acyl side chain (e.g., N propanoylneuraminic acid) into glycoconjugates taking advance of the cellular sialylation machinery. Here, we report that unnatural sialylation of HL60-cells leads to an increased release of intracellular calcium after application of thapsigargin, an inhibitor of SERCA Ca2+-ATPases. Furthermore, this increased intracellular calcium concentration leads to an increased adhesion to fibronectin. Finally, we observed an increase of the lectin galectin-3, a marker of monocytic differentiation of HL60-cells. PMID- 15280025 TI - Staurosporine restores GTPgammaS induced block of rapid endocytosis in chromaffin cells. AB - Fast capacitance measurements demonstrated that chromaffin cells retrieve membrane by several kinetically different pathways. Here, we show that rapid endocytosis is blocked and slow endocytosis reduced by intracellular application of GTPgammaS, an activator of G-proteins, but not by the competitive blocker GDPbetaS. The inhibition of rapid endocytosis by GTPgammaS can be restored with GDPbetaS or staurosporine completely. But only staurosporine partially abolishes the reduction of slow endocytosis by GTPgammaS. Besides triggering exocytosis, GTPgammaS elicits large exo- and endocytotic vesicles that contributed significantly to the total membrane traffic, indicating a third pathway of membrane shuttle. PMID- 15280026 TI - Recycling of intact dense core vesicles in neurites of NGF-treated PC12 cells. AB - Exocytic fusion in neuroendocrine cells does not always result in complete release of the peptide contents from dense core vesicles (DCVs). In this study, we use fluorescence imaging and immunoelectron microscopy to examine the retention, endocytosis and recycling of chromogranin B in DCVs of NGF-treated PC12 cells. Our results indicate that DCVs retained and retrieved an intact core that was available for subsequent exocytic release. The endocytic process was inhibited by cyclosporine A or by substitution of extracellular Ca(2+) with Ba(2+) and the total recycling time was less than 5 min. PMID- 15280027 TI - Inhibition of DNA synthesis by K+-stabilised G-quadruplex promotes allelic preferential amplification. AB - PCR preferential amplification consists of the inefficient amplification of one allele in a heterozygous sample. Here, we report the isolation of a GC-rich human minisatellite, MsH43, that undergoes allelic preferential amplification during PCR. This effect requires the existence of a (TGGGGC)(4) motif that is able to form a G-quadruplex in the presence of K(+). This structure interferes with the DNA synthesis of the alleles harbouring this motif during PCR The present results are the first demonstration that the formation of G-quadruplex can be one of the mechanisms involved in some kinds of preferential amplification. PMID- 15280028 TI - The gun4 gene is essential for cyanobacterial porphyrin metabolism. AB - Ycf53 is a hypothetical chloroplast open reading frame with similarity to the Arabidopsis nuclear gene GUN4. In plants, GUN4 is involved in tetrapyrrole biosynthesis. We demonstrate that one of the two Synechocystis sp. PCC 6803 ycf53 genes with similarity to GUN4 functions in chlorophyll (Chl) biosynthesis as well: cyanobacterial gun4 mutant cells exhibit lower Chl contents, accumulate protoporphyrin IX and show less activity not only of Mg chelatase but also of Fe chelatase. The possible role of Gun4 for the Mg as well as Fe porphyrin biosynthesis branches in Synechocystis sp. PCC 6803 is discussed. PMID- 15280029 TI - Oxygen-dependent H2O2 production by Rubisco. AB - Oxygen and ribulose-1,5-bisphosphate dependent, H(2)O(2) production was observed with several wild type Rubisco enzymes using a sensitive assay. H(2)O(2) and d glycero-2,3-pentodiulose-1,5-bisphosphate, a known and potent inhibitor of Rubisco activity, are predicted products arising from elimination of H(2)O(2) from a peroxyketone intermediate, specific to oxygenase activity. Parallel assays using varying CO(2) and O(2) concentrations revealed that the partitioning to H(2)O(2) during O(2) consumption by spinach Rubisco was constant at 1/260-1/270. High temperature (38 degrees C), which reduces Rubisco specificity for CO(2) versus O(2), increased the rates of H(2)O(2) production and O(2) consumption, resulting in a small increase in partitioning to H(2)O(2) (1/210). Two Rubiscos with lower specificity than spinach exhibited greater partitioning to H(2)O(2) during catalysis: Chlamydomonas reinhardtii (1/200); and Rhodospirillum rubrum (1/150). PMID- 15280031 TI - Heterogeneity based on bending of purple membrane containing bacteriorhodopsin. AB - The first and second derivatives of dielectric spectra have evidenced the existence of two interacting states of purple membrane (PM) that respond differently to the intensity of illuminating light providing, this way, underlying consequences to the heterogeneous behavior of bacteriorhodopsin (bR). It is of particular interest to note that the rotational diffusion coefficient of PM has exhibited non-linearity versus light intensity. The explored non-linearity in electrical properties beers, thereby, on changes in PM size. The non-linear variations in PM bending might initiate, in consequence, variations in the dipole moment (permanent and induced) and dc-conductivity of PM patches. Proposal based on PM bending has been introduced to correlate the light intensity effect to the PM lipid environment. Modulation of the global structure of PM and, in turn, its electrical properties by an external perturbation (e.g., light) could be of interest in biotechnological applications based on optoelectronic properties of bR. PMID- 15280030 TI - Hexose-6-phosphate dehydrogenase determines the reaction direction of 11beta hydroxysteroid dehydrogenase type 1 as an oxoreductase. AB - The impact of hexose-6-phosphate dehydrogenase (H6PDH) on 11beta-hydroxysteroid dehydrogenase (11beta-HSD) type 1 activity was investigated upon coexpression in HEK-293 cells. Confocal microscopy analysis indicated colocalisation of both enzymes at the lumenal side of the endoplasmic reticulum (ER) membrane. Functional analysis in intact cells revealed fivefold stimulation of 11beta-HSD1 oxoreductase activity and sixfold decrease of dehydrogenase activity upon coexpression with H6PDH, without changing kinetic parameters in cell lysates. Thus, H6PDH directly determines the reaction direction of 11beta-HSD1 in intact cells as an oxoreductase without changing intrinsic catalytic properties of 11beta-HSD1 by regenerating NADPH in the ER-lumen. PMID- 15280032 TI - Crystal structure and reactivity of YbdL from Escherichia coli identify a methionine aminotransferase function. AB - The ybdL gene of Escherichia coli codes for a protein of unknown function. Sequence analysis showed moderate homology to several vitamin B(6) dependent enzymes, suggesting that it may bind pyridoxal-5'-phosphate. The structure analysis of YbdL to 2.35 A resolution by protein crystallography verifies that it is a PLP dependent enzyme of fold type I, the typical aspartate aminotransferase fold. The active site contains a bound pyridoxal-5'-phosphate, covalently attached to the conserved active site lysine residue Lys236. The pattern of conserved amino acids in the putative substrate binding pocket of the enzyme reveals that it is most closely related to a hyperthermophilic aromatic residue aminotransferase from the archeon Pyrococcus horikoshii. Activity tests with 10 amino acids as amino-donors reveal, however, a preference for Met, followed by His and Phe, results which can be rationalized by modelization studies. PMID- 15280033 TI - An Arabidopsis mutant disrupted in valine catabolism is also compromised in peroxisomal fatty acid beta-oxidation. AB - Characterisation of the Arabidopsis dbr5 mutant, which was isolated on the basis of 2,4-dichlorophenoxybutyric acid (2,4-DB) resistance, revealed that it is disrupted in the CHY1 gene. CHY1 encodes a peroxisomal protein that is 43% identical to the mammalian beta-hydroxyisobutryl-CoA hydrolase of valine catabolism. We show that 2,4-DB resistance and the associated sucrose dependent seedling growth are due to a large activity decrease of 3-ketoacyl-CoA thiolase, which is involved in peroxisomal fatty acid beta-oxidation. (14)C-feeding studies demonstrate that dbr5 and chy1 seedlings are reduced in valine catabolism. These data support the hypothesis that CHY1 plays a key role in peroxisomal valine catabolism and that disruption of this enzyme results in accumulation of a toxic intermediate, methacrylyl-CoA, that inhibits 3-ketoacyl-CoA thiolase activity and thus blocks peroxisomal beta-oxidation. We also show that CHY1 is repressed in seedlings grown on sugars, which suggests that branched chain amino acid catabolism is transcriptionally regulated by nutritional status. PMID- 15280034 TI - Two cAMP receptor proteins with different biochemical properties in the filamentous cyanobacterium Anabaena sp. PCC 7120. AB - Two open reading frames (ORFs), alr0295 and alr2325, are found to encode putative cAMP receptor proteins (CRPs) in the genome of the filamentous cyanobacterium Anabaena sp. PCC 7120. These ORFs were named cAMP receptor protein-like gene A in Anabaena sp. PCC 7120 (ancrpA) and cAMP receptor protein-like gene B in Anabaena sp. PCC 7120 (ancrpB), respectively, and those translated products were investigated. The equilibrium dialysis measurements revealed that AnCrpA bound with cAMP specifically, while AnCrpB bound with both cAMP and cGMP, though the affinity for cGMP was weak. The binding affinity for cAMP of AnCrpA showed the lowest dissociation constant, approximately 0.8 microM, among bacterial CRPs. A gel mobility shift assay elucidated that AnCrpA and AnCrpB formed a complex with the consensus DNA sequence in the presence of cAMP, although AnCrpB did not have ordinary DNA-binding motifs. PMID- 15280035 TI - Human peroxiredoxin 5 is a peroxynitrite reductase. AB - Peroxiredoxins are an ubiquitous family of peroxidases widely distributed among prokaryotes and eukaryotes. Peroxiredoxin 5, which is the last discovered mammalian member, was previously shown to reduce peroxides with the use of reducing equivalents derived from thioredoxin. We report here that human peroxiredoxin 5 is also a peroxynitrite reductase. Analysis of peroxiredoxin 5 mutants, in which each of the cysteine residues was mutated, suggests that the nucleophilic attack on the O-O bond of peroxynitrite is performed by the N terminal peroxidatic Cys(47). Moreover, with the use of pulse radiolysis, we show that human peroxiredoxin 5 reduces peroxynitrite with an unequalled high rate constant of (7+/-3)x10(7) M(-1)s(-1). PMID- 15280036 TI - Specificity determinants for chemokine recognition identified using eotaxin-MCP-1 chimeras. AB - To identify the elements of two chemokines [monocyte chemoattractant protein-1 (MCP-1) and eotaxin] that control their differential recognition by their respective receptors (CCR2 and CCR3), we have studied the receptor interactions of MCP-1-eotaxin chimeras. Each receptor was found to exhibit a distinct binding preference for proteins containing the amino-terminal region of the cognate chemokine for that receptor. However, other elements dictating chemokine preference were different for the two receptors. In some cases, the influence of replacing a particular region was dependent on the identities of neighboring regions, indicating a complex network of cooperative and/or compensating interactions. PMID- 15280037 TI - Identification of ubiquitin-interacting proteins in purified polyglutamine aggregates. AB - Nuclear aggregates of enhanced green fluorescent protein and nuclear localization signal-fused truncated N-terminal huntingtin containing 150 repeats of glutamine residue were purified from ecdysine-inducible mutant neuro2A cell line by sequential extraction of nuclear soluble proteins. To analyze the aggregate interacting proteins, we subjected the nuclear aggregates to high performance liquid chromatography-mass spectrometry analysis. The resulting data revealed the presence of three new putative aggregate-interacting proteins: ubiquilin 1, ubiquilin 2 and Tollip. These proteins also associated with neuronal intranuclear inclusions in a mouse model of Huntington disease (HD). These aggregate interacting proteins contain ubiquitin-interacting motifs, suggesting that they are recruited to the aggregates where they may lose their normal function. PMID- 15280038 TI - INFgamma stimulates arginine transport through system y+L in human monocytes. AB - Freshly isolated human monocytes transport L-arginine mostly through a sodium independent, NEM insensitive pathway inhibited by L-leucine in the presence, but not in the absence of sodium. Interferon-gamma (IFNgamma) stimulates this pathway, identifiable with system y+L, and markedly enhances the expression of SLC7A7, the gene that encodes for system y+L subunit y+LAT1, but not of SLC7A6, that codes for the alternative subunit y+LAT2. System y+ plays a minor role in arginine uptake by monocytes and the expression of system y+-related genes, SLC7A1 and SLC7A2, is not changed by IFNgamma. These results demonstrate that system y+L is sensitive to IFNgamma. PMID- 15280039 TI - Stemar-13-ene synthase, a diterpene cyclase involved in the biosynthesis of the phytoalexin oryzalexin S in rice. AB - In suspension-cultured rice cells, diterpenoid phytoalexins are produced in response to exogenously applied elicitors. We isolated a cDNA encoding a diterpene cyclase, OsDTC2, from suspension-cultured rice cells treated with a chitin elicitor. The OsDTC2 cDNA was overexpressed in Escherichia coli as a fusion protein with glutathione S-transferase, and the recombinant OsDTC2 was indicated to function as stemar-13-ene synthase that converted syn-copalyl diphosphate to stemar-13-ene, a putative diterpene hydrocarbon precursor of the phytoalexin oryzalexin S. The level of OsDTC2 mRNA in suspension-cultured rice cells began to increase 3 h after addition of the elicitor and reached the maximum after 8 h. The expression of OsDTC2 was also induced in UV-irradiated rice leaves. In addition, we indicated that stemar-13-ene accumulated in the chitin-elicited suspension-cultured rice cells and the UV-irradiated rice leaves. PMID- 15280040 TI - Feeding induces expression of heat shock proteins that reduce oxidative stress. AB - Heat shock proteins (Hsps) are induced in response to various kinds of environmental and physiological stresses. However, it is unclear whether Hsps play roles in protecting cells in the digestive organs against xenobiotic chemicals. Here, we found that feeding induces expression of a set of Hsps specifically in the mouse liver and intestine by activating heat shock transcription factor 1 (HSF1). In the liver, HSF1 is required to suppress toxic effects of electrophiles, which are xenobiotic chemicals causing oxidative stress. We found that overexpression of Hsp27, which elevates cellular glutathione level, promotes survival of culture cells exposed to electrophiles. These results suggest a novel mechanism of cell protection against xenobiotic chemicals in the food. PMID- 15280041 TI - Identification of the catalytic residues in the double-zinc aminopeptidase from Streptomyces griseus. AB - The aminopeptidase from Streptomyces griseus (SGAP) has been cloned and expressed in Escherichia coli. By growing the cells in the presence of 1 M sorbitol at 18 degrees C, the protein was obtained in a soluble and active form. The amino acid sequence of the recombinant SGAP contained four amino acids differing from the previously published sequence. Re-sequencing of the native protein indicated that asparagines 70 and 184 are in fact aspartic acids as in the recombinant protein. Based on the crystal structure of SGAP, Glu131 and Tyr246 were proposed to be the catalytic residues. Replacements of Glu131 resulted in loss of activity of 4-5 orders of magnitude, consistent with Glu131 acting as the general base residue. Mutations in Tyr246 resulted in about 100-fold reduction of activity, suggesting that this residue is involved in the stabilization of the transition state intermediate. PMID- 15280042 TI - Prostatic acid phosphatase degrades lysophosphatidic acid in seminal plasma. AB - Lysophosphatidic acid (LPA) is a lipid mediator with multiple biological activities and is detected in various biological fluids, including human seminal plasma. Due to its cell proliferation stimulatory and anti-apoptotic activities, LPA has been implicated in the progression of some cancers such as ovarian cancer and prostate cancer. Here, we show that prostatic acid phosphatase, which is a non-specific phosphatase and which has been implicated in the progression of prostate cancer, inactivates LPA in human seminal plasma. Human seminal plasma contains both an LPA-synthetic enzyme, lysoPLD, which converts lysophospholipids to LPA and is responsible for LPA production in serum, and its major substrate, lysophosphatidylcholine. In serum, LPA accumulated during incubation at 37 degrees C. However, in seminal plasma, LPA did not accumulate. This discrepancy is explained by the presence of a strong LPA-degrading activity. Incubation of LPA with seminal plasma resulted in the disappearance of LPA and an accompanying accumulation of monoglyceride showing that LPA is degraded by phosphatase activity present in the seminal plasma. When seminal plasma was incubated in the presence of a phosphatase inhibitor, sodium orthovanadate, LPA accumulated, indicating that LPA is produced and degraded in the fluid. Biochemical characterization of the LPA-phosphatase activity identified two phosphatase activities in human seminal plasma. By Western blotting analysis in combination with several column chromatographies, the major activity was revealed to be identical to prostatic acid phosphatase. The present study demonstrates active LPA metabolism in seminal plasma and indicates the possible role of LPA signaling in male sexual organs including prostate cancer. PMID- 15280043 TI - On the disulfide-linker strategy for designing efficacious cationic transfection lipids: an unexpected transfection profile. AB - Herein, employing a previously reported disulfide-linker strategy, we have designed and synthesized a novel cationic lipid 2 with a disulfide-linker and its non-disulfide control analog lipid 1. The relative efficacies of lipids 1 and 2 in transfecting CHO, COS-1 and MCF-7 cells were measured using both reporter gene and whole cell histochemical staining assays. In stark contrast to the expectation based on the disulfide-linker strategy, the control non-disulfide cationic lipid 1 showed phenomenally superior in vitro transfection efficacies to its essentially transfection incompetent disulfide counterpart lipid 2. Results in DNase I protection experiments and the electrophoretic gel patterns in the presence of glutathione, taken together, are consistent with the notion that the success of the disulfide-linker strategy may depend more critically on the DNase I sensitivity of the lipoplexes than on the efficient DNA release induced by intracellular glutathione pool. PMID- 15280044 TI - Nucleoside diphosphate kinase of Mycobacterium tuberculosis acts as GTPase activating protein for Rho-GTPases. AB - Several bacterial pathogens secrete proteins into the host cells that act as GTPase-activating proteins (GAPs) for Rho-GTPases and convert GTP-bound active form to GDP-bound inactive form. However, no such effector molecule has been identified in Mycobacterium tuberculosis. In this study, we show that culture supernatant of M. tuberculosis H(37)Rv harbors a protein that stimulates the conversion of GTP-bound Rho-GTPases to the GDP-bound form. Nucleoside diphosphate kinase (Ndk) was identified as this culture supernatant protein that stimulated in vitro GTP hydrolysis by members of Rho-GTPases. The histidine-117 mutant of Ndk, which is impaired for autophosphorylation and nucleotide-binding activities, shows GAP activity. These results suggest that Ndk of M. tuberculosis functions as a Rho-GAP to downregulate Rho-GTPases, and this activity may aid in pathogenesis of the bacteria. PMID- 15280045 TI - The bacterial type III secretion system-associated pilin HrpA has an unusually long mRNA half-life. AB - Secondary structures affect mRNA stability and may play a role in protein secretion. We have studied the mRNA of hrpA, which codes for the major structural unit of the type III secretion system-associated pilus of Pseudomonas syringae pv. tomato, Erwinia carotovora and Pseudomonas syringae pv. phaseolicola. We show that hrpA mRNA has an unusually long half-life, approximately 33-47 min. We mapped regions in the transcript that affected hrpA mRNA accumulation. Apparently, sequences at both 5' and 3' ends affect accumulation. Altering the hypothetical, stable GC rich loop structure in the 3' end of the transcript decreased transcript levels. PMID- 15280046 TI - Localization and mutagenesis of the sorting signal binding site on sortase A from Staphylococcus aureus. AB - Surface proteins in Gram-positive bacteria are anchored to the cell wall by the action of sortase enzymes. The Staphylococcus aureus sortase A (SrtA) protein anchors proteins by recognizing a cell wall sorting signal containing the amino acid sequence LPXTG. To understand how SrtA binds this sequence, we carried out NMR studies of new peptidyl-cyanoalkene and peptidyl-sulfhydryl inhibitors that contain the sorting signal sequence LPAT. These studies combined with amino acid mutagenesis identified a catalytically important and conserved binding surface formed by residues A118, T180, and I182. Compatible with its recently proposed role as a general base, R197 is also shown to be required for catalysis. PMID- 15280047 TI - Accumulation of the common mitochondrial DNA deletion induced by ionizing radiation. AB - Point mutations and deletions in mitochondrial DNA (mtDNA) accumulate as a result of oxidative stress, including ionizing radiation. As a result, dysfunctional mitochondria suffer from a decline in oxidative phosphorylation and increased release of superoxides and other reactive oxygen species (ROS). Through this mechanism, mitochondria have been implicated in a host of degenerative diseases. Associated with this type of damage, and serving as a marker of total mtDNA mutations and deletions, the accumulation of a specific 4977-bp deletion, known as the common deletion (Delta-mtDNA(4977)), takes place. The Delta-mtDNA(4977) has been reported to increase with age and during the progression of mitochondrial degeneration. The purpose of this study was to investigate whether ionizing radiation induces the formation of the common deletion in a variety of human cell lines and to determine if it is associated with cellular radiosensitivity. Cell lines used included eight normal human skin fibroblast lines, a radiosensitive non-transformed and an SV40 transformed ataxia telangiectasia (AT) homozygous fibroblast line, a Kearns Sayre Syndrome (KSS) line known to contain mitochondrial deletions, and five human tumor lines. The Delta-mtDNA(4977) was assessed by polymerase chain reaction (PCR). Significant levels of Delta-mtDNA(4977) accumulated 72 h after irradiation doses of 2, 5, 10 or 20 Gy in all of the normal lines with lower response in tumor cell lines, but the absolute amounts of the induced deletion were variable. There was no consistent dose-response relationship. SV40 transformed and non-transformed AT cell lines both showed significant induction of the deletion. However, the five tumor cell lines showed only a modest induction of the deletion, including the one line that was deficient in DNA damage repair. No relationship was found between sensitivity to radiation-induced deletions and sensitivity to cell killing by radiation. PMID- 15280048 TI - Efficient generation of transgenic pigs using equine infectious anaemia virus (EIAV) derived vector. AB - Traditional methods of transgene delivery in livestock are inefficient. Recently, human immunodeficiency virus (HIV-1) based lentiviral vectors have been shown to offer an efficient transgene delivery system. We now extend this method by demonstrating efficient generation of transgenic pigs using an equine infectious anaemia virus derived vector. We used this vector to deliver a green fluorescent protein expressing transgene; 31% of injected/transferred eggs resulted in a transgenic founder animal and 95% of founder animals displayed green fluorescence. This compares favourably with results using HIV-1 based vectors, and is substantially more efficient than the standard pronuclear microinjection method, indicating that lentiviral transgene delivery may be a general tool with which to efficiently generate transgenic mammals. PMID- 15280049 TI - Lowered glucose suppressed the proliferation and increased the differentiation of murine neural stem cells in vitro. AB - Cerebral ischemia is known to activate endogenous neural stem cells (NSCs), but its mechanisms remain unknown. Since lowered glucose supply seems to mediate ischemic actions, we examined the effect of low glucose on NSC activities in vitro. Low glucose applied during the proliferation period diminished EGF-induced proliferation of NSCs without affecting subsequent differentiation, but low glucose directly exposed during the differentiation period facilitated the differentiation of NSCs into neurons and astrocytes. These findings suggest that low glucose facilitated NSC differentiation, but it diminished NSC proliferation. Moreover, the effect of low glucose may be dependent on the timing of application. PMID- 15280050 TI - GenAge: a genomic and proteomic network map of human ageing. AB - The aim of this work was to provide an overview of the genetics of human ageing to gain novel insights about the mechanisms involved. By incorporating findings from model organisms to humans, such as mutations that either delay or accelerate ageing in mice, we constructed the gene networks previously related to ageing: namely, the network related to DNA metabolism and the network involving the GH/IGF-1 axis. Gathering data about the interacting partners of these proteins allowed us to suggest the involvement in ageing of a number of proteins through a "guilt-by-association" methodology. To organize our data, we developed the first curated database of genes related to human ageing: GenAge. With over 200 entries, GenAge may serve as a reference database of genes related to human ageing. Moreover, we rendered the first proteomic network map of human ageing, which suggests a relationship between the genetics of development and the genetics of ageing. Our work serves as a framework upon which a systems-biology understanding of ageing can be developed. GenAge is freely available for academic purposes at: http://genomics.senescence.info/genes/. PMID- 15280051 TI - Synthesis of lactoside glycodendrons using photoaddition and reductive amination methodologies. AB - Carbohydrate-based divalent and tetravalent lactoside glycodendrons were constructed in a convergent manner. The dendrons were synthesized beginning with the photoaddition of hepta-O-acetyl-1-thio-beta-lactose, in an anti-Markovnikov manner, to a bis-allyl AB2 trisaccharide to form a divalent dendron. Following two nearly quantitative deprotection steps, the divalent lactoside was coupled to another AB2 trisaccharide by reductive amination to afford a tetravalent dendron. These paucivalent compounds were characterized by NMR spectroscopy and mass spectrometry. PMID- 15280052 TI - Synthesis and co-polymerization of an unsaturated 1,5-anhydro-D-fructose derivative. AB - An unsaturated derivative, 3,6-di-O-acetyl-1,5-anhydro-4-deoxy-D-glycero-hex-3 enopyranos-2-ulose (3), was obtained via regioselective elimination and acetylation of monohydrated 1,5-anhydro-D-fructose (1) in a single step reaction. High yield (80%) was achieved without any dimeric by-products. Its co polymerization to saccharide polymers was investigated with different commercial vinyl co-monomers. Co-polymers were obtained and characterized. PMID- 15280053 TI - Synthesis and biological activities of octyl 2,3-di-O-sulfo-alpha-L-fucopyranosyl (1-->3)-2-O-sulfo-alpha-L-fucopyranosyl-(1-->4)-2,3-di-O-sulfo-alpha-L fucopyranosyl-(1-->3)-2-O-sulfo-alpha-L-fucopyranosyl-(1-->4)-2,3-di-O-sulfo-beta L-fucopyranoside. AB - Octyl 2,3-di-O-sulfo-alpha-L-fucopyranosyl-(1-->3)-2-O-sulfo-alpha-L fucopyranosyl-(1-->4)-2,3-di-O-sulfo-alpha-L-fucopyranosyl-(1-->3)-2-O-sulfo alpha-L-fucopyranosyl-(1-->4)-2,3-di-O-sulfo-beta-L-fucopyranoside, a fucosyl pentasaccharide with a regular structure resembling the repeating unit of a natural sulfated fucan, was chemically synthesized using a convergent '2+3' strategy. Regioselective 3-O-silylation of beta-thiofucopyranoside and AgOTf catalyzed glycosylation of the protected glycosyl trichloroacetimidate facilitated a one-pot trisaccharide synthesis. The synthesized target compound showed good antitumor activity in vivo, and promising anticoagulant activity in vitro. PMID- 15280055 TI - Deoxy and deoxyfluoro analogues of acetylated methyl beta-D-xylopyranoside- substrates for acetylxylan esterases. AB - Four modified substrates for acetylxylan esterases, 2-deoxy, 3-deoxy, 2-deoxy-2 fluoro, and 3-deoxy-3-fluoro derivatives of di-O-acetylated methyl beta-D xylopyranoside were synthesized via 2,3-anhydropentopyranoside precursors. Methyl 2,3-anhydro-4-O-benzyl-beta-D-ribopyranoside was transformed into methyl 2,3 anhydro-4-O-benzyl-beta-D-lyxopyranoside in three steps. The epoxide ring opening of 2,3-anhydropentopyranosides was accomplished either by hydride reduction or hydrofluorination. Methyl beta-D-xylopyranoside 2,3,4-tri-O-, 2,4-di-O-, and 3,4 di-O-acetates, and the prepared diacetate analogues were tested as substrates of acetylxylan esterases from Schizophyllum commune and Trichoderma reesei. Measurement of their rate of deacetylation pointed to unique structural requirements of the enzymes for the substrates. The enzymes differed particularly in the requirement for the trans vicinal hydroxy group in the deacetylation at C 2 and C-3 and in the tolerance to the presence of trans vicinal acetyl groups esterifying the OH group at C-2 and C-3. PMID- 15280054 TI - Accessibility of N-acyl-D-mannosamines to N-acetyl-D-neuraminic acid aldolase. AB - N-Acetyl-D-neuraminic acid (NeuNAc) aldolase is an important enzyme for the metabolic engineering of cell-surface NeuNAc using chemically modified D mannosamines. To explore the optimal substrates for this application, eight N acyl derivatives of D-mannosamine were prepared, and their accessibility to NeuNAc aldolase was quantitatively investigated. The N-propionyl-, N-butanoyl-, N iso-butanoyl-, N-pivaloyl-, and N-phenylacetyl-D-mannosamines proved to be as good substrates as, or even better than, the natural N-acetyl-D-mannosamine, while the N-trifluoropropionyl and benzoyl derivatives were poor. It was proposed that the electronic effects might have a significant influence on the enzymatic aldol condensation reaction of D-mannosamine derivatives, with electron-deficient acyl groups having a negative impact. The results suggest that N-propionyl-, N butanoyl-, N-iso-butanoyl-, and N-phenylacetyl-D-mannosamines may be employed to bioengineer NeuNAc on cells. PMID- 15280056 TI - Crystal architecture and conformational properties of the inclusion complex, neohesperidin dihydrochalcone-cyclomaltoheptaose (beta-cyclodextrin), by X-ray diffraction. AB - The crystal structure of the host-guest noncovalent complex of cyclomaltoheptaose (beta-cyclodextrin, betaCD) with the O-diglycosyl flavonoid neohesperidin dihydrochalcone [(3,5-dihydroxy-4-(3-hydroxy-4-methoxyhydrocinnamoyl)phenyl-2-O (alpha-L-rhamnopyranosyl)-beta-D-glucopyranoside, NDC] has been determined from single-crystal X-ray diffraction data collected at low temperature (130 K), using synchrotron radiation. The crystal data are as follows: a =15.125(5), b =30.523(5), c =41.332(5) Angstroms, orthorhombic, space group C222(1). The structure contains 19 molecules of water, of which 11 appeared well positioned, whereas 9 are disordered over 23-positions. The betaCD-NDC complex is characterized by one aromatic part of NDC deeply inserted into the hydrophobic cavity of the betaCD through the primary OH rim, and it is present in the crystal as a dimer. The dimeric units, formed by head-to-head assemblies of CD molecules, each with its guest, are self-assembled in columns. The stability of the columns is provided by host-guest and guest-guest attractive interactions, thus showing a key role of the guest molecules in the crystal architecture. The guest conformation in the complex is different from that reported in the literature for uncomplexed NDC. The host-induced conformational changes on NDC provide the optimum geometry requirements for the assembly of the dimeric units. PMID- 15280057 TI - The influence of mutanase and dextranase on the production and structure of glucans synthesized by streptococcal glucosyltransferases. AB - Glucanohydrolases, especially mutanase [alpha-(1-->3) glucanase; EC 3.2.1.59] and dextranase [alpha-(1-->6) glucanase; EC 3.2.1.11], which are present in the biofilm known as dental plaque, may affect the synthesis and structure of glucans formed by glucosyltransferases (GTFs) from sucrose within dental plaque. We examined the production and the structure of glucans synthesized by GTFs B (synthesis of alpha-(1-->3)-linked glucans) or C [synthesis of alpha-(1-->6)- and alpha-(1-->3)-linked glucans] in the presence of mutanase and dextranase, alone or in combination, in solution phase and on saliva-coated hydroxyapatite beads (surface phase). The ability of Streptococcus sobrinus 6715 to adhere to the glucan, which was formed in the presence of the glucanohydrolases was also explored. The presence of mutanase and/or dextranase during the synthesis of glucans by GTF B and C altered the proportions of soluble to insoluble glucan. The presence of either dextranase or mutanase alone had a modest effect on total amount of glucan formed, especially in the surface phase; the glucanohydrolases in combination reduced the total amount of glucan. The amount of (1-->6)-linked glucan was reduced in presence of dextranase. In contrast, mutanase enhanced the formation of soluble glucan, and reduced the percentage of 3-linked glucose of GTF B and C glucans whereas dextranase was mostly without effect. Glucan formed in the presence of dextranase provided fewer binding sites for S. sobrinus; mutanase was devoid of any effect. We also noted that the GTFs bind to dextranase and mutanase. Glucanohydrolases, even in the presence of GTFs, influence glucan synthesis, linkage remodeling, and branching, which may have an impact on the formation, maturation, physical properties, and bacterial binding sites of the polysaccharide matrix in dental plaque. Our data have relevance for the formation of polysaccharide matrix of other biofilms. PMID- 15280058 TI - Metal-N-saccharide chemistry: synthesis and structure determination of two Cu(II) complexes containing glycosylamines. AB - Two new complexes [Cu(N,N',N"-(D-Glc)3-tren)Cl]Cl (1) and [Cu(N,N',N"-(maltose) tren)]Cl2.H2O (2), have been synthesized and characterized by elementary analysis, and the IR and UV spectra suggest that complex 1 and complex 2 are arranged in trigonal bipyramidal configuration and square-pyramidal configuration, respectively. The crystal structure of complex 1 has been determined by X-ray diffraction as: a = 9.3476(8), b = 17.4236(13), c = 9.7836(8) angstroms, beta = 91.197 degrees, and V = 1593.1(2) Angstroms3, Z = 2, and R = 0.0325, which shows that three secondary amine groups (N-1, N-2, N-3) of the glycosylamine ligand forms the equatorial plane, and the tertiary amine (N-4) and one Cl- are located at the apical positions. PMID- 15280059 TI - Structure of the O-polysaccharide of Proteus serogroup O34 containing 2-acetamido 2-deoxy-alpha-D-galactosyl phosphate. AB - On mild acid degradation of the lipopolysaccharide of Proteus vulgaris O34, strain CCUG 4669, the O-polysaccharide was cleaved at a glycosyl-phosphate linkage that is present in the main chain. The resultant phosphorylated oligosaccharides and an alkali-treated lipopolysaccharide were studied by sugar and methylation analyses along with 1H and 13C NMR spectroscopy, and the following structure of the branched tetrasaccharide phosphate repeating unit of the O-polysaccharide was established: [carbohydrate structure: see text]The O polysaccharide of Proteus mirabilis strain TG 276 was found to have the same structure and, based on the structural and serological data, this strain was proposed to be classified into the same Proteus serogroup O34. PMID- 15280064 TI - Decrease in tumor apparent permeability-surface area product to a MRI macromolecular contrast medium following angiogenesis inhibition with correlations to cytotoxic drug accumulation. AB - BACKGROUND: New strategies for cancer therapy include the combination of angiogenesis inhibitors with cytotoxins. However, angiogenesis inhibitors may alter tumor microvessel structure and transendothelial permeability thereby reducing tumoral delivery of cytotoxic agents. The aim of this study was to estimate quantitatively the apparent permeability-surface area product (K(PS)) in tumors to a macromolecular contrast medium (MMCM), to follow changes in K(PS) induced by antibodies to vascular endothelial growth factor (anti-VEGF), and to correlate the findings with tumor accumulation of cisplatin, a highly protein bound cytotoxin, and 5-fluorouracil (5-FU), a small unbound cytotoxin. METHODS: Dynamic MRI enhanced with a MMCM (albumin-(Gd-DTPA)(30)) was analyzed using a two compartment tumor tissue model (plasma and interstitial water) to quantitatively estimate K(PS). These estimates of K(PS) were correlated with cytotoxic drug accumulations in the tumors. RESULTS: Anti-VEGF treatment reduced K(PS) to MMCM in tumor tissue from 0.013 mL h(-1) cm(-3) (n = 9) at baseline to 0.003 mL h(-1) cm(-3) (n = 9) 24 h later (p <.05). The K(PS) values correlated significantly (r(2) =.78; p <.0001) with the tumor cisplatin accumulation. No correlation (r(2) =.001; p =.89) was found between K(PS) and tumor accumulation of the substantially smaller 5-FU molecule. CONCLUSIONS: MMCM-enhanced MRI can be used to detect and estimate changes in K(PS) to this contrast agent following a single dose of anti-VEGF antibody. The decline in K(PS) induced by this inhibitor of angiogenesis is associated with reduced tumor concentration of a protein-bound cytotoxin, similar in molecular weight to the contrast agent. MRI assays of microvascular status as performed here may be useful to clinically monitor responses to anti-angiogenesis drugs and to optimize the choice and timing of cytotoxic drug administration. PMID- 15280065 TI - Independence of connexin expression and vasomotor conduction from sympathetic innervation in hamster feed arteries. AB - OBJECTIVE: Vasomotor responses can travel along the wall of resistance microvessels by two distinct mechanisms: cell-to-cell conduction through gap junctions or the release of neurotransmitter along perivascular nerves. It is unknown whether vascular innervation influences the expression of connexin molecules which comprise gap junctions, or the conduction of vasomotor responses. In feed arteries of the hamster retractor muscle (RFA), the authors tested whether sympathetic denervation would alter the expression of connexin isoforms and the conduction of vasomotor responses. METHODS: Using intact vessels with sympathetic innervation and those 7-8 days following denervation surgery, mRNA expression was quantified using real-time PCR, cellular localization of Cx protein was characterized using immunohistochemistry, and vasomotor responses to dilator and constrictor stimuli were evaluated in isolated pressurized RFA. RESULTS: Connexin protein localization and mRNA expression were similar between innervated and denervated vessels. mRNA levels were Cx43 = Cx37 > Cx45 >> Cx40. Vasodilation to acetylcholine conducted >/=2000 microm along innervated and denervated vessels, as did the biphasic conduction of vasoconstriction and vasodilation in response to KCl. Vasoconstriction to phenylephrine conducted <500 microm and was attenuated (p <.05) in denervated vessels. CONCLUSIONS: The profile of connexin expression and the conduction of vasomotor responses are largely independent of sympathetic innervation in feed arteries of the hamster retractor muscle (RFA). PMID- 15280066 TI - Effect of venous air embolization on pulmonary microvascular protein permeability. AB - OBJECTIVE: An increase in pulmonary lymph flow and lymph protein clearance following pulmonary air embolization has been interpreted as evidence of increased pulmonary microvascular permeability. The authors hypothesized that air embolization does not alter the pulmonary microvascular permeability to protein and that this could be demonstrated by determining the effect of air embolization on the pulmonary solvent drag reflection coefficient (sigma(f)). METHODS: Anesthetized dogs were instrumented with left atrial balloon-tipped catheters, pulmonary lymphatic cannulae, and inferior vena caval catheters. Values were determined for pulmonary lymph flow (Q(L)) and the lymph-to-plasma protein concentration ratio (C(L)/C(P)) at baseline, after C(L)/C(P) was decreased to a filtration independent value by raising left atrial pressure via progressive balloon inflation and after 2 h of air embolization into the inferior vena cava with continued left atrial hypertension. RESULTS: Q(L) increased and C(L)/C(P) decreased to a filtration-independent value following induction of left atrial hypertension. Air embolization induced during left atrial hypertension resulted in no significant change in C(L)/C(P). The authors were unable to demonstrate that sigma(f) changed following pulmonary air embolization. CONCLUSIONS: Utilizing the washdown technique, the authors could not find any evidence that pulmonary microvascular protein permeability is altered by air embolization. PMID- 15280067 TI - Nitric oxide measurements in rat mesentery reveal disrupted venulo-arteriolar communication in diabetes. AB - OBJECTIVE: Arteriolar tone is partially controlled by diffusing mediators released by closely paired venules and is reported to depend on venular shear and venular leukocyte adherence. In healthy rat mesentery, venule-initiated arteriolar dilation and consequent enhanced capillary flow appear to be tightly regulated by nitric oxide (NO). In contrast, diabetes inhibits NO-dependent vasodilation and is associated with dysfunctional microcirculation. The objective of this study was to investigate venule-dependent NO in diabetes. METHODS: Arteriolar and venular wall concentrations of NO were measured in control and diabetic (streptozotocin-induced) rat mesentery with fluorescent diaminofluorescein-2-diacetate (DAF-2-DA); tissue NO was measured with DAF-2. Venular leukocyte adherence and microvascular shear rates were also measured. RESULTS: Microvascular NO in diabetic rats was found to be significantly lower (<50%) than in controls. In normal rats, arteriolar NO demonstrated a positive correlation with venular NO and venular shear, and a negative correlation with venular leukocyte adherence. Diabetes eliminated all these correlations. No correlation was present between arteriolar NO and arteriolar shear in either normal or diabetic rats. CONCLUSIONS: Arteriolar NO appears to be enhanced by venular shear in normal but not in diabetic rats. This dysfunction could contribute to poor capillary perfusion in diabetes. PMID- 15280068 TI - Contribution of active membrane processes to conducted hyperpolarization in arterioles of hamster cheek pouch. AB - OBJECTIVE: Conduction of vasodilation triggered by acetylcholine (ACh) in arteriolar networks reflects hyperpolarization and its spread from cell to cell along the vessel wall. The amplitude and distance of the vasomotor response appear greater than can be explained by simple passive decay of the electrical signal. The authors tested the hypothesis that the conduction of hyperpolarization involves active membrane processes as the signal travels along the arteriolar wall. METHODS: Intracellular recordings of membrane potential were made from either the smooth muscle or endothelial cell layer of arterioles of the hamster cheek pouch in vivo. Acetylcholine was delivered onto an arteriole using microiontophoresis at defined distances from the recording site, and transient hyperpolarizations were recorded. The area enclosed by the transients (voltage x time integral below baseline) was measured and compared to the area expected if the hyperpolarization was spreading passively. RESULTS: In 11 of 15 recordings from smooth muscle and 5 of 7 from endothelium, areas of the transients were larger than expected for purely passive spread of the electrical signal. CONCLUSIONS: Conduction of hyperpolarization is enhanced by active membrane processes as the signal travels along the arteriolar wall. Signal augmentation will promote blood flow to tissue regions from which hyperpolarization of arterioles originates. PMID- 15280069 TI - Functional link between ETB receptors and eNOS maintain tissue oxygenation in the normal liver. AB - OBJECTIVE: Endothelins and their receptors play a crucial role in regulating liver microcirculation in pathophysiological conditions. The authors investigated the functional significance of the coupling of ET(B) receptors and eNOS in maintaining regional perfusion and tissue oxygenation in the normal liver. METHODS: The effect of endothelin-1 or the ET(B) agonist IRL1620 on oxygen consumption was determined in isolated perfused liver and isolated hepatocytes. Oxygen delivery to the liver tissue was determined in vivo. Following eNOS or iNOS blockade, either ET-1 or IRL1620 was infused via the portal vein. Hepatic tissue oxygenation, redox state, and microcirculation were investigated by intravital microscopy. Injury was estimated by serum LDH. RESULTS: Although IRL1620 and endothelin-1 increased oxygen consumption in isolated hepatocytes, in intact liver, endothelin decreased oxygen consumption while IRL1620 produced no change. In vivo, ET(B) stimulation modestly altered hepatic tissue P(O(2)), redox potential, and microcirculation. eNOS inhibition and ET(B) activation dramatically reduced microcirculatory blood flow, oxygen supply, and increased LDH release. Inhibition of iNOS resulted in small but not significant changes in these parameters. Concomitant ET(A)/ET(B) receptor activation increased microcirculatory failure and decreased tissue oxygen even without NOS inhibition. In contrast, hepatocellular injury was significantly increased following eNOS inhibition. CONCLUSIONS: Coupling between ET(B) receptor stimulation and eNOS activation decreases sinusoidal constriction and plays a functionally important role in maintaining microcirculation and tissue oxygenation in the normal liver. PMID- 15280070 TI - Microcirculatory heterogeneity in the rat small intestine during compromised flow conditions. AB - OBJECTIVE: To characterize and compare microcirculatory changes in the rat small intestinal tissue layers when temporal or spatial microvascular perfusion heterogeneity is present. METHODS: Intravital videomicroscopy with an orthogonal polarization spectral imaging technique was used to visualize the microcirculation of the intestinal mucosa and the longitudinal muscle during systemic (hemorrhagic shock, endotoxemia, and nitric oxide synthesis inhibition) and local (ischemia-reperfusion) circulatory disorders. The average capillary red blood cell velocity (A-RBCV) was calculated from the relative durations of the observed velocity or as a function of the perfused area, respectively. RESULTS: During hemorrhagic hypotension/resuscitation, timewise (flowmotion) and spatial heterogeneity were found to evolve in the mucosal villi and muscle layer. During resuscitation, the A-RBCV decreased by 40% in the villi and by 60% in the muscle. Reperfusion after a 30-min period of mesenteric ischemia caused a 20% reduction in A-RBCV in both layers, while endotoxin infusion caused a temporary 20% decrease in the mucosa, and a persistent, >25% decrease in the muscle. Nitric oxide synthesis inhibition resulted in spatial heterogeneity within the villi and in a 40% decrease in A-RBCV in both structures. CONCLUSIONS: Calculations of timewise variability and of heterogeneity within and between layers can be used for the comparison of distinct intramural microcirculatory changes. PMID- 15280071 TI - Measurement of muscle microvascular oxygen pressures: compartmentalization of phosphorescent probe. AB - OBJECTIVE: To determine whether the phosphorescent probe Oxyphor R2 (a palladium porphyrin dendrimer) becomes extravasated within normotensive skeletal muscle, R2 perfusion and washout studies were performed using a perfused rat hindlimb preparation. METHODS: Phosphorescence signals were monitored in tibialis anterior muscles after 35 min of R2 blood perfusion and across a subsequent washout period that included vasodilation (sodium nitroprusside, SNP, approximately 3 x 10(-2) M). RESULTS: Two responses were evident: Group 1 (n = 4)--Inflowing blood pressure and vascular conductance remained stable close to initial values and subsequently a marked vasodilation was evident with SNP (vascular conductance; R2 blood perfusion, 0.096 +/- 0.005; washout, pre-SNP, 0.085 +/- 0.005, post-SNP, 0.110 +/- 0.005 mL/min/mmHg, p <.05, for pre- vs. post-SNP). Baseline phosphorescence signals could be monitored up to 99 +/- 36 s post-SNP when the phosphorescence signal disappeared. For these muscles, palladium content was undetectable. Group 2 (n = 3)--Inflowing blood pressure increased 112% and vascular conductance fell approximately 50%. These hindlimbs were unresponsive to SNP, phosphorescence signal was undiminished by washout and SNP, and muscles became edematous. CONCLUSIONS: These results suggest that in normotensive muscle (i.e., Group 1 above), extravasation of phosphorescent probe R2 over 35 min of perfusion is insufficient to yield a detectable phosphorescence signal in skeletal muscle. PMID- 15280072 TI - Acetylcholine-induced vasodilation and reactive hyperemia are not affected by acute cyclo-oxygenase inhibition in human skin. AB - OBJECTIVE: To examine whether prostaglandins are involved in endothelium dependent vasodilatory responses of the skin microcirculation. METHODS: Twenty three young male volunteers were studied on 2 different days 1-3 weeks apart. On each experimental day the forearm skin blood flow response to iontophoretically applied acetylcholine (Ach, an endothelium-dependent vasodilator) was determined with laser Doppler imaging following the intravenous administration of either the cyclo-oxygenase inhibitor lysine acetylsalicylate (L-AS), 900 mg, or the oral intake of indomethacin, 75 mg. Acetylcholine was iontophoresed both in presence and in absence of surface anesthesia. In some subjects, the effects of L-AS on skin reactive hyperemia were also assessed. RESULTS: Acute cyclo-oxygenase inhibition with either drug influenced neither the skin blood flow response to 4 different doses of Ach (0.28, 1.4, 7, and 14 mC/cm2) nor reactive hyperemia. The peak vasodilatory response to Ach was significantly increased by skin anesthesia, regardless of whether the subjects received the cyclo-oxygenase inhibitor or not. For example, the mean response (+/-SD) to the largest dose of Ach (tested in 6 subjects, expressed in perfusion units) were as follows: in absence of anesthesia: L-AS 339 +/- 105, placebo 344 +/- 68; with anesthesia: L-AS 453 +/- 76, placebo 452 +/- 65 (p <.01 for effect of anesthesia). CONCLUSIONS: These data give no support for a contribution of prostaglandins to acetylcholine-induced vasodilation or to reactive hyperemia in the skin microcirculation. In this vascular bed, local anesthesia seems to amplify acetylcholine-induced vasodilation via a prostaglandin-independent mechanism. PMID- 15280073 TI - Impact of the Fahraeus effect on NO and O2 biotransport: a computer model. AB - Nitric oxide (NO) and oxygen (O2) transport in the microcirculation are coupled in a complex manner, since enzymatic production of NO depends on O2 availability, NO modulates vascular tone and O2 delivery, and tissue O2 consumption is reversibly inhibited by NO. The authors investigated whether NO bioavailability is influenced by the well-known Fahraeus effect, which has been observed for over 70 years. This phenomenon occurs in small-diameter blood vessels, where the tube hematocrit is reduced below systemic hematocrit as a plasma boundary layer forms near the vascular wall when flowing red blood cells (rbcs) migrate toward the center of the bloodstream. Since hemoglobin in the bloodstream is thought to be the primary scavenger of NO in vivo, this might have a significant impact on NO transport. To investigate this possibility, the authors developed a multilayered mathematical model for mass transport in arterioles using finite element numerical methods to simulate coupled NO and O2 transport in the blood vessel lumen, plasma layer, endothelium, vascular wall, and surrounding tissue. The Fahraeus effect was modeled by varying plasma layer thickness while increasing core hematocrit based on conservation of mass. Key findings from this study are that (1) despite an increase in the NO scavenging rate in the core with higher hematocrit, the model predicts enhanced vascular wall and tissue NO bioavailability due to the relatively greater resistance for NO diffusion through the plasma layer; (2) increasing the plasma layer thickness also increases the resistance for O2 diffusion, causing a larger P(O2) gradient near the vascular wall and decreasing tissue O2 availability, although this can be partially offset with inhibition of O2 consumption by higher tissue NO levels; (3) the Fahraeus effect can become very significant in smaller blood vessels (diameters <30 microm); and (4) models that ignore the Fahraeus effect may underestimate NO concentrations in blood and tissue. PMID- 15280074 TI - Retention of albumin in the circulation is governed by saturable renal cell mediated processes. AB - OBJECTIVE: To examine the role of renal processing in the turnover of circulating albumin in the model of reversible overload proteinuria. Specifically, this study determines whether proteinuria due to hyperalbuminemia may be attributed to saturation of renal cell processing of albumin or alterations in diffusive/convective transport of albumin across the glomerular capillary wall. METHODS: Overload proteinuria was induced in male Wistar rats by daily intraperitoneal injections of 1.5 g bovine serum albumin (BSA; endotoxin free) for 3 days. Plasma concentration and urinary excretion of total protein and rat serum albumin (RSA) were determined by the biuret assay and radioimmunoassay, respectively. Glomerular size selectivity was assessed by examining the fractional clearance (FC) of neutral [3H]Ficoll using a short-term, steady-state approach. Cellular processing was examined by size exclusion chromatography analysis of urinary [14C]albumin and [14C]transferrin structural integrity. RESULTS: Total protein excretion (n = 4-15) increased 7-fold in BSA-treated rats (58 +/- 26 mg/24 h at baseline vs. 417 +/- 259 mg/24 h at day 3), while intact RSA excretion (n = 4-6) increased 200-fold (0.27 +/- 0.15 mg/24 h vs. 53 +/- 26 mg/24 h) despite a significant decrease in its plasma concentration. Total protein and albumin excretion returned to basal levels by day 7 (82 +/- 16 mg/24 h and 0.93 +/- 0.42 mg/24 h, respectively). FC of [3H]Ficoll of equivalent size to albumin did not change in BSA-treated rats as compared to vehicle controls. Tubular lysosomal processing of proteins was altered at peak proteinuria, as determined by changes in the structural integrity of urinary [14C]albumin, as well as [14C]transferrin, but returned to normal by day 7. CONCLUSIONS: This study demonstrates the lack of proportionality between changes in plasma concentration and urinary excretion of albumin, as well as the lack of change in the glomerular size selectivity to albumin. The profound but reversible changes in the amount and integrity of excreted protein suggest that cell-mediated processes are saturated by albumin concentrations, leading to the development of proteinuria in this model. PMID- 15280075 TI - An adenovirus-mediated gene-transfer model of angiogenesis in rat mesentery. AB - OBJECTIVE: To develop an adenovirus-mediated angiogenesis model, dependent on VEGF, in a system amenable to functional characterization. METHODS: Adenovirus (AdV) expressing VEGF (Ad-VEGF) or GFP (Ad-EGFP) (1-3.3 x 10(8) TCID50/mL), and Monastral blue were injected into the fat pad on either side of a mesenteric connective tissue panel of halothane-anesthetized male Wistar rats after intravital microscopic imaging. The intestine was replaced in the animal, the laparotomy was sutured, and the animal was allowed to recover. Six days later, the same connective tissue panel was identified from the Monastral blue depot and the mesentery was imaged as before, and then excised, fixed, and stained for endothelial cells (GSL isolectin-1B4), proliferating cells, VEGF, VEGF-R2, and actin. The increase in fractional microvascular area (FVA) was measured, and proliferating cell density, sprout density, vessel branch point density, vessel density, and mean vessel length were determined. RESULTS: AdVEGF injection significantly increased the fractional vessel area (2.9 +/- 0.4-fold), proliferating endothelial cell density (1.7 +/- 0.1-fold), sprout density (3.1 +/ 0.3-fold), branch point density (1.9 +/- 0.1-fold), and the microvessel density (1.4 +/- 0.1-fold), and decreased the mean vessel length (0.69 +/- 0.04-fold). VEGF-R2 staining was evident near sprouting tips of endothelial cells, but not on the tip itself, and evidence for arteriogenesis was observed as well as clear evidence of angiogenesis. CONCLUSIONS: Ad-VEGF injection into the fat pad of the rat induced significant angiogenesis in the mesentery. This two-dimensional model of VEGF-induced angiogenesis is amenable to physiological, biochemical, and molecular assessment and may be a useful tool to help understand mechanisms of angiogenesis. PMID- 15280076 TI - Differential expression of E- and P-selectin in the microvasculature of sickle cell transgenic mice. AB - OBJECTIVE: There is a growing body of evidence that endothelial cells assume an inflammatory phenotype in sickle cell disease. The authors determined whether (1) the expression of E- and P-selectin differs between sickle cell transgenic (beta(S)) mice and their wild-type counterparts, and (2) blood platelets and/or neutrophils contribute to the altered selectin expression. METHODS: Expression of E- and P-selectin was measured in different regional vascular beds of wild-type and beta(S) mice (with or without thrombocytopenia or neutropenia) using the dual radiolabeled monoclonal antibody technique. RESULTS: Constitutive expression of P selectin was significantly increased in the heart, lungs, small bowel, large bowel, and penis of beta(S) versus WT mice. While thrombocytopenia reduced P selectin expression in the small bowel and penis of beta(S) mice, neutropenia was associated with a reduction in P-selectin expression only in the penis. E selectin expression was not significantly elevated in any vascular bed except the penis of beta(S) mice. CONCLUSIONS: Sickle cell disease promotes an increased P selectin expression in several vascular beds. An accumulation of platelets may explain the increased P-selectin expression observed in some vascular beds. PMID- 15280077 TI - Influence of ACTH-(1-24) and plasma hyperviscosity on free radical production and capillary perfusion after hemorrhagic shock. AB - OBJECTIVE: The authors investigated the effects of ACTH-(1-24) and a high viscosity solution in the restoration of microvascular function during resuscitation. They injected NG-monomethyl-L-arginine (L-NMMA) and superoxide dismutase (SOD) before ACTH-(1-24) in hamsters resuscitated with the hyperviscous solution to determine the role of ROS and NO in ACTH-(1-24) protective mechanism in the cheek pouch. Hemorrhagic shock (HS) was induced by withdrawing blood to reduce mean arterial pressure (MAP) to 30 mm Hg for 45 min. METHODS: Animals were injected with ACTH-(1-24) and resuscitated with dextran of low molecular weight (70 kDa) and a small amount (4%) of dextran of high molecular weight (500 kDa) plus ACTH-(1-24), or autologous (shed) blood withdrawn during HS. Microvascular effects were characterized by measuring blood flow, perfused capillary length (PCL), arteriolar diameter, and red blood cell (RBC) velocity. ROS were assayed at the beginning and after 45 min of HS and after 10 and 90 min of resuscitation. RESULTS: Resuscitation with either shed blood or dextrans 70/500 resulted in the restoration of MAP, whereas PCL, RBC velocity, and arterial diameter decreased significantly. ROS increased significantly after HS, 10 and 45 min of resuscitation. ACTH-(1-24) plus dextrans 70/500 increased MAP immediately; it increased vasodilation and PCL, and attenuated significantly ROS production and leukocyte adhesion during resuscitation. L-NMMA injected after 30 min of HS did not change the protection exerted by ACTH-(1-24) and dextrans 70/500, while SOD increased their protective effects. CONCLUSIONS: ACTH-(1-24) appears to enhance the protective effects on the endothelium exerted by increased plasma viscosity by significantly decreasing the oxidative stress and the leukocyte adhesion during resuscitation. PMID- 15280078 TI - Mouse cremaster venules are predisposed to light/dye-induced thrombosis independent of wall shear rate, CD18, ICAM-1, or P-selectin. AB - OBJECTIVE: Microvascular adhesion of platelets to endothelium occurs in response to various inflammatory stimuli, and in venules is often accompanied by adherent leukocytes. In a light/dye injury model, platelet adhesion and thrombi occur preferentially in venules, though the reasons for this predisposition are unknown. The authors sought to determine whether lower wall shear rates or leukocyte-endothelial interactions accounted for preferential platelet thrombi formation in venules relative to arterioles. METHODS: A light/dye injury model of microvascular thrombosis was used in the mouse cremaster microcirculation. RESULTS: In wild-type mice (n = 17), the time to form microvascular platelet aggregates was delayed in arterioles by 3.1-fold relative to venules (p <.0001). However, arterioles with spontaneously low wall shear rates, as well as arterioles manipulated to reduce wall shear rate to venous levels, still had delayed thrombosis as compared to venules. Similarly, in animals deficient in CD18, P-selectin, or ICAM-1, the time to form platelet thrombi in arterioles was >3.0-fold higher than in venules. CONCLUSIONS: Mouse cremaster venules are predisposed to light/dye-induced microvascular thrombosis. The data suggest that functional differences between arteriolar and venular endothelial cells (independent of wall shear rate and of CD18, P-selectin, and ICAM-1) account for the venular predisposition to thrombosis. PMID- 15280079 TI - Recovery of nitric oxide from acetylcholine-mediated vasodilatation in human skin in vivo. AB - OBJECTIVES: To investigate the relative contribution of nitric oxide (NO) to the vascular and neural mechanisms underlying the ACh-induced vasodilatation in human skin. METHODS: ACh was delivered to the skin of the forearm of 28 healthy volunteers using intradermal microdialysis. Subsequent changes in tissue levels of NO and histamine were measured in the dialysate outflow and the associated changes in skin blood flux followed with the use of scanning laser Doppler imaging. RESULTS: ACh caused a dose-dependent increase in skin blood flux measured directly above the probe, associated with a twofold increase in dialysate NO. L-NAME (5 mM) delivered simultaneously via the dialysis probe totally blocked the increase in dialysate NO but only partially attenuated (approximately 30%) the ACh-induced increase in blood flux. At concentrations > or =6.25 mM, ACh also induced a widespread flare response, up to 40 mm in width, accompanied by the sensation of itch. The flare was not blocked by L-NAME or the H1 receptor antagonist levocetirizine, but was reduced by C-fiber blockade. Dialysate histamine levels remained unchanged at all times. CONCLUSIONS: These experiments offer further insight into the use of dialysis as an experimental technique in the skin. They provide direct evidence that the skin microvascular response to ACh is only partially mediated by NO. Further they suggest that ACh at higher concentrations can induce an axon-reflex-mediated response that is independent of NO release at the site of dermal provocation or of local histamine release. PMID- 15280080 TI - Telomerase-immortalized lymphatic and blood vessel endothelial cells are functionally stable and retain their lineage specificity. AB - OBJECTIVE: Microvasculature plays an important role in a variety of physiological and pathological processes. The authors have previously shown that primary cultures of human microvascular endothelial cells consist of 2 distinct populations of blood vascular and lymphatic endothelial cells. These subtypes are ephemeral and lose purity through passaging. To generate reproducible in vitro and in vivo experiments stable blood and lymphatic endothelial cell lines are an essential prerequisite. METHODS: In this study they have used human telomerase gene-immortalized nontransformed human microvascular endothelial cell cloned pure cultures of blood and lymphatic endothelial cell subpopulations. Flow cytometry, immunofluorescence, Northern and Western blotting, microarray gene analysis, as well as basic functional assays were used to characterize these clones. RESULTS AND CONCLUSIONS: Immortalized blood and lymphatic subpopulations are stable and functionally specialized cell lineages that expressed pan-endothelial and cell type-specific markers. They are excellent candidates for long-term culture studies on microvascular-related diseases. PMID- 15280081 TI - Myocardial contrast echocardiography in assessing microcirculation in baboons with chagas disease. AB - OBJECTIVE: Microvascular abnormalities have been postulated in the pathogenesis of chagasic cardiomyopathy. The objective of this study was to evaluate the relationship between coronary microcirculation and systolic function impairment in baboons with Chagas disease using myocardial contrast echocardiography (MCE). METHODS: Seventeen seropositive (5 males, 12 females; mean age 20 years) and 13 age- and gender-matched seronegative baboons underwent MCE using intravenous octafluoropropane human albumin microspheres. Color-coding was used to enhance tissue contrast in assessing regional myocardium uniformity and texture. Dipyridamole (0.54 mg/kg) was given to a subset of 4 animals to challenge coronary flow reserve. Systolic indices included left ventricular fractional shortening, velocity of circumferential fiber shortening, and left and right ventricular ejection fractions. RESULTS: Four of the 17 (24%) seropositive primates had decreased fractional shortening (25 +/- 8% vs. 40 +/-5%, p <.005), velocity of circumferential fiber shortening (1.05 +/- 0.36 circ/s vs. 1.84 +/- 0.23 circ/s, p <.0001), and reduced right ventricular ejection fraction (44 +/- 9% vs. 54 +/- 4%, p <.05) compared to other seropositive animals. Seropositive and seronegative groups showed no significant differences on the coronary microcirculation pattern as evaluated by MCE, including the 4 baboons with systolic function impairment. Moreover, coronary flow vasoreactivity resulted in a significant increase in myocardial flow as detected by color-coding masking. CONCLUSIONS: Chagasic heart disease is present in 24% of seropositive baboons spontaneously infected with Trypanosoma cruzi. MCE reveals a discrepancy between coronary microcirculation at rest and alterations in myocardial contractility, suggesting preservation of the microvascular integrity in this unique animal model. PMID- 15280082 TI - K+-induced dilation of hamster cremasteric arterioles involves both the Na+/K+ ATPase and inward-rectifier K+ channels. AB - OBJECTIVE: The mechanism by which elevated extracellular potassium ion concentration ([K+]o) causes dilation of skeletal muscle arterioles was evaluated. METHODS: Arterioles (n = 111) were hand-dissected from hamster cremaster muscles, cannulated with glass micropipettes and pressurized to 80 cm H2O for in vitro study. The vessels were superfused with physiological salt solution containing 5 mM KCl, which could be rapidly switched to test solutions containing elevated [K+]o and/or inhibitors. The authors measured arteriolar diameter with a computer-based diameter tracking system, vascular smooth muscle cell membrane potential with sharp micropipettes filled with 200 mM KCl, and changes in intracellular Ca2+ concentration ([Ca2+]i) with Fura 2. Membrane currents and potentials also were measured in enzymatically isolated arteriolar muscle cells using patch clamp techniques. The role played by inward rectifier K+ (KIR) channels was tested using Ba2+ as an inhibitor. Ouabain and substitution of extracellular Na+ with Li+ were used to examine the function of the Na+/K+ ATPase. RESULTS: Elevation of [K+]o from 5 mM up to 20 mM caused transient dilation of isolated arterioles (27 +/- 1 microm peak dilation when [K+]o was elevated from 5 to 20 mM, n = 105, p <.05). This dilation was preceded by transient membrane hyperpolarization (10 +/-1 mV, n = 23, p <.05) and by a fall in [Ca2+]i as indexed by a decrease in the Fura 2 fluorescence ratio of 22 +/- 5% (n = 4, p <.05). Ba(2+) (50 or 100 microM) attenuated the peak dilation (40 +/- 8% inhibition, n = 22) and hyperpolarization (31 +/- 12% inhibition, n = 7, p <.05) and decreased the duration of responses by 37 +/-11% (n = 20, p < 0.05). Both ouabain (1 mM or 100 microM) and replacement of Na+ with Li+ essentially abolished both the hyperpolarization and vasodilation. CONCLUSIONS: Elevated [K+]o causes transient vasodilation of skeletal muscle arterioles that appears to be an intrinsic property of the arterioles. The results suggest that K+-induced dilation involves activation of both the Na+/K+ ATPase and KIR channels, leading to membrane hyperpolarization, a fall in [Ca2+]i, and culminating in vasodilation. The Na+/K+ ATPase appears to play the major role and is largely responsible for the transient nature of the response to elevated [K+]o, whereas KIR channels primarily affect the duration and kinetics of the response. PMID- 15280083 TI - Effect of fibrinogen on leukocyte margination and adhesion in postcapillary venules. AB - OBJECTIVE: To quantitatively evaluate the role of fibrinogen (Fb) as a determinant of leukocyte (WBC) margination in postcapillary venules in light of its ability to induce red blood cell (RBC) aggregation with reductions in shear rate (gamma) and increase adhesiveness of WBCs to endothelium (EC). METHODS: Red cell aggregation (RCA), WBC margination (flux at the EC), rolling velocity, and adhesion to the EC were measured in rat mesenteric postcapillary venules upon reducing gamma, prior to and following systemic infusion of Fb. Proximal occlusion of feeding microvessels with a blunted probe facilitated reductions in gamma from 600 to 50 s(-1). An index of aggregation (G) was derived from light scattering properties of RBCs, where G was proportional to the number of RBCs per aggregate. WBC margination was measured as the percentage of total luminal WBC flux that rolled on the EC, F*(WBC). RESULTS: For normal levels of Fb (0.07 g%), reductions in gamma resulted in a 4-fold rise in F*(WBC) and no change in G as gamma was reduced to 50 s(-1). Infusion of Fb to achieve a plasma concentration to 0.7 g% caused a modest 20% increase in G and a 2.5-fold increase in F*(WBC) at gamma = 50 s(-1). WBC-EC adhesion appeared to increase significantly, but much less than with infusion of high molecular weight dextran (Dx). With Dx, G increased 3-fold, with reductions in gamma, but F*(WBC) increased only half the amount incurred with Fb at low shear. The greater margination in the presence of Fb results from RBC rouleaux that promote radial migration of WBCs. In contrast, clumps of RBCs resulting from high molecular weight Dx entrain WBCs within plasma gaps along the vessel centerline. CONCLUSIONS: In the presence of Fb, margination of WBCs increases dramatically at low shear due to rouleaux formation, which enhances radial migration of WBCs. This effect is much greater than with Dx because disruption of the much weaker Fb induced rouleaux precludes reductions in H(MICRO), whereas clumping aggregates induced by Dx form plasma gaps. Thus, modest levels of RCA caused by increased Fb may greatly enhance margination and with an enhancement of adhesiveness synergistically promote firm WBC-EC adhesion in the low flow state. PMID- 15280085 TI - Special issue of Microcirculation: examination of the vascular pathobiology of sickle cell anemia. Foreword. PMID- 15280086 TI - The not-so-simple process of sickle cell vasoocclusion. AB - Traditional concepts of sickle cell disease as a monogenically inherited disorder that is understood completely on the basis of polymerization based pathophysiology are more simple that what clinical observations allow. Detailed explications of the determinants of polymerization can be counted, but these do not account for all aspects of sickle cell disease. Neither can all perturbations that count in the course of sickle cell disease be counted as determinants of polymerization. The polymerization based theory that has been extrapolated to describe clinical disease often is not identical to clinical reality. Although contemporary understandings of sickle cell pathophysiology have been described as crazy by those bound to traditional polymerization based understandings, increasingly iconoclastic, seemingly crazy notions are regularly providing important new understandings of sickle cell disease. One of the major challenges to contemporary investigators is to describe new scientific insights in a way that can be understood by others, particularly those reluctant to afford polymerization independent discoveries validity among the interdependent processes that account for sickle cell disease. PMID- 15280087 TI - Polymerization and sickle cell disease: a molecular view. AB - The present molecular-level understanding of polymerization and sickling is reviewed for 2 central questions in sickle hemoglobin pathophysiology, viz., what determines when cells sickle, and what determines when cells get stuck. The description of sickling includes the central aspects of the double nucleation mechanism, as well as recent results on the effects of crowding, with an emphasis on the physiological applicability of this fundamental knowledge. In considering when cells get stuck, new measurements of individual fiber stiffness and the processes of depolymerization are also considered. Finally, a fundamental connection is shown between thermodynamics and rheology. PMID- 15280088 TI - The endothelial biology of sickle cell disease: inflammation and a chronic vasculopathy. AB - A single amino acid substitution in hemoglobin comprises the molecular basis for sickle cell anemia, but evolution of the corresponding clinical disease is extraordinarily complicated and likely involves multiple pathogenic factors. Sickle disease is fundamentally an inflammatory state, with activation of the endothelium, probably through proximate effects of reperfusion injury physiology and chronic molestation by adherent red cells and white cells. The disease also involves enhanced angiogenic propensity, activation of coagulation, disordered vasoregulation, and a component of chronic vasculopathy. Sickle cell anemia is truly an endothelial disease, and it is likely that genetic differences in endothelial function help govern its astonishing phenotypic diversity. PMID- 15280090 TI - Sickle cell vasoocclusion: heterotypic, multicellular aggregations driven by leukocyte adhesion. AB - Homozygous expression of sickle beta-globin alters the function of blood cells and the endothelium, producing a wide spectrum of clinical manifestations. Intravital microscopy studies in sickle cell mice suggest that vasoocclusion is a complex, sequential, multistep phenomenon involving (1) endothelial activation by sickle erythrocyte (SSRBC), (2) leukocyte (WBC) adhesion to the endothelium, and (3) the direct interaction between SSRBCs and adherent WBCs, which leads to reduced blood flow and tissue ischemia. Each of these steps represents a potentially useful therapeutic target. The identification of molecular determinants mediating vasoocclusion will provide new strategies for the prevention and treatment of this debilitating illness. PMID- 15280089 TI - In vivo studies of sickle red blood cells. AB - The defining clinical feature of sickle cell anemia is periodic occurrence of painful vasoocclusive crisis. Factors that promote trapping and sickling of red cells in the microcirculation are likely to trigger vasoocclusion. The marked red cell heterogeneity in sickle blood and abnormal adhesion of sickle red cells to vascular endothelium would be major disruptive influences. Using ex vivo and in vivo models, the authors show how to dissect the relative contribution of heterogeneous sickle red cell classes to adhesive and obstructive events. These studies revealed that (1) both rheological abnormalities and adhesion of sickle red cells contribute to their abnormal hemodynamic behavior, (2) venules are the sites of sickle cell adhesion, and (3) sickle red cell deformability plays an important role in adhesive and obstructive events. Preferential adhesion of deformable sickle red cells in postcapillary venules followed by selective trapping of dense sickle red cells could result in vasoocclusion. An updated version of this 2-step model is presented. The multifactorial nature of sickle red cell adhesion needs to be considered in designing antiadhesive therapy in vivo. PMID- 15280091 TI - The perfusion paradox and vascular instability in sickle cell disease. AB - Sickle cell disease (SCD) exhibits a curious coexistence of contrasting perfusion profiles in the circulatory system: hypoperfusion is endemic in microcirculatory beds occluded by hemoglobin S-containing erythrocytes while hyperperfusion characterizes the systemic (macro)circulation and a number of regional vascular circuits. This review highlights this perfusion paradox of SCD, focusing on forearm blood flow and the renal circulation, and exploring the extent to which alterations in vasoactive systems (such as nitric oxide and prostanoids) are involved. Also reviewed are the mechanisms and pathways that contribute to altered vascular reactivity and vascular instability observed in SCD. Finally, the possibility that the induction of heme oxygenase-1, recently described in SCD, may confer a protective response in the vasculature and other tissues is discussed. PMID- 15280092 TI - Pathophysiology of stroke in sickle cell disease. AB - Stroke affects both motor and cognitive function in patients with sickle cell disease (SCD). Symptomatic stroke is associated with intimal disease of the large cerebral arteries. Silent stroke, defined as cerebral infarction in the absence of overt clinical neurologic symptoms, is often due to microinfarcts suggestive of microvascular disease. While the natural history of stroke in SCD is well described, the pathophysiology remains poorly understood and probably varies with the site of vascular injury. Increased red cell adhesion, oxidative injury of the vessel wall, inflammation, abnormal vasomotor tone regulation, and increased activity of the coagulation system all may contribute to cerebral vasopathology in SCD. PMID- 15280093 TI - Sickle red cell microrheology and sickle blood rheology. AB - The hallmark of the phenotypic expression of sickle cell disease is the remarkable degree of heterogeneity in the clinical manifestations. They vary latitudinally among patients and longitudinally in the same patient. The pathogenesis of sickle cell anemia centers on the sequence of events that occur between polymerization of deoxy hemoglobin S and increased red cell destruction, vasoocclusion, and end organ damage. Cellular dehydration, changes in sickle red blood cell rheology, adhesion of sickle red cells to vascular endothelium, inflammatory response, and tissue injury are some of the factors that contribute to hemolytic anemia, vasoocclusion, and eventual multiorgan damage. The focus of this review is on the rheology of sickle blood and microrheology of sickle RBC. Determinants of sickle RBC rheology and the factors that modulate its severity are discussed. PMID- 15280094 TI - Lymphatic diversion prevents myocardial edema following mesenteric ischemia/reperfusion. AB - OBJECTIVE: Mesenteric ischemia/reperfusion (I/R) is associated with cardiac dysfunction. Mesenteric lymph primes polymorphonuclear leukocytes (PMNs) for increased superoxide release following I/R. We hypothesized that mesenteric I/R causes myocardial edema resulting in myocardial dysfunction, and that diverting mesenteric lymph would preserve myocardial function. METHODS: Two canine groups were studied: lymphatic diversion (LD) and no lymphatic diversion (No LD). Preload recruitable stroke work, +/-dp/dt(max), isovolumic relaxation (tau), cardiac output, and myocardial water content (MWC) were determined. I/R consisted of 60 min of ischemia followed by 180 min of reperfusion. Myocardial myeloperoxidase (MPO) was measured as an index of PMN leukosequestration. In addition, mesenteric lymph harvested after I/R was infused into normal canines and all variables measured. RESULTS: MWC increased from baseline in No LD. Tau and -dp/dt(max) were significantly affected in No LD, but not in LD. After mesenteric I/R, mesenteric lymph primed PMNs for increased superoxide production. Lymph diversion resulted in significantly lower myocardial MPO. With reinfusion of I/R lymph, MWC and tau increased. MPO was also increased post I/R mesenteric lymph reinfusion. CONCLUSIONS: Our data indicate that myocardial dysfunction after mesenteric I/R is due to lymph-induced, PMN-mediated microvascular alterations and myocardial edema. PMID- 15280095 TI - Gender-specific regulation of cardiovascular function: estrogen as key player. AB - This review provides an overview of gender-specific differences in the incidence and development of cardiovascular diseases, including hypertension, atherosclerosis, heart failure and the corresponding myocardial remodeling. The review discusses the possible mechanisms by which estrogen affords a beneficial effect on cardiovascular function via genomic vs non genomic regulation; estrogen receptor-dependent vs estrogen receptor-independent pathways, specific signal transduction cascades, especially those involving protein kinase B (Akt) and mitogen activated protein kinase (MAPK), as well as their downstream targets, such as nitric oxide synthase, cyclooxygenase, cytochrome P450 (CYP), NADPH oxidase and superoxide dismutase. Having considered the essential role of the microcirculation in the control of vascular resistance in vivo, estrogen-related regulation of microvascular function and blood pressure is highlighted. Attention is focused on the effects of estrogen on pressure (myogenic)-dependent and flow/shear stress-dependent mechanisms of arterioles, which contribute significantly to the control of local blood flow and peripheral resistance via alterations in the release of endothelial mediators, such as nitric oxide, prostaglandins and endothelium-derived hyperpolarizing factor. PMID- 15280096 TI - Flow visualization tools for image analysis of capillary networks. AB - OBJECTIVE: Video recordings of red blood cell (RBC) flow through capillary networks contain a considerable amount of information pertaining to oxygen transport through the microcirculation. Image analysis of these video recordings has been widely used to determine RBC dynamics (velocity, lineal density and supply rate) and oxygenation (Brunner et al., 2000; Ellis et al., 1990, 1992; Ellsworth et al., 1987; Klyscz et al., 1997; Pries 1988). However, not all capillaries in a given field of view are suitable for image analysis. Typically, capillary segments that are relatively straight and in sharp focus, and exhibit flow of individual RBCs that are well separated by plasma gaps, are good candidates for analysis. We have developed several image processing tools to aid in the selection of such capillaries for analysis and to obtain quick overviews of RBC flow through the microcirculation. METHODS: Burgess et al. (Microcirc. 2:75, 1995) and Burkell et al. (Annals Biomed. Eng. 24:1, 1996; J. Vasc. Res. 35:2, 1998) have previously introduced mean and variance images to aid in the selection of capillaries for analysis. We have extended their concept and developed similar two dimensional visualization techniques for studies of RBC flow through capillary networks. RESULTS: Five new methods of processing video data were developed. The minimum image highlights all capillaries containing RBCs in a given field of view. The maximum image identifies capillaries that exhibit high lineal density or stopped flow. The range image represents the difference between the maximum and minimum light intensity values that occur at a given pixel over a given time period, and helps to identify capillary segments that are in good focus and are perfused by RBCs and plasma. The difference image represents the cumulative sum of the square of differences in intensity values between consecutive frames and gives an indication of the frequency of passage of RBCs separated by plasma gaps. The transition image represents the number of times the intensity at a given pixel crosses a predefined threshold and indicates the number of RBCs (or trains of RBCs) that passes a given location during the observation period. CONCLUSIONS: The above flow visualization techniques are valuable tools to aid in the study of image focus, network geometry, RBC flow paths and dynamics, that can then be used in identifying capillaries for subsequent (separate) detailed analysis to provide quantitative information about RBC flow. PMID- 15280097 TI - Regulation of leukocyte recruitment by local wall shear rate and leukocyte delivery. AB - OBJECTIVE: To determine the influence of wall shear rate and leukocyte delivery on leukocyte margination, rolling, and adhesion in post-capillary venules. METHODS: Leukocyte-endothelial cell interactions were characterized in cremaster muscle venules of anesthetized mice with video microscopy under control conditions and after 3 h exposure to TNF-alpha. Hemodynamic parameters were measured with fluorescent particle tracking using confocal microscopy. RESULTS: Leukocyte recruitment to the vessel wall and leukocyte rolling increased as a function of wall shear rate (P < 0.05) over the range observed (0-200 s(-1)) in intact post capillary venules. Leukocyte delivery affected recruitment and rolling flux only in activated vessels (P < 0.05). In addition, leukocyte firm adhesion was independent of both wall shear rate and leukocyte delivery, but showed an overall increase in TNF-alpha stimulated tissues (0.9 +/- 0.2 vs. 2.7 +/- 0.6 cells/50 microm). CONCLUSIONS: Leukocytes are delivered to venules in excess of the capacity of the local endothelium to support interactions. Elevated shear forces increase leukocyte recruitment to the vessel wall, which correlates to elevated rolling flux. In contrast, leukocyte firm adhesion is primarily affected by the activation state of the tissue and not by hemodynamic factors. Overall, the capacity of endothelial cells to support leukocyte interactions primarily regulates leukocyte recruitment and is not limited by leukocyte supply. PMID- 15280098 TI - Enhancement of glucocorticoid and mineralocorticoid receptor density in the microcirculation of the spontaneously hypertensive rat. AB - OBJECTIVE: Elevated blood pressure and abnormal physiological parameters in the microcirculation of the spontaneously hypertensive rat (SHR) can be normalized by adrenalectomy. Thus glucocorticoids and mineralocorticoids may have major control over blood pressure status and organ injury mechanisms in SHRs. As background, this study was designed to examine the distribution of the glucocorticoid and mineralocorticoid receptors in a microvascular network. METHODS: Mature SHR and their normotensive controls, the Wistar-Kyoto (WKY) rat, were studied. An immunohistochemical method was developed that provides a comprehensive display of the receptors in all segments of the mesentery microcirculation and the surrounding tissue parenchyma. RESULTS: All cells in the mesentery exhibit immunolabeling of the glucocorticoid receptor with predominant expression in the nuclei of parenchymal and endothelial cells. The mineralocorticoid receptor is expressed also in most cells of the microcirculation and adjacent parenchymal tissue. Both receptors exhibit the highest levels of immunolabel in the wall of the arterioles and venules, with lower levels in capillaries. Compared with WKY rats, the SHRs exhibit significantly enhanced density of glucocorticoid and mineralocorticoid receptors in endothelial cells of arterioles and venules as well as in parenchymal cells. CONCLUSIONS: These results suggest that the enhanced sensitivity of the SHR to glucocorticoids and aldosterone may be in part associated with enhanced glucocorticoid and mineralocorticoid receptor densities in the microcirculation. PMID- 15280099 TI - Glycoprotein IIB/IIIA-inhibition and microcirculatory alterations during experimental endotoxemia--an intravital microscopic study in the rat. AB - OBJECTIVE: Endothelial damage during early endotoxemia has been shown to be leukocyte-independent. Platelet-activating factor (PAF) and serotonin-receptor antagonism are known to reduce leukocyte-independent macromolecular leakage significantly, thereby focusing the field of interest to the platelets. We hypothesized that inhibition of the GP IIb/IIIa receptor, using the unspecific GP IIb/IIIa inhibitor abciximab, would reduce leukocyte-independent endothelial damage during early endotoxemia, and furthermore, that inhibition of the GP IIb/IIIa receptor with abciximab might improve microcirculatory disturbances, seen in a leukocyte-dependent animal model during endotoxemia. METHODS: In male Wistar rats, venular wall shear rate, macromolecular efflux, and leukocyte endothelial interaction were determined in mesenteric postcapillary venules using intravital microscopy at baseline, 60, and 120 min after the start of the experiment. The experiment was divided into two parts. In the first part, we investigated the effects of the GP IIb/IIIa inhibitor abciximab on leukocyte independent endothelial permeability during endotoxemia. In the second part of the experiment, we focused on the effects of the GP IIb/IIIa inhibitor abciximab on microcirculatory disturbances during endotoxemic states, without a modification of the leukocyte-endothelial interaction, putting the main emphasis again on endothelial permeability. RESULTS: GP IIb/IIIa inhibition with abciximab resulted in a significant reduction of macromolecular efflux during leukocyte independent endotoxemia. Both pretreatment with abciximab and post-treatment with abciximab reduced macromolecular leakage during leukocyte-dependent endotoxemia to values comparable to control values, and prevented an increase in leukocyte adherence, that has been reduced to values comparable to control values, too. CONCLUSION: GP IIb/IIIa inhibition, using abciximab, protects against endothelial dysfunction and an increase in leukocyte adherence to the vascular wall during experimental endotoxemia. The protective properties of abciximab on microcirculation seemed to be leukocyte-independent. PMID- 15280100 TI - Expression of cytochrome P450-4A isoforms in the rat cremaster muscle microcirculation. AB - OBJECTIVE: This study sought to identify any specific cytochrome P450 (CYP450) 4A enzyme isoforms expressed in arterioles and/or the surrounding parenchymal tissue of the rat cremaster muscle. METHODS: RT-PCR was used to detect the presence of specific CYP450-4A isoforms in isolated muscle fibers and arterioles from the cremaster muscle of Sprague-Dawley rats; CYP450-4A protein expression was determined by Western blotting. RESULTS: CYP450-4A3 mRNA was expressed in isolated muscle fibers and in cremasteric arterioles, while CYP450-4A8 mRNA was expressed only in cremasteric arterioles. CYP450-4A1 and CYP450-4A2 mRNA were not expressed in arterioles and skeletal muscle cells, although all four isoforms were strongly expressed in the liver. CYP450-4A protein was detected in both the isolated muscle fibers and in the isolated arterioles. CONCLUSIONS: The present study identifies the specific pattern of cytochrome P450-4A isoform expression in arterioles and parenchymal cells of the skeletal muscle microcirculation, and supports the hypothesis that the cytochrome P-450 enzymes may play a role in the regulation of microvascular function in the skeletal muscle microcirculation. PMID- 15280101 TI - In memoriam. PMID- 15280102 TI - Repeat courses of antenatal corticosteroids: what does the evidence show? PMID- 15280103 TI - Chronic prenatal exposure to phenobarbital and long-term behavior effects on mice offspring. AB - OBJECTIVE: To assess the impact of chronic prenatal exposure to phenobarbital on long-term behavior in mice offspring. METHODS: Twenty-eight C3H/He mice were randomized to receive diet chow containing either a daily therapeutic dose of phenobarbital (2.5 mg in 10-g chow) or a placebo for 1 week before mating and throughout gestation. Offspring from eight litters of each treatment group were evaluated using motor function, arousal/motivation, anxiety-provoking and cognition tasks. RESULTS: No significant differences between groups were found in duration of gestation, litter size and birth weights. Fewer counts in a locomotor chamber were observed in phenobarbital-exposed offspring (524 +/- 31 vs. 688 +/- 54 for 60 min, p < 0.02; 4174 +/- 229 vs. 5230 +/- 406 for 22 h, p < 0.05). Initial reactions to a startle were more apparent among phenobarbital-exposed offspring (p < 0.03). Impaired co-ordination of hindlimbs was observed in the phenobarbital-exposed offspring during the wire maneuver (p < 0.001). Fewer entries into the mirrored chamber were observed after phenobarbital exposure (2.1 vs. 4.5; p < 0.05). Exposure to phenobarbital was not found to affect responses to learning and memory tasks (homing, tube runway, water runway, Morris maze). CONCLUSION: Although cognition was unaffected by prenatal exposure to phenobarbital, subtle effects on locomotor activity, hindlimb co-ordination and responses to anxiety-provoking conditions require human correlation. PMID- 15280104 TI - Plasma amino acid concentrations throughout normal pregnancy and early stages of intrauterine growth restricted pregnancy. AB - OBJECTIVES: Assessment of maternal plasma amino acids during normal gestation and in early stages of intrauterine growth restriction (IUGR). STUDY DESIGN: Plasma amino acid concentrations were measured in: (1) non-pregnant women (n=7); (2) normal pregnant women in the first (n=13), second (n=17) and third (n=12) trimester; and (3) pregnant women in the first trimester with later development of IUGR (n=8). Amino acid levels were quantified by electrochemical detection in a reversed-phase high-performance liquid chromatography (HPLC) system. RESULTS: The levels of most essential and non-essential amino acids changed markedly in the first trimester during normal pregnancy and thereafter remained almost constant. In the first trimester of IUGR, a number of both essential and non essential amino acids were significantly different from those observed in normal pregnancies, with values more similar to those observed in non-pregnant women. CONCLUSIONS: Levels of most maternal amino acids decrease and some increase during early gestation reflecting a metabolic adaptation that occurs in normal pregnancies. Pregnancies that later develop IUGR show a lack of these adaptations for a significant number of both essential and non-essential amino acids, suggesting a lack of adaptation. PMID- 15280105 TI - Ultrasound evaluation of intrauterine growth restriction therapy by a nitric oxide donor (L-arginine). AB - OBJECTIVE: There are numerous methods available of treating intrauterine growth restriction but their results are still not satisfactory. Currently, we are conducting a research project whose main aim is based on the use of the nitric oxide (NO) donor L-arginine in growth restriction therapy. The main aim of this study was the ultrasound evaluation of the efficacy of this therapy based on biometric measurements (the estimated fetal weight) compared with the estimated weight of newborn children. STUDY DESIGN: The investigated group comprised two randomly chosen groups of pregnant women with ultrasound-diagnosed intrauterine growth restriction (biometry < 10th centile for gestation age): 78 patients were treated by L-arginine 3 g daily orally for 20 days; and 30 patients, not treated, acted as the control group. RESULTS: The ultrasound estimation of fetal weight at the start and at the end of the treatment showed a mean increase of 642 g (SE 90 g) using the Shepard method, and 648 g (SE 94 g) using the Hadlock method, respectively. By comparison, within the control group a mean value increase of 395 g (SE 77 g) was found, using the Shepard method, and 404 g (SE 82 g) using the Hadlock method, respectively. There was a significant statistical difference when comparing the estimated fetal weight increase in both methods: p=0.008 for the Shepard calculation and p=0.012 for the Hadlock calculation. The weight of the newborn infants was also evaluated: in the treated group the mean value was 2823 g (SE 85 g) and in the untreated group the mean value was 2495 g (SE 147 g). There was a significant (p=0.027) difference, showing a positive effect of the treatment on the weight of newborns. In the treated group the percentage of growth-retarded newborns was 29% while in the untreated group it was 73%. A significant difference has been found (p < 0.01) between both of the groups of newborns. CONCLUSIONS: The ultrasound evaluation of the estimated fetal weight and the birth weight of the newborns showed an improvement: there was an acceleration of fetal development in the L-arginine-treated group of pregnant women as compared with the untreated group. The ultrasound evaluation of the estimated fetal weight is a good diagnostic tool, properly monitoring the efficacy of the L-arginine treatment of the growth-retarded fetuses. PMID- 15280106 TI - Gender differences in amniotic fluid cytokine levels. AB - OBJECTIVE: Placental trophoblast invasion and amniotic fluid cytokine receptor levels have been reported to vary with fetal gender. We investigated whether fetal gender affects amniotic fluid levels of the inflammatory cytokines interleukin (IL)-6 and IL-10 and the pro-angiogenesis cytokine angiogenin. METHODS: Specimens from singleton gestations undergoing mid-trimester amniocentesis for genetic indications were used. Inclusion criteria were (1) outcome information available, (2) no structural or chromosomal anomaly and (3) no conditions associated with preterm delivery. Amniotic fluid IL-6, IL-10 and angiogenin levels were measured by immunoassay. Statistical analysis included the Mann-Whitney U test and Fisher's exact test with p < 0.05 indicating significance. RESULTS: A total of 74 samples were analyzed. Angiogenin levels were significantly lower in amniotic fluid samples from pregnancies with a male than with a female fetus (median (range): 22.2 (5.9-66.4) vs. 32.0 (11.4-159.2) ng/ml, p=0.007), in contrast to no differences in amniotic fluid IL-6 and IL-10 levels (p=0.4 and p=0.1, respectively). In pregnancies with male fetuses delivering preterm (< 37 weeks), angiogenin was also detected at lower levels (p=0.02). There were no gender differences with respect to race, nulliparity or maternal age. CONCLUSION: Angiogenin levels, but not IL-6 or IL-10 levels, are significantly lower in second-trimester amniotic fluid of women with male compared with female fetuses, including those women delivering preterm. PMID- 15280107 TI - An abnormal screening ultrasound: concordance with a tertiary obstetrical ultrasound unit. AB - OBJECTIVE: To determine concordance of ultrasound diagnosis in referrals to a tertiary obstetrical ultrasound unit (TOU) for suspected abnormalities. STUDY DESIGN: Consecutive referrals for "abnormal outside ultrasound" during a 6-month period were compared with the TOU ultrasound diagnosis. Concordance of diagnosis was compared on the basis of organ system involved and referral for single or multiple suspected abnormalities. Chi(2) analysis was used; p < 0.05 was considered significant. RESULTS: Of 104 consecutive referrals reviewed, 42 (40.4%) had no abnormality documented at the TOU. Of the 62 abnormal ultrasound scans at the TOU, 78.3% were concordant. Concordance based on organ system involvement was central nervous system, 30.3%; cardiothoracic, 66.7%; gastrointestinal, 63.6%; genitourinary, 50%. Referrals for a single suspected anomaly were statistically no more likely to have a normal TOU ultrasound scan (40.4%) than those referred for multiple suspected anomalies (36.4%, p > 0.05). CONCLUSIONS: Most referrals to a tertiary center for "abnormal outside ultrasound" will be diagnosed with an abnormality. PMID- 15280108 TI - The predictive value of the 1-h 50-g glucose screen for diagnosing gestational diabetes mellitus in a high-risk population. AB - OBJECTIVE: To determine a value, for a gestational diabetes mellitus (GDM) screening test, above which the glucose tolerance test is obviated. METHODS: A database search of patients delivered at the Medical College of Virginia Hospital (MCV) between April 1991 and April 2002 was undertaken. Subjects were screened using standard methodology: blood glucose level 1 h after a 50-g oral glucose load (1OGT). Subjects with values meeting/exceeding 140 mg/dl underwent 3-h 100-g oral glucose tolerance tests (3OGTT). GDM was diagnosed using criteria of the National Diabetes Data Group (NDDG), with Carpenter-Coustan (CC) criteria for comparison. Receiver-operator characteristic (ROC) curves were generated; areas under the curve (AUC) were calculated. RESULTS: 1OGT results were available for 16898 subjects; 2770 (16.4%) had values meeting/exceeding 140 mg/dl. The NDDG and CC criteria were applied to 1972 subjects with both 1OGT and 3OGTT results available: 419 (21%) and 614 (31%) subjects had GDM, respectively. Positive predictive values for results > or =180 mg/dl and values at 20 mg/dl increments up to 260 mg/dl were: 36, 47, 55, 57 and 63% (NDDG) and 45, 54, 62, 61 and 66% (CC). AUC for NDDG=0.68; AUC for CC=0.64. CONCLUSIONS: GDM cannot be diagnosed with the 1OGT; predictive values are low. A cut-off of 200 mg/dl predicts only 47 54% of GDM cases correctly, and may lead to over-diagnosis. It is inappropriate for GDM to be diagnosed based on the 1OGT. PMID- 15280109 TI - The influence of obstetric intervention on trends in twin stillbirths: United States, 1989-99. AB - OBJECTIVE: Although twin stillbirth rates have declined substantially over the past two decades, the contribution of changes in obstetric interventions to reducing twin stillbirths has not been quantified. METHODS: We carried out a retrospective cohort study of twin live births and stillbirths in the United States between 1989 and 1999 (n=1,102,212). Changes in the rate of stillbirth (> or =22 weeks) before and after adjustment for changes in labor induction, Cesarean delivery and sociodemographic factors were estimated through ecological logistic regression analysis. This analysis was based on aggregating data by each state within the United States. RESULTS: Between 1989 and 1999, rates of labor induction and Cesarean delivery among twin live births increased by 138% (from 5.8% to 13.8%) and 15% (from 48.3% to 55.6%), respectively. These changes were accompanied by a 43% decline in the stillbirth rate between 1989 and 1999 (from 24.4 to 13.9 per 1000 fetuses at risk). After excluding births weighing < 500 g, rates of labor induction among twins at 22-27 weeks', 28-33 weeks' and > or =34 weeks' gestation increased by 95%, 131% and 127%, respectively, between 1989 and 1999. Cesarean delivery rates also increased by 55%, 29% and 2% in these same gestational age categories. The 48% (relative risk (RR) 0.52, 95% confidence interval (CI) 0.49-0.55) decline in stillbirth rate between 1989-91 and 1997-99 was reduced to a 25% (RR 0.75, 95% CI 0.72-0.79) decline after adjustment for changes in labor induction and Cesarean delivery. The decline in the rate of twin stillbirths was larger at later gestational ages (at > or =32 and > or =34 weeks) where the largest absolute increases in labor induction rates were observed. CONCLUSIONS: The use of Cesarean delivery and especially labor induction for twin pregnancies has increased substantially in the United States over the last decade and these changes have been associated with a large decline in the rate of stillbirth among twins. PMID- 15280110 TI - Vaginal birth after Cesarean delivery: predicting success, risks of failure. AB - OBJECTIVES: To identify predictors of successful trial of labor in women after one low transverse Cesarean delivery and no prior deliveries, and to assess perinatal morbidity associated with a failed vaginal birth after Cesarean delivery (VBAC). METHODS: Retrospective chart review of women with one low transverse Cesarean delivery in their first pregnancy who delivered their next pregnancy at our institution. Clinical characteristics and intrapartum data were reviewed to identify predictors of successful VBAC. Perinatal outcomes were reviewed to assess morbidity associated with VBAC attempt and failed VBAC. RESULTS: Of 768 women studied, 522 (68%) attempted VBAC and 344 (66%) of these were successful. Uterine rupture occurred in 0.8% of the VBAC group. On initial examination, women with cervical dilation >1 cm, effacement > 50% and station lower than -1 were more likely to deliver vaginally. Women with successful VBAC had more spontaneous labor (85.2 vs. 76.4%, p=0.02) and less oxytocin use (49.7 vs. 70.8%, p < 0.0001). There were no differences in outcomes between failed and successful VBAC, except more frequent 1-min Apgar scores < 5 (10.1 vs. 4.1%, p=0.01) and increased endometritis (9.6 vs. 2%, p=0.0002) with failed VBAC. Compared with elective repeat Cesarean delivery, VBAC attempt was associated with amnionitis (5.9 vs. 0%, p < 0.0001) and low 1- and 5-min Apgar scores (6.1 vs. 2.4%, p=0.03 and 2.3 vs. 0%, p=0.01, respectively), but not endometritis, admission to a neonatal intensive care unit (NICU), ventilation, intraventricular hemorrhage (IVH) or seizures. Failed VBAC had more amnionitis (7.3 vs. 0%, p < 0.0001), postpartum fever (11.2 vs. 2.4%, p=0.0003) and endometritis (9.6 vs. 2.0, p=0.0007) than elective repeat Cesarean delivery and was associated with low 1- and 5-min Apgar scores (10.1 vs 2.4%, p < 0.001 and 2.8 vs. 0%, p=0.01, respectively), but not NICU admission, ventilation, IVH or seizures. CONCLUSIONS: Favorable initial pelvic examination, spontaneous labor and a lack of oxytocin use are associated with successful VBAC in women with a single prior low transverse Cesarean delivery and no prior vaginal deliveries. While attempted VBAC and failed VBAC have more maternal infectious morbidity and lower Apgar scores, infant outcomes are similar to those of elective repeat Cesarean delivery. PMID- 15280112 TI - STAN S21 fetal heart monitor for fetal surveillance during labor: an observational study in 637 patients. AB - OBJECTIVE: To assess the value of the STAN fetal heart monitor for intrapartum fetal monitoring using cardiotocography (CTG) and fetal electrocardiography (ECG). DESIGN: Prospective observational study. MATERIAL AND METHODS: Between August 2000 and November 2002, 637 high-risk labors were monitored using a STAN S21 fetal heart monitor, providing CTG plus automatic ST analysis of the fetal ECG. Guidelines with recommendations about when to intervene were available. During the study period labor-ward personnel were systematically instructed about the (patho)-physiology of asphyxia and CTG and ST changes during labor. RESULTS: Four hundred and forty-nine recordings were available for analysis of outcome in relation to ST changes. In 61 cases, ST changes requiring intervention occurred > 10 min before birth. In 35 (57%) of these cases, umbilical artery blood pH at delivery was < 7.15. Eighteen (4.0%) neonates were born with metabolic acidosis (umbilical artery pH < 7.05 and extracellular base deficit > 12 mmol/l). Significant ST changes (18-31 min before birth) were present in all five cases with pH < 7.00 and in six of the 13 cases with pH of 7.00-7.04 (false-negative rate 1.6%). Neonatal follow-up showed no adverse outcome. One hundred and ninety two fetal blood samples (121 in the first stage and 71 in the second stage of labor) were taken from 142 women. Fetal scalp blood pH was < 7.15 in ten samples, 7.15-7.19 in 11 samples, 7.20-7.24 in 30 samples and > or =7.25 in 141 samples. ST changes occurred in eight (80%), six (55%), nine (30%) and 15 (11%) of these cases, respectively. In 188 (29.5%) women, outcome could not be analyzed in relation to ST changes because of inadequate recording (time between end of recording and delivery > 20 min or poor signal quality) or the absence of umbilical cord gases. In this group, four (2.1%) neonates with metabolic acidosis were born. In three of these cases the fetal ECG signal was of was poor quality and in one case the recording had ended 60 min before birth. CONCLUSION: ST changes were present in all five cases with severe metabolic acidosis (umbilical artery pH < 7.00). ST changes occurred in 46% of cases with mild metabolic acidosis. CTG plus ST analysis was more specific in detecting fetal acidemia than CTG alone. PMID- 15280111 TI - Ethical decision-making for extremely preterm deliveries: results of a qualitative survey among obstetricians and midwives. AB - OBJECTIVE: To explore the practices, attitudes and feelings of obstetricians and midwives in cases of extreme prematurity. METHODS: A qualitative study was conducted as part of a European Concerted Action (EUROBS) in three tertiary-care maternity units, located in three cities in the northern, southern and central areas of France. Semi-structured interviews lasted an average of 60 min and were tape-recorded. They were independently analyzed by two different researchers using a content analysis. All full-time obstetricians and half of the full-time midwives were eligible for the study. Overall, 17 obstetricians and 30 midwives participated. RESULTS: Both obstetricians and midwives considered that decision making in case of very preterm births raised ethical problems concerning the mother and the fetus. Despite some birth weight and gestational age criteria defined in advance, management around delivery appeared to be decided on a case by-case basis. At birth, the neonatologists made the decisions. They were perceived as being more inclined than the obstetric team to initiate intensive care. If the child was born alive, intensive care was started, in the knowledge that it could be withdrawn later, if appropriate. Parents were sometimes involved in decision-making during pregnancy, in particular when there was no emergency situation. Compared with obstetricians, midwives tended to have a less favorable perception of the neonatologists' practices, and to report less parental involvement in decision-making. CONCLUSIONS: Decisions about the obstetric management and resuscitation of extremely preterm infants are usually made on a case-by-case basis. Parents are sometimes involved in decision-making. Midwives express serious concerns about the current practices. PMID- 15280113 TI - Does maternal smoking influence leptin levels in term, appropriate-for gestational-age newborns? AB - OBJECTIVES: Leptin, a hormone produced in adipose tissue and the placenta, is correlated with neonatal growth. The aim of this study was to investigate the effect of maternal smoking during pregnancy on cord blood leptin concentrations in term, appropriate-for-gestational-age infants. METHODS: Two groups of term, appropriate-for-gestational-age newborns were selected: 19 infants of smoking mothers and 91 infants of non-smoking mothers. Neonatal anthropometric measurements were taken and leptin levels were measured by radioimmunoassay. RESULTS: Leptin concentrations were similar (p=0.915) between the groups. Leptin levels correlated only with ponderal index (p < 0.01) and gestational age of the newborns (p < 0.05). CONCLUSIONS: This study indicates that maternal smoking during pregnancy does not affect cord blood leptin levels in term, appropriate for-gestational-age infants. PMID- 15280114 TI - Birth weight for gestational age centiles for Italian neonates. AB - OBJECTIVE: To provide centiles for birth weight (BW) according to gestational age (GA) and sex for infants born in Italy. METHODS: We used records of the whole neonatal population of Tuscany, a region in Italy, from July 1991 to June 2002 as resulting from the database of the cystic fibrosis neonatal screening program (n=290129). We excluded as unlikely for GA those BW that were more than two interquartile ranges above the 75th centile or below the 25th centile for each GA and gender group. RESULTS: We present the 3rd, 10th, 25th, 50th, 75th, 90th and 97th centiles of BW for GA from the 24th to 43rd week of gestation for male and female Italian neonates, as both tables and smoothed curves. CONCLUSIONS: The large size of the examined population allows us to provide up-to-date, reliable BW for GA centiles for Italian newborns, especially for lower GAs. PMID- 15280115 TI - Pregnancy in a woman with unilateral lung agenesis. AB - Unilateral lung agenesis is a rare congenital condition of unknown etiology. A 33 year-old nullipara with right lung agenesis and scoliosis was admitted to the hospital at 30 weeks of gestation because of oligohydramnios. At 32 weeks she was treated for an upper respiratory tract infection with azithromycin. She went into premature labor at 34 weeks and was delivered by Cesarean for breech presentation. Both mother and infant did well. PMID- 15280116 TI - A checklist to identify the origin of cerebral palsy. PMID- 15280117 TI - Serum concentrations of lipids and apolipoproteins in normal and hyperemetic pregnancies. AB - OBJECTIVE: Hyperemesis gravidarum (HEG) is intractable nausea and vomiting. The purpose of this study was to test the hypothesis that women with HEG have lower cholesterol and triglyceride levels, to find any role in the etiology of reduced risk of spontaneous abortion in hyperemetic patients. STUDY DESIGN: The study group consisted of 39 women with normal ongoing pregnancy and 35 women with HEG. The concentrations of triglycerides, high-density lipoprotein (HDL) cholesterol, low-density lipoprotein (LDL) cholesterol, total cholesterol and apolipoprotein (apo)-A and -B were analyzed. The independent-samples t test, Mann-Whitney U test, chi2 test, Kruskal-Wallis variance analysis and Spearman's correlation were used to examine differences between groups. RESULTS: Serum HDL cholesterol, LDL cholesterol and total cholesterol, apo-A and apo-B were higher in normal pregnancies compared with hyperemetic pregnancies. There were no significant differences in apo-B/apo-A, HDL cholesterol/apo-A and total cholesterol/HDL cholesterol ratios between the hyperemetic patients and controls. A negative correlation was found between total cholesterol and serum thyroxine level. CONCLUSION: We found decreased levels of total cholesterol, LDL cholesterol, apo A and apo-B in hyperemetic patients and the same spontaneous abortion rate in the two groups in our study. PMID- 15280118 TI - Unopposed appetite (orexigenic) mechanisms in the near-term ovine fetus: central leptin does not inhibit sucrose ingestion. AB - OBJECTIVE: Leptin is produced in adipocytes and is present in the term fetus. In the adult, leptin acts centrally to inhibit neuropeptide Y-induced carbohydrate intake. We sought to examine if central leptin alters fetal ingestion of oral sucrose in the near-term ovine fetus. METHODS: Five pregnant ewes and fetuses were prepared with fetal vascular, sublingual and intracerebroventricular (ICV) catheters and esophageal electromyogram electrodes, and studied at 132 +/- 1 days' gestation (term 145-150 days). Following a 2-h baseline period, 10% sucrose was infused sublingually (0.25 ml/min) for the duration of the study. At time 4 h, leptin (0.075 mg/kg) was administered ICV and fetal swallowing was monitored for an additional 6 h. RESULTS: During the basal period, fetal swallowing averaged 0.7 +/- 0.1 swallows/min. Fetal swallowing increased significantly in response to 10% sucrose (1.2 +/- 0.1 swallows/min; p < 0.05). In response to ICV leptin, fetal swallowing remained significantly elevated at 2, 4 and 6 h (1.3 +/- 0.4, 1.4 +/- 0.3 and 1.5 +/- 0.2 swallows/min, respectively; p < 0.05 vs. control). CONCLUSIONS: These results indicate that central leptin inhibition of sucrose ingestion is not functional in the near-term fetus. We speculate that a leptin-mediated anorexigenic response is not present at birth, such that unopposed appetite stimulatory mechanisms in the newborn may facilitate rapid newborn weight gain despite high body fat levels. PMID- 15280119 TI - Non-invasive transabdominal uterine electromyography correlates with the strength of intrauterine pressure and is predictive of labor and delivery. AB - OBJECTIVE: The study was conducted to investigate whether the strength of uterine contractions monitored invasively by intrauterine pressure catheter could be determined from transabdominal electromyography (EMG) and to estimate whether EMG is a better predictor of true labor compared to tocodynamometry (TOCO). STUDY DESIGN: Uterine EMG was recorded from the abdominal surface in laboring patients simultaneously monitored with an intrauterine pressure catheter (n = 13) or TOCO (n = 24). Three to five contractions per patient and corresponding electrical bursts were randomly selected and analyzed (integral of intrauterine pressure; integral, frequency, amplitude of contraction curve on TOCO; burst energy for EMG). The Mann-Whitney test, Spearman correlation and receiver operator characteristics (ROC) analysis were used as appropriate (significance was assumed at a value of p < 0.05). RESULTS: EMG correlated strongly with intrauterine pressure (r = 0.764; p = 0.002). EMG burst energy levels were significantly higher in patients who delivered within 48 h compared to those who delivered later (median [25%/75%]: 96,640 [26,520-322,240] vs. 2960 [1560-10,240]; p < 0.001), whereas none of the TOCO parameters were different. In addition, burst energy levels were highly predictive of delivery within 48 h (AUC = 0.9531; p < 0.0001). CONCLUSION: EMG measurements correlated strongly with the strength of contractions and therefore may be a valuable alternative to invasive measurement of intrauterine pressure. Unlike TOCO, transabdominal uterine EMG can be used reliably to predict labor and delivery. PMID- 15280120 TI - Influence of gestational age and fetal heart rate on the fetal mechanical PR interval. AB - OBJECTIVE: The fetal mechanical PR interval obtained via pulsed Doppler has previously been demonstrated to correlate with electrocardiographic PR interval measured in the neonate. We sought to further analyze the influence of fetal heart rate and gestational age upon the fetal mechanical PR interval. METHODS: We searched our database for mechanical PR intervals, which were obtained during fetal echocardiography performed in our antenatal diagnostic unit. We included fetuses with a normal cardiac structural survey. The mechanical PR interval is measured from the A wave of the mitral valve to the beginning of ventricular systole corresponding to the opening of the aortic valve. Linear regression curves were generated to examine the correlation of mechanical PR interval with gestational age and fetal heart rate. Analysis of variance was used to compare the mean variation across three gestational age groups: 17-21.9 weeks (n = 24), 22-25.9 weeks (n = 52) and 26-38 weeks (n = 20). RESULTS: Mechanical PR intervals were measured in 96 fetuses with normal fetal echocardiography. The mechanical PR interval was 123.9 +/- 10.3 ms (mean +/- SD), with a range of 90-150 ms. Linear regression curves correlating mechanical PR interval with fetal heart rate and gestational age demonstrated a flat slope with R2 = 0.016, p = 0.22 and R2 = 0.0004, p = 0.85, respectively. The mechanical PR interval measured over the three gestational ages was as follows (mean +/- SD): 122.3 +/- 10.5 ms for 17 21.9 weeks; 125.0 +/- 9.6 ms for 22-25.9 weeks; and 123.1 +/- 11.9 ms for 26-38 weeks. Analysis of variance revealed no difference among the mechanical PR interval means measured over the three gestational age groups (p = 0.53). CONCLUSIONS: Fetal mechanical PR interval ranges from 90 to 150 ms in fetuses with sonographically normal fetal cardiac structure and rate. The mechanical PR interval appears to be independent of gestational age and fetal heart rate. PMID- 15280121 TI - The impact of chorionicity on umbilical cord acid-base values in twin gestations. AB - OBJECTIVE: To evaluate the impact of chorionicity on inter-twin differences in acid-base status at birth. METHODS: Records for twin pregnancies delivered at > or = 24 weeks' gestation from 1 January 1990 to 31 June 2000 were reviewed. Collected data included maternal demographics, gestational age, fetal presentation, anesthesia, delivery mode, inter-twin interval, umbilical artery (UA) and venous (UV) acid-base values, Apgar scores and birth weights. The influence of chorionicity on umbilical cord biochemistry was evaluated. (p < 0.05 was considered significant.) RESULTS: Analysis was carried out in 87 twin pairs (29 monochorionic, MC; and 58 dichorionic, DC). MC and DC twins were similar in maternal age (25.5 vs. 28.2 years), estimated gestational age (33.7 vs. 33.6 weeks), Cesarean delivery (55.2 vs. 52.6%), delivery interval (10 vs. 5 min) and respective birth weights (twin A, 1882 vs. 1981; and twin B, 1828 vs. 1872 g). MC first twins had a higher UA pH (7.31 +/- 0.05 vs. 7.26 +/- 0.08; p = 0.0005) than DC first twins. MC first and second twins had higher UA and UV bicarbonate levels than their DC counterparts (DeltapH = 21.7 +/- 5.1 vs. 18.5 +/- 3.1 mmol/l and 22.0 +/- 3.5 vs. 19.6 +/- 2.5 mmol/l, respectively; p = 0.003). MC twins were more discordant in UA pH than DC twins (DeltapH = 0.043 +/- 0.09 vs. 0.003 +/- 0.07; p = 0.009). MC and DC twins had a similar venous pH (DeltapH = 0.01 +/- 0.06 vs. 0.02 +/- 0.06; p = 0.5). CONCLUSIONS: There is a significant association between placental chorionicity and umbilical cord biochemistry in twins. Although it is possible that the mechanism of this finding is related to placental angioarchitecture, it is unlikely to be a result of simple mixing of blood volumes between twins. The physiology of underlying processes requires further study. PMID- 15280122 TI - Second-trimester amniotic fluid interleukin-10 concentration predicts preterm delivery. AB - OBJECTIVE: Interleukin-6 (IL-6) is an inflammatory cytokine that has been shown to be elevated in the amniotic fluid of patients with preterm labor. On the other hand, interleukin-10 (IL-10) is an anti-inflammatory cytokine that has been shown to inhibit the synthesis of other cytokines. We hypothesized that amniotic fluid IL-10 in the early second trimester is low in patients who subsequently develop preterm labor, and because of its deficiency, excessive inflammatory responses associated with IL-6 elevation lead to preterm labor and delivery. STUDY DESIGN: Amniotic fluid IL-6 and IL-10 levels were measured in 96 women who underwent genetic amniocentesis between 15 and 23 weeks' gestation. Levels of IL-6 and IL 10 were measured by immunoassay and correlated with demographic and pregnancy outcome information. RESULTS: Fifteen patients delivered at or before 36 weeks and 81 patients delivered after 36 weeks. There was an inverse correlation between amniotic fluid IL-10 concentration and gestational age at delivery. Similarly, an inverse correlation also existed between amniotic fluid IL-6 concentration and gestational age at delivery. CONCLUSIONS: Both IL-10 and IL-6 levels in second-trimester amniotic fluid obtained at the time of genetic amniocentesis appeared to be higher in patients who subsequently developed preterm delivery. Therefore, low amniotic fluid IL-10 production during the second trimester does not seem to be an etiology for preterm labor. PMID- 15280123 TI - Predictors of successful labor induction with oral or vaginal misoprostol. AB - OBJECTIVE: To identify independent predictors of successful labor induction with oral or vaginal misoprostol. METHODS: Women enrolled in four previous randomized trials involving oral or vaginal misoprostol for cervical ripening and labor induction were included in the present cohort study, with dosing of 25-50 microg every 4 to 6 h vaginally (n = 574) or 50 microg every 4 h orally (n = 207). Multiple logistic regression was performed to identify factors independently associated with successful labor induction -- defined as vaginal delivery within 12 h, vaginal delivery within 24 h and spontaneous vaginal delivery. Predictors of Cesarean birth and the need for only one dose of misoprostol were also identified. Variables included in the models were maternal age, weight, height, parity, gravidity, membrane status, route of misoprostol, gestational age, birth weight, and Bishop score and its individual components. RESULTS: Maternal age, height, weight, parity, birth weight, dilatation, effacement and cervical station were associated with vaginal delivery within 24 h of induction. Maternal age, height, weight, nulliparity, birth weight and route of misoprostol were associated with Cesarean birth, with oral misoprostol being associated with a lower rate of Cesarean birth. The need for only one dose of misoprostol was predicted by maternal height, weight, parity, gestational age, Bishop score and route of misoprostol. CONCLUSION: Characteristics of the woman (height, weight, parity), the fetus (birth weight) and some of the individual components of the Bishop score, were associated with successful labor induction, with oral misoprostol being associated with a lower rate of Cesarean birth. PMID- 15280124 TI - Foscarnet therapy for congenital cytomegalovirus liver fibrosis following prenatal ascites. AB - We report on an infant with multi-system disease including liver fibrosis, right microphthalmia with cataract, interstitial pneumonitis, and hyperechoic lesions in the basal ganglia and in the periventricular and thalamic regions. Prenatal ascites with hepatomegaly concomitantly with detection of cytomegalovirus (CMV) DNA in the amniotic fluid, following recurrent maternal CMV infection, had been shown. Although CMV culture and DNA detection were negative in the urine, the infant was given foscarnet because CMV infection was demonstrated in the liver by DNA detection and immunohistochemical staining. Favorable clinical outcome and absence of CMV in the liver were subsequently shown. Our case suggests that congenital CMV disease following maternal recurrence may not be associated with disseminated infection but only with intracellular infection. The diagnosis should therefore be based on CMV detection in the involved organs. Moreover, this is the first report on the possible efficacy and safety of foscarnet for therapy of immunocompetent infants with congenital CMV disease. PMID- 15280125 TI - The changing face of gastroschisis and omphalocele in southeast Georgia. AB - OBJECTIVE: To document trends in the clinical characteristics of gastroschisis and omphalocele in southeast Georgia, USA, from 1994 to 2002. METHODS: All babies with an abdominal wall defect in a 19-county region were referred to one Perinatal Center for genetic counseling, level II ultrasound scans, pregnancy follow-up and delivery. Karyotyping was offered for omphalocele, advanced maternal age, family history predisposing to aneuploidy, and gastroschisis with an additional anomaly. RESULTS: There were 64 patients, 34 with gastroschisis and 30 with omphalocele. From 1994 to 2002, the birth prevalence of gastroschisis was 1:3600 and omphalocele 1:3400, but from 2000 to 2002, gastroschisis increased to 1:1667, while omphalocele increased to only 1:2709. Gender distribution was different: for gastroschisis the M:F ratio was 1:2.1; for omphalocele the ratio was 1.7:1. In the patients with omphalocele, 90% had an amniocentesis and 9/27 were aneuploid: five had trisomy 18, three had trisomy 13 and one had trisomy 21. Seventy-six per cent of the patients with omphalocele had associated anomalies, but only 17.6% of those with gastroschisis. Mothers whose babies had gastroschisis showed a trend to progressively younger age, while no such trend was observed among mothers whose babies had omphalocele. CONCLUSION: The birth prevalence of abdominal wall defects in general is increasing, but more notably for gastroschisis. Maternal age continues to decrease for gastroschisis. In the study population, gender distribution showed a statistically significant variation between the defects. PMID- 15280126 TI - Methylenetetrahydrofolate reductase 677 C-->T polymorphism and plasma folate in relation to pre-eclampsia risk among Peruvian women. AB - OBJECTIVES: Pre-eclampsia is an important cause of maternal and fetal morbidity and mortality worldwide. Hyperhomocyst(e)inemia in pregnancy is associated with an increased risk of pre-eclampsia in most studies. Nutritional and genetic factors regulate homocyst(e)ine levels. A missense mutation 677 C-->T in the gene for methylenetetrahydrofolate reductase (MTHFR) has been associated with an increased pre-eclampsia risk in some, although not most, previously studied populations. METHODS: To further understand the role of this polymorphism in the etiology of pre-eclampsia, we genotyped a total of 125 pre-eclamptics and 179 normotensive pregnant Peruvian women. RESULTS: The wild-type allele frequency among cases and controls was 54% and 58%, respectively. Twenty percent of cases and 17% of controls were homozygous for the 677 C-->T MTHFR genotype (T/T). After adjustment for confounding by covariates including maternal age, nulliparity, pre pregnancy body mass index and use of prenatal vitamins, women homozygous for the 677 C-->T MTHFR genotype (T/T) experienced a modest, statistically non significant increased risk of pre-eclampsia (adjusted OR 1.6, 95% CI 0.7, 3.8). Maternal folate deficiency was associated with a statistically non-significant doubling in risk of pre-eclampsia in this population (adjusted OR 2.0, 95% CI 0.9, 4.3). CONCLUSIONS: There was no evidence to suggest that pre-eclampsia risk is positively associated with the T/T genotype overall, or in the context of folate deficiency. PMID- 15280128 TI - The impact of route of delivery and presentation on twin neonatal and infant mortality: a population-based study in the USA, 1995-97. AB - OBJECTIVE: We examined whether the route of delivery for near-term (> or = 34 weeks' gestation) twins, as candidates for vaginal delivery, affected neonatal and infant mortality rates. We further evaluated whether these mortality rates were modified by fetal presentation. METHODS: A population-based retrospective cohort study based on the matched multiple births data in the USA (1995-97) was performed. Analyses were restricted to non-malformed liveborn twins delivered at (> or = 34 weeks' gestation. Twins with breech-breech and breech-vertex presentations were excluded, since they are not candidates for vaginal delivery. Neonatal mortality rates (death within the first 27 days) and post-neonatal mortality rates (death between 28 and 365 days) per 1000 twin live births, by route of delivery and fetal presentation, were derived. The associations between neonatal mortality, post-neonatal mortality and the route of delivery for vertex breech versus vertex-vertex presentations were expressed based on relative risks (RR) and 95% confidence intervals (CI) derived from logistic regression models based on the method of generalized estimating equations. RESULTS: Of the 177,622 twins analyzed, 87% (n = 154,531) presented as vertex-vertex. Fifty-five per cent (n = 97,692) of twins were both delivered vaginally, 41% (n = 72,825) were both delivered by Cesarean section and, of the remaining 4% (n = 7,105), the first twin was delivered vaginally and the second by Cesarean section. Twins with vertex-breech presentations delivered by Cesarean-cesarean sections, as well as those with vertex-vertex presentations delivered vaginally, had the lowest neonatal mortality rate (1.6 per 1000 live births). The highest neonatal mortality rate in the vertex-breech pairs occurred with vaginal-Cesarean deliveries (2.7 per 1000 live births). Among twins with vertex-vertex presentations, twins delivered via the vaginal-Cesarean route experienced the highest neonatal mortality (3.8 per 1000 live births). The RR for neonatal mortality in this group was 2.24 (95% CI 1.35, 3.72) compared with twins both delivered vaginally. CONCLUSION: Route of delivery and fetal presentation both confer an impact on twin infant mortality rates. Strategies to reduce discordant routes in complicated vaginal deliveries may lead to improved neonatal survival. PMID- 15280129 TI - Approach for estimating fetal body weight using two-dimensional ultrasound. AB - OBJECTIVE: Estimation of fetal weight is essential in daily obstetric clinical practice. Most formulas for the estimation involve head measurement. However, predicting fetal weight by head measurement is virtually impossible when the fetal head is positioned low in the pelvic cavity. A convenient method for estimating fetal body weight without head measurement is therefore required. METHODS: Women who delivered between August 2001 and June 2002 in our center were the subjects of the present study. All infants were delivered within 48 h of an ultrasound examination. Only thigh measurements were made in an attempt to obtain an estimation formula by conventional two-dimensional ultrasonography in 83 patients. As a parameter, femur length (FL) and the largest cross-sectional area at right angles to the long axis of the thigh were used. The derived formula was compared with an established equation in 58 parturients. RESULTS: The FL value multiplied by the square root of the cross-sectional area of the thigh showed a significant correlation with the actual birth weight. A formula with only two parameters (FL and cross-sectional area of the thigh) was found by linear regression. CONCLUSION: The formula derived using only thigh measurements was found to be convenient among all the established formulas for estimated fetal body weight, and no head measurement was necessary. PMID- 15280130 TI - Serial assessment of amniotic fluid index in uncomplicated term pregnancies: prognostic value of amniotic fluid reduction. AB - OBJECTIVE: Assessment of amniotic fluid volume in association with a non-stress test is a commonly used method to monitor fetal well-being in high-risk pregnancies. The aims of our study were to determine whether oligohydramnios and the trend in amniotic fluid volume have prognostic significance in low-risk pregnancies between 40.0 and 41.6 weeks' gestation. METHODS: Between January 1997 and December 2000, all uncomplicated gestations with a singleton non-anomalous fetus reaching 40.0 weeks' gestation underwent semi-weekly monitoring of amniotic fluid index (AFI) until delivery. Oligohydramnios was defined as an AFI of < or = 5 cm. Changes in AFI were expressed as centimeters per day, and were calculated as: [(last AFI before delivery minus first AFI at 40.0 weeks) / interval in days between the two scans]. Adverse outcome was considered the occurrence of 5-min Apgar score < 7; umbilical artery pH < 7.0; Cesarean section for fetal distress; or fetal death. Comparisons between the groups with favorable and adverse outcomes was performed with chi(2) or Fisher's exact test for categorical variables, and Student's t test for continuous variables. A two-tailed p value < 0.05 was considered significant. RESULTS: A total of 3050 women met the study criteria, and underwent a median number of two (range 1-7) sonographic assessments of AFI after 40.0 weeks, with oligohydramnios detected in 341 women. In 1466 women at least two serial AFI determinations were obtained, allowing computation of an AFI trend. Gestations resulting in adverse perinatal outcome (n = 167, 5.5%) had a significantly higher rate of oligohydramnios (33/167, 19.8% vs. 308/2883, 10.7%, p = 0.001), but a similar rate of reduction in AFI ( -0.65 +/- 0.64 vs. - 0.66 +/- 0.66 cm/day; p = 0.85) than those with favorable outcome. The difference in rate of reduction of AFI between the two groups was not significant, even in the subset of gestations that developed oligohydramnios ( 1.08 +/- 0.87 vs. -1.26 +/- 0.89 cm/day; p = 0.27). CONCLUSION: A sonographic diagnosis of oligohydramnios carries an increased risk of adverse perinatal outcome, even in low-risk pregnancies after 40 weeks. The trend in amniotic fluid volume reduction does not seem to have prognostic significance. PMID- 15280131 TI - Perinatal outcomes in inflammatory bowel disease. AB - OBJECTIVE: To determine whether inflammatory bowel disease (IBD) is associated with increased risk for adverse perinatal outcome. METHODS: A case-control study of 116 singleton pregnancies with IBD compared to 56,398 singleton controls delivered between 1986 and 2001. RESULTS: Patients with IBD were slightly older (32.8 vs. 30.6 years, p < 0.001), more likely to be Caucasian or Asian than Black or Latino (92% vs. 57%, p < 0.001) and have private health insurance (33% vs. 3%, p < 0.001). IBD was associated with an increased risk for labor induction (32% vs. 24%, p = 0.002), chorioamnionitis (7% vs. 3%, p = 0.04) and Cesarean section (32% vs. 22%, p = 0.007), but there were no differences in neonatal outcomes. Subgroup analysis demonstrated an increased risk for low birth weight (LBW) in the ulcerative colitis group vs. the Crohn's disease group (19% vs. 0%, p = 0.002). Patients with prior surgery for IBD had a lower incidence of LBW (0% vs. 12%, p = 0.03). Flares during pregnancy were associated with an increased risk for preterm delivery (27% vs. 8%, p = 0.02) and LBW (32% vs. 3%, p = 0.003). CONCLUSION: IBD was an independent risk factor for Cesarean section but there was no increase in adverse perinatal outcome. Crohn's disease, prior IBD surgery and quiescent disease were associated with a lower risk for LBW. PMID- 15280132 TI - Maternal and neonatal morbidity after elective repeat Cesarean delivery versus a trial of labor after previous Cesarean delivery in a community teaching hospital. AB - OBJECTIVE: To compare maternal and fetal outcomes after elective repeat Cesarean section versus a trial of labor in women after one prior uterine scar. STUDY DESIGN: All women with a previous single low transverse Cesarean section delivered at term with no contraindications to vaginal delivery were retrospectively identified in our database from January 1995 to October 1998. Outcomes were first analyzed by comparing mother-neonate dyads delivered by elective repeat Cesarean section to those undergoing a trial of labor. Secondarily, outcomes of mother-neonatal dyads who achieved a vaginal delivery or failed a trial of labor were compared to those who had elective repeat Cesarean delivery. RESULTS: Of 1408 deliveries, 749/927 (81%) had a successful vaginal birth after a prior Cesarean delivery. There were no differences in the rates of transfusion, infection, uterine rupture and operative injury when comparing trial of labor versus elective repeat Cesarean delivery. Neonates delivered by elective repeat Cesarean delivery were of earlier gestation and had higher rates of respiratory complications (p < 0.05). Mother-neonatal dyads with a failed trial of labor sustained the greatest risk of complications. CONCLUSION: Overall, neonatal and maternal outcomes compared favorably among women undergoing a trial of labor versus elective repeat Cesarean delivery. The majority of morbidity was associated with a failed trial of labor. Better selection of women likely to have a successful vaginal birth after a prior Cesarean delivery would be expected to decrease the risks of trial of labor. PMID- 15280133 TI - Acupuncture plus moxibustion to resolve breech presentation: a randomized controlled study. AB - OBJECTIVE: In many Western countries breech presentation is an indication for elective Cesarean section. In order to correct fetal presentation, the stimulation of the acupoint BL67 by moxibustion, acupuncture or both has been proposed. Since no studies had previously been carried out on Western populations, pregnant Italian women at 33-35 weeks gestational age carrying a fetus in breech presentation were enrolled in a randomized, controlled trial involving an active BL67 point stimulation and an observation group. METHODS: A total of 240 women at 33-35 weeks of gestation carrying a fetus in breech presentation were randomized to receive active treatment (acupuncture plus moxibustion) or to be assigned to the observation group. Bilateral acupuncture plus moxibustion was applied at the BL67 acupoint (Zhiyin). The primary outcome of the study was fetal presentation at delivery. RESULTS: Fourteen cases dropped out. The final analysis was thus made on 226 cases, 114 randomized to observation and 112 to acupuncture plus moxibustion. At delivery, the proportion of cephalic version was lower in the observation group (36.7%) than in the active-treatment group (53.6 %) (p = 0.01). Hence, the proportion of Cesarean sections indicated for breech presentation was significantly lower in the treatment group than in the observation group (52.3% vs. 66.7%, p = 0.03). CONCLUSIONS: Acupuncture plus moxibustion is more effective than observation in revolving fetuses in breech presentation. Such a method appears to be a valid option for women willing to experience a natural birth. PMID- 15280134 TI - The impact of antenatal cocaine use on maternal characteristics and neonatal outcomes. AB - OBJECTIVE: To determine the risk factors and evaluate maternal and neonatal outcomes associated with antenatal cocaine use. METHODS: This was a retrospective case-control study of 200 cocaine-exposed maternal-neonatal pairs and 200 controls from 1991 to 2000. RESULTS: Cocaine-using mothers tended to be older, African American, multiparous and incarcerated and they utilized less prenatal care. However, 79% of Hispanics abusing cocaine were primarily English speaking. Cocaine use correlated with syphilis (36 vs. 1%, p = 0.000) and premature rupture of membranes (23 vs. 0%, p = 0.000), fetal demise (5 vs. 0%, p = 0.004), preterm delivery (40 vs. 6%, p = 0.000). Cocaine-exposed infants delivered earlier (36 vs. 39 weeks, p = 0.000), had lower birth weights (2660 vs. 3305 g, p = 0.000), more respiratory distress syndrome (14 vs. 4%, p = 0.001), congenital syphilis (12 vs. 1%, p = 0.000) and longer hospital stays (10 vs. 3 days, p = 0.000); 75% were placed in foster care or adoption and 37.5% had neonatal withdrawal syndrome. There was a stronger positive correlation between neonatal withdrawal and maternal urine toxicology (rho = 0.443, p = 0.000) than with neonatal urine screen (rho = 0.278, p = 0.003). CONCLUSION: Cocaine use in pregnancy is associated with acculturation, lack of prenatal care, and significant social and obstetric complications resulting in increased neonatal morbidity secondary to prematurity, congenital infection and withdrawal syndrome. PMID- 15280135 TI - Synchronized ventilation of very-low-birth-weight infants; report of 6 years' experience. AB - OBJECTIVES: To evaluate the effects of long-term patient triggered ventilation (PTV) using assist/control or synchronized intermittent mandatory ventilation (SIMV) in very-low-birth-weight infants with respiratory distress. METHODS: Ninety-seven very-low-birth-weight infants who had undergone synchronized ventilation for respiratory distress or insufficiency were assessed from January 1995 to December 2000. Death, oxygen support, pneumothorax development while ventilated, intracranial hemorrhage, necrotizing enterocolitis, periventricular leukomalacia, retinopathy of prematurity and duration of ventilation were noted as the mean outcome measures. RESULTS: The mean birth weight was 1139 +/- 268 g (range 450-1500 g) and the mean gestational age was 29.0 +/- 2.8 weeks (range 23 36 weeks). Eighty-four per cent of 97 infants survived. Antenatal steroids were administered to only 20% of mothers. Surfactant was administered to all of the 67% of infants with respiratory distress syndrome. The mean duration of ventilator support was 4.7 +/- 7.3 days (1-43 days) for survivors and 8.9 +/- 11 days (1-45 days) for infants who died. No respiratory paralysis was necessary in any case during ventilation and pneumothorax was diagnosed in only eight infants. Severe intracranial hemorrhage (grade > or = III) and periventricular leukomalacia developed in 15% and 12% of infants, respectively. Necrotizing enterocolitis (Bell's classification stage > or = 2) and retinopathy of prematurity were noted in two infants. Four infants had evidence of chronic lung disease. The rate of survival without major morbidity was 83.5%. CONCLUSION: Patient-triggered ventilation, initially PTV with Asist/Control and subsequently with SIMV in very-low-birth-weight infants with respiratory distress is feasible, but optimization of trigger and ventilator performance with respect to respiratory diagnosis is essential. PMID- 15280136 TI - The international infections in pregnancy study: group B streptococcal colonization in pregnant women. AB - BACKGROUND: Heavy colonization with group B streptococcus (GBS) has been associated with increased risk of preterm birth and neonatal sepsis; the burden of neonatal GBS disease varies geographically. To determine whether variation in heavy colonization and GBS serotypes could contribute to geographic differences in disease burden, we assessed the prevalence of heavy colonization and the distribution of serotypes in asymptomatic pregnant women in multiple countries. METHODS: Cervical, lower vaginal and urine samples were collected from women attending seven prenatal clinics in six countries. Light colonization was defined as GBS isolation from Lim broth only; heavy colonization was isolation from urine or sheep blood agar plates. Isolates were serotyped using capillary precipitation. RESULTS: GBS was present in 11.3% of 1308 participants (range 7.1 21.7%); 5.0% were heavily colonized (0.4-18.8%) and 6.4% were lightly colonized (2.9-8.0%). Serotypes III and V were most common (both 17.2%). Serotypes VII and VIII were found in one study center. CONCLUSIONS: The prevalence of heavy colonization and GBS serotypes varied significantly among our study centers. Whether this variation could in part explain geographic differences in neonatal morbidity and mortality is a hypothesis that needs further study. PMID- 15280137 TI - Placenta increta; conservative management -- a successful outcome. Case report and literature review. AB - Placenta increta, a rare complication of pregnancy, is associated with significant postpartum hemorrhage often requiring emergency hysterectomy. We report a case of conservative management, with a combination of parenteral methotrexate, serial ultrasound and Doppler assessment, followed by interval manual removal of placenta. PMID- 15280138 TI - Mother-infant health promotion in developing countries: how can the developed world help developing countries? PMID- 15280139 TI - Spontaneous minute ventilation is a predictor of extubation failure in extremely low-birth-weight infants. AB - OBJECTIVE: To validate the percentage of time spent below a target value of spontaneous expiratory minute ventilation (< 125 ml/min per kg) during a 2-h period of continuous positive airway pressure (CPAP) via an endotracheal tube (ETT) as a predictor of failed extubation in preterm infants. METHODS: Forty-one infants intubated for at least 24 h, with birth weight between 500 and 1000 g, who were clinically stable and at ventilator setting compatible with an extubation attempt, were studied during a 2-h period of ETT CPAP. Dynamic lung compliance and total lung resistance were measured during a period of quiet breathing, while tidal volume (Vt), respiratory rate and the corresponding spontaneous expiratory minute ventilation values were calculated for the complete recording period of 2 h using a customized computer program. The time each patient spent below the target spontaneous expiratory minute ventilation value was reported as a percentage of the total recorded time (% spontaneous expiratory minute ventilation < 125 ml/min per kg). Extubation failure was defined as the need for reintubation within 72 h. RESULTS: Eleven out of 41 babies (26.8%) experienced failure of extubation (failure group) while 30 infants (73.2%) were successfully extubated (success group). There were no significant differences in dynamic lung compliance and lung resistance between the two groups, but the mean values of respiratory rate and spontaneous expiratory minute ventilation were significantly lower in the failure group than in the success group: 43 (37-56) breaths/min and 240 (160-353) ml/min per kg vs. 53 (28-67) breaths/min and 309 (223-434) ml/min per kg, respectively (p = 0.0129 and p = 0.0039). Moreover, the babies in whom extubation failed spent a longer time below the target value of spontaneous expiratory minute ventilation when compared with successfully extubated babies (p < 0.0001). Percentage of time spent with spontaneous expiratory minute ventilation < 125 ml/min per kg had a larger area than transcutaneous (Tc)PCO2, TcPO2 and pulse oxymetry saturation (SpO2) under the receiver operator characteristic curves. CONCLUSION: The measurement of spontaneous expiratory minute ventilation prior to extubation could be useful in identifying those babies who are not ready for spontaneous ventilation. PMID- 15280140 TI - In vitro quantification of dexamethasone-induced surfactant protein B expression in human lung cells. AB - OBJECTIVE: To determine whether the effect of a single 48-h exposure to dexamethasone in human lung cells is limited to 7-8 days. STUDY DESIGN: We used the NCI-H441 cell line, in which stability can be maintained beyond 7 days. The outcome was the stimulatory effect of dexamethasone on surfactant protein B (SP B) gene transcription as expressed by SP-B mRNA accumulation. The experiment was conducted five times, in parallel with control. SP-B mRNA was determined at baseline, 48 h after dexamethasone exposure, and at 48-h intervals thereafter, up to 14 days, by quantitative reverse transcription polymerase chain reaction. Comparisons were made by the Mann-Whitney test. RESULTS: In conditions of our experiment, the inductive profile of SP-B mRNA after exposure to dexamethasone demonstrated maximal stimulation at 48 h (13-fold over control). Subsequently, there was a decline in mRNA, with return to near control levels by day 8, suggesting reversibility of dexamethasone action. CONCLUSION: Our data support the view that the surfactant-inducing properties of corticosteroids are limited to 7-8 days. PMID- 15280141 TI - Preliminary estimate for the second-trimester maternal serum screening detection rate of the 45,X karyotype using alpha-fetoprotein, unconjugated estriol and human chorionic gonadotropin. AB - OBJECTIVE: To estimate the detection rate for 45,X pregnancies through second trimester screening using maternal serum alpha-fetoprotein, human chorionic gonadotropin and unconjugated estriol. METHODS: Twenty-two cases of 45,X were ascertained through a cytogenetics database and an additional 51 cases were identified through publications. Serum analyte concentrations were reviewed for cases with fetal hydrops, cystic hygroma alone, and no evidence of edema. Using the statistical characteristics of this sample of affected pregnancies, computer simulations were carried out to determine the proportion of 45,X pregnancies that should be screen-positive for Down syndrome and trisomy 18. The extent to which additional cases of 45,X might be identified using a protocol specifically designed to detect 45,X pregnancies was also estimated. RESULTS: Approximately 54% of all 45,X pregnancies should be identifiable through screening for Down syndrome and trisomy 18. The detection rate for cases with hydrops and/or cystic hygroma was 60%, and without edema 33%. If offered with screening for Down syndrome and trisomy 18, 45,X screening could identify approximately 7% more of the affected pregnancies with an incremental rise of 0.2% in the false-positive rate. CONCLUSIONS: A screening algorithm for 45,X could be developed. However, the number of additional affected pregnancies identified would appear to be too small to justify this screening. PMID- 15280142 TI - Randomized clinical trial comparing two natural surfactant preparations to treat respiratory distress syndrome. AB - OBJECTIVES: Natural surfactant preparations have been shown to reduce the severity and mortality of respiratory distress syndrome (RDS) in preterm infants. The objective of this study was to compare the efficacy of two natural surfactants, namely SF-RI 1 (Alveofact) and barectant (Survanta), on the incidence of chronic lung disease (CLD) and other associated complications of RDS in preterm infants. METHODS: Preterm infants with RDS requiring artificial ventilation were randomly selected to receive an initial dose of either Alveofact or Survanta. The two treatment groups were tested for variation in gas exchange, ventilatory settings and neonatal complications such as CLD and mortality. RESULTS: After 5 days the Survanta-treated infants had a lower fraction of inspired oxygen (FiO2) compared with the Alveofact-treated infants. There were no differences in the ventilatory settings. More infants in the Survanta group were extubated at 3 days and fewer required the use of postnatal steroids. Less CLD and duration of oxygenation were experienced by the Survanta-treated group. CONCLUSIONS: Improved oxygenation and reduced ventilatory requirements were greater with Survanta compared to Alveofact, which in turn was associated with a trend towards a lower incidence of serious pulmonary complications. PMID- 15280143 TI - First-trimester maternal serum activin A in pre-eclampsia and fetal growth restriction. AB - OBJECTIVE: To investigate whether the reported increase in maternal serum activin A concentration in pre-eclampsia is evident from the first trimester. DESIGN: This was a case-control study carried out in antenatal clinics among singleton pregnancies at 10-14 weeks of gestation. METHODS: Activin A concentration was measured in stored maternal serum samples obtained at 11-14 weeks of gestation from 131 women who subsequently developed pre-eclampsia, 77 who developed non proteinuric pregnancy-induced hypertension, 141 with fetal growth restriction in the absence of hypertensive complications and from 494 normotensive controls. RESULTS: Compared to the median activin A level in the control group (1.00 MoM), the median MoM in the patients who subsequently developed pre-eclampsia and pregnancy-induced hypertension (1.49 MoM and 1.32 MoM, respectively) was significantly increased (p < 0.001), and in patients with fetal growth restriction (1.02 MoM) it was not significantly different (p = 0.57). In the pre eclampsia group (n = 131) the disease was considered to be sufficiently severe to necessitate iatrogenic delivery before 35 weeks in 25 patients, and in this group the median MoM was 1.92. CONCLUSION: Maternal serum activin A concentration at 12 weeks of gestation in pregnancies which subsequently develop hypertensive disease is increased, whereas in those complicated by fetal growth restriction it is normal. PMID- 15280144 TI - ST-segment analysis of the fetal electrocardiogram improves fetal heart rate tracing interpretation and clinical decision making. AB - INTRODUCTION: Since its introduction into clinical use, the efficacy of electronic fetal heart rate (FHR) monitoring (EFM) has been questionable. This has been due partly to the marked variation in interpretation of the FHR pattern and subsequent decisions for obstetric intervention, (e.g., the need for prompt delivery). Current application of EFM is limited to the assessment of FHR patterns and uterine contractions. Recent development of higher-order FHR analysis has yielded monitoring systems that can add automated fetal electrocardiographic ST segment analysis to the standard FHR and uterine contraction information. Our goal was to evaluate the effect of adding ST segment analysis to standard FHR information on observer agreement for clinical decision making. METHODS: Seven practitioners who were trained and experienced in combined FHR and ST monitoring reviewed 51 fetal monitor tracings, ranging from 2 to 4 h in length. Reviews were conducted in two sessions and at different times. The first session presented only the FHR and uterine contraction information, following which the participants determined the time at which intervention (decision for operative vaginal or Cesarean section delivery) was indicated. In the second session, the participants were provided with a randomized sequence of the same tracings with the addition of ST segment information, as produced by the STAN monitor system (Neoventa Medical, Gothenburg, Sweden). Observer agreement was based on the proportion of participants who agreed on the need for an intervention, and the per cent agreement on the timing of the intervention within 20 min before or after the median time of intervention. RESULTS: Of the 51 cases included in this study there were ten fetuses with umbilical artery (UA) pH between 7.05 and 7.14, and nine with UA pH of < 7.05. Observer agreement increased significantly for required intervention when the ST segment information was available for tracing analysis as compared with review of the standard tracing alone (0.96 vs. 0.80, p < 0.05) and the timing of intervention (0.92 vs. 0.66, respectively, p < 0.05). Similarly, correct identification for needed interventions on fetuses with abnormal outcomes increased from 86 to 93% while unneeded interventions on normal fetuses decreased from 43 to 6%. CONCLUSION: The addition of ST analysis to standard FHR monitoring improves observer consistency in both the decision for and timing of obstetric interventions. The incorporation of ST segment data with the standard FHR tracing may reduce the number of unneeded obstetric interventions when fetal compromise is absent. PMID- 15280145 TI - Relationship of maternal plasma leptin and risk of pre-eclampsia: a prospective study. AB - OBJECTIVE: We measured maternal plasma leptin concentrations in 55 women with pre eclampsia and 487 normotensive women to determine whether elevated leptin concentrations were associated with the occurrence of pre-eclampsia. METHODS: Maternal blood samples were collected at 13 weeks' gestation, on average. Plasma leptin concentrations were determined using immunoassay. Logistic regression procedures were used to calculate odds ratios (OR) and 95% confidence intervals (95% CI). RESULTS: Leptin concentrations were 78% higher in cases than control subjects (median 34.6 vs. 19.5 ng/ml; p < 0.001). Relative to women with leptin concentrations of < 27.4 ng/ml, those with elevated leptin concentrations (> or = 27.4 ng/ml) experienced a 2.3-fold increased risk of pre-eclampsia (OR 2.3; 95% CI 1.1-4.6). We observed evidence of a strong linear component of trend in risk of pre-eclampsia with increasing maternal plasma leptin concentration. Each 10 ng/ml increase in leptin concentration was associated with a 30% increase in pre eclampsia risk (OR 1.3; 95% CI 1.1-1.5). Overweight women with elevated leptin concentrations experienced the highest risk of pre-eclampsia (OR 6.4; 95% CI 3.1 13.2) as compared with lean women with no leptin elevations. CONCLUSION: Elevated plasma leptin concentration and maternal overweight status appear to be independently associated with an increased risk of pre-eclampsia. PMID- 15280146 TI - Maternal age and risk of fetal death in singleton gestations: USA, 1995-2000. AB - OBJECTIVE: To determine the magnitude of risk for fetal death among singleton pregnancies in relation to maternal age, and to compare the risks with other common indications for fetal testing. STUDY DESIGN: We performed a retrospective cohort analysis of singleton births delivered between 1995 and 2000 using the US linked birth/infant death data. Gestational age at < 24 weeks and fetuses with anomalies were excluded. Fetal death rates at > or = 24 and > or = 32 weeks were calculated among women aged 15-19, 20-24, 25-29, 30-34, 35-39, 40-44 and 45-49 years, as well as for other common indications for testing: chronic and pregnancy induced hypertension, diabetes and small-for-gestational age (SGA). The association between maternal age and fetal deaths was derived after adjusting for potential confounders through multivariable logistic regression models. Relative risks (RR) and 95% confidence intervals (CI) were derived from these models after adjusting for the effects of gravidity, race, marital status, prenatal care, education, smoking and placental abruption. RESULTS: Among the 21,610,873 singleton births delivered at > or = 24 weeks, fetal deaths occurred in 58,580 (2.7 per 1000). Births to young (15-19 years) and older (> or = 35 years) women comprised 12.6% and 11.4%, respectively. Compared with women aged 20-24 years, young women did not experience an increased risk of fetal death. However, increasing rates of fetal death at > or = 24 and at > or = 32 weeks were seen with increasing maternal age. The RR for fetal death at > or = 24 and at > or = 32 weeks among women 35-39 years were 1.21 and 1.31, respectively, while the RRs were 1.62 and 1.67 among women aged 40-44 years. Women 45-49 years were 2.40-fold (95% CI 1.77, 3.27) and 2.38-fold (95% CI 1.64, 3.46) as likely to deliver a stillborn fetus at > or = 24 weeks and > or = 32 weeks, respectively. RRs for fetal death at > or = 24 and > or = 32 weeks for hypertensive disease, diabetes, and SGA ranged between 1.46 and 4.95. CONCLUSION: Fetal deaths are increased among older women (> or = 35 years). Fetal testing in women of advanced maternal age may be beneficial. PMID- 15280147 TI - Efficacy of an acidic vaginal gel on vaginal pH and interleukin-6 levels in low risk pregnant women: a double-blind, randomized placebo-controlled trial. AB - BACKGROUND: Increased interleukin-6 (IL-6) levels and a vaginal pH of > 4.7 are associated with obstetric complications such as preterm delivery and low birth weight. Topical treatments, able to maintain a physiological vaginal pH, could help in the prevention of vaginal infections. STUDY AIM: In a randomized, double blind, placebo-controlled trial, we evaluated the effects of an acidic buffering vaginal gel (Miphil) on vaginal pH and IL-6 levels in pregnant women. PATIENTS AND METHODS: Seventy low-risk women pregnant with a singleton (second trimester) were enrolled in the trial. Thirty-five were randomized to the acidic gel, 2.5 g every 3 days for 12 weeks, and 35 to the corresponding placebo. Vaginal pH and vaginal IL-6 level were measured at baseline and after 12 weeks. Women were then followed until delivery. The main outcome measures were vaginal pH, vaginal pH normalization (pH < 4.5) and vaginal IL-6 levels. RESULTS: Vaginal pH at baseline was 4.6 +/- 0.4 and 4.4 +/- 0.3 in the acidic gel and the placebo group, respectively. At baseline, a total of 40% (14/35) and 22% (8/35) of women in each group, respectively, had a vaginal pH of > or = 4.7. At week 12, the vaginal pH was 4.3 +/- 0.3 in the acidic gel group and 4.3 +/- 0.3 in the placebo group (NS). The acidic gel normalized the vaginal pH in ten out of 14 women (p = 0.04) in comparison with only one out of eight women in the placebo group (NS). The acidic gel induced a significant (p < 0.02) reduction of vaginal IL-6 from 12.0 +/- 7 to 8.9 +/- 5 pg/l (-36%). In the placebo group, IL-6 increased from 9.0 +/- 5 to 13.5 +/- 6.8 pg/l (+50%) (p = 0.05). Birth weight was 2978 +/- 700 g in the placebo group and 3241 +/- 477 g in the acidic gel group (p = 0.06). CONCLUSIONS: The use of the acidic gel in low-risk pregnant women is able to maintain a physiological vaginal ecosystem and prevents the increases of vaginal pH and vaginal IL-6. Prospective and controlled trials are warranted to evaluate whether this acidic gel can reduce obstetric complications linked to vaginal inflammation during pregnancy. PMID- 15280148 TI - The effect of gestational age on trial of labor after Cesarean section. AB - OBJECTIVES: To evaluate the effect of gestational age on the rate of successful vaginal delivery and the rate of uterine rupture in patients undergoing a trial of labor (TOL) after a prior Cesarean delivery. STUDY DESIGN: This was a cohort study including patients with a live singleton fetus undergoing a TOL after a previous low transverse Cesarean delivery between 1988 and 2002. Patients were divided into three groups according to gestational age: 24-36 weeks 6 days, 37-40 weeks 6 days and > or = 41 weeks. Obstetric outcomes, including the rates of successful vaginal delivery and symptomatic uterine rupture, were compared between the groups. Multivariate logistic regression analysis was performed to adjust for potential confounding factors. RESULTS: There were 253, 1911 and 329 patients in each group, respectively. In patients with advanced gestational age (> or = 41 weeks) the rate of uterine rupture was significantly higher (0% vs. 1.0% vs. 2.7%, p = 0.006) and the rate of successful vaginal deliveries was significantly lower (83% vs. 76.9% vs. 62.6%, p < 0.001). After adjusting for confounding variables, advanced gestational age was associated with a lower rate of successful vaginal delivery (odds ratio 0.68, 95% CI 0.51-0.89), and a higher rate of uterine rupture (odds ratio 2.85, 95% CI 1.27-6.42) when compared to 37 40 weeks 6 days. CONCLUSION: Advanced gestational age is associated with higher rates of failed TOL and uterine rupture. PMID- 15280149 TI - Early exploration in perinatal torsion of the spermatic cord: a case report. AB - Perinatal testicular torsion is a rare condition presenting with signs of an acute scrotum. Review of the literature indicates that there is controversy about optimal management, particularly as regards the risks and the benefits of immediate versus delayed surgical treatment. We report a case that was treated by immediate surgical intervention. PMID- 15280150 TI - Cardiac mitochondrial damage and biogenesis in a chronic model of type 1 diabetes. AB - Diabetic cardiomyopathy is a common complication leading to heightened risk of heart failure and death. In the present report, we performed proteomic analysis on total cardiac proteins from the OVE26 mouse model of type 1 diabetes to identify protein changes that may contribute to diabetic cardiomyopathy. This analysis revealed that a surprising high proportion (12 of 20) of the altered proteins that could be identified by mass spectrometry were of mitochondrial origin. All but one of these proteins were upregulated by diabetes. Quantitative RT-PCR, performed for two of these proteins, indicated that part of the upregulation was attributed to increased messenger RNA levels. Morphological study of diabetic hearts showed significantly increased mitochondrial area and number as well as focal regions with severe damage to mitochondria. Diabetic mitochondria also showed reduced respiratory control ratio (9.63 +/- 0.20 vs. 6.13 +/- 0.41, P < 0.0001), apparently due to reduced state 3 rate, and diminished GSH level (5.5 +/- 0.9 vs. 8.2 +/- 2.5 micromol/mg protein, P < 0.05), indicating impaired mitochondrial function and increased oxidative stress. Further examination revealed increased mitochondrial DNA (1.03 +/- 0.18 vs. 0.69 +/- 0.13 relative copy number, P < 0.001) and a tendency to higher protein yield in OVE26 cardiac mitochondria, as well as increased mRNA level for mitochondrial transcription factor A and two mitochondrial encoded proteins. Taken together, these results show that mitochondria are a primary target in the diabetic heart, probably due to oxidative stress, and that this damage coincides with and may stimulate mitochondrial biogenesis. PMID- 15280151 TI - Effects of gender and GH secretory pattern on sterol regulatory element-binding protein-1c and its target genes in rat liver. AB - We investigated whether the sexually dimorphic secretory pattern of growth hormone (GH) in the rat regulates hepatic gene expression of sterol regulatory element-binding protein-1c (SREBP-1c) and its target genes. SREBP-1c, fatty acid synthase (FAS), and glycerol-3-phosphate acyltransferase (GPAT) mRNA were more abundant in female than in male livers, whereas acetyl-CoA carboxylase-1 (ACC1) and stearoyl-CoA desaturase-1 (SCD-1) were similarly expressed in both sexes. Hypophysectomized female rats were given GH as a continuous infusion or as two daily injections for 7 days to mimic the female- and male-specific GH secretory patterns, respectively. The female pattern of GH administration increased the expression of SREBP-1c, ACC1, FAS, SCD-1, and GPAT mRNA, whereas the male pattern of GH administration increased only SCD-1 mRNA. FAS and SCD-1 protein levels were regulated in a similar manner by GH. Incubation of primary rat hepatocytes with GH increased SCD-1 mRNA levels and decreased FAS and GPAT mRNA levels but had no effect on SREBP-1c mRNA. GH decreased hepatic liver X receptor-alpha (LXRalpha) mRNA levels both in vivo and in vitro. Feminization of the GH plasma pattern in male rats by administration of GH as a continuous infusion decreased insulin sensitivity and increased expression of FAS and GPAT mRNA but had no effect on SREBP-1c, ACC1, SCD-1, or LXRalpha mRNA. In conclusion, FAS and GPAT are specifically upregulated by the female secretory pattern of GH. This regulation is not a direct effect of GH on hepatocytes and does not involve changed expression of SREBP-1c or LXRalpha mRNA but is associated with decreased insulin sensitivity. PMID- 15280152 TI - Difference in skeletal muscle function in males vs. females: role of estrogen receptor-beta. AB - Male skeletal muscles are generally faster and have higher maximum power output than female muscles. Conversely, during repeated contractions, female muscles are generally more fatigue resistant and recover faster. We studied the role of estrogen receptor-beta (ERbeta) in this gender difference by comparing contractile function of soleus (mainly slow-twitch) and extensor digitorum longus (fast-twitch) muscles isolated from ERbeta-deficient (ERbeta(-/-)) and wild-type mice of both sexes. Results showed generally shorter contraction and relaxation times in male compared with female muscles, and ERbeta deficiency had no effect on this. Fatigue (induced by repeated tetanic contractions) and recovery of female muscles were not affected by ERbeta deficiency. However, male ERbeta(-/-) muscles were slightly more fatigue resistant and produced higher forces during the recovery period than wild-type male muscles. In fact, female muscles and male ERbeta(-/-) muscles displayed markedly better recovery than male wild-type muscles. Gene screening of male soleus muscles showed 25 genes that were differently expressed in ERbeta(-/-) and wild-type mice. Five of these genes were selected for further analysis: muscle ankyrin repeat protein-2, muscle LIM protein, calsequestrin, parvalbumin, and aquaporin-1. Expression of these genes showed a similar general pattern: increased expression in male and decreased expression in female ERbeta(-/-) muscles. In conclusion, ERbeta deficiency results in increased performance during fatigue and recovery of male muscles, whereas female muscles are not affected. Improved contractile performance of male ERbeta(-/-) mouse muscles was associated with increased expression of mRNAs encoding important muscle proteins. PMID- 15280153 TI - Impaired plasma fatty acid oxidation in extremely obese women. AB - Skeletal muscle from extremely obese individuals exhibits decreased lipid oxidation compared with muscle from lean controls. It is unknown whether this effect is observed in vivo or whether the phenotype is preserved after massive weight loss. The objective of this study was to compare free fatty acid (FFA) oxidation during rest and exercise in female subjects who were either lean [n = 7; body mass index (BMI) = 22.6 +/- 2.2 kg/m(2)] or extremely obese (n = 10; BMI = 40.8 +/- 5.4 kg/m(2)) or postgastric bypass patients who had lost >45 kg (weight reduced) (n = 6; BMI = 33.7 +/- 9.9 kg/m(2)) with the use of tracer ([(13)C]palmitate and [(14)C]acetate) methodology and indirect calorimetry. The lean group oxidized significantly more plasma FFA, as measured by percent fatty acid uptake oxidized, than the extremely obese or weight-reduced group during rest (66.6 +/- 14.9 vs. 41.5 +/- 16.4 vs. 39.9 +/- 15.3%) and exercise (86.3 +/- 11.9 vs. 56.3 +/- 22.1 vs. 57.3 +/- 20.3%, respectively). BMI significantly correlated with percent uptake oxidized during both rest (r = -0.455) and exercise (r = -0.459). In conclusion, extremely obese women and weight-reduced women both possess inherent defects in plasma FFA oxidation, which may play a role in massive weight gain and associated comorbidities. PMID- 15280154 TI - Enhanced circadian ACTH release in obese premenopausal women: reversal by short term acipimox treatment. AB - Several studies suggest that the hypothalamo-pituitary-adrenal (HPA) axis is exceedingly active in obese individuals. Experimental studies show that circulating free fatty acids (FFAs) promote the secretory activity of the HPA axis and that human obesity is associated with high circulating FFAs. We hypothesized that HPA axis activity is enhanced and that lowering of circulating FFAs by acipimox would reduce spontaneous secretion of the HPA hormonal ensemble in obese humans. To evaluate these hypotheses, diurnal ACTH and cortisol secretion was studied in 11 obese and 9 lean premenopausal women (body mass index: obese 33.5 +/- 0.9 vs. lean 21.2 +/- 0.6 kg/m(2), P < 0.001) in the early follicular stage of their menstrual cycle. Obese women were randomly assigned to treatment with either acipimox (inhibitor of lipolysis, 250 mg orally four times daily) or placebo in a double-blind crossover design, starting one day before admission until the end of the blood-sampling period. Blood samples were taken during 24 h with a sampling interval of 10 min for assessment of plasma ACTH and cortisol concentrations. ACTH and cortisol secretion rates were estimated by multiparameter deconvolution analysis. Daily ACTH secretion was substantially higher in obese than in lean women (7,950 +/- 1,212 vs. 2,808 +/- 329 ng/24 h, P = 0.002), whereas cortisol was not altered (obese 36,362 +/- 5,639 vs. lean 37,187 +/- 4,239 nmol/24 h, P = 0.912). Acipimox significantly reduced ACTH secretion in the obese subjects (acipimox 5,850 +/- 769 ng/24 h, P = 0.039 vs. placebo), whereas cortisol release did not change (acipimox 33,542 +/- 3,436 nmol/24 h, P = 0.484 vs. placebo). In conclusion, spontaneous ACTH secretion is enhanced in obese premenopausal women, whereas cortisol production is normal. Reduction of circulating FFA concentrations by acipimox blunts ACTH release in obese women, which suggests that FFAs are involved in the pathophysiology of this neuroendocrine anomaly. PMID- 15280155 TI - Apical potassium channels in the rat connecting tubule. AB - Apical membrane K channels in the rat connecting tubule (CNT) were studied using the patch-clamp technique. Tubules were isolated from the cortical labyrinth of the kidney and split open to provide access to the apical membrane. Cell-attached patches were formed on presumed principal and/or connecting tubule cells. The major channel type observed had a single-channel conductance of 52 pS, high open probability and kinetics that were only weakly dependent on voltage. These correspond closely to the "SK"-type channels in the cortical collecting duct, identified with the ROMK (Kir1.1) gene product. A second channel type, which was less frequently observed, mediated larger currents and was strongly activated by depolarization of the apical membrane voltage. These were identified as BK or maxi-K channels. The density of active SK channels revealed a high degree of clustering. Although heterogeneity of tubules or of cell types within a tubule could not be excluded, the major factor underlying the distribution appeared to be the presence of channel clusters on the membrane of individual cells. The overall density of channels was higher than that previously found in the cortical collecting tubule (CCT). In contrast to results in the CCT, we did not detect an increase in the overall density of SK channels in the apical membrane after feeding the animals a high-K diet. However, the activity of amiloride-sensitive Na channels was undetectable under control conditions but was increased after both 1 day (90 +/- 24 pA/cell) or 7 days (385 +/- 82 pA/cell) of K loading. Thus one important factor leading to an increased K secretion in the CNT in response to increased dietary K is an increased apical Na conductance, leading to depolarization of the apical membrane voltage and an increased driving force for K movement out into the tubular lumen. PMID- 15280157 TI - Dissociation of renal TGF-beta and hypertrophy in female rats with diabetes mellitus. AB - Prepubertal onset of diabetes mellitus (DM) in male rats delays diabetic renal hypertrophy and suppresses renal transforming growth factor-beta (TGF-beta) compared with onset in adults. Because there are sex differences in normal and pathological renal growth, we performed similar experiments in female rats and examined the effects of prior ovariectomy. As in male rats, adult onset of DM increased renal weight approximately 35%, total renal TGF-beta approximately 35%, and mRNA for TGF-beta inducible gene H3 (betaIG-H3) approximately 200%. TGF-beta levels did not increase with DM in prepubertal animals, but renal weight increased approximately 40%, similar to the enlargement seen in adults. In nondiabetic rats, ovariectomy suppressed renal TGF-beta levels by 25-50% in both age groups, but betaIG-H3 was stable in younger animals and increased by approximately 200% in older animals after ovariectomy. Ovariectomy increased kidney weight approximately 10% in both age groups. DM further increased kidney weight by an additional 40% after ovariectomy with an approximately 150% increase in betaIG-H3, even though TGF-beta levels were not significantly increased. Prepubertal (approximately 99% lower), diabetic (approximately 50% lower), and ovariectomized rats (approximately 90% lower) all tended toward lower estradiol levels than intact adults, although not all differences were statistically significant. Both prepubertal onset and ovariectomy suppress TGF-beta in the kidneys of female rats with DM compared with adult-onset animals, but these states have no effect on renal enlargement. Production of the extracellular matrix component betaIG-H3 is dissociated from TGF-beta under these conditions. These observations may help explain some of the sex differences demonstrated in progressive kidney diseases, including DM. PMID- 15280156 TI - PPAR-alpha ligand ameliorates acute renal failure by reducing cisplatin-induced increased expression of renal endonuclease G. AB - Cisplatin injury to the kidney is characterized, in part, by inhibition of substrate oxidation, inflammation, and tubular cell death in the form of apoptosis and necrosis. Recently, we demonstrated that cisplatin-induced inhibition of substrate oxidation can be reversed by the administration of peroxisome proliferator-activated receptor-alpha (PPAR-alpha) ligands, resulting in amelioration of renal function. We therefore hypothesize that by improving fatty acid oxidation in vivo might protect renal function by reducing both apoptosis and necrosis in cisplatin-treated mice. Mice subjected to a single intraperitoneal injection of cisplatin developed acute renal failure (ARF) at days 3 and 4. At day 4 after cisplatin injection mRNA, protein levels and enzyme activity of proapoptotic renal endonuclease G (Endo G) were increased compared with saline-treated mice. In situ hybridization and immunohistochemical studies localized the increased expression of Endo G mRNA to the cytosolic compartment and Endo G protein to the nuclear compartment of proximal tubules in cisplatin treated mice. Pretreatment of PPAR-alpha wild-type mice with PPAR-alpha ligand WY 14643 reduced significantly cisplatin-induced increased protein expression and enzyme activity of Endo G and prevented the nuclear translocation of mitochondrial Endo G. Morphological examination of tubular injury in the PPAR alpha wild-type mice that received PPAR-alpha ligand and cisplatin did show significant amelioration of acute tubular necrosis, as well as a significant reduction in the number of apoptotic cells in the proximal tubule when compared with the cisplatin-treated group. In contrast, in PPAR-alpha-null mice treated with the ligand and cisplatin, Endo G protein expression was not reduced and this was accompanied by lack of protection of kidney function. We conclude that PPAR alpha ligand protects against cisplatin-induced renal injury via a PPAR-alpha dependent mechanism by reducing the expression and enzyme activity of proximal tubule Endo G, which results in amelioration of both proximal tubule cell apoptosis and necrosis. PMID- 15280158 TI - Differential regulation of mesangial cell mitogenesis by cAMP phosphodiesterase isozymes 3 and 4. AB - Mesangial cell (MC) mitogenesis is regulated through "negative cross talk" between cAMP-PKA and ERK signaling. Although it is widely accepted that cAMP inhibits mitogenesis through PKA-mediated phosphorylation of Raf-1, recent studies have indicated that cAMP-mediated inhibition of mitogenesis may occur independently of Raf-1 phosphorylation or without inhibiting ERK activity. We previously showed that MCs possess functionally compartmentalized intracellular pools of cAMP that are differentially regulated by cAMP phosphodiesterases (PDE); an intracellular pool directed by PDE3 but not by PDE4 suppresses mitogenesis. We therefore sought to determine whether there was a differential effect of PDE3 vs. PDE4 inhibitors on the Ras-Raf-MEK-ERK pathway in cultured MC. Although PDE3 and PDE4 inhibitors activated PKA and modestly elevated cAMP levels to a similar extent, only PDE3 inhibitors suppressed MC mitogenesis (-57%) and suppressed Raf 1 kinase and ERK activity (-33 and -68%, respectively). Both PDE3 and PDE4 inhibitors suppressed B-Raf kinase activity. PDE3 inhibitors increased phosphorylation of Raf-1 on serine 43 and serine 259 and decreased phosphorylation on serine 338; PDE4 inhibitors were without effect. Overexpression of a constitutively active MEK-1 construct reversed the antiproliferative effect of PDE3 inhibitors. PDE3 inhibitors also reduced cyclin A levels (-27%), cyclin D and cyclin E kinase activity (-30 and -50%, respectively), and induced expression of the cell cycle inhibitor p21 (+90%). We conclude that the antiproliferative effects of PDE3 inhibitors are mechanistically related to inhibition of the Ras-Raf-MEK-ERK pathway. Additional cell cycle targets of PDE3 inhibitors include cyclin A, cyclin D, cyclin E, and p21. PMID- 15280159 TI - Acute study of interaction among cadmium, calcium, and zinc transport along the rat nephron in vivo. AB - This study investigates the effect in rats of acute CdCl(2) (5 microM) intoxication on renal function and characterizes the transport of Ca(2+), Cd(2+), and Zn(2+) in the proximal tubule (PT), loop of Henle (LH), and terminal segments of the nephron (DT) using whole kidney clearance and nephron microinjection techniques. Acute Cd(2+) injection resulted in renal losses of Na(+), K(+), Ca(2+), Mg(2+), PO(4)(-2), and water, but the glomerular filtration rate remained stable. (45)Ca microinjections showed that Ca(2+) permeability in the DT was strongly inhibited by Cd(2+) (20 microM), Gd(3+) (100 microM), and La(3+) (1 mM), whereas nifedipine (20 microM) had no effect. (109)Cd and (65)Zn(2+) microinjections showed that each segment of nephron was permeable to these metals. In the PT, 95% of injected amounts of (109)Cd were taken up. (109)Cd fluxes were inhibited by Gd(3+) (90 microM), Co(2+) (100 microM), and Fe(2+) (100 microM) in all nephron segments. Bumetanide (50 microM) only inhibited (109)Cd fluxes in LH; Zn(2+) (50 and 500 microM) inhibited transport of (109)Cd in DT. In conclusion, these results indicate that 1) the renal effects of acute Cd(2+) intoxication are suggestive of proximal tubulopathy; 2) Cd(2+) inhibits Ca(2+) reabsorption possibly through the epithelial Ca(2+) channel in the DT, and this blockade could account for the hypercalciuria associated with Cd(2+) intoxication; 3) the PT is the major site of Cd(2+) reabsorption; 4) the paracellular pathway and DMT1 could be involved in Cd(2+) reabsorption along the LH; 5) DMT1 may be one of the major transporters of Cd(2+) in the DT; and 6) Zn(2+) is taken up along each part of the nephron and its transport in the terminal segments could occur via DMT1. PMID- 15280160 TI - Liver X receptor-alpha mediates cholesterol efflux in glomerular mesangial cells. AB - Lipid-mediated injury plays an important role in the pathogenesis of many renal diseases including diabetic nephropathy. Liver X receptor-alpha (LXRalpha) is an intracellular sterol sensor that regulates expression of genes controlling cholesterol absorption, excretion, catabolism, and cellular efflux. The present study was aimed at examining the role of LXRalpha in cholesterol metabolism in glomerular mesangial cells. A 1,561-bp fragment of full-length rabbit LXR cDNA was cloned. The deduced protein sequence exhibited 92.4 and 89.2% identity to human and mouse LXRalpha, respectively. Tissue distribution studies showed that rabbit LXRalpha was expressed in the liver, spleen, and kidney. In situ hybridization and RT-PCR assays further indicated that LXRalpha mRNA was widely expressed in the kidney and present in every nephron segment including the glomeruli. To determine intrarenal regulation of LXRalpha, rabbits were treated with thiazolidinedione (TZD) peroxisome proliferator-activated receptor-gamma (PPARgamma) agonists, which have been previously shown to enhance LXRalpha expression via PPARgamma and increase cholesterol efflux in macrophages. The results showed that glomerular LXRalpha expression was markedly induced by TZDs. In cultured rabbit mesangial cells, LXRalpha mRNA and protein were detected by RT PCR and immunoblotting. Treatment of mesangial cells with a specific LXRalpha agonist, TO-901317, significantly increased basal and apolipoprotein AI-mediated cholesterol efflux and markedly enhanced the promoter activity of an LXRalpha target gene, ATP-binding cassette transporter A1 (ABCA1). In conclusion, LXRalpha is expressed in renal glomeruli and functionally present in mesangial cells where its activation mediates cholesterol efflux via ABCA1. These data suggest that LXRalpha may be a potential therapeutic target for treating lipid-related renal glomerular disease. PMID- 15280161 TI - Determinants of basal nitric oxide concentration in the renal medullary microcirculation. AB - In this study, we modeled the production, transport, and consumption of nitric oxide (NO) in the renal medullary microcirculation under basal conditions. To yield agreement with reported NO concentrations of approximately 60-140 nM in medullary tissues (Zou AP and Cowley AW Jr. Hypertension 29: 194-198, 1997; Am J Physiol Regul Integr Comp Physiol 279: R769-R777, 2000) and 3 nM in plasma (Stamler JS, Jaraki O, Osborne J, Simon DI, Keaney J, Vita J, Singel D, Valeri CR, and Loscalzo J. Proc Natl Acad Sci USA 89: 7674-7677, 1992), the permeabilities of red blood cells (RBCs), vascular walls, and pericytes to NO are all predicted to lie between 0.01 and 0.1 cm/s, and the NO production rate by vasa recta endothelium is estimated to be on the order of 10(-14) mumol.mum( 2).s(-1). Our results suggest that the concentration of NO in RBCs, which is essentially controlled by the kinetics of NO scavenging by hemoglobin, is approximately 0.01 nM, that is, 10(3) times lower than that in plasma, pericytes, and interstitium. Because the basal concentration of NO in pericytes is on the order of 10 nM, it may be too low to active guanylate cyclase, i.e., to induce vasorelaxation. Our simulations also indicate that basal superoxide concentrations may be too low to affect medullary NO levels but that, under pathological conditions, superoxide may be a very significant scavenger of NO. We also found that although oxygen is a negligible NO scavenger, medullary hypoxia may significantly enhance NO concentration gradients along the corticomedullary axis. PMID- 15280162 TI - ACAT inhibition reverses LCAT deficiency and improves plasma HDL in chronic renal failure. AB - Chronic renal failure (CRF) is associated with increased risk of arteriosclerotic cardiovascular disease and profound alteration of plasma lipid profile. Uremic dyslipidemia is marked by increased plasma concentration of ApoB-containing lipoproteins and impaired high-density lipoprotein (HDL)-mediated reverse cholesterol transport. These abnormalities are, in part, due to acquired LCAT deficiency and upregulation of hepatic acyl-CoA:cholesterol acyltransferase (ACAT). ACAT catalyzes intracellular esterification of cholesterol, thereby promoting hepatic production of ApoB-containing lipoproteins and constraining HDL mediated cholesterol uptake in the peripheral tissues. In view of the above considerations, we tested the hypothesis that pharmacological inhibition of ACAT may ameliorate CRF-induced dyslipidemia. 5/6 Nephrectomized rats were treated with either ACAT inhibitor IC-976 (30 mg.kg(-1).day(-1)) or placebo for 6 wk. Sham-operated rats served as controls. Key cholesterol-regulating enzymes, plasma lipids, and creatinine clearance were measured. The untreated CRF rats exhibited increased plasma low-density lipoprotein (LDL) and very LDL (VLDL) cholesterol, unchanged plasma HDL cholesterol, elevated total cholesterol-to-HDL cholesterol ratio, reduced liver microsomal free cholesterol, and diminished creatinine clearance. This was accompanied by reduced plasma LCAT, increased hepatic ACAT-2 mRNA, ACAT-2 protein and ACAT activity, and unchanged hepatic HMG-CoA reductase and cholesterol 7alpha-hydroxylase. ACAT inhibitor raised plasma HDL cholesterol, lowered LDL and VLDL cholesterol, and normalized total cholesterol-to-HDL cholesterol ratio without changing total cholesterol concentration (hence, a shift from ApoB-containing lipoproteins to HDL). This was accompanied by normalizations of hepatic ACAT activity and plasma LCAT. In conclusion, inhibition of ACAT reversed LCAT deficiency and improved plasma HDL level in CRF rats. Future studies are needed to explore the efficacy of ACAT inhibition in humans with CRF. PMID- 15280163 TI - Heterogeneous distribution of chloride channels along the distal convoluted tubule probed by single-cell RT-PCR and patch clamp. AB - The distal convoluted tubule (DCT) is a heterogeneous segment subdivided into early (DCT1) and late (DCT2) parts, depending on the distribution of various transport systems. We do not have an exhaustive picture of the Cl(-) channels on the basolateral side: the presence of ClC-K2 channels is generally accepted, whereas that of ClC-K1 remains controversial. We used here single-cell RT-PCR and patch clamp to probe Cl(-) channel heterogeneity in microdissected mouse DCT at the molecular and functional levels. Our findings show that 63% of the DCT cells express ClC-K2 mRNA, either alone (type 1 cells: 47 and 23% in DCT1 and DCT2, respectively), or combined with ClC-K1, mostly in DCT2 (type 2 cells: 33%), but 37% of DCT1 and DCT2 cells do not express any ClC-K. Patch-clamp experiments revealed that a Cl(-) channel, with 9-pS conductance and Cl(-) > NO(3)(-) = Br(-) anion selectivity sequence, is present in the DCT1 and DCT2 basolateral membranes (87 and 71% of the patches, respectively). This dominant channel is likely to be ClC-K2 in type 1 cells. In type 2 cells, it could be ClC-K2 and/or ClC-K1 homodimers, but also ClC-K1/ClC-K2 heterodimers, or a mixture of all combinations. A second, distinct Cl(-) channel (13% of DCT1 patches, 29% of DCT2 patches) also displayed 9-pS conductance but had a completely different anion selectivity (I(-) > NO(3)(-) > Br(-) > Cl(-)), which was not compatible with that of the ClC-Ks. This indicates that a Cl(-) channel that is unlikely to belong to the ClC family may also be involved in Cl(-) absorption in the DCT2. PMID- 15280164 TI - Six month tracking of microbial growth in a metalworking fluid after system cleaning and recharging. AB - Large volumes of metalworking fluids (MWFs) are used in manufacturing industries for cooling and lubrication of metal pieces and tools during machining. MWFs accumulate microbial growth through continuous recirculation and reuse. We studied the progression of microbial contamination for 6 months after dumping, cleaning and recharging (DCR) of a large semi-synthetic MWF system managed with several biocides. Fresh, uncontaminated fluid was added to the system after extensive cleaning. The following samples were collected and analyzed: pre-DCR fluid (before system cleaning); neat fluid diluted to 6% with water; in use MWF 12 h and 1, 3 and 6 months post-DCR. Samples were analyzed for total microorganism concentrations by direct counting using fluorescence microscopy and by plate counting on various media (R2A, BHI, Middlebrooks and rose bengal under aerobic conditions). In addition, PCR was performed for the detection of mycobacteria. There was a rapid progression in the total bacterial counts as determined by fluorescence microscopy: 5.7 x 10(7) cells/ml in the pre-DCR used fluid, no measurable bacteria in the neat fluid, 6.9 x 10(6) cells/ml after 12 h and 2.2 x 10(6), 3.6 x 10(8) and 6.1 x 10(8) cells/ml after 1, 3 and 6 months, respectively. On average, only 0.2% of the direct count organisms were quantified on R2A cultures. PCR showed the presence of mycobacteria in the used MWF at 3 and 6 months. Mycobacteria were also identified from cultures on Middlebrooks and R2A. This study demonstrates that standard methods for cleaning MWF systems are inadequate since residual bacteria in the system can rapidly repopulate the newly charged MWF. PMID- 15280165 TI - Characteristics of peaks of inhalation exposure to organic solvents. AB - OBJECTIVES: To determine which exposure metrics are sufficient to characterize 'peak' inhalation exposure to organic solvents (OS) during spraying operations. METHODS: Personal exposure measurements (n=27; duration 5-159 min) were collected during application of paints, primers, resins and glues in 15 companies. A MiniRAE Photo-Ionization Detector measured OS concentrations every second. These readings were adjusted for OS composition, which was determined by charcoal tubes. A peak was defined as a period during which exposure exceeded the time weighted average (TWA) exposure. Five second and 1 and 15 min moving average times were considered in defining a peak. The number of peaks per hour, the duration of a peak, the maximum concentration within a peak, the average concentration within a peak, the ratio between maximum and average concentration within a peak and the average time between two peaks were recorded for each sample. Data were analyzed using factor analysis on 13 variables for the 27 samples: TWA concentration of the task and the six peak characteristics based on the 5 s and 1 min moving averaging time. RESULTS: There were three statistically independent sources of correlation among metrics of peak exposure, explaining 87% of the multiple correlation. The first factor reflected the intensity of peak exposure; it was also associated with the TWA. The second and third factors were related to measures of variability (in frequency and intensity) and duration of peaks, respectively. CONCLUSIONS: We present a method for describing peak profiles for inhalation exposure in terms of various distinguishable and independent parameters. Pending development of toxicologically justified peak exposure metrics, such investigations can be of value in identifying exposure metrics for which non-confounded risk estimates can be obtained in epidemiological studies. PMID- 15280166 TI - A method for assessing occupational dermal exposure to permanent hair dyes. AB - Hairdressers have an increased risk of developing occupational skin diseases due to exposure to skin irritants and sensitizers. In the present work a method of assessing dermal exposure to permanent hair dyes was developed. The sampling performance characteristics of hand wash sampling with bag rinsing were studied for five hair dye compounds. The effect of residence time, sample load and different matrices were studied. Thirty volunteers were exposed to a reference solution of these compounds and to commercial hair dye products. The sampling efficiency after 5 min residence time was between 70 and 90% for the dye components in the hair dye products. Sampling efficiency decreases with increasing residence time, making the time of sampling an important factor. Hand wash sampling should be performed as soon as possible after the work task of interest. We conclude that the sampling efficiency is adequate for measurements of dermal exposure to permanent hair dyes. Hand wash sampling with bag rinsing is a useful tool for field studies of dermal exposure assessment in hairdressers. PMID- 15280167 TI - NO and neutrophils during sepsis: NO says "yes" to sequestration but "no" to migration. PMID- 15280168 TI - Progression of asthma: small steps and a long way to go. PMID- 15280169 TI - Angiogenesis and pulmonary fibrosis: feast or famine? PMID- 15280170 TI - Serotonin in pulmonary arterial hypertension. PMID- 15280171 TI - The GOLD classification has advanced understanding of COPD. PMID- 15280172 TI - The GOLD classification has not advanced understanding of COPD. PMID- 15280175 TI - A century of asthma. PMID- 15280177 TI - Mechanisms and limits of induced postnatal lung growth. PMID- 15280178 TI - Severe asthma attacks after sexual intercourse. PMID- 15280180 TI - Familial breast cancer screening: ethical and social implications. PMID- 15280181 TI - Genetic alterations in hereditary breast cancer. AB - Genetic linkage studies have led to the identification of highly penetrant genes as the possible cause of inherited cancer risk in many cancer-prone families. Most women with a family history of breast/ovarian cancer have tumors characterized by alterations in particular genes, mainly BRCA1 and BRCA2, but also CHK2, ATM, STK11 and others. This paper examines the BRCA1 and BRCA2 genes, focusing on the Italian pattern of mutations. The function of these two genes, classified as tumor suppressors, is linked with key metabolic mechanisms such as DNA damage repair, regulation of gene expression and cell cycle control. The pathological BRCA allelic variants may cause alteration of protein function, transcriptional activity and DNA repair; accumulation of the defects leads to widespread chromosome instability that may be directly responsible for cancer formation. In fact, mutations in BRCA1 and BRCA2, conferring a highly increased susceptibility to breast and ovarian cancer, do not lead to cancer by themselves. The current consensus is that these are 'caretaker' genes, which, when inactivated, allow other genetic defects to accumulate. The nature of these other molecular events may define the pathway through which BRCA1 and BRCA2 act. The BRCA mutation spectrum is complex, and the significance of most nucleotide alterations is difficult to understand. Moreover, the mutation pattern seems to be related to ethnicity. The Italian Consortium of Hereditary Breast and Ovarian Cancer has reviewed 1758 families; 23% have been found to be carriers of pathogenetic mutations in BRCA1 or BRCA2. Founder mutations have been described in geographically restricted areas of Italy; a regional founder effect has been demonstrated in Italy for the mutations BRCA1 5083del19 and BRCA2 8765delAG, and a probable new founder mutation has been characterized in Tuscany. The presence of founder mutations has practical implications for genetic testing. PMID- 15280182 TI - External quality assessment for mutation detection in the BRCA1 and BRCA2 genes: EMQN's experience of 3 years. AB - BACKGROUND: The European Molecular Genetics Quality Network (EMQN) was formed in order to improve external quality assessment for molecular genetic testing in Europe. From 1999 to 2002 it received funding from the European Union under the Standards, Measurement and Testing programme (contract no. SMT4-CT98-7515). Since then, its maintenance has been supported through subscription of the participants, and it has been coordinated by the National Genetic Reference Laboratory at Manchester, UK (Rob Elles and Simon Patton; www.emqn.org). MATERIALS AND METHODS: Among other external quality assessment (EQA) schemes, EMQN has provided an EQA scheme for mutation detection in the breast cancer genes, BRCA1 and BRCA2, designed to cover the two important aspects of genetic testing: (i) genotyping and (ii) interpretation and reporting of results. The fourth full scheme was completed in 2003, with data evaluation pending for the 47 participants. RESULTS: Analysis of genotyping data has pinpointed two main types of errors: (i) missing a mutation (in nine of the 17 false results a normal sequence was reported); and (ii) description of the observed sequence change by an incorrect nomenclature. Compared with the more technical process of genotyping, the writing of reports displayed a much wider variation between laboratories. CONCLUSIONS: From the reported data it is clear that external quality control should become an integral part of quality assessment in the laboratory, thus contributing to maintaining confidence in the reliability of genetic testing among patients and health professionals. PMID- 15280183 TI - An overview of the status of imaging screening technology for breast cancer. AB - With breast cancer incidence rates showing no signs of abating, advances in risk stratification and increasing awareness of cancer control, there is interest in expanding the breast imaging arsenal. Mammography is still the standard of care, and a recent meta-analysis of seven large studies supports its value as a screening tool. There is, however, clear need for improved sensitivity and specificity. Imaging of function, metabolism and molecular activity in breast tissue is of potential benefit in addressing these issues. In this article we provide an overview of the current methods of imaging in breast cancer, including mammography, ultrasound, digital mammography, magnetic resonance, positron emission tomography and magnetic resonance spectroscopy. Screening and surveillance should, ideally, be tailored to an individual's cancer risk and breast tissue. Current evidence questions the recent move toward magnetic resonance imaging as a single or multimodality strategy for breast cancer screening. In a high-risk group, the cost effectiveness of technical innovations may be justified. PMID- 15280184 TI - Reducing breast cancer incidence in familial breast cancer: overlooking the present panorama. AB - Familial breast cancer, whether associated or not with particular other breast cancer features (male, early onset, bilateral breast cancer), determines a wide and variable risk of developing breast cancer in the 'unpatients' (unaffected individuals) of these families, particularly in those harboring a genetic predisposition. The antiestrogen tamoxifen has been proposed in different trials to prevent breast cancer in women at risk. The NSABP-P1 study demonstrated that tamoxifen drastically reduced (by approximately 50%) the incidence of breast cancer in women at risk selected according to the Gail score. The preventive effect was particularly consistent in postmenopausal women and in those showing familial breast cancer (three or more affected patients). BRCA1/BRCA2 (BRCA1/2) gene analysis in women accrued in the NSABP-P1 trial who developed breast cancer showed that tamoxifen chemoprevention reduced breast cancer incidence in BRCA2 carriers. Different chemoprevention trials are ongoing to compare different selective estrogen receptor modulators and aromatase inhibitors with tamoxifen. The Italian Consortium of Hereditary Breast Ovarian Cancer recently developed the Aromasin Prevention Study, a multicenter, double-blind, randomized, placebo controlled phase III study evaluating the effect of the aromatase inhibitor exemestane for chemoprevention in postmenopausal women carriers of BRCA1/2 genetic predisposition. Women who are postmenopausal unaffected carriers of BRCA1/2 mutations will be selected by participating institutions and randomly assigned to receive either oral exemestane or oral placebo every day for 3 years in order to reduce the incidence of breast cancer. Genetic counseling and the detection of predisposing BRCA1/2 mutations are mandatory before accrual into the study. Signed informed consents for the performing of BRCA1 and BRCA2 genetic analysis and for enrollment into the study are required. Eligible women will be followed thereafter in order to evaluate the efficacy of exemestane in reducing the incidental rate of breast cancer in unaffected postmenopausal carriers of BRCA1/2 mutations. PMID- 15280185 TI - Decisions and outcomes of genetic testing for inherited breast cancer risk. AB - Since the discovery of breast cancer susceptibility genes and the availability of genetic testing, a substantial amount of research has been conducted to evaluate rates of genetic test acceptance and to understand the psychological and behavioral impact of BRCA1 and BRCA2 (BRCA1/2) genetic test results. This article explores findings related to genetic test acceptance for inherited breast cancer risk and the impact of genetic test results on psychological functioning, cancer prevention and control behaviors, and family communication about genetic testing. Overall, rates of genetic test acceptance were lower than anticipated based on interest in genetic testing reported in early research. While there is limited evidence that genetic testing generates adverse psychological effects, receiving positive BRCA1/2 test results may cause emotional reactions and concerns that are specific to such results. Although early reports suggested that receiving positive BRCA1/2 test results may have a limited impact on cancer screening or prevention behaviors, recent studies have shown that genetic testing for inherited breast cancer risk may increase screening behaviors among mutation carriers. However, utilization of some screening tests remains low among mutation carriers. Additional studies are needed to identify subgroups of participants in genetic testing who may be vulnerable to experiencing testing-specific concerns, and to evaluate the effects of interventions designed to promote behavioral change and address other concerns that may be generated by receiving positive BRCA1/2 test results. PMID- 15280186 TI - Genetic testing and breast cancer: the women's point of view. AB - Breast cancer is the most common cancer among women in Western countries (130,000 cases per year in Europe) and accounts for 20-25% of all malignancies in European women. In the past few years medical journals have focused greater attention on the quality and quantity of information provided to consumers; there is a general consensus amongst physicians on the importance of having better informed consumers. This change in attitude is influencing greatly the physician-patient relationship and political decisions. Breast cancer associations, like the National Breast Cancer Coalition in the USA or EUROPA DONNA, the European Breast Cancer Coalition in Europe, have pushed for involvement in the discussion of any phase of illness, and have a particular interest in preventive medicine. The identification of high-risk women by genetic testing for BRCA1 and BRCA2 mutations is largely debated, in particular regarding patient counseling, and psychosocial and legislative support. This article reports the different points of view raised by women's movements, so that useful suggestions may be provided to improve breast cancer prevention modalities. PMID- 15280187 TI - Intensive radiologic surveillance: a focus on the psychological issues. AB - Although women who carry BRCA1 or BRCA2 mutations have up to an 85% lifetime risk of breast cancer, the majority choose to forego prophylactic mastectomy, which has been proven to markedly lower breast cancer mortality, and opt for lifelong intensive surveillance. Whether surveillance lowers breast cancer mortality in these women is unknown. However, in a formal survey of 34 of these women, 82% indicated a strong belief in the ability of surveillance to find breast cancer at a stage when it is still curable. Since 1997 we have been conducting a study to compare the sensitivity of breast magnetic resonance imaging (MRI), ultrasound, mammography and clinical breast examination (CBE) in women at high risk for hereditary breast cancer. Breast cancer incidence rates have been even higher than predicted for this population. The addition of MRI and ultrasound to conventional surveillance with mammography and CBE significantly improves sensitivity, but at the expense of decreased specificity. Two years ago we began a formal study of distress and breast cancer anxiety. A sample of 25 new and ongoing participants in the surveillance study have completed the Hospital Anxiety and Depression Scale together with the Breast Cancer Worry Scale, up to six times per year over a 2-year period. To date there has been no evidence of any impact of intensive surveillance, including false-positive studies, on anxiety, depression or breast cancer worry. PMID- 15280188 TI - Awareness of breast cancer genetics and interest in predictive genetic testing: a survey of a southern Italian population. AB - BACKGROUND: Before starting a molecular screening program for breast cancer risk and in order to develop ad hoc educational strategies, a population survey in Apulia, Italy, was performed to gather information on women's awareness of breast cancer genetics and their attitude toward genetic testing for breast cancer risk. PATIENTS AND METHODS: A consecutive series of 677 healthy women with or without a family history of breast cancer, who attended the outpatient clinics of Lega Italiana per la Lotta contro i Tumori in Bari, Italy, for preventive visits, were asked to complete a 20-item questionnaire on socio-demographics, risk perception, psychological characteristics and interest in genetic testing for breast cancer predisposing genes. RESULTS: Most women (77%) reported knowing something about the genetics of breast cancer; only 7% of the women were not interested at all in genetic testing. These figures were not significantly different for women with or without a family history of breast cancer. The two most frequently cited reasons for being interested in genetic testing, accounting for more than 50% of collected responses, were 'to learn about your children's risk' and 'to help advance research'. On multiple logistic regression analysis, only older age [odds ratio (OR) 1.9; 95% confidence interval (CI) 1.3-2.9] was associated with women's knowledge of genetic testing. Moreover, marital status (OR 4.0; 95% CI 1.1-14.6) and thinking of cancer (OR 2.2; 95% CI 1.0-4.7) independently predicted the interest in having genetic testing. CONCLUSIONS: Southern Italian women seem highly interested in genetic testing for breast cancer risk. However, their expectations mainly regard their concerns about their children or their altruistic need to help research rather than the idea of a direct clinical benefit. The great interest of the women in genetic testing probably reflects their inappropriate knowledge of the information that genetic testing can provide for breast cancer risk analysis. PMID- 15280189 TI - Costs and benefits of diagnosing familial breast cancer. AB - Costs and benefits are ill-defined terms; this review discusses some interpretations and some ways of assessing costs and benefits. Cost-benefit calculation is a misconception in health care, in that costs should be considered together with what the patients gain in terms of prolonged life and life quality. In the emerging commercialization of Western health care, however, money benefit is to be calculated in order to ask for funding. As professionals we are dealing with inherited breast cancer. We come to know the families and the individual women within these families. We care, we cure, we prevent and we are gaining knowledge. We are doing good, not harm. We are profoundly involved, and we are so deeply and personally involved that we may not even try to be objective in a debate on choosing between our patients and patients cared for by others. As we present our arguments, society has to decide. There is no argument to provide service for other inherited disorders based on ethical standards alone, and to deprive women at risk for breast cancer the life-saving health service they need because of money costs. The money needed is based on a number of rapidly fluctuating factors: the most stable costs are genetic counseling and physical examinations to demonstrate disease, while laboratory costs are rapidly decreasing. The overall conclusion is that economical cost-benefit analysis as an argument for our activity is outside our professional ethics, and should never become a reason for our continued endeavors. PMID- 15280190 TI - Genetic medicine: the balance between science and morality. AB - This article explores the relationship between science and morality with respect to the major changes that genetic knowledge has induced in medicine, as well as in many other spheres of our lives. The following themes are treated: (i) the influence of genetic knowledge on the concepts of normalcy and diversity with respect to health; (ii) the influence of genetic knowledge on the concept of responsibility; (iii) the reciprocal influence of pre-existing biases and genetic knowledge; (iv) the influence of genetic knowledge on the concept of community; and (v) the influence of genetic knowledge on autonomy and trust in the patient doctor relationship. The article does not wish to be prescriptive, but rather to raise open questions. The key philosophical question is to what extent human beings benefit from predictions about the future. The role attributed to genetics is largely overestimated. Genetic knowledge can be perceived as enhancing the control that individuals have on their lives, or as paralyzing the decision process of an individual who may feel predestined to a serious disease. In the case of breast and ovarian cancer, the probabilistic nature of genetics is especially relevant, given the relatively low penetrance of BRCA mutations. "Future things: not our domain. But in this today which unravels in front of us, what shall we do?" (Sophocles, Antigones). PMID- 15280191 TI - Ethical implications of predictive DNA testing for hereditary breast cancer. AB - Predictive medicine offers the possibility of detecting many common diseases that have a genetic basis, such as cancer; however, a genetic alteration might only indicate susceptibility to, not certainty of, disease. Whereas means for identifying a greater susceptibility to disease have been developed, effective interventions have progressed much more slowly. Awareness of one's susceptibility to disease without an actual possibility of intervention can lead to an unacceptable use of such information, or have a dramatic psychological impact on the person involved. Are the risks connected with the knowledge of susceptibility to genetic disease proportional to the benefits that such knowledge may provide? Does the knowledge of one's genetic condition constitute a service to the individual and society, or is this predominantly harmful for the person involved? The problem is vast, and involves medical, psychological, social, political and ethical dilemmas. These dilemmas, common to all predictive medicine, are most evident in predictive DNA testing for hereditary breast cancer. In our analysis, we will first examine the ethical values involved in genetic testing, highlighting the special ethical issues raised by predictive DNA testing for hereditary breast cancer. Next we will deal with genetic counseling, which, in our opinion, is the 'ethos' for ethically justifying predictive DNA testing. PMID- 15280192 TI - Surface fucosylation of human cord blood cells augments binding to P-selectin and E-selectin and enhances engraftment in bone marrow. AB - Murine hematopoietic stem and progenitor cells (HSPCs) home to bone marrow in part by rolling on P-selectin and E-selectin expressed on endothelial cells. Human adult CD34(+) cells, which are enriched in HSPCs, roll on endothelial selectins in bone marrow vessels of nonobese diabetic/severe combined immune deficiency (NOD/SCID) mice. Many human umbilical cord blood (CB) CD34(+) cells do not roll in these vessels, in part because of an uncharacterized defect in binding to P-selectin. Selectin ligands must be alpha1-3 fucosylated to form glycan determinants such as sialyl Lewis x (sLe(x)). We found that inadequate alpha1-3 fucosylation of CB CD34(+) cells, particularly CD34(+)CD38(-/low) cells that are highly enriched in HSPCs, caused them to bind poorly to E-selectin as well as to P-selectin. Treatment of CB CD34(+) cells with guanosine diphosphate (GDP) fucose and exogenous alpha1-3 fucosyltransferase VI increased cell-surface sLe(x) determinants, augmented binding to fluid-phase P- and E-selectin, and improved cell rolling on P- and E-selectin under flow. Similar treatment of CB mononuclear cells enhanced engraftment of human hematopoietic cells in bone marrows of irradiated NOD/SCID mice. These observations suggest that alpha1-3 fucosylation of CB cells might be a simple and effective method to improve hematopoietic cell homing to and engraftment in bone marrows of patients receiving CB transplants. PMID- 15280193 TI - Feasibility of HLA-haploidentical hematopoietic stem cell transplantation between noninherited maternal antigen (NIMA)-mismatched family members linked with long term fetomaternal microchimerism. AB - Based on the hypothesis that long-term fetomaternal microchimerism is associated with acquired immunologic hyporesponsiveness to noninherited maternal antigens (NIMAs) or inherited paternal antigens (IPAs), several groups have recently reported successful cases of non-T-cell-depleted hematopoietic stem cell transplantation (SCT) from HLA-haploidentical family members mismatched for NIMAs. In this study, we examined the outcomes of 35 patients with advanced hematologic malignancies who underwent HLA-2-antigen- or HLA-3-antigen incompatible SCT from a microchimeric NIMA-mismatched donor. After standard intensity or reduced-intensity preparative regimens, all patients had sustained hematopoietic recovery with tacrolimus-based graft-versus-host disease (GVHD) prophylaxis. Grade II/IV acute GVHD occurred in 19 (56%) of 34 evaluable patients, while extensive chronic GVHD developed in 13 (57%) of 23 patients who could be evaluated. Multivariate analysis demonstrated that NIMA mismatch in the GVH direction was associated with a lower risk of severe grade III-IV acute GVHD when compared with IPA mismatch (P = .03). Fifteen patients were alive and 14 of them were disease-free with a median follow-up of 20 (range, 8 to 37) months. These results indicate that T cell-replete SCT from an HLA-haploidentical NIMA mismatched donor can offer durable remission with an acceptable risk of GVHD in selected patients with advanced hematologic malignancies who lack immediate access to a conventional stem cell source. PMID- 15280194 TI - Paradoxical effects of interleukin-18 on the severity of acute graft-versus-host disease mediated by CD4+ and CD8+ T-cell subsets after experimental allogeneic bone marrow transplantation. AB - Administration of exogenous interleukin-18 (IL-18) regulates experimental acute graft-versus-host disease (GVHD) in a Fas-dependent manner when donor CD4(+) T cells are required for mortality after experimental allogeneic bone marrow transplantation (BMT). However, CD4(+) and CD8(+) T cells can induce acute GVHD after clinical allogeneic BMT, and the role of IL-18 in CD8(+)-mediated acute GVHD is unknown. We, therefore, determined the role of IL-18 in GVHD mediated by CD4(+) or CD8(+) T cells across major histocompatibility complex (MHC) class II- and class I-disparate allogeneic BMT, respectively. Administering IL-18 significantly increased survival in CD4(+)-mediated GVHD but reduced survival in CD8(+)-mediated GVHD. This increase in deaths was associated with significantly greater clinical, biochemical, and histopathologic parameters of GVHD damage and was independent of Fas expression on donor T cells. Administering IL-18 significantly enhanced allospecific cytotoxic function and expansion of CD8(+) cells. Endogenous IL-18 was critical to GVHD mediated by CD8(+) donor T cells because IL-18 receptor-deficient donors caused significantly less GVHD but exacerbated CD4(+)-mediated, GVHD-related death. Furthermore, administering anti IL-18 monoclonal antibody significantly reduced CD8(+)-mediated, GVHD-related death. Together these findings demonstrate that IL-18 has paradoxical effects on CD4(+) and CD8(+) cell-mediated GVHD. PMID- 15280195 TI - Peripheral blood stem cell transplantation in 16 patients with POEMS syndrome, and a review of the literature. AB - POEMS syndrome is characterized by peripheral neuropathy (PN), a clonal plasma cell disorder (PCD), organomegaly, endocrinopathy, skin changes, edema, sclerotic bone lesions, and thrombocytosis. Based on the improved response rates observed with peripheral blood stem cell transplantation (PBSCT) in patients with other PCDs, autologous PBSCT may be an attractive treatment option for this syndrome. Sixteen patients with POEMS syndrome have undergone PBSCT at Mayo. Of these patients, 15 had a severe rapidly progressive sensorimotor PN (9 were wheelchair dependent) and 14 were male. Median age was 51 years (range, 19-62 years). The median number of prior therapies was 3 (range, 0-7). From first symptoms and from diagnosis of POEMS the times to transplantation were 42 months and 5 months (ranges, 8-185 months and 2-149 months), respectively. There were 15 patients who had significantly abnormal pretransplant pulmonary function tests. There was one transplant-related death. During the peritransplant period, 5 patients required intubation for respiratory compromise, including one who required intubation during his stem cell mobilization period. Another patient required noninvasive biphasic positive airway pressure throughout his course. Of the 14 evaluable patients, all have had neurologic improvement or stabilization. Other features have improved substantially. PBSCT for POEMS syndrome is effective therapy but may also be associated with significant morbidity. PMID- 15280196 TI - Long-term acquisition of allergen-specific IgE and asthma following allogeneic bone marrow transplantation from allergic donors. AB - Adoptive transfer of allergen-specific immunoglobulin E (IgE) from atopic donors to nonatopic recipients occurs during the first year following bone marrow transplantation (BMT). Mature B- and T-cell clones with allergen-specific memory and hematopoietic progenitor cells are transferred through BMT. The objective of this study was to characterize the long-term rate of allergic sensitization and development of clinical allergic diseases following BMT from atopic donors. A long-term follow-up study was conducted in a cohort of donor and recipient pairs with moderate-to-severe allergic disease in the donor prior to BMT. Assessments of allergen-specific IgE, clinical rhinitis, and asthma were made in the donors prior to BMT and in the recipients with a mean follow-up of 15.5 years after BMT. From an initial cohort of 12 bone marrow transplant recipients who received marrow from allergic donors, 5 long-term survivors were identified. Allergen specific IgE transferred from donor to recipient following BMT frequently persisted, and a high rate of de novo allergic sensitization was observed between 1 and 14 years after BMT. These events were associated with elevation in total IgE, and development of allergic rhinitis and asthma at long-term follow-up. We conclude that marrow-derived immune cells from allergic donors can transfer the predisposition to allergy and asthma. PMID- 15280197 TI - Comparative analysis of T-cell costimulation and CD43 activation reveals novel signaling pathways and target genes. AB - The CD43 lymphocyte surface receptor is involved in the regulation of lymphocyte adhesion and activation. Many CD43 functions remain controversial or unclear, and it is not known to which extent CD43 signaling pathways are shared with or distinct from those used by the T-cell receptor (TCR). Here, we systematically compared signaling events and target gene expression induced by CD43 or T-cell costimulation in primary human peripheral T cells. These studies identify nuclear factor-kappaB (NF-kappaB) p65 serine 468 as a novel inducible phosphorylation site strongly induced by T-cell costimulation and only weakly triggered by CD43 ligation. We also identified CD43 as a novel Jun N-terminal kinase (JNK) activator and a comprehensive analysis of further signaling events suggests that both stimuli use overlapping but also distinct signaling pathways. Microarray analysis of inflammatory genes shows 1 group of genes coregulated by both stimuli and 2 further groups of target genes affected solely by costimulation or primarily by CD43. PMID- 15280198 TI - Mouse CD99 participates in T-cell recruitment into inflamed skin. AB - Human CD99 is a small highly O-glycosylated cell-surface protein expressed on most leukocytes. It was recently found to be expressed at endothelial cell contacts and to participate in the transendothelial migration (TEM) of monocytes in vitro. In order to analyze the physiologic relevance of CD99 in vivo we searched for the mouse homolog. We cloned a mouse cDNA coding for a protein 45% identical in its sequence with human CD99. Based on the cDNA, we generated antibodies against this mouse homolog of CD99, which detected the antigen on most leukocytes, on endothelia of various tissues, and at cell contacts of cultured endothelial cells. Cell aggregation of CD99-transfected Chinese hamster ovary (CHO) cells was completely blocked by anti-CD99 antibodies. The same antibodies inhibited TEM of lymphocytes in vitro, independent of whether T cells or endothelial cells were preincubated with antibodies. In a cutaneous delayed-type hypersensitivity (DTH) reaction, anti-CD99 antibodies inhibited the recruitment of in vivo-activated T cells into inflamed skin as well as edema formation. We conclude that mouse CD99 participates in the TEM of lymphocytes and in their recruitment to inflamed skin in vivo. This establishes CD99 as a valid target for interference with cutaneous inflammatory processes. PMID- 15280199 TI - Single-institute comparative analysis of unrelated bone marrow transplantation and cord blood transplantation for adult patients with hematologic malignancies. AB - Unrelated cord blood transplantation (CBT) has now become more common, but as yet there have been only a few reports on its outcome compared with bone marrow transplantation (BMT), especially for adults. We studied the clinical outcomes of 113 adult patients with hematologic malignancies who received unrelated BM transplants (n = 45) or unrelated CB transplants (n = 68). We analyzed the hematopoietic recovery, rates of graft-versus-host disease (GVHD), risks of transplantation-related mortality (TRM) and relapse, and disease-free survival (DFS) using Cox proportional hazards models. The time from donor search to transplantation was significantly shorter among CB transplant recipients (median, 2 months) than BM transplant recipients (median, 11 months; P < .01). Multivariate analysis demonstrated slow neutrophil (P < .01) and platelet (P < .01) recoveries in CBT patients compared with BMT patients. Despite rapid tapering of immunosuppressants after transplantation and infrequent use of steroids to treat severe acute GVHD, there were no GVHD-related deaths among CB transplant recipients compared with 10 deaths of 24 among BM transplant recipients. Unrelated CBT showed better TRM and DFS results compared with BMT (P = .02 and P < .01, respectively), despite the higher human leukocyte antigen mismatching rate and lower number of infused cells. These data strongly suggest that CBT could be safely and effectively used for adult patients with hematologic malignancies. PMID- 15280200 TI - Hematopoietic cell-derived microparticle tissue factor contributes to fibrin formation during thrombus propagation. AB - Tissue factor (TF) is expressed on nonvascular cells and cells within the vessel wall and circulates in blood associated with microparticles. Although blood-borne TF accumulates into the developing thrombus during thrombus formation, the contribution of blood-borne TF and vessel wall TF to thrombin generation in vivo following vessel injury is unknown. To determine the source and role of blood borne microparticle TF, we studied arterial thrombus formation in a living mouse using intravital microscopy. Platelet, TF, and fibrin accumulation in the developing thrombus was compared in wild-type and low TF mice. Compared to wild type mice, low TF mice formed very small platelet thrombi lacking TF or fibrin. Wild-type and low TF mice received transplants of bone marrow from wild-type and low TF mice. Arterial thrombi in low TF bone marrow/wild-type chimeric mice had decreased size and decreased TF and fibrin levels. Arterial thrombi in wild-type bone marrow/low TF chimeric mice showed decreased platelet thrombus size but normal TF and fibrin levels. This demonstrates that blood-borne TF associated with hematopoietic cell-derived microparticles contributes to thrombus propagation. PMID- 15280201 TI - Cardiomyocyte-specific Bcl-2 overexpression attenuates ischemia-reperfusion injury, immune response during acute rejection, and graft coronary artery disease. AB - After cardiac transplantation, graft damage occurs secondary to ischemia reperfusion injury and acute rejection. This damage ultimately leads to the development of graft coronary artery disease (GCAD), which limits long-term graft survival. Apoptosis is directly involved in graft injury, contributing to the development of GCAD. To assess the role of the antiapoptotic factor Bcl-2 in the process of GCAD, we transplanted hearts from FVB transgenic mice overexpressing human Bcl-2 under the control of alpha-myosin heavy chain promoter into allogenic C57BL/6 mice. Bcl-2 overexpression led to reduced cytochrome c-mediated caspase-9 dependent cardiomyocyte apoptosis and local inflammation (neutrophil infiltration and proinflammatory cytokine production) in cardiac allografts during ischemia reperfusion injury and also led to reduced immune responses (inflammatory cell infiltration, production of T(H)1 cytokines and chemokines, and expression of adhesion molecules) during acute and chronic rejection without affecting host CD4(+) and CD8(+) cell responses in the spleen. Thus, local Bcl-2 expression directly contributes to the modulation of local immune responses in allograft rejection, resulting in attenuated GCAD. In conclusion, our findings suggest that the modulation of Bcl-2 expression by pharmacologic up-regulation or gene transfer may be of clinical benefit in the short- and long-term function of cardiac allografts. PMID- 15280202 TI - Lepirudin for prophylaxis of thrombosis in patients with acute isolated heparin induced thrombocytopenia: an analysis of 3 prospective studies. AB - This analysis of 3 prospective multicenter trials in patients with laboratory confirmed acute heparin-induced thrombocytopenia (HIT) without clinically evident thromboembolic complications (TECs), isolated HIT, assessed the combined individual end points of death, new TECs, and limb amputation. Patients with the same inclusion criteria who did not receive lepirudin or danaparoid served as a contemporaneous control group. Ninety-one patients were treated with lepirudin (intravenous infusion 0.10 mg/kg/h, no bolus, activated partial thromboplastin time [aPTT]-adjusted to 1.5-2.5 times baseline) for a median of 11.0 days (range, 1-68 days). During the observation period (median 24 days), 13 (14.3%) deaths, 4 (4.4%) new TECs, 3 (3.3%) limb amputations (combined 18 [19.8%]), and 13 (14.3%) major bleeding events occurred. In comparison to the control group (N = 47), the combined end point (P = .0281) and new TECs (P = .02) were reduced, and major bleeding was not significantly different between groups (P = .5419). In renal impairment, lepirudin did not reach its steady state within 4 hours, and additional monitoring every 4 hours after start of lepirudin until steady state is reached is recommended. Lepirudin seems to be effective in patients with isolated HIT. Dose reductions in renal impairment are important. Keeping the aPTT in the range corresponding to 600 to 700 microg/L lepirudin during treatment may minimize bleeding complications. PMID- 15280203 TI - Myeloablative allogeneic hematopoietic stem cell transplantation in patients who experience relapse after autologous stem cell transplantation for lymphoma: a report of the International Bone Marrow Transplant Registry. AB - Myeloablative allogeneic hematopoietic stem cell transplantation (allo-HSCT) is increasingly used in patients with lymphoma who experience disease relapse after autologous hematopoietic stem cell transplantation (auto-HSCT) because the allograft is tumor free and may induce a graft-versus-tumor effect. We analyzed 114 patients treated with this approach from 1990 to 1999 to assess disease progression, progression-free survival (PFS), and overall survival (OS). Cumulative incidence of disease progression at 3 years was 52%, whereas treatment related mortality was 22%, lower than previously reported. Three-year probabilities of OS and PFS were 33% and 25%, respectively. With prolonged follow up, however, nearly all patients experienced disease progression, and 5-year probabilities were 24% and 5%, respectively. Complete remission at the time of allo-HSCT and use of total body irradiation (TBI) in patients with non-Hodgkin lymphoma (NHL) were associated with lower rates of disease progression and higher rates of OS. In summary, allo-HSCT is feasible for patients with lymphoma who have relapses after auto-HSCT and can result in prolonged survival for some, but it is usually not curative. Most likely to benefit are patients who have HLA matched sibling donors, are in remission, and have good performance status. PMID- 15280204 TI - Enhanced phagocytosis of ring-parasitized mutant erythrocytes: a common mechanism that may explain protection against falciparum malaria in sickle trait and beta thalassemia trait. AB - High frequency of erythrocyte (red blood cell [RBC]) genetic disorders such as sickle cell trait, thalassemia trait, homozygous hemoglobin C (Hb-C), and glucose 6-phosphate dehydrogenase (G6PD) deficiency in regions with high incidence of Plasmodium falciparum malaria and case-control studies support the protective role of those conditions. Protection has been attributed to defective parasite growth or to enhanced removal of the parasitized RBCs. We suggested enhanced phagocytosis of rings, the early intraerythrocytic form of the parasite, as an alternative explanation for protection in G6PD deficiency. We show here that P falciparum developed similarly in normal RBCs and in sickle trait, beta- and alpha-thalassemia trait, and HbH RBCs. We also show that membrane-bound hemichromes, autologous immunoglobulin G (IgG) and complement C3c fragments, aggregated band 3, and phagocytosis by human monocytes were remarkably higher in rings developing in all mutant RBCs considered except alpha-thalassemia trait. Phagocytosis of ring-parasitized mutant RBCs was predominantly complement mediated and very similar to phagocytosis of senescent or damaged normal RBCs. Trophozoite-parasitized normal and mutant RBCs were phagocytosed similarly in all conditions examined. Enhanced phagocytosis of ring-parasitized mutant RBCs may represent the common mechanism for malaria protection in nonimmune individuals affected by widespread RBC mutations, while individuals with alpha-thalassemia trait are likely protected by a different mechanism. PMID- 15280205 TI - Interleukin-15 enhances immune reconstitution after allogeneic bone marrow transplantation. AB - Interleukin-15 (IL-15) is a gamma-common cytokine that plays an important role in the development, survival, and proliferation of natural killer (NK), NK T, and CD8+ T-cells. We administered IL-15 to recipients of an allogeneic bone marrow transplantation (allo BMT) to determine its effects on immune reconstitution. Posttransplantation IL-15 administration significantly increased donor-derived CD8+ T (mostly CD122(+)CD44(+)CD8+ T-cells), NK, and NK T-cells at day +28 in young and old recipients of allo BMT. This was associated with enhanced T-cell and NK-cell function. IL-15 stimulated homeostatic proliferation of donor CD8+ T cells in recipients of carboxyfluorescein diacetate succinimidyl ester-labeled donor T-cell infusions. Posttransplantation IL-15 administration also resulted in a decrease in apoptotic CD8+ T-cells, an increase in Bcl-2-expressing CD8+ T cells, and an increase in the fraction of Ki67+ proliferative NK and CD8+ T-cells in recipients of allo BMT. IL-15 did not exacerbate graft-versus-host disease (GVHD) in recipients of T-cell-depleted BMT but could aggravate GVHD in some cases in recipients of a T-cell-repleted BMT. Finally, we found that IL-15 administration could enhance graft-versus-leukemia activity. In conclusion, IL-15 can be administered safely to recipients of a T-cell-depleted allo BMT to enhance CD8+ T, NK, and NK T-cell reconstitution. PMID- 15280206 TI - WAVE/Scars in platelets. AB - Using specific antibodies against isoforms of WAVE (WASP [Wiskott-Aldrich syndrome protein] family Verprolin-homologous protein, also called Scar), we demonstrated that human platelets express all 3 isoforms. With the use of an in vitro pull-down technique, the src homology 3 (SH3) domain of insulin receptor substrate p53 (IRSp53) precipitated WAVE2 from platelet lysates more efficiently than did profilin I. The opposite was true for WAVE1, and neither precipitated WAVE3, suggesting that WAVE isoforms have different affinities to these ligands, while the SH3 domain of abl binds to all 3 isoforms. The 3 WAVE isoforms were distributed in the actin-rich Triton X-100-insoluble pellets following platelet aggregation induced by thrombin receptor-activating peptide. We also found that all 3 WAVE isoforms are substrates for calpain in vivo and in vitro. Although portions of these 3 isoforms were commonly distributed in the actin- and actin related protein 2 and 3 (Arp2/3)-rich edge of the lamellipodia in spreading platelets, only WAVE2 remained in the cell fringe following detergent extraction or fixation of the cells. Finally, by mass spectrometry, we found that the proteins, which reportedly interact with WAVE/Scars, are present in platelets. These data suggest that the 3 WAVE isoforms exhibit common and distinct features and may potentially be involved in the regulation of actin cytoskeleton in platelets. PMID- 15280207 TI - Chick Dach1 interacts with the Smad complex and Sin3a to control AER formation and limb development along the proximodistal axis. AB - Based on recent data, a new view is emerging that vertebrate Dachshund (Dach) proteins are components of Six1/6 transcription factor-dependent signaling cascades. Although Drosophila data strongly suggest a tight link between Dpp signaling and the Dachshund gene, a functional relationship between vertebrate Dach and BMP signaling remains undemonstrated. We report that chick Dach1 interacts with the Smad complex and the corepressor mouse Sin3a, thereby acting as a repressor of BMP-mediated transcriptional control. In the limb, this antagonistic action regulates the formation of the apical ectodermal ridge (AER) in both the mesenchyme and the AER itself, and also controls pattern formation along the proximodistal axis of the limb. Our data introduce a new paradigm of BMP antagonism during limb development mediated by Dach1, which is now proven to function in different signaling cascades with distinct interacting partners. PMID- 15280208 TI - Real-time lineage analysis reveals oriented cell divisions associated with morphogenesis at the shoot apex of Arabidopsis thaliana. AB - Precise knowledge of spatial and temporal patterns of cell division, including number and orientation of divisions, and knowledge of cell expansion, is central to understanding morphogenesis. Our current knowledge of cell division patterns during plant and animal morphogenesis is largely deduced from analysis of clonal shapes and sizes. But such an analysis can reveal only the number, not the orientation or exact rate, of cell divisions. In this study, we have analyzed growth in real time by monitoring individual cell divisions in the shoot apical meristems (SAMs) of Arabidopsis thaliana. The live imaging technique has led to the development of a spatial and temporal map of cell division patterns. We have integrated cell behavior over time to visualize growth. Our analysis reveals temporal variation in mitotic activity and the cell division is coordinated across clonally distinct layers of cells. Temporal variation in mitotic activity is not correlated to the estimated plastochron length and diurnal rhythms. Cell division rates vary across the SAM surface. Cells in the peripheral zone (PZ) divide at a faster rate than in the central zone (CZ). Cell division rates in the CZ are relatively heterogeneous when compared with PZ cells. We have analyzed the cell behavior associated with flower primordium development starting from a stage at which the future flower comprises four cells in the L1 epidermal layer. Primordium development is a sequential process linked to distinct cellular behavior. Oriented cell divisions, in primordial progenitors and in cells located proximal to them, are associated with initial primordial outgrowth. The oriented cell divisions are followed by a rapid burst of cell expansion and cell division, which transforms a flower primordium into a three-dimensional flower bud. Distinct lack of cell expansion is seen in a narrow band of cells, which forms the boundary region between developing flower bud and the SAM. We discuss these results in the context of SAM morphogenesis. PMID- 15280209 TI - Cell interactions that affect axonogenesis in the leech Theromyzon rude. AB - The leech nervous system comprises a relatively simple network of longitudinal (connective) and transverse (segmental) nerves. We have followed the normal pattern of axon development in the glossiphoniid leech Theromyzon rude by immunostaining embryonic preparations with antibody to acetylated alpha-tubulin. The dependence of the normal pattern of axon growth on cells in the mesodermal (M) and ectodermal (N, O, P and Q) lineages was examined by selectively ablating subsets of these lineages in developing embryos. We found that ablating mesoderm severely disrupted overall axonogenesis, while various ectodermal ablations induced a range of more specific phenotypes. In particular, formation of the posterior segmental nerve (PP) was abnormal in embryos deficient in primary neuroectoderm (N lineage). More specific ablations demonstrated that a subset of N-derived cells were required for establishing the PP nerve root. Previous studies have shown that the PP nerve root is normally pioneered by an O lineage derived neuron (PD). Our results suggest that the role of the N lineage-derived cells is to induce the migration of neuron P(D) to its normal position in the posterior compartment of the hemiganglion. PMID- 15280210 TI - Interactions between ID and OLIG proteins mediate the inhibitory effects of BMP4 on oligodendroglial differentiation. AB - Bone morphogenetic protein (BMP) signaling inhibits the generation of oligodendroglia and enhances generation of astrocytes by neural progenitor cells both in vitro and in vivo. This study examined the mechanisms underlying the effects of BMP signaling on glial lineage commitment. Treatment of cultured neural progenitor cells with BMP4 induced expression of all four members of the inhibitor of differentiation (ID) family of helix-loop-helix transcriptional inhibitors and blocked oligodendrocyte (OL) lineage commitment. Overexpression of Id4 or Id2 but not Id1 or Id3 in cultured progenitor cells reproduced both the inhibitory effects of BMP4 treatment on OL lineage commitment and the stimulatory effects on astrogliogenesis. Conversely, decreasing the levels of Id4 mRNA by RNA interference enhanced OL differentiation and inhibited the effects of BMP4 on glial lineage commitment. This suggests that induction of Id4 expression mediates effects of BMP signaling. Bacterial two-hybrid and co-immunoprecipitation studies demonstrated that ID4, and to a lesser extent ID2, complexed with the basic-helix loop-helix transcription (bHLH) factors OLIG1 and OLIG2, which are required for the generation of OLs. By contrast, ID1 and ID3 did not complex with the OLIG proteins. In addition, the OLIG and ID proteins both interacted with the E2A proteins E12 and E47. Further, exposure of cultured progenitor cells to BMP4 changed the intracellular localization of OLIG1 and OLIG2 from a predominantly nuclear to a predominantly cytoplasmic localization. These observations suggest that the induction of ID4 and ID2, and their sequestration of both OLIG proteins and E2A proteins mediate the inhibitory effects of BMP signaling on OL lineage commitment and contribute to the generation of astrocytes. PMID- 15280211 TI - Notch inhibits Ptf1 function and acinar cell differentiation in developing mouse and zebrafish pancreas. AB - Notch signaling regulates cell fate decisions in a variety of adult and embryonic tissues, and represents a characteristic feature of exocrine pancreatic cancer. In developing mouse pancreas, targeted inactivation of Notch pathway components has defined a role for Notch in regulating early endocrine differentiation, but has been less informative with respect to a possible role for Notch in regulating subsequent exocrine differentiation events. Here, we show that activated Notch and Notch target genes actively repress completion of an acinar cell differentiation program in developing mouse and zebrafish pancreas. In developing mouse pancreas, the Notch target gene Hes1 is co-expressed with Ptf1-P48 in exocrine precursor cells, but not in differentiated amylase-positive acinar cells. Using lentiviral delivery systems to induce ectopic Notch pathway activation in explant cultures of E10.5 mouse dorsal pancreatic buds, we found that both Hes1 and Notch1-IC repress acinar cell differentiation, but not Ptf1 P48 expression, in a cell-autonomous manner. Ectopic Notch activation also delays acinar cell differentiation in developing zebrafish pancreas. Further evidence of a role for endogenous Notch in regulating exocrine pancreatic differentiation was provided by examination of zebrafish embryos with homozygous mindbomb mutations, in which Notch signaling is disrupted. mindbomb-deficient embryos display accelerated differentiation of exocrine pancreas relative to wild-type clutchmate controls. A similar phenotype was induced by expression of a dominant-negative Suppressor of Hairless [Su(H)] construct, confirming that Notch actively represses acinar cell differentiation during zebrafish pancreatic development. Using transient transfection assays involving a Ptf1-responsive reporter gene, we further demonstrate that Notch and Notch/Su(H) target genes directly inhibit Ptf1 activity, independent of changes in expression of Ptf1 component proteins. These results define a normal inhibitory role for Notch in the regulation of exocrine pancreatic differentiation. PMID- 15280212 TI - Bmp7 regulates branching morphogenesis of the lacrimal gland by promoting mesenchymal proliferation and condensation. AB - The lacrimal gland provides an excellent model with which to study the epithelial mesenchymal interactions that are crucial to the process of branching morphogenesis. In the current study, we show that bone morphogenetic protein 7 (Bmp7) is expressed with a complex pattern in the developing gland and has an important role in regulating branching. In loss-of-function analyses, we find that Bmp7-null mice have distinctive reductions in lacrimal gland branch number, and that inhibition of Bmp activity in gland explant cultures has a very similar consequence. Consistent with this, exposure of whole-gland explants to recombinant Bmp7 results in increased branch number. In determining which cells of the gland respond directly to Bmp7, we have tested isolated mesenchyme and epithelium. We find that, as expected, Bmp4 can suppress bud extension in isolated epithelium stimulated by Fgf10, but interestingly, Bmp7 has no discernible effect. Bmp7 does, however, stimulate a distinct response in mesenchymal cells. This manifests as a promotion of cell division and formation of aggregates, and upregulation of cadherin adhesion molecules, the junctional protein connexin 43 and of alpha-smooth muscle actin. These data suggest that in this branching system, mesenchyme is the primary target of Bmp7 and that formation of mesenchymal condensations characteristic of signaling centers may be enhanced by Bmp7. Based on the activity of Bmp7 in promoting branching, we also propose a model suggesting that a discrete region of Bmp7-expressing head mesenchyme may be crucial in determining the location of the exorbital lobe of the gland. PMID- 15280213 TI - Interaction with eIF5B is essential for Vasa function during development. AB - The DEAD-box RNA helicase Vasa (Vas) is required for germ cell development and function, as well as for embryonic somatic posterior patterning. Vas interacts with the general translation initiation factor eIF5B (cIF2, also known as dIF2), and thus may regulate translation of specific mRNAs. In order to investigate which functions of Vas are related to translational control, we have analyzed the effects of site-directed vas mutations that reduce or eliminate interaction with eIF5B. Reduction in Vas-eIF5B interaction during oogenesis leads to female sterility, with phenotypes similar to a vas null mutation. Accumulation of Gurken (Grk) protein is greatly reduced when Vas-eIF5B interaction is reduced, suggesting that this interaction is crucial for translational regulation of grk. In addition, we show that reduction in Vas-eIF5B interaction virtually abolishes germ cell formation in embryos, while producing a less severe effect on somatic posterior patterning. We conclude that interaction with the general translation factor eIF5B is essential for Vas function during development. PMID- 15280214 TI - Differential cytoplasmic mRNA localisation adjusts pair-rule transcription factor activity to cytoarchitecture in dipteran evolution. AB - Establishment of segmental pattern in the Drosophila syncytial blastoderm embryo depends on pair-rule transcriptional regulators. mRNA transcripts of pair-rule genes localise to the apical cytoplasm of the blastoderm via a selective dynein based transport system and signals within their 3'-untranslated regions. However, the functional and evolutionary significance of this process remains unknown. We have analysed subcellular localisation of mRNAs from multiple dipteran species both in situ and by injection into Drosophila embryos. We find that although localisation of wingless transcripts is conserved in Diptera, localisation of even-skipped and hairy pair-rule transcripts is evolutionarily labile and correlates with taxon-specific changes in positioning of nuclei. We show in Drosophila that localised pair-rule transcripts target their proteins in close proximity to the nuclei and increase the reliability of the segmentation process by augmenting gene activity. Our data suggest that mRNA localisation signals in pair-rule transcripts affect nuclear protein uptake and thereby adjust gene activity to a variety of dipteran blastoderm cytoarchitectures. PMID- 15280215 TI - The development of semicircular canals in the inner ear: role of FGFs in sensory cristae. AB - In the vertebrate inner ear, the ability to detect angular head movements lies in the three semicircular canals and their sensory tissues, the cristae. The molecular mechanisms underlying the formation of the three canals are largely unknown. Malformations of this vestibular apparatus found in zebrafish and mice usually involve both canals and cristae. Although there are examples of mutants with only defective canals, few mutants have normal canals without some prior sensory tissue specification, suggesting that the sensory tissues, cristae, might induce the formation of their non-sensory components, the semicircular canals. We fate-mapped the vertical canal pouch in chicken that gives rise to the anterior and posterior canals, using a fluorescent, lipophilic dye (DiI), and identified a canal genesis zone adjacent to each prospective crista that corresponds to the Bone morphogenetic protein 2 (Bmp2)-positive domain in the canal pouch. Using retroviruses or beads to increase Fibroblast Growth Factors (FGFs) for gain-of function and beads soaked with the FGF inhibitor SU5402 for loss-of-function experiments, we show that FGFs in the crista promote canal development by upregulating Bmp2. We postulate that FGFs in the cristae induce a canal genesis zone by inducing/upregulating Bmp2 expression. Ectopic FGF treatments convert some of the cells in the canal pouch from the prospective common crus to a canal like fate. Thus, we provide the first molecular evidence whereby sensory organs direct the development of the associated non-sensory components, the semicircular canals, in vertebrate inner ears. PMID- 15280216 TI - Compromised generation of GABAergic interneurons in the brains of Vax1-/- mice. AB - The subcortical telencephalon is the major source of GABAergic interneurons that, during development, tangentially migrate to the cerebral cortex, where they modulate the glutamatergic excitatory action of pyramidal cells. The transcription factor Vax1, an intracellular mediator of both Shh and Fgf signaling, is expressed at high levels in the medial and lateral ganglionic eminences (MGE and LGE, respectively), in the septal area (SA), in the anterior entopeduncular area (AEP) and in the preoptic area (POA). We show that Vax1 expression in the neuroepithelium is graded: low in the ventricular zone (VZ) and high in the subventricular zone (SVZ), in a pattern that closely reproduces that of several members of the Dlx and Gsh family of homeobox transcription factors. We provide evidence that Vax1 plays an important role in proliferation and differentiation of MGE, POA/AEP and septum, and that the last structure is completely absent in Vax1-/- mice. We show that the absence of Vax1 causes a severe depletion of GABAergic neurons in the neocortex, ranging from 30% to 44%, depending on the cortical areas considered. Taken together, our data indicate that a loss of function mutation in the Vax1 gene generates abnormalities in basal ganglia subventricular zone development and that it prevents the formation of the septum, impairing GABAergic interneuron generation. PMID- 15280217 TI - The co-repressor hairless has a role in epithelial cell differentiation in the skin. AB - Although mutations in the mammalian hairless (Hr) gene result in congenital hair loss disorders in both mice and humans, the precise role of Hr in skin biology remains unknown. We have shown that the protein encoded by Hr (HR) functions as a nuclear receptor co-repressor. To address the role of HR in vivo, we generated a loss-of-function (Hr-/-) mouse model. The Hr-/- phenotype includes both hair loss and severe wrinkling of the skin. Wrinkling is correlated with increased cell proliferation in the epidermis and the presence of dermal cysts. In addition, a normally undifferentiated region, the infundibulum, is transformed into a morphologically distinct structure (utricle) that maintains epidermal function. Analysis of gene expression revealed upregulation of keratinocyte terminal differentiation markers and a novel caspase in Hr-/- skin, substantiating HR action as a co-repressor in vivo. Differences in gene expression occur prior to morphological changes in vivo, as well as in cultured keratinocytes, indicating that aberrant transcriptional regulation contributes to the Hr-/- phenotype. The properties of the cell types present in Hr-/- skin suggest that the normal balance of cell proliferation and differentiation is disrupted, supporting a model in which HR regulates the timing of epithelial cell differentiation in both the epidermis and hair follicle. PMID- 15280218 TI - Estradiol 3-glucuronide is transported by the multidrug resistance-associated protein 2 but does not activate the allosteric site bound by estradiol 17 glucuronide. AB - beta-estradiol 17-(beta-D-glucuronide) (E217G) is a well known cholestatic agent and substrate of multidrug resistance-associated protein 2 (Mrp2), whereas beta estradiol 3-(beta-D-glucuronide) (E23G) is a noncholestatic regioisomer of E217G with unknown transport properties. The purpose of this study was to compare and contrast the Mrp2-mediated transport of E217G and E23G. The full coding region of rat Mrp2 was cloned into the baculovirus genome, the recombinant baculovirus used to infect Sf9 cells, and ATP-dependent transport of 3H-E23G and 3H-E217G in Sf9 cell membranes was characterized. Mrp2 transported E23G into an osmotically sensitive space, requiring ATP, with S50=55.7 microM, Vmax=326 pmol.mg(-1).min( 1), and a Hill coefficient of 0.88. ATP-dependent Mrp2-mediated E217G transport was markedly stimulated at high E217G concentrations, consistent with positive cooperativity (Hill coefficient 1.5). E217G (5-125 microM) increased S50 but not Vmax for E23G transport, consistent with competitive inhibition. E23G (0.4-400 microM) completely, potently (IC50=14.2 microM), and competitively inhibited E217G transport, but E217G (0.01-250 microM) inhibited only 53% of E23G transport (IC50=33.4 microM). Estriol 16alpha-(beta-D-glucuronide) potently and completely inhibited transport of E23G (IC50=2.23 microM), as did beta-estradiol 3-sulfate 17-(beta-D-glucuronide) (5-50 microM). In summary, E217G binds not only to an Mrp2 transport site, but also to an allosteric site that activates Mrp2 with positive cooperativity, thus activating its own transport and potentially that of other Mrp2 substrates, such as E23G. The noncholestatic E23G is an Mrp2 substrate and competes with E217G for transport, but does not activate the allosteric site. PMID- 15280219 TI - Hepatic transport of PKI166, an epidermal growth factor receptor kinase inhibitor of the pyrrolo-pyrimidine class, and its main metabolite, ACU154. AB - PKI166, a specific inhibitor of the tyrosine kinase activity of two epidermal growth factor receptors, was under development for the treatment of cancer. In preclinical studies PKI166 was mainly cleared by metabolism, and its metabolites were eliminated by biliary excretion, emphasizing the role of liver transport processes for its disposition. Here the transport properties of [14C]PKI166 and its main metabolite [14C]ACU154, an O-glucuronide, were analyzed using 1) Madin Darby canine kidney II (MDCKII) cells stably transfected with human multidrug resistance-associated protein 2 (MRP2) and/or human organic anion-transporting peptide 2 (OATP2) and 2) liver canalicular membrane vesicles (CMVs) prepared from Wistar and mrp2-deficient TR- rats. Analysis of transport through MDCKII cells revealed that [14C]ACU154 was a substrate of MRP2 and OATP2. Rat mrp2 was shown to transport [14C]ACU154 with a Km of approximately 1 microM. [14C]PKI166 efficiently crossed MDCKII cells, particularly toward the apical side, but expression of MRP2 and/or OATP2 did not increase the flux. The effect of PKI166 and ACU154 on transport of [3H]estradiol-17beta-d-glucuronide (EG; via mrp2/MRP2 and OATP2) or [3H]taurocholic acid (TCA; via bile salt export pump (bsep) was analyzed. PKI166 inhibited the transport of [3H]EG by OATP2. ACU154 did strongly inhibit [3H]TCA uptake into CMVs from Wistar but not from TR- rats, demonstrating a dependence of bsep inhibition on mrp2 activity. ATP-dependent uptake of [3H]EG into CMVs from Wistar rats was inhibited by ACU154 but up to 4-fold increased by PKI166. In conclusion, OATP2 and MRP2/mrp2 were identified as transporters involved in ACU154 transport into bile. Both PKI166 and its O-glucuronide ACU154 affected mrp2/MRP2-, OATP2-, and/or bsep-mediated transport processes. PMID- 15280220 TI - A crucial role of uridine/cytidine kinase 2 in antitumor activity of 3'-ethynyl nucleosides. AB - The antitumor 3'-ethynyl nucleosides, 1-(3-C-ethynyl-beta-D ribopentofuranosyl)cytosine (ECyd) and 1-(3-C-ethynyl-beta-D ribopentofuranosyl)uridine (EUrd), are potent inhibitors of RNA polymerases and show excellent antitumor activity against various human solid tumors in xenograft models. ECyd is being investigated in phase I clinical trials as a novel anticancer drug possessing a unique antitumor action. ECyd and EUrd require the activity of uridine/cytidine kinase (UCK) to produce the corresponding active metabolite. The UCK family consists of two members, UCK1 and UCK2, and both UCKs are expressed in many tumor cells. It was unclear, however, whether UCK1 or UCK2 is responsible for the phosphorylation of the 3'-ethynyl nucleosides. We therefore established cell lines that are highly resistant to the 3'-ethynyl nucleosides from human fibrosarcoma HT-1080 and gastric carcinoma NUGC-3. All the resistant cell lines showed a high cross-resistance to ECyd and EUrd. As a result of cDNA sequence analysis, we found that UCK2 mRNA expressed in EUrd-resistant HT 1080 cells has a 98-base pair deletion of exon 5, whereas EUrd-resistant NUGC-3 cells were harboring the point mutation at nucleotide position 484 (C to T) within exon 4 of UCK2 mRNA. This mutation was confirmed by genome sequence analysis of the UCK2 gene. Moreover, the expression of UCK2 protein was decreased in these resistant cells. In contrast, no mutation in the mRNA or differences in protein expression levels of UCK1 were shown in the EUrd-resistant HT-1080 and NUGC-3 cells. These results suggest that UCK2 is responsible for the phosphorylation and activation of the antitumor 3'-ethynyl nucleosides. PMID- 15280221 TI - Ernst Mayr: Genetics and speciation. PMID- 15280223 TI - The molecular population genetics of HIV-1 group O. AB - HIV-1 group O originated through cross-species transmission of SIV from chimpanzees to humans and has established a relatively low prevalence in Central Africa. Here, we infer the population genetics and epidemic history of HIV-1 group O from viral gene sequence data and evaluate the effect of variable evolutionary rates and recombination on our estimates. First, model selection tools were used to specify suitable evolutionary and coalescent models for HIV group O. Second, divergence times and population genetic parameters were estimated in a Bayesian framework using Markov chain Monte Carlo sampling, under both strict and relaxed molecular clock methods. Our results date the origin of the group O radiation to around 1920 (1890-1940), a time frame similar to that estimated for HIV-1 group M. However, group O infections, which remain almost wholly restricted to Cameroon, show a slower rate of exponential growth during the twentieth century, explaining their lower current prevalence. To explore the effect of recombination, the Bayesian framework is extended to incorporate multiple unlinked loci. Although recombination can bias estimates of the time to the most recent common ancestor, this effect does not appear to be important for HIV-1 group O. In addition, we show that evolutionary rate estimates for different HIV genes accurately reflect differential selective constraints along the HIV genome. PMID- 15280222 TI - Mapping sites of positive selection and amino acid diversification in the HIV genome: an alternative approach to vaccine design? AB - A safe and effective HIV-1 vaccine is urgently needed to control the worldwide AIDS epidemic. Traditional methods of vaccine development have been frustratingly slow, and it is becoming increasingly apparent that radical new approaches may be required. Computational and mathematical approaches, combined with evolutionary reasoning, may provide new insights for the design of an efficacious AIDS vaccine. Here, we used codon-based substitution models and maximum-likelihood (ML) methods to identify positively selected sites that are likely to be involved in the immune control of HIV-1. Analysis of subtypes B and C revealed widespread adaptive evolution. Positively selected amino acids were detected in all nine HIV 1 proteins, including Env. Of particular interest was the high level of positive selection within the C-terminal regions of the immediate-early regulatory proteins, Tat and Rev. Many of the amino acid replacements were associated with the emergence of novel (or alternative) myristylation and casein kinase II (CKII) phosphorylation sites. The impact of these changes on the conformation and antigenicity of Tat and Rev remains to be established. In rhesus macaques, a single CTL-associated amino substitution in Tat has been linked to escape from acute SIV infection. Understanding the relationship between host-driven positive selection and antigenic variation may lead to the development of novel vaccine strategies that preempt the escape process. PMID- 15280224 TI - Insertions of mini-Tn10 transposon T-POP in Salmonella enterica sv. typhi. AB - We have mutagenized a clinical strain of Salmonella enterica sv. typhi with mini transposon Tn10dTet (T-POP) to obtain conditional lethal (tetracycline-dependent) mutants with T-POP insertions upstream of essential genes. Generalized transducing phage P22 was used to introduce T-POP from a S. typhimurium donor into a S. typhi recipient. Chromosomal DNA was purified from the mutagenized donor strains, fragmented, and then electroporated into S. typhi to backcross the original T-POP insertions. Four tetracycline-dependent mutants with two distinct terminal phenotypes were found among 1700 mutants with T-POP insertions. When grown in the absence of tetracycline, two of the four tetracycline-dependent mutants arrest at a late stage in the cell cycle, can be rescued by outgrowth in media with tetracycline, and define a reversible checkpoint late in the cell cycle. One of these insertions creates an operon fusion with a gene, yqgF, that is conserved among gram-negative bacteria and likely encodes an essential Holliday junction resolvase. T-POP insertions can be used not only to identify essential S. typhi genes but also to reveal novel phenotypes resulting from the depletion of their products. PMID- 15280225 TI - Changes in the localization of the Saccharomyces cerevisiae anaphase-promoting complex upon microtubule depolymerization and spindle checkpoint activation. AB - The anaphase-promoting complex/cyclosome (APC/C) is an E3 ubiquitin ligase in the ubiquitin-mediated proteolysis pathway (UMP). To understand how the APC/C was targeted to its substrates, we performed a detailed analysis of one of the APC/C components, Cdc23p. In live cells, Cdc23-GFP localized to punctate nuclear spots surrounded by homogenous nuclear signal throughout the cell cycle. These punctate spots colocalized with two outer kinetochore proteins, Slk19p and Okp1p, but not with the spindle pole body protein, Spc42p. In late anaphase, the Cdc23-GFP was also visualized along the length of the mitotic spindle. We hypothesized that spindle checkpoint activation may affect the APC/C nuclear spot localization. Localization of Cdc23-GFP was disrupted upon nocodazole treatment in the kinetochore mutant okp1-5 and in the cdc20-1 mutant. Cdc23-GFP nuclear spot localization was not affected in the ndc10-1 mutant, which is defective in spindle checkpoint function. Additional studies using a mad2Delta strain revealed a microtubule dependency of Cdc23-GFP spot localization, whether or not the checkpoint response was activated. On the basis of these data, we conclude that Cdc23p localization was dependent on microtubules and was affected by specific types of kinetochore disruption. PMID- 15280226 TI - Functional dissection of the gamma-tubulin complex by suppressor analysis of gtb1 and alp4 mutations in Schizosaccharomyces pombe. AB - In fission yeast, gamma-tubulin (encoded by the gtb1+ gene), Alp4 (Spc97/GCP2), and Alp6 (Spc98/GCP3) are essential components of the gamma-tubulin complex. We isolated gtb1 mutants as allele-specific suppressors of temperature-sensitive alp4 mutations. Mutation sites in gtb1 mutants and in several alp4 alleles were determined. The majority of substituted amino acids were mapped to a small area on the predicted surface of the gamma-tubulin molecule that might directly interact with the Alp4 protein. The cold sensitivity of gamma-tubulin mutants was almost completely suppressed by an alpha-tubulin mutation and partially suppressed by a low concentration of thiabendazole, a microtubule assembly inhibitor. Other gtb1 mutants had increased resistance to this drug. Gel filtration and immunoprecipitation analyses suggested that the mutant gamma tubulin formed an altered gamma-tubulin complex with increased stability compared to wild-type gamma-tubulin. In most gtb1 mutants, sexual development was impaired, and aberrant asci that contained an irregular spore shape and number were produced. In contrast, spore formation was not appreciably damaged in some alp4 and alp6 mutants, even at temperatures where vegetative proliferation was substantially defective. These results suggested that the function of the gamma tubulin complex or the requirement of each component of the complex is differentially regulated between the vegetative and sexual phases of the life cycle in fission yeast. In addition, genetic data indicated intimate functional connections of gamma-tubulin with several kinesin-like proteins. PMID- 15280227 TI - Defects arising from whole-genome duplications in Saccharomyces cerevisiae. AB - Comparisons among closely related species have led to the proposal that the duplications found in many extant genomes are the remnants of an ancient polyploidization event, rather than a result of successive duplications of individual chromosomal segments. If this interpretation is correct, it would support Ohno's proposal that polyploidization drives evolution by generating the genetic material necessary for the creation of new genes. Paradoxically, analysis of contemporary polyploids suggests that increased ploidy is an inherently unstable state. To shed light on this apparent contradiction and to determine the effects of nascent duplications of the entire genome, we generated isogenic polyploid strains of the budding yeast Saccharomyces cerevisiae. Our data show that an increase in ploidy results in a marked decrease in a cell's ability to survive during stationary phase in growth medium. Tetraploid cells die rapidly, whereas isogenic haploids remain viable for weeks. Unlike haploid cells, which arrest growth as unbudded cells, tetraploid cells continue to bud and form mitotic spindles in stationary phase. The stationary-phase death of tetraploids can be prevented by mutations or conditions that result in growth arrest. These data show that whole-genome duplications are accompanied by defects that affect viability and subsequent survival of the new organism. PMID- 15280228 TI - Redundant roles for histone H3 N-terminal lysine residues in subtelomeric gene repression in Saccharomyces cerevisiae. AB - The transcription of genes located in subtelomeric regions of yeast chromosomes is repressed relative to the rest of the genome. This repression requires wild type nucleosome levels but not the telomere silencing factors Sir2, Sir3, Sir4, and Rap1. Subtelomeric heterochromatin is characterized by the absence of acetylation or methylation of histone H3 lysine residues, but it is not known whether histone H3 hypoacetylation or hypomethylation is a prerequisite for the establishment of subtelomeric heterochromatin. We have systematically mutated the N-terminal tails of histone H3 and H4 in Saccharomyces cerevisiae and characterized the effects each mutant has on genome-wide expression. Our results show that subtelomeric transcriptional repression is dependent on the histone H3 N-terminal domain, but not the histone H4 N-terminal domain. Mutating lysine-4, lysine-9, lysine-14, lysine-18, lysine-23, and lysine-27 to glycine in histone H3 is also sufficient to significantly reduce subtelomeric gene repression. Individual histone H3 lysine mutations, however, have little effect on subtelomeric gene repression or genome-wide expression, indicating that these six lysine residues have redundant functions. We propose that acetylation and methylation of histone H3 N-terminal lysine residues act as redundant mechanisms to demarcate regions of euchromatin from heterochromatin. PMID- 15280229 TI - A role for DNA polymerase delta in gene conversion and crossing over during meiosis in Saccharomyces cerevisiae. AB - A screen for mutants of budding yeast defective in meiotic gene conversion identified a novel allele of the POL3 gene. POL3 encodes the catalytic subunit of DNA polymerase delta, an essential DNA polymerase involved in genomic DNA replication. The new allele, pol3-ct, specifies a protein missing the last four amino acids. pol3-ct shows little or no defect in DNA replication, but displays a reduction in the length of meiotic gene conversion tracts and a decrease in crossing over. We propose a model in which DNA synthesis determines the length of strand exchange intermediates and influences their resolution toward crossing over. PMID- 15280230 TI - Alleles of the hotspot cog are codominant in effect on recombination in the his-3 region of Neurospora. AB - There are two naturally occurring functional alleles of the recombination hotspot cog, which is located 3.5 kb from the his-3 locus of Neurospora crassa. The presence of the cog+ allele in a cross significantly increases recombination in the his-3 region compared to a cross homozygous for the cog allele. Data obtained shortly after discovery of cog+ suggested that it was fully dominant to cog. However, a dominant cog+ conflicts with observations of hotspots in Saccharomyces cerevisiae and Schizosaccharomyces pombe, in which recombination is initiated independently of homolog interactions, and suggests recombination mechanisms may differ in Neurospora and yeast. We present evidence that cog alleles are codominant in effect on both allelic recombination in his-3 and crossing over between loci flanking his-3. In addition, we show that genetic background variation has at least a twofold effect on allelic recombination. We speculate that variation in genetic background, together with the complexities of recombination in crosses bearing close mutant alleles, accounts for the previous conclusion that cog+ is dominant to cog. PMID- 15280231 TI - Genetic transformation of Neurospora tetrasperma, demonstration of repeat-induced point mutation (RIP) in self-crosses and a screen for recessive RIP-defective mutants. AB - The pseudohomothallic fungus Neurospora tetrasperma is naturally resistant to the antibiotic hygromycin. We discovered that mutation of its erg-3 (sterol C-14 reductase) gene confers a hygromycin-sensitive phenotype that can be used to select transformants on hygromycin medium by complementation with the N. crassa erg-3+ and bacterial hph genes. Cotransformation of hph with PCR-amplified DNA of other genes enabled us to construct strains duplicated for the amplified DNA. Using transformation we constructed self-fertile strains that were homoallelic for an ectopic erg-3+ transgene and a mutant erg-3 allele at the endogenous locus. Self-crosses of these strains yielded erg-3 mutant ascospores that produced colonies with the characteristic morphology on Vogel's sorbose agar described previously for erg-3 mutants of N. crassa. The mutants were generated by repeat-induced point mutation (RIP), a genome defense process that causes numerous G:C to A:T mutations in duplicated DNA sequences. Homozygosity for novel recessive RIP-deficient mutations was signaled by self-crosses of erg-3 duplication strains that fail to produce erg-3 mutant progeny. Using this assay we isolated a UV-induced mutant with a putative partial RIP defect. RIP-induced mutants were isolated in rid-1 and sad-1, which are essential genes, respectively, for RIP and another genome defense mechanism called meiotic silencing by unpaired DNA. PMID- 15280232 TI - Caenorhabditis elegans WASP and Ena/VASP proteins play compensatory roles in morphogenesis and neuronal cell migration. AB - We report here that WASP and Ena/VASP family proteins play overlapping roles in C. elegans morphogenesis and neuronal cell migration. Specifically, these studies demonstrate that UNC-34/Ena plays a role in morphogenesis that is revealed only in the absence of WSP-1 function and that WSP-1 has a role in neuronal cell migration that is revealed only in the absence of UNC-34/Ena activity. To identify additional genes that act in parallel to unc-34/ena during morphogenesis, we performed a screen for synthetic lethals in an unc-34 null mutant background utilizing an RNAi feeding approach. To our knowledge, this is the first reported RNAi-based screen for genetic interactors. As a result of this screen, we identified a second C. elegans WASP family protein, wve-1, that is most homologous to SCAR/WAVE proteins. Animals with impaired wve-1 function display defects in gastrulation, fail to undergo proper morphogenesis, and exhibit defects in neuronal cell migrations and axon outgrowth. Reducing wve-1 levels in either unc-34/ena or wsp-1 mutant backgrounds also leads to a significant enhancement of the gastrulation and morphogenesis defects. Thus, unc 34/ena, wsp-1, and wve-1 play overlapping roles during embryogenesis and unc 34/ena and wsp-1 play overlapping roles in neuronal cell migration. These observations show that WASP and Ena/VASP proteins can compensate for each other in vivo and provide the first demonstration of a role for Ena/VASP proteins in gastrulation and morphogenesis. In addition, our results provide the first example of an in vivo role for WASP family proteins in neuronal cell migrations and cytokinesis in metazoans. PMID- 15280233 TI - lin-35/Rb cooperates with the SWI/SNF complex to control Caenorhabditis elegans larval development. AB - Null mutations in lin-35, the Caenorhabditis elegans ortholog of the mammalian Rb protein, cause no obvious morphological defects. Using a genetic approach to identify genes that may function redundantly with lin-35, we have isolated a mutation in the C. elegans psa-1 gene. lin-35; psa-1 double mutants display severe developmental defects leading to early larval arrest and adult sterility. The psa-1 gene has previously been shown to encode a C. elegans homolog of yeast SWI3, a critical component of the SWI/SNF complex, and has been shown to regulate asymmetric cell divisions during C. elegans development. We observed strong genetic interactions between psa-1 and lin-35 as well as a subset of the class B synMuv genes that include lin-37 and lin-9. Loss-of-function mutations in lin-35, lin-37, and lin-9 strongly enhanced the defects of asymmetric T cell division associated with a psa-1 mutation. Our results suggest that LIN-35/Rb and a certain class B synMuv proteins collaborate with the SWI/SNF protein complex to regulate the T cell division as well as other events essential for larval growth. PMID- 15280234 TI - Association between nucleotide variation in Egfr and wing shape in Drosophila melanogaster. AB - As part of an effort to dissect quantitative trait locus effects to the nucleotide level, association was assessed between 238 single-nucleotide and 20 indel polymorphisms spread over 11 kb of the Drosophila melanogaster Egfr locus and nine relative warp measures of wing shape. One SNP in a conserved potential regulatory site for a GAGA factor in the promoter of alternate first exon 2 approaches conservative experiment-wise significance (P < 0.00003) in the sample of 207 lines for association with the location of the crossveins in the central region of the wing. Several other sites indicate marginal association with one or more other aspects of shape. No strong effects of sex or population of origin were detected with measures of shape, but two different sites were strongly associated with overall wing size in interaction with these fixed factors. Whole gene sequencing in very large samples, rather than selective genotyping, would appear to be the only strategy likely to be successful for detecting subtle associations in species with high polymorphism and little haplotype structure. However, these features severely limit the ability of linkage disequilibrium mapping in Drosophila to resolve quantitative effects to single nucleotides. PMID- 15280235 TI - Nucleotide variation in the Egfr locus of Drosophila melanogaster. AB - The Epidermal growth factor receptor is an essential gene with diverse pleiotropic roles in development throughout the animal kingdom. Analysis of sequence diversity in 10.9 kb covering the complete coding region and 6.4 kb of potential regulatory regions in a sample of 250 alleles from three populations of Drosophila melanogaster suggests that the intensity of different population genetic forces varies along the locus. A total of 238 independent common SNPs and 20 indel polymorphisms were detected, with just six common replacements affecting >1475 amino acids, four of which are in the short alternate first exon. Sequence diversity is lowest in a 2-kb portion of intron 2, which is also highly conserved in comparison with D. simulans and D. pseudoobscura. Linkage disequilibrium decays to background levels within 500 bp of most sites, so haplotypes are generally restricted to up to 5 polymorphisms. The two North American samples from North Carolina and California have diverged in allele frequency at a handful of individual SNPs, but a Kenyan sample is both more divergent and more polymorphic. The effect of sample size on inference of the roles of population structure, uneven recombination, and weak selection in patterning nucleotide variation in the locus is discussed. PMID- 15280236 TI - The direct interaction between ASH2, a Drosophila trithorax group protein, and SKTL, a nuclear phosphatidylinositol 4-phosphate 5-kinase, implies a role for phosphatidylinositol 4,5-bisphosphate in maintaining transcriptionally active chromatin. AB - The products of trithorax group (trxG) genes maintain active transcription of many important developmental regulatory genes, including homeotic genes. Several trxG proteins have been shown to act in multimeric protein complexes that modify chromatin structure. ASH2, the product of the Drosophila trxG gene absent, small, or homeotic discs 2 (ash2) is a component of a 500-kD complex. In this article, we provide biochemical evidence that ASH2 binds directly to Skittles (SKTL), a predicted phosphatidylinositol 4-phosphate 5-kinase, and genetic evidence that the association of these proteins is functionally significant. We also show that histone H1 hyperphosphorylation is dramatically increased in both ash2 and sktl mutant polytene chromosomes. These results suggest that ASH2 maintains active transcription by binding a producer of nuclear phosphoinositides and downregulating histone H1 hyperphosphorylation. PMID- 15280237 TI - Requirement for sex comb on midleg protein interactions in Drosophila polycomb group repression. AB - The Drosophila Sex Comb on Midleg (SCM) protein is a transcriptional repressor of the Polycomb group (PcG). Although genetic studies establish SCM as a crucial PcG member, its molecular role is not known. To investigate how SCM might link to PcG complexes, we analyzed the in vivo role of a conserved protein interaction module, the SPM domain. This domain is found in SCM and in another PcG protein, Polyhomeotic (PH), which is a core component of Polycomb repressive complex 1 (PRC1). SCM-PH interactions in vitro are mediated by their respective SPM domains. Yeast two-hybrid and in vitro binding assays were used to isolate and characterize >30 missense mutations in the SPM domain of SCM. Genetic rescue assays showed that SCM repressor function in vivo is disrupted by mutations that impair SPM domain interactions in vitro. Furthermore, overexpression of an isolated, wild-type SPM domain produced PcG loss-of-function phenotypes in flies. Coassembly of SCM with a reconstituted PRC1 core complex shows that SCM can partner with PRC1. However, gel filtration chromatography showed that the bulk of SCM is biochemically separable from PH in embryo nuclear extracts. These results suggest that SCM, although not a core component of PRC1, interacts and functions with PRC1 in gene silencing. PMID- 15280238 TI - Characterization of conditionally expressed mutants affecting age-specific survival in inbred lines of Drosophila melanogaster: lethal conditions and temperature-sensitive periods. AB - The specific genetic basis of inbreeding depression is poorly understood. To address this question, two conditionally expressed lethal effects that were found to cause line-specific life span reductions in two separate inbred lines of Drosophila melanogaster were characterized phenotypically and genetically in terms of whether the accelerated mortality effects are dominant or recessive. The mortality effect in one line (I4) is potentially a temperature-sensitive semilethal that expresses in adult males only and is partially dominant. The other line (I10) responds as one would expect for a recessive lethal. It requires a cold shock for expression and is cold sensitive. Flies exhibiting this lethal condition responded as pupae and freshly eclosed imagoes. The effect is recessive in both males and females. The expression of the lethal effects in both lines is highly dependent upon environmental conditions. These results will serve as a basis for more detailed and mechanistic genetic research on inbreeding depression and are relevant to sex- and environment-specific effects on life span observed in quantitative trait loci studies using inbred lines. PMID- 15280239 TI - The quantitative genetic basis of male mating behavior in Drosophila melanogaster. AB - Male mating behavior is an important component of fitness in Drosophila and displays segregating variation in natural populations. However, we know very little about the genes affecting naturally occurring variation in mating behavior, their effects, or their interactions. Here, we have mapped quantitative trait loci (QTL) affecting courtship occurrence, courtship latency, copulation occurrence, and copulation latency that segregate between a D. melanogaster strain selected for reduced male mating propensity (2b) and a standard wild-type strain (Oregon-R). Mating behavior was assessed in a population of 98 recombinant inbred lines derived from these two strains and QTL affecting mating behavior were mapped using composite interval mapping. We found four QTL affecting male mating behavior at cytological locations 1A;3E, 57C;57F, 72A;85F, and 96F;99A. We used deficiency complementation mapping to map the autosomal QTL with much higher resolution to five QTL at 56F5;56F8, 56F9;57A3, 70E1;71F4, 78C5;79A1, and 96F1;97B1. Quantitative complementation tests performed for 45 positional candidate genes within these intervals revealed 7 genes that failed to complement the QTL: eagle, 18 wheeler, Enhancer of split, Polycomb, spermatocyte arrest, l(2)05510, and l(2)k02206. None of these genes have been previously implicated in mating behavior, demonstrating that quantitative analysis of subtle variants can reveal novel pleiotropic effects of key developmental loci on behavior. PMID- 15280240 TI - Quantitative trait loci for sexual isolation between Drosophila simulans and D. mauritiana. AB - Sexual isolating mechanisms that act before fertilization are often considered the most important genetic barriers leading to speciation in animals. While recent progress has been made toward understanding the genetic basis of the postzygotic isolating mechanisms of hybrid sterility and inviability, little is known about the genetic basis of prezygotic sexual isolation. Here, we map quantitative trait loci (QTL) contributing to prezygotic reproductive isolation between the sibling species Drosophila simulans and D. mauritiana. We mapped at least seven QTL affecting discrimination of D. mauritiana females against D. simulans males, three QTL affecting D. simulans male traits against which D. mauritiana females discriminate, and six QTL affecting D. mauritiana male traits against which D. simulans females discriminate. QTL affecting sexual isolation act additively, are largely different in males and females, and are not disproportionately concentrated on the X chromosome: The QTL of greatest effect are located on chromosome 3. Unlike the genetic components of postzygotic isolation, the loci for prezygotic isolation do not interact epistatically. The observation of a few QTL with moderate to large effects will facilitate positional cloning of genes underlying sexual isolation. PMID- 15280241 TI - The Mod(mdg4) component of the Su(Hw) insulator inserted in the P transposon can repress its mobility in Drosophila melanogaster. AB - Transposable element P of Drosophila melanogaster is one of the best characterized eukaryotic transposons. Successful transposition requires the interaction between transposase complexes at both termini of the P element. Here we found that insertion of one or two copies of the Su(Hw) insulator in the P transposon reduces the frequency of its transposition. Inactivation of a Mod(mdg4) component of the Su(Hw) insulator suppresses the insulator effect. Thus, the Su(Hw) insulator can modulate interactions between transposase complexes bound to the ends of the P transposon in germ cells. PMID- 15280242 TI - The genetic covariance among clinal environments after adaptation to an environmental gradient in Drosophila serrata. AB - We examined the genetic basis of clinal adaptation by determining the evolutionary response of life-history traits to laboratory natural selection along a gradient of thermal stress in Drosophila serrata. A gradient of heat stress was created by exposing larvae to a heat stress of 36 degrees for 4 hr for 0, 1, 2, 3, 4, or 5 days of larval development, with the remainder of development taking place at 25 degrees. Replicated lines were exposed to each level of this stress every second generation for 30 generations. At the end of selection, we conducted a complete reciprocal transfer experiment where all populations were raised in all environments, to estimate the realized additive genetic covariance matrix among clinal environments in three life-history traits. Visualization of the genetic covariance functions of the life-history traits revealed that the genetic correlation between environments generally declined as environments became more different and even became negative between the most different environments in some cases. One exception to this general pattern was a life history trait representing the classic trade-off between development time and body size, which responded to selection in a similar genetic fashion across all environments. Adaptation to clinal environments may involve a number of distinct genetic effects along the length of the cline, the complexity of which may not be fully revealed by focusing primarily on populations at the ends of the cline. PMID- 15280243 TI - Selective and mutational patterns associated with gene expression in humans: influences on synonymous composition and intron presence. AB - We report the results of a comprehensive study of the influence of gene expression on synonymous codons, amino acid composition, and intron presence and size in human protein-coding genes. First, in addition to a strong effect of isochores, we have detected the influence of transcription-associated mutational biases (TAMB) on gene composition. Genes expressed in different tissues show diverse degrees of TAMB, with genes expressed in testis showing the greatest influence. Second, the study of tissues with no evidence of TAMB reveals a consistent set of optimal synonymous codons favored in highly expressed genes. This result exposes the consequences of natural selection on synonymous composition to increase efficiency of translation in the human lineage. Third, overall amino acid composition of proteins closely resembles tRNA abundance but there is no difference in amino acid composition in differentially expressed genes. Fourth, there is a negative relationship between expression and CDS length. Significantly, this is observed only among genes with introns, suggesting that the cause for this relationship in humans cannot be associated only with costs of amino acid biosynthesis. Fifth, we show that broadly and highly expressed genes have more, although shorter, introns. The selective advantage for having more introns in highly expressed genes is likely counterbalanced by containment of transcriptional costs and a minimum exon size for proper splicing. PMID- 15280244 TI - The GanB Galpha-protein negatively regulates asexual sporulation and plays a positive role in conidial germination in Aspergillus nidulans. AB - We isolated the ganB gene encoding the Galpha-protein homolog from Aspergillus nidulans. To investigate the cellular function of GanB, various mutant strains were isolated. Deletion of constitutively inactive ganB mutants showed conidiation and derepressed brlA expression in a submerged culture. Constitutive activation of GanB caused a reduction in hyphal growth and a severe defect in asexual sporulation. We therefore propose that GanB may negatively regulate asexual sporulation through the BrlA pathway. In addition, deletion or constitutive inactivation of GanB reduced germination rate while constitutive activation led to precocious germination. Furthermore, conidia of a constitutively active mutant could germinate even without carbon source. Taken together, these results indicated that GanB plays a positive role during germination, possibly through carbon source sensing, and negatively regulates asexual conidiation in A. nidulans. PMID- 15280245 TI - Wide-cross whole-genome radiation hybrid mapping of cotton (Gossypium hirsutum L.). AB - We report the development and characterization of a "wide-cross whole-genome radiation hybrid" (WWRH) panel from cotton (Gossypium hirsutum L.). Chromosomes were segmented by gamma-irradiation of G. hirsutum (n = 26) pollen, and segmented chromosomes were rescued after in vivo fertilization of G. barbadense egg cells (n = 26). A 5-krad gamma-ray WWRH mapping panel (N = 93) was constructed and genotyped at 102 SSR loci. SSR marker retention frequencies were higher than those for animal systems and marker retention patterns were informative. Using the program RHMAP, 52 of 102 SSR markers were mapped into 16 syntenic groups. Linkage group 9 (LG 9) SSR markers BNL0625 and BNL2805 had been colocalized by linkage analysis, but their order was resolved by differential retention among WWRH plants. Two linkage groups, LG 13 and LG 9, were combined into one syntenic group, and the chromosome 1 linkage group marker BNL4053 was reassigned to chromosome 9. Analyses of cytogenetic stocks supported synteny of LG 9 and LG 13 and localized them to the short arm of chromosome 17. They also supported reassignment of marker BNL4053 to the long arm of chromosome 9. A WWRH map of the syntenic group composed of linkage groups 9 and 13 was constructed by maximum likelihood analysis under the general retention model. The results demonstrate not only the feasibility of WWRH panel construction and mapping, but also complementarity to traditional linkage mapping and cytogenetic methods. PMID- 15280246 TI - Comparative evolutionary histories of chitinase genes in the Genus zea and Family poaceae. AB - Patterns of DNA sequence diversity vary widely among genes encoding proteins that protect plants against pathogens and herbivores. Comparative studies may help determine whether these differences are due to the strength of selection acting on different types of defense, in different evolutionary lineages, or both. I analyzed sequence diversity at three chitinases, a well-studied component of defense, in two species of Zea and several Poaceae taxa. Although the Zea species are closely related and these genes code for proteins with similar biochemical function, patterns of diversity vary widely within and among species. Intraspecific diversity at chiB, chiI, and Z. mays ssp. parviglumis chiA are consistent with a neutral-equilibrium model whereas chiA had no segregating sites within Z. diploperennis--consistent with a recent and strong selective sweep. Codons identified as having diverged among Poaceae taxa in response to positive selection were significantly overrepresented among targets of selection in Arabis, suggesting common responses to selection in distantly related plant taxa. Divergence of the recent duplicates chiA and chiB is consistent with positive selection but relaxed constraint cannot be rejected. Weak evidence for adaptive divergence of these duplicated downstream components of defense contrasts with strong evidence for adaptive divergence of genes involved in pathogen recognition. PMID- 15280247 TI - Diverse evolutionary mechanisms shape the type III effector virulence factor repertoire in the plant pathogen Pseudomonas syringae. AB - Many gram-negative pathogenic bacteria directly translocate effector proteins into eukaryotic host cells via type III delivery systems. Type III effector proteins are determinants of virulence on susceptible plant hosts; they are also the proteins that trigger specific disease resistance in resistant plant hosts. Evolution of type III effectors is dominated by competing forces: the likely requirement for conservation of virulence function, the avoidance of host defenses, and possible adaptation to new hosts. To understand the evolutionary history of type III effectors in Pseudomonas syringae, we searched for homologs to 44 known or candidate P. syringae type III effectors and two effector chaperones. We examined 24 gene families for distribution among bacterial species, amino acid sequence diversity, and features indicative of horizontal transfer. We assessed the role of diversifying and purifying selection in the evolution of these gene families. While some P. syringae type III effectors were acquired recently, others have evolved predominantly by descent. The majority of codons in most of these genes were subjected to purifying selection, suggesting selective pressure to maintain presumed virulence function. However, members of 7 families had domains subject to diversifying selection. PMID- 15280248 TI - Linkage disequilibrium mapping of Arabidopsis CRY2 flowering time alleles. AB - The selfing plant Arabidopsis thaliana has been proposed to be well suited for linkage disequilibrium (LD) mapping as a means of identifying genes underlying natural trait variation. Here we apply LD mapping to examine haplotype variation in the genomic region of the photoperiod receptor CRYPTOCHROME2 and associated flowering time variation. CRY2 DNA sequences reveal strong LD and the existence of two highly differentiated haplogroups (A and B) across the gene; in addition, a haplotype possessing a radical glutamine-to-serine replacement (AS) occurs within the more common haplogroup. Growth chamber and field experiments using an unstratified population of 95 ecotypes indicate that under short-day photoperiod, the AS and B haplogroups are both highly significantly associated with early flowering. Data from six genes flanking CRY2 indicate that these haplogroups are limited to an approximately 65-kb genomic region around CRY2. Whereas the B haplogroup cannot be delimited to <16 kb around CRY2, the AS haplogroup is characterized almost exclusively by the nucleotide polymorphisms directly associated with the serine replacement in CRY2; this finding strongly suggests that the serine substitution is directly responsible for the AS early flowering phenotype. This study demonstrates the utility of LD mapping for elucidating the genetic basis of natural, ecologically relevant variation in Arabidopsis. PMID- 15280249 TI - The distribution of transgene insertion sites in barley determined by physical and genetic mapping. AB - The exact site of transgene insertion into a plant host genome is one feature of the genetic transformation process that cannot, at present, be controlled and is often poorly understood. The site of transgene insertion may have implications for transgene stability and for potential unintended effects of the transgene on plant metabolism. To increase our understanding of transgene insertion sites in barley, a detailed analysis of transgene integration in independently derived transgenic barley lines was carried out. Fluorescence in situ hybridization (FISH) was used to physically map 23 transgene integration sites from 19 independent barley lines. Genetic mapping further confirmed the location of the transgenes in 11 of these lines. Transgene integration sites were present only on five of the seven barley chromosomes. The pattern of transgene integration appeared to be nonrandom and there was evidence of clustering of independent transgene insertion events within the barley genome. In addition, barley genomic regions flanking the transgene insertion site were isolated for seven independent lines. The data from the transgene flanking regions indicated that transgene insertions were preferentially located in gene-rich areas of the genome. These results are discussed in relation to the structure of the barley genome. PMID- 15280250 TI - Clonal mosaic analysis of EMPTY PERICARP2 reveals nonredundant functions of the duplicated HEAT SHOCK FACTOR BINDING PROTEINs during maize shoot development. AB - The paralogous maize proteins EMPTY PERICARP2 (EMP2) and HEAT SHOCK FACTOR BINDING PROTEIN2 (HSBP2) each contain a single recognizable motif: the coiled coil domain. EMP2 and HSBP2 accumulate differentially during maize development and heat stress. Previous analyses revealed that EMP2 is required for regulation of heat shock protein (hsp) gene expression and also for embryo morphogenesis. Developmentally abnormal emp2 mutant embryos are aborted during early embryogenesis. To analyze EMP2 function during postembryonic stages, plants mosaic for sectors of emp2 mutant tissue were constructed. Clonal sectors of emp2 mutant tissue revealed multiple defects during maize vegetative shoot development, but these sector phenotypes are not correlated with aberrant hsp gene regulation. Furthermore, equivalent phenotypes are observed in emp2 sectored plants grown under heat stress and nonstress conditions. Thus, the function of EMP2 during regulation of the heat stress response can be separated from its role in plant development. The discovery of emp2 mutant phenotypes in postembryonic shoots reveals that the duplicate genes emp2 and hsbp2 encode nonredundant functions throughout maize development. Distinct developmental phenotypes correlated with the developmental timing, position, and tissue layer of emp2 mutant sectors, suggesting that EMP2 has evolved diverse developmental functions in the maize shoot. PMID- 15280252 TI - Quantitative genetic models for describing simultaneous and recursive relationships between phenotypes. AB - Multivariate models are of great importance in theoretical and applied quantitative genetics. We extend quantitative genetic theory to accommodate situations in which there is linear feedback or recursiveness between the phenotypes involved in a multivariate system, assuming an infinitesimal, additive, model of inheritance. It is shown that structural parameters defining a simultaneous or recursive system have a bearing on the interpretation of quantitative genetic parameter estimates (e.g., heritability, offspring-parent regression, genetic correlation) when such features are ignored. Matrix representations are given for treating a plethora of feedback-recursive situations. The likelihood function is derived, assuming multivariate normality, and results from econometric theory for parameter identification are adapted to a quantitative genetic setting. A Bayesian treatment with a Markov chain Monte Carlo implementation is suggested for inference and developed. When the system is fully recursive, all conditional posterior distributions are in closed form, so Gibbs sampling is straightforward. If there is feedback, a Metropolis step may be embedded for sampling the structural parameters, since their conditional distributions are unknown. Extensions of the model to discrete random variables and to nonlinear relationships between phenotypes are discussed. PMID- 15280251 TI - Structure and expression of maize phytochrome family homeologs. AB - To begin the study of phytochrome signaling in maize, we have cloned and characterized the phytochrome gene family from the inbred B73. Through DNA gel blot analysis of maize genomic DNA and BAC library screens, we show that the PhyA, PhyB, and PhyC genes are each duplicated once in the genome of maize. Each gene pair was positioned to homeologous regions of the genome using recombinant inbred mapping populations. These results strongly suggest that the duplication of the phytochrome gene family in maize arose as a consequence of an ancient tetraploidization in the maize ancestral lineage. Furthermore, sequencing of Phy genes directly from BAC clones indicates that there are six functional phytochrome genes in maize. Through Northern gel blot analysis and a semiquantitative reverse transcriptase polymerase chain reaction assay, we determined that all six phytochrome genes are transcribed in several seedling tissues. However, expression from PhyA1, PhyB1, and PhyC1 predominate in all seedling tissues examined. Dark-grown seedlings express higher levels of PhyA and PhyB than do light-grown plants but PhyC genes are expressed at similar levels under light and dark growth conditions. These results are discussed in relation to phytochrome gene regulation in model eudicots and monocots and in light of current genome sequencing efforts in maize. PMID- 15280253 TI - The effects of intraspecific competition and stabilizing selection on a polygenic trait. AB - The equilibrium properties of an additive multilocus model of a quantitative trait under frequency- and density-dependent selection are investigated. Two opposing evolutionary forces are assumed to act: (i) stabilizing selection on the trait, which favors genotypes with an intermediate phenotype, and (ii) intraspecific competition mediated by that trait, which favors genotypes whose effect on the trait deviates most from that of the prevailing genotypes. Accordingly, fitnesses of genotypes have a frequency-independent component describing stabilizing selection and a frequency- and density-dependent component modeling competition. We study how the equilibrium structure, in particular, number, degree of polymorphism, and genetic variance of stable equilibria, is affected by the strength of frequency dependence, and what role the number of loci, the amount of recombination, and the demographic parameters play. To this end, we employ a statistical and numerical approach, complemented by analytical results, and explore how the equilibrium properties averaged over a large number of genetic systems with a given number of loci and average amount of recombination depend on the ecological and demographic parameters. We identify two parameter regions with a transitory region in between, in which the equilibrium properties of genetic systems are distinctively different. These regions depend on the strength of frequency dependence relative to pure stabilizing selection and on the demographic parameters, but not on the number of loci or the amount of recombination. We further study the shape of the fitness function observed at equilibrium and the extent to which the dynamics in this model are adaptive, and we present examples of equilibrium distributions of genotypic values under strong frequency dependence. Consequences for the maintenance of genetic variation, the detection of disruptive selection, and models of sympatric speciation are discussed. PMID- 15280254 TI - Identifying the susceptibility gene(s) in a set of trait-linked genes using genotype data. AB - There are generally three steps to isolate a disease linkage-susceptibility gene: genome-wide scan, fine mapping, and, last, positional cloning. The last step is time consuming and involves intensive laboratory work. In some cases, fine mapping cannot proceed further on a set of markers because they are tightly linked. For years, genetic statisticians have been trying different ways to narrow the fine-mapping results to provide some guidance for the next step of laboratory work. Although these methods are practical and efficient, most of them are based on IBD data, which usually can be inferred only from the genotype data with some uncertainty. The corresponding methods thus have no greater power than one using genotype data directly. Also, IBD-based methods apply only to relative pair data. Here, using genotype data, we have developed a statistical hypothesis testing method to pinpoint a SNP, or SNPs, suspected of responsibility for a disease trait linkage among a set of SNPs tightly linked in a region. Our method uses genotype data of affected individuals or case-control studies, which are widely available in the laboratory. The testing statistic can be constructed using any genotype-based disease-marker disequilibrium measure and is asymptotically distributed as a chi-square mixture. This method can be used for singleton data, relative pair data, or general pedigree data. We have applied the method to simulated data as well as a real data set; it gives satisfactory results. PMID- 15280255 TI - Estimating recombination rates using three-site likelihoods. AB - We introduce a new method for jointly estimating crossing-over and gene conversion rates using sequence polymorphism data. The method calculates probabilities for subsets of the data consisting of three segregating sites and then forms a composite likelihood by multiplying together the probabilities of many subsets. Simulations show that this new method performs better than previously proposed methods for estimating gene conversion rates, but that all methods require large amounts of data to provide reliable estimates. While existing methods can easily estimate an "average" gene conversion rate over many loci, they cannot reliably estimate gene conversion rates for a single region of the genome. PMID- 15280256 TI - Redistribution of gene frequency and changes of genetic variation following a bottleneck in population size. AB - Although the distribution of frequencies of genes influencing quantitative traits is important to our understanding of their genetic basis and their evolution, direct information from laboratory experiments is very limited. In theory, different models of selection and mutation generate different predictions of frequency distributions. When a large population at mutation-selection balance passes through a rapid bottleneck in size, the frequency distribution of genes is dramatically altered, causing changes in observable quantities such as the mean and variance of quantitative traits. We investigate the gene frequency distribution of a population at mutation-selection balance under a joint-effect model of real stabilizing and pleiotropic selection and its redistribution and thus changes of the genetic properties of metric and fitness traits after the population passes a rapid bottleneck and expands in size. If all genes that affect the trait are neutral with respect to fitness, the additive genetic variance (VA) is always reduced by a bottleneck in population size, regardless of their degree of dominance. For genes that have been under selection, VA increases following a bottleneck if they are (partially) recessive, while the dominance variance increases substantially for any degree of dominance. With typical estimates of mutation parameters, the joint-effect model can explain data from laboratory experiments on the effect of bottlenecking on fitness and morphological traits, providing further support for it as a plausible mechanism for maintenance of quantitative genetic variation. PMID- 15280257 TI - Epistasis of quantitative trait loci under different gene action models. AB - Modeling and detecting nonallelic (epistatic) effects at multiple quantitative trait loci (QTL) often assume that the study population is in zygotic equilibrium (i.e., genotypic frequencies at different loci are products of corresponding single-locus genotypic frequencies). However, zygotic associations can arise from physical linkages between different loci or from many evolutionary and demographic processes even for unlinked loci. We describe a new model that partitions the two-locus genotypic values in a zygotic disequilibrium population into equilibrium and residual portions. The residual portion is of course due to the presence of zygotic associations. The equilibrium portion has eight components including epistatic effects that can be defined under three commonly used equilibrium models, Cockerham's model, F2-metric, and F(infinity)-metric models. We evaluate our model along with these equilibrium models theoretically and empirically. While all the equilibrium models require zygotic equilibrium, Cockerham's model is the most general, allowing for Hardy-Weinberg disequilibrium and arbitrary gene frequencies at individual loci whereas F2-metric and F(infinity)-metric models require gene frequencies of one-half in a Hardy Weinberg equilibrium population. In an F2 population with two unlinked loci, Cockerham's model is reduced to the F2-metric model and thus both have a desirable property of orthogonality among the genic effects; the genic effects under the F(infinity)-metric model are not orthogonal but they can be easily translated into those under the F2-metric model through a simple relation. Our model is reduced to these equilibrium models in the absence of zygotic associations. The results from our empirical analysis suggest that the residual genetic variance arising from zygotic associations can be substantial and may be an important source of bias in QTL mapping studies. PMID- 15280258 TI - Genome-wide patterns of nucleotide substitution reveal stringent functional constraints on the protein sequences of thermophiles. AB - To test the hypothesis that the proteins of thermophilic prokaryotes are subject to unusually stringent functional constraints, we estimated the numbers of synonymous and nonsynonymous nucleotide substitutions per site between 17,957 pairs of orthologous genes from 22 pairs of closely related species of Archaea and Bacteria. The average ratio of nonsynonymous to synonymous substitutions was significantly lower in thermophiles than in nonthermophiles, and this effect was observed in both Archaea and Bacteria. There was no evidence that this difference could be explained by factors such as nucleotide content bias. Rather, the results support the hypothesis that proteins of thermophiles are subject to unusually strong purifying selection, leading to a reduced overall level of amino acid evolution per mutational event. The results show that genome-wide patterns of sequence evolution can be influenced by natural selection exerted by a species' environment and shed light on a previous observation that relatively few of the mutations arising in a thermophilic archaeon were nucleotide substitutions in contrast to indels. PMID- 15280260 TI - Does random X-inactivation in mammals reflect a random choice between two X chromosomes? AB - Inactivation of the maternal or paternal X chromosome in a mammalian embryonic XX cell is believed to involve random choice between the two X's. We propose two alternative models. One suggests that choice is not random, while the other is consistent with random choice, but not one between two X's. PMID- 15280259 TI - Linkage disequilibrium as a signature of selective sweeps. AB - The hitchhiking effect of a beneficial mutation, or a selective sweep, generates a unique distribution of allele frequencies and spatial distribution of polymorphic sites. A composite-likelihood test was previously designed to detect these signatures of a selective sweep, solely on the basis of the spatial distribution and marginal allele frequencies of polymorphisms. As an excess of linkage disequilibrium (LD) is also known to be a strong signature of a selective sweep, we investigate how much statistical power is increased by the inclusion of information regarding LD. The expected pattern of LD is predicted by a genealogical approach. Both theory and simulation suggest that strong LD is generated in narrow regions at both sides of the location of beneficial mutation. However, a lack of LD is expected across the two sides. We explore various ways to detect this signature of selective sweeps by statistical tests. A new composite-likelihood method is proposed to incorporate information regarding LD. This method enables us to detect selective sweeps and estimate the parameters of the selection model better than the previous composite-likelihood method that does not take LD into account. However, the improvement made by including LD is rather small, suggesting that most of the relevant information regarding selective sweeps is captured by the spatial distribution and marginal allele frequencies of polymorphisms. PMID- 15280262 TI - Notification that new names and new combinations have appeared in volume 54, part 2, of the IJSEM. PMID- 15280261 TI - Validation list no. 98. Validation of publication of new names and new combinations previously effectively published outside the IJSEM. AB - The purpose of this announcement is to effect the valid publication of the following new names and new combinations under the procedure described in the Bacteriological Code (1990 Revision). Authors and other individuals wishing to have new names and/or combinations included in future lists should send three copies of the pertinent reprint or photocopies thereof to the IJSEM Editorial Office for confirmation that all of the other requirements for valid publication have been met. It is also a requirement of IJSEM and the ICSP that authors of new species, new subspecies and new combinations provide evidence that types are deposited in two recognized culture collections in two different countries (i.e. documents certifying deposition and availability of type strains). It should be noted that the date of valid publication of these new names and combinations is the date of publication of this list, not the date of the original publication of the names and combinations. The authors of the new names and combinations are as given below, and these authors' names will be included in the author index of the present issue and in the volume author index. Inclusion of a name on these lists validates the publication of the name and thereby makes it available in bacteriological nomenclature. The inclusion of a name on this list is not to be construed as taxonomic acceptance of the taxon to which the name is applied. Indeed, some of these names may, in time, be shown to be synonyms, or the organisms may be transferred to another genus, thus necessitating the creation of a new combination. PMID- 15280263 TI - Reclassification of salt-water Bdellovibrio sp. as Bacteriovorax marinus sp. nov. and Bacteriovorax litoralis sp. nov. AB - Bdellovibrios are unique, predatory bacteria with an intraperiplasmic growth and multiplication phase within their prey, which consists of many Gram-negative bacteria. Until recently, all bacteria that exhibited these traits were included in the genus Bdellovibrio. However, analysis of 16S rDNA sequences and other studies have demonstrated substantial genotypic, phenotypic and ecotypic diversity among the organisms in this genus (Baer et al., 2000; Snyder et al., 2002). This has resulted in reclassification of Bdellovibrio stolpii and Bdellovibrio starrii into the newly constructed genus Bacteriovorax (Baer et al., 2000). In this study, examination of marine isolates of Bdellovibrio (designated SJT, AQ and JS5T) has revealed them to be related more closely to the newly designated genus Bacteriovorax. Phylogenetic analysis of 16S rRNA gene sequences revealed that marine isolates SJT, AQ and JS5T clustered in a separate clade from Bdellovibrio bacteriovorus 100T as part of the clade that contains Bacteriovorax spp., indicating a much closer taxonomic relationship to the latter. DNA-DNA hybridization experiments also demonstrated <5 % similarity between Bdellovibrio bacteriovorus 100T and the marine isolates. Distinct differences between the salt water group and Bdellovibrio spp. were also observed by determination of DNA G+C content, salinity growth testing and antibiotic sensitivity analysis. On the basis of the results from the studies described above, it is proposed that marine isolates SJT (=ATCC BAA-682T=DSM 15412T) and JS5T (=ATCC BAA-684T=DSM 15409T) should be classified within the genus Bacteriovorax as the type strains of Bacteriovorax marinus sp. nov. and Bacteriovorax litoralis sp. nov., respectively. PMID- 15280264 TI - Maribacter gen. nov., a new member of the family Flavobacteriaceae, isolated from marine habitats, containing the species Maribacter sedimenticola sp. nov., Maribacter aquivivus sp. nov., Maribacter orientalis sp. nov. and Maribacter ulvicola sp. nov. AB - Six novel gliding, heterotrophic, Gram-negative, yellow-pigmented, aerobic, oxidase- and catalase-positive bacteria were isolated from the green alga Ulva fenestrata, sea water and a bottom sediment sample collected in the Gulf of Peter the Great, Sea of Japan. 16S rRNA gene sequence analysis revealed that the strains studied were members of the family Flavobacteriaceae. On the basis of their phenotypic, chemotaxonomic, genotypic and phylogenetic characteristics, the novel bacteria have been assigned to the new genus Maribacter gen. nov., as Maribacter sedimenticola sp. nov., Maribacter orientalis sp. nov., Maribacter aquivivus sp. nov. and Maribacter ulvicola sp. nov., with the type strains KMM 3903T (=KCTC 12966T=CCUG 47098T), KMM 3947T (=KCTC 12967T=CCUG 48008T), KMM 3949T (=KCTC 12968T=CCUG 48009T) and KMM 3951T (=KCTC 12969T=DSM 15366T), respectively. PMID- 15280265 TI - 'Candidatus Phytoplasma spartii', 'Candidatus Phytoplasma rhamni' and 'Candidatus Phytoplasma allocasuarinae', respectively associated with spartium witches' broom, buckthorn witches'-broom and allocasuarina yellows diseases. AB - Spartium witches'-broom (SpaWB), buckthorn witches'-broom (BWB) and allocasuarina yellows (AlloY) are witches'-broom and yellows diseases of Spartium junceum (Spanish broom), Rhamnus catharticus (buckthorn) and Allocasuarina muelleriana (Slaty she-oak), respectively. These diseases are associated with distinct phytoplasmas. The SpaWB, BWB and AlloY phytoplasmas share <97.5 % 16S rDNA sequence similarity with each other and with other known phytoplasmas, including the closely related phytoplasmas of the apple proliferation group. Also, the SpaWB, BWB and AlloY phytoplasmas each have a different natural plant host. Based on their unique properties, it is proposed to designate the mentioned phytoplasmas as novel 'Candidatus' species under the names 'Candidatus Phytoplasma spartii', 'Candidatus Phytoplasma rhamni' and 'Candidatus Phytoplasma allocasuarinae', respectively. PMID- 15280266 TI - Thiocapsa marina sp. nov., a novel, okenone-containing, purple sulfur bacterium isolated from brackish coastal and marine environments. AB - Four marine, phototrophic, purple sulfur bacteria (strains 5811T, 5812, BM-3 and BS-1) were isolated in pure culture from different brackish to marine sediments in the Mediterranean Sea, the White Sea and the Black Sea. Single cells of these strains were coccus-shaped, non-motile and did not contain gas vesicles. The colour of cell suspensions that were grown in the light was purple-red. Bacteriochlorophyll a and carotenoids of the okenone series were present as photosynthetic pigments. Photosynthetic membrane systems were of the vesicular type. Hydrogen sulfide, thiosulfate, elemental sulfur and molecular hydrogen were used as electron donors during photolithotrophic growth under anoxic conditions; carbon dioxide was utilized as the carbon source. During growth on sulfide, elemental sulfur globules were stored inside the cells. In the presence of hydrogen sulfide, several organic substances could be photoassimilated. Comparative 16S rDNA sequence analysis revealed an affiliation of these four strains to the genus Thiocapsa. Both phylogenetic analysis and the results of DNA DNA hybridization studies revealed that these strains formed a separate cluster within the genus Thiocapsa. Thus, according to phenotypic characteristics and mainly the carotenoid composition, 16S rDNA sequence analysis and DNA-DNA hybridization data, it is proposed that these strains should be classified as a novel species, Thiocapsa marina sp. nov., with strain 5811T (=DSM 5653T=ATCC 43172T) as the type strain. PMID- 15280267 TI - 'Candidatus Phytoplasma asteris', a novel phytoplasma taxon associated with aster yellows and related diseases. AB - Aster yellows (AY) group (16SrI) phytoplasmas are associated with over 100 economically important diseases worldwide and represent the most diverse and widespread phytoplasma group. Strains that belong to the AY group form a phylogenetically discrete subclade within the phytoplasma clade and are related most closely to the stolbur phytoplasma subclade, based on analysis of 16S rRNA gene sequences. AY subclade strains are related more closely to their culturable relatives, Acholeplasma spp., than any other phytoplasmas known. Within the AY subclade, six distinct phylogenetic lineages were revealed. Congruent phylogenies obtained by analyses of tuf gene and ribosomal protein (rp) operon gene sequences further resolved the diversity among AY group phytoplasmas. Distinct phylogenetic lineages were identified by RFLP analysis of 16S rRNA, tuf or rp gene sequences. Ten subgroups were differentiated, based on analysis of rp gene sequences. It is proposed that AY group phytoplasmas represent at least one novel taxon. Strain OAY, which is a member of subgroups 16SrI-B, rpI-B and tufI-B and is associated with evening primrose (Oenothera hookeri) virescence in Michigan, USA, was selected as the reference strain for the novel taxon 'Candidatus Phytoplasma asteris'. A comprehensive database of diverse AY phytoplasma strains and their geographical distribution is presented. PMID- 15280268 TI - Paenibacillus massiliensis sp. nov., Paenibacillus sanguinis sp. nov. and Paenibacillus timonensis sp. nov., isolated from blood cultures. AB - Gram-positive, spore-forming rods were isolated from blood cultures of three different patients. Based on phylogenetic analyses, these strains were placed within the Paenibacillus cluster and specific phenotypic characteristics for each strain were described. Levels of 16S rRNA gene sequence similarity between existing Paenibacillus species and the three novel strains 2301065T, 2301032T and 2301083T were 87.6-94.4, 88.5-95.4 and 87.5-96.0 %, respectively, and anteiso branched C(15 : 0) was the major fatty acid. On the basis of phenotypic data and phylogenetic inference, it is proposed that these strains should be designated Paenibacillus massiliensis sp. nov., Paenibacillus sanguinis sp. nov. and Paenibacillus timonensis sp. nov. The type strains are respectively strain 2301065T (=CIP 107939T=CCUG 48215T), strain 2301083T (=CIP 107938T=CCUG 48214T) and strain 2301032T (=CIP 108005T=CCUG 48216T). PMID- 15280269 TI - Taxonomic characterization of nine strains isolated from clinical and environmental specimens, and proposal of Corynebacterium tuberculostearicum sp. nov. AB - Nine unidentified Gram-positive, lipophilic corynebacteria were isolated from clinical and food samples and subjected to a polyphasic taxonomic analysis. The bacteria were distinguished from Corynebacterium species with validly published names by biochemical tests, fatty acid content and whole-cell protein analysis. Comparative 16S rRNA gene sequence analysis demonstrated unambiguously that the nine strains were related phylogenetically to the species 'Corynebacterium tuberculostearicum' and represented a distinct subline within the genus Corynebacterium. On the basis of both phenotypic and phylogenetic evidence, the formal description of Corynebacterium tuberculostearicum sp. nov. is proposed. The type strain of C. tuberculostearicum is Medalle XT (=LDC-20T=CIP 107291T=CCUG 45418T=ATCC 35529T). PMID- 15280270 TI - Cerasibacillus quisquiliarum gen. nov., sp. nov., isolated from a semi-continuous decomposing system of kitchen refuse. AB - A moderately thermophilic and alkaliphilic bacillus, which had been reported and designated BLx (Haruta et al., 2002), was isolated from a semi-continuous decomposing system of kitchen refuse. Cells of strain BLxT were strictly aerobic, rod-shaped, motile and spore forming. The optimum temperature and pH for growth were approximately 50 degrees C and pH 8-9. Strain BLxT was able to grow at NaCl concentrations from 0.5 to 7.5%, with optimum growth at 0.5% NaCl. The predominant menaquinone was MK-7, and the major fatty acid was iso-C(15 : 0). Phylogenetic analysis showed that strain BLxT was positioned in an independent lineage within the cluster that includes the genera Virgibacillus and Lentibacillus in Bacillus rRNA group 1. Strain BLxT exhibited 16S rDNA similarity of 92.8-94.8% to Virgibacillus species and 92.3% to Lentibacillus salicampi. Phenotypic, chemotaxonomic and phylogenetic analyses supported the classification of strain BLxT in a novel genus and species. Cerasibacillus quisquiliarum gen. nov., sp. nov. is proposed on the basis of phenotypic, chemotaxonomic and phylogenetic data. The type strain is BLxT (DSM 15825T=IAM15044T=KCTC 3815T). PMID- 15280271 TI - Paenibacillus cineris sp. nov. and Paenibacillus cookii sp. nov., from Antarctic volcanic soils and a gelatin-processing plant. AB - Seven strains of aerobic, endospore-forming bacteria were found in soil taken from an active fumarole on Lucifer Hill, Candlemas Island, South Sandwich archipelago, Antarctica, and four strains were from soil of an inactive fumarole at the foot of the hill. Amplified rDNA restriction analysis, 16S rDNA sequence comparisons, SDS-PAGE and routine phenotypic tests support the proposal of two novel species of Paenibacillus, Paenibacillus cineris sp. nov. and Paenibacillus cookii sp. nov., the type strains of which are LMG 18439T (=CIP 108109T) and LMG 18419T (=CIP 108110T), respectively. A further strain, isolated from a gelatin production process, showed more than 99% 16S rDNA sequence similarity to the proposed P. cookii type strain and, although the gelatin isolate was atypical when compared with the fumarole isolates by repeated element primed-PCR, SDS-PAGE and phenotypic analyses, it was shown by DNA-DNA reassociation studies to belong to the same species. Strains of P. cookii produce spreading growth with motile microcolonies. Both species produce swollen sporangia that are typical for the genus, they both show 97.6% 16S rDNA sequence similarity to Paenibacillus azoreducens, they have 51.5-51.6 mol% G+C in their DNA and their major fatty acid is anteiso-C(15 : 0); however, fatty acids C(16 : 0) and anteiso-C(17 : 0) represent, respectively, 18 and 10 % of the total in P. cineris, but 11 and 20% in P. cookii. PMID- 15280272 TI - 'Candidatus Phytoplasma cynodontis', the phytoplasma associated with Bermuda grass white leaf disease. AB - Bermuda grass white leaf (BGWL) is a destructive, phytoplasmal disease of Bermuda grass (Cynodon dactylon). The causal pathogen, the BGWL agent, differs from other phytoplasmas that cluster in the same major branch of the phytoplasma phylogenetic clade in <2.5% of 16S rDNA nucleotide positions, the threshold for assigning species rank to phytoplasmas under the provisional status 'Candidatus'. Thus, the objective of this work was to examine homogeneity of BGWL isolates and to determine whether there are, in addition to 16S rDNA, other markers that support delineation of the BGWL agent at the putative species level. Phylogenetic analyses revealed that the 16S rDNA sequences of BGWL strains were identical or nearly identical. Clear differences that support separation of the BGWL agent from related phytoplasmas were observed within the 16S-23S rDNA spacer sequence, by serological comparisons, in vector transmission and in host-range specificity. From these results, it can be concluded that the BGWL phytoplasma is a discrete taxon at the putative species level, for which the name 'Candidatus Phytoplasma cynodontis' is proposed. Strain BGWL-C1 was selected as the reference strain. Phytoplasmas that are associated with brachiaria white leaf, carpet grass white leaf and diseases of date palms showed 16S rDNA and/or 16S-23S rDNA spacer sequences that were identical or nearly identical to those of the BGWL phytoplasmas. However, the data available do not seem to be sufficient for a proper taxonomic assignment of these phytoplasmas. PMID- 15280273 TI - Shewanella pacifica sp. nov., a polyunsaturated fatty acid-producing bacterium isolated from sea water. AB - Six marine bacterial strains, KMM 3597T, KMM 3775, KMM 3590, KMM 3772, KMM 3605 and KMM 3601, that produce polyunsaturated fatty acids were isolated from sea water samples collected from different locations and depths in Chazhma Bay (Sea of Japan, Pacific Ocean) and characterized to clarify their taxonomic position. The DNA G+C contents of these strains were 39.5-40.3 mol%. The level of DNA hybridization between these strains was conspecific (83-96%), indicating that they represent a single genospecies. 16S rRNA gene sequence-based phylogenetic analysis of the novel strains revealed that Shewanella japonica KMM 3299T was the closest relative (99% similarity). However, DNA-DNA hybridization experiments demonstrated only 45-50% binding with DNA of S. japonica. The novel organisms grew between 4 and 33 degrees C, were neutrophilic and haemolytic, and were able to degrade starch, gelatin, agar and Tween 80. The predominant fatty acids were (%+/-sd): i13 : 0 (9.3+/-1.1); i15 : 0 (33.9+/-1.5); 16 : 0 (8.9+/-1.6); and 16 : 1omega7 (14.8+/-1.1). The fatty acid 20 : 5omega3, formed at 28 degrees C, was present at up to 5.3% total fatty acids. The major isoprenoid quinones were Q7 (21-41%) and Q8 (50-59%). The phylogenetic, genetic and physiological properties of the six strains placed them within a novel species, Shewanella pacifica sp. nov., the type strain of which is R10SW1T (=KMM 3597T=CIP 107849T). PMID- 15280274 TI - Shewanella affinis sp. nov., isolated from marine invertebrates. AB - Four marine bacterial strains, designated KMM 3587T, KMM 3586, KMM 3821 and KMM 3822, were isolated from the sipuncula Phascolosoma japonicum, a common inhabitant of Troitza Bay in the Gulf of Peter the Great (Sea of Japan region), and from an unidentified hydrocoral species collected in Makarov Bay (Iturup Islands), Kuril Islands, North-West Pacific Ocean. The strains were characterized to clarify their taxonomic position. 16S rRNA gene sequences of KMM 3587T and KMM 3586 indicated 99% similarity to Shewanella colwelliana. Despite such a high level of 16S rRNA gene sequence similarity, DNA-DNA hybridization experiments demonstrated only 45-52% binding with DNA of S. colwelliana ATCC 39565T. The DNA G+C contents of the novel strains were 45 mol% and the shared level of DNA hybridization was conspecific (81-97%), indicating that they represent a single genospecies. The novel strains were mesophilic (able to grow at 10-34 degrees C), neutrophilic and haemolytic, and able to degrade gelatin, casein and Tween 20, 40 and 80, but not starch, agar, elastin, alginate or chitin. The major fatty acids were i13 : 0, i15 : 0, 16 : 0, 16 : 1omega7 and 17 : 1omega8 (68.9% of total). The major isoprenoid quinones were Q7 (47-62%) and Q8 (26-47%). Eicosapentaenoic acid was produced in minor amounts. Based on these data, the strains are assigned to a novel species, Shewanella affinis sp. nov. (type strain KMM 3587T=CIP 107703T=ATCC BAA-642T). PMID- 15280275 TI - Methanotorris formicicus sp. nov., a novel extremely thermophilic, methane producing archaeon isolated from a black smoker chimney in the Central Indian Ridge. AB - A novel extremely thermophilic, methane-producing archaeon was isolated from a black smoker chimney at the Kairei field in the Central Indian Ridge. Cells of this isolate were irregular cocci with several flagella; motility was not observed. Growth was observed between 55 and 83 degrees C (optimum of 75 degrees C; 30 min doubling time) and between pH 6.0 and 8.5 (optimum of pH 6.7). The isolate was a strictly anaerobic, methanogenic autotroph capable of using hydrogen and carbon dioxide as sole energy and carbon sources. Formate was utilized as an alternative energy source. The G+C content of the genomic DNA was 33.3 mol%. Phylogenetic analysis based on 16S rRNA gene sequences indicated that the isolate was most closely related to Methanotorris igneus strain Kol 5T. The isolate, however, could be genetically differentiated from this species by DNA DNA hybridization analysis and on the basis of its physiological properties. The name Methanotorris formicicus sp. nov. is proposed for this isolate; the type strain is Mc-S-70T (=JCM 11930T=ATCC BAA-687T). PMID- 15280276 TI - Robiginitalea biformata gen. nov., sp. nov., a novel marine bacterium in the family Flavobacteriaceae with a higher G+C content. AB - Two Gram-negative, chemoheterotrophic, non-motile, rust-coloured, marine strains were isolated from the western Sargasso Sea by high-throughput culturing. Characterization of the two strains by polyphasic approaches indicated that they are members of the same species. Phylogenetic analyses based on 16S rRNA gene sequences using three treeing algorithms revealed that the strains formed a coherent and novel genus-level lineage within the family Flavobacteriaceae. The dominant fatty acids were branched or hydroxy acids, i15 : 0, i15 : 1 and 3-OH i17 : 0 being the most abundant. The higher DNA G+C content of the strains (55-56 mol%) clearly differentiated them from other genera of the family Flavobacteriaceae (27-44 mol%). It is proposed, from the polyphasic evidence, that the strains be placed into a novel genus and a novel species named Robiginitalea biformata gen. nov., sp. nov., with strain HTCC2501T (=ATCC BAA 864T=KCTC 12146T) as the type strain. PMID- 15280277 TI - Salegentibacter holothuriorum sp. nov., isolated from the edible holothurian Apostichopus japonicus. AB - Strain KMM 3524T was isolated from the holothurian Apostichopus japonicus living in the Sea of Japan. The bacterial strain was pigmented, non-motile, Gram negative, strictly aerobic and oxidase-, catalase- and beta-galactosidase positive. From the results of 16S rDNA sequence analysis, strain KMM 3524T was found to be related closely to Salegentibacter salegens (98.1%). DNA-DNA homology between strains KMM 3524T and S. salegens DSM 5424T was 38%; this showed clearly that the holothurian isolate KMM 3524T belongs to a novel species of the genus Salegentibacter for which the name Salegentibacter holothuriorum sp. nov. is proposed, with KMM 3524T (=NBRC 100249T=LMG 21968T) as the type strain. PMID- 15280278 TI - Microbulbifer maritimus sp. nov., isolated from an intertidal sediment from the Yellow Sea, Korea. AB - A Gram-negative, rod-shaped, non-spore-forming, slightly halophilic bacterium (strain TF-17T) was isolated from an intertidal sediment from the Yellow Sea, Korea. Pigment of strain TF-17T was similar to that of Microbulbifer elongatus, but different from those of Microbulbifer hydrolyticus and Microbulbifer salipaludis. Strain TF-17T was distinguishable from M. elongatus by some phenotypic properties, including motility, optimal growth temperature and others. Phylogenetic analysis based on 16S rDNA sequences showed that strain TF-17TT clustered with the type strains of the three Microbulbifer species with validly published names. Strain TF-17T exhibited 16S rDNA sequence similarity levels of 95.1-95.7% to the type strains of the three Microbulbifer species. The predominant respiratory lipoquinone found in strain TF-17T was ubiquinone-8. The major fatty acid was iso-C(15 : 0) and significant amounts of iso-C(11 : 0) 3-OH and iso-C(17 : 1)omega9c were also present. The DNA G+C content of strain TF-17T was 59.9 mol%. Levels of DNA-DNA relatedness between strain TF-17T and the type strains of the three Microbulbifer species were in the range 10.0-13.0%. On the basis of phenotypic and phylogenetic data and genotypic distinctiveness, strain TF-17T (=KCCM 41774T=JCM 12187T) is proposed as the type strain of a novel species of the genus Microbulbifer, Microbulbifer maritimus sp. nov. PMID- 15280279 TI - Treponema putidum sp. nov., a medium-sized proteolytic spirochaete isolated from lesions of human periodontitis and acute necrotizing ulcerative gingivitis. AB - So far, little phenotypic heterogeneity has been detected in cultured oral treponemes with trypsin-like proteolytic activity, and all have been assigned to the species Treponema denticola. However, comparisons of protein patterns and antigen expression in our collection of proteolytic oral treponemes occasionally identified isolates with a unique phenotype; e.g. strain OMZ 830 (=ATCC 700768), which qualified as a 'pathogen-related oral spirochaete' due to the presence of a approximately 37 kDa protein reactive with the Treponema pallidum FlaA-specific mAb H9-2. In addition to such single isolates, a homogeneous group of seven independent strains is described that were highly motile, medium-sized, proteolytic but asaccharolytic spirochaetes and were cultured from human gingivitis, periodontitis and acute necrotizing ulcerative gingivitis in medium OMIZ-Pat supplemented with 1% human serum and antibiotics. Growth of these spirochaetes in OMIZ-Pat was not dependent on, but was stimulated by, human or bovine serum. Carbohydrates were neither required nor stimulatory for growth. The protein and antigen patterns of total cell extracts of these organisms separated by SDS-PAGE were distinct from those of all previously cultured spirochaetes, with highest similarity to T. denticola. The novel spirochaete has a 2 : 4 : 2 arrangement of the periplasmic flagella, similar to T. denticola. However, the flagellin pattern as detected by immunostaining or glycan staining of Western blots readily distinguished the novel group from T. denticola. Also, distinct from reference strains of T. denticola, none of the novel isolates displayed sialidase or dentilisin activities, both of which are expressed by most strains of T. denticola. Trypsin-like activity and other enzymes as detected by API ZYM test were similar to those of T. denticola. The status of a novel species is supported by the 16S rRNA gene sequence, with 98.5% similarity to its closest cultured relative, T. denticola. The name Treponema putidum sp. nov. is proposed (type strain OMZ 758T=ATCC 700334T=CIP 108088T). PMID- 15280280 TI - Mycobacterium parmense sp. nov. AB - The isolation and identification of a novel, slow-growing, scotochromogenic, mycobacterial species is reported. A strain, designated MUP 1182T, was isolated from a cervical lymph node of a 3-year-old child. MUP 1182T is alcohol- and acid fast, with a lipid pattern that is consistent with those of species that belong to the genus Mycobacterium. It grows slowly at 25-37 degrees C, but does not grow at 42 degrees C. The isolate was revealed to be biochemically distinct from previously described mycobacterial species: it has urease and Tween hydrolysis activities and lacks nitrate reductase, 3-day arylsulfatase and beta-glucosidase activities. Comparative 16S rDNA sequencing showed that isolate MUP 1182T represents a novel, slow-growing species that is related closely to Mycobacterium lentiflavum and Mycobacterium simiae. On the basis of these findings, the name Mycobacterium parmense sp. nov. is proposed, with MUP 1182T (=CIP 107385T=DSM 44553T) as the type strain. PMID- 15280282 TI - Polyphasic taxonomic analysis of Bifidobacterium animalis and Bifidobacterium lactis reveals relatedness at the subspecies level: reclassification of Bifidobacterium animalis as Bifidobacterium animalis subsp. animalis subsp. nov. and Bifidobacterium lactis as Bifidobacterium animalis subsp. lactis subsp. nov. AB - The taxonomic standing of Bifidobacterium lactis and Bifidobacterium animalis was investigated using a polyphasic approach. Sixteen representatives of both taxa were found to be phenotypically similar and shared more than 70% DNA-DNA relatedness (76-100%), which reinforces the conclusions of previous studies in which B. lactis and B. animalis were considered to be one single species. However, the results of protein profiling, BOX-PCR fingerprinting, Fluorescent Amplified Fragment Length Polymorphism (FAFLP), and atpD and groEL gene sequence analysis demonstrate that representatives of B. animalis and B. lactis constitute two clearly separated subgroups; this subdivision was also phenotypically supported based on the ability to grow in milk. Given the fact that B. lactis Meile et al. 1997 has to be considered as a junior synonym of B. animalis (Mitsuoka 1969) Scardovi and Trovatelli 1974, our data indicate that the latter species should be split into two new subspecies, i.e. Bifidobacterium animalis subsp. animalis subsp. nov. (type strain R101-8T=LMG 10508T=ATCC 25527T=DSM 20104T=JCM 1190T) and Bifidobacterium animalis subsp. lactis subsp. nov. (type strain UR1T=LMG 18314T=DSM 10140T=JCM 10602T). PMID- 15280281 TI - Oceanicola granulosus gen. nov., sp. nov. and Oceanicola batsensis sp. nov., poly beta-hydroxybutyrate-producing marine bacteria in the order 'Rhodobacterales'. AB - Three Gram-negative, chemoheterotrophic, non-motile, rod-shaped bacterial strains that accumulate poly-beta-hydroxybutyrate granules were isolated from the Bermuda Atlantic Time-series Study site by high-throughput culturing methods and characterized by polyphasic approaches. DNA-DNA hybridization, DNA G+C content and phylogenetic analyses based on 16S rRNA gene sequences divided the three isolates into two distinct genospecies that were clearly differentiated by fatty acid profiles, carbon source utilization patterns, antibiotic susceptibility and biochemical characteristics. The strains utilized a wide range of substrates, including pentoses, hexoses, oligosaccharides, sugar alcohols, organic acids and amino acids. DNA G+C contents were 71.5, 70.9 and 67.3 mol% for strains HTCC2516T, HTCC2523 and HTCC2597T, respectively. The most dominant fatty acid was 18 : 1omega7c in strains HTCC2516T and HTCC2523, and cyclo 19 : 0 in strain HTCC2597T. The type strains HTCC2516T and HTCC2597T were clearly differentiated by the presence or absence of 12 : 0, 12 : 1omega11c, 14 : 0, 15 : 0 and methyl 18 : 1. Phylogenetic analyses indicated that the strains formed a distinct monophyletic lineage within the Roseobacter clade in the order 'Rhodobacterales' of the Alphaproteobacteria, and which did not associate with any of the described genera. Genotypic and phenotypic differences of the isolates from the previously described genera support the description of Oceanicola granulosus gen. nov., sp. nov. with the type strain HTCC2516T (=ATCC BAA-861T=DSM 15982T=KCTC 12143T) and of Oceanicola batsensis sp. nov. with the type strain HTCC2597T (=ATCC BAA 863T=DSM 15984T=KCTC 12145T). PMID- 15280283 TI - Reclassification of Amycolatopsis mediterranei DSM 46095 as Amycolatopsis rifamycinica sp. nov. AB - Previous experiments have suggested that the rifamycin-producing strain DSM 46095 might not belong to Amycolatopsis mediterranei. Analysis of its 16S rRNA gene sequence and construction of a phylogenetic tree showed most similarity to Amycolatopsis kentuckyensis NRRL B-24129T, Amycolatopsis lexingtonensis NRRL B 24129T and Amycolatopsis pretoriensis NRRL B-24133T, but the strain was probably not a member of any of these species. Results from DNA-DNA hybridization experiments and comparison of DNA profiling patterns using pulsed-field gel electrophoresis also supported the assignment of strain DSM 46095 to a novel species. Analyses of phospholipids, fatty acid methyl esters and physiological characteristics also showed that the differences between different isolates of A. mediterranei and A. mediterranei DSM 46095 were as large as those between Amycolatopsis species. Strain DSM 46095 represents a novel species of the genus Amycolatopsis for which the name Amycolatopsis rifamycinica sp. nov. is proposed, with the type strain NT 19T (=DSM 46095T=ATCC 27643T). PMID- 15280284 TI - Identification of the bacterial endosymbionts of the marine ciliate Euplotes magnicirratus (Ciliophora, Hypotrichia) and proposal of 'Candidatus Devosia euplotis'. AB - This paper reports the identification of bacterial endosymbionts that inhabit the cytoplasm of the marine ciliated protozoon Euplotes magnicirratus. Ultrastructural and full-cycle rRNA approaches were used to reveal the identity of these bacteria. Based on analysis of 16S rRNA gene sequences, evolutionary trees were constructed; these placed the endosymbiont in the genus Devosia in the alpha-Proteobacteria. The validity of this finding was also shown by fluorescence in situ hybridization with a Devosia-specific oligonucleotide probe. Differences at the 16S rRNA gene level (which allowed the construction of a species-specific oligonucleotide probe) and the peculiar habitat indicate that the endosymbiont represents a novel species. As its cultivation has not been successful to date, the provisional name 'Candidatus Devosia euplotis' is proposed. The species- and group-specific probes designed in this study could represent convenient tools for the detection of 'Candidatus Devosia euplotis' and Devosia-like bacteria in the environment. PMID- 15280285 TI - Alteromonas stellipolaris sp. nov., a novel, budding, prosthecate bacterium from Antarctic seas, and emended description of the genus Alteromonas. AB - Seven novel, cold-adapted, strictly aerobic, facultatively oligotrophic strains, isolated from Antarctic sea water, were investigated by using a polyphasic taxonomic approach. The isolates were Gram-negative, chemoheterotrophic, motile, rod-shaped cells that were psychrotolerant and moderately halophilic. Buds were produced on mother and daughter cells and on prosthecae. Prostheca formation was peritrichous and prosthecae could be branched. Phylogenetic analysis based on 16S rRNA gene sequences indicated that these strains belong to the gamma Proteobacteria and are related to the genus Alteromonas, with 98.3% sequence similarity to Alteromonas macleodii and 98.0% to Alteromonas marina, their nearest phylogenetic neighbours. Whole-cell fatty acid profiles of the isolates were very similar and included C(16 : 0), C(16 : 1)omega7c, C(17 : 1)omega8c and C(18 : 1)omega8c as the major fatty acid components. These results support the affiliation of these isolates to the genus Alteromonas. DNA-DNA hybridization results and differences in phenotypic characteristics show that the strains represent a novel species with a DNA G+C content of 43-45 mol%. The name Alteromonas stellipolaris sp. nov. is proposed for this novel species; the type strain is ANT 69aT (=LMG 21861T=DSM 15691T). An emended description of the genus Alteromonas is given. PMID- 15280286 TI - Burkholderia unamae sp. nov., an N2-fixing rhizospheric and endophytic species. AB - It was shown recently that the genus Burkholderia is rich in N2-fixing bacteria that are associated with plants. A group of these diazotrophic isolates with identical or very similar 16S rDNA restriction patterns [designated amplified rDNA restriction analysis (ARDRA) genotypes 13, 14 and 15] was selected and a polyphasic taxonomic study was performed, which included new isolates that were recovered from rhizospheres, rhizoplanes or internal tissues of maize, sugarcane and coffee plants. Morphological, physiological and biochemical features, as well as multi-locus enzyme electrophoresis profiles and whole-cell protein patterns, of 20 strains were analysed. In addition, analysis of cellular fatty acid profiles, 16S rDNA sequence analysis and DNA-DNA reassociation experiments were performed with representative strains. The taxonomic data indicated that the strains analysed belong to a novel diazotrophic Burkholderia species, for which the name Burkholderia unamae sp. nov. is proposed. Strain MTl-641T (=ATCC BAA 744T=CIP 107921T), isolated from the rhizosphere of maize, was designated as the type strain. B. unamae was found as an endophyte of plants grown in regions with climates ranging from semi-hot subhumid to hot humid, but not from plants grown in regions with semi-hot or hot dry climates. Moreover, B. unamae was isolated from rhizospheres and plants growing in soils with pH values in the range 4.5 7.1, but not from soils with pH values higher than 7.5. PMID- 15280287 TI - Arenibacter certesii sp. nov., a novel marine bacterium isolated from the green alga Ulva fenestrata. AB - The taxonomic position of a novel, marine, heterotrophic, aerobic, pigmented, non motile bacterium that was isolated from a green alga, Ulva fenestrata, inhabiting the Sea of Japan, was determined. 16S rRNA gene sequence analysis revealed that the strain, KMM 3941T, is a member of the genus Arenibacter. The results of DNA DNA hybridization experiments, supported by phenotypic and chemotaxonomic data, showed that the isolate represents a novel species of the genus Arenibacter, for which the name Arenibacter certesii sp. nov. is proposed. The type strain is KMM 3941T (=KCTC 12113T=CCUG 48006T). PMID- 15280288 TI - Oceanibulbus indolifex gen. nov., sp. nov., a North Sea alphaproteobacterium that produces bioactive metabolites. AB - A water sample from the North Sea was used to isolate the abundant heterotrophic bacteria that are able to grow on complex marine media. Isolation was by serial dilution and spread plating. Phylogenetic analysis of nearly complete 16S rRNA gene sequences revealed that one of the strains, HEL-45T, had 97.4% sequence similarity to Sulfitobacter mediterraneus and 96.5 % sequence similarity to Staleya guttiformis. Strain HEL-45T is a Gram-negative, non-motile rod and obligate aerobe and requires sodium and 1-7% sea salts for growth. It contains storage granules and does not produce bacteriochlorophyll. Optimal growth temperatures are 25-30 degrees C. The DNA base composition (G+C content) is 60.1 mol%. Strain HEL-45T has Q10 as the dominant respiratory quinone. The major polar lipids are phosphatidyl glycerol, diphosphatidyl glycerol, phosphatidyl choline, phosphatidyl ethanolamine and an aminolipid. The fatty acids comprise 18 : 1omega7c, 18 : 0, 16 : 1omega7c, 16 : 0, 3-OH 10 : 0, 3-OH 12 : 1 (or 3-oxo 12 : 0) and traces of an 18 : 2 fatty acid. Among the hydroxylated fatty acids only 3 OH 12 : 1 (or 3-oxo 12 : 0) appears to be amide linked, whereas 3-OH 10 : 0 appears to be ester linked. The minor fatty acid components (between 1 and 7%) allow three subgroups to be distinguished in the Sulfitobacter/Staleya clade, placing HEL-45T into a separate lineage characterized by the presence of 3-OH 12 : 1 (or 3-oxo 12 : 0) and both ester- and amide-linked 16 : 1omega7c phospholipids. HEL-45T produces indole and derivatives thereof, several cyclic dipeptides and thryptanthrin. Phylogenetic analysis of 16S rRNA gene sequences and chemotaxonomic data support the description of a new genus and species, to include Oceanibulbus indolifex gen. nov., sp. nov., with the type strain HEL-45T (=DSM 14862T=NCIMB 13983T). PMID- 15280289 TI - Swaminathania salitolerans gen. nov., sp. nov., a salt-tolerant, nitrogen-fixing and phosphate-solubilizing bacterium from wild rice (Porteresia coarctata Tateoka). AB - A novel species, Swaminathania salitolerans gen. nov., sp. nov., was isolated from the rhizosphere, roots and stems of salt-tolerant, mangrove-associated wild rice (Porteresia coarctata Tateoka) using nitrogen-free, semi-solid LGI medium at pH 5.5. Strains were Gram-negative, rod-shaped and motile with peritrichous flagella. The strains grew well in the presence of 0.35% acetic acid, 3% NaCl and 1% KNO3, and produced acid from l-arabinose, d-glucose, glycerol, ethanol, d mannose, d-galactose and sorbitol. They oxidized ethanol and grew well on mannitol and glutamate agar. The fatty acids 18 : 1omega7c/omega9t/omega12t and 19 : 0cyclo omega8c constituted 30.41 and 11.80% total fatty acids, respectively, whereas 13 : 1 AT 12-13 was found at 0.53%. DNA G+C content was 57.6-59.9 mol% and the major quinone was Q-10. Phylogenetic analysis based on 16S rRNA gene sequences showed that these strains were related to the genera Acidomonas, Asaia, Acetobacter, Gluconacetobacter, Gluconobacter and Kozakia in the Acetobacteraceae. Isolates were able to fix nitrogen and solubilized phosphate in the presence of NaCl. Based on overall analysis of the tests and comparison with the characteristics of members of the Acetobacteraceae, a novel genus and species is proposed for these isolates, Swaminathania salitolerans gen. nov., sp. nov. The type strain is PA51T (=LMG 21291T=MTCC 3852T). PMID- 15280290 TI - Methylobacterium populi sp. nov., a novel aerobic, pink-pigmented, facultatively methylotrophic, methane-utilizing bacterium isolated from poplar trees (Populus deltoides x nigra DN34). AB - A pink-pigmented, aerobic, facultatively methylotrophic bacterium, strain BJ001T, was isolated from internal poplar tissues (Populus deltoidesxnigra DN34) and identified as a member of the genus Methylobacterium. Phylogenetic analyses showed that strain BJ001T is related to Methylobacterium thiocyanatum, Methylobacterium extorquens, Methylobacterium zatmanii and Methylobacterium rhodesianum. However, strain BJ001T differed from these species in its carbon source utilization pattern, particularly its use of methane as the sole source of carbon and energy, an ability that is shared with only one other member of the genus, Methylobacterium organophilum. In addition, strain BJ001T is the only member of the genus Methylobacterium to be described as an endophyte of poplar trees. On the basis of its physiological, genotypic and ecological properties, the isolate is proposed as a member of a novel species of the genus Methylobacterium, Methylobacterium populi sp. nov. (type strain, BJ001T=ATCC BAA 705T=NCIMB 13946T). PMID- 15280291 TI - Alteromonas litorea sp. nov., a slightly halophilic bacterium isolated from an intertidal sediment of the Yellow Sea in Korea. AB - A Gram-negative, motile, non-spore-forming, rod-shaped bacterium, designated strain TF-22T, was isolated from an intertidal sediment in Korea. This organism grew optimally at 30-37 degrees C and in the presence of 2-5% (w/v) NaCl. It did not grow without NaCl or in the presence of more than 14% (w/v) NaCl. Strain TF 22T was characterized chemotaxonomically as having ubiquinone-8 as the predominant respiratory lipoquinone and C(16 : 0), C(16 : 1) omega7c and/or iso C(15 : 0) 2-OH and C(18 : 1) omega7c as the major fatty acids. The DNA G+C content of strain TF-22T was 46.0 mol%. Phylogenetic analyses based on 16S rDNA sequences showed that strain TF-22T falls within the gamma-subclass of the Proteobacteria and forms a coherent cluster with Alteromonas macleodii and Alteromonas marina. Levels of 16S rDNA similarity between strain TF-22T and the type strains of two Alteromonas species were in the range 98.1-98.6%. The level of DNA-DNA relatedness between strain TF-22T and the type strains of two Alteromonas species was 15.7-18.5%. Therefore, on the basis of phenotypic properties, phylogeny and genomic distinctiveness, strain TF-22T should be placed in the genus Alteromonas as a novel species, for which the name Alteromonas litorea sp. nov. is proposed. The type strain is TF-22TT (=KCCM 41775T=JCM 12188T). PMID- 15280292 TI - Hydrocarboniphaga effusa gen. nov., sp. nov., a novel member of the gamma Proteobacteria active in alkane and aromatic hydrocarbon degradation. AB - Novel alkane-degrading strains of bacteria were isolated from soil contaminated with fuel oil from a leaking underground tank in New Jersey, USA. Two phenotypically similar strains (designated AP102 and AP103T) possessed 16S rRNA sequences unique among the majority of known hydrocarbon-degrading bacteria. The 16S rRNA sequences showed a moderate but distant relationship to the genus Nevskia and a substantial similarity to strains that had previously been isolated for growth on phenol (in Japan) and on toluene (in Canada) by other researchers. The hydrocarbon-degrading strains from Japan, Canada and New Jersey showed no resemblance to the typical morphology of Nevskia but did share a striking similarity among themselves in cell morphology, in the unusual appearance of colonies on various solid media and in various physiological properties. A full taxonomic analysis was performed, including DNA-DNA hybridization and nutritional screening with 117 organic compounds as sole sources of carbon and energy. The strains are active in the degradation of important environmental pollutants, and their phenotypic, physiological, metabolic and genomic properties suggest that they are members of a novel taxon in the gamma-Proteobacteria, for which the name Hydrocarboniphaga gen. nov. is proposed, with the single species Hydrocarboniphaga effusa sp. nov. The type strain is AP103T (=ATCC BAA-332T=DSM 16095T). PMID- 15280293 TI - Tsukamurella pseudospumae sp. nov., a novel actinomycete isolated from activated sludge foam. AB - The taxonomic position of two Tsukamurella strains isolated from activated sludge foam was clarified. The organisms, isolates JC85 and N1176T, were found to have chemical and morphological properties typical of members of the genus Tsukamurella. DNA-DNA relatedness studies showed that the strains formed a distinct genomic species that was most closely related to Tsukamurella spumae. The two isolates also share a range of phenotypic properties that distinguishes them from representatives of all species of Tsukamurella with validly published names. It is evident from the data that the two organisms should be classified as a novel Tsukamurella species, Tsukamurella pseudospumae sp. nov. The type strain is N1176T (=DSM 44118T=NCIMB 13963T). PMID- 15280294 TI - Halorubrum tibetense sp. nov., a novel haloalkaliphilic archaeon from Lake Zabuye in Tibet, China. AB - A novel haloalkaliphilic archaeon, strain 8W8T, was isolated from Lake Zabuye, on the Tibetan Plateau, China. On the basis of 16S rRNA gene sequence analysis, strain 8W8T was shown to belong to the genus Halorubrum and was related to Halorubrum vacuolatum (96.7% sequence similarity), Halorubrum saccharovorum (96.0%), Halorubrum lacusprofundi (95.4%) and Halorubrum sodomense (95.3%). The phylogenetic distance from any species within the other genera of Halobacteriales was lower than 90%. The major polar lipids of strain 8W8T were C20C20 and C20C25 derivatives of phosphatidylglycerol phosphate and phosphatidylglycerol phosphate methyl ester. The results of DNA-DNA hybridization and physiological and biochemical tests allowed genotypic and phenotypic differentiation of strain 8W8T from the eight Halorubrum species with validly published names. Therefore, strain 8W8T represents a novel species, for which the name Halorubrum tibetense sp. nov. is proposed, with the type strain 8W8T (=AS 1.3239T=JCM 11889T). PMID- 15280295 TI - 'Candidatus Phytoplasma mali', 'Candidatus Phytoplasma pyri' and 'Candidatus Phytoplasma prunorum', the causal agents of apple proliferation, pear decline and European stone fruit yellows, respectively. AB - Apple proliferation (AP), pear decline (PD) and European stone fruit yellows (ESFY) are among the most economically important plant diseases that are caused by phytoplasmas. Phylogenetic analyses revealed that the 16S rDNA sequences of strains of each of these pathogens were identical or nearly identical. Differences between the three phytoplasmas ranged from 1.0 to 1.5% of nucleotide positions and were thus below the recommended threshold of 2.5% for assigning species rank to phytoplasmas under the provisional status 'Candidatus'. However, supporting data for distinguishing the AP, PD and ESFY agents at the species level were obtained by examining other molecular markers, including the 16S-23S rDNA spacer region, protein-encoding genes and randomly cloned DNA fragments. The three phytoplasmas also differed in serological comparisons and showed clear differences in vector transmission and host-range specificity. From these results, it can be concluded that the AP, PD and ESFY phytoplasmas are coherent but discrete taxa that can be distinguished at the putative species level, for which the names 'Candidatus Phytoplasma mali', 'Candidatus Phytoplasma pyri' and 'Candidatus Phytoplasma prunorum', respectively, are proposed. Strains AP15R, PD1R and ESFY-G1R were selected as reference strains. Examination of available data on the peach yellow leaf roll (PYLR) phytoplasma, which clusters with the AP, PD and ESFY agents, confirmed previous results showing that it is related most closely to the PD pathogen. The two phytoplasmas share 99.6% 16S rDNA sequence similarity. Significant differences were only observed in the sequence of a gene that encodes an immunodominant membrane protein. Until more information on this phytoplasma is available, it is proposed that the PYLR phytoplasma should be regarded as a subtype of 'Candidatus Phytoplasma pyri'. PMID- 15280296 TI - Woodsholea maritima gen. nov., sp. nov., a marine bacterium with a low diversity of polar lipids. AB - Two cauliform bacteria (CM243T and CM251) isolated by J. Poindexter from the Atlantic Ocean were characterized by 16S rRNA gene sequencing, TaqI restriction fragment length polymorphism and single-strand conformation polymorphism analyses of the internally transcribed 16S-23S rDNA spacer (ITS1) region, analysis of fatty acids from cellular lipids, mass spectrometry of polar lipids and physiological properties. The two strains showed very low diversity of polar lipids with diacyl-sulfoquinovosyl glycerols as the predominant lipids. The two bacterial strains were observed to have nearly identical 16S rRNA gene sequences and could not be differentiated by their ITS1 regions. The isolates differed from species of the genus Maricaulis by their 16S rRNA gene sequences, polar lipids and fatty acid patterns. On the basis of the genotypic analyses and estimations of phylogenetic similarities, physiological and chemotaxonomic characteristics, it is proposed that the isolates represent a new genus and species, for which the name Woodsholea maritima gen. nov., sp. nov. (type strain CM243T=VKM B-1512T=LMG 21817T) is proposed. PMID- 15280297 TI - Stenotrophomonas africana Drancourt et al. 1997 is a later synonym of Stenotrophomonas maltophilia (Hugh 1981) Palleroni and Bradbury 1993. AB - Type and reference strains of Stenotrophomonas maltophilia and Stenotrophomonas africana were compared with each other and with the type strains of other Stenotrophomonas species, using SDS-PAGE of whole-cell proteins, DNA-DNA hybridization and extensive biochemical characterization. S. maltophilia LMG 958T and S. africana LMG 22072T had very similar whole-cell-protein patterns and were also biochemically very similar. A DNA-DNA binding level of 70% between both type strains confirmed that S. africana and S. maltophilia represent the same taxon. It is concluded that S. africana is a later synonym of S. maltophilia. PMID- 15280298 TI - Anoxybacillus voinovskiensis sp. nov., a moderately thermophilic bacterium from a hot spring in Kamchatka. AB - A novel moderately thermophilic bacterium, strain TH13T, was isolated from a hot spring in Kamchatka. It was found to be a Gram-positive, facultative aerobe; the straight, non-motile rods grew at 30-64 degrees C (optimum 54 degrees C). The isolate was positive for catalase and oxidase tests and reduced nitrate to nitrite, but was negative for H2S production and growth in more than 3% NaCl (w/v). The isolate grew at pH 7-8, but not at pH values higher than 9. The DNA G+C content was 43.9 mol%. Phylogenetic analysis based on 16S rRNA gene sequencing indicated that strain TH13T was a member of the genus Anoxybacillus. DNA-DNA hybridization revealed a low relatedness (less than 30.2%) between the isolate and its close phylogenetic neighbours Anoxybacillus pushchinoensis and Anoxybacillus flavithermus. On the basis of phenotypic characteristics, phylogenetic data and DNA-DNA hybridization data, it was concluded that the isolate merited classification as a novel species, for which the name Anoxybacillus voinovskiensis sp. nov. is proposed. The type strain of this species is TH13T (=NCIMB 13956T=JCM 12111T). PMID- 15280299 TI - 'Candidatus Phytoplasma', a taxon for the wall-less, non-helical prokaryotes that colonize plant phloem and insects. AB - The trivial name 'phytoplasma' has been adopted to collectively name wall-less, non-helical prokaryotes that colonize plant phloem and insects, which were formerly known as mycoplasma-like organisms. Although phytoplasmas have not yet been cultivated in vitro, phylogenetic analyses based on various conserved genes have shown that they represent a distinct, monophyletic clade within the class Mollicutes. It is proposed here to accommodate phytoplasmas within the novel genus 'Candidatus (Ca.) Phytoplasma'. Given the diversity within 'Ca. Phytoplasma', several subtaxa are needed to accommodate organisms that share <97.5% similarity among their 16S rRNA gene sequences. This report describes the properties of 'Ca. Phytoplasma', a taxon that includes the species 'Ca. Phytoplasma aurantifolia' (the prokaryote associated with witches'-broom disease of small-fruited acid lime), 'Ca. Phytoplasma australiense' (associated with Australian grapevine yellows), 'Ca. Phytoplasma fraxini' (associated with ash yellows), 'Ca. Phytoplasma japonicum' (associated with Japanese hydrangea phyllody), 'Ca. Phytoplasma brasiliense' (associated with hibiscus witches'-broom in Brazil), 'Ca. Phytoplasma castaneae' (associated with chestnut witches'-broom in Korea), 'Ca. Phytoplasma asteris' (associated with aster yellows), 'Ca. Phytoplasma mali' (associated with apple proliferation), 'Ca. Phytoplasma phoenicium' (associated with almond lethal disease), 'Ca. Phytoplasma trifolii' (associated with clover proliferation), 'Ca. Phytoplasma cynodontis' (associated with Bermuda grass white leaf), 'Ca. Phytoplasma ziziphi' (associated with jujube witches'-broom), 'Ca. Phytoplasma oryzae' (associated with rice yellow dwarf) and six species-level taxa for which the Candidatus species designation has not yet been formally proposed (for the phytoplasmas associated with X-disease of peach, grapevine flavescence doree, Central American coconut lethal yellows, Tanzanian lethal decline of coconut, Nigerian lethal decline of coconut and loofah witches' broom, respectively). Additional species are needed to accommodate organisms that, despite their 16S rRNA gene sequence being >97.5% similar to those of other 'Ca. Phytoplasma' species, are characterized by distinctive biological, phytopathological and genetic properties. These include 'Ca. Phytoplasma pyri' (associated with pear decline), 'Ca. Phytoplasma prunorum' (associated with European stone fruit yellows), 'Ca. Phytoplasma spartii' (associated with spartium witches'-broom), 'Ca. Phytoplasma rhamni' (associated with buckthorn witches'-broom), 'Ca. Phytoplasma allocasuarinae' (associated with allocasuarina yellows), 'Ca. Phytoplasma ulmi' (associated with elm yellows) and an additional taxon for the stolbur phytoplasma. Conversely, some organisms, despite their 16S rRNA gene sequence being <97.5% similar to that of any other 'Ca. Phytoplasma' species, are not presently described as Candidatus species, due to their poor overall characterization. PMID- 15280300 TI - Algibacter lectus gen. nov., sp. nov., a novel member of the family Flavobacteriaceae isolated from green algae. AB - Three strains of the marine, gliding, pigmented, facultatively anaerobic, heterotrophic, Gram-negative bacteria were isolated from the green algae Acrosiphonia sonderi (Kutz) Kornm and Ulva fenestrata Ruprecht inhabiting the Sea of Japan. 16S rDNA sequence analysis indicated that the strains were members of the family Flavobacteriaceae, in which they occupied separate lineages. The predominant cellular fatty acids were i15 : 0, a15 : 0, i15 : 1, 15 : 0, 15 : 1omega6c, i15 : 0 3-OH and i17 : 0 3-OH. The DNA base compositions were 31-33 mol% G+C. Based on the phenotypic, genotypic, chemotaxonomic and phylogenetic analyses, the novel bacteria should be placed in a novel taxon as Algibacter lectus gen. nov., sp. nov. with type strain KMM 3902G (=KCTC 12103T=DSM 15365T). PMID- 15280301 TI - Loktanella salsilacus gen. nov., sp. nov., Loktanella fryxellensis sp. nov. and Loktanella vestfoldensis sp. nov., new members of the Rhodobacter group, isolated from microbial mats in Antarctic lakes. AB - A taxonomic study was performed on 26 strains isolated from microbial mats in Antarctic lakes of the Vestfold Hills and the McMurdo Dry Valleys. Phylogenetic analysis based on 16S rRNA gene sequences placed these strains within the Rhodobacter group of the alpha-subclass of the Proteobacteria. Sequence similarity values for the strains with their nearest phylogenetic neighbours (Jannaschia, Octadecabacter and Ketogulonicigenium) ranged between 94.0 and 95.8%. DNA-DNA hybridizations and comparison of repetitive extragenic palindromic DNA-PCR (rep-PCR) fingerprinting patterns revealed that these strains are members of three distinct species. The isolates are Gram-negative, chemoheterotrophic, non-motile rods and their DNA G+C contents range from 59.4 to 66.4 mol%. Whole cell fatty acid profiles are similar and the primary fatty acid in all the strains is 18 : 1 omega7c (74.1-87.7% of total). Genotypic results together with phenotypic characteristics allowed the differentiation of these species from related recognized species of the alpha-Proteobacteria and the strains are assigned to a new genus, Loktanella gen. nov., with three novel species: Loktanella salsilacus sp. nov. (type species), consisting of ten strains with LMG 21507T (=CIP 108322T) as type strain; Loktanella fryxellensis sp. nov., consisting of 12 strains with LMG 22007T (=CIP 108323T) as type strain; and Loktanella vestfoldensis sp. nov., consisting of four strains with LMG 22003T (=CIP 108321TT) as type strain. PMID- 15280302 TI - Bradyrhizobium betae sp. nov., isolated from roots of Beta vulgaris affected by tumour-like deformations. AB - Some varieties of sugar beet, Beta vulgaris, cultivated in northern Spain have large deformations that resemble the tumours produced by Agrobacterium species. In an attempt to isolate the agent responsible for these deformations, several endophytic slow-growing bacterial strains were isolated, the macroscopic morphology of which resembled that of Bradyrhizobium species. These strains were not able to produce tumours in Nicotiana tabacum plants and, based on phylogenetic analysis of their 16S rRNA, they are closely related to the genus Bradyrhizobium. Phenotypic and molecular characteristics of these strains revealed that they represent a species different from all Bradyrhizobium species previously described. Sequence analysis of the 16S-23S rDNA intergenic spacer region indicated that these novel strains form a homogeneous group, related to Bradyrhizobium japonicum, Bradyrhizobium liaoningense and Bradyrhizobium yuanmingense. DNA-DNA hybridization confirmed that these strains represent a novel species of the genus Bradyrhizobium, for which the name Bradyrhizobium betae sp. nov. is proposed. The type strain is PL7HG1T (=LMG 21987T=CECT 5829T). PMID- 15280303 TI - Proposal to elevate the genetic variant MAC-A, included in the Mycobacterium avium complex, to species rank as Mycobacterium chimaera sp. nov. AB - The possibility that the strains included within the Mycobacterium avium complex (MAC), but not belonging either to M. avium or to Mycobacterium intracellulare, may be members of undescribed taxa, has already been questioned by several taxonomists. A very homogeneous cluster of 12 strains characterized by identical nucleotide sequences both in the 16S rDNA and in the 16S-23S internal transcribed spacer was investigated. Similar strains, previously reported in the literature, had been assigned either to the species M. intracellulare on the basis of the 16S rDNA similarity or to the group of MAC intermediates. However, several phenotypical and epidemiological characteristics seem to distinguish these strains from all other MAC organisms. The unique mycolic acid pattern obtained by HPLC is striking as it is characterized by two clusters of peaks, instead of the three presented by all other MAC organisms. All of the strains have been isolated from humans and all but one came from the respiratory tract of elderly people. The clinical significance of these strains, ascertained for seven patients, seems to suggest an unusually high virulence. The characteristics of all the strains reported in the literature, genotypically identical to the ones described here, seem to confirm our data, without reports of isolations from animals or the environment or, among humans, from AIDS patients. Therefore, an elevation of the MAC variant was proposed and characterized here, with the name Mycobacterium chimaera sp. nov.; this increases the number of species included in the M. avium complex. The type strain is FI-01069T (=CIP 107892T=DSM 44623T). PMID- 15280304 TI - Myceligenerans xiligouense gen. nov., sp. nov., a novel hyphae-forming member of the family Promicromonosporaceae. AB - Strain XLG9A10.2T was isolated from an alkaline salt marsh soil in western China. 16S rRNA gene sequence analysis indicated that strain XLG9A10.2T constitutes a distinct lineage within the family Promicromonosporaceae, sharing 94.8-95.1% gene similarity with members of the genus Promicromonospora and 94.4-95.7% similarity with those of Xylanimonas and related genera. The general colony and cell morphology of strain XLG9A10.2T is similar to that of members of Promicromonospora, but differs from members of the genus Xylanimonas in forming a well-developed branching mycelium and production of coccoid spores. Strain XLG9A10.2T shows the peptidoglycan type A4alpha (L-lys<--L-thr<--D-Glu), contains glucose, mannose and galactose as whole cell sugars and has MK-9(H4) and MK-9(H6) as major menaquinones, while phospholipids are phosphatidylglycerol, diphosphatidylglycerol, phosphatidylinositol, three unidentified phospholipids and one unidentified glycolipid. The DNA base composition is 71.9 mol% G+C. On the basis of morphological, chemotaxonomic, metabolic and phylogenetic differences from other species of Promicromonosporaceae, a new genus and species, Myceligenerans xiligouense gen. nov., sp. nov., is proposed. The type strain is XLG9A10.2T (=DSM 15700T=CGMCC 1.3458T.) PMID- 15280305 TI - Nocardioides ganghwensis sp. nov., isolated from tidal flat sediment. AB - A strictly aerobic, non-motile, rod-shaped actinomycete strain, named JC2055T, was isolated from a sediment sample of getbol, the tidal flat of Korea. 16S rDNA analysis revealed that the getbol isolate belonged to the genus Nocardioides with the highest sequence similarity to Nocardioides aquiterrae GW-9T (95.5%). The major menaquinone was MK-8(H4) and the predominant cellular fatty acids were iso 16 : 0 and 17 : 1omega8c. Tuberculostearic acid was absent. The G+C content of DNA was 72 mol%. Based on the morphological, physiological, biochemical and chemotaxonomical data presented in this study, strain JC2055T can be readily differentiated from other validly named Nocardioides species. The name Nocardioides ganghwensis sp. nov. is proposed for the isolate. The type strain is JC2055T (=IMSNU 14028T=KCTC 9920T=JCM 12124T). PMID- 15280306 TI - Taxonomy of Australian clinical isolates of the genus Photorhabdus and proposal of Photorhabdus asymbiotica subsp. asymbiotica subsp. nov. and P. asymbiotica subsp. australis subsp. nov. AB - The relationship of Photorhabdus isolates that were cultured from human clinical specimens in Australia to Photorhabdus asymbiotica isolates from human clinical specimens in the USA and to species of the genus Photorhabdus that are associated symbiotically with entomopathogenic nematodes was evaluated. A polyphasic approach that involved DNA-DNA hybridization, phylogenetic analyses of 16S rRNA and gyrB gene sequences and phenotypic characterization was adopted. These investigations showed that gyrB gene sequence data correlated well with DNA-DNA hybridization and phenotypic data, but that 16S rRNA gene sequence data were not suitable for defining species within the genus Photorhabdus. Australian clinical isolates proved to be related most closely to clinical isolates from the USA, but the two groups were distinct. A novel subspecies, Photorhabdus asymbiotica subsp. australis subsp. nov. (type strain, 9802892T=CIP 108025T=ACM 5210T), is proposed, with the concomitant creation of Photorhabdus asymbiotica subsp. asymbiotica subsp. nov. Analysis of gyrB sequences, coupled with previously published data on DNA-DNA hybridization and PCR-RFLP analysis of the 16S rRNA gene, indicated that there are more than the three subspecies of Photorhabdus luminescens that have been described and confirmed the validity of the previously proposed subdivision of Photorhabdus temperata. Although a non-luminescent, symbiotic isolate clustered consistently with P. asymbiotica in gyrB phylogenetic analyses, DNA-DNA hybridization indicated that this isolate does not belong to the species P. asymbiotica and that there is a clear distinction between symbiotic and clinical species of Photorhabdus. PMID- 15280307 TI - Veillonella montpellierensis sp. nov., a novel, anaerobic, Gram-negative coccus isolated from human clinical samples. AB - Three strains of a hitherto unknown, Gram-negative, anaerobic coccus were isolated from human samples. At the phenotypic level, the isolates displayed all the characteristics of bacteria belonging to the genus Veillonella. Sequence analysis revealed that the three strains shared >99.5% similarity in 16S rDNA sequence and >98.4% similarity in dnaK sequence. The three unknown strains formed a separate subclade that was clearly remote from Veillonella species of human and animal origin. Based on these results, the three strains were considered to represent a novel species within the genus Veillonella, for which the name Veillonella montpellierensis is proposed. The type strain of the species is ADV 281.99T (=CIP 107992T=CCUG 48299T). PMID- 15280308 TI - Marinibacillus campisalis sp. nov., a moderate halophile isolated from a marine solar saltern in Korea, with emended description of the genus Marinibacillus. AB - A Gram-positive, motile, round to ellipsoidal, endospore-forming, rod-shaped bacterial strain, SF-57T, was isolated from a marine solar saltern in Korea. This organism grew between 4 and 39 degrees C, with optimum growth at 30 degrees C. Strain SF-57T grew in the presence of 0.5-15.0% NaCl, with optimum growth at 2-3% NaCl. The peptidoglycan type of strain SF-57T was A1alpha linked directly through l-Lys. In strain SF-57TT, menaquinone-7 (MK-7) was the predominant isoprenoid quinone and anteiso-C(15 : 0) was the major fatty acid. The DNA G+C content was 41.8 mol%. Phylogenetic analyses based on 16S rRNA gene sequences showed that strain SF-57T formed a coherent cluster with Marinibacillus marinus, with a bootstrap resampling value of 100%. The level of 16S rRNA gene sequence similarity between strain SF-57T and M. marinus DSM 1297T was 98.9%. The mean DNA DNA relatedness level between strain SF-57T and the type strain of M. marinus was 20.6%. Based on phenotypic properties, phylogenetic analyses and genomic data, strain SF-57T merits placement in the genus Marinibacillus as a representative of a novel species, for which the name Marinibacillus campisalis sp. nov. is proposed. The type strain is SF-57T (=KCCM 41644T=JCM 11810T). PMID- 15280309 TI - Streptomyces drozdowiczii sp. nov., a novel cellulolytic streptomycete from soil in Brazil. AB - An actinomycete strain, isolated from a Mata Atlantica soil sample, showing cellulolytic activity was subjected to polyphasic taxonomic characterization to determine its identity. Strain M7aT presented morphological and chemotaxonomic characteristics consistent with its assignment to the genus Streptomyces. Phylogenetic analysis of its 16S rDNA sequence revealed that the strain differed from described streptomycetes available in the public databases; the most closely related species was Streptomyces laceyi, with 98.4% nucleotide similarity. It also differed from other cellulolytic strains in its phenotypic characteristics. It is therefore proposed that strain M7aT, a cellulolytic strain with biotechnological potential, represents a novel species, named Streptomyces drozdowiczii sp. nov. The type strain is M7aT (=CBMAI 0498T=CIP 107837T=NRRL B 24297T). PMID- 15280310 TI - Halomonas anticariensis sp. nov., from Fuente de Piedra, a saline-wetland wildfowl reserve in Malaga, southern Spain. AB - Three Halomonas strains, FP34, FP35T and FP36, which were isolated from soil samples taken from Fuente de Piedra, a saline wetland in the province of Malaga in southern Spain, are described. Phylogenetic analyses based on 16S rRNA gene sequences show that the three isolates belong to the genus Halomonas in the gamma Proteobacteria and form an independent genetic line. Phenotypically, they share the characteristics of Halomonas and differ from the most closely related species, Halomonas campisalis, in the following features: they are strictly aerobic and, because of their production of exopolysaccharides, form cream coloured, mucoid colonies; they produce phosphatase and grow within narrow pH and temperature ranges; and they are susceptible to kanamycin and streptomycin. Their G+C content varies between 60.0 and 61.4 mol%. The name Halomonas anticariensis sp. nov. is proposed for these isolates. Strain FP35T (=LMG 22089T=CECT 5854T) is the type strain. The bacterium grows best in 7.5% (w/v) NaCl and does not require magnesium or potassium salts for growth, although they do stimulate growth somewhat when present. Its major fatty acids are 18 : 1omega7c, 16 : 0, 16 : 1omega7c, 15 : 0 iso 2-OH, 12 : 0 3-OH, 12 : 0, 10 : 0 and 19 : 0 cyclo omega8c. Its predominant respiratory lipoquinone is ubiquinone with nine isoprene units (Q 9). PMID- 15280311 TI - Comparative analysis of ribonuclease P RNA of the planctomycetes. AB - The planctomycetes, order Planctomycetales, are a distinct phylum of domain Bacteria. Genes encoding the RNA portion of ribonuclease P (RNase P) of some planctomycete members were sequenced and compared with existing database planctomycete sequences. rnpB gene sequences encoding RNase P RNA were generated by a conserved primer PCR strategy for Planctomyces brasiliensis, Planctomyces limnophilus, Pirellula marina, Pirellula staleyi strain ATCC 35122, Isosphaera pallida, one other Isosphaera strain, Gemmata obscuriglobus and three other strains of the Gemmata group. These sequences were aligned against reference bacterial sequences and secondary structures of corresponding RNase P RNAs deduced by a comparative approach. P12 helices were found to be highly variable in length, as were helices P16.1 and P19, when present. RNase P RNA secondary structures of Gemmata isolates were found to have unusual features relative to other planctomycetes, including a long P9 helix and an insert in the P13 helix not found in any other member of domain Bacteria. These unique features are consistent with other unusual properties of this genus, distinguishing it from other bacteria. Phylogenetic analyses indicate that relationships between planctomycetes derived from RNase P RNA are consistent with 16S rRNA-based analyses. PMID- 15280312 TI - Kribbella solani sp. nov. and Kribbella jejuensis sp. nov., isolated from potato tuber and soil in Jeju, Korea. AB - Two actinomycete strains, DSA1T and HD9T, were isolated from a potato tuber and soil from a potato-cultivating field in Jeju, Korea, respectively. A comprehensive 16S rDNA analysis revealed that the isolates belong to the genus Kribbella and share 97.7-98.6% sequence similarity to Kribbella species. The strains also contained typical chemotaxonomic markers of the genus Kribbella: ll diaminopimelic acid, alanine, glycine and glutamic acid in the cell wall peptidoglycan; mannose, glucose, galactose and ribose as whole cell sugars; diphosphatidylglycerol, phosphatidylcholine and phosphatidylinositol as characteristic phospholipids; and the major menaquinone MK-9 (H4). DNA-DNA hybridization experiments showed that the isolates represented two distinct genomic species. A number of phenotypic properties can be used to differentiate the two isolates from Kribbella species. On the basis of polyphasic evidence, two novel species are proposed: Kribbella solani sp. nov. for strain DSA1T (=KACC 20196T=JCM 12205T) and Kribbella jejuensis sp. nov. for strain HD9T (=KACC 20266T=JCM 12204T). PMID- 15280313 TI - Clover proliferation phytoplasma: 'Candidatus Phytoplasma trifolii'. AB - Clover proliferation phytoplasma (CPR) is designated as the reference strain for the CP phylogenetic group or subclade, on the basis of molecular analyses of genomic DNA, the 16S rRNA gene and the 16S-23S spacer region. Other strains related to CPR include alfalfa witches'-broom (AWB), brinjal little leaf (BLL), beet leafhopper-transmitted virescence (BLTV), Illinois elm yellows (ILEY), potato witches'-broom (PWB), potato yellows (PY), tomato big bud in California (TBBc) and phytoplasmas from Fragaria multicipita (FM). Phylogenetic analysis of the 16S rRNA gene sequences of BLL, CPR, FM and ILEY, together with sequences from 16 other phytoplasmas that belong to the ash yellows (AshY), jujube witches' broom (JWB) and elm yellows (EY) groups that were available in GenBank, produced a tree on which these phytoplasmas clearly clustered as a discrete group. Three subgroups have been classified on the basis of sequence homology and the collective RFLP patterns of amplified 16S rRNA genes. AWB, BLTV, PWB and TBBc are assigned to taxonomic subgroup CP-A, FM belongs to subgroup CP-B and BLL and ILEY are assigned to subgroup CP-C. Genetic heterogeneity between different isolates of AWB, CPR and PWB has been observed from heteroduplex mobility assay analysis of amplified 16S rRNA genes and the 16S-23S spacer region. Two unique signature sequences that can be utilized to distinguish the CP group from others were present. On the basis of unique properties of the DNA from clover proliferation phytoplasma, the name 'Candidatus Phytoplasma trifolii' is proposed for the CP group. PMID- 15280314 TI - Bacillus farraginis sp. nov., Bacillus fortis sp. nov. and Bacillus fordii sp. nov., isolated at dairy farms. AB - Forty-eight bacterial strains were isolated at dairy farms from raw milk, the milking apparatus, green fodder or feed concentrate after a heat treatment of 30 min at 100 degrees C. In this way, spore-forming bacteria with a very high intrinsic heat resistance were selected for. The aerobic spore-forming isolates were subjected to a polyphasic taxonomical study, including repetitive element sequence-based PCR typing, whole-cell protein profiling, 16S rDNA sequence analysis, DNA-DNA hybridizations, DNA base composition, fatty acid analysis, and morphological and biochemical characteristics. A comparison of the REP- and (GTG)5-PCR and whole-cell protein SDS-PAGE profiles resulted in three clusters of similar strains. Analysis of the 16S rDNA sequences and DNA-DNA relatedness data showed that these clusters represented three novel species. The highest 16S rDNA similarity to a recognized species found for the three groups was around 94% with Bacillus lentus and Bacillus sporothermodurans. Further phenotypic characterization supported the proposal of three novel species in the genus Bacillus, Bacillus farraginis, Bacillus fortis and Bacillus fordii. The respective type strains are R-6540T (=LMG 22081T=DSM 16013T), R-6514T (=LMG 22079T=DSM 16012T) and R-7190T (=LMG 22080T=DSM 16014T); their G+C DNA base contents are 43.7, 44.3 and 41.9 mol%, respectively. Although in variable amounts, a predominance of the branched fatty acids iso-C(15 : 0) and anteiso C(15 : 0) was observed in all three novel species. PMID- 15280315 TI - Lactobacillus saerimneri sp. nov., isolated from pig faeces. AB - In studying the composition of the Lactobacillus flora of faeces from pigs fed different diets, isolates with notable differences in their 16S rRNA gene sequence compared to recognized species were found. Phenotypic characteristics together with 16S rRNA gene sequences revealed that the isolates represented a novel species belonging to the Lactobacillus mali subgroup of lactobacilli. The name Lactobacillus saerimneri sp. nov. is proposed (type strain GDA154T=LMG 22087T=DSM 16049T=CCUG 48462T). PMID- 15280316 TI - Bacillus indicus sp. nov., an arsenic-resistant bacterium isolated from an aquifer in West Bengal, India. AB - Strain Sd/3T (=MTCC 4374T=DSM 15820T), an arsenic-resistant bacterium, was isolated from a sand sample obtained from an arsenic-contaminated aquifer in Chakdah district in West Bengal, India (23 degrees 3' N 88 degrees 35' E). The bacterium was Gram-positive, rod-shaped, non-motile, endospore-forming and yellowish-orange pigmented. It possessed all the characteristics that conform to the genus Bacillus, such as it had A4beta murein type (L-orn-D-Asp) peptidoglycan variant, MK-7 as the major menaquinone and iso-C15 : 0 and anteiso-C15 : 0 as the major fatty acids. Based on its chemotaxonomic and phylogenetic characteristics, strain Sd/3T was identified as a species of the genus Bacillus. It exhibited maximum similarity (95%) at the 16S rRNA gene level with Bacillus cohnii; however, DNA-DNA similarity with B. cohnii was 60.7%. Strain Sd/3T also exhibited a number of phenotypic differences from B. cohnii (DSM 6307T). These data suggest that Sd/3T represents a novel species of the genus Bacillus. The name Bacillus indicus sp. nov. is proposed. PMID- 15280317 TI - Saccharothrix algeriensis sp. nov., isolated from Saharan soil. AB - The taxonomic position of a soil isolate, strain SA 233T, recovered from Saharan soil from Algeria was established using a polyphasic approach. This isolate has been previously reported to produce three novel dithiolopyrrolone antibiotics, and preliminary chemotaxonomic and morphological characteristics suggested that it was representative of a member of the genus Saccharothrix. Phylogenetic analysis of the strain from 16S rDNA sequences, along with a detailed analysis of morphological, chemotaxonomic and physiological characteristics, indicates that it belongs to the genus Saccharothrix and represents a novel species that is readily distinguished from all recognized Saccharothrix species. The name Saccharothrix algeriensis sp. nov. is proposed for the isolate, with type strain SA 233T (=NRRL B-24137T=DSM 44581T). PMID- 15280318 TI - Reclassification of Promicromonospora pachnodae Cazemier et al. 2004 as Xylanimicrobium pachnodae gen. nov., comb. nov. AB - The recently described facultatively anaerobic Promicromonospora pachnodae is phylogenetically only moderately related to authentic members of Promicromonospora. P. pachnodae is closely related to Xylanibacterium ulmi and slightly less closely related to Xylanimonas cellulosilytica and Isoptericola variabilis (basonym Cellulosimicrobium variabile). Members of the different genera of Promicromonosporaceae have similar chemotaxonomic properties; they share the same peptidoglycan type (A4alpha) and have similar profiles of polar lipids, menaquinones, fatty acids and whole cell sugars. However, they differ from each other in the detailed amino acid composition of peptidoglycan, a taxonomically significant character that has previously been used in the delineation of actinobacterial genera. Recognized Promicromonospora species and Xylanibacterium ulmi exhibit the L-Lys-L-Ala-D-Glu type, Xylanimonas cellulosilytica and I. variabilis show the L-Lys-D-Asp type, whereas P. pachnodae has the L-Lys-L-Ser-D-Glu type. This property, together with the distinct phylogenetic position of Promicromonospora pachnodae, suggests a novel genus for the xylanolytic organism Xylanimicrobium pachnodae (Cazemier et al. 2004) gen. nov., comb. nov. PMID- 15280319 TI - Chitinibacter tainanensis gen. nov., sp. nov., a chitin-degrading aerobe from soil in Taiwan. AB - Five strains with strong chitinolytic activity were isolated from a soil sample collected from southern Taiwan. The strains shared more than 92% DNA-DNA similarity, indicating membership of the same genospecies. This close relationship was supported by high similarities in fatty acid composition and biochemical characteristics. A 16S rRNA gene sequence analysis indicated that the isolates were members of the class 'Betaproteobacteria', in which they formed an individual subline of descent that was distantly related (<94% similarity) to lineages defined by Formivibrio citricus DSM 6150T and Iodobacter fluviatilis DSM 3764T. On the basis of the phylogenetic and phenotypic distinctness of these novel chitin-degrading organisms, a new genus, Chitinibacter, is proposed, with Chitinibacter tainanensis (type strain, S1T=BCRC 17254T=DSM 15459T) as the type species. PMID- 15280320 TI - Phylogeny of the family Pasteurellaceae based on rpoB sequences. AB - Sequences of the gene encoding the beta-subunit of the RNA polymerase (rpoB) were used to delineate the phylogeny of the family Pasteurellaceae. A total of 72 strains, including the type strains of the major described species as well as selected field isolates, were included in the study. Selection of universal rpoB derived primers for the family allowed straightforward amplification and sequencing of a 560 bp fragment of the rpoB gene. In parallel, 16S rDNA was sequenced from all strains. The phylogenetic tree obtained with the rpoB sequences reflected the major branches of the tree obtained with the 16S rDNA, especially at the genus level. Only a few discrepancies between the trees were observed. In certain cases the rpoB phylogeny was in better agreement with DNA DNA hybridization studies than the phylogeny derived from 16S rDNA. The rpoB gene is strongly conserved within the various species of the family of Pasteurellaceae. Hence, rpoB gene sequence analysis in conjunction with 16S rDNA sequencing is a valuable tool for phylogenetic studies of the Pasteurellaceae and may also prove useful for reorganizing the current taxonomy of this bacterial family. PMID- 15280322 TI - Candida asparagi sp. nov., Candida diospyri sp. nov. and Candida qinlingensis sp. nov., novel anamorphic, ascomycetous yeast species. AB - Among ascomycetous yeasts that were isolated from several nature reserve areas in China, three anamorphic strains isolated from soil (QL 5-5T) and fruit (QL 21-2T and SN 15-1T) were revealed, by conventional characterization and molecular phylogenetic analysis based on internal transcribed spacer and large subunit (26S) rRNA gene D1/D2 region sequencing, to represent three novel species in the genus Candida. Candida qinlingensis sp. nov. (type strain, QL 5-5T=AS 2.2524T=CBS 9768T) was related closely to a teleomorphic species, Williopsis pratensis. The close relatives of Candida diospyri sp. nov. (type strain, QL 21-2T=AS 2.2525T=CBS 9769T) are Candida friedrichii and Candida membranifaciens. Candida asparagi sp. nov. (type strain, SN 15-1T=AS 2.2526T=CBS 9770T) forms a clade with Candida fructus. PMID- 15280321 TI - Curvibasidium cygneicollum gen. nov., sp. nov. and Curvibasidium pallidicorallinum sp. nov., novel taxa in the Microbotryomycetidae (Urediniomycetes), and their relationship with Rhodotorula fujisanensis and Rhodotorula nothofagi. AB - Strains of Rhodotorula fujisanensis (Basidiomycota, Urediniomycetes, Microbotryomycetidae), including the type strain, are sexually compatible and produce clamped mycelium with teliospores. However, as teliospore germination had not been documented, the complete sexual cycle was not known. During the course of this work, the basidial stage of R. fujisanensis was characterized. In addition, mating studies employing isolates that were identified preliminarily as Rhodotorula nothofagi, a species that is related closely to R. fujisanensis, yielded mycelium with teliospores, which formed basidia and basidiospores. The new data were evaluated by using several criteria, including the available molecular phylogenetic framework for the Microbotryomycetidae. Curvibasidium gen. nov. is described here, to accommodate two teleomorphs: Curvibasidium cygneicollum sp. nov. (CBS 4551T), which is described as the sexual stage of R. fujisanensis, and Curvibasidium pallidicorallinum sp. nov. (CBS 9091T), which is related closely to R. nothofagi, but does not represent its sexual stage. PMID- 15280323 TI - Recommended standards for the description of new species of anoxygenic phototrophic bacteria. AB - Recommended standards for the description of new species of the anoxygenic phototrophic bacteria are proposed in accordance with Recommendation 30b of the International Code of Nomenclature of Bacteria. These standards include information on the natural habitat, ecology and phenotypic properties including morphology, physiology and pigments and on genetic information and nucleic acid data. The recommended standards were supported by the Subcommittee on the taxonomy of phototrophic bacteria of the International Committee on Systematics of Prokaryotes. They are considered as guidelines for authors to prepare descriptions of new species. PMID- 15280324 TI - "Promiscuous" anticancer drugs that hit multiple targets may thwart resistance. PMID- 15280325 TI - Aging prisoners stressing health care system. PMID- 15280326 TI - When teens self-treat headaches, OTC drug misuse is frequent result. PMID- 15280327 TI - Acrylamide-food link questions linger. PMID- 15280335 TI - Virtual colonoscopy. PMID- 15280336 TI - Virtual colonoscopy. PMID- 15280337 TI - Virtual colonoscopy. PMID- 15280338 TI - Virtual colonoscopy. PMID- 15280339 TI - Radiation therapy and risk of cardiovascular disease. PMID- 15280340 TI - Radiation therapy and risk of cardiovascular disease. PMID- 15280341 TI - Effect of a decision aid on knowledge and treatment decision making for breast cancer surgery: a randomized trial. AB - CONTEXT: The long-term results of randomized trials have demonstrated equivalent survival rates for mastectomy and breast-conserving therapy for the treatment of early stage breast cancer. Consequently, the choice of treatment should be based on a patient's preferences. OBJECTIVE: To evaluate the impact of a decision aid regarding the different surgical treatment options on patient decision making. DESIGN AND SETTING: A cluster randomized trial for which general surgeons in the communities of central-west, and eastern Ontario, Canada, were randomly assigned to use the decision aid or not in the surgical consultation. Patients received the decision aid or not based on the surgeon seen. PARTICIPANTS: Twenty surgeons participated in the study. Of the 208 eligible women with newly diagnosed clinical stage I or II breast cancer seen by study surgeons, 201 agreed to be evaluated: 94 were assigned to the decision board and 107 to usual practice. Patients were recruited from November 1999 to April 2002. INTERVENTION: The decision board is a decision aid designed to help physicians inform their patients about different treatment options and to enable patients to express a preference for treatment. MAIN OUTCOME MEASURES: Patient knowledge about the surgical treatment of breast cancer; decisional conflict; satisfaction with decision making; and the treatment decision following the consultation. RESULTS: Patients in the decision board group had higher knowledge scores about their treatment options (66.9 vs 58.7; P<.001), had less decisional conflict (1.40 vs 1.62, P =.02), and were more satisfied with decision making (4.50 vs 4.32, P =.05) following the consultation. Patients who used the decision board were more likely to choose BCT (94% vs 76%, P =.03). CONCLUSIONS: The decision board was helpful in improving communication and enabling women to make a choice regarding treatment. Such instruments should be considered by surgeons when communicating the different surgical options to women with breast cancer. PMID- 15280342 TI - Effect of a computer-based decision aid on knowledge, perceptions, and intentions about genetic testing for breast cancer susceptibility: a randomized controlled trial. AB - CONTEXT: As the availability of and demand for genetic testing for hereditary cancers increases in primary care and other clinical settings, alternative or adjunct educational methods to traditional genetic counseling will be needed. OBJECTIVE: To compare the effectiveness of a computer-based decision aid with standard genetic counseling for educating women about BRCA1 and BRCA2 genetic testing. DESIGN: Randomized controlled trial conducted from May 2000 to September 2002. SETTING AND PARTICIPANTS: Outpatient clinics offering cancer genetic counseling at 6 US medical centers enrolled 211 women with personal or family histories of breast cancer. INTERVENTIONS: Standard one-on-one genetic counseling (n = 105) or education by a computer program followed by genetic counseling (n = 106). MAIN OUTCOME MEASURES: Participants' knowledge, risk perception, intention to undergo genetic testing, decisional conflict, satisfaction with decision, anxiety, and satisfaction with the intervention. Counselor group measures were administered at baseline and after counseling. Computer group measures were administered at baseline, after computer use, and after counseling. Testing decisions were assessed at 1 and 6 months. Outcomes were analyzed by high vs low risk of carrying a BRCA1 or BRCA2 mutation. RESULTS: Both groups had comparable demographics, prior computer experience, medical literacy, and baseline knowledge of breast cancer and genetic testing, and both counseling and computer use were rated highly. Knowledge scores increased in both groups (P<.001) regardless of risk status, and change in knowledge was greater in the computer group compared with the counselor group (P =.03) among women at low risk of carrying a mutation. Perception of absolute risk of breast cancer decreased significantly after either intervention among all participants. Intention to undergo testing decreased significantly after either intervention among low-risk but not high-risk women. The counselor group had lower mean scores on a decisional conflict scale (P =.04) and, in low-risk women, higher mean scores on a satisfaction-with-decision scale (P =.001). Mean state anxiety scores were reduced by counseling but were within normal ranges for both groups at baseline and after either intervention, regardless of risk status. CONCLUSIONS: An interactive computer program was more effective than standard genetic counseling for increasing knowledge of breast cancer and genetic testing among women at low risk of carrying a BRCA1 or BRCA2 mutation. However, genetic counseling was more effective than the computer at reducing women's anxiety and facilitating more accurate risk perceptions. These results suggest that this computer program has the potential to stand alone as an educational intervention for low-risk women but should be used as a supplement to genetic counseling for those at high risk. PMID- 15280343 TI - Functional decline in peripheral arterial disease: associations with the ankle brachial index and leg symptoms. AB - CONTEXT: Among individuals with lower-extremity peripheral arterial disease (PAD), specific leg symptoms and the ankle brachial index (ABI) are cross sectionally related to the degree of functional impairment. However, relations between these clinical characteristics and objectively measured functional decline are unknown. OBJECTIVE: To define whether PAD, ABI, and specific leg symptoms predict functional decline at 2-year follow-up. DESIGN, SETTING, AND PARTICIPANTS: Prospective cohort study among 676 consecutively identified individuals (aged > or =55 years) with and without PAD (n = 417 and n = 259, respectively), with baseline functional assessments occurring between October 1, 1998, and January 31, 2000, and follow-up assessments scheduled 1 and 2 years thereafter. PAD was defined as ABI less than 0.90, and participants with PAD were categorized at baseline into 1 of 5 mutually exclusive symptom groups. MAIN OUTCOME MEASURES: Mean annual changes in 6-minute walk performance and in usual paced and fast-paced 4-m walking velocity, adjusted for age, sex, race, prior year functioning, comorbid diseases, body mass index, pack-years of cigarette smoking, and patterns of missing data. RESULTS: Lower baseline ABI values were associated with greater mean (95% confidence interval) annual decline in 6-minute walk performance (-73.0 [-142 to -4.2] ft for ABI <0.50 vs -58.8 [-83.5 to -34.0] ft for ABI 0.50 to <0.90 vs -12.6 [-40.3 to 15.1] ft for ABI 0.90-1.50, P =.02). Compared with participants without PAD, PAD participants with leg pain on exertion and rest at baseline had greater mean annual decline in 6-minute walk performance (-111 [-173 to -50.0] ft vs -8.67 [-36.9 to 19.5] ft, P =.004), usual pace 4-meter walking velocity (-0.06 [-0.09 to -0.02] m/sec vs -0.01 (-0.03 to 0.003] m/sec, P =.02), and fastest-pace 4-meter walking velocity (-0.07 [-0.11 to -0.03] m/sec vs -0.02 [-0.04 to -0.006] m/sec, P =.046). Compared with participants without PAD, asymptomatic PAD was associated with greater mean annual decline in 6-minute walk performance (-76.8 (-135 to -18.6] ft vs -8.67 ( 36.9 to 19.5] ft, P =.04) and an increased odds ratio for becoming unable to walk for 6 minutes continuously (3.63; 95% confidence interval, 1.58-8.36; P =.002). CONCLUSIONS: Baseline ABI and the nature of leg symptoms predict the degree of functional decline at 2-year follow-up. Previously reported lack of worsening in claudication symptoms over time in patients with PAD may be more related to declining functional performance to than lack of disease progression. PMID- 15280344 TI - Diagnosis of intra-amniotic infection by proteomic profiling and identification of novel biomarkers. AB - CONTEXT: Intra-amniotic infection (IAI) is commonly associated with preterm birth and adverse neonatal sequelae. Early diagnosis of IAI, however, has been hindered by insensitive or nonspecific tests. OBJECTIVE: To identify unique protein signatures in rhesus monkeys with experimental IAI, a proteomics-based analysis of amniotic fluid was used to develop diagnostic biomarkers for subclinical IAI in amniotic fluid and blood of women with preterm labor. DESIGN, SETTING, AND PARTICIPANTS: Surface-enhanced laser desorption-ionization/time-of-flight mass spectrometry, gel electrophoresis, and tandem mass spectrometry were used to characterize amniotic fluid peptides in 19 chronically instrumented pregnant rhesus monkeys before and after experimental IAI. Candidate biomarkers were determined by liquid chromatography-tandem mass spectrometry. Polyclonal antibodies were generated from synthetic peptides for validation of biomarkers of IAI. Amniotic fluid peptide profiles identified in experimental IAI were subsequently tested in a cohort of 33 women admitted to Seattle, Wash, hospitals between June 25, 1991, and June 30, 1997, with preterm delivery at 35 weeks or earlier associated with subclinical IAI (n = 11), preterm delivery at 35 weeks or earlier without IAI (n = 11), and preterm contractions with subsequent term delivery at later than 35 weeks (n = 11). MAIN OUTCOME MEASURES: Identification of peptide biomarkers for occult IAI. RESULTS: Protein expression profiles in amniotic fluid showed unique signatures of overexpression of polypeptides in the 3- to 5-kDa and 10- to 12-kDa molecular weight ranges in all animals after infection and in no animal prior to infection. In women, the 10- to 12-kDa signature was identified in all 11 patients with subclinical IAI, in 2 of 11 with preterm delivery without IAI, and in 0 of 11 with preterm labor and term delivery without infection (P<.001). Peptide fragment analysis of the diagnostic peak in amniotic fluid identified calgranulin B and a unique fragment of insulinlike growth factor binding protein 1, which were also expressed in maternal serum. Mapping of other amniotic fluid proteins differentially expressed in IAI identified several immunoregulators not previously described in amniotic fluid. CONCLUSIONS: This proteomics-based characterization of the differential expression of amniotic fluid proteins in IAI identified a distinct proteomic profile in an experimental primate chorioamnionitis model that detected subclinical IAI in a human cohort with preterm labor. These diagnostic protein expression signatures, complemented by immunodetection of specific biomarkers in amniotic fluid and in maternal serum, might have application in the early detection of IAI. PMID- 15280345 TI - Benefits of adding a drug to a single-agent or a 2-agent chemotherapy regimen in advanced non-small-cell lung cancer: a meta-analysis. AB - CONTEXT: Randomized trials have demonstrated that adding a drug to a single-agent or to a 2-agent regimen increased the tumor response rate in patients with advanced non-small-cell lung cancer (NSCLC), although its impact on survival remains controversial. OBJECTIVE: To evaluate the clinical benefit of adding a drug to a single-agent or 2-agent chemotherapy regimen in terms of tumor response rate, survival, and toxicity in patients with advanced NSCLC. DATA SOURCES AND STUDY SELECTION: Data from all randomized controlled trials performed between 1980 and 2001 (published between January 1980 and October 2003) comparing a doublet regimen with a single-agent regimen or comparing a triplet regimen with a doublet regimen in patients with advanced NSCLC. There were no language restrictions. Searches of MEDLINE and EMBASE were performed using the search terms non-small-cell lung carcinoma/drug therapy, adenocarcinoma, large-cell carcinoma, squamous-cell carcinoma, lung, neoplasms, clinical trial phase III, and randomized trial. Manual searches were also performed to find conference proceedings published between January 1982 and October 2003. DATA EXTRACTION: Two independent investigators reviewed the publications and extracted the data. Pooled odds ratios (ORs) for the objective tumor response rate, 1-year survival rate, and toxicity rate were calculated using the fixed-effect model. Pooled median ratios (MRs) for median survival also were calculated using the fixed effect model. ORs and MRs lower than unity (<1.0) indicate a benefit of a doublet regimen compared with a single-agent regimen (or a triplet regimen compared with a doublet regimen). DATA SYNTHESIS: Sixty-five trials (13 601 patients) were eligible. In the trials comparing a doublet regimen with a single-agent regimen, a significant increase was observed in tumor response (OR, 0.42; 95% confidence interval [CI], 0.37-0.47; P<.001) and 1-year survival (OR, 0.80; 95% CI, 0.70 0.91; P<.001) in favor of the doublet regimen. The median survival ratio was 0.83 (95% CI, 0.79-0.89; P<.001). An increase also was observed in the tumor response rate (OR, 0.66; 95% CI, 0.58-0.75; P<.001) in favor of the triplet regimen, but not for 1-year survival (OR, 1.01; 95% CI, 0.85-1.21; P =.88). The median survival ratio was 1.00 (95% CI, 0.94-1.06; P =.97). CONCLUSION: Adding a second drug improved tumor response and survival rate. Adding a third drug had a weaker effect on tumor response and no effect on survival. PMID- 15280346 TI - Health care delivery in the Texas prison system: the role of academic medicine. AB - Faced with explosive growth in its prison population and a legal mandate to improve medical care for incarcerated offenders, the state of Texas implemented a novel correctional managed health care program in 1994. The organizational structure of the program is based on a series of contractual relationships between the state prison system, 2 of the state's academic medical centers, and a separate governing body composed of 9 appointed members, which include 5 physicians. All medical, dental, and psychiatric care for more than 145,000 offenders, incarcerated under the jurisdiction of the Texas Department of Criminal Justice, is provided by the University of Texas Medical Branch and Texas Tech University Health Sciences Center. The health delivery system is composed of several levels of care, including primary ambulatory care clinics in each prison unit, 16 infirmaries at strategic locations throughout the state, several regional medical facilities, and a dedicated prison hospital with a full range of services. Specialized treatment programs have been established at various units for patients with chronic conditions, such as hypertension, diabetes mellitus, major psychiatric disorders, hepatitis, and human immunodeficiency virus infection. Significant improvements in health outcomes have occurred since the managed care program was established. PMID- 15280348 TI - Decision aids from genetics to treatment of breast cancer: long-term clinical utility or temporary solution? PMID- 15280347 TI - Clinical implications of the osteoprotegerin/RANKL/RANK system for bone and vascular diseases. AB - Bone resorption by osteoclasts is coupled with bone formation by osteoblasts, and this balanced process continuously remodels and adapts the skeleton. The receptor activator of nuclear factor kappaB ligand (RANKL) has been identified as an essential cytokine for the formation and activation of osteoclasts. The effects of RANKL are physiologically counterbalanced by the decoy receptor osteoprotegerin (OPG). Estrogen deficiency, glucocorticoid exposure, T-cell activation (eg, rheumatoid arthritis), and skeletal malignancies (eg, myeloma, metastases) enhance the ratio of RANKL to OPG and, thus, promote osteoclastogenesis, accelerate bone resorption, and induce bone loss. Moreover, alterations of the OPG/RANKL/RANK system have been implicated in vascular diseases. RANKL blockade (using OPG or RANK fusion proteins or RANKL antibodies) has prevented bone loss caused by osteoporosis, chronic inflammatory disorders, and malignant tumors in animal models and may emerge as a therapy in humans based on studies in postmenopausal osteoporosis, myeloma bone disease, and osteolytic metastases. This review summarizes the clinical implications of the OPG/RANKL/RANK system for bone and vascular diseases. PMID- 15280349 TI - Can current treatments for advanced non-small-cell lung cancer be improved? PMID- 15280350 TI - Correctional health care systems and collaboration with academic medicine. PMID- 15280351 TI - JAMA patient page. Genetics and breast cancer. PMID- 15280352 TI - Calcium/calmodulin inhibition of transcriptional activity of E-proteins by prevention of their binding to DNA. AB - The Ca2+ sensor protein calmodulin can interact with the DNA binding basic helix loop-helix (bHLH) domain of E12, E47, and SEF2-1 (E2-2), which belong to the E protein subclass of bHLH transcription factors. This interaction inhibits the DNA binding of these bHLH proteins in vitro, and an ionophore that increases intracellular Ca2+ can inhibit transcriptional activation by the E-proteins. Here we have attempted to determine if these phenomena reflect a direct calmodulin dependent inhibition of DNA binding by E-proteins in vivo. We show that calmodulin overexpression inhibits the transcriptional activity of E12, E47, and SEF2-1. We have compared calmodulin effects on DNA binding in vitro and on activation of transcription in vivo using a series of E12 mutants harboring defined alterations within the basic sequence of the bHLH domain that reduce their ability to bind calmodulin to varying degrees. We find a striking direct correlation between the ability of calmodulin to inhibit their DNA binding in vitro and the ability of overexpressed calmodulin or cellular Ca2+ mobilization to inhibit their transcriptional activity in vivo. Furthermore, E12 and overexpressed calmodulin were co-localized in the nucleus, and calmodulin pull down experiments with cell extracts showed a Ca2+-dependent interaction between calmodulin and E12 but not with a calmodulin inhibition-deficient E12 mutant. Chromatin immunoprecipitation showed that calmodulin overexpression leads to decreased binding of E12 and E47, but not a calmodulin inhibition-deficient E12 mutant, to the DNA recognition sequence in vivo. The data suggest that Ca2+ signaling can inhibit the transcriptional activities of E-proteins through direct binding of Ca2+/calmodulin to the basic sequence of E-proteins, resulting in inhibition of their DNA binding. PMID- 15280353 TI - Discrete roles for histone acetylation in human T helper 1 cell-specific gene expression. AB - To better understand the control of T helper (TH) 1-expressed genes, we compared and contrasted acetylation and expression for three key genes, IFNG, TBET, and IL18RAP and found them to be distinctly regulated. The TBET and the IFNG genes, but not the IL18RAP gene, showed preferential acetylation of histones H3 and H4 during TH1 differentiation. Analysis of acetylation of specific histone residues revealed that H3(Lys-9), H4(Lys-8), and H4(Lys-12) were preferentially modified in TH1 cells, suggesting a possible contribution of acetylation of these residues for induction of these genes. On the other hand, the acetylation of IL18RAP gene occurred both in TH1 and TH2 cells the similar kinetics and on the same with residues, demonstrating that selective histone acetylation was not universally the case for all TH1-expressed genes. Histone H3 acetylation of IFNG and TBET genes occurred with different kinetics, however, and was distinctively regulated by cytokines. Interleukin (IL)-12 and IL-18 enhanced the histone acetylation of the IFNG gene. By contrast, histone acetylation of the TBET gene was markedly suppressed by IL-4, whereas IL-12 and IL-18 had only modest effects suggesting that histone acetylation during TH1 differentiation is a process that is regulated by various factors at multiple levels. By treating Th2 cells with a histone deacetylase inhibitor, we restored histone acetylation of the IFNG and TBET genes, but it did not fully restore their expression in TH2 cells, again suggesting that histone acetylation explains one but not all the aspects of TH1 specific gene expression. PMID- 15280354 TI - Ca2+ controls functional expression of the cardiac K+ transient outward current via the calcineurin pathway. AB - The transient outward K+ current (Ito) modulates transmembrane Ca2+ influx into cardiomyocytes, which, in turn, might act on Ito. Here, we investigated whether Ca2+ modifies functional expression of Ito. Whole-cell Ito were recorded using the patch clamp technique in single right ventricular myocytes isolated from adult rats and incubated for 24 h at 37 degrees C in a serum-free medium containing various Ca2+ concentrations ([Ca2+]o). Increasing the [Ca2+]o from 0.5 to 1.0 and 2.5 mM produced a gradual decrease in Ito density without change in current kinetics. Quantitativereverse transcriptase-PCR showed that a decrease of the Kv4.2 mRNA could account for this decrease. In the acetoxymethyl ester form of 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA-AM)-loaded myocytes (a permeant Ca2+ chelator), Ito density increased significantly when cells were exposed for 24 h to either 1 or 2.5 mM [Ca2+]o. Moreover, 24-h exposure to the Ca2+ channel agonist, Bay K8644, in 1 mM [Ca2+]o induced a decrease in Ito density, whereas the Ca2+ channel antagonist, nifedipine, blunted Ito decrease in 2.5 mM [Ca2+]o. The decrease of Ito in 2.5 mM [Ca2+]o was also prevented by co-incubation with either the calmodulin inhibitor W7 or the calcineurin inhibitors FK506 or cyclosporin A. Furthermore, in myocytes incubated for 24 h with 2.5 mM [Ca2+]o, calcineurin activity was significantly increased compared with 1 mM [Ca2+]o. Our data suggest that modulation of [Ca2+]i via L type Ca2+ channels, which appears to involve the Ca2+/calmodulin-regulated protein phosphatase calcineurin, down-regulates the functional expression of Ito. This effect might be involved in many physiological and pathological modulations of Ito channel expression in cardiac cells, as well other cell types. PMID- 15280355 TI - Evidence for a new sub-class of methionine sulfoxide reductases B with an alternative thioredoxin recognition signature. AB - Methionine sulfoxide reductases catalyze the reduction of protein-bound methionine sulfoxide back to methionine via a thioredoxin-recycling process. Two classes of methionine sulfoxide reductases, called MsrA and MsrB, exist that display opposite stereoselectivities toward the sulfoxide function. Although they are structurally unrelated, they share a similar chemical mechanism that includes three steps with 1) formation of a sulfenic acid intermediate with a concomitant release of 1 mol of methionine per mole of enzyme; 2) formation of an intradisulfide Msr bond; and 3) reduction of the oxidized Msr by thioredoxin. In the MsrBs that have been biochemically, enzymatically, and structurally characterized so far, the cysteine involved in the regeneration of the catalytic Cys-117 is Cys-63. Cys-117 is located on a beta strand, whereas the recycling Cys 63 is on a loop near Cys-117. The distance between the two cysteines is compatible with formation of the Cys-117/Cys-63 intradisulfide bond. Analyses of MsrB sequences show that at least 37% of the MsrBs do not possess the recycling Cys-63. In the present study, it is shown that Cys-31 in the Xanthomonas campestris MsrB, which is located on another loop, can efficiently substitute for Cys-63. Such a result implies flexibility of the MsrB structures, at least of the loops on which Cys-31 or Cys-63 are located. The fact that about 25% of the putative MsrBs have no recycling cysteine supports other recycling processes in which thioredoxin is not operative. PMID- 15280356 TI - Induction of apoptosis by X-linked ectodermal dysplasia receptor via a caspase 8 dependent mechanism. AB - X-linked ectodermal dysplasia receptor (XEDAR) is a recently isolated member of the tumor necrosis factor receptor family that is highly expressed during embryonic development and binds to ectodysplasin-A2 (EDA-A2). In this report, we demonstrate that although XEDAR lacks a death domain, it nevertheless induces apoptosis in an EDA-A2-dependent fashion. The apoptosis-inducing ability of XEDAR is dependent on the activation of caspase 8 and can be blocked by its genetic and pharmacological inhibitors. Although XEDAR-induced apoptosis can be blocked by dominant-negative Fas-associated death domain (FADD) protein and FADD small interfering RNA, XEDAR does not directly bind to FADD, tumor necrosis factor receptor-associated death domain (TRADD) protein, or RIP1. Instead, XEDAR signaling leads to the formation of a secondary complex containing FADD, caspase 8, and caspase 10, which results in caspase activation. Thus, XEDAR belongs to a novel class of death receptors that lack a discernible death domain but are capable of activating apoptosis in a caspase 8- and FADD-dependent fashion. XEDAR may represent an early stage in the evolution of death receptors prior to the emergence of the death domain and may play a role in the induction of apoptosis during embryonic development and adult life. PMID- 15280357 TI - Evidence against the rescue of defective DeltaF508-CFTR cellular processing by curcumin in cell culture and mouse models. AB - Curcumin, the yellow colored component of the spice turmeric, has been reported to rescue defective DeltaF508-cystic fibrosis transmembrane conductance regulator (CFTR) cellular processing in homozygous mutant mice, restoring nasal potential differences and improving survival (Egan, M. E., Pearson, M., Weiner, S. A., Rajendran, V., Rubin, D., Glockner-Pagel, J., Canny, S., Du, K., Lukacs, G. L., and Caplan, M. J. (2004) Science 304, 600-602). Because of the implied potential use of curcumin or similar compounds in the therapy of cystic fibrosis caused by the DeltaF508 mutation, we tried to reproduce and extend the pre-clinical data of Egan et al. Fluorometric measurements of iodide influx in Fischer rat thyroid cells expressing DeltaF508-CFTR showed no effect of curcumin (1-40 microm) when added for up to 24 h prior to assay in cells grown at 37 degrees C. Controls, including 27 degrees C rescue and 4 mm phenylbutyrate at 37 degrees C, were strongly positive. Also, curcumin did not increase short circuit current in primary cultures of a human airway epithelium homozygous for DeltaF508-CFTR with a 27 degrees C rescue-positive control. Nasal potential differences in mice were measured in response to topical perfusion with serial solutions containing amiloride, low Cl-, and forskolin. Robust low Cl- and forskolin-induced hyperpolarization of 22 +/- 3 mV was found in wild type mice, with 2.1 +/- 0.4 mV hyperpolarization in DeltaF508 homozygous mutant mice. No significant increase in Cl-/forskolin hyperpolarization was seen in any of the 22 DeltaF508 mice studied using different curcumin preparations and administration regimens, including that used by Egan et al. Assay of serum curcumin by ethyl acetate extraction followed by liquid chromatography/mass spectrometry indicated a maximum serum concentration of 60 nm, well below that of 5-15 microm, where cellular effects by sarcoplasmic/endoplasmic reticulum calcium pump inhibition are proposed to occur. Our results do not support further evaluation of curcumin for cystic fibrosis therapy. PMID- 15280358 TI - ZNF76, a novel transcriptional repressor targeting TATA-binding protein, is modulated by sumoylation. AB - Direct interaction of positive and negative regulators with the general transcription machinery modulates transcription. The TATA-binding protein (TBP) is one target for transcriptional regulators. In this study, we identified ZNF76 as a novel transcriptional repressor that targets TBP. ZNF76 interacts with TBP through both its N and C termini, and both regions are required for ZNF76 to exert its inhibitory function on p53-mediated transactivation. The inhibitory effect of ZNF76 on p53 activity was demonstrated by reporter assays and endogenous target gene expression. We mapped the TBP-interacting region in the C terminus of ZNF76 to a glutamic acid-rich domain, which acts in a dominant negative manner to enhance p53-mediated transactivation in reporter assays. Mutagenesis study for ZNF76 suggests a correlation between interaction with TBP and effect on p53-mediated transactivation, supporting the conclusion that ZNF76 targets TBP for transcriptional repression. Chromatin immunoprecipitation experiments suggest that ZNF76 prevents TBP from occupying the endogenous p21 promoter. ZNF76 is sumoylated by PIAS1 at lysine 411, which is in the minimal TBP interacting region. Overexpression of PIAS1 and SUMO-1 abolishes the interaction between ZNF76 and TBP and partially relieves the repressive effect of ZNF76. These results suggest that ZNF76 functions as a transcriptional repressor through its interaction with TBP and that sumoylation modulates its transcriptional repression activity. PMID- 15280359 TI - Crystal structure and mutagenesis of a protein phosphatase-1:calcineurin hybrid elucidate the role of the beta12-beta13 loop in inhibitor binding. AB - Protein phosphatase-1 and protein phosphatase-2B (calcineurin) are eukaryotic serine/threonine phosphatases that share 40% sequence identity in their catalytic subunits. Despite the similarities in sequence, these phosphatases are widely divergent when it comes to inhibition by natural product toxins, such as microcystin-LR and okadaic acid. The most prominent region of non-conserved sequence between these phosphatases corresponds to the beta12-beta13 loop of protein phosphatase-1, and the L7 loop of toxin-resistant calcineurin. In the present study, mutagenesis of residues 273-277 of the beta12-beta13 loop of the protein phosphatase-1 catalytic subunit (PP-1c) to the corresponding residues in calcineurin (312-316), resulted in a chimeric mutant that showed a decrease in sensitivity to microcystin-LR, okadaic acid, and the endogenous PP-1c inhibitor protein inhibitor-2. A crystal structure of the chimeric mutant in complex with okadaic acid was determined to 2.0-A resolution. The beta12-beta13 loop region of the mutant superimposes closely with that of wild-type PP-1c bound to okadaic acid. Systematic mutation of each residue in the beta12-beta13 loop of PP-1c showed that a single amino acid change (C273L) was the most influential in mediating sensitivity of PP-1c to toxins. Taken together, these data indicate that it is an individual amino acid residue substitution and not a change in the overall beta12-beta13 loop conformation of protein phosphatase-1 that contributes to disrupting important interactions with inhibitors such as microcystin-LR and okadaic acid. PMID- 15280360 TI - Synthesis of mosaic peptidoglycan cross-bridges by hybrid peptidoglycan assembly pathways in gram-positive bacteria. AB - The peptidoglycan cross-bridges of Staphylococcus aureus, Enterococcus faecalis, and Enterococcus faecium consist of the sequences Gly(5), l-Ala(2), and d-Asx, respectively. Expression of the fmhB, femA, and femB genes of S. aureus in E. faecalis led to the production of peptidoglycan precursors substituted by mosaic side chains that were efficiently used by the penicillin-binding proteins for cross-bridge formation. The Fem transferases were specific for incorporation of glycyl residues at defined positions of the side chains in the absence of any additional S. aureus factors such as tRNAs used for amino acid activation. The PBPs of E. faecalis displayed a broad substrate specificity because mosaic side chains containing from 1 to 5 residues and Gly instead of l-Ala at the N-terminal position were used for peptidoglycan cross-linking. Low affinity PBP2a of S. aureus conferred beta-lactam resistance in E. faecalis and E. faecium, thereby indicating that there was no barrier to heterospecific expression of resistance caused by variations in the structure of peptidoglycan precursors. Thus, conservation of the structure of the peptidoglycan cross-bridges in members of the same species reflects the high specificity of the enzymes for side chain synthesis, although this is not essential for the activity of the PBPs. PMID- 15280361 TI - A search for hyperglycosylation signals in yeast glycoproteins. AB - N-oligosaccharides of Saccharomyces cerevisiae glycoproteins are classified as core and mannan types. The former contain 13-14 mannoses whereas mannan-type structures consist of an inner core extended with an outer chain of up to 200-300 mannoses, a process known as hyperglycosylation. The selection of substrates for hyperglycosylation poses a theoretical and practical question. To identify hyperglycosylation determinants, we have analyzed the influence of the second amino acid (Xaa) of the sequon in this process using the major exoglucanase as a model. Our results indicate that negatively charged amino acids inhibit hyperglycosylation, whereas positively charged counterparts promote it. On the basis of the tridimensional structure of Exg1, we propose that Xaa influences the orientation of the inner core making it accessible to mannan polymerase I in the appropriate position for the addition of alpha-1,6-mannoses. The presence of Glu in the Xaa of the second sequon of the native exoglucanase suggests that negative selection may drive evolution of these sites. However, a comparison of invertases secreted by S. cerevisiae and Pichia anomala suggests that hyperglycosylation signals are also subjected to positive selection. PMID- 15280362 TI - Catalase-peroxidases (KatG) exhibit NADH oxidase activity. AB - Catalase-peroxidases (KatG) produced by Burkholderia pseudomallei, Escherichia coli, and Mycobacterium tuberculosis catalyze the oxidation of NADH to form NAD+ and either H2O2 or superoxide radical depending on pH. The NADH oxidase reaction requires molecular oxygen, does not require hydrogen peroxide, is not inhibited by superoxide dismutase or catalase, and has a pH optimum of 8.75, clearly differentiating it from the peroxidase and catalase reactions with pH optima of 5.5 and 6.5, respectively, and from the NADH peroxidase-oxidase reaction of horseradish peroxidase. B. pseudomallei KatG has a relatively high affinity for NADH (Km=12 microm), but the oxidase reaction is slow (kcat=0.54 min(-1)) compared with the peroxidase and catalase reactions. The catalase-peroxidases also catalyze the hydrazinolysis of isonicotinic acid hydrazide (INH) in an oxygen- and H2O2-independent reaction, and KatG-dependent radical generation from a mixture of NADH and INH is two to three times faster than the combined rates of separate reactions with NADH and INH alone. The major products from the coupled reaction, identified by high pressure liquid chromatography fractionation and mass spectrometry, are NAD+ and isonicotinoyl-NAD, the activated form of isoniazid that inhibits mycolic acid synthesis in M. tuberculosis. Isonicotinoyl NAD synthesis from a mixture of NAD+ and INH is KatG-dependent and is activated by manganese ion. M. tuberculosis KatG catalyzes isonicotinoyl-NAD formation from NAD+ and INH more efficiently than B. pseudomallei KatG. PMID- 15280363 TI - Arabidopsis CYP735A1 and CYP735A2 encode cytokinin hydroxylases that catalyze the biosynthesis of trans-Zeatin. AB - Cytokinins (CKs), a group of phytohormones, are adenine derivatives that carry either an isoprene-derived or an aromatic side chain at the N(6) terminus. trans Zeatin (tZ), an isoprenoid CK, is assumed to play a central physiological role because of its general occurrence and high activity in bioassays. Although hydroxylation of isopentenyladenine-type CKs is a key step of tZ biosynthesis, the catalyzing enzyme has not been characterized yet. Here we demonstrate that CYP735A1 and CYP735A2 are cytochrome P450 monooxygenases (P450s) that catalyze the biosynthesis of tZ. We identified the genes from Arabidopsis using an adenosine phosphate-isopentenyltransferase (AtIPT4)/P450 co-expression system in yeast. Co-expression of AtIPT4 and CYP735A enabled yeast to excrete tZ and the nucleosides to the culture medium. In vitro, both CYP735As preferentially utilized isopentenyladenine nucleotides rather than the nucleoside and free base forms and produced tZ nucleotides but not the cis-isomer. The expression of CYP735A1 and CYP735A2 was differentially regulated in terms of organ specificity and response to CK. Root-specific induction of CYP735A2 expression by CK suggests that the trans-hydroxylation is involved in the regulation of CK metabolism and signaling in roots. PMID- 15280364 TI - Histone deacetylase 7 associates with hypoxia-inducible factor 1alpha and increases transcriptional activity. AB - Hypoxia-inducible factor (HIF)-1alpha is a transcription factor that controls expression of genes responsive to low oxygen tension, including vascular endothelial growth factor (VEGF), erythropoietin, and glycolytic enzymes. The activity of HIF-1alpha is regulated by binding to the transcriptional co activator cAMP-response element-binding protein-binding protein (CBP)/p300. Using the yeast two-hybrid screening system, we found that the inhibitory domain of HIF 1alpha strongly interacted with the C-terminal domain of histone deacetylase (HDAC) 7. The o-nitrophenyl beta-d-galactopyranoside assay revealed that regions containing amino acids 735-785 of HIF-1alpha and amino acids 669-952 of HDAC7 were minimum contact sites of the interaction. The binding of HDAC7 with HIF 1alpha was reproduced in HEK293 cells grown under normoxic and hypoxic conditions (2% O(2)). HDAC7 bound solely to HIF-1alpha among other HIF-alpha family members, including HIF-2alpha and HIF-3alpha, whereas HIF-1alpha only interacted with HDAC7 in the class II HDAC family. Although HDAC7 was localized dominantly in the cytoplasm at normal oxygen concentrations, HDAC7 co-translocated to the nucleus with HIF-1alpha under hypoxic conditions. In the nucleus, HDAC7 increased transcriptional activity of HIF-1alpha through the formation of a complex with HIF-1alpha, HDAC7, and p300. Taken together, these results indicate that HDAC7 is a novel transcriptional activator of HIF-1alpha PMID- 15280365 TI - The effects of the polyglutamine repeat protein ataxin-1 on the UbL-UBA protein A1Up. AB - The ataxin-1 interacting ubiquitin-like protein (A1Up) contains an amino-terminal ubiquitin-like (UbL) region, four stress-inducible, heat shock chaperonin-binding motifs (STI1), and an ubiquitin-associated domain (UBA) at the carboxyl terminus of A1Up. Although proteins that have both an UbL and UBA domain are thought to play a crucial role in proteasome-mediated activities, few are characterized, except for hHR23A/B. Similar to other UbL-containing proteins, the UbL of A1Up is essential for the interaction of A1Up with the S5a subunit of the 19S proteasome. Importantly, the interaction with the 19S proteasome was disrupted in the presence of the polyglutamine repeat protein, ataxin-1. The UbL domain of A1Up is ubiquitinated by both Lys(48)-linked and Lys(63)-linked chains. Intact A1Up is stable, suggesting that ubiquitination of A1Up is important for degradation independent targeting of A1Up to the 19S proteasome. The UBA domain of A1Up binds polyubiquitin chains and has a role in the stability of A1Up and in the subcellular localization of A1Up. When the UBA domain was deleted, the localization of A1Up was entirely cytoplasmic, and it co-localized with the proteasome. Interestingly, the interaction between A1Up and mutant ataxin-1-(82Q) increased the half-life of A1Up, whereas nonpathogenic wild-type ataxin-1-(30Q) or ataxin-1-(82Q)-A776 did not. PMID- 15280366 TI - Neuronal apoptosis-inhibitory protein does not interact with Smac and requires ATP to bind caspase-9. AB - The neuronal apoptosis-inhibitory protein (NAIP) is the founding member of the mammalian family of inhibitor of apoptosis (IAP) proteins (also known as BIRC proteins) and has been shown to be antiapoptotic both in vivo and in vitro. The 160-kDa NAIP contains three distinct regions: an amino-terminal cluster of three baculoviral inhibitory repeat (BIR) domains, a central nucleotide binding oligomerization domain (NOD), and a carboxyl-terminal leucine-rich repeat (LRR) domain. The presence of the NOD and LRR domains renders NAIP unique among the IAPs and suggests that NAIP activity is regulated in a manner distinct from that of other members of the family. In this report, we examined the interaction of various regions of NAIP with caspase-9 and Smac. Recombinant NAIPs with truncations of the carboxyl-terminal LRR or NOD-LRR regions bound to caspase-9. In contrast, the full-length protein did not, suggesting some form of structural autoregulation. However, the association of the wild type full-length protein with caspase-9 was observed when interaction analysis was performed in the presence of ATP. Furthermore, mutation of the NAIP ATP binding pocket allowed full-length protein to interact with caspase-9. Thus, we conclude that NAIP binds to caspase-9 with a structural requirement for ATP and that in the absence of ATP the LRR domain negatively regulates the caspase-9-inhibiting activity of the BIR domains. Interestingly, and in contrast to the X-chromosome-linked inhibitor of apoptosis protein (XIAP), NAIP-mediated inhibition of caspase-9 was not countered by a peptide containing an amino-terminal IAP binding motif (IBM). Consistent with this observation was the failure of Smac protein to interact with the NAIP BIR domains. These results demonstrate that NAIP is distinct from the other IAPs, both in demonstrating a ligand-dependent caspase-9 interaction and in demonstrating a distinct mechanism of inhibition. PMID- 15280367 TI - Essential role of B-helix calcium binding sites in annexin V-membrane binding. AB - Crystal structures of annexin V have shown up to 10 bound calcium ions in three different types of binding sites, but previous work concluded that only one of these sites accounted for nearly all of the membrane binding affinity of the molecule. In this study we mutated residues contributing to potential calcium binding sites in the AB and B helices in each of the four domains (eight sites in total) and in DE helices in the first, second, and third domains (three sites in total). We measured the affinity of each protein for phospholipid vesicles and cell membranes by quantitative calcium titration under low occupancy conditions (< 1% saturation of available membrane binding sites). Affinity was calculated from the midpoint and slope of the calcium titration curve and the concentration of membrane binding sites. The results showed that all four AB sites were essential for high affinity binding, as were three of the four B sites (in domains 1, 2, and 3); the DE site in the first domain made a slight contribution to affinity. Multisite mutants showed that each domain contributed additively and independently to binding affinity; in contrast, AB and B sites within the same domain were interdependent. The number of functionally important sites identified was consistent with the Hill coefficient observed in calcium titrations. This study shows an essential and previously unappreciated role for B-helix calcium binding sites in the membrane binding of annexins and indicates that all four domains of the molecule are required for maximum membrane binding affinity. PMID- 15280368 TI - Stress-induced expression of the gamma subunit (FXYD2) modulates Na,K-ATPase activity and cell growth. AB - In kidney, the Na,K-ATPase is associated with a single span protein, the gamma subunit (FXYD2). Two splice variants are differentially expressed along the nephron and have a differential influence on Na,K-ATPase when stably expressed in mammalian cells in culture. Here we used a combination of gene induction and gene silencing techniques to test the functional impact of gamma by means other than transfection. NRK-52E cells (of proximal tubule origin) do not express gamma as a protein under regular tissue culture conditions. However, when they were exposed to hyperosmotic medium, induction of only the gammaa splice variant was observed, which was accompanied by a reduction in the rate of cell division. Kinetic analysis of stable enzyme properties from control (alpha1beta1) and hypertonicity treated cultures (alpha1beta1gammaa) revealed a significant reduction (up to 60%) of Na,K-ATPase activity measured under V(max) conditions with little or no change in the amounts of alpha1beta1. This effect as well as the reduction in cell growth rate was practically abolished when gamma expression was knocked down using specific small interfering RNA duplexes. Surprisingly, a similar induction of endogenous gammaa because of hypertonicity was seen in rat cell lines of other than renal origin: C6 (glioma), PC12 (pheochromocytoma), and L6 (myoblasts). Furthermore, exposure of NRK-52E cells to other stress inducers such as heat shock, exogenous oxidation, and chemical stress also resulted in a selective induction of gammaa. Taken together, the data imply that induction of gammaa may have adaptive value by being a part of a general cellular response to genotoxic stress. PMID- 15280369 TI - Heterooligomeric phosphoribosyl diphosphate synthase of Saccharomyces cerevisiae: combinatorial expression of the five PRS genes in Escherichia coli. AB - The yeast Saccharomyces cerevisiae contains five phosphoribosyl diphosphate (PRPP) synthase-homologous genes (PRS1-5), which specify PRPP synthase subunits 1 5. Expression of the five S. cerevisiae PRS genes individually in an Escherichia coli PRPP-less strain (Deltaprs) showed that a single PRS gene product had no PRPP synthase activity. In contrast, expression of five pairwise combinations of PRS genes resulted in the formation of active PRPP synthase. These combinations were PRS1 PRS2, PRS1 PRS3, and PRS1 PRS4, as well as PRS5 PRS2 and PRS5 PRS4. None of the remaining five possible pairwise combinations of PRS genes appeared to produce active enzyme. Extract of an E. coli strain containing a plasmid-borne PRS1 gene and a chromosome-borne PRS3 gene contained detectable PRPP synthase activity, whereas extracts of strains containing PRS1 PRS2, PRS1 PRS4, PRS5 PRS2, or PRS5 PRS4 contained no detectable PRPP synthase activity. In contrast PRPP could be detected in growing cells containing PRS1 PRS2, PRS1 PRS3, PRS5 PRS2, or PRS5 PRS4. These apparent conflicting results indicate that, apart from the PRS1 PRS3-specified enzyme, PRS-specified enzyme is functional in vivo but unstable when released from the cell. Certain combinations of three PRS genes appeared to produce an enzyme that is stable in vitro. Thus, extracts of strains harboring PRS1 PRS2 PRS5, PRS1 PRS4 PRS5, or PRS2 PRS4 PRS5 as well as extracts of strains harboring combinations with PRS1 PRS3 contained readily assayable PRPP synthase activity. The data indicate that although certain pairwise combinations of subunits produce an active enzyme, yeast PRPP synthase requires at least three different subunits to be stable in vitro. The activity of PRPP synthases containing subunits 1 and 3 or subunits 1, 2, and 5 was found to be dependent on Pi, to be temperature-sensitive, and inhibited by ADP. PMID- 15280370 TI - Induction of hypoxia-inducible factor 1 activity by muscarinic acetylcholine receptor signaling. AB - Hypoxia-inducible factor-1 (HIF-1) is a master regulator of cellular adaptive responses to hypoxia. Levels of the HIF-1alpha subunit increase under hypoxic conditions. Exposure of cells to growth factors, prostaglandin, and certain nitric oxide donors also induces HIF-1alpha expression under non-hypoxic conditions. We demonstrate that muscarinic acetylcholine signals induce HIF 1alpha expression and transcriptional activity in a receptor subtype-specific manner using HEK293 cells transiently overexpressing each of M1-M4 muscarinic acetylcholine receptors. The muscarinic signaling pathways inhibited HIF-1alpha hydroxylation and degradation and induced HIF-1alpha protein synthesis that was confirmed by pulse labeling studies. Muscarinic signal-induced HIF-1alpha protein and HIF-1-dependent gene expression were blocked by treating cells with inhibitors of phosphatidylinositol 3-kinase, MAP kinase kinase, or tyrosine kinase signaling pathways. Dominant-negative forms of Ras and/or Rac-1 significantly suppressed HIF-1 activation by muscarinic signaling. Signaling via M1- and M3- but not M2- or M4-AchRs promote accumulation and transcriptional activation of HIF-1alpha. We conclude that muscarinic acetylcholine signals activate HIF-1 by both stabilization and synthesis of HIF-1alpha and by inducing the transcriptional activity of HIF-1alpha. PMID- 15280371 TI - VPAC receptor modulation of neuroexcitability in intracardiac neurons: dependence on intracellular calcium mobilization and synergistic enhancement by PAC1 receptor activation. AB - Pituitary adenylate cyclase-activating polypeptide (PACAP) and vasoactive intestinal polypeptide (VIP) have been found within mammalian intracardiac ganglia, but the cellular effects of these neuropeptides remain poorly understood. Fluorometric calcium imaging and whole cell patch clamp recordings were used to examine the effects of PACAP and VIP on [Ca2+]i and neuroexcitability, respectively, in intracardiac neurons of neonatal rats. PACAP and VIP evoked rapid increases in [Ca2+]i that exhibited both transient and sustained components. Pharmacological experiments using PAC1 and VPAC receptor selective antagonists demonstrated that the elevations in [Ca2+]i result from the activation of VPAC receptors. The transient increases in [Ca2+]i were shown to be the product of Ca2+ mobilization from caffeine/ryanodine-sensitive intracellular stores and were not due to inositol 1,4,5-trisphosphate-mediated calcium release. In contrast, the sustained [Ca2+]i elevations were dependent on extracellular Ca2+ and were blocked by the transient receptor channel antagonist, 2 aminoethoxydiphenyl borate, which suggests that they are due to Ca2+ entry via store-operated channels. In addition to elevating [Ca2+]i, both PACAP and VIP depolarized intracardiac neurons, and PACAP was further shown to augment action potential firing in these cells. Depolarization of intracardiac neurons by the neuropeptides was dependent on activation of VPAC receptors and the concomitant increases in [Ca2+]i. Although activation of PAC1 receptors alone had no direct effects on neuroexcitability, PAC1 receptor stimulation potentiated the VPAC receptor-induced depolarizations. Furthermore, enhanced action potential firing was only observed upon concurrent stimulation of PAC1 and VPAC receptors, which indicates that these receptors act synergistically to enhance neuroexcitability in intracardiac neurons. PMID- 15280372 TI - Protein kinase Cdelta mediates lysophosphatidic acid-induced NF-kappaB activation and interleukin-8 secretion in human bronchial epithelial cells. AB - Lysophosphatidic acid (LPA), a potent bioactive lipid, elicits many of its biological actions via the specific G-protein-coupled receptors LPA1, LPA2, LPA3, and LPA4. Recently, we have shown that LPA-induced transactivation of platelet derived growth factor receptor-beta is regulated by phospholipase D2 in human bronchial epithelial cells (HBEpCs) (Wang, L., Cummings, R. J., Zhao, Y., Kazlauskas, A., Sham, J., Morris, A., Brindley, D. N., Georas, S., and Natarajan, V. (2003) J. Biol. Chem. 278, 39931-39940). Here, we report that protein kinase Cdelta (PKCdelta) mediates LPA-induced NF-kappaB transcription and interleukin-8 (IL-8) secretion in HBEpCs. Treatment of HBEpCs with LPA increased both IL-8 gene and protein expression, which was coupled to Gi and G(12/13) proteins. LPA caused a marked activation of NF-kappaB in HBEpCs as determined by IkappaB phosphorylation and of NF-kappaB nuclear translocation and a strong induction of NF-kappaB promoter-mediated luciferase activity. Furthermore, LPA-activated PKCdelta and the LPA-mediated activation of NF-kappaB and IL-8 production were attenuated by overexpression of dominant-negative PKCdelta and rottlerin. Intratracheal administration of LPA in mice resulted in elevated levels of macrophage inflammatory protein-2, a murine homolog of IL-8, and an influx of neutrophils in the bronchoalveolar lavage fluid. These results demonstrate for the first time that LPA is a potent stimulator of IL-8 production in HBEpCs, which involves PKCdelta/NF-kappaB signaling pathways. PMID- 15280373 TI - Speriolin is a novel spermatogenic cell-specific centrosomal protein associated with the seventh WD motif of Cdc20. AB - The fundamental mechanisms of mitosis are conserved throughout evolution in eukaryotes, including ubiquitin-mediated proteolysis of cell cycle regulators by the anaphase-promoting complex/cyclosome. The spindle checkpoint protein Cdc20 activates the anaphase-promoting complex/cyclosome in a substrate-specific manner. It is present in the cytoplasm and concentrated in the centrosomes throughout the cell cycle, accumulates at the kinetochores in metaphase, and is no longer detected following anaphase. However, it is unknown whether Cdc20 has the same activities and distribution during meiosis in male germ cells. We found that in mice, Cdc20 accumulates in the cytoplasm of pachytene spermatocytes during meiosis I, is distributed throughout spermatocytes undergoing meiotic division, and is present in the cytoplasm of postmeiotic spermatids. Several proteins bind to and regulate the function of Cdc20 during mitosis. We identified speriolin and determined that it is a novel spermatogenic cell-specific Cdc20 binding protein, is present in the cytoplasm, and is concentrated at the centrosomes of spermatocytes and spermatids and that a leucine zipper domain is required to target speriolin to the centrosome. The seven tandem WD motifs of Cdc20 probably fold into a seven-blade beta-propeller structure, and we determined that they are required for speriolin binding and for localization of Cdc20 to the centrosomes and nucleus, suggesting that speriolin might regulate or stabilize the folding of Cdc20 during meiosis in spermatogenic cells. PMID- 15280374 TI - Disabled-2 is a negative regulator of integrin alpha(IIb)beta(3)-mediated fibrinogen adhesion and cell signaling. AB - Disabled-2 (DAB2) is an adapter protein that is up-reg-ulated during megakaryocytic differentiation of hematopoietic cells and is abundantly expressed in platelets. In this study, the role of DAB2 in integrin alpha(IIb)beta(3) mediated matrix protein fibrinogen adhesion and cell signaling was investigated. In K562 cells differentiating to the megakaryocytic lineage, down-regulation of DAB2 by DAB2 small interfering RNA augmented integrin alpha(IIb)beta(3) activation and resulted in an increase in cell adhesion to fibrinogen. Ectopic expression of DAB2 reversed the DAB2 small interfering RNA effect or, by itself, decreased fibrinogen adhesion of K562 cells. Mutational analysis revealed that a DAB2 Ser(24) phosphorylation mutant (S24A) abrogated the inhibitory function of DAB2. The spatial and temporal association/interaction of DAB2 and platelet integrin alpha(IIb)beta(3) (CD61) in both megakaryocytic cells and platelets led us to examine the effect of Ser(24) phosphorylation on the interaction between DAB2 and integrin beta(3). Through cellular localization and co immunoprecipitation analysis, we demonstrate for the first time that Ser(24) phosphorylation promotes membrane translocation of DAB2 and its subsequent interaction with integrin beta(3), thereby defining a mechanism for DAB2 in regulating integrin alpha(IIb)beta(3) activation and inside-out signaling. Consistent with the effect on fibrinogen adhesion, Ser(24) phosphorylation of DAB2 was also involved in the negative regulation of alpha(IIb)beta(3)-induced T cell factor transcriptional activity. In contrast, the S24A mutant acted like wild-type DAB2 and inhibited both beta-catenin- and plakoglobin-mediated T cell factor transactivation. Hence, DAB2 elicits distinct regulatory mechanisms in alpha(IIb)beta(3) and beta-catenin/plakoglobin signaling in a Ser(24) phosphorylation-dependent and -independent manner, respectively. These findings indicate Ser(24) phosphorylation as a molecular basis for DAB2 acting as a negative regulator in alpha(IIb)beta(3) inside-out signaling and contribute to our understanding of DAB2 in megakaryocytic differentiation and platelet function. PMID- 15280375 TI - Protein kinase A inhibits leukotriene synthesis by phosphorylation of 5 lipoxygenase on serine 523. AB - Leukotrienes (LTs) are lipid messengers generated by leukocytes that drive inflammation and modulate neighboring cell function. The synthesis of LTs from arachidonic acid is initiated by the enzyme 5-lipoxygenase (5-LO). We report for the first time that LT synthesis is inhibited by the direct action of protein kinase A (PKA) on 5-LO. The catalytic subunit of PKA directly phosphorylated 5-LO in vivo and in vitro and inhibited activity in intact cells and in vitro. Mutation of Ser-523 on human 5-LO prevented phosphorylation by PKA and restored LT synthesis. Treatment with PKA activators inhibited LTB(4) synthesis in 3T3 cells expressing wild type 5-LO but not in cells expressing the S523A mutant of 5 LO. The mechanism of inhibition of LTB(4) synthesis did not involve either reduced membrane association of activated 5-LO or redistribution of 5-LO from the nucleus to the cytoplasm. Instead, PKA phosphorylation of recombinant 5-LO inhibited in vitro activity, as did co-transfection of cells with 5-LO plus the catalytic subunit of PKA. Also, substitution of Ser-523 with glutamic acid, mimicking phosphorylation, resulted in the total loss of 5-LO activity. These results indicate that PKA phosphorylates 5-LO on Ser-523, which inhibits the catalytic activity of 5-LO and reduces cellular LT generation. Thus, PKA activation, as can occur in response to adenosine, prostaglandin E(2), beta adrenergic agonists, and other mediators, is a means of directly reducing 5-LO activity and LT synthesis that may be important in limiting inflammation and maintaining homeostasis. PMID- 15280376 TI - Characterization of oligomeric human ATP binding cassette transporter A1. Potential implications for determining the structure of nascent high density lipoprotein particles. AB - The oligomeric structure of ABCA1 transporter and its function related to the biogenesis of nascent apoA-I-containing particles (LpA-I) were investigated. Using n-dodecylmaltoside and perfluoro-octanoic acid combined with non-denaturing gel, the majority of ABCA1 was found as a tetramer in ABCA1-induced human fibroblasts. Furthermore, using chemical cross-linking and SDS-PAGE, ABCA1 dimers but not the tetramers were found covalently linked. Oligomeric ABCA1 was present in isolated plasma membranes as well as in intracellular compartments. Interestingly, apoA-I was found to be associated with both dimeric and tetrameric, but not monomeric, forms of ABCA1. Neither apoA-I nor lipid molecules did affect ABCA1 oligomerization. Immunoprecipitation analysis showed that oligomeric ABCA1 did not contain other associated proteins. We next investigated the relationship between the oligomeric ABCA1 complex and the structure of LpA-I. Lipid-free apoA-I incubated with normal cells generated LpA-I with diameters between 9.5 and 20 nm. Subsequent isolation of LpA-I followed by cross-linking revealed the presence of four and eight apoA-I molecules per particle, whereas apoA-I incubated with ABCA1 mutant (Q597R) cells was unable to form such particles and remained in the monomeric form. These results demonstrate that: 1) ABCA1 exists as an oligomeric complex; and 2) ABCA1 oligomerization was independent of apoA-I binding and lipid molecules. The findings that the majority of ABCA1 exists as a tetramer that binds apoA-I, together with the observation that LpA-I contains at least four molecules of apoA-I per particle, support the concept that the homotetrameric ABCA1 complex constitutes the minimum functional unit required for the biogenesis of high density lipoprotein particles. PMID- 15280377 TI - Regulation of p53 by the ubiquitin-conjugating enzymes UbcH5B/C in vivo. AB - p53 levels are regulated by ubiquitination and 26 S proteasome-mediated degradation. p53 is a substrate for the E3 ligase Mdm2, however, the ubiquitin conjugating enzymes (E2s) involved in p53 ubiquitination in intact cells have not been defined previously. To investigate the E2 specificity of Mdm2 we carried out an in vitro screen using a panel of ubiquitin E2s. Of the E2s tested only UbcH5A, -B, and -C and E2-25K support Mdm2-mediated ubiquitination of p53. The same E2s also support Mdm2 auto-ubiquitination. Small interfering RNA-mediated knockdown of UbcH5B/C causes accumulation of Mdm2 and p53 in unstressed cells. We show that suppression of UbcH5B/C inhibits p53 ubiquitination and degradation. Despite up regulating the level of nuclear p53, UbcH5B/C knockdown does not on its own result in an increase in p53 transcriptional activity or sensitize p53 to activation by the therapeutic drugs doxorubicin and actinomycin D. We provide evidence that Mdm2 is responsible, at least in part, for repression of the transcriptional activity of the accumulated p53. In MCF7 cells levels of UbcH5B/C are reduced by doxorubicin and actinomycin D. This observation and the sensitivity of p53 expression to levels of UbcH5B/C raise the possibility that E2 regulation could be involved in signaling pathways that control the stability of p53. Our data indicate that UbcH5B/C are physiological E2s for Mdm2, which make a significant contribution to the maintenance of low levels of p53 and Mdm2 in unstressed cells and that inhibition of p53 ubiquitination and degradation by targeting UbcH5B/C is not sufficient to up-regulate p53 transcriptional activity. PMID- 15280378 TI - Residues Lys-149 and Glu-153 switch the aminoacylation of tRNA(Trp) in Bacillus subtilis. AB - Tryptophanyl-tRNA synthetase (TrpRS) consists of two identical subunits that induce the cross-subunit binding mode of tRNA(Trp). It has been shown that eubacterial and eukaryotic TrpRSs cannot efficiently cross-aminoacylate the corresponding tRNA(Trp). Although the identity elements in tRNA(Trp) that confer the species-specific recognition have been identified, the corresponding elements in TrpRS have not yet been reported. In this study two residues, Lys-149 and Glu 153, were identified as being crucial for the accurate recognition of tRNA(Trp). These residues reside adjacent to the binding pocket for Trp-AMP and show phylogenic diversities in the charge on their side chains between eubacteria and eukaryotes. Single mutagenesis at Lys-149 or Glu-153 reduced the activity of TrpRS in the activation of Trp. The reduction was less than that caused by the double mutant WBHA (K149D/E153R). It is unusual that E153G had no detectable activity in the activation of Trp unless tRNA(Trp) was added to the reaction. In addition, we successfully switched the species specificity of Bacillus subtilis TrpRS recognition of tRNA(Trp). The affinity of WBHA, K149E and E153K to human tRNA(Trp) was 31-, 13.5-, and 12.9-fold greater than that of wild type B. subtilis TrpRS, respectively. Indeed WBHA and E153K were found to prefer genuine human tRNA(Trp) to their cognate eubacteria tRNA(Trp). PMID- 15280379 TI - ADAM binding protein Eve-1 is required for ectodomain shedding of epidermal growth factor receptor ligands. AB - A disintegrin and metalloproteases (ADAMs) are implicated in the ectodomain shedding of epidermal growth factor receptor (EGFR) ligands in EGFR transactivation. However, the activation mechanisms of ADAMs remain elusive. To analyze the regulatory mechanisms of ADAM activation, we performed yeast two hybrid screening using the cytoplasmic domain of ADAM12 as bait, and identified a protein that we designated Eve-1. Two cDNAs were cloned and characterized. They encode alternatively spliced isoforms of Eve-1, called Eve-1a and Eve-1b, that have four and five tandem Src homology 3 (SH3) domains in the carboxyl-terminal region, respectively, and seven proline-rich SH3 domain binding motifs in the amino-terminal region. The short forms of Eve-1, Eve-1c and Eve-1d, translated at Met-371 are human counterparts of mouse Sh3d19. Northern blot analysis demonstrated that Eve-1 is abundantly expressed in skeletal muscle and heart. Western blot analysis revealed the dominant production of Eve-1c in human cancer cell lines. Knockdown of Eve-1 by small interfering RNA in HT1080 cells reduced the shedding of proHB-EGF induced by angiotensin II and 12-O-tetradecanoylphorbol 13-acetate, as well as the shedding of pro-transforming growth factor-alpha, promphiregulin, and proepiregulin by 12-O-tetradecanoylphorbol-13-acetate, suggesting that Eve-1 plays a role in positively regulating the activity of ADAMs in the signaling of EGFR-ligand shedding. PMID- 15280380 TI - Prostaglandin E2 selectively antagonizes prostaglandin F2alpha-stimulated T-cell factor/beta-catenin signaling pathway by the FPB prostanoid receptor. AB - FP prostanoid receptors are G-protein-coupled receptors that consist of two isoforms named FPA and FPB. Both isoforms activate inositol phosphate second messenger signaling pathways by their endogenous ligand prostaglandin F2alpha (PGF2alpha). Previously we have shown that both isoforms undergo Rho-mediated cell rounding following treatment with PGF2alpha. Following the removal of PGF2alpha, however, FPA-expressing cells return to their original morphology, whereas FPB-expressing cells do not. It was also found that PGF2alpha-could activate T-cell factor (Tcf)/beta-catenin signaling in cells expressing the FPB isoform but not in cells expressing the FPA isoform. We now show that prostaglandin E2 (PGE2) can induce cell rounding and stimulate the formation of inositol phosphates to the same extent as PGF2alpha in cells expressing either the FPA or FPB isoforms. However, PGE2 has much lower efficacy as compared with PGF2alpha for the activation of Tcf/beta-catenin signaling in FPB-expressing cells, and the cell rounding is reversible. Interestingly, pretreatment of FPB expressing cells with PGE2-attenuated PGF2alpha-stimulated Tcf/beta-catenin signaling in a dose-dependent manner while having no effect on PGF2alpha stimulated inositol phosphates formation. Thus, the ratio of endogenous PGE2 and PGF2alpha has the potential to selectively regulate one signaling pathway over another. This represents a novel mechanism for the regulation of cell signaling that is distinct from regulation occurring at the level of the receptor and its effector pathways. PMID- 15280381 TI - Methylation of H3 lysine 4 at euchromatin promotes Sir3p association with heterochromatin. AB - Set1p methylates lysine 4 of histone H3 and can activate transcription by recruiting the chromatin-remodeling factor Isw1p. In addition, Lys-4-methylated H3 is required for maintenance of silencing at the telomeres, rDNA, and HML locus in Saccharomyces cerevisiae. The molecular mechanism underlying the role of Set1p in silencing is not known. Here we report that euchromatic methylation of H3 Lys 4 is necessary to maintain silencing at specific heterochromatic sites. Inactivation of Set1p catalytic activity or mutation of H3 Lys-4 leads to decreased binding of the silent information regulator Sir3p at heterochromatic sites. Concomitantly, there is an increase in the amount of Sir3p bound to genes located in subtelomeric regions. Consistent with this result is the finding that in vitro, Sir3p preferentially binds histone H3 tails when methylation is absent at H3 Lys-4, a situation found in heterochromatin. The inability of Sir3p to bind methylated H3 Lys-4 tails suggests a model whereby H3 Lys-4 methylation prevents Sir3p association at euchromatic sites and therefore concentrates Sir3p at unmodified, heterochromatic regions of the genome. PMID- 15280382 TI - Peroxiredoxin III, a mitochondrion-specific peroxidase, regulates apoptotic signaling by mitochondria. AB - Various proapoptotic stimuli increase the production of superoxide and H(2)O(2) by mitochondria. Whereas superoxide impairs mitochondrial function and is removed by Mn(2+)-dependent superoxide dismutase, the role and metabolism of mitochondrial H(2)O(2) during apoptosis have remained unclear. The effects on apoptotic signaling of depletion of peroxiredoxin (Prx) III, a mitochondrion specific H(2)O(2)-scavenging enzyme, have now been investigated by RNA interference in HeLa cells. Depletion of Prx III resulted in increased intracellular levels of H(2)O(2) and sensitized cells to induction of apoptosis by staurosporine or TNF-alpha. The rates of mitochondrial membrane potential collapse, cytochrome c release, and caspase activation were increased in Prx III depleted cells, and these effects were reversed by ectopic expression of Prx III or mitochondrion-targeted catalase. Depletion of Prx III also exacerbated damage to mitochondrial macromolecules induced by the proapoptotic stimuli. Our results suggest that Prx III is a critical regulator of the abundance of mitochondrial H(2)O(2), which itself promotes apoptosis in cooperation with other mediators of apoptotic signaling. PMID- 15280383 TI - Purification and spectropotentiometric characterization of Escherichia coli NrfB, a decaheme homodimer that transfers electrons to the decaheme periplasmic nitrite reductase complex. AB - Escherichia coli can reduce nitrite to ammonium via a 120-kDa decaheme homodimeric periplasmic nitrite reductase (NrfA) complex. Recent structure-based spectropotentiometric studies are shedding light on the catalytic mechanism of NrfA; however, electron input into the enzyme has not been addressed biochemically. This study reports the first purification of NrfB, a novel 20-kDa pentaheme c-type cytochrome encoded by the nrfB gene that follows the nrfA gene in many bacterial nrf operons. Analyses by gel filtration demonstrated that NrfB purifies as a decaheme homodimer. Analysis of NrfB by UV-visible and magnetic circular dichroism spectroscopy demonstrates that all five NrfB ferric heme irons are low spin and are most likely coordinated by two axial histidine ligands. Spectropotentiometry revealed that the midpoint redox potentials of five ferric hemes were in the low potential range of 0 to -400 mV. Analysis by low temperature EPR spectroscopy revealed signals that arise from two classes of bis His ligated low spin hemes, namely a rhombic trio at g(1,2,3) = 2.99, 2.27, and 1.5 that arises from two hemes in which the planes of histidine imidazole rings are near-parallel and a large g(max) signal at g = 3.57 that arises from three hemes in which the planes of the histidine imidazole rings are near perpendicular. NrfB was also overexpressed as a recombinant protein, which had similar spectropotentiometric properties as the native protein. Reconstitution experiments demonstrated that the reduced decaheme NrfB dimer could serve as a direct electron donor to the oxidized decaheme NrfA dimer, thus forming a transient 20-heme [NrfB](2)[NrfA](2) electron transfer complex. PMID- 15280384 TI - Processing and transport of matrix gamma-carboxyglutamic acid protein and bone morphogenetic protein-2 in cultured human vascular smooth muscle cells: evidence for an uptake mechanism for serum fetuin. AB - Matrix gamma-carboxyglutamic acid protein (MGP) is a member of the vitamin K dependent protein family with unique structural and physical properties. MGP has been shown to be an inhibitor of arterial wall and cartilage calcification. One inhibitory mechanism is thought to be binding of bone morphogenetic protein-2. Binding has been shown to be dependent upon the vitamin K-dependent gamma carboxylation modification of MGP. Since MGP is an insoluble matrix protein, this work has focused on intracellular processing and transport of MGP to become an extracellular binding protein for bone morphogenetic protein-2. Human vascular smooth muscle cells (VSMCs) were infected with an adenovirus carrying the MGP construct, which produced non-gamma-carboxylated MGP and fully gamma-carboxylated MGP. Both forms of MGP were found in the cytosolic and microsomal fractions obtained from the cells by differential centrifugation. The crude microsomal fraction was shown to contain an additional, more acidic Ser-phosphorylated form of MGP believed to be the product of Golgi casein kinase. The data suggest that phosphorylation of MGP dictates different transport routes for MGP in VSMCs. A proteomic approach failed to identify a larger soluble precursor of MGP or an intracellular carrier protein for MGP. Evidence is presented for a receptor mediated uptake mechanism for fetuin by cultured human VSMCs. Fetuin, shown by mass spectrometry not to contain MGP, was found to be recognized by anti-MGP antibodies. Fetuin uptake and secretion by proliferating and differentiating cells at sites of calcification in the arterial wall may represent an additional protective mechanism against arterial calcification. PMID- 15280385 TI - G2A is a proton-sensing G-protein-coupled receptor antagonized by lysophosphatidylcholine. AB - G2A (from G2 accumulation) is a G-protein-coupled receptor (GPCR) that regulates the cell cycle, proliferation, oncogenesis, and immunity. G2A shares significant homology with three GPCRs including ovarian cancer GPCR (OGR1/GPR68), GPR4, and T cell death-associated gene 8 (TDAG8). Lysophosphatidylcholine (LPC) and sphingosylphosphorylcholine (SPC) were reported as ligands for G2A and GPR4 and for OGR1 (SPC only), and a glycosphingolipid psychosine was reported as ligand for TDAG8. As OGR1 and GPR4 were reported as proton-sensing GPCRs (Ludwig, M. G., Vanek, M., Guerini, D., Gasser, J. A., Jones, C. E., Junker, U., Hofstetter, H., Wolf, R. M., and Seuwen, K. (2003) Nature 425, 93-98), we evaluated the proton sensing function of G2A. Transient expression of G2A caused significant activation of the zif 268 promoter and inositol phosphate (IP) accumulation at pH 7.6, and lowering extracellular pH augmented the activation only in G2A expressing cells. LPC inhibited the pH-dependent activation of G2A in a dose dependent manner in these assays. Thus, G2A is another proton-sensing GPCR, and LPC functions as an antagonist, not as an agonist, and regulates the proton dependent activation of G2A. PMID- 15280386 TI - Self-interacting domains in the C terminus of a cation-Cl- cotransporter described for the first time. AB - The first isoform of the Na+-K+-Cl- cotransporter (NKCC1), a widely distributed member of the cation-Cl- cotransporter superfamily, plays key roles in many physiological processes by regulating the ion and water content of animal cells and by sustaining electrolyte secretion across various epithelia. Indirect studies have led to the prediction that NKCC1 operates as a dimer assembled through binding domains that are distal to the amino portion of the carrier. In this study, evidence is presented that NKCC1 possesses self-interacting properties that result in the formation of a large complex between the proximal and the distal segment of the cytosolic C terminus. Elaborate mapping studies of these segments showed that the contact sites are dispersed along the entire C terminus, and they also led to the identification of a critical interacting residue that belongs to a putative forkhead-associated binding domain. In conjunction with previous findings, our results indicate that the uncovered interacting domains are probably a major determinant of the NKCC1 conformational landscape and assembly into a high order structure. A model is proposed in which the carrier could alternate between monomeric and homo-oligomeric units via chemical- or ligand-dependent changes in conformational dynamics. PMID- 15280387 TI - Carbon monoxide promotes Fas/CD95-induced apoptosis in Jurkat cells. AB - A properly functioning immune system is dependent on programmed cell death/apoptosis at virtually every stage of lymphocyte development and activity. Carbon monoxide (CO), an enzymatic product of heme oxyenase-1, has been shown to possess anti-apoptotic effects in a number of different model systems. The purpose of the present study was to expand on this knowledge to determine the role of CO in the well established model of Fas/CD95-induced apoptosis in Jurkat cells, and to determine the mechanism by which CO can modulate T-cell apoptosis. Exposure of Jurkat cells to CO resulted in augmentation in Fas/CD95-induced apoptosis, which correlated with CO-induced up-regulation of the pro-apoptotic protein FADD as well as activation of caspase-8, -9, and -3 while simultaneously down-regulating the anti-apoptotic protein BCL-2. These effects of CO were lost with overexpression of the small interfering RNA of FADD. CO, as demonstrated previously in endothelial cells, was also anti-apoptotic in Jurkat cells against tumor necrosis factor and etoposide. We further demonstrate that this pro apoptotic effect of CO was independent of reactive oxygen species production and involved inhibition in Fas/CD95-induced activation of the pro-survival ERK MAPK. We conclude that in contrast to other studies showing the anti-apoptotic effects of CO, Fas/CD95-induced cell death in Jurkat cells is augmented by exposure to CO and that this occurs in part via inhibition in the activation of ERK MAPK. These data begin to elucidate specific differences with regard to the effects of CO and cell death pathways and provide important and valuable insight into potential mechanisms of action. PMID- 15280388 TI - Metabolic biotinylation as a probe of supramolecular structure of the triad junction in skeletal muscle. AB - Excitation-contraction coupling in skeletal muscle involves conformational coupling between dihydropyridine receptors (DHPRs) in the plasma membrane and ryanodine receptors (RyRs) in the sarcoplasmic reticulum. However, it remains uncertain what regions, if any, of the two proteins interact with one another. Toward this end, it would be valuable to know the spatial interrelationships of DHPRs and RyRs within plasma membrane/sarcoplasmic reticulum junctions. Here we describe a new approach based on metabolic incorporation of biotin into targeted sites of the DHPR. To accomplish this, cDNAs were constructed with a biotin acceptor domain (BAD) fused to selected sites of the DHPR, with fluorescent protein (XFP) attached at a second site. All of the BAD-tagged constructs properly targeted to junctions (as indicted by small puncta of XFP) and were functional for excitation-contraction coupling. To determine whether the introduced BAD was biotinylated and accessible to avidin (approximately 60 kDa), myotubes were fixed, permeablized, and exposed to fluorescently labeled avidin. Upon expression in beta1-null or dysgenic (alpha1S-null) myotubes, punctate avidin fluorescence co-localized with the XFP puncta for BAD attached to the beta1a N- or C-terminals, or the alpha1S N-terminal or II-III loop. However, BAD fused to the alpha1S C-terminal was inaccessible to avidin in dysgenic myotubes (containing RyR1). In contrast, this site was accessible to avidin when the identical construct was expressed in dyspedic myotubes lacking RyR1. These results indicate that avidin has access to a number of sites of the DHPR within fully assembled (RyR1-containing) junctions, but not to the alpha1S C-terminal, which appears to be occluded by the presence of RyR1. PMID- 15280389 TI - Mapping sites of potential proximity between the dihydropyridine receptor and RyR1 in muscle using a cyan fluorescent protein-yellow fluorescent protein tandem as a fluorescence resonance energy transfer probe. AB - Excitation-contraction coupling in skeletal muscle involves conformational coupling between the dihydropyridine receptor (DHPR) and the type 1 ryanodine receptor (RyR1) at junctions between the plasma membrane and sarcoplasmic reticulum. In an attempt to find which regions of these proteins are in close proximity to one another, we have constructed a tandem of cyan and yellow fluorescent proteins (CFP and YFP, respectively) linked by a 23-residue spacer, and measured the fluorescence resonance energy transfer (FRET) of the tandem either in free solution or after attachment to sites of the alpha1S and beta1a subunits of the DHPR. For all of the sites examined, attachment of the CFP-YFP tandem did not impair function of the DHPR as a Ca2+ channel or voltage sensor for excitation-contraction coupling. The free tandem displayed a 27.5% FRET efficiency, which decreased significantly after attachment to the DHPR subunits. At several sites examined for both alpha1S (N-terminal, proximal II-III loop of a two fragment construct) and beta1a (C-terminal), the FRET efficiency was similar after expression in either dysgenic (alpha1S-null) or dyspedic (RyR1-null) myotubes. However, compared with dysgenic myotubes, the FRET efficiency in dyspedic myotubes increased from 9.9 to 16.7% for CFP-YFP attached to the N terminal of beta1a, and from 9.5 to 16.8% for CFP-YFP at the C-terminal of alpha1S. Thus, the tandem reporter suggests that the C terminus of alpha1S and the N terminus of beta1a may be in close proximity to the ryanodine receptor. PMID- 15280390 TI - Changes in plasma membrane properties and phosphatidylcholine subspecies of insect Sf9 cells due to expression of scavenger receptor class B, type I, and CD36. AB - In mammalian cells scavenger receptor class B, type I (SR-BI), mediates the selective uptake of high density lipoprotein (HDL) cholesteryl ester into hepatic and steroidogenic cells. In addition, SR-BI has a variety of effects on plasma membrane properties including stimulation of the bidirectional flux of free cholesterol (FC) between cells and HDL and changes in the organization of plasma membrane FC as indicated by increased susceptibility to exogenous cholesterol oxidase. Recent studies in SR-BI-deficient mice and in SR-BI-expressing Sf9 insect cells showed that SR-BI has significant effects on plasma membrane ultrastructure. The present study was designed to test the range of SR-BI effects in Sf9 insect cells that typically have very low cholesterol content and a different phospholipid profile compared with mammalian cells. The results showed that, as in mammalian cells, SR-BI expression increased HDL cholesteryl ester selective uptake, cellular cholesterol mass, FC efflux to HDL, and the sensitivity of membrane FC to cholesterol oxidase. These activities were diminished or absent upon expression of the related scavenger receptor CD36. Thus, SR-BI has fundamental effects on cholesterol flux and membrane properties that occur in cells of evolutionarily divergent origins. Profiling of phospholipid species by electrospray ionization mass spectrometry showed that scavenger receptor expression led to the accumulation of phosphatidylcholine species with longer mono- or polyunsaturated acyl chains. These changes would be expected to decrease phosphatidylcholine/cholesterol interactions and thereby enhance cholesterol desorption from the membrane. Scavenger receptor-mediated changes in membrane phosphatidylcholine may contribute to the increased flux of cholesterol and other lipids elicited by these receptors. PMID- 15280391 TI - The Caenorhabditis elegans unc-63 gene encodes a levamisole-sensitive nicotinic acetylcholine receptor alpha subunit. AB - The anthelmintic drug levamisole causes hypercontraction of body wall muscles and lethality in nematode worms. In the nematode Caenorhabditis elegans, a genetic screen for levamisole resistance has identified 12 genes, three of which (unc-38, unc-29, and lev-1) encode nicotinic acetylcholine receptor (nAChR) subunits. Here we describe the molecular and functional characterization of another levamisole resistant gene, unc-63, encoding a nAChR alpha subunit with a predicted amino acid sequence most similar to that of UNC-38. Like UNC-38 and UNC-29, UNC-63 is expressed in body wall muscles. In addition, UNC-63 is expressed in vulval muscles and neurons. We also show that LEV-1 is expressed in body wall muscle, thus overlapping the cellular localization of UNC-63, UNC-38, and UNC-29 and suggesting possible association in vivo. This is supported by electrophysiological studies on body wall muscle, which demonstrate that a levamisole-sensitive nAChR present at the C. elegans neuromuscular junction requires both UNC-63 and LEV-1 subunits. Thus, at least four subunits, two alpha types (UNC-38 and UNC-63) and two non-alpha types (UNC-29 and LEV-1), can contribute to levamisole-sensitive muscle nAChRs in nematodes. PMID- 15280392 TI - Angiopoietin-3 is tethered on the cell surface via heparan sulfate proteoglycans. AB - Angiopoietins are a family of factors that play important roles in angiogenesis, and their receptor, Tie-2 receptor tyrosine kinase, is expressed primarily by endothelial cells. Three angiopoietins have been identified so far, angiopoietin 1 (Ang-1), angiopietin-2 (Ang-2), and angiopoietin-3 (Ang-3). It has been established that Ang-1 and Tie-2 play essential roles in embryonic angiogenesis. We have demonstrated recently that, unlike Ang-2, Ang-1 binds to the extracellular matrix, which regulates the availability and activity of Ang-1 (Xu, Y., and Yu, Q. (2001) J. Biol. Chem. 276, 34990-34998). However, the role and biochemical characteristics of Ang-3 are unknown. In our current study, we demonstrated that, unlike Ang-1 and Ang-2, Ang-3 is tethered on cell surface via heparan sulfate proteoglycans (HSPGs), especially perlecan. The cell surface bound Ang-3 is capable of binding to its receptor, Tie-2; suggesting HSPGs concentrate Ang-3 on the cell surface and present Ang-3 to its receptor to elicit specific local reaction. Mutagenesis experiment revealed that the coiled-coil domain of Ang-3 is responsible for its binding to the cell surface. In addition, we demonstrated that the cell surface-bound Ang-3 but not soluble Ang-3 induces retraction and loss of integrity of endothelial monolayer, indicating the binding of Ang-3 to the cell surface via HSPGs is required for this bioactivity of Ang-3. PMID- 15280393 TI - Identification of Elongin C and Skp1 sequences that determine Cullin selection. AB - The multiprotein von Hippel-Lindau (VHL) tumor suppressor and Skp1-Cul1-F-box protein (SCF) complexes belong to families of structurally related E3 ubiquitin ligases. In the VHL ubiquitin ligase, the VHL protein serves as the substrate recognition subunit, which is linked by the adaptor protein Elongin C to a heterodimeric Cul2/Rbx1 module that activates ubiquitylation of target proteins by the E2 ubiquitin-conjugating enzyme Ubc5. In SCF ubiquitin ligases, F-box proteins serve as substrate recognition subunits, which are linked by the Elongin C-like adaptor protein Skp1 to a Cul1/Rbx1 module that activates ubiquitylation of target proteins, in most cases by the E2 Cdc34. In this report, we investigate the functions of the Elongin C and Skp1 proteins in reconstitution of VHL and SCF ubiquitin ligases. We identify Elongin C and Skp1 structural elements responsible for selective interaction with their cognate Cullin/Rbx1 modules. In addition, using altered specificity Elongin C and F-box protein mutants, we investigate models for the mechanism underlying E2 selection by VHL and SCF ubiquitin ligases. Our findings provide evidence that E2 selection by VHL and SCF ubiquitin ligases is determined not solely by the Cullin/Rbx1 module, the target protein, or the integrity of the substrate recognition subunit but by yet to be elucidated features of these macromolecular complexes. PMID- 15280394 TI - Mitochondrial tRNA import in Toxoplasma gondii. AB - Apicomplexan parasites have the smallest known mitochondrial genome. It consists of a repeated element of approximately 6-7 kb in length and encodes three mitochondrial proteins, a number of rRNA fragments, but no tRNAs. It has therefore been postulated that in apicomplexans all tRNAs required for mitochondrial translation are imported from the cytosol. To provide direct evidence for this process we have established a cell fractionation procedure allowing the isolation of defined organellar RNA fractions from the apicomplexan Toxoplasma gondii. Analysis of T. gondii total and organellar RNA by Northern hybridization showed that except for the cytosol-specific initiator tRNAMet all nucleus-encoded tRNAs tested were present in the cytosol and in the mitochondrion but not in the plastid. Thus, these results provide the first experimental evidence for mitochondrial tRNA import in apicomplexans. The only other taxon that imports the whole set of mitochondrial tRNAs are the trypanosomatids. Interestingly, the initiator tRNAMet is the only cytosol-specific tRNA in trypanosomatids, indicating that the import specificity is identical in both groups. In agreement with this, the T. gondii initiator tRNAMet remained in the cytosol when expressed in Trypanosoma brucei. However, in contrast to trypanosomatids, no thio-modifications were detected in the tRNAGln of T. gondii indicating that, unlike what is suggested in Leishmania, they are not involved in regulating import. PMID- 15280395 TI - A platelet secretion pathway mediated by cGMP-dependent protein kinase. AB - Platelet secretion (exocytosis) is critical in amplifying platelet activation, in stabilizing thrombi, and in arteriosclerosis and vascular remodeling. The signaling mechanisms leading to secretion have not been well defined. We have shown previously that cGMP-dependent protein kinase (PKG) plays a stimulatory role in platelet activation via the glycoprotein Ib-IX pathway. Here we show that PKG also plays an important stimulatory role in mediating aggregation-dependent platelet secretion and secretion-dependent second wave platelet aggregation, particularly those induced via Gq-coupled agonist receptors, the thromboxane A2 (TXA2) receptor, and protease-activated receptors (PARs). PKG I knock-out mouse platelets and PKG inhibitor-treated human platelets showed diminished aggregation dependent secretion and also showed a diminished secondary wave of platelet aggregation induced by a TXA2 analog and thrombin receptor-activating peptides that were rescued by the granule content ADP. Low dose collagen-induced platelet secretion and aggregation were also reduced by PKG inhibitors. Furthermore PKG I knockout and PKG inhibitors significantly attenuated activation of the Gi pathway that is mediated by secreted ADP. These data unveil a novel PKG-dependent platelet secretion pathway and a mechanism by which PKG promotes platelet activation. PMID- 15280396 TI - Best practice no 178. Examination of the human placenta. AB - The human placenta is an underexamined organ. The clinical indications for placental examination have no gold standards. There is also inconsistency in the histological reports and the quality is variable. There is great interobserver variability concerning the different entities. Although there are still grey areas in clinicopathological associations, a few mainstream observations have now been clarified. The histopathological examination and diagnosis of the placenta may provide crucial information. It is possible to highlight treatable maternal conditions and identify placental or fetal conditions that can be recurrent or inherited. To achieve optimal benefit from placental reports, it is essential to standardise the method of placenta examination. This article summarises the clinical indications for placenta referral and the most common acknowledged clinicopathological correlations. PMID- 15280397 TI - A review of the heritability of idiopathic nephrolithiasis. AB - Familial aggregations of nephrolithiasis were already noted in the early 19th century and over the intervening years there has been gradual progression in classifying the familial forms of nephrolithiasis. To date, there are at least 10 different monogenic conditions where those affected have a predisposition to nephrolithiasis. However, all of these rare conditions probably account for less than 2% of renal stone formers. This review, rather than considering these clearly defined disorders, concentrates on research into the broad band of stone formers who have a propensity to nephrolithiasis without an obvious discrete genetic basis. PMID- 15280398 TI - Genetic pathways to melanoma tumorigenesis. AB - The incidence of cutaneous malignant melanomas is growing faster than that of any other cancer and therefore posing a major heath threat worldwide. In melanocytic skin tumours, the feasibility of correlating a specific pathological stage with a corresponding genetic alteration provides a remarkable opportunity to study the multistep tumorigenesis model. This multistep melanoma tumorigenesis is best described as a continuum of transformation of the melanocytes, melanocytic dysplasia, and melanoma formation. These steps involve genotypic alterations including loss of tumour suppressor genes, microsatellite instability, and alterations of the mismatch repair system. This review seeks to examine melanoma tumorigenesis based on these genetic changes. PMID- 15280399 TI - Sertoli cell nodules in the undescended testis: a histochemical, immunohistochemical, and ultrastructural study of hyaline deposits. AB - AIMS: To document the morphology, immunohistochemical staining properties, and ultrastructural features of hyaline material in Sertoli cell nodules of undescended testis and contrast them with those of sex cord tumour with annular tubules (SCTAT), which is histologically similar. To highlight the need to distinguish these nodules from other Sertoli cell hyperplasias, such as intratubular Sertoli cell proliferations, which occur in specific clinical contexts. MATERIALS/METHODS: A retrospective study of 46 orchidectomy specimens from cryptorchid testes, 27 of which contained Sertoli cell nodules. Special histochemical stains, immunohistochemical stains for type IV collagen and fibronectin, and ultrastructural examination of the hyaline material were performed using tissue from paraffin wax embedded tissue blocks. RESULTS: The hyaline deposits in SCTAT and Sertoli cell nodules had similar staining patterns periodic acid Schiff (PAS) and PAS-diastase positivity with variable staining of Martius scarlet blue and Masson trichrome. Type IV collagen immunoreactivity was seen in hyaline areas, although fibronectin was negative. Electron microscopy of hyaline areas confirmed a compact matrix identical to components of the basement membrane in the adjacent seminiferous tubules. CONCLUSION: This study describes an unusual form of Sertoli cell proliferation in undescended testes, which must be distinguished from Sertoli cell tumours and other forms of proliferation. In addition, the hyaline material within Sertoli cell nodules in the cryptorchid testis is histochemically, immunohistochemically, and ultrastructurally consistent with both matrix and fibrous components of seminiferous tubule basement membranes. Increased production of basement membrane material, with subsequent invagination into tubules, is the most likely origin of this material. PMID- 15280400 TI - Secular trends of nocardia infection over 15 years in a tertiary care hospital. AB - AIMS: To assess the incidence of nocardia infection over 15 years in a tertiary care hospital. METHODS: Over a 15 year period, Nocardia spp were isolated from 20 patients hospitalised at the Geneva University Hospitals, Switzerland. RESULTS: Sixteen patients had one or more underlying conditions. The median time between symptom onset and diagnosis was 30 days. The most common initial unconfirmed diagnosis was pulmonary tuberculosis (four). The lung was involved in 16 cases, followed by the central nervous system (two) and skin (two); one patient had disseminated infection. The most common species identified was N asteroides. In vitro susceptibility testing was performed on 14 of 20 strains. All strains were susceptible to imipenem and amikacin. Initial treatment with trimethoprim/sulfamethoxazole (TMP/SMX) was started in 14 patients, although five patients had to be switched to another treatment because of side effects or lack of efficacy. A cure was observed in 15 patients, death in three, and relapse or complications in two. CONCLUSIONS: Nocardiosis can become a severe infection and mainly affects profoundly immunocompromised patients. Differential diagnosis often delays the time to diagnosis, which worsens the outcome. New diagnostic tools, such as the polymerase chain reaction, could provide more rapid and reliable results. TMT/SMX was the most commonly prescribed treatment, but needed to be changed for another treatment because of side effects or lack of efficacy in a considerable proportion of patients. Imipenem should be used as an alternative treatment for severely ill patients, and the sulfa combination for less severe infections. PMID- 15280401 TI - Mucin expression in pleomorphic adenoma of salivary gland: a potential role for MUC1 as a marker to predict recurrence. AB - BACKGROUND: Pleomorphic adenoma of the salivary gland (PA) is essentially a benign neoplasm. However, patients with recurrent PA are difficult to manage. There are rare reports on useful immunohistochemical markers to detect a high risk of recurrence when the primary lesions are resected. AIMS: To find a new marker to predict the recurrence of PA. METHODS: Primary lesions of PA were collected from nine patients showing subsequent recurrence and from 40 patients without recurrence during at least 10 years of follow up of the disease. Paraffin wax embedded tumour samples of the two groups were examined for the expression profiles of MUC1 (differentially glycosylated forms), MUC2, MUC4, MUC5AC, and MUC6 using immunohistochemistry. Several clinicopathological factors were also examined. RESULTS: In univariate analysis of the factors examined, MUC1/DF3 high expression (more than 30% of the neoplastic cells stained) in the primary lesions was seen more frequently in patients with recurrence (four of nine) than in those without recurrence (three of 40; p = 0.011). Larger tumour size (more than 3.0 cm) of the primary PA was also a significant (p = 0.035) risk factor for the recurrence of PA. In multivariate analysis, only high expression of MUC1/DF3 was found to be a significant independent risk factor for the recurrence of PA (p = 0.021). CONCLUSIONS: Expression of MUC1/DF3 in PA is a useful marker to predict its recurrence. Those patients with PA showing positive MUC1/DF3 expression should be followed up carefully. PMID- 15280402 TI - Topographical localisation of cagA positive and cagA negative Helicobacter pylori strains in the gastric mucosa; an in situ hybridisation study. AB - BACKGROUND: The cagA gene is a marker for the presence of the cag pathogenicity island, and the presence of cagA positive strains of Helicobacter pylori can identify individuals with a higher risk of developing gastrointestinal diseases. AIMS: To study the interaction between H. pylori cagA(+) and cagA(-) strains and the gastric mucosa. METHODS: Patients with H. pylori associated gastritis and peptic ulcers were studied. Biopsies were obtained from the antrum, corpus, fundus, and incisura for H pylori culture, and for in situ hybridisation studies. From each biopsy, multiple single H. pylori colonies were isolated and propagated for DNA isolation, and cagA was detected by the polymerase chain reaction (PCR). For in situ detection of H. pylori an oligonucleotide specific for an H. pylori common antigen and an oligonucleotide specific for cagA were used as probes. Biotinylated probes were incubated with biopsy sections, developed with streptavidin-horseradish peroxidase, and amplified with the tyramide system. RESULTS: PCR results for cagA in isolated colonies confirmed the in situ hydridisation studies. In situ hybridisation identified cagA(+) bacteria in patients with cagA(+) isolates; cagA(-) bacteria in patients with cagA(-) isolates, and cagA(+) and cagA (-) bacteria in patients with both cagA(+) and cagA(-) isolates. CagA(-) bacteria usually colonised the mucous gel or the apical epithelial surface, whereas cagA(+) bacteria colonised the immediate vicinity of epithelial cells or the intercellular spaces. CONCLUSIONS: These results document a different in vivo interaction between H. pylori cagA(+) or cagA(-) strains and the gastric mucosa. PMID- 15280403 TI - Expression of vascular endothelial growth factor D is associated with hypoxia inducible factor (HIF-1alpha) and the HIF-1alpha target gene DEC1, but not lymph node metastasis in primary human breast carcinomas. AB - BACKGROUND: Vascular endothelial growth factor D (VEGF-D) induces angiogenesis and lymphangiogenesis. Nodal metastasis is recognised as a powerful prognostic marker in breast carcinoma, but the molecular mechanisms underlying this process are unknown. Although it has been suggested that VEGF-D may regulate nodal metastasis, this is based largely on animal models, its role in human disease being unclear. AIMS: To measure the pattern and degree of VEGF-D protein expression in normal and neoplastic human breast tissues. METHODS: The pattern and degree of VEGF-D expression was measured in normal tissue and invasive carcinomas, and expression was correlated with clinicopathological parameters, hypoxia markers, and survival. Because other VEGF family members are affected by oestrogen, whether VEGF-D is regulated by oestrogen in breast cancer cell lines was also assessed. RESULTS: VEGF-D was significantly positively associated with hypoxia inducible factor (HIF-1alpha) (p = 0.03) and the HIF-1alpha regulated gene DEC1 (p = 0.001), but not lymph node status, the number of involved lymph nodes, patient age, tumour size, tumour grade, lymphovascular invasion, oestrogen receptor, progesterone receptor, c-erb-B2, or tumour histology (all p>0.05). There was no significant relation between tumour VEGF-D expression and relapse free (p = 0.78) or overall (p = 0.94) survival. VEGF-D expression was enhanced by oestrogen in MCF-7 and T47D breast cancer cells, and was blocked by hydroxytamoxifen. CONCLUSION: These findings support a role for hypoxia and oestrogen induced VEGF-D in human breast cancer and also suggest that tamoxifen and related oestrogen antagonists may exert some of their antitumour effects through the abrogation of VEGF-D induced function. PMID- 15280404 TI - Intraepithelial lymphocytes in the villous tip: do they indicate potential coeliac disease? AB - BACKGROUND: The counting of intraepithelial lymphocytes (IELs) in the villous tips of architecturally normal small bowel biopsy specimens was proposed as a method to measure mucosal infiltration in gluten sensitive patients. AIMS: To apply this straightforward method in duodenal biopsy specimens from patients affected by potential coeliac disease (PCD) to verify whether it can discriminate these patients from controls. METHODS: Paraffin wax embedded duodenal sections from 11 patients affected by PCD were stained with an antihuman CD3 antibody. Sections from 19 patients affected by treated coeliac disease (TCD) and 17 patients in whom coeliac disease was excluded were stained with the same antibody to serve as controls. The slides were examined blindly. IELs/20 enterocytes in five randomly chosen villous tips were counted. Patients affected by PCD were all on a gluten containing diet. They had an architecturally normal duodenal mucosa and were positive for endomysial antibody. Both TCD and non-coeliac controls were negative for endomysial antibody. RESULTS: The mean villous tip IEL scores were 4.6 (SD, 1.5; range, 1.4-7.8) in non-coeliac controls, 7.9 (SD, 4.0; range, 2.0 18.6) in TCD, and 9.2 (SD, 4.7; range, 5.8-21.8) in patients with PCD. The difference between PCD and non-coeliac controls was significant. CONCLUSIONS: This is a very simple and sufficiently reliable method to count IELs. In patients with an architecturally normal duodenal mucosa, the IEL count in villous tips helps to distinguish between patients with PCD and non-coeliac controls. PMID- 15280405 TI - Proliferative activity in postmenopausal endometrium: the lurking potential for giving rise to an endometrial adenocarcinoma. AB - AIMS: To investigate proliferation in disease free postmenopausal endometrium and that harbouring endometrial adenocarcinoma-is there a dynamic, yet lurking, potential for atrophic endometrium to give rise to endometrial adenocarcinoma? MATERIAL/METHODS: The study comprised 84 disease free endometria from asymptomatic postmenopausal women who had undergone hysterectomy for prolapse, and 50 endometrioid cell type endometrial adenocarcinomas with adjacent uninvolved postmenopausal endometrium. The non-neoplastic tissues were separated histologically into active, inactive, and mixed forms, although only the first two categories were studied immunohistochemically for oestrogen and progesterone receptors (ERs, PRs), epidermal growth factor receptor (EGFR), Ki-67, and angiogenic activity. RESULTS: All postmenopausal endometria were atrophic, but only 42 were inactive; of the remaining samples, 22 were weakly proliferative and 20 were mixed active and inactive. In contrast, the non-neoplastic component of 43 of the 50 endometrial adenocarcinomas examined was of the active form; four specimens were of the pure and 39 of the mixed form. Interestingly, high ER and PR expression was seen in active and inactive endometria, but only the former were EGFR positive and had high proliferative (Ki-67) and angiogenic activity. A similar trend was also shown by the non-neoplastic atrophic endometrium adjacent to endometrial adenocarcinoma. CONCLUSIONS: At least half of the disease free postmenopausal atrophic endometria show a weak proliferative pattern, either diffuse or focal, probably as a response to continuous low level oestrogenic stimulation. These tissues have a latent, although very small, carcinogenic potential, as demonstrated by the immunohistochemical profile and their frequent association with adjacent endometrial adenocarcinoma. PMID- 15280406 TI - Node retrieval in axillary lymph node dissections: recommendations for minimum numbers to be confident about node negative status. AB - AIMS: To determine the minimum number of lymph nodes needed in an axillary lymph node dissection (ALND) specimen to be confident that the axilla is free from metastases. METHODS: The Edinburgh Breast Unit selects patients with large and high grade tumours for ALND; 609 consecutive ALNDs performed between October 1999 and December 2002 were reviewed. Full data about the underlying invasive breast cancer were available for 520 patients. Data were collected regarding number of positive nodes and total number of nodes collected, tumour size and grade, and presence of lymphovascular invasion. RESULTS: Axillary node metastases were seen in 64% of patients. The mean number of positive nodes found was 3.56, with a mean of 17.9 nodes collected. The highest proportion of patients with lymph node metastases were in the group with 16-20 nodes recovered/specimen (68%); specimens with >20 nodes recovered did not have a higher rate of nodal involvement. There was a significant difference between the proportion of metastasis positive specimens in those with 1-15 nodes recovered (58.5%) and those with 16 or more recovered (69.1%). A linear association test showed a direct correlation between the number of nodes collected and presence of node metastasis (p = 0.0005). CONCLUSIONS: Although there is no minimum number of nodes that should be recovered in an ALND specimen, 16 nodes should be regarded as a target to ensure a high level of confidence that the nodes are negative. Node positivity in an ALND specimen appears to obey the law of diminishing returns. PMID- 15280407 TI - Gene expression profiles of giant hairy naevi. AB - BACKGROUND: Congenital neomelanocytic naevi appear in nearly 1% of newborns. Giant hairy naevi (GHN) are uncommon lesions covering large areas of the body. They are of concern because they have the potential to transform into malignant melanomas. AIMS: To describe gene expression profiles of GHN and nearby normal skin from patients with GHN and normal control skin (from patients with cleft lip/palate). METHODS: Tissues from three patients with GHN and two normal controls were studied for differences in gene expression profiles. Total RNA was isolated from normal skin near the hairy naevus, GHN, and skin from normal controls. The RNA samples were subjected to probe labelling, hybridisation to chips, and image acquisition according to the standard Affymetrix protocol. RESULTS: There were 227 genes affected across all samples, as determined by DNA microarray analysis. There was increased expression of 22 genes in GHN compared with nearby normal skin. Decreased expression was noted in 73 genes. In addition, there was increased expression of 36 genes in normal skin near GHN compared with normal control skin, and decreased expression of five genes. Categories of genes affected were those encoding structural proteins, proteins related to developmental processes, cell death associated proteins, transcription factors, growth factors, stress response modulators, and collagen associated proteins. Changes in mRNA expression were checked by reverse transcription polymerase chain reaction. CONCLUSIONS: Genetic profiles of GHN may provide insight into their pathogenesis, including their potential for malignant transformation. Such information may be useful in improving the understanding and management of these lesions. PMID- 15280408 TI - The relation between bone marrow angiogenesis and the proliferation index Ki-67 in multiple myeloma. AB - AIM: Angiogenesis correlates with disease progression in various haematological malignancies. This study investigated the association between microvascular density (MVD) and the Ki-67 proliferation index (Ki-67 PI), bone marrow infiltration, and C reactive protein (CRP) in patients with multiple myeloma. METHODS: Bone marrow MVD was examined in 44 biopsies at diagnosis and 15 in plateau phase by immunostaining the endothelial cells with a monoclonal antibody to CD34. The Ki-67 PI was evaluated by a double immunostaining technique using the monoclonal antibodies MIB-1 and CD38. RESULTS: MVD, Ki-67 PI, bone marrow infiltration, and CRP were significantly higher in pretreatment patients than in controls and decreased in patients achieving plateau phase. MVD significantly correlated with Ki-67 PI and infiltration, and Ki-67 correlated with infiltration. CONCLUSION: In multiple myeloma, apart from being a marker of proliferative activity, Ki-67 is also associated with bone marrow angiogenesis and tumour burden. PMID- 15280409 TI - Aberrant epithelial expression of trefoil family factor 2 and mucin 6 in Helicobacter pylori infected gastric antrum, incisura, and body and its association with antralisation. AB - AIMS: To determine gastric expression of trefoil family factor 2 (TFF2) and MUC6 in Helicobacter pylori positive and negative subjects, and its association with antralisation at the gastric incisura. METHODS: Gastric biopsies from the antrum, incisura, and body of 76 dyspeptic patients without ulcers were used for the determination of H. pylori infection, histological changes, and epithelial TFF2 and MUC6 expression. RESULTS: In the foveola, the rates of TFF2 and MUC6 immunostaining were greater in H. pylori infected (n = 27) than in uninfected patients (n = 49) at the antrum (59.3% v 4.1% for TFF2 and 63.0% v 4.1% for MUC6; both p < 0.001) and incisura (44.4% v 2.0% for TFF2 and 48.1% v 0% for MUC6; both p < 0.001). In the deeper glands, the rates were also greater in H. pylori infected than in uninfected patients at the incisura (85.2% v 22.4% for both TFF2 and MUC6; p < 0.001). Antral-type mucosa was present at the incisura in 28 of the 76 patients. TFF2 and MUC6 expression in the foveola and deeper glands was significantly associated with antral-type mucosa, independent of H. pylori status. CONCLUSIONS: Helicobacter pylori infection increases the expression of TFF2 and MUC6 in the gastric epithelium. Aberrant TFF2 and MUC6 expression is associated with antralisation of gastric incisura. PMID- 15280410 TI - Microvessel density and clinicopathological characteristics in hepatitis C virus and hepatitis B virus related hepatocellular carcinoma. AB - AIMS: To compare intratumorous microvessel density (MVD) and clinicopathological features in two different groups of hepatocellular carcinoma (HCC), namely: hepatitis B virus (HBV) related HCC (B-HCC) and HCV related HCC (C-HCC). METHODS: Fifty consecutive cases each of B-HCC and of C-HCC were studied. Microvessel numbers were assessed by staining for the antigen CD34; in each case, three areas with the highest numbers of microvessels were counted in both the intratumorous and the surrounding non-tumorous tissue; the mean value represented the final MVD. RESULTS: Patients with B-HCC were significantly younger than those with C HCC (mean age, 60.1 (SD, 4.1) v 66.4 (4.3) years); no significant differences were seen for sex or Child's class distribution. The tumour diameter was larger in B-HCCs than in C-HCCs (mean, 5.6 (SD, 1.8) v 3.8 (1.8) cm). Tumour microsatellite formation was significantly higher in C-HCCs (12 v 4 cases). No differences were found for histological subtype, degree of differentiation, tumour encapsulation, and vascular invasion. The mean MVD value was significantly higher in tumorous (mean, 54 (SD, 13.8) v 38 (8.9)) and in the surrounding non tumorous liver tissue (mean, 15 (SD, 4.3) v 7 (3.1)) of C-HCCs. CONCLUSIONS: C HCCs present as smaller tumours in older patients, with a higher incidence of tumour microsatellite formation and higher MVD values both in the tumorous and the non-tumorous areas, suggesting a link between HCV infection, angiogenesis, and hepatocarcinogenesis. PMID- 15280411 TI - Aberrant cellular retinol binding protein 1 (CRBP1) gene expression and promoter methylation in prostate cancer. AB - AIMS: Retinoids are involved in cell growth, differentiation, and carcinogenesis. Their effects depend on cytosolic transport and binding to nuclear receptors. CRBP1 encodes a protein involved in this process. Because altered CRBP1 expression and promoter hypermethylation occur in several tumours, these changes were investigated in prostate tumorigenesis. METHODS: The CRBP1 promoter was assessed by methylation specific polymerase chain reaction on tissue samples from 36 radical prostatectomy specimens (paired normal tissue, adenocarcinoma, and high grade prostatic intraepithelial neoplasia (HGPIN)), 32 benign prostatic hyperplasias (BPHs), and 13 normal prostate tissue samples from cystoprostatectomies. Methylation of DNA extracted from microdissected tissue was examined blindly. CRBP1 expression was assessed by immunohistochemistry on formalin fixed, paraffin wax embedded tissue. RESULTS: Loss of CRBP1 expression was seen in 15 of 36 adenocarcinomas and 18 of 36 HGPINs. Fifteen adenocarcinomas and nine HGPINs showed overexpression, whereas the remainder showed normal expression. BPH displayed normal expression. No significant associations were found between CRBP1 expression and Gleason score or stage. CRBP1 promoter hypermethylation was found in 17 of 36 adenocarcinomas, three of 35 HGPINs, one of 36 normal prostate tissues from the same patients, none of 32 BPHs, and none of 13 normal prostate tissues from cystoprostatectomies. Loss of expression and hypermethylation of CRBP1 were not significantly associated. CONCLUSIONS: Altered CRBP1 expression and hypermethylation are common in prostate carcinoma, although CRBP1 hypermethylation is not an early event in tumorigenesis. Moreover, both adenocarcinoma and HGPIN show frequent CRBP1 overexpression. The molecular mechanisms underlying altered CRBP1 expression in prostate cancer deserve further study. PMID- 15280412 TI - Videos have a role in postgraduate necropsy education. AB - AIMS: This is the first study to investigate the usefulness of structured, scripted videos as an adjunct to the mortuary based training of histopathology trainees in necropsy techniques. METHODS: Four structured and scripted videos describing aspects of necropsy health and safety, evisceration, general dissection techniques, specialist dissection techniques, and reconstruction were shown to histopathology trainees attending the 2001 University of Sheffield short course on the autopsy. Delegates who agreed to participate in the study were asked to complete a short questionnaire seeking Likert-type and free text responses concerning the usefulness of the videos in postgraduate necropsy training. Free text responses were analysed using a themed content analysis. RESULTS: All 38 delegates who viewed the videos agreed to participate in the study. Of these, 35 found the videos enjoyable and 34 found them interesting. Thirty one felt the videos enhanced their learning experience. Advantages of the videos included the ability to learn about specialist techniques rarely encountered in the mortuary, the ability to teach large numbers of students at once, allowing students to learn at their own pace, and as a tool for revision. Repetition between the videos, a lack of interactivity, and a lack of sufficient detail on general necropsy techniques were felt by participants to be the principal disadvantages of this teaching tool. CONCLUSIONS: Videos are an acceptable teaching tool for students. They have a valuable role to play as an adjunct to dissection in teaching junior histopathology trainees about specialist necropsy dissection techniques. PMID- 15280413 TI - Pineal yolk sac tumour with a solid pattern: a case report in a Chinese adult man with Down's syndrome. AB - Intracranial germ cell tumours are rare. The incidence of primary intracranial yolk sac tumour is even more uncommon, with only two reported cases being associated with Down's syndrome in the English literature. This report details the findings of yolk sac tumour in the pineal region affecting a 22 year old Chinese man with Down's syndrome. Histology revealed yolk sac tumour with only a solid pattern, potentially mimicking the more common germinoma in the pineal region. No other germ cell components were identified. This is the third report of intracranial yolk sac tumour manifesting in a patient with trisomy 21. The pathology of this tumour and its differential diagnoses are discussed. PMID- 15280414 TI - Synchronous ileal and colonic adenocarcinomas associated with Crohn's disease: report of a case with a focus on genetic alterations and carcinogenesis. AB - Patients with Crohn's disease have an increased risk of developing intestinal tumours. However, the carcinogenic mechanisms remain poorly understood. To address this question, this report describes an unusual case of Crohn's disease complicated by synchronous small intestinal and colonic adenocarcinomas. Genetic events in both the tumours and their adjacent mucosae were evaluated and the tumorigenesis of these cancers is discussed. PMID- 15280415 TI - Extensive psammomatous calcification of the uterus and cervix associated with a uterine serous carcinoma. AB - This report describes a uterine serous carcinoma with bilateral ovarian metastasis, which was associated with widespread extensive psammomatous calcification of the uterine leiomyomata, the myometrium, and the cervical stroma. These psammoma bodies were not associated with tumour or epithelial elements. This psammomatous calcification is rare, with no previous reports of similar cases. The presence of psammoma bodies is probably related to the serous carcinoma, raising the possibility that psammoma body formation in serous carcinomas is the result of a factor secreted locally by the tumour, rather than the widely held theory that their formation is secondary to necrosis, with subsequent dystrophic calcification within a papillary neoplasm. PMID- 15280416 TI - Increased neutrophil apoptosis in chronic fatigue syndrome. AB - BACKGROUND/AIMS: Many patients with chronic fatigue syndrome (CFS) have symptoms that are consistent with an underlying viral or toxic illness. Because increased neutrophil apoptosis occurs in patients with infection, this study examined whether this phenomenon also occurs in patients with CFS. METHODS: Apoptosis was assessed in patients with CFS in conjunction with concentrations of the anti inflammatory cytokine, transforming growth factor beta1 (TGFbeta1). RESULTS: The 47 patients with CFS had higher numbers of apoptotic neutrophils, lower numbers of viable neutrophils, increased annexin V binding, and increased expression of the death receptor, tumour necrosis factor receptor-I, on their neutrophils than did the 34 healthy controls. Patients with CFS also had raised concentrations of active TGFbeta1 (p < 0.005). CONCLUSIONS: These findings provide new evidence that patients with CFS have an underlying detectable abnormality in their immune cells. PMID- 15280417 TI - Secretory carcinoma of the male breast. PMID- 15280418 TI - Gastric intestinal metaplasia. PMID- 15280419 TI - CD25(+)CD4(+) regulatory T cells from the peripheral blood of asymptomatic HIV infected individuals regulate CD4(+) and CD8(+) HIV-specific T cell immune responses in vitro and are associated with favorable clinical markers of disease status. AB - Human immunodeficiency virus (HIV) disease is associated with loss of CD4(+) T cells, chronic immune activation, and progressive immune dysfunction. HIV specific responses, particularly those of CD4(+) T cells, become impaired early after infection, before the loss of responses directed against other antigens; the basis for this diminution has not been elucidated fully. The potential role of CD25(+)CD4(+) regulatory T cells (T reg cells), previously shown to inhibit immune responses directed against numerous pathogens, as suppressors of HIV specific T cell responses was investigated. In the majority of healthy HIV infected individuals, CD25(+)CD4(+) T cells significantly suppressed cellular proliferation and cytokine production by CD4(+) and CD8(+) T cells in response to HIV antigens/peptides in vitro; these effects were cell contact dependent and IL 10 and TGF-beta independent. Individuals with strong HIV-specific CD25(+) T reg cell function in vitro had significantly lower levels of plasma viremia and higher CD4(+): CD8(+) T cell ratios than did those individuals in whom this activity could not be detected. These in vitro data suggest that CD25(+)CD4(+) T reg cells may contribute to the diminution of HIV-specific T cell immune responses in vivo in the early stages of HIV disease. PMID- 15280420 TI - Deletion of the nucleotide excision repair gene Ercc1 reduces immunoglobulin class switching and alters mutations near switch recombination junctions. AB - The structure-specific endonuclease ERCC1-XPF is an essential component of the nucleotide excision DNA repair pathway. ERCC1-XPF nicks double-stranded DNA immediately adjacent to 3' single-strand regions. Substrates include DNA bubbles and flaps. Furthermore, ERCC1 interacts with Msh2, a mismatch repair (MMR) protein involved in class switch recombination (CSR). Therefore, ERCC1-XPF has abilities that might be useful for antibody CSR. We tested whether ERCC1 is involved in CSR and found that Ercc1(-)(/)(-) splenic B cells show moderately reduced CSR in vitro, demonstrating that ERCC1-XPF participates in, but is not required for, CSR. To investigate the role of ERCC1 in CSR, the nucleotide sequences of switch (S) regions were determined. The mutation frequency in germline Smicro segments and recombined Smicro-Sgamma3 segments cloned from Ercc1(-)(/)(-) splenic B cells induced to switch in culture was identical to that of wild-type (WT) littermates. However, Ercc1(-)(/)(-) cells show increased targeting of the mutations to G:C bp in RGYW/WRCY hotspots and mutations occur at sites more distant from the S-S junctions compared with WT mice. The results indicate that ERCC1 is not epistatic with MMR and suggest that ERCC1 might be involved in processing or repair of DNA lesions in S regions during CSR. PMID- 15280421 TI - Compromised function of regulatory T cells in rheumatoid arthritis and reversal by anti-TNFalpha therapy. AB - Regulatory T cells have been clearly implicated in the control of disease in murine models of autoimmunity. The paucity of data regarding the role of these lymphocytes in human autoimmune disease has prompted us to examine their function in patients with rheumatoid arthritis (RA). Regulatory (CD4(+)CD25(+)) T cells isolated from patients with active RA displayed an anergic phenotype upon stimulation with anti-CD3 and anti-CD28 antibodies, and suppressed the proliferation of effector T cells in vitro. However, they were unable to suppress proinflammatory cytokine secretion from activated T cells and monocytes, or to convey a suppressive phenotype to effector CD4(+)CD25(-) T cells. Treatment with antitumor necrosis factor alpha (TNFalpha; Infliximab) restored the capacity of regulatory T cells to inhibit cytokine production and to convey a suppressive phenotype to "conventional" T cells. Furthermore, anti-TNFalpha treatment led to a significant rise in the number of peripheral blood regulatory T cells in RA patients responding to this treatment, which correlated with a reduction in C reactive protein. These data are the first to demonstrate that regulatory T cells are functionally compromised in RA, and indicate that modulation of regulatory T cells by anti-TNFalpha therapy may be a further mechanism by which this disease is ameliorated. PMID- 15280422 TI - RIP links TLR4 to Akt and is essential for cell survival in response to LPS stimulation. AB - Receptor-interacting protein (RIP) has been reported to associate with tumor necrosis-associated factor (TRAF)2 and TRAF6. Since TRAF2 and TRAF6 play important roles in CD40 signaling and TRAF6 plays an important role in TLR4 signaling, we examined the role of RIP in signaling via CD40 and TLR4. Splenocytes from RIP(-/-) mice proliferated and underwent isotype switching normally in response to anti-CD40-IL-4 but completely failed to do so in response to LPS-IL-4. However, they normally up-regulated TNF-alpha and IL-6 gene expression and CD54 and CD86 surface expression after LPS stimulation. RIP(-/-) splenocytes exhibited increased apoptosis and impaired Akt phosphorylation after LPS stimulation. These results suggest that RIP is essential for cell survival after TLR4 signaling and links TLR4 to the phosphatidylinositol 3 kinase-Akt pathway. PMID- 15280423 TI - Suppressor T cells in human diseases. AB - Although central and peripheral tolerance are important for the regulation of human immune responses to self- and microbial antigens, an important role of suppressor CD4(+) CD25(+) T cells is suggested from the recent investigations of human autoimmune diseases and HIV. These new data provide increasing evidence that altered function of CD4(+) CD25(+) T cells may be an important factor in a wide range of human inflammatory and infectious diseases. PMID- 15280424 TI - Survivin dynamics increases at centromeres during G2/M phase transition and is regulated by microtubule-attachment and Aurora B kinase activity. AB - The inhibitor of apoptosis protein survivin is implicated in two key biological events: in the control of cell proliferation and in the regulation of cell lifespan. Although the details of mitotic roles of survivin are unclear, the protein appears to modulate microtubule function and might participate in regulating the spindle checkpoint. Survivin physically associates with Aurora B, a serine-threonine kinase involved in microtubule attachment to centromeres and regulation of chromosome segregation. Here we have examined the dynamics and localization of a survivin-GFP chimera using high-resolution fluorescence microscopy and photobleaching. Survivin forms a bi-partite structure at the inner centromere that undergoes significant stretching during mitosis. Photobleaching experiments revealed marked changes in rates of survivin turnover at centromeres. These were regulated by stage of the cell cycle, microtubule attachment, and Aurora B kinase activity. We hypothesize that changes in the turnover of survivin at centromeres influence the stability of kinetochore-microtubule attachment and signaling of the spindle checkpoint. PMID- 15280425 TI - Release of a membrane-bound death domain by gamma-secretase processing of the p75NTR homolog NRADD. AB - Neurotrophin receptor alike death domain protein (NRADD) is a death-receptor-like protein with a unique ectodomain and an intracellular domain homologous to p75(NTR). Expression of NRADD results in apoptosis, but only in certain cell types. This paper characterizes the expression and proteolytic processing of the mature 55 kDa glycoprotein. N-terminally truncated NRADD is processed by a gamma secretase activity that requires presenilins and has the same susceptibility to gamma-secretase inhibitors as the secretion of amyloid beta (A beta). The ectodomain of endogenous NRADD is shed by activation of metalloproteinases. Inhibitor studies provide evidence that NRADD is cleaved in two steps typical of regulated intramembrane proteolysis (RIP). Inhibition of gamma-secretase abrogates both the production of the soluble intracellular domain of NRADD and the appearance of NRADD in subnuclear structures. Thus, solubilized death domains with close homology to p75(NTR) might have a nuclear function. Furthermore, presenilin deficiency leads to abnormally glycosylated NRADD and overexpression of presenilin 2 inhibits NRADD maturation, which is dependent on the putative active site residue D366 but not on gamma-secretase activity. Our results demonstrate that NRADD is an additional gamma-secretase substrate and suggest that drugs against Alzheimer's disease will need to target gamma-secretase in a substrate-specific manner. PMID- 15280426 TI - Cdk5 regulates activation and localization of Src during corneal epithelial wound closure. AB - Recent studies have shown that Cdk5, a member of the cyclin-dependent-kinase family, regulates adhesion and migration in a mouse corneal epithelial cell line. Here, we extend these findings to corneal wound healing in vivo and examine the mechanism linking Cdk5 to cytoskeletal reorganization and migration. Cdk5 was overexpressed in the corneal epithelium of transgenic mice under control of the ALDH3 promoter. Elevated Cdk5 expression retarded corneal debridement wound closure in these animals and suppressed remodeling of the actin cytoskeleton. Conversely, the Cdk5 inhibitor, olomoucine, accelerated debridement wound healing in organ cultured eyes of normal mice, caused migrating cells to separate from the epithelial cell sheet, and increased the level of activated Src(pY416) along the wound edge. To explore the relationship between Cdk5 and Src in greater detail, we examined scratch-wounded cultures of corneal epithelial cells. Src was activated in cells along the wound edge and blocking this activation with the Src kinase inhibitor, PP1, inhibited wound closure by 85%. Inhibiting Cdk5 activity with olomoucine or a dominant negative construct, Cdk5T33, increased the concentration of Src(pY416), shifted its subcellular localization to the cell periphery and enhanced wound closure. Cdk5(pY15), an activated form of Cdk5, also appeared along the wound edge. Inhibiting Src activity with PP1 blocked the appearance of Cdk5(pY15), suggesting that Cdk5 phosphorylation is Src dependent. Cdk5 and Src co-immunoprecipitated from scratch-wounded cultures, demonstrating that both kinases are part of an intracellular protein complex. These findings indicate that Cdk5 exerts its effects on cell migration during corneal epithelial wound healing by regulating the activation and localization of Src. PMID- 15280427 TI - Ribosome-translocon complex mediates calcium leakage from endoplasmic reticulum stores. AB - Under resting conditions, the endoplasmic reticulum (ER) intraluminal free calcium concentration ([Ca(2+)](ER)) reflects a balance between active uptake by Ca(2+)-ATPases and passive efflux via 'leak channels'. Despite their physiological importance and ubiquitous leak pathway mechanism, very little is known about the molecular nature of these channels. As it has been suggested that the open translocon pore complex of the ER is permeable to ions and neutral molecules, we hypothesized that the ribosome-bound translocon would be permeable to calcium after treatment with puromycin, a translation inhibitor that specifically releases polypeptide chains. At this time, the translocon channel is left open. We measured the fluctuations in cytoplasmic and luminal calcium concentrations using fluorescent dyes (fura-2 and magfura-2, respectively). The calcium release induced by thapsigargin (a Ca(2+)-ATPase inhibitor) was lower after puromycin treatment. Puromycin also reduced the [Ca(2+)](ER) level when perfused into the medium, but was ineffective after anisomycin pre-treatment (an inhibitor of the peptidyl transferase). Puromycin had a similar effect in the presence of heparin and ryanodine. This puromycin-evoked [Ca(2+)](ER) decrease was specific to the translocon. We conclude that the translocon complex is a major calcium leak channel. This work reveals a new role for the translocon which is involved in the control of the [Ca(2+)](ER) and could therefore supervise many physiological processes, including gene expression and apoptosis. PMID- 15280428 TI - Compartment-specific perturbation of protein handling activates genes encoding mitochondrial chaperones. AB - Protein folding in the mitochondria is assisted by nuclear-encoded compartment specific chaperones but regulation of the expression of their encoding genes is poorly understood. We found that the mitochondrial matrix HSP70 and HSP60 chaperones, encoded by the Caenorhabditis elegans hsp-6 and hsp-60 genes, were selectively activated by perturbations that impair assembly of multi-subunit mitochondrial complexes or by RNAi of genes encoding mitochondrial chaperones or proteases, which lead to defective protein folding and processing in the organelle. hsp-6 and hsp-60 induction was specific to perturbed mitochondrial protein handling, as neither heat-shock nor endoplasmic reticulum stress nor manipulations that impair mitochondrial steps in intermediary metabolism or ATP synthesis activated the mitochondrial chaperone genes. These observations support the existence of a mitochondrial unfolded protein response that couples mitochondrial chaperone gene expression to changes in the protein handling environment in the organelle. PMID- 15280429 TI - Alpha 3 beta 1 integrin promotes keratinocyte cell survival through activation of a MEK/ERK signaling pathway. AB - Inadequate or inappropriate adhesion of epithelial cells to extracellular matrix leads to a form of apoptosis known as anoikis. During various tissue remodelling events, such as wound healing or carcinoma invasion, changes in the physical properties, and/or composition of the extracellular matrix, can lead to anoikis of epithelial cells that lack appropriate receptor-matrix interactions. Laminin-5 is the major ligand for keratinocyte adhesion in the epidermis, and it also promotes keratinocyte survival in vivo and in vitro. Integrins alpha 3 beta 1 and alpha 6 beta 4 are the major receptors for laminin-5; however, specific roles for these integrins in keratinocyte survival have not been determined. In the current study, we exploited keratinocyte cell lines derived from wild-type or alpha 3 integrin knockout mice to reveal a critical role for alpha 3 beta 1 in protecting keratinocytes from apoptosis upon serum withdrawal. We show that alpha 3 beta 1 mediated adhesion to laminin-5 extracellular matrix inhibits proteolytic activation of caspase-3 and TUNEL-staining, both hallmarks of apoptosis. We also show that alpha 3 beta1-mediated adhesion activates focal adhesion kinase (FAK) and extracellular signal-regulated kinase (ERK), and that inhibition of either FAK or ERK signaling leads to apoptosis of keratinocytes attached to laminin-5. alpha 6 beta 4-mediated adhesion to laminin-5 only partially protects cells from apoptosis in the absence of alpha 3 beta 1, and alpha 6 beta 4 is not necessary for cell survival in the presence of alpha 3 beta 1. These results suggest that alpha 3 beta 1 is necessary and sufficient for maximal keratinocyte survival on laminin-5. We propose a model to address the potential importance of alpha 3 beta 1-mediated survival for migrating keratinocytes at the leading edge of a cutaneous wound. PMID- 15280430 TI - Small hepatocytes in culture develop polarized transporter expression and differentiation. AB - Rat small hepatocytes have been shown to proliferate in culture and to form organoids with differentiated hepatocytes in vitro. To evaluate the degree of polarized transporter differentiation of rat small hepatocytes during 9 weeks of culturing, we studied the time-dependent expression and subcellular localization of the major bile salt and organic anion transport systems of hepatocytes [i.e. the basolateral sodium-taurocholate co-transporting protein (Ntcp), organic-anion transporting polypeptide 1b2 (Oatp1b2), the canalicular bile-salt export pump (Bsep) and multidrug-resistance-associated protein 2 (Mrp2)]. Small hepatocytes proliferated and differentiated in culture and formed sharply demarcated colonies as assessed by morphology, alpha-fetoprotein, albumin and Mrp1 expression. Polarized surface transporter expression was evident after 5 weeks of culturing for Ntcp, Oatp1b2 and Mrp2, and after 7 weeks for Bsep. After 9 weeks in culture, the vast majority of matured hepatocytes expressed Ntcp/Oatp1b2 at the basolateral and Bsep/Mrp2 at the canalicular plasma-membrane domains. This polarized transporter expression was accompanied by canalicular secretion of fluorescein-diacetate and cholylglycyl-fluorescein. Furthermore, an anastomizing three-dimensional network of bile canaliculi developed within piling-up colonies. These data demonstrate that cultured rat small hepatocytes acquire a fully differentiated transporter expression phenotype during their development into hepatic 'organoid-like' clusters of mature hepatocytes. Thereby, the time dependent sequence of transporter expression mirrored the ontogenesis of transporter expression in developing rat liver, supporting the concept that small hepatocytes correspond to the hepatocyte lineage derived from embryonic hepatoblasts and/or from a different pool of 'committed hepatocyte progenitor cells'. PMID- 15280431 TI - Ryanodine receptors are expressed and functionally active in mouse spermatogenic cells and their inhibition interferes with spermatogonial differentiation. AB - Ryanodine receptors (RyRs) are intracellular calcium release channels that are highly expressed in striated muscle and neurons but are also detected in several non-excitable cells. We have studied the expression of the three RyR isoforms in male germ cells at different stages of maturation by western blot and RT-PCR. RyR1 was expressed in spermatogonia, pachytene spermatocytes and round spermatids whereas RyR2 was found only in 5- to 10-day-old testis but not in germ cells. RyR3 was not revealed at the protein level, although its mRNA was detected in mixed populations of germ cells. Caffeine, a known agonist of RyRs, was able to induce release of Ca(2+) from intracellular stores in spermatogonia, pachytene spermatocytes and round spermatids, but not spermatozoa. Treatment with high doses of ryanodine, which are known to block RyR channel activity, reduced spermatogonial proliferation and induced meiosis in in vitro organ cultures of testis from 7-day-old mice. In conclusion, the results presented here indicate that RyRs are present in germ cells and that calcium mobilization through RyR channels could participate to the regulation of male germ maturation. PMID- 15280432 TI - Analysis of Smad nucleocytoplasmic shuttling in living cells. AB - Transforming growth factor beta (TGF-beta) signalling leads to phosphorylation and activation of receptor-regulated Smad2 and Smad3, which form complexes with Smad4 and accumulate in the nucleus. The Smads, however, do not seem to reside statically in the cytoplasm in the absence of signalling or in the nucleus upon TGF-beta stimulation, but have been suggested to shuttle continuously between these cellular compartments in both the absence and presence of TGF-beta. Here we investigate this nucleocytoplasmic shuttling in detail in living cells using fusions of Smad2 and Smad4 with enhanced GFP. We first establish that the GFPSmad fusions behave like wild-type Smads in a variety of cellular assays. We go on to demonstrate directly, using photobleaching experiments, that Smad2 and Smad4 shuttle between the cytoplasm and nucleus in both TGF-beta-induced cells and in uninduced cells. In uninduced cells, GFPSmad2 is less mobile in the cytoplasm than is GFPSmad4, suggesting that it may be tethered there. In addition, we show that both GFPSmad2 and GFPSmad4 undergo a substantial decrease in mobility in the nucleus upon TGF-beta stimulation, suggesting that active complexes of Smads are tethered in the nucleus, whereas unactivated Smads are more freely diffusible. We propose that regulated cytoplasmic and nuclear retention may play a role in determining the distribution of Smads between the cytoplasm and the nucleus in both uninduced cells and upon TGF-beta induction. PMID- 15280433 TI - Coordination of cell growth and cell division: a mathematical modeling study. AB - Although there is general agreement that cell growth and division are functionally coordinated, the mechanisms that link the two processes are poorly understood. In this study, we have developed a mathematical model based on current biological concepts of the signaling transduction pathways involved in cell growth, which predicts that cell growth rate is proportional to cell surface area at birth. To investigate the relationship between growth control and cell division, we then applied our mathematical model to three classic experiments measuring cycle time versus cell birth size in fission yeast and Xenopus laevis, and the cell cycle delay in mammalian cells after serum withdrawal. When coupled to a cell cycle exhibiting 'sizer' and 'timer' phases, we show that a simple model in which growth rate is proportional to the cell surface area immediately after division reproduces the experimental observations including the relationship between cycle time and birth size in fission yeast and Xenopus laevis. The model also accounts for the cell cycle delay seen in restriction point experiments performed in HeLa cells. PMID- 15280434 TI - Stress response in yeast mRNA export factor: reversible changes in Rat8p localization are caused by ethanol stress but not heat shock. AB - Ethanol stress (10% v/v) causes selective mRNA export in Saccharomyces cerevisiae in a similar manner to heat shock (42 degrees C). Bulk poly(A)(+) mRNA accumulates in the nucleus, whereas heat shock protein mRNA is exported under such conditions. Here we investigated the effects of stress on mRNA export factors. In cells treated with ethanol stress, the DEAD box protein Rat8p showed a rapid and reversible change in its localization, accumulating in the nucleus. This change correlated closely with the blocking of bulk poly(A)(+) mRNA export caused by ethanol stress. We also found that the nuclear accumulation of Rat8p is caused by a defect in the Xpo1p/Crm1p exportin. Intriguingly, the localization of Rat8p did not change in heat shocked cells, suggesting that the mechanisms blocking bulk poly(A)(+) mRNA export differ for heat shock and ethanol stress. These results suggest that changes in the localization of Rat8p contribute to the selective export of mRNA in ethanol stressed cells, and also indicate differences in mRNA export between the heat shock response and ethanol stress response. PMID- 15280435 TI - Cytoprotective efficacy and mechanisms of the liposoluble iron chelator 2,2' dipyridyl in the rat photothrombotic ischemic stroke model. AB - We examined the efficacy of the liposoluble iron chelator 2,2'-dipyridyl (DP) in reducing histological damage in rats submitted to cerebral ischemia and the mechanisms involved in the potential cytoprotection. For this purpose, DP (20 mg/kg, i.p.) was administered 15 min before and 1 h after induction of cortical photothrombotic vascular occlusion in rat. Histological studies were performed to assess infarct volume (at days 1 and 3 postischemia) and astromicroglial activation (at day 3 postischemia). Damage to endothelial and neuronal cells was evaluated at day 1 postischemia by quantitative measurements of Evans Blue extravasation and N-acetylaspartate levels, respectively. Cerebral blood flow was recorded in the ischemic core by laser-Doppler flowmetry within the 15 min to 2 h period after photothrombosis. At 4-h postischemia, radical oxygen species (ROS) production was evaluated by measuring brain glutathione concentrations. The cortical expression of the proteins heme oxygenase-1 (HO-1) and hypoxia-inducible factor-1alpha (HIF-1alpha) was analyzed by Western blotting at day 1 postischemia. Infarct volume and ischemic damage to endothelial and neuronal cells were significantly reduced by DP treatment. This cytoprotection was associated with a reduction in ROS production, perfusion deficits, and astrocytic activation. DP treatment also resulted in significant changes in HO-1 (+100%) and HIF-1alpha (-50%) protein expression at the level of the ischemic core. These results report the efficacy of the liposoluble iron chelator DP in reducing histological damage induced by permanent focal ischemia. PMID- 15280436 TI - Differential sensitivity of N- and P/Q-type Ca2+ channel currents to a mu opioid in isolectin B4-positive and -negative dorsal root ganglion neurons. AB - Opioids have a selective effect on nociception with little effect on other sensory modalities. However, the cellular mechanisms for this preferential effect are not fully known. Two broad classes of nociceptors can be distinguished based on their growth factor requirements and binding to isolectin B4(IB4). In this study, we determined the difference in the modulation of voltage-gated Ca2+ currents by [D-Ala2,N-Me-Phe4,Gly-ol5]-enkephalin (DAMGO, a specific mu opioid agonist) between IB4-positive and -negative small dorsal root ganglion (DRG) neurons. Whole-cell voltage-clamp recordings were performed in acutely isolated DRG neurons in adult rats. Both 1-10 microM DAMGO and 1 to 10 microM morphine had a greater effect on high voltage-activated Ca2+ currents in IB4-negative than IB4 positive cells. However, DAMGO had no significant effect on T-type Ca2+ currents in both groups. The N-type Ca2+ current was the major subtype of Ca2+ currents inhibited by DAMGO in both IB4-positive and -negative neurons. Although DAMGO had no effect on L-type and R-type Ca2+ currents in both groups, it produced a larger inhibition on N-type and P/Q-type Ca2+ currents in IB4-negative than IB4-positive neurons. Furthermore, double labeling revealed that there was a significantly higher mu opioid receptor immunoreactivity in IB4-negative than IB4-positive cells. Thus, these data suggest that N-and P/Q-type Ca2+ currents are more sensitive to inhibition by the mu opioids in IB4-negative than IB4-positive DRG neurons. The differential sensitivity of voltage-gated Ca2+ channels to the mu opioids in subsets of DRG neurons may constitute an important analgesic mechanism of mu opioids. PMID- 15280437 TI - Haplotype-oriented genetic analysis and functional assessment of promoter variants in the MDR1 (ABCB1) gene. AB - Recently, a number of nucleotide variants have been described in the multidrug resistance 1 (MDR1/ABCB1) gene; however, most studies have focused on the coding region. In the present study, we identified promoter variants of the MDR1 gene and evaluated their phenotypic consequences using a reporter gene assay and the real-time polymerase chain reaction method. Ten allelic variants were detected in the promoter region (approximately 2 kilobases), seven of which were newly identified. Certain mutations occurred simultaneously, and a total of 10 haplotypes were observed. These promoter polymorphisms were found more frequently in Japanese than Caucasians. Some haplotypes were associated with changes in luciferase activity and placental and hepatic mRNA levels. We also determined DNA methylation status in the proximal promoter region of the MDR1 gene. The promoter region around potential binding sites for transcription factors was found to be hypomethylated and thus likely to be independent of the gene expression. Nucleotide and/or haplotype variants not only in the coding region but also in the promoter region of the MDR1 gene may be important for interindividual differences of P-glycoprotein expression. PMID- 15280438 TI - Rotenone induces apoptosis via activation of bad in human dopaminergic SH-SY5Y cells. AB - Chronic complex I inhibition caused by rotenone induces features of Parkinson's disease in rats, including selective nigrostriatal dopaminergic degeneration and Lewy bodies with alpha-synuclein-positive inclusions. To determine the mechanisms underlying rotenone-induced neuronal death, we used an in vitro model of human dopaminergic SH-SY5Y cells. In rotenone-induced cell death, rotenone induced Bad dephosphorylation without changing the amount of Bad proteins. Rotenone also increased the amount of alpha-synuclein in cells showing morphological changes in response to rotenone. Because Bad and alpha-synuclein are known to bind to 14-3-3 proteins, we examined the effects of rotenone on these complexes. Whereas a decreased Bad amount bound to 14-3-3 proteins, rotenone increased alpha-synuclein binding to these proteins. Because dephosphorylation by calcineurin activates Bad, we examined the possible involvement of Bad activation in rotenone-induced apoptosis by using the calcineurin inhibitor tacrolimus (FK506). Tacrolimus suppressed two rotenone-induced actions: Bad dephosphorylation and apoptosis. Furthermore, the inhibition of caspase-9, which functions downstream from Bad, completely suppressed rotenone-induced apoptosis. Our findings demonstrate that Bad activation plays a role in rotenone-induced apoptosis of SH-SY5Y cells. PMID- 15280439 TI - Effects of gamma- and hydroxypropyl-gamma-cyclodextrins on the transport of doxorubicin across an in vitro model of blood-brain barrier. AB - Association between doxorubicin (DOX) and gamma-cyclodextrin (gamma-CD) or hydroxypropyl-gamma-CD (HP-gamma-CD) has been examined to increase the delivery of this antitumoral agent to the brain. The stoichiometry and the stability constant of gamma-CD or HP-gamma-CD and DOX complexes were determined in physiological medium by UV-visible spectroscopy. By using an in vitro model of the blood-brain barrier (BBB), endothelial permeability and toxicity toward the brain capillary endothelial cells of DOX, gamma-CD, and HP-gamma-CD were performed. For each CD, endothelial permeability was relatively low and a disruption of the BBB occurred at 20 microM, 20 mM, and 50 mM DOX, gamma-CD, and HP-gamma-CD, respectively. Increasing amounts of CDs were added to a fixed DOX concentration. Addition of gamma-CD or HP-gamma-CD, up to 15 and 35 mM, respectively, decreased the DOX delivery, probably due to the low complex penetration across the BBB and the decrease in free DOX concentration. Higher CD concentrations increased the DOX delivery to the brain, but this effect is due to a loss of BBB integrity. In contrast to what was observed on Caco-2 cell model with various drugs, CDs are not able to increase the delivery of DOX across our in vitro model of BBB. PMID- 15280440 TI - Long-term ethanol self-administration by cynomolgus macaques alters the pharmacology and expression of GABAA receptors in basolateral amygdala. AB - We have recently demonstrated that chronic ethanol ingestion alters the functional and pharmacological properties of GABAA receptors measured in acutely isolated rat lateral/basolateral amygdala neurons, a limbic forebrain region involved with fear-learning and innate anxiety. To understand relevance of these results in the context of primates, we have examined the effects of long-term ethanol self-administration on basolateral amygdala GABAA receptor pharmacology and expression in cynomolgus macaques (Macaca fascicularis). The impact of this 18-month-long exposure on GABAA receptor function was assessed in acutely isolated neurons from basolateral amygdala with whole-cell patch-clamp electrophysiology. Neurons from control animals expressed maximal current densities that were not significantly different from the maximal current densities of neurons from ethanol-treated animals. However, the GABA concentration-response relationships from ethanol-exposed neurons were significantly right-shifted compared with control neurons. These adaptations were associated with significant alterations in some characteristics of macroscopic current desensitization. To understand the mechanism governing these adaptations, we quantified GABAA alpha subunit mRNAs in basolateral amygdala from the same animals. mRNA levels of the alpha2 and alpha3 subunits were significantly decreased, whereas decreases in alpha1 expression only approached statistical significance. There were no changes in alpha4 mRNA levels. These findings indicate that ethanol-induced alterations in GABAA function may be regulated in part by selective changes in the expression of particular alpha subunits. We conclude that adaptations of basolateral amygdala GABAA receptors after long-term ethanol self-administration by the cynomolgus macaque are similar, but not identical, to those described in rodents after a brief forced ethanol exposure. PMID- 15280441 TI - Comparative study of glucocorticoids, cyclosporine A, and JTE-607 [(-)-Ethyl N[3,5-dichloro-2-hydroxy-4-[2-(4-methylpiperazin-1-yl)ethoxy]benzoyl]-L phenylalaninate dihydrochloride] in a mouse septic shock model. AB - Actions of glucocorticoids, cyclosporine A, and JTE-607 [(-)-ethyl-N-[3,5 dichloro-2-hydroxy-4-[2-(4-methylpiperazin-1-yl)ethoxy]benzoyl]-L-phenylalaninate dihydrochloride], a proinflammatory cytokine inhibitor that does not inhibit interleukin (IL)-2 or interferon-gamma, were compared in a mouse septic shock model induced by cecal ligation and puncture (CLP). CLP caused elevation of macrophage inflammatory protein (MIP)-2 in lung, and MIP-2 and IL-6 in plasma and peritoneal fluid, reaching a peak 4 to 8 h after CLP. Myeloperoxidase (MPO) activity in lung increased and reached a peak 8 to 12 h after CLP. Acute treatment (subcutaneous injections 1 h before and 2 h after CLP) of mice with JTE 607 and methylprednisolone showed significant inhibition of elevated cytokine levels and MPO activity, plus increased survival rate. Similar treatment with cyclosporine A and prednisolone was ineffective. Chronic treatment (subcutaneous injection for seven consecutive days before CLP) of mice with JTE-607 also showed an inhibitory effect on cytokine production, MPO activity and mortality. In contrast, chronic treatment with cyclosporine A and prednisolone did not inhibit cytokine production or MPO activity, but rather exacerbated mortality. These results indicate that JTE-607 has protective effect on mouse mortality induced by CLP, correlating with inhibition of proinflammatory cytokines, whereas the immunosuppressants cyclosporine A and prednisolone do not. This suggests that JTE 607, a multiple cytokine inhibitor that does not cause adverse immunosuppression, is useful for treatment of septic shock. PMID- 15280442 TI - A common antitussive drug, clobutinol, precipitates the long QT syndrome 2. AB - QT prolongation, a classic risk factor for arrhythmias, can result from a mutation in one of the genes governing cardiac repolarization and also can result from the intake of a medication acting as blocker of the cardiac K(+) channel human ether-a-go-go-related gene (HERG). Here, we identified the arrhythmogenic potential of a nonopioid antitussive drug, clobutinol. The deleterious effects of clobutinol were suspected when a young boy, with a diagnosis of congenital long QT syndrome, experienced arrhythmias while being treated with this drug. Using the patch-clamp technique, we showed that clobutinol dose-dependently inhibited the HERG K(+) current with a half-maximum block concentration of 2.9 microM. In the proband, we identified a novel A561P HERG mutation. Two others long QT mutations (A561V and A561T) had been reported previously at the same position. None of the three mutants led to a sizeable current in heterologous expression system. When coexpressed with wild-type (WT) HERG channels, the three Ala561 mutants reduced the trafficking of WT and mutant heteromeric channels, resulting in decreased K(+) current amplitude (dominant-negative effects). In addition, A561P but not A561V and A561T mutants induced a approximately -11 mV shift of the current activation curve and accelerated deactivation, thereby partially counteracting the dominant-negative effects. A561P mutation and clobutinol effects on the human ventricular action potential characteristics were simulated using the Priebe-Beuckelmann model. Our work shows that clobutinol has limited effects on WT action potential but should be classified as a "drug to be avoided by congenital long QT patients" rather than as a "drug with risk of torsades de pointes". PMID- 15280443 TI - Cloning, up-regulation, and mitogenic role of porcine P2Y2 receptor in coronary artery smooth muscle cells. AB - Previous work has shown up-regulation of a UTP-sensitive P2Y receptor in porcine coronary smooth muscle cells (CSMC) of organ-cultured arteries. However, the molecular identity and functional role of this putative receptor remained undefined. Here we report the cloning of the cDNA for this receptor that encodes an open reading frame for a protein of 373 amino acids with the highest homology to the human P2Y(2) receptor (84%). Heterologous expression of this receptor in 1321N1 cells revealed a novel pharmacology in that UTP and ITP were full agonists and UTP was more potent and efficacious than ATP for increasing intracellular [Ca(2+)] and extracellular signal-regulated kinase phosphorylation. Stimulation of subcultured CSMC with UTP, ITP, or ATP induced a concentration-dependent increase in cellular DNA content, protein synthesis, cell number, and proliferating cell nuclear antigen expression, indicating a mitogenic role for P2Y(2) receptors. This was supported by the finding that the treatment of CSMC with antisense oligonucleotides to the cloned cDNA sequence significantly inhibited UTP- and ATP-induced DNA and protein synthesis. In addition, reverse transcription-polymerase chain reaction analysis showed that P2Y(2) receptor mRNA was dramatically increased in cells of organ-cultured arteries compared with freshly harvested arteries, whereas the P2Y(6) receptor mRNA level was unchanged, and the P2Y(4) receptor mRNA was undetectable. This P2Y(2) subtype-specific up regulation was confirmed in cells of coronary arteries stented in vivo. In conclusion, we have cloned the porcine P2Y(2) receptor with novel pharmacology and demonstrated that this receptor is up-regulated in CSMC of in vitro organ cultures or in vivo stented coronary arteries to mediate the mitogenic effects of nucleotides. PMID- 15280444 TI - 5beta-reduced neuroactive steroids are novel voltage-dependent blockers of T-type Ca2+ channels in rat sensory neurons in vitro and potent peripheral analgesics in vivo. AB - T-type Ca(2+) channels are believed to play an important role in pain perception, and anesthetic steroids such as alphaxalone and allopregnanolone, which have a 5alpha-configuration at the steroid A, B ring fusion, are known to inhibit T-type Ca(2+) channels and cause analgesia in a thermal nociceptive model (Soc Neurosci Abstr 29:657.9, 2003). To define further the structure-activity relationships for steroid analgesia, we synthesized and examined a series of 5beta-reduced steroids for their ability to induce thermal antinociception in rats when injected locally into the peripheral receptive fields of the nociceptors and studied their effects on T-type Ca(2+) channel function in vitro. We found that most of the steroids completely blocked T-type Ca(2+) currents in vitro with IC(50) values at a holding potential of -90 mV ranging from 2.8 to 40 microM. T current blockade exhibited mild voltage-dependence, suggesting that 5beta-reduced neuroactive steroids stabilize inactive states of the channel. For the most potent steroids, we found that other voltage-gated currents were not significantly affected at concentrations that produce nearly maximal blockade of T currents. All tested compounds induced dose-dependent analgesia in thermal nociceptive testing; the most potent effect (ED(50), 30 ng/100 microl) obtained with a compound [(3beta,5beta,17beta)-3-hydroxyandrostane-17-carbonitrile] that was also the most effective blocker of T currents. Compared with previously studied 5alpha-reduced steroids, these 5beta-reduced steroids are more efficacious blockers of neuronal T-type Ca(2+) channels and are potentially useful as new experimental reagents for understanding the role of neuronal T-type Ca(2+) channels in peripheral pain pathways. PMID- 15280445 TI - p63 and p73: roles in development and tumor formation. AB - The tumor suppressor p53 is critically important in the cellular damage response and is the founding member of a family of proteins. All three genes regulate cell cycle and apoptosis after DNA damage. However, despite a remarkable structural and partly functional similarity among p53, p63, and p73, mouse knockout studies revealed an unexpected functional diversity among them. p63 and p73 knockouts exhibit severe developmental abnormalities but no increased cancer susceptibility, whereas this picture is reversed for p53 knockouts. Neither p63 nor p73 is the target of inactivating mutations in human cancers. Genomic organization is more complex in p63 and p73, largely the result of an alternative internal promoter generating NH2-terminally deleted dominant-negative proteins that engage in inhibitory circuits within the family. Deregulated dominant negative p73 isoforms might play an active oncogenic role in some human cancers. Moreover, COOH-terminal extensions specific for p63 and p73 enable further unique protein-protein interactions with regulatory pathways involved in development, differentiation, proliferation, and damage response. Thus, p53 family proteins take on functions within a wide biological spectrum stretching from development (p63 and p73), DNA damage response via apoptosis and cell cycle arrest (p53, TAp63, and TAp73), chemosensitivity of tumors (p53 and TAp73), and immortalization and oncogenesis (DeltaNp73). PMID- 15280446 TI - Nucleoside diphosphate kinase A/nm23-H1 promotes metastasis of NB69-derived human neuroblastoma. AB - Nucleoside diphosphate kinase A (NDPK-A), encoded by the nm23-H1 gene, acts as a metastasis suppressor in certain human tumors such as breast carcinoma. However, evidence also points to NDPK-A functioning as a metastasis promoter in other human tumors including neuroblastoma. In fact, amplification and overexpression of nm23-H1 as well as S120G mutation of NDPK-A (NDPK-A(S120G)) have been detected in 14% to 30% of patients with advanced stages of neuroblastoma. To test whether NDPK-A promotes neuroblastoma metastasis, we established stable transfectants and an orthotopic xenograft animal model from the human neuroblastoma NB69 cell line. We demonstrate that overexpressed NDPK-A or NDPK-A(S120G) increased both incidence and colonization of neuroblastoma metastasis in animal lungs without significantly affecting primary tumor development. In vitro, these metastasis associated NDPK-A aberrations abrogated retinoic acid-induced neuronal differentiation while increasing cloning efficiency, cell survival, and colony formation of NB69 derivatives. Furthermore, NDPK-A(S120G) reduced cell adhesion and increased cell migration. Compared with its wild-type, NDPK-A(S120G) appears more effective in promoting neuroblastoma metastasis. Our results provide the first evidence that NDPK-A behaves as a metastasis promoter at least in human neuroblastoma derived from NB69 cells. The findings not only suggest a prognostic value of NDPK-A in neuroblastoma patients but also caution NDPK-A-targeted treatment for patients with different tumor types. PMID- 15280447 TI - Protease-activated receptors (PAR1 and PAR2) contribute to tumor cell motility and metastasis. AB - The effects of the pleiotropic serine protease thrombin on tumor cells are commonly thought to be mediated by the thrombin receptor protease-activated receptor 1 (PAR1). We demonstrate here that PAR1 activation has a role in experimental metastasis using the anti-PAR1 antibodies ATAP2 and WEDE15, which block PAR1 cleavage and activation. Thrombin also stimulates chemokinesis of human melanoma cells toward fibroblast conditioned media and soluble matrix proteins. Thrombin-enhanced migration is abolished by anti-PAR1 antibodies, demonstrating that PAR1 cleavage and activation are required. The PAR1-specific agonist peptide TFLLRNPNDK, however, does not stimulate migration, indicating that PAR1 activation is not sufficient. In contrast, a combination of TFLLRNPNDK and the PAR2 agonist peptide SLIGRL mimics the thrombin effect on migration, whereas PAR2 agonist alone has no effect. Agonist peptides for the thrombin receptors PAR3 and PAR4 used alone or with PAR1 agonist also have no effect. Similarly, activation of PAR1 and PAR2 also enhances chemokinesis of prostate cancer cells. Desensitization with PAR2 agonist abolishes thrombin-enhanced cell motility, demonstrating that thrombin acts through PAR2. PAR2 is cleaved by proteases with trypsin-like specificity but not by thrombin. Thrombin enhances migration in the presence of a cleavage-blocking anti-PAR2 antibody, suggesting that thrombin activates PAR2 indirectly and independent of receptor cleavage. Treatment of melanoma cells with trypsin or PAR2 agonist peptide enhances experimental metastasis. Together, these data confirm a role for PAR1 in migration and metastasis and demonstrate an unexpected role for PAR2 in thrombin dependent tumor cell migration and in metastasis. PMID- 15280448 TI - Interleukin-10 induced activating transcription factor 3 transcriptional suppression of matrix metalloproteinase-2 gene expression in human prostate CPTX 1532 Cells. AB - Aberrant expression of the 72-kDa type IV collagenase [matrix metalloproteinase (MMP)-2] is implicated in the invasion and angiogenesis process of malignant tumors. We investigated the effects of interleukin (IL)-10 on MMP-2 expression in CPTX-1532 human prostate tumor cells. Our results demonstrate that IL-10 significantly inhibited MMP-2 transcription and protein expression induced by a phorbol ester, phorbol 12-myristate 13-acetate. The inhibitory effects of IL-10 on MMP-2 expression correlated with the suppression of MMP-2 promoter activity. To determine the mechanism of IL-10 action, we examined IL-10-dependent promoter activity with luciferase constructs from a 2-kbp promoter region of the human MMP 2 gene. We functionally characterized the promoter fragments by transient transfection experiments with CPTX-1532 cells. The experiments revealed that a cAMP responsive element binding protein (CREB) consensus domain was identified upstream of the 5' transcriptional start site, which was highly responsive to IL 10-dependent down-regulation of promoter luciferase activity. Electrophoretic mobility shift assays combined with antibody "supershift assays" confirmed the data from the luciferase assays. Immunoblot assays of activating transcription factor (ATF) 3 immunoprecipitates with tyrosine specific antibodies revealed that IL-10 stimulated tyrosine phosphorylation of ATF3 to activate binding to the CREB domain and suppress MMP-2 expression. Studies with stable, IL-10 transfected CPTX 1532 subclones further showed that IL-10 failed to suppress MMP-2 expression in ATF3-deficient CPTX-1532 cells, where the ATF3 mRNA was destroyed with a DNAzyme oligonucleotide targeting the 5' region of the mRNA. Finally, reconstitution of ATF3 successfully restored the inhibitory effects of IL-10 on MMP-2 gene expression. Taken together, these data demonstrate the critical role of tyrosine phosphorylated ATF3 and the CREB consensus domain in IL-10 suppression of MMP-2 gene expression in primary human prostate tumor cells. PMID- 15280449 TI - Polo-like kinase 1 inactivation following mitotic DNA damaging treatments is independent of ataxia telangiectasia mutated kinase. AB - Polo-like kinase 1 (Plk1) is an important regulator of several events during mitosis. Recent reports show that Plk1 is involved in both G2 and mitotic DNA damage checkpoints. Ataxia telangiectasia mutated kinase (ATM) is an important enzyme involved in G2 phase cell cycle arrest following interphase DNA damage, and inhibition of Plk1 by DNA damage during G2 occurs in an ATM-/ATM-Rad3-related kinase (ATR)-dependent fashion. However, it is unclear how Plk1 is regulated in response to M phase DNA damage. We found that treatment of mitotic cells with DNA damaging agents inhibits Plk1 activity primarily through dephosphorylation of Plk1, which occurred in both p53 wild-type and mutant cells. Inhibition of Plk1 is not prevented by caffeine pretreatment that inhibits ATM activity and also occurs in ATM mutant cell lines. Furthermore, ATM mutant cell lines, unlike wild type cells, fail to arrest after mitotic DNA damaging treatments. The phosphatidylinositol 3-kinase (PI3K) inhibitor, LY294002, reduces Plk1 dephosphorylation following mitotic DNA damaging treatments, suggesting that the PI3K pathway may be involved in regulating Plk1 activity. Earlier studies showed that inhibition of Plk1 by G2 DNA damage occurs in an ATM-dependent fashion. Our results extend the previous studies by showing that ATM is not required for dephosphorylation and inhibition of Plk1 activity following mitotic DNA damage, and also suggest that Plk1 is not a principal regulator or mediator of the mitotic DNA damage response. PMID- 15280450 TI - Herpes simplex virus 1 has multiple mechanisms for blocking virus-induced interferon production. AB - In response to viral infection, host cells elicit a number of responses, including the expression of alpha/beta interferon (IFN-alpha/beta). In these cells, IFN regulatory factor-3 (IRF-3) undergoes a sequence of posttranslational modifications that allow it to act as a potent transcriptional coactivator of specific IFN genes, including IFN-beta. We investigated the mechanisms by which herpes simplex virus 1 (HSV-1) inhibits the production of IFN-beta mediated by the IRF-3 signaling pathway. Here, we show that HSV-1 infection can block the accumulation of IFN-beta triggered by Sendai virus (SeV) infection. Our results indicate that HSV-1 infection blocks the nuclear accumulation of activated IRF-3 but does not block the initial virus-induced phosphorylation of IRF-3. The former effect was at least partly mediated by increased turnover of IRF-3 in HSV-1 infected cells. Using mutant viruses, we determined that the immediate-early protein ICP0 was necessary for the inhibition of IRF-3 nuclear accumulation. Expression of ICP0 also had the ability to reduce IFN-beta production induced by SeV infection. ICP0 has been shown previously to play a role in HSV-1 sensitivity to IFN and in the inhibition of antiviral gene production. However, we observed that an ICP0 mutant virus still retained the ability to inhibit the production of IFN-beta. These results argue that HSV-1 has multiple mechanisms to inhibit the production of IFN-beta, providing additional ways in which HSV-1 can block the IFN-mediated host response. PMID- 15280451 TI - Integrated self-inactivating lentiviral vectors produce full-length genomic transcripts competent for encapsidation and integration. AB - To make human immunodeficiency virus type 1 (HIV-1)-based vectors safer for use in the research and clinical setting, a significant modification to the HIV-1 genome has been the deletion of promoter and enhancer elements from the U3 region of the long terminal repeat (LTR). Vectors containing this deletion are thought to have no LTR-directed transcription and are called self-inactivating (SIN) lentivectors. Using four distinct approaches, we show that SIN lentivectors continue to have promoter activity near the 5' LTR, which is responsible for the production of full-length vector transcripts. To verify that transcripts derived from the LTR in SIN lentivectors are competent for encapsidation and integration, we transduced a lentiviral packaging cell line with a SIN lentivector and then observed the production of viable vector particles containing full-length SIN lentivector genomes. We have also attempted to identify sequences in the SIN lentivector which are responsible for transcriptional activation at the 5' LTR. Using different segments of the vector LTR and leader region in a promoter assay, we have determined that the residual promoter activity is contained entirely within the leader region and that, although this element is downstream of the transcription initiation site, it is capable of initiating transcription from the 5' end of R in the LTR. Mutation of leader region binding sites for the transcriptional activators downstream binding factor 1 (DBF1) and SP1 reduces transcription from the SIN LTR by up to 80%. Knowledge of the potential for mobilization of HIV-1-derived SIN lentivectors will be important for the design of future gene therapy trials with such vectors. PMID- 15280452 TI - Frequent transmission of cytotoxic-T-lymphocyte escape mutants of human immunodeficiency virus type 1 in the highly HLA-A24-positive Japanese population. AB - Although Japan is classified as a country with a low prevalence of human immunodeficiency virus type 1 (HIV-1), domestic sexual transmission has been increasing steadily. Because 70% of the Japanese population expresses HLA-A24 (genotype HLA-A*2402), we wished to assess the effect of the dominant HLA type on the evolution and transmission of HIV-1 among the Japanese population. Twenty three out of 25 A24-positive Japanese patients had a Y-to-F substitution at the second position [Nef138-10(2F)] in an immunodominant A24-restricted CTL epitope in their HIV-1 nef gene (Nef138-10). None of 12 A24-negative Japanese hemophiliacs but 9 out of 16 patients infected through unprotected sexual intercourse had Nef138-10(2F) (P < 0.01). Two of two A24-positive but none of six A24-negative Australians had Nef138-10(2F). Nef138-10(2F) peptides bound well to the HLA-A*2402 heavy chain; however, Nef138-10(2F) was expressed poorly on the cell surface from the native protein. Thus, HIV-1 with Nef138-10(2F) appears to be a cytotoxic-T-lymphocyte escape mutant and has been transmitted frequently by sexual contact among the highly A24-positive Japanese population. PMID- 15280453 TI - Characterization of recombinant respiratory syncytial viruses with the region responsible for type 2 T-cell responses and pulmonary eosinophilia deleted from the attachment (G) protein. AB - It is essential that preventative vaccines for respiratory syncytial virus (RSV) elicit balanced T-cell responses. Immune responses dominated by type 2 T cells against RSV antigens are believed to cause exaggerated respiratory tract disease and may also contribute to unwanted inflammation in the airways that predisposes infants to wheeze through adolescence. Here we report on the construction and characterization of recombinant RSV (rRSV) strains with amino acids 151 to 221 or 178 to 219 of the attachment (G) glycoprotein deleted (rA2cpDeltaG150-222 or rA2cpDeltaG177-220, respectively). The central ectodomain was chosen for modification because a peptide spanning amino acids 149 to 200 of G protein has recently been shown to prime several strains of naive inbred mice for polarized type 2 T-cell responses, and peripheral blood T cells from most human donors recognize epitopes within this region. Quantitative PCR demonstrated that synthesis of nascent rRSV genomes in human lung epithelial cell lines was similar to that for the parent virus (cp-RSV). Plaque assays further indicated that rRSV replication was not sensitive to 37 degrees C, but pinpoint morphology was observed at 39 degrees C. Both rRSV strains replicated in the respiratory tracts of BALB/c mice and elicited serum neutralization and anti-F-protein immunoglobulin G titers that were equivalent to those elicited by cp-RSV and contributed to a 3.9-log(10)-unit reduction in RSV A2 levels 4 days after challenge. Importantly, pulmonary eosinophilia was significantly diminished in BALB/c mice primed with native G protein and challenged with either rA2cpDeltaG150-222 or rA2cpDeltaG177-220. These findings are important for the development of attenuated RSV vaccines. PMID- 15280454 TI - PKR-dependent and -independent mechanisms are involved in translational shutoff during Sindbis virus infection. AB - The replication of Sindbis virus (SIN) profoundly affects the metabolism of infected vertebrate cells. One of the main events during SIN infection is the strong inhibition of translation of cellular mRNAs. In this study, we used a combination of approaches, including the study of SIN replication in PKR(-/-) mouse embryo fibroblasts or in the presence of an excess of catalytically inactive PKR. We show that the PKR-dependent inhibition of translation is not the only and most likely not the major pathway mediating translational shutoff during SIN infection. The PKR-independent mechanism strongly affects the translation of cellular templates, whereas translation of SIN subgenomic RNA is resistant to inhibition, and this leads to a benefit for viral replication. Our findings suggest that both PKR-dependent and non-PKR-dependent mechanisms of SIN-induced translational shutoff can be manipulated by using SIN replicons expressing mutated SIN nsP2 or kinase-defective PKR. Specifically, we show that expression of heterologous genes from SIN-based and most likely other alphavirus-based replicons can be increased by downregulating both the PKR-dependent and PKR independent translational shutoffs. PMID- 15280455 TI - Development of a DNA vaccine targeting human papillomavirus type 16 oncoprotein E6. AB - Human papillomavirus (HPV), particularly type 16 (HPV-16), is present in more than 99% of cervical cancers. The HPV oncoproteins E6 and E7 are constantly expressed and therefore represent ideal targets for HPV vaccine development. We previously developed DNA vaccines encoding calreticulin (CRT) linked to HPV-16 E7 and generated potent E7-specific CD8(+) T-cell immune responses and antitumor effects against an E7-expressing tumor. Since vaccines targeting E6 also represent an important strategy for controlling HPV-associated lesions, we developed a DNA vaccine encoding CRT linked to E6 (CRT/E6). Our results indicated that the CRT/E6 DNA vaccine, but not a wild-type E6 DNA vaccine, generated significant E6-specific CD8(+) T-cell immune responses in vaccinated mice. Mapping of the immunodominant epitope of E6 revealed that an E6 peptide comprising amino acids (aa) 48 to 57 (E6 aa48-57), presented by H-2K(b), is the optimal peptide and that the region of E6 comprising aa 50 to 57 represents the minimal core sequence required for activating E6-specific CD8(+) T lymphocytes. We also demonstrated that E6 aa48-57 contains cytotoxic T-lymphocyte epitopes naturally presented by E6-expressing TC-1 cells. Vaccination with a CRT/E6 but not a CRT/mtE6 (lacking aa 50 to 57 of E6) DNA vaccine could protect vaccinated mice from challenge with E6-expressing TC-1 tumors. Thus, our data indicate that E6 aa48-57 contains the immunodominant epitope and that a CRT/E6 DNA vaccine may be useful for control of HPV infection and HPV-associated lesions. PMID- 15280456 TI - Initial cleavage of the human immunodeficiency virus type 1 GagPol precursor by its activated protease occurs by an intramolecular mechanism. AB - Processing of the GagPol polyprotein precursor of human immunodeficiency virus type 1 (HIV-1) is a critical step in viral assembly and replication. The HIV-1 protease (PR) is translated as part of GagPol and is both necessary and sufficient for precursor processing. The PR is active only as a dimer; enzyme activation is initiated when the PR domains in two GagPol precursors dimerize. The precise mechanism by which the PR becomes activated and the subsequent initial steps in precursor processing are not well understood. However, it is clear that processing is initiated by the PR domain that is embedded within the precursor itself. We have examined the earliest events in precursor processing using an in vitro assay in which full-length GagPol is cleaved by its embedded PR. We demonstrate that the embedded, immature PR is as much as 10,000-fold less sensitive to inhibition by an active-site PR inhibitor than is the mature, free enzyme. Further, we find that different concentrations of the active-site inhibitor are required to inhibit the processing of different cleavage sites within GagPol. Finally, our results indicate that the first cleavages carried out by the activated PR within GagPol are intramolecular. Overall, our data support a model of virus assembly in which the first cleavages occur in GagPol upstream of the PR. These intramolecular cleavages produce an extended form of PR that completes the final processing steps accompanying the final stages of particle assembly by an intermolecular mechanism. PMID- 15280457 TI - Basic residues of the retroviral nucleocapsid play different roles in gag-gag and Gag-Psi RNA interactions. AB - The Orthoretrovirus Gag interaction (I) domain maps to the nucleocapsid (NC) domain in the Gag polyprotein. We used the yeast two-hybrid system to analyze the role of Alpharetrovirus NC in Gag-Gag interactions and also examined the efficiency of viral assembly and release in vivo. We could delete either or both of the two Cys-His (CH) boxes without abrogating Gag-Gag interactions. We found that as few as eight clustered basic residues, attached to the C terminus of the spacer peptide separating the capsid (CA) and NC domains in the absence of NC, was sufficient for Gag-Gag interactions. Our results support the idea that a sufficient number of basic residues, rather than the CH boxes, play the important role in Gag multimerization. We also examined the requirement for basic residues in Gag for packaging of specific packaging signal (Psi)-containing RNA. Using a yeast three-hybrid RNA-protein interaction assay, second-site suppressors of a packaging-defective Gag mutant were isolated, which restored Psi RNA binding. These suppressors mapped to the p10 or CA domains in Gag and resulted in either introduction of a positively charged residue or elimination of a negatively charged one. These results imply that the structural interactions of NC with other domains of Gag are necessary for Psi RNA binding. Taken together, our results show that while Gag assembly only requires a certain number of positively charged amino acids, Gag binding to genomic RNA for packaging requires more complex interactions inherent in the protein tertiary structure. PMID- 15280458 TI - Diverse hepatitis C virus glycoproteins mediate viral infection in a CD81 dependent manner. AB - We recently reported that retroviral pseudotypes bearing the hepatitis C virus (HCV) strain H and Con1 glycoproteins, genotype 1a and 1b, respectively, require CD81 as a coreceptor for virus-cell entry and infection. Soluble truncated E2 cloned from a number of diverse HCV genotypes fail to interact with CD81, suggesting that viruses of diverse origin may utilize different receptors and display altered cell tropism. We have used the pseudotyping system to study the tropism of viruses bearing diverse HCV glycoproteins. Viruses bearing these glycoproteins showed a 150-fold range in infectivity for hepatoma cells and failed to infect lymphoid cells. The level of glycoprotein incorporation into particles varied considerably between strains, generally reflecting the E2 expression level within transfected cells. However, differences in glycoprotein incorporation were not associated with virus infectivity, suggesting that infectivity is not limited by the absolute level of glycoprotein. All HCV pseudotypes failed to infect HepG2 cells and yet infected the same cells after transduction to express human CD81, confirming the critical role of CD81 in HCV infection. Interestingly, these HCV pseudotypes differed in their ability to infect HepG2 cells expressing a panel of CD81 variants, suggesting subtle differences in the interaction of CD81 residues with diverse viral glycoproteins. Our current model of HCV infection suggests that CD81, together with additional unknown liver specific receptor(s), mediate the virus-cell entry process. PMID- 15280459 TI - Analysis of integration sites of Jaagsiekte sheep retrovirus in ovine pulmonary adenocarcinoma. AB - Ovine pulmonary adenocarcinoma (OPA) is an infectious lung tumor of sheep caused by Jaagsiekte sheep retrovirus (JSRV). To test the hypothesis that JSRV insertional mutagenesis is involved in the oncogenesis of OPA, we cloned and characterized 70 independent integration sites from 23 cases of OPA. Multiple integration sites were identified in most tumors. BLAST analysis of the sequences did not disclose any potential oncogenic motifs or any identical integration sites in different tumors. Thirty-seven of the integration sites were mapped to individual chromosomes by PCR with a panel of sheep-hamster hybrid cell lines. Integration sites were found on 20 of the 28 sheep chromosomes, suggesting a random distribution. However, four integration sites from four different tumors mapped to chromosome 16. By Southern blot hybridization, probes derived from two of these sites mapped to within 5 kb of each other on normal sheep DNA. These sites were found within a single sheep bacterial artificial chromosome clone and were further mapped to only 2.5 kb apart, within an uncharacterized predicted gene and less than 200 kb from a mitogen-activated protein kinase-encoding gene. These findings suggest that there is at least one common integration site for JSRV in OPA and add weight to the hypothesis that insertional mutagenesis is involved in the development of this tumor. PMID- 15280460 TI - Mutations in multiple domains activate paramyxovirus F protein-induced fusion. AB - SER virus, a paramyxovirus that is closely related to simian virus 5 (SV5), is unusual in that it fails to induce syncytium formation. The SER virus F protein has an unusually long cytoplasmic tail (CT), and it was previously observed that truncations or specific mutations of this domain result in enhanced syncytium formation. In addition to the long CT, the SER F protein has nine amino acid differences from the F protein of SV5. We previously observed only a partial suppression of fusion in a chimeric SV5 F protein with a CT derived from SER virus, indicating that these other amino acid differences between the SER and SV5 F proteins also play a role in regulating the fusion phenotype. To examine the effects of individual amino acid differences, we mutated the nine SER residues individually to the respective residues of the SV5 F protein. We found that most of the mutants were expressed well and were transported to the cell surface at levels comparable to that of the wild-type SER F protein. Many of the mutants showed enhanced lipid mixing, calcein transfer, and syncytium formation even in the presence of the long SER F protein CT. Some mutants, such as the I310 M, T438S, M489I, T516V, and N529K mutants, also showed fusion at lower temperatures of 32, 25, and 18 degrees C. The residue Asn529 plays a critical role in the suppression of fusion activity, as the mutation of this residue to lysine caused a marked enhancement of fusion. The effect of the N529K mutation on the enhancement of fusion by a previously described mutant, L539,548A, as well as by chimeric SV5/SER F proteins was also dramatic. These results indicate that activation to a fusogenic conformation is dependent on the interplay of residues in the ectodomain, the transmembrane domain, and the CT domain of paramyxovirus F proteins. PMID- 15280461 TI - Basal core promoter and precore mutations in the hepatitis B virus genome enhance replication efficacy of Lamivudine-resistant mutants. AB - During chronic hepatitis B virus (HBV) infection, mutations in the precore (PC) or basal core promoter (BCP) region affecting HBV e antigen (HBeAg) expression occur commonly and represent the predominant virus species in patients with HBeAg negative chronic hepatitis B. The PC mutation (G1896A+C1858T) creates a translational stop codon resulting in absent HBeAg expression, whereas BCP mutations (A1762T/G1764A) reduce HBeAg expression by transcriptional mechanisms. Treatment of chronic HBV infection with lamivudine (LMV) often selects drug resistant strains with single (rtM204I) or double (rtL180M+rtM204V) point mutations in the YMDD motif of HBV reverse transcriptase. We cloned replication competent HBV vectors (genotype A, adw2) combining mutations in the core (wild type [wt], PC, and BCP) and polymerase gene (wt, rtM204I, and rtL180M/M204V) and analyzed virus replication and drug sensitivity in vitro. Resistance to LMV (rtM204I/rtL180M+rtM204V) was accompanied by a reduced replication efficacy as evidenced by reduced pregenomic RNA, encapsidated progeny DNA, polymerase activity, and virion release. PC mutations alone did not alter virus replication but restored replication efficacy of the LMV-resistant mutants without affecting drug resistance. BCP mutants had higher replication capacities than did the wt, also in combination with LMV resistance mutations. All nine HBV constructs showed similar sensitivities to adefovir. In conclusion, BCP-PC mutations directly impact the replication capacity of LMV-resistant mutants. PC mutations compensated for replication inefficiency of LMV-resistant mutants, whereas BCP mutations increased viral replication levels to above the wt baseline values, even in LMV-resistant mutants, without affecting drug sensitivity in vitro. Adefovir may be an effective treatment when combinations of core and polymerase mutations occur. PMID- 15280462 TI - Effect of membrane curvature-modifying lipids on membrane fusion by tick-borne encephalitis virus. AB - Enveloped viruses enter cells by fusion of their own membrane with a cellular membrane. Incorporation of inverted-cone-shaped lipids such as lysophosphatidylcholine (LPC) into the outer leaflet of target membranes has been shown previously to impair fusion mediated by class I viral fusion proteins, e.g., the influenza virus hemagglutinin. It has been suggested that these results provide evidence for the stalk-pore model of fusion, which involves a hemifusion intermediate (stalk) with highly bent outer membrane leaflets. Here, we investigated the effect of inverted-cone-shaped LPCs and the cone-shaped oleic acid (OA) on the membrane fusion activity of a virus with a class II fusion protein, the flavivirus tick-borne encephalitis virus (TBEV). This study included an analysis of lipid mixing, as well as of the steps preceding or accompanying fusion, i.e., binding to the target membrane and lipid-induced conformational changes in the fusion protein E. We show that the presence of LPC in the outer leaflet of target liposomes strongly inhibited TBEV-mediated fusion, whereas OA caused a very slight enhancement, consistent with a fusion mechanism involving a lipid stalk. However, LPC also impaired the low-pH-induced binding of a soluble form of the E protein to liposomes and its conversion into a trimeric postfusion structure that requires membrane binding at low pH. Because inhibition is already observed before the lipid-mixing step, it cannot be determined whether impairment of stalk formation is a contributing factor in the inhibition of fusion by LPC. These data emphasize, however, the importance of the composition of the target membrane in its interactions with the fusion peptide that are crucial for the initiation of fusion. PMID- 15280463 TI - Identification of protein tyrosine kinases required for B-cell- receptor-mediated activation of an Epstein-Barr Virus immediate-early gene promoter. AB - Epstein-Barr Virus (EBV) is a potentially oncogenic herpesvirus that infects >90% of the world's population. EBV exists predominantly as a latent infection in B lymphocytes, with periodic lytic-cycle reactivation essential for cellular and host transmission. Viral reactivation can be stimulated by ligand-induced activation of B-cell-receptor (BCR)-coupled signaling pathways. The critical first step in the transition from latency to the lytic cycle is the expression of the viral immediate-early gene BZLF1 through the transcription activation of its promoter, Zp. However, the BCR-coupled signal transduction cascade(s) leading to the induction of Zp and the expression of the BZLF1 gene product, Zta, is currently unclear. A major obstacle to delineating the relevant signal transduction events has been the lack of a model of EBV infection that is amenable to genetic manipulation. The use of the avian B-cell line DT40 has proven to be a powerful tool for delineating BCR-mediated signal transduction pathways that appear to be highly conserved between avian and mammalian systems. We demonstrate that the DT40 cell line is a robust and genetically tractable system for the study of BCR-mediated signaling pathways leading to transcriptional activation of BZLF1. Using this system, we demonstrate that activation of Zp requires the BCR-coupled protein tyrosine kinases Syk and Btk and that it is positively regulated by Lyn. Thus, the use of DT40 cells has allowed us to delineate the early signaling components required for BCR-dependent reactivation of latent EBV, and this system is likely to prove useful for further dissection of the downstream signaling cascades involved. PMID- 15280464 TI - Dissecting the requirement for subgenomic promoter sequences by RNA recombination of brome mosaic virus in vivo: evidence for functional separation of transcription and recombination. AB - Previously, we and others mapped an increased homologous recombination activity within the subgenomic promoter (sgp) region in brome mosaic virus (BMV) RNA3. In order to correlate sgp-mediated recombination and transcription, in the present work we used BMV RNA3 constructs that carried altered sgp repeats. We observed that the removal or extension of the poly(U) tract reduced or increased recombination, respectively. Deletion of the sgp core hairpin or its replacement by a different stem-loop structure inhibited recombination activity. Nucleotide substitutions at the +1 or +2 transcription initiation position reduced recombination. The sgp core alone supported only basal recombination activity. The sites of crossovers mapped to the poly(U) region and to the core hairpin. The observed effects on recombination did not parallel those observed for transcription. To explain how both activities operate within the sgp sequence, we propose a dual mechanism whereby recombination is primed at the poly(U) tract by the predetached nascent plus strand, whereas transcription is initiated de novo at the sgp core. PMID- 15280465 TI - Pulmonary collectins modulate strain-specific influenza a virus infection and host responses. AB - Collectins are secreted collagen-like lectins that bind, agglutinate, and neutralize influenza A virus (IAV) in vitro. Surfactant proteins A and D (SP-A and SP-D) are collectins expressed in the airway and alveolar epithelium and could have a role in the regulation of IAV infection in vivo. Previous studies have shown that binding of SP-D to IAV is dependent on the glycosylation of specific sites on the HA1 domain of hemagglutinin on the surface of IAV, while the binding of SP-A to the HA1 domain is dependent on the glycosylation of the carbohydrate recognition domain of SP-A. Here, using SP-A and SP-D gene-targeted mice on a common C57BL6 background, we report that viral replication and the host response as measured by weight loss, neutrophil influx into the lung, and local cytokine release are regulated by SP-D but not SP-A when the IAV is glycosylated at a specific site (N165) on the HA1 domain. SP-D does not protect against IAV infection with a strain lacking glycosylation at N165. With the exception of a small difference on day 2 after infection with X-79, we did not find any significant difference in viral load in SP-A(-/-) mice with either IAV strain, although small differences in the cytokine responses to IAV were detected in SP A(-/-) mice. Mice deficient in both SP-A and SP-D responded to IAV similarly to mice deficient in SP-D alone. Since most strains of IAV currently circulating are glycosylated at N165, SP-D may play a role in protection from IAV infection. PMID- 15280466 TI - Evidence that stable retroviral transduction and cell survival following DNA integration depend on components of the nonhomologous end joining repair pathway. AB - We have previously reported several lines of evidence that support a role for cellular DNA repair systems in completion of the retroviral DNA integration process. Failure to repair an intermediate in the process of integrating viral DNA into host DNA appears to trigger growth arrest or death of a large percentage of infected cells. Cellular proteins involved in the nonhomologous end joining (NHEJ) pathway (DNA-PK(CS)) and the damage-signaling kinases (ATM and ATR) have been implicated in this process. However, some studies have suggested that NHEJ proteins may not be required for the completion of lentiviral DNA integration. Here we provide additional evidence that NHEJ proteins are required for stable transduction by human immunodeficiency type 1 (HIV-1)-based vectors. Our analyses with two different reporters show that the number of stably transduced DNA-PK(CS) deficient scid fibroblasts was reduced by 80 to 90% compared to the number of control cells. Furthermore, transduction efficiency can be restored to wild-type levels in scid cells that are complemented with a functional DNA-PK(CS) gene. The efficiency of stable transduction by an HIV-1-based vector is also reduced upon infection of Xrcc4 and ligase IV-deficient cells, implying a role for these components of the NHEJ repair pathway. Finally, we show that cells deficient in ligase IV are killed by infection with an integrase-competent but not an integrase-deficient HIV-1 vector. Results presented in this study lend further support to a general role for the NHEJ DNA repair pathway in completion of the retroviral DNA integration process. PMID- 15280468 TI - Role of JC virus agnoprotein in DNA repair. AB - The late region of human neurotropic JC virus encodes a small 71-amino-acid agnoprotein that is also found in the polyomaviruses simian virus 40 and BK virus. Several functions of agnoprotein have been identified, including roles in regulating viral transcription and virion maturation. Earlier studies showed that agnoprotein expressed alone induced p21/WAF-1 expression and caused cells to accumulate in the G(2)/M stage of the cell cycle. Here we report that agnoprotein expression sensitized cells to the cytotoxic effects of the DNA-damaging agent cisplatin. Agnoprotein reduced the viability of cisplatin-treated cells and increased chromosome fragmentation and micronucleus formation. Whereas cisplatin treated control cells accumulated in S phase, cells expressing agnoprotein did not, instead becoming aneuploid. Agnoprotein expression correlated with impaired double-strand-break repair activity in cellular extracts and reduced expression of the Ku70 and Ku80 DNA repair proteins. After agnoprotein expression, much of the Ku70 protein was located in the perinuclear space, where agnoprotein was also found. Results from binding studies showed an interaction of agnoprotein with Ku70 which was mediated by the N terminus. The ability of agnoprotein to inhibit double-strand break repair activity when it was added to cellular extracts was also mediated by the N terminus. We conclude that agnoprotein inhibits DNA repair after DNA damage and interferes with DNA damage-induced cell cycle regulation. Since Ku70 is a subunit of the DNA-dependent protein kinase that is responsible both for double-strand break repair and for signaling damage-induced cell cycle arrest, the modulation of Ku70 and/or Ku80 by agnoprotein may represent an important event in the polyomavirus life cycle and in cell transformation. PMID- 15280467 TI - Herpes simplex virus 1 induces cytoplasmic accumulation of TIA-1/TIAR and both synthesis and cytoplasmic accumulation of tristetraprolin, two cellular proteins that bind and destabilize AU-rich RNAs. AB - Herpes simplex virus 1 causes a shutoff of cellular protein synthesis through the degradation of RNA that is mediated by the virion host shutoff (Vhs) protein encoded by the U(L)41 gene. We reported elsewhere that the Vhs-dependent degradation of RNA is selective, and we identified RNAs containing AU-rich elements (AREs) that were upregulated after infection but degraded by deadenylation and progressive 3'-to-5' degradation. We also identified upregulated RNAs that were not subject to Vhs-dependent degradation (A. Esclatine, B. Taddeo, L. Evans, and B. Roizman, Proc. Natl. Acad. Sci. USA 101:3603-3608, 2004). Among the latter was the RNA encoding tristetraprolin, a protein that binds AREs and is known to be associated with the degradation of RNAs containing AREs. Prompted by this observation, we examined the status of the ARE binding proteins tristetraprolin and TIA-1/TIAR in infected cells. We report that tristetraprolin was made and accumulated in the cytoplasm of wild-type virus infected human foreskin fibroblasts as early as 2 h and in HEp-2 cells as early as 6 h after infection. The amounts of tristetraprolin that accumulated in the cytoplasm of cells infected with a mutant virus lacking U(L)41 were significantly lower than those in wild-type virus-infected cells. The localization of tristetraprolin was not modified in cells infected with a mutant lacking the gene encoding infected cell protein 4 (ICP4). TIA-1 and TIAR are two other proteins that are associated with the regulation of ARE-containing RNAs and that normally reside in nuclei. In infected cells, they started to accumulate in the cytoplasm after 6 h of infection. In cells infected with the mutant virus lacking U(L)41, TIA-1/TIAR accumulated in the cytoplasm in granular structures reminiscent of stress granules in a significant percentage of the cells. In addition, an antibody to tristetraprolin coprecipitated the Vhs protein from lysates of cells late in infection. The results indicate that the Vhs-dependent degradation of ARE containing RNAs correlates with the transactivation, cytoplasmic accumulation, and persistence of tristetraprolin in infected cells. PMID- 15280469 TI - Caspases mediate processing of the capsid precursor and cell release of human astroviruses. AB - In this work we have shown that astrovirus infection induces apoptosis of Caco-2 cells, since fragmentation of cellular DNA, cleavage of cellular proteins which are substrate of activated caspases, and a change in the mitochondrial transmembrane potential occur upon virus infection. The human astrovirus Yuc8 polyprotein capsid precursor VP90 is initially processed to yield VP70, and we have shown that this processing is trypsin independent and occurs intracellularly through four cleavages at its carboxy-terminal region. We further showed that VP90-VP70 processing is mediated by caspases, since it was blocked by the pancaspase inhibitor benzyloxycarbonyl-Val-Ala-Asp fluoromethylketone (z-VAD fmk), and it was promoted by the apoptosis inducer TNF-related apoptosis-inducing ligand (TRAIL). Although the cell-associated virus produced in the presence of these compounds was not affected, the release of infectious virus to the cell supernatant was drastically reduced in the presence of z-VAD-fmk and increased by TRAIL, indicating that VP90-VP70 cleavage is important for the virus particles to be released from the cell. This is the first report that describes the induction and utilization of caspase activity by a virus to promote processing of the capsid precursor and dissemination of the viral particles. PMID- 15280470 TI - Continuing evolution of H9N2 influenza viruses in Southeastern China. AB - H9N2 influenza viruses are panzootic in domestic poultry in Eurasia and since 1999 have caused transient infections in humans and pigs. To investigate the zoonotic potential of H9N2 viruses, we studied the evolution of the viruses in live-poultry markets in Hong Kong in 2003. H9N2 was the most prevalent influenza virus subtype in the live-poultry markets between 2001 and 2003. Antigenic and phylogenetic analysis of hemagglutinin (HA) showed that all of the 19 isolates found except one belonged to the lineage represented by A/Duck/Hong Kong/Y280/97 (H9N2). The exception was A/Guinea fowl/NT184/03 (H9N2), whose HA is most closely related to that of the human isolate A/Guangzhou/333/99 (H9N2), a virus belonging to the A/Chicken/Beijing/1/94-like (H9N2) lineage. At least six different genotypes were recognized. The majority of the viruses had nonstructural (and HA) genes derived from the A/Duck/Hong Kong/Y280/97-like virus lineage but had other genes of mixed avian virus origin, including genes similar to those of H5N1 viruses isolated in 2001. Viruses of all six genotypes of H9N2 found were able to replicate in chickens and mice without adaptation. The infected chickens showed no signs of disease, but representatives of two viral genotypes were lethal to mice. Three genotypes of virus replicated in the respiratory tracts of swine, which shed virus for at least 5 days. These results show an increasing genetic and biologic diversity of H9N2 viruses in Hong Kong and support their potential role as pandemic influenza agents. PMID- 15280471 TI - Kaposi's sarcoma-associated herpesvirus ori-Lyt-dependent DNA replication: cis acting requirements for replication and ori-Lyt-associated RNA transcription. AB - Herpesvirus lytic DNA replication requires both the cis-acting element, the origin, and trans-acting factors such as virally encoded origin-binding protein and DNA replication enzymes. Recently, the origins of lytic DNA replication (ori Lyt) in Kaposi's sarcoma-associated herpesvirus (KSHV) have been identified and a virally encoded bZip protein, K8, has been shown to specifically bind to the origin. To map cis-acting elements within KSHV ori-Lyt that are required for DNA replication function and to define the nature of K8 bZip protein binding to the origin, we constructed consecutive internal deletion mutations across the core domain of a KSHV ori-Lyt and tested them for DNA replication function in a transient replication assay. This mutagenesis study allowed the identification of four components within the ori-Lyt, and all were indispensable for ori-Lyt function. The first component contains eight CCAAT/enhancer binding protein (C/EBP) binding motifs that organize as four spaced C/EBP palindromes. Each palindrome contains two head-to-head CCAAT consensus motifs that are separated by a 13- or 12-bp space sequence. Substitution mutagenesis of these C/EBP motifs showed that these C/EBP palindromes are required for both K8 binding and ori-Lyt dependent DNA replication. The second component is an 18-bp AT palindrome, which is essential for ori-Lyt function. The third component was determined to be a 32 bp previously unidentified sequence and is required for DNA replication. The last component consists of an open reading frame 50 (ORF50)/Rta responsive element (RRE) and a TATA box. We showed that the binding of an ORF50/Rta protein to the RRE was essential for ori-Lyt-dependent DNA replication. The presence of a functional RRE and a downstream TATA box suggested that this region serves as an ORF50/Rta-dependent promoter and a transcription event may be necessary for ori Lyt-dependent DNA replication. Using a luciferase reporter system, we demonstrated that the region of the RRE and TATA box constitutes an ORF50/Rta dependent promoter. Furthermore, a polyadenylated RNA of 1.4 kb was identified downstream of the promoter. PMID- 15280472 TI - Characterization of nucleocapsid binding by the measles virus and mumps virus phosphoproteins. AB - We report an analysis of the interaction between the P protein and the RNA associated N protein (N-RNA) for both measles and mumps viruses with proteins produced in a bacterial expression system. During this study, we verified that the C-terminal tail of the N protein is not required for nucleocapsid formation. For both measles and mumps virus N, truncated proteins encompassing amino acids 1 to 375 assemble into nucleocapsid-like particles within the bacterial cell. For measles virus N, the binding site for the P protein maps to residues 477 to 505 within the tail of the molecule, a sequence relatively conserved among the morbilliviruses. For mumps virus N, a binding site for the P protein maps to the assembly domain of N (residues 1 to 398), while no strong binding of the P protein to the tail of N was detected. These results suggest that the site of attachment for the polymerase varies among the paramyxoviruses. Pulldown experiments demonstrate that the last 50 amino acids of both measles virus and mumps virus P (measles virus P, 457 to 507; mumps virus P, 343 to 391) by themselves constitute the nucleocapsid-binding domain (NBD). Spectroscopic studies show that the NBD is predominantly alpha-helical in both viruses. However, only in measles virus P is the NBD stable and folded, having a lesser degree of tertiary organization in mumps virus P. With isothermal titration calorimetry, we demonstrate that the measles virus P NBD binds to residues 477 to 505 of measles virus N with 1:1 stoichiometry. The dissociation constant (K(d)) was determined to be 13 microM at 20 degrees C and 35 microM at 37 degrees C. Our data are consistent with a model in which an alpha-helical nucleocapsid binding domain, located at the C terminus of P, is responsible for tethering the viral polymerase to its template yet also suggest that, in detail, polymerase binding in morbilliviruses and rubulaviruses differs significantly. PMID- 15280473 TI - Suppression of herpes simplex virus 1 in MDBK cells via the interferon pathway. AB - Herpes simplex virus (HSV) normally undergoes productive infection in culture, causing cell destruction and plaque formation. Here we characterize an unusual pattern of HSV type 1 (HSV-1) infection in MDBK cells which surprisingly results in suppression of replication, cell recovery, and maintenance of virus. Compared to Vero cells, MDBK cells supported a normal productive infection at a high multiplicity with complete cell destruction. At low multiplicity, HSV also showed an identical initial specific infectivity in the two cell types. Thereafter, the progression of infection was radically different. In contrast to the rapid plaque expansion and eventual destruction in Vero monolayers, in MDBK cells, after initial plaque formation, plaque size actually decreased and, with time, monolayers recovered. Using a green fluorescent protein (GFP)-VP16-expressing virus, we monitored infection in live individual plaques. After early stages of intense GFP-VP16 expression, expression regressed to a thin boundary at the edge of the plaques and was completely suppressed by 10 days. Cells lacking expression then began to grow into the plaque boundaries. Furthermore, following media replacement, individual cells expressing GFP-VP16 could be observed reinitiating infection. The results indicated the production of a potent inhibitory component during infection in MDBK cells, and we show the continued and prolonged presence of interferon in the medium, at times when there was no longer evidence of ongoing productive infection. We exploited the ability of V protein of simian virus 5 to degrade Stat1 and prevent interferon signaling. We established MDBK cells constitutively expressing the V protein with the resultant loss of Stat1. In comparison to the parental cells, infection in these cells now progressed at a rapid rate with expanding plaque formation. We believe the conclusions have significant implications for the study of HSV-1 and interferon signaling both in culture and in animal models. PMID- 15280474 TI - Spirodiketopiperazine-based CCR5 inhibitor which preserves CC-chemokine/CCR5 interactions and exerts potent activity against R5 human immunodeficiency virus type 1 in vitro. AB - We identified a novel spirodiketopiperazine (SDP) derivative, AK602/ONO4128/GW873140, which specifically blocked the binding of macrophage inflammatory protein 1alpha (MIP-1alpha) to CCR5 with a high affinity (K(d) of approximately 3 nM), potently blocked human immunodeficiency virus type 1 (HIV-1) gp120/CCR5 binding and exerted potent activity against a wide spectrum of laboratory and primary R5 HIV-1 isolates, including multidrug-resistant HIV-1 (HIV-1(MDR)) (50% inhibitory concentration values of 0.1 to 0.6 nM) in vitro. AK602 competitively blocked the binding to CCR5 expressed on Chinese hamster ovary cells of two monoclonal antibodies, 45523, directed against multidomain epitopes of CCR5, and 45531, specific against the C-terminal half of the second extracellular loop (ECL2B) of CCR5. AK602, despite its much greater anti-HIV-1 activity than other previously published CCR5 inhibitors, including TAK-779 and SCH-C, preserved RANTES (regulated on activation normal T-cell expressed and secreted) and MIP-1beta binding to CCR5(+) cells and their functions, including CC-chemokine-induced chemotaxis and CCR5 internalization, while TAK-779 and SCH-C fully blocked the CC-chemokine/CCR5 interactions. Pharmacokinetic studies revealed favorable oral bioavailability in rodents. These data warrant further development of AK602 as a potential therapeutic for HIV-1 infection. PMID- 15280475 TI - Baculovirus vectors elicit antigen-specific immune responses in mice. AB - To characterize the induction of antigen-specific immune response mediated by baculovirus, vectors expressing the E2 glycoprotein of hepatitis C virus or the carcinoembryonic antigen (CEA) under the control of the cytomegalovirus immediate early promoter-enhancer were constructed. Additionally, a baculovirus vector encoding the E2 glycoprotein (Bac-G-E2) and expressing vesicular stomatitis virus glycoprotein (VSV-G) in the viral envelope was generated by inserting the VSV-G coding sequence downstream of the polyhedrin promoter. Mice were subjected to intramuscular, intranasal, or subcutaneous inoculations with Bac-E2 and the cellular immune response was monitored by ELISPOT and intracellular staining. Additionally, humoral response was monitored by titrating anti-E2 antibodies. Induction of a measurable anti-E2 T-cell response was observed only after intramuscular injection and was predominantly CD8(+) specific. The immunogenic properties of baculovirus as vaccine vector were not restricted to E2 because a CEA-specific CD4(+) T-cell response was observed upon intramuscular injection of Bac-CEA. Interestingly, the Bac-G-E2 vector was shown to be a more efficient immunogen than Bac-E2, in view of the 10-fold difference in the minimal dose required to elicit a measurable T-cell response upon intramuscular injection. Induction of inflammatory cytokines such as gamma interferon, tumor necrosis factor alpha, and interleukin-6 was detected as early as 6 h postinjection of Bac G-E2. Most importantly, both vectors elicited CD8(+) T cells with effector function capable of lysing target cells loaded with a hepatitis C virus-specific epitope. Additionally, enhanced NK cytolytic activity was detected in immunized mice. Thus, these results further demonstrate that baculovirus may be considered a useful vector for gene therapy. PMID- 15280477 TI - Papillomavirus-mediated neoplastic progression is associated with reciprocal changes in JAGGED1 and manic fringe expression linked to notch activation. AB - Infection by high-risk human papillomaviruses (HPV) and persistent expression of viral oncogenes E6 and E7 are causally linked to the development of cervical cancer. These oncogenes are necessary but insufficient for complete transformation of human epithelial cells in vivo. Intracellular Notch1 protein is detected in invasive cervical carcinomas (ICC), and truncated Notch1 alleles complement the function of E6/E7 in the transformation of human epithelial cells. Here we investigate potential mechanisms of Notch activation in a human cervical neoplasia. We have analyzed human cervical lesions and serial passages of an HPV type 16-positive human cervical low-grade lesion-derived cell line, W12, that shows abnormalities resembling those seen in cervical neoplastic progression in vivo. Late-passage, but not early-passage, W12 and progression of the majority of human high-grade cervical lesions to ICC showed upregulation of Notch ligand and Jagged1 and downregulation of Manic Fringe, a negative regulator of Jagged1 Notch1 signaling. Concomitantly, an increase in Notch/CSL (CBF1, Suppressor of Hairless, Lag1)-driven reporter activity and a decrease in Manic Fringe upstream regulatory region (MFng-URR)-driven reporter activity was observed in late passage versus early passage W12. Analysis of the MFng-URR revealed that Notch signaling represses this gene through Hairy Enhancer of Split 1, a transcriptional target of the Notch pathway. Expression of Manic Fringe by a recombinant adenovirus, dominant-negative Jagged1, or small interfering RNA against Jagged1 inhibits the tumorigenicity of CaSki, an ICC-derived cell line that was previously shown to be susceptible to growth inhibition induced by antisense Notch1. We suggest that activation of Notch in cervical neoplasia is Jagged1 dependent and that its susceptibility to the influence of Manic Fringe is of therapeutic value. PMID- 15280476 TI - Model for the interaction of gammaherpesvirus 68 RING-CH finger protein mK3 with major histocompatibility complex class I and the peptide-loading complex. AB - The mK3 protein of gammaherpesvirus 68 and the kK5 protein of Kaposi's sarcoma associated herpesvirus are members of a family of structurally related viral immune evasion molecules that all possess a RING-CH domain with ubiquitin ligase activity. These proteins modulate the expression of major histocompatibility complex class I molecules (mK3 and kK5) as well as other molecules like ICAM-1 and B7.2 (kK5). Previously, mK3 was shown to ubiquitinate nascent class I molecules, resulting in their rapid degradation, and this process was found to be dependent on TAP and tapasin, endoplasmic reticulum molecules involved in class I assembly. Here, we demonstrate that in murine cells, kK5 does not affect class I expression but does downregulate human B7.2 molecules in a TAP/tapasin independent manner. These differences in substrate specificity and TAP/tapasin dependence between mK3 and kK5 permitted us, using chimeric molecules, to map the sites of mK3 interaction with TAP/tapasin and to determine the requirements for substrate recognition by mK3. Our findings indicate that mK3 interacts with TAP1 and -2 via their C-terminal domains and with class I molecules via their N terminal domains. Furthermore, by orienting the RING-CH domain of mK3 appropriately with respect to class I, mK3 binding to TAP/tapasin, rather than the presence of unique sequences in class I, appears to be the primary determinant of substrate specificity. PMID- 15280478 TI - Human coronavirus 229E binds to CD13 in rafts and enters the cell through caveolae. AB - CD13, a receptor for human coronavirus 229E (HCoV-229E), was identified as a major component of the Triton X-100-resistant membrane microdomain in human fibroblasts. The incubation of living fibroblasts with an anti-CD13 antibody on ice gave punctate labeling that was evenly distributed on the cell surface, but raising the temperature to 37 degrees C before fixation caused aggregation of the labeling. The aggregated labeling of CD13 colocalized with caveolin-1 in most cells. The HCoV-229E virus particle showed a binding and redistribution pattern that was similar to that caused by the anti-CD13 antibody: the virus bound to the cell evenly when incubated on ice but became colocalized with caveolin-1 at 37 degrees C; importantly, the virus also caused sequestration of CD13 to the caveolin-1-positive area. Electron microscopy confirmed that HCoV-229E was localized near or at the orifice of caveolae after incubation at 37 degrees C. The depletion of plasmalemmal cholesterol with methyl beta-cyclodextrin significantly reduced the HCoV-229E redistribution and subsequent infection. A caveolin-1 knockdown by RNA interference also reduced the HCoV-229E infection considerably. The results indicate that HCoV-229E first binds to CD13 in the Triton X-100-resistant microdomain, then clusters CD13 by cross-linking, and thereby reaches the caveolar region before entering cells. PMID- 15280479 TI - Nitric oxide as an endogenous mutagen for Sendai virus without antiviral activity. AB - Nitric oxide (NO) may affect the genomes of various pathogens, and this mutagenesis is of particular interest for viral pathogenesis and evolution. Here, we investigated the effect of NO on viral replication and mutation. Exogenous or endogenous NO had no apparent antiviral effect on influenza A virus and Sendai virus. The mutagenic potential of NO was analyzed with Sendai virus fused to a green fluorescent protein (GFP) gene (GFP-SeV). GFP-SeV was cultured in SW480 cells transfected with a vector expressing inducible NO synthase (iNOS). The mutation frequency of GFP-SeV was examined by measuring loss of GFP fluorescence of the viral plaques. GFP-SeV mutation frequency in iNOS-SW480 cells was much higher than that in parent SW480 cells and was reduced to the level of mutation frequency in the parent cells by treatment with an NO synthase (NOS) inhibitor. Immunocytochemistry showed generation of more 8-nitroguanosine in iNOS-SW480 cells than in SW480 cells without iNOS transfection. Authentic 8-nitroguanosine added exogenously to GFP-SeV-infected CV-1 cells increased the viral mutation frequency. Profiles of the GFP gene mutations induced by 8-nitroguanosine appeared to resemble those of mutations occurring in mouse lungs in vivo. A base substitution that was characteristic of both mutants (those induced by 8 nitroguanosine and those occurring in vivo) was a C-to-U transition. NO-dependent oxidative stress in iNOS-SW480 cells was also evident. Together, the results indicate unambiguously that NO has mutagenic potential for RNA viruses such as Sendai virus without affecting viral replication, possibly via 8-nitroguanosine formation and cellular oxidative stress. PMID- 15280480 TI - Human cytomegalovirus-encoded interleukin-10 homolog inhibits maturation of dendritic cells and alters their functionality. AB - Interleukin-10 (IL-10) suppresses the maturation and cytokine production of dendritic cells (DCs), key regulators of adaptive immunity, and prevents the activation and polarization of naive T cells towards protective gamma interferon producing effectors. We hypothesized that human cytomegalovirus (HCMV) utilizes its viral IL-10 homolog (cmvIL-10) to attenuate DC functionality, thereby subverting the efficient induction of antiviral immune responses. RNA and protein analyses demonstrated that the cmvIL-10 gene was expressed with late gene kinetics. Treatment of immature DCs (iDCs) with supernatant from HCMV-infected cultures inhibited both the lipopolysaccharide-induced DC maturation and proinflammatory cytokine production. These inhibitory effects were specifically mediated through the IL-10 receptor and were not observed when DCs were treated with supernatant of cells infected with a cmvIL-10-knockout mutant. Incubation of iDCs with recombinant cmvIL-10 recapitulated the inhibition of maturation. Furthermore, cmvIL-10 had pronounced long-term effects on those DCs that could overcome this inhibition of maturation. It enhanced the migration of mature DCs (mDCs) towards the lymph node homing chemokine but greatly reduced their cytokine production. The inability of mDCs to secrete IL-12 was maintained, even when they were restimulated by the activated T-cell signal CD40 ligand in the absence of cmvIL-10. Importantly, cmvIL-10 potentiates these anti-inflammatory effects, at least partially, by inducing endogenous cellular IL-10 expression in DCs. Collectively, we show that cmvIL-10 causes long-term functional alterations at all stages of DC activation. PMID- 15280481 TI - Putative autocleavage of outer capsid protein micro1, allowing release of myristoylated peptide micro1N during particle uncoating, is critical for cell entry by reovirus. AB - Several nonenveloped animal viruses possess an autolytic capsid protein that is cleaved as a maturation step during assembly to yield infectious virions. The 76 kDa major outer capsid protein micro1 of mammalian orthoreoviruses (reoviruses) is also thought to be autocatalytically cleaved, yielding the virion-associated fragments micro1N (4 kDa; myristoylated) and micro1C (72 kDa). In this study, we found that micro1 cleavage to yield micro1N and micro1C was not required for outer capsid assembly but contributed greatly to the infectivity of the assembled particles. Recoated particles containing mutant, cleavage-defective micro1 (asparagine --> alanine substitution at amino acid 42) were competent for attachment; processing by exogenous proteases; structural changes in the outer capsid, including micro1 conformational change and sigma1 release; and transcriptase activation but failed to mediate membrane permeabilization either in vitro (no hemolysis) or in vivo (no coentry of the ribonucleotoxin alpha sarcin). In addition, after these particles were allowed to enter cells, the delta region of micro1 continued to colocalize with viral core proteins in punctate structures, indicating that both elements remained bound together in particles and/or trapped within the same subcellular compartments, consistent with a defect in membrane penetration. If membrane penetration activity was supplied in trans by a coinfecting genome-deficient particle, the recoated particles with cleavage-defective micro1 displayed much higher levels of infectivity. These findings led us to propose a new uncoating intermediate, at which particles are trapped in the absence of micro1N/micro1C cleavage. We additionally showed that this cleavage allowed the myristoylated, N-terminal micro1N fragment to be released from reovirus particles during entry-related uncoating, analogous to the myristoylated, N-terminal VP4 fragment of picornavirus capsid proteins. The results thus suggest that hydrophobic peptide release following capsid protein autocleavage is part of a general mechanism of membrane penetration shared by several diverse nonenveloped animal viruses. PMID- 15280482 TI - Evidence that rabies virus forms different kinds of fusion machines with different pH thresholds for fusion. AB - Fusion of rabies virus with membranes is triggered at a low pH and is mediated by a viral glycoprotein (G). Fusion of rabies virus with liposomes was monitored by using a lipid mixing assay based on fluorescence resonance energy transfer. Fusion was detected below pH 6.4, and its extent increased with H(+) concentrations to be maximal around pH 6.15. The origin of the partial fusion activity of rabies virus under suboptimal pH conditions (i.e., between pH 6.15 and 6.4) was investigated. We demonstrate unambiguously that fusion at a suboptimal pH is distinct from the phenomenon of low-pH-induced inactivation and that it is not due to heterogeneity of the virus population. We also show that viruses that do not fuse under suboptimal pH conditions are indeed bound to the target liposomes and that the fusion complexes they have formed are blocked at an early stage of the fusion pathway. Our conclusion is that along the fusion reaction, different kinds of fusion machines with different pH thresholds for fusion can be formed. Possible explanations of this difference of pH sensitivity are discussed. PMID- 15280483 TI - Vbeta14(+) T cells mediate the vaccine-enhanced disease induced by immunization with respiratory syncytial virus (RSV) G glycoprotein but not with formalin inactivated RSV. AB - Mice immunized with respiratory syncytial virus (RSV) G glycoprotein or with formalin-inactivated RSV (FI-RSV) exhibit severe disease following RSV challenge. This results in type 2 cytokine production and pulmonary eosinophilia, both hallmarks of vaccine-enhanced disease. RSV G-induced T-cell responses were shown to be restricted to CD4(+) T cells expressing Vbeta14 in the T-cell receptor (TCR), and the deletion of these T cells resulted in less severe disease. We therefore examined the role of Vbeta14(+) T cells in FI-RSV-induced disease. BALB/c mice were immunized with vaccinia virus expressing secreted RSV G (vvGs) or with FI-RSV. At the time of challenge with live RSV, mice were injected with antibody to the Vbeta14 component of the TCR. vvGs-immunized mice treated with anti-Vbeta14 had reduced cytokine levels in the lung. Eosinophil recruitment to the lung was also significantly reduced. In contrast, depletion of Vbeta14(+) T cells in FI-RSV-immunized mice had little impact on cytokine production or pulmonary eosinophilia. An analysis of TCR Vbeta chain usage confirmed a bias toward Vbeta14 expression on CD4(+) T cells from vvGs-immunized mice, whereas the CD4(+) T cells in FI-RSV-immunized mice expressed a diverse array of Vbeta chains. These data show that although FI-RSV and vvGs induce responses resulting in similar immunopathology, the T-cell repertoire mediating the response is different for each immunogen and suggest that the immune responses elicited by RSV G are not the basis for FI-RSV vaccine-enhanced disease. PMID- 15280484 TI - Antiretroviral drug resistance mutations in human immunodeficiency virus type 1 reverse transcriptase increase template-switching frequency. AB - Template-switching events during reverse transcription are necessary for completion of retroviral replication and recombination. Structural determinants of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) that influence its template-switching frequency are not known. To identify determinants of HIV-1 RT that affect the frequency of template switching, we developed an in vivo assay in which RT template-switching events during viral replication resulted in functional reconstitution of the green fluorescent protein gene. A survey of single amino acid substitutions near the polymerase active site or deoxynucleoside triphosphate-binding site of HIV-1 RT indicated that several substitutions increased the rate of RT template switching. Several mutations associated with resistance to antiviral nucleoside analogs (K65R, L74V, E89G, Q151N, and M184I) dramatically increased RT template-switching frequencies by two- to sixfold in a single replication cycle. In contrast, substitutions in the RNase H domain (H539N, D549N) decreased the frequency of RT template switching by twofold. Depletion of intracellular nucleotide pools by hydroxyurea treatment of cells used as targets for infection resulted in a 1.8-fold increase in the frequency of RT template switching. These results indicate that the dynamic steady state between polymerase and RNase H activities is an important determinant of HIV-1 RT template switching and establish that HIV-1 recombination occurs by the previously described dynamic copy choice mechanism. These results also indicate that mutations conferring resistance to antiviral drugs can increase the frequency of RT template switching and may influence the rate of retroviral recombination and viral evolution. PMID- 15280485 TI - Matrix gene of influenza a viruses isolated from wild aquatic birds: ecology and emergence of influenza a viruses. AB - Wild aquatic birds are the primary reservoir of influenza A viruses, but little is known about the viruses' gene pool in wild birds. Therefore, we investigated the ecology and emergence of influenza viruses by conducting phylogenetic analysis of 70 matrix (M) genes of influenza viruses isolated from shorebirds and gulls in the Delaware Bay region and from ducks in Alberta, Canada, during >18 years of surveillance. In our analysis, we included 61 published M genes of isolates from various hosts. We showed that M genes of Canadian duck viruses and those of shorebird and gull viruses in the Delaware Bay shared ancestors with the M genes of North American poultry viruses. We found that North American and Eurasian avian-like lineages are divided into sublineages, indicating that multiple branches of virus evolution may be maintained in wild aquatic birds. The presence of non-H13 gull viruses in the gull-like lineage and of H13 gull viruses in other avian lineages suggested that gulls' M genes do not preferentially associate with the H13 subtype or segregate into a distinct lineage. Some North American avian influenza viruses contained M genes closely related to those of Eurasian avian viruses. Therefore, there may be interregional mixing of the two clades. Reassortment of shorebird M and HA genes was evident, but there was no correlation among the HA or NA subtype, M gene sequence, and isolation time. Overall, these results support the hypothesis that influenza viruses in wild waterfowl contain distinguishable lineages of M genes. PMID- 15280486 TI - Chimeras of duck and heron hepatitis B viruses provide evidence for functional interactions between viral components of pregenomic RNA encapsidation. AB - Packaging of hepadnavirus pregenomic RNA (pgRNA) into capsids, or encapsidation, requires several viral components. The viral polymerase (P) and the capsid subunit (C) are necessary for pgRNA encapsidation. Previous studies of duck hepatitis B virus (DHBV) indicated that two cis-acting sequences on pgRNA are required for encapsidation: epsilon, which is near the 5' end of pgRNA, and region II, located near the middle of pgRNA. Later studies suggested that the intervening sequence between these two elements may also make a contribution. It has been demonstrated for DHBV that epsilon interacts with P to facilitate encapsidation, but it is not known how other cis-acting sequences contribute to encapsidation. We analyzed chimeras of DHBV and a related virus, heron hepatitis B virus (HHBV), to gain insight into the interactions between the various viral components during pgRNA encapsidation. We learned that having epsilon and P derived from the same virus was not sufficient for high levels of encapsidation, implying that other viral interactions contribute to encapsidation. Chimeric analysis showed that a large sequence containing region II may interact with P and/or C for efficient encapsidation. Further analysis demonstrated that possibly an RNA-RNA interaction between the intervening sequence and region II facilitates pgRNA encapsidation. Together, these results identify functional interactions among various viral components that contribute to pgRNA encapsidation. PMID- 15280487 TI - Human endogenous retrovirus family HERV-K(HML-5): status, evolution, and reconstruction of an ancient betaretrovirus in the human genome. AB - The human genome harbors numerous distinct families of so-called human endogenous retroviruses (HERV) which are remnants of exogenous retroviruses that entered the germ line millions of years ago. We describe here the hitherto little characterized betaretrovirus HERV-K(HML-5) family (named HERVK22 in Repbase) in greater detail. Out of 139 proviruses, only a few loci represent full-length proviruses, and many lack gag protease and/or env gene regions. We generated a consensus sequence from multiple alignment of 62 HML-5 loci that displays open reading frames for the four major retroviral proteins. Four HML-5 long terminal repeat (LTR) subfamilies were identified that are associated with monophyletic proviral bodies, implying different evolution of HML-5 LTRs and genes. Sequence analysis indicated that the proviruses formed approximately 55 million years ago. Accordingly, HML-5 proviral sequences were detected in Old World and New World primates but not in prosimians. No recent activity is associated with this HERV family. We also conclude that the HML-5 consensus sequence primer binding site is identical to methionine tRNA. Therefore, the family should be designated HERV-M. Our study provides important insights into the structure and evolution of the oldest betaretrovirus in the primate genome known to date. PMID- 15280488 TI - A short peptide at the amino terminus of the Sendai virus C protein acts as an independent element that induces STAT1 instability. AB - The Sendai virus C protein acts to dismantle the interferon-induced cellular antiviral state in an MG132-sensitive manner, in part by inducing STAT1 instability. This activity of C maps to the first 23 amino acids (C(1-23)) of the 204-amino-acid (aa)-long protein (C(1-204)). C(1-23) was found to act as an independent viral element that induces STAT1 instability, since this peptide fused to green fluorescent protein (C(1-23)/GFP) is at least as active as C(1 204) in this respect. This peptide also induces the degradation of C(1-23)/GFP and other proteins to which it is fused. Most of C(1-204), and particularly its amino-terminal half, is predicted to be structurally disordered. C(1-23) as a peptide was found to be disordered by circular dichroism, and the first 11 aa have a strong potential to form an amphipathic alpha-helix in low concentrations of trifluoroethanol, which is thought to mimic protein-protein interaction. The critical degradation-determining sequence of C(1-23) was mapped by mutation to eight residues near its N terminus: (4)FLKKILKL(11). All the large hydrophobic residues of (4)FLKKILKL(11), plus its ability to form an amphipathic alpha-helix, were found to be critical for STAT1 degradation. In contrast, C(1-23)/GFP self degradation did not require (8)ILKL(11), nor the ability to form an alpha-helix throughout this region. Remarkably, C(1-23)/GFP also stimulated C(1-204) degradation, and this degradation in trans required the same peptide determinants as for STAT1. Our results suggest that C(1-204) coordinates its dual activities of regulating viral RNA synthesis and counteracting the host innate antiviral response by sensing both its own intracellular concentration and that of STAT1. PMID- 15280489 TI - Determinants of virulence of classical swine fever virus strain Brescia. AB - Two related classical swine fever virus (CSFV) strain Brescia clones were isolated from blood samples from an infected pig. Virus C1.1.1 is a cell-adapted avirulent variant, whereas CoBrB is a virulent variant. Sequence analysis revealed 29 nucleic acid mutations in C1.1.1, resulting in 9 amino acid substitutions compared to the sequence of CoBrB (476)R. Using reverse genetics, parts of the genomes of these viruses, which contain differences that lead to amino acid changes, were exchanged. Animal experiments with chimeric viruses derived from C1.1.1 and CoBrB (476)R showed that a combination of amino acid changes in the structural and nonstructural regions reduced the virulence of CSFV in pigs. Moreover, the presence of a Leu at position 710 in structural envelope protein E2 seemed to be an important factor in the virulence of the virus. Changing the Leu at position 710 in the CoBrB (476)S variant into a His residue did not affect virulence. However, the (710)His in the C1.1.1/CoBrB virus, together with adaptive mutations (276)R, (476)R, and (477)I in E(rns), resulted in reduced virulence in pigs. These results indicated that mutations in E(rns) and E2 alone do not determine virulence in pigs. The results of in vitro experiments suggested that a high affinity for heparan sulfate of C1.1.1 E(rns) may reduce the spread of the C1.1.1/CoBrB virus in pigs and together with the altered surface structure of E2 caused by the (710)L-->H mutation may result in a less efficient infection of specific target cells in pigs. Both these features contributed to the attenuation of the C1.1.1/CoBrB virus in vivo. PMID- 15280490 TI - Human respiratory coronavirus OC43: genetic stability and neuroinvasion. AB - The complete genome sequences of the human coronavirus OC43 (HCoV-OC43) laboratory strain from the American Type Culture Collection (ATCC), and a HCoV OC43 clinical isolate, designated Paris, were obtained. Both genomes are 30,713 nucleotides long, excluding the poly(A) tail, and only differ by 6 nucleotides. These six mutations are scattered throughout the genome and give rise to only two amino acid substitutions: one in the spike protein gene (I958F) and the other in the nucleocapsid protein gene (V81A). Furthermore, the two variants were shown to reach the central nervous system (CNS) after intranasal inoculation in BALB/c mice, demonstrating neuroinvasive properties. Even though the ATCC strain could penetrate the CNS more effectively than the Paris 2001 isolate, these results suggest that intrinsic neuroinvasive properties already existed for the HCoV-OC43 ATCC human respiratory isolate from the 1960s before it was propagated in newborn mouse brains. It also demonstrates that the molecular structure of HCoV-OC43 is very stable in the environment (the two variants were isolated ca. 40 years apart) despite virus shedding and chances of persistence in the host. The genomes of the two HCoV-OC43 variants display 71, 53.1, and 51.2% identity with those of mouse hepatitis virus A59, severe acute respiratory syndrome human coronavirus Tor2 strain (SARS-HCoV Tor2), and human coronavirus 229E (HCoV-229E), respectively. HCoV-OC43 also possesses well-conserved motifs with regard to the genome sequence of the SARS-HCoV Tor2, especially in open reading frame 1b. These results suggest that HCoV-OC43 and SARS-HCoV may share several important functional properties and that HCoV-OC43 may be used as a model to study the biology of SARS-HCoV without the need for level three biological facilities. PMID- 15280491 TI - Hepatitis C virus infection activates the immunologic (type II) isoform of nitric oxide synthase and thereby enhances DNA damage and mutations of cellular genes. AB - Hepatitis C virus (HCV) infection causes hepatitis, hepatocellular carcinoma, and B-cell lymphomas in a significant number of patients. Previously we have shown that HCV infection causes double-stranded DNA breaks and enhances the mutation frequency of cellular genes, including proto-oncogenes and immunoglobulin genes. To determine the mechanisms, we studied in vitro HCV infection of cell culture. Here we report that HCV infection activated the immunologic (type II) isoform of nitric oxide (NO) synthase (NOS), i.e., inducible NOS (iNOS), thereby inducing NO, which in turn induced DNA breaks and enhanced the mutation frequencies of cellular genes. Treatment of HCV-infected cells with NOS inhibitors or small interfering RNA specific for iNOS abolished most of these effects. Expression of the core protein or nonstructural protein 3 (NS3), but not the other viral proteins, in B cells or hepatocytes induced iNOS and DNA breaks, which could be blocked by NOS inhibitors. The core protein also enhanced the mutation frequency of cellular genes in hepatocytes derived from HCV core transgenic mice compared with that in control mice. The iNOS promoter was activated more than fivefold in HCV-infected cells, as revealed by a luciferase reporter assay driven by the iNOS promoter. Similarly, the core and NS3 proteins also induced the same effects. Therefore, we conclude that HCV infection can stimulate the production of NO through activation of the gene for iNOS by the viral core and NS3 proteins. NO causes DNA breaks and enhances DNA mutation. This sequence of events provides a mechanism for HCV pathogenesis and oncogenesis. PMID- 15280492 TI - Beyond help: direct effector functions of human immunodeficiency virus type 1 specific CD4(+) T cells. AB - The immune correlates of protection in human immunodeficiency virus type 1 (HIV 1) infection remain poorly defined, particularly the contribution of CD4(+) T cells. Here we explore the effector functions of HIV-1-specific CD4(+) T cells. We demonstrate HIV-1 p24-specific CD4(+)-T-cell cytolytic activity in peripheral blood mononuclear cells directly ex vivo and after enrichment by antigen-specific stimulation. We further show that in a rare long-term nonprogressor, both an HIV 1-specific CD4(+)-T-cell clone and CD4(+) T cells directly ex vivo exert potent suppression of HIV-1 replication. Suppression of viral replication was dependent on cell-cell contact between the effector CD4(+) T cells and the target cells. While the antiviral effector activity of CD8(+) T cells has been well documented, these results strongly suggest that HIV-1-specific CD4(+) T cells are capable of directly contributing to antiviral immunity. PMID- 15280493 TI - Internalization of pseudorabies virus glycoprotein B is mediated by an interaction between the YQRL motif in its cytoplasmic domain and the clathrin associated AP-2 adaptor complex. AB - The cytoplasmic domain of pseudorabies virus (PRV) glycoprotein B (gB) contains three putative internalization motifs. Previously, we demonstrated that the tyrosine-based YQRL motif at positions 902 to 905, but not the YMSI motif at positions 864 to 867 or the LL doublet at positions 887 and 888, is required for correct functioning of gB during antibody-mediated internalization of PRV cell surface-bound glycoproteins. In the present study, we demonstrate that the YQRL motif is also crucial to allow spontaneous internalization of PRV gB, and thus, that spontaneous and antibody-mediated internalizations of PRV gB occur through closely related mechanisms. Furthermore, we found that PRV gB colocalizes with the cellular clathrin-associated AP-2 adaptor complex and that this colocalization depends on the YQRL motif. In addition, by coimmunoprecipitation assays, we found that during both spontaneous and antibody-dependent internalization, PRV gB physically interacts with AP-2, and that efficient interaction between gB and AP-2 required an intact YQRL motif. Collectively, these findings demonstrate for the first time that during internalization of an alphaherpesvirus envelope protein, i.e., PRV gB, a specific amino acid sequence in the cytoplasmic tail of the protein interacts with AP-2 and may constitute a common AP-2-mediated mechanism of internalization of alphaherpesvirus envelope proteins. PMID- 15280494 TI - Virus persistence in an animal model of multiple sclerosis requires virion attachment to sialic acid coreceptors. AB - Persistent Theiler's virus infection in the central nervous system (CNS) of mice provides a highly relevant animal model for multiple sclerosis. The low neurovirulence DA strain uses sialic acid as a coreceptor for cell binding before establishing infection. During adaptation of DA virus to growth in sialic acid deficient cells, three amino acid substitutions (G1100D, T1081I, and T3182A) in the capsid arose, and the virus no longer used sialic acid as a coreceptor. The adapted virus retained acute CNS virulence, but its persistence in the CNS, white matter inflammation, and demyelination were largely abrogated. Infection of murine macrophage but not oligodendrocyte cultures with the adapted virus was also significantly reduced. Substitution of G1100D in an infectious DA virus cDNA clone demonstrated a major role for this mutation in loss of sialic acid binding and CNS persistence. These data indicate a direct role for sialic acid binding in Theiler's murine encephalomyelitis virus persistence and chronic demyelinating disease. PMID- 15280495 TI - Porcine endogenous retroviruses infect cells lacking cognate receptors by an alternative pathway: implications for retrovirus evolution and xenotransplantation. AB - A PHQ motif near the amino termini of gammaretroviral envelope glycoprotein surface (SU) subunits is important for infectivity but not for incorporation into virions or binding to cognate receptors. The H residue of this motif is most critical, with all substitutions we tested being inactive. Interestingly, porcine endogenous retroviruses (PERVs) of all three host-range groups, A, B, and C, lack full PHQ motifs, but most members have an H residue at position 10. H10A PERV mutants are noninfectious but were efficiently transactivated by adding to the assays a PHQ-containing SU or receptor-binding subdomain (RBD) derived from a gibbon ape leukemia virus (GALV). A requirement of this transactivation was a functional GALV receptor on the cells. In contrast to this heterologous transactivation, PERV RBDs and SUs were inactive in all tested cells, including porcine ST-IOWA cells. Surprisingly, transactivation by GALV RBD enabled wild type or H10A mutant PERVs of all three host-range groups to efficiently infect cells from humans and rodents that lack functional PERV receptors and it substantially enhanced infectivities of wild-type PERVs, even for cells with PERV receptors. Thus, PERVs can suboptimally infect cells that contain cognate receptors or they can employ a transactivation pathway to more efficiently infect all cells. This ability to infect cells lacking cognate receptors was previously demonstrated only for nontransmissible variant gammaretroviruses with recombinant and mutant envelope glycoproteins. We conclude that some endogenously inherited mammalian retroviruses also have a receptor-independent means for overcoming host range and interference barriers, implying a need for caution in xenotransplantation, especially of porcine tissues. PMID- 15280496 TI - Expression in a recombinant murid herpesvirus 4 reveals the in vivo transforming potential of the K1 open reading frame of Kaposi's sarcoma-associated herpesvirus. AB - Murid herpesvirus 4 (commonly called MHV-68) is closely related to Kaposi's sarcoma-associated herpesvirus (KSHV) and provides an excellent model system for investigating gammaherpesvirus-associated pathogenesis. MHV-76 is a naturally occurring deletion mutant of MHV-68 that lacks 9,538 bp of the left end of the unique portion of the genome encoding nonessential pathogenesis-related genes. The KSHV K1 protein has been shown to transform rodent fibroblasts in vitro and common marmoset T lymphocytes in vivo. Using homologous recombination techniques, we successfully generated recombinants of MHV-76 that encode green fluorescent protein (MHV76-GFP) and KSHV K1 (MHV76-K1). The replication of MHV76-GFP and MHV76-K1 in cell culture was identical to that of MHV-76. However, infection of BALB/c mice via the intranasal route revealed that MHV76-K1 replicated to a 10 fold higher titer than MHV76-GFP in the lungs at day 5 postinfection (p.i.). We observed type 2 pneumocyte proliferation in areas of consolidation and interstitial inflammation of mice infected with MHV76-K1 at day 10 p.i. MHV76-K1 established a 2- to 3-fold higher latent viral load than MHV76-GFP in the spleens of infected mice on days 10 and 14 p.i., although this was 10-fold lower than that established by wild-type MHV-76. A salivary gland tumor was present in one of four mice infected with MHV76-K1, as well as an increased inflammatory response in the lungs at day 120 p.i. compared with that of mice infected with MHV-76 and MHV76-GFP. PMID- 15280497 TI - Vaccinia virus morphogenesis: a13 phosphoprotein is required for assembly of mature virions. AB - The 70-amino-acid A13L protein is a component of the vaccinia virus membrane. We demonstrate here that the protein is expressed at late times of infection, undergoes phosphorylation at a serine residue(s), and becomes encapsidated in a monomeric form. Phosphorylation is dependent on Ser40, which lies within the proline-rich motif SPPP. Because phosphorylation of the A13 protein is only minimally affected by disruption of the viral F10 kinase or H1 phosphatase, a cellular kinase is likely to be involved. We generated an inducible recombinant in which A13 protein expression is dependent upon the inclusion of tetracycline in the culture medium. Repression of the A13L protein spares the biochemical progression of the viral life cycle but arrests virion morphogenesis. Virion assembly progresses through the formation of immature virions (IVs); however, these virions do not acquire nucleoids, and DNA crystalloids accumulate in the cytoplasm. Further development into intracellular mature virions is blocked, causing a 1,000-fold decrease in the infectious virus yield relative to that obtained in the presence of the inducer. We also determined that the temperature sensitive phenotype of the viral mutant Cts40 is due to a nucleotide transition within the A13L gene that causes a Thr(48)-->Ile substitution. This substitution disrupts the function of the A13 protein but does not cause thermolability of the protein; at the nonpermissive temperature, virion morphogenesis arrests at the stage of IV formation. The A13L protein, therefore, is part of a newly recognized group of membrane proteins that are dispensable for the early biogenesis of the virion membrane but are essential for virion maturation. PMID- 15280498 TI - Classic AIDS in a sooty mangabey after an 18-year natural infection. AB - Prevailing theory holds that simian immunodeficiency virus (SIV) infections are nonpathogenic in their natural simian hosts and that lifelong infections persist without disease. Numerous studies have reported that SIV-infected sooty mangabeys (SMs; Cercocebus atys) remain disease free for up to 24 years despite relatively high levels of viral replication. Here, we report that classic AIDS developed after an 18-year incubation in an SM (E041) with a natural SIVsm infection. Unlike that described in previous reports of SIV-related disease in SMs, the SIVsm infecting E041 was not first passaged through macaques; moreover, SM E041 was simian T-cell leukemia virus antibody negative. SM E041 was euthanized in 2002 after being diagnosed with severe disseminated B-cell lymphoma. The plasma virus load had been approximately the same for 16 years when a 100-fold increase in virus load occurred in years 17 and 18. Additional findings associated with AIDS were CD4(+)-cell decline, loss of p27 core antibody, and loss of control of SIVsm replication with disseminated giant cell disease. These findings suggest that the time to development of AIDS exceeds the average lifetime of SMs in the wild and that the principal adaptation of SIV to its natural African hosts does not include complete resistance to disease. Instead, AIDS may develop slowly, even in the presence of high virus loads. However, a long-term relatively high virus load, such as that in SM E041, is consistent with AIDS development in less than 18 years in humans and macaques. Therefore, the results also suggest that SMs have a special mechanism for resisting AIDS development. PMID- 15280499 TI - Heparan sulfate-independent infection attenuates high-neurovirulence GDVII virus induced encephalitis. AB - The high-neurovirulence Theiler's murine encephalomyelitis virus (TMEV) strain GDVII uses heparan sulfate (HS) as a coreceptor to enter target cells. We report here that GDVII virus adapted to growth in HS-deficient cells exhibited two amino acid substitutions (R3126L and N1051S) in the capsid and no longer used HS as a coreceptor. Infectious-virus yields in CHO cells were 25-fold higher for the adapted virus than for the parental GDVII virus, and the neurovirulence of the adapted virus in intracerebrally inoculated mice was substantially attenuated. The adapted virus showed altered cell tropism in the central nervous systems of mice, shifting from cerebral and brainstem neurons to spinal cord anterior horn cells; thus, severe poliomyelitis, but not acute encephalitis, was observed in infected mice. These data indicate that the use of HS as a coreceptor by GDVII virus facilitates cell entry and plays an important role in cell tropism and neurovirulence in vivo. PMID- 15280500 TI - Characterization of the mouse adeno-associated virus AAVS1 ortholog. AB - The nonpathogenic human adeno-associated virus (AAV) has developed a mechanism to integrate its genome into human chromosome 19 at 19q13.4 (termed AAVS1), thereby establishing latency. Here, we provide evidence that the chromosomal signals required for site-specific integration are conserved in the mouse genome proximal to the recently identified Mbs85 gene. These sequence motifs can be specifically nicked by the viral Rep protein required for the initiation of site-specific AAV DNA integration. Furthermore, these signals can serve as a minimal origin for Rep dependent DNA replication. In addition, we isolated the mouse Mbs85 proximal promoter and show transcriptional activity in three mouse cell lines. PMID- 15280501 TI - Ngari virus is a Bunyamwera virus reassortant that can be associated with large outbreaks of hemorrhagic fever in Africa. AB - Two isolates of a virus of the genus Orthobunyavirus (family Bunyaviridae) were obtained from hemorrhagic fever cases during a large disease outbreak in East Africa in 1997 and 1998. Sequence analysis of regions of the three genomic RNA segments of the virus (provisionally referred to as Garissa virus) suggested that it was a genetic reassortant virus with S and L segments derived from Bunyamwera virus but an M segment from an unidentified virus of the genus Orthobunyavirus. While high genetic diversity (52%) was revealed by analysis of virus M segment nucleotide sequences obtained from 21 members of the genus Orthobunyavirus, the Garissa and Ngari virus M segments were almost identical. Surprisingly, the Ngari virus L and S segments showed high sequence identity with those of Bunyamwera virus, showing that Garissa virus is an isolate of Ngari virus, which in turn is a Bunyamwera virus reassortant. Ngari virus should be considered when investigating hemorrhagic fever outbreaks throughout sub-Saharan Africa. PMID- 15280502 TI - Immune escape precedes breakthrough human immunodeficiency virus type 1 viremia and broadening of the cytotoxic T-lymphocyte response in an HLA-B27-positive long term-nonprogressing child. AB - The emergence of cytotoxic T-lymphocyte (CTL) escape mutations in human immunodeficiency virus type 1 (HIV-1) proteins has been anecdotally associated with progression to AIDS, but it has been difficult to determine whether viral mutation is the cause or the result of increased viral replication. Here we describe a perinatally HIV-infected child who maintained a plasma viral load of <400 copies/ml for almost a decade until a nonbinding escape mutation emerged within the immunodominant CTL epitope. The child subsequently experienced a reemergence of HIV-1 viremia accompanied by a marked increase in the number of CTL epitopes targeted. This temporal pattern suggests that CD8 escape can play a causal role in the loss of immune control. PMID- 15280503 TI - RNA interference directed against Poly(ADP-Ribose) polymerase 1 efficiently suppresses human immunodeficiency virus type 1 replication in human cells. AB - We established small interfering RNA (siRNA) directed against poly(ADP-ribose) polymerase 1 (PARP-1) that effectively reduces the expression of PARP-1 in two human cell lines. Established siRNA against PARP-1 significantly suppressed human immunodeficiency virus type 1 (HIV-1) replication, as well as the activation of the integrated HIV-1 long terminal repeat promoter. These results indicate that PARP-1 is required for efficient HIV-1 replication in human cells. We propose that PARP-1 may serve as a cellular target for RNA interference-mediated gene silencing to inhibit HIV-1 replication. PMID- 15280504 TI - The gene encoding the nucleocapsid protein of Gill-associated nidovirus of Penaeus monodon prawns is located upstream of the glycoprotein gene. AB - The ORF2 gene of Gill-associated virus (GAV) of Penaeus monodon prawns resides 93 nucleotides downstream of the ORF1a-ORF1b gene and encodes a 144-amino-acid hydrophilic polypeptide (15,998 Da; pI, 9.75) containing 20 basic (14%) and 13 acidic (9%) residues and 19 prolines (13%). Antiserum to a synthetic ORF2 peptide or an Escherichia coli-expressed glutathione S-transferase-ORF2 fusion protein detected a 20-kDa protein in infected lymphoid organ and gill tissues in Western blots. The GAV ORF2 fusion protein antiserum also cross-reacted with the p20 nucleoprotein in virions of the closely related Yellow head virus. By immuno-gold electron microscopy, it was observed that the ORF2 peptide antibody localized to tubular GAV nucleocapsids, often at the ends or at lateral cross sections. As GAV appears to contain only two structural protein genes (ORF2 and ORF3), these data indicate that GAV differs from vertebrate nidoviruses in that the gene encoding the nucleocapsid protein is located upstream of the gene encoding the virion glycoproteins. PMID- 15280505 TI - Multiplicity of human immunodeficiency virus infections in lymphoid tissue. AB - Human immunodeficiency virus type 1 (HIV-1)-infected splenocytes in humans were recently shown to harbor three to four proviruses per cell on average (A. Jung et al., Nature 418:144, 2002). However, the mechanisms that lead to such extensive multiple infections are not understood. Here, we find by using mathematical analysis that two mechanisms quantitatively capture the distribution of proviral genomes in HIV-1-infected splenocytes, one where multiple genomes are acquired one at a time in a series of sequential infectious contacts of a target cell with free virions and infected cells, and the other where cell-to-cell transmission of multiple virions or genomes results from a single infectious contact of a target cell with an infected cell. The two mechanisms imply different genetic diversities of proviruses within an infected cell and therefore different rates of emergence of drug resistance via recombination. PMID- 15280506 TI - Functional compensation of a detrimental amino acid substitution in a cytotoxic-T lymphocyte epitope of influenza a viruses by comutations. AB - Influenza A viruses accumulate amino acid substitutions in cytotoxic-T-lymphocyte (CTL) epitopes, allowing these viruses to escape from CTL immunity. The arginine to-glycine substitution at position 384 of the viral nucleoprotein is associated with escape from CTLs. Introduction of the R384G substitution in the nucleoprotein gene segment of influenza virus A/Hong Kong/2/68 by site-directed mutagenesis was detrimental to viral fitness. Introduction of one of the comutations associated with R384G, E375G, partially restored viral fitness and nucleoprotein functionality. We hypothesized that influenza A viruses need to overcome functional constraints to accumulate mutations in CTL epitopes and escape from CTLs. PMID- 15280507 TI - SMC1 coordinates DNA double-strand break repair pathways. AB - The SMC1/SMC3 heterodimer acts in sister chromatid cohesion, and recent data indicate a function in DNA double-strand break repair (DSBR). Since this role of SMC proteins has remained largely elusive, we explored interactions between SMC1 and the homologous recombination (HR) or non-homologous end-joining (NHEJ) pathways for DSBR in Saccharomyces cerevisiae. Analysis of conditional single- and double mutants of smc1-2 with rad52Delta, rad54Delta, rad50Delta or dnl4Delta illustrates a significant contribution of SMC1 to the overall capacity of cells to repair DSBs. smc1 but not smc2 mutants show increased hypersensitivity of HR mutants to ionizing irradiation and to the DNA crosslinking agent cis-platin. Haploid, but not diploid smc1-2 mutants were severely affected in repairing multiple genomic DNA breaks, suggesting a selective role of SMC1 in sister chromatid recombination. smc1-2 mutants were also 15-fold less efficient and highly error-prone in plasmid end-joining through the NHEJ pathway. Strikingly, inactivation of RAD52 or RAD54 fully rescued efficiency and accuracy of NHEJ in the smc1 background. Therefore, we propose coordination of HR and NHEJ processes by Smc1p through interaction with the RAD52 pathway. PMID- 15280508 TI - Symmetry elements in DNA structure important for recognition/methylation by DNA [amino]-methyltransferases. AB - The phage T4Dam and EcoDam DNA-[adenine-N6] methyltransferases (MTases) methylate GATC palindromic sequences, while the BamHI DNA-[cytosine-N4] MTase methylates the GGATCC palindrome (which contains GATC) at the internal cytosine residue. We compared the ability of these enzymes to interact productively with defective duplexes in which individual elements were deleted on one chain. A sharp decrease in kcat was observed for all three enzymes if a particular element of structural symmetry was disrupted. For the BamHI MTase, integrity of the ATCC was critical, while an intact GAT sequence was necessary for the activity of T4Dam, and an intact GA was necessary for EcoDam. Theoretical alignment of the region of best contacts between the protein and DNA showed that in the case of a palindromic interaction site, a zone covering the 5'-symmetric residues is located in the major groove versus a zone of contact covering the 3'-symmetric residues in the minor groove. Our data fit a simple rule of thumb that the most important contacts are aligned around the methylation target base: if the target base is in the 5' half of the palindrome, the interaction between the enzyme and the DNA occurs mainly in the major groove; if it is in the 3' half, the interaction occurs mainly in the minor groove. PMID- 15280509 TI - GATA-1 and NF-Y cooperate to mediate erythroid-specific transcription of Gfi-1B gene. AB - Expression of Gfi (growth factor-independence)-1B, a Gfi-1-related transcriptional repressor, is restricted to erythroid lineage cells and is essential for erythropoiesis. We have determined the transcription start site of the human Gfi-1B gene and located its first non-coding exon approximately 7.82 kb upstream of the first coding exon. The genomic sequence preceding this first non coding exon has been identified to be its erythroid-specific promoter region in K562 cells. Using gel-shift and chromatin immunoprecipitation (ChIP) assays, we have demonstrated that NF-Y and GATA-1 directly participate in transcriptional activation of the Gfi-1B gene in K562 cells. Ectopic expression of GATA-1 markedly stimulates the activity of the Gfi-1B promoter in a non-erythroid cell line U937. Interestingly, our results have indicated that this GATA-1-mediated trans-activation is dependent on NF-Y binding to the CCAAT site. Here we conclude that functional cooperation between GATA-1 and NF-Y contributes to erythroid specific transcriptional activation of Gfi-1B promoter. PMID- 15280510 TI - Evolution of divergent DNA recognition specificities in VDE homing endonucleases from two yeast species. AB - Homing endonuclease genes (HEGs) are mobile DNA elements that are thought to confer no benefit to their host. They encode site-specific DNA endonucleases that perpetuate the element within a species population by homing and disseminate it between species by horizontal transfer. Several yeast species contain the VMA1 HEG that encodes the intein-associated VMA1-derived endonuclease (VDE). The evolutionary state of VDEs from 12 species was assessed by assaying their endonuclease activities. Only two enzymes are active, PI-ZbaI from Zygosaccharomyces bailii and PI-ScaI from Saccharomyces cariocanus. PI-ZbaI cleaves the Z.bailii recognition sequence significantly faster than the Saccharomyces cerevisiae site, which differs at six nucleotide positions. A mutational analysis indicates that PI-ZbaI cleaves the S.cerevisiae substrate poorly due to the absence of a contact that is analogous to one made in PI-SceI between Gln-55 and nucleotides +9/+10. PI-ZbaI cleaves the Z.bailii substrate primarily due to a single base-pair substitution (A/T+5 --> T/A+5). Structural modeling of the PI-ZbaI/DNA complex suggests that Arg-331, which is absent in PI SceI, contacts T/A+5, and the reduced activity observed in a PI-ZbaI R331A mutant provides evidence for this interaction. These data illustrate that homing endonucleases evolve altered specificity as they adapt to recognize alternative target sites. PMID- 15280511 TI - Potential interaction between telithromycin and warfarin. AB - OBJECTIVE: To report a case of warfarin-telithromycin interaction resulting in an elevated international normalized ratio (INR) and hemoptysis. CASE SUMMARY: A 73 year-old white man developed an elevated INR and mild hemoptysis as a result of an interaction between warfarin and telithromycin 800 mg/day. The INR increased from 3.1 before telithromycin was started to 11 after 5 days of telithromycin therapy. The INR returned to the therapeutic range 4 days after telithromycin was discontinued. DISCUSSION: Telithromycin is the first member of the macrolide subclass of ketolides and offers potential advantages over traditional macrolides/azalides for community-acquired respiratory tract infections caused by macrolide-resistant pathogens. As of July 16, 2004, bleeding complications and an increased INR as a result of an interaction between warfarin and telithromycin have not been described. Although the mechanism for this interaction remains unknown, it is suspected that it is a result of the inhibition of the metabolism of the R-isomer of warfarin, which is metabolized predominantly by CYP1A2 and less by CYP3A4. Further research is required to elucidate this issue. An objective causality assessment revealed that this adverse drug event as a result of the warfarin and telithromycin interaction was probable. CONCLUSIONS: We recommend close monitoring of INR levels in patients on warfarin who receive telithromycin therapy in an effort to control and prevent bleeding complications. PMID- 15280512 TI - Multidrug-resistant Acinetobacter infections: an emerging challenge to clinicians. AB - OBJECTIVE: To review and evaluate clinically relevant epidemiology, microbiology, and clinical studies regarding the treatment of multidrug-resistant Acinetobacter infections. DATA SOURCES: Pertinent literature was identified by a MEDLINE search (1966-September 2003) and through secondary bibliographies of pertinent articles. STUDY SELECTION AND DATA EXTRACTION: All English-language articles identified from data sources were evaluated for clinical relevance. DATA SYNTHESIS: Acinetobacter baumannii has emerged as a worldwide problem as a nosocomial pathogen in hospitalized patients. Acinetobacter spp. can cause a multitude of infections including pneumonia, bacteremia, meningitis, urinary tract infections, and skin and soft tissue infections, and the mortality associated with these infections is high. Isolates resistant to almost all commercially available antimicrobials have been identified, thus limiting treatment options. The development of new agents and reappraisal of older compounds (ie, polymyxins, ampicillin/sulbactam) are necessary as we consider the optimal treatment of these multidrug-resistant organisms. CONCLUSIONS: There is no simple answer to the treatment of Acinetobacter infections. Eradication of Acinetobacter spp. requires adherence to good infection control practices and prudent antibiotic use, as well as effective antimicrobial therapy. Alternative therapies such as colistin, ampicillin/sulbactam, and tetracycline are potential options, but prospective, randomized, controlled trials are still lacking. PMID- 15280513 TI - CD40 ligand: a novel target in the fight against cardiovascular disease. AB - OBJECTIVE: To discuss the role of CD40 ligand (CD40L) in atherosclerosis and acute coronary syndromes (ACS), as well as describe relevant clinical literature evaluating the effects of pharmacotherapeutic agents on CD40L expression and soluble CD40L levels. DATA SOURCES: A MEDLINE and EMBASE search (1966-September 2003) was conducted using the key terms CD40, CD40 ligand, platelets, inflammation, and drug therapy. Additional primary literature was identified by reviewing the reference lists of relevant original and review papers. STUDY SELECTION AND DATA EXTRACTION: All articles identified in the search were evaluated, and those deemed relevant were incorporated into the review. DATA SYNTHESIS: CD40L is a transmembrane protein expressed on T cells, B cells, mast cells, basophils, eosinophils, natural killer cells, macrophages, endothelial cells, vascular smooth muscle cells, and activated platelets. It is also found in plasma as a soluble protein, sCD40L. As a consequence of CD40L binding to its receptor (CD40), several inflammatory processes are initiated. Studies have demonstrated elevated CD40L levels in patients with hypercholesterolemia and ACS, and elevated sCD40L levels have been associated with increased risk of cardiovascular events. Statins, glitazones, glycoprotein IIb/IIIa inhibitors, and clopidogrel have been demonstrated to effectively reduce CD40L levels both in vitro and in vivo. Abciximab has been shown to reduce the occurrence of death or myocardial infarction during 6 months of follow-up in patients with ACS who had the highest levels of sCD40L. CONCLUSIONS: The proinflammatory and procoagulant protein CD40L represents a novel target in the treatment of atherosclerosis and ACS. A number of therapeutic agents have been shown to modulate the expression of CD40L, findings that could have important clinical applications. PMID- 15280514 TI - Alfuzosin-induced acute hepatitis in a patient with chronic liver disease. AB - OBJECTIVE: To report a new case of probable alfuzosin-induced hepatitis. CASE SUMMARY: An 80-year-old man was evaluated because of jaundice and pruritus. He was diagnosed as having Child-Pugh A chronic liver disease due to hepatitis B virus. Other etiologies of hepatitis were appropriately ruled out, and the hepatitis B was non-replicative. Therefore, elevated liver enzyme levels were ascribed to alfuzosin treatment. DISCUSSION: Although alfuzosin-related mixed type hepatotoxicity has been previously reported, this is the first published case describing probable hepatocellular-type hepatotoxicity resulting from use of alfuzosin in a patient with underlying chronic liver disease. According to the Naranjo probability scale, alfuzosin was a probable cause of the hepatotoxicity. The mechanism of alfuzosin-induced liver damage is unknown. Several features, such as absence of predictable dose-dependent toxicity of alfuzosin in previous studies and absence of hypersensitivity manifestations in our case, are suggestive of a metabolic type of idiosyncratic toxicity. CONCLUSIONS: Alfuzosin rarely causes hepatotoxicity; however, clinicians must be alert for this adverse effect while using alfuzosin. PMID- 15280515 TI - Fatal venous thromboembolism associated with antipsychotic therapy. AB - OBJECTIVE: To describe the occurrence of pulmonary embolism (PE) as a rare adverse effect of clozapine that is treatable, but sometimes fatal, and survey the literature on the subject in the hope of increasing awareness of the potential danger that may result from drug interactions. CASE SUMMARY: A 47-year old woman treated with clozapine and paroxetine was admitted to the hospital with dyspnea and swelling of the leg. The patient was diagnosed as having PE and was treated with intravenous heparin. On hospital day 7, sudden acute respiratory failure developed and the patient died. Postmortem examination confirmed the existence of massive PE. DISCUSSION: The woman had no identifiable risk factors other than receiving a combination of clozapine and paroxetine, with a demonstrated elevated clozapine blood concentration. Use of the Naranjo probability scale revealed a probable likelihood that the adverse reaction was drug related. CONCLUSIONS: The association of antipsychotic drugs and venous thromboembolism has been previously described, but is still a rare finding. This case highlights the importance of monitoring and possibly discontinuing treatment when venous thrombosis is suspected. There should be careful monitoring, especially in patients with risk factors for thrombosis. Finally, antidepressant antipsychotic drug combinations can increase the risk of rare adverse effects, such as venous thromboembolism, even in the absence of other risk factors. PMID- 15280516 TI - Avoiding pregnancy: "A plan" versus plan B. PMID- 15280517 TI - Collagen type IV (alpha3-alpha4) nephropathy: from isolated haematuria to renal failure. PMID- 15280518 TI - Clostridium difficile diarrhoea in the immunosuppressed patient--update on prevention and management. PMID- 15280519 TI - 4-Heptanone is a metabolite of the plasticizer di(2-ethylhexyl) phthalate (DEHP) in haemodialysis patients. AB - BACKGROUND: There is an ongoing discussion about the risks of di(2-ethylhexyl) phthalate (DEHP) exposure for the general population as well as for specific subgroups in various medical settings. Haemodialysis patients certainly belong to the group with the highest exposure taking into account the repeated treatments over a long period of time. Many studies have shown that DEHP metabolites are more active with regard to cellular responses than DEHP itself. Although 4 heptanone has been shown to be a DEHP metabolite in rats, this has never been tested in humans. On the other hand, 4-heptanone was reported to be associated with diabetes mellitus. METHODS: After establishing analytical methods for all postulated metabolites, we analysed (i) plasma samples from 50 patients on haemodialysis and 50 controls; (ii) urine samples from 100 diabetic patients and 100 controls; and (iii) urine samples from 10 controls receiving DEHP intravenously. RESULTS: 4-Heptanone concentrations in urine did not differ between controls (128.6+/-11.4 micro g/l, mean+/- SEM) and diabetic patients (131.2+/-11.6 micro g/l) but were significantly elevated in plasma from haemodialysis patients (95.9+/-9.6 micro g/l) compared with controls (10.4+/-0.5 micro g/l). Exposure to DEHP led to a significant increase (P<0.001) of the metabolite 4-heptanone and all the proposed intermediates in urine of healthy persons within 24 h. CONCLUSIONS: These studies show that 4-heptanone is not associated with diabetes but is a major DEHP metabolite in humans. Studies concerning the toxicity of DEHP in haemodialysis patients and other highly exposed groups should therefore include 4-heptanone together with DEHP and its primary metabolites mono(2-ethylhexyl) phthalate (MEHP) and 2-ethylhexanol. PMID- 15280520 TI - Lack of clinical utility of urine myoglobin detection by microconcentrator ultrafiltration in the diagnosis of rhabdomyolysis. AB - BACKGROUND: In the diagnosis of rhabdomyolysis, the microconcentrator qualitative assay for urine myoglobin (uMb) is often used as a screening tool. The accuracy and clinical utility of this assay in screening patients with rhabdomyolysis have not been examined. METHODS: We conducted a retrospective analysis of the relationship between creatine kinase (CK), serum myoglobin (sMb), the urine qualitative assay for myoglobin and the semi-quantitative assay for urine haem pigments (uH) in patients evaluated for rhabdomyolysis. RESULTS: There were 673 patients with CK and uMb recorded on the same day. The uMb assay had a sensitivity of only 26.4% [95% confidence interval (CI): 23.1-29.7%] and specificity of 96.8% (95% CI: 95.5-98.1%) for the detection of severe rhabdomyolysis, defined as a CK >10 000 U/l. SMb and CK measured simultaneously in 83 patients were highly correlated (R(2) = 0.72 for log-transformed values), suggesting that the negative uMb test was not a result of the absence of sMb. In 241 patients who had CK, uMb and uH measured on the same day, the presence of 'moderate' or 'large' uH in the absence of haematuria, indicating presence of myoglobinuria, had a sensitivity of 81% (95% CI: 76-86%) for the detection of CK >10 000 U/l vs a sensitivity of 22% (95% CI: 17-27%) for the uMb assay. CONCLUSIONS: The microconcentrator-based uMb assay has a poor and clinically inadequate sensitivity in the detection and diagnosis of rhabdomyolysis. PMID- 15280521 TI - Steroid withdrawal after long-term medication for immunosuppressive therapy in renal transplant patients: adrenal response and clinical implications. AB - BACKGROUND: Withdrawal of steroids should be attempted after organ transplantation because of their adverse cardiovascular and metabolic effects. However, immunological, haemodynamic and symptomatic complications may occur due to the suppression of endogenous corticoid hormone synthesis under exogenous steroid intake. We have examined the effect of chronic steroid medication on adrenocortical function, and of steroid withdrawal, in immunologically stable renal transplant patients. METHODS: Sixty-three patients under long-term prednisone therapy (mean+/-SD 36+/-47 months) were assessed regarding basal fasting cortisol concentration and adrenocortical stimulation by the low-dose Synacten test both prior to and after stepwise prednisone withdrawal. Renal graft function (determined as the calculated glomerular filtration rate according to the Cockroft-Gault formula), mean arterial blood pressure and clinical status were evaluated concomitantly. RESULTS: Basal fasting cortisol concentration was clearly suppressed in 14% of patients under long-term steroid medication, and adrenocortical stimulation by the low-dose Synacten test was impaired in 31% after steroid withdrawal. About a third of all patients were symptomatic with fatigue (60%), arthralgias (60%), muscular weakness (20%), loss of appetite (20%), hypotension (15%) or headaches (5%). The incidence of symptoms was much higher in patients with low basal fasting cortisol concentration prior to steroid withdrawal, and after >2 years of steroid medication. Renal graft function, determined as glomerular filtration rate, decreased only slightly overall by approximately 5%, and was more pronounced in symptomatic vs asymptomatic patients (-7 vs -2 ml/min, respectively), as was the fall in mean arterial pressure (-10 vs -4.2 mmHg, respectively). CONCLUSIONS: Adrenal function is impaired in renal transplant patients receiving long-term steroid medication as part of their immunosuppressive regimen. This may lead to mainly symptomatic complications when steroids are withdrawn. The slight decrease in glomerular filtration rate probably can be ascribed mostly to the effect of steroids on systemic renal haemodynamics. It is recommended to consider cessation of steroid medication within 48 months of therapy, and after determination of basal cortisol to identify patients with potential complications. PMID- 15280522 TI - Midodrine appears to be safe and effective for dialysis-induced hypotension: a systematic review. AB - BACKGROUND: Dialysis-induced hypotension is an important complication of haemodialysis. Midodrine is an oral alpha-1 agonist that has been used in several small studies to prevent intradialytic hypotension (IDH). METHODS: The authors searched MEDLINE, EMBASE, ASN conference proceedings, and references of potentially relevant articles, and contacted industry (Shire Pharmaceuticals) for unpublished data. Observational studies, randomized controlled trials, crossover studies and pre- and post-intervention design studies with >/=5 haemodialysis patients were included. Study outcomes assessed were: hypotensive symptoms, changes in systolic and/or diastolic blood pressure, dry weight and length of stay after treatment. Data were abstracted on: study design, patient characteristics, intradialytic changes in blood pressure, nadir blood pressure and symptom improvement with midodrine. Thirty-seven full text articles were retrieved and nine met the selection criteria, in addition to one unpublished study. Midodrine dosing regimens ranged from 2.5 to 10 mg of midodrine given 15 30 min before dialysis. RESULTS: Post-dialysis systolic blood pressure was higher by 12.4 mmHg [95% confidence interval (CI) 7.5-17.7] and diastolic pressure was higher by 7.3 mmHg (95% CI 3.7-10.9) during midodrine treatment vs control. Likewise, the nadir systolic blood pressure was higher by 13.3 mmHg (95% CI 8.6 18.0), with a difference in nadir diastolic pressure of 5.9 mmHg (95% CI 2.7 9.1). Six of 10 studies report improvement in symptoms of IDH, and there were no reported serious adverse events ascribed to midodrine. CONCLUSIONS: This systematic review would suggest that midodrine has a role in the therapy of haemodialysis patients experiencing IDH. This conclusion must be viewed with caution, however, given the quality and sample size of the studies included in this review. PMID- 15280523 TI - Chlamydial infection and progression of carotid atherosclerosis in patients on regular haemodialysis. AB - BACKGROUND: Recent findings have suggested a possible contribution of chlamydial infection to the pathogenesis of atherosclerosis in the general population. However, the role that chlamydial antibody status plays in atherosclerosis generation in haemodialysis (HD) patients remains uncertain. METHODS: We measured carotid artery intima medial thickness (IMT) over 4 years in 100 HD subjects (age: 58+/-10 years; time on HD: 13+/-7 years; male/female: 67/33) and examined potential associations between Chlamydia pneumoniae (Cp) antibody seropositivity and changes in carotid artery IMT. RESULTS: During 4 years, carotid artery IMT increased significantly from 0.62+/-0.13 to 0.73+/-0.12 mm (P< 0.01). IMT progression was significantly and positively correlated with age (r = 0.37, P<0.01), log-transformed C-reactive protein (CRP; r = 0.33, P<0.01) and log transformed interleukin-6 (IL-6; r = 0.22, P<0.04), but inversely correlated with blood creatinine (r = -0.36, P<0.01) and albumin (r = -0.24, P<0.02). IMT increases were more prominent in patients positive for IgA antibodies (0.039+/- 0.022 mm/year, n = 52) compared with those without IgA antibodies (0.025+/-0.032 mm/year, n = 48) (P<0.01). IgA seropositivity did not accelerate IMT progression in patients with increased CRP (>0.11 mg/dl, n = 53), but significantly increased IMT to a greater extent in IgA-positive subjects than in IgA-negative subjects having lower CRP (/=6 weeks were assigned to high-flux polysulphone or high-flux modified cellulose with similar dialyser surface area and permeability characteristics and crossed over twice every 6 weeks. RESULTS: Thirty patients completed the study per protocol. Treatments with high-flux polysulphone and modified cellulose lowered serum triglyceride (by 20% and 10%, respectively; P<0.05) and remnant-like particle cholesterol by 32% (P<0.001) and 11% (NS) after the first 6 weeks of treatment. Oxidized LDL decreased significantly with high-flux polysulphone, but not with modified cellulose. Apolipoproteins CII and CIII were reduced, whereas the ratio CII/CIII was increased (all P<0.05). Acute-phase proteins and LDL and high density lipoprotein cholesterol remained unaffected. CONCLUSIONS: This randomized crossover study demonstrates a potent effect of high-flux haemodialysis on uraemic dyslipidaemia. Polysulphone membrane material showed superiority on oxidatively modified LDL, an indicator of oxidative stress in haemodialysis patients. PMID- 15280525 TI - Association of pelvic arterial calcification with arteriovenous thigh graft failure in haemodialysis patients. AB - BACKGROUND: Arterial calcification is a common problem in patients with chronic kidney disease, and has been associated with adverse clinical outcomes. The goal of the present study was to evaluate whether pelvic artery calcifications are associated with technical failure of arteriovenous thigh grafts in haemodialysis patients. METHODS: From 1 January 1999 to 30 June 2002, thigh grafts were placed in 54 haemodialysis patients who had exhausted all options for permanent vascular access in the upper extremities. Perioperative computed tomography (CT) of the abdomen and pelvis was obtained in 32 of the patients for diagnostic purposes unrelated to vascular access planning. Two radiologists, who were blinded to the graft outcomes, scored the vascular calcifications on CT of the distal aorta, common iliac, external iliac and common femoral arteries on a semi-quantitative 5 point scale. The association between technical graft failure (inability to complete the anastomosis) and the vascular calcification score was analysed. RESULTS: There was a high inter-observer agreement in scoring vascular calcification (kappa = 0.801). Among 26 patients with absent or mild pelvic arterial calcifications (grade 1-2) on CT, none (0%) experienced technical graft failure. In contrast, three of six patients (50%) with moderate to severe calcification (grade 3-5) had technical graft failures (P = 0.004 by Fisher's exact test). The cumulative 1 year graft patency was lower in the group with grade 3-5 calcification (33 vs 81%, P = 0.09). The two groups were similar in age, gender, race, diabetes, duration of dialysis, serum calcium, serum phosphorus and serum parathyroid hormone. CONCLUSION: There is a strong association between pelvic artery calcifications and technical failure of thigh grafts. The presence of moderate to severe vascular calcification is predictive of poor cumulative 1 year graft patency. PMID- 15280526 TI - Antidiuretic action of oxytocin is associated with increased urinary excretion of aquaporin-2. AB - BACKGROUND: The antidiuretic effect of oxytocin in humans is controversial. Urinary excretion of aquaporin-2 (AQP2) can be used as an index of the action of vasopressin on the kidney. We investigated whether exogenous oxytocin affects urinary concentration and urinary AQP2 excretion in human beings. METHODS: Oxytocin was administered intravenously at a rate of 20 mU/min in 10 healthy volunteers, seven patients with central diabetes insipidus (CDI) and three patients with nephrogenic diabetes insipidus (NDI). On the next day, 2 micro g of 1-desamino-8-d-arginine vasopressin (dDAVP) was injected subcutaneously. Two-hour urine was collected before and after the administration of oxytocin and dDAVP, and urinary AQP2 was measured semi-quantitatively by western analysis. RESULTS: Urine volume and free water clearance were decreased, and urine osmolality was increased by the administration of oxytocin or dDAVP in the normal volunteers and CDI patients. Urinary AQP2 excretion was increased by oxytocin infusion in the normal volunteers (from 34+/-12 to 326+/-120 densitometry unit (DU)/2 h) and in the CDI group (from 8+/-2 to 227+/-92 DU/2 h) (P<0.05), but not in the NDI group. dDAVP also had a similar but more potent effect on the urinary excretion of AQP2 in the normal and CDI groups. CONCLUSIONS: Oxytocin has an antidiuretic effect and increases the urinary excretion of AQP2 in humans whose urinary concentration mechanism is preserved. These results suggest that AQP2 might have a regulatory role in the antidiuretic action of oxytocin in humans. PMID- 15280527 TI - The renoprotective effects of structured care in a clinical trial setting in type 2 diabetic patients with nephropathy. AB - BACKGROUND: The RENAAL Study has confirmed the renoprotective effects of Losartan in type 2 diabetes. In this subgroup analysis from the RENAAL Study, we hypothesized that the intensive care received by patients in a clinical trial setting also reduced the rate of decline in renal function through optimization of all risk factors. METHODS: We compared the rate of deterioration in renal function, expressed as the regression coefficient of the monthly serum creatinine (SeCr) reciprocal (beta-1/Cr) in 55 Chinese type 2 diabetic patients before and after entry into the RENAAL Study. RESULTS: Of the 55 patients, 44 had at least three out-patient SeCr measurements both before (2.9+/-2.4 years) and after (3.3+/-0.8 years) entry into the study for evaluation. In the Losartan group (n = 24), the median beta-1/Cr fell from -11.4 x 10(-5) l micro mol(-1) month(-1) before entry into the trial to -4.7 x 10(-5) l micro mol(-1) month(-1) following entry (P = 0.001). The respective figures were -9.1 x 10(-5) and -5.0 x 10(-5) l micro mol(-1) month(-1) (P = 0.01) in the placebo group (n = 20). A decrease in beta-1/Cr was observed in 21 (87.5%) and 14 (70.0%) patients in the Losartan and placebo groups, respectively. Spot urinary albumin-to-creatinine ratio was reduced by 56% (P = 0.001) in the Losartan group but the change was not significant in the placebo group. At the end of the study, patients in both groups had lower blood pressure and better lipid control. The frequency of patient visits to doctors and nurses were doubled. CONCLUSIONS: The rate of renal function decline was significantly reduced in the majority of patients allocated to either Losartan or placebo following entry into the RENAAL study. These results suggest that in patients with diabetic nephropathy, implementation of a structured care protocol in a clinical trial setting facilities intensive treatment of risk factors confering renoprotective effects in addition to those resulting from Losartan treatment. PMID- 15280528 TI - Nephrotic focal segmental glomerulosclerosis in 2004: an update. PMID- 15280529 TI - Can blood flow surveillance and pre-emptive repair of subclinical stenosis prolong the useful life of arteriovenous fistulae? A randomized controlled study. AB - BACKGROUND: Stenosis is the main cause of arteriovenous fistula (AVF) failure. It is unclear, however, if surveillance for stenosis enhances AVF function and longevity and if there is an ideal time for intervention. METHODS: In a 5-year randomized, controlled, open trial we compared blood flow surveillance and pre emptive repair of subclinical stenoses (one or both of angioplasty and open surgery) with standard monitoring and intervention based upon clinical criteria alone to determine if the former prolonged the longevity of mature forearm AVFs. Surveillance with blood pump flow (Qb) monitoring during dialysis sessions and quarterly shunt blood flow (Qa) or recirculation measurements identified 79 AVFs with angiographically proven, significant (>50%) stenosis. The AVFs were randomized to either a control group (intervention done in response to a decline in the delivered dialysis dose or thrombosis; n = 36) or to a pre-emptive treatment group (n = 43). To evaluate a possible relationship between outcome and haemodynamic status of the access, AVFs were divided into functional and failing subgroups, according to Qa values higher or lower than 350 ml/min or the absence or presence of recirculation. RESULTS: A Kaplan-Meier analysis showed that pre emptive treatment reduced failure rate (P = 0.003) and the Cox hazards model identified treatment (P = 0.009) and higher baseline Qa (P = 0.001) as the only variables associated with favourable outcome. Primary patency rates were higher in treatment than in control AVFs in both functional (P = 0.021) and failing subgroups (P = 0.005). They were also higher in functional than in failing AVFs in both control (P<0.001) and treatment groups (P = 0.023). Access survival was significantly higher in pre-emptively treated than in control AVFs (P = 0.050), a higher post-intervention Qa being the only variable associated with improved access longevity (P = 0.044). Secondary patency rates were similar in pre emptively treated and control AVFs in both functional (P = 0.059) and failing subgroups (P = 0.394). They were also similar in functional and failing AVFs in controls (P = 0.082), but were higher in pre-emptively treated functional AVFs than in pre-emptively treated failing AVFs (P = 0.033) or in the entire control group (P = 0.019). CONCLUSIONS: We provide evidence that active blood flow surveillance and pre-emptive repair of subclinical stenosis reduce the thrombosis rate and prolong the functional life of mature forearm AVFs. We also show that Qa is a crucial indicator of access patency and a Qa >350 ml/min portends a superior outcome with pre-emptive action in AVFs. PMID- 15280530 TI - Gadolinium-based contrast media compared with iodinated media for digital subtraction angiography in azotaemic patients. AB - BACKGROUND: To determine whether gadolinium-based contrast media (CM) could be used safely for angiographies in patients with renal dysfunction we investigated renal function after gadobutrol exposure and compared the results with standard iodinated CM (iohexol) in a randomized clinical study. METHODS: Twenty-one patients (aged 67+/-11 years, nine female and 12 male) with severely impaired renal function [mean serum creatinine 3.2+/-1.3 mg/dl, mean glomerular filtration rate (GFR) 31+/-16 ml/min/1.73 m(2)] who needed to have angiography because of severe peripheral vascular disease, renal artery stenosis or aortic aneurysms were randomized to receive in a blinded manner either gadobutrol (Gadovist 1.0 mmol/ml) or iohexol (Omnipaque 350) as contrast agents. GFR was measured by CM clearance (Renalyzer) at baseline and 48 h after CM administration. The primary end point was the mean change of GFR from baseline at 48 h, the secondary one the incidence of CM-induced acute renal failure, defined as a decrease in GFR of >50% from baseline within 48 h of CM administration. RESULTS: In the gadobutrol group (n = 10) we observed a statistically significant decrease in GFR of 10.6+/-13.8 ml/min/1.73 m(2) within 48 h after CM administration (P<0.05, paired t test). The incidence of CM-induced ARF amounted to 50%. In comparison, the iohexol group (n = 11) also showed a statistically significant GFR reduction of 8.7+/-8.8 ml/min/1.73 m(2) (P<0.05, paired t test), and of ARF by 45%. The percentile of differences of GFR decreases between the two groups was not significant (P = 0.70). No patient demonstrated other adverse effects of gadobutrol or iohexol administration, apart from GFR reduction. Despite the decline in GFR, no patient required haemodialysis in the 10 following days. CONCLUSIONS: In our study, gadolinium-based angiography showed no benefit over iohexol angiography with respect to preventing GFR reduction in patients with severely impaired renal function. PMID- 15280531 TI - NF-kappaB activation and overexpression of regulated genes in human diabetic nephropathy. AB - BACKGROUND: Nuclear factor-kappaB (NF-kappaB) regulates genes involved in renal disease progression, such as the chemokines monocyte chemoattractant protein-1 (MCP-1) and RANTES. NF-kappaB is activated in experimental models of renal injury, and in vitro studies also suggest that proteinuria and angiotensin II could be important NF-kappaB activators. It has been proposed that locally produced MCP-1 may be involved in the development of diabetic nephropathy (DN). We examined the hypothesis that NF-kappaB could be an indicator of renal damage progression in DN. METHODS: Biopsy specimens from 11 patients with type 2 diabeties and overt nephropathy were studied by southwestern histochemistry for the in situ detection of activated NF-kappaB. In addition, by immunohistochemistry and/or in situ hybridization, we studied the expression of MCP-1 and RANTES, whose genes are regulated by NF-kappaB. RESULTS: NF-kappaB was detected mainly in cortical tubular epithelial cells and, to a lesser extent, in some glomerular and interstitial cells. A strong upregulation of MCP-1 and RANTES was observed in all the cases, mainly in tubular cells, and there was a strong correlation between the expression of these chemokines and NF-kappaB activation in the same cells, as observed in serial sections (r = 0.7; P = 0.01). In addition, the tubular expression of these chemokines was correlated mainly with the magnitude of the proteinuria (P = 0.002) and with interstitial cell infiltration (P<0.05). CONCLUSIONS: The activation of NF-kappaB and the transcription of certain pro-inflammatory chemokines in tubular epithelial cells are markers of progressive DN. Proteinuria might be one of the main factors inducing the observed pro-inflammatory phenotype. PMID- 15280532 TI - Total chemical synthesis of N-myristoylated HIV-1 matrix protein p17: structural and mechanistic implications of p17 myristoylation. AB - The HIV-1 matrix protein p17, excised proteolytically from the N terminus of the Gag polyprotein, forms a protective shell attached to the inner surface of the plasma membrane of the virus. During the late stages of the HIV-1 replication cycle, the N-terminally myristoylated p17 domain targets the Gag polyprotein to the host-cell membrane for particle assembly. In the early stages of HIV-1 replication, however, some p17 molecules dissociate from the viral membrane to direct the preintegration complex to the host-cell nucleus. These two opposing targeting functions of p17 require that the protein be capable of reversible membrane interaction. It is postulated that a significant structural change in p17 triggered by proteolytic cleavage of the Gag polyprotein sequesters the N terminal myristoyl group, resulting in a weaker membrane binding by the matrix protein than the Gag precursor. To test this "myristoyl switch" hypothesis, we obtained highly purified synthetic HIV-1 p17 of 131 amino acid residues and its N myristoylated form in large quantity. Both forms of p17 were characterized by circular dichroism spectroscopy, protein chemical denaturation, and analytical centrifugal sedimentation. Our results indicate that although N-myristoylation causes no spectroscopically discernible conformational change in p17, it stabilizes the protein by 1 kcal/mol and promotes protein trimerization in solution. These findings support the premise that the myristoyl switch in p17 is triggered not by a structural change associated with proteolysis, but rather by the destabilization of oligomeric structures of membrane-bound p17 in the absence of downstream Gag subdomains. PMID- 15280533 TI - Metastatic cancer DNA phenotype identified in normal tissues surrounding metastasizing prostate carcinomas. AB - Fourier transform-infrared statistical models have the proven ability to identify subtle structural changes in DNA at various stages of tumor development. Using these models, we show evidence for a metastatic cancer DNA phenotype in histologically normal prostate tissues surrounding metastasizing tumors. Strikingly, the DNA base and backbone structures of the metastatic phenotype are indistinguishable from those of the metastasizing prostate tumors but distinctly different from the structure recently reported for the primary cancer DNA phenotype. These findings suggest that the DNA structure linked to the development of metastasis is preordained in progenitor cells relatively early in multistep tumorigenesis. The substantial structural differences found between the primary and metastatic cancer DNA phenotypes suggest that each evolves through a separate pathway. The metastatic phenotype is potentially an early predictor of metastatic disease. Interventions that inhibit its formation would be expected to also inhibit the development of metastatic tumors. PMID- 15280534 TI - Air levels of carcinogenic polycyclic aromatic hydrocarbons after the World Trade Center disaster. AB - The catastrophic collapse of the World Trade Center (WTC) on September 11, 2001, created an immense dust cloud followed by fires that emitted soot into the air of New York City (NYC) well into December. The subsequent cleanup used diesel equipment that further polluted the air until the following June. The particulate air pollutants contained mutagenic and carcinogenic polycyclic aromatic hydrocarbons (PAHs). By using an assay developed for archived samples of fine particles, we measured nine PAHs in 243 samples collected at or near Ground Zero from September 23, 2001, to March 27, 2002. Based on temporal trends of individual PAH levels, we differentiated between fire and diesel sources and predicted PAH levels between 3 and 200 d after the disaster. Predicted PAH air concentrations on September 14, 2001, ranged from 1.3 to 15 ng/m(3); these values are among the highest reported from outdoor sources. We infer that these high initial air concentrations resulted from fires that rapidly diminished over 100 d. Diesel sources predominated for the next 100 d, during which time PAH levels declined slowly to background values. Because elevated PAH levels were transient, any elevation in cancer risk from PAH exposure should be very small among nonoccupationally exposed residents of NYC. However, the high initial levels of PAHs may be associated with reproductive effects observed in the offspring of women who were (or became) pregnant shortly after September 11, 2001. Because no PAH-specific air sampling was conducted, this work provides the only systematic measurements, to our knowledge, of ambient PAHs after the WTC disaster. PMID- 15280535 TI - Transplanted human fetal neural stem cells survive, migrate, and differentiate in ischemic rat cerebral cortex. AB - We characterize the survival, migration, and differentiation of human neurospheres derived from CNS stem cells transplanted into the ischemic cortex of rats 7 days after distal middle cerebral artery occlusion. Transplanted neurospheres survived robustly in naive and ischemic brains 4 wk posttransplant. Survival was influenced by proximity of the graft to the stroke lesion and was negatively correlated with the number of IB4-positive inflammatory cells. Targeted migration of the human cells was seen in ischemic animals, with many human cells migrating long distances ( approximately 1.2 mm) predominantly toward the lesion; in naive rats, cells migrated radially from the injection site in smaller number and over shorter distances (0.2 mm). The majority of migrating cells in ischemic rats had a neuronal phenotype. Migrating cells between the graft and the lesion expressed the neuroblast marker doublecortin, whereas human cells at the lesion border expressed the immature neuronal marker beta-tubulin, although a small percentage of cells at the lesion border also expressed glial fibrillary acid protein (GFAP). Thus, transplanted human CNS (hCNS)-derived neurospheres survived robustly in naive and ischemic brains, and the microenvironment influenced their migration and fate. PMID- 15280536 TI - The inhibitory effect of anandamide on luteinizing hormone-releasing hormone secretion is reversed by estrogen. AB - Because Delta-9-tetrahydrocannabinol (THC) inhibited luteinizing hormone releasing hormone (LHRH) in male rats, we hypothesized that the endocannabinoid, anandamide (AEA), would act similarly. AEA microinjected intracerebroventricularly (i.c.v.) decreased plasma luteinizing hormone (LH) at 30 min in comparison to values in controls (P < 0.001). The cannabinoid receptor 1 (CB1-r)-specific antagonist, [N-(piperidin-1-yl)-1-(2,4-dichlorophenyl)-5-(4 chlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide] (AM251), produced a significant elevation in plasma LH (P < 0.01). AEA (10(-9) M) decreased LHRH release from medial basal hypothalami incubated in vitro. These results support the concept that endogenous AEA inhibits LHRH followed by decreased LH release in male rats. In ovariectomized (OVX) female rats, AEA i.c.v. also inhibited LH release, but in this case AM251 had an even greater inhibitory effect than AEA. In vitro, AEA had no effect on LHRH in OVX rats. It seems that endogenous AEA inhibits LHRH followed by decreased LH release in OVX rats but that AM251 has an inhibitory action in this case. In striking contrast, in OVX, estrogen-primed (OVX-E) rats, AEA i.c.v. instead of decreasing LH, increased its release. This effect was completely blocked by previous injection of AM251. When medial basal hypothalami of OVX-E rats were incubated, AEA increased LHRH release. The synthesized AEA was higher in OVX-E rats than in OVX and males, indicating that estrogen modifies endocannabinoid levels and effects. The results are interpreted to mean that sex steroids have profound effects to modify the response to AEA. It inhibits LHRH and consequently diminishes LH release in males and OVX females, but stimulates LHRH followed by increased LH release in OVX-E-primed rats. PMID- 15280537 TI - Alternate, virus-induced membrane rearrangements support positive-strand RNA virus genome replication. AB - All positive-strand RNA [(+)RNA] viruses replicate their RNA on intracellular membranes, often in association with spherular invaginations of the target membrane. For brome mosaic virus, we previously showed that such spherules serve as compartments or mini-organelles for RNA replication and that their assembly, structure, and function have similarities to the replicative cores of retrovirus and double-stranded RNA virus virions. Some other (+)RNA viruses conduct RNA replication in association with individual or clustered double-membrane vesicles, appressed double membranes, or other structures whose possible relationships to the spherular invaginations are unclear. Here we show that modulating the relative levels and interactions of brome mosaic virus replication factors 1a and 2a polymerase (2apol) shifted the membrane rearrangements associated with RNA replication from small invaginated spherules to large, karmellae-like, multilayer stacks of appressed double membranes that supported RNA replication as efficiently as spherules. Spherules were induced by expressing 1a, which has functional similarities to retrovirus virion protein Gag, or 1a plus low levels of 2apol. Double-membrane layers were induced by 1a plus higher levels of 2apol and were suppressed by deleting the major 1a-interacting domain from 2apol. The stacked, double-membrane layers alternated with spaces that, like spherule interiors, were 50-60 nm wide, connected to the cytoplasm, and contained 1a and 2apol. These and other results suggest that seemingly diverse membrane rearrangements associated with RNA replication by varied (+)RNA viruses may represent topologically and functionally related structures formed by similar protein-protein and protein-membrane interactions and interconverted by altering the balances among those interactions. PMID- 15280538 TI - Translocation of CD28 to lipid rafts and costimulation of IL-2. AB - Stimulation of the CD28 costimulatory receptor can lead to an increased surface lipid raft expression in T lymphocytes. Here, we demonstrate that CD28 itself is recruited to lipid rafts in both Jurkat and peripheral blood T lymphocytes. This recruitment of CD28 is triggered by engagement with either anti-CD28 mAbs or a natural ligand of CD28, B7.2 (CD86). All detectable tyrosine-phosphorylated CD28 is in the lipid raft fractions, as is all of the CD28 associated with phosphatidylinositol 3-kinase, which is recruited to CD28 by tyrosine phosphorylation. Targeting the CD28 cytoplasmic domain to lipid rafts results in its tyrosine phosphorylation, indicating that tyrosine phosphorylation of CD28 may occur after translocation to lipid rafts. Studies with Jurkat cells deficient in Lck and CD45 demonstrate that movement of CD28 into lipid rafts does not require Lck and CD45 and can occur despite reduction of CD28 tyrosine phosphorylation to below the levels of detection. Analysis of murine CD28 mutants reveals a correlation between translocation to lipid rafts and costimulation of IL-2 production. Taken together with the known importance of lipid rafts in T cell activation, these observations suggest that translocation to lipid rafts may play an important role in CD28 signaling. PMID- 15280539 TI - TRIM5alpha mediates the postentry block to N-tropic murine leukemia viruses in human cells. AB - Murine leukemia viruses (MLVs) have been classified as N-tropic (N-MLV) or B tropic (B-MLV), depending on their ability to infect particular mouse strains. The early phase of N-MLV infection is blocked in the cells of several mammalian species, including humans. This block is mediated by a dominant host factor that targets the viral capsid soon after virus entry into the cell has been achieved. A similar block to HIV-1 in rhesus monkey cells is mediated by TRIM5alpha. Here we show that human TRIM5alpha is both necessary and sufficient for the restriction of N-MLV in human cells. Rhesus monkey TRIM5alpha, which potently blocks HIV-1 infection, exhibited only modest inhibition of N-MLV infection. B MLV was resistant to the antiviral effects of both human and rhesus monkey TRIM5alpha; susceptibility to TRIM5alpha-mediated restriction was conferred by alteration of residue 110 of the B-MLV capsid protein to the amino acid found in the N-MLV capsid. Our results demonstrate that species-specific variation in TRIM5alpha governs its ability to block infection by diverse retroviruses. PMID- 15280540 TI - Emigration of monocyte-derived cells from atherosclerotic lesions characterizes regressive, but not progressive, plaques. AB - Some monocytes normally take up residence in tissues as sessile macrophages, but others differentiate into migratory cells resembling dendritic cells that emigrate to lymph nodes. In an in vitro model of a vessel wall, lipid mediators lysophosphatidic acid and platelet-activating factor, whose signals are implicated in promoting atherosclerosis, blocked conversion of monocytes into migratory cells and favored their retention in the subendothelium. In vivo studies revealed trafficking of monocyte-derived cells from atherosclerotic plaques during lesion regression, but little emigration was detected from progressive plaques. Thus, progression of atherosclerotic plaques may result not only from robust monocyte recruitment into arterial walls but also from reduced emigration of these cells from lesions. PMID- 15280541 TI - N-acetylation of hypothalamic alpha-melanocyte-stimulating hormone and regulation by leptin. AB - The central melanocortin system is critical in the regulation of appetite and body weight, and leptin exerts its anorexigenic actions partly by increasing hypothalamic proopiomelanocortin (POMC) expression. The POMC-derived peptide alpha-melanocyte-stimulating hormone (alphaMSH) is a melanocortin 4 receptor agonist, and its potency in reducing energy intake is strongly increased by N acetylation. The reason for the higher biological activity of N-acetylated alphaMSH (Act-alphaMSH) compared with that of N-desacetylated alphaMSH (Des alphaMSH) is unclear, and regulation of acetylation by leptin has not been investigated. We show here that total hypothalamic alphaMSH levels are decreased in leptin-deficient ob/ob mice and increased in leptin-treated ob/ob and C57BL/6J mice. The increase in total alphaMSH occurred as soon as 3 h after leptin injection and was entirely due to an increase in Act-alphaMSH. Consistent with this observation, leptin rapidly induced the enzymatic activity of a N acetyltransferase in the hypothalamus of mice. In 293T cells expressing the melanocortin 4 receptor, Act-alphaMSH is far more potent than Des-alphaMSH in stimulating cAMP accumulation, an effect caused by a dramatically increased stability of Act-alphaMSH. Moreover, Des-alphaMSH is rapidly degraded in the hypothalamus after intracerebroventricular injection in rats and was less potent in inhibiting energy intake. The results suggest that leptin activates a N acetyltransferase in POMC neurons, leading to increased hypothalamic levels of Act-alphaMSH. Due to its increased stability, this posttranslational modification of alphaMSH may play a critical role in leptin action via the central melanocortin pathway. PMID- 15280542 TI - Distinct stoichiometry of BKCa channel tetramer phosphorylation specifies channel activation and inhibition by cAMP-dependent protein kinase. AB - Large conductance voltage- and calcium-activated potassium (BK(Ca)) channels are important signaling molecules that are regulated by multiple protein kinases and protein phosphatases at multiple sites. The pore-forming alpha-subunits, derived from a single gene that undergoes extensive alternative pre-mRNA splicing, assemble as tetramers. Although consensus phosphorylation sites have been identified within the C-terminal domain of alpha-subunits, it is not known whether phosphorylation of all or single alpha-subunits within the tetramer is required for functional regulation of the channel. Here, we have exploited a strategy to study single-ion channels in which both the alpha-subunit splice variant composition is defined and the number of consensus phosphorylation sites available within each tetramer is known. We have used this approach to demonstrate that cAMP-dependent protein kinase (PKA) phosphorylation of the conserved C-terminal PKA consensus site (S899) in all four alpha-subunits is required for channel activation. In contrast, inhibition of BK(Ca) channel activity requires phosphorylation of only a single alpha-subunit at a splice insert (STREX)-specific PKA consensus site (S4(STREX)). Thus, distinct modes of BK(Ca) channel regulation by PKA phosphorylation exist: an "all-or-nothing" rule for activation and a "single-subunit" rule for inhibition. This essentially digital regulation has important implications for the combinatorial and conditional regulation of BK(Ca) channels by reversible protein phosphorylation. PMID- 15280543 TI - A proteomic approach for the discovery of protease substrates. AB - Standardized, comprehensive platforms for the discovery of protease substrates have been extremely difficult to create. Screens for protease specificity are now frequently based on the cleavage patterns of peptide substrates, which contain small recognition motifs that are required for the cleavage of the scissile bond within an active site. However, these studies do not identify in vivo substrates, nor can they lead to the definition of the macromolecular features that account for the biological specificity of proteases. To use properly folded proteins in a proteomic screen for protease substrates, we used 2D difference gel electrophoresis and tandem MS to identify substrates of an apoptosis-inducing protease, granzyme B. We confirmed the cleavage of procaspase-3, one of the key substrates of this enzyme, and identified several substrates that were previously unknown, as well as the cleavage site for one of these substrates. We were also able to observe the kinetics of substrate cleavage and cleavage product accumulation by using the 2D difference gel electrophoresis methodology. "Protease proteomics" may therefore represent an important tool for the discovery of the native substrates of a variety of proteases. PMID- 15280544 TI - Efficient construction of the securine A carbon skeleton. AB - Securamine A is a structurally intriguing alkaloid possessing a pyrroloindole core joined via a modified isoprene subunit to a functionalized imidazole ring. Recent synthetic efforts in this laboratory have resulted in the efficient construction of key lactone 36, which undergoes tandem azide reduction/ring expansion to macrolactam 37. Macrolactam 37 possesses the complete macrocyclic core of securamine A. PMID- 15280545 TI - Glucocorticoid receptor overexpression in forebrain: a mouse model of increased emotional lability. AB - The molecular mechanisms that control the range and stability of emotions are unknown, yet this knowledge is critical for understanding mood disorders, especially bipolar illness. Here, we show that the glucocorticoid receptor (GR) modulates these features of emotional responsiveness. We generated transgenic mice overexpressing GR specifically in forebrain. These mice display a significant increase in anxiety-like and depressant-like behaviors relative to wild type. Yet, they are also supersensitive to antidepressants and show enhanced sensitization to cocaine. Thus, mice overexpressing GR in forebrain have a consistently wider than normal range of reactivity in both positive and negative emotionality tests. This phenotype is associated, in specific brain regions, with increased expression of genes relevant to emotionality: corticotropin-releasing hormone, serotonin, norepinephrine and dopamine transporters, and 5 hydroxytryptamine(1A) receptor. Thus, GR overexpression in forebrain causes higher "emotional lability" secondary to a unique pattern of molecular regulation. This finding suggests that natural variations in GR gene expression can contribute to the fine-tuning of emotional stability or lability and may play a role in bipolar disorder. PMID- 15280546 TI - Prospects for total synthesis: a vision for a totally synthetic vaccine targeting epithelial tumors. AB - Vaccines derived from totally synthetic carbohydrate antigens have been shown to elicit an immune response in both preclinical and clinical settings. The vaccines have been proven safe when administered in human clinical trials and are also competent at inducing antibodies that react with aberrant cells expressing the corresponding carbohydrate antigen. The most well studied vaccines have hitherto focused on single carbohydrate antigens, notwithstanding the known heterogeneity of transformed cells. Advances in synthetic organic chemistry have enabled the preparation and subsequent investigation of vaccines that contain several different tumor-associated carbohydrate antigens in a single molecule. These unimolecular constructs could, in principle, serve as superior mimics of cell surface antigens and hence, as multifaceted cancer vaccines. We report here the synthesis of a pentameric vaccine targeting a specific cancer. The new vaccine contains prostate tumor-associated antigens, Tn, TF, STn, Lewis(y), and Globo-H. To reach our goal, antigen-containing amino acid monomers were assembled in a linear fashion to form a glycopeptide containing the five distinct carbohydrate antigen units. The attachment of a linker to the glycopeptide followed by an extraordinary global deprotection and subsequent conjugation to two different immunogenic carriers, keyhole limpet hemocyanin and N-alpha-palmitoyl-S-[2,3 bis(palmitoyloxy)-(2RS)-propyl]-L-cysteine, resulted in the vaccine constructs. The results described herein indicate that complex unimolecular multivalent vaccines can be efficiently produced in the laboratory. These fully synthetic vaccines have the potential to stimulate a multifaceted immune response against prostate cancer. PMID- 15280547 TI - Twisting macromolecular chains: self-assembly of a chiral supermolecule from nonchiral polythiophene polyanions and random-coil synthetic peptides. AB - The self-assembly of a negatively charged conjugated polythiophene derivative and a positively charged synthetic peptide will create a chiral, well ordered supermolecule. This supermolecule has the three-dimensional ordered structure of a biomolecule and the electronic properties of a conjugated polymer. The molecular complex being formed clearly affects the conformation of the polymer backbone. A main-chain chirality, such as a predominantly one-handed helical structure induced by the acid-base complexation between the conjugated polymer and the synthetic peptide, is seen. The alteration of the polymer backbone influences the optical properties of the polymer, seen as changes in the absorption, emission, and Raman spectra of the polymer. The complexation of the polythiophene and the synthetic peptide also induce a change from random-coil to helical structure of the synthetic peptide. The supermolecule described in this article may be used in a wide range of applications such as biomolecular devices, artificial enzymes, and biosensors. PMID- 15280548 TI - Pauling and Corey's alpha-pleated sheet structure may define the prefibrillar amyloidogenic intermediate in amyloid disease. AB - Transthyretin, beta(2)-microglobulin, lysozyme, and the prion protein are four of the best-characterized proteins implicated in amyloid disease. Upon partial acid denaturation, these proteins undergo conformational change into an amyloidogenic intermediate that can self-assemble into amyloid fibrils. Many experiments have shown that pH-mediated changes in structure are required for the formation of the amyloidogeneic intermediate, but it has proved impossible to characterize these conformational changes at high resolution using experimental means. To probe these conformational changes at atomic resolution, we have performed molecular dynamics simulations of these proteins at neutral and low pH. In low-pH simulations of all four proteins, we observe the formation of alpha-pleated sheet secondary structure, which was first proposed by L. Pauling and R. B. Corey [(1951) Proc. Natl. Acad. Sci. USA 37, 251-256]. In all beta-sheet proteins, transthyretin and beta(2)-microglobulin, alpha-pleated sheet structure formed over the strands that are highly protected in hydrogen-exchange experiments probing amyloidogenic conditions. In lysozyme and the prion protein, alpha-sheets formed in the specific regions of the protein implicated in the amyloidogenic conversion. We propose that the formation of alpha-pleated sheet structure may be a common conformational transition in amyloidosis. PMID- 15280550 TI - Persistent tetrodotoxin-sensitive sodium current resulting from U-to-C RNA editing of an insect sodium channel. AB - The persistent tetrodotoxin (TTX)-sensitive sodium current, detected in neurons of many regions of mammalian brains, is associated with many essential neuronal activities, including boosting of excitatory synaptic inputs, acceleration of firing rates, and promotion of oscillatory neuronal activities. However, the origin and molecular basis of the persistent current have remained controversial for decades. Here, we provide direct evidence that U-to-C RNA editing of an insect sodium channel transcript generates a sodium channel with a persistent current. We detected a persistent TTX-sensitive current in a splice variant of the cockroach sodium channel gene BgNa(v) (formerly para(CSMA)). Site-directed mutagenesis experiments revealed that an F-to-S change at the C-terminal domain of this variant was responsible for the persistent current. We demonstrated that this F-to-S change was the result of a U-to-C RNA editing event, which also occurred in the Drosophila para sodium channel transcript. Our work provides direct support for the hypothesis that posttranscriptional modification of a conventional transient sodium channel produces a persistent TTX-sensitive sodium channel. PMID- 15280549 TI - Rad6-Bre1-mediated histone H2B ubiquitylation modulates the formation of double strand breaks during meiosis. AB - An E2 ubiquitin-conjugating enzyme, Rad6, working with an E3 ubiquitin ligase Bre1, catalyzes monoubiquitylation of histone H2B on a C-terminal lysine residue. The rad6 mutant of Saccharomyces cerevisiae shows a meiotic prophase arrest. Here, we analyzed meiotic defects of a rad6 null mutant of budding yeast. The rad6 mutant exhibits pleiotropic phenotypes during meiosis. RAD6 is required for efficient formation of double-strand breaks (DSBs) at meiotic recombination hotspots, which is catalyzed by Spo11. The mutation decreases overall frequencies of DSBs in a cell. The effect of the rad6 mutation is local along chromosomes; levels of DSBs at stronger hotspots are particularly reduced in the mutant. The absence of RAD6 has little effect on the formation of ectopic DSBs targeted by Spo11 fusion protein with a Gal4 DNA-binding domain. Furthermore, the disruption of the BRE1 as well as substitution of the ubiquitylation site of histone H2B also reduces some DSB formation similar to the rad6. These results suggest that Rad6-Bre1, through ubiquitylation of histone H2B, is necessary for efficient recruitment and/or stabilization of a DSB-forming machinery containing Spo11. Histone tail modifications might play a role in DSB formation during meiosis. PMID- 15280551 TI - In vivo identification of genes that modify ether-a-go-go-related gene activity in Caenorhabditis elegans may also affect human cardiac arrhythmia. AB - Human ether-a-go-go-related gene (HERG) encodes the pore-forming subunit of I(Kr), a cardiac K(+) channel. Although many commonly used drugs block I(Kr), in certain individuals, this action evokes a paradoxical life-threatening cardiac rhythm disturbance, known as the acquired long QT syndrome (aLQTS). Although aLQTS has become the leading cause of drug withdrawal by the U.S. Food and Drug Administration, DNA sequencing in aLQTS patients has revealed HERG mutations only in rare cases, suggesting that unknown HERG modulators are often responsible. By using the worm Caenorhabditis elegans, we have developed in vivo behavioral assays that identify candidate modulators of unc-103, the worm HERG orthologue. By using RNA-interference methods, we have shown that worm homologues of two HERG interacting proteins, Hyperkinetic and K channel regulator 1 (KCR1), modify unc 103 function. Examination of the human KCR1 sequence in patients with drug induced cardiac repolarization defects revealed a sequence variation (the substitution of isoleucine 447 by valine, I447V) that occurs at a reduced frequency (1.1%) relative to a matched control population (7.0%), suggesting that I447V may be an allele for reduced aLQTS susceptibility. This clinical result is supported by in vitro studies of HERG dofetilide sensitivity by using coexpression of HERG with wild-type and I447V KCR1 cDNAs. Our studies demonstrate the feasibility of using C. elegans to assay and potentially identify aLQTS candidate genes. PMID- 15280552 TI - Reproductive tissue selective actions of progesterone receptors. AB - The steroid hormone, progesterone, plays a central coordinate role in diverse events associated with female reproduction. In humans and other vertebrates, the biological activity of progesterone is mediated by modulation of the transcriptional activity of two progesterone receptors, PR-A and PR-B. These receptors arise from the same gene and exhibit both overlapping and distinct transcriptional activities in vitro. To delineate the individual roles of PR-A and PR-B in vivo, we have generated mouse models in which expression of a single PR isoform has been ablated. Analysis of the reproductive phenotypes of these mice has indicated that PR-A and PR-B mediate mostly distinct but partially overlapping reproductive responses to progesterone. While selective ablation of the PR-A protein (PR-A knockout mice, PRAKO mice) shows normal mammary gland response to progesterone but severe uterine hyperplasia and ovarian abnormalities, ablation of PR-B protein (PRBKO mice) does not affect biological responses of the ovary or uterus to progesterone but results in reduced pregnancy associated mammary gland morphogenesis. The distinct tissue-specific reproductive responses to progesterone exhibited by these isoforms are due to regulation of distinct subsets of progesterone-dependent target genes by the individual PR isoforms. This review will summarize our current understanding of the selective contribution of PR isoforms to the cellular and molecular actions of progesterone in reproductive tissues. PMID- 15280553 TI - In vivo and in vitro differentiation of male germ cells in the mouse. AB - Primordial germ cells appear in the embryo at about day 7 after coitum. They proliferate and migrate towards the genital ridge. Once there, they undergo differentiation into germ stem cells, known as 'A spermatogonia'. These cells are the foundation of spermatogenesis. A spermatogonia commit to spermatogenesis, stay undifferentiated or degenerate. The differentiation of primordial germ cells to migratory, postmigratory and germ stem cells is dependent on gene expression and cellular interactions. Some of the genes that play a crucial role in germ cell differentiation are Steel, c-Kit, VASA, DAZL, fragilis, miwi, mili, mil1 and mil2. Their expression is stage specific, therefore allowing solid identification of germ cells at different developmental phases. In addition to the expression of these genes, other markers associated with germ cell development are nonspecific alkaline phosphatase activity, the stage specific embryonic antigen, the transcription factor Oct3/4 and beta1- and alpha6-integrins. Commitment of cells to primordial germ cells and to A spermatogonia is also dependent on induction by the bone morphogenetic protein (BMP)-4. With this knowledge, researchers were able to isolate germ stem cells from embryonic stem cell-derived embryoid bodies, and drive these into gametes either in vivo or in vitro. Although no viable embryos were obtained from these gametes, the prospects are that this goal is not too far from being accomplished. PMID- 15280554 TI - The dynamics of cyclin B1 distribution during meiosis I in mouse oocytes. AB - Cdk1-cyclin B1 kinase activity drives oocytes through meiotic maturation. It is regulated by the phosphorylation status of cdk1 and by its spatial organisation. Here we used a cyclin B1-green fluorescent protein (GFP) fusion protein to examine the dynamics of cdk1-cyclin B1 distribution during meiosis I (MI) in living mouse oocytes. Microinjection of cyclin B1-GFP accelerated germinal vesicle breakdown (GVBD) and, as previously described, overrides cAMP-mediated meiotic arrest. GVBD was pre-empted by a translocation of cyclin B1-GFP from the cytoplasm to the germinal vesicle (GV). After nuclear accumulation, cyclin B1-GFP localised to the chromatin. The localisation of cyclin B1-GFP is governed by nuclear import and export. In GV intact oocytes, cyclin export was demonstrated by showing that cyclin B1-GFP injected into the GV is exported to the cytoplasm while a similar size dextran is retained. Import was revealed by the finding that cyclin B1-GFP accumulated in the GV when export was inhibited using leptomycin B. These studies show that GVBD in mouse oocytes is sensitive to cyclin B1 abundance and that the changes in distribution of cyclin B1 contribute to progression through MI. PMID- 15280555 TI - Spermatogenesis following syngeneic testicular transplantation in Balb/c mice. AB - Transplantation of spermatogonial stem cells in cross-species has been widely used to study the function of Sertoli cells and the effect of phylogenetic distance between donor and recipient animals on the outcome of spermatogonial transplantation, whereas there have been only a few reports on the transplantation of testis tissue. The objective of the present study was to examine the development of grafted testes and the kinetics of spermatogenesis following syngeneic testicular transplantation in both male and female recipient Balb/c mice in an effort to establish an in vivo culture system and to compare the effects of host sex on spermatogenesis. The testes from 5-day-old Balb/c mice were transplanted under the dorsal skin of four-week-old mice. Twenty male and twenty female Balb/c mice were used as the hosts and each host received 4 grafts. The recipient mice were killed at 1, 2, 3, 5, 7, 9, 12 and 15 weeks after transplantation. The graft survival rate and graft size were measured. The status of spermatogenesis was assessed by histological analyses. The expression of the spermatid-specific Protamine-2 gene was examined by RT-PCR. Overall, 70.3% of the testicular grafts in male hosts and 67.2% in female hosts survived. All recovered grafts had increased in volume, some of them had increased by more than 30-fold. The architecture of the seminiferous tubules in female hosts appeared to be better than that in male hosts. The round spermatids were the most advanced germ cells until 15 weeks after transplantation, and no complete spermatozoon was observed in any of the grafts. The expression of protamine-2 was detected in grafts from 5 weeks posttransplantation in both male and female hosts, confirming that the spermatogenic cells differentiated into spermatids. In contrast to grafts, the testes of male hosts had a normal histological appearance. The results showed the schedule of spermatogenesis following syngeneic testicular transplantation in both male and female hosts. This model could be useful for further studies involving the endocrinology of the testis and the mechanisms of spermatogenesis. PMID- 15280556 TI - Induced hyperactivity in boar spermatozoa and its evaluation by computer-assisted sperm analysis. AB - Hyperactivity, a form of sperm motility characterized by vigorous flagellar movements, has been proposed as essential for fertilization in mammals. The objective of the present study was to establish a method for inducing hyperactivity in vitro in boar spermatozoa and to define threshold values to differentiate between hyperactive and non-hyperactive spermatozoa by computer assisted sperm analysis (CASA) as a prerequisite for analyzing the energy metabolism during hyperactivity. In TALP-HEPES medium, non-frozen boar spermatozoa were stimulated to hyperactivity by 50 micromol l(-1) Ca2+ within 15 min at 37 degrees C if 5 micromol l(-1) of the Ca2+ ionophore A23187 was present. If 25% seminal plasma was present, boar spermatozoa required higher Ca2+ concentrations (about 700 micromol l(-1)) for hyperactivity. Under both conditions, immobilization and head-to-head agglutination were low so that hyperactive spermatozoa could be analyzed for at least 40 min. The transition from normal to hyperactive movement was characterized by an increase in flagellar beat angle from 49 degrees +/- 12 degrees to 200 degrees +/- 36 degrees (n = 32) and a decrease in flagellar curvature ratio from 0.89 +/- 0.04 to 0.47 +/- 0.11 (n = 32). For quantification of hyperactive boar sperm, kinematic parameters of hyperactive and non-hyperactive spermatozoa were measured by CASA and statistically evaluated (receiver operating characteristic (ROC) curve analysis). The threshold values of the following four parameters were well suited for differentiating between hyperactive and non-hyperactive boar spermatozoa (ROC curve analysis: > 50% specificity at 100% sensitivity). Hyperactive boar spermatozoa showed mean lateral head displacement > 3.5 microm, curvilinear velocity > 97 microm s(-1), linearity < 32% and wobble < 71%. According to this multiparametric definition, induction of hyperactivity increased significantly (P < 0.0001) the fraction of hyperactive spermatozoa in semen samples from 5.1 +/- 4.3% (n = 13) to 48.3 +/- 6.6% (n = 7) in the absence and to 44.2 +/- 7.6% (n = 10) in the presence of 25% seminal plasma, while the overall percentage of motile spermatozoa did not change significantly. PMID- 15280557 TI - Serotonin antagonist-induced lowering of prolactin secretion does not affect the pattern of pulsatile secretion of follicle-stimulating hormone and luteinizing hormone in the bitch. AB - Dopamine agonists decrease plasma prolactin concentration and shorten the duration of anoestrus in the bitch. In order to determine whether this shortening results from decreased prolactin release or is due to another dopamine agonistic effect on the pulsatile release of FSH and LH, eight anoestrous beagle bitches were treated with a low dose of the serotonin antagonist metergoline (0.1 mg per kg body weight, twice daily) starting 100 days after ovulation. Six-hour plasma profiles of LH and FSH were obtained 7 days before, immediately before, 1 week after, and then at 2-week intervals after the start of the treatment with the serotonin antagonist until signs of pro-oestrus appeared. Plasma prolactin concentration was measured three times weekly from 75 to 142 days after ovulation and thereafter once weekly until the next ovulation, and was observed to decrease significantly after the start of treatment. The length of the interoestrous interval in the treated dogs was, however, not different from that in the preceding pretreatment cycle or from that in a group of untreated bitches. During the first weeks of treatment no changes were observed in the pulsatile plasma profiles of FSH and LH. Four weeks after the start of the treatment with the serotonin antagonist there was an increase in the mean basal plasma FSH concentration and the mean area under the curve for FSH, without a concurrent increase in LH secretion. The increase in FSH secretion continued until late anoestrus. In conclusion, the serotonin antagonist-induced lowering of plasma prolactin concentration was not associated with shortening of the interoestrous interval. The plasma profiles of LH and FSH were similar to those observed during physiological anoestrus, but different from those observed during anoestrus shortened by treatment with a dopamine agonist. Hence the prematurely induced oestrus observed during administration of dopamine agonists cannot be explained by a decreased plasma prolactin concentration but must be due to some other dopamine agonistic effect, probably increased FSH secretion. The observations in this study further strengthen the hypothesis that an increase in circulating FSH is essential for ovarian folliculogenesis and consequently the termination of anoestrus in the bitch. PMID- 15280558 TI - Real-time dynamics of prostaglandin F2alpha release from uterus and corpus luteum during spontaneous luteolysis in the cow. AB - Prostaglandin (PG) F2alpha released from the uterus in a pulsatile fashion is essential to induce regression of the corpus luteum (CL) in the cow. In addition to the uterus, the CL has also been recognized as a site of PGF(2alpha) production. Therefore, this study aimed to determine the detailed dynamics of the releasing profile of CL-derived PGF2alpha together with uterus-derived PGF2alpha during spontaneous luteolysis in the cow. Non-lactating Holstein cows (n = 6) were surgically implanted with a microdialysis system (MDS) on day 15 (oestrus = day 0) of the oestrous cycle. Simultaneously, catheters were implanted to collect ovarian venous plasma ipsilateral to the CL as well as jugular venous plasma. The concentrations of PGF2alpha, 13,14-dihydro-15-keto-PGF2alpha (PGFM) and progesterone in the MDS and plasma samples were determined by enzyme immunoassays. The intra-luteal PGF2alpha secretion slightly increased after the onset of luteolysis (0 h) and drastically increased from 24 h, and was maintained at high levels towards the following oestrus. Furthermore, PGF2alpha was released from the CL into the ovarian vein in a pulsatile manner during spontaneous luteolysis. Also, the fact that intra-luteal secretion of PGF2alpha and PGFM showed a positive correlation indicates the existence of a local metabolic pathway for PGF2alpha in the CL. In conclusion, the present study clarified the real-time dynamics of uterus-derived PGF2alpha and CL-derived PGF2alpha during spontaneous luteolysis in the cow, and gives the first in vivo evidence that the CL releases PGF2alpha during spontaneous luteolysis in the cow. Although the physiological relevance of CL-derived PGF2alpha appears to be restricted to a local role as an autocrine/paracrine factor in the CL, overall results support the concept that the local release of PGF2alpha within the regressing CL amplifies the luteolytic action of PGF2alpha from the uterus. PMID- 15280559 TI - Signal transducer and activator of transcription 3 (Stat3) expression and activation in rat uterus during early pregnancy. AB - Signal transducer and activator of transcription 3 (Stat3), a member of the Stat family, is specifically activated during mouse embryo implantation. The aim of this study was to investigate the expression, activation and regulation of Stat3 in rat uterus during early pregnancy, pseudopregnancy, delayed implantation and artificial decidualization. Stat3 mRNA was highly expressed in the luminal epithelium on day 5 and in the luminal epithelium and underlying stromal cells at implantation sites on day 6 of pregnancy. There was a strong level of Stat3 protein expression and phosphorylation in the stromal cells near the lumen and in the luminal epithelium on day 5 of pregnancy, which was similar to day 5 of pseudopregnancy. In the afternoon of day 6, the strong level of Stat3 phosphorylation was detected only in the luminal epithelium. Stat3 was highly expressed and activated in the decidual cells from days 7 to 9 of pregnancy and under artificial decidualization in the present study. Our results suggest that the strong level of Stat3 activation in the luminal epithelium and underlying stromal cells during the pre-implantation period may be important for establishing uterine receptivity as in mice, and the high level of Stat3 expression and activation in decidual cells may play a role during decidualization. PMID- 15280560 TI - Phagocytosis as a potential mechanism for microbial defense of mouse placental trophoblast cells. AB - Trophoblast giant cells are active phagocytes during implantation and post implantation. Phagocytosis decreases during placental maturation as the phagocytic function of nutrition is gradually replaced by the direct uptake of nutrients by the labyrinth zone trophoblast. We hypothesize that, after placental maturation, trophoblast cells maintain phagocytic functions for purposes other than nutrition. This study employs histological techniques to examine the ability of trophoblast cells to phagocytose microorganisms (yeast or bacteria)--in vivo in females receiving thioglycolate to activate macrophages and in vitro in the presence of phagocytic promoters such as interferon-gamma and complement component C3. Placental trophoblast cells from the second half of gestation show basal phagocytosis that can be dramatically up-regulated by these promoters when microorganisms are inoculated into pregnant animals or introduced into culture systems. Stimulated trophoblast cells phagocytosed organisms more rapidly and in greater numbers than non-stimulated trophoblast exposed to the same numbers of organisms. Taken together, our results indicate that trophoblast cells do not lose their ability to phagocytose during the placentation process, which may imply that trophoblast cells participate in embryonic and fetal innate immune defense through elimination of microorganisms present at the maternal-fetal interface. PMID- 15280561 TI - Expression of the full-length and alternatively spliced equine luteinizing hormone/chorionic gonadotropin receptor mRNAs in the primary corpus luteum and fetal gonads during pregnancy. AB - The full-length equine luteinizing hormone/chorionic gonadotropin (LH/CG) receptor (eLH/CG-RA) cDNA and two alternatively spliced isoforms (eLH/CG-RB,C) were isolated from luteal tissue and characterized using a combination of reverse transcription-polymerase chain reaction (RT-PCR) and 5'-rapid amplification of cDNA ends. The 680-amino acid full sequence of eLH/CG-RA displayed 87-92% homology with other mammalian LH/CG-Rs. The eLH/CG-RB and eLH/CG-RC cDNA isoforms were truncated from the 3'-end of exon X: eLH/CG-RB spliced out of frame into the last exon whereas eLH/CG-RC contained an in-frame stop codon within a divergent sequence. Consequently, both eLH/CG-RB and eLH/CG-RC cDNA isoforms encoded putative proteins without transmembrane and intracellular domains. In order to study the responsiveness of the primary corpus luteum (CL) and fetal gonads to eCG, the expression of eLH/CG-R mRNAs was examined by RT-PCR and Northern blot analysis during early and mid-pregnancy. All three eLH/CG-R cDNA isoforms (eLH/CG RA,B,C) were expressed from day 14 to day 83 of pregnancy in the primary CL and from day 44 to day 222 in fetal gonads. Interestingly, the primary CL at days 89 and 151 expressed only truncated eLH/CG-R cDNA isoforms. The relative values of Northern hybridized major 7, 5.7, 3.9 and 1.8 kb eLH/CG-R mRNA transcripts tended to decrease in the primary CL whereas the unique major 1.8 kb eLH/CG-R mRNA was steadily expressed in fetal gonads during pregnancy. These results show that the expression of eLH/CG-R mRNAs occurs in the fetal gonads before ceasing in the primary CL and suggest that eCG may be involved in the gradual transition from a luteal to a feto-placental output of steroids during equine pregnancy. PMID- 15280562 TI - Placental transport of leucine in a porcine model of low birth weight. AB - Low birth weight is a major factor in neonatal morbidity and mortality in humans and domestic species and is a predictor of physiological disorders in adulthood. This study utilised the naturally occurring variation in pig fetal size within a uterus to test the hypothesis that placental amino acid transport capability is associated with fetal growth. Leucine uptake by trophoblast vesicles prepared from placentas supplying an average-sized fetus and the smallest fetus in the uterus was assessed. On days 45 and 65 of gestation, uptake of leucine by the porcine placenta was predominantly sodium independent and was inhibited by the non-metabolised leucine analogue 2-amino-2-norbornane-carboxylic acid, indicating that uptake occurs via system L. By day 100 the uptake of leucine by placentas supplying average-sized fetuses had changed from being predominantly sodium independent to involving both sodium-dependent (system B0) and -independent (system L) pathways. This change was not seen in placentas supplying the smallest fetus, which continued to display predominantly sodium-independent uptake. In conclusion, these data show gestational- and fetal size-dependent changes in the transport of leucine across the porcine placenta. PMID- 15280563 TI - Seminal plasma regulates endometrial cytokine expression, leukocyte recruitment and embryo development in the pig. AB - In pigs, uterine exposure to the constituents of semen is known to increase litter size but the underlying physiological mechanisms remain undefined. Studies in rodents and humans implicate immune modulating moieties in seminal plasma as likely candidates, acting through enhancing the receptivity of the female tract. In this study, the acute and longer term effects of seminal plasma on cytokine expression and leukocyte abundance in the pig endometrium during early pregnancy have been characterised. The reproductive tracts of gonadotrophin-primed pre pubertal gilts treated with intrauterine infusions of either pooled seminal plasma or phosphate-buffered saline (PBS) were retrieved at 34 h, or on day 5 and day 9 after treatment. Seminal plasma elicited an endometrial inflammatory infiltrate comprised of predominantly macrophages and major histocompatibility complex class II+-activated macrophages and dendritic cells. The abundance of these cells was greatest at the pre-ovulatory (34 h) time-point and their increase relative to PBS-treated tissues was maintained until day 9 after seminal plasma treatment. Seminal plasma induced the expression of the cytokines, granulocyte macrophage colony-stimulating factor, interleukin-6 and monocyte chemoattractant protein-1, and the eicosanoid-synthesising enzyme cyclo-oxygenase 2. Expression was maximal 34 h after treatment but altered expression patterns as a consequence of seminal plasma induction persisted through early pregnancy. These changes were accompanied by altered dynamics in pre-implantation embryo development with an increase in the number of embryos and in their viability after seminal plasma treatment. Together, these findings implicate factors in seminal plasma in programming the trajectory of uterine cytokine expression and leukocyte trafficking during early pregnancy and in regulating pre-implantation embryo development in the pig. PMID- 15280564 TI - Production and localization of activins and activin type IIA and IIB receptors by the human endosalpinx. AB - Fallopian tubes from ten premenopausal women were collected and examined for the presence of inhibin, activin and its type IIA and IIB receptors (ActRIIA and ActRIIB) in the endosalpinx. Immunocytochemistry demonstrated clear staining for the betaA, betaB subunits and ActRIIA and ActRIIB that increased in intensity from the isthmus to the ampulla. No staining for the alpha subunit was observed. Whilst the staining of the betaA subunit and ActRIIA was seen in almost every epithelial cell, staining for the betaB subunit and ActRIIB was more variable. In situ hybridization and RT-PCR confirmed the presence of mRNA for the betaA, betaB subunits and ActRIIA and ActRIIB. These results indicated that the epithelium of the uterine tube is able to synthesize activin but not inhibin and has receptors for activin. Activins may thus act as paracrine regulators of tubal epithelial cell function, and embryonic activity may also bind to epithelial receptor and initiate intracellular processes that alter epithelial cell secretions. PMID- 15280565 TI - Induced radioactivity in CU targets produced by high-energy heavy ions and the corresponding estimated photon dose rates. AB - Irradiation experiments were performed at the Heavy Ion Medical Accelerator in Chiba (HIMAC) facility, National Institute of Radiological Sciences. The radioactive spallation products in a thick Cu target were obtained for Ar(230, 400 MeV per nucleon), Si(800 MeV per nucleon), Ne(100, 230, 400 MeV per nucleon), C(100, 230, 400 MeV per nucleon), He(100, 230 MeV per nucleon), p(100, 230 MeV) ions. The gamma-ray spectra from irradiated Cu samples inserted into the composite Cu target were measured with a high-purity germanium (HPGe) detector. From the gamma-ray spectra, we obtained the spatial distribution of radioactive yields of spallation products of 40 nuclides in the Cu sample in the Cu target. From the spatial distribution of radioactive yields, we estimated the residual activity and photon dose induced in the Cu target. The residual activity and photon dose become larger with the increase in projectile energy per nucleon and the range of the projectile beam for the same projectile energy per nucleon. PMID- 15280566 TI - Development of an automatic smear sampler and evaluation of surface contamination. AB - The surface contamination level of a radiation-controlled area is measured periodically according to atomic energy law and connection regulations. The measurement of surface contamination by an indirect method is subject to various kinds of error depending on the sampling person and consumes much time and effort in the sampling of large nuclear facilities. In this research, an automatic smear sampler is developed to solve these problems. The developed equipment is composed of a rotating sampling part, a sample transferring part, a power supply part a control part, and vacuum part. It improved the efficiency of estimation of the surface contamination level achieved periodically in a radiation-controlled area. Using an automatic smear sampler developed in this research, it is confirmed that radioactive contaminated materials are uniformly transferred to smear paper more than any sampling method by an operator. PMID- 15280567 TI - Multiple radon survey in spa of Loutra Edipsou (Greece). AB - Various investigators are involved with radon research in Greece. Numerous measurements have been reported for different regions around the country. In this study we focus on the city of Loutra Edipsou, a spa centre of about 4000 inhabitants in the northern part of the island of Evia (Greece). This city is built on an area of high geothermal activity, a fact that served as a guide to our study. The aim was to estimate the dose delivered to inhabitants, the potential alpha energy exposure of bathers and spa personnel and to investigate the radon background in this area. The detectors used were of active type. Measurements are reported for dwellings, which were grouped into two different sets: one representing typical summer periods (July and August 2002) and the other typical winter periods (October and November 2002) The Potential Alpha Energy Exposure (PAEE) of the bathers during treatment lies in the range between (45 +/- 3) and (110 +/- 5) mWLM (1 WLM = 12.97 J s m(-3)). The PAEE of the working personnel lies in the range between (34 +/- 3) and (100 +/- 20) mWLM. PMID- 15280568 TI - TNF therapy for spondyloarthropathy: can we marshal the argument? PMID- 15280569 TI - A novel mutation (T61I) in the gene encoding tumour necrosis factor receptor superfamily 1A (TNFRSF1A) in a Japanese patient with tumour necrosis factor receptor-associated periodic syndrome (TRAPS) associated with systemic lupus erythematosus. AB - OBJECTIVE: To identify potential mutations in the tumour necrosis factor receptor superfamily 1A gene (TNFRSF1A) in a Japanese female patient with recurrent fever complicated by systemic lupus erythematosus (SLE), and in her family members. METHODS: DNA sequencing of exons 1-10 of the TNFRSF1A gene was performed to determine mutations that might be associated with the tumour necrosis factor receptor-associated periodic syndrome (TRAPS). Moreover, the TNFRSF1A gene was examined in Japanese patients with autoimmune diseases, including SLE, rheumatoid arthritis (RA), mixed connective tissue disease (MCTD) and Behcet's disease, and in healthy Japanese controls. Enzyme-amplified sensitivity immunoassay (EASIA) analysis was used to assess serum levels of TNF, the 55-kDa TNF receptor (TNFRSF1A) and the 75-kDa TNF receptor (TNFRSF1B). Membrane TNFRSF1A expression was analysed on the surface of peripheral blood mononuclear cells by flow cytometry. RESULTS: A novel mutation, a heterozygous C to T transition in exon 3 which substitutes an isoleucine for a threonine at position 61 (T61I) was detected in the TNFRSF1A gene derived from the genomic DNA of a Japanese female TRAPS patient. Two nieces and one nephew, all with a similar clinical phenotype, also possessed the same TNFRSF1A mutation. We further demonstrated the same mutation in five of 60 SLE patients (8.3%) and in five of 120 healthy individuals (4.2%), with no significant differences. Although high titres of serum TNF and soluble TNFRSF1B protein were observed in this patient, low titres of soluble TNFRSF1A protein were detected. However, a defect in TNFRSF1A shedding in vitro was not observed in monocytes derived from this patient. CONCLUSION: This is the first report of a TRAPS patient associated with SLE with a novel TNFRSF1A mutation (T61I). PMID- 15280570 TI - Rheumatoid arthritis is already expensive during the first year of the disease (the Swedish TIRA project). AB - OBJECTIVE: To calculate direct and indirect costs in early rheumatoid arthritis (RA) and to characterize patients generating high and low costs respectively. METHODS: Two hundred and ninety-seven patients with recent-onset (< or = 12 months) RA were recruited. Clinical/laboratory data and 'health assessment questionnaire' (HAQ) were registered at inclusion and after 3, 6 and 12 months. After 6 and 12 months, the patients completed a questionnaire concerning health care utilization and days lost from work. A cut-off point for direct costs was set at 34,000 Swedish kronor (euro3675) defining one-third of the patients as a high-cost group and two-thirds as low-cost group. Indirect costs were calculated for patients aged <65 yr. RESULTS: Two hundred and eleven patients completed the HAQ on both occasions. Indirect costs exceeded direct costs by a factor of 2.3. Sixty three per cent experienced work disability during the first year and were identified as the 'high-indirect-cost group'. Indirect costs accounted for >70% of total costs. Direct costs included ambulatory health care (76%), hospitalization (12%) and medication (9%). Men aged > or = 65 yr had low costs compared with younger men and women of all ages. In multiple logistic regression tests, HAQ, high levels of IgM rheumatoid factor (IgM RF) and poor hand function increased the odds of entering the high-direct-cost group, and poor hand function and pain increased the odds of entering the high-indirect-cost group. CONCLUSIONS: Substantial costs were incurred during the first year after diagnosis of early RA, mainly due to work disability. Indirect costs were two to three times higher than direct costs. High levels of IgM RF, high HAQ score, poor hand function and pain increased the odds of entering high-cost groups. PMID- 15280571 TI - Factors associated with abnormal Pap results in systemic lupus erythematosus. AB - OBJECTIVE: Previous studies have suggested that women with systemic lupus erythematosus (SLE) are at greater risk for cervical dysplasia than are women in the general population. However, the factors associated with abnormal Pap test results in SLE have not been well studied. We therefore aimed to determine the factors associated with lifetime occurrence of an abnormal Pap test in women with SLE, and the influence of immunosuppressive exposure on the odds of abnormal Pap test results occurring after diagnosis of SLE. METHODS: Data were pooled from SLE cohorts from three centres. Self-report data were available on smoking, reproductive history, use of oral contraceptives (OC), history of sexually transmitted diseases (STDs) and whether the subjects had had cervical dysplasia on Pap testing. Logistic regression was used to examine the effect of these variables on the lifetime odds of cervical dysplasia. We then generated the adjusted odds ratio (OR) for the effect of immunosuppressive exposure on cervical dysplasia occurring after diagnosis of SLE. RESULTS: History of STDs and use of OCs were positively associated with reports of cervical dysplasia in adjusted analyses. The ORs for the effect of immunosuppressives on abnormal Pap test occurrence (adjusted for race, age, smoking, nulliparity, OC use and history of STDs) after diagnosis of SLE was 1.6 (95% CI 1.0, 2.7). CONCLUSIONS: A history of STDs and use of OCs were associated with abnormal Pap reports in this SLE sample. Immunosuppressive exposure may confer further risk to women with SLE. PMID- 15280572 TI - Hepcidin: inflammation's iron curtain. PMID- 15280573 TI - Binding of high-avidity anti-beta2-glycoprotein I antibodies. AB - OBJECTIVES: We studied the influence of different binding conditions on the interaction between high- or low-avidity IgG anti-beta2-glycoprotein I antibodies (anti-beta2-GPI) and beta2-GPI, which was either free in solution or bound to microtitre plates or nitrocellulose membranes. METHODS: Sera from 30 patients with antiphospholipid syndrome and/or systemic lupus erythematosus were selected on the basis of anti-beta2-GPI positivity. Avidity of IgG anti-beta2-GPI was determined by chaotropic ELISA, using increased NaCl concentration during the antibody binding. Immunodetection on nitrocellulose membrane followed reducing or non-reducing sodium dodecyl sulphate-polyacrylamide gel electrophoresis (PAGE) of beta2-GPI. In converted, non-reducing PAGE, the preparation with high-affinity Fab fragments (obtained by papain digestion) was subjected to electrophoresis, while purified beta2-GPI was used as the sample in immunodetection. RESULTS: Anti beta2-GPI antibodies of high, low or heterogeneous (low and high) avidity were found in 5/30, 9/30 and 16/30 sera, respectively. The density of beta2-GPI, which was 20-30 times higher on the nitrocellulose membrane than on the surface of ELISA plates, was not sufficient for the recognition of the antigen by anti-beta2 GPI: 2/5 high-avidity samples reacted only with non-reduced beta2-GPI, 3/9 low avidity samples recognized only denatured and reduced beta2-GPI, and 1/16 samples with heterogeneous-avidity antibodies reacted with reduced and non-reduced beta2 GPI. CONCLUSIONS: Our results suggest that neither high density of the antigen nor high avidity of the antibodies (or Fab fragments) alone was sufficient for the binding of anti-beta2-GPI to beta2-GPI. Some conformational modifications and, consequently, exposed neo-epitopes are required for the recognition of beta2 GPI by polyclonal anti-beta2-GPI antibodies. PMID- 15280574 TI - Cellular domains that contribute to Ca2+ entry events. AB - Stimulation of cell surface receptors that increase phosphatidylinositol 4,5 bisphosphate (PIP2) hydrolysis leads to intracellular Ca2+ release and activation of plasma membrane Ca2+ entry channels. Ca2+ entry via these channels regulates a wide array of physiological functions. The molecular composition of these channels and the mechanisms that activate or inactivate them have not yet been elucidated. Members of the TRPC subfamily of the TRP (transient receptor potential) family of proteins have been recently suggested as molecular components of these channels. In addition, Ca2+ signaling proteins and the signals they generate are compartmentalized and spatiotemporally regulated. Thus, the mechanisms involved in the assembly and trafficking of Ca2+ signaling proteins, including TRPC channels, will determine the regulation of Ca2+ entry and its effect on cellular function. PMID- 15280575 TI - Conformational coupling: a physiological calcium entry mechanism. AB - The entry of external Ca2+ that is activated by inositol 1,4,5-trisphosphate (IP3) may occur through a conformation coupling mechanism. IP3 receptors in the endoplasmic reticulum located in a junctional zone make contact with entry channels in the plasma membrane. IP3 may act directly to stimulate this coupling complex or IP3 could act indirectly by stimulating uncoupled IP3Rs in the vicinity of the junctional zone to induce a localized depletion of the ER store to switch on a store-operated mechanism. At physiological agonist concentrations, the earliest Ca2+ response to receptor activation may be the stimulation of entry, which is then responsible for charging up the internal store to prime the IP3Rs for the large-scale regenerative release of Ca2+ that occurs during each spike. PMID- 15280576 TI - Store-operated channels: diversity and activation mechanisms. AB - This perspective addresses two questions: How many store-operated channels (SOCs) are there, and how many mechanisms can account for SOC activation by depleted stores? Accumulating evidence suggests that the SOC family is not limited to the calcium-selective SOC that is responsible for ICRAC (Ca2+-SOC), but includes poorly selective cation SOCs (cat-SOCs) that may satisfy physiological needs in diverse excitable and nonexcitable cells. A growing number of studies in different cell types support the idea that all the members of SOC family (Ca2+ SOC and cat-SOC) may be activated by depletion of the stores through the same mechanism, which is mediated by calcium influx factor (CIF) and calcium independent phospholipase A2 (iPLA2). A conformational coupling model is also discussed. To account for the most recent findings, we propose that two distinct classes of calcium-conducting channels may exist in plasma membrane, which respond to different signals: SOCs, which are activated by depletion of calcium stores through the CIF-iPLA2 mechanism [no inositol triphosphate (IP3) needed]; and IP3 receptor-operated channels (IP3ROCs), which are activated by IP3 receptor through a direct coupling mechanism (no store depletion is needed). This model, with two separate mechanisms linked to different channels, may resolve many conflicting findings and interpretations and may give a new perspective on the diversity of calcium influx pathways. PMID- 15280577 TI - Receptor-operated cation entry--more than esoteric terminology? AB - Many hormones and neurotransmitters elicit an increase in the intracellular calcium concentration by binding to phospholipase C-linked G protein-coupled receptors. Activated receptors signal to calcium-permeable cation channels in the plasma membrane, which are distinct from those engaged by emptying of intracellular stores of calcium. The TRPC family of the mammalian homologs of the Drosophila transient receptor potential (TRP) cation channel represents likely molecular correlates underlying receptor-operated cation entry. While all TRPC family members are gated in a phospholipase C-dependent manner, the exact activation mechanism still remains elusive, although lipids such as diacylglycerol and polyunsaturated fatty acids are potential diffusible messengers. Functional TRPC channel complexes in the plasma membrane are thought to be composed of four distinct subunits whose stoichiometry and composition under physiological conditions are still largely unknown. However, recent progress in defining the combinatorial rules of TRPC channel assembly may lead to the identification of TRPC-dependent ion fluxes in living cells. Because of the large number of TRP proteins and their frequently overlapping functional characteristics, the central question is whether TRP proteins are functionally interchangeable or whether unique physiological roles can be ascribed to them. Receptor-operated cation entry is critically involved in the control of airway and vascular smooth muscle tone; hence, TRPC proteins are promising new drug targets. PMID- 15280578 TI - Store-operated Ca2+ entry channels: still elusive! AB - Depletion of intracellular Ca2+ stores is believed to trigger Ca2+ entry through a Ca2+- permeable channel in the plasma membrane called the store-operated Ca2+ channel (SOC). This type of Ca2+ entry is thought to play a pivotal role in a plethora of cell functions ranging from gene expression to sensory signal transduction. However, the molecular nature of these channels is still elusive. Many molecular candidates have been described as SOCs, but the candidacy of each has been countered by reports indicating that they are not SOCs. Most of the suggested candidates are members of the recently discovered superfamily of transient receptor potential cation channels (TRPCs). However, no TRP-family channel has yet been incontestably identified as an SOC. Even for the electrophysiologically best-described SOC, the highly Ca2+-permeable channel Ca2+ release-activated Ca2+ channel (CRAC), an acceptable candidate has yet to emerge. This perspective summarizes problems in identifying SOCs, suggests approaches to solving these problems, and discusses concerns over the current tendency to focus exclusively on the hypothesis that TRPCs comprise SOCs. PMID- 15280579 TI - Store-operated calcium channels: how do we measure them, and why do we care? AB - A major mechanism whereby calcium entry into cells is regulated is the store operated or capacitative calcium entry pathway. In this article, two basic issues are discussed: (i) the methods investigators use to measure store-operated entry, and (ii) the role played by the store-operated pathway in responses to hormones and neurotransmitters under physiological conditions. The two topics are considered together because they are closely interrelated; as we begin to ask questions about calcium movements at low concentrations of agonists, the technology to measure these movements becomes increasing challenging. PMID- 15280580 TI - Store-operated calcium entry: a tough nut to CRAC. AB - Store-operated or capacitative Ca2+ entry is a prominent feature of many electrically nonexcitable cell types. It is due to Ca2+ ion permeation pathways in the plasma membrane that are activated after receptor-mediated Ca2+ release from intracellular stores. Despite hundreds of publications on the topic of store operated Ca2+ entry and intense efforts by many dedicated laboratories, neither the molecular nature of the ion permeation pathway nor its activation mechanism is known. Here we review the progress made on the characterization of store operated currents and the challenges encountered in identifying the molecular components of store-operated Ca2+ entry. PMID- 15280581 TI - Toward a consensus on the operation of receptor-induced calcium entry signals. AB - Receptor-induced Ca2+ signals involve both Ca2+ release from intracellular stores and extracellular Ca2+ entry across the plasma membrane. The channels mediating Ca2+ entry and the mechanisms controlling their function remain largely a mystery. Here we critically assess current views on the Ca2+ entry process and consider certain modifications to the widely held hypothesis that Ca2+ store emptying is the fundamental trigger for receptor-induced Ca2+ entry channels. Under physiological conditions, receptor-induced store depletion may be quite limited. A number of distinct channel activities appear to mediate receptor induced Ca2+ entry, and their activation is observed to occur through quite diverse coupling processes. PMID- 15280582 TI - Receptor-activated calcium entry channels--who does what, and when? AB - The recent Science STKE E-Conference on Defining Calcium Entry Signals highlighted many of the outstanding problems and questions regarding the nature and regulation of the receptor-activated entry of Ca2+, particularly as it relates to Ca2+ signaling in nonexcitable cells. Frequently, these stem from the current lack of any clear candidates for the molecular identity of many of the major conductances involved. Moreover, there is considerable confusion in the field, largely as a result of the use of sometimes inappropriate or imprecise methodologies and inconsistent terminology. Nevertheless, much useful information is beginning to be revealed about the biophysical characterization of the fundamental properties of the channels involved and, at least in some cases, the specific conditions under which they are active. As a result, it is becoming clear that cells often contain various Ca2+ entry channels in addition to the ubiquitous store-operated, or capacitative, channels. These different channels are activated in distinct ways and operate under different conditions of stimulation. PMID- 15280583 TI - Culture and phenotyping of chondrocytes in primary culture. AB - The culture of chondrocytes is one of the most powerful tool for exploring the intracellular and molecular features of chondrocyte differentiation and activation. However, chondrocytes tend to dedifferentiate to fibroblasts when they are subcultured, which is a major problem. This chapter describes several protocols for culturing chondrocytes of different anatomical origins (articular and costal chondrocytes) from various species (humans, mice, rabbits, and cattle). All these protocols involve primary cultures in order to limit dedifferentiation. This chapter also describes a new protocol for culturing mouse articular chondrocytes. PMID- 15280584 TI - Culture of chondrocytes in alginate beads. AB - A classic method for the encapsulation and culture of chondrocytes in alginate beads is described. Chondrocytes are released from cartilage matrix by collagenase/dispase digestion and mixed with a solution of 1.25% alginic acid until a homogenous suspension is obtained. The suspension is drawn into a syringe and pushed gently through a needle, so that drops fall into a solution of calcium chloride. Beads form instantaneously and further polymerize after 5 min in the calcium chloride solution. Chondrocytes from any species, including human osteoarthritic chondrocytes, can be cultured with this technique. Under these conditions, chondrocytes maintain a high degree of differentiation. Beads can be dissolved by chelation of calcium with EDTA. In this way, chondrocytes can be recovered and further separated from the matrix by centrifugation. Almost all molecular and biochemical techniques, as well as a number of biological assays, are compatible with the culture of chondrocytes in alginate. PMID- 15280585 TI - Immortalization of human articular chondrocytes for generation of stable, differentiated cell lines. AB - Immortalized chondrocytes of human origin have been developed to serve as reproducible models for studying chondrocyte function. In this chapter, methods for immortalization of primary human chondrocytes with SV40-TAg, HPV-16 E6/E7, and telomerase by retrovirally mediated transduction and selection for neomycin resistance are described. However, stable integration of an immortalizing gene stabilizes proliferative capacity, but not the differentiated chondrocyte phenotype. Thus, strategies for selection of chondrocyte cell lines, involving the maintenance of high cell density and moderation of cell proliferation, are also described. The methods for immortalization and selection are applicable to the development of chondrocyte cell lines using any immortalizing agent. Although immortalized chondrocytes should not be considered as substitutes for primary chondrocytes, they may be useful tools for evaluating and further validating mechanisms relevant to cartilage biology. PMID- 15280586 TI - Culture of immortalized chondrocytes and their use as models of chondrocyte function. AB - Immortalization of chondrocytes increases life span and proliferative capacity but does not necessarily stabilize the differentiated phenotype. Expansion of chondrocyte cell lines in continuous monolayer culture may result in the loss of phenotype, particularly if high cell density is not maintained. This chapter describes strategies for maintaining or restoring differentiated phenotype in established chondrocyte cell lines involving culture in serum-free defined culture medium, in suspension over agarose or polyHEMA, or within alginate or collagen scaffolds. Chondrocyte cell lines have been used successfully to develop reproducible models for studying the regulation of gene expression in experiments requiring large numbers of cells. Thus, approaches for studying transcriptional regulation by transfection of promoter-driven reporter genes and cotransfection of expression vectors for wild-type or mutant proteins are also described. PMID- 15280587 TI - Generation of pluripotent stem cells and their differentiation to the chondrocytic phenotype. AB - It is well documented that adult cartilage has minimal self-repair ability. Current methods for treatment of cartilage injury focus on the relief of pain and inflammation and have met with limited long-term success. In the forefront of new therapeutic approaches, autologous chondrocyte transplantation is still only applied to a very small percentage of the patient population. Our laboratory has focused on cartilage repair using progenitor cells and studied their differentiation into cartilage. Adult mesenchymal stem cells are an attractive candidate as progenitor cells for cartilage repair because of their documented osteogenic and chondrogenic potential, ease of harvest, and ease of expansion in culture; furthermore, their use will obviate the need for harvesting precious healthy cartilage from a patient to obtain autologous chondrocytes for transplantation. However, the need to induce chondrogenic differentiation in the mesenchymal stem cells is superposed on other technical issues associated with cartilage repair; this adds a level of complexity over using mature chondrocytes. This chapter focuses on the methods involved in the isolation of human mesenchymal stem cells and their differentiation along the chondrogenic lineage. Although we have the technology to accomplish chondrogenic differentiation of stem cells, much is still to be learned regarding the regulatory mechanisms controlling the lineage transitions and maturation of the cartilaginous tissue. PMID- 15280588 TI - Semiquantitative analysis of gene expression in cultured chondrocytes by RT-PCR. AB - Reverse transcriptase-polymerase chain reaction (RT-PCR) is a powerful, sensitive, and rapid method to monitor small amounts of nucleic acids. This is of particular interest for small amounts of cells, as in cartilage. We present here two protocols to isolate total RNA and a protocol to study matrix metalloproteinase and type II collagen gene expression from chondrocytes of human origin. Specific gene expression is revealed on an ethidium bromide-containing agarose gel on an ultraviolet plate and normalized to that of a housekeeping gene. PMID- 15280589 TI - Quantification of mRNA expression levels in articular chondrocytes with PCR technologies. AB - Unlike any other technology in molecular biology, the polymerase chain reaction (PCR) has changed the technological armamentarium of molecular scientists working on cartilage, in terms of outstanding sensitivity and accuracy. Four approaches to determine mRNA expression levels by PCR amplification of specific cDNA sequences are currently in use and are discussed in this chapter: conventional PCR with end-point determination, conventional PCR in the logarithmic amplification phase, conventional PCR using internal competitive DNA fragments, and real-time PCR as offered by TaqMan technology and others. The determination of mRNA expression levels by real-time quantitative PCR appears to be the most reliable method for accurate determination of gene expression levels within cartilage and cultured chondrocytes, as in other tissues and cell types. This technology offers outstanding sensitivity and accuracy in terms of determination of the amount of cDNA molecules. However, this method cannot account for factors such as efficiency of RNA isolation and reverse transcription conditions. Thus, normalization of the acquired data is required, with all its limitations as described. PMID- 15280590 TI - RNA extraction from cartilage. AB - The direct isolation of RNA from cartilage has often proved difficult owing to a number of factors. Cartilage has a low cell content and contains an extracellular matrix rich in proteoglycans, which copurify with the RNA as they are large and negatively charged macromolecules. In our laboratory, we are interested in searching for genes differentially expressed in chondrocytes in diverse in vivo situations, for instance during maturation of chondrocytes in the growth plate or during cartilage degeneration. We found that treatment by proteinase K in 1 M guanidinium isothiocyanate prior to cesium trifluoroacetate ultracentrifugation was crucial to increase the yield and purity of RNA extracted from cartilage matrix. This protocol indeed led to reproducible patterns of differential display reverse transcriptase-polymerase chain reaction (RT-PCR) and should be useful for identifying genes differentially expressed by chondrocytes in situ. PMID- 15280591 TI - Gene expression analysis in cartilage by in situ hybridization. AB - In situ hybridization allows detection and localization of specific nucleic acid sequences directly within a cell or tissue. We present an in situ hybridization protocol using double-stranded DNA or single-stranded RNA probes labeled with [32P] to localize and visualize the temporal and spatial distribution of cartilage-characteristic mRNAs. Probes labeled with this high-energy isotope provide good resolution at the tissue level with relatively low background; as a result of the probes that can be obtained that have a higher specificity to emulsion activity, very short exposure times are required. PMID- 15280592 TI - Analysis of differential gene expression in healthy and osteoarthritic cartilage and isolated chondrocytes by microarray analysis. AB - The regulation of chondrocytes in osteoarthritic cartilage and the expression of specific gene products by these cells during early-onset and late-stage osteoarthritis are not well characterized. With the introduction of cDNA array technology, the measurement of thousands of different genes in one small tissue sample can be carried out. Interpretation of gene expression analyses in articular cartilage is aided by the fact that this tissue contains only one cell type in both normal and diseased conditions. However, care has to be taken not to over- and misinterpret results, and some major challenges must be overcome in order to utilize the potential of this technology properly in the field of osteoarthritis. PMID- 15280593 TI - High-efficiency nonviral transfection of primary chondrocytes. AB - The introduction of foreign DNA into mammalian cells is an essential investigative tool in molecular biology. Nonviral approaches to transfection offer the advantage of relatively simple vector design, production, and purification and, for tissue engineering applications, avoid many of the potential risks associated with virus-mediated transfection methods. Unfortunately, primary cells, and in particular chondrocytes, are notoriously refractory to conventional transfection approaches, and optimized transfection efficiencies in these cells are extremely low (1-1.5%). In this chapter, we present three protocols that have proved useful in transfecting primary chondrocytes at high efficiency (~70%). The first uses radiofrequency electroporation, a transfection method that frequently works extremely well in cell types that are difficult to transfect. It should be noted that electroporation is not limited to DNA but that essentially any molecule can be introduced into the cell using this approach. In addition to the primary protocol, we present two additional reliable, albeit less efficient backup protocols, the first using exponential decay electroporation and the second FuGENE 6 transfection. PMID- 15280594 TI - In vitro gene transfer to chondrocytes and synovial fibroblasts by adenoviral vectors. AB - The major requirement of a successful gene transfer is the efficient delivery of an exogenous therapeutic gene to the appropriate cell type with subsequent high or regulated levels of expression. In this context, viral systems are more efficient than nonviral systems, giving higher levels of gene expression for longer periods. For the application of osteoarthritis (OA), gene products triggering anti-inflammatory or chondroprotective effects are of obvious therapeutic utility. Thus, their cognate genes are candidates for use in the gene therapy of OA. In this chapter, we describe the preparation, the use, and the effect of the transduction of chondrocytes or synovial fibroblasts with an adenoviral vector encoding the cDNA for glutamine: fructose-6-phosphate amidotransferase (GFAT). This is intended to serve as an example of a technology that can be used to evaluate the biological effects of overexpression of other cDNAs. PMID- 15280595 TI - Changes of chondrocyte metabolism in vitro: an approach by proteomic analysis. AB - Changes in chondrocyte metabolism in vitro using different support systems and under different culture conditions were studied with a proteomic approach. Qualitative and quantitative modifications in the synthesis of chondrocyte proteins were investigated using two-dimensional (2D) gel electrophoresis. This technique provided a simple way to visualize the most abundant chondrocyte proteins. Proteins were identified after in-gel proteolysis with trypsin and matrix-assisted laser desorption ionization-time of flight mass spectrometry, using peptide mass fingerprinting. Tryptic peptide masses were measured and matched against a computer-generated list from the simulated trypsin proteolysis of a protein database (SwissProt). PMID- 15280596 TI - Analysis of chondrocyte functional markers and pericellular matrix components by flow cytometry. AB - Flow cytometry has been used as a procedure to characterize the phenotype and function of human articular cartilage cells cultured as monolayers or in gelled artificial matrices. Procedures allowing intact cells with their cell-associated matrix, to be obtained have been described. Appropriate monoclonal antibodies have allowed plasma membrane-associated proteins, e.g., growth factors and cytokine receptors, as well as the cell-associated extracellular matrix macromolecules, to be studied. Intracellular compounds have been traced in permeabilized cells after blocking of their intracellular transport and secretion mechanisms. We report the use of fluorescent dye-labeled monoclonal antibodies or specific binding proteins against extracellular matrix compounds such as hyaluronan, aggrecan, types I and II collagen, and fibronectin. The autocrine and paracrine growth factor and cytokine pathways considered include the insulin-like growth factor-1 (IGF-1)/IGF receptor I (IGFRI), and the transforming growth factor-beta1 (TGF-beta1)/TGF-beta receptor II (TGF-betaRII) cascades, as well as the interleukin-1alpha/beta (IL-1alpha/beta)/interleukin-1 receptors I and II (IL 1RI and II) systems. Catabolic enzymes that mediate extracellular matrix turnover, e.g., some matrix metalloproteinases and their natural inhibitors, were also studied. Finally, flow cytometry was used to assess the results of some pharmacological interventions on the aforementioned variables in cultured chondrocytes. PMID- 15280597 TI - A simple and reliable assay of proteoglycan synthesis by cultured chondrocytes. AB - A simple and reliable method to measure proteoglycan synthesis by chondrocytes in culture is described. Confluent chondrocytes in 24-well plates are labeled for 24 72 h with (35)SO4(2-) in the presence of stimulating agents. At the end of treatment, the secretion medium containing radiolabeled neosynthesized secreted proteoglycans (SP) is harvested, and cell-associated proteoglycans (CAP) are extracted with guanidine hydrochloride for 48 h. Aliquots of medium and cell extracts are distributed on Whatman paper and left to dry. SP and CAP are trapped by precipitation with the cationic detergent cetyl pyridinium chloride (CPC), whereas nonincorporated (35)SO4(2-) remains in solution. After drying, each spot corresponding to one well in the plate is cut, and its radioactivity is measured. Counts are proportional to the amount of neosynthesized proteoglycans. In a representative experiment using rabbit chondrocytes, total proteoglycan synthesis (SP plus CAP) was increased 3.5-fold after the addition of 1.34 nM insulin-like growth factor-1 (IGF-1) compared with nonstimulated cells. Further addition of a fourfold molar ratio of IGF binding protein-3 completely abolished this effect. This method can be used to measure proteoglycan synthesis by chondrocytes from many species, including human osteoarthritic chondrocytes, as well as guinea pig, rabbit, and rat chondrocytes. PMID- 15280598 TI - Assays of proteoglycan and collagen degradation in cultures of rabbit cartilage explants. AB - Cultures of cartilage explants have long been used to study the effects of modulators of extracellular matrix degradation. We present a simple and rapid assay system, based on culture of rabbit cartilage explants, which permits study of the effects of protease inhibitors on proteoglycan degradation (caused by either aggrecanases or matrix metalloproteinases [MMPs]), and on collagen degradation. The assay is based on the ability of interleukin-1 to stimulate both aggrecanase activity and synthesis of inactive MMPs, which are then activated by p-aminophenylmercuric acetate for the study of MMP-mediated proteoglycan degradation or by plasmin for the study of collagen degradation. Proteoglycan degradation is quantified as percent release of radioactivity from cartilage explants previously labeled with (35)SO4(2-). Collagen degradation is calculated as percent release of collagen, measured by colorimetric assay of hydroxyproline. PMID- 15280599 TI - Production of antibodies against degradative neoepitopes in aggrecan. AB - The use of synthetic peptides to generate rabbit polyclonal anticatabolic neoepitope antibodies that can be used to study the presence of defined proteolytic cleavage sites in aggrecan is described. Principles of peptide design and methods for preparation and characterization of ovalbumin conjugates are presented along with approaches for the characterization and affinity purification of the resulting antisera. Limitations associated with the use of antipeptide antibodies to study authentic protein neoepitopes are discussed. PMID- 15280600 TI - Immunoassays for collagens in chondrocyte and cartilage explant cultures. AB - Quantitative immunoassays have been developed to measure the content, degradation, and synthesis of types II and IX collagens in hyaline cartilages. Some of these assays and their applications are described in this chapter. These and other assays are commercially available. The applications of these assays are discussed with examples from recent publications. PMID- 15280601 TI - Detection of apoptosis in cartilage in situ and in isolated chondrocytes. AB - Apoptosis is a form of programmed cell death conceptually opposed to necrosis. In view of the inherent difficulty in accurately detecting apoptosis in chondrocytes, this chapter describes complementary techniques that may be used in combination. During apoptotic death, protein and DNA breakdown is accomplished by caspases (cysteine proteases) in a highly regulated manner. Activation of caspases occurs in the initiation and/or the execution phase of certain apoptotic programs and represents an early physiologic marker of apoptosis. Here we present an immunoblotting technique that allows the detection of caspase-3 processing in cultured human chondrocytes. Apoptosis leads to plasma membrane asymmetry and to externalization of phosphatidylserine residues, which are bound with high affinity by annexin V. In the early stages of apoptosis, cells typically have an intact cell membrane. Apoptotic cells will not stain positive with propidium iodide, whereas externalization of phosphatidylserine will be detected by annexin V. Terminal deoxynucleotidyl transferase (TdT)-mediated nick-end labeling (TUNEL) works on the principle that DNA strand breaks (single or double) that occur during apoptosis can be identified by labeling free 3'-hydroxyl termini. Labeled nucleotides are polymerized to these termini in a reaction catalyzed by TdT. The tissue can then be examined histologically for identification of TUNEL-positive cells in situ. PMID- 15280602 TI - Expression, activity, and regulation of MAP kinases in cultured chondrocytes. AB - The mitogen-activated protein (MAP) kinase family consists of extracellular signal-regulated protein kinase (ERK), p38 kinase, and c-Jun N-terminal kinase (JNK) and transduces signals from the extracellular environment to the cytoplasm and nucleus. MAP kinase signaling involves a multistep kinase cascade including MAP kinase kinase kinase (MAPKKK), MAP kinase kinase (MAPKK), and MAP kinase. The MAP kinase subtypes are constitutively expressed in articular chondrocytes and they regulate chondrocyte function, including differentiation, apoptosis, inflammatory responses, and activation of matrix metalloproteinases. Therefore, imbalance or destruction of homeostasis regulating MAP kinase activity is related to the pathogenesis of cartilage diseases such as osteoarthritis. This chapter describes methods for measuring and modulating MAP kinase subtype activity in primary cultured articular chondrocytes and cartilage explants. PMID- 15280603 TI - Mechanical loading of chondrocytes embedded in 3D constructs: in vitro methods for assessment of morphological and metabolic response to compressive strain. AB - Mechanical loading of chondrocytes in 3D constructs has been used to investigate mechanotransduction and its potential for stimulating tissue-engineered cartilage repair. This chapter describes the preparation of 3D agarose or alginate constructs seeded with isolated chondrocytes and specific test rigs for applying gross compressive strain to individual constructs on a confocal microscope or for longer term compression of constructs cultured within an incubator. Experimental methods are described to quantify the level of cell deformation and the elaboration of extracellular matrix. The chapter thus provides an introduction to the experimental techniques used to examine chondrocyte mechanotransduction and downstream cell function. PMID- 15280604 TI - In vitro physical stimulation of tissue-engineered and native cartilage. AB - Because of the limited availability of donor cartilage for resurfacing defects in articular surfaces, there is tremendous interest in the in vitro bioengineering of cartilage replacements for clinical applications. However, attaining mechanical properties in engineered cartilaginous constructs that approach those of native cartilage has not been previously achieved when constructs are cultured under free-swelling conditions. One approach toward stimulating the development of constructs that are mechanically more robust is to expose them to physical environments that are similar, in certain ways, to those encountered by native cartilage. This is a strategy motivated by observations in numerous short-term experiments that certain mechanical signals are potent stimulators of cartilage metabolism. On the other hand, excess mechanical loading can have a deleterious effect on cartilage. Culture conditions that include a physical stimulation component are made possible by the use of specialized bioreactors. This chapter addresses some of the issues involved in using bioreactors as integral components of cartilage tissue engineering and in studying the physical regulation of cartilage. We first consider the generation of cartilaginous constructs in vitro. Next we describe the rationale and design of bioreactors that can impart either mechanical deformation or fluid-induced mechanical signals. PMID- 15280605 TI - Increasing age at diagnosis of celiac disease in Malta. AB - OBJECTIVE: Recent studies have shown that celiac disease is being diagnosed at a progressively later age. This paper analyses trends in age at diagnosis in a closed island population. METHODS: Patient case notes of all known patients with celiac disease were retrieved and demographic information, mode of presentation including symptomatology and diagnostic criteria were obtained. Over the period 1985 to 2000, information was available on 42 patients with celiac disease. All were aged 14 years. RESULT: The mean age at diagnosis showed an increasing age, with a constant disease incidence. A highly significant positive correlation was found for age at diagnosis with time (rho=0.4, p=0.009). CONCLUSION: It is thought that breast feeding, with later introduction of gluten containing products in the diet, leads to a later presentation of celiac disease. In Malta, over the period under study, the breast feeding rate rose from 20% to 60%. Our findings support the hypothesis that breast feeding offers a degree of protection against the early development of celiac disease, without actually reducing the incidence of the disease. PMID- 15280606 TI - Epidemiological study of measles in slum areas of Kolkata. AB - OBJECTIVE: An epidemiological study on measles was conducted among the under five children in slum areas of Kolkata to assess the incidence of measles. METHODS: 20 cluster sampling technique was followed and in each cluster 250 under five children were covered. Paramedical workers identified children of the target of age group who had history of measles in past one year and the medical officer confirmed the diagnosis following standard case definition. RESULTS: Incidence of measles was found as 5.76%. Incidence was equal in both the sex groups, but found more among infants. The incidence of measles gradually declined with the increase of age strengthening the view in favor of early immunization. Amongst the measles cases only 19.7% were immunized with measles vaccine. 100% of measles cases gave history of rash, 98.9% had history of fever, 82.8% reported that the rash started from face and progressed downwards to abdomen and leg. Cough, redness of eye and discoloration of skin were reported by 97.5%, 83.8% and 65.2% measles cases respectively. Only 16.9% and 8.6% children received Vitamin A oil before and after the illness respectively. 49% gave history of exposure to measles cases and 46.6% cases had measles within 2 weeks incubation period. CONCLUSION: The study highlighted the necessity of timely measles vaccine coverage, additional dose at a higher age and Vitamin A supplementation through IEC activities. PMID- 15280607 TI - Demographic profile and outcome analysis of a tertiary level pediatric intensive care unit. AB - OBJECTIVE: To study the profile and outcome of children admitted to a tertiary level pediatric intensive care unit (PICU) in India. METHODS: Prospective study of patient demographics, PRISM III scores, diagnoses, treatment, morbidity and mortality of all PICU admissions. RESULTS: 948 children were admitted to the PICU. Mean age was 41.48 months. Male to female ratio was 2.95:1. Mean PRISM III score on admission was 18.50. Diagnoses included respiratory (19.7%), cardiac (9.7%), neurological (17.9%), infectious (12.5%), trauma (11.7%), other surgical (8.8%).196 children (20.68%) required mechanical ventilation. Average duration of ventilation was 6.39 days. 27 children (30.7 children /1000 admissions) had acute respiratory distress syndrome. Gross mortality was 6.7% (59 patients). PRISMIII adjusted mortality was directly proportional to PRISMIII scores. 49.5% of nonsurvivors had multiorgan failure. Average length of PICU stay was 4.52 +/- 2.6 days. Complications commonly encountered were atelectasis (6.37%), accidental extubation (2%), and pneumothorax (0.9%). Incidence of nosocomial infections was 16.86%. CONCLUSION: Our data appears to be similar with regards to PRISMIII scores and adjusted mortality, length of the PICU stay, and duration of ventilation, to previously published western data. Multiorgan failure remains a major cause of death. As expected, Dengue and malaria were common. Incidence of nosocomial infections was somewhat high. Interestingly, more boys got admitted to the PICU as compared to girls. Clearly more studies are required to assess the overall outcomes of critically ill children in India. PMID- 15280608 TI - Spectrum of holoprosencephaly. AB - OBJECTIVE: To conduct a clinical study of holoprosencephaly (HPE). METHOD: Thirteen cases of HPE were studied regarding their clinical features, family history, and prenatal and imaging studies. Chromosomal analysis was done whenever fresh sample was available. RESULTS: Six cases were antenatally detected by ultrasound; four cases were stillborn. Three cases were identified by neuroimaging done a part of evaluation of developmental delay or cleft lip. Eleven of them had facial anomalies characteristics of HPE. Two of these had subtle facial features and microcephaly. Karyotype was abnormal in 2 of 7 cases studied. CONCLUSION: Most of the cases of HPE present antenatally or at birth. Milder forms like lobar and semilobar can present as developmental delay during infancy. Facial anomalies are usually associated with HPE. Chromosomal study of the case and clinical examination of the parents is essential for providing information regarding risk of recurrence to the family. PMID- 15280609 TI - Pediatric tonsillopharyngitis--an evaluation of cefprozil in Indian patients. AB - OBJECTIVE: The emergence of penicillin resistant strains and the presence of co pathogens have made the treatment of bacterial infections in children a challenge. Streptococcal tonsillopharyngitis, which is a common infection has been well treated with cefprozil, a novel third generation cephalosporin. The aim of the present study was to evaluate cefprozil in pediatric tonsillopharyngitis. An assessment of the clinical cure and bacteriological eradication rates and an overall tolerability was made. METHODS: It was a prospective, open, non comparative multicentric study. 316 children (mean age 6.61 years) with tonsillopharyngitis were included. Patients were given cefprozil susp 15 mg/kg/day in two divided doses a day for 10 days. RESULTS: A clinical cure of 96.6% and bacteriological eradication of 94.29% was achieved with cefprozil. Overall tolerability of cefprozil was assessed by physicians and 46% rated tolerability of cefprozil as excellent, 38% as very good, 10% as good, 6% as fair and none as poor. CONCLUSION: Cefprozil has been found to be an excellent drug of superior microbiological and clinical activity in the treatment of pediatric patients with tonsillopharyngitis. The drug also has an expanded spectrum. PMID- 15280610 TI - Japanese encephalitis in India: an overview. AB - Japanese encephalitis (JE)-epidemics have been reported in many parts of the country. The incidence has been reported to be high among pediatric group with high mortality. The incidence of JE in recent times is showing an increasing trend. It appears that JE may become one of the major public health problems in India, considering the quantum of the vulnerable pediatric population, the proportion of JEV infections among the encephalitic children and wide scattering of JE-prone areas. JE burden can be estimated satisfactorily to some extend by strengthening diagnostic facilities for JE confirmation in hospitals and by maintenance of contact with the nearby referral hospitals to collect the particulars on JE cases. Vaccination proves to be the best to protect the individual against any disease. In the case of JE, it is essential to immunize the pigs (amplifying host) also to interrupt the transmission of the disease. PMID- 15280611 TI - Standard precautions and post exposure prophylaxis for preventing infections. AB - In health care set up, risk of acquiring infection by both patients and health care worker (HCW) from each other is fairly high. Despite progress, hospital acquired infections (HAI) are a problem in both developed and developing countries and are an important cause of death. Many different microbes cause HAI in both patients and HCW; these include various commensals, pathogenic bacteria, viruses, parasites, and fungi. Among these HIV, hepatitis B, and hepatitis C are of major significance to HCW. 'Standard precautions' have now replaced the term 'universal precautions', and are designed to reduce the risk of transmission of microorganisms in health care set-up from both recognized and unrecognized sources. Ultimate aim is to reduce the risk of disease transmission in the healthcare setting, both to the patient and the provider, and thus reduce morbidity. This applies to all patients, regardless of their diagnosis and expands the coverage of universal precautions by recognizing that any body fluid may contain contagious and harmful microorganisms. This article reviews the standard precautions and discusses current guidelines on post exposure prophylaxis (PEP). PMID- 15280612 TI - Evaluation of a child with cerebral palsy. AB - Evaluation of a child with cerebral palsy (CP) requires a multidisciplinary approach with a team of professionals comprising of a pediatrician or pediatric neurologist, occupational therapist, a physiotherapist, child psychologist, and a social worker. The assessment is necessary to confirm the diagnosis, determine the cause, assess the motor function and associated problems. The diagnosis of CP is clinical but selected investigations may be required for ascertaining the cause. Evaluation includes assessment for common medical problems of childhood particularly nutritional disorders and assessment of family functioning. Additional disabilities are common. Routine assessment of vision and hearing is required in children with CP. Since CP is a changing disorder, some limitations may not be evident early in life but manifest in the school age or later. The evaluation of a child with CP is an ongoing process and should be a part of continuing care as the child grows from infancy to adolescence. PMID- 15280613 TI - Cerebral palsy-management. AB - Cerebral Palsy encompasses a heterogeneous group of non-progressive motor disorders caused by injury to the developing brain. Management is best done in a multidisciplinary set up under one roof. Comprehensive assessment of the child to evaluate functional ability and associated problems is followed by an individualized plan of management with long term goals and short term objectives. Participation of the family is pivotal to ensure proper habilitation of the child. A home-based management plan is advocated. Considerable experience, sensitivity and understanding are needed both for breaking the news and counselling the parents of a child with cerebral palsy. PMID- 15280614 TI - Cranial computed tomography in partial motor seizures. AB - OBJECTIVE: To identify the pattern of intracranial structural lesions in developmentally normal children with partial motor seizures by computed tomography and to monitor the behavior of single ring enhancing lesion (SREL) after a period of time with or without treatment. METHODS: Consecutive developmentally normal children between one year and twelve years with partial motor seizures in a tertiary care referral Hospital. After clinical examination and appropriate investigation for tuberculosis and cysticercosis, CT scan was performed. In addition to anticonvulsants, children received antituberculous or anticysticercal therapy if indicated. Repeat CT was performed on children with SREL after 6 months. RESULTS: Computed tomography was abnormal in 102 (68%) children. Majority of the children (75) had SREL. The lesions were located in decreasing order of frequency in the parietal lobe (65), frontal lobe (7), occipital lobe (1), temporal lobe (1) and cerebellum (1). Repeat CT scan was performed on 50 of the 75 children with SREL. Among these, in 41 children who were only on antiepileptic therapy, the SREL had decreased in size in thirty-two whereas in the rest (9), there was no change in the size. CONCLUSION: Awareness of the existence of disappearing SREL lesions is essential to avoid unnecessary treatment with antituberculous or anticysticercal therapy and provides ample justification in treating with anticonvulsant drugs only. PMID- 15280615 TI - Intermittent hyperammonemic encephalopathy in a child with ornithine transcarbamylase deficiency. AB - Ornithine transcarbamylase (OTC) deficiency is an X-linked disorder and the most common inherited cause of hyperammonemia. Clinical manifestations are more severe in hemizygous males who often present in neonatal period. Heterozygous females may be asymptomatic until juvenile or adulthood. Fluctuating concentration of ammonia, glutamine and other excitotoxic amino acids result in a chronic or episodically recurring encephalopathy. The authors report a heterozygous female with OTC deficiency who presented with recurrent encephalopathy. PMID- 15280616 TI - Self induced photosensitive epilepsy. AB - The authors report a case of a 12-year-old girl who had rare self induced photosensitive epilepsy. She used to move her right hand over the right eye while simultaneously rubbing the forehead since the age of 8. During these episodes she was lost in herself. Lately these episodes were followed by brief spell of unconsciousness. The EEG examination, in its third attempt, revealed bilateral multiple symmetric spikes on photic stimulation. She admitted that she often induced the episode herself and derived pleasure out of it. She responded well to Sodium valproate. PMID- 15280617 TI - MURCS association. AB - The MURCS association i.e. MUllerian duct aplasia, Renal aplasia, Cervicothoracic Somite dysplasia is a rare developmental disorder. The authors report a case of MURCS association with supernumerary ribs in a 7-month-old infant who presented with failure to thrive. PMID- 15280618 TI - Mondini dysplasia and pyogenic meningitis. AB - Mondini dysplasia with cerebrospinal fluid leak is a rare cause of recurrent pyogenic meningitis in children. We describe an eleven-year-old female child who presented with the fifth recurrent episode of pyogenic meningitis and unilateral sensorineural deafness. Mondini dysplasia of the inner ear with CSF-perilymph fistula was proven on an HRCT of the temporal bone and MRI. Successful operative intervention was undertaken to close the defect. Though rare, Mondini dysplasia should be considered as a cause of recurrent meningitis in children, especially if they have sensorineural deafness. PMID- 15280619 TI - Tuberous sclerosis complex. PMID- 15280620 TI - Nasogastric feeding tube gastric perforation in a neonate. PMID- 15280621 TI - Association between breast cancer laterality and tumor location, United States, 1994-1998. AB - OBJECTIVE: Cancer is more likely to be diagnosed in the left breast than the right, but the reasons are undetermined. Left-sided predominance has not been evaluated for some demographic groups or by tumor location. METHODS: Laterality was analyzed among 419,935 incident unilateral breast cancers from 26 population based cancer registries covering 40% of the US population. Logistic regression assessed the independent contribution of race, ethnicity, age, histology, stage, and location to laterality. RESULTS: Breast cancer was about 5% more likely to be diagnosed in the left breast than the right, a finding that was generally consistent across demographic groups and tumor types. Left-sided predominance was evident among both younger (<45 years) and older women, and among men with either in situ or invasive disease. Among women, tumors in the upper-outer quadrant, where one-third of cancers are located, occurred with equal frequency in the left and right breast, while those in the lower quadrants were about 10% more likely to occur in the left breast. CONCLUSION: The observation that the left breast is at greater risk of cancer than the right may not apply to tumors arising in the upper-outer quadrant. The identification of physiologic, pathologic, or immunologic differences between the lower, but not upper, left and right breasts may assist in explaining breast cancer laterality. PMID- 15280622 TI - Lifestyle and prostate cancer among older African-American and Caucasian men in South Carolina. AB - OBJECTIVE: We investigated the association between lifestyle and prostate cancer risk among Caucasian and African-American men, separately. METHODS: This population-based case-control study of prostate cancer among men aged 65-79 years was conducted between 2000 and 2002 in South Carolina. Telephone interviews were completed with 416 incident prostate cancer cases ascertained through the South Carolina Central Cancer Registry, and 429 controls identified through the Health Care Financing Administration Medicare beneficiary file (with respective response rates of 71% and 64%). RESULTS: Caucasian men working in production, transportation, and material moving had increased prostate cancer risk (odds ratio [OR] = 2.04, 95% confidence interval [CI] 1.17-3.54), while African American men in the military had reduced prostate cancer risk (OR = 0.19, 95% CI 0.05-0.76). Having five or more prostate specific antigen (PSA) tests within the past five years was associated with prostate cancer among Caucasian men; however, African-American men with prostate cancer tended to have fewer PSA tests. Increasing lycopene consumption was associated with a reduced risk of prostate cancer among Caucasian men (p = 0.03), but not among African-American men. CONCLUSIONS: In this population-based case-control study conducted in South Carolina we did not find marked differences in lifestyle factors associated with prostate cancer by race. PMID- 15280623 TI - Increased risk of developing hepatocellular carcinoma associated with carriage of the TNF2 allele of the -308 tumor necrosis factor-alpha promoter gene. AB - BACKGROUND AND AIMS: Tumor necrosis factor-alpha (TNF-alpha) is a cytokine that may act as an endogenous tumor promoter. A genetic polymorphism of TNF-alpha at position -308 of the promoter region, which includes TNF1 (-308G) and TNF2 ( 308A) alleles, has been found to be associated with susceptibility to various types of cancer. We conducted a study to evaluate the association between this polymorphism and hepatocellular carcinoma (HCC). METHODS: We recruited 74 HCC patients and 289 healthy controls, and determined their -308 TNF-alpha promoter genotypes through polymerase chain reaction followed by electrophoresis. RESULTS: Carriage of the TNF2 allele was associated with an increased risk of HCC (odds ratio [OR] = 3.5; 95% confidence interval [CI]:[2.1, 6.0]), and a trend toward a significant increase in the risk of developing HCC was observed from TNF1/TNF1, TNF1/TNF2, to TNF2/TNF2 genotypes (p < 0.01). After adjustment for gender, age, and markers of hepatitis B and C, the OR of developing HCC associated with TNF2 allele carriage was 5.3 (95% CI: [2.3, 12.1]; p < 0.01) CONCLUSIONS: Carriage of the TNF2 allele is a significant predictor of HCC independent of hepatitis B and C, and therefore it may be used as a biomarker for susceptibility to HCC. PMID- 15280624 TI - Lung cancer incidence trends in black and white young adults by gender (United States). AB - OBJECTIVE: This study analyzed temporal trends in lung cancer incidence rates for young adults, as an indicator of the recent and potential future impact of risk factor trends, by gender for blacks (African Americans) and whites in US geographic areas with high-quality cancer registries. The areas also varied in a tobacco control index (TCI) for 1992-1993. METHODS: Age-standardized incidence rates (ASIRs) were analyzed for lung cancer diagnosed at age 20-44 years from 1973-1976 to 1997-2000 for blacks and whites by gender, using data from the National Cancer Institute's Surveillance, Epidemiology and End Results (SEER) Program. Recent trends (1989-1992 to 1997-2000) also were analyzed for each SEER area, which differed in the TCI. RESULTS: ASIRs declined for black and white men from 1973-1976 to 1997-2000, but not for black or white women after 1985-1988; the gender ratio (men/women) declined to reach 1.0 in whites and 1.4 in blacks. ASIRs decline from 1989-1992 to 1997-2000 for men in all but one of the SEER areas examined, but declines for women were largely limited to SEER areas in California, a state with a high (but not the highest) TCI. Black-white disparities in ASIRs persisted for all SEER areas combined and in each of the areas examined, and increased for women in the Detroit area. CONCLUSIONS: Continued surveillance of ASIRs in young adults is needed, but these data emphasize the need for tobacco control programs to include targeting women and blacks. PMID- 15280625 TI - Impact of body mass index on the risk of total cancer incidence and mortality among middle-aged Japanese: data from a large-scale population-based cohort study -the JPHC study. AB - OBJECTIVES: To determine whether low or high extremes of body mass index (BMI) in otherwise healthy individuals affect mortality only after they develop cancer or affect the likelihood that cancer will occur. METHODS: We conducted a cohort analysis on the possible association between BMI and the risk of total cancer incidence and mortality among a middle-aged Japanese population consisting of a population-based cohort of 88,927 subjects (42,093 men and 46,834 women) with a 10-year follow-up. RESULTS: In men, a U-shaped association between BMI and cancer occurrence was observed, with men with a BMI of 23.0-24.9 having the lowest risk of cancer occurrence (BMI 14.0-18.9: HR = 1.29, 95% CI = 1.08-1.54; BMI 30.0 39.9: HR = 1.22, 95% CI = 0.92-1.61). This tendency did not change substantially after excluding cases diagnosed early during the follow-up period; cancer mortality showed a similar trend but with higher risk values. When analyzed according to smoking category, a low BMI affected cancer occurrence more strongly among current smokers than in never-smokers. Unlike men, no marked fluctuation in risk was observed in women. CONCLUSIONS: A very low BMI seems to have an impact on the total cancer risk in populations with a low average BMI. Therefore, while much attention has been given to the effects of obesity, the health effects of both extreme ends of BMI should be taken into consideration in populations with a low average BMI. PMID- 15280626 TI - A comparison of the sources of cancer mortality in China. AB - OBJECTIVE: To compare the validity of mortality data from available sources in China. MATERIALS, METHODS: Two large-scale surveys have provided accurate national-level rates; the most recent involved deaths occurring in a random 10% sample of the population during 1990-1992. Since then, the only readily available sources are two on-going surveillance systems, which provide annual estimates of mortality--the "Disease Surveillance Points" (DSP) sample survey, and that established by the Center of Health Information and Statistics (CHIS) of the ministry of health, the results of which are published by WHO. They were compared with respect to the representativeness of the populations covered and the rates obtained. RESULTS: Neither source covers a random sample of the Chinese population, with respect to age group, sex, and urban-rural residence, although the DSP population is the more representative of the national population in this respect. Sex and region (urban/rural) specific age-standardized mortality rates from the CHIS dataset were, however, closer to those from the (1990-1992) national survey, than those calculated from DSP data. CONCLUSIONS: The CHIS data is the preferred source for estimation of national mortality, and study of time trends, but requires appropriate weighting (by age, sex, rural/urban residence). The within-stratum estimates are more stable than those of DSP, because of its larger sample size. PMID- 15280627 TI - Association of the California tobacco control program with declines in lung cancer incidence. AB - OBJECTIVE: The California tobacco control program enacted in 1988 has been associated with declines in smoking and heart disease mortality. Since smoking also causes lung cancer, we investigated whether the program was associated with a decline in lung and other cancer incidence. METHODS: Age-adjusted incidence rates of lung and bladder cancer (which are caused by smoking) and prostate and brain cancer (which are not) in the San Francisco-Oakland (SFO) Surveillance Epidemiology End Results (SEER) registry and other eight SEER registries from 1975 to 1999 were fitted in multiple regression analyses accounting for the time lag between program implementation and its effects on cancer incidence. Cigarette consumption over time was also analyzed and related to lung cancer incidence. RESULTS: With a one year lag, the incidence of lung cancer in SFO, relative to eight other SEER registries, fell significantly below that predicted from the pre 1990 rates, by -0.981 (cases/100,000/year)/year (p = 0.001). With a three year lag, the incidence of bladder cancer fell by -0.234 (cases/100,000/year)/year (p = 0.066). No association of the program was observed on prostate or brain cancers in SFO. During the first decade, the Program was associated with about a 6% reduction in lung cancer incidence; state-wide that corresponds to about 11,000 cases avoided. CONCLUSION: A comprehensive tobacco control program is associated with a lower incidence of lung cancer. PMID- 15280628 TI - Mortality among workers employed in the titanium dioxide production industry in Europe. AB - OBJECTIVES: To assess the risk of lung cancer mortality related to occupational exposure to titanium dioxide (TiO2). METHODS: A mortality follow-up study of 15,017 workers (14,331 men) employed in 11 factories producing TiO2 in Europe. Exposure to TiO2 dust was reconstructed for each occupational title; exposure estimates were linked with the occupational history. Observed mortality was compared with national rates, and internal comparisons were based on multivariate Cox regression analysis. RESULTS: The cohort contributed 371,067 person-years of observation (3.3% were lost to follow-up and 0.7% emigrated). 2652 cohort members died during the follow-up, yielding standardized mortality ratios (SMRs) of 0.87 (95% confidence interval [CI] 0.83-0.90) among men and 0.58 (95% CI 0.40-0.82) among women. Among men, the SMR of lung cancer was significantly increased (1.23, 95% CI 1.10-1.38); however, mortality from lung cancer did not increase with duration of employment or estimated cumulative exposure to TiO2 dust. Data on smoking were available for over one third of cohort members. In three countries, the prevalence of smokers was higher among cohort members compared to the national populations. CONCLUSIONS: The results of the study do not suggest a carcinogenic effect of TiO2 dust on the human lung. PMID- 15280629 TI - Implications of oxidative stress and cell membrane lipid peroxidation in human cancer (Spain). AB - Reactive Oxygen Species (ROS) result from cell metabolism as well as from extracellular processes. ROS exert some functions necessary for cell homeostasis maintenance. When produced in excess they play a role in the causation of cancer. ROS mediated lipid peroxides are of critical importance because they participate in chain reactions that amplify damage to biomolecules including DNA. DNA attack gives rise to mutations that may involve tumor suppressor genes or oncogenes, and this is an oncogenic mechanism. On the other hand, ROS production is a mechanism shared by many chemotherapeutic drugs due to their implication in apoptosis control. The ROS mediated cell responses depend on the duration and intensity of the cells exposing to the increased ROS environment. Thus the status redox is of great importance for oncogenetic process activation and it is also implicated in tumor susceptibility to specific chemotherapeutic drugs. Phospholipid Hydroperoxide Glutathione Peroxidase (PH-GPx) is an antioxidant enzyme that is able to directly reduce lipid peroxides even when they are bound to cellular membranes. This article will review the relevance of oxidative stress, particularly of lipid peroxidation, in cell response with special focus in carcinogenesis and cancer therapy that suggests PH-GPx as a potentially important enzyme involved in the control of this processes. PMID- 15280630 TI - Stage of breast cancer diagnosis among medically underserved women in California receiving mammography through a state screening program. AB - OBJECTIVE: This study describes breast cancer stage at diagnosis among California women receiving mammograms through a state-administered screening program in comparison to other California women. METHOD: Linked data from California administered screening programs and the California Cancer Registry were used to identify participants diagnosed with breast cancer between 1994 and 2000. Logistic regression was used to compare the adjusted likelihood of late stage disease among program participants (categorized into four subgroups based on the timing and frequency of mammograms) to non-participants in California diagnosed during the same time period. RESULTS: Program participants were significantly more likely than non-participants to be diagnosed at late stage (adjusted OR 1.2; 95% CI 1.1, 1.3), with the highest risk occurring among those diagnosed 0-1 months after initial mammogram (adjusted OR 1.8; 95% CI 1.6, 2.1). The stage distribution among regularly screened participants was similar to non participants (adjusted OR of late stage disease 0.9; 95% CI 0.7, 1.1). CONCLUSIONS: Although program participants were more likely to be diagnosed at late stage than non-participants, their stage distribution was distinctly different according to their pattern of mammography utilization. This likely reflects differential utilization of program diagnostic and screening services, which should be taken into account in program evaluation. PMID- 15280631 TI - Attenuation of social class and reproductive risk factor associations for Hodgkin lymphoma due to selection bias in controls. AB - OBJECTIVE: Hodgkin lymphoma (HL) risk has been linked with higher social class and lower parity, but our prior population-based case-control study in adult women had unexpected null findings for these variables. Because subject participation was 87% for cases but 65% for random digit-dialing (RDD) controls, we examined representativeness of our controls and the impact of detected bias on prior results. METHODS: Using data from RDD enumeration, abbreviated interviews with nonparticipating controls, and the US census, we compared participating and nonparticipating RDD controls across several age groups and then recomputed odds ratios for risk factor associations adjusted for bias. RESULTS: The 325 RDD control participants were younger, more likely to be white, better educated, and of lower birth order and lower parity than the nonparticipants. Adjustment of odds ratios for bias strengthened previously null findings for education and for parity, breast-feeding and miscarriages in young adult women; these latter changes eliminated previously apparent age modification of risks. CONCLUSIONS: Selection bias in female RDD controls resulted from differential participation by socioeconomic factors, varied with age, and produced underestimations of several associations in young women, including reproductive factors. Thus, our prior conclusions of etiologic irrelevance for some study variables may have been inaccurate. PMID- 15280632 TI - Antihypertensive drug use and the risk of prostate cancer (Canada). AB - PURPOSE: To verify if exposure to antihypertensive drugs was associated to prostate cancer (PC) risk. METHODS: We conducted a matched case-control study using record linkage between two population-based databases. We defined exposure as a binary variable and in terms of timing and cumulative duration of use. We controlled for detection bias and Aspirin use. RESULTS: Among the 2221 cases and 11,105 controls, use of any antihypertensive agent was associated with an adjusted relative risk of PC of 0.98 (CI, 0.88-1.08). Of the different classes of antihypertensives, only beta-blockers (BBs) were associated with a reduction in PC risk (OR = 0.86, CI = 0.77-0.96). In those who cumulated < 1, 1-4, and > or = 4 years of BB use, the risk was 0.89 (0.75-1.05), 0.91 (0.75-1.09), and 0.82 (0.69-0.96), respectively. Also, subjects with > or = 4 years of alpha-blocker (ABs) use had a non-significant 25% reduction in PC risk. CONCLUSIONS: Our results suggest that BBs and long-term use of ABs may prevent PC whereas calcium channel blockers or angiotensin-converting enzyme inhibitors do not influence PC risk. PMID- 15280633 TI - Mammography screening and breast cancer mortality in New South Wales, Australia. AB - OBJECTIVE: To investigate the relationship between utilisation of service mammography screening and breast cancer mortality in New South Wales (NSW) women. Setting : Population-based biennial mammography screening was progressively introduced in NSW from 1988, with active recruitment and re-invitation for women aged 50-69 years, and reached full geographic coverage by 1996. Biennial mammography screening participation has varied widely over time and by municipality. METHODS: Breast cancer mortality by age, period and municipality was obtained from the NSW Central Cancer Registry. Biennial mammography screening rates for the same strata were obtained from the BreastScreen NSW database. Temporal changes in breast cancer mortality for NSW were summarised as annual average declines using Poisson regression. Breast cancer mortality for 1997-2001 was examined in relation to lagged biennial screening rates by municipality, adjusted for age, area socio-economic and geographic indicators, and breast cancer incidence, also using Poisson regression. RESULTS: For the 50-69 year age group, the mean annual breast cancer mortality decline was 0.8% (not significant) for 1988-1994, and 4.4% (p < 0.0001) for 1995-2001. Statistically significant negative associations between breast cancer mortality in 1997-2001 and lagged biennial screening rates were found with the highest significance at a four-year lag for women aged 50-69 years ( p = 0.0003) and also for women aged 50-79 years (p c = 0.0002). From the regression coefficient, a 70% biennial screening rate is associated with 32% lower breast cancer mortality (compared to zero screening). CONCLUSIONS: The effect of population-based mammography screening on breast cancer mortality in NSW inferred using this method is consistent with results of trials and other service studies. This suggests that population-based mammography screening programs can achieve significant reductions in breast cancer mortality with adequate participation. PMID- 15280634 TI - Excess concordance of cancer incidence and lifestyles in married couples (Japan): survival analysis of paired rate data. AB - OBJECTIVE: A population-based cohort study was conducted to examine concordance of cancer incidence and lifestyle in married couples. METHODS: Cancer incidence from 1 September 1986 to 30 June 2000 was followed for a baseline cohort of 2601 community-living married couples aged 40-84 years who were cancer-free and had completed an epidemiologic questionnaire in 1986 in Saitama, Japan. We computed age-adjusted cross-ratios: spouse-with/spouse-without rate ratios as a measure of association for paired rate data, using a bivariate survival analysis method. RESULTS: Concordance of lifestyles for couples was high for dietary habits, and low for smoking and alcohol habits. During the 14 year follow-up, 464 cancer cases occurred (husbands only: 279, wives only: 119, both: 33 couples). Wives whose husbands developed cancer were at an increased risk of developing cancer themselves, compared with those whose husbands had not had cancer (CR: 1.70, 95% CI: 1.12-2.58). Mean age at diagnosis was 68.2 years for husbands and 66.0 years for wives. And correlation of cancer incidence was further seen among couples who shared habits of smoking and/or drinking. CONCLUSIONS: Our data suggest that shared lifestyles including smoking and drinking habits are associated with an excess concordance of cancer incidence among married couples. PMID- 15280635 TI - Maternal dietary risk factors in childhood acute lymphoblastic leukemia (United States). AB - OBJECTIVE: Acute lymphoblastic leukemia (ALL) is the most common childhood cancer, and the second most common cause of mortality in children aged 1-14 years. Recent research has established that the disease can originate in utero, and thus maternal diet may be an important risk factor for ALL. METHODS: The Northern California Childhood Leukemia Study is a population-based case-control study of risk factors for childhood leukemia, including maternal diet. Cases (n = 138) and controls (n = 138) were matched on sex, date of birth, mother's race, Hispanicity, and county of residence at birth. Maternal dietary intake in the 12 months prior to pregnancy was obtained by a 76-item food frequency questionnaire. RESULTS: Consumption of the vegetables (OR = 0.53; 95% CI, 0.33-0.85; p = 0.008), protein sources (OR = 0.40; 95% CI, 0.18-0.90, p = 0.03), and fruits (OR = 0.71; 95% CI, 0.49-1.04; p = 0.08) food groups were inversely associated with ALL. Among nutrients, consumption of provitamin A carotenoids (OR = 0.65, 95% CI, 0.42 1.01; p = 0.05), and the antioxidant glutathione (OR = 0.42; 95% CI, 0.16-1.10; p = 0.08) were inversely associated with ALL. CONCLUSION: Maternal dietary factors, specifically the consumption of vegetables, fruits, protein sources and related nutrients, may play a role in the etiology of ALL. Dietary carotenoids and glutathione appear to be important contributors to this effect. PMID- 15280636 TI - Alcohol drinking may increase risk of breast cancer in men: a European population based case-control study. AB - OBJECTIVE: It has been estimated that alcohol drinking increases the risk of breast cancer in women by approximately 7% for each increment of 10 g alcohol per day. However, the few studies conducted on breast cancer among men have failed to detect an association with quantitative measures of alcohol drinking, even if the alcohol intake is generally higher in men than in women. On the other hand, increased risks of male breast cancer were inconsistently reported in alcoholics or patients with liver cirrhosis. We have investigated the role of alcohol drinking in male breast cancer using data collected in a population-based case control study on seven rare cancers, conducted in Denmark, France, Germany, Italy, and Sweden. METHODS: The cases were 74 histologically verified male breast cancer patients aged 35-70 years. The controls (n = 1432) were selected from population registers, and frequency-matched to the cases by age group and geographic area. To check for consistency, a separate analysis was conducted using as controls the patients with a rare cancer other than male breast recruited simultaneously in the European study (n = 519 men). RESULTS: Based on population controls, the risk of developing breast cancer in men increased by 16% (95% CI: 7-26%) per 10 g alcohol /day (p < 0.001). An odds ratio of 5.89 (95% CI: 2.21-15.69) was observed for alcohol intake greater than 90 g per day, as compared with light consumers (< 15 g per day). Similar associations were observed when other rare cancers patients were used as controls. CONCLUSION: We found that the relative risk of breast cancer in men is comparable to that in women for alcohol intakes below 60 g per day. It continues to increase at high consumption levels not usually studied in women. PMID- 15280637 TI - Body mass index and risk of colorectal cancer in women (United States). AB - OBJECTIVE: Epidemiologic data relating obesity to risk of colorectal cancer in women have been inconclusive. Two recent studies have suggested that the association may be modified by estrogen status; BMI was positively associated with colorectal cancer risk among women with high estrogen exposures [premenopausal women, and postmenopausal women who currently received postmenopausal hormone therapy (PMH)]. We prospectively investigated the role of BMI in colorectal cancer risk along with the modifying effects of estrogen in a large cohort from the Women's Health Study. METHODS: Among 39,876 apparently healthy women aged > or = 45 years at baseline (54% of them were postmenopausal), 37,671 were eligible for the present study. During an average of 8.7 years of follow-up, 202 women had a confirmed diagnosis of colorectal cancer. Baseline BMI was calculated by dividing self-reported weight in kilograms by height in meters squared. RESULTS: The multivariate relative risks (RRs) and 95% confidence interval (CI) of colorectal cancer were 1.72 (1.12-2.66) for 27-29.9 kg/m2, and 1.67 (1.08-2.59) for > or = 30 kg/m2, as compared with BMI < 23 kg/m2 ( p for trend = 0.02). This positive association was seen primarily in the proximal colon ( p for trend = 0.004). When the association was further examined according to PMH use among postmenopausal women, we found that both current and never users with higher BMI were at a greater risk of colorectal cancer ( p for interaction between BMI and PMH use = 0.33). As compared with BMI < 23 kg/m2, the multivariate RRs and 95% CI for 27-29.9 and > or = 30 kg/m2 were 1.98 (0.98-3.99) and 1.41 (0.65-3.06) among current users, and 1.05 (0.42-2.65) and 2.91 (1.40 6.06) among never users. CONCLUSIONS: These data suggest that higher BMI was associated with an elevated risk of colorectal cancer, and the positive relationship was not altered by estrogen exposure among postmenopausal women. PMID- 15280638 TI - Breast cancer risk and the combined effect of environmental estrogens. AB - OBJECTIVE: The present study aimed to determine whether the combined effects of environmental estrogens measured as the total effective xenoestrogen burden (TEXB alpha) are a risk factor for breast cancer over and above the risk potentially linked to specific pesticides. METHODS: We measured the levels of 16 organochlorine pesticides as well as TEXB in adipose tissue of 198 women at the time of breast cancer diagnosis. These were compared with findings in 260 age and hospital matched control women without breast cancer. RESULTS: The median levels of p,p'-DDE (1,1-dichloro-2,2-bis( p -chlorophenyl)ethylene), aldrin, endosulfan ether and lindane (the pesticides detected in > 40% of the study population) were higher in cases than controls, although the differences did not reach statistical significance. After adjusting for potential confounders, the odds ratio (OR) for breast cancer in women with detectable levels of aldrin was 1.55 (95% confidence interval (CI) 1.00-2.40). Among the postmenopausal women, the OR for aldrin and lindane was 1.84 (95% CI 1.06-3.18) and 1.76 (95% CI 1.04-2.98), respectively. Among cases with body mass index (BMI) below the median (28.6 kg/m2), the OR was 3.42 (95% CI 1.22-9.58) for women in the highest quartile of TEXB-alpha versus those in the lowest. The subgroup of leaner postmenopausal women showed an increased risk (OR: 5.67; 95% CI 1.59-20.21) for those in the highest tertile versus those in the lowest. CONCLUSIONS: We found an increased risk for breast cancer in the leaner women, especially in the leaner postmenopausal subgroup, related to the TEXB-alpha. The pesticides aldrin and lindane are also individually associated with risk. PMID- 15280639 TI - Racial differences in oral and pharyngeal cancer treatment and survival in Florida. AB - OBJECTIVE: This study examined racial differences in treatment and survival for blacks and whites in Florida diagnosed with oral or pharyngeal cancer. METHODS: Data for 21,481 malignancies of the oral cavity or pharynx diagnosed from 1988 to 1998 were derived from the Florida Cancer Data System. Type of cancer treatment was compared by race, stratified by anatomic site and summary stage at diagnosis. Kaplan-Meier survival curves were used to compare survival rates and Cox regression models were used to estimate hazard ratios for race. Covariates included age, sex, census tract income, and treatment. RESULTS: Stratifying by tumor site and stage, blacks consistently had poorer survival rates than whites. Across tumor stages, blacks with oral cavity cancer were consistently more likely than whites to have received only radiotherapy and less likely to have received cancer-directed surgery. Trends were similar for pharyngeal cancer, although statistically significant only for regional stages. Across site and stage, blacks consistently had elevated hazard ratios (range: 1.20-1.53) relative to whites. CONCLUSIONS: In Florida, there were racial differences in patient treatment for oral or pharyngeal cancer. Blacks had lower survival rates than whites, but differences in treatment did not entirely account for racial disparities in survival. PMID- 15280640 TI - Estimating regional variation in cancer survival: a tool for improving cancer care. AB - OBJECTIVE: To improve estimation of regional variation in cancer survival and identify cancers to which priority might be given to increase survival. METHODS: Survival measures were calculated for 25 major cancer types diagnosed in each of 17 health service regions in New South Wales, Australia, from 1991 to 1998. Region-specific risks of excess death due to cancer were estimated adjusting for age, sex, and extent of disease at, and years since, diagnosis. Empirical Bayes (EB) methods were used to shrink the estimates. The additional numbers of patients who would survive beyond five years were estimated by shifting the State average risk to the 20th centile. RESULTS: Statistically significant regional variation in the shrunken estimates of risk of excess death was found for nine of the 25 cancer types. The lives of 2903 people (6.4%) out of the 45,047 whose deaths within 5 years were attributable to cancer could be extended with the highest number being for lung cancer (791). CONCLUSIONS: The EB approach gives more precise estimates of region-specific risk of excess death and is preferable to standard methods for identifying cancer sites where gains in survival might be made. The estimated number of lives that could be extended can assist health authorities in prioritising investigation of and attention to causes of regional variation in survival. PMID- 15280641 TI - Incidence of lip cancer in the male Norwegian agricultural population. AB - OBJECTIVE: To explore lip cancer (LC) associations with work environmental exposures in a record-linkage study of Norwegian farmers. We hypothesize immunosuppressive substances (e.g. mycotoxins, pesticides) to influence LC incidence. METHODS: A cohort of 131,243 male Norwegian farmers born 1925-1971 was established by cross-linkage of national registers and followed up through 1999 for incident LC, (ICD-7 site 140) in the Cancer Registry of Norway. Farm production data from agricultural censuses 1969-1979 and meteorological data on solar radiation and fungal forecasts (events of wet and temperate conditions known to favour fungal growth and mycotoxin formation) served as exposure proxies. Adjusted rate ratios (RR) and 95% confidence intervals (CI) were estimated using Poisson regression. RESULTS: We identified 108 LC cases (rate 4.4 per 100,000 person-years). We found LC to be moderately associated with horses on the farm (RR = 1.6, CI = 1.0-2.4), construction work employment (RR = 1.7, CI = 1.1-2.6), pesticide use (RR = 0.7, CI = 0.4-1.0), grain production (RR = 1.3, CI = 0.9-2.1) and increasing levels of fungal forecasts (RR = 1.6, CI = 0.9-2.8 in the highest two quartiles). CONCLUSION: Moderate associations of LC with grain production and fungal forecasts and the negative association with pesticide could possibly be explained by exposure to immunosuppressive mycotoxins. Some of the associations observed could be explained by solar exposure. PMID- 15280642 TI - Estrogen receptor beta (ESR2 ) polymorphisms and endometrial cancer (United States). AB - OBJECTIVE: We hypothesized that variations in the ESR2 gene may influence estrogen exposure in the uterus and thus influence endometrial cancer risk. We validated and screened for variants in the ESR2 gene and examined whether they are associated with endometrial cancer risk. METHODS: We resequenced the promoter and coding regions of the ESR2 gene in 24 endometrial cancer cases, and genotyped the validated/discovered SNPs and intronic dinucleotide CA repeat in a nested case-control study of endometrial cancer (cases = 222, controls = 666) in the Nurses' Health Study (NHS). We also explored statistical interaction between ESR2 genotypes and body mass index (BMI) or hormone replacement therapy (HRT) use among postmenopausal women and cancer risk. RESULTS: Two SNPs were validated [rs1256049 in exon 5 (allelic frequencies = 98% G, 2% A) and rs1271572 in the promoter region (allelic frequencies = 60% G, 40% T)]. After adjusting for potential confounders, we observed no association between ESR2 gene polymorphisms and endometrial cancer risk [rs1256049 (OR = 1.2; 95%CI: 0.7-2.3), rs1271572 (OR = 0.8; 95%CI: 0.5-1.1) and CA repeat (22 repeat allele versus > or = 22 repeat allele, OR = 1.1; 95%CI: 0.7-1.7)]. We also did not observe any significant effect modification of the ESR2 polymorphisms by BMI or HRT use among postmenopausal women. CONCLUSION: Our results indicate that ESR2 polymorphisms may not be associated with endometrial cancer risk. PMID- 15280643 TI - Estimation of the size of the vCJD epidemic. AB - Estimates of the final size of the variant Creuzfeldt-Jakob Disease epidemic have been made by fitting theoretical curves of the incubation period distribution to the histogram of observed annual deaths to 2002, using various assumptions of the mean and standard deviation of this distribution, and also of the efficacy of the Specified Bovine Offals ban of 1989. Unless the mean incubation time is greater than 15 to 20 years the estimates lie in the low hundreds to about a thousand, and the most likely situation, of a mean between 11 and 15 years, gives estimates of about 150 to 500 deaths. Numbers above a few thousands would only occur if the mean incubation period is of the order of 25 to 30 years and reasons are adduced to indicate this is very unlikely. These numbers are not greatly increased if the ban was poorly observed. This method of analysis may be applicable to other situations where a cause that is limited in space and time is expected to have late effects. PMID- 15280644 TI - Candida galli sp. nov., a new yeast from poultry. AB - Six strains of an unknown yeast species, phenotypically resembling Yarrowia lipolytica and isolated from chicken breast and chicken liver, were studied. The investigation of their small (18S) and large (26S) subunit rDNA revealed a robust genetic difference between these strains and the type strain of Y. lipolytica. A consistent difference in the physiological properties, suitable for separation of the two taxa, was also found. The description of the new anamorphic yeast species, Candida galli is given. PMID- 15280645 TI - Cloning and sequence analysis of the glyceraldehyde-3-phosphate dehydrogenase gene from the zygomycetes fungus Rhizomucor miehei. AB - Rhizomucor miehei is important from a biotechnological aspect in consequence of its content of aspartic proteinase, which has high milk-clotting activity. A genomic library of R. miehei NRRL 5901 has been constructed in a phage (Lambda Fix II) vector. The glyceraldehyde-3-phosphate dehydrogenase (gpd) gene was isolated from this library by hybridization of the recombinant phage clones with a gpd-specific gene probe generated by polymerase chain reaction. The complete nucleotide sequence encodes a putative polypeptide chain of 336 amino acids interrupted by 5 introns. The predicted amino acid sequence of this gene shows a high degree of sequence similarity to the glyceraldehyde-3-phosphate dehydrogenase proteins from yeast and filamentous fungi. The promoter region, containing a consensus TATA box, and 246-bp downstream from the putative stop codon were also determined. The possibility of using the gpd promoter in the construction of new transformation vectors is discussed. PMID- 15280646 TI - Characterization of the AINV gene and the encoded invertase from the dimorphic yeast Arxula adeninivorans. AB - The invertase-encoding of AINV gene Arxula adeninivorans was isolated and characterized. The gene includes a coding sequence of 2700 bp encoding a putative 899 amino acid protein of 101.7 kDa. The identity of the gene was confirmed by a high degree of homology of the derived amino acid sequence to that of alpha glucosidases from different sources. The gene activity is regulated by carbon source. In media supplemented with sucrose induction of the AINV gene and accumulation of the encoded invertase in the medium was observed. In addition the extracellular enzyme level is influenced by the morphological status of the organism, with mycelia secreting the enzyme in titres higher than those observed in budding yeasts. The enzyme characteristics were analysed from isolates of native strains as well as from those of recombinant strains expressing the AINV gene under control of the strong A. adeninivorans -derived TEF1 promoter. For both proteins a molecular mass of 600 kDa was determined, a pH optimum at pH 4.5 and a temperature optimum at 55 degrees C. The preferred substrates for the enzyme included the ss-D-fructofuranosides sucrose, inulin and raffinose. Only a weak enzyme activity was observed for the alpha-D-glucopyranosides maltotriose, maltose and isomaltose. Thus the invertase primarily is a ss-fructosidase and not an alpha-glucosidase as suggested by the homology to such enzymes. PMID- 15280647 TI - Bioinformatic analysis of the link between gene composition and expressivity in Saccharomyces cerevisiae and Schizosaccharomyces pombe. AB - The compositional non-randomness was studied in genes of Saccharomyces cerevisiae and Schizosaccharomyces pombe. In both species, codon usage is well correlated with expressivity (measured as the codon adaptation index). Both species generally display higher nucleotide non-randomness in the group of highly expressed genes than in the lowly expressed genes. The highly expressed genes in both species are furthermore characterized by marked peaks in non-randomness at N=3 upstream of start codons, N=2 downstream of start codons and at N=1 and N=7 downstream of stop codons, indicating that these nucleotides may be key elements in translational regulation. Intragenic variation in codon usage was also observed to be linked to expressivity. It is suggested that the firm link between expressivity and codon usage calls for codon optimization. Based on bioinformatic calculations, examples of proteins are given for which codon optimizations might be relevant. PMID- 15280648 TI - Nucleotides downstream of start codons show marked non-randomness in Escherichia coli but not in Bacillus subtilis. AB - This study aimed at measuring the nucleotide non-randomness in the region downstream of start codons in bacterial genes and to see if the non-randomness differs between biased and unbiased genes, in terms of the effective number of codons (Nc) and the codon adaptation index (CAI). In Escherichia coli, there was a marked elevation in nucleotide conservation for the genes having low Nc-values compared to the genes having high Nc-values, i.e the more biased genes showed a higher level of non-randomness. Likewise, the genes displaying high CAI-values showed stronger nucleotide conservation than the genes of low CAI-values. This elevated conservation is visible up to approximately 15-17 nucleotides downstream of the start codon, after which there is little difference. This indicates that there may be distinct selectional mechanisms acting upon the first 5-6 codons within genes in E. coli. In B. subtilis, these effects are less pronounced, if present at all. Furthermore, analyses of codons used in this region were not in support of the hypothesis that the elevation in nucleotide non-randomness is a question of selection for certain optimal codons. PMID- 15280649 TI - Streptacidiphilus jiangxiensis sp. nov., a novel actinomycete isolated from acidic rhizosphere soil in China. AB - The taxonomic position of three acidophilic actinomycetes isolated from acidic rhizosphere soil was established using a polyphasic approach. The morphological and chemical properties of the isolates were found to be consistent with their assignment to the genus Streptacidiphilus. Almost complete 16S rRNA gene sequences determined for the strains were aligned with corresponding sequences of representatives of the genera Kitasatospora, Streptacidiphilus and Streptomyces and phylogenetic trees inferred using three tree-making algorithms. The organisms formed a distinct subclade within the Streptacidiphilus 16S rRNA gene tree. They also shared nearly identical phenotypic profiles and rep-PCR fingerprint patterns that readily distinguished them from representatives of the established species of Streptacidiphilus. It is evident from the genotypic and phenotypic data that the three isolates form a new species in the genus Streptacidiphilus. The name proposed for this new species is Streptacidiphilus jiangxiensis, the type strain is isolate 33214T (= AS 4.1857T = JCM 12277T). PMID- 15280650 TI - The gene encoding the ribulose monophosphate pathway enzyme, 3-hexulose-6 phosphate synthase, from Aminomonas aminovorus C2A1 is adjacent to coding sequences that exhibit similarity to histidine biosynthesis enzymes. AB - In an attempt to understand better the organisation of genes encoding enzymes of the ribulose monophosphate pathway (RuMP), the 3-hexulose 6-phosphate synthase gene (hps) and flanking sequences were cloned from the obligate methylotroph Aminomonas aminovorus C2A1. To date only three hps containing gene clusters from methylotrophs have been characterised and these contain genes encoding other RuMP enzymes. However, hps from A. aminovorus C2A1 was shown to be adjacent to coding sequences for products with sequence similarity to histidine biosynthesis enzymes. Furthermore, none of the hps homologue containing gene clusters, from genome sequences previously analysed or analysed in this paper, were similar in organisation to that of A. aminovorus C2A1. PMID- 15280651 TI - Penicillium persicinum, a new griseofulvin, chrysogine and roquefortine C producing species from Qinghai Province, China. AB - A unique Penicillium isolate from Chinese soil with terverticillate penicilli and ellipsoidal to cylindrical smooth-walled conidia, produces, in addition to the common metabolite ergosterol, copious amounts of an unknown peach-red pigment and the following secondary metabolites: griseofulvin, dechlorogriseofulvin, lichexanthone, roquefortine C, roquefortine D, chrysogine, 2 pyrovoylaminobenzamide, 2-acetyl-quinazolin-4(3H)-one. This isolate, CBS 111235, is described as Penicillium persicinum sp. nov., which belongs to subgenus Penicillium section Chrysogena but is morphologically similar to P. italicum. On the basis of the production of secondary metabolites it resembles P. griseofulvum and P. coprophilum. Sequence data using part of the beta-tubulin gene showed that it is phylogenetically related to P. chrysogenum and P. aethiopicum in section Chrysogena with which it shares both secondary metabolites and ability to grow at 37 degrees C. PMID- 15280652 TI - Genetic variability in the species Rhizopus stolonifer, assessed by random amplified polymorphic DNA analysis. AB - Rhizopus stolonifer is an important post-harvest pathogenic fungus. Recent taxonomic findings based on morphological and growth characteristics led to a dramatic reduction in the number of accepted species within the genus. The aim of this study was to examine this situation with molecular markers. Twenty-nine R. stolonifer strains isolated from various locations and substrates were characterized by random amplified polymorphic DNA (RAPD) analysis. The numerical analysis of the RAPD data revealed four main clusters with extremely high dissimilarity values, but only low or moderate variability was observed within these groups. These results suggest a high genetic heterogeneity in the case of R. stolonifer: isolates of R. stolonifer var. stolonifer, R. stolonifer var. reflexus and R. niveus displayed species-level genetic distances, which gives rise to considerations that they might be separate species. PMID- 15280653 TI - Relationships between Escherichia coli cells and the surrounding medium during survival processes. AB - In Escherichia coli, during survival under adverse conditions, namely starvation and luminous radiation, two things occur. On the one hand organic substances are released into the surrounding medium and on the other there is a transition from the culturable state to viable but non-culturable (VBNC). An analysis of organic molecules released into the surrounding medium showed the presence of proteins, dissolved free amino acids, and dissolved monomeric carbohydrates. The concentration of these substances in the medium changed with exposure time, type of stress and type of molecule. The proteins accumulated in the medium and in some cases their identification revealed the presence of components of the outer membrane. Variations in the concentration of amino acids and carbohydrates point to a twofold process of excretion and uptake. Indeed, cell free supernatants supported the growth of several generations of a population of 10(4) cells ml( 1). The survival of E. coli in supernatants previously colonized by cells in the VBNC state was greater than that observed in the control experiments, with a short delay in the loss of culturability. It was thus clear that organic molecules released into the medium play a role in the transition from culturable to VBNC state. PMID- 15280654 TI - 15th International Chromosome Conference (ICC XV). September 5-10, 2004, London, United Kingdom. Abstracts. PMID- 15280655 TI - Elevated levels of thymidylate synthase linked to neoplastic transformation of mammalian cells. AB - Thymidylate synthase (TS), an enzyme that is essential for DNA synthesis and repair has been identified as an important biomarker for colorectal and other human cancers. The elevated steady-state levels of TS found in many common human malignancies have been thought to represent a secondary event in tumor formation. However, it has recently been demonstrated that the deregulated levels of ectopic TS may also have a causal effect on tumorgenesis since overexpression of human TS transforms immortalized mammalian cells to a malignant phenotype. Since the levels of TS are regulated by E2F-1 and thus are linked to the cell cycle pathway, regulating TS activity may be an important factor for the control of cell cycle progression and for the development of therapeutic strategies and cancer prevention. PMID- 15280656 TI - Mammalian DNA methyltransferase 1: inspiration for new directions. AB - The role of DNA methyltransferase 1, DNMT1, in human cancer cells has recently been contested. In this setting, DNMT1's function as the sole maintenance methyltransferase was based on the assumption that its biological activity is identical to the mouse homologue. However, the application of recent technological advances, including gene targeting and siRNA mediated ablation studies, has cast doubt on this presumed role. Here, we attempt to shed light on these new data within the context of previous experiments. PMID- 15280657 TI - Actvation of NF-kappaB by the API2/MALT1 fusions inhibits p53 dependant but not FAS induced apoptosis: a directional link between NF-kappaB and p53. AB - Interactions between survival pathways and apoptotic cascades play a determinant role in the maintenance of neoplastic clone proliferation and impaired response to apoptosis. Recently, we established a novel interplay between the NF-kappaB survival- and p53 death-pathways in a tumor model system that represents the most common extranodal lymphoid cell neoplasia, MALT (Mucosa Associated Lymphoid Tissue) lymphoma. MALTs are genetically characterized by the t(11;18)(q21;q21) chromosomal translocation that results in API2/MALT1 fusion products. It was shown that distinct API2/MALT1 chimeric proteins function as oncogenes that bilaterally confer a proliferative advantage to the neoplastic clone by activating the NF-kappaB signaling pathway and also inhibiting p53 mediated cell death. Here, we demonstrate that API2/MALT1 mediated inhibition of apoptosis is p53 specific, as distinct API2/MALT1 fusion proteins fail to protect cells from FAS induced cell death. Furthermore, we demonstrate that API2/MALT1 mediated NF kappaB activation does not alter p53 protein levels or subcellular localization suggesting a post-translational or indirect mechanism of p53 deregulation. PMID- 15280658 TI - Is there a link between DNA polymerase beta and cancer? AB - Recent small-scale studies have shown that 30% of human tumors examined to date express DNA polymerase beta variant proteins. One of the DNA polymerase beta colon cancer-associated mutants, K289M, has been shown to synthesize DNA with a lower fidelity than wild-type Pol beta. Thus, the K289M protein could confer a mutator phenotype to the cell, resulting in genomic instability. Another DNA polymerase beta variant identified in colon carcinoma interferes with base excision repair in cells. This may result in unfilled gaps which can serve as substrates for recombination and result in genomic instability. DNA polymerase beta has also been shown to be overexpressed in a variety of tumors. In some cases, overexpression of polymerase beta in cells confers a transformed phenotype to the cells. In other cases, overexpression results in telomere fusions. Thus, mutant forms or aberrant quantities of polymerase beta confer a mutator phenotype to cells. Combined with the small-scale tumor studies, these mechanistic studies implicate variant forms of DNA polymerase beta in the etiology of human cancer. PMID- 15280659 TI - Effects of resveratrol on gene expression in renal cell carcinoma. AB - Studies have shown that Resveratrol (RE) can inhibit cancer initiation, promotion, and progression. However the gene expression profile in renal cell carcinoma (RCC) in response to RE treatment has never been reported. To understand the potential anticancer effect of RE on RCC at molecular level, we profiled and analyzed the expression of 2059 cancer-related genes in a RCC cell line RCC54 treated with RE. Biological functions of 633 genes were annotated based on biological process ontology and clustered into functional categories. Twenty-nine highly differentially expressed genes in RE treated RCC54 were identified and the potential implications of some gene expression alterations in RCC carcinogenesis were identified. RE was also shown to inhibit cell growth and induce cell death of RCC cells. The expression alterations of selected genes were validated using reverse transcription polymerase chain reaction. In addition, the gene expression profiles under different RE treatments were analyzed and visualized using singular value decomposition. The findings from this study support the hypothesis that RE induces differential expression of genes that are directly or indirectly related to the inhibition of RCC cell growth and induction of RCC cell death. In addition, it is apparent that the gene expression alterations due to RE treatment depend strongly on RE concentration. This study provides a general understanding of the overall genetic response of RCC54 to RE treatment and yields insights into the understanding of the cancer preventive mechanism of RE in RCC. PMID- 15280660 TI - Molecular functions of BRCA1 in the DNA damage response. AB - Ten years ago, a concerted effort from several labs resulted in the cloning of BRCA1, the first of two major hereditary breast/ovarian cancer predisposition genes. Since that time, BRCA1 has been linked to several key nuclear functions connected with the prevention of genomic instability. In particular, BRCA1 functions in concert with Rad51, BRCA2 and other genes to control double strand break repair (DSBR) and homologous recombination. Here, we reassess the role of BRCA1 and its associated proteins in this process. PMID- 15280661 TI - TGF-beta targets the Wnt pathway components, APC and beta-catenin, as Mv1Lu cells undergo cell cycle arrest. AB - The highly coordinated interaction of TGF-beta and Wnt signaling pathways is critical for normal development. However, the effects of TGF-beta on APC and beta catenin, two key mediators of Wnt signaling in epithelial cells, have been largely unknown. We determined the effect of TGF-beta on APC and beta-catenin expression in Mv1Lu, a nontransformed epithelial cell line, in which TGF-beta signaling causes a G(1) cell cycle arrest. We found that TGF-beta rapidly reduced APC protein levels through a post-transcriptional mechanism. Further, TGF-beta increased beta-catenin mRNA and protein levels, and increased beta-catenin nuclear accumulation. Finally, retrovirus-mediated overexpression of beta-catenin discernibly enhanced the ability of TGF-beta to induce a G(1) cell cycle arrest. This is the first report demonstrating that TGF-beta mimics the effect of Wnt signaling on beta-catenin in Mv1Lu cells, and that reduction of APC and nuclear accumulation of beta-catenin have cooperative effects on mechanisms that mediate TGF-beta-induced cell cycle arrest. PMID- 15280662 TI - Akt-dependent T198 phosphorylation of cyclin-dependent kinase inhibitor p27kip1 in breast cancer. AB - The localization of the cyclin-dependent kinase inhibitor p27(kip1) is dependent on the phosphorylation of one of three key amino acid residues: S10, T157 and T198. However, it was unclear whether endogenous p27(kip1) is phosphorylated at T198 in the living cell. In the present work we describe the generation and characterization of a polyclonal antibody able to recognize recombinant, transfected as well as endogenous T198-phosphorylated p27(kip1). Using this antibody, we demonstrate that: (1) endogenous p27(kip1) is phosphorylated at T198 in 4 breast cancer cells lines (MCF7, MDA-MB231, MDA- MB436 and MDA-MB468); (2) T198 phosphorylation is increased in breast cancer cells compared with normal mammary epithelial cells (HMEC); (3) T198-phosphorylated p27(kip1) is exclusively cytoplasmic; (4) T198 phosphorylation is dependent on the activity of the PI3K PKB/Akt pathway, being it drastically reduced by the pharmacological PI3K inhibitor LY294002 or stimulated by the constitutive activation of PKB/Akt. Finally, in primary human breast carcinomas, cytoplasmic accumulation of T198 phosphorylated p27(kip1) parallels Akt activation. We conclude that in breast cancer cells p27(kip1) is phosphorylated at T198 in a PI3K/Akt dependent manner and that this phosphorylation may contribute to p27(kip1) cytoplasmic mislocalization observed in breast cancer. PMID- 15280663 TI - Geminin: a major DNA replication safeguard in higher eukaryotes. AB - Eukaryotes have evolved multiple mechanisms to restrict DNA replication to once per cell cycle. These mechanisms prevent relicensing of origins of replication after initiation of DNA replication in S phase until the end of mitosis. Most of our knowledge of mechanisms controlling prereplication complex (preRC) formation are based on studies from yeast and Xenopus, while much less is known for mammalian cells. Here we discuss our recent data demonstrating that Geminin is required for preventing rereplication in human normal and cancer cells. PMID- 15280664 TI - Oxygen sensing by H+: implications for HIF and hypoxic cell memory. AB - Hypoxia and acidosis are common features of several physiological and pathological situations, including cancer and stroke. The HIF (hypoxia-inducible factor) transcription factor plays a seminal role in orchestrating cellular responses to alterations in oxygen availability. HIF is degraded in normal oxygen tension by the VHL (von Hippel-Lindau) tumor suppressor protein but stabilized by hypoxia to activate an array of genes implicated in oxygen homeostasis including vascular endothelial growth factor. Cells respond to a comparatively mild decline in oxygen tension by converting to an anaerobic state of respiration and secreting lactic acid. We recently reported that a decrease in environmental pH triggers sequestration of VHL into the nucleolus neutralizing its ability to degrade HIF. This implies that cells have evolved a parallel mechanism of HIF activation that responds to changes in oxygen levels by sensing extracellular [H+]. Here we discuss the implications of this new VHL regulatory mechanism on oxygen homeostasis and hypoxic cell memory. PMID- 15280665 TI - A novel function for human Rad9 protein as a transcriptional activator of gene expression. AB - The human Rad9 gene is evolutionarily conserved and plays important roles in several fundamental biological processes, including promotion of resistance to DNA damage, mediating cell cycle checkpoints, acting as a proapoptotic element, and maintaining genomic integrity in the presence or even in the absence of exogenous DNA damaging agents. Rad9 protein was recently found to have the capability of activating transcription of p21, a well-known p53 target gene and universal inhibitor of cyclins and CDKs. Rad9 can bind previously defined p53 consensus sequences in the promoter region of p21. Microarray gene expression screening coupled with northern analyses revealed that Rad9 activates transcription of multiple genes, a subset of which is also controlled by p53. In this review, the newly defined ability of Rad9 to transactivate p21 and other downstream target genes is discussed in terms of the activities already associated with this protein. PMID- 15280666 TI - Ubiquitination of PCNA and the polymerase switch in human cells. AB - Replicative DNA polymerases are blocked by damage in the template DNA. To get past this damage, the cell employs specialized translesion synthesis (TLS) polymerases, which have reduced stringency and are able to bypass different lesions. For example, DNA polymerase eta (poleta) is able to carry out TLS past UV-induced cyclobutane pyrimidine dimers. How does the cell bring about the switch from replicative to TLS polymerase? We have shown that, in human cells, when the replication machinery is blocked at DNA damage, PCNA, the sliding clamp required for DNA replication, is mono-ubiquitinated and that this modified form of PCNA has increased affinity for poleta. This provides a mechanism for the polymerase switch. In this Extra-View, we discuss the possible signals that might trigger ubiquitination of PCNA, whether PCNA becomes de-ubiquitinated after TLS has been accomplished and the role of the hREV1 protein in TLS. We point out some apparent differences between mechanisms in Saccharomyces cerevisiae and human cells. PMID- 15280667 TI - Interactions between BMP and Wnt signaling pathways in mammalian cancers. AB - The bone morphogenetic protein (BMP) and Wnt signaling pathways have been extensively studied in the regulation of early embryonic development and in the control of cell proliferation in adult tissues. Genetic interaction between these highly conserved and ubiquitous signaling pathways has been observed in multiple settings in fruit flies, amphibians, zebrafish, and mammals. While the importance of Wnt signaling in carcinogenesis has been well established, more recent work has also implicated BMP signaling in apoptosis and as a negative regulator of proliferation. In this issue of Cancer Biology & Therapy, Nishanian et al. extend these studies and propose interesting potential interactions between BMP and Wnt signaling in transformed mammalian cells that could have important implications for the control of human cancers. PMID- 15280668 TI - PI3K: Missense Mutation Motivates Malignancy. PMID- 15280669 TI - Finding and keeping your partner during meiosis. AB - HIM-3 is a meiosis-specific protein that localizes to the cores of chromosomes from the earliest stages of prophase I until the metaphase to anaphase I transition in Caenorhabditis elegans. him-3 mutations disrupt homolog alignment, synapsis, and recombination and we propose that the association of HIM-3 with chromosome axes is a critical event in meiotic chromosome morphogenesis that is required for the proper coordination of these processes. The presence of HIM-3 like proteins in other eukaryotes, some of which are known to be required for synapsis and recombination, suggests the existence of a conserved class of axis associated proteins that function at the junction of essential meiotic processes. PMID- 15280670 TI - The p53 paddy wagon: COP1, Pirh2 and MDM2 are found resisting apoptosis and growth arrest. AB - For years, the growth inhibitory effects of the tumor suppressor p53 were thought to be antagonized predominantly by the ubiquitin ligase, MDM2. It has long been established that MDM2 physically associates with p53 and targets this tumor suppressor for proteasomal degradation. In light of recent findings, it now appears that MDM2 may not be the only ubiquitin ligase that negatively controls p53 function. Two recently discovered proteins, Pirh2 and COP1, are also believed to facilitate p53 degradation via the ubiquitin-proteasome pathway. Both proteins are upregulated by p53 as well as genotoxic stress and each has been found to directly promote p53 ubiquitination and degradation. Future studies in this field will now face the challenge of elucidating the physiological significance of three molecules all apparently able to independently facilitate p53 degradation and abrogate its function. PMID- 15280671 TI - p53 mutants: the achilles' heel of human cancers? AB - Recent scientific discoveries have thrust mutants of the tumor suppressor protein p53 into the forefront of the war on cancer, and hold out eventual hope for a small molecule drug that will be useful in treating human cancers with mutant p53 protein. PMID- 15280672 TI - The current state of intestinal transplantation. PMID- 15280674 TI - Consensus opinion from the antibody working group on the diagnosis, reporting, and risk assessment for antibody-mediated rejection and desensitization protocols. AB - During the past few decades, much of the experimental and clinical effort in solid-organ transplantation has been directed toward ameliorating or abrogating T cell-mediated responses. As a result, universally understood and accepted nomenclature and diagnostic criteria have evolved. Humoral immunity in transplantation has yet to undergo a similar renaissance. Readers of transplant journals regularly find it difficult and often impossible to interpret data on the diagnosis and management of antibody-mediated rejection. The Antibody Working Group was assembled in an attempt to provide guidelines for the standardization of nomenclature, diagnostic criteria, reporting, antibody profiling, and risk assessment. PMID- 15280675 TI - Toward performing transplantation in highly sensitized patients. AB - Performing transplantation in highly sensitized patients represents a challenge for transplant units and organ exchange organizations. There are various approaches that can be adopted in order to transplant these patients, either through kidney exchange programs or by using desensitization protocols. In this article we discuss a number of these approaches, including the system currently used in the United Kingdom. The system encourages understanding of the patient's antibody profile and has resulted in a threefold increase in the number of highly sensitized patients undergoing transplantation. PMID- 15280676 TI - The acceptable mismatch program as a fast tool for highly sensitized patients awaiting a cadaveric kidney transplantation: short waiting time and excellent graft outcome. AB - There are many highly sensitized patients on the kidney waiting lists of organ exchange organizations because it is difficult to find a crossmatch negative cadaver kidney for these patients. Recently, several protocols have been developed to remove the donor-specific human leukocyte antigen (HLA) antibodies from the serum of these patients before transplantation. These approaches, including the use of intravenous immunoglobulins, plasmapheresis and immunoglobulins (plasmapheresis-cytomegalovirus-immunoglobulin), and immunoabsorption, seem to lead to a certain success rate, although the additional immunosuppression necessary to remove and control the production of donor specific alloantibodies may have its impact on the short-term (infections) and long-term (incidence of cancer) immune surveillance. Furthermore, some of these therapies represent a considerable financial burden for patients and society. In the present report, we advocate selection of crossmatch negative donors on the basis of the Acceptable Mismatch Program, as the first and best option for highly sensitized patients to undergo transplantations. No additional immunosuppression is necessary, and graft survival in this group of "difficult" patients is identical to that of nonsensitized recipients. Because the nature of the HLA polymorphism does not allow all patients to profit from this approach, removal of circulating HLA antibodies can be considered as a rescue therapy for those patients for whom the Acceptable Mismatch Program does not give a solution. PMID- 15280677 TI - Starting a crossover kidney transplantation program in the Netherlands: ethical and psychological considerations. AB - On April 15th, 2003, the first crossover kidney transplantation took place in The Netherlands. In September of the same year, a national database was established to facilitate kidney exchange between two donor-recipient couples. During 2004, kidneys from living donors will be exchanged between the seven university medical centers in The Netherlands. One of the conditions for successfully implementing this program was the need to address the ethical and psychologic implications involved. In this article we will discuss the ethical and psychologic considerations that are accompanying the practical preparations for the first Dutch crossover transplantation program. We identified five topics of interest: the influence of "donation by strangers" on the motivation and willingness of donor-patient couples, the issue of anonymity, the loss of the possibility of "medical excuses" for unwilling donors, the view that crossover is a first step to commercial organ trade, and the interference with existing organ donation programs. We concluded that whether viewed separately or in combination, these issues do not impede the efficient organization of a crossover program or raise worrying ethical issues. PMID- 15280678 TI - Weak humoral posttransplant alloresponse after a well-HLA-matched cadaveric kidney transplantation. AB - BACKGROUND: Screening of donor-specific antibodies or alloantibodies after kidney transplantation has not been performed routinely. The aim of this study was to evaluate the humoral antidonor and alloresponse of immunologically low-risk recipients of cadaveric renal allografts during the first posttransplant year. METHODS: Alloresponse against the donor was analyzed by means of T-cell immunoglobulin (Ig)G and IgM and B cell IgG flow cytometric crossmatch (FCXM) tests with sera from days 0, 21, 90, and 365 posttransplant. In addition, panel reactive anti-human leukocyte antigen (HLA) class I and class II antibodies (PRA I and PRA II) were analyzed using flow cytometric methods. The recipients were treated either with a low initial cyclosporine regimen with single-bolus antithymocyte globulin (ATG) or basiliximab induction or conventional cyclosporine triple therapy. RESULTS: No significant posttransplant anti-HLA class I or class II sensitization was found in the recipients as a whole. Recipients receiving a single-bolus ATG showed significantly higher proportion of PRA I positivity in the day 21 sample compared with the other groups. Flow cytometric donor-specific T- and B-cell IgG alloresponses remained low, but the proportion of T-cell IgM crossmatch-positive recipients increased during the study. Positive T-cell IgM FCXM was found to be associated with acute rejection episodes and cytomegalovirus (CMV) infections. CONCLUSIONS: In immunologically low-risk kidney-graft recipients, positive T-cell IgM FCXM at transplantation was found to be a risk factor for rejection episodes. Conversion of T-cell IgM FCXM to positive was found to be associated with CMV infections. PMID- 15280679 TI - Epstein-Barr virus primary mismatching and HLA matching: key risk factors for post lung transplant lymphoproliferative disease. AB - BACKGROUND: Posttransplant lymphoproliferative disease (PTLD) in lung transplant recipients (LTRs) is potentially lethal with considerable morbidity. The role of donor (D)/recipient (R) HLA matching is unknown. METHOD: We reviewed our LTRs from January 1994, when routine D/R Epstein-Barr virus (EBV) serologic screening was begun, through to January 2000. We examined whether D/R HLA match status influenced the risk of PTLD in EBV D+/R- mismatched LTRs. RESULTS: There were 16 D+/R- EBV-mismatched LTRs, 5 (31%) of whom developed PTLD (from a total of 237 LTRs; 218 survived >30 days). There were only two other cases of PTLD among the non-EBV primary mismatched patients. All patients received baseline immunosuppression of cyclosporine, azathioprine, and prednisolone without cytolytics and ganciclovir prophylaxis if "at risk" from cytomegalovirus. The five PTLD cases were diagnosed 81 to 734 (median 116) days from transplantation; three involved the lung allograft and two others involved lymph nodes. All PTLD patients seroconverted for EBV, whereas 7 of the 11 remaining EBV-mismatched patients who did not develop PTLD did not seroconvert. In the 16 EBV primary mismatched patients, there were 4 of 66 HLA allele matches in the 11 PTLD-free patients versus 15 of 30 matches in the 5 PTLD patients (P<0.001). This resulted in 2 or more HLA (A/B/DR) matches in 4 of 5 patients with PTLD versus 0 of 11 in the PTLD-free group (P=0.003). All PTLD patients were treated with reduced immunosuppression and antiviral therapy. Only two of the five LTRs who developed PTLD died, one with progressive disease despite chemotherapy and the other from chronic allograft rejection. CONCLUSION: A high degree of HLA matching in D/R EBV mismatched LTRs significantly increases the risk of PTLD. PMID- 15280680 TI - Long-term outcome of controlled, non-heart-beating donor liver transplantation. AB - BACKGROUND: Previous reports have established the feasibility of using livers from controlled, non-heart-beating donors (CNHBD) with good immediate graft function. This has been largely borne out of necessity because of the donor shortage. METHODS: Retrospective database review for the last 7 years (1995 2002), encompassing 19 patients receiving CNHBD, with follow-up period of 1,000 +/- 694 days, median 762 days. Detailed review of recipient characteristics, operative and clinical course, immunosuppression, complications, survival rates, and comparison with the results obtained in patients receiving transplants of allografts procured in standard fashion, from heart-beating donors RESULTS: Kaplan-Meier patient survival rates were 100%, 89.5%, and 83.5% at 30 days, 1, and 2 years, respectively, which is not different from recipients of livers procured from heart-beating cadaveric donors (P=0.74, log-rank test). Five patients died at a mean follow-up time of 492 (range 46-1,103) days. The causes of death were related to secondary sclerosing cholangitis (n=1), cardiac failure (n=1), and sepsis (n=3). Two (10.5%) recipients underwent retransplantation, one for primary graft nonfunction and one because of biliary cast syndrome with cholangitis. Significant preservation damage (ALT>2,000) developed in five patients, but this did not affect survival. The incidence of vascular (15.6% vs. 9.6%, P=0.34) and biliary complications (10.55 vs. 13.8%, P=0.68) was no different than for those recipients receiving standard cadaveric donors. CONCLUSIONS: CNHBD safely expands the donor pool with similar long-term results as those obtained in patients receiving organs from brain-dead donors under standard procurement techniques. PMID- 15280681 TI - Efficacy and safety of international donor sharing: a single-center, case controlled study on corneal transplantation. AB - BACKGROUND: To overcome the shortage of donor corneas, we are currently using donor corneas supplied by foreign eye banks. This single-center, case-controlled study was conducted to show the efficacy and safety of corneal transplantation using corneas from foreign donors. METHODS: A retrospective, case-controlled comparison of 118 corneal transplants using foreign donor corneas (foreign group) and domestic donor corneas (domestic group) was performed. The two groups were matched according to original disease, age, severity of preoperative neovascularization, and history of previous grafting. Clinical outcome and incidence of postoperative complications were compared between the two groups. RESULTS: The foreign group had a longer preservation-to-operation time than the domestic group, reflecting the longer transportation time. However, the incidence of clear grafts and postoperative complications such as immunologic rejection, infection, and glaucoma was not significantly different between the two groups. CONCLUSIONS: Corneal transplantations using foreign donor corneas are as effective and safe as those using domestic donor corneas, despite the longer preservation time. PMID- 15280682 TI - Persistence of low levels of alloantibody after desensitization in crossmatch positive living-donor kidney transplantation. AB - BACKGROUND: : Desensitization protocols have been developed to allow successful kidney transplantation in sensitized recipients. However, a detailed analysis of the impact of these protocols on alloantibody has not been performed. METHODS: : We studied 12 living-donor kidney-transplant recipients with positive antihuman globulin-enhanced complement dependent cytotoxicity (AHG-CDC) crossmatches against their donors. Using a variety of crossmatch techniques and single-antigen flowbeads (SAFBs), we characterized the specificity and amount of alloantibody at baseline before desensitization, after desensitization (using plasmapheresis followed by 100 mg/kg intravenous immunoglobulin, and anti-CD20 antibody), and 4 months after transplantation (after splenectomy and on maintenance immunosuppression). RESULTS: : All 12 patients with a positive baseline AHG-CDC crossmatch were AHG-CDC crossmatch negative at the time of transplant (after desensitization). However, despite desensitization, the majority of patients had low-level donor-specific alloantibodies demonstrable on the day of transplantation by both flow crossmatch (FXM 8/12) and SAFBs (10/11). Four months after transplantation, no patient had a positive AHG-CDC crossmatch, but again the majority had persistent low levels of donor-specific alloantibodies by FXM (6/12) and SAFBs (9/11). No patient experienced hyperacute rejection, and the persistence of low levels of donor-specific alloantibodies did not correlate with the development of humoral rejection in the early posttransplant period. CONCLUSIONS: : Despite desensitization, a majority of positive crossmatch transplant recipients demonstrate low levels of donor-specific alloantibodies both on the day of transplant and 4 months after transplantation. The impact of these antibodies appears to be minimal early after transplant, but their long term significance bears further study. PMID- 15280683 TI - Successful withdrawal of steroids in pediatric renal transplant recipients receiving cyclosporine A and mycophenolate mofetil treatment: results after four years. AB - BACKGROUND: Despite their numerous systemic side effects, glucocorticoids (steroids) still form a cornerstone in immunosuppressive regimens in pediatric renal transplant recipients. The addition of mycophenolate mofetil (MMF) to a cyclosporine A (CsA)-based immunosuppressive regimen after renal transplantation may allow steroid withdrawal and amelioration or avoidance of steroid-specific side effects. METHODS: In a retrospective case-control study, covering a mean follow-up period of 46 +/- 2.3 months and 40 patients aged 11.4 +/- 4.9 years, we analyzed the safety and efficacy of steroid withdrawal in pediatric renal transplant recipients receiving CsA micoroemulsion, MMF, and low-dose prednisone treatment. RESULTS: : Steroid withdrawal in all 20 pediatric renal transplant recipients receiving CsA and MMF was successful and not associated with an acute rejection episode; graft function remained stable. At baseline, the degree of growth retardation was comparable between the groups (mean height standard deviation scores [SDSs] -1.60 +/- 0.30 [withdrawal group] and -1.32 +/- 0.39 [case-control group]). After steroid withdrawal, prepubertal patients exhibited a significant catch-up growth with a mean height gain of 1.47 +/- 0.32 SDS, whereas height SDS did not improve in patients receiving steroids. Growth was also improved in pubertal patients who stopped taking steroids. Standardized body mass index in patients who stopped taking steroids decreased significantly by 49% from 0.87 +/- 0.31 SDS to 0.45 +/- 0.30 SDS. After steroid withdrawal, mean arterial blood pressure SDS decreased significantly by 45%. Moreover, the need for antihypertensive medication declined significantly in patients who stopped taking steroids. The white blood cell counts and hemoglobin levels were comparable between the groups. CONCLUSIONS: : This study suggests that steroids can be safely and successfully withdrawn in selected pediatric renal transplant recipients receiving immunosuppressive maintenance therapy consisting of CsA and MMF. PMID- 15280684 TI - Short- and long-term results of liver transplantation in infants aged less than 6 months. AB - BACKGROUND: Despite major surgical and medical advances, it is still a challenge to perform transplantation in small infants. This study, focusing on short- and long-term outcomes, summarizes our 10-year experience with liver transplantation (LTx) in infants aged less than 6 months. PATIENTS AND METHODS: We analyzed 43 patients aged 6 months or less (range: 12-184 days, median: 136 days) whose median weight at the time of LTx was 5.8 kg (range: 2.8-8.0 kg). The reasons for LTx were biliary atresia (n=27; 62.8%), neonatal hepatitis (n=6; 14%), neonatal cholestasis (n=4; 9.3%), and miscellaneous (n=6; 14%). The patients were followed up for a median time of 3 years and evaluated with respect to graft function, physical, and neurodevelopmental outcome. RESULTS: The patient survival was 90.7% after 1 year and 87.2% after 2 years. The graft survival was 86% after 1 year and 82.1% after 2 years. Twelve patients (27.9%) experienced 15 surgical complications requiring intervention, two of whom demonstrated vascular thrombosis (4.7%). Acute early rejection occurred in 15 patients (34.9%), and chronic rejection occurred in 3 patients (7%); 83.3% of the patients had normal liver function test results at the time of evaluation. Complications such as posttransplant lymphoproliferative disease (4.7%) and persistent arterial hypertension (4.7%) were rarely seen. The physical and neurodevelopmental outcomes were good. CONCLUSIONS: LTx in infants aged less than 6 months provides excellent short- and long-term results. Low weight or young age of infants awaiting LTx should not be exclusion criteria for LTx. PMID- 15280685 TI - Natural history, risk factors, and impact of subclinical rejection in kidney transplantation. AB - BACKGROUND: Subclinical rejection (SCR) is defined as histologically proven acute rejection in the absence of immediate functional deterioration. METHODS: We evaluated the impact of SCR in 961 prospective protocol kidney biopsies from diabetic recipients of a kidney-pancreas transplant (n=119) and one kidney transplant alone taken regularly up to 10 years after transplantation. RESULTS: SCR was present in 60.8%, 45.7%, 25.8%, and 17.7% of biopsies at 1, 3, 12, and greater than 12 months after transplantation. Banff scores for acute interstitial inflammation and tubulitis declined exponentially with time. SCR was predicted by prior acute cellular rejection and type of immunosuppressive therapy (P<0.05 0.001). Tacrolimus reduced interstitial infiltration (P<0.001), whereas mycophenolate reduced tubulitis (P<0.05), and the combination effectively eliminated SCR (P<0.001). Persistent SCR of less than 2 years duration on sequential biopsies occurred in 29.2% of patients and was associated with prior acute interstitial rejection (P<0.001) and requirement for antilymphocyte therapy (P<0.05). It resolved by 0.49 +/- 0.33 years and resulted in higher grades of chronic allograft nephropathy (CAN, P<0.05). True chronic rejection, defined as persistent SCR of 2 years or more duration and implying continuous immunologic activation was found in only 5.8% of patients. The presence of SCR increased chronic interstitial fibrosis, tubular atrophy, and CAN scores on subsequent biopsies (P<0.05-0.001). SCR preceded and was correlated with CAN (P<0.001) on sequential analysis. CONCLUSIONS: Histologic evidence of acute rejection in the absence of clinical suspicion resulted in significant tubulointerstitial damage to transplanted kidneys and contributed to CAN. PMID- 15280686 TI - Nitric oxide in early ischaemia reperfusion injury during human orthotopic liver transplantation. AB - BACKGROUND: Altered nitric oxide (NO) metabolism has been shown to contribute to ischemia-reperfusion (IR) injury in animal models. However, similar studies have not been performed in human liver transplantation (LT). In this study, we examined nitrate, nitrite, and nitrosothiols (NOx), NO synthases (endothelial [constitutive] nitric oxide synthase [eNOS] and inducible nitric oxide synthase [iNOS]), and nitrotyrosine in early IR injury after human LT. METHODS: Paired biopsies were obtained from nine donor livers before cold ischemia (retrieval biopsy) and after reimplantation (reperfusion biopsy). Sections were graded for reperfusion injury using the Suzuki score. NO was detected by chemiluminescence after reduction of NOx. Expression of eNOS and iNOS was by Western blot and reverse transcriptase polymerase chain reaction and peroxynitrite by immunodetection of 3-nitrotyrosine. RESULTS: Reperfusion biopsies showed histologic evidence of injury (median Suzuki score: retrieval 2, reperfusion 6, P=0.008) and neutrophil infiltration. NOx was reduced after reperfusion from 5.41 microM/100 mg (median, range 2.17-13.39 microM) to 3.51 microM (1.45-5.66 microM, P=0.05). eNOS protein was reduced after reperfusion from 0.6 units (median, range 0.45-1 unit) in retrieval biopsies to 0.39 units in reperfusion biopsies (range 0.2-0.79 units, P=0.007). There was no change in eNOS or iNOS mRNA expression or iNOS protein. Western blotting showed increased nitrotyrosine formation after reperfusion, median 0.42 (range 0.16-0.87) units in retrieval biopsies and 0.68 (0.29-1.06) units in reperfusion samples (P=0.007) and localized to periportal regions. CONCLUSIONS: iNOS protein may not contribute to early reperfusion injury during human LT. However, reduced NO bioavailability caused by reduced eNOS may contribute significantly to damage at this time point. PMID- 15280687 TI - Ten years of "extended" life: quality of life among heart transplantation survivors. AB - BACKGROUND: Long-term quality of life (QOL) outcome in heart transplant recipients still remains uncertain. This study evaluates the health status and QOL of survivors with associated predictors 10 years after heart transplantation. PATIENTS AND METHODS: A total of 276 patients who underwent heart transplantation in the Department of Cardiac Surgery, University of Pavia, between 1985 and 1992 were included in a cross-sectional study. Patients still alive 10 years after transplantation (n=122) were asked to complete the SF36 questionnaire and then received a full clinical examination. All QOL instruments that were used had acceptable reliability and validity. Descriptive statistics, Kaplan-Meier estimate, correlation coefficients, and general linear regression were used to analyze the data. RESULTS: Survival rates 1, 5, and 10 years after transplantation were 87%, 77%, and 57%, respectively, and the average life expectancy was 9.16 years. The mental QOL of patients 10 years after heart transplantation was similar to that among the general population. The physical QOL was worse among patients when compared with the QOL of the general population, with predictors including older age, being married, the presence of complications, and impaired renal function. CONCLUSIONS: Heart transplantation ensures a relatively high QOL even 10 years after surgery. Predictors of a poor QOL were determined, which may help to identify those patients for whom a poor outcome is likely so treatment can be tailored accordingly. PMID- 15280688 TI - A retrospective review of liver transplant patients treated with sirolimus from a single center: an analysis of sirolimus-related complications. AB - BACKGROUND: Sirolimus (SRL) is a powerful immunosuppressant used primarily in calcineurin inhibitors (CNI)-related nephrotoxicity. However, reports of drug related side effects are increasing. The aim of our report is to review the frequency and timing of these complications within our transplant patient population. METHODS: We retrospectively reviewed the medical records of liver transplanted patients treated with sirolimus between November 1998 and April 2002. The data collected included SRL serum levels, frequency of reported and documented SRL-related side effects, and survival outcomes. Statistical evaluation included Pearson chi-square and the Fisher's exact tests. RESULTS: Overall, 205 patients were identified, with 30 patients removed from the analysis for different reasons. Of the remaining 175 patients, 91 (52%) patients developed a complication other than an increase in serum triglycerides and/or cholesterol. The most frequent complications were: bilateral lower extremity edema (57.1%), dermatitis (25.3%), oral ulcers (24.2%), joint pain (23.0%), pleural effusion (16.5%) and increase in abdominal girth (9.9%). Other complications included: generalized edema (5.5%), pericardial effusion (5.5%), facial edema (2.2%), and upper extremity edema (1.3%). In addition, we reported two cases of hepatic artery thrombosis, one case of wound dehiscence with evisceration that required surgical repair, and one case of skin cancer. Interestingly, we found that a previous history of myocardial ischemia correlates with the development of SRL side effects. CONCLUSIONS: SRL is a powerful immunosuppressant but not devoid of side effects. These results have elevated our level of suspicion when instituting SRL and may help with early recognition and prevention of drug related complications. PMID- 15280689 TI - Proximal tubular dysfunction is associated with chronic allograft nephropathy and decreased long-term renal-graft survival. AB - BACKGROUND: Chronic allograft nephropathy is the major cause of graft loss after the first year of transplantation. Although many conditions are associated with its development, there is no method that can anticipate its risk in patients with good renal function. METHODS: We prospectively studied 92 renal-transplant recipients with good and stable allograft function and correlated the development of chronic allograft nephropathy and graft loss with their levels of urinary retinol binding protein (uRBP). Patients were divided in two groups regarding the level of their tubular protein: high, above 0.400 mg/L, and normal levels, 0.400 mg/L or less. RESULTS: Forty-eight (52%) patients had high levels of uRBP. At the enrollment time, patients with high and normal uRBP had comparable serum creatinine and cyclosporine trough levels. During a 5-year follow-up period, chronic allograft nephropathy was detected in 23 (25%) patients, 19 (82.6%) of whom had high levels of uRBP. Five-year chronic allograft nephropathy-free and graft survivals were significantly worse in patients with higher levels of uRBP than in patients with normal levels (57.5% vs. 89.9% P=0.0004; 70.7% vs. 100%, respectively, P=0.0002). High levels of uRBP were the strongest factor associated with the development of chronic allograft nephropathy (RR=5.3, 95% confidence interval 1.45-19.58, P=0.012). CONCLUSIONS: Among renal-transplant patients with good and stable graft function, high levels of uRBP identify those having a high risk of developing chronic allograft nephropathy. PMID- 15280690 TI - Blood pressure and renal function after kidney donation from hypertensive living donors. AB - BACKGROUND: Rising numbers of patients reaching end-stage kidney disease intensify the demand for expansion of the living-kidney-donor pool. On the basis of low risk in white donors with essential hypertension, our transplant center undertook a structured program of accepting hypertensive donors if kidney function and urine protein were normal. This study reports outcomes of hypertensive donors 1 year after kidney donation. METHODS: We studied detailed measurements of blood pressure (oscillometric, hypertensive therapy nurse [RN], and ambulatory blood pressure monitoring [ABPM]), clinical, and renal characteristics (iothalamate glomerular filtration rate [GFR], urine protein, and microalbumin) in 148 living kidney donors before and 6 to 12 months after nephrectomy. Twenty-four were hypertensive (awake ABPM>135/85 mm Hg and clinic/RN BP>140/90 mm Hg) before donation. RESULTS: After 282 days, normotensive donors had no change in awake ABPM pressure (pre 121 +/- 1/75 +/- 2 vs. post 120 +/- 1/ 5 +/- 1 mm Hg), whereas BP in hypertensive donors fell with both nonpharmacologic and drug therapy (pre 142 +/- 3/85 +/- 2 to post 132 +/- 2/80 +/- 1 mm Hg, P<.01). Hypertensive donors were older (53.4 vs. 41.4 years, P<.001) and had lower GFR after kidney donation (61 +/- 2 vs. 68 +/- 1 mL/min/1.73m, P<.01). After correction for age, no independent BP effect was evident for predicting GFR either before or after nephrectomy. Urine protein and microalbumin did not change in either group after donor nephrectomy. CONCLUSIONS: Our results indicate that white subjects with moderate, essential hypertension and normal kidney function have no adverse effects regarding blood pressure, GFR, or urinary protein excretion during the first year after living kidney donation. Although further studies are essential to confirm long-term safety, these data suggest that selected hypertensive patients may be accepted for living kidney donation. PMID- 15280691 TI - Treatment of cytomegalovirus disease with valganciclovir in renal transplant recipients: a single center experience. AB - Recent data suggest valganciclovir (VGC) to be as effective as ganciclovir for cytomegalovirus (CMV) prophylaxis. The objective of this study was to analyze the effect of oral valganciclovir in renal transplant patients with symptomatic CMV infection. Twenty-one patients with symptomatic CMV infection received VGC in doses adjusted to renal function until resolution of CMV antigenemia. The patients were followed for a mean of 5.5 months. During therapy, CMV antigenemia dropped in all patients from pretreatment positive levels of 5.2 +/- 3.7 to negative values of 0.25 +/- 0.2 positive cells/10,000 PBMC (P<0.001). After cessation of therapy, none of patients developed relapse of CMV antigenemia/symptoms within the follow-up. VGC therapy was well tolerated in all patients and no major adverse effects occurred. This pilot trial showed VGC to be safe and highly effective in antiviral therapy after renal transplantation. However, subsequent multicenter clinical trials for treatment of CMV disease are necessary. PMID- 15280692 TI - Posttransplant lymphoproliferative disease after allogeneic transplantation of the spleen in miniature swine. AB - Spleen transplantation (SpTx) was performed in miniature swine across full major histocompatibility complex barriers to study the tolerogenic effect of the spleen. This study describes the development of posttransplant lymphoproliferative disease (PTLD) after allogeneic SpTx. Recipient pigs underwent whole body irradiation (100 cGy), thymic irradiation (700 cGy), and native splenectomy (day 0), and received a 45-day course of intravenous cyclosporine (trough level 400-800 ng/ml). After SpTx, two of seven pigs developed PTLD (1 donor-type, 1 host-type). These two pigs had greater T cell depletion and higher trough levels of cyclosporine. Early changes that occurred prior to the development of clinical features of PTLD were increased porcine lymphotropic herpesvirus-1 viral loads in blood and tissues, and increased numbers of leukocytes, B cells, and total serum IgM. PTLD can occur after allogeneic SpTx in swine. This model may be useful in studies of the pathogenesis of PTLD. PMID- 15280693 TI - HSP-72 expression in pre-transplant donor kidney biopsies and post-transplant outcome. AB - In experimental transplant models, upregulation of renal heat shock proteins (HSP) represents a new therapeutic tool to improve allograft survival. In this clinical study, we hypothesized that HSP-72 expression in pretransplant donor kidney biopsies predicts posttransplant outcome. Expression of HSP-72 in pretransplant biopsies was assessed by immunohistochemistry and in situ hybridization in 82 consecutive renal transplantations and clinical data were collected prospectively during the first six months posttransplant. Renal tubular expression of HSP-72 was low and not influenced by donor-, graft-, or procedure related risk factors. Neither strength nor pattern of pretransplant HSP-72 staining discriminated allografts with complicated (40%) posttransplant courses from those without complications (60%). The low HSP-72 expression in pretransplant donor kidney biopsies failed to predict delayed graft function or acute rejection. These findings suggest that constitutive HSP-72 gene expression at the time of engraftment does not play a role in graft protection. PMID- 15280694 TI - Possible role of substance P in the ischemia-reperfusion injury in the isolated rabbit lung. AB - The origin of the endothelial damage leading to the ischemia-reperfusion injury after lung transplantation has not been elucidated. We postulated that neurotransmitters released during the preservation of the donor lung might explain this vascular derangement. Thus, in isolated rabbit lungs preserved over 24 hours, we evaluated the release of acetylcholine (ACh) and substance P (SP), the activity of their major degrading enzymes, acetylcholinesterase (AChE) and neutral endopeptidase (NEP), and changes in the capillary permeability. Both neurotransmitters showed the highest release rate in the first 15 minutes, followed by a sharp exponential decrement at 1, 6, 12 and 24 hours. AChE and NEP activities showed no variation at these time intervals. Basal capillary permeability significantly increased (P<0.01) after 24 hours preservation with saline. This increased permeability was avoided (P<0.01) by the SP fragment 4-11 (an SP receptors antagonist), but not by atropine. These results suggest for the first time a pathogenic role of SP in the ischemia-reperfusion injury, and thus the potential usefulness of SP antagonists as additives in the lung preservation solutions should be explored. PMID- 15280695 TI - Drawbacks to technological methods for confirming brain death. PMID- 15280697 TI - Cryptococcal monoarthritis without systemic involvement in a renal transplant patient. PMID- 15280698 TI - Successful treatment of encrusted cystitis and pyelitis with preservation of renal graft. PMID- 15280699 TI - Everolimus-induced drug fever after heart transplantation. PMID- 15280700 TI - Hypophosphatemia and the live liver donor. PMID- 15280701 TI - Resident 80-hour work week. PMID- 15280702 TI - Errors are not diseases: they are symptoms of diseases. PMID- 15280703 TI - Classification and consequences of errors in otolaryngology. AB - OBJECTIVE: To develop a preliminary classification system for errors in otolaryngology. METHODS: A retrospective, anonymous survey was distributed to 2,500 members of the American Academy of Otolaryngology-Head and Neck Surgery (AAO-HNS). Respondents were asked whether an error had occurred in their practice in the last 6 months, and if so, to describe the error, its consequences, and any corrective action taken. RESULTS: There were 466 (18.6%) responses. Two hundred ten (45% of respondents) otolaryngologists reported 216 errors. A classification system for errors in otolaryngology was developed. Errors were classified as related to history and physical (1.4%), differential or final diagnosis (1.4%), testing (10.4%), surgical planning (9.9%), wrong-site surgery (6.1%), anesthesia related (3.3%), wrong drug/dilution on the surgical field (3.8%), technical (19.3%), retained foreign body (0.9%), equipment-related (9.4%), postoperative care (8.5%), medical management (13.7%), nursing/ancillary (0.5%), administrative (6.6%), communication (3.8%), and miscellaneous (0.9%). There were 78 cases of major morbidity and 9 deaths. If these data are representative, there may be more than 2,600 episodes of major morbidity and more than 165 deaths related to medical error in otolaryngology patients annually. CONCLUSIONS: Human error in otolaryngology occurs in all practice components, including diagnostic, treatment, surgical, communication, and administrative. Types of errors reported by otolaryngologists differ from those reported by other specialists. Error classification systems may need to reflect each specialty's realm of practice. Errors in otolaryngology cause appreciable morbidity and mortality. Quantitative study of errors and the development of targeted prevention and amelioration strategies should be a high priority. PMID- 15280704 TI - Diagnosis and surgical management of nasopalatine duct cysts. AB - OBJECTIVES/HYPOTHESIS: Nasopalatine duct cysts are the most common cystic lesion of nonodontogenic origin of the maxilla. The purposes of the study were to review the epidemiology and clinical presentation, to describe the radiographic and pathological findings, and to discuss surgical management of this entity. STUDY DESIGN: Case presentations of two patients with nasopalatine duct cysts at a tertiary care institution with a review of the English medical literature from January 1960 to the present. METHODS: A 69-year-old man presented with an asymptomatic swelling of the premaxilla, and a 17-year-old woman presented with a painful swelling of the hard palate. A computed tomography scan, fine-needle aspiration, and preoperative workup were obtained in both cases. Literature was reviewed with respect to epidemiology, etiology, presentation, diagnostic studies, operative management, and recurrence rates. RESULTS: Computed tomography scan demonstrated midline ovoid cystic lesions in both cases. Fine-needle aspiration of both lesions revealed no evidence of malignancy. Surgical treatment consisted of enucleation in the first case and marsupialization in the second case. Both patients did well with no evidence of recurrence. Nasopalatine duct cyst presents in the fourth to sixth decades of life with a male predilection. Recurrence rates range from 0% to 11%. CONCLUSION: Nasopalatine duct cyst occurs in approximately 1% of the population. Presentation may be asymptomatic or include swelling, pain, and drainage from the hard palate. A well-circumscribed, round, ovoid, or heart-shaped radiolucency is seen on computed tomography. Pathological findings reveal squamous or respiratory cell types, or a combination of these, infiltrated by inflammatory cells. Enucleation is the preferred treatment with low recurrence rates. PMID- 15280705 TI - Respiratory retraining of refractory cough and laryngopharyngeal reflux in patients with paradoxical vocal fold movement disorder. AB - OBJECTIVES/HYPOTHESIS: The objective was to describe a case series of patients with refractory cough and paradoxical vocal fold movement disorder treated with respiratory retraining therapy. STUDY DESIGN: Retrospective review of a case series in a tertiary medical care center. METHODS: Five patients with laryngopharyngeal reflux were identified with refractory cough and paradoxical fold movement disorder on transnasal fiberoptic laryngoscopy by a greater than 50% reduction in airway during inspiration. The were four women and one man (age range, 42-67 y). All patients had normal forced vital capacity and forced expiratory flow but decreased ratio of forced inspiratory volume at 0.5 seconds (FIV(0.5)) to forced inspiratory vital capacity (FIVC) before starting therapy. All patients were treated with more than 6 months of twice-daily proton pump inhibitor therapy with improvement in reflux symptoms but persistent and severe daytime cough. They were subsequently treated with respiratory retraining therapy. Patients were asked to rate subjectively the severity of cough at the onset and conclusion of therapy. All patients underwent pulmonary function testing before and after therapy. Long-term follow-up ranged from 5 to 17 months. RESULTS: Patients received two to seven sessions of respiratory retraining therapy. The mean severity score changed from 9.2 before therapy to 1.3 after therapy. All patients subjectively described an improvement in the severity of their cough. Transnasal flexible laryngoscopy demonstrated improvement in paradoxical vocal fold movement, and pulmonary function testing showed improvement in the FIV(0.5)/FIVC ratio. CONCLUSION: Patients with laryngopharyngeal reflux and refractory cough in the absence of pulmonary disease should be evaluated for paradoxical vocal fold movement disorder. Respiratory retraining therapy may represent an effective therapy for cough in the absence of relief from standard management of laryngopharyngeal reflux. PMID- 15280706 TI - A genomic predictor of oral squamous cell carcinoma. AB - OBJECTIVES/HYPOTHESIS: The objective was to identify a genomic profile that predicts the likelihood of oral squamous cell carcinoma compared with normal oral mucosa in unknown tissue samples. STUDY DESIGN: Using a training set of tissue samples that were histologically classified as oral squamous cell carcinoma or normal mucosa, the authors used principal component analysis to develop a genomic predictor for oral squamous cell carcinoma. On a separate test set of unclassified samples, the authors used the predictor to classify the samples, then evaluated the performance of the predictor using histological diagnosis. METHODS: The authors used a data set consisting of messenger RNA extracted from 29 oral squamous cell carcinoma and 19 normal oral mucosa tissue samples and hybridized to Affymetrix oligonucleotide microarrays containing probe sets for 7070 genes and expressed sequence tags. The samples were divided into a training set of 15 oral squamous cell carcinoma and 10 normal samples and a test set consisting of the remaining samples. Using principal component analysis on the training set, the authors found a composite gene expression vector (principal component vector), which they used to compute likelihood ratios for oral squamous cell carcinoma on the test set. By calculating the contribution of each gene to the principal component vector, the authors identified genes with the greatest predictive value. RESULTS: Using the likelihood ratio, the authors correctly classified all 23 samples in the test set as either oral squamous cell carcinoma or normal. The authors found that many of the most predictive genes are known to be markers of squamous cell carcinoma or normal mucosa. CONCLUSION: Principal component analysis can be used with genomic microarray data to correctly predict the presence of oral squamous cell carcinoma in unknown tissue samples. PMID- 15280707 TI - Effect of magnetic resonance imaging on internal magnet strength in Med-El Combi 40+ cochlear implants. AB - OBJECTIVE: Magnetic resonance imaging (MRI) has been contraindicated when cochlear implants containing an internal magnet are in place because of concerns regarding torque, force, demagnetization, artifacts, induced voltages, and heating. The objective was to determine the magnetic field strength of Med-El Combi 40+ cochlear implant internal magnets after MRI studies. STUDY DESIGN/METHODS: Two fresh cadavers were used to study demagnetization using a repeated measures design and a magnetometer. Pre- and postMRI measurement of magnetic field strength was completed. Five sets of sagittal T1-weighted, axial T1-weighted, and axial T2-weighted sequences were performed on a cadaver at 0.2 Tesla in the device-up and device-down positions. In the other cadaver, 15 sets of sagittal T1-weighted, axial T1-weighted, and axial T2-weighted sequences were performed on a cadaver at 1.5 Tesla were conducted, 5 each with the head oriented at 80, 90, and 100 degrees rotated around the yaw plane (rotated around the z axis). Subsequently, three cochlear implant patients completed 0.2 Tesla MRIs. For these patients, subjective and objective assessment of cochlear implant performance was performed. SETTING: Academic medical center. RESULTS: In the cadaver studies, analysis of variance showed no significant difference in the magnetic field strength after the 0.2 or 1.5 Tesla scans. There was no significant difference in the magnetic field strength for the three patients undergoing 0.2 Tesla MRIs and no adverse consequences, including no changes in telemetry, auditory sensations, nonauditory sensations, and sound quality. CONCLUSIONS: No significant demagnetization of the internal magnet occurred during repeated 1.5 Tesla MRI scans with the head orientations used in this study. In the cochlear implant patients, no significant demagnetization of the internal magnet occurred after a 0.2 Tesla MRI. PMID- 15280708 TI - Swallowing-related quality of life after head and neck cancer treatment. AB - OBJECTIVES: To determine the role of treatment modality in swallowing outcome after head and neck cancer treatment and to identify potential risk factors for posttreatment dysphagia. STUDY DESIGN: Cross-sectional survey of patients with no evidence of disease 12 months or more after the treatment of a stage III or IV squamous cell carcinoma of the oropharynx, larynx, or hypopharynx. METHODS: Potential subjects were stratified by tumor site and tumor T-stage to achieve a balanced comparison between chemoradiation (n = 18) and surgery/radiation (n = 22) groups. Outcome measures included a dysphagia risk factor survey, the MD Anderson Dysphagia Inventory (MDADI), and the Short-Form 36 (SF-36). RESULTS: Patients who received chemoradiation for oropharyngeal primaries demonstrated significantly better scores on the emotional (P =.03) and functional (P =.02) subscales of the MDADI than did patients who underwent surgery followed by radiation. There were no significant differences between chemoradiation and surgery/radiation groups for laryngeal and hypopharyngeal primaries. Additional risk factors for posttreatment dysphagia include prolonged (>2 weeks) nothing by mouth (NPO) status (P =.002) and low SF-36 Mental Health Subscale score (P =.002). CONCLUSION: The study suggests that chemoradiation may provide superior swallowing outcome to surgery/radiation in patients with oropharyngeal primary. Patients with depressed mental health and prolonged feeding tubes may be at higher risk of long-term dysphagia. PMID- 15280709 TI - The transglabellar subcranial approach for nasal dermoids with intracranial extension. AB - OBJECTIVES/HYPOTHESIS: Nasal dermoids are the most common congenital midline nasal lesions. When a midline nasal pit or cyst is identified, scans should be obtained to look for an intracranial connection. The intracranial portion of the lesions has traditionally been approached by the performance of a frontal craniotomy. The transglabellar subcranial approach is a useful technique to resect these lesions and offers several advantages over a traditional craniotomy approach. STUDY DESIGN: Retrospective case review. METHODS: Records of two patients who underwent excision of nasal dermoids by way of a subcranial approach were analyzed. RESULTS: Two patients underwent successful resection of nasal dermoids with intracranial extension by way of a subcranial approach. The patients were 13 months and 19 months old at the time of excision and have been followed for 7 years and 6 years, respectively. There has not been any recurrence of the lesions. There has been no apparent negative impact on facial growth in either of these cases. CONCLUSIONS: The subcranial approach is an effective technique for the resection of nasal dermoids with intracranial extension. These lesions have traditionally been managed with lateral rhinotomy, midface degloving, or external rhinoplasty approaches combined with a frontal craniotomy. The subcranial approach offers excellent exposure, minimizes frontal lobe retraction, reduces the likelihood of cerebrospinal fluid leak, and provides for excellent cosmetic result. This approach was used in two cases with long-term follow-up. The lesions were successfully resected in both cases. Long-term follow up has shown no recurrence or negative effect on craniofacial growth. PMID- 15280710 TI - The status of the contralateral ear in established unilateral Meniere's disease. AB - OBJECTIVES/HYPOTHESIS: To determine the incidence of measurable audiometric changes in the contralateral ear and clinical bilateral Meniere's disease in individuals with longstanding established unilateral Meniere's disease. STUDY DESIGN: Analysis of data in an ongoing prospective study. METHODS: One hundred and one patients who were treated with intratympanic gentamicin installation for disabling unilateral Meniere's disease were followed up for a minimum of 2 years. The mean follow-up time between the initial diagnosis to the last audiogram conducted was 12 +/- 7 (range 2-45) years, and the mean follow-up between the first and last audiograms conducted in our clinic was 5 +/- 3 (range 2-15) years. The incidence of Meniere's disease (clinical diagnosis) as compared with audiometric changes in the contralateral ear was analyzed. RESULTS: Compared with several published reports, the development of bilateral Meniere's disease in our series within the described observation period was much lower (5%). The incidence of contralateral isolated hearing loss in the low frequencies was 16% (average threshold of .25 and .5 kHz of 25 dB or more). The time interval between the initial diagnosis and the onset of contralateral findings ranged from 0 to 26 years. CONCLUSION: In our experience, individuals with disabling longstanding unilateral disease are not likely to develop bilateral disease. However, a small but significant percentage of patients have audiometric changes in the low frequencies of the contralateral ear (.25, .5 kHz). PMID- 15280711 TI - Prospective evaluation of endoscopic approaches to the thyroid compartment. AB - OBJECTIVE: To compare a number of endoscopic approaches to the thyroid compartment. DESIGN: Prospective, nonrandomized, experimental investigation in a porcine model. METHODS: We performed a consecutive series of 13 endoscopic thyroidectomies using 5 distinct approaches. The procedures differed by the direction of the approach, incision placement, and use of facilitative maneuvers. The parameters assessed included procedure duration, estimated blood loss, heart rate, blood pressure, temperature, oxygen saturation, and arterial blood gas levels. The thyroid specimens were weighed and examined histologically. RESULTS: Four of the endoscopic approaches were successfully used for resection of the thyroid (12 of 13 animals). The mean operative times were as follows: superior approach (n = 4), 47 +/- 14.6 minutes; lateral axillary approach (n = 4), 67 +/- 11.8 minutes; and superficial axillary (n = 3), 67.7 +/- 22.3 minutes. The one axillary approach took 84 minutes. The precordial approach (n = 1) lasted 47 minutes and then required open conversion. The overall median estimated blood loss was 0 (range 0-100) mL. The mean change in blood pressure and pH from the beginning to the end of the procedure was -0.5 +/- 24.1 mm Hg and 0.16 +/- 0.07, respectively. The thyroid glands weighed 4.3 +/- 0.9 g and had normal glandular architecture with no evidence of significant tissue trauma or thermal injury. There were no cases of pneumothorax, subcutaneous emphysema, or air embolism. CONCLUSION: A number of approaches to the thyroid compartment are conceivable. The superior approach proved to be the fastest and easiest, whereas the lateral axillary and superficial axillary were the best approaches from a cosmetic and clinical standpoint. PMID- 15280712 TI - Pathology of the olfactory epithelium: smoking and ethanol exposure. AB - OBJECTIVE: To investigate the effects of tobacco smoke on the olfactory epithelium. Cigarette smoking has been associated with hyposmia; however, the pathophysiology is poorly understood. The sense of smell is mediated by olfactory sensory neurons (OSNs) exposed to the nasal airway, rendering them vulnerable to environmental injury and death. As a consequence, a baseline level of apoptotic OSN death has been demonstrated even in the absence of obvious disease. Dead OSNs are replaced by the mitosis and maturation of progenitors to maintain sufficient numbers of neurons into adult life. Disruption of this balance has been suggested as a common cause for clinical smell loss. This current study will evaluate the effects of tobacco smoke on the olfactory mucosa, with emphasis on changes in the degree of OSN apoptosis. STUDY DESIGN: A rat model was used to assess the olfactory epithelium after exposure to tobacco smoke. METHODS: Rats were exposed to tobacco smoke alone (for 12 weeks), smoke plus dietary ethanol (for the final 5 weeks), or to neither (control). Immunohistochemical analysis of the olfactory epithelium was performed using an antibody to the active form of caspase-3. Positive staining for this form of the caspase-3 enzyme indicates a cell undergoing apoptotic proteolysis. RESULTS: Control rats demonstrated a low baseline level of caspase-3 activity in the olfactory epithelium. In contrast, tobacco smoke exposure triggered a dramatic increase in the degree of OSN apoptosis that affected all stages of the neuronal lineage. CONCLUSIONS: These results support the following hypothesis: smell loss in smokers is triggered by increased OSN death, which eventually overwhelms the regenerative capacity of the epithelium. PMID- 15280713 TI - Skin testing in predicting response to nasal provocation with alternaria. AB - OBJECTIVE: Examine the efficacy of epicutaneous and intradermal testing in predicting response to nasal provocation with Alternaria antigen. STUDY DESIGN AND SETTING: Prospective study. Subjects were tested with the Multi-Test II (MT) epicutaneous testing device. Subjects with negative wheals were then tested with a 1:500 weight:volume intradermal injection of Alternaria. They had baseline assessment of nasal cross-sectional area (CSA) using acoustic rhinometry and underwent nasal provocation with increasing Alternaria concentrations. CSA was assessed at each concentration. A nasal allergen provocation score (NAP) of nasal symptoms as well as a nasal visual analogue scale (VAS) were also completed with each concentration. RESULTS: Sensitivity and specificity of MT in predicting nasal response to Alternaria were poor at 42% and 44%, respectively. The addition of intradermal testing increased sensitivity only modestly to 58%. hierarchical linear modeling analysis demonstrated that subjects positive to Alternaria on skin testing did not show a significant reduction in nasal CSA on acoustic rhinometry or significant elevations in two nasal symptom scores with direct nasal provocation. CONCLUSION AND SIGNIFICANCE: Skin testing with either epicutaneous or intradermal testing may not be an accurate or sufficient technique in the assessment of Alternaria reactivity. These results suggest that mold allergies may involve more complex immune mechanisms than simply an immunoglobulin (Ig)E mediated type I immediate hypersensitivity response alone. An alternate model for mold sensitivity, as well as modifications in testing methods, may be required in the evaluation of mold allergy. PMID- 15280714 TI - The resident 80-hour work week: how has it affected surgical specialties? AB - OBJECTIVES: To identify strategies employed by surgical departments to address recently implemented resident duty hour regulations, and to assess resident and faculty acceptance of these changes. METHODS: Attendees to the 2003 Residency Program Coordinator/Administrator Workshop for sub-specialties (Denver, CO) were surveyed. RESULTS: The study population included 46 respondents spanning 9 surgical sub-specialties. Forty-eight percent of programs instituted at least 1 administrative change specifically to comply with duty hour regulations. The most commonly employed strategies were the hiring of nurse practitioners or physician assistants (30%) and the use of Internet-based software to track resident duty hours (30%). Other changes included giving call responsibilities to residents on research rotations (19%), institution of home-call (13%), and assignment of a night-float resident (11%). Perceptions of program coordinators indicated that junior residents and junior faculty accepted changes better than did senior residents and senior faculty (P=.025). CONCLUSION: The resident 80-hour work week is a major health care policy change that has required academic sub-specialty departments to make significant alterations in their administrative structure. Further study is necessary to determine how these changes affect both quality of training and patient care in the short and long term. PMID- 15280715 TI - The utility of routine chest radiography following jet ventilation in elective laryngotracheal surgery. AB - OBJECTIVES/HYPOTHESIS: Jet ventilation is a useful mode of airway management for laryngotracheal surgery. The objective of this study is to evaluate the utility of routine chest radiography following jet ventilation for these cases. STUDY DESIGN: Retrospective chart review. METHODS: Thirty-four elective surgical procedures performed from 1998 to 2002 are reviewed for postoperative airway and pulmonary complications. Data were collected from clinical notes as well as from the results of chest radiographs. This included the need for tracheotomy, reintubation, pneumothorax, pneumomediastinum, and atelectasis. RESULTS: A retrospective chart examination revealed no major complications. Twenty-nine of the 34 cases had postoperative chest radiography (CXR) performed on a routine basis. Atelectasis was present in 12 of 29 CXR (41%); no intervention was required in these cases. CONCLUSIONS: Routine postoperative CXR may not be useful following jet ventilation for elective laryngotracheal surgery. The safety of jet ventilation is discussed. PMID- 15280716 TI - Nasopharyngeal actinomycosis: a rare cause of nasal airway obstruction. AB - Nasopharyngeal actinomycosis is a rare clinical disease. It can occur after nasal trauma or surgical manipulation. It is also reported to occur without prior trauma, making diagnosis difficult. We report a case of nasopharyngeal actinomycosis that presented as nasal airway obstruction causing snoring and mimicking nasopharyngeal carcinoma. To our knowledge, only a few other cases of nasopharyngeal actinomycosis have previously been published, most coming after mucosal trauma. Diagnosis is made by observing the bacteria or its associated sulfur granules in the biopsy specimen. This anaerobic organism is difficult to culture. Treatment consists of wide debridement and prolonged antibiotic therapy, with good prognosis. PMID- 15280717 TI - Chondroitin sulfate hydrogel and wound healing in rabbit maxillary sinus mucosa. AB - OBJECTIVES/HYPOTHESIS: Chondroitin sulfate (CS) is a glycosaminoglycan in the extracellular matrix of all vertebrates. A biocompatible, nonimmunogenic, pliable hydrogel preparation of CS has recently been produced and has shown benefit in wound healing in murine and porcine epidermis. The objective of the current experiment is to compare the wound healing properties of CS hydrogel versus no treatment in wounds of the maxillary sinus mucosa. STUDY DESIGN: Prospective investigation in an animal model. METHODS: A 6 mm wound was created in bilateral maxillary sinuses of 17 New Zealand white rabbits. CS hydrogel (case) and no dressing (control) were randomly assigned to one side each as wound treatment. Wounds were examined ex vivo at 2, 4, 6, 10, and 14 day postinjury intervals. Wound diameter was measured microscopically by a blinded investigator. Representative specimens were examined histologically. RESULTS: The CS disc was partially integrated into the wounds at the 4-day interval and completely integrated by the 6-day interval. The average wound diameters for the case versus control side were similar at 2 days (3.75 mm vs. 4.42 mm) but differed significantly at 4 days (2.86 mm. vs. 3.80 mm., P =.035). At 6 days, the wounds could not be discerned on either the case or control sides. However, histologic analysis revealed accelerated healing with the CS treatment. The treated wounds displayed respiratory epithelium as opposed to the squamous epithelium exhibited on the untreated sides. CONCLUSIONS: Despite some limitations, the New Zealand white rabbit is an effective model for the study of sinonasal wound healing. CS hydrogel accelerates wound healing in sinonasal mucosa at a 4-day endpoint. We propose that the CS hydrogel acts as a surrogate extracellular matrix, serving as a repository for cytokines and growth factors produced by the regenerating mucosa. It may also provide a structural framework for fibroblasts and epithelial regeneration. Further study is necessary to establish this relationship. PMID- 15280718 TI - Is a plexiform neurofibroma pathognomonic of neurofibromatosis type I? AB - OBJECTIVES/HYPOTHESIS: Several prominent textbooks have claimed that a plexiform neurofibroma is pathognomonic for neurofibromatosis type I. This is not in agreement with the National Institutes of Health criteria, which require two signs to be present, one of which can be a plexiform neurofibroma. Is a plexiform neurofibroma pathognomonic for neurofibromatosis type I? STUDY DESIGN: Retrospective chart review. METHODS: Fifty-one patients with surgically resected plexiform neurofibromas between the years 1991 and 2001 were identified, and their charts reviewed. The presence and absence of any manifestation of neurofibromatosis type I as detailed in the National Institutes of Health criteria were recorded. The pathology database at the Hospital for Sick Children (HSC) was searched retrospectively. RESULTS: Five of the 51 patients had a single solitary plexiform neurofibroma. They were followed at the neurofibromatosis clinic at the HSC with regular detailed examinations and no other stigmata were found. CONCLUSION: Although plexiform neurofibromas are highly suggestive of neurofibromatosis type I, they are not pathognomonic as claimed. PMID- 15280719 TI - Cochlear implantation for children with GJB2-related deafness. AB - OBJECTIVES/HYPOTHESIS: Mutations in GJB2 are a common cause of congenital sensorineural hearing loss. Many children with these mutations receive cochlear implants for auditory habilitation. The purpose of the study was to compare the speech perception performance of cochlear implant patients with GJB2-related deafness to patients without GJB2-related deafness. STUDY DESIGN: Retrospective case review. METHODS: Pediatric cochlear implant recipients who have been tested for GJB2 mutation underwent chart review. All patients received cochlear implantation at a tertiary referral center, followed by outpatient auditory habilitation. Charts were reviewed for cause and duration of deafness, age at time of cochlear implantation, intraoperative and postoperative complications, duration of use, and current age. Results of standard tests of speech perception administered as a part of the patients' auditory habilitation were reviewed. RESULTS: Twenty patients with GJB2 mutations were compared with 27 patients without GJB2 mutations. There was no statistical difference between patients with and without GJB2-related congenital sensorineural hearing loss with regard to open-set and closed-set speech recognition performance at 12, 24, and 36 months after cochlear implantation. Surgical complications were uncommon. CONCLUSION: Pediatric patients with congenital sensorineural hearing loss without other comorbid conditions (eg, developmental delay, inner ear malformations) perform well when they receive cochlear implantation and auditory habilitation. The presence or absence of GJB2 mutation does not appear to impact speech recognition performance at 12, 24, and 36 months after implantation. PMID- 15280720 TI - Giant cell arteritis: a new association with benign paroxysmal positional vertigo. AB - OBJECTIVE: To assess the incidence and characteristics of both benign paroxysmal positional vertigo (BPPV) and positional nystagmus in a series of patients with giant cell arteritis (GCA). STUDY DESIGN: Patients diagnosed with GCA between June 1999 and May 2001 at the single hospital for a defined population were examined prospectively. METHOD: Patients included in this study fulfilled the 1990 American College of Rheumatology classification criteria for GCA. Otologic and oculographic studies were performed. Type, frequency, and outcome of positional oculographic findings was assessed. Patients were required to have been examined within 1 week after the onset of corticosteroid therapy. Data found in GCA patients were compared with those observed in an age, sex, and ethnically matched control population. Further studies in patients and controls were performed 3 and 6 months later. RESULTS: Forty-four patients and 44 matched controls were included in this study. Nine (20.5%) GCA patients fulfilled diagnostic criteria of BPPV compared with only 1 (2.3%) of the controls (P =.007). In seven of these nine GCA patients, BPPV was related to the posterior and two to the horizontal semicircular canals, respectively. Horizontal nystagmus was found in seven GCA patients who developed nystagmus in the head hanging position test compared with none in the controls (P =.006). CONCLUSIONS: The present study shows a higher frequency of BPPV in GCA than in matched controls. Because most clinical manifestations in GCA are caused by ischemic complications, our results suggest an ischemic etiology as responsible for BPPV in these elderly patients. According to these results, GCA may constitute a new association with BPPV. PMID- 15280721 TI - Histopathological basis of hearing impairment in Wolf-Hirschhorn syndrome. AB - OBJECTIVES/HYPOTHESIS: To perform histological examination of temporal bones acquired from an infant with Wolf-Hirschhorn syndrome with an emphasis on identifying abnormalities that might be responsible for hearing impairment in this disorder. STUDY DESIGN: Retrospective case review. METHODS: Temporal bones were taken at autopsy from a 10-month-old infant with Wolf-Hirschhorn syndrome. The right-side temporal bone was studied by microdissection. The middle ear was examined, and the inner ear sensory organs dissected for study by light microscopy. The left-side temporal bone was embedded in celloidin, and sections were cut for microscopic examination. RESULTS: Chronic otitis media was observed in both ears. Inflammation, effusion, and adhesions were present in the middle ear space. The malleus was malformed, and the chorda tympani nerve was found to pass through the bone of the malleus bilaterally. There was an area of sharply defined outer hair cell loss in the lower basal turn of the right-side organ of Corti, and defects were noted in the bone of the apical osseous spiral lamina in both cochleae. CONCLUSION: In addition to the presence of otitis media, the likelihood of congenital abnormalities of the middle and inner ear should be considered in the assessment of patients with Wolf-Hirschhorn syndrome with hearing impairment. PMID- 15280722 TI - Effects of long-term nasal continuous positive airway pressure therapy on morphology, function, and mucociliary clearance of nasal epithelium in patients with obstructive sleep apnea syndrome. AB - OBJECTIVES/HYPOTHESIS: The objective was to investigate the possible modification of nasal mucosa function and mucociliary clearance in a group of patients with severe obstructive sleep apnea syndrome receiving mechanical ventilation with long-term nasal continuous positive airway pressure (n-CPAP), without nasal diseases. STUDY DESIGN: The study design was experimental. Eight (six male and two female) nonsmoker patients were selected on the basis of two sleep questionnaires, were identified as needing n-CPAP therapy, and showed normal values of mucociliary transport time, ciliary beat frequency, and anterior rhinomanometry. METHODS: After a full polysomnographic examination, the authors recorded respiratory disturbance index (RDI), apnea/hypopnea index, nadir arterial oxygen saturation, and sleep stage. Every patient underwent pulmonary function test; arterial blood gas analysis; chest radiography; electrocardiography; ear, nose, and throat evaluation with rhinoscopy; anterior rhinomanometry; a saccharine test to measure the mucociliary transport time; and a brushing of nasal epithelium for study of ciliary beat frequency. All patients underwent polysomnographic examination in basal condition with overnight n-CPAP (without humidifier) and repeated this examination after 1 and 6 months with Auto CPAP (Autoset Res Care, Sidney, Australia) to titrate n-CPAP pressure and measure the new respiratory disturbance index. RESULTS: The mean basal respiratory disturbance index (number of respiratory events during sleep per hour of recording time) was 53.7 +/- 21.5 events/h; after 6 months of n-CPAP therapy (mean value, 7.5 +/- 0.7 cm H2O) the respiratory disturbance index was 5.7 +/- 3.76 events/h. Values for nasal resistance, mucociliary transport time, and ciliary beat frequency were normal before and after the ventilatory treatment. CONCLUSION: In the study group of patients with severe obstructive sleep apnea syndrome, the nocturnal use of n-CPAP without humidifier did not modify the function and mucociliary clearance of nasal epithelium. PMID- 15280723 TI - Nontuberculous mycobacteria-induced parotid lymphadenitis successfully limited with clarithromycin and rifabutin. AB - OBJECTIVES/HYPOTHESIS: Nontuberculous mycobacterial adenitis of the parotid gland is often difficult to diagnose. The rarity of these infections in the parotid region and the lack of specific guidelines pose a treatment challenge to the clinician. Three cases of nontuberculous mycobacterial adenitis are presented, with clinical response to antibiotics before surgery. STUDY DESIGN: Retrospective chart review was made of children up to 18 years of age presenting with a parotid mass diagnosed as nontuberculous mycobacterial infection. METHODS: Three patients (age range, 15 to 30 mo) with nontuberculous mycobacteria-induced parotid lymphadenitis were treated with a combination antibiotic regimen of clarithromycin and rifabutin or with clarithromycin alone. RESULTS: All three patients responded clinically to the antibiotic treatment as evidenced by a smaller mass size and resolution of the overlying discoloration. Subsequent parotidectomy or biopsy appeared to be easier to perform because of decreased inflammation and edema and a more readily dissectible facial nerve. CONCLUSION: Children with nontuberculous mycobacteria-induced parotid lymphadenitis should be started on a trial of antibiotic treatment before surgery. Although surgery remains the definitive treatment modality, a larger study of preoperative antibiotic use against nontuberculous mycobacterial adenitis of the parotid in children is necessary. PMID- 15280725 TI - Can maximum phonation time predict voice outcome after thyroplasty type I? AB - BACKGROUND: Thyroplasty type I, as introduced by Isshiki and colleagues almost 30 years ago, has become the gold standard of improving glottal incompetence caused by unilateral vocal fold paralysis. Intraoperative assessment of the adequacy of glottal closure is subjective and based on the perceptual judgment of vocal quality and degree of improvement in glottal gap size. OBJECTIVE/HYPOTHESES: The primary purpose of this study was to investigate whether the intraoperative measurement of maximum phonation time (MPT) is an adequate predictor of voice outcome after thyroplasty type I. To assess this possibility, it was necessary to evaluate the effect of body posture (seated vs. supine) and anesthesia (none vs. light sedation) on the measure of MPT. STUDY DESIGN: A prospective study of 20 individuals with unilateral vocal fold paralysis was undertaken. METHODS: Subjects were assessed at three time points: pre-, intra-, and postoperatively across parameters of breathiness rating, glottal gap size, glottal flow rate, and MPT. RESULTS: Results indicated that MPT was significantly lower in the supine versus seated position. In addition, light sedation resulted in a trend toward lower MPT that was not statistically significant. Finally, the intraoperative measurement of MPT, although lower than a 1-month postoperative measurement, was significantly predictive of the outcome. CONCLUSIONS: The intraoperative measure of MPT appears to be an adequate predictor of the postoperative outcome. PMID- 15280724 TI - Management of stage IV glottic carcinoma: therapeutic outcomes. AB - OBJECTIVES/HYPOTHESIS: The best therapeutic approach for the treatment of stage IV glottic carcinoma is controversial. STUDY DESIGN: A retrospective study. METHODS: A retrospective study of Tumor Research Project data was performed using patients with stage IV glottic squamous cell carcinoma treated with curative intent by five different treatment modalities from 1955 to 1998 at Washington University School of Medicine and Barnes-Jewish Hospital (St. Louis, MO). RESULTS: Ninety-six patients with stage IV glottic carcinoma were treated by five modalities: total laryngectomy (TL) (n = 13), total laryngectomy with neck dissection (TL/ND) (n = 18), radiation therapy alone (RT) (n = 7) (median dose, 69.5 Gy), total laryngectomy combined with radiation therapy (TL/RT) (n = 10), and total laryngectomy and neck dissection combined with radiation therapy (TL/ND/RT) (n = 48). The overall 5-year observed survival (OS) rate was 39%, and the 5-year disease-specific survival (DSS) rate was 45%. The 5-year DSS rates for the individual treatment modalities included the following: TL, 58.3%; TL/ND, 42.9%; RT, 50.0%; TL/RT, 30.0%; and TL/ND/RT, 43.9%. There was no significant difference in DSS for any individual treatment modality (P =.759). The overall locoregional control rate was 69% (66 of 96). The overall recurrence rate was 39% with recurrence at the primary site and in the neck at 19% and 17%, respectively. Recurrence was not related to treatment modality. The 5-year DSS after treatment of locally recurrent cancer (salvage rate) was 30% (3 of 10) and for recurrent neck disease (28 of 67) was 42%. The incidence of delayed regional metastases was 28%; of distant metastasis, 12%; and of second primary cancers, 9%. There was no statistically significant difference in survival between node-negative (N0) necks initially treated (5-y DSS, 31%) versus N0 necks observed and later treated if necessary (5-y DSS, 44%) (P =.685). CONCLUSION: The five treatment modalities had statistically similar survival, recurrence, and complication rates. The overall 5 year DSS for patients with stage IV glottic carcinoma was 45%, and the OS was 39%. The cumulative disease-specific survival (CDSS) was 0.4770 with a mean survival of 10.1 years and a median survival of 3.9 years. Patients younger than age 55 years had better survival (DSS) than patients 56 years of age or older (P =.0002). Patients with early T stage had better survival than patients with more advanced T stage (P =.04). Tumor recurrence at the primary site (P =.0001) and in the neck (P =.014) and distant metastasis (P =.0001) had a deleterious effect on survival. Tumor recurrence was not related to treatment modality. Patients with clear margins of resection had a statistically significant improved survival (DSS and CDSS) compared with patients with close or involved margins (P =.0001). Post treatment quality of life was not significantly related to treatment modality. Patients whose N0 neck was treated with observation and appropriate treatment for subsequent neck disease had statistically similar survival compared with patients whose N0 neck was treated prophylactically at the time of treatment of the primary. A minimum of 7 years of follow-up is recommended for early identification of recurrent disease, second primary tumors, and distant metastasis. None of the standard treatment modalities currently employed has a statistical advantage regarding survival, recurrence, complications, or quality of life. PMID- 15280726 TI - Retrosigmoid versus middle fossa surgery for small vestibular schwannomas. AB - OBJECTIVES/HYPOTHESIS: The objective was to determine the effect of approach, middle fossa versus retrosigmoid, on the hearing and facial nerve outcome of surgery for small vestibular schwannomas. STUDY DESIGN: The study had two parts, a case study of patient data entered into a prospectively designed database at the author's institution, and a meta-analysis of similar published data. METHODS: There were 73 of the author's private practice patients who met the inclusion criteria of intracanalicular vestibular schwannoma and total tumor removal by a retrosigmoid approach. American Academy of Otolaryngology-Head and Neck Surgery standardized hearing and facial nerve classifications of these patients and similar data from 11 other institutions were used to compare results of the two surgical approaches. RESULTS: Median facial nerve results for all institutions were significantly better with the retrosigmoid approach (grade I: 95% for retrosigmoid and 81% for middle fossa). Median hearing results trended toward better outcome with the middle fossa approach (same preoperative hearing class: 48% for middle fossa and 39% for retrosigmoid). Individual institution had an equal or greater effect on outcome than the choice of surgical approach. CONCLUSION: Surgical team accounted for more variability in hearing and facial nerve outcome than did approach. Retrosigmoid approach yielded significantly better facial nerve outcome. The trend toward better hearing outcome with the middle fossa approach may never achieve statistical significance across institutions because of high variability among surgical teams and small numbers of teams reporting results. PMID- 15280728 TI - Surgical outcomes in patients with endolymphatic sac tumors. AB - OBJECTIVE: To determine the surgical outcomes in patients with endolymphatic sac tumors (ELSTs). STUDY DESIGN: Retrospective review of patients at a referral based otology-neurotology practice. METHODS: A review of the records from the House Ear Clinic revealed 16 patients treated for ELSTs from 1971 to 2002. This article reports the treatment outcomes for the 14 patients for whom clinical data were available. RESULTS: Sensorineural hearing loss, tinnitus, and dizziness were the most common presenting signs and symptoms. Six patients presented with facial weakness, and three patients had symptoms characteristic of Meniere's syndrome. One patient suffered from Von Hippel-Lindau disease. Patients underwent microsurgical removal and were followed for an average of 59.6 months. Patients that presented with normal facial function maintained excellent postoperative function, and hearing was preserved in two patients with small tumors. Two patients suffered persistent, progressive disease despite multiple attempts at microsurgical removal and radiotherapy. Both had incomplete resections of their initial tumors. A third patient developed a small recurrent tumor that was successfully managed by a second attempt at microsurgical removal. CONCLUSIONS: Taken together with other reports, these results suggest that ELSTs are best managed by complete surgical resection. This can generally be accomplished with minimal additional morbidity. PMID- 15280727 TI - Advantages of cochlear implantation in prelingual deaf children before 2 years of age when compared with later implantation. AB - OBJECTIVES: To compare the auditory abilities and speech performance of children with a profound prelingual bilateral hearing-impairment when subjected to a cochlear implant (CI) before or after 2 years of age. To analyze the complications that arose during, or as a result of, the implantation process in these groups. DESIGN: Prospective cohort single-subject, repeated-measures study of children with profound bilateral hearing impairment subjected to CI. SETTING: Tertiary referral center with a program of pediatric CI from 1991. PATIENTS: This study analyzed 130 children subjected to multichannel CI for profound prelingual bilateral hearing-impairment in two age groups: 0 to 2 (n = 36) and 2 to 6 years of age (n = 94). INTERVENTIONS: The children were evaluated before, and each year after, the intervention (for up to 5 years) with both closed-set and open-set auditory and speech perception tests. Their speech ability was evaluated according to the Peabody Picture Vocabulary and Reynell general oral expression scales. RESULTS: Auditory and speech perception tests improved significantly in all children after CI, regardless of the follow-up time. The infant's performance was better the earlier the implant was performed. Speech tests showed that the development of children treated before 2 years of age was similar to normal children, and no additional complications were observed when compared with CI in older children. CONCLUSIONS: When performed before 2 years of age, CI offers a quicker and better improvement of performance without augmenting the complications associated with such an intervention. PMID- 15280729 TI - Transnasal endoscopic repair of cerebrospinal fluid rhinorrhea and skull base defect: ten-year experience. AB - OBJECTIVES/HYPOTHESIS: Many reports have advocated the feasibility of using an endoscope for the treatment of cerebrospinal fluid (CSF) rhinorrhea and skull base defect, and diversified endoscopic techniques and repairing materials have recently been proposed. This study determined the effectiveness of endoscopic repair of CSF leaks and interpreted the indications of the lumbar drain. STUDY DESIGN: Retrospective follow-up. METHODS: A total of 39 patients with CSF rhinorrhea (20 traumatic, 13 iatrogenic, and 6 spontaneous) were treated in one institution over a 10-year period. An underlay procedure was used to make eight (20.5%) repairs by way of a turbinate composite graft (4 patients), turbinate mucosa graft (3 patients), or the Dura Substitute (Preclude) (1 patient). A free turbinate graft with the applied fibrin glue was used to repair the defect by way of an overlay procedure in 23 (59.0%) lesions. Abdominal fat was used to repair the other eight (20.5%) lesions. RESULTS: Defects in the cribriform plate and anterior ethmoid sinus comprised the majority (61.5%) of all lesions. The successful rates for overlay, underlay, and fat obliteration procedures were 91.3% (21/23), 87.5% (7/8), and 100% (8/8), respectively. There were no statistical difference between underlay and overlay techniques (P = .792, Student's t test). Lumbar drainage was performed in 18 of 39 (46.2%) cases. Most patients required nasal packing (89.7%) and prophylactic parenteral antibiotics (97.4%). CSF rhinorrhea was resolved during the first attempt in 36 of 39 (92.3%) patients. All leaks were successfully repaired after a second attempt. No major complications were encountered. CONCLUSION: The endoscopic approach is safe and effective for the treatment of CSF rhinorrhea, even in the cases not successfully treated by previous neurosurgical approaches. Lumbar drain was suggested for defects in the frontal and sphenoid sinus and defects associated with meningocele or encephalocele. Complete exposure of the defect, appropriate selection of a fitting graft, as well as accurate placement and stabilization of the graft are critical to the success of repair. PMID- 15280730 TI - Treatment of a proximal accessory nerve injury with nerve transfer. AB - OBJECTIVE AND HYPOTHESIS: This study presents a case report of a patient who sustained an iatrogenic proximal accessory nerve injury that was treated with a medial pectoral to accessory nerve transfer. STUDY DESIGN: Case study. MATERIALS AND METHODS: Chart of one patient who was treated with a medial pectoral to accessory nerve transfer was reviewed. RESULTS: Five months after excision of a branchial cyst that resulted in a very proximal injury to the accessory nerve, this patient underwent a medial pectoral to accessory nerve transfer. At final follow-up, 3 years after surgery, the patient had full abduction overhead with some residual shoulder/scapular discomfort and mild scapular winging. CONCLUSION: The medial pectoral to accessory nerve transfer provides a viable surgical option with good reinnervation of the trapezius muscle in patients with a proximal accessory nerve injury where standard nerve repair or graft techniques are not feasible. PMID- 15280731 TI - Impact of anterior commissure involvement on local control of early glottic carcinoma treated by laser microresection. AB - OBJECTIVES: To analyze the impact of anterior commissure involvement on local control, survival, and laryngeal preservation in patients with early glottic cancer (pT1a-pT2a) treated with unimodality laser microsurgical resection. STUDY DESIGN: Retrospective review of 263 patients with early glottic cancer treated between 1986 and 1996. METHODS: Data on local control and overall survival rates were analyzed and calculated by the Kaplan-Meier method; the larynx preservation rates were given absolutely. RESULTS: Among 158 patients presenting with T1a glottic cancer, the anterior commissure was involved in 28 cases; the 5-year local control rate was 84%, and the larynx preservation rate was 93%. In the 130 cases without anterior commissure involvement, the 5-year local control rate was 90.0% and the corresponding larynx preservation rate 99%. In the T1b category consisting of 30 patients, anterior commissure involvement was observed in 16 patients; the 5-year local control rate was 73%, and the larynx preservation rate was 88%. In the 14 cases without anterior commissure involvement, the 5-year local control rate was 92% and the corresponding larynx preservation rate 100%. Seventy-five patients had T2a glottic carcinomas, with normal vocal cord movement. The anterior commissure was involved in 45 cases; the 5-year local control rate was 79%, and the larynx preservation rate was 93%. In the 30 cases without anterior commissure involvement, the 5-year local control rate was 74.0% and the corresponding larynx preservation rate 97%. CONCLUSIONS: This study shows the effectiveness of laser microsurgery for early glottic carcinoma regardless of anterior commissure involvement at presentation. This method can be performed as an outpatient procedure, even when conducting reresections. PMID- 15280732 TI - Medicolegal analysis of injury during endoscopic sinus surgery. AB - OBJECTIVES: To survey the causes, characteristics, and outcomes of malpractice litigation resulting from injuries sustained during endoscopic sinus surgery (ESS). STUDY DESIGN: A retrospective analysis of United States state and federal civil litigation involving injuries resulting from ESS. METHODS: Sources were state and federal court decisions and jury verdict reports accessed through a computerized legal database. The 41 cases were decided or settled between 1990 and 2003. The cases and reports were analyzed for pertinent data regarding plaintiffs, defendants, allegations of wrongdoing, resulting injury, expert witnesses, and resulting verdict or settlement. Correlation between severity of injury and case outcome was analyzed. RESULTS: All suits reviewed involved ESS. Many cases included multiple causes of action, or types of malpractice, including negligent technique, 31 (76%); lack of informed consent, 15 (37%); and wrongful death, 2 (5%). The defendant-physician specialty was overwhelmingly otolaryngology, 40 (98%). The most common presenting complaint, or indication for surgery, was chronic sinusitis, 30 (73%). The injuries caused by surgery were frequently multiple, including cerebrospinal fluid (CSF) leak, 10 (24%); brain damage, 6 (15%); diplopia, 7 (17%); and death, 2 (5%). The majority of cases reviewed (83%) resulted in a verdict rather than settlement. The result of the verdict or settlement was 17 (41%) in favor of the plaintiff, 23 (56%) in favor of the defendant, and 1 (2%) unknown. The average award was 751,275 dollars, with a median of 410,239 dollars and a range of 61,000 dollars to 2,870,000 dollars. CONCLUSIONS: This is the first study to review malpractice litigation resulting from injuries sustained during ESS and shows a hitherto unexpected pattern between severity of injury and case outcome. PMID- 15280733 TI - Unbiased quantification of the microdissected human Scarpa's ganglion neurons. AB - OBJECTIVE: The objective of the present study was to obtain unbiased estimates of the total number of Scarpa's vestibular ganglion neurons in individuals with normal vestibular function. STUDY DESIGN: Application of unbiased stereology using microdissected human temporal bone specimens. METHODS: Postmortem temporal bones were obtained from five young subjects with no history of audiovestibular disease (age range 42-49 years). The vestibular nerve containing the Scarpa's ganglion was microdissected, embedded in paraffin, and cut into 40 microm serial sections. Unbiased estimates of the total number of neurons were obtained using the optical fractionator technique of stereology. RESULTS: An average of 23,599 (coefficient of variation [CV] = 0.11) vestibular ganglion neurons was obtained. There was no significant difference between the results obtained from the microdissected specimens and results that had been obtained from an age-matched group derived from a previously published report from our laboratory using archival human temporal bone specimens. CONCLUSIONS: This study represents the first report to demonstrate the combination of the microdissection technique and the unbiased stereologic technique in the human temporal bone. This study demonstrates the reliability of the microdissection technique as an alternative method of human temporal bone processing for unbiased stereology. The utility of the microdissection technique is that specimens can be used for quantification, immunohistochemistry, and other powerful applications. PMID- 15280734 TI - Role of cytomegalovirus, Epstein-Barr virus, and human herpes virus-8 in benign lymphoepithelial cysts of the parotid gland. AB - OBJECTIVE: To provide background and evaluate the role of herpesviruses in benign lymphoepithelial cysts (BLC) of the parotid gland. STUDY DESIGN: Case series derived from review of pathology specimens. METHODS: Radiolabeled polymerase chain reaction (PCR) analysis was used to detect for the presence of cytomegalovirus (CMV), Epstein-Barr virus (EBV), and human herpes virus 8 (HHV-8) DNA sequences in 14 paraffin embedded specimens and 1 freshly aspirated BLC specimen. Thirteen normal parotid tissue specimens obtained from paraffin embedded blocks were used as a control group. RESULTS: CMV was detected with nearly equal frequency between the two groups (23% of normal vs. 20% in BLC). HHV 8 was found in 13% of the BLC group and in none of the normal group (P =.4841). There was significant difference in EBV detection between the normal (0%) and the BLC (33%) groups (P =.0437). CONCLUSION: CMV and HHV-8 does not appear to be associated with BLCs. Although EBV is found more frequently in BLC than in normal parotid controls, further studies are needed to elucidate the role of this virus in BLC pathogenesis. PMID- 15280735 TI - Temporary intermaxillary fixation using individualized acrylic splints permits image-data-based surgery of the lower jaw and oropharynx. AB - OBJECTIVES/HYPOTHESIS: Image-data-based surgical navigation is used as a helpful device in the operating room to localize critical structures with a high degree of accuracy. It also enables physicians to plan therapeutic performance. Because it relies on preoperatively acquired computed tomography (CT) or magnetic resonance imaging (MRI) data, there is restricted access for navigation of surgical instruments in areas that show motion uncorrelated with radiologic data. Thus, in the case of moveable structures, for example the lower jaw, navigational procedures could not yet be applied. STUDY DESIGN: We introduce a new technique using individualized intermaxillary splints that fix the mandible in a reproducible aboccluded position at the time of image-data acquisition and surgery. METHODS: Different manufacturing processes were investigated. The feasibility of uni- and bilateral intermaxillary splints was studied under clinical conditions in four patients during different procedures in the mandibular and oropharyngeal regions. RESULTS: The manufacturing of the splints showed was easily performed in a short time. With bilateral fixation, there was a high anatomic target precision of 1.6 to 2.3 mm. CONCLUSIONS: The use of bilateral intermaxillary splints that fix the patient's mandible in a reproducible aboccluded position permits an image-data-based navigated surgical approach to the oropharyngeal and mandibular regions. PMID- 15280736 TI - Inner ear decompression illness. PMID- 15280738 TI - Long-term, tube-free (permanent) tracheostomy in morbidly obese patients. PMID- 15280740 TI - Acoustic tensor tympani response and vestibular-evoked myogenic potential. PMID- 15280743 TI - Isolation, purification, and characterization of poly-N-acetyl glucosamine use as a hemostatic agent. AB - BACKGROUND: A new polymeric material, poly-N-acetyl glucosamine (p-GlcNAc) fiber, has been identified and is effective in achieving hemostasis in surgical procedures and trauma. The p-GlcNAc material is purified from large-scale cultures of a marine microalga. METHODS: Poly-N-acetyl glucosamine materials have been formulated as films, sponges, gels, and microspheres. The polymer's structure has been characterized by chemical composition, carbohydrate analysis, spectroscopic techniques, intrinsic viscosity, and electron microscopy. RESULTS: Carbohydrate analyses indicate that the primary sugar present in p-GlcNAc is N acetyl glucosamine. Elemental analyses yield percentage values for carbon, nitrogen, and hydrogen that support that the polymer is fully acetylated. Molecular weight determinations indicate that the polymer has a molecular weight of 2.0 x 10(6) Da. Fourier transform infrared, nuclear magnetic resonance, and circular dichroism spectral data have defined a unique tertiary structure. Biologic testing demonstrated that p-GlcNAc materials are fully biocompatible. CONCLUSION: The p-GlcNAc fiber has a unique beta-tertiary structure. PMID- 15280744 TI - Poly-N-acetyl glucosamine-mediated red blood cell interactions. AB - BACKGROUND: Investigations were performed to assess the effect of poly-N-acetyl glucosamine (p-GlcNAc) fiber slurry-mediated hemostasis by interactions with red blood cells. METHODS: Red blood cell aggregation studies were performed using test material-coated microscope slides and multiphoton microscopic measurements. Enzymatic removal of red blood cell surface proteins was achieved using trypsin and neuraminidase treatment. Zeta-potential measurements (surface charge) were performed. RESULTS: Red blood cells interact directly with poly-N-acetyl glucosamine polymers through ionic interactions and cell-surface proteins. The effective concentration of poly-N-acetyl glucosamine fiber material for 50% red blood cell aggregation was 0.28 mg/mL. The p-GlcNAc beta-configuration fibers and an alpha-configuration structural modification of the fibers both produced maximal responses because of their zeta-potentials, whereas other chemically modified p-GlcNAcs and chitosans were ineffective. CONCLUSION: Poly-N-acetyl glucosamine-induced red blood cell aggregation is mediated by interactions with red blood cell surface charges. PMID- 15280745 TI - Mechanisms of poly-N-acetyl glucosamine polymer-mediated hemostasis: platelet interactions. AB - BACKGROUND: Investigations were performed to determine whether poly-N-acetyl glucosamine (p-GlcNAc) induces hemostasis by the activation of platelets. METHODS: Platelets were isolated from human blood, fixed in the presence poly-N acetyl glucosamine fibers, and visualized with scanning electron microscopy. Platelet activation surface markers were measured by fluorescence multiphoton microscopy. Platelet aggregation in the presence of p-GlcNAc fibers and integrin receptor blockers was measured. RESULTS: Scanning electron microscopy indicated that contact of platelets with poly-N-acetyl glucosamine fibers resulted in platelet activation. Fluorescent microscopy showed that contact of platelets with the marine polymer increased intracellular levels of free calcium and resulted in surface exposure of platelet phosphatidylserine, P selectin, and the alphaIIbbeta3 integrin. Antibody inhibitors of the platelet alphaIIbbeta3 integrin inhibited p-GlcNAc to stimulate fibrin polymerization. CONCLUSION: Poly N-acetyl glucosamine fiber material promotes hemostasis by the activation of platelets. PMID- 15280746 TI - In vitro effects of poly-N-acetyl glucosamine on the activation of platelets in platelet-rich plasma with and without red blood cells. AB - BACKGROUND: This study was performed to assess the effect of poly-N-acetyl glucosamine fiber slurry on plasma clotting proteins, platelets, and red blood cells in the clotting of the blood. METHODS: Citrate phosphate dextrose whole blood was stored at 22degreesC for 48 hours to prepare platelet-poor plasma, platelet-rich plasma (PRP), and PRP plus red blood cells with hematocrit values of 20%, 35%, and 45% with and without an equal volume of poly-N-acetyl glucosamine fibers (1 mg/mL 0.9% NaCl). RESULTS: Thromboelastogram data show that poly-N-acetyl glucosamine fibers (p-GlcNAc) significantly reduced the R time in platelet-poor plasma, PRP, and PRP supplemented with red blood cells. Poly-N acetyl glucosamine fibers increased, but not significantly, Annexin V and factor X binding to platelets, platelet microparticles, and red blood cell Annexin V binding. Poly-N-acetyl glucosamine fibers increased the production of thromboxane B2 by PRP. CONCLUSION: Poly-N-acetyl glucosamine slurry activates platelets. PMID- 15280747 TI - Application of the poly-N-acetyl glucosamine-derived rapid deployment hemostat trauma dressing in severe/lethal Swine hemorrhage trauma models. AB - BACKGROUND: The Rapid Deployment Hemostat (RDH) Bandage was developed for the rapid control of bleeding caused by trauma. METHODS: An extremity wound involving skin, muscle, bone, and femoral arterial injury and a 1-cm vertical incision in the abdominal aorta in swine were studied to compare the RDH Bandage, a fibrin sealant dressing and gauze to restore hemostasis. The total blood loss was determined and the survival of animals was measured. RESULTS: In the extremity injury model, the RDH Bandage reduced blood loss by 63% compared with the gauze control. In the aorta arterial incision model, the RDH Bandage required a significantly lower compression time to control bleeding compared with gauze and TachoComb. The RDH Bandage was able to stop bleeding from this injury in 100% of the tests. CONCLUSION: The RDH Bandage was superior to a commercially available fibrin bandage in controlling hemorrhage, decreasing blood loss, and increasing survival. PMID- 15280748 TI - Comparison of poly-N-acetyl glucosamine with commercially available topical hemostats for achieving hemostasis in coagulopathic models of splenic hemorrhage. AB - BACKGROUND: The hemostatic quality of the poly-N-acetyl glucosamine (p-GlcNAc) patch was compared with a fibrin sealant, fibrin bandage, and cellulose patch. METHODS: A 2 x 2-cm capsular strip to a depth of 3 mm of the swine spleen was used as a source of bleeding. Splenic lacerations were created in hemophilia B dogs and treated with p-GlcNAc and Surgicel. Wounds were created in rabbits and treated with p-GlcNAc at 37degreesC and after keeping body core temperature at 29degreesC. RESULTS: Poly-N-acetyl glucosamine was able to achieve hemostasis with greater efficacy than either of the fibrin-based bandages. In the hemophilia B dog study, p-GlcNAc significantly outperformed Surgicel, with p-GlcNAc achieving hemostasis in 75% of the treated wounds compared with 17% for the cellulose patch. The hypothermia study demonstrated that p-GlcNAc is equally effective at 29degreesC and at 37degreesC. CONCLUSION: Poly-N-acetyl glucosamine was effective at controlling bleeding in animals with experimentally induced or genetic coagulopathic disorders. PMID- 15280749 TI - Randomized double-blind studies of polysaccharide gel compared with glue and other agents for hemostasis of large veins and bleeding canine esophageal or gastric varices. AB - BACKGROUND: The safety and efficacy of poly-N-acetyl glucosamine (p-GlcNAc) gels were compared with standard agents in three different dog studies to assess abdominal venous collaterals, bleeding esophageal varices, and bleeding gastric varices. METHODS: Adult dogs with prehepatic portal hypertension and large abdominal venous collaterals, esophageal varices, or gastric varices were studied. RESULTS: Significantly higher sclerosis rates were seen with F2 or F4 p GlcNAc gels and standard sclerosants. F2 and F4 gels had high rates of permanent hemostasis, low rates of secondary ulceration, and significant reductions in esophageal and gastric variceal size. These results were either equivalent to or significantly better than the most commonly used gastric varix hemostatic agent (glue) or other sclerosing agents. CONCLUSION: F2 and F4 poly-N-acetyl glucosamine gels are promising therapeutic agents for venous and variceal hemostasis. PMID- 15280750 TI - Evaluation of poly-N-acetyl glucosamine as a hemostatic agent in patients undergoing cardiac catheterization: a double-blind, randomized study. AB - BACKGROUND: This is the first blinded, randomized, placebo-controlled clinical trial to evaluate the efficacy of poly-N-acetyl glucosamine (p-GLcNAc) in improving hemostasis in patients undergoing cardiac catheterization. METHODS: Patients were randomly assigned to have either a placebo-treated (n = 17) or a p GlcNAc-treated (n = 16) 3 x 3-cm patch topically placed at the femoral insertion site at the completion of their catheterization procedure with a mechanical pressure clamp applied over it. The amount of pressure was measured. RESULTS: Although the placebo group had slightly higher clamp pressure applied to the femoral arterial puncture site at the end of the catheterization procedure (189 +/- 47 vs. 149 +/- 49 mm Hg, p = 0.042), the time to effective hemostasis (16 +/- 7 vs. 10 +/- 3 minutes, p = 0.01) was decreased in the p-GlcNAc group by 37%. CONCLUSION: The application of p-GlcNAc patches improved hemostasis at the arterial puncture site in patients undergoing cardiac catheterization. PMID- 15280751 TI - Hemostasis in the absence of clotting factors. AB - BACKGROUND: This study tests whether the hemostatic action of poly-N-acetyl glucosamine (p-GlcNAc) fiber material involves vasoconstrictor release leading to closure of an aortic laceration. METHODS: A 22-gauge cannula was inserted into an infrarenal aortic segment of a rat. Surrounding ligatures were tied, and the aorta was flushed with 60 mL of saline from a reservoir held at 80 cm. A 23-gauge aortic puncture was made. The time taken to empty the reservoir was recorded. RESULTS: Control patches led to an emptying time of 295 seconds, whereas p-GlcNAc patches increased this time to greater than 600 seconds. Ten minutes after patch removal, the emptying time decreased to 330 seconds. The rats were treated intravenously with endothelin receptor antagonists BQ-485 or JKC-301. The emptying time shortened to control values, despite the use of the p-GlcNAc fiber based patch. CONCLUSION: The mechanism of hemostasis by poly-N-acetyl glucosamine involves endothelin release independent of formed elements of blood. PMID- 15280752 TI - Treatment of thoracolumbar burst fractures with variable screw placement or Isola instrumentation and arthrodesis: case series and literature review. AB - BACKGROUND: The controversy of burst fracture surgical management is addressed in this retrospective case study and literature review. METHODS: The series consisted of 40 consecutive patients, index included, with 41 fractures treated with stiff, limited segment transpedicular bone-anchored instrumentation and arthrodesis from 1987 through 1994. RESULTS: No major acute complications such as death, paralysis, or infection occurred. For the 30 fractures with pre- and postoperative computed tomography studies, spinal canal compromise was 61% and 32%, respectively. Neurologic function improved in 7 of 14 patients (50%) and did not worsen in any. The principal problem encountered was screw breakage, which occurred in 16 of the 41 (39%) instrumented fractures. As we have previously reported, transpedicular anterior bone graft augmentation significantly decreased variable screw placement (VSP) implant breakage. However, it did not prevent Isola implant breakage in two-motion segment constructs. Compared with VSP, Isola provided better sagittal plane realignment and constructs that have been found to be significantly stiffer. Unplanned reoperation was necessary in 9 of the 40 patients (23%). At 1- and 2-year follow-up, 95% and 79% of patients were available for study, and a satisfactory outcome was achieved in 84% and 79%, respectively. These satisfaction and reoperation rates are consistent with the literature of the time. CONCLUSIONS: Based on these observations and the loads to which implant constructs are exposed following posterior realignment and stabilization of burst fractures, we recommend that three- or four-motion segment constructs, rather than two motion, be used. To save valuable motion segments, planned construct shortening can be used. An alternative is sequential or staged anterior corpectomy and structural grafting. PMID- 15280753 TI - Cervical laminoplasty: use of a novel titanium plate to maintain canal expansion- surgical technique. AB - Cervical laminoplasty is a technique used to achieve spinal cord decompression in cases of myelopathy or myeloradiculopathy. The most common reason for failure of this technique is restenosis due to hinge closure. Various techniques have been employed to hold the laminar "door" open while the body heals the lamina hinge in the new expanded position. Ideally, a method of achieving laminar fixation should be technically straightforward, provide secure laminar fixation, and be rapid to minimize the risk of iatrogenic injuries, blood loss, and operative time. The authors describe the use of a novel plate designed to accomplish these goals. The technical issues relevant to performing the laminoplasty and securing the laminae are discussed. The plate has been proven biomechanically to be equal or superior to the currently used techniques. The use of this plate will allow the patient to engage in an early active rehabilitation protocol-while minimizing the risk of restenosis of the canal. This may ultimately lead to better preservation of motion and decreased axial neck pain following laminoplasty. PMID- 15280754 TI - Biomechanical study of lumbar pedicle screws in a corpectomy model assessing significance of screw height. AB - BACKGROUND: We tested the hypothesis that a pedicle screw construct's height is an important factor in strengthening a screw-rod system. METHODS: Six corpectomy constructs were made, each using two ultra-high-molecular-weight polyethylene blocks, 6.5-mm pedicle screws, and two 6.35-mm rods. Pedicle screws were placed at +10-, +5-, 0-, and -5-mm depths in relation to the dorsal surface of the corpectomy model. Nondestructive testing was performed in flexion/extension and in torsion. RESULTS: For all modes tested, the screw-rod constructs continued to increase in stiffness as the height of the construct was lowered, and this was statistically significant at all heights tested (P < 0.001). The stiffness increased 232% when comparing flexion at +10 and -5 mm and increased 231% in extension from +10 to -5 mm. The torsional stiffness increased 171% when comparing +10 and -5 mm. CONCLUSIONS: Thus, lower-profile instrumentation systems should be used to take advantage of this by decreasing the size and bulkiness of the implants while increasing the strength of the construct. PMID- 15280755 TI - Probing for thoracic pedicle screw tract violation(s): is it valid? AB - BACKGROUND: Preparation of the thoracic pedicle screw tract is a critical step prior to the placement of screws. The ability to detect pedicle wall violation(s) by probing prior to insertion of thoracic pedicles screws, however, has not been studied. The purpose of this study was to evaluate the inter- and intraobserver agreement and the accuracy in detecting thoracic pedicle screw tract violation(s) among surgeons at various levels of training. METHODS: With use of a straightforward trajectory, under direct visualization, 108 thoracic pedicle screw tracts (54 cadaveric thoracic vertebrae) were prepared in a standard fashion, followed by tapping with a 4.5-mm cannulated tap. A deliberate pedicle violation was randomly created by an independent investigator in either the anterior, the medial, or the lateral wall in 65 pedicles. Following this, four blinded, independent surgeons at various levels of training probed the specimens on three separate occasions to determine if a breach was present (1,296 discrete data points). Surgeon findings were then recorded as breach present or absent and, if present, breach location. The Cohen kappa correlation coefficient (kappa a) and 95% confidence interval were used to assess the accuracy of the observers and the inter- and intraobserver agreement. RESULTS: The mean accuracy over three iterations, the validity in detecting the breach location, and the intraobserver agreement varied by level of training and experience, with the most experienced observer (observer 1) scoring the best and the least experienced observer (observer 4) scoring the worst. The three most senior surgeons had good intraobserver agreement. Interobserver agreement was low between the four observers. CONCLUSIONS: An observer's ability to accurately detect the presence or absence of a pedicle tract violation and the breach location, if present, is dependent on the surgeon's level of training. Probing the pedicle tract prior to placement of pedicle screws in the thoracic spine is likely a learned skill that improves with repetition and experience. PMID- 15280756 TI - Lumbar instability and clinical symptoms: which is the more critical factor for symptoms: sagittal translation or segment angulation? AB - BACKGROUND: The relationship between radiologic instability and its symptoms is controversial. Previous authors consider flexion-extension radiographs to be of little value in evaluating instability; however, the current authors consider the variation of results in evaluating radiologic instability to be the result of limitations in previous researchers' methods. METHODS: In this report, sagittal translation and angulation at the L4-L5 segment were measured in flexion extension films in 1,090 outpatients with low back and/or leg pain using a three landmark measuring method. The symptoms of four groups with and without 3-mm translation and with and without 10 degrees angulation were compared for all the patients and for 280 age-matched patients using a scoring system. The age-matched patients were followed up for 4.6 years. RESULTS: Results showed that patients with > or = 3-mm translation had significantly lower scores, indicating a limitation in their daily activities due to pain, than patients < 3-mm translation; however, no difference was observed between the groups in terms of angulation. The group with > or = 3-mm translation and > or = 10 degrees angulation significantly demonstrated the lowest scores at both evaluations during the initial visit and follow-up. This group had been suffering from low back and/or leg pain the longest and had visited the hospital significantly more often than other groups. CONCLUSION: In conclusion, translation of the lumbar segment has a greater influence than angulation on lumbar symptoms. The presence of both radiologic factors could be an indicator for persistence of the symptoms. PMID- 15280757 TI - Vertebral reconstruction with biodegradable calcium phosphate cement in the treatment of osteoporotic vertebral compression fracture using instrumentation. AB - OBJECTIVE: To assess the efficacy of posterior instrumentation and vertebral reconstruction with biodegradable calcium phosphate cement (CPC) in the treatment of osteoporotic vertebral compression fracture with neurologic deficit. BACKGROUND: Vertebroplasty consists of the injection of polymethylmethacrylate (PMMA) cement into the vertebral body. While PMMA has high mechanical strength, it cures fast and thus allows only a short handling time. Other potential problems of using PMMA injection may include damage to surrounding tissues due to the high polymerization temperature or by the toxic unreacted monomer and the lack of long-term biocompatibility. Bone mineral cements such as calcium carbonate and CPCs have a longer working time and low thermal effect. They are also biodegradable while providing good mechanical strength. However, the viscosity of injectable mineral cements is high, and the infiltration of these cements into the vertebral body has been questioned. Recently, the infiltration properties of CPC have been significantly improved, making it more suitable for injection into the vertebral bodies for vertebral reconstruction. METHODS: Five patients were included in this open prospective study. Inclusion criteria were delayed collapsed vertebral compression fractures responsible for severe pain and neurologic dysfunction necessitating posterior decompression surgery. Of five patients, two were male and three were female with an average age at surgery of 80.4 years (71-85 years) and an average duration of follow-up of 2.5 years (2-3.5 years). Evaluation of clinical data was based on x-ray, Japanese Orthopaedic Association (JOA) score for low back pain (full score is 29 points), and Visual Analog Scale (VAS). RESULTS: The levels of the delayed collapsed vertebrae were T10, L1, and L2 (for one patient each) and L4 (two patients). All patients were in poor condition, for example, renal failure, heart failure, and chronic hepatitis. The average operative time was 2 hours (1 hour 36 minutes to 2 hours 16 minutes), and intraoperative bleeding was 181 mL (85-236 mL). As for clinical symptoms, preoperative JOA score averaged 17.8 points and was improved to 26 points postoperatively, while the preoperative VAS score of 8.6 points improved to 2 points postoperatively. Morphologic evaluation showed preoperative vertebral compression ratio averaged 41% and improved to 74% immediately after the operation and finally settled at 68%. Just one of five cases experienced late vertebral collapse 3 months after the operation. CONCLUSION: Vertebral reconstruction with biodegradable CPC in the treatment of osteoporotic vertebral compression fracture using instrumentation was a safe and useful surgical treatment. PMID- 15280758 TI - Outcome of total en bloc spondylectomy for solitary metastasis of the thoracolumbar spine. AB - BACKGROUND: Total en bloc spondylectomy (TES) was devised to minimize the incidence of local recurrence following resection of spinal tumor. Successful local control with TES has been reported for patients with primary malignant or aggressive benign spinal tumors. As for metastatic spinal tumors, however, only a few surgeons except for the inventor group have reported the outcome of TES. The purpose of this study was to investigate whether TES could provide radical resection of the tumor in patients with solitary spinal metastases. METHODS: Twelve patients underwent TES for a solitary metastatic tumor of the thoracolumbar spine. Primary malignancies included breast cancer in four patients, thyroid cancer in three, renal cell carcinoma in three, lung cancer in one, and unknown in one. All patients were regularly followed up with plain radiographs, computed tomography scans, and magnetic resonance imaging to detect local recurrence. RESULTS: In two of the four cases with paraspinal tumor extension, local recurrences developed at 25 months after surgery. Seven patients have survived for an average of 61 months, while the remaining five died of disseminated metastases with a mean survival of 23 months after surgery. Local recurrences were common in patients with paraspinal extensions. TES for lesions with paraspinal extensions failed to provide curative resection of the tumor. CONCLUSION: Given the great technical demands and potential risks of TES, the indication for TES with spinal metastases should be limited to cases with solitary lesions that do not extend to the paraspinal area. PMID- 15280759 TI - Measurement variability in the assessment of sagittal alignment of the cervical spine: a comparison of the gore and cobb methods. AB - BACKGROUND: Reconstructive procedures of the cervical spine are being performed with increasing frequency. Maintenance of physiologic sagittal alignment is an essential component of reconstructive procedures of the spine. Two methods exist for measuring sagittal alignment in the cervical spine: the Gore and Cobb methods. An experimental study comparing Gore and Cobb measurement techniques for nonspondylotic and spondylotic cervical spines was conducted. The objectives were to assess the intra- and interobserver variability of both the Gore and the Cobb methods of measurement to determine the most reproducible technique for assessing sagittal alignment of the cervical spine. METHODS: With use of C3 and C7 as the end vertebrae, lateral radiographs of 20 nonspondylotic (group 1) and 20 spondylotic (group 2) cervical spines were measured by the Gore and Cobb methods on three different occasions by three orthopaedic surgeons with different levels of experience. RESULTS: For group 1, there was less intra- and interobserver variability for the Gore method than for the Cobb method (P < 0.05). Group 2 measurements were also less variable for the Gore method, although this was not statistically significant. Pooling all three observers, 95% confidence limits for intra- and inter-observer variability for the Gore method were 3 degrees and 6 degrees for group 1 and 4 degrees and 7 degrees for group 2, respectively. For the Cobb method, corresponding values were 4 degrees and 9 degrees for group 1 and 5 degrees and 9 degrees for group 2. Overall, intraobserver measurements were less variable than interobserver measurements (P < 0.01). There were no significant differences in variability based on experience level. CONCLUSION: Measurements of cervical spine sagittal alignment by the Gore method are more reproducible than by the Cobb method. PMID- 15280760 TI - Biomechanical evaluation of relationship of screw pullout strength, insertional torque, and bone mineral density in the cervical spine. AB - BACKGROUND: Understanding of implant failure mechanisms is important in the successful utilization of anterior cervical plates. Many variables influence screw purchase, including the quality of the bone. The purpose of this study was to assess the relationship of screw pullout and screw insertional torque across a wide range of bone mineral densities (BMDs). METHODS: A total of 54 cervical vertebrae in 12 cervical spines were evaluated for BMD using dual-energy x-ray absorptiometry scanning. Actual and perceived peak torques of 3.5-mm anterior cervical screws were determined at each level followed by screw pullout strength testing. RESULTS: A high correlation was observed between screw pullout strength and BMD. However, there was a low correlation of peak insertional torque to pullout strength. CONCLUSION: These findings suggest the quality of the bone is more instrumental in the success or failure of anterior cervical screws than is the insertional torque with which the screws are placed. PMID- 15280761 TI - In vivo porcine intradiscal pressure as a function of external loading. AB - BACKGROUND: Spinal loading during daily activity as it relates to the ability of the intervertebral disc to sustain its integrity has been a major issue in spinal research. The purpose of this investigation was to establish the relationship between the intervertebral disc pressure in the nucleus and the load applied to the motion segment in an in vivo porcine model. METHODS: Nine domestic pigs were used in this study. A miniaturized servohydraulic testing machine was affixed to the lumbar spine via four intrapedicular screws, which were inserted bilaterally into the L2 and L3 vertebrae. A pressure needle was inserted through the lateral part of the L2-L3 disc annulus and into the nucleus pulposus. Force, deformation, and intradiscal pressure data were collected during a loading scheme that consisted of applying a set of constant loads in increasing order, that is, 50, 100, 150, 200, and 250 N. Each load was applied for 30 seconds followed by 30 second restitution. RESULTS: Intradiscal nucleus pressure was found to correlate to the applied load in all cases. Linear regression analyses resulted in the following equation: intradiscal pressure (MPa) = 0.08 + 1.25E(-3)(load, N), r(2) = 0.81, n = 8. Intradiscal pressure was also highly linearly dependent on the stress. The intrinsic intradiscal pressure was found to be 81 +/- 5 kPa. The results also indicated that the pressure within the disc exhibited a creep behavior. CONCLUSION: In conclusion, pressure in the nucleus of the porcine intervertebral disc was linearly related to the applied load and stress. PMID- 15280762 TI - Posterior cervical laminoplasty using a new plating system: technical note. AB - BACKGROUND: Laminoplasty is well described in the Japanese literature as a surgical option for treating ossification of the posterior longitudinal ligament (OPLL). The open door technique has gained increasing popularity in the United States and Europe to treat not only OPLL but also cervical stenotic myelopathy. An obstacle to its widespread use is the lack of a suitable fixation plate to adequately secure the fractured lamina to the lateral mass. Our objective was to demonstrate the advantages of a novel miniplate (Ti-Mesh LP system; Medtronic Sofamor Danek, Memphis, TN, USA) that is ideally suited for fixing the lamina to the lateral mass. METHODS: We used the Ti-Mesh LP miniplate system to perform laminoplasties on five patients, all male, with a mean preoperative Nurick score of 2.8. Four patients had congenital cervical stenosis with myelopathy and one had OPLL. Open door laminoplasties were performed on all patients. The plates were implanted with a claw positioned on the trapdoor lamina and a flat plate on the lateral mass. RESULTS: The system was implanted successfully in all patients. The mean number of levels fixated was 4.4. There were no intraoperative or postoperative complications after >5 months follow-up. CONCLUSIONS: The new Ti Mesh LP cranial miniplate and screw system facilitates posterior cervical laminoplasty procedures by eliminating the need to contour cranial miniplates for use in the cervical spine. Its unique claw construct and angled design are ideal for holding a trapdoor laminoplasty in the open position. We have used this system successfully and without complications to perform decompressive posterior cervical laminoplasties in five patients. PMID- 15280763 TI - Lumbar synovial cysts: a review of diagnosis, surgical management, and outcome assessment. AB - Synovial cysts of the lumbar spine contribute significantly to narrowing of the spinal canal and lateral thecal sac and nerve root compression. Cysts form as a result of arthrotic disruption of the facet joint, leading to degenerative spondylolisthesis in up to 40% of patients. Clinical findings and neurodiagnostic confirmation prompt surgical intervention consisting of varying decompressions with or without primary fusion. Most patients present in their mid-60s, with a male-to-female ratio varying from 2:1 to 1:1. Preoperative symptoms include low back pain, radiculopathy, and neurogenic claudication. Motor and sensory signs usually reflect the anatomic location of the synovial cyst and the level of resultant maximal lumbar stenosis. In descending order of frequency, they are typically found at the L4-L5, L5-S1, L3-L4, and L2-L3 levels. Lumbar synovial cyst surgery includes unilateral or bilateral laminotomies, hemilaminectomies, or laminectomies alone or in combination with in situ or instrumented fusion. Those patients undergoing decompression alone may postoperatively develop progression or the new appearance of olisthy, while those primarily fused rarely show further increase or a new onset of slip. Outcome measures spanning 1- to 2-year postoperative intervals frequently included surgeon-based rather than the current patient-based analysis, the lat-ter including the Medical Outcomes Trust Short Form-36. PMID- 15280764 TI - Spinal manifestations in a patient with congenital insensitivity to pain. AB - Spinal manifestations in congenital insensitivity to pain are relatively uncommon and easily misdiagnosed. We report on a patient with absent protective pain sensation, who developed spinal neuropathic arthropathy. At age 11 years, he presented with a destructive lesion at the L1-L2 level, causing him tingling sensation in both lower limbs. He was treated with combined anteroposterior spinal fusion from T12 to L3 and had full recovery. Five years later, he presented with a long history of clicking in his low back, muscle weakness and paresthesia in both lower extremities during walking, and evidence of Charcot arthropathy at the L4-L5 level, resulting in junctional kyphosis and canal narrowing. Posterior spinal arthrodesis from L3 to the sacrum was performed, due to lack of patient and parental consent for combined anterior decompression/posterior fusion. The patient resumed normal muscle function and his previous level of activities. Spinal complications should be anticipated in this condition and create diagnostic and therapeutic dilemmas. However, surgical management can produce favorable clinical results. PMID- 15280765 TI - An expanding cervical synovial cyst causing acute cervical radiculopathy. AB - Synovial cysts of the cervical spine occur infrequently in the spinal canal and are most often associated with degenerative facet joints. Despite the prevalence of degenerative spinal disease, symptomatic synovial cysts are extremely uncommon. We report a rare case that showed an exacerbation of a cervical radiculopathy due to an acute expansion of the synovial cyst. Magnetic resonance (MR) images originally revealed a small cystic extradural lesion when the patient presented with neck pain and slightly numbness in the right hand. The patient's complaints subsequently subsided after administration of pain killers. However, 2 weeks after this, the patient experienced a spontaneous, sudden, severe radiating pain into the right arm without any accompanying cervical injury. MR images showed that the cyst had become markedly increased in size in the intervening 4 weeks and compressed the spinal cord laterally. Because the arm pain was so severe and neurologic examinations revealed the paralysis of the C8 nerve root, the synovial cyst was excised surgically and a good clinical outcome achieved. Thus, even if symptoms are mild and the size of the synovial cyst is small, acute expansion of the cyst might be rarely observed and careful management, including surgical consideration, is needed. PMID- 15280766 TI - Cauda equina syndrome in lumbar spinal stenosis: case report and incidence in Jutland, Denmark. AB - BACKGROUND: Cauda equina syndrome in spinal stenosis is a rare complication. During a 5-year period (1996-2000), 340 cases of spinal stenosis were diagnosed in the County of South Jutland, Denmark. The annual incidence of spinal stenosis was 272 per million inhabitants. During that period, only one patient with acute cauda equina syndrome in spinal stenosis was diagnosed: that is, the patient described herein. METHODS AND RESULTS: A 74-year-old woman appeared with urinary retention and fecal incontinence for the previous 24 hours. Computed tomography scan showed spinal stenosis from L2 to L4. She underwent an urgent operation and recovered within 5 days from her anal sphincter paresis and within 5 weeks from her bladder paresis. PMID- 15280767 TI - Compartment syndrome of the thigh: an unusual complication after spinal surgery. AB - Compartment syndrome of the thigh is an uncommon pathology, and its acute presentation after spinal surgery is rare. Because a large muscle mass is involved and systemic manifestations of crush syndrome and altered mental status are present, such abnormalities may lead to a delay in appropriate diagnosis and subsequent treatment. A 56-year-old man who was suspected of having a posterior compartment syndrome in the thigh after spinal decompression in the knee-chest position was evaluated clinically and with the use of a catheter for intracompartmental pressure measurement as a tool to help establish the diagnosis and monitor the evolution. Because of sciatic involvement and a demonstrated increase in the pressure in the posterior compartment with myoglobinuria and acute renal failure, prompt longitudinal fasciotomy was performed with excellent neurologic recovery and improvement of both clinical parameters and mental status. PMID- 15280768 TI - A simple bone cyst located in the pedicle of the lumbar vertebra. AB - A simple bone cyst located in the spine is rare. In the current work, we have documented the first case of a simple bone cyst located in the pedicle of the lumbar vertebra. The patient was a 50-year-old woman with low back pain. Radiographs of the lumbar region of the spine showed a well circumscribed radiolucent lesion with surrounding bone sclerosis in the right pedicle of L3. Computed tomography (CT) showed that the right pedicle was completely occupied by the cyst and slightly expanded. The cystic lesion extended to both the vertebral body and the lamina. T1-weighted magnetic resonance imaging (MRI) showed uniform low intensity, and T2-weighted MRI showed uniform very high intensity. Signal enhancement by gadolinium diethylenetriaminepenta-acetic acid was not observed in the lesion. During the operation, the cavity of the cyst was filled with 2 mL of serosanguinous fluid. Subsequent histologic examination showed a thin layer of connective tissue in the inner surface of the cyst. PMID- 15280769 TI - Intradural extramedullary hemangioblastoma differentiated by MR images in the cervical spine: a case report and review of the literature. AB - We report a rare case of hemangioblastoma existing in the intradural extramedullary location diagnosed by magnetic resonance imaging (MRI) examination alone. A 46-year-old man gradually developed numbness in his lower extremities. MR images revealed a tumor shadow located posterior to the spinal cord at C5-C7. Small signal void shadows were continuously observed from C1 to C5 in the region cranial to the tumor, indicating the presence of enlarged vessels. Under microscopic observation, the tumor with accompanying vessels was resected totally via hemilaminectomy. The postoperative course was uneventful, and MR images obtained 4 months after the operative procedure demonstrated total removal of the tumor and the abnormal vessels. In this case, recognizing the abnormally enlarged vessels outside the tumor mass preoperatively led us to the correct diagnosis on MR images. PMID- 15280770 TI - Subarachnoid hematoma, hydrocephalus, and aseptic meningitis resulting from a high cervical myelogram. AB - Despite its reduced use since the advent of magnetic resonance imaging, the high cervical myelogram remains a common diagnostic test in the evaluation of patients whose symptoms suggest cervical stenosis. We report a case of subarachnoid hematoma, hydrocephalus, and aseptic meningitis after a high cervical myelogram. A 52-year-old woman presented with headache, slurred speech, worsened neck pain and stiffness, and diffuse extremity weakness leading to gait instability beginning several hours after a cervical myelogram. Computed tomography (CT) scan revealed a C1-C2 hematoma below the dura, blood in the fourth and lateral ventricles, and hydrocephalus. An external ventricular drain was placed, and cerebrospinal fluid profile was consistent with aseptic meningitis. A suboccipital craniectomy and C1-C2 laminectomies were performed, followed by a C1 C2 durotomy, which revealed a large subarachnoid blood clot in the region of the cisterna magna extending down to the upper aspect of C2, which was evacuated by incising the arachnoid. In the midline at C1, an active source of intramedullary arterial bleeding on the dorsal surface of the spinal cord was coagulated. Spinal subarachnoid hematoma is a rare complication of high cervical myelogram. The extension of blood into the ventricular system with associated hydrocephalus has never been previously reported after myelography. Thus, severe persistent pain after cervical myelography should be evaluated by CT scans of the brain and cervical spine. PMID- 15280771 TI - Identification of the minimal conserved structure of HIV-1 protease in the presence and absence of drug pressure. AB - OBJECTIVE: To define the extent of amino acid protease (PR) conservation in vivo in the absence and presence of pharmacological pressure in a large patient cohort. METHODS: Plasma-derived complete protein PR sequences from a well-defined cohort of 1096 HIV-1 infected individuals (457 drug-naive and 639 under antiretroviral therapy including PR-inhibitors) were obtained and analysed, and are discussed in a structural context. RESULTS: In naive patients, the PR sequence showed conservation (< 1% variability) in 68 out of 99 (69%) residues. Five large conserved regions were observed, one (P1-P9) at the N-terminal site, another (E21-V32) comprised the catalytic active-site, a third (P44-V56) contained the flap, a fourth contained the region G78-N88, and another (G94-F99) contained the C-terminal site. In PR-inhibitor treated patients, the appearance of mutations primarily associated with drug resistance determined a decrease of amino acid invariance to 45 out of 99 residues (45% conservation). The overall degree of enzyme conservation, when compared to the PR sequences in drug-naive patients, was preserved at the N- and C-terminal regions, whereas the other large conserved areas decreased to smaller domains containing, respectively, the active site residues D25-D29, the tip of the flap G49-G52, and the G78-P81 and G86-R87 turns. CONCLUSIONS: Amino acid conservation in HIV PR can be minimally present in 45 residues out of 99. Identification of these invariable residues, with crucial roles in dimer stability, protein flexibility and catalytic activity, and their mapping on the three-dimensional structure of the enzyme will help guide the design of novel resistance-evading drugs. PMID- 15280772 TI - Immune restoration disease after antiretroviral therapy. AB - Suppression of HIV replication by highly active antiretroviral therapy (HAART) often restores protective pathogen-specific immune responses, but in some patients the restored immune response is immunopathological and causes disease [immune restoration disease (IRD)]. Infections by mycobacteria, cryptococci, herpesviruses, hepatitis B and C virus, and JC virus are the most common pathogens associated with infectious IRD. Sarcoid IRD and autoimmune IRD occur less commonly. Infectious IRD presenting during the first 3 months of therapy appears to reflect an immune response against an active (often quiescent) infection by opportunistic pathogens whereas late IRD may result from an immune response against the antigens of non-viable pathogens. Data on the immunopathogenesis of IRD is limited but it suggests that immunopathogenic mechanisms are determined by the pathogen. For example, mycobacterial IRD is associated with delayed-type hypersensitivity responses to mycobacterial antigens whereas there is evidence of a CD8 T-cell response in herpesvirus IRD. Furthermore, the association of different cytokine gene polymorphisms with mycobacterial or herpesvirus IRD provides evidence of different pathogenic mechanisms as well as indicating a genetic susceptibility to IRD. Differentiation of IRD from an opportunistic infection is important because IRD indicates a successful, albeit undesirable, effect of HAART. It is also important to differentiate IRD from drug toxicity to avoid unnecessary cessation of HAART. The management of IRD often requires the use of anti-microbial and/or anti inflammatory therapy. Investigation of strategies to prevent IRD is a priority, particularly in developing countries, and requires the development of risk assessment methods and diagnostic criteria. PMID- 15280773 TI - Preferential in-utero transmission of HIV-1 subtype C as compared to HIV-1 subtype A or D. AB - OBJECTIVE: To determine whether different HIV-1 genotypes present in a single cohort, in Dar es Salaam, Tanzania, showed differences in timing for transmission from mothers to their infants. METHODS: We determined the maternal viral load, transmission time, and the HIV-1 envelope (env) subtype of 253 HIV-1-infected infants enrolled in a randomized double-blind placebo-controlled trial to examine the efficacy of vitamins in decreasing mother-to-child transmission in Tanzania. Classification of HIV-1 positivity in utero was based on PCR results at birth. Infants were classified as intrapartum infected if they scored negative for the sample collected at birth and positive for the sample collected at 6 weeks of age. RESULTS: We found significant differences in the distribution of transmission time according to subtype. A higher proportion of HIV-1 with subtype C env (C-env) was transmitted in utero than HIV-1 with subtype A env (A-env), subtype D env (D-env), or both combined. CONCLUSIONS: The identification of patterns of mother-to-child transmission times among HIV-1 genotypes may be useful in the selection of drug regimens for chemoprophylaxis. Based on our results, the efficacy of regimens administered only at labor may not protect as large a fraction of infants born in geographical regions with subtype C-env epidemics as compared to epidemics in regions where subtypes A-env and D-env predominate in the population. PMID- 15280774 TI - Topical estrogen protects against SIV vaginal transmission without evidence of systemic effect. AB - BACKGROUND: Accumulating data suggest that the state of the vaginal epithelium affects a woman's risk of HIV vaginal transmission and several human and non human primate studies have shown that the rate of HIV or SIV vaginal transmission is decreased when estrogen is dominant. Systemic estrogen can protect against SIV vaginal transmission. OBJECTIVE: To determine the safety and efficacy of topical estrogen in preventing SIV vaginal transmission. DESIGN: The non-human primate model of HIV vaginal transmission was used to assess vaginal estriol cream in ovariectomized macaques. METHODS: Twelve macaques were treated intravaginally with estriol and eight with placebo cream twice a week. The vaginal and systemic effects of estriol were determined by colposcopy and serum luteinizing hormone, levels of which would decline in the presence of systemic estrogen. After 5 weeks of therapy, the animals were challenged vaginally with pathogenic SIVmac251. RESULTS: Vaginal estriol resulted in minimal serum estriol levels and had no effect on serum luteinizing hormone levels. Vaginal epithelia cornified and thickened significantly in response to estriol therapy. One of the estriol treated animals became infected after this single challenge, while six of the control animals became infected (P = 0.0044). CONCLUSIONS: These data demonstrate that topical vaginal estriol can strongly protect against SIV vaginal transmission, while having no detectable systemic effect. These results support the study of topical vaginal estriol in preventing HIV vaginal transmission in at risk women. PMID- 15280775 TI - HIV-1 superinfections in a cohort of commercial sex workers in Burkina Faso as assessed by an autologous heteroduplex mobility procedure. AB - OBJECTIVE: To describe and evaluate a simple procedure to identify HIV-1 co- or super-infections based on a heteroduplex mobility assay (HMA). METHODS: To identify heteroduplexes corresponding to divergent viral populations in a the same individual, HMA was applied to single DNA samples from each subject in a prospective cohort of commercial sex workers (CSW) in Bobo-Dioulasso, Burkina Faso. After denaturation and renaturation of env DNA amplicons, hybridized DNA was separated by electrophoresis through polyacrylamide gel. HIV-1 co-infections were suspected by slow migration of heteroduplex, at a level comparable to that of mixed reference strains. Following electrophoresis, DNA in bands representing heteroduplex was extracted and cloned in a plasmid vector; identification of phylogenetically distinct clones was confirmed by sequencing divergent clones identified through a second HMA step of a pair of two mixed clones. RESULTS: Among 147 HIV-1 infected CSW, four had an autologous HMA profile comparable to low mobility of hybridized DNA from mixed reference strains representing most frequent HIV-1 clades and circulating recombinant forms (CRF) prevalent in Burkina Faso. In two of them, two phylogenetically distinct HIV-1 populations were coexisting. The first woman presented with a CRF02-AG and CRF06-cpx co infection, and the second with a CRF02-AG and a divergent virus co-infection not significantly related to any other known subtypes. In both women, retrospective analyses of stored samples by the same HMA procedure showed acquisition of a second virus concomitent with an increasing plasma HIV RNA. CONCLUSIONS: Autologous HMA procedure followed by acrylamide extraction of heteroduplexes allowed identifying HIV-1 co- and super-infections in a large cohort study. HIV-1 super-infection is not an uncommon phenomenon. PMID- 15280776 TI - Persistence of multidrug-resistant HIV-1 in primary infection leading to superinfection. AB - OBJECTIVE: The authors previous studies documented persistence of multidrug resistance (MDR) acquired in five primary HIV-1 infection (PHI) cases for 1-2 years in the absence of antiretroviral treatment. This study characterizes the evolution of transmitted wild-type (WT) (n = 15), resistant (n = 10), and MDR (n = 6) infections. Long-term persistence of MDR infections (2-7 years), leading to one observed MDR superinfection is documented. METHODS: Genotypic changes in circulating viral quasi-species were evaluated over 1.5-7 years in patients (n = 31) enrolled in the PHI study. Sequencing of reverse transcriptase and protease regions identified nucleotide substitutions in the viral quasi-species and mutations at sites implicated in resistance to antiretroviral drugs. Phylogenetic and clonal analysis were performed to confirm one observed superinfection. RESULTS: Patients acquiring WT, drug-resistant and MDR infections showed little quasi-species evolution (> 99.6% homology) for more than 1.5 years, regardless of route of transmission. Transmitted resistance mutations (other than 184V) persisted for 2-7 years. MDR persistence in two PHI cases contrasted with the corresponding rapid reversion of MDR infections to WT in their partners following treatment interruption. One MDR transmission eliciting low-level viremia resulted in clearance of the original MDR infection followed by re-infection with a second heterologous MDR strain from a different partner. Phylogenetic and clonal analysis of source and index partner confirmed the superinfection. Both MDR species showed approximately 13-fold reductions in replication capacity relative to the homologous WT strain isolated from the source partner. CONCLUSIONS: Genotypic analysis in PHI may identify superinfection and MDR infections that represent important determinants of virological and treatment outcome. PMID- 15280777 TI - Cost-effectiveness of nevirapine to prevent mother-to-child HIV transmission in eight African countries. AB - BACKGROUND: A comprehensive approach to preventing HIV infection in infants has been recommended, including: (a) preventing HIV in young women, (b) reducing unintended pregnancies among HIV-infected women, (c) preventing vertical transmission (PMTCT), and (d) providing care, treatment, and support to HIV infected women and their families. Most attention has been given to preventing vertical transmission based on analysis showing nevirapine to be inexpensive and cost-effective. METHODS: The following were determined using data from eight African countries: national program costs and impact on infant infections; reductions in adult HIV prevalence and unintended pregnancies among HIV-infected women that would have equivalent impact on infant HIV infections averted as the nevirapine intervention; and the cost threshold for drugs with greater efficacy than nevirapine yielding an equivalent cost per DALY saved. RESULTS: Average national annual program cost was 4.8 million dollars. There was, per country, an average of 1898 averted infant HIV infections (2517 US dollars per HIV infection and 84 US dollars per DALY averted). Lowering HIV prevalence among women by 1.25% or reducing unintended pregnancy among HIV-infected women by 16% yielded an equivalent reduction in infant cases. An antiretroviral drug with 70% efficacy could cost 152 US dollars and have the same cost per DALY averted as nevirapine at 47% efficacy. CONCLUSIONS: Cost-effectiveness of nevirapine prophylaxis is influenced by health system costs, low client uptake, and poor effectiveness of nevirapine. Small reductions in maternal HIV prevalence or unintended pregnancy by HIV-infected women have equivalent impacts on infant HIV incidence and should be part of an overall strategy to lessen numbers of infant infections. PMID- 15280778 TI - Immunological and virological study of enfuvirtide-treated HIV-positive patients. AB - OBJECTIVE: To evaluate the predictive value and evolution of immunological and virological parameters related to HIV entry and pathogenesis in patients receiving enfuvirtide (ENF) plus an optimized regimen. METHODS: A phase III clinical trial substudy of ENF in 22 patients measured virus coreceptor use and sensitivity to ENF, levels of chemokines, cytokines and chemokine receptors, CD38 and HLA-DR expression as markers of T cell activation and ex vivo cell death at baseline and at week 32. RESULTS: Treatment including ENF reduced HIV viral load (P < 0.001) and increased the CD4 cell count in patients that responded (RP) to treatment (n = 14). Significant (P < 0.05) increases were noted in the RP group in CXCR4 and CCR5 expression in CD4 cells without major differences in chemokine and interleukin-7 levels. A decrease in CD38 expression in the absence of HLA-DR changes was observed in CD4 cells. Apoptosis of peripheral blood mononuclear cells was significantly reduced in the RP group. Coreceptor use or ENF sensitivity of virus isolated at baseline was not associated with virus resistance or response to treatment, which appeared to be related to the activation state (HLA-DR expression) of CD4 cells at baseline. CONCLUSION: The outcome of ENF-containing treatment could not be associated with HIV coreceptor use at baseline. CD4 cell activation and viral drug resistance were the only markers of treatment response. Changes induced by ENF-containing regimen were seen in HIV coreceptor expression, including an increase in CCR5+CD4+ cells, a decrease in CD38 T cells and a concomitant reduction of T cell apoptosis. PMID- 15280779 TI - Persistence of primary drug resistance among recently HIV-1 infected adults. AB - OBJECTIVES: Primary, or transmitted, drug resistance is common among treatment naive patients recently infected with HIV-1, and impairs response to anti retroviral therapy. We previously observed that patients with secondary resistance (developed in response to anti-retroviral treatment) who chose to stop an anti-retroviral regimen experience rapid overgrowth of drug resistant viruses by wild-type virus of higher pol replication capacity. We sought to determine if primary drug resistance would be lost at a rapid rate, and viral pol replication capacity would increase, in the absence of treatment. METHODS: We tracked drug resistance phenotype, genotype, viral pol replication capacity (single cycle recombinant assay incorporating a segment of the patient pol gene [pol RC]), plasma HIV-1 RNA, and CD4 T cell counts in the absence of treatment among patients in early HIV-1 infection. RESULTS: Six of 22 patients had evidence of primary drug resistance to at least one class of drug; three resistant to protease inhibitors, three resistant to non-nucleoside reverse transcriptase inhibitors, and four resistant to nucleoside reverse transcriptase inhibitors. All six patients maintained evidence of drug resistance for the period of observation. Among patients with baseline primary drug resistance pol RC did not increase over time. CONCLUSION: The selection environment of early infection is determined by immune pressure, and stochastic events, not viral pol replication capacity. In contrast to secondary resistant infections that are rapidly overgrown when therapy is stopped, primary drug resistance persists over time. Surveillance and clinical detection of primary resistance is feasible in the first year of infection. PMID- 15280780 TI - Phenotypic impact of HIV reverse transcriptase M184I/V mutations in combination with single thymidine analog mutations on nucleoside reverse transcriptase inhibitor resistance. AB - OBJECTIVES: To analyse the impact of the M184I/V mutation and individual thymidine-associated mutations (TAM) on nucleoside reverse transcriptase inhibitor (NRTI) phenotypic susceptibility and compare these results with those obtained using commercial and public algorithms. DESIGN: An HIV genotypic/phenotypic database with over 27 000 samples was used to obtain the median fold change (5-95th percentile) in NRTI phenotypic susceptibility for viruses from patients containing individual TAM with or without the M184I or V mutation and for wild-type patient viruses. RESULTS: The resulting data indicated that in vitro, individual TAM do not have an equivalent impact on NRTI resistance, with some individual TAM having little or no impact on NRTI resistance (e.g. M41L or K219Q/E/H/R). In the presence of the M184I/V mutation, re-sensitization to some drugs, including zidovudine, stavudine and tenofovir was observed despite the presence of a TAM. For didanosine and abacavir, the presence of the M184V mutation and a single TAM did not result in a fold-change increase associated with decreased drug susceptibility. Analysis of public and commercial algorithms revealed a lack of concordance regarding the impact of these mutations, and with the observed phenotypic data. CONCLUSION: These analyses should assist in the creation of rules for genotypic drug resistance algorithms for a better reflection of the impact of individual TAM and also the impact of M184I/V on resistance. These data provide additional evidence that retaining lamivudine in those treatment regimens in which TAM can be selected may provide some therapeutic benefit by maintaining the M184V mutation. PMID- 15280781 TI - Differences in HIV RNA levels before the initiation of antiretroviral therapy among 1864 individuals with known HIV-1 seroconversion dates. AB - OBJECTIVE: To assess the effects of sex, risk group, age at and year of seroconversion (SC), and presentation during acute infection on HIV RNA trends before antiretroviral therapy (ART) initiation. METHODS: Multiple HIV RNA measurements from 1864 individuals with reliably estimated dates of SC, aged >/= 15 years at SC were studied using random effects models. Models were adjusted for selective HIV RNA data truncation due to ART initiation or AIDS development and for HIV RNA quantification assay. RESULTS: HIV RNA levels declined precipitously during the first 10 months after SC followed by a slow increase. Women infected heterosexually and through injecting drug use, had an average 34% [95% confidence interval (CI), 2.3-56%] and 46% (95% CI, 17-66%) lower HIV RNA load respectively, compared to men in the same risk group. Among men, those infected heterosexually and by injecting drug use had on average 56% (95% CI, 36-69%) lower HIV RNA levels than homosexual men. Older subjects tended to have higher viral levels. There was no evidence that differences by sex, risk or age group diminished over time, but follow-up was mostly before CD4 cell count had fallen below 200 x 10 cells/l. CONCLUSIONS: HIV RNA levels at the same stage of HIV-1 infection differ significantly by sex, risk group and age at SC. Given the lack of evidence of a survival difference by sex or risk group prior to initiation of effective therapy, further research on differential effects of virus load on treatment-free disease progression is needed, before a conclusion about considering these factors for ART initiation is drawn. PMID- 15280782 TI - Is unsafe sexual behaviour increasing among HIV-infected individuals? AB - BACKGROUND: The number of new diagnoses of HIV infection is rising in the northwestern hemisphere and it is becoming increasingly important to understand the mechanisms behind this trend. OBJECTIVE: To evaluate whether reported unsafe sexual behaviour among HIV- infected individuals is changing over time. DESIGN: Participants in the Swiss HIV Cohort Study were asked about their sexual practices every 6 months for 3 years during regular follow-up of the cohort beginning on 1 April 2000. METHODS: : Logistic regression models were fit using generalized estimating equations assuming a constant correlation between responses from the same individual. RESULTS: At least one sexual behaviour questionnaire was obtained for 6545 HIV-infected individuals and the median number of questionnaires completed per individual was five. There was no evidence of an increase in reported unsafe sex over time in this population [odds ratio (OR), 1.0; 95% confidence interval (CI), 0.96-1.05]. Females (OR, 1.38; 95% CI, 1.19-1.60), 15-30 year olds (OR, 1.26; 95% CI, 1.09-1.47), those with HIV positive partners (OR, 12.58; 95% CI, 10.84-14.07) and those with occasional partners (OR, 3.25; 95% CI, 2.87-3.67) were more likely to report unsafe sex. There was no evidence of a response bias over time, but individuals were less willing to leave questions about their sexual behaviour unanswered or ambiguous (OR, 0.93; 95% CI, 0.90-0.97). CONCLUSIONS: There was no evidence of a trend in unsafe sex behaviour over time. However, several subgroups were identified as being more likely to report unsafe sex and should be targeted for specific interventions. PMID- 15280783 TI - Factors affecting HIV concordancy in married couples in four African cities. AB - OBJECTIVE: To determine risk factors for HIV transmission within married couples in four urban populations in sub-Saharan Africa. METHODS: Data from a cross sectional population-based study were used. Representative random samples approximating 1000 men and 1000 women in each of four cities of Kisumu (Kenya), Ndola (Zambia), Cotonou (Benin), and Yaounde (Cameroon), were interviewed and tested for sexually transmitted infections (STI). Married couples were identified as concordant negative, discordant, or concordant positive for each STI. After excluding concordant HIV negative couples, analysis of behavioural and STI risk factors for HIV positive concordancy was undertaken across the four cities and in each city separately where sample size allowed. RESULTS: Among 221 couples in which at least one member was HIV positive, we found that the only significant risk factor for positive HIV concordancy was herpes simplex type 2 (HSV-2) status. After adjusting for age and city of residence the odds ratio for HIV concordancy compared to couples with neither spouse HSV-2 positive was 3.4 (95% confidence interval, 0.62-18.4) for couples with one partner HSV-2 positive and 8.6 (95% confidence interval, 1.6-45.0) for couples with both partners HSV-2 positive. The same trends were seen in Kisumu and Ndola when they were analysed separately (numbers were small in the other cities). CONCLUSIONS: Although cross sectional studies are not ideal for delineating the sequence of transmission events, this study adds to the evidence that HSV-2 is a key risk factor in promoting HIV transmission. PMID- 15280784 TI - Traditional health practitioners are key to scaling up comprehensive care for HIV/AIDS in sub-Saharan Africa. PMID- 15280785 TI - Nevirapine-containing regimens in HIV-infected naive patients with CD4 cell counts of 200 cells/microl or less. AB - We retrospectively evaluated the effectiveness of nevirapine-containing regimens in 118 naive patients initiating highly active antiretroviral therapy with CD4 cell counts S 200 cells/jl. After 24 months, 51% of patients continued nevirapine, 43 and 83% had viral loads < 50 copies/ml by intent-to-treat and on treatment analyses, and a mean increase of +246 CD4 cells/microl occurred. More than 80% of patients who continued with nevirapine had viral loads < 50 copies/ml and CD4 cell counts > 200 cells/pl. PMID- 15280786 TI - A novel CCR5 mutation selectively affects immunoreactivity and fusogenic property of the HIV co-receptor. AB - A Nepalese heterozygous carrier of a CCR5 mutant, designated 118delF, was characterized. There was a 3 basepair deletion at 352-354 in the CCR5 open reading frame, resulting in the deletion of the phe-118 residue located in the third transmembrane domain. The mutant protein has retained antigen specificity near the third extra-cellular loop (ECL3), but that of ECL2 is markedly reduced. The mutation has also abrogated HIV co-receptor activity. Clinically, the HIV disease had progressed slowly. PMID- 15280787 TI - An update on the impact of HIV/AIDS on life expectancy in the United States. AB - We used the potential gains in life expectancy to quantify the impact of eliminating HIV/AIDS,heart disease and malignant neoplasms on the life expectancy of the population of the USA from 1987 to 1999 by race and sex groups. We previously reported the results from 1987 to 1992,with a focus on the year 1992. This report gives an update to 1999, showing the impact of improvements in the care and treatment of HIV/AIDS in recent years. PMID- 15280788 TI - Severe hepatitis and prolonged hepatitis B virus-specific CD8 T-cell response after selection of hepatitis B virus YMDD variant in an HIV/hepatitis B virus-co infected patient. AB - We describe here a severe flare of hepatitis caused by lamivudine-resistant hepatitis B virus(HBV) in an HIV/HBV co-infected individual.Lamivudine-resistant HBV was detected 6 months before the development of severe hepatitis. Sequencing of the HBV genome isolated from the patients' serum did not identify compensatory mutations in the HBV polymerase that may have restored viral replication. However, a strong HBV-specific CD8 T-cell response was identified and may have resulted in the severe hepatitis. PMID- 15280789 TI - Pegylated liposomal doxorubicin plus highly active antiretroviral therapy versus highly active antiretroviral therapy alone in HIV patients with Kaposi's sarcoma. AB - Twenty-eight HIV patients either naive or failing highly active antiretroviral therapy (HAART)with moderate-advanced Kaposi's sarcoma (KS)were randomly chosen to initiate a new HAART regimen plus pegylated liposomal doxorubicin(PLD) or the new HAART regimen alone. After 48 weeks, better response rates were observed in the HAART plus PLD group (76% versus 20%). In HIV-infected patients with moderate advanced KS, HAART alone may not be enough for KS response. PMID- 15280790 TI - Proximal tubular kidney damage and tenofovir: a role for mitochondrial toxicity? PMID- 15280791 TI - Development of multiple lipomas during treatment with rosiglitazone in a patient with HIV-associated lipoatrophy. PMID- 15280792 TI - Response to Soriano et al., 'Care of patients with hepatitis C and HIV co infection'. PMID- 15280793 TI - Prevalence of non-significant liver fibrosis and rate of fibrosis progression in HIV/hepatitis C virus- co-infected patients: still a role for liver biopsy? PMID- 15280794 TI - Failure of treatment of tuberculous adenitis due to an unexpected drug interaction with rifabutin and efavirenz. PMID- 15280795 TI - [Oxidized dietary lipids may participate in the development of atherosclerosis]. AB - Atherosclerosis is highly correlated with postprandial lipemia and is characterized by elevated triglyceride levels in blood plasma. In normolipidemic individuals, the postprandial period lasts from 4-6 hours. The length of this period depends on the balance between the absorption of fat from the intestine and the removal of chylomicron remnants by the liver. An increasing amount of data suggests that oxidized dietary lipids may participate actively in the development of atherosclerosis. This working hypothesis is supported by numerous data indicating that oxidized lipids are absorbed from the diet in the same way as intact lipids and that they both participate in the synthesis of chylomicrons. Elevated plasma triglyceride levels may lead to rise in plasma low-density lipoproteins (LDL), since chylomicron remnants lower the expression of LDL receptors in the liver. Also the exchange of fatty acids and cholesterol between chylomicrons and LDL as well as high-density lipoproteins (HDL) increases during prolonged lipemia. Increasing interest in preliminary processed and deep-fried food indicates that more attention should be focused on postprandial lipoproteins as a major factor in the development of atherosclerosis, especially as humans are exposed to elevated triglyceride levels for most of their lives PMID- 15280796 TI - [Oxidative stress in hypertension]. AB - Reactive oxygen species (ROS) are involved in the pathogenesis of many cardiovascular diseases such as hypertension. In the circulation, ROS are generated by all vascular cells, i.e. endothelial cells, smooth muscle cells, and fibroblasts. Among the many enzymatic systems that are capable of producing ROS, NAD(P)H oxidase xantine oxidase and uncoupled endothelial nitric oxide synthase have been extensively studied in vascular cells. Enhanced ROS production (especially superoxide anion) causes diminished NO bioavailability and leads to endothelial dysfunction, which occurs for example in impaired vasorelaxation. Superoxide reacts with NO to form peroxynitrite, which can modify proteins and lipids to create nitrotyrosine, and nitrosothiols, isoprostanes, which are also able to modulate vascular tone. Several experimental observations have shown that a free radical scavenger may improve impaired endothelium-dependent vasodilatation and reduce elevated blood pressure in hypertension. PMID- 15280798 TI - [Hydrogen sulfide as a biologically active mediator in the cardiovascular system]. AB - Recent studies suggest that apart from nitric oxide (NO) and carbon monoxide (CO), hydrogen sulfide (H2S) is another inorganic gaseous mediator in the cardiovascular system. H2S is synthesized from L-cysteine by either cystathionine beta-synthase (CBS) or cystathionin gamma--lyase (CSE), both using pyridoxal 5' phosphate (vitamin B6) as a cofactor. CBS is the main H2S-producing enzyme in the brain and CSE is involved in H2S formation in the cardiovascular system. H2S induces hypotension in vivo and vasodilation vitro by opening KATP channels in vascular smooth muscle cells. Chronic administration of CSE inhibitor induces arterial hypertension in the rat. In addition, decreased H2S generation has been demonstrated in the vasculature of spontaneously hypertensive rat, in experimental hypertension induced by NO synthase blockade, and in hypoxia-induced pulmonary hypertension, and administration of exogenous H2S donor has significant therapeutic effects in these models. Deficiency of H2S may contribute to atherogenesis in some patients with hyperhomocysteinemia, in whom the metabolism of homocysteine to cysteine and H2S is compromised by vitamin B6 deficiency. Reduced H2S production in the brain was observed in patients with Alzheimer's disease. On the other hand, excess of H2S may lead to mental retardation in patients with Down's syndrome and may be involved in the pathogenesis of hypotension associated with septic shock. PMID- 15280797 TI - [The role of cytokine gene polymorphisms in organ and haematopoietic stem cell transplantation]. AB - The development of molecular biological techniques during the last decade has led to the recognition of a series of polymorphic sites in the regulatory regions of cytokine-encoding genes. Different alleles are associated with the binding of transcriptional factors and various degrees of cytokine production. Therefore each person has an individual profile of high and low cytokine responses. Some individuals are more susceptible to inflammatory conditions and the development of an immune response after transplantation. It has been documented that in heart transplantation high TNF-alpha/low IL-10 producers had high levels of graft rejection, while in renal transplants high TNF- alpha /high IL-10 producers were characterized with worse prognosis. The polymorphic features of genes encoding cytokines also associate with the outcome of bone marrow transplantation. It was shown that recipient TNFd, IFN- gamma (CA) and IL-10(-1064) microsatellite polymorphisms and IL-10 (-1082) and IL-6 (-174) SNP polymorphisms are associated with acute GvHD manifestation. No relation was found between TNFA (-308) and aGvHD. Recently, the influence of donor polymorphism within the IL-10 and IL-6 genes was documented. Polymorphism of the TNFd, TNFA (-308), TNFA (-238), TNFB ( 252), and IFN- gamma (+874) genes in donors were not related to this complication. In addition, donor and recipient IL-6 gene polymorphism and recipient IFN- gamma and IL-10 alleles were described as risk factors of cGvHD. PMID- 15280799 TI - [Tumor markers of breast cancer]. AB - The most frequent cancer amongst women is that of the breast. Tumor markers may be helpful in the early diagnosis of breast cancer and the initial assessment of the extent of disease, as well as in monitoring tumor growth or volume reduction, and a recurrence of cancer. They have also been used for monitoring the clinical course of chemotherapy and radiotherapy. In this paper we focus on the role of tumor markers such as CA 15-3, CEA, and TPS in breast cancer diagnostics, including cytokines and molecular markers of carcinogenesis. We also show the prognostic significance of markers tested in breast cancer biopsies. PMID- 15280800 TI - [Extracellular superoxide dismutase (EC-SOD)--structure, properties and functions]. AB - EC-SOD catalyzes the dismutation of superoxide radical to hydrogen peroxide and oxygen in the interstitial spaces of tissues and in extracellular fluids (plasma, lymph, and synovial fluid). It eliminates superoxide radicals from the cell environment and prevents the formation of reactive oxygen species and their derivatives. EC-SOD is a secretory, tetrameric glycoprotein containing copper and zinc, with a high affinity to certain glycosaminoglycans, such as heparin and heparan sulfate. It plays an important role in maintaining vascular tone, lung function, and the metabolism of NO, and in the pathology of such diseases as atherosclerosis, diabetes, and arthritis. This paper describes EC-SOD structure, function in tissues, and possibilities of therapy with application of this enzyme. PMID- 15280801 TI - Overview on the mechanisms of drug-induced liver cell death. AB - Drug-induced hepatotoxicity is a significant and still unresolved clinical problem. The limitation in current knowledge regarding mechanisms of hepatic toxicity renders most of the preclinical review process failing and most of drug induced hepatic injury remains unpredictable. Current knowledge on the mechanisms of drug-induced liver cell death is reviewed here. The intervention of both intra and extracellular factors in determining the appearance of drug-induced cell apoptosis or necrosis is also discussed. Finally, the role of both mitochondria and non parenchymal cells are reviewed with respect to approaches useful to manage drug-induced liver injury. PMID- 15280802 TI - Genomic medicine in Mexico. Applications of gene therapy for cirrhosis reversion. AB - Genomic medicine represents a powerful armamentarium to tackle down most of chronic diseases which have not, so far, defeated. Thus, this new and powerful biotechnologic set of weapons enable us to make use of molecular diagnostic to detect silent diseases, otherwise undetectable by conventional analysis. Moreover, elucidation of the complete and final draft of the human genome code will allow, although not in this decade, the design of specific farmaco-genetic treatments for patients on basis of their individual genetic code. Regarding new medical treatments, gene therapy as emerged as a true hope for treatment of many chronic diseases. 636 FDA-.approved clinical protocols are currently undergoing and sooner than later we ll be witness of the results PMID- 15280803 TI - Current role of surgery for the treatment of portal hypertension. AB - Portal hypertension surgery has evolved widely in the last decades. Since the first surgical shunt was done in 1945 for the treatment of recurrent hemorrhage, many surgical options have been developed including selective shunts, low diameter shunts and extensive devascularization procedures. Many of them have been studied and compared showing their advantages and disadvantages, evolving also their role in the therapeutic armamentarium. Surgery is nowadays a second line treatment option (after b blockers and endoscopic therapy), and it's main indication is for patients whose main and only problem is history of bleeding, with good liver function (Child-Pugh A). For emergency situations it has a very limited role and for primary prophylaxis virtually has also no role. Patients with good liver function, electively operated with portal blood flow preserving procedures are the patients that benefit from surgical treatment. Patients with a bad liver function are better candidates for a liver transplant. PMID- 15280804 TI - Evaluation of a third generation anti-HCV assay in predicting viremia in patients with positive HCV antibodies. AB - The most practical screening test for hepatitis C virus antibodies are second and third- generation enzyme immunoassays. We evaluated the usefulness of the third generation microparticle enzyme immunoassay (MEIA) in predicting HCV viraemia in anti-HCV positive patients. Serum samples from 106 patients with positive anti HCV were obtained. To evaluate the diagnostic value of the MEIA test in predicting HCV viraemia, anti-HCV positive patients were categorized in two groups according to the presence or absence of serum HCV-RNA. Among the 106 patients, 26 had non detectable serum HCV-RNA and 80 had detectable HCV-RNA by PCR. The assay automatically calculates a result based on the ratio of sample rate to the cut-of rate for each sample and control (S/CO). When the means of S/CO values for patients with detectable and non detectable HCV-RNA were analyzed, a statistically significant difference was found, (79.3 SD 22.2 vs. 8.2 SD 6.4, respectively) (p 0.0001). We further analyzed the best cut-off value of the S/CO in differentiating viremic from non viremic patients. The S/CO value of 26 showed a sensitivity of 99% and a specificity off 96% in discriminating both categories of HCV infected patients. In conclusion, our data demonstrate that viremic HCV patients had higher S/CO values in the MEIA test in comparison with non viremic patients. Hence, this assay may be used to predict HCV viraemia in anti-HCV positive individuals. PMID- 15280805 TI - Effect of S-nitrosoglutathione (GSNO) added to the University of Wisconsin solution (UW): II) Functional response to cold preservation/reperfusion of rat liver. AB - Livers cold preserved in University of Wisconsin (UW) solution followed by reperfusion suffer ischemia/reperfusion injuries. Microcirculation is the primary target of damage, characterized by sinusoidal perfusion failure due, mainly, to morphological changes of sinusoidal endothelial cells. Here, we demonstrated that the addition of S-nitrosoglutathione (GSNO) to the UW solution before cold storage, as a nitric oxide (NO) donor, attenuated hepatic injuries.Wistar adult rat livers were stored in UW solution (0 degrees C-48 hs) and then reperfused during 60 minutes using the Isolated Perfused Rat Model (IPRL). We assayed four GSNO concentration (50, 100, 250 and 500 mM). NO concentration was estimated calculating the amount of nitrite (NO2-) generated in the UW solution. Injuries during cold preservation were established measuring lactate dehydrogenase (LDH) released to the UW solution. Meanwhile, intrahepatic resistance (IR), LDH released to the perfusate, the effluent/perfusate ratio for K+, bile flow, liver glycogen content and sinusoidal endothelial cell morphology were studied after 1 hour of reperfusion in the IPRL system. In cold preserved livers without GSNO, glycogen content was dramatically reduced, IR increased markedly, LDH released was high, bile flow diminished and sinusoidal endothelial cells appeared rounded and detached from perisinusoidal matrix after reperfusion. The presence of 100 mM GSNO prevented the IR rise and LDH release, improved bile production and partially reduced endothelial cells damages. In conclusion, the addition of 100 mM GSNO to UW solution improved hemodynamic and function capacity of cold preserved/reperfused livers. PMID- 15280806 TI - Extrahepatic manifestations of viral hepatitis. AB - Viral hepatitis has been shown to be associated with various extrahepatic manifestations. These can be seen in both acute and chronic liver disease, may precede or follow overt liver disease. AIMS AND OBJECTS: To study the prevalence of extrahepatic manifestations of viral hepatitis and follow the course of the disease in response to antiviral therapy whenever indicated. METHODS: Prospectively 448 patients of viral hepatitis were evaluated for extrahepatic manifestations and patients of glomerulonephritis (GN), polyarteritis nodosa (PAN) and cryoglobulinemia were tested for viral markers. All patients were investigated for liver and kidney function tests, hematological workup and viral markers such as HBsAg, HBeAg, Anti HBeAg, HCV RNA, IgM anti HAV and IgM anti HEV. Serum electrophoresis and kidney biopsies were done whenever indicated. In 10 cases of hepatitis B glomerulonephritis immunohistochemistry was done on kidney biopsies for demonstration of hepatitis B surface and core antigen. RESULTS: Of total 448 cases 181 (40.4%) had hepatitis B infection, 142 (31.6%) had hepatitis C infection, 86 (19.1%) hepatitis E and 39 (8.7%) had hepatitis A infection. Extrahepatic manifestations were seen in 29 (6.4%) cases and these were cases of GN, PAN, cryoglobulinemia, thrombocytopenia, agranulocytosis, aplastic anemia and pancreatitis. Patients with hepatitis A with extrahepatic manifestations showed complete recovery in both hepatitis and extrahepatic manifestations. Six patients with PAN were treated with interferon of which 4 showed excellent response. Three patients of hepatitis B and hepatitis C related GN were given interferon and 4/6 responded well to treatment. CONCLUSION: Prevalence of extrahepatic manifestations with viral hepatitis was found to be 6.4%. These manifestations recover completely with recovery from viral hepatitis. PMID- 15280807 TI - Caroli's disease and choledochal cyst. PMID- 15280808 TI - Fibrolamellar carcinoma in a young patient. AB - This is a twenty two years old male patient with weight loss, abdominal pain, hepatomegaly and elevated liver function tests. The serological markers for viral B, C hepatitis and tumoral markers were normal. The CT scan demonstrated a hipodense, nodular lesion in the liver and the histological examination was reported as a typical fibrolamellar hepatocarcinoma PMID- 15280809 TI - Drug treatment for portal hypertension. AB - Pharmacological treatment of portal hypertension has played an increasing clinical role in the past 20 years. In the setting of acute variceal bleeding, drug therapy should be considered the initial treatment of choice and can be administered as soon as possible; even during the transfer of the patient to hospital. Several recent trials have reported similar efficacy to emergency sclerotherapy, therefore drug treatment should no longer be considered as a "stop gap" therapy until definitive endoscopic therapy is performed but continued for several days. Antibiotic prophylaxis is an integral part of therapy as it reduces mortality and should be instituted from admission. Non selective b-blockers are the treatment of first choice for secondary and primary prevention. If they are contraindicated or non tolerated banding ligation can be used. There is less evidence for the benefit of ligation for primary prophylaxis. The use of haemodynamic targets for reduction in hepatic venous pressure gradient response need further study, and surrogate markers of pressure response need evaluation PMID- 15280810 TI - From lipid secretion to cholesterol crystallization in bile. Relevance in cholesterol gallstone disease. AB - Failure of cholesterol homeostasis in the body can lead to cholesterol gallstone disease, the most common and costly gastrointestinal disease. The primum movens in cholesterol gallstone formation is the hypersecretion of hepatic cholesterol; this condition leads to bile chronically supersaturated with cholesterol which is prone to rapid precipitation as cholesterol crystals in the gallbladder. Essential topics reviewed here deal with pathways of biliary lipid secretion, cholesterol solubilization and crystallization in bile, according to recent advances. Main in vivo events in cholesterol gallstone disease are also described. PMID- 15280811 TI - Hepatitis C and liver transplantation. AB - Hepatitis C virus-related end-stage liver disease, alone or in combination with alcohol, has become the leading indication for liver transplantation in most transplant programs accounting for approximately half of transplants performed in European centers. The aim of this review is to analyze the factors involved in the results in different groups of patients with HCV underwent to liver transplantation. The groups involved those pretransplantation, post-transplant HCV infection, preventive early post-transplantation and with of recurrent hepatitis C. PMID- 15280812 TI - Hepatic adenomatosis in an Hispanic patient. A case report and review of the literature. AB - Liver adenomatosis (LA) is a rare disease originally defined by Flejou et al. in 1985 from a series of 13 cases. Only 57 cases have been reported in the literature, and all have been documented among Caucasian population. The aim of this study is to review and reappraise the characteristics of this rare liver disease, and to discuss the diagnosis and therapeutic options.LA is defined as the presence of >10 adenomas in an otherwise normal liver parenchyma. Neither female predominance nor a relation with estrogen/progesterone intake has been noted. Natural progression is poorly understood. We describe the clinical presentation, evolution, radiologic studies, histologic characteristics and therapeutic options in a 3rd generation Mexican woman with LA. We also include an updated review of the literature. The natural history and pathogenesis of LA are unclear. The risk of spontaneous hemorrhage or malignant transformation are a major concern. There is controversy regarding the optimal treatment for this disease; treatment options range from conservative medical therapy to surgical resection and even liver transplantation. LA is a rare disease, more common in women, and its outcome and evolution vary. Most often, conservative surgery is indicated. Liver transplantation is indicated only in highly symptomatic and aggressive forms of the disease. PMID- 15280813 TI - The effect of rewarming media composition on the ammonia detoxification ability of cold preserved rat hepatocytes. AB - We examined how different media composition of rewarming solutions affected ammonium detoxification function, urea synthesis and the viability of hepatocytes after 72 hs of cold storage in UW solution. Freshly isolated rat hepatocytes were incubated at 37 C in a cell culture medium (MEM-E) with 3 mM glycine, 5 mM fructose and 2.5 mM adenosine (group 1) and in Krebs-Heinseleit buffer with 3 mM glycine, 5 mM fructose, 2 mM ornithine, 10 mM lactate and adenosine, that was used in two different concentrations: 2.5 mM (group 2) and 10 mM (group 3). We found that freshly isolated cells produced ammonium in group 1 and 2 but the cells were able to diminish ammonium extracellular concentration in group 3. Urea synthesis and ammonium extracellular concentration in group 1 was higher than in group 2. As a result of this observations, we used the Krebs-Heinseleit solution with addition of 10 mM adenosine to determinate the effect of hypothermic preservation on ammonium detoxification and urea synthesis ability of cells. In conclusion the addition of 2.5 mM adenosine into the rewarming medium interfered with the detection of ammonium detoxification of hepatic cells. PMID- 15280814 TI - Hepatic focal fatty change in a chronic hepatitis C patient. PMID- 15280815 TI - National Consensus of Hepatitis C. June 28-29, 2002. PMID- 15280816 TI - The diagnosis and management of common anorectal disorders. PMID- 15280818 TI - Anterior encephalocele with subcutaneous right facial nodule. AB - Encephaloceles consist of heterotopic brain tissue that remains connected to the central nervous system. As such, these lesions can occur anywhere along the midline of the head, neck, and back. The clinical findings associated with an encephalocele are often cutaneous, prompting consultation with a dermatologist. Although abnormalities of the skin overlying the spinal cord are readily recognized by our specialty as markers for dysraphism, head and neck lesions may present a diagnostic challenge. We describe a case of an anterior encephalocele to increase awareness of this disorder and to emphasize the clinical findings that will assist with diagnosis. Our case is of particular interest because of the parasagittal location of the facial nodules and minimal actual midline involvement. PMID- 15280817 TI - Gemcitabine-associated scleroderma-like changes of the lower extremities. AB - Gemcitabine is a nucleosid analog approved for use in the treatment of metastatic urothelial carcinoma of the bladder. We describe an unusual case of scleroderma like changes of the lower extremities after treatment by gemcitabine for metastatic carcinoma of the bladder. The patient developed initial inflammatory edema (3 kg) restricted to the lower extremities and subsequent scleroderma-like changes after 2 cycles of gemcitabine. Cutaneous biopsy specimen revealed diffuse sclerosis without involvment of the fascia or muscle. Discontinuation of gemcitabine resulted in dramatic removal of the edema, softening of the skin, and partial reversibility of the fibrotic process. This is the first case report of a scleroderma-like reaction associated with gemcitabine. This antineoplastic agent must be added to the very limited number of cytostatic agents capable of giving rise to scleroderma-like features. PMID- 15280819 TI - Beta-lactam antibiotic-induced pseudoporphyria. AB - A case of beta-lactam antibiotic-induced pseudoporphyria is presented. A 24-year old African American woman with systemic lupus erythematosus and end-stage renal disease on hemodialysis developed tense bullae on her forehead and cheeks after exposure to ampicillin-sulbactam and cefepime. Histologically, the lesions were similar to porphyria cutanea tarda, but without the associated porphyrin abnormalities. The lesions resolved spontaneously on cessation of the antibiotics. PMID- 15280820 TI - Treatment of Behcet's disease with granulocyte and monocyte adsorption apheresis. AB - The painful orogenital ulcerations of Behcet's disease are among the major symptoms of patients and are often intractable. We assessed the efficacy of granulocyte and monocyte adsorption apheresis therapy in two patients, a 21-year old man with orogenital ulcerations and a 50-year-old woman with genital ulceration and abdominal pain. They underwent 5 and 8 granulocyte and monocyte adsorption apheresis treatments at 5-day intervals, respectively. The painful orogenital ulcerations of the man responded dramatically and the genital ulcer of the woman decreased in size and her abdominal pain was improved. Our results demonstrate that granulocyte and monocyte adsorption apheresis may be useful for treating orogenital ulcerations of Behcet's disease. PMID- 15280821 TI - Dermoscopic features of melanoma on the scalp. AB - There is a need to improve the early detection of melanoma of the scalp because it is characterized by a poorer prognosis compared with melanoma on other body sites. Dermoscopy is a useful tool for the early detection of melanoma but no previous reports on dermoscopic features of scalp melanoma have been published. We describe the first case of melanoma of the scalp seen by dermoscopy exhibiting a multicomponent global pattern with atypical pigment network, irregular streaks, and regression structures. In contrast to the dermoscopic features usually seen in melanoma occurring on the face, the same morphologic type of pigment network usually seen in melanoma of the trunk was observed in our case of scalp melanoma. PMID- 15280822 TI - Periungual lipoma: about three cases. AB - Lipomas are one of the most common benign soft tissue tumors. The usual development sites are the neck, the torso, and the legs. Lipomas of the nail unit are extremely rare. Only five cases have been reported up to now, four in subungual locations and one in the lateral nail fold. We report three cases of peri-ungual lipomas, one on the digit and two on the toes. Two of them exhibited the histological features of perisudoral lipomas. PMID- 15280823 TI - A bullous neutrophilic dermatosis in a patient with severe rheumatoid arthritis and monoclonal IgA gammopathy. AB - A 35-year-old man presented with a 2-month history of itchy grouped vesiculobullae with crusted papules and confluent plaques, distributed symmetrically on the thighs, knees, and elbows. The man had experienced severe disabling seropositive rheumatoid arthritis for 18 years previously. Histologically, subepidermal vesicles were noted, and a dense, diffuse neutrophilic infiltration of the dermis. Direct immunofluorescence studies did not detect IgA deposits. The patient responded well to dapsone (100 mg/d) but not to systemic steroids. This patient's eruption might have been an unusual manifestation of rheumatoid neutrophilic dermatitis. PMID- 15280824 TI - Scrotal calcinosis: is the cause still unknown? AB - Scrotal calcinosis is a rare benign entity of calcified nodules within the scrotal skin. We describe a healthy 25-year-old man with multiple asymptomatic calcified nodules restricted to the scrotum. Histologic studies of multiple nodules showed calcium deposition, and basophilic material with sparse inflammation surrounded by a fibrous capsule. Because the intensive evaluation of our patient failed to reveal a cause, an analysis of all 123 cases found in the literature was conducted. Several plausible origins for scrotal calcinosis are disclosed; however, pathogenesis of this condition remains equivocal and controversial. The most common treatment option is excision of the affected nodules. Our report underscores the need for further investigation of this florid and bizarre disorder. PMID- 15280825 TI - Independent lesions of fixed drug eruption caused by trimethoprim sulfamethoxazole and tenoxicam in the same patient: a rare case of polysensitivity. AB - Polysensitivity in fixed drug eruption is a rare finding that occurs because of chemically unrelated drugs. In cases of polysensitivity, the lesions may occur on identical or separate sites, the latter indicating the role of antigen-specific mechanisms in the site-specificity of fixed drug eruption. I herein report a patient with separate site involvement induced by trimethoprim-sulfamethoxazole and tenoxicam, a drug combination that has not been reported before. Reactivation of old trimethoprim-sulfamethoxazole-specific lesions after a long resting period was another striking feature. PMID- 15280826 TI - Interstitial granulomatous dermatitis secondary to soy. AB - A healthy 58-year-old woman developed an asymptomatic papular eruption of the neck, cheek, abdomen, arms, and flexures. There was an 8-year history of the lesions, which had erupted when the patient started a strict vegetarian diet. Lesions lasted 3 to 5 days, cleared without scarring, and were associated with burning and increased tearing of the eyes. The biopsy specimen showed an interstitial granulomatous dermatitis without vascular injury, collagen alteration, or mononuclear atypia. The eruption cleared when the patient omitted soy products from her diet. It subsequently recurred with intake of even minimal amounts of soy. Interstitial granulomatous dermatitis is a histologic pattern of inflammation that generates a broad differential diagnosis. No previous reports of interstitial granulomatous dermatitis related to soy products are available in the literature. PMID- 15280827 TI - Distant cutaneous metastases of cholangiocarcinoma: report of two cases of a previously unreported condition. AB - Cutaneous metastases develop in 2% to 10% of patients with internal malignancies. Here we present two cases of bile duct carcinoma with distant cutaneous metastases, a condition previously unreported. Scalp tumors were the initial presentation in both cases. PMID- 15280828 TI - Unusual clinical manifestation of linear IgA dermatosis: a report of two cases. AB - Linear IgA dermatosis is a rare autoimmune bullous skin disease with subepidermal blister formation and linear IgA deposits along the basement membrane zone. We describe two female patients showing erythematous annular plaques with scaling at the margin, strictly localized to the palms in one patient, and also found on the soles and buttocks in the second patient. Histology showed numerous neutrophils in the dermis with an admixture of eosinophils, some subepidermal clefting, and occasional papillary microabscesses. Direct immunofluorescence and immunoelectron microscopy revealed in vivo IgA deposition along the basement membrane zone. One patient cleared after treatment with dapsone. The second patient did not respond to dapsone alone and various immunosuppressive treatment regimens. Considerable improvement was achieved with intravenous immunoglobulin therapy combined with corticosteroid and dapsone. PMID- 15280829 TI - Idiopathic calcinosis cutis of the penis. AB - Calcification of the skin occurs in three main forms: dystrophic; metastatic; and idiopathic. Idiopathic calcinosis cutis of the penis is a rare event and only 4 cases have been reported. Herein we present another case in a 19-year-old man and discuss its probable pathogenic origin. PMID- 15280830 TI - A complete and durable clinical response to high-dose dexamethasone in a patient with scleromyxedema. AB - We report a case of a patient with scleromyxedema limited to the skin with an associated IgG lambda monoclonal protein treated successfully with high-dose dexamethasone. We encourage the continued investigation of this complex relationship between the clinical presentation of scleromyxedema and its frequently associated paraproteinemia. PMID- 15280831 TI - Self-resolution of Epstein-Barr virus-associated B-cell lymphoma in a patient with dermatomyositis following withdrawal of mycophenolate mofetil and methotrexate. AB - Self-resolving Epstein-Barr virus (EBV)-associated lymphomas have become more common with the use of immunosuppressive agents in both transplant patients and patients with connective tissue disorders. Immunosuppressive agents are often used for control of dermatomyositis, but their use has not been linked to subsequent malignancy. We present a 46-year-old woman with dermatomyositis, who developed an EBV-associated B-cell lymphoma of the brain while on oral methotrexate, mycophenolate mofetil and low-dose prednisone. The patient's lymphoma gradually resolved "spontaneously" upon discontinuation of the methotrexate and mycophenolate mofetil. The potential for EBV-associated B-cell lymphoma to self-resolve should be recognized by the clinician in order to prevent unnecessary and potentially toxic treatments including radiation therapy or multi-drug chemotherapy. PMID- 15280832 TI - Localized bullous pemphigoid overlying a fistula for hemodialysis. AB - Two cases of bullous pemphigoid exclusively limited to the skin overlying a hemodialysis fistula in patients with chronic renal failure are reported. This is an unusual association, but should be taken into account in the differential diagnosis of bullous disorders among patients on hemodialysis. We hypothesize about the pathogenesis of this process. PMID- 15280833 TI - Absence of Kaposi's sarcoma-associated herpesvirus in lesions of mycosis fungoides in patients with concomitant Kaposi's sarcoma. PMID- 15280834 TI - Papillon-Lefevre syndrome associated with pseudoainhum. PMID- 15280835 TI - Acquired disorders of elastic tissue: Part II. decreased elastic tissue. AB - Elastic fibers in the extracellular matrix are integral components of dermal connective tissue. The resilience and elasticity required for normal structure and function of the skin are attributable to the network of elastic tissue. Advances in our understanding of elastic tissue physiology provide a foundation for studying the pathogenesis of elastic tissue disorders. Many acquired disorders are nevertheless poorly understood owing to the paucity of reported cases. Several acquired disorders in which loss of dermal elastic tissue produces prominent clinical and histopathologic features have recently been described, including middermal elastolysis, papular elastorrhexis, and pseudoxanthoma-like papillary dermal elastolysis, which must be differentiated from more well-known disorders such as anetoderma, acquired cutis laxa, and acrokeratoelastoidosis. Learning objective At the conclusion of this learning activity, participants should have an understanding of the similarities and differences between acquired disorders of elastic tissue that are characterized by a loss of elastic tissue. PMID- 15280836 TI - The reliability of horizontally sectioned scalp biopsies in the diagnosis of chronic diffuse telogen hair loss in women. AB - BACKGROUND: Chronic diffuse telogen hair loss is common in women. Paired 4-mm punch biopsy from the vertex scalp for horizontal and vertical sectioning is commonly used to distinguish between chronic telogen effluvium (CTE) and female pattern hair loss (FPHL). FPHL is now the favored term for androgenetic alopecia in women. OBJECTIVE AND METHODS: To evaluate the reliability of a single horizontally sectioned scalp biopsy in the diagnosis of FPHL, 207 women presenting with chronic diffuse hair loss had three 4-mm punch biopsy specimens taken from immediately adjacent skin on the mid scalp, and all 3 biopsy specimens were sectioned horizontally. Findings were compared with 305 women who underwent two biopsies, with one sectioned horizontally and the other vertically. The terminal to vellus-like hair ratio (T:V) at the mid-isthmus level was used to diagnose FPHL (T:V <4:1), CTE (T:V >8:1), or indeterminate hair loss (T:V=5:1, 6:1, or 7:1). To correlate the histologic diagnosis with the clinical severity, a mid-scalp clinical grading scale was developed. RESULTS: Among the 305 women who had a single horizontal scalp biopsy, 181 (59%) were diagnosed as having FPHL, 54 (18%) having CTE, and 70 (23%) having indeterminate hair loss. Six hundred twenty one horizontal biopsy specimens were assessed from 207 patients. On the basis of consensus over 3 biopsies, 159 (77%) were diagnosed as having FPHL, 44 (21%) having CTE, and the remaining 4 women (2%) as having indeterminate hair loss. Among these 207 women, 114 were assessed clinically as having stage 1 or 2 hair loss. Sixty-nine (60%) were diagnosed as having FPHL on the basis of triple biopsy, 42 (37%) having CTE, and 2 having indeterminate hair loss. Ninety-three were graded as having stage 3, 4, or 5 hair loss. FPHL was diagnosed in 90 women (97%), CTE in 2, and indeterminate hair loss in one. By using each single biopsy as the criterion for diagnosis, 398 (61%) were classified as FPHL, 99 (16%) as CTE, and 124 (20%) as indeterminate. In 493 biopsies (79%), the single biopsy conclusion was identical to the 3 biopsy conclusions. Where disagreement was seen (21%), most were classified as indeterminate, rather than as a wrong diagnosis (3.3%). CONCLUSION: Application of these diagnostic criteria achieved accurate diagnostic definition in 98% of women with triple horizontal biopsies versus 79% with single horizontal biopsy. Ninety-seven percent of women with a mid-scalp clinical grade of 3, 4, or 5 were given a diagnosis of FPHL on triple biopsy. Scalp biopsy for diagnosis should be reserved for women with a mid-scalp clinical grade of 1 or 2. PMID- 15280837 TI - Pilot study of etanercept in patients with relapsed cutaneous T-cell lymphomas. AB - BACKGROUND: Tumor necrosis factor (TNF)-alpha has been implicated in the pathogenesis of cutaneous T-cell lymphoma (CTCL). OBJECTIVE: To assess the toxicity, safety, and efficacy of etanercept (soluble TNF receptor) in patients with relapsed CTCL. METHODS: Etanercept was administered twice weekly at a dose of 25 mg subcutaneously. Patients with improvement after two months could be continued on treatment. RESULTS: Twelve out of the 13 patients enrolled on study were evaluable (Stage I-IIA, 3 patients; Stage IIB-IV disease, 9 patients). The median number of previous therapies was 7 (range, 3-12). Etanercept induced partial remission in one patient (8%) and minor response in one patient (8%), both of whom had Stage IB disease. Most patients experienced no side effects. CONCLUSION: This pilot study suggests that etanercept is safe and generally well tolerated in patients with CTCL. The effect of etanercept in a larger cohort of patients with early disease merits investigation. PMID- 15280838 TI - Association of porphyria cutanea tarda with hereditary hemochromatosis. AB - BACKGROUND: An increased frequency of hereditary hemochromatosis gene mutations occurs in patients with porphyria cutanea tarda. Polymerase chain reaction analysis of peripheral blood for hemochromatosis gene (HFE) mutations is available for clinical use. Early detection and treatment of hereditary hemochromatosis limit disease progression and improve life expectancy. OBJECTIVE: We present 8 patients with porphyria cutanea tarda subsequently found to have hereditary hemochromatosis or mutations in the HFE gene. METHODS: Retrospective review of patients in whom both porphyria cutanea tarda and hereditary hemochromatosis or HFE gene mutations were diagnosed between 1976 and 2000. RESULTS: Eight patients with porphyria cutanea tarda (6 males, 2 females; age range, 4-60 years; mean age at diagnosis of porphyria cutanea tarda, 42 years) were subsequently found to have hepatic iron overload or HFE gene mutations. Two patients had liver biopsy findings compatible with homozygous hereditary hemochromatosis. In the other 6 patients, HFE gene analysis revealed 3 homozygous C282Y, 1 compound heterozygous C282Y/H63D, and 2 heterozygous C282Y mutations. Seven patients (88%) had no specific signs or symptoms of hereditary hemochromatosis at diagnosis. In 5 patients (63%), the diagnosis of hereditary hemochromatosis or HFE gene mutation was initially suspected by the dermatologist. CONCLUSION: Porphyria cutanea tarda can be an important cutaneous marker for patients with mutations of the HFE gene. HFE gene analysis should be done in patients who present with porphyria cutanea tarda. The dermatologist may play a key role in the early diagnosis of subclinical hereditary hemochromatosis in patients who present with porphyria cutanea tarda. PMID- 15280839 TI - Adherence to topical therapy decreases during the course of an 8-week psoriasis clinical trial: commonly used methods of measuring adherence to topical therapy overestimate actual use. AB - INTRODUCTION: Medication nonadherence is common throughout medicine, and research into this area is increasing; however, knowledge about topical medication adherence is limited. METHODS: A total of 30 patients were enrolled in a clinical trial for psoriasis and followed up for 8 weeks using 3 methods of adherence monitoring: electronic monitoring caps; medication logs; and medication usage by weight. RESULTS: Adherence rates calculated from the medication logs and medication weights were consistently higher than those of the electronic monitors (P <.05). Electronically measured adherence rates declined from 84.6% to 51% during the 8-week study (P <.0001). Female sex and increasing age by 1 year predicted improved adherence of 5% and 0.8%, respectively (P <.0001). The number of treatment gaps increased from the first half to the last half of the study, and weekend days were overrepresented in treatment gaps. CONCLUSION: Medication logs and weights do not ensure medication adherence to topical therapy. Electronic monitoring allows a more precise method of adherence measurement. PMID- 15280840 TI - The contribution of the arrector pili muscle and sebaceous glands to the follicular unit structure. AB - BACKGROUND: The evidence of hairs grouped into well-defined follicular units has given a new vision of hair anatomy and pathology. The sebaceous glands and the arrector pili muscle, as part of the pilosebaceous unit, should be viewed as important parts of this organized follicular unit structure. OBJECTIVE: This study was performed to define the morphology and the relationships between the sebaceous glands, muscles, and follicles within the context of the follicular unit structure. METHODS: This study analyzes horizontal, microscopic serial sections of large areas of normal human scalp skin stained with hematoxylin and eosin, Masson's trichrome, and desmin. The course of the arrector pili muscles from their superficial origins to their follicular attachments was followed in each section, which enabled us to match each muscle with its corresponding follicular unit. RESULTS: Serial, horizontal sections show that, at the upper isthmus, the arrector pili muscle is arranged as a muscular unit structure at the periphery of each follicular unit. Then, at a lower level, the muscle divides and encircles the sebaceous gland, and penetrates between the sebaceous lobules towards the follicular attachment zone. CONCLUSION: Microphotographic evaluation of large areas of scalp is important for assessing the anatomical relationships between the sebaceous glands, arrector pili muscles, and hair follicles as components of the follicular unit. We introduce the anatomical concept of a follicular unit served by a muscular unit, which can be identified in horizontal sections made at the upper level of the isthmus. This muscular unit results from the merging of the arrector pili muscles that originate from the hair follicles contained in that particular follicular unit. This anatomical disposition suggests that the arrector pili muscles could play an important role in the integrity of the follicular unit as well as in the secretion of the sebum contents. PMID- 15280841 TI - A multicenter, randomized, double-blind, placebo-controlled study of efficacy and safety of 3 doses of botulinum toxin A in the treatment of glabellar lines. AB - BACKGROUND: Botulinum toxin A (BTX-A) is used to treat glabellar lines but the rigorous demonstration of its efficacy in a well-designed study had never been reported. OBJECTIVE: This study was designed to evaluate the efficacy and the safety of 3 doses of BTX-A in the treatment of glabellar lines. METHODS: A total of 119 patients with moderate to severe glabellar lines at rest were treated with 25, 50, or 75 U of BTX-A (Dysport, Ipsen) or placebo divided into 5 intramuscular glabellar sites. Outcome measures included evaluations of glabellar lines by independent experts from blinded standardized photographs at rest 1 month after treatment, physician evaluations, and patient assessments during a 6-month period. RESULTS: A significant efficacy was reported for the 3 BTX-A groups for at least 3 months after injection (at least P <.015). Investigator and patient evaluations suggested that 50 U was the optimal dose. BTX-A was well tolerated. No blepharoptosis was reported. An evaluation in blinded conditions by independent experts was necessary because the results were overestimated by the investigators. CONCLUSION: BTX-A is an effective and safe treatment for glabellar lines. PMID- 15280842 TI - Statistical reviewing policies in dermatology journals: results of a questionnaire survey of editors. AB - BACKGROUND: Problems with statistical methods and reporting have been noted in articles published in dermatology journals. Conclusions presented in published reports may be misleading if based on inappropriate or misinterpreted statistical analysis. OBJECTIVE: We sought to assess dermatology journal editors' policies and perceptions regarding statistical review of submitted manuscripts. DESIGN: We mailed and e-mailed a questionnaire survey. PARTICIPANTS: A total of 43 dermatology journal editors, representing 35 dermatology journals from the United States and abroad, participated in this study. RESULTS: In all, 32 editors (74.4%), representing 30 journals (85.7%), returned questionnaires. A total of 24 editors (75%) reported having requested statistical reviews on less than 5% of published manuscripts containing original quantitative analysis (ie, excluding reviews and case reports), whereas 3 editors (9.4%) reported having requested statistical reviews on more than 75% of such manuscripts. Most editors reported requesting statistical reviews on a case-by-case basis either after initial favorable review by subject-matter (nonstatistical) reviewers (12 editors; 37.5%) or at the same time that subject-matter review was requested (6 editors; 18.8%). A total of 4 editors (12.5%) reported requesting statistical review for all manuscripts at the same time they are sent for subject-matter review. Another 10 editors (31.3%) said their journals had no general policy on statistical reviewing, and statistical review is almost never needed. For 15 editors (46.9%), ideal statistical reviewing policy was identical to their current policy, whereas 13 (40.6%) favored a more rigorous and 3 (9.4%) a less rigorous policy. CONCLUSIONS: Dermatology journals infrequently perform statistical reviews of submitted manuscripts. Dermatology journal editors' statistical review policies range from no general policy to (most frequently) requesting reviews on a case-by case basis to reviewing all submitted manuscripts. Many editors favor more rigorous statistical reviewing policies for their journals. Increased use of statistical reviewing may increase the reliability of conclusions published in dermatology journals. PMID- 15280843 TI - US prevalence of hyperhidrosis and impact on individuals with axillary hyperhidrosis: results from a national survey. AB - BACKGROUND: The current epidemiologic data on hyperhidrosis are scarce and insufficient to provide precise prevalence or impact estimates. OBJECTIVE: We sought to estimate the prevalence of hyperhidrosis in the US population and assess the impact of sweating on those affected by axillary hyperhidrosis. METHODS: A nationally representative sample of 150,000 households was screened by mailed survey for hyperhidrosis and projected to the US population based on US census data. Ascertainment of hyperhidrosis was based on a question that asked whether participants experienced excessive or abnormal/unusual sweating. RESULTS: The prevalence of hyperhidrosis in the survey sample was 2.9% (6800 individuals). The projected prevalence of hyperhidrosis in the United States is 2.8% (7.8 million individuals), and 50.8% of this population (4.0 million individuals) reported that they have axillary hyperhidrosis (1.4% of the US population). Only 38% had discussed their sweating with a health care professional. Approximately one third of individuals with axillary hyperhidrosis (0.5% of the US population or 1.3 million individuals) reported that their sweating is barely tolerable and frequently interferes, or is intolerable and always interferes, with daily activities. CONCLUSION: Hyperhidrosis affects a much larger proportion of the US population than previously reported. More than half of these individuals have axillary hyperhidrosis, in which sweating can result in occupational, emotional, psychological, social, and physical impairment. PMID- 15280844 TI - A phase II multicenter clinical trial of systemic bexarotene in psoriasis. AB - BACKGROUND: Bexarotene, a novel and unique synthetic P, RXR-selective retinoid, is available as a treatment for cutaneous T-cell lymphoma. In psoriasis, a common retinoid-sensitive disease, no data are available on bexarotene treatment. OBJECTIVE: In this phase II study we investigated the safety, tolerability, and effectiveness of bexarotene in psoriasis at doses of 0.5 to 3.0 mg/kg/day. METHODS: Fifty patients with moderate to severe plaque-type psoriasis were treated with bexarotene in 4 sequential dose-defined panels of 12-13 patients at doses of 1.0, 2.0, 0.5, and 3.0 mg/kg/day for 12-24 weeks. Patients were monitored for safety and clinical efficacy. RESULTS: No serious adverse events related to the drug occurred. Bexarotene was well tolerated in most patients. Most frequently observed adverse events related to bexarotene were hypertriglyceridaemia (56%) and a decrease in free T4 serum levels (54%). Significant improvement of psoriasis after bexarotene at all doses was confirmed by a modified psoriasis area and severity index (mPASI), plaque elevation (PEL), and physician's global assessment (PGA). Overall response rates (> or =50% improvement) for mPASI, PEL, and PGA were 22%, 52%, and 36%, respectively. No significant dose-response effect was established for these parameters. CONCLUSION: The present study indicates an anti-psoriatic effect of bexarotene. Further studies are necessary to assess the optimal dose and the potential for bexarotene as a new therapy for psoriasis. PMID- 15280845 TI - Systemic treatment of psoriatic patients with bexarotene decreases epidermal proliferation and parameters for inflammation, and improves differentiation in lesional skin. AB - BACKGROUND: Bexarotene, a novel synthetic retinoid X receptor (RXR)-selective retinoid, has been reported to have antiproliferative and apoptotic stimulating effects in cutaneous T-cell lymphoma. In benign, hyperproliferative, and retinoid sensitive disorders, such as psoriasis, bexarotene has not been evaluated so far and no information on these parameters is available. OBJECTIVE: In the present study, immunohistochemical parameters for proliferation, differentiation, inflammation, and apoptosis were investigated in a group of bexarotene-treated psoriatic patients. METHODS: Twenty-nine patients with plaque-type psoriasis were treated for 12 weeks with oral bexarotene in four dose-defined treatment panels. Treatment was initiated in the following consecutive order: 1.0 mg/kg/day, 2.0 mg/kg/day, 0.5 mg/kg/day, and 3.0 mg/kg/day. Biopsies for immunohistochemical analysis were taken at the baseline and after 12 weeks of treatment. RESULTS: Significant reductions in Ki-67, keratin 16, transglutaminase, dermal CD4, epidermal CD8, and inflammation scores were seen after bexarotene treatment in combination with a significant increase in keratin 10. No induction of keratin 13 and 19 and no alterations in apoptosis associated p53 expression were observed. Apart from a weak significant dose-response effect for Ki-67, no other significant dose-response effects were seen. CONCLUSION: We have demonstrated efficacy of oral bexarotene in psoriasis in doses up to 3.0 mg/kg/day during 12 weeks of treatment for proliferation, differentiation, and inflammation parameters. Studies investigating higher doses of bexarotene in a larger number of patients are necessary to reveal potentially dose-related immunohistochemical effects of this new rexinoid and to elucidate the role of RXR-signaling in retinoid-associated keratin expression. PMID- 15280846 TI - Effectiveness of small-volume, intralesional, delayed-release triamcinolone injections in orofacial granulomatosis: a pilot study. AB - BACKGROUND: Orofacial granulomatosis (OFG) is an idiopathic disorder characterized by chronic orofacial swellings causing significant cosmetic and functional problems. Treatment with high-volume triamcinolone injections has been shown to be effective but requires nerve block anesthesia and causes a dramatic temporary increase of lip swelling. OBJECTIVE: We have performed a noncomparative open-label pilot study in 7 patients with OFG in order to evaluate the effectiveness of small volumes of extended-release high-concentrate triamcinolone injections in reducing lip swelling and preventing recurrences. METHODS: Seven patients with OFG were studied. Small-volume, intralesional, high-concentrate, extended-release triamcinolone was injected on the basis of a weekly schedule. A standard cycle consisted of 2 or 3 injection sessions over 14 or 21 days, depending on the clinical response. RESULTS: After cycle completion, all patients remained without recurrences or with cosmetically acceptable slight lip enlargement for a mean time of 19 months (range, 8-30 months). No side effects were observed, except in one patient with hypopigmentation of the skin of the upper lip. CONCLUSIONS: Slow-volume, intralesional, high-concentrate, extended release triamcinolone injections appear to be effective in reducing lip enlargement in patients with OFG and do not require nerve block anesthesia or cause a temporary troublesome increase of swelling. A long disease-free period is generally obtained. PMID- 15280847 TI - Granuloma faciale: a clinicopathological study of 11 cases. AB - BACKGROUND: Granuloma faciale is an uncommon disease of unknown etiology that is often misdiagnosed clinically and by general pathologists. OBJECTIVE: To describe the clinicopathological features of a series of patients with granuloma faciale. METHODS: Eleven patients diagnosed with granuloma faciale between 1990 and 2002 were included in the study. RESULTS: Granuloma faciale was diagnosed in 11 patients (9 male and 2 female, mean age 53.45 years). All of them presented facial cutaneous lesions and two of them also developed extrafacial lesions. Histologically, in 8 cases the infiltrate was limited to the upper half of the dermis. Two specimens showed fibrinoid necrosis. Concentric fibrosis around small blood vessels was demonstrated in 5 patients. CONCLUSION: The presence of abundant fibrosis in 5 of our patients similar to that observed in erythema elevatum diutinum suggests that granuloma faciale and erythema elevatum diutinum may be produced by similar or the same pathogenic mechanism. PMID- 15280848 TI - Recognition, diagnosis, and treatment of primary focal hyperhidrosis. PMID- 15280849 TI - Neonatal dermatology. PMID- 15280850 TI - Medical pearl: permethrin can prevent arthropod bites and stings. PMID- 15280852 TI - Hydroxyurea-associated squamous dysplasia. AB - We report 2 cases of squamous dysplasia associated with long-term hydroxyurea therapy. The association between hydroxyurea and multiple aggressive squamous cell carcinomas, Bowen's disease, and multiple actinic keratoses in sun-exposed areas after a variable latency period has been increasingly reported. On reviewing the literature, 17 cases were identified and are reviewed, with emphasis on possible pathogenetic mechanisms of carcinogenicity. Squamous dysplasia, a precursor state to a more aggressive condition of multiple squamous cell carcinomas in photoexposed areas, should be added to the well-known cutaneous toxicities of hydroxyurea therapy. PMID- 15280851 TI - Surgical pearl: tumescent anesthesia reduces pain of axillary laser hair removal. PMID- 15280853 TI - Eccrine nevus. AB - Localized hyperhidrosis on the left forearm of a 7-year-old girl is described. Biopsy revealed an eccrine nevus. The differential diagnosis of localized hyperhidrosis is discussed. PMID- 15280854 TI - Progressive epidermotropic CD8+/CD4- primary cutaneous CD30+ lymphoproliferative disorder in a patient with sarcoidosis. AB - We describe a patient with a CD8+/CD4- primary cutaneous CD30(+) lymphoproliferative disorder with striking epidermotropic histology and coincident cutaneous and systemic sarcoidosis. This patient illustrates the spectrum of clinical and histologic features of CD30+ lymphoproliferative disorders and the need for adequate staging in such cases. This patient's CD30/CD8 coexpression is rare and has clinical and prognostic implications, including mucosally and acrally accentuated lesions and a potentially more aggressive course. Primary cutaneous CD30+ lymphoproliferative disorders have an excellent prognosis; therefore multiagent chemotherapy modalities are generally not indicated. The combination of T-cell lymphoma and sarcoidosis is also rare and may limit treatment options. PMID- 15280855 TI - Pyomyositis. AB - Four patients were admitted to Columbia Presbyterian Medical Center for evaluation of lower extremity pain and swelling. Three patients were initially misdiagnosed with cellulitis and one patient underwent evaluation for dermatomyositis. After consultation by the dermatologist, a correct diagnosis of pyomyositis was made clinically and confirmed by imaging, surgery, or an interventional procedure. Wound, blood, and urine cultures were positive for methicillin-sensitive Staphylococcus aureus in 100%, 50%, and 25% of patients, respectively. After the appropriate diagnosis and treatment, all patients experienced rapid resolution of symptoms and a favorable outcome. PMID- 15280857 TI - Interdigital psoriasis (psoriasis alba): renewed attention for a neglected disorder. PMID- 15280858 TI - Chronic cutaneous graft-versus-host disease in two children responds to UVA1 therapy: improvement of skin lesions, joint mobility, and quality of life. PMID- 15280859 TI - Circumscribed palmar hypokeratosis. PMID- 15280860 TI - Therapy for severe necrotizing vasculitis with infliximab. PMID- 15280861 TI - Thomas B. Fitzpatrick, 1919-2003. PMID- 15280862 TI - Update on clinically significant drug interactions in dermatology. PMID- 15280867 TI - Macrolides in dermatology. PMID- 15280868 TI - Rifampin in dermatology. PMID- 15280869 TI - Quinolones in dermatology. PMID- 15280870 TI - Linezolid, quinupristin/dalfopristin, and daptomycin in dermatology. PMID- 15280871 TI - Topical antimicrobial agents in dermatology. PMID- 15280872 TI - Drug interactions--fact or fiction? PMID- 15280874 TI - Peer review warts and all. PMID- 15280876 TI - Signaling by the sea. AB - In a spectacular location nestled in the hills on Croatia's coast, a group of 300 scientists from around the world gathered to listen to recent advances in cellular signaling and to gaze across the Adriatic Sea during discussions that surely put this work into perspective. Topics discussed ranged from precise structural details of signaling events to animal models for understanding signaling disorders. PMID- 15280877 TI - A master of its sulfate. PMID- 15280878 TI - Reversing DNA damage with a directional bias. PMID- 15280879 TI - Recognizing a phosphate group. PMID- 15280880 TI - Toward understanding GPCR dimers. PMID- 15280881 TI - Junctions on the road to cancer. PMID- 15280884 TI - Energizing depression research. PMID- 15280883 TI - Synergistic interactions between cannabinoids and environmental stress in the activation of the central amygdala. AB - Anxiety and panic are the most common adverse effects of cannabis intoxication; reactions potentiated by stress. Data suggest that cannabinoid (CB1) receptor modulation of amygdalar activity contributes to these phenomena. Using Fos as a marker, we tested the hypothesis that environmental stress and CB1 cannabinoid receptor activity interact in the regulation of amygdalar activation in male mice. Both 30 min of restraint and CB1 receptor agonist treatment (Delta9 tetrahydrocannabinol (2.5 mg/kg) or CP55940 (0.3 mg/kg); by i.p. injection) produced barely detectable increases in Fos expression within the central amygdala (CeA). However, the combination of restraint and CB1 agonist administration produced robust Fos induction within the CeA, indicating a synergistic interaction between environmental stress and CB1 receptor activation. An inhibitor of endocannabinoid transport, AM404 (10 mg/kg), produced an additive interaction with restraint within the CeA. In contrast, fatty acid amide hydrolase (FAAH) inhibitor-treated mice (URB597, 1 mg/kg) and FAAH-/- mice did not exhibit any differences in amygdalar activation in response to restraint compared to control mice. In the basolateral (BLA) and medial amygdala, restraint stress produced a low level of Fos induction, which was unaffected by cannabinoid treatment. Interestingly, the CB1 receptor antagonist SR141716 dose-dependently increased Fos expression in the BLA and CeA. These data suggest the CeA is an important neural substrate subserving the interactions between cannabinoids and environmental stress, and could be relevant to understanding the context dependent emotional and affective changes induced by marijuana intoxication and the role of endocannabinoid signaling in the modulation of amygdalar activity. PMID- 15280885 TI - Is PRG-1 a new lipid phosphatase? PMID- 15280887 TI - Mothering style and methylation. PMID- 15280888 TI - Controlling neuron number: does Numb do the math? PMID- 15280889 TI - Last call for adenosine transporters. PMID- 15280890 TI - A shrewd insight for vision. PMID- 15280892 TI - Communicating results to community residents: lessons from recent ATSDR health investigations. AB - As a public health agency within the US Department of Health and Human Services, the Agency for Toxic Substances and Disease Registry (ATSDR) is responsible for implementing the health-related provisions of the Superfund Act. Much of its work is carried out to address health concerns in communities near sources of environmental contamination, usually in consultation with other local, state, and federal agencies. Over the last decade, ATSDR has considered, supported or conducted health investigations in a variety of different communities across the country. Communication with community residents has been an integral part of the process in all of these activities. The approach to communicating results needs to begin early by developing relationships and clarifying expectations, and it needs to remain flexible. Through examples taken from specific situations, we illustrate many of the lessons we have gained from trying to apply the principles of good community involvement to the design and conduct of health investigations and to the communication of study results. PMID- 15280891 TI - Combined exposure to carbon disulfide and sulfuric acid simultaneously increases the risk of hand dermatitis in rayon industry. AB - OBJECTIVE: To evaluate the association between hand dermatitis (HD) and occupational exposure to CS(2) and to determine whether combined exposure to CS(2) and H(2)SO(4) exhibits a higher risk of HD. METHODS: In all, 110 subjects from a rayon factory were recruited and their exposure was classified into CS(2) exclusively, H(2)SO(4) exclusively, combined exposure, and nonexposure control based on workers' job characteristics. A dermatologist was designated in the diagnosis of HD on palm and dorsal sites for each subject. Other confounding factors including detergent, glove wearing, and participation in wet work were determined using a person-to-person questionnaire interview from 37 randomly selected subjects. RESULTS: Significant elevated odds ratios (ORs) for HD were found in CS(2) exclusively (44.8, P < 0.01) and combined exposure (49.0, P < 0.001) compared with control. Dose-response trends of ORs for HD were found across control, single exposure, and combined exposure for both CS(2) and H(2)SO(4). CONCLUSIONS: HD could occur resulting from occupational exposure to CS(2) alone. This study was unable to affirm that the exposure to H(2)SO(4) alone is associated with HD due to limited H(2)SO(4) exposure subjects. The combined exposure to both CS(2) and H(2)SO(4) simultaneously could increase the risk of HD. The control remedy in preventing dermal contact with either CS(2) or H(2)SO(4) among the rayon workers should be performed immediately. PMID- 15280893 TI - Factor analysis of pesticide use patterns among pesticide applicators in the Agricultural Health Study. AB - Exposure to certain pesticides has been linked with both acute and chronic adverse health outcomes such as neurotoxicity and risk for certain cancers. Univariate analyses of pesticide exposures may not capture the complexity of these exposures since use of various pesticides often occurs simultaneously, and because specific uses have changed over time. Using data from the Agricultural Health Study, a cohort study of 89,658 licensed pesticide applicators and their spouses in Iowa and North Carolina, we employed factor analysis to order to characterize underlying patterns of self-reported exposures to 50 different pesticides. Factor analysis is a statistical method used to explain the relationships between several correlated variables by reducing them to a smaller number of conceptually meaningful, composite variables, known as factors. Three factors emerged for farmer applicators (N=45,074): (1) Iowa agriculture and herbicide use, (2) North Carolina agriculture and use of insecticides, fumigants and fungicides, and (3) older age and use of chlorinated pesticides. The patterns observed for spouses of farmers (N=17,488) were similar to those observed for the farmers themselves, whereas five factors emerged for commercial pesticide applicators (N=4,384): (1) herbicide use, (2) older age and use of chlorinated pesticides, (3) use of fungicides and residential pest treatments, (4) use of animal insecticides, and (5) use of fumigants. Pesticide exposures did not correlate with lifestyle characteristics such as race, smoking status or education. This heterogeneity in exposure patterns may be used to guide etiologic studies of health effects of farmers and other groups exposed to pesticides. PMID- 15280894 TI - Genetic study of the forest pest Tomicus piniperda (Col., Scolytinae) in Yunnan province (China) compared to Europe: new insights for the systematics and evolution of the genus Tomicus. AB - The pine shoot beetle Tomicus piniperda is present throughout Eurasia. In Europe, it is considered as a secondary pest that rarely causes tree mortality, while heavy damage is observed in Yunnan Province (China) where it exhibits a novel aggregative behaviour during shoot attack. To understand why the ecological characteristics of the European and Chinese populations differ so strongly, we conducted an analysis of population genetic structure on 12 populations in Yunnan and one in JiLin using mitochondrial (COI-COII) and nuclear (ITS2 and 28S rDNA) DNA sequences, and compared the results to those obtained in France. We showed that the Yunnan populations differed markedly from French and JiLin populations. For all three markers, the genetic distances measured between the Tomicus from Yunnan and those from France were similar to distances previously observed between species. Similar distances were found between Yunnan and JiLin populations. Conversely, the distances between French and JiLin individuals were substantially lower, falling in the intraspecific range. We concluded that the individuals sampled in Yunnan belong to a new, undescribed species (Tomicus sp. nov.). We also showed that some individuals belong to the species T. brevipilosus that had never been recorded from this region before. Evolution of the genus Tomicus is discussed in the light of these new results. PMID- 15280895 TI - Molecular differentiation within and among island populations of the endemic plant Scalesia affinis (Asteraceae) from the Galapagos Islands. AB - Molecular variance was estimated in seven populations of the endemic species Scalesia affinis within and among islands of the Galapagos. The analysis, based on 157 polymorphic AFLP markers, revealed a high differentiation among populations, of which most was partitioned among islands. In addition, the information content of AFLP markers was tested with sets of discriminant analyses based on different numbers of AFLP markers. This indicated that the markers were highly informative in discriminating the populations. Although one of four populations from the island Isabela was sampled from a volcano 100 km away from the remaining populations, this population resembled the others on Isabela. The partitioning of molecular variance (AFLP) resulted in two unities, one consisting of populations from Isabela and one of populations from Santa Cruz and Floreana. The differentiation in two chloroplast microsatellites was higher than for AFLP markers and equally partitioned among populations within islands as among islands. Thus, gene flow via fruits within islands is as limited as among islands. The lower differentiation within islands in the nuclear AFLP markers may thus indicate that gene flow within islands is mostly accounted for by pollen transfer. S. affinis is the only species in the genus that is not listed in 2000 IUCN Red List of Threatened Species. However, due to prominent grazing and land exploitation, some populations have recently been reduced markedly, which was reflected in lower diversity. As inbreeding depression is present in the species, the rapid bottlenecks are threats to the populations. PMID- 15280896 TI - Additive and nonadditive genetic variation in avian personality traits. AB - Individuals of all vertebrate species differ consistently in their reactions to mildly stressful challenges. These typical reactions, described as personalities or coping strategies, have a clear genetic basis, but the structure of their inheritance in natural populations is almost unknown. We carried out a quantitative genetic analysis of two personality traits (exploration and boldness) and the combination of these two traits (early exploratory behaviour). This study was carried out on the lines resulting from a two-directional artificial selection experiment on early exploratory behaviour (EEB) of great tits (Parus major) originating from a wild population. In analyses using the original lines, reciprocal F(1) and reciprocal first backcross generations, additive, dominance, maternal effects ands sex-dependent expression of exploration, boldness and EEB were estimated. Both additive and dominant genetic effects were important determinants of phenotypic variation in exploratory behaviour and boldness. However, no sex-dependent expression was observed in either of these personality traits. These results are discussed with respect to the maintenance of genetic variation in personality traits, and the expected genetic structure of other behavioural and life history traits in general. PMID- 15280897 TI - Genetic correlations, tradeoffs and environmental variation. AB - Negative genetic correlations among traits are often used as evidence for tradeoffs that can influence evolutionary trajectories in populations. While there may be evidence for negative correlations within a particular environment, genetic correlations can shift when populations encounter different environmental conditions. Here we review the evidence for these shifts by focusing on experiments that have examined genetic correlations in more than one environment. In many studies, there are significant changes in correlations and these can even switch sign across environments. This raises questions about the validity of deducing genetic constraints from studies in one environment and suggests that the interaction between environmental conditions and the expression of genetic covariation is an important avenue for future work. PMID- 15280898 TI - Evolutionary genomics: foxy MHC selection story. PMID- 15280899 TI - A susceptibility gene for premature ovarian failure (POF) maps to proximal Xq28. AB - Terminal deletions of the long arm of the human X chromosome have been described in women with premature ovarian failure (POF). We report here the molecular characterization of an inherited deletion in two affected women and in their mother. The two daughters presented secondary amenorrhea at 17 or 22 years respectively, while the mother was fertile. She had four children, but she eventually had premature menopause at 43 years of age. The fine molecular analysis of the deletion showed that the three women carried an identical deletion. We conclude that the phenotypic difference within the family must be attributed to genetic or environmental factors and not to the presence of different extent deletions. By comparison with other deletions in the region, we map a susceptibility gene for POF to 4.5 Mb, in the distal part of Xq. PMID- 15280900 TI - Reduced genetic structure of the Iberian peninsula revealed by Y-chromosome analysis: implications for population demography. AB - Europe has been influenced by both intra- and intercontinental migrations. Since the Iberian peninsula was a refuge during the Last Glacial Maximum, demographic factors associated with contraction, isolation, subsequent expansion and gene flow episodes have contributed complexity to its population history. In this work, we analysed 26 Y-chromosome biallelic markers in 568 chromosomes from 11 different Iberian population groups and compared them to published data on the Basques and Catalans to gain insight into the paternal gene pool of these populations and find out to what extent major demographic processes account for their genetic structure. Our results reveal a reduced, although geographically correlated, Y-chromosomal interpopulation variance (1.2%), which points to a limited heterogeneity in the region. Coincidentally, spatial analysis of genetic distances points to a focal distribution of Y-chromosome haplogroups in this area. These results indicate that neither old or recent Levantine expansions nor North African contacts have influenced the current Iberian Y-chromosome diversity so that geographical patterns can be identified. PMID- 15280901 TI - Congenital deficiency of alpha feto-protein. AB - Alpha-fetoprotein (AFP) is the main fetus serum glycoprotein with a very low concentration in the adult. AFP deficiency is a rare phenomenon. We studied two families with congenital AFP deficiency and searched for mutations in the AFP gene. We identified one mutation of 2 base deletion in exon 8, in both families, that leads to the congenital deficiency of AFP. The mutation nt930-931delCT (T294fs25X) creates a frameshift after codon 294 that leads to a stop codon after 24 amino acids, thus truncating the normal length of AFP of 609 amino acids. All the affected children were found to be homozygous for the mutation as was one of the fathers. The affected individuals were asymptomatic and presented normal development. This first identification of a mutation in the AFP gene demonstrates for the first time that deficiency of AFP is compatible with human normal fetal development and further reproduction in males. PMID- 15280902 TI - Multivariate QTL linkage analysis suggests a QTL for platelet count on chromosome 19q. AB - Platelet count is a highly heritable trait with genetic factors responsible for around 80% of the phenotypic variance. We measured platelet count longitudinally in 327 monozygotic and 418 dizygotic twin pairs at 12, 14 and 16 years of age. We also performed a genome-wide linkage scan of these twins and their families in an attempt to localize QTLs that influenced variation in platelet concentrations. Suggestive linkage was observed on chromosome 19q13.13-19q13.31 at 12 (LOD = 2.12, P = 0.0009), 14 (LOD = 2.23, P = 0.0007) and 16 (LOD = 1.01, P = 0.016) years of age and multivariate analysis of counts at all three ages increased the LOD to 2.59 (P = 0.0003). A possible candidate in this region is the gene for glycoprotein VI, a receptor involved in platelet aggregation. Smaller linkage peaks were also seen at 2p, 5p, 5q, 10p and 15q. There was little evidence for linkage to the chromosomal regions containing the genes for thrombopoietin (3q27) and the thrombopoietin receptor (1q34), suggesting that polymorphisms in these genes do not contribute substantially to variation in platelet count between healthy individuals. PMID- 15280903 TI - Breast cancer: role of polymorphisms in biotransformation enzymes. AB - We aimed at determining whether any association exists between genetic polymorphisms in epoxide hydrolase (EPHX1), NADPH-quinone oxidoreductase (NQO1), glutathione S-transferases (GSTM1/P1/T1) and individual susceptibility to breast cancer. Polymerase chain reaction-restriction fragment length polymorphism-based genotyping assays were used to determine the frequency of polymorphisms in EPHX1 (exons 3 and 4), NQO1 (exon 6), GSTM1 (deletion), GSTP1 (exon 5), and GSTT1 (deletion) in a case-control study comprised of 238 patients with breast cancer and 313 healthy individuals. The distribution of genotypes in exon 6 of NQO1 was significantly different between the control group and breast cancer cases. Age adjusted odds ratio (OR) for variant genotype NQO1*2/*2 was 3.68 (confidence interval (CI) = 1.41-9.62, P = 0.008). Association of GSTP1*2/*2 genotype as well as that of low EPHX1 activity deduced by combinations of genotypes in exons 3 and 4 with breast cancer was suggestive, but nonsignificant. Individuals simultaneously lacking GSTM1 and carrying at least one GSTP1 variant allele were at significantly higher risk of breast cancer (OR = 2.03, CI = 1.18-3.50, P = 0.010). Combinations of either GSTM1null or GSTP1*2 with low activity of EPHX1 presented significant risk of breast cancer (OR = 1.88, CI = 1.00-3.52, P = 0.049 and OR = 2.40, CI = 1.15-5.00, P = 0.019, respectively) as well. In conclusion, the results suggest that genetic polymorphisms in biotransformation enzymes may play a significant role in the development of breast cancer. PMID- 15280904 TI - Clinical features and molecular analysis of seven British kindreds with hereditary hyperferritinaemia cataract syndrome. AB - Hereditary hyperferritinaemia cataract syndrome (HHCS) is an autosomal dominant disorder characterised by early onset cataracts and increased serum L-ferritin concentration. Affected individuals show nucleotide substitutions in the region of the L-ferritin gene (FTL) that encodes a regulatory sequence within the (mRNA)FTL termed the iron responsive element (IRE). We report the clinical features of seven HHCS kindreds containing 49 individuals with premature cataract. All the probands received diagnoses of HHCS after the incidental discovery of increased serum L-ferritin concentration (median 1420 microg/l; normal range 15-360 microg/l), in most cases during investigation or screening for anaemia. All the probands developed characteristic 'sunflower' morphology cataracts in childhood (median age at diagnosis 5 years), but had no other phenotypic features. All the affected kindreds showed nucleotide substitutions in FTL that were predicted to disrupt function of the (mRNA)FTL IRE. The severity of the clinical phenotype of HHCS was variable both within and between kindreds and showed no clear relationship to FTL genotype. HHCS should be included in the differential diagnosis of hyperferritinaemia and should be carefully distinguished from hereditary haemochromatosis. Measurement of the serum L ferritin concentration should be included in the investigation of all individuals with early onset cataracts. PMID- 15280905 TI - Detection of cardiovascular risk factors by anthropometric measures in Tehranian adults: receiver operating characteristic (ROC) curve analysis. AB - OBJECTIVE: To determine optimal cutoff points of anthropometric measures as cardiovascular indicators in an Iranian adult population. DESIGN: : Population based cross-sectional study. SETTING: Tehran, the capital of Iran. SUBJECTS: A total of 10 522 subjects (4449 men and 6073 women) aged 18-74 y. METHODS: Demographic data were collected and anthropometric indices were measured. Blood pressure was evaluated and hypertension was defined based on JNC VI. Biochemical measurements were done. Diabetes was considered as FBS> or =126 mg/dl (> or =7.0 mmol/l) and dyslipidemia was defined according to ATP II. Risk factors were considered as: hypertension, diabetes, dyslipidemia, 'hypertension or diabetes', 'hypertension and diabetes', 'hypertension or dyslipidemia', 'hypertension and dyslipidemia', ' diabetes or dyslipidemia', 'diabetes and dyslipidemia', 'at least one risk factor' and 'at least two risk factors'. ROC curve analysis was performed to determine optimal cutoff values-where the sensitivity approximates specificity. RESULTS: Younger men (the age category of 18-34 y) had higher WC than women. Men had higher waist-to-hip ratio (WHpR) and lower waist-to-height ratio (WHtR) than women in all age categories. Dyslipidemia, 'hypertension or dyslipidemia', 'diabetes or dyslipidemia' and 'at least one risk factor' were more prevalent risk categories in both genders. Range of areas under ROC curves for BMI and WC was 0.55-0.94 and 0.56-0.93 for men and 0.41-0.94 and 0.53-0.92 for women in various age groups, respectively. Range of areas for WHpR and WHtR in men was between 0.58-0.87 and 0.56-0.94, respectively, and for women varied between 0.53-0.91 and 0.53-0.90 in various age groups, respectively. Cutoff points of BMI for various risk factors were between 24 and 29 kg/m(2) in men and 25-31 kg/m(2) in women. Range of WHpR was between 0.86 and 0.97 in men and between 0.78 and 0.92 in women. Cutoff points for WC and WHtR were between 80 and 93 cm and 0.47 and 0.56 for men and between 79 and 96 cm and 0.50 and 0.63 for women in different age groups to detect various risk factors, respectively. In general, values were lowest for the most prevalent risk factors and highest for less prevalent conditions. CONCLUSION: The results suggested cutoff points of anthropometric measures as indicators of cardiovascular risk factors. It seems that these cut-points are higher for Iranians than for other Asian populations. SPONSORSHIP: Endocrine Research Center, Shaheed Beheshti University of Medical Sciences. PMID- 15280906 TI - Pulque intake during pregnancy and lactation in rural Mexico: alcohol and child growth from 1 to 57 months. AB - OBJECTIVE: To examine maternal intake of a mildly alcoholic beverage (pulque) during pregnancy and lactation, and its potential effect on postpartum child growth and attained size. DESIGN: A prospective cohort study that followed mothers (during pregnancy and lactation) and their offspring (from birth to approximately 57 months of age). SETTING: Six villages in rural, central Mexico. SUBJECTS: Subjects are 58 mother-child pairs. Pulque intake was measured as part of a dietary assessment that was conducted for 2days/month during pregnancy and early lactation. RESULTS: Most mothers consumed pulque during pregnancy (69.0%) and lactation (72.4%). Among pulque drinkers, the average ethanol intake was 125.1 g/week during pregnancy and 113.8 g/week during lactation. Greater pulque intake during lactation, independent of intake during pregnancy, was associated with slower weight and linear growth from 1 to 57 months, and smaller attained size at 57 months. Low-to-moderate pulque intake during pregnancy, in comparison to either nonconsumption or heavy intake, was also associated with greater stature at 57 months. CONCLUSIONS: Pulque intake during lactation may have adversely influenced postnatal growth in this population. Public health interventions are urgently needed in Mexico to reduce heavy intake of pulque by pregnant and lactating women, and to replace intake with foods that provide the vitamins and minerals present in the traditional alcoholic beverage. PMID- 15280907 TI - Prevalence and predictors of vitamin D deficiency in five immigrant groups living in Oslo, Norway: the Oslo Immigrant Health Study. AB - OBJECTIVE: To study the prevalence of vitamin D deficiency and to identify possible predictors of vitamin D deficiency in five main immigrant groups in Oslo. DESIGN: Cross-sectional, population-based. SETTING: City of Oslo. SUBJECTS: In total, 491 men and 509 women with native countries Turkey, Sri Lanka, Iran, Pakistan and Vietnam living in the county of Oslo. RESULTS: Median serum 25(OH)D level (s-25(OH)D) was 28 nmol/l, ranging from 21 nmol/l in women born in Pakistan to 40 nmol/l in men born in Vietnam. Overall prevalence of vitamin D deficiency defined as s-25(OH)D<25 nmol/l was 37.2%, ranging from 8.5% in men born in Vietnam to 64.9% in women born in Pakistan. s-25(OH)D did not vary significantly with age. s-25(OH)D was higher in blood samples drawn in June compared to samples obtained in April, but not significantly for women. Reported use of fatty fish and cod liver oil supplements showed a strong positive association with s-25(OH)D in all groups. Education length was positively associated with s-25(OH)D in women, whereas body mass index (BMI) was inversely associated with s-25(OH)D in women. These two variables were not related to vitamin D deficiency in men. CONCLUSIONS: There is widespread vitamin D deficiency in both men and women born in Turkey, Sri Lanka, Iran, Pakistan and Vietnam residing in Oslo. The prevalence of vitamin D deficiency is higher in women than in men, and it is higher in those born in Pakistan and lower in those born in Vietnam compared to the other ethnic groups. Fatty fish intake and cod liver oil supplements are important determinant factors of vitamin D status in the groups studied. BMI and education length are also important predictors in women. PMID- 15280908 TI - Prevalence of iron deficiency among schoolchildren of different socio-economic status in urban Turkey. AB - OBJECTIVE: To investigate the prevalence of iron deficiency among schoolchildren of different socio-economic status (SES), living in the three largest cities of Turkey. DESIGN: Cross-sectional study. SETTINGS: Primary schools of Istanbul, Ankara and Izmir. SUBJECTS: Schoolchildren aged 12 and 13 y (males: 504; females: 510) from nine primary schools. Inclusion of subjects in the study was on a voluntary basis. METHODS: Data were obtained on children SES, anthropometry, haematological and biochemical indices of iron status and consumption of food items related to dietary iron bioavailability. One-way analysis of variance was mainly applied, for the evaluation of the tested hypotheses, using Bonferroni correction in order to take into account the inflation of Type I error. RESULTS: Iron deficiency (serum ferritin <15 microg/l) prevalence was 17.5% among boys and 20.8% among girls. Furthermore, iron deficiency was significantly more prevalent among boys of lower SES, who were also found to have significantly lower levels of serum iron, serum ferritin, transferrin saturation, mean corpuscular haemoglobin and mean corpuscular haemoglobin concentration compared to those of higher SES. In terms of dietary factors affecting iron bioavailability, low SES boys exhibited significantly higher frequency of tea consumption and lower frequency of citrus fruit, red meat and fish consumption, compared to their higher SES counterparts. CONCLUSION: The prevalence of iron deficiency was relatively high, particularly among lower SES schoolboys. Higher tea and lower citrus fruits, red meat and fish consumption by boys of lower SES may provide an indication about the possible role of certain dietary patterns in the different manifestation of this medical condition among the socio-economic groups. However, further research is needed. PMID- 15280909 TI - Suicide gene therapy of sarcoma cell lines using recombinant adeno-associated virus 2 vectors. AB - Soft-tissue sarcomas are mesenchymal tumors that respond poorly to systemic chemotherapy. Suicide gene therapy may be an alternative treatment strategy. Here we show a high susceptibility of human sarcoma cell lines for recombinant adeno associated virus 2 (rAAV-2) suicide vectors: connective tissue sarcoma (HS-1), fibrosarcoma (HT-1080), Ewing sarcoma (RD-ES), Askin tumor (SK-N-MC), rhabdomyosarcoma (A-204) and soft-tissue sarcoma (WSKL-1). Several vectors containing the thymidine kinase (TK) gene under the control of either the cytomegalovirus promoter or the elongation-factor 1 alpha (EF1alpha) promoter were cloned and tested. Higher expression levels of the transgene were observed in the sarcoma lines when using the EF1alpha-suicide gene-containing vectors. A complete eradication of rAAV-2-EF1alpha-TK/eGFP (TK/enhanced green fluorescent protein fusion gene)-transduced tumor cells was shown following exposure to ganciclovir (2.5 microg/ml) in vitro, while at this dose level > 90% of mock transduced tumor cells survived. Xenotransplantation tumor models (intraperitoneal, subcutaneous) for the human sarcoma cell line HS-1 were established in nonobese diabetic/severe-combined immunodeficient mice. Mice transplanted with rAAV-2-EF1alpha-TK/eGFP-transduced and ganciclovir-exposed tumor cells survived > 5 months while in the nontransduced group all mice had died approximately 1 month after inoculation. These data hold promise for further development of rAAV-2-based suicide gene therapy of sarcomas. PMID- 15280910 TI - Sex differences in cancer risk among germline p53 mutation carriers. PMID- 15280913 TI - Serum CYFRA 21-1 (cytokeratin-19 fragments) is a useful tumour marker for detecting disease relapse and assessing treatment efficacy in breast cancer. AB - The usefulness of serum CYFRA 21-1 (cytokeratin-19 fragments) in monitoring the recurrence of breast cancer and in evaluating therapeutic effects was studied retrospectively. The sera from 173 patients with primary breast cancer or recurrent disease were measured for CYFRA 21-1, carcinoembryonic antigen (CEA), and carbohydrate antigen 15-3 (CA 15-3) levels. The positive rates of serum CYFRA 21-1 for stage IV (n=12) or recurrent disease (n=26) were 83.3 and 84.6%, respectively, while those of serum CEA were 41.7 and 26.9%, and those of serum CA 15-3 were 83.3 and 34.6%. The elevated preoperative levels of serum CYFRA 21-1 decreased to normal levels after curative operation, whereas the levels remained abnormally high after noncurative operation. There was a significantly high frequency of recurrence in patients with elevated levels of serum CYFRA 21-1 preoperatively compared to those with normal levels of the marker preoperatively. The serum CYFRA 21-1 levels were well correlated with response to chemotherapy. The positive rate of serum CYFRA 21-1 alone was higher than that of an assay combining CEA with CA 15-3, in both primary and recurrent cases (28.8 vs 18.8 and 84.6 vs 46.2%, respectively). These observations suggest that serum CYFRA 21-1 may be a reliable marker of recurrence or therapeutic efficacy. PMID- 15280914 TI - Anxiety and support in breast cancer: is this different for affluent and deprived women? A questionnaire study. AB - A postal questionnaire was sent to affluent and deprived women with breast cancer in order to compare psychosocial aspects of care with the purpose of understanding the balance of care and explaining why deprived women have poorer outcomes. Data were collected regarding reported sources of information, SF-36 scores and ongoing causes of anxiety. The results demonstrate that affluent women were more likely to have received information from their hospital specialist (94.8 vs 76.0%) and from a breast care nurse (70.1 vs 40.0%) than deprived women. They were also more likely to have received information from magazines (50.6 vs 33.0%), newspapers (45.5 vs 22.0%) and television news (45.5 vs 26.0%). Deprived women had poorer SF-36 scores than affluent women, and reported greater anxiety about money (12.2 vs 2.8%), other health problems (22.1 vs 8.2%) and family problems (17.5 vs 6.9%). Personal and professional support is clearly important for patients with breast cancer. Health professionals need to be aware of the greater psychological distress demonstrated by deprived women, even some years after diagnosis with breast cancer, and seek to address it. PMID- 15280915 TI - The utility of a multimedia education program for prostate cancer patients: a formative evaluation. AB - A multimedia program (MMP) was developed to educate patients with prostate cancer about their disease. A within-subjects design was used to investigate the changes in levels of cancer-related knowledge, psychosocial functioning, treatment decision-making role and information needs immediately after browsing the MMP. The participants were 67 men recently diagnosed with prostate cancer. Psychosocial functioning was assessed with 20 items describing common emotional states and coping strategies employed by cancer patients. Treatment decision making role was assessed with the Control Preference Scale. A principle component analysis of the 20 psychosocial items yielded three components: distress, positive approach and nonacceptance. After browsing the MMP significant increases in knowledge and reductions in distress were reported. Marital status was significantly associated with knowledge gain. Married men and those attending the study session with their spouse displayed a significant shift towards a more active role in treatment decisions. The majority of information needs were fulfilled by the MMP; however, information related to the likelihood of a cure, treatment side effects, coping strategies and aetiology were not completely satisfied by the MMP. Implications of the findings and suggestions for future work on the design and evaluation of the MMP are discussed. PMID- 15280916 TI - Cervical cancer screening programmes and policies in 18 European countries. AB - A questionnaire survey was conducted by the Epidemiology Working Group of the European Cervical Cancer Screening Network, and the International Agency for Research on Cancer, IARC, between August and December 2003 in 35 centres in 20 European countries with reliable cervical cancer incidence and/or mortality data in databanks held at IARC and WHO. The questionnaire was completed by 28 centres from 20 countries. The final tables included information on 25 centres from 18 countries. Six countries had started screening in the 1960s, whereas 10 countries or regions had started at least a pilot programme by 2003. There were six invitational and nine partially invitational programmes, the rest employing opportunistic screening only. Recommended lifetime number of smears varied from seven to more than 50. Coverage of smear test within the recommended screening interval (usually 3 or 5 years) was above 80% in three countries. Screening registration took place in 13 programmes. Eight programmes reported the rates of screen-detected cervical cancers and precursor lesions. There was wide variation in the CIN3 detection rates. International guidelines and quality assurance protocols are useful for monitoring and evaluating screening programmes systematically. Our survey indicated that the recommendations as currently given are met in only few European countries. Health authorities need to consider stronger measures and incentives than those laid out in the current set of recommendations. PMID- 15280917 TI - Incidence of childhood leukaemia in the vicinity of nuclear sites in France, 1990 1998. AB - Overall, 670 cases (O) of childhood leukaemia were diagnosed within 20 km of the 29 French nuclear installations between 1990 and 1998 compared to an expected number (E) of 729.09 cases (O/E=0.92, 95% confidence interval (CI)=[0.85-0.99]). Each of the four areas defined around the sites showed non significant deficits of cases (0-5 km: O=65, O/E=0.87, CI=[0.67-1.10]; 5-10 km: O=165, O/E=0.95, CI=[0.81-1.10]; 10-15 km: O=220, O/E=0.88, CI=[0.77-1.00]; 15-20 km: O=220, O/E=0.96, CI=[0.84-1.10]). There was no evidence of a trend in standardised incidence ratio with distance from the sites for all children or for any of the three age groups studied. Similar results were obtained when the start-up year of the electricity-generating nuclear sites and their electric nuclear power were taken into account. No evidence was found of a generally increased risk of childhood leukaemia around the 29 French nuclear sites under study during 1990 1998. PMID- 15280918 TI - Individual and joint impact of family history and Helicobacter pylori infection on the risk of stomach cancer: a nested case-control study. AB - We used 202 cases of stomach cancer and 394 controls nested within the Japan Collaborative Cohort Study For Evaluation of Cancer Risk (JACC study) to investigate whether family history has an independent effect on the risk of stomach cancer after controlling for the Helicobacter pylori infection. A positive history of stomach cancer in one or more first-degree relatives was associated with an increased risk of the disease in women, but not in men after controlling for H. pylori infection and other confounding variables. Women with both a family history and H. pylori infection were associated with more than five fold increased risk of the disease (OR 5.10, 95% CI 1.58-16.5) compared to those without these factors. These results suggest the existence of inherited susceptibility to the disease in women, and that measurements of H. pylori infection together with the family history allow meaningful evaluation of risk beyond that provided by either factor alone. PMID- 15280919 TI - Multi-institutional phase II trial of irinotecan, cisplatin, and etoposide for sensitive relapsed small-cell lung cancer. AB - Irinotecan (CPT-11) has been shown to exhibit excellent antitumour activity against small-cell lung cancer (SCLC). A multi-institutional phase II study was therefore conducted to evaluate the efficacy and toxicity of CPT-11 combined with cisplatin (CDDP) and etoposide (ETOP) (PEI regimen) for the treatment of sensitive relapsed SCLC. Patients who responded to first-line chemotherapy but relapsed more than 8 weeks after the completion of first-line therapy (n=40) were treated using the PEI regimen, which consisted of CDDP (25 mg m(-2)) weekly for 9 weeks, ETOP (60 mg m(-2)) for 3 days on weeks 1, 3, 5, 7, and 9, and CPT-11 (90 mg m(-2)) on weeks 2, 4, 6, and 8 with granulocyte colony-stimulating factor support. Five complete responses and 26 partial responses were observed, and the overall response rate was 78% (95% confidence interval 61.5-89.2%). The median survival time was 11.8 months, and the estimated 1-year survival rate was 49%. Grade 3/4 leucocytopenia, neutropenia, and thrombocytopenia were observed in 55, 73, and 33% of the patients, respectively. Nonhaematological toxicities were mild and transient in all patients. In conclusion, the PEI regimen is considered to be highly active and well tolerated for the treatment of sensitive relapsed SCLC. PMID- 15280920 TI - High expression of telomerase is an independent prognostic indicator of poor outcome in hepatoblastoma. AB - Telomerase, an enzyme related with cellular immortality, has been extensively studied in many kinds of malignant tumours for clinical diagnostic or prognostic utilities. Telomerase activity is mainly regulated by the expression of hTERT (human telomerase reverse transcriptase), which is a catalytic component of human telomerase. To evaluate whether the levels of hTERT mRNA provides a molecular marker of hepatoblastoma malignancy, we examined hTERT mRNA expression levels in the primary hepatoblastoma tissues by fluorescent RT-PCR using LightCycler technology and followed up the clinical outcomes in 63 patients listed in the Japanese Study Group of Pediatric Liver Tumor between 1991 and 2002. The hTERT mRNA expression was detected in 61 (96.8%) specimens and their expression levels ranged between 0.1/1000 and 745.1/1000 copies of PBGD gene that was used as an internal control. Among these cases, frozen 39 tumour samples and 14 adjacent noncancerous liver tissues were analysed for semiquantitative telomerase assay. In the 39 tumour samples, the levels of telomerase activity ranged between 0.11 and 2709 TPG and 12 (30.7%) had high telomerase activity (>100 TPG), whereas only nine of 14 noncancerous liver tissue samples showed telomerase activity which was less than 1.0 TPG. The levels of telomerase activity were significantly correlated with the levels of hTERT mRNA expression (P<0.001). The frequency of high hTERT mRNA expression and/or high telomerase activity did not significantly associate with the clinicopathological factors except for stage of disease. The prognosis of the patients with high hTERT mRNA expression was significantly worse than that of others (P<0.01), as was the patients with high telomerase activity (P<0.01). Multivariate analysis indicated that high levels of hTERT mRNA expression as well as telomerase activity are independent prognosis-predicting factors in patients with hepatoblastoma. PMID- 15280921 TI - Cytochrome P450 CYP1B1 activity in renal cell carcinoma. AB - Renal cell carcinoma (RCC) is the most common malignancy of the kidney and has a poor prognosis due to its late presentation and resistance to current anticancer drugs. One mechanism of drug resistance, which is potentially amenable to therapeutic intervention, is based on studies in our laboratory. CYP1B1 is a cytochrome P450 enzyme overexpressed in a variety of malignant tumours. Our studies are now elucidating a functional role for CYP1B1 in drug resistance. Cytochrome P450 reductase (P450R) is required for optimal metabolic activity of CYP1B1. Both CYP1B1 and P450R can catalyse the biotransformation of anticancer drugs at the site of the tumour. In this investigation, we determined the expression of CYP1B1 and P450R in samples of normal kidney and RCC (11 paired normal and tumour and a further 15 tumour samples). The O-deethylation of ethoxyresorufin to resorufin was used to measure CYP1B1 activity in RCC. Cytochrome P450 reductase activity was determined by following the reduction of cytochrome c at 550 nm. The key finding of this study was the presence of active CYP1B1 in 70% of RCC. Coincubation with the CYP1B1 inhibitor alpha-naphthoflavone (10 nM) inhibited this activity. No corresponding CYP1B1 activity was detected in any of the normal tissue examined (n=11). Measurable levels of active P450R were determined in all normal (n=11) and tumour samples (n=26). The presence of detectable CYP1B1, which is capable of metabolising anticancer drugs in tumour cells, highlights a novel target for therapeutic intervention. PMID- 15280922 TI - Nitrogen-containing bisphosphonates inhibit cell cycle progression in human melanoma cells. AB - Cutaneous melanoma is one of the highly malignant human tumours, due to its tendency to generate early metastases and its resistance to classical chemotherapy. We recently demonstrated that pamidronate, a nitrogen-containing bisphosphonate, has an antiproliferative and proapoptotic effect on different melanoma cell lines. In the present study, we compared the in vitro effects of three different bisphosphonates on human melanoma cell lines and we demonstrated that the two nitrogen-containing bisphosphonates pamidronate and zoledronate inhibited the proliferation of melanoma cells and induced apoptosis in a dose- and time-dependent manner. Moreover, cell cycle progression was altered, the two compounds causing accumulation of the cells in the S phase of the cycle. In contrast, the nonaminobisphosphonate clodronate had no effect on melanoma cells. These findings suggest a direct antitumoural effect of bisphosphonates on melanoma cells in vitro and further support the hypothesis of different intracellular mechanisms of action for nitrogen-containing and nonaminobisphosphonates. Our data indicate that nitrogen-containing bisphosphonates may be a useful novel therapeutic class for treatment and/or prevention of melanoma metastases. PMID- 15280923 TI - Dual blockade of EGFR and ERK1/2 phosphorylation potentiates growth inhibition of breast cancer cells. AB - One of the major targets for breast cancer therapy is the epidermal growth factor receptor (EGFR) and related receptors, which signal via different signal transduction pathways including the mitogen-activated protein kinase (MAPK) pathway. This study determined whether there is a correlation between EGFR/HER2 status and MAPK (ERK1/2) phosphorylation in breast cancer cells, and how this affects the response to an inhibitor of the receptors. Expression of EGFR, HER2 and phosphorylated ERK1/2 were measured by immunoblotting in a panel of breast cancer cell lines. Several lines expressed high levels of pERK1/2, with no obvious correlation with the level of EGFR/HER2. The EGFR tyrosine kinase inhibitor PKI166 inhibited growth and induced apoptosis in some cells with high levels of growth factor receptors (MDA-MB-468, SUM149, SKBR3), but was less effective in cells that also had high basal ERK1/2 activity (MDA-MB-231). The combination of an inhibitor of MAPK signalling (U0126) and PKI166 produced significantly more inhibition and apoptosis than either agent alone. This suggests that constitutive activation of the MAPK pathway may bypass inhibition of EGFR/HER2 tyrosine kinases, and lead to insensitivity to agents targeting the receptors. However, inhibiting both EGFR/HER2 and MAPK signalling can result in significant growth inhibition and apoptosis of EGFR-expressing breast cancer cells. PMID- 15280925 TI - CIITA methylation and decreased levels of HLA-DR in tumour progression. PMID- 15280926 TI - Diagnosing melanoma patients entering American Joint Committee on Cancer stage IV, C-reactive protein in serum is superior to lactate dehydrogenase. AB - Lactate dehydrogenase (LDH) in serum has recently been introduced into the American Joint Committee on Cancer (AJCC) staging system for cutaneous melanoma because of its prognostic value. We hypothesised LDH to be of value in discriminating melanoma patients entering AJCC stage IV from patients staying in AJCC stages I, II or III. Lactate dehydrogenase was compared to the acute phase protein C-reactive protein (CRP), which we observed to reflect the course of melanoma metastasis in a previous report. In this prospective study, we measured LDH and CRP in the serum of 91 consecutive melanoma patients progressing into AJCC stage IV in comparison to 125 patients staying in AJCC stages I, II or III. Comparing distributions of the parameters by median values and quartiles by Mann Whitney test, LDH was not significantly elevated in patients entering AJCC stage IV melanoma (P=0.785), whereas CRP was (P<0.001). Analysing the sensitivity and the specificity jointly by the areas under the receiver operating characteristics curves (ROC-AUC), LDH did not discriminate between the defined groups of patients (AUC=0.491; 95% confidence interval, 0.410, 0.581), whereas CRP did (AUC=0.933; 95% confidence interval, 0.900, 0.966; P<0.001). Upon logistic regression analysis to calculate the ROC-AUC values upon the predictive probabilities, LDH provided no additional information to CRP. Choosing a cutoff point of 3.0 mg l( 1), CRP yielded a sensitivity of 0.769 together with a specificity of 0.904 in diagnosing AJCC stage IV entry. Altogether, for first diagnosing AJCC stage IV melanoma, CRP is the superior serum marker when compared to the conventional LDH. PMID- 15280927 TI - UGT1A1 gene variations and irinotecan treatment in patients with metastatic colorectal cancer. AB - SN-38 is the active metabolite of irinotecan and it is metabolised through conjugation by uridine diphosphate glucuronosyl transferase (UGT1A1). The major toxicity of irinotecan therapy is diarrhoea, which has been related to the enzymatic activity of UGT1A1. We examined the influence of the UGT1A1 gene promoter polymorphism in the toxicity profile, in the response rate and in the overall survival (OS) in 95 patients with metastatic colorectal cancer treated with an irinotecan-containing chemotherapy. Genotypes were determined by analysing the sequence of TATA box of UGT1A1 of genomic DNA from the patients. Clinical parameters and genotypes were compared by univariate and multivariate statistical methods. The more frequent adverse effects were asthenia (34 patients), diarrhoea (29 patients) and neutropenia (20 patients). Severe diarrhoea was observed in 7/10 homozygous (70%) and 15/45 heterozygous (33%) in comparison to 7/40 (17%) wild-type patients (P=0.005). These results maintained the statistical significance in logistic regression analysis (P=0.01) after adjustment for other clinical relevant variables. The presence of severe haematological toxicity increased from wild-type patients to UGT1A1(*)28 homozygotes, but without achieving statistical significance. No relationship was found between the UGT1A1(*)28 genotypes and infection, nausea or mucositis. In univariate studies, patients with the UGT1A1(*)28 polymorphism showed a trend to a poorer OS (P=0.09). In the multivariate analysis, the genotype was not related to clinical response or to OS. The role of the UGT1A1 genotype as a predictor of toxicity in cancer patients receiving irinotecan demands the performance of a randomized trial to ascertain whether genotype-adjusted dosages of the drug can help to establish safe and effective doses not only for patients with the UGT1A1(*)28 homozygous genotype but also for those with the most common UGT1A1 6/6 or 6/7 genotype. PMID- 15280929 TI - Fludarabine, adriamycin and dexamethasone (FAD) in newly diagnosed advanced follicular lymphoma: a phase II study by the British National Lymphoma Investigation (BNLI). AB - The optimal first-line treatment for symptomatic patients with advanced stage follicular lymphoma remains unclear. Fludarabine-based combination regimens have been extensively used in relapsed disease and merit consideration as first-line therapy. We here report the results of a phase II study of FAD (fludarabine, adriamycin, dexamethasone) regimen in 30 patients with advanced stage follicular lymphoma requiring treatment. The response rate was in excess of 90% with 39% achieving a complete remission. The major toxicity was myelosuppression, but only 3% of cycles were associated with grade IV leucopenia. The high response rate has not translated into major improvements in failure-free survival and consideration must be given to alternative treatment modalities to consolidate the high rate of initial responses. PMID- 15280928 TI - S100A6 (Calcyclin) is a prostate basal cell marker absent in prostate cancer and its precursors. AB - S100A6 (Calcyclin) is a calcium-binding protein that has been implicated in a variety of biological functions as well as tumorigenesis. The aim of our study was to investigate the involvement of S100A6 during prostate cancer development and progression. Using immunohistochemistry, the expression of S100A6 was examined in benign (n=66), premalignant (n=10), malignant (n=66) and metastatic prostate (n=5) tissues arranged in a tissue-microarray or whole sections as well as in prostate cancer cell lines. The S100A6 immunostaining pattern in tissues was compared with that of cytokeratin 5 (a basal cell marker) and 18 (a benign luminal cell marker). In all cases of benign epithelium, intense S100A6 expression was seen in the basal cell layer with absent staining in luminal cells. In all cases of prostatic adenocarcinoma (matched), metastatic lesions and 3/10 high-grade prostatic intraepithelial neoplasia lesions, an absence of S100A6 was seen. Western blotting and RT-PCR analysis of cell lines showed S100A6 expression to be absent in LNCaP, LNCaP-LN3 and LNCaP-Pro5 but present in Du145, PC3, PC-3M and PC-3M-LN4. LNCaP cells treated with 5-Azacytidine, caused re expression of S100A6 mRNA. Sequencing of bisulphite modified DNA showed CpG methylation within the S100A6 promoter region and exon 1 of LNCaP, LNCaP-LN3 and LNCaP-Pro5 cell lines but not in Du145 cells. Our data suggest that loss of S100A6 protein expression is common in prostate cancer development and may occur at an early stage. The mechanism of loss of expression may involve hypermethylation of CpG sites. The finding of intense S100A6 expression in the basal cells of benign glands but loss of expression in cancer could be useful as a novel diagnostic marker for prostate cancer. PMID- 15280930 TI - A phase I trial of DNA vaccination with a plasmid expressing prostate-specific antigen in patients with hormone-refractory prostate cancer. AB - Prostate-specific antigen (PSA) is a serine protease secreted at low levels by normal luminal epithelial cells of the prostate and in significantly higher levels by prostate cancer cells. Therefore, PSA is a potential target for various immunotherapeutical approaches against prostate cancer. DNA vaccination has been investigated as immunotherapy for infectious diseases in patients and for specific treatment of cancer in certain animal models. In animal studies, we have demonstrated that vaccination with plasmid vector pVAX/PSA results in PSA specific cellular response and protection against tumour challenge. The purpose of the trial was to evaluate the safety, feasibility and biological efficacy of pVAX/PSA vaccine in the clinic. A phase I trial of pVAX/PSA, together with cytokine granulocyte/macrophage-colony stimulating factor (GM-CSF) (Molgramostim) and IL-2 (Aldesleukin) as vaccine adjuvants, was carried out in patients with hormone-refractory prostate cancer. To evaluate the biologically active dose, the vaccine was administered during five cycles in doses of 100, 300 and 900 microg, with three patients in each cohort. Eight patients were evaluable. A PSA-specific cellular immune response, measured by IFN-gamma production against recombinant PSA protein, and a rise in anti-PSA IgG were detected in two of three patients after vaccination in the highest dose cohort. A decrease in the slope of PSA was observed in the two patients exhibiting IFN-gamma production to PSA. No adverse effects (WHO grade >2) were observed in any dose cohort. We demonstrate that DNA vaccination with a PSA-coding plasmid vector, given with GM-CSF and IL-2 to patients with prostate cancer, is safe and in doses of 900 microg the vaccine can induce cellular and humoral immune responses against PSA protein. PMID- 15280931 TI - ATM polymorphisms as risk factors for prostate cancer development. AB - The risk of prostate cancer is known to be elevated in carriers of germline mutations in BRCA2, and possibly also in carriers of BRCA1 and CHEK2 mutations. These genes are components of the ATM-dependent DNA damage signalling pathways. To evaluate the hypothesis that variants in ATM itself might be associated with prostate cancer risk, we genotyped five ATM variants in DNA from 637 prostate cancer patients and 445 controls with no family history of cancer. No significant differences in the frequency of the variant alleles at 5557G>A (D1853N), 5558A>T (D1853V), ivs38-8t>c and ivs38-15g>c were found between the cases and controls. The 3161G (P1054R) variant allele was, however, significantly associated with an increased risk of developing prostate cancer (any G vs CC OR 2.13, 95% CI 1.17 3.87, P=0.016). A lymphoblastoid cell line carrying both the 3161G and the 2572C (858L) variant in the homozygote state shows a cell cycle progression profile after exposure to ionising radiation that is significantly different to that seen in cell lines carrying a wild-type ATM gene. These results provide evidence that the presence of common variants in the ATM gene, may confer an altered cellular phenotype, and that the ATM 3161C>G variant might be associated with prostate cancer risk. PMID- 15280932 TI - Comparative pharmacology of oral fluoropyrimidines: a focus on pharmacokinetics, pharmacodynamics and pharmacomodulation. AB - The main purpose of the present review article was to shed light on the different 5-fluorouracil (5-FU) prodrugs by underlining their respective pharmacological features in terms of metabolic activation, dihydropyrimidine dehydrogenase inhibition, pharmacokinetic profile and biomodulation ability. Oral fluoropyrimidines differ particularly as concerns their pharmacokinetic profile and especially in the delivery of circulating 5-FU. More clinical studies need to be performed incorporating tumour predictive markers during oral fluoropyrimidine based treatment. The new possibilities are to achieve pharmacomodulation of oral fluoropyrimidines, notably for UFT and capecitabine, that open up the prospect of establishing significant novel treatment protocols based on drug combinations. PMID- 15280933 TI - Lung cancer management concerns in New Zealand. PMID- 15280934 TI - Inhaled corticosteroids in asthma action plans--double or quits? PMID- 15280935 TI - Lung cancer treatment in New Zealand: physician's attitudes. AB - AIMS: To determine treatment practices of New Zealand physicians who manage non small cell lung cancer (NSCLC). METHODS: A questionnaire on the treatment of NSCLC was emailed to all respiratory physicians, medical oncologists, and radiation oncologists in New Zealand. Respondents were asked to select the treatment they would offer in six lung cancer case scenarios. RESULTS: Thirty-one (81%) respiratory physicians, 15 (71%) medical oncologists, and 8 (30%) radiation oncologists responded to the questionnaire. Surgery was selected (by all groups) as the best option for early-stage disease NSCLC. Radiotherapy or combination chemo/radiotherapy (for locally advanced disease) was favoured by 37% of respiratory physicians for stage IIIa and 28% for stage IIIb--compared with medical oncologists (100% and 80%) and radiation oncologists (86% and 28%). Chemotherapy for 'fit' patients with advanced disease was favoured by only 11% of respiratory physicians, compared with 67% of medical oncologists and 33% of radiation oncologists. Best supportive care (BSC) was the favoured treatment for patients with advanced disease with poor performance patients. CONCLUSION: This study demonstrates considerable heterogeneity in the choice of treatment for NSCLC between specialities, particularly for locally advanced and advanced disease. These findings suggest international guidelines are not being adhered to, and variations in treatment may potentially have outcome implications for patients. PMID- 15280936 TI - Application of asthma action plans to childhood asthma: national survey repeated. AB - AIMS: Define the way childhood asthma action plans are currently being used in New Zealand; determine New Zealand doctor's recommendations about the use of an increased dose of inhaled steroids in asthma action plans; and determine if there has been any change (during the last 7 years) in the way asthma action plans are used. METHODS: A postal survey was sent to all 297 paediatricians and paediatric registrars in New Zealand, and to a random sample of 500 general practitioners (GPs). The questions related to asthma action plan use, the inclusion of an increased dose of inhaled steroid in those plans, and details of the way doctor's adjusted inhaled steroid dose. Comparisons were made for selected questions between this survey and the same survey conducted in 1995. RESULTS: Valid responses were received from 179 (60%) paediatricians and paediatric registrars, and 233 (47%) GPs. 165 (70.8%) GPs and 137 (76.5%) paediatricians/paediatric registrars indicated that they used written action plans for children with asthma in their care. 184 (61.5%) respondents who used asthma action plans included a step involving an increase in the dose of inhaled steroid, compared to 83.6% in 1995 (p<0.001). GPs in 2002 were less likely to use action plans (p<0.001) and include a step with an increased dose of inhaled steroid (p=0.003) Paediatricians and paediatric registrars in 2002 were just as likely to use action plans (p=0.549), but less likely to include a step with an increased dose of inhaled steroid (p<0.001). GPs in 2002 were significantly more likely (than paediatricians and paediatric registrars) to include a step involving an increased dose of inhaled steroid (p<0.001) There has been a change in the practice of New Zealand GPs, paediatricians, and paediatric registrars--with a decreased tendency to double the dose of inhaled steroids in childhood action plans, thus suggesting doctors are cognisant of conclusions drawn by 'evidence based medicine'. CONCLUSIONS: There has also been a decline in the proportion of asthmatic children receiving a written asthma action plan, and this is inconsistent with recommendations contained in consensus documents. PMID- 15280937 TI - Economic cost of community-acquired pneumonia in New Zealand adults. AB - AIMS: The aim of this study was to evaluate the economic cost of community acquired pneumonia (CAP) in New Zealand adults. Although this is an important illness, there is little published information on the national costs of treatment. Without such information, new treatment options cannot be evaluated in economic terms. METHODS: Costs were estimated from a societal perspective for the adult population (aged 15 years and over) using New Zealand age-specific hospital admission rates (average of 2000-2002), population data (2003), and unit costs (2003) in combination with international data on the proportion of pneumonia cases hospitalised. Univariate and multivariate sensitivity analyses were used to determine the major cost drivers and evaluate uncertainty in the estimates. RESULTS: It was estimated that in 2003 there were 26,826 episodes of pneumonia in adults; a rate of 859 per 100,000 people. The annual cost was estimated to be 63 million dollars, (direct medical costs of 29 million dollars; direct non-medical costs of 1 million dollars; lost productivity of 33 million dollars). CONCLUSIONS: The major generators of costs for community-acquired pneumonia are the number of hospitalisations (particularly for the group aged 65 years and over) and loss of productivity. Intensified prevention and effective community treatment programmes focussing on the 65 years and older age groups should be investigated (as they have the greatest potential to reduce healthcare costs). PMID- 15280938 TI - End of life decision-making by New Zealand general practitioners: a national survey. AB - AIM: To explore type and incidence of medical decisions at the end of life that hasten death made by general practitioners in New Zealand, within the context of access to palliative care. METHOD: An anonymous questionnaire investigating the last death attended in the previous 12 months was sent to 2602 general practitioners (GPs) in New Zealand. RESULTS: From a 48% (1255) response, 88.9% (1116) GPs indicated access to an interdisciplinary pain management or palliative care team. Of those attending a death in the previous 12 months, 63% (693) had made a prior medical decision. These decisions included withdrawing/withholding treatment or increasing pain relief with (a) probability death would be hastened 61.8% (428) or (b) partly or explicitly to hasten death 32.6% (226). Moreover, death was caused by a drug supplied or administered by the GP in 5.6% cases (39), actions consistent with physician-assisted death. CONCLUSION: Physician-assisted death provided by some general practitioners in New Zealand is occurring within the context of available palliative care. PMID- 15280940 TI - Familial primary pulmonary hypertension. PMID- 15280939 TI - Caring for patients and families at the end of life: withdrawal of intensive care in the patient's home. AB - AIM: To describe our experience of transporting 17 intensive care patients home to die. DESIGN: A brief report. SETTING: Mixed medical/surgical intensive care unit (ICU). RESULTS: After discussions with their families, 17 adult patients in whom ongoing care was deemed either inappropriate or futile were transported home. Once there, intensive care modalities of ventilation and vasopressor therapy were withdrawn. The patients were sedated initially with intravenous morphine and if death was not immediately imminent, subcutaneous morphine was administered. In these cases where death took longer than 2 hours, the patients were managed with the assistance of district nurses, the family general practitioner, or staff from the South Auckland Hospice. CONCLUSIONS: All the patients in this report were Maori or Polynesian and all families reported this as a positive experience. Since completion of this report, we have taken our first European patient home to die. PMID- 15280941 TI - Nutritional supplements: friend or foe? PMID- 15280942 TI - Osteomalacia: recovery of bone density. PMID- 15280943 TI - Don't forget about HIV. PMID- 15280944 TI - Safety concerns about nuclear-powered vessels persist. PMID- 15280945 TI - The merger of natural product synthesis and medicinal chemistry: on the chemistry and chemical biology of epothilones. AB - Based on epothilones as powerful natural product leads several promising new anticancer agents have emerged through concerted efforts in chemistry and biology. PMID- 15280946 TI - Microtubule structure and its stabilisation. AB - Microtubules are designed to be dynamically unstable. GTP hydrolysis converts an initially stable polymeric structure into an unstable one in which strain at the interfaces between longitudinal neighbours in the helical lattice of subunits is balanced by lateral interactions. However, stability can be modulated by a variety of factors, including associated proteins and a variety of drug molecules. Stabilising drugs such as Taxol and the assembly-promoting repeat motifs of tau protein occupy a special pocket in beta-tubulin. Microtubule destabilizing drugs such as colchicine alter the longitudinal interfaces of the subunits so that they cannot assemble into a microtubule lattice. These mechanisms are discussed in terms of the atomic structure of the protein. PMID- 15280947 TI - Palladium--phosphinous acid-catalyzed Sonogashira cross-coupling reactions in water. AB - A palladium--phosphinous acid-catalyzed Sonogashira cross-coupling reaction that proceeds in water under air atmosphere in the absence of organic co-solvents has been developed. Disubstituted alkynes have been prepared in up to 91% yield by POPd-catalyzed coupling of various aryl halides including chlorides in the presence of tetrabutylammonium bromide and pyrrolidine or NaOH. PMID- 15280948 TI - Compelling evidence for a stepwise mechanism of the alkaline cyclisation of uridine 3'-phosphate esters. AB - A Bronsted graph with a convex break at pK(a)(Lg)= 12.58 provides compelling evidence for an intermediate in the alkaline cyclisation of uridine 3'-phosphate esters. The transient pentacoordinated oxyphosphorane dianion intermediate collapses to reactant and cyclic uridine 2',3'-monophosphate faster than it can pseudo-rotate and isomerise to the 2'-isomer. PMID- 15280949 TI - Highly enantioselective reaction of lithiated N-Boc-thiazolidine: a new chiral formyl anion equivalent. AB - Reaction of lithiated N-Boc-thiazolidine with various aldehydes in the presence of (-)-sparteine afforded the products with up to 93% ee. The reaction was confirmed to proceed through a dynamic thermodynamic resolution pathway. Each diastereomeric alcohol could be converted to the corresponding optically active 1,2-ethanediols. PMID- 15280950 TI - Indium mediated allylation of quinoline and isoquinoline activated by phenyl chloroformate. AB - Quinoline and isoquinoline activated by phenyl chloroformate were allylated using indium and allyl bromides in THF at room temperature to give the corresponding allyldihydroquinoline and allyldihydroisoquinoline in good to high yields. PMID- 15280951 TI - Synthesis and biological evaluation of caulibugulones A-E. AB - The marine bryozoan metabolites caulibugulone A-E were prepared from a readily available isoquinoline dione. These natural products were found to be potent and selective inhibitors of the dual specificity phosphatase Cdc25B. PMID- 15280952 TI - Nucleophilic and general acid catalysis at physiological pH by a designed miniature esterase. AB - A 31-residue peptide (Art-Est) was designed to catalyse the hydrolysis of p nitrophenyl esters through histidine catalysis on the solvent exposed face of the alpha-helix of bovine pancreatic polypeptide. NMR spectroscopy indicated that Art Est adopted a stable 3-dimensional structure in solution. Art-Est was an efficient catalyst with second order rate constants of up to 0.050 M(-1) s(-1). The activity of Art-Est was a consequence of the increased nucleophilicity of His 22, which had a reduced pK(a) value of 5.5 as a consequence of its interaction with His-18 and the positively charged Arg-25 and Arg-26. Mass spectrometry and NMR spectroscopy confirmed that the Art-Est catalysed hydrolysis of p-nitrophenyl esters proceeded through an acyl-enzyme intermediate. A solvent kinetic isotope effect of 1.8 indicated that the transition state preceding the acyl intermediate was stabilised through interaction with the protonated side-chain of His-18 and indicated a reaction mechanism similar to that generally observed for natural esterases. The involvement in the reaction of two histidine residues with different pK(a) values led to a bell-shaped dependence of the reaction rate on the pH of the solution. The catalytic behaviour of Art-Est indicated that designed miniature enzymes can act in a transparent mechanism based fashion with enzyme-like behaviour through the interplay of several amino acid residues. PMID- 15280953 TI - beta-Pseudopeptide foldamers. The homo-oligomers of (4R)-(2-oxo-1,3-oxazolidin-4 yl)-acetic acid (D-Oxac). AB - A total synthesis in solution and a conformational analysis of the homo-oligomers of (4R)-(2-oxo-1,3-oxazolidin-4-yl)-acetic acid (D-Oxac) to the tetramer level are described. As the D-Oxac building block contains both an oxazolidin-2-one and a beta-amino acid group, it may represent a new type of conformationally constrained tool for the construction of beta-pseudopeptide foldamers. A conformational investigation using NMR and an extensive, unconstrained analysis with a Monte Carlo search to the octamer level, both in water and in chloroform, showed that these homo-oligomers tend to fold in a regular helical structure in a competitive solvent, such as water. Since aqueous solutions are of major relevance for biological systems, these molecules are good candidates for application to these environments. PMID- 15280954 TI - Selective activation of glycosyl donors utilising electrochemical techniques: a study of the thermodynamic oxidation potentials of a range of chalcoglycosides. AB - A series of six chalcoglycosides (phenyl-2,3,4,6-tetra-O-benzoyl-1-seleno-beta-D glucopyranoside, phenyl-2,3,4,6-tetra-O-benzyl-1-seleno-beta-D-glucopyranoside, phenyl-2,3,4,6-tetra-O-benzyl-1-thio-beta-D-glucopyranoside, p-tolyl-2,3,4,6-O benzoyl-1-thio-beta-D-glucopyranoside, p-tolyl-2,3,4,6-O-benzyl-1-thio-beta-D glucopyranoside, and phenyl-2,3,4,6-O-benzyl-beta-D-glucopyranoside) are voltammetrically interrogated in dimethyl sulfoxide, so as to determine their formal (i.e. thermodynamic) redox potentials. The electrochemical oxidation of the chalcoglycoside is shown to follow an overall EC-type mechanism, in which the electro-generated cation radical undergoes an irreversible carbon-chalcogen bond rupture to produce the corresponding glycosyl cation, which may react further. The kinetics of the initial heterogeneous electron transfer process and subsequent irreversible homogeneous chemical degradation of the radical cation are reported, with values for the standard electrochemical rate constant k(0) in the order of 10(-2) cm s(-1) and the first order homogeneous rate constant, k(1), of the order of 10(3) s(-1). The formal oxidation potentials were found to vary according to the identity of the chalcogenide, such that OPh > SPh similar to STol > SePh. PMID- 15280955 TI - Selective electrochemical glycosylation by reactivity tuning. AB - Electrochemical glycosylation of a selenoglycoside donor proceeds efficiently in an undivided cell in acetonitrile to yield beta-glycosides. Measurement of cyclic voltammograms for a selection of seleno-, thio-, and O-glycosides indicates the dependence of oxidation potential on the anomeric substituent allowing the possibility for the rapid construction of oligosaccharides by selective electrochemical activation utilising variable cell potentials in combination with reactivity tuning of the glycosyl donor. A variety of disaccharides are readily synthesised in high yield, but limitations of the use of selenoglycosides as glycosyl donors for selective glycosylation of thioglycoside acceptors are exposed. The first electrochemical trisaccharide synthesis is described. PMID- 15280956 TI - Synthesis of fluorescent oligonucleotide--EYFP conjugate: towards supramolecular construction of semisynthetic biomolecular antennae. AB - A novel species of DNA--protein conjugate was synthesized by chemically linking DNA oligonucleotides to Aequorea victoria green fluorescent protein mutant EYFP. An additional cysteine was added to the C-terminus of the EYFP by genetic engineering and used to covalently attach amino-modified oligonucleotide with the aid of the heterobifunctional crosslinker sSMCC. EYFP maintained its fluorescence upon conjugation. The oligonucleotide provides an additional binding site to the fluorescent protein, and hence, the EYFP conjugate could be specifically hybridized with both complementary DNA-protein conjugates in-solution as well as immobilized at capture oligonucleotides attached to a solid substrate. These studies are paving the way for future applications in the self-assembly of photoactive supramolecular complexes, such as artificial light-harvesting systems. PMID- 15280957 TI - Photochemical and photophysical properties of a poly(propylene amine) dendrimer functionalized with E-stilbene units. AB - A second generation poly(propylene amine) dendrimer functionalized at the periphery with eight E-stilbene and eight 4-tert-butylbenzenesulfonyl units has been prepared. The absorption spectrum, fluorescence spectrum and decay, E<==>Z photoisomerization, and photocyclization of the Z-isomer of the stilbene units have been investigated in air equilibrated acetonitrile solutions. For comparison purposes, a reference compound of the peripheral dendrimer units, namely 4-tert butyl-N-propyl-N-(4-styryl-benzyl)-benzenesulfonamide, has also been studied. The quantum yield of the E-->Z photoisomerization reaction (0.30) and the fluorescence quantum yield of the E isomer (0.014) are substantially smaller for the units appended to the dendrimer compared to those of the reference compound (0.50 and 0.046, respectively). The presence of a red tail and the biexponential decay of the emission band of the dendrimer indicate formation of excimers between the stilbene units appended at the poly(propylene amine) dendritic structure. Under the experimental conditions used (lambda(exc)= 313 nm), a Z/E photostationary state (around 9 : 1 for both reference compound and dendrimer ) is reached in the time scale of minutes. On continuing irradiation, other photoreactions take place in the time scale of hours: the stilbene moiety of compound undergoes photocyclization to phenanthrene (quantum yield 0.015), whereas in dendrimer photocyclization to phenanthrene is accompanied by other processes, including a photoreaction involving the internal amine groups. PMID- 15280958 TI - Electrophilic and oxidative chemistry of pyrene and its non-alternant isomers: theoretical (DFT, GIAO-NMR, NICS) study of protonation carbocations and oxidation dications from pyrene, azupyrene (dicyclopenta[ef,kl]heptalene) and dicyclohepta[ed,gh]pentalene. AB - Mono- and diprotonated carbocations and the two-electron oxidation dications derived from parent pyrene and its nonalternant isomers "azupyrene"(dicyclopenta[ef,kl]heptalene)(DCPH) and dicyclohepta[ed,gh]pentalene (DCHP) were studied by DFT at the B3LYP/6-31G(d) level. The most likely site(s) for mono- and diprotonation were determined based on relative arenium ion energies and the structures of the energetically most favored carbocations were determined by geometry optimization. The NMR chemical shifts for the protonated mono- and dications and the oxidation dications were computed by GIAO-NMR at the B3LYP/6-31G(d)//B3LYP/6-31G(d) level and their charge delocalization paths were deduced based on magnitude of the computed [capital Delta][small delta](13)C values and the NPA-derived changes in charges. Relative aromaticity/antiaromaticity in various rings in the energetically favored mono- and dications was estimated via NICS and [capital Delta]NICS. Calculated NMR chemical shift data for and were compared with the available experimental NMR values. The available data on chemical and physical properties of DCPH and DCHP are extremely limited and biological activity data are non-existent. The present study provides the first glance into their carbocations and oxidation dications, while augmenting and reinforcing the previous stable ion data on the pyrenium cations. PMID- 15280959 TI - Enantioselective synthesis of the dioxabicyclo[3.2.1]octane core of the zaragozic acids via intramolecular Wacker-type cyclisation reactions. AB - Wacker-type cyclisation reactions provide an effective entry into the dioxabicyclo[3.2.1]octane core of the zaragozic acid analogues. PMID- 15280960 TI - Reactions of the natural lignan hydroxymatairesinol in basic and acidic nucleophilic media: formation and reactivity of a quinone methide intermediate. AB - The chemical properties and synthetic modifications of the natural lignan hydroxymatairesinol in basic and acidic nucleophilic media were studied. Hydroxymatairesinol presumably reacts via a quinone methide and a carbonium ion mechanism under basic and acidic conditions, respectively. In these conditions the benzylic hydroxyl group was displaced by nucleophiles yielding new 7 substituted butyrolactone lignans. Reactions in alcoholic basic solutions yielded the 7-alkoxy ethers diastereoselectively. Several previously known lignans as well as new lignans and lignan derivatives were synthesised. The transformations were monitored and the products identified by HPLC-MS and NMR. PMID- 15280961 TI - Triterpenoid total synthesis. Synthesis and absolute configuration of mispyric acid. AB - The first synthesis of mispyric acid (1), an inhibitor of DNA polymerase beta with a novel triterpene skeleton, was achieved by starting from isoprene (2), geraniol (6) and 1,5-dimethoxy-1,4-cyclohexadiene (14). The absolute configuration of the naturally occurring 1 was determined as 2S,4S. PMID- 15280962 TI - Cu(II)-Mediated decomposition of phosphorothionate P=S pesticides. Billion-fold acceleration of the methanolysis of fenitrothion promoted by a simple Cu(II) ligand system. AB - The kinetics of methanolysis of the title compound (3) were studied in the presence of Cu(2+), introduced as Cu(OTf), in the presence of 0.5-1.0 eq. of methoxide and in the presence of 1.0 eq. of a ligand such as bipyridyl (5), phenanthroline (6) or 1,5,9-triazacyclododecane (4). In all cases the active species involve Cu(2+)((-)OCH(3)). In the case of added strong-binding ligands 5 or 6, a plot of the observed rate constant for methanolysis of 3 vs. [Cu(2+)](total) gives a curved line modelled by a process having a [Cu(2+)](1/2) dependence consistent with an active monomeric species in equilibrium with an inactive dimer i.e.[LCu(2+)((-)OCH(3))](2) <==> 2LCu(2+)((-)OCH(3)). In the case of the added strong binding ligand 4, the plot of the observed rate constant for methanolysis of 3 vs.[Cu(2+)](total) gives a straight line consistent with the catalytically active species being Cu(2+)(OCH(3)) which shows no propensity to form inactive dimers. Turnover experiments where the [3] > [Cu(2+)](total) indicate that the systems are truly catalytic. In the optimum case a catalytic system comprising 1 mM of the complex 4Cu(2+)((-)OCH(3)) catalyzes the methanolysis of 3 with a t(1/2) of approximately 58 s accounting for a 1.7 x 10(9)-fold acceleration relative to the background reaction at near neutral (s)(s)pH (8.75). PMID- 15280963 TI - Heck reaction catalyzed by Pd/C, in a triphasic-organic/Aliquat 336/aqueous solvent system. AB - The rate of the Pd/C catalyzed Heck coupling of Ar-I with CH(2)=CH-R is accelerated tenfold by the presence of Aliquat 336 (A336), a well known phase transfer catalyst, and an ionic liquid. Both when conducted in A336 as solvent, and in an isooctane/A336/aqueous triphasic mixture, the Heck reaction of aryl iodides with electron deficient olefins, catalyzed by Pd/C, proceeds with high yields and selectivity. When KOH is used instead of Et(3)N, selective formation of the biphenyl rather than the Heck product, is observed. Aryl bromides react more sluggishly, and only the more activated ones undergo the Heck reaction. In the absence of the olefin, aryl halides possessing an electron withdrawing group are reduced to the corresponding Ar-H. PMID- 15280964 TI - Photochemistry of the three carboxypyridines in water: a pH dependent reaction. AB - The photochemistry of ortho, meta and para-carboxypyridines (pK(a)(1)= 1.0-2.1 and pK(a)(2)= 4.7-5.3) in aqueous medium was studied by laser-flash photolysis and product studies. At pH < pK(a)(1), hydroxylated compounds are produced with low quantum yields. Within the pH range 4-7, ortho and meta isomers undergo dimerization together with decarboxylation with a quantum yield showing a very sharp maximum around pK(a)(2)([small phi](max)= 0.09 and 0.01, respectively) while the para isomer is photostable. End-of-pulse transients assigned to triplet states were detected by laser-flash photolysis at pH < pK(a)(1) and pH > 4. Additionally, the carboxypyridinyl radicals were detected as secondary intermediates at pH < pK(a)(1) and 4 < pH < 7 and the OH-adduct radicals at pH < pK(a)(1). This is in favour of an electron transfer reaction between triplet and starting compound producing a charge transfer species. The radical anion would escape as carboxypyridinyl radical while the radical cation may add water at pH < pK(a)(1) yielding the OH-adduct radical or may undergo decarboxylation at pH > 4. The high quantum yield of phototransformation of the ortho isomer at pH > 4 is due to an easy decarboxylation process. A reaction scheme is proposed accounting for the dependences of [small phi] on both the pH and the carboxypyridines concentration. This study points out the distinct pattern of reactivity of carboxypyridines depending on the ionisation state of starting compounds and isomeric substitution. PMID- 15280965 TI - The natural constituents of historical textile dyes. AB - The sources and structures of dyes used to colour Western historical textiles are described in this tutorial review. Most blue and purple colours were derived from indigo--obtained either from woad or from the indigo plant--though some other sources (e.g. shellfish and lichens) were used. Reds were often anthraquinone derivatives obtained from plants or insects. Yellows were almost always flavonoid derivatives obtained from a variety of plant species. Most other colours were produced by over-dyeing--e.g. greens were obtained by over-dyeing a blue with a yellow dye. Direct analysis of dyes isolated from artefacts allows comparison with the historical record. PMID- 15280966 TI - Combined approach using capillary electrophoresis and NMR spectroscopy for an understanding of enantioselective recognition mechanisms by cyclodextrins. AB - This tutorial review describes the contribution of chiral capillary electrophoresis in combination with other instrumental techniques, especially nuclear magnetic resonance spectroscopy, to a better understanding of the chiral recognition mechanisms by cyclodextrins. Aspects such as affinity pattern of enantiomers towards various cyclodextrins as well as the stoichiometry of the resulting complexes, the equilibrium constants and the structure of complexes are addressed. In addition to the aforementioned techniques, the usefulness of complementary instrumental and molecular modeling techniques for an understanding of the chiral recognition mechanisms of cyclodextrins is also illustrated. PMID- 15280967 TI - Recent progress toward the exploitation of organic radical compounds with photo responsive magnetic properties. AB - Recent studies on the exploitation of novel organic radical compounds that exhibit photo-responsive properties by illumination are described in this tutorial review. Those so far reported consist of a variety of photo-responsive pi-electron moieties such as azobenzene, diarylethene, biindenylidene, terphenoquinone, anthracene, naphthopyran, arylimine, or hexaarylbiimidazoles, bearing stable organic radicals or generating organic radicals as spin centres for magnetic properties. The switching behaviours of their magnetic properties evoked by the stimuli of irradiation are discussed by taking the associated structural changes into consideration. PMID- 15280968 TI - Iodine electrophiles in stereoselective reactions: recent developments and synthetic applications. AB - The regio- and stereocontrolled functionalisation of carbon-carbon double bonds bears enormous potential in organic synthesis. This area has been extensively studied and reviewed as alkenes are amongst the most important starting materials for synthetic chemists, accessible in many varieties and in large quantities. We focus in this tutorial review only on recent developments using iodine electrophiles for the functionalisation of alkenes although transition-mediated reactions and functionalisations with chalcogen electrophiles also play an important role. New synthetic applications using this methodology showing scope and limitations of iodine-mediated processes also within the context of other electrophilic reactions are highlighted. PMID- 15280969 TI - Supramolecular chirality of self-assembled systems in solution. AB - Self-assembly plays an important role in the formation of many (chiral) biological structures, such as DNA, alpha-helices or beta-sheets of proteins. This process, which is the main tool of Supramolecular Chemistry (i.e. the chemistry of the molecular assemblies and of the intermolecular bonds), starts to play a significant role in nanotechnology for the construction of functional synthetic structures of nanometer size. The control of chirality in synthetic self-assembled systems is very important for applications of these systems e.g. in molecular recognition or mimicking of the catalytic activity of enzymes. This tutorial review deals with the most representative contributions in the field of supramolecular chirality. Specifically, the discussion centers on several examples that represent the control over chirality for self-assembled systems in solution. PMID- 15280970 TI - Recent developments in the supramolecular chemistry of terpyridine-metal complexes. AB - This critical review describes recent developments in the field of supramolecular chemistry of terpyridine-metal complexes. The synthesis and characteristics of single as well as multiple homo- and heterometallic complexes is discussed. Furthermore, complexes containing fullerenes, biological building blocks, extended aggregates of different architectures as well as rings are presented. A special emphasis is placed upon the properties (e.g. redox properties, luminescence etc.) of functional systems. Potential applications in optical nano devices, molecular storage units, molecular switches and solar cells are discussed. PMID- 15280971 TI - A prospective, randomized trial of endoscopic band ligation versus endoscopic hemoclip placement for bleeding gastric Dieulafoy's lesions. AB - BACKGROUND AND STUDY AIMS: Dieulafoy's lesion is a rare cause of massive gastrointestinal hemorrhage, most commonly in the proximal stomach. Mechanical endoscopic methods have recently become the standard therapeutic approach. However, there have been few studies comparing the efficacy of different mechanical endoscopic methods in treating gastric Dieulafoy's lesions. This study was therefore carried out to compare the hemostatic efficacy and safety of endoscopic band ligation (EBL) and endoscopic hemoclip placement (EHP) in the treatment of bleeding gastric Dieulafoy's lesions. PATIENTS AND METHODS: Between January 2002 and October 2003, 26 consecutive patients with bleeding gastric Dieulafoy's lesions were prospectively enrolled and were randomly assigned to undergo EBL (13 patients) or EHP (13 patients). Demographic characteristics, endoscopic variables, and outcome parameters, including rates of hemostasis and recurrent bleeding, were analyzed. RESULTS: One O-ring was applied in each case in the EBL group, and the median number of hemoclips applied was one (range one to four) in the EHP group. There were no significant differences between the groups with regard to age, sex, presence of shock, initial hemoglobin level, coagulopathy, concurrent diseases, location of the lesion, type of bleeding stigmata, blood transfusion requirements, or hospitalization periods. Primary hemostasis was achieved in all 26 patients. There was one case of recurrent bleeding in each group; secondary hemostasis was achieved with EBL in one of these patients and by endoscopic epinephrine injection in the other. There were no second episodes of recurrent bleeding, no procedure-related complications, no cases in which surgery was needed, and no bleeding-related deaths in either group. CONCLUSIONS: In this small study, no differences were detected in the efficacy or the safety of EBL vs. EHP in the management of bleeding gastric Dieulafoy's lesions. PMID- 15280972 TI - Endoscopic augmentation of the lower esophageal sphincter for the treatment of gastroesophageal reflux disease: multicenter study of the Gatekeeper Reflux Repair System. AB - BACKGROUND AND STUDY AIMS: The safety and effectiveness of the Gatekeeper Reflux Repair System (Medtronic Europe, Tolochenaz, Switzerland) in the treatment of gastroesophageal reflux disease (GERD) was evaluated. This new, reversible treatment modality involves the endoscopic introduction of expandable polyacrylonitrile-based hydrogel prostheses into the esophageal submucosa to augment the lower esophageal sphincter (LES). PATIENTS AND METHODS: For this study, data from two prospective, nonrandomized European multicenter trials were pooled. Sixty-nine GERD patients with heartburn and regurgitation and abnormal esophageal acid exposure (24-h pH < 4.0 for > 4 % of the total time) who had responded to proton-pump inhibitor (PPI) therapy were recruited, and 68 were treated with up to six prostheses placed at the gastroesophageal junction. Patients underwent esophageal manometry, endoscopy, 24-h pH-metry, and symptom scoring at intake and 1, 3, and 6 months after the procedure. RESULTS: A total of 77 procedures were performed in 67 patients, and a total of 270 prostheses were placed (mean 4.3 per procedure). At 1 and 6 months, 80.4 % and 70.4 % of the prostheses were retained, respectively. At 6 months, 24-h pH-metry outcomes with pH < 4.0 for > 4.0 % of the time decreased from 9.1 % to 6.1 % (n = 45; P < 0.05). Median LES pressure increased significantly from 8.8 mmHg at baseline to 13.8 mmHg at 6 months (n = 42, P < 0.01). Median GERD heartburn-related quality of-life scores improved significantly from 24.0 to 5.0 (n = 53, P < 0.01) in patients no longer receiving PPI therapy. Two serious adverse events (3.0 %) occurred. Both patients recovered uneventfully. Prostheses were endoscopically removed from one patient without any adverse events. CONCLUSIONS: The Gatekeeper Reflux Repair System is a safe endoscopic treatment modality that significantly improves GERD symptoms and has objective effects on acid reflux. PMID- 15280974 TI - Use of self-expandable plastic stents for the treatment of esophageal perforations and symptomatic anastomotic leaks. AB - BACKGROUND AND STUDY AIMS: Extensive anastomotic leaks after esophageal resection and esophageal perforations are a therapeutic challenge. The aim of the present study was to assess the potential of the self-expandable Polyflex plastic stent for the treatment of these conditions. PATIENTS AND METHODS: Between January 2002 and March 2003, nine patients were treated with a self-expandable Polyflex plastic stent for sealing of thoracic esophagoenteric anastomotic leaks following surgical resection (n = 5) or esophageal perforation (n = 4). RESULTS: In all patients the stents were inserted successfully without technical problems. In all but two patients complete sealing of the leak was achieved as demonstrated by radiography with water-soluble contrast media. The stent migration rate was 30 % and repositioning of the migrated stents was possible in all cases. Complete mucosal healing of the esophageal leaks and stent extraction was achieved in six patients. The stents were in situ for an average period of 135 +/- 78 days. Two critically ill patients with anastomotic leaks died in spite of stent insertion due to sepsis and one patient with esophageal perforation died due to the underlying malignant disease. CONCLUSIONS: Our preliminary experience with the self-expanding and removable Polyflex plastic stent for the sealing of anastomotic leaks and esophageal perforations suggests that this stent is a feasible treatment option, in particular, for more extensive esophageal defects, patients with co-morbid conditions, and critically ill patients. PMID- 15280973 TI - Short-term and long-term results of endoscopic balloon dilation for achalasia: 12 years' experience. AB - BACKGROUND AND STUDY AIMS: This retrospective study reports 12 years' experience with pneumatic dilation treatment in patients with achalasia and attempts to define factors capable of predicting failure of endoscopic dilation. PATIENTS AND METHODS: Consecutive patients with achalasia who received endoscopic balloon dilation were studied retrospectively. Repeat dilation was carried out if dysphagia persisted or recurred. A structured symptom score questionnaire (the Eckardt score) was conducted by phone with patients who had received dilation and had been followed up for more than 2 years. Failure was defined as the presence of significant dysphagic symptoms after more than two repeat dilations. Data for the first 2 years (short-term) and for the subsequent follow-up (long-term) were analyzed. RESULTS: From 1989 to 2001, 66 patients underwent endoscopic balloon dilation for achalasia; three perforations (4.5 %) occurred, with no mortalities. Dysphagic symptoms significantly improved 12 weeks after the procedure ( P < 0.05). Fourteen patients (20 %) required a second dilation procedure within a median of 7 months (range 1 - 52 months), and 13 of them underwent repeat dilations within the first 2 years. Five patients (7.5 %) required further surgical or endoscopic therapy. Fifty-eight patients received pneumatic dilation for more than 2 years; 32 (55 %) responded to the questionnaire. The mean dysphagia score was 1.7 (SD 1.2), with only five patients (16 %) having significant dysphagic symptoms during a median follow-up period of 55 months (range 26 - 130 months). The cumulative success rates for pneumatic dilation after 5 and 19 years were 74 % and 62 %, respectively. Cox regression analysis identified small balloon size (30 mm) as the only significant factor capable of predicting failure of endoscopic dilation ( P = 0.009; relative risk 5.3; 95 % confidence interval, 1.7 to 40.9). CONCLUSIONS: Endoscopic balloon dilation is an effective treatment for achalasia, with minimal morbidity (60 % experience long term benefit). PMID- 15280975 TI - Persistence of gastrocutaneous fistula after removal of gastrostomy tubes in children: prevalence and associated factors. AB - BACKGROUND AND STUDY AIMS: The aims of this study were to determine the prevalence of gastrocutaneous fistula after removal of gastrostomy tubes in children and to identify associated risk factors. PATIENTS AND METHODS: The records of children who had undergone removal of gastrostomy tubes between January 1992 and December 2002 were reviewed retrospectively. Persistent gastrocutaneous fistula was defined as the absence of closure of the gastrostomy 1 month after tube removal. Factors that might influence spontaneous closure of the gastrostomy were studied, including age, underlying disease, nutritional status, type of gastrostomy, replacement of the gastrostomy tube by a button, abdominal wall thickness, duration of gastrostomy tube or button placement, and complications related to the presence of the gastrostomy (infection, granulation tissue). RESULTS: A total of 44 patients were included in the study (mean age 20 months, range 1 day to 14 years). Of these, 28 had undergone percutaneous endoscopic gastrostomy and 16 surgical gastrostomy. The mean time to spontaneous closure was 6 +/- 7 days. Persistent gastrocutaneous fistula developed in 11 patients (25 %) and in seven of these patients this required surgical closure (16 %). The mean duration of gastrostomy placement was significantly longer in patients who went on to develop a gastrocutaneous fistula than in patients who did not develop a fistula (39 +/- 19 months vs. 22 +/- 23 months, respectively, P < 0.03). No other significant association was found between the time required for spontaneous closure and the characteristics of patients or the type of gastrostomy. CONCLUSIONS: Persistent gastrocutaneous fistula is common after removal of gastrostomy tubes in children. Surgical closure should be considered when a gastrostomy has not closed spontaneously 1 month after removal of the gastrostomy tube. PMID- 15280976 TI - Accurate diagnosis of pancreas divisum by linear-array endoscopic ultrasonography. AB - BACKGROUND AND STUDY AIMS: During linear-array endoscopic ultrasonography (EUS), the main pancreatic duct can be followed continuously from the major papilla into the pancreatic body in most patients. Often, the duct can also be seen crossing a sonographic border between the ventral and dorsal pancreatic anlagen. It was hypothesized that the presence of either feature excludes pancreas divisum, whereas the absence of these features suggests complete pancreas divisum. PATIENTS AND METHODS: Pancreas divisum was sought during all linear-array EUS examinations conducted between July 1999 and June 2003. Charts were reviewed retrospectively, and patients who underwent endoscopic retrograde pancreatography after, but not before, EUS were included in the study. RESULTS: A total of 162 patients had EUS before ERCP. Adequate evaluation of the pancreatic duct was possible in 78 % of the patients. The prevalence of pancreas divisum was 13.6 %. In patients with adequate duct visualization, the sensitivity, specificity, and positive and negative predictive values for EUS were 95 %, 97 %, 86 %, and 99 %, respectively. The overall accuracy of EUS for identifying pancreas divisum was 97 % in this subgroup. CONCLUSION: Adequate EUS evaluation of pancreas divisum was possible in most cases. Linear-array EUS is a promising diagnostic test for pancreas divisum. PMID- 15280977 TI - Use of the curved linear-array echo endoscope to identify gastrorenal shunts in patients with gastric fundal varices. AB - BACKGROUND AND STUDY AIMS: : Balloon-occluded retrograde transvenous obliteration (B-RTO) has emerged as an effective, minimally invasive treatment for fundal varices. B-RTO requires a spontaneously developed gastrorenal shunt as a pathway for the balloon catheter to reach the fundal varices. We used a curved linear array (CLA) echo endoscope in patients with fundal varices to identify gastrorenal shunts, and compared the detection rate with the gold standard, contrast-enhanced computed tomography (CECT). PATIENTS AND METHODS: A total of 40 patients with fundal varices were examined with both CLA echo endoscopy and CECT. The CECT images were retrospectively and independently evaluated by two gastroenterologists who were unaware of the clinical details, including the results of the CLA echo endoscopy. RESULTS: CLA echo endoscopy identified gastrorenal shunts in 26/40 patients with fundal varices. It visualized the shunt in a longitudinal direction and provided images of the connections of the shunt at both ends, the fundal varices and the left renal vein/branch of the inferior adrenal vein. The kappa index for CLA echo endoscopy and CECT for the identification of gastrorenal shunt was 0.9 (95 % CI, 0.6 to 1.0). When the cutoff point for the diameter of the gastrorenal shunt detected by the CLA echo endoscope was set at equal to or greater than 5 mm, the kappa index was 1.0 (95 % CI, 0.7 to 1.0). CONCLUSIONS: These results suggest that CLA echo endoscopy can successfully identify gastrorenal shunt and provide detailed morphological information. It also efficiently identifies patients suitable for B-RTO, particularly in cases of acute bleeding. It also has considerable potential for providing detailed information with regard to the treatment of gastric varices. PMID- 15280978 TI - Yield of endoscopic ultrasound-guided fine-needle aspiration of bile duct lesions. AB - BACKGROUND AND STUDY AIMS: It is still difficult to differentiate reliably between benign and malignant biliary tract lesions. Endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) has added to the diagnostic power of EUS for other gastrointestinal tumors. A retrospective analysis of experience with FNA sampling of bile duct lesions was therefore carried out. PATIENTS AND METHODS: All EUS-FNA procedures for bile duct masses or strictures were analyzed at our tertiary referral center from May 2000 through October 2002. Data for EUS findings, the results of EUS-FNA, and tissue sampling at surgery were included. EUS-FNA procedures were carried out using a 22-gauge needle. An experienced cytopathologist was present during FNA in all but three cases. Clinical follow-up details were recorded when available for patients in whom a suitable diagnostic gold standard was not available for comparison. RESULTS: A total of 35 patients underwent EUS-FNA of bile duct lesions during the study period. There were no complications. Data for EUS-FNA of bile duct masses or strictures and tissue obtained at surgery were available for 23 patients. If positive cytology at surgical pathology is taken as the gold standard, EUS-FNA has a diagnostic yield for cancer of 100 % (if atypia/inconclusive findings in the FNA sample are regarded as benign). Eleven patients had a definite malignancy on surgical pathology. Of these 11 patients, five had a finding of malignancy on EUS-FNA, giving a sensitivity of 45 % (if FNA cytology reported as atypia/inconclusive is regarded as benign). Twelve patients had findings of no malignancy from tissue obtained at surgery. Of these 12 patients, nine had benign pathology and three had atypia/inconclusive findings in the EUS-FNA sample (specificity of 100 % if atypia/inconclusive findings are considered benign). A further 12 patients did not have surgical specimens for comparison with EUS-FNA results. Four patients had definite findings of malignancy on EUS-FNA alone, and one patient had FNA findings suspicious for malignancy. Seven patients had negative or equivocal EUS FNA results. These 12 patients are described but excluded from further analysis, as a gold standard was not available for comparison. However, clinical follow-up data were available for eight of these 12 patients, and in each case the follow up findings were compatible with previous benign or malignant EUS-FNA findings. CONCLUSIONS: The practice of EUS-FNA has improved the diagnostic yield of EUS. These results suggest that it is a safe and useful procedure for investigating biliary masses or strictures that have hitherto caused considerable diagnostic confusion, especially in patients with negative brush cytology findings. The possibility of false-negative findings remains, but core biopsy needles may improve the situation. The results of further studies are awaited. PMID- 15280979 TI - Intravenous hyoscine as a premedication for colonoscopy: a randomized double blind controlled trial. AB - BACKGROUND AND STUDY AIMS: There is wide variation in the use of antispasmodics to facilitate colonoscopy, both within and between countries, and its use before such procedures remains controversial. The aim of this study was to determine whether there was any objective benefit in using hyoscine as a premedication for colonoscopy in a district general hospital. PATIENTS AND METHODS: Consecutive day case patients undergoing colonoscopy were included in the study. They were prospectively randomly allocated to receive either intravenous hyoscine (n = 61) or intravenous placebo (n = 56) as part of their premedication. RESULTS: Our analysis demonstrated no statistically significant difference between the two groups with respect to the median time from colonoscope insertion to caecal intubation (9.7 minutes in the hyoscine group vs. 8.3 minutes in the placebo group) or the median total procedure time (14.8 minutes in the hyoscine group vs. 13.8 minutes in the placebo group). There was also no statistically significant difference in success rates for caecal intubation between the two groups ( P < 0.06). However a type II error cannot be excluded because of the small sample size. CONCLUSION: This study demonstrated no obvious benefit in the routine use of hyoscine as a premedication for colonoscopy in a district general hospital setting. PMID- 15280980 TI - Eyes wide shut. PMID- 15280981 TI - EUS Meets Voxel-Man: three-dimensional anatomic animation of linear-array endoscopic ultrasound images. AB - Endoscopic ultrasonography (EUS) is a widely used imaging modality in gastroenterology. The development of linear-array endoscopic ultrasound transducers, with facilities for EUS-guided diagnostic and therapeutic procedures, led to increasingly widespread use in different areas of the body. Examiners need to have excellent knowledge of anatomy. Orientation in linear EUS is more difficult and the learning curve is long. In an effort to shorten the training, reducing the risk to the patient and to allow a faster learning of the basic anatomic structures EUS meets VOXEL-MAN, an interactive three-dimensional anatomic simulation program has been developed for linear EUS for the purposes of private and independent study. PMID- 15280982 TI - Oesophagorespiratory fistulas as a complication of self-expanding metal oesophageal stents. AB - Self-expanding metal oesophageal stents are considered by many to be the ideal palliative treatment for malignant dysphagia, but there have been reports of life threatening complications associated with their use. We report three cases in which oesophagorespiratory fistulas were caused by the use of these stents. PMID- 15280983 TI - Endoscopic treatment of a mid-esophageal diverticulum. AB - Mid-esophageal diverticulum is a medical rarity. Only patients in whom the condition is symptomatic should receive treatment. Minimally invasive surgery via a thoracoscopic approach is currently the preferred treatment for the condition. This case report describes a 63-year-old woman with a symptomatic mid-esophageal diverticulum, which was successfully treated by endoscopic diverticulotomy using a needle-knife through a flexible endoscope. PMID- 15280984 TI - External pancreatic fistulas resistant to conventional endoscopic therapy: endoscopic closure with N-butyl-2-cyanoacrylate (Glubran 2). AB - External pancreatic fistulas may follow abdominal surgery or injury. While most respond to conservative management or endoscopic intervention, others might require surgery for complete healing. We report four cases of patients with external pancreatic fistulas that failed to respond to conservative management and drainage. N-butyl-2-cyanoacrylate surgical glue (Glubran 2) was directly injected into the fistulous tract. The fistulas closed within 24 hours of the Glubran 2 injection in three cases (75 %). In patients with external pancreatic fistulas that fail to respond to conservative and endoscopic drainage, injection of Glubran 2 directly into the fistulous tract may lead to closure, thus avoiding the need for surgical intervention. PMID- 15280986 TI - Biliary cannulation and pancreatic guide-wire placement. PMID- 15280987 TI - Botulinum toxin injection after biliary sphincterotomy. PMID- 15280989 TI - Esophageal dislodgment of a reused variceal ligator cap: a complication of endoscopic variceal ligation. PMID- 15280990 TI - Endoscopy-assisted capsule endoscopy in patients with swallowing disorders. PMID- 15280992 TI - Magnification and chromoscopy with the acetic acid test. PMID- 15280994 TI - Large gastric perforation after endoscopic mucosal resection treated by application of metallic clips (video). PMID- 15280995 TI - A reliable method of single-pass endoscopic gastrostomy replacement. PMID- 15280996 TI - Use of a Soehendra stent retriever to treat a pancreatic pseudocyst with EUS guided cystogastrostomy. PMID- 15280997 TI - An alternative method of exchanging an occluded percutaneous transhepatic biliary prosthesis (Yamakawa type). PMID- 15280998 TI - Cecal diverticulitis: an unusual endoscopic finding. PMID- 15280999 TI - Failed colonoscopy due to hernia. PMID- 15281000 TI - Low temperature blocks the stimulatory effect of human chorionic gonadotropin on steroidogenic acute regulatory protein mRNA and testosterone production but not cyclic adenosine monophosphate in mouse Leydig tumor cells. AB - Low temperatures slow down metabolism, partly because the kinetic energy of molecules is reduced and enzymes may be structurally impaired. We now report that relative to its maximal activity at 37 degrees C, adenylate cyclase (AC) still retained 25% functionality (determined as cyclic adenosine monophosphate [cAMP] production) at 4 degrees C in mouse Leydig tumor cells (MLTC-1) in response to 50 IU/L human chorionic gonadotropin (hCG), whereas steroidogenic acute regulatory (StAR) protein mRNA and testosterone production were completely impaired. The incubation of MLTC-1 with the phosphodiesterase inhibitor (3-isobutyl-1 methylxanthine; IBMX) resulted in significantly increased intracellular cAMP concentration at all 3 temperatures, but this had no impact on testosterone production. AC, cAMP, and phosphodiesterase form an important intracellular second-messenger mechanism in many organisms, some that inhabit very low temperature niches. The cold-resistance of AC and phosphodiesterase may thus have evolved to cope with adverse conditions. Although hibernation may lead to decreased steroid hormone production, it is also likely that cold-mediated decreased steroid hormone production induces hibernation. PMID- 15281001 TI - Triple genetic variation in the HNF-4alpha gene is associated with early-onset type 2 diabetes mellitus in a philippino family. AB - Maturity-onset diabetes of the young-type 1 (MODY1) is a form of monogenic type 2 diabetes mellitus (T2DM) with long-term complications due to mutations in the HNF 4alpha gene. The HNF-4alpha gene is involved in hepatic differentiation and expression of genes regulating glucose transport, glycolysis, and lipid metabolism. The abnormal glucose-stimulated insulin secretion in MODY1 subjects may be due to reduced glucose transport and glycolysis. To date, 14 mutations in the HNF-4alpha gene have been identified as a cause of either MODY1 or late-onset type 2 diabetes. So far, no screening has been performed in subjects from the Philippines. We recruited a Philippino family with autosomal dominant early-onset type 2 diabetes and screened the proband for mutations in the genes for HNF 1alpha, GCK, HNF-4alpha, IPF-1, HNF-6, and NGN3. We identified a new missense mutation in exon 5 (V199I) of the HNF-4alpha gene and 2 new single-nucleotide substitutions in intron 4, IVS4-nt4 (G --> A) and IVS4-nt20 (C --> T), all cosegregating with diabetes in the 3 affected available siblings. These variations were not present in 100 normal healthy subjects. Bioinformatic analysis suggests that these variations in the whole, and overall the IVS4-nt4 variation located at splicing site, may affect the splicing potential of intron 4. We have biochemically and clinically characterized the Philippine-1 family. We suggest that the V199I missense mutation located in the ligand binding/dimerization domain of HNF-4alpha contributes to type 2 diabetes in the Philippine-1 family. The intron variations may contribute susceptibility to diabetes. PMID- 15281002 TI - Liver dysfunction induced by bile duct ligation and galactosamine injection alters cardiac protein synthesis. AB - Liver disease has been shown to affect the cardiovascular system and may influence cardiac protein metabolism. This hypothesis was tested by measuring rates of cardiac protein synthesis in 2 models of liver disease in rats. The study consisted of 5 groups--group 1: control, injected with saline and fed ad libitum; group 2: acute liver injury, by dosage with 400 mg/kg galactosamine; group 3: injected with saline and pair-fed to group 2; group 4: chronic liver disease, using bile duct ligation; and group 5: sham-operated and pair-fed to group 4. Rates of cardiac protein synthesis were measured using the flooding dose technique. After 1 week, galactosamine injection caused the following cardiac changes, i.e. group (2) versus (3): an increased RNA content, RNA/DNA ratio, and RNA/protein ratio. However, there was no change in DNA or protein content, or protein/DNA ratio. There was an increase in the fractional rate of protein synthesis, and the absolute synthesis rate. Cellular efficiency was increased, but RNA activity remained unchanged. Comparison of groups 4 and 5 showed that bile duct ligation caused no change in any parameters measured. Although comparison of the ad libitum-fed group 1 with the bile duct ligation group 4 showed reduced cardiac weight, protein, and RNA content, with decreased right ventricular absolute synthesis rates; this was also seen in the pair-fed group 5, suggesting that these effects were due solely to reduced oral intake. Thus, although galactosamine-induced acute liver injury caused marked changes in cardiac biochemistry, bile duct ligation per se did not. This study also illustrates the importance of including a pair-fed group. PMID- 15281003 TI - In vivo glycated low-density lipoprotein is not more susceptible to oxidation than nonglycated low-density lipoprotein in type 1 diabetes. AB - It has been suggested that low-density lipoprotein (LDL) modified by glycation may be more susceptible to oxidation and thus, enhance its atherogenicity. Using affinity chromatography, LDL glycated in vivo (G-LDL) and relatively nonglycated. (N-LDL) subfractions can be isolated from the same individual. The extent of and susceptibility to oxidation of N-LDL compared with G-LDL was determined in 15 type 1 diabetic patients. Total LDL was isolated and separated by boronate affinity chromatography into relatively glycated (G-) and nonglycated (N-) subfractions. The extent of glycation, glycoxidation, and lipoxidation, lipid soluble antioxidant content, susceptibility to in vitro oxidation, and nuclear magnetic resonance (NMR)-determined particle size and subclass distribution were determined for each subfraction. Glycation, (fructose-lysine) was higher in G-LDL versus N-LDL, (0.28 +/- 0.08 v 0.13 +/- 0.04 mmol/mol lysine, P < .0001). However, levels of glycoxidation/lipoxidation products and of antioxidants were similar or lower in G-LDL compared with N-LDL and were inversely correlated with fructose-lysine (FL) concentrations in G-LDL, but positively correlated in N-LDL. In vitro LDL (CuCl2) oxidation demonstrated a longer lag time for oxidation of G LDL than N-LDL (50 +/- 0.16 v 37 +/- 0.15 min, P < .01), but there was no difference in the rate or extent of lipid oxidation, nor in any aspect of protein oxidation. Mean LDL particle size and subclass distribution did not differ between G-LDL and N-LDL. Thus, G-LDL from well-controlled type 1 diabetic patients is not more modified by oxidation, more susceptible to oxidation, or smaller than relatively N-LDL, suggesting alternative factors may contribute to the atherogenicity of LDL from type 1 diabetic patients. PMID- 15281004 TI - Effect of a novel palatinose-based liquid balanced formula (MHN-01) on glucose and lipid metabolism in male Sprague-Dawley rats after short- and long-term ingestion. AB - Postprandial hyperglycemia and hyperinsulinemia are often present in obese subjects with glucose intolerance in whom insufficient early phase insulin secretion and subsequent delayed hyperinsulin response are observed. To address this problem, a novel palatinose-based enteral formula designated as MHN-01 was developed for the prevention of postprandial hyperglycemia and hyperinsulinemia. The effects of MHN-01 on carbohydrate and lipid metabolism in Sprague-Dawley (SD) rats were compared with those of the standard balanced formula (SBF). After a bolus intragastric injection of each formula equivalent to 0.9 g/kg carbohydrate, the peak levels of plasma glucose (PG) and insulin (IRI) in peripheral and portal veins of the MHN-01 group were significantly lower than those of the SBF group. The areas under the curve of PG and IRI in the MHN-01 group were 58.0% and 43.1% of those in the SBF group in the femoral vein and 65.0% and 69.3% in the portal vein, respectively. In the 2-month study, serum levels of IRI and triglyceride in peripheral blood in the MHN-01 group decreased and those in the SBF group increased compared with initial levels. Consequently, both levels in the MHN-01 group were significantly lower than those in the SBF group. In addition, the amount of accumulated fat in abdominal adipose tissue and liver tissue of the MHN 01 group was markedly reduced in comparison to that of the SBF group. Insulin sensitivity, evaluated as glucose infusion rate using the hyperinsulinemic euglycemic clamp technique, in the MHN-01 group was higher than that in the SBF group. Thus, in comparison to SBF, MHN-01 suppressed postprandial hyperglycemia and hyperinsulinemia, reduced visceral fat accumulation, and improved insulin sensitivity. Therefore, human study on the effects of MHN-01 on carbohydrate and lipid metabolism will be recommended to confirm whether MHN-01 may be a useful functional food for the treatment and prevention of insulin resistance. PMID- 15281005 TI - Body fat is the main predictor of fibrinogen levels in healthy non-obese men. AB - Previous studies have demonstrated that circulating levels of C-reactive protein (CRP), a marker of cardiovascular risk, are strictly related to body fatness. Elevated fibrinogen levels are also predictive of future cardiovascular events. The metabolic background of this relationship and the predictors of fibrinogen levels have not been well established. We aimed to evaluate whether fibrinogen levels are associated with body fat content and distribution and to determine the independent predictors of fibrinogen levels in a sample of healthy, non-obese, nonsmoking young adult men. Age, anthropometric measures (body mass index [BMI], waist-to-hip ratio [WHR]), total and regional fat content (determined by dual x ray absorptiometry [DXA]), metabolic variables (total cholesterol [T-Chol], low density lipoprotein cholesterol [LDL-C], and high-density lipoprotein cholesterol [HDL-C]; triglycerides [TG]; glucose and insulin levels; fasting insulin resistance index [FIRI]; blood pressure), interleukin-6 (IL-6), and acute-phase reactants levels (fibrinogen, highly sensitive [hs]-CRP) were determined in 87 healthy nonsmoking, non-obese subjects. Linear regression analysis was used to evaluate the association between body fat, fibrinogen, and metabolic variables, and multiple regression model analysis was used to examine the independent predictors of fibrinogen levels. Eighty-seven (30.5 +/- 3.5 years) non-obese (mean BMI 24.1 +/- 3.5) men were studied. Fibrinogen levels were strongly associated with measures of body fat and with metabolic variables. Total body fat (P < .0001) and LDL-cholesterol (P < .01) were the independent predictors of fibrinogen levels, accounting for 29.5% and 10.9% of its variance, respectively. Total body fat was the best independent predictor of hs-CRP levels, accounting for 32.5 % of its variance. We conclude that in healthy, non-obese subjects, body fat content is the main predictor of fibrinogen levels, as well of hs-CRP levels. These findings support the speculation that there is a direct mechanism by which adipose tissue might regulate the levels of circulating acute-phase reactants. PMID- 15281006 TI - Circulating leptin concentrations can be used as a surrogate marker of fat mass in acute spinal cord injury patients. AB - To determine the acute effect of neurological lesion on body composition, plasma leptin level, and the lipid profile, 7 male patients with acute and complete spinal cord injury (SCI) and 9 able-bodied (AB) males were investigated. At 16, 24, 36, and 48 weeks after injury, plasma leptin level and the lipid profile were analyzed, while whole body (WB) and regional fat mass (FM) and fat-free soft tissue (FFST) were measured by dual-energy x-ray absorptiometry (DXA). At all stages, despite no difference being found between both groups for body mass index (BMI), SCI patients had higher FM at WB (P < .01), lower (P < .01), and upper limbs (P < .05), while FFST was lower at WB (P < .05) and lower limbs (P < .01). The leptin level increased gradually from week 24 and was higher at weeks 16, 36, and 48 in SCI patients than in AB patients (7.0 +/- 3.9; 9.7 +/- 5.1; 10.6 +/- 5.3, respectively, v 3.5 +/- 2.5 ng. mL(-1)). SCI patients had lower high-density lipoprotein-cholesterol (HDL-C) (P < .05) and apolipoprotein (apo) A1 (P < .01), while no difference was found for total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), or ApoB levels. At all stages, leptin was strongly and positively correlated with WB and regional FM % (r > 0.75; P < .05) and with TC, LDL-C, and ApoB levels (r > 0.65; P < .05). Leptin was negatively correlated with FFST and the ApoA1/ApoB ratio (r > -0.75; P < .05). In conclusion, neurological lesion induced an early and acute alteration in body composition and lipid profile. The strong relationship between serum leptin and FM suggests that this hormone can be used as a surrogate marker of FM in acute SCI patients and thus would serve as a good indicator for cardiovascular disease risk. PMID- 15281007 TI - Type 2 diabetes and the genetics of signal transduction: a study of interaction between adenosine deaminase and acid phosphatase locus 1 polymorphisms. AB - Acid phosphatase locus 1 (ACP1) is a highly polymorphic enzyme that has an important role in flavoenzyme activity and in the control of insulin receptor activity and band 3 protein phosphorylation status. Adenosine deaminase (ADA) is a polymorphic enzyme that catalyses the irreversible deamination of adenosine to inosine and has an important role in regulating adenosine concentration. Based on the hypothesis that ACP1 counteracts insulin signaling by dephosphorylating the insulin receptor and that adenosine has an anti-insulin action, we reasoned that low ACP1 activity (low dephosphorylating action on insulin receptor) when associated with high ADA activity (low adenosine concentration) would result in a cumulative effect towards an increased glucose tolerance. On the contrary, high ACP1 activity when associated with low ADA activity would result in a cumulative effect towards a decreased glucose tolerance. A total of 280 adult subjects with type 2 diabetes from the population of Penne (Italy) were studied. There was a nonsignificant trend toward an increase in the proportion of subjects with the complex type with high ACP1 activity and low ADA activity (ie, *B/*B; *A/*C; *B/*C; *C/*C//ADA*1/*2 and *2/*2) in type 2 diabetes relative to that observed in newborn infants from the same population. High ACP1 activity/low ADA activity joint genotype was positively associated with high glycemic levels and with high body mass index (BMI) values. Low ACP1 activity/high ADA activity joint genotype was also positively associated with dyslipidemia. These findings suggest that both ACP1 and ADA contribute to the clinical manifestations of type 2 diabetes and probably also have a marginal influence on susceptibility to the disease. Both additive and epistatic interactions between the 2 systems seem to be operative. PMID- 15281008 TI - The effect of l-carnitine on fat oxidation, protein turnover, and body composition in slightly overweight subjects. AB - We used a combined tracer technique with the stable isotopes 13C and 15N to gain further insight into the metabolic changes that accompany supplementation of L carnitine. The aim of the present study was to investigate whether L-carnitine supplementation can influence fat oxidation, protein turnover, body composition, and weight development in slightly overweight subjects. Twelve volunteers received an individual regular diet either without or with L-carnitine supplementation of 3 g/d for 10 days. Protein turnover and fat oxidation were investigated after administration of [15N]glycine and an [U-13C]algae lipid mixture. The 15N- and 13C-enrichment in urine and breath were measured by isotope ratio mass spectrometry. Body fat mass (BFM), total body water (TBW), and lean body mass (LBM) were calculated by using bioelectric impedance analysis. L carnitine supplementation led to a significant increase in 13C-fat oxidation (15.8% v 19.3%; P = .021) whereas protein synthesis and breakdown rates (3.7 and 3.4 g/kg/d, respectively) remained unchanged, indicating that the increased dietary fat oxidation in slightly overweight subjects was not accompanied by protein catabolism. PMID- 15281009 TI - The effect of cellular retinoic acid binding protein-I expression on the CYP26 mediated catabolism of all-trans retinoic acid and cell proliferation in head and neck squamous cell carcinoma. AB - The aim of this study was to confirm if catabolism of all-trans retinoic acid (RA) is enhanced by type I cellular retinoic acid binding protein (CRABP-I) expression and to investigate the effect of this enhanced catabolism on cell proliferation of the head and neck squamous cell carcinoma (HNSCC) cell line, AMC HN-7. We also analyzed the effects of CRABP-I on RA-induced retinoic acid receptor (RAR) activity. The expression of the CRABP-I in stably transfected AMC HN-7 cell lines (HN7-BPIa and HN7-BPIb) resulted in a lower sensitivity to administered RA compared with that of controls in a clonogenic assay. HN7-BPIs cells showed an increased amount of polar metabolites of RA in thin-layer chromatography. The transcriptional activity of the reporter plasmid RARE(DR5)-tk CAT after the treatment of RA was lesser in HN7-BPIs than in controls. These results suggest that the increased CYP26-mediated catabolism of RA by CRABP-I transfection might decrease the amount of RA that is accessible to the nuclear receptors and make HNSCC cells resistant to RA. PMID- 15281010 TI - Utility of non-high-density lipoprotein cholesterol in hemodialyzed patients. AB - Non-high-density lipoprotein-cholesterol (HDL-C) is proposed as a strong predictor of cardiovascular disease (CVD). Measuring non-HDL-C, as total cholesterol minus HDL-C, is convenient for routine practice because, among other advantages, fasting is not required. There are limited data of non-HDL-C in end stage renal disease patients. We applied non-HDL-C calculation to 50 chronic renal patients receiving maintenance hemodialysis (HD) and 20 healthy subjects, apart from measurement of low-density lipoprotein (LDL), very-low-density lipoprotein (VLDL) HDL, intermediate-density lipoprotein-cholesterol (IDL-C), apoprotein (apo) B, and triglycerides. HD patients presented higher plasma triglycerides and IDL-C and lower HDL-C than the control group, even after adjustment for age (P < .05). VLDL-C increased in HD patients (P < .001) while differences in non-HDL-C did not reach significance (P = .08). To detect which parameter constitutes a better marker of CVD risk among HD patients, a receiver operating characteristic (ROC) analysis was performed considering HD patients in the highest risk group for CVD. In the ROC graphic, the plots of VLDL and IDL-C exhibited the greater observed accuracy and the best performance, while non-HDL-C showed a curve close to the 45 degrees line indicating that this parameter is a poor discriminator for evaluating CVD risk among HD patients. Non-HDL-C calculation, expressing all apo B-containing lipoproteins, may miss the significant contribution of each atherogenic lipoprotein, such as increase in IDL. This observation would not be in agreement with the currently proposed application of non-HDL-C a useful tool for risk assessment among HD patients. PMID- 15281011 TI - Homocysteine and disability in hospitalized geriatric patients. AB - Elevated total homocysteine (tHcy) concentrations have been found to be associated with cardiovascular disease and dementia in old age. The present study was performed to identify the prevalence of hyperhomocysteinemia (HHcy) and to analyze the association between tHcy concentration and sociodemographic characteristics, nutritional parameters, and cognitive and functional status in this sample of hospitalized geriatric patients. A total of 214 patients (77% females) 65+ years old admitted into an acute care geriatric ward of an internal medical department in the Northern Italy were studied. tHcy concentration was measured using a high-performance liquid chromatography with fluorescence detection (HPLC-F). Information about nutrition (body mass index [BMI], serum albumin, cholesterol, and transferrin) was collected on admission. Functional status was investigated with the Basic Activities of Daily Living scale (ADL) and the Instrumental Activities of Daily Living scale (IADL); cognitive and affective status were assessed by the Mini-Mental State Evaluation (MMSE) and the Geriatric Depression Scale (GDS). The mean tHcy concentration was 18.4 +/- 13.1 micromol/L; 74.2% of males and 68.9% of females had HHcy (> 12 micromol/L). Sixty-four percent of patients with normal serum vitamin B12 and folate concentrations had HHcy. Elevated tHcy concentrations were associated with older age, male gender, increasing serum creatinine, lower MMSE score, and disability. The mean tHcy concentration depended on the occurrence of different diseases. Patients affected by atherosclerotic diseases, such as ischemic heart diseases, cerebrovascular diseases, and dementia had higher mean tHcy concentration than those without diagnosed vascular diseases. In multivariate analysis, vitamin B12, folate, serum albumin, creatinine, and disability emerged as factors associated with tHcy, adjusted for age, gender, education, MMSE score, and atherosclerotic diseases. Our results suggest that the prevalence of HHcy in hospitalized patients is very high, even in subjects with normal cobalamin and folate concentrations. High Hcy concentration can be associated with functional impairment. PMID- 15281012 TI - Effect of exercise on postprandial lipemia following a higher calorie meal in Yucatan miniature swine. AB - Exercise has been shown to attenuate the postprandial lipemic (PPL) response to a modest kcal high-fat meal in numerous human studies, but has not been fully examined in swine. In addition, the effects of exercise on a high-fat meal of larger magnitude have not been examined in humans or in swine. Thus, the purpose of this study was to examine the PPL response to a single, high-fat/cholesterol (HFC) meal (approximately 3,000 kcal, 1,300 kcal from fat) and determine if exercise attenuates the PPL response. Sedentary, female Yucatan miniature swine (n = 10) completed 3 PPL trials: (1) pre diet (PRE); (2) post HFC diet (POST); and (3) post HFC diet plus exercise (EX, 45 minutes at 75% heart rate maximum). Blood samples were collected before (0 hour) and at 2, 4, 6, and 8 hours after the single HFC meal for PPL analysis. Postheparin lipoprotein lipase (LPL) activity was assessed at 8 hours. While fasting LPL activity was significantly increased with the HFC diet, the PPL response to the HFC meal did not differ depending on diet. Furthermore, the PPL response was not significantly altered with a single session of exercise, perhaps because of the severity of the HFC meal, the sedentary nature of the swine, or because LPL activity was not elevated after exercise. These findings suggest that administration of a HFC meal of this magnitude (approximately 3,000 kcal, 1,300 kcal from fat) will promote significant elevations in postprandial triglyceride (TG) concentrations, overwhelm the adaptive response to a HFC diet (elevated LPL activity), and attenuate the beneficial effects of a single exercise session on this system. PMID- 15281013 TI - Swim training increases glucose output from liver perfused in situ with glucagon in fed and fasted rats. AB - The effect of endurance swim training (3 hours per day, 5 days/week, for 10 weeks) on hepatic glucose production (HGP) in liver perfused in situ for 60 minutes with glucagon and insulin was studied in Sprague-Dawley rats. The experiments were performed in fed rats and in rats fasted for 24 hours, but with lactate (8 mmol/L) added to the perfusion medium. Liver glycogen content was significantly lower in fasted than fed rats (fasted untrained and trained: 14 +/- 4 and 11 +/- 3 micromol glycosyl U/g of liver wet weight (WW); fed untrained and trained: 205 +/- 11 and 231 +/- 11 micromol glycosyl U/g of liver WW; not significantly different in trained and untrained rats). Glucagon increased HGP in the 4 experimental groups, but the increases were more rapid and pronounced in trained than in untrained rats in both fed and fasted states. HGP values (area under the curve [AUC] in micromol/g of liver WW) were significantly higher in trained fed (112.1 +/- 7.1 v 85.9 +/- 12.2 in untrained rats) than in trained fasted rats (50.8 +/- 4.4 v 34.7 +/- 3.6 in untrained rats). When compared with untrained rats, the total amount of glucose released by the liver in response to glucagon in trained rats was approximately 30% higher in the fed state and approximately 45% larger in the fasted state. These results indicate that endurance training increases the response of both glycogenolysis and gluconeogenesis to glucagon. PMID- 15281014 TI - Fiber type- and fatty acid composition-dependent effects of high-fat diets on rat muscle triacylglyceride and fatty acid transporter protein-1 content. AB - Intramuscular triacylglyceride (TAG) is considered an independent marker of insulin resistance in humans. Here, we examined the effect of high-fat diets, based on distinct fatty acid compositions (saturated, monounsaturated or n-6 polyunsaturated), on TAG levels and fatty acid transporter protein (FATP-1) expression in 2 rat muscles that differ in their fiber type, soleus, and gastrocnemius; the relationship to whole body glucose intolerance was also studied. Compared with carbohydrate-fed rats, the groups subjected to any one of the high-fat diets consistently exhibited enhanced body weight gain and adiposity, elevated plasma free fatty acids and TAG in the fed condition, hyperinsulinemia, and glucose intolerance. TAG content was consistently higher in soleus than in gastrocnemius, but was only significantly elevated by the n-6 polyunsaturated-based diet. FATP-1 levels in soleus were double those in gastrocnemius muscle in carbohydrate-fed animals. High-fat diets caused an elevation in FATP-1 protein content in soleus, but a reduction in gastrocnemius. In conclusion, the hyperinsulinemic hyperlipidemic condition upregulates FATP-1 expression in soleus and downregulates that of gastrocnemius. Hypercaloric saturated, monounsaturated, or n-6 polyunsaturated lipid diets cause equivalent whole body insulin resistance in rats, but only an n-6 polyunsaturated acid-based diet triggers intramuscular TAG accumulation. PMID- 15281015 TI - Reduced action of insulin glargine on protein and lipid metabolism: possible relationship to cellular hormone metabolism. AB - Insulin analogues are used in the treatment of diabetes to mimic physiological insulin secretion. Glargine is used to provide basal insulin levels. Previous work has shown no differences in glucose uptake when glargine was compared to native insulin. The action of insulin on protein and lipid metabolism is studied infrequently, but these important actions should be considered with insulin analogues. In HepG2 cells, protein degradation was inhibited significantly less by glargine (15% over 3 hours) than by insulin (approximately 20% over 3 hours) (P < .05). Lipid metabolism was investigated in 3T3-L1 cells. In these cells glucose oxidation to CO2 was effected equally, but glargine was less potent than insulin at inhibiting epinephrine-stimulated lipolysis (EC50 = 1.4 v 0.35 nmol/L, P < .001) and at stimulating lipogenesis (EC50 = 1.27 v 8.06 nmol/L, P < .01). Since the action of insulin on protein and lipid metabolism has been suggested to be due to the metabolism of the hormone, we compared the cellular handling of 125I[A14]-glargine to 125I[A14]-insulin in HepG2 cells. While binding of glargine to the insulin receptor was identical to insulin, degradation of glargine was reduced compared to insulin (16.3% +/- 0.3% v 21.6% +/- 0.4% degraded/h, P < .01). Less degraded glargine than insulin was released from cells previously loaded with radiolabeled material (50.1% +/- 2.4% v 58.3% +/- 1.4%/2 h, P < .02). The amount of intact glargine released was concomitantly increased compared to insulin (44.8% +/- 2.6% v 35.8% +/- 1.4%/2 h, P < .02). These data provide further evidence for a relationship between insulin metabolism and insulin action on protein and lipid metabolism; however, the clinical relevance of these differences is hard to realize, since for the most part glargine, used as a basal insulin, is administered in addition to other shorter-acting insulin or analogues, and their effects will mask or reduce glargine effects on lipolysis and protein degradation. However, these studies do show that properties of insulin other than glucose metabolism and mitogenesis must be considered when studying insulin analogues. PMID- 15281016 TI - Consistently high plasma high-density lipoprotein-cholesterol levels in children in Spain, a country with low cardiovascular mortality. AB - Coronary heart disease (CHD) mortality is relatively low in Spain compared with other developed countries and has remained low despite an apparent increase in mean plasma cholesterol concentration in adults over the last several years. It is accepted that pathologic processes related to arteriosclerosis development begin in childhood and seem to be related to the presence of cardiovascular risk factors at this age. High-density lipoprotein-cholesterol (HDL-C) levels in children have been inversely correlated with the incidence of coronary heart disease in the different countries studied. Childhood plasma lipoprotein profile might contribute to the low coronary heart disease mortality in Spain. Thus, we analyzed data on lipid levels over time in schoolchildren in Spain in the last decade. Plasma lipid levels were analyzed in prepuberal children (6 to 8 years) in 3 school-based surveys performed by our group in Madrid in 1987, 1993, and 1999. A significant increase in plasma total cholesterol (P < .05) and low density lipoprotein-cholesterol (LDL-C) (P < .01) levels in prepuberal children was observed over the last decade. However, the mean concentration of plasma HDL C remained stable and very high. These high levels of plasma HDL-C in Spanish school children may help to explain why the coronary heart disease mortality rate in Spain is low compared with that in other developed countries. PMID- 15281017 TI - Improvement of glucose homeostasis in obese diabetic db/db mice given Plasmodium yoelii glycosylphosphatidylinositols. AB - We have previously reported that infection with Plasmodium yoelii, Plasmodium chabaudi, or injection of extracts from malaria-parasitized red blood cells induces hypoglycemia in normal mice and normalizes the hyperglycemia in streptozotocin (STZ)-diabetic mice. P yoelii glycosylphosphatidylinositols (GPIs) were extracted in chloroform:methanol:water (CMW) (10:10:3), purified by high performance thin layer chromatography (HPTLC) and tested for their insulin mimetic activities. The effects of P yoelii GPIs on blood glucose were investigated in insulin-resistant C57BL/ks-db/db diabetic mice. A single intravenous injection of GPIs (9 and 30 nmol/mouse) induced a significant dose related decrease in blood glucose (P < .001), but insignificantly increased plasma insulin concentrations. A single oral dose of 2.7 micromol GPIs per db/db mouse significantly lowered blood glucose (P < .01). P yoelii GPIs in vitro (0.062 to 1 micromol/L) significantly stimulated lipogenesis in rat adipocytes in a dose-dependent manner both in the presence and absence of 10(-8) mol/L insulin (P < .01). P yoelii GPIs stimulated pyruvate dehydrogenase phosphatase (PDH-Pase) and inhibited both cyclic adenosine monophosphate (cAMP)-dependent protein kinase A and glucose-6-phosphatase (G6Pase). P yoelii GPIs had no effect on the activity of the gluconeogenic enzymes fructose-1,6-bisphosphatase (FBPase) and phosphoenolpyruvate carboxykinase (PEPCK). This is the first report of the hypoglycemic effect of P yoelii GPIs in murine models of type 2 diabetes. In conclusion, P yoelii GPIs demonstrated acute antidiabetic effects in db/db mice and in vitro. We suggest that P yoelii GPIs, when fully characterized, may provide structural information for the synthesis of new drugs for the management of diabetes. PMID- 15281018 TI - The effects of uncoupling protein-1 genotype on lipoprotein cholesterol level in Korean obese subjects. AB - Uncoupling protein-1 (UCP-1) plays a major role in thermogenesis, and has been implicated in the pathogenesis of obesity and metabolic disorders. The purpose of this study was to estimate the effects of A-3826G polymorphism of the UCP-1 gene on the plasma lipid profiles in 190 Korean obese subjects with a body mass index (BMI) more than 30 kg/m2. Height, weight, BMI, wait-to-hip ratio (WHR), obesity index, and body composition were measured and genotype of UCP-1 was analyzed by polymerase chain reaction (PCR) restriction fragment length polymorphism (RFLP) method. Serum concentrations of fasting glucose, total cholesterol, high-density lipoprotein (HDL) cholesterol, and triglyceride were measured. The frequencies of UCP-1 genotypes were AA type, 22.1%; AG type, 53.7%; and GG type, 24.2%; and the frequency of G allele was 0.51. Among many parameters, diastolic blood pressure (DBP) (P = .023) and low-density lipoprotein (LDL) cholesterol (P = .011) were significantly higher in AG and GG types compared with AA type, whereas HDL cholesterol was significantly lower in GG type compared with other types (P < .05). Atherogenic index was significantly higher in GG type compared with AA type (P = 0.027). LDL-to-HDL cholesterol ratio was significantly increased in the order of AA < AG < GG types (P = .001). When the subjects were divided into a normal group and a hyper-LDL cholesterolemia group by LDL cholesterol level of 3.626 mmol/L (140 mg/dL), the frequency of hyper-LDL cholesterolemia was significantly higher in GG type compared with other types by Fisher's exact (chi square) test (P = .05). When logistic regression analysis was conducted to find the risk factors of hyper-LDL cholesterolemia, the odds ratio was 4.115 (P = .03) for GG type of UCP-1 gene. These results suggest that the GG type of the UCP-1 gene has a strong association with increased LDL cholesterol level and might be a significant risk factor for hyper-LDL cholesterolemia among Korean obese subjects. PMID- 15281019 TI - Upregulation of lipogenic enzymes genes expression in white adipose tissue of rats with chronic renal failure is associated with higher level of sterol regulatory element binding protein-1. AB - Chronic renal failure (CRF) frequently results in hypertriglyceridemia and elevated plasma concentration of very-low-density lipoprotein (VLDL). These abnormalities are thought to be primarily due to depressed lipoprotein lipase and hepatic lipase activities, as well as impaired clearance of plasma lipoproteins. Some results suggest that not only lipoproteins catabolism but also their overproduction might contribute to hypertriglyceridemia in CRF. Because sterol regulatory element binding protein (SREBP) plays an important role in the regulation of lipid homeostasis, increased level of this transcription factor might be involved in modulating lipid metabolism in CRF. The purpose of the present study is to determine whether there is an altered regulation of the SREBP 1 in CRF rats and whether the altered regulation of SREBP-1 is associated with the upregulation of lipogenic enzymes genes expression in CRF rats. In the white adipose tissue (WAT) of CRF rats, marked increases in the microsomal (precursor) and nuclear (mature) forms of SREBP-1 have been found. The increase in SREBP-1 was associated with an increased level of lipogenic enzymes (acetyl-coenzyme A [CoA] carboxylase [ACC], adenosine triphosphate-citrate lyase [ACL], fatty acid synthase [FAS], glucose 6-phosphate dehydrogenase [G6PDH], 6-phosphogluconate dehydrogenase [6PGDH], and malic enzyme [ME]) genes expression. In turn, this was associated with an increased rate of fatty acids synthesis in WAT and a significant increase in plasma triacylglycerol (TAG) and VLDL concentration. Our study indicates that WAT SREBP-1 expression is increased in CRF rats and that SREBP-1 may play an important role in the increased fatty acid synthesis. These results reveal another facet of disturbed lipid metabolism in CRF. PMID- 15281020 TI - Liver fat is not a marker of metabolic risk in lean premenopausal women. AB - We examined the independent associations among abdominal adipose tissue (AT) depots, liver fat, cardiorespiratory fitness (CRF), and metabolic risk factors in 86 lean premenopausal women. We measured abdominal AT and liver fat by computed tomography (CT), and CRF by a maximal treadmill exercise test. Liver fat was not related to any abdominal AT depot, metabolic risk factor, or CRF (P > .10). Visceral AT mass (kilograms) remained a significant (P < .05) predictor of total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), TC/high-density lipoprotein cholesterol (HDL-C), and LDL-C/HDL-C after statistical adjustment for CRF. Abdominal subcutaneous AT mass was also a significant (P < .05) correlate of TC/HDL-C and LDL-C/HDL-C after control for CRF. Visceral AT remained a significant predictor (P < .05) of TC and LDL-C after control for abdominal subcutaneous AT. Conversely, subcutaneous AT did not remain a significant correlate after control for visceral AT. However, the deep subcutaneous AT depot remained significantly associated with LDL-C, TC/HDL-C, and LDL-C/HDL-C after control for visceral AT. In contrast, visceral AT remained correlated with triglycerides (TG) alone, after control for the deep subcutaneous AT. These observations suggest that liver fat is not a determinant of metabolic risk in lean women. Conversely, both visceral and the deep subcutaneous depot are determinants of metabolic risk in premenopausal woman despite the absence of obesity. PMID- 15281021 TI - Dietary composition as a determinant of plasma asymmetric dimethylarginine in subjects with mild hypercholesterolemia. AB - Asymmetric dimethylarginine (ADMA) is an endogenous nitric oxide synthase inhibitor that participates in the regulation of vasodilatory function and is also linked to hypertension, whereas its stereoisomere, symmetric dimethylarginine (SDMA), is biologically inactive. Dietary components influence vascular functions and a high-fat meal seems to increase postprandial plasma ADMA levels. However, it has not been published whether diet influences plasma ADMA levels. In this study, we investigated the impact of diet on plasma ADMA and SDMA levels. Thirty-four mildly hypercholesterolemic, otherwise healthy women (n = 14) and men (n = 20) with a mean age of 46.2 years (range, 35 to 62 years) participated in the study. The subjects were examined twice at intervals of 2 months. Seven-day food records were used to analyze diet and alcohol intake. ADMA was measured by using high-performance liquid chromatography (HPLC)-tandem mass spectrometry. In a multivariate analysis (R2 = 0.20, P < .002), low amount of energy received from carbohydrates (r = -0.31, P = .009) and high plasma triglycerides (r = 0.30, P = .01) were predictors of high ADMA plasma levels. Alcohol drinkers had higher plasma ADMA concentrations than abstainers (0.50 +/- 0.13 v 0.42 +/- 0.11 micromol/L, P = .04). Plasma ADMA correlated with systolic (r = 0.60, P = .005) and diastolic blood pressure (r = 0.53, P = .02) in abstainers but not in alcohol drinkers. Plasma SDMA was not associated with any dietary components or with blood pressure. In conclusion, a high amount of dietary carbohydrates is strongly associated with low levels of plasma ADMA. Concentration of ADMA in plasma seems to be higher in alcohol drinkers than in abstainers. PMID- 15281022 TI - Contractile protein breakdown in human leg skeletal muscle as estimated by [2H3] 3-methylhistidine: a new method. AB - 3-Methylhistidine urinary excretion and net balances across the leg or forearm have been used as markers of contractile protein breakdown in muscle tissue. Here we investigate whether infusion of labeled 3-methylhistidine and the measurement of the arteriovenous dilution of the tracer with unlabeled 3-methylhistidine will result in more consistent and precise measurements of 3-methylhistidine rates of appearance and consequently muscle contractile protein breakdown rates in comparison with conventional arteriovenous concentration difference measurements. Six healthy volunteers were studied in the postabsorptive state and received a primed continuous infusion of 3-[2H3-methyl]- methylhistidine and L-[ring-2H5] phenylalanine for 4 hours. 2H3-3-methylhistidine reached an isotopic steady state after 210 minutes in all subjects. Arteriovenous differences of 3 methylhistidine, measured by high-performance liquid chromatography (HPLC), showed both uptake and release from skeletal muscle, which is theoretically not likely to occur. The enrichment of 2H3-3-methylhistidine was consistently lower in the femoral vein than in the artery, and therefore a constant net release of 3 methylhistidine from the leg was observed. The mean rates of appearance for 3 methylhistidine and phenylalanine were 0.44 +/- 0.30 nmol x min(-1) x 100 mL(-1) and 11.2 +/- 5.7 nmol x min(-1) x 100 mL(-1), respectively. In summary, arteriovenous difference measurement of 2H3-3-methylhistidine enrichment is more reliable than measurement of arteriovenous difference of unlabeled 3 methylhistidine. Consequently, measuring rates of appearance from leg muscle using labeled 3-methylhistidine resulted in more consistent and accurate values of contractile protein degradation rates in human skeletal muscle. PMID- 15281023 TI - Food and red wine do not exert acute effects on vascular reactivity. AB - Experimental hyperglycemia and hyperinsulinemia have been shown to affect vascular reactivity. Chronic red wine consumption is associated with less cardiovascular mortality. Whether ingestion of a natural meal and red wine causes acute changes in vascular homeostasis is poorly understood. The aim of the current study was to clarify whether meal ingestion, with and without red wine, exert acute effects on vascular reactivity in healthy humans. We studied vascular reactivity and forearm nitrite balance in 10 healthy subjects under 3 different circumstances: (1) fasting; (2) after ingestion of a standard natural meal (1,050 kcal); and (3) after the same meal enriched with a glass of red wine. We measured forearm blood flow (FBF) by strain-gauge plethismography during intrabrachial, graded infusion of acetylcholine (ACh), sodium nitroprusside (NP), and norepinephrine (NE). We also measured the forearm balance of nitrite before and during ACh infusion. Despite significant increases in plasma glucose and insulin concentrations, the vasodilatory response to Ach and NP after meal ingestion was not different from the fasting response. Similarly, the vasoconstrictory response to NE was similar postprandially and during fasting. Addition of red wine did not modify the response to any of the vasoactive agents. Finally, the forearm nitrite production during Ach infusion was not different in the 3 experimental settings. Food intake, whether associated or not with red wine, does not affect vascular reactivity in normal human subjects. PMID- 15281032 TI - Summertime: Fresh fruits, fresh vegetables, and a healthy life. PMID- 15281024 TI - Mechanism of dichloroacetate-induced hypolactatemia in humans with or without cirrhosis. AB - Dichloroacetate (DCA) has been used as an experimental treatment for lactic acidosis because it lowers plasma lactic acid concentration. Three potential mechanisms could underlie the hypolactatemic action of DCA, but the dominant mechanism in vivo remains unclear. This study tested whether DCA-induced hypolactatemia occurs via decreased lactate production, increased lactate clearance, or decreased rate of glycolysis in healthy humans and in patients with end-stage cirrhosis. Cirrhosis is associated with decreased hepatic pyruvate dehydrogenase (PDH) content. Six healthy volunteers and 7 cirrhotic patients received a primed, constant infusion of 1-13C-pyruvate and 15N-alanine for 5 hours. DCA (35 mg/kg intravenously) was administered at 2 hours. Plasma isotopic enrichment was measured by gas chromatography/mass spectrometry (GC/MS), and exhaled CO2 enrichment by isotope ratio mass spectrometry. Pyruvate and alanine production rates (Ra) were determined by isotope dilution, and pyruvate oxidation calculated as 13CO2 production from 13C-pyruvate. Ra lactate was calculated as the difference between Ra pyruvate and its disposal by oxidation to CO2 and conversion to alanine. Baseline plasma lactate kinetics in cirrhotic patients did not differ from controls. DCA decreased lactate concentration in both groups by approximately 53%. DCA decreased glycolysis (Ra pyruvate) by 24%, increased the fraction of pyruvate oxidized to CO2 by 26%, and decreased pyruvate transamination to alanine by 25%. DCA also inhibited lactate production by 85%, but decreased plasma lactate clearance by 60% in both groups. DCA reduces plasma lactic acid concentration by inhibiting production, via stimulating pyruvate oxidation and inhibiting glycolysis, rather than increasing clearance. In addition, end-stage cirrhosis does not alter either the mechanism or the magnitude of the metabolic response to DCA. PMID- 15281033 TI - Body piercing in the food service industry. PMID- 15281034 TI - Disordered eating across the life span. PMID- 15281035 TI - Career advancement: Tips for success while on the job hunt. PMID- 15281036 TI - Taking nutrition issues to Capitol Hill. PMID- 15281037 TI - Challenges in the meals on wheels program. PMID- 15281038 TI - Use of thickened liquids in skilled nursing facilities. AB - OBJECTIVE: Long-term care residents are routinely provided with thickened liquids for the management of dysphagia. The objective of this study was to identify the prevalence of thickened liquid use in skilled nursing facilities. DESIGN: Facility-wide data were provided by staff at 252 randomly selected skilled nursing facilities owned by 11 multifacility providers. The sample represented 25,470 residents and approximately 20% of all freestanding skilled nursing facilities nationwide. MAIN OUTCOMES MEASURES: Data regarding prevalence of thickened liquid use and facility characteristics were collected during May 2002. Statistical analysis Descriptive statistics included national and regional averages and national percentile distributions. RESULTS: A mean of 8.3% (range 0% to 28%) of residents were receiving thickened liquids, with considerable variation between Centers for Medicare and Medicaid Services regions. Of those receiving thickened liquids, on average 60% received "nectar/syrup" thick, 33% received "honey" thick, and 6% received "pudding/spoon" thick, although the frequencies with which each thickness was prescribed varied widely between facilities (range 0% to 100%). Thickened water was provided to residents in 91.6% of facilities. Nationally, registered dietitian staffing levels were lower on average than speech language pathologist staffing levels. CONCLUSIONS: Thickened liquids are provided to a significant segment of the skilled nursing facility resident population. In the absence of outcomes-based practice standards to guide administrative decisions related to the provision of thickened liquids, dietetics professionals may find regional and national norms helpful for quality assurance processes and to inform resource management decisions in clinical staffing and foodservice. PMID- 15281039 TI - Improvements in nutritional intake and quality of life among frail homebound older adults receiving home-delivered breakfast and lunch. AB - OBJECTIVE: This study evaluated the influence that expanding a home-delivered meals service to include breakfast and lunch would have on the nutritional status and quality of life of at-risk older adults. DESIGN: This cross-sectional field study compared two groups. The breakfast group (n=167) received a home-delivered breakfast and lunch, 5 days per week. The comparison group (n=214) received a home-delivered lunch 5 days per week. Participants' 24-hour food recall, demographics, malnutrition risk, functional status, and surveys of quality of life as health, loneliness, food enjoyment, food security, and depression were obtained. PARTICIPANTS: Study participants were recruited from five Elderly Nutrition Programs involved in the Morning Meals on Wheels breakfast service demonstration project. They formed a geographically and racially/ethnically diverse sample. Participants ranged in age from 60 to 100 years, were functionally limited, and at high nutritional risk. Most were low income, lived alone, and had difficulty shopping or preparing food. STATISTICAL ANALYSIS: Descriptive statistics were used to assess group comparability. Independent sample t tests were used to examine group differences, with Bonferroni's method used to control for familywise Type I error. RESULTS: Breakfast group participants had greater energy/nutrient intakes (P<.05), greater levels of food security (P<.05), and fewer depressive symptoms (P<.05) than comparison group participants. CONCLUSIONS: The addition of a breakfast service to traditional home-delivered meals services can improve the lives of frail, homebound older adults. Agencies should be encouraged to expand meals programs to include a breakfast service to a targeted population. PMID- 15281040 TI - Differences in psychosocial variables by stage of change for fruits and vegetables in older adults. AB - OBJECTIVE: To describe differences in demographic and psychological variables by stage of change for five servings of fruits and vegetables per day in older adults. DESIGN: Cross-sectional survey with data collected in the participant's home or the project office by interviewers. SUBJECTS: 1,253 community-residing individuals 60 years or older (mean age=75 years) living in East Providence, RI. MEASURES: Stage of change; decisional balance; processes of change; self efficacy; dietary intake measured by the National Institutes of Health (NIH) Fruit and Vegetable Screener, the 5 A Day Screener, and the NIH Fat Scan; demographics; and anthropometrics. STATISTICAL ANALYSES: Variables were compared by stage of change using analysis of variance for continuous variables and chi(2) analysis for categorical variables. Servings of fruits and vegetables were transformed (square root) prior to analyses. RESULTS: There was a strong effect of stage of change on intake measured by the Fruit and Vegetable Screener [F(2, 1203)=109, P<.001, eta(2)=.154] and the 5 A Day Screener [F(2, 1203)=128, P<.001, eta(2)=.175] with a linear increase from precontemplation to action/maintenance. There was an overall stage effect on decisional balance, processes of change, and self-efficacy [Wilks's lambda=.703, F(30, 2132)=13.72, P<.001, multivariate eta(2)=.162], and significant univariate effects for all variables. CONCLUSIONS: Self-assessed servings ("How many servings do you usually eat?") can be used for stage classification for older adults. Interventions for older adults in early stages should focus on increasing perceived benefits of healthful eating and cognitive process use. Self-efficacy as well as behavioral processes seem to be important in attaining maintenance. PMID- 15281041 TI - Factors associated with soft drink consumption in school-aged children. AB - OBJECTIVE: To identify factors associated with nonalcoholic carbonated beverage (soft drink) consumption in children. DESIGN: Mail-in surveys collected by Dragonfly, a children's educational magazine distributed nationally to elementary and middle schools, were analyzed. The survey included questions about frequency of soft drink consumption and factors related to soft drink consumption. SUBJECTS AND PARTICIPANTS: The sample consisted of 560 children, 8 to 13 years old, who completed and mailed in the survey. There was an equal distribution of boys and girls (51% and 49%, respectively). STATISTICAL ANALYSES: Frequency distributions were calculated and chi(2) tests were conducted to determine whether soft drink consumption and related factors varied by sex and age. Multivariate logistic regression analyses were conducted to examine the association of soft drink consumption with each factor after adjustment for potential confounders. RESULTS: Preference for the taste of soft drinks was the strongest predictor in the analysis, with those who reported the strongest taste preference 4.50 times more likely (95% confidence interval=2.89-7.04) to consume soft drinks five or more times per week than those with a lower taste preference. Youth whose parents regularly drank soft drinks were 2.88 times more likely (95% confidence interval=1.76-4.72) to consume soft drinks five or more times per week compared with those whose parents did not regularly drink soft drinks. CONCLUSIONS: Results suggest that several factors may be associated with soft drink intake in school-aged children, most notably taste preferences, soft drink consumption habits of parents and friends, soft drink availability in the home and school, and television viewing. Additional research is needed to verify these findings in a representative sample of children. PMID- 15281042 TI - Heightening awareness about soft drink consumption. PMID- 15281043 TI - The Yale Physical Activity Survey for older adults: predictions in the energy expenditure due to physical activity. AB - OBJECTIVE: To evaluate the accuracy of the Yale Physical Activity Survey (YPAS) for older adults. DESIGN: Fourteen-week strictly controlled diet study. SUBJECTS/SETTING: Eleven men and 17 women, age range 55 to 78 years, spent 10 weeks in an outpatient setting and 4 weeks in an inpatient setting at the General Clinical Research Center, Noll Physiological Research Center, The Pennsylvania State University, University Park. INTERVENTION: Subjects were provided dietary energy to maintain body weight within +/-0.5 kg of baseline weight. The daily menus contained 0.8 g protein per kilogram body weight and nonprotein energy as 60% carbohydrate and 40% fat. MAIN OUTCOME MEASURES: Metabolizable energy intake (MEI) was measured at week 14 from the gross energy contents of food, urine, and feces, with corrections for any body composition changes during the last 6 weeks of weight maintenance. Resting energy expenditure (REE) was determined using indirect calorimetry. The thermic effect of feeding (TEF) was estimated to be 10% of the MEI. The energy expenditure due to physical activity (EEPA) was derived by the formula: EEPA=MEI-REE-TEF. This value was compared with the EEPA estimated from the YPAS. Statistical analyses performed Two-factor analysis of variance with repeated measures and paired t tests. RESULTS: At week 14, after a minimum of 6 weeks of sustained weight stability, the derived EEPA was not different from that estimated using the YPAS for the men and the men and women combined, while the YPAS estimate was more than the measured mean value for the women (P<.05). There was wide variability in the accuracy of the EEPA prediction for individual subjects (range=-637 to 794 kcal). APPLICATIONS/CONCLUSIONS: The YPAS may be used, with caution, to estimate the EEPA for groups of older individuals, and may provide inaccurate estimates of the EEPA in older individuals. PMID- 15281044 TI - Hepatic proteins and nutrition assessment. AB - Serum hepatic protein (albumin, transferrin, and prealbumin) levels have historically been linked in clinical practice to nutritional status. This paradigm can be traced to two conventional categories of malnutrition: kwashiorkor and marasmus. Explanations for both of these conditions evolved before knowledge of the inflammatory processes of acute and chronic illness were known. Substantial literature on the inflammatory process and its effects on hepatic protein metabolism has replaced previous reports suggesting that nutritional status and protein intake are the significant correlates with serum hepatic protein levels. Compelling evidence suggests that serum hepatic protein levels correlate with morbidity and mortality. Thus, serum hepatic protein levels are useful indicators of severity of illness. They help identify those who are the most likely to develop malnutrition, even if well nourished prior to trauma or the onset of illness. Furthermore, hepatic protein levels do not accurately measure nutritional repletion. Low serum levels indicate that a patient is very ill and probably requires aggressive and closely monitored medical nutrition therapy. PMID- 15281045 TI - Growth hormone, glutamine, and modified diet for intestinal adaptation. AB - Many patients who undergo extensive resection of the gastrointestinal tract develop intestinal failure from short-bowel syndrome that results in significant malabsorption of fluid, electrolytes, and other nutrients. This may result in dependence on long-term parenteral nutrition. It has been almost a decade since Byrne and colleagues published their research demonstrating enhanced absorption of nutrients, improved weight gain, and reduction in parenteral nutrition requirements with the administration of a combination of growth hormone, glutamine, and a modified diet. Other researchers have conducted similar studies with inconsistent results. A systematic search on electronic databases and the Internet for the purpose of identifying the evidence published to date on this subject was performed. The analysis suggests administering recombinant human growth hormone alone or together with glutamine with or without a modified diet may be of benefit when the appropriate patients are selected for treatment. PMID- 15281046 TI - Persistent oral health problems associated with comorbidity and impaired diet quality in older adults. AB - Chewing, swallowing, and mouth pain (CSP) are identified as indicators of nutritional risk in older adults. Previous research has shown that oral health problems in community-living older rural adults were associated with increased hospitalization. The purpose of this study was to characterize older adults with self-reported persistent CSP problems at baseline and one-year follow-up. Participants were from the Geisinger Rural Aging Study, either with persistent oral problems (PCSP; n=22) or without problems (NCSP; n=125). Demographic, health, and anthropometric data were collected via home visit; diet information was assessed by five, 24-hour recalls collected over 10 months. PCSP subjects reported almost twice the number of medications (4.2 vs 2.6, respectively, P=.008) and diseases (7.0 vs 4.2, respectively, P=.001), with higher occurrence of type 2 diabetes mellitus, peptic ulcers/gastritis, and angina. PCSP participants had lower Healthy Eating Index scores (66.6 vs 70.6, respectively, P=.04), significantly lower intakes of vitamin A, and higher prevalence of inadequate intakes of vitamins B-6 and A. These results indicate that impaired intake of certain foods and nutrients is associated with persistent oral health problems. Oral status is an important component of overall health and should be monitored for intervention. PMID- 15281047 TI - Reaching low-income families: Focus group results provide direction for a behavioral approach to WIC services. AB - Supplemental Nutrition Program for Women, Infants, and Children (WIC) families were asked to identify motivators and barriers to health behavior change and preferred approaches to nutrition education in WIC. Six focus groups involved a total of 41 English-speaking WIC participants and addressed parenting, family meals, food preparation, and physical activity. The discussions were audiotaped, transcribed, and analyzed using NUD*IST software (Non-Numerical Unstructured Data Indexing, Searching, and Theorizing, version 4.0. Thousand Oaks, CA: Sage Publications Software, 1997). Key barriers to behavior change included inadequate parenting skills, lack of knowledge, unhealthy social environments, lack of time, and lack of social or financial support. Key motivators included feelings of responsibility, concern for child health and development, and positive social support. Participants identified facilitated discussions, support groups, cooking classes, and a WIC Web site as preferred methods of nutrition education. Results provided the foundation for the Healthy Habits nutrition education modules implemented in the Washington State WIC program and can be used to improve future nutrition education in WIC. PMID- 15281048 TI - Using forecasting techniques to predict meal demand in Title IIIc congregate lunch programs. AB - The purpose of this study was to determine which forecasting model would most accurately predict meal demand in Title IIIc congregate lunch programs designed for serving older adults. Forecasting techniques including naive, moving average (three versions) and simple exponential smoothing were applied to data collected over a 4-month period from seven meal sites located in a large urban area. An analysis of the forecasting models using mean absolute deviations and mean squared errors indicated that simple mathematical forecasting techniques provided better predictions of meal demand than did the naive method for all sites. In four of the seven sites, exponential smoothing was the best forecasting model, whereas in the remaining sites, moving average models provided the best forecast. Implications are discussed. PMID- 15281049 TI - Study finds Chapel Hill, NC, soup kitchen serves nutritious meals. AB - Soup kitchens attempt to improve the food security of low-income individuals, but the results of their efforts are rarely researched. We focused our study on the Inter-Faith Council Soup Kitchen (IFC) near the University of North Carolina Chapel Hill (UNC) in Chapel Hill, NC. The IFC uses no centralized nutrition planning and relies heavily on volunteer cooks, yet we found their meals to be highly nutrient-dense when averaged over a 1-month time frame and compared with the Dietary Reference Intakes (DRIs) and the Daily Reference Values (DRVs). In fact, the only nutrients needing improvement were vitamin D, folate, and calcium. The number of servings per meal was also substantially more than one third of the US Department of Agriculture Food Guide Pyramid recommendations, except for dairy at all meals, vegetables at breakfast, and fruit at dinner. PMID- 15281050 TI - Advanced glycoxidation end products in commonly consumed foods. AB - OBJECTIVE: Advanced glycoxidation end products (AGEs), the derivatives of glucose protein or glucose-lipid interactions, are implicated in the complications of diabetes and aging. The objective of this article was to determine the AGE content of commonly consumed foods and to evaluate the effects of various methods of food preparation on AGE production. DESIGN: Two-hundred fifty foods were tested for their content in a common AGE marker (epsilon)N-carboxymethyllysine (CML), using an enzyme-linked immunosorbent assay based on an anti-CML monoclonal antibody. Lipid and protein AGEs were represented in units of AGEs per gram of food. RESULTS: Foods of the fat group showed the highest amount of AGE content with a mean of 100+/-19 kU/g. High values were also observed for the meat and meat-substitute group, 43+/-7 kU/g. The carbohydrate group contained the lowest values of AGEs, 3.4+/-1.8 kU/g. The amount of AGEs present in all food categories was related to cooking temperature, length of cooking time, and presence of moisture. Broiling (225 degrees C) and frying (177 degrees C) resulted in the highest levels of AGEs, followed by roasting (177 degrees C) and boiling (100 degrees C). CONCLUSIONS: The results indicate that diet can be a significant environmental source of AGEs, which may constitute a chronic risk factor for cardiovascular and kidney damage. PMID- 15281051 TI - Is religious orientation associated with fat and fruit/vegetable intake? AB - We explored associations of religious orientation with dietary behavior among participants in the Eating for a Healthy Life Study (EHL), a randomized low-fat, high-fruit/vegetable dietary intervention trial in religious organizations. Data in this report are from baseline telephone surveys of 2,375 people, which assessed dietary behaviors (Fat- and Fiber-Related Diet Behavior Questionnaire) and religiosity (Allport-Ross Religious Orientation Scale). After adjusting for demographic characteristics, higher extrinsic (socially motivated) religious orientation was positively associated with low-fat dietary fat behaviors (P=.0438). No associations were observed for dietary behaviors and intrinsic (life based on religious beliefs) religious orientation. These results support further exploration of religious orientation's potential influence on dietary behaviors and its applicability to dietary interventions. PMID- 15281055 TI - Nephridial and gonoduct distribution patterns in Nerillidae (Annelida: Polychaeta) examined by tubulin staining and cLSM. AB - The distribution and configuration of nephridia and gonoducts are described for seven species from seven genera of the interstitial polychaete family Nerillidae. The ciliated nephridia and gonoducts were identified by tubulin staining and examined with a confocal laser scanning microscope. The following species of the seven to nine-segmented nerillids were examined: Leptonerilla prospera, Nerilla antennata (nine segments); Nerillidium mediterraneum, Trochonerilla mobilis, Gen. sp. A (eight segments); and Aristonerilla brevis, Paranerilla limicola (seven segments). Two of the examined species are hermaphroditic (N. mediterraneum and Gen. sp. A). Segmented nephridia can be found from the first to the last segment, with a total of two to five pairs. One to three pairs of segmented spermioducts are present in all species. One pair of gonoducts is found in all species, except for P. limicola, where they are absent. Nephridia vary in length from half to almost twice the length of a segment and may be curled up in loops. In A. brevis and P. limicola the nephridia are discontinuously ciliated. The distribution and configuration of spermioducts and gonoducts are also variable, although to a lesser extent. The spermioduct distribution is generally consistent within genera and therefore of systematic significance. Nephridia and gonoducts are never found together in the same segments, and the results indicate that gonoducts and nephridia have developed from the same anlagen. The distribution patterns of nephridia and gonoducts are discussed with respect to segmentation, systematics, and development. PMID- 15281056 TI - New unusual miniature-like wing mutation in Drosophila virilis. AB - A new recessive, sex-linked, nonlethal in the homozygote, wing mutation in Drosophila virilis was studied using a hybridological assay, light microscopy, and transmission electron microscopy. The mutants have abnormally small wings; the phenotype is attributed to a cell-autonomous reduction in the size of the epidermal cells of the differentiating wing. The phenotype is also characterized by abnormally oriented wing hairs, wavy wing edge, temperature sensitivity, and some abnormalities in the wing veins. PMID- 15281057 TI - Comparative analysis of masseter fiber architecture in tree-gouging (Callithrix jacchus) and nongouging (Saguinus oedipus) callitrichids. AB - Common marmosets (Callithrix jacchus) and cotton-top tamarins (Saguinus oedipus) (Callitrichidae, Primates) share a broadly similar diet of fruits, insects, and tree exudates. Common marmosets, however, differ from tamarins by actively gouging trees with their anterior teeth to elicit tree exudate flow. During tree gouging, marmosets produce relatively large jaw gapes, but do not necessarily produce relatively large bite forces at the anterior teeth. We compared the fiber architecture of the masseter muscle in tree-gouging Callithrix jacchus (n = 10) to nongouging Saguinus oedipus (n = 8) to determine whether the marmoset masseter facilitates producing these large gapes during tree gouging. We predict that the marmoset masseter has relatively longer fibers and, hence, greater potential muscle excursion (i.e., a greater range of motion through increased muscle stretch). Conversely, because of the expected trade-off between excursion and force production in muscle architecture, we predict that the cotton-top tamarin masseter has more pinnate fibers and increased physiological cross-sectional area (PCSA) as compared to common marmosets. Likewise, the S. oedipus masseter is predicted to have a greater proportion of tendon relative to muscle fiber as compared to the common marmoset masseter. Common marmosets have absolutely and relatively longer masseter fibers than cotton-top tamarins. Given that fiber length is directly proportional to muscle excursion and by extension contraction velocity, this result suggests that marmosets have masseters designed for relatively greater stretching and, hence, larger gapes. Conversely, the cotton top tamarin masseter has a greater angle of pinnation (but not significantly so), larger PCSA, and higher proportion of tendon. The significantly larger PCSA in the tamarin masseter suggests that their masseter has relatively greater force production capabilities as compared to marmosets. Collectively, these results suggest that the fiber architecture of the common marmoset masseter is part of a suite of features of the masticatory apparatus that facilitates the production of relatively large gapes during tree gouging. PMID- 15281058 TI - Life stages and reproductive components of the Marmorkrebs (marbled crayfish), the first parthenogenetic decapod crustacean. AB - Recently, we briefly reported on the first case of parthenogenesis in the decapod Crustacea which was found in the Marmorkrebs or marbled crayfish, a cambarid species of unknown geographic origin and species identity. Curiously, this animal is known only from aquarium populations, where it explosively propagates. By means of light and electron microscopic techniques we have now investigated the reproductive components of this crayfish, using more than 100 specimens ranging from hatchling to repeatedly spawned adult. Additionally, we documented its principal life stages. Our results revealed that the external sexual characters and also the gonads of the marbled crayfish are purely female, making this fast reproducing species a good model for investigating female reproductive features in crayfish. Testicular tissues, ovotestes, or male gonoducts, gonopores, or gonopods were never found, either in small juveniles or large adult specimens, confirming the parthenogenetic nature of this crayfish. Parthenogenesis may have arisen spontaneously or by interspecific hybridization since Wolbachia-like feminizing microorganisms were not found in the ovaries. The external sexual characters of the marbled crayfish are first recognized in Stage 4 juveniles and are structurally complete approximately 2 months after hatching in specimens of approximately 2 cm total length. In the same life stage the ovary is fully differentiated as well, although the oocytes are in previtellogenic and primary vitellogenic stages only. The architecture of the mature ovary and also the synchronous maturation of cohorts of primary vitellogenic oocytes by secondary vitellogenesis are in general agreement with data published on ovaries of bisexual crayfish. New results were obtained with respect to the muscular nature of the ovarian envelope and its extensive proliferation after the first spawning, the distribution of hemal sinuses in the ovarian envelope and in the interstitium around the oogenetic pouches, the high transport activity of the follicle cells, and the colonization of oogenetic pouches by previtellogenic oocytes that originate in the germaria. Investigation of the nuclei of oocytes in the germaria and oogenetic pouches revealed no signs of meiosis, as usually found in females of bisexual decapods, suggesting that parthenogenesis in the marbled crayfish might be an apomictic thelytoky. The detection of new rickettsial and coccidian infections in the ovary and further organs raises fears that the marbled crayfish might endanger native European species by transmission of pathogens once escaped into the wild. PMID- 15281059 TI - Excurrent duct system of the male turtle Chrysemys picta. AB - The epididymis and efferent duct system of the turtle Chrysemys picta were examined. Seminiferous tubules are drained by a series of ducts that form a rete exterior to the tunica albuginea. The rete is located lateral to the testis and consists of anastamosing tubules of varying diameters, lined by a simple epithelium consisting of squamous to cuboidal cells. The rete is highly vascularized. A series of tubules (efferent ductules) connect the rete to the epididymis proper. The efferent ductules are highly convoluted, running between the epididymal tubules and are of varying diameters. The simple columnar epithelium lining these tubules possesses tight junctions, with every third or fourth cell possessing long cilia that protrude into the lumen. The cytoplasm of these epithelial cells contains abundant mitochondria. In the central portion of the efferent ductule, epithelial cells possess granules that appear to be secreted into the lumen by an apocrine process. The epididymis proper is a single, long, highly convoluted tubule that receives efferent ductules along its entire length. It is lined by a pseudostratified epithelium containing several cell types. The most abundant cell (vesicular cell) lacks cilia, but has a darkly staining apical border due to numerous small vesicles immediately beneath the luminal membrane. The small vesicles appear to fuse with each other basally to form larger vesicles. These cells appear to have an absorptive function, and occasionally sperm are embedded in their cytoplasm. The second-most abundant cell is a basal cell found along the basement membrane. The number of these cells fluctuates throughout the year, being most abundant in late summer and early fall. A small narrow cell with an oval nucleus and darkly staining cytoplasm, extending from the basement membrane to the apical surface, is present in small numbers, particularly in the caudal regions of the epididymis. This cell is frequently found in association with another narrow cell having a rounded nucleus and abundant mitochondria in its cytoplasm. PMID- 15281060 TI - Basic fibroblast growth factor, neurofilament, and glial fibrillary acidic protein immunoreactivities in the myenteric plexus of the rat esophagus and colon. AB - The enteric nervous system consists of a number of interconnected networks of neuronal cell bodies and fibers as well as satellite cells, the enteric glia. Basic fibroblast growth factor (bFGF) is a mitogen for a variety of mesodermal and neuroectodermal-derived cells and its presence has been described in many tissues. The present work employs immunohistochemistry to analyze neurons and glial cells in the esophageal and colic enteric plexus of the Wistar rat for neurofilament (NF) and glial fibrillary acidic proteins (GFAP) immunoreactivity as well as bFGF immunoreactivity in these cells. Rats were processed for immunohistochemistry; the distal esophagus and colon were opened and their myenteric plexuses were processed as whole-mount preparations. The membranes were immunostained for visualization of NF, GFAP, and bFGF. NF immunoreactivity was seen in neuronal cell bodies of esophageal and colic enteric ganglia. GFAP immunoreactive enteric glial cells and processes were present in the esophageal and colic enteric plexuses surrounding neuronal cell bodies and axons. A dense net of GFAP-immunoreactive processes was seen in the ganglia and connecting strands of the myenteric plexus. bFGF immunoreactivity was observed in the cytoplasm of the majority of the neurons in the enteric ganglia of esophagus and colon. The two-color immunoperoxidase and immunofluorescence methods revealed bFGF immunoreactivity also in the nucleus of GFAP-positive enteric glial cells. The results suggest that immunohistochemical localization of NF and GFAP may be an important tool in the study of the plasticity in the enteric nervous system. The presence of bFGF in neurons and glia of the myenteric plexus of the esophagus and the colon indicates that this neurotrophic factor may exert autocrine and paracrine actions in the enteric nervous system. PMID- 15281061 TI - Development of the paired fins in the paddlefish, Polyodon spathula. AB - In Polyodon spathula, the pectoral fin radials, with the exception of the metapterygium, are derived from the decomposition of a single continuous cartilage fin plate that is continuous with the scapulocoracoid. This cartilage sheet develops two interior splits to form three precursor pieces, and these decompose in a predictable way to generate the propterygium and radials. The metapterygium is an extension of the scapulocoracoid that segments off of it during early development. To our knowledge, this has not been reported for acipenserids or other basal actinopterygians. In teleosts, the proximal radials also develop from the "break up" of an initially continuous paddle-like sheet of cartilage along the posterior edge of the scapulocoracoid, and in Polypterus and sharks a similar pattern holds. Thus, the pattern observed in Polyodon may represent the basal developmental condition for the gnathostome pectoral fin. The process underlying development of the superficially similar cartilages of the pelvic and pectoral fins is different. In the pectoral fin, the metapterygium is segmented off of the scapulocoracoid and other radials form from the decomposition of the cartilage plate. In contrast, individual rod-like basipterygial elements form in a close one-to-one correspondence with the middle radials of the pelvic fin, but later fuse to form an anterior element that is branched in appearance. To evaluate further claims of similarity among the pectoral and pelvic fin elements of various fishes, the course of the development of these structures must be observed. The pectoral fin and girdle in Polyodon ossifies in a different sequence than that proposed as ancestral (and highly conserved) for actinopterygians: the supracleithrum ossifies significantly before the cleithrum. The later ossification of the cleithrum in Polyodon may be related to the primary use of the caudal fin vs. the pectoral fins in their locomotion. PMID- 15281062 TI - Fine structure and immunocytochemistry of monotreme hairs, with emphasis on the inner root sheath and trichohyalin-based cornification during hair evolution. AB - The fine structure of hairs in the most ancient extant mammals, the monotremes, is not known. The present study analyzes the ultrastructure and immunocytochemistry for keratins, trichohyalin, and transglutaminase in monotreme hairs and compares their distribution with that present in hairs of the other mammals. The overall ultrastructure of the hair and the distribution of keratins is similar to that of marsupial and placental hairs. Acidic and basic keratins mostly localize in the outer root sheath. The inner root sheath (IRS) comprises 4 8 cell layers in most hairs and forms a tile-like sheath around the hair shaft. No cytological distinction between the Henle and Huxley layers is seen as cells become cornified about at the same time. Externally to the last cornified IRS cells (homologous to the Henle layer), the companion layer contains numerous bundles of keratin. Occasionally, some granules in the companion layer show immunoreactivity for the trichohyalin antibody. This further suggests that the IRS in monotremes is ill-defined, as the companion layer of placental hairs studied so far does not express trichohyalin. A cross-reactivity with an antibody against sheep trichohyalin is present in the IRS of monotremes, suggesting conserved epitopes across mammalian trichohyalin. Trichohyalin granules in the IRS consist of a framework of immunolabeled coarse filaments of 10-12 nm. The latter assume a parallel orientation and lose the immunoreactivity in fully cornified cells. Transglutaminase immunolabeling is diffuse among trichohyalin granules and among the parallel 10-12 nm filaments of maturing inner root cells. Transglutaminase is present where its substrate, trichohyalin, is modified as matrix protein. Cornification of IRS is different from that of hair fiber cuticle and from that of the cornified layer of the epidermis above the follicle. The different consistency among cuticle, IRS, and corneous layer of the epidermis determines separation between hair fiber, IRS, and epidermis. This allows the hair to exit on the epidermal surface after shedding from the IRS and epidermis. Based on comparative studies of reptilian and mammalian skin, a speculative hypothesis on the evolution of the IRS and hairs from the skin of synapsid reptiles is presented. PMID- 15281063 TI - Evolutionary transformation from muscular to hydraulic movements in spider (Arachnida, Araneae) genitalia: a study based on histological serial sections. AB - The male genitalia of 107 spider species representing 73 families were serially sectioned and studied with an emphasis on muscles moving the genital bulb. As a rule, most non-Entelegynae have two bulbal muscles, most Entelegyne have none, but many exceptions occur. Variation also occurs with regard to origin and attachment of bulbal muscles. There appears to be a trend towards a shift of the origin from proximal (Liphistius, Atypus) to more distal palpal segments (Haplogynae). In most Entelegynae the muscular movement is replaced by hydraulic movement caused by expanding membranes (hematodochae). Hematodochae probably permit increased bulbal rotation and movements of higher complexity. New evidence is presented arguing against Palpimanidae being representatives of Entelegynae. Bulbal glands other than those discharging into the sperm duct (previously known in Amaurobiidae and Dictynidae only) are described in several entelegyne families. PMID- 15281064 TI - Development of the islets, exocrine pancreas, and related ducts in the Nile tilapia, Oreochromis niloticus (Pisces: Cichlidae). AB - Pancreatic development and the relationship of the islets with the pancreatic, hepatic, and bile ducts were studied in the Nile tilapia, Oreochromis niloticus, from hatching to the onset of maturity at 7 months. The number of islets formed during development was counted, using either serial sections or dithizone staining of isolated islets. There was a general increase in islet number with both age and size. Tilapia housed in individual tanks grew more quickly and had more islets than siblings of the same age left in crowded conditions. The pancreas is a compact organ in early development, and at 1 day posthatch (dph) a single principal islet, positive for all hormones tested (insulin, SST-14, SST 28, glucagon, and PYY), is partially surrounded by exocrine pancreas. However, the exocrine pancreas becomes more disseminated in older fish, following blood vessels along the mesenteries and entering the liver to form a hepatopancreas. The epithelium of the pancreatic duct system from the intercalated ducts to the main duct entering the duodenum was positive for glucagon and SST-14 in 8 and 16 dph tilapia. Individual insulin-immunopositive cells were found in one specimen. At this early stage in development, therefore, the pancreatic duct epithelial cells appear to be pluripotent and may give rise to the small islets found near the pancreatic ducts in 16-37 dph tilapia. Glucagon, SST-14, and some PPY positive enteroendocrine cells were present in the intestine of the 8 dph larva and in the first part of the intestine of the 16 dph juvenile. Glucagon and SST 14-positive inclusions were found in the apical cytoplasm of the mid-gut epithelium of the 16 dph tilapia. These hormones may have been absorbed from the gut lumen, since they are produced in both the pancreatic ducts and the enteroendocrine cells. At least three hepatic ducts join the cystic duct to form the bile duct, which runs alongside the pancreatic duct to the duodenum. PMID- 15281065 TI - Fine structure of marsupial hairs, with emphasis on trichohyalin and the structure of the inner root sheath. AB - The fine structure and cornification of marsupial hairs are unknown. The distribution of keratins, trichohyalin, and transglutaminase in marsupial hairs was studied here for the first time by electron microscopy and immunocytochemistry. The localization of acidic and basic keratins in marsupial hairs is similar to that of hairs in placental mammals, and the keratins are mainly localized in the outer root sheath and surrounding epidermis. Marsupial trichohyalin in both medulla and inner root sheath (IRS) cross-reacts with a trichohyalin antibody that recognizes trichohyalin across placental species, indicating a common epitope(s) among mammalian trichohyalin. Roundish to irregular trichohyalin granules are composed of a network of immunolabeled 10-15 nm-thick coarse filaments within an amorphous matrix in which a weak labeling for transglutaminases is present. This suggests that the enzyme, and its substrate trichohyalin, are associated in mature granules. Transglutaminase labeling mainly occurs in condensing chromatin of mature cells of the outer and inner root sheaths, suggesting formation of the nuclear envelope connected with terminal differentiation of these cells. In mature Huxley or Henle layers the filaments lose the immunolabeling for trichohyalin when they are reoriented into parallel rows linked by short bridges, thus suggesting that the filaments with their reactive epitopes are chemically modified during cornification, as seen in the IRS of hairs of placental mammals. The Huxley layer probably acts as a cushion, absorbing the tensions connected with the distalward movement of the growing hair fiber. Variations in stratification of the Huxley layer are probably related to the diameter of the hair shaft. The cytoplasmic and junctional connections between cells of the Huxley layer and the companion layer and the outer root sheath enhance the grip of the IRS and hair fiber within the follicle. The role of cells of the IRS in sculpturing the fiber cuticle and in the mechanism of shedding that allows the exit of hair on the epidermal surface in mammals are discussed. PMID- 15281066 TI - Constitutive expression of p21H-ras(Val12) in pyramidal neurons results in reorganization of mouse neocortical afferents. AB - The effect of constitutive expression of p21H-ras(Val12) in pyramidal neurons upon the establishment of afferent input has been investigated in the primary somatosensory cortex of transgenic mice. In these animals, relevant transgene expression is confined to cortical pyramidal neurons and starts postnatally at a period when neuronal morphogenesis has been largely completed. We have shown recently that overexpression of p21H-ras(Val12) in these cells results in considerable enlargement of their size and consequently in expansion of the cortex. In the present study we demonstrate that the density of terminals representing intra- or interhemispheric afferents within cortical layers II/III, however, is only slightly decreased. The density of thalamocortical boutons within layer IV is even higher and the number of afferent contacts to transgenic pyramidal neurons is significantly increased compared to the wild-type. The number of catecholaminergic and cholinergic terminals is augmented proportionally to cortical size or even overproportionally, respectively. Along intercortical and striatal fibers arising from p21H-ras(Val12)-expressing pyramidal neurons, frequency of varicosities is significantly increased, but remains unchanged on cortical cholinergic and catecholaminergic axons originating from "nontransgenic" neurons. Additionally, a higher number of multiple synaptic bodies are found in transgenic mice, suggesting subtle effects on synaptic plasticity. It is concluded that the enlargement of pyramidal neurons due to transgenic expression of p21H-ras(Val12) is paralleled by significant changes in the quantity and pattern of afferent connections. Moreover, expression of p21H-ras(Val12) in pyramidal cells induces an enhanced establishment of efferent boutons. PMID- 15281068 TI - RhoA/ROCK and Cdc42 regulate cell-cell contact and N-cadherin protein level during neurodetermination of P19 embryonal stem cells. AB - RhoGTPases regulate actin-based signaling cascades and cellular contacts. In neurogenesis, their action modulates cell migration, neuritogenesis, and synaptogenesis. Murine P19 embryonal stem cells differentiate to neurons upon aggregation in the presence of retinoic acid, and we previously showed that RhoA and Cdc42 RhoGTPases are sequentially up-regulated during neuroinduction, suggesting a role at this very early developmental stage. In this work, incubation of differentiating P19 cells with C3 toxin resulted in decreased aggregate cohesion and cadherin protein level. In contrast, C3 effects were not observed in cells overexpressing recombinant dominant active RhoA. On the other hand, C3 did not affect cadherin in uninduced cells and their postmitotic neuronal derivatives, respectively expressing E- and N-cadherin. RhoA is thus influential on cell aggregation and cadherin expression during a sensitive time window that corresponds to the switch of E- to N-cadherin. Cell treatment with Y27632 inhibitor of Rho-associated-kinase ROCK, or advanced overexpression of Cdc42 by gene transfer of a constitutively active form of the protein reproduced C3 effects. RhoA-antisense RNA also reduced cadherin level and the size of cell aggregates, and increased the generation of fibroblast-like cells relative to neurons following neuroinduction. Colchicin, a microtubule disrupter, but not cytochalasin B actin poison, importantly decreased cadherin in neurodifferentiating cells. Overall, our results indicate that the RhoA/ROCK pathway regulates cadherin protein level and cell-cell interactions during neurodetermination, with an impact on the efficiency of the process. The effect on cadherin seems to involve microtubules. The importance of correct timing of RhoA and Cdc42 functional expression in neurogenesis is also raised. PMID- 15281069 TI - Methanol exposure interferes with morphological cell movements in the Drosophila embryo and causes increased apoptosis in the CNS. AB - Despite the significant contributions of tissue culture and bacterial models to toxicology, whole animal models for developmental neurotoxins are limited in availability and ease of experimentation. Because Drosophila is a well understood model for embryonic development that is highly accessible, we asked whether it could be used to study methanol developmental neurotoxicity. In the presence of 4% methanol, approximately 35% of embryos die and methanol exposure leads to severe CNS defects in about half those embryos, where the longitudinal connectives are dorsally displaced and commissure formation is severely reduced. In addition, a range of morphological defects in other germ layers is seen, and cell movement is adversely affected by methanol exposure. Although we did not find any evidence to suggest that methanol exposure affects the capacity of neuroblasts to divide or induces inappropriate apoptosis in these cells, in the CNS of germ band retracted embryos, the number of apoptotic nuclei is significantly increased in methanol-exposed embryos in comparison to controls, particularly in and adjacent to the ventral midline. Apoptosis contributes significantly to methanol neurotoxicity because embryos lacking the cell death genes grim, hid, and reaper have milder CNS defects resulting from methanol exposure than wild-type embryos. Our data suggest that when neurons and glia are severely adversely affected by methanol exposure, the damaged cells are cleared by apoptosis, leading to embryonic death. Thus, the Drosophila embryo may prove useful in identifying and unraveling mechanistic aspects of developmental neurotoxicity, specifically in relation to methanol toxicity. PMID- 15281067 TI - Hedgehog and Fgf signaling pathways regulate the development of tphR-expressing serotonergic raphe neurons in zebrafish embryos. AB - Serotonin (5HT) plays major roles in the physiological regulation of many behavioral processes, including sleep, feeding, and mood, but the genetic mechanisms by which serotonergic neurons arise during development are poorly understood. In the present study, we have investigated the development of serotonergic neurons in the zebrafish. Neurons exhibiting 5HT-immunoreactivity (5HT-IR) are detected from 45 h postfertilization (hpf) in the ventral hindbrain raphe, the hypothalamus, pineal organ, and pretectal area. Tryptophan hydroxylases encode rate-limiting enzymes that function in the synthesis of 5HT. As part of this study, we cloned and analyzed a novel zebrafish tph gene named tphR. Unlike two other zebrafish tph genes (tphD1 and tphD2), tphR is expressed in serotonergic raphe neurons, similar to tph genes in mammalian species. tphR is also expressed in the pineal organ where it is likely to be involved in the pathway leading to synthesis of melatonin. To better understand the signaling pathways involved in the induction of the serotonergic phenotype, we analyzed tphR expression and 5HT-IR in embryos in which either Hh or Fgf signals are abrogated. Hindbrain 5HT neurons are severely reduced in mutants lacking activity of either Ace/Fgf8 or the transcription factor Noi/Pax2.1, which regulates expression of ace/fgf8, and probably other genes encoding signaling proteins. Similarly, serotonergic raphe neurons are absent in embryos lacking Hh activity confirming a conserved role for Hh signals in the induction of these cells. Conversely, over-activation of the Hh pathway increases the number of serotonergic neurons. As in mammals, our results are consistent with the transcription factors Nk2.2 and Gata3 acting downstream of Hh activity in the development of serotonergic raphe neurons. Our results show that the pathways involved in induction of hindbrain serotonergic neurons are likely to be conserved in all vertebrates and help establish the zebrafish as a model system to study this important neuronal class. PMID- 15281070 TI - Target-derived and locally derived neurotrophins support retinal ganglion cell survival in the neonatal rat retina. AB - Brain-derived neurotrophic factor (BDNF) and neurotrophin-4/5 (NT-4/5) protein and mRNA are found in the neonatal rat retina and also in target sites such as the superficial layers of the superior colliculus. Both neurotrophins support neonatal retinal ganglion cell survival in vitro. In vivo, injections of recombinant BDNF and NT-4/5 reduce naturally occurring cell death as well as death induced by removal of the contralateral superior colliculus. In the latter case, the peak of retinal ganglion cell death occurs about 24 h postlesion. We wished to determine: whether a similar time-course of degeneration occurs after selective removal of target cells or depletion of target-derived trophic factors, and whether ganglion cell viability also depends on intraretinally derived neurotrophins. Retinal ganglion cell death was measured 24 and 48 h following injections of kainic acid or a mixture of BDNF and NT-4/5 blocking antibodies into the superior colliculus and 24 h after intraocular injection of the same antibodies. Retinotectally projecting ganglion cells were identified by retrograde labeling with the nucleophilic dye diamidino yellow. We show that collicular injections of either kainic acid or BDNF and NT-4/5 blocking antibodies significantly increased retinal ganglion cell death in the neonatal rat 24 h postinjection, death rates returning to normal by 48 h. This increase in death was greatest following collicular injections; however, death was also significantly increased 24 h following intravitreal antibody injection. Thus retinal ganglion cell survival during postnatal development is not only dependent upon trophic factors produced by central targets but may also be influenced by local intraretinal neurotrophin release. PMID- 15281071 TI - Cellular bases of activity-dependent paralysis in Drosophila stress-sensitive mutants. AB - Stress-sensitive mutants in Drosophila have been shown to exhibit activity dependent defects in neurotransmission. Using the neuromuscular junction (NMJ), this study investigates synaptic function more specifically in two stress sensitive mutants: stress-sensitive B (sesB), which encodes a mitochondrial ADP/ATP translocase (ANT); and Atpalpha(2206), a conditional mutant of the Na+/K+ ATPase alpha-subunit. Mechanical shock induces a period of brief paralysis in both homozygous and double heterozygous mutants, but further analysis revealed distinct activity-dependent neurotransmission lesions in each mutant. Basal neurotransmission appeared similar to wild-type controls in both mutants under low frequency stimulation. High frequency stimulation, however, caused pronounced synaptic fatigue as well as slow and incomplete synaptic recovery in sesB mutants while Atpalpha(2206) mutants displayed an increase (25-fold) in synaptic failures. Perhaps to compensate for these activity dependent defects, the neuromuscular synapse was found to be overgrown in both mutants. Passive electrotonic stimulation, which initiates synaptic transmission independent of action potentials, ameliorated synaptic failures and resulted in increased neurotransmission amplitude in Atpalpha(2206) mutants. In addition, spontaneous synaptic vesicle fusion rates were increased in Atpalpha(2206) mutants, suggesting that, in the absence of action potential requirements, these synaptic terminals are healthy, if not hyperactive. Dye labeling studies revealed aberrant synaptic vesicle cycling in sesB mutants indicating a reduction of functional synaptic vesicles. We therefore postulate that both stress-sensitive mutants harbor unique neurotransmission defects: Atpalpha(2206) mutants are unable to maintain ionic gradients required during repetitive action potential propagation, and sesB mutants cannot maintain synaptic vesicle cycling during periods of high demand. PMID- 15281072 TI - Exogenous testosterone prevents motoneuron atrophy induced by contralateral motoneuron depletion. AB - Gonadal steroids exhibit neuroprotective and neurotherapeutic effects. The lumbar spinal cord of male rats contains a highly androgen-sensitive population of motoneurons, the spinal nucleus of the bulbocavernosus (SNB), whose morphology and function are dependent on testosterone in adulthood. Unilateral SNB motoneuron depletion induces dendritic atrophy in contralateral SNB motoneurons, but this atrophy is reversed in previously castrated males treated with testosterone. In the present experiment we test the hypothesis that the morphology of SNB motoneurons is protected from atrophy after contralateral motoneuron depletion by exogenous testosterone alone (i.e., with no delay between castration and testosterone replacement). We unilaterally depleted SNB motoneurons by intramuscular injection of cholera toxin conjugated saporin. Simultaneously, some saporin-injected rats were castrated and immediately given replacement testosterone. Four weeks later, contralateral SNB motoneurons were labeled with cholera toxin conjugated HRP, soma sizes were measured, and dendritic arbors were reconstructed. Contralateral SNB motoneuron depletion induced somal atrophy and dendritic retraction, but testosterone treatment prevented both of these effects. Thus, the presence of high-normal levels of testosterone prevents motoneuron atrophy induced by contralateral motoneuron depletion. These data support a therapeutic role for testosterone in preventing atrophy induced by motoneuron injury. PMID- 15281073 TI - Species differences in the regulation of tyrosine hydroxylase in Cnemidophorus whiptail lizards. AB - Evolution of behavioral phenotype involves changes in the underlying neural substrates. Cnemidophorus whiptail lizards enable the study of behavioral and neural evolution because ancestral species involved in producing unisexual, hybrid species still exist. Catecholaminergic systems modulate the expression of social behaviors in a number of vertebrates, including whiptails, and therefore we investigated how changes in catecholamine production correlated with evolutionary changes in behavioral phenotype by measuring the size and number of catecholamine producing (tyrosine hydroxylase-immunoreactive, or TH-ir) cells across the reproductive cycle in females from two related whiptail species. Cnemidophorusuniparens is a triploid, parthenogenetic species that arose from hybridization events involving the diploid, sexual species C. inornatus. Prior to ovulation, females from both species display femalelike receptive behaviors. However, after ovulation, only parthenogenetic individuals display malelike mounting behavior. In all nuclei measured, we found larger TH-ir cells in the parthenogen, a difference consistent with species differences in ploidy. In contrast, species differences in the number of TH-ir cells were nucleus specific. In the preoptic area and anterior hypothalamus, parthenogens had fewer TH-ir cells than females of the sexual species. Reproductive state only affected TH-ir cell number in the substantia nigra pars compacta (SNpc), and C. uniparens individuals had more TH-ir cells after ovulation than when previtellogenic. Thus, species differences over the reproductive cycle in the SNpc are correlated with species differences in behavior, and it appears that the process of speciation may have produced a novel neural and behavioral phenotype in the parthenogen. PMID- 15281075 TI - Bilateral song production in domestic canaries. AB - We studied the mechanism of song production in the outbred common or domestic canary (Serinus canaria). The contribution that each side of the syrinx makes to song was investigated by observing the effect of unilaterally occluding the left or right primary bronchus, followed by section of the ipsilateral branch of the tracheosyringeal nerve. In other birds with a bilaterally intact vocal system we monitored airflow through each side of the syrinx, together with subsyringeal pressure, during spontaneous song. Song production by domestic canaries is not strongly lateralized as it is in the conspecific song-bred waterslager strain. Some syllables are produced entirely on the left or right side of the syrinx, whereas others contain sequential contributions from each side. Low fundamental frequencies are produced with the left syrinx and high frequencies by the right syrinx, increasing the frequency range of domestic canary song compared to that of the waterslager strain. Midrange frequencies can be generated by either side. Syllables at repetition rates below about 25 s(-1) were accompanied by minibreaths, which were usually bilateral. Unilateral minibreaths were typically on the left side. At higher syllable repetition rates, minibreaths were replaced by a respiratory pattern of pulsatile expiration. Our data show that strong unilateral dominance in song production, present in the waterslager strain, is not a trait of the species as a whole and that the pattern of song lateralization can be altered by selective breeding for particular song characteristics. PMID- 15281074 TI - Leech filamin and Tractin: markers for muscle development and nerve formation. AB - The Lan3-14 and Laz10-1 monoclonal antibodies recognize a 400 kDa antigen that is specifically expressed by all muscle cells in leech. We show that the antigen recognized by both antibodies is a member of the filamin family of actin binding proteins. Leech filamin has two calponin homology domains and 35 filamin/ABP repeat domains. In addition, we used the Laz10-1 antibody to characterize the development of the segmentally iterated dorsoventral flattener muscles. We demonstrate that the dorsoventral flattener muscle develops as three discrete bundles of myofibers and that CNS axons pioneering the DP nerve extend only along the middle bundle. Interestingly, the middle dorsoventral muscle anlage is associated with only non-neuronal expression of the L1-family cell adhesion molecule Tractin. This expression is transient and occurs at the precise developmental stages when DP nerve formation takes place. Based on these findings we propose that the middle dorsoventral muscle anlagen provides a substrate for early axonal outgrowth and nerve formation and that this function may be associated with differential expression of distinct cell adhesion molecules. PMID- 15281076 TI - Islet-1 expression in the developing chicken inner ear. AB - The cell types of the inner ear originate from the otic placode, a thickened layer of ectoderm adjacent to the developing hindbrain. The placode invaginates and forms the otic pit, which pinches off as a small vesicle called the otocyst. Presumptive cochleovestibular neurons delaminate from the anterior ventral part of the otocyst and form the cochleovestibular ganglion of the inner ear. Here we show that the LIM/homeodomain protein islet-1 is expressed in cells of the ventral part of the otic placode and that this ventral expression is maintained at the otic pit and the otocyst stages. Auditory and vestibular neurons originate from this islet-1-positive zone of the otocyst, and these neurons maintain islet 1 expression until adulthood. We also demonstrate that islet-1 becomes up regulated in the presumptive sensory epithelia of the inner ear in regions that are defined by the expression domains of BMP4. The up-regulation of islet-1 in developing inner ear hair and supporting cells is accompanied by down-regulation of Pax-2 in these cell types. Islet-1 expression in hair and supporting cells persists until early postnatal stages, when the transcriptional regulator is down regulated in hair cells. Our data is consistent with a role for islet-1 in differentiating inner ear neurons and sensory epithelia cells, perhaps in the specification of cellular subtypes in conjunction with other LIM/homeodomain proteins. PMID- 15281077 TI - Distribution of the mRNAs encoding the thyrotropin-releasing hormone (TRH) precursor and three TRH receptors in the brain and pituitary of Xenopus laevis: effect of background color adaptation on TRH and TRH receptor gene expression. AB - In amphibians, thyrotropin-releasing hormone (TRH) is a potent stimulator of alpha-melanotropin (alpha-MSH) secretion, so TRH plays a major role in the neuroendocrine regulation of skin-color adaptation. We have recently cloned a third type of TRH receptor in Xenopus laevis (xTRHR3) that has not yet been characterized in any other vertebrate species. In the present study, we have examined the distribution of the mRNAs encoding proTRH and the three receptor subtypes (xTRHR1, xTRHR2, and xTRHR3) in the frog CNS and pituitary, and we have investigated the effect of background color adaptation on the expression of these mRNAs. A good correlation was generally observed between the expression patterns of proTRH and xTRHR mRNAs. xTRHRs, including the novel receptor subtype xTRHR3, were widely expressed in the telencephalon and diencephalon, where two or even three xTRHR mRNAs were often simultaneously observed within the same brain structures. In the pituitary, xTRHR2 was expressed selectively in the distal lobe, and xTRHR3 was found exclusively in the intermediate lobe. Adaptation of frog skin to background illumination had no effect on the expression of proTRH and xTRHRs in the brain. In contrast, adaptation of the animals to a white background provoked an 18-fold increase in xTRHR3 mRNA concentration in the intermediate lobe of the pituitary. These data demonstrate that, in amphibians, the effect of TRH on alpha-MSH secretion is mediated through the novel receptor subtype xTRHR3. PMID- 15281078 TI - ERp29, a general endoplasmic reticulum marker, is highly expressed throughout the brain. AB - ERp29 is a recently discovered resident of the endoplasmic reticulum (ER) that is abundant in brain and most other mammalian tissues. Investigations of nonneural secretory tissues have implicated ERp29 in a major role producing export proteins, but a molecular activity remains wanting for this functional orphan. Intriguingly, ERp29 appears to be heavily utilized in the cerebellum, a brain region not conventionally regarded as neurosecretory. To elucidate this functional quandary, we used immunochemical approaches to characterize the regional, cellular, and subcellular distributions of ERp29 in rat brain. Immunohistochemistry revealed ubiquitous expression in neuronal and nonneuronal cells, with a distinctive variation in somatic ERp29 levels. Highly expressing cells were found in diverse locations, implying that ERp29 is not biased towards the cerebellum functionally. Using immunolocalization data mined from the literature, a proteomic profile was developed to assess the functional significance of ERp29's characteristic expression pattern. Surprisingly, ERp29 correlated poorly with classical markers of neurosecretion, but strongly with a variety of major membrane proteins. Together with immunogold localization of ERp29 to somatic ER, these observations led to a novel hypothesis that ERp29 is involved primarily in production of endomembrane proteins rather than proteins destined for export. This study establishes ERp29 as a general ER marker for brain cells and provides a stimulating clue about ERp29's enigmatic function. ERp29 appears to have broad significance for neural pathophysiology, given its ubiquitous distribution and prominence in brain over classical ER residents like BiP and protein disulfide isomerase. PMID- 15281079 TI - Fiber order of the normal and regenerated optic tract of the frog (Rana pipiens). AB - In the normal frog, axons from the peripheral retina arising at the temporal pole course superficially in the middle stream of the diencephalic optic tract. Axons from the nasal pole course in two streams running in the opposite margins of the tract, dorsonasal axons ventrally, ventronasal axons dorsally. Axons from the dorsal and ventral poles of the retina occupy the intervals between the aforementioned middle and marginal streams. Axons from more central regions of the retina tend to occupy deeper levels of the optic tract. The regenerated optic tract does not regain its normal organization, e.g., axons of peripheral nasal origin are spread out widely over the entire width of the tract. However, axons from the temporal pole of the retina do return approximately to their original location in the middle stream. The concentration of temporal axons in the middle stream of the optic tract after regeneration may now be understood in terms of the expression pattern of the ephrin-A class of receptor tyrosine kinase ligands in the cellular matrix of the optic tract. The ephrin-As, which have a repellent effect on growing temporal retinal axons, are concentrated in and along the margins of the diencephalic optic tract and essentially absent from its middle stream. It is proposed here that peripheral temporal axons may be forced into this middle region by their avoidance of the higher levels of ephrin-A expression in the tract margins. In contrast, the growth pattern of regenerating peripheral nasal axons would not be affected by the ephrin-A gradient in the optic tract. PMID- 15281080 TI - Expression of regulatory genes during differentiation of thalamic nuclei in mouse and monkey. AB - Expression patterns of genes implicated in development of the thalamus were examined in mice and monkeys, using in situ hybridization with RNA probes complementary to Cad6, Dlx1, Dlx2, Dlx5, Gbx2, Id2, and Lef1 cDNAs. Expression patterns were related to the evolving cytoarchitecture in mice at birth (P0) and in adulthood, and in fetal monkeys early and late in the period of gestation when thalamic nuclei are becoming histologically differentiated out of a series of pronuclear masses. At the earlier developmental stage, each gene was expressed in a pattern that appeared to be pronucleus-specific and maintained a nucleus specific pattern into adulthood, with the possible exception of Gbx2. Each gene displayed a unique expression pattern in the dorsal thalamus, ventral thalamus, and epithalamus, and no gene was expressed throughout all three divisions or in every nucleus of a division. With the exception of Dlx2, whose expression disappeared at the later time point, all continued to be expressed into adulthood at higher levels and with identical patterns. Despite late appearance of gamma aminobutyric acid (GABA)ergic cells in the dorsal lateral geniculate nucleus of mice, no Dlx genes, which promote formation of a GABAergic phenotype elsewhere, were detected in dorsal thalamus. Each thalamic nucleus was distinguished by expression of a combination of genes, and homologous nuclei in mouse and monkey exhibited the same combination. The presence of a centre median nucleus and four pulvinar nuclei in monkeys was marked by patterns of expression not found in mice. The centre median nucleus was marked by high expression of Id2, which was expressed only weakly in very few nuclei of mice. PMID- 15281081 TI - Laminar organization of the mouse dentate gyrus: insights from BETA2/Neuro D mutant mice. AB - The dentate gyrus of rodents is characterized by a highly laminar organization: above a compact granule cell layer, commissural/associational (C/A) fibers terminate on proximal granule cell dendrites and entorhinal fibers terminate on distal granule cell dendrites in a nonoverlapping manner. To gain insights into mechanisms that underlie the formation of this laminar structure, we studied mice deficient for BETA2/NeuroD, a basic helix-loop-helix transcription factor essential for granule cell differentiation. Anterograde tracing was used to label C/A and entorhinal fibers and combined with confocal double immunofluorescence for calbindin, calretinin, parvalbumin, and reelin to visualize putative target cells. The dentate gyrus of mutant mice contained only few granule cells, which formed a cap-like structure adjacent to area CA3. Despite the severe hypoplasia of the dentate gyrus, the remaining BETA2/NeuroD-deficient granule cells expressed mature markers, extended dendrites into the molecular layer, and extended mossy fibers into area CA3. Entorhinal and C/A fibers terminated in a nonoverlapping manner in the dendritic field overlying the rudiment. Entorhinal fibers terminated in the outermost portion of the dentate gyrus where they surrounded reelin-positive Cajal-Retzius cells, and C/A fibers terminated above and within the dentate rudiment. The laminar termination of C/A fibers was closest to normal in zones of the rudiment in which granule cells were densely packed. These data indicate that granule cells are able to differentiate in the absence of BETA2/NeuroD and suggest that the signals underlying the laminar anatomy of the dentate gyrus are present in the absence of most target cells. PMID- 15281082 TI - Genetic control of sensitivity to hippocampal cell death induced by kainic acid: a quantitative trait loci analysis. AB - Host genetic factors are likely to contribute to differences in individual susceptibility to seizure-induced excitotoxic neuronal damage. Similarly, inbred strains of mice differ in their susceptibility to the kainic acid (KA) model of seizure-induced cell death, but the genes responsible for the differences are not known. Here, we define the inheritance patterns of susceptibility to KA-induced neurodegeneration in the hippocampus by assessing 331 back-cross (N2) progeny of two inbred mouse strains, C57BL/6 and FVB/N, previously shown to display resistance and sensitivity to KA-induced cell death, respectively. Results of phenotypic analysis suggest that the difference in susceptibility between these two strains is conferred by a single dominant gene. Therefore, we used an N2 back cross between the inbred C57BL/6 and FVB/N strains for a genome-wide search for quantitative trait loci (QTLs), which are chromosomal sites containing genes influencing the magnitude of susceptibility. Genome-wide interval mapping in N2 progeny identified a locus on distal chromosome (Chr) 18 with a peak LOD score of 4.9 localized between D18Mit186 and D18Mit4 as having the strongest and most significant effect in this model. QTLs of minor effect were detected on Chr 15 (D15Mit174-D15Mit156) and Chr 4 (D4Mit264-D4Mit91), with peak LOD scores of 3.02 and 2.46, respectively. The three significant QTLs (Chrs 4, 15, 18) together account for nearly 25% of the trait variance for both genders combined. Reduced KA-induced cell death susceptibility was observed in a congenic strain in which the highly susceptible FVB/N strain carried putative resistance alleles from the C57BL/6 strain on Chr 18. PMID- 15281083 TI - The basic helix-loop-helix transcription factor neuroD is expressed in the rod lineage of the teleost retina. AB - Persistent rod genesis in the retinas of teleost fish was first described over 2 decades ago, but little is known regarding the underlying genetic and molecular mechanisms that govern this phenomenon. Because of its function in the developing mammalian retina and persistently mitotic adult tissues, we sought to characterize the cellular expression of the basic helix-loop-helix (bHLH) transcription factor neuroD in the persistently neurogenic retina of adult teleosts. We show here that, in the adult retina of the goldfish, neuroD is expressed by putative amacrine cells, nascent cones, and the mitotically active cells of the rod lineage. neuroD is the first gene shown to be expressed by rod precursors, the immediate antecedents of rod photoreceptors. In contrast to the vertebrate classes described previously, neuroD is not expressed in multipotent progenitors in the teleost retina. Combining neuroD in situ hybridizations with cell-cycle-specific markers suggests that, in rod precursors, neuroD expression is cell cycle specific. PMID- 15281084 TI - Chromatin remodeling and stem cell theory of relativity. AB - The field of stem cell biology is currently being redefined. Stem cell (hematopoietic and non-hematopoietic) differentiation has been considered hierarchical in nature, but recent data suggest that there is no progenitor/stem cell hierarchy, but rather a reversible continuum. The stem cell (hematopoietic and non-hematopoietic) phenotype, the total differentiation capacity (hematopoietic and non-hematopoietic), gene expression as well as other stem cell functional characteristics (homing, receptor and adhesion molecule expression) vary throughout a cell-cycle transit widely. This seems to be dependent on shifting chromatin and gene expression with cell-cycle transit. The published data on DNA methylation, histone acetylation, and also RNAi, the major regulators of gene expression, conjoins very well and provides an explanation for the major issues of stem cell biology. Those features of stem cells mentioned above can be rather difficult to apprehend when a classical hierarchy biology view is applied, but they become clear and easier to understand once they are correlated with the underlining epigenetic changes. We are entering a new era of stem cell biology the era of "chromatinomics." We are one step closer to the practical use of cellular therapy for degenerative diseases. PMID- 15281086 TI - Exogenous metalloporphyrins alter the organization and function of cultured neonatal rat heart cells via modulation of heme oxygenase activity. AB - Heme oxygenase (HO), the enzyme responsible for heme catabolism, has been associated with the function of both skeletal and smooth muscle cells and with protection of the heart against ischemia/reperfusion injury. Exposure of skeletal muscle cultures to heme, the physiological substrate for HO, has been shown to improve differentiation and aerobic metabolism. Little is known, however, about the roles that heme and heme metabolism play in cardiac muscle, and the present study was conducted to examine the effects of exogenous heme on cultured heart cells in the presence or absence of modulators of HO activity. Treatment of neonatal rat ventricular cells with heme resulted in increases in four key indicators: (1) the activity of metabolic enzymes, (2) the rate of spontaneous contraction, (3) the level of myosin heavy chain (MyHC) expressed, and (4) the amount of actin organized as filaments. Treatment with heme while metabolically inhibiting increased HO activity altered these effects such that: (1) increases in enzyme activities were attenuated, (2) spontaneous beating ceased, (3) the level of MyHC was reduced, and (4) the amount of filamentous actin was severely decreased to the point where myofibrils were no longer evident. These results suggest that heme and its catabolites act to modulate aspects of cardiac cell function and organization. PMID- 15281085 TI - Thyroid hormone stimulates osteoclast differentiation by a mechanism independent of RANKL-RANK interaction. AB - It is well known that thyroid hormone excess causes bone loss. However, the precise mechanism of bone loss by thyroid hormone still remains unclear. When T(3) was added to unfractionated bone cells after degeneration of pre-existent osteoclasts, T(3) (1 pM-100 nM) dose-dependently stimulated osteoclast-like cell formation, irrespective of the presence of indomethacin and IL-6 Ab. T(3) increased the expression of osteoprotegerin (OPG) messenger RNA (mRNA), but not of receptor activator of nuclear factor kappaB ligand (RANKL) in unfractionated bone cells, suggesting that the stimulatory effect of T(3) on osteoclast formation was not mediated by the RANKL/OPG system. We next examined the direct effect of T(3) on osteoclast precursors in the absence of osteoblasts, using hemopoietic blast cells derived from spleen cells. T(3) (1 pM-100 nM) dose dependently stimulated osteoclast-like cell formation from osteoclast precursors. OPG did not inhibit T(3)-induced osteoclast formation from osteoclast precursor cells. The polymerase chain reaction (PCR) product corresponding in size to the mouse T(3) receptor alpha1 cDNA was detected in osteoclast precursors from mouse hemopoietic blast cells as well as mouse heart and mouse osteoblastic cell line MC3T3-E1 cells, suggesting that T(3) directly stimulated osteoclast-like cell formation from osteoclast precursors in the absence of osteoblasts. Further, T(3) increased the expression of c-Fos mRNA at 15 min and 24 h and Fra-1 mRNA at 2 and 6 h in osteoclast precursors. Consistent with the increased expression of c-Fos mRNA observed by RT-PCR, the activation of c-Fos occurred in osteoclast precursor cells stimulated by T(3), while the activation of neither NF-kappaB nor MAPKs was observed by immunoblot analysis. Antisense oligodeoxynucleotides (as-ODN) complementary to c-Fos mRNA at 1 microM significantly inhibited T(3)-induced osteoclast-like cell formation from osteoclast precursors in the absence of stromal cells while sense-ODN did not affect T(3)-induced osteoclast-like cell formation. These results indicate that T(3) directly stimulates osteoclast differentiation at least in part by up-regulation of c-fos protein in osteoclast precursor cells. PMID- 15281088 TI - Apotransferrin is internalized and distributed in the same way as holotransferrin in K562 cells. AB - Transferrin (Tf), a naturally existing protein, has received considerable attention in the area of drug targeting since it is biodegradable, non-toxic, and non-immunogenic. The efficient cellular uptake of Tf shows it has potential in the delivery of anti-cancer drugs, proteins, and therapeutic genes into proliferating malignant cells that overexpress transferrin receptor (TfR). In human serum, about 30% of Tf exists in the iron-saturated form (Fe(2)-Tf) and the remainder exists as apotransferrin (apo-Tf). Understanding the uptake of apo-Tf by cells will provide key insights into studies on Tf-mediated drug delivery. In the present study, we investigated visually the transport of apo-Tf into K562 cells and its intracellular localization by laser-scanning confocal microscopy (LSCM) and flow cytometry analysis (FCA). It was found that, like Fe(2)-Tf, apo Tf can be taken up into the cells. The process is time- and temperature dependent, competitively inhibited by Fe(2)-Tf, and significantly abolished by pronase pretreatment. Visual evidence showed that the transport of apo-Tf into K562 cells is a TfR-mediated process. Furthermore, the investigations using optical-slicing technique demonstrated that the distribution of apo-Tf is similar to that of Fe(2)-Tf, both appearing in the perinuclear region in ball-in-bowl shape. PMID- 15281087 TI - MAP kinases differentially regulate the expression of macrophage hyperactivity after thermal injury. AB - Thermal injury increases the capacity of macrophages (Mphi) to produce various inflammatory mediators, (i.e., Mphi hyperactivity), which is believed to be involved in the development of subsequent immunosuppression, sepsis, and multiple organ failure. The signal transduction pathways involved in the expression of Mphi hyperactivity post-burn, however, remain to be clearly elucidated. To study this C57BL/6 female mice were subjected to a 25% TBSA burn and splenic Mphis were isolated 7 days later. LPS-stimulated inflammatory mediator production and MAPK expression (P38 ERK 1/2 and JNK) were determined. Burn injury increased LPS induced P38 MAPK, suppressed JNK activation and ERK 1/2 activation was unaltered. These changes in MAPK activation were paralleled by the increased production of PGE(2), TNF-alpha, IL-1beta, IL-6, and IL-10. Differential sensitivity to the inhibition of the MAPK pathways was observed with regard to the mediator evaluated and the presence or absence of burn injury. In general cytokine production in the burn group was in part resistant to the inhibition of a single MAPK pathway as compared with shams. Thus, burn injury increases cross-talk between the MAPKs pathways, suggesting that alterations MAPK activation and signal transduction contribute to the development Mphi hyperactivity post-injury. PMID- 15281089 TI - Microtubule disassembly induces cytoskeletal remodeling and lung vascular barrier dysfunction: role of Rho-dependent mechanisms. AB - Barrier dysfunction of pulmonary endothelial monolayer is associated with dramatic cytoskeletal reorganization, activation of actomyosin contractility, and gap formation. The linkage between the microtubule (MT) network and the contractile cytoskeleton has not been fully explored, however, clinical observations suggest that intravenous administration of anti-cancer drugs and MT inhibitors (such as the vinca alkaloids) can lead to the sudden development of pulmonary edema in breast cancer patients. In this study, we investigated the crosstalk between MT and actomyosin cytoskeleton and characterized specific molecular mechanisms of endothelial cells (EC) barrier dysfunction induced by MT inhibitor nocodazole (ND). Our results demonstrate that MT disassembly by ND induced rapid decreases in transendothelial electrical resistance (TER) and actin cytoskeletal remodeling, indicating EC barrier dysfunction. These effects involved ND-induced activation of Rho GTPase. Rho-mediated activation of its downstream target, Rho-kinase, induced phosphorylation of Rho-kinase effector EC MLC phosphatase (MYPT1) at Thr(696) and Thr(850) resulting in MYPT1 inactivation. Phosphatase inhibition leaded to accumulation of diphospho-MLC, which induced acto-myosin polymerization, stress fiber formation and gap formation. Inhibition of Rho-kinase by Y27632 abolished ND-induced MYPT1 phosphorylation, MLC phosphorylation, and stress fiber formation. In addition, MT preservation via the MT stabilizer paclitaxel, Rho inhibition (via C3 exotoxin, or dominant negative (DN)-Rho, or DN-Rho-kinase) attenuated ND-induced TER decreases, stress fiber formation and MLC phosphorylation. Collectively, our results demonstrate a leading role for Rho-dependent mechanisms in crosstalk between the MT and actomyosin cytoskeleton, and suggest Rho-kinase and MYPT1 as major Rho effectors mediating pulmonary EC barrier disruption in response to ND-induced MT disassembly. PMID- 15281090 TI - Apoptosis induced by interferon-alpha and antagonized by EGF is regulated by caspase-3-mediated cleavage of gelsolin in human epidermoid cancer cells. AB - We have previously reported that interferon-alpha (IFNalpha) induces apoptosis and EGF can antagonize this effect in human epidermoid cancer KB cells. Since apoptosis occurs together with cytoskeleton reorganization we have evaluated if IFNalpha and EGF could modulate cell remodeling in our experimental conditions. We have found that 48 h 1,000 IU/ml IFNalpha induced structural reorganization of stress fibers and membrane delocalization and partial capping of the actin severing protein gelsolin. The transfection of KB cells with both a wild type (WT) or a C-terminal truncated form of gelsolin caused overexpression of the protein and an increase of both the spontaneous and IFNalpha-induced apoptosis and cell cytoskeletal modifications. In fact, after 48 h of treatment IFNalpha induced 45% of apoptotic cell death in parental cells while an approximately 80% of cell population was apoptotic in transfected cells. These effects occurred together with an increase of the expression and consequent degradation of gelsolin. Again the addition of EGF to IFNalpha-treated transfected cells caused a recovery of the apoptosis. Notably, IFNalpha and EGF did not modify the expression of other molecules associated to cytoskeleton such as focal adhesion kinase and vinculin. In the same experimental conditions IFNalpha induced also gelsolin cleavage that occurred together with caspase-3 activation and release of cytochrome c. All these effects were antagonized by the exposure of IFNalpha treated KB to 10 nM EGF for the last 12 h. Moreover, the specific inhibition of caspase-3 with 20 microM DEVD completely abrogated apoptosis and gelsolin cleavage induced by IFNalpha. In conclusion, our data are the first demonstration that IFNalpha can induce morphological cell changes that are peculiar of apoptosis onset through the caspase-3-mediated cleavage of gelsolin. Furthermore, we have demonstrated that EGF is able to antagonize these effects through the inhibition of caspase-3 activation. PMID- 15281091 TI - Mitogenic signalling by B2 bradykinin receptor in epithelial breast cells. AB - The kinin peptides are released during inflammation and are amongst the most potent known mediators of vasodilatation, pain, and oedema. A role in the modulation or induction of healthy breast tissue growth has been postulated for tissue kallikrein present in human milk. Moreover, tissue kallikrein was found in malignant human breast tissue and bradykinin (BK) stimulates the proliferation of immortalised breast cancer cells. Aim of the present article was to investigate whether BK also exerts mitogenic activity in normal breast epithelial cells and partially characterise the signalling machinery involved. Results show that BK increased up to 2-fold the 24 h proliferation of breast epithelial cells in primary culture, and that the BK B2 receptor (not B1) inhibitor alone fully blocked the BK response. Intracellular effects of B2 stimulation were the following: (a) the increase of free intracellular Ca(2+) concentration by a mechanism dependent upon the phospholipase C (PLC) activity; (b) the cytosol-to membrane translocation of conventional (PKC)-alpha and -beta isozymes, novel PKC delta, -epsilon, and -eta isozymes; (c) the phosphorylation of the extracellular regulated kinase 1 and 2 (ERK1/2); and (d) the stimulation of the expression of c Fos protein. EGF, a well known stimulator of cell proliferation, regulated the proliferative response in human epithelial breast cells to the same extent of BK. The effects of BK on proliferation, ERK1/2 phosphorylation, and c-Fos expression were abolished by GF109203X, which inhibits PKC-delta isozyme. Conversely, Go6976, an inhibitor of PKC-alpha and -beta isozymes, and the 18-h treatment of cells with PMA, that led to the complete down-regulation of PKC-alpha, -beta, epsilon, and -eta, but not of PKC-delta, did not have any effect, thereby indicating that the PKC-delta mediates the mitogenic signalling of BK. Phosphoinositide 3-kinase (PI3K), tyrosine kinase of the epidermal growth factor receptor (EGFR), and mitogen activated protein kinase kinases (MEK) inhibitors were also tested. The results suggest that EGFR, PI3K, and ERK are required for the proliferative effects of BK. In addition, the BK induced cytosol-to-membrane translocation of PKC-delta was blocked by PI3K inhibition, suggesting that PI3K is upstream to PKC-delta. In conclusion, BK has mitogenic actions in cultured human epithelial breast cells; the activation of PKC-delta through B2 receptor acts in concert with ERK and PI3K pathways to induce cell proliferation. PMID- 15281092 TI - Targeted inhibition of the epidermal growth factor receptor-tyrosine kinase by ZD1839 ('Iressa') induces cell-cycle arrest and inhibits proliferation in prostate cancer cells. AB - The epidermal growth factor (EGF) plays a role in the development of prostate cancer, which becomes essential after androgen resistance has emerged. The EGF receptor (EGFR) is therefore a potential target for anticancer therapy. We evaluated the effects of ZD1839 ('Iressa'), an orally active EGFR-tyrosine kinase inhibitor, on prostate cancer cell lines. The effects of ZD1839 were evaluated on the anchorage dependent and independent growth of androgen-responsive (LNCaP) and androgen-independent (DU145 and PC3) cells by a cell proliferation assay, cell counting, and soft agar analysis. Flow cytometric analysis and Western blotting were used to assess the effects on the cell-cycle and on protein expression levels, respectively. ZD1839 caused a dose- and time-dependent growth inhibition in all three cell lines. A dose-dependent supra-additive increase in growth inhibition was observed when ZD1839 was combined with the antiandrogen flutamide or ionizing radiation (IR). The antiproliferative effect of ZD1839 was mainly cytostatic and associated with a block in the G(0)/G(1) phase of the cell-cycle, evident after about 12 h of treatment. In the DU145 cells this block was associated with an increase in expression of the CDK inhibitor p27(Kip1), both in the cytoplasmic and nuclear fractions. The increase in p27(Kip1) was not evident in the LNCaP and PC3 cells. No changes were observed in the expression of cyclin D1 protein. These results demonstrate the antiproliferative effects of ZD1839 on the growth of prostate cancer cells and suggest that inhibition of EGFR associated signal transduction pathway might represent a promising novel therapeutic strategy for the treatment of prostate cancer. PMID- 15281093 TI - N-acetylcysteine inhibits Na+ absorption across human nasal epithelial cells. AB - N-acetylcysteine (NAC) is a widely used mucolytic drug in patients with a variety of respiratory disorders. The mechanism of action is based on rupture of the disulfide bridges of the high molecular glycoproteins present in the mucus, resulting in smaller subunits of the glycoproteins and reduced viscosity of the mucus. Because Na(+) absorption regulates airway surface liquid volume and thus the efficiency of mucociliary clearance, we asked whether NAC affects the bioelectric properties of human nasal epithelial cells. A 24-h basolateral treatment with 10 mM of NAC decreased the transepithelial potential difference and short-circuit current (I(SC)) by 40%, and reduced the amiloride-sensitive current by 50%, without affecting the transepithelial resistance. After permeabilization of the basolateral membranes of cells with amphotericin B in the presence of a mucosal-to-serosal Na(+) gradient (135:25 mM), NAC inhibited 45% of the amiloride-sensitive current. The Na(+)-K(+)-ATPase pump activity and the basolateral K(+) conductance were not affected by NAC treatment. NAC did not alter total cell mRNA and protein levels of alpha-epithelial Na(+) channel (EnaC) subunit, but reduced abundance of alpha-ENaC subunits in the apical cell membrane as quantified by biotinylation. This effect can be ascribed to the sulphydryl (SH) group of NAC, since N-acetylserine and S-carboxymethyl-l-cysteine were ineffective. Given the importance of epithelial Na(+) channels in controlling the thin layer of fluid that covers the surface of the airways, the increase in the fluidity of the airway mucus following NAC treatment in vivo might be in part related to downregulation of Na(+) absorption and consequently water transport. PMID- 15281094 TI - Subcellular trafficking of exogenously expressed interferon-beta in Madin-Darby canine kidney cells. AB - We have recently demonstrated that when IFN-beta was exogenously expressed in epithelial cells, transiently expressed IFN-beta was predominantly secreted from the cell side to which the transfection was performed, while stably expressed one was almost equally secreted to the apical and basolateral sides. In the present study, we analyzed the subcellular transport of IFN-beta using confocal imaging with green fluorescent protein (GFP)-tagged IFN-beta in Madin-Darby canine kidney (MDCK) cells. Stably expressed and transiently expressed human IFN-beta (HuIFN beta)-GFPs were seen in upper regions of the nucleus. In stable HuIFN beta-GFP producing transformants, transiently expressed mouse IFN-beta (MuIFN-beta) was apparently co-localized with the bulk of the constitutive HuIFN beta-GFP proteins at TGN, and a significant quantity of them then appeared to pass into distinct post-TGN vesicles, accepting either type of IFN. Meanwhile, when cells were co transfected with both expression vectors, transiently expressed both IFNs tended to co-localize not only at TGN but in post-TGN vesicles. These results suggest that stably and transiently expressed IFN-betas, albeit co-localized at TGN, were transported through apparently discriminated post-TGN routes. PMID- 15281095 TI - Genes expressed in the human trabecular meshwork during pressure-induced homeostatic response. AB - Physiological pressure inside the eye is maintained by a resistance mechanism provided by the trabecular meshwork tissue. In most cases, prolonged, elevated pressure leads to an eye pathology characterized by retinal ganglion cell (RGC) degeneration, optic nerve damage, and non-remedial blindness. We are investigating the regulation of trabecular meshwork genes in response to elevated pressure. Using perfused organ cultures from postmortem human donors, we have previously demonstrated the presence of a homeostatic mechanism at 2-4 days of pressure insult (Borras et al. 2002, Invest Ophthalmol Vis Sci 43:33-40). Here, we sought to identify trabecular meshwork genes whose expression was altered during this homeostatic period. By macroarray hybridization, we compared the expression profiles of high-pressure (HP) and normal-pressure (NP) treated eyes from the same individual (n = 3 pairs). Our results identified 40 upregulated and 14 downregulated genes. The highest proportion of upregulated genes encoded proteins involved in signal transduction (32%). Among the potentially relevant genes, PIP 5K1C, VIP, tropomodulin, and MMP2 encoded mediators known to influence outflow resistance. Others encoded functions which are new for the trabecular meshwork, but which are intrinsic to unrelated tissues. These new mechanisms appear as they could be of benefit for trabecular meshwork function. Matrix Gla protein (MGP), perlecan, osteomodulin, and osteoblast-specific factor are essential in cartilage and bone physiology whereas spectrin and ICAM4 are specific for blood cells and crucial in maintaining their shape and adhesion. In addition, MGP transcripts were stimulated by extracellular calcium and downregulated by TGF-beta1. We propose that MGP might be an important player in the adaptive homeostatic mechanism by contributing to maintain a softer trabecular meshwork tissue and facilitate aqueous humor outflow. PMID- 15281096 TI - Protease nexin-1: a cellular serpin down-regulated by thrombin in rat aortic smooth muscle cells. AB - Protease nexin-1 (PN-1), a potent inhibitor of serine proteases, is present in vascular cells and forms complexes with thrombin, plasminogen activators, and plasmin. We examined the effect of thrombin on PN-1 expression by rat aortic smooth muscle cells (RASMCs). PN-1 expression was determined by measuring protein and mRNA levels, using respectively immunoblotting and semi-quantitative reverse transcriptase polymerase chain reaction (PCR). Thrombin down-regulated PN-1 expression in a dose- and time-dependent manner. This effect was mediated via the interaction of thrombin with its receptor protease activated receptor (PAR-1) since the peptide thrombin receptor activating peptide (TRAP) reduced PN-1 expression. PN-1 secreted by smooth muscle cells remained essentially associated to cell-surface glycosaminoglycans and was released from the cell surface by heparin. A lower amount of PN-1 was released by heparin from TRAP-stimulated versus unstimulated cells and correlated with a decreased capacity to inhibit thrombin. In addition, the ability to generate peri-cellular plasmin was increased in cells with a low PN-1 expression. Pre-treatment of smooth muscle cells with cycloheximide abolished the reduction of PN-1 expression by thrombin. Furthermore, conditioned media from thrombin-treated cells reproduced the effect of thrombin, suggesting that thrombin acted via the induction of auto/paracrine mediator(s). We observed that fibroblast growth factor-2 (FGF-2)-neutralizing antibodies abolished thrombin effect whereas FGF-2 reproduced it, indicating that FGF-2 is one of the involved mediator. Together, these results indicate that (i) PN-1 modulates the activity of endogenous and exogenous serine proteases in RASMCs, (ii) thrombin down-regulates PN-1 expression and thus may increase its own activity on cells. PMID- 15281097 TI - Temperature-sensitive polymer-conjugated IFN-gamma induces the expression of IDO mRNA and activity by fibroblasts populated in collagen gel (FPCG). AB - Indoleamine 2,3-dioxygenase (IDO) is an intracellular tryptophan-catabolizing enzyme possessing various immunosuppressive properties. Here, we report the use of this enzyme to suppress the proliferation of peripheral blood mononuclear cells (PBMC) co-cultured with IDO-expressing fibroblasts of an allogeneic skin substitute in vitro. Fetal foreskin fibroblasts populated within collagen gel (FPCG) were treated with interferon-gamma (IFN-gamma) conjugated with a temperature-sensitive polymer to induce the expression of IDO mRNA and protein. SDS-PAGE showed successful conjugation of IFN-gamma with the temperature sensitive polymer. Expression of IDO mRNA was evaluated by Northern analysis. IDO enzyme activity was evaluated by the measurement of kynurenine levels. The results of Northern blot analysis showed an induction of IDO mRNA expression when treated with polymer-conjugated IFN-gamma. Kynurenine levels, as a measure of IDO bioactivity, were significantly higher in IFN-gamma-treated fibroblasts than in controls (P < 0.001). In a lasting effect experiment, the expression of IDO mRNA in FPCG treated with polymer-conjugated IFN-gamma was significantly longer than in those treated with free (non-conjugated) IFN-gamma (P < 0.001). IFN-gamma radiolabeling showed a prolonged retention of IFN-gamma within collagen gel in its polymer-conjugated form, compared to its free form. Presence of IDO protein in FPCG was demonstrated by Western analysis even 16 days after removal of the conditioned medium (containing released IFN-gamma). To demonstrate the immunosuppressive effects of IDO on the proliferation of PBMC, IDO-expressing FPCG treated with polymer-conjugated IFN-gamma were co-cultured with PBMC for a period of 5 days. The results showed a significant reduction in proliferation of PBMC co-cultured with IFN-gamma-treated IDO-expressing fibroblasts, compared to those co-cultured with non-IDO-expressing fibroblasts (P < 0.001). The addition of an IDO inhibitor (1-methyl-D-tryptophan) reversed the suppressive effects of IDO on PBMC proliferation. In conclusion, IDO expression in FPCG suppresses the proliferation of immune cells in vitro. The use of a temperature-sensitive polymer further prolongs the effect of IFN-gamma on the expression of IDO. Therefore, modulating IDO levels in situ might be an alternative for prolonging the survival of skin allografts. PMID- 15281098 TI - Activity and expression of Xenopus laevis matrix metalloproteinases: identification of a novel role for the hormone prolactin in regulating collagenolysis in both amphibians and mammals. AB - Prolactin (PRL) has long been implicated in Xenopus metamorphosis as an anti metamorphic and/or juvenilizing hormone. Numerous studies showed that PRL could prevent effects of either endogenous or exogenous thyroid hormone (TH; T(3)). It has been shown that expression of matrix metalloproteinases (MMPs) is induced by TH during Xenopus metamorphosis. Direct in vivo evidence, however, for such anti TH effects by PRL with respect to MMPs has not been available for the early phase of Xenopus development or metamorphosis. To understand the functional role of PRL, we investigated effects of PRL on Xenopus collagenase-3 (XCL3) and collagenase-4 (XCL4) expression in a cultured Xenopus laevis cell line, XL-177. Northern blot analysis demonstrated that XCL3 and XCL4 expression were not detected in control or T(3)-treated cells, but were differentially induced by PRL in a dose- and time-dependent fashion. Moreover, treatment with IL-1alpha as well as phorbol myristate acetate (PMA), a protein kinase C (PKC) activator, or H8, a protein kinase A (PKA) inhibitor, augmented PRL-induced collagenase expression, suggesting that multiple protein kinase pathways and cytokines may participate in PRL-induced collagenase expression. Interestingly, XCL3 expression could be induced in XL-177 cells by T(3), but only when co-cultured with prometamorphic Xenopus tadpole tails (stage 54/55), suggesting that the tails secrete a required intermediate signaling molecule(s) for T(3)-induced XCL3 expression. Taken together, these data demonstrate that XCL3 and XCL4 can be differentially induced by PRL and T(3) and further suggest that PRL is a candidate regulator of TH independent collagenase expression during the organ/tissue remodeling which occurs in Xenopus development. PMID- 15281100 TI - The importance of bioseparations: giving credit where credit is due. PMID- 15281101 TI - Bioseparations. AB - Here we review key applications of separation technology in applied biology. We first sketch out the field as a whole, but then narrow our scope to the processing of fermentation products, particularly to high-value biologicals such as proteins and nucleotides. We go on to provide a qualitative overview describing the importance and general nature of this large field, major trends, and the strategies that have proven most fruitful in evolving effective separation and purification processes. We then give a detailed description of individual separations equipment and the principles governing their operation. We concentrate throughout on making the available literature accessible to the reader; we provide what is hoped to be a representative set of basic references. However, these references, in turn, include some that suggest promising new developments as well as a number of more specialized reviews. We hope that our overall result provides the reader with access to the most relevant literature. PMID- 15281102 TI - Optimized recovery of monoclonal antibodies from transgenic goat milk by microfiltration. AB - The Predictive Aggregate Transport Model for microfiltration is used in combination with optimum fluid mechanics and electrostatics to maximize recovery of a heterologous immunoglobulin (IgG) from transgenic goat milk. The optimization algorithm involved varying pH (6.8-9), transmembrane pressure (2-4.5 psi), milk feed concentration (1-2X), membrane module type (linear vs. helical design), and axial velocity (Reynolds number: 830-1170). Operation in the pressure-dependent regime at low uniform transmembrane pressures (approximately 2 psi) using permeate circulation in co-flow, at the pI of the protein (9 in this case) was used to increase IgG recovery from less than 1% to over 95%. Sodium dodecyl sulfate polyacrylamide gel electrophoresis and attenuated total reflection Fourier transform infrared spectroscopy of the microfiltration permeate samples confirmed that all the fat globules and most of the casein micelles were retained in the MF membrane whereas a large amount of the target IgG was transported through the membrane. Transmembrane pressure and hence permeation flux was kept low (approximately 15 lmh) to maximize IgG membrane transport and thus recovery, due to a sparse deposit on the membrane which facilitated high solute transport. Next, an analytical method was used to optimize the diafiltration process using the aggregate transport model, experimental target protein sieving coefficients and permeation flux (Baruah and Belfort, 2003). The methodology reported here should be generalizable to the recovery of target proteins found in other complex suspensions of biological origin using the microfiltration process. PMID- 15281103 TI - Optimization of ultrafiltration/diafiltration processes for partially bound impurities. AB - Ultrafiltration and diafiltration processes are used extensively for removal of a variety of small impurities from biological products. There has, however, been no experimental or theoretical analysis of the effects of impurity- product binding on the rate of impurity removal during these processes. Model calculations were performed to account for the effects of equilibrium binding between a small impurity and a large (retained) product on impurity clearance. Experiments were performed using D-tryptophan and bovine serum albumin as a model system. The results clearly demonstrate that binding interactions can dramatically reduce the rate of small impurity removal, leading to large increases in the required number of diavolumes. The optimal product concentration for performing the diafiltration shifts to lower product concentrations in the presence of strong binding interactions. Approximate analytical expressions for the impurity removal were developed which can provide a guide for the design and optimization of industrial ultrafiltration/diafiltration processes. PMID- 15281104 TI - Impact of engineering flow conditions on plasmid DNA yield and purity in chemical cell lysis operations. AB - Chemical lysis of bacterial cells using an alkaline solution containing a detergent may provide an efficient scalable means for selectively removing covalently closed circular plasmid DNA from high-molecular-weight contaminating cellular components including chromosomal DNA. In this article we assess the chemical lysis of E. coli cells by SDS in a NaOH solution and determine the impact of pH environment and shear on the supercoiled plasmid and chromosomal DNA obtained. Experiments using a range of plasmids from 6 kb to 113 kb determined that in an unfavorable alkaline environment, where the NaOH concentration during lysis is greater than 0.15 +/- 0.03 M (pH 12.9 +/- 0.2), irreversible denaturation of the supercoiled plasmid DNA occurs. The extent of denaturation is shown to increase with time of exposure and NaOH concentration. Experiments using stirred vessels show that, depending on NaOH concentration, moderate to high mixing rates are necessary to maximize plasmid yield. While NaOH concentration does not significantly affect chromosomal DNA contamination, a high NaOH concentration is necessary to ensure complete conversion of chromosomal DNA to single-stranded form. In a mechanically agitated lysis reactor the correct mixing strategy must balance the need for sufficient mixing to eliminate potential regions of high NaOH concentrations and the need to avoid excessive breakage of the shear sensitive chromosomal DNA. The effect of shear on chromosomal DNA is examined over a wide range of shear rates (10(1)-10(5) s(-1)) demonstrating that, while increasing shear leads to fragmentation of chromosomal DNA to smaller sizes, it does not lead to significantly increased chromosomal DNA contamination except at very high shear rates (about 10(4)-10(5) s(-1)). The consequences of these effects on the choice of lysis reactor and scale-up are discussed. PMID- 15281105 TI - Correlation of diafiltration sieving behavior of lysozyme-BSA mixtures with osmotic second virial cross-coefficients. AB - The role of protein-protein interactions in membrane separations of protein mixtures remains incompletely understood, largely due to the difficulty of characterizing protein self- and, especially, cross-association. Recently, a novel technique, cross-interaction chromatography, has been developed to measure weak protein cross-association in terms of the osmotic second virial cross coefficient. In this work the relationship between protein cross-association and the sieving behavior of lysozyme in the presence of BSA has been investigated. Sieving coefficients were measured using a stirred diafiltration cell over a range of pH and ionic strength, and a striking correlation between the lysozyme sieving and second virial cross-coefficients for BSA/lysozyme mixtures has been found: when the protein cross-interactions are most attractive (negative second virial cross-coefficient), the lysozyme sieving coefficients are lowest, and vice versa. The correlation between the sieving and second virial cross-coefficients may be due to the physically similar environments in the chromatography and filtration experiments since one protein is passed through a concentrated region of the second protein either immobilized on the column or accumulated at the membrane surface, and the migration rate of the mobile protein in both cases is influenced by protein cross-association. This study represents the first time that molecular interactions in binary mixtures have been related directly to filtration behavior, and may provide a useful approach to optimize the separation of other binary protein mixtures. PMID- 15281106 TI - Superparamagnetic adsorbents for high-gradient magnetic fishing of lectins out of legume extracts. AB - This work presents the development, testing, and application in high-gradient magnetic fishing of superparamagnetic supports for adsorption of lectins. Various approaches were examined to produce affinity, mixed mode, and hydrophobic charge induction type adsorbents. In clean monocomponent systems affinity supports created by direct attachment of glucose or maltose to amine-terminated iron oxide particles could bind concanavalin A at levels of up to approximately 280 mg g(-1) support with high affinity ( approximately 1 microM dissociation constants). However, the best performance was delivered by adsorbents featuring coupled tentacular dextran chains displaying a maximum binding capacity of 238 mg g(-1) and a dissociation constant of 0.13 microM. Adsorbents derivatized with mixed mode or hydrophobic charge induction ligands likewise demonstrated very high capacities for both concanavalin A and Lens culinaris agglutinin (> or = 250 mg g(-1)) with dissociation constants in the micromolar range, though neither of these systems showed any selectivity for lectins in leguminous extracts. When the affinity supports were applied to carbohydrate containing legume extracts only the dextran-linked adsorbents supplied sufficient competition to dissolved sugars to selectively bind concanavalin A in an extract of jack beans. The dextran linked supports were employed in a high-gradient magnetic fishing experiment, in which concanavalin A was purified to near homogeneity from a crude, unclarified extract of jack beans. PMID- 15281107 TI - Compatibility of column inlet and adsorbent designs for processing of corn endosperm extract by expanded bed adsorption. AB - Corn has emerged as a viable host for expression of recombinant proteins; targeted expression to the endosperm has received particular attention. The protein extracts from corn endosperm differ from those of traditional hosts in regard to the nature of residual solids and extracted matrix contaminants. Each of these differences presents reasons for considering expanded bed adsorption for product capture and new considerations for limitations of the method. In this work three inlet-flow distribution devices (mesh, glass ballotini, and localized mixing) and six adsorbents with different physical (size and density), chemical (ligand), and base matrix properties were evaluated to determine conditions compatible with processing of crude corn endosperm extract by expanded bed adsorption. Of the inlet devices evaluated, the design with localized mixing at the inlet (as produced commercially by UpFront Chromatography A/S, Copenhagen, DK) allowed solids up to 550 microm into the column without clogging for all flow rates evaluated. A mesh at the inlet with size restriction of either 50 microm or 80 microm became clogged with very small corn particles (< 44 microm). When glass ballotini was used, large particles (550 microm) passed through for high flow rates (570 cm/h), but even small (< 44 microm) particles became trapped at a lower flow rate (180 cm/h). The physical and chemical properties of the resin determined whether solids could be eluted. The denser UpFront adsorbents allowed for complete elution of larger and more concentrated corn solids than the currently available Amersham Streamline adsorbents (Amersham Biosciences, Piscataway, NJ) as a result of the former's higher flow rate for the desired 2x expansion (570 cm/h for UpFront vs. 180 cm/h for Streamline). All corn solids < 162 microm eluted through nonderivatized UpFront resin. Larger corn solids began to accumulate due to their elevated sedimentation velocities. Feeds of < 44 microm solids at 0.45% and 2.0% dry weight successfully eluted through ion exchange adsorbents (DEAE and SP) from UpFront. However, significant accumulation occurred when the solids size increased to a feed of < 96 microm solids, thus indicating a weak interaction between corn solids and both forms of ion exchange ligands. Expanded beds operated with Streamline ion exchange adsorbents (DEAE and SP) did not allow full elution of corn solids of < 44 microm. A hyperdiffuse style EBA resin produced by Biosepra (Ciphergen Biosystems, Fremont, CA) with CM functionality showed a severe interaction with corn solids that collapsed the expanded bed and could not be eliminated with elevated flow rates or higher salt concentration. PMID- 15281108 TI - The influence of biomass on the hydrodynamic behavior and stability of expanded beds. AB - Expanded bed adsorption is an innovative chromatographic technology that allows the introduction of particle-containing feedstock without the risk of blocking the bed. Provided a perfectly classified fluidized bed (termed expanded bed) is formed in the crude feedstock and the biomass is not influencing protein transport towards the adsorbent surface, a sorption performance comparable to packed beds is found. The influence of biomass on the hydrodynamic stability of expanded beds is essential and was investigated systematically in this article. Residence-time distribution analyses were performed using model systems and a yeast suspension under various fluid-phase conditions. It is demonstrated that three factors (biomass/adsorbent interactions, biomass concentration, and flow rate) play an interdependent role disturbing the classified fluidization of an expanded bed. A clear correlation between the degree of aggregative fluidization- obtained by PDE modeling of RTD data--and the expansion behavior of the fluidized bed has been found. Thus, combining three analytical methods, namely cell transmission index analysis, expansion analysis, and RTD analysis provides a solid base for understanding and control of the fluidization behavior and thus further process design during the initial phase of process development. PMID- 15281109 TI - Use of dimensionless residence time to study variations in breakthrough behaviour in expanded beds formed from varied particle size distributions. AB - This work demonstrates an experimental method for studying breakthrough behaviour in expanded beds. The behaviour of beds made with differently sized particles were studied at varying flowrates. The use of a dimensionless residence time measurement allowed a more valid comparison of breakthrough characteristics in expanded bed operation by compensating for the changes in bed volume that occur during expansion. We demonstrate that bed breakthrough behaviour can be compared directly even when the beds contain different-sized particles and hence have different expanded volumes. By utilising this concept we demonstrate that, in the case of the Alcohol Dehydrogenase (ADH) / STREAMLINE Phenyl system used here, there was little or no variation in ADH breakthrough behaviour between beds of differently sized particles operating at flowrates above 100 cm/h. This suggests that the higher specific surface area and hence binding capacity of smaller particles is negated in this case due to mass transfer limitations and the increase in system void volume even at normal operating flowrates of 200-300 cm/h. PMID- 15281110 TI - Evaluation of selectivity changes in HIC systems using a preferential interaction based analysis. AB - It is well established that salt enhances the interaction between solutes (e.g., proteins, displacers) and the weak hydrophobic ligands in hydrophobic interaction chromatography (HIC) and that various salts (e.g., kosmotropes, chaotropes, and neutral) have different effects on protein retention. In this article, the solute affinity in kosmotropic, chaotropic, and neutral mobile phases are compared and the selectivity of solutes in the presence of these salts is examined. Since solute binding in HIC systems is driven by the release of water molecules, the total number of released water molecules in the presence of various types of salts was calculated using the preferential interaction theory. Chromatographic retention times and selectivity reversals of both proteins and displacers were found to be consistent with the total number of released water molecules. Finally, the solute surface hydrophobicity was also found to have a significant effect on its retention in HIC systems. PMID- 15281111 TI - Performance and characterization of a nanophased porous hydroxyapatite for protein chromatography. AB - Nanophased porous hydroxyapatite beads with particle diameters of 25 microm and 30 microm intended for use in protein and biomolecule separation are characterized with respect to chromatographic characteristics. These particles were produced from a hydroxyapatite gel by a controlled spray process yielding microspheres containing hydroxyapatite nanocrystals. By calcification of the microspheres, nanophased porous hydroxyapatite beads were obtained. As a reference material, ceramic hydroxyapatite Types I and II with a particle diameter of 40 microm was chosen. SEM pictures show that the surface of the nanophased hydroxyapatite is very rough compared to ceramic hydroxyapatite Types I and Type II. The calcium-to-phosphorous ratio of this nanophased hydroxyapatite is 1.6, which is slightly below the theoretical ratio of 1.67 of pure hydroxyapatite. The porosity is greater than 60%. An IgG binding capacity of 60.7 mg/ml for Bio-Rad Type I and 36.0 mg/ml for Type II, 42.0 mg/ml for the nanophased material with 25 microm and 19.7 mg/ml for the nanophased material with 30 microm were observed. The nanophased material with 30 microm had the lowest mass transfer resistancy as indicated by the dependency of the dynamic binding capacity on velocity. It is assumed that the mass transport properties are characterized by a low particle diffusion resistancy or by slight intraparticle convection. The material also showed high selectivity for IgG. When culture supernatant with 5% FCS containing 3 mg/ml was loaded, pure IgG could be eluted by linear gradient with increasing sodium phosphate concentration. This nanophased material comprises a novel stationary phase for IgG separation. PMID- 15281112 TI - Chromatofocusing of peptides and proteins using linear pH gradients formed on strong ion-exchange adsorbents. AB - Although it is commonly believed that a column packing used for chromatofocusing must have an "even" buffering capacity in order to produce a linear pH gradient, it is demonstrated here that linear pH gradients suitable for chromatofocusing can be produced on a column packing having a minimal buffering capacity. In particular, if either a strong-acid cation-exchange column packing or a strong base anion-exchange column packing is presaturated with either a weak acid titrated with a strong base, or a weak base titrated with a strong acid, respectively, to the initial pH, then a linear or nearly linear pH gradient can be formed using a polyampholyte elution buffer by taking advantage of the presence of small quantities of weak-acid or weak-base functional groups that generally exist on these types of column packings. Experimental and theoretical studies are used to demonstrate that such systems have potential advantages over traditional chromatofocusing methods in terms of the speed of the separation, the resolution achieved, and the range of applications possible. Among other techniques described, a method for separating tryptic peptides using chromatofocusing and a strong-acid cation-exchange column packing is demonstrated to be a useful alternative to capillary isoelectric focusing and ion-exchange chromatography using a salt gradient for this purpose. PMID- 15281113 TI - Hydrophobic interaction chromatography selectivity changes among three stable proteins: conformation does not play a major role. AB - Interesting retention and selectivity changes have been noted for a number of proteins in hydrophobic interaction chromatography (HIC). In this study, we investigated the degree to which conformational changes may be responsible for selectivity changes of stable proteins. Hydrogen-deuterium isotope exchange detected by mass spectrometry was used to investigate changes in solvent accessibility during adsorption on HIC media. Lysozyme was determined to exhibit EX2 hydrogen exchange kinetics both in solution and adsorbed to Butyl Sepharose 4 Fast Flow and Phenyl Sepharose 6 Fast Flow high sub surfaces. A small, but significant, increase in solvent accessibility was observed upon adsorption. Similar approaches were used to analyze solvent accessibility of three stable proteins with melting temperatures above 50 degrees C exhibiting significant selectivity changes on Butyl Sepharose and Toyopearl Butyl 650M. While all three proteins (lysozyme, chymotrypsinogen A, and ovalbumin) exhibited enhanced exchange while adsorbed, no differences in solvent accessibility on the different adsorbents were observed. More detailed studies of lysozyme showed no significant changes in labeling prior or during elution. These results demonstrate that HIC surfaces examined here do not dramatically alter the structure of these stable proteins and that differences in conformation are not responsible for the selectivity changes observed. Thus, other factors such as different preferred binding orientations or variations between the media pore structure, size, and/or surface chemistry must be responsible. PMID- 15281114 TI - Development and validation of an affinity chromatography step using a peptide ligand for cGMP production of factor VIII. AB - An affinity chromatography step was developed for purification of recombinant B Domain Deleted Factor VIII (BDDrFVIII) using a peptide ligand selected from a phage display library. The peptide library had variegated residues, contained both within a disulfide bond-constrained ring and flanking the ring. The peptide ligand binds to BDDrFVIII with a dissociation constant of approximately 1 microM both in free solution and when immobilized on a chromatographic resin. The peptide is chemically synthesized and the affinity resin is produced by coupling the peptide to an agarose matrix preactivated with N-hydroxysuccinimide. Coupling conditions were optimized to give consistent and complete ligand incorporation and validated with a robustness study that tested various combinations of processing limits. The peptide affinity chromatographic operation employs conditions very similar to an immunoaffinity chromatography step currently in use for BDDrFVIII manufacture. The process step provides excellent recovery of BDDrFVIII from a complex feed stream and reduces host cell protein and DNA by 3-4 logs. Process validation studies established resin reuse over 26 cycles without changes in product recovery or purity. A robustness study using a factorial design was performed and showed that the step was insensitive to small changes in process conditions that represent normal variation in commercial manufacturing. A scaled-down model of the process step was qualified and used for virus removal studies. A validation package addressing the safety of the leached peptide included leaching rate measurements under process conditions, testing of peptide levels in product pools, demonstration of robust removal downstream by spiking studies, end product testing, and toxicological profiling of the ligand. The peptide ligand affinity step was scaled up for cGMP production of BDDrFVIII for clinical trials. PMID- 15281115 TI - Preparative protein purification on underivatized silica. AB - This article describes the use of underivatized silica gel as a preparative stationary phase for process purification of proteins. Although silica has been frequently used as a stationary phase backbone matrix, direct adsorption of proteins on underivatized silica has not been widely exploited for industrial applications. In this study an effort was made to fundamentally understand the interaction mechanisms between a protein and silica surface by using several proteins with a wide range of isoelectric points (pIs) and surface hydrophobicity. Interactions in silica were found to be largely dominated by a combination of ionic and hydrophobic forces. Accordingly, a predictive model was derived for describing linear retention of proteins on silica. Finally, a case study is described investigating the role of silica in an industrial purification process. It was found that the integration of the two modes of interaction confers silica with a unique selectivity that can be very effectively utilized in downstream bioprocessing. PMID- 15281116 TI - Purification of antibody and antibody-fragment from E. coli homogenate using 6,9 diamino-2-ethoxyacridine lactate as precipitation agent. AB - To obtain a more efficient purification process for antibody fragments from an Escherichia coli homogenate, the precipitant, Ethodin (6,9-diamino-2 ethoxyacridine lactate) was introduced to the homogenate. By adding the precipitant a drastic reduction of host cell protein was obtained. The majority of the proteins were recovered in a precipitate with the cell debris, while the antibody or antibody-fragment was recovered in the clarified supernatant. In addition, DNA was also efficiently precipitated when using Ethodin as a precipitation agent. The improved purity of the clarified extract obtained by using the precipitant allows for the use of smaller chromatography columns and may reduce the number of chromatographic steps required in the recovery process. The effect of Ethodin concentration, pH, temperature, and conductivity were investigated. The investigation was performed on two different antibody fragments, e.g., F(ab')(2) molecules and a full-length antibody produced in E. coli. The two F(ab')(2) proteins were F(ab')(2)A and F(ab')(2)B, which have a similar molecular mass (100 kDa) but different isoelectric points (pIs), i.e., 8.9 and 7.5, respectively. The full-length antibody, Ab (the full IgG form of F(ab')(2)B) has a pI of 7.8 and molecular mass of 150 kDa. The investigation showed that the highest purification factors were obtained at neutral pH, low conductivity, and Ethodin concentrations of 0.6%. PMID- 15281117 TI - Potential folding-function interrelationship in proteins. AB - The possibility is addressed that protein folding and function may be related via regions that are critical for both folding and function. This approach is based on the building blocks folding model that describes protein folding as binding events of conformationally fluctuating building blocks. Within these, we identify building block fragments that are critical for achieving the native fold. A library of such critical building blocks (CBBs) is constructed. Then, it is asked whether the functionally important residues fall in these CBB fragments. We find that for over two-thirds of the proteins in our library with available functional information, the catalytic or binding site residues lie within the CBB regions. From the evolutionary standpoint, a folding-function relationship is advantageous, since the need to guard against mutations is limited to one region. Furthermore, conformationally similar CBBs are found in globally unrelated proteins with different functions. Hence, substituting CBBs may lead to designed proteins with altered functions. We further find that the CBBs in our library are conformationally unstable. PMID- 15281118 TI - Statistical sequence analyses of G-protein-coupled receptors: structural and functional characteristics viewed with periodicities of entropy, hydrophobicity, and volume. AB - G-protein-coupled receptors (GPCRs) play a crucial role in signal transduction and receive a wide variety of ligands. GPCRs are a major target in drug design, as nearly 50% of all contemporary medicines act on GPCRs. GPCRs are membrane proteins possessing a common structural feature, seven transmembrane helices. In order to design an effective drug to act on a GPCR, knowledge of the three dimensional (3D) structure of the target GPCR is indispensable. However, as GPCRs are membrane bound, their 3D structures are difficult to obtain. Thus we conducted statistical sequence analyses to find information about 3D structure and ligand binding using the receptors' primary sequences. We present statistical sequence analyses of 270 human GPCRs with regard to entropy (Shannon entropy in sequence alignment), hydrophobicity and volume, which are associated with the alpha-helical periodicity of the accessibility to the surrounding lipid. We found periodicity such that the phase changes once in the middle of each transmembrane region, both in the entropy plot and in the hydrophobicity plot. The phase shift in the entropy plot reflects the variety of ligands and the generality of the mechanism of signal transduction. The two periodic regions in the hydrophobicity plot indicate the regions facing the hydrophobic lipid chain and the polar phospholipid headgroup. We also found a simple periodicity in the plot of volume deviation, which suggests conservation of the stable structural packing among the transmembrane helices. PMID- 15281119 TI - Predictions of protein flexibility: first-order measures. AB - The normal modes of a molecule are utilized, in conjunction with classical conformal vector field theory, to define a function that measures the capability of the molecule to deform at each of its residues. An efficient algorithm is presented to calculate the local chain deformability from the set of normal modes of vibration. This is done by considering each mode as an off-grid sample of a deformation vector field. Predictions of deformability are compared with experimental data in the form of dihedral angle differences between two conformations of ten kinases by using a modified correlation function. Deformability calculations correlate well with experimental results and validate the applicability of this method to protein flexibility predictions. PMID- 15281120 TI - Protein unfolding at interfaces: slow dynamics of alpha-helix to beta-sheet transition. AB - A two-phase sequential dynamic change in the secondary structure of hen egg lysozyme (Lys) adsorbed on solid substrates was observed. The first phase involved fast conversion of alpha-helix to random/turns (within the first minute or at very low coverage or high substrate wettability) with no perceptible change in beta-sheet content. The second phase (1-1200 min), however, involved a relatively slow conversion from alpha-helix to beta-sheet without a noticeable change in random/turns. An important finding of this work is that the concentration of lysozyme in the adsorbed state has a substantial effect on the fractional content of secondary structures. Attenuated total reflection Fourier transform infrared (ATR/FTIR) spectroscopy, along with a newly-developed optimization algorithm for predicting the content of secondary structure motifs, was used to correlate the secondary structure and the amount of adsorbed lysozyme with the surface wettability of six different flat nanoporous substrates. Although three independent variables, surface wettability, solution concentration and time for adsorption, were used to follow the fractional structural changes of lysozyme, the results were all normalized onto a single plot with the amount adsorbed as the universal independent variable. Consequently, lateral interactions among proteins likely drive the transition process. Direct intermolecular force adhesion measurements between lysozyme and different functionalized self-assembled alkanethiol monolayers confirm that hydrophobic surfaces interact strongly with proteins. The lysozyme-unfolding pathway during early adsorption appears to be similar to that predicted by published molecular modeling results. PMID- 15281121 TI - Protein contact prediction using patterns of correlation. AB - We describe a new method for using neural networks to predict residue contact pairs in a protein. The main inputs to the neural network are a set of 25 measures of correlated mutation between all pairs of residues in two "windows" of size 5 centered on the residues of interest. While the individual pair-wise correlations are a relatively weak predictor of contact, by training the network on windows of correlation the accuracy of prediction is significantly improved. The neural network is trained on a set of 100 proteins and then tested on a disjoint set of 1033 proteins of known structure. An average predictive accuracy of 21.7% is obtained taking the best L/2 predictions for each protein, where L is the sequence length. Taking the best L/10 predictions gives an average accuracy of 30.7%. The predictor is also tested on a set of 59 proteins from the CASP5 experiment. The accuracy is found to be relatively consistent across different sequence lengths, but to vary widely according to the secondary structure. Predictive accuracy is also found to improve by using multiple sequence alignments containing many sequences to calculate the correlations. PMID- 15281122 TI - Structural components of SCAN-domain dimerizations. AB - The SCAN or leucine-rich domain has been characterized as a highly conserved sequence in zinc finger transcription factors that mediates selective dimer formation between SCAN-domain-containing proteins. In order to accommodate various SCAN-domain sequence features, a minimal functional folding unit was defined on the premise of proper structural folding and biochemical binding. The 58-amino acid minimal functional units derived from each of four SCAN-domain protein families were subjected to a three-dimensional position-specific scoring matrix (3D-PSSM) and ungapped threading analysis. The resulting fold prediction represented the SCAN-domain's minimal functional unit as a bundle of three alpha helices folded to a core structure. In addition, the minimal functional folding unit biochemically retained the selective dimerization properties of the native proteins. In order to elucidate the structural components within the SCAN-domain that engage in binding interactions, we attempted to correlate the physicochemical helix properties, as represented by a hydropathy profile, with the experimental dimerization selectivities. The amino-terminal helix revealed the highest diversity measure among the three helices of the minimal functional unit and is therefore likely to offer critical surface-exposed binding residues. Indeed, by interchanging the amino-terminal helix between SCAN-domains without alteration of their structural frames consisting of conserved hydrophobic residues, a modulation of binding preferences was demonstrated. The minimal functional folding unit of SCAN-domains may therefore contain within the amino terminal alpha helix structural components that determine selective dimerization patterns and combinatorial control of transcription factors. PMID- 15281123 TI - Backbone-backbone geometry of tertiary contacts between alpha-helices. AB - Knowledge about how secondary-structure elements combine together to form the tertiary structure is crucial for understanding protein folding. We have examined packing solutions for alpha-helices by performing a crystal survey of the underlying backbone-backbone inter-geometry of tertiary contacts. The information content is different from, and complementary to, the results of side-chain to side-chain crystal surveys and studies of helix-helix-crossing angles. Six geometry descriptors were recorded from each tertiary contact in nonredundant data sets of globular and transmembrane proteins. The descriptors included distances, angles, and dihedral angles that together describe completely the underlying geometry of each contact. From the results it is possible to identify differences in the geometry requirements of tertiary contacts between alpha helices in general and transmembrane alpha-helices. The differences become more apparent when the correlation between the different geometry descriptors is analyzed. The results are compared with those of other types of secondary structure. Finally, an investigation of how the geometry of tertiary contacts changes with the amino-acid types involved in the contact is performed using multivariate techniques. The results of this study provide a well-defined overview of the underlying structural framework of tertiary contacts between alpha-helices, in both globular and TM environments, that will have valuable implications for predicting protein tertiary structure. PMID- 15281124 TI - Prediction of protein tertiary structure using PROFESY, a novel method based on fragment assembly and conformational space annealing. AB - A novel method for ab initio prediction of protein tertiary structures, PROFESY (PROFile Enumerating SYstem), is proposed. This method utilizes the secondary structure prediction information of a query sequence and the fragment assembly procedure based on global optimization. Fifteen-residue-long fragment libraries are constructed using the secondary structure prediction method PREDICT, and fragments in these libraries are assembled to generate full-length chains of a query protein. Tertiary structures of 50 to 100 conformations are obtained by minimizing an energy function for proteins, using the conformational space annealing method that enables one to sample diverse low-lying local minima of the energy. We apply PROFESY for benchmark tests to proteins with known structures to demonstrate its feasibility. In addition, we participated in CASP5 and applied PROFESY to four new-fold targets for blind prediction. The results are quite promising, despite the fact that PROFESY was in its early stages of development. In particular, PROFESY successfully provided us the best model-one structure for the target T0161. PMID- 15281125 TI - Markovian Backbone Negentropies: Molecular descriptors for protein research. I. Predicting protein stability in Arc repressor mutants. AB - As more and more protein structures are determined and applied to drug manufacture, there is increasing interest in studying their stability. In this sense, developing novel computational methods to predict and study protein stability in relation to their amino acid sequences has become a significant goal in applied Proteomics. In the study described here, Markovian Backbone Negentropies (MBN) have been introduced in order to model the effect on protein stability of a complete set of alanine substitutions in the Arc repressor. A total of 53 proteins were studied by means of Linear Discriminant Analysis using MBN as molecular descriptors. MBN are molecular descriptors based on a Markov chain model of electron delocalization throughout the protein backbone. The model correctly classified 43 out of 53 (81.13%) proteins according to their thermal stability. More specifically, the model classified 20/28 (71.4%) proteins with near wild-type stability and 23/25 (92%) proteins with reduced stability. Moreover, the model presented a good Mathew's regression coefficient of 0.643. Validation of the model was carried out by several Jackknife procedures. The method compares favorably with surface-dependent and thermodynamic parameter stability scoring functions. For instance, the D-FIRE potential classification function shows a level of good classification of 76.9%. On the other hand, surface, volume, logP, and molar refractivity show accuracies of 70.7, 62.3, 59.0, and 60.0%, respectively. PMID- 15281126 TI - Quantum descriptors for biological macromolecules from linear-scaling electronic structure methods. AB - The characterization of electrostatic and chemical properties at the surface of biological macromolecules is of interest in elucidating the fundamental biological structure-function relationships as well as in problems of rational drug design. This paper presents a set of macromolecular quantum descriptors for the characterization of biological macromolecules in solution that can be obtained with modest computational cost from linear-scaling semi-empirical quantum/solvation methods. The descriptors discussed include: solvent-polarized electrostatic surface potential maps, equilibrated atomic charges, Fukui reactivity indices, approximate local hardness maps, and relative proton potentials. These properties are applied to study the conformational dependence of the electrostatic surface potential of the solvated phosphate-binding protein mutant (T141D), the regioselectivity of the zinc finger domains of HIV-1 nucleocapsid (NC) protein, and the order of pKa values of acidic residues in turkey ovomucoid third domain (OMTKY3) and of the zinc-binding residues in the carboxyl terminal zinc finger of NC. In all cases, insight beyond that obtainable from purely classical models is gained and can be used to rationalize the experimental observations. The macromolecular quantum descriptors presented here greatly extend the arsenal of tools for macromolecular characterization and offer promise in applications to modern structure-based drug design. PMID- 15281127 TI - Constant-pH molecular dynamics using continuous titration coordinates. AB - In this work, we explore the question of whether pK(a) calculations based on a microscopic description of the protein and a macroscopic description of the solvent can be implemented to examine conformationally dependent proton shifts in proteins. To this end, we introduce a new method for performing constant-pH molecular dynamics (PHMD) simulations utilizing the generalized Born implicit solvent model. This approach employs an extended Hamiltonian in which continuous titration coordinates propagate simultaneously with the atomic motions of the system. The values adopted by these coordinates are modulated by potentials of mean force of isolated titratable model groups and the pH to control the proton occupation at particular sites in the polypeptide. Our results for four different proteins yield an absolute average error of approximately 1.6 pK units, and point to the role that thermally driven relaxation of the protein environment in the vicinity of titrating groups plays in modulating the local pK(a), thereby influencing the observed pK1/2 values. While the accuracy of our method is not yet equivalent to methods that obtain pK1/2 values through the ad hoc scaling of electrostatics, the present approach and constant pH methods in general provide a useful framework for studying pH-dependent phenomena. Further work to improve our model to approach quantitative agreement with experiment is outlined. PMID- 15281128 TI - Accurate prediction of solvent accessibility using neural networks-based regression. AB - Accurate prediction of relative solvent accessibilities (RSAs) of amino acid residues in proteins may be used to facilitate protein structure prediction and functional annotation. Toward that goal we developed a novel method for improved prediction of RSAs. Contrary to other machine learning-based methods from the literature, we do not impose a classification problem with arbitrary boundaries between the classes. Instead, we seek a continuous approximation of the real value RSA using nonlinear regression, with several feed forward and recurrent neural networks, which are then combined into a consensus predictor. A set of 860 protein structures derived from the PFAM database was used for training, whereas validation of the results was carefully performed on several nonredundant control sets comprising a total of 603 structures derived from new Protein Data Bank structures and had no homology to proteins included in the training. Two classes of alternative predictors were developed for comparison with the regression-based approach: one based on the standard classification approach and the other based on a semicontinuous approximation with the so-called thermometer encoding. Furthermore, a weighted approximation, with errors being scaled by the observed levels of variability in RSA for equivalent residues in families of homologous structures, was applied in order to improve the results. The effects of including evolutionary profiles and the growth of sequence databases were assessed. In accord with the observed levels of variability in RSA for different ranges of RSA values, the regression accuracy is higher for buried than for exposed residues, with overall 15.3-15.8% mean absolute errors and correlation coefficients between the predicted and experimental values of 0.64-0.67 on different control sets. The new method outperforms classification-based algorithms when the real value predictions are projected onto two-class classification problems with several commonly used thresholds to separate exposed and buried residues. For example, classification accuracy of about 77% is consistently achieved on all control sets with a threshold of 25% RSA. A web server that enables RSA prediction using the new method and provides customizable graphical representation of the results is available at http://sable.cchmc.org. PMID- 15281129 TI - The occurrence of C--H...O hydrogen bonds in alpha-helices and helix termini in globular proteins. AB - A comprehensive database analysis of C--H...O hydrogen bonds in 3124 alpha helices and their corresponding helix termini has been carried out from a nonredundant data set of high-resolution globular protein structures resolved at better than 2.0 A in order to investigate their role in the helix, the important protein secondary structural element. The possible occurrence of 5 --> 1 C--H...O hydrogen bond between the ith residue CH group and (i - 4)th residue C==O with C...O < or = 3.8 A is studied, considering as potential donors the main-chain Calpha and the side-chain carbon atoms Cbeta, Cgamma, Cdelta and Cepsilon. Similar analysis has been carried out for 4 --> 1 C--H...O hydrogen bonds, since the C--H...O hydrogen bonds found in helices are predominantly of type 5 --> 1 or 4 --> 1. A total of 17,367 (9310 of type 5 --> 1 and 8057 of type 4 --> 1) C- H...O hydrogen bonds are found to satisfy the selected criteria. The average stereochemical parameters for the data set suggest that the observed C--H...O hydrogen bonds are attractive interactions. Our analysis reveals that the Cgamma and Cbeta hydrogen atom(s) are frequently involved in such hydrogen bonds. A marked preference is noticed for aliphatic beta-branched residue Ile to participate in 5 --> 1 C--H...O hydrogen bonds involving methylene Cgamma 1 atom as donor in alpha-helices. This may be an enthalpic compensation for the greater loss of side-chain conformational entropy for beta-branched amino acids due to the constraint on side-chain torsion angle, namely, chi1, when they occur in helices. The preference of amino acids for 4 --> 1 C--H...O hydrogen bonds is found to be more for Asp, Cys, and for aromatic residues Trp, Phe, and His. Interestingly, overall propensity for C--H...O hydrogen bonds shows that a majority of the helix favoring residues such as Met, Glu, Arg, Lys, Leu, and Gln, which also have large side-chains, prefer to be involved in such types of weak attractive interactions in helices. The amino acid side-chains that participate in C--H...O interactions are found to shield the acceptor carbonyl oxygen atom from the solvent. In addition, C--H...O hydrogen bonds are present along with helix stabilizing salt bridges. A novel helix terminating interaction motif, X Gly with Gly at C(cap) position having 5 --> 1 Calpha--H...O, and a chain reversal structural motif having 1 --> 5 Calpha-H...O have been identified and discussed. Our analysis highlights that a multitude of local C--H...O hydrogen bonds formed by a variety of amino acid side-chains and Calpha hydrogen atoms occur in helices and more so at the helix termini. It may be surmised that the main-chain Calpha and the side-chain CH that participate in C--H...O hydrogen bonds collectively augment the cohesive energy and thereby contribute together with the classical N--H...O hydrogen bonds and other interactions to the overall stability of helix and therefore of proteins. PMID- 15281130 TI - Combining evolutionary and structural information for local protein structure prediction. AB - We study the effects of various factors in representing and combining evolutionary and structural information for local protein structural prediction based on fragment selection. We prepare databases of fragments from a set of non redundant protein domains. For each fragment, evolutionary information is derived from homologous sequences and represented as estimated effective counts and frequencies of amino acids (evolutionary frequencies) at each position. Position specific amino acid preferences called structural frequencies are derived from statistical analysis of discrete local structural environments in database structures. Our method for local structure prediction is based on ranking and selecting database fragments that are most similar to a target fragment. Using secondary structure type as a local structural property, we test our method in a number of settings. The major findings are: (1) the COMPASS-type scoring function for fragment similarity comparison gives better prediction accuracy than three other tested scoring functions for profile-profile comparison. We show that the COMPASS-type scoring function can be derived both in the probabilistic framework and in the framework of statistical potentials. (2) Using the evolutionary frequencies of database fragments gives better prediction accuracy than using structural frequencies. (3) Finer definition of local environments, such as including more side-chain solvent accessibility classes and considering the backbone conformations of neighboring residues, gives increasingly better prediction accuracy using structural frequencies. (4) Combining evolutionary and structural frequencies of database fragments, either in a linear fashion or using a pseudocount mixture formula, results in improvement of prediction accuracy. Combination at the log-odds score level is not as effective as combination at the frequency level. This suggests that there might be better ways of combining sequence and structural information than the commonly used linear combination of log-odds scores. Our method of fragment selection and frequency combination gives reasonable results of secondary structure prediction tested on 56 CASP5 targets (average SOV score 0.77), suggesting that it is a valid method for local protein structure prediction. Mixture of predicted structural frequencies and evolutionary frequencies improve the quality of local profile-to-profile alignment by COMPASS. PMID- 15281131 TI - The SHS2 module is a common structural theme in functionally diverse protein groups, like Rpb7p, FtsA, GyrI, and MTH1598/TM1083 superfamilies. AB - Using structural comparisons, we identified a novel domain with a simple fold in the bacterial cell division ATPase FtsA, the archaeo-eukaryotic RNA polymerase subunit Rpb7p, the GyrI superfamily, and the uncharacterized MTH1598/Tm1083-like proteins. The fold contains a core of 3 strands, forming a curved sheet, and a single helix in a strand-helix-strand-strand (SHS2) configuration. The SHS2 domain may exist either in single or duplicate copies within the same polypeptide. The single-copy versions of the domain in FtsA and Rbp7p are most closely related, and appear to mediate protein-protein interactions by means of strand 1, and the loop between strand 2 and strand 3 of the domain. We predict that the interactions between FtsA and its functional partners in bacterial cell division are likely to be similar to the interactions of Rbp7p in the archaeo eukaryotic RNA polymerase complex. The dimeric versions typified by the GyrI superfamily appear to have been adapted for small-molecule binding. Sequence profiles searches helped us to identify several new versions of the GyrI superfamily, including a family of secreted forms that is found only in animals and the bacterial pathogen Leptospira. Through sequence-structure comparisons, we predict the positions that are likely to be important for ligand specificity in the GyrI superfamily. In the MTH1598/Tm1083-like proteins, a SHS2 domain is inserted into the loop between strand 1 and helix 1 of another SHS2 domain. This has resulted in a structure that has convergent similarities with the Hsp33 and green fluorescent protein folds. The sequence conservation pattern and its phyletic profile suggest that it might function as an enzyme in some conserved aspect of nucleic acid metabolism. Thus, the SHS2 domain is an example of a simple module that has been adapted to perform an entire spectrum of functions ranging from protein-protein interactions to small-molecule recognition and catalysis. PMID- 15281132 TI - Function-dependent clustering of orthologues and paralogues of cyclophilins. AB - The 18 kDa archetypal cyclosporin-A binding protein, cyclophilin-A, has multiple paralogues in the human genome. Only 18 of those paralogues have been detected as mRNAs or proteins whose masses vary from 18 to 354 kDa, whereas the functional significance of the open reading frames (ORFs) encoding other paralogues of cyclophilin-A remains unknown. The genomes of Drosophila melanogaster, Caenorhabditis elegans, Arabidopsis thaliana, Schizosaccharomyces pombe, and Saccharomyces cerevisiae encode different numbers of the cyclophilin paralogues, some of which are orthologous to the human cyclophilins. A library of novel algorithms was developed and used for computation of the conservation levels for hydrophobicity and bulkiness profiles, and amino acid compositions (AACs) of 303 aligned sequences of cyclophilins. The majority of the paralogues and orthologues encoded in these 6 genomes differ considerably from each other. Some of the orthologues and paralogues have high correlation coefficients (CCFs) for pairwise compared hydrophobicity and bulkiness profiles, and whose AACs differ to a low degree. Convergence of these three properties of the polypeptide chain and apparent conservation of the typical sequence hallmarks and parameters allowed for the clustering of the functionally related orthologues and paralogues of the cyclophilins. The clustering method allowed for sorting out the cyclophilins into several distinct classes. Analyses of the overlapping clusters of sequences permitted delineation of some hypothetical pathways that might have led to the creation of certain paralogues of cyclophilins in the eukaryotic genomes. PMID- 15281133 TI - Effect of glycosylation on the structure of Erythrina corallodendron lectin. AB - The three-dimensional structure of the recombinant form of Erythrina corallodendron lectin, complexed with lactose, has been elucidated by X-ray crystallography at 2.55 A resolution. Comparison of this non-glycosylated structure with that of the native glycosylated lectin reveals that the tertiary and quaternary structures are identical in the two forms, with local changes observed at one of the glycosylation sites (Asn17). These changes take place in such a way that hydrogen bonds with the neighboring protein molecules in rECorL compensate those made by the glycan with the protein in ECorL. Contrary to an earlier report, this study demonstrates that the glycan attached to the lectin does not influence the oligomeric state of the lectin. Identical interactions between the lectin and the non-covalently bound lactose in the two forms indicate, in line with earlier reports, that glycosylation does not affect the carbohydrate specificity of the lectin. The present study, the first of its kind involving a glycosylated protein with a well-defined glycan and the corresponding deglycosylated form, provides insights into the structural aspects of protein glycosylation. PMID- 15281135 TI - Crystal structure of methenyltetrahydrofolate synthetase from Mycoplasma pneumoniae (GI: 13508087) at 2.2 A resolution. PMID- 15281134 TI - Evaluation of the relative stability of liganded versus ligand-free protein conformations using Simplicial Neighborhood Analysis of Protein Packing (SNAPP) method. AB - Many proteins change their conformation upon ligand binding. For instance, bacterial periplasmic binding proteins (bPBPs), which transport nutrients into the cytoplasm, generally consist of two globular domains connected by strands, forming a hinge. During ligand binding, hinge motion changes the conformation from the open to the closed form. Both forms can be crystallized without a ligand, suggesting that the energy difference between them is small. We applied Simplicial Neighborhood Analysis of Protein Packing (SNAPP) as a method to evaluate the relative stability of open and closed forms in bPBPs. Using united residue representation of amino acids, SNAPP performs Delaunay tessellation of the protein, producing an aggregate of space-filling, irregular tetrahedra with nearest neighbor residues at the vertices. The SNAPP statistical scoring function is derived from log-likelihood scores for all possible quadruplet compositions of amino acids found in a representative subset of the Protein Data Bank, and the sum of the scores for a given protein provides the total SNAPP score. Results of scoring for bPBPs suggest that in most cases, the unliganded form is more stable than the liganded form, and this conclusion is corroborated by similar observations of other proteins undergoing conformation changes upon binding their ligands. The results of these studies suggest that the SNAPP method can be used to predict the relative stability of accessible protein conformations. Furthermore, the SNAPP method allows delineation of the role of individual residues in protein stabilization, thereby providing new testable hypotheses for rational site-directed mutagenesis in the context of protein engineering. PMID- 15281138 TI - Spatiotemporal wavelet resampling for functional neuroimaging data. AB - The study of dynamic interdependences between brain regions is currently a very active research field. For any connectivity study, it is important to determine whether correlations between two selected brain regions are statistically significant or only chance effects due to non-specific correlations present throughout the data. In this report, we present a wavelet-based non-parametric technique for testing the null hypothesis that the correlations are typical of the data set and not unique to the regions of interest. This is achieved through spatiotemporal resampling of the data in the wavelet domain. Two functional MRI data sets were analysed: (1) Data from 8 healthy human subjects viewing a checkerboard image, and (2) "Null" data obtained from 3 healthy human subjects, resting with eyes closed. It was demonstrated that constrained resampling of the data in the wavelet domain allows construction of bootstrapped data with four essential properties: (1) Spatial and temporal correlations within and between slices are preserved, (2) The irregular geometry of the intracranial images is maintained, (3) There is adequate type I error control, and (4) Expected experiment-induced correlations are identified. The limitations and possible extensions of the proposed technique are discussed. PMID- 15281139 TI - Temporal dynamics of ipsilateral and contralateral motor activity during voluntary finger movement. AB - The role of motor activity ipsilateral to movement remains a matter of debate, due in part to discrepancies among studies in the localization of this activity, when observed, and uncertainty about its time course. The present study used magnetoencephalography (MEG) to investigate the spatial localization and temporal dynamics of contralateral and ipsilateral motor activity during the preparation of unilateral finger movements. Eight right-handed normal subjects carried out self-paced finger-lifting movements with either their dominant or nondominant hand during MEG recordings. The Multi-Start Spatial Temporal multi-dipole method was used to analyze MEG responses recorded during the movement preparation and early execution stage (-800 msec to +30 msec) of movement. Three sources were localized consistently, including a source in the contralateral primary motor area (M1) and in the supplementary motor area (SMA). A third source ipsilateral to movement was located significantly anterior, inferior, and lateral to M1, in the premotor area (PMA) (Brodmann area [BA] 6). Peak latency of the SMA and the ipsilateral PMA sources significantly preceded the peak latency of the contralateral M1 source by 60 msec and 52 msec, respectively. Peak dipole strengths of both the SMA and ipsilateral PMA sources were significantly weaker than was the contralateral M1 source, but did not differ from each other. Altogether, the results indicated that the ipsilateral motor activity was associated with premotor function, rather than activity in M1. The time courses of activation in SMA and ipsilateral PMA were consistent with their purported roles in planning movements. PMID- 15281140 TI - Monocular visual activation patterns in albinism as revealed by functional magnetic resonance imaging. AB - Human albinism is characterized by a disturbance of the chiasmatic projection system leading to predominant representation of just one eye in the contralateral hemisphere. Patients show congenital nystagmus without perceiving oscillopsia. The purpose of the present study was to demonstrate the consequences of atypical chiasmatic crossing with monocular visual stimulation using functional magnetic resonance imaging (fMRI). Sixteen patients with albinism and fifteen normally pigmented controls were stimulated with a monocular visual activation paradigm using flickering checkerboards. In patients, we observed contralaterally dominated activation of visual cortices correlating to clinical albinism parameters. This confirms albinism as a continuous range of hypopigmentation disorders. Additionally, albinos showed activation of the superior colliculus and of visual motion areas although the stimulus was stationary. Activation of visual motion areas is due probably to congenital nystagmus without a conscious correlate like oscillopsia. PMID- 15281141 TI - Estimation of multiscale neurophysiologic parameters by electroencephalographic means. AB - It is shown that new model-based electroencephalographic (EEG) methods can quantify neurophysiologic parameters that underlie EEG generation in ways that are complementary to and consistent with standard physiologic techniques. This is done by isolating parameter ranges that give good matches between model predictions and a variety of experimental EEG-related phenomena simultaneously. Resulting constraints range from the submicrometer synaptic level to length scales of tens of centimeters, and from timescales of around 1 ms to 1 s or more, and are found to be consistent with independent physiologic and anatomic measures. In the process, a new method of obtaining model parameters from the data is developed, including a Monte Carlo implementation for use when not all input data are available. Overall, the approaches used are complementary to other methods, constraining allowable parameter ranges in different ways and leading to much tighter constraints overall. EEG methods often provide the most restrictive individual constraints. This approach opens a new, noninvasive window on quantitative brain analysis, with the ability to monitor temporal changes, and the potential to map spatial variations. Unlike traditional phenomenologic quantitative EEG measures, the methods proposed here are based explicitly on physiology and anatomy. PMID- 15281143 TI - Radical amination with sulfonyl azides: a powerful method for the formation of C N bonds. AB - A novel reaction for the introduction of an azide moiety by means of a mild radical process is currently under development. Sulfonyl azides are suitable azidating agents for nucleophilic radicals, such as secondary and tertiary alkyl radicals. More electrophilic radicals, such as enolate radicals, do not react with sulfonyl azides. This feature allowed the development of efficient intra- and intermolecular carboazidations of olefins. Due to the versatility of the azido group, this reaction has an important synthetic potential, as already demonstrated by the preparation of the core of several alkaloids, particularly those containing an amino-substituted quaternary carbon center, such as FR901483. PMID- 15281144 TI - Synthesis, characterization, and electronic structure of Ba5In4Bi5: an acentric and one-electron deficient phase. AB - The new ternary phase Ba(5)In(4)Bi(5) was synthesized by direct reaction of the corresponding elements at high temperature. It crystallizes in a noncentrosymmetric space group and represents a new structure type (tetragonal, P4nc with a=10.620(2) and c=9.009(2) A, Z=2). The structure is built of interconnected heteroatomic clusters of In(4)Bi(5), square pyramids with In(4) bases and four exo-bonded bismuth atoms (bond to the In atoms). According to Wade's rule the compound is electron-deficient with one electron per cluster, that is, [In(4)Bi(5)](10-) instead of the expected [In(4)Bi(5)](11-) for a closed shell species. The clusters are discussed also in light of the known heteroatomic deltahedral clusters with the same composition but different charge, [In(4)Bi(5)](3-). Band structure calculations on the new compound suggest substantial participation of barium in the overall bonding of the structure that "accounts" for the electron shortage PMID- 15281145 TI - Metal coordination to the formal P=N bond of an iminophosphorane and charge density evidence against hypervalent phosphorus(V). AB - The iminophosphorane Ph(2)P(CH(2)Py)(NSiMe(3)) (1) was treated with deprotonating alkali metal reagents to give [(Et(2)O)Li[Ph(2)P(CHPy)(NSiMe(3))]] (2), [[Ph(2)P(CH(2)Py)(NSiMe(3))]Li[Ph(2)P(CHPy)(NSiMe(3))]] (3) and [[Ph(2)P(CH(2)Py)(NSiMe(3))]Na[Ph(2)P(CHPy)(NSiMe(3))]] (4). We report their coordination behaviour in solid-state structures and NMR spectroscopic features in solution. Furthermore, we furnish experimental evidence against hypervalency of the phosphorus atom in iminophosphoranes from experimental charge-density studies and subsequent topological analysis. The topological properties, correlated to the results from NMR spectroscopic investigations, illustrate that the formal P=N double bond is better written as a polar P(+)--N(-) single bond. Additionally, the effects of metal coordination on the bonding parameters of the iminophosphorane and the related anion are discussed. PMID- 15281146 TI - Thermodynamic stability of hydrogen-bonded nanostructures: a calorimetric study. AB - The self-assembly of hydrogen-bonded aggregates (rosettes) in solvent mixtures of different polarity has been studied by calorimetry. The C(50) parameter, the concentration when 50 % of the components are incorporated in the assembly, is used to compare assemblies with different stoichiometry. C(50) for the single rosette 1(3).(BuCYA)(3) (1=N,N-di(4-tert-butylphenyl)melamine; BuCYA=n butylcyanuric acid) in 1,2-dichloroethane is 25 microM, whereas for double rosettes 2 a(3).(BuCYA)(6) and 2 b(3).(BuCYA) (2=calix[4]arene-dimelamine) it is 0.7 and 7.1 microM, respectively. DeltaG degrees, DeltaH degrees, and TDeltaS degrees values indicate that the thermodynamics of double rosettes reflect the independent assembly of two individual single rosette structures or two rosettes reinforced by additional stabilizing interactions. In more polar solvents the stability of double rosettes decreases. From the correlation of DeltaG degrees with solvent polarity it is predicted that it should be possible to assemble double rosettes in methanol or water. The assembly of 2 b(3).(BuCYA)(6) in 100 % MeOH was proven by (1)H NMR and CD spectroscopy. PMID- 15281147 TI - Ruthenium complexes of easily accessible tridentate ligands based on the 2-aryl 4,6-bis(2-pyridyl)-s-triazine motif: absorption spectra, luminescence properties, and redox behavior. AB - A family of tridendate ligands 1 a-e, based on the 2-aryl-4,6-di(2-pyridyl)-s triazine motif, was prepared along with their hetero- and homoleptic Ru(II) complexes 2 a-e ([Ru(tpy)(1 a-e)](2+); tpy=2,2':6',2"-terpyridine) and 3 a-e ([(Ru(1 a-e)(2)](2+)), respectively. The ligands and their complexes were characterized by (1)H NMR spectroscopy, ES-MS, and elemental analysis. Single crystal X-ray analysis of 2 a and 2 e demonstrated that the triazine core is nearly coplanar with the non-coordinating ring, with dihedral angles of 1.2 and 18.6 degrees, respectively. The redox behavior and electronic absorption and luminescence properties (both at room temperature in liquid acetonitrile and at 77 K in butyronitrile rigid matrix) were investigated. Each species undergoes one oxidation process centered on the metal ion, and several (three for 2 a-e and four for 3 a-e) reduction processes centered on the ligand orbitals. All compounds exhibit intense absorption bands in the UV region, assigned to spin allowed ligand-centered (LC) transitions, and moderately intense spin-allowed metal-to-ligand charge-transfer (MLCT) absorption bands in the visible region. The compounds exhibit relatively intense emissions, originating from triplet MLCT levels, both at 77 K and at room temperature. The incorporation of triazine rings and the near planarity of the noncoordinating ring increase the luminescence lifetimes of the complexes by lowering the energy of the (3)MLCT state and creating a large energy gap to the dd state. PMID- 15281148 TI - Alkenyl selenols and selenocyanates: synthesis, spectroscopic characterization by photoelectron spectroscopy, and quantum chemical study. AB - Vinyl, allyl, and homoallyl selenols were easily prepared by a chemoselective reduction of the corresponding selenocyanates with aluminum hydrides. Two stable vinyl and five stable allyl conformers of both series were predicted on the potential-energy surface. The interaction of SeH or SeCN groups with the vinyl group has been investigated with UV photoelectron spectroscopy and quantum chemical calculations, using the MP2/cc-pVTZ and B3LYP/cc-pVTZ levels. In the vinyl derivatives, a surprisingly strong direct conjugation of the selenium lone electron pair and the C=C double bond was observed. On the other hand, in allyl position the selenium lone pair is independent on the C=C double bond, and the hyperconjugation between the Se-C bond and the double bond is the ruling effect. Thus is clarified the type and extent of the interaction between the SeH or SeCN group and the unsaturated moiety. PMID- 15281149 TI - Preparation of tripeptide-bridged dicatechol ligands and their macrocyclic molybdenum(VI) complexes: fixation of the RGD sequence and the WKY sequence of urotensin ii in a cyclic conformation. AB - Dicatechol ligands were prepared with caprylic acid (6-H(4)) or the naturally occurring RGD (23-H(4)) or WKY sequences (32-H(4)) as spacers. 6-H(4) was prepared by solution-phase amide coupling chemistry, while 16, the precursor of 23-H(4), was obtained by solution-phase and solid-phase preparation. In the latter case, a polystyrene resin with a hydrazine benzoate linker was used as the solid support. The last coupling step was performed simultaneously with cleavage of the peptide from the resin. The protecting groups of 16 were all removed in one step to yield the free ligand 23-H(4). The WKY-bridged derivative 32-H(4) was obtained by a similar solid-phase synthesis followed by deprotection. The reaction of all three ligands with dioxomolybdenum(VI) bis(acetylacetonate) afforded 19-membered metallamacrocycles in which the short peptides are conformationally fixed in a turn-type structure. Hereby, the side-chain functionalities of the peptides do not interfere in the metal complexation. PMID- 15281150 TI - Hollow boron nitride (BN) nanocages and BN-nanocage-encapsulated nanocrystals. AB - Hollow boron nitride (BN) nanocages (nanospheres, image on the left) and BN nanocage-encapsulated GaN nanocrystals (right) have been synthesized by using a homemade B-N-O precursors. The as-prepared BN hollow nanocages have typically spherical morphologies with diameters ranging from 30 to 200 nm. The nanocages have crystalline structures. Peanutlike nanocages with double walls have also been observed; their internal space is divided into seperated compartments by the internal walls. The method is extended to sheathe nanocrystals with BN nanocages; BN-shell/GaN-core nanostructures have been successfully fabriacted. The method may be generally applicable to the fabrication BN-sheathed nanocrystals. PMID- 15281151 TI - The direct catalytic asymmetric alpha-aminooxylation reaction: development of stereoselective routes to 1,2-diols and 1,2-amino alcohols and density functional calculations. AB - Proline-catalyzed direct asymmetric alpha-aminooxylation of ketones and aldehydes is described. The proline-catalyzed reactions between unmodified ketones or aldehydes and nitrosobenzene proceeded with excellent diastereo- and enantioselectivities. In all cases tested, the corresponding products were isolated with >95 % ees. Methyl alkyl ketones were regiospecifically oxidized at the methylene carbon atom to afford enantiomerically pure alpha-aminooxylated ketones. In addition, cyclic ketones could be alpha,alpha'-dioxidized with remarkably high selectivity, furnishing the corresponding diaminooxylated ketones with >99 % ees. The reaction mechanism of the proline-catalyzed direct asymmetric alpha-aminooxylation was investigated, and we performed density functional theory (DFT) calculations in order to investigate the nature of the plausible transition states further. We also screened other organocatalysts for the asymmetric alpha oxidation reaction and found that several proline derivatives were also able to catalyze the transformation with excellent enantioselectivities. Moreover, stereoselective routes for the synthesis of monoprotected vicinal diols and hydroxyketones were found. In addition, short routes for the direct preparation of enantiomerically pure epoxides and 1,2-amino alcohols are presented. The direct catalytic alpha-oxidation is also a novel route for the stereoselective preparation of beta-adrenoreceptor antagonists. PMID- 15281153 TI - A series of redox active, tetrathiafulvalene-based amidopyridines and bipyridines ligands: syntheses, crystal structures, a radical cation salt and group 10 transition-metal complexes. AB - Amidopyridine and -2,2'-bipyridine derivatives of EDT-TTF and BTM-TTF (EDT=ethylenedithio, BTM=bis(thiomethyl), TTF=tetrathiafulvalene) have been synthesized and crystallographically characterized. In the solid state, the different supramolecular organization of all these donors results from the competition between the intermolecular interactions, that is, van der Waals, hydrogen-bonding, pi-pi stacking, and donor-acceptor interactions. The electron donating properties of the new donors have been investigated by cyclic voltammetry measurements. A radical cation salt, formulated [EDT-TTF-CONH-m Py](.) (+)[PF(6)](-), has been prepared by electrocrystallization and its crystal structure determined by X-ray analysis. Square planar dicationic complexes with the same donor and M(II)L(2) fragments (M=Pd, Pt, L(2)=bis(diphenylphosphino)propane (dppp) or bis(diphenylphosphino)ethane (dppe)) have been synthesized and one of them, containing the Pd(dppp) unit, has been structurally characterized. The conformation of the complex in the crystalline state is anti, with the coexistence of the dl racemic pair of enantiomers. PMID- 15281152 TI - Biquinolino-modified beta-cyclodextrin dimers and their metal complexes as efficient fluorescent sensors for the molecular recognition of steroids. AB - A series of bridged beta-cyclodextrin (beta-CyD) dimers possessing functional tethers of various lengths was synthesized in moderate yield by the treatment of 2,2'-biquinoline- 4,4'-dicarboxylic dichloride with beta-CyD or mono[6 oligo(ethylenediamino)-6-deoxy]-beta-CyDs. The products were 2,2'-biquinoline 4,4'-dicarboxy-bridged bis(6-O-beta-CyD) (8), N,N'-bis(2-aminoethyl)-2,2' biquinoline-4,4'-dicarboxamide-bridged bis(6-amino-6-deoxy-beta-CyD) (9), and N,N'-bis(5-amino-3-azapentyl)-2,2'-biquinoline-4,4'-dicarboxamide-bridged bis(6 amino-6-deoxy-beta-CyD) (10). The reaction of 8-10 with copper perchlorate give their copper(II) complexes 11-13 in satisfactory yields of over 77 %. All the bis(beta-CyD)s 8-13 act as efficient fluorescent sensors and display remarkable fluorescence enhancement upon addition of optically inert steroids. The inclusion complexation behaviors of 8-13 when treated with the representative steroids cholate (14), deoxycholate (15), and glycocholate (16) in aqueous solution at 25 degrees C were investigated by means of UV/Vis spectroscopy, conductivity and fluorescence measurements, circular dichroism spectroscopy, and 2D NMR spectroscopy. The tether length of bis(beta-CyD) 9 allows it to adopt a cooperative host-tether-guest binding mode in which the spacer and guest are co included in the two CyD cavities. As a result of this cooperation, 9 has a stability constant (K(s)) about 2x10(2) times higher than that of monomodified beta-CyD 4 for inclusion complexation with cholate. Metallooligo(beta-CyD)s with four beta-CyD units have enhanced binding abilities compared with monomodified beta-CyDs. These metallo compounds have binding affinities for guest steroids that are up to 50-4.1x10(3) times higher than those of CyDs 2-4. The guest induced fluorescence enhancement of bis(CyD)s opens a new channel for the design of sensor materials. The complex stability constants of these compounds are discussed from the viewpoint of induced-fit interaction and cooperative multiple binding between host and guest. PMID- 15281154 TI - Behaviour of [PdH(dppe)2]X (X=CF3SO3-, SbF6-, BF4-) as proton or hydride donor: relevance to catalysis. AB - The synthesis, characterization and properties of [PdH(dppe)(2)](+)CF(3)SO(3) ( ).0.125 THF (1; dppe=1,2-bis(diphenylphosphanyl)ethane) and its SbF(6) (-) (1') and BF(4) (-) (1") analogues, the missing members of the [MH(dppe)(2)](+)X(-) (M=Ni, Pd, Pt) family, are described. The Pd hydrides are not stable in solution and can react as proton or hydride donors with formation of dihydrogen, [Pd(dppe)(2)](2+) and [Pd(dppe)(2)]. Complexes 1-1" react with carbocations and carbanions by transferring a hydride and a proton, respectively. Such H(-) or H(+) transfer occurs also towards unsaturated compounds, for example, hydrogenation of a C=C double bond. Accordingly, 1 can hydrogenate methyl acrylate to methyl propionate. Complex 1" is an effective (hourly turnover frequency=16) and very selective (100 %) catalyst for the hydrogenation of cyclohexen-2-one to cyclohexanone with dihydrogen under mild conditions. Density functional calculations coupled with a dielectric continuum model were carried out to compute the energetics of the hydride/proton transfer reactions, which were used to rationalize some of the experimental findings. Theory provides strong support for the thermodynamic and kinetic viability of a tetracoordinate Pd complex as an intermediate in the reactions. PMID- 15281155 TI - De novo design, synthesis, and function of semiartificial myoglobin conjugated with coiled-coil two-alpha-helix peptides. AB - The introduction of a flavin chromophore on the myoglobin (Mb) surface and an effective electron-transfer (ET) reaction through the flavin were successfully achieved by utilizing the self-assembly of heterostranded coiled-coil peptides. We have prepared a semiartificial Mb, named Mb-1alphaK, in which an amphiphilic and cationic alpha-helix peptide is conjugated at the heme propionate (Heme 1alphaK). Heme-1alphaK has a covalently bound iron-protoporphyrin IX (heme) at the N terminus of a 1alphaK peptide sequence. This sequence was designed to form a heterostranded coiled-coil in the presence of a counterpart amphiphilic and anionic 1alphaE peptide sequence in a parallel orientation. Two peptides, Fla(1) 1alphaE and Fla(31)-1alphaE, both incorporating a 10-methylisoalloxazine moiety as an artificial flavin molecule, were also prepared (Fla=2-[7-(10 methyl)isoalloxazinyl]-2-oxoethyl). Heme-1alphaK was successfully inserted into apomyoglobin to give Mb-1alphaK. Mb-1alphaK recognized the flavin-modified peptides and a two-alpha-helix structure was formed. In addition, an efficient ET from reduced nicotinamide adenine dinucleotide to the heme center through the flavin unit was observed. The ET rate was faster in the presence of Fla(1) 1alphaE than in the presence of Fla(31)-1alphaE or the equivalent molecule that has no peptide chain. These results demonstrate that the introduction of a functional chromophore on the Mb surface can be achieved by using specific peptide-peptide interactions. Moreover, the dependence of the ET rate on the position of the flavin indicated that the distance between the heme active site and the flavin chromophore was regulated by the three-dimensional structure of the designed polypeptide. PMID- 15281156 TI - Rationally functionalized deltahedral zintl ions: synthesis and characterization of [Ge9-ER3]3-, [R3E-Ge9-ER3]2-, and [R3E-Ge9-Ge9-ER3]4- (E=Ge, Sn; R=Me, Ph). AB - Six new derivatized deltahedral Zintl ions have been synthesized by reactions between the known Zintl ions Ge(9) (n-) with the halides R(3)EX and/or the corresponding anions R(3)E(-) for E=Ge or Sn. This rational approach is based on our previous discovery that these derivatization reactions are based on nucleophilic addition to the clusters. All species were structurally characterized as their salts with potassium countercations sequestered in 2,2,2 crypt or [18]crown-6 ether. The tin-containing anions were characterized also in solutions by (119)Sn NMR spectroscopy. The reaction types for such substitutions and the structures of the new anions are discussed. PMID- 15281157 TI - The partial hydrogenation of benzene to cyclohexene by nanoscale ruthenium catalysts in imidazolium ionic liquids. AB - The controlled decomposition of an Ru(0) organometallic precursor dispersed in 1 n-butyl-3-methylimidazolium hexafluorophosphate (BMI.PF(6)), tetrafluoroborate (BMI.BF(4)) or trifluoromethane sulfonate (BMI.CF(3)SO(3)) ionic liquids with H(2) represents a simple and efficient method for the generation of Ru(0) nanoparticles. TEM analysis of these nanoparticles shows the formation of superstructures with diameters of approximately 57 nm that contain dispersed Ru(0) nanoparticles with diameters of 2.6+/-0.4 nm. These nanoparticles dispersed in the ionic liquids are efficient multiphase catalysts for the hydrogenation of alkenes and benzene under mild reaction conditions (4 atm, 75 degrees C). The ternary diagram (benzene/cyclohexene/BMI.PF(6)) indicated a maximum of 1 % cyclohexene concentration in BMI.PF(6), which is attained with 4 % benzene in the ionic phase. This solubility difference in the ionic liquid can be used for the extraction of cyclohexene during benzene hydrogenation by Ru catalysts suspended in BMI.PF(6). Selectivities of up to 39 % in cyclohexene can be attained at very low benzene conversion. Although the maximum yield of 2 % in cyclohexene is too low for technical applications, it represents a rare example of partial hydrogenation of benzene by soluble transition-metal nanoparticles. PMID- 15281158 TI - Analysis of the 13C NMR spectra of molecules, chiral by isotopic substitution, dissolved in a chiral oriented environment: towards the absolute assignment of the pro-R/pro-S character of enantiotopic ligands in prochiral molecules. AB - We examined and discuss the proton- and deuterium-decoupled carbon-13 1D spectrum of a molecule, chiral by virtue of the isotopic substitution, dissolved in a chiral oriented medium which simultaneously exhibits chiral discrimination, enantiomeric enrichment and isotope effect. Using the 1-deutero-(2',3',4',5',6' pentadeuterophenyl)phenylmethanol orientationally ordered in a chiral nematic liquid crystal as illustrative example, we point out three important features. First, we demonstrate that the absolute assignment of the pro-R/pro-S character may be derived from the absolute configuration of the isotopically chiral analogue. Second, we report evidence that isotopic effect on (13)C chemical shift anisotropy is negligible in a weakly orienting solvent. Third, we definitely establish that the molecular orientation of prochiral C(s) symmetry molecules and their parent compounds that are chiral by virtue of the isotopic substitution is the same. PMID- 15281159 TI - Insights into CO/styrene copolymerization by using Pd(II) catalysts containing modular pyridine-imidazoline ligands. AB - Continuing our studies into the effect that N-N' ligands have on CO/styrene copolymerization, we prepared new C(1)-symmetrical pyridine-imidazoline ligands with 4',5'-cis stereochemistry in the imidazoline ring (5) and 4',5'-trans stereochemistry (6-10) and compared them with our previously reported ligands (1 4). Their coordination to neutral methylpalladium(II) (5 a-10 a) and cationic complexes (5 b-10 b), investigated in solution by NMR spectroscopy, indicates that both the electronic and steric properties of the imidazolines determine the stereochemistry of the palladium complexes. The crystal structures of two neutral palladium precursors [Pd(Me)(2-n)Cl(n)(N-N')] (n=1 for 8 a; n=2 for 9 a') show that the Pd-N coordination distances and the geometrical distortions in the imidazoline ring depend on the electronic nature of the substituents in the imidazoline fragment. Density functional calculations performed on selected neutral and cationic palladium complexes compare well with NMR and X-ray data. The calculations also account for the formation of only one or two stereoisomers of the cationic complexes. The performance of the cationic complexes as catalyst precursors in CO/4-tert-butylstyrene copolymerization under mild pressures and temperatures was analyzed in terms of the productivity and degree of stereoregularity of the polyketones obtained. Insertion of CO into the Pd-Me bond, which was monitored by multinuclear NMR spectroscopy, shows that the N ligand influences the stereochemistry of the acyl species formed. PMID- 15281160 TI - A new system in organooxotin cluster chemistry incorporating inorganic and organic spacers between two ladders each containing five tin atoms. AB - Hydrolysis of dibenzyltin dichloride in ethanol has been found to give an unprecedented carbonate anion (CO(3) (2-))-bridged double-ladder organooxotin cluster, [(R(2)SnO)(3)(R(2)SnOH)(2)(CO(3))](2) (1, R = C(6)H(5)CH(2)), with five tin atoms in each ladder. With the aim of obtaining organooxotin clusters with large cavities suitable for host-guest chemistry, we used 1,1' ferrocenedicarboxylic acid (H(2)L(a)) and hexanedioic acid (H(2)L(b)) to replace the carbonate anions (CO(3) (2-)), and thereby clusters [(R(2)SnO)(3)(R(2)SnOH)(2)L(a)](2) (2) and [(R(2)SnO)(3)(R(2)SnOH)(2)L(b)](2) (3) were obtained. When 1 was treated with benzoic acid (HL(c)) in different stoichiometric ratios (1:4, 1:10), ladder cluster (R(2)SnO)(3)(R(2)SnOH)(2)(L(c))(2) (4) and drum cluster [RSn(O)L(c)](6) (5) were obtained. Through the hydrolysis of Cy(2)SnCl(2) (Cy = C(6)H(11)) and (C(6)H(5)CH(2))(2)SnCl(2), two interesting ethanolate-modified clusters [Cy(2)(C(2)H(5)O)SnOSn(C(2)H(5)O)Cy(2)](2) (6) and [(R(2)SnO)(3)(R(2)SnOH)(R(2)SnOC(2)H(5))(CO(3))](2) (7) were obtained. All the tin atoms in these ladder clusters are five-coordinate surrounded by two alkyl groups and three O atoms, and have distorted trigonal-bipyramidal coordination environments with two carbon atoms and one O atom in the equatorial positions and two O atoms in the axial positions. The structures of all these compounds have been established by single-crystal X-ray diffraction analyses. PMID- 15281161 TI - Ionic liquid analogues formed from hydrated metal salts. AB - A dark green, viscous liquid can be formed by mixing choline chloride with chromium(III) chloride hexahydrate and the physical properties are characteristic of an ionic liquid. The eutectic composition is found to be 1:2 choline chloride/chromium chloride. The viscosity and conductivity are measured as a function of temperature and composition and explained in terms of the ion size and liquid void volume. The electrochemical response of the ionic liquid is also characterised and it is shown that chromium can be electrodeposited efficiently to yield a crack-free deposit. This approach could circumvent the use of chromic acid for chromium electroplating, which would be a major environmental benefit. This method of using hydrated metal salts to form ionic liquids is shown to be valid for a variety of other salt mixtures with choline chloride. PMID- 15281162 TI - Molecular dynamics (MD) simulations and large-angle X-ray scattering (LAXS) studies of the solid-state structure and assembly of isotactic (R)-poly(2,2' dioxy-1,1'-binaphthyl-)phosphazene in the bulk state and in the cast film. AB - The intrachain conformation, molecular structure and interchain assembly of isotactic (R)-poly(2,2'-dioxy-1,1'-binaphthyl)phosphazene (P-DBNP) both in the bulk state (I) and in the cast film (II) were studied by molecular dynamics (MD) simulations of models, as implemented by a bias potential for the analysis of the radial distribution function (RDF) obtained from large-angle X-ray scattering (LAXS) data. The microscopic structure and order extension of the polymer changed from I to II, as qualitatively shown in the shapes of their experimentally measured RDF curves. With the use of a bias potential, the MD simulations provided a much more accurate analysis of the models, as seen in the reproduction of the RDFs. The chiral P-DBNP chain was found to be consistent with helix conformations in both the I and the II samples. The predominant interchain clustering motif was best reproduced with a seven-chain model. In the case of I, the maximum chain length was 18 monomeric -R(2)NP- units, while in the case of the cast film II the chain was more elongated, up to distances of approximately 100 A, equivalent to over 48 monomeric -R(2)NP- units. The seven-chain assembly was accounted for in terms of nonbonded interactions favouring the minimum voids area between the seven tubular structures of the material. The results validate our earlier finding that MD analysis with implementation of a biasing potential for the RDFs can provide quantitative information on the structural and conformational features of amorphous solids. The combined theoretical and experimental approach was found to be a useful tool to detect, locate and evaluate the intra- and intermolecular modifications of materials subsequent to their phase transformation and, as in the present case, changes in their microscopic structures or preparation methods. PMID- 15281163 TI - Does hydrogen bonding matter in crystal engineering? Crystal structures of salts of isomeric ions. AB - The preparation and structure determinations of the crystalline salts [3,3' H(2)bipy][PtCl(4)] (2), [2,2'-H(2)bipy][PtCl(4)] (3) and [1,4'-Hbipy][PtCl(4)] (4) and [3,3'-H(2)bipy][SbCl(5)] (6) and [1,4'-Hbipy][SbCl(5)] (8) are reported. In addition a redetermination of the structure of the metastable salt [4,4' H(2)bipy][SbCl(5)] (5 b) in the corrected space group Pbcm is described. These structures are compared to those of the known salt [4,4'-H(2)bipy][PtCl(4)] (1), the stable triclinic form of [4,4'-H(2)bipy][SbCl(5)] (5 a) and [2,2' H(2)bipy][SbCl(5)] (7). In the case of the salts of the rigid [PtCl(4)](2-) ion, structures 2, 3 and 4 are essentially isostructural despite the differing hydrogen-bonding capability of the cations. Similarly, among the salts of [SbCl(5)](2-) ions, structures 7 and 8 are essentially isostructural. Structure 6 differs from these in having a differing pattern of aggregation of the [SbCl(5)](2-) ions to form polymeric rather than tetrameric units. It is evident that local hydrogen-bonding interactions, although significant, are not the only or even the decisive influence on the crystal structures formed by these salts. These observations are not in good accord with the heuristic "sticky tecton" or supramolecular synthon models for synthetic crystallography or crystal engineering. PMID- 15281164 TI - Thermodynamic analysis of receptors based on guanidinium/boronic acid groups for the complexation of carboxylates, alpha-hydroxycarboxylates, and diols: driving force for binding and cooperativity. AB - The thermodynamics of guanidinium and boronic acid interactions with carboxylates, alpha-hydroxycarboxylates, and diols were studied by determination of the binding constants of a variety of different guests to four different hosts (7-10). Each host contains a different combination of guanidinium groups and boronic acids. The guests included molecules with carboxylate and/or diol moieties, such as citrate, tartrate, and fructose, among others. The Gibbs free energies of binding were determined by UV/Vis absorption spectroscopy, by use of indicator displacement assays. The receptor based on three guanidinium groups (7) was selective for the tricarboxylate guest. The receptors that incorporated boronic acids (8-10) had higher affinities for guests that included alpha hydroxycarboxylate and catechol moieties over guests containing only carboxylates or alkanediols. Isothermal titration calorimetry revealed the enthalpic and entropic contributions to the Gibbs free energies of binding. The binding of citrate and tartrate was investigated with hosts 7-10, for which all the binding events were exothermic, with positive entropy. Because of the selectivity of hosts 8-10, a simple boronic acid (14) was also investigated and determined to be selective for alpha-hydroxycarboxylates and catechols over amino acids and alkanediols. Further, the cooperativity of 8 and 9 in binding tartrate was also investigated, revealing little or no cooperativity with 8, but negative cooperativity with 9. A linear entropy/enthalpy compensation relationship for all the hosts 7-10, 14, and the carboxylate-/diol-containing guests was also obtained. This relationship indicates that increasing enthalpy of binding is offset by similar losses in entropy for molecular recognition involving guanidinium and boronic acid groups. PMID- 15281165 TI - Vibrational and quantum-chemical study of nonlinear optical chromophores containing dithienothiophene as the electron relay. AB - A series of nonlinear optical (NLO) donor-acceptor (D-A) chromophores containing a fused terthiophene, namely dithienothiophene (DTT), as the electron relay, the same donor group, and acceptors of various strengths, has been investigated by means of infrared and Raman spectroscopies, both in the solid state as well as in a variety of solvents, to evaluate the effectiveness of the intramolecular charge transfer from the electron-donor to the electron-acceptor end groups. The Raman spectral profiles of these NLO-phores measured from their dilute solutions have been found to be rather similar to those of the corresponding solids, and thus their intramolecular charge transfer (ICT) shows very little dependence on the solvent polarity. The experimental results obtained for the DTT-containing NLO phore with a 4-(N,N-dibutylamino)styryl end group as the donor and a 2,2 dicyanoethen-1-yl end group as the acceptor differ from those previously obtained for two parent "push-pull" chromophores with the same D-A pair but built-up around either a bis(3,4-ethylenedioxythienyl) (BEDOT) or a bithienyl (BT) electron relay. Vibrational spectroscopy shows that DTT is significantly more efficient as an electron relay than BT (which has the same number of conjugated C=C bonds) or BEDOT (which can be viewed as a rigidified version of BT on account of noncovalent intramolecular interactions between heteroatoms of adjacent rings). Density functional theory (DFT) calculations have also been performed on these NLO-phores to assign their main electronic and vibrational features and to obtain information about their structures. An additional merit of these molecular materials was revealed by the infrared spectra of the DTT-based NLO-phores recorded at different temperatures. Thus, spectra recorded between -170 and 150 degrees C did not show any substantial change, indicating that the materials have a high thermal stability, which is of significance for their use as active components in optoelectronic devices. PMID- 15281166 TI - Toward a 6pi+6pi zwitterion or a bioinhibitors-related OH-substituted aminoquinone: identification of a key intermediate in their pH controlled synthesis. AB - Their pH-controlled reactivity places the N,N'-dialkyl-2-amino-5-lithium alcoholate-1,4-benzoquinonemonoimines [C(6)H(2)(NHCH(2)R') (=NCH(2)R')(=O)(OLi)] 7 (R'=tBu) and 8 (R'=p-C(6)H(4)-tBu) at the crossroads of a new versatile strategy for the preparation of two very different classes of substituted quinones. We describe new 2-(N-alkyl)amino-5-hydroxy-1,4-benzoquinones, which are parent molecules to biologically active substituted aminobenzoquinones, for which changes of the N-substituent will become readily possible. The results of the first X-ray structural determination of such compounds ([C(6)H(2)(NHCH(2)tBu)(OH)(=O)(2)] 13) are also reported and we compare the influence of the number of N-substituents of the C(6) ring on the supramolecular networks resulting from self-assembling of 13, zwitterionic N,N'-dineopentyl-2 amino-5-alcoholate-1,4-benzoquinonemonoiminium [C(6)H(2)(=NHCH(2)tBu)(2)(=O)(2)] 9 and N,N',N",N"'-tetraneopentyl-2,5-diamino-1,4-benzoquinone diimine [C(6)H(2)(NHCH(2)tBu)(2)(=NCH(2)tBu)(2)] 15. PMID- 15281167 TI - Novel stereocontrolled approach to syn- and anti-oxepene-cyclogeranyl trans-fused polycyclic systems: asymmetric total synthesis of (-)-Aplysistatin, (+)-Palisadin A, (+)-Palisadin B, (+)-12-hydroxy-palisadin B, and the AB ring system of adociasulfate-2 and toxicol A. AB - A new stereocontrolled method for the formation of trans-anti cyclogeranyl oxepene systems is described. The demanding stereochemistry is secured by stereoselective coupling of a cyclogeranyl tertiary alcohol with a 1,2 unsymmetrically substituted epoxide, while the formation of the highly strained oxepene is achieved employing ring-closing metathesis. Since the stereochemistry of the trans-fused 6,7-ring system is determined by the epoxide, the method also allows the construction of trans-syn 6,7-ring systems. This approach leads to the synthesis of the AB fragment of Adociasulfate-2 and Toxicol A, for the first time. The flexibility and efficiency of the presented strategy is demonstrated by the total asymmetric synthesis of (-)-Aplysistatin, (+)-Palisadin A, (+)-12 hydroxy-Palisadin B, and (+)-Palisadin B, employing two similar key intermediates. PMID- 15281169 TI - Fighting a flexible virus with flexible drugs. PMID- 15281170 TI - Policy watch. More of the same? PMID- 15281171 TI - Cosmetic surgery: safety first. PMID- 15281172 TI - Gaining control of urinary stress incontinence. PMID- 15281173 TI - It's fast...it's painless...but is it useful? PMID- 15281174 TI - Dry eyes: no longer a crying shame. PMID- 15281175 TI - Should I consider using an insulin pump to take insulin for type 2 diabetes? PMID- 15281176 TI - Dietary omega-3 fatty acids and depression in a community sample. AB - To evaluate the association between omega-3 polyunsaturated essential fatty acids and depression, data regarding prevalence rates of self-reported depression and median daily dietary intakes of these fatty acids were obtained from an age stratified, population-based sample of women (n = 755; 23-97 year) in the Barwon Statistical Division of south-eastern Australia. A self-report questionnaire based on Diagnostic and Statistical Manual-IV criteria was utilised to determine 12-month prevalence rates of depression in this sample, and data from biennial food frequency questionnaires examining seafood and fish oil consumption over a 6 year period were examined. Differences in median dietary intakes of omega-3 fatty acids between the depressed and nondepressed cohorts were analysed and results were adjusted for age, weight and smoking status. No significant differences in median intakes were identified between the two groups of women (median, interquartile range; depressed = 0.09g/day, 0.04-0.18 versus nondepressed = 0.11 g/day, 0.05-0.22, p = 0.3), although overall average intakes of omega-3 fatty acids were lower than recommended and rates of depression within this sample higher than expected, based on previous data. Further research that takes into account ratios of omega-6 to omega-3 polyunsaturated essential fatty acids, as well as other dietary sources of omega-3 fatty acids, is warranted. PMID- 15281177 TI - Multistate licensure. PMID- 15281178 TI - Food, water, and the PVS patient. PMID- 15281179 TI - Emergency contraception revisited: a response to Eugene Diamond. PMID- 15281180 TI - A critique of Hamel and Panicola. PMID- 15281181 TI - Washington insider--the President's Council on Bioethics. PMID- 15281182 TI - Postcoital intervention--from fear of pregnancy to rape crisis. PMID- 15281183 TI - Stem cell research--prospects and problems. PMID- 15281184 TI - Humanity and nanotechnology--judging enhancements. PMID- 15281185 TI - On life-sustaining treatments and the vegetative state--scientific advances and ethical dilemmas. PMID- 15281186 TI - Social contract theory and just decision making: lessons from genetic testing for the BRCA mutations. AB - Decisions about funding health services are crucial to controlling costs in health care insurance plans, yet they encounter serious challenges from intellectual property protection--e.g., patents--of health care services. Using Myriad Genetics' commercial genetic susceptibility test for hereditary breast cancer (BRCA testing) in the context of the Canadian health insurance system as a case study, this paper applies concepts from social contract theory to help develop more just and rational approaches to health care decision making. Specifically, Daniel's and Sabin's "accountability for reasonableness" is compared to broader notions of public consultation, demonstrating that expert assessments in specific decisions must be transparent and accountable and supplemented by public consultation. PMID- 15281187 TI - Oversight of research involving the dead. AB - Research involving the dead, especially heart-beating cadavers, may facilitate the testing of potentially revolutionary and life-saving medical treatments. However, to ensure that such research is conducted ethically, it is essential to: (1) identify appropriate standards for this research and (2) assign institutional responsibility and a mechanism for oversight. Protocols for research involving the dead should be reviewed by a special committee and assessed according to nine standards intended to ensure scientific merit, to protect deceased patients and their families, and to promote institutional integrity and responsibility. Federal regulation of research involving the dead will foster appropriate standards and, equally importantly, help establish the acceptability of such research. PMID- 15281188 TI - Convening a 407 panel for research not otherwise approvable: "Precursors to diabetes in Japanese American youth" as a case study. AB - Subpart D of 45 CFR 46 focuses on research involving children. Section 46.407 addresses research that is not otherwise approvable. The research is not otherwise approvable because either (1) it seeks to enroll healthy children, but offers no prospect of direct benefit and entails more than minimal risk; or (2) it seeks to enroll children with a disorder or condition, but offers no prospect of direct benefit and entails more than a minor increase over minimal risk. According to 46.407, such research can be permissible if it is approved by a panel of experts. Prior to 2000, only two 407 panels had been convened, but in 2001, the Office for Human Research Protections received more than 20 protocols for 407 review. The first, entitled "Precursors to Diabetes in Japanese American Youth," serves here as a case study in human subject protections. PMID- 15281189 TI - Risk standards for pediatric research: rethinking the Grimes ruling. AB - In Grimes v. Kennedy Krieger Institute (KKI), the Maryland Court of Appeals, while noting that U.S. federal regulations include risk standards for pediatric research, endorses its own risk standards. The Grimes case has implications for the debate over whether the minimal risk standard should be interpreted based on the risks in the daily lives of most children (the objective interpretation) or the risks in the daily lives of the children who will be enrolled in a given study (the subjective interpretation). The court's use of the objective interpretation to block studies like the KKI study protects individual children who are worse off than the average child. Unfortunately, this approach also may block research intended to improve the lives of these same individuals. A similar dilemma arises in the context of multinational research, suggesting that a "modified objective standard," proposed to address this dilemma in the multinational setting, may offer a framework for addressing the dilemma in the context of pediatric research as well. PMID- 15281190 TI - Andre Hellegers and Carroll House: architect and blueprint for the Kennedy Institute of Ethics. AB - The Newman programs established at secular colleges and universities provided an opportunity for intellectual, spiritual, and social growth among the Catholic student population. As a young physician and junior medical faculty member, Andre Hellegers took part in the early organization and ongoing work of Carroll House, the Newman Center at the Johns Hopkins Medical Institutions. Helleger's experience at Carroll House enabled him to develop a clear blueprint of an academic center of excellence for the scientific, theological, and philosophical exploration of the many problems that he had seen and foresaw in medicine. That center would become Georgetown's Kennedy Institute of Ethics. PMID- 15281191 TI - Self-infiltration: confusing assigned tasks as self-selected in memory. AB - Two studies examined determinants of self-infiltration (i.e., false self attribution of externally controlled goals or activities). According to Personality Systems Interactions (PSI) theory, a sad mood was expected to reduce access to integrated self-representations and lead to self-infiltration for participants who have an impaired ability to cope with negative affect (i.e., state-oriented participants). Consistent with expectations, state-oriented participants had a tendency toward self-infiltration (as indexed by higher rates of false self-ascription of assigned activities) when reporting higher levels of sadness (Study 1) and after the experimental induction of a sad mood (Study 2). Participants who are able to downregulate negative affect (i.e., action-oriented participants), did not show this tendency. Theoretical and practical implications of the process of self-infiltration are discussed. PMID- 15281192 TI - Digital library projects: beyond the beltway. PMID- 15281193 TI - Genetic research involving human biological materials: a need to tailor current consent forms. PMID- 15281194 TI - What must research subjects be told regarding the results of completed randomized trials? PMID- 15281195 TI - Psychological and social risks of behavioral research. PMID- 15281196 TI - Can underpowered clinical trials be justified? PMID- 15281197 TI - Eugenics and the new genetics. PMID- 15281198 TI - What is wrong in modifying the human germ line? PMID- 15281199 TI - Reappraising genetic screening and testing through the phenomenology of the doctor-patient relationship. PMID- 15281200 TI - Genetic testing and insurance: an international comparison. PMID- 15281201 TI - Is legislative principalism an illusion and perverse policy? A critical opinion of the French regulation and practice of genetic tests. PMID- 15281202 TI - The Council of Ethics' views on therapeutic cloning, 2001. PMID- 15281203 TI - The Council of Ethics' stance on therapeutic cloning, 2001. PMID- 15281204 TI - TRICARE; Individual Case Management Program; Program for Persons with Disabilities; extended benefits for disabled family members of active duty service members; custodial care. Final Rule. AB - The Department is publishing this final rule to implement requirements enacted by Congress in section 701(g) of the National Defense Authorization Act for Fiscal Year 2002 (NDAA-02), which terminates the Individual Case Management Program. The Department withdraws its proposed rule published at 66 FR 39699 on August 1, 2001, regarding the Individual Case Management Program. This rule also implements section 701(b) of the NDAA-02 which provides additional benefits for certain eligible active duty dependents by amending the TRICARE regulations governing the Program for Persons with Disabilities. The Program for Persons with Disabilities is now called the Extended Care Health Option. Other administrative amendments are included to clarify specific policies that relate to the Extended Care Health Option, custodial care, and to update related definitions. PMID- 15281205 TI - The Council of Ethics' views on reproductive cloning, 2001. PMID- 15281206 TI - Human cloning and human dignity: an ethical inquiry--policy recommendations. PMID- 15281207 TI - Genetics and human behaviour: the ethical context--summary and recommendations. PMID- 15281208 TI - Statement on the scope of gene patents, research exemption, and licensing of patented gene sequences for diagnostics. PMID- 15281209 TI - Statement on human genomic databases, December 2002. PMID- 15281210 TI - [Erectile function disorders. Epidemiology, physiology, etiology, diagnosis and therapy]. AB - Erectile dysfunction is a common, age-dependent functional disturbance of men associated to various comorbidities. Interdisciplinary cooperation with neurologists in ca-ses of a suspected neurological aetiology and with psychiatrists in cases with normalorganic diagnostic findings is necessary. Hormone replacement and psychotherapy can cure certain patients. Oral pharmacotherapy is the most effective therapy for erectile dysfunction with the highest patient preference. Oral PDE-5-inhibitors(sildenafil, tadalafil, vardenafil) are superior in effectiveness to centrally acting drugs (apomorphin, yohimbine). Local pharmacotherapy (MUSE, ICI) is a second line therapy in cases of failure or contraindications for oral pharmacotherapy. Vacuum therapy and operative procedures(penile implants) complete the therapeutic options of erectile dysfunction. PMID- 15281211 TI - The conduct of Canadian researchers and institutional review boards regarding substituted consent for research. PMID- 15281212 TI - When a research subject calls with a complaint, what will the institutional review board do? PMID- 15281213 TI - The certificate of confidentiality application: a view from the NIH Institutes. PMID- 15281214 TI - Mortality trend of central nervous system, eye, thyroid, skin, connective tissue and bone in Japan: 1960-2000. PMID- 15281215 TI - Summary of JRC/ESReDA Seminar on Safety Investigation of Accidents, 12-13 May 2003, European Commission, DG JRC-IE, Petten, The Netherlands. PMID- 15281216 TI - Charles Kelman. PMID- 15281217 TI - Sarcomas of the oral and maxillofacial region: a review of 32 cases in 25 years. AB - Thirty-two cases of sarcomas involving the oral and maxillofacial region over a period of 25 years were reviewed. The age range was from 5 months to 77 years with a mean age of 42. The male to female ratio was 3:1. The sarcomas were located in the maxilla including the maxillary sinus (n= 13), mandible (n= 13), buccal mucosa (n= 3), temporomandibular fossa (n= 2), and submandibular region (n= 1). Histologically sarcomas were classified as osteosarcoma (n= 9), malignant fibrous histiocytoma (n= 7), rhabdomyosarcoma (n= 5), fibrosarcoma (n= 3), plasmacytoma (n= 2), leiomyosarcoma (n= 2), angiosarcoma (n= 2), liposarcoma (n= 1), and ameloblastic fibrosarcoma (n= 1). Surgical resection was performed in 29 cases. Local recurrence was found in 10 patients and metastasis in 11 patients. Metastases included five regional lymph node metastases and eight distant metastases. The survival of patients with local recurrence or metastasis was poor. Surgery is the most reliable treatment for sarcomas of the oral and maxillofacial region. Adequate excision with safety surgical margin as the initial therapy is important for better survival. The value of radiation therapy and/or chemotherapy is uncertain. The 5-year survival rate of primary cases was 61%. PMID- 15281218 TI - Obituary. William Ross Adey. PMID- 15281219 TI - Antimutagens, anticarcinogens, and effective worldwide cancer prevention. AB - Cancer is the eventual outcome of the transformation of normal cells by DNA reactive, genotoxic carcinogens and the growth promotion of mutated cells by enhancing factors. It is important to discriminate between genotoxic carcinogens and nongenotoxic chemicals because their mechanisms of action are quite distinct. Their dose-response curves, reversibility, and organ- and species-specificity are also quite distinct. Thus, the mode of action of agents involved in cancer causation and development needs careful analysis. Genotoxic carcinogens are mutagenic, form DNA adducts, and lead to the formation of hydroxy radicals and inappropriate peroxidation reactions that the antioxidants in vegetables, fruits, and tea can effectively decrease. About 35% of known cancers are associated with tobacco use and about 55% with inappropriate nutritional habits. Cancer induction can be decreased by (1) avoiding the formation of carcinogens, (2) reducing their metabolic activation, or (3) increasing their detoxification. Nutritional factors play a major role in cancer prevention. Vegetables, fruits, and the beverage, tea, provide select means to prevent activation of carcinogens or to increase their detoxification. Salting and pickling of certain foods generate direct acting mutagens and carcinogens that affect the stomach and probably the esophagus. Thus, controlling salt use and increasing vegetable and fruit intake can prevent these diseases. Frying and broiling protein foods generate heterocyclic amines that affect the colon, breast, prostate, and pancreas, and their formation is decreased by antioxidants. Heterocyclic amines are converted to reactive components by specific cytochrome P450 enzymes and N acetyltransferases. The tobacco-specific nitrosamines and polycyclic aromatic hydrocarbons undergo specific activation and detoxification processes. These reactions are controlled by antioxidants such as quercetin in vegetables, polyphenols in tea, genistein and daidzin in soy, sulforaphane in broccoli, and 3 methylindole and isothiocyanates in such protective foods. In the Western world, the type and amount of fat play a critical role that operates through specific promoting mechanisms to modify the action of genotoxic carcinogens. In most Western countries where mixed fats and oils are consumed, the total amount of fat at 35 to 40% of calories acts as a powerful promoter. On the other hand, in Italy and Greece, where the monounsaturated oil, olive oil, is used almost exclusively without promoting effect, little enhancement by fat is observed. Also, in the Mediterranean region, meats, mainly veal, are not browned to the point of generating heterocyclic amines. Wheat bran fiber increases stool bulk and eliminates carcinogens and the promoters of colon and breast cancers. Thus, the current base of knowledge on the mechanisms of cancer causation provides effective ways of preventing most types of cancer, the definitive means of cancer control. PMID- 15281220 TI - New approaches to antimutagenesis. AB - One of the most worrisome possibilities in the field of antimutagenesis and anticarcinogenesis is that factors that protect against one mutagen or carcinogen will not be effective against another. Indeed, such specificities are known. Consequently, protective agents found in experimental screens and systems may not be relevant to the human situation. Additionally, the question of dose is also problematic, because factors that can protect against the levels of mutagens or carcinogens present in the human environment may be overwhelmed by the large dose of the agent used in the experiment, a dose necessitated by the need to have an effect against which protection can be judged. We suggest here that the new technologies available for the measurement of somatic mutation and chromosomal damage can be used to study spontaneous events, thus avoiding both problems. PMID- 15281221 TI - Adaptive resistance of Saccharomyces cerevisiae to chronic treatment with genotoxic and nongenotoxic carcinogens. AB - The exposure of mammalian cells or tumors for weeks or months to low nonlethal doses of cytostatic drugs may induce multidrug resistance, which can be enhanced by a variety of DNA-damaging agents. Multidrug resistance to a variety of drugs has been observed. But in yeast, DNA-damaging agents have not yet been tested. As the appearance of resistance is the result of longterm exposure, we decided to extend the application of test substances to a period of up to 400 days. In such long-term experiments S. cerevisiae MP1 adapted to treatment with low doses of mutagens. Consistent results were obtained for both genotoxic and nongenotoxic carcinogenic substances, which implies that there may be a single pathway for carcinogens with different modes of action. PMID- 15281222 TI - Development of test systems for the detection of compounds that prevent the genotoxic effects of heterocyclic aromatic amines: preliminary results with constituents of cruciferous vegetables and other dietary constituents. AB - Over the past decades, strong efforts have been made to identify dietary constituents that protect against the genotoxic effects of heterocyclic aromatic amines (HAAs). However, most of the methods that have been used, in particular in vitro assays that require the addition of exogenous enzyme homogenates, have only a limited predictive value because important protective mechanisms are not adequately represented and may give misleading results. Therefore, we attempted to develop improved test systems, namely assays, with human hepatoma cells and single-cell gel electrophoresis (SCGE) tests with rats. Genotoxicity tests with human derived Hep G2 cells reflect the genotoxic effects of HAAs better than other in vitro systems. They also enable the detection of protective effects since the human derived hepatoma cells possess phase I and phase II enzymes that are involved in the activation/ detoxification of the amines. The most appropriate endpoint for experiments with Hep G2 cells appears to be micronucleus induction, but protocols for other endpoints are available as well. The second promising model is the SCGE ("comet") assay with rats that was used successfully to measure protective effects of constituents of cruciferous vegetables against 2 amino-3-methylimidazo[4,5-flquinoline (IQ) in the liver and in the colon mucosa. The present study describes the experimental design of the new approaches, as well as results obtained with various dietary constituents. PMID- 15281223 TI - Antigenotoxic properties of Terminalia arjuna bark extracts. AB - Compounds possessing antimutagenic properties (polyphenols, tannins, vitamins, etc.) have been identified in fruits, vegetables, spices, and medicinal plants. Terminalia arjuna (Combretaceae), a tropical woody tree occurring throughout India and known locally as Kumbuk, is a medicinal plant rich in tannins and triterpenes that is used extensively in Ayurvedic medicine as a cardiac tonic. The aim of the present collaborative work was to test six solvent extracts from the bark of Terminalia arjuna for antigenotoxic activity using in vitro short term tests. Terminalia arjuna extracts were obtained by sequential extraction using acetone, methanol, methanol + HCl, chloroform, ethyl acetate, and ethyl ether. The antigenotoxic properties of these extracts were investigated by assessing the inhibition of genotoxicity of the directacting mutagen 4 nitroquinoline-N-oxide (4NQO) using the "comet" assay and the micronucleus (MN) test. Human peripheral blood leukocytes were incubated with different concentrations of the six extracts (from 5 to 100 microg/ mL) and with 4NQO (1 and 2 microg/mL, for the "comet" assay and MN test, respectively). Each extract/4NQO combination was tested twice; in each experiment, positive control (4NQO alone) and negative control (1% DMSO) were set. "Comet" assay results showed that acetone and methanol extracts were highly effective in reducing the DNA damage caused by 4NQO, whereas the acidic methanol, chloroform, ethyl acetate, and ethyl ether extracts showed less marked or no antigenotoxic activity. In the MN test, a decrease in 4NQO genotoxicity was observed by testing this mutagen in the presence of acetone, methanol, chloroform, and ethyl acetate extracts, even though the extent of inhibition was not always statistically significant. PMID- 15281224 TI - Antimutagenic property of an herbal medicine, Polygonum multiftorum Thunb. detected by the Tradescantia micronucleus assay. AB - The root extracts of a Chinese herb, Polygonum multiflorum Thunb., have been used for centuries as an internal medicine to improve liver and kidney functions. In this study, we evaluated the antimutagenic property of this drug with the Tradescantia micronucleus (Trad-MCN) assay. The Trad-MCN bioassay is a well established test for chromosome damage induced by physical or chemical agents in terms of micronuclei (MCN) frequency. Inflorescences of the Tradescantia plant cuttings were first exposed to 0.35 Gy soft X-rays (80 kV, 5 mA, 1 mm Al filter, dose rate around 0.50 Gy/min), followed by drug treatments at 1, 3, and 6% concentrations of the aqueous solution for a total recovery period of 24 hours. The positive (X-rays), negative (nutrient solution), and drug control (3% drug solution) groups were maintained in each of the three series of repeated experiments. Flower buds of the treated and control groups were fixed in aceto alcohol (1:3 ratio) in preparation for slides to score MCN frequencies in the early tetrads of the meiotic pollen mother cells. The mean MCN frequencies (MCN/100 tetrads +/- SE) of the positive control (irradiated) was 26.68 +/- 2.49; the negative control was 2.93 +/- 0.50; the PM solution control was 2.06 +/- 0.39, and the 0.35 Gy X-ray plus 6% PM drug treated was 18.76 +/- 1.69. A 45% reduction in chromosome damage was observed. Antimutagenic effects were relatively decreased at lower concentrations of PM. This antimutagenic effect could be attributed to the antioxidant action of PM, enhancement of DNA repair, or the radical elimination from the irradiated plant cells. PMID- 15281226 TI - Ileal effluent as a fermentation substrate: implications for butyrate production in the colon. AB - Fermentation of fiber can lead to an enhanced production of short chain fatty acids (SCFA) and, hence, contribute to the proposed anticarcinogenic properties of butyrate in the colon. The fermentation of fiber isolates and the corresponding ileal effluents has been compared under in vitro conditions. Yield of SCFA per gram of substrate fermented was similar for isolates and fiber enriched effluents (approximately 4.9 mmol/g) and it could be inferred that nonfiber components of effluent also generated SCFA. Butyrate production was highest for glucan-based polymers (approximately 30% total SCFA) and, from the measured acidogenic profile, production of SCFA will occur mainly in the proximal colon. The buffering capacity of ileal effluents during fermentation restrict the potential for a reduction in pH during acidogenesis compared to fiber isolates. This buffering capacity could limit the bioavailability of butyrate in the colon and, hence, the ability to satisfy the proposed antineoplastic properties of butyrate in the colon. PMID- 15281225 TI - Anti-tumor-promoting activities of selected pungent phenolic substances present in ginger. AB - Ginger (Zingiber officinale Roscoe, Zingiberaceae) has been widely used as a dietary spice, as well as in traditional oriental medicine. The rhizome of ginger contains pungent vanillyl ketones, including [6]-gingerol and [6]-paradol, and has been reported to possess a strong anti-inflammatory activity. These pungent substances have a vanilloid structure found in other chemopreventive phytochemicals, including curcumin. In our study, we found anti-tumor-promoting properties of [6]-gingerol and [6]-paradol. Thus, topical application of [6] gingerol or [6]-paradol 30 min prior to 12-O-tetradecanoyl-phorbol-13-acetate (TPA) attenuated the skin papillomagenesis initiated by 7,12 dimethylbenz[a]anthracene in female ICR mice. These substances also significantly inhibited the tumor-promoter-stimulated inflammation, TNF-alpha production, and activation of epidermal ornithine decarboxylase in mice. In another study, [6] gingerol and [6]-paradol suppressed the superoxide production stimulated by TPA in differentiated HL-60 cells. Taken together, these findings suggest that pungent vanilloids found in ginger possess potential chemopreventive activities. PMID- 15281227 TI - Antimutagenic/antioxidant activity of green tea components and related compounds. AB - The ability of green tea components and other antioxidant compounds to function as antimutagens/antioxidants has been well established, and their role in cancer prevention is supported by numerous epidemiological studies. We have utilized modified Ames tests, superoxide scavenging assays, and assays for protection against DNA scissions to compare and contrast the protective effects of various teas and commercial and laboratory-isolated tea components to those produced by compounds such as resveratrol, selenium, curcumin, vitamins C and E, quercetin dihydrate, sulforaphane, ellagic acid dihydrate, glutathione reduced, trolox, butylated hydroxanisole (BHA), butylated hydroxytoluene (BHT), and N-acetyl-L cysteine (NAC). In Ames tests, employing hydrogen peroxide as a mutagen, epigallocatechin gallate (EGCG) produced the highest level of protection of all antioxidants tested. Measurement of protection against DNA scissions produced results that again showed that EGCG produced the strongest protective effects. In scavenging assays using a xanthine-xanthine oxidase (enzymatic system), epicatechin gallate (ECG) showed the highest scavenging potential. In a nonenzymatic (phenazine methosulfate-NADH) oxidizing system, EGCG once again showed the strongest effects. The implications of these and similar results are discussed in relation to cancer prevention and prevention of drug/antibiotic resistance. PMID- 15281228 TI - The protective effect of vitamins C and E against B(a)P-induced genotoxicity in human lymphocytes. AB - Polycyclic aromatic hydrocarbons (PAH) are a well-characterized group of mutagens and carcinogens. Benzo(a)pyrene [B(a)P], the best known compound in the group, exerts its genotoxic activity following metabolic activation, when it acquires the properties of an electrophilic reagent that is capable of interacting with DNA. Reactive oxygen species (ROS) can remerge during the PAH metabolic activation. Because of their antioxidant activity, vitamins C and E are thought to act as antimutagenic agents. We designed an in vitro protocol to study the potential protective effect of vitamins C and E toward B(a)P-induced DNA damage. In this study, we examined peripheral blood lymphocytes obtained from healthy nonsmoking female volunteers, 22 to 25 years of age. The cells were exposed in vitro to 1 microM B(a)P in the presence of 40 microM or 100 microM of vitamin C or, alternatively, to 30 microM or 100 microM of vitamin E. The B(a)P-induced DNA damage and repair were estimated as the generation and removal of single-strand DNA breaks measured by the alkaline version of the single-cell gel electrophoresis (comet) assay. The protective effect of vitamins C and E was demonstrated when the vitamins were applied simultaneously with or after the B(a)P. The background level of DNA damage in the presence of vitamins C and E was lower than in the system without the vitamins. The experiments were conducted according to various protocol schemes of the vitamin treatment and the results offer additional evidence of the antigenotoxic activity of vitamins C and E. The vitamin activity does not appear to be connected with the steps in metabolic activation or DNA repair. It seems that both vitamins act as competitors of DNA molecule in reaction with the reactive oxygen species. PMID- 15281229 TI - Effects of pirfenidone on the generation of reactive oxygen species in vitro. AB - Pirfenidone is a newly developed antifibrotic drug that has been reported to retard the progression of pulmonary fibrosis induced by bleomycin and cyclophosphamide in animal models of lung fibrosis. The present in vitro studies using noncellular and cellular systems evaluated the antioxidant and cytotoxic properties of this drug. The Fenton reaction [Fe(II) + H2O2 --> Fe(III) + *OH + OH-] and the xanthine/xanthine oxidase system were used as sources of hydroxyl (*OH) and superoxide anion (O2*-) radicals, respectively. Electron spin resonance spin trapping was used for free radical detection and measurement. The reaction rate of pirfenidone with *OH was found to be 1.63 x 10(10) M(-1) s(-1), which is comparable to several well-established antioxidants, such as ascorbate, glutathione, cysteine, azide, and lipoic acid. Compared to *OH radicals, the O2*- scavenging was less efficient 42.36 M(-1) s(-1) with pirfenidone. Pirfenidone was also effective in inhibiting zymosan-stimulated chemiluminescence. In a noncellular model of lipid peroxidation, pirfenidone inhibited crystalline silica induced lipid peroxidation. The inhibition of crystalline silica-induced cytotoxic reactions and lipid peroxidation combined with the efficient antioxidant properties of pirfenidone indicate that this agent may express its antifibrotic effects partly through its ability to scavenge reactive oxygen species. PMID- 15281230 TI - Apoptosis of lymphocytes in canine peripheral blood induced by shock vibration and its mechanism. AB - We investigated the mechanism of apoptosis induced by shock vibration in canine peripheral lymphocytes, the T-lymphocyte changes, and the expression of p53 and bax gene products related to apoptosis using the techniques of immuno- and enzyme cytochemistry. We noted obvious apoptosis after delivery of 80, 100, and 200 acceleration of gravity values (G values). The percentage of apoptotic lymphocytes was directly proportional to the G value. On the 3rd day after injury, the number of apoptotic lymphocytes reached the peak value, which was about 5 to 8 times the amount in the control group. On the contrary, on day 3 after injury, T lymphocytes decreased and were about 50% of the control group. On the other hand, we found that the percentage of p53 and bax-positive lymphocytes distinctly increased and, on the 3rd day after injury, their number was, respectively, about 2.3 and 1.8 times that in the control groups, suggesting that they may play an important role in lymphocyte apoptosis. The above-mentioned results provide an important basis for further study of the mechanism of shock vibration injury, its prevention, and treatment. PMID- 15281231 TI - Apoptosis of circulating lymphocytes induced by whole body gamma-irradiation and its mechanism. AB - We studied the apoptosis of mousecirculating lymphocytes and its mechanism induced by 2, 4, 6, and 8 Gy of whole-body gamma-irradiation and the expression of bax and bcl-2 gene products as related to apoptosis. We found that, in the early stage after irradiation (4th-7th day), the percentage of lymphocyte apoptosis increased rapidly. Four hours after 2, 4, 6, and 8 Gy of gamma irradiation, the apoptotic lymphocytes were 2.6, 3.8, 5.5, and 10.4 times those in the controls, respectively. A good correlation was found between the intensity of apoptosis and the radiation dose. As the radiation dose increased, the absolute counts of peripheral lymphocytes decreased sharply; 4 hours after 2, 4, 6, and 8 Gy of gamma-irradiation, the lymphocyte counts were 82, 63, 47, and 22% of the controls, respectively. The peak value of lymphocyte apoptosis was observed on the 7th day after irradiation using the in situ terminal labeling method, and the apoptotic lymphocytes were found to be 16 times the number in the controls. These results were in accordance with those obtained by the May Grunwald-Giemsa staining method. The absolute counts of peripheral lymphocytes dropped to their lowest value on the 7th day after irradiation, suggesting that lymphocyte apoptosis might be the major cause of lymphocytopenia in the early stage after irradiation. The abnormal expression of bax and bcl-2 gene products in irradiated lymphocytes was closely related to apoptosis in peripheral lymphocytes. PMID- 15281232 TI - Comparison of the phenylenediamine isomers bioactivation by the green alga Chlamydomonas reinhardtii. AB - The correlation between the chemical structure of arylamines meta-, orto-, para phenylenediamine (m-PDA, o-PDA, p-PDA), and their mutagenic activity is known. It is accepted that these promutagenic compounds are metabolized to ultimate mutagens in mammals and higher plants. In our previous work, we used the alga Chlamydomonas reinhardtii as the activating organism and the bacteria Salmonella typhimurium and yeast Saccharomyces cerevisiae as the genetic indicators for m PDA activation. In the present work, we used the same activation system for o-PDA and p-PDA activation. Different responses of the yeast and algal wild-type strain and of the repair-deficient strains to the toxic and mutagenic effects of o-PDA and p -PDA were observed. p-PDA had the most toxic effect on both intact yeast and algal cells and in the algal cell/microbe coincubation assays. Concerning repair-deficient algal strains, the recombination-deficient strain was the most sensitive to both compounds tested, indicating that the recombination process played an important role in the DNA repair of arylamines. The rank order of the PDA isomers mutagenicity (including m-PDA) was o-PDA > m-PDA > p-PDA for revertants in intact yeast and forward mutants in algae; m-PDA > o-PDA > p-PDA in the algal cell/S. typhimurium long-term coincubation assay, the algal cell/S. cerevisiae coincubation assay, and the intact S. cerevisiae assay for gene convertants as well. PMID- 15281233 TI - Selective effect of malathion on blood coagulation versus locomotor activity. AB - The objective of this study was to compare the neurotoxic effects of acute malathion exposure with the potential blood coagulation effects. We administered various doses of malathion to Wistar rats by i.p. injection, and then we assessed the effects on blood coagulation screening tests and activity in the open field locomotor test. The activated partial thromboplastin time (APTT), but not the prothrombin time (PT), was significantly prolonged by up to 6.6 seconds (38% of the preinjection value) after the administration of malathion (3-75 mg/kg; 1-25% of the LD50). This effect was observed 5 minutes after injection and persisted for at least 3 hours. In contrast, no effect was observed on the general locomotor activity, movement, rearing, or stereotopy during 3 hours after the i.p. administration of the same doses of malathion. Our data indicate that malathion affects blood clotting before the nervous system (locomotor) function. PMID- 15281234 TI - Modeling and experimental comparison of the differential adsorption of B1 and G1 aflatoxins on mineral aluminosilicate surfaces. AB - Aflatoxins produced by Aspergillus fungus are secondary metabolites undergoing biotransformation rates in hepatic tissues and lipid peroxidation. Although the use of adsorbent materials became a common practice for feed grain detoxification, fundamental studies are needed to clarify the interaction occurring between mineral surfaces and organic molecules. We evaluated the differential adsorption of B1 and G1 on 10 adsorbent materials and compared it in vitro by means of fluorescence emission from solution. Three aluminosilicates showed no adsorption of B1 at all, whereas only one was inactive for G1 adsorption and seven of them showed 15.2 to 77.9% adsorption for B1 and 8.3 to 78% for G1. All these adsorbents were more selective toward G1 rather than B1 aflatoxins. This behavior can be explained by the presence of an additional cyclic ester in G1, which provides a higher electronic density to G1 molecules, thus forming more stable hydrogen bridges with respect to the cyclopentanone ring present in B1. PMID- 15281235 TI - Regulatory molecules in tumor metastasis. AB - The most life-threatening aspects of cancer are invasion and metastasis. The phenomena of metastasis is a multistep process of host tumor interactions. In the present study, we wanted to identify regulatory molecules in tumor metastasis. Our comparative studies between highly metastatic B16F10 and low metastatic B16F1 tumor cells strongly indicate that the function of vitronectin integrin receptor (alphavbeta3) and collagenase enzyme activity (72 kDa) are two of the key factors that control the invasive and metastatic properties of B16 F10 melanoma cells. The decreased expression of nm23 gene product, TIMP-2, and E-cadherin and the increased expression of pp125FAK in highly metastatic B16F10 tumor cells indicate their potential role in metastatic cascade. PMID- 15281236 TI - Experimental hepatic tumorigenicity by environmental hydrocarbon dibenzo[a,l]pyrene. AB - There is an evident need of low-cost vertebrates to be used in experimental carcinogenesis. Medaka (Oryzias latipes) provide a useful vertebrate model system for investigating tissue tropism of carcinogens and the action mechanisms of environmental contaminants posing a potential risk to human health. Juvenile medaka 2 months of age fed diets containing 100 ppm (dry weight basis) dibenzo[a,l]pyrene (DBP) for 28 days responded with hepatic neoplasia predominately of hepatocellular origin. When sampled 9 months following the termination of carcinogen exposure, medaka showed 26% incidence of neoplasia and 25% hepatic neoplasia, compared with 8% total neoplasia and 0% hepatic neoplasia in control fish. The predominant spontaneous neoplasms in this group of medaka were ovarian germ cell tumors. Hepatic neoplasia occurred at a higher incidence in female DBP-treated medaka than in males (11/29 vs 5/36). Nonneoplastic lesions observed in the livers of DBP-exposed fish included spongiosis hepatis, globular hyaline eosinophilic cytoplasmic inclusions in hepatocytes, foci of hepatocellular degeneration, extensive cytomegaly, and karyomegaly of hepatocytes. No activating exon I mutations in the one ras protooncogene examined were detected among six liver neoplasms. These results indicate that medaka are sensitive to the tumorigenic effects of the environmental carcinogen DBP, administered by dietary exposure. PMID- 15281237 TI - Curcumin damages DNA in human gastric mucosa cells and lymphocytes. AB - The naturally occurring pigment curcumin, a major component of the spice turmeric, is reported to be a potent inhibitor of the initiation and promotion of many cancers. Due to its presence in the diet, one of its primary targets is the human gastric mucosa (GM) cells. Using the sensitive single cell electrophoresis method (comet assay), we found that curcumin at of 15, 25, and 50 microM caused DNA damage in GM cells and human peripheral blood lymphocytes. There was no difference between the extent of the damage in both types of cells. Damaged cells were able to recover within a period of 120 minutes. Our results indicate that curcumin may play a dual role in carcinogenesis. PMID- 15281238 TI - Influence of cadmium intoxication on hepatic lipid peroxidation, glutathione level, and glutathione S-transferase and gamma-glutamyl transpeptidase activities: correlation with chromosome aberrations in bone marrow cells. AB - We examined whether there was any correlation between chromosome aberrations (CAs) in bone marrow cells with hepatic lipid peroxidation (LPO), reduced glutathione (GSH) level, glutathione S-transferase (GST), and gamma-glutamyl transpeptidase (GGT) activity after cadmium (Cd) intoxication in both a dose- and a time-dependent manner. Cadmium chloride was administered subcutaneously in doses of 0.5, 1.5, 2.5, and 5.0 mg/kg body weight to Swiss albino Balb/c male mice. The animals were exposed for 8, 16, and 24 days, i.e., 4, 8, and 12 doses, respectively. Biochemical parameters were measured in hepatic tissue for a correlation with chromosome aberrations in bone marrow. With the increment of dose and advancement of time points, the biochemical, as well as the cytogenetic, parameters altered significantly. Hepatic lipid peroxidation and GGT activity increased significantly along with an increased percentage of chromosome aberrations in the bone marrow, but the hepatic reduced glutathione level and GST activity were found to decrease following Cd administration. Up to 5.0 mg Cd/kg body weight, lipid peroxidation did not exhibit threshold levels of toxicity as shown by the two-way (fixed effect) analysis of variance test. In contrast, the observed values of reduced glutathione levels, GST and GGT activity, and chromosome aberrations in bone marrow showed threshold activity levels. Therefore, there might be a relationship between an increase in the frequency of chromosome aberrations, elevated lipid peroxidation, and depleted glutathione levels and GST and GGT activity. The clastogenic efficacy of Cd may be mediated through the biochemical pathways. PMID- 15281239 TI - Apoptosis of the intestinal principal cells of Xenopus larvae exposed to 2,3,7,8 tetrachlorodibenzo-p-dioxin. AB - In our previous work using paraffin-embedded sections, we determined that Xenopus larvae exposed to 200 ppb 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) for 5 days from shortly after fertilization to the early larval stage showed a shortening of the digestive tract and a loss of mucosal epithelium cells due to exfoliation into the gut cavity. In the current study, we ultrastructurally examined the mucosal epithelium of the gut of TCDD-exposed Xenopus larvae 12 days after fertilization. Exfoliated cell structures at the villus tip and in the lumen were equipped with a microvillus portion and occasionally had terminal web-like structures seen by ultramicroscopy. As these exfoliated structures had nuclear fragments with condensed chromatin, they were considered to be apoptotic bodies derived from the principal cells of the epithelium. In addition, many membrane bound cell fragments-identified as apoptotic bodies derived from the principal cells based on their ultrastructural features-were observed at the basal side of the mucosal epithelium. These apoptotic bodies were phagocytized and digested chiefly by the neighboring intact principal cells. No such cells and/or cell fragments showing apoptotic features were observed in the controls. Our observations indicate that marked apoptosis occurs in the intestinal principal cells of TCDD-exposed larvae, which may result in the shortening of the gut. PMID- 15281240 TI - Organochlorine insecticides and PCB residues in fat tissues of autopsied trauma victims in Israel: 1984 to 1986. AB - Between the 1950s and the 1970s, large quantities of organochlorines were used as insecticides in Israel's cowsheds, causing massive contamination of Israel's milk supply. In 32 of 40 autopsied Israeli trauma victims who died between 1984 to 1986, we found three or more types of organochlorines in adipose tissue, and at least one organochlorine residue in all 40 individuals. The cumulative mean organochlorine levels (ppm) in men and women were as follows: DDE = 5.73 and 4.36; DDT = 0.12 and 0.30; HCB = 0.31 and 0.11; beta-HCH = 0.53 and 0.43; PCB = 0.10 and 0.08, respectively. PCB residue levels in fat tissue were lower than those noted in other countries and below the adverse levels associated with health effects. The cumulative mean organochlorine levels in adipose tissue were higher in men than in women in all cases except for DDT. Mean levels for DDE + DDT combined was higher in older than younger people, and all persons' DDT/DDE ratios were less than 1. The DDE levels in adipose tissue were higher than the levels reported in many other countries during the 1970s, 1980s, and 1990s. Our findings suggest that population-wide exposures to organochlorine insecticides come not only from contaminated milk products but other food products as well. The amounts of organochlorines in adipose tissue are compatible with those found in food residue samples during the same period. PMID- 15281241 TI - Apoptosis of hemopoietic cells in irradiated mouse bone marrow. AB - Apoptosis of hemopoietic cells in bone marrow following radiation has been rarely reported, let alone studied quantitatively and pathologically. LACA mice were irradiated with 2.5, 4.0, 5.5, and 7.0 Gy gamma-rays, and the bone marrow was examined at 6 hours, 1 day, and 3 days after radiation. Semi-thin and thin sections were examined by light and electron microscopy. The number and area density of the apoptotic cells were assessed by means of a Cambridge Quantimet 970 Image Analyzer. We found that apoptosis occurred in only a few hemopoietic cells in the control mouse bone marrow, whereas 6 hours after radiation there were many apoptotic hemopoietic cells in each sample of irradiated bone marrow. Compared with controls, both the number and area density of the apoptotic cells markedly increased in the bone marrow of animals in every radiation dose group, and the difference was statistically significant (p < 0.01). Furthermore, the number and area density increased as the radiation dose increased. Our findings suggest that apoptosis is the main mode of radiation-induced hemopoietic cell death. PMID- 15281242 TI - Overexpression of c-fos and Rb proteins in radiation-induced skin ulcers. AB - We established an animal model of radiation-induced skin ulcers in rats irradiated with 35 to 55 Gy y-rays. The pathological changes were observed for 1 year. Immunohistochemical studies were performed on 72 radiation-induced skin ulcer specimens using c-fos and Rb protein polyclonal antibodies. We found that the overexpression rate of c-fos was 45.8% and of Rb was 63.9%. The overexpression of c-fos was mainly seen in the nuclei of activated squamous epithelial cells and in some fibroblasts and endothelial cells of arterioles in the deep part of the skin ulcers. The overexpression of Rb had the same localization. Our results suggest that the changes in c-fos and Rb proteins may be related to the poor healing of radiation-induced skin ulcers. PMID- 15281243 TI - Overexpression of c-erbB-2 and EGF-R proteins in radiation-induced skin ulcers. AB - We established an animal model of radiation-induced skin ulcers in rats irradiated with 35-55Gy gamma-rays. The pathological changes were studied for 1 year. Immunohistochemical studies were performed on 72 radiation-induced skin ulcer specimens using c-erbB-2 and epidermal growth factor receptor (EGF-R) protein antibodies. We found that the overexpression rate of c-erbB2 oncoprotein was 59.7% and of EGF-R was 70.8%. The overexpression of c-erbB-2 was chiefly seen in the activated squamous epithelial cells and in the cytoplasm of fibroblasts, endothelial cells, and leiomyocytes of the arteriolar media in the deep part of the ulcers. The overexpression of EGF-R had the same localizations. It is suggested that the overexpression of c-erbB-2 and EGF-R proteins may be due to the poor healing of the radiation-induced skin ulcers. PMID- 15281244 TI - Overexpression of p53 and MDM2 proteins in rat radiation-induced skin ulcers. AB - We established an animal model of radiation-induced skin ulcer in rats that were locally irradiated with 35 to 55 Gy gamma-rays. The pathological changes were observed for 1 year. Immunohistochemical studies using p53 and MDM2 protein polyclonal antibodies were performed on 72 radiation-induced skin ulcer specimens. The results showed that the overexpression rate of p53 protein was 9.7% and of MDM2 was 19.4%. The overexpression of p53 was chiefly seen in the nuclei of the activated squamous epithelial cells and in the fibroblasts and endothelial cells of arterioles in the deeper part of the skin ulcers. The overexpression of MDM2 had the same localizations. These results suggest that the changes of p53 and MDM2 may be related to the poor healing rate of radiation induced skin ulcers. PMID- 15281245 TI - Transmission electron microscopy findings in the respiratory epithelium of guinea pigs exposed to the polluted air of southwest Mexico City. AB - We evaluated the nature and the extent of the damage to the respiratory epithelium of guinea pigs after a 4-month exposure to the mixture of air pollutants in southwest Mexico City. Guinea pigs were placed outdoors on the roof of our facility, 8 hours daily, from February to May 1995. At the same time, control guinea pigs were kept indoors breathing filtered air. Air pollutants, temperature, and humidity data were obtained from the nearest station of the Environmental Monitoring Net. The airways and lung parenchyma were analyzed after 120 days using transmission electron microscopy (TEM). During the 4-month exposure period, ozone (O3) exceeded the norm during 511 hours, and suspended particles less than 10 microm in diameter (PM10) during 52 hours. Both pollutants reached peak levels of more than twice the norm. TEM revealed no important abnormalities in the control group. In the exposed group, there was loss of cilia, detachment of epithelial cells, and eosinophil and macrophage migration toward alveolar spaces through type I pneumocytes with destruction of their basal membranes. In six guinea pigs in the exposed group, we noted bacteria along the airways, with associated inflammatory response. We explain the colonization of the respiratory epithelium by bacteria as the result of the impairment on the defense mechanism caused by the exposure to environmental O3 and PM10. PMID- 15281246 TI - Retrospective biomonitoring: a hundred years of environmental pollution at selected areas in Slovakia. AB - Tetrade analyses of Calluna vulgaris from herbarium specimens showed significant differences in the frequency of aborted pollen tetrads over the last 100 years within various parts of Slovakia. Specifically, we observed changes in the dynamics of pollution peaking in the year 1965 in heavily polluted industrial area of an aluminium factory in Ziar nad Hronom. PMID- 15281247 TI - Nongonadal neoplasia in patients with Turner syndrome. AB - In a prospective study of 50 patients with retinoblastoma, a 10-year-old girl with unilateral (right eye) retinoblastoma was found to have 45,X karyotype. Because there is increasing evidence of nongonadal neoplasia occurring in patients with Turner syndrome in addition to the gonadal tumor from dysgenetic gonads, we reviewed the occurrence of nongonadal neoplasia in Turner syndrome cases. Of all the nongonadal neoplasia, neurogenic tumors show a preponderence among children and young adults with Turner syndrome. To the best of our knowledge, this is the first reported case of Turner syndrome with retinoblastoma. The available literature strongly suggests that patients with Turner syndrome may be at risk of developing neurogenic tumors. Further studies are necessary to identify the role of some X-linked genes that escape X inactivation in tumorigenesis in patients with Turner syndrome. PMID- 15281248 TI - Novel fiber coated with amide bridged-calix[4]arene used for solid-phase microextraction of aliphatic amines. AB - Solid-phase microextraction (SPME) using a novel fiber coated with 25,27 dihydroxy-26, 28-oxy(2',7'-dioxo-3',6'-diazaoctyl)oxy-p-tert butylcalix[4]arene/hydroxy-terminated silicone oil has been introduced as a rapid and sensitive pretreatment technique coupled to gas chromatography-flame ionization detection (GC-FID) for the detection of aliphatic amines without derivatization. Due to the introduction of the polar amide bridge in calix[4]arene, the new fiber shows good selectivity and sensitivity to the polar aliphatic amines in addition to its high thermal stability (380 degrees C), solvent stability and good reproducibility between fibers. The extraction temperature, extraction time, pH, and ionic strength of the matrix sample were modified to allow for maximum sorption of the analytes onto the fiber. The method proposed in this study showed satisfactory linearity, precision and detection limits. Practical applicability was demonstrated through the determination of trimethylamine (TMA) in fish tissue. Mean recovery of 92.5% (n = 5) was obtained for the fish extracts and the relative standard deviation was 4.9% (n = 5). The results of fish freshness assay indicate the present method is a validated and simple procedure for the simultaneous determination of TMA in fish. PMID- 15281249 TI - Analytical procedure for the in-vial derivatization--extraction of phenolic acids and flavonoids in methanolic and aqueous plant extracts followed by gas chromatography with mass-selective detection. AB - An in-vial simple method for the combined derivatization and extraction of phenolic acids and flavonoids from plant extracts and their direct determination with GC-MS, is described. The method is taking advantage of the beneficial potentials of phase transfer catalysis (PTC). Catalysts in soluble and polymer bound form were tested with the latter being the format of choice due to its high reaction yield and facile separation from the rest of the reaction system. Optimization of experimental conditions was established. Chromatographic separation of eight phenolic acids and four flavonoids methylated via the PTC derivatization step was achieved in 45 min. The detection limits for the described GC-MS(SIM) method of analysis ranged between 2 and 40 ng/ml whereas limits of quantitation fall in the range 5-118 ng/ml, with flavonoids accounting for the lowest sensitivity due to their multiple reaction behavior. Four methanolic extracts from Tilia europea, Urtica dioica, Mentha spicata and Hypericum perforatum grown wild in north-western Greece and four aquatic infusions from commercially available Mentha spicata, Origanum dictamnus, Rosemarinus officinalis and Sideritis cretica were analyzed. Good trueness of the method was demonstrated as no matrix effects were found for the analytes concerned. PMID- 15281250 TI - Trace analysis of phenolic xenoestrogens in water samples by stir bar sorptive extraction with in situ derivatization and thermal desorption-gas chromatography mass spectrometry. AB - A method for the simultaneous measurement of trace amounts of phenolic xenoestrogens, such as 2,4-dichlorophenol (2,4-DCP), 4-tert-butyl-phenol (BP), 4 tert-octylphenol (OP), 4-nonylphenol (NP), pentachlorophenol (PCP) and bisphenol A (BPA), in water samples was developed using stir bar sorptive extraction (SBSE) with in situ derivatization followed by thermal desorption (TD)-gas chromatography-mass spectrometry (GC-MS) analysis. The conditions for derivatization with acetic acid anhydride were investigated. A polydimethylsiloxane (PDMS)-coated stir bar and derivatization reagents were added to 10 ml of water sample and stirring was commenced for 10-180 min at room temperature (25 degrees C) in a headspace vial. Then, the extract was analyzed by TD-GC-MS. The optimum time for SBSE with in situ derivatization was 90 min. The detection limits of 2,4-DCP, BP, OP, NP, PCP and BPA were 2, 1, 0.5, 5, 2 and 2 pg ml(-1), respectively. The method showed good linearity over the concentration ranges of 10, 5, 2, 20, 10 and 10-1000 pg ml(-1) for 2,4-DCP, BP, OP, NP, PCP and BPA, respectively, and the correlation coefficients were higher than 0.99. The average recoveries of those compounds in river water samples were equal to or higher than 93.9% (R.S.D. <7.2%) with correction using the added surrogate standards. This simple, accurate, sensitive and selective method can be used in the determination of trace amounts of phenolic xenoestrogens in river water samples. PMID- 15281251 TI - Determination of quinolones in water samples by solid-phase extraction and liquid chromatography with fluorimetric detection. AB - A method is reported for the determination, in water samples, of 10 quinolones which are used as veterinary drugs. Analytes are isolated from samples by solid phase extraction (SPE) and analysed by reversed-phase high-performance liquid chromatography using fluorimetric detection. A solid-phase extraction procedure based on retention on HBL OASIS cartridges and elution with a mixture of acetonitrile-water in basic medium is suitable for pre-concentration of the analytes. Pre-concentration factors up to 250 can be obtained. The quinolones are separated with an octyl silica-based column and mobile phases consisting of aqueous oxalic acid solutions and acetonitrile mixtures. The attained detection limits of the whole process are in the ng l(-1) level when 250 ml of water sample is processed. Recovery rates, from natural water samples spiked at 2060 ng l(-1) level, range from 70 to 100% and common standard deviation are about 6-12%. PMID- 15281252 TI - Optimization of a matrix solid-phase dispersion method with subsequent clean-up for the determination of ethylene bisdithiocarbamate residues in almond samples. AB - A matrix solid-phase dispersion (MSPD) method with subsequent clean-up has been developed to isolate the ethylene bisdithiocarbamate (EBDC) main metabolites (ethylenethiourea, ETU, and ethylenebis [isothiocyanate] sulphide, EBIS) in almond samples. The optimized experimental set-up configuration involved 0.2 g of almond sample, washed sand as MSPD support and NaOH as defatting agent. A subsequent purification step on alumina using acetonitrile as extraction solvent was enough to remove all interferent matrix components, including the fatty material, and provide clean extracts. Quantitative analysis was performed by reversed phase liquid chromatography (RPLC) with diode-array ultraviolet absorbance (DAD UV) detector. Analytes recoveries were between 76 and 85% with relative standard deviations ranging from 3 to 12%. The low limits of quantification of 0.05 and 0.07 mg kg(-1) achieved for ETU and EBIS, respectively, make the method useful for the determination of EBDC residues on almond samples. PMID- 15281253 TI - Accurate prediction of the retention behaviour of polydisperse macromolecules based on a minimum number of experiments. AB - This study illustrates how retention models can be used to accurately predict the retention behaviour of polydisperse macromolecules in LC separations. It highlights that the number of experiments required can be drastically reduced when the relationship between the model parameters and molecular structure parameters (e.g. molar mass) can be incorporated into one global model. A practical implication of this work is that an appropriate model can then be used for the determination of molar-mass distributions for polydisperse samples. The globalised model can predict retention time as a function of molar mass and gradient slope. Both the original and globalised versions of the model were rigorously validated in terms of the difference between the predicted and experimental retention times. The original model had very low residuals and there was no apparent dependence of the errors on the applied gradient, the molar mass or the retention times. Confidence intervals for the model parameters (S and ln k0) were determined using a bootstrapping analysis of the residual errors in the predicted retention times. Confidence intervals were seen to broaden significantly as the mass of the polymer increased. The parameters were also seen to be highly correlated. For the global model, retention-time residuals remained quite low, even when the number of experiments used to determine the model parameters was reduced from approximately 100 to 10. PMID- 15281254 TI - Analysis of the isolation of a target component using multicomponent isocratic preparative elution chromatography. AB - The separation of a certain target component from a multicomponent mixture using isocratic preparative elution chromatography was studied theoretically. In particular, the important and most complicated case was considered that the target component does not elute in the first or last position. To specify the productivity of collecting this component different options are suggested to identify suitable times for fractionation. Using a conventional Craig model, capable to quantify chromatographic processes, the impact of several essential parameters (e.g. threshold concentration, desired purity, injection volume, separation factor between neighboring components, composition of the mixture) is evaluated for a ternary system based on parametric calculations. The paper provides simple tools to evaluate and optimize the productivity and other objective functions relevant in multicomponent preparative chromatography. PMID- 15281255 TI - Effect of the pH, the concentration and the nature of the buffer on the adsorption mechanism of an ionic compound in reversed-phase liquid chromatography. II. Analytical and overloaded band profiles on Symmetry-C18 and Xterra-C18. AB - In a previous report, the influence of the pH, the concentration, and the nature of the buffer on the retention and overloading behavior of propranolol (pKa = 9.45) was studied on Kromasil-C18 at 2.75 < pH < 6.75, using four buffers (phosphate, acetate, phthalate, and succinate), at three concentrations, 6, 20, and 60 mM. The results showed that the propranolol cation was eluted as an ion pair with the buffer counter-anion. A similar study was carried out with Symmetry C18 and Xterra-C18. Two additional buffers, formate and citrate, were also used. Propranolol elution band profiles were recorded for a small (less than 1 microg) and a large (375 microg) sample size. The results are similar to those obtained with Kromasil and confirm earlier conclusions. The buffer concentration, not its pH, controls the retention time of propranolol, in agreement with the chaotropic model. The retention factor depends also on the nature of the buffer, particularly on its valence, and on the hydrophobicity of the basic anion. With the monovalent anions HCOO- (pH 3.75), H2PO4- (pH 2.75), HOOC-Ph-COO- (pH 2.75), HOOC-CH2-CH2-COO- (pH 4.16), CH3COO- (pH 4.75) and HOOC-CHCOOH-COO- (pH 3.14), at moderate loadings, and for the two larger buffer concentrations, the band profiles are well accounted for by a simple bi-Langmuir isotherm model (no adsorbate-adsorbate interactions). By contrast, these profiles are accounted for by a bi-Moreau isotherm model (i.e., with significant adsorbate-adsorbate interactions) with the bivalent anions -OOC-Ph-COO- (pH 4.75), -OOC-CH2-CH2-COO- (pH 5.61), HPO4(2-) (pH 6.75), and HOOC-CHCOO(-)-COO- (pH 4.77) and with the trivalent anion -OOC-CHCOO(-)-COO- (pH 6.39). The best values of the isotherm parameters were determined using the inverse method. The saturation capacity and the equilibrium constant on the low-energy sites increase with increasing buffer concentration, a result consistent with the formation in the mobile phase of a hydrophobic complex between the propranolol cation and the buffer anion. With bivalent and trivalent anions, adsorbate-adsorbate interactions are strong on the low-energy sites but they remain negligible on the high-energy sites. The density of the high energy sites is lower and the equilibrium constant on the low-energy sites are both higher with the bivalent and the trivalent buffer anions than with the univalent buffer anions. These results are consistent with the formation of a 2:1 and a 3:1 propranolol-buffer complex with the bivalent and the trivalent anions, respectively. PMID- 15281256 TI - Modeling chromatographic columns non-equilibrium packed-bed adsorption with non linear adsorption isotherms. AB - In this work a new mathematical model, based on non-equilibrium conditions, describing the dynamic adsorption of proteins in columns packed with spherical adsorbent particles is used to study the performance of chromatographic systems. Simulations of frontal chromatography, including axial dispersion, for non equilibrium systems with non-linear adsorption isotherms are made and compared to those of the experimentally determined protein A affinity chromatography breakthrough curves of hIgG, gathered from the literature. The non-equilibrium model developed here combines external mass transfer and intra-particle transport by solid (surface) diffusion, and permits the prediction of (time and bed height dependent) interface and average solid concentrations, along with interface and bulk liquid concentrations. The present non-equilibrium approach significantly improved the model predictions of experimentally observed distended breakthrough fronts over local equilibrium based models, and can be used to evaluate the influence of system parameters on the performance of chromatographic packed-bed adsorption columns. PMID- 15281257 TI - Preparative and analytical chromatography of pegylated myelopoietin using monolithic media. AB - Monolithic media were compared with Q- and SP-Sepharose high performance chromatography for preparative purification and with Q- and SP-5PW chromatography for analysis of a pegylated form of myelopoietin (MPO), an engineered hematopoietic growth factor. The use of either monolithic or Sepharose based supports for preparative chromatography produced highly purified pegylated MPO with the monolithic media demonstrating peak resolution and repeatability at flow rates of 1 and 5 ml/min resulting in run times as much as five-fold shorter compared to Sepharose separations. The monolithic disks also resulted in 10-fold shorter run times for the analytical chromatography, however, their chromatographic profiles and peak symmetry were not as sharp compared to their Q 5PW and SP-5PW counterparts. PMID- 15281258 TI - Ion chromatographic determination of hydroxide ion on monolithic reversed-phase silica gel columns coated with nonionic and cationic surfactants. AB - The determination of hydroxide by ion chromatography (IC) is demonstrated using a monolithic octadecylsilyl (ODS)-silica gel column coated first with a nonionic surfactant (polyoxyethylene (POE)) and then with a cationic surfactant (cetyltrimethylammonium bromide (CTAB)). This stationary phase, when used in conjunction with a 10 mmol/l sodium sulfate eluent at pH 8.2, was found to be suitable for the rapid and efficient separation of hydroxide from some other anions, based on a conventional ion-exchange mechanism. The peak directions and detection responses for these ions were in agreement with their known limiting equivalent ionic conductance values. Under these conditions, a linear calibration plot was obtained for hydroxide ion over the range 16 micromol/l to 15 mmol/l, and the detection limit determined at a signal-to-noise ratio of 3 was 6.4 micromol/l. The double-coated stationary phase described above was shown to be superior to a single coating of cetyltrimethylammonium bromide alone, in terms of separation efficiency and stability of the stationary phase. A range of samples comprising solutions of some strong and weak bases was analyzed by the proposed method and the results obtained were in good agreement with those obtained by conventional potentiometric pH measurement. PMID- 15281259 TI - Comprehensive two-dimensional separations of complex mixtures using reversed phase reversed-phase liquid chromatography. AB - A comprehensive two-dimensional reversed-phase reversed-phase liquid chromatographic system for the separation of a complex mixture of oligostyrenes was developed using results from a previous theoretical assessment of the informational similarity, percent synentropy, orthogonality and peak capacity of hypothetically coupled systems. The degree of sample attribute order in the first separation dimension was also used in the development of the experimental two dimensional system. A C18(methanol)/CCZ(acetonitrile) two-dimensional system was chosen for the comprehensive analysis of the oligostyrene mixtures because this system had the lowest solute crowding, highest orthogonality and was observed to have order with respect to a sample attribute in the first separation dimension. The separations achieved were in full agreement with the results from information theory and (a geometric approach to) factor analysis assessments. High sampling rates in the first liquid chromatographic dimension were shown to be impossible or inefficient when the peak capacity and separation time of the second dimension was high or when the aim of the exercise was to isolate individual sample constituents in high yield. PMID- 15281260 TI - Efficient isolation of polyaromatic fraction from aliphatic compounds in complex extracts using dimethylformamide-pentane partitionings. AB - A liquid-liquid partitioning method was optimized for the rapid and quantitative separation of polycyclic aromatic hydrocarbons (PAHs) and polychlorinated biphenyls (PCBs) from aliphatic hydrocarbons in complex primary extracts. This technique was based on the selective extraction of PAHs and PCBs from an aliphatic solvent into dimethylformamide (DMF). Partition experiments demonstrated that the optimal performance was achieved with a DMF (5% H2O)-n pentane binary system. The optimized application of two consecutive DMF (5% H2O) n-pentane treatments to extracts from two different polluted sediments facilitated the elimination of alkanes and unresolved complex mixture by more than 94% while the average recoveries of spiked deuterated-PAHs and 13C labeled PCBs ranged from 84 to 94 and 75 to 96%, respectively. PMID- 15281261 TI - Definition and system implementation of strategies for method development of chiral separations in normal- or reversed-phase liquid chromatography using polysaccharide-based stationary phases. AB - This paper proposes strategies in normal- and reversed-phase liquid chromatography (NP-HPLC or NPLC and RP-HPLC or RPLC), which were developed using three polysaccharide-based stationary phases. Those strategies are implemented in a knowledge-based system for the chiral separation of drug enantiomers. Each strategy includes a screening and an optimisation stage. The screening stage allows a fast evaluation of separation possibilities and enantioselectivity for many drugs in a short period of time, while the optimisation stage gives the opportunity to enhance, if needed, the initially obtained separation. Different examples demonstrate the effectiveness of the strategies for fast method development. PMID- 15281262 TI - Off-line coupling of non-aqueous reversed-phase and silver ion high-performance liquid chromatography-mass spectrometry for the characterization of rice oil triacylglycerol positional isomers. AB - The determination of the triacylglycerol (TAG) profile in real world matrices is rather difficult as these compounds present a complex composition and are characterized by similar physico-chemical properties. This investigation is based on the high-performance liquid chromatography (HPLC) multidimensional determination of the TAG profile in terms of TAG species and positional isomers in a rice oil sample. The off-line bi-dimensional system was attained through the coupling of non-aqueous reversed-phase HPLC and silver ion (Ag)-HPLC. The primary column eluate was fractionated and the fractions of interest were then injected onto the secondary column, allowing the separation of several TAG positional isomers, unresolved in the first dimension. Peak assignment was carried out by combining retention data with atmospheric pressure chemical ionisation (APCI) MS spectra information. The fatty acid distribution along the glycerol backbone, determined by Ag-HPLC, was confirmed through diglyceride ion ratios derived from APCI-MS analysis. Method validation, where both precision and accuracy were measured, was carried out in preliminary applications on standard compounds. The analytical results obtained show that rice oil TAGs follow a distribution which can be considered typical for vegetable oils. PMID- 15281263 TI - Separation of protoberberine quaternary alkaloids from a crude extract of Enantia chlorantha by centrifugal partition chromatography. AB - High-performance centrifugal partition chromatography (HPCPC) has been successfully applied to the separation of four protoberberine quaternary alkaloids, namely palmatine, jatrorrhizine, columbamine and pseudocolumbamine, from a methanolic extract (M1, 1.47 g) of Enantia chlorantha Oliver stem bark. For their isolation, two successive biphasic solvent systems composed of dichloromethane-methanol-water (48:16:36, v/v) were selected. The aqueous-rich phase was the stationary phase and the organic-rich phase was the mobile phase. The first system, containing potassium perchlorate, allowed to isolate 600 mg of palmatine, and to obtain 146 mg of a mixture (M2) containing only jatrorrhizine, columbamine and pseudocolumbamine. The second biphasic system, prepared with water alkalinized with sodium hydroxide, was employed to isolate the M2 components. This system applied to the purification of 70 mg of M2 allowed to obtain 16 mg ofjatrorrhizine and 13 mg of columbamine. To obtain pseudocolumbamine (16 mg), the elution mode was reversed, the aqueous-rich phase becoming the mobile phase, and the organic-rich phase becoming the stationary one. Analytical reversed-phase high-performance liquid chromatography, NMR, high resolution mass spectrometry and UV spectrometry were used to verify the identity and the purity of the isolated compounds. PMID- 15281264 TI - Preparative counter-current chromatography isolation of liensinine and its analogues from embryo of the seed of Nelumbo nucifera GAERTN. using upright coil planet centrifuge with four multilayer coils connected in series. AB - Preparative counter-current chromatography (CCC) isolation of liensinine and its analogues, isoliensinine and neferine from embryo of the seed of Nelumbo nucifera GAERTN. has been successfully performed for the first time using upright coil planet centrifuge with four multilayer coils connected in series with 1600 mL capacity. Two kinds of two-phase solvent systems were applied to preparative CCC isolation. The first was the system composed of light petroleum (b.p. 60-90 degrees C)-ethyl acetate-tetrachloromethane-chloroform-methanol-water (1:1:4:4:6:2, v/v) which was very suitable for fast and small-scale CCC isolation. The second was the system composed of ethyl acetate-tetrachloromethane methanol-water (1:6:4:1, v/v), which was the optimum for large-scale CCC isolation. Using the first system, 1102 mg of the crude alkaloid was purified in one-step separation of 150 min, yielding 350 mg neferine, 100 mg isoliensinine and 95 mg liensinine with over 95% purity. While using the second solvent system, 5850 mg of the crude alkaloid was purified in one-step separation of 9 h, yielding 2545 mg neferine, 698 mg isoliensinine and 650 mg liensinine with over 97% purity. Structures of the compounds were identified by electrospray ionization multiple mass spectrometry, one- and two-dimensional NMR. PMID- 15281265 TI - Chromatographic behavior of epirubicin and its analogues on high-purity silica in hydrophilic interaction chromatography. AB - Hydrophilic interaction chromatography has been applied for the separation of epirubicin and its analogues using high-purity silica column with aqueous-organic mobile phase. Parameters affecting the chromatographic behavior of the solutes such as organic modifier, buffer pH, ionic strength and sample size, have been investigated. Of utmost importance for successful separation of these analogues is the choice of organic modifier, since it impacts both the solvent selectivity and the ionization of silica silanols as well as buffer solution, and consequently the retention behavior of solutes. Acetonitrile was shown to offer superior separation of these analogues to methanol, isopropanol or tetrahydrofuran. Results of the effects of organic modifier, buffer pH and ion strength indicate that the retention mechanism is a mixed-mode of adsorption and ion exchange. In addition, an irreversible adsorption of these compounds was found on silica in the weakly acidic or neutral mobile phases, and the effect of various factors on irreversible adsorption was also preliminarily discussed. More significantly, these basic compounds have exhibited peaks with a slanted front and a sharp tail, a typical overloading peak profile belonging to the behavior of competitive anti-Langmuir isotherm by increasing the sample size at the experimental conditions. PMID- 15281266 TI - Determination of pharmaceutical compounds in surface- and ground-water samples by solid-phase extraction and high-performance liquid chromatography-electrospray ionization mass spectrometry. AB - Commonly used prescription and over-the-counter pharmaceuticals are possibly present in surface- and ground-water samples at ambient concentrations less than 1 microg/L. In this report, the performance characteristics of a combined solid phase extraction isolation and high-performance liquid chromatography electrospray ionization mass spectrometry (HPLC-ESI-MS) analytical procedure for routine determination of the presence and concentration of human-health pharmaceuticals are described. This method was developed and used in a recent national reconnaissance of pharmaceuticals in USA surface waters. The selection of pharmaceuticals evaluated for this method was based on usage estimates, resulting in a method that contains compounds from diverse chemical classes, which presents challenges and compromises when applied as a single routine analysis. The method performed well for the majority of the 22 pharmaceuticals evaluated, with recoveries greater than 60% for 12 pharmaceuticals. The recoveries of angiotensin-converting enzyme inhibitors, a histamine (H2) receptor antagonist, and antihypoglycemic compound classes were less than 50%, but were retained in the method to provide information describing the potential presence of these compounds in environmental samples and to indicate evidence of possible matrix enhancing effects. Long-term recoveries, evaluated from reagent-water fortifications processed over 2 years, were similar to initial method performance. Method detection limits averaged 0.022 microg/L, sufficient for expected ambient concentrations. Compound-dependent matrix effects on HPLC/ESI-MS analysis, including enhancement and suppression of ionization, were observed as a 20-30% increase in measured concentrations for three compounds and greater than 50% increase for two compounds. Changing internal standard and more frequent ESI source maintenance minimized matrix effects. Application of the method in the national survey demonstrates that several pharmaceuticals are routinely detected at 0.010-0.100 microg/L concentrations. PMID- 15281267 TI - Development of a high-performance liquid chromatographic method with electrochemical detection for the determination of hyperforin. AB - An HPLC method with electrochemical detection for the determination of hyperforin extracts without using additional sample precleaning has been developed and validated. The hyperforin solutions were separated isocratically using a mobile phase consisting of 10% ammonium acetate buffer (0.5 M, pH 3.7)-MeOH-acetonitrile (10:40:50, v/v) at a flow rate of 0.8 mL/min. Hyperforin was detected amperometrically with a glassy carbon electrode at a potential of +1.1 V versus Ag/AgCl/3 M potassium chloride reference electrode. Under these conditions, a plot of integrated peak area versus concentration of hyperforin was found to be linear over the range of 0.054-5.4 microg/mL, with a relative standard deviation of 2.2-8.6%. The limit of detection was 0.050 ng on column. The determination of the hyperforin content in a commercially available St. John's Wort preparation exhibited a mean content of 1.56 mg. Recovery experiments led to a mean recovery rate of 97 +/- 5.8%. The proposed method is not time-consuming, sensitive and reproducible and is therefore suitable for routine analysis of hyperforin in herbal medicinal products. PMID- 15281268 TI - Chromatographic behaviour of naproxen-cyclodextrin complexes stationary phase C8 alkyl chain as competitor for the drug release from cyclodextrin cavity. AB - Cyclodextrins are known to alter the absorptivity of the guest molecules, therefore, analytical methods that are based on the spectrophotometric data present accuracy problems. In this work, using RP-HPLC methods for naproxen cyclodextrins quantitation, extensive analytical inaccuracies are detected. Competitive complexation technique is utilised in an attempt to develop an analytical method enabling the determination of naproxen as a free drug. For this reason, stationary phases with silica ligands that can function as competing agents were used, thus contributing to the drug release. The release of the drug from cyclodextrins complexes is achieved by modification of the thermodynamic parameters that determine the stability constant, by changing: the interactions with the mobile phase components (e.g. pH, organic modifier, competitive agents) and the interactions with the stationary phase ligands (C8). After studying the parameters affecting the interaction between the alkyl-chain C8 and naproxen:cyclodextrin complexes, we developed and validated a new specific method for the accurate determination of the drug. Consecutive accumulation of the cyclodextrins molecules on the stationary phase was studied. PMID- 15281269 TI - Effect of alcohols on elution chromatography of trivalent actinides and lanthanides using tertiary pyridine resin with hydrochloric acid-alcohol mixed solvents. AB - Elution chromatography with a tertiary pyridine resin has been used to separate the trivalent actinides (An3+) from the lanthanides (Ln3+) using an alcoholic hydrochloric acid solvent. Trivalent Am and Cm were separated from the Ln by employing a 1 cm(phi) x 10 cm resin column with the mixed solvent system composed of concentrated hydrochloric acid (HCl) and alcohols. The distribution coefficients (Kd) and the separation factors between An and Ln (alpha(An)(Ln)) increased as the alcohol content of the solvent mixture increased. On the other hand, the Kd and alpha(An)(Ln) decreased drastically upon the addition of water to the solvent mixture. Among the four alcohols investigated (methanol, ethanol, n-propanol and n-butanol), the ethanol-HCl mixed solvent system showed the largest Kd and alpha(An)(Ln). The mechanism of adsorption for An and Ln cations on the pyridine resin is discussed in addition to the results presented herein. PMID- 15281270 TI - Influence of the injection technique and the column system on gas chromatographic determination of polybrominated diphenyl ethers. AB - In this paper, we present an investigation of the influence of the gas chromatographic separation system on the determination of polybrominated diphenyl ethers (PBDEs). Capillary columns, retention gaps and press-fit connectors, as well as different injection techniques have been evaluated with respect to yield and repeatability. The split/splitless injection has been optimized and compared to on-column injection, the septum equipped temperature programmable injector (SPI) and the programmable temperature vaporizing (PTV) injector. Furthermore, a comparison of the different operational modes of the PTV injector is presented. The results show that there are large variations in the yield of PBDEs depending on the column and the injection systems. Especially the high molecular weight BDE congeners can be subject to severe discrimination. Unfavorable conditions can lead to a complete loss of nona and deca substituted BDE congeners. PMID- 15281271 TI - Effect of amine mobile phase additives on chiral subcritical fluid chromatography using polysaccharide stationary phases. AB - Increased retention and selectivity in the subcritical fluid chromatography (SFC) of various amine compounds on polysaccharide chiral stationary phases (CSP) was observed upon incorporation of cyclic amines into the modifier. The retention increases are most pronounced with 2-propanol and are almost absent when methanol is used as modifier. This suggests that the effect may arise from a restriction to the modifier access to the binding site required to effect elution. The effect of the amine additives in SFC does not remain after their removal from the mobile phase. Findings were applied to the development of a 5 min separation of amphetamine and methamphetamine enantiomers. PMID- 15281272 TI - Determination of the aggregation threshold of non-UV-absorbing, neutral or charged surfactants by frontal- and vacancy-frontal analysis continuous capillary electrophoresis. AB - Supplementing our recent work on UV-absorbing anionic surfactants, new protocols based on frontal analysis continuous capillary electrophoresis (FACCE) were developed for the investigation of the aggregation threshold of non-UV absorbing anionic, cationic and neutral surfactants, and exemplified with sodium dodecyl sulfate (SDS), tetradecyltrimethylammonium bromide (TTABr) and Brij 35. Contrary to UV-absorbing surfactants, the critical micelle concentration (CMC) determination of non-UV absorbing surfactants requires the use of a marker providing adequate detection capabilities. UV-absorbing markers were selected, according to the charge of the studied surfactant (neutral for SDS and TTABr, anionic for Brij 35). In all cases, the free marker concentration was quantified as a function of the total surfactant concentration. In addition, a modified implementation of FACCE, that we called vacancy FACCE (VFACCE), was employed for the case of the neutral surfactant. VFACCE entails first filling the capillary with the system components to be studied in the background electrolyte, next continuously introducing the plain BGE electrokinetically. The salient theoretical features of FACCE and VFACCE were compared. These new protocols were successfully applied to yield reliable CMC values within short operational time and with low sample consumption. PMID- 15281273 TI - Migration behaviour and separation of tramadol metabolites and diastereomeric separation of tramadol glucuronides by capillary electrophoresis. AB - Capillary electrophoresis with UV detection was used to separate tramadol (TR), a centrally acting analgesic, and its five phase I (M1, M2, M3, M4, M5) and three phase II metabolites (glucuronides of M1, M4 and M5). Several factors were evaluated in optimisation of the separation: pH and composition of the background electrolyte and the influence of a micellar modifier, sodium dodecyl sulfate. Baseline separation of TR and all the analytes was obtained with use of 65 mM tetraborate electrolyte solution at pH 10.65. The lowest concentrations of the analytes that could be detected were below 1 microM for the O-methylated, below 2 microM for the phenolic and ca. 7 microM for the glucuronide metabolites. The suitability of the method for screening of real samples was tested with an authentic urine sample collected after a single oral dose (50 mg) of TR. After purification and five-fold concentration of the sample (solid-phase extraction with Oasis MCX cartridges), the parent drug TR and its metabolites M1, M1G, M5 and M5G were easily detected, in comparison with standards, in an interference free area of the electropherogram. Diastereomeric separation of TR glucuronides in in vitro samples was achieved with 10 mM ammonium acetate-100 mM formic acid electrolyte solution at pH 2.75 and with basic micellar 25 mM tetraborate-70 mM SDS electrolyte solution at pH 10.45. Both separations showed that glucuronidation in vitro produces glucuronide diastereomers in different amounts. The authentic TR urine sample was also analysed by micellar method, but unambiguous identification of the glucuronide diastereomers was not achieved owing to many interferences. PMID- 15281274 TI - Enhanced method performance due to a shorter chromatographic run-time in a liquid chromatography-tandem mass spectrometry assay for paclitaxel. AB - Liquid chromatography-tandem mass spectrometry (LC-MS-MS) is often performed in a high-throughput environment. Unfortunately, with atmospheric pressure ionization (API) techniques, shorter run-times or reduced sample clean-up often result in ion or matrix suppression, which can lead to erroneous results. The present work on the analysis of paclitaxel compares ion suppression, sensitivity and linearity of a high-throughput LC-MS-MS method (0.8 min run-time, method B) to a method with increased separation (2.0 min run-time, method A). An atmospheric pressure chemical ionization (APCI) interface was used for both methods. The high-sample throughput method uses an increased amount of organic solvent in the mobile phase (isocratic, 85% versus 70% of methanol) and a higher flow-rate (600 microl/min versus 400 micro/min). As a result, internal standard (docetaxel) and target analyte (paclitaxel) co-elute, close to, although separated from the solvent front. Ion suppression of both methods was evaluated by comparing pure standard to plasma and plasma containing a vehicle. Sensitivity and linearity were compared by injecting matrix matched calibration samples with both methods. Ion suppression by the vehicle Cremophor EL led to poor data quality for the standard method (A), while for the short method (B), ion suppression was compensated for by the co-eluting internal standard. The short method showed similar linearity but increased sensitivity by at least a factor five. This work provides a strategy to compensate for ion suppression without the use of labeled internal standards. In addition, a better sensitivity and a shorter run-time are noted. PMID- 15281275 TI - Identification of major active constituents in the fingerprint of Salvia miltiorrhiza Bunge developed by high-speed counter-current chromatography. AB - High-speed counter-current chromatography was applied as a method to develop fingerprinting of Salvia miltiorrhiza Bunge, a popular traditional Chinese medicine, in our previous study. Important active constituents that were directly related to the therapy effect should be identified. Each effluent fraction and standard samples (cryptotanshinone, tanshinoneI and tanshinoneIIA) were analyzed by ultraviolet spectroscopy and liquid chromatography-mass spectroscopy. It was concluded from the UV-Vis spectrograms, retention times in LC analysis and mass spectrograms, that fractions 7, 8 and 11 were respectively cryptotanshinone (Mr 296), tanshinone I (Mr 276) and tanshinone IIA (Mr 294). PMID- 15281276 TI - Electrochemical detector for microchip electrophoresis of poly(dimethylsiloxane) with a three-dimensional adjustor. AB - This paper presents an electrochemical detector for poly(dimethylsiloxane) (PDMS) microchip CE with a three-dimensional adjustor which makes it possible to accurately align a separate working electrode that can be easily fabricated in laboratory to the uncertain PDMS microchannel outlet. The substantial influence of the electrode-PDMS microchannel distance was investigated. The optimal electrode-outlet distance was found to be 10 microm for the PDMS microchannel with the width of 50 microm due to its relatively slow electroosmotic flow. Adrenaline and catechol were well separated, with a linear response range from 20 microM to 1 mM, and a detection limit of 2 microM for catechol, using carbon disk electrode (diameter of 300 microm). Furthermore, arginine and histidine can be well separated and detected directly in the PDMS microchannel using a Cu disk electrode (diameter of 150 microm). PMID- 15281277 TI - [File n(o) 52: hTERT]. PMID- 15281278 TI - [pRB, p53, p16INK4a, senescence and malignant transformation]. AB - Recent works aimed at clarifying the respective roles of p16INKa and p14ARF (both located on the same INK4a locus on chromosome 9p21 in man) in malignant transformation come to the conclusion that p16INK4a is the true tumor suppressor gene in man. In mouse, it is the p19ARF knockout that suppresses the barrier protecting cells from malignant transformation. This situation is in agreement with p19ARF- and p16-mediated senescence induced by oncogenic mutated ras (Ras*) in mouse and man respectively. Other results have shown that senescence in human diploid fibroblasts is associated with heterochromatin occurrence that maintains in repressed state E2F1-induced gens required for G1 to S phases transition. Since RB protein is responsible for this chromatin modification, cells with any impaired RB pathway cannot enter into senescence. PMID- 15281279 TI - [Management of emesis in cancer patients]. AB - Nausea and vomiting are often seen in cancer patients. They can be acute (induced by chemotherapy or radiotherapy...), or chronic in patients with advanced disease. A high percentage of patients (70% to 80%) suffer from chemotherapy induced nausea and vomiting. The incidence and severity of these symptoms depend of the nature of chemotherapy, doses, other drugs used in association, and psychological status of the patients. International guidelines have improved the management of nausea and vomiting, with the use of new drugs like setrons. Despite this, optimal care of refractory and delayed nausea and vomiting after chemotherapy is still a matter of debate. Chronic nausea and vomiting concern more than 50% of patients in palliative situation. The origin is often multifactorial. Management consists in aetiologic and symptomatic treatment in order to improve the patients' quality of life. PMID- 15281280 TI - [Update on docetaxel and breast cancer]. AB - Docetaxel (Taxotere) has been developed in breast cancer during the last decade. First its activity in monotherapy was proven in metastatic setting after failure of anthracycline therapy. Then the association with anthracycline demonstrated substantial activity leading to its development in early stages of breast cancer and its incorporation in adjuvant and neoadjuvant settings. Recently the first adjuvant trial comparing the association TAC versus FAC was presented. In the TAC arm, the disease free survival was better comparing with FAC (p = 0.0011) and survival was better in the subgroup with less than four positive lymph nodes. In the neoadjuvant setting, the incorporation of docetaxel after an anthracycline based regimen (protocols Aberdeen and NSABP-B27) led to better clinical response, subsequently to better breast conservation and more important the increase of the pathological response rate. Improvement of survival has been reported in the Aderbeen study but a longer follow-up of the NSABP B27 study is required to confirm the impact of Taxotere in the outcome of breast cancer. The next step will be the development of the combination of the most active chemotherapeutic regimen with targeted therapies according to molecular characteristics of the tumor. The integration of trastuzumab with taxane-based chemotherapy has already demonstrated high activity in metastatic breast cancer with overexpression of HER2 and adjuvant trials are ongoing. PMID- 15281281 TI - [2003 clinical practice guideline: Standards, Options and Recommendations for pain assessment in adult and children with cancer (summary report)]. AB - CONTEXT: The "Standards, Options and Recommendations" (SOR) project, which started in 1993, is a collaboration between the French Federation of Cancer Centers (FNCLCC), the 20 French Regional Cancer Centers, and specialists from French public universities, general hospitals and private clinics. The main objective is the development of clinical practice guidelines to improve the quality of health care and the outcome of cancer patients. OBJECTIVE: To update clinical practice guidelines for the assessment of pain in adult or children with cancer in collaboration with the French society for pain study and treatment. METHOD: The methodology is based on a literature review and critical appraisal by a multidisciplinary group of experts who define the CPGs according to the definitions of the Standards, Options and Recommendations project. Once the guideline has been defined, the document is submitted for review by independent reviewers. RESULTS: This article is a summary version of the full document presenting the clinical practice guidelines with algorithms. The main recommendations concern the means used to evaluate pain and its consequences and their use in specific cases (acute or chronic pain, patients able to communicate or not, children under or over 6 years old). Others recommendations were also established concerning the evaluation ofpsychological, social and family context, the evaluation of pain in hospital or at home, in terminal phase patients and for the establishment of a therapeutic strategy and follow-up of patient with pain. PMID- 15281282 TI - Thrombotic complications of implanted central venous access devices: prospective evaluation. AB - Implanted venous access devices (IVAD) are routinely used in oncologic patients. Thrombotic complication is a source of morbidity. During one year 246 patients with different solid neoplastic diseases received IVAD for chemotherapy administration. Two hundred forty-nine IVAD were placed percutaneously or by surgical cutdown. IVAD were flushed immediately after implantation with 3-5 mL of heparinized saline (100 U/mL). No monthly flush was required. A prospective evaluation of thrombotic complications was realised. in event of catheter dysfunction and/or clinical symptoms of phlebitis, a catheter opacification and/or a Doppler ultrasonography were performed. Twenty-three catheter dysfunctions were noted, corresponding to 13 catheter occlusions. Twelve patients presented clinical symptoms of phlebitis. Eleven venous thrombosis were diagnosed in this group; 10 by echo-Doppler and one by scanography. A unvaried statistic analysis using Fisher's test was performed to detect risk factors. Two factors were identified: the position of catheter tip above T4 (p < 0.001) and mediastinal or cervical lymph nodes larger than 6 cm (p < 0.001). The first increased the risk of catheter occlusion and the second increased the risk of phlebitis. PMID- 15281283 TI - [Anticancer drugs use evaluation: limits of the approved labeling]. AB - A practice survey was performed in Tenon hospital on 396 consecutive patients treated for solid tumors during 4 weeks in november 2002. 33% of anticancer drugs were off label used. The wording heterogeneity of the different anticancer drugs approved labeling and the lack of anticancer drugs in a number of cancers can explain those results. On one hand, randomised comparative clinical trial, considered as the best level of evidence to obtain a label used, is not always possible in cancerology, especially for rare tumors. One the other hand, pharmaceutical firm are not obliged to asked a label used for an anticancer drugs in spite of high level of evidence. So, label used can not be the own references for anticancer drugs prescribing, therapeutic advanced can be realised and disseminated before their taking into account in the label used. PMID- 15281284 TI - [A prospective evaluation of antibiotic prophylaxis efficacy for breast cancer surgery following previous chemotherapy]. AB - Use of antibiotic prophylaxis (AP) in clean breast cancer surgery is still controversial. We assessed the efficacy of preoperative AP in a prospective study of 171 clean breast cancer procedures following previous anticancer chemotherapy. From June 1998 to July 2001, we analyzed 171 procedures. In 133 cases. AB with cefuroxime was performed. Wound infection rate was 3 out of 171 procedures (WI rate of 2/131 with AP compared with 1/37 without AP, p = 1.0). This study suggests that AP is not systematically required in breast cancer surgery following previous anticancer chemotherapy. PMID- 15281285 TI - [For a coordination of the supportive care for people affected by severe illnesses: proposition of organization in the public and private health care centres]. AB - The concept of continuous and global care is acknowledged today by all as inherent to modern medicine. A working group gathered to propose models for the coordination of supportive care for all severe illnesses in the various private and public health care centres. The supportive care are defined as: "all care and supports necessary for ill people, at the same time as specific treatments, along all severe illnesses". This definition is inspired by that of "supportive care" given in 1990 by the MASCC (Multinational Association for Supportive Care in Cancer): "The total medical, nursing and psychosocial help which the patients need besides the specific treatment". It integrates as much the field of cure with possible after-effects as that of palliative care, the definition of which is clarified (initial and terminal palliative phases). Such a coordination is justified by the pluridisciplinarity and hyperspecialisation of the professionals, by a poor communication between the teams, by the administrative difficulties encountered by the teams participating in the supportive care. The working group insists on the fact that the supportive care is not a new speciality. He proposes the creation of units. departments or pole of responsibility of supportive care with a "basic coordination" involving the activities of chronic pain, palliative care, psycho-oncology, and social care. This coordination can be extended, according to the "history" and missions of health care centres. Service done with the implementation of a "unique counter" for the patients and the teams is an important point. The structure has to comply with the terms and conditions of contract (Consultation, Unit or Centre of chronic pain, structures of palliative care, of psycho-oncology, of nutrition, of social care). A common technical organization is one of the interests. The structure has to set up strong links with the private practitioners, the networks, the home medical care (HAD) and the nurses services at home (SSIAD), when they exist, to guarantee the continuity of the supportive care under all its aspects and in order to take into account the preferences of the patients. According to Hospital 2007 propositions, the extended, flexible and general purpose Group of Sanitary Cooperation (GCS) meets the necessities inherent to the structures of supportive care within the territories of health because it can be established between one or several health care centres and the private health professionals, thus favouring the cooperation between public and private health care centres. PSPH and general medicine. PMID- 15281286 TI - [Cancer prevention: congress report]. PMID- 15281287 TI - [Challenge through challenging--how personal development can be successful]. PMID- 15281288 TI - [The nursing case manager]. PMID- 15281289 TI - [Management without nursing directors?]. PMID- 15281290 TI - [MSRA (multi-resistant Staphylococcus A) from the point of view of personal care]. PMID- 15281291 TI - [Brought into the swing]. PMID- 15281292 TI - [Integrated care among geriatric psychiatric nursing for migrants]. PMID- 15281293 TI - [Gender mainstreaming--what relevance for nursing?]. PMID- 15281294 TI - [Care for patients or loss of freedom?]. PMID- 15281295 TI - Temporomandibular joint ankylosis: a multicenter Nigerian study. PMID- 15281296 TI - Awareness of periodontal diseases amongst Nigerian diabetics. AB - This prospective study sample consisted of 261 Nigerians made up of 155 diabetics seen at the medical outpatient clinic of Lagos University Teaching Hospital, Lagos and general hospital, Ikeja, Lagos, Nigeria and 106 health controls randomly selected. The subjects were required to complete a questionnaire containing the biographic data and occupation, and assessment of their knowledge, attitude and practice towards periodontal health and utilisation of appropriate professional care. They were then examined and assessed using the CPITN code. Their ages ranged from 5 to 65 years. Close to three-quarters of the subjects (73.5% for diabetics and 74.5% for controls) were 45 years or older. Furthermore, 6% of the low skilled diabetics new about periodontal disease while only 27.2% of those in the medium and highly skilled group had some knowledge of periodontal disease. This relationship was found to be statistically significant (X2 = 12-22 ; df = 2, p < 0.05). Similarly, statistically significant relationship was demonstrated between diabetics and controls with regards to knowledge about the demonstrated between diabetic and controls with regards to knowledge about the cause/s of periodontal disease and awareness of gum bleeding. 73.6% of the controls had never been to the dentist before unlike 62.1% of diabetics. All subjects in either group who claimed not to bleed scored at least the minimum on the CPITN code. While 52.3% of diabetics and 22.5% of control had a CPITN score of 2,54% of those in the control group and 30% of diabetics recorded a score of 3 on the CPITN code. In conclusion, the level of awareness of periodontal disease is quite low amongst both groups, hence there is a need to promote periodontal health awareness in the general populace as well as clinics. PMID- 15281297 TI - [Topographical and morphological study of the mandibular foramen in black Africans from the Ivory Coast]. AB - Sixty one dry mandibles. have been studied to provide some anatomical informations on the position, shape and size of the mental foramen among adults Black Africans of Ivory Coast. According the results, in the male mandibles, the mental foramen lay 27,31 mm behind the symphyses , 74,75 mm forward the post border of the ramus, 14,89 mm above the lower border and 16,16 mm under the alveolar margin. In the female mandibles, the mental foramen lay 27,16mm behind the symphysis, 69,10 mm forward the post border of the ramus, 14,21 mm above the lower border and 15,66 mm under the alveolar margin. This study confirmed that on the horizontal plane, the mental foramen lay approximatively one quarter of the distance from mandibular symphyses to the post border of the ramus. The margin of mental foramina was elliptic in 66,67 % and 64,52 % of the cases respectively in male and female mandibles. The mean sizes of the long and short axes of forarnina were 5,03 mm and 3,97 mm in the male mandibles. These dimensions were 4,99 mm and 3,87 mm in the female mandibles. PMID- 15281298 TI - Attitude, beliefs and practices of some Nigerian nurses toward teething in infants. AB - The objective of this study was to assess the perceptions of some Nigerian nurses on the various societal beliefs about teething. A cross sectional survey was conducted among 542 nurses in the teaching, general and Local Government hospitals and clinics in Ibadan, a city in south western Nigeria. The outcome of the study revealed that the majority of the nurses believed that loss of appetite, crying, increased salivation and general irritability were a necessary part of the teething process. Furthermore, 82,1%, 61,4% and 27,9 % of them implicated fever, diarrhoea and boils respectively as signs of teething. The older and more experienced nurses and males seemed to ascribe symptoms more with the teething process. From this study, it is clearly evident that there are erroneous beliefs concerning teething persistent among Nigerian nurses. Since the societal beliefs may be harmful to the health of the children, there is a desperate need to address them. In doing this, a health education programme should be formulated to educate these misconceptions among the general public and especially target older and more experienced nurses as well as the males. Nursing and expectant mothers should also be PMID- 15281299 TI - The localised aggressive periodontitis prevalence in Morocco. AB - This work concerns a retrospective study of prevalence records and the clinical characteristics of localised aggressive periodontitis in the breast of consultants in the periodontitis service in Dental Medicine Faculty of Rabat (Morocco) from 1997 to 1999. PMID- 15281300 TI - Oro-facial trauma in amateur secondary school footballers in Ibadan, Nigerian: a study of mouthguards. AB - To assess the awareness and frequency of use of mouth-guards for football among secondary school children in Ibadan, Nigeria, as well as the amount of oro-facial trauma previously associated with football game in these adolescents, a questionnaire - based cross-sectional study was conducted. Six hundred and thirty -one amateur footballers - 465 (73.7%) males and 166 (26.3%) females in the age range of 10 - 19 years (mean age, 15.01 +/- 2.86SD) completed the questionnaire giving a response rate of 90.1%. Majority (58.8%) of the subjects was within the 10-15 years age group. Awareness of mouth-guards was claimed by 69.7% of the respondents but significantly more of them who made the claim were not using the protective device for football games (p<0.05). More boys significantly claimed the usage of mouth-guards than girls (p<0.05). In all, 59.7% agreed that wearing mouth-guard was helpful in oro-facial injury prevention during sports with significantly more boys in agreement than girls (p<0.05). About 36% of the subjects sustained one form of oro-facial trauma or the other with significant sex difference (p<0.05). Secondary schools should serve as good starting points in the campaign for use of mouth-guards for contact sports before some go into professional sports. PMID- 15281301 TI - Caries experience and oral hygiene status of blind, deaf and mentally retarded female children in Riyadh, Saudi Arabia. AB - OBJECTIVE: To determine the caries experience and oral hygiene status in blind, deaf and mentally retarded female children in Riyadh, Saudi Arabia. METHOD: All (N=218) the 6-7-year-old and 11-12-year-old blind, deaf and mentally retarded female children registered with the Presidency of Girls' Education schools in Riyadh were examined for dental caries and oral hygiene in a dental operatory setting. RESULTS: All (100%) the blind 6-7-year-old had caries with a mean dmft score of 6.58 (SD 2.02). The caries prevalence in blind 11-12-year-olds was 88.2% with a mean DMFT score of 3.89 (SD 2.67). Among 6-7-year-old blind children 8.3 %, and in 11-12-year-old blind children 29.4% had good oral hygiene. The caries prevalence in deaf 6-7-year-olds was 95.7% with a mean dmft score of 7.35 (SD 3.51). The caries prevalence in 11-12-year-old deaf children was 93% with a mean DMFT of 5.12 (SD 3.45). Less than one-fifth (17.4%) of the 6-7-year-old deaf children and only 7.0% of 11-12-year-old deaf children had good oral hygiene. The caries prevalence in mentally retarded 6-7-year-old was 93.9% with a mean dmft of 8.00 (SD 4.1). All the mentally retarded 11-12-year-old had carious teeth with a mean DMFT score of 5.81 (SD 2.95). Only 3.1% of the mentally retarded 6-7-year old and none of the mentally retarded 11-12-year-olds had good oral hygiene. CONCLUSIONS: Caries prevalence and severity in all the three groups of female special children were very high, and the number of children with good oral hygiene was very low. PMID- 15281302 TI - [Medical history--basis for modern physicians]. PMID- 15281303 TI - [New routines makes tooth extraction possible during warfarin treatment]. AB - We conclude that it is possible to successfully extract teeth without interrupting or reducing the dose of anticoagulant medication with warfarin. We also conclude that patients with anticoagulant medication are treated differently depending on where they have their dental treatment done. PMID- 15281304 TI - [BRCA1 and BRCA2 have reached the clinical medicine. The 10-year old finding of the genetic mutation makes it now possible to prevent breast cancer]. AB - Increased knowledge of breast cancer genetics has improved the possibilities to predict the future risk of a woman to be diagnosed with breast cancer. In certain families, presymptomatic testing of breast cancer susceptibility genes may be offered, leading to an even more accurate individual risk prediction. As a result, advice regarding follow-up and risk reducing measures may be given to the individuals with the highest risks of cancer. Preventive surgery drastically reduces the risk of having breast or ovarian cancer respectively. The value of increased controls in hereditary high risk women is insufficiently investigated. Further studies are warranted to elucidate the efficacy of chemoprevention in women at a very high risk of breast cancer, e.g. mutation carriers of BRCA1 and BRCA2. PMID- 15281305 TI - [Physical activity as medication against arthrosis--training has a positive effect on the cartilage]. AB - It is well known that exercise, alone or combined with weight reduction, reduces pain and improves function in patients with osteoarthritis. The knowledge of the effects of exercise on cartilage is limited and needs to be improved however. It seems as cartilage adapts to loading as other biological tissues like bone and muscle, and moderate loading seems to be beneficial both for prevention and treatment of osteoarthritis. Too high loads, like elite soccer or repeated knee bendings several hours daily, are associated with increased risk of osteoarthritis. Too high or too low mechanical load decrease the proteoglycan content of the cartilage, indicating not only elite sports but also physical inactivity being a possible risk factor for osteoarthritis development. Muscle weakness may proceed osteoarthritis. Prevention and treatment of osteoarthritis should include regular loading of the cartilage, keeping muscles strong and maintaining normal body weight. PMID- 15281306 TI - [A case report concerning traffic medicine. A fatal accident after renal dialysis]. PMID- 15281307 TI - [The new biology. A brilliant, slandered queen]. PMID- 15281308 TI - [The new biology. Truth, social construction or something in between?]. PMID- 15281309 TI - [Fellow-workers training for motivated specialists. Reflections from two leaders/initiators]. PMID- 15281310 TI - [Guidelines, infarction care and quality]. PMID- 15281311 TI - [Anti-stigmatization campaign--starting at the wrong end]. PMID- 15281312 TI - [The hopeless debate to find out which psychotherapy is the best one is going on and on...]. PMID- 15281313 TI - Introduction: Clonal evolution in bladder cancer: a scientific and clinical perspective. PMID- 15281314 TI - Risk factors in clonal development from superficial to invasive bladder cancer. AB - Bladder cancer has classically been associated with exogenous risk factors, and a large literature has identified risk factors associated with the genesis of transitional cell carcinoma. Only recently have efforts been made to identify host factors and to evaluate possible changes in tumour presentation and biology, including grade and stage, in association with these risk factors. The available literature appears to demonstrate alterations in tumour biology associated with environmental carcinogens. Various studies have suggested a consistent upgrading of bladder cancer stage and grade as a result of cigarette smoking and high risk occupational exposures. It is important, however, that all factors associated with increased risk for bladder cancer be more extensively evaluated in assessing the validity of this concept. PMID- 15281315 TI - Squamous change in bladder cancer and its relevance to understanding clonal evolution in development of bladder cancer. AB - Using conventional morphological assessment, squamous change in bladder epithelium has been observed in 73% of bilharzial associated squamous cancers but only 28% of pure transitional cancers. However, more detailed studies of patients with TCC suggest that the latter figure may be an underestimate, since in one series it was reported to be more than 50%. The most significant risk factor for development of squamous carcinoma in the bladder is chronic persistent bacterial cystitis, although in the areas of the world where bilharzia is endemic this infestation also increases the risk of both squamous bladder cancer and chronic bacterial cystitis. Although it is clear that carcinogens are involved as co factors in transformation from squamous metaplasia to cancer, the fact that in Zimbabwe one author has observed that TCC is more frequent in whites than squamous cancer is in bilharzia infected blacks is evidence that other unidentified risk factors are involved. This is increasing evidence for involvement of HPV subtypes in cervix, oropharynx and lung cancer. As all three of these tumours are associated with squamous metaplasia, there could be a case for investigation of bladder squamous tumours for HPV involvement. This is particularly so given the observation of the "hit and run" type of transient infection in cattle that develop BPV associated tumours and the tenfold difference (30% vs 3%) in frequency of HPV detection in squamous skin tumours developing in immunosuppressed individuals compared with those arising spontaneously. With new technology for cytological screening techniques using dot ELISA and evidence of differences in TP53 mutations that support the involvement of nitrous oxide, it is clear that there is more to learn from study of this tumour type that may be of general interest in understanding the clonal development of cancer. PMID- 15281316 TI - Malignancy associated papillomaviruses and morphology of human bladder cancer. AB - Animal studies in rabbit and cattle have clearly demonstrated the contribution of host genetics, chemical carcinogens and immunosuppression to the conversion of papillomavirus induced benign regressing warts into malignant cancers. More significant is the role of vaccination both with whole tumour cell suspensions with whole virus and viral proteins, particularly L2 molecules, in causing progressing warts to regress. Early results in small scale studies of HPV16 E6/E7 vaccine in patients with cervical cancer have provided evidence that tumour regression can be induced in human papillomavirus induced tumours. These observations provided added impetus for more research to firm up the increasing, but still principally anecdotal, evidence that papillomaviruses may be involved in the pathogenesis of bladder cancer. Studies of carcinomas arising in cattle after BBV 4 infection show absence of fully infectious virus in the majority of tumours, though the tumours have persistent E7, E8 and LCR sequences. As this is all that is required for transformation, it may require in vitro molecular studies in human bladder cancer screening for such elements before final proof of involvement is confirmed. However, even before this is achieved, given the success in animal models of whole tumour cell vaccines, serious thought should be given to how to develop protocols for study of crude tumour cell vaccines in vivo. Such studies would need in vitro assays to seek evidence for specific antitumour immunity, focusing on studies of tumour infiltrating lymphocytes and their T cell receptor polymorphisms. PMID- 15281317 TI - Molecular genetics of bladder cancer: pathways of development and progression. PMID- 15281318 TI - Molecular biological changes in bladder cancer. AB - A large number of potential molecular markers of bladder cancer have been identified, although only a few are truly independent prognostic factors. A number of markers may need to be measured in a single tumour and used as a combination for use in the diagnosis and prognosis of transitional cell carcinoma (TCC). Epidermal growth factor receptor immunoreactivity has been shown to be an independent predictor of survival and stage progression. TP53 may be an independent predictor of recurrence and overall survival in TCC confined to the bladder, and TP53 alterations may predict chemosensitivity in patients who have had TCC treated by radical cystectomy. At present molecular techniques such as fluorescence in situ hybridization and the polymerase chain reaction are restricted to the laboratory, but immunohistochemical methods are available in most hospital pathology departments. There are some discrepancies and conflicting reports of the usefulness of molecular markers in different studies, and these need to be addressed in large, prospective, multi-institutional studies using standardized molecular techniques. PMID- 15281319 TI - Clinical evaluation of immunotherapy: are there differences between papillary and flat in situ bladder cancer? AB - The advantage of BCG immunotherapy over intravesical chemotherapy in superficial bladder cancer has been most apparent in patients with carcinoma in situ (CIS), where complete response is increased from 50% to more than 70% and the proportion of patients remaining disease free for 5 years is increased from 20% to 40%. Similar advantages have been reported using suboptimal BCG treatment schedules in patients with recurrent stage Ta, T1 tumours. BCG provides long term protection from tumour recurrence and, unlike chemotherapy, reduces tumour progression. The observed relative increased sensitivity of CIS to BCG and the occasional failure of BCG to demonstrate significant superiority over mitomycin C in the prevention of tumour appear to be related to the use of suboptimal BCG treatment schedules. With maintenance BCG using 3 weekly instillations at 6 month intervals, patients with papillary tumours fare even better than patients with CIS, and tumour progressio is even further reduceld. Chemotherapy is appropriate for patients who are at very low risk of tumour progression and those who fail to respond to BCG, but overall the results of BCG immunotherapy are superior for patients with either CIS or Ta, T1 transitional cell carcinoma. PMID- 15281320 TI - Clonal development of bladder cancer and its relevance to the clinical potential of HLA antigen and TP53 based gene therapy. AB - In the last decade there has been major progress in understanding the multiple steps involved in cancer cells developing their malignant potential. Study of bladder cancer has provided important information during this period and helped to identify HLA class I and wild type normal TP53 as potential probes for gene therapy studies. Given the magnitude of genetic damage that is associated with the clonal development of bladder cancer, and particularly the number of cellular mechanisms that have been highjacked in the development of terminal metastatic disease, it is obviously highly unlikely that a single gene therapy involving HLA class I alone would work in terminal metastatic disease. However, it seems possible that HLA-B7 treatment of patients with bladder cancer who are candidates for salvage cystectomy would benefit such patients. If it were to work, it could provide a whole new approach to managing superficial tumours. However for patients with extensive metastatic disease, progress could come from investing more effort in uncoverinlg the genetic basis of the chemosensitivity of germ cell tumours and understanding the basis of the normal checkpoints of meiosis and the role of TP53. This could provide a completely new approach to gene therapy that might be exploited even in patients with advanced metastatic disease. Such a tetraploidal construct combined with allogeneic HLA class I and incorporated into a low pathogenic lytic viral construct to induce oncolysis with some form of tissue promoter to focus the cell types in which the genes will be activated could provide ideal effective gene therapy. PMID- 15281321 TI - The role of surgery in the multimodality treatment of bladder cancer. AB - There has been little change in bladder cancer survival for more than 40 years, although earlier diagnosis is now detecting more cases at an early, potentially curable stage. Radical cystectomy remains the most effective single treatment, although in the past the morbidity and mortality of treatment and the age of patients made it less favoured as primary treatment. Progress in continent bladder reconstruction is changing attitudes. However, because tissue damage from preoperative radiation, particularly when combined with chemotherapy, makes such operations less safe in patients with advanced disease, reconstruction is primarily of value in high risk superficial and early invasive cancers, though there remains a need for randomized trials or immediate vs deferred use of these operations to establish when they give most benefit. With new knowledge about the role of trauma released tissue repair cytokines and immunosuppressive effect of prolonged anaesthesia on increasing tumour recurrence after surgery, new approaches such as treatment with TNFA, anti-EGF antibody or neoadjuvant chemo/immunotherapy before TURBT to improve on the benefits of surgery need to be explored in randomized trials in both advanced invasive and early superficial disease. With progress in vaccine and gene therapy on the horizon, the central role of the urologist in both harvesting tumours for molecular diagnosis and monitoring response of local disease to treatment is undisputed. The relative underusage and value of bladder washings cytology to provide cells for such studies is also highlighted. PMID- 15281322 TI - Radiotherapy and chemotherapy for invasive bladder cancer. AB - Invasive bladder cancer is associated with locoregional and distant metastases. To improve the outcome of management, systemic chemotherapy has been combined with locoregional treatment. Programmes have been structured in which chemotherapy is administered before or after definitive radiotherapy or surgery, or in combination with radiotherapy. Most randomized trials to date have failed to define a survival benefit from initial chemotherapy, but evidence is emerging that classical adjuvant chemotherapy may improve survival. New cytotoxic agents, including paclitaxel and gemcitabine, accompanied by an emerging understanding of the factors governing cytotoxic drug resistance, may also lead to better management. PMID- 15281323 TI - Future directions: bladder cancer. PMID- 15281324 TI - Nonsteroidal anti-inflammatory drug-induced reactions and desensitization. PMID- 15281325 TI - Development of an expert system knowledge base: a novel approach to promote guideline congruent asthma care. AB - Existing guidelines for the clinical management of asthma provide a good framework for such tasks as diagnosing asthma, determining severity, and prescribing pharmacological treatment. Guidance is less explicit, however, about establishing a patient-provider partnership and overcoming barriers to asthma management by patients in a way that can be easily adopted in clinical practice. We report herein the first developmental phase of the "Stop Asthma" expert system. We describe the establishment of a knowledge base related to both the clinical management of asthma and the enhancement of patient and family self management (including environmental management). The resultant knowledge base comprises 142 multilayered decision rules that describe clinical and behavioral management in three domains: 1) determination of asthma severity and control; 2) pharmacotherapy, including prescription of medicine for chronic maintenance, acute exacerbation, exercise pretreatment, and rhinitis relief; and 3) patient self-management, including the process of intervening to facilitate the patient's asthma medication management, environmental control, and well-visit scheduling. The knowledge base provides a systematic and accessible approach for intervening with family asthma-related behaviors. PMID- 15281326 TI - The relationship between asthma and obesity in children: is it real or a case of over diagnosis? AB - OBJECTIVE: To determine whether obesity among children is associated with an increased incidence of asthma. DESIGN AND METHOD: Five thousand nine hundred eighty-four children participated in a lung health study in the Ashkelon region, Israel. A lung health questionnaire was completed and they underwent spirometry. Body mass index (BMI) was then calculated for each child. RESULTS: Three hundred two children (5.05%) were above the 95th percentile for BMI and considered obese. Obese children tended to wheeze more than the non-obese children 14.5% vs. 10.5%, respectively (p<0.038). Asthma (physician diagnosis) was diagnosed more often among obese children than non-obese 7.2% vs. 3.9%, respectively (p<0.008). Inhaler use was more prevalent among obese children than non-obese 15.9% vs. 8.8%, respectively (p<0.001). Bronchial hyperreactivity was significantly greater among the non-obese asthmatic children compared with their obese counterparts, 352 (51.4%) vs. 10 (27.8%), respectively (p<0.001). Chest symptoms and asthma were more frequent in obese than non-obese boys. CONCLUSION: Asthma, wheezing, and inhaler use were more common in obese children than in non-obese children. Symptoms were more prevalent among obese boys. Increasing BMI among children is a risk factor for asthma, which may in reality be obesity-related chest symptoms that mimic asthma. PMID- 15281327 TI - Middle lobe syndrome in children with asthma: review of 56 cases. AB - OBJECTIVE: Middle lobe syndrome (MLS) is one of the complications of asthma. Its signs and symptoms are often nonspecific, causing delay in appropriate treatment. We aimed to review our pediatric asthmatic patients and provide differential characteristics between MLS and asthma worsening in order to target early diagnosis. METHOD: File records of all asthmatics (n=3528) seen in our clinic during the last 2 years were retrospectively reviewed to identify the patients with MLS, and a case-control study was undertaken. Files of 56 asthmatic children diagnosed as MLS, with a total of 63 episodes, and 63 matched controls with asthma worsening were analyzed and compared. RESULTS: The incidence of MLS was 1.62% and half were below or at the age of 6. All cases with MLS were documented radiologically, and only 5 of the 63 episodes had physical findings suspicious for MLS. The most affected segments were right middle lobe (50%) and left lingula (26.2%). Although in all cases symptoms cleared, in 23 (36.5%) cases, atelectasis persisted radiologically. Compared to controls, patients with MLS included less atopics (34.9% vs. 59.4%, p<0.05) and fewer boys (52.4% vs. 71.4%, p<0.05), and they reported less frequent dyspnea (57.1% vs. 85.9%), more frequent sputum production (49.2% vs. 7.8%), and longer duration of complaints (22.0+/-6.23 vs. 2.4+/-0.31 days) (p<0.001, for each). Furthermore, the resolution of symptoms took significantly longer (45.2+/-9.3 vs. 3.3+/-0.4 days, p<0.001). CONCLUSION: We conclude that complicating MLS in childhood asthma is more frequent in younger ages, girls, and nonatopics. In most cases, physical findings are not informative, and chest radiographs diagnose most but not all cases. The most suggestive symptoms are unresolving/persisting symptoms during admission and/or following treatment. PMID- 15281328 TI - Attachment of mothers and children with recurrent asthmatic bronchitis. AB - OBJECTIVE: This study on attachment in children with recurrent asthmatic bronchitis and their mothers addresses three issues. The first aim was to test whether children affected by recurrent asthmatic bronchitis more often display an insecure pattern of attachment in comparison with healthy children. The second aim was to verify whether the distribution of adult attachment representations in the mothers of children affected by recurrent asthmatic bronchitis is different from the one shown by the mothers of the healthy comparison group. The third aim was to investigate intergenerational transmission of attachment. METHODS: Sixty Italian children, aged between 2 and 5 years, and their mothers participated in the study. The Adult Attachment Interview and the Attachment Q-Sort were used to assess, respectively, the security of mothers' attachment representations and of mother-child attachment. RESULTS: Children affected by recurrent asthmatic bronchitis appeared to be less secure in comparison with healthy children. Their mothers showed a higher percentage of insecure attachment representations. Finally, the intergenerational transmission of attachment was not influenced by the preclinical condition of the children. CONCLUSIONS: We propose a model of genetic and social transmission of insecure attachments in families struggling with asthma. PMID- 15281329 TI - Asthma management practices at home in young inner-city children. AB - Information on parental asthma management practices for young children is sparse. The objective of this article is to determine if specific caregiver asthma management practices for children were associated with children's asthma morbidity. Caregivers of 100 inner-city children diagnosed with persistent asthma and participating in an ongoing asthma intervention study were enrolled and interviewed to ascertain measures of asthma morbidity, medication use, health care use (acute and primary care), and asthma management practices. Overall, asthma morbidity was high with almost two thirds of caregivers reporting their child having one or more emergency department visits within the last 6 months and 63% receiving specialty care for their asthma. Appropriate medication use was reported predominantly as albuterol and inhaled steroids (78%). However, only 42% of caregivers reported administering asthma medicines when their child starts to cough and less than half (39%) reported having an asthma action plan. There were no significant differences by asthma severity level for any asthma management practice. In conclusion, caregivers lack knowledge regarding cough as an early asthma symptom. Caregivers should be encouraged to review asthma action plans with health care providers at each medical encounter. PMID- 15281330 TI - Successful school-based intervention for inner-city children with persistent asthma. AB - OBJECTIVE: Because children attend school daily, school-based interventions for children with persistent asthma could provide effective disease management for inner-city asthmatic children. The Kunsberg School in Denver, Colorado, enrolls children with chronic diseases, including asthma, into a daily program of school based disease management. This study sought to determine the impact of the Kunsberg program on asthma utilization. METHODS: Children attending Kunsberg (n=18) who received primary care at Denver Health were compared with a group of matched control children who also received primary care at Denver Health, but did not attend Kunsberg (n=36). Asthma-related utilization for an average of 2.9 years before and after Kunsberg enrollment was assessed. RESULTS: The 18 Kunsberg and 36 control subjects were mostly minority children in low-income families, without significant demographic differences between groups. Compared with controls, the Kunsberg cohort experienced fewer hospitalizations (0.5 vs. 0.9 hospitalizations/subject/ year, p=0.05), fewer emergency department (ED) visits (1.4 vs. 2.8 ED visits/ subject/year, p=0.04), and fewer follow-up visits for asthma (3.7 vs. 5.0 visits/subject/ year, p=0.01) in the time period (mean 2.9 years; range 1-6 years) following the intervention. Hospital- and clinic-based asthma utilization costs decreased 80% following enrollment in the school (8122 dollars/year to 1588 dollars/year per child), compared to a 19% decrease in the control group. Among the Kunsberg children with hospitalizations prior to school enrollment (n=8), hospital days decreased from 3.5 days to 0.1 days annually (p<0.01), ED visits decreased from 2.1 to 0.6 visits annually (p=0.02), and follow-up visits decreased from 6.8 to 2.1 visits annually (p=0.02). As part of their school program, 89% of Kunsberg enrollees received inhaled corticosteroids daily on a monitored basis while at school. CONCLUSIONS: The Kunsberg school program improved asthma control and reduced disease severity for at-risk inner city asthmatic children, leading to cost reduction for asthma management. Directly observed controller therapy at school can be an important component of a school-based program for children with chronic conditions. PMID- 15281331 TI - School-based asthma disease management. AB - BACKGROUND: Asthma is the most common chronic childhood illness and the leading cause of missed school days. School is a potential location for establishing an asthma education program for children and their parents/caregivers designed to improve disease management. OBJECTIVE: To determine whether a comprehensive, school-based asthma management program, in addition to a conventional disease management program, can reduce measures of asthma control, student absenteeism, and caregiver lost workdays. METHODS: School nurses recruited parents/caregivers of students with asthma from three urban elementary and middle schools. Children were identified as having asthma by a previous diagnosis from their personal physician. Parents were invited to attend educational sessions about the program. Students received peak flow meters and training in their use and had access to an interactive asthma diary to record symptoms, peak flow, and medicine usage. They received monthly asthma education at school and had access to an online asthma education program and additional handouts. Parents received several educational calls regarding asthma and had a 24-hour, 7-days-a-week emergency number to call if problems arose. RESULTS: At 6 months, missed school days and unscheduled doctor visits were reduced by two thirds (n = 41; p< 0.01 for each). Caregivers' perception of children's activity level increased by 11% (n = 26; p = 0.037). Daytime and nighttime frequency of symptoms dropped by 62% and 34%, respectively (n = 32; p < 0.007 and p<0.03 for each). These trends continued at 12 months, although only reduction in frequency of symptoms attained statistical significance. CONCLUSIONS: A comprehensive, school-based asthma management program can successfully improve asthma control and reduce absenteeism in elementary and middle school students and caregiver lost workdays. PMID- 15281332 TI - Health-related quality of life in children and adolescents with asthma: results from the ESTAR Study. AB - Our aim was to assess the psychosocial well-being of asthmatic children and adolescents, the influencing factors, and to determine the effect of inpatient rehabilitation on their quality of life; 226 asthmatic children and adolescents participated in the inpatient rehabilitation (IG). The comparison group (CG) included 92 asthmatic children and adolescents receiving standard medical treatments. Patients were aged between 8 and 16 years and were predominantly male. The health-related quality of life was measured with the German version of the "Paediatric Asthma Quality of Life Questionnaire." Interviews were carried out for IG 2 weeks before the commencement of their inpatient stay and 1 year after their stay ended. The same time schedule was carried out for CG. All patients reported a mild to moderate impairment of their quality of life. Girls described a slightly lower quality of life than boys. With increasing asthma severity, quality of life decreased. Inpatients described a lower quality of life than CG at enrollment. Inpatient rehabilitation resulted in a greater improvement of quality of life over time for IG than for CG. Gender and severity status had no effect on this time course. The only modestly affected quality of life may reflect the good adaptation to the disease and medical treatment. Children and adolescents in the IG recorded improvements in their quality of life. Differences in quality of life based on gender and disease severity were not shown to influence the improvements. In summary, inpatient rehabilitation results in an improvement of health-related quality of life. Further research concerning the psychosocial situation of children and adolescents in this setting is needed. PMID- 15281333 TI - Exhaled nitric oxide predicts asthma exacerbation. AB - The fraction of exhaled nitric oxide (FeNO) is elevated in asthmatics compared to normal subjects. Many studies have demonstrated that FeNO correlates with other markers of airway inflammation. The purpose of this study was to assess the clinical utility of routine monitoring of FeNO in determining its ability to predict future asthma exacerbations compared with other standard clinical measures of spirometry, peak flows, quality of life score, medication usage, and symptoms. A convenience sample of 22 patients with moderate and severe-persistent asthma in the University of New Mexico Adult Asthma Clinic were evaluated during a routine clinic visit and then noted whether they had an exacerbation within 2 weeks of the initial appointment. Those with an exacerbation had a higher mean FeNO (29.67 ppb +/- 14.48) compared to those who did not (12.92 ppb +/- 5.17), p = 0.002. A nominal logistic regression model to determine those variables that predict asthma exacerbation found that FeNO was the only significant predictor, p = 0.03. Thus, FeNO appears to be a clinically useful tool to assess disease control in this population. PMID- 15281334 TI - Recruitment into a long-term pediatric asthma study during emergency department visits. AB - OBJECTIVE: Asthma is the most common chronic illness in childhood. Recruiting children and their parents into a research study in a busy urban emergency department (ED) is challenging. The purpose of this manuscript is to describe the recruitment process and the results of our recruitment in soliciting children and their parents to participate in an ED-based asthma research study. METHODS: The data for this manuscript came from a National Heart, Lung, and Blood Institute funded study: Study of Asthma Follow-up from the Emergency Department (SAFE). SAFE evaluated an ED-based intervention to link low-income urban children with asthma to their primary care providers. Two persons were assigned specifically to enrolling, which was done from 0700 to 2300 hours Monday through Friday. Data for the analysis come from the web-based database, the master log, and the hospital's patient database. A computerized randomization scheme chose 512 patients from all patients in the master log for more detailed demographic analyses. RESULTS: Five hundred twenty-seven subjects were enrolled between February 1999 and May 2001. There were 9188 children who presented for treatment of an acute asthma exacerbation during this interval. The number of eligible parents was similar to the predicted number. Chart reviews were conducted on a subset of patients presenting to the ED to ensure that the recruitment strategy did not bias the patients enrolled. Demographic characteristics of asthma patients were similar during enrollment and non-enrollment times. Comparison of patients who were enrolled with those who were not enrolled indicated no differences by gender, race, insurance status, age, or socioeconomic status of neighborhood residence. DISCUSSION: The high rate of enrollment was primarily due to the two dedicated enrollers. The enrollers quickly learned how to function within the ED and how to interact with both families and ED staff. Strategies identified by the enrollers as helpful in randomizing subjects included visits with the parents shortly after the physician had initiated treatment so that stability of the child had been achieved. Interacting with the child and showing concern for the comfort of both the child and parent during the ED stay were important as well. CONCLUSION: Recruiting subjects into long-term follow-up studies in the ED setting is a departure from traditional ED studies. The ED enrollment offers the distinct advantage of capturing subjects who are unlikely to present for care in other locations. We were able to successfully recruit low-income urban parents of children with asthma for study of both short-term and long-term outcomes. Careful attention to planning and then integration of enrollers into the ED setting can result in successful recruitment of patients and their parents. The enrollment process successfully captured the subjects of interest without bias. PMID- 15281336 TI - Quality of life in young urban children: does asthma make a difference? AB - The purpose of this study was to determine whether asthma status and severity have an impact on the quality of life of urban elementary school children. Participants were 1292 caregiver-child dyads from six schools serving low-income, ethnic minority, urban families; 53% of the children were female. Caregivers provided data on the children's asthma diagnosis and frequency in the last 12 months of asthma symptoms, use of medication for asthma, emergency room visits, and hospitalizations. Using the KINDL, a generic quality of life instrument, children reported on their health-related quality of life (HRQL). Results revealed a high prevalence of current asthma (18%). No differences were found in HRQL based on having current asthma or the severity of asthma as assessed by proxy measures of health care utilization and limited functioning. These findings are consistent with previous research indicating that HRQL is influenced by several factors other than asthma status and severity. The implications of these results for intervention are discussed. PMID- 15281335 TI - Exposure levels of asthmatic children to allergens, endotoxins, and serine proteases in a tropical environment. AB - A cross-sectional study was conducted in Bayamon, Puerto Rico, to identify and quantify indoor allergens, serine proteases, and bacterial endotoxin present in homes of asthmatic children. A total of 126 dust samples from houses were obtained from the entire mattress and bedside floor. Most of the patients had detectable levels of mite, cockroach, cat, and dog allergens. Mold allergens were found only in bedside floor dust samples. Mouse allergens were not detected. Forty-two percent, 36.5%, and 1.8% of the patients demonstrated exposures to sensitizing levels of mite, Bla g 1 and cat allergens, respectively. The percentage of patients exposed to high levels of allergens capable of triggering asthma symptoms was 33.3% and 26.4% for mite and Bla g 1 allergens. Only dog allergen, bacterial endotoxin, elastase, and trypsin were associated with asthma symptoms. Eighty-nine percent of the asthmatic children were exposed to endotoxin concentrations greater than 100 EU/mg dust, and more than half of the patients were exposed to high levels of serine proteases. Our study indicates that indoor concentrations of allergens traditionally associated with asthma symptoms and severity may not be applicable in tropical environments and highly ventilated households. In fact, in the study population, endotoxins, dog allergen, and serine proteases may play a dominant role in the induction of asthma symptoms. PMID- 15281337 TI - Molecular basis of pituitary development and cytogenesis. AB - The factors that play a role in pituitary development have been identified as transcription factors and growth factors that regulate cell migration, proliferation and differentiation. These factors are expressed in a tightly regulated fashion to ensure the dorso-ventral migration of Rathke's pouch, the proliferation of cells in a correct temporal and spatial fashion, and the cytodifferentiation of the hormone-producing cell types of the mature gland. Dysregulation of these processes results in congenital abnormalities and hormone insufficiency syndromes. Understanding these normal processes of differentiation and proliferation sheds new light on the factors implicated in disordered structure and function of the mature gland during pituitary tumorigenesis. PMID- 15281338 TI - Molecular pathology of the pituitary. Development and functional differentiation of pituitary adenomas. AB - This review article describes functional differentiation of the pituitary cells and pituitary adenomas with special emphasis on transcription factors and co factors. Human pituitary adenomas generally follow the combination of transcription factors and co-factors, which are similar to those of physiologic anterior pituitary cells. On very rare occasions, the single pituitary adenoma produces two hormones, which belong to different cell lineage 'trans-cell lineage'. Basic mechanism for this was considered to be 'aberrant expression' of transcription factors, i.e. NeuroD1 and Pit-1. This was experimentally supported by the induction of GH (mRNA and protein) in AtT-20 cells by transfecting Pit-1 gene. Various mechanisms have been reported for the experimental pituitary oncogenesis. Among these, GHRH has been emphasized as one of oncogenic factors for both human GHomas as well as in the transgenic animals. PMID- 15281339 TI - Cell cycle dysregulation in pituitary oncogenesis. AB - The cell cycle is the process by which cells grow, replicate their genome and divide. The cell cycle control system is a cyclically-operating biochemical device constructed from a set of interacting proteins that induce and coordinate proper progression through the cycle, and includes cyclins, cyclin-dependent kinases (CDK) and their inhibitors (CDKI). There are mainly two families of CDKI, the INK family (INK4a/p16; INK4b/p15; INK4c/p18 and INK4d/p19) and the WAF/KIP family (WAF1/p21; KIP1/p27; KIP2/p57). Progression through the cell cycle is mainly dependent on fluctuations in the concentration of cyclins and CDKI achieved through the programmed degradation of these proteins by proteolysis within the ubiquitin-proteasome system. There is also a transcriptional regulation of cyclin expression, probably dependent on CDK phosphorylation. The p53 family--p53, p63 and p73--function as transcription factors that play a major role in regulating the response of mammalian cells to stressors and damage, in part through the transcriptional activation of genes involved in cell cycle control (e.g. p21), DNA repair, senescence, angiogenesis and apoptosis. Essential for the maintenance of euploidy during mitosis is human securin, identical to the product of the pituitary tumour-transforming gene (PTTG). Loss of regulation at the G1/S transition appears to be a common event among virtually all types of human tumours. Aberrations of one or more components of the pRb/p16/cyclin D1/CDK4 pathway seem to be a frequent event (80%) in pituitary tumours. The role of p27 is rather that of a haploinsufficient gene. p27-/- mice show an increased growth rate, due to increased cellularity, testicular and ovarian cell hyperplasia and infertility, and hyperplasia of the pituitary intermediate lobe with nearly 100% mortality caused by such a benign pituitary tumour. Although the p27 gene was not found to be mutated in human pituitary tumours and its mRNA expression was similar in tumour samples in comparison with normal pituitaries, the load of p27 protein expression in corticotroph adenomas and pituitary carcinomas was shown to be much lower than those in normal pituitary tissue or other types of pituitary adenoma, suggesting that post-translational processing of p27 accelerates its removal from the nucleus. In respect to p27 degradation and its cellular compartmentalization, several pathways have been explored. Malignant tumours are associated with increased nuclear immunostaining for Jun activation binding protein-1 (Jab1) which is responsible for phosphorylated p27 export from the nucleus. Corticotrophinomas are characterized by massively increased phosphorylation of p27 on Thr187, but are not associated with changes in Jab1. Macrophage inhibitory factor (MIF), which binds and inactivates Jab1, was noted to be over-expressed in tumours with abundant Jab1, suggesting that it may be part of a compensatory mechanism to moderate Jab1 activity. Proteasomal degradation of p27 requires its ubiquitylation by the SCF ubiquitin ligase, with specific addressing by the F-box protein Skp2 and its co-factor Cks1. Pituitary tumours with high p27 protein expression showed significantly less Skp2 expression than samples with low p27 immunostaining, suggesting that increased Skp2 could play at least a part in this process. No difference was observed in Cks1 mRNA levels between normal pituitaries and pituitary adenomas. The present data suggest that inhibition of growth and tumour development is sensitive not only to the absolute levels of p27 protein, but also to its cellular compartmentalization. Very recent findings from our group have established up regulation of the serine-threonine kinase Akt in pituitary tumours compared to normal pituitary, which may cause phosphorylation of p27 on Thr157 and cytoplasmic retention of p27. PTTG protein is highly expressed in various human tumours, including pituitary tumours. While its mRNA levels are low in normal pituitary, increases in PTTG transcripts from more than 50% to more than 10-fold were recorded in the majority of a series of pituitary adenomas. Control of the cell cycle is a vital part of the cell's replication machinery. Disruption of this process is commonly seen in pituitary tumours and we are now beginning to identify regulatory elements which are likely to play a major role in pituitary oncogenesis. PMID- 15281340 TI - Role of regulatory factors in pituitary tumour formation. AB - The molecular basis of pituitary tumorigenesis remains controversial, but there are two major theories which have been subject to most investigation: hormonal (usually hypothalamic factors) and/or growth factor overstimulation, or a molecular defect within the pituitary itself. It has been shown, for example, that excessive regulatory hormone stimulation can lead to an increased number of cells in the pituitary in various physiological or pathological states such as pregnancy (lactotrophs), untreated primary hypothyroidism (thyrotrophs and lactotrophs),primary hypoadrenalism (corticotrophs) and ectopic GHRH-secreting tumours (somatotrophs). Animal models also provide data that in the presence of excessive hypothalamic hormone stimulation, adenoma formation can occur. However, evidence in favour of the monoclonal nature of pituitary tumours argues for an intrinsic molecular defect as the primary initiating event in tumour formation. We review the various hormonal factors and their receptors effecting the different types of pituitary cells, such as CRH, AVP and cortisol feedback on corticotrophs; GHRH, Galpha PKA, somatostatin and GH and IGF feedback on somatotrophs; GnRH, LH/FSH, activin and oestrogen feedback on gonadotrophs; dopamine, oestrogen and prolactin feedback on lactotrophs; and TRH, TSH and thyroid hormone feedback on thyrotrophs. The monoclonal origin of adenomas makes it unlikely that hypothalamic factors could initiate pituitary transformation, but they could still create an environment where there is a higher chance that a possible causative tumorigenic mutation may occur in one (or several) of the overstimulated pituitary cells, or enhance the proliferation of an already mutated cell. PMID- 15281341 TI - Growth factors and cytokines: function and molecular regulation in pituitary adenomas. AB - The growth and functions of the anterior pituitary cells are regulated by hypothalamic factors, peripheral hormones and growth factors. However, the expression of numerous growth factors and cytokines, as well as their receptors, within the anterior pituitary suggests that these factors could be locally involved in the control of pituitary development, function and proliferation by auto-/paracrine mechanisms. In the normal pituitary, intrapituitary factors probably play a role in modifying endocrine signals in the pituitary cells. However, since alterations in growth factor production and receptor expression have been found in pituitary adenomas, these factors may also contribute to the pathophysiology and progression of pituitary tumours. The potential roles of the most important growth factors and cytokines in pituitary adenoma pathogenesis are reviewed. PMID- 15281342 TI - Proliferation markers and cell cycle inhibitors in pituitary adenomas. AB - Proliferation markers are widely used in general surgical pathology and also in pituitary pathology. They should help for differing aggressive or rapidly growing tumors from those with slower growth, as cellular atypia is not helpful for identifying aggressive adenomas of the pituitary. Only the number of mitoses is important for prognosis. A lot of markers can be used: antibodies for cyclins A, B, D and E, for proliferating cell nuclear antigen, Ki-67/Mib-1, antibodies for the inhibitory proteins p16, p27, p53, and for DNA topoisomerase IIalpha. A marker for apoptosis and its inhibitors may be also important. From our experiences, Mib-1 is the most reliable marker. The recommendation of this marker in the WHO classification of pituitary adenomas is fully justified. PMID- 15281343 TI - Down-regulation of E-cadherin and catenins in human pituitary growth hormone producing adenomas. AB - Growth hormone (GH)-producing pituitary adenomas can be ultrastructurally divided into two major types: densely granulated and sparsely granulated. The latter type of adenoma characteristically exhibits globular accumulations of cytokeratin filaments known as fibrous bodies, which are immunohistochemically identifiable as juxtanuclear dot-like immunoreactivity. We hypothesize that the formation of fibrous body might be related to dysfunction of adhesion molecules, because of the functional relationship between intermediate filaments and the cadherin catenin complex and frequent observation of loss of cohesiveness of the adenoma cells. Our recent immunohistochemical study showed that expression of E-cadherin and its undercoat proteins, alpha-, beta- and gamma-catenin, in GH cell adenomas with prominent fibrous bodies was significantly reduced compared with GH cell adenomas without fibrous bodies and the normal adenohypophysial cells. Although no mutation of exon 3 of the beta-catenin gene was found in any GH cell adenomas with fibrous bodies, methylation-specific polymerase chain reaction analysis revealed that the E-cadherin promoter region was methylated in 37.5% of these adenomas, two of which displayed total methylation, but not in GH cell adenomas without fibrous bodies. We conclude that the decreased expression of the E cadherin-catenin complex and methylation of the E-cadherin gene promoter region are events associated with the formation of fibrous bodies in GH cell adenomas. It remains to be clarified to explain the mechanism by which down-regulation of adhesion molecules is involved in the abnormal assembly of intermediate filaments. PMID- 15281344 TI - Morphologic changes and molecular regulation of angiogenesis in pituitary adenomas. AB - Angiogenesis, the process of development of a new vasculature, plays a crucial role in tumour growth. In the pituitary, unlike other tissues, vascularization is lower in adenomas compared to the normal gland. Despite this finding, a relationship between increased vascularity and some aspects of tumour behaviour such as size, invasiveness, surgical outcome and malignancy, has been demonstrated. The process of angiogenesis is the result of a balance of stimulating and inhibiting factors. It is likely that an interaction between gene expression (such as pituitary tumour transforming gene), hormonal stimuli including oestrogens, corticosteroids, dopamine, 16-kDa fragments of prolactin and growth hormone, somatostatin analogues, and pro- and anti-angiogenic growth factors (e.g. vascular endothelial growth factor and fibroblast growth factor), determine the final angiogenic phenotype of pituitary tumours, and thus subsequent tumour behaviour. PMID- 15281345 TI - Advances in pituitary pathology: use of novel techniques. AB - Many new techniques are rapidly being developed and applied to the study of normal and neoplastic pituitary tissues. These range from techniques used for the past few decades such as in situ hybridization to more recent developments such as comparative genomic hybridization, laser capture microdissection, DNA and tissue arrays, proteomics, and RNA interference technology. These approaches have been applied to answer many challenging questions about pituitary function and pathophysiology. This chapter reviews many of these recent developments and shows their applications to pituitary biology in order to demonstrate how these new techniques are providing insights about basic aspects, clinical, and pharmacologic knowledge of the pituitary gland and of pituitary tumors. PMID- 15281346 TI - Pituitary tumor transforming gene: an update. AB - Herein we summarize the recent rapid advances in understanding the pituitary tumor transforming gene (PTTG) oncogene. Clinical studies reveal that PTTG binding factor, fibroblast growth factor 2, and vascular endothelial growth factor are elevated in pituitary tumors, and mostly correlate with PTTG levels, also confirming the PTTG role in angiogenesis. PTTG overexpression disrupts mitosis and causes aneuploidy in single live cells and PTTG modulates p53 activity and p53 also mediates DNA damage-induced inhibition of PTTG transcription. Physiological functions of PTTG are revealed by PTTG-null mice who exhibit a variety of cell growth abnormalities including diabetes mellitus secondary to defective beta-cell proliferation. PTTG is therefore an oncogene for pituitary tumors and other neoplasia, and also involved in critical metabolic functions. Further studies are required to address mechanisms for these oncogenic and physiological functions, and more importantly, to understand conditions which determine the switch of PTTG from functioning physiologically to behaving as an oncogene. PMID- 15281347 TI - Pituitary tumour clonality revisited. AB - Allelotype analysis and X chromosome inactivation analysis in women enables the assessment of tissue clonality, and has demonstrated that the majority of sporadic human pituitary adenomas are monoclonal. This implies that these tumours arise from de novo somatic genetic change(s) in a single pituitary cell. However, clonality within any given tumour may be multiple or single, multiple tumours arising on the background of hyperplasia may be of identical or different clonality, multiple 'sporadic' tumours in a tissue may be of differing clonal origin, and finally morphology cannot predict genetic makeup. These general principles may also apply to the pituitary so it is simplistic to assume that monoclonality is inevitable and that pituitary tumours cannot be multiclonal in origin. Indeed, these observations would be entirely compatible with the initiating stimulus resulting in hyperplasia of specific cell subtypes in the pituitary giving rise to a number of different clones each with variable potential to develop into a discrete tumour depending on their rate of cell division/rate of apotosis. Stimuli might include pituitary-specific oncogenes, intrapituitary growth factors, or extrapituitary trophic factors (e.g. hypothalamic releasing hormones). PMID- 15281348 TI - Molecular cytogenetics of pituitary adenomas, assessed by FISH technique. AB - Fluorescent in situ hybridization (FISH) represents a moden molecular pathology technique, alternative to conventional cytogenetics (karyotyping). In addition to metaphase spreads, it can be applied directly to interphase nuclei. The latter makes the FISH technique powerful for pathologists for it integrates molecular genetics and classic cytogenetics and brings them together to a single framework for morphologic evaluation. Interphase FISH can be applied to imprints from fresh tissue or to paraffin sections after proteinase K digestion. Centromeric, telomeric and locus DNA-sequence specific probes can be used to identify aneuploidy or gene mutations. Several protocols combine molecular cytogenetics with classic karyotyping. Other sophisticated, FISH-based protocols have been introduced. Among them, comparative genomic hybridization is very important for it can detect non-balanced chromosomal aberrations of uncultured tumor cells and provide overall genomic information in a single experiment. This review presents the principles and applications of FISH technique for the investigation of the cytogenetic background of pituitary adenomas. PMID- 15281349 TI - Morphology, molecular regulation and significance of apoptosis in pituitary adenomas. AB - Apoptosis represents energy-requiring spontaneous single cell death, with specific morphologic and biochemical features. It is a rapidly processed sequence of events resulting in elimination of damaged cells. Apoptosis occurs in physiological remodeling and proliferative conditions, and also in neoplastic lesions. Several molecules and molecular systems such as bcl-2/bax, Fas/FasL and caspases regulate the apoptotic process. Apoptosis is characterized by a stereotypic pattern of morphologic features, which can be illustrated mostly by electron microscopy. DNA and biochemical assays, based on the specific pattern of nucleosomal fragmentation can detect apoptosis. The in situ labeling techniques are currently used to demonstrate apoptosis in paraffin sections. Several studies of pituitary animal models, cell lines and human pituitaries have been performed during the last 6 years. By electron microscopy, pituitary adenoma cells undergoing apoptosis exhibit a common prototypical pathway of changes. Although the results by the situ labeling techniques are not uniform, apoptosis occurs with low frequency in a subset of pituitary adenomas, in carcinomas and in pituitary hyperplasia. Alternative techniques based on remodeling of cytoskeleton by caspase activity can identify early apoptotic stages. This review presents the principles of apoptosis and summarizes the morphologic and functional changes of apoptosis in pituitary. PMID- 15281350 TI - Somatostatin receptors in pituitary function, diagnosis and therapy. AB - Recent studies have demonstrated that human pituitary adenomas express multiple somatostatin receptor (sst) subtypes. The expression of sst subtypes in human pituitary adenomas is highly variable. This variability in sst subtype expression may explain the variable responsiveness of patients with pituitary adenomas to medical treatment with the sst2-preferring SS-analogs octreotide and lanreotide. In human GH-secreting pituitary adenomas, both sst2 and sst5 are involved in the regulation of GH secretion. In prolactinomas, sst5 receptors are the key receptors in regulating responsiveness to SS. The low abundance of sst2 in prolactinomas explains the lack of efficacy of octreotide in lowering the elevated PRL levels in prolactinoma patients. Octreotide and lanreotide successfully suppress TSH levels, including normalization of thyroid hormone levels in the large majority of patients with TSH-secreting pituitary adenomas, probably due to the high level of sst2, expression in the adenomas. Although sst2 receptors are expressed by a significant proportion of gonadotroph and clinically non-functioning pituitary adenomas, the overall efficacy of sst2-preferring SS analogs seems low in this type of patients. Finally, corticotroph adenomas may express multiple sst subtypes as well. Octapeptide SS-analogs do not lower circulating ACTH levels in patients with untreated pituitary-dependent Cushing's disease, whereas SS and the SS-analog octreotide suppress pathological ACTH release in some patients with Nelson's syndrome. This review discusses the expression and potential role of sst subtypes in the different types of human pituitary adenomas. PMID- 15281352 TI - Recent advances in MEN1 gene study for pituitary tumor pathogenesis. AB - Evidence of multiple endocrine neoplasia type 1 (MEN1) is found in approximately 2.7% of patients with pituitary adenomas. The multicentricity of pituitary adenomas has not yet been proved. Prolactinomas are most frequent in MEN1 pituitary tumors. Pituitary tumors with MEN1 are larger in size and more aggressive than without MEN1. Heterozygous germline mutations of MEN1 gene are responsible for MEN1 disorders. Various types of mutations likely causing loss of the gene function have been identified throughout the entire region in patients with MEN1 and related disorders. However, the function of menin, the product of MEN1 gene, remains to be established. Neither mutation hot spot nor phenotype genotype correlation has been established in classical MEN1. A number of recent studies suggest that somatic mutations in the MEN1 gene do not play prominent role in the pathogenesis of sporadic forms of pituitary adenoma. PMID- 15281351 TI - Pathology and molecular genetics of the pituitary gland in patients with the 'complex of spotty skin pigmentation, myxomas, endocrine overactivity and schwannomas' (Carney complex). AB - Carney complex (CNC) is a familial multiple neoplasia and lentiginosis syndrome with features overlapping those of McCune-Albright syndrome (MAS) and other multiple endocrine neoplasia (MEN) syndromes like MEN type 1 (MEN 1). Pituitary tumors have been described in a number of patients with CNC; all have been growth hormone (GH) and prolactin (PRL)-producing. In at least some patients, pituitary gland involvement is manifested by hyperplastic areas; hyperplasia appears to involve somatomammotrophs only and to precede GH-producing tumor formation, in a pathway similar to that seen in MAS-related pituitary tumors (and in oncogenesis in other CNC tissues). One patient with CNC and advanced acromegaly had a GH producing macroadenoma that showed extensive genetic changes at the chromosomal level. These changes appeared to represent secondary or tertiary genetic 'hits' involved in pituitary oncogenesis and were confirmed at the molecular level. So far, almost half of the patients with CNC have germline-inactivating mutations in the PRKAR1A gene; in their pituitary tumors, the normal allele of the PRKAR1A gene is lost. Loss of heterozygosity suggests that PRKAR1A, which codes for the regulatory subunit type 1alpha of the cAMP-dependent protein kinase A (PKA), may act as a tumor-suppressor gene in pituitary tissue. These data provide evidence for a PKA-induced somatomammotroph hyperplasia in the pituitary tissue of CNC patients; hyperplasia leads to additional genetic changes at the somatic level, which in turn cause the formation of adenomas in some, but not all, patients. PMID- 15281353 TI - [PFLEGE periodical as a mirror of science]. PMID- 15281354 TI - [The relationship between relatives and nurses in the Intensive Care Units]. AB - The relations between the nurses and relatives in the Intensive Care Unit is not always without problems, even if nursing theories unanimously advocate that the nursing staff is responsible for the care of these relatives. Since up to now only marginal empirical examinations concerning the relationship exist, this qualitative, explorative study will look into the nature of the relationship between relatives and nurses in the Intensive Care Unit. Narrative interviews with the relatives and narrative-episodic interviews with the nursing staff made it possible to recreate situations and to detect hidden and not conscious meanings, intentions and behaviour. Key situations could be brought out more clearly which are meaningful to both sides and from which could be extracted the problems which make dealing with one another difficult. Elements which are important in the relationship show themselves in the key situations "first contact", "information", "feelings", "presence and waiting", "dealing with the patient", "feedbacks" and "rules". The interviewed relatives did not experience themselves as part of the nursing work--decisive was that the relative is well taken care of. The nursing staff also experience the support and care of the relatives as an addition to their actual work. PMID- 15281356 TI - [Verbal patient information through nurses--a case of stroke patients]. AB - The article represents results of a theoretical work in the field of nursing education, with the topic: Verbal Patient Information through Nurses--A Case of Stroke Patients. The literature review and analysis show that there is a shortage in (stroke) patient information generally and a lack of successful concepts and strategies for the verbal (stroke) patient information through nurses in hospitals. The authors have developed a theoretical basis for health information as a nursing intervention and this represents a model of health information as a "communicational teach-and-learn process", which is of general application to all patients. The health information takes place as a separate nursing intervention within a non-public, face-to-face communication situation and in the steps-model of the nursing process. Health information is seen as a learning process for patients and nurses too. We consider learning as information production (constructivism) and information processing (cognitivism). Both processes are influenced by different factors and the illness-situation of patients, personality information content and the environment. For a successful health information output, it is necessary to take care of these aspects and this can be realized through a constructivational understanding of didactics. There is a need for an evaluation study to prove our concept of health information. PMID- 15281355 TI - [Autonomy and informed consent in surgical care-patients' and staff perceptions]. AB - The aim of this study was to describe autonomy and informed consent in surgical care. The study is a part of the international BIOMED 2 project "Patients' Autonomy and Privacy in Nursing Interventions" (BIOMED2, BMH4-CT98-3555; 1998 2001) supported by the European Commission. For this study, data of patients (n = 254) and nurses (n = 205) in eleven Berlin hospitals and three hospitals outside Berlin were collected by means of a structured questionnaire. The findings of the study indicate, that information-giving was more positive than decision-making. Patients perceived they were more frequently informed about their surgery than about their care. According to the perceptions of nurses the case was reversed. The perceptions of both groups differed, since from the point of view of nurses, patients' autonomy was more frequently heeded and their consent was sought more often than from the point of view of the patients. Patients admitted as emergencies and in multi-bed rooms perceived their autonomy more negatively than those with a planned surgery or in single rooms. Elderly nurses were more frequently than younger nurses of the opinion to grant patients autonomy. Nurses with a longer working experience in nursing care perceived that patients were more frequently asked their consent. Further, nurses with a higher educational qualification and with a higher occupational status perceived decision-making more negatively. The findings of the present study give implications for clinical practice, nursing education, and for further research. PMID- 15281357 TI - [The work profile of family caregivers of persons affected by HIV/AIDS]. AB - In Switzerland, HIV incidence has increased considerably during the past two years. This fact has raised concern both in the media and in the health care system. However family caregivers have not been considered in these debates, although their figures are increasing as well. In Switzerland, a few research studies were conducted focussing on the situation of family caregivers affected by HIV/AIDS. This particular qualitative research project focuses on the questions as to what kind of work these family caregivers are doing, under which conditions they are working, and which consequences arise from their work. Data were collected by conducting eleven semi-structured interviews, and data analysis was done through a content analysis. Results show that family caregivers provide a wide range and a large amount of family work. Not only are family caregivers working for the HIV-positive persons, but also for the benefit of other family members and friends as well as for themselves. Family caregivers achieve mainly stable life situations for their HIV-positive family member. The discussion focuses on the dynamics of family work which requires constant adaptations in daily life. In addition, ordinary family work needs to be noticed and appreciated in symbolic, social, and financial terms. Finally, implications for nursing practice are considered. PMID- 15281358 TI - [Quality dimensions in patient records--a standardised analysis of nursing home resident records]. AB - 279 resident records of 26 nursing homes were analysed using a standardised instrument as part of a comprehensive study about the quality of nursing homes in Frankfurt on Main. The main questions referred to the completeness of several content indicators and the visibility of the nursing process in the different parts of the records. Selection criteria were records only of residents in care dependency level two or three. From each of the 26 participating nursing homes at least 10 up to a maximum of 14 records depending on the number of residents were chosen. The best part of the documentation is the assessment, especially the body oriented problems and resources and the caring devices. However neither the resident's perspective or the source of information are generally noted. The planning part contains mainly interventions, is not always related to the assessment and refers quite seldom to the support of resources. The report shows even more deficiencies. The worst part is the evaluation. A comparison of extreme groups of nursing homes with a very good resident/staff relationship to those with a very bad resident/staff relationship shows that many parts of the records, but not all, are much better documented in the homes with more staff than in those with less staff. PMID- 15281359 TI - [Evidence-based nursing--a comprehensive term]. AB - The connotation of Evidence Based Nursing, patient centred care, decisions in health care, intuition and evidence based on international published research is presented. A distinction between Evidence Based Medicine and Evidence Based Nursing in terms of evidence and the hierarchy of evidence is necessary. The concept of Evidence Based Nursing for the last few years has been including practical expertise and patient choice. In nursing the selection of research articles for implementation rely not solely on the evidence hierarchy but also on the model of Rycroft-Malone et al. (2002). The term Evidence Based Patient Choice was the answer to the declaration of patient rights. These rights about accurate and detailed information are essential, so that patients can actively be involved in the decision making process about the management of their disease. Nurses, therefore, must reflect and verbalize the intuition behind the decisions in a systematic and institutionalised way to include in their future decisions for similar cases the full range of evidence. PMID- 15281360 TI - [Letter about Pflege issue 2/2004]. PMID- 15281361 TI - [Letter regarding the editorial by Hanna Mayer in Pflege 2004, 17, 70-2]. PMID- 15281362 TI - Synthesis and antiviral activities of new pyrazolo[4,3-c]quinolin-3-ones and their ribonucleoside derivatives. AB - Several new pyrazolo[4,3-c]quinolin-3-one ribonucleosides (5a-g) and their corresponding heterocycle moieties (3a-g) were synthesized and evaluated against vaccinia virus (VV) and herpes simplex virus type 1 (HSV-1). The derivatives 3c and 3d showed modest inhibitory activity against vaccinia virus reaching 70% at a concentration of 100 microM. All heterocyclic compounds (3a-f) showed a modest inhibition against HSV-1, reaching the maximal inhibitory effect around 20-30%. The antiviral effects of most of the pyrazolo[4,3-c]quinolin-3-one ribonucleosides (5a-f) on VV and HSV were not impressive. PMID- 15281363 TI - Glycosylation reaction of unprotected sugars with hydroxyalkylthymine. AB - Under mild conditions (Lewis acid/solvent/room temperature), the reaction of unprotected glucose, deoxyribose or xylose with hydroxylalkylthymine gives selectively nucleoside analogs with a spacer arm between sugar and base moiety. Experimental conditions (Lewis acid, solvent) for this new strategy leading to nucleoside analogs synthesis are discussed. PMID- 15281364 TI - Improved allelic differentiation using sequence-specific oligonucleotide hybridization incorporating an additional base-analogue mismatch. AB - Sequence-specific oligonucleotide hybridization (SSOH, 'dot-blotting') is a widely employed method of typing single nucleotide polymorphisms (SNPs), but it is often compromised by lack of allelic differentiation. We describe a novel improvement to SSOH that incorporates an additional mismatch into the oligonucleotide probe using the universal base analogue 3-nitropyrrole. This method greatly increases allelic differentiation compared to standard SSOH where oligonucleotides contain only SNP-defining base changes. Moreover, stringency of the hybridisation is predictably maintained over a wide range of temperatures, which can be calculated empirically, thus facilitating the genotyping of multiple SNPs using similar conditions. This improved method increases the usefulness of hybridisation-based methods of rapid genotyping of SNPs and may have implications for array methodologies. PMID- 15281365 TI - Solution stability and degradation pathway of deoxyribonucleoside phosphoramidites in acetonitrile. AB - The impuritiy profiles of acetonitrile solutions of the four standard O cyanoethyl-N,N-diisopropyl-phosphoramidites of 5'-O-dimethoxytrityl (DMT) protected deoxyribonucleosides (dG(ib), dA(bz), dC(bz), T) were analyzed by HPLC MS. The solution stability of the phosphoramidites decreases in the order T, dC>dA>dG. After five weeks storage under inert gas atmosphere the amidite purity was reduced by 2% (T, dC), 6% (dA), and 39% (dG), respectively. The main degradation pathways involve hydrolysis, elimination of acrylonitrile and autocatalytic acrylonitrile-induced formation of cyanoethyl phosphonoamidates. Consequently, the rate of degradation is reduced by reducing the water concentration in solution with molecular sieves and by lowering the amidite concentration. Acid-catalyzed hydrolysis could also be reduced by addition of small amounts of base. PMID- 15281366 TI - Enzymatic and hybridization properties of oligonucleotide analogs containing novel phosphoramidate internucleotide linkages. AB - In line with the paradigm, that antisense oligonucleotides should contain minimal structural modifications, in order to minimize the risk of toxicity and antigenicity, we describe here the preparation and the properties of oligonucleotides modified to contain, in addition to phosphodiester bonds, a small number of phosphoramidate internucleotide linkages substituted with aminoethoxyethyl groups in order to convey protection against exo- and endonucleases. Prolonged stability was, in fact, found in model experiments with respective enzymes, as well as in studies done in human blood serum. Regardless of number and position of phosphoramidate linkages, the modified oligonucleotides showed only a slight decrease of Tm in hybridization studies with complementary oligonucleotides. PMID- 15281367 TI - Oligonucleotide-minor groove binder conjugates and their complexes with complementary DNA: effect of conjugate structural factors on the thermal stability of duplexes. AB - Synthetic polycarboxamide minor groove binders (MGB) consisting of N methylpyrrole (Py), N-methylimidazole (Im), N-methyl-3-hydroxypyrrole (Hp) and beta-alanine (beta) show strong and sequence-specific interaction with the DNA minor groove in side-by-side antiparallel or parallel orientation. Two MGB moieties covalently linked to the same terminal phosphate of one DNA strand stabilize DNA duplexes formed by this strand with a complementary one in a sequence-specific manner, similarly to the corresponding mono-conjugated hairpin structures. The series of conjugates with the general formula Oligo-(L-MGB-R)m was synthesized, where m = 1 or 2, L = linker, R = terminal charged or neutral group, MGB = -(Py)n-, -(Im)n- or -[(Py/Im)n-(CH2)3CONH-(Py/Im)n-] and I < n < 5. Using thermal denaturation, we studied effects of structural factors such as m and n, linker L length, nature and orientation of the MGB monomers, the group R and the backbone (DNA or RNA), etc. on the stability of the duplexes. Structural factors are more important for linear and hairpin monophosphoroamidates than for parallel bis-phosphoroamidates. No more than two oligocarboxamide strands can be inserted into the duplex minor groove. Attachment of the second sequence-specific parallel ligand [-L(Py)4R] to monophosphoroamidate conjugate CGTTTATT-L(Py)4R leads to the increase of the duplex Tm, whereas attachment of [-L(Im)4R] leads to its decrease. The mode of interaction between oligonucleotide duplex and attached ligands could be different (stacking with the terminal A:T pair of the duplex or its insertion into the minor groove) depending on the length and structure of the MGB. PMID- 15281368 TI - Synthesis and antiviral evaluation of some novel tricyclic pyrazolo[3,4-b]indole nucleosides. AB - Novel pyrazolo[3,4-b]indole nucleoside analogs were synthesized from the corresponding 3-formyl-2-chloroindole and 3-cyano-2-chloroindole nucleosides by treatment with hydrazine. Very few examples of pyrazolo[3,4-b]indole heterocycles have been published in the literature and this is the first synthesis of nucleoside analogs containing this heterocycle. These new pyrazolo[3,4-b]indole nucleosides were active against human cytomegalovirus and herpes simplex virus type 1, but this activity was not well separated from cytotoxicity. PMID- 15281369 TI - Synthesis and antiviral activity of novel 2',4'-doubly branched carbocyclic nucleosides. AB - A series of 2' and 4'-doubly branched carbocyclic nucleosides 15, 16, 17 and 18 were synthesized starting from simple acyclic ketone derivatives. The required 4' quatemary carbon was constructed using Claisen rearrangement. In addition, the installation of a methyl group in the 2'-position was accomplished using a Grignard carbonyl addition of isopropenylmagnesium bromide. Bis-vinyl was successfully cyclized using a Grubbs' catalyst II. Natural bases (adenine, cytosine) were efficiently coupled by using Pd(0) catalyst. PMID- 15281370 TI - Asbestos dispatches. AB - As the global campaign to ban asbestos gathers momentum, more countries and organizations are restricting the use of chrysotile (white asbestos). Attempting to reverse the fall in global demand, asbestos stakeholders have redoubled marketing efforts in countries such as India where health and safety regulations are not strictly enforced. In the Canadian asbestos heartland, public and media awareness has grown and industry's influence has weakened. In response, chrysotile producers are pressing federal and provincial governments for mandatory increases in consumption, despite new research documenting an asbestos cancer epidemic. PMID- 15281371 TI - Canadian asbestos: a global concern. AB - The impact of the chrysotile (white asbestos) industry at home and abroad was the focus of the first independent conference on asbestos to be held in Canada. In September 2003, Canadian asbestos victims and international experts met to examine a wide range of issues on Capital Hill, Ottawa; victims and their relatives spoke eloquently of the devious means used by asbestos stakeholders to maintain control of the national asbestos agenda. At the conclusion of the conference, Ban Asbestos Canada, a campaigning group to assist victims and raise awareness of Canada's tragic asbestos legacy, was formed. PMID- 15281372 TI - Participatory mapping of occupational hazards and disease among asbestos-exposed workers from a foundry and insulation complex in Canada. AB - A study of former asbestos-exposed foundry and insulation workers was carried out in Sarnia, Ontario, home to Canada's petrochemical industry, using participatory mapping to document past exposures and subsequent diseases. Before it closed, government inspectors had monitored the use of asbestos at the facility, documenting levels that were thousands of times above the current legal limit. The study was undertaken by the Occupational Health Clinics for Ontario Workers (OHCOW) and Canadian Auto Workers (CAW) to provide evidence for worker compensation claims. Using facilitated hazard mapping, former Holmes workers graphically reconstructed their workplaces and detailed their exposures. Using facilitated body mapping, workers recorded and displayed their health problems. The study uncovered a grim pattern of occupational diseases. Following the release of the results, coupled with clinically confirmed diagnoses and corroborating evidence of exposure, hundreds of former Holmes employees and their families received compensation for occupational diseases that had previously gone unrecognized. PMID- 15281373 TI - The Swedish experience with asbestos: history of use, diseases, legislation, and compensation. AB - After World War II, large quantities of asbestos were imported to Sweden and used in construction and ship building. In 1976, the use of asbestos was for practical purposes prohibited. Today, the only exposures are environmental, from asbestos in place and when buildings are demolished or rebuilt, and there are very strict rules for such work. Consequently, it is assumed that the asbestos-related diseases will gradually disappear from society, but due to the long latency time, about 100 mesotheliomas still occur every year in Sweden, and so far there is no certain sign of a decrease in incidence. Compensation is from the state via general insurance and consists basically of compensation for lost income and medical costs. PMID- 15281374 TI - Asbestos, asbestos-related diseases, and compensation claims in The Netherlands. AB - In The Netherlands the number of asbestos-related diseases is increasing. An age cohort model predicts a steep rise in pleural mesothelioma deaths up to 490 cases per year among men, with a total death toll close over 12,000 cases during 2000 2028. In the past decade the number of compensation claims for asbestos-related diseases has more than doubled, with increasingly verdicts in favor of claimants. In addition to the medical information, information about the state of the art of preventive measures in different periods of time plays a decisive role in these claims. The use of asbestos in The Netherlands, the occurrence of asbestos related diseases, the national asbestos regulations, and the position of the claimants in asbestos lawsuits in The Netherlands are reviewed. PMID- 15281375 TI - The Dutch Institute for Asbestos Victims. AB - The primary goal of the Dutch Institute for Asbestos Victims is to compensate mesothelioma victims who have been exposed to asbestos in the workplace, while they are still alive, by acting as a neutral mediator between these victims and their (former) employers or their insurers. Representatives of victims, employees, employers, and insurers have agreed to cooperate in the formation and operation of the Institute. The process of reaching a financial settlement has been collectivized, standardized, pacified, and institutionalized. The difficulty of awarding compensation while victims are still alive has led to the Advance Payment Scheme. PMID- 15281376 TI - Asbestos victims support groups in England: a Manchester and Merseyside perspective. AB - The development, functions, and effectiveness of asbestos victims' support groups in Manchester and Merseyside, England, are described. PMID- 15281377 TI - Asbestos in Scotland. AB - This paper outlines the asbestos hazard in Scotland and draws upon a systematic oral history project to analyze from the workers' perspective the nature of exposure, the limitations of government regulatory initiatives, and the ramifications of contracting asbestos-related diseases for sufferers and their families. Current issues are investigated, stressing the agency of workers, trade unions, sympathetic local councils, and, especially, the victims' pressure groups. The occupational and environmental health threats of asbestos in Scotland remain significant, although recent E.U.- and U.K.-based decisions to ban further use of asbestos together with active campaigning by local activist groups have helped to reduce them. Mesothelioma mortality rates remain high, due to historic exposures, and much work remains to be done to reduce the number and plight of asbestos-exposed workers. PMID- 15281378 TI - Asbestos in Italy. AB - Asbestos-related diseases remain common in Italy due to past exposures that were tolerated by a government distracted and manipulated by multinational asbestos corporations. The incidence of asbestos-related cancers has taken on almost epidemic proportions, for example, in Casale Monferrato in northwest Italy, where Eternit remained in operation until 1985, and in Monfalcone in northeast Italy, where naval dockyards and related activities created pollution. Authorities took action only after public protests, trade union pressures, and campaigning by the families of victims. Now that a ban exists in Italy, it is vital that it be fully enforced to reverse the epidemic of mesothelioma, lung cancer, and other asbestos related diseases. PMID- 15281379 TI - Asbestosis in the Republic of Croatia. AB - Croatians have been exposed to asbestos in the shipbuilding and asbestos-cement industries since 1945. The first cases of asbestosis were reported in 1961; 317 cases were recorded from 1990 to 2000. The Croatian Cancer Registry recorded 248 malignant pleural mesotheliomas between 1991 and 1997, two thirds of which were attributable to occupational exposures to asbestos. The Croatian Asbestosis Patient Association was founded in 1998 to help victims. Croatian law defines the employer's responsibility for work-related health damage and compensation, but average legal proceedings for asbestosis claims take about seven years. Croatian law does not ban the manufacture and import of asbestos. Croatia as a transitional country is subject to socioeconomic pressures. Future approaches to the asbestos issue will depend on revised regulations, which are expected to conform to recommendations of the European Union by 2005. PMID- 15281380 TI - Asbestos banned in Argentina. AB - In 1997, Argentina gave priority to asbestos in its National Plan for the Sound Management of Chemicals, and it was the subject of a Technical Task Force on Occupational Cancer. After five years of public hearings in which government, workers, industry advocates, environmentalists, clinicians, scientists, and consumers participated, it was agreed that asbestos exposure is a risk factor for both workers and the general population, and that Argentina should provide to its people the same protections adopted by many developed countries. Pressure from asbestos industry groups initially delayed the inclusion of chrysotile asbestos in the proposed ban, but on January 1, 2003, the mining and import of all forms of asbestos were banned in Argentina. PMID- 15281381 TI - The phase-out of asbestos in the Australian manufacturing environment. AB - The importation of raw asbestos and asbestos-containing products into Australia was banned at the end of 2003. Despite a high incidence of asbestos-related disease, it was the threat of industrial action that eventually persuaded Australian governments to impose a ban. Significant government-sponsored reports in 1990 and 1999 had not convinced politicians that a phase-out of chrysotile was feasible. An impediment for government and industry was the risk of unemployment of asbestos workers. The threat of future asbestos exposures remains for maintenance workers. An example is given of the need for continued vigilance of workers and their unions before the manufacturing environment can be relatively asbestos-free. PMID- 15281382 TI - The epidemic of asbestos-related diseases in New Zealand. AB - New Zealand is a small country with a big asbestos disease problem. The lack of action on warnings in the 1960s and 1970s has led to epidemics of mesothelioma and asbestosis, which can be clearly documented via the death and cancer registers. In addition, an uncertain number of lung cancers due to asbestos exposure has occurred. The epidemic started in the 1980s, and will eventually have cost the lives of at least 2000 to 3000 workers. Prevention against ongoing exposures from asbestos installed in buildings is essential, and another key issue for New Zealand is to ensure that fair workers' compensation is provided to all victims of asbestos diseases. PMID- 15281383 TI - Asbestos in the sputum, crackles in the lungs, and radiologic changes in women exposed to asbestos. AB - In a rural community in South Africa historically exposed to asbestos environmentally and occupationally, 200 women who had worked with asbestos and applied for medical examination to determine compensable asbestos disease were evaluated. Clinical and radiologic evaluation, sputum collection, and microscopic analysis were done. A questionnaire elicited type of exposure, duration, decade of first work exposure, and environmental exposure. Crackles were present in the lungs of 166 women and asbestos fibers and ferruginous bodies were present in 122. Asbestosis was identified in 26 and plural plaques in 62. Auscultation for crackles (rales) is useful in the initial examination of former asbestos workers in rural communities of developing countries. PMID- 15281384 TI - Collaborative research, participatory solutions: research on asbestos in Kuruman, South Africa. AB - The 1998 South African National Asbestos Summit proposed a post-apartheid asbestos policy for the country. In the areas of environmental rehabilitation, health care, and compensation, it envisioned connecting asbestos mitigation to participatory development. In 2001, the Asbestos Collaborative, an international and interdisciplinary team, conducted follow-up research on the recommendations of the 1998 Summit, researching environmental, health, and compensation issues through consultation of documents and interviews with officials in urban areas and with people in Kuruman, a former crocidolite-mining site with high rates of asbestos-related disease. In Kuruman, local opinion supported the recommendations of the Asbestos Summit, insisting that policies to mitigate the problem of asbestos must also address poverty. In the wake of the 2001 research, a new organization, the Asbestos Interest Group (AIG), has been founded to facilitate grassroots participation in asbestos issues. One success of the AIG has been the settlement of a lawsuit by former workers against the former mining company in Kuruman. PMID- 15281385 TI - Chrysotile asbestos as a cause of mesothelioma: application of the Hill causation model. AB - Chrysotile comprises over 95% of the asbestos used today. Some have contended that the majority of asbestos-related diseases have resulted from exposures to the amphiboles. In fact, chrysotile is being touted as the form of asbestos which can be used safely. Causation is a controversial issue for the epidemiologist. How much proof is needed before causation can be established? This paper examines one proposed model for establishing causation as presented by Sir Austin Bradford Hill in 1965. Many policymakers have relied upon this model in forming public health policy as well as deciding litigation issues. Chrysotile asbestos meets Hill's nine proposed criteria, establishing chrysotile asbestos as a cause of mesothelioma. PMID- 15281386 TI - Open season on Brazil's labor inspectors. PMID- 15281387 TI - Recognition of static and dynamic images of depth-rotated human faces by pigeons. AB - In three experiments, we examined pigeons' recognition of video images of human faces. In Experiment 1, pigeons were trained to discriminate between frontal views of human faces in a go/no-go discrimination procedure. They then showed substantial generalization to novel views, even though human faces change radically as viewpoint changes. In Experiment 2, the pigeons tested in Experiment 1 failed to transfer to the faces dynamically rotating in depth. In Experiment 3, the pigeons trained to discriminate the dynamic stimuli showed excellent transfer to the corresponding static views, but responses to the positive faces decreased at novel viewpoints outside the range spanned by the dynamic stimuli. These results suggest that pigeons are insensitive to the three-dimensional properties of video images. Consideration is given to the nature of the task, relating to the identification of three-dimensional objects and to perceptual classifications based on similarity judgments. PMID- 15281388 TI - Reaction time signatures of discriminative processes: differential effects of stimulus similarity and incentive. AB - In three experiments with pigeons, the similarity of unreinforced test stimuli to a reinforced stimulus and the frequency of reinforcement associated with a stimulus were varied. The stimulus on each trial was a small spot that appeared in different hues or, in Experiment 3, different forms. Differential response frequency and reaction time (RT) patterns emerged: Changes in similarity affected the percentage of stimuli responded to but left the shape of RT distributions about the same, whereas changes in reinforcement shifted RT distributions but had little effect on the percentage of responses. When the similarity and reinforcement variables were applied to the same stimuli (Experiment 2), their effects were largely independent. A generalization procedure (Experiment 3) replicated the similarity effects of the initial discrimination procedure. The RT distributions were modeled by a diffusion process, and implications for a memory instance model were suggested. PMID- 15281389 TI - Stimulus salience and asymmetric forgetting in the pigeon. AB - Pigeons were trained using a symbolic delayed matching-to-sample procedure involving bright versus dim houselight samples. We hypothesized that when sample stimuli differ in salience, increasing the size of the retention interval will affect performance on trials initiated by the more salient sample only. In agreement with this prediction, accuracy following the dim sample did not decline as the retention interval increased, whereas accuracy following the bright sample declined to well below 50% correct. In a second experiment, the less salient (dim) sample from Experiment 1 was arranged as the more salient sample in a sample/no-sample procedure. Accuracy on dim sample trials then declined to well below 50% correct as the retention interval increased, whereas accuracy on no sample trials remained constant. The results suggest that when sample stimuli differ in salience, pigeons may transform the nominal discrimination task into a detection task in which they respond on the basis of the presence or absence of the more salient sample. PMID- 15281390 TI - Time-course of control by specific stimulus features and relational cues during same-different discrimination training. AB - We trained 7 pigeons to discriminate visual displays of 16 same items from displays of 16 different items. The specific stimulus features of the items and the relations among the items could serve as discriminative stimuli. Unlike in most studies of same-different discrimination behavior, we gave a small number of probe tests during each session of acquisition to measure the time-course of control by the learning of specific stimulus features and relational cues. Both the specific stimulus features and relational cues exerted reliable stimulus control, with the specific stimulus features exerting more control during the final three fourths of same-different learning. These findings replicate research suggesting that pigeons encode both the specific stimulus features and relational cues, and for the first time document the time-course of control by each kind of cue. PMID- 15281392 TI - Comparing demand functions when different price manipulations are used: does unit price help? AB - Six hens pecked a key (Experiment 1) or pushed a door (Experiment 2) to obtain food reinforcement. In both experiments and as an analogue of price changes, the response requirements were varied in two ways: by increasing the number of responses required and by increasing the required force of each response. The two price manipulations (response number and response force) had different effects on behavior and produced different-shaped demand functions when the rates of consumption were plotted logarithmically against the price analogues. Irrespective of response topography, when the number of required responses was varied, the data paths appeared linear, with slopes close to -1.0. When the required force of each keypeck and doorpush was varied, the data paths were clearly curved, with increasingly steep downward slopes as the force increased. Using the concept of unit price did not fully remove the different effects of the two price manipulations. Those differences are best attributed to the differing times needed in order to complete each response unit under those price manipulations. PMID- 15281391 TI - The relation of multiple-schedule behavioral contrast to deprivation, time in session, and within-session changes in responding. AB - Pigeons' keypecking was reinforced by food on baseline schedules of multiple variable interval (VI) x VI x and on contrast schedules of multiple VI x VI y. Deprivation of food was varied by maintaining subjects at 75%, 85%, and 95% (+/- 2%) of their free-feeding weights. Positive and negative behavioral contrast were observed. The size of the contrast was not systematically altered by changes in deprivation. Positive and negative contrast were both larger later in the session than they were earlier. Within-session decreases in responding were steeper for the baseline than for the contrast schedules for positive contrast. Within session decreases were steeper for the contrast than for the baseline schedules for negative contrast. These results were predicted by the idea that different amounts of habituation to the reinforcer during the baseline and contrast schedules contribute to behavioral contrast. The results show that contrast occurs under conditions that reduce the effect of the following component. The results support the assumption that positive and negative contrast are produced by symmetrical theoretical variables. PMID- 15281393 TI - Extinction and retraining of simultaneous and successive flavor conditioning. AB - In three experiments, rats received pairings of flavor conditioned stimuli with polycose unconditioned stimuli, in either a simultaneous or a sequential relation. Both temporal relations produced excellent conditioned increases in consumption of the flavors. Separate presentation of the flavors resulted in extinction in both cases. However, restoring the pairing of the flavor with polycose resulted in reconditioning only with the sequential, not with the simultaneous, relation. PMID- 15281394 TI - Memory priming and trial spacing effects in Pavlovian learning. AB - Conditioning trials that are massed in time produce less conditioning than those that are spaced in time. Four experiments with rat subjects examined whether a recent conditioning trial interferes with conditioning on the next trial by temporarily "priming" information in short-term memory (e.g., Wagner, 1978, 1981). We used appetitive conditioning procedures in which priming trials preceded target trials by 60 sec. When the priming trials were nonreinforced presentations of a conditioned stimulus (CS), the CS had to be the same CS as the one on the target trial to interfere with conditioning. When priming trials were actual CS-unconditioned stimulus (US) pairings, the CS identity did not matter, the US was the event that interfered with conditioning on the next trial. Reinforced trials reduced performance in a way that did not depend on context blocking. The results suggest that CS and US priming effects do contribute to conditioning deficits observed with massed trial procedures. The results are consistent with Wagner's (1981) "sometimes opponent process," or SOP, model, although a result that is paradoxical for the model suggests that recent USs may have motivational as well as memory effects. PMID- 15281395 TI - Trial number and compound stimuli temporal relationship as joint determinants of second-order conditioning and conditioned inhibition. AB - Two conditioned lick suppression experiments with rats used feature-negative training (A-footshock trials intermixed with nonreinforced XA presentations) to analyze the role of the number of XA compound presentations and the temporal relationship of the elements within the compound (simultaneous or serial) as determinants of the resulting behavioral control. Second-order conditioning (i.e., excitatory behavioral control by X) was observed to decline as the number of XA compound trials was increased. This decline was more rapid if X and A were presented simultaneously, as opposed to serially (i.e., X before A; Experiment 1). Conditioned inhibition to X, as assessed by a summation test (Experiment 1) and a retardation test (Experiment 2), increased with the number of XA trials and did so more quickly for simultaneous than for serial pairings of X and A. The results help to clarify previously discrepant findings regarding factors that promote excitation versus inhibition with this protocol. PMID- 15281396 TI - Time-place learning in the eight-arm radial maze. AB - Rats (n = 4) searched for food on an eight-arm radial maze. Daily 56-min sessions were divided into eight 7-min time zones, during each of which a different location provided food; locations were randomized across subjects before training. The rats obtained multiple pellets within each time zone by leaving and returning to the correct location. Evidence that the rats had knowledge about the temporal and spatial features of the task includes the following. The rats anticipated locations before they became active and anticipated the end of the currently active locations. The rats discriminated currently active locations from earlier and forthcoming active locations in the absence of food transition cues. After the rats had left the previously active location, they visited the next correct location more often than would be expected by chance in the absence of food transition cues. The rats used handling or opening doors as a cue to visit the first location and timed successive 7-min intervals to get to subsequent locations. PMID- 15281397 TI - In vitro conidial germination in Arthrinium aureum and Arthrinium phaeospermum. AB - This paper describes the microscopic details of conidial germination and the influence of pH, sodium chloride concentration, 3% glucose, 3% saccharose and ultraviolet irradiation on the conidial germination in Arthrinium species. Under laboratory conditions, germination started after an incubation period of 90 minutes in 2% malt extract broth at 25 degrees C. In vitro, the conidia of Arthrinium species have a very low percentage of germination (A. phaeospermum: 7.9%; A. aureum: 15.8%). Conidia of this genus have a characteristic equatorial slit. Conidia may break spontaneously at this slit, releasing their cytoplasmic contents. Arthrinium phaeospermum attains its optimum germination percentage when its conidia are suspended in a sterile saline solution (pH 3.5) in a water bath at 20 degrees C for 15 minutes before being inoculated on 2% malt extract agar. Conidial suspensions of A. aureum may be held in the same conditions, but for 30 minutes. PMID- 15281398 TI - Role of antibiosis in the biological control of spot blotch (Cochliobolus sativus) of wheat by Chaetomium globosum. AB - Chaetomium globosum Kunze, has been identified as a potential antagonist of Cochliobolus sativus (S. Ito & Kurib.) Deschler ex Dastur. (Syn = Drechslera sorokiniana). Production of antifungal compounds by Chaetomium globosum (Cg) and their role in suppression of spot blotch of wheat caused by this fungus under in vitro and in vivo has been evaluated. Interaction between Chaetomium globosum isolates and C. sativus showed mycoparasitism by isolates Cg 1 and Cg 6 whereas isolates Cg 2, Cg 3, Cg 4 and Cg 5 showed antibiosis. Syringe filtered culture extracts of Cg 2 completely inhibited mycelial growth of C. sativus in liquid broth. In vitro bioassays were undertaken by amending the medium with crude extracts and agar diffusion method in order to assess the fungistatic activity of crude extracts from culture filtrates of different isolates of Chaetomium globosum. Significant differences in antagonism between isolates were observed. Antifungal metabolite profiling, on TLC (Thin Layer Chromatography) plates identified 13 compounds in isolate Cg 2, 11 compounds in Cg 3 and 7 compounds in Cg 6. Isolate Cg 1 produced only two faint bands and Cg 5 produced two bands of the same Rf value but of higher intensity. The production of antifungal compounds by isolates was positively correlated with antagonism to C. sativus on seedlings in glasshouse studies. The results showed high antifungal metabolite production by isolate Cg 2, which also gave maximum bioefficacy under laboratory and glasshouse conditions. PMID- 15281399 TI - Frequency and risk factors of dermatophytosis in students living in rural areas in Eskisehir, Turkey. AB - Our study included 2384 students from five villages around Eskisehir, Turkey. We asked every student for their personal identification and also for their sanitation in order to get an idea about dermatophytosis. Samples taken from suspicious lesion were collected and inoculated onto Sabouraud dextrose agar slants. For identification of fungi which were grown, macroscopic appearance of colonies, microscopic examination and biochemical tests were used. We found suspicious lesions in 245 (10.3%) and diagnosed dermatophytosis in 86 (3.6%) of the students. The dermatophyte species were Trichophyton rubrum 37 (43%) at first, Trichophyton mentagrophytes 17 (19.8%), Microsporum canis 11 (12.8%), Microsporum gypseum 8(9.3%), Epidermophyton floccosum 6 (7%), Trichophyton verrucosum 6 (7%) and Trichophyton violaceum 1 (1.1%). Tinea pedis (59.3%) was the most frequent clinic form of dermatophytosis, followed by tinea corporis (22.1%), tinea capitis (9.3%), tinea manum (7.0%) and tinea unguium (2.3%). Older age, male gender, poor hygiene, living in dormitory, low level mother education, history of dermatophytosis within family and sanitary conditions were computed as independently variables associated with dermatophytosis infection. For prevention and control of dermatophyte infection in children living rural areas, field studies should be done and sanitary conditions should be improved. PMID- 15281400 TI - Occurrence of Malassezia species in healthy and dermatologically diseased dogs. AB - The presence of Malassezia spp. yeasts was investigated in dermatological specimens of 224 dogs, 164 dermatologically diseased and 60 normal dogs. Subjects included in the study were of different breed, age, sex and habitat. Malassezia spp. positive cultures were obtained in 142 (63.4%) specimens: 67.6% from dermatologically diseased subjects and 51.6% from healthy dogs. Malassezia pachydermatis, either as a pure culture or in association with lipid-dependent species, was identified in 138 (97%) specimens. Malassezia furfur was identified in 69 (48.6%) specimens and was associated with other Malassezia species in 68 dogs, as a pure culture in one subject: at the best of our knowledge, this species was identified before as the sole species from canine dermatitis. Malassezia sympodialis was identified in 11 (7.7%) specimens, always in association with other species: it was never isolated from kennel dogs. Statistical analysis of data showed a very significant difference (P < 0.01) in the prevalence of isolation of Malassezia spp. between animals with and without dermatological signs, and in the distribution of cultural burden between diseased and healthy dogs. A statistically significant difference (P<0.05) was also detected in the group of animals between 1- and 5-years of age. No significant difference was found between male and female dogs. PMID- 15281401 TI - Gas gangrene and necrotizing fasciitis in the upper extremity. AB - Necrotizing soft tissue infections encompass a wide variety of clinical syndromes resulting from introduction of various pathogens into injured or devitalized tissue. The extent of microbial involvement in such tissue may range from simple contamination to overt and progressive local tissue necrosis, which, if untreated, may lead to septicemia and death. Early differentiation among these infections is not always possible, as there are overlapping classification criteria. These infections exist along a continuum of clinical severity with different etiological agents and associated medical conditions. The often subtle clues heralding the presence of a necrotizing soft tissue infection must be sought so that expeditious surgical debridement and broad-spectrum antibiotic management are initiated. Although experience enables the clinician to make a specific diagnosis based on early findings, aggressive and proper treatment of suspected infections remains the priority. The purpose of the article is to provide an overview of necrotizing soft tissue infections in the upper extremity, focusing on gas gangrene, or clostridial myonecrosis, and necrotizing fasciitis, to facilitate early diagnosis and optimal management of these lethal diseases. PMID- 15281402 TI - Magnetic resonance imaging of the coracoclavicular ligaments: its role in defining pathoanatomy at the acromioclavicular joint. AB - Four patients with acromioclavicular joint injuries (one type II, two type III, one type V), two patients without acromioclavicular joint injury, and a fresh frozen cadaver underwent magnetic resonance imaging (MRI) and plain radiographs. The normal conoid and trapezoid ligaments were easily identified in the cadaver and the two uninjured patients. Magnetic resonance imaging revealed disruption of both coraclavicular ligaments in the three patients with type II and type III injuries. However, the patient with the type V injury had disruption of the trapezoid ligament alone. Thus, the grade of injury, as determined by the change in the coracoclavicular interval onplain radiography and defined by the Rockwood classification system, failed to correlate with the pathoanatomy seen on MRI in two of the four injured patients. These findings suggest that improvements in the classification of these injuries may be necessary. PMID- 15281403 TI - Longitudinal examination of health outcomes associated with botulinum toxin use in children with cerebral palsy. AB - A prospective cohort study with annual follow-up was conducted on 172 children with spastic type cerebral palsy receiving botulinum toxin type A (BTX) injections for spasticity management. A mixed modeling procedure was used to identify changes in both physical functioning outcomes for the child (using the WeeFIM measure) as well as quality of life of the parent caregiver (using the Stein and Reissman Impact on the Family Scale) with increasing utilization of BTX injections. The study found that each additional BTX injection administration was associated with a 2.3% improvement in the WeeFIM compared to the average baseline score (p < .01). Similarly, the study found an improvement of 2.5% compared to baseline in the parent's overall perception of the severity of the child's condition with each additional BTX injection administration (p < .001). These findings suggest that BTX injections may be associated with beneficial outcomes in childhood spasticity. PMID- 15281404 TI - Ankle arthrodesis. AB - This review article deals with ankle fusions, their indications, preoperative assessment, surgical technique, and postoperative treatment along with a discussion of complications and long-term results. PMID- 15281405 TI - The natural history of the periacetabular fragment following Ganz osteotomy. AB - The vascularity of the acetabular free fragment in Ganz osteotomies has remained a concern. This study aims to assess the role of MR imaging in the postoperative evaluation of Ganz osteotomies. Twenty patients (19 females, 1 male), average age 24 years (range, 12-36 years), had sequential magnetic resonance imaging studies of the pelvis at 6 weeks and 6 months following Ganz osteotomies. Normal healing with no evidence of periosteotomy edema was seen in 17 patients at 6 weeks. Three patients showed evidence of reduced vascularity. In two of these, there were focal changes suggestive of subclinical ischemia. The other had gross signal changes in the osteotomy fragment suggestive of diffuse ischemia. The patients with focal changes were asymptomatic and had normal 6-month scans. The patient with diffuse changes complained of persistent groin pain, which resolved after 4 months. The 6-month scan showed some persistent vascular changes. The scan at 1 year showed complete resolution. The study suggests that Ganz osteotomy has minimal effect on the vascularity of the acetabular free fragment. PMID- 15281406 TI - Wear mechanisms in ceramic hip implants. AB - The wear in hip implants is one of the main causes for premature hip replacements. The wear affects the potential life of the prosthesis and subsequent removals of in vivo implants. Therefore, the objective of this article is to review various joints that show lower wear rates and consequently higher life. Ceramics are used in hip implants and have been found to produce lower wear rates. This article discusses the advantages and disadvantages of ceramics compared to other implant materials. Different types of ceramics that are being used are reviewed in terms of the wear characteristics, debris released, and their size together with other biological factors. In general, the wear rates in ceramics were lower than that of metal-on-metal and metal-on-polyethylene combinations. PMID- 15281407 TI - Preliminary experience with a new surgical treatment for dysphagia due to anterior cervical osteophytes. AB - A technique for surgical treatment of anterior cervical osteophytes is presented. A midline trough is created in the osteophytes using a burr under fluoroscopy down to the anterior cervical line. A rongeur is used to remove the remaining osteophytes while protecting the lateral soft tissues. Two patients presented with symptoms of progressive dysphagia secondary to anterior cervical osteophytes. Each underwent surgical ostectomy without complication after failing conservative treatment. This technique provides a safe, effective method to remove anterior cervical osteophytes. PMID- 15281408 TI - Unusual presentation of a giant cell tumor of the tendon sheath in the foot. AB - Giant cell tumor of the tendon sheath has frequently been described in the hand, but it is much less common in the foot. The case report presents an apparent cystic bony lesion of the proximal phalanx of the second toe of the foot. The operative findings and histological examination revealed a giant cell tumor of the tendon sheath that had eroded the phalanx. The case demonstrates that in the differential diagnosis of a cystic lesion of bone, extrinsic soft-tissue lesions should be considered. PMID- 15281409 TI - Treatment of close-range, low-velocity gunshot fractures of tibia and femur diaphysis with consecutive compression-distraction technique: a report of 11 cases. AB - Lower extremity injuries secondary to close-range, low-velocity gunshot wounds are frequently seen in both civilian and military populations. A close-range, low velocity injury produces high energy and often results in comminuted and complicated fractures with significant morbidity. In this study, four femoral, four tibial, and three combined tibia and fibular comminuted diaphyseal fractures secondary to close-range, low-velocity gunshot wounds in 11 military personnel were treated with debridement followed by compression-distraction lengthening using a circular external fixator frame. Fracture union was obtained in all without significant major complications. Fracture consolidation occurred at a mean of 3.5 months. At follow-up of 46.8 months, there were no delayed unions, nonunions, or malunions. Minor complications included four pin-tract infections and knee flexion limitation in two femur fractures. Osteomyelitis and deep soft tissue infection were not observed. This technique provided an alternative to casting, open reduction internal fixation, or intermedullary fixation with an acceptable complication rate. PMID- 15281410 TI - Periprosthetic fracture of the femur after total hip arthroplasty occurring in winter activities: report of two cases. AB - Periprosthetic fractures are uncommon after total hip replacement surgery and are most often associated with loosening or osteolysis. In a review of Mayo Clinic records, the cumulative incidence of femoral fractures after primary uncemented prostheses was only 0.4% (4). No periprosthetic fracture associated with sports participation has been previously reported in the literature. When advising patients about return to sports and recreational activities after total hip replacement, concerns fall into two main categories: 1) wear of the bearing surface(s) and secondary ramifications such as early failure or osteolysis, and 2) dislocation or fracture of the prosthesis or periprosthetic bone. The former concerns have been previously examined (5), but the latter have not been reported to date. These case reports describe a complication that may occur in total hip arthroplasty in those patients who return to winter sports and recreational activities. Although at intermediate follow-up there does not appear to have been irreversible damage for these patients, it is imperative to warn patients that activities that place the patient at risk of trauma may compromise the longevity of the artificial joint. This information can be used in helping patients understand the risks associated with athletic activity after total hip arthroplasty, which is a major goal of current recommendations for advising patients after this type of surgery. PMID- 15281411 TI - Giant cell tumor of tendon sheath simulating giant cell tumor of bone: report of a case. AB - A 24-year-old male patient presented with a painful eccentric lytic lesion of the proximal tibial epiphysis with a soft tissue component. Clinical and radiological assessment led to the tentative diagnosis of aggressive giant cell tumor of bone. The patient was treated with curettage, high-speed burr, and cementation after intraoperative pathology consultation. The final pathological report indicated that the tumor was giant cell tumor of the tendon sheath with bone invasion. Although uncommon, GCTTS should be considered in the differential diagnosis of such lesions when there is a prominent soft tissue component. Although the resection was intralesional, the thermal effect of the cementation of the involved cavity and complete removal of the tendon sheath may allow successful local control conjecture of lesions that otherwise present with clinical and radiographic findings suggesting giant cell tumor of the bone. PMID- 15281412 TI - An American cultural view of the British DSPD proposals. PMID- 15281413 TI - A cautionary lesson from simulated patients. PMID- 15281414 TI - Willingness and competence of depressed and schizophrenic inpatients to consent to research. AB - In this study, the willingness of psychiatric inpatients to volunteer for research and their capacity to consent to and distinguish between protocols offering different levels of risk and benefit were assessed. Twenty-two inpatients with major depressive disorder, 21 inpatients with schizophrenia, and 21 community control subjects were asked to consider participation in a lower risk study offering the potential for direct medical benefit and a higher-risk study offering no direct medical benefit. Consent-related capacities were assessed with the MacArthur Competence Assessment Tool-Clinical Research. Depressed inpatients, while having a greater degree of impairment than control subjects, still demonstrated relatively high decision-making capacity and were able to distinguish levels of risk between studies. Their pattern of preferences did not differ from control subjects. However, they were more likely to decline to participate in the research, being six times more likely to decline the lower risk study and 1.4 times more likely to decline the higher-risk study. Schizophrenic subjects demonstrated greater impairments in decision-making capacity and were even more likely than depressed subjects to decline to participate. PMID- 15281415 TI - Countering countertransference, II: beyond evaluation to cross-examination. AB - Countertransference is a clinical term introduced by Freud in 1909. For years, despite mounting criticism, forensic psychiatrists borrowed this clinical concept to explain their emotional experiences and responses to examinees' emotions and behavior. The authors describe the impact of examinee and nonexaminee factors during evaluations and beyond, including during trial and while providing forensic testimony. The suggestion is made that using the term countertransference in forensic psychiatry can be problematic. The authors delineate the complexities of the term as related to forensic psychiatry and consider modified terms to provide a better explanation of these concepts in forensic contexts. PMID- 15281416 TI - Commentary: "Countertransference" on trial--witness or defendant? PMID- 15281417 TI - Use of the Mini International Neuropsychiatric Interview (MINI) as a screening tool in prisons: results of a preliminary study. AB - The authors describe a pilot study in which the Mini International Neuropsychiatric Interview (MINI) was used to assess a random sample of offenders newly committed to the Iowa Department of Corrections. Following sessions in which correctional personnel were trained to administer the MINI, the instrument was administered to 67 offenders. The interview took from 20 to 105 minutes (mean, 41 minutes) to administer, and all but 13 (19%) offenders were positive for a lifetime MINI disorder. Twenty-six (39%) subjects had a lifetime mood disorder, 20 (30%) a lifetime anxiety disorder, 12 (18%) a lifetime psychotic disorder, and 53 (79%) a substance use disorder. Seven (10%) subjects met criteria for a lifetime attention deficit hyperactivity disorder, while 13 (19%) had a lifetime antisocial personality disorder. Subjects had a mean of 2.8 disorders. The potential use of the MINI as a screening tool in prison settings is discussed. PMID- 15281418 TI - The use of restraint and seclusion in different racial groups in an inpatient forensic setting. AB - The purpose of this study was to determine if physical restraint and/or seclusion had been used with different frequencies in patients of different racial groups in an inpatient forensic psychiatry facility. The method used was a retrospective correlational study of all inpatients (n = 806) treated from January 1993 through August 2000 at Kirby Forensic Psychiatric Center, a maximum-security inpatient forensic facility in Ward's Island, NY, near New York City. Episodes of restraint and/or seclusion were measured in each racial group. The number of violent incidents involving patients of each racial group was also measured. Racial groups at Kirby did not differ significantly from each other in number of violent incidents nor in the number of episodes of restraints. However, Asians and blacks as racial groups were more likely to have been secluded than were other racial groups. This difference did not persist when the number of incidents of seclusion was considered individually rather than for entire racial groups. PMID- 15281419 TI - Delusions, substance abuse, and serious violence. AB - The objective of the study was to learn how delusions, substance abuse, and violence are related. The sample was 90 hospitalized patients with adequate descriptions of mental status when violent. Data sources were risk assessment based on record review and patient and staff interviews. The data include history of violence and substance abuse, diagnosis, and demographic and legal status. Delusions were definitely or questionably present in 73.3 percent and absent in 26.7 percent of violent episodes; 83.5 percent of delusionally violent patients had a history of substance abuse. These results support the importance of substance abuse in relation to violence by psychiatric patients. Delusions alone were infrequently related to violence, but when present appeared almost always to drive the violent behavior. PMID- 15281420 TI - Impact of Winko on absolute discharges. AB - In Canada, case laws have had a significant impact on the way mentally ill offenders are managed, both in the criminal justice system and in the forensic mental health system. The Supreme Court of Canada's decision with respect to Winko has set a major precedent in the application of the test of significant risk to the safety of the public in making dispositions by the Ontario Review Board and granting absolute discharges to the mentally ill offenders in the forensic health system. Our study examines the impact of the Supreme Court of Canada's decision before and after Winko. The results show that the numbers of absolute discharges have increased post-Winko, which was statistically significant, but there could be other factors influencing this increase. PMID- 15281421 TI - Organized psychiatry and the death penalty: an introduction to the special section. PMID- 15281422 TI - Now is the time for AAPL to demonstrate leadership by advocating positions of social importance. AB - The American Academy of Psychiatry and the Law (AAPL) and other medical organizations have not taken a position on the abolition of capital punishment because of a long-standing tradition of remaining neutral on "nonmedical" societal issues that are highly divisive. It is the authors' contention that taking a stand on vital social issues that are clearly in the public interest is wholly consistent with the stated purposes of AAPL and that the time has come for an open and frank discussion by the membership on the merits of altering its policy, with particular focus on eliminating the death penalty. The present article explains why capital punishment can no longer be considered a nonmedical societal issue and why AAPL must awaken to take on controversial matters such as this one. For AAPL to continue to avoid this debate and silence any attempt to organize opposition to the current status quo will only serve to embolden those who argue in favor of the death penalty. Such continued silence betrays any notion of neutrality and is an abdication of the canons of medical ethics we have all sworn to uphold. PMID- 15281423 TI - AAPL and sociopolitical policy. AB - It has been part of the role of medical organizations in Western nations to develop position statements on various sociopolitical issues. Both the American Psychiatric Association and the American Medical Association have established position statements related to social policy, including capital punishment. In 2001, AAPL endorsed a moratorium on capital punishment. Recent calls for AAPL to endorse a recommendation to abolish the death penalty have produced further discussion and a diversity of opinion. The absence of a clear process to develop policy within AAPL complicates both the nature and the resolution of the discussions. Three questions are posed that will help AAPL shape this debate: (1) Do we have anything to contribute? (2) Is there a position consistent with our values? (3) Would a position promote or harm the goals and mandate of the organization? PMID- 15281424 TI - The Royal College of Psychiatrists and the death penalty. AB - The Royal College of Psychiatrists recently issued a revised statement on its position concerning capital punishment. The College proposes to support psychiatrists who refuse to be involved in the capital process, but accepts that some may take up limited involvement in the manner set out in the document. The Royal College is the professional body for psychiatric practitioners in the United Kingdom and Ireland. Almost no public statements are issued from the College without first being deliberated on within at least two of its three major committees. The new document on capital punishment remains in the spirit of the previous ones. The topic of capital punishment is noncontroversial within the British medical profession. In all European countries, capital punishment is against the law, because there is an overarching directive from the Council of Europe (a wide group of nations, wider than the European Union) insisting that it be abolished. PMID- 15281425 TI - Forensic psychiatry and political controversy. AB - This article gives a U.K.-based perspective on the involvement of forensic psychiatry organizations in questions of political controversy. Medical professional bodies are fundamentally concerned to uphold good standards of clinical practice and patient welfare, and to uphold professional medical ethics. In our specialty, when acting as individual expert witnesses, we seek to serve the courts with objectivity and respect for the law. However, as members of our professional bodies we have a legitimate medical concern about how the law affects the mentally disordered as a class. We should articulate a collective view about what treating the mentally disordered justly and appropriately in the legal system means and should challenge the law when it fails to achieve this. PMID- 15281426 TI - Disability and psychotherapy: a response to Bursztajn et al. AB - In their recent Analysis and Commentary article Bursztajn and colleagues argue persuasively that to provide comprehensive independent medical evaluations (IMEs), forensic psychiatrists must take into account the managed care context of a claimant's case. The author of this article agrees with that assessment, but adds that another significant problem in his practice occurs when disability claimants who have severe disabling personality disorders resist psychotherapy and when, as is often the case, there are no experienced psychotherapists available to provide the needed treatment. PMID- 15281427 TI - The question of involuntary treatment of nondangerous defendants who are found to be "incompetent to stand trial". PMID- 15281429 TI - Sarcoidosis and tuberculosis: epidemiological similarities and dissimilarities. A review of a series of studies in a Japanese work population (1941-1996) and the general population (1959-1984). AB - AIM: The aim of this review is to discuss the epidemiological relationship between sarcoidosis and tuberculosis. METHODS: We have used a series of health surveillance data in a Japanese work population of 460,000 employees including 70,000 working in Tokyo (1941-1996) and the data from a nation-wide sarcoidosis survey in the general population (1959-1991). The work population was annually x rayed and tuberculin tested. The data of primary tuberculosis were obtained from a 17 year cohort study of tuberculin positive converters and primary pulmonary sarcoidosis data from the registry of the disease (1952-1996) in the same work population. Hilar lymphadenopathy (HL) was observed as a common marker of the two diseases. RESULTS: 1) Sarcoidosis HL was not detected in the work population until tuberculosis HL decreased. 2) BHL was rare in primary tuberculosis, but occurred in 95.5% of sarcoidosis subjects. 3) In both diseases, HL resolved in a few years, though accompanying extra-pulmonary involvements delayed the resolution of sarcoidosis BHL. 4) The grade of tuberculin sensitivity prior to sarcoidosis was not a risk factor for developing sarcoidosis. 5) Several well documented sarcoidosis cases remained tuberculin-negative before and at the time of diagnosis of the disease and after resolution of pulmonary involvement. 6) Age specific incidence curves showed a mono-modal curve in tuberculosis and a bimodal curve in sarcoidosis. 7) Tuberculosis prevalence was higher in the South of Japan, while sarcoidosis was higher in the North. CONCLUSIONS: These epidemiological dissimilarities do not support a tuberculosis etiology of sarcoidosis. PMID- 15281430 TI - Diffuse panbronchiolitis. AB - Diffuse panbronchiolitis (DPB) is an idiopathic inflammatory disease, largely restricted to Japan, that is characterized by progressive suppurative and obstructive airway disease, which, if left untreated, progresses to bronchiectasis, respiratory failure, and death. The lesion was first described in the early 1960s. In 1969 the name diffuse panbronchiolitis (DPB) was proposed to distinguish it from chronic bronchitis. Diffuse refers to the distribution of the lesions throughout both lungs, and pan refers to the involvement of inflammation in all layers of the respiratory bronchioles. Its distinctive imaging and histologic features, the coexisting sinusitis, and the isolation of Haemophilus influenzae and Pseudomonas aeruginosa in the sputum should enhance disease recognition. Neutrophils and T-lymphocytes, particularly CD8- cells, together with cytokines IL-8 and macrophage inflammatory protein-1 are believed to play key roles in the development of this disease. Significant improvement in the prognosis of this potentially fatal disease has been reported after the use of long-term therapy with macrolide antibiotics, the effect of which is attributed to an anti-inflammatory and immunoregulatory action. PMID- 15281431 TI - Th1 cytokine pattern (IL-12 and IL-18) in bronchoalveolar lavage fluid (BALF) before and after treatment with interferon gamma-1b (IFN-gamma-1b) or colchicine in patients with idiopathic pulmonary fibrosis (IPF/UIP). AB - BACKGROUND AND AIM: Characterization of the biologic effects of Th1 cytokines will enhance the understanding of idiopathic pulmonary fibrosis (IPF) pathogenesis and treatment selection. Th1 response is characterized by increased expression of IFN-gamma, interleukin (IL)-12 and IL-18. The present study aims to evaluate the role of Th1 cytokines and their possible changes in the bronchoalveolar lavage fluid (BALF), before and after treatment with IFN-gamma-1b or colchicine. PATIENTS AND METHODS: We studied prospectively 10 patients (8 male, 2 female) of median age 67 yr with histologically confirmed IPF/UIP. Patients were randomly assigned to receive either IFN-gamma-1b 200 microg sc (5 patients) or colchicine 1 mg qd (5 patients) plus prednisone 10 mg qd. BALF IL-12 and IL-18 levels were measured before and after treatment. RESULTS: BALF IL-12 levels before and after treatment did not differ significantly between the two treatment groups. However, BALF IL-18 levels were significantly decreased after treatment with IFN-gamma-1b (mean +/- SD, 58.4 +/- 15.6 pg/mL vs 42.8 +/- 4.90 pg/mL, p < 0.05). A significant difference was also found after treatment with colchicine (mean +/- SD, 66.8 +/- 36.9 pg/mL vs 42.6 +/- 1.08 pg/mL, p < 0.01). A significant correlation was found between IL-18 BALF levels and the BALF neutrophils (r = 0.75, p = 0.024). CONCLUSION: Our data suggest the potential role of IL-18 as an inflammatory marker in the pathogenetic pathway of IPF such as its possible downregulation by IFN-gamma-1b treatment. Further studies are needed in a higher number of patients in order to define the precise role of both cytokines during the immunoregulatory response with IFN-gamma-1b. PMID- 15281432 TI - BAL cytokine profile in different interstitial lung diseases: a focus on systemic sclerosis. AB - BACKGROUND AND AIM: Fibrosing alveolitis develops in up to 80% of systemic sclerosis patients (SSc) but progression to end stage fibrosis occurs in about 15% of cases. Mechanisms leading to the process remain mostly unknown. We compared cytokine profiles of broncho-alveolar lavage fluids (BAL-f) from patients with SSc associated interstitial lung disease (SSc-ILD) (n. 34), idiopathic pulmonary fibrosis (IPF) (n. 13), stage II sarcoidosis (n. 14) and 9 controls. METHODS: Interleukin (IL) 8, monocyte chemoattractant protein 1 (MCP 1), gamma-interferon (IFN-gamma), IL12, IL18 and IL10 and transforming growth factor-beta (TGF-beta) were assessed by ELISA in concentrated BAL-f. RESULTS: Levels of IL8 and MCP-1 were significantly elevated in SSc-ILD and in IPF as compared with controls (Mann Whitney test p < 0.05), while MCP-1 values were significantly lower in SSc-ILD than in IPF. A significant correlation between neutrophils and IL8 levels (p = 0.047), as well as between eosinophils and MCP-1 levels (p = 0.004) was also observed. IFN-gamma levels were slightly higher than normal only in sarcoidosis (p = 0.06), whereas IL12 levels increased both in sarcoidosis and SSc-ILD (p < 0.05). No differences were found in IL18 and TGF beta levels. Finally, IL10 levels were higher in SSc-ILD and sarcoidosis than in controls and IPF (p < 0.05). CONCLUSION: BAL-f cytokine profile differentiates ILD associated with SSc from IPF. The lower expression of MCP-1 and the higher expression of the anti-fibrotic IL12 and the anti-inflammatory IL10, observed both in sarcoidosis and in SSc-ILD, could account for the better prognosis of these ILDs. Further longitudinal studies are required to confirm whether a different cytokine phenotype may be considered predictive of clinical outcome in SSc-ILD. PMID- 15281433 TI - The organ-specific extrapulmonary presentation of sarcoidosis: a frequent occurrence but a challenge to an early diagnosis. A 3-year-long prospective observational study. AB - BACKGROUND AND AIM: The percentage of organ-specific extrapulmonary presentation of sarcoidosis has usually been estimated on the basis of retrospective studies, with figures varying in the range of 7-22%. We have hypothesized that a prospective study could give a higher figure, and could better outline the variable pattern of extrapulmonary presentation. MATERIAL AND METHODS: A careful examination with particular interest to the possible extrapulmonary signs or symptoms of the disease, even if previously unrecognized, was carried out in 204 consecutive sarcoid patients (mean age 37 +/- 11) seen between 2000 and 2002. Some of them had already visited our clinic in previous years. Their median follow up in our clinic was 34 months, while the median duration of disease was 60 months. RESULTS: An organ-specific extrapulmonary presentation did occur in 73 patients (35.78%). The most frequent extrapulmonary presentation was due to skin lesions (different from erythema nodosum) and to peripheral lymph nodes (8% each). Less frequent were renal stones (4%), uveitis (3%) and others (12%). The extrapulmonary presentation may be a challenge to the diagnosis. In our series an early diagnosis was reached only in 38 of 73 patients. In the other 35 patients, the diagnosis was reached later, after a time interval ranging from 6 months to 20 years. The lung involvement was detected early in 33 patients with extrapulmonary presentation. In 35 other patients the pulmonary involvement appeared after a median time interval of 24 months (range 6-240, interquartiles 15-60). In 5 patients, no pulmonary changes ever appeared after a long term follow up of 20 months, 5 years (2 patients), 10 years and 29 years respectively. CONCLUSION: An organ-specific extrapulmonary presentation is common in sarcoidosis, but may go unrecognized for years. PMID- 15281434 TI - The relationship between fatigue and clinical parameters in pulmonary sarcoidosis. AB - BACKGROUND AND AIM OF THE WORK: Studies on the relationship between fatigue and clinical parameters are sparse. In the present study this relationship was examined in a systematic way. METHODS: Patients with time since diagnosis < or = 2 years, visiting the outpatient clinic of the University Hospital Maastricht (n = 60; 34 untreated, 26 treated) were clinically evaluated and completed the Fatigue Assessment Scale (FAS). A representative sample of the Dutch population (n = 1893) also completed the FAS. Pulmonary disease severity was estimated from lung function test results and measures of metabolic derangement. Acute phase response markers high-sensitivity C-reactive protein (hs-CRP), serum amyloid A (SAA) and sarcoidosis activity parameters, soluble interleukin-2-receptor (sIL2R), and angiotensin-converting enzyme (ACE) were also measured. RESULTS: Only 27% of the sarcoidosis patients were diagnosed as non-fatigued (FAS score < 22), compared to 80% in the control population (n = 1893). In the sarcoidosis patients no sex differences and no differences in fatigue scores between the treated and the untreated groups were found. Patients with fatigue (FAS-score > or = 22) had lower DLCO values (p < 0.05). However, none of the tested clinical or serological parameters appeared to be a significant predictor of fatigue. CONCLUSIONS: In the present study, it was confirmed that fatigue is a major problem in sarcoidosis. The extent of fatigue could not be explained by clinical parameters. Thus, up to now, no clinical or physiological variable seems useful in predicting which patients are fatigued. In this light, the Fatigue Assessment Scale might be considered as a supplementary tool in sarcoidosis. PMID- 15281435 TI - Sleep disturbances associated with periodic leg movements in chronic sarcoidosis. AB - BACKGROUND AND AIM OF THE WORK: Many sarcoidosis patients suffer from fatigue and sleep disturbances. Recently, it was demonstrated that obstructive sleep apnea (OSA) is rather common in sarcoidosis. Moreover, sheet intolerance and painful legs are frequently reported in sarcoidosis patients. These symptoms might interfere with sleep quality. METHODS: In order to determine the relationship between objective and subjective sleep disturbance full polysomnography, including leg EMG analysis, was performed in 46 chronic sarcoidosis patients indicating awakening unrefreshed in the morning. RESULTS: In 20 (44%) patients OSA activity [60% with PLM (n=12), 40% without (n = 8)] was demonstrated, while in 7 patients (15%) significant PLM without OSA were found. In 19 patients (42%) no OSA or PLM activity was present. Moreover restless legs (RLS) were reported by 52% of the patients (45% in OSA; 71% in PLM; 47% in others). Distribution of sleep stages and sleep fragmentation was comparable in all groups. In a healthy snoring control group (n = 102) a prevalence of PLM was found in 13.7% (17.8% in men; 3.4% in women), while RLS were only reported by 1.4% (men) and 6.9% (women). CONCLUSIONS: Sleep disturbance (OSA and/or PLM) and RLS were demonstrated in more than half of the studied sarcoidosis patients. A high prevalence of RLS or PLM (primary and secondary) has not been reported before in sarcoidosis. Further studies are needed to establish whether RLS, OSA and/or PLM might contribute to fatigue and whether fatigue complaints improve after treatment of RLS/PLM/OSA. PMID- 15281436 TI - Transbronchial needle aspiration improves the diagnostic yield of bronchoscopy in sarcoidosis. AB - BACKGROUND AND AIM: Transbronchial needle aspiration (TBNA) is a minimally invasive bronchoscopic procedure that allows sampling of hilar and mediastinal lymph nodes in close contact with the airways. We undertook this study to assess the value of TBNA in the diagnosis of sarcoidosis manifesting with intrathoracic lymphadenopathies (stages I and II), and to compare its yield with that of transbronchial lung biopsy (TBLB). METHODS: The results of bronchoscopy with combined TBNA and TBLB in 32 patients with stage I or II sarcoidosis were retrospectively analyzed. RESULTS: Sensitivity was 65.6% for TBNA (stage I, 82.3 %; stage II, 46.6%), and 62.5% for TBLB (stage 1, 52.9%; stage II, 73.3%). The combination of the two methods was associated with the highest diagnostic yield (93.7% overall sensitivity), and allowed significantly better results over both TBNA alone (93.7% vs 65.6%; p = 0.011) and TBLB alone (93.7% vs 62.5%; p = 0.005). CONCLUSIONS: The results of our study suggest that a diagnostic approach combining TBNA and TBLB is safe and effective in the setting of stage I and II sarcoidosis. It also confirmed the value of TBNA, with excellent diagnostic yields especially in stage I of the disease. PMID- 15281437 TI - Induced sputum as an additional tool in the identification of metal-induced sarcoid-like reaction. AB - BACKGROUND: Aluminium dust exposure produces asthma, chronic bronchitis, pulmonary fibrosis and granulomatous lung diseases. There is an increased risk of mistaken diagnosis of sarcoidosis when other interstitial lung diseases of known origin are occurring. CASE REPORT: We describe a case of a welder working in a stainless steel factory who had been exposed for more than 20 years to a dusty environment containing high levels of hazardous dust. He underwent lung function studies, a beryllium-lymphocyte transformation test (BeLTT), induced sputum (IS) analysis, aluminum-induced blastic proliferation test, and mineralogical and immunologic studies. The lung function tests raised the suspicion of sarcoidosis. T cell subsets recovered from induced sputum disclosed a helper T lymphocyte alveolitis, and transbronchial biopsies showed sarcoid-like epithelioid granulomata. Peripheral blood lymphocytes exhibited blastic transformation in the presence of soluble aluminium compounds. Scanning electron microscope studies from induced sputum-retrieved material showed abundant particles of aluminum. His final diagnosis was sarcoid-like granulomatous-induced aluminium disease. CONCLUSION: We propose an alternative non-invasive approach to identify antigenic metals in occupational exposures. PMID- 15281438 TI - Contemporaneous pulmonary sarcoidosis and chronic myeloid leukemia. PMID- 15281439 TI - Diffuse alveolar hemorrhage due to Legionella pneumonia. PMID- 15281440 TI - Recruitment maneuver and PEEP response in acute exacerbation of nonspecific interstitial pneumonia. PMID- 15281442 TI - [Study on the feasibility of Schistosoma japonicum infection via oral mucosa]. PMID- 15281441 TI - [Screening and expression of recombinant human monoclonal antibody Fab fragments specific to Entamoeba histolytica]. AB - OBJECTIVE: To prepare recombinant human monoclonal antibody Fab fragments specific to the surface antigen of Entamoeba histolytica. METHODS: Total RNA was isolated from lymphocytes which were separated from an asymptomatic E. histolytica cyst carrier. The genes of IgG light chain and Fd region of heavy chain were amplified by a reverse transcriptase PCR and ligated with a plasmid vector. After the genes were introduced into Escherichia coli, the clones expressing Fab fragments specific to the surface antigen of E. histolytica were screened and the product was purified. RESULTS: Thirty thousand clones were screened and one of them was proved positive to the surface antigen of E. histolytica. CONCLUSION: This study demonstrated that the bacterial system can be used to produce recombinant human monoclonal antibody Fab fragments specific to the surface antigen of E. histolytica and they may be applicable for the future diagnosis and treatment of the infection. PMID- 15281443 TI - Effects of tetracycline operator element insertion on Plasmodium falciparum glycophorin binding protein 130 gene promoter activity. AB - OBJECTIVE: To construct tetracycline operator (TetO) modified glycophorin binding protein 130 gene (GBP130) promoter of Plasmodium falciparum and investigate the position effect of insertion of TetO on the promoter activity. METHODS: Cloning of 7-copy of TetO (7cot) sequence into 4 points relative to transcriptional initiation site of GBP130 promoter in pGBPCATdelta2 plasmid (2 upstream and 2 downstream), respectively, produced 4 derivative plasmids, pG/7T (-5), pG/7T ( 2), pG/7T (+2) and pG/7T (+5). After transient transfection, the expression level of reporter gene CAT in both pGBPCATdelta2 and its derivative plasmids was detected and analysed by CAT ELISA. RESULTS: Identification by enzyme restrictions and PCR amplifications, as well as DNA sequencing confirmed that the plasmids were successfully constructed. Transfection of these plasmids and CAT detection suggested that insertion of 7cot into each point of GBP130 promoter enhanced the promoter activity, and downstream location of 7cot in the promoter showed higher promoter activity than those located upstream, with the strongest effect in Ins 4 (point +5). CONCLUSION: Plasmid pG/7T (+5), in which 7cot was inserted into +5 point of GBP130 promoter, can be chosen as the responsive plasmid in establishing tetracycline-controlled transgenic expression system of malarial parasites. PMID- 15281444 TI - [Expression of proteinase cathepsin L1 gene of Schistosoma japonicum in Escherichia coli]. AB - OBJECTIVE: To express the proteinase cathepsin L1 gene of Schistosoma japonicum (SjCL1) in Escherichia coli JM109 cells. METHODS: The SjCL1 gene was amplified from the recombinant plasmid pcDNA3-SjCL1 by PCR. The gene was cloned into a prokaryotic expression vector pGEX4T-1 to construct a recombinant plasmid pGEX SjCL1. The E. coli JM109 cells were transformed with the recombinant plasmid pGEX SjCL1 and the transformants were induced by IPTG to express the recombinant protein, the target protein was then identified by SDS-PAGE and Western blotting. RESULTS: A 1 kb length PCR product was obtained and a recombinant plasmid pGEX SjCL1 was constructed. The expression product was detected by SDS-PAGE and Western blotting and an expression band about 62000 was found. CONCLUSION: The SjCL1 gene is effectively expressed in the E. coli JM109 cells. PMID- 15281445 TI - [Screening of cDNA library of Schistosoma japonicum with sera from rabbits vaccinated with ultraviolet-attenuated schistosomula]. AB - OBJECTIVE: To search new potential schistosomiasis vaccine by screening cDNA library with sera of rabbits vaccinated with attenuated larvae. METHODS: Schistosoma japonicum (Sj) adult worm cDNA library was screened with sera of rabbits vaccinated with ultraviolet-attenuated schistosomula and by sera of infected rabbits, and the inserts of positive clones were amplified and sequenced. RESULTS: Six kinds of Sj genes were obtained after three rounds of screening. Among the genes were glyceraldehyde 3-phosphate dehydrogenase (GAPDH), serine protease inhibitors (serpin), mitochondrion coding region, part of myosin heavy chain gene and two new genes, respectively. CONCLUSION: Screening cDNA library with sera of animals vaccinated with attenuated larvae is an effective way to search new vaccine candidates for schistosomiasis. PMID- 15281446 TI - [Sarcocystis suihominis infection in human and pig population in Guangxi]. PMID- 15281447 TI - [Preparation and identification of monoclonal antibodies against the Region II+ motif in circumsporozoite protein of Plasmodium falciparum]. AB - OBJECTIVE: To develop and identify the monoclonal antibodies (McAbs) against Region II+ motif in circumsporozoite protein of Plasmodium falciparum. METHODS: BALB/c mice were immunized with 12 peptides within Region II+ in circumsporozoite protein of P. falciparum. Spleen cells isolated from the immunized mice were fused with myeloma cell. After three times screening with ELISA, 3 positive hybridoma cell lines were obtained. RESULTS: ELISA test indicated that the McAbs reacted with recombinant circumsporozoite protein fragment containing tandemly repeat region and conserved Region II+. IFA test showed that the McAbs recognized not only the sporozoites of P. falciparum, but also the sporozoifes of P. yoelii. CONCLUSION: McAbs obtained can probe the Region II+ motif in circumsporozoite protein of P. falciparum, which might also recognize that of other Plasmodium species. PMID- 15281449 TI - [Clinical consultation for parasitic diseases inspires the importance of health education]. PMID- 15281450 TI - [RAPD analysis on different isolates of Trichomonas vaginalis]. AB - OBJECTIVE: To study genetic polymorphism of DNA on seven isolates of Trichomonas vaginalis. METHODS: The random amplified polymorphic DNA (RAPD) technique was performed to amplify genomic DNA of the seven T. vaginalis isolates, including Beijing 1, Beijing 2, Chengde, Tangshan, Jiujiang 1, Jiujiang 2 and Jiujiang 3. The DNA bands detected were analyzed by clustering analysis with SPSS software. RESULTS: The percentage of genetic similarity among the seven isolates was from 77.4% to 94.7%, showing a close genetic relationship among them. The percentages between the isolates of Beijing 1 and Tangshan, Jiujiang 1 and Jiujiang 2, Beijing 2 and Jiujiang 3 were 89.2%, 92.1% and 94.7% respectively, while that of Jiujiang 1 and Chengde was 77.4%, indicating a lower homology. CONCLUSION: There are a close genetic relationship and certain gene polymorphism among the seven T. vaginalis isolates; geographical origin plays little role to the genetic characteristics. PMID- 15281448 TI - [Genotype detection of the merozoite surface protein alleles of Plasmodium vivax]. AB - OBJECTIVE: To develop a method for detecting the genotype of Plasmodium vivax merozoite surface protein 1 (PvMSP-1) alleles. METHODS: According to the sequence characteristic of PvMSP-1, nested polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique was used to amplify the polymorphic region of ICB5-ICB6 which contains Q repeats and PvuII restriction site (Sal-1 type). The PCR product was digested by PvuII restriction endonuclease and the digested fragments were observed by 2% agarose gel electrophoresis. The allelic type was determined according to the banding pattern. RESULTS: Bands in size of 400 bp (Belem type) and/or 470 bp (Sal-1 type) appeared in all 98 P. vivax isolates, no band was found in negative control. After PvuII digestion, two Sal-1 type fragments (120 bp and 350 bp) were obtained from 45 samples of 470 bp. Single-band of 400 bp appeared in 3 of 40 samples with 400 bp as Belem type, two bands of 120 bp and 280 bp appeared from other 35 samples as recombination type III, and another 2 bands with 120 bp and 240 bp as Korean isolate. CONCLUSION: The result showed that the nested PCR-RFLP may be applied in the detection and identification of the three PvMSP-1 allelic types in China. PMID- 15281451 TI - [Prevalence of hydatidosis in Heilongjiang Province]. PMID- 15281452 TI - [Soluble expression of Plasmodium falciparum glutamate dehydrogenase in Escherichia coli, and its purification and identification]. AB - OBJECTIVE: To make soluble expression of Plasmodium falciparum (FCC1/HN) glutamate dehydrogenase (GDH) in Escherichia coli, purification and immunocompetence identification of the recombinant non-fusion GDH. METHODS: The GDH gene was cloned into prokaryotic expression vector pET23 (a) to form recombinant expression vector pET23 (a)/GDH. pET23(a)/GDH was transformed into E. coli BL21 (DE3). Induced by IPTG (isopropyl-beta D-thiogalactoside), GDH was highly expressed in the supernatant after sonication. The soluble recombinant GDH was purified by Source-Q and Source-S chromatography. Enzyme-linked immunosorbent assay and Western blotting were carried out to identify the immunocompetence of the purified product. RESULTS: SDS-PAGE analysis showed that the soluble GDH protein accounted for approximately 15% of the total bacterial protein. By two step ion-exchange chromatography, the purity of GDH reached more than 90% and the GDH possessed high antigenicity. CONCLUSION: The soluble expression of GDH results in an integral three-dimensional structure epitope with high biological activity. PMID- 15281453 TI - [Malaria incidence and control in four pilot villages of Xiaochang County in the past nine years]. PMID- 15281454 TI - [Application of EITB in immunodiagnosis of cysticercosis]. AB - OBJECTIVE: To establish a sensitive and specific method EITB for the immunodiagnosis of cysticercosis. METHODS: Two-dimensional electrophoresis was used to analyze and purify the cyst fluid antigen. Western blotting using the sera of patients with cysticercosis, hydatidosis, and other heteroserum was used to screen the specific antigens. EITB using the specific antigen was established and compared with ELISA. The detection effect of serum and blood on filter paper in EITB was also compared. RESULTS: Two specific antigens with isoelectric point (pI) of 9.4 and Mr of 14000 and 16600 were obtained, respectively. EITB method based on these antigens was established with the sensitivity and specificity of 92.5% and 100%, respectively, significantly higher than that of ELISA. The sensitivity of serum and filter paper blood was similar in EITB. CONCLUSION: A sensitive and specific EITB method for immunodiagnosis of cysticercosis was established. PMID- 15281455 TI - [Ultrastructural study on pharyngeal armature of seven species of sandflies in china by scanning electron microscopy]. AB - OBJECTIVE: To observe the ultrastructure of pharyngeal armature of 7 species of sandflies in China. METHODS: The pharyngeal armature of various sandflies were studied by scanning electron microscopy. RESULTS: The pharyngeal armature of sandfly consisted of pointed-teeth with various shape, number and arrangement among different species. CONCLUSION: Such differences may provide the morphological proof for identification of species. PMID- 15281456 TI - [Screening of mimic epitopes of Trichinella spiralis antigen from phage 12-mer peptide library]. AB - OBJECTIVE: To screen special mimic epitopes of Trichinella spiralis antigen from peptide library for exploring new diagnostic antigens. METHODS: Ts-IgG purified from serum of trichinosis patients was used to screen the phage 12-mer peptide library for 5 rounds. 24 clones were picked out randomly to detect the immunoactivity. The sensitivity and specificity of the 6 clones (T1 - T6) whose A values were higher than others were tested by ELISA. RESULTS: The sensitivity of the clones T1 - T6 was the same with larval antigen of Trichinella spiralis (TsA) (positive rate: 100%, P > 0.05), and there was no difference in specificity between T1 - T6 and TsA (negative rate: 0 - 40%, P > 0.05); T3 and T6 did not react with sera from patients of paragonimiasis, showing higher specificity than TsA (P < 0.05); T6 did not react with sera from patients of schistosomiasis, also showing higher specificity than TsA (P < 0.05). CONCLUSION: The mimic antigenic epitopes of Trichinella spiralis have been successfully obtained by screening phage 12-mer peptide library. PMID- 15281457 TI - [Detection of biliverdin reductase activity of Schistosoma japonicum]. AB - OBJECTIVE: To explore the presence of biliverdin reductase (BR) activity in adult worms of Schistosoma japonicum in vitro. METHODS: The soluble fraction was isolated from homogenates of adult worms of S. japonicum, and incubated with biliverdin under different pH conditions and buffers. The time dependency of this enzymatic reaction was also detected. RESULTS: The soluble fraction of the homogenates of adult worms could degrade biliverdin in vitro, the BR activity was 43.30 nmol/(mg x min), with its optimal condition at pH 8.7. The rate of reaction peaked at 15 min of incubation for the BR activity. CONCLUSION: The presence of BR activity in adult worms of S. japonicum was firstly demonstrated. PMID- 15281458 TI - [Studies on Paragonimus proliferus]. AB - OBJECTIVE: To make identification between Paragonimus menglaensis and Paragonimus proliferus. METHODS: Crabs were collected from same area where P. proliferus and P. menglaensi were reported, metacercariae and excysted metacercariae were separated. Adult worms were collected from experimental infection and identified. RESULTS: The metacercaria is large, with an average size of (1.23 +/- 0.087) mm x (1.10 +/- 0.073) mm, covered with a thin and fragile cyst wall; the size of excysted metacercariae is (2.01 +/- 0.71) mm x (0.62 +/- 0.12) mm, with irregular bough-like wrinkles excretory bladder resembling in front of ventral sucker, two pointed and slim distal ends of gut locate at 1/6 of the body from the tail end; the adult worm has large uterine mass, with an average length of 1/4.2 of the whole body. The natural definitive host for P. proliferus is not monkeys, dogs, and cats, but rats. The metacercaria of the reported P. menglaensis has been mixed up with that of P. microrchis from the same crab, excysted metacercaria has been same to that of P. proliferus, and an immature worm has been mistakenly identified as its adult worm. CONCLUSION: P. proliferu is a valid independent species, while P. menglaensis is a mis-identified, invalid one. PMID- 15281459 TI - Classification of the sand-mite family Walchiidae (Acariformes: Trombiculoidea). AB - The priority of Walchiinae Ewing, 1946 based on Walchia Ewing, 1931 or Gahrliepiinae Womersley, 1952 based on Gahrliepia Oudemans, 1912 as Walchia regarded as subgeneric taxon had been a controversy for almost half a century since 1952. Both genera are valid now. Wen (1999) redefined both subfamilial characters and in turn promoted Walchiinae to a full familial status. Walchiidae (Ewing, 1946) Wen, 1999 is characterized by SIF=4B/4Bs/5B/6B-N/B-3/2 2(1)1(0)1(0)0.0000, small to large sized sand-mites, IP=320 - 1220. Scutum is small to large size, extending backward over part of dorsum, and pentagonal with acuminate posterior angle or tongue-shaped. The scutum is never provided with anteromedian setae (AM or vi) and anteromedian projection (A or N=0). Scutal setae have AL and PL pairs basically, frequently in addition with 2 -40 accessories (PPLs), and rarely 1 - 2 pairs of intermedial setae (IM). Sensillae (Sn or sci) are short and expanded. Leg segments are 7.6.6 always without variations. Casting off the anteromedian setae on scutum, increasing the leg segments and reducing the tactile body setae are the plesiomorphic characters of sand-mites, that means Walchiidae in higher advance of evolution than both Trombiculidae (Ewing, 1929) and Leeuwenhoekiidae (Womersley, 1944). It is rationally to unify three families of vertebrate parasitic larvae into a single superfamily, Trombiculoidea nec Welbourn (1991), that separable from superfamily Trombidioidea of arthropod parasitic larvae and standing at most advanced evolution of Parasitengona. Family Walchiidae has 2 subfamilies, Walchiinae Ewing, 1946 sensu Wen 1999 and Gahrliepiinae Womersley, 1952, sensu Wen 1999. Each subfamily contains two tribes, Walchiini (Ewing, 1946) Wen 1984 and Schoengastiellini Wen 1984 for Walchiinae, and Gahrliepiini sensu Wen 1984 and Intermedialiini Wen, 1984 for Gahrliepiinae. Currently this family has 18 genera and 28 subgenera, 248 nominated species and subspecies. Walchiid sand-mites are essentially an Old World family and best developed in the Oriental Region with the center of development in Southeast Asia. PMID- 15281460 TI - [Remote sensing and geographic information system techniques in malaria research]. PMID- 15281461 TI - [Immunostimulatory sequence and DNA vaccine]. PMID- 15281462 TI - [Clinicopathologic analysis of 46 cases of appendiceal schistosomiasis]. PMID- 15281463 TI - [A comparative investigation on chemotherapy in the control of clonorchiasis]. PMID- 15281464 TI - Johnson grass, Sorghum halepense. PMID- 15281465 TI - What can we know about asthma by using administrative databases? PMID- 15281466 TI - Efficacy and safety of sublingual immunotherapy. AB - OBJECTIVE: To review the available published data concerning the use of sublingual immunotherapy (SLIT) in respiratory allergy to primarily evaluate the clinical efficacy and safety of the treatment and to secondarily consider the mechanisms of action and any unresolved questions. DATA SOURCES: Articles in the medical literature (starting from 1986 up to November 2003) derived from searching the MEDLINE database with the keywords sublingual immunotherapy, respiratory allergy, asthma, and rhinitis. Sources included review articles, randomized controlled clinical trials, postmarketing surveillance studies, and relevant reports from meeting proceedings. STUDY SELECTION: Articles concerning safety, efficacy, and mechanisms of SLIT published in English-language, peer reviewed journals. RESULTS: SLIT proved effective and safe in adults and children. As with traditional subcutaneous immunotherapy, SLIT has long-lasting efficacy and a preventive effect on new sensitizations. CONCLUSION: SLIT is a viable alternative to subcutaneous immunotherapy. Its use in pediatric patients seems to be particularly promising. PMID- 15281467 TI - Biological control of fire ants: an update on new techniques. AB - OBJECTIVE: To review the present understanding of biological control methods for imported fire ants (IFAs). DATA SOURCES: We searched MEDLINE, Biological Abstracts, and the US Department of Agriculture Formis Ant Literature database. STUDY SELECTION: All articles published in the last 10 years on biological control of fire ants were selected. RESULTS: The decapitating flies Pseudacteon tricuspis, Pseudacteon curvatus, and Pseudacteon litoralis have been successfully released in the United States. The continued releases of multiple species of decapitating flies will expand the area of impact, applying greater pressure on IFA populations throughout the southern United States. The microsporidium Thelohania solenopsae causes the slow demise of a fire ant colony. The advantages of T. solenopsae as a biological control agent include debilitation of queens, specificity for IFAs, self-sustaining infections, and lower relative tolerance to chemical pesticides. Solenopsis daguerrei has also been shown to have detrimental effects on IFA colony growth, the number of sexual reproductives produced, and the number of host queens in multiple queen colonies; however, this parasite is difficult to rear in the laboratory and to introduce into IFA colonies. CONCLUSIONS: It is unlikely that IFAs can be completely eradicated from the United States. However, technology using chemicals and/or natural control agents could eventually maintain populations at low levels if an integrated approach is used for control. PMID- 15281468 TI - Unusually persistent rhinorrhea in a patient with allergic rhinitis. PMID- 15281469 TI - Rates and characteristics of intensive care unit admissions and intubations among asthma-related hospitalizations. AB - BACKGROUND: A life-threatening attack of asthma that leads to intensive care unit (ICU) admission, intubation, or both identifies patients at high risk of subsequent morbidity and mortality and represents a major cost burden. OBJECTIVE: To assess the rates, characteristics, and costs of ICU admissions and intubations among asthma-related hospitalizations. METHODS: This analysis was performed using a database of 215 hospitals representing more than 3 million annual inpatient visits. Asthma-related hospital admissions were identified by a primary diagnosis code for asthma during 2000. Logistic regression was used to estimate the odds ratios (ORs) for predictors of ICU admission, intubation, and in-hospital mortality. Ordinary least squares regression was used to estimate adjusted mean costs and length of stay. RESULTS: Of 29,430 admissions with a primary diagnosis of asthma, 10.1% were admitted to the ICU and 2.1% were intubated. The risk of in hospital death was significantly greater in patients who were intubated but not admitted to the ICU (OR, 96.20; 95% confidence interval [CI], 50.24-184.20), those who were admitted to the ICU and intubated (OR, 62.69; 95% CI, 38.17 102.96), and patients with more severe comorbidities (OR, 1.53; 95% CI, 1.38 1.70). On average, intubated patients stayed in the hospital 4.5 days longer and incurred more than $11,000 in additional costs; patients admitted to the ICU stayed 1 day longer and accounted for $3,000 in additional costs vs standard admissions. CONCLUSIONS: The inpatient mortality, morbidity, and cost burden of life-threatening asthma in the United States is considerable. This study characterizes patients with asthma at risk of ICU admissions and intubations. Appropriate recognition and treatment are needed to prevent these severe and potentially life-threatening events. PMID- 15281470 TI - Development and validation of school-based asthma and allergy screening questionnaires in a 4-city study. AB - BACKGROUND: Asthma and allergies are commonly undiagnosed in children. Schools provide settings for potentially accessing almost all children for asthma and allergy screening. OBJECTIVE: To evaluate the feasibility and validity of using a questionnaire-based screening tool to identify undiagnosed asthma and respiratory allergies in children in kindergarten to grade 6. METHODS: A student questionnaire (SQ) and a parent questionnaire (PQ) were developed, administered in 4 diverse communities, and validated against standardized clinical assessments. Children without diagnosed asthma and representing a range of symptoms participated in a validation study that consisted of independent, standardized, clinical assessments. Sensitivity, specificity, and predictive values for questionnaire items were evaluated against expert consensus designations. RESULTS: A total of 190 children (age range, 7-13 years) completed the validation study. Affirmative responses to individual questions from either the SQ or PQ regarding asthma and allergy were modestly to moderately predictive of the clinical assessments (odds ratios, generally 2.5-5.0). When considering a positive asthma screen as affirmative responses to 3 of the best 7 SQ asthma questions, the odds ratio for asthma was 9.3 (95% confidence interval, 4.1-21.1), with 80% sensitivity and 70% specificity. Considering the allergy screen as positive based on affirmative response to either of the 2 SQ allergy questions yielded 81% sensitivity and 42% specificity. CONCLUSIONS: Either a 9-item SQ or a 10-item PQ can be used in diverse settings to screen for asthma and respiratory allergies. The SQ, obtained by directly screening students, may provide a sensitive approach for detecting children with previously undiagnosed asthma and allergies. PMID- 15281471 TI - Efficacy and safety of mometasone furoate dry powder inhaler vs fluticasone propionate metered-dose inhaler in asthma subjects previously using fluticasone propionate. AB - OBJECTIVE: To compare the efficacy and safety of mometasone furoate dry powder inhaler (DPI) administered once daily in the evening with fluticasone propionate metered-dose inhaler (MDI) administered twice daily. METHODS: An 8-week, randomized, open-label, parallel-group study compared mometasone furoate DPI, 400 microg every evening (1 puff daily), with fluticasone propionate MDI, two 125 microg puffs twice daily, in 167 adults and adolescents with moderate persistent asthma previously using fluticasone propionate. The primary efficacy variable was the change in forced expiratory volume in 1 second (FEV1) from baseline to the end point. Variables such as response to therapy and subject satisfaction with the inhaler devices were also analyzed. RESULTS: Improvement in FEV1 was noted at the week 2 visit with both treatments. This improvement was maintained at the 4- and 8-week visits and at the end point for both groups. The mean percent change in FEV1 from baseline to the end point was 4.58% with mometasone furoate DPI and 6.98% with fluticasone propionate MDI (P = .35). At the end point, physicians rated 62% of the mometasone furoate DPI group as "improved" or "much improved" compared with 47% of the fluticasone propionate MDI group (P = .007). A significantly greater proportion of subjects in the mometasone furoate DPI group "liked the inhaler a lot" vs subjects in the fluticasone propionate MDI group (46.8% vs 22.4%; P = .01). Both treatments were well tolerated. CONCLUSION: Mometasone furoate DPI, 400 microg every evening, provided comparable efficacy as fluticasone propionate MDI, two 125-microg puffs twice daily, in subjects with moderate persistent asthma previously treated with fluticasone propionate. PMID- 15281472 TI - Effects of butterbur treatment in intermittent allergic rhinitis: a placebo controlled evaluation. AB - BACKGROUND: Butterbur (Petasites hybridus) contains the active ingredient petasin, which exhibits antileukotriene and antihistamine activity. Previous studies of intermittent allergic rhinitis (IAR) have demonstrated a comparable response to butterbur compared with a histamine H1-receptor antagonist on the 36 Item Short-Form Health Survey quality-of-life score. However, there has been no placebo-controlled study of the effects of butterbur use on objective and subjective outcomes in IAR. OBJECTIVE: To evaluate the effects of treatment with butterbur vs placebo on objective and subjective outcomes in IAR. METHODS: A double-blind, placebo-controlled, crossover study was carried out during the grass pollen season in Tayside, Scotland. Thirty-five patients (14 men and 21 women) with IAR received butterbur, 50 mg twice daily, or placebo for 2 weeks. Domiciliary measurements were taken in the morning and evening for peak nasal inspiratory flow (PNIF) (the primary outcome variable), nasal and eye symptoms, and rhinoconjunctivitis-specific quality-of-life score. RESULTS: Butterbur treatment had no significant effect on PNIF, total nasal symptom score, eye symptom score, or quality of life compared with placebo use. Mean (SEM) morning and evening PNIF values were 107 (6) and 114 (6) L/min, respectively, for butterbur vs 105 (6) and 117 (6) L/min for placebo. Mean (SEM) morning and evening total nasal symptom scores (maximum total score, 12) were 3.4 (0.4) and 3.5 (0.4), respectively, for butterbur vs 3.7 (0.3) and 3.8 (0.4) for placebo. CONCLUSIONS: There was no significant clinical efficacy of butterbur use vs placebo use on objective and subjective outcomes in IAR. Further studies are now indicated to investigate the use of butterbur in persistent allergic rhinitis. PMID- 15281473 TI - Effect of budesonide aqueous nasal spray on hypothalamic-pituitary-adrenal axis function in children with allergic rhinitis. AB - BACKGROUND: Intranasal corticosteroids are safe and effective for treating allergic rhinitis in adults. Since children may receive more systemic corticosteroid on a dose-per-weight basis than adults, the safety of corticosteroid therapy in pediatric patients is an important issue. OBJECTIVE: To determine the effects of treatment with budesonide aqueous nasal spray using the recommended once-daily dose for adults and children 6 years and older on hypothalamic-pituitary-adrenal (HPA) axis function in pediatric patients with allergic rhinitis. METHODS: In a 6-week, multicenter, double-blind, placebo controlled study, 78 patients aged 2 to 5 years with allergic rhinitis were treated with budesonide aqueous nasal spray (64 microg/d) or placebo. Mean change in morning plasma cortisol levels from baseline to study end 0, 30, and 60 minutes after low-dose (10-microg) cosyntropin stimulation and mean change in the difference from 0 to 30 minutes and from 0 to 60 minutes after cosyntropin stimulation were used to evaluate HPA axis function. RESULTS: Mean change from baseline to study end in plasma cortisol levels 0, 30, and 60 minutes after cosyntropin stimulation and the difference from 0 to 30 minutes and from 0 to 60 minutes were not significantly different between the treatment and placebo groups (P > .05 for all). At the end of the study, 3 budesonide aqueous nasal spray and 6 placebo patients were classified as having subnormal HPA axis function. The safety and tolerability profile of budesonide aqueous nasal spray was comparable to that of placebo. CONCLUSIONS: Administration of budesonide aqueous nasal spray for 6 weeks was well tolerated and safe and had no measurable suppressive effects on HPA axis function in patients aged 2 to 5 years with allergic rhinitis. PMID- 15281474 TI - Expression of interleukin 4 receptors in bronchial asthma patients who underwent specific immunotherapy. AB - BACKGROUND: Interleukin (IL) 4 and IL-13 are crucial cytokines for the development of allergic reactions and have been shown to modulate the function of monocytes and macrophages. OBJECTIVES: To evaluate the expression of IL-4Rs on peripheral blood monocytes and in the serum of patients with bronchial asthma who underwent specific immunotherapy (SIT). METHODS: The study was performed on 17 asthma patients with a typical clinical history and positive skin prick test results to Dermatophagoides pteronyssinus allergens. Five asthma patients who declined SIT were used as a comparator control group. Ten healthy persons served as negative controls. Flow cytometry analysis was performed on the whole blood samples using labeled monoclonal antibodies against CD14 and CD36 monocyte markers and against the CD124 alpha chain of IL-4R. The serum levels of soluble IL-4R were evaluated using an immunoenzymatic assay. RESULTS: Compared with controls, bronchial asthma patients before SIT had a higher mean +/- SD percentage of CD14-positive cells that coexpressed CD124 (3.5% +/- 1.8% vs 18.6% +/- 7.9%; P < .01). After SIT, the mean +/- SD percentage of CD14 cells coexpressing CD124 decreased to 8.1% +/- 5.1%, which was significantly lower than before SIT (P < .01) but still significantly higher than in controls (P = .01). Changes in CD124 expression were associated with up-regulation of CD14 and down regulation of CD36 expression on peripheral blood monocytes, suggesting that IL 4/IL-13-mediated signaling may be important for regulation of monocyte phenotype and function in asthma patients receiving SIT. CONCLUSIONS: Even short courses of SIT are associated with a decrease in IL-4R expression on peripheral blood monocytes, which may cause decreased IL-4/IL-13-mediated effects in patients who undergo SIT. PMID- 15281475 TI - Respiratory symptoms in greenlanders living in Greenland and Denmark: a population-based study. AB - BACKGROUND: Knowledge of respiratory diseases in an arctic population with increasingly westernized lifestyles provides the opportunity to obtain new information in this field. OBJECTIVE: To investigate the influence of environment and lifestyle on the presence of respiratory symptoms in a genetically homogenous population sample living under widely differing conditions. METHODS: Greenland is a part of Denmark, but its climate is mainly arctic, as opposed to the temperate climate of southern Denmark. A random sample of Inuits who had immigrated to Denmark and Inuits from 3 towns and 4 remote settlements in Greenland were studied. Of the 6,695 invited Inuits, 4,162 (62%) completed a questionnaire concerning respiratory symptoms and risk factors. RESULTS: Of the 4,162 Inuits, 847 (20%) had respiratory symptoms. Bronchitis was more frequent in the areas of Greenland than in Denmark (26% and 20% vs 13%; P < .001), whereas the pattern of asthma was contradictory (6% and 9% vs 10%; P = .057). Bronchitis was associated with living area (P = .01), tobacco consumption (P < .001), and asthma (P = .001), whereas asthma was related to living area (P = .03), hay fever (P < .001), low intake of whale (P = .04), years in Denmark (P = .09), and bronchitis (P < .001). CONCLUSIONS: Inuits' prevalence of bronchitis and asthma differed, with a higher frequency of bronchitis and a lower frequency of asthma in Inuits living in Greenland compared with Denmark. Living conditions or areas, diet, tobacco use, climate, and atopy are important for the presence of symptoms. PMID- 15281476 TI - Safety profile of budesonide inhalation suspension in the pediatric population: worldwide experience. AB - OBJECTIVE: To review the worldwide safety data for budesonide inhalation suspension (Pulmicort Respules) to provide a budesonide inhalation suspension pediatric tolerability profile. DATA SOURCES: Clinical study data were obtained from AstraZeneca safety databases used by the US Food and Drug Administration to support the approval of budesonide inhalation suspension and from postmarketing surveillance reports (January 1, 1990, through June 30, 2002). STUDY SELECTION: Completed parallel-group studies of patients with asthma 18 years and younger. RESULTS: Safety data for budesonide inhalation suspension were pooled from 3 US, 12-week, randomized, double-blind, placebo-controlled studies (n = 1,018); data from their open-label extensions (n = 670) were pooled with data from a fourth US open-label study (n = 335). Data for 333 patients 18 years and younger enrolled in 5 non-US studies also were analyzed. No posterior subcapsular cataracts were reported in any study, and the frequencies of oropharyngeal events and infection with budesonide inhalation suspension were comparable with those of reference treatments. No increased risk of varicella or upper respiratory tract infection was apparent, and budesonide inhalation suspension did not cause significant adrenal suppression in studies assessing this variable. There were small differences in short-term growth velocity between children who received budesonide inhalation suspension and those who received reference treatment in 2 of 5 trials that evaluated this variable. No increased risk of adverse events was apparent from postmarketing reports. CONCLUSIONS: Short- and long-term treatment with budesonide inhalation suspension, using a wide range of doses, is safe and well tolerated in children with asthma. PMID- 15281477 TI - Glutathione and nitrite in induced sputum from patients with stable and acute asthma compared with controls. AB - BACKGROUND: Oxidative stress is believed to play an important role in the pathogenesis of asthma. Determining the reduced glutathione (GSH) and nitric oxide (NO) contents of the airway is useful when investigating oxidative stress in the lung. OBJECTIVE: To explore antioxidant defenses by measuring sputum GSH levels and to evaluate oxidant stress by measuring sputum nitrite (NO2-) levels in asthma patients. METHODS: Sputum GSH, NO2-, cell counts, and plasma NO2- contents were evaluated in 11 patients with stable asthma, 10 patients with acute asthma attacks, and 11 controls. RESULTS: The highest GSH content in sputum samples was in stable asthma patients compared with the other groups (P < .001), and patients with exacerbations of asthma had a greater GSH content than controls (P < .001). Mean sputum NO2- content was significantly lower in controls than in acute (P = .001) and stable (P < .001) asthma patients. There was no significant difference in sputum NO2- contents between acute and stable asthma patients, although there was a trend toward higher levels in acute asthma patients (P = .38). CONCLUSIONS: Sputum induction can be used to obtain bronchial secretions for the evaluation of GSH and NO2- contents. Oxidative stress is chronic and probably less severe in patients with stable asthma. Glutathione and NO2- may serve as markers for determining the extent of the oxidative processes in asthma, which is characterized by chronic airway inflammation. PMID- 15281478 TI - Systemic urticaria in an infant after ingestion of processed food that contained a trace quantity of wheat. AB - BACKGROUND: Wheat is a food allergen that occasionally causes a systemic allergic reaction; however, little is known about the quantities of wheat allergen required to evoke allergic symptoms. OBJECTIVE: To report the case of a wheat allergic boy who experienced systemic urticaria and angioedema within 40 minutes after the ingestion of 9 g of packed rice crackers. METHODS: A skin prick test and IgE immunoblotting with wheat proteins were performed. Contamination of wheat protein in the offending rice cracker and other processed rice crackers from local food retail outlets, with labels that did not mention wheat, was examined with enzyme-linked immunosorbent assaying. RESULTS: A skin reaction to wheat was positive. IgE-bound bands were observed with water- and salt-soluble wheat protein and ethanol-soluble wheat gliadin in immunoblotting. A trace quantity of wheat protein, 1.50 microg/g, was determined in the offending rice cracker. In addition, 3 of 8 other kinds of processed rice crackers were contaminated by wheat protein, with levels ranging from 0.26 to 1.13 microg/g. CONCLUSIONS: Approximately 13.5 microg of wheat protein can elicit a systemic adverse reaction in highly sensitive, wheat allergic individuals. The present study confirms the need for control of contamination of food by nondeclared proteins. PMID- 15281479 TI - A successful challenge in a patient with vancomycin-induced linear IgA dermatosis. AB - BACKGROUND: Linear IgA bullous dermatosis (LABD), a subepidermal, blistering skin disease, is generally believed to be idiopathic. It has been reported in association with multiple medications, including vancomycin. In each case, complete clearance of the skin lesions occurred with discontinued use of the drug. A subsequent rechallenge reproduced the eruption within hours to days. OBJECTIVE: To present a patient with vancomycin-associated LABD who underwent a successful challenge with the antibiotic 4 years after the initial reaction. METHODS: The patient developed blistering lesions over her trunk and extremities 10 days after the initiation of vancomycin for sepsis. A biopsy specimen of a skin eruption was consistent with linear IgA dermatosis. Following discontinued use of the drug, her skin lesions resolved. Four years later, she required vancomycin for osteomyelitis. RESULTS: The patient underwent a vancomycin-graded challenge of 5 doses over 5 days. On day 1, she received 10 mg, and this was increased in a semilog fashion to 1,000 mg on day 5. She had no recurrence of her skin lesions. CONCLUSIONS: This is the first case, to our knowledge, to show a successful rechallenge in a patient with drug-associated LABD. Since the patient did not have a reaction to the challenge, it is possible that the IgA antibodies responsible for drug-induced LABD are only present transiently and diminish over time. PMID- 15281480 TI - Nasal saline: placebo or drug? PMID- 15281481 TI - Federal tax-exemption requirements for joint ventures between nonprofit hospital providers and for-profit entities: form over substance? AB - This article discusses the IRS rule on hospital joint ventures and related legal developments. The central thesis is that the IRS's emphasis on operational control is misplaced from both a legal and a policy perspective, and reflects a decidedly strong preference for the form of a joint venture's governance over the substance of its charitable and community service activities. More specifically, the article challenges the IRS position that the rule is a corollary of existing tax law principles. Additionally, social science research is presented to demonstrate that the rule is not likely to promote, and may in fact undermine, United States health policy objectives. PMID- 15281482 TI - A Dutch perspective: the limits of lawful euthanasia. AB - Dutch author Ubaldus de Vries reviews the current state of the euthanasia law in the Netherlands. The legislation, enacted in 2001, creates a medical exception that allows for euthanasia in cases where patients experience "hopeless and unbearable suffering." A brief history of the Dutch approach to euthanasia is set forth, case law is reviewed, and the unique role of the doctor is examined in seeking to understand the extent of one's right to euthanasia in the Netherlands. Because the courts must determine what constitutes "hopeless and unbearable suffering," Professor de Vries analyzes the judicial interpretation of "suffering" and concludes that judicial interpretation has reached its limits, and thus by implication, the limits of lawful euthanasia have been reached. PMID- 15281483 TI - Breaking through the silence: illegality of performing resuscitation procedures on the "newly-dead". AB - Israeli author Daniel Sperling brings to a light a disturbing practice that is taking place in some teaching hospitals throughout the world--the practice of resuscitation procedures on newly dead patients without the consent of the next of-kin. Mr. Sperling examines some of the policies and procedures in place to prevent such practice and also looks at the ethical principles that should guide such procedures. The paper also reviews the general issue of consent in the context of medical decision-making and discusses potential legal claims that might be available to persons who have not been consulted or informed before such procedures are performed. The evolving jurisprudence surrounding the treatment of the newly dead is analyzed and Mr. Sperling concludes by suggesting ways to improve upon the procedures currently in place at some teaching facilities. PMID- 15281484 TI - Will the Supreme Court finally eliminate ERISA preemption? AB - David Trueman's article reviews the history of ERISA preemption by analyzing seminal Supreme Court cases and predicts the future of ERISA preemption in his analysis of recent federal case law. Traditionally, the ability to hold a managed care entity responsible for its actions has been hampered by a strict interpretation of the preemption clauses of ERISA but as the Supreme Court's jurisprudence has evolved and loosened, several federal courts have allowed suits against managed care companies to go forward. Conflict among the federal circuits has arisen and the Supreme Court has granted certiorari to two cases from Texas in order to clarify ERISA preemption. Mr. Trueman discusses the future of ERISA preemption in light of these decisions. PMID- 15281485 TI - Is there an acceptable answer to rising medical malpractice premiums? AB - This article explores the key issues involved in the attempts at reform of the present medical malpractice system. Investigating the effects that federal tort reform legislation would have on physicians, patients, lawyers, and the medical malpractice insurers, Dr. Gunnar succinctly outlines the issues surrounding the present "crisis in healthcare" and explores the separate interests involved. The article examines the economic forces influencing the medical malpractice insurance industry, reviews previous tort reform, and predicts the future of federal tort reform legislation. Dr. Gunnar concludes by proposing alternatives for malpractice reform. PMID- 15281486 TI - The current medical liability insurance crisis: an overview of the problem, its catalysts and solutions. PMID- 15281487 TI - Medical societies' self-policing of unprofessional expert testimony. PMID- 15281488 TI - An institutional perspective on the medical malpractice crisis. PMID- 15281489 TI - A patient perspective: focusing on compensating harm. PMID- 15281490 TI - An insurance perspective on the medical malpractice crisis. PMID- 15281491 TI - A physician's perspective on the medical malpractice crisis. PMID- 15281492 TI - The influence of allogeneic red blood cell transfusion compared with 100% oxygen ventilation on systemic oxygen transport and skeletal muscle oxygen tension after cardiac surgery. AB - In this study we investigated the effects of allogeneic red blood cell (RBC) transfusion on tissue oxygenation compared with those of 100% oxygen ventilation by using systemic oxygen transport variables and skeletal muscle oxygen tension (PtiO2). Fifty-one volume-resuscitated, mechanically ventilated patients with a nadir hemoglobin concentration in the range from 7.5 to 8.5 g/dL after elective coronary artery bypass grafting were allocated randomly to receive 1 unit (transfusion 1; n = 17) or 2 units (transfusion 2; n = 17) of allogeneic RBCs and ventilation with 40% oxygen or pure oxygen ventilation (100% oxygen; n = 17) and no allogeneic blood for 3 hours. Invasive arterial and pulmonary artery pressures and calculations of oxygen delivery (oxygen delivery index) and consumption indices (oxygen consumption index) were documented at 30-min intervals. PtiO2 was measured continuously by using implantable polarographic microprobes. Systemic oxygen transport variables and PtiO2 were similar between groups at baseline. The oxygen delivery index increased significantly with transfusion of allogeneic RBCs and 100% oxygen ventilation, whereas the oxygen consumption index remained unchanged. Oxygen 100% ventilation increased PtiO2 significantly (from 24.0 +/- 5.1 mm Hg to 34.2 +/- 6.2 mm Hg), whereas no change was found after transfusion of allogeneic RBCs. Peak PtiO2 values were 25.2 +/- 5.2 mm Hg and 26.3 +/- 6.5 mm Hg in the transfusion 1 and 2 groups, respectively. Transfusion of stored allogeneic RBCs was effective only in improving systemic oxygen delivery index, whereas 100% oxygen ventilation improved systemic oxygen transport and PtiO2. This improved oxygenation status was most likely due to an increase in convective oxygen transport with a large driving gradient for diffusion of plasma-dissolved oxygen into the tissue. PMID- 15281493 TI - Utility of intraoperative transesophageal echocardiography for diagnosis of pulmonary embolism. AB - Pulmonary embolism (PE) is associated with significant perioperative morbidity and mortality. Transesophageal echocardiography (TEE) may permit direct visualization of PE or secondary signs of pulmonary artery (PA) obstruction. However, its utility in diagnosing PE in the intraoperative setting has yet to be defined. Therefore, we performed intraoperative TEE examinations in 46 patients immediately before pulmonary embolectomy. TEE examinations were reviewed for signs of thromboemboli within the right, left, and main PA, and secondary signs of acute PA obstruction (right ventricular dysfunction, moderate-to-severe tricuspid regurgitation, leftward bowing of the interatrial septum). The definitive location of thromboemboli was determined from the surgical record. Echocardiographic evidence for the presence of PE was correctly demonstrated in 46% of all patients (n = 21 of 46). However, the sensitivity for direct visualization of thromboemboli at any specific location was only 26%. TEE was least sensitive for thromboemboli in the left PA (17%). TEE evidence of right ventricular dysfunction was observed in 96%, tricuspid regurgitation in 50%, and leftward interatrial septal bowing in 98% of examinations. Therefore, the use of intraoperative TEE to diagnose acute PE via direct visualization is limited. Indirect TEE evidence of PA obstruction may be helpful in supporting a diagnosis of PE. PMID- 15281494 TI - Platelet activity in washed platelet concentrates. AB - Life-threatening anaphylaxis or febrile nonhemolytic transfusion reactions after transfusion of platelet concentrates (PCs) is a serious clinical problem caused by the sensitizing of recipients to plasma components, such as immunoglobulin A, or by cytokines. There is a possible indication for washing of PCs in these thrombocytopenic patients. However, only platelets that show activation after physiological stimulation are useful. We determined the spontaneous and induced activation of platelets before and after washing. We investigated 11 consecutive single-donor-apheresis PCs. After production and leukocyte-depletion the PCs were washed in 15%, acid-citrate-dextrose-solution. The spontaneous and the adenosine diphosphate (ADP)-induced, as well as collagen-induced activation, were determined by flow cytometry. Additionally, ADP- and collagen-induced aggregation were measured. Unwashed platelets (16.1%) were activated spontaneously. The washing of PCs led to a threefold increase of spontaneous activation of platelets (47.4%). Because of increased spontaneous activation after washing we could demonstrate cytometrically a loss of induced activation of washed platelets. Furthermore, washing resulted in an impaired ADP-induced aggregability of platelets. These results have led us to reduce the frequency of washing of PCs in our institution, where the only current indication for washing of PCs is in patients with a history of severe nonhemolytic transfusion reactions. PMID- 15281495 TI - Peroxynitrite decreases hemostasis in human plasma in vitro. AB - Coagulopathy has been associated with clinical scenarios that involve reactive nitrogen species such as peroxynitrite (OONO-). Further, OONO- decreases tissue factor and fibrinogen function in vitro. Thus, we hypothesized that exposure of plasma to the OONO- generated with 3-morpholinosydnonimine (SIN-1), a molecule that produces both nitric oxide and superoxide, would result in a decrease in hemostatic function via diminished coagulation protein activity. Hemostatic function of plasma exposed to SIN-1 (0, 1, 5, and 10 mM for 60 min at 37 degrees C) was assessed with thrombelastography, activated partial thromboplastin time, and prothrombin time in the presence or absence of superoxide dismutase (SOD) or an OONO- scavenger. SIN-1 exposure resulted in a significant (P < 0.05), dose dependent decrease in plasma hemostatic function and concurrent significant (P < 0.05) decreases in activities of factor VII, factor VIII complex, and factor X. Fibrinogen concentration was not affected by SIN-1. Antithrombin and protein C activity also decreased significantly (P < 0.05). Coincubation with SOD or an OONO- scavenger significantly (P < 0.05) attenuated SIN-1 mediated changes in hemostasis and procoagulant/ anticoagulant activity. We conclude that OONO- may decrease hemostatic function in human plasma by nitration of key procoagulants and that OONO- may play a significant role in hemorrhagic states. PMID- 15281496 TI - Recombinant factor VIIa for life-threatening bleeding in high-risk cardiac surgery despite full-dose aprotinin. AB - We report the case of an orthotopic heart transplant in a patient with multiple previous cardiac surgeries. The case was prolonged and complicated by severe coagulopathy and bleeding despite the use of full-dose aprotinin throughout. Bleeding was not controlled after 30 U of platelets, 20 U of fresh frozen plasma, and 10 U of cryoprecipitate. However, after the administration of recombinant factor VIIa 90 microg/kg, the rate of bleeding slowed dramatically and no further factor replacement was required. There was no evidence of unwanted clot formation within the newly transplanted heart or around the intraaortic balloon pump that remained in situ for 72 h postoperatively. With the combined risks of coagulopathy and bleeding as well as acute right ventricular failure with increases in pulmonary vascular resistance, the re-do sternotomy for heart transplant seems to be an ideal situation in which to consider the use of recombinant factor VIIa. PMID- 15281497 TI - Life-threatening mediastinal hematoma caused by extravascular infusion through a triple-lumen central venous catheter. AB - We report a case of life-threatening mediastinal hematoma in a 6-mo-old girl during surgical correction of scaphocephaly. The hematoma was caused by extravascular infusion via the proximal lumen of a dislocated triple-lumen central venous catheter (CVC). Worsening symptoms of hypovolemia and ventilation problems prompted performance of transesophageal echocardiography, which reliably and quickly allowed us to exclude pericardial tamponade and detect a mediastinal hematoma. The anesthesiologist should be alert when a patient with a CVC develops acute cardiopulmonary or respiratory symptoms. Repeated aspirations of blood, especially after major positional changes and before giving large quantities of fluid or blood, should be performed to detect secondary malposition of the CVC. PMID- 15281498 TI - Vasopressin during spinal anesthesia in a patient with primary pulmonary hypertension treated with intravenous epoprostenol. AB - Primary pulmonary hypertension (PPH) is a progressive disease with frequent morbidity and mortality, including the risk of cardiac decompensation and death, during general anesthesia. Administration of IV epoprostenol (Flolan) improves symptoms and survival of patients with PPH and thus is an increasingly used long term treatment for this condition. This therapy is associated with impaired platelet aggregation, which may complicate the perioperative management of patients with PPH. We present a case report of a patient with severe PPH receiving a continuous epoprostenol infusion undergoing skin grafting for a leg ulcer under spinal anesthesia. An IV infusion of vasopressin was given to prevent systemic hypotension resulting from sympathetic blockade while avoiding increases in pulmonary vascular resistance that may have resulted from catecholamine usage. PMID- 15281499 TI - An unusual cause of left ventricular outflow tract obstruction after mitral valve repair. AB - Left ventricular outflow tract (LVOT) obstruction caused by systolic anterior motion is a cause of failed mitral valve repair. Intraoperative transesophageal echocardiography has been very helpful in diagnosing problems with mitral valve repairs intraoperatively, allowing immediate correction. We report an unusual cause of LVOT obstruction attributed to prolapse of the annuloplasty ring into the LVOT. Intraoperative hemodynamics were normal, and the diagnosis would not have been made before leaving the operative suite without the transesophageal echocardiography. PMID- 15281500 TI - Late presentation of esophageal injury after transesophageal echocardiography. AB - Esophageal injury is a rare complication of intraoperative transesophageal echocardiography (TEE) associated with cardiac surgery. We report two cases of delayed presentation (2 and 6 days after surgery) of esophageal injury that were likely due to TEE. The differential diagnosis of postoperative pleural effusion or anemia must include esophageal injury from TEE, even 6 days after the procedure. PMID- 15281501 TI - A comparison of epidural bupivacaine, levobupivacaine, and ropivacaine on postoperative analgesia and motor blockade. AB - In this prospective, randomized, observer-blinded clinical trial, we compared the incidence of unwanted lower extremity motor blockade and the analgesic efficacy between small-dose (0.125%; 0.2 mg x kg(-1) x h(-1)) postoperative epidural infusions of bupivacaine (Group B; n = 28), levobupivacaine (Group L; n = 27), and ropivacaine (Group R; n = 26) in children after hypospadias repair. Motor blockade and pain were assessed at predetermined time points during 48 h by using a modified Bromage scale and the Children's and Infant's Postoperative Pain Scale (CHIPPS). Postoperative analgesia was almost identical in all three study groups (CHIPPS range, 0-3), with no need for the administration of supplemental analgesia in any patient. However, significantly more patients in Group B (n = 6; P = 0.03) displayed signs of unwanted motor blockade during the observation period compared with Group L (n = 0) and Group R (n = 0). In conclusion, significantly less unwanted motor blockade was associated with postoperative epidural infusions of 0.125% levobupivacaine or ropivacaine in children after hypospadias repair as compared with a similar infusion of bupivacaine. However, no difference with regard to postoperative analgesia could be detected among the three different local anesthetics studied. PMID- 15281502 TI - A neuronal mechanism of propofol-induced central respiratory depression in newborn rats. AB - The neural mechanisms of propofol-induced central respiratory depression remain poorly understood. In the present study, we studied these mechanisms and the involvement of gamma-aminobutyric acid (GABA)A receptors in propofol-induced central respiratory depression. The brainstem and the cervical spinal cord of 1- to 4-day-old rats were isolated, and preparations were maintained in vitro with oxygenated artificial cerebrospinal fluid. Rhythmic inspiratory burst activity was recorded from the C4 spinal ventral root. The activity of respiratory neurons in the ventrolateral medulla was recorded using a perforated patch-clamp technique. We found that bath-applied propofol decreased C4 inspiratory burst rate, which could be reversed by the administration of a GABAA antagonist, bicuculline. Propofol caused resting membrane potentials to hyperpolarize and suppressed the firing of action potentials in preinspiratory and expiratory neurons. In contrast, propofol had little effect on resting membrane potentials and action potential firing in inspiratory neurons. Our findings suggest that the depressive effects of propofol are, at least in part, mediated by the agonistic action of propofol on GABAA receptors. It is likely that the GABAA receptor mediated hyperpolarization of preinspiratory neurons serves as the neuronal basis of propofol-induced respiratory depression in the newborn rat. PMID- 15281503 TI - Auscultation of bilateral breath sounds does not rule out endobronchial intubation in children. AB - We performed orotracheal intubation in 153 consecutive pediatric patients undergoing cardiac catheterization. Auscultation of bilateral breath sounds was confirmed. By fluoroscopy, the tip of the endotracheal tube (ETT) was seen in the right mainstem bronchus in 18 patients (11.8%) and in a low position, defined as within 1 cm above the carina, in 29 patients (19.0%). All of the 18 patients with right mainstem intubation were children <120 mo of age, and 7 were infants <12 mo of age (Fisher's exact test; P = 0.013). The age, weight, and ETT size for children who had endobronchial and low tracheal positions were significantly (P < 0.001) less than for those who had midtracheal positions. The failure to diagnose mainstem intubation by auscultation alone may be related to the use of the Murphy eye ETT, which reduces the reliability of chest auscultation in detecting endobronchial intubation. Suggested measures for preventing endobronchial intubation include maintaining increased awareness of the imperfection or lack of accuracy of the auscultatory method, assessing insertion depth by checking the length scale on the tube, and minimizing the patient's head and neck movement after intubation. When extreme flexion or extension of the neck is expected after ETT insertion, the resultant change in ETT final position must be anticipated and taken into consideration when deciding on the depth of ETT insertion. This approach resulted in a decrease in improper tube positioning from 20% when the study was initiated to 7.1% in the last 98 patients. PMID- 15281504 TI - Mucositis and airway obstruction in a pediatric patient. AB - Pediatric patients undergoing induction regimens of chemotherapy may require intubation and mechanical ventilation either for respiratory failure or airway compromise as a complication of their therapy. We describe a case of difficult airway management resulting from pseudomembrane formation in a 16-yr-old girl. The patient was undergoing induction chemotherapy for stage IV rhabdomyosarcoma and developed severe mucositis that led to progressive airway obstruction. PMID- 15281505 TI - The development and application of an instrument for assessing resident competence during preanesthesia consultation. AB - In this study, we aimed to construct, validate, and apply an instrument for assessing resident performance at outpatient preanesthesia consultation (PAC). A focus group and a Delphi panel of experts defined component items of a typical outpatient PAC, which could be used as indicators of competence. Items were incorporated in a checklist, which was further validated in a sample of consultations performed by board-certified anesthesiologists. The resulting instrument contained 37 items, grouped into five domains (physician-patient relationship, medical history, physical examination, patient education, and preanesthesia records), with high construct validity, high discriminant validity, moderate internal consistency, and high probability of inter-raters agreement. The instrument was applied to evaluate the performance of seven first-year residents at 317 consecutive PAC. Data were analyzed by constructing exponentially weighted moving average charts for domain and total scores. Statistically significant differing levels of performance could be consistently detected. Applying exponentially weighted moving average charts to the sequential analysis of the developed checklist scores can reliably assess resident performance at the devised criteria. The Preanesthesia Consultation Scoring Checklist is a potentially useful instrument for both formative and summative assessment of residents during their training in processes involved in outpatient preanesthesia evaluation. PMID- 15281506 TI - Nocturnal arterial oxygen desaturation and episodic airway obstruction after ambulatory surgery. AB - Some patients experience disordered breathing during sleep and arterial oxygen desaturation after major inpatient surgery. We performed this study to determine whether similar events occur after ambulatory surgery. Forty-five ambulatory surgery patients received an unrestricted anesthetic. Continuous unattended nocturnal recordings of breathing pattern and oxygen saturation were made in the patients' homes before surgery and during the first and second postoperative nights. Nine patients had a respiratory disturbance index >10 and/or >1% of recording time with oxygen saturation <90% on at least one study night. These nine patients had a significantly older median age and a significantly larger median body mass index. Their median respiratory disturbance index and median percentage of time with oxygen saturation <90% were significantly higher on the first postoperative night than on the preoperative night. PMID- 15281507 TI - A randomized comparison of a multimodal management strategy versus combination antiemetics for the prevention of postoperative nausea and vomiting. AB - A multimodal management strategy for the prevention of postoperative nausea and vomiting (PONV) appears to be superior to single-drug prophylaxis. We tested the hypothesis that a multimodal PONV prophylaxis regimen incorporating total IV anesthesia (TIVA) with propofol and a combination of ondansetron and droperidol is more effective than a combination of these antiemetics in the presence of an inhaled anesthetic. Ninety patients undergoing laparoscopic cholecystectomy were randomized to one of three groups. Group 1 (multimodal group) received TIVA with propofol, droperidol, and ondansetron. Group 2 (combination group) received droperidol and ondansetron with isoflurane and nitrous oxide for the maintenance of anesthesia. Group 3 (TIVA group) received propofol for the induction and maintenance of anesthesia. The complete response rate (no PONV and no rescue antiemetic) at 2 h after surgery was 90%, 63%, and 66% in Groups 1, 2, and 3, respectively (P < 0.05, Group 1 versus Group 2). At 24 h, the complete response rate was 80%, 63%, and 43% in Groups 1, 2, and 3, respectively (P < 0.05, Group 1 versus Group 3). Patient satisfaction was also greater in the multimodal group than in the other two groups in the postanesthesia care unit (P < 0.05). In conclusion, the multimodal management strategy for PONV was associated with a higher complete response rate and greater patient satisfaction when compared with similar antiemetic prophylaxis with inhaled anesthesia or TIVA with propofol. PMID- 15281508 TI - Inhibition of spinal protein kinase C-epsilon or -gamma isozymes does not affect halothane minimum alveolar anesthetic concentration in rats. AB - Anesthetic effects on receptor or ion channel phosphorylation by enzymes such as protein kinase C (PKC) have been postulated to underlie some aspects of anesthesia. In vitro studies show that anesthetic effects on several receptors are mediated by PKC. To test the importance of PKC for the immobility produced by inhaled anesthetics, we measured the effect of intrathecal injections of PKC epsilon and -gamma inhibitors on halothane minimum alveolar anesthetic concentration (MAC) in 7-day-old and 21-day-old Sprague-Dawley rats. The inhibitors were made as solutions of 100 pmol/5 microL and were given in a volume of 5 microL (7-day-old [P7] rats) or 10 microL (21-day-old [P21] rats). Controls were saline injections or injections of the peptide carrier at the same concentration and volumes; there were six animals in each group. In P7 rats, MAC values (in percentage of an atmosphere) were 1.63 +/- 0.0727 (mean +/- SEM) in saline controls, 1.55 +/- 0.141 in carrier controls, 1.54 +/- 0.0800 in rats given PKC-epsilon, and 1.69 +/- 0.0554 in rats given PKC-gamma. In P21 animals, the values were 1.20 +/- 0.0490, 1.31 +/- 0.0124, 1.27 +/- 0.0367, and 1.15 +/- 0.0483, respectively. Injection of the inhibitors did not change MAC in either age group. These results do not support an anesthetic effect on phosphorylation as a mechanism underlying the capacity of inhaled anesthetics to prevent movement in response to noxious stimulation, and they indirectly support a direct action on receptors or ion channels. PMID- 15281509 TI - Gamma-aminobutyric acidA receptors do not mediate the immobility produced by isoflurane. AB - Many inhaled anesthetics enhance the effect of the inhibitory neurotransmitter gamma aminobutyric acid (GABA), supporting the view that the GABAA receptor could mediate the capacity of inhaled anesthetics to produce immobility in the face of noxious stimulation (i.e., MAC, the minimum alveolar concentration required to suppress movement in response to a noxious stimulus in 50% of subjects). However, only limited in vivo data support the relevance of the GABAA receptor to MAC. In the present study we used two findings to test for the relevance of this receptor to immobilization for isoflurane: 1) differences among anesthetics in their capacity to enhance the response of receptor expression systems to GABA: isoflurane (considerable enhancement), xenon (minimal enhancement), and cyclopropane (minimal enhancement); and 2) studies showing that the spinal cord mediates MAC for isoflurane. If GABAA receptors mediate isoflurane MAC, then their blockade in the spinal cord should increase isoflurane MAC more than cyclopropane or xenon MAC and the MAC increase should be proportional to the in vitro enhancement of the GABAA receptor. To test this thesis, isoflurane, cyclopropane, or xenon MAC was determined in rats during intrathecal infusion of artificial cerebrospinal fluid (aCSF) via chronically implanted catheters. Then MAC was redetermined during infusion of 1 microL/min aCSF containing either 0.6 or 2.4 mg/mL picrotoxin, which noncompetitively blocks GABAA receptors. There was no consistent increase in MAC consequent to increasing the picrotoxin dose from 0.6 to 2.4 microg/min, which suggests that maximal blockade of GABAA receptors in the spinal cord had been achieved. Picrotoxin infusion increased MAC approximately 40% with all anesthetics. This indicates that GABA release in the spinal cord influences anesthetic requirement. However, the increase did not consistently differ among anesthetics and did not correlate with in vitro enhancement of GABAA receptors by these anesthetics. This supports the view that GABAA receptors do not mediate immobilization for isoflurane. PMID- 15281510 TI - 2,6 di-tert-butylphenol, a nonanesthetic propofol analog, modulates alpha1beta glycine receptor function in a manner distinct from propofol. AB - The anesthetic propofol (2,6 diisopropylphenol) mediates some of its effects by activating inhibitory chloride currents in the lower brainstem and spinal cord. The effects comprise direct activation of gamma-aminobutyric acid-A and glycine receptors in the absence of the natural agonist, as well as potentiation of the effect of submaximal agonist concentrations. Replacement of propofol's isopropyl groups by di-tert-butyl groups yields a compound without in vivo anesthetic effects. We have studied the effects of propofol and 2,6 di-tert-butylphenol on chloride inward currents via rat alpha1beta glycine receptors heterologously expressed in human embryonic kidney cells. Propofol, but not 2,6 di-tert butylphenol, directly activated glycine receptors; half-maximal current activation was observed with propofol 114 +/- 27 microM. Both compounds potentiated the effect of a submaximal glycine concentration (10 microM) to a maximum value of 136% +/- 71% (propofol) and 279% +/- 109% (2,6 di-tert butylphenol) of the response to glycine 10 microM. The 50% effective concentration for this effect was 12.5 +/- 6.4 microM and 9.4 +/- 10.2 microM for propofol and 2,6 di-tert-butylphenol, respectively. Propofol and its nonanesthetic structural analog do not differ in their ability to coactivate the glycine receptor but differ in their ability to directly activate the receptor in the absence of the natural agonist. PMID- 15281511 TI - Bupivacaine inhibits thromboxane A2-induced vasoconstriction in rat thoracic aorta. AB - Plasma levels of thromboxane A2 (TXA2), an inflammatory mediator inducing platelet aggregation, bronchoconstriction, and vasoconstriction, are increased in the perioperative period. A major role in the pathogenesis of perioperative thromboembolic and ischemic syndromes is attributed to this prostanoid. Local anesthetics (LA) inhibit signaling of TXA2 receptors expressed in cell models. Therefore, we hypothesized that LA may inhibit vasoconstriction induced by the TXA2 analog U46619 in rat thoracic aorta. Rings (3-mm length) of the rat thoracic aorta were mounted in organ baths and isometric contractile force was measured. Rings, with or without endothelium, were incubated for 60 min in bupivacaine (10( 6) or 10(-5) M) or Krebs-Henseleit solution (control group) and subsequently exposed to cumulative concentrations of U46619 (10(-10) to 10(-6) M). The reversibility of the TXA2-induced vasoconstriction by bupivacaine was also studied. Pretreatment of rings with bupivacaine concentration-dependently diminished TXA2-induced contraction in rat aortic rings. We found no significant differences in relaxing effect of bupivacaine between rings with and without endothelium. Contraction in rings established with U46619 could not be reversed by cumulative concentrations of bupivacaine. Bupivacaine inhibited carbachol induced vascular relaxation. This study provides experimental evidence that bupivacaine is an endothelium-independent inhibitor of TXA2-induced vasoconstriction of rat thoracic aorta. PMID- 15281512 TI - 4-chloro-m-cresol cannot detect malignant hyperthermia equivocal cells in an alternative minimally invasive diagnostic test of malignant hyperthermia susceptibility. AB - Malignant hyperthermia (MH) is an inherited skeletal muscle disorder triggered by commonly used anesthetics. Mutated ryanodine receptors have been identified as molecular targets. The sensitivity of myotubes from individuals classified by the in vitro contracture test as MH susceptible (MHS), normal (MHN), and equivocal (MHEH) was assessed for the Ca2+-releasing activity of 4-chloro-m-cresol (4-CmC) and caffeine. In this study, we sought to determine whether 4-CmC can differentiate the MH status of an individual on the basis of the release of intracellular Ca2+, particularly in regard to MHEH diagnosis. Intracellular Ca2+ concentration was determined photometrically with Fura2. Regions of the ryanodine receptor 1 harboring most of the described MH mutations were sequenced from MHS and MHEH cells. One MH mutation (Gly2434Arg) was found in one MHS individual. Results of the caffeine-induced Ca2+ release in MHS and MHN cells correlated well with the in vitro contracture test results. MHS cells showed a higher sensitivity against caffeine and, to a lesser extent, against 4-CmC. Cells of MHEH individuals showed low sensitivities against both caffeine and 4-CmC, comparable to those of the MHN group. Therefore, with myotubes, caffeine was able to discriminate between MHS and MHN cells, but both caffeine and 4-CmC failed to detect MHEH cells. PMID- 15281513 TI - The pupillary effects of intravenous morphine, codeine, and tramadol in volunteers. AB - Opioid analgesics have pharmacological effects in many organ systems, including the eye. Because the metabolites of morphine and codeine contribute to their overall pharmacological effect pupil diameter measurements were made over a 6-h period. We studied the pupillary effects of IV morphine (0.125 mg/kg), codeine (1 mg/kg), tramadol (1.25 mg/kg), or placebo (10 mL 0.9% w/v sodium chloride) in 10 healthy volunteers. Pupil diameter was measured every 30 min using a pupil densitometer. Comparisons of the change in pupil diameter for each drug were made using analysis of variance with repeated measures. No significant change in pupil diameter was observed after placebo. After IV morphine and codeine administration there was a 26% decrease in pupil diameter (P < 0.001). Over the course of the study period, pupil diameter gradually returned to baseline values. After administration of tramadol there were no significant changes in pupil diameter until 150 min after administration, after which there was a significant reduction for the remainder of the study period (P < 0.01). The changes in pupil diameter may be explained in part by the pharmacokinetic profiles of the opioids studied. Measurement of pupil diameter may have a place in monitoring the central effect of opioids. PMID- 15281514 TI - The recovery profile of reduced diaphragmatic contractility induced by propofol in dogs. AB - Propofol decreases contractility of the diaphragm, but no data are available for its effects on recovery. We studied the recovery profile of reduced diaphragmatic contractility induced by propofol in dogs. Animals were divided into 4 groups of 7 each. Group I, without fatigue, received only maintenance fluid; Group II, without fatigue, was infused with propofol; Group III, with fatigue, received no study drug; Group IV, with fatigue, was infused propofol. Propofol at an anesthetic dose (0.1 mg/kg initial dose plus 6.0 mg x kg(-1) x h(-1)) was administered for 60 min. In Groups III and IV, diaphragmatic fatigue was induced by intermittent supramaximal bilateral electrophrenic stimulation at 20-Hz for 30 min. We assessed diaphragmatic contractility by transdiaphragmatic pressure (Pdi). In group II, Pdi at low-frequency (20-Hz) stimulation decreased to less than baseline (P < 0.05), whereas there was no change in Pdi at high-frequency (100-Hz) stimulation. At 10 min after the end of propofol administration, Pdi at 20-Hz stimulation returned to baseline. When fatigue was established, in Groups III and IV, Pdi at 20-Hz stimulation decreased to less than baseline (P < 0.05), whereas Pdi at 100-Hz stimulation did not change. After administering propofol in Group IV, Pdi at 20-Hz stimulation decreased from fatigued values (P < 0.05). At 20 min after the end of propofol administration, Pdi at 20-Hz stimulation returned to fatigued values. We conclude that reduced contractility in nonfatigued and fatigued canine diaphragm induced by propofol recovers within 20 min after the cessation of administration. PMID- 15281515 TI - Pain during injection of propofol: the effect of prior administration of butorphanol. AB - Propofol causes pain or discomfort on injection in 28%-90% of patients. A number of techniques have been tried for minimizing propofol-induced pain with variable results. We compared the efficacy of butorphanol and lidocaine for prevention of propofol-induced pain. One-hundred-fifty ASA I-II adults, undergoing elective surgery were randomly assigned into 3 groups of 50 each. Group I (NS) received normal saline, Group II (L) received lidocaine 2% (40 mg), and Group III (B) received butorphanol 2 mg. All patients received pretreatment solutions made in 2 mL with normal saline administered over 5 s. One min after pretreatment patients received one-fourth of the total calculated dose of propofol (2.5 mg/kg) over 5 s. Assessment of pain with IV propofol was done by using a four-point scale: 0 = no pain, 1 = mild pain, 2 = moderate pain and 3 = severe pain at the time of propofol injection. In the control Group 39 (78%) patients had pain during propofol injection as compared to 21 (42%) and 10 (20%) in the lidocaine and butorphanol groups, respectively (P < 0.05). Butorphanol was the most effective. We therefore suggest the IV pretreatment with butorphanol 2 mg for attenuation of pain associated with propofol injection. PMID- 15281516 TI - The impact of factor XIII on coagulation kinetics and clot strength determined by thrombelastography. AB - Fibrinogen has been shown to be responsible for most protein-mediated clot strength via thrombelastography. However, factor XIII (FXIII) activity also plays a prominent role in the development of clot strength. Thus, we hypothesized that changes in FXIII activity would significantly increase clot strength. FXIII (0%, 1%, 6.25%, 12.5%, 25%, 50%, and 100% normal activity) was placed in a fixed volume of citrated FXIII-deficient plasma with 1% tissue factor and calcium chloride and underwent thrombelastography for 10 min. We measured the variables reaction time (R; a measurement of clot initiation), alpha (a measure of the rate of clot formation), amplitude (A; a measure of clot strength), and shear elastic modulus (G; a measure of clot strength). FXIII activity significantly decreased R in a pattern of exponential decay (R2 = 0.77; P < 0.001). FXIII activity significantly increased alpha, following a sigmoidal pattern (R2 = 0.88; P < 0.001). Finally, increases in FXIII activity significantly increased A and G in a sigmoidal pattern (R = 0.89; P < 0.001). We concluded that FXHI significantly affects R, alpha, A, and G. Thus, transfusion decision making with protein mediated thrombelastographic patterns must account for the contribution of both fibrinogen and FXIII. PMID- 15281517 TI - A new supraglottic airway, the Elisha Airway Device: a preliminary study. AB - We describe the Elisha Airway Device (EAD), a new reusable supraglottic ventilatory device. Its uniqueness consists of its ability to combine three functions in a single device: ventilation, blind and/or fiberoptic-aided intubation without interruption of ventilation, and gastric tube insertion. This study was performed in 70 ASA status I-II, Mallampati class I-II patients undergoing elective knee arthroscopy and receiving general anesthesia with mechanical ventilation. Anesthesia was induced with fentanyl and propofol and was maintained with isoflurane in N20/oxygen. Neuromuscular blockade was achieved with vecuronium. Blind insertion of the device was successful in 96% of patients, with a mean insertion time of 20 +/- 4 s. In these patients it was possible to maintain oxygenation and ventilation throughout the surgical procedure. Gastric tube insertion was successful in all cases. Endotracheal intubation via the EAD was attempted in 20 patients. Blind intubation was possible during the first and second attempts in 15 and 2 patients, respectively. Fiberoptic intubation was then successful in two of the remaining three patients. The EAD is a new alternative in the evolution of supraglottic ventilatory devices; however, further clinical studies are necessary to evaluate its efficacy. PMID- 15281518 TI - Intrathecal clonidine for postoperative analgesia in elderly patients: the influence of baricity on hemodynamic and analgesic effects. AB - Intrathecal (IT) clonidine is an effective analgesic, but it also produces hemodynamic depression and sedation which are likely to be related to IT clonidine's cephalad spread within the cerebrospinal fluid. We hypothesized that IT clonidine's side effects could be reduced without compromising the duration and quality of analgesia by injecting clonidine IT in a hyperbaric solution and elevating the patient's trunk. We prospectively randomized 30 elderly patients to receive IT 150 microg of either isobaric (ISO) or hyperbaric (HYPER) clonidine for postoperative analgesia after surgical repair of traumatic hip fracture. Hemodynamics, IV fluid administration, visual analog pain scores, sedation scores, and clonidine cerebrospinal fluid levels were recorded at fixed intervals. Patients in the ISO group required significantly more crystalloid fluid administration (median, 2500 mL; range, 1500-3000 mL) than those in the HYPER group (median, 1500; range, 500-3000 mL) to maintain adequate arterial blood pressure (P < 0.01). Also, the decrease in heart rate was significantly more pronounced in the ISO than in the HYPER group (P < 0.01). The duration of analgesia was significantly larger in the ISO (median, 400 min; range, 115-400 min) than in the HYPER (median, 265 min; range, 205-400 min) group (P < 0.05). Sedation scores did not differ between groups. We conclude that increasing the baricity of IT clonidine solution in the conditions of our experiment reduces hemodynamic side effects but also analgesia from IT administered clonidine. PMID- 15281519 TI - Intrathecal clonidine potentiates suppression of tactile hypersensitivity by spinal cord stimulation in a model of neuropathy. AB - Spinal cord stimulation (SCS) may provide pain relief in approximately 60%-70% of well selected patients with pain caused by peripheral nerve injury. We have previously demonstrated that intrathecal (IT) administration of small doses of certain drugs, both in experimental animals and in patients, significantly enhances the pain-relieving effect of SCS. The alpha2-adrenoceptor agonist, clonidine, is extensively used as an adjunct to spinal morphine and is suggested to be particularly effective for neuropathic pain, but its clinical use is limited by side effects such as sedation and hypotension. In this study, we investigated the dose-response characteristics of IT clonidine, and whether a subeffective dose of clonidine could enhance the effect of SCS in nerve-injured rats with tactile hypersensitivity (allodynia). Results showed that clonidine, in doses of 1-20 microg, reduced the hypersensitivity in a dose-dependent manner. In rats in which SCS per se failed to suppress tactile hypersensitivity, the combination of SCS and a subeffective dose of clonidine appeared to be highly synergistic and markedly attenuated the hypersensitivity. These results suggest that small doses of IT clonidine may be combined with SCS in neuropathic pain patients who do not obtain satisfactory relief with SCS alone. PMID- 15281520 TI - Preoperative sciatic nerve block decreases mechanical allodynia more in young rats: is preemptive analgesia developmentally modulated? AB - Postoperative sensitivity to tactile stimuli differs as a function of age. In this study, we hypothesized that preoperative sciatic nerve block (SNB), by providing preemptive analgesia, would result in better analgesia than postoperative SNB in the young rat. With the paw incision model of postoperative pain, male Sprague-Dawley rats, aged 2 or 4 wk, underwent general anesthesia and then received a left SNB with 5 microL/g of 0.5% bupivacaine or normal saline. SNB was performed either before or after surgery. Mechanical allodynia was assessed by using von Frey filaments before and at various times after SNB and surgery. In the 2-wk-old rats, preoperative SNB produced a significant reduction in mechanical allodynia, as reflected by a higher threshold at 2, 5, and 24 h when compared with saline control (P < 0.03). At 24 h, the threshold was 4.0 +/- 0.7 g in the preoperative SNB group compared with 1.6 +/- 0.3 g in the postoperative SNB group (P = 0.004). There was no difference at any time point between the preoperative and the postoperative SNB in the 4-wk-old animals. These results suggest that preoperative SNB in young animals provides a preemptive analgesic effect on mechanical allodynia that is age or developmentally dependent. PMID- 15281521 TI - Pain and sensory dysfunction 6 to 12 months after inguinal herniotomy. AB - Inguinal hernia repair is associated with a 5%-30% incidence of chronic pain, but the pathogenesis remains unknown. We therefore evaluated pain and sensory dysfunction by quantitative sensory testing 6-12 mo after open hemiorrhaphy. Before sensory testing, all patients (n = 72) completed a short-form McGill Pain Questionnaire and a functional impairment questionnaire. Sensory dysfunction in the incisional area was evaluated by quantification of thermal and mechanical thresholds, by mechanical pain responses (von Frey/pressure algometry), and by areas of pinprick hypoesthesia and tactile allodynia. The incidence of chronic pain was 28% (20 of 72). Quantitative sensory testing and pressure algometry did not demonstrate differences between the pain and nonpain groups, except for a small but significant increase in pain response to von Frey hair and brush stimulation in the pain group. Hypoesthesia, or tactile allodynia, in the incisional area was observed in 51% (37 of 72) of the patients, but the incidence did not differ significantly between the pain group and the nonpain group (14 of 20 versus 23 of 52; P > 0.3). We concluded that cutaneous hypoesthesia, or tactile allodynia, is common after inguinal hemiotomy but has a low specificity for chronic postherniotomy pain. Factors other than nerve damage may be involved in the development of chronic posthemiotomy pain. PMID- 15281522 TI - Intraarticular pretreatment with ketamine and memantine could prevent arthritic pain: relevance to the decrease of spinal c-fos expression in rats. AB - To determine whether intraarticular pretreatment with N-methyl-D-aspartic (NMDA) receptor antagonist ketamine or memantine currently used in humans has prophylactic analgesia in arthritic pain, we examined the effects of their intraarticular injection before carrageenan injection into the knee joint on pain related behavior and spinal c-Fos expression in rats. Injection of ketamine (0.2 mg and 1 mg) or memantine (0.1 mg, 0.2 mg, and 1 mg) into the knee joint, but not the abdominal cavity, immediately before carrageenan injection (2%, 40 microL) significantly prevented pain-related behavior. The intraarticular injection of ketamine (1 mg) or memantine (0.2 mg) also suppressed c-Fos expression in the laminae I-II and laminae V-VI at the L3-4 spinal level. Subsequent statistical analyses revealed that the degree of the spinal c-Fos expression was correlated with the extent of the pain-related behavior. These results suggest that peripheral administration of NMDA receptor antagonists has prophylactic analgesic effects in arthritic pain, which might be associated with the decrease of central nociceptive signaling. Because ketamine and memantine are currently used in humans and considered clinically safe, they may have therapeutic value in the treatment of joint pain. PMID- 15281523 TI - Intrathecal catheterization and solvents interfere with cortical somatosensory evoked potentials used in assessing nociception in awake rats. AB - We assessed the objective measurement of central sensitization processes in the awake rat after subcutaneous formalin with cortical somatosensory evoked potentials (CSEPs). Cranial extradural electrodes and intrathecal catheters were implanted in adult male Wistar rats. After 7 days of recovery, CSEPs were induced by electrical stimuli at the tail and recorded before/after the injection of 50 microL of 2% formalin into the hindpaw of rats for 1 h. The drug and tested vehicles were delivered intrathecally 5 min before the injection of formalin. The peak-to-peak amplitude of the P1-N1 (the early positive-negative sequence pair of CSEPs) and the baseline-to-peak amplitude of the N2 (the late negative component of CSEPs) were analyzed. We found that the amplitudes of both signals increased (154.3% +/- 10.9% and 168.7% +/- 9.8%, respectively) from 10 min after formalin injection to the end of the 60-min test period. Pretreatment with intrathecal ketorolac dose-dependently prevented the increases induced by formalin in both measured variables. Moreover, the increases in P1-N1 and N2 were markedly attenuated either by intrathecal polyethylene-10 tubing or by the solvents used for injection, thus indicating the need for distinguishing an impaired nociceptive signal from antinociception when the effects of drugs are evaluated. PMID- 15281524 TI - A european, multicenter, observational study to assess the value of gastric-to end tidal PCO2 difference in predicting postoperative complications. AB - Automated online tonometry displays a rapid, semicontinuous measurement of gastric-to-endtidal carbon dioxide (Pr-etCO2) as an index of gastrointestinal perfusion during surgery. Its use to predict postoperative outcome has not been studied in general surgery patients. We, therefore, studied ASA physical status III-IV patients operated on for elective surgery under general anesthesia and a planned duration of >2 h in a European, multicenter study. As each center was equipped with only 1 tonometric monitor, a randomization was performed if more than one patient was eligible the same day. Patients not monitored with tonometry were assessed only for follow-up. The main outcome measure was the assessment of postoperative functional recovery delay (FRD) on day 8. Among the 290 patients studied, 34% had FRD associated with a longer hospital stay. The most common FRDs were gastrointestinal (45%), infection (39%), and respiratory (35%). In those monitored with tonometry (n = 179), maximum Pr-etCO2 proved to be the best predictor increasing the probability of FRD from 34% for all patients to 65% at a cut-off of 21 mm Hg (2.8kPa) (sensitivity 0.27, specificity 0.92, positive predictive value 64%, negative predictive value 70%). We conclude that intraoperative Pr-etCO2 measurement may be a useful prognostic index of postoperative morbidity. PMID- 15281525 TI - Budget negotiation for industry-sponsored clinical trials. AB - The specialty of anesthesia is well suited to attract industry-sponsored clinical trials and research revenues because of its fundamental contributions to surgery, critical care, and pain medicine. However, the performance and budgeting of industry-sponsored clinical research over the past decade has been significantly altered by the rapid growth of commercially oriented networks of contract research organizations and site-management organizations. Further, the competitive nature of today's clinical research climate can make the planning and negotiating of study budgets and contracts stressful, time consuming, frustrating, and full of pitfalls. Because a clinical trial contract is a fixed price agreement, investigators are obligated to perform the work described in the contract, even if the actual costs exceed the study contract. Successful budgeting for the performance of an industry-sponsored clinical trial thus requires a thorough understanding of the direct and indirect costs associated with performing clinical research. We reviewed budget and contractual considerations for the successful negotiation and performance of industry sponsored clinical research. PMID- 15281526 TI - Colonization and bloodstream infection with single- versus multi-lumen central venous catheters: a quantitative systematic review. AB - There is a controversy as to whether the number of lumens in the central venous catheters may impact the incidence of catheter-related bloodstream infection. We performed a systematic search (MEDLINE, PREMEDLINE, Cochrane Library, EMBASE, BIOSIS Previews, CINAHL, HealthSTAR/Ovid healthstar, bibliographies, any language, to April, 2003) for full reports on randomized comparisons of single lumen and multi-lumen catheters. Trials had to report on dichotomous data of catheter colonization or bloodstream infection. Meta-analyses were performed using a fixed effect model. Data were expressed as odds ratio (OR) and number needed-to-treat (NNT) with 95% confidence interval (CI). Five randomized trials (1987-1995) with data on 255 single-lumen and 275 multi-lumen catheters were analyzed. Average insertion times were 8 to 21 days with multi-lumen catheters and 9 to 24 days with single-lumen catheters. In 4 trials, 23 of 176 (13.1%) multi-lumen and 26 of 177 (14.7%) single-lumen catheters were colonized (OR, 0.92; 95% CI, 0.49-1.72). In 5 trials, bloodstream infection occurred with 23 of 275 (8.4%) multi-lumen and with 8 of 255 (3.1%) single-lumen catheters (OR, 2.58; 95% CI, 1.24-5.37; NNT, 19; 95% CI, 11-75). For every 20 single-lumen catheters inserted, one bloodstream infection will be avoided that would have occurred had multi-lumen catheters been used. The risk of catheter colonization is not decreased. Although these conclusions are based on limited data, single-lumen catheters should be used whenever feasible. PMID- 15281527 TI - Transpectoral ultrasound-guided catheterization of the axillary vein: an alternative to standard catheterization of the subclavian vein. AB - Subclavian vein catheterization is associated with failure and complications because of injury to the nearby lung and subclavian artery. Its position, sandwiched between the clavicle and the first rib, makes sonographic imaging difficult. The medially pointed sonography probe makes it difficult to align the needle as well as image the entire needle. The axillary vein lies outside of the thoracic cage and can be easily imaged in its longitudinal view along with the entire needle, guidewire, dilator, and catheter in real-time. All described techniques of venous access using sonography have used transverse images of veins, and the needle is not completely visualized. Five cases of axillary vein catheterization using longitudinal section images of the vein, and following the needle, guidewire, and line with real-time sonography, are described. The use of longer puncture needles and introducer sheaths is suggested. A larger study is required to assess the potential of this technique. PMID- 15281528 TI - Twenty months' routine use of a new percutaneous tracheostomy set using controlled rotating dilation. AB - After a favorable trial period, we introduced the new percutaneous tracheostomy set, PercuTwist, in February of 2002 for our routine procedures. Over the next 20 mo, 90 procedures were performed with minimal complications. To prospectively evaluate this experience, we collected information on reasons for unit admission, operators' previous experience, the duration of prior tracheal intubation, the time needed for the procedure, the grading of the difficulty, the amount of bleeding, and the complications of the procedure. Twenty-two of 90 (24.4%) procedures were performed by senior consultants with experience; 68 of 90 (75.6%) were safely performed by intensive care residents under close bedside supervision. The mean time needed for the procedure was 13 min 7 s. In only one procedure during the entire study was any difficulty observed during the insertion process. This occurred because the initial skin incision was too small. However, no major bleeding or complications were encountered. PMID- 15281529 TI - Ventilation with smaller tidal volumes: a quantitative systematic review of randomized controlled trials. AB - In this quantitative systematic review we assessed the effects of ventilation with smaller tidal volume (VT) on morbidity and mortality in patients aged 16 yr or older affected by acute lung injury and acute respiratory distress syndrome. Five randomized trials (1202 patients) comparing ventilation using smaller VT and/or low airway driving pressure (plateau pressure 30 cm H2O or less), resulting in VT of 7 mL/kg or less versus ventilation that uses VT in the range of 10 to 15 mL/kg, were identified after a systematic search of The Cochrane Central Register of Controlled Trials, MEDLINE, EMBASE, CINAHL, databases of current research, reference lists, and "gray literature." Mortality at day 28 was significantly reduced by lung-protective ventilation (relative risk [RR], 0.74; confidence interval [CI], 0.61-0.88), whereas beneficial effect on long-term mortality was uncertain (RR, 0.84; CI, 0.68-1.05). The comparison between small and conventional VT was not significantly different if a plateau pressure less than or equal to 31 cm H2O in the control group was used (RR, 1.13; CI, 0.88 1.45). Clinical heterogeneity, such as different lengths of follow-up and higher plateau pressures in control arms in two trials, make the interpretation of the combined results difficult. PMID- 15281530 TI - Does arginine vasopressin influence the coagulation system in advanced vasodilatory shock with severe multiorgan dysfunction syndrome? AB - Arginine vasopressin (AVP) is a potent supplementary vasopressor in advanced vasodilatory shock, but decreases in platelet count have been reported during AVP therapy. In this study we evaluated the effects of AVP infusion on the coagulation system in advanced vasodilatory shock when compared to norepinephrine (NE) infusion alone. Forty-two patients with advanced vasodilatory shock (NE requirements >0.5 microg x kg(-1) x min(-1), mean arterial blood pressure <70 mm Hg) were prospectively randomized to receive an additional AVP infusion (4 U/h) or NE infusion alone. Most patients received coagulation active treatment (fresh frozen plasma, thrombocyte concentrates, coagulation factors, and continuous veno venous hemofiltration with heparin). At baseline and 1, 24, and 48 h after randomization, coagulation laboratory variables and a modified thrombelastography were measured. There were no differences between groups in plasmatic coagulation variables. Although there was no significant difference between groups, platelet count significantly decreased in AVP patients (P = 0.036). There were no differences in results of modified thrombelastography analyses between groups. AVP infusion in advanced vasodilatory shock with severe multiorgan dysfunction syndrome does not increase plasma concentrations of Factor VIII, von Willebrand Factor antigen, and ristocetin Co-Factor but may stimulate platelet aggregation and induce thrombocytopenia. Global coagulation, assessed by modified thrombelastography, is not different from patients receiving NE infusion alone. PMID- 15281531 TI - Supplemental oxygen and carbon dioxide each increase subcutaneous and intestinal intramural oxygenation. AB - Oxidative killing by neutrophils, a primary defense against surgical pathogens, is directly related to tissue oxygenation. We tested the hypothesis that supplemental inspired oxygen or mild hypercapnia (end-tidal PCO2 of 50 mm Hg) improves intestinal oxygenation. Pigs (25 +/- 2.5 kg) were used in 2 studies in random order: 1) Oxygen Study: 30% versus 100% inspired oxygen concentration at an end-tidal PCO2 of 40 mm Hg, and 2) Carbon Dioxide Study: end-tidal PCO2 of 30 mm Hg versus 50 mm Hg with 30% oxygen. Within each study, treatment order was randomized. Treatments were maintained for 1.5 h; measurements were averaged over the final hour. A tonometer inserted in the subcutaneous tissue of the left upper foreleg measured subcutaneous oxygen tension. Tonometers inserted into the intestinal wall measured intestinal intramural oxygen tension from the small and large intestines. Oxygen 100% administration doubled subcutaneous oxygen partial pressure (PO2) (57 +/- 10 to 107 +/- 48 mm Hg, P = 0.006) and large intestine intramural PO2 (53 +/- 14 to 118 +/- 72 mm Hg, P = 0.014); intramural PO2 increased 40% in the small intestine (37 +/- 10 to 52 +/- 25 mm Hg, P = 0.004). An end-tidal PCO2 of 50 mm Hg increased large intestinal PO2 approximately 16% (49 +/- 10 to 57 +/- 12 mm Hg, P = 0.039), whereas intramural PO2 increased by 45% in the small intestine (31 +/- 12 to 44 +/- 16 mm Hg, P = 0.002). Supplemental oxygen and mild hypercapnia each increased subcutaneous and intramural tissue PO2, with supplemental oxygen being most effective. PMID- 15281532 TI - Selective inducible nitric oxide inhibition can restore hemodynamics, but does not improve neurological dysfunction in experimentally-induced septic shock in rats. AB - In this study, we evaluated the time course of changes in inducible nitric oxide synthase (iNOS) in the brain by using the rat model of sepsis induced by cecal ligation and puncture (CLP) and examined whether selective iNOS inhibition can prevent the hemodynamic and neurological changes induced by sepsis. Male Wistar rats were randomly divided into four groups: control, sham, CLP, and CLP + the selective iNOS inhibitor L-N6-(1-iminoethyl)-lysine (L-NIL). Septic shock was induced in the rats by CLP under pentobarbital anesthesia, and then we measured hemodynamic variables, neurological indicators, blood gases, plasma levels of nitrate/nitrite (an indicator of the biosynthesis of NO), and brain iNOS activity and nitrotyrosine levels after 1, 6, 12, and 24 h. Plasma nitrite was increased at 12 and 24 h in the CLP group. The activity of iNOS in the brain was increased at 12 and 24 h after CLP (at 12 h: control, 0.3 +/- 0.05; sham, 0.3 +/- 0.1; CLP, 1.3 +/- 0.08*; CLP + L-NIL, 0.33 +/- 0.1 fmol x mg(-1) x min(-1); at 24 h: control, 0.27 +/- 0.08; sham, 0.31 +/- 0.1; CLP, 1.0 +/- 0.3*; CLP + L-NIL, 0.34 +/- 0.1 fmol x mg(-1) x min(-1); mean +/- SD; *P < 0.05). Brain nitrotyrosine was increased at 24 h after CLP (at 24 h: control, 6.7 +/- 0.4; sham, 6.7 +/- 0.5; CLP, 11.2 +/- 2.8*; CLP + L-NIL, 7.52 +/- 0.5 densitometric units; means +/- SD; *P < 0.01). In contrast, in both the CLP and CLP + L-NIL groups, the consciousness reflex was significantly decreased at 24 h after CLP. Selective iNOS inhibition restored the hemodynamic changes induced by sepsis but could not improve neurological dysfunction. PMID- 15281533 TI - Clinical management of cardiogenic shock associated with prolonged propofol infusion. AB - This case report details the development of cardiogenic shock after craniotomy in a patient sedated with a propofol infusion. The patient survived with the assistance of extracorporeal membrane oxygenation. A literature review summarizes the syndrome of cardiogenic shock associated with prolonged propofol infusion. This is the first report of survival in this syndrome resuiting from mechanical circulatory support. PMID- 15281534 TI - Respiratory distress after intrathecal baclofen withdrawal. AB - We present the case of a 19-yr-old woman with a history of generalized dystonia who developed sudden onset of adductor spasms of the vocal cords and increased dystonia after the interruption or intrathecal baclofen therapy. Her symptoms resolved after intrathecal baclofen was restored. In patients with dystonia receiving intrathecal baclofen therapy, the onset of dyspnea associated with increased muscle tone should prompt the investigation of baclofen withdrawal. PMID- 15281535 TI - Arterio-jugular difference of oxygen content and outcome after head injury. AB - This study investigated AJDO2 (arterio-jugular difference of oxygen content) in a large sample of severely head-injured patients to identify its pattern during the first days after injury and to describe the relationship of AJDO2 with acute neurological severity and with outcome 6 mo after trauma. In 229 comatose head injured patients, we monitored intracranial pressure, cerebral perfusion pressure, and AJDO2. Outcome was defined 6 mo after injury. Jugular hemoglobin oxygen saturation (SjO2) averaged 68%. The mean AJDO2 was 4.24 vol% (SD, 1.3 vol%). There were 80 measurements (4.6%) with SjO2 <55% and 304 (17.6%) with saturation >75%. AJDO2 was higher than 8.7 vol% in 8 measurements (0.4%) and was lower than 3.9 vol% in 718 (42%) measurements. AJDO2 was higher during the first tests and decreased steadily over the next few days. Cases with a favorable outcome had a higher mean AJDO2 (4.3 vol%; SD, 0.3 vol%) than patients with severe disability or vegetative status (3.8 vol%; SD, 1.3 vol%) and patients who died (3.6 vol%; SD, 1 vol%). This difference was significant (P < 0.001). We conclude that low levels of AJDO2 are correlated with a poor prognosis, whereas normal or high levels of AJDO2 are predictive of better results. PMID- 15281536 TI - The effects of the delta-opioid agonist SNC80 on hind-limb motor function and neuronal injury after spinal cord ischemia in rats. AB - Recent investigation suggested neuroprotective efficacy of a delta-opioid agonist in the brain. We investigated the effects of intrathecal treatment with a delta opioid agonist (SNC80) on spinal cord ischemia (SCI) in rats. SCI was induced with an intraaortic balloon catheter. The animals were randomly allocated to one of the following five groups: 1) SNC80 before 9 min of SCI (SNC-9; n = 12), 2) vehicle before 9 min of SCI (V-9; n = 12), 3) SNC80 before 11 min of SCI (SNC-11; n = 10), 4) vehicle before 11 min of SCI (V-11; n = 12), or 5) sham (n = 12). SNC80 (400 nmol) or vehicle was administered 15 min before SCI. Forty-eight hours after reperfusion, hind-limb motor function was assessed by using the Basso, Beattie, Bresnahan (BBB) scale (0 = paraplegia; 21 = normal) and histological assessment of the L4 and L5 spinal segments was performed. BBB scores in the SNC 9 group were higher compared with those in the V-9 group (P < 0.05), whereas there were no differences in BBB scores between the SNC-11 and V-11 groups. There were significantly more normal neurons in the SNC-9 and SNC-11 groups than in the V-9 and V-11 groups (P < 0.05). The results indicate that intrathecal treatment with the delta-opioid agonist SNC80 can attenuate hind-limb motor dysfunction and neuronal injury after SCI in rats. PMID- 15281537 TI - Lower limb wrapping prevents hypotension, but not hypothermia or shivering, after the introduction of epidural anesthesia for cesarean delivery. AB - The decrease of arterial blood pressure and body temperature after epidural or spinal anesthesia is thought to be the result of sympathetic block, which could cause pooling and redistribution of blood into the lower extremities. Studies have demonstrated that leg wrapping with elastic bandages may reduce the incidence of hypotension after spinal anesthesia. We tried to extend these previous observations to epidural anesthesia by testing the hypothesis that leg wrapping with elastic bandages should decrease the incidence of hypotension in patients receiving epidural anesthesia. Moreover, we evaluated the effect of this maneuver as regards hypothermia and shivering. Sixty parturients were randomly allocated to receive either leg wrapping with tight elastic bandages (leg-wrapped group) or not (control group) before anesthesia. Sublingual temperature was observed at five periods: baseline, immediately after epidural anesthesia, abdominal skin disinfection, skin incision, and delivery. Hypotension and shivering during the observation periods were also recorded. The incidence of hypotension was significantly less frequent (P = 0.03) in the leg-wrapped group (23%) compared with the control group (50%). Shivering incidences were similar in both groups (70% versus 70%). Sublingual temperature decreased significantly (P < 0.001) throughout the procedure in each group. However, no differences were found between the two groups at each designated observation, even if compared by the magnitude of temperature decrease. We conclude that although leg wrapping with elastic bandages prevents maternal hypotension after epidural anesthesia, it does not reduce the incidence or magnitude of hypothermia or prevent shivering. PMID- 15281538 TI - Identification of the epidural space: loss of resistance with air, lidocaine, or the combination of air and lidocaine. AB - The ideal technique for identifying the epidural space remains unclear. Five hundred-forty-seven women in labor who requested epidural analgesia were randomly allocated to three groups according to the technique by which the epidural space was identified: 1) loss-of-resistance with air (air; n = 180), 2) loss-of resistance with lidocaine (lidocaine; n = 185), and 3) loss-of-resistance with both air and lidocaine (air-plus-lidocaine; n = 182). We assessed ease of epidural catheter insertion, characteristics of the blockade, quality of analgesia, and complications. The inability to thread the epidural catheter occurred in 16% of the air, 4% of the lidocaine, and 3% of the air-plus-lidocaine patients (P < 0.001). More patients from the air group had unblocked segments (6.6% versus 3.2% and 2.2%, respectively; P < 0.02). The incidence of accidental dural puncture was greater in the air group (1.7% versus 0% in the other two groups; P < 0.02). Pain scores, time to onset of analgesia, upper sensory level, motor blockade, and the incidence of hypotension, transient neurological deficits, postpartum urinary retention, and postdural puncture headache were comparable. Identification of the epidural space with air was more difficult and caused more dural punctures than with lidocaine or air plus lidocaine. Additionally, sequential use of air and lidocaine had no advantage over lidocaine alone. PMID- 15281539 TI - Postoperative analgesia after total knee replacement: the effect of an obturator nerve block added to the femoral 3-in-1 nerve block. AB - Femoral nerve block (FNB) does not consistently produce anesthesia of the obturator nerve. In this single-blind, randomized, controlled study we added a selective obturator nerve block (ONB) to FNB to analyze its influence on postoperative analgesia after total knee replacement (TKR). Before general anesthesia, 90 patients undergoing TKR received FNB (Group 1), FNB and selective ONB (Group 2), or placebo FNB (Group 3). Postoperative analgesia was further provided by morphine IV via patient-controlled analgesia. Analgesic efficacy and side effects were recorded in the first 6 h after surgery. Adductor strength decreased by 18% +/- 9% in Group 1 and by 78% +/- 22% in Group 2 (P < 0.0001). Total morphine consumption was reduced in Group 2 compared with Groups 1 and 3 (P < or = 0.0001). Patients in Group 2 reported lower pain scores than those in Groups 1 and 3 (P = 0.0003). The incidence of nausea was more frequent in Groups 1 and 3 (P = 0.01). We conclude that FNB does not produce complete anesthesia of the obturator nerve. Single-shot FNB does not provide additional benefits on pain at rest over opioids alone in the early postoperative period. The addition of an ONB to FNB improves postoperative analgesia after TKR. PMID- 15281541 TI - Inadvertent cervical epidural catheter placement via the caudal route using electrical stimulation. AB - Inadvertent placement of an epidural catheter in the cervical region via the caudal route is described in an infant who underwent revision of a fundoplication. We attempted electrical stimulation (the Tsui test) via the epidural catheter to confirm correct placement and positioning of the catheter tip. In this case, the epidural catheter was inadvertently advanced to the cervical region, resulting in stimulation of the phrenic nerve. These diaphragmatic twitches were misinterpreted as chest wall twitches, and it was incorrectly assumed that the catheter was in the thoracic region. To avoid misinterpretation of the stimulation level, the catheter should be continuously stimulated while it is advanced. We also recommend that the catheter length be estimated before insertion (although doing so did not help in this case) and that the catheter position be radiographically confirmed after surgery. PMID- 15281540 TI - Muscle weakness and paresthesia associated with epidural analgesia in a patient with an intrathecal neurofibrolipoma as part of a tethered cord syndrome. AB - We report a case of a 75-yr-old female patient in whom motor deficits and paresthesias occurred after lumbar epidural analgesia. These symptoms were eventually found to be due to a tethered cord syndrome. An epidural catheter was inserted for analgesia after colon surgery. The postoperative course was characterized by fluctuating sensory and motor symptoms. A magnetic resonance imaging scan showed an intraspinal mass, which was removed by laminectomy. The presented complication is of major interest because the intraspinal tumor, which must have been present for years, became acutely symptomatic. Tethered cord syndrome is caused by a limited longitudinal mobility of the cord. It is often seen as a part of spinal closure defects and is also associated with intrathecal tumors. Typically, adult patients complain of weak legs, paresthesias of the legs, and urinary incontinence. However, our patient had denied any muscular or neurological problems or urinary incontinence during the preoperative interview. Postoperative electromyogram and electroneurography ascertained chronic neurogenic lesions of multiple lumbar and sacral nerve roots. Three months after the operation, the patient was able to walk 100 m with a crutch. PMID- 15281542 TI - A restrictive use of both autologous donation and recombinant human erythropoietin is an efficient policy for primary total hip or knee arthroplasty. AB - A limitation of preoperative autologous blood donation (PABD) in nonanemics and the use of recombinant human erythropoietin (EPO) in anemics (baseline hematocrit [Hct] < or = 39%) could be an efficient approach of the cost-benefit ratio of transfusion during primary total hip (THA) or knee (TKA) arthroplasties. We evaluated the consequences on transfusion rates and costs of two different applications of a transfusion policy based on personal requirements during primary THA or TKA. This quality assurance observational study compared two prospective successive time periods; each included successive patients treated by the same medical team and standardized care. In Study 1 (n = 182), PABD was indicated if there were insufficient estimated red blood cell reserve and a life expectancy > or = 10 yr, no use of EPO, and identical criteria for any transfusion. Because this policy led to a 50% allogeneic transfusion rate when baseline Hct < or = 37% and autologous blood wastage in the nonanemics (baseline Hct > 39%), 2 refinements were introduced in Study 2 (n = 708): EPO without PABD when baseline Hct < or = 37%, and life expectancy > or = 10 yr, and avoidance of PABD in nonanemics. This novel care induced a marked decrease in transfusion rates (respectively, from 41% to 7%, P < 0.0002, in nonanemics; from 58% to 27%, P < 0.003, in anemics; and from 43% to 12%, P < 0.0001, overall), with no change in allogeneic transfusion (10%) and discharge Hct, and a 39% financial savings. This saving effect is a result of the suppression of PABD in nonanemics, who represent 75% of this surgical population. Although erythropoietin is expensive, it can be used with cost savings in selected patients because the overall cost of transfusion is reduced. PMID- 15281543 TI - The new perilaryngeal airway (CobraPLA) is as efficient as the laryngeal mask airway (LMA) but provides better airway sealing pressures. AB - The Laryngeal Mask Airway (LMA) is a frequently used efficient airway device, yet it sometimes seals poorly, thus reducing the efficacy of positive-pressure ventilation. The Perilaryngeal Airway (CobraPLA) is a novel airway device with a larger pharyngeal cuff (when inflated). We tested the hypothesis that the CobraPLA was superior to the LMA with regard to insertion time and airway sealing pressure and comparable to the LMA in airway adequacy and recovery characteristics. After midazolam and fentanyl administration, 81 ASA physical status I-II outpatients having elective surgery were randomized to receive an LMA or CobraPLA. Anesthesia was induced with propofol (2.5 mg/kg IV), and the airway was inserted. We measured 1) insertion time; 2) adequacy of the airway (no leak at 15-cm-H2O peak pressure or tidal volume of 5 mL/kg); 3) airway sealing pressure; 4) number of repositioning attempts; and 5) sealing quality (no leak at tidal volume of 8 mL/kg). At the end of surgery, gastric insufflation, postoperative sore throat, dysphonia, and dysphagia were evaluated. Data were compared with unpaired Student's t-tests, chi2 tests, or Fisher's exact tests; P < 0.05 was significant. Patient characteristics, insertion times, airway adequacy, number of repositioning attempts, and recovery were similar in each group. Airway sealing pressure was significantly greater with CobraPLA (23 +/- 6 cm H2O) than LMA (18 +/- 5 cm H2O, P < 0.001). The CobraPLA has insertion characteristics similar to the LMA but better airway sealing capabilities. PMID- 15281544 TI - Lightwand-assisted intubation of patients in the lateral decubitus position. AB - In some situations, patients need endotracheal intubation to maintain airway patency while they are constrained in the lateral position. In this study we compared lightwand-guided intubation of 120 randomly enrolled patients placed in the supine, right, or left lateral position. Group S patients were initially placed in the supine position, and subsequent to the artificial airway having been established they were turned to the lateral decubitus position. Group R patients were initially placed in a right decubitus position during induction and intubation. Group L patients were initially placed in a left decubitus position during induction and intubation. The duration of each intubation attempt, the total time to successful intubation, and the incidence of intubation-related intraoral injury, hemodynamic changes, and postoperative sore throat and hoarseness were recorded. Intubation took a similar length of time in the supine (14.5 +/- 13.4 s), left lateral (13.3 +/- 10.2 s), and right lateral positions (15.5 +/- 13.0 s) and resulted in a similar trend in hemodynamic changes. Patients in the lateral and supine positions revealed a comparable incidence of successful first-attempt intubation, sore throat, hoarseness, oral mucosal injury, and dysrhythmia. Insignificantly more esophageal intubations were performed in the lateral position in the first attempt at intubation; however, all patients were correctly intubated shortly after reattempting intubation. We concluded that lightwand-assisted intubation is easily performed and a similar technique may be used whether the patient is in a lateral, recumbent, or a supine position. This alternative technique should be practiced and is recommended for patients who must remain in a lateral position during intubation and surgery. PMID- 15281546 TI - Does the presence of a tracheal bronchus affect the margin of safety of double lumen tube placement? AB - During double-lumen tube (DLT) placement, the anesthesiologist must be mindful of the margin of safety. We determined how this margin is affected by the presence of a tracheal bronchus by elucidating the mathematical relationship between some relevant physical dimensions of the trachea, bronchi, and DLT. Our results suggest that a tracheal bronchus only rarely affects the intrinsic margin of safety of DLT placement. When the tracheal bronchus is located much higher than its most frequently seen location (within 2 cm from the carina), however, there is increased risk that it could be blocked by the tracheal cuff of a left-sided DLT. PMID- 15281545 TI - Mild hyperthermia down-regulates receptor-dependent neutrophil function. AB - Mild hypothermia impairs resistance to infection and, reportedly, impairs phagocytosis and oxidative killing of unopsonized bacteria. We evaluated various functions at 33 degrees-41 degrees C in neutrophils taken from volunteers. Adhesion on endothelial cells was determined using light microscopy. Adhesion molecule expression and receptors, phagocytosis, and release of reactive oxidants were assessed using flow cytometric assays. Adhesion protein CD11b expression on resting neutrophils was temperature-independent. However, up-regulation of CD11b with tumor necrosis factor (TNF)-alpha was increased by hypothermia and decreased with hyperthermia. Neutrophil adhesion to either resting or activated endothelial cells was not temperature-dependent. Bacterial uptake was inversely related to temperature, more so with Escherichia coli than Staphylococcus aureus. Temperature dependence of phagocytosis occurred only wi thopsonized bacteria. Hypothermia slightly increased N-formyl-L-methionyl-L-leucyl-phenylalanine receptors on neutrophils: hyperthermia decreased expression, especially with TNF alpha. N-formyl-L-methionyl-L-leucyl-phenylalanine-induced H2O2 production was inversely related to temperature, especially in the presence of TNF-alpha. Conversely, phorbol-13-myristate-12-acetate, an activator of protein kinase C, induced an extreme and homogenous release of reactive oxidants that increased with temperature. In contrast to nonreceptor-dependent phagocytosis and oxidative killing, several crucial receptor-dependent neutrophil activities show temperature-dependent regulation, with hypothermia increasing function. The temperature dependence of neutrophil function is thus more complicated than previously appreciated. PMID- 15281547 TI - The Society for Pediatric Anesthesia: the first joint meeting of the Society for Pediatric Anesthesia and the Japanese Society of Pediatric Anesthesiology. PMID- 15281548 TI - Comparisons between Level 1 and other fast flow fluid warming systems. PMID- 15281549 TI - Propofol-remifentanil target-controlled anesthesia in a patient with hyperkalemic familial periodic paralysis. PMID- 15281550 TI - Use of small-dose bupivacaine (3 mg vs 4 mg) for unilateral spinal anesthesia in the outpatient setting. PMID- 15281551 TI - Continuous inflation of a punctured cuff during pulmonary surgery. PMID- 15281552 TI - Pneumocephalus following an uneventful lumbar puncture: does the gauge of a spinal needle matter? PMID- 15281553 TI - Monitoring of patients receiving long-term opioid therapy. PMID- 15281554 TI - Termination of idiopathic persistent singultus (hiccup) with suprasupramaximal inspiration. PMID- 15281555 TI - Postoperative intrathecal morphine for analgesia after major orthopedic surgery? PMID- 15281556 TI - Oxygen consumption: validation of the closed-circuit PhysioFlex anaesthesia machine. PMID- 15281557 TI - Malposition of a pulmonary artery catheter in a patient with an inferior vena cava filter. PMID- 15281558 TI - More information from the pressure gauge of oxygen cylinders. PMID- 15281559 TI - Internal or external diameter? PMID- 15281560 TI - A personal experience using Limoge's current during a major surgery. PMID- 15281561 TI - Renal dysfunction after hydroxyethyl starch. PMID- 15281562 TI - The LMA ProSeal may not be the best option for difficult to intubate/ventilate patients. PMID- 15281563 TI - Left ventricular function after cardiopulmonary bypass is related to the length dependent regulation of myocardial function. PMID- 15281564 TI - ProSeal laryngeal mask airway foldover detection. PMID- 15281565 TI - [Ethics]. PMID- 15281566 TI - [A committee for scientific misconduct is necessary]. PMID- 15281567 TI - [Outlook on mankind]. PMID- 15281568 TI - [Autonomy and its limits]. PMID- 15281569 TI - [The need of new concepts]. PMID- 15281570 TI - [Research ethics]. PMID- 15281571 TI - [Research ethics and revised law regarding the Danish Central Scientific Ethical Committee]. PMID- 15281572 TI - [The regional ethical committees. Responsibilities and challenges]. PMID- 15281573 TI - [Attitudes toward and experiences with clinical research among current and potential research subjects]. PMID- 15281574 TI - [Research ethics in a perspective of developing countries]. PMID- 15281575 TI - [Fair distribution and prioritization of health care]. PMID- 15281576 TI - [The need of new and wider perspectives]. PMID- 15281577 TI - [Ethical considerations in the termination of intensive care]. PMID- 15281578 TI - [Research on the dead]. PMID- 15281579 TI - [Death on request?]. PMID- 15281580 TI - [Animal experiments--an ethical dilemma]. PMID- 15281581 TI - [Is clinical research in Denmark directed by drug industry?]. PMID- 15281582 TI - [Asasantin Retard or Persantin Retard+ASA?]. PMID- 15281583 TI - [Interaction between clopidogrel and statins?]. PMID- 15281584 TI - [Arthroscopy, deep infection and results statement]. PMID- 15281585 TI - A call to action. Healthcare executives must translate social values into workable healthcare programs. PMID- 15281586 TI - Are people your priority? How to engage your workforce. PMID- 15281587 TI - Employee retention strategies: lessons from the best. PMID- 15281588 TI - The roadmap to diversity, inclusion, and high performance. PMID- 15281589 TI - Addressing rural ethics issues. The characteristics of rural healthcare settings pose unique ethical challenges. PMID- 15281590 TI - Building green hospitals. Improve your economic viability, environmental impact, and community standing. PMID- 15281591 TI - Starting a corporate college. A New York hospital takes an in-depth approach to employee training. PMID- 15281592 TI - Hospital-sponsored childcare. On-site childcare can help hospitals become employers of choice. PMID- 15281593 TI - What should boards be doing? Or, its my backyard. PMID- 15281594 TI - Upstream investment. A Georgia hospital program focuses on men's health. PMID- 15281595 TI - Pay for performance gaining popularity. This evolving payment methodology holds significant implications for providers. PMID- 15281596 TI - Ask the expert. Online degrees are gaining popularity with students and employers. PMID- 15281597 TI - Make the local connection to ACHE. Partnership between ACHE and chapters enhances programs and services. PMID- 15281598 TI - Cancer trends in Italy: figures from the cancer registries (1986-1997). AB - The Italian Network of Cancer Registries analyzed incidence and mortality cancer trends during the period 1986-1997 Overall, 525,645 incident cancers and 269,902 cancer deaths (in subjects 15 years and older) were included. Age-adjusted rates, joinpoints (points in time where trend significantly changes from linearity) and estimated annual percent changes in rates (EAPC) were computed. Overall cancer incidence was significantly increasing in both sexes and cancer mortality was significantly decreasing (since 1991 among males). Incidence and mortality trends are summarised for single cancer sites. Crude rates are also showed to evaluate the effect of population ageing in terms of diagnostic and therapeutic burden for the National Health System. PMID- 15281599 TI - The Italian Network of Cancer Registries (AIRT). AB - The Italian Network of Cancer Registries (AIRT) built three databases for the study of cancer trends in Italy during 1986-1997: incident cases, deaths and populations observed by the eighteen Italian Cancer Registries active on about 23% of the Italian resident population. The incidence data have been validated through ad hoc programmes and CHECK-IARC. A pool of nine Cancer Registries, that had an almost complete coverage for the study period 1986-1997, has been chosen for the trend analysis (pool AIRT). 525,645 cases and 269,902 deaths have been analysed on a total of 8 million residents. PMID- 15281600 TI - Italian Network of Cancer Registries. Methods. AB - The trend analysis of the data base of the Italian Network of Cancer Registries has had a descriptive approach. The period of study has been from 1986 to 1997. Within this period we have included only nine Registries that were active for at least ten years (pool AIRT). For three Registries few lacking incidence years have been estimated according to their observed data. For the pool of Registries standardised (standard = European population) incidence and mortality rates have been computed with the direct method. The so called joinpoint analysis has been used to detect temporal points of trend change. Trends have been summarised by means of estimated annual percent change (EAPC) of the rate. Age-specific and birth cohort-specific rates have been also computed. PMID- 15281601 TI - Quality and completeness indices. AB - In the present study, time trends of the main quality indices in the pool of the Italian Network of Cancer Registries (pool AIRT) are presented for the period 1986-1997. The percentage of cases with morphological verification increased from 75% to 83% among males and from 77% to 85% among females. Cases known only from death certificates (DCO) decreased over time in both sexes; in recent years they were about 3% of the whole case-series. Mortality incidence ratio decreased over time, from 66% to 54% among males and from 55% to 48% among females. Also cases with unknown primary site showed a decreasing time tendency. Such quality indices are also presented for specific cancer sites and for elderly subjects (75+ years). The overall trends of quality indices showed an improvement in both the activity of Cancer Registries and of the Health System. PMID- 15281602 TI - The use of models for estimating overall incidence trend. AB - Several approaches to evaluate an overall trend including Registries with different length of activity with a defined period of time have been evaluated in the cancer trend study of the Italian Network of Cancer Registries (AIRT). The study used the nine Registries of the pool AIRT in the period 1986-1997. Data for 1986-1993 from the Registry of Ragusa have been excluded to simulate the inclusion of a Registry with partial observation. Four different approaches have been used: Golden standard (GS) as a weighted mean of the observed standardised (European population) rates in the whole pool AIRT; 2) Raw trend (TG) computed with partial data of Ragusa; 3) Model trend (TM) estimated by means of a log linear model including age, registry and period as independent variables and applied on the partial data set; Adjusted trend (TC) computed on complete data for eight Registries, observed 1994-1997 data of Ragusa and estimated data for Ragusa for 1986-1993 computed by (3) model. The trend estimates, as estimated annual percent change of the standardised rates (EAPC) have been compared with GS. The results of this preliminary analysis do not recommend the use of raw trend and indicate adjusted trend as the most reliable method for trend estimate. PMID- 15281603 TI - Head and neck cancers: oral cavity, pharynx, and larynx. AB - The present paper on head and neck cancers analyses oral cavity pharynx and larynx cancers in the database of the Italian Network of Cancer Registries (pool AIRT) during the period 1986-1997. The analysis of 7,234 oral cavity incident cases shows diverging trends between sexes; in fact, rates decrease significantly among males and increase among females. Oral cavity cancer mortality rates (3,032 deaths) significantly decrease in the period 1986-1997 in both sexes. As regards to pharynx (3,728 incident cases and 2,517 deaths), decreasing trends have been observed for both incidence and mortality, statistically significant only among males. Larynx cancers (9,842 incident cases and 3,946 deaths) showed a significant decreasing trend for both incidence and mortality, among males; trends are stable among females. In the present report we refer to head and neck cancers as those arising in the anatomical sites coded in ICDO-2 with C00-C14 (cancer of the lip, oral cavity and pharynx) and C32 (larynx). Such a grouping is justified, at least for epithelial histological types, as these cancers share a common series of risk factors: EBV, HPIV, smoke, alcohol, diet and professional exposures. PMID- 15281604 TI - Upper gastrointestinal tract cancers: oesophagus, stomach, liver, gallbladder and biliary ducts, pancreas. AB - In this paper temporal trend of the upper gastrointestinal tract cancers in the Italian Network of Cancer Registries (pool AIRT) are analysed during the period 1986-1997. Oesophagus cancer (4477 cases and 4226 deaths analysed) showed a decrease in incidence rates that was statistically significant among males and less evident in recent years among females. Mortality is significantly decreasing both among males (since 1993) and females. Stomach cancer (34282 cases and 26430 deaths) had a stable decreasing trend, in both sexes, for both incidence which decreased at a rate of more 3% every year, and of mortality, mean annual rate decrease over 4%. As regards liver cancer (13893 cases 13655 deaths) an increasing incidence trend (up to 1993 among males) has been documented; mortality was stable. Cancers of the biliary tract (6662 cases and 5065 deaths) showed stable rates both in incidence (slightly decreasing among females) than in mortality. Pancreas cancer (13300 cases and 12937 deaths) presented increasing incidence in both sexes with stable mortality rates. PMID- 15281605 TI - Colorectal cancer. AB - Temporal trend analysis for incident cases (n = 60,837) and deaths (n = 28,936) for colorectal cancer in the pool of the Italian Network of Cancer Registries (pool AIRT) in the period 1986-1997, shows a statistically significant increase of incidence rates in both males (mean annual increase of + 1.7%) and females (+ 0. 6%). Mortality rates showed a statistically significant decrease, in both sexes, of -0.7%/year among males and -0.9% among females. PMID- 15281606 TI - Respiratory tract cancers: lung and mesothelioma. AB - The trend analysis of lung cancer in the database of the Italian Network of Cancer Registries (pool AIRT), showed, among males (52,267 incident cases and 46, 726 deaths included in the study) a statistically significant decrease of incidence and mortality in the period 1986-1997; incidence rates decreased by about 1.4%/year and mortality rates by about 1.6%/year. Among females, lung cancer trends are rather different from that of males, according to diverging trends in tobacco smoking exposures; in fact, both incidence (+1.2%/year) and mortality (+0.9%/year) are increasing. Incidence of mesothelioma (1594 cases), showed for the period 1986-1997, a statistically significant increase among both males and females; standardised rates increased, more than 4% every year. As regards to deaths due to pleural malignant cancers (1393 among males and 664 among females) their trend was stable in the analysed period. PMID- 15281607 TI - Skin cancers: melanoma, non-melanoma cancers and Kaposi's sarcoma. AB - The present paper analyses temporal trends for skin cancers including Kaposi's sarcoma in the data set of the Italian Network of Cancer Registries (pool AIRT), during 1986-1997. The estimated annual percent change (EAPC) of the standardised incidence rates for non-melanoma skin cancer (61,586 analysed cases), increased from 1986 to 1997 by 7.5%/year among males and by 5.2%/year among females. Melanoma (8,516 cases and 2,312 deaths) showed increasing incidence, 6.2%/year among males and 5.8%/year among females, and stable mortality. As regards Kaposis sarcoma (1,156 cases), incidence increased among males from 1986 to 1995 by about 13%/year, and then decreased; among females incidence was significantly increasing along the period. PMID- 15281608 TI - Female breast cancer. AB - Breast cancer represents, among females, the most important cancer site, in terms of incidence and mortality. Temporal trends have been computed in the database of the Italian Network of Cancer Registries (pool AIRT) including 58,862 cases and 20,659 deaths during the period 1986-1997. Incidence was increasing and standardised rates increase by, on average, 1.7% every year. Such increase may be partially due to the implementation of organised screening programmes. Mortality was increasing up to 1989, then it statistically significantly decreased. PMID- 15281609 TI - Gynecological cancers: cervix, corpus uteri, ovary. AB - Incidence trend of cervix uteri cancer (5,158 cases), within the Italian Network of Cancer Registries database (pool AIRT), showed for the period 1986-1997 a statistically significant decrease up to 1990. Corpus uteri cancer (9,975 incident cases) incidence rates were stable in the analysed period. As regards uteri cancer mortality, only a small proportion of deaths are coded as cervix (784 deaths) or corpus uteri (886 deaths) whereas the great majority (3,538 deaths) refers to uteri not otherwise specified; mortality for the latter group was significantly decreasing in the analysed period. Ovary cancer showed stable incidence (7,690 incident cases) and mortality (5.040 deaths) rates in the period 1986-1997. PMID- 15281610 TI - Cancers of the male genital organs: prostate and testis. AB - The present paper presents the temporal trend analysis of prostate and testis cancers within the Italian Network of Cancer Registries (pool AIRT) for the period 1986-1997. As regards prostate cancer (27,477 incident cases and 11,189 deaths analysed), incidence showed a strong increasing trend, particularly evident in the period 1991-1994, during which standardised rates increased on an average of 12.1% every year. Mortality showed a slight, but significant, decreasing trend. Testis cancer (1,808 incident cases and 185 deaths) incidence increased significantly from 1986 to 1997 and mortality was stable. PMID- 15281611 TI - Urinary tract cancers: kidney and urinary bladder. AB - The present paper analyses temporal trends for cancers of the kidney including urinary tract and of the urinary bladder, in the Italian Network of Cancer Registries (pool AIRT), for the period 1986-1997. As regards kidney cancer (14,915 cases and 5,857 deaths analysed) a statistically significant increase has been observed in incidence rates. Such increase was present from 1986 to 1991 among males, +5.4%/year, afterwards it became rather stable; among females, the incidence increased along the whole period, +2. 7%/year. Mortality rates were stable along the analysed time period. Bladder cancer incidence (33,283 cases) showed a significant increase in both sexes, + 1. 6%/year among males and +2.5% among females, whereas mortality (9,348 deaths) showed a significant decreasing trend in both males, -2.7%/year, and females, -3.5%/year. PMID- 15281612 TI - Thyroid cancer. AB - Temporal trend for incident cases (n = 5,101) and deaths (n = 1,130) for thyroid cancer in the database of the Italian Network of Cancer Registries (pool AIRT) in the period 1986-1997, showed a statistically significant increase of incidence rates in both males (EAPC = +3.4%) and females (EAPC = +4.6%). Mortality rates showed a statistically significant decrease, for both sexes, of about 4%/year. PMID- 15281613 TI - Lymphomas: Hodgkin and non-Hodgkin lymphomas. AB - Results on lymphomas trend analysis carried out in the database of the Italian Network of Cancer Registries (pool AIRT) in the period 1986-1997 are presented. Hodgkin lymphoma (2,959 cases and 911 deaths included in the analysis) shows a decreasing incidence trend, statistically significant among males. Standardised mortality rates decrease significantly in both sexes, on an average of 7-9%/year. As regards non-Hodgkin lymphomas (16, 470 incident cases and 7,193 deaths), there is a significant increasing trend, in both sexes, both for incidence (more than 3 %/years) and mortality (about 2%/year). PMID- 15281614 TI - Leukemias and multiple myeloma. AB - The present paper analyses temporal trends for leukemias in adulthood and for multiple myeloma in the database of the Italian Network of Cancer Registries (pool AIRT) during 1986-1997. As regards leukemias, in subjects aged 15+, (10,946 analysed cases) the incidence rates trend was rather stable during the period 1986 to 1997, whereas mortality (8,265 deaths included) showed a decrease of about 1-2%/year, statistically significant among males. Multiple myeloma, 6,535 cases and 4,310 deaths, showed a sharp increase in incidence rates that grew in the period at apace of about 3 %/year; mortality was stable. PMID- 15281615 TI - The role of primary prevention. PMID- 15281616 TI - Determination of pantoprazole in human plasma by LC-MS-MS using lansoprazole as internal standard. AB - An analytical method based on liquid chromatography with positive ion electrospray ionization (ESI) coupled to tandem mass spectrometry detection was developed for the determination of pantoprazole (CAS 102625-70-7) in human plasma using lansoprazole (CAS 103577-45-3) as the internal standard. The analyte and internal standard were extracted from the plasma samples by liquid/liquid extraction using diethyl-ether/dichloromethane (70:30; v/v) and chromatographed on a C8 analytical column. The mobile phase consisted of acetonitrile/ water/methanol (57:25:18; v/v/v) + 10 mmol/l acetic acid + 20 mmol/l ammonium acetate. The method has a chromatographic total run time of 4.5 min and was linear within the range 5.0-5,000 ng/ mL. Detection was performed on a triple quadrupole tandem mass spectrometer by Multiple Reaction Monitoring (MRM). The intra- and inter-run precisions calculated from quality control (QC) samples were 4.2 % and 3.2 %, respectively. The accuracies as determined from QC samples were 5.0 % (intra-run) and 2.0 % (inter-run). The method herein described was employed in a bioequivalence study of two tablet formulations of pantoprazole. PMID- 15281617 TI - Ceftazidime determination in serum by high-pressure liquid chromatography. AB - A rapid and sensitive high-pressure liquid chromatographic method with simple sample preparation was developed for the quantitative analysis of the beta-lactam antibiotic ceftazidime (CAS 78439-06-2, Fortum). A good linear relationship was established between the peak area and the amount of ceftazidime injected over a concentration range of 1 to 200 microg/ml. The detection limit of the method was calculated to be 0.9 microg/ml. Stability was shown at 4 degrees C and at -196 degrees C for time periods of 2 h and 84 days, respectively. PMID- 15281618 TI - Complexes of ruthenium(III) with some 2-aminothiazole derivatives/synthesis, properties and pharmacological studies. AB - Four new complexes of Ru(III) with a general formula [Ru(L)2Cl2]Cl, where L = 2 amino-4-phenylthiazole (CAS 2010-06-2), 2-amino-4-methylthiazole (CAS 1603-91-4), ethyl 2-amino-4-methyl-5-thiazolecarboxylate (CAS 7210-76-6) and ethyl 2-amino-4 phenyl-5-thiazolecarboxylate (CAS 64399-23-1), were prepared. The syntheses were carried out in polar medium and inert atmosphere at a molar ratio Ru:L = 1:2 or 1:3. The compounds obtained were characterised by IR-, 1H-NMR- 13C-NMR-, UV-VIS-, EPR spectroscopy, magnetochemical and conductivity measurements. The ligands behaved as bidental, bounding Ru(III) through the nitrogen atoms from the amino group and the heterocycle. The complex of ethyl 2-amino-4-phenyl-5 thiazolecarboxylate showed significant antileukaemic activity on various human cells (IC50 values ranging from 20 to 92 micromol/l). Toxicological studies on mice indicated that such concentrations could be reached without mortality. This compound exhibited a promising antineoplastic potential and needs further pharmacological and toxicological evaluation. PMID- 15281619 TI - In vivo toxicity and antithrombotic profile of the oral formulation of the antileukemic agent, LFM-A13-F. AB - The specific inhibitor of the protein tyrosine kinase, Bruton's tyrosine kinase (BTK), alpha-cyano-beta-hydroxy-beta-methyl-N-(2,5-dibromophenyl)-propenamide (LFM-A13, CAS 244240-24-2), is a chemosensitizing antileukemic agent with antithrombotic properties. Oral formulation of LFM-A13 (LFM-A13-F) did not cause acute, subacute or chronic toxicity in mice at dose levels up to 200 mg/kg. The in vivo antithrombotic activity of LFM-A13 was studied in a mouse model of collagen-induced fatal thromboembolism. Oral doses of LFM-A13-F dose dependently prevented collagen-induced thromboembolism in mice without causing bleeding. LFM A13 could be combined with dipyridamole (CAS 58-32-2) without side effects. These results indicate that LFM-A13 may be particularly useful in the treatment of leukemia patients who are at risk for thromboembolic complications. PMID- 15281620 TI - Biodistribution and pharmacokinetics of variously molecular sized 117mSn(II) polyethyleneiminomethyl phosphonate complexes in the normal primate model as potential selective therapeutic bone agents. AB - In the search for a cure for metastatic bone cancer, 117mSn with its conversion electrons and low energy photons both of discrete energies shows little bone marrow toxicity, providing the opportunity to increase the administered dose. Selective accumulation in lesions would capitalise on this advantage. The 10-30 kDa fraction of the water-soluble polymer polyethyleneimine, functionalised with methyl phosphonate groups (PEI-MP) and labelled with 99mTc, has shown selective uptake into bone tumours. Furthermore using speciation calculations it was predicted that the Sn(II)-PEI-MP complex would remain intact in the blood plasma. Because of this positive indication animal experiments were carried out to test this prediction. This paper relates the labelling, biodistribution and pharmacokinetics of various fractions of 117mSn-(II) PEI-MP in the normal primate model, and points to promising therapeutic possibilities. PMID- 15281622 TI - [Pediatric research is necessary. The children must be guaranteed with a complete protection]. PMID- 15281621 TI - Three-dimensional model of nonsteroidal estrogen receptor ligand binding/electron topological method. AB - Three-dimensional quantitative structure-activity relationship (3D-QSAR) models have been obtained using the electron topological method (ETM) for estrogen (estrone, CAS 53-16-7) receptor (ER) affinities. This method was used to identify regions of the enzyme and inhibitors where steric and electrostatic effects correlate strongly with biological activity. Thirty compounds belong to a series of diethylstilbestrol (CAS 56-53-1, DES) and indenestrol (CAS 24643-94-5) analogues whose affinities for ER have been, theoretically, investigated. After energy minimizations and molecular dynamics calculations were performed to find the ground state conformer for each molecule, quantum chemical properties of each molecule were defined and in ETM their matrices were compared with those of the reference molecule. It is shown that the molecular fragments responsible for this affinity possess fixed electronic and geometric characteristics associated with a distinct arrangement and steric accessibility of an oxygen atom and a group of carbon atoms. Several parameters were modified in order to analyze their influence onto the correlation between binding affinities. The highest correlation coefficient (R2 = 0.930, SE = 0.403) was obtained with the structure of the active fragment and the orientation of aliphatic and phenolic substituents. PMID- 15281623 TI - [Golden ages of the Swedish geriatrics during the 20th century. From the care of chronically ill to a modern specialty--and an international example]. PMID- 15281624 TI - [Survey at the Poison Control Center for the year 2000. Poisoning accidents are common among children but mostly not dangerous]. PMID- 15281625 TI - [A national curriculum in neurology for medical education]. AB - The knowledge within medicine is growing rapidly. It has become more and more difficult to decide what knowledge that has to be taught to medical students during their University Medical Degree (MD) education and what has to be omitted from their study plans. As help for teachers and students, a core curriculum of the education defines what is of importance for all students. However, there is a risk of "curriculum overload" with too much information being put into a short time interval. To avoid this and to define what is really the "core" of a course, a national consensus decision may be useful. As an example of this approach, we here report a new joint Swedish core curriculum in neurology for medical students. Teachers responsible for neurological education at all six universities giving University MD education in Sweden have in January 2003 agreed upon the core curriculum that we present. It is our hope that this method can be useful also for other clinical specialities. PMID- 15281626 TI - [Hormone replacement therapy in the climacteric--from hallelujah to requiem]. PMID- 15281627 TI - [Confusion in somatic hospital departments]. PMID- 15281628 TI - [Extensive initiatives for improved patient safety in the USA]. PMID- 15281629 TI - [An African flower with a Swedish name]. PMID- 15281630 TI - [Are combat sports and medical ethics compatible?]. PMID- 15281631 TI - [Nicholas Culpeter--the 17th century pharmacist who infuriated the medical profession]. PMID- 15281632 TI - [Proposals on the innovation policy by Ostros--correct diagnosis but wrong treatment]. PMID- 15281633 TI - [Ionized calcium is a better basis for decisions than albumin-corrected calcium]. PMID- 15281634 TI - [Arthroscopy with local anesthesia a reality now?]. PMID- 15281635 TI - [Misunderstandings about plant preparations]. PMID- 15281636 TI - [Unfortunate title?]. PMID- 15281637 TI - [Can the physician's driving licence report be of advantage to the patient?]. PMID- 15281638 TI - [The Swedish paradox--a rehabilitation for the medical profession!]. PMID- 15281639 TI - [We are all mortals]. PMID- 15281640 TI - [The issue of bicycle helmet is nt only the traveller's concern]. PMID- 15281641 TI - [Can psychological models explain why good car seats reduce the risk of neck injuries?]. PMID- 15281642 TI - [Continuing education is always the employer's responsibility]. PMID- 15281643 TI - [An explanation concerning the certification of the tampon Ellen]. PMID- 15281644 TI - The veterinary community. PMID- 15281645 TI - Requests more information on intraluminal solution. PMID- 15281646 TI - Comparative assessment of bone mineral measurements obtained by use of dual energy x-ray absorptiometry, peripheral quantitative computed tomography, and chemical-physical analyses in femurs of juvenile and adult dogs. AB - OBJECTIVE: To compare bone mineral measurements obtained by use of dual-energy x ray absorptiometry (DEXA), peripheral quantitative computed tomography (pQCT), and chemical-physical analyses and determine effects of age and femur size on values obtained for the various techniques. SAMPLE POPULATION: Femurs obtained from 15 juvenile and 15 adult large-breed dogs. PROCEDURE: n each femur, 7 regions of interest were examined by use of DEXA to measure the bone mineral content (BMC) and bone mineral density (BMD), and 5 were examined by use of pQCT to measure BMD. Among these, 1 region was examined by both noninvasive methods and an invasive method. Volume of the femur was determined by water displacement. Volumetric bone density (VBD) was calculated. Calcium (Ca), phosphorus (P), total Ca, and total P contents were determined. RESULTS: DEXA- and pQCT-derived results revealed that all values increased with age in juvenile dogs. In adults, VBD and pQCT-derived BMD decreased significantly and DEXA-derived BMD increased with increasing femur length. The pQCT-derived BMD correlated well with VBD and Ca content, whereas DEXA-derived BMC was strongly correlated with Ca content. In juveniles, values correlated regardless of the technique used, whereas in adult dogs, DEXA-derived BMD did not correlate with pQCT-derived BMD, Ca concentration, or VBD unless data were adjusted on the basis of femur length. CONCLUSIONS AND CLINICAL RELEVANCE: DEXA-derived BMD adjusted for femur length yields approximately the same percentage variability in VBD as for pQCT-derived BMD. However, pQCT-derived BMD is still more sensitive for determining variability BMD in Ca concentration, compared with DEXA-derived BMD adjusted for femur length. PMID- 15281647 TI - Investigation of the facilitation of the nociceptive withdrawal reflex evoked by repeated transcutaneous electrical stimulations as a measure of temporal summation in conscious horses. AB - OBJECTIVE: To investigate whether facilitation of the nociceptive withdrawal reflex (NWR) can be evoked and quantified as a measure of temporal summation from the distal aspect of the left forelimb and hind limb in standing nonsedated horses via repeated stimulations of various subthreshold intensities and frequencies. ANIMALS: 10 adult horses. PROCEDURE: Surface electromyographic activity evoked by stimulation of the digital palmar and plantar nerves was recorded from the common digital extensor and cranial tibial muscles. For each horse, the NWR threshold intensity to a single stimulus was determined for the forelimb and hind limb. Repeated stimulations were performed at subthreshold intensities and at frequencies of 2, 5, and 10 Hz. The reflex amplitude was quantified, and the behavioral responses accompanying the stimulations were scored. RESULTS: Repeated stimulations at subthreshold intensities were able to summate and facilitate the NWR in conscious horses. The reflex facilitation was significantly related to the intensity of the repeated stimuli, whereas no effect of stimulation frequency was found. Reaction scores increased significantly for increasing stimulation intensities. CONCLUSIONS AND CLINICAL RELEVANCE: Temporal summation obtained by repeated stimulations of subthreshold intensity appears to represent a new tool for investigating nociceptive pathophysiologic processes in horses; this experimental model may be useful to examine the mode of action and efficacy of analgesic and anesthetic interventions and possibly to assess sensory dysfunction in clinical settings. PMID- 15281648 TI - Assessment of repeatability, reproducibility, and effect of anesthesia on determination of radial and longitudinal left ventricular velocities via tissue Doppler imaging in dogs. AB - OBJECTIVE: To determine left ventricular free wall (LVFW) motions and assess their intra- and interday variability via tissue Doppler imaging (TDI) in healthy awake and anesthetized dogs. ANIMALS: 6 healthy adult Beagles. PROCEDURE: n the first part of the study, 72 TDI examinations (36 radial and 36 longitudinal) were performed by the same observer on 4 days during a 2-week period in all dogs. In the second part, 3 dogs were anesthetized with isoflurane and vecuronium. Two measurements of each TDI parameter were made on 2 consecutive cardiac cycles when ventilation was transiently stopped. The TDI parameters included maximal systolic, early, and late diastolic LVFW velocities. RESULTS: The LVFW velocities were significantly higher in the endocardial than in the epicardial layers and also significantly higher in the basal than in the mid-segments in systole, late diastole, and early diastole. The intraday coefficients of variation (CVs) for systole were 16.4% and 22%, and the interday CV values were 11.2% and 16.4% in the endocardial and epicardial layers, respectively. Isoflurane anesthesia significantly improved the intraday CV but induced a decrease in LVFW velocities, except late diastolic in endocardial layers and early diastolic in epicardial layers. CONCLUSIONS AND CLINICAL RELEVANCE: Left ventricular motion can be adequately quantified in dogs and can provide new noninvasive indices of myocardial function. General anesthesia improved repeatability of the procedure but cannot be recommended because it induces a decrease in myocardial velocities. PMID- 15281649 TI - Effects of feeding meals with various soluble-carbohydrate content on muscle glycogen synthesis after exercise in horses. AB - OBJECTIVES: To determine effects of feeding diets with various soluble carbohydrate (CHO) content on rates of muscle glycogen synthesis after exercise in horses. ANIMALS: 7 fit horses. PROCEDURES: In a 3-way crossover study, horses received each of 3 isocaloric diets (a high soluble CHO [HC] diet, a low soluble CHO [LC] diet, or a mixed soluble CHO [MC] diet). For each diet, horses were subjected to glycogen-depleting exercise, followed by feeding of the HC, LC, or MC diet at 8-hour intervals for 72 hours. RESULTS: Feeding the HC diet resulted in a significantly higher glycemic response for 72 hours and significantly greater muscle glycogen concentration at 48 and 72 hours after exercise, compared with results after feeding the MC and LC diets. Muscle glycogen concentrations similar to baseline concentrations were detected in samples obtained 72 hours after exercise in horses when fed the HC diet. Rate of glycogen synthesis was significantly higher when horses were fed the HC diet, compared with values when horses were fed the MC and LC diets. Glycogen synthase activity was inversely related to glycogen content. Protein content of glucose transporter-4 was the lowest at 72 hours after exercise when horses were fed the HC diet. CONCLUSIONS AND CLINICAL RELEVANCE: Muscle glycogen synthesis was slower after glycogen depleting exercise in horses, compared with synthesis in humans. Feeding HC meals after strenuous exercise hastened replenishment of muscle glycogen content, compared with results for feeding of LC and MC diets, by increasing availability of blood glucose to skeletal muscles. PMID- 15281650 TI - Comparison of hematologic values and transforming growth factor-beta and insulin like growth factor concentrations in platelet concentrates obtained by use of buffy coat and apheresis methods from equine blood. AB - OBJECTIVE: To evaluate the buffy coat and apheresis methods for preparation of platelet concentrates from equine blood by comparing platelet and growth factor concentrations. ANIMALS: 15 mature mixed-breed geldings. PROCEDURE: Whole blood samples were collected and processed by use of a buffy coat or apheresis method to obtain platelet poor and platelet concentrated fractions. The PCV, WBC count, and platelet count were compared among whole blood samples, platelet poor fractions, concentrates obtained by use of the apheresis method (ie, apheresis platelet concentrates), and concentrates obtained by use of the buffy coat method (ie, buffy coat platelet concentrates). Concentrations of transforming growth factor-beta (ie,TGF-beta1 andTGF-beta2) and insulin-like growth factor were compared between buffy coat and apheresis platelet concentrates. RESULTS: Platelet concentrations were 8.9-fold and 5.2-fold greater in buffy coat and apheresis platelet concentrates, respectively, compared with whole blood. Platelet concentrations were 13.1-fold greater in filtered apheresis platelet concentrates, compared with whole blood. TGF-beta1 concentrations were 2.8-fold and 3.1-fold greater in buffy coat and apheresis platelet concentrates, respectively, and TGF-beta1 concentrations were 10.5-fold greater in filtered apheresis platelet concentrates, compared with whole blood. TGF-beta2 concentrations were 3.6-fold greater in apheresis platelet concentrates, compared with whole blood. Platelet concentrations correlated with growth factor concentrations across all blood and platelet fractions. White blood cell counts had a significant positive correlation with TGF-beta1 concentration in buffy coat platelet concentrates. CONCLUSIONS AND CLINICAL RELEVANCE: Platelets and TGF beta1 can be concentrated reliably from equine blood by use of buffy coat or apheresis methods, without modification of the protocols used for humans. PMID- 15281651 TI - Comparative cardiovascular, analgesic, and sedative effects of medetomidine, medetomidine-hydromorphone, and medetomidine-butorphanol in dogs. AB - OBJECTIVE: To compare sedative, analgesic, and cardiopulmonary effects after IV administration of medetomidine (20 microg/kg), medetomidine-hydromorphone (20 microg of medetomidine/kg and 0.1 mg of hydromorphone/kg), and medetomidine butorphanol (20 microg of medetomidine/kg and 0.2 mg of butorphanol tartrate/kg) in dogs. ANIMALS: 6 dogs healthy mixed-breed dogs. PROCEDURE: Instruments were surgically inserted, and heart rate (HR), respiratory rate (RR), systolic arterial pressure (SAP), mean arterial pressure (MAP), diastolic arterial pressure (DAP), mean pulmonary arterial pressure (MPAP), pulmonary capillary wedge pressure (PCWP), central venous pressure (CVP), core body temperature, and cardiac output (CO) were measured 0, 5, 10, 15, 30, 45, and 60 minutes after injection. Cardiac index (CI), stroke volume (SV), stroke index (SI), systemic vascular resistance (SVR), and pulmonary vascular resistance (PVR) were calculated. Arterial samples for blood gas analysis were collected 0, 15, and 45 minutes after injection. Intensity of analgesia, degree of sedation, and degree of muscle relaxation were evaluated at aforementioned time points and 75, 90, 120, 150, 180, and 210 minutes after injection. RESULTS: Administration of medetomidine, medetomidine-hydromorphone, and medetomidine-butorphanol was associated with increases in SAP, MAP, DAP, MPAP, PCWP, CVP, SVR, PVR, core body temperature, and PaCO2 and decreases in HR, CO, CI, SV, SI, RR, pH, and PaO2. Clinically important differences were not detected among treatments. Medetomidine hydromorphone and medetomidine-butorphanol provided a longer duration of sedation and better quality of analgesia, compared with medetomidine alone. CONCLUSIONS AND CLINICAL RELEVANCE: Medetomidine-hydromorphone or medetomidine-butorphanol is associated with improved analgesia and sedation but has cardiopulmonary effects comparable to those for medetomidine alone. PMID- 15281652 TI - Variation in free jumping technique within and among horses with little experience in show jumping. AB - OBJECTIVE: To quantify variation in the jumping technique within and among young horses with little jumping experience, establish relationships between kinetic and kinematic variables, and identify a limited set of variables characteristic for detecting differences in jumping performance among horses. ANIMALS: Fifteen 4 year-old Dutch Warmblood horses. PROCEDURE: The horses were raised under standardized conditions and trained in accordance with a fixed protocol for a short period. Subsequently, horses were analyzed kinematically during free jumping over a fence with a height of 1.05 m. RESULTS: Within-horse variation in all variables that quantified jumping technique was smaller than variation among horses. However, some horses had less variation than others. Height of the center of gravity (CG) at the apex of the jump ranged from 1.80 to 2.01 m among horses; this variation could be explained by the variation in vertical velocity of the CG at takeoff (r, 0.78). Horses that had higher vertical velocity at takeoff left the ground and landed again farther from the fence, had shorter push-off phases for the forelimbs and hind limbs, and generated greater vertical acceleration of the CG primarily during the hind limb push-off. However, all horses cleared the fence successfully, independent of jumping technique. CONCLUSIONS AND CLINICAL RELEVANCE: Each horse had its own jumping technique. Differences among techniques were characterized by variations in the vertical velocity of the CG at takeoff. It must be determined whether jumping performance later in life can be predicted from observing free jumps of young horses. PMID- 15281653 TI - Evaluation of consistency of jumping technique in horses between the ages of 6 months and 4 years. AB - OBJECTIVE: To determine whether differences in jumping technique among horses are consistent at various ages. ANIMALS: 12 Dutch Warmblood horses. PROCEDURE: Kinematics were recorded during free jumps of horses when they were 6 months old (ie, no jumping experience) and 4 years old (ie, the horses had started their training period to become show jumpers). Mean +/- SD height of the horses was 1.40 +/- 0.04 m at 6 months of age and 1.70 +/- 0.05 m at 4 years of age. RESULTS: Strong correlations were found between values from 6-month-old foals and 4-year-old horses for variables such as peak vertical acceleration generated by the hind limbs (r, 0.91), peak rate of change of effective energy generated by the hind limbs (r, 0.71), vertical velocity at takeoff (r, 0.65), vertical displacement of the center of gravity during the airborne phase (r, 0.81), and duration of the airborne phase (r, 0.70). CONCLUSIONS AND CLINICAL RELEVANCE: Although there are substantial anatomic and behavioral changes during the growing period, certain characteristics of jumping technique observed in naive 4-year olds are already detectable when those horses are foals. PMID- 15281654 TI - Investigation of the expression and localization of glucose transporter 4 and fatty acid translocase/CD36 in equine skeletal muscle. AB - OBJECTIVE: To investigate the expression and localization of glucose transporter 4 (GLUT4) and fatty acid translocase (FAT/CD36) in equine skeletal muscle. SAMPLE POPULATION: Muscle biopsy specimens obtained from 5 healthy Dutch Warmblood horses. PROCEDURES: Percutaneous biopsy specimens were obtained from the vastus lateralis, pectoralis descendens, and triceps brachii muscles. Cryosections were stained with combinations of GLUT4 and myosin heavy chain (MHC) specific antibodies or FAT/CD36 and MHC antibodies to assess the fiber specific expression of GLUT4 and FAT/CD36 in equine skeletal muscle via indirect immunofluorescent microscopy. RESULTS: Immunofluorescent staining revealed that GLUT4 was predominantly expressed in the cytosol of fast type 2B fibers of equine skeletal muscle, although several type 1 fibers in the vastus lateralis muscle were positive for GLUT4. In all muscle fibers examined microscopically, FAT/CD36 was strongly expressed in the sarcolemma and capillaries. Type 1 muscle fibers also expressed small intracellular amounts of FAT/CD36, but no intracellular FAT/CD36 expression was detected in type 2 fibers. CONCLUSIONS AND CLINICAL RELEVANCE: In equine skeletal muscle, GLUT4 and FAT/CD36 are expressed in a fiber type selective manner. PMID- 15281656 TI - Diagnostic relevance of qualitative proteinuria evaluated by use of sodium dodecyl sulfate-agarose gel electrophoresis and comparison with renal histologic findings in dogs. AB - OBJECTIVE: To evaluate results of SDS-agarose gel electrophoresis (AGE) of urinary proteins for use in defining glomerular and tubulointerstitial derangements, investigate patterns of high-molecular-weight (HMW) proteins for differentiating among glomerular disorders, and assess low-molecular-weight (LMW) proteins as markers of severity of tubulointerstitial disease in dogs. ANIMALS: 49 dogs with increased serum creatinine concentrations or abnormal renal protein loss. PROCEDURE: Urinary proteins were examined by use of SDS-AGE and differentiated on the basis of molecular weight. The HMW proteins (> or = 69 kd) were considered indicative of glomerular origin, whereas LMW proteins (< 69 kd) were of tubular origin. Renal specimens were examined by use of light microscopy. Glomerular and tubulointerstitial lesions were differentiated by use of the classification for the World Health Organization and semiquantitative grading, respectively. RESULTS: Sensitivity of SDS-AGE was 100% for detection of glomerular lesions and 92.6% for tubulointerstitial lesions; specificity was 40% and 62.5%, respectively. Although HMW urinary proteins were not significantly associated with the type of glomerular lesion, LMW urinary proteins were significantly associated with the grade of tubulointerstitial damage. Detection of 12- or 15-kd proteins or both was highly indicative of a severe tubulointerstitial lesion. CONCLUSIONS AND CLINICAL RELEVANCE: SDS-AGE of urinary proteins in dogs represents a noninvasive test with high sensitivity for identifying glomerular and tubulointerstitial damage, but low specificity limits its validity as a stand-alone test to differentiate between glomerular and tubulointerstitial lesions. The test is particularly useful for identifying dogs with advanced tubulointerstitial disease but cannot be used to characterize glomerular disorders. PMID- 15281655 TI - Characterization of biological activities of feline eosinophil granule proteins. AB - OBJECTIVE: To characterize eosinophil granule-derived proteins in cats. SAMPLE POPULATION: Eosinophils collected via peritoneal lavage from 2 cats. PROCEDURE: The cats were infested orally with Toxocara canis eggs and subsequently challenge exposed with T. canis antigen injected IP to induce peritoneal eosinophilia; eosinophils were collected via peritoneal lavage. Eosinophil granule proteins were acid-extracted, separated by gel-filtration chromatography, and examined for their peroxidase, ribonuclease, and bactericidal activities; the N-terminal sequence of some of these proteins was determined and compared with homologue proteins from other species. RESULTS: 3 protein peaks were separated in the chromatogram. The first peak had both peroxidase and bactericidal activities. The second peak had ribonuclease and bactericidal activities, and the N-terminal sequence of the major protein was homologous with that of proteins of the ribonuclease A superfamily, including eosinophil ribonucleases from humans and other animal species. The third protein peak had bactericidal activity, and the N terminal sequence of the major protein was homologous with that of human and murine major basic proteins. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicated that feline eosinophil granules contain major basic protein and eosinophil-associated ribonuclease and the granule proteins have peroxidase, ribonuclease, and bactericidal activities. In cats, characterization of eosinophil granule proteins may be useful in elucidation of the mechanism of tissue damage in eosinophil-associated diseases and development of new treatment options for those diseases. In addition, the identification of conserved structure and function of eosinophil granule proteins in cats is relevant from an evolutionary viewpoint. PMID- 15281657 TI - Effect of laser shock peening on fatigue life and surface characteristics of stainless steel cortical bone screws. AB - OBJECTIVE: To investigate the effect of laser shock peening on the fatigue life and surface characteristics of 3.5-mm-diameter cortical bone screws. SAMPLE POPULATION: 32 stainless steel, 3.5-mm-diameter cortical bone screws. PROCEDURE: Screws were randomly assigned to an untreated control group or 2 power-density treatment groups, 6 gigawatts (GW)/cm2 and 8.5 GW/cm2, for laser shock peening. Number of cycles to failure and findings on scanning electron microscopy-assisted morphometric evaluation, including the mode of failure, surface debris, surface damage, and thread deformation, were compared between control and treated screws. RESULTS: The 6 GW/cm2 treated screws had a significant (11%) improvement in fatigue life, compared with untreated control screws. The 8.5 GW/cm2 treated screws had a significant (20%) decrease in fatigue life, compared with control screws. A mild but significant increase in thread deformation was evident in all treated screws, compared with control screws. The 8.5 GW/cm2 treated screws had significantly more surface irregularities (elevations and pits), compared with control or 6 GW/cm2 treated screws. CONCLUSION AND CLINICAL RELEVANCE: A modest positive increase in fatigue strength was produced by this design of laser shock peening on the midshaft of cortical bone screws. High laser shock peening power densities were detrimental, decreasing screw fatigue strength probably resulting from structural damage. Greater fatigue life of cortical bone screws can be generated with laser shock peening and could reduce screw breakage as a cause of implant failure; however, future studies will be necessary to address biocompatibility, alternative cleaning techniques, alterations in screw strength and pullout characteristics, and effects on susceptibility to corrosion. PMID- 15281658 TI - Stability, antigenicity, and aggregation of Moraxella bovis cytolysin after purification and storage. AB - OBJECTIVES: To compare stability, antigenicity, and aggregation characteristics of Moraxella bovis cytolysins among isolates from geographically diverse areas. STUDY POPULATION: 8 isolates of M. bovis. PROCEDURE: Filter-sterilized broth culture supernatants of M. bovis were concentrated, diafiltered, and chromatographed. The endotoxin and cytolysin activities in samples were measured. Chromatographed cytolysins of M. bovis were examined by immunoblotting. Hemolytic and leukotoxic activities were measured from samples collected at each step of purification and before and after storage. Hemolysis was measured directly by use of washed bovine erythrocyte targets. Leukotoxicity was measured by use of a 51Cr release assay. RESULTS: Cytolysin was retained by a filter with 100-kd nominal molecular weight limit. Hemolytic activity, leukotoxic activity, and endotoxin were eluted together in void volume of a gel-filtration column (molecular mass exclusion limit = 4 X 10(7) d). Gel-column chromatographed diafiltered retentate had the greatest specific cytolytic activity and the highest endotoxin-to-protein ratio. Frozen diafiltered retentate(-80 degrees C, 4 months) was cytolytic after thawing. Immunoblots of gel-column chromatographed cytolysin contained 4 proteins with molecular masses between 90 and 68 kd. Fractions with high lytic activities also had additional protein bands with molecular masses of 98 and 63 kd. Immunoblots of gel-column chromatographed diafiltered retentate revealed proteins with molecular masses between 90 and 68 kd. CONCLUSIONS AND CLINICAL RELEVANCE: Diafiltered M. bovis cytolysin is aggregated with endotoxin. Antigenicity and cytolytic activities in diafiltered retentate are conserved among M. bovis isolates. Diafiltration could be useful for bulk semipurification of M. bovis cytolysin. Cytolysin-enriched vaccines of M. bovis could be contaminated by endotoxin. PMID- 15281659 TI - Pharmacologic evaluation of neurokinin-2 receptor antagonists in the guinea pig respiratory tract. AB - OBJECTIVE: To evaluate 3 neurokinin-2 (NK2) receptor antagonists on the basis of their ability to block neurokinin A (NKA)-induced contractile responses in various regions of the guinea pig respiratory tract. ANIMALS: 48 clinically normal guinea pigs. PROCEDURE: After euthanasia, the trachea and lungs were removed en bloc. The spirally cut trachea was divided into lower, middle, and upper portions. The main bronchus was spirally cut. A lung strip was cut from the edge of the lung. Tissue strips were mounted in organ baths containing Tyrode solution at 37 degrees C and attached to force transducers interfaced with a polygraph. Lung strips were set at a tension of 1 g; other tissue strips were set at 2 g. After 45 minutes of equilibration, cumulative concentration-response (CR) relationships to graded concentrations of NKA were determined. In the treatment groups, tissues were incubated (30 minutes) with antagonists (MEN 10376, SR 48968, and SR 144190) at 3 concentrations (10(-9), 10(-7), and 10(-5)M) before CR relationships were determined. Effectiveness of SR 48968 against NKA was also tested in vivo. RESULTS: Lung strips failed to contract, but all others responded in a concentration-dependent manner. Bronchial spirals were most sensitive. SR 48968 had the highest pA2 value and effectively blocked NKA. CONCLUSIONS AND CLINICAL RELEVANCE: The bronchial region where airflow resistance is high was the most sensitive to NKA, suggesting the importance of NKA in bronchoconstriction. Nonpeptide antagonists (SR 48968 and SR 144190) were more potent than the peptide antagonist (MEN 10376), indicating their greater therapeutic potential as antiasthmatic agents. PMID- 15281660 TI - Pharmacokinetics of amoxicillin after oral administration in recently weaned piglets with experimentally induced Escherichia coli subtype O149:F4 diarrhea. AB - OBJECTIVE: To measure the effect of Escherichia coli subtype O149:F4-induced diarrhea on the pharmacokinetics of orally administered amoxicillin in affected piglets relative to that of uninfected piglets. ANIMALS: 22 healthy 4-week-old recently weaned Danish crossbred piglets. PROCEDURE: 12 piglets were orally inoculated through gastric intubation with 10(9) CFUs of an E. coli O149:F4 strain and responded by developing diarrhea 12 to 16 hours later. Piglets were dosed with amoxicillin trihydrate solution (20 mg/kg) by gastric intubation. A control group of 10 age-matched piglets without signs of diarrhea was dosed similarly. Blood samples were obtained before amoxicillin administration and at 0.5, 1, 1.5, 2, 3, 6, 12, and 24 hours after amoxicillin administration. The plasma concentration of amoxicillin was analyzed by high-performance liquid chromatography. RESULTS: A significant 39% decrease in the area under the plasma concentration versus time curve of amoxicillin was observed in piglets with diarrhea relative to that of control piglets. The maximum plasma concentration (Cmax) was significantly (52%) lower in piglets with diarrhea, compared with control piglets, while the elimination rate constant, time to reach Cmax, and elimination half-life were unchanged. CONCLUSIONS AND CLINICAL RELEVANCE: Escherichia coli-induced diarrhea may decrease systemic bioavailability of amoxicillin. Escherichia coli bacteria attach to the intestinal epithelial cells. Because it is assumed that the concentration of the antimicrobial at the site of infection reflects the systemic concentration, higher doses of amoxicillin in the treatment of piglets with E. coli O149:F4-induced diarrhea may be appropriate. PMID- 15281661 TI - Assessment of the effects of feed restriction and amino acid supplementation on glucose tolerance in llamas. AB - OBJECTIVE: To assess the effects of prolonged feed deprivation on glucose tolerance, insulin secretion, and lipid homeostasis in llamas. ANIMALS: 9 adult female llamas. PROCEDURE: On each of 2 consecutive days, food was withheld from the llamas for 8 hours. Blood samples were collected before and 5, 15, 30, 45, 60, 120, and 240 minutes after IV injection of dextrose (0.5 g/kg) for determination of plasma insulin and serum glucose, triglyceride, and nonesterified fatty acid concentrations. Between experimental periods, the llamas received supplemental amino acids IV (185 mg/kg in solution). The llamas were then fed a limited diet (grass hay, 0.25% of body weight daily) for 23 days, after which the experimental procedures were repeated. RESULTS: Feed restriction decreased glucose tolerance and had slight effects on insulin secretion in llamas. Basal lipid fractions were higher after feed restriction, but dextrose administration resulted in similar reductions in serum lipid concentrations with and without feed restriction. Insulin secretion was decreased on the second day of each study period, which lessened reduction of serum lipid concentrations but did not affect glucose tolerance. CONCLUSIONS AND CLINICAL RELEVANCE: Despite having a comparatively competent pancreatic response, feed-restricted llamas assimilated dextrose via an IV bolus more slowly than did llamas on full rations. However, repeated administration of dextrose reduced insulin secretion and could promote hyperglycemia and fat mobilization. These findings suggested that veterinarians should use alternative methods of supplying energy to camelids with long-term reduced feed intake or consider administering agents to improve the assimilation of glucose. PMID- 15281662 TI - Assessment of the metabolic effects of hydrocortisone on llamas before and after feed restriction. AB - OBJECTIVE: To evaluate the effects of administration of hydrocortisone on plasma concentration of insulin and serum concentrations of glucose, triglyceride, and nonesterified fatty acids (NEFAs) in llamas before and after feed restriction. ANIMALS: 9 adult female llamas. PROCEDURE: Feed was withheld from llamas for 8 hours. Blood samples were collected before (0 minutes) and 120, 180, 240, and 300 minutes after IV injection of hydrocortisone sodium succinate (1 mg/kg) for determination of plasma insulin concentration and serum concentrations of glucose, triglyceride, and NEFAs. The llamas were then fed a limited diet (grass hay, 0.25% of body weight daily) for 21 days, after which the experimental procedures were repeated. RESULTS: Compared with llamas that were not feed restricted, llamas after feed restriction had significantly higher plasma insulin concentration and serum concentrations of triglycerides and NEFAs. Feed restricted llamas after hydrocortisone injection had a significantly smaller increase in serum glucose concentration, a decrease (rather than an increase) in serum concentration of NEFAs, and no change in blood concentrations of insulin or triglycerides. CONCLUSIONS AND CLINICAL RELEVANCE: Short-acting glucocorticoid hormones did not appear to increase blood lipid concentrations in healthy llamas, regardless of ongoing fat mobilization. Thus, these hormones appear unlikely to be major direct contributors to diseases such as hepatic lipidosis or hyperlipemia. Although administration of hydrocortisone reduced serum concentration of fatty acids in feed-restricted llamas, its use has not been evaluated in sick camelids and cannot be considered therapeutically useful. PMID- 15281663 TI - Evaluation of a technique of inducing hypertensive renal insufficiency in cats. AB - OBJECTIVE: To compare 2 techniques of inducing combined renal insufficiency and systemic hypertension in cats. ANIMALS: 22 cats 6 to 12 months of age. PROCEDURES: Cats were randomly assigned to 1 of 3 groups. Control (C) group cats had 2 intact kidneys, remnant kidney (RK) group cats underwent unilateral partial renal infarction and contralateral nephrectomy, and remnant-wrap (W) group cats underwent unilateral partial renal infarction and partial abtation and wrapping of the contralateral kidney. Systemic arterial blood pressure (BP) was measured continuously by use of implanted radiotelemetric devices. Renal function was assessed via determination of glomerular filtration rate, measurement of serum creatinine and BUN concentrations, and determination of urine protein-to creatinine ratio (UP/C). Serum aldosterone concentration and plasma renin activity were measured on day 75. RESULTS: Systolic BP was significantly higher in groups RK and W than in group C, and systolic BP was significantly higher in group W than in group RK. Serum aldosterone concentration and plasma renin activity were significantly higher in group W, compared with groups C and RK. Glomerular filtration rate was significantly lower in groups RK and W, compared with group C. Histologic indices of renal injury and UP/C were significantly higher in group W, compared with groups C and RK. CONCLUSIONS AND CLINICAL RELEVANCE: Hypertensive renal insufficiency in group W was characterized by marked sustained systemic hypertension, decreased renal function, proteinuria, activation of the renin-angiotensin-aldosterone axis, and renal structural injury. Results support the hypothesis that marked systemic hypertension, activation of the renin-angiotensin-aldosterone axis, and proteinuria may damage the kidney of cats with preexisting renal insufficiency. PMID- 15281664 TI - Estimation of heritability of atopic dermatitis in Labrador and Golden Retrievers. AB - OBJECTIVE: To estimate the heritability of atopic dermatitis in Golden and Labrador Retrievers. ANIMALS: 429 dogs related to 13 dogs with atopic dermatitis. PROCEDURE: Atopic dermatitis was defined on the basis of the type and frequency of clinical signs recorded in the clinical records, and each dog was classified with atopic dermatitis or probable atopic dermatitis or as nonatopic. By use of data from atopic and nonatopic dogs, regression analyses of parental status on offspring status were performed to estimate heritability. RESULTS: There was no difference in the frequency of atopic dermatitis between sexes or between breeds. There was a marked association between the atopic status of the parent and that of the offspring, particularly for sires. By use of data from 32 litters in which the status of both parents was known and considering only those dogs classified with atopic dermatitis or as nonatopic, the heritability (+/- SE) of atopic dermatitis was estimated to be 0.47 (+/- 0.17). CONCLUSIONS AND CLINICAL RELEVANCE: Atopic dermatitis has a strong genetic component, and breeding of dogs with clinical signs of atopic dermatitis should be discouraged. PMID- 15281665 TI - Evaluation of forelimb horseshoe characteristics of thoroughbreds racing on dirt surfaces. AB - OBJECTIVE: To describe forelimb horseshoe characteristics of horses racing on dirt surfaces and determine whether these characteristics vary with region of California, season, horse characteristics, and race-related factors. ANIMALS: 5,730 Thoroughbred racehorses. PROCEDURE: From June 17, 2000, to June 16, 2001, the characteristics of 1 forelimb horseshoe of horses that raced on dirt surfaces at 5 major racetracks in California were recorded. These characteristics included shoe type; toe grab height; and presence of a rim, pad, and heel traction devices (jar caulks, heel stickers, heel blocks, and special nails). Horse and race information was obtained from commercial records. One race/horse was randomly selected. RESULTS: 99% of forelimb horseshoes were aluminum racing plates, 35% had a pad, 23% had a rim, and 8% had a heel traction device. A toe grab was observed on 75% of forelimb horseshoes (14% very low [< or = 2 mm], 30% low [> 2 and < or = 4 mm], 30% regular [> 4 and < or = 6 mm], and 1% high [> 6 and < or = 8 mm]). Forelimb horseshoe characteristics varied with region of California, season, age and sex of the horse, race purse and distance, and track surface condition. Log-linear modeling revealed that all of these factors were significantly interrelated. CONCLUSIONS AND CLINICAL RELEVANCE: Complex interrelationships among forelimb horseshoe characteristics and region, season, age and sex of the horse, and race-related factors need to be considered when evaluating the relationships between injury and horseshoe characteristics in Thoroughbred racehorses. PMID- 15281666 TI - Establishing the Graduate School of Nursing at the Uniformed Services University of the Health Sciences. PMID- 15281667 TI - Outcome of five years of accelerated surveillance in patients at high risk for inherited breast/ovarian cancer: report of a phase II trial. AB - OBJECTIVE: To assess the outcome of accelerated patient surveillance in patients at high risk for inherited breast or ovarian cancer. METHODS: Using stringent inclusion criteria, 57 high-risk patients (7 positive for BRCA1/2 mutations, 39 mutation negative, and 11 unaffected) were recruited from a genetic testing protocol for inherited breast/ovarian cancer and were followed for 5 years (192.5 total patient years). Patients received twice annual physical examinations, imaging studies, measurement of CA125 and CA15-3, psychometric measurements, and unstructured interviews by a psychologist. RESULTS: When mutation (+) and mutation (-) patients were compared, there were no significant differences in the development of disease metastasis, recurrence, or new cancers. No unaffected patients developed cancer. Management of osteoporosis, sexual function, and psychological distress were major concerns. CONCLUSIONS: Our data suggest that all patients with remarkable family history, regardless of their mutation status, may be at substantially increased risk for disease progression and development of new cancers, which is often not ovarian or recurrent breast cancer. Although prophylactic surgery is important in decreasing cancer recurrence in mutation carriers, increased surveillance with physical examinations and psychological support is also valuable and acceptable to such high-risk patients. PMID- 15281668 TI - Experience with directly observed prophylaxis using erythromycin in military trainees exposed to pertussis. AB - Pertussis, once a serious respiratory disease in children, has recently been identified as a common cause of chronic cough in adults. Military personnel are known to be vulnerable to this disease. After a training barracks exposure to pertussis, routine arrangements for contact prophylaxis with erythromycin failed. This experience is reported here as well as that of our subsequent aggressive attempts using directly observed prophylaxis (DOP) with standard erythromycin regimens. No secondary cases occurred. However, many contacts (35%) could not finish a 14-day course despite DOP, mostly because of nausea (85%) or diarrhea (72%). Seventeen (18%) soldiers missed classes because of erythromycin side effects; five required emergency department visits or hospital admission for the same. Sixteen (17%) soldiers were switched to azithromycin because of side effects; all were able to complete a 14-day course without symptoms. High adherence rates with erythromycin administration using DOP are attainable but may trigger unacceptable toxicities; alternative prophylactic regimens should be considered for active duty personnel. PMID- 15281669 TI - Department of Defense West Nile virus surveillance in 2002. AB - The Department of Defense (DoD) has engaged in West Nile virus (WNV) surveillance and response since 1999. In 2002, the three Services continued their cooperative, multidisciplinary approach to the WNV outbreak. Activities included a doubling of mosquito surveillance and vector control responses, extension of and doubling of bird and nonhuman mammal surveillance to all four continental United States regions, expanded diagnostic testing by DoD laboratories, and installation environmental clean up and personnel protection campaigns. Medical treatment facilities conducted passive surveillance and reported possible cases in DoD health care beneficiaries. Efforts were coordinated through active communication within installations, with commands, and with surrounding communities. Undertaken activities complemented each other to maximize surveillance coverage. The surveillance detected WNV on 44 DoD installations. It led directly to vector control and prevention activities, and there were no confirmed cases of WNV reported in the DoD force. This multi-Service effort is a surveillance template for future outbreaks that threaten DoD force health. PMID- 15281670 TI - Hearing health risk in a population of aircraft carrier flight deck personnel. AB - This study evaluated the risk to hearing health associated with duty on the flight deck of a Nimitz class aircraft carrier. Descriptive data includes time weighted average noise exposure and an evaluation of temporary threshold shift for a group of nonaviator flight deck personnel (FD), and a comparison of accrued permanent threshold shift among three shipboard occupational groups that had been matched for years of military service. The study participants included 76 FD personnel in a high-exposure group, 77 engineers in a moderate-exposure group, and 52 administrative personnel who were considered to have low occupational noise exposure. The study found a mean FD time weighted average of 109 dBA over workdays averaging 11.5 hours. Only 2 (4%) of 52 administrative personnel had any appreciable hearing loss (defined as worse than 20 dB at any frequency 1,000 through 4,000 Hz), whereas FD and engineers demonstrated 17% and 27% hearing impairment, respectively. PMID- 15281671 TI - Advance directives and do-not-resuscitate orders on general medical wards versus the intensive care unit. AB - The records of 335 patients admitted to the general medicine wards and to the medical intensive and coronary care unit (MICCU) at Brooke Army Medical Center were retrospectively reviewed to assess the frequency of advance directives and "do not resuscitate" (DNR) designations. Two hundred sixty-seven (79.7%) were admitted to the ward and 68 (20.3%) were admitted to the MICCU. Advance directives were executed in 14.9% of patients. DNR designations were made for 21 (7.9%) patients on the ward and 11 (16.2%) patients in the MICCU (p = 0.064). There were no statistical differences in mean length of stay, presence of advance directives, or documentation of advance directives in ward versus MICCU patients. However, there was a statistical difference in the number of deaths in the MICCU as compared with that on the ward (9.7 vs. 2.7%, p < 0.05). The frequency of advance directives and DNR designations did not differ between ward and MICCU patients in this population, although there was a trend for greater DNR designations in the MICCU environment. PMID- 15281672 TI - A comparison between two systems for pre-employment medical assessment in the Royal Netherlands Army by a randomized, controlled study. AB - In 1998, the basic medical requirements for the Royal Netherlands Army were introduced as a standard for the assessment of the medical suitability of military personnel, consisting of 43 dichotomized points of judgment. This system replaced the old physical capacity, upper limbs, locomotion, hearing, eyesight, and emotional and mental state system, based on the detection of diseases and infirmities. We compared the two different examination systems for their ability to identify suitable recruits. For the latter purpose, we used the two operational measures of availability and health care costs. We performed a randomized, controlled study in which 352 soldiers were monitored for 2 years after being declared fit by one of the pre-employment medical assessment systems in question and having passed their general military training. We found that the pre-employment medical assessment system was the dominant factor for predicting the number of days fit-for-duty, as well as for the health care costs incurred. Those declared fit by the new system showed a statistically significant higher mean number of days fit-for-duty (648 compared with 612) and incurred significantly lower mean health care costs (6396 compared with 746 Euro). In this study, we were not able to uncover the mechanism by which the "basic medical requirements" examination system led to an improvement in outcome. For the present, this mechanism is interpreted because of differences between the two systems. PMID- 15281673 TI - Operational dermatology. AB - Military dermatology encompasses all cutaneous manifestations that present to medical officers in a deployed situation, either in peacetime or in war. Medical officers in a field environment cannot avoid facing cutaneous quandaries. This article briefly highlights the omnipresent threat of cutaneous disease. It follows with a cost-effective look at periodically deploying a dermatologist to Bosnia. Volumes of historical data clearly justify the assignment of a dermatologist as a special consultant staff officer to every corps or theater medical staff. Despite this data, only two U.S. Army units in our present table of organization and equipment will have a dermatologist (60L) available for periodic direct-field consultation and teaching. After review of the historical data and the Stabilization Force-Bosnia statistics presented here, the table of organization and equipment restructuring that is required to meet the challenges of tomorrow will be clear. A dermatologist must be deployed as a theater or division consultant. PMID- 15281674 TI - Benign recurrent lymphocytic meningitis from herpes simplex virus type 2 during a summer outbreak of aseptic meningitis. AB - Meningitis from herpes simplex virus (HSV) may have a clinical presentation similar to other forms of viral meningitis. However, subtle facets of the history and use of the polymerase chain reaction (PCR) can differentiate HSV from other etiologies. During an outbreak of meningitis from enterovirus, a 32-year-old woman presented to the hospital with clinical meningitis, a history of genital HSV infection, and two previous bouts of viral meningitis. Her signs and symptoms as well as lumbar puncture results were similar to patients meeting our case definition for patients with presumed enteroviral meningitis. The cerebral spinal fluid was positive for HSV by PCR, and she was ultimately diagnosed with recurrent meningitis from HSV. We compared her presentation with patients who met our case definition for enteroviral meningitis. A thorough history and use of PCR may assist in differentiating these clinically similar presentations. PMID- 15281675 TI - Perceived versus actual medication regimens among internal medicine patients. AB - Exchange of accurate information between patients and medical providers is imperative for appropriate medication prescribing. We performed an evaluation of medication regimens of patients with information obtained independently from patient-completed surveys and nursing and provider interviews. The actual medication regimen was determined after the clinic encounter via mail-in forms or telephone interviews with patients reporting current medications directly from prescription bottles. Two hundred thirteen patients taking an average of 3.8 prescription medications were enrolled. Patients, nurses, and primary care providers were modestly accurate in reporting the number of medications being taken (kappa, 0.57,0.51, and 0.58, respectively); however, they performed poorly in reporting complete medication regimens as defined by the correct names, doses, and frequencies with 100% accuracy (34%, 26.7%, and 29.3%, respectively). Patients who created their own lists were more accurate than those who relied on memory, lists provided by providers, or discharge summaries. These findings indicate a significant difference between intended versus actual medication regimens at home. PMID- 15281676 TI - Implementing a smallpox vaccination program aboard an aircraft carrier. AB - OBJECTIVE: To determine the feasibility of implementing a smallpox vaccination program aboard an aircraft carrier in conjunction with anthrax vaccination. METHODS: Retrospective review of smallpox vaccination program conducted from January 17, 2003 to February 19, 2003. Morbidity and loss of manpower were the major endpoints. RESULTS: There were 5,204 sailors available for vaccination. There were 243 (4.7%) medical exemptions and 24 administrative exemptions. During the program, 4,931 sailors were vaccinated. There were five reportable complications. Three sailors had autoinoculation, one sailor had localized cellulitis, and one patient had a positive beta human chorionic gonadotropin during vaccination. None of the complications required medical evacuation. Only two sailors required time off from duty. CONCLUSIONS: Smallpox vaccination can be accomplished rapidly and safely aboard an aircraft carrier. There was not an increase in adverse events compared to historical data despite the close-quarter conditions. Smallpox and anthrax vaccinations can be completed simultaneously with minimal morbidity. PMID- 15281677 TI - Classification of the severity of U.S. Army and civilian reports of child maltreatment. AB - This study compares reports of the severity of child maltreatment for the U.S. Army and a civilian jurisdiction, Washington State (WS). Such comparisons can provide important information on risk and protective factors in designing prevention programs. An understanding of the differences facilitates the tailoring of interventions to better fit the characteristics of each community. The ages of the children in the WS cases were significantly older than the cases of the Army children. In both populations, neglect was the most prevalent form of maltreatment, followed in order by physical abuse, sexual abuse, and emotional abuse. The percentages of physical abuse, sexual abuse, and neglect were not statistically different, but the Army classified three times the number of emotional abuse cases as WS. The Army also classified more cases of physical abuse as severe (11%) compared with WS (5%). However, 16% of WS neglect cases were classified severe compared with 3% of Army cases. PMID- 15281678 TI - Childhood exposure to family violence and attrition in the Navy. AB - In a prospective study of U.S. Navy recruits (n = 5,491), we examined the relationship between childhood exposure to family violence (child physical abuse, child sexual abuse [CSA], and domestic violence) and attrition. Overall, 55% of recruits experienced one or more forms of childhood family violence and 34% of recruits attrited within 4 years after enlistment. Considered separately, each form of childhood violence was significantly associated with attrition. When considered simultaneously, all three types of childhood violence were associated with attrition in men, but only CSA was associated with attrition in women. Men and women who experienced all three types of childhood violence were 303% and 139%, respectively, more likely to attrite than recruits who reported no childhood violence. In analyses examining the timing of attrition, CSA was associated with early attrition, whereas child physical abuse and domestic violence were unrelated to timing of attrition. PMID- 15281679 TI - Domestic violence and post-traumatic stress disorder severity for participants of a domestic violence rehabilitation program. AB - Domestic violence has been a long-standing problem for our nation's active duty and military veterans. The purpose of this article is to describe participants of a domestic violence program, the program design to help lessen attrition, and the completers and noncompleters of the program. There was a significant relationship between post-traumatic stress disorder (PTSD) and domestic violence severity for the sample. PTSD severity was also related to reports of domestic violence in the family of origin. Completers and noncompleters were compared on demographic and violence variables and on nine research measures. Completers were more likely younger than 35 years old, employed, had higher self-ratings of relationship mutuality, lower levels of stress and post-traumatic stress, and were regularly court monitored. The results of a logistic regression significantly predicted completers and noncompleters based on age, relationship mutuality, PTSD, and court-monitored status (model chi2 statistic of 31.08, p = 0.0000). PMID- 15281680 TI - Stress, wellness, and mattering among cadets at West Point: factors affecting a fit and healthy force. AB - This study was designed to provide information to assist in planning health promotion and wellness programs for military trainees. During their first semester at West Point, 179 cadets completed measures of holistic wellness, perceived stress, and mattering. The highest wellness scores were in areas of social support, physical wellness, and humor. Significant negative correlations were found between perceived stress and work, realistic beliefs, and stress management. Significant positive correlations resulted between 17 wellness scales and mattering. t tests revealed only one area, work wellness, where the norm group scores exceeded those of the cadets. Cadets' scores exceeded norm group scores for perceived wellness, mattering, and seven of the Wellness Evaluation of Lifestyle scales, including total self-direction. Within-group differences were identified based on gender, for four of the self-direction factors, and for age on the dimension of self-care. Implications for health promotion and wellness from the perspective of holistic wellness are considered. PMID- 15281681 TI - Adequacy of Garrison feeding for Special Forces soldiers during training. AB - This study evaluated whether Special Forces (SF) soldiers training in garrison would meet nutrient intake recommendations using the available garrison dining facility. Dietary intakes were obtained by a visual estimation method and self reported food records from 32 SF and 13 support soldiers for 9 days. Total energy expenditure (TEE) was measured in nine soldiers from each group using doubly labeled water. Mean (+/- SD) total energy expenditure of SF (4,099 +/- 740 kcal/day) was higher than support soldiers (3,361 +/- 939 kcal/day, p < 0.01). Energy intake did not differ between groups. Median energy intake for all soldiers was 3,204 kcal/day. The nutrient intake goals of SF soldiers were not fully met by eating in the dining facility. Extending meal times and providing additional meals or "take out" foods may allow energy needs of SF soldiers (approximately 4,200 kcal/day) to be met, while reducing the reliance on potentially less nutritious outside foods. PMID- 15281682 TI - The effects of a four-day march on the gonadotrophins and mood states of army officers. AB - The effects of daily repeated prolonged exercise on gonadotrophin levels and mood states were studied in six physically active army officers participating in a 4 day march totaling 185 km. We hypothesized that submaximal daily repeated prolonged exercise may disturb the balance of the hypothalamic-pituitary testicular axis, which could be determined from the concentrations of serum luteinizing hormone and follicle-stimulating hormone with time-resolved immunofluorometric assay. In addition, the mood states of the men were followed during the exercise period. The results indicate that soldiers who are in good physical condition and are accustomed to marching are capable of marathon walking on 4 successive days while carrying a 10-kg backpack, without any major adverse effects on the function of the hypothalamic-pituitary-testicular axis or mood states. This study indicates that psychological monitoring and physiological measurements could be of value in following responses to daily repeated prolonged psychophysiological stress in field conditions with soldiers. PMID- 15281683 TI - Intracranial insertion of a nasopharyngeal airway in a patient with craniofacial trauma. AB - Intracranial insertion of a nasopharyngeal airway is an unusual and catastrophic complication of airway management in the patient with a severe closed head injury. We present an unfortunate 43-year-old patient with intracranial insertion of a nasopharyngeal airway during trauma resuscitation. The nasopharyngeal airway was removed. Attempts to resuscitate the patient were continued, but were eventually unsuccessful. Blind nasopharyngeal airway insertion may result in iatrogenic injury when used in the head-injured patient. Oropharyngeal airways may be used to assist with ventilation. However, it is preferable to definitively secure the airway through inline endotracheal intubation or with surgical techniques in this patient population. Should violation of the skull base occur, removal is accomplished in the controlled environment of the operating room. PMID- 15281684 TI - Me, and Walter Reed. PMID- 15281685 TI - Outsourcing that can work for your organization. PMID- 15281686 TI - Behave yourself. PMID- 15281687 TI - Sticky costs. Saving money through prevention. PMID- 15281688 TI - Social work organizations working together. PMID- 15281689 TI - Adolescent substance use: reviewing the effectiveness of prevention strategies. AB - U.S. youths continue to use alcohol, tobacco, and other drugs at alarmingly high rates despite a temporary downward trend in the 1980s. Among an average 500,000 individuals affected annually by substance use, youths (ages 12 to 18) rank as one of the highest groups in morbidity and mortality rates, resulting in many negative consequences. As a result the effectiveness of many prevention strategies has been called into question. This article reviews the extent and social cost of adolescent substance use; standard prevention strategies; prevention under criticism; and salient aspects of successful prevention strategies. Special attention is given to the social influence model of prevention as an effective and amenable model for social work professionals. PMID- 15281690 TI - Harm reduction: a new perspective on substance abuse services. AB - This article provides information on harm reduction, a recent development in substance abuse services in response to the HIV/AIDS epidemic. The author outlines abstinence and harm reduction perspectives and the stages of change model and discusses how these perspectives can be integrated in social work practice. He proposes using harm reduction strategies for individuals for whom the abstinence perspective may not be appropriate. Together, the traditional abstinence and harm reduction perspectives provide a basis for a more comprehensive continuum of care for individuals experiencing problems related to their substance use. PMID- 15281691 TI - Motivational strategies with alcohol-involved older adults: implications for social work practice. AB - Social workers and other health care professionals address problem drinking by older adults inconsistently. Reasons include client-related variables (for example, denial and poor information), practitioner-related factors (for example, inadequate knowledge about addictive behaviors, underdeveloped assessment tools, and limited empirically validated treatment options), and societal factors associated with "ageism." This article explores the nature and extent of problem drinking among older adults and barriers to assistance. The article outlines practice strategies that draw on motivational interviewing principles and a client's motivational readiness to change for reaching out to older adults, assessing their needs, and encouraging them to seek assistance. PMID- 15281692 TI - Methamphetamine abuse and manufacture: the child welfare response. AB - Methamphetamine abuse is on the rise, particularly by women of childbearing age. This article describes the history and effects of methamphetamine use. The authors examine the ways exposure to the manufacture of this drug affects clients and social workers in the course of their work. Because children are frequently found at the scene of a manufacturing laboratory, the child welfare system often becomes involved, and child protective services and other social work agencies need protocols to address the needs of the children and their parents, as well as those of the legal system. In 1997 California created and implemented drug endangered children's units in seven counties to address the needs of children from families that manufacture methamphetamine; these units involve collaborative efforts among child protective workers, district attorneys, physicians, and police officers. A case example provides information about the role of social workers and their collaboration with these multiple systems. PMID- 15281693 TI - Principles for practice with substance-abusing pregnant women: a framework based on the five social work intervention roles. AB - The author discusses components essential to pregnancy-specific substance abuse treatment, based on a review of the literature. Elements and issues related to substance abuse during pregnancy are identified under the five social work intervention roles: teacher, broker, clinician, mediator, and advocate. The concepts and approaches presented in this article can be applied by social workers in residential or outpatient substance abuse treatment programs; hospital prenatal, labor, and delivery units; the child welfare system; public health districts; or community family service centers. PMID- 15281694 TI - Health coverage instability for mothers in working families. AB - Using data from the National Longitudinal Survey of Youth, the authors examined the health insurance coverage stability of 1,667 women in working families over a three-year period (1995-1997). Findings revealed that coverage instability is common. Nearly one-half of low-income women experienced health coverage instability over the three-year study period, and low-income women with poor education, single marital status, low work hours, and frequent job changes were at even greater risk of coverage instability. The findings also imply that women affected by recent welfare reforms are likely to experience widespread health coverage problems. The implications for health care policy development, social work administration, and social work practice are discussed. PMID- 15281695 TI - Justice implications of a proposed Medicare prescription drug policy. AB - Social justice is a core value to the mission of social work. Older people are among the most vulnerable populations for whom social workers are called on to advocate. Although Medicare prescription drug coverage has been a top legislative issue over the past few years, such a benefit expansion has yet to be implemented. This article examines the historical context of Medicare and reviews the proposals for prescription drug coverage, identifying the concerns raised. Literature critiquing the justice dimensions of health care for the elderly population is reviewed. Justice claims are identified and refined, and social justice theories are used in the analysis of the proposed policies. PMID- 15281696 TI - Relationships between social work involvement and hospice outcomes: results of the National Hospice Social Work Survey. AB - In a struggle to balance fiscal realities with hospice philosophy, some hospices have attempted to cut costs by reducing social work involvement. This cross sectional survey of 66 hospices found, however, increased social work involvement was significantly associated with lower hospice costs. Additional benefits included better team functioning, more issues addressed by the social worker on the team, reduced medical services, and fewer visits by other team members, along with increased client satisfaction and lower severity of case. The authors concluded that higher salaries should be paid to a sufficient number of highly educated and experienced social workers. These social workers should be dedicated solely to the hospice social worker position, should participate in intake interviews, and should be supervised by a social worker. PMID- 15281698 TI - Validation of the health care surrogate preferences scale. AB - Recent advances in health care technology have increased the number of health care decisions made by acute care patients and those who act on their behalf, known as health care surrogates. This study reports on the validation of a new measure, the Health Care Surrogate Preferences Scale. Designed to assess the willingness of adults to perform and convey the duties required to communicate patient preferences, the scale offers a promising tool for use by social workers in health care settings. Development, evaluation, application of the new measure, and future research needs are discussed. PMID- 15281697 TI - Evaluation of a group intervention to assist aging parents with permanency planning for an adult offspring with special needs. AB - More than three-fourths of older adults with developmental disabilities and mental illness live in the community with aging parents, the majority of whom do not complete plans for the residential, financial, and legal future of their offspring. The authors used a true experimental design to evaluate the effectiveness of a six-week psychoeducational group intervention with 27 older mothers. Data collected in pre- and posttest telephone interviews were analyzed with repeated measures MANCOVA to test five hypotheses. Significant multivariate effects were found for mothers' knowledge and awareness about permanency planning, confidence and competence to plan, planning activities, and stage of planning. Findings support use of group interventions with older parents and underscore the need for professional education about planning for adults with special needs. PMID- 15281699 TI - Increasing the use of formal services by caregivers of people with dementia. AB - This study evaluated the impact of an information and referral intervention designed to help family caregivers of people with dementia obtain needed health and human services. Caregivers (N = 608) of community-residing people with dementia were surveyed about their need for and use of community services. Of the 608 caregivers, 203 agreed to be referred to a staff member of an Alzheimer's Association chapter to get help finding needed health and community services. The results of a logistic regression indicated that education, marital status, and perceptions about the helpfulness of services were significant predictors of wanting a referral. The information and referral help given by staff of the Alzheimer's Association chapters resulted in a significant increase in the use of human services, but no change in the use of health services. Practice implications are discussed. PMID- 15281700 TI - Dementia diary: a personal and professional journal. AB - This article is not traditional social work writing, because it is in the form of a memoir. It offers a window into lived experience, from which most professional writing is more distant. It explores the last year of life of an elderly woman, seen through the eyes of her daughter-in-law, who is a social worker and social work educator. The article chronicles the day-to-day experiences of progressive memory loss and the challenges of caregiving. Social work can develop similar narratives of other life experiences that can increase its practitioners' understanding and wisdom. PMID- 15281701 TI - Predictors of child custody plans for children whose parents are living with AIDS in New York City. AB - Custody planning among parents living with HIV or AIDS (PLHAs) can buffer the negative impact of parental death. The formal and informal custody plans for 594 children by 253 PLHAs and the relationships among custody plans, parental health, and psychosocial status were examined. About one-half of the parents had no formal custody plan. Parents were more likely to make plans for younger children. In addition, formal custody planning was more likely to occur among parents who reported using positive action, withdrawal-depressive, passive problem solving, social support, or spiritual coping styles and who reported higher parental self esteem. Parent's substance use and emotional distress were not significantly related to custody plans. These data suggest the need for interventions to encourage formal custody planning. PMID- 15281702 TI - A multidimensional conceptual framework for understanding HIV/AIDS as a chronic long-term illness. AB - New treatment advances have radically altered the course of HIV illness and created new challenges for HIV-affected individuals, families, and communities. This article provides a conceptual framework for understanding HIV in the multiple contexts of the client's culture, strengths, life course, and biomedical progression. The article concludes with a discussion of HIV prevention and treatment adherence as key focal points for social workers and their clients. PMID- 15281703 TI - The empowerment model: a critical reflection of empowerment in Chinese culture. AB - The empowerment model has long dominated social work practice in Western countries. Many social workers in Hong Kong use this model regardless of the social or cultural context. In this article the author shares local social work practice experiences in Hong Kong and suggests that the empowerment model may need adaptation in Chinese communities. An empowerment approach in communities influenced by traditional Chinese culture may result in types of pseudo empowerment: pragmatic empowerment, autocratic empowerment, hidden empowerment, lonely romantic empowerment, and fashionable empowerment. A culturally sensitive empowerment model for Chinese communities may have to be gradual and harmonious and empower both the individual and his or her significant others. PMID- 15281704 TI - Surviving as a postmodern social worker: two Ps and three Rs of direct practice. AB - Social workers interested in postmodernism have been provided an abundance of theory, but little to guide them in direct practice in diverse child welfare roles. In this article, two Ps and three Rs of practice based on postmodernist principles are discussed: positioning, power, resource sharing, resistance, and reflection. Professionals working in the delivery of frontline human services are struggling to work both with and in communities to celebrate diversity and localized constructions of reality while fulfilling professional and agency mandates. Informed by the broad theory of postmodernism, the two Ps and three Rs of practice allow workers to be mandated agents of the system, while deconstructing their privileged position. PMID- 15281705 TI - Client-controlled case information: a general system theory perspective. AB - The author proposes a model for client control of case information via the World Wide Web built on principles of general system theory. It incorporates the client into the design, resulting in an information structure that differs from traditional human services information-sharing practices. Referencing general system theory, the concepts of controller and controlled system, as well as entropy and negentropy, are applied to the information flow and autopoietic behavior as they relate to the boundary-maintaining functions of today's organizations. The author's conclusions synthesize general system theory and human services values to lay the foundation for an information-sharing framework for human services in the 21st century. PMID- 15281707 TI - How international is the social work knowledge base? PMID- 15281706 TI - Principled negotiation: a new tool for case advocacy. AB - Many methods of social work practice, including brokering, case advocacy, and cause advocacy, require the social worker to engage in negotiations to resolve disputes. This article demonstrates how principled negotiation, a form of negotiating developed out of the Harvard Negotiation Project at Harvard University and used widely in the business and legal world, can be an effective tool in social work practice. Principled negotiation is especially consonant with the value base of social work because it strives for the just and mutually beneficial resolution of conflicts while acknowledging the value and importance of ongoing relationships. A case example applying the rules of principled negotiation is described in the context of case advocacy. Its application to other social work practice methods is also examined. PMID- 15281708 TI - Social workers and the working and middle classes. PMID- 15281709 TI - Do social workers suppress Evangelical Christians? PMID- 15281710 TI - People with cognitive disabilities. PMID- 15281711 TI - People with cognitive disabilities: the argument from marginal cases and social work ethics. PMID- 15281712 TI - Parents with mental illness. PMID- 15281713 TI - Catholic charities. PMID- 15281714 TI - Catholic charities. PMID- 15281715 TI - Catholic charities. PMID- 15281716 TI - National Committee for Quality Assurance. PMID- 15281717 TI - Fair benefits in international medical research. PMID- 15281718 TI - Research on cognitively impaired adults. PMID- 15281719 TI - Canada bans human cloning. PMID- 15281720 TI - Schiavo: A hard case makes questionable law. PMID- 15281721 TI - A fifteen-year-old translator. PMID- 15281722 TI - On learning humility: a thirty-year journey. PMID- 15281723 TI - Moral standards for research in developing countries: from "reasonable availability" to "fair benefits". PMID- 15281724 TI - Do as I say, not as I do: Why bioethicists should seek informed consent for some case studies. PMID- 15281725 TI - Monitoring and manipulating brain function: new neuroscience technologies and their ethical implications. PMID- 15281727 TI - Reliability science: ensure system success even when components fail. AB - Half of all patients do not receive evidence-based care. The goal: All patients all the time receive the right care at the right time. 'Uncontrollable variables' are no excuse, proponents say. PMID- 15281726 TI - A tale of two conversations. PMID- 15281728 TI - System loop analysis eliminates phlebotomy lines. AB - New process was in place, but lab had been left out of the loop. Interviewing phlebotomists uncovers defects in processes. At least 90% of patients now are seen within 20 minutes of appointment time. PMID- 15281729 TI - Health plans offer rewards for quality improvement. AB - Patient satisfaction, preventive care most commonly used indicators. Goal is to demonstrate value to health insurance purchasers. Trend is 'nascent'; future growth of trend is far from certain. PMID- 15281730 TI - Compliance with protocols may improve outcomes. AB - Top 20% of hospitals had a 14% lower mortality rate. Protocols include use of aspirin, beta blockers, and internal mammary grafting procedure. Variance between top- and bottom-quintile hospitals as high as 22%. PMID- 15281732 TI - Patient safety alert. Finding root causes without blame helps eliminate errors. PMID- 15281731 TI - New clinical guidelines for palliative care published. AB - Great variability found in statements, definitions of palliative care. Authors determine need for benchmarks, gold standard of care. Core care elements, several distinct domains outlined in report. PMID- 15281733 TI - Unprecedented stereospecific synthesis of a novel tetracyclic ring system, a hybrid of tetrahydropyrrolo[2,3-b]indole and tetrahydroimidazo[1,2-a]indole, via a domino reaction upon a tryptophan-derived amino nitrile. AB - Compounds containing a novel tetracyclic ring system, a hybrid of tetrahydropyrrolo[2,3-b]indole and tetrahydroimidazo[1,2-a]indole, are synthesized via an acid-mediated stereospecific domino tautomerization of a tryptophan-derived alpha-amino nitrile. Characterization of these new compounds and preliminary studies on the reactivity of the tetracyclic heterocyclic system are reported. [reaction: see text] PMID- 15281735 TI - Efficient catalyst for the Suzuki-Miyaura coupling of potassium aryl trifluoroborates with aryl chlorides. AB - Palladium-catalyzed Suzuki-Miyaura cross-coupling reactions of aryl- and heteroaryl chlorides with potassium aryl- and heteroaryltrifluoroborates have been accomplished with the supporting ligand S-Phos in good to excellent yield. Hindered biaryls and substrates containing a variety of functional groups can be prepared. Suzuki-Miyaura couplings of a 3-pyridyl boron-based nucleophile with aryl- and heteroaryl chlorides proceed in good to very good yield. [reaction: see text] PMID- 15281734 TI - Novel stereoselective entry to 2'-beta-carbon-substituted 2'-deoxy-4' thionucleosides from 4-thiofuranoid glycals. AB - 2'-Beta-methyl- and 2'-beta-hydroxymethyl-2'-deoxy-4'-thionucleosides have been synthesized through PhSeCl-mediated electrophilic glycosidation using 4 thiofuranoid glycals having carbon substituents at the C2-position as a glycosyl donor. Preparation of these glycals were carried out by means of the C2 lithiation of 1-chloro-4-thiofuranoid glycal with LTMP followed by the Birch reduction of the chlorine atom. [reaction: see text] PMID- 15281736 TI - Selective syntheses of partially etherified derivatives of tetrakis(2 hydroxyphenyl)ethene. An alternative to the calix[4]arene ligand system. AB - Stereoselective syntheses of (E)- and (Z)-1,2-bis(2'-hydroxyphenyl)-bis(2' methoxyphenyl)ethene have been developed, the former by convergent coupling of an orthogonally protected 2,2'-benzophenone derivative and the latter by selective partial dealkylation of tetrakis(2-methoxyphenyl)ethene. Selective single demethylation has also been demonstrated in the 5-tert-butyl series. Thus, divalent and monovalent derivatives of the preorganized tetrakis(2 hydroxyphenyl)ethene ligand system are now available for use in coordination chemistry, analogous to corresponding calix[4]arene systems. [structure: see text] PMID- 15281737 TI - Development of high-load, soluble oligomeric sulfonate esters via ROM polymerization: application to the benzylation of amines. AB - The development of high-load, soluble oligomeric sulfonate esters, generated via ROM polymerization, and their utility in the facile benzylation of an array of amines is reported. These polymeric sulfonate esters exist as free-flowing powders, are stable at refrigerated temperatures, and are readily dissolved in CH(2)Cl(2). Following the benzylation event, purification is attained via simple filtration, followed by solvent removal to deliver the desired benzylated product in good to excellent yield and high purity. [structure: see text] PMID- 15281739 TI - Binding selectivity of cucurbit[7]uril: bis(pyridinium)-1,4-xylylene versus 4,4' bipyridinium guest sites. AB - The binding interactions between the host cucurbit[7]uril (CB7) and a series of linear guests containing bis(pyridinium)-1,4-xylylene and/or 4,4'-bipyridinium residues were investigated by (1)H NMR spectroscopy. CB7 was found to exhibit considerable binding selectivity for bis(pyridinium)-1,4-xylylene over 4,4' bipyridinium sites. New pseudo-rotaxane and rotaxane compounds were synthesized utilizing the host-guest interactions between CB7 and the surveyed guests. [structure: see text] PMID- 15281738 TI - Following an ISES lead: the first examples of asymmetric Ni(0)-mediated allylic amination. AB - An ISES (in situ enzymatic screening) lead pointed to conditions (PMP N protecting group, Ni(cod)(2) catalyst precursor) under which chiral, bidentate phosphines could promote Ni(0)-mediated allylic amination. Therefore, bidentate phosphines bearing central, axial, and planar chirality were examined with two model substrates of interest for PLP-enzyme inhibitor synthesis. In the best case, with (R)-MeO-BIPHEP, vinylglycinol derivative 2 was obtained in 75% ee (97% ee, one recrystallization) from 1. Further manipulation provided a Ni(0)-mediated entry into l-vinylglycine. [reaction: see text] PMID- 15281740 TI - Pi-conjugated dendrimers based on bis(enediynyl)benzene units. AB - We have synthesized a new family of pi-conjugated dendrimers that are based on bis(enediynyl)benzene units by using both divergent and convergent approaches. The compounds at all three generations have strong bluish-green fluorescence, especially the third-generation dendrimer, which has the highest extinction coefficient and quantum efficiency in this series. [structure: see text] PMID- 15281741 TI - Identification of the major tautomer for an etheno adduct of 2,6-diaminopurine by determination of the sign of (n)J(H,C). AB - The major of two solution-state tautomers observed for an etheno product of 2,6 diaminopurine was identified as the tautomer H-1 on the basis of the recognition of the two-bond coupling between the NH proton and C-9a and the three-bond coupling between the NH proton and C-3a. The couplings were distinguished as being over two- or three bonds by determination of the sign of the coupling using two-dimensional heteronuclear NMR, negative in the former case and positive in the latter case. [structure: see text] PMID- 15281742 TI - Synthesis of polycyclic aromatic iodides via ICl-induced intramolecular cyclization. AB - The reaction of 2-(arylethynyl)biphenyls with ICl at -78 degrees C affords substituted polycyclic aromatic iodides in good to excellent yields. The aryl substituents can be either electron-donating or electron-withdrawing groups such as OMe, Me, CHO, CO(2)Et or NO(2) groups. This chemistry has been successfully extended to systems containing a variety of polycyclic and heterocyclic rings. [reaction: see text] PMID- 15281743 TI - BINAP/1,4-diamine-ruthenium(II) complexes for efficient asymmetric hydrogenation of 1-tetralones and analogues. AB - A combined system of a RuCl(2)(binap)(1,4-diamine) complex and t-C(4)H(9)OK in i C(3)H(7)OH catalyzes enantioselective hydrogenation of various 1-tetralone derivatives and some methylated 2-cyclohexenones. Hydrogenation of 2-methyl-1 tetralone under dynamic kinetic resolution gives the cis alcohol with high ee. [reaction: see text] PMID- 15281744 TI - Solvent-dependent diastereoselectivities in reductions of beta-hydroxyketones by SmI2. AB - The reductions of a series of beta-hydroxyketones by SmI(2) were examined in THF, DME, and CH(3)CN using methanol as a proton source. Reductions in THF and DME typically lead to the syn diastereomer with DME providing higher diastereoselectivities. Reductions in CH(3)CN provided the anti diastereomer predominantly. This study reveals that solvation plays an important role in substrate reduction by SmI(2). [reaction: see text] PMID- 15281745 TI - Highly enantioselective copper-catalyzed conjugate addition of diethylzinc to nitroalkenes. AB - Copper-catalyzed asymmetric conjugate addition of diethylzinc to nitroalkenes using new chiral monodentate phosphoramidite ligands proceeds with high enantioselectivity up to 99% ee. [reaction: see text] PMID- 15281746 TI - Allyl, methallyl, prenyl, and methylprenyl ethers as protected alcohols: their selective cleavage with diphenyldisulfone under neutral conditions. AB - Diphenyldisulfone is a mild and efficient reagent for selective cleavage of methylprenyl (2,3-dimethylbut-2-en-1-yl), prenyl (3-methylbut-2-en1-yl), and methallyl (2-methylallyl) ethers. These reaction conditions are compatible with the presence of other protecting groups such as acetals, acetates, and allyl, benzyl, and TBDMS ethers. Exposure of 2,3-dimethylbut-2-en-1-yl and 3-methylbut-2 en1-yl ethers to diphenyldisulfone led to the formation of 2,3-dimethylbuta-1,3 diene and isoprene, respectively. 2-Methylallyl ethers undergo isomerization to 2 methylpropenyl ethers, which are easily hydrolyzed into the corresponding free alcohols and isobutyraldehyde. [reaction: see text] PMID- 15281747 TI - Biosynthesis of indole diterpenes, emindole, and paxilline: involvement of a common intermediate. AB - The key step for construction of the carbon skeleton in the indole diterpenes, paxilline, and emindole DA was examined. Intact incorporation of multiply (2)H labeled 3-geranylgeranylindole into two different fungal metabolites proves 3 geranylgeranylindole to be a biosynthetic intermediate. These results give evidence that indole diterpenes are biosynthesized via epoxidation of a common intermediate, and the subsequent cationic cyclization, analogous to those in the steroid biosynthesis. [structure: see text] PMID- 15281748 TI - Asymmetric allylboration of aldehydes and ketones using 3,3' disubstitutedbinaphthol-modified boronates. AB - Allylboronates derived from 3,3'-disubstituted 2,2'-binaphthols react with aldehydes and ketones to give the expected allylated products with up to >99:1 er. Highest selectivities were observed for aromatic ketones. The bis(trifluoromethyl) derivative is particularly outstanding in terms of reactivity, selectivity, and robustness. [reaction: see text] PMID- 15281749 TI - Novel N-aryl and N-heteroaryl sulfamide synthesis via palladium cross coupling. AB - A novel and efficient synthesis of N-aryl and N-heteroaryl sulfamides via an intermolecular palladium-catalyzed coupling process has been developed. The reactions proceeded with good to excellent yields and were tolerant of a wide range of functional groups. [reaction: see text] PMID- 15281750 TI - Cationic carbohydroxylation of alkenes and alkynes using the cation pool method. AB - The reactions of an N-acyliminium ion pool with alkenes and alkynes gave gamma amino alcohols and beta-amino carbonyl compounds, respectively, after treatment with H(2)O/Et(3)N. The present reaction serves as an efficient method for cationic carbohydroxylation of alkenes and alkynes. When vinyltrimethylsilane was used as an alkene, the reaction was highly diastereoselective and served as an access to an enantiomerically pure alpha-silyl-gamma-amino alcohol. [reaction: see text] PMID- 15281751 TI - Memory of chirality in the transannular cyclization of cyclodecenyl radicals. AB - The transannular cyclization of an enantioenriched cyclodecenyl radical proceeds in a 5-exo fashion to produce scalemic bicyclo[5.3.0]decanes. This cyclization is notable because it demonstrates chirality transfer through conformational memory of an unstabilized secondary radical. [reaction: see text] PMID- 15281753 TI - Total synthesis of cyclomarin C. AB - The total synthesis of cyclomarin C was accomplished through a convergent strategy from a tetrapeptide fragment and a tripeptide one. The developed methods to prepare the needed noncoded amino acids, the proper protection of peptide fragments, and identification of the optimum macrocylization site can be applied to further synthetic studies on other members of cyclomarins. [structure: see text] PMID- 15281752 TI - Reverse fluorous solid-phase extraction: a new technique for rapid separation of fluorous compounds. AB - Fluorous-tagged compounds can rapidly be separated from organic (non-tagged) compounds by the new separation technique of reverse fluorous solid-phase extraction (r-fspe). In a reversal of the roles of solid and liquid phases in standard fluorous spe, a mixture is charged to a polar solid phase (standard silica gel) and then eluted with a fluorous solvent or solvent mixture. The organic components of the mixture are retained, while the fluorous components pass. [structure: see text] PMID- 15281754 TI - Stereoselective chloroacetate aldol reactions: syntheses of acetate aldol equivalents and darzens glycidic esters. AB - Aldol reaction of the Ti-enolate derived from cis-1-tosylamido-2-indanyl chloroacetate with representative aldehydes proceeded in excellent yield and high diastereoselectivities. Removal of chlorine provided alternative access to highly diastereoselective acetate aldol equivalents or the corresponding glycidic ester condensation products. [reaction: see text] PMID- 15281755 TI - Stereocontrolled total synthesis of potent immunosuppressant FR901483. AB - A total synthesis of the potent immunosuppressant FR901483 (1) has been accomplished. The key feature of our convergent synthesis is the stereoselective incorporation of the p-methoxybenzyl and methylamino groups within the core moiety 10. Tricycle 10 was itself constructed by an intramolecular aldol reaction of the symmetrical keto-aldehyde 7. [Structure: see text] PMID- 15281756 TI - Oxidative cross-coupling of beta,beta-difluoroenol silyl ethers with nucleophiles: a dipole-inversion method to difluoroketones. AB - Oxidative cross-coupling of alpha-aryl-beta,beta-difluoroenol silyl ethers with heteroaromatics in the presence of Cu(OTf)(2) in wet acetonitrile proceeds smoothly, affording heteroaryldifluoromethyl aryl ketones in 61-88% yields. Alcohols also react as nucleophiles under the same conditions to provide alkoxydifluoromethyl aryl ketones in 73-80% yields. [reaction: see text] PMID- 15281757 TI - Synthesis of the sialic acid (-)-KDN and certain epimers from (-)-3 dehydroshikimic acid or (-)-quinic acid. AB - (-)-3-Dehydroshikimic acid (3-DHS, 4), a C(7)-building block now available in large quantity from corn syrup, has been converted into the sialic acid (-)-KDN (3) as well as its C-7- and C-8-epimers. (-)-Quinic acid can be used for the same purpose. [structure: see text] PMID- 15281758 TI - Synthesis of quinolines, 2-quinolones, phenanthridines, and 6(5h) phenanthridinones via palladium[0]-mediated Ullmann cross-coupling of 1-bromo-2 nitroarenes with beta-halo-enals, -enones, or -esters. AB - Palladium[0]-mediated Ullmann cross-coupling of 1-bromo-2-nitrobenzene (1 R = H) and its derivatives with a range of beta-halo-enals, -enones, or -esters readily affords the corresponding beta-aryl derivatives, which are converted into the corresponding quinolines, 2-quinolones, phenanthridines, or 6(5H) phenanthridinones on reaction with dihydrogen in the presence of Pd on C or with TiCl(3) in aqueous acetone. [reaction: see text] PMID- 15281759 TI - Generation and utility of tertiary alpha-aminoorganolithium reagents. AB - A general approach to tertiary alpha-aminoorganolithium reagents by reductive lithiation of alpha-aminonitriles has been developed. This class of organolithium nucleophiles reacts efficiently with carbonyl electrophiles or in intramolecular cyclizations with tethered phosphate leaving groups. Transmetalation can be used to produce alpha-aminoorganocuprate reagents that react with alkyl halide electrophiles and in 1,4-additions with enones. These methods establish a new approach for the synthesis of quaternary centers adjacent to nitrogen. [reaction: see text] PMID- 15281760 TI - Asymmetric synthesis of trans-3-amino-4-alkylazetidin-2-ones from chiral N,N dialkylhydrazones. AB - Enantiopure N,N-dialkylhydrazones 3 smoothly react with N-benzyloxycarbonyl-N benzyl glycine as an aminoketene precursor to afford trans-3-amino-4 alkylazetidin-2-ones 4 as single diasteromers. As an exception, hydrazone 3f (R = OBn) affords cis-(3R,4R)-4f under modified conditions. N-N Bond cleavage of cycloadducts 4 afforded free azetidinones 5 in high yields. [structure: see text] PMID- 15281761 TI - Synthesis and characterization of a series of homooligopeptide peroxyesters. AB - A homologous series of stable N(alpha)-phthaloyl peptide peroxyesters based on alpha-aminoisobutyric acid residues was prepared. In each of the six oligomers synthesized, the chain of alpha-amino acids is separated from the peroxyester function by a beta-amino acid. [structure: see text] PMID- 15281762 TI - Novel fluorescent pH sensors based on intramolecular hydrogen bonding ability of naphthalimide. AB - 4-Piperidine-naphthalimide derivatives containing potential acceptors for hydrogen bonding were synthesized, and their fluorescence properties were examined. Some of the compounds with a 2-imino-oxalidin (thiazolidin) side chain at the imide moiety exhibit a strong fluorescence quench and some red shift in weakly acidic conditions, caused by the formation of an intramolecular hydrogen bond. Their pK(a) values were estimated to be about 6.4-7.5. The results showed that these compounds could serve as novel fluorescent pH sensors for further application. [structure: see text] PMID- 15281763 TI - Dimethyl cuprate undergoes C-C bond coupling with methyliodide in the gas phase but dimethyl argenate does not. AB - Multistage mass spectrometry experiments have been used to synthesize and study the reactions of (CH3)2M-(M = Cu and Ag) with methyl iodide in the gas phase. While the dimethylcuprate ion (M = Cu) reacts with CH3I via C-C bond cross coupling, its silver congener is unreactive. The experimental results are consistent with MP2/6-31++G** ab initio calculations, which reveal that the preferred mechanism for Cu involves the formation of a T-shaped Cu transition state. [reaction: see text] PMID- 15281764 TI - Fluorous phase-switching of pyridyl-tagged substrates/products. AB - A bis-monopyridyl Wang-type tag was prepared for application in the hydrocarbon/perfluorocarbon phase-switching of substrates/products. This "catch and-release" strategy relies on the reversible coordination of the tag to a fluorous copper(II)-carboxylate complex. A hydantoin was prepared in high yield and purity by a four-step protocol where the intermediates were purified by the straightforward liquid/liquid phase-switching process. [reaction: see text] PMID- 15281765 TI - Selective mono-O-acylation of C2V-symmetrical calix[4]arenediols with acylisocyanates. AB - Calix[4]arenedialkyl ethers 3 react with an excess of acylisocyanates to give selectively monoacylated products 4. Intramolecular hydrogen bonds and steric effects of the acylcarbamate fragments are most likely responsible for the high selectivity of this monoprotection. [reaction: see text] PMID- 15281766 TI - Ruthenium-catalyzed silyl ether formation and enyne metathesis sequence: synthesis of siloxacycles from terminal alkenyl alcohols and alkynylsilanes. AB - Two consecutive ruthenium-catalyzed reactions have been achieved for the synthesis of siloxacycles from terminal alkenyl carbenols and alkynylsilanes. The metal-catalyzed dehydrogenative condensation between alcohols and silanes, generating molecular hydrogen as the only byproduct, allows for the subsequent enyne metathesis without isolating the intermediate silyl ethers. This system provides a streamlined synthesis of synthetically useful building blocks. [reaction: see text] PMID- 15281768 TI - S-alkyl dithioformates as 1,3-dipolarophiles. Generation of C(2)-unsubstituted penems. AB - S-Alkyl dithioformates, generated by a cycloreversion process, react as 1,3 dipolarophiles with beta-lactam-based azomethine ylids to provide, after (net) elimination of MeSH, C(2)-unsubstituted penems. The overall cycloreversion/cycloaddition sequence was accelerated by microwave irradiation. [reaction: see text] PMID- 15281767 TI - A hierarchy of aryloxide deprotection by boron tribromide. AB - Aryl propargyl ethers and esters are cleaved selectively in the presence of aryl methyl ethers and esters by boron tribromide in dichloromethane. Under the same conditions, allyl ethers undergo very rapid Claisen rearrangement, and benzyl ethers are also cleaved more rapidly than propargyl. A mechanism involving intramolecular delivery of bromide to the propargyl terminus is proposed. [reaction: see text] PMID- 15281769 TI - Synthesis of five-, six-, and seven-membered ring lactams by CpRh complex catalyzed oxidative N-heterocyclization of amino alcohols. AB - A new effective catalytic system consisting of [CpRhCl(2)](2)/K(2)CO(3) (Cp = pentamethylcyclopentadienyl) for the lactamization of amino alcohols has been developed. As an example, the reaction of 3-(2-aminophenyl)-1-propanol in the presence of [CpRhCl(2)](2) (5.0% Rh) and K(2)CO(3) (10%) in acetone gives 3,4 dihydro-2(1H)-quinolinone in an isolated yield of 80%. A variety of five-, six-, and seven-membered benzo-fused lactams are synthesized by this catalytic system. [reaction: see text] PMID- 15281771 TI - Use of polymer-supported Pd reagents for rapid and efficient Suzuki reactions using microwave heating. AB - A rapid and efficient Suzuki coupling protocol has been developed utilizing polymer-supported palladium catalysts and microwave irradiation. It is also shown that solid-phase extraction of excess boronic acids can be rapidly and conveniently accomplished by utilization of a silica-supported carbonate base. [reaction: see text] PMID- 15281770 TI - Asymmetric synthesis of syn-(2R,3S)-and anti-(2S,3S)-ethyl diamino-3 phenylpropanoates from N-(benzylidene)-p-toluenesulfinamide and glycine enolates. AB - Addition of differentially N-protected glycine enolates to enantiopure sulfinimines affords syn- and anti-alpha,beta-diamino esters with high diastereoselectivities and good yields. PMID- 15281772 TI - Synthesis of [60]fullerene adducts bearing carbazole moieties by Bingel reaction and their properties. AB - Carbazole-linked [60]fullerene adducts were successfully prepared by the Bingel reactions using carbazole derivatives bearing one or two ethyl malonate moieties. In the latter cases, specific bisadduct regioisomers were obtained, depending on the spacer unit between two ethyl malonate moieties. [reaction: see text] PMID- 15281773 TI - A stereodivergent approach to amino acids, amino alcohols, or oxazolidinones of high enantiomeric purity. AB - (-)-Menthone, an inexpensive chiral auxiliary, was used to prepare both enantiomers of alpha-amino acids, amino alcohols, or oxazolidinones. The sequence includes the S(N)2' displacement by a cuprate reagent and a Curtius rearrangement as key steps. PMID- 15281774 TI - Synthetic applicability and in situ recycling of a B-methoxy oxazaborolidine catalyst derived from cis-1-amino-indan-2-ol. AB - A procedure is described that greatly simplifies the use of an oxazaborolidine catalyst derived from (1R,2S) cis-1-amino-indan-2-ol. This B-OMe catalyst has been employed in the asymmetric reduction of a number of structurally diverse prochiral ketones, in particular the reduction of alpha-amino acetophenone and its derivatives. A method for reducing the effective catalyst loading by "in situ recycling" is also presented. [structure: see text] PMID- 15281775 TI - Dabco as an inexpensive and highly efficient ligand for palladium-catalyzed Suzuki-Miyaura cross-coupling reaction. AB - An inexpensive and highly efficient Pd(OAc)(2)/Dabco catalytic system has been developed for the cross-coupling of aryl halides with arylboronic acids. A combination of Pd(OAc)(2) and Dabco (triethylenediamine) was observed to form an excellent catalyst, which affords high TONs (turnover numbers; TONs up to 950 000 for the reaction of PhI and p-chlorophenylboronic acid) for Suzuki-Miyaura cross coupling of various aryl iodides and bromides with arylboronic acids. [reaction: see text] PMID- 15281776 TI - Can the nitroso ene reaction proceed concertedly? AB - All nitroso ene reactions so far reported follow exclusively stepwise reaction paths. Herein, we report the first concerted nitroso ene reaction that occurs between o-isotoluene (or its naphthalenic analogues) and nitroso compounds (e.g., nitrosomethane and 4-nitronitrosobenzene). [reaction: see text] PMID- 15281777 TI - A simple and efficient L-prolinamide-catalyzed alpha-selenenylation reaction of aldehydes. AB - An efficient and simple l-prolinamide-catalyzed alpha-selenenylation reaction of aldehydes with N-(phenylseleno)phthalimide has been developed for the efficient preparation of alpha-phenylselenoaldehydes. Such compounds are versatile building blocks for the synthesis of alpha,beta-unsaturated aldehydes, allylic alcohols, and amines. [reaction: see text] PMID- 15281778 TI - Palladium-catalyzed allylalkynylation of benzynes: a highly efficient route to substituted 1-allyl-2-alkynylbenzenes. AB - Benzynes, generated in situ from 2-(trimethylsilyl)aryl triflates and cesium fluoride, undergo allylalkynylation with allylic chlorides and alkynylstannanes in the presence of palladium catalyst to give 1-allyl-2-alkynylbenzenes in good to excellent yields. [reaction: see text] PMID- 15281779 TI - Stereoselective synthesis of 3-alkylideneoxindoles using tandem In-mediated carbometalation and Pd-catalyzed cross-coupling reaction. AB - Efficient methods for stereoselective synthesis of various (E)-, (Z)-, and disubstituted 3-alkylideneoxindoles were investigated using tandem In-mediated carbometalation and ligandless Pd-catalyzed coupling reaction. PMID- 15281780 TI - Efficient Cu-catalyzed asymmetric conjugate additions of alkylzinc reagents to aromatic and aliphatic acyclic nitroalkenes. AB - An efficient and highly enantioselective (up to 95% ee) Cu-catalyzed method for asymmetric conjugate addition (ACA) of alkylzinc reagents to acyclic disubstituted nitroalkenes is presented. Reactions are typically effected at ambient temperature in the presence of 2 mol % chiral dipeptide phosphine and 1 mol % (CuOTf)(2).C(6)H(6). Nitroalkenes bearing aromatic as well as aliphatic substituents readily undergo asymmetric additions. [reaction: see text] PMID- 15281781 TI - Palladium-catalyzed enantioselective ring opening of oxabicyclic alkenes with organozinc halides. AB - Palladium-catalyzed asymmetric ring opening of oxabenzonorbornadienes with readily available organozinc halides under mild conditions in the presence of (S) Pr(i)-PHOX produces the corresponding 1,2-dihydronaphth-1-ols in good yield and high enantioselectivity. [reaction: see text] PMID- 15281782 TI - Thermally and electrochemically controllable self-complexing molecular switches. AB - Self-complexing molecular systems are obtained when an arm component (a pi-donor) is covalently linked to a preformed macrocycle (a pi-acceptor). The resulting self-complexing compounds are not only attractive in relation to their topology, but also for their potential to undergo reversible movements, i.e., the arm can be driven out of or into the cavity of the linked macrocycle in response to temperature or applied voltage. These structurally sensitive changes render them potential thermosensors or electroswitches. PMID- 15281783 TI - Discovery and characterization of catalysts for azide-alkyne cycloaddition by fluorescence quenching. AB - Copper-based catalysts for the 1,3-dipolar cycloaddition of azides and alkynes were screened in parallel fashion using a fluorescence quenching assay. The method was designed to identify systems able to accelerate the coupling of reactants at micromolar concentrations in aqueous mixtures and to obtain quantitative comparisons of their activities. In addition to the tris(triazolylamines) previously reported, two types of compounds (bipy/phen and 2-pyridyl Schiff bases) were found to exhibit significant ligand-accelerated catalysis, with one complex showing especially dramatic rate enhancements. Preliminary explorations of the dependence of reaction rate on pH, ligand:Cu ratio, and Cu concentration are described. PMID- 15281784 TI - On the mechanism of [Rh(CO)2Cl]2-catalyzed intermolecular (5 + 2) reactions between vinylcyclopropanes and alkynes. AB - DFT calculations have been applied to investigate the reaction mechanism of rhodium dimer, [Rh(CO)2Cl]2, catalyzed intermolecular (5 + 2) reactions between vinylcyclopropanes and alkynes. The catalytic species is Rh(CO)Cl and the catalytic cycle is through the sequential reactions of cyclopropyl cleavage of vinylcyclopropane, alkyne insertion (rate-determining step), and a migratory reductive elimination. PMID- 15281785 TI - [60]fullerene-stoppered porphyrinorotaxanes: pronounced elongation of charge separated-state lifetimes. AB - A series of Sauvage-type porphyrinorotaxanes containing [60]fullerene stoppers have been synthesized by a convergent route. Photoinduced energy transfer and electron-transfer reactions in these rotaxanes yield long-distance charge separated radical-pair states, whose lifetimes in solution at ambient temperatures are as high as 32 mus, depending on the distance between the fullerene and zinc porphyrin chromophores. PMID- 15281786 TI - Coordination-driven self-assembly directs a single-crystal-to-single-crystal transformation that exhibits photocontrolled fluorescence. AB - Coordination-driven self-assembly is employed to direct a single-crystal-to single-crystal [2 + 2] photodimerization that exhibits tunable fluorescence. PMID- 15281787 TI - A mechanism-based cross-linker for the identification of kinase-substrate pairs. AB - The reversible phosphorylation of proteins is one of the most important mechanisms for the regulation of signal transduction cascades. Recently, there has been substantial progress made in the identification of new phosphoproteins and phosphorylation sites. Unfortunately, there are very few methods available that allow this information to be used to identify the upstream kinase responsible for the phosphorylation event. Herein, we describe a new method that allows the cross-linking of a substrate of interest to its upstream kinase. This method relies upon a novel, mechanism-based cross-linker and the replacement of the phosphorylated residue with a cysteine residue. The application of this method to a number of kinase-peptide substrate pairs is described. PMID- 15281788 TI - Efficient use of bifunctional acid-base properties for alkylammonium formation in amine-substituted zeolites. AB - The formation of alkylammonium groups in amine-doped zeolites is studied using density functional theory on small clusters representing the chemically active site. The presence of both strong Lewis base and Bronsted acid sites leads to a significant lowering of reaction barriers as opposed to alkoxide formation in full-oxygen zeolites. Furthermore, amine-substituted zeolites suggest novel reaction pathways that are not solely centralized around the aluminum substitution but in which two tetrahedral sites are involved, maximizing use of the zeolitic acid site and its surroundings. An investigation of the proton mobility in these yet to be synthesized materials demonstrates the need for minimizing the amount of Al-NH-Si bridges, as to prevent protonation of the amine group. PMID- 15281789 TI - Isolation, characterization, and theoretical study of La2@C78. AB - A new metallofullerene, La2@C78, has been synthesized by DC arc discharge method, isolated by high-performance liquid chromatography, and characterized by laser desorption time-of-flight mass spectrometry, UV-vis-NIR absorption, differential pulse voltammetry, 13C NMR spectroscopy, and theoretical calculations. The La2@C78/CS2 solution is dark violet and presents several characteristic absorption features at 647, 561, 533, and 386 nm, with an onset around 1000 nm. With respect to empty D3-C78, the capability of La2@C78 as an electron acceptor or donor is stronger. Addition of 1,1,2,2-tetrakis(2,4,6-trimethylphenyl)-1,2 disirane to La2@C78 photochemically, as well as thermally, affords bis- and mono adducts. Theoretical studies and 13C NMR spectroscopic analysis of La2@C78 indicate that it possesses a D3h-C78 cage (78:5). PMID- 15281790 TI - Defect dynamics observed by NMR of quadrupolar nuclei in gallium nitride. AB - Hexagonal and cubic polytypes of bulk gallium nitride powders are characterized by 69,71Ga and 14N MAS NMR at 11.7 T. The (corrected) 71Ga chemical shifts are 333.0 and 357.5 ppm, respectively; the corresponding 14N chemical shifts are 301.8 and -297.0 ppm (all shifts referenced to 1 M gallium nitrate). The 69,71Ga nuclear quadrupole coupling constants (NQCC) in the hexagonal form are axially symmetric and agree with previous single-crystal determinations. The 71Ga MAS NMR satellite pattern envelope of the cubic form has a large Gaussian half-height width of 297 kHz, due to nonzero NQCC values induced by defects. The 14N MAS NMR spinning sideband pattern of the cubic form has a Lorentzian envelope half-height width of 17.5 kHz for the same reason. A sample containing both phases shows an unexpected marked loss of the 71Ga MAS NMR satellite transition intensity expected for the hexagonal phase. Static 71Ga-selective Hahn spin-echo measurements at the perpendicular edge of the powder pattern for the hexagonal form in this sample show a large reduction in T2, especially at higher temperatures. The partial destruction of both spin-echoes and rotational echoes is due to a chemical-exchange type process involving sites having different NQCC values. PMID- 15281791 TI - Doxorubicin accumulation in individually electrophoresed organelles. AB - We report the doxorubicin content in individual organelles following their capillary electrophoretic separation and illustrate that chemical accumulation at the subcellular level is highly heterogeneous. In individual mitochondria from cultured human leukemia cells DOX amount is around 50 zmol, 2 orders of magnitude higher than expected from diffusion during drug treatment, and spans 2 orders of magnitude. PMID- 15281792 TI - Photogenerated polyelectrolyte bilayers from an aqueous-processible photoresist for multicomponent protein patterning. AB - A novel photoresist (PR) that can be processed under mild aqueous conditions was synthesized and used to create photogenerated polyelectrolyte bilayers. Thin films of the PR cast on polycation-coated substrates were exposed to UV irradiation to generate carboxylate groups in the photoresist. The bulk of the UV exposed PR film was dissolved by rinsing with pH 7.4 phosphate-buffered saline, but a polyelectrolyte bilayer formed in situ at the PR/polycation interface on exposure remained bound to the substrate. The UV-exposed photoresist also exhibited pH-dependent solubility; it was soluble in water above pH 6.6, but insoluble at lower pHs. Using these unique properties, two-component protein patterning was achieved using biotinylated PR films under conditions that avoid exposing the proteins to conditions outside the narrow range of physiological pH, ionic strength, and temperature where their stability is greatest. PMID- 15281793 TI - Alkane oxidation via photochemical excitation of a self-assembled molecular cage. AB - The photoexcitation of a self-assembled M6L4-type coordination cage accommodating photochemically inert alkane guests (e.g., adamantane and cyclooctane) led to the regioselective oxidation of the guest within the cage. Under anaerobic conditions, the guest oxidation was accompanied by the stable radical formation as indicated by ESR spectrometry (g = 2.002) and change in solution color (from colorless to blue). These phenomena were shown to be characteristic of the self assembled molecular systems: i.e., from the M6L4 supersetGn assembly, none of the components (M, L, or G) can be eliminated for the unusual oxidation and/or radical formation. PMID- 15281794 TI - Topochemical transketalization reaction driven by hydrogen bonding. AB - An unusual thermal isomerization of 1,2;3,4-di-O-isopropylidene-myo-inositol to 1,2;5,6-di-O-isopropylidene-myo-inositol was observed in the solid state. A detailed study revealed that this ketal migration is a topochemically driven one. X-ray structure revealed that strong intermolecular hydrogen bonding in the crystal preorganizes the hydroxyl and ketal carbon in a transition-state-like arrangement favorable for the reaction. The 5-OH of each molecule faces the trans ketal carbon of a neighboring molecule at a close distance and an angle of approach near to linearity, making a trigonal-bipyramidal-like arrangement in the crystal, which could facilitate the reaction. This is the first report of a topochemical transketalization reaction. PMID- 15281795 TI - Chiral recognition in surface explosion. AB - The vast majority of chiral compounds crystallize into racemic crystals. It has been predicted and was experimentally established as a rule that chiral molecules on surfaces are more easily separated into homochiral domains due to confinement into a plane and lower entropic contributions. We investigated the formation and stability of two-dimensional tartrate crystals on a Cu(110) surface for the racemic mixture for the first time by means of temperature-programmed desorption (TPD), low-energy electron diffraction (LEED), and X-ray photoelectron spectroscopy (XPS). At low coverage, a bitartrate species becomes separated into homochiral domains, while at high coverage a monotartrate species forms a racemic mixture. At the same coverage and lateral arrangement, the thermally induced autocatalytic decomposition reaction occurs for the monotartrate racemate at a lower temperature than for the pure enantiomers. The stereochemistry in this so called "surface explosion" reaction is explained by a higher stability of the enantiopure lattice due to lateral hydrogen-bond formation. The higher stability of the enantiopure two-dimensional lattice is in contrast to the higher stability of racemic three-dimensional tartaric acid crystals but is consistent with the observation that homochirality is preferred in hydrogen-bonded self-assembled biomolecular structures. PMID- 15281796 TI - Control of metal coordination number in de novo designed peptides through subtle sequence modifications. AB - Substitution of an alanine for leucine (shown in light blue) in the hydrophobic interior of designed three-stranded coiled coils allows for the control of metal ion coordination number and geometry. The influence of this perturbation by a noncoordinating residue can be monitored by the dramatic impact on the 113Cd NMR spectrum. The structural effect occurs even when the residue substitution is as much as 7 A from the metal binding site. PMID- 15281797 TI - NMR analysis of enzyme-catalyzed and free-equilibrium mutarotation kinetics of monosaccharides. AB - We have exploited the saturation difference (SD) NMR technique to investigate the mutarotations of two sugars, l-fucose and d-ribose, at equilibrium. The KM and kcat values of Escherichia coli FucU-catalyzed mutarotation of l-fucose have been determined by this technique, and the values of KM for alpha- and beta-forms are shown to be the same. Similarly, the spontaneous mutarotation of d-ribose was investigated by using the same method, and it was found that d-ribose has an exceptionally high spontaneous alpha-to-beta conversion rate only between the furan forms. PMID- 15281798 TI - One ligand fits all: cationic mono(amidinate) alkyl catalysts over the full size range of the group 3 and lanthanide metals. AB - Using a sterically demanding amidinate ancillary ligand and an in situ alkylation procedure, neutral mono(amidinate) dialkyl and cationic mono(amidinate) monoalkyl complexes were prepared for metals spanning the full size range of the group 3 and lanthanide metals. The activity of the cationic monoalkyls in catalytic ethene polymerization was found to vary by over 2 orders of magnitude depending on the metal ionic radius, the intermediate metal sizes being found to be the most effective. PMID- 15281799 TI - Cobalt-mediated two-carbon ring expansion of five-membered rings. Electrophilic carbon-carbon bond activation in the synthesis of seven-membered rings. AB - Electrophilic cobalt(III) mediates an unprecedented two-carbon ring expansion of coordinated five-membered rings, leading to a remarkably general new strategy for the synthesis of seven-membered carbocycles from readily available five-membered ring substrates. The reaction, a metal-mediated [5 + 2] cyclopentenyl/alkyne cycloaddition, proceeds via initial protonation of a cobalt(I) cyclopentadiene complex, followed by rearrangement to an agostic eta3-cyclopentenyl intermediate. The cyclic eta3-allyl residue then undergoes migratory coupling with alkyne followed by carbon-carbon bond activation of the unstrained five-membered ring and recyclization to the ring expanded product, although the order of events and intimate mechanism has not been conclusively established. The reaction is highly selective with respect to which five-membered ring ligand undergoes activation, presumably a consequence of rapid cobalt-mediated interannular hydride transfer and kinetic preference for alkyne insertion into the less substituted cyclopentenyl ring. The alkyne insertion is itself highly regioselective, proceeding via migration to the sterically smaller end of the alkyne. The reaction is sensitive to both the cobalt counterion and the ancillary eta5 cyclopentadienyl substituent but proceeds for a considerable range of alkyl-, aryl-, and trialkylsilyl-substituted terminal and internal alkynes. PMID- 15281800 TI - Biaryl synthesis via direct arylation: establishment of an efficient catalyst for intramolecular processes. AB - In this Communication, we describe direct arylation reactions with improved scope and catalyst activity for the intramolecular formation of biaryl compounds. This was achieved through the establishment of a highly active and robust catalyst system and the subsequent development of a novel phosphine ligand 27. The enhanced catalytic activity extends these transformations to include previously unreactive and poorly reactive substrates, and allows for very low catalyst loadings to be employed-as little as 0.1 mol %. PMID- 15281801 TI - Catalytic enantioselective conjugate addition of carbamates. AB - Catalytic, asymmetric conjugate addition of carbamates to enoyl systems has been realized for the first time, providing a two-step access to virtually enantiopure N-protected beta-amino acids. PMID- 15281802 TI - Stepwise bromination of two acetylene molecules on a butterfly-type tetrairon core and reactivity of the resulting bromoacetylene fragment toward nucleophiles. AB - Treatment of the acetylene-coordinated tetrairon cluster, [(eta5 C5H4Me)4Fe4(HCCH)2]+, with N-bromosuccinimide led to stepwise bromination of two acetylene ligands to form [(eta5-C5H4Me)4Fe4(HCCBr)(HCCH)]+, [(eta5 C5H4Me)4Fe4(HCCBr)2]+, [(eta5-C5H4Me)4Fe4(BrCCBr)(HCCBr)]+, and [(eta5 C5H4Me)4Fe4(BrCCBr)2]+. The reactivity of the bromoacetylene fragment in [(eta5 C5H4Me)4Fe4(HCCBr)(HCCH)]+ toward water, pyridine, and ZnMe2 was also investigated. PMID- 15281803 TI - Catalytic enantio- and diastereoselective aldol reactions of glycine-derived silicon enolate with aldehydes: an efficient approach to the asymmetric synthesis of anti-beta-hydroxy-alpha-amino acid derivatives. AB - We have developed an efficient method for the asymmetric synthesis of anti-beta hydroxy-alpha-amino acid derivatives based on highly enantio- and diastereoselective aldol reactions of the silicon enolate derived from N trifluoroacetylglycinate with aldehydes using a chiral zirconium catalyst. The resulting N-trifluoroacetyl group is easily cleaved under either acidic or basic conditions and can be used directly as a protecting group for further transformations. PMID- 15281804 TI - A powerful selection assay for mixture libraries of DNA alkylating agents. AB - A simple and powerful selection assay that permits the separation (rpHPLC), quantitation (ELSD), and identification (ESI-MS) of thermally released adenine adducts derived from duocarmycin analogues is detailed that can establish the most effective DNA alkylating agents in synthetic combinatorial mixtures. PMID- 15281805 TI - Deoxystreptamine dimers bind to RNA hairpin loops. AB - RNA is known to have multiple roles in critical cellular functions. Thus, there is great potential for RNA-binding small molecules as both therapeutic agents and cellular probes. Unfortunately, the multiple secondary structures that RNA can adopt have caused difficulty in the development of a general paradigm for RNA small molecule binding. In particular, the standard RNA-binding compounds such as aminoglycosides do not generally bind RNA hairpin loops, a widespread and vitally important secondary structural motif. In this manuscript we report that dimers of deoxystreptamine bind to RNA hairpin loops with affinities rivaling that of RNA aminoglycoside interactions. PMID- 15281806 TI - Amino acid N-carboxyanhydrides: activated peptide monomers behaving as phosphate activating agents in aqueous solution. AB - The hydrolysis of valine N-carboxyanhydride (NCA) in aqueous phosphate buffers was shown to proceed through nucleophilic catalysis via an aminoacyl phosphate intermediate that displays phosphorylating capabilities through a potentially prebiotic process that simulates modern biochemical metabolic pathways. PMID- 15281807 TI - Regio- and enantiocontrol in the room-temperature hydroboration of vinyl arenes with pinacol borane. AB - The catalyzed hydroboration of vinyl arenes was carried out using pinacol borane instead of catechol borane, as the former reagent and the product boronates are significantly easier to handle. By careful choice of catalyst, either the branched or the linear product can be obtained in greater than 96% selectivity. Interestingly, common ligands such as BINAP and Josiphos give opposite asymmetric induction with pinacol borane as compared with catechol borane, while P,N-ligands such as Quinap gave the same sense of induction. The hydroboration of 6 methoxynaphthalene proceeded with the greatest regio- (95:5) and enantioselectivity (94:6) of all vinyl arenes examined. The hydroboration product was then employed in a concise synthesis of the nonsteroidal antiinflammatory agent, Naproxen. PMID- 15281808 TI - Shape-shifting tetranuclear oxo-bridged manganese cluster: relevance to photosystem II water oxidase active site. AB - The redox properties of the "dimer-of-dimers" complex, [{Mn2(mu-O)2(tphpn)}2]4+ (1) (where Htphpn = N,N,N',N'-tetra(2-methylpyridyl)-2-hydroxypropane-diamine) were investigated. The structure changes dramatically to an adamantane-shaped core upon one-electron oxidation. On the other hand, the one-electron reduced product of 1, [Mn4O4(tphpn)2]3+, exhibits a hyperfine-structured multiline EPR signal very similar to the so-called S0 state of the tetramanganese cluster, which resides at the Photosystem II water oxidase active site. PMID- 15281809 TI - The disarming effect of the 4,6-acetal group on glycoside reactivity: torsional or electronic? AB - An evaluation of whether the well-known deactivating effect of a 4,6-acetal protection group on glycosyl transfer is caused by torsional or an electronic effect from fixation of the 6-OH in the tg conformation was made. Two conformationally locked probe molecules, 2,4-dinitrophenyl 4,8-anhydro-7-deoxy 2,3,6-tri-O-methyl-beta-D-glycero-D-gluco-octopyranoside (18R) and the L-glycero D-gluco isomer (18S), were prepared, and their rate of hydrolysis was compared to that of the flexible 2,4-dinitrophenyl 2,3,4,6-tetra-O-methyl-beta-D glucopyranoside (21) and the locked 2,4-dinitrophenyl 4,6-O-methylidene-2,3-di-O methyl-beta-D-glucopyranoside (26). The rate of hydrolysis at pH 6.5 was 21 > 18R > 18S > 26, which showed that the deactivating effect of the 4,6-methylene group is partially torsional and partially electronic. A comparison of the rate of acidic hydrolysis of the corresponding methyl alpha-glycosides likewise showed that the probe molecules 17S and 17R hydrolyzed significantly slower than methyl tetra-O-methyl-glucoside 19, confirming a deactivating effect of locking the saccharide in the (4)C(1) conformation. The experiments showed that the hydroxymethyl rotamers deactivate the rate of glycoside hydrolysis in the order tg >> gt > gg. PMID- 15281810 TI - Biosynthesis of lambertellols based on the high specific incorporation of the 13C labeled acetates and their biological properties. AB - Biosyntheses of lambertellols A (1) and B (2) as well as lambertellin (3) were investigated by isotope labeling experiments. Nearly 40% of specific incorporation of [1,2-(13)C(2)]acetate was achieved, and all the carbons in 1 and 2 were labeled. This high incorporation of the labeled acetate was realized by providing INADEQUATE spectra by employing only 0.4 and 0.7 mg of 1 and 2, respectively. Our studies revealed that 1-3 are biogenetically synthesized via loss of two carbons from octameric acetate. A biological assay against Monilinia fructicola revealed those remarkably inhibited hyphal germinations. However, neither of them killed the spores immediately, even in high concentration. These conditions induced the formation of microconidia. PMID- 15281811 TI - Nitroxides scavenge myeloperoxidase-catalyzed thiyl radicals in model systems and in cells. AB - Nitroxide radicals possess important antioxidant activity in live tissues because of their ability to scavenge reactive radicals. Despite the fact that, in cells, damaging free radicals are primarily quenched by glutathione (GSH) with subsequent formation of harmful glutathionyl radical (GS(*)), interactions of nitroxide radicals with GS(*) and thiols have not been studied in detail. In addition, intracellular metabolic pathways leading to the formation of secondary amines from nitroxides are unknown. Here we report that GS(*) radicals react efficiently and irreversibly with nitroxides to produce secondary amines. We developed a sensitive method for the detection of GS(*) based on their specific interaction with Ac-Tempo, a nonfluorescent conjugate of fluorogenic acridine with paramagnetic nitroxide Tempo, and used it to characterize interactions between nitroxide and thiyl radicals generated through phenoxyl radical recycling by peroxidase. During reaction of Ac-Tempo with GS(*), Tempo EPR signals decayed and acridine fluorescence concurrently increased. DMPO and PBN, spin traps for GS(*), inhibited this interaction. Using combined HPLC and mass spectrometry, we determined that 90% of the Ac-Tempo was converted into fluorescent acridine (Ac) piperidine; GSH was primarily oxidized into sulfonic acid. In myeloperoxidase rich HL-60 cells, Ac-piperidine fluorescence was observed upon stimulation of GS(*) generation by H(2)O(2) and phenol. Development of fluorescence was prevented by preincubation of cells with the thiol-blocking reagent N ethylmaleimide as well as with peroxidase inhibitiors. Furthermore, Ac-Tempo preserved intracellular GSH and protected cells from phenol/GS(*) toxicity, suggesting a new mechanism for the free-radical scavenging activity of nitroxides in live cells. PMID- 15281812 TI - Resonance Raman spectroscopy reveals new insight into the electronic structure of beta-hematin and malaria pigment. AB - Resonance Raman spectra of beta-hematin and hemin are reported for a range of excitation wavelengths including 406, 488, 514, 568, 633, 780, 830, and 1064 nm. Dramatic enhancement of A(1g) modes (1570, 1371, 795, 677, and 344 cm(-1)), ring breathing modes (850-650 cm(-1)), and out-of-plane modes including iron-ligand modes (400-200 cm(-1)) were observed when irradiating with 780- and 830-nm laser excitation wavelengths for beta-hematin and to a lesser extent hemin. Absorbance spectra recorded during the transformation of hemin to beta-hematin showed a red shift of the Soret and Q (0-1) bands, which has been interpreted as excitonic coupling resulting from porphyrin aggregation. A small broad electronic transition observed at 867 nm was assigned to a z-polarized charge-transfer transition d(xy) --> e(g)(pi). The extraordinary band enhancement observed when exciting with near-infrared excitation wavelengths in beta-hematin when compared to hemin is explained in terms of an aggregated enhanced Raman scattering hypothesis based on the intermolecular excitonic interactions between porphyrinic units. This study provides new insight into the electronic structure of beta hematin and therefore hemozoin (malaria pigment). The results have important implications in the design and testing of new anti-malaria drugs that specifically interfere with hemozoin formation. PMID- 15281813 TI - Factors influencing the autoxidation of fatty acids: effect of olefin geometry of the nonconjugated diene. AB - Autoxidations of cis,cis, cis,trans, and trans,trans nonconjugated octadecadienoates and pentadecadienes were carried out in the presence of alpha tocopherol to investigate the effect of olefin geometry on this oxidation process and provide insight into the factors that influence the autoxidation of fatty acids. We have found that as the trans character of the diene increases, the amount of O(2) trapping at the central (bis-allylic) position of the pentadienyl radical also increases. In addition, the rate constant for beta-fragmentation (k(beta) approximately 10(6) s(-1)) of the bis-allylic peroxyl radical decreased on going from the cis,cis to the trans,trans diene. We have also found that for the cis,trans nonconjugated dienes, there is a preference for trapping of the pentadienyl radical by O(2) at the transoid end, generating the cis,trans conjugated hydroperoxide as the major product. PMID- 15281814 TI - Synthesis, relaxometric and photophysical properties of a new pH-responsive MRI contrast agent: the effect of other ligating groups on dissociation of a p nitrophenolic pendant arm. AB - Two gadolinium(III) chelates, GdNP-DO3A (1-methlyene-(p-NitroPhenol)-1,4,7,10 tetraazacycloDOdecane-4,7,10-triAcetate) and GdNP-DO3AM (1-methlyene(p NitroPhenol)-1,4,7,10-tetraazacycloDOdecane-4,7,10-triacetAMide), containing a single nitrophenolic pendant arm plus either three acetate or three amide pendant arms were synthesized and characterized. The properties of the gadolinium, terbium, and dysprosium complexes of these ligands were examined as a function of pH. The extent and mechanism of the changes in water relaxivity with pH of each gadolinium complex was found to differ substantially for the two complexes. The water relaxivity of Gd(NP-DO3A) increases from 4.1 mM(-1) s(-1) at pH 9 to 7.0 mM(-1) s(-1) at pH 5 as a result of acid-catalyzed dissociation of the nitrophenol from the lanthanide. The nitrophenol group in Gd(NP-DO3AM) does not dissociate from the metal center even at pH 5; therefore, the very modest increase in relaxivity in this complex must be ascribed to an increase in prototropic exchange rate of the bound water and/or phenolic protons. PMID- 15281815 TI - Mispair-aligned N3T-alkyl-N3T interstrand cross-linked DNA: synthesis and characterization of duplexes with interstrand cross-links of variable lengths. AB - Therapeutic bifunctional alkylating agents generate interstrand cross-links in duplex DNA. As part of our continuing studies on DNA duplexes that contain alkyl interstrand cross-links, we have synthesized a cross-link that bridges the N(3) positions of a mismatched thymidine base pair. This cross-link, which is similar to the N(3)C-alkyl-N(3)C cross-link that has been observed between mismatched cytosine base pairs, was introduced by first incorporating a cross-linked phosphoramidite unit at the 5'-end of an oligonucleotide chain. Fully cross linked duplexes were then synthesized using an orthogonal approach to selectively remove protecting groups, thus allowing construction of the cross-linked duplex via conventional solid-phase oligonucleotide synthesis. Short DNA duplexes with alkyl cross-links of various lengths (two, four, and seven methylene units) were prepared, and their physical properties were studied via UV thermal denaturation and circular dichroism spectroscopy. These linkers were found to stabilize the duplexes by 37, 31, and 16 degrees C for the two-, four-, and seven-carbon linkers, respectively, relative to a non-cross-linked duplex. Circular dichroism spectra suggested that these lesions induce very little deviation in the global structure relative to the non-cross-linked duplex DNA control. Molecular models show that the two-carbon cross-link spans the distance between the N(3) atoms of the T-T mismatch without perturbing the helix structure, whereas the longer linkers, particularly the seven-carbon linker, tend to push the thymines apart, creating a local distortion. This perturbation may account for the lower thermal stability of the seven-carbon versus two-carbon cross-linked duplex. PMID- 15281816 TI - Modular aptameric sensors. AB - We report the first examples of modular aptameric sensors, which transduce recognition events into fluorescence changes through allosteric regulation of noncovalent interactions with a fluorophore. These sensors consist of: (a) a reporting domain, which signals the binding event of an analyte through binding to a fluorophore; (b) a recognition domain, which binds the analyte; and (c) a communication module, which serves as a conduit between recognition and signaling domains. We tested recognition regions specific for ATP, FMN, and theophylline in combinations with malachite green binding aptamer as a signaling domain. In each case, we were able to obtain a functional sensor capable of responding to an increase in analyte concentration with an increase in fluorescence. Similar constructs that consist only of natural RNA could be expressed in cells and used as sensors for intracellular imaging. PMID- 15281817 TI - Amplification of chirality from molecules into morphology of crystals through molecular recognition. AB - We have found a novel type of morphological chiral tuning on inorganic helical crystals through stereochemical recognition of organic molecules. Helical forms consisting of twisted twins emerged from triclinic crystals under diffusion limited conditions. The proportion of the right- and left-handed helices was precisely tuned with the addition of a specified amount of chiral molecules, such as d- and l-glutamic acids. The chiral molecules recognized the enantiomeric surface of the triclinic crystal and then changed the growth behavior of the helical morphology. As a result, the microscopic chiral information, at a molecular level, was amplified into the macroscopic helices consisting of inorganic achiral components. PMID- 15281818 TI - Formation of branched calixarene aggregates-a time-resolved static light scattering study. AB - Mixtures of a calix[4]arene and a naphthyridine derivative dissolved in 1,2 dichlorobenzene form thermoreversible aggregates. The aggregation process was followed by means of time-resolved multiangle light scattering at two different mixing ratios, 1:3 and 1:4, yielding a detailed record of the relative mass, the radius of gyration, and the particle scattering function of the growing aggregates. On the basis of these data, a conclusive model of the structure is presented for the developing aggregates: monomers aggregate to wormlike filaments which form branching points. Formation of branching points proceeds in a frequency and distribution which is similar to the polycondensation of ABC monomers toward non-randomly branched macromolecules (Burchard, W. Macromolecules 1977, 10, 919-927). Thus, aggregation results in hyperbranched-like particles with striking analogies to the polymerization of glucose to amylopectin. PMID- 15281819 TI - Stereochemistry of isoplagiochin C, a macrocyclic bisbibenzyl from liverworts. AB - Cyclic bisbibenzyls, like isoplagiochins C (1) and D (2), are stereochemically intriguing molecules: Although not equipped with any of the traditional stereogenic elements that render molecules conformationally stable per se, they are sometimes isolated in an optically active form and are thus chiral at room temperature. The paper describes quantum chemical calculations, in particular investigations of the conformational space and molecular dynamics simulations, showing that the helicity is a property of the entire molecule, whose ring strain makes the molecule configurationally stable overall, with (formally) three stereogenic elements (two biaryl axes and one helical stilbene unit). Only one of the biaryl axes (the 'upper' one, joining C-12' and C-14) has a stable configuration, leading to a population of four interconverting diastereomers, yet without racemization at room temperature. On the basis of these conformational and dynamic calculations, the circular dichroism spectrum of isoplagiochin C (1) was calculated, leading to the first assignment of the absolute configuration of a cyclic bisbibenzyl. Accordingly, 1 has the P-configuration at the stereochemically stable biaryl axis and constitutes a mixture of diastereomers with respect to the other biaryl axis and the helical stilbene unit. From the temperature dependence of the racemization rates, an enantiomerization barrier of 101.6 kJ/mol was determined. Likewise, for the first time for cyclic bisbibenzyls, the enantiomeric ratio of this natural product was determined, by chromatography on a chiral phase with CD-coupling. Accordingly, 1 from Plagiochila deflexa is not enantiomerically pure, but occurs in a 85:15 ratio in favor of the enantiomer that has the P-configuration at the stereochemically stable axis. PMID- 15281820 TI - Metal-ion sensing fluorophores with large two-photon absorption cross sections: aza-crown ether substituted donor-acceptor-donor distyrylbenzenes. AB - Chromophores based on a donor-acceptor-donor structure possessing a large two photon absorption cross section and one or two mono-aza-15-crown-5 ether moieties, which can bind metal cations, have been synthesized. The influence of Mg(2+) binding on their one- and two-photon spectroscopic properties has been investigated. Upon binding, the two-photon action cross sections at 810 nm decrease by a factor of up to 50 at high Mg(2+) concentrations and this results in a large contrast in the two-photon excited fluorescence signal between the bound and unbound forms, for excitation in the range of 730 to 860 nm. Experimental and computational results indicate that there is a significant reduction of the electron donating strength of the aza-crown nitrogen atom(s) upon metal ion binding and that this leads to a blue shift in the position as well as a reduction in the strength of the lowest-energy two-photon absorption band. The molecules reported here can serve as models for the design of improved two-photon excitable metal-ion sensing fluorophores. PMID- 15281821 TI - Total synthesis of formamicin. AB - The enantioselective total synthesis of the cytotoxic plecomacrolide natural product formamicin (1) is described. Key aspects of this synthesis include the efficient transacetalation reactions of MOM ethers 28 and 38 to form the seven membered formyl acetals 29 and 39, a late-stage Suzuki cross-coupling reaction of the highly functionalized vinyl boronic acid 6 and vinyl iodide 7, a highly beta selective glycosidation reaction of beta-hydroxy ketone 4 with 2,6-dideoxy-2 iodoglucopyranosyl fluoride 3, and the global desilylation of penultimate intermediate 77 mediated by in situ generated Et(3)N.2HF. PMID- 15281822 TI - Nitro-substituted Hoveyda-Grubbs ruthenium carbenes: enhancement of catalyst activity through electronic activation. AB - The design, synthesis, stability, and catalytic activity of nitro-substituted Hoveyda-Grubbs metathesis catalysts are described. The highly active and stable meta- and para-substituted complexes are attractive from a practical point of view. These catalysts operate in very mild conditions and can be successfully applied in various types of metathesis [ring-closing metathesis, cross-metathesis (CM), and enyne metathesis]. Although the presence of a NO(2) group leads to catalysts that are dramatically more active than both the second-generation Grubbs's catalyst and the phosphine-free Hoveyda's carbene, enhancement of reactivity is somewhat lower than that observed for a sterically activated Hoveyda-Grubbs catalyst. Attempts to combine two modes of activation, steric and electronic, result in severely decreasing a catalyst's stability. The present findings illustrate that different Ru catalysts turned out to be optimal for different applications. Whereas phosphine-free carbenes are catalysts of choice for CM of various electron-deficient substrates, they exhibit lower reactivity in the formation of tetrasubstituted double bonds. This demonstrates that no single catalyst outperforms all others in all possible applications. PMID- 15281823 TI - EPR study of dialkyl nitroxides as probes to investigate the exchange of solutes between the ligand shell of monolayers of protected gold nanoparticles and aqueous solutions. AB - EPR spectroscopy has been used to study the interaction of para-substituted benzyl hydroxyalkyl nitroxides with the monolayer of water-soluble protected gold cluster made by a short alkyl chain and a triethylene glycol monomethyl ether unit. The inclusion of nitroxide probes in the more hydrophobic environment of the monolayer gave rise to a reduction of the value of both nitrogen and beta proton hyperfine splittings. The spectra also showed selective line broadening attributed to modulation of the spectroscopic parameters as the result of exchange between free and complexed nitroxide. The rate constants were obtained by analyzing the EPR line shape variations as functions of nanoparticle concentration and temperature. This represents, to the best of our knowledge, the first determination of rate constants for the solubilization of organic substrates in a monolayer-protected cluster. PMID- 15281824 TI - Mechanistic studies of the transfer dehydrogenation of cyclooctane catalyzed by iridium bis(phosphinite) p-XPCP pincer complexes. AB - Reaction of bis(phosphinite) PCP iridium pincer complexes (p-XPCP)IrHCl (5a-f) [X = MeO (5a), Me (5b), H (5c), F (5d), C(6)F(5) (5e), Ar(F)(= 3,5 bis(trifluoromethyl)phenyl) (5f)] with NaOtBu in neat cyclooctane (COA) generates 1:1 mixtures of the respective (p-XPCP)IrH(2) complexes 4a-f and the cyclooctene (COE) olefin complexes (p-XPCP)Ir(COE) (6a-f) at 23 degrees C. At higher temperatures, complexes 4 and 6 are equilibrated because of the degenerate transfer dehydrogenation of COA with free COE (6 + COA right harpoon over left harpoon 4 + 2COE), as was shown by temperature-dependent equilibrium constants and spin saturation transfer experiments at 80 degrees C. At this temperature, the COE complexes 6 exchange with free COE on the NMR time scale with the more electron-deficient complexes 6 exchanging COE faster. The exchange is dissociative and zero order in [COE]. Further analysis reveals that the stoichiometric hydrogenation of COE by complex 4f, and thus the separated back reaction 4f + 2COE --> 6f + COA proceeds at temperatures as low as -100 degrees C with the intermediacy of two isomeric complexes (p-Ar(F)PCP)Ir(H)(2)(COE) (8f, 8f'). COE deuteration with the perdeuterated complex 4f-d(38) at -100 degrees C results in hydrogen incorporation into the hydridic sites of complexes 8f,8f' d(38) but not in the hydridic sites of complex 4f-d(38), thus rendering COE migratory insertion in complexes 8f,8f' reversible and COE coordination by complex 4f rate-determining for the overall COE deuteration. PMID- 15281825 TI - Diastereoselective formation of chiral tris-cyclometalated iridium (III) complexes: characterization and photophysical properties. AB - Chiral, facial tris-cyclometalated Ir(III) complexes, fac-Delta-Ir(pppy)(3), fac Lambda-Ir(pppy)(3), fac-Lambda-IrL (where pppy is (8R,10R)-2-(2'-phenyl)-4,5 pinenopyridine and L is a tripodal ligand comprising three pppy moieties connected through a mesityl spacer) have been synthesized and characterized. In IrL, NMR and CD studies indicate that only one diastereomer is formed, with the Lambda configuration at the metal center, whereas enantiopure pppy yields the fac Lambda- and the fac-Delta-stereoisomer in a ratio 2:3. fac-Lambda-IrL was structurally characterized using X-ray crystallography. The luminescence properties including CPL, of the three complexes and their sensitivity to dioxygen were examined. PMID- 15281826 TI - Design and photochemical characterization of a biomimetic light-driven Z/E switcher. AB - Protonated Schiff bases (PSBs) of polyenals constitute a class of light-driven switchers selected by biological evolution that provide model compounds for the development of artificial light-driven molecular devices or motors. In the present paper, our primary target is to show, through combined computational and experimental studies, that it is possible to approach the design of artificial PSBs suitable for such applications. Below, we use the methods of computational photochemistry to design and characterize the prototype biomimetic molecular switchers 4-cyclopenten-2'-enylidene-3,4-dihydro-2H-pyrrolinium and its 5,5' dimethyl derivative both containing the penta-2,4-dieniminium chromophore. To find support for the predicted behavior, we also report the photochemical reaction path of the synthetically accessible compound 4-benzylidene-3,4-dihydro 2H-pyrrolinium. We show that the preparation and photochemical characterization of this compound (together with three different N-methyl derivatives) provide both support for the predicted photoisomerization mechanism and information on its sensitivity to the molecular environment. PMID- 15281827 TI - Agostic interactions and dissociation in the first layer of water on Pt(111). AB - Recent quantum mechanical (QM) calculations for a monolayer of H(2)O on Ru(0001) suggested a novel stable structure with half the waters dissociated. However, different studies on Pt(111) suggested an undissociated bilayer structure in which the outer half of the water has the OH bonds toward the surface rather than the O lone pair. Since water layers on Pt are important in many catalytic processes (e.g., the fuel cell cathode), we calculated the energetics and structure of the first monolayer of water on the Pt(111) surface using QM [periodic slab using density functional calculations (DFT) with the PBE-flavor of exchange-correlation functional]. We find that the fully saturated surface ((2)/(3) ML) has half the water almost parallel to the surface (forming a Pt-O Lewis acid-base bond), whereas the other half are perpendicular to the surface, but with the H down toward the surface (forming a Pt-HO agostic bond). This leads to a net bond energy of 0.60 eV/water = 13.8 kcal/mol (the standard ice model with the H up configuration of the water molecules perpendicular to the surface is less stable by 0.092 eV/water = 2.1 kcal/mol). We examined whether the partial dissociation of water proposed for Ru(0001) could occur on Pt(111). For the saturated water layer ((2)/(3) ML) we find a stable structure with half the H(2)O dissociated (forming Pt-OH and Pt-H covalent bonds), which is less favorable by only 0.066 eV/water = 1.51 kcal/mol. These results confirm the interpretation of combined experimental (XAS, XES, XPS) and theoretical (DFT cluster and periodic including spectrum calculations) studies, which find only the H down undissociated case. We find that the undissociated structure leads to a vertical displacement between the two layers of oxygens of approximately 0.42 A (for both H down and H up). In contrast, the partially dissociated system leads to a flat structure with a separation of the oxygen layers of 0.08 A. Among the partially dissociated systems, we find that all subsurface positions for the dissociated hydrogen are less favorable than adsorbing on top of the free Pt surface atom. Our results suggest that for less than (1)/(3) ML, clustering would be observed rather than ordered monolayer structures. PMID- 15281828 TI - Supercritical self-assembled monolayer growth. AB - The growth of octadecyltrimethylammonium bromide (C(18)TAB) monolayers on mica was investigated using atomic force microscopy and infrared spectroscopy. A critical temperature was identified below which the monolayer formed via an "islanding" mechanism, that is, nucleation and growth of densely packed two dimensional (2D) islands within a matrix of a disordered dilute phase. However, above the critical temperature, there was no coexistence of 2D phases during film formation. Instead, the monolayer gradually became better ordered, remaining laterally homogeneous throughout. We show that this corresponds to a critical point in a 2D phase diagram of the monolayer. Additional evidence is provided by the in situ observation of 2D phase separation upon cooling an incomplete monolayer from the one-phase to the two-phase region. The lack of coexisting domains (and domain boundaries) during growth above the critical point provides a possible route for the preparation of essentially defect-free monolayers. PMID- 15281829 TI - Intermittent single-molecule interfacial electron transfer dynamics. AB - We report on single-molecule studies of photosensitized interfacial electron transfer (ET) processes in Coumarin 343 (C343)-TiO(2) nanoparticles (NP) and Cresyl Violet (CV(+))-TiO(2) NP systems, using time-correlated single-photon counting coupled with scanning confocal fluorescence microscopy. Fluorescence intensity trajectories of individual dye molecules adsorbed on a semiconductor NP surface showed fluorescence fluctuations and blinking, with time constants distributed from milliseconds to seconds. The fluorescence fluctuation dynamics were found to be inhomogeneous from molecule to molecule and from time to time, showing significant static and dynamic disorders in the interfacial ET reaction dynamics. We attribute fluorescence fluctuations to the interfacial ET reaction rate fluctuations, associating redox reactivity intermittency with the fluctuations of molecule-TiO(2) electronic and Franck-Condon coupling. Intermittent interfacial ET dynamics of individual molecules could be characteristic of a surface chemical reaction strongly involved with and regulated by molecule-surface interactions. The intermittent interfacial reaction dynamics that likely occur among single molecules in other interfacial and surface chemical processes can typically be observed by single-molecule studies but not by conventional ensemble-averaged experiments. PMID- 15281830 TI - Hydration of phenylketene revisited: a counter-intuitive result. AB - In previous work (Can. J. Chem. 1987, 65, 1719-1723 and J. Am. Chem. Soc. 1995, 117, 9165-9171), flash photolysis of diazoacetophenone or phenylhydroxycyclopropenone in aqueous solution was found to produce phenylketene as a short-lived transient species with absorbance at lambda congruent with 260 nm, which decayed with single-exponential kinetics. It has now been discovered that, in the acidity region [H(+)] = 0.000 01 to 0.06 M, this decay is preceded by a faster absorbance rise, and that the overall change conforms well to a double exponential rate law. Analysis of the new data produces rate profiles whose general shapes, as well as the numerical values of their constituent rate constants, plus the form of buffer catalysis, indicate that this newly discovered absorbance rise represents ketonization of phenylacetic acid enol, and that the subsequent absorbance decay represents addition of water to phenylketene. The chemistry of the system, however, requires ketene hydration to precede enol ketonization in a time sequence opposite from that of the absorbance changes. This seemingly counter-intuitive result is nevertheless consistent with the rate law that governs the time evolution of the central species in a two-step rise and decay, such as that observed here. PMID- 15281831 TI - Synthesis of colloidal Mn2+:ZnO quantum dots and high-TC ferromagnetic nanocrystalline thin films. AB - We report the synthesis of colloidal Mn(2+)-doped ZnO (Mn(2+):ZnO) quantum dots and the preparation of room-temperature ferromagnetic nanocrystalline thin films. Mn(2+):ZnO nanocrystals were prepared by a hydrolysis and condensation reaction in DMSO under atmospheric conditions. Synthesis was monitored by electronic absorption and electron paramagnetic resonance (EPR) spectroscopies. Zn(OAc)(2) was found to strongly inhibit oxidation of Mn(2+) by O(2), allowing the synthesis of Mn(2+):ZnO to be performed aerobically. Mn(2+) ions were removed from the surfaces of as-prepared nanocrystals using dodecylamine to yield high-quality internally doped Mn(2+):ZnO colloids of nearly spherical shape and uniform diameter (6.1 +/- 0.7 nm). Simulations of the highly resolved X- and Q-band nanocrystal EPR spectra, combined with quantitative analysis of magnetic susceptibilities, confirmed that the manganese is substitutionally incorporated into the ZnO nanocrystals as Mn(2+) with very homogeneous speciation, differing from bulk Mn(2+):ZnO only in the magnitude of D-strain. Robust ferromagnetism was observed in spin-coated thin films of the nanocrystals, with 300 K saturation moments as large as 1.35 micro(B)/Mn(2+) and T(C) > 350 K. A distinct ferromagnetic resonance signal was observed in the EPR spectra of the ferromagnetic films. The occurrence of ferromagnetism in Mn(2+):ZnO and its dependence on synthetic variables are discussed in the context of these and previous theoretical and experimental results. PMID- 15281832 TI - Synthesis and optical properties of nanorattles and multiple-walled nanoshells/nanotubes made of metal alloys. AB - The galvanic replacement reaction between silver and chloroauric acid has been exploited as a powerful means for preparing metal nanostructures with hollow interiors. Here, the utility of this approach is further extended to produce complex core/shell nanostructures made of metals by combining the replacement reaction with electroless deposition of silver. We have fabricated nanorattles consisting of Au/Ag alloy cores and Au/Ag alloy shells by starting with Au/Ag alloy colloids as the initial template. We have also prepared multiple-walled nanoshells/nanotubes (or nanoscale Matrioshka) with a variety of shapes, compositions, and structures by controlling the morphology of the template and the precursor salt used in each step of the replacement reaction. There are a number of interesting optical features associated with these new core/shell metal nanostructures. For example, nanorattles made of Au/Ag alloys displayed two well separated extinction peaks, a feature similar to that of gold or silver nanorods. The peak at approximately 510 nm could be attributed to the Au/Ag alloy cores, while the other peak was associated with the Au/Ag alloy shells and could be continuously tuned in the spectral range from red to near-infrared. PMID- 15281833 TI - Gas hydrate single-crystal structure analyses. AB - The first single-crystal diffraction studies on methane, propane, methane/propane, and adamantane gas hydrates SI, SII, and SH have been performed. To circumvent the problem of very slow crystal growth, a novel technique of in situ cocrystallization of gases and liquids resulting in oligocrystalline material in a capillary has been developed. With special data treatment, termed oligo diffractometry, structural data of the gas hydrates of methane, acetylene, propane, a propane/ethanol/methane-mixture and an adamantane/methane-mixture were obtained. Cell parameters are in accord with reported values. Host network and guest are subject to extensive disorder, reducing the reliability of structural information. It was found that most cages are fully occupied by a guest molecule with the exception of the dodecahedral cage in the acetylene hydrate which is only filled to 60%. For adamantane in the icosahedral cage a disordered model is proposed. PMID- 15281834 TI - Highly luminescent, triple- and quadruple-stranded, dinuclear Eu, Nd, and Sm(III) lanthanide complexes based on bis-diketonate ligands. AB - The bis(beta-diketone) ligands 1,3-bis(3-phenyl-3-oxopropanoyl)benzene, H(2)L(1) and 1,3-bis(3-phenyl-3-oxopropanoyl) 5-ethoxy-benzene, H(2)L(2), have been prepared for the examination of dinuclear lanthanide complex formation and investigation of their properties as sensitizers for lanthanide luminescence. The ligands bear two conjugated diketonate binding sites linked by a 1,3-phenylene spacer. The ligands bind to lanthanide(III) or yttrium(III) ions to form neutral homodimetallic triple stranded complexes [M(2)L(1)(3)] where M = Eu, Nd, Sm, Y, Gd and [M(2)L(2)(3)], where M = Eu, Nd or anionic quadruple-stranded dinuclear lanthanide units, [Eu(2)L(1)(4)](2-). The crystal structure of the free ligand H(2)L(1) has been determined and shows a twisted arrangement of the two binding sites around the 1,3-phenylene spacer. The dinuclear complexes have been isolated and fully characterized. Detailed NMR investigations of the complexes confirm the formation of a single complex species, with high symmetry; the complexes show clear proton patterns with chemical shifts of a wide range due to the lanthanide paramagnetism. Addition of Pirkle's reagent to solutions of the complexes leads to splitting of the peaks, confirming the chiral nature of the complexes. Electrospray and MALDI mass spectrometry have been used to identify complex formulation and characteristic isotope patterns for the different lanthanide complexes have been obtained. The complexes have high molar absorption coefficients (around 13 x 10(4) M(-1)cm(-1)) and display strong visible (red or pink) or NIR luminescence upon irradiation at the ligand band around 350 nm, depending on the choice of the lanthanide. Emission quantum yield experiments have been performed and the luminescence signals of the dinuclear complexes have been found to be up to 11 times more intense than the luminescence signals of the mononuclear analogues. The emission quantum yields and the luminescence lifetimes are determined to be 5% and 220 micros for [Eu(2)L(1)(3)], 0.16% and 13 micros for [Sm(2)L(1)(3)], and 0.6% and 1.5 micros for [Nd(2)L(1)(3)]. The energy level of the ligand triplet state was determined from the 77 K spectrum of [Gd(2)L(1)(3)]. The bis-diketonate ligand is shown to be an efficient sensitizer, particularly for Sm and Nd. Photophysical studies of the europium complexes at room temperature and 77 K show the presence of a thermally activated deactivation pathway, which we attribute to ligand-to-metal charge transfer (LMCT). Quenching of the luminescence from this level seems to be operational for the Eu(III) complex but not for complexes of Sm(III) and Nd(III), which exhibit long lifetimes. The quadruple-stranded europium complex has been isolated and characterized as the piperidinium salt of [Eu(2)L(1)(4)](2-). Compared with the triple-stranded Eu(III) complex in the solid state, the quadruple-stranded complex displays a more intense emission signal with a distinct emission pattern indicating the higher symmetry of the quadruple-stranded complex. PMID- 15281835 TI - Structure of a surfactant-templated silicate framework in the absence of 3d crystallinity. AB - The structure of a novel molecularly ordered two-dimensional (2D) silicate framework in a surfactant-templated mesophase has been established by using a combination of solid-state nuclear magnetic resonance (NMR) spectroscopy, X-ray diffraction, and quantum chemical and empirical force-field modeling. These materials are unusual in their combination of headgroup-directed 2D crystalline framework ordering, zeolite-like ring structures within the layers, and long range mesoscopic organization without three-dimensional (3D) atomic periodicity. The absence of registry between the silicate sheets, resulting from the liquidlike disorder of the alkyl surfactant chains, has presented significant challenges to the determination of framework structures in these and similar materials lacking 3D crystalline order. Double-quantum (29)Si NMR correlation experiments establish the interactions and connectivities between distinct intra sheet silicon sites from which the structure of the molecularly ordered inorganic framework is determined. PMID- 15281836 TI - Oxygen gas-induced lip-lip interactions on a double-walled carbon nanotube edge. AB - We have investigated adsorption of an O(2) molecule on a double-walled carbon nanotube (DWCNT) edge using density functional theory calculations. An O(2) molecule adsorbs exothermally without an adsorption barrier at open nanotube edges that are energetically favorable with a large adsorption energy of about -9 eV in most cases. Dissociative adsorption of an O(2) molecule induces various spontaneous lip-lip interactions via the bridged carbon atoms, generating the closed tube ends. This explains why the DWCNTs are chemically more stable than the single-walled nanotubes during observed field emission experiments. The field emission takes place via the localized states of the bridged carbon atoms, not via those of the adsorbed oxygen atoms particularly in the armchair nanotubes. We also find that some O(2) precursor states exist as a bridge between tube edges. PMID- 15281837 TI - Hydrogen bonding versus coordination of adsorbate molecules on Ti-silicalites: a density functional theory study. AB - The present study discusses the results of theoretical calculations obtained at the B3LYP/ 6-31G level on the structural, electronic, and energetic properties of Ti-silicalites. Particularly, the relevance of 5T cluster models, either H- or OH terminated, in large-scale calculations has been critically considered. It was shown that an open surface structure with one OH group and a closed-bulk structure with no bonded OH group at the Ti site are responsible for the observed UV-vis properties of Ti-silicalite materials. Both water and methanol can preferably interact with Ti-silicalites through the H-bonding mechanism, while ammonia can form either H-bonded or coordination complexes. The calculations support the existence of highly dispersed Ti sites in a tetrahedral environment only in Ti-silicalites because an increase in the coordination number of the Ti site by next-neighbor lattice oxygens is the energetically less favorable process. PMID- 15281839 TI - DNA logic gates. AB - A conceptually new logic gate based on DNA has been devised. Methoxybenzodeazaadenine ((MD)A), an artificial nucleobase which we recently developed for efficient hole transport through DNA, formed stable base pairs with T and C. However, a reasonable hole-transport efficiency was observed in the reaction for the duplex containing an (MD)A/T base pair, whereas the hole transport was strongly suppressed in the reaction using a duplex where the base opposite (MD)A was replaced by C. The influence of complementary pyrimidines on the efficiency of hole transport through (MD)A was quite contrary to the selectivity observed for hole transport through G. The orthogonality of the modulation of these hole-transport properties by complementary pyrimidine bases is promising for the design of a new molecular logic gate. The logic gate system was executed by hole transport through short DNA duplexes, which consisted of the "logic gate strand", containing hole-transporting nucleobases, and the "input strand", containing pyrimidines which modulate the hole-transport efficiency of logic bases. A logic gate strand containing multiple (MD)A bases in series provided the basis for a sharp AND logic action. On the other hand, for OR logic and combinational logic, conversion of Boolean expressions to standard sum-of product (SOP) expressions was indispensable. Three logic gate strands were designed for OR logic according to each product term in the standard SOP expression of OR logic. The hole-transport efficiency observed for the mixed sample of logic gate strands exhibited an OR logic behavior. This approach is generally applicable to the design of other complicated combinational logic circuits such as the full-adder. PMID- 15281838 TI - Functional vibrational spectroscopy of a cytochrome c monolayer: SEIDAS probes the interaction with different surface-modified electrodes. AB - Electrochemically induced infrared difference spectra of cytochrome c on various chemically modified electrodes (CMEs) are recorded by exploiting the surface enhancement exerted by a granular gold film. We have recently developed surface enhanced infrared difference absorption spectroscopy (SEIDAS), which provides acute sensitivity to observe the minute enzymatic change of a protein on the level of a monolayer. By these means, we demonstrate that the relative band intensities in the potential-induced difference spectra of adsorbed cytochrome c are significantly dependent on the type of CME used (mercaptopropionic acid, mercaptoethanol, 4,4'-dithiodipyridine, or L-cysteine). These differences are attributed to the altered interaction of cytochrome c with the headgroup of the various CMEs leading to variations in surface orientation and relative distance from the surface. Nevertheless, the peak positions of the observed bands are identical among the CMEs employed. This implies that the internal conformational changes induced by the redox reaction of the adsorbed cytochrome c are not disturbed by the interaction with the CME and that full functionality of the protein is retained. Finally, we critically discuss our results within the framework of the different models for cytochrome c adsorption on CMEs. PMID- 15281841 TI - H atom elimination from the pi sigma* state in the photodissociation of phenol. AB - Photodissociation of phenol at 248 nm was studied using multimass ion imaging techniques. Photofragment translational energy distribution of H atom elimination was measured. The results demonstrate that H atom elimination occurs on the pi sigma(*) excited state which has repulsive potential-energy functions with respect to the stretching of OH bond. It supports the recent ab initio calculation. PMID- 15281842 TI - The role of pi sigma* state in intramolecular electron-transfer dynamics of 4 dimethylaminobenzonitrile and related molecules. AB - Evidence is presented which indicates that the photoinduced intramolecular charge transfer (ICT) in 4-dimethylaminobenzonitrile proceeds by a new mechanism in which pi sigma(C triple bond N) (*) state is the intermediate of a consecutive process that takes the initially excited pi pi(*) state to the fully charge separated ICT state. The absence of the ICT-state formation in 4 aminobenzonitrile is attributed to the smaller electron-donor strength of the amino group relative to the dimethylamino group, which hinders the pi sigma(*)- >ICT charge-shift reaction. PMID- 15281843 TI - A finite temperature linear tetrahedron method for electronic structure calculations of periodic systems. AB - A finite-temperature linear tetrahedron method for electronic structure calculations of periodic systems is developed. When compared to widely used simple temperature broadening, the number of k points necessary for accurate integration at finite temperatures is reduced. The utility of the method is demonstrated with benchmark calculations on 1D, 2D, and 3D systems. PMID- 15281844 TI - Quantum hydrodynamics: application to N-dimensional reactive scattering. AB - The quantum hydrodynamic equations associated with the de Broglie-Bohm formulation of quantum mechanics are solved using a new methodology which gives an accurate, unitary, and stable propagation of a time dependent quantum wave packet [B. K. Kendrick, J. Chem. Phys. 119, 5805 (2003)]. The methodology is applied to an N-dimensional model chemical reaction with an activation barrier. A parallel version of the methodology is presented which is designed to run on massively parallel supercomputers. The computational scaling properties of the parallel code are investigated both as a function of the number of processors and the dimension N. A decoupling scheme is introduced which decouples the multidimensional quantum hydrodynamic equations into a set of uncoupled one dimensional problems. The decoupling scheme dramatically reduces the computation time and is highly parallelizable. Furthermore, the computation time is shown to scale linearly with respect to the dimension N=2,...,100. PMID- 15281845 TI - Second order Moller-Plesset perturbation theory based upon the fragment molecular orbital method. AB - The fragment molecular orbital (FMO) method was combined with the second order Moller-Plesset (MP2) perturbation theory. The accuracy of the method using the 6 31G(*) basis set was tested on (H(2)O)(n), n=16,32,64; alpha-helices and beta strands of alanine n-mers, n=10,20,40; as well as on (H(2)O)(n), n=16,32,64 using the 6-31 + + G(**) basis set. Relative to the regular MP2 results that could be afforded, the FMO2-MP2 error in the correlation energy did not exceed 0.003 a.u., the error in the correlation energy gradient did not exceed 0.000 05 a.u./bohr and the error in the correlation contribution to dipole moment did not exceed 0.03 debye. An approximation reducing computational load based on fragment separation was introduced and tested. The FMO2-MP2 method demonstrated nearly linear scaling and drastically reduced the memory requirements of the regular MP2, making possible calculations with several thousands basis functions using small Pentium clusters. As an example, (H(2)O)(64) with the 6-31 + + G(**) basis set (1920 basis functions) can be run in 1 Gbyte RAM and it took 136 s on a 40 node Pentium4 cluster. PMID- 15281846 TI - Replica-exchange extensions of simulated tempering method. AB - In this paper we consider combinations of two well-known generalized-ensemble algorithms, namely, simulated tempering and replica-exchange method. We discuss two examples of such combinations. One is the replica-exchange simulated tempering and the other is the simulated tempering replica-exchange method. In the former method, a short replica-exchange simulation is first performed and the simulated tempering weight factor is obtained by the multiple-histogram reweighting techniques. This process of simulated tempering weight factor determination is faster and simpler than that in the usual iterative process. A long simulated tempering production run is then performed with this weight factor. The latter method is a further extension of the former in which a simulated tempering replica-exchange simulation is performed with a small number of replicas. These algorithms are particularly useful for studying frustrated systems with rough energy landscape. We give the formulations of these two methods in detail and demonstrate their effectiveness taking the example of the system of a 17-residue helical peptide. PMID- 15281847 TI - Non-Markovian theories based on a decomposition of the spectral density. AB - For the description of dynamical effects in quantum mechanical systems on ultrashort time scales, memory effects play an important role. Meier and Tannor [J. Chem. Phys. 111, 3365 (1999)] developed an approach which is based on a time nonlocal scheme employing a numerical decomposition of the spectral density. Here we propose two different approaches which are based on a partial time-ordering prescription, i.e., a time-local formalism and also on a numerical decomposition of the spectral density. In special cases such as the Debye spectral density the present scheme can be employed even without the numerical decomposition of the spectral density. One of the proposed schemes is valid for time-independent Hamiltonians and can be given in a compact quantum master equation. In the case of time-dependent Hamiltonians one has to introduce auxiliary operators which have to be propagated in time along with the density matrix. For the example of a damped harmonic oscillator these non-Markovian theories are compared among each other, to the Markovian limit neglecting memory effects and time dependencies, and to exact path integral calculations. Good agreement between the exact calculations and the non-Markovian results is obtained. Some of the non-Markovian theories mentioned above treat the time dependence in the system Hamiltonians nonperturbatively. Therefore these methods can be used for the simulation of experiments with arbitrary large laser fields. PMID- 15281848 TI - Interpolation of diabatic potential energy surfaces. AB - A method is presented for constructing diabatic potential energy matrices from ab initio quantum chemistry data. The method is similar to that reported previously for single adiabatic potential energy surfaces, but correctly accounts for the nuclear permutation symmetry of diabatic potential energy matrices and other complications that arise from the derivative coupling of electronic states. The method is tested by comparison with an analytic model for the two lowest energy states of H(3). PMID- 15281849 TI - Optimal control theory for a target state distributed in time: optimizing the probe-pulse signal of a pump-probe-scheme. AB - Optimal control theory (OCT) is formulated for the case of a two-color pump-probe experiment. The approach allows to calculate the pump-pulse shape in such a way that the probe-pulse absorption signal is maximized. Since the latter quantity is given by the time-averaged expectation value of a time dependent operator (the probe-pulse field-strength times the dipole operator) a version of OCT has to be used where the target state is distributed in time. The method is applied to a molecular three-level system with the pump-pulse driving the transition from the electronic ground state into the first-excited electronic state and the probe pulse connecting the first-excited state with a higher lying electronic state. Depending on the probe-pulse duration, the vibrational wave packet becomes localized or at least highly concentrated in the Franck-Condon window for the transition into the higher-excited state. The dependence on the probe-pulse duration and on the delay time between the optimized pump-pulse and the probe pulse is discussed in detail. The whole study demonstrates the feasibility of laser pulse induced temporal wave packet localization and the use of spectroscopic quantities as target states in experiments on femtosecond laser pulse control. PMID- 15281850 TI - Memory effect in friction on a particle caused by a system of fixed or moving scatterers with power law potential. AB - The memory function for friction on a particle caused by a system of fixed or moving scatterers is evaluated for power law interaction. For a dilute system the study extends the steady-state calculation based on the Boltzmann equation to the case of frequency dependence due to the dynamics of the scattering process. For a dense gas an Enskog approximation can be used. The power law potential leads to scaling behavior of the dynamical friction coefficient as a function of reduced mass, coupling coefficient, and energy. PMID- 15281851 TI - Non-Markovian theory of open systems in classical limit. AB - A fully classical limit of the recently published quantum-classical approximation [A. A. Neufeld, J. Chem. Phys. 119, 2488 (2003)] is obtained and analyzed. The resulting kinetic equations are capable of describing the evolution of an open system on the entire time axis, including the short-time non-Markovian stage, and are valid beyond linear response regime. We have shown, that proceeding to the classical mechanics limit we restrict the class of allowed correlations between an open system and a canonical bath, so that the initial conditions and the relaxation operator has to be appropriately modified (projected). Disregard of the projection may lead to unphysical behavior, since mechanism of the decay of some correlations is essentially of quantum-mechanical nature, and is not correctly described by classical mechanics. The projection (quantum correction to the kinetics) is particularly important for the non-Markovian regime of relaxation towards canonical equilibrium. The conformity of the developed method to the conventional approaches is demonstrated using a model of Brownian motion (heavy particle in the bath of light ones), for which the obtained non-Markovian equations are reduced to the standard Fokker-Planck equation in phase space. PMID- 15281852 TI - Laplace-transformed diagonal Dyson correction to quasiparticle energies in periodic systems. AB - We present a method to self-consistently evaluate quasiparticle energies of periodic systems within the diagonal approximation for solving Dyson's equation. Our method is based on the Laplace transform of the second-order Moller-Plesset perturbation (MP2) theory kernel in the atomic basis formulation. The overhead computational cost of evaluating the fully self-consistent diagonal correction over the MP2 band energy calculation is negligible. We present numerical benchmark results for the band structure of trans-polyacetylene and compare it with MP2 and other approaches. PMID- 15281853 TI - Theoretical study of nitric oxide adsorption on Au clusters. AB - The adsorption properties of NO molecule on anionic, cationic, and neutral Au(n) clusters (n=1-6) are studied using the density functional theory with the generalized gradient approximation, and with the hybrid functional. For anionic and cationic clusters, the charge transfer between the Au clusters and NO molecule and the corresponding weakening and elongation of the N-O bond are essential factors of the adsorption. The neutral Au clusters have also remarkable adsorption ability to NO molecule. The adsorption energies of NO on the cationic clusters are generally greater than those on the neutral and anionic clusters. PMID- 15281854 TI - Density and wave function analysis of actinide complexes: what can fuzzy atom, atoms-in-molecules, Mulliken, Lowdin, and natural population analysis tell us? AB - Recent advances in computational methods have made it possible to calculate the wave functions for a wide variety of simple actinide complexes. Equally important is the ability to analyze the information contained therein and produce a chemically meaningful understanding of the electronic structure. Yet the performance of the most common wave function analyses for the calculation of atomic charge and bond order has not been thoroughly investigated for actinide systems. This is particularly relevant because the calculation of charge and bond order even in transition metal complexes is known to be fraught with difficulty. Here we use Mulliken, Lowdin, natural population analysis, atoms-in-molecules (AIM), and fuzzy atom techniques to determine the charges and bond orders of UO(2)(2+), PuO(2)(2+), UO(2), UO(2)Cl(4)(2-), UO(2)(CO)(5)(2+), UO(2)(CO)(4)(2+), UO(2)(CN)(5)(3-), UO(2)(CN)(4)(2-), UO(2)(OH)(5)(3-), and UO(2)(OH)(4)(2-). This series exhibits a clear experimental and computational trend in bond lengths and vibrational frequencies. The results indicate that Mulliken and Lowdin populations and bond orders are unreliable for the actinyls. Natural population analysis performs well after modification of the partitioning of atomic orbitals to include the 6d in the valence space. The AIM topological partitioning is insensitive to the electron donating ability of the equatorial ligands and the relative atomic volume of the formally U(VI) center is counterintuitively larger than that of O(2-) in the UO(2)(2+) core. Lastly, the calibrated fuzzy atom method yields reasonable bond orders for the actinyls at significantly reduced computational cost relative to the AIM analysis. PMID- 15281855 TI - Quasiclassical trajectory study of the collision-induced dissociation of CH3SH+ + Ar. AB - Quasiclassical trajectory calculations were carried out to study the dynamics of energy transfer and collision-induced dissociation (CID) of CH(3)SH(+) + Ar at collision energies ranging from 4.34 to 34.7 eV. The relative abundances calculated for the most relevant product ions are found to be in good agreement with experiment, except for the lowest energies investigated. In general, the dissociation to form CH(3)(+) + SH is the dominant channel, even though it is not among the energetically favored reaction pathways. The results corroborate that this selective dissociation observed upon collisional activation arises from a more efficient translational to vibrational energy transfer for the low-frequency C-S stretching mode than for the high-frequency C-H stretching modes, together with weak couplings between the low- and high-frequency modes of vibration. The calculations suggest that CID takes place preferentially by a direct CH(3)(+) + SH detachment, and more efficiently when the Ar atom collides with the methyl group-side of CH(3)SH(+). PMID- 15281857 TI - The first observation of the rhodium monofluoride molecule: jet-cooled laser spectroscopic studies. AB - Rhodium monofluoride has been observed and spectroscopically characterized. RhF molecules were produced under jet-cooled conditions in a laser vaporization molecular beam source by the reaction of a laser-vaporized rhodium plasma with SF(6) doped in helium, and studied with laser-induced fluorescence spectroscopy under both medium and high resolution. More than 25 bands have been observed in laser-induced fluorescence between 18,500 and 24,500 cm(-1) and five of these have been recorded at 200 MHz resolution. All bands of appreciable intensity have been rotationally analyzed. The ground electronic levels has Omega=2, which is attributed to an inverted (3)Pi state from the 2 delta(4)6 pi(3)12 sigma(1) electron configuration. The ground level rotational constants are B=0.272 45 cm( 1), D=1.035 x 10(-7) cm(-1). Very small ground level Lambda doublings are evident in the spectrum. Excited states having Omega=1, 2, and 3 have been identified. Dispersed fluorescence spectroscopy from 11 excited levels has been used to locate a large number of low-lying vibronic states within the energy range up to 8,000 cm(-1). A ground state vibrational interval of approximately 575 cm(-1) is suggested. PMID- 15281856 TI - H atom transfer along an ammonia chain: tunneling and mode selectivity in 7 hydroxyquinoline.(NH3)3. AB - Excitation of the 7-hydroxyquinoline(NH(3))(3) [7HQ(NH(3))(3)] cluster to the S(1) (1)pi pi(*) state results in an O-H-->NH(3) hydrogen atom transfer (HAT) reaction. In order to investigate the entrance channel, the vibronic S(1)<-->S(0) spectra of the 7HQ.(NH(3))(3) and the d(2)-7DQ.(ND(3))(3) clusters have been studied by resonant two-photon ionization, UV-UV depletion and fluorescence techniques, and by ab initio calculations for the ground and excited states. For both isotopomers, the low-frequency part of the S(1)<--S(0) spectra is dominated by ammonia-wire deformation and stretching vibrations. Excitation of overtones or combinations of these modes above a threshold of 200-250 cm(-1) for 7HQ.(NH(3))(3) accelerates the HAT reaction by an order of magnitude or more. The d(2)-7DQ.(ND(3))(3) cluster exhibits a more gradual threshold from 300 to 650 cm( 1). For both isotopomers, intermolecular vibrational states above the threshold exhibit faster HAT rates than the intramolecular vibrations. The reactivity, isotope effects, and mode selectivity are interpreted in terms of H atom tunneling through a barrier along the O-H-->NH(3) coordinate. The barrier results from a conical intersection of the optically excited (1)pi pi(*) state with an optically dark (1)pi sigma(*) state. Excitation of the ammonia-wire stretching modes decreases both the quinoline-O-H...NH(3) distance and the energetic separation between the (1)pi pi(*) and (1)pi sigma(*) states, thereby increasing the H atom tunneling rate. The intramolecular vibrations change the H bond distance and modulate the (1)pi pi(*)<-->(1)pi sigma(*) interaction to a much smaller extent. PMID- 15281858 TI - Geometry change of simple aromatics upon electronic excitation obtained from Franck-Condon fits of dispersed fluorescence spectra. AB - The S(1) state geometries of benzonitrile, p-cyanophenol, o-cyanophenol, chlorobenzene, and p-chlorophenol were determined by Franck-Condon simulations and a fit of the geometry to the vibronic intensities and effective rotational constants in the harmonic limit based on ab initio force constants. PMID- 15281859 TI - Does chlorine peroxide absorb below 250 nm? AB - Low-lying singlet and triplet electronic excited states of ClOOCl are presented. Calculations of the excitation energies and oscillator strengths are reported using excited state coupled cluster response methods, as well as the complete active space self-consistent field method with the full Breit-Pauli spin-orbit operator. These calculations predict that for ClOOCl there should be a weakly absorbing triplet state lying below the lowest absorbing singlet excited state. This state is predicted to have an absorption maximum at about 385 +/- 25 nm. This lowest triplet state is calculated to be dissociative and leads to ClOO+Cl. PMID- 15281860 TI - Far-infrared absorption by collisionally interacting nitrogen and methane molecules. AB - Quantum line shape calculations of the rototranslational enhancement spectra of nitrogen-methane gaseous mixtures are reported. The calculations are based on a recent theoretical dipole function for interacting N(2) and CH(4) molecules, which accounts for the long-range induction mechanisms: multipolar inductions and dispersion force-induced dipoles. Multipolar induction alone was often found to approximate the actual dipole surfaces of pairs of interacting linear molecules reasonably well. However, in the case of the N(2)-CH(4) pair, the absorption spectra calculated with such a dipole function still show a substantial intensity defect at the high frequencies (>250 cm(-1)) when compared to existing measurements at temperatures from 126 to 297 K, much as was previously reported. PMID- 15281861 TI - One-photon mass-analyzed threshold ionization spectroscopy of 1,3,5 trifluorobenzene: the Jahn-Teller effect and vibrational analysis for the molecular cation in the ground electronic state. AB - One-photon mass-analyzed threshold ionization spectrum of 1,3,5-trifluorobenzene was obtained by using vacuum ultraviolet radiation generated by four-wave difference frequency mixing in Kr. The Jahn-Teller parameters for the e' modes (nu(8)-nu(14)) of 1,3,5-C(6)H(3)F(3)(+) in the ground electronic state needed for spectral analysis were taken from the density functional theory results initially and were upgraded through fits to the experimental results. Excellent agreement was achieved between the experimental and calculated Jahn-Teller energy levels. Assignments of the Jahn-Teller inactive modes were accomplished by referring to the calculated frequencies and the selection rule. The ionization energy of 1,3,5 trifluorobenzene determined from the position of the 0-0 band was 9.6359+/-0.0006 eV. PMID- 15281862 TI - Ab initio potential energy surface and rovibrational spectrum of Ar-HCCCN. AB - We report an ab initio intermolecular potential energy surface of the Ar-HCCCN complex using a supermolecular method. The calculations were performed using the fourth-order Moller-Plesset theory with the full counterpoise correction for the basis set superposition error and a large basis set including bond functions. The complex was found to have a planar T-shaped structure minimum and a linear minimum with the Ar atom facing the H atom. The T-shaped minimum is the global minimum with the well depth of 236.81 cm(-1). A potential barrier separating the two minima is located at R=5.57 A and theta=20.39 degrees with the height of 151.59 cm(-1). The two-dimensional discrete variable representation was employed to calculate the rovibrational energy levels for Ar-HCCCN. The rovibrational spectra including intensities for the ground state and the first excited intermolecular vibrational state are also presented. The results show that the spectra are mostly b-type (Delta K(a)=+/-1) transitions with weak a-type (Delta K(a)=0) transitions in structure, which are in good agreement with the recent experimental results [A. Huckauf, W. Jager, P. Botschwina, and R. Oswald, J. Chem. Phys. 119, 7749 (2003)]. PMID- 15281863 TI - An ab initio study of the hyperfine structure in the X2 Pi electronic state of CCCH. AB - The results of an ab initio study of the magnetic hyperfine structure in the X (2)Pi electronic state of CCCH are reported. The potential surfaces for two components of the X (2)Pi electronic state were computed by means of an extensive configuration interaction approach. The electronically averaged hyperfine coupling constants of H and (13)C for (12)C (12)C (12)CH, (13)C (12)C (12)CH, (12)C (13)C (12)CH, and (12)C (12)C (13)CH are obtained as functions of two bending vibrational modes by the density functional theory method. The vibronic wave functions are calculated with the help of a variational approach which takes into account the Renner-Teller effect and spin-orbit coupling. The model Hamiltonian is expressed in terms of the normal bending coordinates. It is found that, due to the generally strong geometry dependence of the hyperfine coupling constants, it is necessary to carry out the vibronic averaging of the corresponding functions in order to obtain the values which can be compared to the results of the measurements. The results of the present study help to reliably interpret the experimental data previously published. They also predict the yet unobserved hyperfine structure in excited vibronic states. PMID- 15281864 TI - On the ground state of titanium phosphide, TiP: a theoretical investigation. AB - Using multireference variational and coupled cluster methods in conjunction with very large core-correlation-consistent basis sets, we have confirmed that the ground state of TiP is of (2)Sigma(+) symmetry with the first excited state A (2)Delta no more than 3.5 kcal/mol higher. We also report full potential energy curves, dissociation energies, bond lengths, dipole moments, and the usual spectroscopic constants. PMID- 15281865 TI - Electronic structures of exohedral lanthanide-C60 clusters. AB - We have studied the electronic structures of several gas phase exohedral lanthanide (Ln)-C(60) clusters, Ln(n)C(60) (Ln=Pr, Ho, Tb, Tm, Eu, and Yb) with n=1-4, by photoionization spectroscopy of the neutrals and photoelectron spectroscopy of their anions. Both of the spectroscopic analyses reveal that most of the Ln atoms preferably take +3 oxidation states, while Eu atoms alone assume +2 oxidation states, and that C(60) accepts up to twelve donated electrons in Ln(n)C(60). An additional photoionization examination of the oxygen atom mixing into the Ln(n)C(60) clusters demonstrated that each oxygen atom reduces two electrons from C(60). This result implies that the number of accepted electrons in C(60) can be varied by a suitable choice of the number of Ln atoms and O atoms. PMID- 15281866 TI - High-level ab initio calculations for the four low-lying families of minima of (H2O)20. I. Estimates of MP2/CBS binding energies and comparison with empirical potentials. AB - We report estimates of complete basis set (CBS) limits at the second-order Moller Plesset perturbation level of theory (MP2) for the binding energies of the lowest lying isomers within each of the four major families of minima of (H(2)O)(20). These were obtained by performing MP2 calculations with the family of correlation consistent basis sets up to quadruple zeta quality, augmented with additional diffuse functions (aug-cc-pVnZ, n=D, T, Q). The MP2/CPS estimates are -200.1 (dodecahedron, 30 hydrogen bonds), -212.6 (fused cubes, 36 hydrogen bonds), 215.0 (face-sharing pentagonal prisms, 35 hydrogen bonds), and -217.9 kcal/mol (edge-sharing pentagonal prisms, 34 hydrogen bonds). The energetic ordering of the various (H(2)O)(20) isomers does not follow monotonically the number of hydrogen bonds as in the case of smaller clusters such as the different isomers of the water hexamer. The dodecahedron lies ca. 18 kcal/mol higher in energy than the most stable edge-sharing pentagonal prism isomer. The TIP4P, ASP-W4, TTM2-R, AMOEBA, and TTM2-F empirical potentials also predict the energetic stabilization of the edge-sharing pentagonal prisms with respect to the dodecahedron, albeit they universally underestimate the cluster binding energies with respect to the MP2/CBS result. Among them, the TTM2-F potential was found to predict the absolute cluster binding energies to within <1% from the corresponding MP2/CBS values, whereas the error for the rest of the potentials considered in this study ranges from 3% to 5%. PMID- 15281867 TI - Complex oscillations in a simple model for the Briggs-Rauscher reaction. AB - Complex oscillations in a simple model of the Briggs-Rauscher reaction mechanism in a continuously stirred tank reactor proposed by Kim et al. [J. Chem. Phys. 117, 2710 (2002)] are investigated numerically. The k(0)-[CH(2)(COOH)(2)](0) phase diagram is constructed first where k(0) is the flow rate and [...](0) is the input concentration. Within the region surrounded by the Hopf bifurcation curve, we find complex oscillation regions which are again separated from the regular oscillation region by the secondary Hopf bifurcation curves. Mixed mode oscillations with an incomplete Farey sequence, periodic-chaotic (or nonperiodic) sequence, and various types of burst oscillations are observed in complex oscillation regions. Also, chaotic burst oscillations, which are due to the transition from one kind of burst to another kind, are reported. PMID- 15281868 TI - Decoherence in an anharmonic oscillator coupled to a thermal environment: a semiclassical forward-backward approach. AB - The decoherence of an anharmonic oscillator in a thermal harmonic bath is examined via a semiclassical approach. A computational strategy is presented and exploited to calculate the time dependence of the purity and the decay of individual matrix elements in the energy representation for a variety of initial states. The time dependence of the decoherence is found to depend on the temperature of the bath, the coupling strength, the initial state of the oscillator, and the choice of quantity measuring the decoherence. Recurrences in the purity and in the off-diagonal matrix elements are observed, as well as the collapse of these matrix elements to the diagonal, providing evidence for the retention of quantum coherence for time scales longer than that indicated by the purity. The results are used to analyze the utility of the Caldeira-Leggett and Redfield models of decoherence and to assess the dependence of dephasing rates on the degree of structure in phase space. In several cases we find that the dephasing dynamics can be described as an initial Zeno-effect regime, followed by a Caldeira-Leggett region, followed by recurrences. PMID- 15281869 TI - Optimal laser control of ultrafast photodissociation of I2- in water: mixed quantum/classical molecular dynamics simulation. AB - A linearized optimal control method in combination with mixed quantum/classical molecular dynamics simulation is used for numerically investigating the possibility of controlling photodissociation wave packets of I(2)(-) in water. Optimal pulses are designed using an ensemble of photodissociation samples, aiming at the creation of localized dissociation wave packets. Numerical results clearly show the effectiveness of the control although the control achievement is reduced with an increase in the internuclear distance associated with a target region. We introduce effective optimal pulses that are designed using a statistically averaged effective dissociation potential, and show that they semiquantitatively reproduce the control achievements calculated by using optimal pulses. The control mechanisms are interpreted from the time- and frequency resolved spectra of the effective optimal pulses. PMID- 15281870 TI - Near infrared study of water-benzene mixtures at high temperatures and pressures. AB - Near-infrared absorption of water-benzene mixtures has been measured at temperatures and pressures in the ranges of 473-673 K and 100-400 bar, respectively. Concentrations of water and benzene in the water-rich phase of the mixtures were obtained from the integrated absorption intensities of the OH stretching overtone transition of water and the CH stretching overtone transition of benzene, respectively. Using these concentrations, the densities of the water rich phase were estimated and compared with the average densities before mixing, which were calculated from literature densities of neat water and neat benzene. It is found that anomalously large volume expansion on the mixing occurs in the region enclosed by an extended line of the three-phase equilibrium curve and the one-phase critical curve of the mixtures, and the gas-liquid equilibrium curve of water. Furthermore, magnitude of the relative volume change increases with decreasing molar fraction of benzene in the present experimental range. It is suggested that dissolving a small amount of benzene in water induces a change in the fluid density from a liquidlike condition to a gaslike condition in the vicinity of the critical region. PMID- 15281871 TI - UV spectra of benzene isotopomers and dimers in helium nanodroplets. AB - We report spectra of various benzene isotopomers and their dimers in helium nanodroplets in the region of the first Herzberg-Teller allowed vibronic transition 6(0)(1) (1)B(2u)<--(1)A(1g) (the A(0) (0) transition) at approximately 260 nm. Excitation spectra have been recorded using both beam depletion detection and laser-induced fluorescence. Unlike for many larger aromatic molecules, the monomer spectra consist of a single "zero-phonon" line, blueshifted by approximately 30 cm(-1) from the gas phase position. Rotational band simulations show that the moments of inertia of C(6)H(6) in the nanodroplets are at least six times larger than in the gas phase. The dimer spectra present the same vibronic fine structure (though modestly compressed) as previously observed in the gas phase. The fluorescence lifetime and quantum yield of the dimer are found to be equal to those of the monomer, implying substantial inhibition of excimer formation in the dimer in helium. PMID- 15281872 TI - The effective fragment potential: small clusters and radial distribution functions. AB - The effective fragment potential (EFP) method for treating solvent effects provides relative energies and structures that are in excellent agreement with the analogous fully quantum [i.e., Hartree-Fock (HF), density functional theory (DFT), and second order perturbation theory (MP2)] results for small water clusters. The ability of the method to predict bulk water properties with a comparable accuracy is assessed by performing EFP molecular dynamics simulations. The resulting radial distribution functions (RDF) suggest that as the underlying quantum method is improved from HF to DFT to MP2, the agreement with the experimental RDF also improves. The MP2-based EFP method yields a RDF that is in excellent agreement with experiment. PMID- 15281873 TI - Compressibility and specific heats of heavier condensed rare gases near the liquid-vapor critical point. AB - This study concerns the way to describe some physical properties of simple liquids by using the Bogoliubov-Born-Green-Kirkwood-Yvon hierarchy. It constructively analyzes the recent contribution of Sarkisov [G. N. Sarkisov, J. Chem. Phys. 119, 373 (2003)] on the structural behavior of a simple fluid near the liquid-vapor critical point. His work, already compared with computer simulation studies, is here brought into direct contact for the heavier condensed rare gases Ar, Kr, and Xe with (a) experiment and (b) earlier theoretical investigations. Directions for future studies then emerge. PMID- 15281874 TI - Thermodynamic derivations of conditions for chemical equilibrium and of Onsager reciprocal relations for chemical reactors. AB - For an isolated chemical reactor, we derive the conditions for chemical equilibrium in terms of either energy, volume, and amounts of constituents or temperature, pressure, and composition, with special emphasis on what is meant by temperature and chemical potentials as the system proceeds through nonequilibrium states towards stable chemical equilibrium. For nonequilibrium states, we give both analytical expressions and pictorial representations of the assumptions and implications underlying chemical dynamics models. In the vicinity of the chemical equilibrium state, we express the affinities of the chemical reactions, the reaction rates, and the rate of entropy generation as functions of the reaction coordinates and derive Onsager reciprocal relations without recourse to statistical fluctuations, time reversal, and the principle of microscopic reversibility. PMID- 15281875 TI - Raman H-bond pair volume for water. AB - The dispersion of the H-bond pair volume Delta V over the decoupled OD and coupled OH-stretching contours from HDO in H(2)O was determined from Raman intensities at pressures to 9700 bar at 301 K. The dispersion of Delta V was determined from -RT[partial differential ln(I(i)/I(REF))/ partial differential P](T) versus omega (in cm(-1)), where i refers to omega's over the stretching contours and I(REF) refers to the reference intensity at the isosbestic frequency. The maximum H-bond pair volume (defined for breakage) is 1.4+/-0.1 cm(3)/mol H-bond, which corresponds to the volume difference between a large dispersion maximum at 2,675 cm(-1) (near the OD stretch omega of HDO in dense supercritical water) and a large, broad minimum centered near 2,375 cm(-1) (just below the OD stretch omega of HDO in lda ice). The average DeltaV is 0.71+/-0.10 cm(3)/mol H-bond. Other minima near 2,625 cm(-1) (OD) and 3550 cm(-1) (OH) refers to bent H-bonds whose angles are approximately 150 deg. Isothermal pressurization of water lowers the molal volume by decreasing the concentration of long, weak H bonds, and increasing the concentrations of bent H-bonds and short, strong, linear H-bonds. Such bending, shortening, and strengthening produces freezing to ice VI near 10 kbar at 301 K. The isobaric temperature derivative of the maximum H-bond volume is (partial differential Delta V/partial differential T)(P)< or =(2 5) x 10(-3) cm(3)/deg mol H-bond. The OH enthalpy dispersion curve for saturated NaBF(4) in water, yields a large maximum at 3,530-3,540 cm(-1) indicating that BF(4) (-) interacts preferentially with the dangling or "free" OH groups of water thus producing weak, strongly bent H-bonds having angles similar to those of the 3,550 cm(-1) high-pressure H-bond bending feature. PMID- 15281876 TI - The mechanism of N2O formation via the (NO)2 dimer: a density functional theory study. AB - Catalytic formation of N(2)O via a (NO)(2) intermediate was studied employing density functional theory with generalized gradient approximations. Dimer formation was not favored on Pt(111), in agreement with previous reports. On Pt(211) a variety of dimer structures were studied, including trans-(NO)(2) and cis-(NO)(2) configurations. A possible pathway involving (NO)(2) formation at the terrace near to a Pt step is identified as the possible mechanism for low temperature N(2)O formation. The dimer is stabilized by bond formation between one O atom of the dimer and two Pt step atoms. The overall mechanism has a low barrier of approximately 0.32 eV. The mechanism is also put into the context of the overall NO + H(2) reaction. A consideration of the step-wise hydrogenation of O(ads) from the step is also presented. Removal of O(ads) from the step is significantly different from O(ads) hydrogenation on Pt(111). The energetically favored structure at the transition state for OH(ads) formation has an activation energy of 0.63 eV. Further hydrogenation of OH(ads) has an activation energy of 0.80 eV. PMID- 15281877 TI - Study of chemically induced pressure generation of hydrogels under isochoric conditions using a microfabricated device. AB - A method is proposed to study the behavior of stimulus-sensitive hydrogels under isochoric conditions. Freedom of swell movement of such a hydrogel was restricted in all directions by enclosing the hydrogel between a micropressure sensor and a porous cover. Water and external stimuli can be applied to the hydrogel through the pores of the cover to provoke swelling, which results in pressure generation measured by the pressure sensor. The method was put to the proof by examining the response of a pH-sensitive hydrogel to changes in pH, ionic strength, and buffer concentrations of the surrounding solution. Both equilibrium and dynamic pressure generation were observed. The results show that higher pressures are obtained by incorporating more ionizable groups into the hydrogel network or by lowering the ionic strength of the external solution. Furthermore it was proven that pressures reach equilibrium faster when less titratable groups are incorporated or at the presence of higher buffer concentrations in the surrounding solution. By using microfabrication techniques the dimensions of the hydrogel could be kept small with the advantage that responses are fast. A DMAEMA-co-HEMA hydrogel with 2.5% protonable groups and a thickness of 15 microm generated a Delta pressure of 0.67 x 10(5) Pa in 12 min when a pH step from 9 to 6 was applied. The presented method is a simple and fast manner to characterize the static and dynamic stimulus dependent behavior of hydrogels. PMID- 15281878 TI - Study of the hydrostatic pressure dependence of the Raman spectrum of single walled carbon nanotubes and nanospheres. AB - We have investigated the behavior of single-walled carbon nanotubes and nanospheres (C(60)) under high hydrostatic pressure using Raman spectroscopy over the pressure range 0.2-10 GPa using a diamond anvil cell. Different liquid mixtures were used as pressure transmission fluids (PTF). Comparing the pressure dependence of the Raman peak positions for the nanotubes and the nanospheres in different PTF leads to the observation of a number of new phenomena. The observed shift in Raman peak position of both radial and tangential modes as a function of applied pressure and their dependence on the PTF chemical composition can be rationalized in terms of adsorption of molecular species from the of PTF on to the surface of the carbon nanotubes and/or nanospheres. The peak shifts are fully reversible and take place at a comparatively modest pressure (2-3 GPa) that is far below pressures that might be required to collapse the nanoparticles. Surface adsorption of molecular species on the nanotube or nanospheres provides a far more plausible rational for the observed phenomena than ideas based on the notion of tube collapse that have been put forward in the recent literature. PMID- 15281879 TI - Vertical diffusion of water molecules near the surface of ice. AB - We studied diffusion of water molecules in the direction perpendicular to the surface of an ice film. Amorphous ice films of H(2)O were deposited on Ru(0001) at temperature of 100-140 K for thickness of 1-5 bilayer (BL) in vacuum, and a fractional coverage of D(2)O was added onto the surface. Vertical migration of surface D(2)O molecules to the underlying H(2)O multilayer and the reverse migration of H(2)O resulted in change of their surface concentrations. Temporal variation of the H(2)O and D(2)O surface concentrations was monitored by the technique of Cs(+) reactive ion scattering to reveal kinetics of the vertical diffusion in depth resolution of 1 BL. The first-order rate coefficient for the migration of surface water molecules ranged from k(1)=5.7(+/-0.6) x 10(-4) s(-1) at T=100 K to k(1)=6.7(+/-2.0) x 10(-2) s(-1) at 140 K, with an activation energy of 13.7+/-1.7 kJ mol(-1). The equivalent surface diffusion coefficients were D(s)=7 x 10(-19) cm(2) s(-1) at 100 K and D(s)=8 x 10(-17) cm(2) s(-1) at 140 K. The measured activation energy was close to interstitial migration energy (15 kJ mol(-1)) and was much lower than diffusion activation energy in bulk ice (52-70 kJ mol(-1)). The result suggested that water molecules diffused via the interstitial mechanism near the surface where defect concentrations were very high. PMID- 15281880 TI - H/D isotopic exchange between water molecules at ice surfaces. AB - H/D isotopic exchange between H(2)O and D(2)O molecules was studied at the surface of ice films at 90-140 K by the technique of Cs(+) reactive ion scattering. Ice films were deposited on a Ru(0001) substrate in different compositions of H(2)O and D(2)O and in various structures to study the kinetics of isotopic exchange. H/D exchange was very slow on an ice film at 95-100 K, even when H(2)O and D(2)O were uniformly mixed in the film. At 140 K, H/D exchange occurred in a time scale of several minutes on the uniform mixture film. Kinetic measurement gave the rate coefficient for the exchange reaction, k(140 K)=1.6(+/ 0.3) x 10(-19) cm(2) molecule(-1) s(-1) and k(100 K)< or =5.7(+/-0.5) x 10(-21) cm(2) molecule(-1) s(-1) and the Arrhenius activation energy, E(a)> or =9.8 kJ mol(-1). Addition of HCl on the film to provide excess protons greatly accelerated the isotopic exchange reaction such that it went to completion very quickly at the surface. The rapid reaction, however, was confined within the first bilayer (BL) of the surface and did not readily propagate to the underlying sublayer. The isotopic exchange in the vertical direction was almost completely blocked at 95 K, and it slowly occurred only to a depth of 3 BLs from the surface at 140 K. Thus, the proton transfer was highly directional. The lateral proton transfer at the surface was attributed to the increased mobility of protonic defects at the molecularly disordered and activated surface. The slow, vertical proton transfer was probably assisted by self-diffusion of water molecules. PMID- 15281881 TI - Molecular dynamics study of the catalyst particle size dependence on carbon nanotube growth. AB - The molecular dynamics method, based on an empirical potential energy surface, was used to study the effect of catalyst particle size on the growth mechanism and structure of single-walled carbon nanotubes (SWNTs). The temperature for nanotube nucleation (800-1100 K), which occurs on the surface of the cluster, is similar to that used in catalyst chemical vapor deposition experiments, and the growth mechanism, which is described within the vapor-liquid-solid model, is the same for all cluster sizes studied here (iron clusters containing between 10 and 200 atoms were simulated). Large catalyst particles, which contain at least 20 iron atoms, nucleate SWNTs that have a far better tubular structure than SWNTs nucleated from smaller clusters. In addition, the SWNTs that grow from the larger clusters have diameters that are similar to the cluster diameter, whereas the smaller clusters, which have diameters less than 0.5 nm, nucleate nanotubes that are approximately 0.6-0.7 nm in diameter. This is in agreement with the experimental observations that SWNT diameters are similar to the catalyst particle diameter, and that the narrowest free-standing SWNT is 0.6-0.7 nm. PMID- 15281882 TI - In situ high P-T Raman spectroscopy and laser heating of carbon dioxide. AB - In situ high P-T Raman spectra of solid CO(2) up to 67 GPa and 1,660 K have been measured, using a micro-optical spectroscopy system coupled with a Nd:YLF laser heating system in diamond anvil cells. A metallic foil was employed to efficiently absorb the incoming Nd:YLF laser and heat the sample. The average sample temperature was accurately determined by detailed balance from the anti Stokes/Stokes ratio, and was compared to the temperature of the absorber determined by fitting the thermal radiation spectrum to the Planck radiation law. The transformation temperature threshold and the transformation dynamics from the molecular phases III and II to the polymeric phase V, previously investigated only by means of temperature quench experiments, was determined at different pressures. The P-T range of the transformation, between 640 and 1,100 K in the 33 65 GPa pressure interval, was assessed to be a kinetic barrier rather than a phase boundary. These findings lead to a new interpretation of the high P-T phase diagram of carbon dioxide. Furthermore, our approach opens a new way to perform quantitative in situ Raman measurements under extremely high pressures and temperatures, providing unique information about phase relations and structural and thermodynamic properties of materials under these conditions. PMID- 15281883 TI - Comparison of random-walk density functional theory to simulation for bead-spring homopolymer melts. AB - Density profiles for a homopolymer melt near a surface are calculated using a random-walk polymeric density functional theory, and compared to results from molecular dynamics simulations. All interactions are of a Lennard-Jones form, for both monomer-monomer interactions and surface-monomer interactions, rather than the hard core interactions which have been most investigated in the literature. For repulsive systems, the theory somewhat overpredicts the density oscillations near a surface. Nevertheless, near quantitative agreement with simulation can be obtained with an empirical scaling of the direct correlation function. Use of the random phase approximation to treat attractive interactions between polymer chains gives reasonable agreement with simulation of dense liquids near neutral and attractive surfaces. PMID- 15281884 TI - Formation and structure of the microemulsion phase in two-dimensional ternary AB+A+B polymeric emulsions. AB - We present an analysis of the structure of the fluctuation-induced microemulsion phase in a ternary blend of balanced AB diblock copolymers with equal amounts of A and B homopolymers. To this end, graphical analysis methods are employed to characterize two-dimensional configuration snapshots obtained with the recently introduced field-theoretic Monte Carlo method. We find that a microemulsion forms when the mean curvature diameter of the lamellar phase coincides roughly with the periodicity of the lamellar phase. Further, we provide evidence to the effect of a subclassification of the microemulsion into a genuine and a defect-driven region. PMID- 15281885 TI - Microphase separation of weakly charged block polyelectrolyte solutions: Donnan theory for dynamic polymer morphologies. AB - A mean-field dynamic density functional theory for the phase behavior of concentrated weakly charged block polyelectrolyte solutions is developed, using the Donnan membrane equilibrium approach to account for electrostatic interactions. In this limit all long-range electrostatic interactions are canceled and the net charge density in any region on a coarse-grained scale is zero. The phase diagram of a model triblock polyelectrolyte in solution as a function of the charge of the solvophilic block and the solvent concentration is established. Different mesoscopic structures (lamellar, bicontinuous, hexagonal, micellar, and dispersed coexisting phases) are formed depending on the copolymer charge asymmetry. It is found that upon changing the charge of the solvophilic copolymer block the polyelectrolyte solution does not follow the lyotropic sequence of phases of this polymer. Upon increase in the charge of the solvophilic blocks, changes in copolymer morphology take place by means of change in curvature of polymeric domains. PMID- 15281886 TI - Dynamical heterogeneities close to a colloidal gel. AB - Dynamical heterogeneities in a colloidal fluid close to gelation are studied by means of computer simulations. A clear distinction between some fast particles and the rest, slow ones, is observed yielding a picture of the gel composed of two populations with different mobilities. Analyzing the statics and dynamics of both sets of particles, it is shown that the slow particles form a network of stuck particles, whereas the fast ones are able to move over long distances. Correlation functions show that the environment of the fast particles relaxes much faster than that of the slow ones, but at short times the bonds between fast particles are longer lived due to the flexibility of their structure. No stringlike motion is observed for the fast particles, but they occupy preferential sites in the surface of the structure formed by the slow ones. PMID- 15281887 TI - Characterization of polyethylene crystallization from an oriented melt by molecular dynamics simulation. AB - Molecular dynamics is used to characterize the process of crystallization for a united atom model of polyethylene. An oriented melt is produced by uniaxial deformation under constant load, followed by quenching below the melting temperature at zero load. The development of crystallinity is monitored simultaneously using molecular-based order parameters for density, energy, and orientation. For crystallization temperatures ranging from 325 to 375 K, these simulations clearly show the hallmarks of crystal nucleation and growth. We can identify multiple nucleation events, lamellar growth up to the limit imposed by periodic boundaries of the simulation cell, and lamellar thickening. We observe a competition between the rate of nucleation, which results in multiple crystallites, the rate of chain extension, which results in thicker lamellae, and the rate of chain conformational relaxation, which is manifested in lower degrees of residual order in the noncrystalline portion of the simulation. The temperature dependence of lamellar thickness is in accord with experimental data. At the higher temperatures, tilted chain lamellae are observed to form with lamellar interfaces corresponding approximately to the [201] facet, indicative of the influence of interfacial energy. PMID- 15281888 TI - Interdiffusion in a polydisperse polymer blend. AB - We present a theoretical description of interdiffusion in a binary blend of polymers that exhibit polydispersity in length. The diffusion equations are formulated in terms of the volume fractions and the chain concentrations of the components. This choice of variables is equivalent to the assumption that the local molecular weight distributions of the components are described by the Flory distribution. The Onsager kinetic coefficients are obtained based on the Green Kubo equation and correspond to the fast-mode interdiffusion theory. As demonstrated by numerical simulations, the resulting equations describe the simultaneous processes of the evolution of blend composition and the relaxation of the local molecular weight distributions of the components. The developed approach can be used to study polymer systems in which the degree of polymerization changes due to interfacial or bulk chemical reactions. PMID- 15281889 TI - Molecular and cellular mechanisms in the pathophysiology of severe head injury. AB - The pathophysiological process following traumatic brain injury is extremely complex and not fully understood. Recent developments have further advanced our knowledge of the cellular and molecular mechanisms that cause this damage. The excitotoxic damage, alterations in calcium homeostasis and free radical induced damage are thought to be the key pathways in this process. It is believed that the final target of all these pathways is the mitochondria, through the alteration in the mitochondrial permeability transition pore. Moreover, the inflammatory response may be important in the exacerbation of secondary damage but its exact role is not very well known. Further advances in our understanding of the cellular and molecular mechanisms will be crucial in the design of new therapies that should improve the prognosis of the traumatic brain injury patients. PMID- 15281890 TI - Microdialysis in neurointensive care. AB - Microdialysis is a technique for sampling the chemistry of the interstitial fluid of tissues and organs in animal and man. It is minimally invasive and simple to perform in a clinical setting. Although microdialysis samples essentially all small molecular substances present in the interstitial fluid the use of microdialysis in neurointensive care has focused on markers of ischemia and cell damage. The lactate/pyruvate ratio is a well-known marker of changes in the redox state of cells caused by ischemia Glycerol is an integral component of cell membranes. Loss of energy due to ischemia eventually leads to an influx of calcium and a decomposition of cell membranes, which liberates glycerol into the interstitial fluid. Thus the lactate/pyruvate ratio and glycerol have become the most important markers of ischemia and cell membrane damage. While the primary insult at the site of the accident is beyond our control, secondary insults during intensive care should be avoided by all means. Therefore, the single most important finding from microdialysis studies is the dramatic difference in the vulnerability of the penumbra surrounding a lesion as compared to normal brain tissue allowing early detection of secondary insults after traumatic brain injury as well as the onset of vasospasm after subarachnoid hemorrhage. PMID- 15281891 TI - Importance of aquaporins in the physiopathology of brain edema. AB - Aquaporins (AQPs) are water channels that mediate the efficient movement of water across the membrane. Among different AQPs, AQP4 is the predominant water channel in the brain and is thought to play a significant role in the physiopathology of brain edema. Brain edema is a major clinical problem since it largely accounts for the cause of death in patients suffering from traumatic brain injury or cerebrovascular accidents. In this review, we have tried to summarize comprehensive information about the pathological events conducting to brain edema. In addition, we also have summarized active investigation in the field of AQPs and cerebral edema to find plausible explanations and to deduce potential future directions for the treatment of this clinical condition. PMID- 15281892 TI - Arterial hyperoxia in severe head injury: a useful or harmful option? AB - There is mounting evidence both from experimental and clinical studies that traumatic brain injury (TBI) is associated with a reduction of aerobic metabolism. This results from a variable combination of impaired substrate delivery and mitochondrial failure. Mitochondria, which are responsible for the production of 95% of cell adenosine triphosphate (ATP), may become compromised after TBI. On the other hand, in the very early period after the primary injury, oxygen delivery may be impaired due to arterial hypoxia and/or to a reduction of cerebral blood flow (CBF). As a consequence, 80-90% of patients who die of head injury show ischemia on histo-pathological examination of the brain tissue. In the absence of an appropriate treatment for TBI, these observations favored the hypothesis that manipulation of brain oxygen delivery could be a therapeutic tool to improve aerobic metabolism. Several strategies were developed, including the increase of cerebral perfusion pressure (CPP) using amines or the increase of arterial partial pressure of oxygen (PaO(2)) through hyperbaric oxygen (HBO) or normobaric hyperoxia. Several experimental and clinical studies, using normobaric hyperoxia, demonstrated an increase in brain tissue oxygen tension and a reduction of brain extracellular lactate levels, but there is no consensus about the biological meaning of these findings. For some authors, they translate an improvement of brain oxidative metabolism, while for others they represent only biological epiphenomena. The current review addresses the rational behind normobaric hyperoxia as a therapeutic application and discusses the experimental and clinical results achieved so far. PMID- 15281893 TI - Cannabinoids as neuroprotective agents in traumatic brain injury. AB - The name Cannabinoid applies to a large and diverse family of compounds including plant derived, synthetic and endogenously produced chemicals, some but not all of which are psychotropic. Cannabinoids of all classes have the ability to protect neurons from a variety of insults that are believed to underlie delayed neuronal death after traumatic brain injury (TBI), including excitotoxicity, calcium influx, free radical formation and neuroinflammation. The pathways and experimental models supporting a neuroprotective role for the various classes of cannabinoids are critically reviewed vis a vis their potential to support the development of a clinically viable neuroprotective agent for human TBI. PMID- 15281894 TI - Endothelins and the role of endothelin antagonists in the management of posttraumatic vasospasm. AB - Increasing evidence implicates endothelin in the pathophysiological development of cerebral vasospasm. This review summarizes background topics such as the structures and biosynthesis of endothelins, the types of endothelin and their receptors, as well as their biological effects. Basic science and clinical observations supportive of the role of endothelins in the spasm associated with stroke and subarachnoid hemorrhage are presented. Similar basic science and clinical observations are presented regarding the role of endothelins in impaired cerebral hemodynamics following traumatic brain injury, cryogenic cortical lesion injury and fluid percussion brain injury. The role of age in the contribution of endothelin to impaired cerebral hemodynamics following fluid percussion brain injury is discussed. Finally, potential mechanisms for endothelin contributions to vasospasm following fluid percussion brain injury such as impaired K+ channel function are described. PMID- 15281895 TI - Moderate hypothermia in the management of severe traumatic brain injury: a good idea proved ineffective? AB - Many drugs with proven efficacy in the preclinical stage have failed to show any benefit in improving the outcome of severe traumatic brain injury (TBI) when tested in controlled clinical trials. Hypothermia is still the most powerful neuroprotective method in experimental models of TBI. Its ability to influence the multiple biochemical cascades that are set in motion after TBI is quite unique. In experimental models hypothermia protects against mechanical neuronal and axonal injury and improves behavioral outcome. Encouraging results from phase II and III clinical trials of hypothermia in TBI reported in the 1990s generated great enthusiasm. However, enthusiasm faded in 2001 after the final report of the multicenter phase III trial in which the neuroprotective effects of moderate hypothermia in TBI were formally tested. This study found no significant effect on outcome in the hypothermia group, leading many clinicians to lose interest in this therapy. The present article reviews the historical background of the use of hypothermia, presents the rationale for using both immediate and deferred hypothermia, and summarizes both experimental and clinical evidence supporting its potential benefits in the management of severe TBI. New technologies using intravascular methods to induce fast hypothermia have recently become available. Cooling either through the intravenous or intra-arterial route is an exciting alternative with great potential. We argue that moderate hypothermia is still the most powerful neuroprotective candidate for severe TBI and that it merits further research and discussion. We also defend the need for further clinical trials to prove or refute its potential for treating high intracranial pressure refractory to first level therapeutic measures. The premature abandonment of hypothermia could close new avenues for improving the devastating effects of TBI. PMID- 15281896 TI - Drug therapy in schizophrenia. AB - Over 40 different antipsychotic medications have been introduced around the world, 21 of which are available in the United States. The conventional antipsychotic drugs introduced in late 50s have two major groups of disadvantages, efficacy and safety. All of the atypical antipsychotic agents have higher 5-HT(2) blocking than D(2) blocking. Atypical antipsychotic agents differ in their receptor action and side effect profile. Among them, clozapine has superior efficacy, and both clozapine and olanzapine have a higher propensity to cause weight gain and possibly diabetes. Quetiapine is difficult to use in acute psychotic states as a result of titration. Ziprasidone and aripiprazole are less sedating, and diabetes as well as weight gain have not been reported with their use. In an acute setting, antipsychotic monotherapy in therapeutic doses is the most useful. AAP drugs are preferred because of the lack of acute EPS symptoms. Intramuscular preparations of haloperidol and ziprasidone are sometimes required to treat acute patients. The goal in acute treatment is to prevent harm to self or others by decreasing excitatory symptoms. Continuing the antipsychotic medication treatment after the acute symptoms are controlled reduces the likelihood of a relapse. The neuroleptic medication should be continued indefinitely. The minimum amount antipsychotic drugs necessary to prevent a relapse should be used, based on clinical decision. PMID- 15281897 TI - Side effects of atypical antipsychotic drugs. AB - Recently there has been increased concern over the side effects of the atypical antipsychotic drugs, including diabetes, hyperlipidemia and obesity. The relationship between diabetes and antipsychotic drugs requires a careful analysis. Patients with schizophrenia are known to suffer from diabetes more often than the general population. In addition, a number of case reports indicate that the conventional antipsychotic as well as atypical antipsychotic drugs produce diabetes. Clozapine and olanzapine, in particular, have been implicated producing diabetes as well as diabetic ketoacidosis. Epidemiological surveys have supplemented the case reports, finding increased incidence of diabetes in patients treated with atypical antipsychotic agents, but these surveys have not yielded consistent results regarding the differential effects of the various atypical antipsychotic drugs. The mechanism by which antipsychotic agents produce diabetes is not elucidated. Weight gain and consequent alteration in triglycerides and cholesterol have been known to occur frequently with olanzapine and clozapine. The ensuing metabolic syndrome itself may cause insulin resistance and diabetes. In the absence of definitive scientific data on the differential effects of antipsychotic drugs in inducing diabetes, clinical prudence and careful monitoring of all patients on atypical antipsychotic drugs is necessary. Aripiprazole and ziprasidone have not been shown to increase weight or produce diabetes, but more information on the diabetogenic effects of ziprasidone and aripiprazole is needed. In order to assess the differential effects of atypical antipsychotic drugs in producing diabetes and the mechanisms by which they produce this reaction, further research is necessary. PMID- 15281898 TI - Antipsychotic polypharmacy. AB - The administration of more than one drug for a single medical condition is considered to be polypharmacy. There are many possible reasons for polypharmacy: (1) psychosis is a chronic disease that cannot be cured; (2) expectations to improve patients' quality of life beyond what drugs can actually do is high; (3) the lack of side effects and interactions can cause physicians to be more daring in terms of potential complications; (4) information from the Internet may cause patients and their families to demand medications; (5) the diluted mental health system allows legal guardians and other mental health professionals to force physicians to provide multiple drugs; (6) many new drugs are available; and (7) physicians are forced to shorten hospitalization days. The 1997 American Psychiatric Association Practice Research Network found that 17% of 146 patients with schizophrenia were treated concurrently with more than one antipsychotic medication. Polypharmacy may increase the risk of adverse effects, drug interactions, noncompliance, and medication errors. It is not wise to use polypharmacy only to prevent side effects and drug and interactions. Our attempts to reduce polypharmacy may fail, as academicians also propagate polypharmacy, and all of the algorithms indicate polypharmacy as an option, putting physicians in a legal and ethical bind. Techniques such as experimental ward, peer review, computer information feedback, and comparing different techniques may temporarily reduce polypharmacy but long-term outcome is not affected. Scientific data on the efficacy of polypharmacy is needed in order to sort out good and bad polypharmacy. PMID- 15281899 TI - The use of antiparkinsonian agents in the management of drug-induced extrapyramidal symptoms. AB - Antipsychotic drugs induce extrapyramidal symptoms such as dystonia, akathisia and parkinsonian symptoms early in treatment, and tardive dyskinesia later in treatment. With the advent of atypical antipsychotic drugs, the incidence of extrapyramidal symptoms has decreased, but the danger still exists. There are many reasons that extrapyramidal symptoms are still a problem. Most often, psychiatrists use doses higher than the recommended dose of atypical antipsychotic agents. For example, the use of 8 to 10 mg of risperidone, 30 to 40 mg of olanzapine, or 1200 to 1500 mg of quetiapine daily is not uncommon. In addition, combinations of both conventional and atypical antipsychotic drugs are used together in many instances. Extrapyramidal symptoms produce unnecessary suffering and add to the health burden; therefore, prompt recognition of these symptoms is necessary. If EPS occur, it is of paramount importance to start an antiparkinsonian agent immediately to provide relief to the patient. In high-risk patients, prophylactic antiparkinsonian therapy is indicated but routine prophylaxis with antiparkinsonian agents is harmful. As only a segment of patients may develop EPS, many patients receive prophylactic medication unnecessarily, and the side effects of antiparkinsonian drugs prescribed without clinical indication may add to the health burden of the patient. If prophylactic antiparkinsonian treatment is initiated, it should be discontinued at least two weeks after its initiation. The long term use of antiparkinsonian treatment is not therapeutically beneficial to the patient, and studies indicate that the gradual withdrawal of antiparkinsonian medication will not produce recurrence of EPS. PMID- 15281900 TI - Treatment of patients with schizophrenia and substance abuse disorders. AB - Approximately half of patients with schizophrenia have a lifetime diagnosis of substance abuse disorders. These dual diagnosis patients are more likely to have poorer outcomes, including more severe psychiatric symptoms with increased hospitalizations, higher utilization of services and frequent homelessness. Assessment and treatment of dually diagnosed patients has evolved over the last twenty years. To date, the strongest evidence for effective management of dual diagnosis patients has been utilization of integrated treatment services, which combines both mental health and substance abuse treatments concurrently. Strategies commonly used include a combination of pharmacological treatment, intensive case management, motivational interviewing, individual and group psychotherapy, and family participation. This chapter summarizes the treatment options available for this population. PMID- 15281901 TI - Treatment considerations in the forensic patient with schizophrenia. AB - Research in new treatments for schizophrenia continues. At the same time, increasing numbers of persons with schizophrenia receive their treatment almost exclusively in correctional settings. Though the literature continues to describe this phenomenon in clearer detail, many questions regarding the characteristics of these mentally disordered offenders and their unique treatment needs remain. Research targeting this growing subset of persons with schizophrenia is limited. Risk factors that lead the person with schizophrenia into the correctional system, and whether the offenders with schizophrenia actually differ from non offenders in terms of their psychopharmacological needs are significant questions that remain unaddressed. Countertransference towards the offender remains a significant issue as well. Given the many limitations on research in this population, pharmacologic strategies for the offender population must be extrapolated from the non-offender population. Traditional depot neuroleptics continue to be underutilized in the treatment of this population. It is argued that both traditional and forthcoming atypical depot neuroleptics may be the best first line agents in this population. PMID- 15281902 TI - Cognitive therapy for schizophrenia: treatment and dissemination. AB - Although medication is the treatment of choice for schizophrenia, most patients continue to experience exhibit residual symptoms despite compliance. Recent studies suggest that cognitive behavioral therapy (CBT) is useful in the treatment of schizophrenia and other psychotic disorders [1]. Results of previous studies suggest that patients treated with CBT, in conjunction with medication, exhibit decreased frequency and severity of delusions, hallucinations and negative symptoms. Finally, CBT has been shown to increase medication compliance in patients with schizophrenia and bipolar disorder. Speaking of the outcome data on CBT, Weiden was recently quoted as saying, "if this were a patented drug trial, you would have heard about [these findings]" [2]. Despite the empirical evidence supporting the use of CBT in patients with schizophrenia, this treatment is relatively unknown to the psychiatric community. This article will review the mechanisms and findings associated with CBT and schizophrenia, and suggest future directions for dissemination in the psychiatric community. PMID- 15281903 TI - Rehabilitation of schizophrenic patients. AB - Schizophrenia is a maldevelopmental disorder of the brain that manifests in positive, negative, cognitive and affective symptoms. Currently, the mainstay of treatment involves pharmacotherapy. The limitations of antipsychotic treatment are that they can only control symptoms and cannot cure the illness, and 20% of patients do not respond, thus leading to the requirement of maintenance treatment. Patients that do respond continue to have disabling residual symptoms such as amotivation and isolation, maladaptive behavior, and impaired social functioning. These symptoms prevent patients from attaining educational, occupational, and social roles. Psychosocial interventions and models of quality of life in schizophrenia are based on the notion that increases in psychosocial functioning will be related to improvement in subjective experiences, such as self-esteem and satisfaction with life. The comparative effect of specific treatment methods and the additional benefits of multiple treatments need to be explored. Diversified techniques have also been employed, such as shaping, cognitive process therapy, mastery-oriented skill training, motivation and enhancement. Issues in designing psychosocial interventions and the role of various professionals in providing such interventions need to be carefully considered. Predictor variables and the indications for particular therapies in an individual need to be explored. Generalizability of the gains made by rehabilitation/recovery is also an important consideration. Patients in jail, chronic mental hospitals, private facilities, and the Veterans Administration system are all different in their ability to benefit, their motivations, and the severity of their psychopathology. Therefore, it is very difficult to generalize findings from one setting to another. PMID- 15281904 TI - Chromosomal aberrations and genomic instability induced by topoisomerase-targeted antitumour drugs. AB - The present review discusses recent evidence on the mechanisms of formation of structural and numerical chromosome aberrations by anti-topoisomerase drugs. Among "cleavable complex"poisoning drugs, DNA topoisomerase II inhibitors induce DNA double strand breaks that lead to chromosomal aberrations independently of the phase of the cell cycle in which the treatment has been performed. Inhibitors of DNA topoisomerase I induce DNA single strand breaks that are transformed in DSB when the trapped "cleavable complex" collides with the replication fork, producing chromatid-type aberrations. Recently, ongoing chromosome condensation and RNA transcription have been shown to play a crucial role in the formation of chromatid-type aberrations by topoisomerase I poisons for treatments in the G(2) phase of the cell cycle. Mutations of single genes are also induced by anti topoisomerase drugs. These consist mostly of deletions, duplications and insertions and are often localized at the topoisomerase cleavable sites. This suggests that alterations at the chromatin level may be responsible for inactivation of gene function after topoisomerase inhibitors. Anti-topoisomerase drugs promote also numerical chromosome aberrations as DNA topoisomerases are involved in chromosome condensation and segregation at mitosis. Polyploid cells are induced as a consequence of the total inhibition of sister chromatid separation before anaphase and aneuploid cells may arise when sister chromatid separation is defective. Gene mutations, chromosomal aberrations and aneuploidy may influence the stability of the genome further producing structural aand numerical aberrations at successive cell cycle divisions. Knowledge of the mechanisms producing gene mutations, chromosome aberrations and genomic instability after drugs interacting with topoisomerases is essential for developing effective therapeutical approaches. PMID- 15281905 TI - Topoisomerase I-DNA complex stability induced by camptothecins and its role in drug activity. AB - The mechanism of cytotoxicity of the camptothecin family of antitumor drugs is thought to be the consequence of a collision between moving replication forks and camptothecin-stabilized cleavable DNA-topoisomerase I complexes. One property of camptothecin analogs relevant to their potent antitumor activity is the slow reversal of the cleavable complexes formed with these drugs. The persistence of cleavable complexes with time may be an essential property for increasing the likelihood of a collision between the replication fork and a cleavable complex, giving rise to lethal DNA lesions. In this paper, we examined a number of camptothecin analogs forming cleavable complexes with distinctly different stabilities. Absolute reaction rate analysis was carried out for each derivative. Our results indicate that the stability of the cleavable complex is dominated by the activation entropy (DeltaS++) of the reversal process. We measured the relative lipophilicity of the CPT analogs by reverse-phase HPLC, but the DeltaS++ of complex reversal is not directly related to the lipophilicity of the CPT analog being used. We suggest that solvent ordering around the 7- through 10 position of the CPT ring may be responsible for reversal rate's dependence on DeltaS++. We demonstrate that the cleavable complex stability conferred by each camptothecin analog is directly correlated with the induction of apoptosis and cytotoxicity to tumor cells. PMID- 15281906 TI - Development of DNA topoisomerase-related therapeutics: a short perspective of new challenges. AB - Antitumor agents targeting DNA and DNA-associated processes are widely used in the treatment of human cancers and produce significant increases in the survival of patients. DNA topoisomerases remain the most significant target of these cytotoxic drugs and constitute a growing family of nuclear enzymes that regulate DNA topology during DNA replication and recombination, DNA transcription, chromosome condensation-decondensation and segregation. Major progress has been attained in recent years in the understanding of the structures of these enzymes and their main cellular functions, hopefully providing new opportunities for pharmacological interventions. New leads and derivatives of known structures have been reported recently, and here they will be discussed highlighting the challenges to find innovative and more effective drugs. Moreover, we will review novel and diverse approaches relevant to the development of new topoisomerase related therapeutics. PMID- 15281907 TI - Recent advances in experimental molecular therapeutics for malignant gliomas. AB - The current lack of effective therapy for malignant gliomas has prompted the development of three primary foci of molecular research: anti-angiogenesis therapy, immunotherapy, and DNA- and RNA-based therapies. Angiogenesis inhibitors, designed to exploit the highly vascularized nature of gliomas, target endothelial cells and/or the extracellular matrix and bypass many of the problems of conventional chemotherapy. There may be easy access to the molecular target (e.g. blood vessels), reduced induction of drug resistance, and general lack of host toxicity. The relatively immunoprivileged status of the brain has also prompted use of immune stimulation as an anti-glioma strategy. Lines of attack include global cytokine therapy, vaccination with specific tumor antigens, dosing with monoclonal antibodies conjugated to radioisotopes or toxins, and ex vivo priming of lymphocytes. With regard to DNA- and RNA-based therapy, numerous oncogenic proteins have been targeted by antisense molecules administered alone or in combination with conventional chemotherapy and radiation. In one tactic, termed "suicide" gene therapy, herpes simplex thymidine kinase has been transfected into glioma cells via a retrovirus; subsequent introduction of ganciclovir causes cytotoxicity in the transduced cells. Although considerable preclinical data have been accumulated, promising results for therapy of human glioma have only recently appeared. PMID- 15281908 TI - Lamellarins, from A to Z: a family of anticancer marine pyrrole alkaloids. AB - The lamellarins form a group of more than 30 polyaromatic pyrrole alkaloids, isolated from diverse marine organisms, mainly but not exclusively ascidians and sponges. These molecules fall in three structural groups, with the central pyrrole ring fused or unfused (lamellarins O-R) to adjacent aromatic rings and with the quinoline moiety containing a 5, 6-single--as in lamellarins I-L--or a double bond, as it is the case for lamellarins D and M which are both potent cytotoxic agents. The family also includes sulphated members, such as the integrase inhibitor lamellarin alpha 20-sulfate. This review presents the origin and structure of the lamellarins and summarizes the various chemical pathways which have been proposed to synthesize all lamellarins and different structurally related marine pyrrole alkaloids, including ningalins, storniamides and lukianols. The mechanisms of actions of these marine products are also discussed. Inhibition of HIV-1 integrase by lamellarin alpha 20-sulfate and human topoisomerase I by lamellarin D and Molluscum contagiosum virus topoisomerase by lamellarin H, along with other effects on nuclear proteins, provide an experimental basis indicating that DNA manipulating enzymes are important targets for the lamellarins. Some of these marine compounds exhibit cytotoxic activities against tumor cells in vitro and are insensitive to Pgp-mediated drug efflux. The structure-activity relationships are discussed. Other compounds in the series, without being strongly cytotoxic, can reverse the multidrug resistance phenotype and thus may be useful to promote the therapeutic activity of conventional cytotoxic drugs toward chemoresistant tumors. A complete description of the chemistry and pharmacological profiles of the lamellarins is presented here to shed light on this undervalued family of marine alkaloids. PMID- 15281909 TI - Effect of prostaglandins on the regulation of tumor growth. AB - Prostaglandins (PGs) are involved in mediating or regulating many physiological as well as pathological processes. Important roles of PGs in the pathophysiology of carcinogenesis offer potentials of targeting PG synthesis and PG receptors in developing novel anti-cancer therapy. Although initial studies suggested direct growth inhibitory role of PGs from in vitro studies, it has been widely demonstrated that in general, PGs stimulate tumor growth. However, cyclopentenone PGs, especially 15d-PGJ2, which can activate peroxisome proliferator activated receptor (PPAR) gamma, exhibited anti-proliferative and proapoptotic effects on many types of cancer cells. But recent studies indicate that growth inhibitory effects of the cyclopentenone PGs might also be a nonspecific effect due to its highly reactive cyclopentenone ring. We have explored the published studies on PGs to specify its known regulatory roles on tumor growth with an objective of targeting the PGs or pathways activated by these lipids in treating cancers. PMID- 15281910 TI - Proteomic analysis for the assessment of diabetic renal damage in humans. AB - Renal disease in patients with Type II diabetes is the leading cause of terminal renal failure and a major healthcare problem. Hence early identification of patients prone to develop this complication is important. Diabetic renal damage should be reflected by a change in urinary polypeptide excretion at a very early stage. To analyse these changes, we used an online combination of CE/MS (capillary electrophoresis coupled with MS), allowing fast and accurate evaluation of up to 2000 polypeptides in urine. Employing this technology, we have examined urine samples from 39 healthy individuals and from 112 patients with Type II diabetes mellitus and different degrees of albumin excretion rate. We established a 'normal' polypeptide pattern in the urine of healthy subjects. In patients with Type II diabetes and normal albumin excretion rate, the polypeptide pattern in urine differed significantly from normal, indicating a specific 'diabetic' pattern of polypeptide excretion. In patients with higher grade albuminuria, we were able to detect a polypeptide pattern indicative of 'diabetic renal damage'. We also found this pattern in 35% of those patients who had low-grade albuminuria and in 4% of patients with normal albumin excretion. Moreover, we could identify several of the indicative polypeptides using MS/MS sequencing. We conclude that proteomic analysis with CE/MS permits fast and accurate identification and differentiation of polypeptide patterns in urine. Longitudinal studies should explore the potential of this powerful diagnostic tool for early detection of diabetic renal damage. PMID- 15281911 TI - Sulpho-conjugation of ethanol in humans in vivo and by individual sulphotransferase forms in vitro. AB - We studied whether ethanol is sulphonated in humans with the perspective of using the urinary excretion of ethyl sulphate after ethanol consumption as a biomarker for SULT (sulphotransferase) activity. We developed a sensitive and selective HPLC-MS/MS method for determining ethyl sulphate in urine. Ten volunteers received a low dose of ethanol (0.1 g/kg of body mass). In general, excretion of ethyl sulphate was maximal in the first or second hour after dosage. Within 8 h, 2.5-6.8 micromol of ethyl sulphate was excreted. A 5-fold increase in the dose of ethanol led to an increase in the amount of ethyl sulphate excreted within 8 h (28-95 micromol) and the presence of this metabolite in urine for at least 24 h. Since ethyl sulphate was still being excreted for a substantial period after the elimination of ethanol, it might be used as a medium-time biomarker for preceding ethanol consumption. We have expressed previously all human SULT forms identified in Salmonella typhimurium. Ethanol sulphonation was studied in cytosolic preparations of these strains. The highest activities were observed with SULT1A2, 1B1 and 1C2, followed by 1A3. Activities were markedly lower with SULT1E1, 1A1 and 2A1, and were negligible with SULT1C1, 2B1a, 2B1b and 4A1. If the expression levels in tissues are additionally taken into account, SULT1A3 might be the predominant form for the sulphonation of ethanol in vivo, although a robust estimate requires further studies. With this limitation, urinary ethyl sulphate excretion appears very promising as a biomarker for SULT activity in vivo. PMID- 15281912 TI - Argpyrimidine, a methylglyoxal-derived advanced glycation end-product in familial amyloidotic polyneuropathy. AB - FAP (familial amyloidotic polyneuropathy) is a systemic amyloid disease characterized by the formation of extracellular deposits of transthyretin. More than 80 single point mutations are associated with amyloidogenic behaviour and the onset of this fatal disease. It is believed that mutant forms of transthyretin lead to a decreased stability of the tetramer, which dissociates into monomers that are prone to unfolding and aggregation, later forming beta fibrils in amyloid deposits. This theory does not explain the formation of beta fibrils nor why they are toxic to nearby cells. Age at disease onset may vary by decades for patients with the same mutation. Moreover, non-mutated transthyretin also forms the same deposits in SSA (senile systemic amyloidosis), suggesting that mutations may only accelerate this process, but are not the determinant factor in amyloid fibril formation and cell toxicity. We propose that glycation is involved in amyloidogenesis, since amyloid fibrils present several properties common to glycated proteins. It was shown recently that glycation causes the structural transition from the folded soluble form to beta-fibrils in serum albumin. We identified for the first time a methylglyoxal-derived advanced glycation end-product, argpyrimidine [N(delta)-(5-hydroxy-4,6-dimethylpyrimidin-2 yl)-L-ornithine] in amyloid fibrils from FAP patients. Unequivocal argpyrimidine identification was achieved chromatographically by amino acid analysis using dabsyl (4-dimethylaminoazobenzene-4'-sulphonyl) chloride. Argpyrimidine was found at a concentration of 162.40+/-9.05 pmol/mg of protein in FAP patients, and it was not detected in control subjects. The presence of argpyrimidine in amyloid deposits from FAP patients supports the view that protein glycation is an important factor in amyloid diseases. PMID- 15281914 TI - Evaluation of the diffusion coefficient for controlled release of oxytetracycline from alginate/chitosan/poly(ethylene glycol) microbeads in simulated gastrointestinal environments. AB - Diffusion studies of OTC (oxytetracycline) entrapped in microbeads of calcium alginate, calcium alginate coacervated with chitosan (of high, medium and low viscosity) and calcium alginate coacervated with chitosan of low viscosity, covered with PEG [poly(ethylene glycol) of molecular mass 2, 4.6 and 10 kDa, were carried out at 37+/-0.5 degrees C, in pH 7.4 and pH 1.2 buffer solutions - conditions similar to those found in the gastrointestinal system. The diffusion coefficient, or diffusivity (D), of OTC was calculated by equations provided by Crank [(1975) Mathematics in Diffusion, p. 85, Clarendon Press, Oxford] for diffusion, which follows Fick's [(1855) Ann. Physik (Leipzig) 170, 59] second law, considering the diffusion from the inner parts to the surface of the microbeads. The least-squares and the Newton-Raphson [Carnahan, Luther and Wilkes (1969) Applied Numerical Methods, p. 319, John Wiley & Sons, New York] methods were used to obtain the diffusion coefficients. The microbead swelling at pH 7.4 and OTC diffusion is classically Fickian, suggesting that the OTC transport, in this case, is controlled by the exchange rates of free water and relaxation of calcium alginate chains. In case of acid media, it was observed that the phenomenon did not follow Fick's law, owing, probably, to the high solubility of the OTC in this environment. It was possible to modulate the release rate of OTC in several types of microbeads. The presence of cracks formed during the process of drying the microbeads was observed by scanning electron microscopy. PMID- 15281913 TI - Characterization of a novel low-molecular-mass dual-specificity phosphatase-3 (LDP-3) that enhances activation of JNK and p38. AB - We have isolated a mouse cDNA for a novel dual-specificity phosphatase designated LDP-3 (low-molecular-mass dual-specificity phosphatase 3). The 450 bp open reading frame encodes a protein of 150 amino acids with a predicted molecular mass of 16 kDa. Northern blot and reverse transcription-PCR analyses show that LDP-3 transcripts are expressed in almost all mouse tissues examined. In vitro analyses using several substrates and inhibitors indicate that LDP-3 possesses intrinsic dual-specificity phosphatase activity. When expressed in mammalian cells, LDP-3 protein is localized mainly to the apical submembrane area. Forced expression of LDP-3 does not alter activation of ERK (extracellular-signal regulated kinase), but rather enhances activation of JNK (c-Jun N-terminal kinase) and p38 and their respective upstream kinases MKK4 (mitogen-activated protein kinase kinase 4) and MKK6 in cells treated with 0.4 M sorbitol. By screening with a variety of stimuli, we found that LDP-3 specifically enhances the osmotic stress-induced activation of JNK and p38. PMID- 15281915 TI - Acculturation strategies, coping process and acculturative stress. AB - Using structural equation modeling, this study examines the influences of motivational factors (Need for Cognitive Closure--NCC--and Decisiveness), coping strategies and acculturation strategies on levels of acculturative stress. Two groups of immigrants in Rome (Croatians n= 156 and Poles n= 179) completed a questionnaire that included scales for the various factors. Although our initial hypothesized model was not confirmed, a modified model showed that the motivational factors of NCC and Decisiveness indirectly influence acculturative stress. The modified model with good fit indices indicated that the relationship between NCC and Decisiveness are mediated by coping strategies and acculturation strategies. Specifically, NCC is associated positively with avoidance coping, which in turn is negatively associated with the host group relationships and positively with the original culture maintenance. The last two dimensions predicted lower levels of acculturative stress. Decisiveness was positively associated with the problem-oriented coping and, negatively, with emotional and avoidance coping. PMID- 15281916 TI - A Swedish translation and validation of the Disgust Scale: a measure of disgust sensitivity. AB - The psychometric properties of a Swedish version of Haidt, McCauley and Rozin's (1994) Disgust Scale were studied. Confirmatory factor analysis of the original model with eight factors (food, animals, body products, sex, body envelope violations, death, hygiene, and magic) provided satisfactory fit to the data (N= 280), significantly better than to the alternative one-factor and five-factor models. As in the US version women scored significantly higher than men. Positive correlations with measures of food neophobia (r= 0.30, p < 0.0001) and nausea frequency (rs= 0.28, p < 0.001) indicate convergent validity. In a separate study (N= 30) a behavioral measure of the willingness to touch, hold, and taste disgusting food objects correlated negatively with the Disgust Scale (r=-0.46, p < 0.01), indicating criterion-related validity. PMID- 15281917 TI - Behavioral and social development of children born extremely premature: 5-year follow-up. AB - A cohort of extremely prematurely born children and matched term controls was assessed at 5 years of age. The parents completed a questionnaire on their behavioral and social development. The purpose was to illuminate whether the children's general intellectual ability and parental sensitivity were associated with behavioral and social development. The index children exhibited more hyperactive behavior and had poorer social skills than the controls. Lower Full Scale IQ (FSIQ) was associated with outward reacting and hyperactive behavior and poorer social skills. Sensitive parenting was associated with less outward reacting and less hyperactive behavior. When controlling for differences in FSIQ and parental sensitivity, the index children persisted to have an increased risk of exhibiting hyperactive behavior but not poorer social skills. The index children with normal intellectual development, however, did not exhibit more behavioral problems or poorer social skills than the control children did. PMID- 15281918 TI - Thinking styles in relation to personality traits: an investigation of the Thinking Styles Inventory and NEO-PI-R. AB - This study is an investigation of the Sternberg-Wagner Thinking Style Inventory (TSI), with regard to cross-cultural replication and relation to the five-factor personality model (FFM). TSI and NEO-PI-R were administered to 107 participants from USA and 114 participants from Norway. Inter-correlations between NEO-PI-R dimensions and TSI-scales and factors were not very strong, few exceeding 0.40, and the correlations were in predicted directions. Joint factor analyses of TSI and NEO-PI-R showed that TSI covers variance that NEO-PI-R does not explain. Thus, it is argued that the thinking styles give an independent contribution beyond FFM dimensions. However, TSI did not relate to FFM in the same manner in the two samples. Finally, the TSI-scales and factors were replicable across samples by Procrustes rotation. The question whether thinking style may be regarded as a valid and reliable construct is discussed. PMID- 15281919 TI - The role of teachers' perceived control and children's characteristics in interactions between 6-year-olds and their teachers. AB - The study aimed at an understanding of child-teacher interactions in school preparatory classrooms. Relations between observed interactions and sex of the child, teachers' ratings of their perceived control, and of children's undercontrolled and overcontrolled problem behaviors, social competence and work efficiency were studied. Thirty-six teachers and 92 six-year-olds, 39 girls and 53 boys, from 19 classrooms were directly observed on 2-5 occasions during a total mean of 60 minutes per child. The results showed that interactions involving teacher support behaviors were the most common, but comparatively less well explained by the predictors. Associations were found between perceived control and two types of teacher command interactions. For teacher commands initiated by child externalizing behavior, the relation with perceived control was shown to hold for boys only. Male sex and rated undercontrolled problems were predictive of more interactions initiated with externalizing behaviors and also of more restrictive teacher responses following child positive behaviors. Overcontrolled children, who had teachers high in perceived control, were more often met with support behavior when they were off-task. It was concluded that teacher perceptions of control and of child behaviors as well as sex of the child contribute to ongoing processes in preschool classrooms and that the chosen methodology could be used to further the search for factors affecting interactions in preschool settings. PMID- 15281920 TI - The effect of low social support on short-term prognosis in patients following a first myocardial infarction. AB - The objectives of the current study were (1) to assess adjustment in patients following a first myocardial infarction (MI) at 9 months compared with 4-6 weeks post-MI, (2) to examine the availability of and satisfaction with social support over time, and (3) to determine separate baseline psychosocial predictors of recurrent cardiac events. A questionnaire assessing post-traumatic stress disorder, anxiety, depression, health complaints, and social support, was distributed to consecutive patients 4-6 weeks and 9 months post-MI. Prior to assessment at follow-up, 8 (7%) of 112 patients had dropped out, and two had died due to cardiac causes. Objective clinical measures were obtained from medical records. There was an improvement in somatic and cognitive symptoms at follow-up, but no change in symptoms of arousal, depression, and anxiety. Half of the patients were afraid of a recurrent MI. There was a significant decrease in social support between baseline and follow-up, and lower social support at baseline was associated with a 10% increased risk of recurrent cardiac events at follow-up (OR: 0.90; 95% CI: 0.84 to 0.97) adjusting for all other variables. Some patients still experienced difficulties with psychosocial adjustment 9 months post-MI despite a reduction in somatic and cognitive symptoms. Social support decreased over time, which may have serious prognostic implications; lower social support at baseline was an independent predictor of recurrent events at 9 months. An important step for future research will be to investigate how social support can be enhanced in patients at risk. PMID- 15281921 TI - The reliability and validity of the Swedish version of the Body Shape Questionnaire. AB - Psychometric properties of the Swedish version of the Body Shape Questionnaire (BSQ) were examined using data from three samples: (1) a sample from the general population (n= 1157), (2) a student sample (n= 124) and (3) a clinical sample (n= 90). Analyses showed that a single factor solution might be a reasonable solution as 32 of the 34 items loaded heavily on the first factor. The derived short 14 item version of the BSQ also showed a coherent structure with all the items loading on one single factor. The BSQ showed high test-retest reliability, very high internal consistency, ranging from 0.94 to 0.97, and high split-half reliability (above 0.93). Furthermore, it showed high validity by correlating highly with the body dissatisfaction subscale of the Eating Disorders Inventory (r= 0.72 and higher), and high discriminant validity. Thus, the Swedish version of the BSQ showed good concurrent and discriminant validity as well as high reliability. PMID- 15281922 TI - Forensic psychology in Iceland: a survey of members of the Icelandic Psychological Society. AB - Forensic psychology is a rapidly growing specialism within psychology. A survey was carried out among the 152 members of the Icelandic Psychological Society (Salfradingafelagi Islands) about their involvement and role in court work. Out of 101 psychologists who responded to the survey, 39 reported having been involved in court work, of whom 34 had testified in court. The great majority of court work of the psychologists involved assessments relating to child-care and custody proceedings, but there was evidence that Icelandic psychologists are increasingly becoming involved in criminal cases concerning criminal responsibility and the reliability of testimony. They are working much more independently of medical colleagues than before. PMID- 15281923 TI - Appraised psychological workload, musculoskeletal symptoms, and the mediating effect of fatigue: a structural equation modeling approach. AB - The aim of the present study was to test two structural models of the relationship between appraised psychological workload and musculoskeletal symptoms from the neck, shoulder, and upper and lower back with different aspects of perceived fatigue as mediating variables. In this cross-sectional study a questionnaire survey was conducted among employees at three Swedish assembly plants (n= 305). The proposed models were tested for one general fatigue dimension--lack of energy--and four specific fatigue dimensions--physical discomfort, physical exertion, lack of motivation, and sleepiness--using structural equation modeling. The results indicate that the role of perceived fatigue in the relationship between appraised workload and musculoskeletal symptoms is different for different aspects of fatigue. The general fatigue dimension, lack of energy, does not mediate the relationship. As regards the specific fatigue dimensions, the relationship is partially mediated by physical discomfort and lack of motivation but not by physical exertion or sleepiness. Appraised psychological workload has a unique effect on musculoskeletal symptoms not mediated by fatigue. PMID- 15281924 TI - Graphic jokes and children's mind: an unusual way to approach children's representational activity. AB - The purpose of this study was to examine the development of representational activity through the comprehension of graphic jokes in 2, 3 and 4-year-old children. In experiment 1 we worked with three kinds of jokes, specifically mentalistic jokes, jokes based on substitution and complex jokes. We found differentiated performances on each kind of joke, as had been expected based on the semiological analysis of the jokes prior to the experiment. The earliest comprehension, at 3 years old, occurred with mentalistic jokes where more than 70% of the total sample was successful. The substitution jokes reached 47% in the three-year-old subjects, and the latest kind of joke to be understood by this group were the complex jokes, with only 31% comprehension. In experiment 2 we wanted to specify the cognitive functioning that was taking place in the comprehension of mentalistic jokes. We found similar successful performances in two mentalistic jokes with both the 3 and 4 year-olds in the study. Children's performances were analyzed from the point of view of processes of redescription which were involved in the understanding of the jokes. We conclude that humor tasks are appropriate instruments to examine development of children's representational abilities. PMID- 15281925 TI - Genetic heterogeneity and functional properties of intestinal bifidobacteria. AB - AIMS: The aim of the present study was to compare several molecular methods for the identification and genotyping of bifidobacteria, and further to investigate genetic heterogeneity and functional properties of bifidobacterial isolates from intestinal samples of Finnish adult subjects. METHODS AND RESULTS: A total of 153 intestinal bifidobacterial isolates were included in initial screening and 34 isolates were further characterized. Identification results obtained with PCR ELISA and ribotyping were well in accordance with each other, while randomly amplified polymorphic DNA (RAPD) gave tentative identification only to Bifidobacterium bifidum and to 65% of the B. longum isolates. The most commonly detected species were B. longum biotype longum followed by B. adolescentis and B. bifidum. In addition, B. animalis (lactis), B. angulatum and B. pseudocatenulatum were found. Ribotyping and pulsed-field gel electrophoresis (PFGE) proved to be discriminatory methods for bifidobacteria, but also RAPD revealed intraspecies heterogeneity. Besides two B. animalis (lactis) isolates with very close similarity to a commercially available probiotic strain, none of the intestinal isolates showed optimal survival in all tolerance (acid, bile and oxygen) or growth performance tests. CONCLUSIONS: Several species/strains of bifidobacteria simultaneously colonize the gastrointestinal tract of healthy Finnish adults and intestinal Bifidobacterium isolates were genetically heterogeneous. Functional properties of bifidobacteria were strain-dependent. SIGNIFICANCE AND IMPACT OF THE STUDY: Applicability of ribotyping with the automated RiboPrinter System for identification and genotyping of bifidobacteria was shown in the present study. PMID- 15281926 TI - A cost-effective cane molasses medium for enhanced cell-bound phytase production by Pichia anomala. AB - AIM: Formulation of an inexpensive cane molasses medium for improved cell-bound phytase production by Pichia anomala. METHODS AND RESULTS: Cell-bound phytase production by Pichia anomala was compared in synthetic glucose-beef extract and cane molasses media. The yeast was cultivated in 250 ml flasks containing 50 ml of the medium, inoculated with a 12 h-old inoculum (3 x 10(6) CFU ml(-1)) and incubated at 25 degrees C for 24 h at 250 rev min(-1). Different cultural parameters were optimized in cane molasses medium in batch fermentation. The cell bound phytase content increased significantly in cane molasses medium (176 U g( 1) dry biomass) when compared with the synthetic medium (100 U g(-1) dry biomass). In fed-batch fermentation, a marked increase in biomass (20 g l(-1)) and the phytase yield (3000 U l(-1)) were recorded in cane molasses medium. The cost of production in cane molasses medium was pound 0.006 per 1000 U, which is much lower when compared with that in synthetic medium (pound 0.25 per 1000 U). CONCLUSIONS: An overall 86.6% enhancement in phytase yield was attained in optimized cane molasses medium using fed-batch fermentation when compared with that in synthetic medium. Furthermore, the production in cane molasses medium is cost-effective. SIGNIFICANCE AND IMPACT OF THE STUDY: Phytase yield was improved in cane molasses when compared with the synthetic medium, and the cost of production was also significantly reduced. This enzyme can find application in the animal feed industry for improving the nutritional status of feed and combating environmental pollution. PMID- 15281927 TI - Paenibacillus isolates possess diverse dextran-degrading enzymes. AB - AIMS: To isolate and identify dextran-degrading organisms from sugar mill and compost samples, and to examine the diversity of the dextranolytic enzymes produced. METHODS AND RESULTS: Fifteen dextranolytic prokaryotes were purified at various temperatures from sugar-mill or compost samples, using indicator plates containing blue dextran. A 16S rRNA gene sequence analysis showed that 12 isolates purified at 40, 50 or 70 degrees C were closely aligned to Paenibacillus spp. The three isolates purified at 60 degrees C had identical 16S rDNA sequences, with highest affinity to Bacillus spp. Liquid culture of the 11 isolates purified at 40 or 50 degrees C produced dextranolytic activity in the spent media with maximal activity at 40 or 45 degrees C under the assay conditions used. Hydrolysis of blue dextran in activity gels showed that the 12 Paenibacillus isolates produced from one to five dextranolytic proteins, ranging from 70 to 120 kDa. Based on 16S rDNA sequence, growth habit in liquid culture and dextranolytic enzyme pattern, the 12 Paenibacillus-like isolates could be differentiated into six distinct groups, one of which was capable of growth at 70 degrees C. CONCLUSIONS: The Bacillales, especially the Paenibacillus, are a valuable environmental repository for dextranolytic enzymes of diverse size and potentially diverse activity. SIGNIFICANCE AND IMPACT OF THE STUDY: Dextranolytic enzymes produced by Paenibacillus spp. are an exploitable resource for those interested in modifying the structure of dextrans. PMID- 15281928 TI - Modelling the growth of Trichoderma virens with limited sampling of digital images. AB - AIMS: Using limited digital image sampling, a model of fungal growth in soil that considers both hyphal production and lysis was constructed for two strains of Trichoderma virens over a range of four temperatures. MATERIALS AND METHODS: A growth model was developed by fitting the radial cross sectional data with a modified form of the Ratkowsky equation to determine maximum growth rate and a modified Arrhenius equation to determine maximal rate of decrease in area covered by mycelia. The parameters obtained from a combined equation were then verified by using the data obtained from the whole colony to determine the appropriateness of the model. CONCLUSIONS: Using a limited data set and a combination of the Ratkowsky and Arrhenius equations, the mycelial coverage of the T. virens colony was determined, relating microscopic hyphal growth to macroscopic colony growth. This model was sufficiently robust to predict growth across four temperatures for a genetically modified and wild-type strain of T. virens. SIGNIFICANCE AND IMPACT OF STUDY: By using simple assumptions for the increase and eventual decline in fungal growth on a resource-limited medium, this model constructs an initial framework onto which additional parameters such as nutrient consumption could be incorporated for prediction of fungal growth. PMID- 15281929 TI - Cloning and partial characterization of zwittermicin A resistance gene cluster from Bacillus thuringiensis subsp. kurstaki strain HD1. AB - AIM: The study seeks to shed light on the aminopolyol, broad-spectrum antibiotic zwittermicin A gene cluster of Bacillus thuringiensis subsp. kurstaki HD1 and to identify any new uncharacterized genes with an eventual goal to establish a better understanding of the resistance gene cluster. METHODS AND RESULTS: We screened 51 serovars of B. thuringiensis by PCR and identified 12 zmaR-positive strains. The zmaR-positive B. thuringiensis subsp. kurstaki HD1 strain displayed inhibition zones against indicator fungal strain Phytophthora meadii and bacterial strain Erwinia herbicola as well as against Rhizopus sp., Xanthomonas campestris and B. thuringiensis subsp. finitimus. The zmaR gene cluster of strain HD1 was partially cloned using a lambda library and was extensively characterized based on the information available from a study performed on a similar group of genes in Bacillus cereus. CONCLUSIONS: Three of the five genes in the zwittermicin gene cluster, including the zmaR gene, had counterparts in B. cereus, and the other two were new members of the B. thuringiensis zmaR gene cluster. SIGNIFICANCE AND IMPACT OF THE STUDY: The two new genes were extensively analysed and the data is presented. Understanding antifungal activity of B. thuringiensis may help us to design suitable Cry toxin delivery agents with antifungal activity as well as enhanced insecticidal activity. PMID- 15281930 TI - A putative new endophytic nitrogen-fixing bacterium Pantoea sp. from sugarcane. AB - AIMS: To isolate and identify endophytic nitrogen-fixing bacteria in sugarcane growing in Cuba without chemical fertilizers. METHODS AND RESULTS: Two N2-fixing isolates, 9C and T2, were obtained from surface-sterilized stems and roots, respectively, of sugarcane variety ML3-18. Both isolates showed acetylene reduction and H2 production in nitrogen-free media. Nitrogenase activity measured by H2 production was about 15 times higher for isolate 9C than for T2 or for Gluconoacetobacter diazotrophicus (PAL-5 standard strain, ATCC 49037). The nifH gene segment was amplified from both isolates using specific primers. Classification of both T2 and 9C was made on the basis of morphological, biochemical, PCR tests and 16S rDNA sequence analysis. CONCLUSIONS: Isolate 9C was identified as a Pantoea species from its 16S rDNA, but showed considerable differences in physiological properties from previously reported species of this genus. For example, 9C can be cultured over a wide range of temperature, pH and salt concentration, and showed high H2 production (up to 67.7 nmol H2 h(-1) 10(10) cell(-1)). Isolate T2 was a strain of Gluconacetobacter diazotrophicus. SIGNIFICANCE AND IMPACT OF THE STUDY: A new N2-fixing endophyte, i.e. Pantoea, able to produce H2 and to grow in a wide range of conditions, was isolated from sugarcane stem tissue and characterized. The strain with these attributes may well be valuable for agriculture. PMID- 15281931 TI - Exopolysaccharides produced by mixed culture of yeast Rhodotorula rubra GED10 and yogurt bacteria (Streptococcus thermophilus 13a + Lactobacillus bulgaricus 2-11). AB - AIMS: The studies of the production of exopolysaccharides by lactose-negative yeast and a yogurt starter co-cultivated in a natural substrate containing lactose may be considered of interest because they reveal the possibilities for high-efficiency synthesis of biopolymers by mixed cultivation. METHODS AND RESULTS: The mixed culture Rhodotorula rubra GED10 + (Streptococcus thermophilus 13a + Lactobacillus bulgaricus 2-11) was cultivated in cheese whey ultrafiltrate (WU) (44.0 g lactose l(-1)) at initial pH 6.0, 28 degrees C, under intensive aeration (air-flow rate 1.0 l l(-1) min(-1), agitation 220 rev min(-1)) in a MBR AG fermentor. The mixed culture manifested the highest activity for synthesis of exopolysaccharides (19.3 g l(-1)) and cell mass (21.0 g l(-1)) at the 84th hour. The yogurt starter synthesized neutral exopolysaccharides, while the mixed culture yeast + yogurt starter produced acidic exopolysaccharides containing uronic acid (6%). The neutral sugar composition was identified as mannose, glucose, galactose, xylose and arabinose. Mannose dominated in the polymer composition (83%) that was produced only by the yeast (97%). CONCLUSIONS: Lactose in the WU can be effectively utilized by a co-culture of lactose-negative yeast yogurt starter for synthesis of exopolysaccharides. SIGNIFICANCE AND IMPACT OF THE STUDY: The present findings propose an alternative use of WU as a cost effective carbohydrate substrate, and suggest that the lactose-negative yeast Rhodotorula rubra can have industrial application as producers of exopolysaccharides. PMID- 15281932 TI - Effects of several inhibitors on pure cultures of ruminal methanogens. AB - AIMS: To examine the effects of five inhibitors of methanogenesis, 2 bromoethanesulphonate (BES), 3-bromopropanesulphonate (BPS), lumazine, propynoic acid and ethyl 2-butynoate, on CH4 production of the ruminal methanogens Methanobrevibacter ruminantium, Methanosarcina mazei and Methanomicrobium mobile. METHODS AND RESULTS: Methanogens were grown in MS medium including 25% (v/v) clarified ruminal fluid. Methane production was measured after 4 and 6 days of incubation. Methanobrevibacter ruminantium was the most sensitive species to BES, propynoic acid and ethyl 2-butynoate. Methanosarcina mazei was the least sensitive species to those chemical additives, and Mm. mobile was intermediate. BPS failed to inhibit any of the methanogens. All three species were almost completely inhibited by 50- and 100%-lumazine saturated media, but the inhibition was somewhat lower with a 25%-lumazine saturated media. CONCLUSIONS: There were important differences among species of methanogens regarding their sensitivity to the different inhibitors. In general, Ms. mazei was the most resistant to inhibitors, Mb. ruminantium the least resistant, and Mm. mobile was intermediate. SIGNIFICANCE AND IMPACT OF THE STUDY: Differences among methanogens regarding their resistance to chemical inhibitors should be considered when designing strategies of inhibition of ruminal methanogenesis, as selection of resistant species may result. PMID- 15281933 TI - Increased stress tolerance of Bifidobacterium longum and Lactococcus lactis produced during continuous mixed-strain immobilized-cell fermentation. AB - AIMS: The effect of immobilization and long-term continuous culture was studied on probiotic and technological characteristics of lactic acid and probiotic bacteria. METHODS AND RESULTS: A continuous culture in a two-stage system was carried out for 17 days at different temperatures ranging from 32 to 37 degrees C, with a first reactor containing Bifidobacterium longum ATCC 15707 and Lactococcus lactis subsp. lactis biovar. diacetylactis MD immobilized separately in gel beads, and a second reactor operated with free cells released from the first reactor. The tolerance of free cells from both strains produced in the effluent medium of both reactors to hydrogen peroxide, simulated gastric and intestinal juices, antibiotics and nisin, and freeze-drying markedly increased with culture time and was generally higher after 6 days than that of stationary phase cells produced during free-cell batch fermentations. The reversibility of the acquired tolerance of B. longum, but not L. diacetylactis, to antibiotics was shown during successive free-cell batch cultures. CONCLUSIONS: Free cells produced from continuous immobilized-cell culture exhibited altered physiology and increased tolerance to various chemical and physico-chemical stresses. SIGNIFICANCE AND IMPACT OF THE STUDY: Continuous culture with immobilized cells could be used to produce probiotic and lactic acid bacteria with enhanced technological and probiotic characteristics. PMID- 15281934 TI - Growth and fermentation patterns of Saccharomyces cerevisiae under different ammonium concentrations and its implications in winemaking industry. AB - AIMS: To study the effects of assimilable nitrogen concentration on growth profile and on fermentation kinetics of Saccharomyces cerevisiae. METHODS AND RESULTS: Saccharomyces cerevisiae was grown in batch in a defined medium with glucose (200 g l(-1)) as the only carbon and energy source, and nitrogen supplied as ammonium sulphate or phosphate forms under different concentrations. The initial nitrogen concentration in the media had no effect on specific growth rates of the yeast strain PYCC 4072. However, fermentation rate and the time required for completion of the alcoholic fermentation were strongly dependent on nitrogen availability. At the stationary phase, the addition of ammonium was effective in increasing cell population, fermentation rate and ethanol. CONCLUSIONS: The yeast strain required a minimum of 267 mg N l(-1) to attain complete dryness of media, within the time considered for the experiments. Lower levels were enough to support growth, although leading to sluggish or stuck fermentation. SIGNIFICANCE AND IMPACT OF THE STUDY: The findings reported here contribute to elucidate the role of nitrogen on growth and fermentation performance of wine yeast. This information might be useful to the wine industry where excessive addition of nitrogen to prevent sluggish or stuck fermentation might have a negative impact on wine stability and quality. PMID- 15281935 TI - Assessment of the rind microbial diversity in a farmhouse-produced vs a pasteurized industrially produced soft red-smear cheese using both cultivation and rDNA-based methods. AB - AIMS: The diversity of the surface flora of two French red-smear soft cheeses was examined by cultivation-dependent and cultivation-independent methods to assess their composition and to evaluate the accuracy of both approaches. METHODS AND RESULTS: Culture-independent methods used involved 16S ribosomal DNA gene cloning and sequencing and single-strand conformation polymorphism analysis (SSCP). The culture-dependent method used involved direct culture and macroscopic observation, polymerase chain reaction of the 16S rRNA gene from DNA extracted from single colonies followed by complete sequencing of the gene. Only few species were recovered by both approaches either in the pasteurized and the farmer cheese. A large diversity of isolates or 16S rDNA sequences related to marine bacteria was identified at the surface of both cheeses. CONCLUSIONS: The results indicated that all three techniques were informative and complementary to allow a more accurate representativeness of the cheese surface biodiversity. SIGNIFICANCE AND IMPACT OF THE STUDY: Cultivation and molecular methods have to be combined in order to obtain an extended view of the bacterial populations of complex ecosystems. PMID- 15281936 TI - Maintenance of pathogenicity during entry into and resuscitation from viable but nonculturable state in Aeromonas hydrophila exposed to natural seawater at low temperature. AB - AIMS: To investigate the fate of Aeromonas hydrophila pathogenicity when cells switch, in nutrient-poor filtered sterilized seawater, between the culturable and nonculturable state. METHODS AND RESULTS: Aeromonas hydrophila ATCC 7966, rendered non culturable within 50-55 days of exposure to marine stress conditions, was tested for its ability to maintain haemolysin and to adhere to McCoy cells. Results showed that pathogenicity was lost concomitantly with culturability, whereas cell viability remained undamaged, as determined by the Kogure cell elongation test. However, this loss is only temporary because, following temperature shift from 5 to 23 degrees C, multiple biological activities of recovered Aer. hydrophila cells, which include their ability to lyse human erythrocytes and to attach and destroy McCoy cells were regained. During the temperature-induced resuscitation, constant total cell counts were observed. Moreover, no significant improvement in recovery yield was obtained on brain-heart infusion (BHI) agar plates amended with catalase. We suggest that in addition to the growth of the few undetected culturable cells, there is repair and growth of some mildly injured viable but nonculturable cells. CONCLUSIONS: The possibility that nonculturable cells of normally culturable Aer. hydrophila in natural marine environment may constitute a source of infectious diseases posing a public health problem was demonstrated. SIGNIFICANCE AND IMPACT OF THE STUDY: These experiments may mimic what happens when Aer. hydrophila cells are released in natural seawater with careful attention to the conditions in which surrounding waters gradually become warmer in late summer/early autumn. PMID- 15281937 TI - Population structure of Salmonella investigated by amplified fragment length polymorphism. AB - AIMS: This study was undertaken to investigate the usefulness of amplified fragment length polymorphism (AFLP) in determining the population structure of Salmonella. METHODS AND RESULTS: A total of 89 strains were subjected to AFLP analysis using the enzymes BglII and BspDI, a combination that is novel in Salmonella. Both species S. bongori and S. enterica and all subsp. of S. enterica were represented with emphasis on S. enterica subsp. enterica using a local strain collection and strains from the Salmonella Reference Collection B (SARB). The amplified fragments were used in a band-based cluster analysis. The tree resulting from the subgroup analysis clearly separated all subgroups with high bootstrap values with the species S. bongori being the most distantly related of the subgroups. The tree resulting from the analysis of the SARB collection showed that some serotypes are very clonal whereas others are highly divergent. CONCLUSIONS: AFLP clearly clustered strains representing the subgroups of Salmonella together with high bootstrap values and the serotypes of subspecies enterica were divided into polyphyletic or monophyletic types corresponding well with multilocus enzyme electrophoresis (MLEE) and sequence-based studies of the population structure in Salmonella. SIGNIFICANCE AND IMPACT OF THE STUDY: AFLP with the enzyme combination BglII and BspDI allows discrimination of individual strains and provides evidence for the usefulness of AFLP in studies of population structure in Salmonella. PMID- 15281938 TI - Identification of P18, a surface protein produced by the fish pathogen Flavobacterium psychrophilum. AB - AIMS: This study was focused on the identification of associated outer membrane proteins which may play a role in the specific interactions between Flavobacterium psychrophilum (the aetiological agent of cold-water disease and rainbow trout fry syndrome in salmonid fish worldwide) and the fish tissues. METHODS AND RESULTS: The surface protein interactions with the outer membrane being mainly ionic, different methods were used for the detachment of proteins from the cell surface of Fl. psychrophilum involving detergent-free buffers or solutions known to perturb the ionic interactions. Such treatments led to the isolation of a surface protein, named P18 in accordance with its relative molecular mass. The expression of P18 was not related to the growth conditions (liquid or solid medium, temperature and aeration) or the strains of Fl. psychrophilum tested here. CONCLUSIONS: Preliminary characterization indicated that P18 is a surface antigen which is not sugar-modified and might be a subunit of a surface layer (i.e. S-layer), one of the most common surface structures on bacteria. SIGNIFICANCE AND IMPACT OF THE STUDY: Data reported here should be used as the basis for further works involving the purification and characterization of P18 to identify the specific roles of such a surface protein, especially the interaction between this protein and the host surface. PMID- 15281939 TI - Development and application of a spiral plating method for the enumeration of Escherichia coli O157 in bovine faeces. AB - AIM: To develop and validate a direct plating method applicable to epidemiological studies for enumerating Escherichia coli O157 in cattle faeces. METHODS AND RESULTS: The spiral plate count method was used to enumerate E. coli O157 in faecal samples. The accuracy and variation of counts was then assessed using faecal samples inoculated with E. coli O157. There was good agreement between inoculated levels of E. coli O157 and those recovered from faeces, particularly when counts were > 10(2) CFU g(-1) of faeces. The method was applied to a small study assessing short-term survival of E. coli O157 in naturally infected cattle faeces. E. coli O157 was found to survive in faeces for over 10 days at concentrations above 10(3) CFU g(-1) of faeces. Populations of E. coli O157 were also found to increase 100-fold in the first few hours after defecation. CONCLUSIONS: The enumeration method is easy to implement and enables a quick throughput of large numbers of samples. The method is accurate and reliable and enables the inherent variation in count data to be explored but needs to be used in combination with a more sensitive method for samples containing < 10(2) CFU g(-1) of faeces. SIGNIFICANCE AND IMPACT OF THE STUDY: The method described is appropriate for enumeration of E. coli O157 in cattle faeces in large-scale epidemiological studies. PMID- 15281940 TI - Microbiological influences in 'blue water' copper corrosion. AB - AIMS: To investigate the influence of micro-organisms associated with copper corrosion on 'blue water' corrosion in drinking water. METHODS AND RESULTS: Laboratory rigs comprising of polycarbonate containers attached to annealed copper plumbing tubes were filled with Melbourne drinking water and sterilized by autoclaving. The copper tubes were inoculated with sterile or nonsterile extracts obtained from corroding copper and allowed to stand for 7 days. The extracts were drained and the tubes flushed and filled with sterile water from the rig. The water within the tubes was removed weekly for analysis and the tubes were refilled with freshly aerated water. The tube water sampled was analysed for pH, total copper and the presence of micro-organisms. Sterile rigs and rigs containing nonsterile water, both without tube inoculums, were used as controls. The results demonstrated that tubes inoculated with nonsterile corrosion extracts showed statistically higher copper release compared with the other rigs. Copper release as blue water was only observed after a lag period of 9 weeks. The internal surfaces of tubes releasing copper showed significant amounts of corrosion products and the presence of biofilm. Bacteria isolated from the corroding tubes included Acidovorax spp. and Sphingomonas sp. CONCLUSIONS: The results demonstrate a microbial role in blue water, as corrosion was induced in new copper tubes by exposure to nonsterile copper corrosion products. SIGNIFICANCE AND IMPACT OF THE STUDY: The potential for micro-organisms present in corrosion products to initiate blue water corrosion presents significant implications for the management of corrosion in distribution systems. PMID- 15281941 TI - Retention and removal of the fish pathogenic bacterium Yersinia ruckeri in biological sand filters. AB - AIMS: To investigate the retention and removal of the fish pathogenic bacterium Yersinia ruckeri in biological sand filters and effects on the microbial community composition. METHODS AND RESULTS: Sand filter columns were loaded (70 mm day(-1)) with fish farm wastewater and a suspension (10(8) CFU ml(-1)) of Y. ruckeri. Bacterial numbers and protozoan numbers were determined by plate counts and epifluorescence microscopy, respectively, and microbial biomass and community composition were assessed by phospholipid fatty acids (PLFA) analysis. Concentrations of Y. ruckeri in the filter effluent decreased from 10(8) to 10(3) 10(5) CFU ml(-1) during the experiment. Numbers of Y. ruckeri in the sand decreased from 10(6) CFU g(-1) dry weight (DW) sand to 10(4) CFU g(-1) DW sand. In contrast, microbial biomass determined with plate counts and total PLFA increased during the whole experiment. Principal component analysis (PCA) revealed a change in microbial community composition with time, with the most pronounced change in surface layers and towards the end of the experiment. Protozoan numbers increased from ca 0-600 cells g(-1) DW sand, indicating the establishment of a moderate population of bacterial grazers. CONCLUSIONS: The removal of Y. ruckeri improved during the experiment. Introduction of Y. ruckeri to the sand filter columns stimulated growth of other micro-organisms, which in turn caused a shift in the microbial community composition in the sand. SIGNIFICANCE AND IMPACT OF THE STUDY: This study increases the understanding of the dynamics of sand filters subjected to a high loading of a pathogenic bacterium and can therefore be used in future work were the overall aim is to provide a more reliable and efficient removal of pathogenic bacteria in biological sand filter systems. PMID- 15281942 TI - Predicting noncompliant levels of ochratoxin A in cereal grain from Penicillium verrucosum counts. AB - AIMS: To model the probability of exceeding the European legislative limit of 5 microg ochratoxin A (OTA) per kilogram grain in relation to Penicillium verrucosum levels and storage conditions, and to evaluate the possibilities of using P. verrucosum colony counts for predicting noncompliant OTA levels. METHODS AND RESULTS: Cereal samples were inoculated with P. verrucosum spores and stored for up to 9 months at temperatures and water activities ranging from 10-25 degrees C and aw 0.77-0.95. A logistic regression analysis showed that the probability of exceeding 5 microg OTA kg(-1) grain was related to colony counts of P. verrucosum and water activity. The sensitivity and specificity of various P. verrucosum count thresholds for predicting noncompliant OTA levels were estimated, using data from the storage trial and natural cereal samples. CONCLUSION: The risk of exceeding 5 microg OTA kg(-1) grain increased with increasing levels of P. verrucosum, and with increasing water activities. A threshold of 1000 CFU P. verrucosum per gram grain is suggested to predict whether or not the legislative limit is exceeded. SIGNIFICANCE AND IMPACT OF THE STUDY: This study has provided a tool to evaluate the levels of P. verrucosum in grain in relation to OTA levels. Hence, mycological analyses can be used to identify cereal samples with high risk of containing OTA levels above the legislative limit. PMID- 15281943 TI - Effect of sunlight on the survival of Salmonella on surfaces. AB - AIMS: To investigate the effect of simulated full-spectrum tropical sunlight on the survival of Salmonella in droplets on surfaces. MATERIALS AND RESULTS: The survival on surfaces of three Zambian strains of Salmonella enterica serovars Enteritidis and Heidelberg was compared with that of a strain of S. enterica serovar Enteritidis phage type (PT) 4 with known characteristics which had been isolated from poultry in the UK. Samples were taken from surfaces every hour for 3 h and after 24 h exposure in either dark or 12 h light/12 h dark cycle conditions. Differences were analysed for significance using a one-way analysis of variance (anova). Results show that there were a significantly higher number of cells surviving on surfaces after 24 h in the dark when compared with populations exposed to a 12 h light/12 h dark cycle. Significantly more cells also survived exposure to sunlight under dirty than clean conditions. CONCLUSIONS: Exposure to sunlight results in a significant decrease in numbers of Salmonella on surfaces. SIGNIFICANCE AND IMPACT OF THE STUDY: Under field conditions exposure of contaminated surfaces to sunlight could be used in place of chemical methods of control as a cheaper way to reduce Salmonella contamination of surfaces. PMID- 15281944 TI - Antilisterial activity of lactic acid bacteria isolated from rigouta, a traditional Tunisian cheese. AB - AIMS: Screening for lactic acid bacteria (LAB) producing bacteriocins and other antimicrobial compounds is of a great significance for the dairy industry to improve food safety. METHODS AND RESULTS: Six-hundred strains of LAB isolated from 'rigouta', a Tunisian fermented cheese, were tested for antilisterial activity. Eight bacteriocinogenic strains were selected and analysed. Seven of these strains were identified as Lactococcus lactis and produced nisin Z as demonstrated by mass spectrometry analysis of the purified antibacterial compound. Polymerase chain reaction experiments using nisin gene-specific primers confirmed the presence of nisin operon. Plasmid profiles analysis suggests the presence of, at least, three different strains in this group. MMT05, the eighth strain of this antilisterial collection was identified, at molecular level, as Enterococcus faecalis. The purified bacteriocin produced by this strain showed a molecular mass of 10 201.33 +/- 0.85 Da. This new member of class III bacteriocins was termed enterocin MMT05. CONCLUSIONS: Seven lactococcal strains producing nisin Z were selected and could be useful as bio-preservative starter cultures. Additional experiments are needed to evaluate the promising strain MMT05 as bio-preservative as Enterococci could exert detrimental or beneficial role in foods. SIGNIFICANCE AND IMPACT OF THE STUDY: Only a few antibacterial strains isolated from traditional African dairy products were described. The new eight strains described herein contribute to the knowledge of this poorly studied environment and constitute promising strains for fermented food safety. PMID- 15281945 TI - Degradation of isooctane by Mycobacterium austroafricanum IFP 2173: growth and catabolic pathway. AB - AIMS: Isooctane (2,2,4-trimethylpentane), a major component of gasoline formulations, is recalcitrant to biodegradation probably because of the quaternary carbon group it contains. Information on the biodegradability of this hydrocarbon is essential to evaluate its fate in the environment. For these reasons, the degradation kinetics and the catabolic pathway of isooctane were investigated in Mycobacterium austroafricanum IFP 2173, the only strain characterized to use it as sole carbon and energy source. METHODS AND RESULTS: The selected strain exhibited a rather moderate maximum growth rate (micromax = 0.053 h(-1)) but degraded isooctane up to 99% with a mineralization yield of 45%, indicating attack of the quaternary carbon group. The GC/MS identification of metabolites, 2,4,4-trimethylpentanoic and dimethylpropanoic (pivalic) acids, which transiently accumulated in the cultures indicated that degradation started from the isopropyl extremity of the molecule and subsequently proceeded by catabolism of the tert-butyl moiety. The degradation of putative metabolic intermediates was investigated. The initial isooctane oxidation system was tentatively characterized. CONCLUSIONS: The isooctane-degrading strain harboured two candidate systems for initial alkane oxidation. Although a cytochrome P450 was induced by isooctane degradation, the functional oxidation system was probably a nonheme alkane monooxygenase as indicated by PCR amplification and RT PCR expression of an alkB gene. SIGNIFICANCE AND IMPACT OF THE STUDY: Isooctane is a recalcitrant branched alkane. A plausible pathway of its degradation by Myco. austroafricanum was put forward. PMID- 15281946 TI - Screening for novel laccase-producing microbes. AB - AIMS: To discover novel laccases potential for industrial applications. METHODS AND RESULTS: Fungi were cultivated on solid media containing indicator compounds that enabled the detection of laccases as specific colour reactions. The indicators used were Remazol Brilliant Blue R (RBBR), Poly R-478, guaiacol and tannic acid. The screening work resulted in isolation of 26 positive fungal strains. Liquid cultivations of positive strains confirmed that four efficient laccase producers were found in the screening. Biochemical characteristics of the four novel laccases were typical for fungal laccases in terms of molecular weight, pH optima and pI. The laccases showed good thermal stability at 60 degrees C. CONCLUSIONS: Plate-test screening based on polymeric dye compounds, guaiacol and tannic acid is an efficient way to discover novel laccase producers. The results indicated that screening for laccase activity can be performed with guaiacol and RBBR or Poly R-478. SIGNIFICANCE AND IMPACT OF THE STUDY: Laccases have many potential industrial applications including textile dye decolourization, delignification of pulp and effluent detoxification. It is essential to find novel, efficient enzymes to further develop these applications. This study showed that relatively simple plate test screening method can be used for discovery of novel laccases. PMID- 15281947 TI - The role of indigenous yeasts in traditional Irish cider fermentations. AB - AIMS: To study the role of the indigenous yeast flora in traditional Irish cider fermentations. METHODS AND RESULTS: Wallerstein laboratory nutrient agar supplemented with biotin, ferric ammonium citrate, calcium carbonate and ethanol was employed together with PCR-restriction fragment length polymorphism analysis of the region spanning the internal transcribed spacers (ITS1 and ITS2) and the 5.8S rRNA gene in the identification of indigenous yeasts at the species level, from traditional Irish cider fermentations. By combining the molecular approach and the presumptive media it was possible to distinguish between a large number of yeast species, and to track them within cider fermentations. The Irish cider fermentation process can be divided into three sequential phases based on the predominant yeast type present. Kloeckera/Hanseniaspora uvarum type yeasts predominate in the initial 'fruit yeast phase'. Thereafter Saccharomyces cerevisiae type yeast dominate in the 'fermentation phase', where the alcoholic fermentation takes place. Finally the 'maturation phase' which follows, is dominated by Dekkera and Brettanomyces type yeasts. H. uvarum type yeast were found to have originated from the fruit. Brettanomyces type yeast could be traced back to the press house, and also to the fruit. The press house was identified as having high levels of S. cerevisiae type yeast. A strong link was noted between the temperature profile of the cider fermentations, which ranged from 22 to 35 degrees C and the yeast strain population dynamics. CONCLUSIONS: Many different indigenous yeast species were identified. The mycology of Irish cider fermentations appears to be very similar to that which has previously been reported in the wine industry. SIGNIFICANCE AND IMPACT OF THE STUDY: This study has allowed us to gain a better understanding of the role of indigenous yeast species in 'Natural' Irish cider fermentations. PMID- 15281948 TI - Mineral and carbon usage of two synthetic pyrethroid degrading bacterial isolates. AB - AIMS: To investigate the biodegrading ability and cometabolism of synthetic pyrethroid (SP) utilizing bacteria in cultures with various minerals and carbon sources. METHODS AND RESULTS: Previously isolated SP-degrading Pseudomonas sp. and Serratia sp. were used in cultures containing either flumethrin SP or cypermethrin SP formulations. The culture media consisted of either (i) water only, (ii) water and sucrose, (iii) mineral broth or (iv) mineral broth and sucrose. The growth of both organisms was greatest in the mineral broth and sucrose medium, but the growth-limiting factor for Pseudomonas sp. strain Circle was the mineral content whereas for Serratia sp. strain White it was the carbon substrate. CONCLUSION: The greatest extent of degradation of both SP-based compounds occurred with Pseudomonas sp. strain Circle but was dependant on the medium. SIGNIFICANCE AND IMPACT OF THE STUDY: This investigation could lead to the development of a relatively inexpensive medium supplement to enhance the microbial biodegradation of undesirable compounds, either in situ or ex situ. In this particular case, for the biodegradation of SPs used in sheep dip. PMID- 15281950 TI - Catastrophic focal epilepsy. AB - Focal epilepsy can present with a rapidly progressing course of intractable epilepsy in children. We present a typical example of such a patient with focal seizures due to a frontal lobe cortical lesion of developmental origin. MRI and SPECT revealed abnormalities in the right frontal lobe. Surgical resection resulted in excellent outcome. PMID- 15281951 TI - Structural MR imaging. AB - Phased-array imaging at 1.5 and 3 T significantly improves image quality compared with that in routine 1.5-T head coil studies. These improvements significantly increase lesion detection in patients with focal epilepsy. Semiautomated image analysis techniques have the potential to improve lesion detection, assess lesion burden more accurately, characterize cortical abnormalities, and determine the location and extent of associated cortical and deep gray nuclei involvement. With increasing interest in understanding the role white matter plays in epileptogenesis and seizure propagation, diffusion tensor imaging may yield useful information about changes in white matter organization. Our current multimodality imaging combines these structural imaging advances with coregistered neurophysiological information obtained with magnetoencephalography and with simultaneous EEG recordings. Our initial experience suggests that multimodality imaging will improve our ability to detect and define the extent of epileptogenic lesions. PMID- 15281952 TI - 1H and 31P spectroscopic imaging of epilepsy: spectroscopic and histologic correlations. AB - Although MRS measurements are useful in assessing the biochemical alterations underlying human epilepsy, to date their use has been limited primarily by three factors: (a) the lack of widespread methods and appropriate hardware for acquiring high-resolution spectroscopic imaging data, (b) difficulties in spectral interpretation associated with metabolic heterogeneity, and (c) difficulties in biological interpretation due to a lack of correlative histologic studies. In this work, we (a) describe approaches to overcome these hurdles, and (b) discuss the biological interpretation of the spectroscopic findings in TLE. PMID- 15281953 TI - Ambiguous language in Wada evaluations. AB - A case is presented with ambiguous cerebral language lateralization, as determined by the intracarotid amobarbital test. Reasons for ambiguous language findings for this test are discussed. PMID- 15281954 TI - fMRI: applications in epilepsy. AB - Functional MRI (fMRI) methods use currently standard MRI scanning hardware to detect changes in regional blood flow and metabolism that accompany regional brain activation. Major applications of fMRI in epilepsy include the localization of task-correlated language and memory function, and the localization of ictal and paroxysmal phenomena. Language lateralization by fMRI provides comparable results to intracarotid amobarbital testing, and memory lateralization by fMRI also shows promise. The recent development of methodologies to allow interpretable electroencephalographic data to be recorded during MRI scanning has opened up new opportunities for combining the spatial resolution of imaging with the temporal resolution of electrophysiology in seizure localization. PMID- 15281955 TI - Ictal SPECT in nonlesional extratemporal epilepsy. AB - PURPOSE: Ictal single-photon emission computed tomography (SPECT) may be a reliable indicator of the ictal onset zone in patients with intractable partial epilepsy who are being considered for epilepsy surgery. The rationale for the illustrated case report is to evaluate the use of an innovation in SPECT imaging in a patient with nonlesional extratemporal epilepsy. METHODS: We investigated the presurgical evaluation and operative outcome in a patient with intractable partial epilepsy. The ictal semiology indicated a "hypermotor" seizure with bipedal automatism. The electroclinical correlation and magnetic resonance imaging (MRI) did not suggest the appropriate localization of the epileptogenic zone. A subtraction periictal SPECT coregistered to MRI (SISCOM) was peformed. RESULTS: SISCOM revealed a region of localized hyperperfusion in the right supplementary sensorimotor area. Chronic intracranial EEG monitoring confirmed the relationship between the localized SISCOM alteration and the ictal onset zone. The patient was rendered seizure free after surgical treatment. CONCLUSIONS: SISCOM may be used to identify potential candidates for surgical treatment of nonlesional extratemporal epilepsy. Periictal imaging may also alter the strategy for intracranial EEG recordings and focal cortical resection. PMID- 15281956 TI - Ictal SPECT. AB - The localizing value of ictal single-photon emission computed tomography (SPECT) performed with cerebral blood flow agents in patients with epilepsy is based on cerebral metabolic and perfusion coupling. Ictal hyperperfusion is used to localize the epileptogenic zone noninvasively, and is particularly useful in magnetic resonance (MR)-negative partial epilepsy and focal cortical dysplasias. Subtraction ictal SPECT coregistered with MRI (SISCOM) improves the localization of the area of hyperperfusion. Ictal SPECT should always be interpreted in the context of a full presurgical evaluation. Early ictal SPECT injections minimize the problem of seizure propagation and of nonlocalization due to an early switch from ictal hyperperfusion to postictal hypoperfusion during brief extratemporal seizures. The degree of thresholding of SISCOM images affects the sensitivity and specificity of ictal SPECT. Ictal hypoperfusion may reflect ictal inhibition or deactivation. Postictal and interictal SPECT studies are less useful to localize the ictal-onset zone. Statistical parametric mapping analysis of groups of selected ictal-interictal difference images has the potential to demonstrate the evolution of cortical, subcortical, and cerebellar perfusion changes during a particular seizure type, to study seizure-gating mechanisms, and to provide new insights into the pathophysiology of seizures. PMID- 15281957 TI - Tuberous sclerosis and multiple tubers: localizing the epileptogenic zone. AB - Tuberous sclerosis complex (TSC) is associated with medically refractory seizures and developmental delay in children. These epilepsies are often resistant to antiepileptic drugs (AEDs), may be quite severe, and usually have a negative impact on the child's neurological and cognitive development. It is believed that functional outcome is improved if seizures can be controlled at an early age. The surgical treatment of intractable epilepsy in children and adults with TSC has gained significant interest in recent years. Previously published studies have shown a potential benefit from resection of single tubers, with most of the results noted in relatively older children. All of these reports support the idea that if a single primary epileptogenic tuber/region can be identified, then a surgical approach is appropriate. However, most children with TSC have multiple potentially epileptogenic tubers, rendering localization challenging, and they are therefore rejected as possible surgical candidates. We have utilized a novel surgical approach using invasive intracranial monitoring, which is typically multistaged and bilateral. Here we present an illustrative case of a young boy with TSC and medically refractory epilepsy who underwent a staged surgical approach. This multistage surgical approach has been useful in identifying both primary and secondary epileptogenic zones in TSC patients with multiple tubers. Multiple or bilateral seizure foci are not necessarily a contraindication to surgery in selected TSC patients. Long-term follow-up will determine whether this approach has durable effects. We await better methods for identifying the epileptogenic zone, both noninvasive and invasive. PMID- 15281958 TI - Optical imaging of epileptiform activity in human neocortex. AB - The surgical outcomes of patients suffering from neocortical epilepsy are not as successful as the surgical outcomes from resections of epilepsy patients with mesial temporal sclerosis. The main difficulty in the treatment of neocortical epilepsy is that current technology has limited accuracy in mapping neocortical epileptogenic tissue. It is known that the optical spectroscopic properties of brain tissue are correlated with changes in neuronal activity. The method of mapping these activity-evoked optical changes is known as imaging of intrinsic optical signals (IIOS). Activity-evoked optical changes measured in neocortex are generated by changes in cerebral hemodynamics (i.e., changes in blood oxygenation and blood volume). Our experimental approach was to acquire high-resolution IIOS maps of epileptiform activity in patients undergoing surgery for medically intractable neocortical epilepsy. Both spontaneous and stimulation-evoked epileptiform activity was monitored. Imaging of intrinsic optical signals was able to localize neocortical epileptic foci precisely by using changes in blood volume in contrast to changes in blood oxygenation. IIOS has the potential to translate from a purely research tool to a new intraoperative approach for the surgical treatment of neocortical epilepsy. PMID- 15281959 TI - Laminar analysis of human neocortical interictal spike generation and propagation: current source density and multiunit analysis in vivo. AB - Multicontact microelectrodes were chronically implanted in epilepsy patients undergoing subdural grid implantation for seizure localization. Current source density and multiple unit activity of interictal spikes (IISs) were sampled every approximately 150 microm in a line traversing all layers of a cortical column. Our data suggest that interictal epileptiform events in humans are initiated by large postsynaptic depolarizations, consistent with the hypothesis that human IISs correspond to animal paroxysmal depolarization shifts. Furthermore, the cortical layer where the initial depolarization occurs may differ according to whether the IIS is locally generated or propagated from a distant location, and among the propagated IISs, whether the IIS is in the direct path of propagation or on the periphery of that path. PMID- 15281960 TI - The utility of magnetoencephalography in the evaluation of secondary bilateral synchrony: a case report. PMID- 15281961 TI - Magnetoencephalography in epilepsy. AB - Magnetoencephalography (MEG)-also known as magnetic source imaging when combined with magnetic resonance imaging-has developed to the point that it has now entered routine clinical application. Epilepsy MEG studies show that it can accurately localize spike sources--both ictal and interictal--as compared to both direct (intracranial EEG) and indirect (imaging abnormalities) measures. Challenges remain with difficulties in detecting complex or deep sources when recording spontaneous cerebral activity. Magnetoencephalography not only provides a novel tool to localize and characterize epileptiform disturbances, it also has an important role in determining the significance of abnormalities seen on both structural and functional imaging. Combined with mapping of normal or eloquent brain function, MEG should ultimately play a major role in the totally noninvasive epilepsy surgery evaluation. PMID- 15281962 TI - Status epilepticus and periictal imaging. AB - Peri- and postictal changes on both anatomic and functional imaging examinations have been recognized for many years. With the wide availability of magnetic resonance imaging and positron emission tomography, a growing range of recognized acute imaging findings have been described. Periictal and postictal findings can be classified as either local or remote, with respect to the site of maximal ictal EEG abnormality. Although many of the findings described are reversible, the factors that determine whether findings will resolve are incompletely understood. This article considers the range of findings that have been described, places them into the context of known or hypothesized pathophysiologic mechanisms, and considers their clinical significance. A framework is proposed for considering the relation between ictal duration and severity, the characteristics of imaging abnormalities, and the mechanism of their underlying pathophysiology. PMID- 15281963 TI - Changing management of retinoblastoma. PMID- 15281964 TI - Rainbows, pots of gold and target pressures in glaucoma. PMID- 15281965 TI - Evaluation of surveillance methods for an epidemiological study of contact lens related microbial keratitis. AB - PURPOSE: To evaluate surveillance methods in a pilot epidemiological study of contact lens related microbial keratitis (MK) cases identified by ophthalmic practitioners in Australia and New Zealand between May and August 2003 inclusive. METHODS: Twelve ophthalmologists and 55 optometrists from rural and metropolitan locations were sent a study information pack with postal reporting forms. After 2 months, practitioners were emailed a link to a website for Internet reporting. After 4 months, practitioners were prompted by email and then by telephone if a response was not received. Passive response rates were the rate of returns after posting information and emailing the website link. Active response rates included personalized email and telephone follow-up. RESULTS: Ten cases of MK were identified by optometrists and five by ophthalmologists. The passive response rates were 79% and 58% for the first and second reporting periods, respectively. There was a lower response rate in the second reporting period compared to the first (P = 0.02). With active surveillance the response rate increased to 97% and 96%. A large proportion of optometrists (62%) and ophthalmologists (55%) used the website for at least one reporting period. Internet reporting was used by all New Zealand practitioners (5/5). CONCLUSIONS: A surveillance study to estimate the incidence of contact lens related MK in Australia and New Zealand is feasible and acceptable. Internet-based reporting offers a reliable, rapid and cost-effective means of running a large scale, international surveillance study. Active surveillance methods are necessary to enhance reporting rates. PMID- 15281966 TI - Retinoblastoma in Victoria, 1976-2000: changing management trends and outcomes. AB - PURPOSE: To describe changes in the management of retinoblastoma in Victoria and to review the effect of newer, conservative treatments on preservation of eyes, visual outcome and mortality by comparing a similar group of patients treated over successive time intervals. METHODS: A retrospective analysis of all cases of retinoblastoma diagnosed and treated in Victoria between 1956 and 2000 was conducted. Historical data on 77 cases previously published by O'Day et al. was used for the period 1956-1976 (series 1). Data on 88 cases recorded in the Royal Children's Hospital (Melbourne) Retinoblastoma Database was used for the period 1976-2000. The latter group was subdivided into those treated between 1976 and 1989 (series 2), prior to the advent of modern eye saving treatments, and those treated subsequent to their introduction from 1990 to 2000 (series 3). RESULTS: In unilateral retinoblastoma, final enucleation rates for 1956-1976 (series 1) and 1976-1989 (series 2) were almost identical, being 98% and 97% of affected eyes, respectively (P = 1.00). Despite the newer treatments used after 1990 (series 3), 88% of affected eyes were still enucleated, representing a statistically similar outcome to series 2 (P = 0.33). In bilateral retinoblastoma, primary enucleation of the more involved eye was similar for series 1 (84%) and 2 (80%) but series 3 (41%) was substantially less than series 2 (P = 0.04) following the increased use of conservative treatments. In series 3, 59% of more involved eyes were treated conservatively compared with 16% (P = 0.007) and 20% (P = 0.04) for series 1 and 2, respectively. Despite attempts at eye salvage, the failure rate was higher in series 3 (29%) yielding a final enucleation rate of 70%, which represented a modest downward trend in the numbers of eyes finally enucleated; 84% (series 1), 73% (series 2) and 70% (series 3) (test for trend, P = 0.33). Bilateral enucleation rates were significantly lower, from 36% and 30% in series 1 and 2, respectively, to 7% in series 3 (test for trend, P = 0.02). As a consequence, more eyes were preserved over time, being 20/50 (40%) in series 1, 15/30 (50%) in series 2 and 21/34 (62%) in series 3. Comparison of visual outcome was hampered by incomplete data in series 1 but it appeared series 2 and 3 achieved better visual acuities with 67% and 62% of preserved eyes in bilateral cases measuring equal to or better than 6/12. Mortality rates in all series were low, being 7.8% in series 1, 4.5% in series 2 and nil in series 3. CONCLUSIONS: Following the introduction of new conservative treatments, there has been an increase in preservation of eyes and improved visual outcome, and a dramatic decrease in numbers of bilateral enucleations without adversely affecting survival. PMID- 15281967 TI - Life expectancy of patients with neovascular glaucoma drained by Molteno implants. AB - PURPOSE: To determine the survival of patients with sighted eyes who had Molteno implants inserted for neovascular glaucoma. METHODS: Patients who had Molteno implants inserted for neovascular glaucoma between October 1977 and March 2001 at Dunedin Hospital, New Zealand, were reviewed. Relative survival analysis was then used to compare survival in this group to survival of the New Zealand population of the same age. RESULTS: A total of 114 patients had 131 Molteno implants inserted for neovascular glaucoma. Neovascular glaucoma was secondary to central retinal vein occlusion in 66 eyes (50%) and diabetes in 42 eyes (32%), and patients lost 52% or 6.53 years of their expected remaining life span. Age, sex and postoperative intraocular pressure control did not predict survival. The only predictor of improved survival was a preoperative visual acuity of 6/48 or better. These patients lost 21% (or 2.43 years) of expected remaining life compared to 62% (or 10.78 years) in those patients who had a presenting visual acuity of 6/60 or worse. CONCLUSIONS: Patients with neovascular glaucoma had a markedly reduced life expectancy, which has improved in recent years when compared to that of the normal sex and age-matched New Zealand population. Patients presenting with better vision (6/48 or better) survived significantly longer than those presenting with poorer vision. PMID- 15281968 TI - Accuracy of the Tonosafe disposable tonometer head compared to the Goldmann tonometer alone. AB - PURPOSE: To assess the accuracy of the Tonosafe disposable prism tonometer head via a randomised controlled prospective trial. METHODS: The intraocular pressure (IOP) of 69 patients (31 men, 38 women) involving 137 eyes was measured using both the Tonosafe disposable and the Goldmann tono-meter head. The average age of patients was 66.5 years (range 23-93 years). The two examiners (PM and SL) who performed tonometry were masked while a separate observer read the IOP measurements. The order was randomised between Tonosafe and Goldmann devices. The minimum interval between the two measurements was 15 min. Patients with corneal and external diseases were excluded from the study. RESULTS: The mean IOP using the Goldmann tonometer head was 17.44 +/- 4.97 mmHg. The mean IOP using the Tonosafe disposable head was 17.58 +/- 5.03 mmHg. The mean difference was 0.14 +/ 1.73 mmHg (95% CI -0.44, 0.16, P= 0.36). Subgroup analysis on eyes with IOP > 21 mmHg showed the Tonosafe disposable head was on average 0.15 +/- 2.40 mmHg higher than the Goldmann tonometer head. There was a high level of correlation between the IOPs obtained with the Goldmann and Tonosafe heads. Pearson's coefficient of correlation was 0.94 (P < 0.0001). CONCLUSION: The Tonosafe disposable prism head was found to be accurate in IOP measurement, even in the higher range. PMID- 15281969 TI - Five-year follow up of selective laser trabeculoplasty in Chinese eyes. AB - PURPOSE: To study the effectiveness and safety of selective laser trabeculoplasty (SLT) on primary open-angle glaucoma and ocular hypertension in Chinese eyes. METHODS: This was a prospective randomized controlled clinical study in which 58 eyes of 29 patients with primary open-angle glaucoma or ocular hypertension were included. One eye of each patient was randomized to receive SLT (Group 1) and the fellow eyes received medical treatment (Group 2). Patients were evaluated after laser treatment at 2 h, 1 day, 1 week, 2 weeks, 1 month, 3 months, 6 months, and then yearly. RESULTS: All patients (13 male, 16 female) were Chinese. The mean age was 51.9 +/- 14.7 years. The mean baseline intraocular pressure was 26.8 +/- 5.6 mmHg in group 1 and 26.2 +/- 4.2 mmHg in group 2 (P = 0.62). The failure rate, defined as intraocular pressure >21 mmHg with maximal medications, was 17.2% in group 1 and 27.6% in group 2 at 5-year follow-up (P = 0.53). Eight eyes (27.6%) in group 1 required medications to control the intraocular pressure to below 21 mmHg. There was no statistically significant difference in the intraocular pressure reductions between the two groups at all time intervals (P > 0.05). The mean number of antiglaucoma medications was significantly lower in the SLT than the medical treatment group up to 5 years of follow up (P < 0.001). Transient post-SLT intraocular pressure spike >5 mmHg was observed in three eyes (10.3%). CONCLUSION: With fewer medications, SLT gives similar intraocular pressure reduction to medical therapy alone in Chinese patients with primary open angle glaucoma or ocular hypertension. PMID- 15281970 TI - Short-term effect of latanoprost on ocular circulation in ocular hypertension. AB - PURPOSE: To determine the short-term effects of latanoprost on retrobulbar circulation in ocular hypertension. METHODS: Forty-six eyes of 23 consecutive bilateral ocular hypertensive patients with an intraocular pressure (IOP) of greater than 22 mmHg were evaluated in a prospective controlled study. All subjects received a single drop of latanoprost 0.005% in one eye and placebo in the fellow control eye. Systemic circulatory parameters, intraocular pressure, blood flow velocities, and resistance indices of the ophthalmic, short posterior ciliary and central retinal arteries were measured using colour Doppler imaging at baseline and 2 h and 8 h after dosing. RESULTS: Latanoprost lowered IOP significantly after 2 h and 8 h (P < 0.01). The mean IOP reduction was 6.7 mmHg 8 h after dosing. At baseline, there were no statistically significant differences in any retrobulbar vessels of eyes that received a single drop of latanoprost when compared with the eyes that received placebo (P > 0.05). Comparisons with baseline and latanoprost conditions revealed that latanoprost did not alter the blood flow velocities and resistance indices in the ophthalmic (P > 0.05), posterior ciliary (P > 0.05) and central retinal (P > 0.05) arteries 2 h and 8 h after dosing. The systolic and diastolic blood pressures (p = 0.74, p = 0.29, respectively) and pulse rate (p = 0.68) remained unchanged over the 8-h period. CONCLUSIONS: This study found that a single drop of latanoprost significantly reduces intraocular pressure 8 h after dosing. However, it does not have any short-term effects on the retrobulbar haemodynamics in ocular hypertensive eyes. PMID- 15281971 TI - Operative revision of non-functioning filtering blebs with 5-fluorouracil to regain intraocular pressure control. AB - PURPOSE: To determine the efficacy of extensive microsurgical needling revision of failed filtering blebs followed by serial 5-fluorouracil subconjunctival injections. METHODS: Thirty-six eyes of 34 consecutive patients with progressive open-angle glaucoma refractory to topical therapy submitted to needling revision as a major procedure. All patients required multiple antiglaucoma medications preoperatively, and had completely flat or densely encapsulated filtering blebs. All patients underwent elaborate needling revision (limbus to superior rectus >8 mm diameter, >3 mm elevation, entry-site sutured with 8-0 vicryl and bleb reformed via paracentesis with viscoelastic) in the operating room, followed by serial 5-fluorouracil. The patients were followed for up to 6 months postoperatively. The main outcome measures were intraocular pressure (IOP) and the number of antiglaucoma medications used. RESULTS: Thirty-one eyes (86%) maintained mean IOP below 15 mmHg postneedling without medication. Overall the mean IOP postneedling was >9 mmHg lower than medicated preoperative levels (P < 0.0001). IOP reduction in encapsulated blebs was marginally superior to that in flat blebs. CONCLUSIONS: Extensive needling revision in the operating room is safe, straightforward, and produces reproducible restoration of filtering function. PMID- 15281972 TI - Systemic and ocular comorbidity of cataract surgical patients in a western Sydney public hospital. AB - BACKGROUND: This study aimed to assess the frequency of major systemic and ocular comorbidities among cataract surgical patients attending a large general public hospital. METHODS: Consecutive patients aged 60+ years who had undergone cataract surgery at Westmead Hospital from July 2001 to December 2002 were included. Preoperative information was obtained from patient medical records. RESULTS: A total of 773 cataract surgical procedures were performed during the 18-month period, including 668 (86.4%) aged 60+ years. Complete data were available for 653 eyes (97.8%); 62.2% were women. The mean age at surgery was 74.6 (+/- 7.2) years. Frequent systemic comorbidities included: angina (20.2%), previous myocardial infarct (15.0%), diabetes (27.5%) and hypertension (56.3%); 12.5% were current smokers. Major preoperative ocular comorbidities included age-related maculopathy (12.6%), diabetic retinopathy (9.0%) and glaucoma (10.6%). Preoperatively, 21.7% of this group had severe visual impairment (visual acuity [VA] <6/60), 30.6% had moderate impairment (VA <6/24 ->or=6/60) and 30.6% had mild impairment (VA <6/12 ->or=6/24); 28.5% had presenting VA >or=6/12. CONCLUSION: The data indicate a high frequency of comorbid systemic and ocular diseases among cataract surgical patients. PMID- 15281973 TI - Cataract surgery in Australia: a profile of patient-centred outcomes. AB - BACKGROUND: Few studies have addressed quality of life or satisfaction outcomes for patients undergoing cataract surgery, particularly in Australia. The purpose of this study was to describe a sample of patients undergoing cataract surgery in typical metropolitan practices in Sydney, with a particular focus on the impact upon quality of life and satisfaction with vision. METHODS: One hundred and eleven patients were recruited prior to surgery and followed through 3 months postoperatively. Patients reported basic demographic information, VF-14 visual disability and SF-36 quality of life information, along with the degree of satisfaction and trouble with vision they experienced. Surgeons' records provided information about type and severity of cataract, refractive error, ocular comorbidity and visual acuity. RESULTS: Patients enjoyed strongly significant improvement in visual acuity, disability, trouble and satisfaction with vision, with a median postoperative Snellen acuity of 6/7.5, and 82% within 1 D refraction. Not wearing glasses was the most commonly stated patient goal for undergoing surgery. Prior to surgery 23% of all driving patients did so illegally due to poor vision; after surgery 21% of non-drivers began driving again, all legally. Nonetheless, quality of life did not improve. Those who failed to achieve improvements in satisfaction with vision were more likely to be female, have lower educational attainment or have high visual function preoperatively. Change in visual acuity was not predictive. CONCLUSIONS: The findings from this study indicate that cataract surgery outcomes in Australia compare well with international standards, and emphasize the inadequacy of visual acuity to measure relevant surgical outcomes. Increased preoperative counselling may be required in those groups less likely to attain high levels of satisfaction. Finally, the role of cataract surgery to improve quality of life must be investigated further, as this is the ultimate goal of the procedure. PMID- 15281975 TI - Auckland proliferative diabetic vitrectomy fellow eye study. AB - BACKGROUND: To review medical and ophthalmic findings of primary diabetic vitrectomy patients to examine indices important in progression to fellow eye surgery. METHODS: A retrospective analysis was undertaken of all diabetic patients undergoing vitreoretinal surgery at Auckland Public Hospital between January 1992 and July 1996. Kaplan-Meier survival analysis was performed along with univariate and multivariate (Cox Proportional Hazards) data analysis. RESULTS: One hundred and fourteen primary diabetic vitrectomy cases were reviewed with mean follow-up duration of 4 years. Thirty-eight per cent (n = 43) of the study group underwent fellow eye surgery at a mean time of 1.6 years after first eye surgery. Fourteen patients were already blind in the fellow eye at baseline, and five patients refused second eye surgery on intention to treat. Thus there were 62 (54%) patients with severe (surgical threshold) fellow eye disease diagnosed within the follow-up period. The presence of either tractional retinal detachment or combined rhegmatogenous/tractional retinal detachment but without vitreous haemorrhage in the presenting eye was, in this series, a risk factor for fellow eye surgery (OR 5.56; 95% CI 1.96-15.8). Maori and Pacific Islander ethnicity was significantly associated with traction retinal detachment (OR 2.23; 95% CI 1.05-4.7). At data analysis 57% (n = 60) of the study patients had died. The mean time to death was 4.3 years, with 84% of these patients having evidence of renal disease at the time of their first eye surgery. Good visual function in at least one eye was maintained in many patients. CONCLUSIONS: A substantial proportion of diabetic vitrectomy patients require fellow eye surgery. Absence of vitreous haemorrhage in the presenting eye (i.e. tractional or combined rhegmatogenous/tractional retinal detachments but without vitreous haemorrhage) was predictive of need for fellow eye surgery. The need for diabetic vitrectomy correlates with poor survival in this study population. PMID- 15281974 TI - Photoscreening for diabetic retinopathy: a comparison of image quality between film photography and digital imaging. AB - PURPOSE: Retinal images from patients attending an urban screening centre before and after the transition from film photography to digital image acquisition were analysed for quality of image. METHODS: A total of 1946 diabetic patients, aged 12-92 years (mean 55.6 +/- 14.88 years), were included in this retrospective study of retinal screening techniques. Each imaging group was subdivided into age matched groups. In all subjects pupils were pharmacologically dilated before photography. The images were reviewed by the same three experienced observers and graded at the time of screening from grade 1 (excellent quality) to grade 4 (unreadable). RESULTS: Of 938 patients in the film group, 31.3% had excellent images, 38.2% good, 22.7% poor and 7.8% were unreadable. Of the 1008 patients in the digital imaging group, 25.3% had excellent images, 46.3% good, 14.6% poor and 13.8% were unreadable. A significant difference was observed in patients over 65 years of age who exhibited a threefold increase in failure rate with digital imaging (33.7% v 11.3%)(P < 0.0001). CONCLUSION: In this study population a statistically significant degradation of image quality was observed in those older than 65 years following transition to digital photography. This has implications for service provision planning. PMID- 15281976 TI - Determination of relative contribution of the superior and inferior canaliculi to the lacrimal drainage system in health using the drop test. AB - PURPOSE: The aim of this study was to explore the use of the 'drop test' as a method of assessing maximal lacrimal outflow capacity, and to measure the relative contribution of the superior and inferior canaliculi to the drainage capacity in normal subjects. METHOD: The drop test involves instilling measured aliquots of normal saline over 3-min periods to raise the tear lake medially. Both eyes were assessed; however, for the right lacrimal drainage system only, sequential insertion and then removal of silicone punctal plugs was performed. After each stage the maximal lacrimal drainage capacity was measured. no punctal plugs were placed in the puncta on the left side. RESULTS: Complete data were collected from 20 subjects with a mean age of 35.6 years. The intraclass correlation coefficient for the five left eye readings was 0.98 (CI 0.96-0.99) and the limits of agreement of a single reading were -22.6 to +93.0 micro L/3 min. Without intervention, no statistically significant difference was found in the mean lacrimal outflow between the left and right eyes (P = 0.16). A statistically significant reduction in outflow resulted from punctal occlusion (P < 0.05). Presenting the proportion of lacrimal outflow as a percentage of the combined values of the superior and inferior canaliculi, 59.9% of outflow occurred through the inferior canaliculus. CONCLUSION: The drop test was found to provide a simple and repeatable method of assessing lacrimal drainage in a minimally invasive manner in the clinical setting. In healthy volunteers in the supine position 60% of maximal lacrimal outflow capacity occurs through the inferior canaliculus. PMID- 15281977 TI - Anti-GM1 antibodies: the cause of otherwise unexplained ophthalmoplegias? AB - PURPOSE: To report four patients with otherwise unexplained ophthalmoplegia who were found to have elevated levels of anti-GM1 antibodies. METHODS: Retrospective chart evaluation of 50 consecutive patients with otherwise unexplained ophthalmoparesis. All patients were tested for anti-GM1 antibodies. RESULTS: Six out of 50 cases (12%) were found to have elevated anti-GM1 antibodies, far greater than the 2.7% found in the general population. Four of the subjects with elevated anti-GM1 antibodies are reported. CONCLUSION: Although the mechanism and relationship between the neural antibodies and ophthalmoparesis is not understood, it is proposed that the presence of anti-GM1 antibodies should be considered in cases of otherwise unexplained ophthalmoplegia. PMID- 15281978 TI - Evaluation of polytetrafluoroethylene suture for frontalis suspension as compared to polybutylate-coated braided polyester. AB - PURPOSE: To evaluate the efficacy of polytetrafluoroethylene (Gore-Tex) suture as compared to polybutylate-coated braided polyester (Ethibond) suture as sling materials for frontalis suspension in bilateral congenital ptosis. METHODS: Frontalis sling surgery by modified Crawford's double triangle technique was performed on 30 patients (60 eyes) with bilateral ptosis. The patients were randomized into two groups depending on the type of suture material used: polytetrafluoroethylene or braided polyester. RESULTS: Polytetrafluoroethylene suture achieved a statistically significant better and more stable ptosis correction, with slightly more lagophthalmos, than braided polyester suture over a mean (+/- SD) follow-up period of 16 +/- 3.24 months. There were more postoperative complications with braided polyester suture, but the difference was not statistically significant. CONCLUSION: This is the first clinical study in which polytetrafluoroethylene in the form of suture has been studied. Polytetrafluoroethylene suture was found to be a safe and effective sling material for frontalis suspension and it can be recommended for clinical use. PMID- 15281979 TI - External ophthalmic findings in multiple endocrine neoplasia type 2B. AB - PURPOSE: To identify external ophthalmic abnormalities in multiple endocrine neoplasia type 2B (MEN2B), which may facilitate early detection and prophylactic thyroidectomy to prevent medullary thyroid carcinoma. METHODS: Three patients with MEN2B were examined by an oculoplastic surgeon and external ophthalmic findings were recorded. RESULTS: All patients had prominent corneal nerves, thickened eyelids, mild ptosis and eversion of the upper eyelids. Two patients displayed eyelid nodules and one exhibited lower lid margin eversion. CONCLUSION: Ptosis and lower lid margin eversion are previously unreported findings in patients with MEN2B. Medullary thyroid carcinoma is the most serious consequence of MEN2B and has a high mortality if untreated. Early diagnosis and prophylactic thyroidectomy may be lifesaving. Gene mutations can be identified but the sporadic tendency of the syndrome emphasizes the importance of early clinical detection. MEN2B is one of a number of systemic malignancies with ophthalmic manifestations. Ophthalmologists should be aware of the external features of this rare but lethal malignancy. PMID- 15281980 TI - Trimethoprim-sulphamethoxazole therapy in Nocardia keratitis. AB - PURPOSE: To describe the clinical features, microbiological features and treatment outcome of nine patients with Nocardia keratitis treated with topical trimethoprim- sulphamethoxazole drops. METHODS: Retrospective review of nine patients with culture-proven Nocardia keratitis. RESULTS: Nine patients with Nocardia keratitis were treated with topical trimethoprim-sulphamethoxazole drops. The average duration of treatment was 25 +/- 9 days. Five of the nine patients presented with superficial ulcers with margins studded with yellowish white discrete pinhead sized infiltration; the other four patients had deep stromal infiltration. Complete healing of the ulcer was achieved in six out of the nine patients with topical trimethoprim- sulphamethoxazole alone or in combination with ciprofloxacin 0.3% eye drops. CONCLUSION: Topical application of trimethoprim-sulphamethoxazole appears to be effective therapy for superficial keratitis due to Nocardia. PMID- 15281981 TI - Two cases of X-linked juvenile retinoschisis with different optical coherence tomography findings and RS1 gene mutations. AB - The optical coherence tomography (OCT) findings, clinical features, and mutations in the RS1 gene of two unrelated patients with X-linked retinoschisis (XLRS) are reported herein. Two Chinese patients with early onset XLRS were given a comprehensive ophthalmologic examination and OCT investigation. The RS1 gene was screened for sequence alterations in all exons and splice regions. The two patients presented with different phenotypic features and OCT findings. One patient with more severe clinical presentation had a RS1 exon 1 deletion and a P193S mutation was found in the other patient with mild macular involvement. OCT demonstrates the markedly different features of XLRS patients with different RS1 mutations. This study strengthens the role of OCT in the diagnosis and monitoring of XLRS. PMID- 15281982 TI - Pseudogerontoxon. AB - Pseudogerontoxon is a lesion that resembles a small segment of arcus senilis or gerontoxon and is seen in many individuals with limbal vernal or atopic keratoconjunctivitis. It is an important clinical finding because pseudogerontoxon is often times the only clinical evidence of previous allergic eye disease. To the authors' knowledge, pseudogerontoxon has only rarely been mentioned, and has never been pictured, in the peer-reviewed literature. Three examples are presented of this important clinical entity. PMID- 15281983 TI - Combined central retinal artery and vein occlusion from orbital inflammatory pseudotumour. AB - Progressive, painful blurred vision and proptosis developed in the left eye of a 32-year-old man over an 8-day period. On initial neuro-ophthalmic evaluation the visual acuity in the left eye was no light perception. Erythema of the periorbital skin, 5 mm of proptosis, and ophthalmoplegia were noted on the left side. Funduscopy revealed occlusion of the central retinal artery and central retinal vein. Computed tomography and magnetic resonance imaging revealed marked distension of the left optic nerve and enhancement of the contents within the left orbit, with clear paranasal sinuses. The diagnosis of orbital inflammatory pseudotumour was made and the orbital signs improved rapidly with 80 mg of prednisone per day. Combined occlusion of the central retinal artery and vein is a rare complication of orbital inflammatory pseudotumour. It is postulated that marked distension of the optic nerve caused mechanical compression of the central retinal vessels leading to the vascular occlusions. PMID- 15281984 TI - Shock-induced anterior ischaemic optic neuropathy after radical prostatectomy. AB - The case is reported of a 61-year-old man who developed shock-induced anterior ischaemic optic neuropathy (SIAION) after undergoing radical prostatectomy. Visual loss began on the third postoperative day. Only one other patient with SIAION after prostatectomy has been reported in the past. SIAION may result in fixed visual loss after radical prostatectomy and ophthalmic evaluation should not be delayed in patients with postoperative visual loss. PMID- 15281985 TI - Upper eyelid oedema in Melkersson-Rosenthal syndrome. AB - Melkersson-Rosenthal syndrome is an uncommon disorder of uncertain aetiology characterized by orofacial oedema, facial nerve palsy and lingua plicata. The triad is seldom seen in its complete form, and oligo-symptomatic or mono symptomatic forms are more common. An unusual case of Melkersson-Rosenthal syndrome involving the left upper eyelid is presented. The pathology, clinical features and management of this disorder are discussed. PMID- 15281986 TI - Churg-Strauss syndrome presenting with conjunctival nodules in association with Candida albicans and ankylosing spondylitis. AB - Churg-Strauss syndrome is a rare diffuse vasculitis of which the ocular manifestations have been well documented. However, reports of conjunctival involvement in Churg-Strauss syndrome are scarce. Such a presentation is described in a man with Candida albicans infection as well as ankylosing spondylitis, and a possible aetiological linkage is established amongst all three. PMID- 15281987 TI - Choroidal neovascularization following hyperopic LASIK surgery. AB - Laser in situ keratomileusis (LASIK) is a well-accepted refractive surgical procedure; however, reports of complications of hyperopic surgery are scarce. This is the first report of bilateral macular choroidal neovascular membrane following bilateral hyperopic LASIK surgery. It highlights important considerations in older patients undergoing the procedure. PMID- 15281988 TI - Three cases of post-traumatic endophthalmitis caused by unusual bacteria. AB - Three cases of post-traumatic endophthalmitis caused by unusual bacteria are presented. The pathogens identified were: (i) Bacillus cereus and Citrobacter freundii; (ii) Pseudomonas fluorescens; and (iii) Chryseobacterium meningosepticum and Stenotrophomonas maltophilia. Two of these pathogens have not previously been reported to cause endophthalmitis. The available literature regarding the individual cases is summarized and a brief discussion of post traumatic endophthalmitis is presented, with reference to a recently published large series at the authors' institution. PMID- 15281989 TI - Orbital metastasis from transitional cell carcinoma of the bladder. AB - Transitional cell carcinoma (TCC) is a common bladder tumour, 10-15% of which will exhibit invasive behaviour. About 50% of patients with invasive TCC will eventually develop distant metastases, usually to lymph nodes, lung or bones. The case is reported of bladder TCC metastasizing to the orbit. PMID- 15281990 TI - Review of Australian indigenous eye health research published in the last decade. PMID- 15281991 TI - Comment on 'Sub-Tenon infiltration or classical analgesic drugs to relieve postoperative pain'. PMID- 15281993 TI - Influence of lesion size on treatment benefit with photodynamic therapy. PMID- 15281994 TI - Editorial comment. PMID- 15281995 TI - The F A Maguire collection at the RANZCOG. PMID- 15281996 TI - Single or multiple embryo transfer following in vitro fertilisation for improved neonatal outcome: a systematic review of the literature. AB - BACKGROUND: The aim of the current review was to determine if single versus two or more embryos, or double versus three or more embryos, transferred to the woman of a subfertile couple at in vitro fertilisation (IVF) maximises the likelihood of pregnancy, while minimising the likelihood of multiple pregnancy and adverse sequelae. METHODS: Studies were identified that reported maternal, infant and cost outcomes following embryo transfer at IVF. RESULTS: Three randomised trials and 17 cohort studies were included. From two randomised trials, single embryo transfer was found to result in decreased incidence of clinical pregnancy, multiple pregnancy and low birthweight. In the cohort studies for single embryo transfer compared with transfer of two or more embryos the incidence of live birth and singleton pregnancies was unchanged, and the incidence of multiple pregnancies and low birthweight was reduced. For double embryo transfer compared with the transfer of three or more embryos, the incidence of clinical pregnancy, live birth, preterm birth and low birthweight babies was reduced. CONCLUSIONS: Information on neonatal and maternal outcomes following transfer of different numbers of embryos is limited. Transfer of one embryo does not alter the likelihood of a singleton pregnancy or birth when compared to transfer of two or more embryos. Transfer of one or two embryos decreases the risk of a multiple pregnancy, preterm birth and low birthweight. Further large, well-designed randomised trials are required to provide maternal and neonatal outcomes of relevance to a couple undergoing IVF. PMID- 15281997 TI - Has a new hazard been added to failed sterilisations? PMID- 15281998 TI - Obstetric medicine, its premise and promise. PMID- 15281999 TI - Effects of coasting on the outcome of intracytoplasmic sperm injection-embryo transfer cycles. AB - OBJECTIVES: To determine the effects of 'coasting' on the outcome of controlled ovarian hyperstimulation (COH) and intracytoplasmic sperm injection-embryo transfer (ICSI-ET). DESIGN: Retrospective study. SETTING: IVF Centre, Ozel Ege Tup Bebek Merkezi, Izmir, Turkey. SAMPLE: Twenty-six coasted and 52 non-coasted COH and ICSI-ET patients were enrolled in this retrospective study. METHODS: Coasted patients were enrolled consecutively during the study period, and two non coasted controls were selected from our database for each coasted patient. Coasting was decided when serum oestradiol level was > or = 4000 pg/mL. Groups were compared using chi2 and Mann-Whitney U-tests for statistical analysis. MAIN OUTCOME MEASURES: Number of oocytes collected, metaphase II (MII) oocytes and cleaving embryos, the fertilisation rate and clinical pregnancy rate were the main outcome measures. RESULTS: Number of oocytes collected, number of MII oocytes, number of cleaving embryos, fertilisation rate and clinical pregnancy rate for the coasted and non-coasted groups were 15.5 +/- 5.2 and 14.0 +/- 7.1, 9.7 +/- 4.8 and 9.3 +/- 3.9, 6.8 +/- 3.9 and 5.8 +/- 3.1, 0.85 +/- 0.18 and 0.78 +/- 0.18, 13/26 and 24/52, respectively; these differences were not statistically significant. None of the patients in the coasted group were hospitalised for signs or findings of severe ovarian hyperstimulation syndrome. CONCLUSIONS: Coasting does not adversely affect the number and the function of mature oocytes and the clinical pregnancy rate. PMID- 15282000 TI - Obstetric and perinatal outcomes in pregnancies associated with illicit substance abuse. AB - OBJECTIVE: To determine the obstetric and perinatal outcomes of women using illicit drugs during pregnancy by substance group. METHOD: A retrospective audit of obstetric and perinatal outcomes in women who used opiates or amphetamines during their pregnancy and delivered at King Edward Memorial Hospital (KEMH), Perth, Australia between December 1997 and April 2000 was performed. Maternal, fetal and neonatal parameters were assessed. These were compared with obstetric and perinatal data recorded by the Health Department of Western Australia (HDWA) for the 25,291 deliveries of 25,677 infants in 1998. RESULTS: Between December 1997 and April 2000 91 opiate-using and 50 amphetamine-using women were identified and included in the analysis. Both groups of drug-using women were younger (opiates P = 0.001, amphetamines P = 0.001) than the general population. There was a higher incidence of aboriginality (P = 0.001) in the amphetamine group. In the opiate-using group multiparity (P = 0.0001) and anaemia (P = 0.0001) were higher. Illicit drug-using women had a higher incidence of hepatitis C (opiates P = 0.001, amphetamines P = 0.003), and a greater need for pharmacological analgesia for labour and delivery (opiates P = 0.007, amphetamines P = 0.042). Their infants were significantly more likely to deliver at less than 37 weeks' gestation (opiates P = 0.0001, amphetamines P = 0.001), to have a birthweight of less than 2.5 kg (P = 0.0001), be small for gestational age and require admission to the special care nursery (P = 0.0001). Infants born to women in the amphetamine group were more likely to have an Apgar score < 7 (P = 0.0001) recorded. Infants of women in the opiate group required more resuscitation (P = 0.05). CONCLUSION: Women who use illicit drugs are more likely to experience adverse obstetric and perinatal outcomes than women in the general population. Differences are seen depending on the type of illicit drug used. These findings need to be replicated in a larger prospective cohort to highlight management requirements of these women and their infants. Further information is required about the effects of amphetamines in pregnancy. PMID- 15282001 TI - Intrapartum pain management at the Royal Hospital for Women. AB - AIMS: To explore use of, and women's satisfaction with, intrapartum pain management at Royal Hospital for Women (RHW), Sydney, Australia. METHODS: From October 2002 to January 2003 women aged over 16 who had been in labour at RHW were given a questionnaire to complete in the first week post-partum regarding their intrapartum pain management. Supplementary information was obtained from patient records. RESULTS: A total of 496 women participated (69% response rate), including 95 birth centre clients. The mean age was 32 years and 73% percent had a normal vaginal delivery. At least one form of pain management ('natural', nitrous oxide, pethidine, epidural, local infiltration of the perineum) was used by 463 (93%) women, with 74% using two or more methods. Labour pain was 'worse' or 'much worse' than expected for 55%. Seventy-two percent were 'very' or 'quite' satisfied with overall pain management. Epidural analgesia had the highest utility scores (89%'very useful') and likelihood of use in subsequent labours (67%), and pethidine the lowest. Factors affecting analgesic use included cervical dilation on admission, labour length, English-speaking background, delivery suite versus birth centre care, parity, and syntocinon use. CONCLUSIONS: Women at RHW use a variety of pain management methods in labour and most use multiple methods. Labour was rated more painful than expected by a majority; however, most were satisfied with their pain management. Labour length and cervical dilation on admission were most predictive of pain management use. PMID- 15282002 TI - Women's antenatal knowledge and plans regarding intrapartum pain management at the Royal Hospital for Women. AB - AIMS: To examine women's knowledge and antenatal plans regarding intrapartum pain management options at Royal Hospital for Women (RHW), Sydney, Australia. METHODS: From October 2002 to January 2003 women aged over 16 who had been in labour were given a questionnaire to complete in the first week post-partum regarding intrapartum pain management. This included questions regarding their antenatal knowledge and predetermined plans. RESULTS: There were 496 participating women (69% response rate). Antenatal pain management information was accessed by 98% of women. Sources most accessed were antenatal classes (55%), multimedia (53%), and friends/relatives (46%). Sixty percent of women felt 'very well informed' antenatally. Women felt better informed antenatally if married/defacto, university educated, privately insured, or receiving birth centre care. Antenatally, 80% planned to use intrapartum pain management: 'natural' methods were most popular (62% planned to use), and pethidine least (49% planned against). The most common determinant against using medical methods was possible maternal side-effects. Intrapartum, 19% used 'unwanted' pain management, mostly (67%) due to increased labour pain. Increased information access was associated with significantly higher use of both 'natural' methods and epidural analgesia, as well as significantly higher satisfaction scores. CONCLUSIONS: Almost all women at RHW access information antenatally about intrapartum pain management, often from informal sources. Demographic factors affected type of information accessed and women's plans. Adequate access to information affected use of, and satisfaction with, pain management. PMID- 15282003 TI - Ethnicity and fetal growth in Fiji. AB - BACKGROUND: Indigenous Fijians and the descendants of Asian Indians constitute the two major ethnic groups in Fiji. There are differences between the two groups in perinatal outcomes. AIMS: To study fetal growth patterns in the two ethnic groups and to ascertain the influence, if any, of ethnicity on fetal growth. METHODS: A longitudinal study was carried out on women with sure dates, regular cycles, no known risk factor complicating pregnancy and having their first antenatal examination before 20 weeks. Symphysis-fundal height, biparietal diameter, abdominal circumference and femur length were measured by the same observer at recruitment and at follow-up visits until delivery. Infant measurements were recorded soon after birth. RESULTS: Indian babies were on average 795 g lighter, had 5.5 days shorter mean length of gestation and slower growth of biparietal diameter and abdominal circumference when compared to Fijian babies. Ethnicity of the mother was significantly associated with the difference in growth even after adjusting for other factors known to influence fetal growth. CONCLUSION: Given the ethnic differences in fetal growth and maturation, it would be appropriate to use ethnicity-specific standards for perinatal care in Fiji. PMID- 15282004 TI - Study of peripheral circulation in non-pregnant, pregnant and pre-eclamptic women using applied potential tomography. AB - BACKGROUND: Profound changes are known to occur in the cardiovascular system during pregnancy, involving an increase in cardiac output and a fall in peripheral resistance. In some women these adaptations may be inappropriate and this may result in pregnancy-induced hypertension and pre-eclampsia. AIMS AND METHODS: The aims of the study were to evaluate the relatively new, non-invasive technique of applied potential tomography (APT) in measurements of peripheral blood flow, to study peripheral blood flow in a sample of non-pregnant, pregnant and pre-eclamptic women, and to investigate whether the adaptive changes in the peripheral circulation are different in pre-eclampsia compared with normal pregnancy. Applied potential tomography was used to assess peripheral vascular reactivity, by monitoring fluid distribution in calf muscles during postural change. RESULTS: The APT technique was able to detect peripheral vasoconstriction in response to an increase in intramural pressure brought about by passive lowering of the leg (peripheral mechanisms). The peripheral vasoconstriction response was found to be more prominent in woman with pre-eclampsia. CONCLUSIONS: The presence of a local reflex in the lower limb had been postulated and the effect of this reflex on the peripheral circulation could be detected using APT, regardless of how it was initiated. In normal pregnant women this reflex was diminished when compared to non-pregnant women, which might contribute to the reduction in peripheral vascular resistance seen in normal pregnancy. This reflex was defective in pre-eclampsia and this lack of adaptation may be a local reflex contributing to the raised peripheral resistance, which in turn may be a factor in high blood pressure in pre-eclampsia. PMID- 15282005 TI - Tissue trauma after vaginal hysterectomy and colporrhaphy versus abdominal hysterectomy: a randomised controlled study. AB - OBJECTIVE: As the magnitude of tissue trauma can be detected by measuring the blood levels of acute phase reactants, we aimed to evaluate tissue trauma markers after abdominal hysterectomy (AH) and vaginal hysterectomy (VH). We hypothesised that VH will be associated with a reduced increase in the level of acute phase reactants than AH. METHODS: Thirty women out of 92 patients scheduled for hysterectomies between June 2002 and June 2003 were randomised into two equal groups (n = 15) of VH and AH. Their levels of C-reactive protein (CRP), alpha1 antitrypsin (alpha1-AT) and myoglobin (M) were analysed preoperatively and on the second, fourth and sixth days. RESULTS: In both methods of hysterectomy, the operating time (85.3 +/- 6.57 min in the VH group vs 69 +/- 7.54 min in the AH group, P < 0.0001), and hospital stay duration (7.2 +/- 2.5 days in the AH group, 3.1 +/- 1.1 days in the VH group, P < 0.0001) were highly significantly different from each other. Demographic parameters and other parameters which may affect tissue trauma markers were not statistically significantly different in each group. Postoperative increases in all markers were markedly high and showed a high statistical difference in both groups (P < 0.05). The postoperative CRP and M values in both groups were significantly higher in the AH group on the second and fourth days and on the sixth day for M only, whereas alpha1-AT levels were only statistically different on the second day. The tissue trauma markers returned to normal levels on the sixth postoperative day for M, although there still was a statistically significant difference, but remained higher than normal for alpha1-AT and CRP. CONCLUSION: Whenever possible, VH should replace AH because this technique leads to a shorter hospital stay and less tissue trauma, enabling patients to return to their normal lives. PMID- 15282006 TI - Presence of a single fetal major anomaly in a twin pregnancy does not increase the preterm rate. AB - OBJECTIVE: To evaluate whether the presence of one major anomaly in a twin pregnancy would affect the perinatal outcome of the unaffected co-twin. MATERIALS AND METHODS: From 1992 May to July 2003, a total of 1400 twin pregnancies were included in the present study and there were 35 pairs of twins with one major anomaly. Major anomaly was defined as the anomaly that had a significant impact on neonatal morbidity and mortality. The perinatal outcomes of the affected and unaffected co-twins, the gestational age of diagnosis of the anomaly and the gestational age of delivery were the parameters for evaluation. RESULT: The incidence of a twin with one major anomaly was 2.5%. Nineteen of 35 (54.3%) affected twins and five of the 35 (14.3%) unaffected co-twins suffered a perinatal death. In the five cases of unaffected co-twins suffering a perinatal death, four were intrauterine fetal deaths and one was a neonatal death. Three of the perinatal deaths of the unaffected co-twins could be attributed to twin-to twin transfusion syndrome. The gestational age at delivery, the perinatal mortality rate, and the incidence of low 5 min Apgar scores of the unaffected co twins were not different from those in twin pregnancies without a major anomaly. CONCLUSION: The perinatal outcomes of the unaffected co-twin were not affected by the fact that its counterpart had one major anomaly, nor were these twin pregnancies at increased risk of preterm labour. PMID- 15282007 TI - Late pregnancy termination within a legislated medical environment. AB - AIMS: To review the indications and outcomes for abortion beyond 20 weeks' gestation within an environment of legislated notifiable pregnancy termination. METHODS: In Western Australia legislation allowing abortion > or = 20 weeks' gestation for serious maternal-fetal conditions was enacted in May 1998. Late abortions are only permitted in a single state institution and are notifiable by law. All pregnancy terminations > or = 20 weeks' gestation performed since this legislation were prospectively identified with the indications and outcomes reviewed. RESULTS: During the study period, 219 women underwent abortion > or = 20 weeks' gestation, representing 0.5% of all abortions in the state. Comparison with 438 contemporanous medical abortions for fetal anomaly at 14-20 weeks' gestations was made. Misoprostol was the primary abortifacient for both. The median maternal age for termination at 14-20 weeks was 32 years (interquartile range (IQR) 27, 36) and 30 years (IQR 26, 34) at > or = 20 weeks' gestation (P < 0.001). There was no significant difference in maternal gravidity or parity. The principal indications for terminations > or = 20 weeks were: karyotypic (28.8%); cardiac anomalies (15.5%) and neural tube defects (11.9%). Cardiac anomalies represented 5.0% of fetal anomaly terminations at 14-20 weeks (P < 0.01). The median time for medical abortion was 15.4 h (IQR 11.5, 23.2) at 14-20 weeks' gestation compared with 18.3 h (IQR 13.3, 26.1) at gestations greater than 20 weeks (P < 0.001). A total of 13.2% of terminations were performed at gestations beyond 24 weeks. CONCLUSIONS: Abortion > or = 20 weeks' gestation under medically regulated legislation is used primarily for serious fetal anomalies. The women are younger and the abortion duration is greater for late pregnancy termination compared with those conducted at earlier gestations. The majority of late terminations occur < 23 weeks' gestation and the incidence has remained stable since the legislation was enacted. PMID- 15282008 TI - Factors identified during the neonatal period associated with risk of cerebral palsy. AB - OBJECTIVE: To identify factors during the neonatal period that are associated with the subsequent development of cerebral palsy (CP). DESIGN: Case-control study. SETTING: Ten hospitals in Victoria, Australia. SAMPLE: Cases were babies with moderate or severe CP identified from the Victorian Cerebral Palsy Register. Controls were matched with cases for year of birth, plurality, sex, birthweight, gestation and hospital of birth. METHODS: A range of neonatal variables was compared between cases and controls, initially in a univariate analysis and subsequently in a logistic regression. The analysis was matched where possible. Where missing data prevented a matched analysis, an adjusted unmatched analysis was performed. MAIN OUTCOME MEASURES: Those neonatal factors making an independent contribution to the risk of CP in both term and preterm infants. RESULTS: Among babies born at term, 73% of cases and 2% of controls were identified by at least one of the following factors: seizures, congenital abnormalities of the brain and elsewhere, 'other lesions', abnormal muscle tone and meconium aspiration. Among babies born preterm, 68% of cases and 26% of controls were identified by the following factors: seizures, intraventricular haemorrhage, periventricular leukomalacia, 'other lesions' and abnormal muscle tone. CONCLUSIONS: The neonatal factors which best identify neonates who will subsequently develop CP are different for term and preterm babies. Babies born at term are identified more efficiently than those born preterm. Among term babies especially, some of these factors are clearly of a long-standing nature and are not associated with delivery. PMID- 15282010 TI - Safer laparoscopic trocar entry: it's all about pressure. AB - This prospective observational study aimed to assess the feasibility of adapting peritoneal hyperdistention to 25 mmHg during laparoscopy in an Australian hospital environment. A total of 1150 consecutive diagnostic or operative laparoscopies were performed. All cases were monitored for early detection of untoward physiological changes. All patients had Veress needle insufflation with distension to 25 mmHg prior to insertion of the primary trocar. No patients experienced any surgical entry complications or adverse clinical effects noted during anaesthetic. The aim of the current study is to assess the feasibility and safety of increasing the peritoneal insufflation pressure to 25 mmHg for primary trocar insertion. PMID- 15282009 TI - First Australian trial of the birth-training device Epi-No: a highly significantly increased chance of an intact perineum. AB - BACKGROUND: A German report suggested significantly better outcomes in terms of perineal care, second stage length and neonatal outcome for users of Epi-No. OBJECTIVE: To carry out a pilot study of the first use of the Epi-No birth training device in Australia for women having their first baby. STUDY POPULATION AND METHODS: Forty-eight primigravidae having their confinement at Birralee Birthing Unit who used the device compared to all other primigravida who delivered during the same period. RESULTS: The study shows a highly significantly improved outcome for the perineum when users are compared to primigravid non-user controls. We could not demonstrate decreased instrumental delivery rates nor a better outcome in term of Apgar scores. DISCUSSION: The Epi-No device should be offered as an option to all primigravidae to use during the late third trimester. PMID- 15282011 TI - Post-mortem Caesarean section performed 30 minutes after maternal cardiopulmonary arrest. PMID- 15282012 TI - Perimortem Caesarean section: a case report. PMID- 15282013 TI - Prevention of recurrent fetal death in utero due to group B streptococcal chorioamnionitis. PMID- 15282014 TI - Benign endocervical cystic glandular change mimicking minimal deviation adenocarcinoma of the cervix on magnetic resonance imaging: a case report. PMID- 15282015 TI - Severe pre-eclampsia complicated by multiple serous effusions. PMID- 15282016 TI - Successful management of chromophobe type renal cell carcinoma in pregnancy. PMID- 15282018 TI - Aggressive angiomyxoma: an important differential diagnosis for a vaginal mass. PMID- 15282017 TI - Ovarian endometrioid carcinoma with yolk sac component in a young patient: a diagnostic and therapeutic dilemma. PMID- 15282019 TI - Successful antenatal treatment of an affected congenital adrenal hyperplasia pregnancy using linkage analysis. PMID- 15282020 TI - Incomplete bladder emptying due to labial fusion in a pubertal girl: a delayed consequence of female circumcision. PMID- 15282021 TI - Surgical anatomy of the posterior division of the internal iliac artery: the important point for internal iliac artery ligation to control pelvic haemorrhage. PMID- 15282022 TI - Re: 'Ultrasound assessment of bladder volume: is it valid after delivery?'. PMID- 15282023 TI - Hepatitis C and pregnancy. PMID- 15282025 TI - Public - private 'partnerships' in health - a global call to action. AB - The need for public-private partnerships arose against the backdrop of inadequacies on the part of the public sector to provide public good on their own, in an efficient and effective manner, owing to lack of resources and management issues. These considerations led to the evolution of a range of interface arrangements that brought together organizations with the mandate to offer public good on one hand, and those that could facilitate this goal though the provision of resources, technical expertise or outreach, on the other. The former category includes of governments and intergovernmental agencies and the latter, the non-profit and for-profit private sector. Though such partnerships create a powerful mechanism for addressing difficult problems by leveraging on the strengths of different partners, they also package complex ethical and process-related challenges. The complex transnational nature of some of these partnership arrangements necessitates that they be guided by a set of global principles and norms. Participation of international agencies warrants that they be set within a comprehensive policy and operational framework within the organizational mandate and involvement of countries requires legislative authorization, within the framework of which, procedural and process related guidelines need to be developed. This paper outlines key ethical and procedural issues inherent to different types of public-private arrangements and issues a Global Call to Action. PMID- 15282026 TI - Small B cell lymphocytic lymphoma presenting as obstructive sleep apnea. AB - BACKGROUND: Most lymphomas that involve the tonsil are large B cell lymphomas. Large B-cell lymphoma is a high grade malignancy which progresses rapidly. Tonsillar lymphoma usually presents as either a unilaterally enlarged palatine tonsil or as an ulcerative and fungating lesion over the tonsillar area. Small lymphocytic lymphomas (SLL) of the Waldeyer's ring are uncommon. CASE PRESENTATION: We report a 41-year-old male who presented with a ten-year history of snoring. Physical examination revealed smooth bilateral symmetrically enlarged tonsils without abnormal surface change or cervical lymphadenopathy. Palatal redundancy and a narrowed oropharyngeal airway were also noted. The respiratory disturbance index (RDI) was 66 per hour, and severe obstruction sleep apnea (OSA) was suspected. No B symptoms, sore throat, odynophagia or dysphagia was found. We performed uvulopalatopharyngoplasty (UPPP) and pathological examination revealed incidental small B-cell lymphocytic lymphoma (SLL). CONCLUSION: It is uncommon for lymphoma to initially present as OSA. SLL is an indolent malignancy and is not easy to detect in the early stage. We conclude that SLL may be a contributing factor of OSA in the present case. PMID- 15282027 TI - Web GIS in practice: an interactive geographical interface to English Primary Care Trust performance ratings for 2003 and 2004. AB - BACKGROUND: On 21 July 2004, the Healthcare Commission http://www.healthcarecommission.org.uk/ released its annual star ratings of the performance of NHS Primary Care Trusts (PCTs) in England for the year ending March 2004. The Healthcare Commission started work on 1 April 2004, taking over all the functions of the former Commission for Health Improvement http://www.chi.nhs.uk/, which had released the corresponding PCT ratings for 2002/2003 in July 2003. RESULTS: We produced two Web-based interactive maps of PCT star ratings, one for 2003 and the other for 2004 http://healthcybermap.org/PCT/ratings/, with handy functions like map search (by PCT name or part of it). The maps feature a colour-blind friendly quadri-colour scheme to represent PCT star ratings. Clicking a PCT on any of the maps will display the detailed performance report of that PCT for the corresponding year. CONCLUSION: Using our Web-based interactive maps, users can visually appreciate at a glance the distribution of PCT performance across England. They can visually compare the performance of different PCTs in the same year and also between 2003 and 2004 (by switching between the synchronised 'PCT Ratings 2003' and 'PCT Ratings 2004' themes). The performance of many PCTs has improved in 2004, whereas some PCTs achieved lower ratings in 2004 compared to 2003. Web-based interactive geographical interfaces offer an intuitive way of indexing, accessing, mining, and understanding large healthcare information sets describing geographically differentiated phenomena. By acting as an enhanced alternative or supplement to purely textual online interfaces, interactive Web maps can further empower organisations and decision makers. PMID- 15282028 TI - "A calorie is a calorie" violates the second law of thermodynamics. AB - The principle of "a calorie is a calorie," that weight change in hypocaloric diets is independent of macronutrient composition, is widely held in the popular and technical literature, and is frequently justified by appeal to the laws of thermodynamics. We review here some aspects of thermodynamics that bear on weight loss and the effect of macronutrient composition. The focus is the so-called metabolic advantage in low-carbohydrate diets--greater weight loss compared to isocaloric diets of different composition. Two laws of thermodynamics are relevant to the systems considered in nutrition and, whereas the first law is a conservation (of energy) law, the second is a dissipation law: something (negative entropy) is lost and therefore balance is not to be expected in diet interventions. Here, we propose that a misunderstanding of the second law accounts for the controversy about the role of macronutrient effect on weight loss and we review some aspects of elementary thermodynamics. We use data in the literature to show that thermogenesis is sufficient to predict metabolic advantage. Whereas homeostasis ensures balance under many conditions, as a general principle, "a calorie is a calorie" violates the second law of thermodynamics. PMID- 15282029 TI - Care-seeking patterns for fatal malaria in Tanzania. AB - BACKGROUND: Once malaria occurs, deaths can be prevented by prompt treatment with relatively affordable and efficacious drugs. Yet this goal is elusive in Africa. The paradox of a continuing but easily preventable cause of high mortality raises important questions for policy makers concerning care-seeking and access to health systems. Although patterns of care-seeking during uncomplicated malaria episodes are well known, studies in cases of fatal malaria are rare. Care-seeking behaviours may differ between these groups. METHODS: This study documents care seeking events in 320 children less than five years of age with fatal malaria seen between 1999 and 2001 during over 240,000 person-years of follow-up in a stable perennial malaria transmission setting in southern Tanzania. Accounts of care-seeking recorded in verbal autopsy histories were analysed to determine providers attended and the sequence of choices made as the patients' condition deteriorated. RESULTS: As first resort to care, 78.7% of malaria-attributable deaths used modern biomedical care in the form of antimalarial pharmaceuticals from shops or government or non-governmental heath facilities, 9.4% used initial traditional care at home or from traditional practitioners and 11.9% sought no care of any kind. There were no differences in patterns of choice by sex of the child, sex of the head of the household, socioeconomic status of the household or presence or absence of convulsions. In malaria deaths of all ages who sought care more than once, modern care was included in the first or second resort to care in 90.0% and 99.4% with and without convulsions respectively. CONCLUSIONS: In this study of fatal malaria in southern Tanzania, biomedical care is the preferred choice of an overwhelming majority of suspected malaria cases, even those complicated by convulsions. Traditional care is no longer a significant delaying factor. To reduce mortality further will require greater emphasis on recognizing danger signs at home, prompter care-seeking, improved quality of care at health facilities and better adherence to treatment. PMID- 15282030 TI - Malaria morbidity and immunity among residents of villages with different Plasmodium falciparum transmission intensity in North-Eastern Tanzania. AB - BACKGROUND: The relationship between the burden of uncomplicated malaria and transmission intensity is unclear and a better understanding of this relationship is important for the implementation of intervention programmes. METHODS: A 6 month longitudinal study monitoring risk factors for anaemia and febrile malaria episodes was conducted among individuals aged below 20 years, residing in three villages of different altitude in areas of high, moderate and low malaria transmission intensity in North-Eastern Tanzania. RESULTS: The burden of anaemia and malarial fever fell mainly on the youngest children and was highest in the village with high transmission intensity. Although a considerable percentage of individuals in all villages carried intestinal worms, logistic regression models indicated that Plasmodium falciparum was the only significant parasitic determinant of anaemia. Interestingly, children who carried low-density parasitaemia at the start of the study had a lower risk of contracting a febrile malaria episode but a higher risk of anaemia during the study period, than children who were slide negative at this point in time. CONCLUSION: Young children living in the high transmission village carried a very high anaemia burden, which could be attributed to malaria. The overall incidence of febrile malaria was also highest in the high transmission village particularly among those under five years of age. These data suggest that in rolling back malaria, available resources in prevention programmes should primarily be focussed on young children, particularly those residing in areas of high malaria transmission. PMID- 15282031 TI - Characterisation of the Escherichia coli membrane structure and function during fedbatch cultivation. AB - BACKGROUND: Important parameters during recombinant protein production in Escherichia coli, such as productivity and protein activity, are affected by the growth rate. This includes the translocation of protein over the membrane to gain better folding capacity or reduced proteolysis. To vary the growth rate two techniques are available: fedbatch and continuous cultivation, both controlled by the ingoing feed rate. RESULTS: During fedbatch cultivation, E. coli contains phosphatidylethanolamine, phosphatidylglycerol, cardiolipin and saturated fatty acids in amounts which are stable with growth rate. However, the levels of cardiolipin are very high compared to continuous cultivation. The reason for fedbatch triggering of this metabolism is not known but hypothesised to result from an additional need for carbon and energy. The reason could be the dynamic and sometimes rapid changes in growth rate to which the fedbatch cell has at all times to adjust. The membrane flexibility, essential for translocation of various components, is however to some degree sustained by production of increased amounts of unsaturated fatty acids in phosphatidylglycerol. The result is a functionally stiff membrane which generally promotes low cell lysis and is constant with respect to protein leakage to the medium. At comparatively high growth rates, when the further stabilising effect of cyclic fatty acids is gone, the high level of unsaturated fatty acids results in a pronounced effect upon sonication. This is very much in contrast to the membrane function in continuous cultivation which shows very specific characteristics as a function of growth rate. CONCLUSIONS: The stiff and unchanging fedbatch membrane should promote a stable behaviour during downstream processing and is less dependent on the time of harvest. However, optimisation of protein leakage can only be achieved in the continuously cultivated cell where leakage is twice as high compared to the constant leakage level in fedbatch. If leakage is undesired, continuous cultivation is also preferred since it can be designed to lead to the lowest values detected. Induction at low growth rate (<0.2 h-1) should be avoided with respect to productivity, in any system, since the specific and total protein production shows their lowest values at this point. PMID- 15282032 TI - Shoulder posture and median nerve sliding. AB - BACKGROUND: Patients with upper limb pain often have a slumped sitting position and poor shoulder posture. Pain could be due to poor posture causing mechanical changes (stretch; local pressure) that in turn affect the function of major limb nerves (e.g. median nerve). This study examines (1) whether the individual components of slumped sitting (forward head position, trunk flexion and shoulder protraction) cause median nerve stretch and (2) whether shoulder protraction restricts normal nerve movements. METHODS: Longitudinal nerve movement was measured using frame-by-frame cross-correlation analysis from high frequency ultrasound images during individual components of slumped sitting. The effects of protraction on nerve movement through the shoulder region were investigated by examining nerve movement in the arm in response to contralateral neck side flexion. RESULTS: Neither moving the head forward or trunk flexion caused significant movement of the median nerve. In contrast, 4.3 mm of movement, adding 0.7% strain, occurred in the forearm during shoulder protraction. A delay in movement at the start of protraction and straightening of the nerve trunk provided evidence of unloading with the shoulder flexed and elbow extended and the scapulothoracic joint in neutral. There was a 60% reduction in nerve movement in the arm during contralateral neck side flexion when the shoulder was protracted compared to scapulothoracic neutral. CONCLUSION: Slumped sitting is unlikely to increase nerve strain sufficient to cause changes to nerve function. However, shoulder protraction may place the median nerve at risk of injury, since nerve movement is reduced through the shoulder region when the shoulder is protracted and other joints are moved. Both altered nerve dynamics in response to moving other joints and local changes to blood supply may adversely affect nerve function and increase the risk of developing upper quadrant pain. PMID- 15282033 TI - Discovery of induced point mutations in maize genes by TILLING. AB - BACKGROUND: Going from a gene sequence to its function in the context of a whole organism requires a strategy for targeting mutations, referred to as reverse genetics. Reverse genetics is highly desirable in the modern genomics era; however, the most powerful methods are generally restricted to a few model organisms. Previously, we introduced a reverse-genetic strategy with the potential for general applicability to organisms that lack well-developed genetic tools. Our TILLING (Targeting Induced Local Lesions IN Genomes) method uses chemical mutagenesis followed by screening for single-base changes to discover induced mutations that alter protein function. TILLING was shown to be an effective reverse genetic strategy by the establishment of a high-throughput TILLING facility and the delivery of thousands of point mutations in hundreds of Arabidopsis genes to members of the plant biology community. RESULTS: We demonstrate that high-throughput TILLING is applicable to maize, an important crop plant with a large genome but with limited reverse-genetic resources currently available. We screened pools of DNA samples for mutations in 1-kb segments from 11 different genes, obtaining 17 independent induced mutations from a population of 750 pollen-mutagenized maize plants. One of the genes targeted was the DMT102 chromomethylase gene, for which we obtained an allelic series of three missense mutations that are predicted to be strongly deleterious. CONCLUSIONS: Our findings indicate that TILLING is a broadly applicable and efficient reverse-genetic strategy. We are establishing a public TILLING service for maize modeled on the existing Arabidopsis TILLING Project. PMID- 15282037 TI - Raman spectroscopy in the deep ocean: successes and challenges. PMID- 15282034 TI - Biochemical enrichment and biophysical characterization of a taste receptor for L arginine from the catfish, Ictalurus puntatus. AB - BACKGROUND: The channel catfish, Ictalurus punctatus, is invested with a high density of cutaneous taste receptors, particularly on the barbel appendages. Many of these receptors are sensitive to selected amino acids, one of these being a receptor for L-arginine (L-Arg). Previous neurophysiological and biophysical studies suggested that this taste receptor is coupled directly to a cation channel and behaves as a ligand-gated ion channel receptor (LGICR). Earlier studies demonstrated that two lectins, Ricinus communis agglutinin I (RCA-I) and Phaseolus vulgaris Erythroagglutinin (PHA-E), inhibited the binding of L-Arg to its presumed receptor sites, and that PHA-E inhibited the L-Arg-stimulated ion conductance of barbel membranes reconstituted into lipid bilayers. RESULTS: Both PHA-E and RCA-I almost exclusively labeled an 82-84 kDa protein band of an SDS PAGE of solubilized barbel taste epithelial membranes. Further, both rhodamine conjugated RCA-I and polyclonal antibodies raised to the 82-84 kDa electroeluted peptides labeled the apical region of catfish taste buds. Because of the specificity shown by RCA-I, lectin affinity was chosen as the first of a three step procedure designed to enrich the presumed LGICR for L-Arg. Purified and CHAPS-solubilized taste epithelial membrane proteins were subjected successively to (1), lectin (RCA-I) affinity; (2), gel filtration (Sephacryl S-300HR); and (3), ion exchange chromatography. All fractions from each chromatography step were evaluated for L-Arg-induced ion channel activity by reconstituting each fraction into a lipid bilayer. Active fractions demonstrated L-Arg-induced channel activity that was inhibited by D-arginine (D-Arg) with kinetics nearly identical to those reported earlier for L-Arg-stimulated ion channels of native barbel membranes reconstituted into lipid bilayers. After the final enrichment step, SDS-PAGE of the active ion channel protein fraction revealed a single band at 82-84 kDa which may be interpreted as a component of a multimeric receptor/channel complex. CONCLUSIONS: The data are consistent with the supposition that the L-Arg receptor is a LGICR. This taste receptor remains active during biochemical enrichment procedures. This is the first report of enrichment of an active LGICR from the taste system of vertebrata. PMID- 15282035 TI - Calcineurin activation influences muscle phenotype in a muscle-specific fashion. AB - BACKGROUND: The calcium activated protein phosphatase 2B, also known as calcineurin, has been implicated as a cell signaling molecule involved with transduction of physiological signals (free cytosolic Ca2+) into molecular signals that influence the expression of phenotype-specific genes in skeletal muscle. In the present study we address the role of calcineurin in mediating adaptations in myosin heavy chain (MHC) isoform expression and muscle mass using 3-month old wild-type (WT) and transgenic mice displaying high-level expression of a constitutively active form of calcineurin (MCK-CN* mice). RESULTS: Slow muscles, e.g., soleus, were significantly larger (by ~24%), whereas fast muscles, e.g., medial gastrocnemius (MG) and tibialis anterior were significantly smaller (by ~26 and ~16%, respectively) in MCK-CN* mice compared to WT. The masses of mixed phenotype muscles, such as the plantaris and the extensor digitorum longus, were not significantly changed from WT. The soleus, plantaris, MG and diaphragm displayed shifts toward slower MHC isoforms, e.g., soleus from WT mice contained ~52% MHC-I, ~39% MHC-IIa, and ~9% MHC-IIx, whereas MCK-CN* mice had ~67% MHC-I, ~26% MHC-IIa, and ~7% MHC-IIx. The specific isoforms that were either up or down regulated were muscle-specific. For instance, the proportion of MHC-IIa was decreased in the soleus and diaphragm, but increased in the plantaris and MG of MCK-CN* mice. Also, the proportion of MHC-IIx was unchanged in the soleus, decreased in the diaphragm and increased in the plantaris and MG of MCK-CN* relative to WT mice. Fast to slow shifts in fiber type proportions were evident for the plantaris, but not the soleus. Fast, but not slow, plantaris fibers of MCK-CN* mice had higher oxidative and lower glycolytic properties than WT. CONCLUSION: These data suggest that calcineurin activation can influence muscle phenotype and that the specific influence of calcineurin activation on the phenotypic and mass characteristics of a muscle is dependent upon the original phenotypic state of the muscle. PMID- 15282038 TI - Apertureless tip-enhanced Raman microscopy with confocal epi illumination/collection optics. AB - It is demonstrated that confocal epi-illumination/collection optics can be effectively used to generate surface-enhanced Raman scattering at the near-field region of a gold-coated tip for an atomic force microscope operated in semi contact tapping mode. When the tip, with a 50-nm apex radius, was illuminated by a highly focused laser beam at 532 nm and approached the isolated diamond particle, with a size of approximately 1 microm, the Raman signal was enhanced by approximately 10(3). This result is in good agreement with numerical simulations performed by the finite difference time-domain method. Since our apertureless microscope is based on readily available conventional components, there is wide room for improvements and modifications by common users in various applications of micro-Raman analysis. PMID- 15282039 TI - Microchip laser-induced breakdown spectroscopy: a preliminary feasibility investigation. AB - A commercial, 7 microJ/pulse, 550 ps microchip laser is used to induce plasma on Pb, Si, Cu, Fe, Ni, Ti, Zn, Ta, and Mo foils and a Si wafer. The measured plasma lifetime is comparable with the duration of the laser pulse (a few ns). The plasma continuum radiation is low, while some of the strong resonance lines (e.g., Zn 213.86 nm) show self-reversal. Quantitative analysis is possible using non-gated detectors but analytical lines should be chosen with care to avoid reduction in the linear dynamic range. The mass removed (0.5-20 ng/pulse) is sufficient to yield spectra that are detectable with portable grating spectrometers equipped with non-gated, non-intensified detector arrays. The spectrum of Cd is detected with a broadband portable spectrometer (200-950 nm). The combination of the broadband spectrometer and the microchip laser is very promising for material identification, especially in field applications. PMID- 15282040 TI - Determination of nitrogen in sand using laser-induced breakdown spectroscopy. AB - The use of laser-induced breakdown spectroscopy (LIBS) to detect a variety of elements in soils has been demonstrated and instruments have been developed to facilitate these measurements. The ability to determine nitrogen in soil is also important for applications ranging from precision farming to space exploration. For terrestrial use, the ideal situation is for measurements to be conducted in the ambient air, thereby simplifying equipment requirements and speeding the analysis. The high concentration of nitrogen in air, however, is a complicating factor for soil nitrogen measurements. Here we present the results of a study of LIBS detection of nitrogen in sand at atmospheric and reduced pressures to evaluate the method for future applications. Results presented include a survey of the nitrogen spectrum to determine strong N emission lines and determination of measurement precision and a detection limit for N in sand (0.8% by weight). Our findings are significantly different from those of a similar study recently published regarding the detection of nitrogen in soil. PMID- 15282041 TI - Controlled dissolution for elemental analysis of sample layers by inductively coupled plasma-mass spectrometry: a feasibility study. AB - Aqueous acid mixtures at room temperature are used to partially dissolve steel samples. The dissolved elements are washed off the surface, diluted, and then determined by inductively coupled plasma-mass spectrometry (ICP-MS) using a magnetic sector mass analyzer. The amount of material removed is measured from the amount of Fe dissolved and increases linearly with HNO(3) concentration in the etch acid. Analyte concentrations in the solid are determined from the signal ratio of analyte ion/Fe(+). The shape of a plot of mass of element removed vs. nitric acid concentration yields information about the efficiency of the removal process and the likely chemical form of the element in the sample. For elements like Mn, Al, and W in steel, these plots have the same linear shape as that for the major element (Fe), and the measured concentrations agree well with the certified values. For problem elements like Nb and Ta, the plots have two linear regions with different slopes, and measured concentrations are lower than the certified values. Laser ablation ICP-MS and scanning electron microscopy (SEM) measurements show these elements to be associated together in the solid in refractory grains that are not dissolved to the same extent as the Fe matrix. For steel, the amount of Fe dissolved corresponds to an average depth of at least 4 microm, or 20 000 atomic layers. PMID- 15282042 TI - Measurements of cavity ringdown spectroscopy of acetone in the ultraviolet and near-infrared spectral regions: potential for development of a breath analyzer. AB - We report a study on the cavity ringdown spectroscopy of acetone in both the ultraviolet (UV) and the near-infrared (NIR) spectral regions to explore the potential for development of a breath analyzer for disease diagnostics. The ringdown spectrum of acetone in the UV (282.4-285.0 nm) region is recorded and the spectrum is in good agreement with those obtained by other spectral techniques reported in the literature. The absorption cross-section of the C-H stretching overtone of acetone in the NIR (1632.7-1672.2 nm) is reported for the first time and the maximum absorption cross-section located at 1666.7 nm is 1.2 x 10(-21) cm(2). A novel, compact, atmospheric cavity with a cavity length of 10 cm has been constructed and implemented to investigate the technical feasibility of the potential instrument size, optical configuration, and detection sensitivity. The detection limit of such a mini cavity employing ringdown mirrors of reflectivity of 99.85% at 266 nm, where acetone has the strongest absorption, is approximately 1.5 ppmv based on the standard 3 criteria. No real breath gas samples are used in the present study. Discussions on the detection sensitivity and background spectral interferences for the instrument development are presented. This study demonstrates the potential of developing a portable, sensitive breath analyzer for medical applications using the cavity ringdown spectral technique. PMID- 15282043 TI - Diode laser based photoacoustic water vapor detection system for atmospheric research. AB - A wavelength modulated, distributed feedback diode laser based photoacoustic water vapor mixing ratio measuring system for atmospheric research applications is presented. Laser modulation parameters were optimized either at 180 or 500 mbar total pressure to enhance the system's sensitivity for low or high pressures (upper troposphere/lower stratosphere or biosphere exchange layer), respectively. A wavelength locking method was developed that ensured sub-picometer absolute (5 x 10(-7) relative) wavelength stability of the laser while consuming minimum additional measurement time. At the calibration of the system, correction factors for the pressure- and temperature-dependence of the photoacoustic signal were determined, which were in turn applied to the calculation of the water vapor mixing ratio from the measured signal during the test operation of the system. The introduced features resulted in reliable, sub-ppm-level water vapor detection even under abrupt gas pressure or temperature variations typical in open atmospheric applications. PMID- 15282044 TI - A faster approach to infrared rheo-optics using a planar array infrared spectrograph. AB - Infrared rheo-optics combines dynamic mechanical analysis with infrared spectroscopy to provide molecular level information about the segmental reorientation and the changes in local environment associated with the dynamic deformation of polymers. Up to now, the application of this technique has been limited by the amount of time necessary to perform the experiments. In this article, we demonstrate that the use of a planar array infrared (PA-IR) spectrograph can accelerate the acquisition time by as much as two orders of magnitude while maintaining a signal-to-noise ratio (SNR) similar to that obtained using step-scan Fourier transform infrared (FT-IR) spectrometry, and by more than three orders of magnitude at the expense of a reduced SNR. The advantages and drawbacks of this new technique are discussed. PMID- 15282045 TI - On-line fermentation monitoring by mid-infrared spectroscopy. AB - A new method for on-line monitoring of fermentations using mid-infrared (MIR) spectroscopy has been developed. The method has been used to predict the concentrations of glucose and ethanol during a baker's yeast fermentations. A completely automated flow system was employed as an interface between the bioprocess under study and the Fourier transform infrared (FT-IR) spectrometer, which was equipped with a flow cell housing a diamond attenuated total reflection (ATR) element. By using the automated flow system, experimental problems related to adherence of CO(2) bubbles to the ATR surface, as well as formation of biofilms on the ATR surface, could be efficiently eliminated. Gas bubbles were removed during sampling, and by using rinsing steps any biofilm could be removed from the ATR surface. In this way, constant measuring conditions could be guaranteed throughout prolonged fermentation times (approximately 8 h). As a reference method, high-performance liquid chromatography (HPLC) with refractive index detection was used. The recorded data from different fermentations were modeled by partial least-squares (PLS) regression comparing two different strategies for the calibration. On the one hand, calibration sets were constructed from spectra recorded from either synthetic standards or from samples drawn during fermentation. On the other hand, spectra from fermentation samples and synthetic standards were combined to form a calibration set. Differences in the kinetics of the studied fermentation processes used for calibration and prediction, as well as the precision of the HPLC reference method, were identified as the main chemometric sources of error. The optimal PLS regression method was obtained using the mixed calibration set of samples from fermentations and synthetic standards. The root mean square errors of prediction in this case were 0.267 and 0.336 g/L for glucose and ethanol concentration, respectively. PMID- 15282046 TI - Use of infrared spectroscopy for the determination of electronegativity of rare earth elements. AB - Infrared spectroscopy has been used to study a series of synthetic agardite minerals. Four OH stretching bands are observed at around 3568, 3482, 3362, and 3296 cm(-1). The first band is assigned to zeolitic, non-hydrogen-bonded water. The band at 3296 cm(-1) is assigned to strongly hydrogen-bonded water with an H bond distance of 2.72 A. The water in agardites is better described as structured water and not as zeolitic water. Two bands at around 999 and 975 cm(-1) are assigned to OH deformation modes. Two sets of AsO symmetric stretching vibrations were found and assigned to the vibrational modes of AsO(4) and HAsO(4) units. Linear relationships between positions of infrared bands associated with bonding to the OH units and the electronegativity of the rare earth elements were derived, with correlation coefficients >0.92. These linear functions were then used to calculate the electronegativity of Eu, for which a value of 1.1808 on the Pauling scale was found. PMID- 15282047 TI - Study of the interactions between sucrose and metal ions (Mg2+ and K+) and their simultaneous quantification in ternary mixture by mid-infrared and 13C nuclear magnetic resonance spectroscopies. AB - In this paper, a ternary aqueous mixture of sucrose and two metal ions (Mg(2+) and K(+)) has been examined by mid-infrared spectroscopy coupled with principal component analysis (PCA) and the partial least-squares regression method (PLS). PCA was first used for the description of Fourier transform mid-infrared (mid FTIR) spectral data of the complex samples. The resulting factorial map, set up with the two most influential component axes, features distinct concentration distribution specific to each component. Prediction equations that linked sucrose, magnesium, and potassium concentrations to the spectral data were established by the partial least-squares regression method. A quite good correlation was obtained between the first 5 axes and the concentration variables, with coefficient values ranging from 0.984 to 0.997. It was thus possible to predict specifically both metal ion concentrations in the ternary mixture with relatively good accuracy. The ternary mixtures of sucrose, Mg(2+), and K(+) were also subjected to (13)C NMR (nuclear magnetic resonance) analysis. From the relative displacements of chemical shifts of the carbon atoms of sucrose, it was possible to determine the influence of each metal ion present in the mixture. PMID- 15282048 TI - Thermodynamics of complexation of dimethyl esters of tere-, iso-, and phthalic acids with alpha- and beta-cyclodextrins. AB - Thermodynamics of inclusion of alpha- and beta-cyclodextrins (CDs) with three diester precursors of polyesters, dimethyl terephthalate (DMT), dimethyl isophthalate (DMI), and dimethyl phthalate (DMP) were studied with different fluorescence spectroscopy techniques. The stoichiometry, association constants, and other thermodynamics parameters were investigated from the fluorescence intensity decrease that takes place upon CD addition and temperature changes. Apart from DMP, which did not form a complex with alphaCD, all other systems showed 1:1 stoichiometry complexes with different stabilities. The complexes formed with the DMT guest are the most stable, whereas the stability constant for DMP:betaCD is the lowest. Binding constants are also larger for complexes formed with betaCD than for those formed with alphaCD. Fluorescence polarization, quenching, and lifetime measurements provide information about the structure of the complexes formed. This geometry explains the values of enthalpy and entropy changes during complexation. Van der Waals interactions seem to be involved in the formation of these complexes. PMID- 15282049 TI - Detection of G-quartet structure in a DNA aptamer stationary phase using a fluorescent dye. AB - The fluorescent porphyrin dye N-methylmesoporphyrin IX (NMM) was used to provide direct evidence of intramolecular G-quartet formation by an oligonucleotide immobilized at the inner surface of a fused silica capillary. The oligonucleotide is the thrombin-binding DNA aptamer, which has been used in several analytical applications, including a stationary phase for open tubular capillary electrochromatography. Spectroscopic studies of the dye in batch solutions of the aptamer and of an oligonucleotide with the same base composition, but in a different, "scrambled" sequence that does not form an intramolecular G-quartet, provided evidence of selective fluorescence enhancement of NMM by the aptamer in the intramolecular G-quartet structure. On-column experiments compared results for injections of NMM onto an aptamer-coated capillary, a capillary coated with the scrambled sequence oligonucleotide, and a bare fused silica capillary. Results show that while NMM adsorbs to both coated capillaries, the selective fluorescence enhancement provides evidence of the intramolecular G-quartet structure on the aptamer-coated capillary. PMID- 15282050 TI - Phosphorescence properties of p-aminobenzoic acid immobilized on a nylon membrane. AB - The properties of nylon as a new material for obtaining room-temperature phosphorescence from p-aminobenzoic acid (PABA) have been investigated. Although PABA shows native phosphorescence when adsorbed on a nylon membrane, a significant improvement in both sensitivity and limit of detection is achieved in the presence of alpha-cyclodextrin and heavy atoms. An additional enhancement of the phosphorescence signal is attained when the measurements are carried out under a nitrogen atmosphere. The analytical figures of merit obtained under the best experimental conditions are: linear calibration range from 0.6 to 6 ng/spot (the lowest value corresponds to the quantitation limit); correlation coefficient, 0.998 for 18 data points; relative standard deviation, 2.2% (n = 5) at a level of 2.4 ng/spot; and limit of detection, 0.2 ng/spot (calculated according to Clayton's definition). PMID- 15282051 TI - Solvatochromism of Nile Red in nonpolar solvents. AB - The absorbance and fluorescence spectra of Nile Red (NR) were examined in a series of nonpolar solvents comprising linear alkanes and a range of poly alpha olefins (PAO). These solvents span a 1000-fold range in viscosity and possess very similar dielectric constants and refractive index properties. A high-energy double peak with vibronic structure is observed in both fluorescence and absorbance spectra, possibly indicating that a locally excited (LE) state is accessed in these solvents. In addition, a red shift in peak position is observed with increasing refractive index; however, it is unaccompanied by any changes in Stokes shift. This shift is attributed to changes in the high-frequency polarizability of the solvent, which is a function of the refractive index. Finally, an increase in quantum yield with viscosity is also observed. PMID- 15282052 TI - Fuzzy logic for identifying pigments studied by Raman spectroscopy. AB - Fuzzy logic and linguistic variables are used for the automatic interpretation of Raman spectra obtained from pigments found in cultural heritage art objects. Featured bands are extracted from a Raman spectrum of a reference pigment and the methodology for constructing the library is illustrated. An unknown spectrum is then interpreted automatically and a process for identifying the corresponding pigment is described. A reference library consisting of 32 pigments was built and the effectiveness of the algorithm was tested by the Raman spectroscopic analysis of 10 pigments that are known to have been extensively used in Byzantine hagiography. Binary mixtures of these pigments were also tested. The algorithm's level of identification was good even though extra peaks, noise, and background signals were encountered in the spectra. PMID- 15282053 TI - Maximum likelihood principal components regression on wavelet-compressed data. AB - Maximum likelihood principal component regression (MLPCR) is an errors-in variables method used to accommodate measurement error information when building multivariate calibration models. A hindrance of MLPCR has been the substantial demand on computational resources sometimes made by the algorithm, especially for certain types of error structures. Operations on these large matrices are memory intensive and time consuming, especially when techniques such as cross-validation are used. This work describes the use of wavelet transforms (WT) as a data compression method for MLPCR. It is shown that the error covariance matrix in the wavelet and spectral domains are related through a two-dimensional WT. This allows the user to account for any effects of the wavelet transform on spectral and error structures. The wavelet transform can be applied to MLPCR when using either the full error covariance matrix or the smaller pooled error covariance matrix. Simulated and experimental near-infrared data sets are used to demonstrate the benefits of using wavelets with the MLPCR algorithm. In all cases, significant compression can be obtained while maintaining favorable predictive ability. Considerable time savings were also attained, with improvements ranging from a factor of 2 to a factor of 720. Using the WT compressed data in MLPCR gave a reduction in prediction errors compared to using the raw data in MLPCR. An analogous reduction in prediction errors was not always seen when using PCR. PMID- 15282054 TI - In-line reaction monitoring of a methyl methacrylate and N, N-dimethylacrylamide copolymerization reaction using near-infrared spectroscopy. AB - Fast and accurate monitoring of monomer concentration during copolymerization reactions is of much interest. It is known that near-infrared spectroscopy (NIRS) can be used to monitor polymerization reactions. Here, a free radical solution copolymerization reaction between methyl methacrylate and N,N-dimethylacrylamide is considered. NIR spectra were measured in-line with a transflectance probe. The spectra of both involved monomers are very similar, making monitoring with NIRS challenging. It is shown that the NIRS calibration can be set up with only a few (5) off-line measured mixtures. Several validation methods for such a NIRS calibration model are discussed and tested. NIRS is used to follow conversion of the two monomers in a copolymerization reaction on-line. PMID- 15282056 TI - SAS Officer Candidate Profiles. PMID- 15282055 TI - Miniature stereo spectral imaging system for multivariate optical computing. PMID- 15282057 TI - [Heart transplantation: new challenges for the XXI century]. PMID- 15282058 TI - [Transradial approach for primary angioplasty. Necessity or tightrope walking?]. PMID- 15282059 TI - [Navigation systems in current electrophysiology]. PMID- 15282060 TI - [Neurohormonal activation in congestive heart failure: does it normalize after heart transplantation?]. AB - INTRODUCTION AND OBJECTIVE: In patients with congestive heart failure, neurohormonal activation plays an important role in disease progression and prognosis. The aim of this study was to document the evolution of neurohormonal activation after heart transplantation. PATIENTS AND METHOD: Thirty-seven patients on the waiting list for heart transplantation were included in the study. Plasma levels of angiotensin II, aldosterone, endothelin, atrial natriuretic peptide and adrenomedullin were measured before heart transplantation and again 1, 4, 9 and 12 months afterwards. Plasma levels of norepinephrine and renin were measured before and 1 month after heart transplantation. RESULTS: The levels of angiotensin II, norepinephrine and renin showed a nonsignificant trend towards reduction. The levels of aldosterone were unchanged, and an increase in endothelin levels was seen 9 and 12 months after transplantation. Plasma levels of atrial natriuretic peptide and adrenomedullin were significantly lower 1, 4, 9 and 12 months after heart transplantation compared to pretransplant levels. CONCLUSIONS: During the first several months after heart transplantation there were no significant reductions in plasma levels of angiotensin II, aldosterone and endothelin, and there were significant reductions soon after surgery in peptides with a predominantly vasodilator effect (atrial natriuretic peptide and adrenomedullin). This unfavorable neurohormonal profile may contribute to the development of posttransplant complications such as edema, arterial hypertension and endothelial dysfunction. PMID- 15282061 TI - [Transradial approach for percutaneous coronary stenting in the treatment of acute myocardial infarction]. AB - INTRODUCTION AND OBJECTIVE: Treatment of acute myocardial infarction by percutaneous coronary intervention with stenting leads to excellent immediate clinical results and a good prognosis. The aim of this study was to compare in this selected population the safety and effectiveness of radial artery access versus femoral artery access. PATIENTS AND METHOD: Between May 2001 and June 2003, 162 consecutive patients with acute myocardial infarction < 12 hours treated by percutaneous stenting were included in an observational study. The radial artery approach was used in 103 patients, and the femoral artery approach in the remaining 59 patients. The success of the procedure, incidence of major adverse cardiac events and local puncture complications were compared in patients treated with the radial artery versus the femoral artery approach. RESULTS: Fluoroscopy time (22.4 [15.4] min vs 24.5 [19.5] min), immediate success of the procedure (96.1% vs 94.9%), and the incidence of major adverse cardiac events (6.8% vs 8.5%) did not differ between the two groups. Bleeding complications due to local puncture were present only in the femoral artery access group (0 vs 5 patients; P= .007). CONCLUSIONS: In selected patients with acute myocardial infarction treated with primary stent implantation, the success rate and clinical safety of the radial artery approach are similar to those of the femoral artery approach, but the incidence of local complications, especially bleeding, is significantly lower in the former. Thus the radial artery approach should become the approach of choice in patients at high risk for bleeding complications. PMID- 15282062 TI - [Ventricular tachycardia ablation guided by LocaLisa system in patients with structural heart disease]. AB - INTRODUCTION: The ablation of ventricular tachycardia is limited by a number of factors that reduce the effectiveness of this intervention in patients with structural heart disease compared to other types of arrhythmia. Recent years have seen the development of several nonfluoroscopic navigation techniques that facilitate the mapping of complex arrhythmogenic substrates. One such technique, the LocaLisa system, has not previously been tested for the ablation of ventricular tachycardia. PATIENTS AND METHOD: A total of 32 patients with structural heart disease were treated at our center with ablation for sustained ventricular tachycardia. In 10 patients the LocaLisa system was used to visualize the catheters during the procedure. We compared the results in the LocaLisa group with those in a control group of 22 patients treated with conventional fluoroscopy-guided ablation. RESULTS: The success rate of ablation was 75% (9/12 procedures) in the LocaLisa group and 68% (17/25 procedures) in the control group (P=NS). In the LocaLisa group, mean total duration of the procedure (243 +/- 84), duration of ablation (86 +/- 56) and fluoroscopy time (46 +/- 19) did not differ significantly from those in the control group (244 +/- 72 min, 79 +/- 58 min, and 43 +/- 27 min, respectively). In the LocaLisa group the trend toward greater hemodynamic intolerance in ventricular tachycardia approached significance (42% in the LocaLisa group vs 24% in the control group, P=.05) and the number of mapping procedures performed during sinus rhythm was significantly higher in the former (33% in the LocaLisa group vs 4% in the control group, P=.03). With the LocaLisa system it was possible to locate and reposition the ablation catheter accurately at the target endocardial sites, as confirmed by electrographic recordings and fluoroscopic verification. CONCLUSIONS: The LocaLisa system helps to delineate the reentry circuit and facilitates accurate catheter repositioning in patients with structural heart disease and ventricular tachycardia. PMID- 15282063 TI - [The effects of rilmenidine on cardiac autonomic function in healthy volunteers]. AB - INTRODUCTION AND OBJECTIVES: Rilmenidine is an antihypertensive drug whose antihypertensive effect occurs via a sympatholytic action on the central nervous system. However, the effects of rilmenidine on autonomic cardiovascular function are not clear. The aim of this study was to evaluate the acute effect of rilmenidine on autonomic cardiac function by measuring heart rate variability. SUBJECTS AND METHOD: A total of 20 healthy men (mean age, 263 years) were included in the study; 1 mg of rilmenidine or placebo was given to participants on different days in a double-blind crossover randomized study protocol. After drug administration, time domain and frequency domain parameters of heart rate variability were determined before and after 2 h with the patient in supine decubitus and during the handgrip exercise with 5-min electrocardiographic recordings. RESULTS: Rilmenidine caused an increase in mean RR values after administration when compared to pre-drug administration recordings with the patient in supine decubitus (929 ms vs 860 ms, P<.05), but this effect was not found in the placebo group. However, there were no differences in other time domain parameters or in any of the frequency domain parameters (normalized low frequency unit, normalized high frequency unit and low frequency/high frequency ratio) with the participant in supine position in either group. In addition, neither rilmenidine nor placebo modified heart rate variability parameters during the handgrip exercise. CONCLUSION: Administration of a single dose of rilmenidine increased vagal tone without affecting vagal modulation in the supine position. The absence of vagal tone increase during the handgrip exercise suggests that this effect of rilmenidine is minimal. PMID- 15282064 TI - [Surgical treatment for hypertrophic obstructive cardiomyopathy]. AB - INTRODUCTION AND OBJECTIVES: Five percent of the patients with hypertrophic obstructive cardiomyopathy (HOCM) have symptoms unresponsive to medical treatment and are candidates for invasive therapy. The objective of this study was to analyze our results with surgical treatment of HOCM during the last 10 years. PATIENTS AND METHOD: Between July 1993 and January 2004 26 patients with HOCM refractory to drug therapy were operated on. An extended septal myectomy was performed, in combination with anterior mitral leaflet plication in 19 cases (73%) and with mitral valve replacement in 5 (19%). Evolution of the grade of dyspnea, left ventricle outflow tract gradient (LVOTG), mitral regurgitation, and systolic anterior motion after surgery was analyzed. RESULTS: Mean follow-up was 63 (37) months. After surgery, a significant reduction in LVOTG (from 96.5 to 19.5 mmHg; P<.001), grade of mitral regurgitation (from 2.54 to 0.69; P<.001) and systolic anterior motion (from 2.92 to 0.23; P<.001) was achieved, which led to improvement in functional class. Hospital mortality and need for pacemaker implantation due to complete heart block after surgery was 3.8% (n=1). There were no cases of iatrogenic ventricular septal defect or mitro-aortic valve injury. Actuarial survival at 5 years was 96% (4%). CONCLUSIONS: Surgery in patients with HOCM yields great clinical improvements with low morbidity and mortality. Simultaneous intervention for both myocardial and valvular components of the disease allows not only reduction in the LVOTG but also correction of mitral regurgitation and abolition of systolic anterior motion. PMID- 15282065 TI - [Update in arrhythmogenic right ventricular cardiomyopathy: genetic, clinical presentation and risk stratification]. AB - Arrhythmogenic right ventricular cardiomyopathy (ARVC), or dysplasia, is a genetic heart muscle disease whose diagnosis is often a challenge for the clinician. It is one of the commonest causes of sudden cardiac death in the young. The classic description of the disease describes the end stage of a process where the myocardium, mainly of the right ventricle, has been substituted by fibrofatty tissue. Thus the early stages of the disease with subtle symptomatology are often missed. Unfortunately the risk of a fatal outcome is no less severe. The genetic basis is under investigations. Disease causing mutations in important cell adhesion genes (plakoglobin, desmoplakin) provide the basis for improved diagnosis and understanding of the pathogenesis. Animal models support the pathogenic theory that alterations on the integrity of the adhesion junction is followed by a cellular death and progressive fibrofatty replacement, the substrate for ventricular arrhythmias. Due the growing complexity and numerous phenotypic variations reported, sometimes in the same family, international registries have been created. The present review aims to summarise the current concepts on ARVC emphasising the genetic studies, the diagnosis, new diagnostic techniques and prognosis. PMID- 15282066 TI - [Risk stratification and prevention of sudden death in patients with heart failure]. AB - Patients with heart failure can die of progressive refractory heart failure or sudden cardiac death. This article reviews the major clinical predictors of sudden death in patients with heart failure due to left ventricular systolic dysfunction. Although earlier studies have identified many independent univariate predictors of reduced survival in these patients, the positive predictive value of most of them is low. Cardioverter defibrillator implantation has been shown to be the most effective therapy in patients resuscitated after cardiac arrest caused by ventricular fibrillation or poorly tolerated ventricular tachycardia. Low left ventricular ejection fraction, low New York Heart Association functional class, unsustained ventricular tachycardia and inducibility of ventricular arrhythmia in electrophysiological studies may also identify high-risk patients who are candidates for cardioverter defibrillator implantation. The role of amiodarone in preventing sudden death in high-risk patients with heart failure seems to be small. Further studies are needed to improve risk stratification criteria to select patients with heart failure who are candidates for cardioverter defibrillator implantation. PMID- 15282067 TI - [Images in cardiology. Aneurysm of the sinus of valsalva, pulmonary stenosis, and right ventricular outflow obstruction]. PMID- 15282068 TI - [Right ventricular dysfunction and ischemia in pulmonary embolism]. AB - Patients with pulmonary embolism and right ventricle dysfunction (determined with clinical, hemodynamic or echocardiographic methods) are a subgroup at high risk for complications. One of the pathogenic factors of right ventricular dysfunction in pulmonary embolism is myocardial ischemia, usually secondary to hemodynamic overload, and sometimes worsened by underlying coronary artery disease. We described a patient with pulmonary embolism and dyskinesia of the right ventricular free wall, related to chronic atherosclerotic occlusion of the right coronary artery proximal to the acute marginal branches that irrigate the free wall. PMID- 15282069 TI - [Nocardia endocarditis in a native mitral valve]. AB - Nocardiosis is an opportunistic infection that usually arises in immunodepressed patients. Cases in immunocompetent patients are uncommon. We report a 53-year-old woman diagnosed as having Nocardia sp. endocarditis in a native mitral valve, which required valve replacement. PMID- 15282070 TI - [Severe thrombocytopenia refractory to platelet transfusions, secondary to abciximab readministration, in a patient previously diagnosed with idiopathic thrombocytopenic purpura. A possible etiopathogenic link]. AB - Acute severe thrombocytopenia is a serious although infrequent complication following abciximab infusion that is usually managed with platelet transfusions. We present a patient who underwent multivessel percutaneous coronary intervention with concomitant abciximab administration. Soon after the procedure the patient developed severe thrombocytopenia that persisted despite multiple platelet transfusions. This patient had been previously diagnosed as having idiopathic thrombocytopenic purpura, although this diagnosis was not recorded in our medical records, and at the time of the intervention the patient had a normal platelet count. He was successfully managed with IgG administration. The clinical and therapeutic implications of this case are discussed. PMID- 15282071 TI - [Asymmetric septal hypertrophy associated with "subclinical hypothyroidism"]. AB - We report a case of hypothyroid cardiomyopathy manifested as reversible asymmetric septal hypertrophy in a 61-year-old white woman diagnosed 20 years previously with primary hypothyroidism. PMID- 15282072 TI - [Malnutrition in hospitalized patients: prevalence and economic impact]. AB - BACKGROUND AND OBJECTIVE: Malnutrition constitutes a serious problem of public health. We intended to know the prevalence of undernourishment in our hospital, as well as to assess the expenses generated by its presence. PATIENTS AND METHOD: A study of cohorts was designed to evaluate the nutritional state of randomly selected patients admitted to a third level hospital. We carried out a protocol of nutritional evaluation and quantified the associated costs including drugs, diet, and hospital stay according to diagnosis-related groups. RESULTS: In agreement with anthropometry and the index of corporal mass, the prevalence of malnutrition was 0.3% and 13.4%, respectively. When analyzing the biochemical markers, the prevalence rose to 65.7%. Patients with malnutrition at the time of admission underwent a 59.9% deterioration of their nutritional state. An increase of costs was observed in relation to the length of hospital stay (68.04% compared with normo-nourished patients). Costs related to consumption of medicines, especially antibiotics, were also higher, as well as costs related to nutritional support. CONCLUSIONS: Malnutrition is a phenomenon frequently observed in a hospital setting. Malnutrition associated costs are significant. PMID- 15282073 TI - [Identification of risk times for dissemination of methicillin-resistant Staphylococcus aureus in an intensive care unit]. AB - BACKGROUND AND OBJECTIVE: The dissemination of methicillin-resistant Staphylococcus aureus (MRSA) in patients admitted to an Intensive Care Unit (ICU) depends, among other reasons, on the time interval between obtention of the first positive sample and the establishment of measures for contact isolation. The objective of this study was to identify the risk intervals for the spread of MRSA in ICU patients and to assess the relationship between these periods and the development of new cases of MRSA acquired in the ICU. MATERIAL AND METHOD: Observational and prospective study, which was carried out in a 18-bed polyvalent ICU during a 49-month period (October 1998-October 2002). The exposure risk period was defined as the time elapsed between obtention of the first positive sample and contact isolation of the index case, and the window period as the time elapsed between recovery of the last negative sample to the first positive sample. Infection sources of MRSA were classified into community-acquired and hospital-acquired (nosocomial extra-ICU and nosocomial intra-ICU infections). RESULTS: MRSA was isolated in 69 (2.73%) of 2,531 patients admitted to the ICU during the study period and in all patients measures of contact isolation were indicated. Community-acquired MRSA was diagnosed in 9 (13%) cases, nosocomial intra-ICU in 29 (42%), and nosocomial extra-ICU in 31 (44.9%). The mean duration of the exposure risk period was 3.1 (SD 2.2) (median 3, range 0-9) days and the window period 2.9 (SD 4.6) (median 1, range 0-28) days. In 18 of the 29 cases of intra-ICU-acquired MRSA (62.1%; 95% CI, 42.3-79.3), the infection was acquired within the exposure risk and/or window periods of other patients with MRSA. CONCLUSIONS: The exposure risk periods and the window periods showed a strong relationship between detection of new cases of intra-ICU colonization and/or infection by MRSA. PMID- 15282074 TI - [Validation study of the Multidimensional Alcohol Craving Scale (MACS)]. AB - BACKGROUND AND OBJECTIVE: Craving is a psychobiological phenomenon characteristic of addictive behaviours, which is difficult to assess due to its subjective nature and multidimensionality. The aim of this study was to validate a multidimensional alcohol craving scale (MACS) entirely developed in Spain. MATERIAL AND METHOD: The validation of the MACS was based on an observational study, carried out in 452 alcohol-dependent patients treated with naltrexone. The scales used as standards were an Analogic Visual Scale (AVS) for alcohol craving and the Obsessive-Compulsive Drinking Scale (OCDS). Validity, reliability and sensitivity were studied for the validation process. RESULTS: The factorial analysis (construct validity) showed that the scale had two factors: desire and behavioural disinhibition (lack of resistance). Their correlation values with the assessment scales were r = 0.56 and r = 0.50, respectively. The scale's internal consistency was alpha = 0.94. The correlation of the scores on the scale with the AVS oscillated between 0.56 and 0.46 (p < 0.01) throughout the course of treatment. CONCLUSIONS: The MACS has good psychometric properties: it measures two craving factors --desire and behavioural disinhibition-- and may be considered as an accurate and precise instrument to use in both research studies and medical and psychological clinical practice. PMID- 15282075 TI - [Risk factors and clinical characteristics of thromboembolic venous disease in young patients: a prospective study]. AB - BACKGROUND AND OBJECTIVE: We aimed to determine the risk factors of thromboembolic disease in young patients and to study the clinical characteristics according to the etiology. PATIENTS AND METHOD: A prospective study of 100 patients under 50 years who were not affected by neoplasias or chronic diseases and who required hospitalization due to thromboembolic disease. The morphological diagnosis was performed by eco-Doppler, flebography, lung gammagraphy or CT scan. Risk factors assessed were antithrombin, protein C and S deficiency, presence of factor V Leiden, prothrombin G20210A, hyperhomocisteinemia, increased PAI-I, increased factor VIII, and presence of antiphospholipid antibodies (APAs). Acquired factors were also evaluated. RESULTS: In 87% of patients, a venous thrombosis was observed in lower limbs. 37% of patients had congenital risk factors and 19% had APAs, whereas in the remaining patients only acquired factors were demonstrated. Most frequent congenital factors were factor V Leiden, pothrombin G20210A, and protein C and S deficiency. Most patients presented several risk factors. A family thrombotic history was significantly more frequent in the group with congenital risk factors. CONCLUSIONS: In 56% of young patients with thromboembolic disease, a congenital etiology or APAs are identified. In these patients the number of acquired factors needed to trigger thrombosis is fewer than in patients in whom a cause is not identified. PMID- 15282076 TI - [Malnutrition in inpatients]. PMID- 15282077 TI - [Craving and alcoholism]. PMID- 15282078 TI - [System for accreditation of continuing medical education in Spain]. PMID- 15282079 TI - [General introduction to clinical chronobiology and the therapeutic manipulation of biological rhythms]. AB - In humans, most clinical variables follow biological rhythms approaching to a mathematical function defined by the cosmic-climatic rhythm. Chronophysiology, chronopathology, chronopharmacology, chronotherapy and phototherapy have common elements frequently studied in a disperse manner, thus making it difficult a global understanding of clinical chronobiology as a unitary and well characterized discipline. This review focuses on medical chronobiology, which is much more technical since it only gathers those directives of clinical chronobiology having a health-care impact on daily practice. PMID- 15282080 TI - [Aspirin use in patients with diabetes mellitus]. PMID- 15282081 TI - [Aspirin use in patients with diabetes mellitus]. PMID- 15282082 TI - [Aspirin use in patients with diabetes mellitus]. PMID- 15282083 TI - [Aspirin use in patients with diabetes mellitus]. PMID- 15282084 TI - [Aspirin use in patients with diabetes mellitus]. PMID- 15282086 TI - Subnanogram on-line column-switching liquid chromatographic-tandem mass spectrometric quantification method for nelfinavir and methoxyphenol metabolite M1 in rat plasma. AB - A new on-line, rapid and sensitive column-switch LC/MS/MS method to measure nelfinavir (NFV), an HIV-1 protease inhibitor, and its major metabolite (M1) in rat plasma was developed. Rat plasma containing the analytes and the internal standard was treated with acetonitrile and the supernatant was processed through an on-line extraction and an analytical columns, with a column-switch device. ESI LC/MS with multiple reaction monitors for appropriate analytes was performed. This assay gave a limit of quantitation (LOQ) of <1 ng/mL for the analytes with 5 min run time. The within-run and between-run precisions were <12 and <10%, respectively. This analytical method was successfully applied to a study to correlate changes in maternal and placental NFV plasma concentrations in rats following NFV exposure in utero. PMID- 15282087 TI - Determination of metoprolol, and its four metabolites in dog plasma. AB - Metoprolol and its metabolites alpha-hydroxymetoprolol, O-desmethylmetoprolol, metoprolol acid and deaminated metoprolol were quantified in dog plasma using an isocratic high-performance liquid chromatographic method (with a BetaBasic Cyano column) with fluorescence detection. A solid phase extraction technique (Oasis HLB, Waters) was used to extract metoprolol and its four metabolites from dog plasma with high recovery rates. The method was shown to be sensitive and reproducible and was used to determine the pharmacokinetic profile of metoprolol in dog. To the best of our knowledge, this is the only method that can extract and analyze metoprolol and its four metabolites in a single assay. PMID- 15282088 TI - Use of reversed phase HP liquid chromatography to assay conversion of N acylglycines to primary fatty acid amides by peptidylglycine-alpha-amidating monooxygenase. AB - Primary fatty acid amides (R-CO-NH2) and N-acylglycines (R-CO-NH-CH2-COOH) are classes of compounds that have only recently been isolated and characterized from biological sources. Key questions remain regarding how these lipid amides are produced and degraded in biological systems. Relative to the fatty acids, little has been done to develop methods to separate and quantify the fatty acid amides and N-acylglycines. We describe reversed phase HPLC methods for the separation of C2-C12 primary fatty acid amides and N-acylglycines and also C12-C22 fatty acid amides. Separation within each class occurs primarily on the basis of simple interactions between the acyl chain and the chromatographic stationary phase, but the polar headgroups on these and related fatty acids and N-acylethanolamides modulate the absolute retention in reversed phase mode. We use these methods to measure the enzyme-mediated, two-step conversion of N-octanoylglycine to octanoamide. PMID- 15282089 TI - Development and validation of a chiral liquid chromatographic method, based on Chiralpak to quantify enantiomers of (+/-)-DRF 2725 in rat plasma: lack of inversion of ragaglitazar (S(-)-DRF 2725) to its antipode in plasma. AB - A selective, accurate and reproducible high-performance liquid chromatographic (HPLC) method for the separation of individual enantiomers of DRF 2725 [R(+)-DRF 2725 and S(-)-DRF 2725 or ragaglitazar] was obtained on a chiral HPLC column (Chiralpak). During method optimization, the separation of enantiomers of DRF 2725 was investigated to determine whether mobile phase composition, flow-rate and column temperature could be varied to yield the base line separation of the enantiomers. Following liquid-liquid extraction, separation of enantiomers of DRF 2725 and internal standard (I.S., desmethyl diazepam) was achieved using an amylose based chiral column (Chiralpak AD) with the mobile phase, n-hexane propanol-ethanol-trifluoro acetic acid (TFA) in the ratio of 89.5:4:6:0.5 (v/v). Baseline separation of DRF 2725 enantiomers and I.S., free from endogenous interferences, was achieved in less than 25 min. The eluate was monitored using an UV detector set at 240 nm. Ratio of peak area of each enantiomer to I.S. was used for quantification of plasma samples. Nominal retention times of R(+)-DRF 2725, S(-)-DRF 2725 and I.S. were 15.8, 17.7 and 22.4 min, respectively. The standard curves for DRF 2725 enantiomers were linear (R(2) > 0.999) in the concentration range 0.3-50 microg/ml for each enantiomer. Absolute recovery, when compared to neat standards, was 70-85% for DRF 2725 enantiomers and 96% for I.S. from rat plasma. The lower limit of quantification (LLOQ) for each enantiomers of DRF 2725 was 0.3 microg/ml. The inter-day precisions were in the range of 1.71 4.60% and 3.77-5.91% for R(+)-DRF 2725, S(-)-DRF 2725, respectively. The intra day precisions were in the range of 1.06-11.5% and 0.58-12.7% for R(+)-DRF 2725, S(-)-DRF 2725, respectively. Accuracy in the measurement of quality control (QC) samples was in the range 83.4-113% and 83.3-113% for R(+)-DRF 2725, S(-)-DRF 2725, respectively. Both enantiomers and I.S. were stable in the battery of stability studies viz., bench-top (up to 6 h), auto-sampler (up to 12 h) and freeze/thaw cycles (n = 3). Stability of DRF 2725 enantiomers was established for 15 days at -20 degrees C. The application of the assay to a pharmacokinetic study of ragaglitazar [S(-)-DRF 2725] in rats is described. It was unequivocally demonstrated that ragaglitazar does not undergo chiral inversion to its antipode in vivo in rat plasma. PMID- 15282090 TI - Epimerization study of the L,L- and L,D-diastereoisomers of the calpain inhibitor MDL 28170 by capillary electrophoresis. AB - MDL 28170, Cbz-(L)-Val-(D,L)-Phe-H, which exists as a mixture of L,L- and L,D diastereoisomers, is a calpain inhibitor currently investigated as a novel therapeutic agent for the treatment of ischemic stroke and head and spinal trauma. This report describes a capillary electrophoresis (CE) method that uses sodium dodecyl sulfate (SDS) micellar electrokinetic conditions for the separation of the L,L- and L,D-diastereoisomers of MDL 28170. The report also describes the applications of this CE method to the study of epimerization of the L,L- and L,D-diastereoisomers in pH 7.4 phosphate buffered saline solution (PBS), rat and human plasma at 37 degrees C. The relative percent-time courses obtained showed interconversion of the diastereoisomers in all three matrices studied. However, the epimerization process in rat and human plasma was found to be at least 50 times faster than that in PBS. The epimerization half-life of the L,L diastereoisomer in rat plasma was approximately 30 min, which is about three-fold faster than the observed elimination half-life of the L,L-diastereoisomer reported in a pharmacokinetic study following intravenous bolus dosing. PMID- 15282091 TI - Colloidal gold-based immunochromatographic assay for detection of botulinum neurotoxin type B. AB - A rapid immunochromatographic assay was developed to detect botulinum neurotoxin type B (BoNT/B). The assay was based on the sandwich format using polyclonal antibody (Pab). The thiophilic gel purified anti-BoNT/B Pab was immobilized to a defined detection zone on a porous nitrocellulose membrane and conjugated to colloidal gold particles that served as a detection reagent. The BoNT/B containing sample was added to the membrane and allowed to react with Pab-coated particles. The mixture was then passed along the porous membrane by capillary action past the Pab in the detection zone, which will bind the particles that had BoNT/B bound to their surface, giving a red colour within this detection zone with an intensity proportional to BoNT/B concentration. In the absence of BoNT/B, no immunogold was bound to the solid-phase antibody. With this method, 50 ng/ml of BoNT/B was detected in less than 10 min. The assay sensitivity can be increased by silver enhancement to 50 pg/ml. The developed BoNT/B assay also showed no cross reaction to type A neurotoxin (BoNT/A) and type E neurotoxin (BoNT/E). PMID- 15282092 TI - Automated solid-phase extraction method for the determination of piperaquine in capillary blood applied onto sampling paper by liquid chromatography. AB - A bioanalytical method for the determination of piperaquine in 100 microL blood applied onto sampling paper, by solid-phase extraction and liquid chromatography, has been developed and validated. Blood spots were cut into small pieces prior to addition of 0.3M perchloric acid, acetonitrile and phosphate buffer containing an internal standard. The liquid phase was loaded onto a mixed phase cation-exchange (MPC) solid-phase extraction column. Piperaquine and the internal standard were analysed by liquid chromatography and separated on a Chromolith Performance (100 mm x 4.6 mm) column with acetonitrile:phosphate buffer pH 2.5, I = 0.1 (8:92, v/v) at the flow of 3.5 mL/min. The UV detection was performed at 345 nm. The intra-assay precision was 12.0% at 0.150 microM, 7.3% at 1.25 microM and 7.3% at 2.25 microM. The inter-assay precision was 1.8% at 0.150 microM, 5.2% at 1.25 microM and 2.8% at 2.25 microM. The lower limit of quantification (LLOQ) was determined to 0.050 microM where the precision was 14.7%. PMID- 15282093 TI - Simple determination of capecitabine and its metabolites by liquid chromatography with ultraviolet detection in a single injection. AB - Capecitabine (N4-pentoxycarbonyl-5'-deoxy-5-fluorocytidine, Xeloda), a prodrug of 5-fluorouracil (5-FU), is an oral tumor-selective fluoropyrimidine carbamate approved in the treatment of colorectal and breast cancer. It has a preferential activation to 5-FU by thymidine phosphorilase (TP) in target tumor tissues through a series of three metabolic steps minimizing the exposure of normal tissues to 5-FU. It offers the potential of less gastrointestinal toxicity and advantages in terms of convenience and quality of life for the patient, in addition to cost-effectiveness as compared with intravenous 5-FU chemotherapy. We developed a high performance liquid chromatography assay for the determination of plasma capecitabine and its nucleoside metabolite concentrations and 5-FU catabolite dihydro-5-fluorouracil in a single step extraction and a single HPLC injection. The retention times of dihydro-5-fluorouracil, 5-FU, 5'-deoxy-5 fluorouridine (5'-DFUR) and capecitabine were 3.6, 4.4, 11.4 and 20.4 min, respectively and the internal standard retention times were 8.7 and 12.2 min for 5-bromouracil (5-BU) and tegafur, respectively. The limit of detection was 0.01 microg/ml for capecitabine and its nucleoside metabolites and the limit of quantification was 0.025 microg/ml. Extraction efficiency was >80% with a single solvent mixture extraction step for all analytes of interest. The assay had good precision, the within-day and between-day standard deviation of the mean (R.S.D.) being <10% in the linear range 0.025-10 microg/ml. The authors conclude that the method described here is ideally suited for the therapeutic monitoring of capecitabine and its metabolites. PMID- 15282094 TI - Determination of NG,NG-dimethyl-L-arginine, an endogenous NO synthase inhibitor, by gas chromatography-mass spectrometry. AB - A fully validated gas chromatographic-mass spectrometric (GC-MS) method for the accurate and precise quantification of NG,NG-dimethyl-L-arginine (asymmetric dimethylarginine, ADMA), an endogenous inhibitor of the NO synthase, in cell culture supernatants and in small volumes of plasma is described. ADMA was concentrated by solid phase extraction and converted to its methyl ester pentafluoropropionic amide derivative. The derivatives were analyzed without any further purification. Using gas chromatography-chemical ionization mass spectrometry, fragment ions at m/z 634 and m/z 640 were obtained for ADMA and for NG,NG-[2H6]-dimethyl-L-arginine ([2H6]-ADMA) as internal standard, respectively. [2H6]-ADMA was synthesized by reaction of L-ornithine fastened at bromcyan agarose with dimethylamine. The limit of detection of the method was 2 fmol, while the limit of quantitation for cell culture supernatants was 0.05 microM. The method was validated in a concentration range of 0-1.2 microM in cell culture medium and 0-2 microM in 50 microl aliquots of human plasma. The precision was > or =97% and the accuracy was determined to be > or =94%. This method is fast, rugged and an alternative to high performance liquid chromatography (HPLC) analysis of ADMA in cell culture supernatants and small volumes of human plasma. PMID- 15282095 TI - Measurements of drug-protein binding by using immobilized human serum albumin liquid chromatography-mass spectrometry. AB - An HPLC/MS based method was used for fast and convenient determination of drug plasma-protein interactions in early drug discovery screening by employing a human serum albumin affinity column. Results from this methodology were compared with data from ultrafiltration or dialysis methods, and good agreement was observed. A compound not suitable for ultrafiltration due to the very high non specific binding to artificial membrane of ultrafiltration device was also successfully analyzed by this method, and the protein binding determined by this chromatography method was very similar to data obtained by dialysis technique employing biological membranes. The immobilized HSA column LC/MS method also proved to be more reproducible and precise compared to ultrafiltration method in drug protein binding measurements. PMID- 15282096 TI - Determination of the cyclic depsipeptide FK228, a histone deacetylase inhibitor, by liquid chromatography-mass spectrometry. AB - An analytical method was developed for the quantitative determination of the novel histone deacetylase inhibitor, depsipeptide FK228 (formerly FR901228; NSC 630176), in human plasma. Calibration curves were constructed in the range of 0.5 100 ng/ml, and were analyzed using a weight factor proportional to the nominal concentration. Sample pretreatment involved a liquid-liquid extraction with ethyl acetate using 500 microl aliquots of plasma. The analyte was separated on a column (50 mm x 4.6 mm i.d.) packed with 3.5 microm C8 material, and eluted with methanol-10 mM ammonium formate (55:45; v/v; pH 8). The column effluent was monitored by mass spectrometry with electrospray ionization. The values for precision and accuracy were always < or =7.88% and <3.33% relative error, respectively. The method was successfully applied to examine the pharmacokinetics of FK228 in a cancer patient. PMID- 15282097 TI - Determination of the investigational anti-cancer drug 5,6-dimethylxanthenone-4 acetic acid and its acyl glucuronide in Caco-2 monolayers by liquid chromatography with fluorescence detection: application to transport studies. AB - 5,6-Dimethylxanthenone-4-acetic acid (DMXAA) is a potent cytokine inducer, with a bioavailability of >70% in the mouse. The aim of this study was to develop and validate HPLC methods for the determination of DMXAA and DMXAA acyl glucuronide (DMXAA-G) in the human intestinal cell line Caco-2 monolayers. The developed HPLC methods were sensitive and reliable, with acceptable accuracy (85-115% of true values) and precision (intra- and inter-assay CV < 15%). The total running time was within 6.8 min, with acceptable separation of the compounds of interest. The limit of quantitation (LOQ) values for DMXAA and DMXAA-G were 14.2 and 24 ng/ml, respectively. The validated HPLC methods were applied to examine the epithelial transport of DMXAA and DMXAA-G by Caco-2 monolayers. The permeability coefficient (Papp) values (overall mean +/- S.D., n = 3-9) of DMXAA over 10-500 microM were independent of concentration for both apical (AP) to basolateral (BL) (4.0 +/- 0.4 x 10(-5)cm/s) and BL-AP (4.3 +/- 0.5 x 10(-5)cm/s) transport, and of similar magnitude in either direction, with net efflux ratio (Rnet) values of 1-1.3. However, the Papp values for the BL to AP transport of DMXAA-G were significantly greater than those for the AP to BL transport, with Rnet values of 17.6, 6.7 and 4.5 at 50, 100 and 200 microM, respectively. Further studies showed that the transport of DMXAA-G was Na+- and energy-dependent, and inhibited by MK-571 [a multidrug resistance associated protein (MRP) 1/2 inhibitor], but not by verapamil and probenecid. These data indicate that the HPLC methods for the determination of DMXAA and DMXAA-G in the transport buffer were simple and reliable, and the methods have been applied to the transport study of both compounds by Caco-2 monolayers. DMXAA across Caco-2 monolayers was through a passive transcellular process, whereas the transport of DMXAA-G was mediated by MRP1/2. PMID- 15282098 TI - Validation of a liquid chromatography assay for the quantification of the Raf kinase inhibitor BAY 43-9006 in small volumes of mouse serum. AB - BAY 43-9006 is a selective Raf-1 kinase inhibitor with antitumor activity against a variety of human cancers. A highly sensitive HPLC method for determination of BAY 43-9006 in small volumes of serum (30 microl) was developed. Sample preparation involved a liquid-liquid extraction procedure with tolnaftate as internal standard followed by linear gradient elution at a reversed phase C18 column and UV detection. The method was selective and the calibration curves were linear over the concentration range of 80-2000 ng/ml. The intra-day accuracy ranged from 99.9 to 107.6% and the inter-day accuracy from 94.6 to 115%. The lower limit of quantitation (LOQ) was 80 ng/ml with an accuracy of 105.8%. Thus, this method has been validated and can be applied for the drug monitoring or pharmacokinetic studies of BAY 43-9006 in small volumes of serum samples. PMID- 15282099 TI - Liquid chromatography determination of the anti-androgen vinclozolin and its metabolites in rat serum. AB - The objective of this study was to develop a chromatographic method for the analysis of the anti-androgen vinclozolin (V) and its metabolites 2-[[(3,5 dichlorophenyl)-carbamoyl]oxy]-2-methyl-3-butenoic acid (M1), 3',5'-dichloro-2 hydroxy-2-methylbut-3-enanilide (M2) and 3,5-dichloroaniline (M3) in rat serum. V, M1-M3 were resolved using an HPLC gradient program with a mobile phase consisting of 60-75% methanol:acetonitrile (70:30) and 0.05 M monobasic sodium phosphate buffer pH 3.3 at 1 ml/min, a C18 column, and monitored at 212 nm. Incubates of 0.01 M monobasic potassium phosphate buffer (PB) pH 7.4 and rat serum were spiked with V and its metabolites and processed by diluting samples (1:4) with 0.1M PB pH 3.3, to limit methodological hydrolysis of analytes, followed by addition of acetonitrile. Recoveries of V, M1 and M2 ranged from 85 to 105%, whereas recovery of M3 was <25%. V was hydrolyzed to M1 and M2 after incubation in PB pH 7.4 and rat serum, with M1 the predominant metabolite. This method was successfully applied in the analysis of V and its metabolites in the serum of a male rat after oral administration of V (100 mg/kg). PMID- 15282100 TI - Field-amplified sample stacking in capillary electrophoresis for the determination of clozapine, clozapine N-oxide, and desmethylclozapine in schizophrenics' plasma. AB - A method of field-amplified sample stacking in capillary electrophoresis is described for the simultaneous determination of clozapine (CZP) and its metabolites, clozapine N-oxide (CNO), and desmethylclozapine (DMC), in human plasma. Plasma (0.2 mL) was extracted with organic solvents (ethyl acetate/n hexane/isopropyl alcohol, 8/1/1 by volume) and centrifuged. An aliquot of supernatant was evaporated and suitably reconstituted with water for CE analysis. An untreated fused-silica capillary was used (31.2 cm; effective length, 20 cm; 50 microm i.d.) for the analysis. The background buffer was phosphate buffer (400 mM, pH 3.0) containing 50% ethylene glycol. The separation voltage was 25 kV with a detection wavelength of 214 nm. In the method validation, the calibration curves were linear (r > or = 0.98) over a range of 50-800 ng/mL for CZP, 30-180 ng/mL for CNO, and 25-600 ng/mL for DMC. The relative standard deviation (R.S.D.) and relative error (R.E.) were all less than 11% for the intra- and inter-day assays. The limits of detection (S/N = 3, electric-driven injection, 99.9s) of CZP, DMC, and CNO were 5, 5, and 10 ng/mL, respectively. After continuing treatment with the CZP tablets, a blood sample from one male schizophrenic patient (41-year-old, 62 kg) who had been receiving ongoing treatment with the CZP tablets was prepared and analyzed. The levels of CZP, DMC, and CNO were determined and the feasibility of the method's application in clinical treatment was proven. PMID- 15282101 TI - Selective and rapid liquid chromatography-tandem mass spectrometry assay of dutasteride in human plasma. AB - A simple, rapid, sensitive and specific liquid chromatography-tandem mass spectrometry method was developed and validated for quantification of dutasteride (I), a potent and the first specific dual inhibitor of 5alpha-reductase, in human plasma. The analyte and internal standard (finasteride (II)) were extracted by liquid-liquid extraction with diethyl ether/dichloromethane (70/30, v/v) using a Glas-Col Multi-Pulse Vortexer. The chromatographic separation was performed on a reverse phase Xterra MS C18 column with a mobile phase of 10 mM ammonium formate/acetonitrile (15/85, v/v, pH adjusted to 3.0 with formic acid). The protonated analyte was quantitated in positive ionization by multiple reaction monitoring with a mass spectrometer. The mass transitions m/z 529.5 --> 461.5 and m/z 373.3 --> 317.4 were used to measure I and II, respectively. The assay exhibited a linear dynamic range of 0.1-25.0 ng/mL for dutasteride in human plasma. The lower limit of quantitation was 100 pg/mL with a relative standard deviation of less than 15%. Acceptable precision and accuracy were obtained for concentrations over the standard curve ranges. A run time of 1.2 min for each sample made it possible to analyze a throughput of more than 400 human plasma samples/day. The validated method has been successfully used to analyze human plasma samples for application in pharmacokinetic, bioavailability or bioequivalence studies. PMID- 15282102 TI - Identification of cereal alkylresorcinol metabolites in human urine-potential biomarkers of wholegrain wheat and rye intake. AB - Alkylresorcinols, phenolic lipids present in high amounts in wholegrain wheat and rye, are of interest as potential biomarkers of the intake of these cereals. Alkylresorcinols are known to be absorbed by humans and animals, but little is known about their metabolism or resulting metabolites. A preliminary human study was carried out to identify alkylresorcinol metabolites in human urine. Urine samples, collected before and after a wheat-bran based meal, were deconjugated with beta-glucuronidase/sulphatase and then extracted with ethyl acetate. Extracts were separated by thin-layer chromatography, and fractions containing alkylresorcinols and possible metabolites were identified by retention on the plate compared to standard compounds, and staining with fast blue B. These fractions were further analysed by gas chromatography-mass spectrometry. Deconjugated human urine after the wheat-bran based meal contained two alkylresorcinol metabolites, 3,5-dihydroxybenzoic acid and 3-(3,5 dihydroxyphenyl)-1-propanoic acid, as well as smaller amounts of unchanged alkylresorcinols, confirming the hypothesis that alkylresorcinols are metabolised in humans via beta-oxidation of their alkyl chain. PMID- 15282103 TI - Determination of tetrabromobisphenol A in human serum by liquid chromatography electrospray ionization tandem mass spectrometry. AB - A method for the determination of tetrabromobisphenol A (TBBPA) in human serum utilizing solid-phase extractions (SPEs) and liquid chromatography (LC) with electrospray ionization tandem MS (MS/MS) has been developed. After purification and concentration of TBBPA using consecutive SPEs on reversed-phase and normal phase cartridges, the serum sample was subjected to LC. TBBPA was separated on a C18 reversed-phase column by gradient elution with a mixture of water, methanol, and acetonitrile as the mobile phase, and then detected with electrospray ionization MS/MS in negative ion mode. 13C12-TBBPA was suitable as an internal standard for the reproducible determination of TBBPA in human serum samples (5 g). The method has been validated in TBBPA concentration range of 5-100 pg per g serum, and the recoveries in the concentration range were higher than 83.3%. The repeatabilities of the proposed method of non-spiked control serum (6.3 pg per g serum) and spiked serum (added 5-100 pg per g serum) were within 10.0% as relative standard deviations. The limit of quantification (LOQ) for TBBPA was 4.1 pg per g serum, which was corresponded to 0.63 fmol on column. PMID- 15282104 TI - Studies of phenytoin binding to human serum albumin by high-performance affinity chromatography. AB - High-performance affinity chromatography was used to study the binding of phenytoin to an immobilized human serum albumin (HSA) column. This was accomplished through frontal analysis and competitive binding zonal elution experiments, the latter of which used four probe compounds for the major and minor binding sites of HSA injected into the presence of mobile phases containing known concentrations of phenytoin. It was found that phenytoin can interact with HSA at the warfarin-azapropazone, indole-benzodiazepine, tamoxifen, and digitoxin sites of this protein. The association constants for phenytoin at the indole benzodiazepine and digitoxin sites were determined to be 1.04 (+/-0.05) x 10(4)M( 1) and 6.5 (+/-0.6) x 10(3)M(-1), respectively, at pH 7.4 and 37 degrees C. Both allosteric interactions and direct binding for phenytoin appear to take place at the warfarin-azapropazone and tamoxifen sites. This rather complex binding system indicates the importance of identifying the binding regions on HSA for specific drugs as a means for understanding the transport of such substances in blood and in characterizing their potential for drug-drug interactions. PMID- 15282105 TI - Large-volume sample stacking-capillary electrophoresis used for the determination of 3-nitrotyrosine in rat urine. AB - Large-volume sample stacking using the electroosmotic flow (EOF) pump technique has been investigated for the quantification of 3-nitrotyrosine in urine of diabetic rats. The best separation conditions for these highly complex samples were obtained using capillary electrophoresis (CE) in the reversed polarity mode (i.e., injecting at the cathode and detecting at the anode) using cetyltrimethylammonium bromide (CTAB) in the running buffer. The optimum CE separation conditions were achieved using a phosphate buffer prepared with 0.15M phosphoric acid and 0.5 mM CTAB adjusted to pH 6.4 with sodium hydroxide. In such CE conditions, the limit of detection (LOD) was 1.77 microM for 3-nitrotyrosine with normal injection mode, meanwhile with the large-volume sample stacking technique a more than 20-fold improvement was observed (i.e., LOD = 0.08 microM was obtained) without noticeable loss of resolution. This value allowed the detection of 3-nitrotyrosine in urine from diabetic rats. To our knowledge, this work is one of the few applications showing the great possibilities of these stacking procedures to analyse biological samples by CE. PMID- 15282106 TI - Discovery stage pharmacokinetics using dried blood spots. AB - Early in the discovery stage, the measurement of drug candidates in biological fluids as a function time provides important information used in decision making for lead optimization. The detection methodology primarily used is liquid chromatography coupled to triple quadrupole mass spectrometry (LC-MS). Sample preparation is an important aspect of these experiments and robotic-based automation is commonly used. The often overlooked aspect of these experiments is the sample collection itself. Typically, several hundred microliters of whole blood is collected and the plasma fraction separated for each time-point. The plasma is then transferred to an appropriate vessel for subsequent aliquoting and processing. We describe a method for performing discovery stage pharmacokinetic analysis using whole blood dried onto filter paper. The use of dried blood spots is a well established technique for neo-natal screening, and its application to early screening of drug candidates proves to be robust, reliable and reproducible. PMID- 15282107 TI - An HPLC method for the determination of lisinopril in human plasma and urine with fluorescence detection. AB - A selective, sensitive and precise HPLC method with fluorimetric detection has been developed for the assay of lisinopril in human plasma and urine. The clean up of the sample was carried out by solid-phase extraction, firstly with C18 cartridge and secondly with a silica-cartridge. After a pre-column derivatization with fluorescamine, the reaction mixture was chromatographed on C18-column with gradient elution, using methanol and 0.02 M phosphate buffer (pH=3.2). The fluorescamine-lisinopril derivative was detected fluorimetrically by monitoring the emission at 477 nm, with excitation at 383 nm. Linear quantitative response curve was generated over a concentration range of 5-200 ng/ml and 25-1000 ng/ml for plasma and urine samples, respectively. The mean recovery of lisinopril from plasma and urine was 63.41 and 74.08%, respectively. Intra-day and inter-day R.S.D. and R.M.E. values at three different concentrations were assessed. The method was applied for pharmacokinetic study in a healthy volunteer after a single oral dose of 20 mg of the drug. PMID- 15282108 TI - Determination of tetracationic zinc(II) phthalocyanine derivative RLP068 in rabbit serum by liquid chromatography-tandem mass spectrometry. AB - The development and validation of a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the determination of the tetracationic zinc(II) phthalocyanine derivative RLP068 in rabbit serum is described. The dodecadeuterated product (RLP068-D12) was used as co-eluting internal standard. RLP068 was isolated from serum samples by solid-phase extraction using weak cationic exchange cartridges (WCX). An oxidative derivatisation was used in order to simplify the peculiar HPLC and MS behaviour of the analyte and thus increasing sensitivity. Liquid Chromatography was carried out on a Polaris C18 Ether column (50 mm x 2.0 mm) with an isocratic run of 0.5% aqueous TFA/methanol. Detection was achieved by means of a Bruker Esquire 3000+ Ion Trap Mass Spectrometer equipped with an ESI source working in positive mode. A Multiple Reaction Monitoring method following the transitions 297.1 --> 282.1 for the analyte and 300.1 --> 282.1 + 285.1 for the internal standard was used. The analytical method was validated over the concentration range 2-65 ng/mL. lower limits of detection (LLOD) and quantification (LLOQ) were respectively 1 and 2 ng/mL. The method is innovative and applicable to pharmacokinetic studies. PMID- 15282109 TI - The use of a monolithic column to improve the simultaneous determination of four cephalosporin antibiotics in pharmaceuticals and body fluids by HPLC after solid phase extraction--a comparison with a conventional reversed-phase silica-based column. AB - The use of a monolithic column (Chromolith, SpeedROD RP-18e, by Merck) was studied on the determination of cephalosporin antibiotics. Results were compared with those from a previously developed analytical method using conventional silica-based analytical column. A rapid, accurate and sensitive method has been developed and validated for the quantitative simultaneous determination of four cephalosporins: Cephalexine and Cephadroxil (first generation), Cefaclor (second generation) and Cefotaxim (third generation) in pharmaceuticals as well as in human blood serum and urine. Hydroflumethiazide (HFM) (3,4-dihydro 6(trifluoromethyl)-2H-1,2,4-benzothiadiazine-7-sulfonamide-1,1-dioxide) was used as an internal standard at a concentration of 1.5 ng/microL. A rectilinear relationship was observed up to 5 ng/microL for the four compounds. Analysis time was less than 4 min. The statistical evaluation of the method was examined by means of within-day repeatability (n=8) and day-to-day precision (n=8) and was found to be satisfactory with high accuracy and precision results. The method was applied to the determination of the cephalosporins in commercial pharmaceuticals and in biological fluids: human blood serum after solid phase extraction and urine simply after filtration and dilution. Recovery of analytes in spiked serum samples was in the range from 88.7 to 107.8%, while for urine samples recovery was from 98.0 to 105.6%. By comparing the figures of merit for the monolithic column and the silica-based one, regarding the determination of the four cephalosporins investigated in the present study, the outstanding efficiency of the monolithic column can be noticed. PMID- 15282110 TI - Analytical method for evaluation of exposure to benzene, toluene, xylene in blood by gas chromatography preceded by solid phase microextraction. AB - Frequency of intentional exposure to organic solvents has been increasing among children and adolescents in Brazil. Analysis of benzene, toluene and xylenes (BTX) in human blood is necessary to diagnose the intentional and accidental exposure to these solvents. A method for BTX determination in blood samples by gas chromatography preceded by solid phase microextration (SPME) from headspace (HS) has been described. SPME has several advantages when compared to other extraction techniques such as simplicity, low cost and solvent-free extraction. The method presents good repeatability (precision was of 2.2-8.0%), accuracy from -4.7 to -9.4%, limit of detection <1.0 ug/mL, linearity from 1.0 to 100 ug/mL for toluene and from 5.0 to 100 ug/mL for the other solvents (R2 > 0.99), which shows to be efficient and adequate for the detection of exposure to BTX in blood samples. PMID- 15282112 TI - Glucuronidation of aliphatic alcohols in human liver microsomes in vitro. AB - The glucuronidation of several short-chained aliphatic alcohols in vitro, with the use of human liver microsomes (HLM) as catalyst, was performed, and the kinetics were studied. The concentrations of the glucuronides were determined by gas chromatography-mass spectrometry after derivatization to the trimethylsilyl compounds. Alcohols from ethanol to pentanols were found to couple with activated glucuronic acid in HLM to a widely varying amount. For analytic reasons the glucuronide of methanol, which is formed after methanol consumption in human beings in vivo, could not be determined. The length of the alkyl chain played the decisive role in the maximum turnover rate of the glucuronidation. The affinity of the alcohols to UDP-glucuronosyltransferase (UDPGT), which catalyzes the glucuronidation reaction, was shown to depend strongly on their structure. Alcohols with a very short alkyl chain and secondary alcohols were glucuronidated much more slowly in comparison with findings for the longer chain primary alcohols and showed less affinity to UDPGT. The alcohols mutually inhibited glucuronidation. However, ethanol inhibited the glucuronidation of isopentanol or n-pentanol only in high concentrations, whereas the two pentanols inhibited each other to a high degree. The glucuronidation of aliphatic alcohols is probably catalyzed by only one of several very similar enzymes of the UDPGT. This finding was indicated by the fact that the Michaelis-Menten constants of the alcohols- with the use of different lots of the HLM from different liver donors--had nearly the same values. PMID- 15282111 TI - Ethanol and acetaldehyde in alcoholic cardiomyopathy: from bad to ugly en route to oxidative stress. AB - Alcoholic cardiomyopathy is characterized by cardiomegaly, disruptions of myofibrillary architecture, reduced myocardial contractility, decreased ejection fraction, and enhanced risk of stroke and hypertension. Although several mechanisms have been postulated for alcoholic cardiomyopathy, including oxidative damage, accumulation of triglycerides, altered fatty acid extraction, decreased myofilament Ca(2+) sensitivity, and impaired protein synthesis, neither the mechanism nor the ultimate toxin has been unveiled. Primary candidates acting as specific toxins of myocardial tissue are ethanol; its first and major metabolic product, acetaldehyde; and fatty acid ethyl esters. Acetaldehyde has been demonstrated to impair directly cardiac contractile function, disrupt cardiac excitation-contractile coupling, and contribute to oxidative damage and lipid peroxidation. Acetaldehyde-elicited cardiac dysfunction may be mediated through cytochrome P450 oxidase, xanthine oxidase, and the stress-signaling cascade. Unfortunately, the most direct approach that can be used to examine toxicity is hampered by the fact that direct intake of acetaldehyde is highly toxic and unsuitable for long-term study. To overcome this obstacle, transgenic mice have been used to alter artificially ethanol/acetaldehyde metabolism, resulting in elevated acetaldehyde concentrations after ethanol ingestion. In this review, we summarize results obtained with the use of transgenic animal models to elucidate the role of acetaldehyde in the mechanism of action in alcoholic cardiomyopathy. PMID- 15282113 TI - Acute ethanol exposure modulates expression of inducible nitric-oxide synthase in human astroglia: evidence for a transcriptional mechanism. AB - Astroglia are important in immunocompetence and response to injury within the CNS. Activated astroglia respond, in part, by expressing inducible nitric-oxide synthase (iNOS) and subsequent catalytic production of nitric oxide. Results from a previous study in our laboratory, in the human A172 astroglial cell line, revealed that induction of iNOS activity by tumor necrosis factor-alpha + interferon-gamma + interleukin-1 beta was inhibited by 24-h exposure to a high ethanol concentration (200 mM), but enhanced by 50 mM ethanol. In the work reported in this article, we tested the working hypothesis that ethanol acts transcriptionally to modulate cytokine-induced expression of the iNOS gene, NOS2A, in human astroglia. Ethanol, 50 or 200 mM, did not directly alter in vitro catalytic activity of the iNOS enzyme, indicating that ethanol does not affect the enzyme directly. Likewise, ethanol exposure after a 12-h cytokine-stimulation period had no effect on in vivo iNOS activity. However, when cells were simultaneously exposed to ethanol and cytokines for 12 h, in vivo iNOS activity was altered. That ethanol must be present during cytokine stimulation to influence iNOS activity is consistent with a transcriptional mechanism of action. In addition, steady-state expression of iNOS protein and NOS2A mRNA levels were modulated in a biphasic manner by ethanol similar to that noted previously for iNOS activity. These findings strongly support the suggestion that ethanol modulates cytokine-induced iNOS expression in A172 cells at a pretranslational site. These findings should be instrumental in the identification of the critical ethanol-sensitive elements involved in the regulation of NOS2A in human astroglia. PMID- 15282114 TI - Devaluation of ethanol reinforcement. AB - Postingestive CNS pharmacologic effects of ethanol are often assumed to provide the major stimuli for development and maintenance of ethanol self-administration in rats. However, there is little direct evidence to support this assumption. In all procedures that have been used to initiate ethanol intake in rats, some type of taste adaptation or taste conditioning could account for the increased and maintained ethanol intake. Thus, it remains critical to demonstrate that increased ethanol intake is related to postingestive CNS actions of ethanol, and not to a positive shift in the hedonic taste value of the solution. Two experiments were performed to examine this question. In both studies, rats were trained to self-administer 20% ethanol by using a sucrose-substitution initiation procedure. The rats were required to press a lever 25 or 30 times to gain access to 20% ethanol for 20 min from a sipper tube. Once initiated, extinction sessions were used to determine the strength of ethanol seeking by measuring the number of lever presses that occurred in 20 min with no presentation of the ethanol solution. After initial training, the rats were split into two groups: one that received pairings of a gavage of ethanol (1 g/kg), followed after 10 min by a lithium chloride (LiCl) injection (paired group), and one that also received ethanol gavage and LiCl injections, but separated by 24 h (unpaired group). This pairing of postingestive effects with the illness induced by LiCl injection has been shown to devalue other food and fluid reinforcers. In Experiment 1, the rats received four pairings, one after the other with no behavioral testing between. In Experiment 2, the rats received three pairings and were tested for devaluation after each pairing. Results from both experiments showed significant decreases in seeking behavior in both groups, but seeking behavior was decreased significantly greater in the paired group, even though neither group had access to ethanol during the extinction testing periods. In Experiment 1, when ethanol became available after the devaluation procedure, the pattern of intake in the paired group was unchanged early in the sipper tube availability period, supporting the suggestion that the devaluation effect was not mediated by taste stimuli. These findings support the assumption that postingestive effects contribute to the reinforcement produced by self-administered ethanol in rats. PMID- 15282115 TI - Effects of ethanol consumption on vasopressin and neuropeptide Y immunoreactivity and mRNA expression in peripheral and central areas related to cardiovascular regulation. AB - Results from previous studies have demonstrated that ethanol influences central neural mechanisms involved in the control of blood pressure. We studied the effects of ethanol consumption on vasopressin and neuropeptide Y immunoreactivity and mRNA expression in the nucleus tractus solitarius and paraventricular hypothalamic nucleus, as well as in the petrosal and nodose ganglia of rats. The ethanol-fed rats received liquid diet ad libitum containing 37.5% ethanol-derived calories (6.7% volume/volume), and the pair-fed rats received the same volume of diet containing isocaloric amounts of maltose-dextrin substituted for ethanol for 3 or 28 days. Arterial blood pressure was evaluated in a separate group of rats, which was unchanged by 3 days, but elevated by 21% after 28 days of ethanol consumption. Vasopressin immunoreactivity and mRNA signal were not detected in the ganglia, nor were they changed in the nucleus tractus solitarius and paraventricular hypothalamic nucleus, by 3 days of ethanol consumption. However, after 28 days of ethanol liquid diet consumption, vasopressin-positive terminals were decreased in the nucleus tractus solitarius and vasopressin immunoreactivity cell bodies and mRNA signal were decreased in the paraventricular hypothalamic nucleus. Neuropeptide Y-immunoreactive terminals were increased in the nucleus tractus solitarius only after 28 days of ethanol liquid diet consumption, but they were decreased in the paraventricular hypothalamic nucleus in rats treated with ethanol for 3 or 28 days. We concluded that the levels of both vasopressin and neuropeptide Y neurotransmitters are changed by long-term ethanol consumption in the neuronal pathways related to control of blood pressure. PMID- 15282116 TI - Mouse striatal transcriptome analysis: effects of oral self-administration of alcohol. AB - Results of recent studies support the notion that substance self-administration is partially a genetically controlled component of addiction tied to habit formation and cellular modification of the striatum. Aiming to define pathways among genomic, neural, and behavioral determinants of addiction, we investigated global striatal gene expression in a paradigm of oral self-administration of alcohol by using genomically very similar alcohol-nonpreferring B6.Cb(5)i(7) alpha 3/Vad (C5A3) and alcohol-preferring B6.Ib(5)i(7)-beta 25A/Vad (I5B25A) quasi-congenic mouse strains and their progenitors, C57BL/6By (B6By) and BALB/cJ. Expression of 12,488 genes and expressed sequence tags (ESTs) was studied by using 24 high-density oligonucleotide microarrays. Transcript signal intensity differences were analyzed with z test after iterative median normalization across groups and Hochberg step-down Bonferroni procedure. As expected, striatal transcriptome differences were far more extensive between the independently derived progenitor strains than between the quasi-congenic strains and their background partner, B6By. However, the genes, which were differentially expressed between the quasi-congenic strains and their background partner, were not subsets of the progenitorial differences and were not located on the chromosome segments introgressed into the quasi-congenic strains from the donor BALB/cJ strain that have been so far defined. Although 25 transcripts showed significantly different expression between the progenitor strains, only two transcripts, phosphatidylserine decarboxylase and a hypothetical 21.2-kDa protein, and one transcript, molybdenum co-factor synthesis 2, showed significantly different expression between C5A3 and I5B25A, and between B6By and I5B25A, respectively. The latter three transcripts are not located on previously identified chromosome segments introgressed from the donor BALB/cJ strain, supporting the suggestion of trans-acting regulatory variations among strains. Exposure to alcohol did not induce statistically significant striatal gene expression changes in any of the mouse strains. In conclusion, the results support the hypothesis that in functional genomic studies the chance of detecting function-relevant genes can be increased by the comparative analysis of quasi-congenic and background strains because the number of functionally irrelevant, differentially expressed genes between genomically similar strains is reduced. Lack of statistically significant alcohol-induced changes in transcript abundance indicated that oral self administration had subtle effects on striatal gene expression and directed attention to important implications for the experimental design of future microarray gene expression studies on complex behaviors. PMID- 15282117 TI - Serum cytokine concentrations in alcohol-dependent individuals without liver disease. AB - In the current study, our aim was to evaluate and investigate the influence of heavy alcohol intake on serum interleukin (IL)-6, IL-8, IL-10, IL-12, and tumor necrosis factor-alpha (TNF-alpha) concentrations. The selection of cytokines was based on their presumptive role in the pathophysiology of alcohol dependence. On admission to the Drug-Free Substance Addiction Detoxification clinic ("ATHENA"), blood samples were obtained from study participants, and serum cytokine concentrations were measured by using a commercial sandwich enzyme-linked immunosorbent assay technique. Alcohol dependence, as diagnosed according to DSM IV [Diagnostic and Statistical Manual of Mental Disorders (4th ed.)] criteria for alcohol dependence and estimated by using the Composite International Diagnostic Interview (CIDI), was characterized by increased serum IL-6 concentration. Interleukin-8, IL-10, IL-12, and TNF-alpha concentrations were comparable to those found in control subjects (P>.05). These results indicate that in alcohol dependent individuals there is a significant increase in the serum IL-6 concentration (P <.05). PMID- 15282118 TI - Inflammatory brain damage in preterm newborns--dry numbers, wet lab, and causal inferences. AB - Epidemiologic observations support the contention that infection, inflammation, and neonatal white matter damage (WMD) are associated. We also have documentation from multiple experimental models that infection/inflammation can damage developing white matter. Based on these observations in humans and animals, we offer causal inferences using widely accepted causal criteria and the multivariable model of causation. As much as we want to, however, we are reluctant to state unequivocally that inflammation causes WMD in humans born much before term. The main reason is that we lack convincing evidence that inflammation precedes WMD (temporal evidence). We also need more (and more detailed) observational studies clarifying the presumed infection --> inflammation --> WMD sequence before we can initiate intervention trials to reduce the risk of WMD. PMID- 15282119 TI - Heart rate variability in response to the sleep-related movements in infants with and without colic. AB - The activity of the autonomic nervous system depends on sleep stage. The imbalance of the autonomic nervous system together with over-reactivity to stimuli has been suggested to be an etiologic factor for infantile colic. This study was designed to estimate the reactivity of the autonomic nervous system to a sleep-time stimulus in non-rapid eye movement (NREM) and rapid eye movement (REM) sleep stages and in colic and control infants. Overnight sleep polygraphic recordings were performed for 12 colic and 14 control infants at the age of 8 weeks. Movements were detected by a static-charge-sensitive bed. Extent of heart rate variability (HRV) was measured in response to spontaneous sleep-related movements. HRV analysis was performed over 2-min segments during NREM and REM sleep before and after 5-36-s long movement periods. Total (0.04-1.0 Hz), low (0.04-0.15 Hz) and high frequency (0.15-1.0 Hz) HRV increased after the movement periods in light NREM sleep (p < 0.001). These changes were not observed in REM sleep. No differences were found between the colic and the control groups in HRV. The observed difference in the response of the HRV between sleep stages is likely to reflect the different characteristics of heart rate control in NREM and REM sleep, but our results do not suggest that colic infants would have abnormal autonomic reactivity to stimuli while asleep. PMID- 15282120 TI - Respiratory characteristics of infants with pulmonary hypoplasia syndrome following preterm rupture of membranes: a preliminary study for establishing clinical diagnostic criteria. AB - BACKGROUND: At present, the diagnosis of pulmonary hypoplasia is based on postmortem findings, and there are no clear clinical diagnostic criteria to facilitate its identification and management. AIM: To characterise the respiratory status of pulmonary hypoplasia syndrome (PHS) following preterm rupture of membranes so as to establish its clinical diagnostic criteria. STUDY DESIGN: Retrospective comparison of respiratory characteristics of six typical PHS infants with six wet lung syndrome (WLS) infants who served as controls. SUBJECTS: The PHS and WLS infants were selected from 1094 patients admitted to a tertiary care neonatal unit over a 6-year period, with criteria based on perinatal history, respiratory signs, X-ray and laboratory findings, and ventilator settings. OUTCOME MEASURES: The compared variables were lung volume index (LVI) calculated from lung dimensions on chest X-ray, ventilatory index (VI), ventilatory efficiency index (VEI), response to artificial surfactant treatment, and ventilation days. RESULTS: In PHS compared to WLS infants, LVI was lower (4.5 +/- 0.5 vs. 9.5 +/- 1.5; p < 0.01), VI was higher (0.108 +/- 0.030 vs. 0.022 +/- 0.005; p < 0.05), and VEI was lower (0.083 +/- 0.012 vs. 0.258 +/- 0.052; p < 0.01) (mean +/- S.E.). Artificial surfactant was given to four PHS infants, but none of them showed respiratory improvement. Ventilation days were 11-79 in three surviving PHS infants and 2-14 in WLS infants. CONCLUSIONS: In this preliminary study, low LVI (< 6.5) and VEI (< 0.15) were the most useful indicators of PHS. PMID- 15282121 TI - Urogenital anomalies in human male fetuses. AB - BACKGROUND: There are few studies in the literature performed in human fetuses evaluating the incidence of genitourinary anomalies. AIMS: Analyze the incidence of congenital urogenital malformations in human male fetuses. STUDY DESIGNS AND SUBJECTS: We analyzed 166 human male fetuses well preserved. The gestational age was determined in weeks post conception (WPC) according to the foot length criterion and ranged from 10 to 35 WPC. The fetuses were dissected with the aid of a stereoscopic lens with 2.5x magnification. We performed abdominal and pelvic incisions to expose the urogenital organs. We studied the incidence of renal, ureteral, vesicle, urethral, testicular, epididymal, vas deferens, prostate and penile anomalies. RESULTS: Of the 166 fetuses, 7 (4.2%) presented some kind of anomaly of the urogenital system. Renal anomalies were found in two fetuses (1.2%). Unilateral renal agenesis was found in a 25 WPC fetus. Horseshoe kidney was found in a 20 WPC fetus. In a 23 WPC fetus (0.6%) the two testes were absent. Epididymal disjunction anomalies were found in four fetuses (2.4%). CONCLUSIONS: The urogenital anomalies in human male fetuses are rare and have an incidence around 4%. PMID- 15282122 TI - Influences of maternal cigarette smoking on infant arousability. AB - Since the reduction in the incidence of the prone sleeping position, maternal cigarette smoking has become the strongest modifiable risk factor for Sudden Infant Death Syndrome (SIDS). This risk is dose dependent. Various mechanisms have been postulated to explain the increased risk of SIDS associated with maternal smoking, among these, impairment of arousal from sleep. This paper reviews the effects of maternal smoking on infant arousability from sleep, cardiorespiratory controls and sleep architecture. Infants exposed to maternal smoking have been shown to have both decreased spontaneous and evoked arousability from sleep. Such impairment of arousal has been demonstrated to be associated with changes in control of autonomic cardiac function. Sleep architecture appears not to be altered by smoking. During arousal, heart rate, blood pressure and breathing movements increase, while gross body movements occur to avoid the stimulus. Any impairment in arousability from sleep could occur when infants are exposed to maternal cigarette smoking, and could possibly contribute to the final pathway to SIDS. PMID- 15282123 TI - Occurrence of Campylobacter in retail foods in Ireland. AB - A surveillance study was carried out to determine the prevalence of Campylobacter in a range of retail foods purchased in three Irish cities over a 20-month period between March 2001 and October 2002. In total 2391 food samples were analysed during this period. Campylobacter was isolated from 444 raw chicken (49.9%), 33 turkey (37.5%) and 11 duck samples (45.8%). Lower isolation rates of 7/221 (3.2%), 10/197 (5.1%) and 31/262 (11.8%) were observed for raw beef, pork and lamb, respectively. One sample of pork pate from 120 samples analysed (0.8%) was Campylobacter-positive. A total of three shellfish samples (oysters) from 129 raw specimens examined (2.3%) were found to contain Campylobacter. Low prevalences of the organism (0.9%) were also isolated from fresh mushrooms. Of 62 raw bulk tank milk samples analysed, Campylobacter was recovered in a single sample (1.6%). Campylobacter was not detected in any of the comminuted pork puddings, prepared vegetables and salads, retail sandwiches or cheeses made from unpasteurised milk. In total, 543 Campylobacter were isolated from all of the food samples analysed, of which 453 (83.4%) were confirmed as Campylobacter jejuni and the remaining 90 (16.6%) as Campylobacter coli. PMID- 15282124 TI - Yeast ecology of Kombucha fermentation. AB - Kombucha is a traditional fermentation of sweetened tea, involving a symbiosis of yeast species and acetic acid bacteria. Despite reports of different yeast species being associated with the fermentation, little is known of the quantitative ecology of yeasts in Kombucha. Using oxytetracycline-supplemented malt extract agar, yeasts were isolated from four commercially available Kombucha products and identified using conventional biochemical and physiological tests. During the fermentation of each of the four products, yeasts were enumerated from both the cellulosic pellicle and liquor of the Kombucha. The number and diversity of species varied between products, but included Brettanomyces bruxellensis, Candida stellata, Schizosaccharomyces pombe, Torulaspora delbrueckii and Zygosaccharomyces bailii. While these yeast species are known to occur in Kombucha, the enumeration of each species present throughout fermentation of each of the four Kombucha cultures demonstrated for the first time the dynamic nature of the yeast ecology. Kombucha fermentation is, in general, initiated by osmotolerant species, succeeded and ultimately dominated by acid-tolerant species. PMID- 15282125 TI - Expression of the catalase gene katA in starter culture Lactobacillus plantarum TISTR850 tolerates oxidative stress and reduces lipid oxidation in fermented meat product. AB - The catalase gene katA of Lactobacillus sakei SR911 was cloned and expressed in Escherichia coli UM2 and Lactobacillus plantarum TISTR850 under strong lactococcal promoter P59 in E. coli-lactococcus expression vector pIL1020. The L. plantarum TISTR850 is a catalase-deficient strain isolated from local fermented meat product. The recombinant L. plantarum TISTR850 was shown to decompose hydrogen peroxide, and catalase activity approximately three times higher that of natural catalase-producing strain L. sakei SR911. The recombinant protein was also detected by in situ activity staining of the catalase enzyme. The recombinant L. plantarum TISTR850 did not accumulate hydrogen peroxide under glucose-limited aerobic conditions and remained viable after 60 h of incubation. The recombinant and host strain L. plantarum TISTR850 were used as starter cultures in the fermented meat product, and lipid oxidation was monitored over a 7-day storage at 20 degrees C determined as thiobarbituric acid-reactive substances (TBARS) value. The lipid oxidation level in the fermented meat product seeded with the catalase genetically modified starter culture L. plantarum TISTR850 was significantly lower than that of the natural catalase-deficient strain. PMID- 15282126 TI - Effect of hydrogen peroxide treatment on microbial quality and appearance of whole and fresh-cut melons contaminated with Salmonella spp. AB - The efficacy of hydrogen peroxide treatment on the inactivation of Salmonella spp. inoculated on the external surface of cantaloupe and honeydew melon was investigated. Salmonella was inoculated onto whole cantaloupe and honeydew melon to a final concentration of 4.65 log(10) CFU/cm(2) and 3.13 log(10) CFU/g, respectively. Inoculated whole melons stored at 5 degrees C for up to 7 days were washed with water, 2.5% and 5% hydrogen peroxide at day 0 and 5. Hydrogen peroxide (2.5% and 5%) treatments of whole melon for 5 min caused a 3 log(10) CFU/cm(2) reduction of the indigenous surface microflora and a 3.0 log(10) CFU/cm(2) reduction in Salmonella spp. on all melon surfaces. The efficacy of the hydrogen peroxide treatments was less when the interval between inoculation and treatment of cantaloupe exceeded 24 h. Unlike cantaloupe fresh-cut pieces, Salmonella was not recovered from fresh-cut pieces prepared from treated whole honeydew melon. Growth of Salmonella occurred in cantaloupe fresh-cut pieces stored at 10 or 20 degrees C, and by 2 weeks, levels reached approximately 1 log CFU/g. A rapid decline in appearance and overall acceptability was observed in fresh-cut pieces prepared from untreated whole cantaloupe. While Salmonella was recovered from fresh-cut pieces from and whole treated cantaloupe, sanitizing the surface of contaminated whole melons with hydrogen peroxide before and after cutting and storage of the fresh-cut pieces at 5 degrees C can enhance the microbial safety and acceptability rating for about 2 weeks after processing. PMID- 15282127 TI - Chitosan kills bacteria through cell membrane damage. AB - The bactericidal activity of chitosan (CS) acetate solution against Escherichia coli and Staphylococcus aureus was evaluated by the enumeration of viable organisms at different incubation times. Morphologies of bacteria treated with CS were observed by transmission electron microscopy (TEM). The integrity of the cell membranes of both species and the permeabilities of the outer membrane (OM) and inner membrane (IM) of E. coli were investigated by determining the release from cells of materials that absorb at 260 nm, changes in the fluorescence of cells treated with the fluorescent probe 1-N-phenylnaphthylamine (NPN) and release of cytoplasmic beta-galactosidase activity. In addition, the interaction of CS with synthetic phospholipid membranes was studied using gel permeation chromatography (GPC), UV-VIS spectrophotometery, Fourier-transform infrared spectroscopy (FT-IR) and thermal analysis. Results showed that CS increased the permeability of the OM and IM and ultimately disrupted bacterial cell membranes, with the release of cellular contents. This damage was likely caused by the electrostatic interaction between NH(3)(+) groups of CS acetate and phosphoryl groups of phospholipid components of cell membranes. PMID- 15282128 TI - Application of central composite design and response surface methodology to the fermentation of olive juice by Lactobacillus plantarum and Debaryomyces hansenii. AB - The objectives of this study were to investigate the effects of NaCl, calcium acetate and calcium lactate in concentrations corresponding to ionic strengths equivalent to 2-10%, w/v salt brines as well as the 50% replacement of NaCl contained in the above mixture by KCl. A central composite design and response surface methodology were used to optimize the maximum specific growth rate of Lactobacillus plantarum ATCC 8014 and Debaryomyces hansenii 2114. The fermentation was carried out in olive juice obtained from Kalamon black olives at 30 degrees C with initial pH 5.0. Mathematical models describing the combined effects of these factors on the maximum specific growth rate of L. plantarum or D. hansenii were established. Both strains in single cultures showed higher maximum specific growth rate in olive juice supplemented with NaCl/KCl, Ca acetate and Ca-lactate. But in mixed culture fermentations of olive juice supplemented with NaCl, Ca-acetate and Ca-lactate, higher specific growth rates were obtained. Under the optimum growth conditions determined for the single culture fermentations, i.e. 378.4 mM NaCl, 34.1 mM Ca-acetate and 39.9 mM Ca lactate, mixed culture fermentation was undertaken by varying the time of inoculation of the yeast strain. When D. hansenii was inoculated 48 h before L. plantarum the maximum specific growth rate of L. plantarum was increased to 0.247 per hour, which was significantly higher compared to L. plantarum alone (0.211 per hour). In mixed culture fermentation of olive juice supplemented with the mixture of NaCl/KCl under similar conditions as above, a maximum specific growth rate of L. plantarum of 0.218 per hour was determined. The optimum conditions determined for mixed culture fermentation are useful in fermentation of black olives Kalamon variety under lower salt content. PMID- 15282129 TI - Predictive modelling of growth and measurement of enzymatic synthesis and activity by a cocktail of Brochothrix thermosphacta. AB - The possibility was examined of developing a predictive model that combined microbial growth (increase in cellular number) and extracellular enzyme activity of a cocktail of three strains of Brochothrix thermosphacta. Estimations of growth and enzyme activity were made within a three-dimensional matrix of conditions: temperature 2-20 degrees C, pH value 4.0-7.5 and water activity (a(w)) 0.95-0.995. A model which predicted growth based on increases in cell number was constructed. No extracellular lipases were detected, but slight proteolytic reactions were observed. Although it was not possible to model protease activity, the growth model and information relating to enzyme activity will be made freely available in a database on the Internet. PMID- 15282130 TI - Application of a molecular beacon-real-time PCR technology to detect Salmonella species contaminating fruits and vegetables. AB - An oligonucleotide probe that becomes fluorescent upon hybridization to the target DNA (molecular beacon; MB) was evaluated in a real-time polymerase chain reaction (PCR) assay to detect the presence of Salmonella species. As few as 1-4 colony-forming units (CFU) per PCR reaction could be detected. The capability of the assay to detect Salmonella species from artificially inoculated fresh-cut produce such as cantaloupe, mixed-salad, cilantro, and alfalfa sprouts was demonstrated. A comparison of two commercially available kits utilizing MB-PCR (iQ-Check, Bio-Rad Laboratories) and conventional Association of Official Analytical Chemists (AOAC)-approved PCR (BAX, Dupont Qualicon) was performed on artificially inoculated produce. As few as 4 CFU/25 g of produce were detected after 16 h of enrichment in buffered peptone broth. These assays could be carried out entirely in sealed PCR tubes, enabling a rapid and high-throughput detection of Salmonella species in a large number of food and environmental samples. This is the first report of the application of MB probe being used for real-time detection of Salmonella species in whole and fresh-cut fruits and vegetables. PMID- 15282131 TI - Factors affecting production of extracellular carbohydrate complexes by Escherichia coli O157:H7. AB - Production of extracellular carbohydrate complexes (ECC) by foodborne pathogens on raw fruits and vegetables may result in protection against removal or inactivation by sanitizers. The influence of environmental conditions on cell growth, the total amount of ECC produced, and the amount of ECC produced on a per cell basis by Escherichia coli O157:H7 strains ATCC 43895 (wild type) and 43895 exopolysaccharides (EPS) (natural mutant, extensive EPS producer) was studied. To determine the effects of pH on the production of ECC on a per cell basis, E. coli O157:H7 was grown aerobically at 12 and 22 degrees C on tryptic soy agar (TSA) acidified to pH 7.0, 6.5, 6.0, 5.5, 5.0, 4.5, and 4.0. Lettuce, alfalfa sprout, cantaloupe, tomato, and apple juice agars (pH 4.46-6.50) were also evaluated for their support of the ECC production. Conditions generally favorable for growth of E. coli O157:H7 were a rich nutrient medium (TSA) vs. heated lettuce juice agar (HLJA) or minimal salts medium (MSM), 22 degrees C compared to 12 degrees C, and an aerobic atmosphere compared to modified atmosphere (1% O(2), 10% CO(2), and 89% N(2)). Conditions favorable for production of ECC on a per cell basis were HLJA, 12 degrees C, and an aerobic atmosphere. This suggests that modified atmosphere packaging of lettuce may not only decrease the growth of E. coli O157:H7 but also its propensity to form biofilm. There was a negative relationship between cell growth and production of ECC on a per cell basis, and environmental conditions that affected the total amount of ECC produced based on initial population reflected a combination of environmental conditions influencing both cell growth and ECC production on a per cell basis. A relative growth index factor (RGIF) was calculated to better understand ECC production as affected by various environmental conditions simultaneously. The production of ECC on a per cell basis by strain 43895-EPS showed a negative linear relationship with pH of TSA at both 12 and 22 degrees C. This strain generally produced a greater amount of ECC on fresh juice agar than on TSA at the same pH, but production of ECC on alfalfa sprout juice agar (FJA, pH 6.45) at 22 degrees C was significantly less than on TSA (pH 6.50). This indicates that nutrient limitation is not based only on nutrient availability. There may be other factors that repress the production of ECC on FJA, and the effects of those factors may be temperature dependent. Further studies will be required to better understand the relationship between nutrient availability and other factors on the production of ECC by E. coli O157:H7 on raw produce. PMID- 15282132 TI - Estimation of the non-isothermal inactivation patterns of Bacillus sporothermodurans IC4 spores in soups from their isothermal survival data. AB - The isothermal survival curves of the heat resistant spores of Bacillus sporothermodurans IC4, in the range of 117-125 degrees C, were determined in chicken, mushroom and pea soups by the capillary method. They were all non-linear with a noticeable upper concavity and could be described by the equation log(10) [N(t)/N(0)]=-b(T)t(n) with a fixed power, n, of the order of 0.7-0.8. The temperature dependence of b(T) could be described by the equation b(T)=log(e)[1+exp[k(T-T(c))]], where T(c) is the temperature where intensive inactivation starts and k is the slope of b(T) at temperatures well above T(c). They were 121-123 degrees C and 0.2-0.4 degrees C(-1), respectively, depending on the soup. These parameters were used to estimate the survival curves of the spores in two non-isothermal heat treatments using the procedure originally proposed by Peleg and Penchina [Crit. Rev. Food Sci. Nutr. 40 (2000) 159]. The results were compared with experimental survival curves, determined by the direct injection method, in another laboratory. There was a general agreement, although not perfect, between the predicted and experimentally observed survival ratios. Also, the isothermal survival parameters, estimated directly from the non isothermal inactivation data using the model, were in general agreement with those calculated from the isothermal data. This suggests that the heat inactivation patterns of B. sporothermodurans IC4 spores in soups can be at least roughly estimated using the same general survival model, which has until now only been experimentally validated for vegetative bacterial cells. PMID- 15282133 TI - Lactobacillus reuteri in bovine milk fermented decreases the oral carriage of mutans streptococci. AB - The effect of Lactobacillus reuteri against one of the major cariogenic organism, Streptococcus mutans, was studied. Yogurt products containing L.reuteri showed a significant growth inhibitory effect against S. mutans, whilst yoghurts with lactobaccilli other than L. reuteri did not show such inhibition. Further, double blind, placebo-controlled trial demonstrated that consuming yogurt with L. reuteri significantly reduced the oral carriage of mutans streptococci, compared with the placebo yogurt. PMID- 15282134 TI - Antimicrobial activity of reuterin in combination with nisin against food-borne pathogens. AB - Antimicrobial activity of reuterin individually or in combination with nisin against different food-borne Gram-positive and Gram-negative pathogens in milk was investigated. Reuterin (8 AU/ml) exhibited bacteriostatic activity against Listeria monocytogenes, whereas its activity was slightly bactericidal against Staphylococcus aureus at 37 degrees C. Higher bactericidal activity was detected against Escherichia coli O157:H7, Salmonella choleraesuis subsp. choleraesuis, Yersinia enterocolitica, Aeromonas hydrophila subsp. hydrophila and Campylobacter jejuni. A significant synergistic effect on L. monocytogenes and a slight additive effect on S. aureus after 24 h at 37 degrees C were observed when reuterin was combined with nisin (100 IU/ml). The combination of reuterin with nisin did not enhance the antimicrobial effect of reuterin against Gram-negative pathogens. PMID- 15282135 TI - Estimation of accordance and concordance in inter-laboratory trials of analytical methods with qualitative results. AB - In this short communication, we discuss alternative ways to estimate accordance and concordance, which are, for qualitative methods, the analogues of the well known concepts repeatability and reproducibility. We argue that estimators in a random framework appear to be more appropriate than the estimators in a fixed framework as they were originally introduced by Langton et al. [International Journal of Food Microbiology 79 (2002) 171]. A simple example of their application is given. PMID- 15282136 TI - The bisphosphonate pamidronate on the surface of titanium stimulates bone formation around tibial implants in rats. AB - Many materials with differing surfaces have been developed for clinical implant therapy in dentistry and orthopedics. We analyzed the quantity of new bone formed in vivo around calcium-immobilized titanium implants with surfaces modified using pamidronate (PAM), a nitrogen-containing bisphosphonate (N-BP), implants of pure titanium, and titanium implants immobilized with calcium ions. New bone formation was visualized using fluorescent labeling (calcein blue and alizarin complexone) with intravenous injection at 1 and 3 weeks after implantation. After 4 weeks, undecalcified sections were prepared, and new bone formation around the implants was examined by morphometry using confocal laser scanning microscopy images. After 1 week, more new bone formed around the PAM-immobilized implant than around the calcium-immobilized and pure titanium implants. This was also seen with the new bone formation after 3 weeks. After 4 weeks, significantly more new bones were formed around the BP-immobilized implant than around the calcium ion implanted and pure titanium implants. The new N-BP-modified titanium surface stimulates new bone formation around the implant, which might contribute to the success of implant therapy. PMID- 15282137 TI - Tethering poly(ethylene glycol)s to improve the surface biocompatibility of poly(acrylonitrile-co-maleic acid) asymmetric membranes. AB - To improve the surface biocompatibility, asymmetric membranes fabricated from poly(acrylonitrile-co-maleic acid)s (PANCMAs) synthesized by water-phase precipitation copolymerization were tethered (or immobilized) with poly(ethylene glycol)s (PEGs) by esterification reaction. Chemical changes on the membrane surface were characterized by Fourier transform infrared spectroscopy and elemental analysis to confirm the immobilization of PEG onto the PANCMA membranes. The hydrophilicity and blood compatibility of the PEG-tethered PANCMA membrane were investigated by water contact angle, water absorption, protein adsorption, plasma platelets adhesion and cell adhesion measurements, and the results were compared with the corresponding PANCMA membranes. It was found that, after the tethering of PEG, the hydrophilicity of the membrane can be improved significantly, and the protein adsorption, platelets adhesion and macrophage attachment on the membrane surface are obviously suppressed. Furthermore, not only the content of maleic acid in PANCMA, which influences the tethering density of PEG, but also the molecular weight of PEG has great effect on the surface modification of PANCMA membranes for biocompatibility. PMID- 15282138 TI - A three-dimensional nanofibrous scaffold for cartilage tissue engineering using human mesenchymal stem cells. AB - The utilization of adult stem cells in tissue engineering is a promising solution to the problem of tissue or organ shortage. Adult bone marrow derived mesenchymal stem cells (MSCs) are undifferentiated, multipotential cells which are capable of giving rise to chondrocytes when maintained in a three-dimensional culture and treated with members of the transforming growth factor-beta (TGF-beta) family of growth factors. In this study, we fabricated a nanofibrous scaffold (NFS) made of a synthetic biodegradable polymer, poly(-caprolactone) (PCL), and examined its ability to support in vitro chondrogenesis of MSCs. The electrospun PCL porous scaffold was constructed of uniform, randomly oriented nanofibers with a diameter of 700 nm, and structural integrity of this scaffold was maintained over a 21-day culture period. MSCs cultured in NFSs in the presence of TGF-beta1 differentiated to a chondrocytic phenotype, as evidenced by chondrocyte-specific gene expression and synthesis of cartilage-associated extracellular matrix (ECM) proteins. The level of chondrogenesis observed in MSCs seeded within NFSs was comparable to that observed for MSCs maintained as cell aggregates or pellets, a widely used culture protocol for studying chondrogenesis of MSCs in vitro. Due to the physical nature and improved mechanical properties of NFSs, particularly in comparison to cell pellets, the findings reported here suggest that the PCL NFS is a practical carrier for MSC transplantation, and represents a candidate scaffold for cell-based tissue engineering approaches to cartilage repair. PMID- 15282139 TI - Feasibility of chitosan-based hyaluronic acid hybrid biomaterial for a novel scaffold in cartilage tissue engineering. AB - In this study, we hypothesized that hyaluronic acid could provide superior biological effects on the chondrocytes in a three-dimensional culture system. To test this hypothesis, we investigated the in vitro behavior of rabbit chondrocytes on a novel chitosan-based hyaluronic acid hybrid polymer fiber. The goal of the current study was to show the superiority of this novel fiber as a scaffold biomaterial for cartilage tissue engineering. Chitosan polymer fibers (chitosan group) and chitosan-based hyaluronic acid hybrid polymer fibers (HA 0.04% and HA 0.07% groups, chitosan coated with hyaluronic acid 0.04% and 0.07%, respectively) were originally developed by the wetspinning method. Articular chondrocytes were isolated from Japanese white rabbits and cultured in the sheets consisting of each polymer fiber. The effects of each polymer fiber on cell adhesivity, proliferation, morphological changes, and synthesis of the extracellular matrix were analyzed by quantitative a cell attachment test, DNA quantification, light and scanning electron microscopy, semi-quantitative RT-PCR, and immunohistochemical analysis. Cell adhesivity, proliferation and the synthesis of aggrecan were significantly higher in the hybrid fiber (HA 0.04% and 0.07%) groups than in the chitosan group. On the cultured hybrid polymer materials, scanning electron microscopic observation showed that chondrocytes proliferated while maintaining their morphological phenotype and with a rich extracellular matrix synthesis around the cells. Immunohistochemical staining with an anti-type II collagen antibody demonstrated rich production of the type II collagen in the pericellular matrix from the chondrocytes. The chitosan-based hyaluronic acid hybrid polymer fibers show great potential as a desirable biomaterial for cartilaginous tissue scaffolds. PMID- 15282140 TI - Analysis and evaluation of a biomedical polycarbonate urethane tested in an in vitro study and an ovine arthroplasty model. Part I: materials selection and evaluation. AB - The polyurethane elastomer (PU) Corethane 80A (Corvita) is being considered as the acetabular bearing material in a novel total replacement hip joint. The biostability of Corethane 80A was investigated in vitro (this work) and in vivo (reported separately) in a fully functioning ovine total hip arthroplasty (THA) model, with the PU as the bearing layer in a prototype compliant layer acetabular cup. The in vitro studies assessed the resistance of Corethane 80A to the main degradation mechanisms observed in PUs: hydrolysis, environmental stress cracking (ESC), metal ion oxidation (MIO) and calcification. The performance of the polycarbonate PU Corethane 80A was assessed alongside three other commercially available biomedical PUs: polyether PUs Pellethane 2363-80A (DOW Chemical) and PHMO-PU (CSIRO, not supplied as a commercial material) as well as polycarbonate PU ChronoFlex AL-80A (CardioTech). Chemical and structural variables that affect the properties of the materials were analysed with particular attention to the nature of the material's hard and soft segments. PU degradation was probed using a range of analytical tools and physical-testing methods, including mechanical testing, differential scanning calorimetry (DSC), Fourier transform infrared spectroscopy (FTIR) and environmental scanning microscopy (ESEM). Corethane 80A displayed the best overall resistance to hydrolysis, ESC, MIO and calcification, followed by ChronoFlex 80A and PHMO-PU. Pellethane 80A was the least stable. This study provides compelling evidence for the biostability and effectiveness of Corethane 80A and points to its suitability for use as a compliant bearing layer in hip arthroplasty, and possibly also other joints. PMID- 15282141 TI - Analysis and evaluation of a biomedical polycarbonate urethane tested in an in vitro study and an ovine arthroplasty model. Part II: in vivo investigation. AB - The polyurethane (PU) elastomer Corethane 80A (Corvita) is being considered as the acetabular bearing material in a novel total replacement hip joint. Its biostability was investigated in vitro (Analysis and evaluation of a biomedical polycarbonate urethane tested in an in vitro study and an ovine arthroplasty model. Part I: material selection and evaluation, Biomaterials, in press) together with three other commercially available biomedical PUs: Pellethane 2363 80A (DOW Chemical), a polyhexamethylene oxide based PU, PHMO-PU (CSIRO, not supplied as a commercial product) and ChronoFlex AL-80A (CardioTech). From the in vitro studies, Corethane 80A displayed the best overall resistance to hydrolysis, ESC, MIO and calcification, followed by ChronoFlex 80A and PHMO-PU, with Pellethane 80A being the least stable. Building on the in vitro investigation, the follow-up in vivo study (reported here) assessed Corethane 80A as the bearing layer in a prototype compliant layer acetabular cup, in a fully functioning ovine total hip arthoplasty (THA) model. PU degradation in the retrieved cups was analysed using a range of analytical and physical-testing methods including mechanical testing, differential scanning calorimetry, Fourier transform infrared spectroscopy and environmental scanning electron microscopy. The Corethane 80A functioned well in the THA model, with the bearing surfaces of the retrieved hip cups showing no significant evidence of biodegradation or wear damage after 3 years in vivo. The findings in this study provide compelling evidence for the biostability and effectiveness of acetabular cups incorporating a Corethane 80A compliant bearing layer. PMID- 15282142 TI - Fixation of distal femoral osteotomies with self-reinforced polymer/bioactive glass rods: an experimental study on rabbits. AB - Two self-reinforced poly(desamino tyrosyl-tyrosine ethyl ester carbonate), poly(DTE carbonate) or self-reinforced poly(DTE carbonate)/bioactive glass rods, (2 mm by 40 mm) were implanted into the dorsal subcutaneous tissue and osteotomies of the distal femur were fixed with these rods (2 mm by 26 mm) in 36 rabbits. The follow-up times varied from three to 100 weeks. After sacrifice, three-point bending and shear tests and molecular weight measurements were performed for subcutaneously placed rods. Radiological, histological, histomorphometrical, microradiographic, and oxytetracycline-fluorescence studies of the osteotomized and intact control femora were performed. The initial mechanical properties were higher with the SR-poly(DTE carbonate) rods, but the SR-poly(DTE carbonate)/bioactive glass rods lost their mechanical properties slower. At 100 weeks the bending strength had decreased to 21% of the initial value with the SR-poly(DTE carbonate) rods and to 49% with the SR-poly(DTE carbonate)/bioactive glass rods. The shear strength had decreased to 10% with the SR-poly(DTE carbonate) rods and to 23% of the initial value with the SR-poly(DTE carbonate)/bioactive glass rods. Two slight displacements and one delayed union and one failure of fixation were seen in the SR-poly(DTE carbonate) group. In the SR-poly(DTE carbonate)/bioactive glass group five delayed unions and seven slight displacements were seen. No signs of osteolysis or foreign body reactions were observed. Signs of resorption of the implants were seen at 100 weeks in the SR poly(DTE carbonate)/bioactive glass group. The present investigation showed that the mechanical strength and fixation properties of SR-poly(DTE carbonate) and SR poly(DTE carbonate)/bioactive glass rods are suitable for fixation of cancellous bone osteotomies in rabbits. PMID- 15282143 TI - The potential of poly(N-isopropylacrylamide) (PNIPAM)-grafted hyaluronan and PNIPAM-grafted gelatin in the control of post-surgical tissue adhesions. AB - Poly(N-isopropylacrylamide)-grafted hyaluronan (PNIPAM-HA) and PNIPAM-grafted gelatin (PNIPAM-gelatin), which exhibit sol-to-gel transformation at physiological temperature, were applied as control of tissue adhesions: tissue adhesion prevention material and hemostatic aid, respectively. The rat cecum, which was abraded using surgical gauze, was coated with PNIPAM-HA-containing PBS (concentration: 0.5 w/v%). The coated solution was immediately converted to an opaque precipitate at body temperature, which weakly adhered to and covered the injured rat cecum. One week after coating, tissue adhesion between the PNIPAM-HA treated cecum and adjacent tissues was significantly reduced as compared with that between non-treated tissue and adjacent tissues. On the other hand, the coating of bleeding spots of a canine liver with PNIPAM-gelatin-containing PBS (concentration: 20 w/v%) resulted in spontaneous gel formation on the tissues and subsequently suppressed bleeding. Although these thermoresponsive tissue adhesion prevention and hemostatic materials are still prototypes at this time, both thermoresponsive biomacromolecules bioconjugated with PNIPAM, PNIPAM-HA and PNIPAM-gelatin, may serve as a tissue adhesion prevention material and hemostatic aid, respectively. PMID- 15282144 TI - Characterization of biodegradable drug delivery vehicles with the adhesive properties of leukocytes II: effect of degradation on targeting activity. AB - The site-specific expression of selectins (P- and E-selectin) on endothelial cells of blood vessels during inflammation provides an opportunity for the targeted delivery of anti-inflammatory drugs to sites of chronic inflammation. It is well documented that the selectins mediate the initial interaction (rolling) of leukocytes in an inflamed vessel by binding to carbohydrate-presenting counter receptors displayed on leukocytes. Previous work in our laboratory has shown that artificial capsules with the adhesive properties of leukocytes can be made by attaching leukocyte adhesive ligands to polymer microspheres (Biomaterials 23(10) (2002) 2167). Specifically, we showed that drug-loaded poly (lactic-co-glycolic acid) (PLGA) microspheres coated with biotinylated-Sialyl LewisX (sLeX), a carbohydrate that serves as a ligand to selectins, mimic the adhesive behavior of leukocytes on selectins in flow chambers, displaying slow rolling under flow, suggesting that these drug-loaded particles can potentially target inflammatory sites in vivo. Since the effectiveness of this delivery system might depend on the degradation of polymer microspheres as well as the degradation of sLeX molecules, we measured the effect of polymer and ligand degradation on the adhesiveness of microspheres over time. We show that degrading sLeX microspheres maintain the ability to recognize selectin surfaces under flow for at least 2 weeks and that the ability to sustain recognition depends upon the extent at which microspheres are loaded. We also show that microsphere rolling velocity increases as microsphere degrade and that this increase is due to a combination of increase in average microsphere size and loss of sLeX molecules on microsphere surface--a result of microsphere degradation confirmed by flow cytometry. Control experiments show that microsphere, not sLeX, degradation limits the lifetime of our targeted delivery system; therefore, factors affecting degradation such as type of polymer, type of drug, extent of drug loading and microsphere size, provide an opportunity for engineering the time-scale of activity for the delivery system. PMID- 15282145 TI - Tissue anti-adhesion potential of ibuprofen-loaded PLLA-PEG diblock copolymer films. AB - This study was designed to evaluate the effect of polyethylene glycol (PEG) and nonsteroidal anti-inflammatory drug (ibuprofen) on the prevention of postsurgical tissue adhesion. For this, poly(L-lactic acid) (PLLA)-PEG diblock copolymers were synthesized by ring opening polymerization of L-lactide and methoxy polyethylene glycol (Mw 5000) of different compositions. The synthesized copolymers were characterized by gel permeation chromatography and 1H-nuclear magnetic resonance spectroscopy. PLLA-PEG copolymer films were prepared by solvent casting. The prepared copolymer films were more flexible and hydrophilic than the control PLLA film, as investigated by the measurements of glass transition temperature, water absorption content, and water contact angle. The drug release behavior from the ibuprofen (10 wt%)-loaded copolymer films was examined by high performance liquid chromatography. It was observed that the drug was released gradually up to about 40% of total loading amount after 20 days, depending on PEG composition; more drug release from the films with higher PEG compositions. In vitro cell adhesions on the copolymer films with/without drug were compared by the culture of NIH/3T3 mouse embryo fibroblasts on the surfaces. For in vivo evaluation of tissue anti adhesion potential, the copolymer films with/without drug were implanted between the cecum and peritoneal wall defects of rats and their tissue adhesion extents were compared. It was observed that the ibuprofen-containing PLLA-PEG films with high PEG composition (particularly PLLA113-PEG113 film with PEG composition, 50 mol%) were very effective in preventing cell or tissue adhesion on the film surfaces, probably owing to the synergistic effects of highly mobile, hydrophilic PEG and anti-inflammatory drug, ibuprofen. PMID- 15282146 TI - Enhanced gene delivery to PC12 cells by a cationic polypeptide. AB - Targeted gene delivery to diseased subtypes of neurons will be beneficial to the success of gene therapy of neurological disorders. We designed a recombinant cationic polypeptide to facilitate gene delivery to neuronal-like PC12 cells that express the nerve growth factor (NGF) receptors. The recombinant polypeptide was composed of a targeting moiety derived from loop 4-containing hairpin motif of NGF and a DNA-binding moiety of 10-lysine sequence and expressed in Escherichia coli. It activated NGF receptor, TrkA and its downstream signaling pathways in PC12 and promoted the survival of neuronally differentiated PC12 cells deprived of serum. The polypeptide could also bind plasmid DNA and enhance polycation mediated gene delivery in NGF receptor-expressing PC12 cells, but not in COS7 cells lacking NGF receptors. The enhancement of gene transfer in PC12 was inhibited by pretreatment of free, unbound polypeptides, suggesting a NGF receptor-specific effect of the polypeptide. These observations demonstrated the concept of using receptor-mediated mechanism for targeted gene delivery to neurons. PMID- 15282147 TI - Antibody microarray for correlating cell phenotype with surface marker. AB - To correlate cell surface markers with the cell phenotype, an antibody microarray prepared by covalently immobilizing antibodies onto a cellulose membrane and subsequent immunocytochemical staining were employed. The direct binding assay of a lymphoblastic leukemia cell line on the microarray showed that the immobilized antibody served to capture cells expressing the specific antigen. The density of bound cells increased linearly with an increasing content of antigen-expressing cells in suspension. The method was further applied to the analysis of surface antigens expressed on neural stem cells. A binding assay was performed with neural cells obtained from the neurosphere culture of the rat fetal striatum on a microarray spotted with eight kinds of antibodies and four different proteins, followed by immunocytochemical staining of cells bound to the microarray using antibodies to the intracellular markers of immature (nestin and vimentin) and mature (beta-tubulin III and glial fibrillary acidic protein) neural cells. As a result, the phenotype of bound cells could be correlated to surface antigen expression, which illustrated the potential of the solid-phase cytometry developed here for the identification of surface markers. PMID- 15282148 TI - Heart fields: one, two or more? PMID- 15282149 TI - Apoptosis and involution in the mammary gland are altered in mice lacking a novel receptor, beta1,4-Galactosyltransferase I. AB - Receptor-mediated cell-extracellular matrix (ECM) interactions are critical regulators of cell survival, and perturbing these signaling pathways can disrupt cellular differentiation and function in a variety of tissues, including the mammary gland. One such receptor is the cell surface-associated, long isoform of beta1,4-galactosyltransferase I (GalT I). Deletion of long GalT I leads to increased mammary ductal branching morphogenesis [Dev. Biol., 244 (2002) 114]. Here, we show that this expansion in the mammary epithelial (ME) cell compartment is accomplished through decreased apoptosis during pregnancy and involution. Decreased apoptosis during involution is concomitant with delayed alveolar collapse, persistent expression of the milk protein gene alpha-lactalbumin and delayed expression of genes associated with the tissue-remodeling phase of involution. Using 3-dimensional in vitro cultures, we show that the decrease in apoptosis is dependent on laminin 1, a ligand for surface GalT I, suggesting that surface GalT I negatively influences ECM-dependent cell survival, a novel function for an ECM receptor. In the best-studied examples, ECM promotes survival through integrin receptor-mediated activation of focal adhesion kinase (FAK). Aggregation of surface GalT I also activates FAK, therefore, we asked if FAK activation was altered in ME from long GalT I null mice. Activated FAK was appropriately localized to focal adhesions in long GalT I null ME. However, FAK activation was constitutively reduced 4.5-fold in long GalT I nulls relative to wild type. Expression of the integrin beta1 subunit was not affected by loss of long GalT I. Collectively, these results suggest that surface GalT I might negatively regulate ME cell survival by linking integrin-independent FAK activation to apoptotic rather than survival signaling events. PMID- 15282150 TI - Regulation of ureteric bud branching morphogenesis by sulfated proteoglycans in the developing kidney. AB - Glycosaminoglycans in the form of heparan sulfate proteoglycans (HSPG) and chondroitin sulfate proteoglycans (CSPG) are required for normal kidney organogenesis. The specific roles of HSPGs and CSPGs on ureteric bud (UB) branching morphogenesis are unclear, and past reports have obtained differing results. Here we employ in vitro systems, including isolated UB culture, to clarify the roles of HSPGs and CSPGs on this process. Microarray analysis revealed that many proteoglycan core proteins change during kidney development (syndecan-1,2,4, glypican-1,2,3, versican, decorin, biglycan). Moreover, syndecan 1, syndecan-4, glypican-3, and versican are differentially expressed during isolated UB culture, while decorin is dynamically regulated in cultured isolated metanephric mesenchyme (MM). Biochemical analysis indicated that while both heparan sulfate (HS) and chondroitin sulfate (CS) are present, CS accounts for approximately 75% of the glycosaminoglycans (GAG) in the embryonic kidney. Selective perturbation of HS in whole kidney rudiments and in the isolated UB resulted in a significant reduction in the number of UB branch tips, while CS perturbation has much less impressive effects on branching morphogenesis. Disruption of endogenous HS sulfation with chlorate resulted in diminished FGF2 binding and proliferation, which markedly altered kidney area but did not have a statistically significant effect on patterning of the ureteric tree. Furthermore, perturbation of GAGs did not have a detectable effect on FGFR2 expression or epithelial marker localization, suggesting the expression of these molecules is largely independent of HS function. Taken together, the data suggests that nonselective perturbation of HSPG function results in a general proliferation defect; selective perturbation of specific core proteins and/or GAG microstructure may result in branching pattern defects. Despite CS being the major GAG synthesized in the whole developing kidney, it appears to play a lesser role in UB branching; however, CS is likely to be integral to other developmental processes during nephrogenesis, possibly involving the MM. A model is presented of how, together with growth factors, heterogeneity of proteoglycan core proteins and glycosaminoglycan sulfation act as a switching mechanism to regulate different stages of the branching process. In this model, specific growth factor HSPG combinations play key roles in the transitioning between stages and their maintenance. PMID- 15282151 TI - Myosin VI is required for structural integrity of the apical surface of sensory hair cells in zebrafish. AB - Unconventional myosins have been associated with hearing loss in humans, mice, and zebrafish. Mutations in myosin VI cause both recessive and dominant forms of nonsyndromic deafness in humans and deafness in Snell's waltzer mice associated with abnormal fusion of hair cell stereocilia. Although myosin VI has been implicated in diverse cellular processes such as vesicle trafficking and epithelial morphogenesis, the role of this protein in the sensory hair cells remains unclear. To investigate the function of myosin VI in zebrafish, we cloned and examined the expression pattern of myosin VI, which is duplicated in the zebrafish genome. One duplicate, myo6a, is expressed in a ubiquitous pattern during early development and at later stages, and is highly expressed in the brain, gut, and kidney. myo6b, on the other hand, is predominantly expressed in the sensory epithelium of the ear and lateral line at all developmental stages examined. Both molecules have different splice variants expressed in these tissues. Using a candidate gene approach, we show that myo6b is satellite, a gene responsible for auditory/vestibular defects in zebrafish larvae. Examination of hair cells in satellite mutants revealed that stereociliary bundles are irregular and disorganized. At the ultrastructural level, we observed that the apical surface of satellite mutant hair cells abnormally protrudes above the epithelium and the membrane near the base of the stereocilia is raised. At later stages, stereocilia fused together. We conclude that zebrafish myo6b is required for maintaining the integrity of the apical surface of hair cells, suggesting a conserved role for myosin VI in regulation of actin-based interactions with the plasma membrane. PMID- 15282152 TI - The role of mechanical forces in dextral rotation during cardiac looping in the chick embryo. AB - Cardiac looping is a vital morphogenetic process that transforms the initially straight heart tube into a curved tube normally directed toward the right side of the embryo. While recent work has brought major advances in our understanding of the genetic and molecular pathways involved in looping, the biophysical mechanisms that drive this process have remained poorly understood. This paper examines the role of biomechanical forces in cardiac rotation during the initial stages of looping, when the heart bends and rotates into a c-shaped tube (c looping). Embryonic chick hearts were subjected to mechanical and chemical perturbations, and tissue stress and strain were studied using dissection and fluorescent labeling, respectively. The results suggest that (1) the heart contains little or no intrinsic ability to rotate, as external forces exerted by the splanchnopleure (SPL) and the omphalomesenteric veins (OVs) drive rotation; (2) unbalanced forces in the omphalomesenteric veins play a role in left-right looping directionality; and (3) in addition to ventral bending and rightward rotation, the heart tube also bends slightly toward the right. The results of this study may help investigators searching for the link between gene expression and the mechanical processes that drive looping. PMID- 15282153 TI - Primordial germ cell migration in the chick and mouse embryo: the role of the chemokine SDF-1/CXCL12. AB - As in many other animals, the primordial germ cells (PGCs) in avian and reptile embryos are specified in positions distinct from the positions where they differentiate into sperm and egg. Unlike in other organism however, in these embryos, the PGCs use the vascular system as a vehicle to transport them to the region of the gonad where they exit the blood vessels and reach their target. To determine the molecular mechanisms governing PGC migration in these species, we have investigated the role of the chemokine stromal cell-derived factor-1 (SDF 1/CXCL12) in guiding the cells towards their target in the chick embryo. We show that sdf-1 mRNA is expressed in locations where PGCs are found and towards which they migrate at the time they leave the blood vessels. Ectopically expressed chicken SDF-1alpha led to accumulation of PGCs at those positions. This analysis, as well as analysis of gene expression and PGC behavior in the mouse embryo, suggest that in both organisms, SDF-1 functions during the second phase of PGC migration, and not at earlier phases. These findings suggest that SDF-1 is required for the PGCs to execute the final migration steps as they transmigrate through the blood vessel endothelium of the chick or the gut epithelium of the mouse. PMID- 15282154 TI - The Drosophila Wnt5 protein mediates selective axon fasciculation in the embryonic central nervous system. AB - The decision of whether and where to cross the midline, an evolutionarily conserved line of bilateral symmetry in the central nervous system, is the first task for many newly extending axons. We show that Wnt5, a member of the conserved Wnt secreted glycoprotein family, is required for the formation of the anterior of the two midline-crossing commissures present in each Drosophila hemisegment. Initial path finding of pioneering neurons across the midline in both commissures is normal in wnt5 mutant embryos; however, the subsequent separation of the early midline-crossing axons into two distinct commissures does not occur. The majority of the follower axons that normally cross the midline in the anterior commissure fail to do so, remaining tightly associated near their cell bodies, or projecting inappropriately across the midline in between the commissures. The lateral and intermediate longitudinal pathways also fail to form correctly, similarly reflecting earlier failures in pathway defasciculation. Panneural expression of Wnt5 in a wnt5 mutant background rescues both the commissural and longitudinal defects. We show that the Wnt5 protein is predominantly present on posterior commissural axons and at a low level on the anterior commissure and longitudinal projections. Finally, we demonstrate that transcriptional repression of wnt5 in AC neurons by the recently described Wnt5 receptor, Derailed, contributes to this largely posterior commissural localization of Wnt5 protein. PMID- 15282155 TI - A sea urchin egg jelly peptide induces a cGMP-mediated decrease in sperm intracellular Ca(2+) before its increase. AB - Speract, a sperm-activating peptide (SAP) from sea urchin eggs, increases the intracellular concentration of Ca(2+) ([Ca(2+)]i) and modulates sperm motility. We measured the initial sperm response to speract using its caged analog and observed, for the first time, a small but significant decrease in sperm [Ca(2+)]i before the increase. Both directions of the [Ca(2+)]i change were completely blocked in high K(+) seawater. Using membrane-permeant caged cyclic nucleotides (cNMP), only cGMP induced the decrease in [Ca(2+)]i although both cGMP and cAMP increased the [Ca(2+)]i. The decrease in the [Ca(2+)]i induced by cGMP was more notable following a second photolytic event, once [Ca(2+)]i had been elevated by an initial flash. This pattern of [Ca(2+)]i change was confirmed in individual sperm. These results together with pharmacological evidence suggest that the initial [Ca(2+)]i decrease is due to a Na(+)/Ca(2+) exchanger activity, stimulated by hyperpolarization mediated by K(+) efflux through cGMP-regulated K(+) channels. PMID- 15282156 TI - UNC-39, the C. elegans homolog of the human myotonic dystrophy-associated homeodomain protein Six5, regulates cell motility and differentiation. AB - Mutations in the unc-39 gene of C. elegans lead to migration and differentiation defects in a subset of mesodermal and ectodermal cells, including muscles and neurons. Defects include mesodermal specification and differentiation as well a neuronal migration and axon pathfinding defects. Molecular analysis revealed that unc-39 corresponds to the previously named gene ceh-35 and that the UNC-39 protein belongs to the Six4/5 family of homeodomain transcription factors and is similar to human Six5, a protein implicated in the pathogenesis of type I myotonic dystrophy (DM1). We show that human Six5 and UNC-39 are functional homologs, suggesting that further characterization of the C. elegans unc-39 gene might provide insight into the etiology of DM1. PMID- 15282157 TI - pha-2 encodes the C. elegans ortholog of the homeodomain protein HEX and is required for the formation of the pharyngeal isthmus. AB - The pha-2 mutant was isolated in 1993 by Leon Avery in a screen for worms with visible defects in pharyngeal feeding behavior. In pha-2 mutant worms, the pharyngeal isthmus is abnormally thick and short and, in contrast to wild-type worms, harbors several cell nuclei. We show here that pha-2 encodes a homeodomain protein and is homologous to the vertebrate homeobox gene, Hex (also known as Prh). Consistent with a function in pharyngeal development, the pha-2 gene is expressed in the pharyngeal primordium of Caenorhabditis elegans embryos, particularly in pm5 cells that form the bulk of the isthmus. We show that in the pha-2 mutant there is a failure of the pm5 cells to elongate anteriorly while keeping their nuclei within the nascent posterior bulb to form the isthmus during the 3-fold embryonic stage. We also present evidence that pha-2 regulates itself positively in pm5 cells, that it is a downstream target of the forkhead gene pha 4, and that it may also act in the isthmus as an inhibitor of the ceh-22 gene, an Nkx2.5 homolog. Finally, we have begun characterizing the regulation of the pha-2 gene and find that intronic sequences are essential for the complete pha-2 expression profile. The present report is the first to examine the expression and function of an invertebrate Hex homolog, that is, the C. elegans pha-2 gene. PMID- 15282158 TI - Drosophila cardiac tube organogenesis requires multiple phases of Hox activity. AB - The segmented Drosophila linear cardiac tube originates from two cell lineages that give rise to the anterior aorta (AA) and the posterior cardiac tube. The three Hox genes of the Bithorax Complex as well as Antennapedia (Antp) have been shown to be expressed in the posterior cardiac tube, while no Hox gene is expressed in the anterior aorta. We show that the cells of the whole tube adopt the anterior aorta identity in the complete absence of Hox function. Conversely, ectopic expression of Antp, Ultrabithorax (Ubx), or abdominal-A (abd-A) transformed the anterior aorta into posterior cardiac tube by all available criteria, indicating an equivalent early function in their ability to direct a posterior cardiac tube lineage. We further demonstrate that Hox genes act in a subsequent step during cardiac tube organogenesis, specifically on the differentiation of posterior cardiac tube myocytes. In addition, while some of these functions are fulfilled equally well by any one of the three Hox genes, some others are specific to a given Hox. Notably, the gene encoding the anion transporter Na+-Driven Anion Exchanger 1 behaves as a Hox differential transcriptional target and is activated by abd-A in the heart and repressed by Ubx in the posterior aorta. This analysis illustrates the mechanisms by which Hox genes can orchestrate organogenesis and, in particular, allows a clear uncoupling of the different phases of Hox activity in this process. PMID- 15282159 TI - In vitro growth and differentiation of mammalian sensory hair cell progenitors: a requirement for EGF and periotic mesenchyme. AB - The sensory hair cells and supporting cells of the organ of Corti are generated by a precise program of coordinated cell division and differentiation. Since no regeneration occurs in the mature organ of Corti, loss of hair cells leads to deafness. To investigate the molecular basis of hair cell differentiation and their lack of regeneration, we have established a dissociated cell culture system in which sensory hair cells and supporting cells can be generated from mitotic precursors. By incorporating a Math1-GFP transgene expressed exclusively in hair cells, we have used this system to characterize the conditions required for the growth and differentiation of hair cells in culture. These conditions include a requirement for epidermal growth factor, as well as the presence of periotic mesenchymal cells. Lastly, we show that early postnatal cochlear tissue also contains cells that can divide and generate new sensory hair cells in vitro. PMID- 15282160 TI - Functional domains and temperature-sensitive mutations in SPE-9, an EGF repeat containing protein required for fertility in Caenorhabditis elegans. AB - The spe-9 gene is required for fertility in Caenorhabditis elegans and encodes a sperm transmembrane protein with an extracellular domain (ECD) that contains 10 epidermal growth factor (EGF) repeats. Deletion analysis reveals that the EGF repeats and the transmembrane domain are required for fertilization. In contrast, the cytoplasmic region of SPE-9 is not essential for fertilization. Individual point mutations in all 10 EGF motifs uncover a differential sensitivity of these sequences to alteration. Some EGF repeats cannot tolerate mutation leading to a complete lack of fertility. Other EGF repeats can be mutated to create animals with temperature-sensitive (ts) fertility phenotypes. All ts mutations were generated by changing either conserved cysteine or glycine residues in the EGF motifs. For two endogenous ts alleles of spe-9, loss of function at nonpermissive temperatures is not due to protein mislocalization or degradation. Additionally, the proper localization of SPE-9 in sperm is not altered in a genetically interacting fertility mutant (spe-13) or a mutant that affects sperm vesicle plasma membrane fusion (fer-1). Like the EGF repeats in the Notch/LIN-12/GLP-1 receptors and their ligands, the EGF repeats in SPE-9 may carry out different functions. Because EGF motifs are found in many proteins in different species, similar experimental strategies could be used to generate useful temperature sensitive mutations in other EGF motif-containing molecules. PMID- 15282161 TI - Multiple roles for the E/Daughterless ortholog HLH-2 during C. elegans gonadogenesis. AB - HLH-2 is the Caenorhabditis elegans ortholog of the Drosophila Daughterless and mammalian E basic helix-loop-helix (bHLH) transcriptional activators that function during diverse events during animal development. HLH-2 has been implicated in cell fate specification in different neural lineages and in the LIN 12/Notch-mediated anchor cell (AC)/ventral uterine precursor cell (VU) decision in the somatic gonad. Here, we show that hlh-2 plays several distinct roles during somatic gonadogenesis. Our analysis suggests that hlh-2 is required to endow specific somatic gonadal cells with the competence to undergo the AC/VU decision, as well as functioning in the AC/VU decision per se; this novel "proAC" role appears to be analogous to the proneural role of Drosophila Daughterless. In addition to its two distinct roles in the specification of the AC, hlh-2 is also required for correct differentiation and function of the AC. hlh-2 also acts at an independent point in the gonadal lineage both to specify distal tip cells (DTCs) and in DTC differentiation and function. PMID- 15282162 TI - Cell autonomy of the mouse claw paw mutation. AB - Mice homozygous for the autosomal recessive mutation claw paw (clp) are characterized by limb posture abnormalities and congenital hypomyelination, with delayed onset of myelination of the peripheral nervous system but not the central nervous system. Although this combination of limb and peripheral nerve abnormalities in clp/clp mice might suggest a common neurogenic origin of the syndrome, it is not clear whether the clp gene acts primarily in the neurone, the Schwann cell or both. In the work described here, we address this question of cell autonomy of the clp mutation through reciprocal nerve grafting experiments between wild-type and clp/clp animals. Our results demonstrate that the clp mutation affects the Schwann cell compartment and possibly also the neuronal compartment. These data suggest that the clp gene product is expressed in Schwann cells as well as neurones and is likely to be involved in direct axon--Schwann cell interactions. Within the Schwann cell, clp affects a myelin-related signaling pathway that regulates periaxin and Krox-20 expression, but not Oct-6. PMID- 15282163 TI - Transcript profiling during preimplantation mouse development. AB - Studies using low-resolution methods to assess gene expression during preimplantation mouse development indicate that changes in gene expression either precede or occur concomitantly with the major morphological transitions, that is, conversion of the oocyte to totipotent 2-cell blastomeres, compaction, and blastocyst formation. Using microarrays, we characterized global changes in gene expression and used Expression Analysis Systematic Explorer (EASE) to identify biological and molecular processes that accompany and likely underlie these transitions. The analysis confirmed previously described processes or events, but more important, EASE revealed new insights. Response to DNA damage and DNA repair genes are overrepresented in the oocyte compared to 1-cell through blastocyst stages and may reflect the oocyte's response to selective pressures to insure genomic integrity; fertilization results in changes in the transcript profile in the 1-cell embryo that are far greater than previously recognized; and genome activation during 2-cell stage may not be as global and promiscuous as previously proposed, but rather far more selective, with genes involved in transcription and RNA processing being preferentially expressed. These results validate this hypothesis-generating approach by identifying genes involved in critical biological processes that can be the subject of a more traditional hypothesis driven approach. PMID- 15282164 TI - Identification of a critical control element directing expression of the muscle specific transcription factor MRF4 in the mouse embryo. AB - Skeletal muscle development in the vertebrate embryo critically depends on the myogenic regulatory factors (MRFs) including MRF4 and Myf5. Both genes exhibit distinct expression patterns during mouse embryogenesis, although they are genetically closely linked with multiple regulatory elements dispersed throughout the common gene locus. MRF4 has a biphasic expression profile, first in somites and later in foetal skeletal muscles. Here, we demonstrate by transgenic analysis that elements within a 7.5-kb promoter fragment of the MRF4 gene are sufficient to drive the embryonic wave of expression very similar to the endogenous gene in somites of mouse embryos. In contrast, a 3-kb fragment of the proximal promoter fails to support expression in the myotome, suggesting that essential cis-acting elements are located between -7.5 and -3 kb upstream of MRF4. Further analysis of this sequence delimits an essential region between -6.6 and -5.6 kb that together with the 3-kb promoter fragment directs transgene expression in the epaxial myotome of all somites during the appropriate developmental period. These data provide evidence that the partly overlapping expression patterns of Mrf4 and Myf5 in somites are controlled by distinct regulatory elements. We also show that 11.4 kb sequence upstream of MRF4, including the promoter and the somitic control region identified in this study, is not sufficient to elicit target specificity towards the strong Myf5 (-58/-48 kb) enhancer, suggesting that additional yet unidentified elements are necessary to convey promoter selectivity and protect the MRF4 gene from this enhancer. PMID- 15282165 TI - Differential control of MHR3 promoter activity by isoforms of the ecdysone receptor and inhibitory effects of E75A and MHR3. AB - MHR3 is an ecdysone-inducible transcription factor whose expression in both Manduca sexta epidermis and the Manduca GV1 cell line is induced by 20 hydroxyecdysone (20E) in vitro. There are four putative ecdysone response elements (EcRE) in the 2.6-kb flanking region of the MHR3 promoter. The most proximal, EcRE1, is necessary for activation of the promoter by 20E in the GV1 cells because the mutation of EcRE1 caused the loss of responsiveness to 20E. Previous studies showed that EcR-B1/USP-1 bound only to EcRE1 and high levels of this complex increased the 20E-induced activation, whereas the presence of high USP-2 prevented this increased activation. When we expressed EcR-A alone or in combination with USP-1 under the control of Autographa californica baculovirus promoter (pIE1hr), the activation of the 2.6-kb promoter by 20E was reduced by about 50%. Moreover, when EcR-A was expressed together with both EcR-B1 and USP 1, it reduced the normal activation caused by EcR-B1 and USP-1 by 50%. Gel mobility shift assays showed no binding of EcR-A/USP-1 to EcRE1. The presence of EcR-A, however, reduced the binding of EcR-B1/USP-1 by about 50%. These findings suggest that EcR-A competes with EcR-B1 for binding of USP-1, leading to a decline in activity of the promoter. In addition, E75A, another ecdysone-induced transcription factor, and MHR3 itself suppressed MHR3 promoter activity by binding to the monomeric response element (MRE2). Therefore, MHR3 can be down regulated both by itself and by E75A. PMID- 15282166 TI - Engrailed and polyhomeotic maintain posterior cell identity through cubitus interruptus regulation. AB - In Drosophila, the subdivision into compartments requires the expression of engrailed (en) and hedgehog (hh) in the posterior cells and of cubitus interruptus (ci) in the anterior cells. Whereas posterior cells express hh, only anterior cells are competent to respond to the hh signal, because of the presence of ci expression in these cells. We show here that engrailed and polyhomeotic (ph), a member of the Polycomb Group (PcG) genes, act concomitantly to maintain the repression of ci in posterior compartments during development. Using chromatin immunoprecipitation (ChIP), we identified a 1 kb genomic fragment located 4 kb upstream of the ci coding region that is responsible for the regulation of ci. This genomic fragment is bound in vivo by both Polyhomeotic and Engrailed. In particular, we show that Engrailed is responsible for the establishment of ci repression early during embryonic development and is also required, along with Polyhomeotic, to maintain the repression of ci throughout development. PMID- 15282167 TI - Identification of evolutionarily conserved promoter elements and amino acids required for function of the C. elegans beta-catenin homolog BAR-1. AB - beta-catenins are conserved transcription factors regulated posttranslationally by Wnt signaling. bar-1 encodes a Caenorhabditis elegans beta-catenin acting in multiple Wnt-mediated processes, including cell fate specification by vulval precursor cells (VPCs) and migration of the Q(L) neuroblast progeny. We took two approaches to extend our knowledge of bar-1 function. First, we undertook a bar-1 promoter analysis using transcriptional GFP reporter fusions and found that bar-1 expression is regulated in specific cells at the transcriptional level. We identified promoter elements necessary for bar-1 expression in several cell types, including a 321-bp element sufficient for expression in ventral cord neurons (VCNs) and a 1.1-kb element sufficient for expression in the developing vulva and adult seam cells. Expression of bar-1 from the 321-bp element rescued the Uncoordinated (Unc) phenotype of bar-1 mutants, but not the vulval phenotype, suggesting that a Wnt pathway may act in ventral cord neurons to mediate proper locomotion. By comparison of the 1.1-kb element to homologous sequences from Caenorhabditis briggsae, we identified evolutionarily conserved sequences necessary for expression in vulval or seam cells. Second, we analyzed 24 mutations in bar-1 and identified several residues required for BAR-1 activity in C. elegans. By phylogenetic comparison, we found that most of these residues are conserved and may identify amino acids necessary for beta-catenin function in all species. PMID- 15282168 TI - NhaA of Escherichia coli, as a model of a pH-regulated Na+/H+antiporter. AB - Na(+)/H(+) antiporters are ubiquitous membrane proteins that are involved in homeostasis of H(+) and Na(+) throughout the biological kingdom. Corroborating their role in pH homeostasis, many of the Na(+)/H(+) antiporter proteins are regulated directly by pH. The pH regulation of NhaA, the Escherichia coli Na(+)/H(+) antiporter (EcNhaA), as of other, both eukaryotic and prokaryotic Na(+)/H(+) antiporters, involves a pH sensor and conformational changes in different parts of the protein that transduce the pH signal into a change in activity. Thus, residues that affect the pH response, the translocation or both activities cluster in separate domains along the antiporter molecules. Importantly, in the NhaA family, these domains are conserved. Helix-packing model of EcNhaA based on cross-linking data suggests, that in the three dimensional structure of NhaA, residues that affect the pH response may be in close proximity, forming a single pH sensitive domain. Therefore, it is suggested that, despite considerable differences in the primary structure of the antiporters from the bacterial NhaA to the mammalian NHEs, their three-dimensional architectures are conserved. Test of this possibility awaits the atomic resolution of the 3D structure of the antiporters. PMID- 15282169 TI - What is the real crystallographic structure of the L photointermediate of bacteriorhodopsin? AB - In the last few years, three laboratories have reported three entirely different crystallographic models for the L photointermediate of bacteriorhodopsin. All are from X-ray diffraction of illuminated crystals that contain L in photostationary states created at similar cryogenic temperatures. This article compares the models and their implications, the crystallographic statistics and the methods used to derive them, as well as their agreement with non-crystallographic information. PMID- 15282170 TI - Thermodynamic and choreographic constraints for energy transduction by cytochrome c oxidase. AB - Cooperative effects are fundamental for electroprotonic energy transduction processes, crucial to sustain much of life chemistry. However, the primary cooperative mechanism by which transmembrane proteins couple the downhill transfer of electrons to the uphill activation (acidification) of protic groups is still a matter of great controversy. To understand cooperative processes fully, it is necessary to obtain the microscopic thermodynamic parameters of the functional centres and relate them to the relevant structural features, a task difficult to achieve for large proteins. The approach discussed here explores how this may be done by extrapolation from mechanisms used by simpler proteins operative in similar processes. The detailed study of small, soluble cytochromes performing electroprotonic activation has shown how they use anti-electrostatic effects to control the synchronous movement of charges. These include negative e( )/H(+) (redox-Bohr effect) cooperativities. This capacity is the basis to discuss an unorthodox mechanism consistent with the available experimental data on the process of electroprotonic energy transduction performed by cytochrome c oxidase (CcO). PMID- 15282171 TI - BetP of Corynebacterium glutamicum, a transporter with three different functions: betaine transport, osmosensing, and osmoregulation. AB - In order to circumvent deleterious effects of hypo- and hyperosmotic conditions in its environment, Corynebacterium glutamicum has developed a number of mechanisms to counteract osmotic stress. The first response to an osmotic upshift is the activation of uptake mechanisms for the compatible solutes betaine, proline, or ectoine, namely BetP, EctP, ProP, LcoP and PutP. BetP, the most important uptake system responds to osmotic stress by regulation at the level of both protein activity and gene expression. BetP was shown to harbor three different properties, i.e. catalytic activity (betaine transport), sensing of appropriate stimuli (osmosensing) and signal transduction to the catalytic part of the carrier protein which adapts its activity to the extent of osmotic stress (osmoregulation). BetP is comprised of 12 transmembrane segments and carries N- and C-terminal domains, which are involved in osmosensing and/or osmoregulation. Recent results on molecular properties of these domains indicate the significance of particular amino acids within the terminal 25 amino acids of the C-terminal domain of BetP for the process of osmosensing and osmoregulation. PMID- 15282172 TI - The protein import and assembly machinery of the mitochondrial outer membrane. AB - The process of mitochondrial protein import has been studied for many years. Despite this attention, many processes associated with mitochondrial biogenesis are poorly understood. Insight into one of these processes, assembly of beta barrel proteins into the mitochondrial outer membrane, will be discussed. This review focuses on recent data that suggest that assembly of beta-barrel proteins into the outer mitochondrial membrane is dependent on a newly identified protein complex termed the sorting and assembly machinery (SAM complex). Members of the SAM complex have been identified in both eukaryotic and prokaryotic organisms, suggesting that the process of beta-barrel assembly into membranes has been conserved through evolution. PMID- 15282173 TI - Inhibition of mitochondrial respiratory complex I by nitric oxide, peroxynitrite and S-nitrosothiols. AB - NO or its derivatives (reactive nitrogen species, RNS) inhibit mitochondrial complex I by several different mechanisms that are not well characterised. There is an inactivation by NO, peroxynitrite and S-nitrosothiols that is reversible by light or reduced thiols, and therefore may be due to S-nitrosation or Fe nitrosylation of the complex. There is also an irreversible inhibition by peroxynitrite, other oxidants and high levels of NO, which may be due to tyrosine nitration, oxidation of residues or damage of iron sulfur centres. Inactivation of complex I by NO or RNS is seen in cells or tissues expressing iNOS, and may be relevant to inflammatory pathologies, such as septic shock and Parkinson's disease. PMID- 15282174 TI - Probing light-induced conformational transitions in bacterial photosynthetic reaction centers embedded in trehalose-water amorphous matrices. AB - The coupling between electron transfer and protein dynamics has been studied in photosynthetic reaction centers (RC) from Rhodobacter sphaeroides by embedding the protein into room temperature solid trehalose-water matrices. Electron transfer kinetics from the primary quinone acceptor (Q(A)(-)) to the photoxidized donor (P(+)) were measured as a function of the duration of photoexcitation from 20 ns (laser flash) to more than 1 min. Decreasing the water content of the matrix down to approximately 5x10(3) water molecules per RC causes a reversible four-times acceleration of P(+)Q(A)(-) recombination after the laser pulse. By comparing the broadly distributed kinetics observed under these conditions with the ones measured in glycerol-water mixtures at cryogenic temperatures, we conclude that RC relaxation from the dark-adapted to the light-adapted state and thermal fluctuations among conformational substates are hindered in the room temperature matrix over the time scale of tens of milliseconds. When the duration of photoexcitation is increased from a few milliseconds to the second time scale, recombination kinetics of P(+)Q(A)(-) slows down progressively and becomes less distributed, indicating that even in the driest matrices, during continuous illumination, the RC is gaining a limited conformational freedom that results in partial stabilization of P(+)Q(A)(-). This behavior is consistent with a tight structural and dynamical coupling between the protein surface and the trehalose water matrix. PMID- 15282175 TI - Induction of the mitochondrial permeability transition by the DNA alkylating agent N-methyl-N'-nitro-N-nitrosoguanidine. Sorting cause and consequence of mitochondrial dysfunction. AB - The alkylating agent N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) alters DNA and stimulates the activity of poly(ADP-ribose) polymerase-1 (PARP-1), a nuclear enzyme involved in DNA repair. The consumption of cellular NAD(+) by PARP-1 is accompanied by ATP depletion, mitochondrial depolarization and release of proapoptotic proteins, but whether a causal relationship exists among these events remains an open question. Most of cellular NAD(+) is stored in the mitochondrial matrix and becomes available for cytosolic and nuclear processes only after its release through the permeability transition pore (PTP), a voltage gated inner membrane channel. Here we have explored whether MNNG affects mitochondrial function upstream of PARP-1 activation. We show that MNNG has a dual effect on isolated mitochondria. At relatively low concentrations (up to 0.1 mM), it selectively sensitizes the PTP to opening, while at higher concentrations (above 0.5 mM) it inhibits carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone (FCCP)-stimulated respiration. MNNG caused PTP opening and activation of the mitochondrial proapoptotic pathway in intact HeLa cells, which resulted in cell death that could be prevented by the PTP inhibitor CsA. We conclude that a key event in MNNG-dependent cell death is induction of PTP opening that occurs independently of PARP-1 activation. PMID- 15282176 TI - Role of calcium signaling in the activation of mitochondrial nitric oxide synthase and citric acid cycle. AB - An apparent discrepancy arises about the role of calcium on the rates of oxygen consumption by mitochondria: mitochondrial calcium increases the rate of oxygen consumption because of the activation of calcium-activated dehydrogenases, and by activating mitochondrial nitric oxide synthase (mtNOS), decreases the rates of oxygen consumption because nitric oxide is a competitive inhibitor of cytochrome oxidase. To this end, the rates of oxygen consumption and nitric oxide production were followed in isolated rat liver mitochondria in the presence of either L-Arg (to sustain a mtNOS activity) or N(G)-monomethyl-L-Arg (NMMA, a competitive inhibitor of mtNOS) under State 3 conditions. In the presence of NMMA, the rates of State 3 oxygen consumption exhibited a K(0.5) of 0.16 microM intramitochondrial free calcium, agreeing with those required for the activation of the Krebs cycle. By plotting the difference between the rates of oxygen consumption in State 3 with L-Arg and with NMMA at various calcium concentrations, a K(0.5) of 1.2 microM intramitochondrial free calcium was obtained, similar to the K(0.5) (0.9 microM) of the dependence of the rate of nitric oxide production on calcium concentrations. The activation of dehydrogenases, followed by the activation of mtNOS, would lead to the modulation of the Krebs cycle activity by the modulation of nitric oxide on the respiratory rates. This would ensue in changes in the NADH/NAD and ATP/ADP ratios, which would influence the rate of the cycle and the oxygen diffusion. PMID- 15282177 TI - Hydration switch model for the proton transfer in the Schiff base region of bacteriorhodopsin. AB - In a light-driven proton-pump protein, bacteriorhodopsin (BR), protonated Schiff base of the retinal chromophore and Asp85 form ion-pair state, which is stabilized by a bridged water molecule. After light absorption, all-trans to 13 cis photoisomerization takes place, followed by the primary proton transfer from the Schiff base to Asp85 that triggers sequential proton transfer reactions for the pump. Fourier transform infrared (FTIR) spectroscopy first observed O-H stretching vibrations of water during the photocycle of BR, and accurate spectral acquisition has extended the water stretching frequencies into the entire stretching frequency region in D(2)O. This enabled to capture the water molecules hydrating with negative charges, and we have identified the water O-D stretch at 2171 cm(-1) as the bridged water interacting with Asp85. We found that retinal isomerization weakens the hydrogen bond in the K intermediate, but not in the later intermediates such as L, M, and N. On the basis of the observation particularly on the M intermediate, we proposed a model for the mechanism of proton transfer from the Schiff base to Asp85. In the "hydration switch model", hydration of a water molecule is switched in the M intermediate from Asp85 to Asp212. This will have raised the pK(a) of the proton acceptor, and the proton transfer is from the Schiff base to Asp85. PMID- 15282178 TI - Mitochondrial diseases. AB - By convention, the term "mitochondrial diseases" refers to disorders of the mitochondrial respiratory chain, which is the only metabolic pathway in the cell that is under the dual control of the mitochondrial genome (mtDNA) and the nuclear genome (nDNA). Therefore, a genetic classification of the mitochondrial diseases distinguishes disorders due to mutations in mtDNA, which are governed by the relatively lax rules of mitochondrial genetics, and disorders due to mutations in nDNA, which are governed by the stricter rules of mendelian genetics. Mutations in mtDNA can be divided into those that impair mitochondrial protein synthesis in toto and those that affect any one of the 13 respiratory chain subunits encoded by mtDNA. Essential clinical features for each group of diseases are reviewed. Disorders due to mutations in nDNA are more abundant not only because most respiratory chain subunits are nucleus-encoded but also because correct assembly and functioning of the respiratory chain require numerous steps, all of which are under the control of nDNA. These steps (and related diseases) include: (i) synthesis of assembly proteins; (ii) intergenomic signaling; (iii) mitochondrial importation of nDNA-encoded proteins; (iv) synthesis of inner mitochondrial membrane phospholipids; (v) mitochondrial motility and fission. PMID- 15282179 TI - Bioenergetics of mitochondrial diseases associated with mtDNA mutations. AB - This mini-review summarizes our present view of the biochemical alterations associated with mitochondrial DNA (mtDNA) point mutations. Mitochondrial cytopathies caused by mutations of mtDNA are well-known genetic and clinical entities, but the biochemical pathogenic mechanisms are often obscure. Leber's hereditary optic neuropathy (LHON) is due to three main mutations in genes for complex I subunits. Even if the catalytic activity of complex I is maintained except in cells carrying the 3460/ND1 mutation, in all cases there is a change in sensitivity to complex I inhibitors and an impairment of mitochondrial respiration, eliciting the possibility of generation of reactive oxygen species (ROS) by the complex. Neurogenic muscle weakness, Ataxia and Retinitis Pigmentosa (NARP), is due to a mutation in the ATPase-6 gene. In NARP patients ATP synthesis is strongly depressed to an extent proportional to the mutation load; nevertheless, ATP hydrolysis and ATP-driven proton translocation are not affected. It is suggested that the NARP mutation affects the ability of the enzyme to couple proton transport to ATP synthesis. A point mutation in subunit III of cytochrome c oxidase is accompanied by a syndrome resembling MELAS: however, no major biochemical defect is found, if we except an enhanced production of ROS. The mechanism of such enhancement is at present unknown. In this review, we draw attention to a few examples in which the overproduction of ROS might represent a common step in the induction of clinical phenotypes and/or in the progression of several human pathologies associated with mtDNA point mutations. PMID- 15282180 TI - Protonmotive cooperativity in cytochrome c oxidase. AB - Cooperative linkage of solute binding at separate binding sites in allosteric proteins is an important functional attribute of soluble and membrane bound hemoproteins. Analysis of proton/electron coupling at the four redox centers, i.e. Cu(A), heme a, heme a(3) and Cu(B), in the purified bovine cytochrome c oxidase in the unliganded, CO-liganded and CN-liganded states is presented. These studies are based on direct measurement of scalar proton translocation associated with oxido-reduction of the metal centers and pH dependence of the midpoint potential of the redox centers. Heme a (and Cu(A)) exhibits a cooperative proton/electron linkage (Bohr effect). Bohr effect seems also to be associated with the oxygen-reduction chemistry at the heme a(3)-Cu(B) binuclear center. Data on electron transfer in cytochrome c oxidase are also presented, which, together with structural data, provide evidence showing the occurrence of direct electron transfer from Cu(A) to the binuclear center in addition to electron transfer via heme a. A survey of structural and functional data showing the essential role of cooperative proton/electron linkage at heme a in the proton pump of cytochrome c oxidase is presented. On the basis of this and related functional and structural information, variants for cooperative mechanisms in the proton pump of the oxidase are examined. PMID- 15282181 TI - Diverse and essential roles of mammalian vacuolar-type proton pump ATPase: toward the physiological understanding of inside acidic compartments. AB - The vacuolar-type H(+)-ATPases (V-ATPase) are a family of multi-subunit ATP dependent proton pumps involved in a wide variety of physiological processes. They are present in endomembrane organelles such as vacuoles, lysosomes, endosomes, the Golgi apparatus, chromaffin granules and coated vesicles, and acidify the luminal pH of these intracellular compartments. They also pump protons across the plasma membranes of specialized cells including osteoclasts and epithelial cells in kidneys and male genital tracts. Here, we briefly summarize our recent studies on the diverse and essential roles of mammalian V ATPase. PMID- 15282182 TI - Mitochondrial diseases and ATPase defects of nuclear origin. AB - Dysfunctions of the F(1)F(o)-ATPase complex cause severe mitochondrial diseases affecting primarily the paediatric population. While in the maternally inherited ATPase defects due to mtDNA mutations in the ATP6 gene the enzyme is structurally and functionally modified, in ATPase defects of nuclear origin mitochondria contain a decreased amount of otherwise normal enzyme. In this case biosynthesis of ATPase is down-regulated due to a block at the early stage of enzyme assembly formation of the F(1) catalytic part. The pathogenetic mechanism implicates dysfunction of Atp12 or other F(1)-specific assembly factors. For cellular energetics, however, the negative consequences may be quite similar irrespective of whether the ATPase dysfunction is of mitochondrial or nuclear origin. PMID- 15282183 TI - Understanding aging: revealing order out of chaos. AB - Aging is often described as an extremely complex process affecting all of the vital parameters of an individual. In this article, we review how understanding of aging evolved from the first analyses of population survival to the identification of the molecular mechanisms regulating life span. Abundant evidence implicates mitochondria in aging and we focus on the three main components of the mitochondrial theory of aging: (1) increased reactive oxygen species (ROS) production, (2) mitochondrial DNA (mtDNA) damage accumulation, and (3) progressive respiratory chain dysfunction. Experimental evidence shows a relationship between respiratory chain dysfunction, ROS damage, and aging in most of the model organisms. However, involvement of the mtDNA mutations in the aging process is still debated. We recently created a mutant mouse strain with increased levels of somatic mtDNA mutations causing a progressive respiratory chain deficiency and premature aging. These mice demonstrate the fundamental importance of the accumulation of mtDNA alterations in aging. We present here an integrative model where aging is provoked by a single primary event leading to a variety of effects and secondary causes. PMID- 15282184 TI - Structural insight into the cooperativity between catalytic and noncatalytic sites of F1-ATPase. AB - F1-ATPase, the catalytic sector of Fo-F1 ATPases-ATPsynthases, displays an apparent negative cooperativity for ATP hydrolysis at high ATP concentrations which involves noncatalytic and catalytic nucleotide binding sites. The molecular mechanism of such cooperativity is currently unknown. To get further insights, we have investigated the structural consequences of the single mutation of two residues: Q173L in the alpha-subunit and Q170Y in the beta-subunit of the F1 ATPase of the yeast Schizosaccharomyces pombe. These residues are localized in or near the Walker-A motifs of each subunit and their mutation produces an opposite effect on the negative cooperativity. The betaQ170 residue (M167 in beef heart) is located close to the binding site for the phosphate-Mg moiety of the nucleotide. Its replacement by tyrosine converts this site into a close state with increased affinity for the bound nucleotide and leads to an increase of negative cooperativity. In contrast, the alphaQ173L mutation (Q172 in beef heart) abolishes negative cooperativity due to the loss of two H-bonds: one stabilizing the nucleotide bound to the noncatalytic site and the other linking alphaQ173 to the adjacent betaT354, localized at the alpha(DP)-beta(TP) interface. The properties of these mutants suggest that negative cooperativity occurs through interactions between neighbor alpha- and beta-subunits. Indeed, in the beef heart enzyme, (i) the alpha(DP)-beta(TP) interface is stabilized by a vicinal alphaR171 betaD352 salt bridge (ii) betaD352 and betaT354 belong to a short peptidic stretch close to betaY345, the aromatic group of which interacts with the adenine moiety of the nucleotide bound to the catalytic site. We therefore propose that the betaY345-betaT354 stretch (beef heart numbering) constitutes a short link that drives structural modifications from a noncatalytic site to the neighbor catalytic site in which, as a result, the affinity for ADP is modulated. PMID- 15282185 TI - "Wages of fear": transient threefold decrease in intracellular ATP level imposes apoptosis. AB - In HeLa cells, complete inhibition of oxidative phosphorylation by oligomycin, myxothiazol or FCCP combined with partial inhibition of glycolysis by DOG resulted in a steady threefold decrease in the intracellular ATP level. The ATP level recovers when the DOG-containing medium was replaced by that with high glucose. In 48 h after a transient (3 h) [ATP] lowering followed by recovery of the ATP level, the majority of the cells commits suicide by means of apoptosis. The cell death does not occur if DOG or an oxidative phosphorylation inhibitor was added separately, treatments resulting in 10-35% lowering of [ATP]. Apoptosis is accompanied by Bax translocation to mitochondria, cytochrome c release into cytosol, caspase activation, reactive oxygen species (ROS) generation, and reorganization and decomposition of chromatin. Apoptosis appears to be sensitive to oncoprotein Bcl-2 and a pancaspase inhibitor zVADfmk. In the latter case, necrosis is shown to develop instead of apoptosis. The cell suicide is resistant to cyclosporine A, a phospholipase inhibitor trifluoroperazine, the JNK and p38 kinase inhibitors, oligomycin, N-acetyl cysteine and mitoQ, differing in these respects from the tumor necrosis factor (TNF)- and H(2)O(2)-induced apoptoses. It is suggested that the ATP concentration in the cell is monitored by intracellular "ATP-meter(s)" generating a cell suicide signal when ATP decreases, even temporarily, below some critical level (around 1 mM). PMID- 15282186 TI - Subunit composition of mitochondrial complex I from the yeast Yarrowia lipolytica. AB - Here we present a first assessment of the subunit inventory of mitochondrial complex I from the obligate aerobic yeast Yarrowia lipolytica. A total of 37 subunits were identified. In addition to the seven central, nuclear coded, and the seven mitochondrially coded subunits, 23 accessory subunits were found based on 2D electrophoretic and mass spectroscopic analysis in combination with sequence information from the Y. lipolytica genome. Nineteen of the 23 accessory subunits are clearly conserved between Y. lipolytica and mammals. The remaining four accessory subunits include NUWM, which has no apparent homologue in any other organism and is predicted to contain a single transmembrane domain bounded by highly charged extramembraneous domains. This structural organization is shared among a group of 7 subunits in the Y. lipolytica and 14 subunits in the mammalian enzyme. Because only five of these subunits display significant evolutionary conservation, their as yet unknown function is proposed to be structure- rather than sequence-specific. The NUWM subunit could be assigned to a hydrophobic subcomplex obtained by fragmentation and sucrose gradient centrifugation. Its position within the membrane arm was determined by electron microscopic single particle analysis of Y. lipolytica complex I decorated with a NUWM-specific monoclonal antibody. PMID- 15282187 TI - Activation by retinoids of the uncoupling protein UCP1. AB - The uncoupling protein from brown adipose tissue (UCP1) is a transporter that catalyzes a regulated discharged of the mitochondrial proton gradient. The proton conductance in UCP1 is inhibited by nucleotides and activated by fatty acids. We have recently shown that all-trans-retinoic acid (ATRA) is a high-affinity activator of UCP1. In the present report, we have set to analyze the structural requirements for the ligands that activate UCP1 and particularly the specificity for different retinoids. For this purpose, we have developed a new protocol to determine the activity of UCP1 in respiring yeast mitochondria that can be adapted for high-throughput screenings. Our results evidence differences between the structural requirements for the activation by fatty acids and retinoids. Thus, although all active retinoids must possess a carboxylate, the introduction of additional polar groups renders them inactive. The linear and rigid structure of these molecules suggests the existence of a long hydrophobic binding pocket. We postulate that the access to the retinoid binding site must occur from the lipid bilayer and this could be at the interface between two transmembrane alpha helices. PMID- 15282188 TI - The quinone chemistry of bc complexes. AB - The quinone chemistry that gives rise to the rather unusual strict bifurcation of electron transfer at the Q(o) site of the cytochrome bc complexes remains controversial. In this article, I review recent ideas and propose a "logic-gated" binding mechanism that combines classical quinone electrochemistry with specific hydrogen bonding requirements and results in a reversible reaction that minimizes unwanted side-reactions that could otherwise undermine the efficiency of the Q cycle proton/electron coupling mechanism. PMID- 15282189 TI - Bioenergetics shapes cellular death pathways in Leber's hereditary optic neuropathy: a model of mitochondrial neurodegeneration. AB - Leber's hereditary optic neuropathy (LHON) was the first maternally inherited disease to be associated with point mutations in mitochondrial DNA and is now considered the most prevalent mitochondrial disorder. The pathology is characterized by selective loss of ganglion cells in the retina leading to central vision loss and optic atrophy, prevalently in young males. The pathogenic mtDNA point mutations for LHON affect complex I with the double effect of lowering the ATP synthesis driven by complex I substrates and increasing oxidative stress chronically. In this review, we first consider the biochemical changes associated with the proton-translocating NADH-quinone oxidoreductase of mitochondria in cybrid cells carrying the most common LHON mutations. However, the LHON cybrid bioenergetic dysfunction is essentially compensated under normal conditions, i.e. in glucose medium, but is unrevealed by stressful conditions such as growing cybrids in glucose free/galactose medium, which forces cells to rely only on respiratory chain for ATP synthesis. In fact, the second part of this review deals with the investigation of LHON cybrid death pathway in galactose medium. The parallel marked changes in antioxidant enzymes, during the time-course of galactose experiments, also reveal a relevant role played by oxidative stress. The LHON cybrid model sheds light on the complex interplay amongst the different levels of biochemical consequences deriving from complex I mutations in determining neurodegeneration in LHON, and suggests an unsuspected role of bioenergetics in shaping cell death pathways. PMID- 15282191 TI - Potentiation of effect of PKA stimulation of Xenopus CFTR by activation of PKC: role of NBD2. AB - Activity of the human (h) cystic fibrosis transmembrane conductance regulator (CFTR) channel is predominantly regulated by PKA-mediated phosphorylation. In contrast, Xenopus (X)CFTR is more responsive to PKC than PKA stimulation. We investigated the interaction between the two kinases in XCFTR. We expressed XCFTR in Xenopus oocytes and maximally stimulated it with PKA agonists. The magnitude of activation after PKC stimulation was about eightfold that without pretreatment with PKC agonist. hCFTR, expressed in the same system, lacked this response. We name this phenomenon XCFTR-specific PKC potentiation effect. To ascertain its biophysical mechanism, we first tested for XCFTR channel insertion into the plasma membrane by a substituted-cysteine-accessibility method. No insertion was detected during kinase stimulation. Next, we studied single-channel properties and found that the single-channel open probability (Po) with PKA stimulation subsequent to PKC stimulation was 2.8-fold that observed in the absence of PKC preactivation and that single-channel conductance (gamma) was increased by approximately 22%. To ascertain which XCFTR regions are responsible for the potentiation, we constructed several XCFTR-hCFTR chimeras, expressed them in Xenopus oocytes, and tested them electrophysiologically. Two chimeras [hCFTR NH2 terminal region or regulatory (R) domain in XCFTR] showed a significant decrease in potentiation. In the chimera in which XCFTR nucleotide-binding domain (NBD)2 was replaced with the hCFTR sequence there was no potentiation whatsoever. The converse chimera (hCFTR with Xenopus NBD2) did not exhibit potentiation. These results indicate that potentiation by PKC involves a large increase in Po (with a small change in gamma) without CFTR channel insertion into the plasma membrane, that XCFTR NBD2 is necessary but not sufficient for the effect, and that the potentiation effect is likely to involve other CFTR domains. PMID- 15282190 TI - Lipopolysaccharide stimulates nitric oxide synthase-2 expression in murine skeletal muscle and C(2)C(12) myoblasts via Toll-like receptor-4 and c-Jun NH(2) terminal kinase pathways. AB - The inducible form of nitric oxide synthase (NOS2) catalyzes the synthesis of nitric oxide (NO) from arginine in response to injury and infection. NOS2 is expressed predominantly by macrophages and lymphocytes. However, skeletal muscle also expresses NOS2 in response to inflammatory stimuli. The present study sought to determine whether lipopolysaccharide (LPS) stimulates NOS2 in skeletal muscle via Toll-like receptor-4 (TLR4). Intraperitoneal injection of LPS in wild-type mice (C3H/HeSnJ) increased NOS2 mRNA fourfold in skeletal muscle, while no change in NOS2 mRNA was observed in C3H/HeJ mice that harbored a mutation in the LPS receptor. NOS2 coimmunoprecipitated with the muscle-specific caveolin-3 protein, suggesting that myofibers per se respond to LPS in vivo. LPS stimulated NOS2 mRNA expression in C(2)C(12) myocytes, and the regulation of NOS2 mRNA was comparable in myoblasts and differentiated myotubes. LPS transiently stimulated the phosphorylation of the interleukin-1 receptor-associated kinase (IRAK-1) in C(2)C(12) cells and decreased the total amount of IRAK-1 both in vitro and in vivo over time. LPS stimulated the expression of an NF-kappabeta reporter plasmid, and this was inhibited by the proteasomal inhibitor MG-132. Both myoblasts and myotubes expressed TLR2 and TLR4 mRNA. Expression of a dominant negative form of TLR4 in C(2)C(12) cells blocked LPS-induced NF-kappabeta reporter activity. SP-600125 [a c-Jun NH(2)-terminal kinase (JNK) inhibitor] also prevented LPS stimulation of NOS2 expression. Moreover, the JNK inhibitor prevented the LPS-induced increase in NO synthesis. These data indicate that LPS increases NOS2 mRNA expression in muscle via a TLR4-dependent mechanism. PMID- 15282192 TI - Biophysical characterization of zebrafish connexin35 hemichannels. AB - A subset of connexins can form unopposed hemichannels in expression systems, providing an opportunity for comparison of hemichannel gating properties with those of intact gap junction channels. Zebrafish connexin35 (Cx35) is a member of the Cx35/Cx36 subgroup of connexins highly expressed in the retina and brain. In the present study, we have shown that Cx35 expression in Xenopus oocytes and N2A cells produced large outward whole cell currents on cell depolarization. Using whole cell, cell-attached, and excised patch configurations, we obtained multichannel and single-channel current recordings attributable to the Cx35 hemichannels (I(hc)) that were activated and increased by stepwise depolarization of membrane potential (V(m)) and deactivated by hyperpolarization. The currents were not detected in untransfected N2A cells or in control oocytes injected with antisense Cx38. However, water-injected oocytes that were not treated with antisense showed activities attributable to Cx38 hemichannels that were easily distinguishable from Cx35 hemichannels by a significantly larger unitary conductance (gamma(hc): 250-320 pS). The gamma(hc) of Cx35 hemichannels exhibited a pronounced V(m) dependence; i.e., gamma(hc) increased/decreased with relative hyperpolarization/depolarization (gamma(hc) was 72 pS at V(m) = -100 mV and 35 pS at V(m) = 100 mV). Extrapolation to V(m) = 0 mV predicted a gamma(hc) of 48 pS, suggesting a unitary conductance of intact Cx35 gap junction channels of approximately 24 pS. Channel gating was also V(m) dependent: open time declined with negative V(m) and increased with positive V(m). The ability to break down the complex gating of intact intercellular channels into component hemichannels in vitro will help to evaluate putative physiological roles for hemichannels in vivo. PMID- 15282193 TI - Real-time three-dimensional imaging of lipid signal transduction: apical membrane insertion of epithelial Na(+) channels. AB - In the distal tubule, Na(+) resorption is mediated by epithelial Na(+) channels (ENaC). Hormones such as aldosterone, vasopressin, and insulin modulate ENaC membrane targeting, assembly, and/or kinetic activity, thereby regulating salt and water homeostasis. Insulin binds to a receptor on the basal membrane to initiate a signal transduction cascade that rapidly results in an increase in apical membrane ENaC. Current models of this signaling pathway envision diffusion of signaling intermediates from the basal to the apical membrane. This necessitates diffusion of several high-molecular-weight signaling elements across a three-dimensional space. Transduction of the insulin signal involves the phosphoinositide pathway, but how and where this lipid-based signaling pathway controls ENaC activity is not known. We used tagged channels, biosensor lipid probes, and intravital imaging to investigate the role of lipids in insulin stimulated Na(+) flux. Insulin-stimulated delivery of intracellular ENaC to apical membranes was concurrent with plasma membrane-limited changes in lipid composition. Notably, in response to insulin, phosphatidylinositol 3,4,5 trisphosphate (PIP(3)) formed in the basolateral membrane, rapidly diffused within the bilayer, and crossed the tight junction to enter the apical membrane. This novel signaling pathway takes advantage of the fact that the lipids of the plasma membrane's inner leaflet are not constrained by the tight junction. Therefore, diffusion of PIP(3) as a signal transduction intermediate occurs within a planar surface, thus facilitating swift responses and confining and controlling the signaling pathway. PMID- 15282195 TI - Effect of creatine on contractile force and sensitivity in mechanically skinned single fibers from rat skeletal muscle. AB - Increasing the intramuscular stores of total creatine [TCr = creatine (Cr) + creatine phosphate (CrP)] can result in improved muscle performance during certain types of exercise in humans. Initial uptake of Cr is accompanied by an increase in cellular water to maintain osmotic balance, resulting in a decrease in myoplasmic ionic strength. Mechanically skinned single fibers from rat soleus (SOL) and extensor digitorum longus (EDL) muscles were used to examine the direct effects on the contractile apparatus of increasing [Cr], increasing [Cr] plus decreasing ionic strength, and increasing [Cr] and [CrP] with no change in ionic strength. Increasing [Cr] from 19 to 32 mM, accompanied by appropriate increases in water to maintain osmolality, had appreciable beneficial effects on contractile apparatus performance. Compared with control conditions, both SOL and EDL fibers showed increases in Ca(2+) sensitivity (+0.061 +/- 0.004 and +0.049 +/ 0.009 pCa units, respectively) and maximum Ca(2+)-activated force (to 104 +/- 1 and 105 +/- 1%, respectively). In contrast, increasing [Cr] alone had a small inhibitory effect. When both [Cr] and [CrP] were increased, there was virtually no change in Ca(2+) sensitivity of the contractile apparatus, and maximum Ca(2+) activated force was approximately 106 +/- 1% compared with control conditions in both SOL and EDL fibers. These results suggest that the initial improvement in performance observed with Cr supplementation is likely due in large part to direct effects of the accompanying decrease in myoplasmic ionic strength on the properties of the contractile apparatus. PMID- 15282194 TI - Detection of intracellular iron by its regulatory effect. AB - Intracellular iron regulates gene expression by inhibiting the interaction of iron regulatory proteins (IRPs) with RNA motifs called iron-responsive elements (IREs). To assay this interaction in living cells we have developed two fluorescent IRE-based reporters that rapidly, reversibly, and specifically respond to changes in cellular iron status as well as signaling that modifies IRP activity. The reporters were also sufficiently sensitive to distinguish apo- from holotransferrin in the medium, to detect the effect of modifiers of the transferrin pathway such as HFE, and to detect the donation or chelation of iron by siderophores bound to the lipocalin neutrophil gelatinase-associated lipocalin (Ngal). In addition, alternative configurations of the IRE motif either enhanced or repressed fluorescence, permitting a ratio analysis of the iron-dependent response. These characteristics make it possible to visualize iron-IRP-IRE interactions in vivo. PMID- 15282196 TI - Parathyroid hormone-related protein regulates apoptosis in lung cancer cells through protein kinase A. AB - Parathyroid hormone-related protein (PTHrP)-(1-34) and PTHrP-(140-173) protect lung cancer cells from apoptosis after ultraviolet (UV) irradiation. This study evaluated upstream signaling in PTHrP-mediated alteration of lung cancer cell sensitivity to apoptosis. The two peptides increased cAMP levels in BEN lung cancer cells by 15-35% in a dose-dependent fashion, suggesting signaling through protein kinase A (PKA). In line with this view, the PKA inhibitor H89 abrogated the protective effects of PTHrP-(1-34) and PTHrP-(140-173) against caspase activation and DNA loss. PKA activation by forskolin, 3-isobutyl-1-methylxanthine (IBMX), or 8-(4-chlorophenylthio)adenosine 3',5'-cyclic monophosphate attenuated and H89 augmented apoptosis after UV exposure as indicated by caspase-3 activation, cell DNA loss, and morphological criteria. Studies with IBMX and varying doses of forskolin indicated that small increases in cAMP, on the order of those generated by IBMX alone and the PTHrP peptides, were sufficient to protect lung cancer cells from apoptosis. In summary, PTHrP-(1-34) and PTHrP-(140 173) stimulate PKA in lung carcinoma cells and protect cells against UV-induced caspase-3 activation and DNA fragmentation. PKA activation by other means also induces resistance to apoptosis, and the protective effect of the PTHrP peptide is blocked by PKA inhibition. Thus PKA appears to have a role in the regulatory effects of PTHrP on lung cancer cell survival. PMID- 15282197 TI - Cells derived from the circulation contribute to the repair of lung injury. AB - Bone marrow (stem/progenitor) cells have been shown to "differentiate" into cells in multiple tissues, including lung. A low number of hematopoietic stem/progenitor cells also circulate in peripheral blood. The physiologic roles of these cells are still uncertain. This study was designed to test, using parabiotic mice that were joined surgically, whether stem/progenitor cells in blood contributed to the regeneration of lung after injury. Parabiotic mice were generated surgically by joining green fluorescent protein transgenic mice and wild-type littermates. These mice developed a common circulation (approximately 50% green cells in blood) by 2 weeks after surgery. The wild-type mouse was either uninjured or lethally irradiated or received intratracheal elastase or the combination of radiation with intratracheal elastase injection. Radiation or the combination of radiation with elastase significantly increased the proportion of bright green cells in the lungs of the wild-type mice. Morphologically, interstitial monocytes/macrophages, subepithelial fibroblast-like interstitial cells, and additionally type I alveolar epithelial cells immunostained for green fluorescent protein in wild-type mice. Approximately 5 to 20% of lung fibroblasts primary cultured from injured wild-type mice were green fluorescent protein expressing cells, indicating their blood derivation. This study demonstrates that stem/progenitor cells in blood contribute to the repair of lung injury in irradiated mice. PMID- 15282198 TI - Acute effects of ozone on mortality from the "air pollution and health: a European approach" project. AB - In the Air Pollution and Health: A European Approach (APHEA2) project, the effects of ambient ozone concentrations on mortality were investigated. Data were collected on daily ozone concentrations, the daily number of deaths, confounders, and potential effect modifiers from 23 cities/areas for at least 3 years since 1990. Effect estimates were obtained for each city with city-specific models and were combined using second-stage regression models. No significant effects were observed during the cold half of the year. For the warm season, an increase in the 1-hour ozone concentration by 10 mug/m3 was associated with a 0.33% (95% confidence interval [CI], 0.17-0.52) increase in the total daily number of deaths, 0.45% (95% CI, 0.22-0.69) in the number of cardiovascular deaths, and 1.13% (95% CI, 0.62-1.48) in the number of respiratory deaths. The corresponding figures for the 8-hour ozone were similar. The associations with total mortality were independent of SO2 and particulate matter with aerodynamic diameter less than 10 mum (PM10) but were somewhat confounded by NO2 and CO. Individual city estimates were heterogeneous for total (a higher standardized mortality rate was associated with larger effects) and cardiovascular mortality (larger effects were observed in southern cities). The dose-response curve of ozone effects on total mortality during the summer did not deviate significantly from linearity. PMID- 15282199 TI - Association of vitamin D receptor genetic variants with susceptibility to asthma and atopy. AB - Genome scans for asthma have identified suggestive or significant linkages on 17 different chromosomes, including chromosome 12, region q13-23, housing the vitamin D receptor (VDR) gene. Through interaction with VDR, 1,25 dihydroxyvitamin D3 mediates numerous biological activities, such as regulation of helper T-cell development and subsequent cytokine secretion profiles. Variants of the VDR have been found to be associated with immune-mediated diseases that are characterized by an imbalance in helper T-cell development, such as Crohn's disease and tuberculosis. The VDR, hence, is a good candidate to be investigated for association with asthma, which is characterized by enhanced helper T-cell type 2 activity. Here, we examined VDR genetic variants in an asthma family-based cohort from Quebec. We report six variants to be strongly associated with asthma and four with atopy (0.0005 < or = p < or = 0.05). Analysis of the linkage disequilibrium pattern and haplotypes also revealed significant association with both phenotypes (0.0004 < or = p < or = 0.01). The findings have been replicated by another research team in a second but not in a third cohort. These results identify VDR variants as genetic risk factors for asthma/atopy and implicate a non-human leukocyte antigen immunoregulatory molecule in the pathogenesis of asthma and atopy. PMID- 15282200 TI - Association of vitamin D receptor gene polymorphisms with childhood and adult asthma. AB - Vitamin D receptor (VDR) polymorphisms have been associated with several immune related diseases, and VDR and vitamin D itself modulate T cell differentiation. VDR maps to chromosome 12q, near a region commonly linked to asthma. We evaluated VDR as part of a 12q positional candidate survey, and in response to observations of VDR polymorphism associations with asthma and atopy in a founder population of Quebec. Twenty-eight loci in 7 positional candidates (7 in VDR) were genotyped in 582 families. Whereas other candidates demonstrated no association, the VDR ApaI polymorphism demonstrated significant transmission distortion, with undertransmission of the C allele in a ratio of 4:5 (p = 0.01). This association was most prominent in girls, in whom distortion was more marked (p = 0.009). Sex specific associations between multiple VDR polymorphisms and immunoglobulin E levels were also observed (p = 0.006-0.01). Asthma associations were replicated in a second cohort (517 females with asthma and 519 matched control subjects): 4 of 6 VDR variants demonstrated significant association (p = 0.02-0.04). The direction of association in this second cohort was opposite to the effects seen in the trios, but similar to findings in the Quebec study. These results suggest that VDR influences asthma and allergy susceptibility in a complex manner. PMID- 15282201 TI - Smoking and tuberculosis among the elderly in Hong Kong. AB - A cohort of 42,655 clients that were first registered with the Elderly Health Service in 2000 were followed prospectively through the tuberculosis (TB) notification registry until the end of 2002. A total of 286 active TB cases (186 culture confirmed) were identified. The annual TB notification rates were 735, 427, and 174 per 100,000 among current smokers, ex-smokers, and never-smokers, respectively (p < 0.001). The trend in TB risk persisted after the control of background characteristics using Cox proportional hazards analysis (adjusted hazard ratios [HRs]: 2.63, 1.41, and 1, p < 0.001). In comparison with never smokers, current smokers had an excess risk of pulmonary TB (adjusted HR, 2.87; 95% confidence interval [CI], 2.00-4.11; p < 0.001), but not extrapulmonary TB (adjusted HR, 1.04; 95% CI, 0.33-3.30; p = 0.95). Among the current smokers, those who developed TB smoked more cigarettes per day than those who did not (13.43, SD 8.76 vs. 10.96, SD 7.87, p = 0.01). A statistically significant dose response relationship was observed with respect to active TB and culture confirmed TB (both p < 0.05). Smoking accounted for 32.8% (95% CI, 14.9-48.0%), 8.6% (95% CI, 3.3-15.1%), and 18.7% (95% CI, 7.7-30.4%) of the TB risk among males, females, and the entire cohort, respectively. Approximately 44.9% (95% CI, 20.7-64.6%) of the sex difference was attributable to smoking. PMID- 15282202 TI - Sphingosine 1-phosphate reduces vascular leak in murine and canine models of acute lung injury. AB - Excessive mechanical stress is a key component of ventilator-associated lung injury, resulting in profound vascular leak and an intense inflammatory response. To extend our in vitro observations concerning the barrier-protective effects of the lipid growth factor sphingosine 1-phosphate (Sph 1-P), we assessed the ability of Sph 1-P to prevent regional pulmonary edema accumulation in clinically relevant rodent and canine models of acute lung injury induced by combined intrabronchial endotoxin administration and high tidal volume mechanical ventilation. Intravenously delivered Sph 1-P significantly attenuated both alveolar and vascular barrier dysfunction while significantly reducing shunt formation associated with lung injury. Whole lung computed tomographic image analysis demonstrated the capability of Sph 1-P to abrogate significantly the accumulation of extravascular lung water evoked by 6-hour exposure to endotoxin. Axial density profiles and vertical density gradients localized the Sph 1-P response to transitional zones between aerated and consolidated lung regions. Together, these results indicate that Sph 1-P represents a novel therapeutic intervention for the prevention of pulmonary edema related to inflammatory injury and increased vascular permeability. PMID- 15282203 TI - The development of emphysema in cigarette smoke-exposed mice is strain dependent. AB - Only 20% of smokers develop chronic obstructive pulmonary disease. An important determinant of susceptibility is genomic variation. We undertook this study to define strains of mice with different susceptibilities for the development of smoking-induced emphysema because they could help identify genetic factors of susceptibility. NZWLac/J, C57BL6/J, A/J, SJ/L, and AKR/J strains were exposed to cigarette smoke for 6 months. Elastance (Htis), the extent of emphysema (mean linear intercept [Lm]), and the inflammatory cell and cytokine response were measured. NZWLac/J had no change in Lm or Htis (resistant). C57BL6/J, A/J, and SJ/L increased Lm, but not Htis (mildly susceptible). AKR/J increased Lm and Htis (super-susceptible). Only AKR/J had significant inflammation comprising macrophages, neutrophils, and T cells. The AKR/J showed an upregulation of Th1 cytokines whereas in the C57BL/6/J and NZWlac/J, cytokines did not change or were downregulated. We conclude that Lm, elastance, and inflammation are features that are needed to phenotype emphysema in mice. The inflammatory cell and cytokine profile may be an important determinant of the phenotype in response to cigarette smoke exposure. The identification of resistant and susceptible strains for the development of emphysema could be useful for genomic studies of emphysema susceptibility in mice and eventually in humans. PMID- 15282204 TI - Alternative methods of titrating continuous positive airway pressure: a large multicenter study. AB - Standard practice for continuous positive airway pressure (CPAP) treatment in sleep apnea and hypopnea syndrome (SAHS) requires pressure titration during attended laboratory polysomnography. However, polysomnographic titration is expensive and time-consuming. The aim of this study was to ascertain, in a large sample of CPAP-naive patients, whether CPAP titration performed by an unattended domiciliary autoadjusted CPAP device or with a predicted formula was as effective as CPAP titration performed by full polysomnography. The main outcomes were the apnea-hypopnea index and the subjective daytime sleepiness. We included 360 patients with SAHS requiring CPAP treatment. Patients were randomly allocated into three groups: standard, autoadjusted, and predicted formula titration with domiciliary adjustment. The follow-up period was 12 weeks. With CPAP treatment, the improvement in subjective sleepiness and apnea-hypopnea index was very similar in the three groups. There were no differences in the objective compliance of CPAP treatment and in the dropout rate of the three groups at the end of the follow-up. Autoadjusted titration at home and predicted formula titration with domiciliary adjustment can replace standard titration. These procedures could lead to considerable savings in cost and to significant reductions in the waiting list. PMID- 15282205 TI - The expression of human mitochondrial ferritin rescues respiratory function in frataxin-deficient yeast. AB - Mitochondrial ferritin (MtF) is structurally and functionally similar to the cytosolic ferritins, molecules designed to store and detoxify cellular iron. MtF expression in human and mouse is restricted to the testis and few tissues, and it is abundant in the erythroblasts of patients with sideroblastic anemia, where it is thought to protect the mitochondria from the damage caused by iron loading. Mitochondria iron overload occurs also in cells deficient in frataxin, a mitochondrial protein involved in iron handling and implicated in Friedreich ataxia. We expressed human MtF in frataxin-deficient yeast cells, a well characterized model of mitochondrial iron overload and oxidative damage. The human MtF precursor was efficiently imported by yeast mitochondria and processed to functional ferritin that actively sequestered iron in the organelle. MtF expression rescued the respiratory deficiency caused by the loss of frataxin protecting the activity of iron-sulfur enzymes and enabling frataxin-deficient cells to grow on non-fermentable carbon sources. Furthermore, MtF expression prevented the development of mitochondrial iron overload, preserved mitochondrial DNA integrity and increased cell resistance to H2O2. The data show that MtF can substitute for most frataxin functions in yeast, suggesting that frataxin is directly involved in mitochondrial iron-binding and detoxification. PMID- 15282206 TI - A common haplotype at the CD36 locus is associated with high free fatty acid levels and increased cardiovascular risk in Caucasians. AB - CD36 is a class B scavenger receptor recognizing a variety of ligands including long-chain fatty acids and modified LDL. We investigated whether genetic variability at this locus is a determinant of free fatty acid (FFA) plasma levels and risk of coronary artery disease (CAD) in Caucasians. Typing of 21 polymorphic markers, evenly spanning the CD36 gene, revealed two linkage disequilibrium (LD) blocks that could be tagged by five polymorphisms (-33137A>G, -31118G>A, 25444G>A, 27645del>ins and 30294G>C). In 585 non-diabetic individuals of Caucasian origin, the 30294G>C polymorphism was significantly associated with FFA levels (P = 0.02)--an effect that was especially visible among men (P = 0.009). A similar association was observed in this gender at -33137 (P = 0.008) and -31118 (P = 0.028). When the five tag polymorphisms were considered together, men carrying the AGGIG haplotype had 31% higher FFA (P = 0.0002) and 20% higher triglycerides (P = 0.025) than non-carriers. The same haplotype was associated with increased risk of CAD in 197 type 2 diabetic individuals from the US (OR = 2.3, 95% CI 1.2-4.2). A similar tendency was observed in a group of 321 type 2 diabetic individuals from Italy (OR = 1.4, 0.9-2.3), resulting in an overall relative risk of 1.6 (1.1-2.3, P = 0.015) in the two populations considered together. By targeted resequencing, we identified a common variant in the CD36 promoter that is in strong LD with the AGGIG haplotype and could be partly responsible for these findings. In conclusion, this comprehensive study of CD36 variability indicates that the common polymorphisms at this locus modulate lipid metabolism and cardiovascular risk in Caucasians. PMID- 15282207 TI - In vivo function of the conserved non-catalytic domain of Werner syndrome helicase in DNA replication. AB - Werner syndrome is a genetic disorder characterized by genomic instability, elevated recombination and replication defects. The WRN gene encodes a RecQ helicase whose function(s) in cellular DNA metabolism is not well understood. To investigate the role of WRN in replication, we examined its ability to rescue cellular phenotypes of a yeast dna2 mutant defective in a helicase-endonuclease that participates with flap endonuclease 1 (FEN-1) in Okazaki fragment processing. Genetic complementation studies indicate that human WRN rescues dna2 1 mutant phenotypes of growth, cell cycle arrest and sensitivity to the replication inhibitor hydroxyurea or DNA damaging agent methylmethane sulfonate. A conserved non-catalytic C-terminal domain of WRN was sufficient for genetic rescue of dna2-1 mutant phenotypes. WRN and yeast FEN-1 were reciprocally co immunoprecipitated from extracts of transformed dna2-1 cells. A physical interaction between yeast FEN-1 and WRN is demonstrated by yeast FEN-1 affinity pull-down experiments using transformed dna2-1 cells extracts and by ELISA assays with purified recombinant proteins. Biochemical analyses demonstrate that the C terminal domain of WRN or BLM stimulates FEN-1 cleavage of its proposed physiological substrates during replication. Collectively, the results suggest that the WRN-FEN-1 interaction is biologically important in DNA metabolism and are consistent with a role of the conserved non-catalytic domain of a human RecQ helicase in DNA replication intermediate processing. PMID- 15282209 TI - Phenotypic analysis of neurofilament light gene mutations linked to Charcot-Marie Tooth disease in cell culture models. AB - Mutations in the neurofilament light (NFL) gene cause Charcot-Marie-Tooth (CMT) disease. There is a wide range of clinical presentations in CMT patients harboring NFL mutations, with patients classified as CMT2E or CMT1F. In this study, we analyzed the effects of five NFL mutations on the assembly and intracellular distribution of intermediate filaments (IFs), and compared the results with those obtained previously for other NFL mutations. Although all NFL mutants affected the formation of IF networks, our data show differential effects on the assembly of IFs depending on the exact nature of the mutation. Defective transport of the mutant NFL subunits was observed for all the CMT-linked NFL mutations, but the characteristics of this defect also depended on the specific mutation. These results show that defects in the assembly and transport of NFs are common to all NFL mutants studied thus far, but the exact nature of the defect appears to be correlated with each mutant genotype. PMID- 15282208 TI - Biochemical analysis of pathogenic ligand-dependent FGFR2 mutations suggests distinct pathophysiological mechanisms for craniofacial and limb abnormalities. AB - Gain-of-function missense mutations in FGF receptor 2 (FGFR2) are responsible for a variety of craniosynostosis syndromes including Apert syndrome (AS), Pfeiffer syndrome (PS) and Crouzon syndrome (CS). Unlike the majority of FGFR2 mutations, S252W and P253R AS mutations and a D321A PS mutation retain ligand-dependency and are also associated with severe limb pathology. In addition, a recently identified ligand-dependent S252L/A315S double mutation in FGFR2 was shown to cause syndactyly in the absence of craniosynostosis. Here, we analyze the effect of the canonical AS mutations, the D321A PS mutation and the S252L/A315S double mutation on FGFR2 ligand binding affinity and specificity using surface plasmon resonance. Both AS mutations and the D321A PS mutation, but not the S252L/A315S double mutation, increase the binding affinity of FGFR2c to multiple FGFs expressed in the cranial suture. Additionally, all four pathogenic mutations also violate FGFR2c ligand binding specificity and enable this receptor to bind FGF10. Based on our data, we propose that an increase in mutant FGFR2c binding to multiple FGFs results in craniosynostosis, whereas binding of mutant FGFR2c to FGF10 results in severe limb pathology. Structural and biophysical analysis shows that AS mutations in FGFR2b also enhance and violate FGFR2b ligand binding affinity and specificity, respectively. We suggest that elevated AS mutant FGFR2b signaling may account for the dermatological manifestations of AS. PMID- 15282210 TI - Sialyltransferase mRNA abundances in B cells are strictly controlled, correlated with cognate lectin binding, and differentially responsive to immune signaling in vitro. AB - Mouse gene knockout studies have provided unimpeachable evidence of immune relevant functions for several sialyltransferase enzymes including ST6Gal I, ST3Gal I, and ST3Gal IV. Such studies cannot, however, identify cellular mechanisms for regulating such activities. In this article we provide evidence that murine B lymphocytes respond to specific immune signals in vitro with tightly regulated changes in the sialic acid composition of the cell surface glycocalyx. These changes are both quantitative and qualitative in nature and are apparently regulated at both the transcriptional and posttranscriptional levels. We used lectin binding and flow cytometry combined with relative real-time PCR to show that MAH and PNA binding are tightly correlated with the abundance of ST3Gal IV and ST3Gal I mRNA, respectively, under several different conditions of B cell stimulation. Finally, we show that although SNA binding and the expression of ST6Gal I coding sequence are not tightly correlated, there is a clear differential control of 5'UTR exon usage in response to different immune signals. PMID- 15282211 TI - Desmethoxyyangonin and dihydromethysticin are two major pharmacological kavalactones with marked activity on the induction of CYP3A23. AB - Kava kava (Piper methysticum), an herbal remedy, is widely used for the treatment of mild to moderate cases of anxiety. The therapeutic activity is presumably achieved through multiple constituents called kavalactones. Recently, kava extracts were shown to induce CYP3A4 and activate human pregnane X receptor (PXR). This study was undertaken to test the ability of purified kavalactones to induce CYP3A23 and activate PXR. Rat hepatocytes were treated with desmethoxyyangonin, dihydrokawain, dihydromethysticin, kawain, methysticin, or yangonin, and the expression of CYP3A23 was monitored. Among the kavalactones, only desmethoxyyangonin and dihydromethysticin markedly induced the expression of CYP3A23 (approximately 7-fold). A similar magnitude of induction was detected with combined six kavalactones at a noninductive concentration when individually used. The induced expression, however, was markedly reduced or completely abolished if dihydromethysticin, desmethoxyyangonin, or both were excluded from the mixtures. Interestingly, regardless of whether dihydromethysticin or desmethoxyyangonin was used alone or together with other kavalactones, similar amounts of total kavalactones were needed to produce comparable induction, suggesting that the inductive activity of dihydromethysticin and desmethoxyyangonin is additively/synergistically enhanced by other kavalactones. In addition, treatment with dihydromethysticin, desmethoxyyangonin, or pregnenolone 16alpha-carbonitrile (PCN) markedly increased the levels of CYP3A23 mRNA, and inhibition of mRNA synthesis abolished the induction. In contrast to PCN, dihydromethysticin and desmethoxyyangonin only slightly activated rat or human PXR. These findings suggest that the induction of CYP3A23 by dihydromethysticin and desmethoxyyangonin involves transcription activation, probably through a PXR-independent or PXR-involved indirect mechanism. PMID- 15282212 TI - Tissue distribution, stability, and pharmacokinetics of Apo2 ligand/tumor necrosis factor-related apoptosis-inducing ligand in human colon carcinoma COLO205 tumor-bearing nude mice. AB - Apo2L/TRAIL [Apo2 ligand/tumor necrosis factor (TNF)-related apoptosis-inducing ligand], a member of the TNF cytokine superfamily, induces cell death by apoptosis in a number of human cancer cells and is a potential agent for cancer therapy. We have characterized the in vitro stability of Apo2L/TRAIL in human serum and the tissue distribution and metabolism of Apo2L/TRAIL in a xenograft model of human colon carcinoma (COLO205). Apo2L/TRAIL was stable after incubation in human serum, with no significant high molecular weight complexes or degradation products observed. After i.v. administration of 125I-Apo2L/TRAIL to mice, a small percentage of the radiolabeled drug was seen as high molecular weight complex or as low molecular weight degradation products in plasma. However, the most abundant radioactive species corresponded to the intact Apo2L/TRAIL monomer, indicative of the relative stability of this recombinant protein in blood. Distribution of 125I-Apo2L/TRAIL to organs and solid xenograft tumors was limited. Intact 125I-Apo2L/TRAIL was detectable in the solid tumor at all time points and was the only tissue in which radioactivity transiently increased over time. Kidney contained the highest levels of radioactivity. Radioactive signal reached a tissue-to-blood ratio of 18 in the kidney cortex region when 125I-Apo2L/TRAIL was given in the presence of excess unlabeled ligand. In contrast to blood, extensive 125I-Apo2L/TRAIL degradation was observed in the kidney and, to a lesser degree, in the solid tumor and other organs, including liver, spleen, and lung. Our studies demonstrated that Apo2L/TRAIL is stable in the circulation, localizes to human solid xenograft tumors, and is primarily eliminated through the kidney. PMID- 15282213 TI - Autonomic asymmetry in migraine: augmented parasympathetic activation in left unilateral migraineurs. AB - Brain autonomic control is asymmetrical, the left hemisphere affecting predominantly parasympathetic function and the right hemisphere affecting predominantly sympathetic function. It is not known whether the extent of autonomic activation is altered in migraine, although the fact that some migraineurs express parasympathetic features such as facial flushing, lacrimation and rhinorrhoea might suggest increased parasympathetic activation. We instilled diluted soapy eyedrops and measured (i) the trigemino-parasympathetic reflex by the vasodilator response of forehead skin bilaterally using photoplethysmography; (ii) the somato-sympathetic reflex by vasoconstriction in the index finger; and (iii) heart rate response. We studied 14 left-sided and 15 right-sided unilateral migraineurs outside attacks. We found that left-side migraineurs had significantly higher bilateral parasympathetic vasodilatation, regardless of the stimulation or measurement side (+60.1 +/- 6.4%) compared with right-side migraineurs (+41.9 +/- 6.4%, P < 0.05). Sympathetic vasoconstriction, however, was similar for the two groups (left, -15.9 +/- 4.2%; right, -17.7 +/- 4.1%, NS). Bradycardia was significantly more pronounced for the left-side migraineurs (interbeat, RR interval increase of +6.2 +/- 1.1% versus +3.1 +/- 1.1%, P < 0.04). We conclude that unilateral left-side migraineurs have increased parasympathetic activation in response to pain compared with right-side migraineurs. Sympathetic responses were similar in the two groups and seemed not to be affected by migraine side. Since cranial parasympathetic activity induces cerebral vasodilatation, this augmentation might be an inherent part of the migraine pathophysiology in these patients. PMID- 15282214 TI - Does anticipation of back pain predispose to back trouble? AB - Limb movement imparts a perturbation to the body. The impact of that perturbation is limited via anticipatory postural adjustments. The strategy by which the CNS controls anticipatory postural adjustments of the trunk muscles during limb movement is altered during acute back pain and in people with recurrent back pain, even when they are pain free. The altered postural strategy probably serves to protect the spine in the short term, but it is associated with a cost and is thought to predispose spinal structures to injury in the long term. It is not known why this protective strategy might occur even when people are pain free, but one possibility is that it is caused by the anticipation of back pain. In eight healthy subjects, recordings of intramuscular EMG were made from the trunk muscles during single and repetitive arm movements. Anticipation of experimental back pain and anticipation of experimental elbow pain were elicited by the threat of painful cutaneous stimulation. There was no effect of anticipated experimental elbow pain on postural adjustments. During anticipated experimental back pain, for single arm movements there was delayed activation of the deep trunk muscles and augmentation of at least one superficial trunk muscle. For repetitive arm movements, there was decreased activity and a shift from biphasic to monophasic activation of the deep trunk muscles and increased activity of superficial trunk muscles during anticipation of back pain. In both instances, the changes were consistent with adoption of an altered strategy for postural control and were similar to those observed in patients with recurrent back pain. We conclude that anticipation of experimental back pain evokes a protective postural strategy that stiffens the spine. This protective strategy is associated with compressive cost and is thought to predispose to spinal injury if maintained long term. PMID- 15282215 TI - Astrocytic degeneration relates to the severity of disease in frontotemporal dementia. AB - The main unifying feature of cases with frontotemporal dementia (FTD) is the pattern of brain atrophy. Surprisingly, there are a variety of underlying histopathologies in cases with the clinical features and typical pattern of atrophy characterizing FTD. This suggests that the degenerative mechanism(s) associated with pyramidal cell loss and gliosis in FTD is likely to be similar in the different histopathological forms of the disease. In this study we tested this hypothesis by analysing a common cell death mechanism, apoptosis, in cases of FTD with either Pick's disease (PiD) (n = 9) or frontotemporal lobar degeneration (FTLD) (n = 7) compared with normal controls (n = 10). Tissue sections from previously analysed cases were stained using anti-activated caspase 3 immunohistochemistry, TUNEL, propidium iodide, and cell- and pathology-specific labels. These markers of apoptosis identified both astrocytes and neurons in regions vulnerable to degeneration in all cases of FTD. However, neuronal apoptosis was rare (<2% of neurons), even at early disease stages where there is considerably less frontotemporal atrophy or pyramidal cell loss. This suggests that other cell death mechanisms account for the progressive neuronal loss in FTD. In contrast, astrocytes with beaded processes and other apoptotic features were very frequent in both PiD and FTLD, with the severity of astrocytosis and astrocytic apoptosis correlating with both the degree of neuronal loss and the stage of disease. These findings provide evidence that astrocytic apoptosis occurs as an early event in different histopathological forms of FTD. Furthermore, this astrocytic apoptosis directly relates to the degree of degeneration in FTD, and becomes the overwhelming pathological feature as the disease progresses. PMID- 15282216 TI - Cognitive dysfunction in patients with cerebral microbleeds on T2*-weighted gradient-echo MRI. AB - Gradient echo T2*-weighted MRI has high sensitivity in detecting cerebral microbleeds, which appear as small dot-like hypointense lesions. Microbleeds are strongly associated with intracerebral haemorrhage, hypertension, lacunar stroke and ischaemic small vessel disease, and have generated interest as a marker of bleeding-prone microangiopathy. Microbleeds have generally been considered to be clinically silent; however, since they are located in widespread cortical and basal ganglia regions and are histologically characterized by tissue damage, we hypothesized that they would cause cognitive dysfunction. We therefore studied patients with microbleeds (n = 25) and a non-microbleed control group (n = 30) matched for age, gender and intelligence quotient. To avoid the confounding effects of coexisting cerebrovascular disease, the groups were also matched for the extent of MRI-visible white matter changes of presumed ischaemic origin, location of cortical strokes, and for the proportion of patients with different stroke subtypes (including lacunar stroke). A battery of neuropsychological tests was used to assess current intellectual function, verbal and visual memory, naming and perceptual skills, speed and attention and executive function. Microbleeds were most common in the basal ganglia but were also found in frontal, parieto-occipital, temporal and infratentorial regions. There was a striking difference between the groups in the prevalence of executive dysfunction, which was present in 60% of microbleed patients compared with 30% of non-microbleed patients (P = 0.03). Logistic regression confirmed that microbleeds (but not white matter changes) were an independent predictor of executive impairment (adjusted odds ratio = 1.32, 95% confidence interval 1.01-1.70, P = 0.04). Patients with executive dysfunction had more microbleeds in the frontal region (mean count 1.54 versus 0.03; P = 0.002) and in the basal ganglia (mean 1.17 versus 0.32; P = 0.048). There was a modest correlation between the number of microbleeds and the number of cognitive domains impaired (r = 0.44, P = 0.03). This study provides novel evidence that microbleeds are associated with cognitive dysfunction, independent of the extent of white matter changes of presumed ischaemic origin, or the presence of ischaemic stroke. The striking effect of microbleeds on executive dysfunction is likely to result from associated tissue damage in the frontal lobes and basal ganglia. These findings have implications for the diagnosis of stroke patients with cognitive impairment, and for the appropriate use of antihypertensive and antiplatelet treatments in these patients. PMID- 15282217 TI - MRI-negative PET-positive temporal lobe epilepsy: a distinct surgically remediable syndrome. AB - Most patients with non-lesional temporal lobe epilepsy (NLTLE) will have the findings of hippocampal sclerosis (HS) on a high resolution MRI. However, a significant minority of patients with NLTLE and electroclinically well lateralized temporal lobe seizures have no evidence of HS on MRI. Many of these patients have concordant hypometabolism on fluorodeoxyglucose-PET ([18F]FDG-PET). The pathophysiological basis of this latter group remains uncertain. We aimed to determine whether NLTLE without HS on MRI represents a variant of or a different clinicopathological syndrome from that of NLTLE with HS on MRI. The clinical, EEG, [18F]FDG-PET, histopathological and surgical outcomes of 30 consecutive NLTLE patients with well-lateralized EEG but without HS on MRI (HS-ve TLE) were compared with 30 consecutive age- and sex-matched NLTLE patients with well lateralized EEG with HS on MRI (HS+ve TLE). Both the HS+ve TLE group and the HS ve TLE patients had a high degree of [18F]FDG-PET concordant lateralization (26 out of 30 HS-ve TLE versus 27 out of 27 HS+ve TLE). HS-ve TLE patients had more widespread hypometabolism on [18F]FDG-PET by blinded visual analysis [odds ratio (OR = + infinity (2.51, -), P = 0.001]. The HS-ve TLE group less frequently had a history of febrile convulsions [OR = 0.077 (0.002-0.512), P = 0.002], more commonly had a delta rhythm at ictal onset [OR = 3.67 (0.97-20.47), P = 0.057], and less frequently had histopathological evidence of HS [OR = 0 (0-0.85), P = 0.031]. There was no significant difference in surgical outcome despite half of those without HS having a hippocampal-sparing procedure. Based on the findings outlined, HS-ve PET-positive TLE may be a surgically remediable syndrome distinct from HS+ve TLE, with a pathophysiological basis that primarily involves lateral temporal neocortical rather than mesial temporal structures. PMID- 15282218 TI - Clinicopathological study of a myelin oligodendrocyte glycoprotein-induced demyelinating disease in LEW.1AV1 rats. AB - Although multiple sclerosis is considered to be an autoimmune disease in the CNS, the immune responses that take place in the CNS and lymphoid organs remain to be elucidated. Here, we have successfully induced various subtypes of experimental autoimmune encephalitis (EAE) in LEW.1AV1 rats carrying RT1(av1) on the Lewis background genes by immunization with recombinant rat myelin oligodendrocyte glycoprotein (MOG) in various solutions with adjuvants. The purpose of the present study was to analyse in more detail the clinical and immunopathological features of MOG-induced EAE in LEW.1AV1 rats. Immunization with high doses of soluble MOG with pertussis toxin induced acute, frequently fatal EAE, whereas medium doses of partially aggregated MOG without pertussis toxin produced relapsing and remitting EAE. Secondary progressive EAE was induced in some rats by immunization with the immunization protocol having an intermediate nature between the above two. The optic nerve (approximately 60% of the immunized rats) and spinal cord (100%) were frequently involved and detectable both clinically and pathologically, while there was no lesion in the cerebrum. Histological examination revealed that, despite variety in the clinical subtypes, progression of the pathological processes was strikingly uniform, i.e. initial inflammation with minimal demyelination followed by predominant demyelination with minimal lymphocyte infiltration. These findings suggest that the lesion during the later stage is maintained by humoral factors. Taken together, this experimental system can serve as a model of neuromyelitis optica. Further analysis will provide useful information to elucidate the pathogenesis and to develop immunotherapy for neuromyelitis optica and multiple sclerosis. PMID- 15282219 TI - Health by association? Social capital, social theory, and the political economy of public health. AB - Three perspectives on the efficacy of social capital have been explored in the public health literature. A "social support" perspective argues that informal networks are central to objective and subjective welfare; an "inequality" thesis posits that widening economic disparities have eroded citizens' sense of social justice and inclusion, which in turn has led to heightened anxiety and compromised rising life expectancies; a "political economy" approach sees the primary determinant of poor health outcomes as the socially and politically mediated exclusion from material resources. A more comprehensive but grounded theory of social capital is presented that develops a distinction between bonding, bridging, and linking social capital. It is argued that this framework helps to reconcile these three perspectives, incorporating a broader reading of history, politics, and the empirical evidence regarding the mechanisms connecting types of network structure and state-society relations to public health outcomes. PMID- 15282220 TI - Adult mortality: time for a reappraisal. AB - BACKGROUND: In many countries, little is known about adult mortality rates. New innovations are necessary to develop reasonable estimates from available information. One readily available resource is household survey data. While birth histories collected in surveys have produced reasonable estimates of child mortality, the use of sibling survival data collected in similar household surveys has not been comprehensively analysed, largely because of concerns of underreporting. METHODS: This paper uses sibling survival schedules from 29 Demographic and Health Surveys (DHS) to generate estimates of under-5 mortality and of the summary measure of adult mortality 45q15-the probability of dying between ages 15 and 59. These are then compared with UN child and adult mortality estimates. RESULTS: Sibling history data collected in these household surveys seems to contain adequate information to estimate adult mortality rates, though there are problems with underreporting. The correlation coefficient between UN estimates and DHS estimates is 0.74 for adult mortality, indicating a strong relationship between the two but suggesting there may be underreporting of adult deaths in the survey data. CONCLUSIONS: Further investigation is necessary to determine the usefulness of household survey data for the estimation of adult mortality. New survey instruments like the World Health Survey have incorporated questions to help correct for underreporting in sibling histories. Further analyses need to be carried out in countries where vital registration data are also available, to determine how well household survey data do in estimating adult mortality and whether improvements in the survey instrument adequately correct for underreporting of deaths. PMID- 15282222 TI - Commentary: Reconciling the three accounts of social capital. PMID- 15282223 TI - Commentary: Is capital the solution or the problem? PMID- 15282224 TI - Commentary: Can subtle refinements of popular concepts be put into practice? PMID- 15282225 TI - Commentary: Social capital, social class, and the slow progress of psychosocial epidemiology. PMID- 15282226 TI - Commentary: "Health by association": some comments. PMID- 15282227 TI - Practical directions to the preservation of health. PMID- 15282228 TI - Preservation of health. 1830. PMID- 15282229 TI - Commentary: On reappraisal of adult mortality. PMID- 15282230 TI - Commentary: Social capital, social epidemiology and disease aetiology. PMID- 15282231 TI - Novel gyrA and parC point mutations in two strains of Acinetobacter baumannii resistant to ciprofloxacin. PMID- 15282232 TI - Optimizing antibiotic prescribing for acute cough in general practice: a cluster randomized controlled trial. AB - OBJECTIVES: To assess the effect of a tailored professional intervention, including academic detailing, on antibiotic prescribing for acute cough. METHODS: In a cluster-randomized controlled before and after study 85 Flemish GPs included adult patients with acute cough consulting in the periods February-April 2000 and 2001. The intervention consisted of a clinical practice guideline for acute cough, an educational outreach visit and a postal reminder to support its implementation in January 2001. Antibiotic prescribing rates and patients' symptom resolution were the main outcome measures. RESULTS: Thirty-six of 42 GPs received the intervention and 35 of 43 GPs served as controls; 1503 patients were eligible for analysis. Only in the intervention group were patients less likely to receive antibiotics after the intervention [OR(adj) (95% CI)=0.56 (0.36 0.87)]. Prescribed antibiotics were also more in line with the guideline in the intervention group [1.90 (0.96-3.75)] and less expensive from the perspective of the National Sickness and Invalidity Insurance Institute [MD(adj) (95% CI)= Euro 6.89 [-11.77-(-2.02)]]. No significant differences were found between the groups for the time to symptom resolution. CONCLUSIONS: An (inter)actively delivered tailored intervention implementing a guideline for acute cough is successful in optimizing antibiotic prescribing without affecting patients' symptom resolution. Further research efforts should be devoted to cost-effectiveness studies of such interventions. PMID- 15282234 TI - There are no paradoxes of adherence and drug resistance to HIV antiretroviral therapy. PMID- 15282233 TI - Which agents should we use for the treatment of multidrug-resistant Mycobacterium tuberculosis? AB - The inappropriate treatment of drug-susceptible tuberculosis can lead to the selection and transmission of multidrug-resistant tuberculosis (MDR-TB), indicating resistance to at least isoniazid and rifampicin. In the treatment of MDR-TB, residual first-line drugs, such as ethambutol, pyrazinamide and streptomycin must be appropriately combined with additional second-line drugs, guided by individual susceptibility patterns. The clinical pharmacology of these second-line antituberculous drugs is reviewed. Fluoroquinolones represent the only substantial therapeutic advance in the last 20 years. Many factors potentially affect the outcome of MDR-TB. Treatment adherence, prior exposure to antituberculous drugs, the number of drugs to which the infection is still susceptible and the time since the first diagnosis of tuberculosis are the most relevant. The management of MDR-TB requires considerable expertise. When initiating or revising therapy for MDR-TB, the process of selecting drugs should rely on prior treatment history, results of susceptibility testing and an evaluation of the patient's adherence. In making drug selection, we propose to follow a hierarchy based on the intrinsic activity against Mycobacterium tuberculosis and the clinical evidence of efficacy of the available active compounds. PMID- 15282235 TI - Susceptibility of capsular Staphylococcus aureus strains to some antibiotics, triclosan and cationic biocides. PMID- 15282236 TI - The rise and fall of triple nucleoside reverse transcriptase inhibitor (NRTI) regimens. AB - Triple nucleoside reverse transcriptase inhibitor (NRTI) regimens have attracted much interest due to their potential to (1) simplify dosing, with potential gains in adherence to treatment, and (2) reduce or even reverse dyslipidaemia associated with protease inhibitor (PI) therapy. A variety of triple NRTI combinations have been investigated, in both antiretroviral-naive and antiretroviral-experienced HIV-infected patients. Many of these trials have generated disappointing results, and some have been prematurely discontinued due to poor efficacy. This article reviews the background to the development of triple NRTI regimens, and the mounting evidence that this approach is suboptimal for antiretroviral-naive patients. Indeed, some triple NRTI regimens should never be used in this population. A role for triple NRTI combinations as a simplification strategy in treatment-experienced patients whose HIV is well controlled has been suggested, but emerging evidence indicates that such an approach can, under adequate selection pressure, lead to the emergence of mutations and viral load rebound. This commentary discusses the factors that appear to influence patients' responses to triple NRTI therapy, and their implications for patient selection. PMID- 15282237 TI - Release of calcium from intracellular stores and subsequent uptake by mitochondria are essential for the candidacidal activity of an N-terminal peptide of human lactoferrin. AB - OBJECTIVES: Earlier studies showed that mitochondrial damage is a hallmark of the candidacidal activity of an N-terminal peptide of human lactoferrin, further referred to as hLF(1-11). Since uptake of Ca(2+) by mitochondria may be essential for their activation, the aim of this study was to define the role of Ca(2+) in killing of Candida albicans by the hLF(1-11) peptide. METHODS: The effect of compounds interfering with Ca(2+) homeostasis on the hLF(1-11)-induced candidacidal activity, changes in mitochondrial membrane potential, and reactive oxygen species production were evaluated using a killing assay, rhodamine 123 staining, and 2',7'-dichlorofluorescein diacetate, respectively. The increase in cellular Ca(2+) content was measured using (45)Ca(2+). RESULTS: Our results revealed that Ruthenium Red, which inhibits the mitochondrial Ca(2+)-uniporter and the voltage-sensitive Ca(2+) release from internal stores, blocked (P<0.05) the hLF(1-11)-induced candidacidal activity as well as changes in the membrane potential of mitochondria, and reactive oxygen species production. Oxalate, which precipitates Ca(2+) in intracellular organelles, decreased (P<0.05) the peptide induced changes in the membrane potential of mitochondria, reactive oxygen species production, and candidacidal activity. Furthermore, the Ca(2+) ionophore ionomycin combined with high CaCl(2) concentrations enhanced the hLF(1-11) induced candidacidal activity. Moreover, hLF(1-11) caused an influx of Ca(2+) from the extracellular medium into C. albicans reaching a three-fold increase at 2 h, whereas no increase was found in unexposed cells. In agreement, the Ca(2+) chelator EGTA blocked the peptide-induced candidacidal activity. CONCLUSIONS: Ca(2+) release from intracellular stores, probably through subsequent mitochondrial Ca(2+) uptake, is essential for the hLF(1-11)-induced candidacidal activity. PMID- 15282238 TI - In vitro effects of nitazoxanide on Echinococcus granulosus protoscoleces and metacestodes. AB - OBJECTIVES: Infection of humans and domestic ruminants with the larval stage (metacestode) of Echinococcus granulosus results in cystic echinococcosis (CE). The metacestode causes a space-occupying lesion in visceral organs, most commonly in the liver. Benzimidazole carbamate derivatives, such as mebendazole and albendazole, are currently used for chemotherapeutic treatment of CE. In human patients, benzimidazoles have to be applied in high doses for extended periods of time, and adverse side effects are frequently observed. In order to evaluate alternative treatment options, the in vitro efficacy of nitazoxanide, a broad spectrum drug used against intestinal parasites and bacteria, was investigated. METHODS: Freshly isolated E. granulosus protoscoleces were subjected to nitazoxanide treatment (1, 5 and 10 microg/mL), and the effects on parasite viability were monitored by Trypan Blue staining and scanning electron microscopy. Protoscolex cultures were maintained further, until metacestode development took place. Metacestodes were then subjected to nitazoxanide treatment (10 microg/mL), and corresponding effects were visualized by scanning and transmission electron microscopy. RESULTS: Dose-dependent protoscolex death within a few days of nitazoxanide treatment was observed. Subsequent in vitro culture of drug-treated protoscoleces confirmed the non-viability of parasites, while further cultivation of non-treated protoscoleces for a period of at least 3 months resulted in stage conversion and the formation of small metacestodes 3-4 mm in diameter. Nitazoxanide had a deleterious effect on these metacestodes, which was comparable to that of albendazole. CONCLUSIONS: Our study indicates a potential for nitazoxanide as an alternative treatment option against CE. PMID- 15282239 TI - Mersacidin eradicates methicillin-resistant Staphylococcus aureus (MRSA) in a mouse rhinitis model. AB - OBJECTIVES: Methicillin-resistant Staphylococcus aureus (MRSA) often colonize the anterior nares, and nasal carriage remains the main source of bacterial dissemination. The aim of this study was to assess the in vivo activity of the lantibiotic mersacidin against MRSA colonizing nasal epithelia. METHODS: The efficiency of mersacidin in the eradication of MRSA was tested employing mice pre treated with hydrocortisone and inoculated intranasally either three or six times with a bacterial suspension. RESULTS: In mersacidin-treated animals, pre colonized with MRSA, bacteria could not be detected in blood, lungs, liver, kidney, spleen or nasal scrapings and there were no lesions manifested after intraperitoneal drug application. Blood samples from infected mice obtained 2 h after mersacidin therapy revealed anti-MRSA activity in a serum bactericidal test. Moreover, elevated interleukin-1beta and tumour necrosis factor-alpha titres were noticed in the pre-infected but not in cured animals. In contrast, mersacidin did not induce differences in the cytokine profiles of treated uninfected control mice. CONCLUSIONS: In the mouse rhinitis model, mersacidin was able to eradicate MRSA colonization. The site of action (epithelium versus blood) of mersacidin needs to be further explored. PMID- 15282240 TI - Frequency and diversity of Class A extended-spectrum beta-lactamases in hospitals of the Auvergne, France: a 2 year prospective study. AB - OBJECTIVES: To evaluate the frequency and diversity of extended-spectrum beta lactamases (ESBLs) produced by Enterobacteriaceae and Pseudomonas aeruginosa in one French region. METHODS: During 2001-2002, all the non-duplicate isolates of P. aeruginosa resistant to ceftazidime and of Enterobacteriaceae intermediate or resistant to ceftazidime and/or cefotaxime and/or aminoglycosides with an AAC(6') I phenotype were collected in nine hospitals of the area. ESBL isoelectric points were determined, bla genes were amplified and sequenced and epidemic isolates were genotyped with ERIC2-PCR. RESULTS: ESBLs were observed in 297 Enterobacteriaceae (0.8%). The most frequent were TEM-3 like (n=152; 51.2%) and TEM-24 (n=115; 38.7%). Four new enzymes were observed, TEM-112 (pI 5.4), TEM-113 (pI 6.3), TEM-114 (pI 5.9) and TEM-126 (pI 5.4). Other TEMs were TEM-8, TEM-12, TEM-16, TEM-19, TEM-20, TEM-21, TEM-29 and TEM-71. The other ESBLs were SHV-4, SHV-5 and SHV-12, CTX-M-1, CTX-M-3, CTX-M-14 and CTX-M-15. In 37 P. aeruginosa (0.7%) only one ESBL was observed, PER-1. Five epidemic strains were detected, Serratia marcescens TEM-3 and four observed in several hospitals, Enterobacter aerogenes TEM-24, Citrobacter koseri TEM-3, Proteus mirabilis TEM-3 and P. aeruginosa PER-1. CONCLUSION: ESBL frequency was lower than in 1998, and CTX-M type frequency higher (2.1% of ESBLs in 2001, 4.9% in 2002). This long-term survey detected new sporadic enzymes (TEM-112, TEM-113, TEM-114 and TEM-126) and interhospital epidemic strains while avoiding any overestimation of ESBL frequency that may otherwise have occurred because of acute epidemics. PMID- 15282241 TI - Cytomegalovirus infection in the era of HAART: fewer reactivations and more immunity. AB - The incidence of cytomegalovirus (CMV) disease, once the most common and highly feared viral complication of AIDS, has dramatically decreased with the advent of highly active antiretroviral therapy (HAART). HAART-associated changes in the epidemiology of CMV disease resulted from the increase in CMV-specific immune responses coupled with the decrease in CMV reactivation. However, CMV disease continues to afflict HIV-infected patients on HAART when CD4+ cell counts fail to rise above 100 cells/mm(3) and when reconstitution of normal CMV-specific immune responses does not occur. The latter scenario may lead to recurrent or de novo CMV end-organ disease, or to the recently described CMV immune recovery vitritis. HAART-associated immune reconstitution offers unique opportunities to investigate the virological and immunological correlates of protection against CMV disease. Although the full extent of CMV-specific immune reconstitution has not been defined thus far, CMV-specific interferon-gamma production has been shown to be significantly associated with protection against CMV reactivation and recurrent disease. PMID- 15282242 TI - Emergence of IMP-4 metallo-beta-lactamase in a clinical isolate from Australia. PMID- 15282244 TI - A new mosaic for women's health. PMID- 15282245 TI - Gender distribution of dentists in Nigeria, 1981 to 2000. AB - In Nigeria, modern dental practice is relatively recent. The dawn can be traced to 1915 when the first government dentist was employed in Lagos, then the country's capital. There are presently four dental schools in the country; each graduates an average of thirty dentists annually. The present study determined the trends in the gender distribution of dental practitioners over the twenty year period from 1981 to 2000 and used available data to project into the future. Data was collected from governmental and nongovernmental publications. The results indicate that there are now 2,598 licensed dentists serving the country's population of 123 million. A vast majority of these dentists work in urban centers, and only about 20 percent work in the rural areas where over 70 percent of Nigerians reside. There has been a male preponderance in the number of practicing dentists: only fifty-eight (15.3 percent) of 379 dental practitioners were female in 1981, though this figure has risen steadily to 35.1 percent of 2,598 dentists at the end of 2000. However, over the twenty-year period, the percentage of females was consistently higher among dental than medical practitioners. In 1981, for example, the percentage female was 15.0 among both dental and medical practitioners, but by the end of 2000 this had increased to 35.1 percent among dental practitioners and only 19.0 percent among medical practitioners. The imbalance in gender distribution of dental practitioners is steadily normalizing, and projections, using current trends, indicate that gender balance will be attained in the year 2015. PMID- 15282246 TI - Gender impact on the socioprofessional identification of women dentists in Bulgaria. AB - Women comprise 73 percent of all dentists in Bulgaria. Almost all of them started their careers as salaried, and now they are self-employed. The purpose of this study was to show how female dentists in Bulgaria met the challenges of the social and health reforms during the period of transition from totalitarian to democratic rule. A field sociological survey was carried out between October 1996 and June 1997, involving a sample of 842 dentists. The questionnaire was designed to give information on dental demography trends, pattern of participation and practice of female dentists, and their career development. The data were treated statistically using an SPSS package. Results show that for the period 1991-96 women dentists owned more than 50 percent of the newly opened private practices in Bulgaria. They were more interested in improving their qualifications and more successful in obtaining specialty status than male dentists. Dentists in Bulgaria by the late 1990 s, with no gender difference, tend to identify themselves as liberal practitioners within the pluralistic model of dental services delivery. The volume of services delivered and profile of women in professional practice and their social and professional mobility are highly competitive, despite the growing problems of maternity leave coverage and funding to re-enter the profession. PMID- 15282247 TI - Dental workforce issues in the United Kingdom. PMID- 15282248 TI - Challenges for leadership of oral health care in Finland. PMID- 15282249 TI - Challenges to the oral health workforce in India. PMID- 15282250 TI - Advancing leadership opportunities for women: the China and Hong Kong scenario. PMID- 15282251 TI - Curriculum and education: a working group report. PMID- 15282252 TI - Women and family health and oral health: a working group report. PMID- 15282253 TI - Practice and community: a working group report. PMID- 15282254 TI - An anatomy of success: proven action steps that lead to achievement. PMID- 15282255 TI - Networking: connecting the dots, building matrices. PMID- 15282256 TI - Marketing women's oral health: lessons from the world of business. PMID- 15282257 TI - Women leading change: the case for oral health. PMID- 15282258 TI - Reversal of hippocampal LTP by spontaneous seizure-like activity: role of group I mGluR and cell depolarization. AB - Memory impairment is a common consequence of epileptic seizures. The hippocampal formation is particularly prone to seizure-induced amnesia due to its prominent role in mnemonic processes. We used the isolated CA1 slice preparation to examine effects of seizure-like activity on hippocampal plasticity, long-term potentiation (LTP), and long-term depression (LTD). Repeated spontaneous ictal events, generated in the presence of antagonists of GABA(A) receptor function, led to a stepwise erasure of LTP (termed spontaneous depotentiation, SDP). SDP could be initiated at various stages of LTP consolidation (tested < or =120 min after the induction of LTP). Renewed tetanic stimulation re-established LTP. SDP was remarkably specific: baseline transmission and other forms of hippocampal plasticity, i.e., Ca(2+)-induced LTP and two forms of LTD [(RS)-3,5 dihydroxyphenyglycine (DHPG) mediated and low-frequency stimulation mediated] were not affected by the same type of seizure activity. SDP was blocked in the presence of the group I mGluR antagonist (S)-4-carboxyphenylglycine. The mGluR1 antagonist (S)-(+)-alpha-amino-methylbenzeneacetic acid blocked approximately 80%, the mGluR5-specific antagonist 2-methyl-6-(phenylethynyl)-pyridine approximately 30% of SDP. Most efficient implementation of SDP was observed during seizures in the combined presence of the group I mGluR agonist DHPG and the GABA(A) antagonist bicuculline. However, similar ictal activity generated in the presence of DHPG alone did not lead to SDP in the vast majority of recordings. Complete disinhibition and at least partial activation of group I mGluR were necessary conditions for the induction of SDP. The depotentiating pharmacological conditions were accompanied by tonic membrane depolarization of CA1 pyramidal cells. Since hyperpolarization (by negative current injection) prevented intracellular SDP under depotentiating pharmacological conditions and depolarization (by positive current injection) led to selective intracellular SDP in the non-depotentiating seizure protocol of DHPG, it is concluded that cell depolarization was a sufficient condition for seizure-like activity to reverse hippocampal LTP. PMID- 15282259 TI - Effects of chronic dorsal column lesions on pelvic viscerosomatic convergent medullary reticular formation neurons. AB - Single medullary reticular formation (MRF) neurons receive multiple somatovisceral convergent inputs originating from many different spinal and cranial nerves, including the pelvic nerve (PN), dorsal nerve of the penis (DNP), and the abdominal branches of the vagus. In a previous study, the input to MRF from the male genitalia was shown to be eliminated with chronic 30-day dorsal hemisection at the T8 spinal level. In this study, the effect of a smaller chronic lesion [dorsal column lesion (DCx)] on MRF neuronal responses was examined. Responses to bilateral electrical stimulation of the DNP remained. MRF neuronal responses to non-noxious (touch/stroke) levels of penile stimulation, however, were eliminated; only responses to noxious pinch remained. No differences were found for the number of neurons responding to noxious distention of the colon between the DCx and control groups. Although no differences were found across these groups for the percent MRF responses to vagal stimulation, the mean response latency for the DCx group was twice the sham-DCx/intact control group. Taken together, these results indicate that the MRF receives at least some of its input from the male genitalia via pathways located within the dorsal columns at the mid-thoracic spinal level. PMID- 15282260 TI - Spontaneous waves in the dentate gyrus of slices from the ventral hippocampus. AB - Spontaneous negative-going potentials occurring at an average frequency of 0.7 Hz were recorded from the dentate gyrus of slices prepared from the temporal hippocampus of young adult rats. These events (here termed "dentate waves") in several respects resembled the dentate spikes described for freely moving rats during immobile behaviors and slow-wave sleep. Action potentials were observed on the descending portion of the in vitro waves and, as expected from this, whole cell recordings established that the waves were composed of depolarizing currents. Dentate waves appeared to be locally generated within the granule cell layer and were greatly reduced by antagonists of AMPA-type glutamate receptors or by lesions to the entorhinal cortex. Simultaneous recordings indicated that the waves were often synchronized in the inner and outer blades of the dentate gyrus. Knife cuts through the perforant path and the commissural/associational system did not eliminate synchronization, leaving electrotonic propagation via gap junctions as its probable cause. In accord with this, cuts that separated the two blades of the dentate eliminated synchronization between them, and a compound that inhibits gap junctions reduced wave activity. Dentate waves were regularly accompanied by sharp waves in field CA3 and were reduced in size by the acetylcholinesterase inhibitor, physostigmine. It is hypothesized that dentate waves occur when spontaneous glutamate release from dentate afferents produces action potentials in neighboring granule cells that then summate electrotonically into a population event; once initiated, the waves propagate, again electrotonically, and thereby engage a significant portion of the granule cell population. PMID- 15282261 TI - Gamma oscillation maintains stimulus structure-dependent synchronization in cat visual cortex. AB - Visual cortical cells demonstrate both oscillation and synchronization, although the underlying causes and functional significance of these behaviors remain uncertain. We simultaneously recorded single-unit activity with microelectrode arrays in supragranular layers of area 17 of cats paralyzed and anesthetized with propofol and N(2)O. Rate-normalized autocorrelograms of 24 cells reveal bursting (100%) and gamma oscillation (63%). Renewal density analysis, used to explore the source of oscillation, suggests a contribution from extrinsic influences such as feedback. However, a bursting refractory period, presumably membrane-based, could also encourage oscillatory firing. When we investigated the source of synchronization for 60 cell pairs we found only moderate correlation of synchrony with bursts and oscillation. We did, nonetheless, discover a possible functional role for oscillation. In all cases of cross-correlograms that exhibited oscillation, the strength of the synchrony was maintained throughout the stimulation period. When no oscillation was apparent, 75% of the cell pairs showed decay in their synchronization. The synchrony between cells is strongly dependent on similar response onset latencies. We therefore propose that structured input, which yields tight organization of latency, is a more likely candidate for the source of synchronization than oscillation. The reliable synchrony at response onset could be driven by spatial and temporal correlation of the stimulus that is preserved through the earlier stages of the visual system. Oscillation then contributes to maintenance of the synchrony to enhance reliable transmission of the information for higher cognitive processing. PMID- 15282262 TI - Learning to control arm stiffness under static conditions. AB - We used a robotic device to test the idea that impedance control involves a process of learning or adaptation that is acquired over time and permits the voluntary control of the pattern of stiffness at the hand. The tests were conducted in statics. Subjects were trained over the course of 3 successive days to resist the effects of one of three different kinds of mechanical loads: single axis loads acting in the lateral direction, single axis loads acting in the forward/backward direction, and isotropic loads that perturbed the limb in eight directions about a circle. We found that subjects in contact with single axis loads voluntarily modified their hand stiffness orientation such that changes to the direction of maximum stiffness mirrored the direction of applied load. In the case of isotropic loads, a uniform increase in endpoint stiffness was observed. Using a physiologically realistic model of two-joint arm movement, the experimentally determined pattern of impedance change could be replicated by assuming that coactivation of elbow and double joint muscles was independent of coactivation of muscles at the shoulder. Moreover, using this pattern of coactivation control we were able to replicate an asymmetric pattern of rotation of the stiffness ellipse that was observed empirically. These findings are consistent with the idea that arm stiffness is controlled through the use of at least two independent co-contraction commands. PMID- 15282263 TI - Timing and laminar profile of eye-position effects on auditory responses in primate auditory cortex. AB - We examined effects of eye position on auditory cortical responses in macaques. Laminar current-source density (CSD) and multiunit activity (MUA) profiles were sampled with linear array multielectrodes. Eye position significantly modulated auditory-evoked CSD amplitude in 24/29 penetrations (83%), across A1 and belt regions; 4/24 cases also showed significant MUA AM. Eye-position effects occurred mainly in the supragranular laminae and lagged the co-located auditory response by, on average, 38 ms. Effects in A1 and belt regions were indistinguishable in amplitude, laminar profile, and latency. The timing and laminar profile of the eye-position effects suggest that they are not combined with auditory signals at a subcortical stage of the lemniscal auditory pathways and simply "fed-forward" into cortex. Rather, these effects may be conveyed to auditory cortex by feedback projections from parietal or frontal cortices, or alternatively, they may be conveyed by nonclassical feedforward projections through auditory koniocellular (calbindin positive) neurons. PMID- 15282264 TI - Estrogen regulation of the cytochrome P450 3A subfamily in humans. AB - This study examines the possible role of estrogen in regulating the expression of the human CYP3A subfamily: CYP3A4, CYP3A5, CYP3A7, and CYP3A43. To accomplish this goal, mRNA was quantified from human livers and endometrial samples, and total CYP3A protein levels were evaluated by Western immunoblot analysis of the liver samples. The human endometrial samples were from premenopausal and postmenopausal women. The premenopausal endometrium was either in the proliferative or secretory phase, whereas for the postmenopausal endometrium samples, the women had been treated with either a placebo or estropipate, an estrogen substitute. After analyses, CYP3A4 mRNA was shown to have lower hepatic expression in females than in males. In the endometrium, CYP3A4 and CYP3A43 are down-regulated by estrogen, whereas CYP3A5 is expressed at higher levels during the secretory phase. CYP3A7 was not detected in the endometrium. In addition, the CYP3A subfamily showed increased mRNA expression in the liver as age increased. The expression levels of total CYP3A protein and total CYP3A mRNA showed good correlation. Despite apparent regulation of CYP3A4 mRNA expression by estrogen, the effects of estrogen may be overshadowed by additional regulators of gene expression. PMID- 15282265 TI - Genetic variants of the human H+/dipeptide transporter PEPT2: analysis of haplotype functions. AB - PEPT2 is a high-affinity H+/dipeptide transporter expressed in kidney, brain, lung, and mammary gland. The physiological role of PEPT2 in kidney is to reabsorb small peptides generated by luminal peptidases. PEPT2 is also a transporter for peptide-like drugs such as penicillins and cephalosporins. We have conducted a haplotype analysis of 27 single nucleotide polymorphisms located in or near exons of the human gene encoding hPEPT2 (SLC15A2), using genotyping data from 247 genomic DNA samples from the Coriell collection. Our analysis reveals that hPEPT2 has a >6-kilobase sequence block with at least 10 abundant polymorphisms in almost complete linkage disequilibrium. As a result, only two main hPEPT2 variants exist (hPEPT2*1 and *2) with several phased amino acid substitutions, present in substantial frequencies in all ethnic groups tested. When expressed in Chinese hamster ovary cells, hPEPT2*1 and *2 displayed similar Vmax values for glycyl-sarcosine (Gly-Sar), but they differed significantly in their Km values (83 +/- 16 and 233 +/- 38 microM, respectively). Moreover, hPEPT2*1 and *2 differed in their pH sensitivity for H+/Gly-Sar transport. In addition, hPEPT2*1 and *2 generated varying levels of mRNA in nine heterozygous kidney tissue samples, including one allele expressing no detectable mRNA, suggesting the presence of cis-acting polymorphisms affecting transcription or mRNA processing. The results indicate that polymorphisms in the gene encoding hPEPT2 can alter substrate transport and therefore could affect drug disposition in vivo. PMID- 15282266 TI - The Drosophila receptor guanylyl cyclase Gyc76C is required for semaphorin-1a plexin A-mediated axonal repulsion. AB - Cyclic nucleotide levels within extending growth cones influence how navigating axons respond to guidance cues. Pharmacological alteration of cAMP or cGMP signaling in vitro dramatically modulates how growth cones respond to attractants and repellents, although how these second messengers function in the context of guidance cue signaling cascades in vivo is poorly understood. We report here that the Drosophila receptor-type guanylyl cyclase Gyc76C is required for semaphorin 1a (Sema-1a)-plexin A repulsive axon guidance of motor axons in vivo. Our genetic analyses define a neuronal requirement for Gyc76C in axonal repulsion. Additionally, we find that the integrity of the Gyc76C catalytic cyclase domain is critical for Gyc76C function in Sema-1a axon repulsion. Our results support a model in which cGMP production by Gyc76C facilitates Sema-1a-plexin A-mediated defasciculation of motor axons, allowing for the generation of neuromuscular connectivity in the developing Drosophila embryo. PMID- 15282268 TI - Blockade of nociceptin/orphanin FQ receptor signaling in rat substantia nigra pars reticulata stimulates nigrostriatal dopaminergic transmission and motor behavior. AB - A multidisciplinary approach was followed to investigate whether the opioid-like peptide nociceptin/orphanin FQ (N/OFQ) regulates the nigrostriatal dopaminergic pathway and motor behavior. Nigrostriatal dopaminergic cells, which express N/OFQ peptide (NOP) receptors, are located in the substantia nigra pars compacta and extend their dendrites in the substantia nigra pars reticulata, thereby modulating the basal ganglia output neurons. In vitro electrophysiological recordings demonstrated that N/OFQ hyperpolarized the dopaminergic cells of the substantia nigra pars compacta and inhibited their firing activity. In vivo dual probe microdialysis showed that N/OFQ perfused in the substantia nigra pars reticulata reduced dopamine release in the ipsilateral striatum, whereas UFP-101 ([Nphe1,Arg14,Lys15]N/OFQ(1-13)-NH2) (a selective NOP receptor peptide antagonist) stimulated it. N/OFQ microinjected in the substantia nigra pars reticulata impaired rat performance on a rotarod apparatus, whereas UFP-101 enhanced it. Electromyography revealed that N/OFQ and UFP-101 oppositely affected muscle tone, inducing relaxation and contraction of triceps, respectively. The selective NOP receptor nonpeptide antagonist J-113397 (1-[3R,4R)-1 cyclooctylmethyl-3-hydroxymethyl-4-piperidyl]-3-ethyl-1,3-dihydro-2H benzimidazol 2-one), either injected intranigrally or given systemically, also elevated striatal dopamine release and facilitated motor activity, confirming that these effects were caused by blockade of endogenous N/OFQ signaling. The inhibitory role played by endogenous N/OFQ on motor activity was additionally strengthened by the finding that mice lacking the NOP receptor gene outperformed wild-type mice on the rotarod. We conclude that NOP receptors in the substantia nigra pars reticulata, activated by endogenous N/OFQ, drive a physiologically inhibitory control on motor behavior, possibly via modulation of the nigrostriatal dopaminergic pathway. PMID- 15282267 TI - Transactivation of Trk neurotrophin receptors by G-protein-coupled receptor ligands occurs on intracellular membranes. AB - Neurotrophins, such as NGF and BDNF, activate Trk receptor tyrosine kinases through receptor dimerization at the cell surface followed by autophosphorylation and intracellular signaling. It has been shown that activation of Trk receptor tyrosine kinases can also occur via a G-protein-coupled receptor (GPCR) mechanism, without involvement of neurotrophins. Two GPCR ligands, adenosine and pituitary adenylate cyclase-activating polypeptide (PACAP), can activate Trk receptor activity to increase the survival of neural cells through stimulation of Akt activity. To investigate the mechanism of Trk receptor transactivation, we have examined the localization of Trk receptors in PC12 cells and primary neurons after treatment with adenosine agonists and PACAP. In contrast to neurotrophin treatment, Trk receptors were sensitive to transcriptional and translational inhibitors, and they were found predominantly in intracellular locations particularly associated with Golgi membranes. Biotinylation and immunostaining experiments confirm that most of the transactivated Trk receptors are found in intracellular membranes. These results indicate that there are alternative modes of activating Trk receptor tyrosine kinases in the absence of neurotrophin binding at the cell surface and that receptor signaling may occur and persist inside of neuronal cells. PMID- 15282269 TI - Slow actions of neuroactive steroids at GABAA receptors. AB - Neuroactive steroids are potent and efficacious modulators of GABA(A) receptor activity and are potent sedatives and anesthetics. These positive modulators of GABA(A) receptors both potentiate the actions of GABA at the receptor and, at higher concentrations, directly gate the channel. The contribution of direct gating to the cellular and behavioral effects of neuroactive steroids is considered of little significance because it has been generally found that concentrations well above those needed for anesthesia are required to gate channels. By studying solitary glutamatergic neurons devoid of synaptic GABA input, we show that direct gating occurs and significantly alters membrane excitability at concentrations < or =100 nm. We propose that the relevance of direct gating has been overlooked partly because of the extremely slow kinetics of receptor activation and deactivation. We show that slow deactivation of directly gated currents does not result from an inherently tight ligand-receptor interaction because the slow deactivation is markedly accelerated by gamma cyclodextrin application. We hypothesize that steroids access the relevant GABA(A) receptor site from a non-aqueous reservoir, likely the plasma membrane, and that it is slow reservoir accumulation and departure that accounts for the slow kinetics of receptor gating by neuroactive steroids. PMID- 15282270 TI - External tufted cells: a major excitatory element that coordinates glomerular activity. AB - The glomeruli of the olfactory bulb are the first site of synaptic processing in the olfactory system. The glomeruli contain three types of neurons that are referred to collectively as juxtaglomerular (JG) cells: external tufted (ET), periglomerular (PG), and short axon (SA) cells. JG cells are thought to interact synaptically, but little is known about the circuitry linking these neurons or their functional roles in olfactory processing. Single and paired whole-cell recordings were performed to investigate these questions. ET cells spontaneously fired rhythmic spike bursts in the theta frequency range and received monosynaptic olfactory nerve (ON) input. In contrast, all SA and most PG cells lacked monosynaptic ON input. PG and SA cells exhibited spontaneous, intermittent bursts of EPSCs that were highly correlated with spike bursts of ET cells in the same but not in different glomeruli. Paired recording experiments demonstrated that ET cells provide monosynaptic excitatory input to PG/SA cells; the ET to PG/SA cell synapse is mediated by glutamate. ET cells thus are a major excitatory linkage between ON input and other JG cells. Spontaneous bursting is highly correlated among ET cells of the same glomerulus, and ET cell activity remains correlated when all fast synaptic activity is blocked. The findings suggest that multiple, synchronously active ET cells synaptically converge onto single PG/SA cells. Synchronous ET cell bursting may function to amplify transient sensory input and coordinate glomerular output. PMID- 15282271 TI - cAMP response element-binding protein is required for stress but not cocaine induced reinstatement. AB - Reinstatement of previously extinguished conditioned place preference (CPP) is precipitated by stress or drug exposure. Here, we show that acute exposure to forced swim stress (FS), in a context distinct from conditioning, induces reinstatement of cocaine CPP in wild-type mice. This behavior is accompanied by a pattern of phosphorylated cAMP response element-binding protein (pCREB) activation in discrete brain regions that is distinct from the pattern observed after cocaine-induced reinstatement. For example, previous cocaine conditioning increases pCREB levels in the amygdala, and acute exposure to FS, but not to cocaine, further augments these changes. In contrast, previous cocaine conditioning does not alter levels of pCREB in the nucleus accumbens, but acute exposure to FS increases pCREB levels in this region on reinstatement day. Furthermore, to determine whether these alterations of CREB are necessary in FS or cocaine-induced reinstatement, we examined the effect of these stimuli on reinstatement behavior in mice deficient in alpha and Delta isoforms of CREB. The CREB(alphaDelta) mutant mice show deficits in FS-induced reinstatement of conditioned place preference. In contrast, they show robust cocaine-induced reinstatement. This deficit in stress but not drug-induced reinstatement indicates a specific requirement for CREB in stress-induced behavioral responses to drugs of abuse. PMID- 15282272 TI - Motoneurons express heteromeric TWIK-related acid-sensitive K+ (TASK) channels containing TASK-1 (KCNK3) and TASK-3 (KCNK9) subunits. AB - Background potassium currents carried by the KCNK family of two-pore-domain K+ channels are important determinants of resting membrane potential and cellular excitability. TWIK-related acid-sensitive K+ 1 (TASK-1, KCNK3) and TASK-3 (KCNK9) are pH-sensitive subunits of the KCNK family that are closely related and coexpressed in many brain regions. There is accumulating evidence that these two subunits can form heterodimeric channels, but this evidence remains controversial. In addition, a substantial contribution of heterodimeric TASK channels to native currents has not been unequivocally established. In a heterologous expression system, we verified formation of heterodimeric TASK channels and characterized their properties; TASK-1 and TASK-3 were coimmunoprecipitated from membranes of mammalian cells transfected with the channel subunits, and a dominant negative TASK-1(Y191F) construct strongly diminished TASK-3 currents. Tandem-linked heterodimeric TASK channel constructs displayed a pH sensitivity (pK approximately 7.3) in the physiological range closer to that of TASK-1 (pK approximately 7.5) than TASK-3 (pK approximately 6.8). On the other hand, heteromeric TASK channels were like TASK-3 insofar as they were activated by high concentrations of isoflurane (0.8 mm), whereas TASK-1 channels were inhibited. The pH and isoflurane sensitivities of native TASK-like currents in hypoglossal motoneurons, which strongly express TASK-1 and TASK-3 mRNA, were best represented by TASK heterodimeric channels. Moreover, after blocking homomeric TASK-3 channels with ruthenium red, we found a major component of motoneuronal isoflurane-sensitive TASK-like current that could be attributed to heteromeric TASK channels. Together, these data indicate that TASK-1 and TASK 3 subunits coassociate in functional channels, and heteromeric TASK channels provide a substantial component of background K(+) current in motoneurons with distinct modulatory properties. PMID- 15282273 TI - Voltage imaging from dendrites of mitral cells: EPSP attenuation and spike trigger zones. AB - To obtain a more complete description of individual neurons, it is necessary to complement the electrical patch pipette measurements with technologies that permit a massive parallel recording from many sites on neuronal processes. This can be achieved by using voltage imaging with intracellular dyes. With this approach, we investigated the functional structure of a mitral cell, the principal output neuron in the rat olfactory bulb. The most significant finding concerns the characteristics of EPSPs at the synaptic sites and surprisingly small attenuation along the trunk of the primary dendrite. Also, the experiments were performed to determine the number, location, and stability of spike trigger zones, the excitability of terminal dendritic branches, and the pattern and nature of spike initiation and propagation in the primary and secondary dendrites. The results show that optical data can be used to deduce the amplitude and shape of the EPSPs evoked by olfactory nerve stimulation at the site of origin (glomerular tuft) and to determine its attenuation along the entire length of the primary dendrite. This attenuation corresponds to an unusually large mean apparent "length constant" of the primary dendrite. Furthermore, the images of spike trigger zones showed that an action potential can be initiated in three different compartments of the mitral cell: the soma-axon region, the primary dendrite trunk, and the terminal dendritic tuft, which appears to be fully excitable. Finally, secondary dendrites clearly support the active propagation of action potentials. PMID- 15282274 TI - Lipid rafts mediate the synaptic localization of alpha-synuclein. AB - Alpha-synuclein contributes to the pathogenesis of Parkinson's disease (PD), but its precise role in the disorder and its normal function remain poorly understood. Consistent with a presumed role in neurotransmitter release and its prominent deposition in the dystrophic neurites of PD, alpha-synuclein localizes almost exclusively to the nerve terminal. In brain extracts, however, alpha synuclein behaves as a soluble, monomeric protein. Using a binding assay to characterize the association of alpha-synuclein with cell membranes, we find that alpha-synuclein binds saturably and with high affinity to characteristic intracellular structures that double label for components of lipid rafts. Biochemical analysis demonstrates the interaction of alpha-synuclein with detergent-resistant membranes and reveals a shift in electrophoretic mobility of the raft-associated protein. In addition, the A30P mutation associated with PD disrupts the interaction of alpha-synuclein with lipid rafts. Furthermore, we find that both the A30P mutation and raft disruption redistribute alpha-synuclein away from synapses, indicating an important role for raft association in the normal function of alpha-synuclein and its role in the pathogenesis of PD. PMID- 15282275 TI - Opposing extracellular signal-regulated kinase and Akt pathways control Schwann cell myelination. AB - Schwann cells are the myelinating glia of the peripheral nervous system, and their development is regulated by various growth factors, such as neuregulin, platelet-derived growth factor (PDGF), and insulin-like growth factor-I (IGF-I). However, the mechanism of intracellular signaling pathways following these ligand stimuli in Schwann cell differentiation remains elusive. Here, we demonstrate that in cultured Schwann cells, neuregulin and PDGF suppressed the expression of myelin-associated protein markers, whereas IGF-I promoted it. Although these ligands activated common downstream signaling pathways [i.e., extracellular signal-regulated kinase (Erk) and phosphatidylinositol-3-kinase (PI3K)-Akt pathways], the profiles of activation varied among ligands. To elucidate the function of these pathways and the mechanisms underlying Schwann cell differentiation, we used adenoviral vectors to selectively activate or inactivate these pathways. We found that the selective activation of Erk pathways suppressed Schwann cell differentiation, whereas that of PI3K pathways promoted it. Furthermore, lithium chloride, a modulator of glycogen synthase kinase-3beta (GSK 3beta) promoted Schwann cell differentiation, suggesting the involvement of GSK 3beta as a downstream molecule of PI3K-Akt pathways. Selective activation of PI3K pathways in Schwann cells by gene transfer also demonstrated increased myelination in in vitro Schwann cell-DRG neuron cocultures and in vivo allogenic nerve graft experiments. We conclude that signals mediated by PI3K-Akt are crucial for initiation of myelination and that the effects of growth factors are primarily dependent on the balance between Erk and PI3K-Akt activation. Our results also propose the possibility of augmenting Schwann cell functions by modulating intracellular signals in light of future cell therapies. PMID- 15282276 TI - Common genetic effects on variation in impulsivity and activity in mice. AB - Impulsivity is a complex psychological construct that impacts on behavioral predispositions in the normal range and has been shown to have a genetic element through the examination of hereditary patterns of abnormal conditions such as attention deficit/hyperactivity disorder and obsessive compulsive disorder. In this study, we took advantage of the isogenic nature of inbred strains of mice to determine the contribution of genes to impulsive behaviors by examining the performance of four separate mouse strains in a novel murine delayed reinforcement paradigm, during which the animals had to choose between rewards that were relatively small but available immediately and larger but progressively delayed rewards. To control for maternal effects, all the mice were cross fostered to a common strain immediately after birth. Under these conditions, we found significant differences between the strains on behaviors indexing impulsive choice and on independent measures of locomotor activity, which subsequent heritability analysis showed could be related, in part, to genetic effects. Moreover, the two aspects of behavior were found to co-vary, with the more active animals also displaying more impulsive behavior. This was not attributable to mundane confounds related to individual task requirements but instead indicated the existence of common genetic factors influencing variation in both impulsivity and locomotor activity. The data are discussed in terms of the coexistence of impulsivity and hyperactivity, interactions between environmental and genetic effects, and possible candidate genes. PMID- 15282277 TI - Mechanosensory activation of a motor circuit by coactivation of two projection neurons. AB - Individual neuronal circuits can generate multiple activity patterns because of the influence of different projection neurons. However, in most systems it has been difficult to identify and assess the relative contribution of all upstream neurons responsible for the activation of any single activity pattern by a behaviorally relevant stimulus. To elucidate this issue, we used the stomatogastric nervous system (STNS) of the crab. The STNS includes the gastric mill (chewing) motor circuit in the stomatogastric ganglion (STG) and no more than 20 projection neurons that innervate the STG. We previously identified at least some (four) of the projection neurons that are activated directly by the ventral cardiac neuron (VCN) system, a population of mechanosensory neurons that activates the gastric mill circuit. Here we show that two of these projection neurons, the previously identified modulatory commissural neuron 1 (MCN1) and commissural projection neuron 2 (CPN2), are necessary and likely sufficient for the initiation/maintenance of the VCN-elicited gastric mill rhythm. Selective inactivation of either MCN1 or CPN2 still enabled a VCN-elicited gastric mill rhythm. However, because MCN1 and CPN2 have different actions on gastric mill neurons, these manipulations resulted in rhythms distinct from each other and from that occurring in the intact system. After removal of both MCN1 and CPN2, VCN stimulation failed to activate the gastric mill rhythm. Selective conjoint stimulation of MCN1 and CPN2, approximating their VCN-elicited activity patterns and firing frequencies, elicited a VCN-like gastric mill rhythm. Thus the VCN mechanosensory system elicits the gastric mill rhythm via its activation of a subset of the relevant projection neurons. PMID- 15282278 TI - GABA, not glutamate, controls the activity of substantia nigra reticulata neurons in awake, unrestrained rats. AB - Substantia nigra pars reticulata (SNr) receives both GABAergic and glutamatergic (GLU) inputs that are believed to act together to regulate neuronal activity in this structure. To examine the role of these inputs, single-unit recording was coupled with iontophoresis of GLU and GABA in rats under two conditions: awake, unrestrained and under chloral hydrate anesthesia. Although GABA potently inhibited SNr cells in both conditions, freely moving rats showed lower sensitivity than anesthetized animals. Likewise, GLU effectively induced excitations in most SNr neurons in anesthetized animals but was much less effective in awake, unrestrained animals in terms of both the number of sensitive cells and the magnitude of GLU-induced excitation. These findings, along with consistent excitations induced by bicuculline in awake, unrestrained rats, suggest that modulation of GABA inhibitory input, not the opposing actions of GLU and GABA, is the primary factor that regulates the activity state of SNr neurons. PMID- 15282279 TI - Dominance hierarchy influences adult neurogenesis in the dentate gyrus. AB - Many mammalian species form dominance hierarchies, but it remains unknown whether differences in social status correspond to structural differences in the brain. Stressful experiences may arise naturally during the establishment of dominance, and stress has been linked to adult neurogenesis in the hippocampus. To determine whether position in a dominance hierarchy leads to changes in adult neurogenesis in the hippocampus, we examined the brains of rats housed in a visible burrow system (VBS), a seminaturalistic environment with opportunities for social interaction. Dominance hierarchies emerged among the males in all colonies within 3 d of living in the VBS. Although cell proliferation in the dentate gyrus did not differ between the groups, more new neurons were observed in the dentate gyrus of the dominant males compared with both subordinates and controls. Dominant and subordinate animals showed similar basal, stress, and recovery from stress levels of corticosterone, as well as similar thymus, adrenal gland, and body weights, suggesting that variables other than stress are responsible for the observed changes in adult neurogenesis. The differences in brain structure persisted among the animals that had no access to the burrow system after the dominance hierarchy stabilized, suggesting that social status rather than living in a complex environment accounts for the effect of dominance on adult neurogenesis. PMID- 15282280 TI - Exclusive postsynaptic action of hypocretin-orexin on sublayer 6b cortical neurons. AB - The hypocretin-orexin (hcrt-orx) neurons are thought to maintain wakefulness because their loss results in narcolepsy. This role may be fulfilled by the excitatory action that the hcrt-orx peptide exerts on multiple brainstem and forebrain systems that, in turn, promote cortical activation. Here, we examined whether hcrt-orx may also exert a postsynaptic excitatory action at the level of the cortex, where hcrt-orx fibers project. However, we found that neurons in layers 2-5 in the primary somatosensory cortex (SSp) were unresponsive to hcrt orx. We then found that although all neurons tested in sublayer 6a were also unresponsive to hcrt-oxr, all those tested in sublayer 6b were highly sensitive to the peptide. The sublayer selectivity of hcrt-oxr was not restricted to the somatosensory cortex, because it was also found to be present in the primary visual cortex, the motor cortex, and the cingulate cortex. In the SSp, in which the hcrt-oxr effect was investigated further, it was demonstrated to be postsynaptic, to result from an interaction with Hcrtr2-OX2 receptors and to depend on the closure of a potassium conductance. Similar to the selectivity of action in the thalamus, where hcrt-oxr excites the nonspecific thalamocortical projection neurons and not the specific sensory relay neurons, here in the cortex, it excites a specific subset of cortical neurons which, through corticocortical projections, may also be involved in promoting widespread cortical activation. PMID- 15282281 TI - Fibroblast growth factor homologous factor 2B: association with Nav1.6 and selective colocalization at nodes of Ranvier of dorsal root axons. AB - Voltage-gated sodium channels interact with cytosolic proteins that regulate channel trafficking and/or modulate the biophysical properties of the channels. Na(v)1.6 is heavily expressed at the nodes of Ranvier along adult CNS and PNS axons and along unmyelinated fibers in the PNS. In an initial yeast two-hybrid screen using the C terminus of Na(v)1.6 as a bait, we identified FHF2B, a member of the FGF homologous factor (FHF) subfamily, as an interacting partner of Na(v)1.6. Members of the FHF subfamily share approximately 70% sequence identity, and individual members demonstrate a cell- and tissue-specific expression pattern. FHF2 is abundantly expressed in the hippocampus and DRG neurons and colocalizes with Na(v)1.6 at mature nodes of Ranvier in myelinated sensory fibers in the dorsal root of the sciatic nerve. However, retinal ganglion cells and spinal ventral horn motor neurons show very low levels of FHF2 expression, and their axons exhibit no nodal FHF2 staining within the optic nerve and ventral root, respectively. Thus, FHF2 is selectively localized at nodes of dorsal root sensory but not ventral root motor axons. The coexpression of FHF2B and Na(v)1.6 in the DRG-derived cell line ND7/23 significantly increases the peak current amplitude and causes a 4 mV depolarizing shift of voltage-dependent inactivation of the channel. The preferential expression of FHF2B in sensory neurons may provide a basis for physiological differences in sodium currents that have been reported at the nodes of Ranvier in sensory versus motor axons. PMID- 15282282 TI - Neuronal nicotinic synapse assembly requires the adenomatous polyposis coli tumor suppressor protein. AB - Normal cognitive and autonomic functions require nicotinic synaptic signaling. Despite the physiological importance of these synapses, little is known about molecular mechanisms that direct their assembly during development. We show here that the tumor-suppressor protein adenomatous polyposis coli (APC) functions in localizing alpha3-nicotinic acetylcholine receptors (nAChRs) to neuronal postsynaptic sites. Our quantitative confocal microscopy studies indicate that APC is selectively enriched at cholinergic synapses; APC surface clusters are juxtaposed to synaptic vesicle clusters and colocalize with alpha3-nAChRs but not with the neighboring synaptic glycine receptors or perisynaptic alpha7-nAChRs on chick ciliary ganglion (CG) neurons. We identify PSD (postsynaptic density)-93, beta-catenin, and microtubule end binding protein EB1 as APC binding partners. PSD-93 and beta-catenin are also enriched at alpha3-nAChR postsynaptic sites. EB1 shows close proximity to and partial overlap with alpha3-nAChR and APC surface clusters. We tested the role of APC in neuronal nicotinic synapse assembly by using retroviral-mediated in vivo overexpression of an APC dominant-negative (APC dn) peptide to block the interaction of endogenous APC with both EB1 and PSD-93 during synapse formation in CG neurons. The overexpressed APC-dn led to dramatic decreases in alpha3-nAChR surface levels and clusters. Effects were specific to alpha3-nAChR postsynaptic sites; synaptic glycine receptor and perisynaptic alpha7-nAChR clusters were not altered. In addition, APC-dn also reduced surface membrane-associated clusters of PSD-93 and EB1. The results show that APC plays a key role in organizing excitatory cholinergic postsynaptic specializations in CG neurons. We identify APC as the first nonreceptor protein to function in localizing nAChRs to neuronal synapses in vivo. PMID- 15282283 TI - Requirement of alpha5-GABAA receptors for the development of tolerance to the sedative action of diazepam in mice. AB - Despite its pharmacological relevance, the mechanism of the development of tolerance to the action of benzodiazepines is essentially unknown. The acute sedative action of diazepam is mediated via alpha1-GABA(A) receptors. Therefore, we tested whether chronic activation of these receptors by diazepam is sufficient to induce tolerance to its sedative action. Knock-in mice, in which thealpha1 ,alpha2-,alpha3-, oralpha(5)-GABA(A) receptors had been rendered insensitive to diazepam by histidine-arginine point mutation, were chronically treated with diazepam (8 d; 15 mg x kg(-1) x d(-1)) and tested for motor activity. Wild-type, alpha2(H101R), and alpha3(H126R) mice showed a robust diminution of the motor depressant drug action. In contrast, alpha5(H105R) mice failed to display any sedative tolerance. alpha1(H101R) mice showed no alteration of motor activity with chronic diazepam treatment. Autoradiography with [3H]flumazenil revealed no change in benzodiazepine binding sites. However, a decrease in alpha5-subunit radioligand binding was detected selectively in the dentate gyrus with specific ligands. This alteration was observed only in diazepam-tolerant animals, indicating that the manifestation of tolerance to the sedative action of diazepam is associated with a downregulation of alpha5-GABA(A) receptors in the dentate gyrus. Thus, the chronic activation of alpha(5)-GABA(A) receptors is crucial for the normal development of sedative tolerance to diazepam, which manifests itself in conjunction with alpha1-GABA(A) receptors. PMID- 15282285 TI - Beta-amyloid peptide at sublethal concentrations downregulates brain-derived neurotrophic factor functions in cultured cortical neurons. AB - The accumulation of beta-amyloid (Abeta) is one of the etiological factors in Alzheimer's disease (AD). It has been assumed that the underlying mechanism involves a critical role of Abeta-induced neurodegeneration. However, low levels of Abeta, such as will accumulate during the course of the disease, may interfere with neuronal function via mechanisms other than those involving neurodegeneration. We have been testing, therefore, the hypothesis that Abeta at levels insufficient to cause degeneration (sublethal) may interfere with critical signal transduction processes. In cultured cortical neurons Abeta at sublethal concentrations interferes with the brain-derived neurotrophic factor (BDNF) induced activation of the Ras-mitogen-activated protein kinase/extracellular signal-regulated protein kinase (ERK) and phosphatidylinositol 3-kinase (PI3 K)/Akt pathways. The effect of sublethal Abeta(1-42) on BDNF signaling results in the suppression of the activation of critical transcription factor cAMP response element-binding protein and Elk-1 and cAMP response element-mediated and serum response element-mediated transcription. The site of interference with the Ras/ERK and PI3-K/Akt signaling is downstream of the TrkB receptor and involves docking proteins insulin receptor substrate-1 and Shc, which convey receptor activation to the downstream effectors. The functional consequences of Abeta interference with signaling are robust, causing increased vulnerability of neurons, abrogating BDNF protection against DNA damage- and trophic deprivation induced apoptosis. These new findings suggest that Abeta engenders a dysfunctional encoding state in neurons and may initiate and/or contribute to cognitive deficit at an early stage of AD before or along with neuronal degeneration. PMID- 15282284 TI - Glycogen synthase kinase-3beta haploinsufficiency mimics the behavioral and molecular effects of lithium. AB - Lithium is widely used to treat bipolar disorder, but its mechanism of action in this disorder is unknown. Several molecular targets of lithium have been identified, but these putative targets have not been shown to be responsible for the behavioral effects of lithium in vivo. A robust model for the effects of chronic lithium on behavior in mice would greatly facilitate the characterization of lithium action. We describe behaviors in mice that are robustly affected by chronic lithium. Remarkably, these lithium-sensitive behaviors are also observed in mice lacking one copy of the gene encoding glycogen synthase kinase-3beta (Gsk 3beta), a well established direct target of lithium. In addition, chronic lithium induces molecular changes consistent with inhibition of GSK-3 within regions of the brain that are paralleled in Gsk-3beta+/- heterozygous mice. We also show that lithium therapy activates Wnt signaling in vivo, as measured by increased Wnt-dependent gene expression in the amygdala, hippocampus, and hypothalamus. These observations support a central role for GSK-3beta in mediating behavioral responses to lithium. PMID- 15282286 TI - A neural representation of pitch salience in nonprimary human auditory cortex revealed with functional magnetic resonance imaging. AB - Pitch, one of the primary auditory percepts, is related to the temporal regularity or periodicity of a sound. Previous functional brain imaging work in humans has shown that the level of population neural activity in centers throughout the auditory system is related to the temporal regularity of a sound, suggesting a possible relationship to pitch. In the current study, functional magnetic resonance imaging was used to measure activation in response to harmonic tone complexes whose temporal regularity was identical, but whose pitch salience (or perceptual pitch strength) differed, across conditions. Cochlear nucleus, inferior colliculus, and primary auditory cortex did not show significant differences in activation level between conditions. Instead, a correlate of pitch salience was found in the neural activity levels of a small, spatially localized region of nonprimary auditory cortex, overlapping the anterolateral end of Heschl's gyrus. The present data contribute to converging evidence that anterior areas of nonprimary auditory cortex play an important role in processing pitch. PMID- 15282287 TI - On the activity of the corticostriatal networks during spike-and-wave discharges in a genetic model of absence epilepsy. AB - Absence seizures are characterized by impairment of consciousness associated with widespread bilaterally synchronous spike-and-wave discharges (SWDs) in the electroencephalogram (EEG), which reflect highly synchronized oscillations in thalamocortical networks. Although recent pharmacological studies suggest that the basal ganglia could provide a remote control system for absence seizures, the mechanisms of propagation of epileptic discharges in these subcortical nuclei remain unknown. In the present study, we provide the first description of the electrical events in the corticostriatal pathway during spontaneous SWDs in the genetic absence epilepsy rats from Strasbourg (GAERS), a genetic model of absence epilepsy. In corticostriatal neurons, the SWDs were associated with suprathreshold rhythmic depolarizations in-phase with local EEG spikes. Consistent with this synchronized firing in their excitatory cortical afferents, striatal output neurons (SONs) exhibited, during SWDs, large-amplitude rhythmic synaptic depolarizations. However, SONs did not discharge during SWDs. Instead, the rhythmic synaptic excitation of SONs was shunted by a Cl(-)-dependent increase in membrane conductance that was temporally correlated with bursts of action potentials in striatal GABAergic interneurons. The reduced SON excitability accompanying absence seizures may participate in the control of SWDs by affecting the flow of cortical information within the basal ganglia circuits. PMID- 15282288 TI - Promotion of axon regeneration by myelin-associated glycoprotein and Nogo through divergent signals downstream of Gi/G. AB - Several myelin-derived proteins have been identified as components of the CNS myelin that prevents axonal regeneration in the adult vertebrate CNS. Activation of RhoA has been shown to be an essential part of the signaling mechanism of these proteins. Here we report an additional signal, which determines whether these proteins promote or inhibit axon outgrowth. Myelin-associated glycoprotein (MAG) and Nogo trigger the intracellular elevation of Ca2+ as well as the activation of PKC, presumably mediated by G(i)/G. Neurite outgrowth inhibition and growth cone collapse by MAG or Nogo can be converted to neurite extension and growth cone spreading by inhibiting conventional PKC, but not by inhibiting inositol 1,4,5-triphosphate (IP3). Conversely, neurite growth of immature neurons promoted by MAG is abolished by inhibiting IP3. Activation of RhoA is independent of PKC. Thus, a balance between PKC and IP3 is important for bidirectional regulation of axon regeneration by the myelin-derived proteins. PMID- 15282289 TI - Regulation of an Aplysia bag-cell neuron cation channel by closely associated protein kinase A and a protein phosphatase. AB - Ion channel regulation by closely associated kinases or phosphatases has emerged as a key mechanism for orchestrating neuromodulation. An exemplary case is the nonselective cation channel that drives the afterdischarge in Aplysia bag cell neurons. Initial studies showed that this channel is modulated by both a closely associated PKC and a serine/threonine protein phosphatase (PP). In excised, inside-out patches, the addition of ATP (a phosphate source) increases open probability (P(O)) through PKC, and this is reversed by the PP. Previous work also reported that, in certain cases, ATP can decrease cation channel P(O). The present study characterizes and provides a mechanism for this decreased P(O) ATP response. The kinetic change for channels inhibited by ATP was identical to the previously reported effect of exogenously applied protein kinase A (PKA) (i.e., a lengthening of the third closed-state time constant). The decreased P(O) ATP response was blocked by the PKA inhibitor peptide PKA(6-22), and its reversal was prevented by the PP inhibitor microcystin-LR. Furthermore, PKA(6-22) did not alter the increased P(O) ATP response. This suggests that both PKA and a PP are closely associated with these cation channels, but PKA and PKC are not simultaneously targeted. After an afterdischarge, the bag cell neurons are refractory and fail to respond to subsequent stimulation. The association of PKA with the cation channel may contribute to this decrease in excitability. Altering the constituents of a regulatory complex, such as exchanging PKA for PKC, may represent a general mechanism to precisely control ion channel function and excitability. PMID- 15282290 TI - Brain-derived neurotrophic factor mRNA and protein are targeted to discrete dendritic laminas by events that trigger epileptogenesis. AB - Dendritic targeting of mRNA and local protein synthesis are mechanisms that enable neurons to deliver proteins to specific postsynaptic sites. Here, we demonstrate that epileptogenic stimuli induce a dramatic accumulation of BDNF mRNA and protein in the dendrites of hippocampal neurons in vivo. BDNF mRNA and protein accumulate in dendrites in all hippocampal subfields after pilocarpine seizures and in selected subfields after other epileptogenic stimuli (kainate and kindling). BDNF accumulates selectively in discrete dendritic laminas, suggesting targeting to synapses that are active during seizures. Dendritic targeting of BDNF mRNA occurs during the time when the cellular changes that underlie epilepsy are occurring and is not seen after intense stimuli that are non-epileptogenic, including electroconvulsive seizures and high-frequency stimulation. MK801, an NMDA receptor antagonist that can prevent epileptogenesis but not acute seizures, prevents the dendritic accumulation of BDNF mRNA, indicating that dendritic targeting is mediated via NMDA receptor activation. Together, these results suggest that dendritic accumulation of BDNF mRNA and protein plays a critical role in the cellular changes leading to epilepsy. PMID- 15282291 TI - Role of DNA replication proteins in double-strand break-induced recombination in Saccharomyces cerevisiae. AB - Mitotic double-strand break (DSB)-induced gene conversion involves new DNA synthesis. We have analyzed the requirement of several essential replication components, the Mcm proteins, Cdc45p, and DNA ligase I, in the DNA synthesis of Saccharomyces cerevisiae MAT switching. In an mcm7-td (temperature-inducible degron) mutant, MAT switching occurred normally when Mcm7p was degraded below the level of detection, suggesting the lack of the Mcm2-7 proteins during gene conversion. A cdc45-td mutant was also able to complete recombination. Surprisingly, even after eliminating both of the identified DNA ligases in yeast, a cdc9-1 dnl4 Delta strain was able to complete DSB repair. Previous studies of asynchronous cultures carrying temperature-sensitive alleles of PCNA, DNA polymerase alpha (Pol alpha), or primase showed that these mutations inhibited MAT switching (A. M. Holmes and J. E. Haber, Cell 96:415-424, 1999). We have reevaluated the roles of these proteins in G(2)-arrested cells. Whereas PCNA was still essential for MAT switching, neither Pol alpha nor primase was required. These results suggest that arresting cells in S phase using ts alleles of Pol alpha-primase, prior to inducing the DSB, sequesters some other component that is required for repair. We conclude that DNA synthesis during gene conversion is different from S-phase replication, involving only leading-strand polymerization. PMID- 15282292 TI - Efficient and error-free replication past a minor-groove N2-guanine adduct by the sequential action of yeast Rev1 and DNA polymerase zeta. AB - Rev1, a member of the Y family of DNA polymerases, functions in lesion bypass together with DNA polymerase zeta (Pol zeta). Rev1 is a highly specialized enzyme in that it incorporates only a C opposite template G. While Rev1 plays an indispensable structural role in Pol zeta-dependent lesion bypass, the role of its DNA synthetic activity in lesion bypass has remained unclear. Since interactions of DNA polymerases with the DNA minor groove contribute to the nearly equivalent efficiencies and fidelities of nucleotide incorporation opposite each of the four template bases, here we examine the possibility that unlike other DNA polymerases, Rev1 does not come into close contact with the minor groove of the incipient base pair, and that enables it to incorporate a C opposite the N(2)-adducted guanines in DNA. To test this idea, we examined whether Rev1 could incorporate a C opposite the gamma-hydroxy-1,N(2)-propano 2'deoxyguanosine DNA minor-groove adduct, which is formed from the reaction of acrolein with the N(2) of guanine. Acrolein, an alpha,beta-unsaturated aldehyde, is generated in vivo as the end product of lipid peroxidation and from other oxidation reactions. We show here that Rev1 efficiently incorporates a C opposite this adduct from which Pol zeta subsequently extends, thereby completing the lesion bypass reaction. Based upon these observations, we suggest that an important role of the Rev1 DNA synthetic activity in lesion bypass is to incorporate a C opposite the various N(2)-guanine DNA minor-groove adducts that form in DNA. PMID- 15282293 TI - Kinetochores prevent repair of UV damage in Saccharomyces cerevisiae centromeres. AB - Centromeres form specialized chromatin structures termed kinetochores which are required for accurate segregation of chromosomes. DNA lesions might disrupt protein-DNA interactions, thereby compromising segregation and genome stability. We show that yeast centromeres are heavily resistant to removal of UV-induced DNA lesions by two different repair systems, photolyase and nucleotide excision repair. Repair resistance persists in G(1)- and G(2)/M-arrested cells. Efficient repair was obtained only by disruption of the kinetochore structure in a ndc10-1 mutant, but not in cse4-1 and cbf1 Delta mutants. Moreover, UV photofootprinting and DNA repair footprinting showed that centromere proteins cover about 120 bp of the centromere elements CDEII and CDEIII, including 20 bp of flanking CDEIII. Thus, DNA lesions do not appear to disrupt protein-DNA interactions in the centromere. Maintaining a stable kinetochore structure seems to be more important for the cell than immediate removal of DNA lesions. It is conceivable that centromeres are repaired by postreplication repair pathways. PMID- 15282294 TI - Biochemical and phenotypic abnormalities in kynurenine aminotransferase II deficient mice. AB - Kynurenic acid (KYNA) can act as an endogenous modulator of excitatory neurotransmission and has been implicated in the pathogenesis of several neurological and psychiatric diseases. To evaluate its role in the brain, we disrupted the murine gene for kynurenine aminotransferase II (KAT II), the principal enzyme responsible for the synthesis of KYNA in the rat brain. mKat-2( /-) mice showed no detectable KAT II mRNA or protein. Total brain KAT activity and KYNA levels were reduced during the first month but returned to normal levels thereafter. In contrast, liver KAT activity and KYNA levels in mKat-2(-/-) mice were decreased by >90% throughout life, though no hepatic abnormalities were observed histologically. KYNA-associated metabolites kynurenine, 3 hydroxykynurenine, and quinolinic acid were unchanged in the brain and liver of knockout mice. mKat-2(-/-) mice began to manifest hyperactivity and abnormal motor coordination at 2 weeks of age but were indistinguishable from wild type after 1 month of age. Golgi staining of cortical and striatal neurons revealed enlarged dendritic spines and a significant increase in spine density in 3-week old mKat-2(-/-) mice but not in 2-month-old animals. Our results show that gene targeting of mKat-2 in mice leads to early and transitory decreases in brain KAT activity and KYNA levels with commensurate behavioral and neuropathological changes and suggest that compensatory changes or ontogenic expression of another isoform may account for the normalization of KYNA levels in the adult mKat-2(-/-) brain. PMID- 15282295 TI - Budding yeast silencing complexes and regulation of Sir2 activity by protein protein interactions. AB - Gene silencing in the budding yeast Saccharomyces cerevisiae requires the enzymatic activity of the Sir2 protein, a highly conserved NAD-dependent deacetylase. In order to study the activity of native Sir2, we purified and characterized two budding yeast Sir2 complexes: the Sir2/Sir4 complex, which mediates silencing at mating-type loci and at telomeres, and the RENT complex, which mediates silencing at the ribosomal DNA repeats. Analyses of the protein compositions of these complexes confirmed previously described interactions. We show that the assembly of Sir2 into native silencing complexes does not alter its selectivity for acetylated substrates, nor does it allow the deacetylation of nucleosomal histones. The inability of Sir2 complexes to deacetylate nucleosomes suggests that additional factors influence Sir2 activity in vivo. In contrast, Sir2 complexes show significant enhancement in their affinities for acetylated substrates and their sensitivities to the physiological inhibitor nicotinamide relative to recombinant Sir2. Reconstitution experiments showed that, for the Sir2/Sir4 complex, these differences stem from the physical interaction of Sir2 with Sir4. Finally, we provide evidence that the different nicotinamide sensitivities of Sir2/Sir4 and RENT in vitro could contribute to locus-specific differences in how Sir2 activity is regulated in vivo. PMID- 15282296 TI - BRCA1 can modulate RNA polymerase II carboxy-terminal domain phosphorylation levels. AB - A high incidence of breast and ovarian cancers has been linked to mutations in the BRCA1 gene. BRCA1 has been shown to be involved in both positive and negative regulation of gene activity as well as in numerous other processes such as DNA repair and cell cycle regulation. Since modulation of the RNA polymerase II carboxy-terminal domain (CTD) phosphorylation levels could constitute an interface to all these functions, we wanted to directly test the possibility that BRCA1 might regulate the phosphorylation state of the CTD. We have shown that the BRCA1 C-terminal region can negatively modulate phosphorylation levels of the RNA polymerase II CTD by the Cdk-activating kinase (CAK) in vitro. Interestingly, the BRCA1 C-terminal region can directly interact with CAK and inhibit CAK activity by competing with ATP. Finally, we demonstrated that full-length BRCA1 can inhibit CTD phosphorylation when introduced in the BRCA1(-/-) HCC1937 cell line. Our results suggest that BRCA1 could play its ascribed roles, at least in part, by modulating CTD kinase components. PMID- 15282297 TI - Regulation of the maintenance of peripheral T-cell anergy by TAB1-mediated p38 alpha activation. AB - In anergic T cells, T-cell receptor (TCR)-mediated responses are functionally inactivated by negative regulatory signals whose mechanisms are poorly understood. Here, we show that CD4(+) T cells anergized in vivo by superantigen Mls-1(a) express a scaffolding protein, transforming growth factor beta-activated protein kinase 1-binding protein 1 (TAB1), that negatively regulates TCR signaling through the activation of mitogen-activated protein kinase p38 alpha. TAB1 was not expressed in naive and activated CD4(+) T cells. Inhibition of p38 activity in anergic T cells by a chemical inhibitor resulted in the recovery of interleukin 2 (IL-2) and the inhibition of IL-10 secretion. T-cell hybridoma 2B4 cells transduced with TAB1-containing retrovirus (TAB1-2B4 cells) showed activated p38 alpha, inhibited extracellular signal-regulated kinase (ERK) activity, culminating in reduced IL-2 levels and increased IL-10 production. The use of a p38 inhibitor or cotransfection of a dominant-negative form of p38 in TAB1-2B4 cells resulted in the recovery of ERK activity and IL-2 production. These results imply that TAB1-mediated activation of p38 alpha in anergic T cells regulates the maintenance of T-cell unresponsiveness both by inhibiting IL-2 production and by promoting IL-10 production. PMID- 15282298 TI - The in vivo activity of Ime1, the key transcriptional activator of meiosis specific genes in Saccharomyces cerevisiae, is inhibited by the cyclic AMP/protein kinase A signal pathway through the glycogen synthase kinase 3-beta homolog Rim11. AB - Phosphorylation is the main mode by which signals are transmitted to key regulators of developmental pathways. The glycogen synthase kinase 3 family plays pivotal roles in the development and well-being of all eukaryotic organisms. Similarly, the budding yeast homolog Rim11 is essential for the exit of diploid cells from the cell cycle and for entry into the meiotic developmental pathway. In this report we show that in vivo, in cells grown in a medium promoting vegetative growth with acetate as the sole carbon source (SA medium), Rim11 phosphorylates Ime1, the master transcriptional activator required for entry into the meiotic cycle and for the transcription of early meiosis-specific genes. We demonstrate that in the presence of glucose, the kinase activity of Rim11 is inhibited. This inhibition could be due to phosphorylation on Ser-5, Ser-8, and/or Ser-12 because in the rim11S5AS8AS12A mutant, Ime1 is incorrectly phosphorylated in the presence of glucose and cells undergo sporulation. We further show that this nutrient signal is transmitted to Rim11 and consequently to Ime1 by the cyclic AMP/protein kinase A signal transduction pathway. Ime1 is phosphorylated in SA medium on at least two residues, Tyr-359 and Ser-302 and/or Ser-306. Ser-302 and Ser-306 are part of a consensus site for the mammalian homolog of Rim11, glycogen synthase kinase 3-beta. Phosphorylation on Tyr-359 but not Ser-302 or Ser-306 is essential for the transcription of early meiosis specific genes and sporulation. We show that Tyr-359 is phosphorylated by Rim11. PMID- 15282299 TI - Activation of RBL-2H3 mast cells is dependent on tyrosine phosphorylation of phospholipase D2 by Fyn and Fgr. AB - Both phospholipase D1 (PLD1) and PLD2 regulate degranulation when RBL-2H3 cells are stimulated via the immunoglobulin E receptor, Fc epsilon RI. However, the activation mechanism for PLD2 is unclear. As reported here, PLD2 but not PLD1 is phosphorylated through the Src kinases, Fyn and Fgr, and this phosphorylation appears to regulate PLD2 activation and degranulation. For example, only hemagglutinin-tagged PLD2 was tyrosine phosphorylated in antigen-stimulated cells that had been made to express HA-PLD1 and HA-PLD2. This phosphorylation was blocked by a Src kinase inhibitor or by small interfering RNAs directed against Fyn and Fgr and was enhanced by overexpression of Fyn and Fgr but not by other Src kinases. The phosphorylation and activity of PLD2 were further enhanced by the tyrosine phosphatase inhibitor, Na(3)VO(4). Mutation of PLD2 at tyrosines 11, 14, 165, or 470 partially impaired, and mutation of all tyrosines blocked, PLD2 phosphorylation and activation, although two of these mutations were detrimental to PLD2 function. PLD2 phosphorylation preceded degranulation, both events were equally sensitive to inhibition of Src kinase activity, and both were enhanced by coexpression of PLD2 and the Src kinases. The findings provide the first description of a mechanism for activation of PLD2 in a physiological setting and of a role for Fgr in Fc epsilon RI-mediated signaling. PMID- 15282300 TI - Hematopoietic cell fate and the initiation of leukemic properties in primitive primary human cells are influenced by Ras activity and farnesyltransferase inhibition. AB - The Ras pathway transduces divergent signals determining normal cell fate and is frequently activated in hematopoietic malignancies, but the manner in which activation contributes to human leukemia is poorly understood. We report that a high level of activated H-Ras signaling in transduced primary human hematopoietic progenitors reduced their proliferation and enhanced monocyte/macrophage differentiation. However, the exposure of these cells to a farnesyltransferase inhibitor and establishment of a moderate level of Ras activity showed increased proliferation, an elevated frequency of primitive blast-like cells, and progenitors with enhanced self-renewal capacity. These results suggest that the amplitude of Ras pathway signaling is a determinant of myeloid cell fate and that moderate Ras activation in primitive hematopoietic cells can be an early event in leukemogenesis. PMID- 15282301 TI - Upstream regulatory role for XIAP in receptor-mediated apoptosis. AB - X-linked inhibitor of apoptosis (XIAP) is an endogenous inhibitor of cell death that functions by suppressing caspases 3, 7, and 9. Here we describe the establishment of Jurkat-derived cell lines stably overexpressing either full length XIAP or a truncation mutant of XIAP that can only inhibit caspase 9. Characterization of these cell lines revealed that following CD95 activation full length XIAP supported both short- and long-term survival as well as proliferative capacity, in contrast to the truncation mutant but similar to Bcl-x(L). Full length XIAP was also able to inhibit CD95-mediated caspase 3 processing and activation, the mitochondrial release of cytochrome c and Smac/DIABLO, and the loss of mitochondrial membrane potential, whereas the XIAP truncation mutant failed to prevent any of these cell death events. Finally, suppression of XIAP levels by RNA interference sensitized Bcl-x(L)-overexpressing cells to death receptor-induced apoptosis. These data demonstrate for the first time that full length XIAP inhibits caspase activation required for mitochondrial amplification of death receptor signals and that, by acting upstream of mitochondrial activation, XIAP supports the long-term proliferative capacity of cells following CD95 stimulation. PMID- 15282302 TI - The T-cell receptor beta variable gene promoter is required for efficient V beta rearrangement but not allelic exclusion. AB - To investigate the role of promoters in regulating variable gene rearrangement and allelic exclusion, we constructed mutant mice in which a 1.2-kb region of the V beta 13 promoter was either deleted (P13(-/-)) or replaced with the simian virus 40 minimal promoter plus five copies of Gal4 DNA sequences (P13(R/R)). In P13(-/-) mice, cleavage, rearrangement, and transcription of V beta 13, but not the flanking V beta gene segments, were significantly inhibited. In P13(R/R) mice, inhibition of V beta 13 rearrangement was less severe and was not associated with any apparent reduction in V beta 13 cleavage. Expression of a T cell receptor (TCR) transgene blocked cleavages at the normal V beta 13 recombination signal sequence junction and V beta 13 coding joint formation of both wild-type and mutant V beta 13 alleles. However, a low level of aberrant V beta 13 cleavage was consistently detected, especially in TCR transgenic P13(R/R) mice. These findings suggest that the variable gene promoter is required for promoting local recombination accessibility of the associated V beta gene segment. Although the promoter is dispensable for allelic exclusion, it appears to suppress aberrant V beta cleavages during allelic exclusion. PMID- 15282303 TI - Expression of telomerase RNA template, but not telomerase reverse transcriptase, is limiting for telomere length maintenance in vivo. AB - Telomerase consists of two essential components, the telomerase RNA template (TR) and telomerase reverse transcriptase (TERT). The haplo-insufficiency of TR was recently shown to cause one form of human dyskeratosis congenita, an inherited disease marked by abnormal telomere shortening. Consistent with this finding, we recently reported that mice heterozygous for inactivation of mouse TR exhibit a similar haplo-insufficiency and are deficient in the ability to elongate telomeres in vivo. To further assess the genetic regulation of telomerase activity, we have compared the abilities of TR-deficient and TERT-deficient mice to maintain or elongate telomeres in interspecies crosses. Homozygous TERT knockout mice had no telomerase activity and failed to maintain telomere length. In contrast, TERT(+/-) heterozygotes had no detectable defect in telomere elongation compared to wild-type controls, whereas TR(+/-) heterozygotes were deficient in telomere elongation. Levels of TERT mRNA in heterozygous mice were one-third to one-half the levels expressed in wild-type mice, similar to the reductions in telomerase RNA observed in TR heterozygotes. These findings indicate that both TR and TERT are essential for telomere maintenance and elongation but that gene copy number and transcriptional regulation of TR, but not TERT, are limiting for telomerase activity under the in vivo conditions analyzed. PMID- 15282304 TI - A histone methyltransferase is required for maximal response to female sex hormones. AB - RIZ1 is an estrogen receptor (ER) coactivator but is also a histone lysine methyltransferase that methylates lysine 9 of histone H3, an activity known to repress transcription. We show here that target organs of mice deficient in RIZ1 exhibit decreased response to female sex hormones. RIZ1 interacted with SRC1 and p300, suggesting that the coactivator function of RIZ1 may be mediated by its interaction with other transcriptional coactivators. In the presence of estrogen, RIZ1 binding to estrogen target genes became less direct and followed the binding of ER to DNA and RIZ1 methyltransferase activity on H3-Lys 9 was inhibited, indicating derepression may play a role in estrogen induction of gene transcription. Reducing RIZ1 level correlated with decreased induction of pS2 gene by estrogen in MCF7 cells. The data suggest that a histone methyltransferase is required for optimal estrogen response in female reproductive tissues and that estrogen-bound ER may turn a transcriptional repressor into a coactivator. PMID- 15282305 TI - RPAP1, a novel human RNA polymerase II-associated protein affinity purified with recombinant wild-type and mutated polymerase subunits. AB - We have programmed human cells to express physiological levels of recombinant RNA polymerase II (RNAPII) subunits carrying tandem affinity purification (TAP) tags. Double-affinity chromatography allowed for the simple and efficient isolation of a complex containing all 12 RNAPII subunits, the general transcription factors TFIIB and TFIIF, the RNAPII phosphatase Fcp1, and a novel 153-kDa polypeptide of unknown function that we named RNAPII-associated protein 1 (RPAP1). The TAP tagged RNAPII complex is functionally active both in vitro and in vivo. A role for RPAP1 in RNAPII transcription was established by shutting off the synthesis of Ydr527wp, a Saccharomyces cerevisiae protein homologous to RPAP1, and demonstrating that changes in global gene expression were similar to those caused by the loss of the yeast RNAPII subunit Rpb11. We also used TAP-tagged Rpb2 with mutations in fork loop 1 and switch 3, two structural elements located strategically within the active center, to start addressing the roles of these elements in the interaction of the enzyme with the template DNA during the transcription reaction. PMID- 15282306 TI - Phosphorylation of Y845 on the epidermal growth factor receptor mediates binding to the mitochondrial protein cytochrome c oxidase subunit II. AB - When co-overexpressed, the epidermal growth factor receptor (EGFR) and c-Src cooperate to cause synergistic increases in EGF-induced DNA synthesis, soft agar colony growth, and tumor formation in nude mice. This synergy is dependent upon c Src-mediated phosphorylation of a unique tyrosine on the EGFR, namely, tyrosine 845 (Y845). Phenylalanine substitution of Y845 (Y845F) was found to inhibit EGF induced DNA synthesis without affecting the catalytic activity of the receptor or its ability to phosphorylate Shc or activate mitogen-activated protein kinase. These results suggest that synergism may occur through alternate signaling pathways mediated by phosphorylated Y845 (pY845). One such pathway involves the transcription factor Stat5b. Here we describe another pathway that involves cytochrome c oxidase subunit II (CoxII). CoxII was identified as a specific binding partner of a pY845-containing peptide in a phage display screen. EGF dependent binding of CoxII to the wild type but not to the mutant Y845F-EGFR was confirmed by coimmunoprecipitation experiments. This association also required the kinase activity of c-Src. Confocal microscopy, as well as biochemical fractionation, indicated that the EGFR translocates to the mitochondria after EGF stimulation, where it colocalizes with CoxII. Such translocation required the catalytic activity of the receptor but not phosphorylation of Y845. However, ectopic expression of the Y845F-EGFR prevented the EGF from protecting MDA-MB-231 breast cancer cells from adriamycin-induced apoptosis, whereas two mutants of Stat5b, a dominant-interfering mutant (DNstat5b) and a tyrosine mutation at 699 (Y699F-Stat5b) did not. Taken together, these data suggest that, through the ability of EGFR to translocate to the mitochondria, the binding of proteins such as CoxII to pY845 on the EGFR may positively regulate survival pathways that contribute to oncogenesis. PMID- 15282307 TI - Keratins modulate c-Flip/extracellular signal-regulated kinase 1 and 2 antiapoptotic signaling in simple epithelial cells. AB - Among the large family of intermediate filament proteins, the keratin 8 and 18 (K8/K18) pair constitutes a hallmark for all simple epithelial cells, such as hepatocytes and mammary cells. Functional studies with different cell models have suggested that K8/K18 are involved in simple epithelial cell resistance to several forms of stress that may lead to cell death. We have reported recently that K8/K18-deprived hepatocytes from K8-null mice are more sensitive to Fas mediated apoptosis. Here we show that upon Fas, tumor necrosis factor alpha receptor, or tumor necrosis factor alpha-related apoptosis-inducing ligand receptor stimulation, an inhibition of extracellular signal-regulated kinase 1 and 2 (ERK1/2) activation sensitizes wild-type but not K8-null mouse hepatocytes to apoptosis and that a much weaker ERK1/2 activation occurs in K8-null hepatocytes. In turn, this impaired ERK1/2 activation in K8-null hepatocytes is associated with a drastic reduction in c-Flip protein, an event that also holds in a K8-null mouse mammary cell line. c-Flip, along with Raf-1, is part of a K8/K18-immunoisolated complex from wild-type hepatocytes, and Fas stimulation leads to further c-Flip and Raf-1 recruitment in the complex. This points to a new regulatory role of simple epithelium keratins in the c-Flip/ERK1/2 antiapoptotic signaling pathway. PMID- 15282308 TI - Mrc1 is required for sister chromatid cohesion to aid in recombination repair of spontaneous damage. AB - The SRS2 gene of Saccharomyces cerevisiae encoding a 3'-->5' DNA helicase is part of the postreplication repair pathway and functions to ensure proper repair of DNA damage arising during DNA replication through pathways that do not involve homologous recombination. Through a synthetic gene array analysis, genes that are essential when Srs2 is absent have been identified. Among these are MRC1, TOF1, and CSM3, which mediate the intra-S checkpoint response. srs2 Delta mrc1 Delta synthetic lethality is due to inappropriate recombination, as the lethality can be suppressed by genetic elimination of homologous recombination. srs2 Delta mrc1 Delta synthetic lethality is dependent on the role of Mrc1 in DNA replication but independent of the role of Mrc1 in a DNA damage checkpoint response. mrc1 Delta, tof1 Delta and csm3 Delta mutants have sister chromatid cohesion defects, implicating sister chromatid cohesion established at the replication fork as an important factor in promoting repair of stalled replication forks through gap repair. PMID- 15282309 TI - Inhibition of nuclear import by the proapoptotic protein CC3. AB - We report here that the normal cellular protein CC3/TIP30, when in excess, inhibits nuclear import in vitro and in vivo. CC3 binds directly to the karyopherins of the importin beta family in a RanGTP-insensitive manner and associates with nucleoporins in vivo. CC3 inhibits the nuclear import of proteins possessing either the classical nuclear localization signal or the M9 signal recognized by transportin. CC3 also inhibits nuclear translocation of transportin itself. Cells modified to express higher levels of CC3 have a slower rate of nuclear import and, as described earlier, show an increased sensitivity to death signals. A mutant CC3 protein lacking proapoptotic activity has a lower affinity for transportin, is displaced from it by RanGTP, and fails to inhibit nuclear import in vitro and in vivo. Together, our results support a correlation between the ability of CC3 to form a RanGTP-resistant complex with importins, inhibit nuclear import, and induce apoptosis. Significantly, a dominant-negative form of importin beta1 shown previously to inhibit multiple transport pathways induces rapid cell death, strongly indicating that inhibition of nuclear transport serves as a potent apoptotic signal. PMID- 15282310 TI - Loss of the Sall3 gene leads to palate deficiency, abnormalities in cranial nerves, and perinatal lethality. AB - Members of the Spalt gene family encode putative transcription factors characterized by seven to nine C2H2 zinc finger motifs. Four genes have been identified in mice--Spalt1 to Spalt4 (Sall1 to Sall4). Spalt homologues are widely expressed in neural and mesodermal tissues during early embryogenesis. Sall3 is normally expressed in mice from embryonic day 7 (E7) in the neural ectoderm and primitive streak and subsequently in the brain, peripheral nerves, spinal cord, limb buds, palate, heart, and otic vesicles. We have generated a targeted disruption of Sall3 in mice. Homozygous mutant animals die on the first postnatal day and fail to feed. Examination of the oral structures of these animals revealed that abnormalities were present in the palate and epiglottis from E16.5. In E10.5 embryos, deficiencies in cranial nerves that normally innervate oral structures, particularly the glossopharyngeal nerve (IX), were observed. These studies indicate that Sall3 is required for the development of nerves that are derived from the hindbrain and for the formation of adjacent branchial arch derivatives. PMID- 15282311 TI - Defective extraembryonic angiogenesis in mice lacking LBP-1a, a member of the grainyhead family of transcription factors. AB - LBP-1a and CP2 are ubiquitously expressed members of the grainyhead transcription factor family, sharing significant sequence homology, a common DNA binding motif, and modulating a range of key regulatory and structural genes. We have reported previously that CP2-null mice are viable with no obvious abnormality. LBP-1a provides redundant function in this context. We show here that mice lacking LBP 1a expression develop intrauterine growth retardation at embryonic day 10.5, culminating in death 1 day later. No focal intraembryonic cause for this CP2 independent defect is evident. In contrast, a significant reduction in the thickness of the labyrinthine layer of the placenta is observed in LBP-1a(-/-) animals. However, expression of trophoblast differentiation markers is unperturbed in this context, and complementation studies utilizing tetraploid wild-type cells failed to rescue or ameliorate the LBP-1a(-/-) phenotype, excluding a primary trophoblast defect. An explanation for these observations is provided by the prominent angiogenic defect observed in the mutant placentas. LBP 1a(-/-) allantoic blood vessels fail to penetrate deeply and branch into the complex embryonic vasculature characteristic of the normal placenta. Interestingly, a similar defect in angiogenesis is observed in the yolk sac vasculature, primary endothelial cell-lined capillary tubes, although present, failed to connect into a characteristic intricate vascular network. Collectively, these results demonstrate that LBP-1a plays a critical role in the regulation of extraembryonic angiogenesis. PMID- 15282312 TI - Oxidative stress sensor Keap1 functions as an adaptor for Cul3-based E3 ligase to regulate proteasomal degradation of Nrf2. AB - Transcription factor Nrf2 is a major regulator of genes encoding phase 2 detoxifying enzymes and antioxidant stress proteins in response to electrophilic agents and oxidative stress. In the absence of such stimuli, Nrf2 is inactive owing to its cytoplasmic retention by Keap1 and rapid degradation through the proteasome system. We examined the contribution of Keap1 to the rapid turnover of Nrf2 (half-life of less than 20 min) and found that a direct association between Keap1 and Nrf2 is required for Nrf2 degradation. In a series of domain function analyses of Keap1, we found that both the BTB and intervening-region (IVR) domains are crucial for Nrf2 degradation, implying that these two domains act to recruit ubiquitin-proteasome factors. Indeed, Cullin 3 (Cul3), a subunit of the E3 ligase complex, was found to interact specifically with Keap1 in vivo. Keap1 associates with the N-terminal region of Cul3 through the IVR domain and promotes the ubiquitination of Nrf2 in cooperation with the Cul3-Roc1 complex. These results thus provide solid evidence that Keap1 functions as an adaptor of Cul3 based E3 ligase. To our knowledge, Nrf2 and Keap1 are the first reported mammalian substrate and adaptor, respectively, of the Cul3-based E3 ligase system. PMID- 15282313 TI - Rereplication by depletion of geminin is seen regardless of p53 status and activates a G2/M checkpoint. AB - Genomic DNA replication is tightly controlled to ensure that DNA replication occurs once per cell cycle; loss of this control leads to genomic instability. Geminin, a DNA replication inhibitor, plays an important role in regulation of DNA replication. To investigate the role of human geminin in the maintenance of genomic stability, we eliminated geminin by RNA interference in human cancer cells. Depletion of geminin led to overreplication and the formation of giant nuclei in cells that had wild-type or mutant p53. We found that overreplication caused by depletion of geminin activated both Chk1 and Chk2, which then phosphorylated Cdc25C on Ser216, resulting in its sequestration outside the nucleus, thus inhibiting cyclin B-Cdc2 activity. This activated G(2)/M checkpoint prevented cells with overreplicated DNA from entering mitosis. Addition of caffeine, UCN-01, or inhibitors of checkpoint pathways or silencing of Chk1 suppressed the accumulation of overreplicated cells and promoted apoptosis. From these results, we conclude that geminin is required for suppressing overreplication in human cells and that a G(2)/M checkpoint restricts the proliferation of cells with overreplicated DNA. PMID- 15282314 TI - Glut4 storage vesicles without Glut4: transcriptional regulation of insulin dependent vesicular traffic. AB - Two families of transcription factors that play a major role in the development of adipocytes are the CCAAT/enhancer-binding proteins (C/EBPs) and the peroxisome proliferator-activated receptors (PPARs), in particular PPAR gamma. Ectopic expression of either C/EBP alpha or PPAR gamma in NIH 3T3 fibroblasts results in the conversion of these cells to adipocyte-like cells replete with fat droplets. NIH 3T3 cells ectopically expressing C/EBP alpha (NIH-C/EBP alpha) differentiate into adipocytes and exhibit insulin-stimulated glucose uptake, whereas NIH 3T3 cells ectopically expressing PPAR gamma (NIH-PPAR gamma) differentiate but do not exhibit any insulin-stimulated glucose uptake, nor do they express any C/EBP alpha. The reason for the lack of insulin-responsive glucose uptake in the NIH PPAR gamma cells is their virtual lack of the insulin-responsive glucose transporter, Glut4. The NIH-PPAR gamma cells express functionally active components of the insulin receptor-signaling pathway (the insulin receptor, IRS 1, phosphatidylinositol 3-kinase, and Akt2) at levels comparable to those in responsive cell lines. They also express components of the insulin-sensitive vesicular transport machinery, namely, VAMP2, syntaxin-4, and IRAP, the last of these being the other marker of insulin-regulated vesicular traffic along with Glut4. Interestingly, the NIH-PPAR gamma cells show normal insulin-dependent translocation of IRAP and form an insulin-responsive vesicular compartment as assessed by cell surface biotinylation and sucrose velocity gradient analysis, respectively. Moreover, expression of a Glut4-myc construct in the NIH-PPAR gamma cells results in its insulin-dependent translocation to the plasma membrane as assessed by immunofluorescence and Western blot analysis. Based on these data, we conclude that major role of C/EBP alpha in the context of the NIH-PPAR gamma cells is to regulate Glut4 expression. The differentiated cells possess a large insulin-sensitive vesicular compartment with negligible Glut4, and Glut4 translocation can be reconstituted on expression of this transporter. PMID- 15282315 TI - Depletion of the 110-kilodalton isoform of poly(ADP-ribose) glycohydrolase increases sensitivity to genotoxic and endotoxic stress in mice. AB - Poly(ADP-ribosylation) is rapidly stimulated in cells following DNA damage. This posttranslational modification is regulated by the synthesizing enzyme poly(ADP ribose) polymerase 1 (PARP-1) and the degrading enzyme poly(ADP-ribose) glycohydrolase (PARG). Although the role of PARP-1 in response to DNA damage has been studied extensively, the function of PARG and the impact of poly(ADP-ribose) homeostasis in various cellular processes are largely unknown. Here we show that by gene targeting in embryonic stem cells and mice, we specifically deleted the 110-kDa PARG protein (PARG(110)) normally found in the nucleus and that depletion of PARG(110) severely compromised the automodification of PARP-1 in vivo. PARG(110)-deficient mice were viable and fertile, but these mice were hypersensitive to alkylating agents and ionizing radiation. In addition, these mice were susceptible to streptozotocin-induced diabetes and endotoxic shock. These data indicate that PARG(110) plays an important role in DNA damage responses and in pathological processes. PMID- 15282316 TI - Abnormal cardiac development in the absence of heart glycogen. AB - Glycogen serves as a repository of glucose in many mammalian tissues. Mice lacking this glucose reserve in muscle, heart, and several other tissues were generated by disruption of the GYS1 gene, which encodes an isoform of glycogen synthase. Crossing mice heterozygous for the GYS1 disruption resulted in a significant underrepresentation of GYS1-null mice in the offspring. Timed matings established that Mendelian inheritance was followed for up to 18.5 days postcoitum (dpc) and that approximately 90% of GYS1-null animals died soon after birth due to impaired cardiac function. Defects in cardiac development began between 11.5 and 14.5 dpc. At 18.5 dpc, the hearts were significantly smaller, with reduced ventricular chamber size and enlarged atria. Consistent with impaired cardiac function, edema, pooling of blood, and hemorrhagic liver were seen. Glycogen synthase and glycogen were undetectable in cardiac muscle and skeletal muscle from the surviving null mice, and the hearts showed normal morphology and function. Congenital heart disease is one of the most common birth defects in humans, at up to 1 in 50 live births. The results provide the first direct evidence that the ability to synthesize glycogen in cardiac muscle is critical for normal heart development and hence that its impairment could be a significant contributor to congenital heart defects. PMID- 15282317 TI - Regulation of the rapsyn promoter by kaiso and delta-catenin. AB - Rapsyn is a synapse-specific protein that is required for clustering acetylcholine receptors at the neuromuscular junction. Analysis of the rapsyn promoter revealed a consensus site for the transcription factor Kaiso within a region that is mutated in a subset of patients with congenital myasthenic syndrome. Kaiso is a POZ-zinc finger family transcription factor which recognizes the specific core consensus sequence CTGCNA (where N is any nucleotide). Previously, the only known binding partner for Kaiso was the cell adhesion cofactor, p120 catenin. Here we show that delta-catenin, a brain-specific member of the p120 catenin subfamily, forms a complex with Kaiso. Antibodies against Kaiso and delta-catenin recognize proteins in the nuclei of C2C12 myocytes and at the postsynaptic domain of the mouse neuromuscular junction. Endogenous Kaiso in C2C12 cells coprecipitates with the rapsyn promoter in vivo as shown by chromatin immunoprecipitation assay. Minimal promoter assays demonstrated that the rapsyn promoter can be activated by Kaiso and delta-catenin; this activation is apparently muscle specific. These results provide the first experimental evidence that rapsyn is a direct sequence-specific target of Kaiso and delta-catenin. We propose a new model of synapse-specific transcription that involves the interaction of Kaiso, delta-catenin, and myogenic transcription factors at the neuromuscular junction. PMID- 15282318 TI - Dp1 is largely dispensable for embryonic development. AB - E2F/DP complexes activate or repress the transcription of E2F target genes, depending on the association of a pRB family member, thereby regulating cell cycle progression. Whereas the E2F family consists of seven members, the DP family contains only two (Dp1 and Dp2), Dp1 being the more highly expressed member. In contrast to the inactivation of individual E2F family members, we have recently demonstrated that loss of Dp1 results in embryonic lethality by embryonic day 12.5 (E12.5) due to the failure of extraembryonic lineages to develop and replicate DNA properly. To bypass this placental requirement and search for roles of Dp1 in the embryo proper, we generated Dp1-deficient embryonic stem (ES) cells that carry the ROSA26-LacZ marker and injected them into wild-type blastocysts to construct Dp1-deficient chimeras. Surprisingly, we recovered mid- to late gestational embryos (E12.5 to E17.5), in which the Dp1 deficient ES cells contributed strongly to most chimeric tissues as judged by X Gal (5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside) staining and Western blotting. Importantly, the abundance of DP2 protein does not increase and the expression of an array of cell cycle genes is virtually unchanged in Dp1 deficient ES cells or chimeric E15.5 tissues with the absence of Dp1. Thus, Dp1 is largely dispensable for embryonic development, despite the absolute extraembryonic requirement for Dp1, which is highly reminiscent of the restricted roles for Rb and cyclins E1/E2 in vivo. PMID- 15282319 TI - Scrambled prion domains form prions and amyloid. AB - The [URE3] prion of Saccharomyces cerevisiae is a self-propagating amyloid form of Ure2p. The amino-terminal prion domain of Ure2p is necessary and sufficient for prion formation and has a high glutamine (Q) and asparagine (N) content. Such Q/N-rich domains are found in two other yeast prion proteins, Sup35p and Rnq1p, although none of the many other yeast Q/N-rich domain proteins have yet been found to be prions. To examine the role of amino acid sequence composition in prion formation, we used Ure2p as a model system and generated five Ure2p variants in which the order of the amino acids in the prion domain was randomly shuffled while keeping the amino acid composition and C-terminal domain unchanged. Surprisingly, all five formed prions in vivo, with a range of frequencies and stabilities, and the prion domains of all five readily formed amyloid fibers in vitro. Although it is unclear whether other amyloid-forming proteins would be equally resistant to scrambling, this result demonstrates that [URE3] formation is driven primarily by amino acid composition, largely independent of primary sequence. PMID- 15282320 TI - Vanin-1-/- mice exhibit a glutathione-mediated tissue resistance to oxidative stress. AB - Vanin-1 is an epithelial ectoenzyme with pantetheinase activity and generating the amino-thiol cysteamine through the metabolism of pantothenic acid (vitamin B(5)). Here we show that Vanin-1(-/-) mice, which lack cysteamine in tissues, exhibit resistance to oxidative injury induced by whole-body gamma-irradiation or paraquat. This protection is correlated with reduced apoptosis and inflammation and is reversed by treating mutant animals with cystamine. The better tolerance of the Vanin-1(-/-) mice is associated with an enhanced gamma-glutamylcysteine synthetase activity in liver, probably due to the absence of cysteamine and leading to elevated stores of glutathione (GSH), the most potent cellular antioxidant. Consequently, Vanin-1(-/-) mice maintain a more reducing environment in tissue after exposure to irradiation. In normal mice, we found a stress induced biphasic expression of Vanin-1 regulated via antioxidant response elements in its promoter region. This process should finely tune the redox environment and thus change an early inflammatory process into a late tissue repair process. We propose Vanin-1 as a key molecule to regulate the GSH dependent response to oxidative injury in tissue at the epithelial level. Therefore, Vanin/pantetheinase inhibitors could be useful for treatment of damage due to irradiation and pro-oxidant inducers. PMID- 15282321 TI - Edd, the murine hyperplastic disc gene, is essential for yolk sac vascularization and chorioallantoic fusion. AB - EDD is the mammalian ortholog of the Drosophila melanogaster hyperplastic disc gene (hyd), which is critical for cell proliferation and differentiation in flies through regulation of hedgehog and decapentaplegic signaling. Amplification and overexpression of EDD occurs frequently in several cancers, including those of the breast and ovary, and truncating mutations of EDD are also observed in gastric and colon cancer with microsatellite instability. EDD has E3 ubiquitin ligase activity, is involved in regulation of the DNA damage response, and may control hedgehog signaling, but a definitive biological role has yet to be established. To investigate the role of Edd in vivo, gene targeting was used to generate Edd knockout (Edd(Delta/Delta)) mice. While heterozygous mice had normal development and fertility, no viable Edd-deficient embryos were observed beyond E10.5, with delayed growth and development evident from E8.5 onward. Failed yolk sac and allantoic vascular development, along with defective chorioallantoic fusion, were the primary effects of Edd deficiency. These extraembryonic defects presumably compromised fetal-maternal circulation and hence efficient exchange of nutrients and oxygen between the embryo and maternal environment, leading to a general failure of embryonic cell proliferation and widespread apoptosis. Hence, Edd has an essential role in extraembryonic development. PMID- 15282323 TI - Cluster analysis of mass spectrometry data reveals a novel component of SAGA. AB - The SAGA histone acetyltransferase and TFIID complexes play key roles in eukaryotic transcription. Using hierarchical cluster analysis of mass spectrometry data to identify proteins that copurify with components of the budding yeast TFIID transcription complex, we discovered that an uncharacterized protein corresponding to the YPL047W open reading frame significantly associated with shared components of the TFIID and SAGA complexes. Using mass spectrometry and biochemical assays, we show that YPL047W (SGF11, 11-kDa SAGA-associated factor) is an integral subunit of SAGA. However, SGF11 does not appear to play a role in SAGA-mediated histone acetylation. DNA microarray analysis showed that SGF11 mediates transcription of a subset of SAGA-dependent genes, as well as SAGA independent genes. SAGA purified from a sgf11 Delta deletion strain has reduced amounts of Ubp8p, and a ubp8 Delta deletion strain shows changes in transcription similar to those seen with the sgf11 Delta deletion strain. Together, these data show that Sgf11p is a novel component of the yeast SAGA complex and that SGF11 regulates transcription of a subset of SAGA-regulated genes. Our data suggest that the role of SGF11 in transcription is independent of SAGA's histone acetyltransferase activity but may involve Ubp8p recruitment to or stabilization in SAGA. PMID- 15282322 TI - Deletion of mouse rad9 causes abnormal cellular responses to DNA damage, genomic instability, and embryonic lethality. AB - The fission yeast Schizosaccharomyces pombe rad9 gene promotes cell survival through activation of cell cycle checkpoints induced by DNA damage. Mouse embryonic stem cells with a targeted deletion of Mrad9, the mouse ortholog of this gene, were created to evaluate its function in mammals. Mrad9(-/-) cells demonstrated a marked increase in spontaneous chromosome aberrations and HPRT mutations, indicating a role in the maintenance of genomic integrity. These cells were also extremely sensitive to UV light, gamma rays, and hydroxyurea, and heterozygotes were somewhat sensitive to the last two agents relative to Mrad9(+/+) controls. Mrad9(-/-) cells could initiate but not maintain gamma-ray induced G(2) delay and retained the ability to delay DNA synthesis rapidly after UV irradiation, suggesting that checkpoint abnormalities contribute little to the radiosensitivity observed. Ectopic expression of Mrad9 or human HRAD9 complemented Mrad9(-/-) cell defects, indicating that the gene has radioresponse and genomic maintenance functions that are evolutionarily conserved. Mrad9(+/-) mice were generated, but heterozygous intercrosses failed to yield Mrad9(-/-) pups, since embryos died at midgestation. Furthermore, Mrad9(-/-) mouse embryo fibroblasts were not viable. These investigations establish Mrad9 as a key mammalian genetic element of pathways that regulate the cellular response to DNA damage, maintenance of genomic integrity, and proper embryonic development. PMID- 15282325 TI - EWS/FLI-1 silencing and gene profiling of Ewing cells reveal downstream oncogenic pathways and a crucial role for repression of insulin-like growth factor binding protein 3. AB - Ewing tumors are characterized by abnormal transcription factors resulting from the oncogenic fusion of EWS with members of the ETS family, most commonly FLI-1. RNA interference targeted to the junction between EWS and FLI-1 sequences was used to inactivate the EWS/FLI-1 fusion gene in Ewing cells and to explore the resulting phenotype and alteration of the gene expression profile. Loss of expression of EWS/FLI-1 resulted in the complete arrest of growth and was associated with a dramatic increase in the number of apoptotic cells. Gene profiling of Ewing cells in which the EWS/FLI-1 fusion gene had been inactivated identified downstream targets which could be grouped in two major functional clusters related to extracellular matrix structure or remodeling and regulation of signal transduction pathways. Among these targets, the insulin-like growth factor binding protein 3 gene (IGFBP-3), a major regulator of insulin-like growth factor 1 (IGF-1) proliferation and survival signaling, was strongly induced upon treating Ewing cells with EWS/FLI-1-specific small interfering RNAs. We show that EWS/FLI-1 can bind the IGFBP-3 promoter in vitro and in vivo and can repress its activity. Moreover, IGFBP-3 silencing can partially rescue the apoptotic phenotype caused by EWS/FLI-1 inactivation. Finally, IGFBP-3-induced Ewing cell apoptosis relies on both IGF-1-dependent and -independent pathways. These findings therefore identify the repression of IGFBP-3 as a key event in the development of Ewing's sarcoma. PMID- 15282324 TI - Estrogens and progesterone promote persistent CCND1 gene activation during G1 by inducing transcriptional derepression via c-Jun/c-Fos/estrogen receptor (progesterone receptor) complex assembly to a distal regulatory element and recruitment of cyclin D1 to its own gene promoter. AB - Transcriptional activation of the cyclin D1 gene (CCND1) plays a pivotal role in G(1)-phase progression, which is thereby controlled by multiple regulatory factors, including nuclear receptors (NRs). Appropriate CCND1 gene activity is essential for normal development and physiology of the mammary gland, where it is regulated by ovarian steroids through a mechanism(s) that is not fully elucidated. We report here that CCND1 promoter activation by estrogens in human breast cancer cells is mediated by recruitment of a c-Jun/c-Fos/estrogen receptor alpha complex to the tetradecanoyl phorbol acetate-responsive element of the gene, together with Oct-1 to a site immediately adjacent. This process coincides with the release from the same DNA region of a transcriptional repressor complex including Yin-Yang 1 (YY1) and histone deacetylase 1 and is sufficient to induce the assembly of the basal transcription machinery on the promoter and to lead to initial cyclin D1 accumulation in the cell. Later on in estrogen stimulation, the cyclin D1/Cdk4 holoenzyme associates with the CCND1 promoter, where E2F and pRb can also be found, contributing to the long-lasting gene enhancement required to drive G(1)-phase completion. Interestingly, progesterone triggers similar regulatory events through its own NRs, suggesting that the gene regulation cascade described here represents a crossroad for the transcriptional control of G(1)-phase progression by different classes of NRs. PMID- 15282326 TI - A plant snoRNP complex containing snoRNAs, fibrillarin, and nucleolin-like proteins is competent for both rRNA gene binding and pre-rRNA processing in vitro. AB - In eukaryotes the primary cleavage of the precursor rRNA (pre-rRNA) occurs in the 5' external transcribed spacer (5'ETS). In Saccharomyces cerevisiae and animals this cleavage depends on a conserved U3 small nucleolar ribonucleoprotein particle (snoRNP), including fibrillarin, and on other transiently associated proteins such as nucleolin. This large complex can be visualized by electron microscopy bound to the nascent pre-rRNA soon after initiation of transcription. Our group previously described a radish rRNA gene binding activity, NF D, that specifically binds to a cluster of conserved motifs preceding the primary cleavage site in the 5'ETS of crucifer plants including radish, cauliflower, and Arabidopsis thaliana (D. Caparros-Ruiz, S. Lahmy, S. Piersanti, and M. Echeverria, Eur. J. Biochem. 247:981-989, 1997). Here we report the purification and functional characterization of NF D from cauliflower inflorescences. Remarkably NF D also binds to 5'ETS RNA and accurately cleaves it at the primary cleavage site mapped in vivo. NF D is a multiprotein factor of 600 kDa that dissociates into smaller complexes. Two polypeptides of NF D identified by microsequencing are homologues of nucleolin and fibrillarin. The conserved U3 and U14 snoRNAs associated with fibrillarin and required for early pre-rRNA cleavages are also found in NF D. Based on this it is proposed that NF D is a processing complex that assembles on the rDNA prior to its interaction with the nascent pre rRNA. PMID- 15282327 TI - Protein kinase C delta blocks immediate-early gene expression in senescent cells by inactivating serum response factor. AB - Fibroblasts lose the ability to replicate in response to growth factors and become unable to express growth-associated immediate-early genes, including c-fos and egr-1, as they become senescent. The serum response factor (SRF), a major transcriptional activator of immediate-early gene promoters, loses the ability to bind to the serum response element (SRE) and becomes hyperphosphorylated in senescent cells. We identify protein kinase C delta (PKC delta) as the kinase responsible for inactivation of SRF both in vitro and endogenously in senescent cells. This is due to a higher level of PKC delta activity as cells age, production of the PKC delta catalytic fragment, and its nuclear localization in senescent but not in low-passage-number cells. The phosphorylation of T160 of SRF by PKC delta in vitro and in vivo led to loss of SRF DNA binding activity. Both the PKC delta inhibitor rottlerin and ectopic expression of a dominant negative form of PKC delta independently restored SRE-dependent transcription and immediate-early gene expression in senescent cells. Modulation of PKC delta activity in vivo with rottlerin or bistratene A altered senescent- and young-cell morphology, respectively. These observations support the idea that the coordinate transcriptional inhibition of several growth-associated genes by PKC delta contributes to the senescent phenotype. PMID- 15282329 TI - Heat capacity changes in RNA folding: application of perturbation theory to hammerhead ribozyme cold denaturation. AB - In proteins, empirical correlations have shown that changes in heat capacity (DeltaC(P)) scale linearly with the hydrophobic surface area buried upon folding. The influence of DeltaC(P) on RNA folding has been widely overlooked and is poorly understood. In addition to considerations of solvent reorganization, electrostatic effects might contribute to DeltaC(P)s of folding in polyanionic species such as RNAs. Here, we employ a perturbation method based on electrostatic theory to probe the hot and cold denaturation behavior of the hammerhead ribozyme. This treatment avoids much of the error associated with imposing two-state folding models on non-two-state systems. Ribozyme stability is perturbed across a matrix of solvent conditions by varying the concentration of NaCl and methanol co-solvent. Temperature-dependent unfolding is then monitored by circular dichroism spectroscopy. The resulting array of unfolding transitions can be used to calculate a DeltaC(P) of folding that accurately predicts the observed cold denaturation temperature. We confirm the accuracy of the calculated DeltaC(P) by using isothermal titration calorimetry, and also demonstrate a methanol-dependence of the DeltaC(P). We weigh the strengths and limitations of this method for determining DeltaC(P) values. Finally, we discuss the data in light of the physical origins of the DeltaC(P)s for RNA folding and consider their impact on biological function. PMID- 15282328 TI - Synthesis and processing of tRNA-related SINE transcripts in Arabidopsis thaliana. AB - Despite the ubiquitous distribution of tRNA-related short interspersed elements (SINEs) in eukaryotic species, very little is known about the synthesis and processing of their RNAs. In this work, we have characterized in detail the different RNA populations resulting from the expression of a tRNA-related SINE S1 founder copy in Arabidopsis thaliana. The main population is composed of poly(A) ending (pa) SINE RNAs, while two minor populations correspond to full-length (fl) or poly(A) minus [small cytoplasmic (sc)] SINE RNAs. Part of the poly(A) minus RNAs is modified by 3'-terminal addition of C or CA nucleotides. All three RNA populations accumulate in the cytoplasm. Using a mutagenesis approach, we show that the poly(A) region and the 3' end unique region, present at the founder locus, are both important for the maturation and the steady-state accumulation of the different S1 RNA populations. The observation that primary SINE transcripts can be post-transcriptionally processed in vivo into a poly(A)-ending species introduces the possibility that this paRNA is used as a retroposition intermediate. PMID- 15282330 TI - Transcriptionally competent chromatin assembled with exogenous histones in a yeast whole cell extract. AB - We describe a cell-free chromatin assembly system derived from the yeast Saccharomyces cerevisiae, which efficiently packages DNA into minichromosomes in a reaction dependent on exogenous core histones and an ATP-regenerating system. Both supercoiled and relaxed plasmid DNA serve as templates for nucleosomal loading in a gradual process that takes at least 6 h for completion at 30 degrees C. Micrococcal nuclease digestion of the assembled minichromosomes displays an extended nucleosomal ladder with a repeat length of 165 bp. The purified minichromosomes contain the four core histones in stoichiometric proportion and exhibit phased nucleosomes over the mouse mammary tumour virus (MMTV) promoter. The progesterone receptor and NF1 synergize on these minichromosomes resulting in efficient cell-free transcription. The ease of manipulation and the potential use of yeast strains carrying mutations in the chromatin handling machinery make this system suitable for detailed mechanistic studies. PMID- 15282331 TI - Recent origin of a hominoid-specific splice form of neuropsin, a gene involved in learning and memory. AB - Neuropsin is a secreted-type serine protease involved in learning and memory. The type II splice form of neuropsin is abundantly expressed in the human brain but not in the mouse brain. We sequenced the type II-spliced region of neuropsin gene in humans and representative nonhuman primate species. Our comparative sequence analysis showed that only the hominoid species (humans and apes) have the intact open reading frame of the type II splice form, indicating that the type II neuropsin originated recently in the primate lineage about 18 MYA. Expression analysis using RT-PCR detected abundant expression of the type II form in the frontal lobe of the adult human brain, but no expression was detected in the brains of lesser apes and Old World monkeys, indicating that the type II form of neuropsin only became functional in recent time, and it might contribute to the progressive change of cognitive abilities during primate evolution. PMID- 15282333 TI - Conservation and coevolution in the scale-free human gene coexpression network. AB - The role of natural selection in biology is well appreciated. Recently, however, a critical role for physical principles of network self-organization in biological systems has been revealed. Here, we employ a systems level view of genome-scale sequence and expression data to examine the interplay between these two sources of order, natural selection and physical self-organization, in the evolution of human gene regulation. The topology of a human gene coexpression network, derived from tissue-specific expression profiles, shows scale-free properties that imply evolutionary self-organization via preferential node attachment. Genes with numerous coexpressed partners (the hubs of the coexpression network) evolve more slowly on average than genes with fewer coexpressed partners, and genes that are coexpressed show similar rates of evolution. Thus, the strength of selective constraints on gene sequences is affected by the topology of the gene coexpression network. This connection is strong for the coding regions and 3' untranslated regions (UTRs), but the 5' UTRs appear to evolve under a different regime. Surprisingly, we found no connection between the rate of gene sequence divergence and the extent of gene expression profile divergence between human and mouse. This suggests that distinct modes of natural selection might govern sequence versus expression divergence, and we propose a model, based on rapid, adaptation-driven divergence and convergent evolution of gene expression patterns, for how natural selection could influence gene expression divergence. PMID- 15282332 TI - Divergence of conserved non-coding sequences: rate estimates and relative rate tests. AB - In many eukaryotic genomes only a small fraction of the DNA codes for proteins, but the non-protein coding DNA harbors important genetic elements directing the development and the physiology of the organisms, like promoters, enhancers, insulators, and micro-RNA genes. The molecular evolution of these genetic elements is difficult to study because their functional significance is hard to deduce from sequence information alone. Here we propose an approach to the study of the rate of evolution of functional non-coding sequences at a macro evolutionary scale. We identify functionally important non-coding sequences as Conserved Non-Coding Nucleotide (CNCN) sequences from the comparison of two outgroup species. The CNCN sequences so identified are then compared to their homologous sequences in a pair of ingroup species, and we monitor the degree of modification these sequences suffered in the two ingroup lineages. We propose a method to test for rate differences in the modification of CNCN sequences among the two ingroup lineages, as well as a method to estimate their rate of modification. We apply this method to the full sequences of the HoxA clusters from six gnathostome species: a shark, Heterodontus francisci; a basal ray finned fish, Polypterus senegalus; the amphibian, Xenopus tropicalis; as well as three mammalian species, human, rat and mouse. The results show that the evolutionary rate of CNCN sequences is not distinguishable among the three mammalian lineages, while the Xenopus lineage has a significantly increased rate of evolution. Furthermore the estimates of the rate parameters suggest that in the stem lineage of mammals the rate of CNCN sequence evolution was more than twice the rate observed within the placental amniotes clade, suggesting a high rate of evolution of cis-regulatory elements during the origin of amniotes and mammals. We conclude that the proposed methods can be used for testing hypotheses about the rate and pattern of evolution of putative cis-regulatory elements. PMID- 15282334 TI - Molecular evolution of sex-biased genes in Drosophila. AB - Studies of morphology, interspecific hybridization, protein/DNA sequences, and levels of gene expression have suggested that sex-related characters (particularly those involved in male reproduction) evolve rapidly relative to non sex-related characters. Here we report a general comparison of evolutionary rates of sex-biased genes using data from cDNA microarray experiments and comparative genomic studies of Drosophila. Comparisons of nonsynonymous/synonymous substitution rates (d(N)/d(S)) between species of the D. melanogaster subgroup revealed that genes with male-biased expression had significantly faster rates of evolution than genes with female-biased or unbiased expression. The difference was caused primarily by a higher d(N) in the male-biased genes. The same pattern was observed for comparisons among more distantly related species. In comparisons between D. melanogaster and D. pseudoobscura, genes with highly biased male expression were significantly more divergent than genes with highly biased female expression. In many cases, orthologs of D. melanogaster male-biased genes could not be identified in D. pseudoobscura through a Blast search. In contrast to the male-biased genes, there was no clear evidence for accelerated rates of evolution in female-biased genes, and most comparisons indicated a reduced rate of evolution in female-biased genes relative to unbiased genes. Male-biased genes did not show an increased ratio of nonsynonymous/synonymous polymorphism within D. melanogaster, and comparisons of polymorphism/divergence ratios suggest that the rapid evolution of male-biased genes is caused by positive selection. PMID- 15282336 TI - Golgi enzymes are enriched in perforated zones of golgi cisternae but are depleted in COPI vesicles. AB - In the most widely accepted version of the cisternal maturation/progression model of intra-Golgi transport, the polarity of the Golgi complex is maintained by retrograde transport of Golgi enzymes in COPI-coated vesicles. By analyzing enzyme localization in relation to the three-dimensional ultrastructure of the Golgi complex, we now observe that Golgi enzymes are depleted in COPI-coated buds and 50- to 60-nm COPI-dependent vesicles in a variety of different cell types. Instead, we find that Golgi enzymes are concentrated in the perforated zones of cisternal rims both in vivo and in a cell-free system. This lateral segregation of Golgi enzymes is detectable in some stacks during steady-state transport, but it was significantly prominent after blocking endoplasmic reticulum-to-Golgi transport. Delivery of transport carriers to the Golgi after the release of a transport block leads to a diminution in Golgi enzyme concentrations in perforated zones of cisternae. The exclusion of Golgi enzymes from COPI vesicles and their transport-dependent accumulation in perforated zones argues against the current vesicle-mediated version of the cisternal maturation/progression model. PMID- 15282335 TI - Insulin and wnt1 pathways cooperate to induce reserve cell activation in differentiation and myotube hypertrophy. AB - During ex vivo myoblast differentiation, a pool of quiescent mononucleated myoblasts, reserve cells, arise alongside myotubes. Insulin/insulin-like growth factor (IGF) and PKB/Akt-dependent phosphorylation activates skeletal muscle differentiation and hypertrophy. We have investigated the role of glycogen synthase kinase 3 (GSK-3) inhibition by protein kinase B (PKB)/Akt and Wnt/beta catenin pathways in reserve cell activation during myoblast differentiation and myotube hypertrophy. Inhibition of GSK-3 by LiCl or SB216763, restored insulin dependent differentiation of C2ind myoblasts in low serum, and cooperated with insulin in serum-free medium to induce MyoD and myogenin expression in C2ind myoblasts, quiescent C2 or primary human reserve cells. We show that LiCl treatment induced nuclear accumulation of beta-catenin in C2 myoblasts, thus mimicking activation of canonical Wnt signaling. Similarly to the effect of GSK-3 inhibitors with insulin, coculturing C2 reserve cells with Wnt1-expressing fibroblasts enhanced insulin-stimulated induction of MyoD and myogenin in reserve cells. A similar cooperative effect of LiCl or Wnt1 with insulin was observed during late ex vivo differentiation and promoted increased size and fusion of myotubes. We show that this synergistic effect on myotube hypertrophy involved an increased fusion of reserve cells into preexisting myotubes. These data reveal insulin and Wnt/beta-catenin pathways cooperate in muscle cell differentiation through activation and recruitment of satellite cell-like reserve myoblasts. PMID- 15282337 TI - Insufficient folding of type IV collagen and formation of abnormal basement membrane-like structure in embryoid bodies derived from Hsp47-null embryonic stem cells. AB - Hsp47 is a molecular chaperone that specifically recognizes procollagen in the endoplasmic reticulum. Hsp47-null mouse embryos produce immature type I collagen and form discontinuous basement membranes. We established Hsp47-/- embryonic stem cell lines and examined formation of basement membrane and production of type IV collagen in embryoid bodies, a model for postimplantation egg-cylinder stage embryos. The visceral endodermal cell layers surrounding Hsp47-/- embryoid bodies were often disorganized, a result that suggested abnormal function of the basement membrane under the visceral endoderm. Rate of type IV collagen secretion by Hsp47-/- cells was fourfold lower than that of Hsp47+/+ cells. Furthermore, type IV collagen secreted from Hsp47-/- cells was much more sensitive to protease digestion than was type IV collagen secreted from Hsp47+/+ cells, which suggested insufficient or incorrect triple helix formation in type IV collagen in the absence of Hsp47. These results indicate for the first time that Hsp47 is required for the molecular maturation of type IV collagen and suggest that misfolded type IV collagen causes abnormal morphology of embryoid bodies. PMID- 15282338 TI - A Ras-like GTPase is involved in hyphal growth guidance in the filamentous fungus Ashbya gossypii. AB - Characteristic features of morphogenesis in filamentous fungi are sustained polar growth at tips of hyphae and frequent initiation of novel growth sites (branches) along the extending hyphae. We have begun to study regulation of this process on the molecular level by using the model fungus Ashbya gossypii. We found that the A. gossypii Ras-like GTPase Rsr1p/Bud1p localizes to the tip region and that it is involved in apical polarization of the actin cytoskeleton, a determinant of growth direction. In the absence of RSR1/BUD1, hyphal growth was severely slowed down due to frequent phases of pausing of growth at the hyphal tip. During pausing events a hyphal tip marker, encoded by the polarisome component AgSPA2, disappeared from the tip as was shown by in vivo time-lapse fluorescence microscopy of green fluorescent protein-labeled AgSpa2p. Reoccurrence of AgSpa2p was required for the resumption of hyphal growth. In the Agrsr1/bud1Delta deletion mutant, resumption of growth occurred at the hyphal tip in a frequently uncoordinated manner to the previous axis of polarity. Additionally, hyphal filaments in the mutant developed aberrant branching sites by mislocalizing AgSpa2p thus distorting hyphal morphology. These results define AgRsr1p/Bud1p as a key regulator of hyphal growth guidance. PMID- 15282339 TI - Visualization of regulated exocytosis with a granule-membrane probe using total internal reflection microscopy. AB - Secretory granules labeled with Vamp-green fluorescent protein (GFP) showed distinct signatures upon exocytosis when viewed by total internal reflection fluorescence microscopy. In approximately 90% of fusion events, we observed a large increase in fluorescence intensity coupled with a transition from a small punctate appearance to a larger, spreading cloud with free diffusion of the Vamp GFP into the plasma membrane. Quantitation suggests that these events reflect the progression of an initially fused and spherical granule flattening into the plane of the plasma membrane as the Vamp-GFP simultaneously diffuses through the fusion junction. Approximately 10% of the events showed a transition from puncta to ring like structures coupled with little or no spreading. The ring-like images correspond quantitatively to granules fusing and retaining concavity (recess of approximately 200 nm). A majority of fusion events involved granules that were present in the evanescent field for at least 12 s. However, approximately 20% of the events involved granules that were present in the evanescent field for no more than 0.3 s, indicating that the interaction of the granule with the plasma membrane that leads to exocytosis can occur within that time. In addition, approximately 10% of the exocytotic sites were much more likely to occur within a granule diameter of a previous event than can be accounted for by chance, suggestive of sequential (piggy-back) exocytosis that has been observed in other cells. Overall granule behavior before and during fusion is strikingly similar to exocytosis previously described in the constitutive secretory pathway. PMID- 15282340 TI - Defective epidermal barrier in neonatal mice lacking the C-terminal region of connexin43. AB - More than 97% of mice in which the C-terminal region of connexin43 (Cx43) was removed (designated as Cx43K258stop) die shortly after birth due to a defect of the epidermal barrier. The abnormal expression of Cx43K258stop protein in the uppermost layers of the epidermis seems to perturb terminal differentiation of keratinocytes. In contrast to Cx43-deficient mice, neonatal Cx43K258stop hearts show no lethal obstruction of the right ventricular outflow tract, but signs of dilatation. Electrocardiographies of neonatal hearts reveal repolarization abnormalities in 20% of homozygous Cx43K258stop animals. The very rare adult Cx43K258stop mice show a compensation of the epidermal barrier defect but persisting impairment of cardiac function in echocardiography. Female Cx43K258stop mice are infertile due to impaired folliculogenesis. Our results indicate that the C-terminally truncated Cx43K258stop mice lack essential functions of Cx43, although the truncated Cx43 protein can form open gap junctional channels. PMID- 15282341 TI - Protein-protein interactions governing septin heteropentamer assembly and septin filament organization in Saccharomyces cerevisiae. AB - Mitotic yeast (Saccharomyces cerevisiae) cells express five related septins (Cdc3, Cdc10, Cdc11, Cdc12, and Shs1) that form a cortical filamentous collar at the mother-bud neck necessary for normal morphogenesis and cytokinesis. All five possess an N-terminal GTPase domain and, except for Cdc10, a C-terminal extension (CTE) containing a predicted coiled coil. Here, we show that the CTEs of Cdc3 and Cdc12 are essential for their association and for the function of both septins in vivo. Cdc10 interacts with a Cdc3-Cdc12 complex independently of the CTE of either protein. In contrast to Cdc3 and Cdc12, the Cdc11 CTE, which recruits the nonessential septin Shs1, is dispensable for its function in vivo. In addition, Cdc11 forms a stoichiometric complex with Cdc12, independent of its CTE. Reconstitution of various multiseptin complexes and electron microscopic analysis reveal that Cdc3, Cdc11, and Cdc12 are all necessary and sufficient for septin filament formation, and presence of Cdc10 causes filament pairing. These data provide novel insights about the connectivity among the five individual septins in functional septin heteropentamers and the organization of septin filaments. PMID- 15282342 TI - Targets of fibroblast growth factor 1 (FGF-1) and FGF-2 signaling involved in the invasive and tumorigenic behavior of carcinoma cells. AB - Fibroblast growth factor (FGF)-1 and -2 have potent biological activities implicated in malignant tumor development. Their autocrine and nonautocrine activity in tumor progression of carcinoma was investigated in the NBT-II cell system. Cells were manipulated to either produce and be autocrine for FGF-1 or -2 or to only produce but not respond to these factors. The autocrine cells are highly invasive and tumorigenic and the determination of specific targets of FGF/fibroblast growth factor receptor (FGFR) signaling was assessed. In vitro studies showed that nonautocrine cells behave like epithelial parental cells, whereas autocrine cells have a mesenchymal phenotype correlated with the overexpression of urokinase plasminogen activator receptor (uPAR), the internalization of E-cadherin, and the redistribution of beta-catenin from the cell surface to the cytoplasm and nucleus. uPAR was defined as an early target, whereas E-cadherin and the leukocyte common antigen-related protein-tyrosine phosphatase (LAR-PTP) were later targets of FGF signaling, with FGFR1 activation more efficient than FGFR2 at modulating these targets. Behavior of autocrine cells was consistent with a decrease of tumor-suppressive activities of both E cadherin and LAR-PTP. These molecular analyses show that the potential of these two growth factors in tumor progression is highly dependent on specific FGFR signaling and highlights its importance as a target for antitumor therapy. PMID- 15282343 TI - Induction of vascular smooth muscle alpha-actin gene transcription in transforming growth factor beta1-activated myofibroblasts mediated by dynamic interplay between the Pur repressor proteins and Sp1/Smad coactivators. AB - The mouse vascular smooth muscle alpha-actin (SMA) gene enhancer is activated in fibroblasts by transforming growth factor beta1 (TGFbeta1), a potent mediator of myofibroblast differentiation and wound healing. The SMA enhancer contains tandem sites for the Sp1 transcriptional activator protein and Puralpha and beta repressor proteins. We have examined dynamic interplay between these divergent proteins to identify checkpoints for possible control of myofibroblast differentiation during chronic inflammatory disease. A novel element in the SMA enhancer named SPUR was responsible for both basal and TGFbeta1-dependent transcriptional activation in fibroblasts and capable of binding Sp1 and Pur proteins. A novel Sp1:Pur:SPUR complex was dissociated when SMA enhancer activity was increased by TGFbeta1 or Smad protein overexpression. Physical association of Pur proteins with Smad2/3 was observed as was binding of Smads to an upstream enhancer region that undergoes DNA duplex unwinding in TGFbeta1-activated myofibroblasts. Purbeta repression of the SMA enhancer could not be relieved by TGFbeta1, whereas repression mediated by Puralpha was partially rescued by TGFbeta1 or overexpression of Smad proteins. Interplay between Pur repressor isoforms and Sp1 and Smad coactivators may regulate SMA enhancer output in TGFbeta1-activated myofibroblasts during episodes of wound repair and tissue remodeling. PMID- 15282345 TI - Doctors and torture. PMID- 15282346 TI - In the name of public health--Nazi racial hygiene. PMID- 15282344 TI - PAK kinases Ste20 and Pak1 govern cell polarity at different stages of mating in Cryptococcus neoformans. AB - Sexual identity and mating are linked to virulence of the fungal pathogen Cryptococcus neoformans. Cells of the alpha mating type are more prevalent and can be more virulent than a cells, and basidiospores are thought to be the infectious propagule. Mating in C. neoformans involves cell-cell fusion and the generation of dikaryotic hyphae, processes that involve substantial changes in cell polarity. Two p21-activated kinase (PAK) kinases, Pak1 and Ste20, are required for both mating and virulence in C. neoformans. We show here that Ste20 and Pak1 play crucial roles in polarized morphogenesis at different steps during mating: Pak1 functions during cell fusion, whereas Ste20 fulfills a distinct morphogenic role and is required to maintain polarity in the heterokaryotic mating filament. In conclusion, our studies demonstrate that PAK kinases are necessary for polar growth during mating and that polarity establishment is necessary for mating and may contribute to virulence of C. neoformans. PMID- 15282347 TI - Mind reading. PMID- 15282348 TI - Treating hepatitis C in "difficult-to-treat" patients. PMID- 15282349 TI - Myosin at the heart of the problem. PMID- 15282350 TI - Efficacy of MRI and mammography for breast-cancer screening in women with a familial or genetic predisposition. AB - BACKGROUND: The value of regular surveillance for breast cancer in women with a genetic or familial predisposition to breast cancer is currently unproven. We compared the efficacy of magnetic resonance imaging (MRI) with that of mammography for screening in this group of high-risk women. METHODS: Women who had a cumulative lifetime risk of breast cancer of 15 percent or more were screened every six months with a clinical breast examination and once a year by mammography and MRI, with independent readings. The characteristics of the cancers that were detected were compared with the characteristics of those in two different age-matched control groups. RESULTS: We screened 1909 eligible women, including 358 carriers of germ-line mutations. Within a median follow-up period of 2.9 years, 51 tumors (44 invasive cancers, 6 ductal carcinomas in situ, and 1 lymphoma) and 1 lobular carcinoma in situ were detected. The sensitivity of clinical breast examination, mammography, and MRI for detecting invasive breast cancer was 17.9 percent, 33.3 percent, and 79.5 percent, respectively, and the specificity was 98.1 percent, 95.0 percent, and 89.8 percent, respectively. The overall discriminating capacity of MRI was significantly better than that of mammography (P<0.05). The proportion of invasive tumors that were 10 mm or less in diameter was significantly greater in our surveillance group (43.2 percent) than in either control group (14.0 percent [P<0.001] and 12.5 percent [P=0.04], respectively). The combined incidence of positive axillary nodes and micrometastases in invasive cancers in our study was 21.4 percent, as compared with 52.4 percent (P<0.001) and 56.4 percent (P=0.001) in the two control groups. CONCLUSIONS: MRI appears to be more sensitive than mammography in detecting tumors in women with an inherited susceptibility to breast cancer. PMID- 15282351 TI - Peginterferon Alfa-2a plus ribavirin for chronic hepatitis C virus infection in HIV-infected patients. AB - BACKGROUND: Hepatitis C virus (HCV) infection is highly prevalent and is associated with substantial morbidity and mortality among persons infected with the human immunodeficiency virus (HIV). We compared the efficacy and safety of pegylated interferon alfa-2a (peginterferon alfa-2a) plus either ribavirin or placebo with those of interferon alfa-2a plus ribavirin for the treatment of chronic HCV infection in patients who were also infected with HIV. METHODS: A total of 868 persons who were infected with both HIV and HCV and who had not previously been treated with interferon or ribavirin were randomly assigned to receive one of three regimens: peginterferon alfa-2a (180 microg per week) plus ribavirin (800 mg per day), peginterferon alfa-2a plus placebo, or interferon alfa-2a (3 million IU three times a week) plus ribavirin. Patients were treated for 48 weeks and followed for an additional 24 weeks. The primary end point was a sustained virologic response (defined as a serum HCV RNA level below 50 IU per milliliter at the end of follow-up, at week 72). RESULTS: The overall rate of sustained virologic response was significantly higher among the recipients of peginterferon alfa-2a plus ribavirin than among those assigned to interferon alfa 2a plus ribavirin (40 percent vs. 12 percent, P<0.001), or peginterferon alfa-2a plus placebo (40 percent vs. 20 percent, P<0.001). Among patients infected with HCV genotype 1, the rates of sustained virologic response were 29 percent with peginterferon alfa-2a plus ribavirin, 14 percent with peginterferon alfa-2a plus placebo, and 7 percent with interferon alfa-2a plus ribavirin. The corresponding rates among patients infected with HCV genotype 2 or 3 were 62 percent, 36 percent, and 20 percent. Neutropenia and thrombocytopenia were more common among patients treated with regimens that contained peginterferon alfa-2a, and anemia was more common among patients treated with regimens containing ribavirin. CONCLUSIONS: Among patients infected with both HIV and HCV, the combination of peginterferon alfa-2a plus ribavirin was significantly more effective than either interferon alfa-2a plus ribavirin or peginterferon alfa-2a monotherapy. PMID- 15282352 TI - Peginterferon Alfa-2a plus ribavirin versus interferon alfa-2a plus ribavirin for chronic hepatitis C in HIV-coinfected persons. AB - BACKGROUND: Chronic hepatitis C virus (HCV) infection is a cause of major complications in persons who are also infected with the human immunodeficiency virus (HIV). However, the treatment of HCV infection in such persons has been associated with a high rate of intolerance and a low rate of response. We conducted a multicenter, randomized trial comparing peginterferon plus ribavirin with interferon plus ribavirin for the treatment of chronic hepatitis C in persons coinfected with HIV. METHODS: A total of 66 subjects were randomly assigned to receive 180 microg of peginterferon alfa-2a weekly for 48 weeks, and 67 subjects were assigned to receive 6 million IU of interferon alfa-2a three times weekly for 12 weeks followed by 3 million IU three times weekly for 36 weeks. Both groups received ribavirin according to a dose-escalation schedule. At week 24, subjects who did not have a virologic response (those who had an HCV RNA level greater than or equal to 60 IU per milliliter) underwent liver biopsy, and medications were continued in subjects with either a virologic response or histologic improvement. RESULTS: Treatment with peginterferon and ribavirin was associated with a significantly higher rate of sustained virologic response (an HCV RNA level of less than 60 IU per milliliter 24 weeks after completion of therapy) than was treatment with interferon and ribavirin (27 percent vs. 12 percent, P=0.03). In the group given peginterferon and ribavirin, only 14 percent of subjects with HCV genotype 1 infection had a sustained virologic response (7 of 51), as compared with 73 percent of subjects with an HCV genotype other than 1 (11 of 15, P<0.001). Histologic responses were observed in 35 percent of subjects with no virologic response who underwent liver biopsy. CONCLUSIONS: In persons infected with HIV, the combination of peginterferon and ribavirin is superior to the combination of interferon and ribavirin in the treatment of chronic hepatitis C. These regimens may provide clinical benefit even in the absence of virologic clearance. The marked discrepancy in the rates of sustained virologic response between HCV genotypes indicates that strategies are needed to improve the outcome in persons infected with HCV genotype 1. PMID- 15282353 TI - Mutation of perinatal myosin heavy chain associated with a Carney complex variant. AB - BACKGROUND: Familial cardiac myxomas occur in the hereditary syndrome Carney complex. Although PRKAR1A mutations can cause the Carney complex, the disorder is genetically heterogeneous. To identify the cause of a Carney complex variant associated with distal arthrogryposis (the trismus-pseudocamptodactyly syndrome), we performed clinical and genetic studies. METHODS: A large family with familial cardiac myxomas and the trismus-pseudocamptodactyly syndrome (Family 1) was identified and clinically evaluated along with two families with trismus and pseudocamptodactyly. Genetic linkage analyses were performed with the use of microsatellite polymorphisms to determine a locus for this Carney complex variant. Positional cloning and mutational analyses of candidate genes were performed to identify the genetic cause of disease in the family with the Carney complex as well as in the families with the trismus-pseudocamptodactyly syndrome. RESULTS: Clinical evaluations demonstrated that the Carney complex cosegregated with the trismus-pseudocamptodactyly syndrome in Family 1, and genetic analyses demonstrated linkage of the disease to chromosome 17p12-p13.1 (maximum multipoint lod score, 4.39). Sequence analysis revealed a missense mutation (Arg674Gln) in the perinatal myosin heavy-chain gene (MYH8). The same mutation was also found in the two families with the trismus-pseudocamptodactyly syndrome. Arg674 is highly conserved evolutionarily, localizes to the actin-binding domain of the perinatal myosin head, and is close to the ATP-binding site. We identified nonsynonymous MYH8 polymorphisms in patients with cardiac myxoma syndromes but without arthrogryposis. CONCLUSIONS: We describe a novel heart-hand syndrome involving familial cardiac myxomas and distal arthrogryposis and demonstrate that these disorders are caused by a founder mutation in the MYH8 gene. Our findings demonstrate novel roles for perinatal myosin in both the development of skeletal muscle and cardiac tumorigenesis. PMID- 15282354 TI - Clinical practice. Use of lasers for vision correction of nearsightedness and farsightedness. PMID- 15282355 TI - Bipolar disorder. PMID- 15282356 TI - Images in clinical medicine. The chemosis of trichinosis. PMID- 15282357 TI - Case records of the Massachusetts General Hospital. Weekly clinicopathological exercises. Case 24-2004. A 48-year-old man with recurrent gastrointestinal bleeding. PMID- 15282358 TI - Breast cancer screening with MRI--what are the data for patients at high risk? PMID- 15282359 TI - Beyond fast track for drug approvals. PMID- 15282360 TI - Cord-blood transplants from unrelated donors in Hurler's syndrome. PMID- 15282361 TI - The clinical-skills examination. PMID- 15282362 TI - Fibromuscular dysplasia. PMID- 15282363 TI - Case 15-2004: cancer therapy and sperm banking. PMID- 15282364 TI - Transient improvement of spinocerebellar ataxia with zolpidem. PMID- 15282365 TI - Images in clinical medicine. Electrocardiographic changes in extreme hypothermia. PMID- 15282366 TI - Progressive supranuclear palsy. AB - In this case study, we describe the symptoms, neurological examination, clinical course, and neuropathology of a patient with progressive supranuclear palsy (PSP). PSP is a relatively uncommon neurodegenerative disorder with many features similar to those of Parkinson's disease. It is characterized by slow motor function, ocular movement abnormalities, dystonia, and cognitive disabilities. PSP is largely a sporadic disorder caused by accumulation of the protein tau in diverse regions of the central nervous system. It is thus classified as one of several tauopathies. The exact cause of the disease remains unknown, and treatment is often limited. The following case provides a framework to explore the manifestations of PSP, as well as the progress made in understanding the nature of this challenging disorder. PMID- 15282367 TI - (Data)base desires. PMID- 15282368 TI - High anxiety. PMID- 15282369 TI - Growing pains. PMID- 15282370 TI - Noncompetitive allosteric inhibitors of the inflammatory chemokine receptors CXCR1 and CXCR2: prevention of reperfusion injury. AB - The chemokine CXC ligand 8 (CXCL8)/IL-8 and related agonists recruit and activate polymorphonuclear cells by binding the CXC chemokine receptor 1 (CXCR1) and CXCR2. Here we characterize the unique mode of action of a small-molecule inhibitor (Repertaxin) of CXCR1 and CXCR2. Structural and biochemical data are consistent with a noncompetitive allosteric mode of interaction between CXCR1 and Repertaxin, which, by locking CXCR1 in an inactive conformation, prevents signaling. Repertaxin is an effective inhibitor of polymorphonuclear cell recruitment in vivo and protects organs against reperfusion injury. Targeting the Repertaxin interaction site of CXCR1 represents a general strategy to modulate the activity of chemoattractant receptors. PMID- 15282371 TI - Eshkol-Wachman movement notation in diagnosis: the early detection of Asperger's syndrome. AB - The diagnostic criteria of Asperger's syndrome (AS), considered a part of the autistic spectrum disorder, are still unclear. A critical marker, which distinguishes AS from autism, is the presence of language. The ability of a child with AS to acquire and use language early results in the fact that AS usually is diagnosed much later than autism. Autism is not usually diagnosed until around the age of 3, whereas AS usually is not diagnosed until the child is 6 or 7 years of age. In the present article, using Eshkol-Wachman movement notation, we present evidence that abnormal movement patterns can be detected in AS in infancy. This finding suggests that AS can be diagnosed very early, independent of the presence of language. As shown earlier by us, almost all of the movement disturbances in autism can be interpreted as infantile reflexes "gone astray"; i.e., some reflexes are not inhibited at the appropriate age in development, whereas others fail to appear when they should. This phenomenon appears to apply to AS as well. Based on preliminary results, a simple test using one such reflex is proposed for the early detection of a subgroup of children with AS or autism. PMID- 15282372 TI - Genome-wide molecular dissection of serotype M3 group A Streptococcus strains causing two epidemics of invasive infections. AB - Molecular factors that contribute to the emergence of new virulent bacterial subclones and epidemics are poorly understood. We hypothesized that analysis of a population-based strain sample of serotype M3 group A Streptococcus (GAS) recovered from patients with invasive infection by using genome-wide investigative methods would provide new insight into this fundamental infectious disease problem. Serotype M3 GAS strains (n = 255) cultured from patients in Ontario, Canada, over 11 years and representing two distinct infection peaks were studied. Genetic diversity was indexed by pulsed-field gel electrophoresis, DNA DNA microarray, whole-genome PCR scanning, prophage genotyping, targeted gene sequencing, and single-nucleotide polymorphism genotyping. All variation in gene content was attributable to acquisition or loss of prophages, a molecular process that generated unique combinations of proven or putative virulence genes. Distinct serotype M3 genotypes experienced rapid population expansion and caused infections that differed significantly in character and severity. Molecular genetic analysis, combined with immunologic studies, implicated a 4-aa duplication in the extreme N terminus of M protein as a factor contributing to an epidemic wave of serotype M3 invasive infections. This finding has implications for GAS vaccine research. Genome-wide analysis of population-based strain samples cultured from clinically well defined patients is crucial for understanding the molecular events underlying bacterial epidemics. PMID- 15282373 TI - Environmental mutagenesis during the end-Permian ecological crisis. AB - During the end-Permian ecological crisis, terrestrial ecosystems experienced preferential dieback of woody vegetation. Across the world, surviving herbaceous lycopsids played a pioneering role in repopulating deforested terrain. We document that the microspores of these lycopsids were regularly released in unseparated tetrads indicative of failure to complete the normal process of spore development. Although involvement of mutation has long been hinted at or proposed in theory, this finding provides concrete evidence for chronic environmental mutagenesis at the time of global ecological crisis. Prolonged exposure to enhanced UV radiation could account satisfactorily for a worldwide increase in land plant mutation. At the end of the Permian, a period of raised UV stress may have been the consequence of severe disruption of the stratospheric ozone balance by excessive emission of hydrothermal organohalogens in the vast area of Siberian Traps volcanism. PMID- 15282374 TI - An antiproliferative factor from interstitial cystitis patients is a frizzled 8 protein-related sialoglycopeptide. AB - Approximately 1 million people in the United States suffer from interstitial cystitis, a chronic painful urinary bladder disorder characterized by thinning or ulceration of the bladder epithelial lining; its etiology is unknown. We have identified a glycosylated frizzled-related peptide inhibitor of cell proliferation that is secreted specifically by bladder epithelial cells from patients with this disorder. This antiproliferative factor (APF) profoundly inhibits bladder cell proliferation by means of regulation of cell adhesion protein and growth factor production. The structure of APF was deduced by using ion trap mass spectrometry (MS), enzymatic digestion, lectin affinity chromatography, and total synthesis, and confirmed by coelution of native and synthetic APF derivatives on microcapillary reversed-phase liquid chromatography (microRPLC)/MS. APF was determined to be an acidic, heat-stable sialoglycopeptide whose peptide chain has 100% homology to the putative sixth transmembrane domain of frizzled 8. Both synthetic and native APF had identical biological activity in normal bladder epithelial cells and T24 bladder cancer cells. Northern blot analysis indicated binding of a probe containing the sequence for the frizzled 8 segment with mRNA extracted from cells of patients with interstitial cystitis but not controls. APF is therefore a frizzled-related peptide growth inhibitor shown to contain exclusively a transmembrane segment of a frizzled protein and is a potential biomarker for interstitial cystitis. PMID- 15282375 TI - Time-resolved energy transfer in DNA sequence detection using water-soluble conjugated polymers: the role of electrostatic and hydrophobic interactions. AB - We have investigated the energy transfer processes in DNA sequence detection by using cationic conjugated polymers and peptide nucleic acid (PNA) probes with ultrafast pump-dump-emission spectroscopy. Pump-dump-emission spectroscopy provides femtosecond temporal resolution and high sensitivity and avoids interference from the solvent response. The energy transfer from donor (the conjugated polymer) to acceptor (a fluorescent molecule attached to a PNA terminus) has been time resolved. The results indicate that both electrostatic and hydrophobic interactions contribute to the formation of cationic conjugated polymers/PNA-C/DNA complexes. The two interactions result in two different binding conformations. This picture is supported by the average donor-acceptor separations as estimated from time-resolved and steady-state measurements. Electrostatic interactions dominate at low concentrations and in mixed solvents. PMID- 15282376 TI - Genome-wide discovery of loci influencing chemotherapy cytotoxicity. AB - Little is known about the heritability of chemotherapy activity or the identity of genes that may enable the individualization of cancer chemotherapy. Although numerous genes are likely to influence chemotherapy response, current candidate gene-based pharmacogenetics approaches require a priori knowledge and the selection of a small number of candidate genes for hypothesis testing. In this study, an ex vivo familial genetics strategy using lymphoblastoid cells derived from Centre d'Etude du Polymorphisme Humain reference pedigrees was used to discover genetic determinants of chemotherapy cytotoxicity. Cytotoxicity to the mechanistically distinct chemotherapy agents 5-fluorouracil and docetaxel were shown to be heritable traits, with heritability values ranging from 0.26 to 0.65 for 5-fluorouracil and 0.21 to 0.70 for docetaxel, varying with dose. Genome-wide linkage analysis was also used to map a quantitative trait locus influencing the cellular effects of 5-fluorouracil to chromosome 9q13-q22 [logarithm of odds (LOD) = 3.44], and two quantitative trait loci influencing the cellular effects of docetaxel to chromosomes 5q11-21 (LOD = 2.21) and 9q13-q22 (LOD = 2.73). Finally, 5-fluorouracil and docetaxel were shown to cause apoptotic cell death involving caspase-3 cleavage in Centre d'Etude du Polymorphisme Humain lymphoblastoid cells. This study identifies genomic regions likely to harbor genes important for chemotherapy cytotoxicity using genome-wide linkage analysis in human pedigrees and provides a widely applicable strategy for pharmacogenomic discovery without the requirement for a priori candidate gene selection. PMID- 15282377 TI - Hematopoietic cells and osteoblasts are derived from a common marrow progenitor after bone marrow transplantation. AB - Bone and bone marrow are closely aligned physiologic compartments, suggesting that these tissues may represent a single functional unit with a common bone marrow progenitor that gives rise to both osteoblasts and hematopoietic cells. Although reports of multilineage engraftment by a single marrow-derived stem cell support this idea, more recent evidence has challenged claims of stem cell transdifferentiation and therefore the existence of a multipotent hematopoietic/osteogenic progenitor cell. Using a repopulation assay in mice, we show here that gene-marked, transplantable marrow cells from the plastic nonadherent population can generate both functional osteoblasts/osteocytes and hematopoietic cells. Fluorescent in situ hybridization for the X and Y chromosomes and karyotype analysis of cultured osteoblasts confirmed the donor origin of these cells and excluded their generation by a fusion process. Molecular analysis demonstrated a common retroviral integration site in clonogenic hematopoietic cells and osteoprogenitors from each of seven animals studied, establishing a shared clonal origin for these ostensibly independent cell types. Our findings indicate that the bone marrow contains a primitive cell able to generate both the hematopoietic and osteocytic lineages. Its isolation and characterization may suggest novel treatments for genetic bone diseases and bone injuries. PMID- 15282378 TI - Asthma prevalence in adults: good news? PMID- 15282379 TI - Short burst oxygen therapy for relief of breathlessness in COPD. PMID- 15282380 TI - Regulation: the art of control? Regulatory T cells and asthma and allergy. PMID- 15282381 TI - Antiviral agents and corticosteroids in the treatment of severe acute respiratory syndrome (SARS). PMID- 15282382 TI - Increase in diagnosed asthma but not in symptoms in the European Community Respiratory Health Survey. AB - BACKGROUND: Information on the epidemiology of asthma in relation to age is limited and hampered by reporting error. To determine the change in the prevalence of asthma with age in young adults we analysed longitudinal data from the European Community Respiratory Health Survey. METHODS: A self-administered questionnaire was completed by 11 168 randomly selected subjects in 14 countries in 1991-3 when they were aged 20-44 years and 5-11 years later from 1998 to 2003. Generalised estimating equations were used to estimate net change in wheeze, nocturnal tightness in chest, shortness of breath, coughing, asthma attacks in the last 12 months, current medication, "diagnosed" asthma, and nasal allergies. RESULTS: Expressed as change in status per 10 years of follow up, subjects reporting asthma attacks in the previous 12 months increased by 0.8% of the population (95% CI 0.2 to 1.4) and asthma medication by 2.1% (95% CI 1.6 to 2.6), while no statistically significant net change was found in reported symptoms. Reported nasal allergies increased, especially in the youngest age group. CONCLUSIONS: As this cohort of young adults has aged, there has been an increase in the proportion treated for asthma but not in the proportion of those reporting symptoms suggestive of asthma. Either increased use of effective treatments has led to decreased morbidity among asthmatic subjects or those with mild disease have become more likely to label themselves as asthmatic. PMID- 15282383 TI - Vitamin E supplements in asthma: a parallel group randomised placebo controlled trial. AB - BACKGROUND: Increased dietary vitamin E intake is associated with a reduced incidence of asthma, and combinations of antioxidant supplements including vitamin E are effective in reducing ozone induced bronchoconstriction. A study was undertaken to investigate the effect of supplementation with vitamin E for 6 weeks on bronchial hyperresponsiveness in atopic adults with asthma. METHODS: 72 participants from a clinical trial register of adults with asthma were randomised to receive 500 mg natural vitamin E or matched placebo for 6 weeks in a placebo controlled, double blind parallel group clinical trial. Inclusion criteria included age 18-60 years, maintenance treatment of at least one dose of inhaled corticosteroid per day, a positive skin prick test to one of three common allergens, and bronchial hyperresponsiveness to methacholine (defined as a dose provoking a 20% fall in forced expiratory volume in 1 second (FEV(1)) (PD(20)) of 12.25 micromol). Secondary outcomes were FEV(1), forced vital capacity, mean morning and evening peak flow, symptom scores, bronchodilator use, and serum immunoglobulin E levels. RESULTS: In the primary intention to treat analysis the change in PD(20) was similar in the vitamin E and placebo groups with a mean difference of +0.25 doubling doses of methacholine (95% confidence interval -0.67 to +1.16 greater with vitamin E). There was no effect of vitamin E supplementation on any other measure of asthma control, either in the intention to treat or per protocol analysis. There was also no effect of vitamin E supplementation on serum immunoglobulin levels. CONCLUSION: Dietary supplementation with vitamin E adds no benefit to current standard treatment in adults with mild to moderate asthma. PMID- 15282384 TI - Fluticasone induces T cell apoptosis in the bronchial wall of mild to moderate asthmatics. AB - BACKGROUND: Cytokines which signal via the gamma chain of the interleukin (IL)-2 receptor and the interferons (IFNs) have been shown to enhance T cell survival in vitro by rescuing cells from apoptosis. METHODS: A study was undertaken to determine whether treatment with inhaled fluticasone propionate (FP; 250 microg twice daily) for 2 weeks could modulate production of IL-15 or IFN-beta and thereby affect T cell survival in bronchial tissue of 10 patients with mild/moderate asthma. Bronchial biopsy specimens were taken before and on completion of treatment. RESULTS: The mean (95% CI) number of T cells per unit area decreased in the asthmatic group following 2 weeks of treatment with FP (from 7.0 (5.6 to 8.4) to 4.5 (4.0 to 5.1); p = 0.001). There was an increase in the percentage of T cells undergoing apoptosis following FP treatment as assessed by T cell/TUNEL staining (from 4.5 (2.6 to 6.4) to 8.7 (6.6 to 10.8); p = 0.0001). The percentage of cells staining for IL-15 and IFN-beta in the lamina propria, determined by an alkaline phosphatase biotin streptavidin technique, decreased significantly from baseline values of 31.6 (23.4 to 39.7) to 19.6 (12.5 to 26.7), p = 0.039 for IL-15 and from 18.9 (13.5 to 24.4) to 9.5 (5.9 to 13.1), p = 0.007 for IFN-beta following 2 weeks of treatment with FP. However, only the decrease in the percentage of cells staining for IL-15 was significantly correlated with an increased number of apoptotic T cells following treatment (p = 0.008). CONCLUSION: These findings support a novel mechanism for the ability of inhaled corticosteroids to decrease T cell numbers, possibly by downregulation of the cytokine IL-15. PMID- 15282385 TI - Cross tolerance to salbutamol occurs independently of beta2 adrenoceptor genotype 16 in asthmatic patients receiving regular formoterol or salmeterol. AB - BACKGROUND: The development of tolerance following the use of long acting beta(2) agonists in asthmatic patients with either the homozygous arginine (Arg-16) or glycine (Gly-16) genotypes is poorly documented, especially in relation to the acute reliever response to salbutamol in constricted airways. A study was undertaken to evaluate the Arg-16 and Gly-16 genotypes for the acute salbutamol response following methacholine bronchial challenge between the first and last doses of formoterol (FM) and salmeterol (SM) combination inhalers. METHODS: Parallel groups of 10 matched homozygous Arg-16 and 10 homozygous Gly-16 patients completed a randomised, double blind, double dummy, crossover study. Following a 1 week washout period, patients received treatment for 2 weeks with either inhaled budesonide (BUD) 200 micro g + FM 6 micro g (two puffs twice daily) or inhaled fluticasone propionate (FP) 250 micro g + SM 50 micro g (one puff twice daily). After washouts and randomised treatments (1 hour after the first and last inhalation) a methacholine challenge was performed followed by salbutamol 200 micro g, with recovery over 30 minutes (the primary outcome). RESULTS: Washout values for forced expiratory volume in 1 second (FEV(1)), methacholine hyperreactivity, and salbutamol recovery were similar for both treatments and genotypes. Pre-challenge FEV(1) values for both genotypes did not differ significantly between the first and last doses of each treatment. Salbutamol recovery as mean (SE) area under the 30 minute time-response curve was significantly delayed (p<0.05) equally in both genotype and treatment groups. There were no differences in salbutamol recovery in either genotype or treatment group. CONCLUSION: Acute salbutamol recovery in methacholine constricted airways was significantly delayed to a similar degree in both genotypes due to cross tolerance induced by FM or SM. PMID- 15282386 TI - Effect of oxygen on recovery from maximal exercise in patients with chronic obstructive pulmonary disease. AB - BACKGROUND: The effects of oxygen on recovery from exercise in patients with chronic obstructive pulmonary disease (COPD) are not clearly known. A study was undertaken to determine whether oxygen given after maximal exercise reduced the degree of dynamic hyperinflation and so reduced the perception of breathlessness. METHODS: Eighteen patients with moderate to severe COPD performed maximal symptom limited exercise on a cycle ergometer. During recovery they received either air or oxygen at identical flow rates in a randomised, single blind, crossover design. Inspiratory capacity, breathing pattern data, dyspnoea intensity, and leg fatigue scores were collected at regular intervals during recovery. At a subsequent visit patients underwent a similar protocol but with a face mask in situ to eliminate the effects of instrumentation. RESULTS: When oxygen was given the time taken for resolution of dynamic hyperinflation was significantly shorter (mean difference between air and oxygen 6.61(1.65) minutes (95% CI 3.13 to 10.09), p = 0.001). Oxygen did not, however, reduce the perception of breathlessness during recovery nor did it affect the time taken to return to baseline dyspnoea scores in either the instrumented or non-instrumented state (mean difference 2.11 (1.41) minutes (95% CI -0.88 to 5.10), p = 0.15). CONCLUSIONS: Oxygen reduces the degree of dynamic hyperinflation during recovery from exercise but does not make patients feel less breathless than breathing air. This suggests that factors other than lung mechanics may be important during recovery from exercise, or it may reflect the cooling effect of both air and oxygen. PMID- 15282387 TI - Peripheral muscle endurance and the oxidative profile of the quadriceps in patients with COPD. AB - BACKGROUND: Based on previously reported changes in muscle metabolism that could increase susceptibility to fatigue, we speculated that patients with chronic obstructive pulmonary disease (COPD) have reduced quadriceps endurance and that this will be correlated with the proportion of type I muscle fibres and with the activity of oxidative enzymes. METHODS: The endurance of the quadriceps was evaluated during an isometric contraction in 29 patients with COPD (mean (SE) age 65 (1) years; forced expiratory volume in 1 second 37 (3)% predicted) and 18 healthy subjects of similar age. The electrical activity of the quadriceps was recorded during muscle contraction as an objective index of fatigue. The time at which the isometric contraction at 60% of maximal voluntary capacity could no longer be sustained was used to define time to fatigue (Tf). Needle biopsies of the quadriceps were performed in 16 subjects in both groups to evaluate possible relationships between Tf and markers of muscle oxidative metabolism (type I fibre proportion and citrate synthase activity). RESULTS: Tf was lower in patients with COPD than in controls (42 (3) v 80 (7) seconds; mean difference 38 seconds (95% CI 25 to 50), p<0.001). Subjects in both groups had evidence of electrical muscle fatigue at the end of the endurance test. In both groups significant correlations were found between Tf and the proportion of type I fibres and citrate synthase activity. CONCLUSION: Isometric endurance of the quadriceps muscle is reduced in patients with COPD and the muscle oxidative profile is significantly correlated with muscle endurance. PMID- 15282388 TI - COPD increases the risk of squamous histological subtype in smokers who develop non-small cell lung carcinoma. AB - BACKGROUND: Squamous cell carcinoma has a stronger association with tobacco smoking than other non-small cell lung cancers (NSCLC). A study was undertaken to determine whether chronic obstructive pulmonary disease (COPD) is a risk factor for the squamous cell carcinoma histological subtype in smokers with surgically resectable NSCLC. METHODS: Using a case-control design, subjects with a surgically confirmed diagnosis of squamous cell carcinoma were enrolled from smokers undergoing lung resection for NSCLC in the District Hospital of Ferrara, Italy. Control subjects were smokers who underwent lung resection for NSCLC in the same hospital and had a surgically confirmed diagnosis of NSCLC of any histological type other than squamous cell. RESULTS: Eighty six cases and 54 controls (mainly adenocarcinoma, n = 50) were enrolled. The presence of COPD was found to increase the risk for the squamous cell histological subtype by more than four times. Conversely, the presence of chronic bronchitis was found to decrease the risk for this histological subtype by more than four times. Among patients with chronic bronchitis (n = 77), those with COPD had a 3.5 times higher risk of having the squamous cell histological subtype. CONCLUSIONS: These data suggest that, among smokers with surgically resectable NSCLC, COPD is a risk factor for the squamous cell histological subtype and chronic bronchitis, particularly when not associated with COPD, is a risk factor for the adenocarcinoma histological subtype. PMID- 15282389 TI - Childhood smoking is an independent risk factor for obstructive airways disease in women. AB - OBJECTIVE: To assess whether starting to smoke in childhood increases the risk of obstructive airways disease (OAD) in adult life. METHODS: A retrospective cohort analysis was undertaken of 12 504 current and ex-smokers in the EPIC-Norfolk cohort. The main exposure was starting to smoke during childhood (age <16 years). Three definitions of OAD were used: doctor diagnosed asthma, doctor diagnosed bronchitis/emphysema, and "any OAD" (doctor diagnosed asthma or bronchitis/emphysema, or taking medication used in the treatment of OAD). RESULTS: Childhood smokers had significantly more pack years of exposure and poorer lung function than subjects who started to smoke in adulthood (>/=16 years). Compared with starting in adulthood, starting to smoke in childhood was associated with a greater risk of bronchitis/emphysema in female smokers (OR 1.79, 95% CI 1.25 to 2.56) and ex-smokers of both sexes (OR 1.29, 95% CI 1.07 to 1.55 in men and OR 1.40, 95% CI 1.05 to 1.85 in women), and of "any OAD" in female smokers (OR 1.72, 95% CI 1.24 to 2.38) and male and female ex-smokers (OR 1.20, 95% CI 1.03 to 1.40 in men and 1.34, 95% CI 1.07 to 1.57 in women). After adjustment for pack years, childhood smoking was associated with poorer lung function (FEV(1) 92.3% predicted in adult smokers and 89.5% in childhood smokers, p = 0.03) and a greater risk of bronchitis/emphysema (adjusted OR 1.55, 95% CI 1.08 to 2.24) and for "any OAD" (OR 1.54, 95% CI 1.10 to 2.13) in female smokers but not in male and female ex-smokers. CONCLUSION: Starting to smoke in childhood is associated with an increased risk of airways disease because of the extra pack years smoked. In women, childhood smoking is itself an independent risk factor for the development of airways disease. PMID- 15282390 TI - Expression of glucocorticoid receptors alpha and beta in steroid sensitive and steroid insensitive interstitial lung diseases. AB - BACKGROUND: Sensitivity to glucocorticoids may be related to the concentration of glucocorticoid receptors alpha (GRalpha) and beta (GRbeta). A study was undertaken to assess GRalpha and GRbeta expression in steroid insensitive interstitial lung disease (idiopathic pulmonary fibrosis (IPF)) and steroid sensitive interstitial lung diseases (sarcoidosis and cryptogenic organising pneumonia (COP)). METHODS: Lung tissue was obtained from control subjects and from patients with IPF, sarcoidosis, and COP. Pulmonary function tests were carried out at the time of lung biopsy and every 3 months. GRalpha and GRbeta expression was evaluated by both competitive RT-PCR and immunohistochemistry. Data are presented as median and 25-75th percentile. RESULTS: GRalpha mRNA expression (10(5) cDNA copies/ micro g total RNA) was higher in patients with steroid sensitive interstitial lung diseases (10.0; 7.8-14.9; n = 11) than in patients with IPF (4.4; 3.2-6.6; n = 19; p<0.001). GRbeta expression was at least 1000 times lower than that of GRalpha and did not differ between the three groups. A negative correlation was found between GRalpha mRNA levels and the fibrotic pathology score of the tissue (r = -0.484, p<0.01) and a positive correlation was found between GRalpha mRNA levels and improvement in forced vital capacity (r = 0.633; p<0.01) after treatment of patients with glucocorticoids. Immunoreactivity for GR protein was also higher in patients with sarcoidosis and COP than in those with IPF. CONCLUSION: The variable response of some interstitial lung diseases to steroid treatment may be the result of differences in the expression of GRalpha. PMID- 15282391 TI - Effects of breathing pattern and inspired air conditions on breath condensate volume, pH, nitrite, and protein concentrations. AB - BACKGROUND: The effects of breathing pattern and inspired air conditions on the volume and content of exhaled breath condensate (EBC) were investigated. METHODS: Total exhaled water (TEW), EBC volume, pH, nitrite and protein concentrations were measured in three groups of 10 healthy subjects breathing into a condenser at different target minute ventilations (Vm), tidal volumes (Vt), and inspired air conditions. RESULTS: The volumes of both TEW and EBC increased significantly with Vm. For Vm 7.5, 15 and 22.5 l/min, mean (SD) EBC was 627 (258) microl, 1019 (313) microl, and 1358 (364) microl, respectively (p<0.001) and TEW was 1879 (378) microl, 2986 (496) microl, and 4679 (700) microl, respectively (p<0.001). TEW was significantly higher than EBC, reflecting a condenser efficiency of 40% at a target Vm of 7.5 l/min which reduced to 29% at Vm 22.5 l/min. Lower Vt gave less TEW than higher Vt (26.6 v 30.7 microl/l, mean difference 4.1 (95% CI 2.6 to 5.6), p<0.001) and a smaller EBC volume (4.3 v 7.6 microl/l, mean difference 3.4 (95% CI 2.3 to 4.5), p<0.001). Cooler and drier inspired air yielded less water vapour and less breath condensate than standard conditions (p<0.05). Changes in the breathing pattern had no effect on EBC protein and nitrite concentrations and pH. CONCLUSION: These results show that condensate volume can be increased by using high Vt and increased Vm without compromising the dilution of the sample. PMID- 15282392 TI - Effect of oral bisphosphonates on bone mineral density and body composition in adult patients with cystic fibrosis: a pilot study. AB - BACKGROUND: Approximately two thirds of adult patients with cystic fibrosis have reduced bone mineral density and up to one quarter have osteoporosis at one or more sites. Any bone mineral deficits are likely to be exacerbated in patients following lung transplantation by their immunosuppressive regimen. Vertebral collapse and rib fractures will impair the ability to cough and the efficacy of physiotherapy treatments. METHODS: Patients attending the Leeds Regional Adult Cystic Fibrosis Unit with either osteopenia or osteoporosis on dual energy x ray absorptiometry (DXA) scanning were offered treatment with oral bisphosphonates after exclusion of abnormal vitamin D, calcium, or phosphate levels, abnormal thyroid function, or hypogonadism. Those declining treatment or patients with a normal initial DXA scan formed the control group. A second DXA scan was performed after a mean of 2.4 years in the treatment group and 2.9 years in the non treatment group. Patients in the active group were asked to complete a short questionnaire detailing their adherence to treatment. RESULTS: The medians of the differences in annual changes in bone parameters between treatment and control groups showed significant differences in bone mineralisation in favour of the treatment group at the lumbar spine (L2-L4), the femoral neck, and for total body measurements. There were no significant differences in weight, height, or body composition in either patient group. Most treated patients stated that they adhered to treatment most of the time. CONCLUSION: Treatment with oral bisphosphonates may improve bone mineralisation in adult patients with cystic fibrosis. The results of this pilot study need to be further explored in a randomised controlled trial. PMID- 15282393 TI - Paradoxical reactions during tuberculosis treatment in patients with and without HIV co-infection. AB - BACKGROUND: It has been suggested that deterioration of tuberculosis (TB) during appropriate treatment, termed a paradoxical reaction (PR), is more common and severe in HIV positive individuals on highly active antiretroviral therapy (HAART). METHOD: A study was undertaken to determine the frequency of PR and its associated features in a population of HIV+TB+ patients and a similar sized group of HIV-TB+ individuals. RESULTS: PR occurred in 28% of 50 HIV+TB+ patients and 10% of 50 HIV-TB+ patients. Disseminated TB was present in eight of 13 HIV+TB+ patients and four of five HIV-TB+ patients with PR. In 28 HIV+TB+ patients starting HAART, PR was significantly associated with commencing HAART within 6 weeks of starting antituberculosis treatment (p = 0.03) and was more common in those with disseminated TB (p = 0.09). No association was found between development of PR and baseline CD4 count or CD4 response to HAART. CONCLUSIONS: PR is common in HIV infected and uninfected individuals with TB. Early introduction of HAART and the presence of disseminated TB appear to be important in co-infected patients. PMID- 15282394 TI - alpha1-Antitrypsin deficiency . 5: intravenous augmentation therapy: current understanding. AB - The biochemical and clinical efficacy of intravenous augmentation therapy in alpha(1)-antitrypsin deficiency is reviewed, adverse events experienced with this treatment are considered, and its cost effectiveness is discussed. PMID- 15282396 TI - Music: a new cause of primary spontaneous pneumothorax. AB - Most cases of primary spontaneous pneumothorax are thought to be caused by air leaks at so-called "emphysema-like changes" or in areas of pleural porosity at the surface of the lung. Environmental pressure swings may cause air leaks as a result of transpulmonary pressure changes across areas of trapped gas in the distal lung. This is the first report of music as a specific form of air pressure change causing pneumothorax (five episodes in four patients). While rupture of the interface between the alveolar space and pleural cavity in these patients may be linked to the mechanical effects of acute transpulmonary pressure differences caused by exposure to sound energy in association with some form of distal air trapping, we speculate that repetitive pressure changes in the high energy-low frequency range of the sound exposures is more likely to be responsible. Exposure to loud music should be included as a precipitating factor in the history of patients with spontaneous pneumothorax. PMID- 15282397 TI - Consumption of milk fat and reduced asthma risk in pre-school children. PMID- 15282395 TI - Acute effects of cigarette smoke on inflammation and oxidative stress: a review. AB - Compared with the effects of chronic smoke exposure on lung function and airway inflammation, there are few data on the acute effects of smoking. A review of the literature identified 123 studies investigating the acute effects of cigarette smoking on inflammation and oxidative stress in human, animal, and in vitro models. An acute smoking model is a relatively easy and sensitive method of investigating the specific effects of cigarette smoke on oxidative stress and inflammation. Acute smoke exposure can result in tissue damage, as suggested by increased products of lipid peroxidation and degradation products of extracellular matrix proteins. Acute cigarette smoke has a suppressive effect on the number of eosinophils and several inflammatory cytokines, possibly due to the anti-inflammatory effect of carbon monoxide. An acute smoking model can supplement other ways of studying the effects of smoking and is an as yet underinvestigated method for intervention studies in smoking related diseases. PMID- 15282398 TI - BIS/BTS SARS guidelines. PMID- 15282399 TI - Urinary leukotriene LTE4 levels in non-responders to antileukotriene therapy. PMID- 15282400 TI - Rhomboideus major muscle metastasis as an initial clinical manifestation of pulmonary adenocarcinoma. PMID- 15282401 TI - Hepatic gene expression and lipid homeostasis in C57BL/6 mice exposed to hydrazine or acetylhydrazine. AB - Hydrazine (HD) and acetylhydrazine (AcHD) are metabolites of the antituberculosis drug isoniazid (INH) that have been implicated in INH-induced liver damage. The hepatotoxicity of AcHD and HD were compared in adult male C57Bl/6J mice by evaluating hepatic histopathology, plasma biochemistry, and hepatic gene expression. By all measures, HD had significantly greater effects than AcHD. There was no evidence of liver damage following exposure to AcHD (300 mg/kg, po). However, HD at this dose caused marked hepatic necrosis, macrovesicular degeneration, and steatosis. Lipid accumulation was initiated 2 h after HD exposure, with hepatic macrovesicular degeneration evident after 4 h, and severe necrosis by 36 h. Gene expression profiles were compared 24 h following 100 mg/kg po of HD or AcHD. HD changed the hepatic expression of more genes than AcHD, particularly lipid synthesis, transport, and metabolism genes that may be involved in steatosis. Hepatic expression of genes regulated by peroxisome proliferator activated receptors (PPAR) and sterol regulatory element binding protein (SREBP) transcription factors was increased only by HD. The hepatotoxicty and hepatic gene expression profile of HD, but not AcHD, indicate that exposure to HD initiates a process whereby the production and intracellular transport of hepatic lipids is favored over the removal of fatty acids and their metabolites. PMID- 15282402 TI - Oral and dermal exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) induces cutaneous papillomas and squamous cell carcinomas in female hemizygous Tg.AC transgenic mice. AB - Tg.AC mice develop epidermal papillomas in response to treatment with dermally applied nongenotoxic and complete carcinogens. The persistent environmental contaminant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is a multi-site rodent carcinogen and tumor promoter that induces the formation of papillomas in Tg.AC mice. To examine the dose-response relationship and compare dermal and oral routes of exposure for TCDD-induced skin papillomas, female Tg.AC mice were exposed dermally to average daily doses of 0, 2.1, 7.3, 15, 33, 52, 71, 152, and 326 ng TCDD/kg/day or 0, 75, 321, and 893 ng TCDD/kg body weight by gavage for 26 weeks. The incidence of cutaneous papillomas was increased in a dose-dependent manner, and tumors developed earlier with higher exposure to TCDD regardless of route of administration. Increased incidences of cutaneous squamous cell carcinomas were observed in mice exposed to dermal (> or =52 ng/kg) and oral (893 ng/kg) TCDD. Higher gavage doses than dermal exposure doses were required to induce papillomas and squamous cell carcinomas. Despite a linear correlation between administered dose and terminal skin concentrations, the incidence of tumor formation was lower in the gavage study than in the dermal study with respect to mean terminal skin TCDD concentrations. These studies demonstrate that, although Tg.AC mice are less responsive to TCDD by gavage than by dermal exposure, the induction of skin neoplasms is a response to systemic exposure and not solely a local response at the site of dermal application. Differences in response between the routes of exposure may reflect pharmacokinetic differences in the delivery of TCDD to the skin over the duration of the study. PMID- 15282403 TI - Time course of cII gene mutant manifestation in the liver, spleen, and bone marrow of N-ethyl-N-nitrosourea-treated Big Blue transgenic mice. AB - The time between treatment and the appearance of mutants (mutant manifestation time) is a critical variable for in vivo transgenic mutation assays. There are, however, limited data describing the optimal sampling time for detecting mutations in various tissues of mutagen-treated animals. In this study, we investigated the time course of cII gene mutant induction in the liver, spleen, and bone marrow of Big Blue transgenic mice treated with N-ethyl-N-nitrosourea (ENU). Six-month-old female mice were treated with a single dose (120 mg/kg) of ENU, and the animals were sacrificed, and the cII mutant frequencies (MFs) were determined at 1, 3, 7, 15, 30, and 120 days after the treatment. The MFs in the liver cII gene of ENU-treated mice increased with time after the treatment, while the MFs for concurrent controls remained constant. The liver cII MFs in ENU treated mice were significantly increased at day 30 and 120 (p < 0.01), with the largest increase at day 120. The spleen cII MFs in ENU-treated mice were increased significantly at day 7 and later (p < 0.01), and reached a plateau at day 30. In the bone marrow, the cII MFs in ENU-treated mice were increased significantly at all sampling times (p < 0.01), with the maximum MF at day 3. These results confirm that the time after treatment required to reach the maximum MF is tissue specific, with the approximate time for the maximum ENU-induced cII MF response being: bone marrow, 3 days; spleen, 14-30 days; and liver, more than 30 days. PMID- 15282404 TI - ARNT2 is not required for TCDD developmental toxicity in zebrafish. AB - ZfAHR2 has been identified as the receptor that is essential for mediating the developmental toxicity caused by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in zebrafish. One form of zfARNT2, zfARNT2b, forms a functional heterodimer with zfAHR2 that specifically recognizes XREs in gel shift experiments and induces XRE driven transcription in COS-7 cells treated with TCDD. However, it has not been demonstrated that zfARNT2b acts as the physiological dimerization partner for zfAHR2 to mediate TCDD toxicity in developing zebrafish. An antisense morpholino targeted against zfARNT2 (zfarnt2-MO) along with a line of mutant zebrafish lacking expression of the zfarnt2 gene have been used to test the hypothesis that zfARNT2 mediates the developmental toxicity of TCDD. Injection of the zfarnt2-MO decreased expression of the zfARNT2 protein but did not provide any protection against the formation of pericardial edema at 72 hpf. In addition, in TCDD dose response studies the zfarnt2(-/-) embryos showed no protection against three endpoints of TCDD toxicity observed at 96 hpf: pericardial edema, reduced trunk blood flow, and shortened lower jaw. Finally, immunostaining results at 96 hpf demonstrate that the zfarnt2(-/-) embryos show a similar pattern of TCDD-induced zfCYP1A expression as WT embryos. These results demonstrate that zfARNT2 is not essential for mediating TCDD developmental toxicity in zebrafish and suggest that alternate dimerization partner(s) exist for zfAHR2 in vivo. PMID- 15282405 TI - Nuclear factor kappaB activity determines the sensitivity of kidney epithelial cells to apoptosis: implications for mercury-induced renal failure. AB - Nuclear factor kappa B (NF-kappaB) is a thiol-dependent transcriptional factor that promotes cell survival and protects cells from apoptotic stimuli. Numerous studies have demonstrated increased sensitivity to apoptosis associated with inhibition of NF-kappaB activation in various cell types. We have previously demonstrated that mercuric ion (Hg(2+)), one of the strongest thiol-binding agents known, impairs NF-kappaB activation and DNA binding at low microM concentrations in kidney epithelial cells. In the present studies we investigated the hypothesis that inhibition of NF-kappaB activation by Hg(2+) and other selective NF-kappaB inhibitors would increase the sensitivity of kidney epithelial (NRK52E) cells to apoptogenic agents to which these cells are normally resistant. Fewer than 10% of untreated cells in culture were found to be apoptotic when evaluated by DNA fragmentation (TUNEL) assay. Treatment of cells with Hg(2+) in concentrations up to 5 microM or with tumor necrosis factor-alpha (TNF) (300 units/ml) did not significantly increase the proportion of apoptotic cells, compared with untreated controls. However, when TNF was given following Hg(2+) pretreatment (0.5 to 5 microM for 30 min), the proportion of cells undergoing apoptosis increased by 2- to 6-fold over that seen in untreated controls. Kidney cells pretreated with specific NF-kappaB inhibitors (Bay11-7082 or SN50) prior to TNF also showed a significant increase in apoptosis. Increased sensitivity to apoptotic cell death following these treatments was significantly attenuated in cells transfected with a p65 expression vector. In studies in vivo, rats pretreated by intraperitoneal injection with Hg(2+) (0.75 mg/kg) 18 h prior to administration of bacterial lipopolysaccharide (LPS) (10 mg/kg) displayed impaired NF-kappaB activation and an increased mitochondrial cytochrome c release in kidney cortical cells. These findings are consistent with the view that prevention of NF-kappaB activity in vitro or in vivo enhances the sensitivity of kidney cells to apoptotic stimuli to which these cells are otherwise resistant. Since apoptosis is known to play a seminal role in the pathogenesis of renal failure caused by toxicant injury to tubular cells, the present findings suggest that inhibition of NF-kappaB activity may define a molecular mechanism underlying the pathogenesis of Hg(2+) toxicity in kidney cells. PMID- 15282406 TI - Roles for epoxidation and detoxification of coumarin in determining species differences in clara cell toxicity. AB - Coumarin-induced mouse Clara cell toxicity is thought to result from the local formation of coumarin 3,4-epoxide (CE). However, this toxicity is not observed in the rat, indicating species differences in coumarin metabolism. The purpose of the present work was to characterize the in vitro kinetics of coumarin metabolism in mouse, rat, and human whole lung microsomes, and to determine whether species differences in coumarin-induced Clara cell toxicity correlate with coumarin epoxidation or detoxification. In B6C3F1 mouse lung microsomes, coumarin was metabolized to CE, which in the absence of glutathione spontaneously rearranges to o-hydroxyphenylacetaldehyde (o-HPA). The K(m) and V(max) for o-HPA formation were 155 microM and 7.3 nmol/min/mg protein, respectively. In contrast, the K(m) and V(max) were 2573 microM and 1.75 nmol/min/mg protein, respectively, in F344 rat lung microsomes. Since the intrinsic clearance through the epoxidation pathway was 69 times higher in the mouse, the epoxidation rate was shown to correlate with species sensitivity to toxicity. To determine whether detoxification reactions contribute to species differences in toxicity, the fate of CE and o-HPA were examined. Detoxification of CE via conjugation with glutathione was evaluated in lung cytosol from mice and rats, and the K(m) of this reaction was approximately 800 microM in both species, whereas the V(max) was 3.5 and 6 nmol/min/mg protein, respectively, indicating that conjugation is faster in the rat. Oxidation of o-HPA to o-hydroxyphenylacetic acid (o-HPAA) was examined in lung cytosol from mice and rats. The K(m) of this reaction was approximately 1.5 microM in both species, whereas the V(max) was 0.08 and 0.33 nmol/min/mg protein in mice and rats, respectively, indicating that oxidation is faster in the rat. While the rate of epoxidation correlates with species sensitivity to coumarin, it is likely that Clara cell toxicity is modulated by CE and o-HPA detoxification. In contrast to rodent lung microsomes, bioactivation of coumarin to o-HPA did not occur in 16 different human lung microsomes, which suggests metabolism-dependent toxicity in the human lung is unlikely following low level coumarin exposure. PMID- 15282407 TI - Xenoestrogenicity in in vitro assays is not caused by displacement of endogenous estradiol from serum proteins. AB - The possibility that compounds tested for estrogenicity can compete for binding places on serum proteins and cause an increase of available and very potent endogenous estrogens is of great interest for both the in vitro assay results and the prediction of risk for humans. In in vitro assays, small amounts of estradiol remaining after the charcoal stripping of serum applied in the culture medium could be displaced by the tested compounds, leading to an estrogenic response that might be falsely attributed to the test compound. We have studied the stripping efficiency of charcoal and measured whether reported xenoestrogens can displace estradiol from serum in an in vitro assay using negligible depletion solid phase microextraction (nd-SPME). Possible competition was also studied with a mathematical exposure model, from which the predictions were compared to the measurements. We found that the common charcoal stripping procedure removed 99% of initially present estradiol. Additionally, our results with charcoal adsorption indicate that charcoal is not useful for serum protein binding assays, as it adsorbs more than the free fraction of ligand. Although the competition model predicted a displacement of estradiol from the serum proteins at the higher applied doses of xenoestrogen, the measurements showed no displacement. Therefore, we conclude that estrogenic responses in the in vitro assay applied here are not caused by displacement of remaining estradiol in the stripped serum. The possibility remains, however, that our displacement hypothesis does apply for estrogen sulfates, as these are present in much higher concentrations than estradiol in stripped serum. PMID- 15282408 TI - Accumulation of PBDE-47 in primary cultures of rat neocortical cells. AB - A number of recent studies have examined the neurotoxic actions of polybrominated diphenyl ethers (PBDEs) using in vitro cell culture models. However, there are few data reporting the final concentration of PBDEs in cells after in vitro exposure to these compounds. To address this issue, the present study examined the concentration-dependent and time-dependent accumulation of 2,2',4,4' tetrabromodiphenyl ether (PBDE-47) in primary cultures of rat neocortex. Mixed cultures of neuronal and glial cells were prepared from the neocortex of newborn rats and grown for 7 days in vitro. The cells were then exposed to freshly prepared serum-free culture medium containing (14)C-PBDE-47. Radiolabel associated with the cells or remaining in the medium was determined by liquid scintillation spectrometry. Exposure to 0.01-3.0 microM PBDE-47 for 60 min resulted in a concentration-dependent accumulation in cells. At each concentration, approximately 15% of the applied PBDE-47 was associated with the cells, resulting in a 100-fold magnification of the applied concentration (e.g., a 60-min exposure to 1 microM resulted in an approximate 100 microM concentration in the cells); 55% of the PBDE remained in the medium and 30% was associated with the plastic culture dish. Exposure to 1 microM PBDE-47 resulted in a linear increase in PBDE-47 in cells with time for the first 60 min, which began to saturate at 120 min. Addition of serum proteins to the medium decreased accumulation; at 10% serum in the medium, only 3% of the applied PBDE-47 was associated with the cells and 96% remained in the media after 60 min. The total volume of exposure also influenced accumulation of PBDE-47. Doubling the volume of serum-free exposure medium (from 2 ml to 4 ml) but leaving the concentration constant (1 microM) resulted in a 1.5-fold increase in PBDE-47 concentration in the cells. These data show that a number of factors, including duration of exposure, volume of exposure, and concentration of serum proteins in the medium, can influence the accumulation of PBDE-47 in cells in vitro. For this highly lipophilic compound, use of medium concentration underestimates tissue concentration by up to two orders of magnitude. Thus, accurate information on the tissue concentration for in vitro experiments should be determined empirically. PMID- 15282409 TI - No-observed effect levels for carcinogenicity and for in vivo mutagenicity of a genotoxic carcinogen. AB - To elucidate the relationship between in vivo carcinogenic and mutagenic potentials of genotoxic carcinogens, low doses were tested in the livers of Big Blue transgenic rats with 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx). Male Big Blue rats were fed a diet containing 0.001, 0.01, 0.1, 1, 10, or 100 ppm of MeIQx for 16 weeks, and the frequencies of lacI mutants and glutathione S transferase placental form (GST-P) positive foci in the liver were determined. The mutation frequencies significantly increased at doses of 10 and 100 ppm, and GST-P positive foci significantly increased at a dose of 100 ppm. However, no statistical increases in both frequencies were observed at lower doses. MeIQx most frequently induced G frameshifts, followed by G to T transversions. Thus, no observed effect level (NOEL) was demonstrated for both carcinogenicity in terms of preneoplastic lesion induction and in vivo mutagenicity of MeIQx, and the NOEL for in vivo mutagenicity was lower than that for carcinogenicity. PMID- 15282410 TI - Compartmentation of Nrf-2 redox control: regulation of cytoplasmic activation by glutathione and DNA binding by thioredoxin-1. AB - Nrf-2 is a redox-sensitive transcription factor that is activated by an oxidative signal in the cytoplasm but has a critical cysteine that must be reduced to bind to DNA in the nucleus. The glutathione (GSH) and thioredoxin (TRX) systems have overlapping functions in thiol/disulfide redox control in both the cytoplasm and the nucleus, and it is unclear whether these are redundant or have unique functions in control of Nrf-2-dependent signaling. To test whether GSH and Trx-1 have distinct functions in Nrf-2 signaling, we selectively modified GSH by metabolic manipulation and selectively modified Trx-1 expression by transient transfection. Cytoplasmic activation of Nrf-2 was measured by its nuclear translocation and nuclear activity of Nrf-2 was measured by expression of a luciferase reporter construct containing an ARE4 from glutamate cysteine ligase. Results showed that tert-butylhydroquinone (TBHQ), a transcriptional activator that functions through Nrf-2/ARE, promoted Nrf-2 nuclear translocation by a type I (thiylation) redox switch which was regulated by GSH not by Trx-1. In contrast, the ARE reporter was principally controlled by nuclear-targeted Trx-1 and not by GSH. The data show that the GSH and TRX systems have unique, compartmented functions in the control of transcriptional regulation by Nrf-2/ARE. PMID- 15282411 TI - Should we treat patients with moderately severe stenosis of the left main coronary artery and negative FFR results? AB - BACKGROUND: The prognostic importance of significant left main coronary artery disease is unequivocal. However, moderate lesions of the left main coronary artery (LMCA) are sometimes found in patients presenting significant stenosis in other coronary arteries or equivocal symptoms. The ability of myocardial fractional flow reserve (FFR) to predict coronary events could be useful in the decision-making process in these patients. The present study was designed to investigate the occurrence of cardiac events in patients with coronary syndromes and LMCA stenosis of moderate severity in whom FFR failed to show an haemodynamic significant repercussion of the LMCA. METHODS AND RESULTS: We studied 27 consecutive patients (mean age 62.7 +/- 10.5 years) with moderate stenoses (30 50%) of the LMCA. In seven patients who presented significantly reduced FFR (< 0.75) at the LMCA level (Group A), coronary revascularization of this vessel was performed. In 20 patients with negative FFR (greater than or equal to 0.75) at the LMCA level (Group B), the LMCA stenosis was not revascularized, being the revascularization procedures (if any) limited to other arteries with significant obstructions. During a mean follow-up period of 26.2 +/- 12.1 months, clinical events occurred in 3 patients in the whole group. One patient with positive FFR died during coronary bypass surgery. Two group B patients were surgically revascularized 4 months and 4 years after the initial coronariography. CONCLUSIONS: Patients with coronary lesions of moderate severity on the LMCA and negative FFR may constitute a subgroup of good prognosis in the follow-up. Our findings suggest that negative FFR is a potentially useful indicator of the likelihood of cardiac events, and thus represents a useful aid in clinical decision-making in the hemodynamics laboratory. PMID- 15282412 TI - Plaque that makes a patient vulnerable. PMID- 15282413 TI - Clinical and angiographic safety and efficacy trial with a new coronary stent: the RESTOR study of the R Stent. AB - The RESTOR trial (R Stent Efficacy and Safety Trial by ORBUS) is an efficacy and safety evaluation of the R Stent for treatment of patients with a single de novo coronary lesion < 25 mm in length in a coronary artery of 2.75-4.0 mm diameter. This new stent utilizes a patented dual helix design for radial strength and flexibility. The aim of the study was to assess major adverse cardiac events (MACE) and angiographic outcome at 6 months after implantation. From May to December 2000 a total of 121 patients with symptomatic stable or unstable angina pectoris or documented silent ischemia and a significant single, de novo coronary lesion (average reference vessel diameter 2.84 +/- 0.54 mm, average lesion length 10.53 +/- 3.70 mm) were included in two Dutch centers. All patients were treated with clopidogrel 75 mg/day for 1 month and with aspirin greater than or equal to 100 mg/day. The angiographic success rate (< 30% diameter stenosis post procedure) was 98.3%. Procedural success (angiographic success without in hospital MACE) was 95.9%. The 6-month MACE rate was 12.4%. 101 of the 121 patients had an angiographic follow-up at 6 months. Minimal lumen diameter pre /post-procedure and at follow-up was 0.98 +/- 0.37, 2.64 +/- 0.38 and 1.85 +/- 0.68 mm, respectively. The resulting binary restenosis rate in this population was 20.8%. The coronary R Stent is safe and effective as a primary device for the treatment of native coronary lesions in patients with stable or unstable angina pectoris, and well suitable as a platform for a drug eluting stent. PMID- 15282414 TI - Early and late clinical outcomes after rotational atherectomy with stenting versus rotational atherectomy with balloon angioplasty for complex coronary lesions. AB - Limited data are available on the effect of rotational atherectomy plus stenting versus rotational atherectomy plus balloon angioplasty for complex coronary lesions. We compared the early and late clinical outcomes between rotational atherectomy plus stenting (158 patients, 171 lesions) and rotational atherectomy plus balloon angioplasty (165 patients, 186 lesions) for complex lesions. Baseline characteristics were similar between the two groups. The procedural success rate was similar between the 2 groups (94% in rotational atherectomy plus stenting versus 96% in rotational atherectomy plus balloon angioplasty; p = 0.54). There were no significant differences in the in-hospital complications between the 2 groups. During mean follow-up of 40.4 +/- 20.2 months, fourteen patients died: 6 in rotational atherectomy plus stenting and 8 in rotational atherectomy plus balloon angioplasty. Target lesion revascularization was similar between the 2 groups (20% in rotational atherectomy plus stenting versus 24% in rotational atherectomy plus balloon angioplasty; p = 0.46). Three-year event (death, nonfatal myocardial infarction and target lesion revascularization)-free survival rate was 79 +/- 4% in the rotational atherectomy plus stenting group and 75 +/- 3% in the rotational atherectomy plus balloon angioplasty group (p = 0.44). In conclusion, rotational atherectomy followed by stenting or balloon angioplasty is associated with favorable long-term outcomes. Compared with rotational atherectomy plus balloon angioplasty, routine stenting after rotational atherectomy does not provide additional benefits in the clinical outcomes in complex coronary lesions. PMID- 15282415 TI - Rotastenting for "complex lesions": benefit unlikely with bare metal stents. PMID- 15282416 TI - The current status of stent placement in small coronary arteries < 3.0 mm in diameter. AB - Data accrued to date indicate that stent placement in small vessels (< 3.0 mm reference diameter) suffers from the same disadvantage as other non-stent interventional devices in that the restenosis rate is significantly higher than observed following intervention involving large vessels. Randomized trials comparing systematic bare metal stenting versus conventional balloon angioplasty in the setting of small coronary arteries, however, show that the former therapeutic modality is probably superior to the latter treatment in its acute and mid-term angiographic and clinical results. Balloon angioplasty, even if performed optimally with resultant stent-like luminal outcome, yields a restenosis rate that is at best equivalent to that observed with stent placement. Stent performance is influenced profoundly by stent design and configuration. Tubular and corrugated stents are better than coil or meshwire stent design. Stents with thin struts appear to yield a lower restenosis rate compared with thick-strut stents. Coating the surface of stents with gold, phosphorylcholine or heparin does not appear to confer any additional long-term benefit compared with bare stainless-steel stents. On the other hand, impregnation of stents with anti proliferative drugs, with or without a carrier polymer, has produced a significantly lower risk of restenosis, without an increase in stent thrombosis rate, compared with uncoated metal stents in multiple randomized trials. However, whether the clinico-anatomic benefits of drug-eluting stents can be sustained for several years and whether there are any long-term deleterious effects from the antiproliferative drug or carrier polymer remains unclear at this stage. PMID- 15282417 TI - Contemporary treatment of small-vessel disease. PMID- 15282418 TI - Hybrid revascularization using percutaneous coronary intervention and robotically assisted minimally invasive direct coronary artery bypass surgery. AB - Hybrid revascularization (HR) combines staged percutaneous coronary intervention (PCI) on stenoses in the non-left anterior descending (LAD) territories with minimally invasive direct coronary artery bypass (MIDCAB) using the left internal thoracic artery (LITA) to the LAD. The LITA-to-LAD graft, which has a 5-year patency rate of 95%, is the major determinant of the long-term survival for patients. Thus, HR aims to perform full revascularization without compromising the survival advantage of the LITA-to-LAD graft, while preserving the minimally invasive advantages associated with the percutaneous treatment of symptomatic coronary stenoses. We investigated whether HR was a valid alternative to conventional coronary artery bypass graft surgery in patients with multivessel coronary artery disease. We also present our early experiences with HR using a combined approach of advanced PCI and robotically-assisted MIDCAB. PMID- 15282419 TI - Hybrid revascularization: another step forward in coronary revascularization. PMID- 15282420 TI - Current applications for nicardipine in invasive and interventional cardiology. AB - Interventional coronary procedures, such as rotational atherectomy and coronary artery bypass graft (CABG) stenting, are associated with a risk of postoperative vasoconstriction, which can lead to sequelae that produce morbidity and even death. Vasodilators, such as calcium channel blockers, sodium nitroprusside and adenosine, are often administered to prevent or reverse these sequelae, and have proven effective for this purpose. The injectable dihydropyridine calcium channel blocker, nicardipine, has several unique properties that make it an effective option for preventing and/or reversing microvascular or conductance vessel vasoconstriction. In this review, we describe the evidence-based uses of nicardipine injection in interventional cardiology and cardiac catheterization procedures. In comparison to other calcium channel blockers, nicardipine injection appears to be potentially safer, easier to administer and capable of producing a more predictable response. This drug has potential advantages in preventing or reversing the no-reflow phenomenon that sometimes occurs after interventions in coronary bypass grafts. Nicardipine may also be effective when administered in a flush solution with other drugs during rotational atherectomy and clot debulking with or without distal protection devices. It also can control the hypertension that occurs after sternotomy and cardiac surgery, and in the cardiac catheterization laboratory. In summary, the dihydropyridine calcium channel blocker, nicardipine, has a wide range of useful applications during cardiovascular interventions. PMID- 15282421 TI - Successful exclusion of a left main coronary artery aneurysm with a PTFE-covered coronary stent. PMID- 15282422 TI - The bifurcation stent. PMID- 15282423 TI - An overview of antithrombins in peripheral vascular interventions. PMID- 15282424 TI - Novel implication of combined stent crushing and intravascular ultrasound for dislodged stents. PMID- 15282425 TI - Comparison of 5 French versus 6 French guiding catheters for transradial coronary intervention: a prospective, randomized study. AB - We compared 5 versus 6 French (Fr) guiding catheters in coronary intervention using the transradial approach. Smaller guiding catheters may have advantages over larger ones in transradial coronary intervention. However, there is uncertainty about how small is small enough, and when smaller would become too small. Eligible patients were randomized between the 5 and 6 Fr groups before the procedure. The primary endpoint was procedural success. A total of 216 patients were randomized. Procedural success was obtained in 95% of the 6 Fr group versus 90% of the 5 Fr group (p = 0.25). Most of the failures in the 5 Fr group were because of cross-over to the 6 Fr group. Crossover to the 5 Fr group occurred in 1 patient in the 6 Fr group (0.9%; p = 0.05) because of a small radial artery. Transradial intervention using 5 Fr guiding catheters necessitates crossover to a 6 Fr catheter in 6.8% of cases, and offers no clear advantages over the 6 Fr technique. PMID- 15282426 TI - Comparison of safety and efficacy between first and second generation of angio seal closure devices in interventional patients. AB - Arterial closure devices are safe and effective in selected patients, with complication rates similar to or lower than manual compression. The purpose of this study was to compare the safety and efficacy of the first- and new generation Angio-Seal devices in patients undergoing PCI. This study found that the new Angio-Seal STS Platform device can secure hemostasis after PCI in a safe and effective manner similar to the old device. The new platform is easier for the operator and for the patients. PMID- 15282427 TI - To hold or to fold - what's your best bet. PMID- 15282428 TI - Traditional versus automated injection contrast system in diagnostic and percutaneous coronary interventional procedures: comparison of the contrast volume delivered. AB - Contrast injection with a manual stopcock-manifold system is the standard technique during diagnostic coronary angiography or percutaneous coronary intervention (PCI). The ACIST Injection System is a new automatic injection device that allows online hemodynamic monitoring, as well as control of injection rate and amount of contrast to be delivered. The aim of the study was to compare the amount of contrast media delivered using the two methods, i.e., the traditional (manual injection plus standard power injection for the left ventriculography using the ANGIOMAT-6000) versus the ACIST System programmed injector. A total of 453 consecutive patients underwent diagnostic cardiac catheterization and/or PCI at our institution. Patients were randomly assigned to either automated contrast injection with the ACIST device (n = 253) or to conventional contrast injection using a stopcock-manifold system and contrast injection by hand syringe (n = 200). In the diagnostic catheterization group, the mean quantity of contrast volume was significantly lower in the ACIST group compared to the control group (130 +/- 60 ml versus 257 +/- 64 ml, respectively; 97.4% more contrast media; p < 0.001). When the data were analyzed for patients who underwent diagnostic catheterization plus PCI, the mean quantity of contrast volume was 228 +/- 90 ml versus 350 +/- 94 ml, respectively (53.8% more contrast media; p < 0.001). For patients who underwent PCI alone, the mean quantity of contrast volume was 175 +/- 76 ml versus 275 +/- 100 ml, respectively (57.3% more contrast media; p = 0.009). When only the total volume of contrast media delivered to the patient was considered (not including the contrast wasted outside), the results were very similar. CONCLUSION: There was a significant reduction in the total volume of contrast media used (amount injected to the patient as well as the amount wasted) and in the net amount of contrast delivered to the patient with the ACIST power device when compared to the traditional method of manual contrast injection. PMID- 15282429 TI - Results of intracoronary beta-brachytherapy administered by 60 mm transfer device/radiation source train: a subgroup analysis from the RENO registry. AB - To investigate the safety and efficacy of a 60 mm transfer device, delivering 60 mm radiation source train, in the treatment of coronary lesions by b brachytherapy employing the BetaCath system (Novoste, Norcross, Georgia). METHODS AND RESULTS: As part of the REgistry NOvoste (RENO), the first large-scale registry of intracoronary beta-radiation applied in routine clinical practice, 46 centers registered 1,098 consecutive patients undergoing brachytherapy with the BetaCath system. Of these, 49 patients with 56 lesions were treated with a 60 mm transfer device/radiation source train (TD/RST) in at least 1 vessel, constituting the study population. With 75.4% in-stent restenosis (ISR), 3.6% graft lesions, long lesions (30.9 +/- 14.7 mm) and 19% diabetes, the cohort had a high-risk for recurrence. The in-hospital major adverse cardiac event (MACE) rate was 4.1%. The 6-month follow-up revealed 2.0% death, 4.1% myocardial infarction, 8.2% target vessel revascularization, 12.2% MACE, 82.6% improved angina, 16.7% binary restenosis and 4.1% late thrombosis. The results were comparable to all other patients in the registry treated with standard source lengths of 30 mm and 40 mm, although much longer lesions were treated by the 60 mm device (18.4 +/- 11.3 mm versus 30.9 +/- 14.7 mm; p < 0.0001). In the ISR subgroup (mean lesion length, 32.03 +/- 14.99 mm), the 6-month MACE rate was 12.8%, while the angiographic restenosis rate was 16.0% and the late thrombosis rate was 2.6%. CONCLUSION: Beta-brachytherapy with 60 mm TD/RST was safe, feasible and effective in this broad population of high-risk patients presenting in day-to-day practice. Its efficacy in long-segment ISR, where conventional interventional strategies have poor outcome rates, is particularly noteworthy. PMID- 15282430 TI - Survival following renal artery stent revascularization: four-year follow-up. AB - To evaluate survival following renal artery stenting, 72 consecutive patients (Mean age 69 +/-10 years, 72% African-American) were retrospectively evaluated over a 4-year period. Complete follow-up was available in 97% patients (70/72). Twelve patients (12/70 = 17%) died; 2 procedure-related (renal stent = 1, coronary stent = 1), one cancer-related and 9 cardiac. The overall survival at a mean follow-up of 31 +/- 13 months was 83%. Eight patients (8/70 = 11%) were eligible for 1 year, 25/70 (36%) for 2 year, 16/70 (23%) for 3 year, 21/70 (30%) for 4 years (or longer) follow up. Five patients died within 1 year, three during the second year, three during the third year and 1 patient after the forth year. Abnormal baseline serum creatinine, male gender, bilateral renal artery stenosis and systolic dysfunction were associated with statistically significant lower survival. PMID- 15282431 TI - Renal artery stenting: continuing to evaluate the benefits. PMID- 15282432 TI - Management of a patient with severe coronary artery disease and cardiogenic shock after prolonged cardiopulmonary resuscitation. PMID- 15282433 TI - Inflammation and atherosclerosis in acute coronary syndromes. AB - We review the role of inflammation in coronary artery disease, particularly its conversion from a chronic to an acute illness. An overview is provided of the various biochemical reactions that are grouped under the heading of inflammation and which lead to the development and progression of atherosclerotic vascular disease and its clinical consequences, especially acute coronary syndromes. The potential role of inflammatory markers in identifying patients at risk and for primary and secondary prevention of events is explored. The impact of current pharmacologic therapies on inflammation and possible future medicines are also discussed. PMID- 15282434 TI - Utility of intravascular ultrasound in the diagnosis of ambiguous calcific left main stenoses. AB - Advances in coronary angiographic imaging have been recently realized with the advent of so-called flat panel technology. However, some of the limitations of coronary angiography, including the so-called pseudo-thrombus appearance of focal highly calcific coronary artery lesions remain. This case highlights the utility of intravascular ultrasound in determining the pathobiology of coronary artery lesions in select cases. PMID- 15282435 TI - Percutaneous transcatheter closure of coronary-pulmonary artery fistula using polytetrafluoroethylene-covered graft stents. PMID- 15282436 TI - The rapid diagnosis of pseudoaneurysm formation in left ventricular free wall rupture. AB - Left ventricular rupture with subsequent pseudoaneurysm formation is an uncommon but potentially catastrophic complication of acute myocardial infarction. We describe a patient with suspected myocardial rupture in whom the diagnosis was rapidly established with the novel use of contrast echocardiography in an emergency room setting. Contrast echocardiography is compared to other modalities in diagnosing this rare, potentially fatal, condition. PMID- 15282437 TI - Isolated right ventricular infarction: percutaneous coronary intervention in three different types of clinical presentation. PMID- 15282438 TI - Pharmacodynamics and bactericidal activity of gatifloxacin in experimental pneumonia caused by penicillin-resistant Streptococcus pneumoniae. AB - BACKGROUND: Antimicrobial resistance rates for Streptococcus pneumoniae continue to increase worldwide, and resistance to nearly every major class of antimicrobials used to treat pneumococcal infections has been reported. Gatifloxacin (GFLX) is one of the quinolones that have strong activity against S. pneumoniae. METHODS: We compared the bacteriological, pharmacological and histopathological effects of orally administered GFLX with those of levofloxacin (LVFX) and ciprofloxacin (CPFX) in a murine model of pneumonia caused by penicillin-resistant S. pneumoniae (PRSP). RESULTS: Treatment with GFLX resulted in a significant decrease in the number of viable bacteria (control, CPFX, LVFX, and GFLX: 6.48 +/- 0.36, 6.44 +/- 0.27, 5.51 +/- 0.15, and 4.89 +/- 0.28 log10 CFU/lung, respectively, mean +/- SD). A significant decrease in mortality was observed in the GFLX-treated group in comparison with the other groups. Histopathological examination revealed that inflammatory changes in GFLX-treated mice were less marked than in the other mice. CONCLUSION: Our results suggest that orally administered GFLX is effective in PRSP pneumonia. The pharmacokinetic profiles also reflected the effectiveness of GFLX. PMID- 15282439 TI - Gemcitabine combined with infusional 5-fluorouracil and high-dose leucovorin for the treatment of advanced carcinoma of the pancreas. AB - BACKGROUND: 5-Fluorouracil (5-FU) and gemcitabine are the major active drugs in the treatment of pancreatic cancer. METHODS: Twenty-two patients with advanced pancreas cancer were treated with a new chemotherapy regimen consisting of infusional 5-FU and high-dose leucovorin with gemcitabine (GEMFUFOL). RESULTS: A total of 200 cycles of chemotherapy were administered. The response rate was 27.3%, all responses being partial. The median survival time and 1-year survival rate were, respectively, 13 months and 60.4%. The toxicity was very low and severe hematological toxicity was exceptional. CONCLUSION: The GEMFUFOL regimen can be an active regimen for the treatment of advanced pancreatic cancer and has a low toxicity. PMID- 15282440 TI - Ocular penetration of grepafloxacin after intravitreal administration in albino and pigmented rabbits. AB - Ocular penetration of grepafloxacin into several ocular tissues was determined in albino and pigmented rabbits following a single intravitreal administration. After administration, grepafloxacin was detected in all ocular tissues studied in both breeds of rabbits. The superior mean penetration ratios were found in the chorioretina and lens of albino rabbits, and in the chorioretina, iris and lens of pigmented rabbits. A significantly greater penetration of grepafloxacin was found in the chorioretina and iris of the pigmented rabbits than in those of the albino rabbits. As a final conclusion, grepafloxacin detected in different ocular structures could attain therapeutic concentrations against a variety of ocular conditions. PMID- 15282441 TI - Chemotherapy delays progression of motor neuron disease in the SOD1 G93A transgenic mouse. AB - BACKGROUND: A significant proliferation of glial cells occurs in the spinal cord and brainstem of SOD1 G93A transgenic mice with familial amyotrophic lateral sclerosis (ALS). Since activated glia may contribute to motor neuron degeneration, we tested whether inhibition of gliosis using low-dose chemotherapy is beneficial in this mouse model. METHODS: Mice were administered fortnightly intraperitoneal injections of 0.1 mg/kg vincristine (VIN) or saline commencing at postnatal day 68 before disease onset. Mice were sacrificed at end-stage disease, and spinal cords were examined for histology. RESULTS: Survival of VIN-treated mice was significantly increased at 132.0 +/- 4.1 days compared to control animals at 117.8 +/- 2.1 days (p < 0.05). Furthermore, analysis of microglia and astrocyte populations suggests a reduction in the former following VIN therapy. CONCLUSION: This study suggests that chemotherapy may offer an alternative therapy or co-therapy for ALS. PMID- 15282442 TI - Cardiovascular complications in acromegaly. AB - Cardiovascular morbidity and mortality are increased in acromegaly. In fact, GH and IGF-I excess induces a specific cardiomyopathy. The early stage of acromegaly is characterized by the hyperkinetic syndrome (high heart rate and increased systolic output). Frequently, concentric biventricular hypertrophy and diastolic dysfunction occur in acromegaly, leading to an impaired systolic function ending in heart failure if the disease is untreated or unsuccessfully untreated. Besides, abnormalities of cardiac rhythm and of valves have been also described in acromegaly. The coexistence of other complications, such as arterial hypertension and diabetes, aggravates the acromegalic cardiomyopathy. The suppression of GH/IGF-I following an efficacious therapy could decrease left ventricular mass and improve cardiac function. In conclusion, a careful evaluation of cardiac function, morphology and activity seems to be mandatory in acromegaly. PMID- 15282443 TI - Slaying the metabolic syndrome. Are we battling the Hydra or the Chimera? AB - Metabolic syndrome is the latest moniker for an alliance of pathologic conditions that conspire to amplify the risk of atherosclerosis or type 2 diabetes. Several recent advances in the understanding of this condition have led to its widespread adoption in clinical practice, prompted by influential practice guidelines. In particular, guidelines relating to the management of hyperlipidemia and hypertension afford a prominent role for metabolic syndrome, as a risk factor and a target of therapy. In many ways, the scientific evidence base has not kept pace with the demand for treatment options, rendering therapy nebulous. Most prominently, we do not know whether to adopt a strategy based on a multi-pronged attack, or whether to concentrate our greatest efforts on attacking a critical weakness. We review the limited data available regarding metabolic syndrome, with a focus on expert opinion gleaned from clinical guidelines, and offer advice to the clinician from our own experience with this population. PMID- 15282444 TI - Natriuretic peptides and the management of heart failure. AB - Hyperactivation of the renin-angiotensin-aldosterone system (RAAS), heightened sympathetic drive and uncontrolled synthesis of inflammatory cytokines, exacerbates ventricular contractile dysfunction in heart failure patients. The pathophysiological consequences include excessive fluid retention, increased peripheral vascular resistance, and endothelial dysfunction. Consequently, the demand for additional work by the failing myocardium in the presence of a greater afterload cannot be sustained. Therapeutically exploiting the natriuretic peptide system may represent a physiological approach to dampen the deleterious effects of the neuroendocrine systems and inflammatory cytokines. In both patients and animal models of heart failure, pharmacologically increasing plasma natriuretic peptide levels ameliorated vascular tone, renal and endothelial function, and ventricular contractility. Based on these observations, the following review will highlight the therapeutic benefits of the natriuretic peptide system in heart failure. PMID- 15282445 TI - Polycystic ovary syndrome and cardiovascular disease. AB - The aim of the present paper is to analyze recent literature concerning the incidence of cardiovascular complications in women suffering from polycystic ovary syndrome (PCOS). The study takes into consideration all the studies that have been published to date in the international literature in order to clarify whether or not PCOS is able to determine an early onset or whether it is responsible for a higher global incidence of cardiovascular complications in adult age. The main difficulty lies in the absence of prospective studies owing to the long period of time existing between the diagnosis of PCOS and cardiovascular disease which notoriously has a long latency period. Much attention has been paid in the literature, on the other hand, to the analysis of the incidence of cardiovascular risk factors in women suffering from PCOS. Although epidemiological studies have not evidenced an increased incidence of death from cardiovascular events in women suffering from PCOS, the above conclusions might well be invalidated by a patient selection bias, by obsolete diagnostic criteria or by medical or surgical therapies that could influence the outcome of the disease and which are not considered as a confusion factor. Undoubtedly, all the data available up to the present suggest that PCOS possesses the intrinsic conditions that lead to an increased incidence of factors predisposing to cardiovascular diseases. Future longitudinal studies of a prospective nature might be useful for understanding whether the higher incidence of predisposing factors might also lead to greater expectation of cardiovascular events or whether medical therapies or other factors (improvement in endocrine symptomatology with the menopause?) may prevent the increase in the expected incidence of these events. PMID- 15282446 TI - Thyroid hormone and the cardiovascular system. AB - Thyroid hormone is an important regulator of cardiac function and cardiovascular hemodynamics. Triiodothyronine, (T(3)), the physiologically active form of thyroid hormone, binds to nuclear receptor proteins and mediates the expression of several important cardiac genes, inducing transcription of the positively regulated genes including alpha-myosin heavy chain (MHC) and the sarcoplasmic reticulum calcium ATPase. Negatively regulated genes include beta-MHC and phospholamban, which are down regulated in the presence of normal serum levels of thyroid hormone. T(3) mediated effects on the systemic vasculature include relaxation of vascular smooth muscle resulting in decreased arterial resistance and diastolic blood pressure. In hyperthyroidism, cardiac contractility and cardiac output are enhanced and systemic vascular resistance is decreased, while in hypothyroidism, the opposite is true. Patients with subclinical hypothyroidism manifest many of the same cardiovascular changes, but to a lesser degree than that which occurs in overt hypothyroidism. Cardiac disease states are sometimes associated with the low T(3) syndrome. The phenotype of the failing heart resembles that of the hypothyroid heart, both in cardiac physiology and in gene expression. Changes in serum T(3) levels in patients with chronic congestive heart failure are caused by alterations in thyroid hormone metabolism suggesting that patients may benefit from T(3) replacement in this setting. PMID- 15282447 TI - Violence-related behaviors among high school students--United States, 1991-2003. AB - Homicide and suicide are responsible for approximately one fourth of deaths among persons aged 10-24 years in the United States. Two of the national health objectives for 2010 are to reduce the prevalence of physical fighting among adolescents to < or =32% and to reduce the prevalence of carrying a weapon by adolescents on school property to < or =4.9%. To examine changes in violence related behaviors among high school students in the United States during 1991 2003, CDC analyzed data from the national Youth Risk Behavior Survey (YRBS). This report summarizes the results of that analysis, which indicated that most violence-related behaviors decreased during 1991-2003; however, students increasingly were likely to miss school because they felt too unsafe to attend. In addition, in 2003, nearly one in 10 high school students reported being threatened or injured with a weapon on school property during the preceding 12 months. Schools and communities should continue efforts to establish physical and social environments that prevent violence and promote actual and perceived safety in schools. PMID- 15282448 TI - Racial/ethnic disparities in neonatal mortality--United States, 1989-2001. AB - Neonatal mortality (i.e., death at age <28 days) accounts for approximately two thirds of infant deaths in the United States. During 1989-2001, neonatal mortality rates (NMRs) declined; however, 2002 preliminary data indicated an increase. To characterize trends in neonatal mortality by gestational age and race/ethnicity, CDC analyzed linked birth/infant death data sets for 1989--1991 and 1995-2001 (2002 linked data were not available). This report summarizes the results of that analysis, which indicated that 1) extremely preterm infants (i.e., born at <28 weeks' gestation) accounted for 49%-58% of neonatal deaths during 1989-2001 and 2) racial/ethnic disparities persisted despite NMR declines among infants of all gestational ages. Public health practitioners, researchers, and clinicians can use these results to determine the efficacy of prevention programs at a national level and consider new studies and programs aimed at reducing preterm births and NMR disparities among racial/ethnic populations. PMID- 15282449 TI - National, state, and urban area vaccination coverage among children aged 19-35 months--United States, 2003. AB - Each annual birth cohort in the United States comprises approximately 4 million infants. Maintaining the gains in vaccination coverage achieved during the 1990s among these children poses a continuing challenge for public health practitioners. The National Immunization Survey (NIS) provides estimates of vaccination coverage among children aged 19-35 months for each of the 50 states and 28 selected urban areas. This report summarizes NIS results for 2003, which indicated substantial increases nationwide in coverage with > or =1 dose of varicella vaccine (VAR) and > or =3 doses of pneumococcal conjugate vaccine (PCV) and the highest coverage ever for all vaccines; however, wide variability in coverage continues among states and urban areas. Continued vigilance is needed to maintain high levels of coverage, and sustained efforts will be required to reduce geographic disparities in coverage. PMID- 15282450 TI - West Nile virus activity--United States, July 21-27, 2004. AB - During July 21-27, a total of 83 cases of human West Nile virus (WNV) illness were reported from 13 states (Alabama, Arizona, Arkansas, California, Colorado, Florida, Illinois, Iowa, Missouri, New Mexico, Ohio, Pennsylvania, and South Dakota). PMID- 15282451 TI - Regional myocardial ischemia in hypertrophic cardiomyopathy: impact of myectomy. AB - OBJECTIVE: Chest pain is a common finding in patients with hypertrophic cardiomyopathy and can be observed in 40% to 50% of all patients. However, the pathogenesis of these ischemia-like symptoms is still unclear. METHODS: Twenty two patients with hypertrophic cardiomyopathy and 15 controls underwent positron emission tomography for evaluation of regional myocardial perfusion and coronary flow reserve (hyperemic/baseline myocardial blood flow). Myocardial perfusion (mL/min/g) was measured using [(13)N]ammonia at rest and during hyperemia with dipyridamole (0.56 mg/kg intravenously). Regional coronary flow reserve was assessed in 3 planes (apical, midventricular, basal) in 4 regions (septal, anterior, lateral, inferior). Patients were divided into 2 groups: group 1 consisted of 11 patients treated with surgical myectomy (age 56 +/- 10 years) and group 2 consisted of 11 patients treated medically (age 53 +/- 13 years). RESULTS: Mean global coronary flow reserve was 3.87 +/- 0.92 in controls but 2.31 +/- 0.40 in operated (P <.001 vs controls) and 1.76 +/- 0.58 in medically treated patients (P <.001 vs controls, P <.05 vs operated). Similarly, septal coronary flow reserve was 4.19 +/- 1.22 in controls but significantly reduced in operated patients (2.26 +/- 0.48, P <.001 vs controls) and in medically treated patients (1.76 +/- 0.58; P <.001 vs controls). However, septal flow reserve was significantly higher in operated patients than in patients with medically treated hypertrophic cardiomyopathy (+37%, P <.05), mainly due to a reduced resting myocardial perfusion. CONCLUSIONS: Global and regional myocardial perfusion is reduced in patients with hypertrophic cardiomyopathy. However, myectomy may have a beneficial effect on septal perfusion and flow reserve. Thus, ischemia seems to play an important role in the symptomatology and pathophysiology of hypertrophic cardiomyopathy. PMID- 15282452 TI - Toll-like receptor 4 mediates ischemia/reperfusion injury of the heart. AB - BACKGROUND: Restoration of blood flow to the ischemic heart may paradoxically exacerbate tissue injury (ischemia/reperfusion injury). Toll-like receptor 4, expressed on several cell types, including cardiomyocytes, is a mediator of the host inflammatory response to infection. Because ischemia/reperfusion injury is characterized by an acute inflammatory reaction, we investigated toll-like receptor 4 activation in a murine model of regional myocardial ischemia/reperfusion injury. We used C3H/HeJ mice, which express a nonfunctional toll-like receptor 4, to assess the pertinence of this receptor to tissue injury after reperfusion of ischemic myocardium. METHODS: Wild-type mice (C3H/HeN) or toll-like receptor 4 mutant mice (C3H/HeJ) were subjected to 60 minutes of regional myocardial ischemia followed by 2 hours of reperfusion. At the end of reperfusion, the area at risk and the myocardial infarct size were measured as the end point of myocardial ischemia/reperfusion injury. Myocardial mitogen activated protein kinase activation was measured by Western blotting, and nuclear translocation of nuclear factor-kappaB and activator protein-1 was determined by electrophoretic mobility shift assay. Ischemia/reperfusion-injured myocardium was also assessed by ribonuclease protection assay for expression of inflammatory mediators (tumor necrosis factor-alpha, interleukin-1beta, monocyte chemotactic factor-1, and interleukin-6). RESULTS: The area at risk was similar for all groups after myocardial ischemia/reperfusion injury. There was a 40% reduction in infarct size (as a percentage of the area at risk) in C3H/HeJ mice compared with C3H/HeN mice (P =.001). Within the myocardium, significant activation of c-Jun N terminal kinase, p38, and extracellular signal-regulated kinase was observed in both strains after ischemia and during reperfusion as compared with an absence of mitogen-activated protein kinase activation during sham operations; however, c Jun N-terminal kinase activity, but not p38 or extracellular signal-regulated kinase activity, was significantly reduced in C3H/HeJ mice (P <.05). In both groups, nuclear factor-kappaB and activator protein-1 nuclear translocation occurred in the myocardium during myocardial ischemia/reperfusion injury, but, by densitometric analysis, nuclear translocation of nuclear factor-kappaB and activator protein-1 was significantly decreased in C3H/HeJ mice compared with C3H/HeN mice. Interleukin-1beta, monocyte chemotactic factor-1, and interleukin-6 were detectable in reperfused ischemic myocardium but were not detected in sham operated myocardium; the expression of each of these mediators was significantly decreased in the myocardial tissue of C3H/HeJ mice when compared with expression in the control C3H/HeN mouse strain. CONCLUSIONS: Our data suggest that toll-like receptor 4 may mediate, at least in part, myocardial ischemia/reperfusion injury. Inhibition of toll-like receptor 4 activation may be a potential therapeutic target to attenuate ischemia/reperfusion-induced tissue damage in the clinical setting. PMID- 15282453 TI - Nitric oxide attenuates cardiomyocytic apoptosis via diminished mitochondrial complex I up-regulation from cardiac ischemia-reperfusion injury under cardiopulmonary bypass. AB - OBJECTIVE: This study tested the hypothesis that cardioplegic solution supplemented with a nitric oxide donor agent attenuates postischemic cardiomyocytic apoptosis by reduction of mitochondrial complex I up-regulation during global cardiac arrest under cardiopulmonary bypass. METHODS: Twenty-four anesthetized dogs supported by total vented bypass were divided evenly into 4 groups (n = 6) and subjected to 60 minutes of hypothermic ischemia followed by 4 degrees C multidose crystalloid cardioplegic solution infusion. Hearts received either standard crystalloid cardioplegic solution (control), crystalloid cardioplegic solution supplemented with 2 mmol/L L-arginine (L-Arg group), crystalloid cardioplegic solution supplemented with 400 micromol/L N(G) monomethyl-L-arginine (L-NMMA group), or crystalloid cardioplegic solution supplemented with 100 micromol/L of NO donor compound (3-morpholinosydnonimine; SIN-1 group). After 60 minutes of cardioplegic arrest, the heart was reperfused for a total of 240 minutes after discontinuation of bypass. The occurrence of cardiomyocytic apoptosis was assessed by terminal deoxynucleotidyl transferase mediated dUTP-biotin nick end labeling and Western blot analysis of caspase-3. RESULTS: The occurrence of cardiomyocytic apoptosis was significantly reduced in SIN-1 and L-Arg groups compared with the control group. Mitochondrial complex I mRNA was up-regulated in the control group, and its expression was significantly higher in the L-NMMA group but significantly reduced in the SIN-1 and L-Arg groups. Western blot analysis of Bcl-2 and cytochrome c, an index of mitochondrial damage in postischemic myocardium, revealed a similar pattern. CONCLUSION: Nitric oxide-supplemented crystalloid cardioplegic solution diminished postischemic cardiomyocytic apoptosis after global cardiac arrest under cardiopulmonary bypass, possibly via prevention of mitochondrial complex I up-regulation. PMID- 15282454 TI - Recombinant hirudin enhances cardiac output and decreases systemic vascular resistance during reperfusion after cardiopulmonary bypass in a porcine model. AB - OBJECTIVE: Cardiopulmonary bypass and surgical stress are accompanied by a systemic inflammatory response and activation of coagulation. Thrombin forms fibrin and activates platelets and neutrophils. Consequently, disseminated microthrombosis might increase capillary vascular resistance and thus impair reperfusion. We hypothesized that recombinant hirudin, a direct inhibitor of thrombin, could attenuate coagulation and enhance microvascular flow during reperfusion. METHODS: Twenty pigs undergoing 60 minutes of aortic clamping and 75 minutes of normothermic perfusion were randomized in a blinded setting to receive an intravenous bolus of recombinant hirudin (10 mg, 0.4 mg/kg; n = 10) or placebo (n = 10) 15 minutes before aortic declamping and then continued with an intravenous 135-minute infusion of recombinant hirudin (3.75 mg/h, 0.15 mg/kg) or placebo. Thrombin-antithrombin complexes, activated clotting times, and several hemodynamic parameters were measured before cardiopulmonary bypass, after weaning from cardiopulmonary bypass, and at 30, 60, 90, and 120 minutes after aortic declamping. Intramucosal pH and Pco(2) were measured from the luminal surface of ileum simultaneously with arterial gas analysis at 30-minute intervals. RESULTS: Recombinant hirudin inhibited thrombin formation after aortic declamping; at 120 minutes, thrombin-antithrombin complexes levels (microg/L, mean +/- SD) were 75 +/- 21 and 29 +/- 44 (P <.001) for placebo and pigs receiving recombinant hirudin, respectively. When compared with the placebo group, pigs receiving recombinant hirudin showed significantly higher stroke volume, cardiac output, and lower systemic vascular resistance at 60 and 90 minutes after aortic declamping (P <.05). Based on arteriomucosal Pco(2) and pH differences, progressive worsening of intestinal microcirculatory perfusion occurred in the placebo group but not in the recombinant hirudin group. CONCLUSION: Infusion of thrombin inhibitor recombinant hirudin during reperfusion was associated with attenuated postischemia left ventricular dysfunction and decreased vascular resistance. Consequently microvascular flow was improved during ischemia reperfusion injury. Control of thrombin formation during reperfusion may be a feasible approach to improve oxygen delivery to reperfused vascular beds. PMID- 15282455 TI - Platelet activation and aggregation profile in prolonged external ventricular support. AB - BACKGROUND: Platelet function plays a major role in the understanding of thromboembolic events in prolonged mechanical support. We studied the platelet activation, platelet aggregation profile, and efficacy of aspirin in patients in whom an external ventricular assist device had been implanted. PATIENTS AND METHODS: Fifteen patients were studied prospectively up to 6 weeks after implantation of the same type of ventricular assist device. Platelet function was studied weekly before daily aspirin administration. Aspirin efficacy was tested ex vivo by measuring platelet aggregation triggered by arachidonic acid. Flow cytometry was used to quantify the spontaneous and induced (adenosine diphosphate stimulation) expression of glycoproteins alphaIIbbeta3, Ibalpha, and CD62P on platelet membranes. The plasma levels of von Willebrand factor (von Willebrand factor activity and von Willebrand factor antigen) and fibrinogen were also determined. RESULTS: Six of the 15 patients (26%) maintained an arachidonic acid induced platelet aggregation despite daily aspirin treatment (250 mg). CD62P values remained increased during a 5-week postoperative period. Spontaneous levels of glycoproteins alphaIIbbeta3 and Ibalpha on platelet membranes remained within a normal range with a preserved reactivity. The plasma levels of fibrinogen and von Willebrand factor remained increased during the entire study period. CONCLUSION: In patients with an implanted external ventricular assist device, the platelet activation profile displays a persistent activation with a preserved reactivity associated with a persistent high inflammatory state and endothelial activation. PMID- 15282456 TI - Ischemic preconditioning or heat shock pretreatment ameliorates neuronal apoptosis following hypothermic circulatory arrest. AB - OBJECTIVE: Hypothermic circulatory arrest has been widely used in complex cardiac and aortic surgery. Stroke and/or neurologic injury can occur after prolonged hypothermic circulatory arrest, possibly due to apoptosis. Ischemic preconditioning has been widely used as a neuroprotective tool, but its application in neuronal injury under hypothermic circulatory arrest has never been studied. METHODS: Forty male New Zealand white rabbits were placed on closed chest cardiopulmonary bypass, subjected to hypothermic circulatory arrest, and rewarmed to normothermia. Experimental groups were treated with heat shock or ischemic preconditioning before hypothermic circulatory arrest. Hippocampal CA1 neurons were analyzed histopathologically. Apoptosis was confirmed by TUNEL assay and Western blot analysis, and serum S-100beta levels, c-Fos and Bcl-2 antibodies, and caspase-3 and heat shock protein 70 levels were measured. RESULTS: After 2-hour hypothermic circulatory arrest and 4-hour reperfusion, apoptosis was observed in hippocampal CA1 neurons with elevation of serum S 100beta levels, which could be ameliorated by ischemic preconditioning or heat shock manipulations. TUNEL-positive nuclear expression of caspase-3 increased after hypothermic circulatory arrest (3.08% +/- 0.71%, P <.001) and was diminished with ischemic preconditioning (1.61% +/- 0.42%) and heat shock (1.72% +/- 0.38%) manipulations. Ischemic preconditioning or heat shock manipulations produced diverse patterns of heat shock protein 70, c-Fos, and Bcl-2 protein expression, suggesting that these manipulations provide neuroprotection via different pathways. CONCLUSIONS: Ischemic preconditioning and heat shock can attenuate hippocampal CA1 neuronal apoptosis after prolonged hypothermic circulatory arrest under cardiopulmonary bypass. The expression of heat shock protein 70 may not play a major role in the first window of ischemic preconditioning-induced neuroprotection. PMID- 15282457 TI - Can extra protamine eliminate heparin rebound following cardiopulmonary bypass surgery? AB - OBJECTIVES: Heparin rebound, the reappearance of anticoagulant activity after adequate neutralization with protamine, is thought to contribute to excessive postoperative bleeding after cardiac surgery. We have previously demonstrated that a significant amount of heparin is bound nonspecifically to plasma proteins and is incompletely neutralized by protamine. The aim of this study was to investigate whether clinically important bleeding attributable to heparin rebound can be eliminated by infusion of small amounts of additional protamine for 6 hours postoperatively and whether this treatment can reduce mediastinal blood loss. METHODS: Three hundred patients undergoing elective cardiac surgery were randomized to receive either a continuous infusion of protamine sulphate (25 mg/h for 6 hours) postoperatively or saline placebo. Serial blood samples were obtained to measure thrombin clotting time and anti-factor Xa activity. Heparin bound nonspecifically to plasma proteins was measured after displacement with a chemically altered heparin with low affinity to antithrombin. Mediastinal blood loss and transfusion requirements were recorded. RESULTS: Heparin rebound was demonstrated in every patient in the placebo group as reflected by increased thrombin clotting time, anti-factor Xa activity, and protein-bound heparin between 1 and 6 hours after surgery. In contrast, heparin rebound was eliminated in the protamine infusion group. The thrombin clotting time was normalized and both heparin concentration and protein-bound heparin were almost undetectable (P <.001). There was a modest 13% reduction in postoperative bleeding but this did not reduce blood transfusions. No adverse events were attributable to the extra protamine. CONCLUSIONS: Postoperative protamine infusion was able to almost totally abolish heparin rebound. In the context of this study, protamine infusion resulted in reduced postoperative bleeding but the magnitude was insufficient to alter transfusion requirements. PMID- 15282458 TI - Interaction of temperature with hematocrit level and pH determines safe duration of hypothermic circulatory arrest. AB - OBJECTIVE: Previous studies have demonstrated that both hematocrit level and pH influence the protection afforded by deep hypothermic circulatory arrest. The current study examines how temperature modulates the effect of hematocrit level and pH in determining a safe duration of circulatory arrest. The study also builds on previous work investigating the utility of near-infrared spectroscopy as a real-time monitor of cerebral protection during circulatory arrest. METHODS: Seventy-six piglets (9.3 +/- 1.2 kg) underwent circulatory arrest under varying conditions with continuous monitoring by means of near-infrared spectroscopy (hematocrit level of 20% or 30%; pH-stat or alpha-stat strategy; temperature of 15 degrees C or 25 degrees C; arrest time of 60, 80, or 100 minutes). Neurologic recovery was evaluated daily by a veterinarian, and the brain was fixed in situ on postoperative day 4 to be examined on the basis of histologic score in a blinded fashion. RESULTS: Multivariable analysis of total histologic score revealed that higher temperature, lower hematocrit level, more alkaline pH, and longer hypothermic circulatory arrest duration were predictive of more severe damage to the brain (P <.01). Regression modeling revealed that higher temperature exacerbated the disadvantage of a lower hematocrit level and longer arrest times but not pH strategy. Normalized oxyhemoglobin nadir time, derived from near-infrared spectroscopy, was positively correlated with neurologic recovery on the fourth postoperative day and with total histologic injury score (P <.0001). CONCLUSION: Hematocrit level and pH, as well as temperature, determine the safe duration of hypothermic circulatory arrest. Near-infrared spectroscopy is a useful real-time monitor of safe duration of circulatory arrest. PMID- 15282459 TI - Total aortic arch replacement with a branched graft and limited circulatory arrest of the brain. AB - BACKGROUND: Total replacement of the aortic arch is commonly performed with either antegrade perfusion of the brachiocephalic arteries by means of direct cannulation or with an interval of hypothermic circulatory arrest of at least 30 to 40 minutes. We present a technique with a branched graft that uses antegrade brain perfusion without the need for direct cannulation of the brachiocephalic arteries or a separate perfusion circuit, with only a brief period of circulatory arrest of the brain. METHODS: Twelve patients underwent resection of the aortic arch through either a midline sternotomy (4 patients) or a bilateral anterior thoracotomy (8 patients). The right axillary artery was used for arterial return and for brain perfusion. After establishing hypothermic circulatory arrest, the brachiocephalic arteries were detached from the aorta, flushed, and occluded with clamps. Hypothermic perfusion of the brain was established through the right axillary artery, and the brachiocephalic arteries were sequentially attached to the limbs of a branched aortic graft. Flow to the brain was then established in the antegrade direction through the axillary artery. RESULTS: The mean duration of circulatory arrest of the brain at a mean nasopharyngeal temperature of 16 degrees C was 8.8 minutes (range, 6-13 minutes). The subsequent period of hypothermic (20 degrees C-22 degrees C) brain perfusion, during which the 3 branches of the graft were attached to the brachiocephalic arteries, averaged 35 minutes (range, 23-44 minutes). All the patients survived the procedure and were discharged from the hospital. No patient sustained a permanent neurologic deficit. One patient had lethargy for 2 days, with full recovery. Nine of the 12 patients were extubated within 72 hours. CONCLUSIONS: This technique obviates the need for direct cannulation of the brachiocephalic arteries and for a separate perfusion circuit and requires only a brief period of circulatory arrest of the brain. PMID- 15282460 TI - Does off-pump total arterial grafting increase the incidence of intraoperative graft failure? AB - BACKGROUND: Early graft failure is a common cause of cardiac mortality and morbidity after coronary artery bypass grafting, but there is little information on its natural incidence. Furthermore, there is particular concern about graft patency in off-pump coronary artery bypass grafting and total arterial grafting. METHODS: We performed a prospective observational study to assess intraoperative graft patency in patients undergoing off-pump and on-pump coronary artery bypass grafting, who also underwent total arterial grafting. We used an intraoperative imaging system, SPY (Novadaq Technologies Inc), based on the fluorescent properties of indocyanine green dye. RESULTS: We assessed the intraoperative graft patency of 533 conduits in 200 patients. The mean number of grafts was 2.7 per patient. Of these patients, 155 (78%) had off-pump coronary artery bypass grafting, and 45 (22%) had on-pump coronary artery bypass grafting. Overall, 161 (80%) had total arterial grafting, with composite arterial grafting performed in 120 (60%) patients. Fluorescence, confirming graft patency, was observed in all but 8 (1.5%) conduits in 8 (4%) patients, necessitating graft revision. Six (3.9%) and 2 (4.4%) of these patients, respectively, had off-pump coronary artery bypass grafting and on-pump coronary artery bypass grafting. CONCLUSION: Intraoperative fluorescence imaging demonstrated a low (1.5%) but well-defined incidence of intraoperative graft failure, which affects around 4% of patients. This emphasizes the need for routine assessment of graft patency. Intraoperative fluorescence imaging permits detection and revision of failed grafts in the operating room. We found no difference in the incidence of failed grafts when comparing on-pump and off-pump total arterial grafting. PMID- 15282461 TI - Angiogenic growth factors and/or cellular therapy for myocardial regeneration: a comparative study. AB - BACKGROUND: Locally delivered angiogenic growth factors and cell implantation have been proposed for patients with myocardial infarcts without a possibility of percutaneous or surgical revascularization. The goal of this study was to compare the effects of these techniques in an experimental model of myocardial infarct. METHODS: Left ventricular myocardial infarction was created in 27 sheep by ligation of 2 coronary arteries. Three weeks after creation of the infarct, animals were randomized into 4 groups. In group 1, sheep received a culture medium injection to the infarct area (control group); group 2 underwent autologous myoblast implantation; group 3 received vascular endothelial growth factor; and group 4 received injection of both vascular endothelial growth factor and myoblasts. Evaluation included serum troponin IC levels, echocardiography (2 dimensional and color kinesis), and immunohistologic studies for quantitative analysis of capillaries (3 months after surgery). RESULTS: Four animals died of refractory ventricular fibrillation during myocardial infarction; 2 died after surgery because of stroke and 2 because of infections. Serum troponin increased to 45.6 +/- 4.7 ng/mL at postinfarction day 2. Echocardiography at 3 months showed a significant limitation of left ventricular dilation in the cell group (57 +/- 11.1 mL) and in the cell plus vascular endothelial growth factor group (58.6 +/- 6.6 mL: control group, 74.4 +/- 11.2 mL; vascular endothelial growth factor group, 68.1 +/- 3.4 mL). Color kinesis echography showed important improvements of regional fractional area change in the cell group (from 13.6% +/- 0.8% to 21.1% +/- 1.5%) and in the cell plus vascular endothelial growth factor group (from 12.8% +/- 0.9% to 18.7% +/- 2.3%). The number of capillaries increased in the peri-infarct region of the vascular endothelial growth factor group (1036 +/- 75: control group, 785 +/- 31; cell group, 830 +/- 75; cell plus vascular endothelial growth factor group, 831 +/- 83). CONCLUSIONS: In the cell therapy groups, regional ventricular contractility improved and heart dilatation was limited compared with either vascular endothelial growth factor or control; thus, postischemic remodeling was reduced. Angiogenesis was demonstrated in the vascular endothelial growth factor group, without improvement of ventricular function and remodeling. To improve local conditions for cell survival, further studies are warranted on prevascularization of myocardial scars with angiogenic therapy. PMID- 15282462 TI - Thoracic surgical operations in patients enrolled in a computed tomographic screening trial. AB - OBJECTIVE: Screening for lung cancer with computed tomography may detect cancers at an earlier stage but may also result in overdiagnosis. We reviewed the thoracic surgical operations performed on patients enrolled in our computed tomographic screening program. METHODS: From January 1999 through December 2002, screening computed tomography for lung cancer was performed annually on 1520 participants. All participants were at least 50 years old and smoked more than 20 pack/y. We found 3130 indeterminate pulmonary nodules in 1112 participants (73%). Fifty-five participants (3.6%) underwent 60 thoracic operations for a variety of indications. The medical records of these 55 patients were reviewed. RESULTS: Indications for operation included suspicious pulmonary nodules, mediastinal adenopathy, and a spontaneous pneumothorax. Operations performed included a lobectomy in 37 cases, wedge resection in 11, segmentectomy in 6, video-assisted thoracoscopic surgical talc pleurodesis in 1, bilobectomy in 2, mediastinoscopy in 2, and anterior mediastinotomy in 1. Benign disease was found in 10 patients (18.1%), and lung cancer was found in 45 (81.9%), 2 of whom had metachronous lung cancers. Cell types were adenocarcinoma in 15 cancers, bronchioloalveolar cell carcinoma in 13, squamous cell in 13, carcinoid in 2, small cell in 2, and large cell and undifferentiated non-small cell in 1 case each. Twenty-eight cancers were classified as stage IA, 4 as IB, 4 as IIA, 1 as IIB, 4 as IIIA, 3 as IIIB, 1 as IV, and 2 as limited small cell carcinoma. Complications occurred in 27% of patients. Operative mortality was 1.7%. CONCLUSION: Computed tomographic screening finds a large number of indeterminate pulmonary nodules in smokers 50 years old or older, most of which are observed and not operated on. Although 47 cancers were detected thus far in this highly selected group of patients, this represents only 1.5% of the pulmonary nodules identified. PMID- 15282463 TI - Clinical outcomes after separate and composite replacement of the aortic valve and ascending aorta. AB - OBJECTIVES: We sought to compare the clinical profile and outcomes of operations for aortic valve disease and ascending aortic aneurysm in patients treated with aortic valve replacement and supracoronary replacement of the ascending aorta or composite replacement of the aortic valve and ascending aorta (Bentall operation). METHODS: From 1990 through 2001, 133 patients had aortic valve replacement and supracoronary replacement of the ascending aorta, and 452 patients had Bentall operations. Aortic valve replacement and supracoronary replacement of the ascending aorta was performed in patients who had aortic valve disease and dilation of the ascending aorta, whereas the Bentall operation was performed in patients with aortic root abnormality and ascending aortic aneurysm. Mean follow-up was 4.6 +/- 3.1 years and was 100% complete. RESULTS: Patients who had aortic valve replacement and supracoronary replacement of the ascending aorta were older (61 +/- 13 vs 52 +/- 16 years, P <.001) and more likely to have aortic stenosis, coronary artery disease, and mitral valve disease than those who had Bentall operations. The use of mechanical valves was equal in both groups (42% for aortic valve replacement and supracoronary replacement of the ascending aorta and 43% for the Bentall operation). Operative mortality was 5% for patients undergoing aortic valve replacement and supracoronary replacement of the ascending aorta and 4% for patients undergoing the Bentall operation (P =.45). Survival at 10 years was 57% +/- 8% for patients undergoing aortic valve replacement and supracoronary replacement of the ascending aorta and 74% +/- 4% for patients undergoing the Bentall operation (P =.04), but the type of operation had no effect on survival. Older age, moderate or severe left ventricular dysfunction, active endocarditis, previous cardiac surgery, and coronary artery disease were independent predictors of death. The freedom from reoperation at 10 years was 95% +/- 5% for patients undergoing aortic valve replacement and supracoronary replacement of the ascending aorta and 94% +/- 3% for patients undergoing the Bentall operation (P =.18). Reoperations were mostly because of tissue valve failure or endocarditis. The risk of valve-related complications was the same in both groups. No patient required reoperation for aortic root aneurysm after having aortic valve replacement and supracoronary replacement of the ascending aorta. CONCLUSIONS: Aortic valve replacement and supracoronary replacement of the ascending aorta and the Bentall operation provide comparable long-term results. The Bentall operation is more appropriate for patients with aortic root abnormality and a dilated ascending aorta, whereas aortic valve replacement and supracoronary replacement of the ascending aorta is a perfectly acceptable operation for patients with aortic valve disease, normal or mildly dilated aortic sinuses, and a dilated ascending aorta. PMID- 15282464 TI - Quality of life after aortic valve replacement with tissue and mechanical implants. AB - OBJECTIVES: We sought to determine whether changes in quality of life at 18 months following aortic valve replacement differ depending on the use of tissue valves or mechanical valves. METHODS: We prospectively studied 73 patients with tissue valve replacements and 53 patients with mechanical valve replacements performed from April 1998 through March 1999 at Yale-New Haven Hospital. Quality of life was measured at baseline and at 18 months using the Medical Outcomes Trust Short Form 36-Item Health Survey. RESULTS: Baseline unadjusted mean quality of-life scores were lower in tissue valve recipients than in mechanical valve recipients and, for both groups, were generally lower than US population norms. At 18 months postoperatively, quality-of-life scores were greatly improved in both groups and were comparable to population norms (ie, within one-half a standard deviation). After adjusting for baseline quality of life, age, and other prognostic factors in an analysis of covariance, improvements in quality-of-life scores for tissue valve recipients versus mechanical valve recipients were similar. Of 10 (8 domains and 2 summary) scales examined, the only significant difference between the 2 groups was for the improvement in role limitations due to physical problems (Role Physical), which was more favorable in patients with mechanical valve implants (P =.04). CONCLUSIONS: The use of tissue valve implants versus mechanical valve implants has little influence on improvement in quality of life at 18 months following aortic valve replacement. Thus, decisions about whether to choose a tissue valve or mechanical valve implant should depend upon other factors such as rates of complications and differences in the life span of the implants. PMID- 15282465 TI - Initial experience with the Heartstring proximal anastomotic system. AB - OBJECTIVE: Manipulation of the aorta has been shown to be associated with postoperative neurologic events after surgical myocardial revascularization when the aorta is diseased. The Heartstring proximal anastomotic system (Guidant, Indianapolis, Ind) is a device designed to assist in the performance of proximal anastomoses with minimal aortic manipulation. We describe our initial experience with this product. METHODS: Twelve patients with a diseased aorta who underwent off-pump myocardial revascularization and had their proximal anastomoses performed with the Heartstring device were studied for operative and postoperative outcomes and surgical technique. RESULTS: The mean age of the patients was 76 +/- 7 years. The estimated EuroSCORE perioperative mortality was 12% +/- 8%. The median number of distal anastomoses was 3. In all patients, a saphenous vein graft was anastomosed to the aorta using the Heartstring device. In 5 patients, the radial artery was used as a conduit and connected proximally to a left internal thoracic artery as a Y graft, to a saphenous vein graft as a "horseshoe," or on the hood of a saphenous vein graft. The operative and postoperative courses were uneventful. Three seals developed cracks and were not used. One seal developed an unravel in its periphery but was used successfully. CONCLUSIONS: The Heartstring proximal anastomotic system is a device that allows the surgeon to perform standard proximal clampless anastomoses. Elderly patients with a diseased aorta may benefit from this device. PMID- 15282466 TI - Late incidence and predictors of persistent or recurrent heart failure in patients with mitral prosthetic valves. AB - OBJECTIVE: The study's objective was to examine factors associated with persistent or recurrent congestive heart failure after mitral valve replacement. METHODS: Patients who underwent mitral valve replacement with contemporary prostheses (N = 708) were followed with annual clinical assessment and echocardiography. Cox proportional hazard models were developed to evaluate the impact of demographic, comorbid, and valve-related variables on the occurrence of congestive heart failure after mitral valve replacement, defined as the composite outcome of New York Heart Association class III or IV symptoms or death caused by congestive heart failure postoperatively. Factors associated with all-cause mortality were also examined. Models were bootstrapped 1000 times. RESULTS: The total follow-up was 3376 patient-years (mean 4.8 +/- 3.7 years, range 60 days to 17.1 years). Freedom from New York Heart Association III or IV symptoms or death caused by congestive heart failure was 96.1% +/- 0.8%, 82.7% +/- 1.7%, 66.4% +/- 3.0%, and 38.8% +/- 6.9% at 1, 5, 10, and 15 years, respectively. Preoperative New York Heart Association class, left ventricular grade, atrial fibrillation, coronary artery disease, smoking, persistent tricuspid regurgitation, and redo status predicted congestive heart failure postoperatively (all P <.05). Patients who underwent mitral valve replacement for pure mitral stenosis had less congestive heart failure events after surgery than those with regurgitation or mixed disease. Prosthesis size and elevated transprosthesis gradients were not predictive of freedom from congestive heart failure after mitral valve replacement. Atrial fibrillation, persistent tricuspid regurgitation, and surgical referral for mitral valve replacement at an advanced functional stage were also risk factors for all-cause mortality. CONCLUSIONS: This study identifies the incidence of and risk factors for congestive heart failure and death late after mitral valve replacement. Although prosthesis size has no effect, other potentially modifiable factors such as atrial fibrillation, persistent tricuspid regurgitation, and late surgical referral have a negative impact on freedom from congestive heart failure and overall survival after mitral valve replacement. PMID- 15282467 TI - Health-related quality of life after coronary artery bypass grafting: a gender analysis using the Duke Activity Status Index. AB - OBJECTIVE: Our objectives were to document the preoperative and postoperative functional status of patients undergoing coronary artery bypass grafting, to examine factors that influence functional recovery, and to determine whether gender differences exist in the preoperative and postoperative functional status with the Duke Activity Status Index. METHODS: One thousand eight hundred twenty five patients undergoing isolated coronary artery bypass grafting had baseline and follow-up quality-of-life surveys. Mean follow-up from baseline to postoperative Duke Activity Status Index was 8.0 months for women and men. The influence of 47 variables, in addition to baseline scores on postoperative functional status, was examined with logistic ordinal modeling. An ordinal model for the follow-up score was determined by means of backward selection, with variables retained if they satisfied the criterion of a P value of less than.05. RESULTS: Median baseline Duke Activity Status Index scores (women, 21.5; men, 32.2; P <.001) and first follow-up scores (women, 42.7; men, 58.2; P <.001) were lower in women than in men. Patients who were older and those who had chronic obstructive pulmonary disease, myocardial infarction, stroke, diabetes, vascular disease, postoperative serious infection, and return to the operating room had lower postoperative scores. After adjusting for these factors, women still had lower follow-up scores (odds ratio for men, 2.1 [95% confidence interval, 1.7 2.6]; P <.001). CONCLUSIONS: A number of preoperative factors, operative variables, and postoperative events are associated with functional recovery after coronary revascularization. In addition, female gender is associated with more postoperative functional impairment after adjusting for these perioperative variables. PMID- 15282468 TI - Medical and surgical outcome of tricuspid regurgitation caused by flail leaflets. AB - OBJECTIVE: We sought to evaluate the medical and surgical outcome of tricuspid regurgitation caused by flail leaflets. METHODS: We analyzed the cause, clinical presentation, outcome, and natural history of 60 patients with tricuspid regurgitation caused by flail leaflets, a cause of mostly severe and organic tricuspid regurgitation, diagnosed by means of echocardiography between 1980 and 2000. RESULTS: The main cause was traumatic (62%). Clinical presentation was often severe: 57% were symptomatic, 33% had a history of congestive heart failure, and 40% had a history of atrial fibrillation. Compared with expected survival of the US matched population, excess mortality (39% +/- 10% at 10 years or 4.5% yearly, P <.01) was observed. Even patients asymptomatic at presentation experienced high tricuspid-related event rates (at 10 years, 75% +/- 15% had symptoms or heart failure, atrial fibrillation, surgical intervention, or death). In those patients severe enlargement of right-sided chambers was predictive of poor outcome (at 5 years: 86% +/- 9% vs 39% +/- 11%, P <.01) independent of cause (P =.31). The poor medical outcome was further confirmed by high event rates (69% +/- 9% at 15 years) in the natural history beginning from the flail's occurrence. Tricuspid operations were performed in 33 patients (55% +/- 7% at 5 years), with valve repair in 82%, low mortality (3%), and, despite frequently refractory atrial fibrillation, symptomatic improvement in 88%. CONCLUSION: Tricuspid regurgitation caused by flail leaflets is a serious disease associated with excess mortality and high morbidity. Tricuspid valve repair can often be performed with low risk, allowing symptomatic improvement. These results suggest that surgical intervention should be considered early in the course of the disease before the occurrence of irreversible consequences. PMID- 15282469 TI - Residual dissection of the brachiocephalic arteries: significance, management, and long-term outcome. AB - OBJECTIVES: Residual dissection of the brachiocephalic arteries after operations for acute type A dissection is considered a benign condition that does not expose patients to late neurologic events. This retrospective study, conducted on an outpatient clinic basis between June 1995 and May 2003, had the objectives of evaluating the consequences of residual dissection of the brachiocephalic arteries, investigating the long-term outcomes of patients with this condition, and illustrating our approach to the condition. METHODS: Forty-two of 137 patients with spontaneous aortic dissection were identified as having residual dissection of the brachiocephalic arteries. There were 30 men and 12 women, with median age of 64.8 years. Patients were followed for a median time of 3.17 years (25th-75th percentile, 1.43-4.40 years; maximum, 7.5 years). The main outcome was the occurrence of cerebral ischemic events (transient ischemic attack or stroke) or death. The functional consequences of brachiocephalic artery dissection were studied by using duplex scanning and transcranial Doppler ultrasonography. RESULTS: Twenty-four focal neurologic complications occurred in 13 of 42 patients (incidence, 30.9%); major strokes occurred in 6 patients, and none were fatal. Minor strokes occurred in 12 patients. In all patients the damaged territory was dependent on a dissected artery. Kaplan-Meier (90-months) freedom from focal neurologic events was 55.7% (95% confidence interval, 33.7%-72.9%). Mean time of freedom from focal neurologic events was 64.5 months (95% confidence interval, 53.1-75.9 months). Positive transcranial Doppler monitoring for microembolic signals was 24.1%, and patients with clinical symptoms had higher microembolic signal counts than did those without symptoms (8.4/h vs 1.9/h, P <.001). Reduced cerebrovascular reactivity to hypercapnia, calculated by using the breath-holding index values, was associated with severely impaired brachiocephalic artery perfusion. The multivariable model for predictors of late stroke (minor and major) included the following variables: microembolic signal count (1 signal/h increase; relative risk, 1.27 [95% CI, 1.12-1.77]), breath-holding index (0.10 increase; relative risk, 0.91 [95% CI, 0.87-0.94]), and the presence of at least one carotid axis with a thrombosed false channel (relative risk, 0.82 [95% CI, 0.64-0.93]). Sixteen operations were performed in 12 patients to relieve residual dissection. CONCLUSIONS: These results suggest an increased risk of ischemic events ipsilateral to the dissected arteries. Strict follow-up and identification of subjects at risk implies the exact knowledge of vessel anatomy and perfusion status. Ultrasonographic transcranial Doppler examination plays an important role in the clinical work-up of these patients. PMID- 15282470 TI - Repair of an isolated huge congenital left ventricular diverticulum. PMID- 15282471 TI - Fontan operation for hypoplastic left heart syndrome with absent aortic valve. PMID- 15282472 TI - Congenital tracheoesophageal fistulas presenting in adults: presentation of two cases and a synopsis of the literature. PMID- 15282473 TI - Clinical-pathologic conference in general thoracic surgery: pulmonary artery fibrohistiocytic tumor in a child. PMID- 15282474 TI - Poly-adenosine diphosphate-ribose polymerase inhibition for myocardial protection: pathophysiologic and physiologic considerations. PMID- 15282475 TI - Interventional atrial incision in the Fontan operation: novel prophylaxis or iatrogenic substrate for intra-atrial reentrant tachycardia? PMID- 15282476 TI - Ventral cardiac denervation: is it truly an effective prophylaxis against atrial fibrillation after coronary artery bypass grafting? PMID- 15282477 TI - Use of enoxaparin in cardiac surgery. PMID- 15282478 TI - Reconstruction of double-outlet right ventricular outflow tract comprising a pulmonary artery flap in a child with an anomalous coronary artery. PMID- 15282479 TI - Tricuspid valve in hypoplastic left heart syndrome. PMID- 15282480 TI - The return of the coronary stapler: will a new technique overcome an old obstacle? PMID- 15282481 TI - Value of intraoperative pleural lavage in staging non-small cell lung cancer. PMID- 15282482 TI - The pseudorestrictive pattern of transmitral Doppler flow pattern after conversion of atrial fibrillation to sinus rhythm: is atrial or ventricular dysfunction to blame? AB - Patients with paroxysmal atrial fibrillation (AF) who have recently converted from AF to sinus rhythm often exhibit a restrictive Doppler pattern in the transmitral flow (TMF) velocity. However, the mechanism of this phenomenon has not been well defined. We evaluated the temporal change of TMF pattern and hemodynamics after conversion of AF to in sinus rhythm in an animal model. Eight open-chest dogs underwent 3 hours of pacing-induced AF. TMF velocities and pressure data were acquired at baseline (sinus rhythm), immediately after conversion of AF, and every 10 minutes thereafter. Early diastolic TMF velocity was increased immediately after conversion and recovered to the baseline value in 20 minutes. Atrial systolic TMF velocity was reduced after AF and recovered to baseline value in 20 to 30 minutes. Early diastolic/atrial systolic TMF velocity was increased after conversion, and recovered to baseline value in 20 to 30 minutes. The mean left atrial (LA) pressure increased immediately, 10 and 20 minutes after the conversion of AF to sinus rhythm. The left ventricular end diastolic pressure was increased and positive left ventricular dP/dt and tau were decreased immediately after AF, whereas they recovered within 10 minutes. In conclusion, a pseudorestrictive pattern of TMF after AF occurred as a result of transient LA mechanical functional impairment and increased LA pressure caused by LA stunning. Transient left ventricular diastolic dysfunction also effected the TMF velocity immediately after the conversion from AF to sinus rhythm, although it recovered faster than LA mechanical dysfunction. PMID- 15282483 TI - Clinical and echocardiographic predictors of left atrial appendage dysfunction in patients with mitral stenosis in sinus rhythm. AB - BACKGROUND: Mitral stenosis (MS) causes left atrial (LA) appendage (LAA) dysfunction resulting in reduced LAA flow velocities. Low LAA peak emptying velocity (PEV), determined by transesophageal echocardiography, is a risk for thrombus formation and systemic embolism. OBJECTIVE: We sought to investigate various clinical and echocardiographic predictors of low LAA blood flow velocities. METHODS: A total of 44 patients with newly diagnosed MS were classified into two groups on the basis of the presence of high (PEV > or = 46 cm/s) or low (PEV < 46 cm/s) LAA flow profile on Doppler transesophageal echocardiography. LAA flow velocities were measured to be 27.38 +/- 8.17 cm/s in patients with LAA dysfunction and 70.75 +/- 16.71 cm/s in high-flow profile (P <.0001). Simultaneous 12-lead electrocardiogram was used to measure P waves. RESULTS: P maximum, P dispersion, and LA diameter were significantly higher in patients with low LAA PEV (n = 32) than in those with high LAA PEV (111.87 +/- 16.93 vs 96.66 +/- 14.97, P =.0084; 73.12 +/- 20.7 vs 49.16 +/- 9.96, P <.0001; 46.06 +/- 4.384 vs 38.08 +/- 7.42 mm, P =.004; respectively). Patients with MS and low LAA blood flow had smaller mitral valve area compared with those with high LAA blood flow velocity (1.48 +/- 0.431 vs 1.85 +/- 0.442 cm(2), P =.02). Male sex, spontaneous echocontrast, and thrombus were more frequent in patients with low LAA PEV [7 [21.87%] vs 5 [41.66%], P =.026; 21 [65.62%] vs 4 [33.3%], P =.088; 4 [12.5%] vs none; respectively]. Mild MS was more frequent in patients with high blood flow velocity [6 [27.2%] vs 14 [63.6%], P =.03]. CONCLUSION: At linear regression analysis, only P-wave dispersion and LA diameter predicted the LAA mechanical dysfunction reflected as low LAA PEVs. PMID- 15282484 TI - Effects of percutaneous mitral commissurotomy on longitudinal left ventricular dynamics in mitral stenosis: quantitative assessment by tissue velocity imaging. AB - OBJECTIVE: We hypothesized that mitral annular velocities would improve immediately after relief of mitral stenosis and that serial assessment could be used as an index for quantifying functional changes after percutaneous mitral commissurotomy (PMC). METHODS: Longitudinal left ventricular annular velocities were quantified by spectral pulsed wave Doppler tissue velocity imaging in 25 patients (16 women; mean age [+/-SD], 29.2 +/- 8.6 years) who had isolated mitral stenosis and were in sinus rhythm, and were compared with 30 age- and sex-matched control subjects. Echocardiography was performed 1 to 24 hours before PMC and 48 to 72 hours after, and changes in velocities from the lateral and septal corners of the mitral annulus in early diastole, late diastole, isovolumic contraction, and ejection were recorded. RESULTS: Systolic and diastolic mitral annular velocities were significantly less in patients with mitral stenosis than in control subjects. After PMC, peak annular velocity of systolic excursion in ejection and peak annular velocity in early diastole showed significant improvement. The change in peak annular velocity in early diastole in the lateral wall correlated well with improvement in the mitral valve orifice area by planimetry (ratio of mitral valve orifice area, 1.92 +/- 0.42; ratio of peak annular velocity in early diastole, 1.36 +/- 0.22; r = 0.65; P <.001). CONCLUSION: Serial evaluation of changes in mitral annular velocities by Doppler tissue imaging aids clinical assessment of immediate improvement in left ventricular function after PMC. PMID- 15282485 TI - Prosthetic mitral regurgitation can be mimicked by Doppler color flow mapping: avoiding misdiagnosis. AB - OBJECTIVES: We sought to characterize a region of apparent systolic flow resembling mitral regurgitation (MR) in patients with mechanical disk mitral prostheses as artifact. BACKGROUND: Diagnosing MR in the presence of mechanical prostheses is challenging. Occasionally, important MR is suggested by a substantial region of systolic Doppler color flow in an acoustically shadowed region of the left atrium when, in fact, only trace MR exists. We hypothesized this pseudo-MR is caused by acoustic mirroring of the left ventricular outflow tract (LVOT) flow by sound reflected off the prosthesis, projecting flow into the left atrium because of longer transit time. METHODS: We reviewed 19 patients with mechanical mitral valves and trace MR by transesophageal echocardiography who had transthoracic echocardiography studies within 1 week (group A), and prospectively studied 22 consecutive patients by transthoracic echocardiography with subtle transducer angulation variation to detect pseudo-MR and characterize it by pulsed Doppler (group B). RESULTS: In group A, 12 of 19 patients had evidence of pseudo MR on review of their transthoracic echocardiograms, absent by transesophageal echocardiography. In group B, this pseudo-MR signal was present in 13 of 22 patients, with velocity and timing by pulsed Doppler comparable with LVOT flow (onset at 125 +/- 27 milliseconds vs 135 +/- 11 milliseconds from QRS, P = not significant). The angle between the mitral plane and the LVOT, which determines whether this mirroring can occur, was smaller for patients with pseudo-MR. CONCLUSION: Artifactual pseudo-MR is often seen with mechanical mitral prostheses. Its behavior and sensitivity to geometric relationships are consistent with mirroring of LVOT flow. Practically, potential misdiagnosis can be readily avoided by pulsed Doppler sampling, sparing the patient further procedures. PMID- 15282486 TI - Noninvasive measurement of systemic vascular resistance using Doppler echocardiography. AB - BACKGROUND: Systemic vascular resistance (SVR) is an integral therapeutic component of patients with heart failure and shock. We hypothesized that the ratio of the peak mitral regurgitant velocity (MRV) (m/s) to left ventricular outflow time-velocity integral (TVI(LVOT)) (cm) by Doppler would provide a noninvasive correlate of SVR. METHODS: SVR was correlated to MRV/TVI(LVOT) in 33 patients undergoing right heart catheterization. Receiver operating characteristic curves were generated to determine the best-balanced sensitivity and specificity to identify SVR > 14 Wood units (WU) and <10 WU. RESULTS: MRV/TVI(LVOT) correlated well with SVR (r = 0.842, 95% confidence interval 0.7 0.92, P <.001, Y = 0.459 + 49.397*X). By receiver operating characteristics, MRV/TVI(LVOT) > 0.27 had a 70% sensitivity and a 77% specificity to identify SVR > 14 WU. MRV/TVI(LVOT) < 0.2 had a 92% sensitivity and a 88% specificity to identify SVR < 10 WU. CONCLUSION: Doppler echocardiography provides a reliable noninvasive assessment of SVR. PMID- 15282487 TI - The peak atrioventricular pressure gradient to transmitral flow relation: kinematic model prediction with in vivo validation. AB - Physiologists and cardiologists estimate peak transvalvular pressure gradients (DeltaP) by Doppler echocardiographic imaging of peak flow velocities using the simplified Bernoulli relationship: DeltaP (mm Hg) = 4V(2) (m/s). Because left ventricular filling is initiated by mechanical suction, V can be predicted by the motion of a simple harmonic oscillator by the parametrized diastolic filling formalism that characterizes E-wave contours by 3 unique simple harmonic oscillator parameters: initial displacement (x(o) cm); spring constant (k g/s(2)); and damping constant (c g/s). Parametrized diastolic filling predicts peak atrioventricular pressure gradient as kx(o), the peak simple harmonic oscillator force. For validation, simultaneous (micromanometric) left ventricular pressure and E-wave data from 19 patients were analyzed. Model-predicted peak gradient (kx(o)) was compared with actual gradient (DeltaP(cath)) and with 4V(2). Multiple linear regression results for all patients yielded highly significant relation between kx(o) and DeltaP(cath) (kx(o) = m(1)DeltaP(cath) + b(1), where m(1) = 40.7 +/- 8.0 dyne/mm Hg, b(1) = 1540 +/- 116 dyne, r(2) = 0.97, P <.001). Regression analysis showed no significant correlation between 4V(2) and DeltaP(cath) (4V(2) = m(2)DeltaP(cath) + b(2), where m(2) = 0.01 +/- 0.03, m(2)/s(2)/mm Hg and b(2) = 2.07 +/- 0.44 m(2)/s(2), P = nonsignificant). We conclude that E-wave analysis by parametrized diastolic filling predicts peak atrioventricular gradients reliably and more accurately than 4V(2). PMID- 15282488 TI - Evaluation of ventricular synchrony using novel Doppler echocardiographic indices in patients with heart failure receiving cardiac resynchronization therapy. AB - Cardiac resynchronization therapy improves hemodynamics in selected patients with heart failure. Mechanic asynchrony parameters that may guide patient selection or therapy optimization are still being investigated. A biventricular (BiV) pacemaker was implanted in 34 patients with dilated ischemic, idiopathic, or valvular cardiomyopathy, and a QRS duration of > or =130 milliseconds. Two dimensional standard and Doppler tissue echocardiography was performed during right ventricular (RV), left ventricular (LV), BiV, and no pacing in a random and blinded manner. LV and BiV pacing increased stroke volume (P <.02 for both) and ejection fraction (P <.001 for both). Regional contractility assessed by displacement, strain rate, and peak systolic strain was improved in some segments (P <.05) during LV and BiV pacing. A homogenization of segmental contractions was observed during LV and BiV pacing as evaluated by net systolic displacement and segmental myocardial performance index. LV and BiV pacing provides benefits that can be quantified by echocardiography. PMID- 15282489 TI - Propagation velocity of mitral late flow toward the apex in healthy participants. AB - The transit time of mitral late flow (A wave) to the outflow tract (A-Ar interval) has been demonstrated to be shorter in the presence of increased ventricular stiffness. However, the A-Ar interval is just a measure of time duration and the data of intraventricular A-wave propagation velocity are still unavailable. In this study we presented the aging trend in various intracardiac Doppler signals, including the mitral A-wave propagation velocity toward the apex (APV(apex)). It was measured as the slope of the first aliasing velocity line segment of the mitral A wavefront at color M-mode Doppler analysis. Age correlated significantly with peak velocity of mitral early flow (E wave), peak A velocity, velocity ratio of E to A wave, deceleration time of E wave, and mitral E-wave propagation velocity toward the apex, but not with the A-Ar interval (r = 0.262, P =.066) or the APV(apex) (r = -0.047, P =.748). Neither the A-Ar interval nor the APV(apex) was different between the young and the elderly groups. Furthermore, there was no significant correlation between the APV(apex) and the A Ar interval (r = -0.135, P =.348). In conclusion, aging has a major adverse impact on myocardial relaxation associated with a minor one on the noninvasive indices of ventricular compliance. The APV(apex) is age-independent and, moreover, there is no significant correlation between the APV(apex) and the A-Ar interval. PMID- 15282490 TI - Distinctive changes in end-diastolic wall thickness and postsystolic thickening in viable and infarcted myocardium. AB - OBJECTIVES: In this study, we sought to compare the magnitude of changes in end diastolic wall thickness (WT(ed)) and postsystolic thickening (PST) in a swine model of stunning and reperfused acute myocardial infarction, and to explore the relationship between WT(ed) and PST. METHODS: Twenty-six pigs were subjected to left anterior descending coronary artery occlusion followed by reperfusion to induce stunning (n = 6), nontransmural (n = 8), or transmural (n = 12) myocardial infarction. Myocardial wall thickness was measured using intracardiac echocardiography. Transmural extent of necrosis (TEN) was quantified by triphenyltetrazolium chloride technique. RESULTS: During the first minutes of reperfusion, a marked increase in WT(ed) occurred in the myocardial walls with nontransmural and transmural infarct (42% and 102%, respectively) but less in those with stunning (19%). PST persisted at reperfusion in walls with stunning and nontransmural infarct (23% and 26%, respectively). In transmurally infarcted walls, PST progressively decreased either during occlusion (5/12 pigs) or shortly after reperfusion (7/12 pigs). PST at reperfusion was virtually absent when TEN was >70%. Both PST and the increase in WT(ed) at reperfusion correlated well with TEN (P <.0001 for both). Changes in PST at reperfusion were weakly correlated with changes in WT(ed). CONCLUSIONS: A marked increase in WT(ed) after reperfusion and absence of PST indicate transmural myocardial infarction. Presence of PST at reperfusion indicates viable tissue in more than 30% of wall thickness. The results suggest that amplitude of PST is modulated predominantely by factors related to the severity of ischemia and, to a smaller extent, by changes in wall thickness. PMID- 15282491 TI - Assessment of right ventricular function during exercise with quantitative Doppler tissue imaging in children late after repair of tetralogy of Fallot. AB - Doppler tissue imaging (DTI) has been developed to assess ventricular wall-motion velocity quantitatively for patients with various types of heart disease. This technique has a possibility of assessing right ventricular (RV) function reserve during exercise. To investigate RV function during exercise using DTI, 21 patients (9.3 +/- 3.3 years) who had undergone operation for tetralogy of Fallot at 1 to 3 years of age and 19 age-matched healthy children were studied. Echocardiography combined with DTI was performed at rest and during supine bicycle submaximal exercise. DTI of tricuspid annulus movement during systole (Sa) was obtained from a 4-chamber view. RV pressure was estimated by maximal tricuspid regurgitation (TR) velocity. The peak value of the first derivation of RV pressure (peak dP/dt) was measured from the continuous wave Doppler-derived TR profile. Adequate spectral Doppler recordings of TR were obtained in all participants. However, 9 healthy children and 2 patients with tetralogy of Fallot were excluded from the study because of an inability to determine the entire spectral TR velocity envelope during exercise. Therefore, data were analyzed in 29 participants. At rest, the mean RV pressure for patients was higher than that in control subjects (27 +/- 4 vs 18 +/- 3 mm Hg, P <.01). The mean Sa and RV peak dP/dt for patients were lower than those in control subjects (6.7 +/- 1.6 vs 8.8 +/- 1.7 cm/s and 464 +/- 77 vs 550 +/- 80 mm Hg/s, P <.01, respectively). Sa and RV peak dP/dt in the two groups increased significantly during exercise. However, the magnitude of increases in Sa and peak dP/dt was significantly less for patients than in control subjects (37 +/- 16 vs 66 +/- 19% and 42 +/- 10 vs 80 +/ 13%, P <.01, respectively). The magnitude of increase in Sa correlated with that in RV peak dP/dt (r = 0.84, P <.01). Results of DTI show high correlation with RV peak dP/dt during exercise. This technique has a potential as a useful indicator of the effect of exercise on RV systolic function. An insufficient increase in Sa suggests impaired response to exercise of RV in patients with tetralogy of Fallot. PMID- 15282492 TI - Real-time 3-dimensional echocardiography for quantification of the difference in left ventricular versus right ventricular stroke volume in a chronic animal model study: Improved results using C-scans for quantifying aortic regurgitation. AB - OBJECTIVE: The purpose of our study was to test the applicability of calculating the difference between left ventricular (LV) and right ventricular (RV) stroke volume (SV) for assessing the severity of aortic (Ao) regurgitation (AR) using a real-time 3-dimensional (3D) echocardiographic (RT3DE) imaging system. METHODS: The Ao valve was incised in 5 juvenile sheep, 6 to 10 weeks before the study, to produce AR (mean regurgitant fraction = 0.50). Simultaneous hemodynamic and RT3DE images were obtained on open-chest animals with Ao and pulmonary flows derived by Ao and pulmonary electromagnetic flowmeters balanced against each other. Four stages (baseline, volume loading, sodium nitroprusside, and angiotensin infusion) were used to produce a total of 16 different hemodynamic states. Epicardial scanning was done with a 2.5-MHz probe to sequentially record first the RV and then the LV cavities. Cavity volumes from the 3D echocardiography data were determined from angled sector planes (B-scans) and parallel cutting planes (C scans, which are planes perpendicular to the direction of the volume interrogation). AR volumes were determined from 3D images by computing and then subtracting RV SVs from LV SVs and then these were compared with electromagnetic flowmeter-derived SV and regurgitant volumes. RESULTS: There was close correlation between RV and LV SVs of the RT3DE and electromagnetic methods (C scans: LV, r = 0.98, standard error of the estimate [SEE] = 2.62 mL, P =.0001; RV, r = 0.89, SEE = 2.67 mL, P <.0001; and B-scans: LV, r = 0.95, SEE = 3.55 mL, P =.0001; RV, r = 0.77, SEE = 2.78 mL, P =.0003). Because of the small size of the RV in this model, the correlation was closer for C-scans than B-scans for RV SV. AR volume estimation also showed that C-scan (r = 0.93, SEE = 4.23 mL, P <.0001) had closer correlation than B-scan (r = 0.89, SEE = 4.87 mL, P <.0001). However, B-scan-derived AR fraction showed closer correlation than did C-scan (r = 0.82 vs r = 0.85, respectively). CONCLUSION: In this animal model, RT3DE imaging had the ability to reliably quantify both LV (B- and C-scans) and RV SVs and to assess the severity of AR. PMID- 15282494 TI - Ultrasonic tissue characterization of the mouse myocardium: successful in vivo cyclic variation measurements. AB - BACKGROUND: Measurements of the systematic variation of backscattered ultrasonic energy from myocardium during the heart cycle (cyclic variation) have been successfully used to characterize a wide spectrum of cardiac pathologies in large animal models and human subjects. The purpose of this study was to evaluate the feasibility of extending cyclic variation measurements to the study of genetically manipulated mouse models of cardiac diseases as a method for developing further insights into the disease-altered properties of the myocardium and its characterization with ultrasound. METHODS: Parasternal long-axis images of the heart were obtained in 9 wild-type mice under light anesthesia using a commercial imaging system with a 15-MHz nominal center frequency linear array. Images of a tissue-mimicking phantom and the mouse hearts were obtained for a series of specific receiver gains for each of a series of specific dynamic range settings. Analyses of these data formed the basis for gray-scale image calibration. Cyclic variation measurements were obtained by determining the average gray-scale value for a region of interest placed in the midmyocardium of the posterior wall for each frame acquired during 4 cardiac cycles and converting these mean gray-scale values to backscatter values expressed in decibels using the determined calibration. Results are expressed in terms of the magnitude and time delay of cyclic variation. To evaluate repeatability of these measurements the same group of mice underwent the identical imaging protocol 2 weeks after the first study. RESULTS: The mean magnitude of cyclic variation was found to be 4.6 +/- 0.2 dB with a corresponding normalized time delay of 1.02 +/- 0.03 for data averaged over all dynamic range settings. There was no significant difference among results obtained with each of the dynamic range settings. A comparison of these results with those from data acquired 2 weeks after the initial study showed no significant difference. CONCLUSION: This study represents the first reported measurement of cyclic variation in mice and demonstrates that reliable cyclic variation measurements can be obtained among individual animals and over different time points and, hence, forms the basis for subsequent investigations addressing specific cardiac pathologies and effects arising from myocardial anisotropy. PMID- 15282493 TI - Semiautomatic detection of left ventricular contours in contrast-enhanced echocardiographic images: Comparison with magnetic resonance imaging. AB - The aim of this study was to evaluate the accuracy of a semiautomatic contour detection method for left ventricular (LV) volume calculation in contrast enhanced echocardiographic images. In 26 patients, LV volumes were automatically measured by magnetic resonance imaging and second harmonic echocardiography after intravenous Levovist administration. LV cavity edges were manually drawn and semiautomatically outlined using the active contour algorithm, improved by a nonlinear anisotropic filter; LV volumes were calculated by the modified Simpson's rule. Manual and semiautomatic analysis of echocardiographic images lasted 45 +/- 6 and 20 +/- 8 seconds, respectively. Contrast echocardiography volumes were smaller than those by magnetic resonance imaging (mean difference: 16 mL for manual and 18 mL for automatic analysis). LV volumes by echocardiography closely related with those by magnetic resonance imaging using both manual (r = 0.955) and semiautomatic (r = 0.945) analysis; the correlation was closer for end-systolic than for end-diastolic volumes. In conclusion, this method provides a fast measure of LV volumes in contrast-enhanced images while reducing operator dependency. PMID- 15282495 TI - High-frequency ultrasound database profiling growth, development, and cardiovascular function in C57BL/6J mouse fetuses. AB - BACKGROUND: High-frequency ultrasound is effective for noninvasive phenotypic analysis of cardiovascular development and function in mutagenized fetal mice. However, lacking is a normative database of echocardiographic variables for monitoring growth and cardiovascular function. METHODS: C57BL/6J fetal mice were scanned in utero using an ultrasound system with a 15-MHz linear phased-array transducer. Pregnant mothers were anesthetized with 1% isoflurane mixed with 100% oxygen. Quantitative variables for monitoring fetal growth and cardiac function were obtained in several imaging planes and modalities. RESULTS: Fetal growth measurements increased linearly. Inflow velocities showed significant A-wave dominance. The E wave progressively increased during development. Cardiac function was best assessed through M-mode analysis, but short-axis images were difficult to obtain. Spectral Doppler was readily obtained and the myocardial performance index was calculated. CONCLUSIONS: These results provide an essential foundation for the evaluation of cardiovascular defects in mutagenized and transgenic mice. PMID- 15282496 TI - Spontaneous inversion of the left atrial appendage. AB - An interesting case of spontaneous inversion of the left atrial appendage was caused by an acute decrease in ventricular volume and the shortening of diastole during supraventricular tachycardia, creating a negative pressure invaginating the left atrial appendage. The return of normal in sinus rhythm and left-sided diastolic pressures reversed the left atrial appendage to a normal position, sparing the patient unnecessary surgery. PMID- 15282497 TI - An apparently immobile mitral valve prosthesis leaflet. PMID- 15282498 TI - Ruptured aneurysm of the left sinus of Valsalva discovered 41 years after a decelerational injury. AB - We present an unusual case of ruptured aneurysm of the left sinus of Valsalva discovered 41 years after a car accident. Echocardiography was a key element in establishing the diagnosis of this rare anomaly, which is often congenital but has also been reported as acquired or traumatic in origin. PMID- 15282499 TI - Infective endocarditis of the aortic valve complicated by massive pericardial effusion and rupture of a sinus of valsalva into the right atrium. AB - Infective endocarditis (IE) of the aortic valve is most commonly associated with perivalvular invasion and intracardiac fistula formation, which sometimes give rise to unpredictable clinical events. Massive pericardial effusion and aorticocardiac fistula are very rare complications of IE. We present a case in which IE of the aortic valve was initially associated with massive pericardial effusion requiring drainage and later complicated by rupture of a sinus of Valsalva with a fistula into the right atrium without formation of aneurysm. To our knowledge, the concurrent occurrence of these two events in a patient with IE has not been previously reported. PMID- 15282500 TI - Combination of acceleration and collision involving the left atrial appendage limbus as a mechanism of hemolytic anemia in the setting of periprosthetic mitral valve regurgitation. PMID- 15282501 TI - Giant thrombus trapped in foramen ovale with pulmonary embolus and stroke. AB - We describe the case of a young man who, while he was in coma because of a traffic accident, had first a pulmonary embolus and immediately afterwards had a systemic (cerebral) embolus. A transesophageal echocardiographic image revealed a giant thrombi trapped in foramen ovale protruding in right and left ventricles, diagnosing, thus, a paradoxical embolism. The relationship between patent foramen ovale and pulmonary embolism has been reported in some series. Elevated right chamber pressure caused by pulmonary hypertension could favor the establishment of a right-to-left shunt, causing, in some cases, paradoxical embolisms. We review the clinical implications. PMID- 15282502 TI - Assessment of mitral valve leaflet perforation as a result of infective endocarditis by 3-dimensional real-time echocardiography. PMID- 15282504 TI - A Kubler-Ross year. PMID- 15282505 TI - A 79-year-old man with an impalement injury of his face. PMID- 15282506 TI - A comparison of wait times and patients leaving without being seen when licensed nurses versus unlicensed assistive personnel perform triage. AB - INTRODUCTION: In today's health care environment, nurses are sometimes substituted with unlicensed assistive personnel (UAP), despite the lack of research into the possible negative effects. This study compared the wait time and the number of patients who left without being seen (LWBS) between triage systems that use nurses (including licensed practical nurses, nurses with an associate's degree, and nurses with a baccalaureate degree) versus UAP. METHODS: This study was a comparative descriptive retrospective chart review. The sample consisted of 2 parts: Part 1 involved charts for every fifth patient presenting to the emergency department during the second shift in April 1997 (N = 323) and April 1998 (N = 281). Part 2 included the monthly number of patients who LWBS between January 1997 and April 1998. Data were collected from department generated reports with use of investigator-designed forms. Specific time increments were calculated and compared between triage systems via 2-tailed t tests. The number of patients who LWBS was analyzed for trends. RESULTS: Wait time decreased by 73 minutes and the number of patients who LWBS decreased by 85% when nurses performed triage. DISCUSSION: Results of this study show that a triage system using nurses provided more services during triage and were associated with less wait time after triage and a decreased likelihood of patients LWBS compared with the triage system using UAP. PMID- 15282507 TI - Nonemergent ED patients referred to community resources after medical screening examination: characteristics, medical condition after 72 hours, and use of follow up services. AB - INTRODUCTION: Limited access to medical care has resulted in large numbers of patients seeking primary care for non-emergent emergency conditions in emergency departments. This influx of patients is contributing to overcrowding and delays in care for patients with emergencies. In response, a system was implemented in which persons with non-emergent medical conditions, following a medical screening examination, did not receive further ED assessment or treatment and instead were referred to community resources. The purpose of this study was to describe the characteristics of individuals who were referred to community-based services, their condition after 72 hours, and their use of follow-up health care services. METHODS: All referred cases (n=225) were reviewed for chief complaint and demographics. Phone contact was attempted after 72 hours to determine the person's condition and if community resources were utilized. RESULTS: Of the 225 cases, 52% were female, with a mean age of 33 years. Their most common chief complaints were extremity problems (16%), toothache (9%), and medication refill (8%). Follow-up phone contact was successful with 82 people (37%) an average of 14 days after their ED visit. The majority (55%) reported their condition had improved; 39%were unchanged, and 6% were worse. Thirty-one people(40%) accessed community resources and 8 (26%) returned to another emergency department. No clinically significant associations were found between patient demographics and use of community resources. DISCUSSION: Most people who were successfully contacted for follow-up and who had come to the emergency department with non emergent chief complaints did not access community resources and their condition frequently improved. Additional studies, with improved follow-up, are needed before suggesting that referring individuals to community resources is an acceptable practice. PMID- 15282508 TI - Complementary and alternative medicine use among urban ED patients: prevalence and patterns. AB - INTRODUCTION: High rates of complementary and alternative medicine (CAM) use are well documented in the general population without clear clinical benefits. Published studies examining prevalence and patterns of CAM use in emergency patients, however, are limited. The objectives of this study were to describe the prevalence and patterns of CAM use in urban ED patients. METHODS: This was a descriptive study of a convenience sample of 174 patients presenting to the emergency department of a level I, urban, Catholic, tertiary teaching center, with an annual ED census of 43,000. RESULTS: CAM use in our study group was high (47%). Although no sociodemographic predictors of CAM users were found, CAM users were more likely to have chronic conditions (P =.044). One third did not disclose CAM use. Prayer (28%), music therapy (11%), and meditation (10%) were the most frequently used types of CAM reported. DISCUSSION: Patients should be questioned routinely about CAM use, given the high rates of use and low disclosure rates. Knowledge of potential positive and negative effects of CAM, interactions with conventional treatments, and sensitivity toward patients' decisions to opt for CAM are imperative. Spiritual support, where available, should be considered for at least some ED patients. The 3 most common types of CAM reported by ED patients at our level I trauma center were prayer/spirituality, music therapy, and meditation. PMID- 15282509 TI - ED overcrowding: successful action plans of a Southern California community hospital. PMID- 15282510 TI - A nurse-initiated pain management advanced triage protocol for ED patients with an extremity injury at a level I trauma center. PMID- 15282511 TI - Improving ED patient satisfaction when triage nurses routinely communicate with patients as to reasons for waits: one rural hospital's experience. PMID- 15282513 TI - Encouraging the inclusion of children in grief after a sudden death: memory bags. PMID- 15282514 TI - ED brochure for recruitment of emergency nurses. PMID- 15282516 TI - Thrombolytic therapy: no room for error. PMID- 15282522 TI - A pocket guide to one emergency department: the 60-page OHSU ED At A Glance. PMID- 15282523 TI - Pediatric influenza and "universal respiratory etiquette": preparing and protecting your staff and the public. PMID- 15282524 TI - Dermal exposure to hydrofluoric acid causing significant systemic toxicity. PMID- 15282525 TI - California ENA joins coalition and ballot initiative to increase funding for emergency care in California. PMID- 15282526 TI - Living with nurse staffing ratios: early experiences. PMID- 15282527 TI - PDAs for the trauma nurse: help or hindrance? PMID- 15282528 TI - A 6-month-old with bilateral swollen, painful, and deformed hands. PMID- 15282529 TI - The Mortimer M. Bortin Lecture: to destroy by the reaction of immunity: the search for separation of graft-versus-leukemia and graft-versus-host. AB - The graft-versus-leukemia effect of allogeneic blood or marrow transplantation is a dramatic example of the power of the immune system to eradicate malignant disease. In this personal essay, adapted from the inaugural Mortimer M. Bortin Lecture presented at the 2004 Tandem BMT Meetings, the author recounts early efforts by Bortin and others to manipulate the graft-versus-leukemia effect and separate it from the potentially fatal complications of graft-versus-host disease. PMID- 15282530 TI - Augmentation of antitumor immune responses after adoptive transfer of bone marrow derived from donors immunized with tumor lysate-pulsed dendritic cells. AB - We demonstrated previously that tumor lysate-pulsed dendritic cells (TP-DC) could mediate a specific and long-lasting antitumor immune response against a weakly immunogenic breast tumor during early lymphoid reconstitution. The purpose of this study was to examine the potential therapeutic efficacy of bone marrow transplants from TP-DC-vaccinated donors. In 2 aggressive metastatic models, bone marrow transplantation with donor bone marrow cells from TP-DC-immunized mice mediated a tumor-specific immune response in the recipient, and this caused regressions of preexisting tumor metastases. After vaccination with TP-DC, donors harbored increased numbers of both activated CD4+ and CD8+ T-cell populations in the bone marrow. Adoptive transfer of T cells purified from the bone marrow of TP DC-vaccinated mice led to a reduction in preestablished lung metastases, whereas depletion of T cells from bone marrow abolished this effect. By using T cells derived from the bone marrow of TP-DC-vaccinated major histocompatibility complex class I and class II knockout mice, the effector cells required for the observed antitumor effect were determined to be major histocompatibility complex class I restricted CD8+ T cells. Additionally, the tumor burden in TP-DC-immunized transplant recipients could be reduced further by repetitive TP-DC immunizations after bone marrow transplantation. Collectively, these results demonstrate an important therapeutic role of bone marrow from TP-DC-immunized donors and raise the potential for this approach in patients with advanced cancer. PMID- 15282531 TI - Tumor regression by anti-CD40 and interleukin-2: role of CD40 in hematopoietic cells and organ-specific effects. AB - CD40 stimulation can synergize with interleukin (IL)-2 for antitumor responses against mouse metastatic renal cell carcinomas, with coincident increases in tumor-specific CD8+ T-cell responses and dendritic cell numbers in both the spleen and liver. Because CD40 is present on various hematopoietic-derived cells, endothelial cells, and some tumors themselves, this study was performed to determine whether the antitumor effects of CD40 stimulation and IL-2 were primarily mediated by CD40+ hematopoietic-derived cells. Bone marrow chimeras were created by reconstituting lethally irradiated CD40+/+ recipients with bone marrow from CD40-/- or CD40+/+ mice. Chimeric mice were then implanted orthotopically with renal cancer cells, followed by treatment with anti-CD40 agonist monoclonal antibodies and IL-2. Immune parameters of the spleen and liver were assessed after therapy and correlated with antitumor responses. The antitumor effects in the CD40-/- bone marrow transplantation chimeras were almost completely abrogated after treatment, and this shows that hematopoietically derived CD40+ cells are the principal targets for CD40 stimulation in this model. Although both spleen and liver showed reductions in CD8+ T-cell and dendritic cell expansion in the CD40-/- versus CD40+/+ chimeras after therapy, only the liver exhibited no significant increases in either CD8+ T cells or dendritic cells after treatment. CD40 cells on hematopoietic cells are the primary target for anti-CD40 and IL-2 therapy. The results also suggest that the immunologic events in the liver may be more revealing that those in lymphoid organs with regard to critical events related to responses after therapy. PMID- 15282532 TI - Donor antigen-presenting cells regulate T-cell expansion and antitumor activity after allogeneic bone marrow transplantation. AB - Delayed immunologic recovery after allogeneic bone marrow transplantation (BMT) represents a major cause of morbidity and mortality that limits the overall success of the transplantation procedure. Recent clinical data suggest that a subset of donor dendritic cells may inhibit the graft-versus-tumor activity of donor T cells. We studied the immunoregulatory activity of donor dendritic cells in allogeneic BMT between major histocompatibility complex-disparate strains of mice. Bone marrow grafts enriched or depleted of CD11b- and CD11b+ dendritic cell subsets by immunomagnetic cell sorting were combined with small numbers of congenic splenic T cells. Recipients of CD11b-depleted bone marrow had significant posttransplantation expansion of donor spleen-derived CD4+ memory T cells compared with recipients of unmanipulated bone marrow. CD11b depletion enhanced the antitumor activity of the splenic donor T cells without producing significant graft-versus-host disease and resulted in long-term survival after a supralethal dose of T-cell leukemia administered after BMT. Expansion of donor spleen-derived T cells was proportional to the number of CD11b- dendritic cells in the bone marrow graft and was associated with increased levels of serum interferon-gamma. Thus, manipulating the content of donor antigen-presenting cells in allogeneic BMT is a novel strategy to activate donor memory T cells and enhance allogeneic graft-versus-leukemia effects with minimal graft-versus-host disease. PMID- 15282533 TI - Evaluation of a CD25-specific immunotoxin for prevention of graft-versus-host disease after unrelated marrow transplantation. AB - Donor T cells activated by recipient alloantigens cause graft-versus-host disease (GVHD) after hematopoietic cell transplantation. Activated T cells express CD25, among other components of the interleukin-2 receptor. We conducted a phase I/II study to determine whether administration of CD25-specific antibody conjugated to ricin toxin A could reduce the risk of grade III or IV GVHD after marrow transplantation from HLA-matched unrelated donors. All patients received methotrexate and cyclosporine after the transplantation. The immunotoxin was given to 36 patients for 4 consecutive days beginning approximately 36 hours after the marrow infusion was completed. Fourteen (40%) of the 35 patients who could be evaluated developed grade III or IV GVHD. In a contemporaneous population of 121 patients who received marrow from HLA-matched unrelated donors and were given methotrexate and cyclosporine without the immunotoxin, the incidence of grades III and IV GVHD was 24%. Cyclosporine blocked the induction of CD25 expression on alloactivated T cells in vitro but had no detectable effect on CD25 expression by T-regulatory cells. Taken together, these results are consistent with the hypothesis that cyclosporine protected alloactivated donor T cells from the effects of the immunotoxin, whereas the CD25+ T-regulatory cells remained susceptible, causing an unexpected exacerbation of acute GVHD. PMID- 15282534 TI - Early central nervous system complications after reduced-intensity stem cell transplantation. AB - To investigate clinical characteristics of early central nervous system (CNS) complications after reduced-intensity stem cell transplantation (RIST), we reviewed the medical records of 232 patients who had undergone RIST for hematologic diseases at our institutions between September 1999 and June 2003. All patients had received purine analog-based preparative regimens. Stem cell sources comprised granulocyte colony-stimulating factor-mobilized blood from HLA identical or 1 locus-mismatched related donors (n = 151), unrelated bone marrow (n = 44), or unrelated cord blood (n = 37). Graft-versus-host disease prophylaxis incorporated cyclosporine with or without methotrexate. Diagnosis of CNS complications was based on clinical, radiologic, and microbiological findings. CNS complications occurred in 18 patients (7.8%), with a median onset of 22 days, and were infectious (n = 1), metabolic (n = 15), or cerebrovascular (n = 2). Symptoms included seizures (n = 7), visual disturbance (n = 2), headache (n = 8), nausea (n = 8), vomiting (n = 6), impaired consciousness (n = 16), and hemiparesis (n = 3). Complications improved promptly in 10 patients, and 8 patients died without improvement within 30 days. Multivariate analysis with logistic regression identified umbilical cord blood transplantation as a significant risk factor for early CNS complications (odds ratio, 14.5; 95% confidence interval, 3.7-56.9; P <.0001). CNS complications are a significant problem after RIST, particularly with umbilical cord blood. Limbic encephalopathy is an unrecognized subtype of neurotoxicity after umbilical cord blood transplantation. PMID- 15282535 TI - Adult recipients of umbilical cord blood transplants after nonmyeloablative preparative regimens. AB - We report the outcome of 13 patients with advanced malignancies who underwent nonmyeloablative conditioning therapy followed by infusion of partially matched unrelated cord blood cells. The median age of these patients was 49 years, and their median weight was 65.7 kg. The median nucleated cell dose infused was 2.07 x 10(7)/kg. Eight of the 13 patients demonstrated donor chimerism between 4 weeks and 6 months, and subsequent conversion to full donor chimerism was achieved in 5 patients. Three patients were alive and free of disease at 158 to 1054 days, with a median survival of 288 days after transplantation. The 100-day event-free survival is 69%, and overall survival is 77%. At 1 year, the event-free and overall survival was 43%. Treatment-related mortality observed within the first 100 days after transplantation was low: 1 previously extensively pretreated patient died of multiorgan failure. This result provides a basis for further exploring this potentially curative approach to selected patients who lack matched related or unrelated hematopoietic stem cell donors. PMID- 15282541 TI - Biological role and structural mechanism of twinfilin-capping protein interaction. AB - Twinfilin and capping protein (CP) are highly conserved actin-binding proteins that regulate cytoskeletal dynamics in organisms from yeast to mammals. Twinfilin binds actin monomer, while CP binds the barbed end of the actin filament. Remarkably, twinfilin and CP also bind directly to each other, but the mechanism and role of this interaction in actin dynamics are not defined. Here, we found that the binding of twinfilin to CP does not affect the binding of either protein to actin. Furthermore, site-directed mutagenesis studies revealed that the CP binding site resides in the conserved C-terminal tail region of twinfilin. The solution structure of the twinfilin-CP complex supports these conclusions. In vivo, twinfilin's binding to both CP and actin monomer was found to be necessary for twinfilin's role in actin assembly dynamics, based on genetic studies with mutants that have defined biochemical functions. Our results support a novel model for how sequential interactions between actin monomers, twinfilin, CP, and actin filaments promote cytoskeletal dynamics. PMID- 15282542 TI - ATR functions as a gene dosage-dependent tumor suppressor on a mismatch repair deficient background. AB - The ataxia-telangiectasia mutated and rad3-related (ATR) kinase orchestrates cellular responses to DNA damage and replication stress. Complete loss of ATR function leads to chromosomal instability and cell death. However, heterozygous ATR mutations are found in human cancers with microsatellite instability, suggesting that ATR haploinsufficiency contributes to tumorigenesis. To test this possibility, we generated human cell line and mouse model systems in which a single ATR allele was inactivated on a mismatch repair (MMR)-deficient background. Monoallelic ATR gene targeting in MLH1-deficient HCT 116 colon carcinoma cells resulted in hypersensitivity to genotoxic stress accompanied by dramatic increases in fragile site instability, and chromosomal amplifications and rearrangements. The ATR(+/-) HCT 116 cells also displayed compromised activation of Chk1, an important downstream target for ATR. In complementary studies, we demonstrated that mice bearing the same Atr(+/-)/Mlh1(-/-) genotype were highly prone to both embryonic lethality and early tumor development. These results demonstrate that MMR proteins and ATR functionally interact during the cellular response to genotoxic stress, and that ATR serves as a haploinsufficient tumor suppressor in MMR-deficient cells. PMID- 15282543 TI - MAD1 and c-MYC regulate UBF and rDNA transcription during granulocyte differentiation. AB - The regulation of cell mass (cell growth) is often tightly coupled to the cell division cycle (cell proliferation). Ribosome biogenesis and the control of rDNA transcription through RNA polymerase I are known to be critical determinants of cell growth. Here we show that granulocytic cells deficient in the c-MYC antagonist MAD1 display increased cell volume, rDNA transcription and protein synthesis. MAD1 repressed and c-MYC activated rDNA transcription in nuclear run on assays. Repression of rDNA transcription by MAD1 was associated with its ability to interact directly with the promoter of upstream binding factor (UBF), an rDNA regulatory factor. Conversely, c-MYC activated transcription from the UBF promoter. Using siRNA, UBF was shown to be required for c-MYC-induced rDNA transcription. These data demonstrate that MAD1 and c-MYC reciprocally regulate rDNA transcription, providing a mechanism for coordination of ribosome biogenesis and cell growth under conditions of sustained growth inhibition such as granulocyte differentiation. PMID- 15282544 TI - A complex network of RNA-RNA interactions controls subgenomic mRNA transcription in a tombusvirus. AB - Eukaryotic (+)-strand RNA viruses utilize a wide variety of gene expression strategies to achieve regulated production of their viral proteins. A common mechanism used by many is to transcribe viral subgenomic (sg) mRNAs. Transcription of sg mRNA2 in tombusviruses allows for expression of the p19 suppressor of gene silencing and p22 movement proteins. We have investigated the mechanism of transcription of this sg mRNA in Tomato bushy stunt virus and have determined that this process is facilitated by no less than three different RNA modules that are located throughout the viral genome. These RNA units perform distinct tasks and function via long-distance RNA-RNA interactions. Systematic deconstruction of the RNA network and analysis of related RNA promoter elements allowed us to identify fundamental properties necessary for productive sg mRNA2 transcription. Collectively, our results (i) establish specific roles for the different RNA components of a multipartite RNA-based control system, (ii) support a premature termination mechanism for tombusvirus sg mRNA transcription and (iii) reveal a close mechanistic relationship between sg mRNA transcription, viral RNA replication and RNA recombination. PMID- 15282545 TI - The response regulator 2 mediates ethylene signalling and hormone signal integration in Arabidopsis. AB - Hormones are important regulators of plant growth and development. In Arabidopsis, perception of the phytohormones ethylene and cytokinin is accomplished by a family of sensor histidine kinases including ethylene-resistant (ETR) 1 and cytokinin-response (CRE) 1. We identified the Arabidopsis response regulator 2 (ARR2) as a signalling component functioning downstream of ETR1 in ethylene signal transduction. Analyses of loss-of-function and ARR2 overexpressing lines as well as functional assays in protoplasts indicate an important role of ARR2 in mediating ethylene responses. Additional investigations indicate that an ETR1-initiated phosphorelay regulates the transcription factor activity of ARR2. This mechanism may create a novel signal transfer from endoplasmic reticulum-associated ETR1 to the nucleus for the regulation of ethylene-response genes. Furthermore, global expression profiling revealed a complex ARR2-involving two-component network that interferes with a multitude of different signalling pathways and thereby contributes to the highly integrated signal processing machinery in higher plants. PMID- 15282546 TI - Exportin 7 defines a novel general nuclear export pathway. AB - Most transport pathways between cell nucleus and cytoplasm are mediated by nuclear transport receptors of the importin beta family. These receptors are in continuous circulation between the two compartments and transfer cargo molecules from one side of the nuclear envelope to the other. RanBP16 is a family member from higher eukaryotes of so far unknown function. We now show that it exports p50RhoGAP from the nucleus and thereby confines this activity to the cytoplasm. It also accounts for nuclear exclusion of 14-3-3sigma, which in turn is known to anchor, for example, cyclin-dependent kinases in the cytoplasm. Our data further suggest that RanBP16 exports several additional cargoes. It thus appears to be a nuclear export mediator with broad substrate specificity and we will therefore refer to it as exportin 7 (Exp7). Finally, we demonstrate that Exp7-dependent nuclear export signals differ fundamentally from the leucine-rich, CRM1-dependent ones: First, they are not just short linear sequences, but instead include folded motifs. Second, basic residues are critical for Exp7 recruitment. PMID- 15282547 TI - MicroRNA control of PHABULOSA in leaf development: importance of pairing to the microRNA 5' region. AB - MicroRNAs (miRNAs) are approximately 22-nucleotide noncoding RNAs that can regulate gene expression by directing mRNA degradation or inhibiting productive translation. Dominant mutations in PHABULOSA (PHB) and PHAVOLUTA (PHV) map to a miR165/166 complementary site and impair miRNA-guided cleavage of these mRNAs in vitro. Here, we confirm that disrupted miRNA pairing, not changes in PHB protein sequence, causes the developmental defects in phb-d mutants. In planta, disrupting miRNA pairing near the center of the miRNA complementary site had far milder developmental consequences than more distal mismatches. These differences correlated with differences in miRNA-directed cleavage efficiency in vitro, where mismatch scanning revealed more tolerance for mismatches at the center and 3' end of the miRNA compared to mismatches to the miRNA 5' region. In this respect, miR165/166 resembles animal miRNAs in its pairing requirements. Pairing to the 5' portion of the small silencing RNA appears crucial regardless of the mode of post transcriptional repression or whether it occurs in plants or animals, supporting a model in which this region of the silencing RNA nucleates pairing to its target. PMID- 15282548 TI - Visualization of RNA-protein interactions in living cells: FMRP and IMP1 interact on mRNAs. AB - Protein expression depends significantly on the stability, translation efficiency and localization of mRNA. These qualities are largely dictated by the RNA-binding proteins associated with an mRNA. Here, we report a method to visualize and localize RNA-protein interactions in living mammalian cells. Using this method, we found that the fragile X mental retardation protein (FMRP) isoform 18 and the human zipcode-binding protein 1 ortholog IMP1, an RNA transport factor, were present on common mRNAs. These interactions occurred predominantly in the cytoplasm, in granular structures. In addition, FMRP and IMP1 interacted independently of RNA. Tethering of FMRP to an mRNA caused IMP1 to be recruited to the same mRNA and resulted in granule formation. The intimate association of FMRP and IMP1 suggests a link between mRNA transport and translational repression in mammalian cells. PMID- 15282549 TI - LRIG1 restricts growth factor signaling by enhancing receptor ubiquitylation and degradation. AB - Kekkon proteins negatively regulate the epidermal growth factor receptor (EGFR) during oogenesis in Drosophila. Their structural relative in mammals, LRIG1, is a transmembrane protein whose inactivation in rodents promotes skin hyperplasia, suggesting involvement in EGFR regulation. We report upregulation of LRIG1 transcript and protein upon EGF stimulation, and physical association of the encoded protein with the four EGFR orthologs of mammals. Upregulation of LRIG1 is followed by enhanced ubiquitylation and degradation of EGFR. The underlying mechanism involves recruitment of c-Cbl, an E3 ubiquitin ligase that simultaneously ubiquitylates EGFR and LRIG1 and sorts them for degradation. We conclude that LRIG1 evolved in mammals as a feedback negative attenuator of signaling by receptor tyrosine kinases. PMID- 15282550 TI - True reversal of Mu integration. AB - We describe a high-temperature (75 degrees C) transition in the Mu integration complex that causes efficient and true reversal of the integration reaction. A second reversal pathway, first described as 'foldback' reversal for the HIV integrase, was also observed upon disassembly/reassembly of the Mu complex at normal temperatures. Both true and foldback reversal severed only one or the other of the two integrated Mu ends, and each exhibited distinct metal ion specificities. Our results directly implicate an altered transposase configuration in the Mu strand transfer complex that inhibits reversal, thereby regulating the directionality of transposition. PMID- 15282553 TI - Sequencing is believing? PMID- 15282552 TI - Pax7 directs postnatal renewal and propagation of myogenic satellite cells but not their specification. AB - The paired-box transcription factor Pax7 has been claimed to specify the muscle stem cell lineage since inactivation of Pax7 led to a failure to detect muscle satellite cells. Here we show that muscles of juvenile Pax7(-/-) mice at P11 contain a reduced but substantial number of satellite cells. Neither juvenile nor adult Pax7(-/-) mice displayed a significant reduction in the number and size of myotubes, indicating that the remaining number of satellite cells sufficed to allow normal postnatal muscle growth. The number of satellite cells in Pax7 mutant mice declined strongly during postnatal development, although single satellite cells were readily identified in adult Pax7 mutant mice. Muscle regeneration was impaired in adult Pax7 mutant mice. Our results clearly indicate an essential function of Pax7 for renewal and maintenance of muscle stem cells and exclude an exclusive role of Pax7 in satellite cell specification. PMID- 15282554 TI - Immunology needs the mind. PMID- 15282551 TI - PKN3 is required for malignant prostate cell growth downstream of activated PI 3 kinase. AB - Chronic activation of the phosphoinositide 3-kinase (PI3K)/PTEN signal transduction pathway contributes to metastatic cell growth, but up to now effectors mediating this response are poorly defined. By simulating chronic activation of PI3K signaling experimentally, combined with three-dimensional (3D) culture conditions and gene expression profiling, we aimed to identify novel effectors that contribute to malignant cell growth. Using this approach we identified and validated PKN3, a barely characterized protein kinase C-related molecule, as a novel effector mediating malignant cell growth downstream of activated PI3K. PKN3 is required for invasive prostate cell growth as assessed by 3D cell culture assays and in an orthotopic mouse tumor model by inducible expression of short hairpin RNA (shRNA). We demonstrate that PKN3 is regulated by PI3K at both the expression level and the catalytic activity level. Therefore, PKN3 might represent a preferred target for therapeutic intervention in cancers that lack tumor suppressor PTEN function or depend on chronic activation of PI3K. PMID- 15282555 TI - Working with dangerous bugs. AB - As part of the national effort in the US to protect civilians from bioterrorist attacks, the US National Institute of Allergy and Infectious Diseases (NIAID) was charged with the development of diverse research resources. The NIAID Resources for Biodefense Research program is forging new collaborations between immunologists and infectious disease experts and is reinvigorating research in the general area of immune protection against pathogenic infection. PMID- 15282556 TI - Bioinformatics for the 'bench biologist': how to find regulatory regions in genomic DNA. AB - The combination of bioinformatic and biological approaches constitutes a powerful method for identifying gene regulatory elements. High-quality genome sequences are available in public databases for several vertebrate species. Comparative cross-species sequence analysis of these genomes shows considerable conservation of noncoding sequences in DNA. Biological analyses show that an unexpectedly high number of the conserved sequences correspond to functional cis-regulatory regions that influence gene transcription. Because research biologists are often unfamiliar with the bioinformatic resources at their disposal, this commentary discusses how to integrate biological and bioinformatic methods in the discovery of gene regulatory regions and includes a tutorial on widely available comparative genomics programs. PMID- 15282557 TI - Paradise lost and paradigm found. PMID- 15282558 TI - How NOD-ing off leads to Crohn disease. PMID- 15282559 TI - Mutual attraction: does it benefit the host or the bug? PMID- 15282560 TI - Self help for T cells. PMID- 15282563 TI - Tribal culture versus genetics. PMID- 15282562 TI - Protein kinase C and beyond. AB - Protein kinase C molecules regulate both positive and negative signal transduction pathways essential for the initiation and homeostasis of immune responses. There are multiple isoforms of protein kinase C that are activated differently by calcium and diacylglycerol, and these are activated mainly by antigen receptors in T cells, B cells and mast cells. Additionally, mammals express several other diacylglycerol binding proteins that are linked to a network of key signal transduction pathways that control lymphocyte biology. Diacylglycerol and protein kinase C regulate a broad range of gene transcription programs but also modulate integrins, chemokine responses and antigen receptors, thereby regulating lymphocyte adhesion, migration, differentiation and proliferation. PMID- 15282564 TI - States versus gases. PMID- 15282566 TI - Energy labs halt classified research amid security fears. PMID- 15282565 TI - Lawyers blast nuclear pact as a breach of disarmament treaty. PMID- 15282567 TI - Joint suits aim to weed out agencies' red tape. PMID- 15282568 TI - Sea snapshots will map frequency of freak waves. PMID- 15282570 TI - Swedish enthusiasm peps up plans for neutron source. PMID- 15282569 TI - Dinosaur eggs escape sale as smuggling claims unearthed. PMID- 15282571 TI - Winged messenger set to follow ancient mariner to Mercury. PMID- 15282572 TI - Russian bid to drill Antarctic lake gets chilly response. PMID- 15282573 TI - Tough talker quits Congress for bioindustry. PMID- 15282576 TI - What's in a name? PMID- 15282574 TI - Biologists lobby China's government for funding reform. PMID- 15282578 TI - If you can lose a driving licence, why not a PhD? PMID- 15282577 TI - When two tribes go to war. PMID- 15282579 TI - Species problem solved 100 years ago. PMID- 15282586 TI - Why can't planets be like stars? PMID- 15282580 TI - Tight budget should fund benefits, not more posts. PMID- 15282587 TI - Neurobiology: odorant receptors make scents. PMID- 15282588 TI - Superconductivity: why the temperature is high. PMID- 15282589 TI - Cell biology: how to build a cell junction. PMID- 15282590 TI - Evolutionary biology: oceans of bacteria. PMID- 15282591 TI - Nuclear physics: not-so-magic numbers. PMID- 15282592 TI - Self-assembly: towards precision micelles. PMID- 15282594 TI - Molecular biology: cohesins slip sliding away. PMID- 15282596 TI - Animal communication: ground squirrel uses ultrasonic alarms. AB - Apart from echolocation and the pursuit of prey by bats, the function of ultrasound in animal communication is poorly understood. This is mainly because of the broad range of responses that it can evoke and the widely varied contexts in which it is used (for example, in rodents of the Muridae family it may indicate distress in infants or a sexual or predatory encounter in adults). Here we find that a purely ultrasonic signal is produced in the wild by a rodent of the Sciuridae family, Richardson's ground squirrel, and show that its function is to warn conspecifics of impending danger. To our knowledge, ultrasonic alarm calls have not previously been detected in any animal group, despite their twin advantages of being highly directional and inaudible to key predators. PMID- 15282597 TI - The nonlinear nature of friction. AB - Tribology is the study of adhesion, friction, lubrication and wear of surfaces in relative motion. It remains as important today as it was in ancient times, arising in the fields of physics, chemistry, geology, biology and engineering. The more we learn about tribology the more complex it appears. Nevertheless, recent experiments coupled to theoretical modelling have made great advances in unifying apparently diverse phenomena and revealed many subtle and often non intuitive aspects of matter in motion, which stem from the nonlinear nature of the problem. PMID- 15282598 TI - Solar chromospheric spicules from the leakage of photospheric oscillations and flows. AB - Spicules are dynamic jets propelled upwards (at speeds of approximately 20 km s( 1)) from the solar 'surface' (photosphere) into the magnetized low atmosphere of the Sun. They carry a mass flux of 100 times that of the solar wind into the low solar corona. With diameters close to observational limits (< 500 km), spicules have been largely unexplained since their discovery in 1877: none of the existing models can account simultaneously for their ubiquity, evolution, energetics and recently discovered periodicity. Here we report a synthesis of modelling and high spatial-resolution observations in which numerical simulations driven by observed photospheric velocities directly reproduce the observed occurrence and properties of individual spicules. Photospheric velocities are dominated by convective granulation (which has been considered before for spicule formation) and by p modes (which are solar global resonant acoustic oscillations visible in the photosphere as quasi-sinusoidal velocity and intensity pulsations). We show that the previously ignored p-modes are crucial: on inclined magnetic flux tubes, the p-modes leak sufficient energy from the global resonant cavity into the chromosphere to power shocks that drive upward flows and form spicules. PMID- 15282599 TI - A universal scaling relation in high-temperature superconductors. AB - Since the discovery of superconductivity at elevated temperatures in the copper oxide materials there has been a considerable effort to find universal trends and correlations amongst physical quantities, as a clue to the origin of the superconductivity. One of the earliest patterns that emerged was the linear scaling of the superfluid density (rho(s)) with the superconducting transition temperature (T(c)), which marks the onset of phase coherence. This is referred to as the Uemura relation, and it works reasonably well for the underdoped materials. It does not, however, describe optimally doped (where T(c) is a maximum) or overdoped materials. Similarly, an attempt to scale the superfluid density with the d.c. conductivity (sigma(dc)) was only partially successful. Here we report a simple scaling relation (rho(s) proportional, variant sigma(dc)T(c), with sigma(dc) measured at approximately T(c)) that holds for all tested high-T(c) materials. It holds regardless of doping level, nature of dopant (electrons versus holes), crystal structure and type of disorder, and direction (parallel or perpendicular to the copper-oxygen planes). PMID- 15282600 TI - Magnetic phase control by an electric field. AB - The quest for higher data density in information storage is motivating investigations into approaches for manipulating magnetization by means other than magnetic fields. This is evidenced by the recent boom in magnetoelectronics and 'spintronics', where phenomena such as carrier effects in magnetic semiconductors and high-correlation effects in colossal magnetoresistive compounds are studied for their device potential. The linear magnetoelectric effect-the induction of polarization by a magnetic field and of magnetization by an electric field provides another route for linking magnetic and electric properties. It was recently discovered that composite materials and magnetic ferroelectrics exhibit magnetoelectric effects that exceed previously known effects by orders of magnitude, with the potential to trigger magnetic or electric phase transitions. Here we report a system whose magnetic phase can be controlled by an external electric field: ferromagnetic ordering in hexagonal HoMnO3 is reversibly switched on and off by the applied field via magnetoelectric interactions. We monitor this process using magneto-optical techniques and reveal its microscopic origin by neutron and X-ray diffraction. From our results, we identify basic requirements for other candidate materials to exhibit magnetoelectric phase control. PMID- 15282601 TI - Demixing in simple fluids induced by electric field gradients. AB - Phase separation in liquid mixtures is mainly controlled by temperature and pressure, but can also be influenced by gravitational, magnetic or electric fields. However, the weak coupling between such fields and concentration fluctuations limits this effect to extreme conditions. For example, mixing induced by uniform electric fields is detectable only at temperatures that are within a few hundredths of degree or less of the phase transition temperature of the system being studied. Here we predict and demonstrate that electric fields can control the phase separation behaviour of mixtures of simple liquids under more practical conditions, provided that the fields are non-uniform. By applying a voltage of 100 V across unevenly spaced electrodes about 50 micro m apart, we can reversibly induce the demixing of paraffin and silicone oil at 1 K above the phase transition temperature of the mixture; when the field gradients are turned off, the mixture becomes homogeneous again. This direct control over phase separation behaviour depends on field intensity, with the electrode geometry determining the length-scale of the effect. We expect that this phenomenon will find a number of nanotechnological applications, particularly as it benefits from field gradients near small conducting objects. PMID- 15282602 TI - Evidence of power-law flow in the Mojave desert mantle. AB - Studies of the Earth's response to large earthquakes can be viewed as large rock deformation experiments in which sudden stress changes induce viscous flow in the lower crust and upper mantle that lead to observable postseismic surface deformation. Laboratory experiments suggest that viscous flow of deforming hot lithospheric rocks is characterized by a power law in which strain rate is proportional to stress raised to a power, n (refs 2, 3). Most geodynamic models of flow in the lower crust and upper mantle, however, resort to newtonian (linear) stress-strain rate relations. Here we show that a power-law model of viscous flow in the mantle with n = 3.5 successfully explains the spatial and temporal evolution of transient surface deformation following the 1992 Landers and 1999 Hector Mine earthquakes in southern California. A power-law rheology implies that viscosity varies spatially with stress causing localization of strain, and varies temporally as stress evolves, rendering newtonian models untenable. Our findings are consistent with laboratory-derived flow law parameters for hot and wet olivine--the most abundant mineral in the upper mantle -and support the contention that, at least beneath the Mojave desert, the upper mantle is weaker than the lower crust. PMID- 15282604 TI - Cambrian origins and affinities of an enigmatic fossil group of arthropods. AB - Euthycarcinoids are one of the most enigmatic arthropod groups, having been assigned to nearly all major clades of Arthropoda. Recent work has endorsed closest relationships with crustaceans or a myriapod-hexapod assemblage, a basal position in the Euarthropoda, or a placement in the Hexapoda or hexapod stem group. Euthycarcinoids are known from 13 species ranging in age from Late Ordovician or Early Silurian to Middle Triassic, all in freshwater or brackish water environments. Here we describe a euthycarcinoid from marine strata in Argentina dating from the latest Cambrian period, extending the group's record back as much as 50 million years. Despite its antiquity and marine occurrence, the Cambrian species demonstrates that morphological details were conserved in the transition to fresh water. Trackways in the same unit as the euthycarcinoid strengthen arguments that similar traces of subaerial origin from Cambro Ordovician rocks were made by euthycarcinoids. Large mandibles in euthycarcinoids are confirmed by the Cambrian species. A morphology-based phylogeny resolves euthycarcinoids as stem-group Mandibulata, sister to the Myriapoda and Crustacea plus Hexapoda. PMID- 15282605 TI - Social parasitism by male-producing reproductive workers in a eusocial insect. AB - The evolution of extreme cooperation, as found in eusocial insects (those with a worker caste), is potentially undermined by selfish reproduction among group members. In some eusocial Hymenoptera (ants, bees and wasps), workers can produce male offspring from unfertilized eggs. Kin selection theory predicts levels of worker reproduction as a function of the relatedness structure of the workers' natal colony and the colony-level costs of worker reproduction. However, the theory has been only partially successful in explaining levels of worker reproduction. Here we show that workers of a eusocial bumble bee (Bombus terrestris) enter unrelated, conspecific colonies in which they then produce adult male offspring, and that such socially parasitic workers reproduce earlier and are significantly more reproductive and aggressive than resident workers that reproduce within their own colonies. Explaining levels of worker reproduction, and hence the potential of worker selfishness to undermine the evolution of cooperation, will therefore require more than simply a consideration of the kin selected interests of resident workers. It will also require knowledge of the full set of reproductive options available to workers, including intraspecific social parasitism. PMID- 15282603 TI - Fine-scale phylogenetic architecture of a complex bacterial community. AB - Although molecular data have revealed the vast scope of microbial diversity, two fundamental questions remain unanswered even for well-defined natural microbial communities: how many bacterial types co-exist, and are such types naturally organized into phylogenetically discrete units of potential ecological significance? It has been argued that without such information, the environmental function, population biology and biogeography of microorganisms cannot be rigorously explored. Here we address these questions by comprehensive sampling of two large 16S ribosomal RNA clone libraries from a coastal bacterioplankton community. We show that compensation for artefacts generated by common library construction techniques reveals fine-scale patterns of community composition. At least 516 ribotypes (unique rRNA sequences) were detected in the sample and, by statistical extrapolation, at least 1,633 co-existing ribotypes in the sampled population. More than 50% of the ribotypes fall into discrete clusters containing less than 1% sequence divergence. This pattern cannot be accounted for by interoperon variation, indicating a large predominance of closely related taxa in this community. We propose that such microdiverse clusters arise by selective sweeps and persist because competitive mechanisms are too weak to purge diversity from within them. PMID- 15282606 TI - Neurons compute internal models of the physical laws of motion. AB - A critical step in self-motion perception and spatial awareness is the integration of motion cues from multiple sensory organs that individually do not provide an accurate representation of the physical world. One of the best-studied sensory ambiguities is found in visual processing, and arises because of the inherent uncertainty in detecting the motion direction of an untextured contour moving within a small aperture. A similar sensory ambiguity arises in identifying the actual motion associated with linear accelerations sensed by the otolith organs in the inner ear. These internal linear accelerometers respond identically during translational motion (for example, running forward) and gravitational accelerations experienced as we reorient the head relative to gravity (that is, head tilt). Using new stimulus combinations, we identify here cerebellar and brainstem motion-sensitive neurons that compute a solution to the inertial motion detection problem. We show that the firing rates of these populations of neurons reflect the computations necessary to construct an internal model representation of the physical equations of motion. PMID- 15282607 TI - Median bundle neurons coordinate behaviours during Drosophila male courtship. AB - Throughout the animal kingdom the innate nature of basic behaviour routines suggests that the underlying neuronal substrates necessary for their execution are genetically determined and developmentally programmed. Complex innate behaviours require proper timing and ordering of individual component behaviours. In Drosophila melanogaster, analyses of combinations of mutations of the fruitless (fru) gene have shown that male-specific isoforms (Fru(M)) of the Fru transcription factor are necessary for proper execution of all steps of the innate courtship ritual. Here, we eliminate Fru(M) expression in one group of about 60 neurons in the Drosophila central nervous system and observe severely contracted courtship behaviour, including rapid courtship initiation, absence of orienting and tapping, and the simultaneous occurrence of wing vibration, licking and attempted copulation. Our results identify a small group of median bundle neurons, that in wild-type Drosophila appropriately trigger the sequential execution of the component behaviours that constitute the Drosophila courtship ritual. PMID- 15282608 TI - Structural determinants for generating centromeric chromatin. AB - Mammalian centromeres are not defined by a consensus DNA sequence. In all eukaryotes a hallmark of functional centromeres--both normal ones and those formed aberrantly at atypical loci--is the accumulation of centromere protein A (CENP-A), a histone variant that replaces H3 in centromeric nucleosomes. Here we show using deuterium exchange/mass spectrometry coupled with hydrodynamic measures that CENP-A and histone H4 form sub-nucleosomal tetramers that are more compact and conformationally more rigid than the corresponding tetramers of histones H3 and H4. Substitution into histone H3 of the domain of CENP-A responsible for compaction is sufficient to direct it to centromeres. Thus, the centromere-targeting domain of CENP-A confers a unique structural rigidity to the nucleosomes into which it assembles, and is likely to have a role in maintaining centromere identity. PMID- 15282609 TI - Low-populated folding intermediates of Fyn SH3 characterized by relaxation dispersion NMR. AB - Many biochemical processes proceed through the formation of functionally significant intermediates. Although the identification and characterization of such species can provide vital clues about the mechanisms of the reactions involved, it is challenging to obtain information of this type in cases where the intermediates are transient or present only at low population. One important example of such a situation involves the folding behaviour of small proteins that represents a model for the acquisition of functional structure in biology. Here we use relaxation dispersion nuclear magnetic resonance (NMR) spectroscopy to identify, for two mutational variants of one such protein, the SH3 domain from Fyn tyrosine kinase, a low-population folding intermediate in equilibrium with its unfolded and fully folded states. By performing the NMR experiments at different temperatures, this approach has enabled characterization of the kinetics and energetics of the folding process as well as providing structures of the intermediates. A general strategy emerges for an experimental determination of the energy landscape of a protein by applying this methodology to a series of mutants whose intermediates have differing degrees of native-like structure. PMID- 15282610 TI - Up for review. PMID- 15282612 TI - Nuts and bolts. Seeking feedback. PMID- 15282615 TI - Epidural anesthesia. PMID- 15282614 TI - Cell cycle regulation of central spindle assembly. AB - The bipolar mitotic spindle is responsible for segregating sister chromatids at anaphase. Microtubule motor proteins generate spindle bipolarity and enable the spindle to perform mechanical work. A major change in spindle architecture occurs at anaphase onset when central spindle assembly begins. This structure regulates the initiation of cytokinesis and is essential for its completion. Central spindle assembly requires the centralspindlin complex composed of the Caenorhabditis elegans ZEN-4 (mammalian orthologue MKLP1) kinesin-like protein and the Rho family GAP CYK-4 (MgcRacGAP). Here we describe a regulatory mechanism that controls the timing of central spindle assembly. The mitotic kinase Cdk1/cyclin B phosphorylates the motor domain of ZEN-4 on a conserved site within a basic amino-terminal extension characteristic of the MKLP1 subfamily. Phosphorylation by Cdk1 diminishes the motor activity of ZEN-4 by reducing its affinity for microtubules. Preventing Cdk1 phosphorylation of ZEN-4/MKLP1 causes enhanced metaphase spindle localization and defects in chromosome segregation. Thus, phosphoregulation of the motor domain of MKLP1 kinesin ensures that central spindle assembly occurs at the appropriate time in the cell cycle and maintains genomic stability. PMID- 15282616 TI - Acinetobacter skin abscess in a neonate. AB - Although Acinetobacter is usually a species of low virulence, it is becoming increasingly more important as a cause of hospital outbreaks, particularly on intensive care units. Antibiotic resistance can develop rapidly. This organism has not been reported to cause skin abscesses previously. We describe a case of a neonate who developed an Acinetobacter abscess on our neonatal intensive care unit. PMID- 15282617 TI - A neonate with severe thrombocytopenia and radio-ulnar synostosis. AB - Bone marrow failure syndromes can be associated with abnormalities of the forearms. We observed a neonate with congenital thrombocytopenia who had bilateral radio-ulnar synostosis and fifth finger clinodactly. We performed an evaluation of the mechanism causing the thrombocytopenia using a combination of direct and indirect measures of thrombopoiesis. These tests indicated decreased platelet production. This entity of congenital hyporegenerative thrombocytopenia with bilateral radio-ulnar synostosis and fifth-finger clinodactly is an uncommon but easily recognizable form of congenital amegakaryocytic thrombocytopenia (CAMT). This entity can be distinguished from the TAR syndrome (thrombocytopenia and absent radii) by the distinctive orthopedic issues, different underlying genetic mutations, and a more worrisome prognosis for CAMT than for TAR. PMID- 15282618 TI - Dialysis in the elderly. PMID- 15282619 TI - Physicians' negative views of direct-to-consumer advertising (DTCA): the international evidence grows. PMID- 15282620 TI - Interest and participation in selected sports among New Zealand adolescents. AB - AIM: The current study aims to describe participation and interest in 18 selected popular sports among a large sample of New Zealand adolescents with the goal of identifying opportunities for increasing youth participation in physical activity. METHODS: Multi-stage cluster sampling was used to select 82 secondary schools and classes of Year 10 and 12 students within schools from six geographical regions throughout New Zealand. Students completed self-administered written questionnaires. RESULTS: The school response rate was 58%, with physical activity data available for 1730 females and 1704 males, including 637 students who self-identified as Maori. The greatest gaps between expressed interest and actual participation were reported for rugby union, rugby league, basketball, soccer and surfing (among both sexes); dance and volleyball (among females); and skateboarding (among males). When diversity of participation was modelled, increased diversity was associated with being male, having an income from part time work, and having greater diversity of interest in the selected sports. CONCLUSION: For several sports there are substantial groups of young people who express interest but do not participate. The challenge for public health is to turn this interest into increased participation in health-promoting physical activity--by providing appropriate opportunities and support for participation. PMID- 15282621 TI - Finnish physicians show little support for consumer advertising of prescription drugs. AB - AIM: To study Finnish physicians' opinions of direct-to-consumer advertising of prescription drugs (DTCA) and other drug information sources for patients. METHOD: A survey was sent to all working-aged physicians (n=16,698; response rate 85%). RESULTS: Almost all physicians were against full DTCA, but half would allow advertising indirectly via patients organisations or healthcare units; 18% were against all types of DTCA. Thirty-six percent generally considered drug advertisements to patients and consumers to be harmful or useless. CONCLUSION: Further discussion of DTCA and other means of disseminating drug information are needed. PMID- 15282622 TI - Factors associated with not breastfeeding exclusively among mothers of a cohort of Pacific infants in New Zealand. AB - AIMS: This study investigated the association between not breastfeeding exclusively (among mothers of a cohort of Pacific infants in New Zealand) and several maternal, sociodemographic, and infant care factors. METHODS: The data were gathered as part of the Pacific Islands Families (PIF) Study. Infant feeding information was obtained through interviews with mothers (6 weeks post-birth) and from hospital records for 1247 of the 1365 biological mothers. RESULTS: Factors significantly associated with not exclusively breastfeeding at hospital discharge included smoking, unemployment prior to pregnancy, years in New Zealand, not seeing a midwife during pregnancy, caesarean delivery, and twin birth status. Factors significantly associated with cessation (before 6 weeks post-birth) of exclusive breastfeeding (for mothers who initially breastfed exclusively) included smoking, employment prior to pregnancy, being in current employment, high parity, dummy use, not receiving a visit from Plunket, infant not discharged at the same time as the mother, infant not sharing the same room as the parent(s) at night, regular childcare, and having a home visit for the infant from a traditional healer. CONCLUSIONS: Aside from smoking, different factors were associated with initiation and maintenance of exclusive breastfeeding. Identification of risk factors should assist targeting women who are at heightened risk of not breastfeeding exclusively. PMID- 15282623 TI - Comparison of Maori and non-Maori maternal and fetal iron parameters. AB - AIMS: To investigate the effect of maternal iron stores on the fetus in Maori and non-Maori neonates. METHODS: Paired samples of maternal venous and fetal cord blood were compared for haemoglobin, iron, and ferritin. Women were included who had no medical complications and were delivering by elective caesarian section at Hastings Memorial Hospital. RESULTS: The study involved 124 participants, of whom 31 were Maori. The mothers in our study had normal iron status or were mildly-to moderately anaemic. Maori mothers had significantly lower haemoglobin levels compared to non-Maori; however there was no significant difference in maternal levels of iron or ferritin. Cord blood parameters for Maori neonates were not different for haemoglobin or iron, however ferritin was significantly lower. When Maori and non-Maori were analysed together, no statistical relationship was found between maternal and fetal cord blood for haemoglobin, iron, and ferritin levels. CONCLUSIONS: Our study suggested that, when analysing our study population of mothers with normal iron status or mild-to-moderate anaemia, iron stores in the fetus were not adversely affected by maternal haemoglobin, ferritin, or iron levels. However, separating for ethnicity, Maori mothers had significantly lower serum haemoglobin values than non-Maori. Furthermore, Maori neonates had significantly lower cord ferritin levels than non-Maori. It is possible that the lower ferritin values seen in Maori neonates compared to non Maori may ultimately contribute to higher rates of anaemia in these infants. PMID- 15282624 TI - The epidemic of elderly patients with dialysis-requiring end-stage renal disease in New Zealand. PMID- 15282625 TI - Ethnicity and body fatness in New Zealanders. AB - In light of alarming rises in the prevalence of obesity worldwide, tackling the obesity 'epidemic' is now a national health priority in many countries. Increasingly, population measures that provide accurate estimates of body fatness are required to assist public health organisations in identifying at-risk groups and developing appropriate preventative strategies. Body mass index (BMI) remains the most cost-effective and practical tool in this regard. The World Health Organization (WHO) has issued universal BMI standards for defining 'overweight' and 'obesity' in adults (BMI > or =25 kg x m(-2), and > or =30 kg x m(-2), respectively) based on the risk of obesity-related disease in Europeans. Although widely used, there is mounting evidence suggesting that these standards are not appropriate for all populations. Research indicates that the associations between BMI, percent body fat (%BF), and health risks can vary across different ethnicities. Accordingly, ethnic-specific and country-specific BMI cut-offs for overweight and obesity may be necessary to attain valid prevalence estimates. In New Zealand, this area is largely unexplored in both young people and Asian populations. There is a need for large-scale longitudinal studies investigating the relationships between excessive body fatness and related health outcomes across all major ethnic groups in New Zealand. PMID- 15282627 TI - Chlamydia in New Zealand. PMID- 15282626 TI - The assessment and management of primary antibody deficiency. PMID- 15282628 TI - Duration of methicillin-resistant Staphylococcus aureus colonisation in hospitalised patients. PMID- 15282629 TI - An in vitro assay to study the recruitment and substrate specificity of chromatin modifying enzymes. AB - Post-translational modifications of core histones play an important role in regulating fundamental biological processes such as DNA repair, transcription and replication. In this paper, we describe a novel assay that allows sequential targeting of distinct histone modifying enzymes to immobilized nucleosomal templates using recombinant chimeric targeting molecules. The assay can be used to study the histone substrate specificity of chromatin modifying enzymes as well as whether and how certain enzymes affect each other's histone modifying activities. As such the assay can help to understand how a certain histone code is established and interpreted. PMID- 15282630 TI - Occurrence, biological activity and synthesis of drimane sesquiterpenoids. AB - In this review the names, structures and occurrence of all new drimanes and rearranged drimanes which have been published between January 1990 and January 2003 have been collected. Subjects that have been treated are biosynthesis, analysis, biological activities, with special attention to cytotoxic activity and antifeedant and insecticidal activity and mode of action. An important part of the review deals with the synthesis of drimanes. This part has been subdivided into syntheses by transformation of natural products, syntheses starting from chiral compounds obtained by enzymatic resolution, syntheses by cationic polyolefin cyclizations, syntheses from trans-decalones, syntheses by radical cyclizations and syntheses by cycloaddition reactions. The review contains about 350 references. PMID- 15282631 TI - Antioxidants in Chinese herbal medicines: a biochemical perspective. AB - Recently, intense interest has focused on the antioxidant properties of natural products. In particular, Chinese herbal medicines (CHM) have become hot topics for life science researchers since many are reported to possess cardioprotective compounds, many of which remain to be identified. Indeed, the exact mechanisms by which CHM work remain unknown. Although many of these herbal remedies are undoubtedly efficacious, few have been scientifically investigated for their active chemical constituents and biological activities. We have previously reported higher activities of antioxidant defence enzymes such as superoxide dismutase, catalase, glutathione peroxidase and glutathione S-transferases in the liver of rats treated with the herb Salvia miltiorrhiza in a model of acute myocardial infarction. Using well established in vitro antioxidant assays employing 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS) and diphenyl-l-picrylhydrazyl (DPPH) we have shown that in addition to elevating endogenous antioxidant enzyme activity, Salvia miltiorrhiza and other CHM traditionally used for cardiovascular disorders (such as Rhizoma ligustici, Herba leonuri, Radix achyranthis bidentatae, and Camellia sinensis) contain potent antioxidant moieties in addition to their phenolic constituents. Furthermore, these novel non-phenolic components are effective inhibitors of oxidative reactions mediated by the inflammatory oxidants, peroxynitrite,hypochlorous acid and hydroxyl radical as well as iron-dependent lipid peroxidation. In this review, we discuss the various antioxidant properties of CHM in the context of their biochemical mechanisms. PMID- 15282632 TI - Directed evolution of enzymes for product chemistry. AB - This review discusses the achievements of directed evolution of enzymes relevant to natural and unnatural product chemistry focusing on the period 1998-2004, citing 156 references. A few selected papers published before 1998 are also cited for their fundamental relevance, together with additional reviews covering other aspects of directed evolution applied to closely related topics. An overview of the latest methods applied in directed evolution is given in the introduction, together with a range of approaches for the selection of variants of enzymes with desired catalytic properties. All classes of enzymes are systematically covered. PMID- 15282633 TI - Recent developments in automated structure elucidation of natural products. AB - Advancements in the field of Computer-Assisted Structure Elucidation (CASE) of Natural Products achieved in the past five years are discussed. This process starts with a dereplication procedure, supported by structure-spectrum databases. Both commercial and free products are available to support the procedure. A number of new programs,as well as advancements in existing ones, are presented. Finally, the option to validate the result by an independent procedure, a high quality ab initio quantum mechanical calculation, is discussed. PMID- 15282634 TI - Metabolites from symbiotic bacteria. AB - This review describes natural products that are shown or suspected to be synthesized by symbiotic bacteria. It includes 349 references and covers the literature in this field through 2003. PMID- 15282635 TI - Anthocyanins and other flavonoids. AB - More than 450 new flavonoid structures, reported from January 2001 until December 2003, are reviewed. They comprise anthocyanidins, flavones, flavonols, chalcones, dihydrochalcones, aurones, flavanones and dihydroflavonols, both as aglycones and as glycosides. The biological activity of some of the compounds is briefly discussed. There are 289 cited references. PMID- 15282642 TI - Aspirin and indomethacin for the prevention of experimental port-site metastases. AB - BACKGROUND: Nonsteroidal antiinflammatory drugs have been shown to have antitumor and chemopreventative effects. We investigated the potential of these drugs to inhibit port-site and intraperitoneal metastases. METHODS: The antiproliferative effect of aspirin and indomethacin on tumor cells was measured in vitro and in vivo. The in vivo experiments used DA rats to measure the effects of aspirin and indomethacin on the development of port-site metastases and the proliferation of intraperitoneal tumor cells after laparoscopy. RESULTS: In vitro, aspirin and indomethacin had an antiproliferative effect on tumor cells, inhibiting cell division and killing cells in a concentration and time-dependent manner. Orally administered aspirin and indomethacin, at the maximum tolerated dose, did not reduce the rate of intraperitoneal tumor cell division and had no effect on peritoneal metastases, or the number or size of port-site metastases. CONCLUSIONS: Despite promising in vitro studies, this study does not suggest there will be any clinical therapeutic value associated the use of aspirin or indomethacin for the prevention of the spread of tumor following the spillage of cells into the peritoneal cavity at laparoscopic surgery. PMID- 15282645 TI - Teaching, training, and clinical surgery: are we making a difference? PMID- 15282649 TI - An overview of the heparin-induced thrombocytopenia syndrome. AB - Heparin-induced thrombocytopenia (HIT) is caused by heparin-dependent, platelet activating IgG antibodies that increase thrombin generation in vivo, producing a prothrombotic phenotype. In addition to platelet activation, there is in vitro evidence that activation of endothelium and monocytes occurs, perhaps directly by HIT antibodies, but more likely through activated platelet (or microparticle) endothelial-leukocyte interactions. Patients with cardiac disease receiving heparin present important diagnostic and therapeutic issues when unexpected thrombocytopenia arises. Concomitant vascular disease burden and intravascular catheter use further increase risk of HIT-associated arterial thrombosis in this patient population. Whether arterial thrombosis simply reflects the "hypercoagulability state" of HIT interacting with diseased or injured arteries, or whether arterial "white clots" reflect additional prothrombotic effects of HIT via endothelial and monocyte activation, remains uncertain. Patients with HIT can also develop deep-vein thrombosis, which can progress to limb loss if coumarin (warfarin) leads to severe protein C depletion (coumarin-induced venous limb gangrene). Therapy for patients strongly suspected to have HIT should focus on inhibiting thrombin (or its generation) pharmacologically. Two direct thrombin inhibitors (lepirudin, argatroban) are approved for treating HIT. When using these agents, coumarin anticoagulation should be delayed pending substantial resolution of thrombocytopenia, before cautiously introducing overlapping coumarin therapy. PMID- 15282650 TI - Case studies of HIT in cardiovascular medicine. AB - We present three clinical cases that illustrate some of the key features of the diagnosis and management of immune-mediated heparin-induced thrombocytopenia (HIT). The importance of having a high clinical suspicion for HIT in the appropriate clinical setting is emphasized. Early therapeutic decisions should be based on a clinical diagnosis, with laboratory tests serving as confirmation. Low molecular-weight and unfractionated heparins are to be strictly avoided in patients with HIT. Identification bracelets or necklaces may be useful to reduce inappropriate administration of these agents to patients with HIT presenting with acute coronary syndromes. PMID- 15282651 TI - Prevalence of anti-PF4/heparin antibodies and the HIT syndrome in cardiovascular medicine. AB - Heparin-induced thrombocytopenia (HIT) is a potentially life-threatening adverse effect of heparin treatment. HIT is mediated by antibodies directed at complexes that form between heparin and platelet factor 4 in plasma and are located on the platelet surface and on endothelium. HIT occurs in 1 to 2% of patients receiving unfractionated heparin (UFH) and with lower frequency in patients receiving low molecular-weight heparins and heparinoids. Despite recent insights into the mechanisms of HIT, there remain important unresolved issues. For example, the reason for the wide difference in the frequency of HIT in patients treated with UFH in various clinical trials is not yet clear. There are patient population dependent differences in the risk for HIT immunoglobulin G and the development of thrombotic episodes. The complex nature of this syndrome may relate to the composition of the responsible antigen. Patients with HIT need a more accurate evaluation of platelet counts and a better assessment of clinical evidence for thrombosis. Alternative anticoagulant therapy approaches are being studied, but there is at this time no firm clinical evidence for which treatment is best for patients with HIT and HIT-related thromboses. PMID- 15282652 TI - Direct thrombin inhibitors in the treatment of immune-mediated heparin-induced thrombocytopenia. AB - Heparin was first discovered in 1916 and at present is used in more than 12 million patients a year. In the 1950s, several physicians noticed an uncommon paradoxical phenomenon in which heparin appeared to function as a procoagulant instead of an anticoagulant. This phenomenon is now known as the immune-mediated heparin-induced thrombocytopenia (HIT) and thrombosis syndrome (HITTS). Our understanding of this syndrome has evolved over the last 2 to 3 decades, and therapeutic options are arising. This article will focus on the most extensively studied therapy for HIT, which is the class of drugs known as the direct thrombin inhibitors. Specifically, we will focus on the mechanisms by which direct thrombin inhibitors may be useful in this syndrome, the evidence for their use, and the unique characteristics of the two FDA-approved agents in this class, lepirudin and argatroban. PMID- 15282653 TI - Percutaneous interventions in patients with immune-mediated heparin-induced thrombocytopenia. AB - The use of unfractionated heparin, the traditional antithrombotic agent during percutaneous coronary interventions (PCI), is associated with the risk of heparin induced thrombocytopenia, a rare but often fatal clinical condition. This article focuses on several issues related to heparin-induced immune-mediated thrombocytopenia (HIT, type II) and alternative modes of periprocedural anticoagulation in patients with suspected or known HIT. The hypercoagulable state characterizing HIT, along with mechanical plaque disruption resulting from PCI place patients with HIT at particular risk of thrombosis during PCI. Given that a diagnosis of HIT precludes any further use of heparin, other treatment modalities are essential. Direct thrombin inhibitors are the drugs of choice in this challenging situation. These agents offer several advantages as anticoagulants for patients with HIT: (1) the ability to inhibit both thrombin that is bound to fibrin (clot-bound thrombin) and fluid-phase free thrombin; (2) rapid achievement of steady state; and (3) no cross-reactivity with HIT antibodies. Recent data on the use of bivalirudin, lepirudin, and argatroban in the setting of PCI in patients with HIT are encouraging. Optimal dosing regimens for argatroban, lepirudin, and bivalirudin should be further established in PCI patients. PMID- 15282654 TI - The use of direct thrombin inhibitors in cardiovascular surgery in patients with heparin-induced thrombocytopenia. AB - One of the most important adverse drug reactions that physicians encounter is the life- and limb-threatening prothrombotic syndrome known as heparin-induced thrombocytopenia (HIT). Unfractionated heparin (UFH), administered during cardiopulmonary bypass (CPB), is highly immunogenic. Heparin-dependent antibodies can develop in 25 to 50% of UFH-treated cardiac surgery patients within 5 to 10 days. These antibodies can activate platelets and are considered the causative agents of HIT. HIT is a relatively common complication, occurring in 1 to 3% of cardiovascular surgery patients when UFH administration is continued postoperatively. It is strongly associated with new thromboembolic events leading to limb amputation and death. In acute or recent (< 100 days) HIT, alternative anticoagulatory regimens are needed during CPB surgery for prevention of HIT related thrombosis. Treatment options for such patients now generally include the use of alternative anticoagulants such as lepirudin, bivalirudin, or danaparoid, as well as a combined treatment with platelet-function inhibitors and heparin. In patients with a history of HIT and no detectable antibodies, heparin is currently the safest approach for high-dose anticoagulation during CPB. Before and after surgery, however, alternative anticoagulants should be used. The risk of clinical HIT after heart surgery could potentially be reduced by using low-molecular weight heparins for postsurgery anticoagulation. PMID- 15282655 TI - Bivalirudin in PCI: an overview of the REPLACE-2 trial. AB - The Randomized Evaluation in PCI Linking Angiomax to Reduced Clinical Events (REPLACE)-2 trial is one of the largest acute randomized controlled trials evaluating the efficacy of two anticoagulant strategies during contemporary urgent or elective percutaneous coronary intervention (PCI). The direct thrombin inhibitor, bivalirudin, with provisional use of glycoprotein IIb/IIIa (GP IIb/IIIa) inhibitor was compared to low-dose unfractionated heparin (UFH) plus planned GP IIb/IIIa inhibitor. At 30-day follow-up, the primary quadruple composite endpoint (death, myocardial infarction (MI), urgent repeat revascularization, or in-hospital major bleeding) occurred in 9.2% of patients in the bivalirudin group versus 10.0% of patients in the UFH plus GP IIb/IIIa inhibitor group. The secondary triple composite endpoint (death, MI, urgent repeat revascularization) occurred in 7.6% of patients in the bivalirudin group compared with 7.1% of patients in the UFH plus GP IIb/IIIa inhibitor group. Both endpoints met formal statistical criteria for noninferiority to UFH plus GP IIb/IIIa inhibitor. By imputed comparison from historic GP IIb/IIIa trials between bivalirudin versus UFH alone, REPLACE-2 demonstrated that bivalirudin was superior to UFH alone with respect to the quadruple and triple composite endpoints. Furthermore, bivalirudin plus provisional GP IIb/IIIa blockade was associated with a significant reduction in in-hospital bleeding (2.4% vs. 4.1%; p < 0.001). At 6 months' follow-up, there was no significant difference in rates of death, MI, or revascularization between the two groups. Furthermore, there was no evidence that the early, nonsignificant 0.5% excess non-Q-wave MI in the bivalirudin group translated into later mortality. There was a trend toward decreased mortality at 6 months in the bivalirudin arm (0.95% vs. 1.35%; p = 0.148). The relative efficacy of bivalirudin versus UFH plus GP IIb/IIIa inhibitor was similar in several high-risk subgroups, including patients with diabetes mellitus or prior MI, women, the elderly (age > 65 years), and patients undergoing PCI of bypass grafts. Bivalirudin represents an exciting alternative to UFH plus GP IIb/IIIa inhibitor in patients undergoing urgent and elective PCI with similar suppression of ischemic events, fewer bleeding complications, and the potential for greater cost savings and ease of administration. PMID- 15282656 TI - Bivalirudin, blood loss, and graft patency in coronary artery bypass surgery. AB - A safe and effective alternative is needed for patients in whom unfractionated heparin (UFH) or protamine is contraindicated (e.g., those with heparin-induced thrombocytopenia or allergy to protamine). Furthermore, choice of anticoagulant may influence graft patency in coronary surgery and may therefore be important even when there is no contraindication to UFH. Direct thrombin inhibitors have several potential advantages over UFH, demonstrated in acute coronary syndromes. However, there are also potential difficulties with their use related to lack of reversal agents and paucity of clinical experience in monitoring their anticoagulant activity at the levels required for cardiac surgery with cardiopulmonary bypass (CPB). In the first prospective randomized trial of an alternative to heparin in cardiac surgery, we compared bivalirudin (a short acting direct thrombin inhibitor) with UFH in 100 patients undergoing off-pump coronary artery bypass (OPCAB) surgery. Blood loss for the 12 hours following study drug initiation in the bivalirudin group was not significantly greater than in the heparin group. Median graft flow was significantly higher in the bivalirudin group. We concluded that anticoagulation for OPCAB surgery with bivalirudin was feasible without a clinically important increase in perioperative blood loss. A larger study is needed to investigate the impact of improved graft patency on other clinical outcomes after cardiac surgery. PMID- 15282657 TI - Role of anti-PF4/heparin antibodies in recurrent thrombotic events after acute coronary syndromes. AB - We postulated that patients with recent acute coronary syndromes and antibodies to the platelet factor 4/heparin complex would have an increased risk of myocardial infarction (MI), even in the absence of thrombocytopenia. We analyzed sera from patients enrolled in the placebo/unfractionated heparin arm of the GUSTO IV-ACS trial who had a high likelihood of prior heparin exposure. We selected 109 patients without thrombocytopenia with the 30-day primary endpoint (death, MI, or revascularization) and 109 age-, gender-, and race-matched controls who did not achieve the primary endpoint. Twenty-three of 218 patients (10.6%) had anti-PF4/heparin antibodies. Patients with anti-PF4/heparin were more likely to have death or MI (30.4% vs. 11.3%, p = 0.011) or MI (21.7% vs. 6.2%, p = 0.008) than patients who were negative for the antibody. Antibody-positive patients had higher levels of sVCAM-1 (892 +/- 263 microg/L vs. 780 +/- 228 microg/L; p = 0.04) and sICAM-1 (246 +/- 50 microg/L vs. 222 +/- 71 microg/L; p = 0.02) than antibody-negative patients. In a multiple logistic regression model that included inflammatory markers and clinical risk factors, antibodies to PF4/heparin were a strong predictor of 30-day MI (odds ratio, 9.0; 95% confidence intervals, 2.1 to 38.6; p < 0.01), with IL-6 being the only other predictor (odds ratio, 1.1; 95% confidence intervals, 1.0 to 1.2; p = 0.03). Antibodies to the platelet factor 4/heparin complex are a novel, independent predictor of MI at 30 days in patients presenting with acute coronary ischemic syndromes. Antibodies to PF4/heparin are a stronger predictor of MI than clinical characteristics or inflammation markers. PMID- 15282658 TI - Interactions of platelet factor 4 with the vessel wall. AB - Platelet factor 4 (PF4) is a platelet-specific protein that is stored in platelet alpha granules and released following platelet activation. PF4 was the first chemokine that was isolated, but unlike other chemokines, it may not have a clear role in inflammation. Gathering evidence suggests that unlike other chemokines that bind to specific receptors, PF4's biology depends on its unusually high affinity for heparan sulfates and other negatively charged molecules at concentrations attained in the immediate vicinity of activated platelets. There has been one report that PF4 binds to CXCR3B, a chemokine receptor isoform that may be present in some vascular beds, but the biological relevance of this single observation is not clear. We propose that the main biological role of PF4 and the basis for its presence in the alpha granules of all known mammalian platelets is to neutralize surface heparan sulfate side-chains of glycosaminoglycans and to optimize thrombus development at sites of vascular injury. In addition, the binding of PF4 to surface glycosaminoglycans may also underlie its angiostatic and proatherogenic properties. Additionally, PF4 binds to several other proteins that are central to thrombosis, angiogenesis, and atherogenesis. These interactions may also contribute to its biological and pathobiological effects. Certainly, future studies using in vivo models to test biological relevance of each of these proposed mechanisms by which PF4 interacts with the vasculature are needed, as are studies to define the importance of PF4 binding to CXCR3B. PMID- 15282659 TI - A critical evaluation of assays for detecting antibodies to the heparin-PF4 complex. AB - Heparin-induced thrombocytopenia (HIT) is a potentially catastrophic complication of heparin therapy. The syndrome is the result of the production of an antibody to the complex that forms between heparin and platelet factor 4 (H-PF4) released from activated platelets. IgG antibodies bind to platelet Fc receptors and cause platelet activation, resulting in thrombocytopenia and greatly increased risk of thrombosis. Tests for the H-PF4 antibody can be classified into functional assays (which rely on the demonstration of platelet activation) and immunoassays (which detect the presence of an antibody without regard for its functional ability). The functional assays have a greater specificity for clinical HIT, but require normal donor platelets and are relatively un-standardized. The immunoassays have the advantage of better standardization and do not require normal platelets, but may give positive results in the absence of clinical HIT. The choice of test is usually dictated by what is possible for a given laboratory in terms of instrumentation, expertise, and interest. For most institutions this will be a commercially available enzyme-linked immunosorbent assay. Although HIT is a true clinicopathological syndrome, there are several reasons why its diagnosis still rests primarily on clinical grounds. First, laboratory tests may not be available locally. Second, they may not be available in a sufficiently timely manner. Finally, available tests are not completely sensitive or specific for the condition. There is, therefore, a continuing need to develop more rapid testing strategies with greater specificity for clinical HIT. PMID- 15282660 TI - Recombinant platelet factor 4 for heparin neutralization. AB - Protamine sulfate has been used for many years to reverse the effects of unfractionated heparin, but it can cause hemodynamic changes and other serious side effects. Platelet factor 4 (PF4) is a naturally occurring protein synthesized in megakaryocytes and eventually stored in the alpha granules of platelets for later release. Although the complete physiologic role of PF4 is unknown, it is highly effective for the neutralization of heparin anticoagulation. Several preliminary animal studies and trials using blood obtained from cardiopulmonary bypass circuits suggested recombinant PF4 (rPF4) would be an effective alternative to protamine. In the first open-label, phase 1 human study, patients received rPF4 in doses of 0.5, 1.0, 2.5, or 5.0 mg/kg over 3 minutes to reverse heparin anticoagulation after diagnostic cardiac catheterization. There were no important hemodynamic changes and the rPF4 was highly effective in neutralizing heparin. Serial measurements of rPF4 levels showed a monophasic elimination pattern with a serum half-life of 25.5 +/- 13.5 minutes that was independent of dose administered. A randomized and blinded trial comparing rPF4 to protamine confirmed the safety and effectiveness of rPF4. Although rPF4 was initially being evaluated as a clinical alternative to protamine, it is not currently being developed for general clinical use. PMID- 15282661 TI - Platelet factor 4: an inhibitor of angiogenesis. AB - Platelet factor 4 (PF4) is an antiangiogenic ELR-negative chemokine. PF4 inhibits endothelial cell proliferation and migration and angiogenesis in vitro and in vivo. Three different mechanisms have been proposed to explain PF4's antiangiogenic effects. First, PF4 may bind proteoglycans and interfere with the proteoglycan-bystander effect on growth factor activity. Second, PF4 is able to interact directly with angiogenesis growth factors such as fibroblast growth factors or vascular endothelial growth factors and inhibits their interaction with cell surface receptors. Third, PF4 may activate cell surface receptors on endothelial cells and induce inhibitory signals. Recently, one such receptor, CXCR3-B, was identified. In cardiovascular disease, PF4 may possibly intervene in collateral vessel formation, plaque neovascularization, heparin-induced thrombocytopenia and stent endothelialization. Several PF4 fragments such as PF4 CTF and modified molecules have been made that exhibit antiangiogenesis properties and may serve as leads for further therapeutic development. PMID- 15282662 TI - Hematologic differences in heterophile-positive and heterophile-negative infectious mononucleosis. AB - Infectious mononucleosis (IM) due to all causes is characterized by atypical lymphocytosis. We sought to compare hematologic parameters of infectious mononucleosis due to Epstein-Barr virus (EBV) infection (heterophile antibody (HA) positive) with mononucleosis due to other causes. Mono-Latex Slide Agglutination Test results and complete blood counts (CBC) of 147 patients with mononucleosis were retrospectively analyzed. Leukocyte count, absolute lymphocyte count, and presence of atypical lymphocytes in EBV-positive and EBV-negative groups were statistically compared. We analyzed 68 EBV-positive and 79 EBV negative cases. EBV-positive patients were significantly younger than EBV negative patients were. Mean total WBC count and mean absolute lymphocyte count were significantly higher in EBV-positive patients. Absolute lymphocytosis, absolute leukocytosis, and atypical lymphocytosis were also significantly more frequent in EBV-positive patients. Leukopenia was more frequently seen in EBV negative patients. PMID- 15282663 TI - Increasing dose intensity of anthracycline antibiotics improves outcome in patients with acute myelogenous leukemia. AB - To understand the effect of dose concentration in the overall survival of AML, we conducted a study on the efficacy and toxicity of a drug combination where the dose of daunorubicin was intensified. For this analysis, the outcome of patients entered into two consecutive prospective trials was compared. Inclusion criteria in both arms were identical and consisted of primary AML in adults. Treatment protocol for Cape Town Regimen 4 (CTR-IV) comprised of cytarabine infusion (100 mg/m(2)) and etoposide (100 mg/m(2)), injection daily for 7 days in combination with daunorubicin (45 mg/m(2)) on days 1, 2, and 3. Patients achieving remission were given two further courses of the same chemotherapy and received allogeneic or autologous transplantation. CTR-V was a similar treatment program, except that daunorubicin was escalated on days 1, 2, and 3 to 75 mg/m(2) during induction and to 60 mg/m(2) during a single consolidation. Patients were also offered stem cell transplantation. Between 1990 and 1997, 78 patients (median age 33; range 13-67 years) fulfilled entry criteria and received CTR-IV. From 1998 onwards, 35 patients (median age 36; range 15-66 years) were prospectively enlisted into the CTR-V trial. The patient population in CTR-V had fewer Caucasian individuals (P = 0.02) and had significantly lower presentation hemoglobin (P = 0.0002). Following initiation of induction chemotherapy, 40 patients failed to respond. Among these, 10 patients demised before day 28. Another 30 (25/69 CTR-IV and 5/32 in CTR-V groups; P = 0.01) had leukemia that was resistant to chemotherapy, and all died. Remission was achieved in 59% of patients treated with CTR-IV and 77% of those receiving CTR-V (P = 0.03). CR occurred with a single course in 64% versus 88% (P = 0.02), respectively. There were no differences in the toxicity profile between these two combinations. Disease recurred in 50% and 28% (P = 0.07) of patients. For the 113 individuals, median follow up is 254 (range 19-4,451) and 304 (12 1,702; P = 0.03) days. Survival is 23% and 40%, respectively, favoring patients treated with CTR-V (log rank; P = 0.03). Cox regression analysis showed that treatment group (P < 0.001), FAB type, hemoglobin level, and platelet count were independent factors for response to chemotherapy. Older age and not undergoing myeloablative therapy were the only adverse factors for survival. We conclude that increase in the treatment dose of daunorubicin in patients with AML led to a higher remission rate, particularly with a single course of chemotherapy and had an equivalent toxicity profile. This therapeutic modification is also likely to result in substantial reduction in patient stay in hospital and in the overall expenditure. PMID- 15282664 TI - Primary upper-extremity deep vein thrombosis: high prevalence of thrombophilic defects. AB - Primary deep venous thrombosis of the upper extremity (UEDVT) is an unusual disorder. Limited data are available on the contribution of hypercoagulable status in the pathogenesis of this disease. This study aims to report the prevalence of inherited and acquired thrombophilic risk factors (TF) in patients with primary (effort-related and spontaneous) UEDVT. From 1993 to 2002, 31 patients (17 females, median age 38.8 years, range 16-60 years; and 14 males, median age 31.4 years, range 20-56 years) with primary UEDVT (n = 15 effort related and n = 16 spontaneous) were referred for screening of hypercoagulable status. Nineteen (61.3%) patients had at least one coagulation abnormality. The most common acquired TF were antiphospholipid antibodies (31% lupus anticoagulant and 12.9% anticardiolipin antibodies). Factor V Leiden (12.9%) and prothrombin G20210A mutation (20%) were the most prevalent genetic risk factors. Five patients (16.1%) had high plasma homocysteine levels, and one patient (4.7%) had protein S deficiency. Effort-related UEDVT was associated with male gender (P = 0.04) and younger age (P = 0.02). There was no significant difference in the prevalence of acquired or inherited TF between patients with effort-related or spontaneous UEDVT. A local anatomic abnormality was detected in seven patients (22.5%), and the prevalence of TF was significantly lower within this group (P = 0.006). The incidence of TF in patients without an anatomic abnormality was 75% (RR 5.25). This study found a high prevalence of an underlying thrombophilic status in spontaneous and effort-related UEDVT. Hypercoagulable status may play a significant role in both groups. Screening for local anatomical abnormalities and thrombophilia should be included in the evaluation of primary UEDVT. PMID- 15282665 TI - High prevalence of anti-prothrombin antibody in patients with deep vein thrombosis. AB - The present study was designed to determine the prevalence of lupus anticoagulant (LA) antibody and several antibodies for antiphospholipid syndrome (APS) in patients with deep vein thrombosis (DVT)/pulmonary embolism (PE) (n = 48), cerebral thrombosis (CT, n = 30), systemic lupus erythematosus (SLE, n = 22), and idiopathic thrombocytopenic purpura (ITP, n = 30). The presence of antibodies was examined by using the respective ELISA kits. LA was positive in 38.6% of patients with DVT/PE, suggesting that LA is one of the most important risk factors in DVT/PE. The highest prevalence of anti-beta(2) glycoprotein I (beta(2)GPI) IgG was in CT and SLE, followed by DVT, and none in ITP and healthy volunteers (control, n = 40), suggesting that it is related to thrombosis, particularly arterial thrombosis. The highest prevalence of anti-prothrombin (aPT) IgG antibody was in DVT, followed by CT and SLE, and none in ITP and the control, suggesting that it is related to thrombosis, especially venous thrombosis. The highest prevalence of antiphospholipid (aPL) IgG was in DVT, CT, and SLE, but 0% in ITP and control. On the other hand, aPL IgM, anti-annexin V IgG, and anti annexin V IgM were positive in patients both with and without thrombosis, suggesting that they are not related to thrombosis. Our results indicated that among the anti-phospholipid antibodies, LA is the most sensitive marker for APS while anti-beta(2)GPI IgG, aPT IgG, and aPL IgG are risk factors for thrombosis. In particular, aPT IgG is a significant marker for DVT/PE. PMID- 15282666 TI - Increased levels of soluble ICAM-1 in the plasma of sickle cell patients are reversed by hydroxyurea. AB - Increased adhesive events between the blood vessel endothelium and red and white cells play a central role in the initiation of vasoocclusive crisis in sickle cell disease (SCD). Soluble VCAM-1 levels are increased in the plasma of sickle cell patients and may be reduced during hydroxyurea (HU) therapy. Reports regarding any changes in soluble ICAM-1 (sICAM-1) levels in sickle cell patients, however, are conflicting, and as yet no beneficial effect of HU upon levels has been observed. Thus, we sought to thoroughly investigate changes in sICAM-1 levels in SCD patients and the effect of HU therapy (20-30 mg/kg/day). Plasma sVCAM-1 levels were significantly higher in steady-state SCD patients than in normal controls (766 +/- 86 ng/mL vs. 325 +/- 38 ng/mL, respectively, P < 0.0001). sVCAM-1 levels were decreased in patients on HU therapy (543 +/- 69 ng/mL) compared to those not taking HU; however, this difference was not significant. Plasma sICAM-1 levels were significantly increased in steady-state SCD patients compared to normal individuals (285 +/- 20 ng/mL vs. 202 +/- 16 ng/mL, respectively, P = 0.002), and HU therapy significantly reduced sICAM-1 levels in patients (217 +/- 12, P = 0.027) to levels approaching those of healthy individuals. sVCAM-1 levels inversely correlated with fetal hemoglobin levels in SCD patients, while a nonsignificant inverse trend was observed between sICAM-1 levels and fetal hemoglobin. In conclusion, plasma sICAM-1 levels were significantly increased in SCD patients, and this increase was reversed by hydroxyurea therapy, possibly reflecting reduced endothelial activation in patients taking HU. Such an event may benefit patients by reducing adhesive interactions between white cells and the endothelium. PMID- 15282667 TI - Association of functional thrombin-activatable fibrinolysis inhibitor (TAFI) with conventional cardiovascular risk factors and its correlation with other hemostatic factors in a Spanish population. AB - Our aim was to determine the associations of functional thrombin-activatable fibrinolysis inhibitor (TAFI) levels in plasma with conventional cardiovascular risk factors, sex and age, and possible correlations with other hemostatic factors in a Spanish population. We included 303 individuals from a Spanish population. Hemostatic factors such as von Willebrand Factor, VII ag, VIIIc, XIc, XIIc, APCR, protein S, protein C, antithrombin, fibrinogen, and t-PA antigen were assayed. The functional TAFI assay was based on the activation of plasma TAFI with thrombin-thrombomodulin, and the measure of TAFIa activity on the hippuryl Arg substrate. There were no statistical differences in mean values of functional TAFI among the various female age groups or among the different male age groups, with or without cardiovascular risk factors. Only women younger than 30 years of age showed lower levels of functional TAFI compared to older women. No differences were found among men of different ages. Adjusted for sex and age, hemostatic factors did not show a correlation with functional TAFI levels in plasma. Women with hypercholesterolemia showed higher levels of TAFI; other conventional cardiovascular risk factors did not modify functional TAFI levels either in men or in women. We also found no correlation of functional TAFI levels related to any other hemostatic factors. PMID- 15282668 TI - Successful treatment of idiopathic thrombocytopenic purpura-like syndrome in a cancer patient with low-dose interferon: case report and review of the literature. AB - Idiopathic thrombocytopenic purpura (ITP)-like syndrome is a rare complication of carcinomas, and its treatment usually precedes cancer therapy in order to make further procedures safe. We describe the case of a 78-year-old man with a small cell lung cancer of extended stage, associated with ITP-like syndrome, which proved resistant to treatment with corticosteroids and vincristine, short responsive to IV immunoglobulin, but quickly and steadily responsive to low-dose interferon (ld IFN, 3 million IU, sc, twice weekly, for 8 weeks), until the patient's death, due to his primarily chemoresistant cancer. This case is, to our knowledge, the second reported ITP-like syndrome in a cancer patient who had been successfully treated with ld IFN. The excellent and cost-effective therapeutic index of ld IFN makes it an attractive alternative treatment in patients with this specific complication and calls for further investigation regarding its potential use as a first-line treatment. PMID- 15282669 TI - ABL oncogene amplification with p16(INK4a) gene deletion in precursor T-cell acute lymphoblastic leukemia/lymphoma: report of the first case. AB - Gene amplification is a relatively rare event in hematologic malignancies. The ABL gene on chromosome band 9q34 is a proto-oncogene and is the well-known translocation partner of the BCR gene on 22q11 giving rise to t(9;22)(q34;q11), which is the hallmark of chronic myeloid leukemia and is the most common chromosomal abnormality in adult acute lymphoblastic leukemia (ALL). Amplification of ABL is an exceedingly rare event, with only less than 5 cases reported in the literature. The p16(INK4a) (or CDKN2A) gene on 9p21 is a tumor suppressor gene, and deletion thereof is recently recognized as one of the most common genetic abnormalities in ALL. The authors herein describe an 8-year-old male patient with precursor T-cell ALL harboring both ABL gene amplification and p16(INK4a) gene deletion. Fluorescence in situ hybridization (FISH) analysis using BCR/ABL probes revealed five or more ABL signals, indicating amplification in 51.5% of interphase nuclei. FISH using p16(INK4a) gene probes showed heterozygous p16(INK4a) deletion in 71.0%. On conventional cytogenetic analysis, however, only 10 metaphases were available, which showed the normal karyotype, 46,XY[10], serving no evidence for the findings on FISH. This is the first report of an ALL case with ABL amplification, and the authors speculate that both ABL proto-oncogene amplification and the p16(INK4a) tumor suppressor gene deletion have been implicated in leukemogenesis in the present case, although whether the ABL amplification truly contributes to the leukemogenesis or merely an epiphenomenon representing underlying genomic instability remains to be determined. PMID- 15282670 TI - Prolonged fever of unknown origin and hemophagocytosis evolving into acute lymphoblastic leukemia. AB - Hemophagocytic syndrome (HPS) is an unusual acute syndrome presenting with fever, hepatosplenomegaly, and cytopenias. The hallmark of HPS is the accumulation of activated macrophages that engulf hematopoietic cells in the reticuloendothelial system. Most cases of HPS in adults are secondary to infection or malignancy, and thus investigation of the underlying disease is necessary. We describe a patient with prolonged fever, HPS, and chromosomal abnormalities in the bone marrow who underwent thorough evaluation for the cause of his symptoms. A final diagnosis of acute lymphoblastic leukemia (ALL) was established in a fourth, repeated bone marrow biopsy performed more than 2 months after the first presenting symptom appeared. This unusual case demonstrates the importance of cytogenetic abnormalities found in cases of HPS and the importance of repeated testing when an underlying disease is suspected. PMID- 15282671 TI - Successful control of Epstein-Barr virus (EBV)-infected cells by allogeneic nonmyeloablative stem cell transplantation in a patient with the lethal form of chronic active EBV infection. AB - Chronic active Epstein-Barr virus infection (CAEBV) is a heterogeneous EBV related disorder, ranging from mild/moderate forms to rapidly lethal disorders. The lethal form of CAEBV is characterized by multiple organ failure, hemophagocytic syndrome, and development of lymphomas. Allogeneic stem cell transplantation is considered as the only potentially curative treatment for the lethal form of CAEBV, but it is not always desirable because of the high incidence of regimen-related toxicities. A 17-year-old female with CAEBV, who was refractory to conventional therapies and considered to be unable to receive a myeloablative regimen because of multiple organ dysfunction, underwent allogeneic nonmyeloablative stem cell transplantation (allo-NST) before developing a hematological malignancy. She has been well without any signs of CAEBV for 27 months after allo-NST, and we confirmed that specific cytotoxic T lymphocyte activity against EBV was reconstituted. This outcome suggests that allo-NST can control CAEBV by reconstituting the host immunity against EBV. PMID- 15282672 TI - Cardiobacterium hominis endocarditis associated with very severe thrombocytopenia and platelet autoantibodies. AB - Severe thrombocytopenia is a life-threatening condition. It is often associated with immune-mediated platelet destruction or myeloablative chemotherapy. Infective endocarditis has been associated with thrombocytopenia, which, as in sepsis, tends to be mild and is often the result of several pathological mechanisms. We report a case of Cardiobacterium hominis endocarditis associated with very severe thrombocytopenia and bleeding in a patient who refused platelet transfusion. Platelet autoantibodies directed against glycoprotein (Gp) IIb/IIIa and Gp Ib/IX were detected during active infection using a glycoprotein-specific assay. Successful treatment of C. hominis endocarditis was associated with loss of platelet autoantibodies and recovery of the platelet count. This report illustrates that the development of platelet autoantibodies can contribute to very severe thrombocytopenia in occasional patients with infective endocarditis. PMID- 15282673 TI - Severe hemolytic anemia associated with Hb Volga [beta27(B9)Ala-->Asp]: GCC-->GAC at codon 27 in a Turkish family. AB - A boy presented at age 4 years with severe congenital hemolytic anemia characterized by highly elevated reticulocyte count (30-50%) and prominent basophilic stippling. Hb had been 4 g/dL at age 7 months. The patient was on a monthly transfusion regimen up to the age of 7 years, when he underwent splenectomy. After removal of the spleen, his Hb stabilized at 11 g/dL. No abnormal pattern was detected in hemoglobin electrophoresis at pH 9 and 6. In vitro globin synthesis revealed the presence of an abnormal beta-chain in front of the gamma-chain. The beta(A)/beta(X) ratio was 0.77 at 30 min and 0.74 at 2 hr of incubation. Molecular analysis revealed that the patient had GCC-->GAC alteration at codon 27 (beta27(B9)Ala-->Asp) causing the abnormal hemoglobin Volga. The beta-cDNA derived from the beta-Hb Volga allele could be differentiated from HbA beta-cDNA on silver-stained gel. No imbalance in the mRNA of beta(A)/beta(Hb Volga) ratio was observed. PMID- 15282674 TI - Heparin-induced thrombocytopenia presenting with thrombosis of multiple saphenous vein grafts and myocardial infarction. AB - We report herein a patient with coronary artery disease that developed heparin induced thrombocytopenia after coronary artery bypass graft with resulting thrombosis of multiple saphenous vein grafts and myocardial infarction after heparin exposure. The patient required lepirudin and a cardiac catheterization with placement of stents. PMID- 15282675 TI - Desferrioxamine related maculopathy: a case report. AB - Desferrioxamine is used for the treatment of chronic iron overload, acute iron poisoning, and certain anaemias. Ocular toxicity secondary to prolonged treatment with desferrioxamine may result in night blindness, visual field constriction, cataract, pigmentary retinopathy and optic neuropathy. To avoid such complications an ophthalmic screening has been suggested for patients taking desferrioxamine. We report an 81-year-old patient who developed irreversible ocular toxicity despite undergoing ophthalmic screening. PMID- 15282676 TI - Donor cell myelodysplastic syndrome after allogeneic stem cell transplantation responding to donor lymphocyte infusion: case report and literature review. AB - Allogeneic hematopoietic stem cell transplantation (SCT) is a potentially curative treatment for patients with myelodysplastic syndrome (MDS). Relapses after transplantation however, are not uncommon and are usually due to re emergence of a recipient derived, neoplastic, stem cell clone. We report a unique case of MDS recurring 5 months after non-myeloablative, sibling, allogeneic SCT. Interestingly, chimerism analysis at relapse showed hematopoiesis to be entirely of donor origin confirming donor cell MDS. Donor lymphocyte infusion (DLI) produced a hematological response lasting several months. Our review of the literature shows donor-derived MDS to be very rare, with only four such cases described previously. In this report, we describe the details of our case and discuss putative mechanisms underlying the genesis of donor cell MDS and the observed response to DLI. PMID- 15282677 TI - A case of familial thrombocytosis: possible role of altered thrombopoietin production. AB - Familial thrombocytosis (FT) is an inherited disorder with clinical presentations similar to essential thrombocytosis (ET). In several pedigrees, overproduction of thrombopoietin (TPO) has been shown to be responsible for the disease. We report herein three cases of thrombocytosis in three successive generations. All cases had increased serum TPO levels. Sequence analysis of TPO gene and transmembrane domain of c-MPL, known as the TPO receptor, revealed no mutations. Platelet c-MPL expression was similar or slightly increased as compared to normal volunteers. These data suggest that altered regulation of the TPO gene might be involved in the pathogenesis of FT. PMID- 15282678 TI - Comparison of subcutaneous infusion needles for transfusion-dependent thalassemia patients by the intrapersonal cross-over assessment model. AB - Needle-induced trauma is one of the major contributing factors for poor compliance in patients with thalassaemia major on iron chelation therapy. A new generation of needles is currently available on the market, but their theoretical advantages have not been tested clinically. We performed a study to compare the pros and cons of the representative prototypes from each of the new (Thalaset needle) and old (butterfly scalp vein needle) generations of needles. Patients with thalassemia major who had been receiving subcutaneous iron chelation therapy for at least 2 years were recruited. Patients using butterfly needles were instructed to switch to the newer form of needle (Thalaset) for 2.5 months and then to change back to butterfly needles for another 2.5 months. Comparison was done by the intrapersonal cross-over model using three identical sets of questionnaires collected at the beginning of the study and after the use of Thalaset and butterfly needles, respectively. Fifty-four (22 females; 32 males) patients were included in the statistical analysis. The median age was 24.1 years (range = 7.6-47.2 years). Local reactions such as pain, itchiness, tenderness, and swelling were significantly different among the three evaluation periods and were all in favor of the Thalaset needle (all with P < 0.001), even after adjusting for the intention-to-treat calculation. The Thalaset needle is significantly better than the butterfly needle in reducing needle-related trauma. It induced fewer local skin reactions such as pain, itchiness, tenderness, and swelling. However, recommendations for its routine clinical use require further cost-effectiveness analysis. PMID- 15282679 TI - Rapid detection of common Chinese glucose-6-phosphate dehydrogenase (G6PD) mutations by microarray-based assay. AB - Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common human enzymopathy, affecting more than 200 million people worldwide. To date more than 123 mutations in the G6PD gene have been discovered, among which 12 point mutations are found in the Chinese. Setting up a simple and accurate method for detecting these mutations is not only useful for diagnosing G6PD deficiency under some circumstances that it is difficult to measure the activity of the enzyme, but also for studying the frequency of the G6PD genotypes. The purpose of this study was to develop a simple, inexpensive and accurate method for detecting these common mutations. Microarray-based assay was described in this study. Samples from 198 G6PD-deficient persons were investigated. The DNA sequencing data supported the results obtained by microarray-based assay. Thus, we concluded that the microarray-based assay is a rapid, simple, inexpensive, and accurate method for detecting the most common G6PD gene mutations among the Chinese. This method involves the selective amplification of human G6PD gene with specific oligonucleotide primers, fragmentation and labeling of PCR products, followed by hybridization with allele-specific oligonucleotide (ASO) probes on chip. PMID- 15282680 TI - Peripheral blood mitosis in association with Mycoplasma pneumoniae infection. PMID- 15282681 TI - Adherence to deferoxamine therapy: heeding Hippocrates and Osler. PMID- 15282682 TI - Mutations in PTPN11 are rare in adult myelodysplastic syndromes and acute myeloid leukemia. PMID- 15282683 TI - The use of alternative delivery systems and new technologies in the treatment of patients with eating disorders. AB - OBJECTIVE: The purpose of the current article is to review the literature regarding the use of alternative delivery systems, such as telemedicine, and new technologies, such as the use of hand-held computers, in the treatment of patients with eating disorders. METHOD: The literature is reviewed in the following areas: self-help (supervised and unsupervised), telemedicine, telephone therapy, e-mail, internet, computer software, CD-ROMs, portable computers, and virtual reality techniques. RESULTS: A growing literature suggests a number of alternative delivery systems hold promise, in particular permitting patients to access services who otherwise would not be able to receive treatment. Although most of these areas are early in their development, a growing literature supports the utility of several of these approaches. DISCUSSION: Although the literature in this area is limited, and the research base is small, a number of these technologies appear to hold substantial promise for the treatment of patients with eating disorders. PMID- 15282684 TI - A case series evaluation of guided self-help for bulimia nervosa using a cognitive manual. AB - OBJECTIVE: The current study examined the usefulness of a new, cognitive-based self-help manual for bulimia nervosa. METHOD: Twenty people were provided with assessment and six sessions of guided self-help using the manual. Participants were assessed for eating-related behaviors and attitudes and psychopathology at pretreatment, posttreatment, and at the 3-month follow-up. Assessment instruments included the Eating Disorder Examination, Symptom Checklist-90-Revised, Rosenberg Self-Esteem Scale, Screening Test for Co-morbid Personality Disorders, and The University of Rhode Island Change Assessment. Data from 15 people were available at posttreatment and from 13 people at follow-up. RESULTS: Using intention-to treat analyses, binge eating, vomiting, four of the five eating attitudes and self-esteem significantly improved between pretreatment and posttreatment. At follow-up, there was continued improvement on all measures, with the exception of binge eating. DISCUSSION: Guided self-help using cognitive techniques is a promising first-line treatment for bulimia nervosa, with further evaluation required in a randomized, controlled trial with long-term follow-up. PMID- 15282685 TI - Secondary prevention for eating disorders: the impact of education, screening, and referral in a college-based screening program. AB - OBJECTIVE: The first National Eating Disorders Screening Program (NEDSP), conducted on more than 400 college campuses in 1996, was an educational and two stage screening program designed to detect potentially clinically significant disordered eating attitudes and behaviors and offer referrals for further evaluation when warranted. The current study assessed the impact of the NEDSP on participants. METHOD: A randomly selected subset of this sample (n = 289) was contacted approximately 2 years after the NEDSP to assess the impact of the program on knowledge and treatment-seeking behaviors for eating disorders. RESULTS: For greater than 80% of the participants, the program made participants aware of the danger of eating disorders and the availability of treatment. Of those who received a recommendation for further clinical evaluation of disordered eating (n = 109), nearly one half (47.7%) followed up on this recommendation and kept at least a first appointment and 39.4% actually sought treatment subsequent to the NEDSP. DISCUSSION: The results of the current study suggest that educational and screening programs may be a promising strategy for secondary prevention of eating disorders. They also suggest that awareness of the risks of disordered eating and available treatment may not be sufficient to motivate individuals to adhere to recommendations to seek treatment. Clinicians should, therefore, be vigilant for nonadherence to treatment recommendations and proactive in facilitating treatment. PMID- 15282686 TI - The contribution of anxiety and food restriction on physical activity levels in acute anorexia nervosa. AB - OBJECTIVE: Excessive exercise is present in 40%-80% of anorexia nervosa (AN) patients. Hyperactivity often plays a role in developing and maintaining AN and represents an obstacle to weight gain in refeeding. Interconnections among caloric restriction, psychopathology, and physical activity in humans with AN are poorly investigated. METHODS: Physical activity and food restriction during the last 3 months and status of body image/slimness ideal were assessed by the Structured Interview of Anorexia and Bulimia Nervosa (SIAB) in 30 adolescent patients with acute AN at admission to inpatient treatment. Anxiety, depression, and obsessive-compulsiveness were assessed with the Symptom Check-List-90-Revised (SCL-90-R). A regression model based on the independent variables body mass index, food reduction, body image/slimness ideal, anxiety, depression, and obsessive-compulsiveness was calculated to determine the relevant prediction variables of physical activity. RESULTS: The regression model explained 64% (R(2) = .64, p = .000) of the variance of physical activity. Only food restriction (p = .006) and anxiety (p = .004) contributed significantly to the variance. DISCUSSION: Our results indicate that anxiety symptoms and food restriction synergistically contribute to increased levels of physical activity in the acute phase of AN. PMID- 15282687 TI - Impulsivity as a risk factor for eating disorder behavior: assessment implications with adolescents. AB - OBJECTIVE: The aims of the current study were to determine if impulsivity serves as a risk factor for eating disorder behavior and to examine whether different risk outcomes are obtained depending on the assessment strategy used to measure impulsivity. METHOD: Three independent studies are reported, each of which examined the relationship of impulsivity and eating disorder behavior in a prospective longitudinal design with adolescent subjects recruited from both public and private schools. Individuals displaying eating disorder behavior at initial assessments were not included in the analyses, to ensure that we were testing the role of impulsivity in the onset of eating disorder behavior. RESULTS: Trait impulsivity, measured with traditional personality scales, failed to predict the onset of eating disorder behavior in all three studies. However, when behavioral constructs associated with impulsivity, such as delinquency or substance abuse, were examined, they significantly predicted the onset of eating disorder behavior in most of the analyses conducted. DISCUSSION: These results provide moderate support for the idea that impulsivity serves as a risk factor for the onset of eating disorder behavior. However, this is only true when more objective behavioral measures were utilized. PMID- 15282688 TI - Comparison of the child and parent forms of the Questionnaire on Eating and Weight Patterns in the assessment of children's eating-disordered behaviors. AB - OBJECTIVE: The assessment of eating-disordered behaviors in middle childhood is challenging. Frequently, both child and parents are queried about the child's eating behavior. However, no direct comparisons between parent and child reports of child eating disturbance have been published. We compared results from the adolescent and parent versions of the Questionnaire on Eating and Weight Patterns (QEWP-A and QEWP-P, respectively) in a nontreatment sample of overweight and normal weight children. METHOD: The QEWP-A and QEWP-P were administered to 142 overweight (body mass index [BMI] > or = 85th percentile) and 121 normal weight (BMI 15th-84th percentile) children, age 9.7 +/- 1.9 years, recruited from the community. RESULTS: The QEWP-A and QEWP-P showed good agreement for the absence of eating-disordered behavior but were not concordant in terms of the number or type of binge eating, overeating episodes, or compensatory weight control behaviors in the past 6 months. Children categorized by their own reports (QEWP A) as engaging in no overeating, simple overeating, or binge eating behaviors did not differ significantly in body composition or in eating and general psychopathology. Children categorized according to their parents' reports (QEWP P) as engaging in binge eating had significantly greater body adiposity, eating disordered cognitions, body dissatisfaction, and parent-reported problems (all ps <.001) than children engaging in no overeating or simple overeating according to the QEWP-P. DISCUSSION: Child and parent reports of eating behaviors are not concordant regarding the presence of binge eating or compensatory behaviors. Further investigation of the utility of these questionnaires is needed before either can serve as a surrogate for a clinical interview. PMID- 15282689 TI - Temporal stability of the Eating Disorder Examination Questionnaire. AB - OBJECTIVES: The current study examined the stability and internal consistency of the Eating Disorder Examination Questionnaire (EDE-Q) in a general population sample. METHODS: The EDE-Q was administered to a community sample of women aged 18-45 on two occasions, with a median test-retest interval of 315.0 days. RESULTS: Pearson correlations between items of the EDE-Q assessing attitudinal features of eating disorder psychopathology ranged from 0.57 for the Restraint subscale to 0.77 for the Eating Concern subscale. The stability of items addressing eating disorder behaviors was much lower, with phi coefficients for the occurrence of objective bulimic episodes, subjective bulimic episodes, and use of exercise as a compensatory behavior of 0.44, 0.24, and 0.31, respectively, and Kendall's tau b correlations of 0.44, 0.28, and 0.31, respectively, for the frequency of these behaviors, across occasions. The internal consistency of the EDE-Q was high, with a Cronbach alpha coefficient for the global scale of 0.93, compared with a value of 0.90 for the Eating Disorder Examination interview. DISCUSSION: Items of the EDE-Q assessing attitudinal features of eating disorder psychopathology demonstrate a high degree of temporal stability, whereas the stability of items addressing eating disorder behaviors is much lower. In the case of compensatory eating disorder behaviors, low stability is likely to reflect actual trait variation, whereas the low stability of binge eating behaviors, in particular subjective bulimic episodes, is likely to reflect both trait variation and measurement error. The high internal consistency of EDE-Q items supports its use as a screening instrument in two-phase epidemiologic studies. PMID- 15282690 TI - Daily stress, coping, and dietary restraint in binge eating. AB - OBJECTIVE: The current study evaluated whether psychological stress, use of specific coping strategies, and trait dietary restraint would prospectively predict binge eating episodes. METHOD: After completing a baseline measure of restraint, 46 binge eating college women kept daily diaries assessing depressed affect, stress, coping, and binge eating for 30 days. RESULTS: Regardless of level of depressed mood, higher stress was associated with increased risk of same day binge eating; distraction coping was associated with increased risk of future binge eating; social support was associated with decreased risk of same-day binge eating; and women with high versus low dietary restraint showed different patterns of relationship for stress, coping, and binge eating. DISCUSSION: Vulnerability to binge eating in women who differ in terms of dietary restraint level may vary as a function of their coping responses to stress. Results highlight the complexities of stress and coping in binge eating. PMID- 15282691 TI - Similarity in young women's eating attitudes: self-selected versus artificially constructed groups. AB - OBJECTIVE: The current study explored similarity (in terms of eating attitudes, depression, and anxiety) among new versus established groups of young women. METHOD: Three hundred and thirty-two female students (living in 80 apartments) participated in the study. They were either living in a newly formed "no-choice" apartment or a "choice" apartment where they had chosen their housemates. All participants completed the Eating Disorders Inventory and the Hospital Anxiety and Depression Scale. The groups were compared using similarity indices (showing attitude spread per apartment). RESULTS: Consistent with our hypothesis, the choice groups held more similar eating-related attitudes and depression levels than the no-choice groups. Specifically, the choice groups were significantly more similar in their levels of ineffectiveness, interpersonal distrust, and social insecurity. DISCUSSION: In a similar way to depression, eating attitudes may be shared among relatively close groups of women. PMID- 15282692 TI - Bulimia symptoms and other risk behaviors during pregnancy in women with bulimia nervosa. AB - OBJECTIVE: The current study examined the change in bulimic symptoms as well as alcohol, drug, and tobacco use during pregnancy in subjects with bulimia nervosa. METHOD: A self-report questionnaire was used to collect retrospective data on eating disorder symptoms and substance use during pregnancy from 129 participants in a long-term study of bulimia nervosa. Follow-up data were collected for a total of 322 pregnancies occurring over a 10-15-year period. RESULTS: Overall, subjects reported that body dissatisfaction worsened, but binge eating and purging improved during pregnancy. However, the number of women completely abstinent from bulimic symptoms did not change significantly with pregnancy. Body esteem often worsened with pregnancy, particularly for women with active bulimia symptoms. Self-reported alcohol use significantly declined with pregnancy. DISCUSSION: In general, bulimia nervosa symptoms decreased during pregnancy, although the number of women completely abstinent did not change significantly. PMID- 15282693 TI - The associations between laxative abuse and other symptoms among adults with anorexia nervosa. AB - OBJECTIVE: The purpose of the current study was to examine the association between laxative abuse and other symptoms and features among adult patients presenting with anorexia nervosa. METHOD: One hundred and seventeen patients with anorexia nervosa were studied. Laxative abusers and nonabusers were compared. RESULTS: Compared with nonabusers, laxative-abusing patients had higher ratings on the Ineffectiveness, Body Dissatisfaction, and Drive for Thinness subscales on the Eating Disorders Inventory (EDI), as well as more depressive and somatization symptoms. There was an association between laxative abuse and low self-esteem. DISCUSSION: Laxative abuse appears to be associated with especially severe psychopathology and low self-esteem among subjects with anorexia nervosa. PMID- 15282694 TI - Associations among aspects of impulsivity and eating factors in a nonclinical sample. AB - OBJECTIVE: Impulsivity has been well documented as a factor in the behavior of eating-disordered populations and is likely to influence normal eating patterns as well. The current study attempted to clarify the relationship between the three elements of impulsivity (nonplanning, attentional, and motor) and the three factors of eating (cognitive restraint, disinhibition, and hunger) in a nonclinical sample. METHOD: Data were collected from a sample (N = 112) of volunteer participants from the community who answered two self-report instruments related to impulsivity (the Barratt Impulsiveness Scale, Version 11) and eating (the Eating Inventory). Partial correlations were performed on the data to control for age, sex, and education. Cognitive restraint did not correlate with any element of impulsivity. Disinhibition was positively correlated with both attentional impulsivity (r = .40, p < .001) and motor impulsivity (r = .32, p < .01). Attentional impulsivity was also positively correlated with hunger (r = .24, p < .05). DISCUSSION: The lack of association between cognitive restraint and impulsivity suggests that they are functionally distinct. Disinhibition is most closely associated with impulsivity, consistent with findings from clinical samples. Further clarification of the relationship between impulsivity and eating in nonclinical populations could facilitate a better understanding of the relationship between personality variables and normal eating behavior. PMID- 15282695 TI - Pancreatitis causing death in bulimia nervosa. AB - OBJECTIVE: We report the case of a 19-year-old woman with bulimia nervosa who died of acute hemorrhagic pancreatitis. Our objective is to raise awareness that because the symptoms of both conditions are very similar, the pre-existence of an eating disorder should not distract physicians from the possibility that potentially lethal acute pancreatitis may coexist. METHOD: The study includes autopsy results and a review of the literature. RESULTS: Pancreatitis usually presents with abdominal pain, nausea, and vomiting. DISCUSSION: In patients with eating disorders who may already have exhibited these symptoms pancreatitis may not be considered. Elevated serum amylase values may occur in subjects with bulimia nervosa without pancreatitis. If the serum amylase value is elevated, pancreatitis can be confirmed by measuring the levels of serum lipase, trypsinogen, pancreatic isoenzyme of amylase, or by abdominal computerized tomography (CT). PMID- 15282696 TI - Cerebellar atrophy in a patient with anorexia nervosa. AB - Reversible cerebral atrophy (pseudoatrophy) is observable in patients with anorexia nervosa. However, it is extremely rare to see marked cerebellar atrophy. OBJECTIVES: We report on a patient who developed cerebellar atrophy after the severe deterioration of cardiac and respiratory functions resulting from undernutririon. RESULTS: A 30-year-old Japanese woman was admitted to the Wakayama Medical University Hospital (Wakayama, Japan) because of unsteadiness of gait. She had a 7-year history of anorexia nervosa and had been admitted to an emergency hospital because of asthenic shock resulting from severe undernutrition at the age of 28. On admission to our hospital, neurologic examination revealed dysarthria and cerebellar ataxia of the trunk and lower extremities without nystagmus. A brain magnetic resonance imaging scan demonstrated marked atrophy of the cerebellum. DISCUSSION: Because her cerebellar ataxia appeared during severe deterioration of her general condition, and there has been no subsequent progression, it is possible that her cerebellar atrophy was induced by undernutrition. PMID- 15282697 TI - Jewish ethnicity and Black race: contrasting influences on the prognosis of breast cancer. AB - BACKGROUND AND OBJECTIVES: For reasons that are incompletely understood, race and ethnicity are associated with differing mortality from breast cancer. This investigation was undertaken to evaluate racial and ethnic influences on survival of women with breast cancers and to identify influential factors. METHODS: A population of 766 racially and ethnically defined women with newly diagnosed invasive breast cancers in a clinical practice were prospectively evaluated and observed on a continuing basis. Survivals were compared with multiple demographic and clinicopathologic variables. RESULTS: Compared to White, non-Jewish patients, no significant difference was found in survival of Jewish patients. Stage of cancers at diagnosis and measures of tumor biology were also comparable. By contrast, the survival of Black patients was significantly worse than that of White patients. Advanced tumor stage at diagnosis, unfavorable tumor biology and poor ancillary health all appeared to contribute to the poor survival of Black women. CONCLUSION: No adverse influence of Jewish ethnicity was found on the prognosis of breast cancer. The poor prognosis of Black women compared to White women appeared to be the result of multiple untoward influences. PMID- 15282698 TI - Esophageal cancer: outcomes of surgery, neoadjuvant chemotherapy, and three dimension conformal radiotherapy. AB - Neoadjuvant chemotherapy and radiation are being utilized with increasing frequency in the multimodal treatment of esophageal cancer, although their effects on morbidity, mortality, and survival remain unclear. The objective of this study was to determine the outcome of multimodal treatment in patients with localized esophageal cancer treated at a single institution. Between 1995 and 2002, 118 patients underwent treatment for localized esophageal cancer, utilizing surgery alone, chemoradiation alone, or surgery following neoadjuvant chemoradiation. There was no statistically significant difference in morbidity, mortality, or length of stay between the patients who received multimodal therapy when compared to surgery alone. A surgical resection after down-staging was possible in 9 out of 28 patients (32%) with a clinically non-resectable tumor (T4 or M1a). Forty-seven percent of the patients who received neoadjuvant therapy had a complete pathologic response with a 3-year survival of 59% as compared to only 20 months in those patients who did not achieve a complete response (P = 0.037). Neoadjuvant chemotherapy administered concomitantly with conformal radiotherapy can be performed safely in the treatment of esophageal cancer, without increasing the operative morbidity, mortality, or length of stay. The higher complete response rates to neoadjuvant treatment (as compared to other reports) may be due to the use of three-dimensional conformal radiation therapy or the novel use of weekly carboplatin and paclitaxel. PMID- 15282699 TI - IV-DSA is a new diagnostic tool for axillary lymph node metastasis in breast cancer patients. AB - BACKGROUND AND OBJECTIVE: The aim of this study is to know whether intravenous digital subtraction angiography (IV-DSA) is useful to detect axillary lymph node metastasis of breast cancer and to evaluate the anigiogenesis of lymph nodes in the axilla. PATIENTS AND METHODS: Forty three primary breast cancer patients (N0: 26 cases, N1: 5 cases, N2: 2 cases) who underwent IV-DSA between January and November 2000 were included in the study. Infinix CB apparatus (Toshiba, Japan) was used to collect IV-DSA images and when a mass became stained in the axilla, it was considered to be metastatic. The angiogenesis was studied by examining microvessel density (MVD) after lymph node immunostaining for factor VIII. Primary tumor was detected by IV-DSA in all 43 cases. RESULTS: Axillary lymph node metastases were detected by IV-DSA in 34.9% of cases (15/43), and by pathology in 37.2% (16/43). The sensitivity, specificity, and accuracy of the diagnostic method were 75.0% (12/16), 88.9% (24/27), and 83.7% (36/43), respectively. MVD, calculated after immunostaining for factor VIII, was significantly lower in the in metastatic region of lymph nodes identified by DSA (88.5 +/- 35.0) than in metastasis-free lymph nodes (141.1 +/- 34.0, P < 0.0001). CONCLUSIONS: IV-DSA is useful in the diagnosis of axillary lymph node metastasis of breast cancer. Our results suggest that the primary factors involved in the mechanism of DSA display may be different from high/low MVC values. PMID- 15282700 TI - Intrahepatic cholangiocarcinoma diagnosed preoperatively as hepatocellular carcinoma. AB - BACKGROUND: Some cases of mass-forming intrahepatic cholangiocarcinoma (ICC) are diagnosed as hepatocellular carcinoma (HCC) based on preoperative imaging and clinical findings. We investigated the backgrounds of such cases. METHODS: Sixty seven patients with mass-forming ICC underwent surgery from 1980 to 2002. Twenty four of these patients received a diagnosis of HCC preoperatively. We compared the group diagnosed as HCC and that diagnosed as ICC. ICC was diagnosed histopathologically in all 67 patients. RESULTS: The specific clinical findings included high rates of associated hepatitis C virus infection, high levels of serum alpha fetoprotein, lower levels of serum CA19-9, small dimension of the tumor, hypervascular staining on angiography or computed tomography, lower rates of lymph node metastasis, and high rates of HCC occurrence in the group diagnosed as HCC. None of the patients underwent extrahepatic bile duct resection and most patients did not undergo lymph node dissection in the group diagnosed as HCC. The rates of mucus secretion and the ductal expression of mucin core protein-1 (MUC1) were significantly different between the subgroups. The cumulative survival rates were significantly better in the group diagnosed as HCC than in the group diagnosed as ICC. CONCLUSION: Patients with ICC given a preoperative diagnosis of HCC had distinct clinical features and could be treated with the same operation as HCC patients. PMID- 15282702 TI - Extent of hepatic resection does not correlate with toxicity following adjuvant chemotherapy. AB - BACKGROUND: In patients with liver metastases from colorectal cancer, survival can be increased by hepatic resection. Treatment with hepatic arterial infusion (HAI) and systemic chemotherapy following resection may further increase survival and decrease recurrence, but may also result in hepatic and systemic toxicity. This article will address the question of whether large hepatic resections resulting in a greater loss of healthy liver predisposes patients to developing toxicity from the subsequent chemotherapeutic regimens. DESIGN: Retrospective analysis of 88 patients who underwent liver resection of colorectal metastases followed by adjuvant HAI and systemic chemotherapy and whose computerized tomography (CT) scans were done at Memorial Sloan-Kettering Cancer Center (MSKCC). Liver volumes were calculated from CT scans and used to determine the percentage change in healthy liver volume between the pre- and post-operative CT scans. Hepatic and systemic toxicities were defined according to the Common Toxicity Criteria of the National Cancer Institute. RESULTS: Patients experienced a mean percentage decrease in healthy liver tissue of 17% (range: 57% decrease to 32% increase) at an estimated 1 month after resection and at the initiation of chemotherapy. In a logistic regression model using percentage change in the healthy liver volume as a continuous variable, no significant association was revealed between percentage of healthy liver resected and diarrhea (P = 0.47), leukopenia (P = 0.37), neutropenia (P = 0.31), high bilirubin (P = 0.27), or alkaline phosphatase (P = 0.79). CONCLUSIONS: A greater loss of healthy liver following resection of hepatic metastases from colorectal cancer does not seem to predispose to the development of toxicity from adjuvant HAI and systemic chemotherapy. PMID- 15282703 TI - Incidentally discovered gallbladder cancer: role of cryotherapy. AB - BACKGROUND AND OBJECTIVES: Carcinoma of the gallbladder is the most common malignancy of the biliary tree. There is no standard surgical approach to the treatment of gallbladder cancer. Extended cholecystectomy with partial or complete hepatic lobectomy has resulted in high morbidity and poor long-term survival. This report describes cryotherapy of the gallbladder fossa as an alternative to liver resection in the treatment of gallbladder cancer. METHODS: A group of patients underwent cryotherapy to the gallbladder fossa for local control of gallbladder cancer. Portal lymph node dissection was performed at the same time and patients with positive lymph nodes received post-operative chemotherapy and radiation; negative node patients were observed. RESULTS: Seven patients underwent surgery. Gallbladder cancer had been incidentally found in all patients. Median hospital stay was 4 days. Five patients had positive nodes. Complications included biliary stricture, pleural effusion, congestive heart failure exacerbation, and pulmonary embolus. Mean disease-free survival for patients with T2 tumors was 11.7 months. Mean disease-free survival for patients with T3 tumors was 16.3 months. Mean disease-free survival for lymph node positive patients was 14.8 months compared to 13 months for node negative patients. No local recurrences have been detected in the gallbladder fossa. CONCLUSIONS: This study suggests cryotherapy to the gallbladder fossa done concomitantly with portal lymph node dissection may represent a safe and effective alternative to hepatic resection in the treatment of gallbladder cancer. PMID- 15282704 TI - Vascular endothelial growth factor in esophageal cancer. AB - Vascular endothelial growth factor (VEGF) plays a crucial role in angiogenesis of many solid malignancies. The influence of angiogenesis and VEGF expression on progression and recurrence of esophageal cancer has been investigated over the last years. This article reviews the prognostic significance of VEGF expression, microvessel density (MVD), and lymphangiogenic factors in squamous cell carcinoma (SCC), Barrett's dysplasia, and adenocarcinoma (AC) of the esophagus, their predictive value for treatment response to chemo-radiotherapy and new anti angiogenic treatment strategies. PMID- 15282705 TI - Commentary on Barranger et al., Contralateral axillary lymph node drainage in breast cancer: a case report: unusual lymph drainage pathways in patients with breast cancer. PMID- 15282706 TI - Re: Gipponi M, Solari N, Di Somma FC, et al.: New fields of application of the sentinel lymph node biopsy in the pathologic staging of solid neoplasms: review of literature and surgical perspectives. J Surg Oncol 2004: 85:171-179. PMID- 15282707 TI - Identification of one of the least well understood 5-HT receptors (5-HT5A) in the spinal cord. PMID- 15282708 TI - 5-HT5A receptor localization in the rat spinal cord suggests a role in nociception and control of pelvic floor musculature. AB - The 5-HT5A receptor is a seven-transmembrane receptor negatively coupled to adenylate cyclase, whose activation opens K+ channels. The 5-HT5A receptor may thus exert an inhibitory effect on neuronal activity. However, the function of this receptor is still largely unknown, in particular at the spinal level, and this is partly due to lack of specific ligands. Immunocytochemistry using specific anti-5-HT5A antibodies reveals a particularly dense labeling in the two superficial layers of the dorsal horn, suggesting that the 5-HT5A receptor may be involved in the spinal modulation of pain. In addition, a very intense staining in the lumbar dorsolateral nucleus (Onuf nucleus) in both males and females suggests that the 5-HT5A receptor is also involved in micturition through the control of urethral sphincter muscles. Colchicine pretreatment allows the staining of numerous cell bodies in lamina II. Fewer labeled cell bodies are seen in laminae I and III-VI, in the lateral spinal nucleus, and in lamina X. Electron microscope examination of 5-HT5A receptor immunoreactivity in spinal cords from untreated animals confirmed the postsynaptic labeling in all regions studied (dorsal horn, dorsolateral nucleus, and lamina X). The morphological heterogeneity of labeled dorsal horn cell bodies suggests that they belong to functionally distinct neurons (projection neurons and interneurons). In the lumbar dorsolateral nucleus, the labeling is preferentially localized on dendrites, suggesting that in this nucleus 5-HT preferentially acts at the dendritic level. Finally, the dense labeling of postsynaptic specializations suggests that the receptor may be in stock before being addressed to the synaptic differentiation. PMID- 15282709 TI - Amygdaloid inputs define a caudal component of the ventral striatum in primates. AB - The ventral striatum mediates goal-directed behavior through limbic afferents. One well-established afferent to the ventral striatum is the amygdaloid complex, which projects throughout the shell and core of the nucleus accumbens, the rostral ventromedial caudate nucleus, and rostral ventromedial putamen. However, striatal regions caudal to the anterior commissure also receive inputs from the amygdala. These caudal areas contain histochemical and cytoarchitectural features that resemble the shell and core, based on our recent studies. Specifically, there is a calcium binding protein (CaBP)-poor region in the lateral amygdalostriatal area that resembles the "shell." To examine the idea that the caudal ventral striatum is part of the "classic" ventral striatum, we placed small injections of retrograde tracers throughout the caudal ventral striatum/amygdalostriatal area and charted the distribution of specific amygdaloid inputs. Amygdaloid inputs to the CaBP-poor zone in the lateral amygdalostriatal area arise from the basal nucleus, the magnocellular subdivision of the accessory basal nucleus, the periamygdaloid cortex, and the medial subdivision of the central nucleus, resembling that of the shell of the ventral striatum found in our previous studies. There are also amygdaloid inputs to CaBP positive areas outside the shell, which originate mainly in the basal nucleus. Taken together, the "limbic-related" striatum forms a continuum from the rostral ventral striatum through the caudal ventral striatum/lateral amygdalostriatal area based on histochemical and cellular similarities, as well as inputs from the amygdala. PMID- 15282710 TI - Mesostriatal and mesolimbic projections of midbrain neurons immunoreactive for estrogen receptor beta or androgen receptors in rats. AB - The dopamine (DA) inputs to the caudate putamen, the nucleus accumbens, and the amygdala in rats are sensitive to circulating estrogens and androgens. One mechanism for the hormone modulation of these systems may be via actions at cognate intracellular estrogen and androgen receptors. However, although it is known that specific subsets of midbrain DA neurons are immunopositive for estrogen receptor beta (ERbeta) or androgen receptors (ARs), it is not known where these receptor-bearing cells project. To address this issue, we combined double-label immunocytochemistry with retrograde tract tracing to identify the forebrain projections of ERbeta- or AR-immunoreactive (IR) midbrain neurons. Specifically, Fluoro-Gold and/or cholera toxin were injected into discrete subregions of the caudate-putamen, the nucleus accumbens, or the amygdala. Evaluations of the resultant midbrain labeling revealed that ERbeta-IR neurons sent collateral projections mainly to both the ventral caudate-putamen and the amygdala, but not to the dorsal caudate or nucleus accumbens. In contrast, AR-IR neurons projected either to the amygdala or the nucleus accumbens but not to the caudate-putamen. The organization of these forebrain projections concurs with some of the known hormone sensitivities of mesostriatal and mesolimbic DA systems in rats and provides an anatomical model that predicts separate influences for androgens and estrogens over mesostriatal and mesolimbic DA systems. PMID- 15282711 TI - Anatomical and functional organization of somatosensory areas of the lateral fissure of the New World titi monkey (Callicebus moloch). AB - The organization of anterior and lateral somatosensory cortex was investigated in titi monkeys (Callicebus moloch). Multiunit microelectrode recordings were used to identify multiple representations of the body, and anatomical tracer injections were used to reveal connections. (1) Representations of the face were identified in areas 3a, 3b, 1, S2, and the parietal ventral area (PV). In area 3b, the face was represented from chin/lower lip to upper lip and neck/upper face in a rostrocaudal sequence. The representation of the face in area 1 mirrored that of area 3b. Another face representation was located in area 3a. Adjoining face representations in S2 and PV exhibited mirror-image patterns to those of areas 3b and 1. (2) Two representations of the body, the rostral and caudal ventral somatosensory areas (VSr and VSc), were found in the dorsal part of the insula. VSc was roughly a reversal image of the S2 body representation, and VSr was roughly a reversal of PV. (3) Neurons in the insula next to VSr and VSc responded to auditory stimuli or to both auditory and somatosensory stimuli. (4) Injections of tracers within the hand representations in areas 3b, 1, and S2 revealed reciprocal connections between these three areas. Injections in areas 3b and 1 labeled the ventroposterior nucleus, whereas injections in S2 labeled the inferior ventroposterior nucleus. The present study demonstrates features of somatosensory cortex of other monkeys in titi monkeys, while revealing additional features that likely apply to other primates. PMID- 15282713 TI - Projections from the amygdaloid complex to the piriform cortex: A PHA-L study in the rat. AB - Projections from the amygdala to the piriform cortex are proposed to provide a pathway via which the emotional system can modulate the processing of olfactory information as well as mediate the spread of seizure activity in epilepsy. To understand the details of the distribution and topography of these projections, we injected the anterograde tracer Phaseolus vulgaris-leucoagglutinin into different nuclear divisions of the amygdaloid complex in 101 rats and analyzed the distribution and density of projections in immunohistochemically processed preparations. The heaviest projections from the amygdala to the piriform cortex originated in the medial division of the lateral nucleus, the periamygdaloid and sulcal subfields of the periamygdaloid cortex, and the posterior cortical nucleus. The heaviest terminal labeling was observed in layers Ib and III of the medial aspect of the posterior piriform cortex. Lighter projections to the posterior piriform cortex originated in the dorsolateral division of the lateral nucleus, the magnocellular and parvicellular divisions of the basal and accessory basal nuclei, and the anterior cortical nucleus. The projections to the anterior piriform cortex were light and originated in the dorsolateral and medial divisions of the lateral nucleus, the magnocellular division of the basal and accessory basal nuclei, the anterior and posterior cortical nuclei, and the periamygdaloid subfield of the periamygdaloid cortex. The results indicate that only selective amygdaloid nuclei or their subdivisions project to the piriform cortex. In addition, substantial projections from several amygdaloid nuclei converge in the medial aspect of the posterior piriform cortex. Via these projections, the amygdaloid complex can modulate the processing of olfactory information in the piriform cortex. In pathologic conditions such as epilepsy, these connections might provide pathways for the spread of seizure activity from the amygdala to extra-amygdaloid regions. PMID- 15282712 TI - Distribution of vasoactive intestinal peptide and pituitary adenylate cyclase activating polypeptide receptors (VPAC1, VPAC2, and PAC1 receptor) in the rat brain. AB - To examine the distributions of VIP/PACAP receptors (VPAC1, VPAC2, and PAC1 receptors) in the brain and to identify the cell types that express these receptors, we performed immunohistochemistry and double immunofluorescence in the rat brain with specific antibodies. The immunohistochemistry revealed that the receptors had distinctive, complementary, and overlapping distribution patterns. High levels of the VPAC1 receptor were expressed in the cerebral cortex, hippocampal formation, deep cerebellar nuclei, thalamus, hypothalamus, and brainstem. The VPAC2 receptors were concentrated in the cerebral cortex, hippocampal formation, amygdalar regions, cerebellar cortex, deep cerebellar nuclei, hypothalamus, and brainstem. On the other hand, the PAC1 receptors had a more restricted distribution pattern in the brain, and high levels of the PAC1 receptors were confined to the cerebellar cortex, deep cerebellar nuclei, epithalamus, hypothalamus, brainstem, and white matter of many brain regions. Also, many fibers expressing the PAC1 receptors were observed in various areas, i.e., the thalamus, hypothalamus, and brainstem. The double immunofluorescence showed that the VIP/PACAP receptors were confined to the neuroglia as well as the neurons. All three types of the VIP/PACAP receptors were expressed in the astrocytes, and the PAC1 receptors were also expressed in the oligodendrocytes. These findings indicate that VIP and PACAP exert their functions through their receptors in specific locations in different combinations. We hope that this first demonstration of the distributions of the VIP/PACAP receptors provides data useful in the investigation of the mechanisms of the many functions of VIP and PACAP in the brain, which require further elucidation. PMID- 15282714 TI - Tyrosine hydroxylase expression is affected by sexual vigor and social environment in male Cnemidophorus inornatus. AB - Although the distribution of catecholamine-synthesizing cells has been described for a variety of taxa, less is known about the functional significance of particular populations in nonmammalian species, especially reptiles. To understand the role of these populations in the display of social behaviors in lizards, we studied the interactive effects of sexual vigor (sexually vigorous vs. sluggish) and social condition (housing in isolation vs. with females) on the number and somal areas of cells expressing tyrosine hydroxylase (TH), a rate limiting enzyme in catecholamine synthesis, in male whiptail lizards, Cnemidophorus inornatus. We found that, regardless of social condition, sexually vigorous males had more TH-immunoreactive (TH-ir) cells in the dorsal hypothalamus (DH) relative to sluggish males. Sexually vigorous males also had more TH-ir cells in the substantia nigra pars compacta (SNpc), but this difference was significant only among males housed with females. Sexually vigorous males that had been housed with females had smaller TH-ir cells in the preoptic area (POA) than vigorous males housed in isolation. On the other hand, no significant differences were found in the anterior hypothalamus. These results highlight the regional heterogeneity in the plasticity of TH expression and suggest that, just as in other species, the DH, SNpc, and POA might be involved in the expression of social behaviors and in behavioral plasticity following social experiences in lizards. PMID- 15282716 TI - Fine-needle sampling in malignant phyllodes tumors: clinicopathologic study of 22 cases seen at the Institut Curie. AB - The preoperative cytological diagnosis of malignant phyllodes tumor (MPT) is challenging due to the heterogeneity of its clinical, radiological, and morphological presentation. To better define the cytopathological characteristics of MPT, we reviewed 22 examples seen at the Institut Curie. The original cytologic diagnosis was benign breast tumor in four cases (18.2%), suspicious in seven cases (31.8%) (low-grade phyllodes tumor in six cases needing histological evaluation, suspicious of sarcoma in one case), and malignant in 11 cases (50%). Smears were composed of different proportions of clusters of epithelial cells (68.2%), phyllodes fragments (31.8%), spindle cells within stromal tissue (31.8%), isolated spindle-shaped or round cells (45.5%), and bipolar naked nuclei (22.7%). Giant cells and mitotic figures were also occasionally seen. The cytological findings on smears were correlated with histopathological observations. One of the difficulties to reach an accurate cytological diagnosis for MPT is the frequent overwhelming of clearly malignant sarcomatous cell by the presence of largely predominant clusters of epithelial cell. PMID- 15282717 TI - Significance of MiB-1 staining in smears with atypical glandular cells. AB - MiB-1 immunostaining may facilitate recognition of developing adenocarcinoma in cervical smear screening. In a retrospective analysis of prospectively collected data of 170 patients with atypical endocervical glandular cells and with repeat smears, archival Papanicolaou-stained smears were restained for MiB-1 and classified for the presence of preneoplastic changes of MiB-1 positive epithelial fragments. The results of classification based on MiB-1 positive epithelial fragments corresponded "roughly" with cytomorphological diagnoses. Of the 38 patients in which the primary smear was found to be MiB-1-positive, 12 of these persisted in the repeat smear. Thirty-eight of the 132 repeat smears of patients originally negative for MiB-1 pathology were positive. Of the total 50 MiB-1 positive repeat smears, four showed adenocarcinoma in situ on cytomorphological grounds. MiB-1 staining enhances detection of (pre)neoplastic changes. This approach does not destroy the morphology of the original smear and can be applied to routine material. PMID- 15282718 TI - Diagnostic limitations in testicular cytopathology: to what extent is fine-needle aspiration reliable for the diagnosis of epidermoid cyst of the testis? AB - This article describes the cytologic and histologic findings of a epidermoid cyst of the testis diagnosed by means of fine-needle aspiration (FNA) cytology. The gross and cytologic features are creamy aspirate, squamous cells, squamae, and fragments of granulomatous tissue. The cytologic features are fairly typical and similar to those observed in cutaneous epidermoid cysts; however, in this setting, the differential diagnosis should be carried out mainly with teratoma and dermoid cysts. The patient's age and precise location of the mass are paramount in the differential diagnosis. We believe that FNA is a reliable tool for the diagnosis of testicular epidermoid cysts, but the differential diagnosis with dermoid cysts should be based on histology. PMID- 15282719 TI - Cytologic diagnosis of invasive lobular carcinoma: factors associated with negative and equivocal diagnoses. AB - Fine-needle aspiration biopsy (FNAB) of invasive lobular carcinoma (ILC) is associated with notoriously high rates of false negative and equivocal diagnoses. To identify causative factors, we reviewed the cytologic features of presurgical FNAB smears of ILC and correlated the cytologic findings with the number of passes, tumor size, mammographic findings, and the histologic characteristics of the tumor. Smear cellularity, presence of single intact epithelial cells, nuclear size, nuclear atypia, palpability of the tumor, and histologic type of ILC (classic versus nonclassic) were statistically significant in establishing an unequivocally positive diagnosis. We also found that the cytologic cellularity of the lesion does not reflect the actual cellularity of the tumor but instead is an indicator of the architectural arrangement of the neoplastic cells; tumors that form epithelial cell groups, such as in nonclassic ILC, tend to yield more cellular aspirates that are diagnostic for carcinoma. In contrast, classic ILC, in which single neoplastic cells are embedded in fibrous stroma, is more likely to yield a paucicellular smear with subtle atypia and rare single intact epithelial cells. As such, an inconclusive diagnosis in a certain percentage of classic ILC cases may be unavoidable. PMID- 15282720 TI - Effectiveness of AutoPap system location-guided screening in the evaluation of cervical cytology smears. AB - Location-guided screening is a feature of the AutoPap primary screening software. Areas of the smear most likely to contain an abnormality are identified for prompt review by the cytotechnologist, thereby facilitating diagnostic accuracy and reducing laboratory workload. A two-armed retrospective study comprising 6,000 conventional smears was undertaken to compare this approach with the current practice of full manual screening of conventional smears. Discrepant diagnoses between the two study arms were subject to an internal discrepancy review process to determine the final truth diagnosis. Analysis of the results show that AutoPap location-guided screening is at least equivalent to current practice when detecting high-grade or suspected high-grade smears. However, the device does not detect low-grade abnormalities, unsatisfactory smears, an endocervical component or organisms, as well as standard screening. The device also assigns a numerical score to each slide, with abnormal smears allocated a higher rank. Slide ranking was found to be of value in triaging abnormal smears for prompt screening and reporting. The performance of the primary screening software was found to be comparable to previous studies, with the majority of abnormal smears being selected by the instrument for manual review. PMID- 15282721 TI - Cyst fluid analysis for the differential diagnosis of pancreatic cysts. AB - Pancreatic cystic neoplasms comprise a pathologically heterogeneous group with many shared clinical features. We assessed the reliability of cyst fluid analysis for the differential diagnosis of pancreatic cysts. Cyst fluid was obtained by fine-needle aspiration from 78 pancreatic cysts. The lesions studied consisted of 17 mucinous cystic tumors (MCTs), 13 serous cystadenomas (SCAs), 5 solid pseudopapillary tumors (SPTs), 8 intraductal papillary mucinous tumors (IPMTs), 6 ductal adenocarcinomas (ACAs) with cystic degeneration, and 29 pseudocysts (PCs). Epithelial cells were observed in 27 (81%) of 33 successful aspirates of cystic neoplasms. Cytologic diagnosis was possible in 5 (31%) out of 16 MCTs. Mucicarmine staining was positive in five out of nine MCTs, one out of one ACA, and one out of two IPMTs, but in none of the SCAs, SPTs, or PCs. Cyst fluid carcinoembryonic antigen (CEA) levels of more than 467 ng/mL had a 87% sensitivity and a 98% specificity for detecting MCTs, and amylase levels of more than 479 U/L had a 73% sensitivity and a 90% specificity for detecting PCs. In conclusion, cyst fluid analysis for cytology, mucin staining, CEA, and amylase levels are useful in the differential diagnosis of pancreatic cysts. PMID- 15282722 TI - How stereotactic core-needle biopsy affected breast fine-needle aspiration utilization: an 11-year institutional review. AB - To determine the effect of stereotactic core-needle biopsy (SCNB) on the utilization of breast fine-needle aspirate (FNA) biopsy, we retrospectively reviewed 1,568 cases of breast FNAs that were obtained from 1,188 patients between the years 1990 and 2000. There were 378 positive and atypical cases and 497 negative and unsatisfactory cases in the pre-SCNB group (between 1990 and 1996; 7 years); and 225 positive and atypical cases and 468 negative and unsatisfactory cases in the post-SCNB group (between 1997 and 2000; 4 years). The average number of cases per year in the pre- and post-SCNB groups was 125 and 173, respectively. While the average positive/atypical cases per year in both groups remained relatively constant, the average negative/unsatisfactory cases per year were significantly increased in the post-SCNB group (117 in the post SCNB vs. 71 in the pre-SCNB). The increase in this group was due to a true increase in the negative diagnoses, since unsatisfactory rate decreased in the post-SCNB group (12.6% in the post-SCNB vs. 9.3% in the pre-SCNB). The sensitivity, specificity, positive predictive value, and negative predictive value were 90%, 79%, 92%, and 82% in the pre-SCNB group and 93%, 86%, 91%, and 90% for the post-SCNB group, respectively. In conclusion, the implementation of SCNB did not result in a decrease in the total number of breast FNAs; however, the distribution of cases changed. FNA is increasingly used to complete the triple test in clinically and radiographically negative cases. PMID- 15282723 TI - Echinococcal cyst of the liver. PMID- 15282724 TI - Cytologic features of central giant-cell granuloma of the jaw. AB - In this present series, we studied in detail the cytologic features of five histopathologically verified cases of central giant-cell granuloma (CGCG). All the patients in this series were female, with an age range of 11-60 years. There were three cases with involvement of the lower jaw and two cases had upper jaw involvement. Cytology smears showed dispersed single cells in the background. Nuclei of the individual cells were round to ovoid with fine chromatin and inconspicuous nucleoli. The cytoplasm of these cells was moderate in amount with indistinct cell borders. Many randomly scattered multinucleated giant cells with 10-20 nuclei were present in the background. Combination of clinical features, radiologic pictures, and cytologic features may be helpful for diagnosis of CGCG on fine-needle aspiration cytology. PMID- 15282725 TI - Fine-needle aspiration cytology of bilateral renal malakoplakia. AB - Isolated bilateral renal malakoplakia in the absence of concomitant involvement of the urinary tract is a rare occurrence. We report imaging, cytologic, and histologic findings of such a case diagnosed initially by fine-needle aspiration (FNA) cytology. A 26-yr-old female presented with pain in the left flank, fever, anorexia, and weight loss for the past 2 mo. A left lumbar lump was palpable on physical examination. Imaging studies showed an enlarged nonfunctioning left kidney and a small lesion in the right kidney with preserved function. FNA from both kidneys yielded purulent material positive for E. coli on culture. The smears were inflammatory, with a predominance of neutrophilic polymorphs and numerous histiocytes along with some intracellular and extracellular Michaelis Guttman bodies, which were highlighted with the use of a PAS stain. Histology of the nephrectomy specimen showed ill-defined nodules, composed of foamy histiocytes intermingled with neutrophils, plasma cells, and many variably sized concentric laminated bodies. The right-side lesion resolved with the use of broad spectrum antibiotics in conjunction with ascorbic acid and bethanecol. When imaging studies are suggestive of a chronic inflammatory process, renal malakoplakia must always be considered in the differential diagnosis even if aspirated material shows a predominance of polymorphonuclear leukocytes. The use of special stains like PAS, Von Kossa, and Perl's helps in reaching the correct diagnosis. PMID- 15282726 TI - Cytopathology of chalazia. AB - Chalazion is a common inflammatory condition of the eyelid, usually treated on the basis of clinical diagnosis alone. Preoperative exclusion of malignancy in chalazia with atypical clinical presentation could prevent unwarranted surgery. This is a retrospective study of aspirates from 16 patients with chalazia having an atypical clinical presentation. Smears were stained with May-Grunwald Giemsa. Two broad patterns of granulomatous inflammation reflecting the spectrum of changes in the course of the disease were seen. Nine smears had mixed-cell granulomas consisting of neutrophils, lymphocytes, plasma cells, macrophages, giant cells, and granulation tissue. Seven smears had suppurating granulomas characterized by epithelioid cell granulomas with numerous neutrophils in a proteinaceous background. Fine-needle aspiration cytology of chalazia with atypical clinical presentation provides a rapid, safe, and reliable means of documenting the diagnosis and excluding malignancy. PMID- 15282727 TI - Eosinophilic metaplastic atypia in exfoliated cells of ovarian endometriosis: a potential cytodiagnostic pitfall in peritoneal fluids. AB - Endometriosis is a protean clinical condition that is defined pathologically by the presence of endometrial glands and stroma with associated evidence of bleeding occurring outside of the usual anatomy of the endometrium/uterine cavity. It is one of the most common benign gynecologic disorders, affecting approximately 5% of all women. The prevalence of the condition may increase to as high as 30% in infertile premenopausal women. Despite extensive study, the etiology of this condition remains an enigma. Cytopathologists may encounter abnormal cells arising from endometriosis in cervicovaginal cytology samples, nongynecologic exfoliative cytology cases, needle aspiration biopsy slides, and in histologic materials and cell blocks. We report a case of eosinophilic epithelial metaplasia present in cytospins made from a peritoneal washing taken from a perimemopausal woman. This patient was concomitantly histologically documented to have ovarian surface endometriosis with similar metaplastic alteration. The case highlights the potential for misdiagnosis of such processes as being potentially malignant and stresses the need for cytohistologic correlation whenever possible. PMID- 15282728 TI - Diagnosis of granulocytic sarcoma on pleural effusion cytology: report of a case. AB - Granulocytic sarcoma is a rare manifestation of myeloid leukemia and is even rarer as a primary presentation. Granulocytic sarcoma may affect any organ and has many modes of presentation. Diagnosis by cytology is possible as long as the diagnosis is considered. Diagnosis by cytology of body cavity effusion fluid has rarely been described. We present an unusual case of granulocytic sarcoma in which pleural effusion was a part of the initial presentation. Cytologic examination of the pleural effusion was able to make the definitive diagnosis of granulocytic sarcoma. PMID- 15282729 TI - Beware of chasing crystals in sputum samples. PMID- 15282730 TI - Inflammatory fibroid polyps are not inflammatory myofibroblastic tumors. PMID- 15282731 TI - Aspiration cytology diagnosis of a primary amyloid tumor of thoracic vertebra: a case report. PMID- 15282732 TI - The role of maternal toxicity in lovastatin-induced developmental toxicity. AB - The role of maternal toxicity in lovastatin-induced developmental toxicity in rats was examined in a series of studies. The first study administered lovastatin at 100, 200, 400, or 800 mg/kg/day (mkd) orally to mated rats from Gestation Day (GD) 6 through 20. Maternal toxicity was observed as transient dose-related body weight losses at the initiation of dosing; there were also deaths and/or morbidity at 400 and 800 mkd. These toxicities occurred in conjunction with forestomach lesions. Mean fetal weights were decreased in all groups (-5 to 16%), and the incidence of skeletal malformations, variations, and incomplete ossifications was increased. The 2 highest doses produced the most severe maternal and developmental effects. Using the same dosages, the second study avoided gestational maternal weight losses and morbidity by starting treatment 14 days before mating with dosing continued to GD 20. There were transient dose related body weight losses after the start of dosing and deaths in the 400- and 800-mkd groups; however, there was no evidence of maternal toxicity during gestation. Developmental toxicity was evident only as slight, but generally significant (p< or =0.05) decreases in mean fetal weights in groups given > or =200 mkd (-2 to -5%). Significantly, no skeletal abnormalities were observed. A third study administered the pharmacologically active metabolite of lovastatin subcutaneously at dose levels that matched oral maternal drug exposures. In the high-dose group, maternal weight gain and mean fetal weight were slightly decreased but there were no treatment-related skeletal abnormalities. Finally, a series of toxicokinetic studies assessed whether the 2 different developmental toxicity profiles were due to differences in drug exposure between the developmentally toxic and non-toxic dosing regimes. The data showed that groups with no skeletal abnormalities had maternal and embryonic/fetal drug concentrations similar to or even greater than the groups with fetal abnormalities. These results indicate that fetal skeletal abnormalities observed at lovastatin dose levels > or =100 mkd are not due to a direct teratogenic effect, but are the result of excessive maternal toxicity, which most likely involves a nutritional deficiency associated with forestomach lesions and reduced maternal food intake. PMID- 15282733 TI - Effects of protein deficient diets on the developmental toxicity of inorganic arsenic in mice. AB - BACKGROUND: Inorganic arsenic, when given by injection to pregnant laboratory animals (mice, rats, hamsters), has been shown to induce malformations. Arsenic methylation may be a detoxification step, and diets deficient in protein are a poor source of methyl donors and may possibly result in impaired arsenic methylation. Human health effects from chronic arsenic exposure have been reported mainly in populations with low socioeconomic status. Individuals in such populations are likely to suffer from malnutrition, which can compromise embryonic/fetal development and diminish arsenic methylating capacity. We sought to determine if dietary protein deficiency affects the developmental toxicity of inorganic arsenic. METHODS: Mated females were randomly assigned to one of 12 treatment groups. Experimental groups received either AsIII or AsV i.p. on Gestation Day 8 (GD 8, plug=GD 0) and were maintained on a 5%, 10%, or 20% protein custom mixed diet from GD 1 until sacrifice. Controls received the custom diets alone, were given AsIII or AsV i.p. on GD 8 with Teklad LM-485 rodent diet, or were fed the LM-485 diet alone. Test females were sacrificed on GD 17, and their litters were examined for mortality and developmental defects. RESULTS: Arsenic plus dietary protein deficiency decreased maternal weight gain and increased the incidences of exencephaly, ablepharia, and skeletal defects, such as malformed vertebral centra, fused ribs, and abnormal sternebrae (bipartite, rudimentary, or unossified). CONCLUSIONS: These results demonstrate that dietary protein deficiency enhances the developmental toxicity of inorganic arsenic, possibly by impairment of arsenic methylation. PMID- 15282734 TI - Long-term alcohol exposure prior to conception results in lower fetal body weights. AB - BACKGROUND: It is well known that alcohol consumption during pregnancy can result in lower birth weight babies but many women stop consuming alcohol prior to conception as a part of pregnancy planning. The purpose of this study was to determine whether alcohol consumption prior to conception may also have an effect on fetal development. METHODS: Male and female C57BL/6J mice at 4, 6, or 8 weeks of age received either a single administration of alcohol (3.0 g/kg) via intragastric gavage (IG) each day for at least 60 days, or an isovolumetric IG administration of sterile water. After 60 treatment days, males and females within each age and treatment group were mated overnight. Females continued to receive daily alcohol treatments until conception. Males continued to receive treatments until all females were successfully mated. At conception, females were isolated and left undisturbed. On embryonic day 14, fetus number, size, and weight was determined. RESULTS: Maternal food consumption, body weight at conception, and delay to conception onset did not differ between the two treatment groups or among the three age groups. Fetal body weights did not differ among the three age groups. Fetuses from females treated with alcohol had lower body weights compared to those treated with water. Male treatments did not seem to affect fetal body weight. CONCLUSIONS: Fetal growth and development can be affected by alcohol consumption prior to the time of conception. Alcohol consumption prior to conception is a potential risk factor to fetal outcome and an important consideration for those females planning to have children. PMID- 15282735 TI - Reproductive toxicity assessment of lasofoxifene, a selective estrogen receptor modulator (SERM), in male rats. AB - BACKGROUND: Lasofoxifene is a nonsteroidal selective estrogen receptor modulator (SERM) with greater than 100-fold selectivity against all other steroid receptors and is a potentially superior treatment for postmenopausal osteoporosis. The purpose of this study was to evaluate the effects of lasofoxifene on male reproduction in rats in light of the known effects of estrogen modulating compounds on male reproductive ability. METHODS: Lasofoxifene was administered to adult male rats at doses of 0.1, 1, 10, and 100 mg/kg for 66-70 consecutive days. After 28 days of dosing, male rats were cohabited with untreated female rats. Female rats were euthanized on gestation day 14 and a uterine examination was carried out for evaluation of reproductive parameters and embryo viability. Male rats were euthanized after 66-70 days of dosing and epididymal sperm motility and concentration were assayed. The testes, epididymides, prostate, and seminal vesicles were weighed and microscopically examined. RESULTS: The duration of cohabitation was increased for 100 mg/kg males by 0.7 days. The number of males copulating and the number of implantation sites produced per copulation were reduced in the 10 and 100 mg/kg groups. Weights of the seminal vesicles and epididymides were reduced for all groups, although the testes weight and epididymal sperm motility and concentration were not affected by treatment. There were no microscopic findings in the male reproductive tissues. CONCLUSION: The changes in male fertility and reproductive tissue weights after exposure to lasofoxifene are consistent with those previously described for estrogen receptor modulating compounds. PMID- 15282736 TI - Reproductive toxicity assessment of lasofoxifene, a selective estrogen receptor modulator (SERM), in female rats. AB - BACKGROUND: Lasofoxifene is a nonsteroidal selective estrogen receptor modulator (SERM). With high affinity to the alpha and beta human estrogen receptors and greater potency than other SERMs, lasofoxifene is potentially a superior treatment for postmenopausal osteoporosis. In light of the known effects of estrogen-modulating compounds on female reproductive indices, two studies were conducted to evaluate the effects of lasofoxifene on female rat cyclicity, reproduction, and parturition. METHODS: One study evaluated effects of lasofoxifene on estrous cyclicity, and the second study assessed effects on implantation and parturition. In the cyclicity study, lasofoxifene was administered to female rats at doses of 0.1, 0.3, and 1.0 mg/kg/day for 14 consecutive days. After treatment, there was a 3-week reversibility phase followed by a mating phase. In the implantation study, lasofoxifene was administered to pregnant female rats at doses of 0.01, 0.03, and 0.1 mg/kg/day for 7 consecutive days (gestation day [GD] 0-6). Some animals were euthanized on GD 21, and the remainder of the group was allowed to deliver the F1 generation. Several developmental indices were evaluated in the F1 pups through post-natal day (PND) 21. RESULTS: In the cyclicity study, all lasofoxifene-treated females were anestrous by Study Day 7 (1.0 mg/kg) or 9 (0.3 and 0.1 mg/kg). The reversibility phase resulted in restoration of normal estrous cycles by the end of 1 (0.1 mg/kg) or 2 weeks (0.3 and 1.0 mg/kg). During the mating phase, no adverse effects occurred in pregnancy success or reproductive parameters. In the implantation study, all doses of lasofoxifene increased pre- and post implantation losses, increased gestation length, and reduced litter size. None of the developmental parameters measured on the F1 generation was adversely affected. CONCLUSION: Lasofoxifene reversibly altered the estrous cycle and inhibited implantation, consistent with what would be expected from a member of the SERM class. PMID- 15282737 TI - Embryo/fetal toxicity assessment of lasofoxifene, a selective estrogen receptor modulator (SERM), in rats and rabbits. AB - BACKGROUND: The purpose of this study was to evaluate the effects of lasofoxifene, a selective estrogen receptor modulator (SERM), on rat and rabbit fetal development. METHODS: Lasofoxifene was administered orally to rats (1, 10, 100 mg/kg) between gestation days (GD) 6-17, and in rabbits (0.1, 1, 3 mg/kg) between GD 6-18. Maternal body weight and food consumption were monitored throughout pregnancy. Fetuses were delivered by Cesarean section on GD 21 in rats, and GD 28 in rabbits, to evaluate fetal viability, weight, and morphology. Drug concentrations in maternal plasma were measured in a separate cohort of animals at several time points commencing on GD 17 (rats) and 18 (rabbits). On GD 18 (rat) and GD 19 (rabbit) drug concentrations were measured in maternal plasma and in fetal tissue 2 hr post dosing to determine the fetal to maternal drug ratio. RESULTS: In rats, there were dose-related declines in maternal weight gain and food consumption. Post implantation loss was significantly increased at dosages of 10 and 100 mg/kg, and the number of viable fetuses was decreased at 100 mg/kg. The placental weights increased, whereas fetal weights decreased in a dose-dependent manner. Lasofoxifene-related teratologic findings were noted at 10 and 100 mg/kg and included imperforate anus with hypoplastic tails, dilatation of the ureters and renal pelvis, misaligned sternebrae, hypoflexion of hindpaw, wavy ribs, and absent ossification of sternebrae. In rabbits, neither maternal weight gain nor food consumption were affected during treatment. Between GD 26-28, there was a dose-dependent increased incidence of red discharge beneath the cages. At 1 and 3 mg/kg, resorptions and post-implantation loss increased. There were no significant external or visceral effects, but 3 mg/kg there was an increased incidence of supernumerary ribs. Although the maternal plasma Cmax and AUC(0-24) were dose-dependent, the exposures in the rat were many orders of magnitude greater than in the rabbit even for the same 1 mg/kg dose. The single time point fetal/maternal drug ratio was higher in the rat (1.3-0.78) than in the rabbit (0.21-0.16). CONCLUSION: In general, both maternal and fetal effects of lasofoxifene were similar to those reported with other SERMs. Although the incidence or severity of these effects was, in some instances, greater in the rat than in the rabbit, the doses and the resultant maternal and fetal exposures were many orders of magnitude higher in the rat, suggesting the rabbit to be more sensitive to the toxicological effects of lasofoxifene. PMID- 15282738 TI - Pre- and postnatal development studies of lasofoxifene, a selective estrogen receptor modulator (SERM), in Sprague-Dawley rats. AB - BACKGROUND: Lasofoxifene is a nonsteroidal selective estrogen receptor modulator (SERM) developed for the treatment of postmenopausal osteoporosis. The purpose of these studies was to evaluate the effects of lasofoxifene on the postnatal development, behavior, and reproductive performance of offspring of female rats given lasofoxifene during organogenesis and lactation. METHODS: Two range-finding studies were conducted to determine the effects of lasofoxifene at doses from 0.01-10 mg/kg on parturition and lactation in pregnant rats and on the early postnatal development of the offspring, and to optimize the dosing regimen. Maternal milk and plasma were sampled for concentrations of lasofoxifene on Lactation Days 4, 7, and 14. In the pre- and postnatal development study, lasofoxifene was administered to pregnant and lactating rats by oral gavage at dose levels of 0.01, 0.03, and 0.1 mg/kg on Gestation Days 6-17 and Lactation Days 1-20. Maternal body weight and food consumption were measured throughout pregnancy, and body weight was measured throughout lactation. Parturition was monitored closely. The F1 offspring were measured for viability, body weight, anogenital distance, the appearance of postnatal developmental indices and reflex behaviors, sensory function, in an age-appropriate functional observational battery, motor activity, auditory startle, passive avoidance, and the Cincinnati Water Maze. The F1 generation was assessed for reproductive function, and the F2 offspring were measured for body weight and viability throughout the lactation period. RESULTS: In the range-finding studies, indications of maternal toxicity included decreased body weight and food consumption, increased length of gestation, prolonged parturition, dystocia, and increased offspring mortality at birth. Concentrations of lasofoxifene in maternal plasma were similar to those in milk, increased with increasing dose, and remained consistent over a 10-day period. In the pre- and postnatal development study, maternal body weights and food consumption were decreased in all treated groups during gestation. Length of gestation was increased, parturition was prolonged, and dystocia was noted in the dams in the 0.1 mg/kg group. There was increased pup mortality in the F1 litters in the 0.1 mg/kg group and all treated groups had decreased offspring body weights beginning at 1 week of age, continuing into the postweaning period and, for the F1 males, into adulthood. Female F1 offspring in the 0.03 and 0.1 mg/kg groups had increased body weights as adults. There were delays in the age of appearance of preputial separation in the males in the 0.1 mg/kg group and vaginal opening in the females in all treated groups. Body temperature was decreased by <0.5 degrees C after weaning for male and female offspring in the 0.1 mg/kg group. The sensory, behavioral, and functional measures, including the tests of learning and memory, were unaffected by treatment. Mating success was lower for the F1 animals in the 0.1 mg/kg group, but there were no effects on the reproductive parameters. Mating, reproduction, and maternal behavior of the F1 animals in the 0.01 and 0.03 mg/kg groups and the survival and body weights of the F2 offspring in all treated groups through Postnatal Day 21 were unaffected by treatment. CONCLUSION: The maternal findings in this study were related to the pharmacologic activity of lasofoxifene. Inhibition of growth of the F1 offspring after perinatal exposure to lasofoxifene was observed, but there were no significant effects on the sensory, behavioral, or functional measures, including learning and memory. There were no effects on the F2 generation. The findings are consistent with those reported for at least one other SERM. The findings of this study do not suggest increased risk for the primary indication of use in postmenopausal women. PMID- 15282739 TI - Postnatal closure of membranous ventricular septal defects in Sprague-Dawley rat pups after maternal exposure with trimethadione. AB - BACKGROUND: Congenital membranous ventricular septal defects (VSD) have been shown to close during postnatal development in rats [Solomon et al., Teratology 55:185-194, 1997]. Although they may differ in size, spontaneous and treatment related VSD are histologically similar; however, the postnatal fate of treatment induced VSD is not known. The objective of this study was to determine if treatment-induced VSD persist throughout postnatal development. METHODS: Groups of 40 female rats were given oral doses of trimethadione (TMD) at 400 mg/kg/day (200 b.i.d.) or 600 mg/kg/day (300 b.i.d.) on Gestation Days (GD) 9 and 10. Twenty dams in each group were designated for Cesarean section and 20 were allowed to deliver and rear their offspring to Postnatal Day (PND) 21. The integrity of the ventricular septum was evaluated in fetuses (GD 21) and pups (PND 21). RESULTS: The incidence of membranous VSD was 0.6, 7.6, and 49.8% per litter in the Control, 400, and 600 mg/kg groups, respectively, on GD 21. Both the incidence and severity of VSD increased with dose. The VSD at 400 mg/kg were small in size and initially detected by the presence of blood flowing through the defect from the closed right ventricle. In the 600 mg/kg dose group, the VSD, although still membranous, were larger and more readily detected without the need to examine the blood flow. At 600 mg/kg, not only were the VSD larger than those in the Control or the 400 mg/kg group, 10.1% per litter of the affected fetuses had other vessel anomalies associated with the VSD, which were incompatible with pup survival. On PND 21, VSD was noted in 0.3, 0, and 6.4% per litter evaluated in the Control, 400, and 600 mg/kg groups, respectively. This demonstrates that the small, isolated treatment-related VSD can resolve postnatally; however, the closure of the larger or more severe VSD may be prolonged or may not occur at all. Although TMD exposure reduced group mean fetal weights at both dose levels, there was no difference between the mean weight of fetuses with VSD and those fetuses without VSD in the same group. CONCLUSION: Treatment-induced VSD close postnatally, and appears to be a delay in cardiac development not associated with fetal weight. The timing of closure and survivability during closure is dependent on the severity of the VSD. Further characterization of the two sizes of VSD may provide diagnostic clarity; however, the current data support the smaller VSD as a variation with no significant impact on viability and growth, and the more severe VSD to be a malformation. PMID- 15282740 TI - Concern over decreased training in embryology and developmental/reproductive toxicology. PMID- 15282742 TI - Efficient temporally controlled targeted somatic mutagenesis in hepatocytes of the mouse. PMID- 15282741 TI - Folate modulates Hox gene-controlled skeletal phenotypes. AB - Hox genes are well-known regulators of pattern formation and cell differentiation in the developing vertebrate skeleton. Although skeletal variations are not uncommon in humans few mutations in human HOX genes have been described. If such mutations are compatible with life, there may be physiological modifiers for the manifestation of Hox gene-controlled phenotypes, masking underlying mutations. Here we present evidence that the essential nutrient folate modulates genetically induced skeletal defects in Hoxd4 transgenic mice. We also show that chondrocytes require folate for growth and differentiation and that they express folate transport genes, providing evidence for a direct effect of folate on skeletal cells. To our knowledge, this is the first report of nutritional influence on Hox gene-controlled phenotypes, and implicates gene-environment interactions as important modifiers of Hox gene function. Taken together, our results demonstrate a beneficial effect of folate on skeletal development that may also be relevant to disorders and variations of the human skeleton. PMID- 15282743 TI - Inner hair cell Cre-expressing transgenic mouse. AB - Cochlear hair cells of the inner ear are mechanosensory transducers critical for sound reception in mammals. A mouse with a specific expression of Cre recombinase activity in hair cells is essential for hair cell-specific gene targeting. Here we report a transgenic mouse in which Cre activity is detected in inner hair cells, not in supporting cells, in the cochlea. The Cre activity was visualized with both X-gal staining and beta-galactosidase immunostaining in progeny of a cross between our Cre line and the reporter ROSA26R line. In inner hair cells, the Cre activity started at postnatal day 14 and was maintained throughout adulthood. Starting at postnatal day 50, a few outer hair cells in the outermost row of cochlear apical and middle turns displayed the Cre activity. In vestibular hair cells and spiral ganglia, the Cre activity was also detected. Cre activity was present in cells widely distributed throughout brain, testis, and retina, but was absent in many other tissues such as kidney, heart, liver, and intestine. This Cre mouse line can thus be used for conditional gene targeting in mature inner hair cells of the cochlea. genesis 39:173-177, 2004. PMID- 15282744 TI - Transgenic mice that express Cre recombinase in osteoclasts. AB - To study the physiological control of osteoclasts, the bone resorbing cells, we generated transgenic mice carrying the Cre recombinase gene driven by either the tartrate-resistant acid phosphatase (TRAP) or cathepsin K (Ctsk) promoters. TRAP Cre and Ctsk-Cre transgenic mouse lines were characterized by breeding with LacZ ROSA 26 (R26R) reporter mice and immunohistochemistry for Cre recombinase. The Cre transgene was functional in all lines, with Cre-mediated recombination occurring primarily in the long bones, vertebrae, ribs, and calvaria. Histological analyses of the bones demonstrated that functional Cre protein was present in 1) osteoclasts (Ctsk-Cre); 2) osteoclasts, columnar proliferating, and hypertrophic chondrocytes (TRAP-Cre line 4); and 3) round proliferating chondrocytes (TRAP-Cre line 3). In conclusion, we generated transgenic mouse lines that will enable the deletion of floxed target genes in osteoclasts, which will be valuable tools for studying the regulation of osteoclast function. PMID- 15282745 TI - Tissue-specific and inducible Cre-mediated recombination in the gut epithelium. AB - We generated two complementary systems for Cre-mediated recombination of target genes in the mouse digestive epithelium and tested them with a Cre-reporter mouse strain. Cre was expressed under the control of a 9 kb regulatory region of the murine villin gene (vil-Cre). Genetic recombination was initiated at embryonic day (E) 9 in the visceral endoderm, and by E12.5 in the entire intestinal epithelium, but not in other tissues. Cre expression was maintained throughout adulthood. Furthermore, transgenic mice bearing a tamoxifen-dependent Cre recombinase (vil-Cre-ERT2) expressed under the control of the villin promoter were created to perform targeted spatiotemporally controlled somatic recombination. After tamoxifen treatment, recombination was detectable throughout the digestive epithelium. The recombined locus persisted for 60 days after tamoxifen administration, despite rapid intestinal cell renewal, indicating that epithelial progenitor cells had been targeted. The villin-Cre and villin-Cre-ERT2 mice provide valuable tools for studies of cell lineage allocation and gene function in the developing and adult intestine. PMID- 15282746 TI - Nuclear beta-catenin-dependent Wnt8 signaling in vegetal cells of the early sea urchin embryo regulates gastrulation and differentiation of endoderm and mesodermal cell lineages. AB - The entry of beta-catenin into vegetal cell nuclei beginning at the 16-cell stage is one of the earliest known molecular asymmetries seen along the animal-vegetal axis in the sea urchin embryo. Nuclear beta-catenin activates a vegetal signaling cascade that mediates micromere specification and specification of the endomesoderm in the remaining cells of the vegetal half of the embryo. Only a few potential target genes of nuclear beta-catenin have been functionally analyzed in the sea urchin embryo. Here, we show that SpWnt8, a Wnt8 homolog from Strongylocentrotus purpuratus, is zygotically activated specifically in 16-cell stage micromeres in a nuclear beta-catenin-dependent manner, and its expression remains restricted to the micromeres until the 60-cell stage. At the late 60-cell stage nuclear beta-catenin-dependent SpWnt8 expression expands to the veg2 cell tier. SpWnt8 is the only signaling molecule thus far identified with expression localized to the 16-60-cell stage micromeres and the veg2 tier. Overexpression of SpWnt8 by mRNA microinjection produced embryos with multiple invagination sites and showed that, consistent with its localization, SpWnt8 is a strong inducer of endoderm. Blocking SpWnt8 function using SpWnt8 morpholino antisense oligonucleotides produced embryos that formed micromeres that could transmit the early endomesoderm-inducing signal, but these cells failed to differentiate as primary mesenchyme cells. SpWnt8-morpholino embryos also did not form endoderm, or secondary mesenchyme-derived pigment and muscle cells, indicating a role for SpWnt8 in gastrulation and in the differentiation of endomesodermal lineages. These results establish SpWnt8 as a critical component of the endomesoderm regulatory network in the sea urchin embryo. PMID- 15282747 TI - Targeted insertion of an IRES Cre into the Hnf4alpha locus: Cre-mediated recombination in the liver, kidney, and gut epithelium. AB - The Hnf4alpha gene belongs to a family of trancriptional regulators required for liver development and function. Hnf4alpha is also expressed in other tissues, including the newly formed visceral endoderm of the early postimplantation embryo, and later in embryogenesis in the gut epithelium and the kidney. The regulatory sequences involved in controlling expression of Hnf4alpha at these diverse sites are not clearly understood. Here we used homologous recombination to introduce Cre recombinase coding sequences into the endogenous Hnf4alpha locus. Crossing Hnf4alpha(Creex2/+) mice with R26R partners allowed us to follow the pattern of Cre-mediated recombination. Our results show that recombination of the reporter allele closely follows endogenous Hnf4alpha expression, but with a slight temporal delay. Thus, the Hnf4alpha(Creex2) strain should prove useful for conditionally deleting gene activity in the liver, gut epithelium, or kidney. PMID- 15282748 TI - Thyrocyte-specific expression of Cre recombinase in transgenic mice. AB - A transgenic mouse line that expresses Cre recombinase under control of the human thyroid peroxidase (TPO) gene promoter was established. The activity and specificity of the TPO-driven Cre recombinase were examined by using Northern blotting and by crossing with the ROSA26 reporter transgenic mouse line. In the latter mice, Cre-mediated recombination occurred only in the thyrocytes, and recombination commenced around embryonic day 14.5, at the time during thyroid organogenesis when TPO expression begins. This study demonstrates that the TPO Cre transgenic mouse is a powerful tool to specifically delete loxP-inserted (floxed) genes in thyrocytes and will be of great value in the study of thyrocyte specific genes during development and/or in adult thyroids. PMID- 15282749 TI - Germline transmission and efficient DNA recombination in mouse embryonic stem cells mediated by adenoviral-Cre transduction. AB - Following gene targeting, a loxP-neo-loxP cassette was introduced into ES cells. The presence of a selectable marker such as neo in the targeted allele may result in gene interference in flox mice or unexpected phenotypes due to genetic ambiguity in direct knockout mice. Typically, the neo cassette is selectively removed by transient expression of the Cre recombinase in targeted ES cell. However, this method involves a tedious process of selecting, expanding, and screening ES cell clones which may compromise germline competency. Here, we describe a novel method of combining adenovirus-Cre mediated gene recombination with ES gene targeting to facilitate efficient loxP-neo-loxP removal in ES cells. We demonstrate that adenovirus-Cre infected ES cells can retain their germline competency. The procedures described here facilitate a rapid genetic manipulation of ES cells to obtain neo-free knockout animals, multiple gene targeting, homozygous mutant ES cells ideal for in vitro characterization, or Rag-deficient blastocyst complementation. PMID- 15282750 TI - Peptide and protein characterization by high-rate electron capture dissociation Fourier transform ion cyclotron resonance mass spectrometry. AB - The analytical utility of the electron capture dissociation (ECD) technique, developed by McLafferty and co-workers, has substantially improved peptide and protein characterization using Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS). The limitations of the first ECD implementations on commercial instruments were eliminated by the employment of low-energy electron injection systems based on indirectly heated dispenser cathodes. In particular, the ECD rate and reliability were greatly increased, enabling the combination of ECD/FTICR-MS with on-line liquid separation techniques. Further technique development allowed the combination of two rapid fragmentation techniques, high rate ECD and infrared multiphoton dissociation (IRMPD), in a single experimental configuration. Simultaneous and consecutive irradiations of trapped ions with electrons and photons extended the possibilities for ion activation/dissociation and led to improved peptide and protein characterization. The application of high rate ECD/FTICR-MS has demonstrated its power and unique capabilities in top-down sequencing of peptides and proteins, including characterization of post translational modifications, improved sequencing of peptides with multiple disulfide bridges and secondary fragmentation (w-ion formation). Analysis of peptide mixtures has been accomplished using high-rate ECD in bottom-up mass spectrometry based on mixture separation by liquid chromatography and capillary electrophoresis. This paper summarizes the current impact of high-rate ECD/FTICR MS for top-down and bottom-up mass spectrometry of peptides and proteins. PMID- 15282751 TI - Novel benzyl rearrangements in electrospray ionization multistage tandem mass spectra of benzyl 2',3'- didehydro-2',3'-dideoxythymidin-5'-yl H-phosphonate. AB - Several alkyl 2',3'-didehydro-2',3'-dideoxythymidin-5'-yl H-phosphonates were synthesized and analyzed by electrospray ionization multistage tandem mass spectrometry (ESI-MS(n)). Two kinds of novel benzyl rearrangement reactions were observed in ESI - MS(2) of [M + H](+), [M + Na](+) and [M + K](+) of benzyl 2',3' didehydro-2',3'-dideoxythymidin-5' yl H-phosphonate. Results from tandem mass spectrometry, high-resolution mass spectrometry and control experiments showed that the benzyl migration could undergo a four-membered cyclic rearrangement reaction, and benzyl was essential in the process. PMID- 15282752 TI - Study of peptide-sugar non-covalent complexes by infrared atmospheric pressure matrix-assisted laser desorption/ionization. AB - Infrared atmospheric pressure matrix-assisted laser desorption/ionization quadrupole ion trap mass spectrometry was applied to the study of siglec binding to oligosaccharide ligands. Peptides were designed to mimic the binding sites of three members of the siglec family: sialoadhesin, MAG and CD22. These peptides were tested for their ability to complex with their carbohydrate ligands 3' sialyllactose (3'SL) and 6'-sialyllactose (6'SL). All peptides demonstrated the ability to bind to the carbohydrates, with the peptide representing sialoadhesin maintaining its binding specificity for 3'SL in preference to 6'SL. This technique can be used to study other protein-sugar interactions and can be expanded to create high-throughput screening techniques. PMID- 15282753 TI - Borate-nucleotide complex formation depends on charge and phosphorylation state. AB - Flow injection analysis with electrospray ionization mass spectrometry was used to investigate borate-nucleotide complex formation. Solutions containing 100 microM nucleotide and 500 microM boric acid in water-acetonitrile-triethylamine (50:50:0.2, v/v/v; pH 10.3) showed that borate complexation with nicotinamide nucleotides was significantly influenced by the charge on the nicotinamide group and the number of phosphate groups on the adenine ribose. Borate binding decreased in the order of NAD(+), NADH, NADP(+) and NADPH. To investigate the relationship between complex formation and phosphorylation, association constants (K(A)) of borate-adenine (AMP, ADP, ATP), -guanine (GMP, GDP, GTP), -cytidine (CMP, CDP, CTP) and -uridine (UMP, UDP, UTP) complexes were compared. The results showed that the number of nucleotide phosphate groups was inversely proportional to the relative abundance of the borate complexes, with the K(A) of borate nucleotide complex decreasing in the order mono-, di- and tri-phosphates (AMP approximately GMP approximately CMP approximately UMP > ADP approximately GDP approximately CDP approximately UDP > GTP > ATP approximately CTP approximately UTP). At pH 7.4, using ammonium bicarbonate buffer, only borate-NAD(+) complex was observed. This indicates that the borate-NAD(+) complex may be the most physiologically relevant of those studied. PMID- 15282754 TI - Collision-induced dissociation tandem mass spectrometry of desferrioxamine siderophore complexes from electrospray ionization of UO2(2+), Fe3+ and Ca2+ solutions. AB - Desferrioxamine (DEF) is a trihydroxamate siderophore typical of those produced by bacteria and fungi for the purpose of scavenging Fe(3+) from environments where the element is in short supply. Since this class of molecules has excellent chelating properties, reaction with metal contaminants such as actinide species can also occur. The complexes that are formed can be mobile in the environment. Because the natural environment is extremely diverse, strategies are needed for the identification of metal complexes in aqueous matrices having a high degree of chemical heterogeneity, and electrospray ionization mass spectrometry (ESI-MS) has been highly effective for the characterization of siderophore-metal complexes. In this study, ESI-MS of solutions containing DEF and either UO(2)(2+), Fe(3+) or Ca(2+) resulted in generation of abundant singly charged ions corresponding to [UO(2)(DEF - H)](+), [Fe(DEF - 2H)](+) and [Ca(DEF - H)](+). In addition, less abundant doubly charged ions were produced. Mass spectrometry/mass spectrometry (MS/MS) studies of collision-induced dissociation (CID) reactions of protonated DEF and metal-DEF complexes were contrasted and rationalized in terms of ligand structure. In all cases, the most abundant fragmentation reactions involved cleavage of the hydroxamate moieties, consistent with the idea that they are most actively involved with metal complexation. Singly charged complexes tended to be dominated by cleavage of a single hydroxamate, while competitive fragmentation between two hydroxamate moieties increased when the doubly charged complexes were considered. Rupture of amide bonds was also observed, but these were in general less significant than the hydroxamate fragmentations. Several lower abundance fragmentations were unique to the metal examined: abundant loss of H(2)O occurred only for the singly charged UO(2)(2+) complex. Further, NH(3) was eliminated only from the singly charged Fe(3+) complex; this and fragmentation of C-C and C-N bonds derived from neither the hydroxamate nor the amide groups suggested that Fe(3+) insertion reactions were competing with ligand complexation. In no experiments were coordinating solvent molecules observed, attached either to the intact complexes or to the fragment ions, which indicated that both intact DEF and its fragments were occupying all of the coordination sites around the metal centers. This conclusion was based on previous experiments that showed that undercoordinated UO(2)(2+) and Fe(3+) readily added H(2)O and methanol in the ESI quadrupole ion trap mass spectrometer that was used in this study. PMID- 15282755 TI - Quantitative analysis of 5 alpha-reductase inhibitors in DU145 cells using matrix assisted laser desorption/ ionization time-of-flight mass spectrometry and high performance liquid chromatography/tandem mass spectrometry. AB - N-(Dicyclohexyl)acetylpiperidine-4-benzylidene-4-carboxylic acid (1) is an excellent in vitro inhibitor of 5 alpha-reductase (5 alpha R). Compound 1 showed, however, much lower inhibition activity of 5 alpha R in vivo than in vitro, which might be caused by poor membrane permeability. The methyl ester of 1 (1a) was therefore tested as a model prodrug to see if it has better permeability properties than the corresponding acid 1. It was also monitored that this methyl ester was cleaved into the active compound 1 within the DU145 cells. Quantitative matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) and high-performance liquid chromatography/tandem mass spectrometry (HPLC/MS/MS) methods were established with reliable linearity factors (0.996 for MALDI-TOFMS and 0.998 for HPLC/MS/MS) and reproducibility (relative standard deviation = 6.5% for MALDI-TOFMS and 2.8% for HPLC/MS/MS). The samples for MS analysis were effectively prepared from the cell homogenates using solid-phase extraction, with a high recovery of 90% on average. The intracellular amount of 1a (1.7 nmol) was much higher than that of 1 (0.032 nmol) in DU145 cells after 6 h of incubation. After incubation with the ester (1a), the cleaved acid (1) was detected within the cells. The concentration of acid 1 (0.045 nmol) in this experiment was higher than the acid content (0.032 nmol) after direct incubation with 1. Surprisingly, high amounts of the cleaved compound 1 were found outside the cells after 6 h of incubation with 1a. PMID- 15282756 TI - Identification of low molecular weight proteins isolated by 2-D liquid separations. AB - Proteins with molecular mass (M(r)) <20 kDa are often poorly separated in 2-D sodium dodecyl sulfate polyacrylamide gel electrophoresis. In addition, low-M(r) proteins may not be readily identified using peptide mass fingerprinting (PMF) owing to the small number of peptides generated in tryptic digestion. In this work, we used a 2-D liquid separation method based on chromatofocusing and non porous silica reversed-phase high-performance liquid chromatography to purify proteins for matrix-assisted laser desorption/ionization time-of-flight mass spectrometric (MALDI-TOFMS) analysis and protein identification. Several proteins were identified using the PMF method where the result was supported using an accurate M(r) value obtained from electrospray ionization TOFMS. However, many proteins were not identified owing to an insufficient number of peptides observed in the MALDI-TOF experiments. The small number of peptides detected in MALDI TOFMS can result from internal fragmentation, the few arginines in its sequence and incomplete tryptic digestion. MALDI-QTOFMS/MS can be used to identify many of these proteins. The accurate experimental M(r) and pI confirm identification and aid in identifying post-translational modifications such as truncations and acetylations. In some cases, high-quality MS/MS data obtained from the MALDI-QTOF spectrometer overcome preferential cleavages and result in protein identification. PMID- 15282758 TI - An atmospheric pressure chemical ionization study of the positive and negative ion chemistry of the hydrofluorocarbons 1,1-difluoroethane (HFC-152a) and 1,1,1,2 tetrafluoroethane (HFC-134a) and of perfluoro-n-hexane (FC-72) in air plasma at atmospheric pressure. AB - A report is given on the ionization/dissociation behavior of the title compounds within air plasmas produced by electrical corona discharges at atmospheric pressure: both positive and negative ions were investigated at different temperatures using atmospheric pressure chemical ionization mass spectrometry (APCI-MS). CHF(2)CH(3) (HFC-152a) undergoes efficient ionic oxidation to C(2)H(5)O(+), in which the oxygen comes from water present in the plasma. In contrast, CF(3)CH(2)F (HFC-134a) does not produce any characteristic positive ion under APCI conditions, its presence within the plasma being revealed only as a neutral ligand in ion-molecule complexes with ions of the background (H(3)O(+) and NO(+)). Analogously, the perfluorocarbon FC-72 (n-C(6)F(14)) does not produce significant positive ions at 30 degrees C: at high temperature, however, it undergoes dissociative ionization to form many product ions including C(3)F(6)(+), C(2)F(4)(+), C(n)F(2n+1)(+) and a few families of oxygen containing cations (C(n)F(2n+1)OH(2)(+), C(n)F(2n)OH(+), C(n)F(2n-1)O(+), C(n)F(2n 1)O(2)H(2)(+), C(n)F(2n-2)O(2)H(+)) which are suggested to derive from C(n)F(2n+1)(+) in a cascade of steps initiated by condensation with water followed by steps of HF elimination and H(2)O addition. Negative ions formed from the fluoroethanes CHF(2)CH(3) and CF(3)CH(2)F (M) include complexes with ions of the background, O(2)(-)(M), O(3)(-)(M) and some higher complexes involving also water, and complexes of the fluoride ion, F(-)(H(2)O), F(-)(M) and higher complexes with both M and H(2)O also together. The interesting product O(2)( )(HF) is also formed from 1,1-difluoroethane. In contrast to the HFCs, perfluoro n-hexane gives stable molecular anions, M(-), which at low source temperature or in humidified air are also detected as hydrates, M(-)(H(2)O). In addition, in humidified air F(-)(H(2)O)(n) complexes are also formed. The reactions leading to all major positive and negative product ions are discussed also with reference to available thermochemical data and relevant literature reports. The effects on both positive and negative APCI spectra due to ion activation via increasing V(cone) are also reported and discussed: several interesting endothermic processes are observed under these conditions. The results provide important information on the role of ionic reactions in non-thermal plasma processes. PMID- 15282760 TI - Matrix-dependent cationization in MALDI mass spectrometry. AB - The matrix dependence in cationization processes, the competition between cationization and protonation and the question of whether gas-phase cation transfer or attachment of free cations dominates in matrix-assisted laser desorption/ionization mass spectrometry were studied. Two different sample preparation methods were employed, the dried-droplet sample preparation and a mixture of solid matrix, analyte and salt. The latter ensures that the formation of cation adducts takes place in the gas phase. By monitoring the suppression of matrix signals for different matrices, it was found that matrices with high gas phase metal ion binding energies require high analyte concentrations for matrix suppression to occur. By comparing the mass spectra obtained using sinapinic acid or sinapinic methyl ester as a matrix, a correlation between cationization and deprotonation of matrix molecules was found. It is also demonstrated that attachment of free gas-phase cations, rather than cation transfer from the cationized matrix, is the predominant process in cationization. PMID- 15282761 TI - Fragmentation study of peptide acetals and aldehydes using in-source collision induced dissociation. AB - The fragmentation of peptide acetals and peptide diols, corresponding to the hydrated form of the peptide aldehyde, is dominated by the successive losses of two molecules of MeOH and water, respectively. Using model peptides, the fragmentation mechanism, with respect to the loss of methanol and water, was elucidated. The first loss was certainly charge-directed whereas the second probably occurred via the nucleophilic attack of the nitrogen of an amine on the C-terminal carbon leading to a cyclic ion. PMID- 15282763 TI - Current literature in mass spectrometry. PMID- 15282762 TI - Simple, sensitive and rapid liquid chromatographic/electrospray ionization tandem mass spectrometric method for the quantification of lacidipine in human plasma. AB - A simple, sensitive and rapid liquid chromatographic/electrospray ionization tandem mass spectrometric method was developed and validated for the quantification of lacidipine in human plasma using its structural analogue, amlodipine, as internal standard (IS). The method involves a simple single-step liquid-liquid extraction with tert-butyl methyl ether. The analyte was chromatographed on an Xterra MS C(18) reversed-phase chromatographic column by isocratic elution with 20 mM ammonium acetate buffer-acetonitrile (10:90, v/v; pH 6) and analyzed by mass spectrometry in the multiple reaction monitoring mode. The precursor to product ion transitions of m/z 456.4 --> 354.4 and m/z 409.3 --> 238.3 were used to measure the analyte and the I.S., respectively. The chromatographic run time was 1.5 min and the weighted (1/x(2)) calibration curves were linear over the range 0.1-25 ng ml(-1). Lacidipine was sensitive to temperature in addition to light. The method was validated in terms of accuracy, precision, absolute recovery, freeze-thaw stability, bench-top stability and re injection reproducibility. The limit of detection and lower limit of quantification in human plasma were 50 and 100 pg ml(-1), respectively. The within- and between-batch accuracy and precision were found to be well within acceptable limits (<15%). The analyte was stable after three freeze-thaw cycles (deviation <15%). The average absolute recoveries of lacidipine and amlodipine (IS) from spiked plasma samples were 51.1 +/- 1.3 and 50.3 +/- 4.9%, respectively. The assay method described here could be applied to study the pharmacokinetics of lacidipine. PMID- 15282764 TI - Quantification of mephenytoin and its metabolites 4'-hydroxymephenytoin and nirvanol in human urine using a simple sample processing method. AB - A reliable and easy to use liquid chromatography/tandem mass spectrometry (LC/MS/MS) method without the use of sample extraction was developed for the simultaneous quantification of urinary concentrations of mephenytoin, a standard phenotyping substrate for the cytochrome P450 enzyme CYP2C19, and its phase I metabolites 4'-hydroxymephenytoin and nirvanol. Fifty microL of urine were diluted with a buffered beta-glucuronidase solution and incubated at 37 degrees C for 6 h followed by addition of methanol, containing the internal standard 4' methoxymephenytoin. The chromatographic separation was achieved using a 100 x 3 mm, 5 micro Thermo Electron Aquasil C18 column with a gradient flow, increasing the organic fraction (acetonitrile/methanol 50:50) of the mobile phase from 10 to 90%. Quantification by triple-stage mass spectrometry (TSQ Quantum, Thermo Electron) was accomplished by negative electrospray ionization in the selected reaction monitoring mode. Linearity was observed for all substances in the concentration range 15-10 000 ng/mL. The lower limit of quantification (LLOQ) was 20 ng/mL for 4'-hydroxymephenytoin and 30 ng/mL for nirvanol and mephenytoin, respectively. Intra- and inter-day inaccuracy did not exceed 9.5% for all substances from LLOQ to 10 000 ng/mL. Intra- and inter-day precision were in the range of 0.8-10.5%. The method was validated according to international ICH and FDA guidelines and successfully applied for phenotyping of Caucasian male volunteers who received an oral dose of 50 mg mephenytoin. PMID- 15282765 TI - Quantitation of drug metabolites in the absence of pure metabolite standards by high-performance liquid chromatography coupled with a chemiluminescence nitrogen detector and mass spectrometer. AB - Quantitative information on drug metabolites with pharmacological or toxicological activities is of great interest during the drug discovery and development process. Because the analyte response with mass spectrometry can change significantly due to small variations in chemical structure, pure standards are required to construct standard curves for quantitation. However, for most programs at the discovery stage, pure metabolite standards are not available. In this work, an evaluation was conducted using a chemiluminescent nitrogen detector (CLND) as a calibrator to obtain the response factor ratio on a mass spectrometer generated from a metabolite and its parent compound in biological fluids. Using the response factor ratio obtained from the CLND, the metabolite could be quantified with the liquid chromatography/tandem mass spectrometry (LC/MS/MS) response obtained from the parent drug's standard curve. For this evaluation, oxazepam and temazepam were chosen as a 'drug/metabolite' pair. Temazepam was treated as the methylated metabolite of oxazepam. A spiked dog urine sample with a known concentration of oxazepam and unknown concentration of temazepam was injected onto the HPLC system and detected by both the CLND and MS/MS. Taking advantage of the equimolar response feature of the CLND, a response factor ratio between temazepam and oxazepam on the mass spectrometer was obtained by comparing the peak areas generated on the CLND and the mass spectrometer. From this ratio, temazepam was quantified using the oxazepam standard curve. The difference between the concentration of temazepam obtained from the reconstructed standard curve and the concentration obtained directly from a real temazepam standard curve was within 13% except the least concentrated standard (31%). This methodology has been successfully applied to measure quantities of the metabolite of a proprietary compound in a dog pharmacokinetic (PK) study. PMID- 15282766 TI - A computational and experimental study of cation affinity (Na+) of nucleobases and modified nucleobases by electrospray ionization ion trap mass spectrometry. AB - Gas-phase Na+ affinities of modified or unmodified nucleobases were determined theoretically at the density functional theory level, with the B3P86 functional and the 6-31 + G* basis set, and experimentally using electrospray ionization ion trap mass spectrometry (ESI-ITMS) and the kinetic method. For the calculations, the sodium cation affinities (SCA) were obtained from energies of the most stable complexes of the free nucleobases. Experimentally and theoretically relative scales of cation affinities were determined using eight modified and unmodified nucleobases and a very good agreement was obtained. PMID- 15282767 TI - Determination of abamectin in soil samples using high-performance liquid chromatography with tandem mass spectrometry. AB - Abamectin, which is comprised of a mixture of avermectins B1a and B1b, is a natural pesticide used as an anti-parasitic agent in livestock, ornamental, and agricultural crops, which can potentially be transported to aquatic systems. These compounds are highly toxic to both aquatic vertebrates and invertebrates at low concentrations in water. This investigation developed high-performance liquid chromatography/tandem mass spectrometry (HPLC/MS/MS) techniques to support automated extraction by an accelerated solvent extraction (ASE) system and chromatographic techniques to measure residues of avermectins in complex soil samples. HPLC along with atmospheric pressure chemical ionization (APCI) MS/MS was used for separation and determination of avermectin isomers in soil samples. Average method recovery for abamectin by UV was 91%, while detection by MS/MS resulted in a 68% recovery for abamectin. Individual method recoveries by MS/MS were 53.6% for avermectin B1a and 36.8% for avermectin B1b. The use of tandem technology eliminated matrix interferences and resulted in an approximately eight fold increase in sensitivity. PMID- 15282768 TI - The reproducible acquisition of comparative liquid chromatography/tandem mass spectrometry data from complex biological samples. AB - An in-depth study of the reproducibility of data acquired for comparative proteomics analysis using a prototype two-stage heated laminar flow chamber fitted to a commercial high-performance liquid chromatography/tandem mass spectrometry (HPLC/MS/MS) instrument was undertaken. The study is based on 24 replicate samples from four independent membrane preparations derived from two matched breast cancer cell lines. Variation and reproducibility in the data were evaluated at several levels highlighting the relative efficiency and variability of the acquisition routines used. Specifically, variation in the number and relative intensities of chromatographic peaks eluted from the LC column, precursor ion selection and sequence identification were evaluated. On average, approximately 6500 chromatographic peaks were generated for each acquisition with a corresponding coefficient of variance (CV) of less than 20%. Precursor ion selection and sequence identification averaged 1380 and 780 events per acquisition sample, respectively, with corresponding CVs of less than 10% for each. The reproducibility in the precursor ion selection was typically better than 60% between similar replicates. Using protein and peptide internal standards, it was found that the CV in retention time across the gradient between two acquisition pairs was typically less than 5%, whereas the average intensity ratio was 1.0 (expected) with a CV approaching 20%. An evaluation of the intensity ratios calculated from endogenous peptide sequences, identified across the acquisition set, indicated a CV of approximately 30%. Similarly, the CV associated with the top 1000 peptides indicated a mean and median of 28.4 and 26.95%. For a given acquisition pair it was also found that approximately 11% of the chromatographic peaks eluting from the column were linked to a sequence or identified. For these experiments, less than 10% of the peak pairs had absolute ratios greater than 2.0 and of those only approximately 10% had sequences linked to them. For each matched acquisition set on average 406 proteins were identified with a CV of less than 10%. Of the proteins that were identified approximately 30% had at least one predicted trans-membrane domain, indicating a four-fold increase over a crude homogenate sample with only minor enrichment. During these experiments it was found that the interface did not significantly alter the relative charge state distribution of ions, nor did it introduce significant interference from background ions. The interface was capable of 24-hour acquisition cycles. PMID- 15282769 TI - Determination of amphetamine in rat brain by in vivo microdialysis and ion pairing liquid chromatography with electrospray tandem mass spectrometry. AB - An ion-pairing liquid chromatography/electrospray tandem mass spectrometry (LC/ES MS/MS) method with in vivo microdialysis for the determination of amphetamine in rat brain has been developed. A microdialysis probe was surgically implanted into the striatum of the rat and artificial cerebrospinal fluid (aCSF) was used as the perfusion medium. Samples were collected and then analyzed off-line by LC/ES MS/MS. A reversed-phase C18 column was employed for LC separation and MS/MS was utilized for detection. Trifluoroacetic acid (TFA) was added to the mobile phase (acetonitrile/water) as an ion-pairing reagent. Detection was by ES-MS/MS directly, and no post-column addition of organic modifier was needed. Dual linear ranges were determined from 0.1-0.5 microg/mL and 0.005-0.1 microg/mL, respectively. The detection limit, based on a signal-to-noise ratio of 3, was 0.001 microg/mL (5 nM). Good precision and accuracy were obtained. The applicability of this newly developed method was demonstrated by continuous monitoring of amphetamine concentrations in rat brain. Amphetamine reached a maximum concentration of 0.086 +/- 0.017 microg/mL over 20-40 min after a single 3.0 mg/kg intraperitoneal administration. PMID- 15282770 TI - A simple, rapid and sensitive method for determination of aldehydes in human blood by gas chromatography/mass spectrometry and solid-phase microextraction with on-fiber derivatization. AB - Aldehydes are considered potential markers for enhanced oxidative stress and have been proposed as a diagnostic measure of cancer status. Do to their volatility and activity, it is very difficult to accurately measure aldehydes in human blood. In the present work, gas chromatography/mass spectrometry (GC/MS) and solid-phase microextraction (SPME) with on-fiber derivatization was developed for determination of aldehydes in human blood. O-(2,3,4,5,6 Pentafluorobenzyl)hydroxylamine hydrochloride (PFBHA) in aqueous solution was first adsorbed by a SPME fiber, and then the aldehydes in blood samples were headspace extracted by the SPME fiber and rapidly derivatized with PFBHA on the SPME fiber. Finally, the oximes formed were desorbed and detected by GC/MS in electron ionization (EI) mode. Validation of the present method was carried out, and the method was applied to quantitative analysis of the aldehydes in lung cancer blood. The results demonstrated that GC/MS and SPME with on-fiber derivatization is a simple, rapid, sensitive and solvent-free method for the determination of aldehydes in human blood. PMID- 15282771 TI - Phosphoric acid enhances the performance of Fe(III) affinity chromatography and matrix-assisted laser desorption/ionization tandem mass spectrometry for recovery, detection and sequencing of phosphopeptides. AB - An integrated analytical strategy for enrichment, detection and sequencing of phosphorylated peptides by matrix-assisted laser desorption/ionization (MALDI) tandem mass spectrometry (MS/MS) is reported. o-Phosphoric acid was found to enhance phosphopeptide ion signals in MALDI-MS when used as the acid dopant in 2,5-dihydroxybenzoic acid (2,5-DHB) matrix. The effect was largest for multiply phosphorylated peptides, which exhibited an up to ten-fold increase in ion intensity as compared with standard sample preparation methods. The enhanced phosphopeptide response was observed during MALDI-MS analysis of several peptide mixtures derived by proteolytic digestion of phosphoproteins. Furthermore, the mixture of 2,5-DHB and o-phosphoric acid was an excellent eluant for immobilized metal affinity chromatography (IMAC). Singly and multiply phosphorylated peptide species were efficiently recovered from Fe(III)-IMAC columns, reducing sample handling for phosphopeptide mapping by MALDI-MS and subsequent phosphopeptide sequencing by MALDI-MS/MS. The enhanced response of phosphopeptide ions in MALDI facilitates MS/MS of large (>3 kDa) multiply phosphorylated peptide species and reduces the amount of analyte needed for complete characterization of phosphoproteins. PMID- 15282772 TI - Negative electrospray ionization low-energy tandem mass spectrometry of hydroxylated fatty acids: a mechanistic study. AB - Recently, we reported that by converting olefinic fatty acids to their saturated vicinally 1,2-di-hydroxylated derivatives, abundant ions indicative for hydroxyl group locations are produced by negative electrospray ionization low-energy tandem mass spectrometry, allowing the assignment of the olefinic site in the native fatty acid. In this report the mechanisms whereby the characteristic ions are produced are investigated. The mono-hydroxylated fatty acid, 12 hydroxyoctadecanoic acid, served as a model for the more complex 12,13 dihydroxyoctadecanoic acid, and fragmentation mechanisms accounting for the most abundant product ions generated from their deprotonated molecules are proposed. In general, three different mechanisms are proposed to operate in the formation of the observed product ions: (i) step-wise charge-remote homolytic cleavages, (ii) step-wise charge-proximate homolytic cleavages, and (iii) concerted charge directed rearrangement reactions involving bond formation(s) and heterolytic cleavages. Support for the proposed mechanisms was achieved by investigating the deuterium- and oxygen-18-labeled isotopomers of both compounds. PMID- 15282773 TI - Glycopeptide analysis by matrix-assisted laser desorption/ionization tandem time of-flight mass spectrometry reveals novel features of horseradish peroxidase glycosylation. AB - We explored matrix-assisted laser desorption/ionization (MALDI) tandem time-of flight (TOF/TOF) mass spectrometry for the analysis of N-glycosylated peptides, using horseradish peroxidase (HRP) as a test case. Two different types of cleavage were observed in the TOF/TOF fragmentation spectra: Firstly, cleavages of peptide bonds yielded fragments with the attached N-glycans staying intact, thus revealing information on peptide sequence and glycan attachment site. Secondly, fragmentation of the glycan moiety was characterized by cleavage of glycosidic bonds as well as a (0,2)X-ring fragmentation of the innermost N acetylglucosamine of the chitobiose core. Loss of the complete N-glycan moiety occurred by cleavage of both the N-glycosidic bond and the side-chain amide group of the N-glycosylated asparagine, yielding a characteristic peak doublet with a mass difference of 17 Da, which revealed the individual masses of the N-glycan and the peptide moiety. Analysis of a HRP tryptic digest at the sub-picomole level allowed the characterization of various N-glycosylated peptides including those with internal disulfide linkages, a glycopeptide linked via a disulfide bond to another peptide, and a 5 kDa glycopeptide carrying two N-glycans. The potential of our approach was illustrated by the detection of the following novel features of HRP glycosylation: (i) The conjugation of a xylosylated trimannosyl N glycan without core-fucosylation to site Asn316, showing for the first time unambiguously the occupation of this site; and (ii) A disaccharide N acetylhexosamine1deoxyhexose1 attached to N-glycosylation sites Asn285 and Asn298, which might represent a Fuc(alpha1-3)GlcNAc- moiety arising from the processing of N-glycans by a horse-radish endoglycosidase during biosynthesis of HRP. PMID- 15282774 TI - High-pressure ion trap mass spectrometry. AB - The effects of buffer gas pressure on ion trap stability, mass resolution/calibration, and choice of mass scanning are described. Pressure effects were treated phenomenologically by adding a drag term to the ion equations of motion. The resulting collisional damping enlarges the mass dependent stability region but reduces the region in which mass-selective resonance ejection can be performed. The pressure effects can be reduced by increasing the frequency of the alternating quadrupole field. PMID- 15282775 TI - Simultaneous quantitation of dexmedetomidine and glucuronide metabolites (G-Dex-1 and G-Dex-2) in human plasma utilizing liquid chromatography with tandem mass spectrometric detection. AB - Dexmedetomidine (Dex) (Precedex) is a novel lipophilic imidazole derivative with a high affinity for alpha-2 adrenergic receptors, which exhibits sedative, analgesic-sparing, and sympatholytic properties. The pharmacological effects and therapeutic benefits of this drug have drawn continued interest from the medical community. Here we report a liquid chromatography/tandem mass spectrometry (LC/MS/MS) method to simultaneously measure the concentrations of dexmedetomidine and its glucoronide metabolites, G-Dex-1 and G-Dex-2, in human plasma samples. A solid-phase extraction method was developed to effectively extract Dex, G-Dex-1, and G-Dex-2 from plasma matrices. An isocratic chromatographic method was developed to achieve baseline separation of G-Dex-1 and G-Dex-2. The linear dynamic range evaluated was 19.08-1908.56 pg/mL for Dex, 65.17-6518.17 pg/mL for G-Dex-1, and 29.42-2943.28 pg/mL for G-Dex-2. The linear correlation coefficient (r) ranged from 0.9944-0.9979 for Dex, from 0.9966-0.9984 for G-Dex-1, and from 0.9939-0.9966 for G-Dex-2. The intra-assay coefficient of variation (CV) was between 2.5-12.5% for Dex, between 5.2-11.0% for G-Dex-1, and between 3.5-12.1% for G-Dex-2. The inter-assay precision of QC samples give % CV ranges from 6.5 9.3% for Dex, from 7.1-10.6% for G-Dex-1, and from 8.2-10.2% for G-Dex-2. The inter-assay accuracies ranged from 102.0-109.3% for Dex, from 95.4-105.6% for G Dex-1, and from 98.7-115.0% for G-Dex-2. PMID- 15282776 TI - Probing the molecular weight distributions of non-boiling petroleum fractions by Ag+ electrospray ionization mass spectrometry. AB - This work explores the possibility of Ag+ electrospray ionization mass spectrometry (ESI-MS) to determine the molecular weight distributions of non boiling petroleum fractions. Information about the molecular weight distributions is needed for fundamental studies on the nature of heavy crude oils and bitumens and for the development of novel recovery and processing methods. The method does not depend on thermal processes for the introduction of the fractions into the gas phase of the mass spectrometer, which is a considerable advantage over most other ionization methods. The Ag+ electrospray mass spectra of the fractions analyzed by using a toluene/methanol/cyclohexane (60:28:12%) solvent system display bimodal distributions in the ranges m/z approximately 300 to approximately 3000 and m/z 3000 to approximately 20,000. The abundances of the high molecular weight peak distributions can be reduced by in-source collisional activation experiments. Comparisons with the results obtained for model heteroatom-containing compounds (molecular weight < 600 Da) and high molecular weight polystyrene standards (up to one million Da) indicate that the majority of the structures in the saturate, naphthenoaromatic and polar aromatic fractions, and a significant portion of the asphaltenes, are small molecules. However, a considerable portion of the asphaltenes and some portion of the other fractions contain high molecular weight structures bound by covalent or strong non-covalent bonds. The results obtained by the Ag+ ESI method in this study for the saturate, aromatic, and polar fractions in a bitumen are in qualitative agreement with published molecular weight average results obtained for Cold Lake bitumen fractions analyzed by conventional gel permeation chromatography and field desorption mass spectrometry. Further work is needed to study the nature of the bonds and the interactions of the molecules in the asphaltene fractions by Ag+ ESI-MS. PMID- 15282777 TI - Selective digestion and novel cleanup techniques for detection of benzo[a]pyrene diol epoxide-DNA adducts by capillary electrophoresis/mass spectrometry. AB - Benzo[a]pyrene (BP) is a ubiquitous environmental polycyclic aromatic hydrocarbon (PAH) which, upon metabolic conversion to reactive benzo[a]pyrene-7,8-diol-9,10 epoxide (BPDE), has been found to attach covalently to DNA. Given the low level of DNA adducts typically present in vivo or in vitro, an essential first step prior to capillary electrophoresis/mass spectrometry (CE/MS) (or liquid chromatography/mass spectrometry (LC/MS)) analysis of the DNA digests is the removal of the bulk non-adducted nucleotides, enzymes or salts, and isolation of enriched adducts. This report focuses on the development of novel sample handling methods aimed at facilitating the analysis of BPDE-DNA adducts by CE/MS. This approach involves a simple variation on the digestion procedure, in combination with the use of metal affinity ZipTips for the more efficient cleanup of BPDE-DNA adducts formed in vitro for subsequent CE/MS analysis. The previously described digestion procedure, consisting of micrococcal nuclease, spleen phosphodiesterase and nuclease P1, allows for selective dephosphorylation of normal nucleotides, while leaving adducted nucleotides intact. Metal affinity ZipTips, typically used for selective extraction of phosphopeptides, were used here for extraction of adducted nucleotides. The utility of metal affinity SPE was tested on mixtures of dG and dGp, wherein nucleotide extracts contained no detectable nucleosides by CE/UV analysis. An in vitro BPDE-DNA incubation was then digested using the above procedure. Metal affinity solid-phase extraction (SPE) was subsequently used for the selective isolation of phosphorylated components, i.e., adducted nucleotides, from the mixture of enzymes and non-adducted nucleosides. SPE extracts were enriched in nucleotide adducts and analyzed using sample stacking and CE/MS. This method has several advantages over previously described cleanup procedures for dGp-BPDE adducts: fast, simple, uses commercially available materials, no need for excessive dilution (small scale), the suitability for use with automation, and possible applicability to other bulky hydrophobic adducts. PMID- 15282778 TI - Validation of a real-time monitoring method for aniline in freshwater by high performance liquid chromatography on porous graphitic carbon/electrospray ionization tandem mass spectrometry. AB - Aniline is an anthropogenic organic compound widely used in polymer, rubber, pharmaceutical and dye industries but also used in biodegradability assays of chemical compounds as a positive biodegradation standard. By the two approaches, the rapid determination of aniline is necessary because of the high toxicity of aniline on hemoglobin. A high-performance liquid chromatography/tandem mass spectrometry (HPLC/MS/MS) method for the determination of aniline in water is described here. This method, using benzylamine as internal standard, was validated. No time-consuming sample preparation was needed. A rapid separation (7 min between two chromatographic runs) of aniline and benzylamine was performed on a Hypercarb porous graphitic carbon column using a gradient of methanol and 100 mM formic acid. The obtained limits of detection and quantification were 10 and 1 ng/mL, respectively. The response for aniline was quadratic. We show that this problem could be circumvented by showing that the [calculated concentration = f(introduced concentration)] function was linear. The linearity range was 10-1000 ng/mL. An example of an application consisting of an aniline 42-day degradation kinetic in water was demonstrated. PMID- 15282779 TI - Electrospray ionization mass spectrometric characterization and quantitation of xanthine derivatives using isotopically labelled analogues: an application for equine doping control analysis. AB - Isotope-dilution mass spectrometry has been employed successfully in numerous fields of analytical chemistry enabling the establishment of fast and reliable procedures. In equine sports, xanthine derivatives such as caffeine and theobromine are prohibited, and doping control laboratories analyze horse urine specimens regarding these illicit performance-enhancing drugs. Theobromine has to exceed a threshold level of 2 microg/mL, hence a robust and reliable quantitation is required. Stably deuterated theobromine and caffeine were synthesized by the reaction of xanthine or theobromine with iodomethane-d3 in the presence of N methyl-N-trimethylsilyltrifluoroacetamide or potassium carbonate in acetonitrile, respectively. Both compounds were characterized by nuclear magnetic resonance spectroscopy and electrospray ionization tandem mass spectrometry, and a robust and fast assay for the qualitative and quantitative analysis of theobromine in equine urine samples was validated. Urine specimens were extracted by means of solid-phase extraction cartridges, and concentrated extracts were analyzed by liquid chromatography interfaced to a triple-quadrupole mass spectrometer. In addition, the dissociation behavior of deuterated analogues to caffeine and theobromine allowed proposals for fragmentation routes of xanthine derivatives after atmospheric pressure ionization and collisionally activated dissociation. PMID- 15282780 TI - Influence of flooding on delta15N, delta18O, 1delta15N and 2delta15N signatures of N2O released from estuarine soils--a laboratory experiment using tidal flooding chambers. AB - The influence of flooding on N2O fluxes, denitrification rates, dual isotope (delta18O and delta15N) and isotopomer (1delta15N and 2delta15N) ratios of emitted N2O from estuarine intertidal zones was examined in a laboratory study using tidal flooding incubation chambers. Five replicate soil cores were collected from two differently managed intertidal zones in the estuary of the River Torridge (North Devon, UK): (1) a natural salt marsh fringing the estuary, and 2 a managed retreat site, previous agricultural land to which flooding was restored in summer 2001. Gas samples from the incubated soil cores were collected from the tidal chamber headspaces over a range of flooding conditions, and analysed for the delta18O, delta15N, 1delta15N and 2delta15N values of the emitted N2O. Isotope signals did not differ between the two sites, and nitrate addition to the flooding water did not change the isotopic content of emitted N2O. Under non-flooded conditions, the isotopic composition of the emitted N2O displayed a moderate variability in delta18O and 2delta15N delta values that was expected for microbial activity associated with denitrification. However, under flooded conditions, half of the samples showed strong and simultaneous depletions in 1delta15N and delta18O values, but not in 2delta15N. Such an isotope signal has not been reported in the literature, and it could point towards an unidentified N2O production pathway. Its signature differed from denitrification, which was generally the N2O production pathway in the salt marsh and the managed retreat site. PMID- 15282781 TI - Mass spectrometric analysis of ceramide perturbations in brain and fibroblasts of mice and human patients with peroxisomal disorders. AB - In this study, the levels and composition of ceramides in brains of newborn mice lacking peroxisomes (Pex5-/-, Zellweger mice) were analyzed using normal-phase high-performance liquid chromatography/atmospheric pressure chemical ionization mass spectrometry (HPLC/APCI-MS). Total ceramide compositions were found to be comparable to that of control animals. However, a minor ceramide species, containing hexacosanoic/hexacosenoic acid as the amide fatty acid, was 9-fold increased. Also, in the sphingomyelin-derived ceramides this species was elevated. Subsequent analysis of extracts from fibroblasts of Pex5-/- mice and mice with a defective peroxisomal beta-oxidation (lacking D-specific multifunctional protein 2 (MFP2)), revealed, again, a similar rise in this particular ceramide. Further, this ceramide was elevated in human X-ALD fibroblasts as well. Whether C26:1/0-ceramide is linked to some of the pathology seen in Zellweger syndrome remains to be investigated. However, an increase in this sphingolipid can be considered as a diagnostic criterion for diseases caused by defects in peroxisome biogenesis or peroxisomal beta-oxidation. PMID- 15282782 TI - Determination of glycopeptide structures by multistage mass spectrometry with low energy collision-induced dissociation: comparison of electrospray ionization quadrupole ion trap and matrix-assisted laser desorption/ionization quadrupole ion trap reflectron time-of-flight approaches. AB - Multistage mass spectrometry, as implemented using low-energy collision-induced dissociation (CID) analysis in three-dimensional (3D) quadrupole ion traps (QITs), has become a powerful tool for the investigation of protein glycosylation. In addition to the well-known combination of QITs with electrospray ionization (ESI), also a matrix-assisted laser desorption/ionization -quadrupole ion trap--reflectron time-of-flight (MALDI-QIT-rTOF) mass spectrometer has recently become available. This study systematically investigates the differences between these types of instrument, as applied to characterization of glycopeptides from human antithrombin. The glycopeptides were obtained by tryptic digestion followed by lectin-affinity purification. Some significant differences between the ESI-QIT and MALDI-QIT-rTOF approaches appeared, most of them are causally related to the desorption/ionization process. The combination of a vacuum MALDI source with an ion-trap analyzer accentuates some characteristic differences between MALDI and ESI due the longer time frame needed for the trapping process. In contrast to ESI, MALDI generated ions that exhibited considerable metastable fragmentation during trapping. The long time span of the QIT process (ms range) compared with that for conventional rTOF experiments (micros range) significantly magnified the extent of this metastable fragmentation. With the investigated glycopeptides, a complete depletion of the terminal sialic acids of the glycopeptides as well as a variety of other fragment ions was already found in the MS1 spectra from the MALDI-QIT-rTOF instrument. The positive ion low-energy CID spectra (MS2) of the selected glycopeptides obtained using the two different QIT equipped instruments were found to be quite similar. In both approaches, fragmentation of the glycan and peptide structures occurred sequentially, allowing unambiguous sequence determination. In the case of ESI-QIT MS, fragmentation of the glycan structure occurred at the MS2 stage and fragmentation of the peptide structure was obtained only at the MS3 stage, which indicates the necessity of multistage CID experiments for complete structure elucidation. The MALDI-QIT-rTOF instrument yielded both kinds of fragments at the MS2 stage but without mutual interference. PMID- 15282783 TI - Identification of variant forms of the neuroendocrine peptide galanin. AB - Galanin is a neuroendocrine peptide widely distributed in the central and peripheral nervous systems and in endocrine tissues. Using radioimmunoassays, chromatographic separations, and tandem mass spectrometry, we have identified a series of five variant forms of galanin purified from porcine upper intestine. The modified variants include three beta-aspartyl-shifted forms, an oxidized form containing Trp monooxide, and an N-terminally truncated form. All were found to be C-terminally amidated. At least the beta-aspartyl-shifted forms could be of native occurrence. Mechanistic explanations of the mass spectrometric fragmentation patterns of the different forms are suggested. PMID- 15282784 TI - Correlations between air levels of hexahydrophthalic anhydride (HHPA) and HHPA adducted albumin tryptic peptides in nasal lavage fluid from experimentally exposed volunteers. AB - Organic acid anhydrides (OAAs) are low molecular weight, reactive compounds extensively used in industry. Exposure to these compounds may lead to allergic symptoms such as rhinitis and asthma. It is important to develop better and more informative methods for assessment of exposure to OAAs. The aim of this study was to develop a method for analysis of specific hexahydrophthalic anhydride (HHPA) adducted tryptic peptides of human serum albumin (HSA) in nasal lavage (NAL). Furthermore, these peptides were evaluated as biomarkers of exposure. The proteins in the NAL samples were reduced, alkylated and digested with trypsin and the obtained peptides were analyzed using liquid chromatography/tandem mass spectrometry. The total amount of hydrolyzable HHPA in an HHPA-HSA conjugate was used for calibration. A deuterium-labeled HHPA-HSA conjugate was used as internal standard. Five volunteers were exposed to 10, 40 and 80 microg/m3 of HHPA in an exposure chamber and NAL samples were collected before and after exposure. Acceptable precisions of the assay at 13-14% were found for three adducted peptides. The mean levels of these three peptides for the five subjects ranged between 5-22, 15-75 and 33-125 pmol/mL NAL for the exposures at 10, 40 and 80 microg/m3, respectively. High correlations between air levels and the measured peptides were found on an individual basis but there were large inter-individual differences ranging between 63 and 110% for the three peptides. The large differences remained after protein adjustments. It was possible to detect exposures below 10 microg/m3 with the method. Thus, these adducted peptides may be used as biomarkers of exposure, which may better estimate the risk than previous biomarkers developed for OAAs. PMID- 15282785 TI - Liquid chromatographic analysis of nucleosides and their mono-, di- and triphosphates using porous graphitic carbon stationary phase coupled with electrospray mass spectrometry. AB - Analysis of nucleosides and nucleotides is desirable in many biological studies, but the task is analytically challenging due to the high polarity of the analytes. In this study, resolution of mixtures containing nucleosides and their mono-, di- and triphosphates was achieved using a porous graphitic carbon (PGC) stationary phase, Hypercarb, under conditions suitable for liquid chromatography/mass spectrometry (LC/MS). Different organic mobile phases and modifiers were evaluated and the separation of 16 nucleosides and nucleotides was optimized using gradient elution with a water/acetonitrile mobile phase containing ammonium acetate and diethylamine as modifiers. The ammonium acetate concentration proved to be critical for retention and diethylamine was found to improve the peak shapes of di- and triphosphates for mass spectrometric detection. A variety of silica-based columns designed for polar compound separation were also tested using optimized LC conditions and compared with results obtained with the Hypercarb column. Only the Hypercarb column provided separations suitable for accurate quantitation of mixed nucleosides and their phosphates. PMID- 15282786 TI - Electron capture dissociation Fourier transform ion cyclotron resonance mass spectrometry in the electron energy range 0-50 eV. AB - Electron capture dissociation (ECD) of polypeptide cations was obtained with pencil and hollow electron beams for both sidekick and gas-assisted dynamic ion trapping (GADT) using Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS) with an electrostatic ion transfer line. Increasing the number of trapped ions by multiple ICR trap loads using GADT improved the ECD sensitivity in comparison with sidekick ion trapping and ECD efficiency in comparison with single ion trap load by GADT. Furthermore, enhanced sensitivity made it possible to observe ECD in a wide range of electron energies (0-50 eV). The degree, rate and fragmentation characteristics of ECD FTICR-MS were investigated as functions of electron energy, electron irradiation time, electron flux and ion trapping parameters for this broad energy range. The results obtained show that the rate of ECD is higher for more energetic (>1 eV) electrons. Long electron irradiation time with energetic electrons reduces average fragment ion mass and decreases efficiency of formation of c- and z-type ions. The obtained dependencies suggest that the average fragment ion mass and the ECD efficiency are functions of the total fluence of the electron beam (electron energy multiplied by irradiation time). The measured electron energy distributions in low-energy ECD and hot ECD regimes are about 1 eV at full width half maximum in employed experimental configurations. PMID- 15282787 TI - Flow-rate characterization of microfabricated polymer microspray emitters. AB - Microfabricated polymer microspray emitters are characterized in terms of applicable flow rates, temporal and spectral signal-to-noise ratios (SNRs), and solution composition. First, microspray emitters can be operated with 50% methanol/49% water/1% acetic acid from 250 nL/min up to 7 microL/min, with better SNRs above 1 microL/min. Interestingly, even at the lowest flow rates tested, they compare well with nanospray capillaries in terms of mass spectral performances. Secondly, they can be operated with acetonitrile from 10 up to 99% (v/v), with flow rates from 250 nL/min up to 4 microL/min. Even if the mass spectral performances (especially the spectral SNRs) vary with the acetonitrile content, this study validates such microfabricated microspray emitters as interfaces between liquid-phase separations and electrospray mass spectrometers. PMID- 15282788 TI - Trace level quantification of deuterated 17beta-estradiol and estrone in ovariectomized mouse plasma and brain using liquid chromatography/tandem mass spectrometry following dansylation reaction. AB - A sensitive liquid chromatography/tandem mass spectrometry (LC/MS/MS) method coupled with dansylation was developed for the simultaneous quantification of exogenously administered deuterated 17beta-estradiol-d4 (E2-d4) and its metabolite, estrone-d4 (E1-d4), in mouse plasma and brain homogenates. The dansylation reaction was simple, fast, and sensitive, and a lower limit of quantification of 50 pg/mL was achieved by using 50 microL of mouse plasma. Interference from endogenous 17beta-estradiol and estrone in plasma and brain samples was minimized by the use of deuterated-E2 as well by utilizing ovariectomized (OVX) mice. The recovery of dansylated derivative exceeded 83% and the reaction was completed within approximately 3 min. The intra- and inter-day assay precision were better than 12.9% and assay accuracy ranged between 92-104% for E1-d4 and E2-d4 in plasma, respectively. The absorption of E2-d4 at both 1 and 3 mg/kg P.O. was rapid, reaching peak plasma concentrations (Cmax) at 5 min post-dose that was the earliest time point obtained, and were 1.1 and 13.8 ng/mL, respectively; the Cmax values for the estrone metabolite, E1-d4, were 1.1 and 43.2 ng/mL, respectively. The area-under-the-plasma-time curve (AUC(0-2 h)) values were determined to be 0.65 and 2.90 ng. h/mL for E2-d4 and 0.77 and 6.74 ng. h/mL for E1-d4, respectively, at 1 and 3 mg/kg. The mean brain-to-plasma ratio for E1-d4 and E2-d4 after P.O. administration of E2-d4 to the OVX mice at 1 and 3 mg/kg indicated that both E1-d4 and E2-d4 were present in the brain as well as in the circulation. PMID- 15282789 TI - Determination of alkanolamines in cattails (Typha latifolia) utilizing electrospray ionization with selected reaction monitoring and ion-exchange chromatography. AB - Selected reaction monitoring (SRM) with electrospray ionization was used as a specific detection technique for the analysis of alkanolamines in plant tissue extracts. Ion-exchange chromatography was used as the method of separation. Quantification was based on monitoring the loss of either H2O or 2(H2O) from the protonated molecule [M+H]+. The method provided increased selectivity for all analytes and better detection limits for three of the six analytes investigated compared with an earlier method using selected ion monitoring with liquid chromatography. Instrumental detection limits ranged from 6-300 pg injected for monoethanolamine (MEA), monoisopropanolamine (MIPA), diethanolamine (DEA), methyldiethanolamine (MDEA), diisopropanolamine (DIPA), and triethanolamine (TEA). Method robustness and selectivity were demonstrated by the determination of DIPA and a known transformation product MIPA in over 35 plant extract samples derived from a laboratory study of plant uptake mechanisms. PMID- 15282790 TI - Formation of iminium ions by fragmentation of a2 ions. AB - Tandem mass spectrometric experiments have been carried out on the protonated amides H-Gly-Ala-NH2, H-Ala-Gly-NH2, H-Ala-Val-NH2, H-Val-Ala-pNA, H-Leu-Phe-NH2, H-Phe-Leu-NH2, H-Phe-Tyr-NH2 and H-Tyr-Phe-NH2 with particular emphasis on the fragmentation of the isomeric a2 ions derived therefrom. Primary fragmentation reactions of the protonated amides involve formation of the y1" and b2 ions with further fragmentation of the b2 ion to form the a2 ion which fragments to form iminium ions. Collision-induced dissociation studies of the mass-selected a2 ions were carried out. For the Gly-Ala, Ala-Gly and Val-Ala a2 ions, weak signals were observed corresponding to loss of CO from the a2 ion. With the exception of the Gly-Ala, Ala-Gly and Val-Ala a2 ions, both possible iminium ions (a1 and the internal iminium ion) are observed with the most abundant being that formed by proton attachment to the imine of higher proton affinity. The results provide strong support for the recently proposed (El Aribi et al. J. Am. Chem. Soc. 2003; 125: 9229) mechanism of fragmentation of a2 ions which involves elimination of CO from the a2 ion to form a proton-bound complex of two imines. Based on this mechanism ab initio calculations of the total energies of the a2 ions and the transition states for fragmentation have been carried out giving the energy barrier for fragmentation of each a2 ion. The experimental results are interpreted in terms of these energetics data, unimolecular rate constants calculated by using the RRKM theory, and the imine proton affinities. PMID- 15282792 TI - Synthesis and identification of two potential oxidation degradants of oxymetazoline. AB - Two potential oxidation degradants of oxymetazoline have been isolated by liquid chromatography and monitored by electrospray single quadrupole mass spectrometry. The structures of the products are shown to be oxymetazoline N-oxide and hydroxyamine by multiple-stage fragmentation ion trap mass spectrometry. The product ion spectra were installed in a library database and the library was used to examine an aged commercial product; one of the degradants was detected, but at a level of less than 0.1% of the parent. PMID- 15282793 TI - RScore: a peptide randomicity score for evaluating tandem mass spectra. AB - RScore, a new criterion of randomicity for evaluating tandem mass (MS/MS) spectra, is described. RScore is defined as the relative quality in cross correlation and matched intensity percentage of a potentially positive peptide to those of other possible candidates for the same spectrum. By utilizing RScore combined with less stringent SEQUEST score filters, the number of true positive peptides can be increased and the number of false positives in datasets from a known protein mixture can be reduced compared with current SEQUEST parameters used alone. This algorithm is simple and adds little overheads to SEQUEST computation. PMID- 15282794 TI - Electron ionization mass spectrometric study of antifungal 1,2,4-trithiols. PMID- 15282795 TI - Kel-F discs improve storage time of canopy air samples in 10-mL vials for CO2 delta13C analysis. PMID- 15282797 TI - A history of research on yeasts 7: enzymic adaptation and regulation. PMID- 15282798 TI - HXT5 expression is under control of STRE and HAP elements in the HXT5 promoter. AB - Hexose transporter (Hxt) proteins transport glucose across the plasma membrane in the yeast Saccharomyces cerevisiae. Recently, we have shown that expression of HXT5 is regulated by the growth rate of the cells. Because gene expression is regulated by binding of specific transcription factors to regulatory elements in the promoters of genes, the presence of putative regulatory elements in the promoter of HXT5 was determined by computer-assisted analysis. This revealed the presence of two putative stress-responsive elements (STREs), one putative post diauxic shift (PDS) element and two putative Hap2/3/4/5p (HAP) complex binding elements. The involvement of these elements was studied by using mutations in a HXT5 promoter-LacZ fusion construct. Growth during various conditions that result in low growth rates of yeast cells revealed that the STRE most proximal to the translation initiation site seemed to be involved in particular in regulation of HXT5 expression during growth at decreased growth rates. In addition, the HAP elements seemed to be required during growth on non-fermentable carbon sources. The PDS element and, to a lesser extent, the other STRE showed particular involvement in regulation of HXT5 expression during growth on ethanol. Finally, it was shown that the PKA pathway, which is known to be involved in expression of STRE-regulated genes, was also involved in regulation of HXT5 expression. A possible mechanism by which expression of HXT5 could be regulated by the transcriptional regulatory elements in the promoter is discussed. PMID- 15282799 TI - Actin distribution is disrupted upon expression of Yersinia YopO/YpkA in yeast. AB - Pathogenic Yersinia spp. use a Type III secretion system to inject effector proteins directly into host cells. The effector proteins interfere with normal cellular signalling and disrupt cytoskeletal structures that are required for phagocytosis of the invader. Here we show that Saccharomyces cerevisiae responds to one of these effector proteins, YopO/YpkA, essentially as has been observed in mammalian model systems. Expression of YopO in yeast results in inhibition of growth on solid medium. YopO expression confers a cytotoxic effect upon yeast with only 6% of cells remaining viable after 3 h of expression. Moreover, there is no obvious cell cycle arrest associated with cytotoxicity. YopO expression disrupts the normal distribution of actin in yeast. Actin is no longer observed exclusively at sites of cell growth. Concurrent with disruption of actin, we find YopO localizes to the cell periphery, where it may disrupt actin localization. Inactivation of the YopO kinase has no apparent effect upon the deleterious effects of YopO. Collectively, our results show that the yeast model system that has been constructed meets all requirements to serve as an excellent genetic model to exploit examination of YopO and its intracellular targets. PMID- 15282800 TI - Phenotypic characterization of glucose repression mutants of Saccharomyces cerevisiae using experiments with 13C-labelled glucose. AB - In the field of metabolic engineering and functional genomics, methods for analysis of metabolic fluxes in the cell are attractive as they give an overview of the phenotypic response of the cells at the level of the active metabolic network. This is unlike several other high-throughput experimental techniques, which do not provide information about the integrated response a specific genetic modification has on the cellular function. In this study we have performed phenotypic characterization of several mutants of the yeast Saccharomyces cerevisiae through the use of experiments with (13)C-labelled glucose. Through GC MS analysis of the (13)C incorporated into the amino acids of cellular proteins, it was possible to obtain quantitative information on the function of the central carbon metabolism in the different mutants. Traditionally, such labelling data have been used to quantify metabolic fluxes through the use of a suitable mathematical model, but here we show that the raw labelling data may also be used directly for phenotypic characterization of different mutant strains. Different glucose derepressed strains investigated employed are the disruption mutants reg1, hxk2, grr1, mig1 and mig1mig2 and the reference strain CEN.PK113-7D. Principal components analysis of the summed fractional labelling data show that deleting the genes HXK2 and GRR1 results in similar phenotype at the fluxome level, with a partial alleviation of glucose repression on the respiratory metabolism. Furthermore, deletion of the genes MIG1, MIG1/MIG2 and REG1 did not result in a significant change in the phenotype at the fluxome level. PMID- 15282801 TI - Efficient gene targeting in Kluyveromyces lactis. AB - Integration of a DNA fragment in a host genome requires the action of a double strand break (DSB) repair mechanism. Homologous recombination (HR) is initiated by binding of Rad52p to DNA ends and results in targeted integration. Binding of the Ku heterodimer (Ku70p/Ku80p) results in random integration via non-homologous end joining (NHEJ). In contrast to Saccharomyces cerevisiae, the budding yeast Kluyveromyces lactis shows variable, but in general low, gene targeting efficiency. To study and to improve gene targeting efficiency, K. lactis has been used as a model. The KlRAD51, KlRAD52 and KlKU80 genes have been isolated and deletion mutants for these genes have been constructed. Efficiency of gene targeting was determined at the KlADE2 locus using targeting constructs with different lengths of homologous flanking sequences. In wild-type K. lactis, the gene targeting efficiency ranged from 0% with 50 to 88% with 600 bp flanks. The Klku80 mutant, however, showed >97% gene targeting efficiency independently of the size of the homologous flanks. These results demonstrate that deletion of the NHEJ mechanism results in a higher gene targeting efficiency. Furthermore, increased gene targeting efficiency was achieved by the transformation of wild type K. lactis with the KlADE2 deletion construct in the presence of excess small DNA fragments. Using this method, PCR-generated deletion constructs containing only 50 bp of homologous flanking sequences resulted in efficient targeted gene replacement. PMID- 15282802 TI - Localization of proteins that are coordinately expressed with Cln2 during the cell cycle. AB - The localization of proteins can give important clues about their function and help sort data from large-scale proteomic screens. Forty-five proteins were tagged with the GFP variant YFP. These proteins were chosen because they are encoded by genes that display strong cell cycle-dependent expression that peaks in G(1). Most of these proteins localize to either the nucleus or to sites of cell growth. We are able to assign new cellular component GO terms to ASF2, TOS4, RTT109, YBR070C, YKR090W, YOL007C, YOL019W and YPR174C. We also have localization data for 21 other proteins. Noteworthy localizations were found for Rfa1p, a member of the DNA replication A complex, and Pri2p and Pol12p, subunits of the alpha-DNA polymerase : primase complex. In addition to its nuclear localization, Rfa1p assembled into cytoplasmic foci adjacent to the nucleus in cells during the G(1)-S phase transition of the cell cycle. Pri2 and Pol12 took on a beaded appearance at the G(1)-S transition and later in the cell cycle were enriched in the nuclear envelope. A new spindle pole body/nuclear envelope component encoded by YPR174 was identified. The cell cycle-dependent abundance of Tos4p mirrored Yox1p and these two proteins were the only proteins that were found exclusively at the G(1)-S phase of the cell cycle. A complete list of localizations, along with images, can be found at our website (http://www.yeastrc.org/cln2/). PMID- 15282803 TI - Sensitivity-encoded coronary MRA at 3T. AB - Long scan times are still a main limitation in free-breathing navigator-gated 3D coronary MR angiography (MRA). Unlike other MRI applications, high-resolution coronary MRA has not been amenable to acceleration by parallel imaging techniques due to signal-to-noise ratio (SNR) concerns. In the present work, mitigating SNR limitations by the transition to higher static magnetic field strength is proposed, thus enabling scan time reduction by the parallel sensitivity encoding (SENSE) technique. The study reports the implementation and evaluation of free breathing navigator-gated 3D coronary MRA with SENSE at 3T. Results from 11 healthy subjects indicate that the approach permits significant scan time reduction in MRA of the left and right coronary systems. Quantitative image analysis and visual grading suggest that two-fold scan acceleration can be accomplished at nearly preserved image quality. The additional experiments appear to demonstrate that parallel MRA equally permits enhancing volume coverage and spatial resolution while maintaining scan time. PMID- 15282804 TI - Multiple spin-echo spectroscopic imaging for rapid quantitative assessment of N acetylaspartate and lactate in acute stroke. AB - Monitoring the signal levels of lactate (Lac) and N-acetylaspartate (NAA) by chemical shift imaging can provide additional knowledge about tissue damage in acute stroke. Despite the need for this metabolic information, spectroscopic imaging (SI) has not been used routinely for acute stroke patients, mainly due to the long acquisition time required. The presented data demonstrate that the application of a fast multiple spin-echo (MSE) SI sequence can reduce the measurement time to 6 min (four spin echoes per echo train, 32 x 32 matrix). Quantification of Lac and NAA in terms of absolute concentrations (i.e., mmol/l) can be achieved by means of the phantom replacement approach, with correction terms for the longitudinal and transversal relaxation adapted to the multiple spin-echo sequence. In this pilot study of 10 stroke patients (symptom onset < 24 hr), metabolite concentrations obtained from MSE-SI add important information regarding tissue viability that is not provided by other sequences (e.g., diffusion-weighted imaging (DWI) and perfusion-weighted imaging (PWI)). Metabolic changes extended beyond the borders of the apparent diffusion coefficient (ADC) lesion in nine of the 10 patients, showing a rise in Lac concentrations up to 18 mmol/l, while NAA levels sometimes dropped below the detection level. Considerable differences among the patients in terms of the Lac concentrations and the size of the SI-ADC mismatch were observed. PMID- 15282805 TI - Effects of physiologic challenge on the ADC of intracellular water in the Xenopus oocyte. AB - The biophysical determinants of the intracellular water apparent diffusion coefficient (ADC) in mammalian tissues are poorly understood. Model systems that are more amenable to physical measurements may provide insights into the behavior of more complex systems. Toward that end, we used MRI to evaluate the effects of altered microtubule concentration, nuclear breakdown, and ATP depletion on intracellular water ADC in the Xenopus oocyte. Water ADC did not change in response to polymerization of microtubules with taxol or depolymerization with nocodazole. Water ADC did not change following the breakdown of the nucleus in healthy cells. Short-term depletion of ATP (approximately 20% of normal levels following 4 hr of exposure to sodium azide and 2-deoxy-D-glucose) was not associated with a change in intracellular ADC. Long-term depletion of ATP (approximately 20% of normal levels following 2 days of exposure to antimycin A) was associated with a significant decrease in intracellular water ADC. These findings suggest that intracellular water diffusion in oocytes is not dependent on the state of microtubule polymerization or short-term ATP depletion, although long-term ATP depletion is associated with changes that lead to a decrease in intracellular water ADC. PMID- 15282806 TI - Simultaneous measurement of arterial input function and tumor pharmacokinetics in mice by dynamic contrast enhanced imaging: effects of transcytolemmal water exchange. AB - A noninvasive technique for simultaneous measurement of the arterial input function (AIF) for gadodiamide (Omniscan) and its uptake in tumor was demonstrated in mice. Implantation of a tumor at a suitable location enabled its visualization in a cardiac short axis image. Sets of gated, low-resolution saturation recovery images were acquired from each of five tumor-bearing mice following intravenous administration of a bolus of contrast agent (CA). The AIF was extracted from the signal intensity changes in left ventricular blood using literature values of the CA relaxivity and a precontrast T1 map. The time dependent 1H2O relaxation rate constant (R1 = 1/T1) in the tumor was modeled using the BOLus Enhanced Relaxation Overview (BOLERO) method in two modes regarding the equilibrium transcytolemmal water exchange system: 1) constraining it exclusively to the fast exchange limit (FXL) (the conventional assumption), and 2) allowing its transient departure from FXL and access to the fast exchange regime (FXR), thus designated FXL/FXR. The FXL/FXR analysis yielded better fittings than the FXL-constrained analysis for data from the tumor rims, whereas the results based on the two modes were indistinguishable for data from the tumor cores. For the tumor rims, the values of Ktrans (the rate constant for CA transfer from the vasculature to the interstitium) and ve (volume fraction of the tissue extracellular and extravascular space) returned from FXL/FXR analysis are consistently greater than those from the FXL-constrained analysis by a factor of 1.5 or more corresponding to a CA dose of 0.05 mmole/kg. PMID- 15282807 TI - Proton MRI of lung parenchyma reflects allergen-induced airway remodeling and endotoxin-aroused hyporesponsiveness: a step toward ventilation studies in spontaneously breathing rats. AB - Proton signals from lung parenchyma were detected with the use of a gradient-echo sequence to noninvasively obtain information on pulmonary function in models of airway diseases in rats. Initial measurements carried out in artificially ventilated control rats revealed a highly significant negative correlation between the parenchymal signal and the partial pressure of oxygen (pO2) in the blood, for different amounts of oxygen administered. The magnitude of the signal intensity variations caused by changes in the oxygen concentration was larger than expected solely from the paramagnetic properties of molecular oxygen. Inhomogeneous line-broadening induced by lung inflation may explain the observed signal amplification. Experiments carried out in spontaneously breathing animals challenged with allergen or endotoxin revealed parenchymal signal changes that reflected the oxygenation status of the lungs and were consistent with airway remodeling or hyporesponsiveness. The results suggest that proton MRI of parenchymal tissue is a sensitive tool for probing the functional status of the lung in rat models of respiratory diseases. The method is complementary to the recently described noninvasive assessment by MRI of pulmonary inflammation in small rodents. Overall, these techniques provide invaluable information for profiling anti-inflammatory drugs in models of airway diseases. PMID- 15282808 TI - Assessment of tumor blood perfusion by high-resolution dynamic contrast-enhanced MRI: a preclinical study of human melanoma xenografts. AB - A noninvasive method to obtain high-resolution images of tumor blood perfusion is needed for individualized cancer treatments. In this study we investigated the potential usefulness of dynamic contrast-enhanced MRI (DCE-MRI), using human melanoma xenografts as models of human cancer. Gadopentetate dimeglumine (Gd DTPA) was used as the contrast agent, and DCE-MRI was performed at a voxel size of 0.5 x 0.2 x 2.0 mm3 with spoiled gradient-recalled sequences. We obtained images of E. F (where E is the extraction fraction, and F is perfusion) by subjecting DCE-MR images to Kety analysis. We obtained highly reproducible E. F images, which we verified by imaging heterogeneous tumors twice. We hypothesized that the extraction fraction of Gd-DTPA would be high and would not vary significantly in tumor tissue, implying that E. F should be a well-suited parameter for describing tumor blood perfusion. Observations consistent with this hypothesis were made by comparison of E. F-images with immunostained histological preparations from the imaged sections. The E. F images mirrored the histological appearance of the tumor tissue perfectly. Quantitative studies showed that E. F was highest in nonhypoxic tissue with high microvascular density, second highest in nonhypoxic tissue with low microvascular density, third highest in hypoxic tissue, and lowest in necrotic tissue. Moreover, the radial heterogeneity in E. F was almost identical to that in the blood supply, as assessed by the use of Na99mTcO4 as a perfusion tracer. Taken together, our observations show that high resolution images reflecting tumor blood perfusion can be obtained by DCE-MRI. PMID- 15282809 TI - Imaging oxygen consumption in forepaw somatosensory stimulation in rats under isoflurane anesthesia. AB - The cerebral metabolic rate of oxygen (CMRO2) was dynamically evaluated on a pixel-by-pixel basis in isoflurane-anesthetized and spontaneously breathing rats following graded electrical somatosensory forepaw stimulations (4, 6, and 8 mA). In contrast to alpha-chloralose, which is the most widely used anesthetic in forepaw-stimulation fMRI studies of rats under mechanical ventilation, isoflurane (1.1-1.2%) provided a stable anesthesia level over a prolonged period, without the need to adjust the ventilation volume/rate or sample blood gases. Combined cerebral blood flow signals (CBF) and blood oxygenation level-dependent (BOLD) fMRI signals were simultaneously measured with the use of a multislice continuous arterial spin labeling (CASL) technique (two-coil setup). CMRO2 was calculated using the biophysical BOLD model of Ogawa et al. (Proc Natl Acad Sci USA 1992;89:5951-5955). The stimulus-evoked BOLD percent changes at 4, 6, and 8 A were, respectively, 0.5% +/- 0.2%, 1.4% +/- 0.3%, and 2.0% +/- 0.3% (mean +/- SD, N = 6). The CBF percent changes were 23% +/- 6%, 58% +/- 9%, and 87% +/- 14%. The CMRO2 percent changes were 14% +/- 4%, 24% +/- 6%, and 43% +/- 11%. BOLD, CBF, and CMRO2 activations were localized to the forepaw somatosensory cortices without evidence of plateau for oxygen consumption, indicative of partial coupling of CBF and CMRO2. This study describes a useful forepaw-stimulation model for fMRI, and demonstrate that CMRO2 changes can be dynamically imaged on a pixel-by-pixel basis in a single setting with high spatiotemporal resolution. PMID- 15282810 TI - Quantification of vessel wall motion and cyclic strain using cine phase contrast MRI: in vivo validation in the porcine aorta. AB - Artery wall motion and strain play important roles in vascular remodeling and may be important in the pathogenesis of vascular disease. In vivo observations of circumferentially nonuniform wall motion in the human aorta suggest that nonuniform strain may contribute to the localization of vascular pathology. A velocity-based method to investigate circumferential strain variations was previously developed and validated in vitro; the current study was undertaken to determine whether accurate displacement and strain fields can be calculated from velocity data acquired in vivo. Wall velocities in the porcine thoracic aorta were quantified with PC-MRI and an implanted coil and were then time-integrated to compute wall displacement trajectories and cyclic strain. Displacement trajectories were consistent with observed aortic wall motion and with the displacements of markers in the aortic wall. The mean difference between velocity based and marker-based trajectory points was 0.1 mm, relative to an average pixel size of 0.4 mm. Propagation of error analyses based on the precision of the computed displacements were used to demonstrate that 10% strain results in a standard deviation of 3.6%. This study demonstrates that it is feasible to accurately quantify strain from low wall velocities in vivo and that the porcine thoracic aorta does not deform uniformly. PMID- 15282811 TI - Prebolus quantitative MR heart perfusion imaging. AB - The purpose of this study was to present the prebolus technique for quantitative multislice myocardial perfusion imaging. In quantitative MR perfusion studies the maximum contrast agent dose is limited by the requirement to determine the arterial input function (AIF). The prebolus technique consists of two consecutive contrast agent administrations. The AIF is determined from a first low-dose bolus, while a second, high-dose bolus allows the measurement of the myocardium with improved signal increase. The results of the prebolus technique using a multislice saturation recovery trueFISP sequence in healthy volunteers are presented. In comparison to a standard dose of 3 ml Gd-DTPA, perfusion values are maintained while the signal increase in the concentration time courses was considerably improved, accompanied by reduced standard deviations of the obtained perfusion values (0.72 +/- 0.13 ml/g/min for 1 ml/8 ml and 0.67 +/- 0.10 ml/g/min for 1 ml/12 ml Gd-DTPA, respectively). PMID- 15282812 TI - Factors affecting the accuracy of pressure measurements in vascular stenoses from phase-contrast MRI. AB - In this work the effects of noise, resolution, and velocity (flow) on the measurement of intravascular pressure from phase-contrast (PC) MRI are discussed. To elucidate these effects, we employed an axisymmetric geometry that enabled us to calculate pressures in <2 min on a Sun Ultra SPARC 10 workstation. To determine the effects of vascular stenoses, we fabricated several stenotic phantom geometries (with 50%, 75%, and 90% area stenoses), and performed both MRI and computational fluid dynamics (CFD) simulations for various flow rates for these phantom geometries. Noise with Gaussian statistics was added to the velocity field obtained from the CFD simulations. The pressure maps obtained directly from CFD simulations for our phantom geometries were compared with pressure maps derived by our algorithm when 1) the input was noise-corrupted velocity data from CFD, and 2) the input was PC-MRI data collected from the phantoms. The quantitative effects of noise, resolution, and flow rate on the accuracy of pressure measurements were determined. We found that for flow rates below the Reynolds number for turbulent flow, resolution is a more significant determinant of accuracy than SNR. Furthermore, if other parameters remain constant, increased flow rates may result in decreased accuracy. PMID- 15282813 TI - UNFOLD-SENSE: a parallel MRI method with self-calibration and artifact suppression. AB - This work aims at improving the performance of parallel imaging by using it with our "unaliasing by Fourier-encoding the overlaps in the temporal dimension" (UNFOLD) temporal strategy. A self-calibration method called "self, hybrid referencing with UNFOLD and GRAPPA" (SHRUG) is presented. SHRUG combines the UNFOLD-based sensitivity mapping strategy introduced in the TSENSE method by Kellman et al. (5), with the strategy introduced in the GRAPPA method by Griswold et al. (10). SHRUG merges the two approaches to alleviate their respective limitations, and provides fast self-calibration at any given acceleration factor. UNFOLD-SENSE further includes an UNFOLD artifact suppression scheme to significantly suppress artifacts and amplified noise produced by parallel imaging. This suppression scheme, which was published previously (4), is related to another method that was presented independently as part of TSENSE. While the two are equivalent at accelerations < or = 2.0, the present approach is shown here to be significantly superior at accelerations > 2.0, with up to double the artifact suppression at high accelerations. Furthermore, a slight modification of Cartesian SENSE is introduced, which allows departures from purely Cartesian sampling grids. This technique, termed variable-density SENSE (vdSENSE), allows the variable-density data required by SHRUG to be reconstructed with the simplicity and fast processing of Cartesian SENSE. UNFOLD-SENSE is given by the combination of SHRUG for sensitivity mapping, vdSENSE for reconstruction, and UNFOLD for artifact/amplified noise suppression. The method was implemented, with online reconstruction, on both an SSFP and a myocardium-perfusion sequence. The results from six patients scanned with UNFOLD-SENSE are presented. PMID- 15282814 TI - Non-Fourier-encoded parallel MRI using multiple receiver coils. AB - This paper describes a general theoretical framework that combines non-Fourier (NF) spatially-encoded MRI with multichannel acquisition parallel MRI. The two spatial-encoding mechanisms are physically and analytically separable, which allows NF encoding to be expressed as complementary to the inherent encoding imposed by RF receiver coil sensitivities. Consequently, the number of NF spatial encoding steps necessary to fully encode an FOV is reduced. Furthermore, by casting the FOV reduction of parallel imaging techniques as a dimensionality reduction of the k-space that is NF-encoded, one can obtain a speed-up of each digital NF spatial excitation in addition to accelerated imaging. Images acquired at speed-up factors of 2x to 8x with a four-element RF receiver coil array demonstrate the utility of this framework and the efficiency afforded by it. PMID- 15282815 TI - Fast three-dimensional k-space trajectory design using missile guidance ideas. AB - Three-dimensional (3D) k-space trajectories are needed to acquire volumetric images in MRI. While scan time is determined by the trajectory efficiency, image quality and distortions depend on the shape of the trajectories. There are several 3D trajectory strategies for sampling the k-space using rectilinear or curve schemes. Since there is no evidence about their optimality in terms of image quality and acquisition time, a new design method based on missile guidance ideas is explored. Since air-to-air missile guidance shares similar goals and constraints with the problem of k-space trajectory design, a control approach for missiles is used to design a 3D trajectory. The k-space is divided into small cubes, and each one is treated as a target to be sampled. The main goal is to cover the entire space as quickly and efficiently as possible, with good performance under different conditions. This novel design method is compared to other trajectories using simulated and real data. As an example, a trajectory that requires 0.11 times the number of shots needed by the cylindrical 3DFT acquisition was designed. This trajectory requires more shots (1.66 times) than the stack of spirals, but behaves better under nonideal conditions, such as off resonance and motion. PMID- 15282816 TI - Self-navigated motion correction using moments of spatial projections in radial MRI. AB - Interest in radial MRI (also known as projection reconstruction (PR) MRI) has increased recently for uses such as fast scanning and undersampled acquisitions. Additionally, PR acquisitions offer intrinsic advantages over standard two dimensional Fourier transform (2DFT) imaging with respect to motion of the imaged object. It is well known that aligning each spatial domain projection's center of mass (calculated using the 0th and 1st moments) to the center of the field of view (FOV) corrects shifts caused by in-plane translation. In this work, a previously unrealized ability to determine the in-plane rotational motion of an imaged object using the 2nd moments of the spatial domain projections in conjunction with a specific projection angle acquisition time order is reported. We performed the correction using only the PR data itself acquired with the newly proposed projection angle acquisition time order. With the proposed view angle acquisition order, the acquisition is "self-navigating" with respect to both in plane translation and rotation. We reconstructed the images using the aligned projections and detected acquisition angles to significantly reduce image artifacts due to such motion. The theory of the correction technique is described, and its effectiveness is demonstrated in phantom and in vivo experiments. PMID- 15282817 TI - Coherence-induced artifacts in large-flip-angle steady-state spin-echo imaging. AB - High-resolution imaging of trabecular bone aimed at analyzing the bone's microarchitecture is preferably performed with spin-echo-type pulse sequences. Unlike gradient echoes, spin-echoes are immune to artifactual broadening of trabeculae caused by local static field gradients near the bone-bone marrow interface and signal loss from chemical shift dephasing at k-space center. However, the previously practiced 3D fast large-angle spin-echo (FLASE) pulse sequence was found to be prone to a low-frequency modulation artifact in both the readout and slice direction. The artifact is caused by deviations in the effective flip angle of the nonselective 180 degrees pulse, which converts a fraction of the phase-encoded transverse magnetization to longitudinal magnetization. The latter recurs as transverse magnetization in the subsequent pulse sequence cycle forming a spurious stimulated echo. The objective of this work was to perform a k-space analysis of this steady-state artifact and propose two modifications of the original 3D FLASE that effectively remove it. The results of the simulations were in exact agreement with the experiments and the proposed remedy was found to eliminate the artifact. PMID- 15282818 TI - Imaging of myocardial infarction for diagnosis and intervention using real-time interactive MRI without ECG-gating or breath-holding. AB - Current methods for MRI of infarcted myocardium require ECG-gating and breath holding during contrast-enhanced segmented k-space inversion-recovery (IR) imaging. However, ECG-gating can be problematic in MRI, and breath-holding can be difficult for some patients. This work demonstrates that infarcted tissue can be visualized without ECG-gating or breath-holding with the use of intermittent inversion pulses during real-time (RT) interactive imaging with steady-state free precession (SSFP). The sequence generates a RT image stream containing a myocardium-nulled image every few frames, which allows nearly simultaneous observation of both infarcted regions and wall motion. First-pass perfusion and wall motion can be simultaneously observed with minor parameter modifications. This method may reduce diagnostic scan time, expand the target population, improve patient comfort, and facilitate targeted, interventional treatment of infarcted myocardium. Supplementary material for this article can be found on the MRM website at http://www.interscience.wiley.com/jpages/0740 3194/suppmat/index.html. PMID- 15282819 TI - Multiprocessor scheduling implementation of the simultaneous multiple volume (SMV) navigator method. AB - The simultaneous multiple volume (SMV) approach in navigator-gated MRI allows the use of the whole motion range or the entire scan time for the reconstruction of final images by simultaneously acquiring different image volumes at different motion states. The motion tolerance range for each volume is kept small, thus SMV substantially increases the scan efficiency of navigator methods while maintaining the effectiveness of motion suppression. This article reports a general implementation of the SMV approach using a multiprocessor scheduling algorithm. Each motion state is regarded as a processor and each volume is regarded as a job. An efficient scheduling that completes all jobs in minimal time is maintained even when the motion pattern changes. Initial experiments demonstrated that SMV significantly increased the scan efficiency of navigator gated MRI. PMID- 15282820 TI - Off-resonance effects in the transient response of SSFP sequences. AB - A previously developed eigenvector formalism is adapted to off-resonance in the transient response of quasiperiodic steady-state free precession (SSFP) sequences, including TrueFISP as a special case. The effective relaxation rates for essentially parallel and perpendicular deviations from the steady state are determined analytically in leading order perturbation theory. The latter are a known cause of oscillatory artifacts and therefore constitute the main target of a variety of preparation techniques. In addition, the former also play a dominating role in applications such as inversion recovery (IR) TrueFISP, which intentionally measure far away from the equilibrium. For both components, the approach toward equilibrium turns out to depend sensitively on field inhomogeneities, especially for smaller ratios of T2/T1. For the perpendicular deviations, the calculations show that--except very close to banding artifacts, where the steady-state signal is almost zero--field inhomogeneities additionally increase their effective relaxation rate almost as much as in the free induction decay (FID). The analytical results are tested against numerical simulation and MR measurements. PMID- 15282821 TI - Electrodynamics and ultimate SNR in parallel MR imaging. AB - The purpose of this article is to elucidate inherent limitations to the performance of parallel MRI. The study focuses on the ultimate signal-to-noise ratio (SNR), which refers to the maximum SNR permitted by the electrodynamics of the signal detection process. Using a spherical model object, it is shown that the behavior of the ultimate SNR imposes distinct limits on the acceleration rate in parallel imaging. For low and moderate acceleration, the ultimate SNR performance is nearly optimal, with geometry factors close to 1. However, for high reduction factors beyond a critical value, the ultimate performance deteriorates rapidly, corresponding to exponential growth of the geometry factor. The transition from optimal to deteriorating performance depends on the electrodynamic characteristics of the detected RF fields. In the near-field regime, i.e., for low B0 and small object size, the critical reduction factor is constant and approximately equal to four for 1D acceleration in the sphere. In the far-field wave regime the critical reduction factor is larger and increases both with B0 and object size. Therefore, it is concluded that parallel techniques hold particular promise for human MR imaging at very high field. PMID- 15282822 TI - Contribution of oxygenation to BOLD contrast in exercising muscle. AB - The potential physiological and therapeutic applications of functional MRI (fMRI) in skeletal muscle will depend on our ability to identify factors that may contribute to fluctuations in the BOLD signal. Until now, interpretations of signal changes in fMRI studies of muscle have mostly relied on the increase in muscle T2 associated with osmotically driven fluid shifts. However, recent studies have documented increases in BOLD signal intensity (SI) after single contractions, coinciding with increases in muscle hemoglobin saturation. In this study, the factors that contribute to variations in the intensity of the BOLD signal in exercising muscle are further addressed. For this purpose, BOLD imaging was performed during and after a moderate electrical stimulation was applied to the sciatic nerve in mice. In addition, oxygen pressure (pO2), blood flow, and skeletal muscle T2 (fast and slow components: T2 and T'2, respectively) were monitored. A comparison between mice lacking eNOS (eNOS-/- mice) and their wild type (WT) littermates was performed. In WT mice, the BOLD SI, as well as muscle oxygenation and T'2, were significantly increased for a prolonged time in response to this moderate exercise protocol. Blood flow immediately dropped after the electrical stimulation was stopped. In eNOS-/- mice, the high BOLD SI did not persist after the exercise protocol ended. This finding correlates well with the evolution of muscle oxygenation, which progressively decreases after stimulation in eNOS-/- mice. However, T'2 remained high for a prolonged time after stimulation. We therefore concluded that the maintenance of BOLD SI in moderately exercising skeletal muscle depends mainly on changes in pO2, rather than on blood flow or T2 effects. PMID- 15282823 TI - Metabolite and diffusion changes in the rat brain after Leksell Gamma Knife irradiation. AB - Our study describes the time course of necrotic damage to the rat brain resulting from Leksell Gamma Knife (LGK) irradiation at a dose that was previously considered to be subnecrotic. A lesion induced in the rat hippocampus by 35 Gy irradiation was monitored by MRI, MRS, and DW-MRI for 16 months. T2-weighted images revealed a large hyperintense area with an increased apparent diffusion coefficient of water (ADCw), which occurred 8 months after irradiation, accompanied by metabolic changes (increase of lactate (Lac) and choline (Cho), and decrease of creatine (Cr) and N-acetyl aspartate (NAA), as determined by MRS) that indicated an edema. In two animals, the hyperintensity persisted and a postnecrotic cavity connected to enlarged lateral ventricles developed. In the rest of the animals, the hyperintensity started to decrease 9 months post irradiation (PI), revealing hypointense areas with a decreased ADCw. Histology confirmed the MRI data, showing either scar formation or the development of a postnecrotic cavity. PMID- 15282824 TI - Histochemical detection of ultrasmall superparamagnetic iron oxide (USPIO) contrast medium uptake in experimental brain ischemia. AB - Recently, macrophage infiltration in different central nervous system (CNS) pathologies has been visualized with ultrasmall particles of iron oxide (USPIO) as a new cell-specific contrast medium for MRI. However, validation of these findings at the histological level has been hampered by the fact that the in situ detection of iron uptake by conventional Prussian blue staining is not sensitive enough to detect low amounts of iron in the brain. Here, an improved method for the histochemical detection of USPIO uptake in ischemic brain lesions is reported. The procedure relies on the sequential enhancement of Prussian blue staining by diaminobenzidine and silver/gold impregnation. After photothrombotic cortical brain infarction, this method allowed sensitive in situ detection of iron-laden macrophages which matched both macrophage immunostaining and USPIO induced signal alterations in high-resolution 7 T MRI. This staining method provides a basis for correlative histological assessment of USPIO-enhanced MRI in a broad spectrum of CNS pathologies. PMID- 15282825 TI - Long component time constant of 23Na T*2 relaxation in healthy human brain. AB - Signal intensity in 23Na images is altered in pathologic conditions such as ischemia and may provide information regarding tissue viability complementary to MR diffusion and perfusion imaging. However, the multicomponent transverse relaxation of 23Na (spin 3/2) complicates the determination of tissue sodium concentration from 23Na images with nonzero echo-time. The purpose of this study was to measure the long component time constant of tissue sodium T*2 relaxation in the healthy human brain at 4 T. Multiecho gradient-echo 23Na images (10 echo times ranging from 3.8-68.7 ms) were acquired in five healthy human volunteers. T*2 was quantified on a pixel-by-pixel basis using a nonnegative least squares fitting routine using 100 equally spaced bins between 0.5-99.5 ms and parametric maps were produced representing components between 0.5-3, 3.1-50, 50.1-98, and 98.1-99.5 ms. The long T*2 component of tissue sodium (average +/- standard deviation) varied between cortex (occipital = 22.0 +/- 2.4 ms), white matter (parietal = 18.2 +/- 1.9 ms), and subcortical gray matter (thalamus = 16.9 +/- 2.4 ms). These results demonstrate considerable regional variability and establish a foundation for future characterization of 23Na T*2 in conditions such as cerebral ischemia and cancer. PMID- 15282826 TI - fMRI of the lumbar spinal cord during a lower limb motor task. AB - This study applied spinal fMRI to the lumbar spinal cord during lower limb motor activity. During active ankle movement, activity was detected in the lumbar spinal cord motor areas and sensory areas bilaterally. During passive ankle movement, activity was detected in the motor and sensory areas in lower lumbar spinal cord segments and motor activity in higher lumbar spinal cord segments. Spinal fMRI detects patterns of activity consistent with known physiology and can be used to reliably assess activity in the lumbar spinal cord during lower limb motor stimulation. This study affirms spinal fMRI as an effective tool for assessing spinal cord function and increases its potential as a clinical tool. PMID- 15282827 TI - Breath-hold water and fat imaging using a dual-echo two-point Dixon technique with an efficient and robust phase-correction algorithm. AB - A two-point Dixon technique using a novel phase-correction algorithm and commercially available dual-echo fast gradient-echo pulse sequence is presented. The phase-correction algorithm determines the directional rather than phase distribution of signals due to field inhomogeneities. Specifically, a region growing scheme uses precalculated spatial gradients of the signal phase to guide the growth sequence, so there is no need to manually select the seeds or use an empirical angular threshold. Further, the determination of the signal direction of a given pixel is based on both the amplitude and phase of the surrounding pixels, the direction of which has already been determined. The advantages of this algorithm include its easy implementation, computational efficiency, and robustness in the presence of pixels with large phase uncertainty. The feasibility and usefulness of the technique are demonstrated in vivo with artifact-free water and fat images of an entire abdomen in a single breath-hold. PMID- 15282828 TI - Microcirculation and microvasculature in breast tumors: pharmacokinetic analysis of dynamic MR image series. AB - The purpose of this study was to quantify microcirculation and microvasculature in breast lesions by pharmacokinetic analysis of Gd-DTPA-enhanced MRI series. Strongly T1-weighted MR images were acquired in 18 patients with breast lesions using a saturation-recovery-TurboFLASH sequence. Concentration-time courses were determined for blood, pectoral muscle, and breast masses and subsequently analyzed by a two-compartment model to estimate plasma flow and the capillary transfer coefficient per unit of plasma volume (F/VP, KPS/VP) as well as fractional volumes of the plasma and interstitial space (fP, fI). Tissue parameters determined for pectoral muscle (fP = 0.04 +/- 0.01, fI = 0.09 +/- 0.01, F/VP = 2.4 +/- 1.3 min(-1), and KPS/VP = 1.2 +/- 0.5 min(-1)) and 10 histologically proven carcinomas (fP = 0.20 +/- 0.07, fI = 0.34 +/- 0.16, F/VP = 2.4 +/- 0.7 min(-1), and KPS/VP = 0.86 +/- 0.62 min(-1)) agreed reasonable well with literature data. Best separation between malignant and benign lesions was obtained by the ratio KPS/F (0.35 +/- 0.17 vs. 1.23 +/- 0.65). The functional imaging technique presented appears promising to quantitatively characterize tumor pathophysiology. Its impact on diagnosis and therapy management of breast tumors, however, has to be evaluated in larger patient studies. PMID- 15282829 TI - Echo-planar BOLD fMRI of mice on a narrow-bore 9.4 T magnet. AB - The feasibility of BOLD fMRI in association with electrical somatosensory stimulation on spontaneously breathing, isoflurane-anesthetized mice was investigated using spin-echo, echo-planar imaging (EPI) on a vertical narrow-bore 9.4 T magnet. Three experiments were performed to derive an optimal fMRI protocol. In Experiment 1 (n = 9), spin-echo BOLD responses to 10% CO2 challenge under graded isoflurane (0.25-1.25%) ranged from 10 +/- 2% to 3.5 +/- 0.9%; the optimal BOLD contrast-to-noise ratio peaked at 0.75% isoflurane. In Experiment 2 (n = 6), hindpaw somatosensory stimulations using 1-7 mA under 0.75% isoflurane revealed the optimal BOLD response was at 6 mA. In Experiment 3 (n = 5), BOLD responses to 4 and 6 mA stimulation under 0.75% and 1% isoflurane were evaluated in detail, confirming the optimal conditions in Experiment 2. These results demonstrated that BOLD fMRI using single-shot, spin-echo EPI in a mouse somatosensory stimulation model could be routinely performed on high-field, vertical, narrow-bore magnets. This protocol might prove useful for fMRI studies of transgenic mice. PMID- 15282830 TI - Rapid T2 estimation with phase-cycled variable nutation steady-state free precession. AB - Variable nutation SSFP (DESPOT2) permits rapid, high-resolution determination of the transverse (T2) relaxation constant. A limitation of DESPOT2, however, is the presence of T2 voids due to off-resonance banding artifacts associated with SSFP images. These artifacts typically occur in images acquired with long repetition times (TR) in the presence of B0 inhomogeneities, or near areas of magnetic susceptibility difference, such that the transverse magnetization experiences a net phase shift during the TR interval. This places constraints on the maximum spatial resolution that can be achieved without artifact. Here, a novel implementation of DESPOT2 is presented incorporating RF phase-cycling which acts to shift the spatial location of the bands, allowing reconstruction of a single, reduced artifact-image. The method is demonstrated in vivo with the acquisition of a 0.34 mm3 isotropic resolution T2 map of the brain with high precision and accuracy and significantly reduced artifact. PMID- 15282831 TI - Development of a compact MRI system for trabecular bone volume fraction measurements. AB - A compact MRI system for measuring trabecular bone volume fraction (TBVF) of the calcaneus was developed with the use of a 0.21 T permanent magnet and portable MRI console. The entire system weighed < 600 kg and was installed in a 2 m x 2 m space. Two cross-sectional spin-echo images of a heel acquired with external reference phantoms (total measurement time = 5 min) were used to quantify the TBVF of the calcaneus. The linearity and reproducibility of the measurements were evaluated by means of proton density-adjusted phantoms. Comparative measurements with quantitative ultrasound (QUS) in groups of healthy female volunteers showed a relatively high positive correlation (R(2) = 0.4539, 0.2693) between TBVF and the speed of sound (SOS). These results demonstrate the potential of this new system for measuring bone density. Magn Reson Med 52:440-444, 2004. PMID- 15282833 TI - Challenges in implementing evidence-based treatment practices for co-occurring disorders in the criminal justice system. AB - The presence of adults with mental health and substance abuse disorders within the criminal justice system has become increasingly evident over the past decade. Interventions and treatment services have been designed and research conducted in an effort to establish evidence-based practices that effectively address the complex needs of this population. However, adopting and implementing these evidence-based interventions and practices within the real-world setting of criminal justice environments is challenging. This article reviews the research literature related to evidence-based treatment practices for offenders with co occurring mental health and substance abuse disorders and explores the inherent challenges of fitting these interventions and services within criminal justice settings. PMID- 15282834 TI - Women with co-occurring substance use and mental disorders (COD) in the criminal justice system: a research review. AB - Associated with the dramatic increase in the numbers of women entering the criminal justice system is the recognition of the prominent role of co-occurring substance use and mental disorders (COD) in the lives of female offenders. This article reviews current research examining the prevalence and range of COD among female offenders, the variety of psychosocial problems faced by the female offender with COD, and the multiple treatment needs of women with COD who are under criminal justice supervision. Women with COD can enter the criminal justice system at several different points and, because both substance use and mental disorders carry significant risk of relapse, effective treatment approaches must address both disorders. The paper concludes with a discussion of several important treatment issues and provides suggestions regarding an agenda for future treatment and research. PMID- 15282835 TI - Antipsychotic medication adherence, cocaine use, and recidivism among a parolee sample. AB - This study examined the independent and interactive associations between cocaine use and antipsychotic medication adherence in predicting 12 month criminal recidivism among a sample of mentally ill parolees (N = 200). Consistent with prior research, cocaine use (based on hair assays) was associated with more than a threefold increase (relative to non-cocaine users) in the likelihood of a parolee being returned to custody during the follow-up period. Although medication adherence (based on urine specimens) was not independently associated with a significant reduction in recidivism risk, the interaction between cocaine use and medication adherence was significant, revealing a disproportionate impact of medication adherence specific to cocaine users. Prediction models of recidivism based on self-reported measures of medication adherence and cocaine use revealed only marginally significant trends for cocaine use, no effect for adherence, and no significant interaction between these two predictors. PMID- 15282836 TI - Modified TC for MICA offenders: crime outcomes. AB - The study randomly assigned male inmates with co-occurring serious mental illness and chemical abuse (MICA) disorders to either modified therapeutic community (MTC) or mental health (MH) treatment programs. On their release from prison, MICA inmates who completed the prison MTC program could enter the MTC aftercare program. The results, obtained from an intent-to-treat analysis of all study entries, showed that inmates randomized into the MTC group had significantly lower rates of reincarceration compared with those in the MH group. The results also show that differences between the MTC + aftercare and comparison group across a variety of crime outcomes (i.e. any criminal activity, and alcohol or drug related criminal activity) are consistent and significant, and persist after an examination of various threats to validity (e.g. initial motivation, duration of treatment, exposure to risk). This study provides some support for the effectiveness of the prison TC only condition. The findings are encouraging and consonant with other studies of integrated prison and aftercare TC programs for substance abusing non-MICA offenders, although qualified by the possibility that selection bias (i.e. differences in motivation on entry into aftercare) may be operating. Nevertheless, given the available evidence and the need for effective programming for MICA offenders, program and policy makers should strongly consider developing integrated prison and aftercare modified TC programs for MICA offenders. PMID- 15282837 TI - One year return to custody rates among co-disordered offenders. AB - The extent to which therapeutic community (TC) methods meet the treatment needs of offenders with substance abuse disorders and co-occurring psychiatric disorders in prison is largely unknown. Very little research has been conducted with this population. The purposes of this study were to generate profiles of co disordered drug offenders entering TC treatment in prison and to assess their post-release reincarceration rates, compared with drug offenders without psychiatric disorders. Extensive intake interview data for over 8,500 men and women who received treatment in one of 16 prison-based TCs in California were analyzed to produce profiles of co-disordered participants. Intake data come from a 5 year process and outcome evaluation of the California Department of Corrections' (CDC's) treatment initiative. Post-release reincarceration rates come from the CDC's Offender Based Information System. Compared with non psychiatric disordered drug offenders, co-disordered offenders had substantially more severe substance abuse and criminal histories, in addition to their psychiatric impairment, at treatment entry. Logistic regression results indicated that, compared with drug offenders without psychiatric illness, co-disordered offenders were significantly more likely to be reincarcerated during the first year of their parole. These results suggest that prison treatment programs may need to use more comprehensive diagnostic assessments at intake to assess the diverse mental health needs of drug offenders with co-occurring psychiatric disorders and to develop treatment approaches suitable for this population. PMID- 15282838 TI - Effects of diversion on adults with co-occurring mental illness and substance use: outcomes from a national multi-site study. AB - This quasi-experimental non-equivalent comparison group study examines outcomes for participants in eight programs conducting criminal justice diversion for people with co-occurring serious mental illness and substance use disorders compared with jail detainees eligible for diversion, but who were processed through standard criminal justice methods without diversion. Nearly 2000 participants were interviewed at baseline, and 1500 at 3 month and 1300 at 12 month follow-up to baseline. In these interviews, outcome measures of re-arrest, mental health functioning, substance abuse, quality of life, and service utilization were obtained. Those diverted were more likely to have received mental health counseling, mental health medication, and mental health hospitalization than those not enrolled in a diversion program, but were equally likely to have received substance abuse counseling. Overall, the differences in proportions receiving services between the two groups were small, even when these differences were statistically significant. The effect associated with diversion differed somewhat across the individual sites. However, overall cross-site pooled analyses revealed no outcome differences between groups on measures of mental health symptoms, substance use, criminal justice recidivism, or quality of life. Although the immediate benefit of diversion as an access mechanism to community treatment is indicated in pooled cross-site results, such access was driven by more coercive (pre-booking and court) models and results suggest that effecting substantially greater access to services or services use did not occur. The findings also suggest that mental health, substance abuse, and criminal justice outcomes remain dependent on the treatment intervention received, perhaps moderated by type of diversion intervention, rather than on a generic and initial diversion event. PMID- 15282839 TI - Dual diagnosis in an Australian forensic psychiatric hospital: prevalence and implications for services. AB - Despite a growing awareness of the unique needs of psychiatric patients with co occurring substance abuse and dependence disorders (i.e. dual diagnosis), there is a dearth of research investigating the prevalence of dual diagnosis in forensic psychiatric populations. Similarly, little work has been done to determine the implications of dual diagnosis for forensic psychiatric patients. Patients at the Thomas Embling Hospital in Victoria, Australia, were assessed to determine the prevalence of substance abuse disorders and mental illnesses within this population. Results reveal that the majority of patients (approximately 74%) have a lifetime substance abuse or dependence disorder. Information was collected concerning patients' criminal histories and the Level of Service Inventory, Revised, was completed for each patient who participated. Results suggest that patients with both major mental illnesses and substance abuse disorders have more extensive criminal histories and demonstrate a higher level of risks and needs when compared with patients with major mental illness alone. The implications for the development and delivery of effective forensic mental health services that address both co-occurring disorders are also discussed. PMID- 15282840 TI - Correctional treatment for co-occurring disorders: results of a national survey. AB - The significant expansion of correctional populations in the last decade, coupled with the gradual erosion of community treatment infrastructure and health insurance, have brought greater attention to the needs of offenders who have co occurring mental health and substance use disorders. Individuals with co occurring disorders frequently cycle through acute care facilities in the community and increasingly are placed in jails or prisons. Approximately 16% of inmates in correctional facilities have major mental health disorders (Bureau of Justice Statistics, 1999), and a large majority of these inmates have co occurring substance use disorders. Few studies have examined the effectiveness of correctional treatment for co-occurring disorders, and there is little information available regarding clinical and programmatic approaches used with this population. The current study provides findings from a comprehensive national survey of co-occurring disorder treatment programs in correctional settings. A total of 20 co-occurring disorder treatment programs from 13 state correctional systems were identified and surveyed. Many of the programs featured modified therapeutic communities, but there was significant diversity in the duration of treatment and type of services provided. Several unique structural and clinical modifications to treatment have been developed in these settings. Implementation of co-occurring disorder treatment programs has led to enhanced collaboration with prison health services and community supervision and treatment agencies, and greater use of interdisciplinary staff to provide outreach and case management services. Research is now being conducted to examine outcomes in several of these correctional treatment programs. PMID- 15282842 TI - Transition planning and recidivism among mentally ill juvenile offenders. AB - OBJECTIVES: To determine the extent to which transition planning and community service would predict lower levels of recidivism among mentally ill juvenile offenders. DESIGN: Review of legal, medical and social service records including examination of 3-month period following community release. PARTICIPANTS: Juvenile offenders incarcerated for 6 months or more (N = 44) transitioning to community. MEASURES: Child and Adolescent Functional Assessment Scale (CAFAS), sum of different documented pre- and post-release discharge planning contacts, documented community service contacts. OUTCOMES: Re-offense during the first year post-release. RESULTS: Documented community service contacts in the 3 months following discharge were rare for mental health (20.5%), substance abuse (38.6%), financial assistance (29.5%), and school placement (34.1%). The average number of different pre-release and post-release discharge planning contacts was also low, M(SD)=1.86(1.68) and M(SD)= 2.34(1.71) respectively. Post-release discharge planning and the receipt of financial assistance and mental health services were all associated with lower levels of reoffending. CONCLUSIONS: Community transition planning, including the coordination and provision of community services, is an essential component of community reintegration for juvenile offenders and is associated with lower rates of recidivism during the first year post-discharge. PMID- 15282841 TI - Treatment retention of dually diagnosed offenders in an institutional therapeutic community. AB - This study investigated treatment retention in an institutional therapeutic community (ITC) for dually diagnosed male inmates. Twenty-five percent of admissions successfully completed the residential substance abuse treatment program. Graduates participated for an average of 9 months, while terminations participated for nearly 4 months. One-third of the terminations participated for at least 3 months and 15% participated for 6 months or more. A striking result was the relationship between retention and risk ratings and psychopathy scores. Both measures assess behavior and interaction styles over a range of situations. The presence of a single type of negative behavior (i.e. the nature of the committing offense) did not predict completion, but a rating based on multiple samples of behavior was a significant predictor of retention. PMID- 15282843 TI - IAVI begins trials of AIDS vaccine. PMID- 15282844 TI - AlphaVax receives NIAID contract to support development of HIV vaccine. PMID- 15282845 TI - Audience attitude toward the free lunch. PMID- 15282846 TI - Polymyositis: an overdiagnosed entity. PMID- 15282847 TI - Polymyositis: an overdiagnosed entity. PMID- 15282848 TI - [Drug-induced headache and pain reduction medication with amitriptyline in an 11 year old pupil]. AB - We report on an 11-year-old pupil who present-ed to a neurological practice in the company of his parents because of migraine and strong headaches. Precise anamnesis of drugs yielded the diagnosis of a drug-induced headache with underlying migraine disease. The boy was afflict-ed with a hereditary taint, as both parents also suffered from migraine. The mother was also continuously in treatment because of a chronic pain disorder. Therapy with up to 50 mg per day of amitriptyline enabled the boy to stop consuming analgesics without suffering from head-aches. Moreover, under 47.5 mg per day of metoprolol, the migraine symptoms did not re-occur. PMID- 15282849 TI - Contamination. Company to issue global dialysis alert. PMID- 15282850 TI - Global AIDS experts call for shift in HIV prevention. PMID- 15282852 TI - Group urges G-8 to honor vows. PMID- 15282851 TI - World leaders plan to speed up development of HIV vaccine. PMID- 15282853 TI - Exposure. Class action based on "fear factor" dismissed on appeal. PMID- 15282854 TI - Prison settles suit as judge decries care of AIDS inmates. PMID- 15282855 TI - Recommended steps in HIV prevention plan developed by global leaders. PMID- 15282856 TI - Politics. VIP group assails Bush's AIDS, other policies. PMID- 15282857 TI - Protest. Activists: Reagan ceremonies recolor history. PMID- 15282859 TI - HIV testing order overturned in carjacking case. PMID- 15282858 TI - Court-ordered testing upheld for child sex offender. PMID- 15282860 TI - Prisons. HIV-positive inmates in supermax have due process rights. PMID- 15282861 TI - Global prisons. Report: China approves special jails for AIDS victims. PMID- 15282862 TI - Schools. Districts, teachers cannot challenge unfunded HIV mandate. PMID- 15282863 TI - Order for HIV testing to hinge on fluid transmission. PMID- 15282864 TI - Advertisements. FDA orders Abbott to change misleading ads for Norvir. PMID- 15282865 TI - Simplifying antiretroviral therapy. AB - The substantial benefits conferred by HAART require strict patient adherence. Many of the initial HAART regimens consisted of a number of large pills that needed to be taken several times daily, sometimes with meal restrictions. The development of once-daily antiretroviral agents has eased some of the burden associated with the intense, difficult schedules of early HAART regimens.. The majority of regimens currently used in treatment-naive HIV-positive patients contain a mixture of agents that are taken on a once- and twice-daily basis. Although this is an improvement over past regimens, the asynchronous administration of pills throughout the day still presents a scheduling challenge for most patients. The newest advance in simplifying antiretroviral therapy is the use of regimens in which all pills are taken at the same time once a day. Choosing drugs for a fully once-daily regimen requires awareness of a number of factors, including pharmacokinetics, potency, durability of response, resistance and safety. At this time, there are a limited number of combinations that can be used as a fully once-daily regimen and few clinical trials evaluating such combinations. Results from initial clinical trials using simplified, once-daily regimens in treatment-naive patients have been promising. Additional studies should add to this experience and provide guidance on the role and timing of such regimens in the management of patients with HIV disease. PMID- 15282866 TI - Plasma cell disorders in HIV-infected patients: from benign gammopathy to multiple myeloma. AB - Plasma cell disorders are not uncommonly reported in young patients with HIV infection. These disorders range from benign polyclonal hypergammaglobulinemia to indeterminate monoclonal gammopathy of unknown significance (MGUS) to malignant dyscrasias, including multiple myeloma and plasma cell leukemia. Hypergammaglobulinemia and oligoclonal banding had been the most frequently reported disorders in the pre-HAART era. In HIV-infected persons, the incidence of MGUS is reported to be around 2.5%, with an approximate 4.5-fold increased risk of multiple myeloma. Many of these HIV-infected patients had been treated with alkylator-based regimens, and these reports predate the current widespread use of thalidomide-dexamethasone combination treatment in multiple myeloma. Although the optimal therapy for an HIV-infected person might with plasma cell dyscrasia is yet to be defined, in the current era of HAART the otherwise healthy HIV-infected patient might be tested like an HIV-negative person. Consequently, treatment with immunomodulatory agents (eg, thalidomide) and proteasome inhibitors (eg, bortezomib) may also be worth considering. High-dose chemotherapy with an autologous peripheral blood stem cell transplant is increasingly being considered as consolidation therapy in the younger non-HIV-infected myeloma patient. In the next few years, it is anticipated that these approaches will be applied more frequently to HIV-infected persons with myeloma. PMID- 15282867 TI - Simplifying suppression: switching therapies to improve HIV disease management. PMID- 15282868 TI - Tri-infection: successful treatment of a patient infected with HIV and hepatitis B and C viruses--a case report. AB - There is a growing abundance of literature on persons with HIV/AIDS who are coinfected with either hepatitis B or C virus. However, there is little published information on HIV-infected persons coinfected with both hepatitis B and C virus. This Case Report describes a patient with HIV infection and hepatitis B and C with biopsy-proven cirrhosis. The patient was successfully treated, achieving a sustained virologic response with respect to hepatitis C and seroconversion with respect to hepatitis B. PMID- 15282869 TI - Editorial comment: triple infection with HIV and hepatitis B and C viruses- lesson in combination therapy. PMID- 15282871 TI - [Association of HIV and parotid cysts]. AB - Parotid cystics are included in salivary glands affection, found with persons infected by the human immunodeficiency virus. This study reports 28 cases registered in the oto-rhino-laryngology service of Brazzaville university hospital. The interested age is between 32 and 57 years old. The HIV profile reveals a prevailing of HIV1 (20 cases). The dia-gnosis is clinical, echographic and by the exploratory puncture. The therapeuratic approach (iterative puncture and compressive dressing) adopted to these brittle patients and with limited resource, permitted to get satisfactory results. PMID- 15282872 TI - [Comparative metric study of the edentulous dental arch in black Africans and white Europeans]. AB - The current 89 samples comparative study shows that the mandible arch is different statistically in Blacks and Whites populations. African Blacks mandible dimensions are superior considering both inter retromolar tubercles measurements and the arch length. Therefore, the impression trays manufacturing should be reconsidered for Blacks prosthetic rehabilitation. PMID- 15282873 TI - Commentary: Willingness and competence of depressed and schizophrenic inpatients to consent to research. PMID- 15282874 TI - [Clinical-pharmacological aspects to accelerate the development process from the preclinical to the clinical phase/2nd communication: promising strategies]. AB - To improve the transition from research to development a critical evaluation of the individual project by research and disease area teams is required to include input from pharmacology, toxicology, pharmacokinetics, galenics, clinical pharmacology, clinical as well as regulatory experts and marketing. Decisions on the individual development strategy should be made prior to the start of development and all projects should be reviewed at predefined stages throughout the product development life cycle. This ensures consistency of decision-making not only during the development of individual products but throughout the entire development pipeline. Studies in the exploratory stage of drug development should be designed for decision making in contrast to later clinical trials in the confirmatory stage that require power for proof-of-safety and proof-of-efficacy. The more thorough and profound studies have been carried out during this exploratory stage of drug development, the earlier a decision can be made on the continuation or discontinuation of further development, thus saving development time and money and assessing and considerably reducing the risk for the patients and increasing the success rate of the project in the later confirmatory effectiveness trial with an adequate number of subjects receiving the new therapy under typical conditions of use. Strategies which may be helpful to improve the quality of decisions in drug discovery and drug development are: discovery experiments should be done to critically evaluate the compound, the "killer" experiments should be done as early as possible, continuous effort on preclinical disease models is necessary to improve predictability of efficacy in patients ("humanized" research): genomic technology should be used to identify novel, disease-related targets and to characterise preclinical test systems, improvement of knowledge and experience concerning the relevance of new technologies for the clinical picture, genotyping of clinical trial patients to select patient groups which are likely to respond to treatment (pharmacogenomics), modelling and simulation of preclinical and clinical trials, integration of pharmacokinetic and pharmacodynamic principles into drug development, assessment of the interaction potential (CYP-450, trasporter proteins and others), increasing use of biomarker/surrogate marker for rapid clinical feedback, involvement of the target population as soon as possible, applying statistical data analysis techniques for proving effectiveness, co-operation with high quality centers. To reach this goal clinical pharmacology must be fully integrated in the whole process from the candidate selection to its positioning within the market. PMID- 15282875 TI - Proceedings of the International Workshop on Outcomes in Osteoarthritis. Indianapolis, Indian, USA, December 13-14, 2002. PMID- 15282876 TI - Has the sun set on the golden age of NHS pensions? PMID- 15282877 TI - GPs to spend their pennies how and when they like. PMID- 15282878 TI - Primary care ideas. In the balance. PMID- 15282879 TI - Introduction: The origins and implications of a growing shortage of cardiologists. PMID- 15282880 TI - The myth of the Chinese culture, the myth of Chinese medical ethics. PMID- 15282881 TI - Religion-based arguments in the public arena: a Catholic perspective on euthanasia, Compassion in Dying v. State of Washington and Quill v. Vacco. PMID- 15282882 TI - Ryan White care amendments: mandatory HIV testing of newborns and a woman's right to privacy. PMID- 15282883 TI - The question remains: are there terminally ill patients who have a constitutional right to physician assistance in hastening the dying process. PMID- 15282884 TI - Persistent vegetative state: medical, ethical, religious, economic and legal perspectives. PMID- 15282885 TI - Undue economic influence on physician-assisted suicide. PMID- 15282886 TI - Correction to accreditation decision in Early Survey Policy Option 2. PMID- 15282888 TI - Corrections to 2004 requirements. PMID- 15282887 TI - Physician-assisted suicide: misconceptions and implications from a physician's perspective. PMID- 15282889 TI - Does early surgical repair of hip fractures improve patient outcomes? PMID- 15282890 TI - Physician assisted suicide under Jewish law. PMID- 15282891 TI - Aquinas and morphine: notes on double effect at the end of life. PMID- 15282892 TI - Rhetoric vs. reality: employer views on consumer-driven health care. AB - Because of rising premiums, employers are investigating new health insurance approaches that maintain workers' broad choice of providers while raising awareness of health care costs through increased patient financial responsibility. Employers' knowledge of new health plan products, including consumer-driven health plans and tiered-provider networks, has grown considerably in recent years, according to findings from the Center for Studying Health System Change's (HSC) 2002-03 site visit to 12 nationally representative communities. But employers are concerned that consumer-driven health plans would take considerable effort to implement without much cost savings. They also are skeptical that tiered-provider networks can adequately capture both cost and quality information in a way that is understandable to patients. PMID- 15282893 TI - Australia's Northern Territory: the first jurisdiction to legislate voluntary euthanasia, and the first to repeal it. PMID- 15282894 TI - Defining "health": three visions and their ramifications. PMID- 15282895 TI - Autonomy, informed consent, and genetic testing: an uneasy tension. PMID- 15282896 TI - Law, medicine, and trust. PMID- 15282897 TI - The governance of human genetics: policy discourse and constructions of public trust. AB - The collection of practices now commonly understood as 'biotechnology' poses a challenge to traditional mechanisms of regulating science and technology, just as it challenges traditional practices of science. The task of regulation is to reconcile the often conflicting political demands of protecting science, economy and the public interest. Public trust is the key measure of political success or failure. The purpose of this paper is to use policy discourse analysis as a vehicle for exploring the politics of the relationship between human genetics governance and public trust. An analysis of 30 policy documents produced six identifiable discourse streams relevant to public trust. These findings will be discussed, and an analysis of their impact on effective governance presented in conclusion. PMID- 15282898 TI - Anatomy of a media event: how arguments clashed in the 2001 human cloning debate. AB - This paper studies the distinctive role that staged media events play in the public understanding of genetics: they can focus the attention of the media, scientists and the public on the risks and benefits of genetic advances, in our case, cloning; they can accelerate policy changes by exposing scientific, legal and ethical uncertainties; the use of images, metaphors, cliches, and cultural narratives by scientists and the media engaged in this event can reinforce stereotypical representations of cloning, but can also expose fundamental clashes in arguments about cloning. The media event staged by two fertility experts in 2001 is here analysed as a case study. PMID- 15282899 TI - Clarification to scoring of management of information requirement for hospitals. PMID- 15282900 TI - 2003 sentinel event data. PMID- 15282901 TI - Slow-burning strategies see London drag behind in smoking cessation. PMID- 15282902 TI - On location. Birmingham and the Black Country. Chronic disease replaces waiting lists as new focus. PMID- 15282903 TI - Public health ideas. Raise a glass to public health. PMID- 15282904 TI - ETOX 11. Proceedings of the 11th European Workshop Conference on Bacterial Protein Toxins. Czech Republic, June 28-July 2, 2003. PMID- 15282905 TI - Ethics and practice: two worlds? The example of genetic counselling. AB - The aim of this paper is to work out the relationship between ethics and practice with reference to genetic counselling. First, the most important principles with respect to genetic counselling and to counsellor-client-interaction, are explained briefly. Then, we discuss what these principles might mean, when applied to the practice of counselling. To do so, we also look at some empirical data. Finally, we draw some conclusions. PMID- 15282906 TI - Affect- and self-based models of relationships between daily events and daily well-being. AB - The present study examined affect- self-based explanatory models of relationships between daily events and daily well-being. Twice a week for up to 10 weeks, participants described the events that occurred each day and provided measures of their daily affect, self-esteem, and depressogenic thinking. Participants also provided trait-level measures of affect, depression, and self-esteem. Measures of daily well-being representing each model covaried jointly and independently with daily negative and positive events. Positive events buffered the effects of negative events on daily self-esteem and daily depressogenic thinking, whereas there was no buffering effect for daily affect. More depressed people were more reactive to positive events, and those higher in trait PA were less reactive to negative events. Buffering effects for self-esteem were pronounced for those with lower trait self-esteem, and buffering effects for daily depressogenic adjustment were now more pronounced for those with higher trait negative affect. The results suggest that affect- and self-based models provide complementary perspectives on relationships between psychological well-being and daily events. PMID- 15282907 TI - The Australian joint inquiry into the Protection of Human Genetic Information. AB - The Australian Law Reform Commission (ALRC) and the Australian Health Ethics Committee are currently engaged in an inquiry into the Protection of Human Genetic Information. In particular, the Attorney-General and the Minister for Health and Ageing have asked us to focus, in relation to human genetic information and tissue samples, on how best to ensure world's best practice in relation to: privacy protection; protection against unlawful discrimination; and the maintenance of high ethical standards in medical research and clinical practice. While initial concerns and controversies have related mainly to aspects of medical research (e.g. consent; re-use of samples) and access to private insurance coverage, relevant issues arise in a wide variety of contexts, including: employment; medical practice; tissue banks and genetic databases; health administration; superannuation; access to government services (e.g. schools, nursing homes); law enforcement; and use by government authorities (e.g. for immigration purposes) or other bodies (e.g. by sports associations). Under the Australian federal system, it is also the case that laws and practices may vary across states and territories. For example, neonatal genetic testing is standard, but storage and retention policies for the resulting 'Guthrie cards' differ markedly. Similarly, some states have developed highly linked health information systems (e.g. incorporating hospitals, doctors' offices and public records), while others discourage such linkages owing to concerns about privacy. The challenge for Australia is to develop policies, standards and practices that promote the intelligent use of genetic information, while providing a level of security with which the community feels comfortable. The inquiry is presently reviewing the adequacy of existing laws and regulatory mechanisms, but recognizes that it will be even more important to develop a broad mix of strategies, such as community and professional education, and the development of official standards and industry codes that reflect emerging international best practice in the area. PMID- 15282908 TI - From energy efficiency under stress to rapid development and a long life in natural populations. AB - Simplistically, high fitness depends upon high energy efficiency in the stressful habitats of organisms in the wild. Rapid development and high survival to adulthood should be followed by long-lived stress-resistant genotypes under this reductionist model. Empirical evidence is very limited because of the common use of benign laboratory environments, which remains a major difficulty in understanding relationships between life-history traits under more natural settings. Heterozygotes tend to show greater energy efficiency than do corresponding homozygotes especially in stressful environments, which leads the above connections among fitness traits. In particular rapid development and increased longevity should be correlated, and underwritten by the availability of metabolic energy. Empirical work is needed based upon severe stresses where a small environmental perturbation cases lethality; drought, heat and nutritional inadequacy are suitable candidate stresses. The importance of viewing fitness in energy terms is emphasized throughout. A byproduct is the potential to fuse functional and evolutionary biology under stressful environments. PMID- 15282909 TI - Rosy and Jim: the mystery of the double helix. PMID- 15282910 TI - Governing genetics: reifying choice and progress. AB - The governance of genetics involves a wide range of policy networks and covers a considerable array of genetic technoscience. Despite this apparent diversity, the uniformity of some genetic governance requires investigation. Reviewing policy documents on genetic patenting and embryonic stem cell research, I shall argue that policy networks often conceive of the ethical aspects of these practices in similar ways. In particular, I shall argue that individual choice and medico scientific progress are common rhetorical devices in their frameworks. I shall end by commenting upon the implications of these trends for the future. PMID- 15282911 TI - The politics of cloning: mapping the rhetorical convergence of embyros and stem cells in parliamentary debates. AB - In April 2001, the 1990 Human Fertilisation and Embryology Act (HFE Act) was amended to allow stem cell research to use human embryos. By identifying what Mulkay calls "discursive regularities" [Mulkay, M. (1993) Rhetorics of hope and fear in the great embryo debate, Social Studies of Science, 23, p. 723], this paper examines the rhetorical strategies of and manoeuvrings over the meanings of stem cells, cloning, and embryos within the parliamentary context. I focus upon the "return to the embryo question" and the significance" of this for the stem cell debates in terms of form and content. This feeds into an analysis of the ways in which two specific groups are discursively invoked and constructed--those with diseases and disabilities who have been identified as likely to benefit from stem cell therapies, and couples undergoing fertility treatment who are needed to donate spare embryos. In doing so, I draw upon similar analyses of the earlier embryo debates--those of Mulkay, Franklin, Kirejcyk and Spallone--leading up to the establishment of the 1990 HFE Act. In conjunction with these analyses, I am able to identify parallels between the rhetorical devices mobilized and the legislative outcomes. PMID- 15282912 TI - Biological weapons, genetics and social analysis: emerging responses, emerging issues--I. AB - Recent terrorist attacks in the USA have generated significant attention in many countries to the threats posed by biological weapons. In response to these events and the spectre of future attacks, bioscientists and professional organizations have begun or intensified asking questions about the possible malign applications of their research. As Part I of a two-part article, this paper surveys how genetics might contribute to the development of novel forms of weaponry. It is further argued that the dilemmas and difficulties facing bioscientists pose pressing and thorny questions for the hitherto agendas and orientations of those concerned with the social, ethical and political implications of genetics. Part II will examine the emerging responses initiated by biomedical organizations and spokespersons in the US and the UK. This will be done with a view to asking how scientific and medical research communities are defining and policing notions of professionalism, responsibility and accountability. On the basis of this, suggested lines for future social analysis will be offered. PMID- 15282913 TI - International ethics and equity. PMID- 15282914 TI - Empirical research in bioethics: the method of "oppositional collaboration". PMID- 15282915 TI - Moral intuitions as a source for empirical ethics. PMID- 15282916 TI - Delays in implementing the lessons from empirical studies on bioethics to ethics committees in Asia. PMID- 15282917 TI - Global inverse care: what to do about it. PMID- 15282918 TI - International regulations and medical research in developing countries: double standards or differing standards? PMID- 15282919 TI - Research ethics in Russia. PMID- 15282920 TI - Ethics committees in Northern Europe. PMID- 15282921 TI - Ethics committees in Finland: their levels, methods, and point. PMID- 15282922 TI - Ethical decision making in committee. A view from Germany. PMID- 15282923 TI - The development of clinical ethics support in the United Kingdom. PMID- 15282924 TI - Ethics committees in the United States. PMID- 15282925 TI - Research ethics committees in Argentina and South America. PMID- 15282926 TI - Building a new culture of ethics in research involving humans in Brazil. PMID- 15282927 TI - Passing of a giant. PMID- 15282928 TI - Ethics committees in Australia and New Zealand: a critique. PMID- 15282929 TI - Ethics committees in Asia. PMID- 15282930 TI - Research ethics committees--the future. PMID- 15282931 TI - The end of an era: Wayne Streilein and his impact on transplantation immunology. PMID- 15282932 TI - A nonscientific tribute to another scientist. PMID- 15282933 TI - The ethical implications of direct-to-consumer pharmaceutical advertising. PMID- 15282934 TI - The role of perceived specificity level of failure events in self-enhancement and in constructive self-criticism. AB - Three studies tested the role played by perceived specificity level of failures in self-enhancement and in constructive self-criticism. The first study demonstrated that perceived specificity level of events can serve as a self protection mechanism (N = 137). The second study, based on retrospective reports of past failures and their implications, showed that perception of failures as specific induced a higher level of constructive self-criticism (N = 171). The third study tested reactions to failures induced in the laboratory. It was found that self-improvement processes are more pronounced and negative emotional reaction is weaker in failures that are specific (shape perception test) than in those that are global (intelligence test). Statistical control over perceived severity of the failure diminished the difference between the two conditions in negative emotional reaction but not in self-improvement (N = 84). PMID- 15282935 TI - Association between atopy and cryptorchidism. PMID- 15282936 TI - Eosinophilic meningitis secondary to allergic Aspergillus sinusitis. PMID- 15282937 TI - Soluble decoy receptor 3: increased levels in atopic patients. PMID- 15282938 TI - Cysteinyl leukotrienes and collagen type I synthesis in asthma. PMID- 15282939 TI - Association between TARC C-431T and atopy and asthma in children. PMID- 15282940 TI - Exhaled air temperature and eosinophil airway inflammation in allergic asthmatic children. PMID- 15282941 TI - Regarding "Trichosporon pullulans infection in 2 patients with chronic granulomatous disease". PMID- 15282942 TI - Exposure-dose-response: critical considerations in toxicology and immunology alike. PMID- 15282943 TI - Cellular automata and its advances to drug therapy for HIV infection. AB - This paper gives an over view of the use of cellular automata (CA) model of drug therapy for HIV infection. Nonuniform CA is employed to simulate drug treatment of HIV infection, where each computational domain may contain different CA rules, in contrast to normal uniform CA models. Ordinary (or partial) differential equation models are insufficient to describe the two extreme time scales involved in HIV infection (days and decades), as well as the implicit spatial heterogeneity. Zorzenon and Coutinho [Phy Rev Lett, 16 (2001) 1] reported a cellular automata approach to simulate three-phase patterns of human immunodeficiency virus (HIV) infection consisting of primary response, clinical latency and onset of acquired immunodeficiency syndrome (AIDS). But here we present a related model, based on non-uniform CA to study the dynamics of drug therapy of HIV infection. The main aim in this model is to simulate the four phases (acute, chronic, drug treatment responds and onset of AIDS). The results shown here indicate that both simulations (with and without treatments) evolve to the relatively same steady state (characteristics of Wolfram's class II behavior). Different kinds of drug therapies can also be simulated in this model, which can be found useful for developing a proper drug therapy. PMID- 15282944 TI - Nitrogen fixation and carbon metabolism in legume nodules. AB - A large amount of energy is utilized by legume nodules for the fixation of nitrogen and assimilation of fixed nitrogen (ammonia) into organic compounds. The source of energy is provided in the form of photosynthates by the host plant. Phosphoenol pyruvate carboxylase (PEPC) enzyme, which is responsible for carbon dioxide fixation in C4 and crassulacean acid metabolism plants, has also been found to play an important role in carbon metabolism in legume root nodule. PEPC mediated CO2 fixation in nodules results in the synthesis of C4 dicarboxylic acids, viz. aspartate, malate, fumarate etc. which can be transported into bacteroids with the intervention of dicarboxylate transporter (DCT) protein. PEPC has been purified from the root nodules of few legume species. Information on the relationship between nitrogen fixation and carbon metabolism through PEPC in leguminous plants is scanty and incoherent. This review summarizes the various aspects of carbon and nitrogen metabolism in legume root nodules. PMID- 15282945 TI - Effect of simultaneous exposure to lead and cadmium on gonadotropin binding and steroidogenesis on granulosa cells: an in vitro study. AB - Effects of lead (Pb) and cadmium (Cd) both alone or in combination on the binding of LH and FSH on isolated granulosa cells were studied. Granulosa cells isolated from proestrous rats were incubated (in vitro) with lead acetate and/or cadmium acetate (0.03 microM of Pb or Cd) for 1 hr. LH binding was dropped to 84% in Pb treated cells, 72.5% in Cd treated cells and 74.8% in combined metal treated cells compared to control. FSH binding dropped to 85.5% in Pb treated cells, 71.16% in Cd treated cells and 72.5% in combined metal treated cells compared to control. Activity of 17beta Hydroxy Steroid Dehydrogenase (17betaHSDH), a key steroidogenic enzyme was reduced by 52% in Cd and 37% in combined metal exposed cells whereas Pb exposed cells showed 31% reduction in the enzyme activity. Pretreatment with SH groups protectants (glutathione [GSH], dithiothretol [DTT]) and zinc caused an ameriolation in enzyme activity whereas Zn pretreatment showed an increase in gonadotropin binding in metal exposed cells. These results suggest that both Pb and Cd can cause a reduction in LH and FSH binding, which significantly alters steroid production in vitro and exerts a direct influence on granulosa cell function. PMID- 15282946 TI - Inhibitory effect of manganese on contraction of isolated rat aorta. AB - The effects of MnCl2 on vascular smooth muscle contraction induced by noradrenaline (NA) and KCl were investigated. Rings segments from rat aorta were isolated and changes in isometric tension recorded. MnCl2 (10 microM and 1 mM) significantly attenuated the contractile responses to NA and KCI. There were also reductions in the contractile responses to CaCl2 in NA- and KCl-stimulated rings, after pretreatment with MnCl2. The magnitude of the phasic contraction to NA was significantly reduced in presence of MnCl2. The results suggest that MnCl2 inhibits vascular smooth muscle contraction by influencing a Ca2+-mediated mechanism. PMID- 15282947 TI - Experimental pathogenicity evaluation of Mycoplasma canadense from bovine mastitis in vitro and in vivo laboratory models. AB - Mycoplasma canadense, a clinical isolate from milk of a mastitic buffalo, was experimentally tested for its pathogenic potential in hamster tracheal ring and rabbit fallopian tube explant organ cultures (in vitro) and rat and rabbit mammary gland (in vivo) models. The activity percentage reduction in M. canadense infected hamster tracheal rings was 99.1% in comparison to 16.4% in control rings. Mycoplasma canadense, also induced complete ciliostasis at 11-day post infection in rabbit fallopian tube explants. Histopathological lesions in these infected organ cultures were loss of cilia, desquamation or denudation of epithelium, infiltration of inflammatory cells and proliferation of macrophages as well as oedema in lamina propria. At the end of the experiments, M. canadense organisms were reisolated in pure colonies from the infected but not the control organ cultures. In the rat and rabbit mammary glands, M. canadense organisms persisted upto 6-day and 7-day postinfection, respectively and caused histopathological changes suggestive of subacute to chronic mastitis during the experimental period. The results indicate that the tested M. canadense clinical isolate was virulent. PMID- 15282948 TI - Potentiation of insecticidal activity of Bacillus thuringiensis subsp. kurstaki HD-1 by proteinase inhibitors in the American bollworm, Helicoverpa armigera (Hubner). AB - The effect of crude proteinase inhibitor extracts from seeds of different crop plants (black gram, chickpea, chickling vetch, finger millet, French bean, green gram, horse gram, lentil, pea and soybean) on the insecticidal activity of B. thuringiensis var. kurstaki HD-1 was investigated against neonate larvae of H. armigera by diet incorporation method. The larval mortality due to crude proteinase inhibitors alone (5% seed weight equivalent) ranged from 4.1 to 19.1%; the maximum mortality with finger millet and the minimum with pea var. DDR-23. A mixture of B. thuringiensis var. kurstaki HD-1 (10 ppm) and proteinase inhibitor (5% seed weight equivalent) was synergistic in larval mortality with respect to proteinase inhibitors of pea var. DMR-16, chickling vetch var. RLK-1098 and B101 212, lentil var. ILL-8095 and L-4076, soybean var. PK-1042, PK-416 and Pusa-22, chickpea var. Pusa-413, French bean (Chitra) and black gram; and antagonistic with respect to those of finger millet, horse gram and kidney bean. The larval growth reduction with crude proteinase inhibitors alone ranged from 17.9 to 53.1%; the maximum growth reduction with soybean var. PK-1042 and minimum with lentil var. L-4076. A mixture of B. thuringiensis var. kurstaki and proteinase inhibitor was synergistic in growth reduction with respect to proteinase inhibitors of lentil var. ILL-8095, and L-4626 and antagonistic with respect to that of finger millet. The midgut proteinase inhibition with crude seed extracts (3.3% seed weight equivalent) ranged from 9.3 to 60.9% and was negatively correlated with larval mortality. These results showed that interactive effect of B. thuringiensis var. kurstaki HD-1 and proteinase inhibitors in the larvae of H. armigera depended upon the quality and quantity of proteinase inhibitors, which vary widely in different plants. PMID- 15282949 TI - Development and mechanisms of resistance to Bacillus thuringiensis endotoxin Cry1Ac in the American bollworm, Helicoverpa armigera (Hubner). AB - The American bollworm, H. armigera, evolved 31-fold resistance to selection pressure of B. thuringiensis endotoxin Cry1Ac within six generations. The Cry1Ac selected larvae of H. armigera showed cross-resistance to Cry1Aa and Cry1Ab both in terms of mortality and growth reduction. Studies on mechanisms of resistance to Cry1Ac showed that proteases of resistant insects degraded Cry1Ac faster than those of susceptible insects, which led to the relative unavailability of toxin of about 58 kDa for binding and perforation of midgut epithelial membrane of the target insect. Besides, resistant and susceptible populations of H. armigera differed in the binding of their receptors with Cry1Ac toxin. These results suggest the possibility of both mechanisms existing in imparting resistance. These findings mandate the necessity of B. thuringiensis resistance management for usage of B. thuringiensis either as a conventional insecticide or through transgenic crops. PMID- 15282950 TI - Protective effect of Terminalia chebula against experimental myocardial injury induced by isoproterenol. AB - Cardioprotective effect of ethanolic extract of Terminalia chebula fruits (500 mg/kg body wt) was examined in isoproterenol (200 mg/kg body wt) induced myocardial damage in rats. In isoproterenol administered rats, the level of lipid peroxides increased significantly in the serum and heart. A significant decrease was observed in the activity of the myocardial marker enzymes with a concomitant increase in their activity in serum. Histopathological examination was carried out to confirm the myocardial necrosis. T. chebula extract pretreatment was found to ameliorate the effect of isoproterenol on lipid peroxide formation and retained the activities of the diagnostic marker enzymes. PMID- 15282951 TI - Antiinflammatory and antiulcer activities of phytic acid in rats. AB - Maximum antiinflammatory activity of phytic acid (PA) was seen at an oral dose of 150 mg/kg in the carrageenan induced rat paw edema model. Although PA showed ability to prevent denaturation of proteins, it showed less antiinflammatory activity than ibuprofen. Ability of PA to bring down thermal denaturation of proteins might be a contributing factor in the mechanism of action against inflammation. PA, at all the doses tested, showed significant protection from ulcers induced by ibuprofen, ethanol and cold stress, with a maximum activity at 150 mg/kg. There was a significant increase in gastric tissue malondialdehyde levels in ethanol treated rats but these levels decreased following PA pretreatment. Moreover, pretreatment with PA significantly inhibited various effects of ethanol on gastric mucosa, such as, reduction in the concentration of nonprotein sulfhydryl groups, necrosis, erosions, congestion and hemorrhage. These results suggested that gastro-protective effect of PA could be mediated by its antioxidant activity and cytoprotection of gastric mucosa. PMID- 15282953 TI - A partial sequence of lipoxygenase gene from genomic DNA of aromatic rice (Oryza sativa L.). AB - Aromatic (Bas-370, PB-1) and non-aromatic (Pusa-677, Pusa-834) rice were selected for the characterization and for distribution of lipoxygenase (Lox) genes. Polymorphism was observed when genomic DNA of rice varieties was hybridized with a heterologous lipoxygenase probe. A distinct polymorphic fragment (approximately 1.2 kb) was found in Bas-370. Sub-genomic library of Bas-370 was constructed and screened with LoxA probe. The smallest putative clone (pBas-14) of approximately 1.2 kb was sequenced. Complete nucleotide and deduced amino acid sequence showed the clone was 1134 bp long and comprised of 378 amino acid residues. PCR amplification of genomic DNA from four rice varieties with a soybean Lox primer also showed a polymorphic fragment of size approximately 600 bp (amplicon) in aromatic varieties that was sequenced directly. Nucleotide sequence alignment between pBas-14 and amplicon concluded that the amplicon was a part of the insert pBas-14. PMID- 15282952 TI - Phytochemical studies and effect on urine volume of Glossostemon bruguieri Desf. constituents. AB - Two new flavonoids, takakin 7-O-glucoside (1) and (2) bucegin 7-O-glucoside, and six other known compounds (3-8), takakin, isosctullarien, its 7-O-glucoside, takakin 8-O-glucoside, xanthotoxin and esculetin, were separated and identified from Glossostemon bruguieri. The new compounds were characterized using modern spectroscopic techniques, including UV spectroscopy, proton nuclear resonance (1HNMR), carbon thirteen nuclear resonance (13CNMR), homomolecular quantum coherance (HMQC), heteromolecular bonding connectivity (HMBC) and chemical ionization mass spectra (CI). The effect on rats urine volume of the plant powder, its ethanolic extract, (500 mg kg(-1)) along with four of the purified compounds (1,4-6), (100 mg kg(-1)) are described. Eight groups of albino rats (200-300 g body weight) (n=5 for each group) were used in the tests for a one time treatment, and other seven groups (150-180 g body weight) (n=5 for each group) were tested using the same dose with repeated administration for 15 days. The rat sera were collected and used to determine liver and kidney functions based on alanine amino transferase (ALT) and aspartate amino transferase (AST) for both single and repeated administration. Levels of urea, creatinine and uric acid were determined for both sets of experiments. The toxic effects of both the powder and its alcoholic extract were also studied on mice to determine their LD50, both materials proved to be non-toxic up to 2500 mg kg(-1) body weight. PMID- 15282954 TI - Role of phenolics and boron in reproductive success in seasonally transient sterile Tecoma stans L. AB - Quantitative and qualitative analysis of phenolics and boron in stigma of transient sterile Tecoma stans L. during seedless (May-July), partially seedbearing (August-November, April) and seedbearing periods (December-March) was made. UV absorption profile of stigmatic exudates indicated the presence of simple phenolics. Total phenolics were higher in stigma during seedless period. Thin layer chromatographic analysis of stigmatic extracts exhibited only three principal spots. Mass spectrophotometry showed the presence of derivatives of cinnamic acid, namely, caffeic acid in these spots. Quantity of boron in stigma during seedless period was lowest but the difference with other periods was not significant. It was suggested that the accumulation of higher quantity of caffeic acid in the stigma during seedless period due to high temperature (40 degrees-45 degrees C) could lead to inhibition of pollen germination in vivo, thereby rendering the plants seedless. This was confirmed by inhibition of in vitro pollen germination in the basal medium containing higher quantity of caffeic acid. PMID- 15282955 TI - Production of lipase in a fermentor using a mutant strain of Corynebacterium species: its partial purification and immobilization. AB - Extracellular Corynebacterium lipase was produced using a 2.5 L Chemap fermentor using 1300 ml fermentation medium at temperature 33 degrees C, agitator speed 50 rpm, aeration rate 1 VVM having KLa 16.21 hr(-1). Crude lipase was purified by salting out method followed by dialysis and immobilized using calcium alginate gel matrix followed by glutaraldehyde cross linking Purification process increased specific activity of enzyme from 2.76 to 114.7 IU/mg. Activity of immobilized enzyme was 107.31 IU/mg. Optimum temperature for purified and immobilized enzyme activity were 65 degrees and 50 degrees C respectively. Optimum pH was 8.0 in both the cases, Km and Vmax value for purified lipase were 111.1 micromol/min and 14.7% respectively. Ca2+ (5 mM) was found to be stimulator for enzyme activity. Immobilized lipase retained 68.18% of the original activity when stored for 40 days. PMID- 15282956 TI - Effect of glyphosate toxicity on growth, pigment and alkaline phosphatase activity in cyanobacterium Anabaena doliolum: a role of inorganic phosphate in glyphosate tolerance. AB - Response of glyphosate toxicity on photoautotrophic cyanobacterium A. doliolum and its mutant strain was investigated. Chlorophyll a content of both the wild type and mutant strain in the presence of glyphosate (N-phosphonomethyl glycine) initially showed an increasing trend when supplemented with Pi and a declining tendency under the Pi-starved condition. The results suggested that both the wild type and mutant strains were more sensitive to glyphosate in the absence of phosphate. Alkaline phosphatase activity of wild type strain in the presence of Pi, enhanced in response to addition of glyphosate (40 microg/ml), but the activity remained unaltered by addition of glyphosate in the Pi-starved cells, whereas the alkaline phosphatase activity in the mutant strain under both Pi starved as well as unstarved conditions was stimulated (approximately 5.4 and 3.1 fold, respectively) by addition of glyphosate. The results on alkaline phosphatase activity indicated a glyphosate-induced depletion in the phosphate content of the cells, particularly in the mutant strain, as evident from the stimulated activity of alkaline phosphatase. It is suggested that enzyme activity in the Pi-starved wild type cells may not be influenced any further by glyphosate, as cellular phosphate reserve might not be available for further depletion. PMID- 15282957 TI - Hydrolysis of organophosphorus compounds by an esterase isozyme from insecticide resistant pest Helicoverpa armigera. AB - Esterase activity of resistant and susceptible H. armigera were compared in gels with different substrate such as naphthyl acetate, naphthyl phosphate, paraoxon and monocrotophos. Whole body extract of resistant H. armigera hydrolyzed paraoxon, monocrotophos and naphthyl phosphate in gels. Resistant H. armigera showed high esterase, phosphatase and paraoxon hydrolase activity compared to susceptible ones. PMID- 15282958 TI - Ferric reductase, superoxide dismutase and alkaline phosphatase activities in siderophore producing fungi. AB - Enzymes associated with release of iron from internalized ferrated siderophore (ferrisiderophore reductase), with damage to the cell at high iron concentration (superoxide dismutase) and siderophore synthesis (alkaline phosphatase), were examined in 3 test fungi viz., Aspergillus sp. ABp4, Aureobasidium pullulans and Rhizopus sp. Extracellular ferrisiderophore reductase activity was present in all the three fungi, but Aureobasidium pullulans, that showed the highest activity (84.3 microM min(-1)), was the only one to produce intra-cellular ferric reductase (147.9 microM min(-1)). Superoxide dismutase was produced by Aureobasidium pullulans and Rhizopus sp., but not by Aspergillus sp. ABp4, that showed intra-cellular enzyme activity in case of ferric reductase and alkaline phosphatase. Maximum SOD activity was seen in Aureobasidium pullulans both extra cellularly (93.83 ng ml(-1)) and intra-cellularly (57.14 ng ml(-1)). All the test fungi examined, produced intra-cellular alkaline phosphatase. There was no extracellular alkaline phosphatase. Among the three fungi, Aureobasidium pullulans showed highest alkaline phosphatase activity (129.9 microM min(-1)) and Aspergillus sp. ABp4 the least (76.4 microM min(-1)). PMID- 15282959 TI - Vitamin E prevents nonylphenol-induced oxidative stress in testis of rats. AB - In the present study we have investigated if administration of nonylphenol induced oxidative stress in various subcellular fractions of adult rat testis and the effect of vitamin E on reactive oxygen species mediated nonylphenol toxicity. Male rats were administered orally with nonylphenol at 1, 10 and 100 microg/kg body weight per day for 45 days with and without supplementation of vitamin E (20 mg/kg body weight). In nonylphenol-treated rats the activities of antioxidant enzymes superoxide dismutase and glutathione reductase decreased significantly while the levels of lipid peroxidation increased significantly in the crude homogenate and in the mitochondrial and microsome-rich fractions of testis. Co administration of nonylphenol and vitamin E did not cause changes in the activities of antioxidant enzymes in various subcellular fractions of rat testis. The results suggest that graded doses of nonylphenol elicit depletion of antioxidant defence system in rat testis, indicating nonylphenol induced oxidative stress in the testis of rats which could be reversed by the administration of vitamin E. PMID- 15282960 TI - Role of Arogh, a polyherbal formulation to mitigate oxidative stress in experimental myocardial infarction. AB - Antioxidant role of Arogh in isoproterenol induced myocardial infarction in rats has been studied. The activity of heart tissue antioxidants like glutathione, superoxide dismutase, catalase, glutathione peroxidase and glutathione-s transferase were significantly decreased in isoproterenol administered group. The activity of ceruloplasmin and levels of glutathione, vitamins E and C were also found to be substantially decreased in serum with a concomitant rise in lipid peroxide levels after isoproterenol exposure to rats. The synergistic effect of Arogh pretreatment, significantly suppressed the alterations induced by isoproterenol alone in rats. PMID- 15282961 TI - Effect of different temperature on starch synthase activity in excised grains of wheat cultivars. AB - Excised grains of wheat (Triticum aestivum) varieties HD 2285 (relatively tolerant) and HD 2329 (susceptible type) were incubated for 1 hr at 15 degrees, 25 degrees, 35 degrees and 45 degrees C. In an another treatment, excised grains were incubated for 1 hr at increasing temperature (15 degrees, 25 degrees, 35 degrees and 45 degrees C) continuously, thus exposing the grains to gradual rise in temperature. The above treated grains were then analysed for the activity of soluble starch synthase (SSS) and granule bound starch synthase (GBSS) assayed at 20 degrees C. SSS activity decreased as the pre-exposure temperature was higher, though the tolerant variety showed lesser decrease. Decrease in SSS activity was lesser when excised grains were exposed to gradual rise in temperature from 15 degrees to 45 degrees C as compared to direct exposure to 45 degrees C. Pre exposure of excised grains to different temperatures however, had no significant effect on GBSS activity. PMID- 15282962 TI - Molecular detection of drug resistance in Mycobacterium leprae. PMID- 15282963 TI - Inter-rater reliability of WHO 'disability' grading. AB - The World Health Organization 'disability' grading system was introduced in 1960. It is mainly used as an indicator for early diagnosis or reporting. Disability grades are usually aggregated at national levels. Comparison of data with previous years or comparison of data between programmes may show that patients report earlier for treatment, alternatively, are diagnosed earlier, that is without, or with fewer 'disabilities'. Despite its long and universal use as an epidemiological parameter, the WHO disability grading has not been the subject of reliability studies. In this study, three testers unfamiliar with the grading prior to the study each graded 65 (former) leprosy affected persons. The weighted kappa ranged from 0.87 to 0.89 (95% CI 0.73-1.00) for the highest score and from 0.90 to 0.96 (95% CI 0.90-0.99) for the EHF (Eye, Hand, Foot) score, indicating excellent reliability. The study shows that with limited training and little experience a high degree of reliability in grading 'disabilities' between testers is attainable. PMID- 15282964 TI - Nerve thickening in leprosy patients and risk of paralytic deformities: a field based study in Agra, India. AB - This paper examines the extent of nerve thickening among leprosy patients detected in the field in Agra district. All the clinically diagnosed leprosy patients were examined in detailed for thickening of local cutaneous nerves and peripheral nerve trunks. In each case all the major nerve trunks in both upper and lower extremities, forehead and neck were examined for thickening. Nerve thickening was found in 94% of multibacillary (MB) patients and among 52% paucibacillary (PB) patients. Nerve thickening was found to be more in males, in prevalent cases than in new (untreated) cases and increased significantly with age and delay in diagnosis (P<0.001). Visible deformities of grade > or =2 were found in 10% (58/573) of the leprosy patients; paralytic deformity accounted for 78% (45/58). Claw hand alone or in combination was seen in 82% (37/45) of patients with paralytic deformities. Risk (odd ratio) for deformities was observed to be high (15-18 times) with increasing number of nerves among patients with neuritic leprosy but correlated with delay in diagnosis of over 5 years. Likewise, deformities were more often seen in those with skin lesions, provided they had > or =3 thickened nerves. Findings suggest that early detection and treatment is useful in preventing deformities. PMID- 15282965 TI - Impairments in multibacillary leprosy; a study from Brazil. AB - This is a retrospective cohort study of 103 multibacillary leprosy patients (18% BB, 48% BL and 34% LL) followed during and after treatment, in a tertiary referral centre with an outpatient clinic in an endemic area in Brazil, for an average period of 65 months since the start of multidrug therapy (24-dose MDT). The objective of the study was to identify the role of overt neuritis (presence of pain in a peripheral nerve trunk, with or without enlargement or neural function damage), in the development of impairments. They were evaluated using the World Health Organization disability grade before treatment, at the end of the treatment, and at the end of the follow-up period. Thirty-four percent of patients presented overt neuritis during MDT, and 45% had overt neuritis episodes during the follow-up period; the most commonly affected nerves were ulnar, fibular and posterior tibial nerves, and the neuritic episodes were carefully treated with steroid therapy and physiotherapy. Impairments were associated with: affected (painful and/or thick) nerves at diagnosis (P < 0.005); delay in diagnosis (P = 0.010); impairments already present at the start of treatment (P = 0.00041 at the end of MDT, and P = 0.000013 at the end of follow-up); occurrence of overt neuritis episodes during MDT (P = 0.0016) or the whole follow-up (P = 0.015). These data draw attention to the importance of early diagnosis and of good neurological examination throughout the follow-up, as well as suggest the importance of neuritis in the induction of impairments in multibacillary leprosy. PMID- 15282966 TI - The fulfilment of health care needs of leprosy patients from Kaski District, Nepal. AB - Hospital records of 142 leprosy patients from Kaski district in Western Nepal were reviewed to assess their use of leprosy related health services and the fulfilment of these needs. Use of services was reviewed from diagnosis until release from treatment. Voluntary muscle and sensory testing were on average done 15.2 times per patient Of MB patients, 65.5% had longer intervals between testing than recommended. A course of prednisolone was indicated in 40% of cases, but 10% of needed courses were not given. Twenty-eight percent needed protective footwear. Of the cohort, 10% had complicated ulcers and 28% had at least one admission. Paralytic impairments that could be corrected were present in 10% of the cohort. PMID- 15282967 TI - Analysis on the detection of new leprosy cases before, during and after the year of leprosy elimination campaigns. AB - To analyse the impact on of case finding of leprosy elimination campaigns (LECs), data on newly detected leprosy cases in a leprosy endemic area were collected before, during and after the year of LEC. The number of new leprosy cases detected during the year of LEC was significantly higher than previously. The number of newly detected cases after the year of LEC was similar to that of detected before the year of LEC in counties with persisting case finding activities. However, the number of newly detected cases after the year of LEC significantly decreased in counties without active case finding activities. The average distance from the homes of leprosy cases detected during LEC to the leprosy control unit at the count town was 62.8 km, which is farther than that of other leprosy cases detected before and after the year of LEC. The average time from disease onset to diagnosis of leprosy cases detected after the year of LEC shortened. The results also showed that carrying out LECs is unlikely to have a significant impact on the trend of case finding within a short time in local areas, but it may improve some indicators of leprosy patients and so promote leprosy control in local areas. PMID- 15282968 TI - Leprosy control in the Republic of Yemen: co-operation between government and non government organizations, 1989-2003. AB - Although the prevalence rate of leprosy in the Republic of Yemen has dropped below the WHO elimination level of less than one case per 10,000 of the population, it is still regarded as a serious public health problem calling for continued vigilance, notably in the detection and treatment of hidden and undiagnosed cases. In the past, religious misinterpretation has generated adverse behaviour patterns towards people affected by leprosy, characterized by aggression, negligence and isolation. Until about 1982, following a visit of a leprologist (Dr S. K. Noordeen) from the World Health Organization, there was no leprosy control programme and attempts to establish one remained ineffective until in 1989, when an agreement was signed between the Ministry of Public Health and Population and the German Leprosy Relief Association. This led to the development of a leprosy control programme in four governorates, later extended to the rest of the country. This paper describes the progress made in the control of leprosy in the Yemen, 1989-2003, by the Ministry of Health and Population and the GLRA, in association with two local societies. PMID- 15282969 TI - Intra-abdominal, crystal-storing histiocytosis due to clofazimine in a patient with lepromatous leprosy and concurrent carcinoma of the colon. AB - We report a case with abdominal complications of clofazimine treatment which included blackish discolouration of the lymph nodes, omentum and peritoneum. A 44 year-old female with lepromatous leprosy and a history of adverse reaction to clofazimine 2 years previously, presented with rectosigmoid junction adenocarcinoma. Laparotomy revealed an inoperable tumour with pigmentation of the bowel, serosa and peritoneum. A second operation had o be performed for transverse loop colostomy and a mesenteric lymph node biopsy sent for frozen section showed typical clofazimine crystals. Despite widespread use for many years in the treatment of leprosy, this drug is not known to be carcinogenic and this case provides no evidence for an association or link between its use and the patient's cancer. Apart from its use in leprosy, clofazimine may be used in the treatment of disseminated Mycobacterium avium-intracellulare infection, Buruli ulcer due to M. ulcerans and occasionally in other mycobacterial infections. An awareness of the rare side-effect described above may help in the clinical assessment and management of such cases, including the avoidance of unnecessary laparotomy. PMID- 15282970 TI - Bullous erythema nodosum leprosum: a case report from Nepal. AB - A patient with lepromatous leprosy, while on WHO multidrug therapy (MDT) for multibacillary disease, was diagnosed as having dapsone syndrome with recurrent episodes of bullous lesions on the lower extremities for 4-5 years. The lesions were associated with high-grade fever. Examination revealed multiple hypopigmented macules on the limbs. Multiple atrophic scars were also found on the buttocks and lower limbs. Bilateral ulnar, radial cutaneous and lateral popliteal nerves were thickened. On day 10 of WHO-MB-MDT he developed a flaccid bulla on the lower leg. Skin slit smear showed a bacterial index (BI) of 3+ and the histopathology was consistent with type II reaction. High dose corticosteroid therapy was started but he continued to have new lesions, and was therefore referred to a centre where thalidomide was available. Clinical response was good and he remained symptom-free after gradual reduction in dosage. ENL should be differentiated from bullous drug reactions, pemphigus vulgaris, bullous pemphigoid and other blistering diseases. PMID- 15282971 TI - A case of post-partum borderline tuberculoid leprosy complicated by a median nerve abscess, peptic ulceration and rifampicin-induced haemolytic renal failure. AB - We report a case of borderline tuberculoid leprosy complicated by a median nerve abscess, acute renal failure secondary to rifampicin-induced haemolysis and duodenal ulceration secondary to steroid use. Rifampicin induced hameolysis is a rare and probably under-reported complication of leprosy multi-drug therapy. It should be considered when patients complain of flu-like symptoms after taking their monthly rifampicin. PMID- 15282972 TI - Tuberculoid leprosy confined to glans penis in two cases. PMID- 15282973 TI - A strategy to improve the ml flow test for detection of anti-phenolic glycolipid 1 antibodies. PMID- 15282974 TI - Potential impacts of poverty alleviation activities on reducing leprosy transmission. PMID- 15282975 TI - [The stages of comprehensive periodontal therapy--diagnosis-driven treatment planning]. AB - The rationale and the stages of the comprehensive periodontal treatment are overviewed. The consecutive phases are logically following each other. Basically three major therapeutical categories can be established, such as: 1. absolutely healthy from periodontal point of view, 2. gingivitis, 3. periodontitis. Within the periodontitis group the basic principles and strategies are totally different for the management of chronic and aggressive periodontitis cases. Principally the comprehensive periodontal therapy can be divided into four main phases: phase I. initial or cause related therapy, phase II. surgical therapy, phase III. periodontal reconstruction, and phase IV. periodontal maintenance. The classic non-surgical and surgical techniques can cure inflammation, arrest the progression of periodontal destruction and restore gingival-periodontal health, and the outcome is quite predictable for gingivitis and chronic periodontitis. The long-lasting results are dependent on the patients' motivation and maintenance. The late therapeutical results and the maintenance of aggressive periodontitis cases are less predictable. Technically the vertical component of the bone loss can be fully corrected by regenerative surgical techniques. So far the horizontal bone loss should be considered as an irreversible feature as no periodontal regenerative techniques are available to restore the lost alveolar crest and interdental septa. PMID- 15282976 TI - [Oral manifestations in Crohn's disease and dental management]. AB - Crohn's disease is a systemic chronic inflammatory disorder, which may involve any part of the gastrointestinal tract, including the oral cavity. Bowel symptoms are the predominant manifestations, however during the course of the disease a lot of extra-intestinal complications may occur. The prevalence rates of oral manifestations varied between 5 to 20 per cent, but in pediatric patients the prevalence is much higher: 48 to 80 per cent. The review gives an overview of the oral findings observed in patients with Crohn's disease, and the potential implications of the disease for dental management are discussed. PMID- 15282977 TI - [An incidence rate study of different types of partial dentures made in Szeged in the year 2001]. AB - The prosthetic rehabilitation in cases of partial edentia is influenced by several objective and subjective factors. The goal of this study was to examine the correlations and differences among patients' sex, age, the type and material of the denture, type of anchorage and the color of the artificial teeth regarding the prepared dentures. The data relating to the prostheses and analyzed in this study were collected from the work-sheets of the technical laboratories and upon these statistical analyses were made according to the above given points of view. Researchers found that 65% of the dentures were made for women and the age-group of 50-70-year-old patients were most in need for prosthetic rehabilitation. Altogether the rate of fix prostheses was lower than the rate of removable partial dentures, which were prepared with metal framework and clasps. The authors concluded that resin-veneered bridges were overtaken by the metal-ceramic bridges and also the removable partial dentures with metal framework overshadowed the resin baseplate prostheses. PMID- 15282978 TI - [Microbiological testing of the artificial gingival margin in dentures]. AB - In everyday practice dental laboratories try to reproduce the natural form of sulcus gingivae at the transitional area between artificial teeth and gingiva of removable dentures, even on esthetically less important areas. Aim of these investigations were to examine how artificial recreation of the sulcus gingivae influences plaque retention, and what is the microbiological relevance of these. Investigations were carried out on the vestibular side of removable dentures of 32 randomly selected patients treated at the Department of Prosthodontics at the Faculty of Dentistry, Semmelweis University. Microbiological samples were taken from each patient using the same method. Samples were taken from the left upper first molars' artificial gingival margin using sterile paper points. Paper points were then transported in Eppendorf-tubes, in 2 ml of physiological saline solution, and processed within a two-hour period of time. Series dilutions were made of the sample solutions, then surface-streaked on Subaraud and Gentamycin, blood-agar, eosin-methylene blue and Mitis Salivarius culture enriched with Bacitracin. Subaraud culture was induced under aerob conditions, at room temperature for two days, then the total amount of fungi quantified. After pure culturing Candida albicans ID-culture was used for identification, and BioMerieux ATB automatic equipment to identify different Candida species. From pure cultures identification was carried out with Gram-staining, Neisser-staining, catalase, oxidase and also with other biochemical reactions. Blood-agar was used to determine total germ count, and normal commensal pharyngeal and oral bacteria. After collecting the microbiological samples, the conventional shape of the dental margin of gingiva was abolished on one side of the dentures and a smooth transition was created between denture teeth and the artificial gingiva in the molar and premolar region. During our investigations only blastomycetes were found. Besides most common Candida albicans, Candida glabrata, tropicallis, spherica and lambica were also identified. Our patients did not miss artificial sulcus gingivae, had no aesthetic complaints about smooth transition between artificial teeth and gingiva. Microbiological investigations of the samples and the comparative analysis showed, that on the smooth transitional areas compared to conventionally shaped sulci significantly less gram-negative, gram-positive bacteria and oral fungi were found, and there was no plaque formation. PMID- 15282979 TI - [Background data about the high dental fear scores of Hungarian 8-15-year-old primary school children]. AB - Free associations (coupling) of 139 Hungarian primary school children about their teeth was collected. Dental fear (DAS, DFS) and general anxiety scores were measured. Typical dental events (i.e.: loss of deciduous teeth, simple and traumatising dental treatments, tooth fractures) were coupled by the participants in 41.0% of the cases. Functions and importance of the teeth and oral hygiene were described in 20.1% of the cases. Simple, grotesque, or magical stories and tales about teeth were found in 8.6% of the cases. No answer was given in 30.2% of the cases. Highest dental fear and general anxiety scores were found in the group coupled traumatising dental treatment. Traumatising loss of deciduous teeth was caused by the dentist or by the father of the child, and was associated with higher dental fear and general anxiety comparing to simple loss of deciduous teeth. Higher dental fear and general anxiety scores were found in the group coupled functions and importance of the teeth comparing to the group coupled simple, grotesque, or magical stories and tales, or the group giving no answer. PMID- 15282980 TI - Treating psoriasis as a T-cell mediated disease. PMID- 15282981 TI - Perspective. Economists: big Medicare changes can't wait...but probably will. PMID- 15282982 TI - [Consumer drug advertising has negative effects on the patient-physician relationship]. PMID- 15282983 TI - [From surgeon assistant to independent specialist. The role of anesthesiologists and intensive care physicians in emergency medicine of the 20th century]. PMID- 15282984 TI - [APS I--a severe autoimmune disease with endocrine and non-endocrine symptoms]. AB - Autoimmune polyglandular syndrome type I (APS I) is an autosomal recessive disorder characterized by a combination of autoimmune manifestations affecting endocrine and non-endocrine organs. APS I usually presents in childhood. The three most common manifestations are chronic mucocutaneous candidiasis, hypoparathyroidism and Addison's disease. At least two of these must be present to fulfill the diagnostic criteria of this syndrome. The spectrum of other associated diseases includes gonadal insufficiency, alopecia, vitiligo and chronic active hepatitis. APS I is caused by a mutation in the AIRE-gene (autoimmune regulator) located on chromosome 21. Analysis of specific autoantibodies against intracellular enzymes, particularly enzymes in the synthesis of steroids and neurotransmittors, can be used in the diagnosis of APS I and to predict different manifestations of the disease. PMID- 15282985 TI - [Femoral nerve block as pain relief in hip fracture. A good alternative in perioperative treatment proved by a prospective study]. AB - Almost 25% of all patients with hip fracture experience temporary confusion pre- and directly postoperatively due to trauma, advanced age, transport between units, and the use of analgesics, 35-50% of the patients suffer temporary or chronic decubitus. Analgesics often lead to nausea. A femoral nerve block can interrupt sensory impulses from the hip joint and provide complete pain relief without affecting the CNS, thus making preoperative care easier and postoperative rehabilitation can be started earlier. 80 consecutive patients with hip fracture were randomized to femoral nerve block or pharmacological treatment only. Paracetamol and tramadol were the standard analgesics used. All patients were followed up with regard to pain, duration of the block, number of analgesics doses, temporary confusion and time for postoperative mobilization. Pain was estimated by the patients using the visual analogue scale (VAS). A nerve block was performed to block the femoral nerve, the lateral femoral cutaneous nerve and the obturator nerve with 30 ml of ropivacaine 7.5 mg/ml. Mental status was evaluated with Pfeiffer-test. All patients experienced relatively intense pain on admission with an average VAS of 6. After nerve block the VAS was 2. Pain relief was the same in the control group. Pain relief was sustained for 15 hours. The time for mobilization after surgery was significantly lower, 23 hours compared to 36 for the control group. There was a lower number of patients temporarily confused in the block group compared to the control group, however no significant differences were seen. Femoral nerve block provides adequate pain relief, equivalent to pharmacological treatment in most patients. The time for postoperative mobilization was shorter and less temporary confusion was seen. There were no complications in this group, making nerve block a good alternative to traditional pharmacological preoperative treatment for patients with hip fractures. PMID- 15282986 TI - [Emotional reactions common sequelae of stroke]. AB - Emotional reactions are important sequelae of stroke. Mood disorders, such as depression, anxiety, post-traumatic stress syndrome and emotionalism, occur during the first post-stroke year, each of them in approximately 20-30% of patients. They often overlap, and prevalence estimates differ on account of differences in definitions; study populations; exclusion criteria and time of assessment. The risk seems to be greatest, at least for depression, in the first months after stroke. Some patients recover spontaneously but symptoms persist in up to one third. Pharmacological treatment can have a positive effect that also applies to rehabilitation, quality of life and cardiovascular mortality. However, study findings are not uniform and conclusive therapeutic and preventive intervention trials on mood disorders after stroke are urgently needed. PMID- 15282987 TI - [Better scientific basis is necessary for traffic safety]. PMID- 15282988 TI - [Health insurance money can be used to increase medical research support]. PMID- 15282989 TI - [Adaptation of working hours to EU directives does not require more physicians!]. PMID- 15282990 TI - [Biological control--a better way of controlling infections]. PMID- 15282991 TI - [The sick Sweden--a challenge to all!]. PMID- 15282992 TI - Two epidemics: are we getting fatter as we sleep less? PMID- 15282993 TI - A novel combination therapy for primary insomnia? PMID- 15282994 TI - Rapid maxillary expansion in obstructive sleep apnea--hope on the horizon? PMID- 15282995 TI - REM sleep deprivation induces changes in coping responses that are not reversed by amphetamine. AB - STUDY OBJECTIVE: Although sleep deprivation often occurs during stressful or threatening situations, the effects of sleep loss on defensive and coping behaviors are not well known. The purpose of the present study was to measure the effects of selective rapid eye movement (REM) sleep deprivation (RSD) on responses elicited by threatening situations and to assess the extent to which RSD-induced changes are reversed by amphetamine. DESIGN: Animals were divided into 3 groups; home-cage control, apparatus control, and REM sleep-deprived groups. The flowerpot method was used to produce RSD for up to 5 days. One set of rats was tested in the elevated plus maze, open field, shock-induced freezing, and analgesia tests. A second set of rats was evaluated for locomotor activity. A third set of animals was assessed in the defensive burying test. For the amphetamine studies, groups of home-cage control and REM sleep-deprived rats received an intraperitoneal injection of amphetamine prior to administration of the shock-induced freezing test or the defensive burying test. SETTING: Sleep Research Laboratory at UW-Madison. PARTICIPANTS AND INTERVENTIONS: 186 male Long Evans rats approximately 3 months old. MEASUREMENTS AND RESULTS: RSD increased the proportion of time spent in open arms of the elevated plus maze and center of the open field, decreased freezing time, and reduced defensive burying. Amphetamine did not reverse RSD-induced changes in freezing or burying responses. CONCLUSIONS: RSD causes widespread abnormalities in coping and defensive responses in threatening situations; these deficits are not reversed and, in some cases, may be exacerbated by amphetamine. PMID- 15282996 TI - Lesions of the suprachiasmatic nucleus eliminate the daily rhythm of hypocretin-1 release. AB - STUDY OBJECTIVES: Hypocretins (orexins) are involved in the sleep disorder narcolepsy. While hypocretin-1 has a daily oscillation, little is known regarding the relative contribution of circadian and homeostatic components on hypocretin release. The effect of lesions of the suprachiasmatic nucleus (SCN) on hypocretin 1 in the cerebrospinal fluid (CSF) was examined. DESIGN: SCN-ablated (SCNx) and sham-operated control rats were implanted with activity-temperature transmitters. Animals were housed individually under 1 of 3 lighting conditions: 12-hour:12 hour light:dark cycle (LD), constant light (LL), and constant darkness (DD). Lesions were verified histologically and shown not to affect hypocretin containing cells. Hypocretin-1 concentrations in the CSF were determined every 4 hours using radioimmunoassays. MEASUREMENTS AND RESULTS: Control animals displayed robust circadian (LL, DD) and diurnal (LD) fluctuations in CSF hypocretin-1, locomotor activity, and temperature. Peak CSF hypocretin-1 was at the end of the active period. Activity, temperature, and CSF hypocretin-1 were arrhythmic in SCNx animals in LL and DD. In LD, a weak but significant fluctuation in activity and temperature but not CSF hypocretin-1 was observed in SCNx animals. We also explored correlations between CSF hypocretin-1, CSF corticosterone, and locomotor activity occurring prior to CSF sampling in arrhythmic SCNx rats under constant conditions. Significant correlations between hypocretin-1 and activity were observed both across and within animals, suggesting that interindividual and time-of-the-day differences in activity have significant effects on hypocretin release in arrhythmic animals. No correlation was found between CSF hypocretin-1 and corticosterone. CONCLUSIONS: Hypocretin-1 release is under SCN control. Locomotor activity influences the activity of the hypocretin neurons. PMID- 15282997 TI - Sleep homeostasis in Drosophila melanogaster. AB - STUDY OBJECTIVES: The fruit fly Drosophila melanogaster is emerging as a promising model system for the genetic dissection of sleep. As in mammals, sleep in the fruit fly is a reversible state of reduced responsiveness to the external world and has been defined using an array of behavioral, pharmacologic, molecular, and electrophysiologic criteria. A central feature of mammalian sleep is its homeostatic regulation by the amount of previous wakefulness. Dissecting the mechanisms of homeostatic regulation is likely to provide key insights into the functions of sleep. Thus, it is important to establish to what extent sleep homeostasis is similar between mammals and flies. This study was designed to determine whether in flies, as in mammals, (1) sleep rebound is dependent on prior time awake; (2) sleep deprivation affects the intensity, in addition to the duration, of sleep rebound; (3) sleep loss impairs vigilance and performance; (4) the sleep homeostatic response is conserved among different wild-type lines, and between female and male flies of the same line. DESIGN: Motor activity of individual flies was recorded at 1-minute intervals using the infrared Drosophila Activity Monitoring System during 2 baseline days; during 6, 12, and 24 hours of sleep deprivation; and during 2 days of recovery. Sleep was defined as any period of uninterrupted behavioral immobility lasting > 5 minutes. Sleep continuity was measured by calculating the number of brief awakenings, the number and duration of sleep episodes, and a sleep continuity score. Vigilance before and after sleep deprivation was assessed by measuring the escape response triggered by 2 different aversive stimuli. SETTING: Fly sleep research laboratory at UW-Madison. PARTICIPANTS AND INTERVENTIONS: Adult flies of the Canton-S (CS) strain and 116 other wild-type lines (> or = 16 female and > or = 16 male flies per line). MEASUREMENTS AND RESULTS: In wild-type CS flies, as in mammals, the amount of sleep recovered after sleep deprivation was dependent on prior time awake. Relative to baseline sleep, recovery sleep in CS flies was less fragmented, with longer sleep episodes, and was associated with a higher arousal threshold. Sleep deprivation in CS flies also reduced performance. Sleep duration and continuity increased after 24 hour of sleep deprivation in all the other wild-type lines tested. CONCLUSION: The sleep homeostatic response in fruit flies is a stable phenotype and shares most of, if not all, the major features of mammalian sleep homeostasis, thus supporting the use of Drosophila as a model system for the genetic dissection of sleep mechanisms and functions. PMID- 15282998 TI - Sleep as a tool for evaluating autonomic drive to the heart in cardiac transplant patients. AB - STUDY OBJECTIVES: The aim of this study was to investigate the autonomic drive to the heart in cardiac transplant patients (CTP) using heart rate (HR) and HR variability (HRV) analysis during non-rapid eye movement (NREM)-rapid eye movement (REM) sleep cycles, in particular during arousal associated with the emergence from slow wave sleep (SWS). In healthy subjects, this arousal is characterized by a pronounced HR surge, and HRV is lower during SWS than during the subsequent "active" sleep stage 2 and REM sleep. PARTICIPANTS: The participants were 24 adults, 14 CTP (men, n = 11; women, n = 3; mean age, 62.2 +/ 2.2 years; time after transplantation, 4-14 years) and 10 control subjects (men, n = 7; women, n = 3; mean age, 61.0 +/- 1.8 years). DESIGN: The subjects underwent polygraphic sleep, cardiac, and respiratory recordings during an experimental night. HR was measured during the arousal. HRV was estimated from the R-R intervals in 5-minute stationary segments preceding and following arousal, ie, during SWS and active sleep stage 2 from the first 2 complete NREM REM sleep cycles. RESULTS: In controls, HR increased during arousal associated with the emergence from SWS during the 2 sleep cycles (P < .05). Sleep-stage dependent increases of all HRV indexes were observed in the 2 sleep cycles. Concerning CTP, 5 of them displayed a smaller HR increase at arousal, whereas 9 other patients had no HR variation. This distinction between the 2 groups of CTP was confirmed by HRV analysis. The patients with HR reactivity to arousal presented significant sleep-stage-dependent increases in global HRV and sympathetic HRV indexes, whereas the nonreactive group was characterized by an inability of HRV to change with sleep-stage alternation. Sympathetic HRV indexes were significantly higher in the reactive patients than in nonreactive patients, but high frequency power reflecting parasympathetic activity did not differ. However, the absolute HRV indexes were greatly decreased in both groups of patients compared to controls. CONCLUSION: HR reactivity during arousal associated with the emergence from SWS, corroborated by HRV surrounding arousal, may suggest a partial improvement of the sympathetic drive to the heart in some CTP, with no indication of increased parasympathetic activity. Other signs of reinnervation have to be identified to validate this hypothesis. PMID- 15282999 TI - Scatterplot analysis of EEG slow-wave magnitude and heart rate variability: an integrative exploration of cerebral cortical and autonomic functions. AB - STUDY OBJECTIVES: To explore interactions between cerebral cortical and autonomic functions in different sleep-wake states. DESIGN: Active waking (AW), quiet sleep (QS), and paradoxical sleep (PS) of adult male Wistar-Kyoto rats (WKY) on their daytime sleep were compared. PARTICIPANTS: Ten WKY. INTERVENTIONS: All rats had electrodes implanted for polygraphic recordings. One week later, a 6-hour daytime sleep-wakefulness recording session was performed. MEASUREMENTS AND RESULTS: A scatterplot analysis of electroencephalogram (EEG) slow-wave magnitude (0.5-4 Hz) and heart rate variability (HRV) was applied in each rat. The EEG slow-wave-RR interval scatterplot from all of the recordings revealed a propeller-like pattern. If the scatterplot was divided into AW, PS, and QS according to the corresponding EEG mean power frequency and nuchal electromyogram, the EEG slow wave-RR interval relationship became nil, negative, and positive for AW, PS, and QS, respectively. A significant negative relationship was found for EEG slow-wave and high-frequency power of HRV (HF) coupling during PS and for EEG slow wave and low-frequency power of HRV to HF ratio (LF/HF) coupling during QS. The optimal time lags for the slow wave-LF/HF relationship were different between PS and QS. CONCLUSIONS: Bradycardia noted in QS and PS was related to sympathetic suppression and vagal excitation, respectively. The EEG slow wave-HRV scatterplot may provide unique insights into studies of sleep, and such a relationship may delineate the sleep-state-dependent fluctuations in autonomic nervous system activity. PMID- 15283000 TI - The association between short sleep duration and obesity in young adults: a 13 year prospective study. AB - STUDY OBJECTIVES: Obesity has become a major health problem with increasing prevalence. Given the limited availability of effective treatment of weight problems, the identification of potentially modifiable risk factors may lead to preventive approaches to obesity. The objective of this study was to test the hypothesis that short sleep duration is associated with obesity and weight gain during young adulthood. DESIGN: Prospective single-age cohort study of young adults. Information was derived from 4 interviews when participants were ages 27, 29, 34, and 40 years. SETTING: Community setting. PARTICIPANTS: 496 young adults. MEASUREMENTS AND RESULTS: Trained health professionals administered a semistructured interview for psychiatric and medical conditions and health habits. This study showed an association between short sleep duration and obesity (at age 27 years, odds ratio: 7.4, 95% confidence interval: 1.3-43.1) and a negative association between sleep duration and body mass index in young adults. These associations persisted after controlling for a variety of potentially confounding variables, including family history of weight problems, levels of physical activity, and demographic variables. Associations between sleep duration and obesity diminished after age 34 years. There was a trend (P = .08) for average change rate of weight gain to be negatively associated with average change rate of sleep duration. CONCLUSIONS: Because sleep duration is a potentially modifiable risk factor, these findings might have important clinical implications for the prevention and treatment of obesity. PMID- 15283001 TI - Circadian variation in neuroendocrine response to L-dopa in patients with restless legs syndrome. AB - STUDY OBJECTIVE: To investigate circadian changes in dopaminergic function by means of a neuroendocrine challenge (growth hormone and prolactin responses to an acute oral administration of L-dopa) in patients with idiopathic restless legs syndrome (RLS) and controls. DESIGN: Randomized administration of the L-dopa neuroendocrine challenge. SETTING: Sleep disorders laboratory at a 500-bed academic hospital. PATIENTS OR PARTICIPANTS: Twelve patients diagnosed with idiopathic RLS and 12 age- and sex-matched healthy controls. INTERVENTIONS: Following a comprehensive evaluation that included nocturnal polysomnographic study, all participants underwent the L-dopa neuroendocrine challenge on 2 occasions (11 am and 11 pm). Subjects were previously randomly assigned to the time of first challenge (11 am or 11 pm). On each occasion, subjects took 200 mg of L-dopa (plus 50 mg carbidopa) by mouth. Blood was drawn 20 minutes and 5 minutes before administration of the drug, as well as 15, 30, 45, 60, 75, 90, 102, and 120 minutes after administration. RESULTS: Prechallenge levels of plasma values of growth hormone or prolactin did not differ in the 2 subject groups. Following only the nighttime administration of L-dopa, RLS patients manifested a more pronounced inhibition of prolactin release and an increase in growth hormone secretion. Prolactin plasma levels were significantly correlated to the periodic limb movement index on the polysomnogram. CONCLUSIONS: These findings may reflect enhanced circadian variations in dopaminergic function and support an increased sensitivity at night of dopamine receptors in patients with RLS. PMID- 15283002 TI - Long-term safety and efficacy of cabergoline for the treatment of idiopathic restless legs syndrome: results from an open-label 6-month clinical trial. AB - STUDY OBJECTIVES: To assess the safety and efficacy of cabergoline in the treatment of idiopathic restless legs syndrome (RLS) patients. DESIGN: Open-label intervention study for 26 weeks; no control group. SETTING: 302 patients (73% women, aged 61 +/- 11 years) from 37 German neurologic outpatient departments and private practices. INTERVENTION: Cabergoline was upwardly titrated over 4 weeks to individually optimized dosages. Median treatment duration was 181 days. The median daily dose of cabergoline was 1.5 mg. MEASUREMENTS: Drug safety was assessed by adverse events; efficacy was evaluated with the RLS-6 and the International RLS Rating Scales. RESULTS: In 48% of the study participants, investigators reported adverse events suspected to be drug related. Most adverse events were mild and transient and related to the gastrointestinal system (nausea: 16.6%) or the central nervous system (dizziness: 7.0%, headache: 4.6%). Premature dropout from the study occurred in 54 patients (17.9%), in 17 patients (3.0%) due to a drug-related adverse event. The severity of RLS symptoms at night, at bedtime, and during the day, as well as the International RLS Rating Scale total score improved during therapy. Satisfaction with sleep was increased (all P values < .001). In 5% of all patients, RLS symptoms worsened, and in a further 6.3%, response to therapy was poor. In 9 patients (3.0%) between 1 and 3 criteria for augmentation were noted. CONCLUSIONS: Long-term therapy with cabergoline is a safe and well-tolerated treatment option for the great majority of patients with idiopathic RLS. The treatment was efficacious both for nighttime and daytime symptoms in this indication and may carry a low risk of augmentation. PMID- 15283003 TI - Executive function in sleep apnea: controlling for attentional capacity in assessing executive attention. AB - STUDY OBJECTIVES: As the effects of general slowness and decreased attentional capacity on higher executive attention have not been fully taken into account in the sleep apnea literature, we statistically controlled for basic attentional performance in evaluating executive attention per se in sleep apnea patients. DESIGN: A case-controlled design was used with comparison of basic and executive attentional tasks. PARTICIPANTS: Thirty-six polysomnographically diagnosed patients (mean apnea-hypopnea index = 60.5 +/- SD 31.6) participated, together with 32 healthy controls. MEASUREMENTS AND RESULTS: Neuropsychological tests included Trail Making part A and B, Symbol Digit Modalities (SDMT), Digit Span forward and backward, Stroop Color-Word, Five-Point design fluency, and an Attentional Flexibility task. Patients' vigilance data indicated time-on-task decrements after 10 minutes. Moreover, their performance was significantly reduced on the SDMT (effect size d = 0.93), the Digit Span forward task (d = 0.44), the number of errors on the basic 2-choice reaction time subtest of the Attentional Flexibility task (d = 0.74) and the mean RT on the actual Attentional Flexibility subtest (d = 0.54). It has been argued that the latter poor performance was probably primarily related to the task's phonologic loop component of working memory rather than to an attentional switching deficit per se. No other performance differences were found between patients and healthy controls. CONCLUSIONS: In addition to vigilance decrements, attentional capacity deficits clearly emerge, ie, slowed information processing and decreased short term memory span. However, no specific clinical indications for executive attentional deficits--such as disinhibition, distractibility, perseveration, attentional switching dysfunction, decreased design fluency, or an impaired central executive of working memory--are found in patients with severe sleep apnea. Their cognitive performance seems very similar to the cognitive decline found after sleep loss and qualitatively different from patients with chronic obstructive pulmonary disease, suggesting sleepiness as the primary factor in a parsimonious explanation for the attention deficits in sleep apnea, without the need to assume prefrontal brain damage. PMID- 15283004 TI - A clinical decision rule to prioritize polysomnography in patients with suspected sleep apnea. AB - STUDY OBJECTIVES: To derive and validate a clinical decision rule that can help to prioritize patients who are on waiting lists for polysomnography, DESIGN: Prospective data collection on consecutive patients referred to a sleep center. SETTING: The Newcastle Sleep Disorders Centre, University of Newcastle, NSW, Australia. PATIENTS: Consecutive adult patients who had been scheduled for initial diagnostic polysomnography. MEASUREMENTS AND RESULTS: Eight hundred and thirty-seven patients were used for derivation of the decision rule. An apnea hypopnoea index of at least 5 was used as the cutoff point to diagnose sleep apnea. Fifteen clinical features were included in the analyses using logistic regression to construct a model from the derivation data set. Only 5 variables- age, sex, body mass index, snoring, and stopping breathing during sleep--were significantly associated with sleep apnea. A scoring scheme based on regression coefficients was developed, and the total score was trichotomized into low-, moderate-, and high-risk groups with prevalence of sleep apnea of 8%, 51%, and 82%, respectively. Color-coded tables were developed for ease of use. The clinical decision rule was validated on a separate set of 243 patients. Receiver operating characteristic analysis confirmed that the decision rule performed well, with the area under the curve being similar for both the derivation and validation sets: 0.81 and 0.79, P =.612. CONCLUSION: We conclude that this decision rule was able to accurately classify the risk of sleep apnea and will be useful for prioritizing patients with suspected sleep apnea who are on waiting lists for polysomnography. PMID- 15283005 TI - The prevalence of parasomnias in preadolescent school-aged children: a Turkish sample. AB - STUDY OBJECTIVES: To survey the prevalence of parasomnias in a population of children aged 7 to 11 years and to determine whether parasomnias are associated with medical and neurobehavioral properties. DESIGN: Parents and children completed a pediatric sleep questionnaire that contains 27 items developed by the authors to assess parasomnias in children. Parents and children were also interviewed about the children's medical and sociofamilial history, schooling, psychological difficulties, medication intake, and the history of psychomotor and psychosocial development. SETTING: NA PARTICIPANTS: 971 preadolescent school-aged children from 4 locations in Turkey participated in the study. RESULTS: We found a 14.4% prevalence of parasomnia in preadolescent school-aged children. Almost every sixth child had about at least 1 parasomnia. When we examined parasomnias separately, bruxism, nocturnal enuresis, and night terrors were the most common parasomnias among both girls and boys. The prevalence of parasomnias was higher in the 9- and 10-year-old age groups than in the other age groups. Girls and boys did not differ. Children with parasomnias had higher rates of past physical illness, delays in toilet raining, behavior disturbances, adjustment problems, and learning difficulties. CONCLUSIONS: These results suggest that the prevalence of parasomnias was high in the 9- and 10-year-old age groups. Parasomnias are associated with a history of physical illness and neurobehavioral abnormalities. PMID- 15283006 TI - Hypersynchronous delta sleep EEG activity and sudden arousals from slow-wave sleep in adults without a history of parasomnias: clinical and forensic implications. AB - STUDY OBJECTIVES: To determine the frequency of classical markers of non-rapid eye movement (NREM) parasomnias--hypersynchronous delta sleep (HSD) electroencephalogram waves and sudden arousals from slow-wave sleep (SWS)--in patients without histories of somnambulism or other NREM parasomnias. DESIGN: Retrospective review. SETTING: Sleep disorders center laboratory. PATIENTS: 82 consecutive patients without a history of parasomnias who underwent diagnostic polysomnograms; 57 men and 25 women, mean age 48+/-13.3 years, were included without regard to diagnosis or findings. All patients had at least 30 seconds of stage 3 or 4 sleep during the polysomnogram. MEASUREMENTS AND RESULTS: The primary diagnosis of all but 4 patients was obstructive sleep apnea (mean respiratory disturbance index, 30 +/- 23.6 [range, 2.7-117] per hour of sleep). Polysomnograms were then reviewed for the presence of HSD and SWS arousals. A total of 235 arousals (mean, 2.9 +/- 2.7; range, 0-14) from stage 3 or 4 sleep were noted. Eight-five percent of all patients had at least 1 SWS arousal and 45% had 3 or more SWS arousals; 85.1% of all arousals from SWS were secondary to sleep-disordered breathing, and 5.9% were secondary to leg movements. At least 1 episode of HSD (mean, 1.4 +/- 1.6; range, 0-9) was noted in 65.8% of patients. CONCLUSIONS: HSD and SWS arousals were a common finding in patients without clinical histories of sleepwalking or other parasomnias but who were found to have frequent respiratory-related arousals during sleep. HSD and SWS arousals thus have a low specificity for NREM parasomnias and, without further research, are not useful for the objective confirmation of parasomnias in clinical evaluations and in the forensic evaluation of sleepwalking as a legal defense. PMID- 15283007 TI - The effects of modafinil and cognitive behavior therapy on sleep continuity in patients with primary insomnia. AB - BACKGROUND: Daytime fatigue, if not frank sleepiness, is a common symptom among patients with insomnia, one that is exacerbated during acute treatment with cognitive behavior therapy (CBT). The present study was undertaken to assess whether modafinil could be used to reduce daytime fatigue, sleepiness, or both in patients with primary insomnia and whether the pharmacologic augmentation of wakefulness might produce improved sleep by itself or in combination with CBT. METHODS: 30 subjects with primary insomnia were enrolled in this study and were randomly assigned to 1 of 3 treatment conditions: (1) placebo plus CBT, (2) 100 mg modafinil plus CBT, or (3) 100 mg modafinil plus a contact control (monitor only condition). Subjects were continuously monitored with sleep diaries from study intake until study end (10 weeks) and were evaluated on a weekly basis for changes in sleepiness. RESULTS: The mean age of the group was 41.3 years (SD, 13.4), and 70.4% of subjects were women. All 3 groups exhibited mean sleep latency and wake after sleep-onset times that were more than 30 minutes in duration. The mean pretreatment sleep profiles did not significantly differ. Modafinil, when administered alone, did not significantly affect the patients' sleep profiles. A trend, however, was evident for improved sleep latency. Modafinil, as an adjunct to CBT, tended to (1) reduce daytime sleepiness as measured by the Epworth Sleepiness Scale and (2) enhance compliance with CBT. With respect to the latter, subjects in the modafinil plus CBT group more reliably adhered to the prescribed phase delay in bedtime than did the placebo plus CBT group. DISCUSSION: These data suggest that modafinil may be used to diminish the negative side effects of CBT (increased daytime sleepiness) and may increase subject compliance with therapy. Whether enhanced daytime function mediates the change in adherence and whether reduced sleepiness and enhanced compliance translate to less patient attrition in the clinical setting remain to be evaluated. PMID- 15283008 TI - Prevalence of reported insomnia and its consequences in a survey of 5,044 adolescents in Kuwait. AB - STUDY OBJECTIVES: To examine the psychometric characteristics of a short scale to assess insomnia complaints and their consequences, to estimate the prevalence rates of reported insomnia, and to explore the sex differences in reported insomnia. METHODS: 5,044 male and female nonclinical secondary-school students in the State of Kuwait participated. Their ages ranged from 14 to 19 years. An insomnia scale (IS) comprising 12 items was administered in group sessions. It has acceptable test-retest and alpha reliabilities and good convergent validity. Two factors were disclosed: consequence of insomnia and difficulty in initiating and maintaining sleep. Point prevalence rate was computed as the summation of the percentages of responses in the two options: "Much" and "Very much" on each item during the most recent month. RESULTS: The prevalence of the 12 IS items ranged from 6.4% to 31.7% in boys, and between 6.5% and 35.9% among girls. The highest reported insomnia complaint was early morning awakening, ie, 31.7% in boys, and 35.9% in girls. It was found that 14.6% of boys and 20.3% of girls reported difficulty initiating sleep, while 8.6% of boys and 15.7% of girls reported difficulty maintaining sleep. Girls had higher mean scores in most of the IS items. CONCLUSIONS: Adolescents in the present sample have reported high rates of insomnia. There is a great need to agree upon the methodology, especially the assessment tool, for epidemiology of sleep disorder research. PMID- 15283009 TI - Association of daytime sleepiness with COMT polymorphism in patients with parkinson disease: a pilot study. AB - STUDY OBJECTIVES: To evaluate an association between catechol-O-methyltransferase (COMT) genotype and subjective daytime sleepiness in patients with Parkinson disease. DESIGN: Structured questionnaire study. SETTING: Tertiary Parkinson disease care center and sleep outpatients' department at the university hospital neurology department. PARTICIPANTS: All nondemented patients with idiopathic Parkinson disease who had been part of a previous study of D4-receptor polymorphisms in 1997 were eligible to participate. From the original sample of 113 patients, 46 participated in the study, 22 met exclusion criteria, and 43 were not available. INTERVENTIONS: Not applicable. MEASUREMENTS AND RESULTS: In this study, 46 patients were included (27 men, 19 women; 68.4 +/- 9.9 years of age; symptomatic disease duration, 12.2 +/- 5.2 years; Hoehn and Yahr stage in "on" of 2.6 +/- 0.8). Out of the 46 patients, 13 had LL genotype, 22 LH, and 11 HH. The Epworth Sleepiness Scale scores were 9.5 +/- 4.8 in LL, 8.5 +/- 4.7 in LH, and 6.8 +/- 3.1 in HH (mean +/- SD) (NS). LL and LH were grouped together. The Epworth Sleepiness Scale score was 11 or more in 40% of the LL+LH group, compared to 9.1% of the HH group (P = .039). The levodopa or dopamine-agonist doses and types did not differ between the LL+LH group versus the HH group. CONCLUSIONS: These preliminary data suggest an association of the Lallele and daytime sleepiness in patients with Parkinson disease. PMID- 15283010 TI - Decreased sleep spindles and spindle activity in midlife women with fibromyalgia and pain. AB - OBJECTIVES: To compare sleep-spindle incidence (number of spindles per minute of non-rapid eye movement [NREM] stage 2 sleep) and duration, spindle wave time (seconds per epoch in NREM stage 2 sleep), spindle frequency activity, and pain measures (pressure pain threshold, number of tender points, skinfold tenderness) between midlife women with fibromyalgia (FM) and moderate to high pain to a control group of sedentary women without pain. A second goal was to explore the extent to which pain pressure thresholds, age, and depression explain the variance in spindle incidence. DESIGN: A cross-sectional descriptive study. SETTING: A university-based sleep research laboratory and a referral clinic for chronic fatigue and pain. PARTICIPANTS: Thirty-seven medication-free women with FM (mean age, 44.9 +/- 8 years) and 30 women with self-reported good sleep and no pain (mean age, 44.1 +/- 7.7 years) completed a psychiatric interview and the Beck Depression Inventory prior to 2 consecutive nights of polysomnography, with pain measures obtained in the morning. Time domain analysis of spindle incidence and spectral analysis of spindle frequency activity were conducted on night 2 of polysomnography recordings. INTERVENTIONS: NA. RESULTS: Women with FM had fewer mean spindles per minute of NREM stage 2 sleep and lower mean spindle time per epoch of NREM stage 2 sleep (both P values < .02), but mean spindle duration, although slightly shorter, was not statistically significantly different (P < .06) compared to control women. Women with FM had a lower mean pressure pain threshold, a higher average number of positive tender points, and higher skinfold tenderness compared to control women (all P values < .001). Group differences in spindle frequency activity were found after controlling for age, depression, and psychiatric diagnosis in a general linear model (P < .02). One-way analysis of variance revealed significantly lower spindle activity in the 3 frequency bins (12-12.5 Hz, 13-13.5 Hz, 14-14.5 Hz) at C3 (all P values < .04), Fz (all P values < .02), and Cz (all P values < .02). Finally, after controlling for age and depression, pain pressure threshold significantly predicted spindles per minute and spindle time per epoch of NREM stage 2 sleep (r2 = .26; P < .001). CONCLUSIONS: Women with FM and pain have fewer sleep spindles and reduced electroencephalogram power in spindle frequency activity compared to control women of similar age. These data imply that some aspect of thalamocortical mechanisms of spindle generation might be impaired in FM. PMID- 15283011 TI - Symptoms of insomnia among adolescents in the lower Rio Grande Valley of Texas. AB - STUDY OBJECTIVES: To estimate the prevalence of symptoms of insomnia among adolescents living along the United States-Mexico border and to examine whether ethnicity and birthplace affect risk for such symptoms. DESIGN: Cross-sectional school-based survey using a version of the 2001 Youth Risk Behavior Survey, modified to elicit data on sleep problems. SETTING: A probability sample of 13 high schools selected from more than 40 high schools in the 4 southernmost counties in Texas-the Lower Rio Grande Valley-contiguous with Mexico. PARTICIPANTS: All ninth-grade students who agreed to participate (n = 5,118). MEASUREMENTS: The sleep module consisted of queries about trouble initiating asleep, trouble maintaining sleep, early morning waking, nonrestorative sleep, quality of sleep, and amount of sleep. RESULTS: Symptoms of insomnia were common, with 12.4% of respondents meeting symptom criteria for insomnia almost every day of the past month. Females were more likely to report insomnia, as were youths reporting lower socioeconomic status. Crude odds ratios suggested foreign-born and those who identified themselves as "Mexican" rather than "Mexican American" were at lower risk of insomnia. However, multivariate analyses eliminated these differences. CONCLUSIONS: More comparative research is needed to ascertain whether and how ethnic culture affects risk for disordered sleep. PMID- 15283012 TI - Rapid maxillary expansion in children with obstructive sleep apnea syndrome. AB - OBJECTIVE: To evaluate the effect of rapid maxillary expansion on children with nasal breathing and obstructive sleep apnea syndrome. METHOD: Recruitment of children with maxillary contraction, without of adenoid hypertrophy, with a body mass index < 24 kg/m2, with obstructive sleep apnea syndrome demonstrated by polysomnography, and whose parents signed informed consent. Otolaryngologic and orthognathic-odontologic evaluation with clinical evaluation, anterior rhinometry and nasal fibroscopy, panoramic radiographs, anteroposterior and laterolateral telecephalometry were performed at entry and follow-up. INTERVENTION: Rapid maxillary expansion (ie, active phase of treatment) was performed for 10 to 20 days; maintenance of device (for consolidation) and orthodontic treatment on teeth lasted 6 to 12 months. RESULTS: 31 children (19 boys), mean age 8.7 years, participated in the study. The mean apnea-hypopnea index was 12.2 events per hour. At the 4-month follow-up, the anterior rhinometry was normal, and all children had an apnea-hypopnea index < 1 event per hour. The mean cross-sectional expansion of the maxilla was 4.32 +/- 0.7 mm. There was a mean increase of the pyriform opening of 1.3 +/- 0.3 mm. CONCLUSION: Rapid maxillary expansion may be a useful approach in dealing with abnormal breathing during sleep. PMID- 15283013 TI - Some children with growing pains may actually have restless legs syndrome. AB - STUDY OBJECTIVES: Growing pains may be an important clue to the diagnosis of childhood restless legs syndrome (RLS). However, there are no previous studies to determine whether a subpopulation of children with growing pains meet the diagnostic criteria for RLS. The purpose of this study is to determine if some children with growing pains meet diagnostic criteria for RLS and to compare the polysomnographic characteristics of these children to controls. DESIGN/PARTICIPANTS/MEASUREMENTS: Eleven children from a pediatric neurology clinic with an emphasis on attention-deficit/hyperactivity disorder (ADHD) and with a diagnosis of growing pains were referred. They were interviewed with the parent to determine if their symptoms of growing pains met criteria for definite RLS. Those who met clinical criteria for RLS underwent polysomnography, and the results of their polysomnographic studies were compared to those of a control group (10 children, mean age 9.7 years). SETTING: Academic medical center. RESULTS: Ten (mean age 10.4 years) of the 11 children with growing pains met clinical criteria for RLS. In 4 of 8 families of these 10 children, 1 parent had RLS. Six of the 10 children had ADHD. There were no differences in the polysomnographic findings between the growing-pain and control groups, and none of the children with RLS had what is considered to be a clinically significant number of periodic limb movements of sleep. There were no differences in the polysomnographic findings between the "growing-pain ADHD" and "growing-pain non ADHD" subgroups. The growing pains were severe enough for the patients and family to ask for treatment in 4 cases, and carbidopa/levodopa was utilized. CONCLUSIONS: Some children diagnosed with growing pains meet diagnostic criteria for RLS, and a family history of RLS is common in these children. In some cases symptoms are severe enough to warrant treatment. PMID- 15283014 TI - Spectral analysis of the sleep electroencephalogram during adolescence. AB - OBJECTIVES: To describe developmental changes of the human sleep electroencephalogram (EEG) during adolescence using EEG spectral analysis and specifically to compare the nocturnal dynamics of slow-wave activity (EEG spectral power 0.6-4.6 Hz, a marker for sleep homeostatic pressure) in prepubertal and mature adolescents. DESIGN: After 10 nights on a fixed 10-hour sleep schedule without daytime naps, participants were studied during a 10-hour baseline night. SETTING: Data were collected in a 4-bed sleep research laboratory. PARTICIPANTS: Eight prepubertal children (pubertal stage Tanner 1; mean age 11.3 years, SD +/- 1.2, 4 boys) and 8 mature adolescents (Tanner 5; mean age 14.1 years, +/- 1.3, 3 boys). INTERVENTIONS: Not applicable. MEASUREMENTS: All-night polysomnography was performed. Sleep stages were scored according to conventional criteria. EEG power spectra (of derivation C3/A2) were calculated using a fast Fourier transform routine. RESULTS: A reduction of non-rapid eye movement (NREM) sleep stage 4 (by 40.1%) and greater amounts of stage 2 sleep (19.7%) were found in mature compared to prepubertal adolescents. NREM sleep EEG power was lower in the frequency ranges < 7 Hz, 11.8 to 12.6 Hz, and 16.2 to 16.8 Hz in mature adolescents. A reduction of rapid eye movement sleep spectral power was present in the frequency ranges < 8.6 Hz and 9.6 to 15 Hz for mature compared to prepubertal adolescents. Slow-wave activity showed identical dynamics within individual NREM sleep episodes and across the night in both developmental groups. CONCLUSIONS: The homeostatic recuperative drive during sleep remains unchanged across puberty. The decline of slow-wave sleep during adolescence may reflect developmental changes of the brain rather than changes of sleep regulatory processes. PMID- 15283015 TI - Detection of obstructive sleep apnea in pediatric subjects using surface lead electrocardiogram features. AB - STUDY OBJECTIVES: To investigate the feasibility of detecting obstructive sleep apnea (OSA) in children using an automated classification system based on analysis of overnight electrocardiogram (ECG) recordings. DESIGN: Retrospective observational study. SETTING: A pediatric sleep clinic. PARTICIPANTS: Fifty children underwent full overnight polysomnography. INTERVENTION: N/A. MEASUREMENTS AND RESULTS: Expert polysomnography scoring was performed. The datasets were divided into a training set of 25 subjects (11 normal, 14 with OSA) and a withheld test set of 25 subjects (11 normal, 14 with OSA). Features, calculated from the ECG of the 25 training datasets, were empirically chosen to train a modified quadratic discriminant analysis classification system. The selected configuration used a segment length of 60 seconds and processed mean, SD, power spectral density, and serial correlation measures to classify segments as apneic or normal. By combining per-segment classifications and using receiver operator characteristic analysis, a per-subject classifier was obtained that had a sensitivity of 85.7%, specificity of 90.9%, and accuracy of 88% on the training datasets. The same decision threshold was applied to the withheld datasets and yielded a sensitivity of 85.7%, specificity of 81.8%, and accuracy of 84%. The positive and negative predictive values were 85.7% and 81.8%, respectively, on the test dataset. CONCLUSIONS: The ability to correctly identify 12 out of 14 cases of OSA (with the 2 false negatives arising from subjects with an apnea hypopnea index less than 10) indicates that the automated apnea classification system outlined may have clinical utility in pediatric patients. PMID- 15283016 TI - Older people awaken more frequently but fall back asleep at the same rate as younger people. AB - STUDY OBJECTIVES: To determine whether the increased wake within a bedrest episode in healthy older people is due to an increased number and/or increased duration of awakenings by evaluating the rates of transition between sleep and wake bouts within a bedrest episode. DESIGN: Analysis of previously reported polysomnographic data from 13 older and 11 younger healthy individuals scheduled to sleep at many different phases of the endogenous circadian cycle during conditions of forced desynchrony (18.7 hours wake: 9.3 hours bedrest) of the circadian and wake-bedrest cycles. SETTING: General clinical research center PATIENTS OR PARTICIPANTS: None. INTERVENTIONS: None. MEASUREMENTS AND RESULTS: Older subjects had an approximately 2.7-fold increased rate of awakening from sleep but the same rate of falling back asleep as younger subjects. These differences between young and older individuals were observed at most circadian phases and throughout the bedrest episodes. In addition, the circadian variation in transition rates was greater in younger than older subjects. CONCLUSIONS: These results suggest that the reduced consolidation of sleep within a bedrest episode is due to difficulties remaining asleep, rather than falling asleep once awake, and is a primary change in sleep with aging. PMID- 15283017 TI - The maintenance of wakefulness test in normal healthy subjects. AB - STUDY OBJECTIVES: The Maintenance of Wakefulness Test (MWT) examines an individual's ability to stay awake in an environment of decreased sensory stimulation. Only 1 previous study has systematically examined the MWT in normal healthy subjects. SETTING: Sleep disorders unit laboratory PARTICIPANTS AND DESIGN: 31 subjects (mean age 48.5 years, SD 9.6; 8 women) were randomly selected via the telephone directory within a 30-km radius of the test centers. They answered a general screen for health complaints (respiratory, cardiovascular, and psychiatric disorders) and sleep difficulties (snoring). INTERVENTIONS: N/A. MEASUREMENTS AND RESULTS: Overnight polysomnography and a 40-minute MWT the following day were performed on all subjects. Mean sleep latency to the first epoch of unequivocal sleep during the 40-minute trial MWT was 36.9 +/- 5.4 (SD) minutes. The lower normal limit, defined as 2 SD below the mean, was therefore 26.1 minutes. Mean sleep latency for the first 20 minutes of the trial (with sleep latency defined as time to the first appearance of 1 epoch of stage 1 sleep or a 10-second microsleep) was 18.6 +/- 2.3 minutes, with a lower normal limit of 14.0 minutes. CONCLUSIONS: The mean results are consistent with previously published normative data. However, the SDs found in this study are smaller, and, thus, the lower normal limit suggested here is 4 to 6 minutes longer. The subjects in this study were randomly selected from the general population and may, therefore, be a truer representation of the normal population than in the previous study in which subjects were recruited via hospital advertisements and word of mouth. PMID- 15283018 TI - Effect of varying recording cable weight and flexibility on activity and sleep in mice. AB - STUDY OBJECTIVES: Sleep in mice has typically been determined from electroencephalograms and electromyograms recorded via cables in tethered animals. However, the relatively small physical size of mice can produce concerns in recording with cables that may not be seen in larger animals. To examine the influence of implantation and tethering on mice, we recorded activity and sleep in 2 strains while they were attached to 3 cable configurations that varied in weight and flexibility. DESIGN: Activity was recorded prior to surgery and after surgery without tethering. Afterward, the mice were habituated to 3 cable configurations (light [L]: 1.5 g; medium [M]: 2.2 g; heavy [H]: 3.0 g), and activity and sleep were recorded for 2 consecutive days under each configuration. SETTING: N/A. PATIENTS OR PARTICIPANTS: Studies were conducted in 2 mouse strains that differ significantly in levels of spontaneous activity (more-active strain: BALB/cJ [C]; less-active strain: DBA/2J [D2]). INTERVENTIONS: N/A. MEASUREMENTS AND RESULTS: Significant postsurgery reductions in activity in nontethered mice were found only in the more-active C strain. Activity in both strains was reduced in a graded manner as cable weight increased and flexibility decreased. In contrast, changes in sleep were not graded across cables, and changes in rapid eye movement sleep showed more variability. In addition, the effects of varying cables were not consistent across strains. CONCLUSIONS: The differential impact that variations in the weight and flexibility of recording cables can have on the amount and patterns of sleep is an important consideration in conducting and interpreting sleep studies in mice. PMID- 15283019 TI - Sleep neurobiology for the clinician. AB - Many neurochemically distinct systems interact to regulate wakefulness and sleep. Wakefulness is promoted by brainstem and hypothalamic neurons producing acetylcholine, norepinephrine, dopamine, serotonin, histamine, and orexin/hypocretin. Each of these arousal systems is capable of increasing wakefulness, but coordinated activity in all these pathways is required for complete alertness and cortical activation. Neurons in the pons and preoptic area control rapid eye movement and non-rapid eye movement sleep. Mutual inhibition between these wake- and sleep-regulating regions likely helps generate discrete behavioral states. An up-to-date understanding of these systems should allow clinicians and researchers to better understand the effects of drugs, lesions, and neurologic disease on sleep and wakefulness. PMID- 15283020 TI - Hypersomnia associated with isotretinoin in a patient with recurrent major depressive disorder and acne vulgaris. PMID- 15283021 TI - [The role of multidetector computed tomography in the diagnosis in 3 patients with ischaemic heart complaints]. AB - Multidetector CT (MDCT) can provide important information before or after coronary angiography (CAG). This is illustrated by three cases. In a 21-year-old female with ventricle fibrillation CAG demonstrated an anomalous right coronary artery, the exact course of which could not be evaluated. MDCT demonstrated a course between the aorta and pulmonary trunk with vessel compression during systole. After a bypass operation and subsequent pacemaker implantation, she was asymptomatic at follow-up six weeks after hospital discharge. In a 46-year-old male with chest pain, MDCT showed triple vessel disease after which percutaneous coronary intervention (PCI) with stent implantation of the three main branches was performed. Two months after discharge, the patient did not report any complaints. In a 51-year-old male scheduled for PCI of the left anterior descending coronary artery (LAD), MDCT detected a LAD thrombus with 90% occlusion prior to PCI. The occlusion was confirmed during CAG and treated with angioplasty. The next day he was discharged. MDCT offers a practical solution for different cardiac problems through its high diagnostic value. PMID- 15283022 TI - [Practice makes perfect. The favourable effect of experience on the outcome of care]. AB - The number of times that a certain procedure is performed in a hospital per year affects the outcome following this procedure. Such an effect on postoperative mortality has clearly been shown for many procedures in different countries. It is most outspoken for high-risk gastrointestinal surgery, e.g. pancreatectomy and oesophagectomy, and to a lesser degree for surgery for abdominal aneurysms. The experience of the surgical team, which includes surgeons, anaesthetists, nurses and paramedical staff, is responsible for one half of the beneficial effect and the surgeon for the other half. The percentage varies for different procedures, but teamwork and the hospital are important factors in all high-risk procedures. Pancreatectomies, oesophagectomies and high-risk vascular surgery should be concentrated in dedicated centres with a sufficient volume to obtain a good outcome of surgical care. PMID- 15283023 TI - [Multidetector computed tomography of the coronary arteries]. AB - In the past decade, improvements in CT techniques have enabled non-invasive visualization of the coronary arteries. Multidetector CT (MDCT) is currently the generally accepted technique for the follow-up of coronary stents and by-pass grafts, and for the evaluation of anomalous coronary arteries and coronary artery disease. Both the degree of stenosis, as well as plaque composition can be determined by MDCT. Plaque composition has proven to be a more important predictor for acute coronary syndromes than the degree of stenosis. In addition, MDCT has less risks of complication and lower costs. Limitations of MDCT are: sensitivity to rhythm- and breathing artefacts, a lower spatial and time resolution than coronary angiography (CAG), and difficulties in coronary evaluation close to high density structures such as calcifications and stents. Coronary angiography is still indicated when functional information has to be obtained about coronary flow. MDCT should be considered in all cases in which diagnostic CAG is performed. PMID- 15283024 TI - [Fatigue in neuromuscular disease]. AB - Chronic fatigue is a symptom of diseases such as cancer, multiple sclerosis, Parkinson's and cerebrovascular disease. Fatigue can also be present in people with no demonstrable somatic disease. If certain criteria are met, chronic fatigue syndrome may be diagnosed in these cases. Fatigue is a multi-dimensional concept with physiological and psychological dimensions. The 'Short Fatigue Questionnaire' consisting of 4 questions is a tool to measure fatigue with a high degree of reliability and validity. Within the group of neuromuscular disorders, fatigue has been reported by patients with post-polio syndrome, myasthenia gravis, and Guillain-Barre syndrome. The percentage of neuromuscular patients suffering from severe fatigue (64%) is comparable with that of patients with multiple sclerosis, a disease in which fatigue is an acknowledged symptom. Now that reliable psychological and clinical neurophysiological techniques are available, a multidisciplinary approach to fatigue in patients with well-defined neuromuscular disorders may contribute towards the elucidation of the pathophysiological mechanisms of chronic fatigue, with the ultimate goal being to develop methods of treatment for fatigue in neuromuscular patients. PMID- 15283025 TI - [Diagnostic image (196). A newborn with fever and a swollen finger]. AB - A 5-day-old girl was presented with monoarthritis of the proximal interphalangeal joint of the right fourth digit, without signs of septic illness or meningitis, due to group B Streptococcus agalactiae. PMID- 15283026 TI - [From gene to disease; primary open-angle glaucoma and three known genes: MYOC, CYP1B1 and OPTN]. AB - Primary open-angle glaucoma (POAG) is a group of multifactorial diseases that affects 1.5% of the population. If untreated, the disease leads to irreversible damage to the visual system. The clinical features of POAG are excavation of the optic disc and visual field defects, probably due to degeneration of retinal ganglion cells. Important risk factors for POAG are older age, elevated intraocular pressure, the presence of POAG in relatives, and still largely unknown molecular genetic factors. The clinical, genetic and pathological heterogeneity most likely reflects the complex heterogeneous situation at the molecular level. The three genes known to be involved in POAG (MYOC, CYP1B1 and OPTN) account for up to 18% of the POAG cases. These findings result in new possibilities for the presymptomatic molecular diagnosis of POAG. PMID- 15283028 TI - [Three patients with complications following piercing of the auricular cartilage]. AB - In three patients, a 21-year-old man and females aged 16 and 18 years, piercing through the auricular cartilage was followed by an infection. Treatment left behind a residual deformity, for which a reconstruction was carried out with a satisfactory result. The risk of infection following piercing through the avascular cartilage of the helix and tragus of the pinna is greater than following piercing of non-cartilaginous tissue such as the ear lobe. (Peri)chondritis leads to chondral necrosis and subsequent deformities. It is important to recognise the early features of perichondritis, which include local heat, erythema and pain, before swelling appears. Treatment should focus on eradicating Pseudomonas aeruginosa and Staphylococcus aureus. Surgical intervention is required at the earliest sign of an abscess. Reconstruction for a post-piercing deformity can be considered at a later stage. PMID- 15283027 TI - [Growth charts for height, weight and body-mass index of twins during infancy]. AB - OBJECTIVE: To determine the magnitude of the growth retardation in Dutch monozygotic and dizygotic twins during infancy in comparison with the Dutch reference growth charts for general population infants from 1997 and to construct reference growth charts for twins. DESIGN: Descriptive. METHOD: The growth of twins was studied using longitudinal data on over 4000 Dutch twin pairs from birth until 2.5 years of age. The LMS method was used to obtain growth charts for height, weight and body-mass index (BMI) for twin pairs during infancy. Centiles were estimated by the Box-Cox power curve (L), the median curve (M) and the coefficient of variation curve (S). RESULTS: From birth until the age of half a year, the average height and weight of twin pairs were at about the 10th percentile of the Dutch reference population. One year later this difference had decreased to about the 25th percentile, and when the twin pairs were between 1.5 and 2.5 years of age the difference was further decreased to the 35th percentile. The BMI deviated less from that of the reference population: during the first half a year the BMI of twin pairs was at about the 25th percentile. Subsequently, the BMI improved, but remained slightly below the median of the reference population at the age of about two years. Approximately half (50% for height, 58% for weight) of the growth retardation from birth until 1.5 years was attributable to gestational age. Between 1.5 years and 2.5 years of age, this difference was reduced to one third: 33% for both height and weight. Thus, a substantial part of the growth difference could not be explained by gestational age. CONCLUSION: Correcting for gestational age alone is not sufficient to make possible a comparison of the growth of twin pairs with the growth of general population infants. The development of twins can, however, be followed by means of the reference growth charts designed by the authors. PMID- 15283029 TI - [Young adults with Ewing's sarcoma]. AB - Ewing's sarcoma was diagnosed in three men, one aged 22 and two aged 30. The disease was diagnosed by biopsy and chromosome investigations (t(11;22) translocation). In the youngest patient with localised disease, supplementary radiotherapy was withheld in view of the good results of induction chemotherapy, surgery and consolidation chemotherapy. However, four months later, there was a localised recurrence, again followed by induction chemotherapy, chemotherapy at high dosage, stem cell transplantation, radiotherapy and finally surgical intervention, after which a complete remission was achieved. The 30-year-old man with localised disease was given induction chemotherapy, surgery, consolidation chemotherapy and radiotherapy; 14 months after the diagnosis he was in good condition. The other 30-year-old man had metastases in TXII and both lungs. Despite intensive therapy he died 8 months after diagnosis. Ewing's sarcoma is a musculoskeletal malignancy that occurs in children and adolescents but also in young adults. It generally manifests itself as a painful swelling originating in bone or soft tissue. There are often accompanying symptoms such as weight loss and fever. In 20-25% of cases there are already metastases (to the lungs, bone and bone marrow) by the time of diagnosis. The diagnosis and treatment of this rare, therapy-sensitive disease should take place in a study setting and in co operation with a multidisciplinary sarcoma working group. PMID- 15283030 TI - [Summary of the practice guideline 'Acute cough' from the Dutch College of General Practitioners]. PMID- 15283031 TI - [The practice guideline 'Hormonal contraception' (second revision) from the Dutch College of General Practitioners; a response from the perspective of obstetrics & gynaecology]. PMID- 15283032 TI - [The 2002 recommendations on the prophylaxis of infectious endocarditis]. PMID- 15283033 TI - [Sudden death by coronary spasm: a particular and often unrecognised clinical entity]. AB - The authors report 3 cases of resuscitated sudden death in which the investigations clearly showed coronary spasm. This was demonstrated by systematic coronary angiography with an ergometric test. Two patients underwent electrophysiological investigations which were normal. The three patients were prescribed long-term calcium antagonist therapy and one of them underwent coronary angioplasty. With a follow-up of 6 months to 3 years, there was no clinical recurrence or documented arrhythmia. A review of the literature shows that this is a cause of sudden death which is probably underestimated and unrecognised. Electrophysiological investigations often give disappointing results and medical therapy is the keystone of treatment. Coronary angioplasty and implantation of an automatic defibrillator are second-line treatments reserved to forms refractory to medical therapy. PMID- 15283034 TI - [Traumatic aortic regurgitation: diagnostic, management and treatment]. AB - Traumatic aortic valve regurgitation is a rare complication of non-penetrating thoracic trauma. The most frequent lesion is the isolated injury of the non coronary cusp. Actually, the transoesophageal echocardiography is the procedure of choice to confirm the diagnosis and to reveal the associated cardiovascular lesions. Surgical management with early operation is the best policy, however this surgery can be delayed for treatment of other life-threatening injuries. Up today, aortic valve replacement (AVR) was recommended to repair traumatic aortic valve regurgitation; nevertheless, in the recent international literature, the number of cases reports with conservative surgery (CS) is increasing: 10 AVR (group I) and 10 CS (group II). Analysis of the post-operative and long term periods shows good results: it confirms the excellent clinical evolution in the group I (mean time of follow-up: 18.2 +/- 16.3 months), and reveals satisfactory results in the group II for patients with isolated lesion (mean time of follow up: 29.1 +/- 30.7 months). In conclusion, each time the traumatic aortic regurgitation is due to an isolated lesion, the conservative surgery should be performed in order to avoid aortic valve replacement and its potential complications especially in young patients with healthy valves. However, the aortic valve replacement is the safest technique for complex or multiple injuries of the aortic valve. PMID- 15283035 TI - [Echographic calculation of left ventricular mass in TM mode]. AB - TM sections of the left ventricle (LV) have only been validated in fundamental (F) imaging. We were interested in evaluating the repercussions of new imaging techniques, harmonic (H) and colour tissue Doppler (CTD), on the TM measurement of left ventricular mass (LVM), the reference imaging being F imaging. METHOD: We performed a prospective study, including 26 patients with a valid TM section. The LV and LVM parameters in F, H and CTD mode according to the Penn and ASE conventions as well as the inter-observer reproducibility were studied. RESULTS: The correlations for the LVM measurements between F and H, and between F and CTD were high whichever convention was used (r>0.95, p<0.0001). For each observer, the LVM in H and in CTD was always greater than the LVM in F with both conventions (p<0.02). A false diagnosis of LV hypertrophy was made in 27% of patients in H and in 15% of patients in CTD. The best inter-observer reproducibility was obtained in H: the average inter-observer difference (gr.) was 23+/-15 for H, 32+/-19 for F and 59+/-18 for CTD. CONCLUSION: H and CTD imaging entail an overestimation of LVM, essentially by overestimation of the parietal thickness of the LV. The inter-observer reproducibility was excellent in H and poor in CTD. The use of the harmonic mode for LVM calculation must be validated using new formulae. PMID- 15283036 TI - [New French recommendations for the prophylaxis of infectious endocarditis]. AB - New French recommendations on infective endocarditis (IE) prevention were recently published and mark a turning point in the history of antibiotic prophylaxis. Endocarditis is an evolving disease, and its clinical and microbiological profile dramatically changes over time. The French surveys that were conducted in 1991 and 1999 showed variations in underlying heart disease with a decrease in native valvular disease and an increase of IE in patients without previously known heart disease. Moreover, the distribution of responsible microorganisms dramatically changed over time, with a marked decrease of oral streptococci. In addition, some dogmas are now challenged. First of all, the part of responsibility of dental procedures is debated, as dental bacteraemia possibly responsible of endocarditis are more likely due to daily manoeuvres such as tooth brushing or chewing gum than to occasional dental procedures. Moreover, as suggested by case-control studies, efficacy--or lack of efficacy--of antibiotic prophylaxis is far from being clinically proved. For all these reasons, the proportion of theoretically avoidable endocarditis seems very low, and the benefit of largely and systematically applied antibiotic prophylaxis may be discussed, not only in terms of financial cost but also in terms of microbiological threat of emergence of antibiotic resistant bacteria. So, the general idea of those new recommendations was to maintain the principle and the modalities of antibiotic prophylaxis, but to limit its indications to situations at high benefit to risk ratio, i.e. procedures at high risk in patients at high risk. Depending on the situation, antibiotic prophylaxis may be either recommended or become optional and decision-making factors are defined. Furthermore, the importance of general prophylaxis was emphasised, concerning more specifically oral and cutaneous hygiene, and patients and practitioners' education, such as, for example, recommendations on blood cultures to be performed before any antibiotic treatment in case on fever occurring in a patient at risk during the 3 months following a procedure at risk. PMID- 15283037 TI - [Specific cyclo-oxygenase-2 inhibitors in cardiovascular pathology]. AB - Cyclo-oxygenase catalyses the conversion of arachidonic acid and O2 into prostaglandins. Its two isoenzymes, COX-1 and COX-2, have different functions. COX-1 is expressed in constituent form in most tissues where it controls the production of arachidonic acid metabolites which maintain the physiological tissue integrity. By contrast, COX-2 is only induced in response to inflammatory stimuli, resulting in the production of inflammation mediating prostaglandins. Conventional non-steroidal anti-inflammatories inhibit both enzymes in a non specific manner. The recent development of specific COX-2 inhibitors, which retain the same anti-inflammatory efficacy but do not have the gastric side effects of conventional treatment related to COX-1 inhibition, gives them a greater safety margin. However, coronary events are observed in patients treated with COX-2 inhibitors. This risk, seemingly confirmed at least at higher dosages, has been attributed to a disequilibrium in the balance of thromboxane A2/prostacyclin (TxA2/PGI2) which they induce. Paradoxically, COX-2 inhibitors also have several favorable effects on atheromatous plaque progression and its inflammatory component, not only in vitro but also therapeutically, including situations where platelet activation and arterial thrombosis are predominant, such as in acute coronary syndrome. The salt retention induced by COX-2 inhibitors could also be the origin of an increase in blood pressure, and in susceptible subjects it could provoke cardiac decompensation. These multiple cardiovascular risks tinge the safety profile of COX-2 inhibitors, especially in elderly subjects and those with multiple pathology, for whom extra surveillance is required. PMID- 15283038 TI - [Is there an indication for the association of betablockers and angiotensin II receptor antagonists in cardiac failure?]. AB - ACE inhibitors initially developed as vasodilators are effective by their anti hormonal action. Antagonists of the receptors of angiotensin II (ARA II) should provide an equivalent or better blockade of the rennin-angiotensin system (absence of tolerance). Clinical trials have shown equivalent haemodynamic effects of the two classes, equal functional tolerance but mortality studies have shown more variable results. None have shown the superiority of ARA II over ACE inhibitors and the demonstration of their equivalence has just been reported with high doses in the post-infarction period. A deleterious effect of ARA II in association with betablockers was reported in two mortality studies but has not been confirmed in the most recent trials. The difficulty is to determine the roles of the association of ARA II-ACE inhibitors, ARA II-antialdosterones or of the association of all three classes of molecules. PMID- 15283039 TI - [Ischaemic mitral insufficiency]. AB - Ischaemic mitral insufficiency (IMI) due to regurgitation of an anatomically normal valve, due to dysfunction directly related to myocardial ischaemia, is observed in over 20% of post-infarction patients and is associated with a doubling of the risk of death. The responsibility of ventricular remodelling with displacement of the papillary muscles in the genesis of IMI has been demonstrated experimentally. 3-D echocardiography has improved our understanding of the central role of geometrical changes of the subvalvular apparatus. The inconsistent results of surgery using an undersized mitral annulus have led to the search for alternative techniques. The correction of mitral insufficiency at coronary bypass surgery is a current topic of research. The application of new techniques of mitral valvuloplasty seems more effective and should provide an answer to this problem. PMID- 15283040 TI - [Prophylaxis of infectious endocarditis]. PMID- 15283041 TI - PPAR alpha, fibrates, lipid metabolism and inflammation. AB - Peroxisome proliferator-activated receptors (PPARs) are lipid-activated transcription factors belonging to the nuclear receptor superfamily. Three PPAR isotypes have been characterised: alpha, beta/delta and gamma. Identification of selective ligands and gene targeting studies have allowed determination of subtype-specific functions and the therapeutic potential of these receptors. Thus, PPAR alpha plays a prominent role in several physiological processes including the control of lipid and lipoprotein metabolism, glucose homoeostasis and the inflammatory response. Here, we update the potential of PPAR alpha activation in the treatment of metabolic disorders and related cardiovascular complications. PMID- 15283042 TI - Contribution of PAI-1 in cardiovascular pathology. AB - Plasminogen activator inhibitor-1 (PAI-1), is a serpin whose major function is to negate plasminogen activation and impair fibrinolysis. It occurs in plasma and tissues. Studies in genetically modified mice indicate that PAI-1 might be involved in thrombosis, vascular healing and atherosclerosis although contradictory findings have been obtained in the latter two processes. Differences between results depend on the types and the lengths of the models and underline the fact that besides its role in regulating fibrinolysis, PAI-1 plays a role in several cellular processes independent of plasminogen activation. In patients, high plasma PAI-1 levels worsen the prognosis of myocardial infarction in the acute phase and have been considered as a risk factor for coronary heart disease. The predictive capacity of PAI-1 is mainly related to several metabolic covariates which constitute the metabolic syndrome (MS). This syndrome is associated with increased cardiovascular morbidity and mortality and is becoming one of the major health problems as its prevalence is growing rapidly. Accelerated atherothrombotic process in the MS is attributed not only to metabolic abnormalities but also to a specific inflammatory state which leads to increased plasma PAI-1 levels. Modifying PAI-1 expression by PAI-1 inhibitors may open a new field of research and may reveal the true role of PAI-1 in atherosclerotic and insulin resistance processes. PMID- 15283043 TI - Angiogenesis therapy with human tissue kallikrein for the treatment of ischemic diseases. AB - Angiogenesis is essential for the repair of wounds and tissues damaged by ischemia. The regenerative process is tightly regulated by master angiogenic factors, cytokines and the downstream mediator NO. In addition, modulators of vascular growth, such as COX-2-generated prostanoids, contribute to the process by stabilizing the hypoxia-inducible factor and stimulating the expression of VEGF. Recently, we discovered that human tissue kallikrein, a member of the serine proteinase superfamily, possesses potent angiogenic effects. It has been categorized as a pleiotropic angiogenic agent acting via enzymatic cleavage of kininogen and subsequent release of kinin peptides. Kinins bind G-protein coupled receptors, subtype B1 and B2, and exert proliferative effects on endothelial cells via an IP3K-Akt-NO mediated mechanism independent of VEGF. In addition, kinins stimulate the release of angiogenic prostacyclin. Gene transfer of human tissue kallikrein rescues ischemic tissues in otherwise normal mice, as well as in hypertensive or diabetic animals. In addition, prophylactic gene delivery of tissue kallikrein to diabetic skeletal muscles prevents the development of microangiopathy and stimulates collateralization, thus protecting from the consequences of supervening arterial occlusion. PMID- 15283044 TI - [Brugada syndrome and supraventricular arrhythmias]. AB - The author reports the case of a 46-year old patient diagnosed with idiopathic ventricular fibrillation (Brugada syndrome) further to induction of class Ic antiarrhythmic therapy for the management of paroxystic ventricular fibrillation. It would appear that this diagnosis is increasingly frequent in young patients with Brugada syndrome shown to be minimal or intermittent on electrocardiograms. Atrial arrhythmia was the only rhythmic pathology objectively evidenced in this patient and the author was consequently led to reconsider its prevalence in patients presenting this syndrome both in the literature and according to his personal experience. PMID- 15283045 TI - [Right atrial thrombus--a complication of central venous catheters]. AB - We describe the development, in three days, of a pediculate mass hanging on the right atrial lateral wall in a 39-year-old woman with a subclavian venous catheterization. She was a current smoker and alcoholic but without drug addict. The hypothesis of a non valvular right atrial infective endocarditis was considered at first, but subsequent events directed the diagnosis towards a thrombus, which was resorbed by heparin. We discuss the incidence, the complications, the treatment and the differential diagnosis of thrombus caused by a central venous catheter. The prevention of right atrial thrombus caused by a central venous catheter depends on the position of the central venous catheter tip, either in the superior vena cava or at the superior vena cava-right atrium junction. A more distal position is a frequent source of thrombotic and embolic complications. PMID- 15283046 TI - Significance of chest computed tomography findings in the evaluation and treatment of persistent gestational trophoblastic neoplasia. AB - OBJECTIVE: To evaluate whether chest computed tomography (CT) findings in patients with persistent gestational trophoblastic neoplasia (GTN) and a negative chest roentgenogram (CXR) significantly influence clinical outcome and to determine potential clinical predictors of pulmonary micrometastasis STUDY DESIGN: The charts of 201 patients with nonmetastatic GTN (International Federation of Gynecology and Obstetrics [FIGO] stage I) receiving primary treatment with methotrexate (MTX) infusion between December 1985 and December 2000 were reviewed, and data were collected on age, gravidity and parity, FIGO stage, histologic diagnosis, metastatic disease, radiologic findings, surgery, presenting human chorionic gonadotropin (hCG) level, total number of chemotherapy courses and chemotherapeutic agents required to reach remission, and time to remission. The chi2, regression, Kaplan-Meier and log-rank tests were utilized to evaluate the correlation of chest CT with CXR findings, histology of antecedent pregnancy, presenting hCG level, chemotherapeutic requirements and time to remission. RESULTS: Of 30 patients with a negative CXR, 13 (43.3%) had chest CT positive for micrometastasis. Histology of the antecedent pregnancy and mean presenting hCG did not correlate with the chest CT result. There was no significant difference between patients with positive or negative chest CT results in the requirement for > 1 dose of MTX or for additional chemotherapeutic agents. There also was no significant difference in time to remission by chest CT status. Regression analysis using histologic diagnosis, presenting hCG level, age, gravidity and parity as covariates did not reveal any clear risk factors for pulmonary micrometastasis. CONCLUSION: It has been suggested that GTN patients with micrometastases identified on chest CT only are at increased risk of requiring > 1 dose of MTX or of requiring additional chemotherapeutic agents. Our data suggest that chest CT alone is not predictive of clinical outcome. Furthermore, the presence of micrometastases does not correlate with hCG level or histologic diagnosis, and there are no clear risk factors for pulmonary micrometastases. PMID- 15283047 TI - Molecular biology of gestational trophoblastic neoplasia: a review. AB - Gestational trophoblastic diseases are interrelated conditions characterized by abnormal growth of chorionic tissues with varying propensitiesfor local invasion and metastases. These diseases are characterized by altered expression of several growth regulatory factors and oncogenes. On the basis of the expression of various oncogenes and growth factors, partial mole appears to be more like normal placenta, while complete mole seems to be more like choriocarcinoma. These results may have both prognostic and therapeutic consequences and provide insight into the relationship between normal placenta and gestational trophoblastic diseases. PMID- 15283048 TI - Inappropriate management of women with persistent low hCG results. AB - The USA hCG Reference Service is a consulting service with a specialized clinical laboratory aiding physicians in the interpretation of conflicting or nonrepresentative human chorionic gonadotropin (hCG) results. We have consulted on 189 cases with persistent low levels of hCG but no evidence of pregnancy or tumor. Quiescent gestational trophoblastic disease (GTD) was identified in 121 cases by the absence of invasive trophoblast antigen and nonresponse to chemotherapy (64 cases with a history of hydatidiform mole or gestational trophoblastic neoplasia (GTN) and 57 cases following antecedent pregnancy). Another 61 Reference Service cases hadfalse positive hCG, and we observed 7 cases with low levels of hCG of pituitary origin (hCG subsequently suppressed by estrogen-progesterone medication). Most disturbing is that the majority of these cases (68%) received needless therapy for assumed GTN/choriocarcinoma/placental site trophoblastic tumor before consultation with the Reference Service. One hundred twenty-eight of the 189 patients (77 of 121 with quiescent GTD, 48 of 61 withfalse positive hCG and 3 of 7 with pituitary hCG) underwent therapy ranging from single-agent chemotherapy (117 cases), to EMA-CO combination chemotherapy (etoposide, methotrexate, actinomycin D alternating with cyclophosphamide and vincristine) (16 cases), to hysterectomy and/or bilateral salpingo-oophorectomy (31 cases). False positive hCG and pituitary hCG would obviously not respond to these treatments, and no treated cases of quiescent GTD responded to chemotherapy orfully responded to hysterectomy. The continued needless treatment of patients with quiescent GTD, even after multiple publications, is entirely avoidable. Unfortunately, the number of needlessly treated cases referred to the Reference Service is increasing. PMID- 15283049 TI - What we know about low-level hCG: definition, classification and management. AB - OBJECTIVE: To critically assess the literature on the syndrome of low-level "real" human chorionic gonadotropin (hCG), to add new cases from our practice, to enumerate the investigations that are essential in the management of these patients and to offer a working classificationfor use by physicians encountering the condition. STUDY DESIGN: We report our experience with 9 patients with low level hCG treated at the Yale Trophoblast Center and discuss London and Sheffield patients as well as reports from the USA hCG Reference Service. RESULTS: One of the 9 Yale patients had developed placental site trophoblastic tumor metastatic to the lung. Following resection and 18 months of observation, she then had a successful pregnancy, has remained without evidence of disease and has negative hCG. The other patients continue to be observed. The experience from England shows that 2 of 14 patients with detectable hCG in urine and serum developed overt trophoblastic neoplasia and were treated successfully. The others are being followed. None have developed gestational trophoblastic neoplasia, 3 have regular menstrual periods, and 1 has had 2 pregnancies. The USA hCG Reference Laboratory has had 114 consultations. Sixty-three patients had real hCG and were followed for 6 months to 6 years. Forty of the 63 (63%) received single agent or combination chemotherapy, and 10 underwent hysterectomy, also. hCG persisted in spite of therapy. Four of the 63 (6%) eventually developed overt trophoblastic neoplasia and were then treated effectively; their hCG became negative. In these 4 patients whose hCG rose significantly and who did require therapy, the proportion of hyperglycosylated hCG became > or =80% of total hCG. In contrast, the proportion of hyperglycosylated hCG was always very low in the 63 quiescent cases. CONCLUSION: Active therapy with chemotherapy or surgery for persistent, elevated, low-level, real hCG is counterproductive. Therapy should be initiated only if overt trophoblastic neoplasia appears. All patients with low-level, real hCG require sophisticated imaging to exclude the presence of extrauterine sites of trophoblast, such as trophoblastic metastases or pituitary adenoma. They require long-term follow-up with periodic clinical examination, imaging and frequent hCG testing with an assay that measures all aspects of the hCG molecule. PMID- 15283050 TI - Salvage chemotherapy for high-risk gestational trophoblastic tumor. AB - OBJECTIVE: To evaluate the efficacy and safety of etoposide/methotrexate/actinomycin D (MEA regimen) as initial chemotherapy and 5 fluorouracil/actinomycin D (FA regimen) as salvage chemotherapy for high-risk gestational trophoblastic tumor (GTT). STUDY DESIGN: From 1985 to 2001, 36 patients with World Health Organization (WHO)--defined high-risk GTT were treated with MEA or FA at Chiba University Hospital. Thirty-three patients were initially treated with MEA. FA was administered to 11 patients; 1 had had no previous chemotherapy, 7 had developed drug resistance to MEA, 1 had relapsed following MEA, and 2 had relapsed following etoposide/methotrexate/actinomycin D/ cyclophosphamide/vincristine (EMA/CO) combination chemotherapy. RESULTS: The primary remission rate with MEA was 69.7% (23 of 33). With FA the survival rate was 81.8% (9 of 11) for a mean follow-up period of 11.5 years. Two patients died due to multidrug resistance, and 2 patients relapsed subsequently. The 2 relapse cases were successfully salvaged again with MEA. The toxicity of FA was evaluated in 89 cycles. Myelosuppression seemed to be the dose-limiting toxicity, and the incidence of WHO grade 4 leukocytopenia and thrombocytopenia were 5.6% and 3.4%, respectively. CONCLUSION: Although etoposide-containing chemotherapy is currently the most effective and well tolerated regimen for high-risk GTT, 20-30% of patients develop drug resistance to these regimens. Salvage combination chemotherapy with FA is effective for refractory patients, and the toxicity is predictable and manageable. PMID- 15283051 TI - EMA/EP chemotherapy for chemorefractory gestational trophoblastic tumor. AB - OBJECTIVE: To evaluate the results of etoposide/ methotrexate/actinomycin D/etoposide/cisplatin (EMA/ EP) chemotherapy in patients with chemorefractory gestational trophoblastic tumor (GTT). STUDY DESIGN: Fifteen patients with chemorefractory GTT were treated with EMA/EP. RESULTS: Twelve of the 15 cases were choriocarcinoma, and the last 3 were metastatic placental site trophoblastic tumor (PSTT): International Federation of Gynecology and Obstetrics (FIGO) stage I, 2 cases; stage III, 10 cases; stage IV, 3 cases. Seven cases have FIGO score 7 10; the scores of the remaining 8 cases were > 10. Fifteen patients received a total of 93 cycles of the study regimen. The median number of courses for each patient was 6.2. Eleven cases (73.3%) achieved complete remission, while 3 (20%) had partial remission; 1 case (6.7%) showed no response. The main complications of EMA/EP chemotherapy were myelosuppression and gastrointestinal symptoms. CONCLUSION: The EMA/EP regimen is effective for chemorefractory GTT, and the chemotherapeutic results can be improved when combined with surgery and arterial infusion chemotherapy in selected patients. The EMA/EP regimen should be considered for primary management of metastatic PSTT. PMID- 15283052 TI - Placental site trophoblastic tumor: a review of 7 cases and their implications for prognosis and treatment. AB - OBJECTIVE: To review cases of placental site trophoblastic tumor (PSTT) for prognostic factors and treatment implications. STUDY DESIGN: Between 1982 and 2003, 7 cases of PSTT were treated at the Brewer Trophoblastic Disease Center. Pathology and operative reports, patient records and laboratory results were reviewed. Data collected included patient age, presenting symptoms, human chorionic gonadotropin (hCG) levels, type of antecedent pregnancy, International Federation of Gynecology and Obstetrics (FIGO) stage at diagnosis, mitotic count and immunohistochemical expression of the tumor, treatments, response to treatments and length of survival. RESULTS: The mean age of patients was 34 years. The most frequent presenting symptom was vaginal bleeding (72%). The antecedent pregnancy was a normal, term vaginal delivery in 4 patients, spontaneous or elective abortion in 2 and unknown in 1. The mean interval from last pregnancy to diagnosis of PSTT was 3.2 years (range, 4 months-8 years). Serum hCG levels at the time of diagnosis ranged from 2 to 456 mIU/mL (mean, 130). All patients initially underwent surgery (hysterectomy and/or other procedures), and those with metastatic disease also received chemotherapy (most commonly etoposide, methotrexate, actinomycin D/etoposide, cisplatin [EMA/EP]). The 4 patients with recurrent or advanced disease had additional surgical procedures (thoracotomy, excision of vaginal metastases or laparotomy with intraperitoneal and retroperitoneal disease resection) as well as multiple other types of chemotherapy (e.g., bleomycin, etoposide and cisplatin; ifosfamide, carboplatin and etoposide; carboplatin/paclitaxel). Survival was 57% overall: 75%for the 4 patients with FIGO stage I disease and 33% for the 3 with FIGO stage IV. The 2 patients with mitotic counts < 2 per 10 high power fields survived. Three patients were alive and without evidence of disease for 17 years, 16 years and 8 months; 1 patient was alive with recurrent metastatic disease at 20 months; and 3 patients were dead of disease 13, 30 and 33 months after diagnosis. CONCLUSION: Advanced FIGO stage, long interval from last known pregnancy to diagnosis and high mitotic count were adverse prognostic indicators for survival in PSTT. All patients with PSTT should undergo initial hysterectomy with other surgical procedures, as indicated. Chemotherapy, usually EMA/EP, should be used in patients with advanced PSTT and may be considered in patients with FIGO stage I disease with length of time from antecedent pregnancy >2 years or high mitotic PMID- 15283054 TI - Atypical squamous cells, cannot exclude HSIL: a review of original vs. duplicate thin-layer slides. AB - OBJECTIVE: To assess the utility of consensus review of original ThinPrep slides (Cytyc Corp., Boxborough, Masssachusetts) vs. duplicate slides and to identify the significance of the number of representative cells in classifying atypical squamous metaplastic cells, cannot exclude high grade squamous intraepithelial lesion (ASC-H). STUDY DESIGN: A duplicate ThinPrep slide was prepared from 19 cases of ASC-H. Both original and duplicate slides underwent blinded review by 4 pathologists. Consensus cytologic interpretations (75% agreement) were noted and compared with follow-up diagnoses. Dots with atypical metaplastic cells from each slide were counted and correlated with the consensus cytologic interpretation. RESULTS: Following review of 19 original and 19 duplicate slides, consensus interpretations were reached in 14 and 15 cases, respectively. Nine cases of squamous intraepithelial lesion (SIL) were confirmed by biopsies. Consensus interpretation of original and duplicate slides predicted 7 and 6 cases of SIL, respectively. Consensus interpretation of duplicate slides predicted 60% of histologically benign cases. Two-thirds of nonconsensus cases were histologically benign. Consensus interpretations for high grade SIL were reached more often on slides with greater numbers of atypical squamous metaplastic cells. Fewer abnormal cells were present on the duplicate slides. CONCLUSION: Consensus reviews of original and duplicate slides are equally useful in classifying atypical squamous cells, cannot exclude high grade squamous intraepithelial lesion, but may not be practical in all circumstances. HPV DNA testing of all such cases may better detect clinically significant cases. PMID- 15283053 TI - Floxuridine-containing regimens in the treatment of gestational trophoblastic tumor. AB - OBJECTIVE: To analyze the efficacy of floxuridine (FUDR)-containing regimens in the treatment of gestational trophoblastic tumor (GTT). STUDY DESIGN: Seventy four patients with GTT, 47 with invasive mole and 27 with choriocarcinoma were treated with FUDR-containing regimens. Clinical staging of the disease was: 33 cases of stage I, 3 cases of stage II, 31 cases of stage IIIa, 6 cases of stage IIIb and 1 case of stage IV. RESULTS: The complete response rate of FUDR containing regimens in the treatment of GTT was 91.9% (68 of 74 cases). Six patients, of whom 3 showed signs of drug resistance and 3 showed myelosuppression, had their regimens changed to non-FUDR-containing regimens, and all achieved a complete response. All 7 patients with advanced disease (>IIIb) achieved a complete response. The major adverse event with FUDR-containing regimens was myelosuppression and gastrointestinal toxicity: third- and fourth degree neutropenia in 26% and thrombocytopenia in 6.2%, third-degree vomiting in 57.1% and third-degree diarrhea in 4.3%. CONCLUSION: FUDR-containing regimens are efficient for the treatment of GTT even for patients with advanced or drug resistant disease. PMID- 15283055 TI - Autologous endometrial coculture in patients with IVF failure: outcome of the first 1,030 cases. AB - OBJECTIVE: To analyze the effectiveness of autologous endometrial coculture (AECC) in improving embryo quality and pregnancy rates in 1,030 consecutive cycles of in vitro fertilization-embryo transfer (IVF-ET) utilizing AECC from January 1996 to December 2001. STUDY DESIGN: Embryos from each of 1,030 patients allocated to growth on AECC were analyzed for outcome. All patients had previously undergone failed IVF cycles. During a luteal phase biopsy (5-12 days after the luteinizing hormone surge) performed prior to the treatment cycle, glandular (G) and stromal (S) endometrial cells were isolated by enzymatic digestion and separated based on differential sedimentation rates. These cells were cryopreserved, then plated as a 50%/50% combination of G and S cells prior to embryo exposure. The conditioned medium was changed every 2 days. Embryos were randomly grown on endometrial coculture (ECC) or conventional media if > 6 oocytes were normally fertilized. Otherwise, all embryos were grown on AECC. RESULTS: The patients' mean age was 36.9 (+/-3.1) years. The patients had on average a history of 3.1 (+/- 1.7) failed prior attempts. When comparing a previous cycle (same institution only), the cleaved embryos on day 3 were of an improved quality (6.8+/-1.2 vs. 5.5+/-1.0 blastomeres and 14.6% +/- 9.3 vs. 27.2% +/- 9.8 fragmentation, P <.05). Twenty-two (2.13%) patients did not undergo ET secondary to poor embryonic development. Overall positive and clinical pregnancy rates of 49.8% and 41.5% were noted, respectively. Age remained the most important predictor of outcome. CONCLUSION: We demonstrated a significant improvement in embryo quality with ECC. We also demonstrated that patients with a poor prognosis secondary to prior IVF failures can have a good outcome when utilizing AECC. PMID- 15283056 TI - T allele for VEGF gene-460 polymorphism at the 5'-untranslated region: association with a higher susceptibility to endometriosis. AB - OBJECTIVE: To investigate whether vascular endothelial growth factor (VEGF) gene 5'-UTR-460 polymorphism could be used as a marker of susceptibility to endometriosis. STUDY DESIGN: Women were divided into 2 groups, endometriosis (n = 122) and nonendometriosis (n = 131). Polymorphisms for VEGF were detected by polymerase chain reaction and BstUI (New England Biolabs, Beverly, Massachusetts) restriction enzyme analysis. Genotypes and allelic frequencies between the groups were compared. RESULTS: Proportions of different VEGF polymorphisms in the groups were significantly different. Proportions of cuttable (C) homozygote/heterozygote/ uncuttable (T) homozygotefor VEGF in the groups were 0/44.3/55.7% and 0/63.4/36.6%, respectively. A higher percentage of T/F homozygote and T allele was present in the endometriosis population. The proportions of C/T alleles in the groups were 22.1/77.9% and 31.7/68.3%, respectively. CONCLUSION: T/T homozygotes and the T allele of the VEGF-460 gene are associated with a higher risk of endometriosis. Heterozygotes and C allele are related to the lower risk of endometriosis formation. VEGF polymorphism likely contributes to the pathogenesis of endometriosis and may become a useful markerfor predicting endometriosis susceptibility. PMID- 15283057 TI - Role of obesity in the surgical management of advanced-stage ovarian cancer. AB - OBJECTIVE: To assess whether obese women with advanced-stage ovarian cancer undergoing primary cytoreduction surgery were at increased risk of suboptimal cytoreduction and complications during the operative and postoperative period as compared to nonobese women. STUDY DESIGN: A retrospective, case-control study of all cases of advanced-stage epithelial ovarian cancer managed surgically at Johns Hopkins Hospital between January 1, 1990, and December 31, 1999. RESULTS: Obese patients were as likely as nonobese patients to undergo optimal cytoreduction at surgery. Obese patients were more likely than nonobese patients to be high-risk anesthesia candidates and more likely than nonobese patients to have tumors >20 cm at surgery. Obese patients were not at greater risk of surgical or postoperative complications than were nonobese patients. CONCLUSION: Obesity is not a risk factor for suboptimal surgical management of advanced-stage ovarian cancer. PMID- 15283058 TI - Maternal serum level of placental growth factor in diabetic pregnancies. AB - OBJECTIVE: To determine if maternal serum placental growth factor (PlGF) concentration at 11-14 weeks in pregnancies complicated by diabetes, including those with preexisting diabetes and those that developed gestational diabetes subsequently, differed from that in normal, uncomplicated pregnancies. STUDY DESIGN: PlGF concentration was measured in stored maternal serum samples obtained at 11-14 weeks of gestation from 82 women with diabetic pregnancies, including 32 with preexisting diabetes and 50 with gestational diabetes, and 400 normal controls. Gestational diabetes mellitus was diagnosed with the 75-g oral glucose tolerance test and by applying World Health Organization criteria. RESULTS: When expressed as the multiple of the median and adjusted for gestation, the log transformed values of PlGF were significantly higher (P < .05) in the diabetic group (mean multiple of the median, [MoM] 1.14; SD 0.11) as compared to the controls (mean MoM, 1.00; SD, 0.11). On further comparison between the diabetic groups with the controls, significant differences were found in the groups with gestational diabetes (mean MoM, 1.15; SD, 0.12) and non-insulin dependent diabetes (mean MoM, 1.16; SD, 0.09) but not in those complicated by insulin dependent diabetes (mean MoM, 1.09; SD, 0.10). CONCLUSION: These findings suggest that in the first trimester of pregnancy the level of maternal PIGF was already significantly increased not only in pregnancies complicated by noninsulin dependent diabetes but also in those with gestational diabetes. The implications of these findings remain to be explored. PMID- 15283059 TI - Predictors of cervical dysplasia after the loop electrosurgical excision procedure in an inner-city population. AB - OBJECTIVE: To identify factors associated with persistence or clearance of cervical intraepithelial neoplasia (CIN) following loop electrosurgical excision procedure (LEEP) in high-risk patients. STUDY DESIGN: In a retrospective database review, we identified 343 patients who had 2 LEEP procedures or LEEP followed by hysterectomy for CIN at Grady Memorial Hospital. We compared margin status, endocervical curettage (ECC) at LEEP and follow-up cytology for patients characterized as having persistent or nonpersistent dysplasia. RESULTS: Seventy nine percent (71/90) of patients with positive LEEP margins had persistent disease vs. 50% (45/90) with negative margins (odds ratio [OR]=3.7, 95% confidence interval [CI] 1.9-7.2, P<.0001). Ninety-one percent (29/32) with positive margins and positive ECC had persistent disease vs. 47% (26/55) with negative margins and negative ECC (OR=10.8, 95% CI 2.9-39.6, P<.0001). Sixty eight percent (149/218) with at least 1 positive Pap smear following LEEP had persistent disease vs. 37% (11/30) with all negative follow-up Pap smears (OR = 3.7, 95% CI 1.7-8.3, P = .0007). CONCLUSION: Although the risk of persistent CIN increased with positive LEEP margins, ECC and cytology, these variables, when negative, offered no ensurance of a future disease-free state in this high-risk population. PMID- 15283060 TI - Altered IGF-I receptor expression in chorioamniotic cells in premature rupture of fetal membranes. AB - OBJECTIVE: To investigate the presence and distribution of type I insulinlike growth factor receptor (IGF-IR) in the cells of the chorioamniotic membrane and to search for any alterations occurring in IGF-IR expression in premature rupture of membranes (PROM) patients. STUDY DESIGN: Fetal membranes collected at delivery from 42 pregnancies between 36 and 40 gestational weeks were included in the study. Seventeen of 42 cases had premature rupture of membranes, and 25 cases had intact membranes prior to delivery. Paraffin sections of thefetal membranes were stained with IGF-IR antibody by the streptavidin-biotin-immunoperoxidase method. The staining was scored and compared statistically between PROM and control cases. RESULTS: The fetal membranes of PROM cases had significantly reduced IGF IR expression in chorionic trophoblastic cells when compared with the control group (P = .006, X2). CONCLUSION: Our immunohistochemical findings revealed that chanlges in IGF-IR levels in choriolzic amniotic cells may play a pathogenetic role in PROM cases, but the mechanism is speculative and needs further investigation. PMID- 15283061 TI - Polycystic ovary syndrome after menopause: a case report. AB - BACKGROUND: Polycystic ovary syndrome (PCOS) is a common endocrine disorder characterized by characteristic ovarian morphology on ultrasound, hyperandrogenism and chronic anovulation. Little is known about zhat happens to ovarian morphology or testosterone production in women with PCOS after menopause, and it is unclear how these women should be treated for the complaint of hirsutism. CASE: A 65-year-old woman presented with elevated testosterone levels, hirsutism and insulin resistance. Laparoscopic bilateral salpingo-oophorectomy (BSO) was performed, and following the procedure the testosterone levels normalized, although the ovaries were normal histologically. CONCLUSION: Laparoscopic BSO is a relatively safe, definitive treatment for postmenoapusal women with PCOS. PMID- 15283062 TI - Categorical perception depends on the discrimination task. AB - Speech sounds are said to be perceived categorically. This notion is usually operationalized as the extent to which discrimination of stimuli is predictable from phoneme classification of the same stimuli. In this article, vowel continua were presented to listeners in a four-interval discrimination task (2IFC with flankers, or 4I2AFC) and a classification task. The results showed that there was no indication of categorical perception at all, since observed discrimination was found not to be predictable from the classification data. Variation in design, such as different step sizes or longer interstimulus intervals, did not affect this outcome, but a 2IFC experiment (without flankers, or 2I2AFC) involving the same stimuli elicited the traditional categorical results. These results indicate that the four-interval task made it difficult for listeners to use phonetic information and, hence, that categorical perception may be a function of the type of task used for discrimination. PMID- 15283063 TI - Large errors, but no depth compression, in walked indications of exocentric extent. AB - Observers can sight a target 20 m away or more and then walk to it accurately without vision. In contrast to this good performance, this article shows that walked indications of the exocentric separation of two locations exceed the required values by over 70% when vision is obscured. Significantly, these large errors are coupled with a robust lack of depth foreshortening, even under conditions in which visual matches and verbal estimates of extent exhibit strong evidence of depth compression. This article presents evidence that the overshooting errors are due largely to recalibration of locomotor control produced by prolonged exposure to nonvisual walking. The robust lack of depth foreshortening, meanwhile, could reflect a corresponding isotropy in the spatial representation controlling the walking response. More research is needed to confirm this interpretation, however. PMID- 15283064 TI - On the concordance among empirical confusion matrices for visual and tactual letter recognition. AB - In this article, we examine the concordance among 19 empirical confusion matrices for visual and tactual recognition of capital letters of the alphabet. As a measure of concordance, we employed an index based on within-stimulus triads of letters. Unlike correlation measures of agreement that are based on a one-to-one matching of matrix elements, the selected index directly captures the internal structures of the confusion matrices prior to the comparison. Permutation tests revealed statistically significant concordance among 166 of 171 pairs of matrices in the study. Concordance of confusion structure among tactual matrices tended to be somewhat stronger than concordance among the visual matrices. PMID- 15283065 TI - Covert shifts of attention precede involuntary eye movements. AB - There is considerable evidence that covert visual attention precedes voluntary eye movements to an intended location. What happens to covert attention when an involuntary saccadic eye movement is made? In agreement with other researchers, we found that attention and voluntary eye movements are tightly coupled in such a way that attention always shifts to the intended location before the eyes begin to move. However, we found that when an involuntary eye movement is made, attention first precedes the eyes to the unintended location and then switches to the intended location, with the eyes following this pattern a short time later. These results support the notion that attention and saccade programming are tightly coupled. PMID- 15283066 TI - The sampling distributions of Gaussian ROC statistics. AB - For a discrimination experiment, a plot of the hit rate against the false-alarm rate--the ROC curve--summarizes performance across a range of confidence levels. In many content areas, ROCs are well described by a normal-distribution model and the z-transformed hit and false-alarm rates are approximately linearly related. We examined the sampling distributions of three parameters of this model when applied to a ratings procedure: the area under the ROC (Az), the normalized difference between the means of the underlying signal and noise distributions (da), and the slope of the ROC on z-coordinates (s). Statistical bias (the degree to which the mean of the sampling distribution differed from the true value) was trivial for Az, small but noticeable for da, and substantial for s. Variability of the sampling distributions decreased with the number of trials and was also affected by the number of response categories available to the participant and by the overall sensitivity level. Figures in the article and tables available on line can be used to construct confidence intervals around ROC statistics and to test statistical hypotheses. PMID- 15283067 TI - Imitation in shadowing words. AB - Imitation of shadowed words was evaluated using Goldinger's (1998) AXB paradigm. The first experiment was a replication of Goldinger's experiments with different tokens. Experiment 1's AXB tests showed that shadowed words were judged to be better imitations of target words than were baseline (read) counterparts more often than chance (.50). Order of presentation of baseline and shadowed words in the AXB test also significantly influenced judgments. Degree of prior exposure to token words did not significantly influence judgments of imitation. Experiment 2 employed modified target tokens with extended voice onset times (VOTs). In addition to AXB tests, VOTs of response tokens were compared across baseline and shadowing conditions. The AXB tests revealed shadowed words to be better imitations of target tokens than baseline, without an influence of AXB presentation order. Differences between baseline and shadowing VOTs were greater when VOTs were extended. The implications of spontaneous imitation in nonsocial settings are considered. PMID- 15283068 TI - Long-term memory for elementary visual percepts: memory psychophysics of context and acquisition effects. AB - In the first phase of each of two experiments, participants learned to associate a set of labels (i.e., consonant-vowel-consonant [CVC]) with a set of line lengths by using a paired-associate learning procedure. In the second phase of each experiment, these learned labels were used as memorial standards in the method of constant stimuli. Psychometric functions and the associated indices of discriminative performance (i.e., Weber fractions [WFs], just noticeable difference, and point of subjective equality) were then obtained for the remembered standards. In Experiment 1, WFs (i.e., the indices of memory precision) obtained with remembered standards were found to be higher (i.e., had poorer discriminability) than were WFs obtained with perceptual standards. In addition, WFs obtained with the remembered standards exhibited serial position effects (i.e., poorer discriminability for central items in the memory ensemble) and systematically varied with set size (i.e., the number of standards in the memory set), but WFs obtained with perceptual standards did not depend on serial position or set size. In Experiment 2, increasing the number of acquisition trials reduced WFs and diminished serial position effects. In addition, WFs did not vary systematically with the "physical" spacing between the standards in memory, but they did with the ordinal spacing. The results are consistent with a noisy analogue representation of remembered magnitudes, whereby central items in a memory ensemble are subject to lateral inhibition and thus reduced discriminability. Finally, presentation order effects, as defined by the classic time-order error, were observed with purely perceptual comparisons but not with comparisons involving a remembered standard. This latter finding is inconsistent with a strong form of the functional equivalence view of perception and memory. PMID- 15283069 TI - Perceptual grouping in change detection. AB - Detection of an item's changing of its location from one instance to another is typically unaffected by changes in the shape or color of contextual items. However, we demonstrate here that such location change detection is severely impaired if the elongated axes of contextual items change orientation, even though individual locations remain constant and even though the orientation was irrelevant to the task. Changing the orientations of the elongated stimuli altered the perceptual organization of the display, which had an important influence on change detection. In detecting location changes, subjects were unable to ignore changes in orientation unless additional, invariant grouping cues were provided or unless the items changing orientation could be actively ignored using feature-based attention (color cues). Our results suggest that some relational grouping cues are represented in change detection even when they are task irrelevant. PMID- 15283070 TI - What is learned in spatial contextual cuing--configuration or individual locations? AB - With the use of spatial contextual cuing, we tested whether subjects learned to associate target locations with overall configurations of distractors or with individual locations of distractors. In Experiment 1, subjects were trained on 36 visual search displays that contained 36 sets of distractor locations and 18 target locations. Each target location was paired with two sets of distractor locations on separate trials. After training, the subjects showed perfect transfer to recombined displays, which were created by combining half of one trained distractor set with half of another trained distractor set. This result suggests that individual distractor locations were sufficient to cue the target location. In Experiment 2, the subjects showed good transfer from trained displays to rescaled, displaced, and perceptually regrouped displays, suggesting that the relative locations among items were also learned. Thus, both individual target-distractor associations and configural associations are learned in contextual cuing. PMID- 15283071 TI - Are three-sample tasks less sensitive than two-sample tasks? Memory effects in the testing of taste discrimination. AB - In order to provide insights into why discrimination protocols with three stimuli sometimes tend to be less sensitive than protocols with two stimuli, two experiments were conducted. In these experiments, the relative effects of memory decay and memory interference were investigated. Both experiments involved purified water and/or solutions of low NaCl concentration. In Experiment 1, three protocols were compared: the traditional same-different test (Protocol 1), the same protocol with a rinse between the two samples (Protocol 2), and Protocol 2 with an added time delay between the first sample and the intermediate rinse (Protocol 3.) The decrease in measured d' values as time delay increased indicated that memory decay might be a factor for tests with three stimuli, such as the triangle method, rendering it less sensitive than tests with two stimuli, such as the same-different method. In Experiment 2, four protocols were compared: the traditional same-different test, the two-rinse same-different test, the triangle test, and what will be called duo same-different test. The experimental design allowed the individual consideration of memory decay and interference effects. From this last experiment, the substantial effect of memory interference was uncovered. Further experimentation will be necessary to estimate the exact relative effects of memory interference and memory decay. PMID- 15283072 TI - The Ebbinghaus illusion modulates visual search for size-defined targets: evidence for preattentive processing of apparent object size. AB - Five search experiments investigated whether the apparent size of objects is, like their retinal size, coded in preattentive vision. Observers searched for a target circle that was either larger or smaller than distractor circles, with both types of test circles surrounded by context circles modulating apparent size (i.e., the Ebbinghaus illusion). The size ratio between the test and the context circles was manipulated in such a way that the test circles were surrounded by, for example, smaller context circles (making the larger target appear even larger) or by larger context circles (making the smaller distractors appear even smaller). Under optimal conditions, the detection reaction times were independent of the number of test circles in the display, and the Ebbinghaus illusion facilitated the detection of the target even in comparison with control conditions without any context circles. This finding is consistent with preattentive, spatially parallel processing of apparent size. PMID- 15283073 TI - Gender and lexical access in Bulgarian. AB - Two procedures were used to explore the effects of semantic and grammatical gender on the recognition and processing of Bulgarian nouns, in relation to other factors that are known to affect lexical access. This study in a three-gender language was modeled on previous work in Italian, a two-gender language (Bates, Devescovi, Pizzamiglio, D'Amico, & Hernandez, 1995). Words were presented auditorily in randomized lists in two tasks: (1) repeat the word as quickly as possible and (2) determine the grammatical gender of the noun as soon as possible and indicate the decision by pressing a button. Reaction times in both tasks were influenced by phonological factors, word frequency, and irregularity of gender marking, but semantic and grammatical gender affected only gender monitoring. The significant contribution of semantic gender to processing in Bulgarian contrasts with previous findings for Italian. Also, we obtained an interaction between sex of the subject and noun gender, reflecting a bias toward one's own grammatical gender "counterpart" (especially for females). Reanalysis of the prior study in Italian showed a similar interaction but confirmed no effects of the semantic gender of the noun, suggesting that these two natural gender effects can dissociate. Possible reasons for cross-linguistic differences are discussed, with implications for comparative studies of gender and lexical access. PMID- 15283074 TI - The time course of attention in a simple auditory detection task. AB - What is the time course of human attention in a simple auditory detection task? To investigate this question, we determined the detectability of a 20-msec, 1000 Hz tone presented at expected and unexpected times. Twelve listeners who expected the tone to occur at a specific time after a 300-msec narrowband noise rarely detected signals presented 150-375 msec before or 100-200 msec after that expected time. The shape of this temporal-attention window depended on the expected presentation time of the tone and the temporal markers available in the trials. Further, though expecting the signal to occur in silence, listeners often detected signals presented at unexpected times during the noise. Combined with previous data, these results further clarify the listening strategy humans use when trying to detect an expected sound: Humans seem to listen specifically for that sound, while ignoring the background in which it is presented, around the time when the sound is expected to occur. PMID- 15283075 TI - Threshold estimation in two-alternative forced-choice (2AFC) tasks: the Spearman Karber method. AB - The Spearman-Karber method can be used to estimate the threshold value or difference limen in two-alternative forced-choice tasks. This method yields a simple estimator for the difference limen and its standard error, so that both can be calculated with a pocket calculator. In contrast to previous estimators, the present approach does not require any assumptions about the shape of the true underlying psychometric function. The performance of this new nonparametric estimator is compared with the standard technique of probit analysis. The Spearman-Karber method appears to be a valuable addition to the toolbox of psychophysical methods, because it is most accurate for estimating the mean (i.e., absolute and difference thresholds) and dispersion of the psychometric function, although it is not optimal for estimating percentile-based parameters of this function. PMID- 15283076 TI - Individual and relational conceptions of self in India and the United States. PMID- 15283077 TI - Self in learning among Chinese children. PMID- 15283078 TI - Multiplicity of ethnic identification during middle childhood: conceptual and methodological considerations. PMID- 15283079 TI - Within-culture complexities: multifaceted and interrelated autonomy and connectedness characteristics in late adolescent selves. PMID- 15283080 TI - The coactive construction of selves in cultures. PMID- 15283081 TI - Commentary: beyond individualism and collectivism--a problem, or progress? PMID- 15283082 TI - [Providing new knowledge]. PMID- 15283083 TI - [Problems of strain and coping of breast cancer patients--demands on nursing concepts of care]. AB - Over the last decades the number of breast cancer patients has dramatically increased. Attention has been drawn especially in research literature in the field of psychological medicine to the patients ' severe problems of strain and coping. In comparison to that, at least German intervention research in nursing science has to make up for lost time. This essay is, above all, a report on research literature. It summarizes study findings concerning emotional and physical problems associated with the disease and treatment for breast cancer. They result in a need for nursing care and support. Some practical conclusions for the nursing profession will be drawn in order to evolve comprehensive concepts of care both at a cognitive-emotional and tactile level of perception. PMID- 15283084 TI - [HIV/AIDS family care-giving in Switzerland in the context of silence]. AB - Based on critical hermeneutics, this qualitative study presents insights about care giving experiences of HIV affected families in the German speaking part in Switzerland. Eleven families (12 women, 5 men) were interviewed, and members of different families participated in group-conversations. Critical reflection of the results highlighted that these HIV-families offered family caregiving in a context of silence. Two perspectives of this silence are presented: 1. HIV families do not share their experiences in their closer and larger environments because of stigmatization, changing family constellations, and traditional images of family care. Instead, HIV-families create silent circles. 2. Professional discourses about family care are almost absent in the Swiss health care system because of its focus on the needs of individual patients with health care providers, in particular nurses, offering no systematic health services tailored to family caregivers. The results highlight, that it is paramount for nurses to take responsibility and to create programs that will better serve HIV-families. PMID- 15283085 TI - [Patient records: supporting interprofessional communication in hospital]. AB - Complete and continuous documentation in patient records is an important condition for adequate communication with patients, between the professions concerned and to ensure the quality of the following working steps in care provision. Part of a German research project concerning the interprofessional communication in hospital was therefore to analyse the use of the documentation system. 54 users were asked about practical aspects of their documentation system and 450 patient records were evaluated. The analysis focused on the medical and nursing documentation of admission, process and discharge. Deficits that need to be improved appeared first of all in the practical aspects of the documentation system, the flow of information between the professions, in specific gaps of medical and nursing admission, documentation of process and discharge. Quality management is asked to improve and develop the documentation in collaboration with the users and to consider specific problems when introducing computer based records. PMID- 15283086 TI - [How do nurses experience and describe quality of care in everyday nursing]. AB - This study examines how nurses experience and describe quality of care in a hospital. Applying a qualitative approach, semi structured interviews were conducted with eight nurses. The data were analysed using a seven phase process of analysis outlined by Colaizzi (1978). Fifteen clusters of topics were found which describe the dimensions of care as seen by the nurses. These clusters of topics could be further segmented into three main topics which are identical with the three aspects of structure, process and outcome quality as suggested by Donabedian (1966). The requirement criteria of the structure and process quality were experienced either as facilitating or inhibiting. The beliefs and the experience of the quality of care in everyday nursing practice were described in a subjective but comprehensive manner The quality facilitating factors quoted most frequently are professional knowledge and the perception of the process of caring. The limited personnel resources due to the vacant positions were experienced as the most inhibiting factor The outcome requirements were mostly related to the issue of satisfaction/dissatisfaction of nurses and of patients and to the issue of examining the quality of nursing. We were able to formulate some recommendations for nursing practice as well as for further studies. PMID- 15283087 TI - [Nursing management in conflict between ethics and economy: a qualitative study in institutional and semi community care of older people]. AB - The starting point of this explorative study is the current problematic development in institutional and semi community care of older people. The relationship between economy and ethics is a permanent tug of war for nursing managers, who have to fulfil not only the needs of inhabitants and professional standards, but also the restrictive financial framework. This study was based on the following questions: Which are the relevant values for decisions of nursing managers and which are important ethical principles to direct their behaviour? Which dilemmas are significant? The examination group consisted of eleven nursing managers from the field of institutional and semi community care. Qualitative, problem-based interviews were taken with them. The results show the high sensibility of this group in context with ethical questions, which express the following principles: The principle of untouchable dignity, the principle of caring, the principle of assurance of the inhabitants' economic rights and the principle of openness and transparency. The significant dilemmas concern business management, employees, medical service of health insurance companies and doctors. Initial stages of an integrative business-ethical approach could be found in the majority of care centres. By contrast, a classical division into economy and ethics was found in the other care centres. PMID- 15283088 TI - [Editorial]. PMID- 15283089 TI - [Health care between art and science--experience and problems of professional jargon in nursing and health care]. AB - As health care increasingly focuses its emphasis on the standards of science (objectivization, formulation, standardization and quantification of practical knowledge specific to this profession), the question, whether essential parts of professional experience are adequately included in this process or are rather an implicit part of it, has to be brought up. This article compares the structures of rational propositional knowledge (being the dominant approach within the medical conception of the world) with practical knowledge of health care on an epistemological level. It is further suggested to look for linguistic access to practical knowledge within health care not only via a rational technical terminology--which does not include essential parts of this knowledge--, but also via an aesthetically structured language. Literary texts as to their aesthetic use of language, on the other hand, offer a way to reflect and impart tacit knowledge as well as the opportunity to reflect ethically on beliefs, which are- mostly unconsciously--the driving force behind medical and nursing action. PMID- 15283090 TI - ["However we are different"--research of nursing experience at migrants on the background of a dynamic culture understanding]. AB - At the beginning of this study was the assumption that cultural differences have hardly been taken into consideration in Germany. How do people from a different cultural background experience nursing in German hospitals? The actual relationship with some group of patients from another cultural background in the hospital ward is to be described vividly and properly. Therefore this study is following the applied phenomenology that nursing experience should be examined by problem oriented interviews. The results should be discussed in the context of present-day literature on science of nursing. The dominating topic of the interviews is "being different"-"being foreign". Perception and acceptance of foreign understanding of reality is the starting point for culturally adapted nursing. Professional nursing means individually planned nursing, considering the given context and environment. Therefore professional nurses have to grasp and conduct according to the individual experience of reality of patients. PMID- 15283091 TI - [An encounter with foreign parts is not the objective of the journey. The reality of the return]. AB - It is not only the experience of the "outside", the self-chosen possibility of disarrangement, of moving away from previously-binding sensory structures that determines the transcultural experience. It depends much more on a second step, of returning to oneself-- having returned from foreign parts, to re-integrate oneself into one's own culture. This means that the journey itself into foreign areas, the encounter with medical and nursing systems of other cultures, is in fact only half the journey; the complete journey is one which returns back into one's own (nursing) culture. Only when the nurse has succeeded--after his/her return--in newly regarding his/her own nursing activity--that is, with different eyes--in an expanded context and in recognizing the resulting changes, has he/she reached the real objective of his/her journey. PMID- 15283092 TI - [Quality of life of people with schizophrenia in the supported group homes: assessing the individual]. AB - The study assessed the individual quality of life of people with schizophrenia who live in the supported group homes. Forty residents in seven group homes were interviewed using The Schedule for the Evaluation of Individual Quality of Life: A Direct Weighting Procedure Instrument (SEIQoL-DW). The SEIQoL-DW assesses the individualised QoL using semi-structured interview technique. The definition underpinning the SEIQoL-DW is that a person's QoL is what he or she determines it to be. The respondents defined the areas that are important to them, rated their current situation on each LQ-area as well as the relative importance of each area by using visual analogue scale (VAS 0-100). The mean QoL score was 64.71 (SD 19.05, range 6.81-93.49). A total of 18 QoL areas were formed of 200 individually nominated cues. The areas most frequently nominated by respondents were "human relations", "social life", "work", "leisure time", "finances and other material desires", "quality of living space", "autonomy" and "health". It showed clearly that nominated QoL-areas had for each respondent individual meaning as well as individual importance in his or her life. A positive correlation was found between length of stay in supported home and global quality of life. In addition, the respondents who had lived in supported home for a longer period rated their current situation higher with respect to autonomy, work and finding the meaning of life than respondents with shorter duration of stay. The individual nature of QoL was reflected in differences in QoL-areas defined, differences in levels of satisfaction, and differences in the relative importance of each LQ-area to individuals. The measures were generally acceptable, with all respondents showing great interest and being able of completing the SEIQoL-DW. This study showed also that instruments such as the SEIQoL-DW might have a therapeutic application for people with psychiatric disorders. PMID- 15283093 TI - [Effects of health political measures on health and well-being of women postpartum and their newborns]. AB - For decades politicians have tried to reduce or stabilize the costs of health supply in Germany. The shortening of the average lengths of stay in hospitals seems to be one of the most popular measures. This development can be seen above all in the field of obstetric care. In the last 15 years the average stay of women after birth in hospitals decreased from 5.6 days to 3.8 days in parts of northern Germany. Based on the criticism that through the implemenetation of new health policies the formulation of qualitative and patient- and population oriented aims is disregarded this article deals with the question of the effects of the shortening of hospital stay on health and well-being of women postpartum and their newborns. It is mainly based on a critical analysis of international studies. PMID- 15283094 TI - [The care dependency scale, an assessment instrument]. AB - The Care Dependency Scale, an instrument for the assessment of patients'care dependency, has been translated into German. The scale was tested on (inter-) rater reliability and criterion and construct validity in a hospital population on geriatric, surgical and paediatric wards. As the results of this study were very satisfying, positive recommendations regarding the suitability of the scale for use in the German nursing care situation could be made. However further psychometric testing of the scale is important, for instance in other populations. A final conclusion is that the scale may be used in care settings in German-speaking countries. PMID- 15283095 TI - The impact of advances in developmental biology on the management of neonatal surgical anomalies. AB - While advances in the clinical management of various congenital anomalies in pediatric surgery have led to new and exciting therapeutic modalities, our understanding of the mechanisms responsible for these defects lags far behind. In a new era of developmental biology, the prospect of unlocking some of these mysteries has become a real possibility. Advances in gene sequencing has allowed us to create new phenotypes that closely mimic those seen in patients, and has created a setting where we are now better able to understand and develop new therapeutic interventions. Here we discuss the implications of some of the molecular mechanisms underlying various congenital anomalies encountered in pediatric surgery, and how continued research will impact the future of these disease processes. PMID- 15283096 TI - Potential tissue-engineering applications for neonatal surgery. AB - Tissue engineering attempts to build neotissue from its cellular building blocks. This neotissue can then be used for reconstructive surgical applications such as replacement of a congenitally abnormal heart valve or repair of a craniofacial abnormality. Since its inception in the late 1980s, tissue engineering has sparked the interests of physicians and scientists alike because of its great potential. Significant progress has been made in this burgeoning branch of science. This article reviews some of the ongoing preclinical and clinical tissue engineering research as it applies to neonatology. PMID- 15283097 TI - The CDH Study Group and advances in the clinical care of the patient with congenital diaphragmatic hernia. AB - Congenital diaphragmatic hernia (CDH) occurs in 1 of every 2000 to 4000 births and accounts for 8% of all major congenital anomalies. Recurrence risk for a subsequent pregnancy is estimated at 2%. The mortality rate for CDH when diagnosed antenatally, varies with fetal age and with the presence or absence of hydramnios and degree of pulmonary hypoplasia. The prognosis has improved dramatically in recent years, primarily due to advances in neonatal and surgical interventions. Neonatal survival rates with an antenatal diagnosis now exceed 80% in some centers. Treatment for infants with CDH reflects other pediatric surgical problems in that a majority of the clinical research that shapes treatment is retrospective in nature. Because CDH is a relatively rare disease, using a compilation of cases, such as the CDH database provides, greatly aids our understanding of this disease process. Moreover, the application of a quality assessment scale provides the practitioner with a knowledge base to critically evaluate the published retrospective data. PMID- 15283098 TI - New approaches to managing congenital diaphragmatic hernia. AB - A number of new techniques have been studied for managing newborns with congenital diaphragmatic hernia and respiratory insufficiency. Among these have been the techniques of delayed approach to the repair of the diaphragmatic hernia; permissive hypercapnia; nitric oxide and surfactant administration; intratracheal pulmonary ventilation; liquid ventilation; perfluorocarbon-induced lung growth; and lung transplantation. These interventions are at various stages of development and evaluation of effectiveness. All, however, are being explored in the hopes of improving outcome in patients with congenital diaphragmatic hernia who continue to have significant morbidity and mortality in the newborn period. PMID- 15283099 TI - Recent advances in fetal surgery. AB - Fetal surgery is now an accepted modality for treatment of a variety of lethal and non-lethal congenital conditions. It represents a new, fast-moving frontier of medicine in which cooperative mulitdisciplinary effort and input are required to assure both fetal and maternal welfare. A wide range of therapeutic strategies from percutaneous to open invasive techniques has led to a complex list of different procedures for different diseases. This review identifies the most common disease entities managed by fetal intervention, examines the evolution in development of techniques to those currently used, and describes the prospective, randomized trials presently underway that are designed to establish the safety and determine true efficacy of treatment. Fetal surgery as a (multi)discipline continues to strive to minimize maternal and fetal risk. Undoubtedly, as tocolytic therapy and neonatal intensive efforts improve, fetal therapy will expand. PMID- 15283100 TI - Minimally invasive surgery in the neonate: review of current evidence. AB - Minimally invasive surgery has been one of the most important surgical advances in the last 15 years. The development of smaller instruments has allowed pediatric surgeons to apply this rapidly evolving technology to neonates. Congenital neonatal deformities including tracheoesophageal fistula, patent ductus arteriosus, duodenal atresia and anorectal malformations are now being managed with minimally invasive surgery. This article summarizes the status of these techniques in neonates. PMID- 15283101 TI - Current issues in the management of necrotizing enterocolitis. AB - Necrotizing enterocolitis (NEC) is the most common surgical emergency in the neonatal intensive care unit and remains a major cause of death in neonates. Although the pathophysiology of NEC has not been completely elucidated, progress has been made in the characterization of the molecular events which may take place during an episode of ischemia. This possible initiating event is followed by a complex cascade of inflammatory mediators active in NEC: epidermal growth factor, platelet-activating factor, and, nitric oxide. Additionally, unique characteristics of the premature gut are thought to be crucial to the development of NEC. The diagnosis of NEC continues to be based on clinical and radiographic features. Several new laboratory tests are under investigation for the purposes of earlier diagnosis, but none have prevailed at this time. Both exploratory laparotomy, with intestinal resection and peritoneal drainage are widely practiced. Mortality rates remain high and have improved little over the last couple of decades. Therefore, prevention remains crucial in order to decrease the incidence of NEC. Cautious feeding regimens, the use of maternal breast milk, passive immunization, and the use of probiotics have all been suggested but not proven as possible preventive methods. Although many advances have been made, significant opportunity remains to improve our understanding of the disease process and to develop better strategies for prevention and treatment. PMID- 15283102 TI - Operating on critically ill neonates: the OR or the NICU. AB - Advances in neonatal care have resulted in the survival of smaller infants with more complicated medical problems. From a surgical standpoint this has required novel approaches to patient care. Surgical care has evolved in many respects. Procedures performed on premature infants range from elective, minor procedures to major, emergent lifesaving interventions. The emergent nature of these surgical interventions has led to controversies in management. Certain conditions require surgical procedures that are commonly performed at the bedside by pediatric surgical specialists. Under other circumstances, the specific details of management are less uniform with wide variability in approach by different practitioners. The rationale in these cases is primarily driven by personal preference with a paucity of supportive data in the published literature to either support or contradict individual opinion. Nevertheless, the role of bedside procedures appears to be expanding. If these procedures are to be undertaken, significant planning is required to ensure a good outcome for the patient. Prospective data are needed determine which patients may benefit from this approach. PMID- 15283103 TI - Ethical issues in the management of neonatal surgical anomalies. AB - This article provides a framework for thinking about three areas in neonatal surgery that contain potential moral and ethical concerns for pediatric surgeons and the parents of a newborn and/or fetus with a surgical anomaly. The utilization of life-sustaining therapy for neonates has made survival possible for many infants with serious birth defects. Sometimes the use of these treatments is problematic in terms of their actual benefit to the infant and the potential for enhancing their future quality of life. Second, the prenatal diagnosis of congential anomalies has made counseling of the prospective parents a routine part of pediatric surgical practice and raises the issue of how best to advise and support a couple whose fetus has a significant birth defect. Finally, pediatric surgeons have a responsibility to their patients and society to provide the highest quality of care. This may involve participation in multi institutional clinical trials, so that the optimal care of a surgical neonate with a congenital or acquired condition is ascertained by rigorous prospective research evaluation. PMID- 15283104 TI - [Also in Denmark are cultural and social conditions of significance when it comes to the extent of violence against women]. PMID- 15283105 TI - [Chronic subdural hematoma--more complicated than expected]. PMID- 15283106 TI - [Chronic subdural hematoma. Physiopathology and treatment]. PMID- 15283107 TI - [Lyme borreliosis--the most frequent vector-borne infection in Denmark]. PMID- 15283108 TI - [Should chemotherapy or endocrine therapy be used as first choice in patients with advanced breast cancer? A status article based on the Cochrane analysis "Chemotherapy alone versus endocrine therapy alone for metastatic breast cancer"]. PMID- 15283109 TI - [Violence against Danish and immigrant women in Aarhus]. PMID- 15283110 TI - [Infectious endocarditis in the county of Frederiksborg, 1990-2000. Clinical findings and prognostic aspects]. PMID- 15283111 TI - [Incidence and diagnosis of infectious endocarditis in Frederiksborg county, 1990 2000]. PMID- 15283112 TI - [Ocular emergencies referred to the duty ophthalmologist]. PMID- 15283113 TI - [Cancer patients' experiences with unconventional treatment]. PMID- 15283114 TI - [Dietary fibers in food and protection against colorectal cancer in the European Prospective Investigation into Cancer and Nutrition (EPIC) study]. PMID- 15283115 TI - [Depression and cancer risk. A registry-based study in Denmark, 1969-1993]. PMID- 15283116 TI - [Rare manifestation of embryonal rhabdomyosarcoma]. PMID- 15283117 TI - [Emphysema: a high-resolution CT diagnosis]. PMID- 15283118 TI - [Picture of the month: orbital myeloma]. PMID- 15283119 TI - [Medical error!]. PMID- 15283120 TI - [Musculoskeletal diseases]. PMID- 15283121 TI - [Fatal outcome of pneumococcal pneumonia and following conclusions]. PMID- 15283122 TI - [Physicians, patients and newspapers]. PMID- 15283123 TI - beta-xylosidase activity and expression of a beta-xylosidase gene during strawberry fruit ripening. AB - Strawberry fruit shows a marked softening during ripening and the process is associated with an increment of pectin solubility and a reduction of the molecular mass of hemicelluloses. In this work, we report the activity of beta xylosidase and the expression of a beta-xylosidase gene in strawberry fruit. We have cloned a cDNA fragment encoding a putative beta-xylosidase (FaXyl1) from a cDNA library obtained from ripe strawberry fruit. The analysis of the deduced amino acid sequence revealed that FaXyl1 is closely related to other beta xylosidases from higher plants. The expression of FaXyl1 was strongly associated to the receptacle tissue although a low expression level was detected in achenes and ovaries. The accumulation of FaXyl1 mRNA is ripening-related, starting in white fruit, reaching the maximum at 25-50% red fruit and decreasing thereafter. The total beta-xylosidase enzyme activity was detected in all ripening stages with the maximum in 25-50% red fruit. The low activity level detected in immature stages, where no expression of FaXyl1 was found, suggests the presence of other beta-xylosidases-like genes. Both the expression of FcaXyl1 and the total beta xylosidase activity were down regulated by auxins, as occurs for most of the ripening-related processes in strawberry fruit. A putative role of FaXyl1 and beta-xylosidase is discussed. PMID- 15283124 TI - L-Myo-inositol 1-phosphate synthase in the aquatic fern Azolla filiculoides. AB - L-Myo-inositol 1-phosphate synthase (INPS EC 5.5.1.4) catalyzes the conversion of D-glucose 6-phosphate to L-myo-inositol 1-phosphate. INPS is a key enzyme involved in the biosynthesis of phytate which is a common form of stored phosphates in higher plants. The present study monitored the increase of INPS expression in Azolla filiculoides resulting from exposure to inorganic phosphates, metals and salt stress. The expression of INPS was significantly higher in Azolla plants that were grown in rich mineral growth medium than those maintained on nutritional growth medium. The expression of INPS protein and corresponding mRNA increased in plants cultured in minimal nutritional growth medium when phosphate or Zn2+, Cd2+ and NaCl were added to the growth medium. When employing rich mineral growth medium, INPS protein content increased with the addition of Zn2+, but decreased in the presence of Cd2+ and NaCl. These results indicated that accumulation of phytate in Azolla is a result of the intensified expression of INPS protein and mRNA, and its regulation may be primarily derived by the uptake of inorganic phosphate, and Zn2+, Cd2+ or NaCl. PMID- 15283125 TI - The catecholamine potentiates starch mobilization in transgenic potato tubers. AB - In human and animal cells, the catecholamines are involved in glycogen mobilization. Since the compounds are found in a potato, their function in starch mobilization was hypothesized. In order to verify this hypothesis, the transgenic potato plants Solanum tuberosum L. cv. Desiree overexpressing tyrosine decarboxylase (TD EC 4.1.1.25) cDNA from parsley has been generated. The cDNA expression was judged by the northern blot analysis and the enzyme activity measurements. Four independent transgenic lines with the highest TD mRNA expression were selected and analyzed. The expected substantial decrease in tyrosine content was followed by significant increase in tyramine and dramatic enhancement of norepinephrine synthesis was detected. The level of L-3,4 dihydroxyphenylalanin (L-Dopa) was only slightly increased and dopamine significantly decreased in most cases in these plants. The increase in norepinephrine was accompanied by changes in carbohydrate metabolism. The significant increase in glucose and sucrose and the decrease in starch content were characteristic features of TD overexpressed transgenic potato tubers. The features mentioned above indicate that catecholamines potentiate starch mobilization in potato plants in common with animal cells. The decrease in tyrosine content in transgenic plants is also compensated by significant increase in chlorogenic acid synthesis thus potentially increasing the antioxidant capacity of transgenic tubers. The glycoalkaloids content is changed in the transformants. This may originate from glucose accumulation and glycolysis activation. The obtained transgenic potato provides material for further detailed studies of the physiological function of catecholamines in plants. PMID- 15283127 TI - Limitation of photosynthetic carbon metabolism by dark chilling in temperate and tropical soybean genotypes. AB - In the experiments reported in this paper, we characterised the physiological and biochemical factors involved in the chilling-induced inhibition of photosynthetic carbon metabolism in soybean [Glycine max (L.) Merr.] genotypes of temperate and tropical adaptation. Plants of Maple Arrow (temperate genotype) and Java 29 (tropical genotype) were exposed to a single night at 8 degrees C. Dark chilling resulted in the inhibition of diurnal CO2 assimilation rate and decreased stomatal conductance in both genotypes. Further analysis, however, revealed a difference in the response of the two genotypes. Stomatal limitation was largely responsible for the inhibition of CO2 assimilation in Maple Arrow, whereas mesophyll limitation dominated the inhibition in Java 29. The results indicate that inhibition of stromal fructose-1,6-bisphosphatase (sFBPase; EC 3.1.3.11) activity and impaired electron transport capacity were responsible for the decrease in ribulose-1,5-bisphosphate (RuBP) regeneration capacity in Java 29. Sucrose-phosphate synthase (SPS; EC 2.4.1.14) activity was progressively inhibited during the light period in this genotype and might impose an additional constraint on photosynthesis. Maple Arrow appears to possess, at least with respect to photosynthetic carbon metabolism, physiological and biochemical characteristics that contribute towards its superior dark chilling tolerance. PMID- 15283126 TI - SNF1-related protein kinase (snRK1) phosphorylates class I heat shock protein. AB - The nucleotide sequence of cBSnIP2, a cDNA that had been cloned from a barley (Hordeum vulgare) seed endosperm cDNA library by two-hybrid screening with barley SNF1-related protein kinase (SnRKI) was determined. It was found to contain a complete open reading frame encoding a class I heat shock protein. Transcripts corresponding to the cDNA (renamed cBHSP17) were detectable in RNA isolated from barley seeds harvested in mid-development but not RNA from roots or leaves. BHSP17 protein was expressed in Escherischia coli and shown to be phosphorylated by SnRKI from barley endosperm and spinach leaf. It was found to be a less effective substrate than 3-hydroxy-3-methylglutaryl-Coenzyme A, a previously identified substrate of SnRK1. However, a specific phosphorylation site at serine 35 was identified by solid phase sequencing of RP-HPLC-purified peptides after phosphorylation by spinach SnRK1. PMID- 15283128 TI - Light-responsive subtilisin-related protease in soybean seedling leaves. AB - Protease C1 (E.C. 3.4.21.25), the soybean (Glycine max L. Merrill) proteolytic enzyme responsible for initiating the degradation of soybean storage proteins in seedling cotyledons appears at even higher levels in seedling leaves. This was manifested at the mRNA level through northern blot analysis, at the protein level through western blot analysis, through determination of enzyme activity, and also through isolation and partial sequencing of active leaf enzyme. Comparison of cDNA and amino acid sequences, as well as characterization of enzyme activity, is consistent with the leaf enzyme being identical to or highly similar to the cotyledon enzyme. Protease C1 mRNA and protein are also present in stems of soybean seedlings, but is very low to absent in the roots. This presence in the aerial tissues is consistent with the higher steady state level of gene expression at both the mRNA and protein levels when the seedlings are grown in a 12-h light: 12-h dark photoperiod as compared to seedlings grown in continuous darkness. Transfer of dark-grown seedlings to light is followed by marked elevation in protease C1 protein as seen in western blots. PMID- 15283129 TI - Six genes strongly regulated by mercury in Pisum sativum roots. AB - Suppression subtractive hybridisation was used to isolate heavy metal-induced genes from Pisum sativum roots hydroponically exposed to 5 microM HgCl2 and 10 microM EDTA. Six genes were induced out of which one, PsHMIP6B, was novel. The other genes (PsSAMT, PsI2'H, PsNDA, PsAPSR, PsPOD) had not previously been isolated from pea and sequenced. All six genes were also induced after exposure to 5 microM HgCl2 in the absence of EDTA. The induction pattern was in some cases different for the two Hg species, demonstrating a quicker response to-free Hg2+ than Hg-EDTA. The stress-specificity of the gene regulation was investigated by hydroponically adding 5 microM Cd2+. Most Hg-induced cDNAs were also induced by Cd2+ but to a smaller extent than after Hg exposure. In addition, the gene expression was also probed for tissue specificity, which showed that all six genes were expressed in roots and not in leaves. PMID- 15283130 TI - Copper deficiency induced expression of Fe-superoxide dismutase gene in Matteuccia struthiopteris. AB - Iron-superoxide dismutase (Fe-SOD) activity was not detected in extracts from the leaves of ferns, Equisetum arvense and Matteuccia struthiopteris. To know why ferns lack Fe-SOD activity, the Fe-SOD like gene (MsFeSOD1) was isolated from M. struthiopteris and its expression was investigated with a focus on the metals Fe and Cu using the prothalli of the fern. The expression of MsFeSOD1 mRNA was induced by a deficiency of Cu, but Fe-SOD activity was not detected. The recombinant protein of MsFeSOD1 produced in E. coli showed Fe-SOD activity. These findings suggest that the fern Fe-SOD like gene was transcriptionally regulated by Cu but an additional mechanism is involved in the formation of an active enzyme. PMID- 15283131 TI - Cytological changes and alterations in polyamine contents induced by cadmium in tobacco BY-2 cells. AB - Changes in cell viability, proliferation, cell and nuclear morphology including nuclear and DNA fragmentation induced by 0.05 and 1 mM CdSO4 (Cd2+) in tobacco BY 2 cell line (Nicotiana tabacum L.) were studied in the course of 7 days. Simultaneously changes in endogenous contents of both free and conjugated forms of polyamines (PAs) were investigated for 3 days. The application of 0.05 mM Cd2+ evoked decline of cell viability to approximately 60% during the first 24 h of treatment. Later on degradation of cytoplasmic strands, formation of the stress granules and vesicles, modifications in size and shape of the nuclei, including their fragmentation, were observed in the surviving cells. Their proliferation was blocked and cells elongated. Beginning the first day of treatment TUNEL positive nuclei were detected in cells cultivated in medium containing 0.05 mM Cd2+. Treatment with highly toxic 1 mM Cd2+ induced fast decrease of cell viability (no viable cells remained after 6-h treatment) and cell death occurred before DNA cleavage might be initiated. The exposure of tobacco BY-2 cells to 0.05 mM Cd2+ resulted in a marked accumulation of total PAs (represented by the sum of free PAs and their perchloric acid (PCA)-soluble and PCA-insoluble conjugates) during 3-day treatment. The increase in total PA contents was primarily caused by the increase in putrescine (Put) concentration. The accumulation of free spermidine (Spd) and spermine (Spm) at 12 and 24 h in 0.05 mM Cd2+ treated BY-2 cells and high contents of Spd and especially Spm determined in dead cells after I mM Cd2+ application was observed. The participation of PA conjugation with hydroxycinnamic acids and PA oxidative deamination in maintaining of free PA levels in BY-2 cells under Cd2+-induced oxidative stress is discussed. PMID- 15283132 TI - Characterization and evolutionary relationship of methionine-rich legumin-like protein from buckwheat. AB - We have isolated and characterized a full-length cDNA for legumin-like storage polypeptide from buckwheat seed (Fagopyrum esculentum Moench) and compared its deduced amino acid sequence with those from different representatives of dicots, monocots and gymnosperms. The cDNA sequence was reconstructed from two overlapping clones isolated from a cDNA library made on mRNA of buckwheat seed at the mid-maturation stage of development. Analysis of the deduced amino acid sequence revealed that this specific buckwheat storage polypeptide should be classified in the methionine-rich legumin subfamily present in the lower angiosperm clades, a representative of which was first characterized in Magnolia salicifolia (clone B 14). The fact that a methionine-rich legumin coexists together with methionine-poor legumins in buckwheat should be an important element regarding the evolutionary position of buckwheat. This may also be supporting evidence that the B14 ortholog was not lost in evolution but was protected under pressure of an increased need for sulfur. Using primers designed from characterized cDNA, we also isolated its corresponding gene from buckwheat genomic DNA and analyzed the characteristic exon/intron structure. The firstly identified two-intron structure of buckwheat legumin gene is an important contribution to study of methionine-rich legumins in lower angiosperms. PMID- 15283133 TI - Increased production of beta-1,3 glucanase and proteins in Bipolaris sorokiniana pathosystem treated using commercial xanthan gum. AB - Barley plants (cultivars Embrapa 127, 128 and 129) treated with xanthan gum, and with different time intervals between the administration of the inducer and the pathogen. demonstrated induction of resistance against Bipolaris sorokiniana. Induction was shown to have local and systemic action. In order to prove the resistance effect, biochemical analyses were performed to quantify proteins and the enzymatic activity of beta-1,3 glucanase. Results demonstrated that barley plants treated with the inducer, showed an increase in the concentration of proteins, as well as in the activity of the enzyme beta-1,3 glucanase, when compared with the extract from healthy plants. In infected plants, protein concentrations decreased and enzymatic activity was lower than in healthy plants. Results suggest that barley plants treated with xanthan gum developed mechanisms responsible for induced resistance, which are still unknown. The most important macromolecule in the defense mechanism was demonstrated to be PR-protein, due to its accumulation and concentration of proteins. However, it may not be the only macromolecule responsible for the resistance effect. PMID- 15283134 TI - [Target molecules of molecular target therapy of cancer]. PMID- 15283135 TI - [Molecular targeting drugs--present status and future development]. AB - Development of molecular targeting drugs is a recent highlight in cancer therapeutic field. One can look for 'drugable' target(s) from many molecular targets specific in malignant characteristics of human cancers. Drugs targeting various malignancy-linked molecules such as EGF receptor and its family proteins. Bcr-abl, CD20, Ras and others are now approved or under clinical trials against cancer patients. These molecular targeting drugs will provide a novel and useful therapeutic strategy, but, at the same time, we have many problems to overcome. We should continue our further efforts to answer following problems: (1) How therapeutic efficacy of molecular targeting drugs could be determined in patients in evidence-based manner?; (2) What is promising molecular target for development of drug?; (3) How combination therapy of molecular targeting drug with other cytotoxic drugs should be designed? PMID- 15283136 TI - [Targeting molecular mechanisms for cancer therapy]. AB - Cancer development is considered by the dysregulated proliferation of transformed cells. In recent years, dramatic and remarkable insight into the molecular mechanisms of this phenomenon have come from basic cancer research. Detailed knowledge of the molecular mechanisms and pathophysiology of cancer allows therapeutic agents to be designed that specifically target aberrant cellular processes. There are currently a number of targeted treatment strategies with less toxicity being evaluated. In the future, it is likely that this mechanistic and targeted approach will have a significant impact on clinical oncology and the diagnostic evaluation of tumors. PMID- 15283137 TI - [International estimation of molecular target drug]. AB - Development of tumor molecular biology makes a lot of molecular target drugs. Now molecular target drugs get constant estimation, and development competition is globally hot. Even if they have promising data in preclinical studies, it is very difficult to prove survival benefits in phase III studies. We have to understand a characteristic of molecular target drug based on Translational Research. Translational Research is an extremely important breakthrough to molecular target drug development/clinical application. PMID- 15283138 TI - [Clinical evaluation of molecular-target based drug]. PMID- 15283139 TI - [TGF-beta signaling pathway in pancreatic cancer cells]. AB - TGF-beta is a multifunctional cytokine which regulates cell growth, extracellular matrix deposition, cell differentiation, and immunosuppression etc. The signal is mainly mediated through Smad pathway to inhibit epithelial cell growth. In 50% of pancreatic cancer, Smad4/DPC4 gene is deleted or mutated, which might cause pancreatic carcinogenesis and be associated with highly invasive and metastatic character of the disease. On the other hand, TGF-beta signal itself has recently been shown to act in favor of cancer cells, especially in the late phase of tumor progression. In Smad4-inactivated pancreatic cancer cells, TGF-beta signal regulates a number of genes involved in tumor suppression and progression. The regulated genes and signaling pathways of TGF-beta signal should be investigated to obtain an effective therapeutic target molecule for pancreatic cancer. PMID- 15283141 TI - [Novel angiogenesis inhibitors for molecular target therapy of cancer]. AB - Angiogenesis, neovascularization from pre-existing vasculature, is essential to allow growth of primary solid tumors and to enable metastasis. Recent biological studies in both tumor and endothelial cells have begun to present a wide variety of molecular targets for developing angiogenesis inhibitors. Therefore, angiogenesis inhibitors including anti-angiogenic agents as well as anti-vascular targeting agents have become promising drugs in cancer chemotherapy. However current unsolved problems in anti-angiogenic therapy are the lack of surrogate markers for therapeutic efficacy, as well as of establishment of effective combinations with other therapeutic approaches including conventional anticancer therapy, radiotherapy, and immunotherapy. This article focuses on the promising drugs with anti-angiogenic activity and their molecular targets under clinical trials, as well as the significance of clinical evaluation for anti-angiogenic therapies. PMID- 15283140 TI - [Novel molecularly targeted peptides interfering the specific signal transduction of cancer metastasis]. AB - Metastatic spread of tumor cells is one of the major risk factors affecting clinical prognosis. There are several sequential steps in this process including cell migration into the surrounding host tissue, infiltration and penetration into vessels for dissemination, attachment to capillary beds of distant organs, cell extravasation and subsequent growth in tissues. Some of the mechanisms involving normal cell migration observed during development are believed to be closely associated with the properties of tumor cell invasion. Thus, the so called 'cell migration genes' may be frequently utilized by cancer cells to allow for metastatic spread of disease. We have isolated one of the candidate genes homologous to a C. elegans cell migration gene, that overexpressed specially in clinical cancer tissues. Based on in vitro analysis of the signal transduction, a novel peptide interfering the signals provides an attractive target for therapeutic intervention against the cancer invasion and metastasis. PMID- 15283142 TI - [Development of novel angiogenesis inhibitors targeting VEGF (vascular endothelial growth factor) for cancer chemotherapy]. AB - Recent progress in cancer biology has revealed that angiogenesis is a promising target for new anticancer drugs. Angiogenesis is tightly regulated by the balance between stimulatory and inhibitory angiogenic factors, and the imbalance of these regulators causes dysfunction of angiogenesis. Vascular endothelial growth factor (VEGF) is one of the best characterized pro-angiogenic factors, and multiple strategies have been studied to inhibit the pathway; i. e. production and secretion of VEGF receptor, VEGF binding to its receptor, tyrosine kinase activity of VEGF, and signaling pathway downstream induced by VEGF. In this article, the summary of function of VEGF family as well as recent promising drugs under clinical trials including bevacizumab (Avastin), a humanized monoclonal antibody developed against VEGF, and several small molecule inhibitors targeting VEGF function are described. PMID- 15283143 TI - [Telomere and telomerase as targets for anti-cancer drugs]. AB - Telomerase is a hopeful molecular target of anti-cancer drugs because it is practically specific and essential for survival and growth of cancer cells. Cancer cells in vitro ceased proliferation after introduction of dominant negative cDNA or RNAi of telomerase catalytic subunit (hTERT) or anti-sense RNA of template RNA subunit (hTR) of telomerase. Some reverse-transcriptase inhibitors, compounds that stabilize G-quartet structure of telomere DNA and other natural or synthetic compounds were reported to inhibit telomerase activity and proliferation of cancer cells in vitro. Vectors carrying cDNA that produces mutant hTR partly blocked proliferation of cancer cells probably through inhibition of sequence-specific binding of telomere proteins. Telomere binding proteins are possible targets of anti-cancer drugs that modify telomere structure and block recruitment of telomerase. Combination of inhibitors with different action mechanisms accelerated telomere shortening and reduced period of time required for cancer cell killing. Repressor of hTERT expression, menin, is a possible target in future. PMID- 15283144 TI - [Telomerase inhibitor, telomestatin, a specific mechanism to interact with telomere structure]. AB - A novel telomerase inhibitor, telomestatin, isolated from Streptomyces anulatus is the most potent telomerase inhibitor so far. Telomestatin specifically inhibited telomerase without affecting reverse transcriptases and polymerases. In addition, telomestatin induced telomere shortening, but its ratio was extremely faster than that observed in physiological telomere shortening. These results suggested the existence of other mechanisms to inhibit telomerase. Telomeres consist of guanine rich sequences which compose a characteristic three dimensional structure designated as G-quadruplex. Stabilization of G-quadruplex structure inhibited the catalysis of not only telomerase but also other DNA interacting molecules. Telomestatin potently stabilized G-quadruplex structure in a specific manner. G-quadruplex structure is also involved in a lot of oncogene promoters. Thus, telomestatin provide the novel therapeutic molecular target for cancer chemotherapy. PMID- 15283145 TI - [Mode of action and molecular target of ECH, a specific inhibitor of death receptor-dependent apoptosis]. AB - ECH (epoxycyclohexenone) specifically blocks death receptor-mediated apoptosis induced by anti-Fas antibody, Fas ligand, or TNF-alpha, whereas it has no effect on death receptor independent apoptosis induced by staurosporine, MG-132, C2 ceramide, or UV irradiation. ECH blocks the activation of pro-caspase-8 in the death-inducing signaling complex (DISC), even though recruitment of FADD and pro caspase-8 is not affected. In Fas ligand treated cells, ECH is only able to inhibit the activation of pro-caspase-8 and it has no effect on the already activated caspase-8. ECH has a relatively higher affinity to pro-caspase-8, although it directly binds both pro- and active-form of caspase-8. In conclusion, ECH targets pro-caspase-8 and blocks the self-activation of pro-caspase-8 in the DISC, and thus selectively inhibits death receptor-mediated apoptosis. Moreover novel non-peptide inhibitors, RKTS-33 & RKTS-34 that are chemically synthesized derivatives of ECH have been developed. PMID- 15283146 TI - [Heat shock protein 70 binding protein 1 induces enhanced apoptotic response against anticancer drugs in tumor cells]. AB - HspBP1 was originally identified and characterized as a novel Hsp70/Hsc70 interacting protein that inhibited the Hsp70/Hsc70 chaperone activity. In this respect, Hsps have been shown to influence cell-death pathway through the interaction with key components of the apoptotic machinery. In this report, we have examined the effect of HspBP1 overexpression on the sensitivity of tumor cells to anticancer drug-induced apoptotic response. Analysis with HspBP1 deletion mutant genes showed that C-terminus 47 amino acid residues were essential for the specific interaction of HspBP1 with Hsp70/Hsc70. Overexpression of HspBP1 induced enhanced apoptotic response against anticancer drugs, but HspBP1 lacking C-terminus 47 amino acid residues did not. These results support the notion that inhibition of the function of Hsp70 enhances the therapeutic efficacy of chemotherapy. PMID- 15283147 TI - [The mechanisms of the resistance to molecular targeting agents]. AB - Increasing knowledge of the mechanism of the initiation and progression of various cancers is the catalyst for developing new anticancer therapeutics that target specific molecules expressed in cancer cells. STI571 (imatinib mesylate) is an example of the successful development of a rationally designed and targeted agent. Its target is the constitutively active tyrosine kinase, BCR-ABL in chronic myelogenous leukemia (CML). Clinical studies with STI571 in CML demonstrated that many patients with advanced stage disease respond initially but then relapse. Drug resistance is associated with the reactivation of BCR-ABL signal transduction. Another targeted protein-tyrosine kinase inhibitor that was approved for clinical use is ZD1839 (Iressa). ZD1839 is an orally active and selective inhibitor for epidermal growth factor receptor (EGFR) tyrosine kinase. HER2 is overexpressed in 25-30% of breast cancers and associated with shorter time to relapse and lower survival rate. Specific targeting of these cancers can be accomplished with Herceptin directed against the extracellular domain of the HER2 protein. However, even in the selected group of patients with high levels of HER2, the response to Herceptin is limited in magnitude and duration. The mechanisms of the resistance to these targeted agents were reviewed. PMID- 15283148 TI - [Trastuzumab treatment for breast cancer]. AB - Trastuzumab is a recombinant humanized monoclonal antibody directed against the HER-2. HER-2 is overexpressed in 20-25% of invasive breast cancer and known as poor prognostic factor and its status of a tumor is related to response to trastuzumab. Combination therapy with trastuzumab and chemotherapy increases response rate, time to progression, and survival thus trastuzumab is perceived to be the first line treatment for HER-2 overexpressed metastatic breast cancer. The optimal duration and period of trastuzumab treatment are still unknown and result of large scale randomized trial will be expected. PMID- 15283149 TI - [Gefitinib in non-small cell lung cancer]. PMID- 15283150 TI - [Rituximab and hematological malignancy]. AB - After genetic recombination with murine and human cDNA, monoclonal antibody has been produced in chimeric form. CD20 is a B cell antigen, which can be a target molecule for antibody therapy. Rituximab is an anti-CD20 monoclonal antibody that has an efficacy against B cell lymphomas. Combination therapy with CHOP regimen as well as single therapy will give rise to new treatment strategy for CD20+B cell lymphomas. Mechanism is not clarified, but induction of apoptosis, complement-mediated cell toxicity, and antibody-dependent cell toxicity are considered. Synergy with chemotherapy agents such as CHOP, ICE, DHAP and stem cell transplantation will help the patients who suffer lymphomas, CLL and Waldenstrom's macroglobulinemia. Treatment with rituximab is well tolerated and we must be careful about infusion reaction after administration. PMID- 15283151 TI - [Imatinib therapy for patients with chronic myelogenous leukemia]. AB - Imatinib mesylate (imatinib), a selective inhibitor of BCR-ABL tyrosine kinase, has shown excellent efficacy in patients with chronic myelogenous leukemia (CML) in the chronic phase, however, it does not in those in the accelerated phase or blastic crisis. In patients with CML who have undergone allogeneic stem cell transplantation, imatinib has the capability to induce hematological and even molecular response, and provides a prolonged survival among those in the chronic and accelerated phases. It has been demonstrated that major cytogenic response is a surrogate marker for survival in cases receiving imatinib. It has also been demonstrated that a genome-wide cDNA microarray enables the prediction of sensitivity to imatinib. The acquired resistance in patients who failed to respond to imatinib seemed to be induced by several point mutations in the BCR ABL gene, which were likely to affect the binding of imatinib with BCR-ABL. Polyclonal cells which harbor distinct mutations in a single patient seemed to be selected in vivo under the selective pressure of imatinib, indicating the rationale of combined treatment with other types of agents. Recently, SPIRIT (STI571 Prospective International Randomized Trials) have been conducted, in which the efficacy of imatinib monotherapy, and imatinib combined with interferon or cytarabine were compared. New agents which inhibit the signaling pathway related to BCR-ABL, such as adaphostin (NSC680410), farnesyltransferase inhibitor SCH66336, MAP kinase inhibitor PD184352, PD98059, U0126, and antibiotic geldanamycin, have shown excellent activity combined with imatinib in vitro. PMID- 15283152 TI - [The effect of imatinib for gastrointestinal tumor]. AB - Long-term survival in patients with gastrointestinal stromal tumors (GIST) was very rare. Recently treatment with imatinib mesylate, a molecular targeted agent that inhibits the KIT tyrosine kinase receptor showed 81.6% of outstanding clinical response (PR 53.7%, SD 27.9%). Toxicities with daily dose of 400 mg and 600 mg, all patients had grade 1 or 2 toxicity. Grade 3 or higher toxicities occurred in 21.1% including edema, neutropenia, nausea, dermatitis, hepatitis and gastrointestinal hemorrhage. The drug was relatively safe overall. PMID- 15283153 TI - [Hairy cell leukemia therapy by cladribine]. AB - Cladribine (2-chlorodeoxyadenosine: 2-CdA), a purine nucleoside analog, is a safe and very effective treatment for patients with hairy cell leukemia (HCL). 2-CdA was approved in USA and Japan, and is first-choice drug against symptomatic HCL. It is administered at a dose of 0.09 mg/kg daily as a continuous intravenous infusion over 7 days. Structure, mechanism of action, clinical data of efficacy and safety of 2-CdA, and indications for therapy for HCL are reviewed. PMID- 15283154 TI - [Resistance to target-based therapy and its circumvention]. AB - Intrinsic and acquired resistance to molecular target therapy critically limits the outcome of cancer treatments. Target levels including quantitative and gene alteration should be determinants for the resistance. Downstream of the target molecules, drug metabolism, and drug transport influences the tumor sensitivity to molecular target therapy. The mechanisms of resistance to antibody therapy have not been fully clarified. Correlative clinical studies using these biomarkers of resistance are extremely important for circumvention of clinical resistance to target based therapy. PMID- 15283155 TI - [Potentiation of radiation-induced cell killing by histone deacetylase inhibitor]. AB - HDAC inhibitor has been focused as a molecular target agent for cancer treatment because of unique pharmacological activity. HDAC inhibitors induce cancer cells to undergo growth arrest, differentiation or induce apoptosis in vitro, and also inhibit the growth of transplanted tumors in vivo, but the enhancement effect or interaction of HDAC inhibitors with other chemotherapeutic agents or radiation has not yet been fully investigated. Biade et al examined the effect of trichostatin A (TSA), and reported synergistic enhancement of radiosensitivity in human cancer cell lines. We also confirmed the similar results in vitro by using TSA in combination with radiation. The main purpose of this paper is to review the current status of HDAC inhibitor in clinical use, and evaluate the potential possibility for the potentiation of radiation-induced cell killing including the results of our research. PMID- 15283156 TI - [Overexpression of COX-2 and a potential clinical application of its inhibitors in lung cancer]. AB - It has been shown that a significantly increased expression of COX-2 is frequently present in lung cancers. Recent studies suggested that an increase in the expression of COX-2 may play a significant role in carcinogenesis in addition to its well-known role in inflammatory reactions. Interestingly, increased COX-2 expression was associated with a shortened survival of patients who underwent surgical resection of early stage adenocarcinoma, while several lines of in vitro and in vivo evidence suggest its potential role in invasion and metastasis. These findings suggest that the use of a selective COX-2 inhibitor may be a promising therapeutic approach in the treatment of lung cancer. PMID- 15283157 TI - [Proteasome inhibitor, bortezomib (Vercade), potently inhibits the growth of adult T-cell leukemia cells]. AB - The proteasome is a multicatalytic proteinase complex responsible for the degradation of most intracellular proteins, including crucial to cell cycle regulation, cell growth, and apoptosis. Bortezomib (Vercade; formerly known as PS 341) is a novel dipeptide boronic acid that is the first proteasome inhibitor to have progressed to clinical trials. Below we discuss the mechanism of bortezomib for cancer therapy, review the clinical data, and introduce the potent effect on ATL cells. PMID- 15283158 TI - [Telomerase activity in gastrointestinal, bladder and breast carcinomas and their clinical applications]. AB - Telomere ends are known to be shortened at every division of the cells. Telomerase is a ribonucleoprotein which compensate for the telomere ends and indispensable for the immortalization of the cells. It is reported that the enzyme is activated in a variety of cancer cells. In the present report, the enzyme was activated in more than 70% of gastrointestinal, bladder and breast cancer by TRAP assay. Moreover, in order to elucidate whether the enzyme activity is useful for the non-invasive detection of exfoliated cancer cells in the colon bowel washings, urine and fine needle aspirate of the breast tumors, TRAP assay was performed. In most of the cases, the positive findings were observed which support that the enzyme activity is useful for the clinical application. The expression of mRNA and protein for hTERT was detected in cancer cells, however, the assay contains the pitfall including the Taq inhibitor or telomerase inhibitor. Moreover, the activity is also detected although it is weak, in the benign or premalignant lesions. Further analysis is required for the clinical application of the method. PMID- 15283159 TI - [The mechanisms of gastric cancer development produced by the combination of Helicobacter pylori with Sterigmatocystin, a mycotoxin]. AB - It is considered that about sixty-five percent of people are suffering from Helicobacter pylori infection in our country. In the East Asian countries including Japan, such fungus as aspergillus are ubiquitously found in the environment as a contaminant in human food stuffs and animal feeds. Sterigmatocystin is a mycotoxin and a precursor of aflatoxin which is produced by Aspergillus versicolor. The mechanisms of gastric carcinoma development induced by the combination of Helicobacter pylori infection with Sterigmatocystin, a mycotoxin are shown and discussed. We revealed that Sterigmatocystin-treated cells exhibited an absence of P53-mediated G1 arrest with induction of MDM2 at 12 and 24 hours of treating time. Furthermore, it was revealed that long term treatment with Sterigmatocystin enhanced dominantly the development of intestinal metaplasia, and of precancerous lesions of gastric mucosa in Helicobacter pylori infected Mongolian gerbils. It has been reported that the accumulations of P53 nucleotide substitutions in the H. pylori-infected monkeys were increased as the score of gastric atrophy increased, nevertheless no mutations were noted in the H. pylori-uninfected monkeys. The mechanisms of the development of gastric cancer produced by combination of H.P. with ST remain to be unclear. Further study concerning the mechanisms must be carried out. PMID- 15283160 TI - [Japanese guideline for the management of gastric ulcer]. AB - Japanese guideline for the management of gastric ulcer prepared by a research group supported by government funding was published in 2003 as a concise book for use in general practice. Overall management strategy classified based on etiology of ulcer was shown as a flow chart. Recommendation of treatment strategy for NSAID ulcer with PPI, and for H. pylori-positive gastric ulcer with eradication therapy are in line with internationally acknowledged consensus. A number of so called mucoprotective drugs most of which are available in Japanese market alone, however, were given low priority when choosing single regimen, and their use in combination with acid suppressant, the most popular ulcer regimens in Japan, was not recommended due to lack of reliable evidence. Though it will take some time for general physicians to widely adopt this guideline, it undoubtedly will serve for reasonable gastric ulcer management in Japan. PMID- 15283161 TI - [Detection of micro mutation in dystrophin gene of DMD female carrier]. AB - We attempted to identify a mutation in dystrophin gene in a female patient who was suspected a Duchenne muscular dystrophy (DMD) carrier with muscle weakness of upper limbs and congestive heart failure. We examined the mutation hot spots in DMD gene, exon 3, 6, 8, 13, 17, 19, 43, 44, 45, 47, 48, 49, 50, 52, 60 by multiplex PCR which had been a diagnostic screening strategy, and detected an extra band in exon 43 product. We also detected an extra band in exon 43 products by SSCP analysis for detection of small mutations which could not be detected by multiplex PCR. As a result of sequencing a PCR product of an exon 43, we confirmed an allele having the insertion of a 2 base of AT in Intron42, which is described for the first time. Although we can not conclude that this insertion is responsible for DMD, but it may cause abnormal splicing. In carrier detection of DMD without genetic information of proband, it is difficult to detect mutations by multiplex PCR solely. Therefore, SSCP of PCR products are recommended to detect mutation in DMD carrier. PMID- 15283162 TI - [An attempt for standardization of serum CA19-9 levels, in order to dissolve the gap between three different methods]. AB - It is well known that serious method-related differences exist in results of serum CA19-9, and the necessity of standardization has been pointed out. In this study, differences of serum tumor marker CA19-9 levels obtained by various immunoassay kits (CLEIA, FEIA, LPIA and RIA) were evaluated in sixty-seven clinical samples and five calibrators and the possibility to improve the inter methodological differences were observed not only for clinical samples but also for calibrators. We supposed an assumed standard material using by a calibrator. We calculated the serum levels of CA19-9 when using the assumed standard material for three different measurement methods. We approximate the CA19-9 values using by this method. It is suggested that the obtained CA19-9 values could be approximated by recalculation with the assumed standard material would be able to correct between-method and between-laboratory discrepancies in particular systematic errors. PMID- 15283163 TI - [Diabetic nephropathy and plasminogen activator inhibitor 1 in urine samples]. AB - Plasminogen activator inhibitor-1 (PAI-1) may contribute to renal fibrosis because of its involvement in matrix (ECM) accumulation through inhibition of plasmin-dependent ECM degradation. The aim of this study is to determine urinary PAI-1 concentrations and its intrarenal localization in patients with various renal diseases and to identify inducers for PAI-1 expression in human cultured proximal renal tubular cells (HRCs). Urinary PAI-1 concentrations were significantly higher in patients with overt diabetic nephropathy (DN, n=36) than in proliferative glomerulonephritis (PGN, n=8), nephrotic syndrome (NS, n=10) and healthy controls (n=12). Urinary PAI-1 concentrations (ng/gCr) were directly correlated with urinary N-acetyl glucosaminidase (NAG) levels (r=0.58, p<0.05). As for intrarenal localization of PAI-1 antigen, strong stainings for PAI-1 were observed in proximal tubular cells of renal biopsy samples from patients with DN, while no stainings for PAI-1 were found in renal tissues of PGN or NS. Immunoblot analysis revealed the presence of PAI-1 protein in whole cell lyzates from HRCs grown to semiconfluency. Exposure of growth-arrested HRCs with hypoxia (1% O2) or TNF-alpha (10 ng/ml) for 24 hours increased the secretion rate of PAI-1 protein by about 2.0-fold, while 24-hour treatment with high glucose (450 mg/dl) did not increase PAI-1 secretion at all, compared with that of the control cells under normal glucose (100 mg/dl) and normoxia (18% O2). These findings suggest that PAI 1 expression is upregulated especially in the proximal renal tubular cells of DN, which may be explained partially by hypoxia and inflammatory cytokines but not high glucose. PMID- 15283164 TI - [Self-testing using by OTC test kits and mail testing services--it's future prospect and problems]. AB - Under the certification of the Ministry of Health, Labor and Welfare, OTC medicine (Over the counter medicine: medicine for general tests) such as test strips for urine protein and urine glucose as well as urinary pregnancy tests are presently available at pharmacies and drug stores for use as self-testing by ordinary people. Meanwhile, mail health check-up testing services have recently been making a rapid progress in the market. In this system, users purchase "sample collecting kits" at large-scale pharmacies, directly or indirectly, and then after collecting samples (urine, blood, etc) by themselves, they put the samples onto the mail testing service process consisting of the linkage of convenience stores--couriers--testing agencies affiliated with clinical facilities. The samples are tested in the testing agencies and the results of the testing are returned to individual user. This innovative system allows users to control and manage their own health conditions. Future prospects and problems over the system will be discussed here. PMID- 15283166 TI - [Future of clinical laboratory medicine--review of the system in United States]. AB - As the clinical laboratory medicine is the integral bridge between clinical patient care and basic science, both the high quality patient care service and the active laboratory research challenges are indispensable. This article introduces the state of the department of laboratory medicine in M.D. Anderson Cancer Center (MDACC), Houston, Texas, USA. The department of laboratory medicine in MDACC processes over 4 million tests per year and performs over 1,800 different tests and assays with good cost effectiveness. The laboratory staff carry a heavy patient care service load in addition to laboratory support for the clinical research protocols in the other clinical divisions. Research projects in the laboratory medicine are aimed primarily at establishing new or improved methodologies and procedures for clinical laboratory testing, and evaluation of prototypes for new analytic instruments. Generally, the translation of the research findings into effective clinical laboratory tests is the important approach. The more strength should be on the productive research collaboration with basic science and clinician colleagues. PMID- 15283165 TI - [Laboratory medicine in the field of emergency medicine and disaster medicine- Old-And-New tools of emergency medical service]. AB - We reviewed the characteristics of the field of laboratory medicine concerning emergency medicine and disaster medicine. Gram's stain was apt to be made light of by clinicians, but it has been reviewed again. A modern sophisticated analysis system is expected for the consequence management of NBC terrorism and mass poisoning, but we would like to emphasize the importance of the basic physical strength of laboratory medicine, such as Gram's stain, to the meaning of old-and new technology. PMID- 15283167 TI - [On-site screening of toxic substances in clinical laboratory and poisoning information system]. AB - An accurate screening system is required to treat acute poisoning patients in clinical toxicology. However, the medical center analysis of poisonous substances using machines is not sufficient. Moreover, the handling and maintenance of such machines are tedious and costly. To improve these problems and employ effective information, we have developed a simple detection method and constructed a support system using the Internet. Various support systems have been attempted for the training of analysts who can cope with a poisoning incident (accident) involving toxic substances. Our simple detection method for toxic substances in the medical center was developed without using expensive analysis apparati. As technical support for the analysts of medical laboratories, the following items were completed: 1) training for analysts, 2) research of analytical techniques in the medical centers (accuracy management), 3) creation of an analysis manual, 4) construction of on-line analysis manuals, 5) construction of the poisoning information system on the Internet, 6) construction of a system for requesting analysis of poisoning, 7) a quick-detection method for toxic substances and 8) examination of the insurance application. PMID- 15283168 TI - [Analysis of the interaction between DnaA protein which is involved in the initiation of chromosomal replication in E. coli and its specifically binding DNA]. AB - DnaA protein binds specifically to a group of binding sites called DnaA boxes within the bacterial replication origin to induce local unwinding of duplex DNA. DnaA domain IV comprises 94 amino acid residues and is required for DNA binding. At first, the backbone assignments of DnaA domain IV in the complexes were determined using several nuclear magnetic resonance (NMR) spectra. Using NMR analysis, we investigated the interaction between DnaA domain IV and DnaA box R1. The 1H-15N HSQC spectrum of DnaA domain IV showed prominent chemical shift perturbations on six residues (Arg399, Ala404, Leu422, Asp433, Thr435 and Thr436). Through homology modeling, we located all of these residues on one side of the surface of the DnaA domain IV molecule. Moreover, we confirmed that these residues in DnaA domain IV bind to DnaA box R1 by mutation analysis. Finally, we compared the chemical shift perturbation of the 1H-15N HSQC spectrum in the presence of the DnaA box with that in the presence of a non-specific oligonucleotide that has a reduced affinity for DnaA, and the results suggested that Leu422 imparts specificity in binding with DnaA box R1. PMID- 15283169 TI - [A new role of pathologists in medical risk management]. AB - Safety management is the prime issue for health professionals. How can we pathologists to improve their risk sense to prevent medical errors? Various autopsy cases such as drug adverse events, unexpected sudden deaths and/or multiple organ failures after operations have made us aware of faulty systems of health care. Thus, the scope of pathology must be extended from the traditional tissue and cell pathology to the implementation of preventive pathology, by participation in morbidity and mortality conferences. PMID- 15283170 TI - [Clinical efficacy assessment of diagnostic tests: from the standpoint of analysis of prognostic factors]. AB - The usefulness of laboratory tests has been assessed by indices such as diagnostic sensitivity, specificity and predictive values. This paper describes the means by which usefulness is evaluated by analysis of prognostic factors, including risk factors and clinical outcome of therapy. It is essential to follow appropriate research designs and to make use of appropriate statistical analyses. Some of the efficient means for analyzing prognostic values include prospective investigation, as represented by cohort studies which follow characteristics of groups for a period of time, and retrospective investigation, as represented by case-controlled studies which evaluate characteristics of groups in a retrospective manner. The appropriate method of analysis for sample data which contain time factors and a relationship between cause and outcome may be multiple logistic regression analysis. This analytical method can present the effects of each factor on a given disease by odds ratios, and can also calculate the occurrence probability of the disease for each case. For example, we analyzed the prognostic factors that govern the effects of interferon therapy on hepatitis type C. We found that the genetic type and quantity of the virus are the major prognostic factors determining the efficacy of interferon therapy, which is in good accord with previous studies. Furthermore, we were able to identify some factors which have not been recognized before. Thus, appropriate research designs not only provide new insights into therapeutic approaches but can also improve the usefulness of laboratory tests. PMID- 15283171 TI - [Quality assurance of analytical methods to guarantee the reliability of medical decision levels for interpretation of clinical laboratory data]. AB - We sought to investigate the effects of measurement errors on the evaluation of biological variations in healthy subjects. To this end, we analyzed the allowable limits of analytical error which can guarantee the reliability of medical decision levels for interpretation of clinical laboratory data. As a conclusion, we suggest that one-half or less of biological intra-individual variations is appropriate as the criterion for an allowable limit of error to be applied in health check-ups, and this value is in agreement with those in previous reports. If this criteria as the marker for intra-laboratory imprecision is met, a given institute should be able to evaluate time series change follow up of individual data. If the reference interval of laboratory data for disease screening is shared by different institutes, the criterion of 1/4 or less of a biological inter pulse intra-individual variation should be appropriate. This criterion appears to be the goal to be achieved for analytical inter-laboratory variations. On the other hand, as for the criterion for measurement errors which guarantees disease identification based on the cut-off value, a criterion of 1/4 or less of biological inter-pulse intra-individual variation appears to be appropriate, taking into consideration measurement errors which did not influence false positive or false-negative rates of disease identification. The value turned out to be the same as the limit for the screening of disease. In this study, we considered allowable limits of error in the vicinity of reference value concentrations. However, it will be necessary to set separately the criteria for data in abnormal ranges. PMID- 15283172 TI - [Immune regulation by malaria parasites]. PMID- 15283173 TI - [Double anterior (anterolateral and anteromedial) thigh flaps for reconstruction of head and neck defects]. AB - Although the anterolateral and anteromedial thigh flaps have such disadvantages as anatomical variations in the lateral circumflex femoral system and its cutaneous perforators, they have a sufficient number of cutaneous perforators on the anterior aspect of the thigh in many cases, meaning double anterior thigh flaps can be elevated from the ipsilateral thigh. We report 8 cases in which single-pedicle double anterior thigh flaps were transferred for reconstruction of head and neck defects following resection of head and neck tumors from January 1995 to March 2001 at Okayama Saiseikai General Hospital. Eight double anterior thigh flaps were classified into the following 3 types by perforator derivation: Type I flaps consisting of double anterolateral thigh flaps with a single vascular source were elevated in 4 cases. Type II flaps consisting of anterolateral thigh flap and anteromedial thigh flap supplied by separate branches from a single vascular source were elevated in 2 cases. Type III flaps, in which two anterior thigh flaps were harvested separately and constructed into a single flap with microvascular anastomosis, were elevated in 2 cases. Double anterior thigh flaps were combined with vascularized bone, and 2 flaps supplied by independent long vascular pedicles from a single vascular source could be 3 dimensionally arranged. All flaps completely survived in 7 cases and functionally and aesthetically acceptable results were obtained except in 1 case whose flap was lost to major necrosis after infection. We concluded that the single-pedicle double-flap technique using anterior thigh flaps is useful in 3-dimensional reconstruction of massive complex head and neck defects since sufficient tissue implant with preserved blood flow and free flap arrangement is made available. PMID- 15283174 TI - [Laminin gamma2 chain expression in squamous cell carcinomas of the tongue and its clinical relevance]. AB - Laminin gamma2 chain (LNgamma2) expression and its clinical relevance were examined in squamous cell carcinomas of the tongue. When tumor cells were attached to each other and showed expansive growth, LNgamma2 was expressed only in the peripheral cells of the tumor nests (peripheral expression). In contrast, when tumor cells showed infiltrative growth diminishing cell-cell adhesion, LNgamma2 expression was diffusely observed in almost all of the cells (diffuse expression). Patients with caricinoma cells of the primary lesions showing peripheral LNgamma2 expression were classified as the peripheral expression type, whereas patients with carcinoma cells showing diffuse LNgamma2 expression at least in part of the invasive fronts were classified as the diffuse expression type. Among 30 patients with tongue carcinomas of more than stage II, 19 patients were the peripheral expression type and 11 patients were the diffuse expression type. The 3-year disease-specific survival rates for the peripheral type and diffuse type were 64% and 34%, respectively. LNgamma2 may play an important role in growth and invasion of tongue carcinomas. In particular, it seems likely that the diffuse LNgamma2 expression in carcinoma cells has a significant relevance to the malignant characteristics of infiltrative carcinoma cells. In addition, LNgamma2 expression may be a useful prognostic factor for the patients with carcinomas of the tongue. PMID- 15283175 TI - [Head and neck reconstruction using laparoscopically harvested omentum]. AB - Since the need for laparotomy in harvesting the omentum is the most significant drawback, the omentum has not been the tissue of choice for reconstructive surgery. To compensate for this drawback, we started laparoscopic harvesting of the omentum and clarified the advantages and disadvantages of this procedure. Ten patients underwent laparoscopic harvesting of the omentum by abdominal surgeons, followed by reconstruction of head and neck defects. Surgery was conducted in 5 cases of defect reconstructions for parotid gland tumor surgery and 5 of oropharyngeal defect after cancer surgery. The average harvesting time was 107 minutes (55-140 minutes) and used the omentum and different amounts and length of the vascular pedicle. Although the omentum was successfully transplanted in 9 of 10 cases, 2 cases showed partial peripheral necrosis and 1 total necrosis. With the advantage of laparoscopic harvesting of the omentum, we could obtain appropriate omental size for the defect size. Especially after total parotidecomy, the omentum was useful to fill in the defect, reducing the patients' worries about postoperative deformity. In one case, the omentum was used to treat Frey syndrome, successfully relieving the symptoms. In oropharyngeal reconstruction, the omentum is used to fill dead space and prevented postoperative infection. Although mild abdominal pain was observed a few days after surgery, no major abdominal complications such as intestinal perforation or ileus occurred in the 8 to 39 months following laparoscopic harvest of the omentum. Since the omentum is pliable and easily fills a complicated defect, the omentum is considered satisfactory for reconstructing defects of the lateral face after parotid tumor surgery and small defects after oropharyngeal tumor surgery. PMID- 15283176 TI - [A case report on congenital anosmia]. AB - We report 2 cases of congenital anosmia, in a 13-year-old girl and the other in a 10-year-old boy. They reported having no concept of "smell". The girl has no complications but the boy has congenital microphthalmia and is completely blind. They showed scale-out results on both T & T olfactometry and intravenous Alinamin test. Brain MRI detected hypoplasia or lack of the olfactory bulbs, tracts, and olfactory sulci in the frontal lobe of the brain in both patients. Neither had endocrinal dysfunction. In the boy, we biopsied the nasal mucosa in the olfactory cleft and found it had no olfactory epithelial cells at all. We found MRI to be the most useful imaging for diagnosing congenital olfactory disturbance. PMID- 15283177 TI - Management of spontaneous dissection of the superior mesenteric artery. PMID- 15283178 TI - Associations among lifestyle status, serum adiponectin level and insulin resistance. AB - OBJECTIVE: The aim of this study was to determine whether lifestyle status affects the insulin resistance index or serum adiponectin level, which may be responsible for the development of insulin resistance syndrome that sometimes indicates lifestyle-related diseases. METHODS: A cross-sectional study was performed. PATIENTS: Seven hundred thirty-eight males aged 30 to 65 years who had regular health checkups in our office were enrolled. Each subject's lifestyle status, level of serum adiponectin, and serum insulin level were assessed by a self-administered questionnaire based on Breslow's lifestyle index, enzyme-linked immunosorbent assay, and radioimmunoassay, respectively. Moreover, their insulin resistance indexes were assessed by the homeostasis model. RESULTS: One-way ANOVA demonstrated an inverse correlation between Breslow's index and the logarithmic insulin resistance index (p<0.0001), and a tendency of a correlation between Breslow's index and the logarithmic serum adiponectin level (p=0.0681). Multiple logistic regression analyses demonstrated that, among the seven lifestyle items in Breslow's index, body mass index of more than 26.1 kg/m2 and insufficient exercise style had 8.9 times and 2.1 times the risks for insulin resistance and the former also had 3.2 times the risk for hypoadiponectinemia. Partial correlation coefficients of these correlations were 0.336 (p<0.0001), 0.107 (p=0.0013), and 0.165, (p<0.0001), respectively. CONCLUSION: Unhealthy lifestyles may cause hypoadiponectinemia and insulin resistance followed by insulin resistance syndrome, i.e. lifestyle-related diseases. These findings present reasonable explanations for the relationships between lifestyles and lifestyle related diseases. Improvement of unhealthy lifestyles, especially the control of body weight, may have beneficial effects against the development of lifestyle related diseases. PMID- 15283179 TI - Hepatopulmonary syndrome in a patient with primary biliary cirrhosis. AB - A 54-year-old Japanese woman with primary biliary cirrhosis (PBC) was admitted to our hospital due to hepatic coma and refractory pleural effusion. The physical examination revealed clubbed fingers and collateral veins. The patient had an increased alveolar-arterial oxygen gas tension difference. The levels of anti mitochondrial antibody (AMA) and AMA M2 was 80 times normal. A technetium 99m labeled macro-aggregated human albumin scintigram showed uptake in the spleen and the kidneys. A diagnosis of hepatopulmonary syndrome (HPS) was made. HPS may be overlooked because of the lack of symptoms. We conclude that closer attention should be paid to the occurrence of HPS. PMID- 15283180 TI - Spontaneous esophageal submucosal hematoma in which the course could be observed endoscopically. AB - A 66-year-old man was hospitalized after vomiting blood after inducing vomiting using his fingers due to laryngeal discomfort. Upper digestive tract endoscopy revealed a large, dark red mass that connected from the upper esophagus to the lower esophagus. Esophageal submucosal hematoma was diagnosed using endoscopy, X ray images, a small-diameter ultrasonic probe, and chest CT scanning. Pain from the epigastrium to the larynx disappeared after 3 days. Melena occurred on Day 3. Endoscopic examination revealed that the hematoma had collapsed over a wide area. Endoscopic examination after one week showed that the mucous membrane covering the hematoma had peeled away revealing an extensive shallow ulcer in the esophagus. Endoscopic examination after one month confirmed the ulcer had scarred and healed. PMID- 15283182 TI - Spontaneous dissection of the superior mesenteric artery in four cases treated with anticoagulation therapy. AB - Reports of spontaneous dissection of the superior mesenteric artery are rare. Diagnosis in the acute stage has been considered difficult, but we encountered four cases from November 1998 to November 2001. All four cases were diagnosed using abdominal CT scanning in the acute stage and could be treated conservatively. All patients were provided anticoagulation therapy upon fasting. The mean period of continuous abdominal pain was 10.2 days, the mean period of fasting was 27.2 days, and the mean number of in-hospital days was 44.5. There is no established opinion on treatment, but conservative treatment is considered possible if there are no symptoms or if it has not been aggravated. PMID- 15283181 TI - Recurrent gastric hemorrhaging with large submucosal hematomas in a patient with primary AL systemic amyloidosis: endoscopic and histopathological findings. AB - A 64-year-old woman who suffered intractable gastric ulcers with hemorrhaging showed huge submucosal hematomas in her stomach on the endoscopic examination. Since gastric mucosal biopsy revealed amyloid deposition and IgG lambda type M protein was detectable in her serum, she was diagnosed as having primary AL systemic amyloidosis. The gastric hemorrhages did not improve despite intensive medication, so total gastrectomy was performed, resulting in an unfavorable outcome. Massive deposition of amyloid with A lambda immunoreactivity was seen on the submucosal vessels in her stomach. This is a rare primary AL systemic amyloidosis case showing recurrent and fatal gastric submucosal hematomas. PMID- 15283183 TI - Intussusception of the appendix that reduced spontaneously during follow-up in a patient on hemodialysis therapy. AB - A 63-year-old man on long-term hemodialysis therapy was hospitalized due to right lower abdominal pain. CT scan demonstrated a multiple concentric structure in the ileocecal region. Colonoscopy showed a polyp-like tumor arising from the expected location of the appendix, with a dimple at the top. Barium enema study revealed a submucosal tumor-like filling defect in the cecum with non-filling of the appendix. A diagnosis of intussusception of the appendix (IA) was made. During the follow-up, IA reduced spontaneously. The present case report is the first description of IA in a patient on hemodialysis therapy. Furthermore, spontaneous reduction of IA is indeed rare. PMID- 15283184 TI - Histopathologically-diagnosed splenic metastasis in a hepatocellular carcinoma case with adrenal metastasis. AB - We encountered a patient with hepatocellular carcinoma (HCC), with adrenal gland metastasis, in whom splenic metastasis was diagnosed histopathologically. A 59 year-old man visited our hospital in May 2001 with chief complaints of abdominal distension and pretibial pitting edema. Multiple HCCs associated with HCV positive liver cirrhosis were detected. Transarterial embolization (TAE) was performed a total of 4 times for HCCs. A left adrenal gland metastatic lesion was detected and it was found to increase in diameter from 3 cm to 6 cm over a four month period; left adrenalectomy was performed in June 2002. Because of marked splenomegaly and findings of hypersplenism, the spleen was also resected. Although no metastatic lesions were evident on macroscopic examination of the spleen, a small metastatic lesion from moderately differentiated HCC, approximately 0.5 mm in diameter, was detected histopathologically. Splenic metastasis from HCC is rare, usually occurring with metastases involving other organs. Our patient also had adrenal gland metastasis. Therefore, hematogenous metastasis to the congested spleen via the systemic circulation was suspected. PMID- 15283185 TI - Scintigraphic evaluation of cardiac metabolism in multicentric Castleman's disease. AB - A 72-year-old woman had insidious onset of heart failure, and was diagnosed as multicentric Castleman's disease. She underwent myocardial imaging with technetium-99m tetrofosmin, 1-123 beta-methyl-iodophenyl pentadecanoic acid (BMIPP). Technetium-99m tetrofosmin studies showed almost normal uptake of the left ventricular myocardium indicating normal myocardial perfusion. 1-123 BMIPP showed reduced uptake in the apical segment of the myocardium, indicating regional fatty acid metabolic abnormalities. PMID- 15283186 TI - Thymic squamous cell carcinoma producing parathyroid hormone-related protein and CYFRA 21-1. AB - A 54-year-old man was admitted to our hospital because of dyspnea. Radiographic examination showed an anterior mediastinal mass and pericardial effusion. Serum calcium and parathyroid hormone-related protein (PTHrP) levels were elevated, and serum CYFRA 21-1 level was extremely high. Results of percutaneous needle biopsy under computed tomography guidance led to a diagnosis of moderately differentiated squamous cell carcinoma. Immunohistological staining showed the tumor cells to be positive for PTHrP and cytokeratin monoclonal antibodies. Postmortem findings were considered to indicate thymic carcinoma. Thymic carcinoma is rare, but our case indicates that thymic squamous cell carcinoma can be identified in terms of paraneoplastic hypercalcemia. PMID- 15283187 TI - ANCA-related crescentic glomerulonephritis in a patient with scleroderma without marked dermatological change and malignant hypertension. AB - A 64-year-old woman with scleroderma without marked dermatological change developed anti-neutrophil cytoplasmic autoantibody (ANCA)-related renal failure. She had neither malignant hypertension nor elevation of plasma renin concentration. Renal biopsy showed crescentic glomerulonephritis (pauci-immune type) and the myeloperoxidase-specific anti-neutrophil cytoplasmic autoantibody (MPO-ANCA) titer was found to be elevated to 757 IU/ml. Methylprednisolone pulse therapy followed by oral prednisolone effectively suppressed renal failure and lowered the MPO-ANCA titer. We believe this is a rare case of ANCA-related renal failure in a patient with scleroderma without marked dermatological change. PMID- 15283188 TI - Legionella micdadei pneumonia diagnosed by culture isolation and DNA-dNA hybridization from bronchial lavage fluid. AB - An 80-year-old man was admitted because of dyspnea on effort. We suspected an acute exacerbation of chronic heart failure and idiopathic interstitial pneumonia caused by right-sided pneumonia. A nodular shadow in right upper lobe spread and consolidated into the airspace, and it failed to improve despite administration of meropenem trihydrate, vancomycin hydrochloride and clindamycin. A definitive diagnosis of Legionella micdadei pneumonia was made on the basis of this organism being isolated in culture from bronchial lavage fluid and subsequent identification of Legionella micdadei using DNA-DNA hybridization. The airspace consolidation gradually improved following treatment with intravenous erythromycin and minocycline hydrochloride. PMID- 15283189 TI - Acute myelogeneous leukemia (M5a) that demonstrated chromosomal abnormality of robertsonian 13;21 translocation at onset. AB - A 27-year-old woman had congenital lissencephaly syndrome and mental retardation. She had a fever of unknown origin and visited her local physician. Blood test indicated leukocytosis, so she was referred to our hospital for detailed examination. She was diagnosed to have acute myelogeneous leukemia (M5a). The chromosome analysis in blast cells revealed Robertsonian 13;21 translocation. Complete remission was obtained by induction chemotherapy. As normal karyotype (46, XX) was observed in the chromosome analysis of bone marrow cells after remission, it was considered that the patient had acquired Robertsonian 13;21 translocation complicated by acute myelogeneous leukemia. PMID- 15283190 TI - Minimal change nephrotic syndrome after allogenic hematopoietic stem cell transplantation. AB - We describe a 24-year-old man who developed minimal change nephrotic syndrome after allogenic hematopoietic stem cell transplantation (HSCT). One year after undergoing allogenic peripheral blood stem cell transplantation (PBSCT), this patient presented with proteinuria. He also presented with skin and lip lesions considered to be chronic graft-versus host disease. Observation of renal biopsy specimens by light microscopy revealed minor glomerular abnormalities. However, electron microscopy disclosed focal mesangial interposition, irregular thickening of the glomerular basement membrane and subendothelial loosening. Two years after HSCT, the patient developed nephrotic syndrome. Long use of cyclosporine improved the proteinuria and hypoalbuminemia within 12 months. PMID- 15283191 TI - Graves' disease accompanied by anti-myeloperoxidase antibody-related nephropathy and autoimmune hepatitis. AB - A 17-year-old woman was admitted because of proteinuria, microhematuria and liver dysfunction with increased antinuclear antibody and anti-myeloperoxidase antibody (MPO-ANCA). Fourteen months' previously, urinalysis and liver function showed normal range. At that time she suffered from tachycardia and weight reduction, diagnosed as Graves' disease, she was given propylthiouracil for treatment of her Graves' disease. The histological finding of renal biopsy was compatible with minor glomerular abnormalities. Liver biopsy finding was compatible with autoimmune hepatitis. After we had administered prednisolone, liver function returned to normal range and urine protein became negative. Then we performed subtotal thyroidectomy, and she was not given propylthiouracil. MPO-ANCA decreased gradually. PMID- 15283192 TI - Tuberculous liver abscess not associated with lung involvement. AB - Hepatic tuberculosis is one of the uncommon forms of extrapulmonary tuberculosis. We report a 78-year-old woman who developed tuberculous liver abscesses with splenic abscess not associated with pulmonary foci. Ultrasonography and computed tomography of the abdomen showed the low-density lesions in the liver and spleen. Histopathology of specimens obtained by percutaneous needle biopsy revealed coagulation necrosis and epithelioid cells but not tumor cells, suggesting tuberculosis infection in the liver and spleen. Systemic chemotherapy with anti tuberculous agents led to the improvement of the lesions in the liver as well as spleen. Although tuberculous liver abscess is a very rare case, it should be included in the differential diagnosis of unknown hepatic mass lesions. PMID- 15283193 TI - Primary malignant lymphoma of bone marrow. PMID- 15283194 TI - The desire to do something good. PMID- 15283195 TI - Microfluidics. Big things in narrow channels. PMID- 15283196 TI - Influence of matrix application timing on spectral reproducibility and quality in SELDI-TOF-MS. PMID- 15283197 TI - DNA ligation by selection. PMID- 15283198 TI - Improved efficacy of whole genome amplification from bacterial cells. PMID- 15283199 TI - Nucleic acid stain-dependent single strand conformation polymorphisms. PMID- 15283200 TI - Microsoft Excel spreadsheet-based animal colony management for genetically altered animals. PMID- 15283201 TI - Screening assays for heart function mutants in Drosophila. AB - The rapid life cycle and genetic tractability of Drosophila make it an ideal organism for large-scale genetic screens. Here we describe a novel assay for pupal heart rate and rhythmicity as well as techniques to measure adult cardiac stress response. These assays can be powerfully combined to concurrently screen for both mutations affecting cardiac function and mutations affecting the age dependent decline in adult cardiac stress response. Mutations identified in such screens have the potential to contribute greatly to the understanding of both congenital heart disease and the regulation of age-dependent decline in cardiac function in the human population. PMID- 15283202 TI - Locking of 3' ends of single-stranded DNA templates for improved Pyrosequencing performance. AB - In Pyrosequencing, a DNA strand complementary to a single-stranded DNA (ssDNA) template is synthesized, whereby each incorporated nucleotide yields detectable light, and the light intensity is proportional to the incorporated nucleotides. Correct data interpretation (i.e., signal-to-noise ratio of light intensities) is hampered by artifacts due to the formation of secondary structures of single stranded templates. Critical among these is the looping back of the template's nonbiotinylated 3' end to itself In the resulting structure, the 3' end functions as a primer, the extension of which results in background signals. We present two ways of preventing the self-priming of a template's 3' end: (i) the use of a modified oligonucleotide, called blOligo, which is complementary to the template's 3' end and (ii) the extension of the template's 3' end with a ddNMP. In contrast to unprotected 3' ends of ssDNA templates, causing inconsistent results, we show that protecting the 3' end of an ssDNA template using either blOligos or ddNMP enables the correct interpretation of signals and results in reliable quantification. PMID- 15283203 TI - Quantification of donor microchimerism in sex-mismatched porcine allotransplantation by competitive PCR. AB - The persistence of donor cells in recipient circulation and peripheral tissues post-transplantation has been demonstrated in solid organ allotransplantation and xenotransplantation models. Although this state of microchimerism has been postulated as the basis for graft acceptance, chimerism has not been directly linked to the maintenance of peripheral tolerance or prevention of rejection. Studies have demonstrated that the qualitative presence or absence of donor microchimerism bears no association with graft acceptance. Our preliminary work suggests that there is a threshold chimerism necessary for the induction of donor specific hyporesponsiveness. Because the kinetics of donor cell accumulation and distribution in allograft recipients are largely unknown, quantitative analyses are needed to evaluate chimerism's significance to donor-specific tolerance. We developed a quantitative, competitive PCR assay to precisely measure the amount of chimerism in male to female transplant pairs by targeting the sex-determining region of the Y chromosome (SRY gene). Traditionally, this technique requires that serial known amounts of an SRY-specific competitive template (CT) be coamplified with a constant amount of sample DNA to determine the equivalence point of the relative band intensities of the PCR products. However running a panel of PCRs with CT amounts above and below the equivalence point to generate a standard curve for ever' sample is laborious. Here we describe the generation of a single standard curve that permits the rapid and reliable quantification of microchimerism after coamplification of sample DNA with a single amount of CT. PMID- 15283204 TI - In utero detection of T7 phage after systemic administration to pregnant mice. AB - The phage is used as a scaffold to display recombinant libraries of peptides, which provides the means to rescue and amplify peptides that bind target macromolecules. Many reports showed that the T7 phage display method can be used to obtain a ligand-binding peptidefor tissue-targeted therapies in adult animals. In utero tissue targeting of fetal tissues may help in the correction of many genetic and metabolic diseases. Here we demonstrate the distribution and detection of T7 phage displaying the C-X7-C peptide library in mouse fetal tissues after systemic injection of T7 phage into pregnant mouse tail vein. T7 phage was recovered from fetal tissues 15 min after injection of T7 phage. Our results suggest that T7 phage may be a useful tool in selecting the tissue specific ligand-binding peptide for fetal tissues. This approach may be helpful in designing in utero tissue-targeted therapies. PMID- 15283205 TI - "Lumen digestion" technique for isolation of aortic endothelial cells from heme oxygenase-1 knockout mice. AB - Endothelial cell dysfunction plays a critical role in the pathogenesis of cardiovascular diseases. Gene targeted mutant, knockout. or transgenic mice are widely used in the laboratory investigation of these disorders. We describe a simple and reproducible "lumen digestion" technique to isolate aortic endothelial cells from mice that would be useful for researchers in endothelial cell biology. We used wild-type, homozygote, or heterozygote heme oxygenase-1 null mice from which the aorta is isolated and removed under anesthesia. After cauterizing all the branches, both ends of the aorta are cannulated using an Intramedic PE-20 tube. After flushing the aorta with phosphate-buffered saline (PBS), the lumen is repeatedly instilled (five times) with 50 microL 0.25% trypsin in PBS, incubated for 2 min, and flushed with PBS. The outflow is collected in endothelial cell media with 20% fetal bovine serum. After centrifugation, the endothelial cells in the pellet are resuspended in media and plated in a 24-well tissue culture dish. Following culture for 2 to 3 weeks, the cells demonstrate typical cobblestone appearance, stain positive for the endothelial marker CD31, and are capable of low-density lipoprotein uptake. Following challenge with oxidized lipids, heme oxygenase-1 deficient endothelial cells demonstrate increased susceptibility to cell injury. PMID- 15283206 TI - RNA interference by osmotic lysis of pinosomes: liposome-independent transfection of siRNAs into mammalian cells. AB - The osmotic lysis of pinosomes procedure has been adapted to deliver small interfering RNAs (siRNAs) into cells in culture. Under hypertonic conditions, siRNAs were internalized into pinosomes. A subsequent osmotic shock in hypotonic buffer disrupted the pinosomes and caused the release of siRNAs into the cell cytoplasm. Both steps could be demonstrated directly using fluorescein-labeled siRNAs and confocal laser-scanning microscopy. Uptake by the pinocytosis/osmotic lysis procedure is concentration- and time-dependent. At an siRNA concentration of 0.4 microM, treatment for 40 or 80 min results in silencing efficiencies of 60% and 90%, respectively, after 44 h. A double treatment resulted in approximately equal silencing efficiencies but in reduced viability. This method has been used on a variety of human and murine cell lines including HEK293, HeLa SS6, and SW3T3 cells. Targets such as lamin A/C and Eg5 were effectively silenced. Novel silencing data are provided for Ki67, one of the few reliable prognostic markers for tumor patients. The new procedure avoids certain technical problems encountered with commercial transfection reagents while yielding silencing efficiencies that are comparable to those obtained with liposome mediated siRNA transfection. PMID- 15283207 TI - Construction of recombinant fowlpox viruses carrying multiple vaccine antigens and immunomodulatory molecules. AB - Here we describe plasmid vectors and selection protocols developed to allow the construction of recombinant fowlpox viruses (rFPVs) with up to three insertions of foreign DNA in the viral genome. Transient dominant selection allows the construction of recombinant viruses that do not retain the selection markers and can therefore be used for the insertion of additional genes at other sites in the viral genome. A SYBR Green real-time PCR sequence detection assay was applied to the identification of recombinant viruses from individual plaques, eliminating the need for amplification and hybridization from the transient dominant protocol and resulting in significant savings in time at each round of plaque purification. Dominant selection techniques allow more rapid recombinant virus construction; however, as the markers are retained along with the gene of interest, they can only be used to generate the final recombinant. rFPV vaccines constructed using these techniques have reached preclinical nonhuman primate and phase I human clinical trials in prime/boost vaccination studies as human immunodeficiency virus (HIV) therapeutic andprophylactic vaccines. PMID- 15283208 TI - Validation of housekeeping genes for normalizing RNA expression in real-time PCR. AB - Analysis of RNA expression using techniques like real-time PCR has traditionally used reference or housekeeping genes to control for error between samples. This practice is being questioned as it becomes increasingly clear that some housekeeping genes may vary considerably in certain biological samples. We used real-time reverse transcription PCR (RT-PCR) to assess the levels of 13 housekeeping genes expressed in peripheral blood mononuclear cell culture and whole blood from healthy individuals and those with tuberculosis. Housekeeping genes were selected from conventionally used ones and from genes reported to be invariant in human T cell culture. None of the commonly used housekeeping genes [e.g., glyceraldehyde-phosphate-dehydrogenase (GAPDH)] were found to be suitable as internal references, as they were highly variable (>30-fold maximal variability). Furthermore, genes previously found to be invariant in human T cell culture also showed large variation in RNA expression (>34-fold maximal variability). Genes that were invariant in blood were highly variable in peripheral blood mononuclear cell culture. Our data show that RNA specifying human acidic ribosomal protein was the most suitable housekeeping gene for normalizing mRNA levels in human pulmonary tuberculosis. Validations of housekeeping genes are highly specific for a particular experimental model and are a crucial component in assessing any new model. PMID- 15283209 TI - Nonspecific enhancement of gene expression by compounds identified in high throughput cell-based screening. PMID- 15283210 TI - Aptamer-dependent full-length cDNA synthesis by overlap extension PCR. AB - Sequencing of the human genome in combination with computational annotation has provided tremendous data on predicted genes. However, for most of them, no corresponding cDNAs are available yet. Furthermore, even where cDNA clones were obtained, gene transcripts often have many different splice variants that are not covered by current gene collections. For direct synthesis of cDNA clones corresponding to predicted genes, new splice variants, or any other gene of interest, we established optimal PCR conditions for the direct amplification of exons from genomic DNA, which require a specific Taq aptamer. PCR products comprising differently tagged exons were concatenated by overlap extension into full-length cDNAs. To prove the effectiveness of the approach, the 1900-bp full length open reading frame of the human mitochondrial aldehyde dehydrogenase (ALDH2) gene was synthesized in a two-step reaction comprising all 13 exons. Thus, our conditions are of general value for in vitro synthesis of cDNAs and alternative splice variants from genomic DNA. PMID- 15283211 TI - Array comparative genomic hybridization with cyanin cis-platinum-labeled DNAs. AB - Fluorescent cis-platinum compounds that react with the N7 atom of guanine are useful for labeling nucleic acids influorescence hybridization applications. Here we report that cyanin (CyN) cis-platinum labeling of DNA samples for array comparative genomic hybridizations (arrayCGH) can be achieved reproducibly and reliably. We demonstrate that degrees of labeling of approximately 1% of all nucleotides in test and reference DNA samples with CyN3- and CyN5-cis-platinum produces arrayCGH signal-to-background ratios ranging from 30 to 40. The arrayCGH results achieved during analyses of mouse and human tumor samples were comparable to those achieved using enzymatic labeling. Thus, we conclude that Cy-cis platinum labeling is an alternative to enzymatic labeling for arrayCGH. PMID- 15283212 TI - Assessment of multiple displacement amplification in molecular epidemiology. AB - Well-characterized epidemiological resources are generated with great effort, yet associated patient DNA samples can be limiting. The efficacy of the whole genome amplification (WGA) method, termed multiple displacement amplification (MDA), was assessed for detecting heterozygous sequence variants, mutation scanning, and PCR for challenging segments. Fifteen common polymorphisms from 10 genes located on 8 chromosomes were genotyped by direct sequencing of 300 PCR products from 115 high quality MDA-amplified DNA samples extracted by different methods. The GC content of these analyzed segments ranges from 30% to 69%. Genotyping results demonstrate 100% accuracy. For heterozygotes, the relative intensity of peaks generated by the two alleles is highly similar for genomic and MDA-amplified genomic DNA, independent of GC content. In contrast, one of four heterozygous loci was mistyped when lower quality MDA-amplified DNA samples were used. The results of single-stranded conformation polymorphism (SSCP)-type of mutation scanningfor seven MDA-amplified DNA samples in four genes were concordant with the genomic DNA samples. PCR on MDA-amplified DNA was routinely successful for challenging 10 and 12-kb segments with GC content ranging from 30% to 80%, demonstrating that rather long segments, which are difficult to amplify with PCR, are amplified well with MDA. These results suggest that MDA is an effective method of WGA with utility in molecular epidemiology. Quality control of the MDA-amplified DNA is critical for high performance. PMID- 15283213 TI - Major ocular complications after organ transplantation. PMID- 15283214 TI - Posterior capsule opacification: a review of the aetiopathogenesis, experimental and clinical studies and factors for prevention. AB - Posterior capsule opacification (PCO, secondary cataract, after cataract) is a nagging postsurgical complication following extracapsular cataract surgery (ECCE) and intraocular lens (IOL) implantation. PCO should be eliminated since it has deleterious sequelae and Neodynium: Yttrium Aluminium Garnet (Nd: YAG) laser treatment often is an unnecessary financial burden on the health care system. PCO following cataract surgery could be a major problem, since patient follow-up is difficult and the Nd:YAG laser is not always available. Advances in surgical techniques, IOL designs/biomaterials have been instrumental in bringing about a gradual and unnoticed decrease in the incidence of PCO. We strongly believe that the overall incidence of PCO and hence the incidence of Nd:YAG laser posterior capsulotomy is now rapidly decreasing - from 50% in the 1980s and early 1990s to less than 10% currently. Superior tools, surgical procedures, skills and appropriate IOL designs have all helped to significantly reduce this complication. In this article, we review the aetio pathogenesis, experimental and clinical studies and propose surgical and implant-related factors for PCO prevention. Careful application and utilisation of these factors by surgeons could lead to a significant reduction is secondary cataract, the second most common cause of visual loss worldwide. PMID- 15283215 TI - Ex-vivo potential of cadaveric and fresh limbal tissues to regenerate cultured epithelium. AB - PURPOSE: To evaluate and compare the ex-vivo growth potential and formation of cultured corneal epithelium from residual corneo-limbal rings obtained from the operating room after penetrating keratoplasty, and fresh limbal tissues from patients undergoing routine cataract surgery. METHODS: With the approval of the Institutional Review Board and informed consent from patients, 1-2 mm of limbal tissues from 15 patients and 31 tissues from the cadaveric limbal ring preserved in MK medium (16 tissues) and Optisol (15 tissues) were used for the study. Donor data included age, time lapse between death and collection, collection and preservation and preservation and culture. Tiny bits of the limbal tissue were explanted on the de-epithelialised human amniotic membrane prepared following standard guidelines, and cultured using Human Corneal Epithelial cell medium. Radial growth from the explant was observed and measured by phase contrast microscopy over 2-4 weeks. After adequate confluent growth, whole mount preparation of the membrane was made and stained with haematoxylin and eosin. Part of the membrane was fixed in formalin and processed for routine histologic examination. The sections were stained with haematoxylin and eosin. RESULTS: Forty-six tissues were evaluated from 42 eyes (15 from patients, 31 from cadaveric eyes) with a mean age of 55.3 years +/- 21.23 years (range 18 years - 110 years). The growth pattern observed was similar in all the positive cases with clusters of cells budding from the explant over 24-72 hours, and subsequent formation of a monolayer over the next 2-3 weeks. The stained whole mount preparation showed a radial growth of cells around explants with diameter ranging from 5 to 16mm. Histologic evaluation of the membrane confirmed the growth of 2-3 cell-layered epithelium over the amniotic membrane. Cultivated epithelium around explant cell cultures was observed in 100% (15/15) of limbal tissue obtained from patients, as against 56% (9/16) of MK medium preserved tissues and 46.7% (7/15) of Optisol preserved tissues. This was statistically significant (P=0.0131) There was no significant statistical difference in the growth properties, i.e, the mean percentage of fragments showing growth (P=0.229) or the mean diameter of growth (P=0.479) in the cultures obtained from fresh and preserved tissues. The time lapse at various stages between death and utilisation and donor age had no significant influence on the growth potential of the limbal tissues. CONCLUSION: The potential for generating cultured corneal epithelium from fresh limbal tissues obtained from living subjects is higher than that observed with preserved tissues. It would also be worthwhile to address the factors that could further enhance the proliferative potential of the cadaveric tissues obtained from eye banks. PMID- 15283216 TI - Pattern of uveitis in a referral eye clinic in north India. AB - PURPOSE: To report the pattern of uveitis in a north Indian tertiary eye center. METHODS: A retrospective study was done to identify the pattern of uveitis in a uveitis clinic population of a major referral center in north India from January 1996 to June 2001. A standard clinical protocol, the "naming and meshing" approach with tailored laboratory investigations, was used for the final diagnosis. RESULTS: 1233 patients were included in the study; 641 (51.98%) were males and 592 (48.01%) females ranging in age from 1.5 to 75 years. The anterior uveitis was seen in 607 patients (49.23%) followed by posterior uveitis (247 patients, 20.23 %), intermediate uveitis (198 patients, 16.06%) and panuveitis (181 patients, 14.68%). A specific diagnosis could be established in 602 patients (48.82%). The infective aetiology was seen in 179 patients, of which tuberculosis was the commonest cause in 125 patients followed by toxoplasmosis (21 patients, 11.7%). Non-infectious aetiology was seen in 423 patients, of which ankylosing spondylitis was the commonest cause in 80 patients followed by sepigionous choroidopathy (62 patients, 14.65%). CONCLUSION: Tuberculosis and toxoplasmosis were the commonest form of infective uveitis, while ankylosing spondylitis and serpiginous choroidopathy were commonly seen as the non-infective causes of uveitis in North India. PMID- 15283217 TI - Sterile endophthalmitis in vitrectomised eyes due to suspected heat resistant endotoxins in the infusion fluid. AB - PURPOSE: To report on to the possibility of development of severe postoperative sterile endophthalmitis due to heat-resistant bacterial endotoxins in commercially available infusion fluids METHODS: A case study of 4 eyes that had previously undergone vitreoretinal surgery, which developed clinical endophthalmitis within 18 hours of surgery and two eyes that had vitreous surgery with intraocular gas and did not develop clinical endophthalmitis following intraocular surgery on three consecutive operative days RESULTS: The vitreous samples were sterile, both for bacteria and fungi. The only common supply in all cases was a new batch of Ringer's lactate infusion fluid. Though the Ringer's lactate solutions in the same batch were also sterile, the infusion fluids contained abnormally high levels of bacterial endotoxins detected by gel clot method. CONCLUSION: Commercially available infusion fluid may be sterile, yet contain endotoxins from killed bacteria. This could cause postoperative sterile endophthalmitis. PMID- 15283218 TI - Corneal endothelial safety of intracameral preservative-free 1% xylocaine. AB - PURPOSE: To evaluate the effect of intracameral preservative-free 1% xylocaine on the corneal endothelium as an adjuvant to topical anaesthesia during phacoemulsification and Acrysof foldable IOL implantation. MATERIAL & METHODS: This is a prospective, controlled, randomised, double-masked study. 106 patients with soft to moderately dense (Grade 1-3) senile cataract and corneal endothelial cell density of >1500/mm2 were randomised to the xylocaine group (n=53) and control group(n=53). Central endothelial specular microscopy and ultrasound corneal pachymetry were performed preoperatively. On the first postoperative day the eyes were evaluated for corneal oedema and Descemet's folds. Ultrasound corneal pachymetry was performed at 1, 3 and 12 months. Specular microscopy was performed at 3 and 12 months. Cell loss was expressed as a percentage of preoperative cell density. Six patients could not complete one year follow-up. Chi-square and paired t test (2 tail) statistical tests were applied for analysis. RESULTS: Four (7.54%) patients in the xylocaine group and 5 (9.43%) in the control group had a few Descemet's folds associated with mild central stromal oedema. Corneal thickness increased from 549.3micro +/- 37.2micro to 555.5micro +/- 36.5micro in the xylocaine group and from 553.1micro +/- 36.2micro to 559.3micro +/- 40.5micro in the control group at the one-month postoperative visit. Thickness returned to the preoperative level in xylocaine group 549.6micro +/- 34.5micro and control group 554.7micro +/- 41.1micro at three months. (P=0.484) The percentage of cell loss was 4.47 +/- 2.53% in the xylocaine group and 4.49 +/- 3.09% in the control group at one year. (P=0.97) CONCLUSION: Intracameral preservative-free 1% xylocaine does not appear to affect corneal endothelium adversely during phacoemulsification. PMID- 15283219 TI - Determination of carbonyl group content in plasma proteins as a useful marker to assess impairment in antioxidant defense in patients with Eales' disease. AB - PURPOSE: Formation of protein carbonyl groups is considered an early biomarker for the oxidant/antioxidant barrier impairment in various inflammatory diseases. We evaluated the intensity of free radical reactions in patients with Eales' disease, an idiopathic inflammatory condition of the retina. METHODS: Twenty patients with Eales' disease in active vasculitis stage, 15 patients with Eales' disease in healed vasculitis stage and 20 healthy control subjects were recruited for the study. Plasma protein carbonyl groups,plasma glutathione (GSH) superoxide dismutase (SOD) activity and thiobarbituric acid reactive substances (TBARS) were determined in erythrocytes. RESULTS: Plasma protein carbonyl content was elevated by a factor of 3.5 and 1.8 respectively in active and healed vasculitis stages. The increase of carbonyl group content in active and healed stage of patients with Eales' disease correlated with diminished SOD activity and GSH content. There was also increased accumulation of TBARS in active and healed vasculitis stages of Eales' disease, and this correlated with diminished SOD activity. CONCLUSION: Our results showed that protein carbonyl group content increases with severity of Eales' disease. The increase in carbonyl content correlated with diminished antioxidant status. This confirms an earlier report that free radical mediated tissue damage occurs in Eales' disease. The determination of protein carbonyl content may be used as a simple biomarker to monitor the efficacy of antioxidant supplementation in controlling retinal vasculitis in patients with Eales' disease. PMID- 15283220 TI - Trace elements iron, copper and zinc in vitreous of patients with various vitreoretinal diseases. AB - PURPOSE: To measure the concentrations of iron, copper and zinc in human vitreous and to interpret their levels with various vitreoretinal diseases like proliferative diabetic retinopathy, retinal detachment, intraocular foreign body, Eales' disease and macular hole. METHODS: Undiluted vitreous fluid collected during pars plana vitrectomy was used to measure trace elements using an atomic absorption spectrophotometer. RESULTS: The level of vitreous iron increased threefold in Eales' disease (1.85 +/- 0.36 pg/ml), 2.5-fold in proliferative diabetic retinopathy (1.534 +/- 0.17 pg/ml) and 2.3-fold in eyes with intraocular foreign body (1.341 +/- 0.25 pg/ml) when compared with macular hole (0.588 +/- 0.16 pg/ml). This was statistically significant (P < 0.05). Zinc was found to be low in Eales' disease (0.57 +/- 0.22 pg/ ml) when compared with other groups, though the difference was not statistically significant. CONCLUSION: The increased level of iron with decreased zinc content in Eales' disease confirms the earlier reported oxidative stress mechanism for the disease. In proliferative diabetic retinopathy and intraocular foreign body the level of iron increases. This is undesirable as iron can augment glycoxidation, which can lead to increased susceptibility to oxidative damage, in turn causing vitreous liquefaction, posterior vitreous detachment and ultimately retinal detachment and vision loss. PMID- 15283221 TI - Optic disc imaging by Heidelberg retinal tomogram in congenital optic disc anomaly. AB - We evaluated two cases of congenital optic disc anomaly with the Heidelberg Retinal Tomograph (HRT) that could be mistaken for glaucomatous optic disc. One was an optic disc coloboma with a visual field defect and the other had an optic disc pit without a visual field defect. HRT was abnormal only in the eye with optic disc pit with normal fields. While HRT can be a valuable adjunct to disc evaluation and follow-up, it cannot be used in isolation in the differentiation of abnormal from normal optic discs. PMID- 15283222 TI - Low endothelial cell count and clear corneal grafts. AB - Specular microscopic study on clear corneal grafts indicates that at times surprisingly low endothelial cell density can maintain the grafted cornea in a relatively dehydrated state. The critical limit of the endothelial cell count for corneal decompensation is thought to be 700 cells/mm2. This communication reports 13 cases of clear corneal graft with endothelial cell count below 700 cells/mm2. PMID- 15283223 TI - Seborrheic keratosis of the conjunctiva. AB - Seborrheic keratosis can simulate a malignant melanoma and should form the differential diagnosis of a malignant melanoma. Histopathology is confirmatory. PMID- 15283224 TI - Recurrent mucinous carcinoma of the eyelid. AB - A case of recurrent mucinous carcinoma of the lid in a 40-year-old male is reported. Clinical differential diagnosis and histopathologic features are discussed. PMID- 15283225 TI - Late recurrent uveitis after phacoemulsification. AB - It is now assumed that recurrent late onset uveitis after phacoemulsification with intraocular lens (IOL) is due to indolent infection. Fifteen such cases were observed after uncomplicated phacoemulsification with-in-the-bag IOL implant. These cases were considered noninfective and treated medically with good visual recovery. PMID- 15283226 TI - Uveal metastasis from a bronchial carcinoid tumour. AB - A rare case of bronchial carcinoid tumour metastasis to the ciliary body and the choroid with clinical, diagnostic and histopathological correlation is reported. PMID- 15283227 TI - Perceptions of eye diseases and eye care needs of children among parents in rural south India: the Kariapatti Pediatric Eye Evaluation Project (KEEP). AB - We conducted 24 focus group discussions for parents and grandparents as part of a population-based survey of ocular morbidity to determine awareness and perceptions of eye diseases in children among parents and guardians of children in a rural south Indian population. Focus group discussions were conducted separately for mothers, fathers and grandparents. They were audiotaped and subsequently transcribed to the local language and English. Content analysis of the focus group discussions was done to identify key concepts, and this yielded five broad areas of interest relating to awareness and attitudes towards: 1) eye problems in children, 2) specific eye diseases in children, 3) vision problems in children, 4) existing health practices, and 5) utilization of services. Vision impairment did not figure in the top ten eye problems cited for children. There was a predominant belief that children below 4 years should not wear spectacles. Strabismus was considered as untreatable and was seen as a sign of good luck. Differing advice provided by the medical community for the same condition was an issue. The discussions also brought out that eye doctors were approached last for eye care, after traditional healers and general physicians. The discussions raise several issues of relevance that eye care programs need to address for better community involvement with programs. This will require a far greater focus than the current curative focus adopted by most programs. PMID- 15283228 TI - Laser range finder can cause retinal injury. PMID- 15283229 TI - Fluticasone propionate raises IOP in susceptible individuals. PMID- 15283230 TI - Rhino-orbito-cerebral mucormycosis. PMID- 15283231 TI - Prolapsed intraocular aspergilloma masquerading as malignant melanoma. PMID- 15283232 TI - Much ado about ciprofloxacin and Acinetobacter. PMID- 15283233 TI - The Sweep-VEP: a faster estimation of visual acuity in preverbal children. PMID- 15283234 TI - A vaccine against coccidioidomycosis is justified and attainable. AB - Coccidioides is a fungal pathogen of humans which can cause a life-threatening respiratory disease in immunocompetent individuals. Recurrent epidemics of coccidioidal infections in Southwestern United States has raised the specter of awareness of this soil-borne microbe, particularly among residents of Arizona and Southern California, and has galvanized research efforts to develop a human vaccine against coccidioidomycosis. In this review, we discuss the rationale for such a vaccine, examine the features of host innate and acquired immune response to Coccidioides infection, describe strategies used to identify and evaluate vaccine candidates, and provide an update on progress toward development of a vaccine against this endemic pathogen. PMID- 15283235 TI - Molecular cloning and characterization of a cDNA encoding the Paracoccidioides brasiliensis 135 ribosomal protein. AB - A 630 bp cDNA encoding an L35 ribosomal protein of Paracoccidioides brasiliensis, designated as Pbl35, was cloned from a yeast expression library. Pbl35 encodes a polypeptide of 125 amino acids, with a predicted molecular mass of 14.5 kDa and a pI of 11.0. The deduced PbL35 shows significant conservation in respect to other described ribosomal L35 proteins from eukaryotes and prokaryotes. Motifs of ribosomal proteins are present in PbL35, including a bipartite nuclear localization signal (NLS) that could be related to the protein addressing to the nucleolus for the ribosomal assembly. The mRNA for PbL35, about 700 nucleotides in length, is expressed at a high level in P. brasiliensis. The PbL35 and the deduced amino acid sequence constitute the first description of a ribosomal protein in P. brasiliensis. The cDNA was deposited in GenBank under accession number AF416509. PMID- 15283236 TI - Ascospore-derived isolate of Arthroderma benhamiae with morphology suggestive of Trichophyton verrucosum. AB - Sixty-one ascospores were isolated from an ascocarp produced by the mating of two Arthroderma benhamiae strains, RV 26678 and KMU4169, that differed in their mitochondrial DNA (mtDNA) restriction fragment length polymorphism (RFLP) patterns and in the sequences of their nuclear ribosomal internal transcribed spacer (ITS) regions. RV 26678 is a genetically typical A. benhamiae isolate, while KMU4169, though morphologically indistinguishable from A. benhamiae, is an isolate with a deviating ITS sequence and with a mtDNA RFLP profile identical to that of T. verrucosum. One of the 61 progeny ascospores formed a colony, KMU5-46, that was quite different from both parental isolates. KMU5-46 is a faviform colony morphologically similar to Trichophyton verrucosum, although its mtDNA RFLP patterns and ITS sequences were identical to those of A. benhamiae parental strain RV 26678. The morphological alteration manifested in KMU5-46, as well as this isolate's complete loss of sexual response, indicates the possibility that the asexual T. verrucosum and the sexual A. benhamiae are conspecific. PMID- 15283237 TI - Molecular epidemiology of Cryptococcus neoformans isolates from AIDS patients of the Brazilian city, Rio de Janeiro. AB - A high biodiversity of Cryptococcus neoformans isolates is known to exist in some Brazilian urban areas, raising the possibility that patients may encounter multiple inoculum sources in their daily life. C. neoformans isolates from two groups of AIDS patients with cryptococcosis from Rio de Janeiro were studied by polymerase chain reaction (PCR) fingerprinting and randomly amplified polymorphic DNA (RAPD) analysis. The first group contained 60 serial isolates obtained from 19 patients over periods ranging from 18 to 461 days; the intent was to determine whether the original strain persisted or whether reinfection with a new strain occurred. The second group was made up of 22 isolates from 11 patients, and consisted of a pair of isolates collected from blood and cerebrospinal fluid from each patient either before or shortly after treatment was initiated. The aim was to determine if the patient was infected by different strains simultaneously. All isolates were subtyped by PCR fingerprinting, using minisatellite (M13), and microsatellite [(GACA)4 and (GTG)5] specific primers, and RAPD analysis employing the combined primers 5SOR and CN1. The majority of isolates were C. neoformans var. grubii, specifically, molecular types VNI or VNII, but numerous distinguishable subtypes were found. Only three isolates were C. n. var. gattii (molecular types VGI or VGII). Except in two cases, all isolates obtained from the same patient showed identical PCR profiles independent of time of isolation or body site. Almost all patients, however, carried unique genotypes not found in any other patient. Our results confirm that persistent cryptococcal infection is caused by relapse rather than reinfection, but they also show that in exceptional cases, patients may be infected with more than one C. neoformans strain. PMID- 15283238 TI - Determination of keratin degradation by fungi using keratin azure. AB - Azure dye-impregnated sheep's wool keratin (keratin azure) was incorporated in a high pH medium and overlaid on a keratin-free basal medium. The release and diffusion of the azure dye into the lower layer indicated production of keratinase. Fifty-eight fungal taxa, including 49 members of the Arthrodermataceae, Gymnoascaceae and Onygenaceae (Order Onygenales), were assessed for keratin degradation using this method. The results were comparable to measures of keratin utilization reported in studies using tests based on the perforation or erosion of human hair in vitro. PMID- 15283239 TI - Molecular cloning and characterization of a cDNA encoding the N-acetyl-beta-D glucosaminidase homologue of Paracoccidioides brasiliensis. AB - A cDNA encoding the N-acetyl-beta-D-glucosaminidase (NAG) protein of Paracoccidioides brasiliensis, Pb NAG1, was cloned and characterized. The 2663 nucleotide sequence of the cDNA consisted of a single open reading frame encoding a protein with a predicted molecular mass of 64.73 kDa and an isoeletric point of 6.35. The predicted protein includes a putative 30-amino-acid signal peptide. The protein as a whole shares considerable sequence similarity with 'classic' NAG. The primary sequence of Pb NAG1 was used to infer phylogenetic relationships. The amino acid sequence of Pb NAG1 has 45, 31 and 30% identity, respectively, with homologous sequences from Trichoderma harzianum, Aspergillus nidulans and Candida albicans. In particular, striking homology was observed with the active site regions of the glycosyl hydrolase group of proteins (family 20). The expected active site consensus motif G X D E and catalytic Asp and Glu residues at positions 373 and 374 were found, reinforcing that Pb NAG1 belongs to glycosyl hydrolase family 20. The nucleotide sequence of Pb nag1 and its flanking regions have been deposited, along with the amino acid sequence of the deduced protein, in GenBank under accession number AF419158. PMID- 15283240 TI - Reservoir of Candida albicans infection in a vascular bypass graft demonstrates a stable karyotype over six months. AB - We retrospectively analyzed five Candida albicans isolates from two infection episodes in a single patient 6 months apart. Using contour-clamped homogeneous field electrophoresis (CHEF), random amplified polymorphic DNA (RAPD) fingerprinting, and restriction fragment length polymorphism (RFLP) complex probe 27A as means of molecular typing, we demonstrate an unvarying genotype amongst the infection-causing C. albicans strains. Several months later, the patient yielded C. glabrata in a yeast survey of oral and rectal sites. The preponderance of C. glabrata and lack of C. albicans isolated from normal flora sites suggests that this patient harbored the prior C. albicans bloodstream isolate on a Gore Tex graft for 6 months prior to the second episode of fungemia. PMID- 15283241 TI - Chromoblastomycosis caused by Rhinocladiella aquaspersa. AB - An unusual case is presented featuring chromoblastomycosis lesions localised in three different sites. The patient was a 52-year-old male farm worker from Barra do Corda, State of Maranhao, Brazil, who had had the disease for 2 years. Physical examination revealed extensive plaques situated on the left leg, left arm, forehead, and on the left side of the face. Direct examination of biopsies showed numerous sclerotic cells. The fungus was recovered in culture and identified on the basis of the characteristic conidiation as Rhinocladiella aquaspersa. PMID- 15283242 TI - Onychomycosis caused by an isolate conforming to the description of Trichophyton raubitschekii. AB - Trichophyton raubitschekii is currently regarded as a synonymous name betokening a variant form of Trichophyton rubrum. Nonetheless, isolates conforming to this morphotaxonomic concept have morphological, physiological and clinical features very different from those of typical T. rubrum. Isolates are mainly obtained from subjects originating from certain tropical and subtropical countries, and are mainly obtained from upper body skin infections, rarely from onychomycosis. In this paper the authors report the first known Italian case of onychomycosis caused by such an isolate. The patient, a male student from Cameroon, had a typical fingernail tinea unguium, without any other sign of skin or nail infection. 'T. raubitschekii' was identified on morphological and physiological grounds by the following features: velvety colony surface, brownish pigment, abundant macroconidia and microconidia, and positive urease activity. Such isolates may prove very difficult to identify correctly, especially in areas like Italy where T. rubrum is normally seen only as isolates presenting a strongly differing phenotype. PMID- 15283243 TI - Cutaneous infection caused by Alternaria in patients receiving tacrolimus. AB - Tacrolimus, an immunosuppressant used in organ transplant surgery, is inhibitory to some medically important fungi but also may obstruct azole monotherapy in the immunocompromised patient. We observed a case of cutaneous phaeohyphomycosis caused by Alternaria alternata in a liver transplant recipient who had been under tacrolimus immunosuppression for 6 months post-transplantation. At the onset of the infection, the patient presented with an increased whole-blood tacrolimus level. After a simple surgical excision the patient was discharged from the hospital without antifungal treatment but with an adjusted tacrolimus dosage. Literature review on fungal infections in patients receiving tacrolimus suggested these patients experience cutaneous and deep mould infections that are more frequent, severe and therapy-refractory than those seen in patients with other types of immunosuppression. PMID- 15283244 TI - Antifungal susceptibility of 50 Candida isolates from invasive mycoses in Chile. AB - We determined the antifungal susceptibility of 50 Candida isolates from invasive mycoses in intensive care unit patients in Chile. Candida albicans (27 isolates) and C. parapsilosis (12 isolates) were the most common etiologic species. Candida glabrata (five isolates) was isolated only from children. Our data are consistent with those of recent Brazilian and Argentinean studies but differ from those of some US, Canadian and Norwegian studies, in which the relatively azole-resistant C. glabrata was the predominant non-C. albicans species seen. All isolates in this study were susceptible to amphotericin B. Itraconazole and fluconazole resistance were observed in 6 and 4% of the isolates, respectively. PMID- 15283245 TI - Histoplasma capsulatum yeast cells attach and agglutinate human erythrocytes. AB - The ability of yeast cells of Histoplasma capsulatum to attach and agglutinate human erythrocytes has been described. This is the first report involving these yeasts in the hemagglutination phenomenon. Results revealed that the yeast cells were able to bind to erythrocytes irrespective of blood groups and to agglutinate them when a high density of yeast cells was used. Assays on the inhibition of yeast attachment to erythrocytes were also performed, using sugar-treated yeast cells. Results indicate that galactose (Gal), mainly the beta-anomer, specially inhibited yeast attachment. Disaccharides (Gal-derivatives) and glycosaminoglycans containing Gal residues, mainly chondroitin sulfate C, promote this type of inhibition. In addition, preliminary data of inhibition assays also involved a probable ionic strength driven mechanism mediated by sialic acid and heparan sulfate, suggesting that yeast binding to erythrocytes could be associated with negative charges of both molecules. PMID- 15283246 TI - Quantitative electron probe microanalysis of rough targets: testing the peak-to local background method. AB - Rough samples with topography on a scale that is much greater than the micrometer dimensions of the electron interaction volume present an extreme challenge to quantitative electron beam x-ray microanalysis with energy-dispersive x-ray spectrometry. Conventional quantitative analysis procedures for flat, bulk specimens become subject to large systematic errors due to the action of geometric effects on electron scattering and the x-ray absorption path compared with the ideal flat sample. The best practical approach is to minimize geometric effects through specimen reorientation using a multiaxis sample stage to obtain the least compromised spectrum. When rough samples must be analyzed, corrections for geometric factors are possible by the peak-to-local background (P/B) method. Correction factors as a function of photon energy can be determined by the use of reference background spectra that are either measured locally or calculated from pure element spectra and estimated compositions. Significant improvements in accuracy can be achieved with the P/B method over conventional analysis with simple normalization. PMID- 15283247 TI - Metrics for focusing in extremely noisy scanning electron microscopy condition. AB - Finding a best focused image in very noisy condition is an extremely difficult task for the SEM user. If a performance, which is much higher than that of an expert in focusing, can be achieved in a computer-controlled scanning electron microscope (SEM), it will be very helpful for our field due to the many possible applications. To accomplish this work, we employ a powerful metric-the covariance obtained by a special scanning method. It can select the best focused image from a series of SEM images acquired by altering the focus of the objective lens under an extremely noisy SEM image condition. The noise immunity of the present method is quantitatively evaluated, and it is further improved based on the obtained evaluation result. PMID- 15283248 TI - Practical method for high-resolution imaging of polymers by low-voltage scanning electron microscopy. AB - Morphologic characterization of polymers by scanning electron microscopy (SEM) is often made difficult by their sensitivity to electron beam damage. We describe here a specimen preparation method for the imaging of polymer blends by low voltage SEM (LV-SEM) that improves their stability in the electron beam and hence facilitates focusing and recording of high magnification images. Its application to nanosized core-shell latexes embedded in a polymethylmethacrylate matrix and semi-crystalline polypropylene/ethylene-propylene rubber blends is discussed. PMID- 15283249 TI - A method for imaging single clay platelets by scanning electron microscopy. AB - A method for preparing and observing clay platelets for size and shape analysis using scanning electron microscopy (SEM) was developed. Samples of the clay platelets were prepared by polyelectrolyte-assisted adsorption onto a pyrolytic graphite surface. The use of graphite as a substrate was advantageous because of the low number of secondary electrons emitted from it during imaging by SEM. The resulting low background noise allowed the emission from the approximately 1 nm thick clay sheets to be clearly visualized. Images of centrifuged montmorillonite showed large exfoliated platelets with lateral dimensions between 200 and 600 nm. In contrast, uncentrifuged montmorillonite appeared to contain a large amount of unexfoliated clusters. Although it was not possible to obtain high-quality images of the smaller sheets of Laponite RD, the images of this material did contain size features comparable to the approximately 30 nm2 size reported previously using light scattering, as well as transmission electron and atomic force microscopies. PMID- 15283250 TI - Image signal-to-noise ratio estimation using the autoregressive model. AB - In the last two decades, a variety of techniques for signal-to-noise ratio (SNR) estimation in scanning electron microscope (SEM) images have been proposed. However, these techniques can be divided into two groups: first, SNR estimators of good accuracy, but based on impractical assumptions; second, estimators based on realistic assumptions but of poor accuracy. In this paper we propose the implementation of autoregressive (AR)-model interpolation as a solution to the problem. Unlike others, the proposed technique is based on a single SEM image and offers the required accuracy and robustness in estimating SNR values. PMID- 15283251 TI - Reduction in acquisition time of scanning electron microscopy image using complex hysteresis smoothing. AB - Complex hysteresis smoothing (CHS), which was developed for noise removal of scanning electron microscopy (SEM) images some years ago, is utilized in acquisition of an SEM image. When using CHS together, recording time can be reduced without problems by about one-third under the condition of SEM signal with a comparatively high signal-to-noise ratio (SNR). We do not recognize artificiality in a CHS-filtered image, because it has some advantages, that is, no degradation of resolution, only one easily chosen processing parameter (this parameter can be fixed and used in this study), and no processing artifacts. This originates in the fact that its criterion for distinguishing noise depends simply on the amplitude of the SEM signal. The automation of reduction in acquisition time is not difficult, because CHS successfully works for almost all varieties of SEM images with a fairly high SNR. PMID- 15283252 TI - [Formation mechanisms of the fibrous capsule around hepatic and splenic hydatid cyst]. AB - OBJECTIVE: To investigate the formative mechanisms of the fibrous capsule around hepatic and splenic hydatid cyst. METHODS: HE stain was used to observe the histopathologic changes of the fibrous capsules around hepatic and splenic hydatid cysts and the adjacent parenchyma. The expression of collagen IV, fibronectin (FN), laminin (LN), transforming growth factor-beta1 (TGF-beta1) and tumor necrosis factor-alpha (TNF-alpha) were detected by using immunohistochemistry or in situ hybridization in fibrous capsules around 40 hepatic hydatid cysts and 15 splenic hydatid cysts including adjacent parenchyma. RESULTS: A special delaminated phenomenon was observed in the fibrous capsules around hepatic hydatid cysts. Granuloma-like pathologic changes were found in the fibrous capsules near worm and there were many depressed Glisson capsules and hepatic vein system in the fibrous capsules near hepatic parenchyma while these phenomena could not be observed in splenic hydatid cysts. All parameters were highly expressed in the fibrous capsules near hepatic parenchyma and the difference between the two fibrous capsules was significant (P<0.01). CONCLUSION: The mechanisms of forming fibrous capsule around hepatic and splenic hydatid cysts are different. The fibrous capsule around hepatic hydatid cyst is a granuloma-like structure, covered by the pressed Glisson system and hepatic vein system with a small gap between them; while the fibrous capsule around splenic hydatid cyst is formed by granuloma-like tissue covering the worm and there is no gap between the capsule and the splenicparenchyma. PMID- 15283253 TI - [A case of echinococcosis in Liuzhou City of Guangxi]. PMID- 15283254 TI - [Construction of DNA vaccine of Trichinella spiralis muscle larvae and its expression in mice]. AB - OBJECTIVE: To construct and express recombinant plasmid containing the structural gene encoding Mr 31 000 antigen of Trichinella spiralis muscle larvae. METHODS: The target gene TspE1 was amplified by RT-PCR, cloned into pUC18 vector, and sub cloned into the eukaryotic expression vector pcDNA3. BALB/c mice were immunized with the purified recombinant plasmid pcDNA3-TspE1 by gene-gun bombardment. The expression of recombinant plasmid in the skin tissue was observed by HE staining and immunohistochemical staining. RESULTS AND CONCLUSION: The recombinant plasmid pcDNA3-TspE1 was successfully constructed and expressed in the BALB/c mice. PMID- 15283255 TI - [A case report of Phthirus pubis infestation of eyelids in an infant]. PMID- 15283256 TI - [Effect of ecological treatment by Musca domestica larvae to pig manure on the oviposition and larvae hatching rate]. AB - OBJECTIVE: To observe the change of housefly (Musca domestica) breeding in the pig manure treated ecologically with its larvae. METHODS: The number of eggs and the hatching rate of larvae in the treated manure were compared with that in the untreated manure. RESULTS: The number of eggs laid in the treated manure accounted for only 17.7% of the total eggs, while those in the untreated manure accounted for 82.3%. The hatching rate in the treated manure was 41.4 %, but 85.1% in the untreated manure. CONCLUSION: There is a significant reduction of eggs laid and of their hatching rate in the pig manure treated ecologically by housefly larvae. PMID- 15283257 TI - [Western blotting analysis of specific antigens from different components of Echinococcus metacestodes]. AB - OBJECTIVE: To analyze antigens for searching specific antigenic components for immunodiagnosis of echinococcosis. METHODS: Fourteen crude antigens from different tissues (cyst fluid, protoscoleces, laminated layer and germinal layer) of Echinococcus granulosus and E. multilocularis metacestodes and other 4 species of cestodes were analyzed by Western blotting. The differences of protein bands were compared for the 14 crude antigens by reacting with pooled sera from cystic echinococcosis (CE) and alveolar echinococcosis (AE) patients. RESULTS: Eleven protein bands from the antigens reacted nonspecifically with sera from both CE and AE patients were Mr 130000, 100000, 94000, 80000, 75000, 66000, 62000, 52000, 38000, 32000, 24000. The highly specific protein bands recognized by AE sera were Mr 120000, 109000, 86000, 59000, 43000, 28000, 20000, 18000, and by CE sera were Mr 41000, 40000, 22000, 16000 and 12000. CONCLUSION: Different antigens shared by the two species of Echinococcus were examined and potential antigenic proteins specific for AE or CE sera were found, providing useful information for further identifying specific antigens for immunodiagnosis. PMID- 15283258 TI - [A case of myiasis in conjunctiva]. PMID- 15283259 TI - [Transmission-blocking vaccine candidate of Plasmodium vivax Pvs25 is highly conservative among Chinese isolates]. AB - OBJECTIVE: To investigate the genetic diversity of Plasmodium vivax transmission blocking vaccine candidate antigen (TBV) Pvs25, with P. vivax isolates from Hubei and Zhejiang Provinces, and to compare the genetic polymorphism of Pvs25 with that from Bangladesh. METHODS: The parasite DNA used for the genetic polymorphism assay was obtained from dried filter paper blood spots. The genes were PCR amplified and the products were purified and sequenced directly. RESULTS: 45 complete new sequences were analyzed. Only 3 nucleotide changes were found that would result in amino acid substitutions in Pvs25 in comparison with the sequence from P. vivax Sal-I strain. The measurement of nucleotide diversity (pi) was remarkably similar for the two populations, indicating that DNA sequences and deduced amino acid sequences were highly homologous among the geographically dispersed isolates or isolates from the same geographical region. CONCLUSION: The results suggest that Pvs25 has limited antigenic polymorphism, especially compared with candidate antigens expressed by hepatic and erythrocytic stage, which may support the development and application of Pvs25-based transmission blocking vaccine in China. PMID- 15283260 TI - [Prophylactic effect of artesunate against experimental infection of Schistosoma mansoni]. AB - OBJECTIVE: To study the prophylactic effect of artesunate against the infection of Schistosoma mansoni in mice and its optimal scheme for preventing schistosomiasis mansoni. METHODS: BALB/c mice were infected by tail dipping method with S. mansoni cercariae. Mice were administered orally with artesunate at different developmental stage of the parasite, with different regimens. The reduction rates of total and female worms, the number of eggs in the liver and intestine, and the fecundity were calculated and treated statistically. RESULTS: The optimal dosage of artesunate to prevent murine schistosomiasis was 300 mg/kg. The parasite was found to be especially susceptible to artesunate in its schistosomula stage of 14 and 21 d after infection, resulting in worm reduction rate of 84% and 93% respectively compared with control. High protection was reached with worm reduction rate of 99% by the regimens of 300 mg/kg once a week for 4 consecutive weeks beginning 14 d after infection. The fecundity was significantly suppressed, suggesting that the drug inhibited sexual maturation of female worms. The effective protection could also be gained with prolonged interval time of two weeks with worm reduction rate of 97% and 96% beginning 14 or 21 d after infection. CONCLUSION: Artesunate kills schistosomula and reduces the fecundity of females effectively, the infected mice do not develop schistosomiasis mansoni when treated with artesunate. It's proposed that an optimal scheme for field use be the first administration 14 or 21 days after infection with 1 or 2 weeks interval. PMID- 15283261 TI - [Canonical correlation and redundancy analysis on the indices of abdominal ultrasonography of schistosomiasis japonica]. AB - OBJECTIVE: To explore the correlation of the ultrasound indices of liver and spleen in schistosomiasis japonica and with infection frequency, infection time and EPG. METHODS: The canonical correlation was applied to analyze the correlation of the hepatosplenic ultrasound indices in schistosomiasis japonica, and the correlation of the ultrasound indices with infection frequency, infection time and EPG. The proportions of variation related to each other in liver and spleen were analyzed by the redundancy analysis. RESULTS: The correlation coefficients of the first pair correlation canonical variable were 0.7842, 0.5483 and 0.5800, 0.4220, respectively, in males and females without infection, males and females with infection history (P<0.01). The correlation coefficients of the first pair correlation canonical variable were 0.6063, 0.5215 and 0.6595, 0.3849, respectively, in male negatives and female negatives, male positives and female positives (P<0.01). In groups of males and females without infection, the variations of liver ultrasound indices related with the variation of spleen ultrasound indices were 43.5% and 17.5% respectively, and in groups infection history, they were 22.1% and 11.4% respectively. In male and female negative groups, the variations of liver ultrasound indices related with the variation of spleen ultrasound indices were 26.8% and 16.8% respectively, and in positive groups, they were 27.6% and 10.7% respectively. The infection frequency, infection time and EPG in the stool-positive groups were not significantly related with the canonical variables of liver and spleen ultrasound indices (P>0.05). CONCLUSION: There is a significant canonical correlation between the liver and spleen ultrasound indices in schistosomiasis japonica, especially in males. In groups with infection history, the infection frequency, infection time and EPG of those stool-positives are not relevant to the canonical variables of liver and spleen ultrasound indices. PMID- 15283262 TI - [Studies on the characteristic of interferon-gamma mediating resistance in mice infected with Schistosoma japonicum]. AB - OBJECTIVE: To investigate the molecular characteristic of interferon-gamma mediating protective immunity against schistosomiasis japonica in mice. METHODS: CD4+ T cells were isolated from spleens of mice infected with Schistosoma japonicum at different time-points. The cDNA microarray technique combined with RT-PCR was used to explore IFN-gamma inducible GTPase family gene expression profile of CD4+ T cell. IGTP, a representative IFN-gamma, inducible GTPase having vital anti-infection activity, was amplified from spleen of BALB/c mice using RT PCR, then cloned into pGEM(r)-T easy vector for sequencing. RESULTS: IFN-gamma inducible GTPase family had the similar characteristic over the course of S. japonicum infection. The gene expression of these members were up-regulated or had little change at 3 wk post-infection, then down-modulated from 6 wk to 13 wk post-infection, which was also confirmed by RT-PCR. As for IGTP, two inserts were identified after sequencing. One was 142 bp shorter than another, but the fragment was lost due to low annealing temperature. CONCLUSION: There is a dramatic inhibition of IFN-gamma pathway and IFN-gamma-dependent anti-infective immunity during the infection of S. japonicum. PMID- 15283263 TI - [Ultrastructural changes in the midgut epithelium of Ixodes sinensis after infesting the rabbits immunized by purified ixodic protein]. AB - OBJECTIVE: To observe the ultrastructural changes in the midgut epithelium of Ixodes sinensis after infesting rabbits immunized with Mr 105000 purified tick antigen. METHODS: New Zealand rabbits were inoculated with Mr 105000 purified antigen by means of mutiple intradermal injection in foot pad, groin and back. Each immunized rabbit was infested by 30 female Ixodes sinensis. At 24 hours, 48 hours, 72 hours, 5 days and 8 days after infestation, three Ixodes sinensis in each group were observed for ultrastructural changes in the epithelium of their midgut. RESULTS: Histological examinations showed that with the time going, digestive cells of the ticks after infesting hosts became more and larger with dense and regularly arranged microvilli, enriched organella, distinct unit membrane structure, and the appearance of tubli, small vacuole, numerous lipid droplets and hematin granules. These cells also developed a highly infolded basal lamina, forming a labyrinth system. The digestive cells of immunized group were however greatly damaged, whose number and volume were significantly different from control groups. From 24 to 48 hours after infestation, the midgut epithelium of Ixodes snenss showed pathological changes with the basal lamina becoming thinner, looser and broken; digestive cells damaged and vacuolated; microvilli decreased, shortened and irregularly arranged; the mitochondria swollen and its crests reduced, shortened and even with myeloid changes; the rough endoplasmic reticulum dilated; lipid droplets and hematin granules decreased; phagocytic and pinocytic activity weakened; and basal labyrinth system vacuolated. From 72 hours to 8 days after infestation, cells were severely damaged, organella were denatured and necrotic, nuclei showed pyknosis and cells lysed. CONCLUSION: The rabbits immunized with Mr 105000 purified ixodic protein have acquired the adoptive immunity against Ixodes sinensis; in the anti-tick immunity described above, the midgut of Ixodes sinensis is the major affected site. PMID- 15283264 TI - [Development of hybrid strains from geographic isolates of Eimeria tenella and their immunoprotection]. AB - OBJECTIVE: To cultivate hybrid strains from three geographic isolates of Eimeria tenella and to explore the possibility of developing vaccine candidates. METHODS: Three parental strains were selected from five geographic isolates of E. tenella through immune experiment, and hybrid strains were cultivated. The genomic DNA of the three parental strains and their filial generation were analyzed by random amplified polymorphic DNA (RAPD) technique with 30 optimization primers screened from 200 primers, the hybrid strains were isolated from the filial generation by RAPD. Chicken were inoculated with hybrid strains, and challenged with different strains to compare the immunogenicity and immunoprotection. RESULTS: Immunogenicity and immunoprotection of the three strains isolated from Guangzhou, Baoding and Changchun were stronger than those of other strains. Hybridization was performed to cultivate hybrid strain. Two hybrid strains were isolated from Changchun x Baoding and Guangzhou x F1 by RAPD. The result of immune experiment proved that immunoprotecion of F1 and F2 were higher than their parental strains. CONCLUSION: Two hybrid strains have been cultivated from the three geographic isolates of E. tenella, with the immunogenicity of their parental strains. Chicken immunized by F2 strain have shown strong resistance against the infection of the geographic strains, with an average protection rate of 84%. PMID- 15283265 TI - [A case of cerebral sparganosis mansoni]. PMID- 15283266 TI - [Intra-cellular signal pathway and synthesis of prostaglandin E2 during invasion of macrophage by Toxoplasma gondii]. AB - OBJECTIVE: To investigate the signal transduction pathway of arachidonic acid (AA) and prostaglandin E2 (PGE2) synthesis in macrophage invaded by Toxoplasma gondii. METHODS: Synthesis of AA and PGE2, expression of COX-2 mRNA and protein following stimulation infection by Toxoplasma gondii were evaluated in RAW264.7 cells by ELISA, RT-PCR and Western blotting after treatment with calcium channel blocker verapamil, chelator of extracellular calcium EGTA and inhibitor of CaM trifluoperazine (TFP), selective PKC inhibitor H7. RESULTS: Production of AA and PGE2 induced by tachyzoite was significantly inhibited by EGTA, TFP and BAPTA/AM, and the PGE2 production was inhibited by H7, with a reduced expression of COX-2 mRNA and protein in a dose-dependent manner. CONCLUSION: The parasite down regulates macrophage functions by affecting PKC signaling pathways, and triggers a biochemical cascade whose signals ultimately conduct to the secretion of immunosuppressive molecules PGE2. PMID- 15283267 TI - [Expression of adenylate kinase of Schistosoma japonicum and evaluation on the immunoreactivity of the recombinant protein]. AB - OBJECTIVE: To express and purify the Schistosoma japonicum adenylate kinase (AK) protein of which the cDNA sequence was subcloned into the pET32a(+) soluble expression plasmid, and to evaluate its immunoreactivity. METHODS: The recombinant plasmid was transformed into E. coli BL21, and induced with IPTG for expression. The bacterial lysis was conducted by ultrasonication and the supernatant was analyzed by SDS-PAGE after boiling and centrifugation. The target protein was purified with the Ni-NTA agarose. The proteins on the gel were transferred to the PVDF film and then the immunoreactivity was tested by Western blotting using anti-6-His antibody, sera from rabbits 6 weeks after infected with cercariae or UV-attenuated cercariae. The purified protein was coated for ELISA to test the cercariae infected rabbit serum and the normal rabbit serum. RESULTS: The molecular weight of the target fusion protein was Mr 40 000 after being induced with IPTG. The fused protein showed a single band when reacted with anti 6-His antibody, the cercariae infected rabbit serum and attenuated cercariae infected rabbit serum. CONCLUSION: The AK protein is expressed as a fused protein with thioredoxin and its molecular weight is about Mr 40000. This protein has a positive immunoreactivity with sera of rabbits infected with cercariae and UV attenuated cercariae. PMID- 15283269 TI - [Pseudo-parasitism of Limax flavus in human body]. PMID- 15283268 TI - [Relationship between the Demodex and bacteria infection in human rosacea]. AB - OBJECTIVE: To understand the relationship between Demodex and bacteria infection in rosacea (brandy nose), and to find effective means for the treatment. METHODS: Cellophane tape was used to detect Demodex on the nasolabial grooves and the face; sebum and tissue on face was scraped and cultured to examine bacteria under microscope. The hospital-made anti-rosacea lotion was used on the affected part two times a day for 7 days. RESULTS: It was found that 193 (74.2%) of 260 cases with rosacea were infected by Demodex and 209 (80.4%) of the patients were infected by bacteria. The overall effective rate of the treatment for rosacea was 73.5%. CONCLUSION: Bacteria infection in rosacea is an important factor inducing rosacea. The curative effect of the anti-rosacea lotion is good. PMID- 15283270 TI - [Studies on the relationship between the level of cytokine and liver function in patients with clonorchiasis sinensis]. AB - OBJECTIVE: To explore the relationship between cytokine level and liver function among patients infected with Clonorchis sinensis. METHODS: 47 patients were divided into three groups according to the degree of Child-Pugh liver function grade: 20 in group A (3-4 scores), 15 in group B (5-6 scores) and 12 in group C (7-9 scores). Interleukin 2 (IL-2), soluble IL-2 receptor (sIL-2R), interleukin 8 (IL-8) and tumor necrosis factor (TNF-alpha) were examined by radioimmunoassay (RIA) and enzyme-linked immunosorbent assay (ELISA). Automatic biochemical analyzer was employed for the determination of serum level of total bilirubin (TBL), albumin (ALB) and alanine aminotransferase (ALT). Data were analyzed with SAS statistic software. RESULTS: Serum levels of sIL-2R, IL-8 and TNF-alpha from patients were significantly higher than those obtained from healthy people (P<0.05, P<0.05, P<0.01), whereas the IL-2 level was significantly lower than the former (P<0.01). With the affected degree of the liver, serum levels of sIL-2R, IL-8 and TNF-alpha increased, in contrast to the decrease of IL-2 level. The differences were significant between groups A and C (P<0.05). The level of sIL-2R and TNF-alpha directly correlated with that of TBL (r=0.331 P<0.05, r=0.518 P<0.01) and ALT (r=0.475 P<0.01, r=0.285 P<0.05) respectively, but inversely correlated with the level of ALB (r=-0.319 P<0.05, r=-0.665 P<0.01). CONCLUSION: The infection of Clonorchis sinensis results in the reduction of cellular immune function of the patients. Certain relationship exists between serum cytokine level and liver function. Two cytokines, sIL-2R and TNF-alpha, are involved in the process of pathology. PMID- 15283271 TI - [Advances in research of molecules related to the immune evasion of schistosomes]. PMID- 15283272 TI - [Imported falciparum malaria and misdiagnosis in Beijing: retrospective analysis of fifteen cases]. PMID- 15283273 TI - [Surveillance of schistosomiasis after transmission interruption in Jiaxing City]. PMID- 15283274 TI - [Sero-epidemiological investigation of schistosomiasis in human population of lake and swamp region]. PMID- 15283275 TI - [Diseases of the throat and phonosurgery]. PMID- 15283276 TI - [Oropharyngeal measure in patients with obstructive sleep apnea-hypopnea syndrome before and after uvulopalatopharyngoplasty]. AB - OBJECTIVE: To explore the relationship between the extent of enlarged oropharynx and efficiency through measuring the anterior-posterior and transverse diameter of oropharynx of patients with obstructive sleep apnea-hypopnea syndrome (OSAHS) before and after uvulopalatopharyngoplasty (UPPP). METHODS: Thirty eight patients with OSAHS were studied. The following indexes were measured before and after UPPP: width of uvula base, length of uvula, distance between uvula and posterior pharyngeal wall (DBUP), distance between anterior pillar (DBPP), apnea-hypopnea index (AHI), body mass index (BMI) and SaO2. RESULTS: The preoperative DBAP and DBPP were significantly less than those of normal adults (P < 0.05). DBUP, length of uvula and width of uvula base has no significant difference between preoperative patients and normal adults (P > 0.05). There was no significance difference in DBAP. DBPP and DBUP between postoperative patients and normal adults (P > 0.05). The preoperative AHI, IBM, minimal SaO2, mean SaO2, DBUP, DBPP, DBAP, length of uvula and width of uvula base has no significant difference between good responders and nonresponders (P > 0.1). CONCLUSIONS: Transverse diameter of OSAHS patients is shorter than that of normal adults but anterior posterior diameter of OSAHS patients has no difference compared with normal adults. Transverse diameter could be enlarged by UPPP. Not only anatomical abnormality but also other factors will contribute to the effect of UPPP. PMID- 15283277 TI - [Surgery for velopharyngeal insufficiency: a review of 219 modified pharyngoplasties]. AB - OBJECTIVE: Two-hundreds and nineteen velopharyngeal insufficiency cases treated with a modified pharyngoplasty were retrospectively analyzed, the effectiveness of the surgery was studied. METHODS: Two-hundreds and nineteen patients (111 males, 108 females, range from 4 to 33 years, mean age 14.2 years) with velopharyngeal insufficiency were studied. Among 219 patients, 44 with congenital velopharyngeal insufficiency, 21 with fistula after palatoplasty and 154 with velopharyngeal insufficiency after primary palatoplasty. All cases had special examination, Chinese speech intelligibility test and blowing test before and after modified pharyngoplasty. RESULTS: Hypernasality in all cases were reduced and blowing test increased with various degrees. The Chinese speech intelligibility rose from 41% to 63%. CONCLUSIONS: The modify pharyngoplasty is a safe and effective method for velopharyngeal insufficiency of patient. PMID- 15283278 TI - [Long-term follow-up of autogenous fat injection for unilateral vocal cord paralysis]. AB - OBJECTIVE: To study the long-term results of autogenous fat injection for unilateral vocal cord paralysis. METHODS: Twenty cases with unilateral vocal cord paralysis were treated by autogenous fat injection into the thyroarytenoid muscle to achieve medialization. The patients were divided into 3 groups by hoarse degree before operation, all of them were followed more than 12 months with serial video laryngoscope and voice evaluation. The ratio between paralyzed vocal cord upper surface and that of the normal vocal cord were adopted as the measurement for the vocal cord volume changes before and after operation. RESULTS: 1. The volume of paralyzed vocal cord was increased. The degree of hoarse and normalized noise energy (NNE) were evaluated by objective methods after operation. 2. The hoarse symptom was less severe after operation than that before operation. The cure cases 3 to 6 months and over 12 months after operation were nearly the same. 3. NNE of over 12 months and 3 to 6 months after operation were not significantly different, but the postoperative NNE were different with that before operation. CONCLUSIONS: Autologous fat injection was an effective method for treating unilateral vocal cord paralysis, and the long term effects were reliable. PMID- 15283279 TI - [Anteroposterior cricoid split interposition grafting for severe glottic and subglottic stenosis]. AB - OBJECTIVE: To study the effect of anterior and posterior cricoid splitting interposition grafting for severe glottic and subglottic stenosis. METHODS: This is a retrospective study, from 1991 to 2001 years, 25 patients (male 15, female 10, aged 9 to 46 years) with severe glottic and subglottic stenosis were operated with anterior and posterior cricoid splitting interposition grafting method at Tangdu Hospital. All of 25 patients were tracheostomy dependent before reconstruction. 19 patients had previously undergone 1 to 7 (average 2) surgical procedures. The surgical technique consisted of laryngotracheostomy, cricoid lamina midline vertical incision; rib cartilage graft (17 cases), muscular fasciae, perichondrium or split-thickness skin graft (15 cases), pedicle arytenoid cartilage graft (2 cases) and thyroid cartilage graft (1 case) interposition and silicon T-tube stenting for 3 to 6 months. RESULTS: Twenty-four patients (96%) were successfully decannulated and got an effective phonation. One patient failed decannulation. The follow-up period ranged from 1 to 10 years. All of the 24 patients had a stable airway and effective phonation. CONCLUSIONS: The anteroposterior cricoid split interposition graft technique was a safe and effective method for the treatment of severe glottic and subglottic stenosis. Careful split of the cricoid, avoiding injury of esophageal musculature, careful hemostasis, a tight suture graft and using stent were the keys of successful operation. PMID- 15283280 TI - [Expression of transforming growth factor-beta1 and c-fos and p27 in human middle ear cholesteatoma]. AB - OBJECTIVES: To analyze the expression of transforming growth factor-beta1 (TGF beta1), cyclin dependent kinase inhibitor (p27) and c-fos in human middle ear cholesteatomas and to investigate the correlation between their expression and the ability of erosion of cholesteatoma. METHODS: Immunohistochemical staining (SP method) of 31 cholesteatomas and 11 external ear canal skin samples from patients and 10 external ear canal skin samples from healthful men which were taken intraoperatively, was performed for c-fos, TGF-beta1 and p27 positivity. The signals representing the expression of c-fos, TGF-beta1 and p27 were observed under microscope and scanned into a computer by an image scanner. The gray-scale of positive signals were quantitated by image processing computer. RESULTS: The percentage of positive expression of TGF-beta1 and c-fos in cholesteatoma were 87.1% and 83.9%, respectively. Their expression tended to be increased greatly compared with which in the skins of the control groups. Positive p27 signals were not observed in cholesteatomas and external ear skin tissues. It showed statistically significant correlation between expression of c-fos and the ability of erosion of cholesteatoma. There was correlation between the express ion of TGF beta1 and the ability of erosion of cholesteatoma too. But there was no correlation between the expression of c-fos and TGF-beta1. CONCLUSION: The expression of c-fos in cholesteatoma was significally higher compared with which in the skin of external auditory of cholesteatoma patients and healthful men, which indicate that c-fos plays an important role in the hyperproliferative of cholesteatoma. The expression of TGF-beta1 was significant higher in cholesteatoma, which indicate that cytokines such as TGF-beta1 play a great role in the etiology of cholesteatoma. PMID- 15283281 TI - [Effect of ventilation tube insertion on otitis media with effusion in cleft palate children]. AB - OBJECTIVE: To explore the effect of eardrum tubing in the repair of cleft palate on alleviating the otitis media with effusion (OME) and hearing loss in cleft palate patients. METHODS: Nineteen ears of 19 cleft palate children with OME and hearing loss were treated with the ventilation tube insertion in the repair of the cleft palates, while the untreated opposite ears were selected as the control group. All patients were followed up from 2 weeks to 18 months postoperatively and their middle ear condition and hearing thresholds were reevaluated by otoscopy and pure-tone audiometry. RESULTS: Significant differences were found in the incidences of hearing loss between pre and postoperative patients in treated ears, and there are no differences in the untreated ears. Postoperative hearing thresholds become lower than that before the operation and no serious complications were found in the treated ears. CONCLUSIONS: The ventilation tube insertion in medial ear is safe and effective to restore the hearing impaired by OME in the cleft palate patients. It can be used as a regular management for OME and hearing loss in cleft palate children. PMID- 15283282 TI - [Treating defective damage of facial nerve with great auricular nerve grafting covered by pediculated fascial tube]. AB - OBJECTIVE: To study the curative effect of grafting great auricular nerve with pediculated fascial tube in defective damage of facial nerve. METHODS: All the 7 patients in this study were treated by grafting great auricular nerve covered by pediculated fascial tube near facial nerve trunk. RESULTS: Four cases with otogenic facial paralysis had grade III - IV recovery of facial nerve function from two to two and half years after the nerve grafting operation. Three patients post-traumatic facial paralysis had grade III recovery of facial nerve function 2 years after the nerve grafting operation. CONCLUSIONS: The grafting of pediculated fascial tube surrounded great auricular nerve can provide a biological environment with better blood supply for the plerosis and regeneration of nerve and can accelerate the functional recovery of nerves after the nerve grafting. PMID- 15283283 TI - [Immune responses against cancers in cervical lymph nodes of laryngeal carcinoma]. AB - OBJECTIVE: Densities of dendritic cells (DC) and hyperplastic follicular response in cervical lymph nodes were performed to prove their roles in immune responses against cancers. METHODS: Paraffin blocks were prepared for staining with monoclonal antibodies against CD45RO, CD20 and S-100 proteins,in 157 lymph nodes obtained from elective cervical lymphadenectomy in 47 cases of laryngeal squamous cell carcinomas. RESULTS: Quantitative analysis showed that the patients who survived longer than 5 years had significant higher number of follicles and higher extent of infiltration by DCs in the lymph nodes than those who less than 5 years (P < 0.001). According to negative or positive lymph node metastasis, there were statistically significant differences between two groups (P < 0.001). The patients who possessed T cell increase type follicular reaction had significant higher five-year survival rate ( P < 0.01) and lower lymph node metastasis rate (P < 0.05) than those who possessed T cell decrease type reaction. CONCLUSION: DCs and hyperplastic follicular response may be more directly involved in the host immune reaction against tumor. The classification of follicular reaction, the densities of DCs and follicular reaction, can serve as important indicators in assessing prognosis of laryngeal carcinomas. PMID- 15283284 TI - [Quality of life, personality, coping style in the patients with advanced laryngeal cancer]. AB - OBJECTIVE: To evaluate the relationship between the life quality of advanced laryngeal squamous cell cancer patients and their personality, coping style and other psychological factors. METHODS: The life quality were measured for 2 sub groups of advanced laryngeal cancer patients and the normal control group. The Eysenck personality questionnaire (EPQ), coping style questionnaires (CSQ) and University of Washington-quality of life (UW-QOL) were used for life quality evaluation. The 2 sub-groups of the patients on the worse speech, job and ability (group I), the better speech, job and ability (group II), and normal control. RESULTS: (1) UW-QOL score: the total, activity, recreation, job and speech scores of group II were significantly higher than those of group I (P < 0.01). Group II was better than group I in appearance (P < 0.05). (2) EPQ score: the P and N scores in group II were lower than that in group I (P < 0.05). The E scores in group II were significantly higher than that in group I (P < 0.01). (3) CSQ score: the problem-saving factor and help-seeking factor in group II were more significantly lower than that in group I (P < 0.01). The self-blaming factor in group II were higher than that in group I (P < 0.05). (4) There was positive correlation between total scores of QOL and the problem-saving factor, help seeking factor of CSQ, the E scores of EPQ (P < 0.05), there was negative correlation between total scores of QOL and the P scores, the N scores of EPQ (P < 0.05). CONCLUSIONS: The advanced laryngeal cancer patients of group II have better life quality, and their personalities showed more extroversive, stable feeling, adaptable, mature coping styles. The above characteristics may have good effects on the prognosis of advanced laryngeal cancer. PMID- 15283285 TI - [Clinical analysis of 71 cases of multiple primary cancers in head and neck squamous carcinomas]. AB - OBJECTIVE: To study the location, treatment, life status of multiple primary cancers (MPCs) in head and neck squamous cell carcinomas. METHODS: The clinical data of 71 head and neck squamous carcinoma patients with MPCs were retrospectively analyzed. RESULTS: MPCs were seen in head and neck regions in 27 cases and in remote organs in 42 cases, two of which were triplicate primary cancers. Four cases were synchronous MPCs, including one patient with synchronous triplicate primary cancer. Other 67 cases were heterochronous MPCs, including one patient with heterochronous triplicate primary cancer. Of 67 heterochronous MPCs, the time interval between index tumor presentation and diagnosis of MPCs was eight months to twelve years. MPCs occurred in seventy percent index oral cavity squamous cancers, which were located in head and neck regions, and in sixty-two percent index hypopharynx cancers and seventy-nine percent index laryngeal cancers, which were located in remote organs. The incidence of MPCs in esophagus and lung was higher than that in other remote organs. Among the various MPCs in this serials, the incidence of the disease appeared to be the highest in esophagus, accounting for twenty-four percent of all cases. The total three- and five-year survival rates were 32.4% and 22.5%, respectively. Of all MPCs patients, the three-year survival rate for patients who received different therapies for their MPCs was obviously higher than that of untreated patients (P < 0.01, Chi-square test). CONCLUSIONS: Esophageal carcinoma is the most common second primary cancer among the various MPCs of the head and neck squamous carcinomas. Oral cavity cancers tend to have more MPCs in the head and neck regions, and laryngeal and hypopharyngeal cancers are easily to be associated with MPCs in the remote organs. Regular follow-up and early diagnosis with effective treatment can help to improve the survival of MPC patients in head and neck squamous cell carcinomas. PMID- 15283286 TI - [Surgical treatment of advanced thyroid carcinoma with aero-digestive invasion]. AB - OBJECTIVE: To study the surgical management of the advanced thyroid carcinoma with aero-digestive invasion. METHODS: To analyze 18 cases of advanced thyroid carcinoma which had aerodigestive invasion retrospectively, the patients were treated from 1988 to 1998. RESULTS: The rate of aerodigestive invasion occur was 3.5% (18/516), The inner cavity invaded rate was 2.7% (14/516). The most invaded organ was trachea; The others were esophagus and larynx, pharynx; Two and/or more organs invaded rates were 44. 4% (8/18). The cases that survived 1, 3, 5 and 10 year were 17, 14, 9 and 6. The 5 years survival rates of the differentiated thyroid cancer (61.5%) were more higher than the undifferentiated thyroid cancer (20.0%), but there were no statistical difference between them (P > 0.05) and the same in clinical stage (P > 0.05). CONCLUSIONS: Intraluminal extension is a more serious problem in which usually extensive resection of the aerodigestive tract is required. Effective surgical treatment combined with postoperative auxiliary management for this kind of patients may achieve a good long term results. PMID- 15283287 TI - [Evaluation of the clinical studies of prevention and treatment of sudden deafness in China]. AB - OBJECTIVE: To summarize and evaluate the current situation on sudden deafness in China by retrospective review of the articles appeared in Chinese journals in the past 3 years. METHODS: The terms of sudden deafness and therapy were used to search all of the articles on the therapy of sudden deafness through the Chinese bio-medicine literature data base and Chinese journal net from 2000 - 2002. All of the articles were analyzed according to the standards of evidence-based medicine. RESULTS: One hundred and seventy-six articles were collected. 126 were on the curative effect of medicine, of which 26 (20.6%) based on the diagnosis and therapy standards settled by the otolaryngology branch of Chinese medical association in 1996, 89 (70.6%) were controlled clinical trials and 36 (28.5%) were randomized controlled trials. Most of the articles mentioned the 5 items diagnosis based and not detailed weather hypertension, diabetes and hyperlipemia were excluded. CONCLUSIONS: There are many articles on the therapy of sudden deafness. However, the inconsistent understanding of the prognosis and evaluation and unstuck trial design limited the therapy, research and the standards of literature on sudden deafness to progress further. PMID- 15283288 TI - [The current development of rhinology in China: still a lot of work to be done]. PMID- 15283289 TI - [Clinical analysis of 58 cases intraoperative image-guidance in nasal endoscopic surgery]. AB - OBJECTIVE: To evaluate the availability and our experience of intraoperative image-guidance in endoscopic nasal surgery. METHODS: Fifty-eight cases of endoscopic nasal surgery with intraoperative image-guidance were retrospectively reviewed, including 39 cases of chronic sinusitis with or without nasal polyp; 3 cases of necrotizing maxillary sinusitis; 6 cases of sphenoid sinus cyst; 2 cases of nasopharyngeal angiofibroma; 1 case of cranio-nasal meningioma; 1 case of traumatic cerebrospinal rhinorrhea; 1 case of traumatic optic nerve lesion. All patients were operated on with Brain Lab operation imaging navigation system and nasal endoscope. RESULTS: The preoperative preparatory time would take 10-20 minutes, including coordination, head holder localization, conventional instrument registration. In our cases, the localization accuracy between 3-D image landmarks of navigation system and actual anatomical landmarks was less than 1.5 mm. The optic nerve and other anatomical points could be orientated accurately in intraoperative procedures. No complication occurred. CONCLUSIONS: Nasal endoscope combined with image-guidance systems provides accurate anatomical localization of nasal cavity, sinuses and anterior skull base with enlarged operation field. It is possible for surgeons to observe the surrounding important anatomical structures during endoscopic nasal surgery. It could increase the effectiveness and decrease surgical complications, especially in complicated cases. PMID- 15283290 TI - [Imaging-navigated endoscopic sinus surgery]. AB - OBJECTIVE: To evaluate the advantages and disadvantages of different type of image-guidance system in endoscopic sinus surgery. METHODS: Fifty-three endoscopic sinus surgery were performed under different type of image-guidance system, there were 24 chronic sinusitis with or without nasal polyp, 4 juvenile nasopharyngeal angiofibroma, 8 pituitary adenoma, 9 ethmoid ossifying fibroma, 2 nasopharyngeal mixed tumor, 1 nasal leiomyoma, 3 fungal sinusitis, and 2 inverting papilloma. RESULTS: In all cases, the preoperative time was 15-30 minutes, the registration rate were 1.3-2.0, the localization accuracy was within 1 mm. Compared with the traditional endoscopic sinus surgery, the operating time was similar, without obvious difference. No complication occurred. CONCLUSION: All types of image-guidance system could work well with endoscopic system, each of them had its own shortages. Every type of image-guidance system could identify the borders and critical anatomical structures in the corresponding CT data, especially in cases in which anatomical landmarks were no longer present, with anatomical variation, intranasal and anterior skull base tumor. Combined with endoscopic surgery, the image-guided endoscopic surgery provided accurate tumor resection while preserving normal tissue, increased surgical effectiveness, decreased overall surgical complications. It is believed that the image-guidance system is a useful tool for endoscopic sinus surgery. PMID- 15283291 TI - [Effect of endoscopic sinus surgery on asthmatic patients with chronic sinusitis and nasal polyps]. AB - OBJECTIVES: To explore the effect of endoscopic sinus surgery (ESS) in patients with chronic rhinosinusitis and asthma. METHODS: Forty-two patients with asthma who underwent ESS and control group were investigated. Interleukin-4 (IL-4), interferon-gamma (IFN-gamma), soluble interleukin-2 receptor (sIL-2R) and soluble CD23 (sCD23) levels in culture supernatant of peripheral blood mononuclear cell (PBMC) were determined by enzyme-linked immunosorbent assay (ELISA). RESULTS: Of these 42 patients 32 (76%), their asthma was relieved greatly after ESS comparing with that before ESS. 14 of 21 patients (67%) taken regular medication for asthma before ESS reported that less medicine was used after operation. CONCLUSION: This study demonstrates that ESS has a favorable effect on asthma in patients with symptomatic chronic sinusitis. PMID- 15283292 TI - [Expression and distribution of Fas and Fas-L in the nasal polyps]. AB - OBJECTIVES: To confirm the expression and distribution of Fas and Fas-L in the nasal polyps and to illustrate the role of the Fas/Fas-L system in the pathogenesis of human nasal polyps. METHODS: Investigating the transcripts of the Fas/Fas-L gene in 30 human nasal polyp tissues and 30 nasal turbinate mucosa specimens using reverse transcription-polymerase chain reaction. Localization of Fas/Fas-L was performed with immunohistochemistry. RESULTS: The transcripts of the Fas/Fas-L gene were detected at similar levels in both polyps and nasal mucosa. There was a significant overexpression of Fas-L protein on nasal polyps (epithelium: 25 +/- 21, glands: 19 +/- 14) compared to nasal mucosa (epithelium: 14 +/- 13, glands: 12 +/- 10), (t = 1.66, P < 0.01), while Fas was overexpressed on nasal mucosa (epithelium: 17 +/- 11, glands: 17 +/- 13) compared to nasal polyps (epithelium: 13 +/- 10, glands: 11 +/- 9), (t = 1.98, P < 0.01). Fas-L positive cells were localized on the epithelial layers of cystically dilated glands and the down-growing epithelium of nasal polyps. Fas positive cells were localized on the cilia of the epithelial of nasal mucosa and mainly on the infiltrative cells. CONCLUSION: Fas/Fas-L may play an important role on the pathogenesis of human nasal polyps, including cystic degeneration of submucosal glands, apoptosis and conferring of immune privilege to nasal polyp formation. PMID- 15283293 TI - [An improved animal model of autoimmune inner ear disease]. AB - OBJECTIVE: To establish an experimental autoimmune inner ear disease model, which could exhibit high reproducibility and be adopted for detailed immunological analysis. METHODS: Extraction of guinea pig inner ear antigens (IEAg). The inbred mice were given a single subcutaneous injection of diluted solution of pertussis and an emulsion containing equal parts of CFA and IEAg. The ABR threshold shifts were evaluated. The antibody level to IEAg in serum was detected by ELISA. Inner ear specimen were examined by light microscopy with hematoxylin and eosin staining. The infiltrated cells within cochlea were clarified with immunohistochemical techniques. RESULTS: The ABR thresholds of IEAg-sensitized animals were elevated significantly. Histological changes in cochlea were significant. Inflammatory cell infiltration was clearly observed in the cochlea of the animals following sensitization with IEAg. Degeneration of the spiral ganglion cells, which characterizes a decrease in cell numbers, and formation of endolymphatic hydrops were often seen too. Serum anti-IEAg levels after inoculation were significantly increased in the IEAg sensitised groups. Most of the infiltrated lymphocytes in scala tympani were CD4+ T cells. CONCLUSIONS: The experimental autoimmune inner ear disease can be induced by a single inoculation of IEAg-CFA emulsion and pertussis in inbred C57BL/6 mice. PMID- 15283294 TI - [Promoter hypermethylation of DNA repair gene O6-methylguanine DNA methyltransferase in laryngeal squamous cell carcinoma]. AB - OBJECTIVE: O6-methylguanine DNA methyltransferase (MGMT) is a DNA repair protein. It plays a important role in the repair of DNA damage caused by the exposure to methylating agents. Hypermethylation of CpG islands in the promoter regions result in MGMT genes loss of transcription. This study explored the hypermethylation of CpG islands in the promoter regions of MGMT genes in laryngeal squamous cell carcinoma. METHODS: Methylation-specific PCR and semiquantify RT-PCR were used to study the promoter methylation of the MGMT gene and loss of transcription in laryngeal carcinoma tissues, tissues adjacent to the tumor and normal laryngeal tissues. RESULTS: Among the 46 laryngeal carcinoma, MGMT gene hypermethylation was present in 16 cases (34.8%). MGMT hypermethylation was not detected in all tissues adjacent to the tumors and 5 normal tissues. There is no statistical significant difference of promoter hypermethylation among the samples of different histological grade (chi2 = 3.130, P = 0.077) or samples from patients with different TNM stages (chi2 = 3.957, P = 0.138). No expression of MGMT mRNA was found in all hypermethylated laryngeal carcinoma tissues. Expression of MGMT mRNA was detected in all unmethylated laryngeal carcinoma tissues, tissues adjacent to the tumors and 5 normal tissues. Expression of MGMT mRNA in unmethylated laryngeal carcinoma tissues is higher than tissues adjacent to the tumors (t = 30.540, P < 0.01) and normal tissues (t = 31.882, P < 0.01). There is no statistical significant difference of MGMT mRNA expression between tissues adjacent to the tumors and normal tissues (t = 0.419, P = 0.677). CONCLUSION: MGMT gene promoter hypermethylation is associated with loss of its transcription. PMID- 15283295 TI - [The expression and significance of bcl-2 and bax in each phase of the cell cycle in laryngeal carcinoma]. AB - OBJECTIVE: To detect the expression of bcl-2 and bax in each phase of cell cycle in laryngeal carcinoma, and explore the relationship between the expression of bcl-2 and bax in each phase of the cell cycle and the occurrence, development and prognosis in the carcinoma of larynx. METHODS: The immunohistochemical method, TUNEL technique and flowcytometry (FCM) parameter analyses were combined to detect the apoptosis and the expression of bcl-2 and bax in each phase of the cell cycle in 15 polyps of vocal cord and 387 laryngeal carcinomas. RESULTS: Total bcl-2 expression and bcl-2 expression in G0G1 stage in laryngeal carcinoma was significantly higher than that in polyp of vocal cord. In contrast, the total bax expression and the bax expression in each phase of cell cycle in laryngeal carcinoma were all lower than that in polyp of vocal cord. The total apoptosis index in laryngeal carcinoma was obviously lower than that in polyp of vocal cord, and this phenomenon was mainly caused by the decrease of the apoptosis in G0G1 phase. The bcl-2, bax expression and the apoptosis wasn't notably related to clinical stage, clinical type and T grade. In poor-differentiated squamous carcinoma, the bcl-2 expression in S and G2M phase was obviously higher than that in well-differentiated and the moderate-differentiated squamous carcinoma. The total apoptosis index, the apoptosis in S phase and the apoptosis in G2M phase were obviously enhanced both in the group of recurrence and in the patients who died in 5 years after the operation, in the same samples, the significant increasing of bcl-2 expression in S and G2M phase was detected. CONCLUSIONS: The significant decreasing of the apoptosis in G0G1 phase caused by high expression of bcl-2 was an important affair in the initial stage of laryngeal carcinoma. Accompanying the significant increasing of the total apoptosis index, the apoptosis in S phase and the apoptosis in G2M phase could be regard as an indicator that the cancer of larynx was malignant with poor prognosis and need adjuvant therapy. The decreasing of bax expression may play a role in the occurrence of laryngeal carcinoma. PMID- 15283296 TI - [The clinical value of quantitative analysis of plasma Epstein-Barr virus DNA in patients with nasopharyngeal carcinoma]. AB - OBJECTIVE: To elucidate the clinical value of quantitative analysis of plasma cell-free EBV-DNA in patients with nasopharyngeal carcinoma (NPC) . METHODS: Using fluorescence quantitative polymerase chain reaction (FQ-PCR), the copies of EBV-DNA in 66 plasma samples of pre- and post-treated NPC patients were examined, and compared with 30 normal controls. The relationships between the copies of free EBV-DNA and tumor stage, metastases load, as well as short-term therapeutic response were analyzed. RESULTS: The positive percentage and copies of plasma EBV DNA of NPC patients (66.7%, Median 522.0) were significantly higher than those of normal controls (10.0%, Median 0). Among the pre-treated patients, plasma EBV-DNA copies in stage III-IV patients (Median 3130.0) were higher than those were in stage I-II (Median 400.0), and the copies of N2-3 patients (Median 3250.0) were higher than those were in N0-1 patients (Median 400.0). And then the level of post-treated patients (Median 0) was lower than that of pre-treated patients (Median 522.0). Depending on the evaluation of therapeutic response, post-treated patients could be classified as two different categories, patients whose tumor regressed satisfyingly and those with remaining lesions. It was found that EBV DNA copies of the former (Median 0) were lower than those of the latter (Median 2083.0). CONCLUSIONS: FQ-PCR determination of plasma EBV-DNA may become a promising marker to monitor the course of this disease. PMID- 15283297 TI - [The surgical treatment and reconstructive methods in management of hypopharyngeal cancer]. AB - OBJECTIVE: To study the surgical treatment and the long-term effect of different reconstructive methods in surgical treatment of hypopharyngeal cancer. METHODS: A retrospective review of 303 (265 males, 38 females, ages ranging from 32 to 77 years) cases with hypopharyngeal cancer that were treated with surgical treatment from 1965 to 1998 were accomplished. Of the 303 cases, 130 cases were treated with different reconstructive methods and 173 cases without reconstruction. Of the 130 (stage II, 5; III, 16; IV, 109) cases, 94 were originated from pyriform sinus, 18 from posterior pharyngeal wall and 18 from postcricoid. Fifteen patients were reconstructed with free jejunum. Eighty-one patients were performed total pharyngo-larygo-oesophagectomy and gastric pull-up. Ten patients were treated with vascularized colon. Twenty patients were reconstructed with pectorals major myocutaneous flap. Four patients were reconstructed with other methods. Of the 173 (stage I, 7; II, 12; III, 51; IV, 103) cases, 160 were originated from pyriform sinus, 8 from posterior pharyngeal wall and 5 from postcricoid. RESULTS: The overall 3 and 5 years survival rate of 130 patients with reconstruction were 43.2%, 36.4% respectively. The overall 5 years survival rate of 173 patients with no reconstruction was 47.7%. The overall rate of regular swallowing was over 80%. The mortality rate in period of surgery of gastric pull-up and pectorals major myocutaneous flap were 8.6% and 15% respectively. No patients with free jejunum and vascularized colon died. The overall complication rate in nineties was lower than before 1990 (chi2 = 13.457, P = 0.004). The highest complication rate occurred in patients with pectorals major myocutaneous flap. In contrast to other reconstruction methods, the rate of success of swallowing was higher in patients with free jejunum. CONCLUSIONS: In selected patients these reconstruction techniques enable functional rehabilitation of swallowing. Even high survival rate was obtained after extended partial pharyngolaryngectomy. PMID- 15283298 TI - [Analysis of risk factors in the prediction of distant metastases of head and neck squamous cell carcinomas]. AB - OBJECTIVE: To investigate the risk factors related with distant metastases (DM) from head and neck squamous cell carcinomas (HNSCC). METHODS: A retrospective study was carried out to review the histopathological data from 532 HNSCC patients treated in Bethune International Peace Hospital from February 1978 to February 1998. The incidence and the risk factor for DM were evaluated in a model that included the following factors: sex, age, clinical staging, T and N staging, site of primary tumor, depth of primary tumor infiltration, histological grade of primary tumor, presence of cervical lymph node metastasis, number of positive neck nodes and levels involved, and presence of extracapsular nodal spread. Univariate chi2 test and multivariate stepwise logistic regression model were used for the analysis. Statistical analysis of overall survival was performed using Kaplan-Meier method. RESULTS: Sixty cases (11.3%) presented distant metastases in 532 patients of head and neck squamous cell carcinomas. In a univariate analysis, it was confirmed that the following variables correlated to DM, i.e., clinical staging (P = 0.0126), T classification (P = 0.0082), site of primary tumor (P = 0.0011), depth of primary tumor infiltration (P = 0.0005) , presence of cervical metastasis (P = 0.0057), number of positive neck nodes (P = 0.0149) and levels involved (P = 0.0034), presence of extracapsular nodal spread (P = 0.0118). In a multivariate analysis, the most significant risk factors for DM were the site of primary tumor and the depth of primary tumor infiltration. Kaplan-Meier analysis showed that overall survival rates of 60 HNSCC patients who presented distant metastases were 51.7% at 1 year, 13.3% at 3 years, 6.5% at 5 years, respectively. CONCLUSION: The site of primary tumor and the depth of primary tumor infiltration are the key risk factors in determining the development of DM in HNSCC patients. Patients with laryngeal and hypopharyngeal carcinomas and patients with primary tumor infiltrating muscular, bone or cartilage level have the highest risk of developing DM. PMID- 15283299 TI - The effect of cysteine analogues on the excretion of urinary sulphate in the rat following cysteine administration. AB - A major pathway for the production of sulphate within the mammalian body is known to be via the oxidative degradation of the sulphur moiety within the amino acid, L-cysteine. The ability of two structurally similar sulphur-containing drugs, the anti-rheumatic agent, D-penicillamine, and the mucoactive compound, S carboxymethyl-L-cysteine, to interfere with this sulphate production was investigated. Co-administration to the male rat of D-penicillamine (p.o.) and S carboxymethyl-L-cysteine (p.o.) with [35S]-L-cysteine (i.p.) led to a significant decrease in the subsequent urinary elimination of inorganic sulphate whilst having no measurable effect on organic sulphate excretion. The co-administration of L-valine, an amino acid not containing sulphur, had no effect. It is not known where, within the complex sequence of events surrounding the degradation of cysteine to sulphate, that D-penicillamine or S-carboxymethyl-L-cysteine may interact. PMID- 15283300 TI - Intravenous pharmacokinetics and metabolism of the reactive oxygen scavenger alpha-phenyl-N-tert-butyl nitrone (PBN) in the cynomolgus monkey. AB - The pharmacokinetics and metabolism of the antioxidant and reactive oxygen scavenger alpha-phenyl-N-tert-butyl nitrone (PBN) was examined in the male cynomolgus monkey after intravenous administration. Following an i.v. bolus dose of 5 mg/kg, plasma concentrations of PBN declined in a bi-exponential fashion. PBN demonstrated a moderate plasma clearance (CL(p) = 27.02 +/- 6.46 ml/min/kg) and a moderate volume of distribution at steady state (Vd(ss) = 1.70 +/- 0.23 l/kg), resulting in a terminal elimination half-life of 0.76 +/- 0.25 h. The corresponding area under the curve (AUC(0-infinity)) was 3.20 +/- 0.77 microg h/ml. Scale-up of the in vitro microsomal intrinsic clearance data for PBN afforded a blood clearance (CLb) value of 22 ml/min/kg, which was in reasonable agreement with the observed in vivo CLb. Monkey liver microsomes catalyzed the NADPH-dependent monohydroxylation of PBN to the corresponding alpha-4 hydroxyphenyl-N-tert-butylnitrone (4-HOPBN) metabolite. The formation of 4-HOPBN and its corresponding O-glucuronide was also discernible upon qualitative analysis of pooled (0-24 h) monkey plasma and urine samples. Less than 5% of the administered dose was excreted as unchanged PBN in the urine, suggesting that P450-catalyzed metabolism constituted the major route of PBN clearance in the primate. In conclusion, the pharmacokinetic attributes and the clearance mechanism of PBN in the cynomolgus monkey is similar to that observed in the Sprague-Dawley rat. PMID- 15283301 TI - Effect of gemfibrozil on the metabolism of pitavastatin--determining the best animal model for human CYP and UGT activities. AB - A series of studies was conducted to determine the best animal model for human CYP and UGT activities. The investigation focused primarily on the interactions occurring in the CYP- or UGT-mediated metabolism of pitavastatin, and involved in vitro and in vivo experiments. We found that the best animal models for human CYP mediated hydroxylation and UGT-mediated lactonization of pitavastatin were rats and dogs, respectively. In addition, a large difference in the metabolic properties of pitavastatin was found between monkeys and humans. In the presence of gemfibrozil, the CYP- or UGT-mediated metabolism of pitavastatin was inhibited in vitro. However, gemfibrozil treatment had no inhibitory effect on the AUC of pitavastatin and its lactone form in rats and dogs. We conclude that the plasma level of pitavastatin would not be increased by co-administration of gemfibrozil in humans. PMID- 15283302 TI - Effects of Chinese, Japanese and Western tea on hepatic P450 enzyme activities in rats. AB - Previous studies have reported that green tea effectively protects against cancers caused by various dietary carcinogens. As P450 enzymes are the major system responsible for the metabolism of many carcinogens, we hypothesise that tea consumption may alter the catalytic activities of P450 enzymes. We conducted this study to screen the effects of four different teas on the activities of P450 enzymes. Tea solutions (2.5%) were prepared by adding boiling water to tea leaves and filtering. Female Wistar rats were divided into five groups (n = 4 each); each had free access to tea solutions while the control group was supplied with water for 4 weeks. Animals were sacrificed and livers were removed for preparation of microsomes. Enzyme activities were determined by incubation of liver microsomes with the appropriate CYP substrate. The activity of CYP1A1 in livers from rats receiving Oolong (Chinese) tea (185 +/- 63 pmol/mg/min), Japanese green tea (197 +/- 22 pmol/mg/min) and Earl Grey tea (228 +/- 40 pmol/mg/min) was significantly higher (p < 0.05) than in the control group (94 +/ 34 pmol/mg/min), whereas no change was observed in the activity of CYP1A2 in any of tested animals. The hepatic activity of CYP2D6 was greater only in rats drinking Earl Grey tea compared to the controls (235 +/- 37 vs 161 +/- 41 pmol/mg/min, p < 0.05). There were also significant increases (p < 0.05) in the activity of CYP3A in livers of animals given Oolong tea (653 +/- 174 vs 382 +/- 114 pmol/mg/min) and Earl Grey tea (751 +/- 202 pmol/mg/min), while Jasmine and Japanese green tea had no significant effect. These results indicate that not all types of tea cause alterations in liver CYP enzymes as some elevated activities and some did not. Further studies are needed to determine whether there is a relationship between the effect of tea on CYP activities and anti-carcinogenesis. PMID- 15283303 TI - Metabolism of CDRI-85/92, a new potent anti-ulcer agent, involving cis-trans conversion. AB - CDRI 85/92, an anti-ulcer drug, is a new proton pump inhibitor, currently in an advanced stage of drug development. To know more about the drug it was our objective to delineate/identify the metabolic pathway as well as the enzymes responsible for the formation of metabolites. Metabolism of CDRI-85/92 (cis-5 styryl-2-oxazolidinone-4-carboxylic acid) was investigated in rat liver cellular fractions (S9, microsomes and cytosol) using reverse-phase HPLC and mass spectrometry techniques. Two major metabolites were produced by rat liver S9 fractions and reducing factor generating system from either untreated rats or phenobarbitone (PB)-pretreated rats. Incubation of CDRI-85/92 with postmitochondrial fraction (S9) for 24 h resulted in a cis to trans conversion (metabolite M2). Further cis-trans metabolizing capacity was measured separately in the cytosolic and microsomal fractions. Incubation with the cytosolic fraction resulted in an increased rate of cis-trans conversion, while the microsomal fraction showed no cis to trans conversion, thereby restricting the cis to trans conversion to Phase II enzymes, which are mainly located in the cytosol. Studies with PB-pretreated rat liver S9 fractions resulted in an increased rate of cis to trans conversion. Another metabolite was also present (M1) which was identified as an oxygenated metabolite by mass spectrometry. The major urinary metabolite from CDRI-85/92-treated Sprague-Dawley rats (20 mg/kg p.o.) was identified as M2. Studies using sulfobromophthalein and N-ethylmaleimide, as specific inhibitors of GST, showed a complete absence of metabolism, thus indicating the involvement of GST in the metabolism of CDRI-85/92. This study will be helpful in providing clues about factors influencing the bioavailability of CDRI-85/92 as well as drug drug interactions. PMID- 15283304 TI - Impact of truncated area under the curve on failed bioequivalence studies: a computer simulation analysis. AB - The common measures used in a bioequivalence study are area under the curve (AUC) and the maximum plasma concentration. Estimation of AUC requires frequent blood samples. For long half-life drugs, sampling for long periods of time may become cumbersome. To resolve this issue some investigators have suggested the use of truncated AUC in bioequivalence studies for long half-life drugs. The suggested length of time for the truncated AUC is 72 hours. Many studies have been conducted to show that truncated AUC till 72 hours is a suitable approach. However, the suitability of truncated AUC for failed bioequivalence study has not been demonstrated. This report of simulated plasma concentration versus time data evaluates the suitability of truncated AUC for failed bioequivalence study of two hypothetical drugs. The results of the study indicate that the truncated approach for the estimation of the AUC for long half-life drugs in bioequivalence studies may be useful but it also increases the probability of accepting drugs as being bioequivalent when they are not. PMID- 15283305 TI - Estimation of absorption rate constant (ka) following oral administration by Wagner-Nelson, Loo-Riegelman, and statistical moments in the presence of a secondary peak. AB - The objective of this study was to evaluate the performance of Wagner-Nelson, Loo Reigelman, and statistical moments methods in determining the absorption rate constant(s) in the presence of a secondary peak. These methods were also evaluated when there were two absorption rates without a secondary peak. Different sets of plasma concentration versus time data for a hypothetical drug following one or two compartment models were generated by simulation. The true ka was compared with the ka estimated by Wagner-Nelson, Loo-Riegelman and statistical moments methods. The results of this study indicate that Wagner Nelson, Loo-Riegelman and statistical moments methods may not be used for the estimation of absorption rate constants in the presence of a secondary peak or when absorption takes place with two absorption rates. PMID- 15283306 TI - In vitro susceptibility of Candida species isolated from cancer patients to some antifungal agents. AB - This study was undertaken to study the resistance of Candida species isolated from oropharyngeal swabs of cancer patients to ketoconazole (KET), fluconazole (FLU), amphotericin B (AmpB), and flucytosine (FCU). The most common species identified was C. albicans, followed by C. tropicalis, C. glabrata, C. famata, C. krusei, C. kefyr, and C. gulliermondii. The minimum inhibitory concentration (MIC) of the antifungal agents was evaluated by RPMI 1640 medium with microdilution method. There were no C. albicans strains resistant to KET, FLU and AmpB. All Candida isolates were found highly susceptible to AmpB (MIC AmpB < 1 microg/ml), followed by KET (MIC KET < or =8 microg/ml), FLU (MIC FLU < or =8 microg/ml) and FCU (MIC FCU < or =4 microg/ml). The main conclusion of this study is that prophylactic therapy planned according to typing and antifungal susceptibility will contribute to the prevention of invasive fungal infections in immunosuppressied oncology patients. PMID- 15283307 TI - Guideline for the management of chronic obstructive pulmonary disease (COPD): 2004 revision. AB - OBJECTIVE: To revise the South African Guideline for the Management of Chronic Obstructive Pulmonary Disease (COPD) in the light of new insights into the disease and the value of new treatment approaches and drugs. New aspects considered include: A growing awareness of the impact of COPD in South Africa, and the urgent need for prevention strategies. The role of concurrent exposures to domestic and occupational atmospheric pollution, and previous lung infections including tuberculosis. The need to consider as goals of treatment both prevention of exacerbations and improvement of quality of life (health status) of patients with COPD. The development of both long-acting beta2-agonist and anticholinergic drugs for use in COPD. Emerging evidence on a limited role for inhaled corticosteroids in the treatment of COPD. RECOMMENDATIONS: These include primary and secondary prevention; early diagnosis; staging of severity; assessment of reversibility with bronchodilator and, in some, responsiveness to corticosteroids; use of bronchodilators and other forms of treatment; rehabilitation; and treatment of complications. Advice is provided on the management of acute exacerbations, and the approach to air travel, prescribing long-term oxygen, and lung surgery including lung volume reduction surgery. Prevention, both primary and secondary, remains the most cost-effective measure in the management of COPD, and deserves more emphasis, particularly on the part of health care professionals. Primary prevention involves reducing public exposure to cigarette and other forms of smoke, and reduction of atmospheric pollution, and secondary prevention limits exposure and resultant progression in those with established disease. Spirometry is essential for the diagnosis of COPD and in staging severity. In addition, a new classification of severity that considers other indices of functional impairment is provided. Treatment involves a progression from 'as-needed' bronchodilators, through the addition of other more effective bronchodilators, usually in combination, in more severe stages. The importance of assessing potential reversibility in every patient with persistent symptoms, and of the limited role of oral and inhaled corticosteroids (ICS), is emphasised. These approaches also reduce exacerbations and may result in cost savings and improved prognosis. A practical low-cost approach to rehabilitation is proposed. OPTIONS: Treatment recommendations are based on the following: the recommendations of the Global Obstructive Lung Disease (GOLD) initiative, which provides an evidence-based comprehensive and up-to-date review of treatment options; independent evaluation of the level of evidence in support of some of the new treatment trends; and consideration of factors that influence COPD management in South Africa, including lung co-morbidity and drug availability and cost. OUTCOMES: The use of bronchodilators is driven by the presence of symptoms, but regular assessment of benefit, based on objective criteria, is essential. Several forms of treatment reduce exacerbations, the most effective of these is smoking cessation. EVIDENCE: Working group of clinicians and clinical researchers following detailed literature review, particularly of studies performed in South Africa, and the GOLD guidelines. BENEFITS, HARMS AND COSTS: The guideline pays particular attention to cost-effectiveness in South Africa, and promotes the initial use of less costly options. It rejects empirical use of corticosteroids both oral and inhaled, and promotes smoking cessation, and selection of treatment based on objective evidence of benefit. It also rejects a nihilistic or punitive approach, even in those who are unable to break the smoking addiction. VALIDATION: The COPD Working Group comprised experienced pulmonologists representing all university departments in South Africa and some from private practice. All contributed to the development of the previous version of the South African guideline, and attend international meetings. One (JRJ) represents South Africa on the GOLD Guideline Executive. GUIDELINE SPONSOR: The meeting of the Working Group of the South African Thoracic Society was sponsored by an unrestricted educational grant from Boehringer Ingelheim (South Africa) (Pty) Ltd. PMID- 15283308 TI - Guideline for office spirometry in adults, 2004. AB - OBJECTIVE: To provide clinical guidelines for office spirometry in South Africa. OPTIONS: More stringent guidelines are required for diagnostic laboratories and research. OUTCOMES: To minimise variations in standard practice and improve the quality and usefulness of spirometry in the clinical setting. EVIDENCE: Recommendations are based on key international publications as well as research publications regarding reference values for South Africans. BENEFITS, HARM AND COSTS: The medical, social and economic benefits and costs of standardisation of office spirometry in South Africa were considered in the recommendations. VALIDATION: The document has been reviewed and endorsed by the South African Thoracic Society. CONCLUSIONS: The indications for spirometry must be specific and clear. Spirometry equipment must meet internationally accepted performance standards and carry proof of validation. Equipment must be regularly calibrated and maintained. Individuals performing spirometry must be adequately trained and demonstrate a high level of competence. Subject preparation, testing and quality control of results must be carried out according to published guidelines. Finally, test results must be interpreted according to current diagnostic guidelines, taking into account the purpose of the test, appropriateness of reference values and the clinical evaluation. PMID- 15283309 TI - [The Spanish National Center for Transplants and Regenerative Medicine]. PMID- 15283310 TI - [Strategy of the candidate gene in arterial hypertension: linkage and association study]. PMID- 15283311 TI - [Diagnosis and folow up of vascular calcifications in chronic renal failure]. PMID- 15283312 TI - [Arterial hypertension: effectiveness and cost-effectiveness of its measurement with 24 hour ambulatory blood pressure monitoring]. PMID- 15283313 TI - [Cost-efectiveness of Ibersartan in type II diabetic nephropathy with hypertension. A Spanish perspective]. AB - BACKGROUND: In the Irbesartan Diabetic Nephropathy Trial (IDNT), treatment with irbesartan demonstrated 23% and 20% reductions in the combined endpoint of doubling of serum creatinine (DSC), end-stage renal disease (ESRD) or death in patients with hypertension, type 2 diabetes and overt nephropathy compared to amlodipine and control respectively. A simulation model was developed to project long-term cost consequences of the IDNT in the Spanish setting. METHODS: A Markov model simulated progression from nephropathy to DSC, ESRD and death in patients with hypertension, type 2 diabetes and overt nephropathy. Treatment-specific probabilities were derived from IDNT. Country-specific ESRD-related data were retrieved from published sources. Delay in onset of ESRD, life expectancy and mean lifetime costs were calculated for patients with baseline age 59 years. Future costs were discounted at 6% per annum, and clinical benefits were discounted at 0% and 6% per annum. Extensive sensitivity analyses were performed. RESULTS: Onset of ESRD was delayed with irbesartan by 1.41 and 1.35 years versus amlodipine and control respectively. When a 25-year (lifetime) horizon was considered, delay in ESRD onset led to anticipated improvements in life expectancy (discounted at 6% shown in brackets) of 0.46 (0.21) years versus amlodipine and 0.75 (0.37) years versus control. Irbesartan was associated with cost savings of 13,673 Euro and 7,632 Euro patient versus amlodipine and control respectively. The results were robust under a wide range of plausible assumptions. CONCLUSIONS: Treating patients with hypertension, type 2 diabetes and overt nephropathy using irbesartan was both cost- and life-saving compared to amlodipine and control in the Spanish setting. PMID- 15283314 TI - [Evaluation of individual risk and mortality related factors in acute renal failure]. AB - OBJECTIVES: To evaluate the Individual Severity Score (ISS) as a predictor of mortality in patients with acute renal failure (ARF) and to determine mortality associated factors. METHODS: Clinical records of patients with de diagnosis of ARF at the Hospital Arzobispo Loayza (HAL) of Lima (Peru) between the years 1995 and 2000 revised. The optimal cut point for sensitivity and specificity of the ISS was assessed using ROC curve analysis. RESULTS: 128 clinical records of patients with ARF were revised. The mortality rate was 10,94 % (14/128). Patients who died were significatively older and showed more frequently hypotension, jaundice, need of mechanical ventilation and coma. The optimal cut point for the ISS was 0,74 (sensitivity of 71% and specificity of 100%). The area under the curve value was 0,91 (CI95 0,81-1,03). CONCLUSION: The ISS was shown to be a cheap, easy clinical tool with an elevated discriminant ability in the study population. Mortality associated factors were similar as previously reported. PMID- 15283315 TI - [Vascular access surveillance with blood flow monitoring: a prospective study with 65 patients]. AB - BACKGROUND: Periodic intra-access blood flow rate (QA) monitoring is the preferred method for vascular access (VA) surveillance (NKF-K/DOQI, update 2000). OBJECTIVES: 1) To determine the ultrafiltration (UF) method accuracy for early detection of VA stenosis. 2) To evaluate the hemodynamic effect of elective VA intervention (angioplasty or surgery). 3) To define the impact of periodic QA monitoring using the UF method combined by elective VA intervention on VA thrombosis. PATIENTS AND METHODS: We prospectively monitored QA during hemodialysis (HD) in 65 ESRD (mean age 64.9 +/- 11.4 years, 20% diabetes) patients over 1 year period. All patients undergoing HD in the Hospital de Mollet by arteriovenous fistula (89.2%) or graft 10.8%. QA was measured at least every 4 months by the UF method using the Crit Line III Monitor. Fifty (77%) patients were included at the beginning of the study period and the remaining 15 (23%) were added later when they started HD. All patients with absolute QA <700 ml/min or decreased >20% from baseline met criteria of positive evaluation (PE) and were referred for angiography (AG) plus subsequent preventive intervention (angioplasty or surgery) if VA stenosis >50%. We also studied 94 not QA monitored patients since the beginning of the study period (mean age 64.6 +/- 13.7 years; 12.8% diabetes) that undergoing HD simultaneous in the Institut Nefrologic Granollers. RESULTS: We performed 200 QA measurements in 509 months of follow-up. The overall mean QA was 1176.7 +/- 491.8 ml/min (range, 380.5-2904.0 ml/min). Three patients (4.6%) thrombosed VA. Nineteen (29.2%) patients had PE; none of them clotted VA. The AG was performed in 84.2% (16/19) patients with PE and all of them (16/16) showed VA stenosis > or =50%; 31.2% (5/16) were lost to follow-up (3 death, 2 transplantation); of the remaining explored patients (11/16), 72.7% (8/11) underwent intervention (3 angioplasty, 5 surgery). The mean QA increased from 577.2 +/- 108.2 ml/min to 878.1 +/- 264.4 ml/min postintervention (p=0.005). The positive predictive value, negative predictive value, sensitivity and specificity of UF method for VA stenosis were 84.2%, 93.5%, 84.2% and 93.5%, respectively. VA thrombosis rate in our 50 beginners QA monitored patients (mean age 64.5 +/- 1 1.4 years; 20% diabetes) was lower (2/50, 4%) compared to 94 not QA monitored patients (16/94, 17%) (p=0.024). CONCLUSIONS: 1) QA monitoring using the UF method allows an early diagnosis of VA stenosis. 2) Serial QA measurement by UF method can be used in assessing the functional response to corrective VA intervention. 3) Periodic VA surveillance by QA measurements using the UF method combined with elective intervention results in reduced VA thrombosis. PMID- 15283316 TI - [Diagnosis and prevention of tuberculosis in hemodialysis patients. a new old problem?]. AB - Patients with chronic renal failure undergoing hemodialysis are at increased risk of developing tuberculosis because of impaired cellular immunity. Most cases are due to reactivation of disease and are known to develop the complication of extrapulmonary tuberculosis more frequently than the general population, and this makes the disease difficult to diagnose, delaying the establishment of appropiate therapy. We evaluated 39 patients undergoing hemodialysis treatment in the course of a 3 year period, therewere. Tuberculosis control program was developed to evaluate all patients newly admitted to the renal dialysis program. All of the patients were skin-tested initially with use 2 U.I. tuberculin PPD RT23, and 16 patients had positive results. Three cases of tuberculosis -registered that developed in patients, and in exposed patients, 3 patients with negative test result 2 months later had positive test. No relationship was found between the results of the tests and age, sex, renal disease, other pathologies or previous tuberculosis and albumin figures. However, positive patients had higher levels of total leukocytes. The period from undergoing hemodialysis until mantoux testing was performed was lower in those patients who resulted positive: 9.4+/-13.6 months versus 32.9+/-39 months in negative patients. The use of mantoux testing is important as a sieve system in hemodialysis patients as long as is associated with other diagnosis techniques, detecting that way those cases at risk of developing tuberculosis and latting perform further studies and isoniacida prophylaxis. Establishing isoniacida prevention in hemodialysis patients can avoid the development of tuberculosis in patients at risk as well as prompt detection makes easier the establishment of right therapy. PMID- 15283317 TI - [Impact of a quality program in hemodialysis]. AB - One of the requirements of a health care quality management system is to be able to established clinical performance measures (CPM) for its key organisation processes. We described some of the performance measurement that has been used in our hemodialysis unit, since the implementation in the year 2001, of a Quality Management System (QMS). We analyze and compare the effect that the introduction of a ISO 9002 based QMS had in our CPM during the period 2001-2002 (post QMS) vs. the two previous years -1999-2000- (pre QMS). METHODS: We defined several CPM for assessment of hemodialysis adequacy and medical management that covered : Anemia, iron status renal osteodystrophy, hemodialysis prescription and nutritional status , follow up of the established guidelines for vascular access care and prevention of nosocomial infections water quality and general performance outcome like annual crude mortality rate and hospitalization (express as hospital days/patient year). RESULTS: No significant difference was found between both periods regarding annual crude mortality( pre QMS 8.37% vs post QMS 8.95%) or the hospitalization rate ( pre 0.47 patient-days vs. post 0.52 patient-days) . There was a significant difference after implementation of the quality system in the average hemoglobin levels (pre 11.3 +/- 1.5 vs. post 11.9+ +/- .5 p <0,001). Ferritin levels (pre 220 +/- 162 vs. post 313 +/- 373 p<0.01), albumin levels (pre 3.61 +/- 0.46 vs. post 3.82 +/- 0.56 p<0.001) and KTV>1.2 (pre 1.41 +/- 0.26 vs. post 1.50 +/- 0.33 p<0.001). The transferring saturation index (TSI) was unchanged (pre 27.98 +/- 14.39 vs.. post 29.4 +/- 16.66 p=0.11). There was a significant decrease in the average PTH levels (pre 234.9 +/- 285 vs. post 174 +/ 174 p< 0.0001) PTH>300 pg/ml (pre 23.7% vs. post 16.4% p<0.001) calcium levels (pre 10.02 +/- 0.99 vs. 9.83 +/- 0.88 p<0.001), phosphorus (pre 5.50 +/- 1.55 vs. post 5.01 +/- 1.47 p<0.001) as well as serum calcium levels >11 mg/dl (pre 14.6% vs. post 11% p<0.001) and phosphorus >6 g/dl (pre 34% post 21.5% p<0.001). Although the average serum potassium levels decrease (pre 5.51 +/- 0.85 vs. post 5.40 +/- 0.87), the percentage of patients with potassium over 6.5 meq/l was similar in both periods (pre 11.5% vs. post 10. 1%). The number of native A-V vascular access was similar in both periods and above the current DOQI Recommendations. Nevertheless, there was a gradual decrease in native A-V fistula, associated with an increase on the use of permanent catheters. The number of incident patients with a permanent catheter as the only vascular access for hemodialysis increased from 0% in the year 1999, 2000 and 2001 to 6.98% in 2002. There was no hepatitis B and C seroconversion detected in both periods. CONCLUSION: From our study we concluded that regular follow-up of quality performance measurement associated with an ongoing corrective action, promotes an improvement of the outcome measures results. PMID- 15283318 TI - [Parathyroid function in patients with gland autotransplantation]. PMID- 15283319 TI - [Colitis due to cytomegalovirus in a dialysis patient]. AB - Cytomegalovirus (CMV) is an ubiquitous agent and a pathogen in all age groups. Although CMV disease in normal adults is not very usual, the virus is well known to produce severe symptoms, mostly in immunocompromised patients. Chronic hemodialysed patients constitute a risk population for developing CMV infection, nevertheless, clinical manifestations are not usual. One chronic renal failure patient who developed acute and severe colitis due to CMV infection is presented. Of interest are, the rarity of this case, the favorable clinical course after the treatment and the differential diagnosis with other gastrointestinal disorders frequently found in renal patients. PMID- 15283320 TI - [Acute hyperphosphatemia secondary to phosphate administration for bowel preparation]. AB - We report a 75-years-old woman, stable on a three-weekly hemodialysis program over a period of 3 years, who develop acute hyperphosphatemia secondary to phosphate administration for bowel preparation. The quick clinical diagnosis and the treatment with intensive hemodialysis resulted in a correction of hyperphosphatemia, hypocalcemia, acidemia and other electrolyte abnormalities. The phosphate cathartics are contraindicated in patients with severe renal insufficient or in dialysis program. Our case shows the severe side effects secondary to injudicious use of sodium phosphate cathartics. PMID- 15283321 TI - [Corynebacterium urealyticum in renal trasplantation. CT and sonography imaging characteristics of encrusted cistitis and pielitis]. AB - PURPOSE: Its described three cases of Corynebacterium urealyticum (CU) infection in patients with renal transplantation and one of its most serious consequences: encrusted pyelitis and cystitis. It is explained the principal keys for its diagnosis, based in the appearance of alkaline pH in in urine analysis (alkaline urine), positives urinary cultures for CU, and the CT and US studies revealed the characteristic images of calcifications in the wall of renal pelvis and bladder. PATIENTS: Three male patients with renal transplantation and CU infection that caused encrusted pyelitis in two of the cases and encrusted cystitis in one case. RESULTS: Calcifications of the urinary tract were noticed in CT in all the patients. In two cases bladder stones were linear, and in the third case they were fundamentally coarse and placed in pelvis. The diagnosis suspicion showed by the images was confirmed by the use of prolonged urine cultures, necessary for detecting CU. All the patients were treated with vancomycin, with success in two of the cases and, finally needing surgery, and after loss of the graft, in the other case. CONCLUSION: Encrusted pyelitis and cystitis are cronic and severe infections of the urinary tract. Calcic struvite incrustations in the urothelium are characteristics of this infection. CT is a choice technique for the diagnosis and followup of the calcifications after treatment. PMID- 15283322 TI - [Post-partum focal and segmental glomerulosclerosis. Primary nephropathy or pre eclampsia related?]. PMID- 15283323 TI - [Erythropoietin resistance as initial presentation of ANCA-positive intra alveolar hemorrhage in a hemodialysis patient]. PMID- 15283324 TI - [Malignant hypertension and irreversible kidney failure associated with oral contraception pills intake]. PMID- 15283325 TI - [Renal insufficiency and Rokitansky syndrome]. PMID- 15283326 TI - [Stressors, stress reactions, and survival of bacteria (a review)]. AB - Recent data on the molecular mechanisms of stress responses of bacteria are reviewed, with emphasis on their reactions to a variety of stressors (heat, oxidation, cold, osmotic shock, etc.). Mechanisms underlying the phenomenon of sensoring are discussed. It is shown that cross-resistance to stressors and cell to-cell communication of bacteria, mediated by chemical metabolites, affect their survival in food products. Stress-antiagonizing activity of bacteria is discussed in relation to food product biotechnology. PMID- 15283327 TI - [Transformation of phospholipids by cabbage phospholipase D in mixed micelles containing 3-[(3-cholamidopropyl)dimethylammonio]propanesulfonate]. AB - We compared the activities of cabbage phospholipase D during hydrolysis and transesterification of phosphatidylcholine in mixed micelles of surface-active compounds with various physicochemical properties. Mixed micelles of phospholipids and 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (ratio, 1:2) were among the best substrates. Hydrolysis and transphosphatidylation were studied in micelles containing 3-[(3 cholamidopropyl)dimethylammonio]-1-propanesulfonate. Mixed micelles of phosphatidylcholine and 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate may serve as a new substrate for measurements of phospholipase D activity and preparative isolation of phospholipids using this enzyme. PMID- 15283328 TI - [Isolation and partial characterization of DNA topoisomerase I from the nucleoids of white mustard chloroplasts]. AB - DNA topoisomerase was isolated for the first time from nucleoids of white mustard (Sinapis alba L.) chloroplasts. The enzyme had a molecular weight of 70 kDa; it was ATP-independent, required the presence of mono- (K+) and bivalent (Mg2+) cations, and was capable of relaxing both negatively and positively supercoiled DNA. These results suggest that the enzyme isolated belongs to type IB DNA topoisomerases. PMID- 15283329 TI - [Conversion of L-cystine and L-cysteine to taurin by the enzyme systems of liver cells]. AB - The kinetics of conversion of sulfur-containing amino acids L-cystine and L cysteine to taurin by the enzyme system of cattle liver cells was studied, and a mathematical model was developed. It was shown that L-cystine and L-cysteine conversion obeyed the Michaelis-Menten equations of serial-sequential conversions with regard to inhibition by the final product and inactivation. The yield of taurin under the optimized conditions of L-cystine and L-cysteine conversion (temperature, 40 degrees C; pH 1.5 and 3.0, respectively; and addition of enzyme preparations in five equal portions at 2-h intervals) was in the range 80-85% of the substrate weight. PMID- 15283330 TI - Cloning and expression of the Vitreoscilla hemoglobin gene in Enterobacter aerogenes: effect on cell growth and oxygen uptake. AB - The hemoglobins found in unicellular organisms show a greater chemical reactivity, protect cells against oxidative stress and hence have been implicated in a wider variety of potential functions than those traditionally associated with animal and plant hemoglobins. There are well-documented studies showing that bacteria expressing Vitreoscilla hemoglobin (VHb), the first prokaryotic hemoglobin characterized, have better growth and oxygen uptake rates than VHb counterparts. PMID- 15283331 TI - [Study of Ralstonia eutropha culture producing polyhydroxyalkanoates on products of coal processing]. AB - Kinetic indices of growth, polyhydroxyalkanoate (PHA) accumulation, and gas exchange have been studied in a culture of the carbon monoxide-resistant hydrogen strain Ralstonia eutropha B-5786 grown on a gaseous substrate (GS) obtained by lignite gasification. The GS was shown to be suitable for PHA production. To increase the degree of GS consumption, various modes of gas supply to the culture were tested. Based on the results, an algorithm was developed for calculating and controlling gas-exchange parameters in the PHA-accumulating culture of Ralstonia eutropha, grown on a new GS allowing high polymer yields (up to 75%) and degrees of the substrate utilization (up to 90%). PMID- 15283332 TI - [Antibacterial effects of water-soluble low-molecular-weight chitosans on different microorganisms]. AB - Low-molecular-weight chitosans with a viscosity-average molecular weight (Mv) of 5 to 27 kDa and equal degree of deacetylation (DD, 85%) were highly active against Pseudomonas aureofaciens, Enterobacter agglomerans, Bacillus subtilis, and Bifidobacterium bifidum 791, causing death of 80 to 100% of cells. An exception to this tendency was Escherichia coli, for which the rate of cell death, induced by the 5-kDa chitosan, was 38%. The antibacterial effect was manifested as early as 10 min after incubation of 12-kDa chitosan with B. subtilis or E. coli cells. Candida krusei was almost insensitive to the above crab chitosans. However, Candida krusei was highly sensitive to chitosans with Mv 5, 6, 12, 15.7, and 27 kDa: the minimum inhibitory concentration (MIC) varied from 0.06 to 0.005%. Chitosans with M, 5, 12, and 15.7 kDa exerted an antibacterial effect on Staphylococcus aureus. Chitosans with Mv 5, 15.7, and 27 kDa had no effect on Bifidobacterium bifidum ATCC 14893. The antibacterial effect of the 4-kDa chitosan on E. coli and B. bifidum 791 increased with DD in the range 55-85%. PMID- 15283334 TI - [Low-temperature microbial degradation of crude oil products differing in the extent of condensation]. AB - Out of the 30 strains capable of oil degradation at 4-6 degrees C, four were selected by the ability to degrade 40% of the oil substrate present in the growth medium: Rhodococcus spp. DS-07 and DS-21 and Pseudomonas spp. DS-09 and DS-22. We studied the activity of these strains as degraders of oil products of various condensation degrees (crude oil, masut, petroleum oils, benzene resins and ethanol-benzene resins) at 4-6 degrees C. The maximum degrees of degradation of masut and ethanol-benzene resins were observed in Pseudomonas spp. DS-22 (17.2% and 5.2%, respectively). The maximum degradation of petroleum oils and benzene resins was observed in Rhodococcus spp. DS-07 (40% and 16.6%, respectively). The strains provide a basis for developing biodegrader preparations applicable to bioremediation of oil-polluted sites under the conditions of cold climate. PMID- 15283333 TI - [Degradation of 2,4-dinitrophenol by free and immobilized cells of Rhodococcus erythropolis HL PM-1]. AB - Degradation of 2,4-dinitrophenol (2,4-DNP) by the cells of Rhodococcus erythropolis HL PM-1 was studied. The enzymes involved in 2,4-DNP degradation were inducible, and their resynthesis took place during the process. Cell immobilization by embedding into agar gels decreased the degrader activity. Maximum rates of 2,4-DNP degradation by free and immobilized cells were 10.0 and 5.4 nmol/min per mg cells, respectively. The concentration dependence of 2,4-DNP degradation was typical of substrate inhibition kinetics. The immobilized cells were used in a model reactor designed for 2,4-DNP biodegradation. Its maximum capacity was 0.45 nmol/min per mg cells at a volumetric flow rate of 20 h-1. The reactor operated for 14 days without losing capacity; its half-lifetime equaled 16 days. PMID- 15283335 TI - [Intensification of microbial degradation of crude oil and oil products in the presence of perfluorodecalin]. AB - The possibility of using perfluorinated organic compounds for growing microorganisms and degrading xenobiotics has been demonstrated for the first time with perfluorodecalin (PFD), a gas-transporting component of the blood substitute Perftoran. In particular, this is promising for intensifying microbial degradation of oil and oil products and production of biodegrader biomass in synthetic mineral media. Addition of PFD to a mineral medium with crude oil and masut increased 4.5-10.2 times maximum concentrations and growth rates of all bacterial strains under study (Pseudomonas, Rhodococcus, and Bacillus genera). The degree of oil product consumption was increased 8.7-12.7 times. PMID- 15283336 TI - [Zeta-potential of n-alkane emulsion droplets and its role in substrate transport into yeast cells]. AB - Zeta-potential of n-alkane droplets, formed by fatty acids, were studied in model systems of culture liquid of yeasts (Candida maltosa) capable of utilizing n alkanes. The value of zeta-potential was found to depend on the droplet size. The negative zeta-potential of submicron droplets was so high that it prevented the droplets from being coagulated with cells possessing a high negative zeta potential. The dominant role of submicron n-alkane droplets in the kinetics of yeast growth could be accounted for by the existence of a mechanism regulating contact interactions of individual cells with the droplets followed by uptake of the substrate. PMID- 15283337 TI - [Selection and study of potent lactose-fermenting yeasts]. AB - Whey-fermenting Kluyveromyces cultures were revealed among 105 yeast strains assimilating lactose. Eighteen most potent strains isolated from milk products fermented galactose, sucrose, and raffinose, in addition to lactose. Many yeast strains fermented inulin. Most strains were resistant to cycloheximide and grew in medium containing glucose, NaCl, and ethanol at concentrations of up to 50, 11 12, and 10-12%, respectively (4 degrees C). Three strains had mycocinogenic activity. After fermentation of whey with selected yeast strains at 30 degrees C for 2-3 days, ethanol concentration was 4-5%. PMID- 15283338 TI - [Degradation of the herbicide atrazine by the soil mycelial fungus INBI 2-26(-)- a producer of cellobiose dehydrogenase]. AB - Nonsporulating mycelial fungi producing cellobiose dehydrogenase (CDH) and isolated from soils of South Vietnam with high residual content of dioxins are capable of growing on a solid medium in the presence of high atrazine concentrations (to 500 mg/l). At 20 and 50 mg/l atrazine, the area of fungal colonies was 1.5-1.2-fold larger, respectively, compared with control colonies of the same age, whereas development of the colonies at 500 mg/l atrazine was delayed by 5 days, compared with controls grown in the absence of atrazine. Surface cultivation of the fungus on a minimal medium with glucose as a sole source of carbon and energy decreased the initial concentration of atrazine (20 mg/l) 50 times in 40 days; in addition, no pronounced sorption of atrazine by mycelium was detected. This was paralleled by accumulation in the culture medium of extracellular CDH; atrazine increased the synthesis of this enzyme two- to threefold. Accumulation of beta-glucosidase (a mycelium-associated enzyme) and cellulases preceded the formation of CDH. PMID- 15283339 TI - [Penicillium expansum, a resident fungal strain of the orbital complex Mir, producing xanthocillin X and questiomycin A]. AB - It was demonstrated that the fungus Penicillium expansum 2-7, a resident strain of the orbital complex Mir, which became dominating at the end of a long-term space flight, formed biologically active secondary metabolites (antibiotics). Using physicochemical methods, these metabolites were identified as xanthocyllin X and questiomycin A. Time courses of their biosyntheses during growth and development of the producer culture were studied. Addition of zinc to the culture medium affected both the growth of the culture and the biosyntheses of the antibiotics. The concentrations of zinc in the medium, optimum for xanthocyllin X and questiomycin A production, amount to 0.3 and 3.0 mg/l, respectively. PMID- 15283340 TI - [Isolation of cell membranes from Saccharomyces cerevisiae for evaluation of the protein composition]. AB - We describe a simple method for the isolation of membrane fractions from Saccharomyces cerevisiae yeasts, containing the complex of plasma membranes and cell walls. The method is based on cell disruption on an INBI flow disintegrator. This device spares subcellular structures, which simplifies the isolation of cell membranes. The membrane fraction obtained by this method was suitable for studies of protein composition of these structures by means of two-dimensional electrophoresis. PMID- 15283341 TI - [Hydrolysis of chitosan in lactic acid]. AB - Effects of molecular weight and degree of acetylation on the hydrolysis of chitosan in dilute lactic acid were studied. It was demonstrated that the higher were the values of both parameters, the more rapid were the decreases in viscosity and viscosity-average molecular weight of chitosan. PMID- 15283342 TI - [Determination of polyphenolic complex in wines by electrochemical methods and using the enzymes tyrosinase and laccase]. AB - Several red wines were studied to find a correlation between physicochemical parameters characterizing the antioxidant status of wine and total content of phenols in samples. The content of dissolved oxygen (its value varied from 0.75 to 3.28 mg/ml), pH (3.10-3.63), redox potential (-186 to -106 mV), mass concentration of free and total sulfur dioxide (10-30 and 36-200 mg/dm3, respectively), absorption spectra, and total phenol content were determined. The wines fell into two main groups-with a relatively low (1850-2050 mg/dm3) and high (2300-2900 mg/dm3) contents of polyphenols. It was demonstrated that physicochemical parameters (except for the content of sulfur dioxide) correlate with the total phenol content in the wines studied. PMID- 15283343 TI - [Effects of treatment with a composite preparation (2-chloroethylphosphonic acid and methacide) or butylated hydroxyanisole on ethylene release in apples]. AB - We studied the effect of a Russian composite preparation (2-chloroethylphosphonic acid and methacide) and butylated hydroxyanisole on ethylene release in whole fruit and peel disks of two apple cultivars, Antonovka obyknovennaya (Antonovka) and Simirenko's rennet (Simirenko). Treatment with the composite preparation was followed by an increase in ethylene release from whole apples and peel disks. The development of microbial infection (fruit rot) in whole apples became less pronounced after the treatment. Treatment of whole apples with the antioxidant butylated hydroxyanisole (BHA) increased the intensity of ethylene release during the first subsequent days; thereafter, ethylene release decreased and was 10-15% lower than in the control on days 10-12. In model experiments, BHA decreased ethylene release from apple peel disks below control levels as early as on day 1 after the treatment. Antonovka apples gave quick responses to the treatment. In the late-ripening Simirenko apples, the response persisted for a longer period. Our results suggest that treatment with physiologically active preparations affects ethylene release, ripening, and preservation of apples in storage. PMID- 15283344 TI - [Composition and structure of galactomannan from the seed of Gleditsia ferox Desf]. AB - Galactomannan, a heteropolysaccharide with a molecular weight of 1660 kDa, was isolated form the seed of Gleditsia ferox Desf., introduced in Russia, with a yield of 18.9%. Its aqueous solutions were optically active ([alpha]D = +30.5 degrees) and highly viscous ([eta] = 1430 ml/g). Analysis of the heteropolysaccharide using chemical, enzymatic, and chromatographic procedures showed that it consists of D-mannopyranose and D-galactopyranose residues (molar ratio, 2.54:1). The main chain of this galactomannan consists of 1,4-beta-D mannopyranose residues, 39.2% of which are substituted at C6 with single residues of alpha-D-galactopyranose. The probability of occurrence of mannobiose units differentially substituted with galactose was determined by 13C-NMR data and equaled, respectively, 0.37, 0.47, and 0.16 for non-substituted Man-Man units, monosubstituted Gal(Man-Man) and (Man-Man)Gal units taken together, and for the disubstituted Gal(Man-Man)Gal units. PMID- 15283345 TI - [II International Congress in Moscow "Biotechnology: status and prospects of development"]. PMID- 15283346 TI - Hypoplastic left heart syndrome: conventional surgical management. AB - Surgical palliation for hypoplastic left heart syndrome has become standard therapy in many centers throughout the world. Significant improvements in management, from preoperative diagnosis and intraoperative technique to postoperative care, have resulted in a dramatic increase in survival. With this improved survival, however, come new challenges and unforeseen problems. The first stage of the reconstructive surgery remains the highest risk procedure for the reason that establishing a stable circulation that is dependent on a single, volume-overloaded right ventricle with the potential for jeopardized coronary blood flow remains a surgical challenge. In this review, current management strategies for the conventional reconstructive surgical approach are examined. PMID- 15283347 TI - Practical use of alpha blockade strategy in the management of hypoplastic left heart syndrome following stage one palliation with a Blalock-Taussig shunt. AB - Alpha blockade with phenoxybenzamine has been used in the postoperative management after the Norwood operation. The principle of this approach is the increase in systemic cardiac output by maximal dilation of the systemic circulation. This effect results in a more stable parallel circulation through prevention of fluctuations in systemic vascular resistance in the early postoperative period. We have incorporated this approach in our postoperative management protocol and have observed a decrease in sudden cardiac collapse. This article describes the practical use of alpha blockade strategy, its clinical application, and the most common clinical scenarios observed during the initial hours following stage one palliation with a Blalock-Taussig shunt for hypoplastic left heart syndrome in our cardiac surgery unit. PMID- 15283348 TI - Routine use of mechanical ventricular assist following the Norwood procedure. AB - Conventional postoperative management after the Norwood procedure in patients with hypoplastic left heart syndrome suffers from three main shortfalls. First, the early postoperative care is often labor-intensive and ironically (despite sometimes heroic efforts), when babies die, health care providers often feel like failures, and in the worst scenarios, surgeons or other physicians create cultures of blame. Second, hospital survival is inconsistent in most centers, especially the ones with small surgical volume and limited experience. Third, survivors often show evidence of significant neurologic impairment. To address these postoperative problems, we have adopted the strategy of routinely placing all our patients with hypoplastic left heart syndrome on mechanical circulatory support immediately after their Norwood procedure. No attempt was made to balance the systemic and pulmonary circulation. Because an oxygenator was not used in the circuit, a much lower level of anticoagulation was used. Once the lactate level normalized, the amount of mechanical circulatory support was weaned. Since January of 2001, 23 patients have been managed using this strategy. The average time of mechanical circulatory support was approximately 3 days and has decreased to 2 days in more recent experience. The overall incidence of complications was 22%, and overall hospital survival was 87%. Neurodevelopmental testing before the Glenn procedure was normal for all patients tested. Routine postoperative use of mechanical ventricular assist device can support the increased cardiac output demands of infants following Norwood procedure and results in a stable postoperative convalescence. This strategy can simplify postoperative management, lead to excellent hospital survival, and possibly augment cerebral oxygen delivery resulting in improved neurologic outcomes for these patients. PMID- 15283349 TI - Right ventricle-pulmonary artery shunt in first-stage palliation of hypoplastic left heart syndrome. AB - This article reviews our experience using a prosthetic conduit between the right ventricle (RV) and the pulmonary artery (PA), in lieu of the more traditional aortopulmonary shunt, for infants undergoing surgical palliation of hypoplastic left heart syndrome. Thirty-three consecutive infants underwent Norwood procedure between February 1998 and November 2003, using an RV-PA conduit. There were 31 hospital survivors (94%) and 25 patients have undergone a stage II bidirectional Glenn anastomosis with an additional two deaths. Nine patients have undergone completion Fontan. This technique provides reproducible results, simplifies postoperative management, and improves outcome, especially for "low volume" programs. PMID- 15283350 TI - Home monitoring of infants after stage one palliation for hypoplastic left heart syndrome. AB - Despite improved early results with the Norwood procedure (stage one palliation), patients remain with at-risk anatomy and interstage mortality continues to be a limitation of staged single ventricle palliation. Retrospective analyses have implicated residual or recurrent anatomic lesions as well as intercurrent illness as causes of interstage mortality. We hypothesized that potentially life threatening anatomic lesions and illnesses would be manifest before serious physiologic impact by alteration in arterial saturation, failure to gain weight or in the case of dehydration, acute weight loss. As a result, we developed a home monitoring program of daily weights and oxygen saturations to earlier identify those patients at increased risk for interstage death. Frequent monitoring of these physiologic variables between stage one and two palliation identified life-threatening anatomic lesions and illness and permitted timely intervention that ultimately improved survival. All 36 survivors of the stage one palliation discharged from the hospital and entered into the home monitoring program survived the interstage period. PMID- 15283352 TI - Fontan completion without surgery. AB - An ideal Fontan procedure would minimize complications while maximizing flow dynamics through the circuit. We report our early experience with a new combined surgical/transcatheter approach which enables a nonoperative, transcatheter Fontan completion. The conceptual rationale of this management strategy, as well as surgical and catheterization techniques, are discussed. PMID- 15283351 TI - Neurodevelopmental outcomes in hypoplastic left heart syndrome. AB - Infants with hypoplastic left heart syndrome (HLHS) are now surviving to school age and early adulthood because of surgical palliation with either staged reconstruction or transplantation. Although in some respects these patients are doing remarkably well, recent investigations have shown that they are at risk for cognitive, neuromotor, and psychosocial problems. This article examines the neurodevelopmental outcome for children and adolescents with HLHS. In addition, hemodynamic and genetic factors unique to this patient population that may impact longer-term outcome are explored. The continued evolution of the pre-, intra- and post-operative care of the patient with HLHS has significantly improved early survival. Concomitant changes in care, such as selective cerebral perfusion and neurologic monitoring, aimed at reducing neurologic insult may also favorably impact later neurodevelopmental status. PMID- 15283353 TI - Heart transplantation for the failing Fontan circulation. AB - The failing Fontan circulation presents difficult treatment challenges. When Fontan revision and or intervention for treatable arrhythmias are not feasible, heart transplantation is the only therapeutic option. Particular challenges presented by these patients include limited ability to assess hemodynamics, complex anatomy, multiple prior procedures, and unique underlying pathologic states. These issues complicate the decision-making process for further surgical intervention verses transplantation. The pretransplant evaluation, transplant operation, and postoperative management are more problematic for these patients compared with most patients undergoing transplantation. Consequently, failing Fontan patients constitute one of the highest risk heart transplant subsets. PMID- 15283354 TI - Tetralogy of Fallot with absent pulmonary valve syndrome. AB - Tetralogy of Fallot with absent pulmonary valve syndrome is a rare variant of tetralogy of Fallot marked by severe pulmonary insufficiency in utero and significant, sometimes massive, dilation of the pulmonary arteries. This syndrome can be associated with airway abnormalities that can cause respiratory compromise. Patients who present with respiratory symptoms early in life continue to present a major clinical challenge associated with relatively high mortality rates, typically in the 20% to 30% range. A variety of surgical techniques have been developed to address absent pulmonary valve syndrome that may improve long term outcomes. PMID- 15283355 TI - Total cavopulmonary connection to one lung. AB - The status of pulmonary circulation is of utmost importance to the success of the modified Fontan operation. Pulmonary artery distortion, hypoplasia of the total pulmonary vascular bed, and elevated pulmonary vascular resistance are all risk factors for adverse outcome after modified Fontan operations. In cases of irreparable acquired atresia of one or the other branch pulmonary artery, clinicians are forced to contemplate the possibility of total cavopulmonary connection to one lung. The combined experience of the authors with 12 cases suggests that the likelihood of operative survival following Fontan's operation to one lung is predicted based on the usual hemodynamic parameters: pulmonary artery pressure and flow, ventricular end diastolic pressure, transpulmonary gradient, and pulmonary vascular resistance. In this series, there were no operative mortalities among patients undergoing Fontan's operation to one lung, all of whom meet the usual criteria for hemodynamic acceptability. There may, however, be a higher incidence of protein-losing enteropathy than in Fontan patients with normal pulmonary vascular beds. All possible means of resuscitating the lost elements of the pulmonary vascular bed and re-establishing pulmonary artery continuity should be attempted to minimize pulmonary vascular capacitance of patients undergoing Fontan's operation. It is clear, however, that the presence of only one pulmonary artery does not in and of itself preclude satisfactory outcome. PMID- 15283356 TI - The role of aortic translocation in the management of complex transposition of the great arteries. AB - The surgical management of transposition of the great arteries with a ventricular septal defect and left ventricular outflow tract obstruction still remains a significant challenge despite major advances in pediatric cardiovascular surgery. Almost four decades after its inception, the Rastelli procedure continues to be the standard surgical technique used in the management of these patients, despite less than optimal long-term results. Aortic translocation with biventricular outflow tract reconstruction is an alternative surgical technique that creates anatomically sound connections between the ventricles and their respective great arteries, which could result in better long-term outcomes. This technique is especially useful in patients with transposition of the great arteries/ventricular septal defect/left ventricular outflow tract obstruction who have an inlet or restrictive ventricular septal defect, a small right ventricle, a straddling atrioventricular valve, or an aberrant coronary artery crossing the right ventricular outflow tract. PMID- 15283357 TI - Pulmonary atresia, ventricular septal defect, and multiple aorta pulmonary collateral arteries. AB - Pulmonary atresia with ventricular septal defect and major aorta pulmonary collaterals arteries is a rare and complex congenital cardiac defect. There is considerable variability in the anatomy, morphology, and geometry of the native pulmonary arteries and the collateral vessels. While the ultimate goal of therapy is a biventricular correction with complete unifocalization, establishment of right ventricular to pulmonary arterial continuity, and closure of all intracardiac defects, achieving this endpoint can be frustrating and difficult. A carefully considered approach for each individual patient is required. Patients with appropriate anatomy may undergo a definitive single-stage unifocalization and biventricular correction in early infancy. Patients with less favorable anatomy will require a more eclectic approach. While our knowledge of the genetics of this defect is rudimentary, further advances in genetic understanding and technology hold tremendous promise for the development of future therapies. PMID- 15283358 TI - Surgical outcome for congenital heart malformations in the adult age: a multicentric European study. AB - Congenital heart diseases are currently treated in the pediatric age. However, a conspicuous number of patients requires treatment in the adult age. This study has been undertaken by members of the European Congenital Heart Surgeons Association with the aim of evaluating the impact of cardiac surgery in this particular age group. We have collected data from 1,247 patients who underwent 1,287 operations during a 5-year period between January 1, 1997 and December 31, 2001. Patients were divided into three groups: (1) palliative procedures (4.4%), operation performed to improve patients' clinical status without restoring normal anatomy or physiology; (2) repair (79.3%), operation performed to achieve an anatomic or physiologic repair with separation of the pulmonary from the systemic circulation (included in this group are also Fontan-type repair and one and a half ventricle repair); (3) reoperation (16.3%), all the reoperations performed after repair (either anatomic or physiologic). Hospital mortality (within 30 days) was 2.4% (range, 0% to 15.3% in different centers). Kaplan-Meier estimates shows a 94% survival at 4 years, which is higher for repair (95%) as compared with reoperations (92%) or palliations (88%). Surgery for congenital heart disease in the adult age is a safe, beneficial, and low-risk treatment that modifies patients' natural history by improving their clinical status. PMID- 15283359 TI - Left ventricular outflow tract obstruction. AB - Left ventricular outflow tract obstruction can occur at the supravalvar, valvar, or subvalvar level. Each level of obstruction is associated with distinct symptomatology, natural history, and operative approach. Reconstructive techniques can usually be used with low operative risk and excellent immediate and longer-term outcomes. Valve replacement for valvar obstruction is advised when reconstruction is not possible. The Ross procedure has greatly improved the results of valve replacement in children. PMID- 15283360 TI - Mitral valve replacement with the pulmonary autograft: Ross II procedure with Kabanni modification. AB - We report our experience with pulmonary autograft replacement of the mitral valve in eight patients. Hospital mortality of 0% was reported, and there has been one autograft failure requiring replacement (mean follow-up 12 +/- 6 months). We also review the world's experience with this technique. Mitral valve replacement with a pulmonary autograft (Ross II) is safe, reproducible, and possible warranted for selected patients. PMID- 15283361 TI - Conduit selection for right ventricular outflow tract reconstruction: contemporary options and outcomes. AB - Reconstruction of the right ventricular outflow tract, with the establishment of an unobstructed pathway between the right ventricle and the pulmonary arteries, is a task that the congenital heart surgeon frequently faces. In situations where this outflow tract is congenitally absent, or the pulmonary valve has been used to replace a dysfunctional left ventricular outflow tract, a conduit is usually required to establish pulmonary blood flow. Cryopreserved homografts are currently favored for this, though these nonviable valved allografts have certain limitations. The following review will further define the problem of right ventricular outflow tract reconstruction, with an emphasis on conduit selection and possible alternatives to conduit repair. PMID- 15283362 TI - Neurologic monitoring for special cardiopulmonary bypass techniques. AB - Low flow hypothermic cardiopulmonary bypass, deep hypothermic circulatory arrest, and regional low-flow cerebral perfusion are special techniques used to facilitate complex intracardiac and aortic surgery in neonates and infants. Each carries a risk of cerebral hypoxia and neurologic morbidity. Neurologic monitoring in the form of near-infrared spectroscopy for cerebral oxygenation, transcranial Doppler ultrasound, and the bispectral index electroencephalogram can monitor the brain during these techniques to determine the minimum acceptable bypass flow rates or maximum acceptable duration of deep hypothermic circulatory arrest. The use of this monitoring has the potential to improve long-term neurologic and developmental outcome. PMID- 15283363 TI - Periventricular leukomalacia following neonatal and infant cardiac surgery. AB - The dramatic reduction in surgical mortality following repair of congenital heart defects has been accompanied by increasing recognition of adverse neurodevelopmental sequelae in some children. Neurodevelopmental abnormalities following neonatal and infant cardiac surgery include mild cognitive impairment, expressive speech and language abnormalities, impaired attention and executive function, impaired visual and spatial motor skills, as well as learning disabilities. These defects result in a significant need for early intervention, as well as rehabilitative and special education services. Central nervous system injury following repair of congenital heart defects results from a complex interaction of patient specific and environmental factors. Recent studies suggest that cerebral white matter injury characterized by periventricular leukomalacia (PVL) is common in infants with congenital heart disease particularly following cardiac surgery. Studies at our institution show that this occurs in greater than 50% of neonates following cardiac surgery, but is rare in older infants. Prolonged exposure to cardiopulmonary bypass (with or without deep hypothermic circulatory arrest) is a risk factor for PVL, possibly secondary to the systemic inflammatory response to cardiopulmonary bypass. Hypotension and hypoxemia in the early postoperative period, especially diastolic hypotension, significantly increase the risk of PVL. Future studies are needed to determine significance of PVL as a marker for long-term developmental dysfunction. PMID- 15283364 TI - Pediatric myocardial protection: a cardioplegic strategy is the "solution". AB - This article describes the experimental infrastructure and subsequent successful clinical application of a comprehensive cardioplegic strategy that limits intraoperative injury and improves postoperative outcomes in pediatric patients. The infant heart is at high risk of damage from poor protection as a result of preoperative hypertrophy, cyanosis, and ischemia. These factors may also make the immature (pediatric) heart more sensitive to cardioplegic arrest compared with the mature (adult) heart. The preoperative factors of cyanosis and pressure volume overload are discussed, followed by the infrastructure of the strategies of warm induction and reperfusion with substrate enhancements, multidose cardioplegia, and a "modified" integrated approach to allow ischemia only when visualization is needed in pediatric surgeries. The importance of using a blood cardioplegia solution, with reduced calcium, increased magnesium, and low perfusion pressure are also shown. A practical clinical framework based on these experimentally proven principles is then presented to allow the surgeon to apply these strategies clinically. The results of using these principles are depicted in a series of 567 patients, including 93 patients with hypoplastic left heart syndrome. Applications of these concepts should improve the safety of protection of the infant heart and reduce postoperative morbidity and mortality. PMID- 15283365 TI - Heparin-induced thrombocytopenia (HIT) in pediatric cardiac surgery: an emerging cause of morbidity and mortality. AB - Unfractionated heparin (UFH) is immunogenic, and heparin-dependent antibodies can be demonstrated 5 to 10 days postoperatively in 25% to 50% of adult postcardiac surgery patients. In a minority of these cases (1% to 3% if UFH is continued longer than 1 week) these antibodies strongly activate platelets, causing thrombocytopenia and massive thrombin generation (HIT syndrome). HIT is an intensely procoagulant disorder, and in adult cardiac surgery patients carries both significant thrombotic morbidity (38% to 81%) and mortality (28%). Despite the ubiquitous use of UFH in pediatric intensive care units, and the repeated and sustained exposures to UFH in neonates and young children with congenital heart disease, HIT has been infrequently recognized and reported in this patient population. However, emerging experience at our institution and elsewhere suggests that HIT is significantly under-recognized in pediatric congenital heart disease patients, and may in fact have an incidence and associated thrombotic morbidity and mortality in this patient group comparable to that seen in adult cardiac surgery patients. This article will review HIT in pediatric patients with congenital heart disease and emphasize the special challenges posed in clinical recognition, laboratory diagnosis, and treatment of HIT in this patient group. We will also outline our experience with the off-label use of the direct thrombin inhibitor, argatroban, in pediatric patients with HIT. PMID- 15283366 TI - Practice patterns in neonatal cardiopulmonary bypass. AB - This article reviews practice patterns of numerous congenital heart surgeons, as collected from surveys (the Congenital Heart Surgeon's Society) and from audience response at the American Association of Thoracic Surgery (May 2003) and the Society of Thoracic Surgeons (January 2004). The information shows that there are numerous practice patterns, with unanimity only in the use of cardioplegia, although the delivery, type, and timing of doses varies. Hypothermic circulatory arrest continues to be used by the majority of congenital heart surgeons, although strategies for delivery have evolved in a way that reflects research contributions over the past decade. PMID- 15283367 TI - Risk adjustment for congenital heart surgery: the RACHS-1 method. AB - The new health care environment has increased the need for accurate information about outcomes after pediatric cardiac surgery to facilitate quality improvement efforts both locally and globally. The Risk Adjustment for Congenital Heart Surgery (RACHS-1) method was created to allow a refined understanding of differences in mortality among patients undergoing congenital heart surgery, as would typically be encountered within a pediatric population. RACHS-1 can be used to evaluate differences in mortality among groups of patients within a single dataset, such as variability among institutions. It can also be used to evaluate the performance of a single institution in comparison to other benchmark data, provided that complete model parameters are known. Underlying assumptions about RACHS-1 risk categories, inclusion and exclusion criteria, and appropriate and inappropriate uses are discussed. PMID- 15283368 TI - The Aristotle score for congenital heart surgery. AB - The aim of the Aristotle project was to develop a new method of evaluation of quality of care in congenital heart surgery based on the complexity of the surgical procedures. Involving a panel of expert surgeons, the project started in 1999 and included 50 pediatric surgeons from 23 countries representing International Scientific Societies. The complexity was based on the procedures as defined by the Society of Thoracic Surgeons (STS)/European Association for Cadiothoracic Surgery (EACTS) International Nomenclature and was undertaken in two steps: The first step was establishing the Basic Score, which adjusts only the complexity of the procedures and is based on three factors: the potential for mortality, the potential for morbidity, and the anticipated technical difficulty. The second step was the development of the Comprehensive Score, which further adjusts the complexity according to the specific patient characteristics. The Aristotle score allows precise scoring of the complexity for 145 congenital heart surgery procedures. One interesting concept coming out of this study is that complexity is a constant and precise value for a given patient regardless of the center where he is operated. The Aristotle method allows proposing the following equation of quality of care: Complexity FN Outcome = Performance. The Aristotle score, electronically available, was introduced in the EACTS and STS databases. A validation process, designed to evaluate its predictive value, is being developed. PMID- 15283369 TI - Let the data speak for themselves? AB - No medical discipline has been more shaped, driven, and scrutinized by outcomes data than cardiac surgery. Unlike high-volume operations for acquired heart disease, congenital heart disease is considerably more heterogeneous, many anomalies are rare, and outcomes after surgical correction are highly variable. How, then, can outcome of institutional programs be compared fairly? Growing in popularity among congenital heart surgeons are methods of comparison that rely fundamentally on expert opinion about perceived complexity of treatment. They may be broadly calibrated using administrative or registry outcomes data. This approach, one of two suggested by Aristotle, characterized pre-Newtonian science, in which observed data played a secondary role. This contrasts sharply with the second approach suggested by Aristotle and revived by Newton in the 18th century that places data at its center: "Let the data speak for themselves." The latter is the basis for contemporary methods of risk-adjusted comparisons. The proposed international collection of a uniform set of congenital heart surgery data elements, a well-conceived and internationally accepted ontology of congenital heart disease, accurate understanding of established incremental risk factor concepts and their role in risk adjustment, advent of powerful data analysis techniques that include new types of predictive modeling, and wide understanding of risk-adjusted comparison suggest there is ample motivation and opportunity for letting data speak for themselves. There is no evidence that a data-centric approach, based on Aristotle's and Newton's ideas that liberated 18th century science, has failed and should be abandoned. PMID- 15283370 TI - HDA dissolution need not hold back public health. Small risk-taking team to take agenda forward is what is needed now. PMID- 15283371 TI - Devil-spotting in the detail. Tory plans for PCT's may give succour to those who undervalue their worth. PMID- 15283372 TI - Hang on in there. PMID- 15283373 TI - Worthy Winston's on the ball in health debate. PMID- 15283374 TI - Day-case surgery. PMID- 15283375 TI - All at C. AB - Hepatitis C is officially recognised as a major public health problem, but DoH delays in tackling the problem mean it has not been on PCTs' priority lists. A public awareness campaign due to be launched by the DoH is set to change this by encouraging people to come forward for testing. Services will need to invest in IT for improved surveillance and respond to the patient influx this may create. PMID- 15283376 TI - Assault of the earth. AB - The NHS needs more incentives to take sustainable development to heart. Achievements by trusts are largely due to personal commitment, and are not systematic. A centrally backed programme of sustainable development could save money and would benefit staff, patients and visitors. PMID- 15283377 TI - Funding and crisis resolution teams dominate mental health trust chief executives' concerns. Sins of commission. PMID- 15283378 TI - Beware the hazards when offloading a freehold, say Mark Paget Skelin and David Firth. Tenants' extra. PMID- 15283379 TI - Introduction of a balanced scorecard will enable HR professionals to gauge performance in their organisation more accurately, says Mary-Louise harding. Another brick in the wall. PMID- 15283380 TI - Pancreas and islet transplantation. AB - The aim of pancreas and islet transplantation is to establish the same status of glucose control that is provided by endogenous secretion of insulin from a healthy native pancreas in order to improve the quality of life and ameliorate secondary diabetic complications in patients with type I insulin-dependent diabetes mellitus (IDDM). Islet transplantation is, theoretically, an ideal solution for patients with IDDM since it is not a major procedure, can be performed radiologically and can be repeated several times without any major discomfort to the patient, but despite experimental and clinical efforts over the past 25 years, long term and consistent insulin independence has not yet been achieved. Pancreas transplantation is indicated for patients with IDDM following also additional selection criteria. In a suitable candidate, the evaluation is also needed to determine the type of pancreas transplantation, based mainly on the degree of nephropathy. Details of the recipient operation together with the anti-reject procedures and actual global results are described analytically. Similar considerations are dedicated to the islet transplantation procedure. PMID- 15283381 TI - [Risk factors of surgical wound infection]. AB - Surgical Site Infection (SSI) continues to be a major source of morbidity following operative procedures. The aging of the population means that not only will the number of operations likely increase, but the National Nosocomial Infections Surveillance (NNIS) Risk Index, which standardizes the risk of SSI for an aging population, will be greater. The NNIS report for 1986-1996 described an SSI rate of 2.6% for all operations at the reporting hospitals. It seems likely that overall SSI rates are likely to be greater than reported. All surgical wounds are contaminated by bacteria, but only a minority actually demonstrate clinical infection. The SSI are the biological summation of several factors: the inoculum of bacteria introduced into the wound during the procedure, the unique virulence of contaminants, the microenvironment of each wound, and the integrity of the patients host defense mechanisms. Risk factors were studied in single and multivariate analyses. Although an SSI rate of zero may not be achievable, continued progress in understanding the biology of infection at the surgical site and consistent applications of proven methods of prevention will allow us to further reduce the frequency, cost, and morbidity associated with SSI. PMID- 15283382 TI - Treatment of "locally advanced" well-differentiated thyroid carcinomas. AB - PURPOSE: To analyse thyroid carcinomas having an extrathyroid extension in order to identify the principal prognostic factors and outline an effective therapeutic strategy. METHODS: We selected a sample of 160 patients suffering from locally advanced "well differentiated thyroid carcinoma (T4) who had undergone surgery at the Department of Surgery of University of Rome "La Sapienza". The sample was subdivided into three groups: T4, limited type I, and extensive type II, T4 microcarcinomas. RESULTS: We obtained excellent results with the T4 microcarcinomas, above all in patients under the age of 45, with a 94.5% survival rate, compared with 88% in patients aged over 45. In the extensive type II T4 carcinoma we obtained a survival rate of 29.4% in patients aged over 45 years. CONCLUSIONS: Age, combined with an aggressive histological variant (Sclerosing and tall-cell papillary carcinoma), is an important factor in prognosis. The radicality of surgical excision is considered an important prognostic factor, although the results reported in the literature are contradictory. Aggressive surgery can free from the disease a high percentage of patients over the age of 50 even with T4. We deem it fundamental to perform total thyroidectomy in all advanced cases of thyroid neoplasm and to extend neoplasm excision to the adjacent tissues, even involving justified surgical demolition. PMID- 15283383 TI - [Hypoparathyroidism following total thyroidectomy: prognostic value of intraoperative parathyroid hormone assay]. AB - Transient and definitive hypoparathyroidism represent a frequent complication after thyroid surgery. Recently some authors proposed the use of intraoperative parathyroid hormone assay for the rapid detection of this complication. In this paper the authors describe the data obtained from 42 total thyroidectomies with intraoperative measurements of parathyroid hormone. When parathormone decrement was over 75% during thyroidectomy, the hypocalcemic symptomatology was found in all cases during postoperative observation. The authors emphasize intraoperative PTH dosage for immediate identification of patients at risk for postoperative hypoparathyroidism. In this cases parathyroid autotransplantation is suggested to prevent postoperative hypoparathyroidism. PMID- 15283384 TI - [Endometriosis of the abdominal wall (authors' experience)]. AB - Endometriosis is a very common gynecological disease, although the abdominal wall localization is a rare clinical problem in the everyday practice and this explains the incomplete reports in literature and the difficulty of a standard treatment. The authors report four cases of abdominal wall endometriosis, underlining how the surgical therapy represents the golden standard of decisive treatment, thanks also to prosthetic reconstruction techniques who allow more radical demolitions. On the contrary, the medical treatment should be reserved to selected cases such as especially the less symptomatic umbilical primitive diseases. PMID- 15283385 TI - [Urgent management of obstructing colo-rectal cancer: authors' experience]. AB - PURPOSE: The aim of this retrospective study is to compare the different surgical approaches in obstructing colo-rectal cancer in terms of mortality, morbidity and quality of life. MATERIALS AND METHODS: We observed 379 patients with colorectal cancer, 354 of which underwent surgical treatment, 189 M (53.4%) and 165 F (46.6%), with a median age of 72.6 years. Complicated tumors were 150 (42.4%), with 126 obstructions (84%). For 95 obstructing left-sided colorectal cancers we performed: 9 defunctioning colostomies; 62 two-stages operations: 55 Hartmann's procedures, 5 primary anastomosis with colostomy; 2 primary anastomosis with on table wash-out and ileostomy; 24 single-stage operations: 17 primary anastomosis with on table wash-out and 7 colectomy. RESULTS: The overall operative mortality rate was 8.7% (11/126). The overall leak rate was 8% (5/62), 12.9% (4/31) in left colon and 3.2% (1/31) in right colon, all treated conservatively. The wound infection rate was 23.8% (30/126). DISCUSSION AND CONCLUSIONS: Obstructing colo rectal cancer is associated with a high operative mortality and a worse prognosis. Defunctioning colostomy can be regarded as a valid option only in extreme circumstances. Hartmann's operation has indicated in case of metastatic disease, unsure anastomosis, simultaneous colonic perforation. The gold-standard is primary anastomosis, as colonic resection with on table wash-out or subtotal/total colectomy, in case of largely distended colon or synchronous lesions. PMID- 15283386 TI - [Prolassomucosectomy according to Longo. Intermediate results]. AB - We present results obtained in our first series of 25 patient treated by prolassomucosectomy for haemorrhoidal prolapse with a follow up of 18-36 months. Control of the disease and functional results proved to be optimal. After small early haemorrhages from the suture line, we started adding stitch sutures with haemostatic intent to all three vascular pedicles. Early or late additional complication were not observed. PMID- 15283387 TI - [Minimally invasive video-assisted thyroidectomy--techniques and results over 4 years of experience (1999-2002)]. AB - INTRODUCTION: This study reviews four years of Minimally Invasive Video Assisted Thyroidectomy (MIVAT) technique and compares the results to those of traditional thyroid surgery. MATERIALS AND METHODS: Between 1999 and 2002, a series of 427 patients were submitted to MIVAT at our Department. Selection criteria were: thyroid nodule maximum diameter of 3.5 cm, total thyroid volume under 25 cc, no signs associated thyroiditis, diagnosis of benign thyroid disease or "low risk" thyroid tumor, no evidence of nodal disease of the neck. RESULTS: We operated on 362 females and 65 males and the mean age of the population was 39.6 years (range 10-77). A total thyroidectomy was performed in 208 cases, and 219 patients underwent a single-side procedure. Mean operative time was 30.4 minutes for lobectomy (range 20-140 minutes) and 50.2 for total thyroidectomy (range 35-140). Complications were represented by definitive recurrent nerve palsy in 3 patients (0.7%) and one case of definitive hypoparathyroidism (0.4%). A wound infection is reported in 3 cases and we had no major bleeding that required surgical revision. A conversion to open procedure was performed in 5 cases (1.2%); mean hospitalisation was 1.28 days (range: 1-4). CONCLUSIONS: This series demonstrates that MIVAT is not different to conventional open surgery in terms of complications, radicality of the procedure and operative time. Moreover, even if not statistically proved, MIVAT appears to offer some advantages in terms of cosmetic results and postoperative pain. In conclusion, we believe that MIVAT is a perfectly reproducible and safe technique for both benign and low-risk malignant thyroid disease, when correct indications are strictly followed. PMID- 15283388 TI - [Radiofrequency-assisted hepatic resection--first experience]. AB - AIMS: Intra-operative blood loss during liver, resection remains a major concern due to its association with higher postoperative complications and shorter long term survival. The aim of this study was to assess the feasibility and safety of a novel concept for liver resection using a Radio-Frequency energy assisted technique. METHODS: From January 2001 to January 2002, 27 patients were operated on using Radio-Frequency energy assisted liver resection. Radio-Frequency energy was applied along the resection edge to create a 'zone of desiccation' prior to resection with a scalpel. RESULTS: Median resection time was 47.5 minutes (range 30-110). The median blood loss during resection was 30 mls (range 15-992) and mean pre-operative and post-operative haemaglobin values were 13.5 g/dL (SD+1.7) and 11.6 g/dL (SD+1.4) respectively. No blood transfusion was registered, nor was any mortality observed. There was one post-operative complication: a sub-phrenic abscess. Median post-operative stay was 8 days (range 5-86). CONCLUSION: Liver resection assisted by RF energy is feasible, easy and safe. This novel technique offers a new method for 'transfusion-free' resection without the need for sutures, ties, staples, tissue glue or admission to Intensive Care Unit. PMID- 15283389 TI - [Experimental study of the effect of the Argon Beam Coagulator on organic tissues from the viewpoint of surgical utilization]. AB - The Argon Beam Coagulator has gained his space in surgery thanks to its operative characteristics, that are very useful in sealing the bleeding parenchymal tissue with minimal injury to the surroundings. The aim of the present study is that of evaluate the physical properties of the instrument in its coagulation action. The experimental study with the Birtcher 6000 Argon Beam Coagulator has been designed to measure the top temperature that develops right where the Argon beam meets the tissue, while operating. Using a laser guided telethermometer, the searching of that temperature was uneasy right on the operatory field because of the unwilling movements of the operator hand and those of the patient himself. Therefore a similar protocol was made on a piece of meat coming from the butcher. At the longest application of the beam coagulation on the same point the developed temperature was never higher than 100 degrees C whilst a complete coagulation effects. The advantage of the Argon Beam coagulation are therefore to seal the diffuse bleeding without injury to this adjacent tissues, never exposed to a temperature higher than that of boiling water. The advantages are also evident in comparison with the more usual spray electrosurgery that is instead characterized by a wide carbonisation also with this spray option. PMID- 15283390 TI - [Regression of primary low-grade gastric mucosa-associated lymphoma by eradication of Helicobacter pylori infection: case report]. AB - OBJECTIVE: The Authors report their experience in diagnosis and treatment of one case of primary low-grade gastric lymphoma of mucosa associated lymphoid tissue (MALT); recent international literature review. EXPERIMENTAL DESIGN: Complete clinical report. Diagnostic, clinical and prognostic indication, evaluation of effectiveness of eradication therapy and short follow-up. SETTING: Operative Unit of General and Thoracic Surgery. University "Paolo Giaccone" of Palermo. INTERVENTION: Treatment of H. Pylori infection (lansoprazole, amoxicillin and metronidazole twice a day for 14 days; after that, lansoprazole for another 4 weeks), according to international guide-lines. RESULTS: H. Pylori was completed eradicated. Disappearance and total regression of the lymphomatous tissue was observed. No relapse were recorded at short follow-up. CONCLUSIONS: Our reports confirm the recent anecdotal reports on regression of gastric MALT lymphoma after eradication of H. Pylori and indicates that the growth of these extranodal lymphomas may depend on H. Pylori. PMID- 15283392 TI - Intussusception of the small bowel due to Peutz-Jeghers syndrome: a case report. AB - The Peutz-Jeghers syndrome is an autosomal dominant disorder characterized by hamartomatous polyposis of the gastrointestinal tract, melanin pigmentation of the skin and mucous membranes, and an increased risk for cancer. The incidence of surgical complications in these patients is relatively rare, and correlates with the size and location of the polyps. Herein we report the case of a 27-year-old woman presented with episodes of abdominal pain, abdominal distention and intermittent vomiting. Moreover, multiple pigmentation of the mouth was also noted. A preoperative diagnosis of a double jejunal intussusception and jejunal occlusion was based on the findings of small bowel enema and computed tomography. The diagnosis was confirmed at laparotomy. PMID- 15283391 TI - [Gastrointestinal stromal tumor (GIST): a rare case of severe gastric haemorrhage]. AB - INTRODUCTION: A rare case of gastrointestinal stromal tumor (GIST) provoking a severe gastric bleeding is reported. Case report and results. The Authors report on the case of a 53-year-old male patient who presented with hematemesis, melena and severe acute anemia (Hb: 6 g/dl). EGDS disclosed a protruding lesion centrally ulcerated, of 7 cm in size, localized in the upper half of gastric corpus. Multiple endoscopic biopsies were negative for neoplastic changes. Because of further gastric bleeding, a total gastrectomy with "Roux-en-Y" reconstruction was performed in urgency GIST of smooth muscle was diagnosed by histological and immunohistochemical postoperative examination. DISCUSSION: Gist are neoplasms arising from connective tissue elements of gastrointestinal wall, which represent about 2% of GI-tract malignant tumor. Tumor size of 5 cm or greater, elevated mitotic count, lack of histological differentiation are significantly associated with a shorter recurrence-free survival. GIST-s are rarely cause of an inarrestable gastric bleeding. For gastroenterological surgeons it is critical to select the most suitable surgical procedure. In our case, 9 months after gastrectomy the patient is well, in spite the severe clinical background and the malignancy degree. CONCLUSION: The GIST-s have to be taken into account in the differential diagnosis of GI-tract tumors if endoscopic biopsies are negative for malignancy. In our opinion, total or partial gastrectomy fro gastric should be preferred. Moreover, a close follow up is recommended. PMID- 15283393 TI - [Mesenteric cysts: a case report]. AB - Mesenteric cysts are lesions of rare chech, they can manifest or their painful abdominal pathology or comprehensive by effect mass. Today with imaging methods, especially the U.S., diffuse with capillarity in the hospitals, it possible to point at them also by chance for others types of problems. When possible, the FNAB, is diriment by diagnostic point of view. New imaging methods give an aimed operative planning, that spares to the patient a blindly operation on an unprepared intestine. PMID- 15283394 TI - Large extrapleural hematoma in an anticoagulated patient after a thoracic blunt trauma. AB - We report the 7th case of a traumatic extrapleural hematoma that developed in an anticoagulated patient with a thoracic blunt trauma and rib fractures, and required an emergency surgical treatment. Extrapleural hematoma is a rare and life-threatening condition characterized by a collection of blood between the pleura parietalis and the endothoracic fascia. Related symptoms and chest x-ray findings are not characteristic and may present several hours after the injury, leading to delayed diagnosis and treatment. Etiological, surgical and prognostic implications of this finding are briefly discussed. PMID- 15283395 TI - Abrikossoff's tumour: report of a rare case in anal perianal region. AB - The authors report a rare case of a 47-year old man suffered from Crohn's disease and depression with multiple prominent lesions in the anal and perianal region. The biopsy of these lesions showed the presence of Abrikossoff's tumour. This tumour is very rare in the anal region and usually the lesions are small (0.5-3 cm) and solitary. The authors report this case because they considered it an interesting case for the localization and the appearance of the lesions. PMID- 15283396 TI - Cutaneous melanoma with neurofibromatosis type 1: rare association? A case report and review of the literature. AB - Neurofibromatosis (NF) is a relatively common disorder characterized by cutaneous pigmented maculas, multiple neurofibromas and Lisch nodules (pigmented iris hamartomas). This disorder is retained being a neurocristopathy. Melanocytes are neural crest derivates too. Twenty-six patients with neurofibromatosis associated to cutaneous malignant melanoma have been reported till now, but data on association between these two pathologies are lacking. One more case of malignant cutaneous melanoma in a patient with neurofibromatosis is reported and the hypothesis of a more frequent association than usually believed of these two pathologies is discussed. PMID- 15283397 TI - [Fournier's gangrene: case report and review of recent literature]. AB - OBJECTIVE: The authors report their experience in diagnosis and treatment of one case of Fournier's gangrene; recent international literature review. EXPERIMENTAL DESIGN: Complete clinical report. Diagnostic, clinical and prognostic indication, evaluation of effectiveness of surgical treatment (debridement and necrosectomy) and follow-up; comparison between indications and multidisciplinary approach proposed by international literature. SETTING: Operative Unit of General and Thoracic Surgery. University "Paolo Giaccone" of Palermo. INTERVENTION: Repeated surgical treatment previous multimodal approach, according to international guide lines. RESULTS: Complete recovery with "restitutio ad integrum". No relapse were recorded at follow up. CONCLUSIONS: Fournier's gangrene is an uncommon and aggressive synergistic fasciitis of the perineum and genital organs, which may bring the patient to death; it is a true surgical emergency. The disease can no longer be considered to be idiopathic; in most cases a urologic, colorectal or cutaneous source can be identified. Despite antibiotics and aggressive debridement, the mortality rate remains high, particularly in the elderly, in patients with renal failure, and in patients with extensive disease. The presentation is highly variable, necessitating a high index of suspicion. High risk patients include diabetics, alcoholics and debilitated and immunosuppressed individuals. As the AIDS population increases, the incidence of Fournier's gangrene may increase as well. In questionable cases, imaging modalities should be performed to allow early diagnosis and to reduce misses diagnosis. Broad spectrum antibiotics (while waiting for the results of culture and antibiogram effectuated on tissue specimens obtained during necrosectomy) and aggressive debridement remain the hallmarks of treatment. Hyperbaric oxygen therapy and improved local wound care may decrease the extent of tissue destruction. The surgical operation has to be performed in emergency to avoid a rapid spread of tissue necrosis and a possible development towards septic shock. Reconstructive techniques afford better cosmetic results. With early recognition, prompt treatment, improved wound care and reconstructive efforts, the mortality rates and cosmetic results should continue to improve. PMID- 15283398 TI - [Memory of Professor Orazio Campione]. PMID- 15283399 TI - [State-of-the-art in the diagnosis and therapy of the MEN1 and MEN2 syndromes]. PMID- 15283400 TI - [Functional outcomes and quality of life in patients with anterior resection for rectal cancer. Does preoperative manometry predict the feasibility of a J pouch?]. AB - INTRODUCTION: The goal in the treatment of rectal cancer is the recovery of the disease with the best fecal continence and quality of life. The Authors compared quality of life and manometric results in patients treated with neo-adjuvant chemotherapy and rectal low anterior resection (LRA). METHODS: From January 1998 to March 2002 50 patients with advanced (T3-T4) rectal cancer underwent neo adjuvant chemotherapy. Subsequently 41 of them underwent LRA with colon pouch (19) or without the pouch (22). After few months the quality of life was evaluated through a questionnaire. Later they underwent manometric evaluation measuring resting, squeeze and rectal compliance. RESULTS: The manometric results and the questionnaire scores agreed in 75% of patients. In detail, patients with hypotonic sphincter had a better (one could say good) quality of life if a LAR with pouch had been performed respect to the patients without pouch. CONCLUSION: Performing LAR with colon pouch after neoadjuvant chemotherapy in patients with hypotonic sphincter improves quality of life. Preoperative anorectal manometry could select patient who would benefit from pouch construction. PMID- 15283401 TI - [Laparoscopic colectomy in the treatment of colon cancer: prospective study]. AB - The Authors report their experience on laparoscopic colectomy in 38 patients treated between June 2001-September 2003 in General Surgery and Organ Transplantation Department of University of Parma, Italy. The patients were 23 male and 15 female, with average age 58.4 years. All patients were studied with TC and colonoscopy performed by the surgeon. The conversion rate was 15.8% and the average hospital stay 6.9 days (range 6-15 days). The patient's general clinical conditions and the results showed that the laparoscopic colectomy is a safe surgical option. PMID- 15283402 TI - [Synchronous colorectal and renal tumors: two case reports]. AB - The simultaneous presence of more than one primary malignant tumour in the same patient is a relatively rare event, with values going from 2.7% to 6.8%. In this paper two synchronous colorectal-renal cases surgically treated are described; in the first case with only one resolutive operation for the two tumours, in the second with two consecutive operations. In both cases, the patients had no familiarity for colorectal or renal malignant tumours. Both cases have paradigmatic peculiarities: in the first the discovery of the renal neoplasm has been possible in election during the normal execution of the diagnostic protocol for sigmoido-rectal neoplasm. The second case shows the other possible modality of the outset of the two tumours: the occlusive urgency which never allows to discover immediately the renal tumour (unless a laparotomic exploration makes it perceptible). PMID- 15283403 TI - [Intraoperative injuries of the superior laryngeal nerve in thyroid surgery]. AB - The intraoperative injury of the external branch of the Superior Laryngeal Nerve (SLN) is a disregarded complication of thyroid surgery that becomes a serious one for particular professional groups. The Authors retrospectively evaluate 124 cases of total thyroidectomies performed by the same surgeon from 1999 to 2002. Clinically evaluated SLN lesions (hypomobility of the vocal cord) was registered in six patients (4.8%). Routine identification of SLN and meticulous dissection of the superior thyroid vessels must be the technical benchmark to avoid nerve injury, even if bulky disease, anatomic variations and previous surgery may increase the chance of nerve damaging. PMID- 15283404 TI - Nipple leiomyoma in man: a case report. AB - We describe a rare case of a man, 38 year old, with a nipple leiomyoma, and report the presentation as a small nodule of the areola spreading the nipple, the symptoms, the clinical signs, the treatment that includes a complete excision; free margins should be histologically established to prevent recurrence. PMID- 15283405 TI - [Spigelian hernia. Diagnosis and surgical treatment]. AB - Spigelian hernia (SH) is a rare partial abdominal wall defect; its manifestation is rare. Seven cases were observed--4 females and 3 males with mean age of 56.5 years (range 38-65)--in 857 patients operated for hernia (0.8%) between 1995 and 2003. Ultrasound examination avoid the diagnosis and marked the fascial defect, measuring diameter and sac contents. In all cases a surgical approach with an epicritic incision has be done and the fascia defect closed with properitoneal and subfascial polypropylene mesh (Prolene Hernia System, PHS). All patients underwent to follow-up demonstrating no recurrences or complications like mesh suppuration or dislocation. PMID- 15283406 TI - [Combined video-thoracoscopic, surgical and endovascular treatment in a case of ascites and recurrent bilateral pleural effusion]. AB - Among recurrent pleural effusions a role of remarkable importance is held by those combined with ascitis due to the difficulty of their treatment, even using widely tested techniques. The incidence of such pathology varies from 4% to 6% of patients suffering from cirrhotic pathology, reaching 10% in cases with advanced illnesses. Pleural effusions involve the right emithorax more frequently than the left one, but it can show up bilaterally too. Its etiopathogenesis is tied up to the direct passage of ascitic liquid into the chest and, during the past years, numerous theories have been described to explain this migration. The Authors report the case of a patient with interesting considerations for the diagnostic difficulties and the peculiarity of the treatment performed. PMID- 15283407 TI - [Use of prosthetic materials in incisional hernias: our clinical experience]. AB - Incisional hernias are fairly frequent complications of surgery. Such complications are presented by variable percentages ranging from 1% to 8% for non complicated laparotomies and from 10% to 15% in case of infection. The therapeutic possibilities are abdomino-plastic with direct suture and the use of prosthesis. The percentage of recurrency varies from 30% to 50% and from 0% to 19% respectively. From May 2000 to September 2003, the Authors operated 64 patients. In 30 cases they opted for a direct suture approach, in 34 cases a prosthesis was used. Of the 34 patients, 18 were treated using polypropylene mesh placed below the rectus muscles and above the peritoneum (Rives technique); 10 with PTFEe (Polytetrafluoroethylene expanded) prosthesis placed in intraperitoneal site and 6 were treated with SIS prosthesis (Small Intestine Submucosa), 3 placed using the Rives technique and 3 in intraperitoneal site. Nine patients had associated operations. The use of prosthesis in the management of incisional hernias has brought a considerable reduction in the percentages of recurrency, especially in cases of associated pathologies. The availability of SIS in the market has permitted the use of this prosthesis even in case of contaminated and/or infected fields. In a follow-up ranging from 3 years to 3 months (mean 18 months) the Authors observed 2 cases of recurrence in the group of direct suture. Despite the advantages offered by the use of prosthesis, we can affirm that there is no "universal" prosthesis. Their selection should be evaluated on the bases of the site and the dimensions of the incisional hernia, the possibility of infection, the surgical technique and the site of prosthetic placement in the abdominal wall. PMID- 15283408 TI - [Learning curve and early complications of totally implantable venous access devices: is tutoring the solution for the problem?]. AB - During the last years operators implanting totally implantable venous access devices (TIVADs), type of access, and kind of complications are changed. Aim of this work is to evaluate the incidence of early complications during the learning curve of residents in surgery or inexperienced surgeons, besides considering the tutoring to evaluate its rule to prevent early complications. TIVADs, implanted by residents in surgery or by inexperienced surgeons in the Department of Surgical Science Organ Transplantation and Advanced Technologies of University of Catania from January 1995 to October 2003, have been considered for the present study. Age and sex of the patients, indication, type of surgical access and early complications of the TIVADs have been considered. Early complications are those complications that occur within 30 day after the implant. Ninety-five TIVADs were implanted by surgical approach in 95 patients: 58 males (61%) and 37 females (39%), with a mean age of 55 years (range 31-79). Inexperienced surgeons performed 40 implants (42%) instead resident in surgery implanted 55 TIVADs (58%). The migration out of the vein of a catheter 20 days after the operation was the only complication recorded. TIVADs implant by cut-down technique represent the way to avoid early complications. The learning curve is short and the tutoring is limited at the first phase. PMID- 15283409 TI - Studies of the transmission routes of Ornithobacterium rhinotracheale and immunoprophylaxis to prevent infection in young meat turkeys. AB - The importance and prevention of the horizontal as well as the vertical transmission of Ornithobacterium rhinotracheale were investigated. In our first experiment we observed that specific-pathogen-free broiler chickens that were placed in hatching incubators at a commercial turkey hatchery during hatch showed respiratory tract lesions at postmortem examination that were positive for O. rhinotracheale by bacteriology and immunohistology. It appeared that vertical transmission occurred and that horizontal transmission of O. rhinotracheale is possible. In a second experiment, the turkeys derived from vaccinated parents showed significantly fewer respiratory tract lesions at postmortem examination at 16 days of age than the birds derived from nonvaccinated parents. In a third experiment, all vaccinated young birds, regardless of the vaccination state of their parents, showed significantly fewer respiratory tract lesions at 6 wk of age. We concluded that vaccination of the breeders reduces vertical transmission and that vaccination of the progeny is needed to resist challenge at 6 wk of age. PMID- 15283410 TI - Effect of BioSentry 904 and ethylenediaminetetraacetic acid-tris disinfecting during incubation of chicken eggs on microbial levels and productivity of poultry. AB - Proper sanitation practices and the use of efficacious disinfectants in a hatchery have an effect on chick quality. Aerosol bacterial counts, egg moisture loss, hatchability, chick quality, and broiler productivity were evaluated when egg surfaces were contaminated by immersion of each egg into a broth medium containing a field isolate of Pseudomonas aeruginosa and incubated with exposure to one of three disinfectant treatments administered by fine spray: distilled water, BioSentry 904 (904), and a 1:1 ratio of 904 and ethylenediaminetetraacetic acid (EDTA)-Tris. The aerosol bacteria levels were statistically greater on day 21 of incubation in the group treated with distilled water than in those receiving disinfectants. Overall hatch of fertile eggs and egg moisture loss were comparable among all three treatments. At 1 day of age, the chicks incubated with 904 had a statistically lower yolk sac contamination rate than those incubated with 904+EDTA-Tris or distilled water. The 2-wk mortality rates, body weights, feed conversion ratios, yolk sac weights, and yolk sac contamination rates were all similar among the three treatments. PMID- 15283411 TI - Nitric oxide production by macrophages stimulated with Coccidia sporozoites, lipopolysaccharide, or interferon-gamma, and its dynamic changes in SC and TK strains of chickens infected with Eimeria tenella. AB - Nitric oxide (NO) is an important mediator of innate and acquired immunities. In the studies reported here, we quantified NO produced in vitro by chicken leukocytes and macrophages and in vivo during the course of experimental infection with Eimeria, the causative agent of avian coccidiosis, and identified macrophages as the primary source of inducible NO. Eimeria tenella-infected chickens produced higher levels of NO compared with noninfected controls. In Eimeria-infected animals, SC chickens produced greater amounts of NO compared with infected TK chickens, particularly in the intestinal cecum, the region of the intestine infected by E. tenella. Macrophages that were isolated from normal spleen were a major source of NO induced by interferon (IFN)-gamma, lipopolysaccharide (LPS), and E. tenella sporozoites. Macrophage cell line MQ NCSU produced high levels of NO in response to Escherichia coli or Salmonella typhi LPS, whereas the HD-11 macrophage cell line was more responsive to IFN gamma. These findings are discussed in the context of the genetic differences in SC and TK chickens that may contribute to their divergent disease phenotypes. PMID- 15283412 TI - Relationship of messenger RNA reverse transcriptase-polymerase chain reaction signal to Campylobacter spp. viability. AB - SUMMARY. Discriminating viable from dead cells is of importance in the development of bacterial detection methods. A positive reverse transcriptase polymerase chain reaction (RT-PCR) amplification signal was tested as a potential predictor of chick colonization. Some researchers have suggested that the presence of messenger RNA (mRNA) may not correlate with cell viability. Chicken colonization by cells that have positive mRNA signal but that are noncultivable would provide a correlation in cell viability and persistence of mRNA. The role of a viable but noncultivable (VBNC) form of Campylobacter spp. for colonization of poultry could be verified by such an mRNA signal. The levels of four strains of Campylobacter spp., previously isolated from poultry feces, declined progressively over time, and loss of cultivability occurred after 6 to 7 wk incubation in phosphate-buffered saline (PBS) at 4 C. Cold-stored, noncultivable and heat-inactivated (60 C for 10 min) Campylobacter spp. produced inconsistent amplified products from RT-PCR assay, depending on the target transcripts and strains used, although all fresh cultures showed mRNA signals. For the most part, signals of mRNA species from VBNC and heat-killed Campylobacter spp. AH-1, AH-2, and CH-3 persisted. RT-PCR amplification of transcripts originating from the tkt and cmp genes and a 256-base pair amplicon (from a previously described putative haem-copper oxidase) provided consistent signals, whereas transcripts from the flaA gene did not. Presumed VBNC and heat-inactivated Campylobacter spp., which produced positive mRNA signal but was not cultivable by conventional culture based methods, did not establish colonization in the intestine of chicks 7 days after challenge. These results lead us to question the correlation between mRNA durability with cell viability as well as the significance of the VBNC cells in environmental transmission of Campylobacter spp. PMID- 15283413 TI - Evaluation of the infectivity, length of infection, and immune response of a low pathogenicity H7N2 avian influenza virus in specific-pathogen-free chickens. AB - The H7N2 subtype of avian influenza virus (AIV) field isolate (H7N2/chicken/PA/3779-2/97), which caused the 1997-98 AIV outbreak in Pennsylvania, was evaluated for its infectivity, length of infection, and immune response in specific-pathogen-free (SPF) chickens. The composite findings of three clinical trials with various concentrations of virus indicated that this H7N2 subtype contained minimal pathogenicity for chickens. The concentration of the virus in the inoculum proved critical in the establishment of a productive infection in a chicken. Seven-day-old SPF chickens were not infected when inoculated with 10(0.7-2.0) mean embryo lethal dose (ELD50) of the H7N2 virus per bird. At this dose level, the immune response to this virus was not detected by the hemagglutination-inhibition (HI) test. Nonetheless, chickens at ages of 5 and 23 wk old tested were successfully infected when exposed to 10(4.7-5.7) ELD50 of H7N2 infectious doses per bird by various routes of administration and also by direct contact. Infected birds started shedding virus as early as 2 days postinoculation, and the period of virus shedding occurred mostly within 1 or 2 wk postinoculation (WPI). This H7N2 subtype of AIV induced a measurable immune response in all birds within 2 wk after virus exposure. Antibody titers were associated with AIV infectious doses and age of exposure of birds. Challenge of these infected birds with the same H7N2 virus at 5 and 10 WPI indicated the infective virus was recoverable from cloacal swabs at 3 days postchallenge and disappeared thereafter. In these challenged birds, the antibody levels as measured by the HI test spiked within 1-2 wk. PMID- 15283414 TI - The influence of reovirus sigma C protein diversity on vaccination efficiency. AB - Avian reovirus (ARV) is a disease agent that causes economic losses in the poultry industry. The available vaccines do not confer full protection. One possible reason is the existence in the field of many virulent serotypes with no cross protection. Several ARV strains have been isolated in Israel in the last few years. In this study, we investigated the diversity of the sigma C protein of ARV because this is the most variable protein in the virus and it induces the production of neutralizing antibodies. Sigma C from two virulent isolates was sequenced, cloned, and expressed. The protein sequence differed from the attenuated vaccine strain (strain 1133) but was similar to a U.S. virulent strain (strain 1733). Those differences led to a change in the antigenic index of the protein, mainly at three sites. Sera of infected birds in a field trial and of birds in a controlled experiment vaccinated with the recombinant sigma C protein showed high titers in enzyme-linked immunosorbent assay to the recombinant protein and lower titers to the attenuated vaccine strain. This means that sigma C can be used as a diagnostic tool for the detection of antibodies relevant for protection and in the future as a subunit vaccine. The results of this study highlight the need to reconsider vaccinations against ARV in terms of the strains to be used and of the method of identifying protective antibodies transferred to progeny. PMID- 15283415 TI - Salmonella enterica serovar typhimurium colonization of the crop in the domestic turkey: influence of probiotic and prebiotic treatment (Lactobacillus acidophilus and lactose). AB - Acute colonization of the crop of the domestic turkey by Salmonella enterica serovar typhimurium (ST) was examined. The influences of preharvest probiotic and prebiotic treatment with lactobaccilli and lactose on crop colonization with ST were also investigated. Prior to Salmonella challenge, poults received 2.5% lactose and Lactobacillus acidophilus (1.9 x 10(9) organisms/liter) in the only source of drinking water from 1 day old to termination. At 3-wk-old, turkey poults were challenged with ST (1.7 X 10(8) colony-forming units [CFU]/ml) before their natural nocturnal fast to determine the potential effects of supplementation on crop colonization when the crop was engorged and subsequently undergoing emptying. Crop ingesta and tissue were collected at time points 30 min and 4, 8, and 24 hr postchallenge and ST levels were determined. High levels of ST were detected in the crop. For instance, for the poults not receiving lactose or lactobacilli, 30 min after ST challenge, there were 4.4 x 10(7) CFU in the crop ingesta and 5.3 x 10(5) CFU in the crop wall. Ingesta ST levels dropped dramatically to 1.0 x 10(6) CFU after 4 hr as the crop emptied. Crop wall ST levels were steady during the nocturnal crop evacuation. Immunohistochemical staining demonstrated ST in close association with the crop epithelium. Treatment with lactose and L. acidophilus supplementation did not reduce ST colonization. PMID- 15283416 TI - Construction, characterization, and evaluation of the vaccine potential of three genetically defined mutants of avian pathogenic Escherichia coli. AB - The delta galE, delta purA, and delta aroA derivatives of avian septicemic Escherichia coli EC99 strain (O78 serogroup) were constructed with a suicide vector containing the pir-dependent R6K replicon and the sacB gene of Bacillus subtilis. The resultant isogenic mutants were stable and lacked approximately 45%, 36%, and 52% of the genes for galE, purA, and aroA, respectively. The delta purA and delta aroA mutants did not grow on minimal medium, whereas the delta galE mutant grew on minimal medium but was sensitive to galactose-induced lysis. The reversion frequencies of all three mutants were <10(-12). The mutants were highly attenuated for virulence as determined by administration of approximately 10(7) colony-forming units of each mutant to 1-day-old chicks by the subcutaneous route. Chickens were vaccinated with the mutants by spray (droplet size approximately 20 microm) at 1 and 14 days of age to determine safety, immunogenicity, and efficacy. The mutants were found to be safe. Seven days after a second vaccination, immunoglobulin (Ig)Y antibodies to E. coli in serum and air sac washings were detected by enzyme-linked immunosorbent assay. In both serum and air sac washings, IgY antibodies were significantly higher in chickens vaccinated with the mutants as compared with phosphate-buffered saline-treated controls but were significantly lower compared with chickens that were vaccinated with the parent strain. In serum, but not in air sac washings, IgY antibodies were significantly lower in chickens vaccinated with the mutants compared with the parent strain. The vaccinated chickens were given infectious bronchitis virus intranasally at 17 days of age and were challenged with homologous (EC99 strain) or heterologous (O2 serogroup) E. coli 4 days later. Chickens that received wild type EC99 strain or its mutant derivatives were protected from homologous but not from heterologous challenge. This study indicates that the delta galE, delta purA, and delta aroA mutants are safe and moderately immunogenic but the protection conferred by the mutants is serogroup specific. PMID- 15283417 TI - Proventriculitis in broiler chickens: effects of immunosuppression. AB - Proventriculitis in broilers causes carcass condemnation when swollen proventriculi tear during evisceration. The cause of this proventriculitis is unknown, but several infectious agents have been associated with it. One such agent, infectious bursal disease virus (IBDV), has been implicated as a cause of proventriculitis, but a direct effect of this virus on the proventriculus has not been proven. The role of IBDV in proventriculitis may be indirect as a result of its ability to cause immunosuppression. The objective of this study was to understand how immunosuppression affects the incidence of proventriculitis in broiler chickens. Immunosuppression was induced in commercial and specific pathogen-free broiler chickens using chemicals (cyclophosphamide and cyclosporin) or virus (IBDV). All groups were then exposed to a proventricular homogenate produced from diseased birds. At 7 and 14 days postinoculation, the incidence of proventriculitis in these groups was compared to that produced by homogenate exposure in immunocompetent broilers. All birds exposed to the proventricular homogenate from diseased birds developed proventriculitis. Cyclophosphamide and IBDV, both B cell suppressors, did not significantly affect the incidence or characteristics of the proventriculitis observed, although they did have an effect on the size of the proventriculus at 7 days postinoculation. Chickens immunosuppressed with cyclosporin, a T cell suppressor, developed more severe lesions and had a higher incidence of proventriculitis. These findings indicate that both B and T cells are involved in the immune response against proventriculitis, but cell-mediated immunity appears to have a more important role in controlling the disease. IBDV affects both humoral and cellular immunity in the chicken, so although under experimental conditions it didn't have a major effect on proventriculitis, it may explain why control of IBDV in the field seems to reduce the incidence of proventriculitis. PMID- 15283418 TI - Inhibition of resistance plasmid transfer in Escherichia coli by ionophores, chlortetracycline, bacitracin, and ionophore/antimicrobial combinations. AB - Medicinal feed additives bacitracin, chlortetracycline (CTC), laidlomycin, lasalocid, and salinomycin inhibited the transfer of multiresistance-conferring plasmid pBR325 (Tet(r) Amp(r) Cp(r), 6.0 kb) into selected gram-negative strains with the use of an in vitro model. High concentrations of ampicillin-sensitive competence-pretreated Escherichia coli HB 101 cells were exposed to 10% (v/v) of 1:10 dimethyl sulfoxide/agent : water containing test mixtures for 0.5 hr prior to plasmid addition and transforming conditions. Transformation was inhibited for all antimicrobials and showed a positive association wich higher concentration. Additional testing of ionophore compounds separately and in combination with bacitracin, chlortetracycline, lincomycin, roxarsone, tylosin, and virginiamycin at representative feed concentrations demonstrated 80.6% to >99.9% inhibition (P < 0.001) of resistance transfer. Bacitracin alone inhibited transformation within the range of 50-500 ppm. No increase in resistance transfer was observed when poultry-derived and reference gram-negative isolates having low or no transformation efficiency were additionally tested. The results suggest that these compounds, at relevant concentrations used in animal feed, may interfere with cell envelope-associated DNA uptake channels or other transformation competence mechanisms. Through these mechanisms, ionophores and cell membrane interactive feed agents such as CTC and bacitracin may act to inhibit resistance transfer mechanisms within poultry and livestock. PMID- 15283419 TI - The effects of water supplementation with vitamin E and sodium salicylate (Uni Sol) on the resistance of turkeys to Escherichia coli respiratory infection. AB - The objective of this study was to determine the prophylactic efficacy of two commercial products, soluble vitamin E and soluble sodium salicylate (Uni-Sol), in an Escherichia coli respiratory challenge. The drinking water of male turkey poults was nonsupplemented or supplemented with either vitamin E or Uni-Sol or a combination of both at dosages recommended by the manufacturer. There were 110 birds in each of the four treatments, housed in four floor pens per treatment. At 5 wk of age, birds in half of the pens were challenged with an air sac inoculation of approximately 50 colony-forming units of E. coli. Water treatment commenced 5 days before challenge and continued for 2 wk after challenge, when birds were necropsied. All water treatments prevented the decrease in body weight due to E. coli challenge; however, either vitamin E or Uni-Sol alone, but not the combination of the two, decreased body weight in nonchallenged controls. Either vitamin E or Uni-Sol treatment alone, but not the combination of the two, significantly decreased mortality and air sacculitis scores of challenged birds, and all treatments decreased the isolation rates of E. coli from the liver. All treatments protected liver, spleen, and bursa weights (relative to body weight) from the effects of E. coli challenge, and Uni-Sol alone or vitamin E with Uni Sol protected relative heart weights from the effect of challenge. Uni-Sol treatment alone increased the main effect mean total leukocyte counts and the number and percent of lymphocytes. Uni-Sol in combination with vitamin E increased the number of lymphocytes of challenged birds. Uni-Sol alone decreased the main effect mean heterophil/lymphocyte ratio (H/L) ratio, whereas vitamin E alone increased the H/L ratio of challenged birds. These results indicate that treatment of turkey poults with vitamin E or Uni-Sol prior to and during the stressful events that can lead to colisepticema may decrease disease incidence and mortality. PMID- 15283421 TI - Phenotypic and genotypic characterization of Salmonella arizonae from an integrated turkey operation. AB - Fifty cases submitted between 2000 and 2002 were selected for retrospective analysis to evaluate possible relationships between Salmonella arizonae isolated from breeder flocks, hatching eggs, and meat bird flocks belonging to a single turkey integrator. In all the meat bird cases selected for this study, arizonosis was the primary diagnosis. In birds under 1 month of age, clinical signs and pathologic changes were observed in older birds. The Salmonella arizonae isolates were analyzed by antibiotic resistance pattern and serotype and genotyped by pulsed-field gel electrophoresis (PFGE). Serotyping and PFGE yielded similar results, but the antibiotic resistance patterns did not correspond to either serotyping or PFGE typing. The presence of common pulsed-field patterns in breeder flocks, eggs, and meat bird flocks suggested that S. arizonae was being transmitted vertically from the breeder flock. PMID- 15283420 TI - Development of a virosome vaccine against avian metapneumovirus subtype C for protection in turkeys. AB - An avian metapneumovirus (aMPV) virosome vaccine was prepared and tested for protection of turkeys by aMPV challenge. The vaccine was produced using a detergent-based (Triton X-100) extraction of aMPV subtype C followed by detergent removal with SM2 Bio-Beads. Western blot and virus-neutralization analysis confirmed that the aMPV virosomes contained both the fusion and attachment glycoproteins. Specific-pathogen-free turkeys were immunized either intranasally (i.n.) or intramuscularly (i.m.) with two doses of the aMPV virosome vaccine. Vaccination decreased clinical signs of disease following virulent challenge, and IN vaccination was superior to i.m. vaccination in reducing clinical signs. Decreases in viral load in the respiratory tract were observed in turkeys receiving i.n. vaccination with aMPV virosomes compared to unvaccinated poults. Increased virus-neutralizing antibody levels against aMPV were observed in birds vaccinated with virosomes. These results demonstrate that immunization of turkeys with aMPV virosomes can be an effective strategy for control of disease. PMID- 15283422 TI - Multiple antimicrobial resistance region of a putative virulence plasmid from an Escherichia coli isolate incriminated in avian colibacillosis. AB - Infections due to Escherichia coli have been costly to the poultry industry, but the exact virulence mechanisms used by these organisms to cause disease in birds remain undefined. Several factors have been shown to contribute to the virulence of avian E. coli, and many of the genes encoding these factors have been found on large conjugative plasmids. Because of the occurrence of antimicrobial resistance genes on these same plasmids, it is possible that the use of antimicrobial agents may select for persistence of E. coli containing such plasmids. In the present study, a subclone of one of these plasmids was identified as likely containing some virulence and antimicrobial resistance genes. In an effort to better understand the relationship between virulence and resistance in these plasmids, this subclone was sequenced and the sequence analyzed. Analysis of this 30 kilobase (kb) region of plasmid pTJ100 revealed a mosaic of virulence genes, insertion sequences, antimicrobial resistance cassettes, and their remnants. Many of the resistance genes found in this region were expressed under laboratory conditions, indicating that certain antimicrobial agents, including disinfectants, antibiotics, and heavy metals, could promote selection of E. coli containing such plasmids in the production environment. Also, analysis of the G + C content of this clone indicated that it is the likely consequence of a complex evolution with components derived from various sources. The occurrence of many mobile elements in conjunction with antimicrobial resistance and virulence genes in this 30-kb region may indicate that the genetic constitution of the clone is quite plastic. Although further study will be required to better define this plasmid's role in avian E. coli virulence, the sequence described here is, to our knowledge, the longest known contiguous sequence of a ColV plasmid yet presented. Analysis of this sequence indicates that this clone and its parent plasmid may be important to the pathogenesis of avian colibacillosis and the evolution of avian E. coli virulence. PMID- 15283423 TI - Effect of a variant infectious bursal disease virus (E/Del) on Salmonella typhimurium infection in commercial broiler chickens. AB - The effect of infectious bursal disease virus (IBDV) on Salmonella typhimurium (ST) infections in broilers was investigated in terms of Salmonella shedding and persistence, pathogenicity, and isotype specific humoral immune responses. Thirty six, 1-day-old, straight-run commercial broiler chickens that were Salmonella negative by polymerase chain reaction (PCR) and culture were divided into two groups of 18 chicks each (ST and ST-IBDV). One group (ST-IBDV) of chicks received the E/Del strain of IBDV (10(5.0) median tissue culture infective dose [TCID50]/ml) through the ocular and cloacal routes divided into doses of 50 microl each at 2 days of age. Both groups were then inoculated with 10(8) colony forming units (CFU)/ml nalidixic acid-resistant ST in the drinking water at 3 days of age. Environmental Salmonella counts were higher in the ST-IBDV group at 2 and 3 wk postinfection (PI) compared to the ST group. ST carriage in the cecal contents between the ST and ST-IBDV groups was not statistically different. The ST-IBDV group had a single mortality at 10 days postinfection compared to none in the ST group. The ST-IBDV group had significantly lower bursa to body weight ratios at 4 and 6 wk, as well as higher bursal lesion scores than the ST group at 2, 4, and 6 wk PI. The ST group had significant increase in serum IgG from 2 to 6 wk PI in comparison to the ST-IBDV group, which had no significant changes over time. Both IgA and IgM were significantly increased at 4 and 6 wk relative to 2 wk levels. There was an IBDV-induced failure of anti-Salmonella IgG seroconversion over time in ST-IBDV. Both groups continued to shed high levels of Salmonella up to the end of the study despite high antibody levels in the ST group and an unimpaired IgM and IgA production in the ST-IBDV group, indicating a limited influence of humoral immunity on Salmonella clearance. PMID- 15283424 TI - Development of a protective index to rank effectiveness of multiple treatments within an experiment: application to a cross-protection study of several strains of Eimeria maxima and a live vaccine. AB - Vaccination of chickens with live oocysts has become a more widely used method for controlling avian coccidiosis as resistance to anticoccidial medication increases. However, some coccidia strains are not useful in multispecies vaccines because antigenic variation has made them generally less protective. In order to experimentally test a number of strains for the best cross protection, we have devised an evaluation method using four independently measured variables: weight gain, lesion score, plasma carotenoids, and plasma NO2- + NO3-. These values, when measured at 6 days postchallenge, tend to be significantly correlated. A protective index (PX) is calculated for each chicken using the following algorithm: PX = (Ngain + Ncarotenoids) - (Nsqrls + N[NO2- + NO3-]), where the prefix N indicates values for a variable normalized against a mean of that variable from a control group. Nsqrls values are normalized values of the square roots of lesion scores. The PX can then be treated as a dependent variable. In this study, mean PX values of unchallenged groups cluster around 0. Mean PX values of protected chickens are statistically close to those from unchallenged groups, whereas unprotected chickens have highly negative mean PX values. PMID- 15283425 TI - Molecular characterization of low pathogenicity H7N3 avian influenza viruses isolated in Italy. AB - The complete coding regions of the surface glycoproteins, nucleoprotein (NP), polymerase 2 (PB2), and matrix (M) of A/turkey/214845/02 and A/turkey/220158/99 (H7N3) low pathogenicity avian influenza (LPAI) viruses isolated in October 2002 in Italy were amplified and sequenced to determine the epidemiologic relationships with an A/turkey/Italy/4603/99 (H7N1/4603/99) LPAI virus isolated during the 1999-2001 epizootic in Italy. The hemagglutinin (HA) of H7N3 viruses showed 97.8% nucleotide similarity with A/turkey/Italy/4603/99 (H7N1), and NP, M, and PB2 gene similarities were 93.6%, 98.2%, and 96.2%, respectively. Phylogenetic analyses of HA, PB2, and M genes showed that H7N3 and H7N1 viruses were closely related. Sequence analysis revealed a 23 amino acid deletion in the stalk of the neuraminidase of H7N3 viruses and a unique deletion of amino acid glycine in position 17 in the NP gene of H7N1 virus. PMID- 15283426 TI - Interactions of butyric acid- and acetic acid-treated Salmonella with chicken primary cecal epithelial cells in vitro. AB - In vitro studies of the interaction between pathogenic bacteria and the chicken intestinal epithelium are hampered by the lack of a host- and tissue-specific in vitro model. Therefore, a reproducible method for isolation and cultivation of chicken primary cecal epithelial cells was developed. Cecal crypts were isolated and cultured in vitro to form a semiconfluent layer of epithelial cells. Incubation of Salmonella enteritidis with these cells resulted in invasion. Pretreatment of the Salmonella bacteria with butyric acid resulted in a significant decrease of invasion of the bacteria in the chicken cecal epithelial cells, whereas pretreatment with acetic acid increased invasiveness. These interactions of S. enteritidis with primary chicken cecal epithelial cells were similar to the interactions with other epithelial cell types. PMID- 15283427 TI - Determination of the acute 50% lethal dose T-2 toxin in adult bobwhite quail: additional studies on the effect of T-2 mycotoxin on blood chemistry and the morphology of internal organs. AB - Three experiments were conducted to assess mortality rate, blood chemistry, and histologic changes associated with acute exposure to T-2 mycotoxin in adult bobwhite quail. In Experiment 1, adult quail were orally dosed with T-2 toxin to determine the lethal dose that resulted in 50% mortality of the affected population (LD50), and that dose was determined to be 14.7 mg of T-2 toxin per kilogram of body weight (BW). A second experiment was performed to study the effects of 12-18 mg/kg BW T-2 toxin on blood chemistry and liver enzyme profiles. Posttreatment uric acid, aspartate aminotransferase, lactic dehydrogenase, and gamma glutamyltransferase increased as compared with pretreatment values. In contrast, posttreatment plasma total protein, cholesterol, and triglyceride levels numerically decreased as compared with pretreatment values. Changes in blood chemistry values were consistent with liver and kidney damage after T-2 toxin exposure. In Experiment 3, histologic analyses of bone marrow, spleen, liver, small intestine, kidney, and heart were conducted on birds dosed in Experiment 2. Marked lymphocyte necrosis and depletion throughout the spleen, thymus, bursa, and gut-associated lymphoid tissue in the small intestine were observed in birds dosed with 15 and 18 mg/kg BW T-2 toxin. Necrosis of liver and lipid accumulation as a result of malfunctioning hepatocytes were also observed. Little or no morphologic change was observed in bone marrow and heart tissue. The LD50 for adult bobwhite quail as found in this study is two to three times higher than that reported for other species of commercial poultry. Results from these data confirm previous reports of immunosuppressive and/or cytotoxic effects of T 2 toxin in other mammalian and avian species. T-2 toxin may have a negative impact on the viability of wild quail populations. PMID- 15283428 TI - Detection of avian polyomavirus infection by polymerase chain reaction using formalin-fixed, paraffin-embedded tissues. AB - Avian polyomavirus infection in psittacines was diagnosed in tissues by the use of polymerase chain reaction (PCR) test. The tissues used in the procedure were either formalin-fixed tissues embedded in paraffin blocks or fresh tissues (heart, liver, and spleen) collected from the psittacines during necropsy. DNA was extracted from these tissues and was tested with the published primers for avian polyomavirus VP1 gene in the PCR that yielded an amplicon of 550 base pair size, which was then visualized by electrophoresis. The amplicon size was consistent with avian polyomavirus. The PCR test was found to be an effective method for identifying avian polyomavirus infection in both formalin-fixed, paraffin-embedded and fresh tissues from psittacine birds of different age groups. PMID- 15283429 TI - The normal electrocardiogram of the unanesthetized peregrine falcon (Falco peregrinus brookei). AB - The mean duration and amplitudes of the lead II electrocardiogram were determined in the peregrine falcon (Falco peregrinus brookei) using 10 birds ranging in age from 1 to 5 yr. Electrocardiograms were performed on unanesthetized falcons in order to avoid the anesthesia effect on the electrocardiogram, by a method which seems to induce a tonic immobility-like reaction. All the falcons had a normal sinus rhythm, with a mean heart rate of 268 beats per minute. Mean durations of PR, ST, QT, and RR intervals were higher (but not statistically significant) in females than in males, except for the ST segment, with similar values in both sexes. P-wave deflections were positive in I, II, III, aVL, and aVF and negative in aVR. The normal patterns of wave forms of the QRS complexes in all leads were of QS and rS types, except for aVR and aVL, which presented an R configuration. The mean electrical axis was negative, with an average of -99.9 degrees. T-wave deflections were positive in I, II, III, and aVF leads II and negative in aVR and aVL. The data collected in this study may serve as a guide for electrocardiographic monitoring of peregrine falcons. PMID- 15283430 TI - Pathogenicity and molecular analysis of an infectious bursal disease virus isolated from Malaysian village chickens. AB - The characteristics of the pathogenic infectious bursal disease virus (IBDV) that infected avian species other than commercial chickens were largely unknown. In this study, by using in vivo and molecular methods, we had characterized an IBDV isolate (named 94268) isolated from an infectious bursal disease (IBD) outbreak in Malaysian village chickens--the adulterated descendant of the Southeast Asian jungle fowl (Gallus bankiva) that were commonly reared in the backyard. The 94268 isolate was grouped as the very virulent IBDV (vvIBDV) strain because it caused severe lesions and a high mortality rate in village chickens (>88%) and experimentally infected specific-pathogen-free chickens (>66%). In addition, it possessed all of the vvIBDV molecular markers in its VP2 gene. Phylogenetic analysis using distance, maximum parsimony, and maximum likelihood methods revealed that 94268 was monophyletic with other vvIBDV isolates and closely related to the Malaysian vvIBDV isolates. Given that the VP2 gene of 94268 isolate was almost identical and evolutionarily closely related to other field IBDV isolates that affected the commercial chickens, we therefore concluded that IBD infections had spread across the farm boundary. IBD infection in the village chicken may represent an important part of the IBD epidemiology because these birds could harbor the vvIBDV strain and should not be overlooked in the control and prevention of the disease. PMID- 15283432 TI - The presence of nicotinamide adenine dinucleotide-independent Haemophilus paragallinarum in Mexico. AB - Two isolates of Haemophilus paragallinarum were obtained from a layer chicken in Mexico. The isolates were confirmed as H. paragallinarum by polymerase chain reaction and conventional biochemical identification. The isolates were nicotinamide adenine dinucleotide (NAD) independent-growing on blood agar without the need of a nurse colony as well as on a complex medium that lacked both NAD and chicken serum. Both isolates were pathogenic, causing the typical clinical signs of infectious coryza in susceptible chickens. One isolate was Page serovar B/Kume serovar B-1 and the other isolate was Page serovar C/Kume serovar C-2. The isolates were associated with a field outbreak that involved an egg drop of 20% over a 3-wk period and a doubling of weekly mortality (from 0.1% to 0.2%). This is the first report of NAD-independent H. paragallinarum outside South Africa and is the first time that NAD-independent H. paragallinarum of serovar B has been reported. PMID- 15283431 TI - Mortality factors, environmental contaminants, and parasites of white-tailed sea eagles from Greenland. AB - Twelve white-tailed sea eagles (Haliaeetus albicilla groenlandicus) found dead between 1997 and 2000 in Greenland were examined to investigate the health status, including the causes of death and the burden of organochlorine contaminants and potentially toxic heavy metals. The determined causes of death were unspecific trauma (n = 6), lead poisoning (n = 2) with 36 and 26 ppm lead in the liver tissue, infectious diseases (n = 1), injuries sustained during intraspecific conflict (n = 1), and gunshot (n = 1). One lead poisoned eagle had a single lead shot pellet in its gizzard. No diagnosis could be made in one case because of decomposition of the carcass. Four of the investigated eagles were injured with lead shot or bullet fragments; one of the birds was killed with about 69 lead shots. Levels of organochlorine pesticides, polychlorinated biphenyls, mercury, and cadmium in organs were moderate. The parasite fauna consisted of one coccidian and three helminth species. The acanthocephalas Profiliocollis botulus and Corynosoma suduche as well as the nematode Stegophorus stellaepolaris are all new records for the white-tailed sea eagle. PMID- 15283433 TI - Mild infectious laryngotracheitis in broilers in the southeast. AB - During 2001, a mild infectious laryngotracheitis virus (ILTV) infection occurred in broiler flocks in the southeastern United States. Clinical signs included mild tracheitis, swollen sinuses, and conjunctivitis, with no increased mortality and minimal serologic response. Infrequent intranuclear inclusion bodies with or without syncytial cell formation were observed in eyelid, trachea, and larynx and in the chorioallantoic membrane of infected embryos. Immunohistochemistry and a nested infectious laryngotracheitis polymerase chain reaction (ILT PCR) were utilized to confirm the presence of ILTV nucleic acid in fixed tissues. In addition, 2-wk-old specific-pathogen-free (SPF) birds inoculated with field material exhibited the mild signs observed in broilers in the field. Tracheal swabs and tissues taken from these SPF birds were also positive by nested ILT PCR. Restriction fragment length polymorphism analysis of ILT PCR products indicated that ILT virus associated with mild respiratory disease in the southeast is related to the chicken embryo origin vaccine type strains. PMID- 15283434 TI - Mycobacteriosis in an American bald eagle (Haliaeetus leucocephalus). AB - Avian mycobacteriosis is an important disease in companion, captive, exotic, and wild birds worldwide. Mycobacterium avium is the most widely distributed and pathogenic organism causing tuberculous lesions in birds. Multiple factors including age, stress, immune status, and preexisting disease determine the pathogenicity of M. avium, and the disease can manifest itself in a variety of forms. Mycobacteriosis can cause severe losses in zoo aviaries, including the loss of rare and endangered bird species. We report a case of systemic avian mycobacteriosis in an adult, free-living male American bald eagle (Haliaeetus leucocephalus) that presented to the Diagnostic Center for Population and Animal Health in November 2003. PMID- 15283435 TI - Recent trends. PMID- 15283436 TI - MCOs: cost centers or profit partners for employers? AB - Although health insurance has traditionally been a cost center for employers, and they are ever striving to reduce such costs, MCOs have the opportunity to become a means for employers to save costs rather then suffer more. By focusing on the total health-related costs affecting businesses' bottom lines, MCOs can become "account managers, " where the "account" reflects all the costs that they can positively affect through their own initiatives, and thereby become true solutions rather than costs. This method will enable them to differentiate themselves from competitors and generate "solution pricing"-based new revenue focusing on outcomes versus coverage. This article covers the rationale for and basics of such a strategy. PMID- 15283437 TI - Have "evergreen" provider contracts lost their bloom? AB - An "evergreen contract" is one that automatically renews itself from year to year, unless one of the contracting parties acts at specified intervals (which may be annually or as long as several years) to give notice in the manner required to terminate the otherwise perpetual agreement. Such contracts are in contrast to fixed-term agreements, where both parties must affirmatively agree to extend the term beyond the initial specified term. Such "evergreen contracts" are legally valid, but for compelling reasons health plans and providers may want to consider terminating their evergreen provider contracts now in favor of a contract with a fixed term. PMID- 15283438 TI - Towards improving health care delivery for people with physical disabilities: findings from focus groups with health care consumers in Minnesota. AB - Specialized managed care programs such as the MnDHO program can improve the health care experience for people with disabilities if they include seamless care coordination among disability competent providers and promote consumer participation. Findings indicate greater satisfaction with care coordination, consumer participation, and provider knowledge in the AXIS/UCare complete program. PMID- 15283439 TI - Guide for clinical trials disappointing. PMID- 15283440 TI - The American Psychiatric Association's Resource Document on Mental Retardation and Capital Sentencing: implementing Atkins v. Virginia. PMID- 15283441 TI - Spinal mechanisms contributing to joint pain. AB - Nociceptive input from the joint is processed in different types of spinal cord neurons. A proportion of these neurons are only activated by mechanical stimulation of the joint and other deep tissue, e.g. adjacent muscles. Other neurons are activated by mechanical stimulation of joint, muscles and skin. The majority of the neurons are wide dynamic-range neurons (small responses to innocuous pressure to deep tissue and stronger and graded responses to noxious mechanical stimulation). Importantly, neurons with joint input show pronounced hyperexcitability during development of joint inflammation (enhanced responses to mechanical stimulation of the inflamed joint as well as to healthy adjacent deep structures, reduction of mechanical threshold in high threshold neurons and expansion of the receptive field). Thus inflammation induces neuroplastic changes in the spinal cord which alter nociceptive processing. This state of hyperexcitability is maintained during persistent inflammation. The neurons are under strong control of descending inhibition which increases at least during the acute phase of inflammation. Several transmitters and mediators contribute to the generation and maintenance of inflammation-induced spinal hyperexcitability including glutamate, substance P, neurokinin A, CGRP, prostaglandins and probably others. The latter compounds show enhanced release and an altered release pattern during inflammation in the joint. PMID- 15283442 TI - Activation of sensory neurons in the arthritic joint. AB - Joints are richly innervated with a range of sensory nerve fibres that convey information to the central nervous system about forces exerted on articular tissues by both low and high threshold mechanical stimuli. High threshold nociceptive afferents terminate primarily in the synovium and periosteum, and normally respond only to movement of the joint beyond the working limits. Following joint damage, two factors combine to alter the mechanical sensitivity of articular nociceptors. Firstly, physical changes (joint effusion and tissue oedema) alter the resting and movement-induced forces exerted on the joint tissues and secondly, inflammatory mediators released within the damaged tissue sensitize articular nociceptive afferents by binding to receptors on the nerve endings. These factors result in a reduction of the mechanical threshold for activation of articular nociceptors such that manipulation of the joint within the normal range is easily sufficient to activate them. Acute and chronic animal models of joint inflammation have been used to study the mechanisms of articular nociceptor sensitization and a number of inflammatory mediators and their receptors have been implicated. The focus of this paper will be to introduce some of the important issues involved in the sensitization of nociceptive articular afferents. PMID- 15283443 TI - Neuromuscular aspects of osteoarthritis: a perspective. AB - Osteoarthritis (OA) represents failure of the diarthrodial joint and may be due to a primary abnormality in any of the tissues of the joint, e.g. articular cartilage, subchondral bone, synovium, periarticular muscle, or sensory nerves whose termini lie within the joint. Neuropathic arthropathy, due to severe sensory neuropathy, causes severe and rapid breakdown of joints. We have shown that interruption of sensory input from the ipsilateral hind limb strikingly accelerates progression of OA after anterior cruciate ligament transection in the dog; a clinical correlate exists in humans with diabetic neuropathy who sustain even minor joint trauma. Knee OA in humans is accompanied by defects in proprioception, although it is not clear whether the neurological abnormality is primary or a consequence of intra-articular pathology. The magnitude of the load on a joint and, especially, the rate of impulsive loading, influence development of OA. It is relevant, therefore, that quadriceps weakness may precede development of knee OA in some people, insofar as this may diminish the effectiveness of protective muscular reflexes and thereby increase deleterious joint loading. Individuals vary with respect to how they load their joints, perhaps because of genetic differences in central program generators. PMID- 15283444 TI - Current perspectives on the clinical presentation of joint pain in human OA. AB - Pain is the commonest symptom of osteoarthritis (OA), the principal reason why individuals seek medical care and a major determinant of other outcomes such as disability and joint replacement. Most studies have examined knee OA: little is known about other sites. Community studies indicate only a modest relationship between structural change on X-ray and reporting of pain. Many community subjects, for example, fail to complain of pain despite extensive X-ray change, while others report pain with normal X-rays. Pain severity of patients attending hospital is even less related to X-ray change, being more dependent on body mass index (BMI), coping strategies and psychosocial variables. Many patients can identify more than one type of pain. It is increasingly clear that OA pain is heterogeneous, being classifiable on the basis of location, precipitating factors, response to anti-inflammatory and steroid medication and the effects of local anaesthetic. This potential to classify OA pain represents a useful tool with which to test hypotheses regarding structural origin of pain. PMID- 15283445 TI - Joint mechanics in osteoarthritis. AB - The primary goal of our research has been to quantify the in vivo loading of normal and osteoarthritic (OA) joints, and to determine the corresponding biological responses. Much of the research in this area has been performed using articular cartilage explants. We feel that, although critically important to our understanding of cartilage mechanics and biology, these experiments may not be directly transferable to interpreting the in vivo joint mechanics and elucidating the detailed mechanisms of onset and progression of OA. Therefore, we have attempted to measure the loading of the knee in freely moving feline and lapine models of OA. We have found that, upon anterior cruciate ligament transection in the cat, knee joints are more flexed, muscle forces are decreased and muscle control patterns are destroyed. Articular cartilage initially becomes thicker, softer and more permeable, resulting in generally increased joint contact areas and decreased peak pressures in the initial stages of joint degeneration compared to control values. Based on our results, we speculate that unloading of the joint (rather than overloading), combined with poor muscular control and weakness, might constitute risks for the onset of joint degeneration. PMID- 15283446 TI - Characterization of joint pain in human OA. AB - Characterization and differentiation of joint pain is difficult. Though degenerative changes in joints are frequent causes of pain in hip and knee, these changes are not always painful, and other possible causes of pain must also be considered. In degenerative changes in the spine, the problem is even more complex, as peripheral neuropathic pain, caused by mechanical compression and/or leakage of cytokines irritating nerve roots may be difficult to differentiate from nociceptive pain from intervertebral joints, discs or muscles. We know now that nociceptive pain has often referred to areas of pain with numbness and parestesthethic sensations, previously regarded as characteristic for neurogenic pain. Furthermore, in patients with painful coxarthrosis quantitative sensory testing (QST) has shown disturbed sensory thresholds not only in regions adjacent to the affected hip but also contralaterally. These sensory disturbances, previously noted in neuropathic pain, normalized after successful surgery with relief of pain, thus confirming the relation to the hip joint. Patients with painful coxarthrosis also have moderately increased substance P activity in cerebrospinal fluid. Thus the findings show some similarities with fibromyalgic patients with highly increased substance P in cerebrospinal fluid and sensory disturbances. In conclusion, joint pain has a profound impact on the sensory system and need a multimodal approach. PMID- 15283447 TI - The role of inflammatory mediators on nociception and pain in arthritis. AB - Pain is the most common complaint of individuals with osteoarthritis but the cause of symptoms in this disorder remains unclear. Quantitative sensory testing reveals that in patients with chronic joint disease there is diffuse and persistent alteration of nociceptive (pain) pathways, irrespective of the level of activity of the underlying disease. Inflammatory mediators contribute to this plasticity either by directly activating high threshold receptors or more commonly by sensitizing nociceptive neurons to subsequent everyday stimuli. This involves early post-translational modification of receptors/ion channels and later, longer-lasting transcription-dependent mechanisms involving changes to the chemical phenotype of the neuron. Included amongst these changes are the increased production and release of various pro- and anti-inflammatory neuropeptides which have diverse actions on both circulating and resident cell populations. These neurally derived mediators act synergistically with cytokines and growth factors to contribute to ongoing tissue injury. It is becoming apparent that the interaction between a damaged joint and the sensory nervous system is far from straightforward and that activity arising from such interactions may produce not only pain but may also influence the subsequent course of the underlying disease. PMID- 15283448 TI - Molecular events of chronic pain: from neuron to whole animal in an animal model of osteoarthritis. AB - A novel animal model of osteoarthritis has been established for studies on the disease process as well as on mechanisms underlying the symptoms of osteoarthritis, principally pain and fatigue. The model is established by cutting the anterior cruciate ligament in the rat to introduce instability of the joint, removing the medial meniscus to induce further derangement of the joint, and exercising the rat on a modified rota-rod to generate weight-bearing mobility of the joint; this exercise can be regulated in terms of duration to govern severity of the model. The model exhibits many of the characteristics of osteoarthritis in humans, including bone and cartilage remodelling, infiltration of the joint tissues by immune cells and alterations in sensory mechanisms. This model will find application in development of novel interventions for the treatment of osteoarthritis and its symptoms as well as development of diagnostic tools for early detection. PMID- 15283449 TI - Phantoms in rheumatology. AB - This paper examines rheumatology pain and how it may relate to amputee phantom limb pain (PLP), specifically as experienced in rheumatoid arthritis, fibromyalgia and complex regional pain syndrome (CRPS). Clinical findings, which suggest cortical sensory reorganization, are discussed and illustrated for each condition. It is proposed that this sensory reorganization generates pain and altered body image in rheumatology patients in the same manner as has previously been hypothesized for amputees with PLP; that is via a motor/sensory conflict. The correction of this conflict through the provision of appropriate visual sensory input, using a mirror, is tested in a population of patients with CRPS. Its analgesic efficacy is assessed in those with acute, intermediate and chronic disease. Finally, the hypothesis is taken to its natural conclusion whereby motor/sensory conflict is artificially generated in healthy volunteers and chronic pain patients to establish whether sensory disturbances can be created where no pain symptoms exists and exacerbated when it is already present. The findings of our studies support the hypothesis that a mismatch between motor output and sensory input creates sensory disturbances, including pain, in rheumatology patients and healthy volunteers. We propose the term 'ominory' to describe the central monitoring mechanism and the resultant sensory disturbances as a dissensory state. PMID- 15283450 TI - Bone pain and pressure in osteoarthritic joints. AB - Intraosseous hypertension has been associated with a deep aching bone pain, particularly at rest, in subsets of patients with osteoarthritis of the hip and knee. The pathophysiology of this problem remains uncertain, but intraosseous phlebography implicates outflow impairment at relatively distal venous sites. Although the issue has been controversial, intraosseous pressures rise normally, and painlessly, when epiphyseal bone is loaded and these pulses may be mechanically meaningful in the distribution and transmission of impact energy. Increased outflow resistance may amplify the episodic pressure response with subsequent intravasation of epiphyseal fat leading to 'marrow oedema' and altered mechanics. The relationship between persisting pain and pressure is an old but convincing association. Its precise mechanism in osteoarthritis remains in need of an adequate explanation. PMID- 15283451 TI - Structural associations of osteoarthritis pain: lessons from magnetic resonance imaging. AB - For many years the search for structural associations of osteoarthritis (OA) pain were based on conventional radiographic imaging that predominantly visualizes bone. As well as being tomographic, magnetic resonance imaging (MRI) has the ability to directly visualize all the structures of a joint, including soft tissue and cartilage. Initial MRI studies focused on cartilage assessment, but recently there has been a growing body of work examining the correlation of structural findings with pain in OA and their relation to structural progression. Painful OA knees have more MRI-detected abnormalities and these pathologies are often correlated making individual contributions difficult to assess. However, in large cohort studies, both synovial hypertrophy and large synovial effusions were demonstrated to be more frequent in patients with OA knee pain. Similarly MRI determined subchondral bone marrow oedema lesions (BME), particularly large ones, are associated with OA knee pain. Meniscal tears in OA knees, although common, have not been linked with pain. Improved, reliable quantification of the structural features and the rapid advances in MRI technology can only improve structure-pain understanding. PMID- 15283452 TI - The role of TRP channels in sensory neurons. AB - Two parallel processes characterize the contemporary pain field. Firstly, enormous progress is being made in the discovery of the cellular and molecular mechanisms responsible for the pathogenesis of pain and secondly, there is a growing appreciation that multiple mechanisms contribute to common clinical pain syndromes. The aim of this chapter is to provide a short overview how transient receptor potential (TRP) channels could contribute to acute and chronic pain states. TRP channels of the vanilloid family (TRPV1, TRPV2, TRPV3, TRPV4) are excited by heat stimuli whereas TRPM8 and ANKTM1 are cold responsive. TRPV1 and ANKTM1 are mediating the pungency of nociceptor-specific chemicals such as capsaicin or mustard oil. Sensitization of TRPV1 is an important mechanisms for heat hyperalgesia and thus the generation of chronic pain symptoms. PMID- 15283453 TI - Mechanisms that generate and maintain bone cancer pain. AB - Although bone cancer pain can be severe and is relatively common, as it frequently arises from metastases from breast, prostate, and lung tumours, very little is known about the basic mechanisms that generate and maintain this chronic pain. To begin to define the mechanisms that give rise to bone cancer pain, we have developed mouse and rat models using the intramedullary injection and containment of tumour cells into the femur. These tumour cells induced bone remodelling as well as ongoing and movement evoked pain behaviours similar to that found in patients with bone cancer pain. In addition there is a significant reorganization of the spinal cord that received sensory input from the cancerous bone and this reorganization generated a neurochemical signature of bone cancer pain that is both dramatic and significantly different from that observed in mouse and rat models of chronic neuropathic or inflammatory pain. These models have provided insight into the mechanisms that drive cancer pain and have begun to allow the development of mechanism-based therapies. Together these advances should reduce tumour-induced pain and suffering and significantly improve the quality of life of cancer patients. PMID- 15283454 TI - Symmetry, T cells and neurogenic arthritis. AB - Symmetry in clinical disease occurs more commonly than expected by chance and is unexplained. In this paper we focus on symmetry in arthritis and describe the neurogenic hypothesis. Neuropeptides are anatomically relevant to systemic arthritis and have been shown to have modulating effects on both the immune and circulatory systems. Neural networks project bilaterally and are involved in the development and propagation of inflammatory disease. These putative pathological neuro-feedback loops may derive from the existence of biologically protective symmetrical mechanisms. PMID- 15283455 TI - Lessons from fibromyalgia: abnormal pain sensitivity in knee osteoarthritis. AB - Fibromyalgia (FM) is a disorder that is characterized by widespread, musculoskeletal pain and abnormal pain sensitivity at multiple anatomic sites. Laboratory studies involving psychophysical and neuroimaging methods suggest that central augmentation of low intensity stimulation may contribute to abnormal pain sensitivity in FM. Recently, several investigators, using similar laboratory methods, have shown that patients with knee or hip osteoarthritis (OA) exhibit abnormal pain sensitivity or abnormal pain inhibition at anatomic sites distal to affected joints. Consistent with animal models of central sensitization, differences between patients and healthy controls in pain processing and pain inhibition at these distal sites are eliminated after nociceptive input is eliminated following total joint replacement surgery. This paper reviews these findings from our laboratory and those of independent investigators. It also presents verbal, psychophysical and neuroimaging data concerning ethnic group differences in affective and cognitive pain responses among patients with knee OA. We suggest that central sensitization as well as centrally-mediated cognitive and affective factors influence the pain responses of patients with knee OA. In addition, ethnic group differences in pain cognition and affect may contribute to differences among these groups in preferences for healthcare interventions such as total joint replacement. PMID- 15283456 TI - Synthesis and cytotoxicity of 2,5-dihydroxychalcones and related compounds. AB - A series of 2, 5-dihydroxychalcones and related compounds were synthesized, and their cytotoxicities against tumor cell lines and human umbilical venous endothelial cells (HUVEC) evaluated. It was found that chalcones, with electron withdrawing substituents on an A ring, exhibited significant cytotoxicities. Among the synthesized compounds, 2'-chloro-2, 5-dihydroxychalcone (9) was most potent, with an IC50 value as low as 0.31 microg/mL. This compound also exhibited a significant cytotoxic selectivity toward HUVEC. PMID- 15283457 TI - Flavonoids from Spatholobus suberectus. AB - Two pterocarpans [(6aR,11aR)-maackiain, (6aR,11aR)-medicarpin], one flavanone [(2S)-7-hydroxy-6-methoxy-flavanone], one isoflavan (sativan) and two isoflavones (pseudobaptigenin, genistein) were isolated from the Spatholobus suberectus (Leguminosae). Their chemical structures were determined by comparison of their spectroscopic parameters of CD, EIMS, 1D-NMR and 2D-NMR with those reported in the literatures. All of these compounds are reported for the first time from this plant through the present study. PMID- 15283458 TI - Isolation of flavonoids and a cerebroside from the stem bark of Albizzia julibrissin. AB - From the EtOAc fraction of the MeOH extract of Albizzia julibrissin (Leguminosae), a rare 5-deoxyflavone (geraldone, 1), isookanin (2), luteolin (3), an isoflavone (daidzein, 4), five prenylated flavonoids [sophoflavescenol (5), kurarinone (6), kurarinol (7), kuraridin (8) and kuraridinol (9)], a cerebroside (soya-cerebroside I, 10), and (-)-syringaresinol-4-O-beta-D-glucopyranoside (11) were isolated and characterized on the basis of spectral data. Compounds 2, 3, and 11, showed 1,1-diphenyl-2-picrylhydrazyl radical scavenging activity. PMID- 15283459 TI - Norisoprenoids and hepatoprotective flavone glycosides from the aerial parts of Beta vulgaris var. cicla. AB - (+)-Dehydrovomifoliol (1), 3-hydroxy-5alpha,6alpha-epoxy-beta-ionone (2), vitexin 7-O-beta-D-glucopyranoside (3), and vitexin 2''-O-beta-D-glucopyranoside (4) were isolated as new constituents from the aerial parts of Beta vulgaris var. cicla. Compounds 3 and 4 demonstrated hepatoprotective activity with values of 65.8 and 56.1%, respectively, in primary cultured rat hepatocytes with CCl4-induced cell toxicity, compared to controls. This was comparable to that of silibinin (69.8 %) which was used as a positive control. PMID- 15283460 TI - A new indolinepeptide from Paecilomyces sp. J300. AB - A new indolinepeptide (3) was isolated, together with two known compounds, a cerebroside (1) and an alloxazine (2), from silkworm larvae infected with Paecilomyces sp. J300. On the basis of spectroscopic data, their structures were elucidated as (4E, 8E, 2S, 2'R, 3R)-N-2'-hydroxyhexadecanoyl-1-O-beta-D glucopyranosyl-9-methyl-4, 8-sphingadienine (1), 7,8-dimethylalloxazine (2) and 3beta,5-dihydroxy-1-N-methyl-indoline-2beta-carbonyl amino-D-alanyl-erythro-beta hydoxyisoleucinyl-glycine (3). PMID- 15283461 TI - Antioxidant properties of flavone-6(4')-carboxaldehyde oxime ether derivatives. AB - The in vitro antioxidant properties of some flavone-6(4)-carboxaldehyde oxime ether derivatives (Ia-f, IIa-f) were determined by their effects on the rat liver microsomal NADPH-dependent lipid peroxidation (LP) levels by measuring the formation of 2-thiobarbituric acid reactive substances. The free radical scavenging properties of the compounds were also examined in vitro by determining their capacity to scavenge superoxide anions and interact with the stable free radical 2, 2-diphenyl-1-picrylhydrazyl (DPPH). The most active compounds, IIb (Flavone-4'-carboxaldehyde-O-ethyl oxime) and Id (Flavone-6-carboxaldehyde-O-[2 (1-pyrolidino) ethyl] oxime), caused 98 and 79% inhibition of superoxide anion production and DPPH stable free radical at 10(-3) M, respectively. PMID- 15283462 TI - An antioxidant hispidin from the mycelial cultures of Phellinus linteus. AB - In the course of screening for reactive oxygen species scavengers from natural products, an antioxidant was isolated from the mycelial culture broth of Phellinus linteus and identified as hispidin. The hispidin content was reached its maximum level at 12 days after onset of inoculation. About 2.5 mg/mL of hispidin was produced by P. linteus in a yeast-malt medium (pH 5.8, 25 degrees C). Hispidin inhibited 22.6 and 56.8% of the super oxide anion radical, 79.4 and 95.3% of the hydroxyl radical, and 28.1 and 85.5% of the DPPH radical at 0.1 and 1.0 mM, respectively. The positive control alpha-tocopherol scavenged 25.6 and 60.3%, 74.6 and 96.3%, and 32.7 and 77.5% of each radical, respectively, at the same concentrations. However, hispidin showed no significant activity on the hydrogen peroxide radical. PMID- 15283463 TI - Tissue factor inhibitory sesquiterpene glycoside from Eriobotrya japonica. AB - Tissue factor (TF, tissue thromboplastin) is a membrane bound glycoprotein, which accelerates the blood clotting, activating both the intrinsic and the extrinsic pathways to serve as a cofactor for activated factor VII (VIIa). The TF-factor VIIa complex (TF/VIIa) proteolytically activates factors IX and X, which leads to the generation of thrombin and fibrin clots. In order to isolate TF inhibitors, by means of a bioassay-directed chromatographic separation technique, from the leaves of Eriobotrya japonica Lindley (Rosaceae), a known sesquiterpene glycoside (2) and ferulic acid (3) were isolated as inhibitors that were evaluated using a single-clotting assay method for determining TF activity. Another sesquiterpene glycoside (1) was also isolated but was inactive in the assay system. Compound 3 was yielded by alkaline hydrolysis of compound 2. The structures of compounds 1, 2, and 3 were identified by means of spectral analysis as 3-O-alpha-L rhamnopyranosyl-(1-->4)-alpha-L-rhamnopyranosyl-(1-->2)-[alpha-L-rhamnopyranosyl (1-->6)]-beta-D-glucopyranosyl nerolidol (1), 3-O-alpha-L-rhamnopyranosyl-(1-->4) alpha-L-rhamnopyranosyl-(1-->2)-[alpha-L-(4-trans-feruloyl)-rhamnopyranosyl-(1- >6)]-beta-D-glucopyranosyl nerolidol (2) and ferulic acid (3), respectively. Compounds 2 and 3 inhibited 50% of the TF activity at concentrations of 2 and 369 microM/TF units, respectively. PMID- 15283464 TI - A cholesterol biosynthesis inhibitor from Rhizopus oryzae. AB - A bile acid derivative, methyl cholate (1), was isolated from EtOAc extract of the fungus Rhizopus oryzae as a cholesterol biosynthesis inhibitor. It showed moderate inhibitory activity on cholesterol biosynthesis in human Chang liver cells. Compound 1 exhibited inhibitory effect on the later step of cholesterol biosynthesis, indicating that its action mode is different from that of statins that act on the HMG-CoA reductase. PMID- 15283465 TI - Antiinflammatory activity of hyperin from Acanthopanax chiisanensis roots. AB - The chloroform and the ethyl acetate fractions from the roots of Acanthopanax chiisanensis exhibited a significant inhibition of prostaglandin E2 (PGE2) production in rat peritoneal macrophages stimulated by the protein kinase C activator, 12-O-tetradecanoylphorbol 13-acetate (TPA). Hyperin was isolated as an active principle from the ethyl acetate fraction. It suppressed not only PGE2 production but also nitric oxide (NO) production in vitro in a concentration dependent manner, their IC50, being 24.3 and 32.9 microM, respectively. Hyperin also caused a significant inhibition of increase in acetic acid-induced vascular permeability in mice in vivo. PMID- 15283466 TI - Recombinant adenoviral vector containing tumor-specific L-plastin promoter fused to cytosine deaminase gene as a transcription unit: generation and functional test. AB - The expression of therapeutic transgenes in recombinant adenoviral vectors is a major cause of toxicity in dividing cancer cells as well as non dividing normal cells. To solve the problem of toxicity to normal cells, we have reported on a recombinant adenoviral vector system (AdLP-) in which the expression of the transgene is directed by the tumor-specific L-plastin promoter (LP) (Chung et al., 1999). The object of this study was to generate a recombinant adenoviral vector system which would generate tumor cell specific expression of cytosine deaminase (CD) gene. We report the construction of a replication-incompetent adenoviral vector in which CD is driven by the L-plastin promoter (AdLPCD). Infection of 293 cells by AdLPCD generated the functional CD protein as measured by HPLC analysis for the conversion of 5-Fluorocytosine (5-FC) to 5-Fluorouracil (5-FU). HPLC analysis in conjunction with counting radioactivity for [6-3H]-5FC and [6-3H]-5FU demonstrated vector dose-dependent conversion of 5-FC to 5-FU in AdLPCD infected ovarian cancer cells. The results from present and previous studies (Peng et al., 2001; Akbulut et al., 2003) suggest that the use of the AdLPCD/5-FC system may be of value in the treatment of cancer including microscopic ovarian cancer in the peritoneal cavity. PMID- 15283467 TI - Antiproliferative effect of trichostatin A and HC-toxin in T47D human breast cancer cells. AB - Histone deacetylase inhibitors are new class of chemotherapeutic drugs able to induce tumor cell apoptosis and/or cell cycle arrest. Trichostatin A, an antifungal antibiotic, and HC-toxin are potent and specific inhibitors of histone deacetylase activity. In this study, we have examined the antiproliferative activities of trichostatin A and HC-toxin in estrogen receptor positive human breast cancer, T47D cells. Both trichostatin A and HC-toxin showed potent antiproliferative efficacy and cell cycle arrest at G2/M in T47D human breast cancer cells in a dose-dependent manner. Trichostatin A caused potent apoptosis of T47D human breast cancer cells and trichostatin A-induced apoptosis might be involved in an increase of caspase-3/7 activity. HC-toxin evoked apoptosis of T47D cells and HC-toxin induced apoptosis might not be mediated through direct increase in caspase-3/7 activity. We have identified potent activities of antiproliferation, apoptosis, and cell cycle arrest of trichostatin A and HC toxin in estrogen receptor positive human breast cancer cell line T47D. PMID- 15283468 TI - The acute effect of trimetazidine on the high frequency fatigue in the isolated rat diaphragm muscle. AB - The objective of this study was to determine the acute effect of trimetazidine (TMZ) on the pre-fatigue, fatigue and post-fatigue contractile characteristics and tension-frequency relationships of isolated rat diaphragm muscle. Muscle strips were taken from the ventral-costal aspects of the diaphragm muscle of rats killed by decapitation. The muscle strips were suspended in organ baths containing Krebs solution, with a gas mixture of 95% O2 and 5% CO2 at 37 degrees C and pH 7.35-7.45. After determining the thermoregulation and optimum muscle length the muscles were subjected to direct supramaximal stimulation with 0.05 Hz frequency square pulses for periods of 0.5 msec to obtain control values. After adding 5 x 10(-6) and 5 x 10(-5) M trimetazidine solution to the respective bath media, the contractile parameters of the muscles were recorded. The contractile parameters were also recorded for both the trimetazidine and trimetazidine-free media after application of the high frequency fatigue protocols. Later, the tension-frequency relationship was determined by applying stimulating pulses of 10, 20, 50 and 100 Hz to the muscle strips. Whilst the twitch tension obtained from the 5 x 10(-6) and 5 x 10(-5) M trimetazidine media showed numerical increases compared to that of the controls, these were not statistically significant (p>0.05). The contraction time exhibited a dose dependent increase (p<0.001), whilst the contraction and relaxation rates did not differ significantly. The isometric contraction forces obtained with the different stimulating frequencies showed a significant increase in the tetanic contraction only at 100 Hz (p<0.05). A comparison of the pre- and post-fatigue twitch tensions in the trimetazidine media showed the post- fatigue twitch tensions to be significantly higher than those of the pre-fatigue contraction forces (p<0.05). In the 5 x 10(-6) and 5 x 10(-5) M trimetazidine media the increases in the post-fatigue contraction force were 22 and 30%, respectively. These results demonstrated that in isolated rat diaphragm muscle, TMZ significantly limited the mechanical performance decrease during fatigue. It is our opinion that trimetazidine contributed to the observed fatigue tolerance by eliminating the factors of fatigue, due to preservation of intracellular calcium homeostasis, provision of the ATP energy levels needed by ATPase dependent pumps and especially by keeping the intracellular pH within certain limits. PMID- 15283470 TI - Water soluble cyclosporine monomethoxy poly(ethyleneglycol) conjugates as potential prodrugs. AB - The highly water-soluble monomethoxypoly(ethyleneglycol) (mPEG) prodrugs of cyclosporin A (CsA) were synthesized. These prodrugs were prepared by initially preparing intermediate in the form of carbonate at the 3'-positions of CsA with chloromethyl chloroformate, in the presence of a base to provide a 3'-carbonated CsA intermediate. Reaction of the CsA intermediate with mPEG derivative in the presence of a base provides the desired water-soluble prodrugs. As a model, we chose molecular weight 5 kDa mPEG in the reaction with CsA to give water soluble prodrugs. To prove that the prodrug is decomposed in the body to produce CsA, the enzymatic hydrolysis test was conducted using human liver homogenate at 37 degrees C. The prodrug was decomposed in human liver homogenate to produce the active material, CsA, and the hydrolysis half-life (t(1/2)) of the prodrug, KI 306 was 2.2 minutes at 37 degrees C. However, a demonstration of non-enzymatic conversion in pH 7.4 phosphate buffer was provided by the fact that the half-life (t(1/2)) is 21 hours at 37 degrees C. The hydrolysis test in rat whole blood was also conducted. The hydrolysis was seen with half-life (t(1/2)) of about 9.9, 65.0, 14.2, 3.4, 2.1 9.5, and 1.6 minutes for KI-306, 309, 312, 313, 315, 316, and 317, respectively. This is the ideal for CsA prodrug. The pharmacokinetic study of the prodrug, KI-306, in comparison to the commercial product (Sandimmune Neoral Solution) was also carried out after single oral dose. Each rat received 7 mg/kg of CsA equivalent dose. Especially, the prodrug KI-306 exhibits higher AUC and Cmax than the conventional Neoral. The AUC and Cmax were increased nearly 1.5 fold. The kinetic value was also seen with Tmax of about 1.43 and 2.44 hours for KI-306 and Neoral, respectively. PMID- 15283469 TI - Effects of dopamine HCl on structural parameters of bovine brain membranes. AB - Fluorescence probes located in different membrane regions were used to evaluate the effect of dopamine.HCl on the structural parameters (transbilayer lateral mobility, annular lipid fluidity, protein distribution, and thickness of the lipid bilayer) of synaptosomal plasma membrane vesicles (SPMV), which were obtained from the bovine cerebral cortex. An experimental procedure was used based on selective quenching of 1,3-di(1-pyrenyl)propane (Py-3-Py) by trinitrophenyl groups, and radiationless energy transfer from the tryptophan of membrane proteins to Py-3-Py and energy transfer from Py-3-Py monomers to 1 anilinonaphthalene-8-sulfonic acid (ANS) was also utilized. Dopamine.HCl increased both the bulk lateral mobility and annular lipid fluidity, and it had a greater fluidizing effect on the inner monolayer than on the outer monolayer. Furthermore, the drug had a clustering effect on membrane proteins. PMID- 15283471 TI - Preparation and characterization of enrofloxacin/carbopol complex in aqueous solution. AB - Since the bitter taste of enrofloxacin apparently limit the patient compliance in the oral formulations of the antibacterial agent, the masking of the taste is essential for the improvement of the therapeutic effectiveness. Therefore, this study was carried out to examine the feasibility of taste masking of enrofloxacin by the retardation of its dissolution rate using the formation of complex between the drug and Carbopol. The complexation between Carbopol and enrofloxacin was confirmed by turbidity, UV spectrophotometry, wide angle X-ray diffraction, and differential scanning calorimetry. The enrofloxacin content in the complexes was 34% (Carbo-enrofloxacin complex I) and 57% (Carbo-enrofloxacin complex II) depending on the preparation method. The dissolution rate of enrofloxacin from the complex increased as the pH was reduced. The dissolution rate of enrofloxacin from the Carbo-enrofloxacin complex I was significantly lower than that of the enrofloxacin powder. Therefore, these observations suggest that Carbo enrofloxacin complex I can be used to mask the taste of enrofloxacin. PMID- 15283472 TI - HPLC determination and pharmacokinetics of endogenous acetyl-L-carnitine (ALC) in human volunteers orally administered a single dose of ALC. AB - Acetyl-L-carnitine (ALC), a naturally occurring endogenous compound, has been shown to improve the cognitive performance of patients with senile dementia Alzheimer's type, and to be involved in cholinergic neurotransmission. Because ALC is an endogenous compound, validation of the analytical methods of ALC in the biological fluids is very important and difficult. This study was presented validation and correction for plasma ALC concentrations and pharmacokinetics after oral administration of ALC to human volunteers. ALC concentrations in human plasma were corrected by subtracting the concentration of blank plasma from each sample. Precision and accuracy (bias %) for uncorrected ALC concentrations were below 2.6 and 6.5% for intra-days, and 4.0 and 9.4% for inter-days, respectively. Precision and accuracy (bias %) for corrected ALC concentrations were below 10.9 and 6.0% for intra-days, and 10.5 and 16.9% for inter-days, respectively. Quantitation limit was 0.1 microg/mL. After oral administration of a 500 mg ALC tablet to 8 healthy volunteers, the principle pharmacokinetic parameters were 4.2 h of the half-life (t(1/2,beta)), the area under the curve (AUC(0-8)) of 9.88 microg.h/mL, and 3.1 h of the time (Tmax) to reach Cmax. This study first describes the pharmacokinetic study after oral administration of a single dose of ALC in human volunteers. PMID- 15283473 TI - Lost in time: a historical frame, elementary processing units and the 3-second window. AB - The main topics of time and timing in psychology, cognitive neuroscience and biology have been formulated already in the nineteenth century. Unfortunately, time and timing as a challenging topic has been put to rest for quite some time, but has become a central issue again during the last years. It has become clear, that perceptual or cognitive processes can only be understood if the dimension of time is taken more seriously. The reduction of complexity in neuronal systems is for instance, achieved by temporal integration mechanisms which are independent of the content of a percept or a cognitive act but are presemantical operations. It is essential to distinguish between content functions and logistical functions that provide presemantically defined temporal frames for mental activity. PMID- 15283474 TI - Decision processes in models of timing. AB - The article discusses the role played by decision mechanisms in the leading model of timing, scalar expectancy theory. Examples of the roles played by decision mechanisms in explanations of behaviour on temporal generalization and bisection are presented. Decision mechanisms for different timing tasks often have a common form (thresholded normalized difference, TND), where differences between durations are "normalized" (i.e., divided) by another duration value, then compared with a threshold. The TND principle provides a rule for both similarity and identity judgements of duration. The role of threshold mechanisms in timing is discussed, and it is shown that some procedural manipulations appear to specifically alter threshold values. Finally, problems in modelling the decision processes involved in verbal estimation are discussed. PMID- 15283475 TI - Prospective and retrospective duration judgments: an executive-control perspective. AB - Most theorists propose that when a person is aware that a duration judgment must be made (prospective paradigm), experienced duration depends on attention to temporal information, which competes with attention to nontemporal information. When a person is not aware that a duration judgment must be made until later (retrospective paradigm), remembered duration depends on incidental memory for temporal information. In the present article we describe two experiments in which durations involved with high-level, executive-control functions were judged either prospectively or retrospectively. In one experiment, the executive function involved resolving syntactic ambiguity in reading. In another experiment, it involved controlling the switching between tasks. In both experiments, there was a unique cost to the operation of control high-level, executive functions which was manifested by prospective reproductions shortening a finding that supports an attentional model of prospective timing. In addition, activation of executive functions produced contextual changes that were encoded in memory and resulted in longer retrospective reproductions, a finding that supports a contextual-change model of retrospective timing. Thus, different cognitive processes underlie prospective and retrospective timing. Recent findings obtained by some brain researchers also support these conclusions. PMID- 15283476 TI - The neural correlates of cognitive time management: a review. AB - Cognitive time management is an important aspect of human behaviour and cognition that has so far been understudied. Functional imaging studies in recent years have tried to identify the neural correlates of several timing functions, ranging from simple motor tapping to higher cognitive time estimation functions. Several regions of the frontal lobes, in particular dorsolateral prefrontal cortex (DLPFC), inferior prefrontal cortex (IFC), anterior cingulate gyrus (ACG) and the supplementary motor area (SMA), alongside non-frontal brain regions such as the inferior parietal lobes, the cerebellum and the basal ganglia have been found to be involved in tasks of motor timing and time estimation. In this paper we review and discuss the involvement of these brain regions in different tasks of cognitive time management, illustrating it with own findings on motor timing and time perception tasks using functional magnetic resonance imaging (fMRI). The review shows that the same brain regions are involved in both motor timing and time estimation, suggesting that both functions are probably inseparable and mediated by common neural networks. PMID- 15283477 TI - Time and language--critical remarks on diagnosis and training methods of temporal order judgment. AB - A lively discussion concerning the causal relation between auditory temporal processing and phoneme identification has evolved over the last decades. Subjects with language impairments not only show deficits in the identification of stop consonant vowel syllables, but also have problems detecting the temporal order of acoustic stimuli. Recently published studies claim that an improvement in phoneme discrimination can be achieved through the training of temporal-processing abilities. Critical assessment of these studies often reveals the following weaknesses: first, the diagnostic and training methods vary between studies, which makes comparisons difficult. Second, usually only mean differences between groups or before/after treatment are presented. The success in diagnosis and training of individuals or subgroups is not documented. Third, only few diagnostic measures employed have been tested for reliability. Furthermore, the tests have not been designed according to modern psychometric methods. Fourth, several training modules are used in parallel. The effects of temporal-processing training cannot be isolated. Possible approaches for detecting the possible causal relation between the time and the language domain are discussed. PMID- 15283478 TI - Individual differences in temporal information processing in humans. AB - This article reviews some of our investigations concerning individual differences in temporal information processing. Two different levels of temporal information processing are discussed, namely the low-frequency (i.e., a few seconds time range) and the high-frequency processing level (i.e., some tens of milliseconds range) of temporal information with respect to various experimental paradigms. Evidence has been obtained indicating that the processing of temporal information on these two levels can be influenced by various subject-related factors, out of which age, gender, developmental disorders, auditory experience and localisation of damage in the brain seem to be the most significant. PMID- 15283479 TI - Time perception depends on accurate clock mechanisms as well as unimpaired attention and memory processes. AB - We report a series of studies aimed at characterizing the relationships between duration judgments and slowing down of the internal clock, attention and memory deficits. Different groups of participants (elderly people, patients with Parkinson's disease, patients with severe traumatic brain injury, and patients with temporal lobe lesions) performed a duration reproduction task and a duration production task in two conditions: a control counting condition and a concurrent reading condition. Participants were also administered reaction time tasks, tapping tasks, and a battery of attention and memory tests. The results allow us to characterize the relationships between cognitive deficits and impaired duration reproductions and productions in each group. Moreover, results as a whole clarify the respective weight of processing speed, attention and memory in both tasks, and allow better insight into the theoretical models of psychological time. PMID- 15283480 TI - Measurement of temporal-order judgment in children. AB - Abnormal auditory temporal processing might be an underlying deficit in language disabilities. The auditory temporal-order threshold, one measure for temporal processing abilities, is defined as the shortest time interval between two acoustic events necessary for a person to be able to identify the correct temporal order. In our study, we examined the reliability of the auditory temporal-order threshold during a one-week period and over a time interval of four months in normally developing children aged 5 to 11 years. The results of our method show that children younger than 7 years have difficulties performing the task successfully. The reliability of the assessment of the temporal-order threshold during a period of one week is only moderate, and its stability over a time interval of four months is low. The results show that auditory-order thresholds in children have to be treated with caution. A high temporal-order threshold does not necessarily predict disabilities in temporal processing. PMID- 15283481 TI - Reproduction of auditory and visual standards in monochannel cochlear implant users. AB - The temporal reproduction of standard durations ranging from 1 to 9 seconds was investigated in monochannel cochlear implant (CI) users and in normally hearing subjects for the auditory and visual modality. The results showed that the pattern of performance in patients depended on their level of auditory comprehension. Results for CI users, who displayed relatively good auditory comprehension, did not differ from that of normally hearing subjects for both modalities. Patients with poor auditory comprehension significantly overestimated shorter auditory standards (1, 1.5 and 2.5 s), compared to both patients with good comprehension and controls. For the visual modality the between-group comparisons were not significant. These deficits in the reproduction of auditory standards were explained in accordance with both the attentional-gate model and the role of working memory in prospective time judgment. The impairments described above can influence the functioning of the temporal integration mechanism that is crucial for auditory speech comprehension on the level of words and phrases. We postulate that the deficits in time reproduction of short standards may be one of the possible reasons for poor speech understanding in monochannel CI users. PMID- 15283482 TI - Aging and the time course of inhibition of return in a static environment. AB - Age-related differences on the time course of inhibition of return (IOR), a phenomenon that refers to a slowed response time for targets appearing at a previously attended location, were examined in 30 young and 30 elderly adults. Stimulus onset asynchronies (SOAs) between peripheral cues and targets were systematically manipulated on a detection task with a double-cue procedure to capture the onset and offset of IOR. Results show that IOR in elderly people developed 50 ms later as compared to young adults, at an approximately 200 ms cue target interval. The magnitude of IOR for elderly people was also weaker than that for young adults during short SOAs. Similar magnitude and dissipation of IOR at an approximately 3.5 s cue-target interval during long SOAs were observed for both young and elderly people. Possible reasons underlying the age effects on the time course of IOR and the involvement of temporal processing mechanisms are discussed. PMID- 15283483 TI - Two types of anticipation in synchronization tapping. AB - The time perception mechanism in anticipatory timing control was investigated in a synchronization tapping task. An especially negative asynchrony phenomenon in which the tap onset precedes the stimulus onset was used as an anticipatory response. In this experiment, to clarify the effects of higher brain functions, such as attention, a dual-task method was applied and a word memory task was used as a secondary task. The results revealed two types of anticipatory mechanisms from the standpoint of attentional resources involved in time perception. One is the anticipatory tapping that is influenced by attention and seen in the interstimulus-onset interval (ISI) range of 1,800 to 3,600 ms. In this region, the magnitude of synchronization error (SE) between tap onset and stimulus onset was scaled by the ISI. The other is the automatic anticipation that is not affected by attention and is seen in the 450 to 1,500 ms range. SE in this region was constant and independent of the ISI. Accordingly, this anticipatory timing mechanism in synchronous tapping is thought to be a dual process including the attention processing of temporal information and the embodied automatic anticipation. PMID- 15283484 TI - Visual perception in space and time--mapping the visual field of temporal resolution. AB - To characterize temporal aspects of information processing in the human visual field, we studied the topographical distribution of temporal and non-temporal performance parameters in 95 normally sighted subjects. Visual field maps of double-pulse resolution thresholds (DPR) (the minimum detectable temporal gap between two light stimuli) and simple visual reaction times (RT) (measuring the speed of reaction to a light stimulus) were compared to maps of luminance thresholds determined by standard perimetry. Thus, for the first time, the topography of a visual variable without temporal constraints (perimetry) could be compared to visual variables in the temporal domain, with (RT) and without (DPR) motor reaction. The goal of the study was to obtain and to describe the pattern of co-variation of performance indicators. In all three measures, performance was best in the central visual field and dropped significantly towards the periphery. Although the correlation between DPR and RT was significant, shared variance was low, and we observed large topographical differences between these two temporal performance variables. In contrast, DPR and perimetric thresholds correlated more substantially, and visual field maps were similar. The Gestalt of DPR maps shares characteristics of basic visual processing (e.g., light sensitivity), but it also reflects top-down influences, i.e., from spatial attention. Although the correlation between DPR and RT suggests common characteristics between these two temporal variables, the topographic distributions reveal significant differences, indicating separate underlying processing mechanisms. PMID- 15283485 TI - Long-term quality of life measures after functional endoscopic sinus surgery. AB - BACKGROUND: Chronic rhinosinusitis (CRS) is a common disease that has a significant impact on quality of life (QOL). The aim of this study was to evaluate the longer-term effects of combined medical and surgical therapy for CRS on overall health status and QOL. METHODS: We used a prospective study that utilized the Short-Form 36 Survey at baseline presentation and at a mean time of 3 years post-functional endoscopic sinus surgery to assess the general health status of patients who presented for their initial visitfrom 1996 to 1998. Of the 200 randomly selected patients, 150 respondents completed follow-up surveys (a 75% response rate). RESULTS: Eighty-nine (59.3%) women and 61 (40.7%) men were included in the study. Baseline QOL scores indicated significant differences between patients with CRS and published norms in 6/8 subscale parameters (role physical, bodily pain, general health, social function, vitality, and mental health). Significant improvement in all six categories was maintained at the end of the study period (p < 0.05) with QOL scores within limits of published norms for the general population. CONCLUSION: Our data indicate that functional endoscopic sinus surgery, combined with appropriate postoperative care, is effective at maintaining a significant improvement in the overall general health status of patients for at least 3 years after surgical intervention and that the overall scores return to a range of normative values for the general population. PMID- 15283486 TI - Anosmia after intranasal zinc gluconate use. AB - BACKGROUND: Zinc is an essential mineral. Beneficial zinc absorption takes place via enteral, parenteral, or cutaneous routes. However, direct application to the olfactory epithelium has been reported to cause loss of smell. Recently, intranasal zinc gluconate has been recommended as a treatment for the common cold. Severe posttreatment hyposmia and anosmia have been observed. METHODS: The case report of a typical patient is presented and analyzed in detail, followed by a series of patients with severe hyposmia or anosmia after the use of intranasal zinc gluconate. RESULTS: Although interindividual variation in drug response and drug effect is apparent, the severe hyposmia or anosmia appears to be long lasting or permanent in some cases. The mechanism of olfactory loss is thought to be the direct action of the divalent zinc ion on the olfactory receptor cell. CONCLUSIONS: Zinc ions are toxic to olfactory epithelium. Reports of severe hyposmia with parosmia or anosmia have occurred after intranasal use of zinc gluconate. PMID- 15283487 TI - The nasal valve: a review of the anatomy, imaging, and physiology. AB - BACKGROUND: The nasal valve region has remained difficult to define in clinical practice in part because of lack of integration between physiological data and anatomic-surgical findings. METHODS: In this review, we summarize the anatomic, physiological, and imaging data regarding this complex area of airflow regulation. RESULTS: There is no singular resistive focus or singular valve structure to explain all of the reported findings. CONCLUSION: We conclude that there is a nasal valve region that begins approximately at the limen nasi and continues for several millimeters within the nasal cavum beyond the piriform aperture. Intranasal pressure measurements reflect distributed resistance across this nasal valve region. The geometry and anatomic constitution of the nasal valve region change greatly from its entrance to its distal aspect. To refer consistently to the component portions of the nasal valve region, we suggest the terms cartilaginous valve segment and bony valve segment for use in reporting future studies. PMID- 15283488 TI - Multi-use Venturi nasal atomizer contamination in a clinical rhinologic practice. AB - INTRODUCTION: Cross-contamination of Venturi atomizers has been reported. METHODS: In phase I, 16 atomizers were sterilized and refilled with either 1% lidocaine or 0.1% Tyzine (day 0). During phase II, atomizers were wiped with isopropyl alcohol wipes between uses. In both phases, on days 7, 14, 21, and 28, the contents of the atomizer were sprayed onto two culture media. If a culture was found positive, cultures from the nozzle, lumen, and solution were taken and cultured in depth. RESULTS: Twelve lidocaine bottles and three Tyzine bottles were initially positive. There were more positive cultures from lidocaine bottles than Tyzine bottles at 2 weeks (p = 0.02). After wiping with isopropyl alcohol, contamination was significantly reduced in the lidocaine bottles at 2 weeks (p = 0.02). CONCLUSION: Microbial contamination of questionable clinical significance may occur with nasal atomizers. Regardless of this significance, wiping the devices with isopropyl alcohol can eliminate microbial growth for a 2-week interval. PMID- 15283490 TI - Role of nasal endoscopy in patients undergoing endoscopic sinus surgery. AB - BACKGROUND: The goal of this study was to determine the role of nasal endoscopy in patients undergoing endoscopic sinus surgery and to examine the correlation between nasal endoscopy and pre- and postoperative computed tomography (CT) grade, symptom scores, and the patients' asthma status. METHODS: A retrospective analysis of prospectively collected data from 254 consecutive patients treated with endoscopic sinus surgery was performed from 1999 to 2002. There were 131 men and 123 women, with a mean age of 48 years. All patients had at least 1-year follow-up. All patients received preoperative CT scans, which were graded as per Lund-MacKay, completed the sinonasal outcome test 20 questionnaires, and underwent endoscopic nasal examination at both the preoperative and 1-year postoperative visits. The correlation between the endoscopy score, CT grade, and sinonasal outcome test 20 findings was calculated. RESULTS: Nasal endoscopy indicated a statistically significant correlation with preoperative CT grade in this patient population. There was no correlation between endoscopic findings and patient symptom scores. Nasal endoscopy in patients with asthma (n = 47) did not correlate with either CT grade or patient symptom scores. In patients without asthma (n = 207), nasal endoscopy correlated with CT grade but not with symptom scores. CONCLUSION: Nasal endoscopy findings do not correlate with symptom scores in patients undergoing endoscopic sinus surgery for chronic rhinosinusitis. Nasal endoscopy findings correlated with CT grade in patients without asthma, but no such correlation was found in patients with asthma. PMID- 15283489 TI - Resection of the inferior superior turbinate: does it affect olfactory ability or contain olfactory neuronal tissue? AB - BACKGROUND: One approach to the sphenoid sinus involves resection of the inferior portion of the superior turbinate. There is general agreement from anatomic investigations that this area contains olfactory mucosa. This study will determine if olfactory tissue can be found in the superior turbinate mucosa of patients with chronic sphenoiditis and what effect its removal has on a patient's olfactory ability. METHODS: The inferior one-third of the superior turbinate removed during endoscopic sphenoidotomy was stained with olfactory marker protein antibody, a marker for mature olfactory tissue. The specimens were graded for content of olfactory neuronal elements. All patients underwent uninasal 12-item smell identification testing before surgery and at least 3 weeks after surgery. RESULTS: Fifty-five superior turbinate samples were taken from 31 patients. Nine (16%) of 55 samples contained olfactory neuronal elements that stained with olfactory marker protein. When comparing the pre- and postoperative smell test results, 52% of the nostrils had no more than a one-item change, 35% of the nostrils had a more than one-item improvement, and only 12% had more than a one item loss. None of the nostrils with a loss of olfactory ability after the surgery showed olfactory neuronal elements in their superior turbinate specimens. CONCLUSION: There is olfactory mucosa in approximately one-sixth of the superior turbinate specimens removed during the endoscopic transethmoidal sphenoidotomy procedure. Although 12% of the patients had a loss of olfactory ability in this study, none of the loss could be attributed to excision of olfactory tissue. PMID- 15283491 TI - Histopathological features of nasal polyps with asthma association: an immunohistochemical study. AB - BACKGROUND: Myofibroblasts are related to airway remodeling and may play an important role in the pathogenesis of the histological features of nasal polyps associated with asthma. The aim of this study was to analyze and compare the eosinophilic cationic protein, transforming growth factor (TGF) beta, and myofibroblasts in the nasal polyps of patients with chronic sinusitis associated with and without asthma. METHODS: Nasal polyp samples were obtained during endoscopic sinus surgery and were classified into asthma and nonasthma groups. Immunohistochemistry was performed using antibodies against activated eosinophils, TGF-beta, and myofibroblasts. RESULTS: The asthma group showed an increased number of activated eosinophils, TGF-beta, and myofibroblasts compared with the nonasthma and control groups. We found no correlation of asthma and aspirin intolerance with the immunohistochemical findings. CONCLUSION: The increased number of myofibroblasts in the nasal polyps of the asthma group may be responsible for the extracellular matrix accumulation, polyp formation, and polyp recurrence. PMID- 15283492 TI - Quantitative computer-aided computed tomography analysis of sphenoid sinus anatomical relationships. AB - BACKGROUND: This study describes a novel computer-generated anatomic symmetry plane as a framework for the quantitative description of sphenoid sinus anatomy. The aim of this study was to (1) determine relationships and distances between a midline sphenoid reference point (called the central sphenoid point [CSP]) and lateral sphenoid wall structures and (2) assess the incidence of anterior clinoid process (ACP) pneumatization and pterygoid recess (PR) pneumatization. METHODS: Axial computed tomography (CT) scans (1-mm slice thickness) were obtained on a VolumeZoom CT scanner (Siemens Medical, Erlangen, Germany). Mathematically derived anatomic symmetry planes were created using custom postprocessing software. A standardized review of each CT scan using surgical planning software (CBYON Suite version 2.6; CBYON, Mountain View, CA) was performed. The CSP was defined as a reference point in the midline sagittal plane at the intersection of the vertical sellar face and the horizontal sellar floor. RESULTS: A total of 128 sides in 64 cadaveric specimens were available for review. The incidences of ACP pneumatization and PR pneumatization were 23.4 and 37.5%. The mean distances from the CSP to the left optic canal midpoint, the left ACP entrance point, and the left PR lateral wall were 17.2, 15.6, and 27.6 mm, respectively. The corresponding distances from the CSP on the right side were 17.3, 15.8, and 28.0 mm, respectively. Measurements from the maxillary spine to the optic canal midpoint, ACP entrance point, and PR lateral wall on each side were performed also. CONCLUSION: This approach provides both quantitative and qualitative understanding of sphenoid osteology and may be coupled with intraoperative surgical navigation to reduce the risks of sphenoid surgery. Both PR and ACP pneumatization are surprisingly common. Because the CSP-derived relationships may be referenced during endoscopic surgical navigation, they may provide greater clinical utility than traditional alternatives. This paradigm may facilitate a greater understanding of sphenoid anatomy and enhance surgical safety and precision. PMID- 15283493 TI - Congenital nasal piriform aperture stenosis or bony inlet stenosis: a report of three cases. AB - BACKGROUND: Congenital nasal piriform aperture stenosis is an uncommon disorder causing nasal obstruction in neonates. Findings on computed tomography (CT) greatly facilitate the diagnosis. METHODS: Three cases of congenital nasal piriform aperture stenosis are reported, with an emphasis on CT findings. CONCLUSIONS: At birth, a diameter of the piriform aperture at the level of the inferior nasal meatus of > or =5 mm on CT may indicate a good chance of success using conservative treatment. PMID- 15283494 TI - The effect of the modified endoscopic Lothrop procedure on the mucociliary clearance of the frontal sinus in an animal model. AB - BACKGROUND: The surgical management of recalcitrant frontal sinus disease has been a dilemma for otolaryngologists for many years. Although the osteoplastic flap with obliteration has been the gold standard of treatment for years, the modified endoscopic Lothrop (MEL) procedure recently has been advocated as an alternative. However, little is known about the effect of this procedure on the mucociliary drainage of the frontal sinuses postoperatively and this animal study addresses this issue. METHODS: Fourteen sheep underwent the MEL procedure. The sheep were randomized regarding the use of postoperative irrigation via minitrephines. Each sheep had a nuclear medicine gamma-scintigraphy frontal sinus clearance study via minitrephines performed on each frontal sinus preoperatively and then 3 months postoperatively. Then, the results of these studies were compared. RESULTS: The scans revealed a trend toward faster clearance times postoperatively. However, this decrease was not statistically significant. Importantly, there was no trend or significant increase in clearance times postoperatively. Also, the use of postoperative irrigation was associated with a nonsignificant trend toward faster clearance times postoperatively. CONCLUSION: The MEL procedure has no adverse effects on the mucociliary clearance of the frontal sinus at 3 months postoperatively. Irrigation of the frontal sinus in the immediate postoperative period showed a trend toward improved postoperative mucociliary function at 3 months. PMID- 15283495 TI - Guidelines for managing life-threatening food allergies in Massachusetts schools. AB - During the past decade, prevalence of food allergies among children increased. Caring for children with life-threatening food allergies has become a major challenge for school personnel. Prior to 2002, Massachusetts did not provide clear guidelines to assist schools in providing a safe environment for these children and preparing for an emergency response to unintended allergic reactions. In 2001, the Asthma and Allergy Foundation of America/New England Chapter, Massachusetts Department of Education, Massachusetts Department of Public Health, Massachusetts School Nurse Organization, parents, and other professional organizations forged a successful collaboration to develop guidelines for managing life- threatening food allergies in schools. The guidelines assist schools by providing information on food allergies and anaphylaxis, emphasizing the need for team planning and development of an individualized health care plan, giving guidance on strategies to prevent accidental exposure to specific allergens in school settings, and offering information on emergency responses should unintended exposures occur. The collaborative process for developing the guidelines, which continued during the distribution and implementation phases, set a tone for successful multidisciplinary teamwork in local schools. PMID- 15283496 TI - Chronic disease medication administration rates in a public school system. AB - Anecdotal reports suggest school nurses and staff treat increasing numbers of public school students with chronic diseases. However, professionals know little about actual disease burden in schools. This study measured prevalence of chronic disease medication administration rates in a large, urban midwestern school district. Data from daily medication logs were recorded by school nurses during a single week. Medications and administrations were sorted by disease type. Prevalence rates were calculated for six chronic diseases: asthma, diabetes, seizures, attention deficit/hyperactivity disorder, other mental/behavioral disorders, and other diseases/conditions. Separate rates stratified by school grade, poverty level, and type of school were calculated. Overall, 3.12% of students received medication for chronic diseases, including 2.13% for psychiatric/mental disorders and 1.91% for attention deficit/hyperactivity disorder alone. These rates were lower than estimates from other states. Factors that contributed to this finding are reviewed. PMID- 15283497 TI - Adolescent and parent assessments of diabetes mellitus management at school. AB - This study explored opinions, concerns, and recommendations regarding care of Type 1 diabetes in schools. Thirty adolescent females and their parents participated in semi- structured, individual interviews that were audiotaped, transcribed, coded, and qualitatively analyzed. Responses emerged in three categories: knowledge/training of school staff; foods offered/available at school; and school rules. Participants expressed concerns that school personnel, particularly classroom teachers, possess limited knowledge of diabetes; that healthy food/beverage options are limited in the cafeteria, vending machines, and classrooms; and that school rules impede self-care of diabetes. Implications for enhancing diabetes management at school are noted. PMID- 15283498 TI - Evaluation of the National School Health Coordinator Leadership Institute. AB - In 1999 the American Cancer Society (ACS) launched the National School Health Coordinator Leadership Institute, a groundbreaking initiative designed to enhance and invigorate school health in the nation's schools by training individual school health coordinators to act as change agents. The Institute consisted of three, week-long summer training sessions, and three, shorter midyear "booster" sessions, with the various sessions designed to assist one cohort of participants (n = 50) to build leadership capacities to coordinate school health. This evaluation examined the effects of the Institute as measured through a panel survey of participants - school health coordinators or their equivalents - over an 18-month period. Findings suggest that substantial progress was made in institutionalizing the school health coordination function in the target school districts. Specifically, in contrast to when the training began, clear, written position descriptions are common among program participants, most have functioning school health councils in their school districts, and program trainees appear to be moving aggressively to enhance the infrastructure for school health coordination through planning, setting priorities, and assessing needs. Program trainees report spending increased time on school health coordination, and they are active in spreading the word about coordinated school health programs. Trainees viewed the Institute as relevant to their needs, though not all aspects of the program are viewed as equally useful. As possible shortcomings, the training needed more emphasis on funding for school health coordination, and a low level of evaluation skills existed among coordinators. PMID- 15283500 TI - Perspectives. Lower premiums? Medicare PPOs? Don't bet on it, analysts say. PMID- 15283501 TI - Four predictions for the future of health care philanthropy: what lies ahead? PMID- 15283499 TI - Attempted suicide and associated health risk behaviors among Native American high school students. AB - Suicide represents the second-leading cause of death among American Indian/Alaska Native (AI/AN) youth aged 15-24 years. Data from the 2001 Bureau of Indian Affairs (BIA) Youth Risk Behavior Survey were used to examine the association between attempted suicide among high school students and unintentional injury and violence behaviors, sexual risk behaviors, tobacco use, and alcohol and other drug use. The study included students in BIA-funded high schools with 10 or more students enrolled in grades 9-12. Overall, 16% of BIA high school students attempted suicide one or more times in the 12 months preceding the survey. Females and males who attempted suicide were more likely than females and males who did not attempt suicide to engage in every risk behavior analyzed: unintentional injury and violence behaviors, sexual risk behaviors, tobacco use, and alcohol and other drug use. These data enable educators, school health professionals, and others who work with this population to better identify American Indian youth at risk for attempting suicide by recognizing the number and variety of health risk behaviors associated with attempted suicide. PMID- 15283502 TI - Present a worthy case--showcase your vision, not your need. PMID- 15283503 TI - A CEO's perspective. PMID- 15283504 TI - The legacy of AHP continues. PMID- 15283505 TI - A perspective on hospital foundation amalgamations. PMID- 15283506 TI - "Wouldn't it be neat to keep all this information on a PDA?". PMID- 15283507 TI - A better bedside manner. Hospital caregivers treat patients and complete clinical documentation with ease, thanks to point-of-care and mobile workstations. PMID- 15283508 TI - ISABEL at the helm. A web-based diagnosis system speeds clinical decisions for pediatric physicians. PMID- 15283509 TI - Calling for automation. Computerized call center helps school of medicine and a teaching affiliate hospital improve switchboard, information directory and after hours answering services. PMID- 15283510 TI - IT is coming to a physician near you. Proactivity on the part of health plans may be the biggest boon to physician adoption of it. PMID- 15283511 TI - Intervenability: another measure of health risk. By coupling predictive modeling with evidence-based medicine, health plans can identify patients who will benefit the most from care management intervention. PMID- 15283512 TI - Greater than the sum of the parts. Eighteen New York physician practices gain centralized patient information database with ASP-hosted system. PMID- 15283513 TI - Wireless hotlist. PMID- 15283514 TI - Targeting the highest-risk population to complement disease management. PMID- 15283515 TI - An ethicist's commentary on social concern for animal suffering immediately before death. PMID- 15283516 TI - Perioperative use of selective alpha-2 agonists and antagonists in small animals. AB - Alpha-2 agonists are the only single class of anesthetic drugs that induce reliable, dose-dependent sedation, analgesia, and muscle relaxation in dogs and cats. Used at low doses, as adjuncts to injectable and inhalational anesthetics, selective alpha-2 agonists dramatically reduce the amount of anesthetic drug required to induce and maintain anesthesia. This reduction in anesthetic requirements is achieved without significant depression of pulmonary function and with limited effects on cardiovascular function. Selective alpha-2 agonists can also be used postoperatively to potentiate the analgesic effects of opioids and other drugs. Given the nearly ideal pharmacodynamic profile and reversibility of alpha-2 agonists, these drugs will play a central role in balanced approaches to anesthesia and the management of perioperative pain in healthy dogs and cats. PMID- 15283517 TI - Comparison of medetomidine-ketamine and dexmedetomidine-ketamine anesthesia in golden-headed lion tamarins. AB - The cardiovascular, respiratory, and anesthetic effects of medetomidine-ketamine (20 microg/kg bodyweight [BW] and 10 mg/kg BW) (MK group) or dexmedetomidine ketamine (10 microg/kg BW and 10 mg/kg BW) (DK group) were studied in golden headed lion tamarins. Heart rate decreased after administration of both combinations; this reduction was statistically greater in the DK group than in the MK group after 15 and 45 minutes. Systolic arterial pressure decreased in a similar way in both groups, except at 15 minutes, when systolic arterial pressure was significantly lower in the DK group. Diastolic arterial pressure, mean arterial pressure, respiratory rate, and rectal temperature were progressively reduced in all groups. Sedation time was significantly shorter and anesthesia time was significantly longer in the DK group compared with MK group. Anesthetic quality and analgesia scores were significantly greater at 5 and 15 minutes in the DK group compared with the MK group. The administration of dexmedetomidine ketamine is as safe and effective as the administration of medetomidine-ketamine in tamarins. PMID- 15283518 TI - Prevalence of antibodies to bluetongue virus and Anaplasma marginale in Montana yearling cattle entering Alberta feedlots: Fall 2001. AB - A serologic survey was conducted in yearling cattle imported into Alberta feedlots from Montana during October 2001 to estimate the prevalence of antibodies to bluetongue virus (BTV) and Anaplasma marginale in Montana yearling cattle. The apparent prevalence of antibodies to BTV when the competitive enzyme linked immunosorbent assay (cELISA) was used was 0.37% (21/5608). Test positive cELISA samples were also all positive when tested by virus neutralization (VN) and they reacted to 1 or more BTV serotypes, including 2, 10, 11, 13, and 17. The apparent prevalence of antibodies to A. marginale when a recombinant cELISA (rcELISA) was used with a positive cutoff at 30% inhibition was 1.93% (108/5608). When the rcELISA positive cutoff was at 42% inhibition, the apparent prevalence was 0.73% (41/5608). After the reported sensitivity and specificity of the test had been accounted for, the A. marginale antibody results were consistent with a population that was either free of exposure or had a very low prevalence for A. marginale. PMID- 15283519 TI - Androgen insensitivity syndrome in a thoroughbred mare (64, XY--testicular feminization). AB - A Thoroughbred mare was presented for stallion-like behavior. Reproductive and ultrasonographic evaluation, testosterone assays, and karyotyping confirmed a diagnosis of androgen insensitivity syndrome (64, XY--testicular feminization). Surgery to remove abdominal testicles was successful in alleviating the behavioral abnormality. This condition is discussed with reference to the current literature. PMID- 15283520 TI - Hemoperitoneum caused by the rupture of a granulosa cell tumor in a Holstein heifer. AB - A case of a hemoperitoneum caused by the rupture of a granulosa cell tumor in a 9 month-old Holstein heifer is reported. Management of hemorrhagic shock in cattle is discussed. PMID- 15283521 TI - Challenging diagnosis--icterus associated with a single perforating duodenal ulcer after long-term nonsteroidal antiinflammatory drug administration in a dog. AB - A dog developed icterus, vomiting, and anorexia 2 wk after orthopedic surgery and treatment with meloxicam for approximately 1 y. Exploratory laparotomy revealed a single perforated duodenal ulcer. The most likely cause of the hyperbilirubinemia was intrahepatic cholestasis resulting from peritonitis associated with the perforation. PMID- 15283522 TI - Fatal trichuris spp. infection in a Holstein heifer persistently infected with bovine viral diarrhea virus. AB - Whipworms (Trichuris spp.) were identified in the colon of a recently purchased, 10-month-old dairy heifer that died suddenly. A skin test was positive for bovine viral diarrhea virus (BVDV). Signs of BVDV occurred in other heifers in the group, but fecal flotations were negative for whipworm eggs. PMID- 15283523 TI - Heat stroke in a great Pyrenees dog. AB - A 2-year-old, male Great Pyrenees presented with a history and clinical signs suggesting heat exhaustion. Treatment with intravenous fluids, antibiotics, and surface cooling was unsuccessful, and euthanasia was elected. Histological evaluation of the dog's tissues revealed lesions consistent with severe hyperthermia and shock. PMID- 15283524 TI - Saskatchewan. Unusual winter outbreak of anthrax. PMID- 15283525 TI - Practical bond considerations: guilt and the treatment process. PMID- 15283526 TI - Tibial plateau leveling osteotomy--part I. PMID- 15283527 TI - CVMA National Benchmarking Program--part 5. PMID- 15283528 TI - Palliative care: an opportunity for mental health professionals. PMID- 15283530 TI - Palliative care for families: remembering the hidden patients. AB - Families of patients receiving palliative care are profoundly affected by the challenges of the illness. They observe care that the patient receives, provide care for the patient, and receive support from health professionals in the form of information, counselling, or practical assistance. As they witness and participate in the patient's care, they judge the quality of care that the patient receives. They often see themselves as the patient's care advocates and may harbour regret and guilt if they believe that the patient did not have the best possible care. The illness experience profoundly affects family members' psychological and physical health; recognition of this has coined the term "hidden patients." This article briefly synthesizes empirical work that suggests how to best support families in a palliative care context. We discuss how to define the family, emphasizing a systems approach to family care. We describe the impact of the illness on the family in terms of family members' health, family communication issues, psychological issues, needs for information, physical care demands, and family costs of caring. PMID- 15283529 TI - Structured interview assessment of symptoms and concerns in palliative care. AB - OBJECTIVE: Assessment in palliative care requires a multidimensional review of physical symptoms and psychosocial concerns in a format appropriate for patients with advanced illness. In this study, we describe the initial development and validation of a structured interview for assessing common symptoms and concerns faced by terminally ill individuals. METHOD: We constructed a 13-item Structured Interview for Symptoms and Concerns (SISC) based on a review of end-of-life issues and administered it to 69 patients receiving palliative care for advanced cancer. Along with the interview, each participant completed visual analog scales (VAS) addressing the same constructs. Test-retest and interrater reliability were determined, as was the concordance between interview ratings and VAS scores. RESULTS: Overall, the interview items had excellent interrater reliability (intraclass correlations were > 0.90) and at least moderate temporal stability (test-retest correlations ranged from 0.50 to 0.90). Concurrent validity was evident in the good concordance between interview items and VAS measures (correlations were > 0.70). The SISC was also sensitive to individual differences between subgroups of participants who did or did not meet diagnostic criteria for anxiety or depressive disorders. CONCLUSIONS: This study demonstrates that structured interviews provide a reliable and valid approach to assessment in palliative care and may be an appropriate alternative for some research applications. PMID- 15283531 TI - Psychotherapeutic interventions at the end of life: a focus on meaning and spirituality. AB - Medical and psychological discourse on end-of-life care has steadily shifted over the years from focusing primarily on symptom control and pain management to incorporating more person-centred approaches to patient care. Such approaches underscore the significance of spirituality and meaning making as important resources for coping with emotional and existential suffering as one nears death. Though existential themes are omnipresent in end-of-life care, little has been written about their foundations or import for palliative care practitioners and patients in need. In this article, we explore the existential foundations of meaning and spirituality in light of terminal illness and palliative care. We discuss existential themes in terms of patients' awareness of death and search for meaning and practitioners' promotion of personal agency and responsibility as patients face life-and-death issues. Viktor Frankl's existential logotherapy is discussed in light of emerging psychotherapeutic interventions. Meaning-centred group therapy is one such novel modality that has successfully integrated themes of meaning and spirituality into end-of-life care. We further explore spiritual and existential themes through this meaning-oriented approach that encourages dying patients to find meaning and purpose in living until their death. PMID- 15283532 TI - Prevalence studies of substance-related disorders: a systematic review of the literature. AB - OBJECTIVE: To present the results of a systematic review of literature published between January 1, 1980, and December 31, 2000, that reports epidemiologic estimates of substance-related disorders. METHOD: We conducted a literature search of substance-related epidemiologic studies, using medline and HealthSTAR databases and applying a set of predetermined inclusion and exclusion criteria to identify relevant studies. We extracted and analyzed prevalence and incidence data for heterogeneity. RESULTS: A total of 19 prevalence studies of substance related disorders met inclusion criteria for this review. Heterogeneity analyses revealed significant variability across 1-year and lifetime prevalence of both alcohol and other substance use disorders. The corresponding 1-year and lifetime pooled rates were 6.6 per 100 and 13.2 per 100, respectively, for alcohol use disorders and 2.4 per 100 and 2.4 per 100, respectively, for other substance use disorders. We observed variability among countries and also among regions within the same country. In contrast to other drug problems, alcohol use disorders were substantially more common, were more likely to occur among male subjects, and were more likely to be associated with abuse symptoms. For other drugs, dependence was consistently more prevalent than abuse. CONCLUSIONS: Studies using rigorous and comparable methodologies report significant variability in rates of alcohol and other substance use disorders. These data suggest that different policies and regional practices are associated with variability in rates of disorders. Policy-makers and health planners require regular, regionally sensitive estimates of prevalence rates to respond effectively to unique patterns of need in their constituencies. PMID- 15283533 TI - Stress and psychological impact on SARS patients during the outbreak. AB - OBJECTIVE: To examine stress and psychological impact in severe acute respiratory syndrome (SARS) patients during the 2003 outbreak. SARS is a novel, highly infectious pneumonia, and its psychological impact is still unclear. METHOD: At the peak of the outbreak, SARS patients (n = 79) and healthy control subjects (n = 145) completed the Perceived Stress Scale (PSS) and documented a range of psychological responses. Groups were balanced for age, sex, education, and living circumstances. RESULTS: Stress was significantly higher in SARS patients than in healthy control subjects. Stress correlated significantly with negative psychological effects. Of SARS patients, 39% (n = 30) were infected health care workers; these individuals reported significantly more fatigue and worries about health than did other patients. Of patients, 25% (n = 20) requested psychological follow-up. CONCLUSIONS: General stress and negative psychological effects are increased in SARS patients, particularly among infected health care workers. This may increase the risk of mood and stress-related disorders. Functional impairment is apparent in the postrecovery phase. PMID- 15283535 TI - Schizophrenia: the quest for a minimum sense of identity to ward off delusional disorder. AB - OBJECTIVE: This study was designed to analyze the language of patients with schizophrenia exhibiting negative symptoms during a 3-month period. METHOD: The computer-assisted ALCESTE method was used to simultaneously analyze the subjects' oral behaviour and speech patterns at various levels. RESULTS: The tested subjects had very specific speech patterns. Most significantly, analysis of the underlying syntactic processes showed that the patients exhibited a sense of identity, however minimum, based on their own pathologies and on the surrounding world. In our previous study, no such characteristics were observed in the discourse of schizophrenia patients with delusions (exhibiting positive symptoms). This suggests that the minimum sense of identity that develops in patients with schizophrenia allows them to avoid positive symptoms. CONCLUSION: In studies of language production by subjects suffering from schizophrenia, it is necessary to distinguish between patients with positive symptoms and those with negative symptoms. The speech patterns of these 2 groups have to be analyzed separately, which has not been done previously, since the groups differ in too many respects. PMID- 15283534 TI - Psychological effects of the SARS outbreak in Hong Kong on high-risk health care workers. AB - OBJECTIVE: To quantify stress and the psychological impact of severe acute respiratory syndrome (SARS) on high-risk health care workers (HCWs). METHOD: We evaluated 271 HCWs from SARS units and 342 healthy control subjects, using the Perceived Stress Scale (PSS) to assess stress levels and a structured list of putative psychological effects of SARS to assess its psychological effects. Healthy control subjects were balanced for age, sex, education, parenthood, living circumstances, and lack of health care experience. RESULTS: Stress levels were raised in both groups (PSS = 18) but were not relatively increased in the HCWs. HCWs reported significantly more positive (94%, n = 256) and more negative psychological effects (89%, n = 241) from SARS than did control subjects. HCWs declared confidence in infection-control measures. CONCLUSIONS: In HCWs, adaptive responses to stress and the positive effects of infection control training may be protective in future outbreaks. Elevated stress in the population may be an important indicator of future psychiatric morbidity. PMID- 15283536 TI - Diabetes, tardive dyskinesia, parkinsonism, and akathisia in schizophrenia: a retrospective study applying 1998 diabetes health care guidelines to antipsychotic use. AB - BACKGROUND: This study examines the links among diabetes, tardive dyskinesia (TD), and other extrapyramidal symptoms (EPS) in schizophrenia outpatients treated with typical and atypical antipsychotics. OBJECTIVES: Using a retrospective chart review, we compared 30 schizophrenia patients with diabetes mellitus (DM) with 30 schizophrenia patients, matched for age and sex, with no DM. We compared prevalence and severity of parkinsonism, akathisia, TD, dystonia, and antipsychotic type (that is, typical vs atypical). RESULTS: We found no statistically significant differences between the DM group and the non-DM group prevalence and severity of EPS, including TD. CONCLUSION: We did not find DM and TD association to be significant in the era of atypical antipsychotics, possibly because of their antidyskinetic effect. PMID- 15283537 TI - The psychosocial effects of being quarantined following exposure to SARS: a qualitative study of Toronto health care workers. AB - OBJECTIVE: To examine the psychosocial effects on health care workers of being quarantined because of exposure to severe acute respiratory syndrome (SARS). METHOD: We used semistructured qualitative interviews. RESULTS: We identified 3 major themes concerning psychosocial effects: loss, duty, and conflict. CONCLUSIONS: Quarantined workers experienced stigma, fear, and frustration. We highlight the need for clear and easily accessible information on dealing with infectious diseases. Practical advice on coping and stress management techniques for health care workers are needed in preparation for potential future outbreaks of infectious diseases. PMID- 15283538 TI - Hard times and good friends: negative life events and social support in patients with seasonal and nonseasonal depression. AB - OBJECTIVE: Although a relatively large body of research has now accumulated concerning the relation between negative life events, social support, and major depressive disorder (MDD), little is known about the relation between seasonal affective disorder and these psychosocial variables. This study aimed to compare baseline levels of negative life events (NLEs) and perceived social support (SS) in patients with seasonal and nonseasonal depression. METHOD: Canadian patients with winter seasonal affective disorder (SAD) (n = 26) and nonseasonal recurrent MDD (n = 66) completed measures of recent NLEs (the List of Threatening Experiences) and perceived SS (the Social Support Survey) prior to treatment. RESULTS: No significant between-group differences were observed in mean number of NLEs experienced or in quality of SS. Perceived SS was impaired in both groups, compared with patients with chronic medical conditions. CONCLUSIONS: The results of this study complement those of previous research reporting increased incidence of NLEs and decreased SS in primary care patients with high seasonality in the UK. Future research is required to determine the causal relation between these psychosocial risk factors and SAD and to assess whether they have an effect on, or are affected by, treatment interventions for SAD. PMID- 15283539 TI - Combined transcranial magnetic stimulation and right unilateral electroconvulsive therapy in patients with treatment-refractory depression. PMID- 15283540 TI - Re: treatment noncompliance with orally disintegrating olanzapine tablets. PMID- 15283541 TI - Quetiapine in the management of psychosis secondary to huntington's disease: a case report. PMID- 15283542 TI - Ziprasidone-induced lupus erythematosus. PMID- 15283543 TI - Lorazepam-induced prolongation of the QT interval in a patient with schizoaffective disorder and complete AV block. PMID- 15283544 TI - Healthcare reform in Canada: the Romanow Report. PMID- 15283545 TI - U.K. bioethics, U.K. metabioethics: organ sales and the justification of bioethical methods. PMID- 15283546 TI - Healthcare ethics in Finland: a follow-up. PMID- 15283547 TI - Dealing drugs with the Bush. PMID- 15283548 TI - Administrative legislation in Japan: guidelines on scientific and ethical standards. PMID- 15283549 TI - The Estonian healthcare system and the genetic database project: from limited resources to big hopes. PMID- 15283550 TI - Developments regarding ethical issues in medicine in the Republic of Croatia. PMID- 15283551 TI - Reproductive health and research ethics: hot issues in Argentina. PMID- 15283552 TI - Abortion in Brazilian bioethics. PMID- 15283553 TI - CQ sources/bibliography. PMID- 15283554 TI - Response to "The creation lottery" by Julian Savulescu and John Harris (CQ Vol 13, No 1). The creation lottery and method in bioethics: a comment on Savulescu and Harris. PMID- 15283555 TI - Response to "The rise and fall of death: the plateau of futility" by Lawrence J. Schneiderman, Holly Teetzel, and Todd Gilmer (CQ Vol 12, No 3). Correcting false impressions. PMID- 15283556 TI - Response to "What constitutes a just match? A reply to Murphy" by D. Micah Hester (CQ Vol 12, No 1). Of need, justice, and random acts of education. PMID- 15283557 TI - The moral authority of symbolic appeals in biomedical ethics. PMID- 15283558 TI - Electron capture dissociation in a radio frequency ion trap. AB - We report on the first evidence of electron capture dissociation (ECD) in a radio frequency (rf) ion trap. Peptide ions, [substance P]2+, trapped in a two dimensional, linear rf ion trap were cleaved by electrons injected along the central axis of the trap. Along the axis, the rf field component was zero and a magnetic field of 50 mT was applied. This electron injection scheme keeps the energy of the electrons below 1 eV, preventing them from heating by the rf field. The present ECD efficiency is approximately 4% by irradiation of electron current of 0.2 microA for 80 ms. ECD in rf traps may open high-throughput and low-cost ECD applications to obtain molecular structure information complementary to collision-induced dissociation. PMID- 15283559 TI - Derivatization of surface-bound peptides for mass spectrometric detection via threshold single photon ionization. AB - Chemical derivatization of peptides allows efficient F2 laser single photon ionization (SPI) of Fmoc-derivatized peptides covalently bound to surfaces. Laser desorption photoionization mass spectrometry using 337-nm pulses for desorption and 157.6-nm pulses for threshold SPI forms large ions identified as common peptide fragments bound to either Fmoc or the surface linker. Electronic structure calculations indicate the Fmoc label is behaving as an ionization tag for the entire peptide, lowering the ionization potential of the complex below the 7.87-eV photon energy. This method should allow detection of many molecular species covalently or electrostatically bound to surfaces. PMID- 15283560 TI - Mass spectrometric imaging of lipids in brain tissue. AB - The spatial distributions of various specific lipids in freeze-dried mouse brain sections were monitored using time-of-flight secondary ion mass spectrometry (TOF SIMS). Mouse brain sections were prepared by cryosectioning, rinsing in 0.15 M NH3HCOO, and freeze-drying, after which the samples were analyzed directly by TOF SIMS, using Au3+ ions as primary ions. Positive and negative TOF-SIMS spectra of the tissue surface contained peaks from quasimolecular ions of a variety of specific lipids, including cholesterol, sulfatides, phosphatidylinositols, and phosphatidylcholines. Images showing the spatial signal intensity distributions of specific ions were recorded across analysis areas ranging from 100 x 100 microm(2) to 9 x 9 mm(2). The results demonstrate a highly complementary localization of cholesterol and phosphatidylcholine over dimensions from millimeter to micrometer range. Characteristic spatial distributions of several other lipids, including sulfatides and phosphatidylinositols, were observed. Principal component analysis was used to localize regions of the sample surface that show common spectral features. Spectra from different such regions showed large variations in lipid ion signals, indicating large variations in the lipid composition in different regions. PMID- 15283561 TI - Spongiform immobilization architecture of ionotropy polymer hydrogel coentrapping alcohol oxidase and horseradish peroxidase with octadecylsilica for optical biosensing alcohol in organic solvent. AB - An organic-phase optical alcohol biosensor consisting of alcohol oxidase and horseradish peroxidase coimmobilized in a spongiform hydrogel matrix of hydroxethyl carboxymethyl cellulose, an adduct of 3-methoxy-4-ethoxy benzaldehyde, 4-tert-butylpyridinium acetohydrazone, silica gel particles, and octadecylsilica particles in conjunction with an optical oxygen transducer has been successfully fabricated. The novel enzyme entrapment structure was mainly characterized with desirable solvent permeability, high efficiency of mass transfer for reactants, and good accessibility and stability of the immobilized enzymes. The biosensor could work in water-miscible solvent such as a solvent mixture of acetonitrile and phosphate aqueous buffer, as well as hydrophobic organic solvent such as n-hexane. The biosensor had the highest sensitivity to methanol in both solvent systems. Under the stop-flow mode, the biosensor had the analytical working ranges from 80 microM to 90 mM methanol in n-hexane and 0.10 to 90 mM methanol in acetonitrile/buffer. When the biosensor functioned in n hexane, it could take benzaldehyde as an alcohol substrate and was free from any pH disturbance. In the presence of coimmobilized horseradish peroxidase, the operational life of the biosensor was 60 assays and the shelf life was longer than two weeks. The biosensor has been satisfactorily applied to the determination of methanol in commercial gasoline-methanol blend samples. PMID- 15283562 TI - Detection of chemicals by a reporter immunoassay: application to fluoride. AB - This report describes a concept in which an immunoassay is used indirectly to quantify a nonantigenic very low molecular weight compound participating in a chemical reaction with a haptenic reporter. The detection limit of each reagent is, therefore, governed only by the affinity of the antibodies toward the reporter. Fluoride was used as a model, and silylated estradiol was used as a reporter. Upon silylation with N-O-bis(trimethylsilyl)trifluoroacetamide (BSTFA) or N-O-bis(dimethylterbutylsilyl) trifluoroacetamide (MTBSTFA), estradiol is no longer recognized by antibodies specific to estradiol. After reaction with hydrofluoric acid (HF) or fluoride salts (KF, CsF, NaF), its immunoreactivity is restored, and native estradiol is formed and is detected by immunoassay. The level of synthesized estradiol is dependent on the concentration of fluoride. A fluoride detection limit of 0.3 microg/L (15 nM) is obtained. Potential interference with other acids has been eliminated by choosing the silyl group (trimethylsilyl vs tert-butyldimethylsilyl) and by selecting optimal reaction conditions for the desilylation. The method has been applied to the detection of fluoride salts in natural waters (range 0.28-9.0 mg/L) and in an atmosphere artificially contaminated with HF between 8 and 160 microg/m(3) in the parts-per billion range. This indirect immunoassay combines simplicity and high sensitivity and, therefore, can be used in field monitoring. Finally, the extension of the concept to other chemicals is discussed. PMID- 15283563 TI - A flow injection kinase assay system based on time-resolved fluorescence resonance energy-transfer detection in the millisecond range. AB - A flow injection analysis (FIA) system for biochemical assays using time-resolved fluorescence resonance energy transfer (TR-FRET) in the millisecond time scale was developed. As a model system, we studied a kinase assay, measuring the phosphorylation of poly(GT)-biotin (substrate) by a receptor tyrosine kinase (epidermal growth factor receptor). A streptavidin labeled with XL665 (SA-XL665) the acceptor-was coupled to the biotin moiety, and an antiphosphotyrosine antibody labeled with europium cryptate (Ab-EuK)-the donor-was coupled to the phosphorylated tyrosine group(s). Long-lived FRET can only occur if the substrate is successfully phosphorylated. For the time-resolved detection of such long lived luminescence phenomena in a flow system, the repetition rate of the excitation source plays a crucial role. Good results were obtained for a small sized commercially available quadrupled Nd:YAG laser emitting at 266 nm with a repetition rate of 7.8 kHz and a pulse width of 0.3 ns. The long-lived emissions of the donor at 625 nm and that of the acceptor at 665 nm were monitored simultaneously with two photomultipliers, using a delay time of 50 micros and a gate time of 75 micros to exclude background fluorescence interferences. In the FIA experiments, the Ab-EuK concentration was 6 nM and the substrate concentration and SA-XL665 concentrations were 7 nM. By monitoring the intensity changes at 625 and 665 nm, the inhibition of tyrosine kinase by tyrphostin AG1478 was studied and an IC(50) value of 5.1 +/- 0.4 nM obtained. PMID- 15283564 TI - Realization and characterization of porous gold for increased protein coverage on acoustic sensors. AB - Immunosensors show great potential for the direct detection of biological molecules. The sensitivity of these affinity-based biosensors is dictated by the amount of receptor molecules immobilized on the sensor surface. An enlargement of the sensor area would allow for an increase of the binding capacity, hence a larger amount of immobilized receptor molecules. To this end, we use electrochemically deposited "gold black" as a porous sensor surface for the immobilization of proteins. In this paper, we have analyzed the different parameters that define the electrochemical growth of porous gold, starting from flat gold surfaces, using different characterization techniques. Applied potentials of -0.5 V versus a reference electrode were found to constitute the most adequate conditions to grow porous gold surfaces. Using cyclic voltammetry, a 16 times increase of the surface area was observed under these electrochemical deposition conditions. In addition, we have assessed the immobilization degree of alkanethiols and of proteins on these different porous surfaces. The optimized deposition conditions for realizing porous gold substrates lead to a 11.4-fold increase of thiol adsorption and a 3.3-fold increase of protein adsorption, using the quartz crystal microbalance (QCM-D) as a biological transducer system. Hence, it follows that the high specific area of the porous gold can amplify the final sensitivity of the original flat surface device. PMID- 15283565 TI - Direct determination of the "organic extent" of tin species in environmental samples by X-ray absorption near-edge structure spectroscopy. AB - A direct method for the speciation of Sn compounds in solid environmental samples by X-ray absorption near-edge structure spectroscopy (XANES) has been developed. It was found that the method can provide the "organic extent", the average number of organic ligands bound to Sn, for environmental samples. For Sn XANES at the L(III), L(I), and K edges, systematic variations were found in the spectra for butyl-, phenyl-, and methyl-substituted Sn compounds depending on the organic extent. A quantitative relationship between the organic extent and the characteristics in the XANES spectra was determined based on the peak position, peak area ratio, and peak width. The detection limit was better than 10 microg/g Sn when using the K edge, which is sensitive enough for some environmental samples, e.g., sediments, biological samples, and antifouling paints, and the sensitivity will be better if a more intense X-ray source such as an undulator or Wiggler beamline is used. The present XANES method is totally nondestructive, having the advantage that no complicated pretreatment procedures are needed, whereas such procedures are essential in conventional chromatographic analyses, which may cause experimental error by alteration of Sn species and poor recovery during analyses. Although the XANES method only provides the average number of organic ligands, the direct speciation using XANES will be helpful for estimating roughly the ratio of organic and inorganic Sn species, which can be used to study organotin transformations in sediment cores and inspection of organotin compounds in antifouling paints. In particular, micro-XANES analysis based on the present method is a promising tool in obtaining the distribution of organotin species in biological samples and specific phases in sediments. PMID- 15283566 TI - Selective mercury determination after membrane complexation and total reflection X-ray fluorescence analysis. AB - A new method for low mercury concentration analysis in drinking waters is presented. Membranes containing a few micrograms of various complexing reagents were produced on the surface of quartz glasses (reflectors). The reflectors were immersed in water solutions containing various concentrations of inorganic mercury salts at low concentrations (1-40 ng/mL). The membranes were left to equilibrate in 5-500 mL of solution for many hours; when the equilibration stage was finished they were cleaned with ultrapure water and left to dry. Analyses were performed by total reflection X-ray fluorescence (TXRF). The effects of various experimental parameters (complexing agent, equilibrium time, sample volume, etc.) as well as the selectivity of the membranes were studied. The complexing reagent dithizone with a PVC-based membrane gave the best results. The limit of quantitation was 0.8 ng/mL. PMID- 15283567 TI - Localization of the O-glycosylated sites in peptides by fixed-charge derivatization with a phosphonium group. AB - The present study demonstrates that matrix-assisted laser desorption ionization/postsource decay (MALDI/PSD) analysis of the molecular cation of glycopeptides derivatized at their amino terminus with a phosphonium group cleaves peptide backbone without removing the glycan. The predictable a-type fragment ions retain the glycan moiety, enabling unambiguous localization of O glycans on the peptide chain. In contrast, collision-activated dissociation tandem mass spectrometry analysis carried out on the doubly charged protonated phosphonium cation results in the predominant loss of the sugar moiety from the peptide. This result supports the previously proposed charge-induced fragmentation mechanism of the sugar-peptide bond. MALDI/PSD analysis of glycopeptides converted to their acetyl phosphonium derivatives is an effective alternative to electron capture dissociation, as illustrated by the positioning of up to three GalNac residues along the full tandem repeat peptide sequence derived from the MUC 5AC mucin. PMID- 15283568 TI - Determination of dissociation constants for protein-ligand complexes by electrospray ionization mass spectrometry. AB - A fully automated biophysical assay based on electrospray ionization mass spectrometry (ESI-MS) for the determination of the dissociation constants (KD) between soluble proteins and low molecular mass ligands is presented. The method can be applied to systems where the relative MS response of the protein and the protein-ligand complexes do not reflect relative concentrations. Thus, the employed approach enables the use of both electrostatically and nonpolar bound complexes. The dynamic range is wider than for most biological assays, which facilitates the process of establishing a structure-activity relationship. This fully automated ESI-MS assay is now routinely used for ligand screening. The entire procedure is described in detail using hGHbp, a 25-kDa extracellular soluble domain of the human growth hormone receptor, as a model protein. PMID- 15283569 TI - A method for extraction and quantification of Ginkgo terpene trilactones. AB - A method was developed for the extraction and quantification of pharmacologically active terpene trilactones (ginkgolides, bilobalide) from the tissues of Ginkgo biloba L. and pharmaceutical ginkgo products by RP-HPLC, based on the theory of terpene trilactones ionization. Four ginkgolides (GA, GB, GC, GJ) and bilobalide (BB) from both the ginkgo leaves and commercially available ginkgo extracts were quantitatively extracted by using this method. The recovery rate of the method was 97.5-100% with RSD of 1.2-2.8%. The detection limit was 0.05-0.1 microg, and the linear range was 0.1-12 microg. This detection limit represents a marked improvement over previously reported methods, suggesting the new method is a viable technique for routine analysis of ginkgo terpene trilactones in natural and commercial samples. The method reported by van Beek et al. in 1991 (van Beek, T. A.; Scheeren, H. A.; Rantio T.; Melger, W. C.; Lelyveld, G. P. J. Chromatogr. 1991, 543, 375-387.) was used as a reference method to monitor the accuracy of extraction and analysis in this study. SSI-MS technique was used to identify isolated target components. Carbohydrase treatment and solubility of terpene trilactones in various solvents were also discussed. PMID- 15283570 TI - Gold nanoparticles as selective and concentrating probes for samples in MALDI MS analysis. AB - MALDI mass spectrometry is used widely in various fields because it has the characteristics of speed, ease of use, high sensitivity, and wide detectable mass range, but suppression effects between analyte molecules and interference from the sample matrix frequently arise during MALDI analysis. The suppression effects can be avoided if target species are isolated from complicated matrix solutions in advance. Herein, we proposed a novel method for achieving such a goal. We describe a strategy that uses gold nanoparticles to capture charged species from a sample solution. Generally, ionic agents, such as anionic or cationic stabilizers, encapsulate gold nanoparticles to prevent their aggregation in solution. These charged stabilizers at the surface of the gold particles are capable of attracting oppositely charged species from a sample solution through electrostatic interactions. We have employed this concept to develop nanoparticle based probes that selectively trap and concentrate target species in sample solutions. Additionally, to readily isolate them from solution after attracting their target species, we used gold nanoparticles that are adhered to the surface of magnetic particles through S-Au bonding. A magnet can then be employed to isolate the Au@magnetic particles from the solution. The species trapped by the isolated particles were then characterized by MALDI MS after a simple washing. We demonstrate that Au@magnetic particles having negatively charged surfaces are suitable probes for selectively trapping positively charged proteins from aqueous solutions. In addition, we have employed Au@magnetic particle-based probes successfully to concentrate low amounts of peptide residues from the tryptic digest products of cytochrome c (10(-7) M). PMID- 15283571 TI - Thermodynamic analysis of cyclosporin a binding to cyclophilin a in a lung tumor tissue lysate. AB - We report on the application of SUPREX (stability of unpurified proteins from rates of H/D exchange) to the analysis of a protein-ligand binding interaction under the ex vivo solution conditions of a human lung tumor tissue lysate. A SUPREX-derived binding free energy (i.e. DeltaDeltaG(f) value) and dissociation constant (i.e., K(d) value) were determined for the binding of cyclosporin A (CsA) to a cyclophilin A (CypA) sample in which the protein was a component of a tissue lysate derived from fresh frozen lung tumor. The DeltaDeltaG(f) and K(d) values determined by SUPREX for CsA binding to CypA in this unpurified protein sample, 4.7 +/- 0.8 kcal/mol and 77 +/- 17 nM, respectively, were comparable to the those obtained when SUPREX was used to analyze the binding of CsA to a highly purified CypA sample, 4.2 +/- 1.0 kcal/mol and 32 +/- 20 nM, respectively. Moreover, the SUPREX-derived K(d) values determined in this work were both in the range of those previously reported for the CypA-CsA complex. The results of this proof-of-principle work validate the extension of SUPREX to the thermodynamic analysis of proteins and protein-ligand binding interactions in the unpurified, ex vivo conditions of human tissue lysates,and they represent the first K(d) measurement on a protein-ligand complex under such conditions PMID- 15283572 TI - Comparison of different mass spectrometric techniques combined with liquid chromatography for confirmation of pesticides in environmental water based on the use of identification points. AB - Three mass spectrometric techniques have been used and compared for the confirmation of the presence of several pesticides that had been detected in environmental water samples by a previously reported SPE-LC-MS/MS screening method. The 2002/657/EC European Comission Decision establishes the need to obtain at least three identification points (IPs) in order to confirm organic residues and contaminants in live animals and animal products. In this paper, a similar approach has been applied for confirmation of pesticides in water samples, using triple quadrupole mass spectrometry (QqQ), time-of-flight mass spectrometry (TOF), and hybrid quadrupole time-of-flight mass spectrometry (QTOF) to achieve the required IPs. The number of IPs collected, the sensitivity, and the practical advantages and disadvantages of these techniques have been discussed. In summary, the QqQ instrument allowed the confirmation of detected pesticides even at very low concentrations (ng/L) achieving between four and five IPs when adding confirmatory transitions. The direct confirmation with a TOF instrument was only feasible for those compounds showing sufficient sensitivity, isotopic pattern, or easy in-source fragmentation. In other cases, the required IPs could be reached by adding IPs earned with this technique to those obtained from the MS/MS screening method. Finally, the use of a QTOF instrument allowed obtaining up to 20 IPs in a single run at relatively high concentrations (submicrograms per liter) as no "ion shopping" was required. Additionally, the application of TOF and QTOF techniques made it possible to detect some nontarget organic contaminants, which were not included in the screening method. PMID- 15283573 TI - Simultaneous quantification of opiates, cocaine, and metabolites in hair by LC APCI-MS/MS. AB - A quantitative LC-APCI-MS/MS method for simultaneous measurement of opiates, cocaine, and metabolites in hair was developed and validated. Cocaine and opiates were extracted from pulverized hair via sonication in methanol at 37 degrees C for 3 h. Samples were cleaned up using solid-phase extraction. LC separation was achieved in 20 min with identification and quantification by selected reaction monitoring. Calibration by linear regression analysis utilized deuterated internal standards and a weighting factor of 1/x (R(2) > 0.998). Limits of quantification (LOQ) ranged from 17 to 50 pg/mg for cocaine and metabolites and were 83 pg/mg for opiates. Standard curves were linear from the LOQ to 5000 pg/mg for cocaine and metabolites, except benzoylecgonine (2500 pg/mg). Opiate standard curves were linear from the LOQ to 12500 pg/mg. Accuracy ranged from 84 to 115% for all quantitative analytes. Precision, % relative standard deviation, was less than 11.0% for all analytes. Methanolic sonication produced less than 5% hydrolysis of cocaine and 6-acetylmorphine. The method should be useful for studying cocaine and opiate distribution into hair. PMID- 15283574 TI - Discordance between net analyte signal theory and practical multivariate calibration. AB - Lorber's concept of net analyte signal is reviewed in the context of classical and inverse least-squares approaches to multivariate calibration. It is shown that, in the presence of device measurement error, the classical and inverse calibration procedures have radically different theoretical prediction objectives, and the assertion that the popular inverse least-squares procedures (including partial least squares, principal components regression) approximate Lorber's net analyte signal vector in the limit is disproved. Exact theoretical expressions for the prediction error bias, variance, and mean-squared error are given under general measurement error conditions, which reinforce the very discrepant behavior between these two predictive approaches, and Lorber's net analyte signal theory. Implications for multivariate figures of merit and numerous recently proposed preprocessing treatments involving orthogonal projections are also discussed. PMID- 15283575 TI - Determining oxygen diffusion coefficients in polymer films by lifetimes of luminescent complexes measured in the frequency domain. AB - Polymer films doped with luminescent ruthenium complexes are proving to be important oxygen sensors. We describe a technique using lifetime measurements in the frequency domain for determining the diffusion coefficient of oxygen through various polymer supports. These fundamental measurements will allow for more rational design of improved sensors. Three types of polymers were doped with [Ru(4,7-diphenyl-1,10-phenanthroline)3]Cl2. We monitored the luminescence versus time after applying a step increase in the oxygen pressure at the surface of the film. We modeled the decrease in apparent lifetime as a function of time using the diffusion coefficient of oxygen in the polymer as the only adjustable parameter. The model accurately predicted the lifetime versus time curves, and diffusion coefficients agreed well with those obtained from intensity measurements. The advantages and disadvantages of the lifetime technique to those used earlier are discussed. PMID- 15283576 TI - Elimination of dimer formation in InIIIporphyrin-based anion-selective membranes by covalent attachment of the ionophore. AB - The spontaneous hydroxy-bridged dimer formation of metalloporphyrins in ion selective membranes gives rise to a short sensor lifetime (typically days), triggered by solubility problems, the occurrence of a super-Nernstian response slope, and a pH cross response. This dimer formation is eliminated here by covalent attachment of the ionophore to the polymer matrix. Specifically, two different indium(III)porphyrins containing polymerizable groups, the chloride selective chloro(3-[18-(3-acryloyloxypropyl)-7,12-bis(1-methoxyethyl)-3,8,13,17 tetramethylporphyrin-2-yl]propyl ester)indium(III) and the nitrite-selective Chloro(5-(4-acryloyloxyphenyl)-10,15,20-triphenylporphyrinato)indium(III), were synthesized and copolymerized with methyl methacrylate and decyl methacrylate. The covalent attachment of the ionophore to the polymer matrix indeed prevents the metalloporphyrin from forming dimeric species, as confirmed by UV/visible spectroscopy. The ion-selective membranes with grafted indium porphyrin showed Nernstian response slopes to chloride, nitrite, perchlorate, and thiocyanate anions, with a selectivity comparable to membranes with freely dissolved or underivatized metalloporphyrin. The membranes containing grafted ionophores showed a lifetime of at least two months, apparently since crystallization of the poorly soluble dimeric species may no longer occur. This is one of the first examples where the covalent attachment of an ionophore drastically improves on a number of important sensor characteristics. PMID- 15283577 TI - Stability of the inner polyaniline solid contact layer in all-solid-state K+ selective electrodes based on plasticized poly(vinyl chloride). AB - A simple and powerful method based on UV-visible spectroscopy is presented for studying the stability of the inner electrically conducting polyaniline (PANI) solid contact (SC) layer in all-solid-state ion-selective electrodes (ISE). The influence of the plasticized poly(vinyl chloride) (PVC) membrane (ISM) composition and the pH of the sample solution on the stability of the solid contact is reported. PANI is used as a model compound in this study, but the method presented is universal and can be applied to different types and combinations of SCs and ISMs. It provides a tool for finding the best combination of conducting polymer and ISM for solid contact ISEs. PANI is deposited electrochemically either on glassy carbon or quartz glass covered with a thin layer of tin oxide, and a K+-selective ISM is deposited on top of the PANI layer. The short-term stability of the inner PANI layer is good for all membrane types in buffer solutions with pH 2, 6, and 9, indicating that the outer plasticized PVC membrane hinders the emeraldine salt-emeraldine base transition of the highly pH sensitive PANI layer. The solid contact K+-selective electrodes studied showed a Nernstian response of 58.2 +/- 0.1 mV/log aK. Significant differences are observed in the long-term stability of the inner PANI layer between the different membrane types. This indicates that water uptake of the PVC membrane and its permeability to OH- ions are critical parameters affecting the stability of the PANI layer. The solid contact electrodes based on PANI may require a composition of the PVC membrane different from those typically used in conventional ISEs with an inner solution. PMID- 15283578 TI - Transport model of chemical secretion process for tracking exocytotic event dynamics using electroanalysis. AB - A unified model is developed to analyze the key features of the chemical secretion process observed in experimental studies of various vesicles with application to electroanalytical measurements of vesicular exocytosis. The intimately coupled dynamics and kinetics are simultaneously resolved based on continuum fluid flow, mass transport, and linear elasticity theories combined with biomembrane mechanics. We report three case studies of exocytosis, including a large electroporated granule of the mast cell, a small and clear synaptic vesicle, and a medium size vesicle in the chromaffin cell. The simulation results for each case are compared with electroanalytical measurements from the literature. The results provide a theoretical ground for defining the rate controlling step(s) of an exocytotic sequence, allowing interpretation of electroanalysis data. Thus, it provides a tool for theoretical verification of competing hypotheses of what controls/limits messenger release during exocytosis. Simulations show that the pore size, the pore opening velocity, and the swelling dynamics of the granule matrix play the key roles in controlling the messenger release kinetics. PMID- 15283579 TI - Determination of hereditary mutations in the BRCA1 gene using archived serum samples and capillary electrophoresis. AB - Analysis of DNA variation in biological samples most frequently utilizes the polymerase chain reaction (PCR) performed on extracted genomic DNA, followed by visualization of alleles using various methodologies. Few reports have demonstrated that amplification of DNA from plasma and serum samples is possible. We have performed DNA amplification on a large set of serum samples (n = 2955). Here, we report that known hereditary mutations in the BRCA gene can efficiently be analyzed in serum samples collected and stored over several decades. Fragments were PCR-amplified following a short initial denaturation of the serum sample in a standard microwave oven. Fragment analysis was subsequently performed using a DNA capillary-sequencing instrument. The PCR success rates were fragment- and size-dependent ranging from 83.2% to 97.9%. Of the 11,820 polymerase chain reactions performed, the overall PCR success rate was 91.3% (10,796/11,820), which is comparable to PCR performed on genomic DNA. The advantage of the method described herein is its ability to utilize archival material stored in serum biobanks for long periods of time. PMID- 15283580 TI - Direct detection of biomolecules in a capillary electrophoresis-chemiluminescence detection system. AB - Direct detection of biomolecules, such as alpha-amino acids, peptides, and proteins, was accomplished using a capillary electrophoresis-chemiluminescence detection system, in which a luminol-hydrogen peroxide-Cu(II)-catalyzed chemiluminescence reaction was utilized. Biomolecules migrated in the capillary, where they mixed with luminol and the Cu(II) catalyst included in the running buffer. The capillary outlet was inserted into a batch-type chemiluminescence detection cell with hydrogen peroxide-supplemented electrolyte solution. Chemiluminescence was observed at the tip of the capillary outlet. The chemiluminescence peak from biomolecules appeared due to the enhancement of Cu(II) catalytic activity for luminol-hydrogen peroxide chemiluminescence. The Cu(II) was more catalytically active when it interacted with biomolecules forming Cu(II)-biomolecule complexes. In this study, biomolecules were directly separated and detected in a capillary electrophoresis-chemiluminescence detection system. Twenty alpha-amino acids, 4 peptides, and 11 proteins were examined. Most of them were detected with satisfactory CL intensity response. Glutamic acid, an alpha amino acid, was detected at concentrations ranging from 2.0 x 10(-7) to 1.2 x 10( 5) M with a detection limit (S/N = 3) of 1.0 x 10(-7) M (0.6 fmol). Glycylglycine, a peptide, was detected at concentrations ranging from 1.7 x 10( 7) to 1.2 x 10(-5) M with a detection limit (S/N = 3) of 1.7 x 10(-7) M (0.9 fmol). Hemoglobin, a heme protein, in which the heme structure was independently catalytically active, was detected at concentrations ranging from 1.2 x 10(-7) to 1.0 x 10(-5) M with a detection limit (S/N = 3) of 1.2 x 10(-7) M (0.6 fmol). Representative mixtures of alpha-amino acids and peptides were well detected with superior separation. PMID- 15283581 TI - Reflective interferometric detection of label-free oligonucleotides. AB - New chip-based methods for the detection of unmodified biomolecular targets have significant potential as enabling technology in fundamental biology and biomedical analysis. We report a method based on changes in reflectivity from specially fabricated substrates that is capable of detecting the binding of as little as an average of 0.2 nm (i.e., a fraction of a monolayer) of biomolecules. We demonstrate the method on detection of femtomole quantities of untagged oligonucleotides in an array format, showing that the amount of target bound can be determined quantitatively. The simplicity of the approach promises to make it broadly applicable for any biomolecule for which suitable molecular recognition chemistry is available. PMID- 15283582 TI - Hadamard transform capillary electrophoresis combined with absorption spectrometry. AB - A novel injection device for applying absorption spectrometry to Hadamard transform (HT) capillary electrophoresis is described. A small hole, at the center part of the capillary, functions as an inlet port for the sample. The hole is immersed in a sample solution and the end of the capillary that is usually employed for sample introduction is immersed in a buffer solution. An ultraviolet absorption detector is placed between the sample injection port and the other end of the capillary filled with a buffer solution. A high potential is continuously applied between the injection port and the end of the capillary, which allows the sample solution to be introduced into the separation capillary. By application of a higher potential modulated according to a Hadamard code between both ends of the capillary, the buffer solution is injected into the separation capillary. In some preliminary experiments, this injection device was utilized to introduce a single sample segment into a capillary. As expected, a single peak was observed in the electropherogram for a sample containing a single component. This device was then employed for multiple sample injection in HT capillary electrophoresis. An 8-fold improvement in the S/N ratio was observed when the HT technique was used, in which a 255-order of a Hadamard matrix was used, as expected from theory. The present approach was also utilized for the sensitive detection of a sample comprised of multiple components. PMID- 15283583 TI - Two-dimensional protein separation with advanced sample and buffer isolation using microfluidic valves. AB - Methods are described to achieve more efficient multidimensional protein separation in a microfluidic channel. The new methods couple isoelectric focusing (IEF) with high ionic strength electrophoretic separations by active microvalve control in a microchip. Several experiments demonstrating independent 2D separation were performed, and critical parameters for optimal chip performance were identified, including channel passivation, electroosmosis control, and IEF linearity control. This strategy can be used for integration of different heterogeneous separation techniques, such as IEF, capillary electrophoresis, and liquid chromatography. This new device can be ideal for preseparation and preconcentration of complex biomolecule samples for a streamlined biomolecule analysis using mass spectrometry. PMID- 15283584 TI - Chromatographic preconcentration coupled to capillary electrophoresis via an in line injection valve. AB - A preconcentration-capillary electrophoresis (CE) system using a small precolumn in combination with an in-line injection valve is presented. The advantage of the present design is the ability to perform the sample preconcentration fully independently from the CE separation and to prevent sample matrix and washing solvents from entering the CE capillary. With a micro injection valve, sample could be effectively introduced into the CE system in an in-line fashion without seriously affecting the CE separation efficiency. Breakthrough volume, desorption efficiency, and elution volume for the C18 microcolumn (5 x 0.5 mm i.d.) were established, yielding values of 750 microL, 70%, and 0.9-1.1 microL, respectively, using enkephalin peptides. The time between the start of the desorption of the analytes from the precolumn and the injection into the CE system was also studied in order to achieve optimal sensitivity and separation efficiency. The performance of the complete system was demonstrated by the preconcentration and separation of an enkephalin mixture. Using a sample volume of 250 microL and a CE injection voltage of -15 kV for 12 s, linearity was observed over 2 orders of magnitude, and detection limits (S/N = 3) were in the 5 10 ng/mL range. A 1000-fold sensitivity enhancement is obtained using this setup, as compared to a regular CE setup. For 100 ng/mL samples, repeatabilities (RSDs) of migration time and peak area were 1.2 and 11%, respectively. PMID- 15283585 TI - Metal oxide-based monolithic complementary metal oxide semiconductor gas sensor microsystem. AB - A fully integrated gas sensor microsystem is presented, which comprises for the first time a micro hot plate as well as advanced analog and digital circuitry on a single chip. The micro hot plate is coated with a nanocrystalline SnO2 thick film. The sensor chip is produced in an industrial 0.8-microm CMOS process with subsequent micromachining steps. A novel circular micro hot plate, which is 500 x 500 microm(2) in size, features an excellent temperature homogeneity of +/-2% over the heated area (300-microm diameter) and a high thermal efficiency of 6.0 degrees C/mW. A robust prototype package was developed, which relies on standard microelectronic packaging methods. Apart from a microcontroller board for managing chip communication and providing power supply and reference signals, no additional measurement equipment is needed. The on-chip digital temperature controller can accurately adjust the membrane temperature between 170 and 300 degrees C with an error of +/-2 degrees C. The on-chip logarithmic converter covers a wide measurement range between 1 kOmega and 10 MOmega. CO concentrations in the sub-parts-per-million range are detectable, and a resolution of +/-0.1 ppm CO was achieved, which renders the sensor capable of measuring CO concentrations at threshold levels. PMID- 15283586 TI - Molecule by molecule direct and quantitative counting of antibody-protein complexes in solution. AB - We have used two-color fluorescence coincidence detection to directly count individual protein-antibody complexes of protein G or herpes simplex virus labeled with one or more red- and blue-excited antibodies. This allowed quantitative measurement of the concentration of the protein-antibody complexes over 3 orders of magnitude down to the femtomolar level. Single molecule measurements in diluted serum are also possible. The sample preparation is simple, takes place in solution, and requires no separation. Both the antibody affinity and complex dissociation rate are important in determining the sensitivity of the method. At present, the sensitivity limit of 50 fM is determined by the encounter rate of the labeled analyte with the probe volume. This method can be used to detect and quantitate proteins and to measure the stoichiometry, equilibrium constant, and dissociation rate of protein-protein complexes at low concentrations. PMID- 15283587 TI - Raman microscopic analysis of single microbial cells. AB - We demonstrate the utility of the Raman confocal microscope to generate a spectral profile from a single microbial cell and the use of this approach to differentiate bacterial species. In general, profiles from different bacterial taxa shared similar peaks, but the relative abundances of these components varied between different species. The use of multivariate methods subsequently allowed taxa discrimination. Further investigations revealed that the single-cell spectra could be used to differentiate between growth phases of a single species, but these differences did not obscure the overall interspecies discrimination. Finally, we tested the efficacy of the method as a means to identify cells responsible for the uptake of a specific substrate. A single strain was grown in media containing incrementally varying ratios of (13)C(6) to (12)C(6) glucose, and it was found that (13)C incorporation shifted characteristic peaks to lower wavenumbers. These findings suggest that Raman microscopy has significant potential for studies requiring the taxonomic identity and functioning of single microbial cells to be determined. PMID- 15283588 TI - Direct observation of single native DNA molecules in a microchannel by differential interference contrast microscopy. AB - Direct observation of single native DNA molecules in a microchannel was monitored without fluorescence-dye labeling. At a PDMS/glass microchip, the image of individual lambda-DNA molecules appear sharp and distinct in Nomarski differential interference contrast microscopy. Intercalator dyes affected the physical properties and dynamic behavior of individual DNA molecules. From the migration velocities in the microchannel it is evident that native DNA molecules migrated faster than DNA molecules labeled with the intercalator YOYO-1. This is because YOYO-1 increases the molecular weight and size of lambda-DNA and decreases the charge. The electric field strength and pH also affected the dynamics of single DNA molecules. We also observed that YOYO-labeled DNA was more stretched out compared to native DNA. PMID- 15283589 TI - Nanoparticle-based in vivo investigation on blood-brain barrier permeability following ischemia and reperfusion. AB - The blood-brain barrier (BBB) represents a significant impediment to a large variety of central nervous system-active agents. In the current study, we applied fluorescent polystyrene nanospheres (20 nm) to study the BBB permeability following cerebral ischemia and reperfusion. A microdialysis probe was implanted in the cerebral cortex of an anesthetized rat injected with fluorescent polystyrene nanospheres. The circulating nanospheres extravasating to the brain extracellular fluids were collected by the probe. Fluorescence intensity in the microdialysates throughout the course of cerebral ischemia/reperfusion was measured. Cerebral ischemia and reperfusion induced transient accumulations of extracellular nanospheres in the brain. The accumulation of nanospheres may result from their extravasation from the blood vessels. The concurrent cerebral oxygen levels monitored using oxygen-dependent quenching of phosphorescence decreased following ischemia and returned to their original levels after reperfusion. In conclusion, we demonstrated that high temporal resolution measurements of BBB permeability in vivo can be obtained using fluorescence polystyrene nanospheres and that these data correlate with changes of cerebral oxygen concentration. This present investigation indicates that nanoparticles have potential clinical applications involving drug delivery and determination of therapeutic efficacy and on-site diagnosis. PMID- 15283590 TI - Analysis of protein phosphorylation by hypothesis-driven multiple-stage mass spectrometry. AB - We describe a strategy, which we term hypothesis-driven multiple-stage mass spectrometry (HMS-MS), for the sensitive detection and identification of phosphopeptides derived from enzymatic digests of phosphoproteins. In this strategy, we postulate that any or all of the potential sites of phosphorylation in a given protein may be phosphorylated. Using this assumption, we calculate the m/z values of all the corresponding singly charged phosphopeptide ions that could, in theory, be produced by the enzyme employed for proteolysis. We test ions at these m/z values for the presence of phosphoserine or phosphothreonine residues using tandem mass spectrometry (MS(2)) in a vacuum MALDI ion trap mass spectrometer, where the neutral loss of the elements of H(3)PO(4) (98 Da) provides a sensitive assay for the presence of phosphopeptides. Subsequent MS(3) analysis of the (M + H - 98)(+) peaks allows us to confirm or reject the hypotheses that the putative phosphopeptides are present in the sample. HMS-MS was successfully applied to the detection and identification of phosphopeptides from substrates of the Saccharomyces cerevisiae cyclin-dependent kinase (Cdk) Cdc28, phosphorylated in vitro (Ipl1) and in vivo (Orc6), basing hypothesis formation on the minimal Cdk consensus phosphorylation motif Ser/Thr-Pro. The method was also used to find in vitro phosphopeptides from a domain of the Drosophila melanogaster protein PERIOD, hypothesizing possible phosphorylations of all Ser/Thr residues without assuming a consensus motif. Our results demonstrate that HMS-MS is a sensitive, highly specific tool for systematically surveying proteins for Ser/Thr phosphorylation, and represents a significant step toward our goal of comprehensive phosphorylation mapping. PMID- 15283591 TI - High sensitivity and analyte capture with desorption/ionization mass spectrometry on silylated porous silicon. AB - Silylation chemistry on porous silicon provides for ultrahigh sensitivity and analyte specificity with desorption/ionization on silicon mass spectrometry (DIOS MS) analysis. Here, we report that the silylation of oxidized porous silicon offers a DIOS platform that is resistant to air oxidation and acid/base hydrolysis. Furthermore, surface modification with appropriate hydrophobic silanes allows analytes to absorb to the surface via hydrophobic interactions for direct analyte extraction from complex matrixes containing salts and other nonvolatile interferences present in the sample matrix. This enables rapid cleanup by simply spotting the sample onto the modified DIOS target and removing the liquid phase containing the interferences. This approach is demonstrated in the analysis of protein digests and metabolites in biofluids, as well as for the characterizing of inhibitors from their enzyme complex. An unprecedented detection limit of 480 molecules (800 ymol) for des-Arg(9)-bradykinin is reported on a pentafluorophenyl-functionalized DIOS chip. PMID- 15283592 TI - Detection of quadruplex DNA structures in human telomeres by a fluorescent carbazole derivative. AB - Single-stranded telomeric DNA tends to form a four-base-paired planar structure termed G-quadruplex. This structure was easily formed in vitro in the presence of monovalent cations. However, the existence of this structure in native human telomeres is unclear. Here we address this important question through the distinctive properties of 3,6-bis(1-methyl-4-vinylpyridinium)carbazole diiodide (BMVC) upon binding to various DNA structures. Although the fluorescence of BMVC increases significantly in the presence of DNA, BMVC has high sensitivity and binding preference to quadruplex d(T(2)AG(3))(4) over duplex DNA. In addition, the fluorescent emissions were characterized around 575 nm for quadruplex d(T(2)AG(3))(4) and 545 nm for most of duplex DNA. The 575-nm fluorescence emissions were detected in the mixtures of 2 nM BMVC with the chromosomal DNA that were extracted from human cells, suggesting the presence of quadruplex structure in human nucleus. Further analyzing the BMVC fluorescence at the ends of metaphase chromosomes and other regions of chromosomes, we detected the quadruplex-binding BMVC fluorescence at telomere-proximal regions. Together these results provide the first evidence for the presence of quadruplex structures in human telomeres. PMID- 15283593 TI - Liquid membrane operations in a microfluidic device for selective separation of metal ions. AB - A three-phase flow, water/n-heptane/water, was constructed in a microchannel (100 microm width, 25-microm depth) on a glass microchip (3 cm x 7 cm) and was used as a liquid membrane for separation of metal ions. Surface modification of the microchannel by octadecylsilane groups induced spontaneous phase separation of the three-phase flow in the microfluidic device, which allows control of interfacial contact time and off-chip analysis using conventional analytical apparatus. Prior to the selective transport of a metal ion through the liquid membrane in the microchannel, the forward and backward extraction of yttrium and zinc ions was investigated in a two-phase flow on a microfluidic device using 2 ethylhexyl phosphonic acid mono-2-ethylhexyl ester (commercial name, PC-88A) as an extractant. The extraction conditions (contact time of the two phases, pH, extractant concentration) in the microfluidic device were examined. These investigations demonstrated that the conventional methodology for solvent extraction of metal ions is applicable to solvent extraction in a microchannel. Finally, we employed the three-phase flow in the microchannel as a liquid membrane and observed the selective transport of Y ion through the liquid membrane. In the present study, we succeeded, for the first time, in the selective separation of a targeted metal ion from an aqueous feed solution to a receiving phase within a few seconds by employing a liquid membrane formed in a microfluidic device. PMID- 15283594 TI - Importance and reduction of the sidewall-induced band-broadening effect in pressure-driven microfabricated columns. AB - The influence of the detailed design of the sidewall region upon the over-all band-broadening in microfabricated packed-bed or collocated monolithic support structure (COMOSS) columns has been investigated using computational fluid dynamics (CFD) simulation techniques. It is shown that, under unretained solute conditions, very small structural variations of the order of only 5% of the particle diameter can give rise to a 4-fold increase of the band-broadening. A comprehensive study has been made to quantify this effect as a function of the fluid velocity, the particle diameter, the channel widths, and of course, the sidewall region design. Because the sidewall effect can be fully attributed to a mismatch between the flow rates in the column center and in the sidewall region, it is fortunately also quite straightforward to avoid it. A very simple design, yielding band-broadening values identical to that of a hypothetical sidewall-less column for all possible values of the flow velocity, the particle diameter, or the channel width is proposed. PMID- 15283595 TI - Semiautomated high-throughput extraction and cleanup method for the measurement of polybrominated diphenyl ethers and polybrominated and polychlorinated biphenyls in breast milk. AB - A semiautomated extraction and cleanup method has been developed to measure eight polybrominated diphenyl ethers (PBDEs), 2,2',4,4',5,5'-hexabromobiphenyl (BB 153), and 2,2',4,4',5,5'-hexachlorobiphenyl (CB-153). The method employs solid phase dispersion on diatomaceous earth in a solid-phase extraction cartridge followed by automated addition of internal standards ((13)C-labeled). Extraction is then performed using an automated modular solid-phase extraction system. The extraction procedure includes drying the sample on diatomaceous earth by pressurized nitrogen and eluting target analytes and lipids with dichloromethane. Lipid content is determined gravimetrically. Lipid determinations performed using this method are compared with other standard methods and with a certified reference material. A relative standard deviation of 7.9% was obtained for 130 determinations of the lipid content in a breast milk quality control sample. Final analytical determination of target analytes was performed by gas chromatography-isotope dilution high-resolution mass spectrometry. Relative standard deviations for the measurements of target analytes for which a labeled internal standard was available were below 10% for analytes at concentrations above 1 ng/g of lipid. Mean recoveries of the (13)C-labeled internal standards ranged from 60 to 89% for the eight PBDE congeners; 74 and 113% were recovered for BB-153 and CB-153, respectively. PMID- 15283596 TI - Automated monitoring of phosphatidylcholine biosyntheses in Plasmodium falciparum by electrospray ionization mass spectrometry through stable isotope labeling experiments. AB - The metabolic pathways contributing to phosphatidylcholine biosyntheses in Plasmodium falciparum, the malaria-causing parasite, was explored by electrospray ionization mass spectrometry. Phosphatidylcholine produced by the CDP-choline pathway and by the methylation of phosphatidylethanolamine was identified and quantified through isotopic labeling experiments. A straightforward method based on cone voltage directed in-source fragmentations and relative abundance measurement of endogenous versus deuterated specific fragment ions was developed for simple and rapid automated data acquisition. Such high-throughput analytical protocol allowed us to measure the relative contribution of two different metabolic pathways leading to phosphatidylcholine without performing technically more demanding and time-consuming MS/MS or LC/MS experiments. PMID- 15283597 TI - Enrichment of cysteine-containing peptides from tryptic digests using a quaternary amine tag. AB - A new strategy for specifically targeting cysteine-containing peptides in a tryptic digest is described. The method is based on quantitatively derivatizing cysteine residues with a quaternary amine tag (QAT). Tags were introduced into proteins following reduction of disulfide bonds through derivatization of cysteine residues with (3-acrylamidopropyl)trimethylammonium chloride. After trypsin digestion, derivatized cysteine-containing peptides were enriched by strong cation exchange chromatography. The method was validated using model peptides and a protein. The QAT strategy has several advantages over other methods for the selection of cysteine-containing peptides. One is that it increases the ionization efficiency of cysteine-containing peptides. The other is that chromatographic selection is achieved with simple, robust cation exchange chromatography columns. As a result, this new strategy provides a simple way to facilitate enrichment of cysteine-containing peptides, thereby reducing sample complexity in bottom-up proteomics. PMID- 15283598 TI - Controlled layer-by-layer formation of ultrathin TiO2 on silver island films via a surface sol-gel method for surface-enhanced Raman scattering measurement. AB - A surface sol-gel process has been demonstrated to be an effective method for the surface modification of silver island films as unique SERS substrates for monitoring molecular adsorption on a dielectric titania surface. This layer-by layer approach allows control of the thickness of the dielectric surface with a monolayer precision on silver surfaces. The enhancement of Raman scattering from adsorbed Rhodamine 6G molecules is inversely proportional to the thickness of the titania film, which is consistent with the decay of electromagnetic enhancement. Despite a reduction in the sensitivity of the film, a substantial improvement in the film was achieved as a result of the enhanced stability of this substrate compared to the silver island film without a TiO(2) coating. PMID- 15283599 TI - Detection of oligosaccharides labeled with cyanine dyes using matrix-assisted laser desorption/ionization mass spectrometry. AB - The sensitivity of oligosaccharides in mass spectrometry lags far behind that of peptides. This is a critical factor in realizing the high-throughput analysis of posttranslational modifications in proteomics. We here described that hydrazide derivatives of cyanine dyes (Cy3, Cy5) with a positive charge made excellent labeling reagents for the detection of oligosaccharides by matrix-assisted laser desorption/ionization mass spectrometry. Cy3-labeled standard N-glycan could be detected at 200 amol on the MALDI target plate in reflectron mode without any purification procedures after the labeling reaction, which may meet the level of sensitivity required in proteome research. Despite the general recognition that the production of signals of oligosaccharides under MALDI conditions would be highly dependent on the matrix, most of the known N-glycans from chicken ovalbumin could be detected upon Cye derivatization nearly independent of the kind of matrix tested (e.g., nor-harman, 2,5-dihydroxybenzoic acid and alpha cyano-4-hydroxycinnamic acid) without spoiling the signal strength. Postsource decay afforded simple spectra mainly consisting of Y-type fragment ions, thus simplifying the sequence analysis. In-source decay afforded a similar fragmentation pattern only when acidic matrixes were used. In addition, this derivatization technique was successfully applied to the profiling of N-glycans of gel-separated glycoproteins. PMID- 15283600 TI - Nitric oxide-releasing sol-gel particle/polyurethane glucose biosensors. AB - A hybrid sol-gel/polyurethane glucose biosensor that releases nitric oxide is developed and characterized. The biosensor consists of a platinum electrode coated with four polymeric membranes including the following: (1) sol-gel with immobilized glucose oxidase (GOx); (2) polyurethane to protect the enzyme; (3) NO donor-modified sol-gel particle-doped polyurethane; and (4) polyurethane. This configuration was developed due to the drastic reduction in sensitivity observed for NO donor-modified sol-gel film-based glucose sensors. For the hybrid sol gel/polyurethane biosensor, sol-gel particles are first modified with the NO donor and then incorporated into a polyurethane layer that is coated onto the preimmobilized GOx electrode. In this manner, the GOx layer is not exposed to the harsh conditions necessary to impart NO release ability to the biosensor, and only a minimal decrease in sensitivity due to the NO release is observed. The glucose response of the NO-releasing glucose biosensor and its NO generation profiles are reported. In addition, the stability of the sol-gel particles in the supporting polyurethane membrane is discussed. PMID- 15283601 TI - Coumarin tags for improved analysis of peptides by MALDI-TOF MS and MS/MS. 1. Enhancement in MALDI MS signal intensities. AB - The goal of this study was the development of N-terminal tags to improve peptide identification using high-throughput MALDI-TOF MS and MS/MS. The proposed tags, commercially available fluorescent derivatives of coumarin, can be advantageous for peptide analysis in both MS and MS/MS modes. This paper, part 1, will focus on the influence of derivatization on the intensities of MALDI-TOF MS signals of peptides. Labeling peptides with tags containing the coumarin core was found to enhance the intensities of peptide peaks (in some cases over 40-fold) in MALDI TOF MS using CHCA and 2,5-DHAP matrixes. The signal enhancement was found to be peptide- and matrix-dependent, being the most pronounced for hydrophilic peptides. No correlation was found between the UV absorptivity of the tags at the excitation wavelengths typical for UV-MALDI and the magnitude of the signal enhancement. Interestingly, peptides labeled with Alexa Fluor 350, a coumarin derivative containing a sulfo group (i.e., bearing strong negative charge), showed a 5-15-fold increase in intensity of MALDI MS signal in the positive ion mode, relative to the underivatized peptides, when 2,5-DHAP was used as the matrix. The Alexa Fluor 350 tag yielded a significantly higher signal relative to that for the CAF tag, likely due to the increased hydrophobicity of the coumarin structure. With 2,5-DHB, a decrease of MALDI MS signal was observed for all coumarin-labeled peptides, again relative to the unlabeled species. These findings support the hypothesis that derivatization with coumarin, a relatively hydrophobic structure, improves incorporation of hydrophilic peptides into hydrophobic MALDI matrixes, such as CHCA and 2,5-DHAP. PMID- 15283602 TI - A reversible gel for chiral separations. AB - The use of a guanosine gel as a chiral selector in capillary electrophoresis is introduced. Guanosine gels are reversible organized media that are formed through the self-association of guanosine compounds. Their degree of organization and their physicochemical properties can be modulated through variations in guanosine monomer concentration, pH, temperature, and cation content. Baseline resolution of the d and l enantiomers of propranolol was achieved using a reversible biogel formed by 5'-guanosine monophosphate as the run buffer in capillary electrophoresis. Conditions were optimized to provide enantiomeric resolution of 2.1-2.3 in less than 5 min. The reversibility of the gel network offers potential advantages for chiral separations, including the possibility of using thermal or chemical dissociation of the gel network to remove the nucleoside monomers from the separated enantiomers, thereby eliminating the chiral selector as a source of physical contamination of the enantiomerically pure products and spectral background in UV absorbance detection. PMID- 15283603 TI - Hydration of halide anions in ion-exchange resin and their dissociation from cationic groups. AB - The local structures of Cl- and Br- in an anion-exchange resin have been investigated by X-ray absorption fine structure (XAFS). The resins, which have been equilibrated under various partial water vapor pressures to allow the anions to have various hydration numbers, are provided for XAFS measurements. The XAFS spectra indicate that two scattering groups around the counteranion are present, that is, water molecules and an ion-exchange group. Regression analyses allow the separation of the contributions from these two scattering groups; thus, the average hydration number (N) is determined. The hydration number linearly increases with increasing the number of water molecules (n) adsorbed by an ion exchange pair (an ion-exchange group and a counteranion) until the ion-exchange pair adsorbs ca. 3 water molecules, indicating that all of the adsorbed water molecules coordinate the counteranion. However, an increase in N with increasing n becomes small as n exceeds 3; N finally reaches 3.9 (+/-0.4) for Cl- and ca. 3.4 (+/-0.5) for Br-. Detailed studies of the water adsorption isotherms imply that the maximum hydration number of these anions is three when they are bound by the ion-exchange groups, and as more water molecules are supplied, they are dissociated from the ion-exchange groups; ca. 40% of total counteranions are dissociated from the ion-exchange groups. PMID- 15283604 TI - Solid-phase microextraction coupled to gas chromatography/mass spectrometry for determining polycyclic aromatic hydrocarbon-micelle partition coefficients. AB - Solid-phase microextraction (SPME) coupled to gas chromatography with MS detection has been employed to study the partition coefficients of PAHs to ionic and nonionic micelles. The results obtained in this work for seven PAHs, using 85 microm polyacrylate- and 100-microm poly(dimethylsiloxane)-coated fibers and anionic (sodium dodecyl sulfate), cationic (cetyltrimethylammonium bromide), and nonionic (polyoxyethylene-10-lauryl ether) surfactants, indicate that SPME is a viable method for estimating the partition coefficients of PAHs to micelle. The procedure could also be potentially extended to the measurement of partition coefficients between a wide variety of semi- or nonvolatile compounds and micellar media. PMID- 15283605 TI - Smart polyoxometalate-based nitrogen monoxide sensors. AB - An electrochemical sensor design for selective NO detection is presented based on a polyoxometalate (POM) cluster immobilized on an electrode through a polyelectrolyte matrix. It is suggested that the POM can electrocatalyze the reduction of NO. The reduction current is proportional to the NO concentration in the investigated concentration window ranging from 1 nM to 10 microM. The sensitivity of the device can be adjusted by the number of immobilized layers. The response to possible interfering reagents such as nitrate and nitrite can be controlled through the multilayer design. By a predominant negatively charged outer surface, the response to these ions is markedly reduced. PMID- 15283606 TI - Use of room temperature ionic liquids in gas sensor design. AB - The attainable steady-state limiting currents and time responses of membrane covered and membrane-independent gas sensors incorporating different electrode and electrolyte materials have been compared. A new design comprising a membrane free microelectrode modified with a thin layer of a room temperature ionic liquid is considered. While the use of ionic liquid as electrolyte eliminates the need for a membrane and added supporting electrolyte, the slower diffusion of analyte within the more viscous medium results in slower time responses. Such sensors do, however, have potential application in more extreme operating conditions, such as high temperature and pressure, where traditional solvents would volatise. PMID- 15283607 TI - Electrophoretic concentration of proteins at laser-patterned nanoporous membranes in microchips. AB - Laser-patterning of nanoporous membranes at the junction of a cross channel in a microchip is used to integrate protein concentration with an electrokinetic injection scheme. Upon application of voltage, linear electrophoretic concentration of charged proteins is achieved at the membrane surface because buffer ions can easily pass through the membrane while proteins larger than the molecular weight cutoff of the membrane (>5700) are retained. Simple buffer systems can be used, and the concentration results constitute outward evidence that the uniformity of buffer ion concentration is maintained throughout the process. Local and spatially averaged concentration are increased by 4 and 2 orders of magnitude, respectively, upon injection with moderate voltages (70-150 V) and concentration times (100 s). The degree of concentration is limited only by the solubility limit of the proteins. The porous polymer membrane can be used repeatedly as long as care is taken to avoid protein precipitation. PMID- 15283609 TI - Records--the Achilles' heel of school nursing: answers to bothersome questions. AB - This article addresses practice issues related to school health records and school nursing documentation. Because the issues have been posed by practicing school nurses, the article is in Question and Answer (Q and A) format. Specifically, the questions addressed concern the following: ownership and storage location of student health records when the school nurse is contracted from a community health agency rather than employed by the school district; documentation of sensitive health information on students' health records including pregnancy, drug and alcohol abuse, mental illness, history of suicide attempt, and HIV status; inclusion of medical diagnoses and current medications on a student's Individual Educational Program (IEP); and Health Insurance Portability and Accountability Act (HIPAA)-permitted communications between school nurses and health care providers related to students' immunization status, regarding a student's treatment needs in school, and via facsimile (e.g., records of immunizations, completed physical examination forms, and medical orders). HIPAA, the Family Educational Records and Privacy Act (FERPA), and other laws are addressed as appropriate, and resources for obtaining further information are included. PMID- 15283610 TI - Data integrity: beware of viruses. AB - School nurses and health office employees are the creators and caretakers of legal documentation. School nurses have an ethical and legal obligation to protect the integrity of electronic student health records. Although there are many threats to data integrity, from inadequate hardware to electrical surges, one of the most pervasive threats to data is computer viruses. There are many precautions that can be taken to protect electronic student health data from viruses in the school health office. PMID- 15283611 TI - School nurses' perceptions of empowerment and autonomy. AB - The purpose of this study was to explore Kanter's Theory of Structural Power in Organizations, using school nurses and to answer the research question of whether there is a relationship between empowerment and autonomy in school nurses. This study found a positive relationship between the nurses' perceptions of empowerment and autonomy. The school nurses surveyed perceived themselves to have a high degree of autonomy and a moderate degree of empowerment, and they reported that their access to informal power structures was higher than their access to formal power structures in their school systems. School nurses can benefit by understanding factors that can increase their empowerment in the workplace. They need to understand the organizational structure of their workplace to increase their effectiveness and job satisfaction. PMID- 15283612 TI - The impact of homelessness on the health of families. AB - Qualitative research using the symbolic interactionism framework and grounded theory methodology was employed to discover the perceived health problems and dangers that homeless families with children endure. Data were collected using semistructured interviews from 34 homeless volunteer participants with 87 children. An in-depth analysis of the data using the constant comparative method led to recurrent descriptors and patterns, which were synthesized under four themes: external locus of control, deterrents to health, economic barriers, and lack of support. The findings indicate that there is an urgent need for preventive approaches to alleviate homelessness and its attendant health problems. Recommendations consistent with current federal, state, and local strategies and plans for the early identification and prevention of homelessness are presented. School nurses have an important role as advocates, health educators, and coordinators to promote realistic strategies, programs, and policies in the delivery of services to homeless families. PMID- 15283613 TI - Teens' knowledge of risk factors for sports injuries. AB - Youth participation in sports has increased greatly over the past 20 years. Consequently, there has been a rise in the number of sports injuries. A study was conducted to determine teen's level of physical activity, knowledge about risk factors for sports injuries, use of protective equipment, and parental involvement. Two groups of teens, one of which was required to take a physical education class, were given a self-administered, written survey. The study found that the teens in this small Virginia town have a high level of involvement in sports and other physical activity and good general knowledge of sports injury prevention. Improvement is needed in the use of protective equipment when participating in informal sports activities and in the provision of sports injury prevention education to parents. As advocates for student health, school nurses are in a unique position to educate students, parents, staff, and the community about prevention of sports-related injuries. PMID- 15283614 TI - Identifying and intervening in relational aggression. AB - Chronic victimization by bullies has been associated with academic failure in adolescence, as well as adjustment difficulties, depression, and suicidal ideation. Relational aggression is a form of bullying that is a problem for adolescent girls. It often takes the form of damaging peer relationships and includes verbal assaults such as teasing or name calling, as well as psychological attacks such as gossip, social exclusion, and strategic friendship manipulations. A girl's ability to identify these indirect attacks may be imperative for her to enact an effective defense. Because many students do not recognize relational aggression as a form of bullying, their experiences often go unreported to parents or teachers. School nurses may be the front line of defense. With this in mind, school nurses must be informed about bullying behaviors, equipped to identify these behaviors, and prepared to intervene with victims as well as perpetrators of bullying. PMID- 15283615 TI - Increasing immunization compliance. AB - School nurses often have the responsibility to ensure that students meet all immunization requirements for school entry and school attendance. In large inner city school districts, many obstacles exist which make this task daunting and often result in lengthy absences and exclusions for students. It is critical that school nurses find creative and systematic ways to meet these challenges, which include working parents, lack of access to primary care, lack of transportation, cost of immunizations, poor compliance and follow-up, myths regarding immunizations, and the impact of the Health Insurance Portability and Accountability Act (HIPAA) legislation. This article describes an immunization project that removed most of those barriers for high-risk students and gave the school nurses tools to succeed in achieving higher levels of immunization compliance in inner-city schools. Since the immunization project's conception, compliance in the district has risen from an overall level of 50-60% to 90-100%, along with better record-keeping and the prevention of exclusions. PMID- 15283616 TI - The female athlete triad. AB - The Female Athlete Triad is a syndrome of the interrelated components of disordered eating, amenorrhea, and osteoporosis. Sometimes inadvertently, but more often by willful dietary restriction, many female athletes do not ingest sufficient calories to adequately fuel their physical or sport activities, which can disrupt menstrual functioning, thereby increasing their risk of bone loss. Although its prevalence is unknown, the Female Athlete Triad is believed to affect many athletes at all ages and all sport competition levels. Even though the Triad affects athletes in all sports, girls and women in sports that emphasize a thin or small body size or shape appear to be most at risk. This article focuses on the risks of the Female Athlete Triad for middle- and high school-age female athletes as well as the unique issues related to the identification, management, and treatment of the various components of the Triad in this special adolescent subpopulation. PMID- 15283617 TI - Children's mental health and school success. AB - An integrative review of literature was undertaken to examine the impact of children's mental health on their school success. The literature confirmed a confluence of problems associated with school performance and child and adolescent mental health. Poor academic functioning and inconsistent school attendance were identified as early signs of emerging or existing mental health problems during childhood and adolescence. Among the goals of school nursing is to provide a process for identification and resolution of students' health needs as they affect educational achievement. Thus, it is within the scope of practice and goals of school nursing to also address children's mental health needs, as they affect school performance. This review of literature supports the conclusion that school nursing is well positioned to respond to the need for mental health promotion, illness prevention, and early intervention related to children's mental health. PMID- 15283618 TI - Starting a journal club. PMID- 15283619 TI - Lumps, bumps, and things that go itch in your office! PMID- 15283620 TI - Chronic fatigue syndrome: lack of association between pain-related fear of movement and exercise capacity and disability. AB - BACKGROUND AND PURPOSE: Patients who experience pain, a symptom of chronic fatigue syndrome (CFS), often exhibit kinesiophobia (irrational fear of movement). The purpose of this study was to examine whether pain-related fear of movement is associated with exercise capacity, activity limitations, or participation restrictions in patients with CFS who experience widespread pain. SUBJECTS AND METHODS: Sixty-four subjects met the inclusion criteria. All subjects fulfilled the 1994 Centers for Disease Control and Prevention case definition for CFS and experienced widespread myalgias or arthralgias. The subjects completed the Tampa Scale for Kinesiophobia-Dutch Version (TSK-DV) and the Dutch Chronic Fatigue Syndrome-Activities and Participation Questionnaire (CFS-APQ). They then performed a maximal exercise test on a bicycle ergometer. Heart rate was monitored continuously by use of an electrocardiograph. Ventilatory factors were measured through spirometry. Correlations between the TSK-DV scores and both the exercise capacity data and the CFS-APQ scores were assessed using the Spearman rank correlation coefficient. Using the Mann-Whitney U test, the TSK-DV scores were compared between subjects who performed a maximal exercise stress test and those who did not perform the test. RESULTS: Forty-seven subjects (73.4%) attained a total score of greater than 37 on the TSK-DV, indicating high fear of movement. Neither the exercise capacity data nor the CFS APQ scores indicated a correlation with the TSK-DV scores (n=64). Subjects who did not perform a maximal exercise capacity test had more fear of movement (median TSK-DV score=43.0, interquartile range=10.3) compared with those who did perform a maximal exercise capacity test (median TSK-DV score=38.0, interquartile range=13.2; Mann-Whitney U-test score=322.5, z=-1.974, P=.048), but the correlation analysis was unable to reveal an association between exercise capacity and kinesiophobia in either subgroup. DISCUSSION AND CONCLUSION: These results indicate a lack of correlation between kinesiophobia and exercise capacity, activity limitations, or participation restrictions, at least in patients with CFS who are experiencing widespread muscle or joint pain. PMID- 15283621 TI - Predictors of exercise behavior in patients with rheumatoid arthritis 6 months following a visit with their rheumatologist. AB - BACKGROUND AND PURPOSE: When factors that influence exercise behavior are known, health care professionals can more likely design and modify patient education materials targeted to promote exercise behavior. This study aimed to identify predictors of exercise behavior in patients with rheumatoid arthritis 6 months after a visit with their rheumatologist. SUBJECTS AND METHODS: Twenty-five rheumatologists and 132 patients with rheumatoid arthritis participated. One hundred thirteen patients (85.6%) completed the 6-month follow-up. Rheumatologists and patients completed baseline questionnaires and were audiotaped during a subsequent visit. Physical function and exercise behavior were ascertained via questionnaire 6 months following the visit. Multivariate logistic regression identified predictors of exercise behavior at 6 months. Eighty-nine patients (79%) were female. The average age was 54.8 years (SD=14.4, range=20-94). The mean duration of illness was 9.8 years (SD=8.7, range= <1-35). Patients were moderately impaired (mean Medical Outcomes Study 36-Item Short-Form Health Survey [SF-36] function score=49.3, SD=27.5). Thirty-four patients (27%) were exercising 6 months after visiting their rheumatologist. More than 50% of the rheumatologists had 5 or more years of clinical experience, 18 (72%) were male, and 10 (42%) reported they exercised regularly. RESULTS: Predictors of exercise behavior at 6 months were patients' past history of exercise (odds ratio=6.8, 95% confidence interval=3.1-15) and rheumatologists' current exercise behavior (odds ratio=0.26, 95% confidence interval=0.09-0.77). DISCUSSION AND CONCLUSION: Patients were nearly 7 times more likely to exercise 6 months after visiting their rheumatologist if they participated in exercise in the past. If a patient's rheumatologist was currently performing aerobic exercise, the patient was 26% more likely to be engaged in exercise at follow-up. These data may be useful in understanding patient motivation to participate in exercise. PMID- 15283622 TI - Diagnosis of lower-extremity deep vein thrombosis in outpatients with musculoskeletal disorders: a national survey study of physical therapists. AB - BACKGROUND AND PURPOSE: Prompt identification of outpatients who may have proximal lower-extremity deep vein thrombosis (PDVT) is important, in part, because of the risk of pulmonary embolism. The purposes of our study were to determine the degree of accuracy of physical therapists' estimates of the probability of PDVT in hypothetical patient vignettes and to determine whether physical therapists would contact the referring physician about the hypothetical patients' condition as recommended in published evidence. SUBJECTS AND METHODS: A survey instrument consisting of 6 vignettes was sent to a nationally representative random sample of 1,500 physical therapists. The clinical decision rule developed by Wells and colleagues served as the gold standard for PDVT probability. RESULTS: A total of 969 (65% response rate) physical therapists completed the survey. We found no evidence of nonresponse bias. For the 2 high probability vignettes, 87% and 64% of the physical therapists underestimated the probability of PDVT. For the 2 high-probability cases, 32% and 27% of the physical therapists reported that they would not have contacted the referring physician. For the 2 moderate-probability cases, 15% and 30% of the physical therapists would not have contacted the referring physician. Therapist experience, certification status, place of practice, and region of the country did not explain the findings. DISCUSSION AND CONCLUSION: The care of outpatients who are at risk for PDVT could potentially be improved by use of the clinical decision rule developed by Wells and colleagues, although more study is warranted. PMID- 15283623 TI - Diagnosis of lower-extremity deep vein thrombosis in outpatients. PMID- 15283624 TI - Exercise as an intervention for cancer-related fatigue. PMID- 15283625 TI - What is the evidence regarding specific methods of pelvic floor exercise for a patient with urinary stress incontinence and mild anterior wall prolapse? PMID- 15283626 TI - Laparoscopic cholecystectomy and outpatient surgery. PMID- 15283627 TI - Quality of life (GIQLI) and laparoscopic cholecystectomy usefulness in patients with gallbladder dysfunction or chronic non-lithiasic biliary pain (chronic acalculous cholecystitis). AB - OBJECTIVE: The aim of this study was to evaluate the incidence, clinical features and role of laparoscopic cholecystectomy (LC) in patients with chronic acalculous cholecystitis (CAC) in comparison with a control group of patients who underwent cholecystectomy for chronic calculous cholecystitis (CCC). MATERIAL AND METHODS: Prospective evaluation of 34 patients with CAC in contrast with 297 patients with CCC. OUTCOME MEASURES: Clinical presentation, quality of life using the Gastrointestinal Quality of Life Index (GIQLI), usefulness derived from the therapeutic procedure as measured in quality of life units by GIQLI, and clinical efficacy at one year of follow-up. RESULTS: The incidence of complicated biliary disease was higher in CAC (27%), in comparison with CCC (13.8%). The histological study of the excised gallbladder revealed a higher incidence of cholesterolosis associated with chronic cholecystitis in the CAC group (64.9%). GIQLI showed significant differences between preoperative and postoperative measurements in both groups. The associated usefulness of LC was similar in both groups (73 versus 67.3 percent), confirming an important increase in quality of life for both categories. CONCLUSIONS: The incidence of CAC is 11 per cent with a high association with cholesterolosis. Quality of life and LC usefulness are similar to those of patients with CCC. Due to the fact that cholecistogammagraphy is a technique not available in daily clinical practice, and that oral cholecystography and dynamic ultrasound are reliable when a positive result is obtained, extended clinical evaluation is still the most reliable indicator for cholecystectomy. PMID- 15283630 TI - Gastrointestinal motility and perception disorders re-visited. PMID- 15283628 TI - Laparoscopic cholecystectomy in the treatment of biliary lithiasis: outpatient surgery or short stay unit? AB - OBJECTIVE: Analysis of clinical and surgical factors in a series of patients subjected to laparoscopic cholecystectomy in an outpatient unit and their relationship with time of discharge and patient acceptance. PATIENTS AND METHOD: Eighty one consecutive patients underwent to elective laparoscopic cholecystectomy during year 2002 within S.A.S. (Andalusian Health Service) from a surgical waiting list. Retrospective and comparative study between two groups: group A includes patients discharged between 24 and 48 hours after intervention; group B includes patients discharged in less than 24 hours. We analyse the clinical and surgical characteristics and post-operative outcome of both groups of patients. RESULTS: Group A was composed of 53 patients and group B of 28 patients. Factors of clinical significance which determined discharge after 24 hours included: early post-surgical incidences or complications (p = 0.017), inability to tolerate oral diet (p = 0.002), and doubts and feelings insecurity of patients regarding discharge by traditional means 62.3% (p = 0.0003). CONCLUSIONS: Outpatient laparoscopic cholecystectomy is a safe and reliable procedure with a high acceptance rate and few complications. Perhaps traditional culture has to be changed to obtain better results. PMID- 15283631 TI - Hypertransaminasemia in patients with negative viral markers. PMID- 15283632 TI - Experience with granulocytapheresis in Crohn's disease. AB - OBJECTIVE: To describe our experience with granulocyte apheresis to induce remission in patients with active Crohn's disease refractory to conventional treatment. We summarize the results previously obtained with this technique. CONCLUSIONS: Granulocyte apheresis is a safe and well tolerated therapeutic modality that can be a valid therapeutic alternative in the induction of remission in inflammatory bowel disease, although controlled clinical trials must be conducted to define long-term efficacy, as well as to establish "optimal patient" selection, re-treatment interval, and number of sessions. PMID- 15283633 TI - Hereditary hyperferritinemia-cataract syndrome. Study of a new family in Spain. AB - The hyperferritinemia-cataract syndrome, inherited as a Mendelian dominant trait, is due to mutations in the 5' non-coding region of the ferritin light chain gene that modifies the shape of the IRE (iron responsive element) region, which loses its normal function of regulating the synthesis of ferritin light chains. Excess of light chains results in complexes that accumulate into the lens giving rise to early cataracts. We present a Spanish family with seven affected members through three generations. A genetic study reveals a substitution of a single base (C- >T) at position 33 in the IRE sequence in the index case and in one affected brother, whereas a non-affected sister shows the normal sequence. The hyperferritinemia-cataract syndrome was identified in 1995 and is still poorly understood. Clinicians should suspect it when treating any subject with early cataracts, even more if they are familial, or in patients with very high levels of ferritinemia without evidence of iron overload. There are no known consequences of the syndrome other than cataracts, and its proper diagnosis carries a favorable prognosis and eliminates the risk of unnecessary phlebotomies. PMID- 15283634 TI - Apoptosis: a rapid and silent form of death. PMID- 15283635 TI - [Haemoperitoneum: unusual complication of the gallbladder perforation]. PMID- 15283636 TI - [Liver transplant in patients co-infected with hepatitis C virus and human immunodeficiency virus]. PMID- 15283637 TI - [Prevalence and risk factors associated to malnutrition in elderly inpatients]. PMID- 15283638 TI - [Prevalence and risk factors associated to malnutrition in elderly inpatients]. AB - BACKGROUND: Malnutrition is a frequent clinical finding in elderly inpatients that is deleterious to the physiologic function of several body's organs or systems that is accompanied of an increase of the length of hospitalization, economic burden and mortality. Protein-energy malnutrition risk factors in elderly patients are not well defined. METHOD: A transversal study of inpatients over 70 years in a internal medicine ward was undertaken in order to know the protein-energy malnutrition's prevalence a possible risk factors. It was considered that a patient suffered from had protein-energy malnutrition if he or she had a tricipital skinfold thickness or an arm circumference under 10th percentile and/or had low levels of two of the following clinical parameters: plasmatic albumin (under 3.5 mg/100 ml). Plasmatic transferrin (under 150 mg/100 ml) or a lymphocytic cell count (under 1,500 cells/ml). RESULTS: A hundred and five patients were included. Mean age was 83.0 +/- 6.4 years with a predominance of the female sex (61%). Thirty-three patients (31%) were sent from nursing homes. A 35% were very dependent and frail elderly patients. Fifty-eight patients presented at hospital with protein-energy malnutrition (prevalence = 57.1%; CI 95% 47.1%-66.8%). Chronic obstructive pulmonary disease (COPD) and a acute infectious disease were independent risk factors associated protein-energy malnutrition (prevalence ratio of 1.4 and 0.5, respectively). CONCLUSIONS: Protein-energy malnutrition's prevalence in elderly inpatients is very important. Infectious diseases and COPD are positively and negatively associated with protein-energy malnutrition. PMID- 15283639 TI - [Cost distribution of highly active antiretroviral therapy]. AB - The efficacy of highly active antiretroviral therapy (HAART) implies a high pharmaceutical cost. Deficient patient adhesion to therapy means that not all individuals benefit from treatment, while some patients are in non-advanced stages of the disease where such management may not be necessary. A study is made of the cost distribution of HAART. MATERIAL AND METHODS: An analysis was made of 199 HIV-infected adults subjected to HAART at least once during 2001. The medication history was used to calculate the cost of HAART in the global group, in the patients who abandoned treatment (group A), in the group in which viral load was not controlled (group B), and in the patients starting with or reaching a CD4+ count > 500 cells/mm3 (groups C1 and C2, respectively). RESULTS: The global annual cost of HAART (discounting returned medication) was 961,720 Euro (21.3% of the global hospital pharmacological expenditure), group A accounting for 6.3%, group B 14.26%, and subgroups C1 and C2 for 12.1 and 8.1, respectively. COMMENTS: In our center, almost 40% of antiretroviral drug expenditure may be considered of little use. PMID- 15283640 TI - [Impact of the long active beta-2 agonists inhaled therapy on the quality of life in asthmatic patients]. AB - OBJECTIVE: To assess the impact of the addition of inhaled therapy with long acting beta2-agonists (formoterol) on the health related quality of life (HRQL) in asthmatic patients treated with inhaled corticosteroids and poorly controlled. METHODS: Observational prospective naturalistic study with a cohort of 343 ambulatory patients during 6 months. We collected socio-demographic, clinical and lung function variables. The HRQL was assessed by means of a generic scale (SF 36) and a specific one (Marks's AQLQ). RESULTS: They were evaluated 268 patients (78%). mean age 48.6 years (SD 16.28), 64.15% women. Women and the subjects with worse socioeconomic situation had poor basal HRQL scores. During the follow-up an average improvement in FEV1 of 0.26 l (SD 0.44; p <0.001) was observed. The average change in the AQLQ was -1.92 (-2.09-1.75) being the larger improvement in the breathlessness domain. The average change in SF-36 was 5.13 (4.75-5.51) for physical standardized component and 5.23 (4.83-5.63) for mental component. The effect size was "large" with the measurement of the AQLQ and "moderate" with SF 36. The AQLQ and SF-12 scores are strongly correlated. basally and after the treatment (r > 0.5). CONCLUSION: To add long acting beta2-agonists to the asthmatic patients treated with inhaled corticosteroids and poorly controlled, under usual clinical practice conditions, is associated with an improvement of the HRQL perception measured by means of generic and specific instruments. PMID- 15283641 TI - [Inflammatory pseudotumor of the spleen. Review and a new case report]. AB - Splenic inflammatory pseudotumour has been defined as a benign tumour reactive lesion with unspecified reparative or inflammatory alterations. It is a mimicking tumour often confused with other pathologies, primarily, lymphoproliferative disorders. Though its aetiology remains unknown, these tumours are often associated to infectious agents such as the Epstein-Barr virus. We describe a case of a 29 year-old patient with a history of infectious mononucleosis, followed by fever, weight loss, and splenomegaly. Splenectomy detected a homogeneous, whitish 2 cm node diagnosed as splenic inflammatory pseudotumour. The clinical features and diagnosis of this disorder are discussed in relation to the histopathological findings. PMID- 15283642 TI - [Sinistral portal hypertension with bleeding gastric varices as initial manifestation of renal-cell carcinoma]. AB - We report a clinic case of renal-cell carcinoma presenting as sinistral portal hypertension; a clinical syndrome consisting of esplenic vein thrombosis manifested as isolated gastric varices with patent portal vein and normal hepatic function. The most frequent cause of this syndrome is pancreatic pathology. Renal cell carcinoma is characterized by a wide variety of symptoms as initial manifestation. In our case, the patient developed a massive gastrointestinal bleeding secondary to isolated gastric varices caused by splenic vein thrombosis due to extrinsic compression by a hypernephroma that infiltrated the pancreas. PMID- 15283643 TI - [Diffuse erythema with lung and inguinal masses as the initial manifestation of a large cell neuroendocrine tumor of the lung]. AB - Large cell neuroendocrine carcinoma of the lung is defined as poorly differentiated and high-grade neuroendocrine tumor that is morphologically and biologically between atypical carcinoid and small cell lung carcinoma. The prognosis of this type of tumor is poor, specially in advanced disease. We report on a case with atypical presentation, with high blood levels of histamine as a previously unreported association, and IV stage, in which the diagnosis was made after biopsy of an inguinal mass. PMID- 15283644 TI - [Lactic acidosis in diabetic patient treated with metformin]. AB - We present a case of metabolic acidosis in a man, recently diagnosed with type 2 diabetes mellitus under treatment with metformin. Metformin (along with Fenformin and Butformin) is an oral antihyperglycemic agent belonging to the biguanide group employed in the treatment of non insulin dependent diabetes (NIDDM). Its main use is in association with other oral agents in obese diabetic patients with difficult metabolic control. In some of these patients, clearly beneficial developed lactic acidosis, specially in those who have predisposing factors (respiratory failure, liver disease or cardiovascular disease) and/or those who require high doses. For this reason we describe its pharmacokinetics, therapeutic indications and its correct use in this type of diabetic patient. PMID- 15283645 TI - [Perioperative management of chronic medications not related with surgical procedures]. AB - The correct management of chronic medications not related with surgical procedures in the perioperative period has a relevant place because each year millions of patients around the world undergo surgical procedures. For this reason the assistencial team should be aware of the importance of continuate or discontinuate determinate drugs during perioperative period because some of them are considered an important risk factor in the development of complications. The key is to differentiate necessary from unnecessary medication. This is a complex aspect, little studied, which difficult clinical decisions and favours the coexistence of several trends of clinical practice. The purpose of this review is to describe the factors that determinate the continuity or suspension of chronic medications which are not related with surgery in the perioperative period and to provide practice recommendations in lights of available publications. PMID- 15283646 TI - [Internal Medicine: present and future]. PMID- 15283647 TI - [Hydropneumothorax in an immunocompetent patient]. PMID- 15283648 TI - [Pharyngeal infection by Mycoplasma pneumoniae in a young man. A case report]. PMID- 15283649 TI - [Liver toxicity associated with flutamide]. PMID- 15283650 TI - [Pleural effusion as a manifestation of ovarian hyperstimulation syndrome, after treatment with triptoreline for uterus myomata]. PMID- 15283651 TI - [Metabolic syndrome prevalence in type 2 diabetic patients]. PMID- 15283652 TI - [Liver infiltrative model and bone pain]. PMID- 15283653 TI - [Herpes simplex esophagitis associated to Candida albicans in an immunocompetent host]. PMID- 15283654 TI - [Experience of thyroid carcinoma in a town hospital: review of 21 cases]. PMID- 15283655 TI - [Is there complete permeability in the venous thrombosis when the anticoagulant treatment finishes in the deep venous thrombosis?]. PMID- 15283656 TI - [Consultation-liaison Internal Medicine]. PMID- 15283657 TI - The accidental plant pathologist. AB - This article presents the experiences of a woman in academic plant pathology from the 1950s to today. Topics include the social climate for women in science, personal and professional developments and research discoveries, public policy issues in agriculture and biotechnology affecting plant pathology, and projections for the future of plant pathology. PMID- 15283658 TI - Tobacco mosaic virus: a model system for plant biology. AB - Tobacco mosaic virus (TMV) has had an illustrious history for more than 100 years, dating to Beijerinck's description of the mosaic disease of tobacco as a contagium vivum fluidum and the modern usage of the word "virus." Since then, TMV has been acknowledged as a preferred didactic model and a symbolic model to illuminate the essential features that define a virus. TMV additionally emerged as a prototypic model to investigate the biology of host plants, namely tobacco. TMV also exemplifies how a model system furthers novel, and often unexpected, developments in biology and virology. Today, TMV is used as a tool to study host pathogen interactions and cellular trafficking, and as a technology to express valuable pharmaceutical proteins in tobacco. The history of TMV illustrates how pragmatic strategies to control an economically important disease of tobacco have had unexpected and transforming effects across platforms that impinge on plant health and public health. PMID- 15283659 TI - Assessment and management of soil microbial community structure for disease suppression. AB - Identification of the biological properties contributing to the function of suppressive soils is a necessary first step to the management of such systems for use in the control of soilborne diseases. The development and application of molecular methods for the characterization and monitoring of soil microbial properties will enable a more rapid and detailed assessment of the biological nature of soil suppressiveness. Although suppressive soils have provided a wealth of microbial resources that have subsequently been applied for the biological control of soilborne plant pathogens, the full functional capabilities of the phenomena have not been realized in production agricultural ecosystems. Cultural practices, such as the application of soil amendments, have the capacity to enhance disease suppression, though the biological modes of action may vary from that initially resident to the soil. Plants have a distinct impact on characteristics and activity of resident soil microbial communities, and therefore play an important role in determining the development of the disease suppressive state. Likewise, plant genotype will modulate these same biological communities, and should be considered when developing strategies to exploit the potential of such a natural disease control system. Implementation of consistently effective practices to manage this resource in an economically and environmentally feasible manner will require more detailed investigation of these biologically complex systems and refinement of currently available methodologies. PMID- 15283660 TI - Analysis of disease progress as a basis for evaluating disease management practices. AB - The relationship between epidemiology and disease management is long-standing but sometimes tenuous. It may seem self-evident that improved understanding of epidemic processes will lead to more effective control practices but this remains a testable proposition rather than demonstrated reality. A wide range of models differing in mathematical sophistication and computational complexity has been proposed as a means of achieving a greater understanding of epidemiology and carrying this through to improved management. The potential exists to align these modeling approaches to evaluation of control practices and prediction of the consequent epidemic outcomes, but these have yet to make a major impact on practical disease management. For the immediate future simpler pragmatic approaches for analysis of disease progress, using nonlinear growth functions and/or integrated measures such as area under disease progress curves, will play a key role in informing tactical and strategic decisions on control treatments. These approaches have proved useful in describing control effectiveness and, in some cases, optimizing or changing control practices. PMID- 15283661 TI - Evolution of plant parasitism among nematodes. AB - Despite extraordinary diversity of free-living species, a comparatively small fraction of nematodes are parasites of plants. These parasites represent at least three disparate clades in the nematode tree of life, as inferred from rRNA sequences. Plant parasites share functional similarities regarding feeding, but many similarities in feeding structures result from convergent evolution and have fundamentally different developmental origins. Although Tylenchida rRNA phylogenies are not fully resolved, they strongly support convergent evolution of sedentary endoparasitism and plant nurse cells in cyst and root-knot nematodes. This result has critical implications for using model systems and genomics to identify and characterize parasitism genes for representatives of this clade. Phylogenetic studies reveal that plant parasites have rich and complex evolutionary histories that involve multiple transitions to plant parasitism and the possible use of genes obtained by horizontal transfer from prokaryotes. Developing a fuller understanding of plant parasitism will require integrating more comprehensive and resolved phylogenies with appropriate choices of model organisms and comparative evolutionary methods. PMID- 15283662 TI - Lessons learned from the genome analysis of ralstonia solanacearum. AB - Ralstonia solanacearum is a devastating plant pathogen with a global distribution and an unusually wide host range. This bacterium can also be free-living as a saprophyte in water or in the soil in the absence of host plants. The availability of the complete genome sequence from strain GMI1000 provided the basis for an integrative analysis of the molecular traits determining the adaptation of the bacterium to various environmental niches and pathogenicity toward plants. This review summarizes current knowledge and speculates on some key bacterial functions, including metabolic versatility, resistance to metals, complex and extensive systems for motility and attachment to external surfaces, and multiple protein secretion systems. Genome sequence analysis provides clues about the evolution of essential virulence genes such as those encoding the Type III secretion system and related pathogenicity effectors. It also provided insights into possible mechanisms contributing to the rapid adaptation of the bacterium to its environment in general and to its interaction with plants in particular. PMID- 15283663 TI - Management and resistance in wheat and barley to fusarium head blight. AB - Fusarium head blight (FHB) is a devastating disease of wheat and barley worldwide. Resistant cultivars could reduce damage from FHB. Chinese wheat cultivar Sumai 3 and its derivatives represent the greatest degree of resistance to FHB known. A major quantitative trait locus (QTL) on chromosome 3BS and other minor QTL for FHB resistance have been identified in these cultivars and used in wheat-breeding programs worldwide. Many breeding lines with the 3BS resistance QTL and improved agronomic traits have been developed. In barley, only limited sources of FHB resistance are available, especially in six-rowed barley, and none of them contains a DON level low enough to meet the safety requirement of the brewing industry. Several QTL have been identified for lower FHB severity, DON content, and kernel discoloration and used to enhance FHB resistance in barley. Marker-assisted selection for FHB resistance QTL on 3BS of wheat and on 2H of barley is in progress. PMID- 15283664 TI - Comparative genomics analyses of citrus-associated bacteria. AB - Xylella fastidiosa 9a5c (XF-9a5c) and Xanthomonas axonopodis pv. citri (XAC) are bacteria that infect citrus plants. Sequencing of the genomes of these strains is complete and comparative analyses are now under way with the genomes of other bacteria of the same genera. In this review, we present an overview of this comparative genomic work. We also present a detailed genomic comparison between XF-9a5a and XAC. Based on this analysis, genes and operons were identified that might be relevant for adaptation to citrus. XAC has two copies of a type II secretion system, a large number of cell wall-degrading enzymes and sugar transporters, a complete energy metabolism, a whole set of avirulence genes associated with a type III secretion system, and a complete flagellar and chemotatic system. By contrast, XF-9a5c possesses more genes involved with type IV pili biosynthesis than does XAC, contains genes encoding for production of colicins, and has 4 copies of Type I restriction/modification system while XAC has only one. PMID- 15283665 TI - Systemic acquired resistance. AB - Systemic acquired resistance (SAR) is a mechanism of induced defense that confers long-lasting protection against a broad spectrum of microorganisms. SAR requires the signal molecule salicylic acid (SA) and is associated with accumulation of pathogenesis-related proteins, which are thought to contribute to resistance. Much progress has been made recently in elucidating the mechanism of SAR. Using the model plant Arabidopsis, it was discovered that the isochorismate pathway is the major source of SA during SAR. In response to SA, the positive regulator protein NPR1 moves to the nucleus where it interacts with TGA transcription factors to induce defense gene expression, thus activating SAR. Exciting new data suggest that the mobile signal for SAR might be a lipid molecule. We discuss the molecular and genetic data that have contributed to our understanding of SAR and present a model describing the sequence of events leading from initial infection to the induction of defense genes. PMID- 15283666 TI - Molecular aspects of plant virus transmission by olpidium and plasmodiophorid vectors. AB - The genome structures of a large number of viruses transmitted by olpidium and plasmodiophorid vectors have been determined. The viruses are highly diverse, belonging to 12 genera in at least 4 families. Plasmodiophorids are now classified as protists rather than true fungi. This finding, along with the recognition of the great variety of viruses transmitted by olpidium and plasmodiophorid vectors, will likely lead to an elaboration of the details of in vitro and in vivo transmission mechanisms. Recent progress in elucidating the interaction between Cucumber necrosis virus (CNV) and its zoospore vector suggests that specific sites on the capsid as well as on the zoospore are involved in transmission. Moreover, some features of CNV/zoospore attachment are similar to poliovirus/host cell interactions, suggesting evolutionary conservation of functional features of plant and animal virus capsids. PMID- 15283667 TI - Microbial diversity in soil: selection microbial populations by plant and soil type and implications for disease suppressiveness. AB - An increasing interest has emerged with respect to the importance of microbial diversity in soil habitats. The extent of the diversity of microorganisms in soil is seen to be critical to the maintenance of soil health and quality, as a wide range of microorganisms is involved in important soil functions. This review focuses on recent data relating how plant type, soil type, and soil management regime affect the microbial diversity of soil and the implication for the soil's disease suppressiveness. The two main drivers of soil microbial community structure, i.e., plant type and soil type, are thought to exert their function in a complex manner. We propose that the fact that in some situations the soil and in others the plant type is the key factor determining soil microbial diversity is related to the complexity of the microbial interactions in soil, including interactions between microorganisms and soil and microorganisms and plants. A conceptual framework, based on the relative strengths of the shaping forces exerted by plant and soil versus the ecological behavior of microorganisms, is proposed. PMID- 15283668 TI - Microbial dynamics and interactions in the spermosphere. AB - The spermosphere represents a short-lived, rapidly changing, and microbiologically dynamic zone of soil surrounding a germinating seed. It is analogous to the rhizosphere, being established largely by the carbon compounds released into the soil once the seed begins to hydrate. These seed exudations drive the microbial activities that take place in the spermosphere, many of which can have long-lasting impacts on plant growth and development as well as on plant health. In this review, I discuss the nature of the spermosphere habitat and the factors that give rise to its character, with emphasis on the types of microbial activities in the spermosphere that have important implications for disease development and biological disease control. This review, which represents the first comprehensive synthesis of the literature on spermosphere biology, is meant to illustrate the unique nature of the spermosphere and how studies of interactions in this habitat may serve as useful experimental models for testing hypotheses about plant-microbe associations and microbial ecology. PMID- 15283669 TI - Biological control of chestnut blight with hypovirulence: a critical analysis. AB - Most hypovirulence in the chestnut blight fungus, Cryphonectria parasitica, is associated with infection by fungal viruses in the family Hypoviridae. Hypovirulence has controlled chestnut blight well in some locations in Europe and in Michigan in the United States. In contrast, with few exceptions, biological control has failed almost completely in eastern North America. Therapeutic treatment of individual cankers is successful in most cases, but the success of hypovirulence at the population level depends on the natural spread of viruses. Characteristics of three interacting trophic levels (virus, fungus, and tree), plus the environment, determine the success or failure of hypovirulence. Vegetative incompatibility restricts virus transmission, but this factor alone is a poor predictor of biological control. Any factor reducing the rate of chestnut blight epidemics enhances hypovirus invasion. Overall, however, not enough is understood about the epidemiological dynamics of this system to determine the crucial factors regulating the establishment of hypovirulence in chestnut forests. PMID- 15283670 TI - Integrated approaches for detection of plant pathogenic bacteria and diagnosis of bacterial diseases. AB - Disease diagnosis is based on a number of factors, including laboratory tests for pathogen identification. Rapid development of genomic techniques for characterization of bacteria over the past decade has greatly simplified and improved pathogen detection and identification, but DNA-based methods have not yet entirely replaced traditional culture and phenotypic tests in the plant industry. The first section of this review focuses on rapid immunodiagnostic and DNA-based detection methods for known bacterial pathogens in plants or plant products, which often manifest no symptoms of disease. The second section covers the broader topic of disease diagnosis and new methods for identifying and characterizing bacteria. PMID- 15283671 TI - Type III secretion system effector proteins: double agents in bacterial disease and plant defense. AB - Many phytopathogenic bacteria inject virulence effector proteins into plant cells via a Hrp type III secretion system (TTSS). Without the TTSS, these pathogens cannot defeat basal defenses, grow in plants, produce disease lesions in hosts, or elicit the hypersensitive response (HR) in nonhosts. Pathogen genome projects employing bioinformatic methods to identify TTSS Hrp regulon promoters and TTSS pathway targeting signals suggest that phytopathogenic Pseudomonas, Xanthomonas, and Ralstonia spp. harbor large arsenals of effectors. The Hrp TTSS employs customized cytoplasmic chaperones, conserved export components in the bacterial envelope (also used by the TTSS of animal pathogens), and a more specialized set of TTSS-secreted proteins to deliver effectors across the plant cell wall and plasma membrane. Many effectors can act as molecular double agents that betray the pathogen to plant defenses in some interactions and suppress host defenses in others. Investigations of the functions of effectors within plant cells have demonstrated the plasma membrane and nucleus as subcellular sites for several effectors, revealed some effectors to possess cysteine protease or protein tyrosine phosphatase activity, and provided new clues to the coevolution of bacterium-plant interactions. PMID- 15283672 TI - Plant virus satellite and defective interfering RNAs: new paradigms for a new century. AB - Although many subviral RNAs reduce or intensify disease symptoms caused by the helper virus, only recently have clues concerning the mechanism of disease modulation been revealed. New models for DI RNA-mediated reduction in helper virus levels and symptom attenuation include DI RNA enhancement of posttranscriptional gene silencing (PTGS), which is an antiviral defense mechanism in plants. Symptom enhancement by the satRNA of Cucumber mosaic virus is caused by minus-strand induction of the programmed cell death pathway. In contrast, symptom enhancement by satC of Turnip crinkle virus is due to satC interference with virion formation, leading to increased levels of free coat protein, which is the viral suppressor of PTGS. Mutualism between satRNA and helper virus can be seen for the satRNA of Groundnut rosette virus, which contributes to the virus by allowing virion assembly. These novel findings are leading to re-evaluation of the relationships between subviral RNAs, helper viruses, and hosts. PMID- 15283673 TI - Chemical biology of multi-host/pathogen interactions: chemical perception and metabolic complementation. AB - The xenognostic mechanisms of two multi-host pathogens, the causative agent of crown gall tumors Agrobacterium tumefaciens and the parasitic plant Striga asiatica, are compared. Both organisms are general plant pathogens and require similar information prior to host commitment. Two mechanistic strategies, chemical perception and metabolic complementation, are used to ensure successful host commitment. The critical reactions at host-parasite contact are proton and electron transfer events. Such strategies may be common among multi-host pathogens. PMID- 15283674 TI - Two-dimensional IR correlation spectroscopy of mutants of the beta-glycosidase from the hyperthermophilic archaeon Sulfolobus solfataricus identifies the mechanism of quaternary structure stabilization and unravels the sequence of thermal unfolding events. AB - Beta-glycosidase from the hyperthermophilic archaeon Sulfolobus solfataricus is a homotetramer with a higher number of ion pairs compared with mesophilic glycoside hydrolases. The ion pairs are arranged in large networks located mainly at the tetrameric interface of the molecule. In the present study, the structure and thermal stability of the wild-type beta-glycosidase and of three mutants in residues R488 and H489 involved in the C-terminal ionic network were examined by FTIR (Fourier-transform IR) spectroscopy. The FTIR data revealed small differences in the secondary structure of the proteins and showed a lower thermostability of the mutant proteins with respect to the wild-type. Generalized 2D-IR (two-dimensional IR correlation spectroscopy) at different temperatures showed different sequences of thermal unfolding events in the mutants with respect to the wild-type, indicating that punctual mutations affect the unfolding and aggregation process of the protein. A detailed 2D-IR analysis of synchronous maps of the proteins allowed us to identify the temperatures at which the ionic network that stabilizes the quaternary structure of the native and mutant enzymes at the C-terminal breaks down. This evidence gives support to the current theories on the mechanism of ion-pair stabilization in proteins from hyperthermophilic organisms. PMID- 15283675 TI - Evaluation of angiotensin-converting enzyme (ACE), its homologue ACE2 and neprilysin in angiotensin peptide metabolism. AB - In the RAS (renin-angiotensin system), Ang I (angiotensin I) is cleaved by ACE (angiotensin-converting enzyme) to form Ang II (angiotensin II), which has effects on blood pressure, fluid and electrolyte homoeostasis. We have examined the kinetics of angiotensin peptide cleavage by full-length human ACE, the separate N- and C-domains of ACE, the homologue of ACE, ACE2, and NEP (neprilysin). The activity of the enzyme preparations was determined by active site titrations using competitive tight-binding inhibitors and fluorogenic substrates. Ang I was effectively cleaved by NEP to Ang (1-7) (kcat/K(m) of 6.2x10(5) M(-1) x s(-1)), but was a poor substrate for ACE2 (kcat/K(m) of 3.3x10(4) M(-1) x s(-1)). Ang (1-9) was a better substrate for NEP than ACE (kcat/K(m) of 3.7x10(5) M(-1) x s(-1) compared with kcat/K(m) of 6.8x10(4) M(-1) x s(-1)). Ang II was cleaved efficiently by ACE2 to Ang (1-7) (kcat/K(m) of 2.2x10(6) M(-1) x s(-1)) and was cleaved by NEP (kcat/K(m) of 2.2x10(5) M(-1) x s(-1)) to several degradation products. In contrast with a previous report, Ang (1-7), like Ang I and Ang (1-9), was cleaved with a similar efficiency by both the N- and C-domains of ACE (kcat/K(m) of 3.6x10(5) M(-1) x s(-1) compared with kcat/K(m) of 3.3x10(5) M(-1) x s(-1)). The two active sites of ACE exhibited negative co-operativity when either Ang I or Ang (1-7) was the substrate. In addition, a range of ACE inhibitors failed to inhibit ACE2. These kinetic data highlight that the flux of peptides through the RAS is complex, with the levels of ACE, ACE2 and NEP dictating whether vasoconstriction or vasodilation will predominate. PMID- 15283676 TI - Ubiquitination of the peroxisomal import receptor Pex5p. AB - Proteins harbouring a peroxisomal targeting signal of type 1 (PTS1) are recognized by the import receptor Pex5p in the cytosol which directs them to a docking and translocation complex at the peroxisomal membrane. We demonstrate the ubiquitination of Pex5p in cells lacking components of the peroxisomal AAA (ATPases associated with various cellular activities) or Pex4p-Pex22p complexes of the peroxisomal protein import machinery and in cells affected in proteasomal degradation. In cells lacking components of the Pex4p-Pex22p complex, mono ubiquitinated Pex5p represents the major modification, while in cells lacking components of the AAA complex polyubiquitinated forms are most prominent. Ubiquitination of Pex5p is shown to take place exclusively at the peroxisomal membrane after the docking step, and requires the presence of the RING-finger peroxin Pex10p. Mono- and poly-ubiquitination are demonstrated to depend on the ubiquitin-conjugating enzyme Ubc4p, suggesting that the ubiquitinated forms of Pex5p are targeted for proteasomal degradation. Accumulation of ubiquitinated Pex5p in proteasomal mutants demonstrates that the ubiquitination of Pex5p also takes place in strains which are not affected in peroxisomal biogenesis, indicating that the ubiquitination of Pex5p represents a genuine stage in the Pex5p receptor cycle. PMID- 15283677 TI - Staphylococcus aureus DNA ligase: characterization of its kinetics of catalysis and development of a high-throughput screening compatible chemiluminescent hybridization protection assay. AB - DNA ligases are key enzymes involved in the repair and replication of DNA. Prokaryotic DNA ligases uniquely use NAD+ as the adenylate donor during catalysis, whereas eukaryotic enzymes use ATP. This difference in substrate specificity makes the bacterial enzymes potential targets for therapeutic intervention. We have developed a homogeneous chemiluminescence-based hybridization protection assay for Staphylococcus aureus DNA ligase that uses novel acridinium ester technology and demonstrate that it is an alternative to the commonly used radiometric assays for ligases. The assay has been used to determine a number of kinetic constants for S. aureus DNA ligase catalysis. These included the K(m) values for NAD+ (2.75+/-0.1 microM) and the acridinium-ester labelled DNA substrate (2.5+/-0.2 nM). A study of the pH-dependencies of kcat, K(m) and kcat/K(m) has revealed values of kinetically influential ionizations within the enzyme-substrate complexes (kcat) and free enzyme (kcat/K(m)). In each case, the curves were shown to be composed of one kinetically influential ionization, for k(cat), pK(a)=6.6+/-0.1 and kcat/K(m), pK(a)=7.1+/-0.1. Inhibition characteristics of the enzyme against two Escherichia coli DNA ligase inhibitors have also been determined with IC50 values for these being 3.30+/-0.86 microM for doxorubicin and 1.40+/-0.07 microM for chloroquine diphosphate. The assay has also been successfully miniaturized to a sufficiently low volume to allow it to be utilized in a high-throughput screen (384-well format; 20 microl reaction volume), enabling the assay to be used in screening campaigns against libraries of compounds to discover leads for further drug development. PMID- 15283678 TI - Batch cooling crystallization and pressure filtration of sulphathiazole: the influence of solvent composition. AB - Currently there is a great interest in new process analytical approaches to increase the process understanding of pharmaceutical unit operations. In the present study, the influence of the solvent composition on the material properties and, further, on the filtration characteristics, of different crystal suspensions obtained through an unseeded batch-cooling-crystallization process was studied. Sulphathiazole, which is an antibiotic agent with multiple polymorphic forms, was produced by performing laboratory-scale cooling crystallization experiments from five different mixtures of water and propan-1-ol (n-propanol). The size, shape and polymorphic composition of the crystals produced were characterized with a scanning electron microscope, with a novel automated image analyser and with an X-ray powder diffractometer. All of the monitored crystal properties were found to clearly differ between the samples obtained from different solvents. The crystals produced in the batch-cooling crystallization experiments were separated from the crystallizing solvents using a batch-type pressure Nutsche filter, and the filtration characteristics of the suspensions were evaluated on the basis of average filter-cake porosities and average specific cake resistances, which were determined from the experimentally obtained filtration data. Comparison between the calculated filtration characteristics revealed that considerable differences existed between the different suspensions, and it could therefore be concluded that the pressure filtration process was influenced by the composition of the crystallizing solvent. The filterability of all the studied sulphathiazole suspensions was considered to be rather good on the basis of the relatively low cake porosities (0.51-0.63), which were accompanied with low average specific cake resistances [(8.7 x 10(7))-(1.2 x 10(9)) m/kg]. PMID- 15283679 TI - Is estrogen receptor alpha key to controlling bones' resistance to fracture? AB - The ability of bones to withstand functional loading without damage depends upon their cell populations establishing and subsequently maintaining a mass and architecture that are appropriately robust for the purpose. In women, the rapid loss of bone associated with the menopause represents a steplike decline in the effectiveness of this process with consequent increase in bone fragility. In men, loss of bone tissue and reduction in bone strength are more gradual and the increased incidence of fragility fractures occurs later. In both sexes, bone mass is associated with levels of bioavailable estrogen. This poses the major question as to how the presence or concentration of the reproductive hormone estrogen influences the relationship between bone mass and bone loading. In this paper, we briefly review evidence of the mechanism(s) by which the mechanical strains engendered by loading influence bone cells to establish and maintain structurally competent bone architecture. We highlight the finding that at least one strain related cascade responsible for adaptive control of bone architecture is mediated through estrogen receptor (ER) alpha, the number and activity of which are regulated by estrogen. We hypothesize that a major contributor to the rapid loss of bone mass that occurs in females, and the slower age-related fall in males and females, is reduced effectiveness of ER-mediated processing of strain-related information by resident bone cells. PMID- 15283680 TI - The adaptive response of bone to mechanical loading in female transgenic mice is deficient in the absence of oestrogen receptor-alpha and -beta. AB - Postmenopausal osteoporosis represents a failure of the response by which bone cells adapt bone mass and architecture to be sufficiently strong to withstand loading without fracture. To address why this failure should be associated with oestrogen withdrawal, we investigated the ulna's adaptive response to mechanical loading in adult female mice lacking oestrogen receptor-alpha (ERalpha(-/-)), those lacking oestrogen receptor-beta (ERbeta(-/-)) and their wild-type littermates. In wild-type mice, short periods of physiologic cyclic compressive loading of the ulna in vivo over a 2-week period stimulates new bone formation. In ERalpha(-/-) and ERbeta(-/-) mice this osteogenic response was respectively threefold and twofold less (P<0.05). In vitro, primary cultures of osteoblast like cells derived from these mice were subjected to a single short period of mechanical strain. Twenty-four hours after strain the number of wild-type cells was 61+/-25% higher than in unstrained controls (P<0.05), whereas in ERalpha(-/-) cells there was no strain-related increase in cell number. However, the strain related response of ERalpha(-/-) cells could be partially rescued by transfection with functional human ERalpha (P<0.05). ERbeta(-/-) cells showed a 125+/-40% increase in cell number following strain. This was significantly greater than in wild types (P<0.05).These data support previous findings that functional ERalpha is required for the full osteogenic response to mechanical loading and particularly the stage of this response, which involves an increase in osteoblast number. ERbeta appears to depress the ERalpha-mediated strain-related increase in osteoblast number in vitro, but in female transgenic mice in vivo the constitutive absence of either ERalpha or ERbeta appears to diminish the osteogenic response to loading. PMID- 15283681 TI - Analysis of spatial and temporal expression patterns of bone morphogenetic protein family members in the rat uterus over the estrous cycle. AB - Recent studies have demonstrated that bone morphogenetic proteins (BMPs) play fundamental roles in female fertility. This is particularly evident in terms of the ovary. One major question that is just beginning to be addressed is the role of BMPs in the non-pregnant uterus. To help fill this gap, we used in situ hybridization to investigate the expression of BMP family members in the rat uterus over the estrous cycle. We found that the endometrial/uterine cycle is accompanied by the expression of several components of the BMP pathway - including ligands, receptors and antagonists. The mRNAs encoding BMP receptors are expressed in the epithelial (BMP-RIA, -RIB and -RII), periluminal stroma (BMP RIA and -RII) and smooth muscle cells (BMP-RIA and -RII). The expression of all three receptors showed clear cyclic variations. The mRNAs encoding BMP ligands were highly expressed in the periluminal stroma (BMP-2 and -7) and glandular epithelium (BMP-7). The expression of BMP-2, but not BMP-7, was cyclical. Notably, the periluminal stroma expressed noggin mRNA. In the blood vascular system, BMP-4, -6 and -RII mRNAs were expressed in myometrial endothelial cells. Interestingly, follistatin, noggin, and BMP-4, -6 and -7 mRNAs were expressed in eosinophilic leukocytes, suggesting unexpected roles for eosinophil-derived BMPs in uterine function. We conclude that BMP ligands, receptors and antagonists are expressed in spatially and temporally restricted patterns that are consistent with a physiological role for these regulatory molecules in promoting uterine cellular processes including cell proliferation, differentiation and apoptosis during the cycle. PMID- 15283682 TI - Epithelial c-jun and c-fos are temporally and spatially regulated by estradiol during neonatal rat oviduct differentiation. AB - Expression of transcription factors binding to the activating protein-1 (AP-1) site is induced by estrogens in association with epithelial proliferation in the uterus, but, in the oviduct, the relationship between cell proliferation and differentiation and AP-1 transcription factors is not well understood. In the developing rat oviduct, we found that proliferation and differentiation of epithelial cells were region-dependently regulated by 17beta-estradiol (E2). To determine the role of AP-1 transcription factors in the development of rat oviduct, we performed immunohistochemistry for epithelial c-jun and c-fos proteins in E2-untreated and -treated newborn rats. E2 increased the expression of c-jun and c-fos during proliferation of undifferentiated epithelial cells, but diminished both proteins during accelerated differentiation of ciliated epithelial cells. A pure estrogen receptor (ER) antagonist, ICI 182,780, inhibited changes in their expression during both cell proliferation and differentiation. Importantly, no reduction of c-jun was noted in the epithelial cells of the foxj1-deficient oviduct, which lacks cilia development. This study shows that c-jun and c-fos are regulated during epithelial cell proliferation and differentiation in a region-specific manner. This provides critical information for understanding the molecular and cellular mechanisms of the development of the neonatal oviduct. PMID- 15283683 TI - Effects of peroxisome proliferator activated receptors-alpha and -gamma agonists on estradiol-induced proliferation and hyperplasia formation in the mouse uterus. AB - It is suggested that the action of peroxisome proliferator-activated receptors (PPARs) cross-talks with estrogen signaling in the uterus. However, it is not known how PPAR agonists affect estrogen-dependent processes in the uterus, especially proliferation and morphogenetic changes. The effects of agonists of PPAR-alpha and -gamma on proliferative and morphogenetic reactions in the uterus under short- and long-term estrogen treatments were therefore examined. Ovariectomized mice were treated with estradiol dipropionate (4 micro g/100 g, s.c., once a week) or vehicle and rosiglitazone (PPAR-gamma agonist) or fenofibrate (PPAR-alpha agonist) or with no additional treatment for 2 days or for 30 days. Treatment with estradiol and PPAR agonists for 2 days did not affect uterine mass. In mice treated with estradiol and rosiglitazone for 2 days, proliferation was enhanced and levels of estrogen receptors-alpha and beta catenin were decreased in all uterine tissues. Treatment with estradiol and fenofibrate for 2 days had the opposite effects on the parameters tested. In animals treated with estradiol and rosiglitazone for 30 days, uterine mass was increased, abnormal uterine glands and atypical endometrial hyperplasia were found more often and levels of estrogen receptors-alpha and beta-catenin were decreased. In animals treated with estradiol and fenofibrate for 30 days, uterine mass was decreased, most of the uterine glands had a normal structure, no cases of atypical hyperplasia were diagnosed, proliferative activity was declined and the levels of estrogen receptors-alpha and beta-catenin were markedly higher. Treatment with rosiglitazone or fenofibrate did not affect the serum estradiol level in the mice which received estradiol together with PPAR agonists for 30 days. Thus, rosiglitazone exerted the proliferative and morphogenetic effects of estradiol, but fenofibrate had the opposite effect. The actions of rosiglitazone and fenofibrate are associated with changes in the expression of estrogen receptors-alpha and beta-catenin in the uterus. PMID- 15283684 TI - Characterization of the concentration gradient of prostaglandin H synthase 2 mRNA throughout the pregnant baboon uterus. AB - The present study was designed to determine the effect of the spatial gradient from the cervix to the uterine fundus on the control of local prostaglandin H synthase (PGHS) 2 mRNA expression. We performed total cesarean hysterectomies during the last trimester in 12 pregnant baboons, 7 not in labor and 5 in labor, and examined PGHS2 mRNA expression throughout the uterus. PGHS2 mRNA abundance was quantified by in situ hybridization and northern blot analysis in the uterine fundus, lower uterine segment and the different segments of the cervix. Quantitative northern blot and in situ analysis demonstrated a gradient of PGHS2 mRNA expression, with the highest levels at the level of the lower portion of the cervix and decreased expression through the mid- and upper portion of the cervix and lower uterine segment; the lowest levels of expression were seen in the uterine fundus. Moreover, cellular localization of PGHS2 mRNA and protein demonstrated high levels of expression in the cervical glandular epithelial cells with only occasional staining of smooth muscle cells in pregnant baboons. Decreased PGHS2 mRNA concentration gradient from the cervical external os to the fundus suggests that prostaglandin (PG) production in the uterus and cervix strongly depends on anatomical relations. This increased local PG production activity may be critical to pregnancy-associated lower uterine segment elongation, cervical softening and effacement in primate labor. These data provide a compelling biological basis for the use of PGHS2 inhibitors in the prophylaxis of preterm birth and cervical incompetence. PMID- 15283685 TI - Alteration of prostaglandin production and agonist responsiveness by n-6 polyunsaturated fatty acids in endometrial cells from late-gestation ewes. AB - We investigated the effect of n-6 polyunsaturated fatty acids (PUFAs) on prostaglandin (PG) production by the uterus. A mixed population of endometrial cells (epthelium and stroma) from late-gestation ewes were cultured in defined medium containing linoleic acid (LA, 18:2, n-6), gamma-linolenic acid (GLA, 18:3, n-6) or arachidonic acid (AA, 20:4, n-6) in concentrations of 0 (control), 20 or 100 microM. After 45 h in test medium with or without added PUFAs, cells were challenged with control medium (CM), oxytocin (OT, 250 nM), lipopolysaccharide (LPS, 0.1 micro g/ml) or dexamethasone (DEX, 5 microM) for 22 h in the continued presence of the same concentration of PUFA and the medium was collected for measurement of PGF(2alpha) and PGE(2). Supplementation with LA inhibited the production of PGF(2alpha) but did not alter PGE(2), whereas GLA and AA increased production of both PGs. All PUFA supplements thus increased the ratio of PGE(2) to PGF(2alpha) (E:F ratio) two- to threefold. In control cells, OT and LPS challenges stimulated the production of PGF(2alpha) and PGE(2). In all challenge groups, the concentrations of PGF(2alpha) in response to PUFAs followed the same pattern - LA C, intron 4b --> a and Glu298 --> Asp) in patients with angiographic CAD (coronary artery disease), and/or prior MI (myocardial infarction) and a group of healthy population-based controls. However, the findings of this study appear to contradict previous studies published on the eNOS polymorphisms, and this commentary will attempt to resolve the inconsistency in such genetic association studies. PMID- 15283698 TI - Inflammatory and redox responses to ischaemia/reperfusion in human skeletal muscle. AB - The objective of this study was to identify cellular and plasma marker(s) of post I/R (ischaemia/reperfusion) in patients undergoing elective knee surgery where a tourniquet was used to facilitate a bloodless surgical field. We evaluated the inflammatory and redox response by measuring the mRNA levels of ICAM-1 (intercellular cell-adhesion molecule-1), MnSOD (manganese superoxide dismutase), GST-mu (glutathione transferase-mu) and Cu/ZnSOD (copper/zinc superoxide dismutase) in the operated muscle and blood cells pre-operatively (pre tourniquet) and at various times after reperfusion (tourniquet release). We also measured plasma concentrations of IL (interleukin)-6, IL-8, sICAM-1 (soluble ICAM 1), IL-1beta and TNF-alpha (tumour necrosis factor-alpha) using ELISA. Our results show a strong induction of MnSOD and GST-mu in granulocytes (but not in mononuclear cells or muscle) after reperfusion (2 and 4 h). There was no change in the mRNA level of Cu/ZnSOD after reperfusion. An up-regulation of membrane ICAM-1 in muscle and a decrease in sICAM-1 in plasma were detected after reperfusion. Plasma IL-6 and IL-8 levels (but not TNF-alpha or IL-1beta) increased significantly over baseline at 2 and 4 h after reperfusion. Elevated expression of ICAM-1 in muscle, MnSOD and GST-mu in granulocytes and increased levels of plasma IL-6 and IL-8 may be considered as phase- and cell-specific markers of post-I/R of skeletal muscle in humans. PMID- 15283699 TI - Spermidine/spermine-N1-acetyltransferase-2 (SSAT2) acetylates thialysine and is not involved in polyamine metabolism. AB - Spermidine/spermine-N1-acetyltransferase (SSAT1) is a short-lived polyamine catabolic enzyme inducible by polyamines and polyamine analogues. Induction of SSAT1 plays an important role in polyamine homoeostasis, since the N1-acetylated polyamines can be excreted or oxidized by acetylpolyamine oxidase. We have purified a recombinant human acetyltransferase (SSAT2) that shares 45% identity and 61% homology with human SSAT1, but is only distally related to other known members of the GNAT (GCN5-related N-acetyltransferase) family. Like SSAT1, SSAT2 is widely expressed, but did not turn over rapidly, and levels were unaffected by treatments with polyamine analogues. Despite similarity in sequence to SSAT1, polyamines were found to be poor substrates of purified SSAT2, having K(m) values in the low millimolar range and kcat values of <0.01 s(-1). The kcat/K(m) values for spermine and spermidine for SSAT2 were <0.0003% those of SSAT1. Expression of SSAT2 in NIH-3T3 cells was not detrimental to growth, and did not reduce polyamine content or increase acetylpolyamines. These results indicate that SSAT2 is not a polyamine catabolic enzyme, and that polyamines are unlikely to be its natural intracellular substrates. A promising candidate for the physiological substrate of SSAT2 is thialysine [S-(2-aminoethyl)-L-cysteine], which is acetylated predominantly at the epsilon-amino group with K(m) and kcat values of 290 muM and 5.2 s(-1). Thialysine is a naturally occurring modified amino acid that can undergo metabolism to form cyclic ketimine derivatives found in the brain and as urinary metabolites, which can undergo further reaction to form antioxidants. SSAT2 should be renamed 'thialysine N(epsilon)-acetyltransferase', and may regulate this pathway. PMID- 15283700 TI - A novel member of the GCN5-related N-acetyltransferase superfamily from Caenorhabditis elegans preferentially catalyses the N-acetylation of thialysine [S-(2-aminoethyl)-L-cysteine]. AB - The putative diamine N-acetyltransferase D2023.4 has been cloned from the model nematode Caenorhabditis elegans. The 483 bp open reading frame of the cDNA encodes a deduced polypeptide of 18.6 kDa. Accordingly, the recombinantly expressed His6-tagged protein forms an enzymically active homodimer with a molecular mass of approx. 44000 Da. The protein belongs to the GNAT (GCN5-related N-acetyltransferase) superfamily, and its amino acid sequence exhibits considerable similarity to mammalian spermidine/spermine-N1-acetyltransferases. However, neither the polyamines spermidine and spermine nor the diamines putrescine and cadaverine were efficiently acetylated by the protein. The smaller diamines diaminopropane and ethylenediamine, as well as L-lysine, represent better substrates, but, surprisingly, the enzyme most efficiently catalyses the N acetylation of amino acids analogous with L-lysine. As determined by the k(cat)/K(m) values, the C. elegans N-acetyltransferase prefers thialysine [S-(2 aminoethyl)-L-cysteine], followed by O-(2-aminoethyl)-L-serine and S-(2 aminoethyl)-D,L-homocysteine. Reversed-phase HPLC and mass spectrometric analyses revealed that N-acetylation of L-lysine and L-thialysine occurs exclusively at the amino moiety of the side chain. Remarkably, heterologous expression of C. elegans N-acetyltransferase D2023.4 in Escherichia coli, which does not possess a homologous gene, results in a pronounced resistance against the anti-metabolite thialysine. Furthermore, C. elegans N-acetyltransferase D2023.4 exhibits the highest homology with a number of GNATs found in numerous genomes from bacteria to mammals that have not been biochemically characterized so far, suggesting a novel group of GNAT enzymes closely related to spermidine/spermine-N1 acetyltransferase, but with a distinct substrate specificity. Taken together, we propose to name the enzyme 'thialysine N(epsilon)-acetyltransferase'. PMID- 15283701 TI - Synergistic role of specificity proteins and upstream stimulatory factor 1 in transactivation of the mouse carboxylesterase 2/microsomal acylcarnitine hydrolase gene promoter. AB - Mouse carboxylesterase 2 (mCES2), a microsomal acylcarnitine hydrolase, is thought to play some important roles in fatty acid (ester) metabolism, and it is therefore thought that the level of transcription of the mCES2 gene is under tight control. Examination of the tissue expression profiles revealed that mCES2 is expressed in the liver, kidney, small intestine, brain, thymus, lung, adipose tissue and testis. When the mCES2 promoter was cloned and characterized, it was revealed that Sp1 (specificity protein 1) and Sp3 could bind to a GC box, that USF (upstream stimulatory factor) 1 could bind to an E (enhancer) box, and that Sp1 could bind to an NFkappaB (nuclear factor kappaB) element in the mCES2 promoter. Co-transfection assays showed that all of these transcription factors contributed synergistically to transactivation of the mCES2 promoter. Taken together, our results indicate that Sp1, Sp3 and USF1 are indispensable factors for transactivation of the mCES2 gene promoter. To our knowledge, this is the first study in which transcription factors that interact with a CES2 family gene have been identified. The results of the present study have provided some clues for understanding the molecular mechanisms regulating mCES2 gene expression, and should be useful for studies aimed at elucidation of physiological functions of mCES2. PMID- 15283704 TI - Diagnosis of infantile spasms, Lennox-Gastaut syndrome, and progressive myoclonic epilepsy. AB - The epilepsies of childhood are distinguished by an interesting dichotomy between the benign and catastrophic disorders. Approximately 50% of children outgrow childhood epilepsy as they mature; although the disorder is disruptive for children and families alike, it is not considered a medical disaster. The catastrophic epilepsies of childhood, in contrast, are associated with significant morbidity and mortality. Infantile spasms, Lennox-Gastaut syndrome, and the progressive myoclonic epilepsies are correlated with significant disability and a multiplicity of underlying etiologies. Accurate diagnosis of both the syndrome and the etiology is very important for treatment purposes, as well as for family education, since many of the disorders have a significant genetic component. PMID- 15283702 TI - Glycoprotein VI/Fc receptor gamma chain-independent tyrosine phosphorylation and activation of murine platelets by collagen. AB - We have investigated the ability of collagen to induce signalling and functional responses in suspensions of murine platelets deficient in the FcRgamma (Fc receptor gamma) chain, which lack the collagen receptor GPVI (glycoprotein VI). In the absence of the FcRgamma chain, collagen induced a unique pattern of tyrosine phosphorylation which was potentiated by the thromboxane analogue U46619. Immunoprecipitation studies indicated that neither collagen alone nor the combination of collagen plus U46619 induced phosphorylation of the GPVI-regulated proteins Syk and SLP-76 (Src homology 2-containing leucocyte protein of 76 kDa). A low level of tyrosine phosphorylation of phospholipase Cgamma2 was observed, which was increased in the presence of U46619, although the degree of phosphorylation remained well below that observed in wild-type platelets (approximately 10%). By contrast, collagen-induced phosphorylation of the adapter ADAP (adhesion- and degranulation-promoting adapter protein) was substantially potentiated by U46619 to levels equivalent to those observed in wild-type platelets. Collagen plus U46619 also induced significant phosphorylation of FAK (focal adhesion kinase). The functional significance of collagen-induced non-GPVI signals was highlighted by the ability of U46619 and collagen to induce the secretion of ATP in FcRgamma chain-deficient platelets, even though neither agonist was effective alone. Protein tyrosine phosphorylation and the release of ATP were abolished by the anti-(alpha2 integrin) antibodies Ha1/29 and HMalpha2, but not by blockade of alphaIIbbeta3. These results illustrate a novel mechanism of platelet activation by collagen which is independent of the GPVI-FcRgamma chain complex, and is facilitated by binding of collagen to integrin alpha2beta1. PMID- 15283705 TI - Basic science behind the catastrophic epilepsies. AB - The major catastrophic epileptic syndromes of childhood include infantile spasms, Lennox-Gastaut syndrome, and the progressive myoclonus epilepsies (PMEs). Although each of these syndromes manifests in an age-specific manner and is defined by distinct electroclinical features, they are all refractory to medical therapy and are invariably associated with psychomotor deficits, and in the most severe cases, either epileptic encephalopathy or progressive neurodegeneration. While much has been written about the clinical features and natural history of the catastrophic epilepsies, very little is known about the underlying pathophysiology. Progress in our understanding and treatment of these conditions has been hampered by the lack of suitable animal models in which putative mechanisms and novel targets for intervention could be rigorously studied. Nevertheless, recent clinical and basic investigations have identified certain mechanisms that may be relevant to their pathogenesis. In this review, three major hypotheses regarding the pathophysiology of infantile spasms are highlighted: the corticotropin-releasing hormone (CRH) hypothesis, the N-methyl-D aspartate (NMDA) hypothesis, and the serotonin-kynurenine hypothesis. One or more of these mechanisms may be relevant in part to later-onset catastrophic epilepsies since infantile spasms can persist into later childhood and, like Lennox-Gastaut syndrome, well into adulthood. There is a profound need to develop more relevant animal models of the developmental encephalopathic epilepsies to truly develop better therapeutic strategies for these catastrophic disorders. PMID- 15283706 TI - Pharmacologic treatment of the catastrophic epilepsies. AB - Treatment of the catastrophic epilepsies [infantile spasms (IS), Lennox-Gastaut syndrome (LGS), and progressive myoclonic epilepsy (PME)] remains a challenge to clinicians. For IS, adrenocorticotropic hormone has traditionally been the drug of choice in the United States but may be associated with serious side effects in some patients. Vigabatrin has shown promise in treating IS patients, particularly those with tuberous sclerosis. However, the drug is associated with visual field loss and is not commercially available in the United States. Newer antiepilepsy drugs (AEDs), such as zonisamide, topiramate (TPM), and lamotrigine (LTG), may be useful in patients with IS. Although LTG, TPM, and felbamate are approved in the United States for the treatment of LGS, the overall effectiveness of therapy in patients with LGS is poor. For PME, valproate is a first-line treatment. Zonisamide and levetiracetam also show promise. Supplementation with certain cofactors to correct deficiencies and increase mitochondrial function may be useful in some patients with PME, but response to such therapy is not well documented. Advances in our understanding of the etiologies, mechanisms, and genetics underlying the catastrophic epilepsies may facilitate more effective pharmacologic interventions. PMID- 15283707 TI - Nonpharmacologic treatment of the catastrophic epilepsies of childhood. AB - The catastrophic epilepsy syndromes of childhood are initially treated with a pharmacologic intervention in most cases. However, due to the poor response patients often have to pharmacologic interventions, nonpharmacologic treatment options are an important part of a comprehensive treatment plan for this group of children. Additionally, nonpharmacologic therapy may offer a method to minimize associated morbidity and mortality. This article discusses the use of epilepsy surgery, the ketogenic diet, and vagus nerve stimulation in the treatment of patients with infantile spasms, Lennox-Gastaut syndrome, and progressive myoclonic epilepsy. Efficacy of the nonpharmacologic treatment options, as measured by reduction in seizure frequency, as well as by developmental progress or behavioral improvement, varies according to the specific catastrophic epilepsy disorder and the treatment option. PMID- 15283708 TI - Following catastrophic epilepsy patients from childhood to adulthood. AB - As patients with catastrophic epilepsies move from childhood to adulthood, evolving and innovative therapeutic regimens are often required. However, the goal of providing the best quality of life while minimizing both seizures and side effects remains the same. Clinicians can develop appropriate care plans by being aware of patients' changing needs. Clinical symptoms of the catastrophic epilepsies may change over time; by understanding the natural history of a patient's condition, clinicians can help ease the transition from childhood to adulthood. Additionally, as children with catastrophic epilepsies become adults, medical issues (e.g., medication side effects, tolerance, and dependence) and nonmedical issues (e.g., guardian/caretaker issue, group home applications, and respite care options) must be considered when developing strategies for patient care. Regular assessment of patients, the development of emergency plans, and maintenance of consistency in the delivery of care are also important issues to consider. Finally, a multidisciplinary care plan that incorporates resources from health-care practitioners, social service professionals, and community agencies can be valuable in optimizing treatment for patients with catastrophic epilepsies. PMID- 15283709 TI - Primary prevention is better than cure. PMID- 15283710 TI - Paternalism, pragmatism or self protection? PMID- 15283711 TI - Optimal staffing of helicopter emergency medical services is controversial. PMID- 15283712 TI - Is the ACEM training programme adequate? PMID- 15283713 TI - Injury patterns and preventability in prehospital motor vehicle crash fatalities in Victoria. AB - OBJECTIVE: To examine the pattern of anatomical injury in victims of motor vehicle crashes who die prior to reaching hospital. Cases were identified where death was an unexpected outcome. METHODS: A retrospective review of autopsy case records including police reports, of all persons who died in motor vehicle crashes between 1 January 1998 and 31 December 1999 and underwent full autopsy at the Victorian Institute of Forensic Medicine (VIFM). Those cases where the victim died in the prehospital phase were examined. Abbreviate Injury Scores and Injury Severity Scores were calculated in each case. Bull's probit analysis was used to identify unexpected deaths. RESULTS: There were 352 motor road crash fatalities identified that underwent autopsy at the VIFM in the study period. Two hundred and six of these were prehospital deaths involving motor vehicles, which satisfied specified criteria. 82% (95% CI: 77.7-86.3%) of cases had Abbreviated Injury Scores of 5 (critical) or 6 (incompatible with life). 80.1% (95% CI: 75.7 84.5%) had an Injury Severity Score greater than 40. 36.9% (95% CI: 34.5-39.3%) of cases had the maximum Injury Severity score of 75. 88.8% (95% CI: 85-92.7%) of cases sustained a head injury and 83.9% (95% CI: 79.8-88.2%) a chest injury. Possibly preventable fatality was identified in 30 (14.6% 95% CI: 13.9-15.3%) cases. CONCLUSION: In motor vehicle crash fatalities, most victims who die before reaching hospital do so because of major injury, with the head and chest the commonest regions involved. A large proportion of these injuries could be considered unsurvivable regardless of treatment. Earlier intervention or retrieval of such patients is unlikely to influence outcome in the majority of cases. PMID- 15283714 TI - Does potassium concentration measured on blood gas analysis agree with serum potassium in patients with diabetic ketoacidosis? AB - OBJECTIVE: The aims of this study were to define the maximum clinically acceptable difference between potassium concentrations on different samples and to determine the degree of agreement between potassium concentration measured on blood gas analysis and serum for patients with diabetic ketoacidosis (DKA). METHODS: This project comprised two sub studies. In the first, 15 emergency physicians, intensivists and endocrinologists were surveyed and asked to mark on a line with markings at 0.5 mmol/L intervals, the maximum clinically acceptable differences (both above and below the 'true' value) between potassium concentration measured on different samples. The maximum clinically acceptable difference was calculated as the median of responses. The second study was a retrospective agreement study. Patients with an ED diagnosis of DKA were identified from a computer database. They were eligible for inclusion if they had both blood gas analysis including potassium concentration and serum potassium concentration and pH was less than 7.3. Data collected included potassium concentration on serum and blood gas samples, pH, serum glucose concentration and time of sample collections. Data were analysed using bias plot and Spearman correlation analyses. RESULTS: The maximum clinically acceptable difference was defined as 0.5 mmol/L for both over and underestimation of potassium concentration. Fifty patients were studied with a median pH of 7.17 and median serum glucose of 29.5 mmol/L. Difference in potassium concentration between samples ranged from -0.9-2.9 mmol/L. 80% of sample pairs had a difference within the maximum clinically acceptable difference defined previously. The magnitude of difference between samples correlated with serum glucose (P = 0.0033, coefficient 0.41) but not with pH. CONCLUSION: This study suggests that potassium concentration derived from blood gas analysis may not be an acceptable substitute for serum potassium concentration in patients with diabetic ketoacidosis, particularly at higher serum glucose concentrations. PMID- 15283715 TI - Incorrect instruction in the use of the Valsalva manoeuvre for paroxysmal supra ventricular tachycardia is common. AB - OBJECTIVE: Success rates for the Valsalva manoeuvre (VM) in treatment of paroxysmal supraventricular tachycardia (SVT) vary with performance technique. This study aimed to assess whether ED doctors instruct their patients to perform the recommended VM technique (supine position for 15 s). METHODS: A multicentre, observational study of 35 ED registrars and 17 emergency physicians. Each doctor was asked to describe how he/she would instruct a patient in SVT to perform the VM. RESULTS: Only five (9.6%) doctors would position their patient correctly and 31 (59.6%) would incorrectly instruct their patient to assume a sitting or semirecumbent position. Only five (9.6%) doctors would give specific instructions to blow for at least 15 s and 34 (65.4%) would instruct their patient to blow 'as long as you can'. Only four (7.4%) doctors would use a sphygmomanometer to measure intrathoracic pressure during the VM. There were no significant differences (P > 0.05) between the registrar and physician group responses for any study endpoint. CONCLUSION: Few ED doctors correctly instruct their patients in the VM technique recommended for management of SVT. Hence, maximal vagal tone and SVT conversion rates may not be achieved in many cases. The use of the recommended VM technique is encouraged. PMID- 15283716 TI - Do radiographs requested from a paediatric emergency department in New Zealand need reporting? AB - OBJECTIVES: To ascertain whether plain radiographs interpreted by doctors working in a paediatric ED need reporting by radiologists. METHODS: Prospective observational study based in a paediatric teaching hospital. RESULTS: The overall disagreement rate was 26.1%. The clinically significant disagreement rate was 4.8%. There was no relationship between seniority or confidence of ED doctor, and likelihood of disagreement. Disagreement was more likely where radiographs were interpreted during normal working hours (P < 0.05). Chest films, then abdominal films, then skeletal films were more likely to result in disagreement (P < 0.0001). However, this relationship was reversed when considering rates of clinically significant disagreement (P < 0.05). The highest risk of disagreement was in chest films initially thought to be normal, and the lowest in skeletal films initially thought to be abnormal. CONCLUSION: All radiographs requested on patients discharged from the paediatric ED should be reported. Reporting need not be immediate. PMID- 15283717 TI - Family member presence during resuscitation in the emergency department: An Australian perspective. AB - OBJECTIVE: The practice of family member presence during resuscitation in the ED has attracted widespread attention over the last few decades. Despite the recommendations of international organizations, clinical staff remain reluctant to engage in this practice in many EDs. This paper separates the evidence from opinion to determine the current state of knowledge about this practice. METHODS: A search strategy was developed and used to locate research based publications, which were subsequently reviewed for the strength of evidence providing the basis for recommendations. RESULTS: The literature was examined to reveal what patients and their family members want; the outcomes of family presence during resuscitation for patients and their family members; staff views and practices regarding family presence during resuscitation. Findings suggest that providing the opportunity to be with their critically ill family member is both important to and beneficial for families, however, disparity in staff views has been identified as a major obstacle to family presence during resuscitation. Examination of published guidelines and staff practices described in the literature revealed consistent elements. CONCLUSION: Although critics point to the lack of rigour in this body of literature, the current state of knowledge suggests merit in pursuing future research to examine and measure effects of family member presence during resuscitation on patients, family members and healthcare providers. PMID- 15283718 TI - Emergency management of the morbidly obese. AB - OBJECTIVES: To identify the difficulties encountered with the emergency management of morbidly obese patients and formulate recommendations to streamline care. METHODS: An English language literature search was undertaken using Medline (1966-2003) with key words 'morbid obesity' 'anaesthesia' 'imaging' 'obesity' 'emergency' 'transportation' 'retrieval' 'critical illness' and 'monitoring'. Potential articles were selected for content applicable to emergency medicine based on title and abstract and reviewed in detail. Reference lists were manually searched for further relevant articles. In view of the very limited systematic study in this area, all information deemed by the authors' to be of assistance to the emergency physician was included regardless of evidence level. Additional information was sought from standard critical care textbooks and their bibliographies and through personal communication with local ambulance and retrieval services. The authors' unpublished personal experience in providing emergency care to the morbidly obese was included for aspects of management not documented in medical literature. RESULTS: Obesity levels and associated health problems are rapidly rising in Australia. Few studies were identified dealing with critical illness in the morbidly obese and none specifically addressing ED management. Problems identified included size related logistical issues, and limitations of physical assessment, monitoring and routine investigations. Invasive procedures, intubation and ventilation can be particularly problematic, and modified techniques may be required. Limited data indicates a poorer outcome from critical illness most marked in the case of blunt traumatic injury. CONCLUSION: Very obese patients present a variety of logistical and medical challenges for EDs. A series of recommendations are made based on available data. Further studies in this area would be desirable to more specifically address ED issues. PMID- 15283719 TI - The role of physician staffing of helicopter emergency medical services in prehospital trauma response. AB - The crewing of Helicopter Emergency Medical Service (HEMS) for scene response to trauma patients is generally considered to be controversial, particularly regarding the role of physicians. This is reflected in HEMS in Australia with some services utilizing physician crewing for all prehospital missions. Others however, use physicians for selected missions only whilst others do not use physicians at all. This review seeks to determine whether the literature supports using physicians in addition to paramedics in HEMS teams for prehospital trauma care. Studies were excluded if they compared physician teams with basic life support teams (BLS) teams rather than paramedics. Ambulance officers were considered to be paramedics where they were able to administer intravenous fluids and use a method of airway management beyond bag-valve-mask ventilation. Studies were excluded if the skill set of the ambulance team was not defined, the level of staffing of the helicopter service was not stated, team composition varied without reporting outcomes for each team type, patient outcome data were not reported, or the majority of the transports were interhospital rather than prehospital transports. PMID- 15283720 TI - Drotrecogin alfa: a role in emergency department treatment of severe sepsis? AB - Human protein C is a serine protease that circulates in the blood as an inactive zymogen. It is converted to its active form by interaction with thrombomodulin on the endothelial wall. Activated protein C has a significant role in maintaining haemostasis, and is a major mechanism of controlling microvascular thrombosis. Recent reports describe the use of drotrecogin alfa (recombinant activated protein C) in severe sepsis, a condition relevant to emergency medicine. This review describes the physiology of the protein C pathway and its importance in sepsis. It will also focus on the use of drotrecogin alfa in sepsis, and its use in the ED. PMID- 15283721 TI - Why don't trainees pass the emergency medicine fellowship examination? AB - The pass rate in the Australasian College for Emergency Medicine Fellowship exams between 1996 and 2003 inclusive averaged 61%, substantially lower than that of other specialties with comparable training structures. The explanation for this pass rate, which many would judge as unacceptably low, is likely to be multi factorial. Possible factors that should be considered include trainee selection, training programme structure, the impact of the Director of Emergency Medicine Training, examination preparation, examination validity and examination reliability. Each of these potential factors needs to be addressed in a systematic fashion in the context of inevitable and increasing internal and external scrutiny of the outcomes of our training programme. PMID- 15283722 TI - The first year of a formal emergency medicine training programme in Papua New Guinea. AB - OBJECTIVE: To describe a programme catalyzing the development of emergency medicine in Papua New Guinea (PNG). METHODS: Five emergency physicians rotated through a new position of Senior Lecturer in Emergency Medicine in the University of PNG during 2003. The position was established as a consequence of emergency physician input supported by AusAID in 2002. RESULTS: Fifth (final)-year medical students and medical officers in the Emergency Department at Port Moresby General Hospital undertook formal and bedside problem based learning. The first trainees for a Master of Medicine in Emergency Medicine programme were inducted and supported. Emergency department management was provided with specialist input. Research projects were initiated, dealing with snakebite, chloroquine toxicity and HIV/AIDS. The first year of an emergency nursing curriculum was supported. CONCLUSIONS: There is now considerable enthusiasm for the development of emergency medicine as the hospital generalists' specialty. Emergency nursing training has also made a start. Limitations on resources will require flexibility to sustain the project. Further support by emergency physicians will be needed. PMID- 15283723 TI - Audit as a learning tool in postgraduate emergency medicine training. AB - OBJECTIVE: Educational activities for emergency medicine trainees need to be clinically focused, relevant, and ideally have the capacity to change practice and patient outcomes. It is proposed that the use of audit methods in educational sessions may address these learning needs. The aims of this project were to involve emergency medicine trainees in undertaking audits of ED patient care, and to evaluate the use of this technique in fulfilling training needs. METHODS: Trainees were given clinical topics on which to develop a presentation at weekly education sessions within the ED. This presentation included a brief clinical audit concerning an aspect of the same topic. The audit question addressed one of the issues identified as standard of care in the trainee presentation and investigated our department's level of compliance with that standard. At the end of a 6-month period, a questionnaire was given to all trainees involved, either as presenters or attendees at an audit presentation. RESULTS: Trainees performing audits reported that this method was most useful for demonstrating the limitations of coding, giving a greater appreciation of poor documentation in medical records, and improving their presentation skills. Most trainees attending audit based presentations reported that this educational method gave them a greater appreciation of systems based practice, actually changed their clinical practice, and was more useful than traditional lectures. CONCLUSIONS: Audit should be a key component of emergency medicine education. Trainees perceive the technique as useful addressing a number of training needs in a clinical context. PMID- 15283724 TI - Responding to formal complaints about the emergency department: lessons from the service marketing literature. AB - The ability to respond to formal complaints is a necessary part of emergency medicine practice. In spite of the significance of formal complaints there is little guidance within the medical literature to understand why patients complain or how to provide satisfaction to individuals who complain. Practitioners are usually left to their own devices in the style and substance of complaint responses even when working within a defined complaint management system. This article draws on relatively abundant literature in the service marketing field to provide an understanding of dissatisfaction, complaining and complaint handling. Having developed an appropriate theoretical framework the article provides guidance for applying these concepts in dealing with formal complaints. PMID- 15283725 TI - Phenytoin overdose complicated by prolonged intoxication and residual neurological deficits. AB - This report describes a case of massive phenytoin deliberate self-poisoning, notable for delayed peak serum concentrations, multiple general complications, and permanent cerebellar injury. A 38-year-old 70 kg male patient presented to the ED after ingestion of at least 10 g of phenytoin 12-16 h earlier. Marked cerebellar dysfunction and persistent vomiting were observed, with an initial serum phenytoin concentration of 181 micromol/L. Initial conservative treatment (activated charcoal, whole bowel irrigation), and later attempts at charcoal haemoperfusion were unsuccessful. The serum phenytoin concentration peaked on day 15 (354 micromol/L). The patient developed seizures followed by a prolonged depression in conscious state requiring intubation. Multiple medical sequelae occurred and the patient was discharged to a rehabilitation facility 100 days after admission exhibiting signs consistent with permanent cerebellar dysfunction. PMID- 15283730 TI - Urgent removal of nasal foreign bodies. PMID- 15283732 TI - A nasal foreign body that progressed. PMID- 15283733 TI - What's the surprise? PMID- 15283735 TI - The impact of the SCL-90 on the validity of the DSM-IV neurotic or stress-related disorders. PMID- 15283736 TI - Platelet hyperactivity in clinical depression and the beneficial effect of antidepressant drug treatment: how strong is the evidence? AB - OBJECTIVE: Platelet hyperactivity is thought to contribute to the increased coronary artery disease (CAD) risk in depression. This study reviewed the evidence for hyperactive platelets and for effects of antidepressant drug treatment on platelet 'stickiness' in clinical depression. METHOD: By means of PubMed electronic library search, 34 studies in English were identified (1983 2003) and critically reviewed. RESULTS: In depression, flow cytometry studies allowing detection of subtle platelet activation states consistently found at least one platelet activation marker to be increased, while the bulk of platelet aggregation studies did not suggest increased platelet aggregability. Platelets seem to be more activated in depressed patients with CAD than in depressed individuals without CAD. The selective serotonin reuptake inhibitors normalized platelet hyperactivity in four studies. CONCLUSION: Data on platelet activity in depression are inconclusive. To resolve this issue and its clinical implications, studies in larger sample sizes controlling for confounders of platelet functioning and prospectively designed are needed. PMID- 15283737 TI - Is depression related to subsequent diabetes mellitus? AB - OBJECTIVE: To compare the risk of developing diabetes mellitus (DM) in the general population between subjects who had a depression and subjects who never had a depression. METHOD: Retrospective cohort design. People with depression were diagnosed with a depression between 1975 and 1990; controls never had a depression. Both groups were followed for a diagnosis of type II diabetes until 2000. Data on 1334 depressed and 66 670 non-depressed subjects were available from a large general practice-based database. RESULTS: No overall relation was found, but among males below age 50 there was a 78% increase in the rate of development of DM compared with non-depressed patients (hazard ratio 1.78, 95% CI: 1.21-2.62). CONCLUSION: Depression in males between the age of 20 and 50 years is related to an increased risk of developing DM. PMID- 15283738 TI - Changes in brain metabolism associated with remission in unipolar major depression. AB - OBJECTIVE: Functional brain correlates of remission in patients with major depressive disorder (MDD) are measured with positron emission tomography (PET) and 18F-fluorodeoxyglucose. METHOD: Glucose metabolism was measured in patients (n = 41) with moderate to severe MDD during acute depression and in the remitted state defined as a period of asymptomatic condition over 12 weeks. Data analyses used a region-of-interest (ROI) approach and statistical parametric mapping (SPM). RESULTS: There were significant decreases in metabolism upon remission with respect to the baseline scan in left prefrontal, anterior temporal and anterior cingulate cortex and bilateral thalamus (SPM analysis) and bilateral putamen and cerebellum (SPM and ROI analyses). There was a significant asymmetry in prefrontal and anterior cingulate cortex metabolism with lower metabolism in the left hemisphere that persisted despite clinical remission. CONCLUSION: These findings support the hypothesis that selective monoamine reuptake inhibition leads to an attenuation of a brain circuit that mediates depressive symptomatology. PMID- 15283739 TI - Smoking and depressive symptoms in Chinese elderly in Hong Kong. AB - OBJECTIVE: To examine the association between smoking and depressive symptoms among Chinese elderly in Hong Kong. METHOD: Cross-sectional data on smoking and depressive symptoms from 56,167 Chinese elderly aged 65 or over in Hong Kong were analysed using logistic regression. RESULTS: Current smokers and former smokers were more likely to have depressive symptoms than never smokers. The adjusted odds ratios (OR) and 95% confidence interval (CI) in males and females respectively were 1.62 (1.34-1.96) and 1.43 (1.20-1.70) for current smokers, and were 1.18 (0.99-1.40) and 1.29 (1.12-1.47) for former smokers. Former smokers were less likely to have depressive symptoms than current smokers (OR = 0.80, 95% CI: 0.70-0.92). CONCLUSION: Smoking is positively associated with depressive symptoms in Chinese elderly. Health care workers should be vigilant about the detection of depressive symptoms in elderly smokers. PMID- 15283740 TI - Development of an antidepressant compliance questionnaire. AB - OBJECTIVE: The development of the Antidepressant Compliance Questionnaire (ADCQ), assessing patients' attitudes and beliefs on depression and antidepressants. METHOD: A 51-item questionnaire was applied to 85 psychiatric out-patients with a DSM-IV diagnosis of major depressive disorder (MDD). This data set was used to assess psychometric properties of the ADCQ. The questionnaire was also applied to 272 primary care out-patients with MDD. RESULTS: A principal component analysis revealed four dimensions with good internal consistency and acceptable test retest reliability: 'perceived doctor-patient relationship', 'preserved autonomy', 'positive beliefs on antidepressants' and 'partner agreement', resulting in a final questionnaire comprising 33-items. Responses were independent from depression severity and patient age. The response patterns of both psychiatric and primary care patients are provided and illustrate the many erroneous beliefs on antidepressants. CONCLUSION: The ADCQ has good psychometric properties; further investigation should investigate whether this questionnaire is predictive of patient compliance. PMID- 15283741 TI - Functional disability and depression in the general population. Results from the Netherlands Mental Health Survey and Incidence Study (NEMESIS). AB - OBJECTIVE: Data on the temporal relationships between duration of depression and recovery and functional disability are sparse. These relationships were examined in subjects from the general population (n = 250) with newly originated episodes of DSM-III-R major depression. METHOD: The Netherlands Mental Health Survey and Incidence Study is a prospective epidemiological survey in the adult population (n = 7076), using the Composite International Diagnostic Interview (CIDI). Duration of depression and duration of recovery over 2 years were assessed with a life chart interview. Functional disabilities were assessed with the MOS-SF-36 and with absence days from work. RESULTS: Functional disabilities and absence days in depressed individuals were not found to be associated with duration of depression. Functioning in daily activities improved with longer duration of recovery but social functioning not. CONCLUSION: Functioning deteriorates by actual depressive symptomatology and comorbid anxiety but not by longer duration of depression. After symptomatic recovery, functioning improves to premorbid level, irrespective of the length of the depression. Improvements in daily activities and work can be expected with longer duration of recovery. PMID- 15283742 TI - Is SCL-90R helpful for the clinician in assessing DSM-IV symptom disorders? AB - OBJECTIVE: To investigate associations between six of the Symptom Check-List 90R (SCL-90R) subscales and specific DSM-IV symptom disorders in a sample of patients with high comorbidity of axis I and axis II disorders. METHOD: SCL-90R questionnaires from 1202 patients admitted for treatment in the Norwegian Network of Psychotherapeutic Day Hospitals. Mean score differences on subscales among diagnostic categories were investigated. With diagnoses as external criteria, cut off scores for different diagnostic groups were used to calculate diagnostic efficacy. Multiple regressions were conducted in order to disentangle variance of the subscales accounted for by symptom disorders. Frequency distributions of subscales for patients with and without symptom disorders were compared. RESULTS: Poor diagnostic efficacy was found for most of the subscales, except for the phobic anxiety subscale. The strongest association with each subscale was with its associated symptom disorder, except for dysthymia. CONCLUSION: Diagnostic inferences about DSM-IV symptom disorders based on SCL-90R should be conducted with care. PMID- 15283743 TI - The SCL-90 and SCL-90R versions validated by item response models in a Danish community sample. AB - OBJECTIVE: To evaluate the internal validity of the subscales of the combined SCL 90 and SCL-90R, the SCL-92, by item response analyses as compared with several previously reported factor analyses of this questionnaire in the literature. METHOD: The SCL-92 questionnaire was mailed to an age- and gender-stratified random sample of Danish citizens. The sample comprised 2040 individuals. The internal structure of the nine factors of the SCL-92 questionnaire was evaluated by Mokken-Loevinger analysis and Rasch analysis. RESULTS: In total, 1153 persons or 58% returned the questionnaire fully completed. Mokken analysis found all scales apart from the psychoticism scale acceptable. The Rasch analysis found most of the subscales to be robust. Minor problems were seen for the scales of phobic anxiety, obsession-compulsion and depression. Analysis of the Global Severity Index showed that the Rasch model was rejected for the full 92-item scale, but not for a scale consisting of the 63 items from the non-psychotic subscales. Spearman correlations among the subscales were all positive (range 0.34-0.79) and so were correlations between each of the subscales and the Global Severity Index (range 0.55-0.91). CONCLUSION: In this sample from the Danish general population the non-psychotic subscales, i.e. the subscales covering psychological distress were observed to function well. In a general population sample, the 63 non-psychotic items primarily appear to reflect one broad dimension of distress. PMID- 15283744 TI - Measuring disordered personality functioning: to love and to work reprised. AB - OBJECTIVE: Current limitations to diagnosing and measuring the personality disorders encouraged a set of studies seeking to provide an alternate approach to modeling and measuring disordered personality function. METHOD: A large set of self-reported descriptors of disordered personality function were factor analyzed in a sample of patients with clinician-diagnosed personality dysfunction, generating 11 lower-order and two higher-order constructs. Subjects and non clinical controls also completed a measure of personality styles underpinning formalized personality disorder groupings. Properties of the refined self-report (SR) measure were assessed in an independent sample of patients with a clinically diagnosed personality disorder. RESULTS: Limitations in 'cooperativeness' and 'coping' formed the higher-order constructs defining disordered personality function, with these constructs relevant to all personality styles. Analyses of SR, corroborative witness (CW) and clinician-rated data in an independent sample supported measuring disordered personality function by our derived 20-item SR measure, and exposed limitations to clinician-based assessment. CONCLUSION: Study findings build to a multi-axial strategy for measuring personality disorder, involving separate dimensional assessment of both disordered personality function and of personality style. PMID- 15283747 TI - cAMP controls human renin mRNA stability via specific RNA-binding proteins. AB - It is now recognized that post-transcriptional mechanisms are pivotal to renin production. These involve factors that modulate renin mRNA stability. In 2003 new data has emerged from work in Australia and Germany that has identified several of the, as many as, 20 or so proteins involved. These include CP1 (hnRNP E1), HuR, HADHB, dynamin, nucleolin, YP-1, hnRNP K and MINT-homologous protein. Cyclic AMP (cAMP) is a crucial regulator of renin secretion as well as transcriptional and post-transcriptional control of expression. Many of the RNA-binding proteins that were identified responded to forskolin, increasing in amount by two to 10 fold. The cAMP mechanisms that regulate renin mRNA target, at least in large part, other genes that presumably encode some of these proteins. The increase in the expression of these then facilitates, sequentially, renin mRNA stabilization and destabilization. Our data, using a battery of different techniques, confirm that CP1 and HuR stabilize renin mRNA, whereas HADHB causes destabilization. These proteins target cis-acting C-rich sequences (in the case of CP1) and AU rich sequences (HuR) in the distal region of the 3'-untranslated region of renin mRNA. We found HADHB was enriched in juxtaglomerular cells and that that within Calu-6 cells HADHB, HuR and CP1 all localized in nuclear subregions, as well as cytoplasm (HADHB and CP1) and mitochondria (HADHB) commensurate with the role each plays in control of renin mRNA stability. The specific proteins that bind to human renin mRNA have begun to be revealed. Cyclic AMP upregulates the binding of several of these proteins, which in turn affect renin mRNA stability and thus overall expression of renin. PMID- 15283748 TI - Controlling the release and production of renin. AB - The renin-angiotensin system (RAS) plays a pivotal role for a variety of cardiovascular functions. The diversity of renin actions is reflected by its complex control. The major stimulus for the release of renin from the vesicles in juxtaglomerular cells is determined by stretch, as induced by changes in arterial pressure. The production of renin underlies a vastly complex control network, which takes place at different levels, such as transcription and translation. With regard to transcription, important regions for binding transcription factors have been identified several years ago, but the conservation of nucleotide sequences throughout different species suggests that there might be additional binding regions of importance. At the post-transcriptional level, the modulation of renin mRNA stability is seems pivotal. The half-life of renin mRNA appears to be controlled by the interaction between several regulatory proteins, most of which are well known in other systems. Moreover, in addition to the modulation of mRNA stability, the translation efficiency seems to play a key role in determining the amount of renin to be produced. PMID- 15283749 TI - Cellular mechanism of renin release. AB - In this review we aim to give a comprehensive overview over the current knowledge of the cellular control of renin release. We hereby focus on the inhibitory effects of calcium on the exocytosis of renin. After a short introduction into general aspects of the regulation of renin release, including a brief summary on the role of the second messengers cAMP and cGMP, we will discuss parts of the literature on the effects of calcium on the renin system together with recent studies from our laboratory, investigating putative calcium influx and extrusion pathways of juxtaglomerular cells. Finally, as the precise mechanisms by which calcium inhibits the exocytosis of renin are far from being understood, we will present some hypotheses on the intracellular events being involved in the suppression of renin release by calcium. PMID- 15283750 TI - Membrane potential and cation channels in rat juxtaglomerular cells. AB - The relationship between membrane potential and cation channels in juxtaglomerular (JG) cells is not well understood. Here we review electrophysiological and molecular studies of JG cells demonstrating the presence of large voltage-sensitive, calcium-activated potassium channels (BK(Ca)) of the ZERO splice variant, which is also activated by cAMP. These channels explain the hyperpolarization, which has been observed after stimulation of renin release with cAMP. In addition, there is now evidence that JG cells express functional L type voltage-dependent calcium channels (Ca(v) 1.2), which in situations with strong depolarization lead to calcium influx and inhibition of renin release. In most in vivo situations the membrane potential is probably protected against depolarization by the BK(Ca) channels. PMID- 15283751 TI - Structure of renal afferent arterioles in the pathogenesis of hypertension. AB - Renal vascular resistance is increased in essential hypertension, as in genetic models of hypertension. Here we review the evidence that this is at least in part due to structural changes in the afferent arterioles. Rat studies show that the renal afferent arteriole is structurally narrowed in young and adult spontaneously hypertensive rats (SHR). Furthermore, in the second generation of crossbred SHRs/normotensive rats (SHR/WKY F(2)-hybrids), a narrowed afferent arteriole lumen diameter at 7 weeks is a predictor of later development of high blood pressure. The reduced lumen diameter of resistance vessels is accompanied by a decrease in media cross-sectional area in SHR and could therefore be due to inhibited growth. Evidence from a primate model of hypertension has shown a negative correlation between left ventricular hypertrophy and afferent arteriole diameter, but apparently no relation to blood pressure. In SHR, the antihypertensive effect of angiotensin converting enzyme (ACE) inhibitors is mediated through renal vascular mechanisms, while ACE inhibitors (like AT(1) antagonists) have a more persistent effect on blood pressure after treatment withdrawal compared with other antihypertensive drugs. Taken together, the evidence suggests that structural narrowing of the renal afferent arteriole could be an important link in the pathogenesis of primary hypertension, at least in the SHR. PMID- 15283752 TI - Systolic pressure and the myogenic response of the renal afferent arteriole. AB - The transmission of elevated blood pressure to the glomerulus and pressure induced glomerular injury play central roles in the pathogenesis of kidney disease and its progression to end-stage renal failure. The renal afferent arteriole sets the pre-glomerular resistance and pressure-induced or 'myogenic' afferent arteriolar vasoconstriction is a primary mechanism protecting the glomerulus from the damaging effects of hypertension. The systolic pressure, being the highest level of pressure attained and most frequent pressure oscillation impacting on the renal vasculature, potentially represents the most damaging component of the blood pressure. Indeed, recent studies indicate that elevations in systolic blood pressure are more closely linked to kidney disease than are elevations in diastolic pressure. However, the current view, derived from dynamic studies of autoregulation, is that the renal vasculature responds passively to pressure signals presented at rates exceeding the myogenic operating frequency (0.2-0.3 Hz in the rat). Thus existing concepts do not explain the mechanisms that normally protect the kidney from elevations in the systolic pressure which are presented at the heart rate (6 Hz in the rat). A recent study from our laboratory addressed this issue. Using a modelling approach and direct measurements of myogenic responses, we found that the afferent arteriole is able to sense and appropriately adjust tone in response to changes in systolic pressure, presented at the heart rate. Key kinetic attributes allowing this vessel to respond in this manner appear to be a very short delay in activation, an unusually rapid rate of vasoconstriction and a longer delay in vasodilation. The present review summarizes this work and presents recent findings addressing the determinants of the myogenic vasoconstriction in the afferent arteriole. PMID- 15283753 TI - Rapid non-genomic effects of aldosterone on rodent vascular function. AB - The main role of aldosterone is to maintain body sodium homeostasis by promoting salt reabsorption in the collecting ducts of the kidney. In the cardiovascular system, aldosterone may be harmful in a number of disease states by inducing fibrosis and vascular dysfunction. The present review describes novel results from several laboratories, which show that aldosterone also has beneficial effects in the cardiovascular system by stimulating the production of nitric oxide (NO) from the endothelium. The effect of aldosterone is seen within minutes, and is not inhibited by blockers of gene transcription, thus pointing to a non-genomic mechanism. Furthermore, this potentially beneficial effect is observed at low physiological concentrations of aldosterone (0.1-10 pm). The effect is mediated by the classical mineralocorticoid receptor, and it involves heat shock protein 90, phosphatidylinositol (PI)-3 kinase, protein kinase B, endothelial nitric oxide synthase, and liberation of NO. It is proposed that in healthy individuals with a functioning NO system, the detrimental effects of aldosterone on cardiovascular function are balanced by activation of the potentially beneficial effect of NO. However, in situations with endothelial dysfunction, such as congestive heart failure and hypertension, the negative effects of aldosterone are unopposed and inhibition of aldosterone is warranted. PMID- 15283754 TI - Calcium handling in afferent arterioles. AB - The cytosolic intracellular calcium concentration ([Ca(2+)](i)) is a major determining factor in the vascular smooth muscle tone. In the afferent arteriole it has been shown that agonists utilizing G-protein coupled receptors recruit Ca(2+) via release from intracellular stores and entry via pathways in the plasma membrane. The relative importances of entry vs. mobilization seem to differ between different agonists, species and preparations. The entry pathway might include different types of voltage sensitive Ca(2+) channels located in the plasmalemma such as dihydropyridine sensitive L-type channels, T-type channels and P/Q channels. A role for non-voltage sensitive entry pathways has also been suggested. The importance of voltage sensitive Ca(2+) channels in the control of the tone of the afferent arteriole (and thus in the control of renal function and whole body control of extracellular fluid volume and blood pressure) sheds light on the control of the membrane potential of afferent arteriolar smooth muscle cells. Thus, K(+) and Cl(-) channels are of importance in their role as major determinants of membrane potential. Some studies suggest a role for calcium activated chloride (Cl(Ca)) channels in the renal vasoconstriction elicited by agonists. Other investigators have found evidence for several types of K(+) channels in the regulation of the afferent arteriolar tone. The available literature in this field regarding afferent arterioles is, however, relatively sparse and not conclusive. This review is an attempt to summarize the results obtained by others and ourselves in the field of agonist induced afferent arteriolar Ca(2+) recruitment, with special emphasis on the control of voltage sensitive Ca(2+) entry. Outline of the Manuscript: This manuscript is structured as follows: it begins with an introduction where the general role for [Ca(2+)](i) as a key factor in the regulation of the tone of vascular smooth muscles (VSMC) is detailed. In this section there is an emphasis is on observations that could be attributed to afferent arteriolar function. We then investigate the literature and describe our results regarding the relative roles for Ca(2+) entry and intracellular release in afferent arterioles in response to vasoactive agents, with the focus on noradrenalin (NA) and angiotensin II (Ang II). Finally, we examine the role of ion channels (i.e. K(+) and Cl(-) channels) for the membrane potential, and thus activation of voltage sensitive Ca(2+) channels. PMID- 15283755 TI - Plasma renin in mice with one or two renin genes. AB - AIM: In the present study we have investigated whether the presence of a second renin gene exerts an overriding influence on plasma renin such that mice with two renin genes have consistently higher renin levels than mice with only one renin gene. METHODS: Plasma renin was determined as the rate of angiotensin I generation using a radioimmunoassay (RIA) kit with (plasma renin concentration, PRC) or without (plasma renin activity, PRA) the addition of purified rat angiotensinogen as substrate. RESULTS: In male 129SvJ, DBA/2 and Swiss Webster mice, strains possessing both Ren-1 and Ren-2, PRC (ng Ang I mL(-1) h(-1)) averaged 178 +/- 36, 563 +/- 57 and 550 +/- 43 while PRA was 2.9 +/- 0.5, 3.6 +/- 0.8 and 7.8 +/- 1.2. In male C57BL/6, C3H and BALB/c mice that express only Ren 1, PRC averaged 426 +/- 133, 917 +/- 105 and 315 +/- 72, and PRA was 3.4 +/- 1.0, 6.9 +/- 1.7 and 4.5 +/- 1.2. In the two renin gene A1AR-/- mice compared with the one renin gene A1AR+/+, PRC averaged 538 +/- 321 and 415 +/- 159 while PRA averaged 3.2 +/- 1.1 and 4.4 +/- 1.4 ng Ang I mL(-1) h(-1). Aldosterone levels showed no significant differences between one renin (C57BL/6, C3H and BALB/c) and two renin (129SvJ, DBA/2 and Swiss Webster) gene mice. Furthermore, by quantitative real-time polymerase chain reaction (RT-PCR) we found no correlation between the number of renin genes and whole kidney renin mRNA levels from one and two renin gene mice. CONCLUSION: Our data show that baseline plasma renin is not systematically higher in mice with two renin genes than in one renin gene mice. Thus, the presence of a second renin gene does not seem to be a major determinant of differences in PRC between different mouse strains. PMID- 15283757 TI - Renal autoregulation in P2X1 knockout mice. AB - Autoregulation of renal blood flow is an established physiological phenomenon, however the signalling mechanisms involved remain elusive. Autoregulatory adjustments in preglomerular resistance involve myogenic and tubuloglomerular feedback (TGF) influences. While there is general agreement on the participation of these two regulatory pathways, the signalling molecules and effector mechanisms have not been identified. Currently, there are two major hypotheses being considered to explain the mechanism by which TGF signals are transmitted from the macula densa to the afferent arteriole. The adenosine hypothesis proposes that the released adenosine triphosphate (ATP) is hydrolysed to adenosine and this product stimulates preglomerular vasoconstriction by activation of A(1) receptors on the afferent arteriole. Alternatively, the P2 receptor hypothesis postulates that ATP released from the macula densa directly stimulates afferent arteriolar vasoconstriction by activation of ATP-sensitive P2X(1) receptors. This hypothesis has emerged from the realization that P2X(1) receptors are heavily expressed along the preglomerular vasculature. Inactivation of P2X(1) receptors impairs autoregulatory responses while afferent arteriolar responses to A(1) adenosine receptor activation are retained. Autoregulatory behaviour is markedly attenuated in mice lacking P2X(1) receptors but responses to adenosine A(1) receptor activation remain intact. More recent experiments suggest that P2X(1) receptors play an essential role in TGF-dependent vasoconstriction of the afferent arteriole. Interruption of TGF-dependent influences on afferent arteriolar diameter, by papillectomy or furosemide treatment, significantly attenuated pressure-mediated afferent arteriolar vasoconstriction in wild-type mice but had no effect on the response in P2X(1) knockout mice. Collectively, these observations support an essential role for P2X(1) receptors in TGF-mediated afferent arteriolar vasoconstriction. PMID- 15283756 TI - Nitric oxide counteracts angiotensin II induced contraction in efferent arterioles in mice. AB - AIM: Efferent arterioles (Ef) are one of the final control elements in glomerular haemodynamics. The influence of nitric oxide (NO) on Ef remains ambiguous. METHODS: To test the hypothesis that endothelial NO plays an important role in this context, afferent arterioles (Af) and Ef of wild-type mice (WT), and Ef of mice lacking the endothelial NO synthetase [eNOS(-/-)] were perfused. Perfusion was performed in Ef via Af (orthograde) as well as from the distal end of Ef (retrograde), which provides an estimate for the importance of substances derived from the glomerulus. Angiotensin II (Ang II) was added in doses ranging from 10( 12) to 10(-6) mol L(-1) to the bath solution. RESULTS: Ang II reduced the luminal diameter of Af to 68 +/- 7 and in Ef to 55 +/- 8% during orthograde, and to 35 +/ 6% during retrograde perfusion (10(-6) mol L(-1) Ang II) in WT. Pre-treatment with N(G)-Nitro-L-arginine-methylester (l-NAME) (10(-4) mol L(-1)) increased the Ang II sensitivity in retrograde (17 +/- 9%) and orthograde perfused Ef (19 +/- 9%). The Ang II sensitivity was enhanced in eNOS(-/-) mice compared with WT, too. Already at a dose of Ang II 10(-9) mol L(-1), luminal diameters diminished to 8 +/- 7 and 7 +/- 4%. CONCLUSION: The increased Ang II sensitivity during L-NAME pre-treatment and in eNOS(-/-) mice indicates a strong counteraction of endothelial derived NO on Ang II induced contraction in Ef. Moreover, Ef are similarly sensitive to Ang II during either retrograde or orthograde perfusion in the absence of NO effects, suggesting that NO mediates, at least in part, the action of potential vasodilatory substances from the glomerulus. PMID- 15283758 TI - Compensation of proximal tubule malabsorption in AQP1-deficient mice without TGF mediated reduction of GFR. AB - AIM: By crossing aquaporin 1 (AQP1)-/- and adenosine 1 receptor (A1AR)-/- mice, we generated an animal model that combines a proximal tubular absorption defect with absence of tubuloglomerular feedback (TGF) regulation of glomerular filtration rate (GFR). The aim of studies in these animals was to determine whether a TGF-induced reduction of GFR is a prerequisite for preventing potentially fatal fluid losses. METHODS AND RESULTS: In contrast to AQP1 deficient mice, AQP1/A1AR-/- mice were found to have a normal GFR. TGF responses were abolished in these animals, in contrast to AQP1-/- mice in which TGF responses of single nephron glomerular filtration rate (SNGFR) were left-shifted. Proximal tubule fluid absorption in AQP1/A1AR-/- mice was reduced to levels previously reported for AQP1-/- mice. However, SNGFR was significantly higher in AQP1/A1AR-/- than AQP1-/- mice (10.6 +/- 0.8 nL min(-1) vs. 5.9 +/- 0.7 nL min( 1)). As a consequence of the normal GFR and the reduced proximal reabsorption distal fluid delivery was markedly higher in the double knockout compared with normal or AQP1-/- mice (5.5 +/- 0.5 nL min(-1) vs. 2.35 +/- 0.3 nL min(-1) in AQP1-/-). Despite the approximate doubling of distal fluid and Cl delivery, AQP1/A1AR-/- mice have a normal salt excretion, normal arterial blood pressure, and only a small increase in plasma renin concentration. CONCLUSION: The ability to compensate for proximal tubule malabsorption without a TGF-induced reduction of GFR attests to a remarkable adaptability of distal tubule transport mechanisms. PMID- 15283759 TI - Current mechanisms of macula densa cell signalling. AB - Macula densa cells couple renal haemodynamics, glomerular filtration and renin release with tubular fluid salt and water reabsorption. These cells detect changes in tubular fluid composition through a complex of intracellular signalling events that are mediated by membrane transport pathways. Increases in luminal fluid sodium chloride concentration result in alterations in cell sodium chloride concentration, cytosolic calcium, cell pH, basolateral membrane depolarization and cell volume. Macula densa signalling then involves the production and release of specific paracrine signalling molecules at their basolateral membrane. Upon moderate increases in luminal sodium chloride concentration macula densa cells release increasing amounts of ATP and decreasing amounts of prostaglandin E(2), thereby increasing afferent arteriolar tone and decreasing the release of renin from granular cells. On the other hand, further increases in luminal concentration stimulate the release of nitric oxide, which serve to prevent excessive tubuloglomerular feedback vasoconstriction. Paracrine signalling by the macula densa cells therefore controls juxtaglomerular function, renal vascular resistance and participates in the regulation of renin release. PMID- 15283760 TI - Mechanisms for macula densa cell release of renin. AB - The juxtaglomerular apparatus in the kidney is important in controlling extracellular fluid volume and renin release. The fluid load to the distal tubule is first sensed at the macula densa site via the entry of NaCl, through a Na, K, 2Cl co-transport mechanism. The next step is unclear, but there is recent evidence of an increased macula densa cell calcium concentration with a reduction in fluid load to the macula densa. An increase in macula densa cell calcium could activate phospholipase A2 to release arachidonic acid, the rate-limiting step in the formation of prostaglandins. Recent evidence suggests that the prostaglandin formed is PGE2, a potent stimulator for renin release. Recent evidence has also shown that adenosine has an important function in the juxtaglomerular apparatus. It stimulates calcium release in afferent arteriolar smooth muscle cells, leading to contraction of the afferent arteriole as part of the tubuloglomerular feedback mechanism, and inhibits renin release. Thus, renin release from the afferent arteriole is mediated partly through formation of PGE2, and partly through the reduction of adenosine formation that inhibits renin production. PMID- 15283761 TI - Mechanisms underlying the antihypertensive functions of the renal medulla. AB - There is good evidence that the renal medulla plays a pivotal role in long-term regulation of blood pressure. 'Renal medullary' blood pressure regulating systems have been postulated to involve both exocrine (pressure natriuresis/diuresis) and endocrine [renal medullary depressor hormone (RMDH)] functions. However, recent studies indicate that pressure diuresis/natriuresis dominates the antihypertensive renal response to increased renal perfusion pressure, suggesting little physiological role for a putative RMDH in compensatory responses to acutely increased blood pressure. The medullary circulation appears to play a key role in mediating pressure diuresis, although the precise mechanisms involved remain controversial. Counter-regulatory vasodilator mechanisms (e.g. nitric oxide), at least partly mediated through cross-talk between the vasculature and the tubular epithelium, protect the medullary circulation from the vasoconstrictor effects of hormonal factors such as angiotensin II. These mechanisms also appear to contribute to compensatory responses to increased salt intake in salt-resistant individuals. Failure of these mechanisms predisposes the organism towards the development of hypertension, appears to underlie the development of some forms of experimental hypertension, and may even contribute to the pathogenesis of essential hypertension. PMID- 15283762 TI - Insights into angiotensin II receptor function through AT2 receptor knockout mice. AB - Angiotensin II signals via at least two receptors termed AT1 and AT2. The function of the AT1 receptor is well defined, while that of the AT2 receptor is still shrouded in uncertainty. AT2 gene-deficient (-/-) mice have been helpful in unravelling the function of the AT2 receptor. We have studied AT2-/- and AT2+/+ mice with classical physiological techniques developed for the rat. We found that although AT2-/- mice have normal glomerular filtration rate, the pressure natriuresis relationship in these mice, compared with AT2+/+ mice, is shifted rightward. Moreover, medullary blood flow fails to increase with increased perfusion pressure while the AT1 receptor expression in the kidneys is increased. We used telemetry and found that AT2-/- mice have about 10 mmHg higher blood pressures than AT2+/+ mice and that their circadian rhythm is disturbed. Moreover, their baroreflexes, as measured by spectral analyses, differs from AT2+/+ controls. The cardiac function of AT2-/- mice is remarkably preserved and the differences are subtle. However, if the mice are given l-NAME hypertension, they exhibit an end-systolic pressure-volume relationship that reveals decreased contractility and probable increased vascular stiffness. Furthermore, the hearts of AT2-/- mice hypertrophy more in response to l-NAME than those of AT2+/+ mice and perivascular fibrosis is increased. DOCA-salt treatment also shows a more rightward pressure-natriuresis relationship in AT2-/- compared with AT2+/+ mice. The renal iNOS expression is increased with DOCA-salt treatment. Our findings support the notion that the AT2 receptor signals antiproliferative and antifibrotic effects and that its presence results in lower blood pressures and lesser responses to secondary forms of hypertension. Technical advances that have allowed us to adapt methods for the rat to the much smaller mouse have facilitated our studies. PMID- 15283763 TI - Volume natriuresis vs. pressure natriuresis. AB - Body fluid regulation depends on regulation of renal excretion. This includes a fast vasopressin-mediated water-retaining mechanism, and slower, complex sodium retaining systems dominated by the renin-angiotensin aldosterone cascade. The sensory mechanisms of sodium control are not identified; effectors may include renal arterial pressure, renal reflexes, extrarenal hormones and other regulatory factors. Since the pioneering work of Guyton more than three decades ago, pressure natriuresis has been in focus. Dissociations between sodium excretion and blood pressure are explained as conditions where regulatory performance exceeds the precision of the measurements. It is inherent to the concept, however, that sudden transition from low to high sodium intake elicits an arterial pressure increase, which is reversed by the pressure natriuresis mechanism. However, such transitions elicit parallel changes in extracellular fluid volume thereby activating volume receptors. Recently we studied the orchestration of sodium homeostasis by chronic and acute sodium loading in normal humans and trained dogs. Small increases in arterial blood pressure are easily generated by acute sodium loading, and dogs appear more sensitive than humans. However, with suitable loading procedures it is possible - also acutely - to augment renal sodium excretion by at least one order of magnitude without any change in arterial pressure whatsoever. Although pressure natriuresis is a powerful mechanism capable of overriding any other controller, it seems possible that it is not operative under normal conditions. Consequently, it is suggested that physiological control of sodium excretion is neurohumoral based on extracellular volume with neural control of renin system activity as an essential component. PMID- 15283764 TI - Sodium coupled bicarbonate transporters in the kidney, an update. AB - Recently five genes have been cloned, which code for sodium dependent bicarbonate transport proteins. These genes belong to the SLC4A gene family. This short review summarizes our knowledge of these gene products with respect to their renal distribution and function. The best characterized members are the SLC4A4 and SLC4A7. SLC4A4 codes for an electrogenic Na(+), HCO(3) (-)-cotransporter (NBCe1), which is present in the basolateral membranes of proximal tubules and is responsible for the bicarbonate efflux here, and thus about 80% of the renal bicarbonate reabsorption. SLC4A7 codes for an electroneutral NBC (called NBC3 and NBCn1), which is present basolaterally in the thick ascending limb and the distal part of the collecting ducts and in intercalated cells (either apically or basolaterally) in the connecting and collecting tubules. In the thick ascending limb NBCn1 may be important for NH(4) (+) reabsorption. SLCA5 codes for an electrogenic NBC (called NBC4 and NBCe2), which based on RT-PCR is located to the kidney but the exact localization awaits a good antibody. This is also the case for the SLC4A8 and SLC4A10 gene products, which are sodium dependent Cl(-), HCO(3) (-) exchangers. The recent development in this field substantially increases our understanding of the complex renal regulation of acid base status. PMID- 15283765 TI - Pharmacotyping of hypokalaemic salt-losing tubular disorders. AB - Long standing confusion exists in the terminology of hypokalaemic salt-losing tubulopathies (SLTs). SLTs are autosomal recessively transmitted and characterized by normotensive secondary hyperreninism/hyperaldosteronism with hypokalaemic metabolic alkalosis. Historically, four phenotypical variants have been described: (1) the (classic) Bartter syndrome (cBS), (2) the hypomagnesaemic hypocalciuric Gitelman syndrome (GS), (3) the hypercalciuric hyperprostaglandin-E syndrome (HPS) or antenatal Bartter syndrome (aBS) and (4) the hyperprostaglandin E-syndrome with sensorineural deafness (HPS + SND). The latter two syndromes are the most severe variants with antenatal manifestation with polyhydramnios and life-threatening course of salt- and water-loss. Defects in five renal membrane proteins involved in electrolyte reabsorption have been identified: In HPS patients mutations in (1) either the furosemide-sensitive sodium-potassium chloride cotransporter NKCC2, or (2) in the potassium channel ROMK have been identified, and (3) HPS + SND is caused by mutations in the beta-subunit of the chloride channels ClC-Kb and -Ka (named barttin), all mimicking the major pharmacological effects of furosemide with minor potassium-wasting in ROMK patients as seen in patients treated with simultaneous furosemide and amiloride, and minor calcium-wasting in Barttin-patients resembling the combination of furosemide and thiazides. (4) cBS is caused by mutations in the chloride channel ClC-Kb with similar clinical characteristics as seen under combination of thiazides and furosemide, (5) GS is caused by mutations in the thiazide-sensitive sodium-chloride cotransporter NCCT resembling the effect of long-term thiazide administration. CONCLUSION: The combination of pharmacology and genetics suggests a new terminology for the above described SLTs: Furosemide-like-SLT for HPS caused by NKCC2-mutations, furosemide/amiloride-like-SLT for HPS caused by ROMK mutations, furosemide/thiazide-like-SLT for HPS + SND, thiazide/furosemide-like SLT for cBS, and thiazide-like-SLT for GS. PMID- 15283766 TI - The role of cyclooxygenases and prostanoid receptorsin furosemide-like salt losing tubulopathy: the hyperprostaglandin E syndrome. AB - Hyperprostaglandin E syndrome/antenatal Bartter syndrome is characterized by NaCl wasting and volume depletion, juxtaglomerula hypertrophy, hyperreninism and secondary hyperaldosteronism. Primary causes are mutations in the gene for Na-K 2Cl-cotransporter, NKCC2, or for potassium channel, ROMK, responsible for medullary NaCl malabsorption. Most intriguing aspect of the syndrome is the association with a massively increased renal prostaglandin production which contributes substantially to the clinical picture of the patients. Therefore the term hyperprostaglandin E syndrome has been introduced. It is unclear how prostaglandins aggravate the NaCl transport deficiency. Aspects to prostaglandin synthesis and receptor-mediated function within the kidney in patients suffering from hyperprostaglandin E syndrome/antenatal Bartter syndrome will be discussed. PMID- 15283767 TI - The renin-angiotensin system in kidney development. AB - All components of the renin-angiotensin system (RAS) are highly expressed in the developing kidney in a pattern suggesting a role for angiotensin II in renal development. In support of this notion, pharmacological interruption of angiotensin II type-1 (AT(1)) receptor signalling in animals with an ongoing nephrogenesis produces specific renal abnormalities characterized by papillary atrophy, abnormal wall thickening of intrarenal arterioles, tubular atrophy associated with expansion of the interstitium, and a marked impairment in urinary concentrating ability. Similar changes in renal morphology and function develop also in mice with targeted inactivation of genes encoding renin, angiotensinogen, angiotensin-converting enzyme, or both AT(1) receptor isoforms simultaneously. Taken together, these results clearly indicate that an intact signalling through AT(1) receptors is a prerequisite for normal renal development. The present report mainly reviews the renal abnormalities induced by blocking the RAS pharmacologically in experimental animal models. In addition, pathogenetic mechanisms are discussed. PMID- 15283768 TI - The role of the RAS in programming of adult hypertension. AB - The aetiology of cardiovascular disease originally included two components: a genetic component and an environmental or lifestyle component. Increasing epidemiologic evidence has been accumulating during the last decades indicating the importance of a third component: the influence of the environment during foetal development. Poor living conditions resulted in a high infant mortality and influenced the incidence of cardiovascular diseases in adulthood despite better living conditions (A. Forsdahl. Br J Prev Soc Med 1977; 31, 91-95). An association between pre-natal growth pattern and the rate of death from cardiovascular disease in adulthood was reported (D.J. Barker, P.D. Winter, C. Osmond, B. Margetts & S.J. Simmonds. Lancet 1989; 2, 577-580). Men from Hartfordshire (UK), born between 1911 and 1930 were investigated. The investigations showed that men with the lowest weight at birth and at 1 year of age had the highest risks of death from cardiovascular disease (D.J. Barker, P.D. Winter, C. Osmond, B. Margetts & S.J. Simmonds. Lancet 1989; 2, 577-580). These findings suggested that factors in the perinatal environment could programme an individual for later risk of development of cardiovascular disease compared with someone born with a normal weight. Numerous studies have since confirmed these initial findings of an inverse relationship between early growth pattern and cardiovascular disease in adulthood. PMID- 15283769 TI - Cyclooxygenase-2 and the renal renin-angiotensin system. AB - In the kidney, cyclooxygenase-2 (COX-2) is expressed in the macula densa/cTALH and medullary interstitial cells. The macula densa is involved in regulating afferent arteriolar tone and renin release by sensing alterations in luminal chloride via changes in the rate of Na(+)/K(+)/2Cl(-) cotransport, and administration of non-specific cyclooxygenase inhibitors will blunt increases in renin release mediated by macula densa sensing of decreases in luminal NaCl. High renin states [salt deficiency, angiotensin converting enzyme (ACE) inhibitors or angiotensin receptor blockers, diuretic administration or experimental renovascular hypertension] are associated with increased macula densa/cTALH COX-2 expression. Furthermore, there is evidence that angiotensin II and/or aldosterone may inhibit COX-2 expression. In AT1 receptor knockout mice, COX-2 expression is increased similar to increases with ACE inhibitors or AT1 receptor blockers. Direct administration of angiotensin II inhibits macula densa COX-2 expression. Previous studies demonstrated that alterations in intraluminal chloride concentration are the signal for macula densa regulation of tubuloglomerular feedback and renin secretion, with high chloride stimulating tubuloglomerular feedback and low chloride stimulating renin release. When cultured cTALH or macula densa cells were incubated in media with selective substitution of chloride ions, COX-2 expression and prostaglandin production were significantly increased. A variety of studies have indicated a role for COX-2 in the macula densa mediation of renin release. In isolated perfused glomerular preparations, renin release induced by macula densa perfusion with a low chloride solution was inhibited by a COX-2 inhibitor but not a COX-1 inhibitor. In vivo studies in rats indicated that increased renin release in response to low-salt diet, ACE inhibitor, loop diuretics or aortic coarctation could be inhibited by administration of COX-2-selective inhibitors. In mice with genetic deletion of COX-2, ACE inhibitors or low-salt diet failed to increase renal renin expression, although renin significantly increased in wild type mice. In contrast, in COX-1 null mice there were no significant differences in either the basal or ACE inhibitor-stimulated level of renal renin activity from plasma or renal tissue compared with wild type mice. In summary, there is increasing evidence that COX-2 expression in the macula densa and surrounding cortical thick ascending limb cells is regulated by angiotensin II and is a modulator of renal renin production. These interactions of COX-2 derived prostaglandins and the renin angiotensin system may underlie physiological and pathophysiological regulation of renal function. PMID- 15283770 TI - The renin-angiotensin system in kidney development: role of COX-2 and adrenal steroids. AB - Recent data from studies in rodents with targeted gene disruption and pharmacological antagonists have shown that the renin-angiotensin-aldosterone system (RAAS) and cyclooxygenase type-2 (COX-2) are necessary for late stages of kidney development. The present review summarizes data on the developmental changes of RAAS and COX-2 and the pathways by which they are activated; their possible interplay and the mechanisms by which they affect kidney development. Intrarenal and circulating renin and angiotensin II (ANG II) are stimulated at birth in most mammals. In rats, renin and ANG II stay significantly elevated in the suckling period while aldosterone stabilizes at an adult level. COX-2 is stimulated in thick ascending limb of Henle's loop in the suckling period at a time when urine concentrating ability is not developed. Data suggest that this induction is mediated by combined low plasma glucocorticoid concentration and by a low NaCl intake. Studies with selective inhibitors of COX-2 and COX-2 null mice show that COX-2 activity stimulates renin secretion from JG-cells during postnatal kidney development and that lack of COX-2 activity leads to pathological change in cortical architecture and eventually to renal failure. In the postnatal period, ANG II initiates and maintains pelvic and ureteric contractions necessary for urine flow. Lack of ANG II in the neonatal period is thought to cause injury by a chronic increase of renal pelvic pressure. Aldosterone is crucial for survival and growth in the neonatal period through its effects on sodium reabsorption and the intrarenal sensitivity to aldosterone is increased in the postnatal period. Final maturation of the kidney occurs through an intimate interplay between a low dietary sodium intake and a non-responsive HPA-axis which stimulates cortical COX-2 activity. COX-2 supports increased activity of the RAAS and may contribute to a low concentrating ability. PMID- 15283771 TI - Exploring type I angiotensin (AT1) receptor functions through gene targeting. AB - The renin-angiotensin system (RAS) modulates a diverse set of physiological processes including development, blood pressure, renal function and inflammation. The principal effector molecule of this system, angiotensin II, mediates most of these actions. The classically recognized functions of the RAS are triggered via the type 1 (AT(1)) class of angiotensin receptors. Pharmacological blockade of the AT(1) receptor lowers blood pressure and slows the progression of cardiovascular and renal diseases. Gene-targeting technology provides an experimental approach for precisely dissecting the physiological functions of the RAS. Here, we review how gene-targeting experiments have elucidated AT(1) receptor functions. PMID- 15283772 TI - Transgenic mice for studies of the renin-angiotensin system in hypertension. AB - Hypertension is a polygenic and multi-factorial disorder that is extremely prevalent in western societies, and thus has received a great deal of attention by the research community. The renin-angiotensin system has a strong impact on the control of blood pressure both in the short- and long-term, making it one of the most extensively studied physiological systems. Nevertheless, despite decades of research, the specific mechanisms implicated in its action on blood pressure and electrolyte balance, as well as its integration with other cardiovascular pathways remains incomplete. The production of transgenic models either over expressing or knocking-out specific components of the renin-angiotensin system has given us a better understanding of its role in the pathogenesis of hypertension. Moreover, our attention has recently been refocused on local tissue renin-angiotensin systems and their physiological effect on blood pressure and end-organ damage. Herein, we will review studies using genetic manipulation of animals to determine the role of the endocrine and tissue renin-angiotensin system in hypertension. We will also discuss some untraditional approaches to target the renin-angiotensin system in the kidney. PMID- 15283773 TI - Genetic targeting of the brain renin-angiotensin system in transgenic rats: impact on stress-induced renin release. AB - The advance of genetic technologies to permit tissue-specific targeted gene manipulation allowed the development of transgenic models with alterations of the renin-angiotensin (RAS) solely in the brain. We have used such methodology to develop a transgenic rat with a brain specific alteration of the RAS [TGR(ASrAOGEN)], in order to elucidate a causative role for the brain RAS and its relevance in different pathophysiological processes. The TGR(ASrAOGEN) rats have decreased levels of angiotensinogen (AOGEN) throughout the brain because of an antisense inhibition of the astroglial AOGEN synthesis. In this review we aimed at summarizing the experience obtained from utilizing the TGR(ASrAOGEN) rat model to study the brain RAS and present novel results providing evidence for the involvement of this system in stress-induced renin release. PMID- 15283774 TI - Consomic rat model systems for physiological genomics. AB - A consomic rat strain is one in which an entire chromosome is introgressed into the isogenic background of another inbred strain using marker-assisted selection. The development and physiological screening of two inbred consomic rat panels on two genetic backgrounds (44 strains) is well underway. Consomic strains enable one to assign traits and quantitative trait loci (QTL) to chromosomes by surveying the panel of strains with substituted chromosomes. They enable the rapid development of congenic strains over a narrow region and enable one to perform F2 linkage studies to positionally locate QTL on a single chromosome with a fixed genetic background. These rodent model systems overcome many of the problems encountered with segregating crosses where even if linkage is found, each individual in the cross is genetically unique and the combination of genes cannot be reproduced or studied in detail. For physiologists, consomics enable studies to be performed in a replicative or longitudinal manner to elucidate in greater detail the sequential expression of genes responsible for the observed phenotypes of these animals. They often provide the best available inbred control strains for physiological comparisons with the parental strains and they enable one to assess the impact of a causal gene region in a genome by allowing comparisons of the effect of replacement of a specific chromosome on a disease susceptible or a resistant genomic background. Consomic rat strains are proving to be a unique scientific resource that can greatly extend our understanding of genes and their role in the regulation of complex function and disease. PMID- 15283775 TI - Structuring health needs assessments: the medicalisation of health visiting. AB - This paper draws on Foucault to understand the changing discourse and impact of structured 'health needs assessments' on health visiting practice. Literature about this activity makes little mention of the long-standing social purposes of health visiting, which include surveillance of vulnerable and invisible populations, providing them, where needed, with help and support to access protective and supportive services. Instead, the discourse has been concerned primarily with an epidemiological focus and public health, which is associated with risk factors and assessments. The use of pre-defined needs assessment schedules suggests that health visiting activity can be sanctioned and clients' needs serviced only if they reach the threshold of pre-determined, epidemiologically-defined risk. Their effect on practice is examined through a conversation analysis of ten health visitor/client interactions using two different structured needs assessment tools. The study indicates that the health visitors, like their clients, were controlled by institutional expectations of their role; analysis of their conversations shows how they achieved the requirements of the organisational agenda. Structuring client needs and health visiting practice through the use of formal needs assessment tools emphasises the epidemiological focus of the health service above the need to arrange support for vulnerable individuals. In this respect, it serves as a marker in the continued medicalisation of health visiting. PMID- 15283776 TI - Accounting for Irish Catholic ill health in Scotland: a qualitative exploration of some links between 'religion', class and health. AB - This paper considers the ways in which accounts from Glasgow Catholics diverge from those of Protestants and explores the reasons why people leave jobs, including health grounds. Accounts reveal experiences distinctive to Catholics, of health-threatening stress, obstacles to career progression within (mainly) private-sector organisations, and interactional difficulties which create particular problems for (mainly) middle class men. This narrows the employment options for upwardly mobile Catholics, who may then resort to self-employment or other similarly stressful options. The paper considers whether the competence of Catholics or Catholic cultural factors are implicated in thwarting social mobility among Catholics or, alternatively, whether institutional sectarianism is involved. We conclude that, of these options, theories of institutional sectarianism provide the hypothesis which currently best fits these data. In Glasgow, people of indigenous Irish descent are recognisable from their names and Catholic background and are identified as Catholic by others. Overt historical exclusion of Catholics from middle class employment options now seems to take unrecognised forms in routine assumptions and practices which restrict Catholic employment opportunities. It is argued that younger Catholics use education to overcome the obstacles to mobility faced by older people and circumvent exclusions by recourse to middle class public-sector employment. This paper aims to link historical, structural and sectarian patterns of employment experience to accounts of health and work, and in so doing to contribute to an explanation for the relatively poor health of Catholic Glaswegians with Irish roots. PMID- 15283777 TI - A continuum of risk? The management of health, physical and emotional risks by female sex workers. AB - This paper describes the findings from a 10-month ethnographic study of the female sex industry in a large British city. I argue that sex workers construct a continuum of risk which prioritizes certain types of dangers depending on the perceived consequences and the degree of control individuals consider they have over minimising the likelihood of a risk occurring. Although health-related matters are a real concern to many women, because they generally have comprehensive strategies to manage health risks at work, this risk category is given a low priority compared with other risks. The risk of violence is considered a greater anxiety because of the prevalence of incidents in the sex work community. However, because of comprehensive screening and protection strategies to minimise violence, this type of harm is not given the same level of attention that emotional risks receive. By using a continuum of risk to understand how sex workers perceive occupational hazards in prostitution, further understanding can be gained about the nature of risk in prostitution, sex workers' routines and the organisational features of the sex industry. In addition, the implications for health policy are discussed, suggesting that the emotional consequences of selling sex should be considered as much as the tangible, physical risks of prostitution. PMID- 15283778 TI - After the euphoria: HIV medical technologies from the perspective of their prescribers. AB - This paper focuses on the relationship of HIV medical technologies to current styles of medical practice and highlights issues posed by the technologies for those working and/or living with HIV. The paper examines HIV anti-retroviral combination therapies and associated tests from the perspective of their prescribers. The prescribers were interviewed during the later part of 2002 at three London HIV clinics. Their comments, considered in light of other recent studies in the field, suggest that current therapies are part of a transitional phase in the epidemic which informs the identification and negotiation of known risks and uncertainty. An undetermined but extended life expectancy, afforded by anti-retroviral therapies, is understood against risk of iatrogenic diseases and/or viral drug resistance. The tension arising in this situation of unwanted and even uncertain phenomena poses ethical dilemmas and affects doctor/patient relations. Indeed, it also contributes to a reconfiguring of the lived experience of managing HIV. While the new technologies have offered considerable advances in the medical management of HIV, they are altering the nature of HIV medicine both materially and socially. The scenario is further complicated by the uneven allocation of resources and different patient health and disease states. The heterogeneity of resources, disease states and technological effects points to the need for ongoing and extended evaluation as the relationship between these and the everyday practice of medicine continues to change. PMID- 15283779 TI - Entangled identities and psychotropic substance use. AB - This paper reports the findings of a grounded theory study investigating drug users' concerns and experiences of their oral health. The aim of this paper is to demonstrate how the findings relate to various strands of literature which focus on processes and discourses of recovery from problematic drug use (biographical reconstruction), the chronic illness literature (biographical disruption), public/private discourses and the myth of addiction. Data were collected from four focus groups containing a total of 25 participants, and 15 in-depth interviews. Participants were recruited from drug detoxification programmes (27), recovery units following detoxification (9) and a drug rehabilitation unit (4). Data analysis revealed that the core concern of drug users' was talking about the 'entangled' nature of their identity whilst they were on drugs. Such 'entangled identities' emerged through what appeared to be a gradual sedimentation process of drug-using habits and routines that replaced those of the everyday self. Other concerns were distancing one's self from the drug using self (involving expressions of disgust) and recovery processes (disentangling). The paper discusses each of these core problems in the light of the literature on the recovery from drug use, the chronic illness literature and the myth of addiction. It concludes by briefly reflecting on problematic psychotropic substance use as another form of biographical disruption formed on the basis of a dialectic between private discourses of the entangled self and public discourses of addiction. It suggests that further work should be conducted in these areas. PMID- 15283780 TI - The use of technology at home: what patient manuals say and sell vs. what patients face and fear. AB - Over the past 15 years, the use of specialised medical equipment by patients at home has increased in most industrialised countries. Adopting a conceptual framework that brings together two research perspectives, i.e. the sociology of technology and the sociology of illness, this paper empirically examines why and how patients use health technology at home and in the broader social world. Our study compares and contrasts the use of four interventions: antibiotic intravenous therapy, parenteral nutrition, peritoneal dialysis and oxygen therapy. We conducted interviews with patients (n = 16) and caregivers (n = 6), and made direct observations of home visits by nurses (n = 16). The content and structure of patient manuals distributed by major manufacturers and hospitals were analysed (n = 26). The aim of our study was to determine how technology was supposed to be used versus how it was actually used. This study shows that patients are deeply ambivalent about the benefits and drawbacks of technology, and that these advantages and disadvantages are shaped by the various places in which the technology is used. While technology can be pivotal in making patients autonomous and able to participate in the social world, it also imposes heavy restrictions that are intimately interwoven with the nature of the particular disease and with the patient's personal life trajectory. PMID- 15283781 TI - A disputed occupational boundary: operating theatre nurses and Operating Department Practitioners. AB - Traditionally, surgeons (and to a lesser extent anaesthetists) have been assisted primarily by nurses. This role has been threatened in recent years, in the UK NHS (and elsewhere), by a relatively new profession, that of the Operating Department Practitioner (ODP). The ODP profession is still in the process of establishing itself as a 'full' profession within UK health care. While occupational boundary disputes between professions are common in health care, it is unusual for them to become as overt as the dispute we will analyse in this paper. Drawing on fieldwork observations and interviews conducted in operating theatres, as well as documentary sources, we will show how this dispute arose, how it is manifested at both the micro and the macro level, and how both groups involved justify their positions, drawing on surprisingly similar rhetorical strategies. A further unusual feature of this dispute is the fact that, unlike many attempts by managers to substitute one type of labour for another, issues of cost are relatively unimportant, as both theatre nurses and ODPs earn similar salaries. PMID- 15283783 TI - Pseudoacne of the nasal crease: a new entity? AB - In this article we describe a clinical entity appearing in seven preadolescent patients who presented with chronic red papules within a prominent nasal crease. Milia were also noted in the nasal crease, but there was no evidence of acne vulgaris. The duration of symptoms was 4 months to 2 years, and lesions ranged from inflamed red papules, which were treated with topical antiinflammatory medications, to scarred white papules requiring excision. Histologic evaluation of two lesions revealed keratin granulomas that were likely derived from ruptured, inflamed milia. Due to its similarity in appearance to acne vulgaris, but different pathogenesis and clinical course, we suggest naming this newly described entity "pseudoacne of the nasal crease." PMID- 15283784 TI - Epidermal nevus syndromes: clinical findings in 35 patients. AB - Of the patients with epidermal nevi, 10-18% may have disorders of the eye, nervous, and musculoskeletal systems. A predisposition to malignant neoplasms in ectodermal and mesodermal structures may also be found. There are six different epidermal nevus syndromes described so far: Proteus, congenital hemidysplasia with ichthyosiform nevus and limb defect syndrome, phakomatosis pigmentokeratotica, sebaceous nevus, Becker nevus, and nevus comedonicus. Thirty five patients with epidermal nevus syndrome seen at the National Institute of Pediatrics in Mexico City during a 31-year period are described. This syndrome represented 7.9% of 443 patients with epidermal nevi; its relative frequency was 1 case per 11,928 pediatric patients and 1 case per 1080 dermatologic patients. Nine epidermal nevus syndrome patients (26%) had Proteus syndrome. Sebaceous nevus syndrome was found in six patients (17%), while the nevus comedonicus syndrome was found in three (8%). Two patients were diagnosed with phakomatosis pigmentokeratotica and one patient with congenital hemidysplasia with ichthyosiform nevus and limb defect syndrome. This is the first report of phakomatosis pigmentokeratotica and congenital hemidysplasia with ichthyosiform nevus and limb defect syndrome in Mexican patients. One patient had an inflammatory linear verrucous epidermal nevus with systemic involvement. Thirteen patients (37%) had keratinocytic nevi with systemic involvement. We propose the keratinocytic nevus syndrome to be defined as the association of a keratinocytic nevus with neuronal migration and/or musculoskeletal disorders in addition to a higher risk for mesodermal neoplasms. PMID- 15283785 TI - Lichen striatus: description of 89 cases in children. AB - Lichen striatus (LS) is a benign, self-limited, linear, inflammatory dermatosis of unknown etiology that usually affects children. We analyzed 89 cases in regard to age of appearance, sex, race, symptoms, seasonal incidence, localization of lesions and affected side of the body, and presence of atopy. Lesions predominated on the inferior limbs, with no preponderance of any age, and were asymptomatic in the majority of the instances. There was no difference in the incidence of LS in regard to the season of the year. A possible association of lichen sclerosus with atopy and pruritus was observed. PMID- 15283786 TI - Tinea capitis in Kwa-Zulu Natal, South Africa. AB - Tinea capitis is the most common dermatophyte infection in children. The hair involvement can be classified as endothrix, ectothrix, or favus, and the clinical appearance is variable. The goal of this study was to determine the demography, etiology, and clinical patterns of tinea capitis in South Africa. A prospective, cross-sectional study was conducted over a 1-year period. All cases were classified clinically and subject to Wood light examination, microscopy, and culture. One hundred patients were studied. The male:female ratio was 1.4:1. The mean age was 4.6 years (range 1-11 years). Trichophyton violaceum was isolated in 90% of positive cultures. Wood light was positive in one patient with Microsporum gypseum. The most common clinical variety was the "black dot" type, seen in 50% of patients. Twenty percent of the children presented with more than one clinical type simultaneously. We concluded that the most common cause of tinea capitis in South Africa is T. violaceum. The presentation is variable. PMID- 15283787 TI - A syndrome of hemimaxillary enlargement, asymmetry of the face, tooth abnormalities, and skin findings (HATS). AB - Hemimaxillofacial dysplasia and segmental odontomaxillary dysplasia appear to be the same syndrome, having the common features of unilateral abnormalities of bone, teeth, gums, and skin. Oral manifestations are the hallmark of this condition. Those affected are generally recognized in childhood and may have partial anodontia, abnormal spacing of the teeth, delayed eruption, and gingival thickening of the affected segment. Reported cutaneous manifestations include facial asymmetry, Becker's nevus, "hairy nevus," lip hypopigmentation, discontinuity of the vermilion border, depression of the cheek, and erythema. The oral lesions do not appear to be progressive. We describe a child with features consistent with hemimaxillofacial dysplasia/segmental odontomaxillary dysplasia. Findings of a biopsy specimen from the cheek confirmed the presence of a Becker's nevus. Cutaneous findings reported in the previous 31 cases are reviewed and summarized. The acronym HATS (hemimaxillary enlargement, asymmetry of the face, tooth abnormalities, and skin findings) is introduced to reflect the spectrum of abnormalities in bone, teeth, and skin that may be seen in this developmental disorder. PMID- 15283788 TI - Subungual chondroblastoma in a 9-year-old girl. AB - Chondroblastoma is an uncommon primary bone tumor, mainly found in the epiphyses of long bones. We describe a 9-year-old girl who presented with a chondroblastoma as a subungual mass in the fifth toe. Radiographs showed an expansive, calcified tumor of the distal phalanx. Histologic examination after excision revealed chondroid differentiation, active mitosis, multinucleated giant cells, calcification, and necrosis. There was no recurrence of the lesion after surgical excision. The purpose of this report is to document this unusual event that occurred in such a short bone as the distal phalanx of the fifth toe, mimicking a dermatologic entity. PMID- 15283789 TI - Unilateral aplasia cutis congenita on the leg. AB - We present the case of a newborn male with aplasia cutis congenita on the extensor side of the right leg, with unilateral absence of skin on the lower limb. There was no abnormality in pregnancy or birth and there was no associated malformation or skin disease such as blistering or nail pathology. The management of this large ulcer was conservative, using silver sulfadiazine ointment, and healing occurred within 3 months. The follow-up after 21 months showed little scar formation and no handicap regarding function and appearance. The psychomotor development was normal. According to the classification outlined by Frieden, we classified this condition as type VII aplasia cutis congenita. PMID- 15283790 TI - Aleukemic congenital leukemia cutis. AB - A newborn girl had typical "blueberry muffin" skin lesions, which showed histopathologic features of myelomonocytic leukemia cutis. We could not demonstrate leukemic infiltration of bone marrow in four aspirates. Her course was complicated with primary pulmonary hypertension, which led to death at 7 months of age. We emphasize the persistence of skin lesions in the absence of bone marrow infiltration by leukemia throughout the course of the disease. PMID- 15283791 TI - A subungual nevus in a Filipino child. AB - A 2.5-year-old Filipino girl with a progressively growing, black, subungual lesion is described. A biopsy was performed because of the progressive increase in size of the lesion and its periungual involvement. Histologic examination revealed a junctional nevus. To our knowledge, this represents the first report of subungual nevus in a Filipino child. The need for histologic diagnosis prior to definitive surgery cannot be overemphasized. PMID- 15283792 TI - Palmoplantar eccrine hidradenitis: seven new cases. AB - Palmoplantar eccrine hidradenitis is a self-limited disease characterized by painful erythematous papules and nodules of abrupt onset on the soles, and less frequently on the palms, of young individuals in good health. We describe seven children, four girls and three boys, between 4 and 12 years of age, with characteristic cutaneous and histopathologic findings of palmoplantar eccrine hidradenitis. All patients had complete resolution of their lesions within 2-4 weeks without treatment, however, one child experienced recurrences. All skin biopsy specimens showed a deep dermal mixed infiltrate with abundant neutrophils surrounding eccrine sweat glands, the histologic hallmark of the disease. Palmoplantar eccrine hidradenitis is a distinct clinical entity in which physical activity, excessive sweating, and prolonged wetness are possible triggering factors. The regression of the lesions is usually rapid, with complete clearance after 1 month, although there may be recurrent episodes. PMID- 15283793 TI - Miliary neonatal hemangiomatosis with fulminant heart failure and cardiac septal hypertrophy in two infants. AB - Miliary neonatal hemangiomatosis is a rare, life-threatening condition associated with cutaneous and multiorgan involvement. We report two infants with this condition who had fulminant cardiac failure and cardiac septal hypertrophy. The first was a 5-day-old boy who presented with increasing numbers of cutaneous hemangiomata associated with worsening cardiac failure. Magnetic resonance imaging (MRI) showed extensive hepatic hemangioma. Despite treatment with systemic corticosteroids and subcutaneous interferon alfa-2b his disease progressed. Hepatic artery embolization was unsuccessful. The infant died of congestive cardiac failure at 6 weeks of age. Postmortem examination showed a massively enlarged cardiac interventricular septum and biventricular hypertrophy. The second patient was a 1-week-old girl who also had cutaneous hemangioma and cardiac decompensation. MRI showed extensive intrahepatic involvement. She was treated early with corticosteroids and interferon alpha, which resulted in involution of the cutaneous and hepatic lesions. Cardiac septal hypertrophy did not persist at late follow-up, and the association of miliary neonatal hemangiomatosis and cardiac septal hypertrophy has not yet been established. PMID- 15283795 TI - Plica neuropathica: a mystery. AB - Plica neuropathica is a rare, acquired condition of sudden onset characterized by irreversible matting of hair. The exact mechanism is not understood, but multiple factors such as physical conditions, chemical agents, and behavioral factors play main roles. Cutting of the affected hair and avoiding all possible triggering factors are the only effective treatments. We report a 14-year-old boy with this condition. PMID- 15283794 TI - Malignant melanoma associated with lichen sclerosus in the vulva of a 10-year old. AB - Malignant melanoma of the vulva in childhood is a rare neoplasm. Lichen sclerosus of the vulva in childhood is also a rare disease. The association of these two rare lesions in the vulva of young girls is extremely rare. We present a 10-year old white girl with malignant melanoma associated with lichen sclerosus of the vulva. She had dark pigmentation of both the labia minora and posterior fourchette. The inner labia majora and fourchette showed whitish, glistening areas of skin. Histologic examination found mostly an in situ lentiginous/mucosal melanoma with focal invasion to a depth of 0.44 mm in the left upper labium majus. All specimens showed evidence of lichen sclerosus. Partial vulvectomy was performed, and no metastases were detected at the time of treatment. The patient has been disease free for the 12 months after treatment. It is critical for physicians to realize that melanoma can occur in children, and although rare, can occur in the vulva. We feel that lichen sclerosus in this instance may represent a pattern of host immune response to melanoma. PMID- 15283796 TI - Elejalde syndrome: report of a case and review of the literature. AB - Elejalde syndrome is a rare autosomal recessive condition, with only 10 reported cases through 2001. It is characterized by silvery hair, pigment abnormalities, and profound central nervous system dysfunction. The differential diagnosis includes Griscelli and Chediak-Higashi syndromes, which present with silvery hair, pigment abnormalities, central nervous system alterations, and severe immunologic dysfunction. We report a 6-year-old girl with Elejalde syndrome and review Elejalde, Griscelli, and Chediak-Higashi syndromes. PMID- 15283797 TI - Milia-like idiopathic calcinosis cutis. AB - Milia-like idiopathic calcinosis cutis is a rare entity. Only 17 cases have been reported so far. Two-thirds of these have been associated with Down syndrome. We report the fifth case occurring in a child without Down syndrome. Milia-like idiopathic calcinosis cutis has long been regarded as a peculiar subtype of idiopathic calcinosis cutis. The pathogenesis of the disorder remains unclear. PMID- 15283798 TI - Severe monilethrix associated with intractable scalp pruritus, posterior subcapsular cataract, brachiocephaly, and distinct facial features: a new variant of monilethrix syndrome? AB - Monilethrix is a rare developmental hair shaft defect characterized by small elliptical node-like deformities with increased hair fragility resulting in partial or diffuse alopecia. The disorder is usually transmitted in an autosomal dominant fashion with incomplete penetrance and variable expressivity, but autosomal recessive inheritance has also been reported. It is thought to be without systemic involvement, whereas keratosis pilaris and follicular papules are almost invariably associated features. We describe an instance of monilethrix in a 9-year-old boy from consanguineous parents, characterized by universal dystrophic alopecia associated with intractable scalp pruritus, diffuse keratosis pilaris, and bilateral posterior subcapsular cataracts. His disease was further characterized by physical underdevelopment and distinct features of hypertelorism, a wide-based nose, long philtrum, relatively large mouth with thick lower lip, enlarged forehead, small, receding chin, short neck, and rounded (ultrabrachycranial) skull. The findings in our patient suggest that "monilethrix syndrome" is an appropriate term for defining the instances of monilethrix associated with other abnormalities. We conclude that our patient may represent a new and severe, autosomal recessive variant of monilethrix syndrome. PMID- 15283799 TI - Aplasia cutis congenita after methimazole exposure in utero. AB - We describe a patient who was exposed to the antithyroid drug methimazole during the first 6 weeks of gestation and was born prematurely with scalp and skull defects associated with facial asymmetry. A review of the literature seems to support the hypothesis that methimazole is a potential teratogen. Although the risk of birth defects is low with clinically applied doses of the drug, it cannot be regarded as safe and should therefore be avoided in the treatment of pregnant women. PMID- 15283800 TI - Calcipotriene and corticosteroid combination therapy for vitiligo. AB - Corticosteroids and photochemotherapy, using a combination of psoralen and ultraviolet A (PUVA) exposure, are the most widely prescribed therapies for vitiligo. These treatments are not uniformly effective and many patients have inadequate responses. Calcipotriene has been shown to be effective in adults and children with psoriasis when used as monotherapy and in combination with corticosteroids and phototherapy. We hypothesized that since the mechanisms of action for calcipotriene and corticosteroids are different, patients may develop more repigmentation with a combination of the two agents, while decreasing the side effects from both agents. Twelve patients with vitiligo (average age 13.1 years) were advised to use topical corticosteroids in the morning and topical calcipotriene in the evening. Of the 12 patients, 83% responded to therapy, with an average of 95% repigmentation by body surface area. Four of the patients who responded had previously failed trials of topical corticosteroids alone. All of the patients in this group had repigmentation. Eyelid and facial skin responded best to this therapy. None of the patients had adverse reactions to the treatment. Our results show that topical calcipotriene in combination with corticosteroids can repigment vitiligo, even in those patients who were previous topical corticosteroid failures. PMID- 15283801 TI - Itraconazole in the treatment of tinea capitis caused by Microsporum canis: experience in a large cohort. AB - Mycotic scalp infection caused by Microsporum canis is one of the more recalcitrant disorders, with increasing incidence during the last decade. We report our experience with administration of itraconazole in 163 children (86 girls, 77 boys) with M. canis tinea capitis. Fifty-five patients had previous treatment with terbinafine without success. In all children, the dosage of itraconazole was adjusted according to body weight, with 5 mg/kg/day given in a continuous regimen either as a capsule (116 patients) or an oral suspension (47 patients). In all children, there was both clinical and mycologic cure after a mean treatment period of 39 +/- 12 days (range 10-77 days). Eleven children (6.7%) had side effects: diarrhea in five children, cutaneous eruption in four, and abdominal pain in two. Itraconazole was effective and safe for the treatment of M. canis tinea capitis. PMID- 15283802 TI - What syndrome is this? Peutz-Jeghers syndrome. PMID- 15283803 TI - Congenital plantar nodule in an infant. PMID- 15283804 TI - Stiff skin syndrome is highly heterogeneous, and congenital fascial dystrophy is its distinct subset. PMID- 15283807 TI - Re: Candida albicans: an immune response modifier. PMID- 15283808 TI - Juvenile pityriasis rubra pilaris. PMID- 15283809 TI - Linear juvenile xanthogranuloma. PMID- 15283810 TI - The unidentified parasite: a probable case of north american cuterebrid myiasis in a pediatric patient. PMID- 15283811 TI - Giant nevus spilus and centrofacial lentiginosis. PMID- 15283812 TI - Pachyoncyhia congenita type 2. PMID- 15283813 TI - Additional aspects of keratitis, ichthyosis, and deafness (KID) syndrome. PMID- 15283815 TI - Cutaneous mastocytosis in children: an Indian experience. PMID- 15283816 TI - Xanthelasmoid mastocytosis in flexural areas. PMID- 15283817 TI - Symmetrical lividity in an infant. PMID- 15283818 TI - Tracheal intubation without intravenous access. PMID- 15283819 TI - What does analgesia mask? PMID- 15283820 TI - Anesthesia for thymectomy in children with myasthenia gravis. PMID- 15283821 TI - Low doses of rocuronium during remifentanil-propofol-based anesthesia in children: comparison of intubating conditions. AB - BACKGROUND: In this prospective double-blind study, intubation conditions were compared at 90 s following two different low doses of rocuronium during remifentanil and propofol anesthesia in children undergoing ambulatory procedures. METHODS: Forty-four children (ASA I-II, aged 3-12 years) undergoing day case ENT surgery were premedicated with midazolam 0.5 mg x kg(-1). Following atropine 10 microg x kg(-1), remifentanil infusion 0.5 microg x kg(-1) x min(-1) was started. After 60 s, anesthesia was induced with propofol 2.5 mg x kg(-1). Immediately after a bolus dose of propofol, the children received rocuronium doses of 0.15 mg x kg(-1) (group I, n = 22) or 0.3 mg x kg(-1) (group II, n = 22) in a randomized manner, after which an infusion of propofol 6 mg x kg(-1) h(-1) was added to the infusion of remifentanil 0.5 microg x kg(-1) min(-1) for maintenance of anesthesia. Intubating conditions were evaluated 90 s after rocuronium administration applying the Copenhagen Scoring System which included components of laryngoscopy, vocal cord movement and reaction to intubation. Hemodynamic values were recorded at predetermined time intervals. RESULTS: Excellent, good and poor intubation conditions were 18.2, 40.9 and 40.9% in group I and 40.9, 54.5 and 4.5% in group II. Clinically acceptable intubating conditions (excellent and good) were significantly higher in group II (95.5%) than in group I (59.1%) (P = 0.004). Mean values of heart rate and blood pressure did not differ significantly between groups. No children required any intervention for hemodynamic instability and/or muscle rigidity. CONCLUSIONS: The results suggest that 0.3 mg x kg(-1) of rocuronium may be a better low dose than 0.15 mg x kg(-1) of rocuronium for clinically acceptable intubating conditions in pediatric ambulatory surgery during remifentanil-propofol-based anesthesia at the doses used in the study. PMID- 15283822 TI - Buprenorphine blocks withdrawal in morphine-dependent rat pups. AB - BACKGROUND: Infants placed on extracorporeal membrane oxygenation (ECMO) or mechanical ventilation often need continuous morphine infusions for pain relief and sedation. The resulting physical dependence requires an additional 2-3-week hospital stay to taper the morphine to avoid withdrawal. Buprenorphine effectively blocks abstinence in dependent adults, and in infants it could accelerate or eliminate the tapering schedule, thereby enabling earlier hospital dismissals. METHODS: Morphine-dependent infant rats were used in this study to determine the effectiveness of buprenorphine in blocking abstinence. Postnatal day-14 (P14) rats were implanted with osmotic minipumps that delivered saline (1 microl x h(-1)) or morphine (2 mg x kg(-1) h(-1)) for 72 h. The minipumps were then removed to allow the rats to undergo spontaneous morphine withdrawal. RESULTS: The withdrawal period lasted approximately 72 h out of a 96-h observation period. The following signs were significant during these hours: wet dog shakes, 1-72 h; abdominal stretches, 1-72 h; forepaw tremors, 1-24 h; splayed hind-limbs, 1-72 h; ptosis, 4-72 h; and evoked vocalization, 4 and 8 h. A single 1 mg x kg(-1) buprenorphine dose significantly decreased wet-dog shakes from 1 to 72 h, abdominal stretches from 1 to 48 h, forepaw tremors and splayed hind-limbs 1-8 h, and ptosis and evoked vocalization at 4 and 8 h. Repeated administration of 1 mg x kg(-1) buprenorphine before pump removal and at 24, 48 and 72 h resulted in a greater magnitude of blockade of abstinence throughout the 96-h observation period. CONCLUSIONS: Buprenorphine may prove to be a suitable drug for treating opioid withdrawal in human infants. PMID- 15283823 TI - Endoscopic intratracheal carbon dioxide measurements during pediatric flexible bronchoscopy. AB - BACKGROUND: CO2 monitoring is recommended for thoracic telescopic procedures and for spontaneous breathing general anesthesia in children. During flexible bronchoscopy (FB) in children, the various currently available methods of CO2 measurements are limited. The CO2 falls and increases have been reported in FB but it is unknown whether airway lesions predispose to CO2 change. The aim of this study was to describe and validate endoscopic intratracheal CO2 measurements in children undergoing FB under spontaneously breathing GA. METHODS: Endtidal CO2 (P(E)CO2) measurements at the start (Start-CO2) and end (End-CO2) of FB on 100 consecutive children were performed using a newly designed endoscopic intratracheal method. To validate the method blood gas sampling was simultaneously performed in 28 children and results analyzed using the Bland and Altman method, intraclass correlation and 95% range for repeatability. RESULTS: End-CO2 and CO2-change (End-CO2 minus Start-CO2) were significantly different in children with airway lesions (CO2 change: no lesion = 3 mmHg, extrathoracic airway lesion = 4.5, intrathoracic airway lesion = 8, P = 0.038). There was no significant difference in Start-CO2 values among the groups. CO2-change in those aged < or =12 months was similar to those >12 months. Intratracheal CO2 measurements were comparable with arterial blood values in the Bland and Altman plots. The intraclass correlation was 0.69 and 95% range for repeatability was 3.7-4.17 mmHg. CONCLUSIONS: Midtracheal P(E)CO2 provides a useful estimate of P(a)CO2 for monitoring the respiratory status of children undergoing FB. The presence of airway lesions rather than age is associated with significant increased PCO2 rise. PMID- 15283824 TI - The use of recalcified citrated whole blood -- a pragmatic approach for thromboelastography in children. AB - BACKGROUND: Thromboelastography (TEG) is an established way of monitoring the coagulation status of children and adults requiring blood products during surgery. Serial measurements are performed using a nearside machine and blood product prescription may be titrated against changes in TEG. There may also be useful applications when the patient is remote from the TEG machine but these are limited because TEG is usually performed on fresh native whole blood within 6 min of venepuncture. Citrated whole blood can be used for TEG if transport time is more than 6 min. We wished to establish whether TEG parameters for citrated whole blood were comparable with those of native whole blood in healthy children. METHODS: Blood was obtained from 14 healthy children undergoing minor surgical procedures, at the time of intravenous cannula insertion for anaesthesia. Each sample was divided: TEG was performed on part of the sample in its fresh native state at 6 min and second portion of the sample was citrated, kept at room temperature and TEG was performed at 30 min after recalcification. RESULTS: There was a significant difference in TEG parameters (r, k, alpha, MA and LY30) for fresh native whole blood and recalcified citrated whole blood (paired t-test). CONCLUSIONS: The normal range for fresh native whole blood TEG parameters is well established, which is routinely used in practice. There was a significant difference between TEG parameters for fresh native whole blood and citrated whole blood. We recommend that a specific normal range be established for citrated whole blood to enable it to be used in clinical practice. PMID- 15283825 TI - Evaluation of adding preoperative or postoperative rectal paracetamol to caudal bupivacaine for postoperative analgesia in children. AB - BACKGROUND: Our aim was to investigate whether effects of caudal analgesia could be extended by preoperative or postoperative rectal paracetamol administration in children undergoing surgical repair of hypospadias. METHODS: The group consisted of 60 ASA I boys, aged 3-12 years, who were operated for surgical repair of hypospadias. The patients were randomized into three groups: patients in group I received rectal paracetamol (20-25 mg x kg(-1)) just before the operation. Group II received only caudal bupivacaine. Group III patients received rectal paracetamol (20-25 mg x kg(-1)) at the end of the operation. Pain was assessed by Children's Hospital of Eastern Ontario Pain Scale (CHEOPS) and the degree of sedation was evaluated. During the first 24 h, time to the patients' first analgesic requirement and the number of supplementary analgesics needed were recorded. RESULTS: There was no difference between the demographic and haemodynamic data of the three groups. In addition, the duration of surgery and anaesthesia, pain scores and sedation scores of the groups were not significantly different. CONCLUSIONS: Addition of preoperative or postoperative rectal paracetamol in the doses used did not show an effect on the duration and intensity of postoperative analgesia obtained by caudal bupivacaine. PMID- 15283826 TI - Minimum effective dose of dexamethasone after tonsillectomy. AB - BACKGROUND: The minimum effective dose of dexamethasone in conjunction with 50 microg x kg(-1) ondansetron was evaluated in the treatment for vomiting after elective tonsillectomy or adenotonsillectomy. METHODS: A total of 102 healthy children between 2 and 12 years of age participated in this prospective, randomized, double-blind study. A single intravenous (i.v.) dose of dexamethasone (50, 100, 150 microg x kg(-1), maximum dose 8 mg) with ondansetron (50 microg x kg(-1)) was administered just before the end of surgery. Equal volumes of normal saline were given to the control group. General anaesthesia was induced and maintained by inhalation of N2O/O2 and sevoflurane. All other preoperative and postoperative medications (including a supplementary dose of antiemetics if necessary), anaesthesia and surgical techniques were standardized. RESULTS: No significant differences were observed between groups in postoperative vomiting on the day of surgery and the next day, or in the need for postoperative pain medication and supplementary doses of antiemetics (P > 0.05). CONCLUSIONS: These results indicate that surgical technique and anaesthetic management used in this study could be the cause of the lower incidence of nausea and vomiting. Assessment of nausea and vomiting in a prospective study with larger groups of patients may reflect different results. PMID- 15283827 TI - Propofol or propofol--alfentanil anesthesia for painful procedures in the pediatric oncology ward. AB - BACKGROUND: For children with cancer receiving curative treatment, the pain of diagnostic and therapeutic procedures is often worse than that of the disease itself. In order to evaluate if light propofol anesthesia in the pediatric oncology ward (POW) could improve the management of procedure pain and anxiety, a questionnaire was developed. METHODS: After prolonged EMLA application, 65 propofol anesthetics were performed successfully in 28 children during lumbar puncture and/or bone marrow aspiration in the POW, with short recovery time and without major adverse events. The questionnaire was mailed to the parents of the 28 children who were included in the survey. RESULTS: The return of questionnaire compliance was 89% (25 of 28), 12 females and 13 males, mean age was 7 years (range 2-16). Among those who replied, the diagnoses were acute lymphatic leukemia in 21, lymphoma in two and tumor in the other two. In the questionnaire, all parents/patients reported advantages with anesthesia in the POW compared with the operating room. In the list of stated advantages, 88% marked 'familiar nurses and doctors', 84% 'familiar environment', 80% 'closer to own room', 68% 'the child more calm', 72% 'shorter waiting-time', 60% 'faster recovery', 44% 'shorter fasting-time' and 44% 'parents more calm', as benefits. For future procedures requiring anesthesia to reduce pain, discomfort and/or anxiety, 92% of the parents/patients preferred anesthesia in the POW. CONCLUSIONS: If anesthesia is chosen for invasive procedures, this study suggest that propofol anesthesia in the POW is preferred by parents and children. PMID- 15283828 TI - Cuff compliance of pediatric and adult cuffed tracheal tubes: an experimental study. AB - BACKGROUND: Tracheal mucosal damage related to tracheal intubation has been widely described in pediatric and adult patients. High volume-low pressure cuffs (HVLPC) are being advertised as safe to avoid this particularly unpleasant complication. Compliances of these supposed pediatric and adult HVLPC are not mentioned by manufacturers and still remain unknown. METHODS: The compliance of HVLPC was measured in vitro and defined as the straight portion of the pressure volume curve. Cuff pressure was measured after incremental 0.1 ml filling volumes of air for sizes 3.0-8.0 of internal diameter of Rusch and Mallinckrodt tracheal tubes. Compliances were assessed in air and in a rigid tube. The filling volume to achieve a 25-mmHg intracuff pressure was also measured. RESULTS: In air, each 0.1 ml step almost linearly increased cuff pressure by 1 mmHg (size 8.0) to 9 mmHg (size 3). In air, the volume needed to maintain a cuff pressure < 25 mmHg was small for sizes 3-5.5 (0.35-2 ml). The 25 mmHg inflated cuff volume and compliance were decreased within a rigid tube, especially for adult sizes. In a rigid tube simulating a trachea, the compliances of almost every Rusch tracheal tube were statistically higher than those of the Mallinckrodt. CONCLUSION: We conclude that the tested tracheal tube cuffs have low compliance and cannot be defined as high volume-low pressure. PMID- 15283829 TI - Axillary brachial plexus block for treatment of severe forearm ischemia after arterial cannulation in an extremely low birth-weight infant. AB - Severe limb ischemia after arterial catheterization in neonates and premature infants is a well-recognized problem. The usual treatment of ischemic injuries includes removal of the catheter and elevation of the effected limb. If unsuccessful, tissue necrosis and loss may follow. We report the case of a 700 g infant with severe distal forearm ischemia after right radial and ulnar artery catheterization. Immediate removal of the arterial line did not improve ischemia. Thirty-six hours later a brachial plexus block via the axillary approach with 0.5 ml bupivacaine 0.125% was performed resulting in rapid improvement, restricting ischemia eventually to fingers II-V as well as the distal part of the thumb. Brachial plexus blockade and active vasodilatation in tiny neonates after severe local ischemia are discussed. PMID- 15283830 TI - Chediak-Higashi syndrome in the intensive care unit. AB - Chediak-Higashi Syndrome is a rare autosomal recessive disease characterized by recurrent infections, giant cytoplasmic granules and oculocutaneous albinism. We describe the clinical and laboratory findings of a patient with Chediak-Higashi syndrome who was diagnosed and treated in the intensive care unit because of bleeding tendency after surgery. PMID- 15283831 TI - Anesthesia for proteus syndrome. AB - Proteus syndrome is a complex disorder comprising malformations and overgrowth of multiple tissues. The disorder is highly variable affecting tissues in a mosaic pattern. A 2-year-old boy with Proteus syndrome, with epidermal verrucal naevus, hyperplastic lesions of connective tissue, hyperostosis, overgrowth of tubular bones, bilateral inguinal hernia, and juvenile intestinal polyposis was scheduled for vertebral magnetic resonance imaging (MRI) for further evaluation of malignancies. In addition to the pathological findings of this syndrome, potential complications such as difficult intubation, pulmonary hypertension, and pulmonary thromboembolism necessitate a careful preoperative and anesthetic preparation. MRI was performed under general anesthesia. There were no anesthetic complications. There are few previous reports on anesthesia in a patient with Proteus syndrome. PMID- 15283832 TI - Anesthetic management of a patient with myotonic dystrophy for a Nissen fundoplication and gastrostomy. AB - A 16-month-old baby with myotonic dystrophy underwent an open Nissen fundoplication and gastrostomy insertion under general anesthesia with an epidural. Postoperative care was managed on the pediatric intensive care unit for the first 6 h. She was then discharged to the ward, where she continued to make an uncomplicated recovery. Other anesthetic management that has been used in children with myotonic dystrophy is discussed. PMID- 15283833 TI - A rare cause of upper airway obstruction in a 5-year-old girl: a laryngeal web. PMID- 15283834 TI - Anesthetic management of a child with xeroderma pigmentosum. PMID- 15283836 TI - Anesthetic management in a child with Coffin-Siris syndrome. PMID- 15283837 TI - Treacher Collins syndrome with choanal atresia: one way to handle the airway. PMID- 15283838 TI - Anesthetic management of a strabismus patient with phenylketonuria. PMID- 15283840 TI - Duplicate publication. Dexmedetomidine in pediatrics: controlled studies needed. PMID- 15283839 TI - Anesthetic management of a strabismus patient with phenylketonuria. PMID- 15283843 TI - On 'reactivity' versus 'tolerance'. AB - In Burnet's review on 'The impact of ideas on immunology' he considers himself an observer of nature using biochemical and molecular analysis for more detailed understanding, a description that applies also to me. I use three examples- repertoire selection of T cells, rules of immune reactivity versus non-reactivity and immunological memory--to illustrate the difficulties we all have in probing nature's immunological secrets and in critically testing immunologists' ideas. At one end of the spectrum of biological research one may argue everything is possible and therefore all results are correct, if correctly measured. But perhaps it is more important to always ask again and again what is frequent and enhances survival versus what is rare and an exception. At the same time one must keep in mind that special situations and special tricks may well be applied for medical benefits, although they may have little impact on physiology and species survival. I will attempt to use disease in virus-infected mice to obtain some answers to what I consider to be important immunological questions with the hope of improving the ratio of answers that are right for the right experimental reasons versus those that are right for the wrong reasons. Some of these experiments falsify hypotheses, previous experiments and interpretations and therefore are particularly important in correcting misleading concepts. They should help to find out which half of immunological ideas and truths in immunological text books written today are likely to be wrong. Ideas are important in immunology, but are often rather demagogically handled and therefore may cost us very dearly indeed. Evaluating immunity to infections and tumours in vivo should help prevent us from getting lost in immunology. PMID- 15283844 TI - Prostanoids and their receptors that modulate dendritic cell-mediated immunity. AB - Dendritic cells (DC) are essential for the initiation of immune responses by capturing, processing and presenting antigens to T cells. In addition to their important role as professional APC, they are able to produce immunosuppressive and pro-inflammatory prostanoids from arachidonic acid (AA) by the action of cyclooxygenase (COX) enzymes. In an autocrine and paracrine fashion, the secreted lipid mediators subsequently modulate the maturation, cytokine production, Th cell polarizing ability, chemokine receptor expression, migration, and apoptosis of these extremely versatile APC. The biological actions of prostanoids, including their effects on APC-mediated immunity and acute inflammatory responses, are exerted by G protein-coupled receptors on plasma membrane. Some COX metabolites act as anti-inflammatory lipid mediators by binding to nuclear receptors and modulating DC functions. Although the role of cytokines in DC function has been studied extensively, the effects of prostanoids on DC biology have only recently become the focus of investigation. This review summarizes the current knowledge about the role of prostanoids and their receptors in modulating DC function and the subsequent immune responses. PMID- 15283845 TI - Investigation of the immunocompetent cells that bind early pregnancy factor and preliminary studies of the early pregnancy factor target molecule. AB - Early pregnancy factor (EPF) is a secreted protein with immunosuppressive and growth factor properties. It has been shown to suppress the delayed-type hypersensitivity response in mice as well as acute and chronic forms of experimental autoimmune encephalomyelitis in rats and mice, respectively. In previous studies, we have demonstrated that EPF binds to a population of lymphocytes and we hypothesized that it mediates its suppressive effects by binding to CD4+ T cells. In the present study, we isolated monocytes and subpopulations of lymphocytes and labelled them with fluoresceinated EPF in order to determine which populations bind EPF. We demonstrated that EPF binds specifically to CD4+, CD8+, CD14+ (monocytes) and CD56+ NK cells but not to CD19+ B cells. The identity of the molecule(s) on the cell surface that is targeted by EPF is unknown, but as EPF is an extracellular homologue of the intracellular protein chaperonin 10 (Cpn10), we examined the possibility that the EPF receptor is a membrane-associated form of chaperonin 60 (Cpn60), the functional associate of Cpn10 within the cell. The EPF target molecule on lymphocytes was visualized by chemical cross-linking of exogenous iodinated Cpn10 to cells and probed with anti-Cpn60. The effect of anti-Cpn60 on activity in the EPF bioassay, the rosette inhibition test, was also examined. In both instances, no specific interaction of this antibody and the putative receptor was observed. It was concluded that the cell surface molecule targeted by EPF is unlikely to be a homologue of Cpn60. PMID- 15283846 TI - Changes in the immune functions and susceptibility to Listeria monocytogenes infection in mice fed dietary lipids. AB - The direct examination of the effects that fish oil diets (composed of long-chain n-3 polyunsaturated fatty acids) exert on immune system function indicates a reduction of host natural resistance to infectious diseases mainly because of a suppression of immune function generated by the fatty acids contained in this diet. Here, we evaluated the concentration of IL-12, IL-4, prostaglandin E2 and leukotriene B4 in the serum from BALB/c mice receiving four different diets. Each group was fed a diet that differed only in the source of fat: a low-fat diet (2.5% by weight), an olive oil diet (20% by weight), a fish oil diet (20% by weight) or a hydrogenated coconut oil diet (20% by weight). Mice were fed for 4 weeks and then infected with the intracellular pathogen Listeria monocytogenes. An initial reduction in the Th1-type response as a result of a decrease in IL 12p70 secretion, an inefficient action of IL-4 (Th2-type response) and no modification of pro-inflammatory lipid-mediator production could be, at least in part, the key events responsible for the inadequate elimination of L. monocytogenes from the spleens of mice fed a fish oil diet. Furthermore, our results suggest that the type of dietary lipids may affect the circulating concentration of IL-12p70 and IL-4, leading to a modulation in the protective cellular immune response to L. monocytogenes infection. PMID- 15283847 TI - Mycobacterium tuberculosis induces high production of nitric oxide in coordination with production of tumour necrosis factor-alpha in patients with fresh active tuberculosis but not in MDR tuberculosis. AB - Mycobacterium tuberculosis is an intracellular pathogen that readily survives and replicates in human macrophages. Host cells have developed various mycobactericidal and immunoregulatory mechanisms, such as the production of nitric oxide and inflammatory cytokines to control intracellular replication of M. tuberculosis. Inducible nitric oxide synthase (iNOS) is transcriptionally under the control of IFN-gamma and TNF-alpha. IL-12 provides a crucial link between activated mononuclear phagocytes and T cells by regulating the production of IFN-gamma. In this study, we investigated the production of nitric oxide (NO), TNF-alpha and IL-12 by the peripheral blood monocytes (PB Mn) of patients suffering from multidrug-resistant tuberculosis (MDR-TB). The cells were infected with M. tuberculosis and stimulated with IFN-gamma or activated with mycobacterial subcellular components. The results were compared with those from cases of newly diagnosed TB and healthy controls. Nitric oxide production was significantly depressed in PB Mn from MDR-TB patients. Infected monocytes from newly diagnosed TB patients produced significantly higher levels of NO as compared to those from MDR-TB patients or normal controls. The subcellular fraction of M. tuberculosis-like whole cell lysate (WCL), culture filtrate protein (CFP) and lipoarabinomannan (LAM) induced higher concentrations of NO release in PB Mn from newly diagnosed TB patients as compared to those from MDR TB patients. Cell culture supernatant from PB Mn assayed at 48 h after infection or stimulation demonstrated significantly depressed release of TNF-alpha and IL 12 from MDR-TB cases as compared to the fresh cases. We observed a definite correlation between nitric oxide release and TNF-alpha production, irrespective of low or high production in MDR-TB or fresh cases, respectively. The present data suggest that peripheral blood monocytes of MDR-TB patients typically show signs of immunosuppression. Whether such immunodepression is the cause or the effect of MDR-TB merits further investigation. PMID- 15283848 TI - Pseudomonas aeruginosa-induced production of free radicals by IFNgamma plus TNFalpha-activated human endothelial cells: mechanism of host defense or of bacterial pathogenesis? AB - We have previously shown that human umbilical vein endothelial cells (HUVEC) can be activated by IFNgamma plus TNFalpha to kill intracellular (IC) Pseudomonas aeruginosa through production of reactive oxygen intermediate, but the cumulative effects of cytokine activation and bacterial infection on host cells has not been extensively addressed. In this study we investigated the fate of IFNgamma plus TNFalpha-activated HUVEC that have harboured IC bacteria for up to 24 h. At 10 h, the endothelial cell killing of P. aeruginosa isolates exceeded 90%. IC bacteria enhanced the expression of inducible nitric oxide synthase (iNOS) and induced overproduction of NO and superoxide by infected HUVEC. P. aeruginosa IC infection also induced a slight decrease in the cellular level of reduced glutathione (GSH). Overproduction of NO correlated with a marked peroxidation of plasma membrane lipids and decline in HUVEC viability. Treatment of cells with the antioxidant alpha-lipoic acid significantly increased the survival of infected cells. Our data suggest that with the failure of adequate scavenger mechanisms, oxidant radicals overproduced in response to bacterial infection were highly toxic to host cells. Therefore, instead of contributing to defence against infectious agents, the upregulation of free radicals production by endothelial cells in response to cytokine activation would be detrimental to the host. PMID- 15283849 TI - Germline transcription of multiple TCR-Vbeta genes in cloned T-cell lines. AB - The functional significance of germline transcription of T cell receptor (TCR) beta chain variable (V) region genes is under investigation. The accepted model is that transcriptional activation of germline TCR genes is associated with the rearrangement process during T-cell development. By this model, germline expression of a subset of TCR-Vbeta genes might be expected in early T cells which have not yet undergone rearrangement. Germline transcription of TCR-Vbeta genes was analysed using the reverse transcriptase (RT)-PCR in a clonal T-cell precursor line C1-V13D, a clonal pre-B cell line RAW112 and a mature T helper cell line D10.G4.1. Evidence is presented for germline transcription of TCR Vbeta8.2 and TCR-Vbeta2.1 genes in all three cell lines, although expression in RAW112 was very weak. C1-V13D cells expressed very high levels of the whole range of transcripts including Vbeta2.1, Vbeta5.1, Vbeta5.2, Vbeta6.1, Vbeta7.1, Vbeta8.1, Vbeta8.2, Vbeta8.3 and Vbeta13.1. However, D10.G4.1 cells expressed a subset of transcripts with apparently lower levels of expression, including Vbeta2.1, Vbeta5.1, Vbeta5.2, Vbeta6.1, Vbeta8.2 and Vbeta8.3. These results raise questions about the significance and possible function of germline transcripts and/or their encoded products in early lymphoid cells and in T cells at different stages of development. PMID- 15283850 TI - Bioassay detects soluble MAdCAM-1 in body fluids. AB - Mucosal addressin cell adhesion molecule (MAdCAM-1) is a key player in mediating the infiltration of leucocytes into chronically inflamed tissues. Five anti MAdCAM-1 monoclonal antibodies (mAb), designated 17F5, 201F7, 314G8, 377D10 and 355G8, were generated by fusion of P3 x 63Ag8.653 myeloma cells with spleen cells from BALB/c mice immunized with recombinant human MAdCAM-1-Fc. The latter four mAb recognize the ligand-binding first Ig domain, and block T -cell adhesion to MAdCAM-1. The non-blocking mAb 17F5 recognizes the mucin domain. Extensive analysis of a large panel of paraffin-embedded human tissues revealed that the 314G8 mAb detected MAdCAM-1 on venules in the spleen and small intestine. MAdCAM 1 was strongly expressed in the synovium of osteoarthritis patients, predominantly on the endothelial lining of blood vessels, but also within the vessel lumen. An ELISA, based on mAb 377D10 and 355G8, was developed to determine whether soluble MAdCAM-1 was present in body fluids, and to measure the levels present. The assay detected soluble MAdCAM-1 in the serum and urine of healthy donors, at levels similar to those of soluble forms of the related CAM, ICAM-1 and VCAM-1. The anti-MAdCAM-1 antibodies and assay developed here may be useful therapeutically in the treatment of inflammation in humans. Similarly, they may be useful diagnostically to monitor the presence and levels of MAdCAM-1. PMID- 15283851 TI - Double-stranded RNA enhances the expression of galectin-9 in vascular endothelial cells. AB - Treatment of cells with double-stranded RNA (dsRNA) in vitro mimics viral infection and regulates expression of various genes. We addressed the mechanisms of leucocyte traffic across the vascular endothelium induced by dsRNA. The present study focused on the expression of galectin-9, which is one of key molecules in the regulation of the interaction between vascular wall and white blood cells. Human umbilical vein endothelial cells (HUVEC) in culture were treated with polyinosinic-polycytidylic acid (poly IC), and expression of mRNA and protein of galectin-9 was analysed by reverse transcription polymerase-chain reaction (RT-PCR) and Western blotting. Poly IC enhanced the expression of galectin-9 mRNA and protein in concentration- and time-dependent manners. This effect of poly IC was almost completely suppressed by the pretreatment with 2 aminopurine, an inhibitor of dsRNA-dependent kinase. Poly IC treatment of HUVEC also enhanced the adherence of EoL-1 cells to the cells, which was inhibited by co-treatment with lactose. We conclude that poly IC upregulates galectin-9 expression in the vascular endothelium and this may explain part of the mechanism for leucocyte traffic through the vascular wall. PMID- 15283852 TI - Phenotypic and functional changes of circulating monocytes and polymorphonuclear leucocytes from elderly persons. AB - The function and phenotype of monocytes and granulocytes in the elderly is consistently remodelled. Because leucocyte adhesion molecules play important roles in mediating a wide variety of leucocyte functions, age-related changes in their expression on granulocyte and monocyte surfaces could be partially responsible for immune dysfunctions during senescence. Considering the central role of innate immunity in the process of immunosenescence and the involvement of cell adhesion molecules (CAM) in the great majority of leucocyte functions, we studied the expression of CD50 and CD62L adhesion molecules in peripheral blood granulocytes and monocytes from healthy elderly and young subjects. We show here that the percentage of granulocytes and monocytes expressing CD62L is decreased in the elderly, whereas its density expression is unchanged on both cell types. A downregulation of the density expression of CD50 at a per cell level characterizes granulocytes in the elderly, whereas CD50 expression on monocytes from old subjects shows a peculiar attitude: its density expression decreases whereas the number of positive cells is expanded. The downregulation of this receptor on granulocytes from aged people could determine a state of hyperactivation contributing to the proinflammatory status of the elderly, while the lower expression on monocytes could therefore contribute to the impaired antigen presentation in the elderly. On the other hand, the increased number of CD50 positive monocytes in the elderly, despite its decreased density expression at a per cell level, could be interpreted as an attempt to counteract the inability to mount strong immune responses. Both CD50 and CD62L changes in ageing polymorphonuclear (PMN) cells allow recognition as non-self or senescent self to permit macrophages in the liver and spleen to remove them from the circulation. The increased proportion of granulocytes and monocytes lacking CD62L and the downregulation of CD50 intensity expression on both cell types may suggest a state of in vivo activation. Therefore, CD50 and CD62L shedding from the cell surface of activated granulocytes and monocytes could be interpreted as a tentative to counteract the dangerous effects of an excessive chronic inflammation in the elderly. However, the increased proportion of CD62L negative granulocytes in the elderly leads to an impairment in cell adhesion which is the first line of response to acute inflammatory stimuli. This phenomenon likely contributes to the increased susceptibility to acute infections of elderly people. PMID- 15283853 TI - Effect of IL-12 and soluble IL-4 receptor on the herpesvirus infection in human SCID chimeras whose Th2 cells predominate. AB - Th2 cytokines, commonly detected in burn patients, have been shown as inhibitors for the generation of Th1 cells that are essential for the host's resistance against herpes simplex virus type 1 (HSV-1) infection. In this study, the possibility of immunological treatment through the regulation of Th1/Th2 responses was examined in two kinds of human severe combined immunodeficiency (SCID) chimera models reflecting human immune functions. SCID mice injected with a mixture of PBMC from a healthy donor and Th2 cells experimentally generated from the same healthy PBMC (Th2 SCID chimeras) were more susceptible to HSV-1 infection when compared with SCID mice injected with healthy donor PBMC (healthy SCID chimeras). When Th2 SCID chimeras were individually treated with human IL-12 (hIL-12) or human soluble IL-4 receptor (hsIL-4R), hIFN-gamma was not produced in their sera after antihuman CD3 mAb stimulation. However, hIFN-gamma production in sera of Th2 SCID chimeras treated with the combination therapy of hIL-12 and hsIL 4R was recovered at levels observed in healthy SCID chimeras. When Th2 SCID chimeras infected with HSV-1 were treated with saline, hIL-12, hsIL-4R or a combination of hIL-12 and hsIL-4R, 13%, 13%, 25% or 100% of them survived, respectively. Also, Th1 responses (hIFN-gamma production) were demonstrated in Th2 SCID chimeras that became resistant against HSV-1 infection after the combination treatment. These results suggest that individuals whose Th2 cells predominated may be immunologically controlled by the combination treatment between a Th1 response inducer and a Th2 response inhibitor. PMID- 15283854 TI - An arthritogenic monoclonal antibody to type II collagen, CII-C1, impairs cartilage formation by cultured chondrocytes. AB - Antibodies to type II collagen (CII) cause articular damage in collagen-induced arthritis (CIA) in mice as judged by passive transfer to naive animals of mAb to CII. We tested the hypothesis that mAb degrade cartilage structure by reacting with functionally important regions of the collagen molecule by examining the effects of an arthritogenic mAb to CII, CII-C1, on cultured bovine chondrocytes at high density, at days 7 and 14. The effects were compared of CII-C1, an isotype-matched control mAb, or medium alone, on chondrocyte proliferation and viability, cell morphology, matrix structure by light and electron microscopy, and matrix synthesis by metabolic labelling with 3H-proline for collagen or 35SO4 for proteoglycans. Chondrocytes in culture remained viable, proliferated, and produced an extracellular matrix in which CII was the major collagen. The addition of CII-C1, but not a control mAb, increased the synthesis of CII and proteoglycan, and caused disorganization of the extracellular matrix and thin collagen fibrils ultrastructurally. Moreover, using a cell-free assay, CII-C1 inhibited the normal self-assembly of collagen fibrils from CII in solution. The finding that the mAb to CII, CII-C1 has striking degradative effects in vitro on cartilage synthesis suggests that antibodies to collagen perpetuate the chronic phase of CIA and that, in mice at least, such antibodies are an important component of pathogenesis. PMID- 15283856 TI - Evaluation of immunotherapy to reverse sequestration in the treatment of severe Plasmodium falciparum malaria. AB - Sequestration and the attachment of Plasmodium falciparum malaria-infected RBC to venous endothelial cells involves parasite-encoded ligands interacting with up to nine host receptors. Antisequestration immunotherapy as an adjunct to quinine did not alter the dynamics of parasite clearance or prove beneficial for the patient. Estimated concentrations of antibody likely to reverse adherence in patients were based on the concentrations of parasite ligands, host receptors and patient equivalents derived from in vitro observations. Calculations presented here indicate that concentrations in excess of a fivefold increase in antibody concentrations used in the immunotherapy trial and equivalent to doubling normal peripheral blood antibody concentrations are anticipated for the successful reversal of sequestration to occur. It is suggested that immunotherapy aimed at either parasite ligands or host receptors to reverse sequestration in the treatment of severe malaria infections is unlikely to be successful given the complexity and number of receptors and ligands and the calculated concentrations of antibodies required. PMID- 15283855 TI - The CD11/CD18 (beta2) integrins modulate neutrophil caspase activation and survival following TNF-alpha or endotoxin induced transendothelial migration. AB - Neutrophils (PMN) are short-lived cells but their survival is often prolonged in inflammation. The beta2 (CD11/CD18) integrins are involved in PMN migration into inflammation but their role in PMN survival is not well understood. We investigated the role of beta2 integrins in PMN caspase activation, a key enzyme cascade in apoptosis. After 20 h, caspase activation (Western blotting) was markedly decreased in PMN cultured on fibrinogen, a ligand for Mac-1 (CD11b/CD18), but not on fibronectin or albumin. In the presence of TNF-alpha or endotoxin (LPS), blockade of CD18 (beta2 chain) with mAb markedly increased caspase activation in PMN on fibrinogen. PMN which migrated through endothelium in vitro in response to TNF-alpha, LPS, IL-1alpha, IL-8 or C5a contained 58% fewer active caspase positive PMN after 20 h than non-migrated PMN remaining on the endothelium. When beta2 (CD18) integrin or lymphocyte function antigen (LFA) 1 (CD11a) plus Mac1 (CD11b) were blocked by mAb (intact or Fab'), the proportion of migrated PMN (but not of non-migrated PMN) with active caspases was significantly increased (2-4-fold) and this was associated with accelerated PMN apoptosis and death. Thus, engagement of ligands on extracellular matrix and endothelium by the beta2 integrins Mac-1 and LFA-1 plays a role in delaying apoptosis in PMN recruited in response to LPS and TNF-alpha. Inhibition of beta2 integrin function may not only inhibit PMN infiltration, but also accelerate PMN clearance from inflamed tissue. PMID- 15283860 TI - Phylogenetic relationships of typical antbirds (Thamnophilidae) and test of incongruence based on Bayes factors. AB - BACKGROUND: The typical antbirds (Thamnophilidae) form a monophyletic and diverse family of suboscine passerines that inhabit neotropical forests. However, the phylogenetic relationships within this assemblage are poorly understood. Herein, we present a hypothesis of the generic relationships of this group based on Bayesian inference analyses of two nuclear introns and the mitochondrial cytochrome b gene. The level of phylogenetic congruence between the individual genes has been investigated utilizing Bayes factors. We also explore how changes in the substitution models affected the observed incongruence between partitions of our data set. RESULTS: The phylogenetic analysis supports both novel relationships, as well as traditional groupings. Among the more interesting novel relationship suggested is that the Terenura antwrens, the wing-banded antbird (Myrmornis torquata), the spot-winged antshrike (Pygiptila stellaris) and the russet antshrike (Thamnistes anabatinus) are sisters to all other typical antbirds. The remaining genera fall into two major clades. The first includes antshrikes, antvireos and the Herpsilochmus antwrens, while the second clade consists of most antwren genera, the Myrmeciza antbirds, the "professional" ant following antbirds, and allied species. Our results also support previously suggested polyphyly of Myrmotherula antwrens and Myrmeciza antbirds. The tests of phylogenetic incongruence, using Bayes factors, clearly suggests that allowing the gene partitions to have separate topology parameters clearly increased the model likelihood. However, changing a component of the nucleotide substitution model had much higher impact on the model likelihood. CONCLUSIONS: The phylogenetic results are in broad agreement with traditional classification of the typical antbirds, but some relationships are unexpected based on external morphology. In these cases their true affinities may have been obscured by convergent evolution and morphological adaptations to new habitats or food sources, and genera like Myrmeciza antbirds and the Myrmotherula antwrens obviously need taxonomic revisions. Although, Bayes factors seem promising for evaluating the relative contribution of components to an evolutionary model, the results suggests that even if strong evidence for a model allowing separate topology parameters is found, this might not mean strong evidence for separate gene phylogenies, as long as vital components of the substitution model are still missing. PMID- 15283861 TI - Improving the scaling normalization for high-density oligonucleotide GeneChip expression microarrays. AB - BACKGROUND: Normalization is an important step for microarray data analysis to minimize biological and technical variations. Choosing a suitable approach can be critical. The default method in GeneChip expression microarray uses a constant factor, the scaling factor (SF), for every gene on an array. The SF is obtained from a trimmed average signal of the array after excluding the 2% of the probe sets with the highest and the lowest values. RESULTS: Among the 76 U34A GeneChip experiments, the total signals on each array showed 25.8% variations in terms of the coefficient of variation, although all microarrays were hybridized with the same amount of biotin-labeled cRNA. The 2% of the probe sets with the highest signals that were normally excluded from SF calculation accounted for 34% to 54% of the total signals (40.7% +/- 4.4%, mean +/- sd). In comparison with normalization factors obtained from the median signal or from the mean of the log transformed signal, SF showed the greatest variation. The normalization factors obtained from log transformed signals showed least variation. CONCLUSIONS: Eliminating 40% of the signal data during SF calculation failed to show any benefit. Normalization factors obtained with log transformed signals performed the best. Thus, it is suggested to use the mean of the logarithm transformed data for normalization, rather than the arithmetic mean of signals in GeneChip gene expression microarrays. PMID- 15283859 TI - The Retinome - defining a reference transcriptome of the adult mammalian retina/retinal pigment epithelium. AB - BACKGROUND: The mammalian retina is a valuable model system to study neuronal biology in health and disease. To obtain insight into intrinsic processes of the retina, great efforts are directed towards the identification and characterization of transcripts with functional relevance to this tissue. RESULTS: With the goal to assemble a first genome-wide reference transcriptome of the adult mammalian retina, referred to as the retinome, we have extracted 13,037 non-redundant annotated genes from nearly 500,000 published datasets on redundant retina/retinal pigment epithelium (RPE) transcripts. The data were generated from 27 independent studies employing a wide range of molecular and biocomputational approaches. Comparison to known retina-/RPE-specific pathways and established retinal gene networks suggest that the reference retinome may represent up to 90% of the retinal transcripts. We show that the distribution of retinal genes along the chromosomes is not random but exhibits a higher order organization closely following the previously observed clustering of genes with increased expression. CONCLUSION: The genome wide retinome map offers a rational basis for selecting suggestive candidate genes for hereditary as well as complex retinal diseases facilitating elaborate studies into normal and pathological pathways. To make this unique resource freely available we have built a database providing a query interface to the reference retinome 1. PMID- 15283862 TI - Cancer immunotherapy: avoiding the road to perdition. AB - The hypothesis that human cancers express antigens that can be specifically targeted by cell mediated immunity has become a scientifically justifiable rationale for the design and clinical testing of novel tumor-associated antigens (TAA). Although a number of TAA have been recognized and it has been suggested that they could be useful in the immunological treatment of cancer, the complexity of human beings leads us to reflect on the need to establish new criteria for validating their real applicability. Herein, we show a system level based approach that includes morphological and molecular techniques, which is specifically required to improve the capacity to produce desired results and to allow cancer immunotherapy to re-emerge from the mist in which it is currently shrouded. PMID- 15283863 TI - A randomised controlled trial to measure the effect of chest pain unit care upon anxiety, depression, and health-related quality of life [ISRCTN85078221]. AB - BACKGROUND: The chest pain unit (CPU) has been developed to provide a rapid and accurate diagnostic assessment for patients attending hospital with acute, undifferentiated chest pain. We aimed to measure the effect of CPU assessment upon psychological symptoms and health-related quality of life. METHODS: We undertook a single-centre, cluster-randomised controlled trial. Days (N = 442) were randomised in equal numbers to CPU or routine care. Patients with acute chest pain, undiagnosed by clinical assessment, ECG and chest radiograph, were recruited and followed up with self-completed questionnaires (SF-36 and HADS) at two days and one month after hospital attendance. RESULTS: Patients receiving CPU assessment had significantly higher scores on the physical functioning (difference 5.1 points; 95% CI 1.1 to 9.0), vitality (4.6; 1.3 to 8.0), and general health (5.7; 2.3 to 9.2) dimensions of the SF-36 at two days, and significantly higher scores on all except the emotional role dimension at one month. They also had significantly lower depression scores on the HADS depression scale at two days (0.93; 0.34 to 1.51) and one month (1.0; 0.36 to 1.66). However, initially lower anxiety scores at two days (0.89; 0.21 to 1.56) were not maintained at one month (0.48; -0.26 to 1.23). CPU assessment was associated with reduced prevalence (OR 0.71; 95% CI 0.52 to 0.97) and severity (6.5 mm on 100 m visual analogue scale; 95% CI 2.2 to 10.8) of chest pain at one month, but no significant difference in the proportion of patients taking time off work (OR 0.82; 95% CI 0.54 to 1.04). CONCLUSION: CPU assessment is associated with improvements in nearly all dimensions of quality of life and with reduced symptoms of depression. PMID- 15283864 TI - Emerging resistance among bacterial pathogens in the intensive care unit--a European and North American Surveillance study (2000-2002). AB - BACKGROUND: Globally ICUs are encountering emergence and spread of antibiotic resistant pathogens and for some pathogens there are few therapeutic options available. METHODS: Antibiotic in vitro susceptibility data of predominant ICU pathogens during 2000-2 were analyzed using data from The Surveillance Network (TSN) Databases in Europe (France, Germany and Italy), Canada, and the United States (US). RESULTS: Oxacillin resistance rates among Staphylococcus aureus isolates ranged from 19.7% to 59.4%. Penicillin resistance rates among Streptococcus pneumoniae varied from 2.0% in Germany to as high as 20.2% in the US; however, ceftriaxone resistance rates were comparably lower, ranging from 0% in Germany to 3.4% in Italy. Vancomycin resistance rates among Enterococcus faecalis were < or = 4.5%; however, among Enterococcus faecium vancomycin resistance rates were more frequent ranging from 0.8% in France to 76.3% in the United States. Putative rates of extended-spectrum beta-lactamase (ESBL) production among Enterobacteriaceae were low, <6% among Escherichia coli in the five countries studied. Ceftriaxone resistance rates were generally lower than or similar to piperacillin-tazobactam for most of the Enterobacteriaceae species examined. Fluoroquinolone resistance rates were generally higher for E. coli (6.5% - 13.9%), Proteus mirabilis (0-34.7%), and Morganella morganii (1.6-20.7%) than other Enterobacteriaceae spp (1.5-21.3%). P. aeruginosa demonstrated marked variation in beta-lactam resistance rates among countries. Imipenem was the most active compound tested against Acinetobacter spp., based on resistance rates. CONCLUSION: There was a wide distribution in resistance patterns among the five countries. Compared with other countries, Italy showed the highest resistance rates to all the organisms with the exception of Enterococcus spp., which were highest in the US. This data highlights the differences in resistance encountered in intensive care units in Europe and North America and the need to determine current local resistance patterns by which to guide empiric antimicrobial therapy for intensive care infections. PMID- 15283865 TI - Failed attempts at experimental transplantation and transmission of nocturnally periodic simian Loa from monkey to man. AB - This paper describes unsuccessful attempts to induce a nocturnally-periodic infection with simian Loa in a human volunteer (the author of this paper) by means of 1. Transplanting adult simian Loa worms from a wild drill (Mandrillus leucophaeus) to man; and 2. Infecting the same volunteer by sub-cutaneous inoculation with infective larvae of simian Loa from a laboratory-bred, experimentally infected Chrysops silacea. PMID- 15283866 TI - Molecular dissection of the human antibody response to the structural repeat epitope of Plasmodium falciparum sporozoite from a protected donor. AB - BACKGROUND: The circumsporozoite surface protein is the primary target of human antibodies against Plasmodium falciparum sporozoites, these antibodies are predominantly directed to the major repetitive epitope (Asn-Pro-Asn-Ala)n, (NPNA)n. In individuals immunized by the bites of irradiated Anopheles mosquitoes carrying P. falciparum sporozoites in their salivary glands, the anti-repeat response dominates and is thought by many to play a role in protective immunity. METHODS: The antibody repertoire from a protected individual immunized by the bites of irradiated P. falciparum infected Anopheles stephensi was recapitulated in a phage display library. Following affinity based selection against (NPNA)3 antibody fragments that recognized the PfCSP repeat epitope were rescued. RESULTS: Analysis of selected antibody fragments implied the response was restricted to a single antibody fragment consisting of VH3 and VkappaI families for heavy and light chain respectively with moderate affinity for the ligand. CONCLUSION: The dissection of the protective antibody response against the repeat epitope revealed that the response was apparently restricted to a single VH/VL pairing (PfNPNA-1). The affinity for the ligand was in the microM range. If anti repeat antibodies are involved in the protective immunity elicited by exposure to radiation attenuated P. falciparum sporozoites, then high circulating levels of antibodies against the repeat region may be more important than intrinsic high affinity for protection. The ability to attain and sustain high levels of anti (NPNA)n will be one of the key determinants of efficacy for a vaccine that relies upon anti-PfCSP repeat antibodies as the primary mechanism of protective immunity against P. falciparum. PMID- 15283867 TI - Brain choline concentrations may not be altered in euthymic bipolar disorder patients chronically treated with either lithium or sodium valproate. AB - BACKGROUND: It has been suggested that lithium increases choline concentrations, although previous human studies examining this possibility using 1H magnetic resonance spectroscopy (1H MRS) have had mixed results: some found increases while most found no differences. METHODS: The present study utilized 1H MRS, in a 3 T scanner to examine the effects of both lithium and sodium valproate upon choline concentrations in treated euthymic bipolar patients utilizing two different methodologies. In the first part of the study healthy controls (n = 18) were compared with euthymic Bipolar Disorder patients (Type I and Type II) who were taking either lithium (n = 14) or sodium valproate (n = 11), and temporal lobe choline/creatine (Cho/Cr) ratios were determined. In the second part we examined a separate group of euthymic Bipolar Disorder Type I patients taking sodium valproate (n = 9) and compared these to controls (n = 11). Here we measured the absolute concentrations of choline in both temporal and frontal lobes. RESULTS: The results from the first part of the study showed that bipolar patients chronically treated with both lithium and sodium valproate had significantly reduced temporal lobe Cho/Cr ratios. In contrast, in the second part of the study, there were no effects of sodium valproate on either absolute choline concentrations or on Cho/Cr ratios in either temporal or frontal lobes. CONCLUSIONS: These findings suggest that measuring Cho/Cr ratios may not accurately reflect brain choline concentrations. In addition, the results do not support previous suggestions that either lithium or valproate increases choline concentrations in bipolar patients. PMID- 15283868 TI - Western medical ethics taught to junior medical students can cross cultural and linguistic boundaries. AB - BACKGROUND: Little is known about teaching medical ethics across cultural and linguistic boundaries. This study examined two successive cohorts of first year medical students in a six year undergraduate MBBS program. METHODS: The objective was to investigate whether Arabic speaking students studying medicine in an Arabic country would be able to correctly identify some of the principles of Western medical ethical reasoning. This cohort study was conducted on first year students in a six-year undergraduate program studying medicine in English, their second language at a medical school in the Arabian Gulf. The ethics teaching was based on the four-principle approach (autonomy, beneficence, non-malfeasance and justice) and delivered by a non-Muslim native English speaker with no knowledge of the Arabic language. Although the course was respectful of Arabic culture and tradition, the content excluded an analysis of Islamic medical ethics and focused on Western ethical reasoning. Following two 45-minute interactive seminars, students in groups of 3 or 4 visited a primary health care centre for one morning, sitting in with an attending physician seeing his or her patients in Arabic. Each student submitted a personal report for summative assessment detailing the ethical issues they had observed. RESULTS: All 62 students enrolled in these courses participated. Each student acting independently was able to correctly identify a median number of 4 different medical ethical issues (range 2 9) and correctly identify and label accurately a median of 2 different medical ethical issues (range 2-7) There were no significant correlations between their English language skills or general academic ability and the number or accuracy of ethical issues identified. CONCLUSIONS: This study has demonstrated that these students could identify medical ethical issues based on Western constructs, despite learning in English, their second language, being in the third week of their medical school experience and with minimal instruction. This result was independent of their academic and English language skills suggesting that ethical principles as espoused in the four principal approach may be common to the students' Islamic religious beliefs, allowing them to access complex medical ethical reasoning skills at an early stage in the medical curriculum. PMID- 15283869 TI - The usefulness of the Korean version of modified Mini-Mental State Examination (K mMMSE) for dementia screening in community dwelling elderly people. AB - BACKGROUND: We assessed whether the Korean version of modified Mini-Mental State Examination (K-mMMSE) has improved performance as a screening test for cognitive impairment or dementia in a general population compared with the Korean Mini Mental State Examination (K-MMSE). METHODS: Screening interviews were conducted with people aged 65 and over in Noam-dong, Namwon-city, Jeonbuk province. There were 522 community participants, of whom 235 underwent clinical and neuropsychological examination for diagnosis of dementia and Cognitive Impairment No Dementia (CIND). Sensitivity, specificity and areas under the receiver operating characteristic (ROC) curves for the K-mMMSE and the K-MMSE were the main outcome measures. RESULTS: Cronbach's alpha for the K-mMMSE was 0.91, compared with 0.84 for the K-MMSE. The areas under the ROC curves in identifying all levels of CIND or dementia were 0.91 for the K-mMMSE and 0.89 for the K-MMSE (P < 0.05). For the K-mMMSE, the optimal cut-off score for a diagnosis of CIND was 69/70, which had a sensitivity of 0.86 and a specificity of 0.79, while, for a diagnosis of dementia, the optimal cut-off score of 59/60 had a sensitivity of 0.91 and a specificity of 0.78. The K-mMMSE also had a high test-retest reliability (r = 0.89). CONCLUSION: Our findings indicate that the K-mMMSE is more reliable and valid than the K-MMSE as a cognitive screen in a population based study of dementia. Considering the test characteristics, the K-MMSE and modified version are expected to be optimally used in clinical and epidemiologic fields. PMID- 15283870 TI - Diagnostic accuracy of ultrasonography compared to unenhanced CT for stone and obstruction in patients with renal failure. AB - BACKGROUND: To determine accuracy of ultrasound (US) kidney, ureter and bladder (KUB) compared to un-enhanced helical CT (UHCT) in patients with renal failure in the diagnosis of stone and obstruction. METHODS: This is a case controlled study conducted in the period from June 2000 to July 2003 at a university hospital. All patients had both US and UHCT scan. Patients with serum creatinine >/= 1.8 mg/dl were included in the study. Only direct visualization of stone was considered as confirmatory. In both the studies, UHCT and US, presence of stone and obstruction were noted. The relevant biochemicals, radiological and clinical records of all the patients were analyzed. Data was analyzed using commercially available software. RESULTS: During the period of study 864 patients had UHCT for evaluation of the urinary tract in patients presenting with flank pain. Out of these 34 patients had both UHCT and US done within a span of one day and had serum creatinine of >/=1.8 mg/dl. Mean age was 48 +/-15.8 years and 59% of patients were males. UHCT identified renal stones in 21 (62%), whereas 17 of these were identified on US, with a sensitivity of 81%. Of the four patients with renal stones missed on US, three were identified on plain x-ray; the mean size of stones missed was 6.3 mm. Of the 22 (65%) patients with ureteric stone on UHCT, US could only identify 10; a further 7 were identified on x-ray KUB, giving a sensitivity of 45% (US alone) and 77% (US with x-ray KUB). CONCLUSIONS: US is sensitive and specific for renal stones, 81% and 100% and for hydronephrosis, 93% and 100%, respectively. Its sensitivity to pick ureteric stone (46%) and to identify hydroureter (50%) is low. Addition of x-ray KUB abdomen increases the sensitivity for ureteric stones to 77%. PMID- 15283871 TI - A randomised controlled trial and cost effectiveness study of systematic screening (targeted and total population screening) versus routine practice for the detection of atrial fibrillation in the over 65s: (SAFE) [ISRCTN19633732]. AB - BACKGROUND: Atrial fibrillation (AF) has been recognised as an important independent risk factor for thromboembolic disease, particularly stroke for which it provides a five-fold increase in risk. This study aimed to determine the baseline prevalence and the incidence of AF based on a variety of screening strategies and in doing so to evaluate the incremental cost-effectiveness of different screening strategies, including targeted or whole population screening, compared with routine clinical practice, for detection of AF in people aged 65 and over. The value of clinical assessment and echocardiography as additional methods of risk stratification for thromboembolic disease in patients with AF were also evaluated. METHODS: The study design was a multi-centre randomised controlled trial with a study population of patients aged 65 and over from 50 General Practices in the West Midlands. These purposefully selected general practices were randomly allocated to 25 intervention practices and 25 control practices. GPs and practice nurses within the intervention practices received education on the importance of AF detection and ECG interpretation. Patients in the intervention practices were randomly allocated to systematic (n = 5000) or opportunistic screening (n = 5000). Prospective identification of pre-existing risk factors for AF within the screened population enabled comparison between high risk targeted screening and total population screening. AF detection rates in systematically screened and opportunistically screened populations in the intervention practices were compared to AF detection rate in 5,000 patients in the control practices. PMID- 15283872 TI - X-linked immunodeficiencies. AB - Recent advances in molecular genetics have allowed identification of at least seven genes involved in X-linked immunodeficiencies. This has resulted not only in improved diagnostic possibilities but also in a better understanding of the pathophysiology of these disorders. In some cases, mutations in the same gene have been shown to cause distinct clinical and immunologic phenotypes, demonstrating a strong genotype-phenotype correlation. Identification of the molecular basis of these diseases has permitted creation of disease-specific registries, with a better characterization of the clinical and immunologic features associated with the various forms of X-linked immunodeficiencies. Additionally, gene therapy has been attempted in X-linked severe combined immune deficiency (XSCID), with clear evidence of successful correction of the pathology, and the appearance of severe adverse effects. PMID- 15283873 TI - The genetics of hypogammaglobulinemia. AB - Etiologies for human hypogammaglobulinemias are diverse and include genetic and nongenetic causes. Although recent reviews focus on the complex genetics of common variable immunodeficiency, in this review, we survey different causes of hypogammaglobulinemias and discuss possible mechanisms. PMID- 15283874 TI - Complement-induced impairment of the innate immune system during sepsis. AB - The complement system is an integral part of innate immunity and is chiefly responsible for controlling bacterial infections, especially those involving gram negative organisms. To accomplish this task, serum proteins engage in a series of enzymatic cascades. The cleaved proteins assemble pores on membranous structures, which lead to cell lysis. During this process, powerful inflammatory mediators are produced, including the anaphylatoxins, C5a, C3a, and the membrane attack complex (MAC). Under systemic inflammatory conditions, an overactive complement system may compromise the effectiveness of innate immunity. We review the detrimental effects that are caused by uncontrolled complement activation during sepsis. PMID- 15283875 TI - Virus subversion of protective immunity. AB - The major histocompatibility (MHC) class I antigen presentation pathway plays a pivotal role in immunity to viruses. MHC class I molecules are expressed on the cell surface of all nucleated cells and present peptides derived from intracellular proteins to cytotoxic T lymphocytes (CTLs), which then eliminate virally infected cells. However, many viruses have evolved proteins to inhibit the MHC class I pathway, thus enabling virally infected cells to escape CTL lysis. In this review, we summarize recent findings about viral inhibition of the MHC class I pathway. PMID- 15283878 TI - Methods for aeroallergen sampling. AB - Air sampling provides information about the bioaerosol composition of the atmosphere. Principal methods of volumetric sample collection include impaction, impingement, and filtration. Many instruments have been developed based on these collection methods. The most widely used devices are slit impactors, rotating arm impactors, and sieve impactors. Samples can be analyzed by various methods, with microscopy and culturing the most important approaches; however, immunoassays, molecular methods such as polymerase chain reaction, and other new techniques are becoming more widely used to analyze samples. PMID- 15283879 TI - Biology of tree pollen allergens. AB - More than 25% of the population suffer from type I allergy. Pollens from trees of the Fagales, Oleaceae, and Cupressaceae belong to the most potent and frequent allergen sources. During the past 15 years, the nature of the most important allergens has been identified by molecular biological techniques, and recombinant allergens equivalent to the natural allergens have been produced. These advances provide insight into the biological functions of important allergens and allow the development of novel forms of diagnosis and therapy. In this review, we focus on Fagales allergens to illustrate the impact of recombinant allergens on diagnosis and therapy. We discuss structural similarities as a molecular basis for cross-reactivities and develop diagnostic concepts by using species-specific marker allergens as well as highly cross-reactive allergens. The identification of the allergen recognition profiles of patients with recombinant allergens allows a more precise selection of patients for available forms of allergy treatment. Moreover, we describe novel recombinant allergen-based forms of specific immunotherapy. PMID- 15283880 TI - Biology of weed pollen allergens. AB - Weeds represent a heterogeneous group of plants, usually defined by no commercial or aesthetic value. Important allergenic weeds belong to the plant families Asteraceae, Amaranthaceae, Urticaceae, Euphorbiaceae, and Plantaginaceae. Major allergens from ragweed, mugwort, feverfew, pellitory, goosefoot, Russian thistle, plantain, and Mercurialis pollen have been characterized to varying degrees. Four major families of proteins seem to be the major cause of allergic reactions to weed pollen: the ragweed Amb a 1 family of pectate lyases; the defensin-like Art v 1 family from mugwort, feverfew, and probably also from sunflower; the Ole e 1 like allergens Pla l 1 from plantain and Che a 1 from goosefoot; and the nonspecific lipid transfer proteins Par j 1 and Par j 2 from pellitory. As described for other pollens, weed pollen also contains the panallergens profilin and calcium-binding proteins, which are responsible for extensive cross reactivity among pollen-sensitized patients. PMID- 15283881 TI - Cross-reactivity of pollen allergens. AB - Pollen cross-allergenicity has practical implications on the management of inhalant allergy, in both evaluation and therapy, especially with allergen vaccine immunotherapy. The study of cross-reactivity among pollen allergens has expanded beyond investigation of crude extracts to the characterization and cloning of numerous pollen proteins. In this review, the interrelationships between these pollen allergens in the context of botanical systematics are examined, to provide a framework for cross-reactivity understanding. Recommendations for choices in evaluation and therapy are given. PMID- 15283883 TI - The International Bladder Cancer Bank: proposal for a new study concept. AB - At present, results of marker studies are often inconsistent and sometimes contradictory. Recognized problems include multiple different methods of performing the assays, different subsets of patients and different endpoints, leading to incompatible datasets. Although there has been discussion of establishing general methodological principles and guidelines (analogous to those for clinical trials) for design, conduct, analysis, and reporting of marker studies, these have not been widely implemented. There are no well-recognized prototypes or examples that the urologic researcher can use to model future marker studies. We will discuss our plans to establish a multi-institutional bladder cancer data base and virtual tumor bank as a resource for participating institutions to evaluate the biological and prognostic significance of potential markers for bladder cancer. Samples will be identified and stored at each participating institution and will be available for analysis. A standard, minimal set of patient and pathologic information will be collected. The use of common software, as part of this proposal will facilitate the data transfer of updated patient information to a central database. All contributing centers will have access to summarized information, also to simplify the process of finding collaborating partners. Prospectively collected, consistent datasets with available long-term follow-up, should provide information sooner than with a conventional prospective study. Furthermore, the quality of these data and samples may be superior to that of retrospectively collected data and samples. The proposed International Bladder Cancer Bank of specimens and data will be an effective tool during all phases of marker development. PMID- 15283882 TI - Thunderstorm asthma. AB - Thunderstorms have often been linked to epidemics of asthma, especially during the grass flowering season; however, the precise mechanisms explaining this phenomenon are unknown. Evidence of high respirable allergen loadings in the air associated with specific meteorologic events combined with an analysis of pollen physiology suggests that rupture of airborne pollen can occur. Strong downdrafts and dry, cold outflows distinguish thunderstorm rain from frontal rain. The weather system of a mature thunderstorm likely entrains grass pollen into the cloud base, where pollen rupture would be enhanced, then transports the respirable-sized fragments of pollen debris to ground level where outflows distribute them ahead of the rain. The conditions occurring at the onset of a thunderstorm might expose susceptible people to a rapid increase in concentrations of pollen allergens in the air that can readily deposit in the lower airways and initiate asthmatic reactions. PMID- 15283884 TI - Testicular microlithiasis: a review and its association with testicular cancer. AB - Testicular microlithiasis (TM) is an entity of unknown etiology that results in the formation of intratubular calcifications. It is of concern to the urologist because of its possible association with intratubular germ cell neoplasia and testicular germ cell cancer. Although commonly present in patients with germ cell tumors, there appears to be no definitive association with TM and cancer. Therefore, follow-up at this time should be dictated based on risk factors for developing testis cancer more than on the presence of TM. PMID- 15283885 TI - Management of clinical T1 bladder transitional cell carcinoma by radical cystectomy. AB - High-grade bladder cancer involving the lamina propria is considered superficial disease. This spectrum is generally treated with TUR plus intravesical therapy. However, significant understaging jeopardizes long-term survival and improvements and radical surgery represents a provocative alternative. We evaluated disease free and cancer-specific survival (CSS) in our cohort of patients with high-grade T1 tumors. A total of 318 patients with bladder cancer underwent radical cystectomy between 1990 and 2000 at our institution. Of these, 66 had cT1 tumors with or without Carcinoma in-situ (CIS). Our multidisciplinary bladder cancer database was queried to perform a multivariate analysis on clinical parameters such as: age, race, sex, cystectomy year, intravesical therapy, angiolymphatic invasion and tumor upstage in relation to recurrence and survival. The clinical stage was accurate in 44 of the cases (66%). However, 27% were upstaged by cystectomy and 12% of the cT1 + CIS patients had nodal disease. Patients with cT1 tumors plus CIS had a significantly worse CSS. Those with persistent disease after an initial course of BCG therapy appeared to have worse CSS also. At a median follow up of 4 years, overall cancer-specific mortality was 22%, however, pathologic T1 +/- CIS had 92% CSS at 10 years. Our data suggests that some cT1 bladder cancer tumors have assiduous clinical courses evidenced in staging discrepancies. For high-grade tumors, early cystectomy and orthotopic diversion increases life expectancy significantly and should be carry out early rather than late. PMID- 15283886 TI - The impact of pathology review on treatment recommendations for patients with adenocarcinoma of the prostate. AB - This study was designed to estimate the frequency with which changes in Gleason score because of a genitourinary pathologist's review changed prostate cancer treatment recommendations. The study cohort consisted of 602 patients who presented to a genitourinary oncologist for a second opinion after being diagnosed with prostate cancer based on a needle biopsy at a nonacademic institution from 1989 through 2001. Each of the prostate biopsy specimens was sent for review by a genitourinary pathologist. Applying the rule that low-risk patients would receive monotherapy, and intermediate or high-risk patients would receive combined modality therapy, the frequency with which treatment recommendations were changed by pathology review was calculated. Pathology review by a genitourinary pathologist changed the Gleason score by at least 1 point in 44% of cases. Upgrades were more common than downgrades and accounted for 81% [95% confidence interval: 76-86%] of the changes. Patients' risk category was increased in 10.8% of cases and was decreased in 3.4%. Risk category was changed from low risk to intermediate or high risk in 8.2%, but was changed from intermediate or high risk to low risk in only 0.9%. Genitourinary pathology review would have changed management in approximately 10% of men, mainly in the direction of combined therapy over monotherapy. PMID- 15283887 TI - Continuing trends in pathological stage migration in radical prostatectomy specimens. AB - Prostate-specific antigen (PSA) screening has resulted in a profound clinical stage migration. Extracapsular extension (ECE) presents a poor prognosis after radical prostatectomy (RP). In this study the trends in rate of ECE for cancers detected by PSA screening between 1987, when PSA screening became routine in the United States, and 2001, were examined. The clinical outcome of patients (total 1505; 888 clinical Tlc, 614 clinical T2, and 3 clinical T3) with prostate cancer diagnosed by PSA screening and treated with RP without neoadjuvant hormonal therapy was analyzed. The primary outcome variable was ECE rate with respect to year of treatment for a given tumor stage, preoperative PSA level, biopsy Gleason score, and surgical Gleason score. Logistic regression analysis was used to identify predictors of ECE. Biochemical relapse-free survival (bRFS) by year of treatment was analyzed by Kaplan-Meier Curve. Rate of ECE decreased from 65.8 to 25.2% during the 15-year study duration. Multivariate analysis of clinical tumor stage, age, preoperative serum PSA level, and Gleason score confirmed that year of treatment was an independent predictor of ECE. Six-year bRFS rates (by years of treatment) were 75.1% for 1987 to 1994 and 82.6% for 1995 to 2001 (P-value = 0.0022). PSA screening has resulted in a downward pathological stage migration. These observations demonstrate improved biochemical failure rates in more recently treated patients. PMID- 15283888 TI - The assessment of PTEN tumor suppressor gene in combination with Gleason scoring and serum PSA to evaluate progression of prostate carcinoma. AB - OBJECTIVE: The purpose of the study was to determine if the tumor suppressor gene phosphate and tensin homolog (PTEN) (mutated in multiple advanced cancers 1) in combination with Gleason scoring and serum prostate specific antigen (PSA) could be employed to better predict the progression of prostate carcinoma. MATERIALS AND METHODS: The study group consisted of 43 patients with benign prostate hyperplasia (BPH), 15 with organ confined prostate carcinoma (OCPCa), and 18 with advanced prostate carcinoma (APCa). Prostate tissue samples were obtained from radical prostatectomy, transurethral resection, and TRUS guided trans-rectal needle biopsy and then evaluated for biomarker expression. The clinical stage was assessed according to tumor node metastasis classification and grade according to Gleason system. Serum PSA was measured by conventional techniques and Western blotting analysis was used to determine PTEN expression in the primary tissue. Multivariate analysis was performed to analyze whether these markers could individually predict the progression of prostate carcinoma. RESULTS: APCa patients displayed higher Gleason scores and serum PSA levels. But much lower PTEN expression was detected in prostate of APCa patients compared to patients with BPH or OCPCa. Hormone refractory (HR) and hormone sensitive (HS) APCa cases did not yield any significant differences in terms of Gleason scoring, serum PSA and PTEN expression. PSA levels were significantly higher in patients with OCPCa or APCa compared to patients with BPH. CONCLUSION: Our results suggested that both PTEN and serum PSA appeared to be useful as independent markers to depict the nature of tumor behavior as benign or malign. In addition, PTEN also appeared to be useful as an independent marker to predict the progression of prostate carcinoma. PMID- 15283889 TI - Microarray analysis of differential gene expression in androgen independent prostate cancer using a metastatic human prostate cancer cell line model. AB - Progression to androgen independence (AI) leading to uncontrolled cell growth is the main cause of death in prostate cancer. While almost all patients with metastatic prostate cancer will initially respond to anti-androgen treatments, the majority will fail hormonal treatments in less than 2 yrs. Both genetic and epigenetic alterations in gene expression contribute significantly to the development of AI. To investigate this we have used an in vitro cell line model of AI prostate cancer from which we have identified a number of differentially expressed genes associated with progression to AI in prostate cancer. We used an in vitro cell line model of AI prostate cancer, to study differential gene expression using cDNA microarray analysis and corroborated the microarray results with Ribonuclease Protection Assay (RPA). Approximately 4480 out of 7075 (63.3%) cDNA cloned genes were differentially expressed, of which, 6 genes were differentially expressed by at least fivefold. RPA was used to corroborate the microarray results for the five most highly differentially expressed genes. Using an in vitro cell line model and microarray analysis we have identified a number of candidate genes for further investigation in AI prostate cancer. PMID- 15283891 TI - Clinical proteomics: Applications for prostate cancer biomarker discovery and detection. AB - The science of proteomics comprises much more than simply generating lists of proteins that change in expression as a cause of or consequence of pathophysiology. The goal of proteomics should be to characterize the information flow through the intercellular protein circuitry that communicates with the extracellular microenvironment and then ultimately to the serum/plasma macroenvironment. Serum proteomic pattern diagnostics is a new type of proteomic concept in which patterns of ion signatures generated from high dimensional mass spectrometry data are used as diagnostic classifiers. This recent approach has exciting potential for clinical utility of diagnostic patterns because low molecular weight metabolites, peptides, and protein fragments may have higher accuracy than traditional biomarkers of cancer detection. Intriguingly, we now have discovered that this diagnostic information exists in a bound state, complexed with circulating highly abundant carrier proteins. These diagnostic fragments may one day be harvested by circulating nanoparticles, designed to absorb, enrich, and amplify the repertoire of diagnostic biomarkers generated even at the critical, initial stages of carcinogenesis. PMID- 15283892 TI - Selective capture of prostatic basal cells and secretory epithelial cells for proteomic and genomic analysis. AB - Basal cells play an undefined role in signaling the growth and differentiation of normal secretory epithelial cells in the human prostate. Because basal cells disappear during malignant transformation, we hypothesize that loss of basal cell function may have a permissive role in progression of prostate intraepithelial neoplasia into invasive carcinoma. We describe an immuno-laser capture microdissection approach to selectively capture basal cells. Using surface enhanced laser desorption/ionization time-of-flight mass spectrometry, we identified several protein candidates selectively expressed in microdissected basal cells. We also demonstrate that the RNA derived form this technique is an excellent source for gene-array studies. Thus, we provide evidence that proteomic and microgenomic techniques can be successfully applied to investigate the expression profiles of basal and secretory cells after immuno-capture. PMID- 15283894 TI - The science of early detection. AB - Most clinical judgment and clinical intuition derives from observations made on patients who suffer from a disease or medical condition. However, the target population for cancer screening is healthy people who would not seek out a health professional unless convinced to do so by advertising or public messages. Extrapolation of clinical observations to the target population for screening can be very misleading and even harmful. This is because powerful screening biases and confounding effects--such as selection bias, lead-time bias, length-bias sampling, and overdiagnosis--can mislead even the most astute clinician. This article will discuss those biases, review methods to avoid them, and provide useful resources to the clinician or health scientist. PMID- 15283893 TI - The early detection research network surface-enhanced laser desorption and ionization prostate cancer detection study: A study in biomarker validation in genitourinary oncology. AB - Prostate-specific antigen (PSA) screening has led to a dramatic increase in prostate cancer detection with a concurrent stage migration. Although the test has revolutionized prostate cancer detection by identifying disease that is potentially curable in the majority of men, only 25% of men receiving test results of PSA > 4 ng/ml will have prostate cancer and many men receiving a normal PSA will have disease, including high-grade disease. There is a need for improved biomarkers for detecting prostate cancer. One such method of cancer detection is surface-enhanced laser desorption and ionization (SELDI). The Early Detection Research Network (EDRN) validation study for SELDI for prostate cancer is described. In a three-stage study, the portability and reproducibility of the technique will be determined; the predictive algorithm will be refined in a multi institutional case-control population; followed by ultimate validation in the context of a prospective trial with complete disease ascertainment. The unique aspect of the EDRN SELDI validation study is the novel use of two groups of cancer cases: those cases with higher-risk disease (Gleason > or = 7) and those cases with lower-risk disease (Gleason < or = 6). This study will allow the first evaluation of a predictive algorithm that includes prognosis in disease screening. The EDRN SELDI prostate cancer biomarker validation study is a rigorous evaluation of a new detection method for prostate cancer. The methodologies used for this evaluation will prove useful for guiding future biomarker studies in this challenging disease. PMID- 15283895 TI - Incorporating predictions of individual patient risk in clinical trials. AB - A risk prediction model is a statistical technique that gives a predicted probability of a certain event for an individual patient. Prediction models outperform the traditional risk classification systems that work by assigning patients into risk groups based on the presence or absence of particular risk factors, such as stage of disease. As such, risk prediction models have a number of important possible uses in clinical trials. For Phase II studies, prediction models can help adjust comparisons with historical control groups for differences in case mix. For Phase III studies, prediction models can ensure that accrued patients are at sufficiently high risk. This improves statistical power and avoids unethical inclusion of low-risk patients. We also propose that prediction models could potentially be used for applying the results of Phase III trials to individual patients. Clinical decisions could be informed by individualized estimates of treatment benefit, rather than by average treatment effects. PMID- 15283896 TI - Prostate cancer screening: clinical applications and challenges. AB - Prostate cancer is a serious illness warranting appropriate screening measures. However, current screening tests that include prostate-specific antigen and digital rectal examination must be proven to save lives to be considered truly legitimate and appropriate public health tools. Even though these tests are associated with the diagnosis of disease, important questions remain as to how well these tests identify all disease and whether screening leads to interventions that save lives. Prostate cancer is undoubtedly a killer, yet there appear to be large numbers of detectable prostate cancers that are of little threat to life. Some men with this grade of cancer receive curative treatment, even though their disease does not require treatment. Some studies suggest that more than three of four men with screen-detected localized disease may not need treatment. One of the great challenges of cancer communications is how to convey the hope of prostate cancer screening while adequately acknowledging the boundaries of our knowledge. The current absence of an appropriate informed consent tool points to the necessity to develop an easy to understand informed consent that allows men to evaluate screening decisions with a clear understanding of what is known, what is not known, and what is believed to be true about prostate cancer screening. PMID- 15283897 TI - The prostate, lung, colon, and ovarian (PLCO) cancer screening trial: Status and promise. AB - The Prostate, Lung, Cancer, and Ovarian Cancer Screening Trial is a randomized, multicenter, study evaluating whether screening for prostate, lung, colon, and ovarian cancer will reduce cancer-specific mortality. More than 155,000 participants have been identified and will be monitored through 2015. A biorepository of screened individuals and control participants has been created. Together, the data derived from the intervention and control arms and material in the biorepository provide a valuable resource to allow identification and testing of novel biomarkers for human disease. PMID- 15283898 TI - The person-years saved model and other methodologies for assessing the population impact of cancer-prevention strategies. AB - The results of the Prostate Cancer Prevention Trial spurred debate because finasteride was found to reduce the period prevalence of prostate cancer by about 25% while also increasing the rate of high-grade prostate cancers. Assessing how finasteride would impact mortality at the population level is key to evaluating the public health implications of these results. Any model for evaluating the impact of chemoprevention at the population level will involve methods for assigning weights to competing outcomes of the chemoprevention. The person-years saved model, which uses survival to weigh outcomes, assesses the impact on population mortality and has particular strengths. The person-years saved model shows that more than 300,000 person-years would be saved during a period of 10 years with the widespread use of finasteride, assuming no change in the rate of high-grade prostate cancers. The rate of high-grade prostate cancers in the population, about 20% in any given year, would have to nearly triple to 60% for the net positive impact of finasteride to be zero. The person-years saved model shows that the administration of finasteride is likely to result in a net positive impact of finasteride on population mortality, even with an increase in the rate of high-grade prostate cancers. Future models may use other outcomes to assess population impact, including economic, quality-of-life, or various combinations of outcomes. PMID- 15283899 TI - Delivering prostate cancer prevention messages to the public: how the National Cancer Institute (NCI) effectively spread the word about the Prostate Cancer Prevention Trial (PCPT) results. AB - The Prostate Cancer Prevention Trial was the first clinical trial to show that a direct intervention (5 mg of finasteride daily for 7 years) could reduce a man's risk of developing prostate cancer. Initial results also suggested that men taking finasteride had an increased risk of developing what appeared to be higher grade disease (Gleason score 7-10). The National Cancer Institute has a congressional mandate to communicate health information to the public and has established methods to reach the public directly and to reach information intermediaries in the media, professional societies, and advocacy groups. The groundbreaking yet complicated results of the Prostate Cancer Prevention Trial were widely disseminated by National Cancer Institute using the social marketing and public-relations strategies and tactics detailed here. PMID- 15283910 TI - Investigating protein folding, misfolding and nonnative states: experimental and theoretical methods. PMID- 15283911 TI - Experimental investigation of protein folding and misfolding. AB - Newly synthesised proteins need to fold, often to intricate and close-packed structures, in order to function. The underlying mechanism by which this complex process takes place both in vitro and in vivo is now becoming understood, at least in general terms, as a result of the application of a wide range of biophysical and computational methods used in combination with the techniques of biochemistry and protein engineering. It is increasingly apparent, however, that folding is not only crucial for generating biological activity, but that it is also coupled to a wide range of processes within the cell, ranging from the trafficking of proteins to specific organelles to the regulation of cell growth and differentiation. Not surprisingly, therefore, the failure of proteins to fold appropriately, or to remain correctly folded, is associated with a large number of cellular malfunctions that give rise to disease. Misfolding, and its consequences such as aggregation, can be investigated by extending the types of techniques used to study the normal folding process. Application of these techniques is enabling the development of a unified description of the interconversion and regulation of the different conformational states available to proteins in living systems. Such a description proves a generic basis for understanding the fundamental links between protein misfolding and its associated clinical disorders, such as Alzheimer's disease and Type II diabetes, and for exploring novel therapeutic strategies directed at their prevention and treatment on a rational basis. PMID- 15283912 TI - Rapid mixing methods for exploring the kinetics of protein folding. AB - Information on the time-dependence of molecular species is critical for elucidating reaction mechanisms in chemistry and biology. Rapid flow experiments involving turbulent mixing of two or more solutions continue to be the main source of kinetic information on protein folding and other biochemical processes, such as ligand binding and enzymatic reactions. Recent advances in mixer design and detection methods have opened a new window for exploring conformational changes in proteins on the microsecond time scale. These developments have been especially important for exploring early stages of protein folding. PMID- 15283913 TI - Methods to study protein folding by stopped-flow FT-IR. AB - Stopped-flow mixing coupled with time-resolved Fourier transform infrared (FT-IR) spectroscopy represents a new experimental approach to explore protein folding events, which has become possible only recently with the development of appropriate techniques. Here, we discuss experimental apparatus that are capable of initiating and monitoring protein folding processes on the millisecond to minute timescale. The strongest point of the FT-IR approach as a structure specific probe is that a complete spectrum is available for each time point of measurement. In this way, several spectral windows are accessible simultaneously for the observation of the unfolding or the formation of different secondary structure elements and also events that can be attributed to changes in tertiary structure. One specific advantage of the infrared technique is the ability to monitor directly the kinetics of processes involving beta-sheet structures, which is exceptionally difficult to do with other techniques. PMID- 15283914 TI - The analysis of protein folding kinetic data produced in protein engineering experiments. AB - Over the past decade, the "protein engineering method" has been used to investigate the folding pathways of more than 20 different proteins. This method involves measuring the folding and unfolding rates of mutant proteins with single amino acid substitutions spread across the sequence. Comparison of folding rates of the mutant proteins to that of the wild-type protein allows the calculation of the phi value, which can be used to evaluate the stabilizing contribution of an amino acid side chain to the structure of the folding transition state. Here, we review the methodology for analysing data collected in protein engineering folding kinetics studies. We discuss the calculation of folding rates and kinetic m values, the estimation of errors in folding kinetics experiments, phi value calculation including potential pitfalls of the analysis, Bronsted plots, detecting Hammond behaviour, and the analysis of curved chevron plots. PMID- 15283915 TI - Hydrogen exchange methods to study protein folding. AB - The measurement of amino acid-resolved hydrogen exchange (HX) has provided the most detailed information so far available on the structure and properties of protein folding intermediates. Direct HX measurements can define the structure of tenuous molten globule forms that are generally inaccessible to the usual crystallographic and NMR methods (C. Redfield review in this issue). HX pulse labeling methods can specify the structure, stability and kinetics of folding intermediates that exist for less than 1 s during kinetic folding. Native state HX methods can detect and characterize folding intermediates that exist as infinitesimally populated high energy excited state forms under native conditions. The results obtained in these ways suggest principles that appear to explain the properties of partially folded intermediates and how they are organized into folding pathways. The application of these methods is detailed here. PMID- 15283916 TI - Protein folding studied by real-time NMR spectroscopy. AB - Real-time NMR spectroscopy developed to a generally applicable method to follow protein folding reactions. It combines the access to high resolution data with kinetic experiments allowing very detailed insights into the development of the protein structure during different steps of folding. The present review concentrates mainly on the progress of real-time NMR during the last 5 years. Starting from simple 1D experiments, mainly changes of the chemical shifts and line widths of the resonances have been used to analyze the different states populated during the folding reactions. Today, we have a broad spectrum of 1D, 2D, and even 3D NMR methods focusing on different characteristics of the folding polypeptide chains. More than 20 proteins have been investigated so far by these time-resolved experiments and the main results and conclusions are discussed in this report. Real-time NMR provides comprehensive contributions for joining experiment and theory within the 'new view' of protein folding. PMID- 15283917 TI - Photo-CIDNP NMR methods for studying protein folding. AB - Chemically induced dynamic nuclear polarization (CIDNP) is a nuclear magnetic resonance phenomenon that can be used to probe the solvent-accessibility of tryptophan, tyrosine, and histidine residues in proteins by means of laser induced photochemical reactions, resulting in significant enhancement of NMR signals. CIDNP offers good sensitivity as a surface probe of protein structure and is particularly powerful in time-resolved NMR measurements. Real-time, rapid injection protein refolding experiments permit the observation of changes in the accessibility of specific residues during the folding process. CIDNP pulse labeling gives information on the accessibility of residues in partially structured proteins (e.g., molten globule states) whose NMR spectra are broad and poorly resolved. Heteronuclear two-dimensional (15)N-(1)H CIDNP techniques allow identification of surface-accessible residues with improved resolution and sensitivity. These methods offer residue-specific structural and kinetic information on transient folding intermediates and other partially folded states of proteins that are not readily available from more routine NMR techniques. PMID- 15283918 TI - Methods to study protein dynamics and folding by mass spectrometry. AB - It is clear that merely knowing the structure of a protein alone is not sufficient to fully understand its behavior: knowledge also of the dynamic events that occur within proteins is vital to elucidate their function and folding. In recent years, mass spectrometry has come to the forefront as a powerful biophysical method, which can shed light both on the structure and dynamics of proteins. Hydrogen exchange monitored by mass spectrometry is a highly sensitive marker of the backbone dynamics in solution that, combined with gas phase dissociation techniques, can provide a high resolution tool to locate the dynamic regions of a protein. Additionally, charge state distributions in electrospray mass spectra yield insight into the nature and population of alternate structural states present at equilibrium. In this paper, we describe several applications of these methodologies to probe the dynamic events key to the structure, folding, and biological functions of proteins. PMID- 15283919 TI - Atomic force microscopy: mechanical unfolding of proteins. AB - Mechanical unfolding of proteins using atomic force microscopy is becoming a routine biophysical technique. Mechanistic investigations in this rapidly evolving field are beginning to resolve the factors that contribute to the behaviour of biological macromolecules under force. Here we describe the force mode apparatus, the experimental set-up, tractable systems of study, and the analysis of the resultant force data. Finally we summarise some of the recent achievements and limitations of this technique. PMID- 15283920 TI - Methods for molecular dynamics simulations of protein folding/unfolding in solution. AB - All atom molecular dynamics simulations have become a standard method for mapping equilibrium protein dynamics and non-equilibrium events like folding and unfolding. Here, we present detailed methods for performing such simulations. Generic protocols for minimization, solvation, simulation, and analysis derived from previous studies are also presented. As a measure of validation, our water model is compared with experiment. An example of current applications of these methods, simulations of the ultrafast folding protein Engrailed Homeodomain are presented including the experimental evidence used to verify their results. Ultrafast folders are an invaluable tool for studying protein behavior as folding and unfolding events measured by experiment occur on timescales accessible with the high-resolution molecular dynamics methods we describe. Finally, to demonstrate the prospect of these methods for folding proteins, a temperature quench simulation of a thermal unfolding intermediate of the Engrailed Homeodomain is described. PMID- 15283921 TI - Using nuclear magnetic resonance spectroscopy to study molten globule states of proteins. AB - Nuclear magnetic resonance (NMR) spectroscopy is a powerful technique for the study of the structure, dynamics, and folding of proteins in solution. It is particularly powerful when applied to dynamic or flexible systems, such as partially folded molten globule states of proteins, which are not usually amenable to X-ray crystallography. In this article, NMR methods suitable for the detailed characterisation of molten globule states are described. The specific method used to study the molten globule is determined by the quality of the NMR spectrum obtained. Molten globules are characterised by significant levels of secondary structure. Site-specific hydrogen-deuterium exchange experiments can be used to identify residues located in regions of secondary structure in the molten globule. If spectra characterised by sharp peaks are observed for the molten globule then information about secondary structure can be obtained by analysis of (1)H(alpha), (13)C(alpha), (13)C(beta), and (13)CO chemical shifts; this can be supplemented by (15)N relaxation studies. For molten globules characterised by extremely broad peaks (15)N-edited NMR experiments carried out in increasing concentrations of denaturants can be used to study the relative stabilities of different regions of structure. Examples of the application of these methods to the study of the low pH molten globule states of alpha-lactalbumin and apomyoglobin are presented. PMID- 15283922 TI - High-pressure NMR spectroscopy for characterizing folding intermediates and denatured states of proteins. AB - Extensive structural studies using high-pressure NMR spectroscopy have recently been carried out on proteins, which potentially contribute to our understanding of the mechanisms of protein folding. Pressure shifts the conformational equilibrium from higher to lower volume conformers. If the pressure is varied, starting from the folded native structure, in many cases we observe intermediate conformers before the onset of total unfolding. This enables the investigation of details of the structure and thermodynamic characteristics of various intermediate conformers of proteins under equilibrium conditions. We can also examine pressure effects on the structure and stability of some typical denatured states such as helical denatured, molten globule, and unfolded states. The high pressure NMR method can also be used to investigate association/dissociation equilibria of oligomeric or aggregated proteins. Beside direct observation of kinetic intermediates upon pressure jump, NMR structural investigations of equilibrium conformers under pressure provide information about the structures of kinetic intermediates during folding/unfolding reactions. PMID- 15283923 TI - Computational methods for generating models of denatured and partially folded proteins. AB - Partially folded and denatured proteins can give important insights into protein folding, misfolding, and aggregation. Such non-native states of proteins are however very difficult to characterise in detail as they are dynamic, heterogeneous systems comprising of ensembles of interconverting conformers. This article describes methods that produce models for non-native proteins in atomic detail. A variety of molecular dynamics based protocols are discussed together with some recent procedures that include restraints from experimental data. These models provide an important framework for interpreting experimental data from studies of non-native states using nuclear magnetic resonance spectroscopy, fluorescence, circular dichroism, and small angle scattering techniques. PMID- 15283924 TI - Techniques to study amyloid fibril formation in vitro. AB - Amyloid fibrils are ordered aggregates of peptides or proteins that are fibrillar in structure and contribute to the complications of many diseases (e.g., type 2 diabetes mellitus, Alzheimer's disease, and primary systemic amyloidosis). These fibrils can also be prepared in vitro and there are three criteria that define a protein aggregate as an amyloid fibril: green birefringence upon staining with Congo Red, fibrillar morphology, and beta-sheet secondary structure. The purpose of this review is to describe the techniques used to study amyloid fibril formation in vitro, address common errors in the collection and interpretation of data, and open a discussion for a critical review of the criteria currently used to classify a protein aggregate as an amyloid fibril. PMID- 15283925 TI - Hormone replacement therapy in cancer survivors. AB - OBJECTIVE: Thousands of women are treated each year for cancer; many of these are already in menopause, while other younger patients will go into early menopause due to surgery, or chemotherapy, or the need for radiotherapy to the pelvic region. In most cases the oncologist and the gynaecologist would advise these women against the use of HRT. The purpose of this paper is to review biological and clinical evidences in favour and against HRT use in the different tumours and to propose an algorithm that can help choosing the treatment for the single woman. METHODS: We performed a systematic literature review through April 2002 concerning: (1) biological basis of hormonal modulation of tumour growth; (2) epidemiological data on the impact of HRT on different cancers risk in healthy women; (3) safety of HRT use in cancer survivors; (4) alternatives to HRT. RESULTS: With the exception of meningioma, breast and endometrial cancer, there is no biological evidence that HRT may increase recurrence risk. In women with previous breast and endometrial cancer HRT is potentially hazardous on a biological basis, even if published data do not show any worsening of prognosis. CONCLUSIONS: Even if a cautious approach to hormonal-dependent neoplasias is fully comprehensible and the available alternative treatment should be taken into greater consideration, the reticence to prescribe HRT in women previously treated for other non hormone-related tumours has neither a biological nor a clinical basis. An algorithm based on present knowledge is proposed. PMID- 15283926 TI - Skin contamination by oestradiol gel--a remarkable source of error in plasma oestradiol measurements during percutaneous hormone replacement therapy. AB - OBJECTIVES: To study the consequence of skin contamination by oestradiol gel on circulating plasma oestradiol levels. METHODS: We studied ten healthy, hysterectomized postmenopausal women who had used percutaneous oestradiol gel for at least 2 years. After wash-out period percutaneous dose of 1.5 mg 17beta oestradiol was administered once a day in the evening. The gel was applied with a bare or gloved hand to an arm or thigh according to the schedule. Blood samples for assay of plasma oestradiol concentrations were collected from both cubital veins 12 h after gel administration, at baseline and every time after using the gel, for 2 weeks. RESULTS: Plasma oestradiol concentrations were significantly higher in the gel-contaminated samples: in the cubital vein of the gel-applying arm and in the cubital vein of the forearm on which the gel had been spread. CONCLUSIONS: Skin contamination by topical 17beta-oestradiol can distort plasma oestradiol measurements by giving much higher oestradiol concentrations than in reality there are in the systemic circulation. This has an important meaning when tailoring individual oestrogen therapy. PMID- 15283927 TI - Comparison of the effects of tibolone and estrogen replacement therapy on echocardiographic basic cardiac functions in post-menopausal women: a randomized placebo controlled study. AB - OBJECTIVES: This study is designed to investigate and compare the effects of synthetic steroid tibolone and HRT on systolic and diastolic heart functions in post-menopausal women. METHODS: This prospective, randomized placebo controlled double blind study was conducted in a university clinic. Fifty-eight non-smoking, otherwise healthy post-menopausal women who did not receive any kind of HRT at least for 3 years within the onset of menopause were included in the study. The patients were randomly allocated to either 2.5 mg per day tibolone (OD, n = 18), daily combined 0.625 mg of conjugated estrogens 2.5 mg-1 of medroxy progesterone acetate pill (EP, n = 20) or a vitamin pill (n= 20) in a double blinded fashion. Their basic systolic and diastolic functions were investigated with HP Sonos-1000 echocardiography using standard positions and windows before and 6 months after the initiation of HRT. RESULTS: Mean age, weight, length of post-menopausal period, heart rate, systolic and diastolic pressures were similar between the groups. At the initiation of the study all groups had similar echocardiographic measurements. However, at the end of 6 months, left ventricular end-systolic and diastolic volumes were decreased significantly compared to pretreatment and placebo in both EP and OD treated groups. (55.5 +/- 18.4 and 53.7 +/- 19.1.8 ml; 109.9 +/-19.9 and 110.7 +/- 20.8 ml versus 74.5 +/- 14.9 and 142.7 +/- 19.1 ml, respectively; P < 0.05). Improvement in diastolic functions was significant in EP/OD groups compared to pre-treatment period and the placebo groups (E/A 1.34 +/ 0.1 and 1.38 +/- 0.1 versus 1.18 +/-.09, deceleration time 204 +/- 11.1 and 202.8 +/- 27.1 ms versus 237.6 +/- 26.9 ms, respectively). Besides increase in left ventricular mass adjusted for height, decrease in left ventricular relative wall thickness, and systemic vascular resistance were significant in EP and OD treated groups than placebo and the pre-treatment measurements. Although improved in both OD and EP groups, the changes in systolic and diastolic functions were significantly higher in the OD treated group. Based on our preliminary results, we may conclude that both EP and OD regimens may improve cardiac performance and age related dysfunctions. CONCLUSION: The present results may further support that both OD and EP exert many direct effects on cardiovascular system other than metabolic changes regarding lipoproteins. The greater improvement in the OD group may be explained by its weak androgenic activity which is consistent with the in vitro data that androgens are potent relaxing agents on coronary arteries and restores cardiac myosin isoenzyme and ATPase patterns which mandates further clinical studies. PMID- 15283928 TI - Effect of HRT on hormone responses to resistance exercise in post-menopausal women. AB - PURPOSE: The purpose of this study was to evaluate the effect of hormone replacement therapy (HRT) on the acute and chronic hormonal responses to resistance exercise in post-menopausal women. METHODS: Thirty-two post-menopausal women were recruited for this study; 16 who were currently using HRT and 16 who were not using HRT. Subjects in both the HRT and NHRT groups were randomly assigned to either a resistance training group (N = 16; 8 HRT and 8 NHRT) or a control group (N = 16; 8 HRT and 8 NHRT). The training group completed a supervised resistance training program three times a week for 12 weeks. To evaluate changes in hormone levels, resting blood samples were drawn at weeks 0, 4, and 13 of the program. In addition, at weeks 0 and 13, post-exercise blood samples were drawn in order to examine the hormone response to an acute bout of resistance exercise. Samples were analyzed for serum growth hormone (GH), insulin like growth factor-1 (IGF-1), testosterone, estradiol, dehydroepiandrosterone (DHEA), and cortisol. RESULTS: There were no significant changes in resting hormone levels between weeks 0, 4, and 13 of the training program. There was a significant week-by-group interaction for DHEA (P < 0.05 ) and cortisol (P < 0.05 ) with the NHRT-training group having a greater post-exercise increase in DHEA and cortisol after training. Overall, the post-exercise GH levels were significantly greater than pre-exercise (P < 0.05 ) or recovery levels (P < 0.01). There were no significant differences between HRT and NHRT groups in the acute hormone response to exercise. CONCLUSION: These results indicate that HRT will not have an effect on the acute or chronic hormone response to a recreational resistance training program in post-menopausal women. PMID- 15283930 TI - Prevalence of disability and associated social and health-related factors among the elderly in Spain: a population-based study. AB - OBJECTIVES: To estimate the prevalence of disability and its association with morbidity and other social and health-related factors among Spain's non institutionalized elderly population. METHODS: Cross-sectional survey, by home based personal interview, covering a sample of 4000 subjects representative of the non-institutionalized Spanish population aged 60 years and over. The relationship between disability and social and health-related study variables was studied using logistic regression. RESULTS: A total of 72.9% of subjects reported some type of disability: 59.1% in agility, 51.6% in mobility, 40.1% in instrumental activities and 19.1% in activities of daily living. After adjusting for all relevant variables, disability showed to be significantly more frequent for: female gender (OR = 1.83; 1.53-2.19); more advanced age (OR = 4.54; 3.27 6.32); low/no educational level (OR = 2.01; 1.67-2.42); deteriorated cognitive status (OR = 1.67; 1.24-2.23); at least two chronic diseases (OR = 2.54; 2.01 3.20); poor perceived health status (OR = 3.02; 2.48-3.69); little physical activity (OR = 2.57; 1.94-3.42); and greater use of hospital care (OR = 1.34; 1.10-1.64). CONCLUSIONS: Prevalence of disability among Spain's non institutionalized elderly population is very high. This might be explained by a greater number of chronic diseases, a higher percentage of subjects with low educational level and a higher proportion of community-dwelling elderly persons than in Anglo-Saxon countries. PMID- 15283929 TI - Benefits of soy germ isoflavones in postmenopausal women with contraindication for conventional hormone replacement therapy. AB - OBJECTIVE: To evaluate the effects of isoflavones on vasomotor symptoms and blood lipids in postmenopausal women with contraindication for conventional hormone replacement therapy (HRT). METHODS: This prospective, double-blind and placebo controlled study included 50 postmenopausal women randomly divided into two groups: 25 women on soy germ isoflavones (60 mg per day, capsules) and 25 women on placebo. Inclusion criteria included: non-vegetarian, non-asian women whose last menstruation dated at least 12 months prior to the beginning of the study, with FSH > 40 mIU/ml, hot flushes and contraindication for HRT, not using tamoxifen or antibiotic and no disease of the gastrointestinal tract. For 6 months, the Kupperman menopausal index (KMI), the vaginal cytological maturation value (MV) and both hormonal and lipid profiles were assessed. The t-test and analysis of variance (ANOVA) were employed to compare the two groups. RESULTS: In both groups, a decreased KI rate was observed. However, isoflavone was significantly superior to placebo in reducing hot flushes (44% versus 10%, respectively) (P < 0.05). After 6 months, the isoflavone group showed increased estradiol levels with unchanged FSH, LH, and vaginal cytology, and a reduction of 11.8% in LDL and an increase of 27.3% in HDL (P < 0.05 ). In the placebo group, just a reduction in MV was observed after 6 months (P < 0.05 ). CONCLUSIONS: Soy germ isoflavone exerted favorable effects on vasomotor symptoms and lipid profile, showing itself to be an interesting alternative therapy for the postmenopausal women with contraindication for conventional HRT. PMID- 15283931 TI - Anticardiolipin antibodies during hormone replacement therapy in healthy postmenopausal women. AB - OBJECTIVE: The aim of the study was to investigate IgG and IgM anticardiolipin (aCL) antibodies in the course of hormone replacement therapy (HRT). SUBJECTS AND METHODS: Thirty clinically healthy postmenopausal women with no history of previous thrombotic events or autoimmune disease were divided in two groups: control group (n = 12, mean age 52.9 +/- 4.5 years, and 6.2 +/- 3.6 years duration of amenorrhea) and a second group (n = 18, mean age 53.6 +/- 3.5 years, and 5.7+/- 4.5 years of amenorrhea) who were allocated to HRT, containing 2 mg 17 beta estradiol plus 1mg norethisterone acetate daily for 6 months. ACL antibodies of IgG and IgM isotype were assessed using ELISA and the Kupperman menopausal index (KI) was calculated at baseline and after 3 and 6 months of treatment. RESULTS: HRT had a beneficial effect on climacteric symptoms, evaluated by KI (baseline versus 3rd month and 3rd month versus 6th month, P < 0.001 ). The KI did not change in the control group. The values of IgG at the three studied periods did not change significantly and were 14.1 +/- 4.2, 13.1 +/- 4.9 and 13.4 +/- 3.7 in the HRT group and 12.7 +/- 3.1, 13.7 +/-1.8 and 13.1 +/- 3.8 GPL, respectively, in the control group. IgM aCL antibodies increased during HRT and were as follows: 7.7 +/- 4.8 at baseline, 12.9 +/- 5.6 at 3rd month and 9.3 +/- 3.2 MPL at 6th month. In the control group, IgM were 8.0 +/- 2.8; 7.9 +/- 2.3 and 7.1 +/- 2.3 MPL, respectively. The differences between the two groups were significant at the third and the 6th month (P < 0.01 and P < 0.05 ). CONCLUSION: These data suggest that HRT is associated with elevation of IgM ACA in healthy postmenopausal women. As IgG aACA are considered more pathogenic, it seems unlikely that the early prothrombogenic effects of HRT can be assigned to ACA. PMID- 15283932 TI - Hormone replacement therapy and health behavior in postmenopausal polio survivors. AB - OBJECTIVES: Little is known about menopause and hormone replacement therapy (HRT) use in women with disabilities. The objectives of this study were to explore the health behaviors, health outcomes, and efficacy of HRT in a group of postmenopausal polio survivors and to compare selected outcomes to nationally representative cohorts. METHODS: One hundred and thirty-one postmenopausal polio survivors completed self-report surveys on health behaviors, HRT use, functional status, and psychosocial well-being. During a physical examination, fasting cholesterol and body mass index (BMI) were collected. Independent sample t-tests and Chi-square analysis were used to compare HRT users and non-users on health behaviors and health outcomes; logistic regression was used to predict HRT use. RESULTS: Prevalence of HRT use was 58%. Only BMI predicted HRT use (OR = 0.30, CI: 0.11-0.81). HRT users had better high density lipoprotein (HDL), low density lipoprotein, total cholesterol/HDL ratios, lower BMIs, were more confident when communicating with their physicians, more likely to discuss menopause with their physician, and experienced greater overall stress. HRT was not associated with health behavior, health-related quality of life, mood, or life satisfaction. Compared to non-disabled women, more of these women had higher total cholesterol, obesity, more sleeping problems, and were less likely to vigorously exercise or smoke. CONCLUSIONS: HRT did not confer substantial benefits in these postmenopausal polio survivors to warrant them using HRT at a higher rate than their non-disabled peers. Comparisons to their non-disabled peers suggested they may be at higher risk for adverse health problems associated with postmenopause. PMID- 15283933 TI - Sexual function, menopause and hormone replacement therapy (HRT). AB - OBJECTIVE: To assess the prevalence of female sexual dysfunction in premenopausal and postmenopausal women with and without hormone replacement therapy (HRT). To determine the relationship between menopause and sexual activity, and the impact of HRT on sexual function. METHODS: A cross-sectional analysis of 231 Colombian born women, aged 40-62 years. Sexual function was measured by self-questionnaire. The analysis was performed by using the chi2-test and multivariate regression analysis. The sexual function was divided in five domains: desire, arousal, lubrication, orgasm and pain; in addition, questioning about sexual satisfaction was included in the research. RESULTS: In the study 38.1% of women showed sexual dysfunction in the desire, and 25% in the arousal, these two being the most affected domains. Even though menopause marginally decreases all stages of sexual function, this association was statistically significant only for the lubrication and pain domains. HRT improves sexual function in the orgasm, lubrication and pain domains in a statistically significant manner. The level of sexual satisfaction was better on postmenopausal women with HRT than ones without HRT. Age negatively influences almost all sexual function domains in a significant manner. CONCLUSIONS: Menopause affects in a negative manner some domains of female sexual function. HRT improves some factors of the sexual function during menopause but it not improves desire and arousal which were the most affected domains. There is a negative association between age and female sexual response in middle-aged women. PMID- 15283934 TI - Clinical usefulness of endometrial screening by ultrasound in asymptomatic postmenopausal women. AB - OBJECTIVE: The aim of the present study was to evaluate the clinical usefulness of routine use of endometrial ultrasound in asymptomatic, bleeding-free postmenopausal women. METHODS: We retrospectively reviewed the data of 850 postmenopausal women subjected to hysteroscopy, focusing our attention on those cases (148) with an ultrasound indication of endometrial thickening. RESULTS: In 850 postmenopausal women, we identified 27 (3.2%) endometrial adenocarcinomas. In these subjects, the indication for office hysteroscopy was abnormal uterine bleeding in 24 (24/27; 88.9%) cases; pathological pap smear with abnormal endometrial cells in 2 (2/27; 7.4%) cases and thickened endometrium upon transvaginal ultrasound (tvUS) only in one (1/27; 3.7%) patient. On the other hand, 148 hysteroscopies were performed on the basis of the tvUS indication in otherwise asymptomatic (bleeding free) postmenopausal women; only 1(0.7%) of these presented an adenocarcinoma. CONCLUSION: Our findings show that the use of tvUS as a screening tool for endometrial pathology in asymptomatic postmenopausal women generates 93.2% false positive results, so that most of these women undergo this second level invasive procedure uselessly. Our data suggest that, in asymptomatic postmenopausal women, endometrial ultrasound evaluation is not worthwhile as a screening tool, such as it is considered in common clinical practice. The present results call for a larger prospective trial to further elucidate this controversial issue. PMID- 15283935 TI - Influence of psycho-social factors on climacteric symptoms. AB - BACKGROUND: It has been suggested that psycho-social factors may be crucial in the development of climacteric symptoms. MATERIAL AND METHODS: In order to evaluate the effect of psycho-social and biological factors on menopausal symptoms, Greene (climacterical symptoms), Cooper (psychosomatic symptoms of stress), Smilkstein (family dysfunction), Duke-UNC (social support) and Israel (life events) tests were passed to 300 Chilean women between 40 and 59 years of age. Data were evaluated with ANOVA, chi2 and logistic regression using the Epi info package. RESULTS: Perimenopausal women had a significant increase in stress and climacteric symptoms; however comparing with pre and postmenopausal women, tests for life events, family dysfunction or social support did not show any differences. A history of premenstrual syndrome was the main risk predictor f or climacteric symptoms (OR: 3.6, IC: 1.5-8.5; P < 0.03 ), followed by perimenopausal state (OR: 2.9, IC: 1.4-6.0; P < 0.001 ) and negative life events (OR: 2.3, IC: 1.0-5.3; P < 0.05 ). The psycho-social factors were predictors for anxiety and depression; on the other hand, perimenopausal state was a risk factor for somatic and vasomotor symptoms. During premenopause, women with regular cycles and vasomotor symptoms have more psychological symptoms and stress. CONCLUSION: Climacteric symptoms that appear in the perimenopause are more intense in those women who have a biological predisposition such as premenstrual syndrome and are modulated by psycho-social factors. PMID- 15283936 TI - A research on the level of urine neopterin to see if it may provide a vital clue for a provisional diagnosis of breast cancer in menopausal women. AB - OBJECTIVES: To search the relation between the level of neopterin in urine and breast cancer developed in menopause. METHODS: In this study, urine samples were collected from randomly selected menopausal and post-menopausal women who attended hospital, and from a control group. The participants were classified into two representative sample groups and a control group; 1st group: 30 menopausal and post-menopausal women whose ages varied from 45 to 80 and who suffered from breast cancer but had no viral infections, chronic inflammatory diseases and smoking habit; 2nd group: 30 menopausal and post-menopausal women aged between 48 and 63 with no complaint of any type of tumors, viral infections or chronic inflammatory diseases and with no smoking habit; 3rd group: A control group with 20 women aged between 20 and 28, who did not take part in sexual activities yet, and had no complaint of any type of tumors, viral infections or chronic inflammatory diseases and had no smoking habit. Urinalysis was carried out for each sample in order to measure the level of neopterin. RESULTS: The mathematical results of neopterin levels for the groups showed that group I was significantly higher than group II and III (P = 0.0001 ); group II was significantly higher than group III (P = 0.003 ). CONCLUSIONS: If the results of this study were not confounded by another factor, then can we deduce that this relationship can be used as a risk factor that should warrent further investigation of breast cancer during the care and treatment of menopausal women. PMID- 15283937 TI - Does hormone therapy increase allergic reactions and upper gastrointestinal problems? Results from a population-based study of Swedish woman. The women's health in the Lund area (WHILA) study. AB - OBJECTIVES: To delineate the use of various drugs particularly pertaining to allergy and upper gastrointestinal problems in relation to hormone status in middle aged women. METHODS: An analysis from a population-based study on women born between 1935 and 1945 and lived in the Lund area southern Sweden. Of 10,766 women, 6,917 provided complete data sets; in turn 5,673 were assessed for the use of medication in this study. Among the cohort, 9% of women were premenopausal (PM), 54% were postmenopausal without hormone replacement therapy (PM0) and 37% were current hormone replacement therapy users (PMT). RESULTS: There were 7 (1.3%) women in PM, 11 (0.4%) in PM0 and 21 (1.0%) in PMT group who used loratadine regularly. There was a significant difference between the PM and PM0 groups and also between the PM0 and PMT groups in the use of loratadine (P < 0.05 ). Among 21 loratadine users in PMT group 4 (19%) used transdermal patches and 17 (81%) used oral HRT. The result for omeprazole use was as follows: 4 (0.8%) of PM group, 39 (1.3%) of PM0 group and 42 (2.0%) of PMT group. The use of omeprazole was significantly higher in the PMT group than in the PM (P = 0.05 ) and PM0 group (P < 0.05 ). There was no relation between the use of omeprazole and smoking or alcohol consumption. CONCLUSIONS: Use of hormone replacement therapy seems to be related to a higher frequency of omeprazole and loratadine use, which implies that hormone replacement therapy, may be associated with more upper gastrointestinal symptoms as well as allergy. PMID- 15283938 TI - Tibolone, oral or transdermal hormone replacement and colour Doppler analysis: a prospective, randomised pilot study. AB - OBJECTIVE: To compare the plasma thromboxane, the plasma viscosity and the Doppler flow modifications induced by tibolone and by oral or transdermal continuous combined hormone replacement therapy. METHODS: Forty-two post menopausal patients underwent either on: oral daily treatment with tibolone (2.5 mg) (Group I; n= 14); or continuous oral administration of 0.625 mg conjugated equine estrogens + medroxyprogesterone 5 mg per day (Group II; n = 14 ); or continuous estradiol transdermal supplementation, at a dose of 50 microg per day, + medroxyprogesterone 5 mg per day (Group III; n = 14 ). The duration of the study was 6 months and the patients were submitted to transvaginal ultrasonographic evaluation of pelvic organs; Doppler analysis of the uterine, internal carotid and ophthalmic arteries; thromboxane and plasma viscosity assays in basal condition, and at 1, 3 and 6 months from the beginning of the study. RESULTS: Although the endometrial thickness increased significantly, there were no cases in which it exceeded the normal range (< or = 5 mm). In all the three groups, the pulsatility index of the uterine, internal carotid and ophthalmic arteries significantly decreased during the therapy showing a reduced impedance since the first month of treatment. Similar variations were observed for the peak systolic blood flow velocity of the internal carotid and ophthalmic arteries. Hormone replacement therapy and tibolone induced a deep, significant and rapid decrease in plasma thromboxane and plasma viscosity levels. CONCLUSIONS: Hormone replacement therapy and tibolone seem to have beneficial effects on vascular and hemorrheological parameters. PMID- 15283939 TI - A comparison of the central effects of different progestins used in hormone replacement therapy. AB - OBJECTIVE: To evaluate the central effect exerted by different progestins used for hormone replacement therapy. METHODS: Randomised, placebo-controlled study. One hundred-twenty postmenopausal women on continuous hormonal replacement therapy with transdermal estradiol (50 microg per day) associated, for 10 days every 28 days, with four different progestins: dydrogesterone (DYD; 10 mg per day; n = 20), medroxyprogesterone acetete (MPA; 10 mg per day; n = 20), nomegestrol acetate (NMG; 5 mg per day; n = 20) or norethisterone acetate (NETA; 10 mg per day; n = 20). Other 40 women, 10 for each treatment group, were used as controls and were monitored for a single cycle of 28 days during the administration of transdermal estradiol plus placebo. Morning basal body temperature (BBT) was monitored for 28 days. Anxiety, by the state-trait anxiety inventory, and depression, by the self-evaluation depression scale of Zung, were evaluated just prior to and in the last 2 days of the 10-day progestins adjunct. RESULTS: All progestins except DYD increased (P < 0.0001) BBT by 0.3-0.5 degrees C. Anxiety was decreased by DYD (- 2.3 + 1.1; P < 0.01) and MPA (- 1.5 + 0.5; P < 0.01), but not by NMG or NETA. Depression did not significantly increase during progestins and actually decreased during MPA (- 3.0 + 0.7; P < 0.01). Only the effect of DYD on anxiety and that of MPA on depression were significant versus the control group (P < 0.05 ). CONCLUSIONS: Different progestins exert different central effects. DYD has the peculiarity of not increasing BBT and of decreasing anxiety, which is also decreased by MPA. Depression is not negatively affected by the tested progestins and it may be ameliorated by MPA. The present data may help to individualise the progestin choice of hormone replacement therapy. PMID- 15283940 TI - Rat tail skin temperature regulation by estrogen, phytoestrogens and tamoxifen. AB - OBJECTIVE: Develop a rat model for the evaluation of estrogenic agents on estrogen deficiency-induced changes in thermoregulation. METHODS: OVX rats are impaired in thermoregulation which manifests itself as an elevation in basal tail skin temperature (TST) and are less able to respond to temperature changes than intact rats. RESULTS: Administration of estrogen subcutaneously to estrogen depleted rats either as depot formulation, biodegradable pellets, or daily injections, suppressed the increased TST. OVX rats maintained on a diet devoid of phytoestrogens had a higher TST by several degrees than OVX rats fed normal chow, offering greater ability to test estrogenic agents on thermoregulation. Depletion of estrogen in intact rats via chronic administration of leuprolide acetate, a GnRH agonist, also increased TST, which was in turn suppressed by estrogen. In intact rats, tamoxifen exhibited estrogen antagonistic activity elevating TST, while in OVX rats, tamoxifen acted as an agonist by suppressing TST. CONCLUSION: OVX rats kept on a diet devoid of phytoestrogens are a sensitive model for estrogen-dependent thermoregulation. PMID- 15283941 TI - Carotid and uterine vascular resistance in short-term hormone replacement therapy postmenopausal users. AB - OBJECTIVE: To compare the short-term effects of oral hormone replacement therapy (HRT) and placebo on carotid and uterine vascular impedance. METHODS: 80 postmenopausal women selected from the outpatient clinic of the Hospital Leonor Mendes de Barros in Sao Paulo, Brazil, were randomized to oral HRT (estradiol 2 mg/norethisterone acetate 1 mg-Kliogest(r)) or placebo. Carotid and uterine arteries pulsatility indices (PIs) were assessed by color Doppler at baseline, after 4 and 12 weeks of treatment. Seventy-six women completed the trial, 38 in each group. RESULTS: The carotid PI did not decrease significantly in either group. In the uterine arteries, the drop in PI was steeper and greater for HRT women. Drops occurred despite the supposed counteracting effect of norethisterone acetate. In placebo group, there was no significant difference between 4 and 12 weeks of treatment compared with the baseline. The results did not change when analyzed in a real treatment approach. CONCLUSION: Oral continuous HRT are effective at 12 weeks in reducing impedance to flow in uterine, but not in carotid circulation. These results suggest that the effects of HRT vary by vascular site, and do not have a detectable short-term vascular effect in the carotid area. PMID- 15283942 TI - Imaginary pregnancy 10 years after abortion and sterilization in a menopausal woman: a case report. PMID- 15283943 TI - Alcohol dependence and the alcohol Stroop paradigm: evidence and issues. AB - AIMS: Firstly to test alcohol abusers attentional bias towards alcohol-related stimuli. Secondly, to shed light onto other factors that may influence the alcohol Stroop effect by considering variables including mood status in the analyses. Finally, to examine severity of dependence on Stroop performance. DESIGN: Repeated measures with alcohol versus control group as the between participant factors and within participant factors were the reaction times to different types of stimuli. Standard multiple regression was used to determine predictors of Stroop performance. A repeated measures design was used with severity of dependence as the between participant factors and Stroop reaction times as the within participant factors. SETTING: South and East London, UK. PARTICIPANTS: Sixty-four alcoholics in treatment and 64 community controls from general practice participated in the study. MEASUREMENTS: Alcohol dependence severity was measured using the SADQ, mood was measured with the POMS-SF and a computerised emotional Stroop task was employed to measure attentional bias. FINDINGS: Regardless of demographic factors and mood status, alcoholics responded significantly slower to alcohol-related than neutral words when compared to controls. When severity of alcohol dependence was used as between participant factors, no significant differences were found with Stroop performance between high and low alcohol severity groups. CONCLUSIONS: Alcohol-related stimuli are distracting to heavy users of alcohol, independent of demographic, mood and dependence status. Findings offer insight into the development of alcohol dependence and the issues that surround the alcohol Stroop paradigm. PMID- 15283944 TI - A pilot trial of topiramate for the treatment of cocaine dependence. AB - BACKGROUND: Both GABAergic and glutamatergic neurons appear to be important modulators of the brain reward system and medications that affect GABA and glutamatergic neurotransmission may reduce the rewarding properties of cocaine and reduce cocaine craving. Topiramate, an anticonvulsant, raises cerebral GABA levels, facilitates GABAergic neurotransmission and inhibits glutametergic activity at AMPA/kainite receptors. Thus, it may be useful for treating cocaine dependence. METHODS: The efficacy of topiramate for cocaine dependence was tested in a 13-week, double-blind, placebo-controlled pilot trial (n = 40). Topiramate was titrated gradually over 8 weeks to a dose of 200 mg daily. The primary outcome measure was cocaine abstinence verified by twice weekly urine benzoylecgonine tests (UBT). RESULTS: Eighty-two percent of subjects completed the trial. Analysis of the UBT using a GEE model showed that after week 8, when the dose titration was completed, topiramate-treated subjects were more likely to be abstinent from cocaine compared to placebo-treated subjects (Z = 2.67, P = 0.01). Topiramate-treated subjects were also more likely to attain 3 weeks of continuous abstinence from cocaine (chi2 = 3.9, d.f. = 1, P = 0.05). CONCLUSION: Topiramate may be effective for the treatment of cocaine dependence. PMID- 15283945 TI - Characteristics of men with substance use disorder consequent to illicit drug use: comparison of a random sample and volunteers. AB - OBJECTIVE: Men qualifying for substance use disorder (SUD) consequent to consumption of an illicit drug were compared according to recruitment method. It was hypothesized that volunteers would be more self-disclosing and exhibit more severe disturbances compared to randomly recruited subjects. METHODS: Personal, demographic, family, social, substance use, psychiatric, and SUD characteristics of volunteers (N = 146) were compared to randomly recruited (N = 102) subjects. RESULTS: Volunteers had lower socioceconomic status, were more likely to be African American, and had lower IQ than randomly recruited subjects. Volunteers also evidenced greater social and family maladjustment and more frequently had received treatment for substance abuse. In addition, lower social desirability response bias was observed in the volunteers. SUD was not more severe in the volunteers; however, they reported a higher lifetime rate of opiate, diet, depressant, and analgesic drug use. CONCLUSIONS: Volunteers and randomly recruited subjects qualifying for SUD consequent to illicit drug use are similar in SUD severity but differ in terms of severity of psychosocial disturbance and history of drug involvement. The factors discriminating volunteers and randomly recruited subjects are well known to impact on outcome, hence they need to be considered in research design, especially when selecting a sampling strategy in treatment research. PMID- 15283946 TI - Neonatal abstinence syndrome in methadone-exposed infants is altered by level of prenatal tobacco exposure. AB - Maternal tobacco consumption during pregnancy has been associated with lower birth weight infants, preterm births, intrauterine growth retardation, smaller head circumference and increase in morbidity, yet few studies have examined the role tobacco has on the opiate neonatal abstinence syndrome (NAS). This study examined the effect of prenatal tobacco exposure on NAS for infants born to mothers maintained on methadone during gestation. Twenty-nine pregnant women and their newborn infants participated in this study. Tobacco exposure was based on maternal self-report with 16 women reporting cigarette consumption of 10 or less per day and 13 reporting smoking 20 cigarettes or more a day. The onset, peak, and duration of NAS were examined. Results showed that infants born to mothers who reported smoking 20 or more cigarettes per day had significantly higher NAS peak scores of 9.8 versus 4.8, and took longer to peak (113.0 h versus 37.8 h), than light smokers of 10 or fewer cigarettes per day. We concluded that tobacco use in conjunction with methadone plays an important role in the timing and severity of NAS in prenatally exposed infants. PMID- 15283947 TI - Correlates of attempted suicide among young injection drug users in a multi-site cohort. AB - The purpose of this study was to determine the prevalence and correlates of attempted suicide among young injection drug users (IDUs) from six study sites in five US cities. Two thousand two hundred and nineteen participants 15-30 years of age underwent interviewer-administered questionnaires relating to self-reported drug use, sociodemographics, suicidal ideation and attempts, and exposure to violence. The 6-month prevalence of suicidal ideation and attempts was 35.8% (n = 795) and 7% (n = 156), respectively. Compared to those not reporting a recent (past 6 months) suicide attempt, those attempting suicide were more likely to have a lifetime history of mental health facility admission or sexual abuse. Participants receiving drug treatment at the time of the baseline interview (53.2% versus 37.1%, odds ratio [OR] = 1.93, 95% confidence interval [CI]: 1.39, 2.67) were also more likely to report a recent attempt; as were those reporting a history of experiencing violence. These associations persisted after adjusting for age, sex, race/ethnicity, study site, and other significant covariates by multiple logistic regression. These data suggest that increased access to drug treatment, community mental health, and violence prevention programs may decrease suicidal behavior among young injection drug users. PMID- 15283948 TI - Evaluation of the reinforcing effects of atomoxetine in monkeys: comparison to methylphenidate and desipramine. AB - Atomoxetine is a selective norepinephrine (NE) reuptake blocker that has recently been marketed for the treatment of attention deficit hyperactivity disorder. The purpose of the present study was to evaluate the self-administration of atomoxetine in an animal model predictive of abuse liability in humans. Rhesus monkeys (N = 5) were prepared with chronic intravenous catheters and allowed to self-administer cocaine or saline during alternating baseline sessions. When behavior was stable, atomoxetine (0.03-3.0 mg/kg per injection), desipramine (0.1 3.0 mg/kg per injection), methylphenidate (0.001-0.1 mg/kg per injection), or their vehicles were substituted for baseline conditions. Methylphenidate consistently maintained responding above the levels maintained by its vehicle. Atomoxetine and desipramine failed to reliably maintain self-administration above vehicle levels in four of five individual monkeys. These results predict that atomoxetine, in contrast to methylphenidate but like desipramine, will lack reinforcing effects and abuse potential in humans. PMID- 15283949 TI - Cognitive impairments in sober alcoholics: performance on selective and divided attention tasks. AB - AIMS: The aim of this study was to investigate performance on neuropsychological tasks and tasks measuring different aspects of attention in a representative sample of sober alcoholics. METHODS: The study followed a between-groups design whereby sober alcoholics were compared to a matched non-alcoholic control group. The alcoholics were recruited from a six-week residential rehabilitation unit for addicts. A total of 98 alcoholics (64 males) and 30 non-alcoholic controls (22 males) participated in the study. The alcoholics were assessed on four standard neuropsychological tasks and three attention tasks from cognitive experimental literature targeted towards measuring attention. RESULTS: In comparison to a non alcoholic control group the alcoholics were significantly impaired on all neuropsychological tasks, the divided attention task and the Stroop task. However, a normal pattern of performance for the alcoholics appears in the Eriksen task, a measure of selective attention. Alcoholics also showed elevated levels of negative affect at treatment intake. CONCLUSION: It is concluded that a representative sample of alcoholics show specific deficits of attention as opposed to a general decline of attention at treatment intake. Thus, sober alcoholics appear to be as efficient as controls at selecting on the basis of location. Nonetheless when they are required to select on the basis of semantic information, as in the Stroop task, or required to respond to two independent sources of information, as in the divided attention task, they are at a deficit. This study has added to previous research by highlighting the specific attentional processes impaired in alcoholics. PMID- 15283950 TI - Effects of age at first substance use and psychiatric comorbidity on the development of substance use disorders. AB - In this paper, we examine the effects of age at first substance use, and history of psychiatric disorders, on the development of substance use disorder (SUD) by age 16. We use a prospective, longitudinal design to disaggregate the effects of age at first use and time since first use on the development of adolescent SUD. Second, we test the hypothesis that adolescent SUD is an unlikely progression from early substance use unless children also show other early conduct problems. A population sample of 1,420 children from the Great Smoky Mountains Study (GSMS) was assessed annually between ages 9 and 16. Logistic regression models were applied within the hierarchical Bayesian framework, where the covariate effects were described by time-varying parameters having a first-order auto-regressive prior distribution. Posterior analyses based on a Gibbs sampling approach revealed that, controlling for years of exposure, the risk of transition to SUD increased with age at onset for onsets before age 13, but began to fall for onset at 14. Among users, use alone, without early conduct problems, led to a 11% prevalence of SUD by age 16. Past conduct disorder (CD) had a strong additive effect at ages 13-15, but at age 16, when substance use and abuse became more normative, the excess risk from prior CD decreased. Boys, but not girls, with a history of depression were at increased risk of SUD. Anxiety increased the risk of SUD in girls at age 16, but not before that. Results only partially support the study hypothesis; early use was a major predictor of adolescent SUD even in the absence of CD. PMID- 15283951 TI - Long-term effects of ketamine: evidence for a persisting impairment of source memory in recreational users. AB - RATIONALE: Ketamine is an N-methyl-d-aspartate (NMDA)-receptor antagonist that is increasingly being used as a recreational drug. Previous research has shown gross generalised verbal memory impairments persisting 3 days after ketamine use in chronic users, however episodic memory has not specifically investigated in this population. OBJECTIVE: To determine whether ketamine, on the night of drug use (day 0) and 3 days later, is associated with impaired episodic memory as assessed by a source memory task. METHODS: Twenty ketamine users and 20 poly-drug controls were compared on a source memory task both on day 0 and 3. Participants also completed questionnaires on both days indexing schizophrenic-like and dissociative symptoms. RESULTS: On day 0, ketamine abusers were impaired on both source memory and item recognition and scored more highly on schizophrenic and dissociative symptom scales compared to poly-drug controls. On day 3 ketamine abusers only displayed source memory impairments and these positively correlated with the level of schizophrenic-like symptoms on day 0. No differences on day 3 in schizophrenic-like or dissociative symptoms were observed. CONCLUSIONS: Ketamine abusers exhibit a persisting deficit in source memory on day 3 but not in item recognition. These findings suggest that repeated use of ketamine produces chronic impairments to episodic memory. PMID- 15283952 TI - Behavioural activation as predictor of substance use: mediating and moderating role of attitudes and social relationships. AB - Alcohol, tobacco, and drug use were investigated in 4,501 Russian youths aged 14 25 years. The participants also filled out the short forms of the Gray-Wilson Personality Questionnaire and the Eysenck Personality Questionnaire along with questions about attitudes and social relationships. Behavioural Activation (BAS) was the best personality predictor of substance use. Its influence was mediated by disobedience to adults, affiliation with peers (Outings) and tolerant attitude toward illegal activity. BAS was negatively associated with subjective well-being and educational aspiration (Learn). Extraversion was the second strongest predictor of substance use with its influence being mostly mediated by Outings. Besides, Extraversion was positively associated with some protective factors such as subjective well-being, Learn and good relationship with parents. Effects of Neuroticism and Behavioural Inhibition (BIS) on substance use were weak and gender-specific. In females BIS provided a degree of protection while in males it increased the risk of substance use. The personality factors interacted so that BAS and Extraversion tended to mutually increase the impact of each other, while BIS diminished the effect of BAS. Among attitude variables, Outings acted as the most potent predictor of substance use. Relationship with parents was a protective factor, which acted most strongly in adolescents with higher Psychoticism and Extraversion. PMID- 15283953 TI - Ethanol does not alter the binding of the gamma-hydroxybutyric acid (GHB) receptor ligand [3H]NCS-382 in the rat brain. AB - We investigated the effect of ethanol on the binding of the gamma-hydroxybutyric acid (GHB) receptor ligand [3H]NCS-382 in the rat cerebral cortex and hippocampus. Ethanol (50-100 mM) did not alter the binding of [3H]NCS-382. Furthermore, acute (3g/kg, p.o.) as well as chronic (9-15 g/kg/day p.o. for 6 days) administration of ethanol also did not have any significant effect on the binding of [3H]NCS-382 in the rat cerebrocortical and hippocampal membranes. These observations suggest that ethanol does not interact directly with the GHB receptor in vitro or in vivo, and GHB receptor may not be involved in the pharmacological effects of ethanol. PMID- 15283955 TI - Sleep and rest facilitate auditory learning. AB - Sleep is superior to waking for promoting performance improvements between sessions of visual perceptual and motor learning tasks. Few studies have investigated possible effects of sleep on auditory learning. A key issue is whether sleep specifically promotes learning, or whether restful waking yields similar benefits. According to the "interference hypothesis," sleep facilitates learning because it prevents interference from ongoing sensory input, learning and other cognitive activities that normally occur during waking. We tested this hypothesis by comparing effects of sleep, busy waking (watching a film) and restful waking (lying in the dark) on auditory tone sequence learning. Consistent with recent findings for human language learning, we found that compared with busy waking, sleep between sessions of auditory tone sequence learning enhanced performance improvements. Restful waking provided similar benefits, as predicted based on the interference hypothesis. These findings indicate that physiological, behavioral and environmental conditions that accompany restful waking are sufficient to facilitate learning and may contribute to the facilitation of learning that occurs during sleep. PMID- 15283954 TI - Temporal relationship between the age of onset of phobic disorders and development of substance dependence in adolescent psychiatric patients. AB - AIMS: To investigate the age of onset of phobic disorders in relation to later development of substance dependence in a sample of adolescent psychiatric patients. DESIGN, SETTING AND PARTICIPANTS: Clinical sample of 238 adolescents (age 12-17) admitted to psychiatric inpatient hospitalization between April 2001 and July 2003. MEASUREMENTS: Psychiatric diagnoses and onset ages obtained from the schedule for affective disorders and schizophrenia for school aged children present and lifetime (K-SADS-PL). FINDINGS: Logistic regression analyses revealed that adolescents with phobic disorders had a 4.9-fold risk for comorbid substance dependence compared to those without phobia. The mean onset age was 11.4 and 14.4 years for phobias and comorbid substance dependence, respectively. Boys (13.7 years) had a statistically significantly lower onset age for substance dependence than girls (15.4 years). Over one-half of the adolescents with phobic disorders had developed substance dependence within three years after the onset of phobia. CONCLUSIONS: We found that phobias might influence the development of secondary substance dependence within a few years from the onset of phobia already in adolescence. PMID- 15283956 TI - Inhibition of nicotinic acetylcholine receptors by apolipoprotein E-derived peptides in rat hippocampal slices. AB - Apolipoprotein E (ApoE) is a well-known genetic risk factor for Alzheimer's disease (AD). Dysfunctions in cholinergic signaling, and in particular in the function of neuronal nicotinic acetylcholine receptors (nAChRs), have also been linked with AD and cognition. To address whether there is a link between ApoE and nAChR function, we used electrophysiological techniques to test the effects of synthetic ApoE-mimetic peptides derived from the low-density lipoprotein receptor (LDLR) binding domain for the ability to modulate nAChR activity in hippocampal interneurons. ApoE(133-149) completely inhibited ACh-evoked responses in a dose dependent manner, yielding an IC(50) value of 720+/-70 nM. A shorter peptide spanning residues 141-148 mimicked this effect while a second peptide spanning residues 133-140 was without effect, indicating that the arginine-rich domain is responsible for nAChR interaction. Inhibition of ACh-evoked responses was voltage independent, and displayed partial receptor specificity as no effect on glycine- or GABA-evoked responses occurred. These results demonstrate that peptides derived from the LDLR binding domain of ApoE block the function of nAChRs in hippocampal slices, an interaction that may have implications for AD. PMID- 15283957 TI - The role of IL-1beta in stress-induced sensitization of proinflammatory cytokine and corticosterone responses. AB - Proinflammatory cytokines often sensitize neuronal, hormonal, and behavioral responses to subsequent challenge. Recently, it was observed that exposure to inescapable tailshock enhances peripheral and central proinflammatory cytokine and corticosterone (CORT) responses to subsequent immune challenge up to 4 days later. Thus, we examined the role of central interleukin-1beta (IL-1beta) in stress-induced sensitization of proinflammatory cytokine and CORT responses to a subsequent immune challenge. Rats were administered IL-1 receptor antagonist (IL 1ra) or vehicle into the intra-cisterna magna 1 h prior to tailshock (100, 1.6 mA 5 s shocks) exposure. Twenty-four hours later, rats were challenged i.p. with 10 microg/kg lipopolysaccharide (LPS) and killed 1 h later. IL-1ra had no effect on basal proinflammatory cytokines, but completely blocked the stress-induced enhancement in central and pituitary IL-1beta and plasma IL-6 release following LPS challenge. IL-1ra had no effect on stress-induced enhancement in CORT responses following LPS challenge. Additional rats were administered i.c.v. hrIL 1beta or vehicle and returned to their home cage. Twenty-four hours later, rats were challenged i.p. with either saline or 10 microg/kg LPS and killed 1 h later. Central hrIL-1beta administration significantly elevated central IL-1beta levels and plasma CORT following LPS challenge compared with vehicle-injected controls. These data demonstrate that elevations in central IL-1beta, whether stress induced or exogenously administered, are sufficient for sensitizing central IL 1beta and CORT responses to subsequent immune challenge. However, during times of stress, exogenous central IL-1ra administration only blocked sensitization of subsequent central IL-1beta responses, not CORT responses, suggesting other factors during the stress response can sensitize CORT responses. PMID- 15283958 TI - Loss of cortical acetylcholine enhances amphetamine-induced locomotor activity. AB - Cholinergic disturbances have been implicated in schizophrenia. In a recent study we found that intracerebroventricular (i.c.v.) delivery of the immunotoxin 192 IgG-saporin, that effectively destroys cholinergic projections from the basal forebrain to hippocampus and cortex cerebri, leads to a marked facilitation of amphetamine-induced locomotor activity in adult rats. The aim of the present experiments was to evaluate the contribution of the septohippocampal versus the basalocortical cholinergic projections for the amphetamine hyper-response seen previously in i.c.v. 192 IgG-saporin injected rats. Since i.c.v. delivery of 192 IgG-saporin also destroys a population of Purkinje neurons in cerebellum, this cell loss needs to be taken into consideration as well. Cortex cerebri and hippocampus were selectively cholinergically denervated by intraparenchymal injections of 192 IgG-saporin into nucleus basalis magnocellularis and the medial septum/diagonal band of Broca, respectively. Selective loss of Purkinje cells in cerebellum was achieved by i.c.v. delivery of OX7 saporin. Possible effects of these three lesions on spontaneous and amphetamine-induced locomotor activity were assessed in locomotor activity cages. We find that selective cholinergic denervation of cortex cerebri, but not denervation of hippocampus or damage to cerebellum can elicit dopaminergic hyper-reactivity similar to that seen in previous i.c.v. 192 IgG-saporin experiments. Our data are compatible with the hypothesis that disturbances of cholinergic neurotransmission in cortex cerebri may be causally involved in forms of schizophrenia. PMID- 15283959 TI - Timing of administration mediates the memory effects of intraseptal carbachol infusion. AB - Medial septal neurons innervate the entire hippocampal formation. This input provides a potent regulation of hippocampal formation physiology (e.g. theta) and memory function. Medial septal neurons are rich in cholinergic receptors and thus are potential targets for the development of cognitive enhancers. Direct intraseptal infusion of cholinomimetics alters hippocampal physiology and can produce either promnestic or amnestic effects. Several variables (e.g. age of animal, integrity of septohippocampal circuits, task difficulty) may influence treatment outcome. We have previously demonstrated that intraseptal carbachol (12.5-125 ng) infusion immediately after the sample session of a delayed-non match-to-sample radial maze paradigm produces a dose-dependent amnesia. The present study examined whether manipulating the timing of intraseptal carbachol infusion with respect to the sample session would alter the amnestic effect. A within-subjects design was used to examine the effect of intraseptal carbachol (125 ng/0.5 microl) in a delayed-non-match to sample radial maze task. During a sample session, rats retrieved rewards from six of 12 maze arms. At the test session (3 h later), only the alternate set contained reward and entries into the sample set arms constituted errors. Intraseptal carbachol was administered: 1) 30 min prior; 2) immediately prior; 3) immediately after and 4) 90 min after the sample session. Intraseptal carbachol prior to the sample had no effect on any index of accuracy. Infusion immediately after the sample, or delayed 90 min into the retention interval, produced an acute amnesia. These findings demonstrate that the timing of treatment is a critical variable in determining the memory effects of septohippocampal manipulations and that dynamic changes in cholinergic tone are important for memory. PMID- 15283960 TI - Modulation of hippocampal cell proliferation, memory, and amyloid plaque deposition in APPsw (Tg2576) mutant mice by isolation stress. AB - Tg2576 transgenic mice (mice overexpressing the "Swedish" mutation in the human amyloid precursor protein 695) demonstrated a decreased capacity for cell proliferation in the dentate gyrus of the hippocampus compared with non transgenic littermates at 3 months, 6 months and 9 months of age. Isolation stress induced by individually housing each mouse from the time of weaning further decreased hippocampal cell proliferation in Tg2576 mice as well as in non transgenic littermates at 6 months of age. Decreases in hippocampal cell proliferation in isolated Tg2576 mice were associated with impairments in contextual but not cued memory. Fluoxetine administration increased cell proliferation and improved contextual memory in isolated Tg2576 mice. Further, isolation stress accelerated the age-dependent deposition of beta-amyloid 42 plaques in Tg2576 mice. Numerous beta-amyloid plaques were found in isolated but not non-isolated Tg2576 mice at 6 months of age. These results suggest that Tg2576 mice, a mouse model of Alzheimer disease, have an impaired ability to generate new cells in the dentate gyrus of the hippocampus and that the magnitude of this impairment can be modulated by behavioral interventions and drugs known to have effects on hippocampal neurogenesis in normal rodents. Unexpectedly, isolation stress also appeared to accelerate the underlying process of beta amyloid plaque deposition in Tg2576 mice. These results suggest that stress may have an impact on the underlying disease process associated with Alzheimer's disease. PMID- 15283961 TI - c-fos Changes following an aggressive encounter in female California mice: a synthesis of behavior, hormone changes and neural activity. AB - Although there has been growing interest in the neuroanatomical and physiological mechanisms underlying aggressive behavior, little work has focused on possible mechanisms controlling natural plasticity in aggression. In the current study, we used naturally occurring changes in aggression level displayed by female Peromyscus californicus across the estrous cycle and parallel changes in c-fos expression to examine possible brain regions involved in mediating this plasticity. We found that c-fos expression was increased in females exposed to a conspecific female intruder compared with control females in numerous brain regions thought to be involved in the control of aggression. More importantly, we found that c-fos increased in the bed nucleus of the stria terminalis (BNST) and ventral lateral septum (LSv) only in the more aggressive, diestrous females, and not in the less aggressive, proestrous and estrous females. Conversely, c-fos increased in the medial amygdala (MeA) across all stages of estrus compared with controls, suggesting the MeA is not involved in mediating changes in individual levels of aggression. Moreover, we found correlations between several measures of aggression and c-fos expression in the BNST and LSv but not the MeA, again suggesting a role in mediating aggression plasticity for the former two but not the latter brain region. We further hypothesize that the BNST and the LSv may be involved more generally in mediating natural changes in aggression, such as increases often observed after individuals win aggressive interactions against conspecifics. PMID- 15283962 TI - Glutamate transporters regulate excitability in local networks in rat neocortex. AB - Excitatory postsynaptic currents (EPSCs) in the neocortex are principally mediated by glutamate receptors. Termination of excitation requires rapid removal of glutamate from the synaptic cleft following release. Glutamate transporters are involved in EPSC termination but the effect of uptake inhibition on excitatory neurotransmission varies by brain region. Epileptiform activity is largely mediated by a synchronous synaptic activation of cells in local cortical circuits, presumably associated with a large release of glutamate. The role of glutamate transporters in regulating epileptiform activity has not been addressed. Here we examine the effect of glutamate transport inhibition on EPSCs and epileptiform events in layer II/III pyramidal cells in rat neocortex. Inhibiting glutamate transporters with DL-threo-beta-benzyloxyaspartic acid (TBOA; 30 microM) had no effect on the amplitude or decay time of evoked, presumably alpha-amino-3-hydroxyl-5-methyl-isoxazolepropionic acid-mediated, EPSCs. In contrast, the amplitude and duration of epileptiform discharges were significantly enhanced. TBOA resulted also in a decreased threshold for evoking epileptiform activity and an increased probability of occurrence of spontaneous epileptiform discharges. TBOA's effects were not inhibited by the group I and II metabotropic glutamate receptors antagonist (S)-alpha-methyl-4 carboxyphenylglycine or the kainate receptor antagonist [(3S,4aR, 6S, 8aR)-6-((4 carboxyphenyl)methyl-1,2,3,4,4a,5,6,7,8,8a-decahydroisoquinoline-3-carboxylic acid]. D-(-)-2-amino-5-phosphonovaleric acid could both prevent excitability changes by TBOA and block already induced changes. Dihydrokainate (300 microM) had effects similar to TBOA suggesting involvement of the glial transporter GLT 1. Inhibiting glutamate transport increases local network excitability under conditions where there is an enhanced release of glutamate. Our results indicate that uptake inhibition produces an elevation of extracellular glutamate levels and activation of N-methyl-D-aspartate receptors. PMID- 15283963 TI - Ca(2+)-calmodulin signalling pathway up-regulates GABA synaptic transmission through cytoskeleton-mediated mechanisms. AB - We investigated the role of calcium (Ca(2+))/calmodulin (CaM) signaling pathways in modulating GABA synaptic transmission at CA1 pyramidal neurons in hippocampal slices. Whole-cell pipettes were used to record type A GABA receptor (GABA(A)R) gated inhibitory postsynaptic currents (IPSCs) and to perfuse intracellularly modulators in the presence of glutamate receptor antagonists. GABA(A)R-gated IPSCs were enhanced by the postsynaptic infusions of adenophostin (1 microM), a potent agonist of inositol-1,4,5-triphosphate receptor (IP(3)R) that induces Ca(2+) release. The enhancement was blocked by co-infusing either 1,2-bis(2 aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid (10 mM) or CaM-binding peptide (100 microM). Moreover, the postsynaptic infusion of Ca(2+)-CaM (40/10 microM) enhanced both evoked and spontaneous GABA(A)R-gated IPSCs. The enhancement was attenuated by co-infusing 100 microM CaM-KII(281-301), an autoinhibitory peptide of CaM-dependent protein kinases. These results indicate that postsynaptic Ca(2+) CaM signaling pathways essentially enhance GABAergic synaptic transmission. In the investigation of synaptic targets for the enhancement, we found that IP(3)R agonist-enhanced GABA(A)R-gated IPSCs were attenuated by co-infusing colchicine (30 microM), vincristine (3 microM) or cytochalasin D (1 microM) that inhibits tubulin or actin polymerization, implying that actin filament and microtubules are involved. We conclude that postsynaptic Ca(2+)-CaM signaling pathways strengthen the function of GABAergic synapses via a cytoskeleton-mediated mechanism, probably the recruitment of receptors in the postsynaptic membrane. PMID- 15283964 TI - Phosphorylation of tau at THR212 and SER214 in human neuronal and glial cultures: the role of AKT. AB - We have reported recently that the microtubule-associated protein tau is phosphorylated in vitro by Akt, an important kinase in anti-apoptotic signaling regulated by insulin and growth factors. We also established that Akt phosphorylates tau separately at T212 and S214, two sites previously shown to be phosphorylated by glycogen synthase kinase 3beta (GSK3beta) and protein kinase A (PKA), respectively. In the present studies, we examined the relationship between Akt and T212/S214 in primary cultures of human neurons and astrocytes, and evaluated the contribution of two other kinases. In intact cells, we found a very low content of active (phospho-S473) form of Akt. We also found a low content of phospho-S214 but not phospho-T212 of tau, suggesting that only phospho-S212 may depend on Akt activity in situ. We upregulated Akt activity using two experimental models: treatment with a protein phosphatase inhibitor, okadaic acid, and transfection with a constitutively active Akt gene construct (c-Akt). Under these conditions, phosphorylation of tau at T212 and S214 was regulated independently, with little change or downregulation of phospho-T212 and dynamic upregulation of phospho-S214. Our studies revealed that Akt may influence the phospho-S214 content in a meaningful manner. They also revealed that PKA may only partially contribute to the phosphorylation of S214. In comparison, okadaic acid treatment severely depleted the content of GSK3beta and downregulated the remaining GSK3beta activity by Akt-dependent inhibition, consistent with minimal changes in phospho-T212. In summary, these results strongly suggest that in primary cultures, Akt selectively phosphorylates tau at S214 rather than T212. Our studies raise the possibility that tau S214 may participate in Akt-mediated anti-apoptotic signaling. PMID- 15283965 TI - Differential pH and capsaicin responses of Griffonia simplicifolia IB4 (IB4) positive and IB4-negative small sensory neurons. AB - Protons play a key role in nociception caused by inflammation and ischaemia, but little is known about the relative sensitivities of different dorsal root ganglion (DRG) neurons. We have therefore examined the responses in vitro of rat DRG cells classified according to whether or not they bind Griffonia simplicifolia IB4 (IB4), a lectin which is widely used to distinguish between two major populations of small diameter neurons. Under voltage-clamp conditions, proton-activated inward currents were found in approximately 90% of small DRG neurons and showed one of three waveforms: transient, sustained or mixed. The majority of IB4-positive (IB4+) neurons (63%) gave rise to sustained inward currents that were sensitive to capsazepine. In contrast, the most prevalent waveform in small IB4-negative (IB4-) neurons (69%) was a mixed response containing transient and sustained components. The transient component was inhibited by amiloride whilst the sustained component showed a variable sensitivity to capsazepine. We also found that more IB4+ cells responded to capsaicin and, on average, gave rise to a larger magnitude of response than small IB4- neurons, consistent with their higher prevalence and greater amplitude of vanilloid receptor 1 (TRPV1)-like acid responses. The increase in intracellular Ca(2+) induced by capsaicin was also slightly greater in IB4+ neurons and in these cells its magnitude correlated with the level of TRPV1 immunoreactivity. Our data suggest that acid-sensing ion channels (ASICs) and TRPV1 are the major acid-sensitive receptors in small IB4- neurons, whilst TRPV1 is the predominant one in IB4+ neurons. Because ASIC-like responses were approximately 10-fold more sensitive to changes in H(+) than TRPV1-like responses, we speculate that small IB4- rather than IB4+ neurons play an essential role in sensing acid. Our results also highlight differences in capsaicin responses between IB4+ and IB4- small neurons and reveal the close link between capsaicin responses and levels of TRPV1 expression. PMID- 15283966 TI - Moderate increases in intracellular calcium activate neuroprotective signals in hippocampal neurons. AB - Although large increases in neuronal intracellular calcium concentrations ([Ca(2+)](i)) are lethal, moderate increases in [Ca(2+)](i) of 50-200 nM may induce immediate or long-term tolerance of ischemia or other stresses. In neurons in rat hippocampal slice cultures, we determined the relationship between [Ca(2+)](i), cell death, and Ca(2+)-dependent neuroprotective signals before and after a 45 min period of oxygen and glucose deprivation (OGD). Thirty minutes before OGD, [Ca(2+)](i) was increased in CA1 neurons by 40-200 nM with 1 nM-1 microM of a Ca(2+)-selective ionophore (calcimycin or ionomycin-"Ca(2+) preconditioning"). Ca(2+) preconditioning greatly reduced cell death in CA1, CA3 and dentate during the following 7 days, even though [Ca(2+)](i) was similar (approximately 2 microM) in preconditioned and control neurons 1 h after the OGD. When pre-OGD [Ca(2+)](i) was lowered to 25 nM (10 nM ionophore in Ca(2+)-free medium) or increased to 8 microM (10 microM ionophore), more than 90% of neurons died. Increased levels of the anti-apoptotic protein protein kinase B (Akt) and the MAP kinase ERK (p42/44) were present in preconditioned slices after OGD. Reducing Ca(2+) influx, inhibiting calmodulin, and preventing Akt or MAP kinase p42/44 upregulation prevented Ca(2+) preconditioning, supporting a specific role for Ca(2+) in the neuroprotective process. Further, in continuously oxygenated cultured hippocampal/cortical neurons, preconditioning for 30 min with 10 nM ionomycin reduced cell death following a 4 microM increase in [Ca(2+)](i) elicited by 1 microM ionomycin. Thus, a zone of moderately increased [Ca(2+)](i) before a potentially lethal insult promotes cell survival, uncoupling subsequent large increases in [Ca(2+)](i) from initiating cell death processes. PMID- 15283967 TI - Expression of aquaporin water channels in mouse spinal cord. AB - Aquaporins (AQPs) are membrane proteins involved in water transport in many fluid transporting tissues. Aquaporins AQP1, AQP4, and AQP9 have been identified in brain and hypothesized to participate in brain water homeostasis. Here we use reverse transcriptase-polymerase chain reaction (RT-PCR), Western blotting and immunohistochemistry to describe the expression and immunolocalization of AQPs in adult mouse spinal cord. AQP4 was expressed in glial cells, predominantly in gray matter, and in astrocytic end-feet surrounding capillaries in spinal cord white matter. AQP9 expression extensively co-localized with glial fibrillary acidic protein-immunoreactive astrocytes, located predominantly in the white matter. AQP5 was detected by RT-PCR but not by immunohistochemical analysis. Interestingly, AQP8 was detected primarily in ependymal cells lining the fluid filled central canal. The aquaporin expression pattern in spinal cord suggests involvement in water homeostasis and diseases associated with abnormal water fluxes such as spinal cord injury and syringomyelia. PMID- 15283968 TI - Bi-directional associations between galanin and luteinizing hormone-releasing hormone neuronal systems in the human diencephalon. AB - Evidence suggests that galanin plays an important role in the regulation of reproduction in the rat. Galanin is colocalized with luteinizing hormone (LH) releasing hormone (LHRH) in a subset of LHRH neurons in female rats and galanin immunoreactive (galanin-IR) nerve terminals innervate LHRH neurons. Recent studies indicate that galanin may control gonadal functions in rats at two different levels: (i) via direct modulation of pituitary LH secretion and/or (ii) indirectly via the regulation of the hypothalamic LHRH release. However, the morphological substrate of any similar modulation is not known in human. In the present series of experiments we first mapped the galanin-IR and LHRH-IR neural elements in human brain, utilizing single label immunohistochemistry. Then, following the superimposition of the maps of these systems, the overlapping sites were identified with double labeling immunocytochemistry and examined in order to verify the putative juxtapositions between galanin-IR and LHRH-IR structures. LHRH and galanin immunoreactivity were detected mainly in the medial basal hypothalamus, in the medial preoptic area and along the diagonal band of Broca. Careful examination of the IR elements in the overlapping areas revealed close, bi-directional contacts between galanin-IR and LHRH-IR structures, which have been verified in semithin plastic sections. These galanin-LHRH and LHRH-galanin juxtapositions were most numerous in the medial preoptic area and in the infundibulum/median eminence of the human diencephalon. In conclusion, the present study is the first to reveal bi-directional juxtapositions between galanin- and LHRH-IR neural elements in the human diencephalon. These galanin LHRH and LHRH-galanin contacts may be functional synapses, and they may be the morphological substrate of the galanin-controlled gonadal functions in humans. PMID- 15283969 TI - Differential subcellular and subsynaptic distribution of GABA(A) and GABA(B) receptors in the monkey subthalamic nucleus. AB - The activation of GABA receptor subtype A (GABA(A)) and GABA receptor subtype B (GABA(B)) receptors mediates differential effects on GABAergic and non-GABAergic transmission in the basal ganglia. To further characterize the anatomical substrate that underlies these functions, we used immunogold labeling to compare the subcellular and subsynaptic localization of GABA(A) and GABA(B) receptors in the subthalamic nucleus (STN). Our findings demonstrate major differences and some similarities in the distribution of GABA(A) and GABA(B) receptors in the monkey STN. The immunoreactivity for GABA(A) receptor alpha1 subunits is mostly bound to the plasma membrane, whereas GABA(B) R1 subunit alpha1 immunoreactivity is largely expressed intracellularly. Plasma membrane-bound GABA(A) alpha1 subunit aggregate in the main body of putative GABAergic synapses, while GABA(B) R1 receptors are found at the edges of putative glutamatergic or GABAergic synapses. A large pool of plasma membrane-bound GABA(A) and GABA(B) receptors is extrasynaptic. In conclusion, these findings demonstrate a significant degree of heterogeneity between the distributions of the two major GABA receptor subtypes in the monkey STN. Their pattern of synaptic localization puts forward interesting questions regarding their mechanisms of activation and functions at GABAergic and non-GABAergic synapses. PMID- 15283970 TI - Identification of neural pathways involved in genital reflexes in the female: a combined anterograde and retrograde tracing study. AB - The medial preoptic area (MPOA) is important for reproductive behavior in females. However, the descending pathways mediating these responses to the spinal motor output are unknown. The MPOA does not directly innervate the spinal cord. Therefore, pathways mediating MPOA-induced changes in sexual behavior must relay in the brain. The nucleus paragigantocellularis (nPGi) projects heavily to spinal circuits involved in female sexual reflexes and is involved in the tonic inhibition of genital reflexes. However, the periaqueductal gray (PAG) is also important for female sexual behavior. The present study examined the hypothesis that the MPOA output relays through PAG and the nPGi before descending to the spinal cord. We used anterograde and retrograde tracing techniques to examine the descending pathways and relay sites from the MPOA to the spinal cord and the nPGi in the female rat. Injection of biotinylated dextran amine into the MPOA produced dense labeling in specific regions of the PAG and Barrington's nucleus; anterogradely labeled fibers terminated close to neurons retrogradely labeled from the spinal cord in the PAG, Barrington's nucleus, nPGi, lateral hypothalamus and paraventricular nucleus (PVN). Anterogradely labeled fibers and varicosities were also found close to neurons retrogradely labeled from the nPGi in the PAG, lateral hypothalamus and PVN. These results suggest that the major MPOA output relays in the PAG and nPGi before descending to innervate spinal circuits regulating female genital reflexes and that the MPOA plays a multifaceted role in female reproductive behavior through its modulation of PAG output systems. PMID- 15283971 TI - Cloning and characterization of alpha9 subunits of the nicotinic acetylcholine receptor expressed by saccular hair cells of the rainbow trout (Oncorhynchus mykiss). AB - alpha9/alpha10 Subunits are thought to constitute the nicotinic acetylcholine receptors mediating cholinergic efferent modulation of vertebrate hair cells. The present report describes the cloning and sequence analysis of subunits of the alpha9-containing receptor of a hair-cell layer from the saccule of the rainbow trout (Oncorhynchus mykiss). A major alpha9 subunit, termed alpha9-I, displayed typical features of a nicotinic alpha subunit, with total coding sequence of 572 amino acids including a 16 amino-acid signal peptide. It possessed an extended cytoplasmic loop between membrane-spanning regions M3 and M4, compared with mammalian homologs. Transcript for alpha9-I was robustly expressed in the saccular hair cell layer and less prominently in trout olfactory mucosa, spleen, pituitary gland, and liver, as determined by reverse transcription-polymerase chain reaction. alpha9-I cDNA was not detected in trout brain, skeletal muscle, retina, and kidney. The alpha9-I nicotinic receptor protein was immunolocalized, with an affinity-purified antibody directed against a trout alpha9-I epitope, to hair-cell and neural sites in the saccular hair-cell layer. Foci were found at basal and basolateral membrane sites on hair cells as well as on afferent nerve. Receptor clustering was observed in hair cells bordering non-sensory epithelium. Since in higher vertebrates the alpha9 is reported to associate with another nicotinic subunit, alpha10, we examined the possibility of expression of additional nicotinic subunits in trout saccular hair cells. Message for another nicotinic subunit, termed alpha9-II, was found to be expressed in the hair cells, although more difficult to amplify than alpha9-I. In contrast to alpha9-I, alpha9 II was expressed in brain, as well as in olfactory mucosa, less prominently in pituitary gland and liver, but not in spleen, skeletal muscle, retina, or kidney. The cloned alpha9-II had a total coding sequence of 550 amino acids, which included a 17-amino-acid signal peptide, and an extended M3-M4 loop. A third nicotinic subunit message, termed alpha9-III, was PCR-amplified from trout olfactory mucosa where it was strongly expressed. However, message for alpha9-III was not detected in hair cells. Message for alpha9-III was moderately expressed in trout brain, retina, and pituitary gland but not in trout spleen, skeletal muscle, liver, and kidney. Thus, alpha9-I and alpha9-II may together contribute to the formation of the hair-cell nicotinic receptor of teleosts, where no ortholog of alpha10 appears to exist. The current work is, to our knowledge, the first description of alpha9 coding sequences directly from a vertebrate hair cell source. Further, the generality of hair cell expression of subunits for the alpha9-containing nicotinic cholinergic receptor has been extended to fishes, suggesting a similar efferent mechanism across all vertebrate octavolateralis sensory systems. PMID- 15283972 TI - Non-angiotensin II [(125)I] CGP42112 binding is a sensitive marker of neuronal injury in brainstem following unilateral nodose ganglionectomy: comparison with markers for activated microglia. AB - Previously we reported that a non-angiotensin II [(125)I] CGP42112 binding site is up-regulated in rat brainstem nuclei as a result of unilateral nodose ganglionectomy. In the present study, we compared non-angiotensin II [(125)I] CGP42112 binding with microglia/macrophage activation following nodose ganglionectomy, using both in vitro autoradiography and immunohistochemistry. Specific [(125)I] CGP42112 binding was observed in the nucleus of the solitary tract (NTS) and revealed an AT(2) receptor component as well as a non-angiotensin II receptor component. Subsequent to unilateral nodose ganglionectomy, [(125)I] CGP42112 binding in the ipsilateral NTS was increased approximately two-fold and was also induced in the ipsilateral dorsal motor nucleus (DMX) and the nucleus ambiguus (n.amb). This non-angiotensin II [(125)I] CGP42112 binding site was displaced by CGP42112 but not other ligands. Increased [(3)H] PK11195 binding (a known marker of reactive gliosis) was also observed in the same brainstem nuclei as non-angiotensin II [(125)I] CGP42112 binding after nodose ganglionectomy. The similarity in binding patterns between [(125)I] CGP42112 and [(3)H] PK11195 was shown to be primarily due to retrograde degeneration in the ipsilateral NTS, DMX and n.amb, as both radioligands were localized to similar cellular targets within the interstial space and over cellular debris. Immunohistochemical data confirmed reactive gliosis within the ipsilateral NTS, DMX and n.amb, following nodose ganglionectomy, which was predominantly characterized by an increase in OX-42 immunoreactivity (a marker for activated microglia/macrophages), with only a small increase in glial fibrillary acidic protein immunoreactivity (a marker of astrogliosis) detected. These data demonstrate for the first time that non angiotensin II [(125)I] CGP42112 binding is associated with activated microglia, as well as macrophages, following unilateral nodose ganglionectomy. Furthermore, these studies also demonstrate the potential use of non-angiotensin II [(125)I] CGP42112 binding as a marker for quantitating inflammatory events which occur as a result of damage to the CNS. PMID- 15283973 TI - Plastic interactions between hand and face cortical representations in patients with trigeminal neuralgia: a somatosensory-evoked potentials study. AB - Neurophysiological and neuroimaging studies suggest that pain may play a major role in determining cortical somatosensory rearrangements even in the adult brain. The re-organizational power of pain, however, has been tested in models in which massive deafferentation co-existed with pain (e.g. in phantom pain). Moreover, information on whether spinal and brainstem changes contribute to pain related plasticity in humans is meagre. We used the non-invasive somatosensory evoked potentials technique in patients with right primary trigeminal neuralgia and no clinical signs of large-diameter fibers of trigeminal deafferentation to assess whether pain may induce plastic changes at multiple levels in the somatosensory system. Subcortical and cortical potentials evoked by stimulation of the right median and posterior tibial nerves ipsilateral to the facial pain were compared with those obtained following stimulation of the left median and tibial nerves and with those obtained in a control group tested in comparable conditions. Amplitudes of parietal N20 and P27 and frontal N30 potentials observed following stimulation of the right median nerve ipsilateral to the facial pain were greater than those of the left median nerve and showed a positive correlation with magnitude of pain. This right-left asymmetry was absent following stimulation of the patients' tibial nerves and in control subjects. No changes were found in spinal N13 and brainstem P14. That facial pain is associated with neuroplastic changes within the somatic cortical representation of the hand suggests a pain-related topographic cortical reorganisation. PMID- 15283974 TI - An in vivo profile of beta-endorphin release in the arcuate nucleus and nucleus accumbens following exposure to stress or alcohol. AB - The aim of the present study was to determine the effects of distinct categories of stressors on beta-endorphin (beta-EP) release in the arcuate nucleus (ArcN) and nucleus accumbens (NAcb) using in vivo microdialysis. Adult male rats were implanted with a cannula aimed at either the NAcb or the ArcN. On the day of testing, a 2 mm microdialysis probe was inserted into the cannula, and artificial cerebrospinal fluid was infused at 2.0 microl/min. After three baseline collections, animals either had a clothespin applied to the base of their tail for 20 min (a physical/tactile stressor), were exposed to fox urine odour for 20 min (a psychological stressor/species-specific threat), or were administered 2.4 g ethanol/kg body weight, 16.5% w/v, i.p. (a chemical/pharmacological stressor) with control animals receiving an equivalent volume of saline. Both tail-pinch and fox odour significantly increased beta-EP release from the ArcN (P<0.05), whilst only tail-pinch enhanced beta-EP release from the NAcb (P<0.01). On the other hand, alcohol stimulated beta-EP release in the NAcb as compared with saline-treated controls (P<0.01), but not in the ArcN. Although the increase in extracellular beta-EP produced by the other stressors was relatively rapid, there was a 90-min delay before alcohol administration caused beta-EP levels to exceed that of saline-injected controls. In conclusion, the fact that physical and fear inducing psychological stressors stimulate beta-EP release in the ArcN and only physical stressors stimulate beta-EP release in the NAcb, indicates that stressors with different properties are processed differently in the brain. Also, an injection of alcohol caused a delayed increase of beta-EP in the NAcb but not the ArcN, indicating that alcohol may recruit a mechanism that is, at least partially, distinct from stress-related pathways. PMID- 15283975 TI - Abnormal motor behavior and vestibular dysfunction in the stargazer mouse mutant. AB - In stargazer mutant mice, a mutation in the gene encoding stargazin results in absence epilepsy, cerebellar ataxia, and a characteristic abnormal motor syndrome. The main goal of the current studies was to characterize the nature and source of the abnormal motor behavior. Because the stargazer motor syndrome resembles that of other rodents with vestibular dysfunction, the motor abnormalities were compared with those of normal mice treated with toxins known to damage the vestibular system. Quantitative open field assessments revealed that the stargazer mice display a motor syndrome very similar to that exhibited by mice with toxin-induced vestibulopathy. However, stargazer mice also displayed several additional behaviors, such as ataxic gait and sustained extensor movements of the neck. In addition, stargazer mice performed worse than mice with toxin-induced vestibulopathy in most standard tests of motor function. Motor function was also impaired on each of four behavioral tests sensitive to vestibular function. Because of the close associations between the vestibular and auditory systems, tests of auditory function were also employed. The stargazer mutants exhibited relatively normal auditory brainstem evoked responses but no apparent acoustic startle reflex. Histological examination of vestibular sensory epithelium at the light and electron microscopic levels confirmed the existence of abnormalities in the stargazer mutants. These results imply a previously unrecognized role for stargazin in the normal functions of the vestibular system and indicate that some, but not all, of the abnormal motor syndrome of stargazer mice can be attributed to vestibular dysfunction. PMID- 15283976 TI - Thyroid hormone (T3) and its acetic derivative (TA3) protect low-density lipoproteins from oxidation by different mechanisms. AB - Triiodothyronine (T3) and triiodothyroacetic acid (TA3) are thyroid compounds that similarly protect low-density lipoprotein (LDL) against oxidation induced by the free radical generator 2,2'-azobis-[2-amidinopropane] dihydrochloride (AAPH). However, TA3 is more antioxidant than T3 on LDL oxidation induced by copper ions (Cu2+), suggesting that these compounds act by different mechanisms. Here we measured conjugated diene production kinetics during in vitro human LDL (50 mg LDL-protein per l) oxidation induced by various Cu2+ (0.5-4 microM) or AAPH (0.25 2 mM) concentrations in the presence of T3, TA3, butylated hydroxytoluene (BHT) (a free radical scavenger) or ethylenediaminetetracetic acid (EDTA) (a metal chelator). From the kinetics were estimated: length of the lag phase (Tlag), maximum velocity of conjugated diene production (Vmax), and maximum amount of generated dienes (Dmax). Thyroid compound effects on these oxidation parameters were compared to those of the controls BHT and EDTA. In addition we measured by atomic absorption spectrometry copper remaining in LDL after a 30 min incubation of LDL with Cu2+ and the compounds followed by extensive dialysis, i.e. copper bound to LDL. As expected, LDL-copper was decreased by EDTA in a concentration dependent manner, whereas it was not affected by BHT. T3 increased LDL-copper whereas TA3 slightly decreased it. The whole data suggest that T3 and TA3 are free radical scavengers that also differently disturb LDL-copper binding, an essential step for LDL lipid peroxidation. The most likely mechanisms are that T3 induces new copper binding sites inside the LDL particle, increasing the LDL copper amount but in a redox-inactive form, whereas TA3 blocks some redox-active copper binding sites highly implicated in the initiation and the propagation of lipid peroxidation. Alternatively, we also found that a little amount of copper is tightly bound in LDL, which may be essential for the propagation of lipid peroxidation induced by free radical generators. PMID- 15283977 TI - Disclosing the subterranean treasury of plants. AB - Roots are crucial for plant growth and development but, so far, their beneficial qualities have not been used in genetic engineering strategies. To exploit this missed opportunity, a thorough understanding is required of the genetic mechanisms that control gene expression in response to different cues. A global map of gene expression for the Arabidopsis root has recently been published that consists of high-resolution spatial- and temporal-expression profiles. This new dataset will be a valuable starting point for understanding the hidden organ of plants. PMID- 15283978 TI - Extracting novel information from gene expression data. AB - Data from high throughput technologies, such as DNA microarrays, necessitated the development of new computational methodologies for analyzing the high dimensional information contained within the gene expression data. Liao's group suggested the use of network component analysis to predict transcription factor activities by integrating gene expression data from Escherichia coli with known connectivity information between their genes and transcription factors. This introduces an approach for obtaining novel information from gene expression data. PMID- 15283979 TI - Bacteriophage biocontrol of plant pathogens: fact or fiction? AB - Bacterial resistance due to the misuse of antibiotics has become a global issue and alternative methods are being developed that might decrease the use of antimicrobials in agricultural settings. Bacteriophage therapy represents a novel way to control the growth of plant-based bacterial pathogens. Although this method shows promise, a recent paper by Gill and Abedon has shown that the complex bacteriophage-host interactions in the plant environment must be investigated further. PMID- 15283980 TI - Dkk-1-mediated expansion of adult stem cells. AB - Despite the wealth of research on stem-cell biology and therapy, there have been few advances in the ex vivo expansion of adult stem cells. Fetal bovine serum, which can elicit an immune reaction in hosts receiving cellular products, remains essential to the culture methods. Recent work by Prockop et al. suggests that Dickkopf-1 (Dkk-1) could stimulate the proliferation of mesenchymal stem cells (MSCs) while maintaining an undifferentiated phenotype. The use of Dkk-1 in cell processing laboratories might advance cell-expansion technology and greatly increase the clinical uses of MSCs. PMID- 15283981 TI - Stem cells: a lesson from dedifferentiation. PMID- 15283982 TI - Challenges for RNAi in vivo. AB - Synthetic small interfering RNA (siRNA) has become a valuable tool for investigating gene function in cell culture. This success has led to high expectations for siRNA as a tool for in vivo investigation and as a platform for therapeutic development. siRNA in cell culture owes much of its success to years of development of traditional antisense oligonucleotides, and in vivo applications will also benefit from previous experience in this regard. However, the duplex nature of siRNA presents significant obstacles that will need to be overcome. Here, we discuss the current status of in vivo siRNA technology and describe some of the barriers to widespread application of RNAi-mediated gene silencing in mammals. PMID- 15283983 TI - What is disrupting IFN-alpha's antiviral activity? AB - Despite advances in treatment strategies for hepatitis C virus (HCV), a significant proportion of patients fail to achieve viral clearance following treatment with pegylated interferon (IFN)-alpha plus ribavirin. Many of these individuals show elevated levels of tumor necrosis factor (TNF)-alpha compared with normal controls, and recent data have implicated this cytokine in the negative regulation of IFN-alpha. Although a therapeutic opportunity for TNF alpha antagonists might exist for reducing inflammation in chronic HCV disease, further exploration is required to identify the key mediators of responsiveness to IFN-alpha. In particular, the interplay should be clarified between host response factors [e.g. IFN-alpha, IFN-gamma, suppressor of cytokine signaling (SOCS), TNF-alpha and others] and pathogen-associated molecular patterns [PAMPs, e.g. lipopolysaccharide (LPS) and CpG DNA] in HCV disease; this information might guide future therapies aimed at improving IFN-alpha responsiveness. PMID- 15283984 TI - Comparison of network-based pathway analysis methods. AB - Network-based definitions of biochemical pathways have emerged in recent years. These pathway definitions insist on the balanced use of a whole network of biochemical reactions. Two such related definitions, elementary modes and extreme pathways, have generated novel hypotheses regarding biochemical network function. The relationship between these two approaches can be illustrated by comparing and contrasting the elementary modes and extreme pathways of previously published metabolic reconstructions of the human red blood cell (RBC) and the human pathogen Helicobacter pylori. Descriptions of network properties generated by using these two approaches in the analysis of realistic metabolic networks need careful interpretation. PMID- 15283985 TI - When biotech proteins go off-patent. AB - The patents of the first generation of biopharmaceuticals derived from recombinant DNA such as interferons, growth hormone and epoietins are expiring, opening up the possibility for competitors to introduce biosimilar products. The concept of generics that applies to classical drugs and allows market admission on limited documentation cannot be extrapolated to these "off-patent biologics". Physicochemical characterization, bioassays and animals studies do not predict completely the efficacy and safety of therapeutic proteins. Clinical studies will nearly always be necessary to obtain marketing authorization for off-patent biologics. Immunogenicity is considered to be the main problem with therapeutic proteins. The recent upsurge of pure red cell aplasia (PRCA), a severe form of anemia associated with the use of epoietin-alpha, highlights both the unpredictability and the severe consequences of immunogenicity. A risk-based approach can be used to evaluate the potential induction of antibodies by off patent biologics. PMID- 15283986 TI - Biomarker discovery and validation: technologies and integrative approaches. AB - The emerging field of biomarkers has applications in the diagnosis, staging, prognosis and monitoring of disease progression, as well as in the monitoring of clinical responses to a therapeutic intervention and the development and delivery of personalized treatments to reduce attrition in clinical trials. Moreover, biomarkers have a positive impact on health economics. The word "biomarker" has been used extensively across therapeutic areas and many disciplines, and its nature takes into consideration clinical, physiological, biochemical, developmental, morphological and molecular measures. In drug trials, biomarkers have been proposed for use in efficacy determination and patient population stratification, in deducing pharmacokinetic-pharmacodynamic relationships and in safety monitoring. The interfacing and integration of different technologies for data collection and analysis are pivotal to biomarker identification, characterization, validation and application. "Integrative functional informatics" represents a novel direction in such technology integration. PMID- 15283987 TI - Towards microbial tissue engineering? AB - Tissue engineering involves the creation of multicellular tissues from individual cells. It was previously perceived that tissues were only formed by higher organisms such as plants and animals. However, it is now known that multicellular systems of microorganisms, such as microbial colonies, biofilms, flocs and aggregates, can also show extensive spatial organization. Here, we discuss methods that can be used to spatially organize microorganisms--bacteria, in particular--into tissue-like materials with defined internal architectures. Some potential uses of such "microbial tissues" are covered. PMID- 15283988 TI - Design and development of oral drugs for the prophylaxis and treatment of smallpox infection. AB - Smallpox was eradicated by the World Health Organization (WHO) vaccination campaign in the 1970s and the variola virus was restricted to repositories in the United States and Russia. Recently, however, concerns have arisen about the possible existence of variola outside these sites and the potential for using the virus as a weapon of bioterror. The world population now has little residual immunity to smallpox and supplies of the smallpox vaccine are being reconstituted. Large numbers of individuals with various skin diseases or immunosuppression owing to AIDS or organ transplantation medications, or who are pregnant or have heart disease might not be ideal candidates for vaccination with the current live vaccines. It would be useful to have an orally active drug that could be self-administered in case of an outbreak of smallpox. PMID- 15283989 TI - Expression of phosphacan and neurocan during early development of mouse retinofugal pathway. AB - We have investigated whether the two major brain chondroitin sulfate (CS) proteoglycans (PGs), phosphacan and neurocan, are expressed in patterns that correlate to the axon order changes in the mouse retinofugal pathway. Expression of these proteoglycans was examined by polyclonal antibodies against phosphacan and N- and C-terminal fragments of neurocan. In E13-E15 mouse embryos, when most optic axons grow in the chiasm and the optic tract, phosphacan and neurocan were observed in the inner regions of the retina. In the chiasm and the tract, phosphacan but not neurocan was expressed prominently at the midline and in the deep parts of the tract. Both proteoglycans were observed on the chiasmatic neurons, which have been shown to regulate axon divergence at the chiasmatic midline and the chronotopic fiber ordering in the tract, but phosphacan appeared to be the predominant form that persists to later developmental stages. Intense staining of both proteoglycans was also observed in a strip of glial-like elements in lateral regions of the chiasm, partitioning axons in the stalk from those in the tract. We conclude that phosphacan but not neurocan is likely the major carrier of the CS glycosaminoglycans that play crucial functions in axon divergence and age-related axon ordering in the mouse optic pathway. Furthermore, localization of these carrier proteins in the optic pathway raises a possibility that these two proteoglycans regulate axon growth and patterning not only through the sulfated sugars but also by interactions of the protein parts with guidance molecules on the optic axons. PMID- 15283990 TI - Temporal changes in cerebral antioxidant enzyme activities after ischemia and reperfusion in a rat focal brain ischemia model: effect of dietary fish oil. AB - This study investigated the neuroprotective effects of dietary supplementation of fish oil on both brain infarction and the activities of antioxidant enzymes. Male Sprague-Dawley rats (4-weeks old) were divided into two groups and received either a regular diet (RD) or a fish-oil-supplemented diet (FOD) for 6 weeks prior to middle cerebral artery (MCA) occlusion. The infarction volume of the brain was calculated using image analysis after staining. Antioxidant enzymes were measured before ischemia (BI), after 2 h of ischemia (AI) and after 24 h (24hR), 48 h (48hR) and after 7 days (7dR) of reperfusion. The infarction volume of the brain was significantly smaller in the FOD group than in the RD group after 24 h of reperfusion (p<0.05). Before ischemia, the levels of lipid peroxide and the glutathione peroxidase (GPx) activity were higher in the FOD group than in the RD group. During reperfusion, the catalase (CAT) activity in the FOD group remained at the preischemia level until after 48 h of reperfusion, while those in the RD group did not. The Mn-superoxide dismutase (SOD) activity and GPx activity were higher in the FOD group than in the RD group only after 2 h of ischemia. In the fatty acid analysis, the ratio of docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) were higher in the FOD group than in the RD group (p<0.05). Our results demonstrate that supplementing the diet with fish oil could decrease the cerebral infarction volume following ischemia and reperfusion (I/R) partly by working directly as an antioxidant and partly by modulating antioxidant enzyme activities. PMID- 15283991 TI - Infant rats with chronic neonatal isolation experience show decreased extracellular serotonin levels in ventral striatum at baseline and in response to cocaine. AB - Previously, we demonstrated that the early life stress of neonatal isolation enhances extracellular dopamine (DA) levels in ventral striatum in response to psychostimulants in infant rats. Yet, neonatal isolation does not alter baseline DA levels. DA levels are affected by serotonin (5-HT) and striatal levels of this transmitter are also enhanced by cocaine. Other early life stresses are reported to alter various 5-HT neural systems. Thus, the purpose of this study is to test whether neonatal isolation alters ventral striatal 5-HT levels at baseline or in response to cocaine. Litters were subjected to neonatal isolation (1-h individual isolation/day on postnatal days 2-9) or to non-handled conditions and pups assigned to one of three cocaine doses (0, 2.5, or 5.0 mg/kg) groups. On postnatal day 10, probes were implanted in the ventral striatum. Dialysate samples obtained over a 60-min baseline period and for 120 min post cocaine injections were assessed for levels of 5-HT and its metabolite, 5-HIAA. ISO decreased ventral striatal 5-HT levels at baseline and after cocaine administration but did not alter 5-HIAA levels. These data add to the literature on the immediate effects of early life stress on 5-HT systems by showing alterations in the ventral striatal system. Because serotonergic effects in this neural area are associated with reward and with emotion and affect regulation, the results of this study suggest that early life stress may be a risk factor for addiction and other psychiatric disorders. PMID- 15283992 TI - Endothelial cell-astrocyte interactions and TGF beta are required for induction of blood-neural barrier properties. AB - We sought to establish a blood-neural barrier (BNB) model of astrocyte contact with endothelial cells (EC) to test the hypothesis that transforming growth factor beta (TGF beta) promotes an EC barrier-phenotype. Astrocyte-EC contact induced BNB properties in EC. Transendothelial resistance was augmented by direct contact between astrocytes-EC, but not by astrocyte-conditioned medium or astrocyte-EC coculture conditioned medium. Coculture of EC and astrocytes led to significant increase in endothelial occludin levels and junctional localization. EC gamma-glutamyl-transferase (GGT) activity was increased by direct contact with astrocytes, by conditioned medium from cocultures or by TGF beta1. Coculture inhibited EC proliferation with no effect on astrocyte proliferation. A neutralizing antibody to TGF beta decreased GGT activity in cocultures and increased cell number. Whereas total TGF beta was not significantly altered by coculture, activated TGF beta increased in astrocyte-EC cocultures. In summary, astrocyte-EC contact induces BNB characteristics in EC and locally activated TGF beta is responsible for part of the induction. PMID- 15283993 TI - Postnatal ontogeny of natriuretic peptide systems in the rat hypothalamus. AB - Our study has attempted to clarify the developmental profile of atrial natriuretic peptide (ANP) and C-type natriuretic peptide (CNP) along with the expression of their receptors in the rat hypothalamus. Radioimmunoassay (RIA) of dissected hypothalamic tissue revealed that ANP rose from 167 +/- 50 pg/mg protein immediately after birth to 516 +/- 78 pg/mg protein in the next 24 h and to 928 +/- 100 pg/mg protein by postnatal day (PD) 5. A second increment of ANP in the hypothalamus was noted between PD 10 and PD 20 (from 780 +/- 110 to 2,650 +/- 136 pg/mg protein). These changes were not gender-related and consistent with a rise of ANP mRNA. Diethylstilbestrol treatment of immature rats increased hypothalamic ANP concentration from 2.11 +/- 0.24 to 2.97 +/- 0.44 ng/mg protein (P<0.001), but equine chorionic gonadotropin had no effect, indicating that estrogen is a potential stimulus of ANP only at supra-physiological concentrations. CNP, the most abundant natriuretic peptide in the brain, gradually increased in the developing hypothalamus, but did not plateau at PD 20. Reverse transcription-polymerase chain reaction analysis of ANP receptor mRNA demonstrated higher guanylyl cyclase (GC) A, no changes in GC-B, and lower C receptor levels in adult compared to newborn rats. In conclusion, we have shown that hypothalamic ANP undergoes a dramatic rise after birth, and progresses further until the 3rd postnatal week. ANP and CNP changes in the developing hypothalamus can influence brain maturation. PMID- 15283994 TI - Developmental expression of rat torsinA transcript and protein. AB - A GAG deletion in the gene (TOR1A) for torsinA is associated with childhood-onset generalized dystonia (DYT1). Environmental factors may contribute to development of the phenotype since mutations in TOR1A are clinically penetrant in less than 40% of cases. Median age of onset is 10 and appearance of dystonia after 28 is rare. As a step towards understanding the temporal window of DYT1 disease penetrance, we have examined torsinA transcript and protein expression in rats from the embryonic period through adulthood. With relative quantitative multiplex real-time RT-PCR, we detected torsinA transcript in both neural (cerebellar cortex, striatum, cerebral cortex, thalamus and hippocampus) and non-neural (liver, kidney and heart) tissues at each developmental time point tested (embryonic day 20 [E20], postnatal day 1 [P1], P7, P14, P36, 6 months, 1.5 years). Levels of torsinA transcript were highest at E20 or P1 in all tissues examined except for the cerebellum where transcript levels peaked at P14. Early postnatal levels of torsinA transcript were over three times higher than those seen in adult rats. With quantitative radioactive in situ hybridization, torsinA transcript was widely distributed in brain at all ages with levels peaking at P14 in both cerebellum and striatum. TorsinA-immunoreactivity (IR) was present in neurons throughout the brain. TorsinA-IR was detected in perikarya, dendrites and axons but not nuclei. At P14, prominent expression of torsinA was noted in both striatal cholinergic interneurons and cerebellar Purkinje cells. Our results suggest that torsinA may contribute to postnatal maturational events in the brain such as dendritic arborization and synaptogenesis. Furthermore, the time course of torsinA expression in discrete components of motor networks is compatible with the temporal window of clinical penetrance in DYT1 mutation carriers. PMID- 15283995 TI - Glutamate receptor subunit expression after spinal cord injury in young rats. AB - To investigate the possibility that glutamate receptor levels in the spinal cord are altered following injury to young rats, we used a previously characterized model of spinal cord contusion that produces a reliable injury in rats at postnatal day 14-15. Quantitative Western blot analysis was used to measure relative amounts of protein for several glutamate receptor subunits acutely (24 h) and chronically (28 days) after spinal cord injury (SCI). Acutely after injury significant decreases were observed in the GluR1, GluR2, and GluR4 subunits of the alpha-amino-3-hydroxy-5-methyl-4-isoxazole proprionate (AMPA) receptor, and the NR2A and NR2B subunits, but not the NR1 subunit, of the N-methyl-d-aspartate (NMDA) receptor. However, 28 days after injury only one subunit (GluR4) was shown to be altered. These widespread changes that occur acutely in receptor subunit expression may be an attempt to protect cells from glutamate-induced death. The injured spinal cord in these young animals, however, appears to have the capacity to regulate receptor subunit levels to normal within a month of injury. PMID- 15283996 TI - Hemorrhagic hydrocephalus (hhy): a novel mutation on mouse chromosome 12. AB - A novel mouse hemorrhagic hydrocephalus mutation (hhy) inherited in an autosomal recessive manner on chromosome 12 has been found at the Osaka Prefecture University. The hhy homozygous mutant had dilated lateral ventricles and a communicating aqueduct, with no histological abnormalities either in the subarachnoid space or in the choroid plexus. Multiple hemorrhages in the meninges and throughout the brain parenchyma of the mutant were relevant to advanced stages of hydrocephalus. Subcortical heterotopia was detected unexceptionally in the mutants. Thus, the hhy mutation is characterized by three different abnormalities, i.e. hydrocephalus, intracranial hemorrhage and subcortical heterotopia. PMID- 15283997 TI - Maternal separation suppresses TGF alpha mRNA expression in the prefrontal cortex of male and female neonatal C57BL/6 mice. AB - Male C57BL/6 mice that undergo maternal separation (MS) early in life demonstrate higher levels of anxiety upon reaching adulthood compared to normally reared offspring. This study reports that neonatal males and females that undergo MS have reduced mRNA levels of transforming growth factor-alpha (TGF alpha) in the prefrontal cortex, an area of the brain implicated in emotionality, compared to normally reared animals. TGF alpha expression was unaffected by MS in the hippocampus. These data indicate that MS leads to a brain region-specific suppression of TGF alpha expression early in development. PMID- 15283998 TI - Effects of photoperiod on the development of the central glutamate system in tilapia, Oreochromis mossambicus. AB - The effect of photoperiod (light/dark cycle) on the development of the central glutamate system was investigated in tilapia, Oreochromis mossambicus. Tilapia, at 0, 5, and 10 days posthatching were respectively divided into three equal groups to be kept in different photoperiods: 12/12 h, 24/0 h (full day), and 0/24 h (full night). Neither the full-day nor the full-night photoperiod showed any influence on the development of the central glutamate system, including glutamate content and mRNA expression of glutamate receptor 3 alpha, in the developing tilapia brain. These results suggest that neither constant light nor dark photoperiods affected the influence of the central glutamate system on brain sex differentiation in tilapia during the early developing period. PMID- 15283999 TI - Ovarian expression of chemokines and their receptors. AB - Recent studies suggest involvement of the immune system, including leukocytes and cytokines/chemokines, in various ovarian functions such as ovulation. Using the RT-PCR method, we examined expression of various chemokines and their receptors in normal mouse ovaries. Among seventeen examined chemokines (17 CC types and two CXC types), expressions of CC types MCP-1 and RANTES, and CXC type IP-10 were detected at high levels, while most CC types expressed at variable or low levels. Only five chemokines were not detected in the ovary. We next examined expression of chemokine receptors. CCR1 and CCR2, which are the receptors for MCP-1 and RANTES, were also expressed at constitutively high levels while others were not detectable. We further showed that a significant part of expression of both detected chemokines and receptors originated from peripheral blood leukocytes (PBL) circulating in the ovary. However, ovarian tissue was the major contributor of expression. Constitutive expression of several chemokines and their receptors suggests frequent migrations/movements of leukocytes in the ovary, which may be involved in ovarian functions other than ovulation. PMID- 15284000 TI - Expression and localization of cell adhesion molecules in human fetal membranes during parturition. AB - There is increasing evidence to support the view that human parturition represents an inflammatory process. We have previously demonstrated that parturition is associated with leukocyte invasion and pro-inflammatory cytokine production in the cervix and myometrium. Furthermore, we have shown that several cell adhesion molecules are upregulated in these tissues during labor. In fetal membranes, previous studies have shown intercellular adhesion molecule-1 (ICAM-1) upregulation in association with labor. The role of other adhesion molecules has not been explored. The aims of this study were, therefore, to determine the expression of ICAM-1, platelet endothelial cell adhesion molecule (PECAM), vascular cell adhesion molecule (VCAM) and E-selectin in pre- and post-laboring amnion and choriodecidua and to identify cell types responsible for their expression. Biopsies of fetal membranes were obtained from pregnant women delivered by caesarean section before the onset of labor (n = 8) and following spontaneous vaginal delivery (n = 8). Cell adhesion molecules were identified using immunohistochemistry and messenger RNA expression quantified using Northern analysis. We found that following labor, ICAM-1 mRNA expression was significantly upregulated in amnion and choriodecidua (P < 0.05). PECAM mRNA expression was also increased in choriodecidua (P < 0.05). The main cell types responsible for adhesion molecule expression were leukocytes, amniotic epithelial cells and endothelial cells. The upregulation of ICAM-1 and PECAM mRNA expression in fetal membranes following labor provides further evidence that fetal membranes play an important role in the inflammatory process of parturition. PMID- 15284001 TI - Homocysteine levels in women with antiphospholipid syndrome and normal fertile controls. AB - OBJECTIVE: Recent studies have identified hyperhomocysteinemia as a risk factor for both recurrent pregnancy loss and thrombosis. Antiphospholipid syndrome, an autoimmune disorder, is also characterized by recurrent pregnancy loss and thrombosis. Thus, our purpose was to determine if hyperhomocysteinemia is more common in patients with APS than normal fertile controls. METHODS: Plasma, sera and whole blood were obtained from two groups of women: (1) 22 with well characterized antiphospholipid syndrome; and (2) 41 healthy fertile controls. Levels of fasting homocysteine, vitamin B16, vitamin B12, folate and the incidence of the C677/T mutation of the methylene tetrahydrofolate reductase genotype (C677T/MTHFR) were determined. RESULTS: The proportion of individuals with hyperhomocysteinemia and fasting plasma homocysteine levels were similar in women with APS and controls. Levels of vitamin B6, vitamin B12, folate and the incidence of C677/MTHFR were also similar in the two groups. CONCLUSION: Hyperhomocyteinemia and the C677T/MTHFR mutation are not common in women with antiphospholipid syndrome. Abnormal homocysteine metabolism is unlikely to play a major role in the pathogenesis of antiphospholipid syndrome. PMID- 15284002 TI - Immune effects of surgical menopause and estrogen replacement therapy in peri menopausal women. AB - The complex relationship between sex hormones and immune function suggests that sex hormone deficiency and estrogen replacement therapy (ERT) in post-menopausal women may have pleiotropic effects on immune function. For this reason, we aimed to investigate short-term effects of surgical menopause and ERT on immunity profile in peri-menopausal women. Seventeen healthy peri-menopausal women who were to undergo total abdominal hysterectomy and bilateral salpingo-oopherectomy (TAH + BSO) for uterine myoma were enrolled into this study. Three blood samples were collected from each patient: 1-day prior to surgery, 30 days after the operation (before ERT) and 30 days after transdermal ERT. The percentages of peripheral blood lymphocyte subpopulations, serum interleukin-4 (IL-4) and interferon-gamma (IFN-gamma) concentrations were determined by flow-cytometry and ELISA, respectively. Following TAH + BSO, the percentage of CD8+ cells was increased ( P < 0.001 ) while the percentage of CD19+ cells, serum IL-4, and IFN gamma concentration and the ratio of CD4+ to CD8+ cells were decreased ( P < 0.001, P < 0.001, P < 0.002, and P < 0.05 respectively). After ERT, this trend reversed and a decrease in the CD8+ cells ( P < 0.001 ), increase in the CD19+ cells percentages ( P < 0.02 ) and increase in serum IFN-gamma concentration ( P < 0.002 ) were observed. Although an increasing trend in the CD4+ to CD8+ ratio occurred by ERT, this was not significant. However, the decrease in the serum IL 4 concentration after TAH + BSO was not reversed by ERT. Hormone deficiency in post-menopausal women may cause an impaired immune response, and ERT can restore this phenomenon. Estrogen seems to have an important role in the regulation of immune function. PMID- 15284004 TI - Global gene expression profiling of human endometrial receptivity. AB - Scientific knowledge on the molecular changes that occur during the window of implantation is fundamental for the understanding of human reproduction. To gain a global molecular understanding of human endometrial receptivity, we have compared gene expression profiles of pre-receptive (day LH + 2) versus receptive (LH + 7) in well characterized human endometrial biopsies. The samples were analyzed using the Affymetrix HG-95A array, a high density oligonucleotide microarray comprising more than 12,000 genes. In this work, we present part of our results and a comparison with similar works published in the literature. Identified genes include not only genes previously documented to be involved in implantation but also genes for which a role in endometrial receptivity, or even endometrial expression, has not been previously described. Collectively, these studies identify new candidate markers that may be used to diagnose unequivocally the receptive endometrium. PMID- 15284005 TI - The molecular signature of polycystic ovary syndrome (PCOS) theca cells defined by gene expression profiling. AB - Polycystic ovary syndrome (PCOS) is characterized by increased ovarian androgen secretion, anovulatory infertility due to arrested folliculogenesis, and is frequently found in association with insulin resistance and obesity. Characterization of PCOS theca cells demonstrated that elevated expression of the steroidogenic enzymes 17alpha hydroxylase/17,20 lyase (CYP17) and P450 side chain cleavage enzyme (CYP11A1) play a role in increased androgen production by 3beta hydroxysteroid dehydrogenase in the PCOS theca cell. However, the gene networks and signal transduction pathways which cause the altered expansion of the steroid enzymes remain to be determined. In order to identify these gene networks and/or signaling pathways, we carried out global gene expression profiling of normal and PCOS theca cells using subtractive suppressive hybridization and oligonucleotide microarray analysis. These analyses demonstrated that approximately 2% of genes expressed in the theca cell exhibit altered mRNA abundance in PCOS. Characterization of these genes revealed that retinoic acid synthesis and Wnt signal transduction are altered in the PCOS theca cell. In addition, the transcription factor GATA6, which regulates the promoter activity of CYP17 and CYP11A, was increased in the PCOS compared to normal theca cells. Thus, global gene expression profiling has identified potential pathways which may determine the PCOS theca cell phenotype. PMID- 15284006 TI - Proteomics in reproductive medicine: the technology for separation and identification of proteins. AB - With the near completion of the human genome project, reproductive biology is poised to enter the vastly more complex arena of proteomics. Proteomics involves the identification, characterization and quantitation of all proteins present in a cell at a particular metabolic state. Although the number of genes in the human genome is estimated to be about 27,000 +/- 5000, the number of proteins produced by humans is unknown, with estimates ranging as high as 1,500,000 distinct molecular entities. In order to address problems in the early stages of reproduction, proteomics must be scaled down to work with very few numbers of cells, termed zeptoproteomics. Mass spectrometry has rapidly become the key technology in proteomics, enabling rapid and facile identification and quantitation of femtomole and attomole quantities of a protein. PMID- 15284007 TI - Bioinformatics in reproductive biology--functional annotation based on comparative sequence analysis. AB - Recent studies of the genomes of a variety of model organisms have provided an unprecedented opportunity to identify and characterize all signaling molecules in the human genome. Regardless of the approaches used to decipher gene characteristics and their role in physiology, pairwise sequence comparison represents the fundamental bioinformatic tool for initial functional annotation of newly identified genes. Because genes evolved from duplication and adapted to different evolutionary niches for each organism during speciation, detailed sequence analysis could provide additional information on the biochemical and biological characteristics of novel genes. In addition, the integration of sequence-based gene discovery with phylogeny-based function prediction leads to a more complete understanding of the signaling pathways. For example, detailed pairwise sequence analysis has led to the identification of (1) stresscopin (SCP) and stresscopin-related peptides (SRP) as the CRH-related genes, (2) multiple relaxin-like factor genes, and (3) the novel glycoprotein hormone subunit family genes, alpha2 and beta5. Furthermore, based on the understanding that ligands and receptors coevolved during evolution, we have identified a variety of novel extracellular signaling polypeptides including (1) stresscopin and stresscopin related peptides as selective ligands for the type 2 CRH receptor, (2) the pregnancy hormone, relaxin, and related peptides that activate two orphan G protein-coupled receptors (GPCRs), LGR7 and LGR8, and (3) alpha2 and beta5 that form a heterodimer capable of activating the TSH receptor. Thus, detailed studies on the characteristics and evolution of gene sequences have provided an inroad to the elucidation of novel signaling polypeptides and the associated signal transduction pathways. PMID- 15284008 TI - Cardiovascular effects of the toxin(s) of the Australian paralysis tick, Ixodes holocyclus, in the rat. AB - An extract of toxin(s) from the Australian paralysis tick, Ixodes holocyclus, produced positive inotropic responses in rat left ventricular papillary muscles and positive contractile responses in rat thoracic aortic rings. There was no measurable chronotropic response in rat right atria, but positive inotropic concentrations in papillary muscles produced arrhythmias in right atria. Positive inotropic responses were attenuated by verapamil, but unaffected by metoprolol, cimetidine, pyrilamine, tetrodotoxin and pinacidil. Microelectrode studies on isolated left ventricular papillary muscles demonstrated that the extract prolonged action potential duration at 20, 50 and 90% of repolarisation and delayed ventricular papillary muscle relaxation. Cardiovascular tissues isolated from rats with experimentally induced tick paralysis showed no myocardial damage as identified by histological and ultrastructural examination. The basal rate and force of contraction of isolated cardiac tissues were lower from tick-paralysed than normal rats. Concentration-response curves to dobutamine and calcium chloride were similar between tissues from tick-paralysed and normal rats. Thus, the Australian paralysis tick, I. holocyclus, produces one or more toxins with direct cardiovascular effects which mimic the effects produced by direct blockade of cardiac and vascular K+ channels. PMID- 15284009 TI - Identification of crotasin, a crotamine-related gene of Crotalus durissus terrificus. AB - Crotamine is a cationic peptide (4.9 kDa, pI 9.5) of South American rattlesnake, Crotalus durissus terrificus' venom. Its presence varies according to the subspecies or the geographical locality of a given species. At the genomic level, we observed the presence of 1.8 kb gene, Crt-p1, in crotamine-positive specimens and its absence in crotamine-negative ones. In this work, we described a crotamine-related 2.5 kb gene, crotasin (Cts-p2), isolated from crotamine negative specimens. Reverse transcription coupled to polymerase chain reaction indicates that Cts-p2 is abundantly expressed in several snake tissues, but scarcely expressed in the venom gland. The genome of crotamine-positive specimen contains both Crt-p1 and Cts-p2 genes. The present data suggest that both crotamine and crotasin have evolved by duplication of a common ancestor gene, and the conservation of their three disulfide bonds indicates that they might adopt the same fold as beta-defensin. The physiological function of the crotasin is not yet known. PMID- 15284010 TI - Effects of Lonomia obliqua (lepidoptera, saturniidae) toxin on clotting, inflammatory and antibody responsiveness in genetically selected lines of mice. AB - Lines of mice genetically selected for high (H) or low (L) antibody response and for maximal (AIRMAX) or minimal (AIRMIN) acute inflammatory reaction, in which the opposite extreme potentialities have been clearly defined, offer an appropriate model for investigating the environmental and genetic factors acting on innate and adaptative immunobiological functions. This model has been successfully employed to study the resistance or susceptibility against pathogens and/or toxins. It had been demonstrated that the skin contact with Lonomia obliqua caterpillar bristles induces local inflammation and may elicit severe hemorrhagic disorders. In the present study, blood coagulation time, and the acute inflammatory reaction were scored 24 h after injection of the Lonomia bristles crude extract in a subcutaneous dorsal air pouch. The acute inflammation was determined by the leukocyte concentration in the local exudates. The highest interline differences were observed between the AIRMAX (10(6) cells/ml) and AIRMIN (2 x 10(5) cells/ml) and this distinct expression involves the number of monocytes, eosinophils and mainly neutrophils. Regarding coagulation, the highest interline difference was observed between the HIII and LIII mice, and the F1)[LIII x HIII] hybrids showed the overdominance of the fast clotting character. The adaptative immune response was evaluated by comparing the anti-Lonomia bristle extract IgG titer among the lines: the antibody titers were higher in the H lines than in the L ones and equivalent in the AIRMAX and AIRMIN mice, in accordance to the phenotype profiles generated by the distinct selective processes. The genetically selected mice lines-AIRMAX, AIRMIN, HI, HIII, HG, LIII and LG-showed an almost continuous distributions for inflammation, coagulation time and IgG antibody titers, being the interline variances always higher than the intraline ones for the individually measured phenotypes. Altogether, these results suggest the independent polygenic regulation of these traits, being indicative of the genetic control to Lonomia toxin innate and adaptative sensitivity in humans. PMID- 15284011 TI - Inhibitory effects of human alpha2-macroglobulin and mouse serum on the PSGL-1 and glycoprotein Ib proteolysis by a snake venom metalloproteinase, triflamp. AB - Triflamp, a 28 kDa snake venom metalloproteinase purified from the venom of Trimeresurus flavoviridis, possesses the proteolytic activities toward P-selectin glycoprotein ligand-1 (PSGL-1), glycoprotein Ib (GPIb) and fibrinogen. In human whole blood preparation, however, triflamp (6 microg/ml) failed to cleave neutrophil PSGL-1 and platelet GPIb. Human alpha2-macroglobulin (alpha2M) was mainly responsible for the neutralization of the proteolytic effects of triflamp on PSGL-1, GPIb and fibrinogen. Human alpha2M neutralized triflamp at a stoichiometry about 1.1:1 (molar basis) determined by azocaseinolysis. SDS-PAGE analysis revealed that triflamp cleaved the bait-region of alpha2M. Western blot demonstrated that triflamp interacted with the C-terminal half-subunits of truncated alpha2M resulting in the formation of high-molecular-weight species of alpha2M-triflamp complexes. In the presence of competing nucleophile, 0.2 M methylamine, the proteolytic activity of triflamp was conserved. In vivo we found that mice neutrophils were resistant to the cleavage of PSGL-1 by triflamp. However, mouse PSGL-1 and GPIb were susceptible to be cleaved by triflamp in washed mouse neutrophil and platelet preparation, respectively. Similarly, mouse serum was also responsible for the inactivation of the proteolytic activity of triflamp. This study provides direct evidences for the reasonable explanation regarding the reduced proteolytic activity of triflamp toward its substrates in whole blood preparation and in vivo model. PMID- 15284012 TI - Comparative concentrations of brevetoxins PbTx-2, PbTx-3, BTX-B1 and BTX-B5 in cockle, Austrovenus stutchburyi, greenshell mussel, Perna canaliculus, and Pacific oyster, Crassostrea gigas, involved neurotoxic shellfish poisoning in New Zealand. AB - Previously, we found brevetoxins PbTx-3, BTX-B5 and BTX-B1 in cockle, Austrovenus (A.) stutchburyi, PbTx-2, PbTx-3 and BTX-B1 in Pacific oyster, Crassostrea (C.) gigas and PbTx-3 and BTX-B1 in greenshell mussel, Perna (P.) canaliculus following outbreak of neurotoxic shellfish poisoning (NSP) in New Zealand by isolation and/or liquid chromatography coupled with tandem mass spectrometry (LC MS/MS). In this study, procedures for quantitative determination of PbTx-2 and BTX-B5 were developed and those for PbTx-3 and BTX-B1 were further examined by LC MS/MS. In mass spectrometry with an electrospray ionization interface operating in the positive or negative ion mode, the protonated ions [M+H]+ of PbTx-2 (m/z 895), [M+H]+ of PbTx-3 (m/z 897), [M-H]- of BTX-B5 (m/z 909), and [M-Na]- of BTX B1 (m/z 1016) were generated abundantly, when 0.1% formic acid-acetonitrile was used as the mobile phase for column chromatography. The product ions of m/z 877, 725, 111 and 80 from PbTx-2, PbTx-3, BTX-B5 and BTX-B1 were identified, respectively, allowing unambiguous confirmation of these toxins by selective reaction monitoring LC-MS/MS analysis. High levels of PbTx-3 and BTX-B5 were detected in C. gigas, of PbTx-3, BTX-B1 and BTX-B5 in A. stutchburyi, and of PbTx 2, PbTx-3 and BTX-B5 in P. canaliculus by this LC-MS/MS method. PMID- 15284013 TI - Neutralization of venoms from two Southern Pacific Rattlesnakes (Crotalus helleri) with commercial antivenoms and endothermic animal sera. AB - The Southern Pacific Rattlesnake (Crotalus helleri) is found in southwestern California (USA), southward through north Baja California (MX) into the northern part of southern Baja California (MX). In this study, the venoms from two Southern Pacific Rattlesnakes were characterized. The two venoms were different in color, concentration, and enzyme activities. Two commercial antivenoms neutralized both C. helleri venoms differently. Antivipmyn (Fab2H) and CroFab (FabO) neutralized both venoms but had different ED50. Four times more Fab2H antivenom was required to neutralize the C. helleri venom No. 011-084-009 than the venom from the snake No. 010-367-284. The hemorrhagic activity of two C. helleri venoms were neutralized differently by endothermic animal sera having a natural resistance to hemorrhagic activity of snake venoms. Opossums and Mexican ground squirrel sera did not neutralize the hemorrhagic activity of the venom No. 010-367-284. The sera of gray woodrats and hispid cotton rats neutralized all hemorrhagins in both C. helleri venoms. This is the first reported case in which opossum serum has not neutralized hemorrhagic activity of pit viper venom. Differences in the compositions of C. helleri venoms and their ability to be neutralized may help explain why snakebites are a difficult medical problem to treat and why effective polyvalent antivenoms are difficult to produce. PMID- 15284014 TI - A comparative study of biological activities of crotoxin and CB fraction of venoms from Crotalus durissus terrificus, Crotalus durissus cascavella and Crotalus durissus collilineatus. AB - In Brazil, the Crotalus durissus terrificus subspecie is the most studied, particularly concerning its crotoxin. Crotoxin is the major toxic component of the South American rattlesnake Crotalus durissus venom. It is composed of two different subunits, CA called crotapotin and CB weakly toxic phospholipase A2 with high enzymatic activity. In this paper, we decided to make a study of the main toxic characteristics of crotoxin (CTX) and CB fraction from the other subspecies, Crotalus durissus cascavella and of Crotalus durissus collilineatus, in comparison with those of C. d. terrificus. Ours results have shown that the venoms presented similar chromatographic profiles and the purified fractions were free of contaminants. Regarding the toxic activities, the DL50 of the crotoxins showed no significant differences between the subspecies. The smaller toxicity of CB indicated that the toxicity of the crotoxin complex depends on the interaction between CA and CB. CTX and fraction CB of the three species of Crotalus showed negligible proteolytic activity. C. d. terrificus CTX presented higher PLA2 activity when compared with the others two subspecies. The oedema induced by CB developed later than the CTX and reached its peak 3 h after the injection. The myotoxic activity was determined by assaying serum CK levels. Mice injected with CTX of C. d. terrificus presented greater myotoxic activity compared to the others. The myotoxic activity of CB from the three subspecies was lower than the activity of the crotoxin, reinforcing the idea that the fraction CA increases the toxicity of CB. PMID- 15284015 TI - Efficacy of two different doses of antilonomic serum in the resolution of hemorrhagic syndrome resulting from envenoming by Lonomia obliqua caterpillars: a randomized controlled trial. AB - The hemorrhagic syndrome caused by contact with Lonomia obliqua caterpillars has reached epidemic proportions in southern Brazil. This study aimed at assessing the efficacy of two different dosages of an antilonomic serum (SALon) in the treatment of patients who had accidental contact with L. obliqua caterpillars. A randomized, prospective controlled trial was conducted at Sao Vicente de Paulo Hospital, a tertiary hospital in Passo Fundo, Brazil. From January 2000 to April 2002, 44 patients with grade I or II hemorrhagic syndrome were randomly assigned to either group A: 22 patients treated with 10.5 mg total dose (three vials of SALon) or group B: 22 patients treated with 17.5 mg total dose (five vials of SALon). Treatment efficacy was assessed according to time necessary for blood coagulation to go back to normal, incidence of adverse reactions, and hospitalization time. There were no differences in clinical picture and laboratory findings between the two groups. Mean time for coagulation to go back to normal was 15.3+/-6.6 and 19.1+/-8 h in groups A and B, respectively. Adverse reactions and hospitalization time were similar in both groups. SALon was effective and well tolerated. This study demonstrated that three vials are as effective as five vials of SALon for the treatment of envenoming by Lonomia caterpillars. PMID- 15284016 TI - In vitro neuromuscular activity of 'colubrid' venoms: clinical and evolutionary implications. AB - In this study, venoms from species in the Colubrinae, Homalopsinae, Natricinae, Pseudoxyrhophiinae and Psammophiinae snake families were assayed for activity in the chick biventer cervicis skeletal nerve muscle preparation. Boiga dendrophila, Boiga cynodon, Boiga dendrophila gemincincta, Boiga drapiezii, Boiga irregularis, Boiga nigriceps and Telescopus dhara venoms (10 microg/ml) displayed postsynaptic neuromuscular activity as evidenced by inhibition of indirect (0.1 Hz, 0.2 ms, supramaximal V) twitches. Neostigmine (5 microM) reversed the inhibition caused by B. cynodon venom (10 microg/ml) while the inhibitory effects of Psammophis mossambicus venom (10 microg/ml) spontaneously reversed, indicating a reversible mode of action for both venoms. Trimorphodon biscutatus (10 microg/ml) displayed irreversible presynaptic neurotoxic activity. Detectable levels of phospholipase A2 activity were found only in T. biscutatus, T. dhara and P. mossambicus venoms. The results demonstrate a hitherto unsuspected diversity of pharmacological actions in all lineages which may have implications ranging from clinical management of envenomings to venom evolution. PMID- 15284017 TI - Degradation of microcystin-LR toxin by Fenton and Photo-Fenton processes. AB - This study reports a laboratory investigation of the degradation of microcystin LR using Fenton (Fe2+ + H2O2) and Photo-Fenton processes. The effect of hydrogen peroxide concentration on the Fenton reaction rate was investigated at constant Fe2+ concentrations. It was observed that at low concentrations of hydrogen peroxide (0.25-0.5 mM), the extent of microcystin-LR degradation was low, even after prolonged reaction time (up to 600 min). Higher H2O2 concentrations (2.5-5 mM) resulted in higher degradation rates that yielded microcystin-LR degradation as high as 60% in approximately 180 min. However, the highest degradation efficiency of the toxin was achieved during the Photo-Fenton process in which UV radiation was involved. In the Photo-Fenton process, the removal efficiency of microcystin-LR reached 84% in the first 25 min and 100% in approximately 35-40 min of irradiation. These results are encouraging for the application of efficient UV-based advanced oxidation technologies for toxin removal from drinking water sources. PMID- 15284018 TI - Bothrops venom induces direct renal tubular injury: role for lipid peroxidation and prevention by antivenom. AB - Acute renal failure (ARF) is one of the most serious complications of Bothrops snakebites. Pathogenesis of ARF in snakebite envenomation may involve hemodynamic disturbances, immunologic reactions and direct nephrotoxicity. This study aimed at evaluating Bothrops jararaca venom direct toxicity on isolated rat renal proximal tubules (PT). PT was kept oxygenated and subjected to hypoxia (H, 15 min) and reoxygenation (R, 45 min). Bothropic antivenom effects, role of extracellular calcium and peroxide production were also evaluated. Cell injury was determined by LDH release (%) and peroxide production determined by xylenol orange method. B. jararaca venom caused tubular injury (LDH 31.6 vs. 17.2%, P <0.05), which was prevented by simultaneous or delayed antivenom administration, but not with low extracellular calcium medium. Venom increased tubules peroxide production: 2.21 vs. 1.27 microM/mg protein (P <0.05) which was also prevented by antivenom administration. Venom toxic concentration did not enhance H/R injury. In contrast, non-toxic venom concentration afforded protection (LDH 41.3 vs. 51.5%, P <0.05). In conclusion, B. jararaca venom caused direct injury to normoxic renal tubules, but not to hypoxic/reoxygenated tubules. Tubular toxicity is independent of extracellular calcium and mediated in part by lipid peroxidation. Venom induced tubular injury was prevented by simultaneous or delayed antivenom administration. PMID- 15284019 TI - Extracts from two marine sponges lower cyclin B1 levels, cause a G2/M cell cycle block and trigger apoptosis in SW-13 human adrenal carcinoma cells. AB - Marine sponges have been shown to produce metabolites with cell growth- and endocrine-altering activities. We tested extracts from two species: the 'brown variable sponge' (Anthosigmella varians) and the 'West Indian bath sponge' (Spongia barbara), for effects on the cell cycle regulatory protein, cyclin B1; cell cycle growth-phase (sub-G1/apoptosis, G1, S, and G2/M); and cell survival in SW-13 human adrenal carcinoma cultures. Polyacrylamide gel electrophoresis studies indicated a 70-90% reduction in cyclin B1 levels by treatment with these agents. Microscopic examination of cultures with DAPI staining showed dense and fragmented DNA fluorescence, characteristic of apoptosis, in both sponge extract treated cultures but not in controls. Flow cytometry analysis showed a 16-fold increase in the percentage of cells entering apoptosis (sub-G1 phase of cell cycle) by treatment with Anthosigmella varians extract (p <0.01) and a 10-fold increase using Spongia barbara extract (p <0.01) During this same time, the percentage of cells in G2/M was increased 1.6-fold by Anthosigmella varians extract (p <0.01) and 2.0-fold by Spongia barbara extract (p <0.01) Cell growth/survival studies also indicated a time-dependent decline in the percentage confluence of cell cultures exposed to Anthosigmella varians or Spongia barbara extracts. These experiments demonstrate that some species of marine sponges have metabolites which are capable of interfering with the mammalian cell cycle and with the survival of human adrenal carcinoma cells in culture. PMID- 15284020 TI - Effects of a myotoxic Asp49 phospholipase A2 (ACL-I PLA2) isolated from Agkistrodon contortrix laticinctus snake venom on water transport in the isolated toad urinary bladder. AB - An Asp49 PLA2 (ACL-I PLA2) was purified from the venom of Agkistrodon contortrix laticinctus by gel filtration and cation-exchange chromatography. It has a relative molecular mass of 14,000, and its N-terminal sequence has more than 65% of identity with other snake venom PLA2s. ACL-I PLA2 injected into the Tibialis anterior muscle of rats and mice at doses of 0.3 and 1.6 mg/kg, respectively, induced muscle fiber necrosis, cellular infiltration and edema 3 and 48 h after injection. The effect of the purified enzyme on water permeability was tested in the isolated toad urinary bladder. Water flow through the membrane was measured gravimetrically in bag preparations of the bladder. ACL-I PLA2 (20 nM) did not significantly alter the water permeability in the bladder preparations, whereas ACL myotoxin (ACLMT), a Lys49 PLA2 isolated from the same venom, at similar concentration significantly increased (81%) the water permeability. However, both toxins inhibited the AVP-stimulated water permeability. These results strongly suggest that PLA2 activity is not involved in the ACLMT effect on water transport and the effect of ACL-I PLA2 myotoxin on membrane permeability is mediated by mechanisms that are different in comparison to ACLMT. PMID- 15284021 TI - Complete amino acid sequence and phylogenetic analysis of a long-chain neurotoxin from the venom of the African banded water cobra, Boulengerina annulata. AB - The sequence of a long-chain neurotoxin (71 amino acid residues, 10 half cystines) from the venom of the African banded water cobra (Boulengerina annulata) was determined by Edman degradation. It exhibits high sequence similarity with long-chain neurotoxins from the venoms of four species of African cobras (genus Naja), which are collectively more similar to the Boulengerina toxin than to those of Asian Naja species. These results are discussed in the light of phylogenetic hypotheses on the relationships of African cobras. PMID- 15284022 TI - Metalloprotease disintegrin-mediated ectodomain shedding of EGFR ligands promotes intestinal epithelial restitution. AB - EGF receptor (EGFR) promotes intestinal epithelial restitution, an important early process in the reepithelialization of ulcers. During epithelial restitution, the mechanism of EGFR activation is not known. We evaluated the role of TNF-converting enzyme (TACE), a metalloprotease disintegrin that proteolytically processes plasma membrane-anchored EGFR ligand precursors into their mature active forms, in wound-induced EGFR activation and epithelial restitution. With the use of scrape-wounded rat intestinal epithelial-1 (RIE-1) cell monolayers to model epithelial ulceration and restitution, we observed the rapid wound-dependent release of EGFR ligands into culture medium. RIE-1 cells express TACE, and treatment with phorbol ester, an established TACE stimulus, triggered the extracellular release of an EGFR ligand, transforming growth factor alpha. Blockade of TACE using TNF processing inhibitor (TAPI-1), a specific hydroxamate inhibitor of metalloprotease disintegrins, prevented release of EGFR ligands from wounded RIE-1 cell monolayers. The restitution of wounded RIE-1 cell monolayers was also dose-dependently inhibited by TAPI-1, establishing the role of metalloprotease disintegrins in this process. These results have established a mechanism of EGFR activation in wounded intestinal epithelium and show an important functional role for metalloprotease disintegrin-mediated ectodomain shedding during intestinal epithelial restitution. Therefore, activation of the TACE-EGFR system might promote the healing of intestinal tract ulcers in patients. PMID- 15284023 TI - Electroneutral sodium absorption and electrogenic anion secretion across murine small intestine are regulated in parallel. AB - Electrolyte transport processes of small intestinal epithelia maintain a balance between hydration of the luminal contents and systemic fluid homeostasis. Under basal conditions, electroneutral Na(+) absorption mediated by Na(+)/H(+) exchanger 3 (NHE3) predominates; under stimulated conditions, increased anion secretion mediated by CFTR occurs concurrently with inhibition of Na(+) absorption. Homeostatic adjustments to diseases that chronically affect the activity of one transporter (e.g., cystic fibrosis) may include adaptations in the opposing transport process to prevent enterosystemic fluid imbalance. To test this hypothesis, we measured electrogenic anion secretion (indexed by the short circuit current) across NHE3-null [NHE3(-)] murine small intestine and electroneutral Na(+) absorption (by radioisotopic flux analysis) across small intestine of mice with gene-targeted disruptions of the anion secretory pathway, i.e., CFTR-null [CFTR(-)] or Na(+)-K(+)-2Cl(-) cotransporter-null [NKCC1(-)]. Protein expression of NHE3 and CFTR in the intestinal epithelia was measured by immunoblotting. In NHE3(-), compared with wild-type small intestine, maximal and bumetanide-sensitive anion secretion following cAMP stimulation was significantly reduced, and there was a corresponding decrease in CFTR protein expression. In CFTR(-) and NKCC1(-) intestine, Na(+) absorption was significantly reduced compared with wild-type. NHE3 protein expression was decreased in the CFTR(-) intestine but was unchanged in the NKCC1(-) intestine, indicating that factors independent of expression also downregulate NHE3 activity. Together, these data support the concept that absorptive and secretory processes determining NaCl and water movement across the intestinal epithelium are regulated in parallel to maintain balance between the systemic fluid volume and hydration of the luminal contents. PMID- 15284024 TI - EGFR-induced cell migration is mediated predominantly by the JAK-STAT pathway in primary esophageal keratinocytes. AB - The epidermal growth factor receptor (EGFR) activates several signaling cascades in response to epidermal growth factor stimulation. One of these signaling events involves tyrosine phosphorylation of signal transducer and activator of transcription (STAT), whereas another involves activation of the phosphatidylinositol 3-OH kinase pathway. Two possibilities for STAT activation exist: a janus kinase (JAK)-dependent and a JAK-independent mechanism. Herein, we demonstrate that EGFR overexpression in primary esophageal keratinocytes activates STAT in a JAK-dependent fashion with the functional consequence of enhanced cell migration, which can be abolished by use of a JAK-specific inhibitor, AG-490. We determined the mechanisms underlying the signal transduction pathway responsible for increased cell migration. Stimulation of EGFR induces Tyr701 phosphorylation of STAT1 and initiates complex formation of STAT1 and STAT3 with JAK1 and JAK2. Thereafter, the STATs translocate to the nucleus within 15 min. In addition, we found that activation of this signaling pathway results in matrix metalloproteinase-1 (MMP-1) activity. By contrast, Akt activation does not impact the EGFR-STATs-JAKs complex formation and nuclear translocation of the STATs with subsequent MMP-1 activity, although Akt activation may contribute to cell migration through an independent mechanism. Taken together, we find that the recruitment of the STAT-JAK complex by EGFR is responsible for keratinocyte migration that, in turn, might be mediated by MMP-1 activation. PMID- 15284025 TI - Social networks and peer education. PMID- 15284026 TI - Tobacco harm reduction involves more than cigarette harm reduction. PMID- 15284027 TI - Updating Derryberry's priorities and the role of health education. PMID- 15284028 TI - Access denied, science denied. PMID- 15284029 TI - A pioneer of chemical dependency treatment: Dr Mondanaro takes no prisoners. PMID- 15284030 TI - Tracking gender-based human rights violations in postwar Kosovo. AB - Four years have passed since the institution of the cease-fire in Yugoslavia, and questions remain as to how Kosovar women are faring in the country's postwar reconstruction. Reports, albeit fragmented, suggest that violence against women began to increase in 1998 and 1999. This trend continued through 2001, even while rates of other major crimes decreased. Despite considerable local efforts to address the conditions of women, there remains a lack of systematic data documenting the scope and frequency of violent acts committed against women. A centralized surveillance system focused on tracking human rights abuses needs to be established to address this critical need for empirically based reports and to ultimately guide reform efforts. PMID- 15284031 TI - A cigarette manufacturer and a managed care company collaborate to censor health information targeted at employees. AB - A review of internal tobacco company documents showed that the tobacco company Philip Morris and the insurance company CIGNA collaborated to censor accurate information on the harm of smoking and on environmental tobacco smoke exposure from CIGNA health newsletters sent to employees of Philip Morris and its affiliates. From 1996 to 1998, 5 of the 8 CIGNA newsletters discussed in the internal tobacco documents were censored.We recommend that accrediting bodies mandate that health plans not censor employee-directed health information at the request of employers. PMID- 15284032 TI - The decline in maternal mortality in Sweden: the role of community midwifery. AB - The maternal mortality rate in Sweden in the early 20th century was one third that in the United States. This rate was recognized by American visitors as an achievement of Swedish maternity care, in which highly competent midwives attend home deliveries. The 19th century decline in maternal mortality was largely caused by improvements in obstetric care, but was also helped along by the national health strategy of giving midwives and doctors complementary roles in maternity care, as well as equal involvement in setting public health policy. The 20th century decline in maternal mortality, seen in all Western countries, was made possible by the emergence of modern medicine. However, the contribution of the mobilization of human resources should not be underestimated, nor should key developments in public health policy. PMID- 15284033 TI - The control of rickets. 1925. PMID- 15284035 TI - WIC participation, breastfeeding practices, and well-child care among unmarried, low-income mothers. AB - We estimated the effect of Special Supplemental Nutrition Program for Women, Infants, and Children (WIC) participation in 1999 to 2000 on breastfeeding initiation and duration and well-child care. We applied multivariate regression to a sample of 2136 unmarried, low-income, urban mothers from the Fragile Families and Child Wellbeing Study. WIC participation was associated with small increases in the probabilities of initiating breastfeeding and having had at least 4 well-child visits since birth-behaviors that benefit infants beyond the newborn period-but not with breastfeeding duration. PMID- 15284036 TI - Condom use and the risk of recurrent pelvic inflammatory disease, chronic pelvic pain, or infertility following an episode of pelvic inflammatory disease. AB - Among 684 sexually active women with pelvic inflammatory disease (PID) followed up for a mean of 35 months, we related contraceptive use to self-reported PID recurrence, chronic pelvic pain, and infertility. Persistent use of condoms during the study reduced the risk of recurrent PID, chronic pelvic pain, and infertility. Consistent condom use (about 60% of encounters) at baseline also reduced these risks, after adjustment for confounders, by 30% to 60%. Self reported persistent and consistent condom use was associated with lower rates of PID sequelae. PMID- 15284037 TI - The benefit of health insurance coverage of contraceptives in a population-based sample. AB - This study estimated the value of contraceptives, through a random-digit-dialed survey of willingness to pay for health insurance coverage of contraceptives among 659 Washington State adults. People valued contraceptives at 5 times the actuarial cost; in general, women and reproductive-aged persons were willing to pay more, but low-income men highly valued contraceptives. Most respondents (85%) said that contraceptives should be covered by health insurance plans. The full benefit of contraceptives exceeds their cost. PMID- 15284038 TI - Young males' sexual education and health services. AB - This study examined the basis for one hospital's decision to restructure its teen family planning clinical services. We examined results of surveys conducted from 1998 to 2003 with more than 2000 mostly African American eighth-grade boys. Most young males wanted to postpone sexual intercourse, but an even greater percentage were willing to use a method of protection. The hospital determined that it needed to give the same in-hospital clinical and counseling support to young males as it gives to young females. PMID- 15284039 TI - Do men know that they have had a prostate-specific antigen test? Accuracy of self reports of testing at 2 sites. AB - This study determined the accuracy of self-reports of prostate-specific antigen (PSA) testing. Men (N = 402) attending 2 outpatient clinics were asked: "Did you have a PSA test today?" and their medical records were checked. Concordance, sensitivity, and false-negative values were 65%, 67%, and 33%, respectively, at 1 clinic site and 88%, 64%, and 36% at the other. The accuracy of self-reports of PSA testing should be interpreted with caution. PMID- 15284040 TI - Insurance coverage of smoking cessation treatment for state employees. AB - Public health experts recommend that health insurance include coverage for smoking cessation treatment as an evidence-based strategy to reduce smoking. As employers, states can implement this policy for more than 5 million individuals nationwide. This study identified the extent to which states require smoking cessation treatment insurance coverage for their employees; of 45 states, 29 required coverage for at least 1 US Public Health Service (PHS)-recommended treatment, and only 17 of 45 provided coverage that was fully consistent with PHS recommendations. PMID- 15284041 TI - Expanded state-funded family planning services: estimating pregnancies averted by the Family PACT Program in California, 1997-1998. AB - OBJECTIVES: The California Family Planning, Access, Care, and Treatment Program was implemented in 1997 to provide family planning services for uninsured, low income women and men. We estimated the impact on fertility of providing 500 000 women with contraceptives. METHODS: Paid claims and medical record review data were used to estimate pregnancies averted. Pregnancies women experienced while enrolled in the program and pregnancies they would have experienced given methods used before enrollment were modeled as a Markov process. RESULTS: One year of Family Planning, Access, Care, and Treatment services averted an estimated 108 000 unintended pregnancies that would have resulted in 50 000 unintended births and 41 000 induced abortions. CONCLUSIONS: Providing contraceptives to low income, medically indigent women significantly reduced the number of unintended pregnancies in California. PMID- 15284042 TI - Fertility and parental consent for minors to receive contraceptives. AB - OBJECTIVES: I examined the effect of imposing a requirement for parental consent before minors can receive medical contraceptives. METHODS: Birth and abortions among teens, relative to adults, in a suburban Illinois county that imposed a parental consent requirement in 1998 were compared with births and abortions in nearby counties during the period 1997-2000. RESULTS: The relative proportion of births to women under age 19 years in the county rose significantly compared with nearby counties, whereas the relative proportion of abortions to women under age 20 years declined insignificantly, with a relative increase in the proportion of pregnancies (births and abortions) to young women in the county. CONCLUSIONS: Imposing a parental consent requirement for contraceptives, but not abortions, appears to raise the frequency of pregnancies and births among young women. PMID- 15284043 TI - Physician assistants as providers of surgically induced abortion services. AB - OBJECTIVES: We compared complication rates after surgical abortions performed by physician assistants with rates after abortions performed by physicians. METHODS: A 2-year prospective cohort study of women undergoing surgically induced abortion was conducted. Ninety-one percent of eligible women (1363) were enrolled. RESULTS: Total complication rates were 22.0 per 1000 procedures (95% confidence interval [CI] = 11.9, 39.2) performed by physician assistants and 23.3 per 1000 procedures (95% CI = 14.5, 36.8) performed by physicians (P =.88). The most common complication that occurred during physician assistant-performed procedures was incomplete abortion; during physician-performed procedures the most common complication was infection not requiring hospitalization. A history of pelvic inflammatory disease was associated with an increased risk of total complications (odds ratio = 2.1; 95% CI = 1.1, 4.1). CONCLUSIONS: Surgical abortion services provided by experienced physician assistants were comparable in safety and efficacy to those provided by physicians. PMID- 15284044 TI - Psychosocial factors and preterm birth among African American and White women in central North Carolina. AB - OBJECTIVES: We assessed associations between psychosocial factors and preterm birth, stratified by race in a prospective cohort study. METHODS: We surveyed 1898 women who used university and public health prenatal clinics regarding various psychosocial factors. RESULTS: African Americans were at higher risk of preterm birth if they used distancing from problems as a coping mechanism or reported racial discrimination. Whites were at higher risk if they had high counts of negative life events or were not living with a partner. The association of pregnancy-related anxiety with preterm birth weakened when medical comorbidities were taken into account. No association with preterm birth was found for depression, general social support, or church attendance. CONCLUSIONS: Some associations between psychosocial variables and preterm birth differed by race. PMID- 15284045 TI - Trends in prenatal care use and low birthweight in southeast Brazil. AB - OBJECTIVES: We investigated trends in prenatal care use and its association with low birthweight in a developing country. METHODS: We examined data from 2 southeast Brazilian cohort surveys, 1 conducted in 1978-1979 and the other in 1994. RESULTS: Socioeconomic inequalities in prenatal care use increased during the 15-year period of 1979-1994. Although prenatal care use increases paralleled increases in low birthweight rate during this period, having no prenatal care was associated with higher risk of low birthweight in both surveys. Inadequate prenatal care use was also associated with higher risk of low birthweight in 1978 1979 only. CONCLUSIONS: Increasing low birthweight rates among women who adequately used prenatal care may be causing a bias by reducing the estimates of the effect of inadequate prenatal care use on low birthweight rates. PMID- 15284046 TI - Implications of family income dynamics for women's depressive symptoms during the first 3 years after childbirth. AB - OBJECTIVES: We examined within-person associations between changes in family income and women's depressive symptoms during the first 3 years after childbirth. METHODS: Data were analyzed for 1351 women (mean baseline age = 28.13 years) who participated in the National Institute of Child Health and Human Development Study of Early Child Care. Nineteen percent of these women belonged to an ethnic minority, and 35% were poor at some time during the study. RESULTS: Changes in income and poverty status were significantly associated with changes in depressive symptoms. Effects were greatest for chronically poor women and for women who perceived fewer costs associated with their employment. CONCLUSIONS: Given that women head most poor households in the United States, our findings indicate that reductions in poverty would have mental health benefits for women and families. PMID- 15284047 TI - Racial/ethnic differences in the prevalence of depressive symptoms among middle aged women: The Study of Women's Health Across the Nation (SWAN). AB - OBJECTIVES: We examined racial/ethnic differences in significant depressive symptoms among middle-aged women before and after adjustment for socioeconomic, health-related, and psychosocial characteristics. METHODS: Racial/ethnic differences in unadjusted and adjusted prevalence of significant depressive symptoms (score >/= 16 on the Center for Epidemiologic Studies Depression [CES-D] Scale) were assessed with univariate and multiple logistic regressions. RESULTS: Twenty-four percent of the sample had a CES-D score of 16 or higher. Unadjusted prevalence varied by race/ethnicity (P <.0001). After adjustment for covariates, racial/ethnic differences overall were no longer significant. CONCLUSIONS: Hispanic and African American women had the highest odds, and Chinese and Japanese women had the lowest odds, for a CES-D score of 16 or higher. This variation is in part because of health-related and psychosocial factors that are linked to socioeconomic status. PMID- 15284048 TI - Association between childhood socioeconomic status and coronary heart disease risk among postmenopausal women: findings from the British Women's Heart and Health Study. AB - OBJECTIVES: We assessed the association between childhood socioeconomic status (SES) and coronary heart disease among postmenopausal women. METHODS: We conducted a cross-sectional analysis of 3444 women aged 60 to 79 years. RESULTS: There was an independent linear association between childhood and adult SES and coronary heart disease. The association between childhood SES and coronary heart disease was attenuated when we adjusted for insulin resistance syndrome, adult smoking, physical activity, biomarkers of childhood nutrition, and passive smoking. CONCLUSIONS: The association between adverse childhood SES and coronary heart disease is in part mediated through insulin resistance, which may be influenced by poor childhood nutrition, and in part through the association between childhood SES and adult behavioral risk factors. PMID- 15284049 TI - Breast and cervical cancer screening among Latinas and non-Latina whites. AB - OBJECTIVES: We examined whether Latinas differ from non-Latinas in having undergone recent mammography, clinical breast examination, or Papanicolaou testing, as well as the contribution of sociodemographic and health care variables to screening. METHODS: We used data from the 1991 National Health Interview Survey Health Promotion and Disease Prevention supplement. RESULTS: Latinas were less likely than non-Latina Whites to have undergone mammography (odds ratio [OR] = 0.71; 95% confidence interval [CI] = 0.57, 0.88), but this difference was attenuated when we controlled for socioeconomic factors (OR = 0.90; 95% CI = 0.70, 1.15). Latinas did not differ from Whites on Papanicolaou tests or clinical breast examinations. Quality of and access to health care predicted screening. CONCLUSIONS: Latina ethnicity does not predict breast and cervical cancer screening behavior independent of sociodemographic and structural factors. PMID- 15284050 TI - Quality of care for women undergoing a hysterectomy: effects of insurance and race/ethnicity. AB - OBJECTIVE: We assessed the quality of hospital care for women who underwent a hysterectomy to compare Medicaid-covered women with privately insured women and minority women with White women. METHODS: We evaluated medical decisions, inpatient care, quality of inpatient care, and outcomes. RESULTS: Quality of hospital care was equivalent for Medicaid-covered women compared with privately insured women and for non-Hispanic Black women compared with White women. Medicaid-covered women (40%) and Black women (68%) were more likely to have a complication compared with privately insured women and White women, respectively. CONCLUSIONS: Increased complications after hysterectomy may result in increased economic burdens to Medicaid. Further studies of the racial/ethnic and sociodemographic issues are needed so that disparities may be adequately addressed. PMID- 15284051 TI - Gender differences in physical disability among an elderly cohort. AB - OBJECTIVES: We analyzed the role of sociodemographic factors, chronic-disease risk factors, and health conditions in explaining gender differences in disability among senior citizens. METHODS: We compared 1348 men and women (mean age = 79 years) on overall disability and compared their specific activities of daily living, instrumental activities of daily living (IADL), and mobility limitations. Analysis of covariance adjusted for possible explanatory factors. RESULTS: Women were more likely to report limitations, use of assistance, and a greater degree of disability, particularly among IADL categories. However, these gender differences were largely explained by differences in disability-related health conditions. CONCLUSIONS: Greater prevalence of nonfatal disabling conditions, including fractures, osteoporosis, back problems, osteoarthritis and depression, contributes substantially to greater disability and diminished quality of life among aging women compared with men. PMID- 15284052 TI - Weapons in the lives of battered women. AB - OBJECTIVES: We assessed weapon use in intimate partner violence and perspectives on hypothetical firearm policies. METHODS: We conducted structured in-person interviews with 417 women in 67 battered women's shelters. RESULTS: Words, hands/fists, and feet were the most common weapons used against and by battered women. About one third of the battered women had a firearm in the home. In two thirds of these households, the intimate partner used the gun(s) against the woman, usually threatening to shoot/kill her (71.4%) or to shoot at her (5.1%). Most battered women thought spousal notification/consultation regarding gun purchase would be useful and that a personalized firearm ("smart gun") in the home would make things worse. CONCLUSIONS: A wide range of objects are used as weapons against intimate partners. Firearms, especially handguns, are more common in the homes of battered women than in households in the general population. PMID- 15284053 TI - Gender differences in substance use treatment entry and retention among prisoners with substance use histories. AB - OBJECTIVES: We examined gender similarities and differences in the predictors of substance use treatment entry and of the combination of treatment entry and completion. METHODS: The sample consisted of 2219 male and female program participants. Maximum likelihood probit estimation was used to identify background and attitudinal characteristics predictive of substance use treatment entry and retention. RESULTS: We observed gender similarities and differences in predictors of treatment entry and the combination of treatment entry and completion. Many of the factors that attract individuals to treatment are the same ones that keep individuals in treatment. CONCLUSIONS: Attitudinal predictors namely, motivation to change-showed the greatest consistency between genders and between predictors of treatment entry and predictors of treatment entry and completion. PMID- 15284054 TI - The potential protective effect of youth assets on adolescent alcohol and drug use. AB - OBJECTIVES: We examined the association between adolescent alcohol and drug use and 9 youth assets in a low-income, inner-city population. METHODS: An in-person survey of 1350 adolescents and parents assessed youth assets and risk behaviors. We analyzed data with chi(2) tests and logistic regression analyses. RESULTS: When we controlled for appropriate variables, there were significant positive relationships between several youth assets and nonuse of alcohol and drugs. Furthermore, youths who possessed all of the statistically significant youth assets were 4.44 times more likely to report nonuse of alcohol and 5.41 times more likely to report nonuse of drugs compared with youths who possessed fewer youth assets. CONCLUSIONS: Our study supports the view that specific youth assets may protect youths from alcohol and drug use. PMID- 15284055 TI - Health care system changes and reported musculoskeletal disorders among registered nurses. AB - OBJECTIVES: We evaluated the impact of health care system changes on nurses' health, and we studied reported musculoskeletal disorders associated with these changes. METHODS: This cross-sectional study (n = 1163) defined a musculoskeletal disorder case as moderate pain that lasted at least 1 week or occurred monthly during the past year. Nurses were asked about changes in the health care system in the past year, and responses to 12 changes were summed and were categorized as low, moderate, or high changes. RESULTS: When the changes were summed, the adjusted odds ratios for musculoskeletal disorders for more than 6 versus 0 to 1 changes were (1) neck: 4.45 (95% confidence interval [CI] = 1.97, 10.08), (2) shoulder: 2.63 (95% CI = 1.17, 5.91), and (3) back: 3.42 (95% CI = 1.61, 7.27). CONCLUSIONS: The adverse impact on health caused by the changing health care system must be addressed to prevent further injuries among nurses. PMID- 15284056 TI - Function and response of nursing facilities during community disaster. AB - OBJECTIVES: We sought to describe the role and function of nursing facilities after disaster. METHODS: We surveyed administrators at 144 widely dispersed nursing facilities after the Los Angeles Northridge earthquake. RESULTS: Of the 113 (78%) nursing facilities that responded (11 365 beds), 23 sustained severe damage, 5 closed (625 beds), and 72 lost vital services. Of 87 nursing facilities implementing disaster plans, 56 cited problems that plans did not adequately address, including absent staff, communication problems, and insufficient water and generator fuel. Fifty-nine (52%) reported disaster-related admissions from hospitals, nursing facilities, and community residences. Nursing facilities received limited postdisaster assistance. Five months after the earthquake, only half of inadequate nursing facility disaster plans had been revised. CONCLUSIONS: Despite considerable disaster-related stresses, nursing facilities met important community needs. To optimize disaster response, community-wide disaster plans should incorporate nursing facilities. PMID- 15284057 TI - Graphic Canadian cigarette warning labels and adverse outcomes: evidence from Canadian smokers. AB - OBJECTIVES: We assessed the impact of graphic Canadian cigarette warning labels. METHODS: We used a longitudinal telephone survey of 616 adult smokers. RESULTS: Approximately one fifth of participants reported smoking less as a result of the labels; only 1% reported smoking more. Although participants reported negative emotional responses to the warnings including fear (44%) and disgust (58%), smokers who reported greater negative emotion were more likely to have quit, attempted to quit, or reduced their smoking 3 months later. Participants who attempted to avoid the warnings (30%) were no less likely to think about the warnings or engage in cessation behavior at follow-up. CONCLUSIONS: Policymakers should not be reluctant to introduce vivid or graphic warnings for fear of adverse outcomes. PMID- 15284058 TI - Overexpression of human heme oxygenase-1 attenuates endothelial cell sloughing in experimental diabetes. AB - Heme oxygenase (HO)-1 represents a key defense mechanism against oxidative injury. Hyperglycemia produces oxidative stress and various perturbations of cell physiology. The effect of streptozotocin (STZ)-induced diabetes on aortic HO activity, heme content, the number of circulating endothelial cells, and urinary 8-epi-isoprostane PGF2alpha (8-Epi) levels in control rats and rats overexpressing or underexpressing HO-1 was measured. HO activity was decreased in hyperglycemic rats. Hyperglycemia increased urinary 8-Epi, and this increase was augmented in rats underexpressing HO-1 and diminished in rats overexpressing HO 1. The number of detached endothelial cells and O2- formation increased in diabetic rats and in hyperglycemic animals underexpressing HO-1 and decreased in diabetic animals overexpressing HO-1 compared with controls. These data demonstrate that HO-1 gene transfer in hyperglycemic rats brings about a reduction in O2- production and a decrease in endothelial cell sloughing. Upregulation of HO-1 decreases oxidant production and endothelial cell damage and shedding and may attenuate vascular complications in diabetes. PMID- 15284059 TI - Intravenous administration of mesenchymal stem cells improves cardiac function in rats with acute myocardial infarction through angiogenesis and myogenesis. AB - Mesenchymal stem cells (MSCs) are pluripotent cells that differentiate into a variety of cells, including cardiomyocytes and endothelial cells. However, little information is available regarding the therapeutic potency of systemically delivered MSCs for myocardial infarction. Accordingly, we investigated whether intravenously transplanted MSCs induce angiogenesis and myogenesis and improve cardiac function in rats with acute myocardial infarction. MSCs were isolated from bone marrow aspirates of isogenic adult rats and expanded ex vivo. At 3 h after coronary ligation, 5 x 10(6) MSCs (MSC group, n=12) or vehicle (control group, n=12) was intravenously administered to Lewis rats. Transplanted MSCs were preferentially attracted to the infarcted, but not the noninfarcted, myocardium. The engrafted MSCs were positive for cardiac markers: desmin, cardiac troponin T, and connexin43. On the other hand, some of the transplanted MSCs were positive for von Willebrand factor and formed vascular structures. Capillary density was markedly increased after MSC transplantation. Cardiac infarct size was significantly smaller in the MSC than in the control group (24 +/- 2 vs. 33 +/- 2%, P <0.05). MSC transplantation decreased left ventricular end-diastolic pressure and increased left ventricular maximum dP/dt (both P <0.05 vs. control). These results suggest that intravenous administration of MSCs improves cardiac function after acute myocardial infarction through enhancement of angiogenesis and myogenesis in the ischemic myocardium. PMID- 15284060 TI - Ionic mechanisms mediating the myogenic response in newborn porcine cerebral arteries. AB - Mechanisms that underlie autoregulation in the newborn vasculature are unclear. Here we tested the hypothesis that in newborn porcine cerebral arteries intravascular pressure elevates wall tension, leading to an increase in intracellular calcium concentration ([Ca2+]i) and a constriction that is opposed by pressure-induced K+ channel activation. Incremental step (20 mmHg) elevations in intravascular pressure between 10 and 90 mmHg induced an immediate transient elevation in arterial wall [Ca2+]i and a short-lived constriction that was followed by a smaller steady-state [Ca2+]i elevation and sustained constriction. Pressures between 10 and 90 mmHg increased steady-state arterial wall [Ca2+]i between approximately 142 and 299 nM and myogenic (defined as passive-active) tension between 25 and 437 dyn/cm. The relationship between pressure and myogenic tension was strongly Ca2+ dependent until forced dilation. At low pressure, 60 mM K+ induced a steady-state elevation in arterial wall [Ca2+]i and a constriction. Nimodipine, a voltage-dependent Ca2+ channel blocker, and removal of extracellular Ca2+ similarly dilated arteries at low or high pressures. 4 Aminopyridine, a voltage-dependent K+ (Kv) channel blocker, induced significantly larger constrictions at high pressure, when compared with those at low pressure. Although selective Ca2+-activated K+ (KCa) channel blockers and intracellular Ca2+ release inhibitors induced only small constrictions at low and high pressures, a low concentration of caffeine (1 microM), a ryanodine-sensitive Ca2+ release (RyR) channel activator, increased KCa channel activity and induced dilation. These data suggest that in newborn cerebral arteries, intravascular pressure elevates wall tension, leading to voltage-dependent Ca2+ channel activation, an increase in wall [Ca2+]i and Ca2+-dependent constriction. In addition, pressure strongly activates Kv channels that opposes constriction but only weakly activates KCa channels. PMID- 15284061 TI - Graded alterations of RBC aggregation influence in vivo blood flow resistance. AB - Although the effects of red blood cell (RBC) aggregation on low-shear rate blood viscosity are well known, the effects on in vivo flow resistance are still not fully resolved. The present study was designed to explore the in vivo effects of RBC aggregation on flow resistance using a novel technique to enhance aggregation: cells are covalently coated with a block copolymer (Pluronic F-98) and then suspended in unaltered plasma. RBC aggregation was increased in graded steps by varying the Pluronic concentration during cell coating and was verified by microscopy and erythrocyte sedimentation rate (ESR), which increased by 200% at the highest Pluronic level. RBC suspensions were perfused through an isolated in situ guinea pig hindlimb preparation while the arterial perfusion pressure was held constant at 100 mmHg via a pressure servo-controlled pump. No significant effects of enhanced RBC aggregation were observed when studies were conducted in preparations with intact vascular control mechanisms. However, after inhibition of smooth muscle tone (using 10(-4) M papaverin), a significant change in flow resistance was observed in a RBC suspension with a 97% increase of ESR. Additional enhancements of RBC aggregation (i.e., 136 and 162% increases of ESR) decreased flow resistance almost to control values. This was followed by another significant increase in flow resistance during perfusion with RBC suspensions with a 200% increase of ESR. This triphasic effect of graded increases of RBC aggregation is most likely explained by an interplay of several hemodynamic mechanisms that are triggered by enhanced RBC aggregation. PMID- 15284062 TI - Effect of soluble epoxide hydrolase inhibition on epoxyeicosatrienoic acid metabolism in human blood vessels. AB - We investigated the effects of soluble epoxide hydrolase (sEH) inhibition on epoxyeicosatrienoic acid (EET) metabolism in intact human blood vessels, including the human saphenous vein (HSV), coronary artery (HCA), and aorta (HA). When HSV segments were perfused with 2 micromol/l 14,15-[3H]EET for 4 h, >60% of radioactivity in the perfusion medium was converted to 14,15 dihydroxyeicosatrienoic acid (DHET). Similar results were obtained with endothelium-denuded vessels. 14,15-DHET was released from both the luminal and adventitial surfaces of the HSV. When HSVs were incubated with 14,15-[3H]EET under static (no flow) conditions, formation of 14,15-DHET was detected within 15 min and was inhibited by the selective sEH inhibitors N,N'-dicyclohexyl urea and N-cyclohexyl-N'-dodecanoic acid urea (CUDA). Similarly, CUDA inhibited the conversion of 11,12-[3H]EET to 11,12-DHET by the HSV. sEH inhibition enhanced the uptake of 14,15-[3H]EET and facilitated the formation of 10,11-epoxy-16:2, a beta oxidation product. The HCA and HA converted 14,15-[3H]EET to DHET, and this also was inhibited by CUDA. These findings in intact human blood vessels indicate that conversion to DHET is the predominant pathway for 11,12- and 14,15-EET metabolism and that sEH inhibition can modulate EET metabolism in vascular tissue. PMID- 15284063 TI - Rat heart is a site of leptin production and action. AB - Leptin, the 16-kDa peptide hormone product of the ob gene, is produced primarily by adipocytes and was initially thought to exert its effects exclusively through actions on the hypothalamus via distinct leptin receptors termed OB-R. However, recent data show that leptin is produced elsewhere and that receptors are present in many other tissues. Using real-time PCR, we determined whether leptin and its receptors are present in the rat heart and demonstrated regional distribution patterns and gender differences as well as the effect of ischemia and reperfusion. Gene expression of leptin and its receptors (OB-Ra, OB-Rb, and OB Re) was identified in myocytes and whole heart homogenates from all regions of the heart of male and female rats, with the highest abundance in left and right atria of male and female rats, respectively. No differences in regional distribution of OB-R were evident in male rat hearts. In female rats, expression was highest in right atria for all three isoforms and was significantly greater than in male rats. Ischemia and reperfusion significantly downregulated leptin and OB-R expression, although this was more pronounced in male rat hearts. Leptin release in the coronary effluent was also detected using ELISA, although this was generally unaffected by global ischemia and reperfusion. Our results demonstrate for the first time the presence of the leptin system, including the peptide and its receptors, in all regions of the rat heart. In view of emerging evidence for cardiac effects of leptin, it is proposed that the heart is a target for leptin action and that the peptide modulates function through a paracrine- or autocrine dependent manner. PMID- 15284064 TI - Brief episode of STZ-induced hyperglycemia produces cardiac abnormalities in rats fed a diet rich in n-6 PUFA. AB - Diabetic patients are particularly susceptible to cardiomyopathy independent of vascular disease, and recent evidence implicates cell death as a contributing factor. Given its protective role against apoptosis, we hypothesized that dietary n-6 polyunsaturated fatty acid (PUFA) may well decrease the incidence of this mode of cardiac cell death after diabetes. Male Wistar rats were first fed a diet rich in n-6 PUFA [20% (wt/wt) sunflower oil] for 4 wk followed by streptozotocin (STZ, 55 mg/kg) to induce diabetes. After a brief period of hyperglycemia (4 days), hearts were excised for functional, morphological, and biochemical analysis. In diabetic rats, n-6 PUFA decreased caspase-3 activity, crucial for myocardial apoptosis. However, cardiac necrosis, an alternative mode of cell death, increased. In these hearts, a rise in linoleic acid and depleted cardiac glutathione could explain this "switch" to necrotic cell death. Additionally, mitochondrial abnormalities, impaired substrate utilization, and enhanced triglyceride accumulation could have also contributed to a decline in cardiac function in these animals. Our study provides evidence that, in contrast to other models of diabetic cardiomyopathy that exhibit cardiac dysfunction only after chronic hyperglycemia, n-6 PUFA feeding coupled with only 4 days of diabetes precipitated metabolic and contractile abnormalities in the heart. Thus, although promoted as being beneficial, excess n-6 PUFA, with its predisposition to induce obesity, insulin resistance, and ultimately diabetes, could accelerate myocardial abnormalities in diabetic patients. PMID- 15284065 TI - Exercise hyperventilation, dyspnea sensation, and ergoreflex activation in lone atrial fibrillation. AB - Lone atrial fibrillation may be associated with daily life disability and exercise limitation. The extracardiac pathophysiology of these effects is poorly explored. In 35 subjects with lone atrial fibrillation (mean age 67 +/- 7 yr), we investigated pulmonary function, symptom-limited cardiopulmonary exercise performance, muscle ergoreflex (handgrip exercise) contribution to ventilation, and brachial artery flow-mediated dilation (as a measure of endothelial function) before and after (average interval 20 +/- 5 days) restoring sinus rhythm with external cardioversion. Respiratory volumes and lung diffusing capacity at rest were within normal limits during both atrial fibrillation and after restoring sinus rhythm. Cardioversion was associated with the following changes: a decrease of the slope of exercise ventilation vs. CO2 production (from 35 +/- 5 to 29 +/- 3; P <0.01) and of dyspnea sensation (Borg score from 4 to 2) and an increase of peak oxygen uptake (Vo2; from 16 +/- 4 to 20 +/- 5 ml.min(-1).kg(-1); P <0.01), Vo2 at anaerobic threshold (from 11 +/- 2 to 13 +/- 2 ml.min(-1).kg(-1); P <0.05), and O2 pulse (from 8 +/- 3 to 11 +/- 3 ml/beat; P <0.01). After cardioversion, the observed improvement in ventilatory efficiency was accompanied by a significant peak end-tidal CO2 increase (from 33 +/- 2 to 37 +/- 2 mmHg; P <0.01) and no changes in dead space-to-tidal volume ratio (from 0.23 +/- 0.03 to 0.23 +/- 0.02; P=not significant). In addition, the ergoreflex contribution to ventilation was remarkably attenuated, and the brachial artery flow-mediated dilatation was significantly augmented (from 0.32 +/- 0.07 to 0.42 +/- 0.08 mm; P <0.01). Ten patients had atrial fibrillation relapse and, compared with values after restoration of regular sinus rhythm, invariably showed worsening of endothelial function, exercise ventilatory efficiency, and muscle ergoreflex contribution to ventilation. In subjects with lone atrial fibrillation, an impairment in ventilatory efficiency appears to be involved in the pathophysiology of exercise limitation, and to be primarily related with a demodulated peripheral control of ventilation. PMID- 15284067 TI - Coupled pacing improves cardiac efficiency during acute atrial fibrillation with or without cardiac dysfunction. AB - Coupled pacing (CP), a method for controlling ventricular rate during atrial fibrillation (AF), consists of a single electrical stimulation applied to the ventricles after each spontaneous activation. CP results in a mechanical contraction rate approximately one-half the rate during AF. Paired stimulation in which two electrical stimuli are delivered to the ventricles has also been proposed as a therapy for heart failure. Although paired stimulation enhances contractility, it greatly increases energy consumption. The primary hypothesis of the present study is that CP improves cardiac function during acute AF without a similar increase in energy consumption because of the reduced rate of ventricular contractions. In a canine model, CP was applied during four stages: sinus rhythm (SR), acute AF, cardiac dysfunction (CD), and AF in the presence of cardiac dysfunction. The rate of ventricular contraction decreased in all four stages as the result of CP. In addition, we determined the changes in external cardiac work, myocardial oxygen consumption, and myocardial efficiency in the each of four stages. CP partially reversed the effects of AF and CD on external cardiac work, whereas myocardial oxygen consumption increased only moderately. In all stages but SR, CP increased myocardial efficiency because of the marked increases in cardiac work compared with the moderate increases in total energy consumed. Thus this pacing therapy may be a viable therapy for patients with concurrent atrial fibrillation and heart failure. PMID- 15284066 TI - Cobalt chloride induces delayed cardiac preconditioning in mice through selective activation of HIF-1alpha and AP-1 and iNOS signaling. AB - Acute systemic hypoxia induces delayed cardioprotection against ischemia (I) reperfusion (R) injury via inducible nitric oxide synthase (iNOS)-dependent mechanism. Because CoCl2 is known to elicit hypoxia-like responses, we hypothesized that this chemical would mimic the delayed preconditioning effect in the heart. Adult male mice were pretreated with CoCl2 or saline. The hearts were isolated 24 h later and subjected to 20 min of global I and 30 min of R in Langendorff mode. Myocardial infarct size (% of risk area; mean +/- SE, n=6 8/group) was reduced in mice pretreated with 30 mg/kg CoCl2 (16.1 +/- 3.1% vs. 27.6 +/- 3.3% with saline control; P <0.05) without compromising postischemic cardiac function. Higher doses of CoCl2 failed to induce similar protection. Electrophoretic mobility gel shift assay demonstrated significant enhancement in DNA binding activity of hypoxia-inducible factor 1alpha (HIF-1alpha) and activator protein 1 (AP-1) in nuclear extracts from CoCl2-treated hearts. Activation of HIF-1alpha and AP-1 was evident at 30 min and sustained for the next 4 h after CoCl2 injection. In contrast, CoCl2-induced protection was independent of NF-kappaB activation because no DNA binding or p65 translocation was observed in nuclear extracts. Also, CoCl2-induced cardioprotection was preserved in p50 subunit NF-kappaB-knockout (KO) mice (11.1 +/- 3.0% vs. 25.1 +/- 5.0% in saline-treated p50-KO mice; P <0.05). The infarct-limiting effect of CoCl2 was absent in iNOS-KO mice (20.9 +/- 3.0%). We conclude that in vivo administration of CoCl2 preconditions the heart against I/R injury. The delayed protective effect of CoCl2 is achieved through a distinctive signaling mechanism involving HIF-1alpha, AP-1, and iNOS but independent of NF-kappaB activation. PMID- 15284068 TI - Mathematical modeling of vascular endothelial layer maintenance: the role of endothelial cell division, progenitor cell homing, and telomere shortening. AB - Maintenance of the endothelial cell (EC) layer of the vessel wall is essential for proper functioning of the vessel and prevention of vascular disorders. Replacement of damaged ECs could occur through division of surrounding ECs. Furthermore, EC progenitor cells (EPCs), derived from the bone marrow and circulating in the bloodstream, can differentiate into ECs. Therefore, these cells might also play a role in maintenance of the endothelial layer in the vascular system. The proliferative potential of both cell types is limited by shortening of telomeric DNA. Accelerated telomere shortening might lead to senescent vascular wall cells and eventually to the inability of the endothelium to maintain a continuous monolayer. The aim of this study was to describe the dynamics of EC damage and repair and telomere shortening by a mathematical model. In the model, ECs were integrated in a two-dimensional structure resembling the endothelium in a large artery. Telomere shortening was described as a stochastic process with oxidative damage as the main cause of attrition. Simulating the model illustrated that increased cellular turnover or elevated levels of oxidative stress could lead to critical telomere shortening and senescence at an age of 65 yr. The model predicted that under those conditions the EC layer could display defects, which could initiate severe vascular wall damage in reality. Furthermore, simulations showed that 5% progenitor cell homing/yr can significantly delay the EC layer defects. This stresses the potential importance of EPC number and function to the maintenance of vascular wall integrity during the human life span. PMID- 15284069 TI - Feasibility of cardiac microimpedance measurement using multisite interstitial stimulation. AB - This study was designed to test the hypothesis that analyses of central interstitial potential differences recorded during multisite stimulation with a set of interstitial electrodes provide sufficient data for accurate measurement of cardiac microimpedances. On theoretical grounds, interstitial current injected and removed using electrodes in close proximity does not cross the membrane, whereas equilibration of intracellular and interstitial potentials occurs distant from electrodes widely separated. Multisite interstitial stimulation should therefore give rise to interstitial potential differences recorded centrally that depend on intracellular and interstitial microimpedances, allowing independent measurement. Simulations of multisite stimulation with fine (25 microm) and wide (400 microm) spacing in one-dimensional models that included Luo-Rudy dynamic membrane equations were performed. Constant interstitial and intracellular microimpedances were prescribed for initial analyses. Discrete myoplasmic and gap junctional components were prescribed intracellularly in later simulations. With constant microimpedances, multisite stimulation using 29 total electrode combinations allowed interstitial and intracellular microimpedance measurements at errors of 0.30% and 0.34%, respectively, with errors of 0.05% and 0.40% achieved using 6 combinations and 10 total electrodes. With discrete myoplasmic and junctional components, comparable accuracy was maintained following adjustments to the junctions to reflect uncoupling. This allowed uncoupling to be quantified as relative increases in total junctional resistance. Our findings suggest development of microfabricated devices to implement the procedure would facilitate routine measurement as a component of cardiac electrophysiological study. PMID- 15284070 TI - Control of myocardial oxygen consumption in transgenic mice overexpressing vascular eNOS. AB - Our objective was to investigate the potential role of selective endothelial nitric oxide (NO) synthase (eNOS) overexpression in coronary blood vessels in the control of myocardial oxygen consumption (MVO2). Transgenic (Tg) eNOS overexpressing mice (eNOS Tg) (n=22) and wild-type (WT) mice (n=24) were studied. Western blot analysis indicated greater than sixfold increase of eNOS in cardiac tissue. Echocardiography in awake mice indicated no difference in cardiac function between WT and eNOS Tg; however, systolic pressure in eNOS Tg mice decreased significantly (126 +/- 2.3 to 109 +/- 2.3 mmHg; P <0.05), whereas heart rate (HR) was not different. Total peripheral resistance (TPR) was also decreased (9.8 +/- 0.8 to 7.6 +/- 0.4 4 mmHg.ml(-1).min; P <0.05) in eNOS Tg. Furthermore, female eNOS Tg mice showed even lower TPR (7.2 +/- 0.4 mmHg.ml(-1).min) compared with male eNOS mice (8.6 +/- 0.5, mmHg.ml.min(-1); P <0.05). Left ventricular slices were isolated from WT and eNOS Tg mice. With the use of a Clark-type oxygen electrode in an airtight bath, MVO2 was determined as the percent decrease during increasing doses (10(-10) to 10(-4) mol/l) of bradykinin (BK), carbachol (CCh), forskolin (10(-12) to 10(-6) mol/l), or S-nitroso-N-acetyl penicillamine (SNAP; 10(-7) to 10(-4) mol/l). Baseline MVO2 was not different between WT (181 +/- 13 nmol.g(-1).min(-1)) and eNOS Tg (188 +/- 14 nmol.g(-1).min(-1)). BK decreased MVO2 (10(-4) mol/l) in WT by 17% +/- 1.1 and 33% +/- 2.7 in eNOS Tg (P < 0.05). CCh also decreased MVO2, 10(-4) mol/l, in WT by 20% +/- 1.7 and 31% +/- 2.0 in eNOS Tg (P <0.05). Forskolin (10(-6) mol/l) or SNAP (10(-4) mol/l) also decreased MVO2 in WT by 24% +/- 2.8 and 36% +/- 1.8 versus eNOS 31% +/- 1.8 and 37% +/- 3.5, respectively. N-nitro-L-arginine methyl ester (10(-3) mol/l) inhibited the MVO2 reduction to BK, CCh, and forskolin by a similar degree (P <0.05), but not to SNAP. Thus selective overexpression of eNOS in cardiac blood vessels in mice enhances the control of MVO2 by eNOS-derived NO. PMID- 15284071 TI - Functional effects of enhancing or silencing adenosine A2b receptors in cardiac fibroblasts. AB - Cardiac fibroblasts (CF) express adenosine (ADO) receptors, and pharmacological evidence suggests the possible involvement of the A2 (A2a and A2b) receptor (A2aR and A2bR) subtypes in inhibiting cell functions involved in fibrosis. The main objective of this study was to define the contributions of A2a and/or A2b receptors in modulating ADO-induced decreases in CF functions. For this purpose, CF were either treated pharmacologically or had the A2aR or A2bR levels modified through the use of recombinant adenovirus or siRNA. The assessment of mRNA expression in adult rat CF yielded evidence for A1R, A2bR, A2a), and A3R. Endogenously or exogenously enhanced ADO significantly inhibits CF proliferation, collagen, and protein synthesis. A2R and A2aR agonists, although capable of inhibiting CF protein and collagen synthesis, were unable to define the contributions derived from A2aR or A2bR. Overexpression of A2bR in CF yielded significant decreases in basal levels of collagen and protein synthesis and correlated with increases in cAMP levels. However, at higher doses of ADO receptor agonists, significant increases in protein and collagen synthesis were observed. CF with underexpression of A2bR yielded increases in protein and collagen synthesis. In contrast, A2aR underexpression did not modify ADO-induced decreases in CF protein or collagen synthesis. In conclusion, results derived from the molecular manipulation of receptor levels indicate that A2bR are critically involved in ADO-mediated inhibition of CF functions. PMID- 15284072 TI - c-Fos expression in rat brain stem and spinal cord in response to activation of cardiac ischemia-sensitive afferent neurons and electrostimulatory modulation. AB - The purpose of this study was to identify central neuronal sites activated by stimulation of cardiac ischemia-sensitive afferent neurons and determine whether electrical stimulation of left vagal afferent fibers modified the pattern of neuronal activation. Fos-like immunoreactivity (Fos-LI) was used as an index of neuronal activation in selected levels of cervical and thoracic spinal cord and brain stem. Adult Sprague-Dawley rats were anesthetized with urethane and underwent intrapericardial infusion of an "inflammatory exudate solution" (IES) containing algogenic substances that are released during ischemia (10 mM adenosine, bradykinin, prostaglandin E2, and 5-hydroxytryptamine) or occlusion of the left anterior descending coronary artery (CoAO) to activate cardiac ischemia sensitive (nociceptive) afferent fibers. IES and CoAO increased Fos-LI above resting levels in dorsal horns in laminae I-V at C2 and T4 and in the caudal nucleus tractus solitarius. Dorsal rhizotomy virtually eliminated Fos-LI in the spinal cord as well as the brain stem. Neuromodulation of the ischemic signal by electrical stimulation of the central end of the left thoracic vagus excited neurons at the cervical and brain stem level but inhibited neurons at the thoracic spinal cord during IES or CoAO. These results suggest that stimulation of the left thoracic vagus excites descending inhibitory pathways. Inhibition at the thoracic spinal level that suppresses the ischemic (nociceptive) input signal may occur by a short-loop descending pathway via signals from cervical propriospinal circuits and/or a longer-loop descending pathway via signals from the nucleus tractus solitarius. PMID- 15284073 TI - Effects of nitric oxide synthase inhibitor on decrease in peripheral arterial stiffness with acute low-intensity aerobic exercise. AB - We previously reported that even low-intensity, short-duration acute aerobic exercise decreases arterial stiffness. We aimed to test the hypothesis that the exercise-induced decrease in arterial stiffness is caused by the increased production of NO in vascular endothelium with exercise. Nine healthy men (age: approximately 22-28 yr) performed a 5-min single-leg cycling exercise (30 W) in the supine position under an intravenous infusion of NG-monomethyl-L-arginine (L NMMA; 3 mg/kg during the initial 5 min and subsequent continuous infusion of 50 mug.kg(-1).min(-1) in saline) or vehicle (saline) in random order on separate days. The pulse wave velocity (PWV) from the femoral to posterior tibial artery was measured on both legs before and after the infusion at rest and 2 min after exercise. Under the control condition, exercised leg PWV significantly decreased after exercise (P <0.05), whereas nonexercised leg PWV did not show a significant change throughout the experiment. Under L-NMMA administration, exercised leg PWV was increased significantly by the infusion (P <0.05) but decreased significantly after the exercise (P <0.05). Nonexercised leg PWV increased with L-NMMA administration and maintained a significantly higher level during the administration compared with baseline (before the infusion, all P <0.05). The NO synthase blockade x time interaction on exercised leg PWV was not significant (P=0.706). These results suggest that increased production of NO is not a major factor in the decrease of regional arterial stiffness with low-intensity, short duration aerobic exercise. PMID- 15284074 TI - Central Tempol alters basal sympathetic nerve discharge and attenuates sympathetic excitation to central ANG II. AB - In the present study, we established dose-response relationships between central administration of 4-hydroxy-2,2,6,6-tetramethylpiperidine 1-oxyl (Tempol, a superoxide dismutase mimetic) and the level of renal sympathetic nerve discharge (SND) and tested the hypothesis that intracerebroventricular (icv) Tempol pretreatment would attenuate centrally mediated changes in SND produced by icv ANG II administration. Urethane-chloralose-anesthetized, baroreceptor-denervated, normotensive rats were used. We found that icv Tempol administration produced dose-dependent sympathoinhibitory, hypotensive, and bradycardic responses. Mean arterial pressure and SND values were significantly increased after icv ANG II (150 ng/kg) administration, and these responses were abrogated after icv pretreatment with Tempol (75 micromol/kg) or losartan. Brain superoxide levels tended to be higher in ANG II-treated rats compared with rats treated with Tempol and ANG II. Tempol pretreatment did not prevent increases in SND level that were produced by acute heat stress, which indicates specificity in the effect of Tempol in reducing sympathoexcitation. These results demonstrate that icv Tempol administration influences central sympathetic neural circuits in a dose-dependent manner and attenuates SND responses to central ANG II infusion. PMID- 15284075 TI - Differential regulation of human lung epithelial and endothelial barrier function by thrombin. AB - Lung epithelial and endothelial barrier dysfunction is critical to the physiologic derangement observed in acute lung injury, but remains poorly understood. We utilized human alveolar epithelial (A549) and endothelial cells (EC) to study cytoskeletal remodeling, myosin light chain (MLC) phosphorylation and barrier regulation evoked by the edemagenic agent, thrombin. Thrombin challenged human EC monolayers demonstrated increased MLC phosphorylation, actin stress fiber formation and loss of barrier integrity reflected by decreased transmonolayer electrical resistance (TER). In contrast, thrombin produced prominent circumferential localization of actin fibers, increased MLC phosphorylation and increased TER across epithelial monolayers, consistent with barrier protection. Reductions in MLC phosphorylation induced by cell pretreatment with pharmacological inhibitors of MLC kinase (ML-7) and Rho kinase (Y-27632) significantly attenuated thrombin-mediated TER changes and MLC phosphorylation in both lung cell types. Thrombin-produced, time-dependent activation of Rho GTPase in both epithelial and EC, whereas Rac GTPase activation was observed only in A549 cells. Molecular inhibition of Rac activity by adenoviral transfer of dominant-negative Rac mutant abolished thrombin-induced TER increases in alveolar epithelial cells. Finally, A549 cells, but not endothelium, demonstrated increased levels of tight junction proteins (ZO-1 and occludin) after thrombin at the cell-cell interface areas linked to thrombin elicited barrier protection. These results demonstrate differential pulmonary endothelial and alveolar epithelial barrier regulation via unique actomyosin remodeling and cytoskeletal interactions with tight junction complexes, which confer selective barrier responses to edemagenic stimuli. PMID- 15284076 TI - Emphysema lung tissue gene expression profiling. AB - Emphysema occurs in a subgroup of patients with chronic obstructive pulmonary disease and patients with the genetic defect of alpha(1)-antitrypsin deficiency who have a smoking history of many years' duration. Emphysema is generally the result of a chronic and progressive destruction of the alveolar structures, which is believed to be driven by chronic inflammation, infections, oxidative stress, and an imbalance of protease and antiprotease activity. Here, we use microarray technology to characterize the gene expression profile of lung tissue samples obtained from patients with advanced emphysema and that obtained from healthy subjects. We hypothesized that the gene expression profile of emphysema lung tissue is distinct when compared with the expression profile of normal lungs. We report that severely emphysematous tissue is characterized by a global decrease in gene expression and by an increased abundance of transcripts encoding proteins involved in inflammation, immune responses, and proteolysis. Whereas the gene expression profile is to some degree shared between "usual" emphysema and alpha(1)-antitrypsin deficiency-related emphysema, there are statistically significant differences in the modulation of groups of genes associated with protein and energy metabolism, and immune function, which allow distinction between these two emphysema types on the lung tissue level. PMID- 15284077 TI - Surfactant protein D binding to terminal alpha1-3-linked fucose residues and to Schistosoma mansoni. AB - Pulmonary surfactant protein (SP)-D is an important component of the innate immune system of the lung, which is thought to function by binding to specific carbohydrates on the surface of viruses and unicellular pathogens. SP-D has been shown to have a relatively high affinity for the monosaccharides mannose, glucose, and fucose. However, there is limited information on SP-D binding to complex carbohydrate structures, and binding of SP-D to fucose in the context of an oligosaccharide has not yet been investigated. In this study, we used surface plasmon resonance spectroscopy to examine the potential of SP-D to bind to various synthetic fucosylated oligosaccharides, and identified Fucalpha1-3GalNAc and Fucalpha1-3GlcNAc elements as strong ligands. These types of fucosylated glycoconjugates are presented at the surface of Schistosoma mansoni, a parasitic worm that, during development, transiently resides in the lung. In line with the findings by surface plasmon resonance, we found that SP-D can bind to larval stages of S. mansoni, demonstrating for the first time that SP-D interacts with multicellular lung pathogens. PMID- 15284078 TI - Localization of plasminogen activator activity within normal and injured lungs by in situ zymography. AB - During inflammatory lung injury, the fibrinolytic activity that is normally present within bronchoalveolar lavage (BAL) fluid (BALF) is often suppressed due to increased levels of inhibitors, including plasminogen activator inhibitor (PAI)-1. Despite this suppression, BALF frequently contains fibrin degradation products, indicating persistence of fibrinolytic activity within the lung. To address this discrepancy and determine the sites where plasminogen activation is occurring, we developed an in situ zymographic technique for frozen sections of lung tissue that localizes plasminogen activator activity at the cellular level. After validating the method using enzyme inhibitors and mice with genetic manipulations of their plasminogen system genes, we applied the technique to lungs of normal and bleomycin-exposed mice. In normal mice, plasminogen activator activity was localized to bronchial epithelial cells, cells of the alveolar walls, and alveolar macrophages. After bleomycin exposure, in situ zymography showed that, despite loss of fibrinolytic activity within BALF, abundant enzymatic activity was associated with aggregates of inflammatory cells. PAI-1 deficient mice that are protected from bleomycin-induced fibrosis had preserved plasminogen activator activity in BALF and increased tissue activity, as determined by in situ zymography. We conclude that analysis of BALF does not adequately reflect the fibrinolytic activity that persists within microenvironments of the lung during inflammation. PMID- 15284079 TI - Intracellular-specific colocalization of prostaglandin E2 synthases and cyclooxygenases in the brain. AB - Prostaglandin E2 (PGE2) is the major primary prostaglandin generated by brain cells. However, the coordination and intracellular localization of the cyclooxygenases (COXs) and prostaglandin E synthases (PGESs) that convert arachidonic acid to PGE2 in brain tissue are not known. We aimed to determine whether microsomal and cytosolic PGES (mPGES-1 and cPGES) colocalize and coordinate activity with either COX-1 or COX-2 in brain tissue, particularly during development. Importantly, we found that cytosolic PGES also associates with microsomes (cPGES-m) from the cerebrum and cerebral vasculature of the pig and rat as well as microsomes from various cell lines; this seemed dependent on the carboxyl terminal 35-amino acid domain and a cysteine residue (C58) of cPGES. In microsomal membranes from the postnatal brain and cerebral microvessels of mature animals, cPGES-m colocalized with both COX-1 and COX-2, whereas mPGES-1 was undetectable in these microsomes. Accordingly, in this cell compartment, cPGES could coordinate its activity with COX-2 and COX-1 (partly inhibited by NS398); albeit in microsomes of the brain microvasculature from newborns, mPGES-1 was also present. In contrast, in nuclei of brain parenchymal and endothelial cells, mPGES-1 and cPGES colocalized exclusively with COX-2 (determined by immunoblotting and immunohistochemistry); these PGESs contributed to conversion of PGH2 into PGE2. Hence, contrary to a previously proposed model of exclusive COX-2/mPGES-1 coordination, COX-2 can coordinate with mPGES-1 and/or cPGES in the brain, depending on the cell compartment and the age group. PMID- 15284080 TI - FHL5, a novel actin-binding protein, is highly expressed in eel gill pillar cells and responds to wall tension. AB - Supporting evidence for the contractile nature of fish branchial pillar cells was provided by demonstrating the presence of actin fibers and a novel four-and-a half LIM (FHL) protein in which expression is specific for contractile tissues and sensitive to the tension applied to the pillar cell. When eel gill sections were stained with rhodamine-phalloidin, a selective fluorescent probe for fibrous actin, a strong bundle-like staining was observed around collagen columns in pillar cells, suggesting the presence of abundant actin fibers. A cDNA clone encoding a novel member of the actin-binding FHL family, FHL5, was isolated from a subtracted cDNA library of eel gill. Northern analysis revealed that FHL5 mRNA is highly expressed only in gills, heart, and skeletal muscle. In gills, FHL5 was found to be confined to pillar cells by immunohistochemistry. Confocal fluorescence microscopy showed that FHL5 is present in both cytosol and nucleus; within the cytosol, a large portion of FHL5 is colocalized with the phalloidin positive actin bundles. Furthermore, transfection of myogenic C2C12 cells with FHL5 cDNA demonstrated, in addition to its interaction with actin stress fibers, a nuclear shuttling activity of FHL5. The mRNA and protein levels were found to be elevated on 1) transfer of eels from seawater to freshwater, 2) volume expansion by infusion of isotonic dextran-saline, and 3) constriction of gill vasculature by bolus injection of endothelin-1. These results suggest contractile nature of pillar cells and a role of FHL5 in maintaining the integrity and regulating the dynamics of pillar cells. PMID- 15284081 TI - Acclimatization to neurological decompression sickness in rabbits. AB - Diving acclimatization refers to a reduced susceptibility to acute decompression sickness (DCS) in individuals undergoing repeated compression-decompression cycles. We demonstrated in a previous study that the mechanism responsible for this acclimatization is similar to that of stress preconditioning. In this study, we investigated the protective effect of prior DCS preconditioning on the severity of neurological DCS in subsequent exposure to high pressure in rabbits. We exposed the rabbits (n = 10) to a pressure cycle of 6 absolute atmospheres (ATA) for 90 min, which induced signs of neurological DCS in 60% of the animals. Twenty-four hours after the pressure cycle, rabbits with DCS expressed more heat shock protein 70 (HSP70) in the lungs, liver, and heart than rabbits without signs of disease or those in the control group (n = 6). In another group of rabbits (n = 24), 50% of animals presented signs of neurological DCS after exposure to high pressure, with a neurological score of 46.5 (SD 19.5). A course of hyperbaric oxygen therapy alleviated the signs of neurological DCS and ensured the animals' survival for 24 h. Experiencing another pressure cycle of 6 ATA for 90 min, 50% of 12 rabbits with prior DCS preconditioning developed signs of DCS, with a neurological score of 16.3 (SD 28.3), significantly lower than that before hyperbaric oxygen therapy (P = 0.002). In summary, our results show that the occurrence of DCS in rabbits after rapid decompression is associated with increased expression of a stress protein, indicating that the stress response is induced by DCS. This phenomenon was defined as "DCS preconditioning." DCS preconditioning attenuated the severity of neurological DCS caused by subsequent exposure to high pressure. These results suggest that bubble formation in tissues activates the stress response and stress preconditioning attenuates tissue injury on subsequent DCS stress, which may be the mechanism responsible for diving acclimatization. PMID- 15284082 TI - Measuring committed preadipocytes in human adipose tissue from severely obese patients by using adipocyte fatty acid binding protein. AB - To understand the significance of the reported depot differences in preadipocyte dynamics, we developed a procedure to identify committed preadipocytes in the stromovascular fraction of fresh human adipose tissue. We documented that adipocyte fatty acid binding protein (aP2) is expressed in human preadipocyte clones capable of replication, indicating that can be used as a marker of committed preadipocytes. Because aP2 expression can be induced in macrophages, stromovascular cells were also stained for the macrophage marker CD68. We found aP2+CD68- cells (designated as committed preadipocytes) that did not have lipid droplets (true preadipocytes) and that did have lipid droplets < 6.5 microm in diameter (very immature adipocytes). Adipose tissue from subcutaneous, omental, and mesenteric depots was obtained from nine patients undergoing bariatric surgery for measurement of stromovascular cell number, the number of committed preadipocytes (aP2+CD68-), aP2+ macrophages (aP2+CD68+), and aP2- macrophages (aP2-CD68+). The number of committed preadipocytes did not differ significantly between depots but varied >20-fold among individuals. Total cell number, stromovascular cell number, and the number of aP2- macrophages was less (P < 0.05) in subcutaneous than in omental fat (means +/- SE, in millions: subcutaneous, 2.3 +/- 0.3, 1.4 +/- 0.3, and 0.17 +/- 0.08; and omental, 4.8 +/- 0.7, 3.8 +/- 0.5, and 0.34 +/- 0.06); mesenteric depot was intermediate. These data indicate that the cellular composition of adipose tissue varies between depots and between individuals. The ability to quantify committed preadipocytes in fresh adipose tissue should facilitate study of adipose tissue biology. PMID- 15284083 TI - Effects of aging on cardiac and skeletal muscle AMPK activity: basal activity, allosteric activation, and response to in vivo hypoxemia in mice. AB - Although a diminished ability of tissues and organisms to tolerate stress is a clinically important hallmark of normal aging, little is known regarding its biochemical basis. Our goal was to determine whether age-associated changes in AMP-activated protein kinase (AMPK), a key regulator of cellular metabolism during the stress response, might contribute to the poor stress tolerance of aged cardiac and skeletal muscle. Basal AMPK activity and the degree of activation of AMPK by AMP and by in vivo hypoxemia (arterial Po2 of 39 mmHg) were measured in cardiac and skeletal muscle (gastrocnemius) from 5- and 24-mo-old C57Bl/6 mice. In the heart, neither basal AMPK activity nor its allosteric activation by AMP was affected by age. However, after 10 min of hypoxemia, the activity of alpha2 AMPK, but not alpha1-AMPK, was significantly higher in the hearts from old than from young mice (P < 0.005), this difference being due to differences in phosphorylation of alpha2-AMPK. Significant activation of AMPK in the young hearts did not occur until 30 min of hypoxemia (P < 0.01), stress that was poorly tolerated by the old mice (mortality = 67%). In contrast, AMPK activity in gastrocnemius muscle was unaffected by age or hypoxemia. We conclude that the age associated decline in hypoxic tolerance in cardiac and skeletal muscle is not caused by changes in basal AMPK activity or a blunted AMPK response to hypoxia. Activation of AMPK by in vivo hypoxia is slower and more modest than might be predicted from in vitro and ex vivo experiments. PMID- 15284084 TI - System beta and system A amino acid transporters in the feline endotheliochorial placenta. AB - There is no knowledge of the transport mechanisms by which solutes cross the cat placenta or any other endotheliochorial placenta. Here, we investigated whether the amino acid transport systems beta and A are present in the cat placenta using a placental fragment uptake technique. Data were compared with studies in the human placenta, in which the presence of these two transport systems has been well established. A time course of [(3)H]taurine (substrate for system beta) and [(14)C]MeAIB (nonmetabolizable substrate for system A) uptake was determined in the term cat and human placental fragments in the presence and absence (choline substituted) of Na(+), and further studies were carried out over 15 min. Taurine uptake into both cat and human placenta fragments was found to be Na(+) and Cl(-) dependent, and Na(+)-dependent taurine uptake was blocked by excess beta-alanine. MeAIB uptake was found to be Na(+) dependent, and Na(+)-dependent MeAIB uptake was blocked by excess MeAIB or glycine. Western blotting and immunohistochemistry performed on cat and human placenta showed expression of TAUT and ATA2 (SNAT2), proteins associated with system beta and system A activity, respectively. This study therefore provides the first evidence of the presence of amino acid transport systems beta and A in the cat placenta. PMID- 15284085 TI - Contrasting effects of oral versus transdermal estrogen on serum amyloid A (SAA) and high-density lipoprotein-SAA in postmenopausal women. AB - OBJECTIVES: Previous studies indicated that oral estrogen increased C-reactive protein by a first-pass hepatic effect. In this study, we determine whether the route of estrogen administration influences serum amyloid A (SAA), another acute phase protein produced by the liver, and the SAA content of the high-density lipoprotein (HDL-SAA) in postmenopausal women. METHODS AND RESULTS: In 29 postmenopausal women without coronary heart disease, we conducted a randomized crossover placebo-controlled study to compare effects of transdermal versus oral estrogen on SAA and HDL-SAA. SAA, apolipoprotein A-I, HDL, and HDL-SAA were measured before and after 8 weeks of transdermal estradiol (100 microg per day), oral-conjugated estrogens (0.625 mg per day), or placebo. We found that oral estrogen significantly increased levels of SAA, HDL, and HDL-SAA, whereas transdermal estrogen reduced both SAA and HDL-SAA but had no effect on HDL in the same women. CONCLUSIONS: Oral estrogen increased SAA and altered HDL composition to contain a higher level of SAA by a first-pass hepatic mechanism. Because elevated SAA levels predict adverse prognosis in healthy postmenopausal women, and elevated HDL-SAA levels have been shown to interfere with HDL function, the route of administration may be an important consideration in minimizing side effects of estrogen replacement therapy on cardiovascular outcomes. PMID- 15284086 TI - Can exercise training with weight loss lower serum C-reactive protein levels? AB - OBJECTIVE: C-reactive protein (CRP), an obesity-related inflammatory marker, is a promising predictor for cardiovascular disease and may be a mediator for atherogenesis. It has been reported that diet-induced weight loss lowered CRP levels. However, the effect of exercise training, another therapy that can reduce weight, on CRP is still unclear. We examined effects of exercise training with weight loss on CRP levels and conventional cardiovascular risks. METHODS AND RESULTS: A total of 227 apparently healthy women were recruited, and 199 subjects (average age 52 years) completed a 2-month weight reduction program consisting of supervised aerobic exercises. After the program, weight was reduced from 65.8 to 62.8 kg (P<0.0001), and all conventional variables were remarkably improved. Similarly, CRP levels were significantly decreased, from 0.63 (0.28 to 1.19) to 0.41 (0.18 to 0.80) mg/L (P<0.0001). However, in contrast to other variables, the changes in CRP levels were not proportionally associated with the extent of weight reduction. In the quartile analysis of % weight reduction, the largest weight reduction quartile did not show significant decreases in CRP levels, whereas moderate quartile showed remarkable CRP decreases. CONCLUSIONS: Exercise training with weight reduction disproportionately lowered CRP levels. Considering inflammatory status, there might be an optimal pace of exercise with weight loss. PMID- 15284087 TI - Hepatic lipase, lipoprotein metabolism, and atherogenesis. AB - The role of hepatic lipase as a multifunctional protein that modulates lipoprotein metabolism and atherosclerosis has been extensively documented over the last decade. Hepatic lipase functions as a lipolytic enzyme that hydrolyzes triglycerides and phospholipids present in circulating plasma lipoproteins. Hepatic lipase also serves as a ligand that facilitates lipoprotein uptake by cell surface receptors and proteoglycans, thereby directly affecting cellular lipid delivery. Recently, another process by which hepatic lipase modulates atherogenic risk has been identified. Bone marrow transplantation studies demonstrate that hepatic lipase present in aortic lesions markedly alters aortic lesion formation even in the absence of changes in plasma lipids. These multiple functions of hepatic lipase, which facilitate not only plasma lipid metabolism but also cellular lipid uptake, can be anticipated to have a major and complex impact on atherogenesis. Consistently, human and animal studies support proatherogenic and antiatherogenic roles for hepatic lipase. The concept of hepatic lipase as mainly a lipolytic enzyme that reduces atherogenic risk has evolved into that of a complex protein with multiple functions that, depending on genetic background and sites of expression, can have a variable effect on atherosclerosis. PMID- 15284088 TI - Expression of angiopoietin-2 in endothelial cells is controlled by positive and negative regulatory promoter elements. AB - OBJECTIVE: Angiopoietin-2 (Ang-2) is a non-signal transducing ligand of the endothelial receptor tyrosine kinase Tie-2. Ang-2 is produced by endothelial cells and acts as an autocrine regulator mediating vascular destabilization by inhibiting Angiopoietin-1-mediated Tie-2 activation. To examine the transcriptional regulation of Ang-2, we studied the Ang-2 promoter in endothelial cells and nonendothelial cells. METHODS AND RESULTS: The human Ang-2 promoter contains a 585-bp region around the transcriptional start site (-109 to +476) that is sufficient to control endothelial cell-specific and cytokine-dependent Ang-2 expression. Strong repressor elements of Ang-2-promoter activity are located in the 5'-region of the promoter and in the first intron. The Ets family transcription factors Ets-1 and Elf-1 act as strong enhancers of endothelial cell Ang-2-promoter activity. Ets-binding sites -4 and -7 act as positive regulators, whereas Ets-binding site -3 acts as negative regulator. Demethylation experiments revealed that the Ang-2 gene (in contrast to the Tie-2 gene) is not controlled by imprinting. CONCLUSIONS: The data determine unique positive and negative regulatory mechanisms of endothelial cell Ang-2 expression and provide further evidence for the critical role of Ang-2 as a key autocrine regulator of vascular stability and responsiveness. PMID- 15284089 TI - PECAM-1 interacts with nitric oxide synthase in human endothelial cells: implication for flow-induced nitric oxide synthase activation. AB - OBJECTIVE: We have previously shown that fluid shear stress (FSS) triggers endothelial nitric oxide synthase (eNOS) activity in endothelial cells and that the mechanotransduction mechanisms responsible for activation discriminate between rapid changes in FSS and FSS per se. We hypothesized that the particular sublocalization of eNOS at the cell-cell junction would render it responsive to activation by FSS temporal gradients. METHODS AND RESULTS: In human umbilical vein endothelial cells (HUVECs), immunofluorescence revealed strong eNOS membrane staining at the cell-cell junction colocalizing with platelet/endothelial cell adhesion molecule-1 (PECAM-1). In PECAM-1-/- mouse aorta, eNOS junctional localization seen in the wild type was absent. Similarly, junctional staining was lost in wild-type aorta near intercostal artery branches. eNOS/PECAM-1 association in HUVECs was confirmed by coimmunoprecipitation. When HUVECs were subjected to a 0.5s impulse of 12 dynes/cm2, a transient disruption of the eNOS/PECAM-1 complex was observed, accompanied by an increase in eNOS activity (cGMP production). Ramped flow did not trigger complex dissociation or an increase in cGMP production. In a cell-free system, a direct inhibition of eNOS activity by PECAM-1 is shown. CONCLUSIONS: These results suggest that eNOS is complexed with PECAM-1 at the cell-cell junction and is likely involved in the modulation of eNOS activity by FSS temporal gradients but not by FSS itself. PMID- 15284090 TI - Mast cells in neovascularized human coronary plaques store and secrete basic fibroblast growth factor, a potent angiogenic mediator. AB - OBJECTIVE: Intraplaque neovascularization and hemorrhage may facilitate plaque progression. We studied expression of basic fibroblast growth factor (bFGF), a potent angiogenic mediator, by mast cells (MCs) in human coronary plaques with increasing degrees of atherosclerosis. METHODS AND RESULTS: Normal and atherosclerotic coronary segments were collected from 30 autopsied subjects. Immunohistochemical methods were used to detect MCs, bFGF, and microvessels. Both adventitial and intimal MCs showed intracytoplasmic granular staining for bFGF, and bFGF-positive extracellular granules were observed close to the MCs. Increased numbers of bFGF-positive MCs were detected in neovascularized areas of plaques, and there was a positive correlation between numbers of bFGF-positive MCs and microvessels in both the intima and adventitia. In plaques, the highly neovascularized areas contained increased numbers of bFGF-positive MCs compared with the adjacent nonvascularized areas, where only few MCs were present. Importantly, the proportion of intimal MCs expressing bFGF increased with increasing severity of atherosclerosis. CONCLUSIONS: The present work reveals a novel source of bFGF in human coronary arteries, the intimal and adventitial MCs. The association of bFGF-positive MCs with microvessels and with the severity of atherosclerosis suggests that coronary MCs, by releasing bFGF, may play a role in angiogenesis and progression of coronary plaques. PMID- 15284091 TI - Antimonocyte chemoattractant protein-1 gene therapy attenuates graft vasculopathy. AB - OBJECTIVE: Accelerated coronary arteriosclerosis remains a major problem in the long-term survival of cardiac transplant recipients. However, the pathogenesis of graft vasculopathy is poorly understood, and there is no effective therapy. Transplant arteriosclerosis is characterized by early mononuclear cell attachment on the transplanted vessel followed by development of concentric neointimal hyperplasia. Early and persistent expression of monocyte chemoattractant protein 1 (MCP-1) in cardiac allografts has been implicated for the pathogenesis of transplant arteriosclerosis. METHODS AND RESULTS: We investigated whether anti MCP-1 gene therapy can inhibit the development of intima hyperplasia in a mouse model of cardiac transplantation. Either the dominant-negative form of MCP-1 (7ND) or control vector was transfected into the skeletal muscles of B10.D2 mice. Cardiac allografts from DBA/2 mice were transplanted heterotopically into B10.D2 mice. 7ND gene transfer was associated with a significant reduction of the number of mononuclear cells accumulating in the lumen of the graft coronary arteries at 1 week and an attenuation of the development of the lesion at 8 weeks (intima/media ratio 0.79+/-0.05 versus 0.48+/-0.04). CONCLUSIONS: The MCP 1/chemokine receptor 2 (CCR2) signaling pathway plays a critical role in the pathogenesis of graft vasculopathy. This new anti-MCP-1 gene therapy might be useful to treat graft vascular disease. PMID- 15284092 TI - Discovering patterns to extract protein-protein interactions from full texts. AB - MOTIVATION: Although there are several databases storing protein-protein interactions, most such data still exist only in the scientific literature. They are scattered in scientific literature written in natural languages, defying data mining efforts. Much time and labor have to be spent on extracting protein pathways from literature. Our aim is to develop a robust and powerful methodology to mine protein-protein interactions from biomedical texts. RESULTS: We present a novel and robust approach for extracting protein-protein interactions from literature. Our method uses a dynamic programming algorithm to compute distinguishing patterns by aligning relevant sentences and key verbs that describe protein interactions. A matching algorithm is designed to extract the interactions between proteins. Equipped only with a dictionary of protein names, our system achieves a recall rate of 80.0% and precision rate of 80.5%. AVAILABILITY: The program is available on request from the authors. PMID- 15284093 TI - NMRb: a web-site repository for raw NMR datasets. AB - The development of NMR in structural proteomics requires the availability of automatic structure determination methods. Many researchers are commonly confronted with the lack of raw datasets during the validation step of such methods. In order to increase test possibilities, the NMRb web-site offers a database of NMR raw datasets, ordered by spectral characteristics. AVAILABILITY: NMRb is available from: http://nmrb.cbs.cnrs.fr. SUPPLEMENTARY INFORMATION: General organization of NMRb figure, relational model organization, and XML structure files are available from http://nmrb.cbs.cnrs.fr/nmrb-doc.html. PMID- 15284094 TI - Advances to Bayesian network inference for generating causal networks from observational biological data. AB - MOTIVATION: Network inference algorithms are powerful computational tools for identifying putative causal interactions among variables from observational data. Bayesian network inference algorithms hold particular promise in that they can capture linear, non-linear, combinatorial, stochastic and other types of relationships among variables across multiple levels of biological organization. However, challenges remain when applying these algorithms to limited quantities of experimental data collected from biological systems. Here, we use a simulation approach to make advances in our dynamic Bayesian network (DBN) inference algorithm, especially in the context of limited quantities of biological data. RESULTS: We test a range of scoring metrics and search heuristics to find an effective algorithm configuration for evaluating our methodological advances. We also identify sampling intervals and levels of data discretization that allow the best recovery of the simulated networks. We develop a novel influence score for DBNs that attempts to estimate both the sign (activation or repression) and relative magnitude of interactions among variables. When faced with limited quantities of observational data, combining our influence score with moderate data interpolation reduces a significant portion of false positive interactions in the recovered networks. Together, our advances allow DBN inference algorithms to be more effective in recovering biological networks from experimentally collected data. AVAILABILITY: Source code and simulated data are available upon request. SUPPLEMENTARY INFORMATION: http://www.jarvislab.net/Bioinformatics/BNAdvances/ PMID- 15284095 TI - Quantifying reproducibility for differential proteomics: noise analysis for protein liquid chromatography-mass spectrometry of human serum. AB - SUMMARY: Using replicated human serum samples, we applied an error model for proteomic differential expression profiling for a high-resolution liquid chromatography-mass spectrometry (LC-MS) platform. The detailed noise analysis presented here uses an experimental design that separates variance caused by sample preparation from variance due to analytical equipment. An analytic approach based on a two-component error model was applied, and in combination with an existing data driven technique that utilizes local sample averaging, we characterized and quantified the noise variance as a function of mean peak intensity. The results indicate that for processed LC-MS data a constant coefficient of variation is dominant for high intensities, whereas a model for low intensities explains Poisson-like variations. This result leads to a quadratic variance model which is used for the estimation of sample preparation noise present in LC-MS data. PMID- 15284096 TI - Discovery of meaningful associations in genomic data using partial correlation coefficients. AB - MOTIVATION: A major challenge of systems biology is to infer biochemical interactions from large-scale observations, such as transcriptomics, proteomics and metabolomics. We propose to use a partial correlation analysis to construct approximate Undirected Dependency Graphs from such large-scale biochemical data. This approach enables a distinction between direct and indirect interactions of biochemical compounds, thereby inferring the underlying network topology. RESULTS: The method is first thoroughly evaluated with a large set of simulated data. Results indicate that the approach has good statistical power and a low False Discovery Rate even in the presence of noise in the data. We then applied the method to an existing data set of yeast gene expression. Several small gene networks were inferred and found to contain genes known to be collectively involved in particular biochemical processes. In some of these networks there are also uncharacterized ORFs present, which lead to hypotheses about their functions. AVAILABILITY: Programs running in MS-Windows and Linux for applying zeroth, first, second and third order partial correlation analysis can be downloaded at: http://mendes.vbi.vt.edu/tiki-index.php?page=Software. SUPPLEMENTARY INFORMATION: Supplementary information can be found at: URL to be decided. PMID- 15284097 TI - CLANS: a Java application for visualizing protein families based on pairwise similarity. AB - SUMMARY: The main source of hypotheses on the structure and function of new proteins is their homology to proteins with known properties. Homologous relationships are typically established through sequence similarity searches, multiple alignments and phylogenetic reconstruction. In cases where the number of potential relationships is large, for example in P-loop NTPases with many thousands of members, alignments and phylogenies become computationally demanding, accumulate errors and lose resolution. In search of a better way to analyze relationships in large sequence datasets we have developed a Java application, CLANS (CLuster ANalysis of Sequences), which uses a version of the Fruchterman-Reingold graph layout algorithm to visualize pairwise sequence similarities in either two-dimensional or three-dimensional space. AVAILABILITY: CLANS can be downloaded at http://protevo.eb.tuebingen.mpg.de/download. PMID- 15284098 TI - The Baumgartner-Weiss-Schindler test for the detection of differentially expressed genes in replicated microarray experiments. AB - MOTIVATION: An important application of microarray experiments is to identify differentially expressed genes. Because microarray data are often not distributed according to a normal distribution nonparametric methods were suggested for their statistical analysis. Here, the Baumgartner-Weiss-Schindler test, a novel and powerful test based on ranks, is investigated and compared with the parametric t test as well as with two other nonparametric tests (Wilcoxon rank sum test, Fisher-Pitman permutation test) recently recommended for the analysis of gene expression data. RESULTS: Simulation studies show that an exact permutation test based on the Baumgartner-Weiss-Schindler statistic B is preferable to the other three tests. It is less conservative than the Wilcoxon test and more powerful, in particular in case of asymmetric or heavily tailed distributions. When the underlying distribution is symmetric the differences in power between the tests are relatively small. Thus, the Baumgartner-Weiss-Schindler is recommended for the usual situation that the underlying distribution is a priori unknown. AVAILABILITY: SAS code available on request from the authors. PMID- 15284099 TI - The 'subsequent artificial neural network' (SANN) approach might bring more classificatory power to ANN-based DNA microarray analyses. AB - MOTIVATION: Human decisions often proceed in two steps. Initially those most preferred are chosen followed by a subsequent choice of these preferences. Applying one artificial neural network (ANN), a classification is limited to the preselection process. The final categorization is only possible by a subsequent ANN that distinguishes the pre-chosen classes. Existing strategies using coupled ANNs are discussed and a new approach particularly suited for multiclass classification problems is introduced ('Subsequent ANN', SANN). RESULTS: Evaluating a simulated data base comprising 3 classes, classification results of SANN were obviously superior to those achieved by ANN. To evaluate a real-world data base the microarray benchmark GCM (14 classes) was chosen. The ANN results reached 72%, comparable to previous results. Using SANN, up to 81% of the tumors were correctly classified. AVAILABILITY: Programs used in this work and numerical results are available upon request. PMID- 15284100 TI - Accurate detection of aneuploidies in array CGH and gene expression microarray data. AB - MOTIVATION: Chromosomal copy number changes (aneuploidies) are common in cell populations that undergo multiple cell divisions including yeast strains, cell lines and tumor cells. Identification of aneuploidies is critical in evolutionary studies, where changes in copy number serve an adaptive purpose, as well as in cancer studies, where amplifications and deletions of chromosomal regions have been identified as a major pathogenetic mechanism. Aneuploidies can be studied on whole-genome level using array CGH (a microarray-based method that measures the DNA content), but their presence also affects gene expression. In gene expression microarray analysis, identification of copy number changes is especially important in preventing aberrant biological conclusions based on spurious gene expression correlation or masked phenotypes that arise due to aneuploidies. Previously suggested approaches for aneuploidy detection from microarray data mostly focus on array CGH, address only whole-chromosome or whole-arm copy number changes, and rely on thresholds or other heuristics, making them unsuitable for fully automated general application to gene expression datasets. There is a need for a general and robust method for identification of aneuploidies of any size from both array CGH and gene expression microarray data. RESULTS: We present ChARM (Chromosomal Aberration Region Miner), a robust and accurate expectation maximization based method for identification of segmental aneuploidies (partial chromosome changes) from gene expression and array CGH microarray data. Systematic evaluation of the algorithm on synthetic and biological data shows that the method is robust to noise, aneuploidal segment size and P-value cutoff. Using our approach, we identify known chromosomal changes and predict novel potential segmental aneuploidies in commonly used yeast deletion strains and in breast cancer. ChARM can be routinely used to identify aneuploidies in array CGH datasets and to screen gene expression data for aneuploidies or array biases. Our methodology is sensitive enough to detect statistically significant and biologically relevant aneuploidies even when expression or DNA content changes are subtle as in mixed populations of cells. AVAILABILITY: Code available by request from the authors and on Web supplement at http://function.cs.princeton.edu/ChARM/ PMID- 15284101 TI - Primer design and marker clustering for multiplex SNP-IT primer extension genotyping assay using statistical modeling. AB - MOTIVATION: The optimization of the primer design is critical for the development of high-throughput SNP genotyping methods. Recently developed statistical models of the SNP-IT primer extension genotyping reaction allow further improvement of primer quality for the assay. RESULTS: Here we describe how the statistical models can be used to improve primer design for the assay. We also show how to optimize clustering of the SNP markers into multiplex panels using statistical model for multiplex SNP-IT. The primer set failure probability calculated by a model is used as a minimization function for both primer selection and primers clustering. Three clustering algorithms for the multiplex genotyping SNP-IT assay are described and their relative performance is evaluated. We also describe the approaches to improve the speed of primer design and clustering calculations when using the statistical models. Our clustering decreases the average failure probability of the marker set by 7-25%. The experimental marker failure rate in the multiplex reaction was reduced dramatically and success rate can be achieved as high as 96%. AVAILABILITY: The primer design using statistical models is freely available from www.autoprimer.com. PMID- 15284102 TI - DNAFSMiner: a web-based software toolbox to recognize two types of functional sites in DNA sequences. AB - DNAFSMiner (DNA Functional Sites Miner) is a web-based software toolbox to recognize functional sites in nucleic acid sequences. Currently in this toolbox, we provide two software: TIS Miner and Poly(A) Signal Miner. The TIS Miner can be used to predict translation initiation sites in vertebrate DNA/mRNA/cDNA sequences, and the Poly(A) Signal Miner can be used to predict polyadenylation [poly(A)] signals in human DNA sequences. The prediction results are better than those by literature methods on two benchmark applications. This good performance is mainly attributable to our unique learning method. DNAFSMiner is available free of charge for academic and non-profit organizations. AVAILABILITY: http://research.i2r.a-star.edu.sg/DNAFSMiner/ CONTACT: huiqing@i2r.a-star.edu.sg. PMID- 15284103 TI - Modeling interactome: scale-free or geometric? AB - MOTIVATION: Networks have been used to model many real-world phenomena to better understand the phenomena and to guide experiments in order to predict their behavior. Since incorrect models lead to incorrect predictions, it is vital to have as accurate a model as possible. As a result, new techniques and models for analyzing and modeling real-world networks have recently been introduced. RESULTS: One example of large and complex networks involves protein-protein interaction (PPI) networks. We analyze PPI networks of yeast Saccharomyces cerevisiae and fruitfly Drosophila melanogaster using a newly introduced measure of local network structure as well as the standardly used measures of global network structure. We examine the fit of four different network models, including Erdos-Renyi, scale-free and geometric random network models, to these PPI networks with respect to the measures of local and global network structure. We demonstrate that the currently accepted scale-free model of PPI networks fails to fit the data in several respects and show that a random geometric model provides a much more accurate model of the PPI data. We hypothesize that only the noise in these networks is scale-free. CONCLUSIONS: We systematically evaluate how well different network models fit the PPI networks. We show that the structure of PPI networks is better modeled by a geometric random graph than by a scale-free model. SUPPLEMENTARY INFORMATION: Supplementary information is available at http://www.cs.utoronto.ca/~juris/data/data/ppiGRG04/ PMID- 15284104 TI - Exploring protein fold space by secondary structure prediction using data distribution method on Grid platform. AB - MOTIVATION: Since the newly developed Grid platform has been considered as a powerful tool to share resources in the Internet environment, it is of interest to demonstrate an efficient methodology to process massive biological data on the Grid environments at a low cost. This paper presents an efficient and economical method based on a Grid platform to predict secondary structures of all proteins in a given organism, which normally requires a long computation time through sequential execution, by means of processing a large amount of protein sequence data simultaneously. From the prediction results, a genome scale protein fold space can be pursued. RESULTS: Using the improved Grid platform, the secondary structure prediction on genomic scale and protein topology derived from the new scoring scheme for four different model proteomes was presented. This protein fold space was compared with structures from the Protein Data Bank, database and it showed similarly aligned distribution. Therefore, the fold space approach based on this new scoring scheme could be a guideline for predicting a folding family in a given organism. PMID- 15284105 TI - Ontologies for behavior. AB - SUMMARY: Although dozens of biological ontologies have been created and deployed, relatively little attention has been given to using ontologies to represent behavior. Ontologies for two different behavior systems are described here. One ontology was a translation of a published ethogram, and the second was coded from video clips in a comparative study of jumping spider courtship. AVAILABILITY: http://mesquiteproject.org/ontology/. PMID- 15284106 TI - DIVAA: analysis of amino acid diversity in multiple aligned protein sequences. AB - MOTIVATION: Multiple alignments of proteins are an effective way of identifying conserved amino acids that provide clues to functional relationships among proteins. Quantitation of the abundances of amino acids found at each position in a sequence motif can provide a basis for understanding the structural and functional constraints at each point. Distribution of information across a motif has been used previously, but the non-intuitive nature of the analysis has limited its impact. RESULTS: Here, we introduce a quantitative measure of amino acid sequence diversity (DIVAA) that has a simple, intuitive meaning. Diversity, as a measure of sequence conservation or variation, is inextricably linked to the probability of selecting identical pairs from a distribution. We demonstrate its utility through the analysis of four populations: ATP-binding P-loops, hypervariable domains of kappa light chains, signal sequences, and the N- and C- termini of proteins. DIVAA provides a simple means to generate hypotheses concerning the contribution of individual residues to the functional and evolutionary relationships among proteins. AVAILABILITY: Access to DIVAA software is available at RELIC (http://relic.bio.anl.gov). PMID- 15284107 TI - MedlineR: an open source library in R for Medline literature data mining. AB - SUMMARY: We describe an open source library written in the R programming language for Medline literature data mining. This MedlineR library includes programs to query Medline through the NCBI PubMed database; to construct the co-occurrence matrix; and to visualize the network topology of query terms. The open source nature of this library allows users to extend it freely in the statistical programming language of R. To demonstrate its utility, we have built an application to analyze term-association by using only 10 lines of code. We provide MedlineR as a library foundation for bioinformaticians and statisticians to build more sophisticated literature data mining applications. AVAILABILITY: The library is available from http://dbsr.duke.edu/pub/MedlineR. PMID- 15284108 TI - Functional assessment and specific depletion of alloreactive human T cells using flow cytometry. AB - Human T-cell alloreactivity plays an important role in many disease processes, including the rejection of solid organ grafts and graft-versus-host disease (GVHD) following allogeneic stem cell transplantation. To develop a better understanding of the T cells involved in alloreactivity in humans, we developed a cytokine flow cytometry (CFC) assay that enabled us to characterize the phenotypic and functional characteristic of T cells responding to allogeneic stimuli. Using this approach, we determined that most T-cell alloreactivity resided within the CD4(+) T-cell subset, as assessed by activation marker expression and the production of effector cytokines (eg, tumor necrosis factor alpha [TNF]alpha) implicated in human GVHD. Following prolonged stimulation in vitro using either allogeneic stimulator cells or viral antigens, we found that coexpression of activation markers within the CD4(+) T-cell subset occurred exclusively within a subpopulation of T cells that significantly increased their surface expression of CD4. We then developed a simple sorting strategy that exploited these phenotypic characteristics to specifically deplete alloreactive T cells while retaining broad specificity for other stimuli, including viral antigens and third-party alloantigens. This approach also was applied to specifically enrich or deplete human virus-specific T cells. PMID- 15284109 TI - Mutations of the SBDS gene are present in most patients with Shwachman-Diamond syndrome. AB - Shwachman-Diamond Syndrome (SDS) is a rare multisystem disorder characterized by exocrine pancreatic insufficiency, bone marrow dysfunction, and metaphyseal chondrodysplasia. Recent studies show that mutations of SBDS, a gene of unknown function, are present in the majority of patients with SDS. In the present study, we show that most, but not all, patients classified based on rigorous clinical criteria as having SDS had compound heterozygous mutations of SBDS. Full-length SBDS protein was not detected in leukocytes of SDS patients with the most common SBDS mutations, consistent with a loss-of-function mechanism. In contrast, SBDS protein was expressed at normal levels in SDS patients without SBDS mutations. These data confirm the absence of SBDS mutations in this subgroup of patients and suggest that SDS is a genetically heterogeneous disorder. The presence (or absence) of SBDS mutations may define subgroups of patients with SDS who share distinct clinical features or natural history. PMID- 15284110 TI - Distinct roles of ADP receptors in von Willebrand factor-mediated platelet signaling and activation under high flow. AB - We have investigated the role of adenosine diphosphate (ADP) receptors in the adhesion, activation, and aggregation of platelets perfused over immobilized von Willebrand factor (VWF) under high shear stress. Blocking P2Y(1) prevented stable platelet adhesion and aggregation, indicative of a complete inhibition of alpha(IIb)beta(3) activation, and decreased the duration of transient arrests from 5.9 seconds +/- 2.8 seconds to 1.2 seconds +/- 0.8 seconds; in contrast, blocking P2Y(12) inhibited only the formation of larger aggregates. Moreover, blocking P2Y(1) decreased the proportion of platelets showing early intracytoplasmic Ca(++) elevations (alpha/beta peaks) from 20.6% +/- 1.6% to 14.6% +/- 1.5% (P < .01), and the corresponding peak ion concentration from 1543 nM +/- 312 nM to 1037 nM +/- 322 nM (P < .05); it also abolished the Ca(++) elevations seen in firmly attached platelets (gamma peaks). Blocking P2Y(12) had no effect on these parameters, and did not enhance the effect of inhibiting P2Y(1). Inhibition of phospholipase C had similar consequences as the blocking of P2Y(1), whereas inhibition of Src family kinases abolished both type alpha/beta and gamma Ca(++) oscillations, although the former effect required a higher inhibitor concentration. Our results demonstrate that, under elevated shear stress conditions, ADP signaling through P2Y(1) may contribute to the initial stages of platelet adhesion and activation mediated by immobilized VWF, and through P2Y(12) to sustained thrombus formation. PMID- 15284111 TI - Expression and analysis of a split premature termination codon in FGG responsible for congenital afibrinogenemia: escape from RNA surveillance mechanisms in transfected cells. AB - Congenital afibrinogenemia, the most severe form of fibrinogen deficiency, is characterized by the complete absence of fibrinogen. The disease is caused by mutations in 1 of the 3 fibrinogen genes FGG, FGA, and FGB, clustered on the long arm of human chromosome 4. The majority of cases are due to null mutations in the FGA gene although one would expect the 3 genes to be equally implicated. However, most patients studied so far are white, and therefore the identification of causative mutations in non-European families is necessary to establish if this finding holds true in all ethnic groups. In this study, we report the identification of a novel nonsense mutation (Arg134Xaa) in the FGG gene responsible for congenital afibrinogenemia in 10 patients from Lebanon. Expression studies in COS-7 cells demonstrated that the Arg134Xaa codon, which is encoded by adjacent exons (TG-intron 4-A) affected neither mRNA splicing nor stability, but led to the production of an unstable, severely truncated fibrinogen gamma chain that is not incorporated into a functional fibrinogen hexamer. PMID- 15284113 TI - A GTPase-activating protein binds STAT3 and is required for IL-6-induced STAT3 activation and for differentiation of a leukemic cell line. AB - We previously identified a guanosine triphosphatase (GTPase)-activating protein (GAP) male germ cell Rac GAP (MgcRacGAP) that enhanced interleukin-6 (IL-6) induced macrophage differentiation of murine M1 leukemia cells. Later, MgcRacGAP was found to play crucial roles in cell division. However, how MgcRacGAP enhanced IL-6-induced differentiation remained elusive. Here we show that MgcRacGAP enhances IL-6-induced differentiation through enhancement of signal transducer and activator of transcription-3 (STAT3) activation. MgcRacGAP, Rac, and STAT3 formed a complex in IL-6-stimulated M1 cells, where MgcRacGAP interacted with Rac1 and STAT3 through its cysteine-rich domain and GAP domain. In reporter assays, the wild-type MgcRacGAP enhanced transcriptional activation of STAT3 while a GAP-domain deletion mutant (DeltaGAP) did not significantly enhance it, suggesting that the GAP domain was required for enhancement of STAT3-dependent transcription. Intriguingly, M1 cells expressing DeltaGAP had no effect on the differentiation signal of IL-6, while forced expression of MgcRacGAP rendered M1 cells hyperresponsive to the IL-6-induced differentiation. Moreover, knockdown of MgcRacGAP by RNA interference profoundly suppressed STAT3 activation, implicating MgcRacGAP in the STAT3-dependent transcription. All together, our data not only reveal an important role for MgcRacGAP in STAT3 activation, but also demonstrate that MgcRacGAP regulates IL-6-induced cellular differentiation in which STAT3 plays a pivotal role. PMID- 15284112 TI - The addition of rituximab to a combination of fludarabine, cyclophosphamide, mitoxantrone (FCM) significantly increases the response rate and prolongs survival as compared with FCM alone in patients with relapsed and refractory follicular and mantle cell lymphomas: results of a prospective randomized study of the German Low-Grade Lymphoma Study Group. AB - In follicular lymphoma (FL) and mantle cell lymphoma (MCL) the monoclonal antibody rituximab may improve the prognosis when combined with chemotherapy. This was investigated in a prospective randomized study in patients with relapsed disease. A total of 147 patients were randomized to receive 4 courses of chemotherapy with 25 mg/m(2) fludarabine on days 1 to 3, 200 mg/m(2) cyclophosphamide on days 1 to 3, and 8 mg/m(2) mitoxantrone on day 1 (FCM), alone or combined with rituximab (375 mg/m(2); R-FCM). Of 128 evaluable patients, 62 were randomized for FCM and 66 for R-FCM. R-FCM revealed an overall response rate of 79% (33% complete remission [CR], 45% partial remission [PR]) as compared with 58% for FCM alone (13% CR, 45% PR; P = .01), with similar results in a subgroup analysis of FL (94% vs 70%) and MCL (58% vs 46%). In the total group, the R-FCM arm was significantly superior concerning progression-free survival (PFS; P = .0381) and overall survival (OS; P = .0030). In FL PFS was significantly longer in the R-FCM arm (P = .0139) whereas in MCL a significantly longer OS was observed (P = .0042). There were no differences in clinically relevant side effects in both study arms. Hence, the addition of rituximab to FCM chemotherapy significantly improves the outcome of relapsed or refractory FL and MCL. PMID- 15284114 TI - Determination of relapse risk based on assessment of minimal residual disease during complete remission by multiparameter flow cytometry in unselected patients with acute myeloid leukemia. AB - Quantification of minimal residual disease (MRD) reveals significant prognostic information in patients treated for acute myeloid leukemia (AML). The application of multiparameter flow cytometry (MFC) for MRD assessment has resulted in significant prognostic information in selected cases in previous analyses. We analyzed MRD in unselected patients with AML in complete remission (CR) after induction (n = 58) and consolidation (n = 62) therapies. By using a comprehensive panel of monoclonal antibodies we identified at least one leukemia-associated aberrant immunophenotype (LAIP) in each patient. The degree of reduction between diagnosis and CR in LAIP-positive cells (log difference [LD]) as a continuous variable was significantly related to relapse-free survival (RFS) both after induction (P = .0001) and consolidation (P = .000 08) therapies, respectively. The LD determined after consolidation therapy was the only parameter related to overall survival (OS) (P = .005). Separation of patients based on the 75th percentile of LD after consolidation therapy resulted in groups with highly different RFS (83.3% versus 25.7%, P = .0034) and OS (87.5% versus 51.4%, P = .0507) at 2 years. Multivariate analysis identified LD as an independent prognostic factor for RFS at both checkpoints. MFC-based quantification of MRD reveals important prognostic information in unselected patients with AML in addition to cytogenetics and should be further evaluated and used in clinical trials. PMID- 15284115 TI - Hairy cell leukemia: at the crossroad of somatic mutation and isotype switch. AB - Hairy cell leukemia (HCL) commonly expresses multiple immunoglobulin isotypes, a feature rare in other B-cell malignancies or in normal B cells. In HCL, there is no phenotypic evidence for subpopulations, and single cells from one previous case contained transcripts for several isotypes. This raises the questions of the differentiation status of the cell of origin and of posttransformation events. We have investigated 9 cases, all expressing multiple immunoglobulin isotypes. Multiple tumor-derived variable-(diversity)-joining-constant mu delta, gamma, alpha (V(D)J-Cmu, delta, gamma, alpha) transcripts were confirmed in single cells of a further case. All cases were negative for germinal center (GC)-associated markers CD27 and CD38. Seven of 9 cases had mutated V(H) genes, with low levels of intraclonal heterogeneity, but 2 of 9 were unmutated, indicative of pre-GC origin. Eight of 9 cases expressed activation-induced cytidine deaminase (AID), a molecule essential for somatic mutation and isotype switch. All cases expressed germ line heavy-chain I exon (I(H))-C(H) transcripts which paralleled surface immunoglobulin (sIg) isotype. Significantly, no circle transcripts indicative of deletional recombination of switched isotypes were detectable in 9 of 9 cases. These data indicate heterogeneity in the cell of origin in terms of mutational status, but reveal common features of AID expression and isotype-switching events occurring prior to deletional recombination. Both mutational and switching events may be influenced by environmental factors at extrafollicular sites. PMID- 15284116 TI - Endocytosis, intracellular sorting, and processing of exosomes by dendritic cells. AB - Exosomes are nanovesicles released by leukocytes and epithelial cells. Although their function remains enigmatic, exosomes are a source of antigen and transfer functional major histocompatibility complex (MHC)-I/peptide complexes to dendritic cells (DCs) for CD8(+) T-cell activation. Here we demonstrate that exosomes also are internalized and processed by immature DCs for presentation to CD4(+) T cells. Endocytosed exosomes are sorted into the endocytic compartment of DCs for processing, followed by loading of exosome-derived peptides in MHC-II molecules for presentation to CD4(+) T cells. Targeting of exosomes to DCs is mediated via milk fat globule (MFG)-E8/lactadherin, CD11a, CD54, phosphatidylserine, and the tetraspanins CD9 and CD81 on the exosome and alpha(v)/beta(3) integrin, and CD11a and CD54 on the DCs. Circulating exosomes are internalized by DCs and specialized phagocytes of the spleen and by hepatic Kupffer cells. Internalization of blood-borne allogeneic exosomes by splenic DCs does not affect DC maturation and is followed by loading of the exosome-derived allopeptide IEalpha(52-68) in IA(b) by host CD8alpha(+) DCs for presentation to CD4(+) T cells. These data imply that exosomes present in circulation or extracellular fluids constitute an alternative source of self- or allopeptides for DCs during maintenance of peripheral tolerance or initiation of the indirect pathway of allorecognition in transplantation. PMID- 15284117 TI - Erythroblasts secrete the nonclassical HLA-G molecule from primitive to definitive hematopoiesis. AB - The initial steps of primitive hematopoiesis and endothelial vascular formation in the human embryo remain to be defined. Here, we report the identification of a novel marker, namely the nonclassical HLA-G class I molecule, which targets both primitive erythroid cells of the yolk sac and endothelial cells from developing vessels. Moreover, HLA-G was present in its soluble form in the erythropoietic lineage in all organs sustaining primitive to definitive erythropoiesis (ie, aorta-gonad-mesonephros, liver, spleen, and bone marrow). The alternatively spliced transcript coding the soluble HLA-G5 molecule was detected in erythroid cells. The corresponding intron 4-retaining 37-kDa HLA-G5 isoform was secreted from the erythroid progenitor stage to the reticulocyte but was lost in mature erythrocytes and in endothelial cells from differentiated vessels. This study constitutes the first description of an HLA class I antigen expression on the primitive erythroid lineage and provides a way of seeking both primitive and definitive erythropoiesis using HLA-G5. This new marker, previously known by its immunotolerogeneic properties, may be involved in erythroid differentiation, angiogenesis, or both. PMID- 15284118 TI - FIP1L1-PDGFRA fusion: prevalence and clinicopathologic correlates in 89 consecutive patients with moderate to severe eosinophilia. AB - A novel oncogenic mutation (FIP1L1-PDGFRA), which results in a constitutively activated platelet-derived growth factor receptor-alpha (PDGFRA), has been invariably associated with a primary eosinophilic disorder. The current study examines both the prevalence and the associated clinicopathologic features of this mutation in a cohort of 89 adult patients presenting with an absolute eosinophil count (AEC) of higher than 1.5 x 10(9)/L. A fluorescence in situ hybridization (FISH)-based strategy was used to detect FIP1L1-PDGFRA in bone marrow cells. None of 8 patients with reactive eosinophilia displayed the abnormality, whereas the incidence of FIP1L1-PDGFRA in the remaining 81 patients with primary eosinophilia was 14% (11 patients). None (0%) of 57 patients with the hypereosinophilic syndrome (HES) but 10 (56%) of 19 patients with systemic mast cell disease associated with eosinophilia (SMCD-eos) carried the specific mutation. The bone marrow mast cell infiltration pattern in FIP1L1-PDGFRA(+) SMCD eos was distinctly diffuse with loose tumoral aggregates. Treatment with low-dose imatinib (100 mg/d) produced complete and durable responses in all 8 FIP1L1 PDGFRA(+) cases treated. In contrast, only 40% partial response rate was seen in 10 HES cases. FIP1L1-PDGFRA is a relatively infrequent but treatment-relevant mutation in primary eosinophilia that is indicative of an underlying systemic mastocytosis. PMID- 15284119 TI - Monocyte-like and mature macrophages produce CXCL13 (B cell-attracting chemokine 1) in inflammatory lesions with lymphoid neogenesis. AB - The homeostatic chemokine CXCL13 (also called B cell-attracting chemokine 1 [BCA 1] or B-lymphocyte chemoattractant [BLC]) is constitutively expressed in secondary lymphoid tissue and initiates lymphoid neogenesis when expressed aberrantly in mice. CXCL13 has also been detected in chronic inflammation associated with human lymphoid neogenesis, suggesting a pathogenic role. Follicular dendritic cells (FDCs) are generally considered to be the major source of CXCL13 both in normal and aberrant lymphoid tissue. We show here, instead, that most CXCL13-expressing cells in rheumatoid arthritis and ulcerative colitis are of monocyte/macrophage lineage. They are located in irregular lymphoid aggregates within an FDC network, but also within and near smaller collections of B cells in diseased tissue where no FDCs are detected. Some of these CXCL13 expressing cells are CD14(+), suggesting derivation from recently extravasated monocytes. Interestingly, monocytes from healthy donors stimulated in vitro with lipopolysaccharide secrete CXCL13. This induced production is enhanced after in vitro maturation of the monocytes toward macrophages but markedly decreased after maturation toward dendritic cells. Together, our findings strongly suggest that newly recruited monocytes/macrophages play a role for lymphoid neogenesis in human inflammatory diseases. Circulating monocytes are therefore potential candidates for future targeted therapy of chronic inflammation. PMID- 15284120 TI - SDF-1 involvement in endothelial phenotype and ischemia-induced recruitment of bone marrow progenitor cells. AB - Chemokine stromal derived factor 1 (SDF-1) is involved in trafficking of hematopoietic stem cells (HSCs) from the bone marrow (BM) to peripheral blood (PB) and has been found to enhance postischemia angiogenesis. This study was aimed at investigating whether SDF-1 plays a role in differentiation of BM derived c-kit(+) stem cells into endothelial progenitor cells (EPCs) and in ischemia-induced trafficking of stem cells from PB to ischemic tissues. We found that SDF-1 enhanced EPC number by promoting alpha(2), alpha(4), and alpha(5) integrin-mediated adhesion to fibronectin and collagen I. EPC differentiation was reduced in mitogen-stimulated c-kit(+) cells, while cytokine withdrawal or the overexpression of the cyclin-dependent kinase (CDK) inhibitor p16(INK4) restored such differentiation, suggesting a link between control of cell cycle and EPC differentiation. We also analyzed the time course of SDF-1 expression in a mouse model of hind-limb ischemia. Shortly after femoral artery dissection, plasma SDF 1 levels were up-regulated, while SDF-1 expression in the bone marrow was down regulated in a timely fashion with the increase in the percentage of PB progenitor cells. An increase in ischemic tissue expression of SDF-1 at RNA and protein level was also observed. Finally, using an in vivo assay such as injection of matrigel plugs, we found that SDF-1 improves formation of tubulelike structures by coinjected c-kit(+) cells. Our findings unravel a function for SDF 1 in increase of EPC number and formation of vascular structures by bone marrow progenitor cells. PMID- 15284121 TI - Rh autoantigen presentation to helper T cells in chronic lymphocytic leukemia by malignant B cells. AB - Chronic lymphocytic leukemia (CLL) is frequently associated with autoimmune diseases directed against constituents of the blood, including hemolytic anemia (AIHA). We hypothesized that CLL cells predispose to hematologic autoimmunity by acting as aberrant antigen-presenting cells (APCs). Initially, it was confirmed that all studied patients with AIHA secondary to CLL harbored activated helper T (T(H)) cells specific for epitopes on the dominant red blood cell (RBC) autoantigens in primary AIHA, the Rh proteins. Rh-specific T(H) cells were also detected in a number of patients with CLL who, although they did not have AIHA, had low levels of anti-RBC antibody in their sera. Fractionation of putative APC populations from the peripheral blood of patients by negative selection showed that CD5+ CLL cells are the most effective cell type in processing and presenting purified Rh protein to autoreactive T(H) cells. This ability was confirmed using positively selected CD5+ CLL cells. Thus, our study provides the first evidence for malignant cells driving an autoimmune response by acting as aberrant APCs. PMID- 15284122 TI - Mutations of the Wiskott-Aldrich Syndrome Protein (WASP): hotspots, effect on transcription, and translation and phenotype/genotype correlation. AB - The Wiskott-Aldrich syndrome (WAS) is an X-linked recessive immune deficiency disorder characterized by thrombocytopenia, small platelet size, eczema, recurrent infections, and increased risk of autoimmune disorders and malignancies. X-linked thrombocytopenia (XLT) is an allelic variant of WAS which presents with a milder phenotype, generally limited to thrombocytopenia. WAS and XLT are caused by mutations of the Wiskott-Aldrich syndrome protein (WASP) gene which encodes a 502-amino acid protein, named WASP. WASP is thought to play a role in actin cytoskeleton organization and cell signaling. Here, we report the identification of 141 unique mutations, 71 not previously reported, from 227 WAS/XLT families with a total of 262 affected members. When possible we studied the effects of these mutations on transcription, RNA splicing, and protein expression. By analyzing a large number of patients with WAS/XLT at the molecular level we identified 5 mutational hotspots in the WASP gene and have been able to establish a strong association between genotype and phenotype. PMID- 15284123 TI - Epigenetic silencing of the immunoglobulin heavy-chain gene in classical Hodgkin lymphoma-derived cell lines contributes to the loss of immunoglobulin expression. AB - Immunoglobulin production is impaired in Hodgkin and Reed-Sternberg (HRS) cells of classical Hodgkin lymphoma (cHL) in spite of functional clonal rearrangements. The presence of "crippling" mutations in coding and regulatory regions, as well as down-regulation of B-cell-specific transcription factors, has been suggested as a potential reason for the lack of immunoglobulin (Ig) chain gene transcription. We have investigated the impact of epigenetic silencing in suppressing Ig heavy (H)-chain expression. Chromatin immunoprecipitation (ChIP) was used to analyze transcription factor binding to octamer motifs present in the IgH regulatory regions. Transcription factors were bound to these motifs in control cell lines, however, they were absent in the cHL-derived cell lines KMH2, L1236, and L428. Ectopic expression of octamer-binding transcription factor (Oct2) and/or B-cell Oct binding protein/Oct-binding factor (BOB.1/OBF.1) did not result in any measurable binding to these sites. Increased histone 3 Lysine 9 (H3 K9) methylation was observed in the promoter region of the IgH locus in L428 and L1236 cells. This is a typical feature of heterochromatic, transcriptionally silent regions. Treatment of cHL-derived cell lines with the DNA demethylating agent 5-aza-2'-deoxycytidine (5-aza-dC) partially reactivated IgH transcription and affected chromatin modifications. Our results suggest an important role of epigenetic silencing in the inhibition of IgH transcription in HRS cells. PMID- 15284124 TI - Clinical research under the cosh again. PMID- 15284126 TI - Fine needle aspiration cytology in cancer diagnosis. PMID- 15284127 TI - The future of psychotherapy in the NHS. PMID- 15284128 TI - Scandals have eroded US public's confidence in drug industry. PMID- 15284130 TI - Report calls for more care for people wanting to die at home. PMID- 15284131 TI - WHO wants more palliative care for Europeans. PMID- 15284132 TI - French surgeons are set to strike. PMID- 15284133 TI - Sexually transmitted diseases continue to rise. PMID- 15284135 TI - Life expectancy in Great Britain rises--but later years are still spent in poor health. PMID- 15284136 TI - Bush signs law to protect US from bioterrorism. PMID- 15284138 TI - Rape victims in Sudan face life of stigma, says report. PMID- 15284139 TI - Fertilisation authority rules in favour of embryo selection for tissue donation. PMID- 15284140 TI - Government confirms second case of vCJD transmitted by blood transfusion. PMID- 15284141 TI - WHO uses work on tuberculosis in Lima as model for tackling AIDS. PMID- 15284143 TI - Audit report criticises India's slow progress on AIDS. PMID- 15284146 TI - New US Medicare policy changes ruling that obesity is not an illness. PMID- 15284148 TI - Indian doctor's decision to "self test" AIDS vaccine decried as unethical. PMID- 15284149 TI - Photo-onycholysis associated with the use of doxycycline. PMID- 15284150 TI - Advice to parents has limited effect--where next? PMID- 15284151 TI - Corticosteroids for HELLP (haemolysis, elevated liver enzymes, low platelets) syndrome. PMID- 15284153 TI - Burns reconstruction. PMID- 15284154 TI - Obstacles to conducting epidemiological research in the UK general population. PMID- 15284155 TI - The other face of research governance. PMID- 15284156 TI - View from the research and development office. PMID- 15284157 TI - Bureaucracy of ethics applications. PMID- 15284158 TI - Better support for investigators is essential. PMID- 15284159 TI - Research ethics paperwork: what is the plot we seem to have lost? PMID- 15284160 TI - Ethical review of research into rare genetic disorders. PMID- 15284161 TI - Fine needle aspiration of hepatic colorectal metastases: BMJ enters arena of tabloid journalism. PMID- 15284162 TI - Fine needle aspiration of hepatic colorectal metastases: damage is done now. PMID- 15284163 TI - Fine needle aspiration of hepatic colorectal metastases: fine needle aspiration has its place. PMID- 15284164 TI - Social class and elective caesareans in the NHS: analysis is not really about social class. PMID- 15284165 TI - Social class and elective caesareans in the NHS: Dr Foster is cheap and offensive. PMID- 15284166 TI - Treatment of minor burns: dressings do not need to stick. PMID- 15284168 TI - Bush plans to screen whole US population for mental illness: an unlikely public healthcare initiative. PMID- 15284169 TI - Bush plans to screen whole US population for mental illness: Bush's sanity test is revealed. PMID- 15284170 TI - Bush plans to screen whole US population for mental illness: is road of initiative paved with good intentions? PMID- 15284171 TI - Treatment of minor burns: treatment of burns is controversial. PMID- 15284172 TI - Good news is often ignored. PMID- 15284173 TI - Hope of prevention training in South Asia. PMID- 15284174 TI - Campaign to revitalise academic medicine: don't believe us. PMID- 15284175 TI - Campaign to revitalise academic medicine: is the bubble due to burst for medical research funding? PMID- 15284176 TI - Doctors are not scientists but we still need science. PMID- 15284177 TI - Inhibition of human lung cancer cell growth by angiotensin-(1-7). AB - Angiotensin-(1-7) [Ang-(1-7)] is an endogenous peptide hormone of the renin angiotensin system with vasodilator and anti-proliferative properties. Human adenocarcinoma SK-LU-1 and A549 cells as well as non-small lung cancer SK-MES-1 cells were treated with serum in the presence and absence of Ang-(1-7), to determine whether Ang-(1-7) inhibits the growth of lung cancer cells. Ang-(1-7) caused a significant reduction in serum-stimulated growth in all three lung cancer cell lines. Treatment with Ang-(1-7) resulted in both a dose- and time dependent reduction in serum-stimulated DNA synthesis in all three cell lines, with IC(50)'s in the sub-nanomolar range. The Ang-(1-7) receptor antagonist [D Ala(7)]-Ang-(1-7) blocked the attenuation of the serum-stimulated DNA synthesis of SK-LU-1 cells by Ang-(1-7), while neither AT(1) nor AT(2) angiotensin receptor subtype antagonists prevented the response to the heptapeptide. MAS mRNA and protein, a receptor for Ang-(1-7), was detected in the three lung cancer cell lines, suggesting that the anti-proliferative effect of Ang-(1-7) in the cancer cells may be mediated by the non-AT(1), non-AT(2), AT((1-7)) receptor MAS. Other angiotensin peptides [Ang I, Ang II, Ang-(2-8), Ang-(3-8) and Ang-(3-7)] did not attenuate mitogen-stimulated DNA synthesis of SK-LU-1 cells, demonstrating that Ang-(1-7) selectively inhibits SK-LU-1 cancer cell growth. Pre-treatment of SK-LU 1 cells with 10 nM Ang-(1-7) reduced serum-stimulated phosphorylation of extracellular signal-regulated kinase (ERK)1 and ERK2, indicating that the anti proliferative effects may occur, at least in part, through inhibition of the ERK signal transduction pathway. The results of this study suggest that Ang-(1-7) inhibits lung cancer cell growth through the activation of an angiotensin peptide receptor and may represent a novel chemotherapeutic and chemopreventive treatment for lung cancer. PMID- 15284178 TI - Habitual consumption of fruits and vegetables: associations with human rectal glutathione S-transferase. AB - The glutathione (GSH)/glutathione S-transferase (GST) system is an important detoxification system in the gastrointestinal tract. A high activity of this system may benefit cancer prevention. The aim of the study was to assess whether habitual consumption of fruits and vegetables, especially citrus fruits and brassica and allium vegetables, is positively associated with parameters reflecting the activity of the GSH/GST enzyme system in human rectal mucosa. GST enzyme activity, GST isoenzyme levels of GST-alpha (A1-1, A1-2 and A2-2), -mu (M1 1) and -pi (P1-1), and GSH levels were measured in rectal biopsies from 94 subjects. Diet, lifestyle, GSTM1 and GSTT1 null polymorphisms were assessed. Mean GST enzyme activity was 237 nmol/min/mg protein (SD = 79). Consumption of citrus fruits was positively associated with GST enzyme activity [difference between high and low consumption: 28.9 (95% confidence interval (CI) = 9.3-48.6) nmol/min/mg protein], but was not associated with the other parameters. A positive association with brassica vegetables was found among carriers of the GSTM1-plus genotype [difference between high and low consumption: 22.6 (95% CI = 0.2-45.0) nmol/min/mg protein], but not among GSTM1-null individuals (-25.8 nmol/min/mg protein, 95% CI = -63.3-11.8). This is in line with a positive association between consumption of brassica vegetables and GSTM isoenzyme level [difference between high and low consumption: 67.5%, 95% CI = (6.8-162.7)]. Consumption of allium vegetables was not associated with GST enzyme activity, but negatively with GSTP1-1 levels [difference between high and low consumption: 23.3%, 95% CI = (-35.5; -8.6)]. Associations were similar among those with the GSTT1-plus and GSTT1-null genotype. In conclusion, variations in habitual consumption of fruits, particularly citrus fruits, and of vegetables, in particular brassica vegetables, among those with the GSTM1-plus genotype, may contribute to variations in human rectal GST enzyme activity. PMID- 15284179 TI - Oxidative damage-related genes AKR1C3 and OGG1 modulate risks for lung cancer due to exposure to PAH-rich coal combustion emissions. AB - Lung cancer rates among men and particularly among women, almost all of whom are non-smokers, in Xuan Wei County, China are among the highest in China and have been causally associated with exposure to indoor smoky coal emissions that contain very high levels of polycyclic aromatic hydrocarbons (PAHs). As such, this population provides a unique opportunity to study the pathogenesis of PAH induced lung cancer that is not substantially influenced by the large number of other carcinogenic constituents of tobacco smoke. Aldo-keto reductases (AKRs) activate PAH dihydrodiols to yield their corresponding reactive and redox-active o-quinones, which can then generate reactive oxygen species that cause oxidative DNA damage. We therefore examined the association between single nucleotide polymorphisms (SNPs) in four genes (AKR1C3-Gln5His, NQO1-Pro187Ser, MnSOD Val16Ala and OGG1-Ser326Cys) that play a role in the generation, prevention or repair of oxidative damage and lung cancer risk in a population-based, case control study of 119 cases and 113 controls in Xuan Wei, China. The AKR1C3 Gln/Gln genotype was associated with a 1.84-fold [95% confidence interval (CI) = 0.98-3.45] increased risk and the combined OGG1-Cys/Cys and Ser/Cys genotypes were associated with a 1.93-fold (95% CI = 1.12-3.34) increased risk of lung cancer. Subgroup analysis revealed that the effects were particularly elevated among women who had relatively high cumulative exposure to smoky coal. SNPs in MnSOD and NQO1 were not associated with lung cancer risk. These results suggest that SNPs in the oxidative stress related-genes AKR1C3 and OGG1 may play a role in the pathogenesis of lung cancer in this population, particularly among heavily exposed women. However, due to the small sample size, additional studies are needed to evaluate these associations within Xuan Wei and other populations with substantial exposure to PAHs. PMID- 15284180 TI - Generation of S phase-dependent DNA double-strand breaks by Cr(VI) exposure: involvement of ATM in Cr(VI) induction of gamma-H2AX. AB - Certain hexavalent chromium [Cr(VI)] compounds are implicated as occupational respiratory carcinogens. Cr(VI) induces a broad spectrum of DNA damage, but Cr(VI)-induced DNA double-strand breaks (DSBs) have not been reported. Previously we found that Cr(VI) activates the ataxia telangiectasia mutated (ATM) kinase. ATM is activated specifically in response to DSBs. Therefore, the objective of this study was to investigate DSB induction by Cr(VI) exposure with the overarching hypothesis that S phase-dependent DSBs are produced by Cr(VI) exposure. To test this hypothesis, normal human fibroblasts were treated with either Cr(VI) or neocarzinostatin (NCS). DSBs were analyzed by both comet assay under neutral conditions, which detects primarily DNA DSBs, and phosphorylation of histone H2AX (gamma-H2AX) and the resultant formation of nuclear foci, which are considered to be indicative of DSBs. Induction of DSBs was observed after Cr(VI) exposure, however, the Cr(VI)-induced DSBs were abrogated by G(1) synchronization. Furthermore, our data showed that Cr(VI)-induced DSBs were only observed in the S phase population, whereas no significant DSBs were observed in Cr(VI)-treated G(1) synchronized cells. In contrast, NCS-induced DSBs were equally distributed in all cell cycle phases in both asynchronous and G(1) synchronized cells. Moreover, Cr(VI)-induced gamma-H2AX foci formation was restricted to PCNA-positive cells, whereas NCS-induced gamma-H2AX foci formed in both PCNA-positive and PCNA-negative cells. These results indicate that Cr(VI) induced DSBs are S phase-dependent. Finally, our data showed that Cr(VI)-induced gamma-H2AX production was significantly decreased in ATM(-/-) cells compared with ATM(+/+) cells. Taken together, these results suggest that Cr(VI)-induced activation of ATM involves the formation of S phase-dependent DSBs. Examining the mechanism of Cr(VI)-induced DSBs will aid in understanding the interrelated mechanisms of Cr(VI) toxicity and carcinogenesis. PMID- 15284181 TI - Mutant K-rasV12 increases COX-2, peroxides and DNA damage in lung cells. AB - K-ras is frequently mutated in lung adenocarcinomas. Recent discovery that wild type K-ras is tumor suppressive in the lung raises a question: how is mutant K ras aggressively oncogenic? We hypothesized that mutant K-ras might lead to generation of reactive oxygen species (ROS) and DNA damage, contributing to malignant transformation. We stably transfected human mutant K-ras(V12) into non transformed peripheral mouse lung epithelial cells (E10 line). Constitutively active mutant K-ras(V12) in E10 cells led to a highly significant (P < 0.001) increased level of peroxides, and a corresponding increase in the amount of DNA strand-break damage, compared with the parental line E10 and the vector control. Levels of superoxide were not increased, suggesting a direct source of peroxides, such as cyclooxygenase-2 (COX-2). COX-2 protein and activity measured as prostaglandin E(2) level were up-regulated in cells expressing mutant K-ras(V12); COX-2 activity correlated with K-ras activity (K-ras p21-GTP). Both peroxide generation and DNA single strand breaks were significantly reduced by pre treatment with COX-2-specific inhibitor SC 58125, confirming COX-2 as the source of the ROS. COX-2 has been repeatedly implicated in lung cancer, and is known to be regulated by ras and to release ROS. Our data suggest that up-regulation of COX-2, with a consequent increase in peroxides and DNA damage, contributes to the dominant oncogenicity of mutant K-ras. PMID- 15284182 TI - IL-6-induced survival of colorectal carcinoma cells is inhibited by butyrate through down-regulation of the IL-6 receptor. AB - Colorectal carcinoma cells are characterized by over-expression of IL-6 and the IL-6 receptor, an autocrine loop that promotes the development of many tumors. To determine the importance of this pathway, we examined the role that IL-6 plays in the biology of 228 and RKO colorectal tumor cells. IL-6 induced prominent tyrosine phosphorylation of the transcription factor STAT1 in both cell types. Furthermore, IL-6 exerts functional effects in these cells in that it inhibited apoptosis induced by Fas ligation, and up-regulated Bcl-xl, a STAT target gene, which can promote cell survival. Butyrate, a compound formed in the intestines of people who consume a high-fiber diet, may confer protection against the development of colorectal cancer. Given the potential importance of IL-6 in the pathogenesis of colorectal tumors, we tested the hypothesis that butyrate acts by inhibiting IL-6-induced signaling events in colorectal carcinoma cells. Following treatment with butyrate, the activation of STAT1 in response to IL-6, but not interferon-gamma, was completely lost. Butyrate induced a prominent decrease of mRNA and cell surface expression of the IL-6 receptor alpha (IL-6Ralpha) chain. Introduction of a soluble form of the IL-6Ralpha chain restored IL-6-induced STAT1 activation and resistance to apoptosis of butyrate treated cells. These experiments indicate that IL-6 may play an important role in the pathogenesis of colorectal cancers, and that butyrate may exert its protective effect by specifically blocking IL-6-induced signaling events. PMID- 15284183 TI - Association of the thymidylate synthase polymorphisms with esophageal squamous cell carcinoma and gastric cardiac adenocarcinoma. AB - Polymorphisms in the untranslated regions (UTRs) of the thymidylate synthase (TS) gene, which may modulate TS transcription and expression, have been associated with susceptibility and prognosis of several tumors. However, their effects on the development and clinical staging of esophageal squamous cell carcinoma (ESCC) and gastric cardiac adenocarcinoma (GCA) have not been assessed so far. In this study, the 28-bp tandem repeat and the G/C single nucleotide polymorphism in the TS 5'UTR, the 6-bp deletion (6 bp-) polymorphism in the TS 3'UTR, were genotyped in 465 cancer patients (232 ESCC, 233 GCA) and 348 control subjects in North China. The genotype and allelotype distribution of the TS variants in ESCC, GCA patients and controls did not show significant difference. However, the frequency of the 6 bp-/2R haplotype in ESCC and GCA patients was marginally or significantly lower than that in controls (P = 0.05 and 0.006, respectively). Thus, the 6 bp-/2R significantly reduced the risk to ESCC and GCA, compared with the 6 bp-/3G haplotype [odds ratio (OR) = 0.61 and 0.48, 95% confidence interval (CI) = 0.37-1.00 and 0.28-0.81, respectively]. In addition, the 6 bp+/3G haplotype in ESCC patients was also significantly less common than in controls (P = 0.002). Compared with the 6 bp-/3G haplotype, the 6 bp+/3G significantly reduced the risk to ESCC (OR = 0.30, 95% CI = 0.14-0.67). Moreover, the TS 2R/3G genotype frequency in ESCC patients with and without lymphatic metastasis was significantly different (27.1 versus 4.9%, P < 0.001). Therefore, the 2R/3G genotype had an approximately 11-fold increase in the risk of lymphatic metastasis of ESCC, compared with the 3G/3G genotype (95% CI = 2.67-49.74). The results suggested that the TS polymorphisms might be associated with the susceptibility to ESCC and GCA, and the 2R/3G genotype might be a candidate marker to predict the potential of lymphatic metastasis in ESCC. PMID- 15284184 TI - The role of human papillomavirus in oral carcinogenesis. AB - Human papillomavirus (HPV) infection with high-risk types 16 and 18 has widely been reported as one of the prominent mechanisms behind the development of cervical squamous cell carcinoma. Links between HPV and oral cavity cancer have been suggested as well, based on epidemiologic and molecular means, though the association is less well-established. It is likely that HPV plays a role in oral cavity carcinogenesis, though only in a small subset of cases. The difficulty in providing true causal evidence of HPV's role in oral cancer lies in our lack of understanding of the significance of mechanisms by which HPV leads to oral carcinogenesis, as well as limitations in the molecular analysis of HPV. Further studies are necessary for the contribution of HPV in oral cavity malignancy to be better demonstrated. PMID- 15284185 TI - The embryonic origins of left-right asymmetry. AB - The bilaterally symmetric body plan of vertebrates features several consistent asymmetries in the placement, structure, and function of organs such as the heart, intestine, and brain. Deviations from the normal pattern result in situs inversus, isomerisms, or heterotaxia (independent randomization), which have significant clinical implications. The invariance of the left-right (LR) asymmetry of normal morphology, neuronal function, and phenotype of several syndromes raises fascinating and fundamental questions in cell, developmental, evolutionary, and neurobiology. While a pathway of asymmetrically expressed signaling factors has been well-characterized in several model systems, very early steps in the establishment of LR asymmetry remain poorly understood. In particular, the origin of consistently oriented asymmetry is unknown. Recently, a candidate for the origins of asymmetry has been suggested: bulk transport of extracellular morphogens by rotating primary cilia during gastrulation. This model is appealing because it 'bootstraps' morphological asymmetry of the embryo from the intrinsic structural (molecular) chirality of motile cilia. However, conceptual and practical problems remain with this hypothesis. Indeed, the genetic data are also consistent with a different mechanism: cytoplasmic transport roles of motor proteins. This review outlines the progress and remaining questions in the field of left-right asymmetry, and focuses on an alternative model for 'Step 1' of asymmetry. More specifically, based on wide ranging data on ion fluxes and motor protein function in several species, it is suggested that laterality is driven by pH/voltage gradients across the midline, which are established by chiral movement of motor proteins with respect to the cytoskeleton. PMID- 15284186 TI - Chemotaxis-guided movements in bacteria. AB - Motile bacteria often use sophisticated chemotaxis signaling systems to direct their movements. In general, bacterial chemotactic signal transduction pathways have three basic elements: (1) signal reception by bacterial chemoreceptors located on the membrane; (2) signal transduction to relay the signals from membrane receptors to the motor; and (3) signal adaptation to desensitize the initial signal input. The chemotaxis proteins involved in these signal transduction pathways have been identified and extensively studied, especially in the enterobacteria Escherichia coli and Salmonella enterica serovar typhimurium. Chemotaxis-guided bacterial movements enable bacteria to adapt better to their natural habitats via moving toward favorable conditions and away from hostile surroundings. A variety of oral microbes exhibits motility and chemotaxis, behaviors that may play important roles in bacterial survival and pathogenesis in the oral cavity. PMID- 15284187 TI - Adverse drug reactions in the orofacial region. AB - A wide spectrum of drugs can sometimes give rise to numerous adverse orofacial manifestations, particularly dry mouth, taste disturbances, oral mucosal ulceration, and/or gingival swelling. There are few relevant randomized double blind controlled studies in this field, and therefore this paper reviews the data from case reports, small series, and non-peer-reviewed reports of adverse drug reactions affecting the orofacial region (available from a MEDLINE search to April, 2003). The more common and significant adverse orofacial consequences of drug therapy are discussed. PMID- 15284188 TI - Implantology and the severely resorbed edentulous mandible. AB - Patients with a severely resorbed edentulous mandible often suffer from problems with the lower denture. These problems include: insufficient retention of the lower denture, intolerance to loading by the mucosa, pain, difficulties with eating and speech, loss of soft-tissue support, and altered facial appearance. These problems are a challenge for the prosthodontist and surgeon. Dental implants have been shown to provide a reliable basis for fixed and removable prostheses. This has resulted in a drastic change in the treatment concepts for management of the severely resorbed edentulous mandible. Reconstructive, pre prosthetic surgery has changed from surgery aimed to provide a sufficient osseous and mucosal support for a conventional denture into surgery aimed to provide a sufficient bone volume enabling implants to be placed at the most optimal positions from a prosthetic point of view. The aim of this paper is to review critically the literature on procedures related to the severely resorbed edentulous mandible and dental implant treatment. The study includes the transmandibular implant, (short) endosseous implants, and reconstructive procedures such as distraction osteogenesis, augmentation of the mandibular ridge with autogenous bone, and bone substitutes followed by the placement of implants. The number of patients participating in a study, the follow-up period, the design of the study, the degree of mandibular resorption, and the survival rate of the dental implants all are considered evaluation parameters. Although numerous studies have described the outcome results of dental implants in the edentulous mandible, there have been few prospective studies designed as randomized clinical trials that compare different treatment modalities to restore the severely resorbed mandible. Therefore, it is not yet possible to select an evidence-based treatment modality. Future research has to be focused on long-term, detailed follow-up clinical trials before scientifically based decisions in treating these patients can be made. This will contribute to a higher level of care in this field. PMID- 15284189 TI - Modifications in the biophysical properties of connexin43 channels by a peptide of the cytoplasmic loop region. AB - Connexin43 (Cx43) channels reside in at least 3 states: closed, open, or residual. It is hypothesized that the residual state results from the interaction of an intracellular "gating element" with structures at the vestibule of the pore. Recently, we showed in vitro that there is an intramolecular interaction of the carboxyl-terminal domain (referred to as "CT") with a region in the cytoplasmic loop of Cx43 (amino acids 119 to 144; referred to as "L2"). Here, we assessed whether the L2 region was able to interact with the gating particle in a functional channel. Cx43 channels were recorded in the presence of a peptide corresponding to the L2 region, delivered via the patch pipette. This manipulation did not modify unitary conductance, but decreased the frequency of transitions into the residual state, prolonged open time, and altered the voltage dependence of the channel in a manner analogous to that observed after truncation of the CT domain. The latter correlated with the ability of the peptide to bind to the CT domain, as determined by mirror resonance spectroscopy. Overall, we propose that the L2 acts as a "receptor" that interacts with a flexible intracellular gating element during channel gating. The full text of this article is available online at http://circres.ahajournals.org. PMID- 15284190 TI - Renovascular hypertension in mice with brain-selective overexpression of AT1a receptors is buffered by increased nitric oxide production in the periphery. AB - We recently established a new transgenic mouse model with brain-restricted overexpression of angiotensin II (Ang II) type 1a receptors (NSE-AT(1a)) to unmask the role of the brain renin-angiotensin system in hypertension. To test the hypothesis that these mice would exhibit an early exacerbation of renovascular hypertension, NSE-AT(1a) and nontransgenic (NT) mice underwent 2 kidney-1-clip (2K1C) surgery and blood pressure (BP) and heart rate (HR) were recorded continuously by radiotelemetry for 28 days. Results show that NSE-AT(1a) mice developed hypertension much more rapidly than NT, and this was not attributable to genotype-related differences in plasma or brain Ang II levels. A marked bradycardia accompanied this early increase in BP in NSE-AT(1a) mice, as did a substantial cardiovascular region-specific downregulation of AT(1) receptor binding in brain but not in kidney. As BP reached its plateau in NT ( approximately 1 week after clip), hypertension began to abate and eventually stabilized at significantly lower levels in NSE-AT(1a) mice despite marked elevations in Ang II levels in brain stem and hypothalamus at these later time points. This hypertension reversal and the bradycardia were prevented by chronic infusion of the nitric oxide synthase (NOS) blocker l-NAME. These data, along with evidence showing enhanced NOS expression and NO-mediated compensatory responses in 2K1C NSE-AT(1a) peripheral arteries during this later phase, suggest that activation of endogenous NO systems plays an important role in buffering the maintenance of hypertension caused by overexpression of AT(1a) receptors in the brain. PMID- 15284191 TI - Thrombospondin-2 is essential for myocardial matrix integrity: increased expression identifies failure-prone cardiac hypertrophy. AB - Cardiac hypertrophy can lead to heart failure (HF), but it is unpredictable which hypertrophied myocardium will progress to HF. We surmised that apart from hypertrophy-related genes, failure-related genes are expressed before the onset of failure, permitting molecular prediction of HF. Hearts from hypertensive homozygous renin-overexpressing (Ren-2) rats that had progressed to early HF were compared by microarray analysis to Ren-2 rats that had remained compensated. To identify which HF-related genes preceded failure, cardiac biopsy specimens were taken during compensated hypertrophy and we then monitored whether the rat progressed to HF or remained compensated. Among 48 genes overexpressed in failing hearts, we focused on thrombospondin-2 (TSP2). TSP2 was selectively overexpressed only in biopsy specimens from rats that later progressed to HF. Moreover, expression of TSP2 was increased in human hypertrophied hearts with decreased (0.19+/-0.01) versus normal ejection fraction (0.11+/-0.03 [arbitrary units]; P<0.05). Angiotensin II induced fatal cardiac rupture in 70% of TSP2 knockout mice, with cardiac failure in the surviving mice; this was not seen in wild-type mice. In TSP2 knockout mice, angiotensin II increased matrix metalloproteinase (MMP)-2 and MMP-9 activity by 120% and 390% compared with wild-type mice (P<0.05). In conclusion, we identify TSP2 as a crucial regulator of the integrity of the cardiac matrix that is necessary for the myocardium to cope with increased loading and that may function by its regulation of MMP activity. This suggests that expression of TSP2 marks an early-stage molecular program that is activated uniquely in hypertrophied hearts that are prone to fail. PMID- 15284192 TI - Role for a novel signaling intermediate, phosphatidylinositol 5-phosphate, in insulin-regulated F-actin stress fiber breakdown and GLUT4 translocation. AB - The cellular functions and regulation of phosphatidylinositol (PtdIns) 5 phosphate (5-P), the newest addition to the family of phosphoinositides (PIs), are still elusive. Here we have examined a plausible role of PtdIns 5-P as a signaling intermediate in acute insulin action. A wortmannin-insensitive transient increase of PtdIns 5-P mass levels that peaked at 10 min, and declined 20-30 min after insulin stimulation, was observed in both Chinese hamster ovary (CHO)-T cells stably expressing the insulin receptor and 3T3-L1 adipocytes. Similarly to insulin, found to induce a rapid disassembly of Texas-Red phalloidin labeled actin stress fibers in CHO-T cells, microinjected PtdIns 5-P, but not other PIs, decreased the number and length of F-actin stress fibers in this cell type to a magnitude seen in response to insulin. Likewise, increases of PtdIns 5 P by ectopic expression of the PtdIns 5-P-producing enzyme PIKfyve yielded a similar effect. As with insulin, the PtdIns 5-P-induced loss of actin stress fibers was independent of PI 3-kinase activation. Furthermore, sequestration of functional PtdIns 5-P, either by ectopic expression of 3xPHD domains that bind selectively PtdIns 5-P or by microinjecting the GST-3xPHD fusion peptide, abrogated insulin-induced F-actin stress fiber disassembly in CHO-T cells. In 3T3 L1 adipocytes, microinjected PtdIns 5-P, but not other PIs, partially mimicked insulin's effect of translocating enhanced green fluorescent protein-GLUT4 to the cell surface. Conversely, insulin-induced myc-GLUT4 vesicle dynamics was arrested in the presence of coexpressed enhanced green fluorescent protein-3xPHD. Involvement of PIKfyve membrane recruitment, but not activation, and/or a decrease in PtdIns 4,5-bisphosphate levels are likely to be among the mechanisms underlying the insulin-induced PtdIns 5-P increase. Together, these results identify PtdIns 5-P as a novel key intermediate for insulin signaling in F-actin remodeling and GLUT4 translocation. PMID- 15284193 TI - Cooperative activation of lipolysis by protein kinase A and protein kinase C pathways in 3T3-L1 adipocytes. AB - In the present study, we investigate the coherence of signaling pathways leading to lipolysis in 3T3-L1 adipocytes. We observe two linear signaling pathways: one well known, acting via cAMP and protein kinase A (PKA) activation, and a second one induced by phorbol 12-myristate 13-acetate treatment involving protein kinase C (PKC) and MAPK. We demonstrate that both the PKA regulatory subunits RIalpha and RIIbeta are expressed in 3T3-L1 adipocytes and are responsible for the lipolytic effect mediated via the cAMP/PKA pathway. Inhibition of the PKA pathway by the selective PKA inhibitor Rp-8-CPT-cAMPS does not impair lipolysis induced by PKC activation, and neither PD98059 nor U0126, as known MAPK kinase inhibitors, changes the level of glycerol release caused by PKA activation, indicating no cross-talk between these two pathways when only one is activated. However, when both are activated, they act synergistically on glycerol release. Additional experiments focusing on this synergy show no involvement of MAPK phosphorylation and cAMP formation. Phosphorylation of hormone-sensitive lipase is similar upon stimulation of either pathway, but we demonstrate a difference in the ability of both PKA and the PKC pathway activation to phosphorylate perilipin, which in turn may be an explanation for the different maximal lipolytic effect of both pathways. PMID- 15284194 TI - Leptin enhances oocyte nuclear and cytoplasmic maturation via the mitogen activated protein kinase pathway. AB - Recent studies have suggested that leptin has a central role in female reproduction, including ovarian function. The leptin receptor (Ob-R) has six isoforms and can signal through either the MAPK or the Janus-activated kinase/signal transducer and activator of transcription signal-transduction pathway, depending on the isoform. Expression of Ob-R has been reported in human and mouse oocytes; however, the physiological role of leptin during follicular development and oocyte maturation is largely unknown. In the current study, expression of Ob-R during oocyte growth and maturation was investigated in porcine oocytes from small, medium, and large follicles and in oocytes in the germinal vesicle (GV), GV breakdown, and metaphase II (MII) stages at both the mRNA and protein levels. The proportion of oocytes expressing Ob-R was maximal in oocytes from medium follicles and at the GV breakdown stage (P < 0.05), whereas the proportion of oocytes expressing the long isoform, Ob-Rb, was found to be consistently low throughout growth and maturation. When included in oocyte maturation medium, leptin significantly increased the proportion of oocytes reaching MII (P < 0.01), elevated cyclin B1 protein content in MII-stage oocytes (P < 0.05), and enhanced embryo developmental potential (P < 0.05), suggesting that leptin plays a role in both nuclear and cytoplasmic maturation. During oocyte maturation, leptin increased phosphorylated MAPK content by 2.8-fold (P < 0.05), and leptin-stimulated oocyte maturation was blocked when leptin-induced MAPK phosphorylation was suppressed by a specific MAPK activation inhibitor, U0126 (P < 0.01), demonstrating that leptin enhances nuclear maturation via activation of the MAPK pathway. PMID- 15284195 TI - Prenatal exposure to interleukin-6 results in hypertension and increased hypothalamic-pituitary-adrenal axis activity in adult rats. AB - During pregnancy, systemic inflammatory responses induce cytokines that may stress the fetus and contribute to cardiovascular and neuroendocrine dysfunction in adulthood. We evaluated the effects of early and late prenatal exposure to IL 6 on mean systolic arterial pressure (MSAP) and hypothalamic-pituitary-adrenal (HPA) axis regulation in male and female rats at 5-24 wk of age. MSAP and ACTH and corticosterone levels were measured basally and in response to a novel environment, immobilization stress, and stimulation with corticotropin-releasing factor (CRF) and ACTH. In addition, mRNA expression and protein levels of glucocorticoid receptor, mineralocorticoid receptor, CRF receptor type 1, and CRF were estimated in brain areas thought to mediate central effects of corticosteroids on the HPA axis and on central neuroendocrine regulation of MSAP. Both early and late prenatal IL-6 exposure led to hypertension, which was evident in females at 5 wk of age. In adult rats, basal ACTH and corticosterone levels were elevated, the responses to stress and stimulation tests were of extended duration, and circadian rhythm during the light period was flattened and reversed. Mineralocorticoid receptor and glucocorticoid receptor mRNA expression was reduced in the hippocampus, the CRF level was increased in the hypothalamus, and CRF receptor type 1 mRNA expression was increased in the pituitary. These findings suggest that fetal stress induced by prenatal exposure to IL-6 leads to hypertension and dysregulation of HPA axis activity during adulthood. PMID- 15284196 TI - Influence of diabetes on the exacerbation of an inflammatory response in cardiovascular tissue. AB - Coronary artery disease results from an inflammatory process in blood vessels of afflicted individuals. This process is accelerated with diabetes for reasons that are largely unknown. Recent evidence indicates that infection at sites remote from the heart leads to bacteremia and endotoxemia, thereby stimulating systemic inflammation, which represents an important risk factor for atherosclerosis. We examined the inflammatory response of the heart/aorta of diabetic db/db mice that develop type II diabetes. Subcutaneous inoculation of lipopolysaccharide was used to mimic a local infection. This stimulated an up-regulation of adhesion molecules, cytokines, and chemokines via an endotoxemia that was significantly more rapid and more pronounced in the diabetic compared with normal mice. The 13- to 30-fold induction of key proinflammatory molecules in the heart/aorta of diabetic mice even exceeded that at the site of inoculation. Given that infection, bacteremia, and endotoxemia are relatively frequent events in humans, these results identify a putative mechanism for increased cardiovascular heart disease in diabetes. PMID- 15284198 TI - Molecular and functional characterization of sex hormone binding globulin in zebrafish. AB - SHBG (sex hormone binding globulin) transports androgens and estrogens in the blood of vertebrates including fish. Orthologs of SHBG in fish are poorly defined, and we have now obtained a zebrafish SHBG cDNA and characterized the zebrafish SHBG gene and protein through molecular biological, biochemical, and informatics approaches. Amino-terminal analysis of zebrafish SHBG indicated that its deduced precursor sequence includes a 25-residue secretion polypeptide and exhibits 22-27% homology with mammalian SHBG sequences and 41% with a deduced fugufish SHBG sequence. The 356-residue mature zebrafish SHBG (39,243 Da) sequence comprises a tandem repeat of laminin G-like domains typical of SHBG sequences; contains three N-glycosylation sites; and exists as a 105,000 +/- 8700 Da homodimer. Zebrafish SHBG exhibits a high affinity and specificity for sex steroids. An RT-PCR indicated that SHBG mRNA first appears in zebrafish larva, and SHBG mRNA was localized within the liver and gut at this stage of development by whole-mount in situ hybridization. In adult fish, SHBG mRNA was found in liver, testis, and gut. In the liver, immunoreactive SHBG was present in hepatocytes and concentrated in intrahepatic bile duct cells, whereas in the testis it was confined to cells surrounding the seminiferous tubule cysts. In the intestine, immunoreactive SHBG was present in the stroma and epithelial cells of the villous projections and the surrounding muscle. The production and presence of SHBG in the gut of developing and adult zebrafish suggests a novel role for this protein in regulating sex steroid action at this site. PMID- 15284197 TI - Susceptibility rather than resistance to hyperthyroidism is dominant in a thyrotropin receptor adenovirus-induced animal model of Graves' disease as revealed by BALB/c-C57BL/6 hybrid mice. AB - We investigated why TSH receptor (TSHR) adenovirus immunization induces hyperthyroidism more commonly in BALB/c than in C57BL/6 mice. Recent modifications of the adenovirus model suggested that using adenovirus expressing the TSHR A subunit (A-subunit-Ad), rather than the full-length TSHR, and injecting fewer viral particles would increase the frequency of hyperthyroidism in C57BL/6 mice. This hypothesis was not fulfilled; 65% of BALB/c but only 5% of C57BL/6 mice developed hyperthyroidism. TSH binding inhibitory antibody titers were similar in each strain. Functional TSHR antibody measurements provided a better indication for this strain difference. Whereas thyroid-stimulating antibody activity was higher in C57BL/6 than BALB/c mice, TSH blocking antibody activity was more potent in hyperthyroid-resistant C57BL/6 mice. F(1) hybrids (BALB/c x C57BL/6) responded to A-subunit-Ad immunization with hyperthyroidism and TSHR antibody profiles similar to those of the hyperthyroid-susceptible parental BALB/c strain. In contrast, ELISA of TSHR antibodies revealed that the IgG subclass distribution in the F(1) mice resembled the disease-resistant C57BL/6 parental strain. Because the IgG subclass distribution is dependent on the T helper 1/T helper 2 cytokine balance, this paradigm can likely be excluded as an explanation for susceptibility to hyperthyroidism. In summary, our data for BALB/c, C57BL/6, and F(1) strains suggest that BALB/c mice carry a dominant gene(s) for susceptibility to induction of a thyroid-stimulating antibody/TSH blocking antibody balance that results in hyperthyroidism. Study of this genetic influence will provide useful information on potential candidate genes in human Graves' disease. PMID- 15284199 TI - Novel expression of gonadotropin subunit genes in oocytes of the gilthead seabream (Sparus aurata). AB - It is widely believed that FSH and LH, which are known to play key roles in controlling the production of functional oocytes in vertebrates, are synthesized and secreted exclusively by the anterior pituitary. Here we present evidence for the novel expression of FSHbeta, LHbeta, and the common glycoprotein-alpha (Cgalpha) in the gilthead seabream ovary. Using in situ hybridization and immunocytochemistry, FSHbeta was detected in primary-growth and secondary-growth I oocytes, LHbeta was found in secondary-growth oocytes, and Cgalpha was observed in both primary and secondary-growth oocytes. Northern blot analyses demonstrated that Fshbeta transcript is 0.6 kb in both pituitary and ovary, whereas the ovarian Lhbeta transcript (1.1 kb), unexpectedly, is longer than the known pituitary Lhbeta transcript (0.6 kb). Sequence analyses revealed that ovarian Lhbeta is driven by a different promoter than pituitary Lhbeta, which generates an additional 459 bases at the distal portion of the 5'-untranslated region of the ovarian Lhbeta. Furthermore, using in vitro ovarian fragment incubation, we demonstrated that mammalian GnRH analog agonist enhanced the expression of ovarian Fshbeta (up to 2.7-fold), Lhbeta (up to 1.4-fold), Cgalpha (up to 1.8 fold), and the secretion of ovarian LH (up to 2.2-fold). In contrast, GnRH antagonist, analog E, suppressed the secretion of ovarian LH. Our findings suggest that a GnRH-gonadotropin axis is present in the gilthead seabream ovary and that FSH and LH, the well-characterized pituitary hormones, may have prominent novel roles in teleost intraovarian communication between oocytes and ovarian follicle cells. PMID- 15284200 TI - Chromosomes 6 and 13 harbor genes that regulate pubertal timing in mouse chromosome substitution strains. AB - Variation in the onset of puberty among inbred strains of mice suggests that quantitative trait loci (QTLs) affect neurological and hormonal aspects of sexual maturation. Taking a novel approach toward identifying factors that regulate the hypothalamic-pituitary-gonadal (HPG) axis, we evaluated pubertal timing [as assessed by vaginal opening (VO)] in two inbred strains of mice, A/J and C57BL/6J (B6), and in a panel of chromosome substitution strains (CSSs) generated from A/J and B6 mice. In each CSS, a single chromosome from A/J has been substituted in a homozygous fashion for the corresponding chromosome in B6, partitioning the A/J genome into 22 strains with a common host (B6) background. VO occurred significantly earlier in A/J compared with B6 mice. Although the majority of the CSSs assessed had a timing of VO that was similar to the progenitor B6 strain, CSSs for chromosomes 6 and 13 each displayed significantly earlier time of VO than B6 mice. F1 (B6 x CSS) mice for chromosomes 6 and 13 displayed phenotypes that were intermediate between the CSS and B6 strains, suggesting that the trait was inherited in a codominant manner. These findings demonstrate that chromosomes 6 and 13 harbor QTLs that control the timing of VO. Identification of the responsible genes may reveal factors that regulate the maturation of the HPG axis and determine the timing of puberty. PMID- 15284201 TI - Cytotrophoblasts up-regulate soluble fms-like tyrosine kinase-1 expression under reduced oxygen: an implication for the placental vascular development and the pathophysiology of preeclampsia. AB - Sufficient cytotrophoblast (CT) invasion into the uterine wall and subsequent remodeling of maternal uterine vasculature is critical to establish uteroplacental circulation. The production of vascular endothelial growth factor (VEGF) family molecules is confirmed in placental cells including CTs, but it is not elucidated how the VEGF system in CTs is controlled by oxygen tension and how it is involved in the development of placental circulation. To address this, we explored the effect of oxygen tension on the expression of VEGF, placenta growth factor (PlGF), and their antagonist, soluble fms-like tyrosine kinase-1 (sFlt-1) using ELISA and real-time PCR in a primary CT cell culture. For comparison, the same was conducted in parallel using other cells comprising placenta, such as human umbilical vein endothelial cells (HUVECs) and villous fibroblasts (VFs). Reduced oxygen resulted in a pronounced increase in sFlt-1 mRNA amount and sFlt-1 release into the culture media in CTs, whereas this was not the case with HUVECs and VFs. Free (not bound to sFlt-1) VEGF was not detected in CT culture media regardless of oxygen concentration, even though VEGF expression was stimulated by reduced oxygen in CTs, which was similar to the stimulation in HUVECs and VFs. Free PlGF was also diminished in CT culture media by reduced oxygen. These results implicate that CTs possess a unique property to enhance sFlt-1 production under reduced oxygen, which could consequently antagonize angiogenic activity of VEGF and PlGF. The presented findings might provide a framework with which to understand the mechanism of uterine vascular remodeling and its perturbations as exemplified in preeclampsia. PMID- 15284202 TI - Effects of peptide YY[3-36] on short-term food intake in mice are not affected by prevailing plasma ghrelin levels. AB - The gut-derived hormones peptide YY[3-36] (PYY[3-36]) and ghrelin are believed to influence similar hypothalamic circuits, albeit with opposing actions on energy balance. Thus, we carried out a series of studies to evaluate the interaction of these hormones on short-term food intake responses in mice. Intraperitoneal PYY[3 36] injection reduced short-term food intake by up to 50% in overnight-fasted mice and in postabsorptive animals during the early and late light cycle. This effect was not sensitive to the prevailing endogenous plasma acyl-ghrelin concentrations, which ranged from high physiological (overnight-fasted, 1252 +/- 108 pg/ml) to low levels (late light cycle, 402 +/- 33 pg/ml). PYY[3-36] administration did not reduce plasma total or acyl-ghrelin concentration in conjunction with its anorexigenic actions. Ghrelin increased short-term food intake by up to 1.8-fold in mice treated ip in the early light cycle, but was ineffective in animals treated after an overnight fast or during the late light cycle. Ghrelin did not increase food intake or GH secretion unless plasma levels were increased above high physiological fasting values. The anorexigenic effect of PYY[3-36] over a range of doses was not compromised by coinjection of ghrelin, and PYY[3-36] reduced food intake in agouti mice, which lack fully functional melanocortin signaling. These results in mice support a model in which 1) PYY[3 36] diminishes short-term food intake at least in part through mechanisms distinct from the neuropeptide Y/proopiomelanocortin neural circuits engaged by ghrelin; and 2) a reduction in circulating ghrelin is not requisite for the anorexigenic effects of PYY[3-36]. PMID- 15284203 TI - The inhibition of gastric ghrelin production by food intake in rats is dependent on the type of macronutrient. AB - Ghrelin is a peptide mainly produced by the stomach that increases food intake and body weight. Ghrelin expression increases with fasting and is diminished by re-feeding, but although the expression of this hormone is regulated by the feeding state, the relation with diet composition is not yet well established. We have studied the inhibitory effect of the intake of two different macronutrients (fat and carbohydrates) on ghrelin production by the stomach in fasted rats, as well as the relation with another important signal in the regulation of energy balance, leptin. Ghrelin mRNA expression in the gastric mucosa was determined by Northern blotting, and leptin mRNA expression was determined by Northern blotting in the adipose tissue and by RT-PCR in the stomach; circulating and gastric concentrations of ghrelin and leptin were measured by enzyme immunosorbent assay and ELISA, respectively. Our results showed an increase in ghrelin mRNA levels in response to 14 h of fasting. Food intake for 20 min after the fast produced a decrease in ghrelin mRNA expression that was recovered in 45 min in rats that ate the fat diet, whereas levels remained low when rats ate the carbohydrate diet. Serum ghrelin followed a similar tendency. The decrease in ghrelin expression by feeding was associated with an increased expression of gastric leptin only when animals ate carbohydrates. We conclude that the inhibition of ghrelin production by the stomach after re-feeding of fasted rats is dependent on diet composition and can be related to the different satiating capacity of the ingested macronutrients, which is higher for carbohydrates than fat. PMID- 15284204 TI - Regulation of insulin-stimulated muscle glucose uptake in the conscious mouse: role of glucose transport is dependent on glucose phosphorylation capacity. AB - Previous work suggests that normal GLUT4 content is sufficient for increases in muscle glucose uptake (MGU) during hyperinsulinemia, because glucose phosphorylation is the more formidable barrier to insulin-stimulated MGU. It was hypothesized that a partial ablation of GLUT4 would not impair insulin-stimulated MGU when glucose phosphorylation capacity is normal but would do so when glucose phosphorylation capacity is increased. Thus, chow-fed C57BL/6J mice with a GLUT4 partial knockout (GLUT4(+/-)), hexokinase II overexpression (HK(Tg)), or both (HK(Tg) + GLUT4(+/-)) and wild-type littermates were studied. Carotid artery and jugular vein catheters were implanted for sampling and infusions at 4 months of age. After a 5-d recovery, 5-h fasted mice (n = 8-11/group) underwent a 120-min saline infusion or insulin clamp (4 mU/kg.min insulin with glucose maintained at 165 mg/dl) and received a 2-deoxy[(3)H]glucose bolus to provide an index of MGU (R(g)) for the soleus, gastrocnemius, and superficial vastus lateralis. Basal R(g) from all muscles studied from saline-infused mice were not changed by any of the genetic modifications. HK(Tg) mice had augmented insulin-stimulated R(g) in all muscles studied compared with remaining genotypes. Insulin-stimulated R(g) was not impaired in any of the muscles studied from GLUT4(+/-) mice. However, the enhanced insulin-stimulated R(g) created by HK overexpression was ablated in HK(Tg) + GLUT4(+/-) mice. Thus, a 50% reduction of normal GLUT4 content in the presence of normal HK activity does not impair insulin-stimulated MGU. However, when the glucose phosphorylation barrier is lowered by HK overexpression, GLUT4 availability becomes a limitation to insulin-stimulated MGU. PMID- 15284205 TI - Expression and the biological activities of insulin-like growth factor-binding protein related protein 1 in rat uterus during the periimplantation period. AB - IGF binding protein-related protein 1 (IGFBP-rP1) is highly expressed in the rat uterus around the time of implantation. In the present study, we determined the periimplantation localization of IGFBP-rP1 mRNA and assessed the effects of recombinant IGFBP-rP1 on the proliferative and prostacyclin (PGI(2))-producing abilities of cultured endometrial cells early in pregnancy. IGFBP-rP1 mRNA was detected at high levels in endometrial stromal cells close to the smooth muscle of interimplantation sites around the time of implantation but absent from decidual zones surrounding the embryo. Differential uterine IGFBP-rP1 expression was also recognized in the delayed implanting pregnant model, but the level of mRNA decreased as decidual tissues formed in the decidualization model. Recombinant IGFBP-rP1 inhibited the proliferation of endometrial stromal cells in vitro and arrested them in the G(1) phase of the cell cycle. Furthermore, IGFBP rP1 significantly stimulated PGI(2) synthesis and cyclooxygenase II mRNA expression in myometrial cells, both of which are essential molecules for successful implantation. These data suggest that IGFBP-rP1 is an implantation associated protein and that it modulates the proliferation of rat uterine cells and their production of PGI(2) during the periimplantation period. PMID- 15284206 TI - Role of the insulin-like growth factor I decline in the induction of atrogin 1/MAFbx during fasting and diabetes. AB - In catabolic conditions, atrogin-1/MAFbx, a muscle-specific ubiquitin-ligase required for muscle atrophy, is increased, and concentrations of IGF-I, a growth factor known to have antiproteolytic action, are reduced. To define the relationship between the decline in IGF-I and the induction of atrogin-1/MAFbx, we studied the effect of IGF-I replacement on atrogin-1/MAFbx mRNA in rats fasted for 51 h and in rats made diabetic with streptozotocin (STZ). Fasting produced a 5.8-fold increase in atrogin-1/MAFbx (P < 0.001). This was attenuated to a 2.5 fold increase by injections of IGF-I (P < 0.05 vs. fasting). Animals with STZ induced diabetes experienced a 15.1-fold increase in atrogin-1/MAFbx (P < 0.001). Normalization of their circulating IGF-I concentrations by IGF-I infusion blunted the induction of atrogin-1/MAFbx to 6.3-fold (P < 0.05 vs. STZ diabetes without IGF-I). To further delineate the regulation of atrogin-1/MAFbx by IGF-I, we studied a model of cultured muscle cells. We observed that IGF-I produced a time- and dose-dependent reduction of atrogin-1/MAFbx mRNA, with a 50% effective dose of 5 nm IGF-I, a physiological concentration. The degradation rate of atrogin 1/MAFbx mRNA was not affected by IGF-I, suggesting that the reduction of atrogin 1/MAFbx mRNA by IGF-I is a transcriptional effect. Exposure of muscle cells in culture to dexamethasone increased atrogin-1/MAFbx mRNA with a 50% effective dose of 10 nm, a pharmacological concentration. In the presence of dexamethasone, IGF I at physiological concentrations retained its full inhibitory effect on atrogin 1/MAFbx mRNA. We conclude that IGF-I inhibits atrogin-1/MAFbx expression and speculate that this effect might contribute to the antiproteolytic action of IGF I in muscle. PMID- 15284207 TI - Transgenic mice overexpressing human fibroblast growth factor 23 (R176Q) delineate a putative role for parathyroid hormone in renal phosphate wasting disorders. AB - Fibroblast growth factor 23 (FGF23) is a recently characterized protein likely involved in the regulation of serum phosphate homeostasis. Increased circulating levels of FGF23 have been reported in patients with renal phosphate-wasting disorders, but it is unclear whether FGF23 is the direct mediator responsible for the decreased phosphate transport at the proximal renal tubules and the altered vitamin D metabolism associated with these states. To examine this question, we generated transgenic mice expressing and secreting from the liver human FGF23 (R176Q), a mutant form that fails to be degraded by furin proteases. At 1 and 2 months of age, mice carrying the transgene recapitulated the biochemical (decreased urinary phosphate reabsorption, hypophosphatemia, low serum 1,25 dihydroxyvitamin D(3)) and skeletal (rickets and osteomalacia) alterations associated with these disorders. Unexpectantly, marked changes in parameters of calcium homeostasis were also observed, consistent with secondary hyperparathyroidism. Moreover, in the kidney the anticipated alterations in the expression of hydroxylases associated with vitamin D metabolism were not observed despite the profound hypophosphatemia and increased circulating levels of PTH, both major physiological stimuli for 1,25-dihydroxyvitamin D(3) production. Our findings strongly support the novel concept that high circulating levels of FGF23 are associated with profound disturbances in the regulation of phosphate and vitamin D metabolism as well as calcium homeostasis and that elevated PTH levels likely also contribute to the renal phosphate wasting associated with these disorders. PMID- 15284208 TI - The epidermal growth factor-like growth factor amphiregulin is strongly induced by the adenosine 3',5'-monophosphate pathway in various cell types. AB - We examined the cAMP-mediated regulation of the epidermal growth factor-like growth factor amphiregulin (AR) in T cells and observed a strong cAMP-induced up regulation of AR mRNA in a time- and concentration-dependent manner independent of T cell activation. This regulation may be mediated in part through activation of a cAMP-responsive element in the AR promoter, because the cAMP-responsive element conferred cAMP responsiveness to a luciferase reporter in Jurkat TAg cells. Similar effects of AR mRNA induction were seen in T cells treated with cAMP-elevating agents such as prostaglandin E(2) and forskolin as well as with the phosphodiesterase inhibitors rolipram and isobutylmethylxanthine. Furthermore, the induction of AR mRNA by cAMP was strongly suppressed by a protein kinase A type I-selective inhibitor, whereas treatment with an exchange protein directly activated by cAMP-specific agonist did not increase AR levels. In addition, an increase in AR gene transcripts by cAMP was seen in MCF-7 mammary carcinoma cells and H295R adrenal cells. Moreover, the potent cAMP-mediated induction of AR mRNA resulted in increased secretion (5-fold) of AR from T cells. Furthermore, supernatants from cAMP-stimulated T cells containing secreted AR induced phosphorylated MAPK in OVCAR-3 carcinoma cells. In conclusion, our data suggest that AR is under strong regulation by the cAMP pathway in various cell types, and that prostaglandin E(2)- and cAMP-induced AR secretion from T cells may be highly relevant in a microenvironment consisting of tumor cells and infiltrated immune cells, because AR by activating the MAPK pathway through a paracrine route may contribute to proliferation of tumor cells and thus add to neoplastic processes. PMID- 15284209 TI - Transcription suppression of peroxisome proliferator-activated receptor gamma2 gene expression by tumor necrosis factor alpha via an inhibition of CCAAT/ enhancer-binding protein delta during the early stage of adipocyte differentiation. AB - TNFalpha is known to inhibit adipocyte differentiation and induce insulin resistance. Moreover, TNFalpha is known to down-regulate peroxisome proliferator activated receptor (PPAR)gamma2, an adipocyte-specific nuclear receptor of insulin-sensitizer thiazolidinediones. To clarify molecular mechanisms of TNFalpha- mediated PPARgamma2 down-regulation, we here examined the effect of TNFalpha on transcription regulation of PPARgamma2 gene expression during the early stage of adipocyte differentiation. 3T3-L1 preadipocytes (2 d after 100% confluent) were incubated in a differentiation mixture (dexamethasone, insulin, 3 isobutyl-1-methlxanthine), with or without 50 ng/ml TNFalpha, for 24 h. TNFalpha significantly decreased PPARgamma2 expression both at mRNA and protein levels (to approximately 40%), as well as aP2 mRNA expression. The mouse PPARgamma2 gene promoter region (2.2-kb) was isolated and was used for luciferase reporter assays by transient transfection. TNFalpha significantly suppressed PPARgamma2 gene transcription (to approximately 50%), and deletion analyses demonstrated that the suppression was mediated via CCAAT/enhancer-binding protein (C/EBP) binding elements at the -320/-340 region of the promoter. Moreover, TNFalpha significantly decreased expression of C/EBPdelta mRNA and protein levels (to approximately 40%). EMSA, using 3T3-L1 cells nuclear extracts with the -320/-340 region as a probe, demonstrated the binding of C/EBPdelta to the element, which was significantly decreased by TNFalpha treatment. Overexpression of CEBP/delta prevented the TNFalpha-mediated suppression of PPARgamma2 transactivation. Taken together, TNFalpha suppresses PPARgamma2 gene transcription by the inhibition of C/EBPdelta expression and its DNA binding during the early stage of adipocyte differentiation, which may contribute to the inhibition of adipocyte differentiation, as well as the induction of insulin resistance. PMID- 15284210 TI - Ghrelin inhibits the proliferative activity of immature Leydig cells in vivo and regulates stem cell factor messenger ribonucleic acid expression in rat testis. AB - Ghrelin has emerged as putative regulator of an array of endocrine and nonendocrine functions, including cell proliferation. Recently, we provided evidence for the expression of ghrelin in mature, but not in undifferentiated, Leydig cells of rat and human testis. Yet testicular actions of ghrelin, other than modulation of testosterone secretion, remain unexplored. In the present study we evaluated the effects of ghrelin on proliferation of Leydig cell precursors during puberty and after selective elimination of mature Leydig cells by treatment with ethylene dimethane sulfonate. In these settings, intratesticular injection of ghrelin significantly decreased the proliferative activity of differentiating immature Leydig cells, estimated by 5 bromodeoxyuridine labeling. This response was selective and associated, in ethylene dimethane sulfonate-treated animals, with a decrease in the mRNA levels of stem cell factor (SCF), i.e. a key signal in spermatogenesis and a putative regulator of Leydig cell development. Thus, the effects of ghrelin on SCF gene expression were evaluated. In adult rats, ghrelin induced a significant decrease in SCF mRNA levels in vivo. Such an inhibitory action was also detected in vitro using cultures of staged seminiferous tubules. The inhibitory effect of ghrelin in vivo was dependent on proper FSH input, because it was detected in hypophysectomized rats only after FSH replacement. Overall, it is proposed that acquisition of ghrelin expression by Leydig cell precursors during differentiation may operate as a self-regulatory signal for the inhibition of the proliferative activity of this cell type through direct or indirect (i.e. SCF mediated) mechanisms. In addition, we present novel evidence for the ability of ghrelin to modulate the expression of the SCF gene, which may have implications for the mode of action of this molecule in the testis as well as in other physiological systems. PMID- 15284211 TI - Exposure to CB-153 and p,p'-DDE and male reproductive function. AB - BACKGROUND: During the last decades, there has been concern that exposure to endocrine disruptors, such as persistent organochlorine pollutants (POPs), may contribute to an impairment of male reproductive function. To investigate whether exposure to 2,2'4,4'5,5'-hexachlorobiphenyl (CB-153) and 1,1-dichloro-2,2-bis (p chlorophenyl)-ethylene (p,p'-DDE) affects semen quantity and quality and reproductive hormones, 195 Swedish fishermen, aged 24-65 years, were investigated. METHODS: The men provided semen samples which were analysed in a mobile laboratory unit. Blood samples and information relating to lifestyle, medical and reproductive history were obtained. RESULTS: The subjects had a median CB-153 serum level of 193 ng/g lipid (range 39-1460) and a median p,p'-DDE serum level of 240 ng/g lipid (range 334-2251). When CB-153 was categorized into quintiles, the subjects in the quintile with the highest concentration (> 328 ng/g lipid), tended to have decreased sperm motility compared with the subjects in the lowest quintile (< 113 ng/g lipid). The age-adjusted mean difference was 9.9% (95% confidence interval -1.0 to 21% P = 0.08). We found no significant associations between p,p'-DDE and semen characteristics or reproductive hormones. CONCLUSION: The association between CB-153 and sperm motility, although not formally significant, is of interest considering the possible endocrine disrupting effects of polychlorinated biphenyls (PCBs). PMID- 15284213 TI - CTG amplification in the DM1PK gene is not associated with idiopathic male subfertility. AB - BACKGROUND: Polymorphism in the CTG triplet number in the myotonic dystrophy type 1 (DM1PK) gene has been proposed as being associated with idiopathic azoospermia. The aim of this study was to investigate whether the CTG trinucleotide amplification in the DM1PK gene is associated with male subfertility. METHODS: We evaluated 107 subfertile patients, male partners of infertile couples, affected by non-obstructive azoospermia (n = 38) and oligoasthenoteratozoospermia (OAT) (n = 69), and 102 men with proven fertility. Main outcome measures were CTG repeat size in the DM1PK gene, testicular volume, sperm concentration, rapid progressive motility, normal morphology, serum FSH levels, testicular histology and Johnsen score. RESULTS: In subfertile males, no minimal mutation or mutation carriers were found. The difference in the number of CTG repeat lengths between the groups was not statistically significant (P = 0.825). There was no correlation between the number of CTG repeats and the clinical parameters of subfertile patients: testicular volume, sperm concentration, rapid progressive motility, normal morphology, FSH level, testicular histology and Johnsen score. CONCLUSIONS: The number of CTG repeats in the normal or mutational range of DM1PK gene is associated with neither idiopathic male subfertility nor with clinical characteristics of male subfertility. PMID- 15284212 TI - Ovarian stimulation by concomitant administration of cetrorelix acetate and HMG following Diane-35 pre-treatment for patients with polycystic ovary syndrome: a prospective randomized study. AB - BACKGROUND: Patients with polycystic ovary syndrome (PCOS) may need a longer period of pituitary downregulation to suppress the elevated serum LH and androgen levels effectively during IVF treatment using the GnRH agonist long protocol. We proposed a stimulation protocol incorporating Diane-35 and GnRH antagonist (Diane/cetrorelix protocol) and compared it with the GnRH agonist long protocol for PCOS patients. METHODS: Part I of the study was an observational pilot study to evaluate the hormonal change as a result of the Diane/cetrorelix protocol (n = 26). Part II of the study was a prospective randomized study comparing the Diane/cetrorelix protocol (n = 25) and the GnRH agonist long protocol (n = 24). In the Diane/cetrorelix protocol, patients were pre-treated with three cycles of Diane-35, followed by 0.25 mg of cetrorelix on cycle day 3. From day 4, cetrorelix and gonadotrophin were administered concomitantly until the day of HCG injection. RESULTS: Serum LH, estradiol and testosterone levels were suppressed comparably in both protocols at the start of gonadotrophin administration. Serum LH was suppressed at constant levels without a premature LH surge in the Diane/cetrorelix protocol. The clinical results for both protocols were comparable, with significantly fewer days of injection, lower amounts of gonadotrophin used and lower estradiol levels on the day of HCG injection following the Diane/cetrorelix protocol. Furthermore, there was no significant difference in clinical pregnancy outcome between the two stimulation protocols. CONCLUSIONS: The Diane/cetrorelix protocol has a similar pregnancy outcome to the GnRH agonist long protocol for women with PCOS undergoing IVF treatment. PMID- 15284214 TI - The effect of HRT on cerebral haemodynamics and cerebral vasomotor reactivity in post-menopausal women. AB - BACKGROUND: Cerebral vasomotor reactivity (CVR) is an index of cerebrovascular dilatory capacity which can readily be assessed using trans-cranial Doppler ultrasound. Impaired CVR is associated with elevated risk of stroke. We performed a randomized, double-blind placebo-controlled trial to investigate the effect of two HRT preparations upon CVR. METHODS: We examined middle cerebral artery mean flow velocity (MFV), internal carotid artery pulsatility index (PI) and CVR to an i.v. acetazolamide bolus using ultrasound in three groups of post-menopausal women randomized to oral estradiol 1 mg+norethisterone 0.5 mg (group N), estradiol 1 mg+dydrogesterone 5 mg (group D) or placebo (group P). The MFV, PI and CVR were measured before and after 3 months treatment. RESULTS: Thirty-eight post-menopausal women were recruited (N=12, D=14, P=12); mean (SE) age was 56.7 (4) years. Neither HRT preparation affected CVR [% (SE) change from baseline N +4.2 (11); D +3.8 (5.5); P +4.0 (3.8); all comparisons P = NS]. PI was significantly reduced in recipients of dydrogesterone [% (SE) change from baseline D -5.4% (4.6); N +12.3 (6.9); P +11.6 (6.9). P=0.025]. Middle cerebral artery velocity was significantly increased following dydrogesterone treatment compared with placebo [% (SE) change from baseline D +6.8 (3.4) N +3.9 (4.2) P 4.6% (3.4) P=0.03 for D versus P]. CONCLUSION: HRT did not alter CVR. The reduced PI and increased MFV suggest HRT-induced intracranial vasodilatation, which is more apparent in dydrogesterone recipients. Differences may exist between progestogens with regard to changes in intracranial haemodynamics. PMID- 15284215 TI - A European multicentre prospective randomized study to assess the use of assisted hatching with a diode laser and the benefit of an immunosuppressive/antibiotic treatment in different patient populations. AB - BACKGROUND: Assisted hatching (AH) techniques, designed for facilitating the embryo escape out of the zona pellucida (ZP) have been used in IVF centres since 1992. The initial indications for AH were patient's age, ZP thickness, high basal FSH and repeated IVF failures. Several retrospective and prospective studies assessing AH in these indications have given disparate results. Our aims were to evaluate the benefits of AH and immunosuppressive/antibiotic treatment (IA) in patients with either a poor prognosis of success, previous implantation failures or transfers of cryopreserved embryos. METHODS: Four IVF centres allocated 426 patients, randomized for AH and IA, into four groups of AH indications between 1997 and 1999. AH was performed with a diode laser. ZP thickness, opening size and embryo score were recorded. Outcome measures were implantation and delivery rates. RESULTS: Patients coming for a first or third transfer of cryopreserved embryos and poor prognosis patients admitted for a first trial did not benefit from AH. Even patients with repeated implantation failures of fresh embryos did not gain significantly from AH. CONCLUSIONS: Among AH indications, absence of implantation after several transfers of good quality embryos remains the strongest patient selection criterion. Prescription of an immunosuppressive/antibiotic treatment is essential. PMID- 15284216 TI - The role of structural disorder in the function of RNA and protein chaperones. AB - Chaperones are highly sophisticated protein machines that assist the folding of RNA molecules or other proteins. Their function is generally thought to require a fine-tuned and highly conserved structure: despite the recent recognition of the widespread occurrence of structural disorder in the proteome, this structural trait has never been generally considered in molecular chaperones. In this review we give evidence for the prevalence of functional regions without a well-defined 3-D structure in RNA and protein chaperones. By considering a variety of individual examples, we suggest that the structurally disordered chaperone regions either function as molecular recognition elements that act as solubilizers or locally loosen the structure of the kinetically trapped folding intermediate via transient binding to facilitate its conformational search. The importance of structural disorder is underlined by a predictor of natural disordered regions, which shows an extremely high proportion of such regions, unparalleled in any other protein class, within RNA chaperones: 54.2% of their residues fall into disordered regions and 40% fall within disordered regions longer than 30 consecutive residues. Structural disorder also prevails in protein chaperones, for which frequency values are 36.7% and 15%, respectively. In keeping with these and other details, a novel "entropy transfer" model is presented to account for the mechanistic role of structural disorder in chaperone function. PMID- 15284217 TI - Loss of metal transcription factor-1 suppresses tumor growth through enhanced matrix deposition. AB - Metal transcription factor-1 (MTF-1) is a ubiquitous transcriptional regulator and chromatin insulator with roles in cellular stress responses and embryonic development. The studies described herein establish for the first time the involvement of MTF-1 in tumor development. Genetically manipulated ras transformed mouse embryonic fibroblasts (MEFs), wild-type (MTF-1+/+), or nullizygous for MTF-1 (MTF-1-/-) were used to develop fibrosarcoma tumors. Loss of MTF-1 resulted in delayed tumor growth associated with increased matrix collagen deposition and reductions in vasculature density. Molecular consequences of MTF-1 loss include increased expression and activation of the transforming growth factor-beta1 (TGF-beta1) and tissue transglutaminase (tTG), two proteins with documented roles in the production and stabilization of extracellular matrix (ECM). Our findings support the hypothesis that MTF-1 enhances the ability of the developing tumor mass to evade fibrosis and scarring of the tumor, a critical step in tumor cell proliferation. PMID- 15284218 TI - Improved survival in experimental sepsis with an orally administered inhibitor of apoptosis. AB - The pathophysiology of sepsis involves excessive lymphocyte apoptosis, which correlates with adverse outcomes, and disordered cytokine production, which may promote host injury. As the protease inhibitor (PI) class of antiretroviral agents is known to prevent apoptosis in vitro, we evaluated their effect on survival, lymphocyte apoptosis, and consequent cytokine production in mice with sepsis induced by cecal ligation and perforation. Mice pretreated with PIs have improved survival (67%; P<0.0005) compared with controls (17%) and a significant (P<0.05) reduction in lymphocyte apoptosis. Even mice receiving therapy beginning 4 h after perforation demonstrated improved survival (50%; P<0.05) compared with controls. PI therapy is also associated with an increase in the Th1 cytokine TNF alpha (P<0.05) early in sepsis and a reduction in the Th2 cytokines IL-6 and IL 10 (P<0.05) late in sepsis; despite no intrinsic antibacterial effects, PI also reduced quantitative bacterial blood cultures. The beneficial effects of PI appear to be specific to lymphocyte apoptosis, as lymphocyte-deficient Rag1-/- mice did not experience benefit from treatment with PI. Thus, inhibition of lymphocyte apoptosis by PI is a candidate approach for the treatment of sepsis. PMID- 15284219 TI - Central role for aldose reductase pathway in myocardial ischemic injury. AB - Aldose reductase (AR), a member of the aldo-keto reductase family, has been implicated in the development of vascular and neurological complications of diabetes. Recently, we demonstrated that aldose reductase is a component of myocardial ischemic injury and that inhibitors of this enzyme protect rat hearts from ischemia-reperfusion injury. To rigorously test the effect of aldose reductase on myocardial ischemia-reperfusion injury, we used transgenic mice broadly overexpressing human aldose reductase (ARTg) driven by the major histocompatibility complex I promoter. Hearts from these ARTg or littermate mice (WT) (n=6 in each group) were isolated, perfused under normoxic conditions, then subjected to 50 min of severe low flow ischemia followed by 60 min of reperfusion. Creatine kinase (CK) release (a marker of ischemic injury) was measured during reperfusion; left ventricular developed pressure (LVDP), end diastolic pressure (EDP), and ATP were measured throughout the protocol. CK release was significantly greater in ARTg mice compared with the WT mice. LVDP recovery was significantly reduced in ARTg mice compared with the WT mice. Furthermore, ATP content was higher in WT mice compared with ARTg mice during ischemia and reperfusion. Infarct size measured by staining techniques and myocardial damage evaluated histologically were also significantly worse in ARTg mice hearts than in controls. Pharmacological inhibition of aldose reductase significantly reduced ischemic injury and improved functional recovery in ARTg mice. These data strongly support key roles for AR in ischemic injury and impairment of functional and metabolic recovery after ischemia. We propose that interventions targeting AR may provide a novel adjunctive approach to protect ischemic myocardium. PMID- 15284220 TI - Biological characterization of angiopoietin-3 and angiopoietin-4. AB - The angiopoietin (Ang) family of growth factors includes Ang1, Ang2, Ang3, and Ang4, all of which bind to the endothelial receptor tyrosine kinase Tie2. Ang3 (mouse) and Ang4 (human) are interspecies orthologs. In experiments with human endothelial cell lines, Ang3 was identified as an antagonist of Tie2 and Ang4 was identified as an agonist of Tie2. However, the biological roles of Ang3 and Ang4 are unknown. We examined the biological effect of recombinant Ang3 and Ang4 proteins in primary cultured endothelial cells and in vivo in mice. Recombinant Ang3 and Ang4 formed disulfide-linked dimers. Ang4 (400 ng/mL) markedly increased Tie2 and Akt phosphorylation in primary cultured HUVECs whereas Ang3 (400 ng/mL) did not produce significant changes. Accordingly, Ang4, but not Ang3, induced survival and migration in primary cultured HUVECs. Unexpectedly, intravenously administered Ang3 (30 microg) was more potent than Ang4 (30 microg) in phosphorylating the Tie2 receptor in lung tissue from mice in vivo. Accordingly, Ang3 was more potent than Ang4 in phosphorylating Akt in primary cultured mouse lung microvascular endothelial cells. Ang3 and Ang4 both produced potent corneal angiogenesis extending from the limbus across the mouse cornea in vivo. Thus, Ang3 and Ang4 are agonists of Tie2, but mouse Ang3 has strong activity only on endothelial cells of its own species. PMID- 15284221 TI - Inhibition of aldose reductase attenuates TNF-alpha-induced expression of adhesion molecules in endothelial cells. AB - Increased expression of adhesion molecules by the activated endothelium is a critical feature of vascular inflammation associated with several disease states such as atherosclerosis. However, mechanisms regulating the endothelial induction of adhesion molecules are not entirely clear. Herein we report that inhibition of the polyol pathway enzyme aldose reductase (AR) prevents the increase in ICAM-1 and VCAM-1 in human umbilical vein endothelial cells (HUVECs) and decreases monocyte adhesion to these cells. In TNF-alpha-stimulated HUVECs, treatment with AR inhibitors sorbinil and tolrestat diminished NF-kappaB activity, phosphorylation and degradation of Ikappa-Balpha, and the nuclear translocation of NF-kappaB. Inhibition of AR abrogated TNF-alpha-induced activation and membrane translocation of PKC, and antisense ablation of AR prevented both TNF alpha-induced PKC and NF-kappaB activation. However, inhibition of AR did not prevent phorbol ester-induced activation of PKC or NF-kappaB, indicating that inhibition of AR does prevents events upstream of PKC activation. These results identify a novel regulator of endothelial activation and suggest that AR is an obligatory mediator of TNF-alpha signaling leading to an increase in the expression of adhesion molecules and increased binding of monocytes to the endothelium. PMID- 15284222 TI - A novel mechanism of regulation of cardiac contractility by mitochondrial functional state. AB - It is generally considered that mitochondria regulate cardiac cell contractility by providing ATP for cellular ATPases and by participating in Ca2+ homeostasis. However, other possible mechanisms by which mitochondria can influence contractility have been largely overlooked. Here, we demonstrate that inhibition of the mitochondrial electron transport chain strongly increases Ca2+-dependent and independent isometric force development in rat ventricular fibers with selectively permeabilized sarcolemma. This effect is unrelated to the ATP generating activity of mitochondria or Ca2+ homeostasis. Furthermore, various conditions that increase K+ accumulation in the mitochondrial matrix (activation of ATP- or Ca2+-dependent K+ channels as well as inhibition of the K+ efflux pathway via the K+/H+ exchanger) induce a similar mechanical response. Modulators of mitochondrial function that augment isometric force also cause swelling of mitochondria in the vicinity of myofibrils in situ, as shown by confocal microscopy. Osmotic compression of intracellular structures abolishes the effect of mitochondria-induced force modulation, suggesting a mechanical basis for the interaction between the organelles. These findings suggest a novel mechanism for cellular regulation of myofibrillar function, whereby increases in mitochondrial volume can impose mechanical constraints inside the cell, leading to an increase in force developed by myofibrils. PMID- 15284223 TI - Molecular and functional analyses of two new calcium-activated chloride channel family members from mouse eye and intestine. AB - Two new calcium-activated chloride channel (CLCA) family members, mCLCA5 and mCLCA6, have been cloned from mouse eye and intestine, respectively. mCLCA5 is highly homologous to hCLCA2, and mCLCA6 is highly homologous to hCLCA4. mCLCA5 is widely expressed with strong expression in eye and spleen, whereas mCLCA6 is primarily expressed in intestine and stomach. mCLCA6 is also expressed as a splice variant lacking exon 8 and part of exon 10 in intestine and stomach. Transfection of tsA201 cells with enhanced green fluorescent protein-tagged versions of the three cDNAs reveals protein products of 155 and 65 kDa for mCLCA5 and mCLCA6 and 145 and 65 kDa for the mCLCA6 splice variant. In vitro translation of mCLCA5 generates a 90-kDa protein that does not appear to be glycosylated. mCLCA6 also generates a 90-kDa protein that is glycosylated to a 110-kDa product, whereas the mCLCA6 splice variant generates an 80-kDa product that is 100 kDa after glycosylation. Treatment of enhanced green fluorescent protein-tagged mCLCA6 with PNGase F (peptide: N-glycosidase F) to remove N-linked glycosyl groups shows a reduction in size of the 65 kDa product to 60 kDa. Consistent with the hypothesis that mCLCA5, mCLCA6, and its splice variant encode calcium activated chloride channels, in HEK293 cells expressing CLCAs ionomycin-evoked increases in intracellular calcium stimulated a current that reversed near Cl(-) equilibrium potential, E(Cl). Furthermore, these currents were inhibited by the chloride channel blocker niflumic acid. Given the prominent role of hCLCA2 in cancer cell adhesion and the unique high level of expression of hCLCA4 in brain, the identification of their murine counterparts presents the opportunity to clarify the role of CLCAs in disease and normal cell physiology. PMID- 15284224 TI - Decay-accelerating factor induction on vascular endothelium by vascular endothelial growth factor (VEGF) is mediated via a VEGF receptor-2 (VEGF-R2)- and protein kinase C-alpha/epsilon (PKCalpha/epsilon)-dependent cytoprotective signaling pathway and is inhibited by cyclosporin A. AB - Decay-accelerating factor (DAF), a membrane-bound complement regulatory protein, is up-regulated on endothelial cells (ECs) following treatment with vascular endothelial growth factor (VEGF), providing enhanced protection from complement mediated injury. We explored the signaling pathways involved in this response. Incubation of human umbilical vein ECs with VEGF induced a 3-fold increase in DAF expression. Inhibition by flk-1 kinase inhibitor SU1498 and failure of placental growth factor (PlGF) to up-regulate DAF confirmed the role of VEGF-R2. The response was also blocked by pretreatment with phospholipase C-gamma (PLCgamma) inhibitor U71322 and protein kinase C (PKC) antagonist GF109203X. In contrast, no effect was seen with nitric oxide synthase inhibitor N(G)-monomethyl-l-arginine (l-NMMA). Use of PKC agonists and isozyme-specific pseudosubstrate peptide antagonists suggested a role for PKCalpha and -epsilon in VEGF-mediated DAF up regulation. This was confirmed by transfection of ECs with PKCalpha and -epsilon dominant-negative constructs, which in combination completely abrogated induction of DAF by VEGF. In contrast, LY290042, a phosphoinositide 3-kinase (PI3K) inhibitor, significantly augmented DAF expression, suggesting a negative regulatory role for phosphoinositide 3-kinase. The widely used immunosuppressive drug cyclosporin A (CsA) inhibited DAF induction by VEGF in a dose-dependent manner. The VEGF-induced DAF expression was functionally effective, significantly reducing complement-mediated EC lysis, and this cytoprotective effect was reversed by CsA. These data provide evidence for a VEGF-R2-, phospholipase C gamma-, and PKCalpha/epsilon-mediated cytoprotective pathway in ECs. This may represent an important mechanism for the maintenance of vascular integrity during chronic inflammation involving complement activation. Moreover, inhibition of this pathway by CsA may play a role in CsA-mediated vascular injury. PMID- 15284225 TI - Proteolytic shedding of the extracellular domain of photoreceptor cadherin. Implications for outer segment assembly. AB - Photoreceptor cadherin (prCAD) is a distinctive cadherin family member that is concentrated at the base of rod and cone outer segments and is required for their structural integrity. During retinal development, prCAD localizes to the site of the future outer segment before rhodopsin or other phototransduction proteins. In vivo, prCAD undergoes a single proteolytic cleavage that releases the ectodomain as a soluble fragment. The C-terminal fragment containing the transmembrane and cytosolic domains remains associated with the outer segment. In rds(-/-) retinas, in which outer segment assembly is severely disrupted because of the absence of retinal degeneration slow (RDS)/peripherin, an essential outer segment structural protein, the level of prCAD is increased, whereas the levels of other outer segment proteins are decreased relative to wild type retinas. Additionally, the ratio of intact:cleaved prCAD polypeptides is increased in rds(-/-) retinas. These data imply that prCAD ectodomain cleavage is an integral part of the outer segment assembly process, and they further suggest that outer segment assembly might be driven, at least in part, by the near irreversibility of proteolysis. PMID- 15284226 TI - The stability of the nuclear lamina polymer changes with the composition of lamin subtypes according to their individual binding strengths. AB - The nuclear lamina, which provides a structural scaffolding for the nuclear envelope, consists largely of a polymer of the intermediate filament lamin proteins. Although different cell types contain distinctive relative amounts of the major lamin subtypes (A, C, B1, and B2), the functions of this variation are not understood. We have investigated the possibility that subtype variation affects lamina stability. We find that homotypic and heterotypic binding interactions of lamin B2 are substantially less resistant to chemical dissociation in vitro than those between the other lamin subtypes, whereas lamin A interactions are the most stable. Surprisingly, removal of the central four fifths of the rod domain did not substantially weaken the interactions of lamins A and B2, suggesting that other regions also strongly contribute to their binding interactions. In contrast, this rod deletion strongly destabilizes the binding interactions of lamins B1 and C. Consistent with the binding studies, lamins are more readily solubilized by chemical extraction from cells enriched for lamin B2 than from cells enriched for lamin A. This suggests that the distinctive ensemble of heterotypic lamin interactions in a particular cell type affects the stability of the lamin polymer, and, correspondingly, could be relevant to tissue-specific properties of the lamina including its involvement in disease. PMID- 15284228 TI - A heteromeric complex of the two nucleotide binding domains of cystic fibrosis transmembrane conductance regulator (CFTR) mediates ATPase activity. AB - The cystic fibrosis transmembrane conductance regulator (CFTR) protein is a member of the ABC superfamily of transporter proteins. Recently, crystal structures of intact, prokaryotic members of this family have been described. These structures suggested that ATP binding and hydrolysis occurs at two sites formed at the interface between their nucleotide binding domains (NBDs). In contrast to the prokaryotic family members, the NBDs of CFTR are asymmetric (both structurally and functionally), and previous to the present studies, it was not clear whether both NBDs are required for ATP hydrolysis. In order to assess the relative roles of the two NBDs of human CFTR, we purified and reconstituted NBD1 and NBD2, separately and together. We found that NBD1 and NBD2 by themselves exhibited relatively low ATPase activity. Co-assembly of NBD1 and NBD2 exhibited a 2-3-fold enhancement in catalytic activity relative to the isolated domains and this increase reflected enhanced ATP turnover (V(max)). These data provide the first direct evidence that heterodimerization of the NBD1 and NBD2 domains of CFTR is required to generate optimal catalytic activity. PMID- 15284227 TI - MAP kinase phosphatase 3 (MKP3) interacts with and is phosphorylated by protein kinase CK2alpha. AB - Mitogen-activated protein (MAP) kinases play a central role in controlling a wide range of cellular functions following their activation by a variety of extracellular stimuli. MAP kinase phosphatases (MKPs) represent a subfamily of dual specificity phosphatases, which negatively regulate MAP kinases. Although ERK2 activity is regulated by its phosphorylation state, MKP3 is regulated by physical interaction with ERK2, independent of its enzymatic activity (Camps, M., Nichols, A., Gillieron, C., Antonsson, B., Muda, M., Chabert, C., Boschert, U., and Arkinstall, S., (1998) Science 280, 1262-1265; Farooq, A., Chaturvedi, G., Mujtaba, S., Plotnikova, O., Zeng, L., Dhalluin, C., Ashton, R., and Zhou, M. M. (2001), Mol. Cell 7, 387-399; Zhou, B., and Zhang, Z. Y. (1999) J. Biol. Chem. 274, 35526-35534). The interaction of ERK2 and MKP3 allows the reciprocal cross regulation of their catalytic activity. Indeed, MKP3 acts as a negative regulator on ERK2-MAP kinase signal transduction activity, representing thus a negative feedback for this MAPK pathway. To identify novel proteins able to complex MKP3, we used the yeast two-hybrid system. Here we report that MKP3 and protein kinase CK2 form a protein complex, which can include ERK2. The phosphatase activity of MKP3 is then slightly increased in vitro, whereas in transfected cells, ERK2 dephosphorylation is reduced. In addition, we demonstrated that CK2 selectively phosphorylates MKP3, suggesting cross-regulation between CK2alpha and MKP3, as well as a modulation of ERK2-MAPK signaling by CK2alpha via MKP3. PMID- 15284229 TI - Rab4 is an essential regulator of lysosomal trafficking in trypanosomes. AB - Rapid endocytosis and recycling of surface proteins are important processes common to most nucleated eukaryotic cells. The best characterized membrane recycling routes are mediated by the small GTPases Rab4 and Rab11, but the precise roles that these pathways play have not been fully elucidated. The protozoan Trypanosoma brucei has a highly developed endocytic system that is similar to that found in metazoans, albeit with an accelerated rate of membrane turnover. We have used this organism to investigate the function of the trypanosome orthologue of Rab4 (TbRAB4) by a combination of RNA interference, microscopy, and quantitative trafficking assays. RNA interference-mediated suppression of TbRAB4 expression inhibited the growth of trypanosomes without affecting receptor-mediated endocytosis or ligand recycling. Ultrastructural analysis indicated a major defect in membrane transport events. The accumulation of fluorescent dextran, a fluid-phase marker, was blocked in cells lacking TbRAB4 protein. Since most fluid-phase markers are transported to the lysosome in T. brucei, the effects of TbRAB4 RNA interference on lysosomal function were investigated. By immunofluorescence, the major lysosomal protein p67 became progressively dispersed in cells lacking the TbRAB4 protein. Pulse-chase analysis demonstrated that initial proteolytic cleavage and glycan processing of p67 were unaffected but that cells failed to accumulate the later p67 proteolyzed products associated with the lysosome. To confirm the role of TbRAB4 in lysosomal trafficking, a constitutively active mutant, TbRAB4QL, was expressed. TbRAB4QL was closely associated with an enlarged multivesicular body that contained p67. In addition, cells expressing TbRAB4QL showed increased fluid-phase uptake when compared with the parental line. Taken together, these data suggest that TbRAB4 is involved in regulation of fluid-phase traffic to the lysosome in T. brucei but not in receptor-mediated endocytosis or recycling. These data have implications for the role of Rab4 in other cell systems. PMID- 15284230 TI - Self-assembly of the cytoskeletal glial fibrillary acidic protein is inhibited by an isoform-specific C terminus. AB - The predominant isoform of glial fibrillary acidic protein (GFAP), GFAPalpha, is the characteristic building block of the cytoskeletal intermediate filaments in astrocytes. Isoform GFAPepsilon, produced by alternative splicing of the GFAP gene, includes a new tail domain that confers a presenilin binding capacity. We here show that the GFAPepsilon tail prevents GFAPepsilon homodimerization and homomeric filament formation, whereas the ability to form heterodimers and filaments with GFAPalpha is retained. Furthermore, GFAPepsilon shows decreased affinity for several GFAPalpha-interacting proteins. A GFAPepsilon tail mutation that results in gain of GFAPepsilon dimerization and filament formation abolishes presenilin binding. This mutation also abolishes interaction between the tail and the coiled-coil domain of GFAPepsilon. Together, this indicates that direct interaction between the coiled-coil and tail domains may serve as an inhibitory mechanism for homomeric dimerization and filament formation. We propose that the GFAPepsilon isoform represents a new functionally distinct component of GFAP intermediate filaments. PMID- 15284231 TI - Fluorescence anisotropy studies on the Ku-DNA interaction: anion and cation effects. AB - DNA non-homologous end joining starts with the binding of Ku heterodimers to double strand breaks. In this work, we characterized the thermodynamics of the Ku DNA interaction by fluorescence anisotropy of the probe-labeled DNA. We determined that the microscopic dissociation constant (kd) for the binding of Ku to a DNA binding site of the proper length (>20 bp) ranges from 22 to 29 nm at 300 mm NaCl. The binding isotherms for DNA duplexes with two or three heterodimers were analyzed with two independent models considering the presence and absence of overlapping binding sites. This analysis demonstrated that there is no or very weak nearest-neighbor cooperativity among the Ku molecules. These models can most likely be applied to study the interaction of Ku with duplexes of any length. Furthermore, our salt dependence studies indicated that electrostatic interactions play a major role in the binding of Ku to DNA and that the kd decreases approximately 60-fold as the salt concentration is lowered from 300 to 200 mm. The slope (Gammasalt) of the plot of log kd versus log[NaCl] is 12.4 +/- 0.1. This value is among the highest reported in the literature for a protein-DNA interaction and suggests that approximately 12 ions are released upon formation of the Ku-DNA complex. In addition, comparison of the slope values measured upon varying the type of cation and anion indicated that approximately nine cations and three anions are released from DNA and Ku, respectively, when the complex is formed. PMID- 15284232 TI - Alternative activation of STAT1 and STAT3 in response to interferon-gamma. AB - Interferon-gamma (IFNgamma) is a pluripotent cytokine whose major biological effects are mediated through a pathway in which STAT1 is the predominant and essential transcription factor. STAT3 can also be activated weakly by IFNgamma, but the mechanism of activation and function of STAT3 as a part of the interferon response are not known. Here we show that STAT3 activation is much stronger and more prolonged in STAT1-null mouse embryo fibroblasts than in wild-type cells. In response to IFNgamma, SRC-family kinases are required to activate STAT3 (but not STAT1) through tyrosine phosphorylation, whereas the receptor-bound kinases JAK1 and JAK2 are required to activate both STATs. Tyrosine 419 of the IFNgamma receptor subunit 1 (IFNGR1) is required to activate both STATs, suggesting that STAT1 and STAT3 compete with each other for the same receptor phosphotyrosine motif. Activated STAT3 can replace STAT1 in STAT1-null cells to drive the transcription of certain genes, for example, socs-3 and c/ebpdelta, which have gamma-activated sequence motifs in their promoters. Work from Ian Kerr's laboratory reveals that the gp130-linked interleukin-6 receptor, which usually activates STAT3 predominantly, activates STAT1 efficiently when STAT3 is absent. Because STAT1 and STAT3 have opposing biological effects (STAT3 is an oncogene, and STAT1 is a tumor suppressor), the reciprocal activation of these two transcription factors in response to IFNgamma or interleukin-6 suggests that their relative abundance, which may vary substantially in different normal cell types, under different conditions or in tumors is likely to have a major impact on how cells behave in response to different cytokines. PMID- 15284233 TI - MEK1 and MEK2, different regulators of the G1/S transition. AB - The ERK cascade is activated by hormones, cytokines, and growth factors that result in either proliferation or growth arrest depending on the duration and intensity of the ERK activation. Here we provide evidence that the MEK1/ERK module preferentially provides proliferative signals, whereas the MEK2/ERK module induces growth arrest at the G1/S boundary. Depletion of either MEK subtype by RNA interference generated a unique phenotype. The MEK1 knock down led to p21cip1 induction and to the appearance of cells with a senescence-like phenotype. Permanent ablation of MEK1 resulted in reduced colony formation potential, indicating the importance of MEK1 for long term proliferation and survival. MEK2 deficiency, in contrast, was accompanied by a massive induction of cyclin D expression and, thus, CDK4/6 activation followed by nucleophosmin hyperphosphorylation and centrosome over-amplification. Our results suggest that the two MEK subtypes have distinct ways to contribute to a regulated ERK activity and cell cycle progression. PMID- 15284234 TI - Dissecting the protein-protein interface between beta-lactamase inhibitory protein and class A beta-lactamases. AB - beta-Lactamase inhibitory protein (BLIP) binds and inhibits a diverse collection of class A beta-lactamases at a wide range of affinities. Alanine-scanning mutagenesis was previously performed to identify the amino acid sequence requirements of BLIP for inhibiting TEM-1 beta-lactamase and SME-1 beta lactamase. Two hotspots of binding energy, one from each domain of BLIP, were identified (Zhang, Z., and Palzkill, T. (2003) J. Biol. Chem. 278, 45706-45712). This study has been extended to examine the amino acid sequence requirements of BLIP for binding to the SHV-1 beta-lactamase, which is a poor binding substrate (Ki= 1.1 microm), and the Bacillus anthracis Bla1 enzyme (Ki= 2.5 nm). The two hotspots previously identified as important for binding TEM-1 and SME-1 beta lactamase were also found to be important for binding Bla1. The hotspot from the second domain of BLIP, however, does not make substantial contributions to SHV-1 binding. This may explain why BLIP binds to SHV-1 beta-lactamase with much weaker affinity than to the other three enzymes. Three regions, including two loops that insert into the active pocket of TEM-1 beta-lactamase and the Glu-73-Lys-74 buried charge motif, exhibit strikingly different effects on the binding affinity of BLIP toward the various enzymes when mutated and, therefore, act as specificity determinants. Analysis of double mutants of BLIP that combine specificity-determining residues suggests that these residues contribute to the poor affinity between the second domain of BLIP and SHV-1 beta-lactamase. PMID- 15284235 TI - Structural analysis of DNA interactions with biogenic polyamines and cobalt(III)hexamine studied by Fourier transform infrared and capillary electrophoresis. AB - Biogenic polyamines, such as putrescine, spermidine, and spermine are small organic polycations involved in numerous diverse biological processes. These compounds play an important role in nucleic acid function due to their binding to DNA and RNA. It has been shown that biogenic polyamines cause DNA condensation and aggregation similar to that of inorganic cobalt(III)hexamine cation, which has the ability to induce DNA conformational changes. However, the nature of the polyamine.DNA binding at the molecular level is not clearly established and is the subject of much controversy. In the present study the effects of spermine, spermidine, putrescine, and cobalt(III)hexamine on the solution structure of calf thymus DNA were investigated using affinity capillary electrophoresis, Fourier transform infrared, and circular dichroism spectroscopic methods. At low polycation concentrations, putrescine binds preferentially through the minor and major grooves of double strand DNA, whereas spermine, spermidine, and cobalt(III)hexamine bind to the major groove. At high polycation concentrations, putrescine interaction with the bases is weak, whereas strong base binding occurred for spermidine in the major and minor grooves of DNA duplex. However, major groove binding is preferred by spermine and cobalt(III)hexamine cations. Electrostatic attractions between polycation and the backbone phosphate group were also observed. No major alterations of B-DNA were observed for biogenic polyamines, whereas cobalt(III)hexamine induced a partial B --> A transition. DNA condensation was also observed for cobalt(III)hexamine cation, whereas organic polyamines induced duplex stabilization. The binding constants calculated for biogenic polyamines are K(Spm) = 2.3 x 10(5) M(-1), K(Spd) = 1.4 x 10(5) M(-1), and K(Put) = 1.02 x 10(5) M(-1). Two binding constants have been found for cobalt(III)hexamine with K(1) = 1.8 x 10(5) M(-1) and K(2) = 9.2 x 10(4) M(-1). The Hill coefficients indicate a positive cooperativity binding for biogenic polyamines and a negative cooperativity for cobalt(III)hexamine. PMID- 15284236 TI - A gain of function mutation in the activation loop of platelet-derived growth factor beta-receptor deregulates its kinase activity. AB - The platelet-derived growth factor receptors (PDGFRs) are receptor tyrosine kinases implicated in multiple aspects of cell growth, differentiation, and survival. Recently, a gain of function mutation in the activation loop of the human PDGFRalpha has been found in patients with gastrointestinal stromal tumors. Here we show that a mutation in the corresponding codon in the activation loop of the murine PDGFRbeta, namely an exchange of asparagine for aspartic acid at amino acid position 849 (D849N), confers transforming characteristics to embryonic fibroblasts from mutant mice, generated by a knock-in strategy. By comparing the enzymatic properties of the wild-type versus the mutant receptor protein, we demonstrate that the D849N mutation lowers the threshold for kinase activation, causes a dramatic alteration in the pattern of tyrosine phosphorylation kinetics following ligand stimulation, and induces a ligand-independent phosphorylation of several tyrosine residues. These changes result in deregulated recruitment of specific signal transducers. The GTPase-activating protein for Ras (RasGAP), a negative regulator of the Ras mitogenic pathway, displayed a delayed binding to the mutant receptor. Moreover, we have observed enhanced ligand-independent ERK1/2 activation and an increased proliferation of mutant cells. The p85 regulatory subunit of the phosphatidylinositol 3 '-kinase was constitutively associated with the mutant receptor, and this ligand-independent activation of the phosphatidylinositol 3'-kinase pathway may explain the observed strong protection against apoptosis and increased motility in cellular wounding assays. Our findings support a model whereby an activating point mutation results in a deregulated PDGFRbeta with oncogenic predisposition. PMID- 15284237 TI - The replicative regulator protein geminin on chromatin in the HeLa cell cycle. AB - Geminin is believed to have a major function in the regulation of genome replication and cell proliferation. Published evidence shows that geminin specifically interacts with Cdt1 to block its function in the assembly of prereplication complexes. However, in proliferating HeLa cells geminin and Cdt1 are co-expressed during a relatively short time at the G(1)-to-S phase transition. Under these conditions, nearly all Cdt1 and a major part of geminin are bound to chromatin and reside at the same or closely adjacent sites as shown here by chromatin immunoprecipitation. Cdt1 is rapidly degraded early in S phase, but geminin remains bound to the chromatin sites. One function that chromatin bound geminin could perform is to prevent access to Cdt1 that may escape S phase dependent degradation or is synthesized in excess. Indeed, Cdt1 continues to be synthesized in HeLa cells in S phase but never accumulates because of the efficient degradation. Therefore, geminin can be eliminated by RNA interference without detectable effects on cell cycle parameters. PMID- 15284238 TI - Deamidation affects structural and functional properties of human alphaA crystallin and its oligomerization with alphaB-crystallin. AB - To determine the effects of deamidation on structural and functional properties of alphaA-crystallin, three mutants (N101D, N123D, and N101D/N123D) were generated. Deamidated alphaB-crystallin mutants (N78D, N146D, and N78D/N146D), characterized in a previous study (Gupta, R., and Srivastava, O. P. (2004) Invest. Ophthalmol. Vis. Sci. 45, 206-214) were also used. The biophysical and chaperone properties were determined in (a) homoaggregates of alphaA mutants (N101D, N123D, and N101D/N123D) and (b) reconstituted heteroaggregates of alpha crystallin containing (i) wild type alphaA (WT-alphaA): WT-alphaB crystallins, (ii) individual alphaA-deamidated mutants:WT-alphaB crystallins, and (iii) WT alphaA:individual alphaB-deamidated mutant crystallins. Compared with the WT alphaA, the three alphaA-deamidated mutants showed reduced levels of chaperone activity, alterations in secondary and tertiary structures, and larger aggregates. These altered properties were relatively more pronounced in the mutant N101D compared with the mutant N123D. Further, compared with heteroaggregates of WT-alphaA and WT-alphaB, the heteroaggregates containing deamidated subunits of either alphaA- or alphaB-crystallins and their counterpart WT proteins showed higher molecular mass, altered tertiary structures, lower exposed hydrophobic surfaces, and reduced chaperone activity. However, the heteroaggregate containing WT-alphaA and deamidated alphaB subunit showed lower chaperone activity, smaller oligomers, and 3-fold lower subunit exchange rate than heteroaggregate containing deamidated alphaA- and WT-alphaB subunits. Together, the results suggested that (a) both Asn residues (Asn-101 and Asn-123) are required for the structural integrity and chaperone function of alphaA crystallin and (b) the presence of WT-alphaB in the alpha-crystallin heteroaggregate leads to packing-induced structural changes which influences the oligomerization and modulate chaperone activity. PMID- 15284239 TI - Active mutants of the human p38alpha mitogen-activated protein kinase. AB - Mitogen-activated protein (MAP) kinases compose a family of serine/threonine kinases that function in many signal transduction pathways and affect various cellular phenotypes. Despite the abundance of available data, the exact role of each MAP kinase is not completely defined, in part because of the inability to activate MAP kinase molecules individually and specifically. Based on activating mutations found in the yeast MAP kinase p38/Hog1 (Bell, M., Capone, R., Pashtan, I., Levitzki, A., and Engelberg, D. (2001) J. Biol. Chem. 276, 25351-25358), we designed and constructed single and multiple mutants of human MAP kinase p38alpha. Single (p38D176A, p38F327L, and p38F327S) and subsequent double (p38D176A/F327L and p38D176A/F327S) mutants acquired high intrinsic activity independent of any upstream regulation and reached levels of 10 and 25%, respectively, in reference to the dually phosphorylated wild type p38alpha. The active p38 mutants have retained high specificity toward p38 substrates and were inhibited by the specific p38 inhibitors SB-203580 and PD-169316. We also show that similar mutations can render p38gamma active as well. Based on the available structures of p38 and ERK2, we have analyzed the p38 mutants and identified a hydrophobic core stabilized by three aromatic residues, Tyr-69, Phe-327, and Trp 337, in the vicinity of the L16 loop region. Upon activation, a segment of the L16 loop, including Phe-327, becomes disordered. Structural analysis suggests that the active p38 mutants emulate the conformational changes imposed naturally by dual phosphorylation, namely, destabilization of the hydrophobic core. Essentially, the hydrophobic core is an inherent stabilizer that maintains low basal activity level in unphosphorylated p38. PMID- 15284240 TI - The ALX Src homology 2 domain is both necessary and sufficient to inhibit T cell receptor/CD28-mediated up-regulation of RE/AP. AB - Activation of naive T cells occurs when two signals are received. The first signal is received through the T cell antigen receptor (TCR), and a second costimulatory signal is primarily provided by CD28. We have recently identified a novel adaptor molecule, ALX, which is expressed exclusively in hematopoietic cells. ALX contains several sites for potential protein-protein interaction, including an Src homology 2 (SH2) domain, four PXXP polyproline sequences, and two likely sites of tyrosine phosphorylation. Overexpression of ALX inhibits the transcriptional activation of the interleukin 2 promoter during T cell activation, specifically affecting CD28-mediated activation of the RE/AP element of the interleukin 2 promoter. To understand how ALX functions downstream of CD28, we generated a panel of site-directed mutants as well as truncations in which potential protein-binding sites were mutated or absent. We found that the ALX SH2 domain is both necessary and sufficient to mediate inhibition of RE/AP activation. Mutation of the SH2 domain did not affect ALX expression, relative localization in the cytoplasm and nucleus, phosphorylation, or a mobility shift in response to TCR signaling alone. However, an activation-induced mobility shift triggered by CD28 was reduced in the ALX SH2 domain mutant. In addition, the isolated ALX SH2 domain was found to associate with a phosphoprotein from Jurkat T cells on TCR/CD28 stimulation. Therefore, the ALX SH2 domain plays a critical role in ALX function downstream of CD28. PMID- 15284241 TI - Caspase-3-induced truncation of type 1 inositol trisphosphate receptor accelerates apoptotic cell death and induces inositol trisphosphate-independent calcium release during apoptosis. AB - Inositol 1,4,5-trisphosphate receptor-deficient (IP3RKO) B-lymphocytes were used to investigate the functional relevance of type 1 inositol 1,4,5-trisphosphate receptor (IP3R1) and its cleavage by caspase-3 in apoptosis. We showed that inositol 1,4,5-trisphosphate receptor-deficient cells were largely resistant to apoptosis induced by both staurosporine (STS) and B-cell receptor (BCR) stimulation. Expression of either the wild-type IP3R1 or an N-terminal deletion mutant (Delta1-225) that lacks inositol 1,4,5-trisphosphate-induced Ca2+ release activity restored sensitivity to apoptosis and the consequent rise in free cytosolic Ca2+ concentration ([Ca2+]i). Expression of caspase-3-non-cleavable mutant receptor, however, dramatically slowed down the rate of apoptosis and prevented both Ca2+ overload and secondary necrosis. Conversely, expression of the "channel-only" domain of IP3R1, a fragment of the receptor generated by caspase-3 cleavage, strongly increased the propensity of the cells to undergo apoptosis. In agreement with these observations, caspase inhibitors impeded apoptosis and the associated rise in [Ca2+]i. Both the staurosporine- and B-cell receptor-induced apoptosis and increase in [Ca2+]i could be induced in nominally Ca2+-free and serum-free culture media, suggesting that the apoptosis-related rise in [Ca2+]i was primarily because of the release from internal stores rather than of influx through the plasma membrane. Altogether, our results suggest that IP3R1 plays a pivotal role in apoptosis and that the increase in [Ca2+]i during apoptosis is mainly the consequence of IP3R1 cleavage by caspase-3. These observations also indicate that expression of a functional IP3R1 per se is not enough to generate the significant levels of cytosolic Ca2+ needed for the rapid execution of apoptosis, but a prior activation of caspase-3 and the resulting truncation of the IP3R1 are required. PMID- 15284242 TI - Zonation of labeling of lipogenic acetyl-CoA across the liver: implications for studies of lipogenesis by mass isotopomer analysis. AB - Measurement of fractional lipogenesis by condensation polymerization methods assumes constant enrichment of lipogenic acetyl-CoA in all hepatocytes. mass isotopomer distribution analysis (MIDA) and isotopomer spectral analysis (ISA) represent such methods and are based on the combinatorial analyses of mass isotopomer distributions (MIDs) of fatty acids and sterols. We previously showed that the concentration and enrichment of [13C]acetate decrease markedly across the dog liver because of the simultaneous uptake and production of acetate. To test for zonation of the enrichment of lipogenic acetyl-CoA, conscious dogs, prefitted with transhepatic catheters, were infused with glucose and [1,2 13C2]acetate in a branch of the portal vein. Analyses of MIDs of fatty acids and sterols isolated from liver, bile, and plasma very low density lipoprotein by a variant of ISA designed to detect gradients in precursor enrichment revealed marked zonation of enrichment of lipogenic acetyl-CoA. As control experiments where no zonation of acetyl-CoA enrichment would be expected, isolated rat livers were perfused with 10 mm [1,2-13C2]acetate. The ISA analyses of MIDs of fatty acids and sterols from liver and bile still revealed a zonation of acetyl-CoA enrichment. We conclude that zonation of hepatic acetyl-CoA enrichment occurs under a variety of animal models and physiological conditions. Failure to consider gradients of precursor enrichment can lead to underestimations of fractional lipogenesis calculated from the mass isotopomer distributions. The degree of such underestimation was modeled in vitro, and the data are reported in the companion paper (Bederman, I. R., Kasumov, T., Reszko, A. E., David, F., Brunengraber, H., and Kelleher, J. K. (2004) J. Biol. Chem. 279, 43217-43226). PMID- 15284243 TI - In vitro modeling of fatty acid synthesis under conditions simulating the zonation of lipogenic [13C]acetyl-CoA enrichment in the liver. AB - In the companion report (Bederman, I. R., Reszko, A. E., Kasumov, T., David, F., Wasserman, D. H., Kelleher, J. K., and Brunengraber, H. (2004) J. Biol. Chem. 279, 43207-43216), we demonstrated that, when the hepatic pool of lipogenic acetyl-CoA is labeled from [13C]acetate, the enrichment of this pool decreases across the liver lobule. In addition, estimates of fractional synthesis calculated by isotopomer spectral analysis (ISA), a nonlinear regression method, did not agree with a simpler algebraic two-isotopomer method. To evaluate differences between these methods, we simulated in vitro the synthesis of fatty acids under known gradients of precursor enrichment, and known values of fractional synthesis. First, we synthesized pentadecanoate from [U-13C3]propionyl CoA and four gradients of [U-13C3]malonyl-CoA enrichment. Second, we pooled the fractions of each gradient. Third, we diluted each pool with pentadecanoate prepared from unlabeled malonyl-CoA to simulate the dilution of the newly synthesized compound by pre-existing fatty acids. This yielded a series of samples of pentadecanoate with known values of (i) lower and upper limits for the precursor enrichment, (ii) the shape of the gradient, and (iii) the fractional synthesis. At each step, the mass isotopomer distributions of the samples were analyzed by ISA and the two-isotopomer method to determine whether each method could correctly (i) detect gradients of precursor enrichment, (ii) estimate the gradient limits, and (iii) estimate the fractional synthesis. The two-isotopomer method did not identify gradients of precursor enrichment and underestimated fractional synthesis by up to 2-fold in the presence of gradients. ISA uses all mass isotopomers, correctly identified imposed gradients of precursor enrichment, and estimated the expected values of fractional synthesis within the constraints of the data. PMID- 15284244 TI - Reverse signaling through membrane-bound interleukin-15. AB - The results from this study implicate membrane-anchored interleukin (IL)-15 constitutively expressed on the cell surface of PC-3 human prostate carcinoma cells and interferon-gamma-activated human monocytes in reverse signaling upon stimulation with soluble IL-15 receptor-alpha or anti-IL-15 antibodies, mediating the outside-to-inside signal transduction that involves the activation of members of the MAPK family (ERK and p38) and focal adhesion kinase. The presence of membrane-bound IL-15 was not dependent on the expression of the trimeric IL-15 receptor complex by these cells and resisted treatment with acidic buffer or trypsin. Reverse signaling through membrane-bound IL-15 considerably increased the production of several pro-inflammatory cytokines by monocytes, such as IL-6, IL-8, and tumor necrosis factor-alpha, thereby indicating the relevance of this process to the complex immunomodulatory function of these cells. Furthermore, stimulation of transmembrane IL-15 also enhanced the transcription of IL-6 and IL 8 in the PC-3 cell line and promoted migration of PC-3 cells as well as LNCaP human prostate carcinoma cells stably expressing IL-15 on the cell surface. Thus, IL-15 can exist as a biologically active transmembrane molecule that possesses dual ligand-receptor qualities with a potential to induce bidirectional signaling. This fact highlights a new level of complexity in the biology of IL-15 and offers novel important insights into our understanding of the cellular responses modulated by this pleiotropic cytokine. PMID- 15284245 TI - Prostratin antagonizes HIV latency by activating NF-kappaB. AB - A subset of quiescent memory CD4 T cells harboring integrated but transcriptionally silent proviruses poses a currently insurmountable barrier to the eradication of the human immunodeficiency virus (HIV) in infected patients. Induction of HIV gene expression in these latently infected cells by immune activating agents has been proposed as one approach to confer sensitivity to antiretroviral therapy. Interest has recently focused on the non-tumor-promoting phorbol ester, prostratin, as a potential agent to activate latent HIV proviruses. Using multiple Jurkat T cell lines containing integrated but transcriptionally latent HIV proviruses (J-Lat cells), we now demonstrate that prostratin effectively activates HIV gene expression in these latently infected cells. We further show that prostratin acts by stimulating IKK-dependent phosphorylation and degradation of IkappaBalpha, leading to the rapid nuclear translocation of NF-kappaB and activation of the HIV-1 long terminal repeat in a kappaB enhancer-dependent manner. In contrast, NFAT and AP-1 are not induced by prostratin. Using chromatin immunoprecipitation assays to identify host transcription factors recruited to the latent HIV-1 promoter in living cells, we find that prostratin induces RelA binding. Analysis of potential upstream signal transducers demonstrates that prostratin stimulates membrane translocation of classical, novel, and atypical protein kinase C (PKC) isoforms. Studies with isoform-specific PKC inhibitors suggest that the novel PKCs play a particularly prominent role in the prostratin response. These findings provide new insights into the molecular pathway through which prostratin antagonizes HIV latency highlighting a central role for the action of NF-kappaB. PMID- 15284246 TI - Distinct roles of two structurally closely related focal adhesion proteins, alpha parvins and beta-parvins, in regulation of cell morphology and survival. AB - Proteins at cell-extracellular matrix adhesions (e.g. focal adhesions) are crucially involved in regulation of cell morphology and survival. We show here that CH-ILKBP/actopaxin/alpha-parvin and affixin/beta-parvin (abbreviated as alpha- and beta-parvin, respectively), two structurally closely related integrin linked kinase (ILK)-binding focal adhesion proteins, are co-expressed in human cells. Depletion of alpha-parvin dramatically increased the level of beta-parvin, suggesting that beta-parvin is negatively regulated by alpha-parvin in human cells. Loss of PINCH-1 or ILK, to which alpha- and beta-parvin bind, significantly reduced the activation of Rac, a key signaling event that controls lamellipodium formation and cell spreading. We were surprised to find that loss of alpha-parvin, but not that of beta-parvin, markedly stimulated Rac activation and enhanced lamellipodium formation. Overexpression of beta-parvin, however, was insufficient for stimulation of Rac activation or lamellipodium formation, although it was sufficient for promotion of apoptosis, another important cellular process that is regulated by PINCH-1, ILK, and alpha-parvin. In addition, we show that the interactions of ILK with alpha- and beta-parvin are mutually exclusive. Overexpression of beta-parvin or its CH(2) fragment, but not a CH(2) deletion mutant, inhibited the ILK-alpha-parvin complex formation. Finally, we provide evidence suggesting that inhibition of the ILK-alpha-parvin complex is sufficient, although not necessary, for promotion of apoptosis. These results identify Rac as a downstream target of PINCH-1, ILK, and parvin. Furthermore, they demonstrate that alpha- and beta-parvins play distinct roles in mammalian cells and suggest that the formation of the ILK-alpha-parvin complex is crucial for protection of cells from apoptosis. PMID- 15284247 TI - Identification of CC chemokine receptor 7 residues important for receptor activation. AB - The binding pocket of family A GPCRs that bind small biogenic amines is well characterized. In this study we identify residues on CC chemokine receptor 7 (CCR 7) that are involved in agonist-mediated receptor activation but not in high affinity ligand binding. The mutations also affect the ability of the ligands to induce chemotaxis. Two of the residues, Lys3.33(137) and Gln5.42(227), are consistent with the binding pocket described for biogenic amines, while Lys3.26(130) and Asn7.32(305), are found at, or close to, the cell surface. Our observations are in agreement with findings from other peptide and chemokine receptors, which indicate that receptors that bind larger ligands contain contact sites closer to the cell surface in addition to the conventional transmembrane binding pocket. These findings also support the theory that chemokine receptors require different sets of interactions for high affinity ligand binding and receptor activation. PMID- 15284248 TI - Repression of hsp90beta gene by p53 in UV irradiation-induced apoptosis of Jurkat cells. AB - Tumor suppressor p53 has been implicated in cell stress response and determines cell fate of either growth arrest or apoptosis. Heat shock proteins (Hsps) expressed under stress usually confer survival protection to the cell or interruption in the apoptotic pathways. Although Hsp90 can physically interact with p53, whether or not the hsp90 gene is influenced downstream of p53 in UV irradiation-induced apoptosis remains unclear. We have found that the level of p53 is elevated with the decline of Hsp90 in UV-irradiated cells and that malfunction of Hsp90, as inhibited by geldanamycin, enhances the p53-involved UV irradiation-induced apoptosis. In addition, the expression of the hsp90beta gene was reduced in both UV-irradiated and wild type p53-transfected cells. These results suggest a negative correlation between the trans factor p53 and a chaperone gene hsp90beta in apoptotic cells. Mutation analysis demonstrated that the p53 binding site in the first exon was indispensable for p53 regulation on the hsp90beta gene. In addition, with p53 bound at the promoter of the hsp90beta gene, mSin3a and p300 were differentially recruited in UV irradiation-treated or untreated Jurkat cells in vivo. The evidence of p53-repressed hsp90beta gene expression in UV-irradiated cells shed light on a novel pathway of Hsp90 in the survival control of the stressed cells. PMID- 15284249 TI - Developing a system to assess the quality of cancer care: ASCO's national initiative on cancer care quality. PMID- 15284250 TI - Understanding cancer treatment and outcomes: the Cancer Care Outcomes Research and Surveillance Consortium. PMID- 15284251 TI - Managing accrual in cooperative group clinical trials. PMID- 15284252 TI - A randomized trial comparing defined-duration with continuous irinotecan until disease progression in fluoropyrimidine and thymidylate synthase inhibitor resistant advanced colorectal cancer. AB - PURPOSE: Irinotecan given until disease progression is an accepted standard treatment for advanced colorectal cancer (CRC) resistant to fluoropyrimidines. It is not known whether a predefined period of irinotecan treatment would result in similar duration of disease control. We performed a multicenter phase III trial to compare the two policies of defined-duration versus continuous irinotecan treatment. PATIENTS AND METHODS: Three hundred thirty-three eligible patients with advanced CRC progressing on or within 24 weeks of completing fluoropyrimidine-based chemotherapy were prospectively registered. After receiving eight cycles of irinotecan given at 350 mg/m2 once every 3 weeks, 55 patients with responding or stable disease were randomly assigned to stop irinotecan (n = 30) or continue until disease progression (n = 25). Registered patients were not randomly assigned predominantly due to disease progression (n = 236) and intolerable toxicity (n = 38). RESULTS: From the time of random assignment, there were no differences in failure-free survival (P = .999) or overall survival (P = .11) between the two arms. No difference was seen in mean global health status quality-of-life score between the two arms at 12 weeks after random assignment. No grade 3 diarrhea and febrile neutropenia was seen in the continue-irinotecan arm after random assignment. CONCLUSION: For most patients, the decision to continue on irinotecan beyond 24 weeks is influenced by disease progression or treatment-related toxicity. However, for 17% of patients in whom this decision is clinically relevant, there seems to be little benefit from continuing irinotecan, though the drug was well tolerated without any deterioration in quality of life. PMID- 15284253 TI - Ongoing monoclonal B-cell proliferation is not common in gastric B-cell lymphoma after combined radiochemotherapy. AB - PURPOSE: Gastric marginal-zone B-cell lymphoma (MZBCL) of the mucosa-associated lymphoid tissue (MALT) is associated with chronic Helicobacter pylori gastritis. Stable complete remission (CR) can be induced by H pylori eradication. Whether this is paralleled by cure of the lymphoma remains unclear. Persisting monoclonal bands for immunoglobulin heavy chain variable region (VH) representing the lymphoma clone have been described in up to 50% of patients in CR. This retrospective study investigated whether this phenomenon also occurs after radiochemotherapy. PATIENTS AND METHODS: Biopsy samples of 20 patients receiving chemotherapy with cyclophosphamide, doxorubicin, vincristine, and prednisone and irradiation were analyzed before and after therapy. Study patients had Ann Arbor stage I/II primary gastric cancer, including four cases of MZBCL of MALT type, 12 cases of diffuse large-cell lymphomas (DLCL), and four cases of mixed MALT type/DLCL. Polymerase chain reaction (PCR) for VH rearrangement was performed. Monoclonal PCR products were cloned and sequenced. RESULTS: Fourteen of 20 patients had a monoclonal or oligoclonal band distribution at diagnosis converted into polyclonal pattern after radiochemotherapy. Of the remaining six patients, two were lost to follow-up. One patient did not respond and died of progressive disease. PCR in this patient showed persistent B-cell clonality. In three patients, the initial PCR showed a polyclonal pattern and thus could not be evaluated during follow-up. CONCLUSION: In contrast with H pylori eradication alone, radiochemotherapy results in clearing of monoclonal cells during follow up. This may result in better elimination of residual lymphoma cells. Further study is needed to determine whether this translates into lower risk of relapse. PMID- 15284254 TI - Utility of fine-needle aspiration as a diagnostic technique in lymphoma. AB - PURPOSE: To evaluate, from a clinician's perspective, the sensitivity and specificity of fine-needle aspiration (FNA) as a technique for the diagnosis of lymphoma. PATIENTS AND METHODS: Medical records of 470 new patients seen in one lymphoma specialist's clinic from January 1998 through December 2002 were reviewed. Ninety-nine (21%) of the 470 patients underwent a total of 115 FNA procedures, which were assessed by more than 70 different pathologists in 32 different pathology departments. Subsequent excisional biopsies were performed in 67 of these patients and interpreted by a single hematopathology group without independent review. RESULTS: Of 115 FNA procedures, 93 were completed for the initial evaluation of lymphoma and 22 were done for assessment of relapsed disease. Of the 93 FNA attempts at initial diagnosis, only 27 (29%) were given a specific and complete histologic diagnosis using an accepted classification system (Working Formulation, Revised European-American Classification of Lymphoid Neoplasms, WHO). For the 22 FNAs done for recurrent disease, only nine (41%) were classified using an accepted system. Sixty-seven (72%) of the 93 FNAs performed for the evaluation of initial disease had subsequent excisional biopsies. Among these paired comparisons, only eight (12%) of 67 FNA diagnoses were correlated with the subsequent excisional biopsy diagnosis. Immunophenotyping was completed on 24 of the 67 paired FNAs. Seven of the 24 FNAs with immunophenotyping (29%) were correlated with subsequent histology on excisional biopsy. Only one (2%) of 43 FNA diagnoses, based on morphology alone, was correlated with subsequent excisional biopsy diagnosis. CONCLUSION: Overall, FNA for lymphoma diagnosis is not helpful, not cost effective, and in addition may misguide treatment. PMID- 15284255 TI - Phase II study of neoadjuvant carboplatin and paclitaxel followed by radiotherapy and concurrent cisplatin in patients with locoregionally advanced nasopharyngeal carcinoma: therapeutic monitoring with plasma Epstein-Barr virus DNA. AB - PURPOSE: To assess the efficacy of neoadjuvant paclitaxel and carboplatin (TC) followed by concurrent cisplatin and radiotherapy (RT) in patients with locoregionally advanced nasopharyngeal carcinoma (NPC) and to monitor treatment response with plasma Epstein-Barr virus (EBV) DNA. PATIENTS AND METHODS: Thirty one patients with International Union Against Cancer stages III and IV undifferentiated NPC had two cycles of paclitaxel (70 mg/m2 on days 1, 8, and 15) and carboplatin (area under the curve 6 mg/mL/min on day 1) on a 3-weekly cycle, followed by 6 to 8 weeks of cisplatin (40 mg/m2 weekly) and RT at 66 Gy in 2-Gy fractions. Plasma EBV DNA was measured serially using the real-time quantitative polymerase chain reaction method. Results All patients completed planned treatment. Response to neoadjuvant TC was as follows: 12 patients (39%) achieved partial response (PR) and 18 achieved (58%) complete response (CR) in regional nodes; five patients (16%) achieved PR and no patients achieved CR in nasopharynx. At 6 weeks after RT, one patient (3%) achieved PR and 30 patients (97%) achieved CR in regional nodes, and 31 patients (100%) achieved CR in nasopharynx; 29 patients (93%) had EBV DNA level of less than 500 copies/mL. Neoadjuvant TC was well tolerated, and the most common acute toxicity of cisplatin plus RT was grade 3 mucositis (55%). At median follow-up of 33.7 months (range, 7 to 39.3 months), six distant and three locoregional failures occurred. Plasma EBV DNA level increased significantly in eight of nine patients who experienced treatment failure but did not increase in those who did not. The 2 year overall and progression-free survival rates were 91.8% and 78.5%, respectively. CONCLUSION This strategy was feasible and resulted in excellent local tumor control. Serial plasma EBV DNA provides a noninvasive method of monitoring response in NPC. PMID- 15284256 TI - Cisplatin, fluorouracil, and leucovorin induction chemotherapy followed by concurrent cisplatin chemoradiotherapy for organ preservation and cure in patients with advanced head and neck cancer: long-term follow-up. AB - PURPOSE: The poor functional outcome in patients with advanced head and neck squamous cell carcinoma (HNSCC) with surgery and radiation has led to alternative approaches to advanced disease. We conducted a phase II study of induction chemotherapy followed by concurrent chemoradiotherapy for organ preservation in patients with advanced resectable and unresectable (nasopharyngeal) tumors. PATIENTS AND METHODS: Forty-two patients with stage III to IV resectable HNSCC and nasopharyngeal tumors received induction chemotherapy with two courses of cisplatin (20 mg/m2/d continuous infusion [CI]), fluorouracil (800 mg/m2/d CI), and leucovorin (500 mg/m2/d CI; PFL) for 4 days followed by concurrent therapy with cisplatin (100 mg/m2/d on days 1 and 22) and approximately 70 Gy of external beam radiotherapy. RESULTS: Response to induction chemotherapy included partial response rate of 52% and complete response rate of 24%. The most common grade 3 or 4 toxicity was neutropenia (59%). After cisplatin chemoradiotherapy the complete response rate was 67%. Toxicities of cisplatin chemoradiotherapy consisted of grade 3 or 4 mucositis (79%) and neutropenia (51%). At a median follow-up of 71.5 months, 43% of the patients are still alive and disease-free. The 5-year progression-free survival (PFS) rate was 60%, and the 2- and 5-year overall survival (OS) rates were 67% and 52%, respectively. Three patients died of second primaries. Late complications of treatment included xerostomia and hoarseness. One patient had persistent dysphagia and required laser epiglotectomy 108 months after treatment. CONCLUSION: Induction chemotherapy with PFL followed by concurrent cisplatin chemoradiotherapy is well tolerated and results in a good likelihood of organ preservation and excellent PFS and OS. PMID- 15284257 TI - Long-term cardiac follow-up in relapse-free patients after six courses of fluorouracil, epirubicin, and cyclophosphamide, with either 50 or 100 mg of epirubicin, as adjuvant therapy for node-positive breast cancer: French adjuvant study group. AB - PURPOSE To evaluate long-term cardiac function in patients without disease who had received six cycles of fluorouracil 500 mg/m(2), epirubicin 50 mg/m(2), and cyclophosphamide 500 mg/m(2) (FEC 50) or the same regimen with epirubicin 100 mg/m(2) (FEC 100) as adjuvant chemotherapy for node-positive breast cancer in the French Adjuvant Study Group-05 trial. PATIENTS AND METHODS One hundred fifty patients (FEC 50, n = 65; FEC 100, n = 85) who were without disease and who gave their informed consent were enrolled for long-term cardiac assessment. The assessment included cardiac events occurring after the end of chemotherapy, vital signs, concomitant disease, ECG, isotopic left ventricular ejection fraction (LVEF), and echographic parameters. Abnormal files were blindly reviewed by cardiologists and oncologists. Results The median follow-up time was 102 months. After FEC 100, LVEF was less than 50% in five patients (radioisotopic method), and two patients experienced congestive heart failure (CHF) that was possibly related to treatment. Asymptomatic left ventricular dysfunction (LVD) was experienced in 18 patients after FEC 100 and in one patient after FEC 50. In these patients, treatment causality was probable in eight patients. Two additional years after this assessment, all 18 patients were still asymptomatic. CONCLUSION After more than 8 years of follow-up, the cardiac toxicity observed after adjuvant treatment with FEC 100 comprised two cases of well-controlled CHF and 18 cases of asymptomatic LVD. In the majority of women with primary breast cancer, the benefits of treatment with FEC 100 in terms of disease-free and overall survival outweigh the risks, and cardiac risk factors should be carefully evaluated in patient selection. PMID- 15284258 TI - Evaluation of biologic end points and pharmacokinetics in patients with metastatic breast cancer after treatment with erlotinib, an epidermal growth factor receptor tyrosine kinase inhibitor. AB - PURPOSE: To evaluate changes in epidermal growth factor receptor (EGFR) phosphorylation and its downstream signaling in tumor and surrogate tissue biopsies in patients with metastatic breast cancer treated with erlotinib, an EGFR tyrosine kinase inhibitor, and to assess relationships between biomarkers in tumor and normal tissues and between biomarkers and pharmacokinetics. PATIENTS AND METHODS: Eighteen patients were treated orally with 150 mg/d of erlotinib. Ki67, EGFR, phosphorylated EGFR (pEGFR), phosphorylated mitogen-activated protein kinase (pMAPK), and phosphorylated AKT (pAKT) in 15 paired tumor, skin, and buccal mucosa biopsies (at baseline and after 1 month of therapy) were examined by immunohistochemistry and analyzed quantitatively. Pharmacokinetic sampling was also obtained. RESULTS: The stratum corneum layer and Ki67 in keratinocytes of the epidermis in 15 paired skin biopsies significantly decreased after treatment (P = .0005 and P = .0003, respectively). No significant change in Ki67 was detected in 15 tumors, and no responses were observed. One was EGFR-positive and displayed heterogeneous expression of the receptor, and 14 were EGFR-negative. In the EGFR-positive tumor, pEGFR, pMAPK, and pAKT were reduced after treatment. Paradoxically, pEGFR was increased in EGFR-negative tumors post-treatment (P = .001). Although markers were reduced in surrogate and tumor tissues in the patient with EGFR-positive tumor, no apparent associations were observed in patients with EGFR-negative tumor. CONCLUSION: Erlotinib has inhibitory biologic effects on normal surrogate tissues and on an EGFR-positive tumor. The lack of reduced tumor proliferation may be attributed to the heterogeneous expression of receptor in the EGFR-positive patient and absence of target in this cohort of heavily pretreated patients. PMID- 15284259 TI - Patient-physician concordance: preferences, perceptions, and factors influencing the breast cancer surgical decision. AB - PURPOSE: This study explored patient preferences for involvement in the breast cancer treatment decision and concordance between patients' and physicians' views on decisional role. The impact of demographic and psychosocial characteristics on patients' decisional role was also examined. PATIENTS AND METHODS: Women with stage I or II breast cancer who were candidates for either mastectomy or lumpectomy were recruited from a university breast cancer treatment center. Patient interviews were obtained before meeting the surgical oncologist and again after the treatment decision was made but before surgical intervention. Clinician responses were obtained after the consultation. RESULTS: The 101 participants were generally white (97%), married (80%), and well-educated. They reported moderate levels of depression and anxiety but good social support and self efficacy in communicating with their physician. Before the consultation, 47% of women reported a preference for shared decision making; afterwards, 61% felt they had primary responsibility for the decision. Only 38% of patients agreed with the physician's assessment of how the treatment decision was made. In regression analyses, higher education was significantly associated with patients' preferred level of control (P = .01). There was a trend toward women with greater self efficacy desiring more active decisional roles (P = .08). Patient preference for decision making did not impact time in the patient-physician encounter, but more influence did increase satisfaction. CONCLUSION: Limited concordance between patient preference and patient perception and between patient and physician perception in how the treatment decision was made suggests the need for better communication between patient and clinician during a critical treatment encounter for breast cancer patients. PMID- 15284260 TI - Differential prognostic impact of comorbidity. AB - PURPOSE: Cancer patients with concurrent comorbid conditions have worse outcomes than patients with no comorbidities. We hypothesized that the prognostic impact of comorbidities would be greatest for patients with cancers associated with a long natural history and least in patients with aggressive cancers. PATIENTS AND METHODS: Using the Barnes-Jewish Hospital Oncology Data Services cancer registry, we grouped 11,558 patients with breast, lung, colon, or prostate cancer by morphologic stage at diagnosis and then determined the 1-year overall survival rate for each group. Overall, severity of comorbidity was assessed from chart review and classified into one of four groups: none, mild, moderate, or severe. The relative prognostic impact of comorbidity was measured by the hazard ratio and adjusted for the prognostic impact of age, race, and sex. RESULTS: One-year overall survival rate ranged from 20% for 1,005 patients with distant spread of lung cancer to 98% for 3,325 patients with localized prostate cancer. Adjusted hazard ratio of moderate/severe comorbidity (relative to none/mild) ranged from 1.04 to 4.48. The correlation between overall survival rate and severity of comorbidity was statistically significant (r2 = 0.56; P < .001). The proportion of variance in outcome explained by comorbidity ranged from less than 1% to almost 9%, depending on tumor site and stage. CONCLUSION: Concurrent comorbidities had the greatest prognostic impact among groups with the highest survival rate and the least impact in groups with the lowest survival rate. These findings can be used to help determine the role comorbidity information should play in studies of cancer outcomes. PMID- 15284261 TI - High levels of circulating insulin-like growth factor-I increase prostate cancer risk: a prospective study in a population-based nonscreened cohort. AB - PURPOSE: Insulin-like growth factor-I (IGF-I) stimulates proliferation and inhibits apoptosis in prostate cancer cells, and IGF-I has been associated with increased prostate cancer risk in some, but not all, epidemiologic studies. SUBJECTS AND METHODS: We extended our previous case-control study nested in the Northern Sweden Health and Disease Cohort, a population-based cohort from a region where little prostate specific antigen (PSA) screening is done. Levels of IGF-I and IGF binding protein-3 (IGFBP-3) were measured in prediagnostic blood samples from a total of 281 men who were subsequently diagnosed with prostate cancer after recruitment (median, 5 years after blood collection) and from 560 matched controls. RESULTS: Logistic regression analyses showed increases in prostate cancer risk with increasing plasma peptide levels, up to an odds ratio (OR) for top versus bottom quartile of IGF-I of 1.67 (95% CI, 1.02 to 2.71; Ptrend = .05), which was attenuated after adjustment for IGFBP-3 to an OR of 1.47 (95% CI, 0.81 to 2.64; P (trend) =.32). For men younger than 59 years at recruitment, OR for top versus bottom quartile of IGF-I was 4.12 (95% CI, 1.01 to 16.70; Ptrend = .002), which was significantly stronger than for men older than 59 years (P (interaction) = .006). For men with advanced cancer, OR for top versus bottom quartile of IGF-I was 2.87 (95% CI, 1.01 to 8.12; Ptrend = .10). CONCLUSION: Our data add further support for IGF-I as an etiologic factor in prostate cancer and indicate that circulating IGF-I levels measured at a comparatively young age may be most strongly associated with prostate cancer risk. PMID- 15284262 TI - Phase III study of cisplatin with or without paclitaxel in stage IVB, recurrent, or persistent squamous cell carcinoma of the cervix: a gynecologic oncology group study. AB - PURPOSE: To determine whether cisplatin plus paclitaxel (C+P) improved response rate, progression-free survival (PFS), or survival compared with cisplatin alone in patients with stage IVB, recurrent, or persistent squamous cell carcinoma of the cervix. PATIENTS AND METHODS Eligible: patients with measurable disease, performance status (PS) 0 to 2, and adequate hematologic, hepatic, and renal function received either cisplatin 50 mg/m2 or C+P (cisplatin 50 mg/m2 plus paclitaxel 135 mg/m2) every 3 weeks for six cycles. Tumor measurements and quality-of-life (QOL) assessments were obtained before each treatment cycle. RESULTS: Of 280 patients entered, 6% were ineligible. Among 264 eligible patients, 134 received cisplatin and 130 received C+P. Groups were well matched with respect to age, ethnicity, PS, tumor grade, disease site, and number of cycles received. The majority of all patients had prior radiation therapy (cisplatin, 92%; C+P, 91%). Objective responses occurred in 19% (6% complete plus 13% partial) of patients receiving cisplatin versus 36% (15% complete plus 21% partial) receiving C+P (P = .002). The median PFS was 2.8 and 4.8 months, respectively, for cisplatin versus C+P (P < .001). There was no difference in median survival (8.8 months v 9.7 months). Grade 3 to 4 anemia and neutropenia were more common in the combination arm. There was no significant difference in QOL scores, although a disproportionate number of patients (cisplatin, n = 50; C+P, n = 33) dropped out of the QOL component, presumably because of increasing disease, deteriorating health status, or early death. CONCLUSION C+P is superior to cisplatin alone with respect to response rate and PFS with sustained QOL. PMID- 15284263 TI - Duration of response to second-line, platinum-based chemotherapy for ovarian cancer: implications for patient management and clinical trial design. AB - PURPOSE: Limited information is available regarding the influence of the duration of a prior response on the length of a subsequent response to platinum chemotherapy in recurrent ovarian cancer. PATIENTS AND METHODS: We retrospectively reviewed the medical records of women with ovarian cancer treated at the Cleveland Clinic from 1993 through April 2003 who received two or more platinum-based regimens for recurrence of the malignancy. Patients were considered to have responded to second-line therapy if they satisfied specific criteria, including favorable effects on both measurable or assessable disease. RESULTS: A total of 211 platinum-based regimens were administered to 176 women with recurrent ovarian cancer during this time period, with a response being observed in 125 treatment episodes (59%). Only four (3%) of 121 currently assessable secondary responses were of longer duration than the prior response in a specific individual. In three of these four cases, the platinum-based regimen used in the second-line approach included a drug that had not been used in that patient's primary chemotherapy program. CONCLUSION: The length of a prior response to platinum-based therapy in ovarian cancer is highly predictive of the upper limit of the duration of response to a subsequent platinum program, assuming the same or similar drugs are used. Knowledge of this clinical parameter may assist in developing optimal management for an individual patient and may potentially be exploited in clinical trial designs examining novel maintenance strategies with both cytotoxic and cytostatic agents in women who achieve a secondary response to a platinum-based regimen. PMID- 15284264 TI - Her-2/neu overexpression and amplification in uterine papillary serous carcinoma. AB - PURPOSE: Uterine papillary serous carcinoma (UPSC) is an aggressive subtype of endometrial cancer characterized by early metastasis, resistance to therapy, and a high mortality rate. Little is known about the biology of these tumors. Smaller studies suggest that Her-2/neu may be involved in the tumorigenesis of this disease. The purpose of this study was to evaluate the protein expression and gene amplification of Her-2/neu in UPSC and to determine its prognostic value. PATIENTS AND METHODS: Tumor tissue from 68 patients with UPSC treated at The University of Texas M.D. Anderson Cancer Center from 1989 to 2002 was available. Her-2/neu expression was evaluated by immunohistochemistry (IHC). Overexpression was defined as complete membrane staining in greater than 10% of the cells. In tumors with overexpression of Her-2/neu by IHC, fluorescence in situ hybridization (FISH) was performed to assess gene amplification. Clinical and pathologic information was obtained from medical records. RESULTS: Twelve (18%) of 68 tumors demonstrated Her-2/neu overexpression. Of these, only two showed gene amplification. When evaluating all 68 patients, Her-2/neu overexpression was associated with a poorer overall survival (OS; P = .008). In our multivariate Cox proportional hazards models, Her-2/neu IHC overexpression, lymph node status, and stage were each associated with OS (P < or = .05). CONCLUSION: Positive IHC overexpression of Her-2/neu was seen in 18% of UPSCs but was rarely correlated with Her-2/neu gene amplification. Overexpression of Her-2/neu was associated with a worse overall prognosis. The use of trastuzumab (Herceptin; Genentech, South San Francisco, CA) in women with UPSC should be further evaluated in a clinical trial setting. PMID- 15284265 TI - Temozolomide as initial treatment for adults with low-grade oligodendrogliomas or oligoastrocytomas and correlation with chromosome 1p deletions. AB - PURPOSE: To determine the response rate of low-grade oligodendroglial tumors (LGOT) to temozolomide (TMZ) as initial treatment and to evaluate the predictive value of chromosome 1p deletion on the radiologic response. PATIENTS AND METHODS: Adult patients with pathologically proven LGOT with progressive disease on magnetic resonance imaging (MRI) were eligible for the study. TMZ was administered at the starting dose of 200 mg/m2/d for 5 days, repeated every 28 days. Response was evaluated clinically and by central review of MRIs. Chromosome 1p and 19q deletions were detected by the loss of heterozygosity technique. RESULTS: Sixty consecutive patients were included in the study. At the time of analysis, the median number of TMZ cycles delivered was 11. Clinically, 51% of patients improved, particularly those with uncontrolled epilepsy. The objective radiologic response rate was 31% (17% partial response and 14% minor response), whereas 61% of patients had stable disease and 8% experienced disease progression. The median time to maximum tumor response was 12 months (range, 5 to 20 months). Myelosuppression was the most frequent side effect, with grade 3 to 4 toxicity in 8% of patients. Loss of chromosome 1p was associated with objective tumor response (P < .004). CONCLUSION: TMZ is well tolerated and provides a substantial rate of response in LGOT. Chromosome 1p loss is correlated with radiographic response and could be a helpful marker for guiding therapeutic decision making in LGOT. PMID- 15284266 TI - Cardiovascular status in long-term survivors of Hodgkin's disease treated with chest radiotherapy. AB - PURPOSE: Cardiovascular status was assessed in 48 Hodgkin's disease (HD) survivors at a median of 14.3 years (range, 5.9 to 27.5 years) after diagnosis because they may be at increased risk for cardiovascular abnormalities. PATIENTS AND METHODS: Patients completed the Short-Form 36 quality-of-life instrument and were screened by echocardiography, exercise stress testing, and resting and 24 hour ECG. RESULTS: All patients received mediastinal irradiation (median, 40.0 Gy; range, 27.0 to 51.7 Gy) at a median age of 16.5 years (range, 6.4 to 25.0 years). Four patients received an anthracycline. Although every patient described their health as good or better, and none had symptomatic heart disease at screening, all but one had cardiac abnormalities on screening. Restrictive cardiomyopathy was suggested by reduced average left ventricular (LV) dimension (P < .001) and mass (P < .001), without increased LV wall thickness. Significant valvular defects were present in 42%; 75% had conduction defects. One survivor developed complete heart block shortly after the study visit. Autonomic dysfunction was suggested by a monotonous heart rate in 57%, persistent tachycardia in 31%, and blunted hemodynamic responses to exercise in 27%. Peak oxygen uptake (VO2max) during exercise, a predictor of mortality in heart failure, was significantly reduced (< 20 mL/kg/m2) in 30% of survivors. VO2max was correlated with increasing fatigue, increasing shortness of breath (both, r = -0.35; P =. 02), and decreasing physical component score on the SF-36 (r = 0.554; P = .00017). CONCLUSION: A variety of unsuspected, clinically significant cardiovascular abnormalities are common in long-term survivors of HD who are treated at a young age with mediastinal irradiation. We recommend serial, comprehensive cardiac screening of HD survivors who fit this profile. PMID- 15284267 TI - Using health-related quality of life measures to predict cardiac function in survivors exposed to anthracyclines. AB - PURPOSE: As the number of pediatric cancer survivors increases, so does the number of survivors previously exposed to anthracyclines as part of their cancer therapy. Because screening is costly, some have suggested that health-related quality of life (HRQL) measures might be useful in focusing screening tests on those patients with cases most likely to display positive findings. This study reports on the predictive ability of HRQL measures to detect patients with abnormalities on serial cardiac testing. METHODS: Using 127 patients from the ACE Inhibitor after Anthracycline (AAA) Trial, this study compared serial measures of the Short Form-36 (SF-36; for ages > 13 years) and Child Health Questionnaire Child Form 87 (CHQ-CF87; for ages < or = 13 years) to serial cardiac performance tests including echocardiographic shortening fraction, left ventricular end systolic wall stress (LVESWS), LVESWS-index, and maximal cardiac index (MCI; a measure of cardiac output at peak exercise). RESULTS: Generally, there was no clinically or statistically significant correlation between any HRQL measure and any cardiac function measure except between MCI and vitality and physical functioning. For each of these measures, the correlation between MCI was statistically significant (P < .006), but each HRQL subscale could explain no more than 7% of the variation in MCI. HRQL measures were not predictive of any other cardiac function measure. CONCLUSION: HRQL measures should not be used in isolation as a screen for cardiac function abnormalities in patients exposed to anthracylines who already have a mild degree of ventricular dysfunction. Patient history appears to be no substitute for cardiac testing in this cohort. PMID- 15284268 TI - Preliminary results from a phase II trial of conformal radiation therapy and evaluation of radiation-related CNS effects for pediatric patients with localized ependymoma. AB - PURPOSE: We conducted a phase II trial of conformal radiation therapy (CRT) for localized childhood ependymoma to determine whether the irradiated volume could be reduced to decrease CNS-related side effects without diminishing the rate of disease control. PATIENTS AND METHODS: Between July 1997 and January 2003, 88 pediatric patients (median age, 2.85 +/- 4.5 years) received CRT in which doses (59.4 Gy to 73 patients or 54.0 Gy after gross-total resection to 15 patients younger than 18 months) were administered to the gross tumor volume and a margin of 10 mm. Patients were categorized according to extent of resection (underwent gross total resection, n = 74; near-total resection, n = 6; subtotal resection, n = 8), prior chemotherapy (n = 16), tumor grade (anaplastic, n = 35), and tumor location (infratentorial, n = 68). An age-appropriate neurocognitive battery was administered before and serially after CRT. RESULTS: The median length of follow up was 38.2 months (+/- 16.4 months); the 3-year progression-free survival estimate was 74.7% +/- 5.7%. Local failure occurred in eight patients, distant failure in eight patients, and both in four patients. The cumulative incidence of local failure as a component of failure at 3 years was 14.8% +/- 4.0%. Mean scores on all neurocognitive outcomes were stable and within normal limits, with more than half the cohort tested at or beyond 24 months. CONCLUSION: Limited volume irradiation achieves high rates of disease control in pediatric patients with ependymoma and results in stable neurocognitive outcomes. PMID- 15284269 TI - Long-term silicone central venous catheters impregnated with minocycline and rifampin decrease rates of catheter-related bloodstream infection in cancer patients: a prospective randomized clinical trial. AB - PURPOSE: To evaluate the efficacy of long-term nontunneled silicone catheters impregnated with minocycline and rifampin (M-R) in reducing catheter-related bloodstream infections. PATIENTS AND METHODS: This prospective, randomized, double-blind clinical trial was conducted at M.D. Anderson Cancer Center, a tertiary care hospital in Houston, TX. All patients in the trial had a malignancy. RESULTS: Between September 1999 and May 2002, 356 assessable catheters were used: 182 M-R and 174 nonimpregnated. The patients' characteristics were comparable between the two study groups. The mean (+/- standard deviation) duration of catheterization with M-R catheters was comparable to that of nonimpregnated catheters (66.21 +/- 30.88 v 63.01 +/- 30.80 days). A total of 17 catheter-related bloodstream infections occurred during the course of the study. Three were associated with the use of M-R catheters and 14 were associated with the nonimpregnated catheters, with a rate of catheter-related bloodstream infection of 0.25 and 1.28/1,000 catheter-days, respectively (P = .003). Gram-positive cocci accounted for the majority of the organisms causing the infections. There were no allergic reactions associated with M-R catheters. CONCLUSION: Long-term nontunneled central venous catheters impregnated with minocycline and rifampin are efficacious and safe in reducing catheter-related bloodstream infections in cancer patients. PMID- 15284270 TI - Assessing quality of life during chemotherapy for pleural mesothelioma: feasibility, validity, and results of using the European Organization for Research and Treatment of Cancer Core Quality of Life Questionnaire and Lung Cancer Module. AB - PURPOSE: To assess the feasibility and validity of using the European Organization for Research and Treatment of Cancer (EORTC) Core Quality of Life Questionnaire (QLQ-C30) and Lung Cancer Module (QLQ-LC13) to describe health related quality of life (HRQL) in patients with pleural mesothelioma undergoing combination chemotherapy, to identify the most impaired aspects of HRQL, and to assess the impact of chemotherapy on HRQL. PATIENTS AND METHODS: Fifty-three patients received cisplatin on day 1 and gemcitabine on days 1, 8, and 15 of a 28 day cycle for a maximum of six cycles. HRQL was assessed using the EORTC QLQ-C30 and QLQ-LC13. RESULTS: Compliance was 100% at baseline but subsequently decreased. At baseline, role function and social function were the most impaired domains, and the worst-rated symptoms were fatigue, dyspnea, pain, insomnia, appetite loss, and cough. Dyspnea, pain, insomnia, and cough improved with chemotherapy, although functional domains and chemotherapy-related symptoms deteriorated. Fatigue remained unchanged. Few patients reported hemoptysis. Functional domains and symptoms scales from the QLQ-C30 demonstrated predictive validity for survival. The predictive value of QLQ-LC13 pain scores was improved by combining three pain items into a single score. Dyspnea scores were correlated strongly with lung function as measured by forced vital capacity. CONCLUSION: This study supports the validity of the EORTC QLQ-C30 and LC13 as outcome measures for trials of chemotherapy in mesothelioma. Although the most prominent symptoms reported were concordant with clinical experience, impairments in role and social function and insomnia were worse than expected. Future research should focus on how best to apply, analyze, and interpret existing, validated HRQL instruments in mesothelioma research and practice, not on the development of new ones. PMID- 15284271 TI - Review of determinants of patients' preferences for adjuvant therapy in cancer. AB - PURPOSE: Many studies have determined cancer patients' preferences for adjuvant therapy, for example, by asking patients the extent of benefit they would need in order to accept the therapy. However, little is known about the determinants that influence these preferences. Our research goal was to explore which determinants underlie patients' preferences by means of a literature review. METHODS: PubMed searches were conducted to identify studies in which cancer patients' preferences for adjuvant therapy had been elicited by means of a treatment preference instrument. Twenty-three papers were evaluated with regard to reported relationships between preferences and potential determinants. A total of 40 determinants were recorded and classified into one of seven categories: (1) treatment-related determinants, (2) sociodemographic characteristics and current quality of life, (3) clinical characteristics, (4) measurement instrument-related determinants, (5) time-related determinants, (6) cognitive/affective determinants, and (7) specialist-related determinants. RESULTS: The benefit and toxicity of treatment, experience of the treatment, and having dependents (eg, children) living at home were important determinants of patients' preferences. Furthermore, qualitative data suggested that cognitive/affective and specialist related determinants might have a large impact on patients' treatment preferences. CONCLUSION: Our results show that patients' preferences cannot fully be explained on the basis of treatment-related determinants and patient and clinical characteristics. More research is needed in the area of cognitive/affective and specialist-related determinants because of the lack of quantitative results. Furthermore, we recommend carrying out larger studies in which the (internal) relationships between determinants and preferences are assessed in the context of a cognitive cost-benefit model. PMID- 15284272 TI - Uncommon manifestations of common malignancies: case 1. Soft-tissue metastases from malignant pleural mesothelioma. PMID- 15284273 TI - Uncommon manifestations of common malignancies: case 2. Chronic lymphocytic leukemia and multiple myeloma in the same patient. PMID- 15284274 TI - Uncommon manifestations of common malignancies: case 3. Malignant melanoma arising from a spinal nerve root. PMID- 15284275 TI - Reassurance. PMID- 15284276 TI - Phase III study of letrozole versus tamoxifen as first-line therapy of advanced breast cancer in postmenopausal women: analysis of survival and update of efficacy from the international letrozole breast cancer group. PMID- 15284278 TI - Free circulating DNA: good as a diagnostic marker in lung cancer? PMID- 15284280 TI - Oxaliplatin toxicity masquerading as recurrent colon cancer. PMID- 15284282 TI - Overview: mechanisms of hypertension: cells, hormones, and the kidney. PMID- 15284281 TI - A cyclooxygenase metabolite of anandamide causes inhibition of interleukin-2 secretion in murine splenocytes. AB - Arachidonyl ethanolamine, which is commonly known as anandamide, was the first endogenous compound to be identified that binds to the cannabinoid receptors. Anandamide mimics many of the physiological effects of Delta(9) tetrahydrocannabinol (Delta(9)-THC), including hypothermia, antinociception, immobility, catalepsy, and immune modulation. In the present studies, we show that anandamide caused a concentration-dependent inhibition of interleukin-2 in primary splenocytes. The CB1 and CB2 antagonists, SR141716A [N-(piperidin-1-yl)-5 (4-chlorophenyl)-1-(2,4-dichlorphenyl)-4-methyl-H-pyrazole-3 carboxyamidehydrochloride] and SR144528 [N-[(1S)-endo-1,3,3, trimethylbicyclo[2,2,1]heptan-2-yl]-5-(4-chloro-3-methylphenyl)-1-(4 methylbenzyl)-pyrazole-3-carboxamide], when used in combination, did not antagonize the inhibition of interleukin-2 by anandamide. Additionally, neither UCM707 [N-(3-furanylmethyl)-5Z,8Z,11Z,14Z-eicosatetraenamide], the inhibitor of the putative anandamide membrane transporter (AMT), nor methyl arachidonoyl fluorophosphonate (MAFP), the inhibitor of fatty acid amidohydrolase (FAAH), were able to affect the inhibitory activity of anandamide upon interleukin-2. Interestingly, arachidonic acid caused a concentration-dependent inhibition of interleukin-2 secretion (IC(50) = 10.3 microM), which was similar to that of structurally related anandamide (IC(50) = 11.4 microM). The inhibition of interleukin-2 by anandamide and arachidonic acid was partially reversed by pretreatment with the nonspecific cyclooxygenase inhibitors, flurbiprofen and piroxicam. Moreover, NS398 [N-[2-(cyclohexyloxy)-4-nitrophenyl] methanesulfonamide], a cyclooxygenase-2-specific inhibitor, also attenuated the inhibitory effects of anandamide and arachidonic acid upon interleukin-2 secretion. Finally, pretreatment with a peroxisome proliferator-activated receptor gamma (PPARgamma)-specific antagonist, T0070907 [2-chloro-5-nitro-N-4 pyridinyl-benzamide], partially antagonized anandamide-mediated suppression of IL 2 secretion. Collectively, the aforementioned studies suggest that inhibition of interleukin-2 secretion by anandamide is independent of CB1/CB2 and the AMT/FAAH system. Additionally, these studies also suggest that inhibition of interleukin-2 is mediated by a PPARgamma, which is activated by a cyclooxygenase-2 metabolite of anandamide. PMID- 15284283 TI - Ischemic nephropathy: where are we now? AB - Identification and reversing the loss of kidney function beyond occlusive disease of the renal arteries poses a major clinical challenge. Recent studies indicate that atherosclerotic renal artery stenosis develops as a function of age and is commonly associated with other microvascular disease, including nephrosclerosis and diabetic nephropathy. The risks of renal artery stenosis are related both to declining kidney function and to accelerated cardiovascular disease, with increased morbidity and mortality. Newer drugs, including agents that block the renin-angiotensin system, have improved the level of BP control for renovascular hypertension. Progressive renovascular disease during medical therapy can produce refractory hypertension, congestive heart failure, and renal failure with tubulointerstitial fibrosis. Recent studies indicate a complex interplay of oxidative stress, endothelial dysfunction, and activation of fibrogenic cytokines as a result of experimental atherosclerosis and renal hypoperfusion. Advances in imaging and interventional devices offer major new opportunities to prevent progressive loss of kidney function. Recent series indicate that although 25 to 30% of patients with impaired renal function can recover glomerular filtration after revascularization, many have no apparent change in kidney function and 19 to 25% experience a significant loss of kidney function, in some cases as a result of atheroemboli. To select patients who are most likely to benefit from vascular intervention, clinicians should understand the pathophysiology of developing ischemic nephropathy and the potential hazards of revascularization in the setting of diffuse atherosclerotic disease. Further research should be directed toward identification of critical disease, regulation of fibrogenesis, and the interaction with other atherosclerotic processes. PMID- 15284284 TI - Endothelial dysfunction. AB - Endothelial dysfunction is characterized by a shift of the actions of the endothelium toward reduced vasodilation, a proinflammatory state, and prothrombic properties. It is associated with most forms of cardiovascular disease, such as hypertension, coronary artery disease, chronic heart failure, peripheral artery disease, diabetes, and chronic renal failure. Mechanisms that participate in the reduced vasodilatory responses in endothelial dysfunction include reduced nitric oxide generation, oxidative excess, and reduced production of hyperpolarizing factor. Upregulation of adhesion molecules, generation of chemokines such as macrophage chemoattractant peptide-1, and production of plasminogen activator inhibitor-1 participate in the inflammatory response and contribute to a prothrombic state. Vasoactive peptides such as angiotensin II and endothelin-1; the accumulation of asymmetric dimethylarginine, an endogenous nitric oxide inhibitor; hypercholesterolemia; hyperhomocysteinemia; altered insulin signaling; and hyperglycemia can contribute to these different mechanisms. Detachment and apoptosis of endothelial cells (anoikis) are associated phenomena. Endothelial dysfunction is an important early event in the pathogenesis of atherosclerosis, contributing to plaque initiation and progression. Reductions in circulating endothelial progenitor cells that participate in regeneration of the endothelium participate in endothelial pathophysiology. The severity of endothelial dysfunction has been shown to have prognostic value for cardiovascular events. Correction of endothelial dysfunction may be associated with reduced cardiovascular risk. Circulating endothelial progenitor cells may represent a potential therapeutic approach for endothelial dysfunction. PMID- 15284285 TI - Mechanisms of hypertension: the expanding role of aldosterone. AB - Hypertension is a common disorder that affects a large heterogeneous patient population. Subgroups can be identified on the basis of their responses to hormonal and biologic stimuli. These subgroups include low-renin hypertensives and nonmodulators. Aldosterone, the principal human mineralocorticoid, is increasingly recognized as playing a significant role in cardiovascular morbidity, and its role in hypertension has recently been reevaluated with studies that suggest that increased aldosterone biosynthesis (as defined by an elevated aldosterone to renin ratio) is a key phenotype in up to 15% of individuals with hypertension. It was reported previously that a polymorphism of the gene (C to T conversion at position -344) encoding aldosterone synthase is associated with hypertension, particularly in individuals with a high ratio. However, the most consistent association with this variant is a relative impairment of adrenal 11beta-hydroxylation. This review explores the evidence for this and provides a hypothesis linking impaired 11beta-hydroxylation and hypertension with a raised aldosterone to renin ratio. It is also speculated that there is substantial overlap between this group of patients and previously identified low-renin hypertensives and nonmodulators. Thus, these groups may form a neurohormonal spectrum reflecting different stages of hypertension or indeed form sequential steps in the natural history of hypertension in genetically susceptible individuals. PMID- 15284286 TI - Differential regulation of basolateral Cl-/HCO3- exchangers SLC26A7 and AE1 in kidney outer medullary collecting duct. AB - SLC26A7 is a recently identified Cl(-)/HCO(3)(-) exchanger that co-localizes with AE1 on the basolateral membrane of Alpha intercalated cells (A-IC) in outer medullary collecting duct (OMCD). The purpose of these studies was to determine whether AE1 and SLC26A7 are differentially regulated in OMCD in pathophysiologic states. Toward this end, the expression and regulation of AE1 and SLC26A7 was examined in water deprivation, a condition known to increase the osmolality of the medulla. Rats were subjected to 3 d of water deprivation while having free access to food. Northern hybridizations demonstrated that in the outer medulla, the mRNA expression of SLC26A7 increased by approximately 300% (P < 0.01 versus control; n = 3), whereas the expression of AE1 decreased by approximately 50% (P < 0.05 versus control, n = 3) in water-deprived rats. Immunoblot analysis studies demonstrated that in the outer medulla, SLC26A7 abundance increased by approximately 3.5-fold (P < 0.02 versus control; n = 3), whereas the AE1 abundance decreased by approximately 55% (P < 0.05 versus control) in water deprivation. The expression of SLC26A7 remained unchanged in the kidney cortex and stomach in water deprivation, indicating the specificity of SLC26A7 upregulation in outer medulla. In situ hybridization indicated the exclusive expression of SLC26A7 in the outer medulla and double immunofluorescence labeling confirmed the co-localization of AE1 and SLC26A7 on the basolateral membrane of A IC cells in OMCD. It is concluded that AE1 and SLC26A7 are differentially regulated in OMCD in water deprivation. On the basis of these results and previous functional studies indicating the activation of SLC26A7 activity by high osmolality, it is proposed that SLC26A7 may play an important role in bicarbonate reabsorption and or cell volume regulation in OMCD (specifically under hypertonic conditions). PMID- 15284287 TI - The renal-specific transporter mediates facilitative transport of organic anions at the brush border membrane of mouse renal tubules. AB - The renal secretion of organic anions across the proximal tubules is achieved by a coordination of uptake and efflux transporters. This study reports the expression, localization, and functional properties of mouse renal-specific transporter (RST). Mouse RST mRNA is predominantly expressed in the kidney and localized on the brush border membrane of mouse kidney proximal tubules. Mouse RST-expressing HEK293 cells exhibited saturable uptake of p-aminohippurate (Km approximately 234 microM), which was increased by an increase in K(+) concentration or in the presence of Ba(2+) and ouabain and decreased by diethylpyrocarbonate, a histidine modifier. An increase in K(+) concentration enhanced the uptake of benzylpenicillin, 2,4-dichlorophenoxyacetate, and dehydroepiandrosterone sulfate, suggesting polyspecific substrate specificity of mouse RST. Vectorial transport of 2,4-dichlorophenoxyacetate was observed in the basal-to-apical direction in rat organic anion transporter 3-expressing LLC-PK1 cells (rOat3-LLC); however, coexpression of mouse RST in rOat3-LLC caused a 1.3 fold increase in the basal-to-apical transport. In addition, the basal-to-apical transport of benzylpenicillin and urate was 3- and 2.5-fold greater than that in the opposite direction in the double-transfected cells, respectively, whereas their transepithelial transport in vector- or rOat3-LLC was symmetrical. Furthermore, the basal-to-apical transport of benzylpenicillin was saturable and reduced by increasing extracellular K(+) concentration and ouabain. These results suggest that mouse RST mediates the efflux of organic anions including urate and works as exit for organic anions in the proximal tubules. In addition to the kidney, mouse RST was detected in the brain capillaries and the choroid plexus, and it may also play a role in efflux transport of organic anions across the barriers of the central nervous system. PMID- 15284288 TI - Homocysteine and the renal epithelial transport and toxicity of inorganic mercury: role of basolateral transporter organic anion transporter 1. AB - The epithelial cells that line the renal proximal tubule have been shown to be the primary cellular targets where mercuric ions gain entry, accumulate, and induce pathologic effects in vivo. Recent data have implicated at least one of the organic anion transport systems in the basolateral uptake of inorganic mercury (Hg). With the use of a line of type II MDCK cells transfected stably with the human organic anion transporter 1 (hOAT1), the hypothesis that hOAT1 can transport mercuric conjugates of homocysteine (Hcy) was tested. Indeed, MDCK II cells expressing a functional form of hOAT1 gained the ability to transport the mercuric conjugate 2-amino-4-(3-amino-3-carboxy-propylsulfanylmercuricsulfanyl) butyric acid (Hcy-S-Hg-S-Hcy). In addition, p-aminohippurate and the dicarboxylates adipate and glutarate (but not succinate or malonate) inhibited individually the uptake of Hcy-S-Hg-S-Hcy in a concentration-dependent manner. Furthermore, a direct relationship between the uptake of Hcy-S-Hg-S-Hcy and the induction of cellular injury and death was demonstrated in the hOAT1-expressing MDCK II cells only. These data represent the first line of direct evidence implicating one of the organic anion transporters in the uptake of a mercuric conjugate of Hcy in a mammalian cell. Thus, mercuric conjugates of Hcy are potential transportable substrates of OAT1. More important, the findings from the present study implicate the activity of OAT1 in the uptake and toxicity of Hg (when in the form of Hcy-S-Hg-S-Hcy in the extracellular compartment) in proximal tubular epithelial cells in vivo. PMID- 15284289 TI - High ambient glucose enhances sensitivity to TGF-beta1 via extracellular signal- regulated kinase and protein kinase Cdelta activities in human mesangial cells. AB - High ambient glucose activates intracellular signaling pathways to induce cytokines such as TGF-beta1 in the extracellular matrix accumulation of diabetic nephropathy. These same pathways also may directly modulate TGF-beta1 signaling. R-Smad phosphorylation, association with Smad4, and nuclear accumulation after TGF-beta1 treatment (1.0 ng/ml) were significantly higher in mesangial cells that were conditioned to 20 mM glucose for 72 h than mesangial cells in 6.5 mM glucose, suggesting that high glucose enhanced responsiveness to TGF-beta1. Neither TGF-beta1 bioactivity nor TGF-beta receptor binding was significantly different between in 6.5 and 20 mM glucose-conditioned cultures. Furthermore, adding a neutralizing anti-TGF-beta1 antibody during glucose conditioning did not affect the enhanced Smad responsiveness, indicating that enhancement likely did not result from increased TGF-beta expression. In contrast, a mitogen-activated protein (MAP) kinase/extracellular signal-regulated kinase (ERK) kinase (MEK)/ERK inhibitor, PD98059, completely abrogated the effect of high glucose. Glucose stimulation of ERK was inhibited by the general protein kinase C (PKC) inhibitor calphostin C and by the PKCdelta-specific inhibitor rottlerin, whereas Go6976, an inhibitor of conventional PKC, had no effect on ERK activity. Specificity of the PKC inhibitors was further verified by PKCbeta and delta kinase assay. High glucose increased expression of several PKC isozymes, but only PKCdelta showed proportionally increased membrane translocation and kinase activity in cells that were conditioned to 20 mM glucose. Finally, both ERK and PKCdelta inhibition during glucose conditioning abrogated enhanced alpha1(I) collagen mRNA and promoter induction by TGF-beta1. Taken together, these data strongly suggest that heightened ERK and PKCdelta activity in high ambient glucose conditions interact with the Smad pathway, leading to enhanced responsiveness to TGF-beta1 and increased extracellular matrix production in mesangial cells. PMID- 15284290 TI - Siah-1 interacts with the intracellular region of polycystin-1 and affects its stability via the ubiquitin-proteasome pathway. AB - Autosomal dominant polycystic kidney disease, characterized by extensive formation of renal cysts and progressive renal failure, is a genetic disorder caused by mutations in the PKD1 and PKD2 genes. The PKD1 gene product, polycystin 1, is a transmembrane protein with its N-terminus facing the extracellular region and C-terminus facing the cytoplasm. Polycystin-1 seems to be involved in regulating cell growth and maturation, but the precise mechanisms are not yet well defined. For investigating the function of the intracellular region of polycystin-1, the C-terminal cytoplasmic fragment of polycystin-1, PKD1-C, was used as bait in two-hybrid screening, and a polycystin-1-binding protein, the human homologue of Drosophila Seven in Absentia (Siah-1), which has a RING domain and promotes the ubiquitin-dependent proteasome pathway, was identified. It was shown that PKD1-C interacts with Siah-1 in vivo. In addition, interaction with Siah-1 induces the degradation of PKD1-C, shortening its half-life. PKD1-C and CD4 chimeric proteins, which are attached to the plasma membrane, also show similar results. Furthermore, ubiquitination and degradation of PKD1-C are increased in the presence of Siah-1, and overexpression of Siah-1 protein promotes the degradation of polycystin-1 via the ubiquitin-proteasome pathway. These results suggest that polycystin-1 is regulated by Siah-1 through the ubiquitin-dependent proteasome pathway. PMID- 15284291 TI - Specific Cre/Lox recombination in the mouse proximal tubule. AB - The present work reports for the first time the construction of a transgenic mouse strain with specific expression of Cre recombinase in the kidney proximal tubule. A Cre/loxP strategy was developed using sglt2 promoter to drive Cre recombinase expression in transgenic mice. The mouse sglt2 5' region consisting of the first exon, the first intron, and part of the second exon was cloned upstream of a nucleotide sequence encoding the Cre recombinase. Transgenic mice were generated by pronuclear injection, and tissue specificity of Cre expression was analyzed using reverse transcription-PCR. The iL1-sglt2-Cre mouse line scored positive for kidney transcription of Cre but not for the other tissues analyzed. Within the kidney, Cre transcripts were demonstrated to be restricted to the proximal tubule only. iL1-sglt2-Cre mice were bred with ROSA26-LacZ reporter mice that contained a loxP-flanked stop sequence upstream of the LacZ gene. X-gal staining and immunohistochemistry using specific antibodies (anti-megalin, anti Tamm-Horsfall, anti-NaCl co-transporter, and anti-aquaporin 2) revealed that sglt2 drives Cre functional expression specifically in proximal tubules. The iL1 sglt2-Cre mouse therefore represents a powerful tool for Cre-LoxP-mediated conditional expression in the renal proximal tubule. PMID- 15284292 TI - An acidic peptide sequence of nucleolin-related protein can mediate the attachment of calcium oxalate to renal tubule cells. AB - Crystals that form in tubular fluid must be retained in the kidney to become stones. Nucleolin-related protein (NRP) is found on the surface of inner medullary collecting duct (IMCD) cells in culture (cIMCD) and selectively adsorbs to calcium oxalate (CaOx). We proposed that NRP mediates attachment to the renal tubular epithelium of Ca stone crystals through an electrostatic interaction with a highly acidic region (acidic fragment [AF]) similar to those of other proteins that have been reported to affect urinary crystal formation. The current studies demonstrate that nucleolin is expressed on both apical and basolateral cell surfaces of cIMCD, reaching a peak in the late stages of mitosis and gradually declining to undetectable levels with maturation of the polarized epithelium. Scraping areas of mature monolayers stimulated the cells surrounding the defects to migrate and proliferate so as to repair them, and these areas demonstrate surface NRP expression and enhanced attachment of CaOx monohydrate crystals. Surface expression of the NRP AF was produced by cloning the NRP AF into a display vector. Transfected cIMCD demonstrating copious surface expression of AF enhanced CaOx attachment 6.7-fold compared with control cIMCD, whereas cells transfected with a vector without the AF did not differ from control. AF was also cloned into a replication-deficient adenovirus and expressed in 293 cells, resulting in AF secretion into the nutrient medium. This medium inhibited CaOx attachment to cIMCD, compared with conditioned medium from cells infected with wild-type virus. These results demonstrate that surface-bound AF can mediate CaOx attachment and that secreted AF can inhibit attachment. These results support the notion that surface-associated NRP could mediate attachment of CaOx to the renal tubule epithelium, thereby causing retention of crystals that might eventually become kidney stones. PMID- 15284293 TI - Coupled induction of iNOS and p53 upregulation in renal resident cells may be linked with apoptotic activity in the pathogenesis of progressive IgA nephropathy. AB - In situ hybridization, immunohistochemistry, and TUNEL staining were applied to renal biopsy specimens of immunoglobulin A nephropathy (IgAN) patients to determine the expression of nitric oxide synthase (iNOS) (mRNA and protein), p53, and their potential roles in renal cell apoptosis in relation to the development of pathologic lesions. Fifty-one cases were categorized into four subgroups (A-D) according to the presence of progressive histopathological features. A cell type specific and differential overexpression of iNOS mRNA and protein was demonstrated in glomerular cells in subgroups (A-C) and was found to correlate well with the upregulation of p53 protein by glomerular endothelium and epithelium in early- and advanced-stage disease. In the tubulointerstitium, induction of iNOS products was evident in damaged tubules in late-stage disease, in parallel with the upregulation of p53 protein levels in these tubules. Increased TUNEL staining observed in glomeruli with progressive lesions and tubules with degenerative changes positively correlated with the expression levels of iNOS and p53 in glomerular endothelium, epithelium, and their overexpression in damaged tubules. Clinicopathologic correlations demonstrated that induction of iNOS products in renal cells was associated with indices of poor renal prognosis in human IgAN. The coupled induction of iNOS and p53 upregulation in intrinsic renal cells of IgAN may be linked with both pro- and anti-apoptotic activities, thus playing an important role in mediating progressive renal injury and determining renal outcome in human IgAN. PMID- 15284294 TI - Podocyte flattening and disorder of glomerular basement membrane are associated with splitting of dystroglycan-matrix interaction. AB - The transmembrane component of the dystroglycan complex, a heterodimer of alpha- and beta-dystroglycan, was recently localized at the basal cell membrane domain of podocytes, and it was speculated that it serves as a device of the podocyte for maintaining the complex podocyte foot process architecture, and for regulating the exact position of its ligands, the matrix proteins laminin and agrin, in the glomerular basement membrane (GBM). The redistribution of dystroglycan in two experimental rat models of foot process flattening and proteinuria-i.e., podocyte damage induced by polycationic protamine sulfate perfusion, and reactive oxygen species (ROS)-associated puromycin aminonucleoside nephrosis-was examined. In both experimental diseases, aggregation and reduced density of alpha-dystroglycan by endocytosis by podocytes was observed. In in vitro solid-phase binding assays, protamine and ROS competed with the binding of alpha-dystroglycan with purified laminin and a recombinant C-terminal fragment of agrin that contains the dystroglycan-binding domain. These changes were associated with disorder of the fibrillar components of the lamina rara externa of the GBM, as confirmed quantitatively by fractal analysis. These results indicate that both polycation and ROS induce similar changes in the distribution of podocyte alpha-dystroglycan that involve competitive disruption of alpha dystroglycan/matrix protein complexes, endocytosis of the liberated receptor by podocytes, and disorganization of the matrix protein arrangement in the lamina rara externa. This links functional damage of the dystroglycan complex with structural changes in the GBM. PMID- 15284295 TI - Mitogenic signaling of urokinase receptor-deficient kidney fibroblasts: actions of an alternative urokinase receptor and LDL receptor-related protein. AB - The urokinase receptor (uPAR) attenuates myofibroblast recruitment and fibrosis in the kidney. This study examined the role of uPAR and its co-receptor LDL receptor-related protein (LRP) in the regulation of kidney fibroblast proliferation and extracellular signal-regulated kinase (ERK) signaling. Compared with uPAR+/+ cells, uPAR-/- kidney fibroblasts were hyperproliferative. UPAR-/- fibroblast proliferation was 60% inhibited by an ERK kinase inhibitor. LRP protein was reduced and extracellular accumulation of urokinase-type plasminogen activator (uPA) and plasminogen activator inhibitor type 1 (PAI-1) proteins were greater in uPAR-/- cultures. Addition of functional uPA protein or LRP antisense RNA significantly increased ERK signaling and cell mitosis in both genotypes. Enhanced uPAR-/- fibroblast proliferation was reversed by a recombinant nonfunctional uPA peptide. The density of cell-bound fluor-uPA was similar between uPAR-/- and uPAR+/+ fibroblasts (78 +/- 6 versus 92 +/- 16 units). These data suggest that uPAR-deficient kidney fibroblasts express lower levels of its scavenger co-receptor LRP, resulting in greater extracellular accumulation of uPA and PAI-1. Enhanced proliferation of uPAR-/- fibroblasts seems to be mediated by uPA-dependent ERK signaling via an alternative urokinase receptor. PMID- 15284296 TI - Circumvention of normal constraints on granule protein gene expression in peripheral blood neutrophils and monocytes of patients with antineutrophil cytoplasmic autoantibody-associated glomerulonephritis. AB - Granulopoiesis-related genes are distinctively upregulated in peripheral leukocytes of patients with antineutrophil cytoplasmic autoantibodies (ANCA) associated glomerulonephritis. Affymetrix microarrays identified the upregulation of nine neutrophilic primary granule genes, including myeloperoxidase (MPO) and proteinase 3 (PR3), plus five secondary granule genes. Coordinate expression of granulocyte maturation marker CD35, measured by TaqMan PCR, and positive in situ staining for PR3 transcripts in polymorphic neutrophils and monocytes indicate that these genes are expressed in "mature" cells. Increased transcripts correlated with disease activity and absolute neutrophil values but not with "left shift," drug regimen, cytokine levels, hematuria, proteinuria, ANCA titer, serum creatinine, gender, or age. Upregulation of PR3 and MPO transcripts was specifically associated with ANCA disease (n = 56) as these changes were not detected in patients with ESRD (n = 25) or systemic lupus erythematosus (n = 17), as determined by TaqMan PCR. This is the first report of this phenomenon in nonneoplastic cells. The data raise the hypothesis that, in addition to the presence of anti-MPO or anti-PR3 autoantibodies, a second critical component in the cause of this disease is the reactivation of once-silenced genes leading to increased antigen availability. PMID- 15284297 TI - Erythropoietin protects the kidney against the injury and dysfunction caused by ischemia-reperfusion. AB - Erythropoietin (EPO) is upregulated by hypoxia and causes proliferation and differentiation of erythroid progenitors in the bone marrow through inhibition of apoptosis. EPO receptors are expressed in many tissues, including the kidney. Here it is shown that a single systemic administration of EPO either preischemia or just before reperfusion prevents ischemia-reperfusion injury in the rat kidney. Specifically, EPO (300 U/kg) reduced glomerular dysfunction and tubular injury (biochemical and histologic assessment) and prevented caspase-3, -8, and 9 activation in vivo and reduced apoptotic cell death. In human (HK-2) proximal tubule epithelial cells, EPO attenuated cell death in response to oxidative stress and serum starvation. EPO reduced DNA fragmentation and prevented caspase 3 activation, with upregulation of Bcl-X(L) and XIAP. The antiapoptotic effects of EPO were dependent on JAK2 signaling and the phosphorylation of Akt by phosphatidylinositol 3-kinase. These findings may have major implications in the treatment of acute renal tubular damage. PMID- 15284298 TI - Accelerated nephropathy in diabetic apolipoprotein e-knockout mouse: role of advanced glycation end products. AB - Hyperlipidemia not only may be relevant to cardiovascular disease in diabetes but may also play a role in the development and progression of diabetic nephropathy. Furthermore, there is increasing evidence that advanced glycation end products (AGE) play an important role in diabetic renal disease. The objectives of this study were first to characterize renal injury in diabetic apolipoprotein E knockout (apo E-KO) mice and second to explore the role of AGE in the development and progression of renal disease in this model. Diabetes was induced by injection of streptozotocin in 6-wk-old apo E-KO mice. Diabetic animals received no treatment or treatment with the inhibitor of AGE formation aminoguanidine (1 g/kg per d) or the cross-link breaker [4,5-dimethyl-3-(2-oxo2-phenylethyl)-thiazolium chloride] ALT-711, which cleaves preformed AGE (20 mg/kg per d) for 20 wk. Nondiabetic apo E-KO mice as well as nondiabetic and diabetic C57BL/6 mice served as controls. Compared with nondiabetic apo E-KO mice, induction of diabetes in apo E-KO mice resulted in accelerated renal injury characterized by albuminuria and glomerular and tubulointerstitial injury. These abnormalities were associated with increased expression of collagen type I and type IV and transforming growth factor-beta1 (TGF-beta1), increased alpha-smooth muscle actin immunostaining and macrophage infiltration, and increased serum and renal AGE. The two treatments, which attenuated renal AGE accumulation in a disparate manner, were associated with less albuminuria, structural injury, macrophage infiltration, TGF-beta1, and collagen expression. The accelerated renal injury that was observed in diabetic apo E-KO mice was attenuated by approaches that inhibit renal AGE accumulation. PMID- 15284299 TI - Interactions between angiotensin II and NF-kappaB-dependent pathways in modulating macrophage infiltration in experimental diabetic nephropathy. AB - NF-kappaB-dependent pathways play an important role in macrophage infiltration and kidney injury. NF-kappaB is regulated by angiotensin II (AII). However, the role of this pathway in diabetic nephropathy has not been clearly delineated. First, the activation of NF-kappaB, monocyte chemoattractant protein-1 (MCP-1), and macrophage infiltration in the diabetic kidney were explored, in a temporal manner. The active subunit of NF-kappaB, p65, was elevated in the diabetic animals in association with increased MCP-1 gene expression and macrophage infiltration. Second, the effects of treatment for 4 wk with the AII type 1 receptor antagonist valsartan, the AII type 2 receptor antagonist PD123319, or pyrrolidine dithiocarbamate, an inhibitor of NF-kappaB and on these parameters were assessed. These treatments were associated with a reduction in p65 activation, MCP-1 gene expression, and macrophage infiltration. These findings demonstrate a role for activation of NF-kappaB, in particular the p65 subunit, in the pathogenesis of early renal macrophage infiltration in experimental diabetes. In the context of the known proinflammatory effects of AII, it is postulated that the renoprotection conferred by angiotensin II receptor antagonism is at least partly related to the inhibition of NF-kappaB-dependent pathways. PMID- 15284301 TI - From Finland to fatland: beneficial effects of statins for patients with chronic kidney disease. PMID- 15284300 TI - Involvement of peripheral benzodiazepine receptor in the oxidative stress, death signaling pathways, and renal injury induced by ischemia-reperfusion. AB - The peripheral benzodiazepine receptor (PBR) is a critical component of the mitochondrial permeability transition pore, which is involved in the regulation of cell death. In the present study we investigated the role of PBR in the regulation of signaling pathways leading to apoptotic and necrotic damage and renal dysfunction in a rat model of ischemia-reperfusion. Renal ischemia reperfusion led to extended tubular apoptosis and necrosis that were associated with peroxidative damage, high levels of proapoptotic Bax expression, and low levels of antiapoptotic Bcl-2 expression, cleavage of death substrate, poly(ADP ribose) polymerase (PARP), and activation of a key effector of apoptosis, caspase 3. Rat pretreatment with a novel PBR antagonist, SSR180575, significantly decreased postreperfusion oxidative stress and tubular apoptosis and necrosis. This effect was associated with inhibition of caspase-3 activation and PARP cleavage, upregulation of Bcl-2, and downregulation of Bax. Furthermore, inhibition of PBR accelerated the recovery of normal renal function, as assessed by measurement of levels of plasma creatinine and blood urea nitrogen. These findings reveal a role for PBR as a modulator of necrotic and apoptotic cell death induced by ischemia-reperfusion and suggest that regulation of PBR may provide new therapeutic implications for the prevention of acute renal failure. PMID- 15284302 TI - Focal segmental glomerulosclerosis in nephrotic adults: presentation, prognosis, and response to therapy of the histologic variants. AB - The histopathologic diagnosis of primary focal segmental glomerulosclerosis (FSGS) has come to include a number of histologic lesions (variants), but the prognostic significance of these discrete lesions is controversial because published information regarding the presentation, course, and response to treatment is limited. A retrospective analysis was conducted of 87 nephrotic adult patients with biopsy-proven primary FSGS. Patients were categorized on the basis of histologic criteria into those with a classic scar (36 patients), the cellular or collapsing lesion (40 patients), or the tip lesion (11 patients) of FSGS to evaluate differences in presentation, response to therapy, and clinical outcomes. The clinical features at biopsy were similar among the three groups with the exception that patients with the tip lesion were older and patients with the collapsing lesion had more severe proteinuria. Over the course of follow-up, 63% of patients treated attained remission and the response to steroid therapy was similar among the groups (classic scar 53% versus collapsing lesion 64% versus tip lesion 78%; P = 0.45). The overall renal survival was significantly better for patients who entered remission compared with patients who did not enter remission (92% versus 33% at 10 yr; P < 0.0001). The renal survival at 10 yr for patients who entered remission was similar among the three groups (classic scar 100% versus tip lesion 100% versus collapsing lesion 80%; P = 0.61). In patients who did not enter remission, the renal survival at 10 yr was significantly worse for patients with collapsing lesion and tip lesion (classic scar 49% versus tip lesion 25% versus collapsing lesion 21%; P = 0.002). In conclusion, the prognosis for nephrotic FSGS patients who enter remission is excellent regardless of the histologic lesion. Because the remission rate after treatment is similar among patients with the histologic variants, response to therapy cannot be predicted on the basis of histology alone. Thus, nephrotic patients with primary FSGS should receive a trial of therapy irrespective of the histologic lesion when not contraindicated. PMID- 15284303 TI - Association between smoking and chronic renal failure in a nationwide population based case-control study. AB - For determining whether smoking is associated with an increased risk for chronic renal failure (CRF) overall and by type of renal disease, smoking data were analyzed from a nationwide population-based case-control study. Eligible as cases were native 18- to 74-yr-old Swedes whose serum creatinine for the first time and permanently exceeded 3.4 mg/dl (men) or 2.8 mg/dl (women). A total of 926 cases (78% of all eligible) and 998 control subjects (75% of 1330 randomly selected subjects from the source population), frequency matched to the cases by gender and age within 10 yr, were included. A face-to-face interview and a self administered questionnaire provided information about smoking habits and other lifestyle factors. Logistic regression models estimated odds ratios (OR) as measures of relative risk for disease-specific types of CRF among smokers compared with never-smokers. Despite a modest and nonsignificant overall association, the risk increased with high daily doses (OR among smokers of >20 cigarettes/d, 1.51; 95% confidence interval [CI], 1.06 to 2.15), long duration (OR among smokers for >40 yr, 1.45; 95% CI, 1.00 to 2.09), and a high cumulative dose (OR among smokers with >30 pack-years, 1.52; 95% CI, 1.08 to 2.14). Smoking increased risk most strongly for CRF classified as nephrosclerosis (OR among smokers with >20 pack-years, 2.2; 95% CI, 1.3 to 3.8), but significant positive associations were also noted with glomerulonephritis. This study thus suggests that heavy cigarette smoking increases the risk of CRF for both men and women, at least CRF classified as nephrosclerosis and glomerulonephritis. PMID- 15284304 TI - Inflammation, residual kidney function, and cardiac hypertrophy are interrelated and combine adversely to enhance mortality and cardiovascular death risk of peritoneal dialysis patients. AB - C-reactive protein (CRP), the prototype marker of inflammation, and cardiac hypertrophy are important prognostic indicators in dialysis patients. Residual renal function (RRF) has also been shown to influence survival of peritoneal dialysis (PD) patients. This study examined the relations between inflammation, RRF, and left ventricular hypertrophy (LVH) and determined whether inflammation, RRF, and LVH combine adversely to predict the outcomes of PD patients. A prospective observational study was performed in 231 chronic PD patients. Left ventricular mass index (LVMi), residual glomerular filtration rate (GFR), CRP, hemoglobin, serum albumin, and BP were determined at study baseline and related to outcomes. On univariate analysis, age (P = 0.002), dialysis duration (P = 0.004), coronary artery disease (P < 0.001), pulse pressure (P < 0.001), hemoglobin (P < 0.001), serum albumin (P = 0.032), log-CRP (P < 0.001), and GFR (P < 0.001) were significantly associated with log-LVMi. Log-CRP was positively correlated with pulse pressure (R = 0.218, P = 0.001) and negatively correlated with GFR (R = -0.272, P < 0.001). Multivariate analysis showed that log-CRP (P = 0.008) and RRF (P = 0.003) remained associated with log-LVMi independent of hemoglobin, serum albumin, arterial pulse pressure, and coronary artery disease. After follow-up for 30 +/- 14 mo, 34.2% patients had died. CRP, RRF, and LVMi each were significantly predictive of all-cause mortality and cardiovascular death. Kaplan-Meier analysis showed a significant increase in all-cause (P < 0.0001) and cardiovascular mortality (P < 0.0001) as the number of risk factors, namely CRP >/=50th percentile, no RRF, and LVMi>/= 50th percentile increased with the 2-yr all-cause mortality and cardiovascular death reaching as high as 61% and 46%, respectively, for patients who had all three risk factors. Compared with patients with none of the three risk factors, those with all three risk factors had an adjusted hazards ratio of 6.94 (P < 0.001) and 5.43 (P = 0.001) for all cause mortality and cardiovascular mortality, respectively. In conclusion, inflammation, RRF, and LVH are interrelated and combine adversely to increase mortality and cardiovascular death risk of PD patients. PMID- 15284305 TI - Renal function, digoxin therapy, and heart failure outcomes: evidence from the digoxin intervention group trial. AB - Renal dysfunction is a common complication for patients with heart failure, but its association with clinical outcomes has not been fully characterized. We evaluated the association of glomerular filtration rate (GFR) with heart failure survival and the effect of digoxin on heart failure outcomes across GFR strata. A secondary analysis from the Digitalis Intervention Group trial was conducted of 6800 outpatients with systolic heart failure. Renal function was categorized as estimated GFR (expressed in ml/min per 1.73 m(2)). All-cause mortality (mean, 3 yr) was inversely proportional to GFR (GFR >60, 31% mortality; GFR 30 to 60, 46% mortality; GFR <30, 62% mortality; P < 0.001). Among patients with a GFR <50, lower GFR were associated with greater adjusted mortality risk (GFR <30: hazard ratio [HR], 2.06, 95% confidence interval [CI], 1.69 to 2.51; GFR 30 to 40: HR, 1.42, 95% CI, 1.22 to 1.67; GFR 40 to 50: HR, 1.22, 95% CI, 1.07 to 1.39; GFR 50 to 60: HR, 1.00, referent). In contrast, participants with GFR 60 to 70 had similar risk (HR, 1.00; 95% CI, 0.88 to 1.14) compared with GFR 50 to 60, and those with GFR >70 had a slightly lower mortality hazard (0.89; 95% CI, 0.78 to 1.00). Linear spline analyses confirmed that GFR = 50 was the appropriate risk threshold; above 50, GFR had no association with mortality, whereas below 50, mortality risk increased sharply with declining GFR (spline coefficient, P < 0.0001). Digoxin efficacy did not differ by level of GFR (P = 0.19 for interaction). Renal dysfunction is strongly associated with mortality in stable outpatients with heart failure, notably in patients with estimated GFR <50 ml/min per 1.73 m(2). The effect of digoxin did not differ by level of renal function. PMID- 15284306 TI - Pure red cell aplasia secondary to epoetin alpha responding to Darbepoetin alpha in a patient on peritoneal dialysis. AB - Recombinant Human Erythropoietin (EPO) is extensively used for anemia in renal failure patients. It is normally safe and effective, improving symptoms of anemia. We report here a case of renal anemia in a patient undergoing peritoneal dialysis (PD) for end stage renal failure from renovascular disease. He initially responded well to Epoetin alpha (Eprex) but subsequently developed EPO antibodies and pure red cell aplasia (PRCA), becoming blood transfusion dependent. Subsequently, he responded to Darbepoetin alpha (Aranesp), without any complications in the presence of persisting EPO antibodies. This positive response, which restored hemoglobin values to normal, occurred despite general belief that any form of EPO will cross-react to EPO antibodies. This is the first case report where PRCA with EPO antibodies responded well to another EPO preparation without intervention from immunosuppression therapy. PMID- 15284307 TI - Mineral metabolism, mortality, and morbidity in maintenance hemodialysis. AB - Mortality rates in ESRD are unacceptably high. Disorders of mineral metabolism (hyperphosphatemia, hypercalcemia, and secondary hyperparathyroidism) are potentially modifiable. For determining associations among disorders of mineral metabolism, mortality, and morbidity in hemodialysis patients, data on 40,538 hemodialysis patients with at least one determination of serum phosphorus and calcium during the last 3 mo of 1997 were analyzed. Unadjusted, case mix adjusted, and multivariable-adjusted relative risks of death were calculated for categories of serum phosphorus, calcium, calcium x phosphorus product, and intact parathyroid hormone (PTH) using proportional hazards regression. Also determined was whether disorders of mineral metabolism were associated with all-cause, cardiovascular, infection-related, fracture-related, and vascular access-related hospitalization. After adjustment for case mix and laboratory variables, serum phosphorus concentrations >5.0 mg/dl were associated with an increased relative risk of death (1.07, 1.25, 1.43, 1.67, and 2.02 for serum phosphorus 5.0 to 6.0, 6.0 to 7.0, 7.0 to 8.0, 8.0 to 9.0, and >/=9.0 mg/dl). Higher adjusted serum calcium concentrations were also associated with an increased risk of death, even when examined within narrow ranges of serum phosphorus. Moderate to severe hyperparathyroidism (PTH concentrations >/=600 pg/ml) was associated with an increase in the relative risk of death, whereas more modest increases in PTH were not. When examined collectively, the population attributable risk percentage for disorders of mineral metabolism was 17.5%, owing largely to the high prevalence of hyperphosphatemia. Hyperphosphatemia and hyperparathyroidism were significantly associated with all-cause, cardiovascular, and fracture-related hospitalization. Disorders of mineral metabolism are independently associated with mortality and morbidity associated with cardiovascular disease and fracture in hemodialysis patients. PMID- 15284308 TI - Does the phosphate binder lanthanum carbonate affect bone in rats with chronic renal failure? AB - Adequate control of phosphate levels remains an important issue in patients with chronic renal failure (CRF). Lanthanum carbonate has been proposed as a new phosphate binder. Previous studies have shown a high phosphate binding capacity (>97%) and low gastrointestinal absorption of lanthanum, without serious toxic side effects in the presence of a normal renal function (NRF). Because of lanthanum's physicochemical resemblance to calcium, the possible effects of it on bone have to be considered. The aim of this study was to investigate the effects of lanthanum carbonate on bone histology in NRF and CRF rats after oral administration of the compound with doses of 100, 500, or 1000 mg/kg per d for 12 wk. Bone histomorphometry showed that CRF animals that received vehicle developed secondary hyperparathyroidism. Urinalysis of lanthanum-loaded CRF animals showed a dose-dependent decrease in urinary phosphorus excretion, which was clearly more pronounced in the CRF groups compared with NRF animals. Phosphatemia, however, remained normal. Lanthanum carbonate administration induced a dose-dependent decrease in bone formation rate and increase in osteoid area in CRF animals. Three of seven animals in the CRF-1000 group and one of eight animals in the NRF 100 group were classified as having a mineralization defect. The number of cuboidal osteoblasts, however, was not affected, indicating that bone changes were not due to a toxic effect of lanthanum on the osteoblast. Furthermore, lanthanum concentrations in the femur remained low and did not correlate with histomorphometric parameters. These findings suggest that the administration of high doses of phosphate binder (1000 mg/kg per d lanthanum carbonate), in combination with decreased 25-(OH) vitamin D(3) in the uremic state, resulted in phosphate depletion and followed by an increased mobilization of phosphorus out of bone and/or reduced incorporation into bone. There was no evidence that lanthanum had a direct toxic effect on osteoblasts. PMID- 15284309 TI - Angiotensin converting enzyme genotype and chronic allograft nephropathy in protocol biopsies. AB - Genotype DD of the angiotensin-converting enzyme (ACE) is not associated with an increased incidence of native renal diseases, although it could modulate progression to renal failure in patients who already display chronic lesions. Because its role in renal allograft degeneration is not well characterized, whether ACE genotype was associated with the prevalence of chronic allograft nephropathy (CAN) was studied, in a group of protocol biopsies from 180 patients, or with the incidence of CAN in 152 patients with at least two sequential biopsies. As a control group, ACE genotype was also studied in 41 donors and 72 healthy subjects. For analyzing the influence of ACE genotype in graft survival, patients were grouped into six categories (II-normal biopsy, ID-normal, DD normal, II-CAN, ID-CAN and DD-CAN). Finally, relative renal ACE mRNA levels were measured in 67 cases by real-time PCR using the delta threshold cycle method. ACE DD genotype was more frequent in patients who received a transplant than in control subjects (43.3% versus 30.1%, P = 0.026), but prevalence (DD = 42.7% versus non-DD = 42.2%) or incidence (DD = 24.6% versus non-DD = 29.9%) of CAN was not different regarding recipient ACE genotype. Furthermore, patients with the ACE-DD genotype and CAN had the poorest graft survival (II-normal = 100%, ID normal = 91%, DD-normal = 84%, II-CAN = 100%, ID-CAN = 66%, and DD-CAN = 36%; P = 0.034) and higher ACE mRNA levels than non-DD and CAN (DD = -3.36 +/- 2.35 versus non-DD = -5.65 +/- 1.72-fold in ACE copies; P = 0.012). It is concluded that ACE DD genotype is not associated with an increased prevalence or incidence of CAN but is actually associated with higher ACE mRNA levels and poorer graft survival in patients who already display CAN. PMID- 15284310 TI - Kidney-specific gene targeting. PMID- 15284311 TI - Erythropoietin is more than just a promoter of erythropoiesis. PMID- 15284312 TI - Some immaculate misconceptions. PMID- 15284313 TI - Oral history -- Walter Holland. PMID- 15284314 TI - Cost-effectiveness of organized versus opportunistic cervical cytology screening in Hong Kong. AB - BACKGROUND: To assess the cost-effectiveness of alternative cervical cancer screening strategies to inform the design and implementation of a government sponsored population-based screening programme in Hong Kong. METHODS: Cost effectiveness analysis using a computer-based model of cervical carcinogenesis was performed. Strategies included no screening, opportunistic screening (status quo), organized screening using either conventional or liquid-based cytology conducted at different frequencies. The main outcome measures were cancer incidence reduction, years of life saved (YLS), lifetime costs and incremental cost-effectiveness ratios. Data were from local hospitals and laboratories, clinical trials, prospective studies and other published literature. RESULTS: Compared with no screening, a simulation of the current situation of opportunistic screening using cervical cytology produced a nearly 40 per cent reduction in the lifetime risk of cervical cancer. However, with organized screening every 3, 4 and 5 years, corresponding reductions with conventional (and liquid-based) cytology were 90.4 (92.9), 86.8 (90.2) and 83.2 per cent (87.3 per cent) compared with no screening. For all cytology-based screening strategies, opportunistic screening was more costly and less effective than an organized programme of screening every 3, 4 and 5 years. Every 3-, 4- and 5-year screening cost $12,300, $7100 and $800 per YLS, each compared with the next best alternative. CONCLUSIONS: Compared with the status quo of opportunistic screening, adopting a policy of organized, mass cervical screening in Hong Kong can substantially increase benefits and reduce costs. PMID- 15284315 TI - Development and preliminary examination of the predictive validity of the Falls Risk Assessment Tool (FRAT) for use in primary care. AB - BACKGROUND: There is no validated assessment of an older person's risk of falling that is easily applied in primary care. We aimed to develop a two-part tool for use in primary care or the community. Part 1 includes a rapid assessment of the individual's risk of falling for administration by clinical or non-clinical staff. Part 2 (for clinical staff) includes guidance on further assessment, referral and interventions. We assessed the predictive validity of part 1. METHODS: The tool was developed by an expert panel following the updating of an existing systematic review of community-based prospective studies identifying risk factors for falling and modified in accordance with the feedback from extensive piloting. We assessed predictive validity by a questionnaire survey sent at baseline and 6 months to a random sample of 1000 people aged over 65 in one Primary Care Group area. RESULTS: Five items were included in part 1: history of any fall in the previous year, four or more prescribed medications, diagnosis of stroke or Parkinson's disease, reported problems with balance, inability to rise from a chair without using arms. The presence of three or more risk factors had a positive predictive value for a fall in the next 6 months of 0.57 (95 per cent confidence interval 0.43-0.69). Less than three risk factors had a negative predictive value of 0.86 (0.82-0.89), and a specificity of 0.92 (0.88-0.94). CONCLUSION: The tool may be useful for identifying people who would benefit from further assessment of their risk of falling and appropriate intervention. PMID- 15284316 TI - Iodine nutritional status of adults during a period of salt iodization. AB - BACKGROUND: To measure the iodine nutritional status of island adults in Zhoushan, China. METHODS: A comparison study was carried out in eight villages. These were selected from iodized salt and non-iodized salt districts of Zhoushan island by random sampling. The Mann-Whitney test was used to compare the urinary iodine concentration and dietary iodine intake between the two districts. RESULTS: The median of urinary iodine concentration in the non-iodized salt group was 90 micrograms/l, which was lower than 194 micrograms/l in the iodized salt group (u = 14.673; p < 0.000), whereas the median of daily dietary iodine intake in the two groups was 128 and 147 micrograms, respectively (u = 1.847; p = 0.065). There was no significant correlation between dietary iodine intake and urinary iodine concentration (p = 0.095). CONCLUSIONS: Salt iodization is necessary. Special characteristics of the island diet should also be considered. PMID- 15284317 TI - A health needs assessment of street-based prostitutes: cross-sectional survey. AB - BACKGROUND: Research with prostitutes has tended to concentrate on sexual health rather than wider health issues, and has failed to differentiate between street based prostitutes and off-street workers. Little is known about the general health and background of street-based sex workers, the group likely to have the greatest needs. METHODS: An interview-based survey amongst street-based sex workers in central Bristol was employed. RESULTS: Seventy-one women were interviewed. All reported chronic health problems. Sexually transmitted infections were between nine and 60 times more common than the general population. Many women (44 per cent; n = 31) had experienced sexual abuse and 38 per cent (n = 27) had been in care. Women who had experienced care left school earlier (14.1 versus 15.5 years; p < 0.0001 unpaired t-test) and were less likely to have their own children at home [1/18 (5.5 per cent) versus 8/25 (32 per cent); p = 0.06) The stillbirth rate was 50/1000. Most (97 per cent; n = 69) had been offered more money for unprotected sex. Half (51 per cent; n = 36) had unprotected sex in the last week. All had drug or alcohol dependency problems. In the last week, 22 per cent (n = 9/41) of injecting drug users had shared needles and 59 per cent (n = 24/41) had shared injecting equipment, despite most (96 per cent; n = 39/41) knowing the risks. CONCLUSIONS: The health and social inequalities experienced by this group are much worse than any group highlighted in the 'Tackling Health Inequalities Review 2002' and appear cross generational. In neither that report nor the Sexual Health and HIV Strategy report are sex workers identified as a particularly high priority group. There is the potential for their needs to continue to be unmet. PMID- 15284318 TI - Use of complementary or alternative medicine in a general population in Great Britain. Results from the National Omnibus survey. AB - BACKGROUND: A representative sample of the adults in England, Scotland and Wales was interviewed to estimate levels of use of complementary or alternative medicines (CAMs) and their socio-economic correlates. METHODS: The Omnibus survey is a multi-purpose survey carried out in the United Kingdom by the Office for National Statistics on behalf of non-profit making organizations. The survey is carried out in 2 out of 3 months each quarter using a stratified random, probability sample of households. An eight-question module was added to the interview schedule of the survey in March 2001. Topics included practitioner based use of 23 named CAM therapies in the past 12 months. The resulting data were analysed in conjunction with socio-economic and demographic variables. RESULTS: A response rate of 65 per cent (1794/2761) was achieved. An estimated 10.0 per cent of the population [95 per cent confidence interval (CI) 8.7-11.5 per cent] had received any CAM therapy from a practitioner in the past year. No individual therapy was used by more than 2 per cent of the sample. An estimated 6.5 per cent (95 per cent CI 5.4-7.6) had used one of the five main therapies: acupuncture, homeopathy, chiropractic, osteopathy or herbal medicine. Estimates of CAM use were similar in England, Scotland and Wales. There was a significant positive association between CAM use and non-manual social class (p < 0.002), age left full-time education (p < 0.001), and gross income over pounds sterling 15,600 (p < 0.001). More than half (52 per cent) of the respondents that had used CAM in the past year had not told their general practitioner. CONCLUSIONS: Strong correlations between the use of CAM and gross socio-economic indicators are demonstrated in the survey. Repeated national surveys of this type could provide a useful vehicle for collecting information about changing patterns of CAM use on a routine basis. PMID- 15284319 TI - NHS Direct derived data: an exciting new opportunity or an epidemiological headache? AB - NHS Direct, a national telephone helpline for health advice, was established in 1998 to provide health information and advice to callers and refer them to an appropriate service. This article briefly describes the nature of the NHS Direct call record and discusses issues relevant to the use of the data for disease surveillance and epidemiological purposes. Clinical decision support software [the NHS Clinical Assessment System (NHS CAS)] is used by NHS Direct to collect callers' demographic details and direct them to the appropriate level of care. Data relating to NHS Direct calls provide a timely snapshot of symptoms occurring in the community and are summarized in 'off the shelf' NHS CAS reports. Adapting the system to provide customized data extracts requires considerable development work. When interpreting NHS Direct derived data, particular attention should be given to the age distribution of callers, NHS Direct demand surges, call 'networking' and changes to the NHS CAS clinical algorithms. An increasingly rich source of baseline data, growing body of published work, and a more 'bedded down' NHS Direct service will further our understanding and acceptance of the value of the NHS Direct call record. PMID- 15284320 TI - Completeness of ascertainment by cancer registries: putting bounds on the number of missing cases. AB - BACKGROUND: When comparing cancer incidence or mortality rates between different regions, it is important to know how complete the registration data are on which these figures are based. A number of ways of estimating completeness have been proposed, but it is often difficult to say how precise these estimates are. We describe a computer program developed to produce measures of precision for estimates of completeness obtained by one such method, the flow method. METHODS: The program works by resampling the required data sets, and repeatedly calculating completeness estimates until convergence of the standard errors occurs. It was tested on colorectal tumours from a single health district, and empirical confidence limits for 1 and 5 year completeness were compared with those obtained by applying various normalizing transformations and a beta distribution. The method was then applied to tumours of the head and neck, breast and lung and the results compared with those from a capture-mark-recapture exercise carried out 4 years previously. RESULTS: The sampling distribution was close to normal for 1 year completeness, but much less so for 5 year completeness, as assessed by quantile plots. Approximation by a beta distribution was better than by normalizing transformation. Although there were differences between the results produced by the flow method and capture-recapture, the flow method is more reproducible and easier to apply. CONCLUSION: It is now possible to estimate confidence limits for the results of the flow method, and thus determine whether comparative results between registries are likely to be affected by sampling error. PMID- 15284321 TI - Quality of age data in patients from developing countries. AB - BACKGROUND: Age misreporting is common in demographic studies but the prevalence and magnitude of age misreporting in clinical cohorts is unknown. We analysed single-year age distribution and terminal digit preference in cancer patients from developing countries. METHOD: Age distribution was analysed by plotting a single-year age of 3874 cancer patients from 72 different countries, mainly from the Indian subcontinent and the Middle East, who resided in the UAE at the time of cancer diagnosis. Preference for age ending with digits '0' and '5' was evaluated using Whipple's index (WI), which has value 100 in cohorts without preference. Preference for all 10 terminal digits was expressed as the difference between the found and expected frequencies using Myers blended method and was graphed. RESULTS: Age data quality was low in cancer patients from the Indian subcontinent (WI = 177) and Middle Eastern countries (WI = 113-204). Females of all nationalities supplied better quality of age data (lower WI) than males. Preference for age ending with digits '0' and '5' was found in all populations except the UAE male citizens who did not show preference for terminal digit '0'. CONCLUSION: Age data quality in this cohort of patients from developing countries was low. Preference for age ending with numbers '0' and '5' is common. In studies conducted in developing countries, age data quality should be analysed as it may bias results and weaken the power of the study. PMID- 15284322 TI - Which patients spend more than 4 hours in the Accident and Emergency department? AB - OBJECTIVES: The NHS Plan has a target that no patient should spend longer than 4 hours in Accident and Emergency (A & E) by the end of 2004. The aim of this study is to describe the attendance characteristics of patients spending less than and more than 4 hours total time in A & E. METHODS: Data were collected from 10 A & E departments in the West Midlands NHS region for the period 1 April 2001 to 31 March 2002. Patients were split into three groups; those spending less than 4 hours, between 4 and 8 hours and over 8 hours in A & E. The groups were compared in terms of their attendance characteristics, these being demography, temporal patterns, arrival mode and disposal. The data were also entered into a multinomial logistic regression using SPSS. RESULTS: Overall, 83.0 per cent (range 76.7 - 94.0 per cent) of patients spent less than 4 hours in A & E ; 3.6% per cent (range 0.3-8.6 per cent) spent longer than 8 hours in A & E. The risk factors for spending over 4 hours in A & E were requiring admission, arriving by ambulance, arriving during the night, increasing age and higher levels of deprivation. Being admitted had the greatest effect on time spent in A & E, with a patient being 2.64 times more likely to spend 4-8 hours and 4.84 times more likely to spend over 8 hours in the department. CONCLUSIONS: This study points to admission and service provision at night as factors leading to long periods in A & E. However, these results can only act as a guide as the problems are different in different Trusts and each should analyse their problem before taking action. PMID- 15284323 TI - Comparing performance between coronary intervention centres requires detailed case-mix adjusted analysis. AB - This study compares 12 month clinical outcomes and procedural costs at two interventional centres with significant differences in crude mortality and revascularization outcomes between 1997 and 1998. Percutaneous coronary intervention (PCI) registry data on 1046 consecutive patients treated contemporaneously at two university centres were linked to hospital discharge and death data to provide 12 month follow-up information on survival and repeat revascularization. Costs were determined by detailed analysis of equipment use, length of stay and staff from 100 contemporary cases at each centre to derive a procedural cost model. This model was then applied retrospectively to estimate cost per procedure. Stents were used more frequently at one centre (56 versus 26 per cent, chi(2) test, p < 0.001) resulting in greater procedural cost [mean (SE), pounds sterling 1970 (34) versus pounds sterling 1521 (39), t-test, p < 0.001). One year repeat target vessel PCI was significantly greater at the centre using more stents (10.3 versus 5.6 per cent, chi(2) test, p = 0.005) and the need for any repeat revascularization (PCI or coronary artery by-pass surgery) was also significantly greater at this centre (18.4 versus 10.8 per cent, chi(2) test, p < 0.001). Cox regression revealed that after correction for case-mix the difference in the need for repeat target vessel PCI between the two centres was no longer significant (p = 0.15). In the two centres studied, crude differences in cost per case, mortality and the need for revascularization were largely accounted for by significant differences in case-mix. Comparison of outcomes and costs between centres should not be published without careful adjustment for differences in case-mix. PMID- 15284324 TI - Results and cost of meeting the National Service Framework for Coronary Heart Disease requirement for 12 month follow-up after acute coronary events. AB - BACKGROUND: The National Service Framework (NSF) for Coronary Heart Disease (CHD) set standards, targets and milestones. In the case of acute myocardial infarction (AMI) or coronary revascularization, Milestone 3 of Standard 12 requires a 12 month audit of exercise and smoking habit and of body mass index (BMI) for patients who have attended cardiac rehabilitation (CR). The targets are that 50 per cent of patients should be exercising regularly, not smoking and have a BMI of <30 kg/m(2). The purpose of this study was to find out whether the targets are realistic and to measure the cost of retrieving the data. METHODS: A postal questionnaire was used to follow up all the patients who attended our CR programme over a 12 month period. The project was costed. RESULTS: Four hundred and three CHD patients who had attended the programme between April 2001 and March 2002 were sent questionnaires 12 months after their index event. Their diagnoses were AMI in 147 (36.5 per cent), coronary artery surgery in 157 (39 per cent) and angioplasty in 99 (24.5 per cent). Completed questionnaires were received from 358 (89 per cent). Of the responders, 69 per cent were exercising regularly, 91.6 per cent were not smoking (73 per cent had been non-smokers before their index cardiac event) and 79 per cent had a BMI of <30 kg/m(2)(the figure at the start of rehabilitation had been 79 per cent). The cost of performing the audit was pounds sterling 1204. CONCLUSION: This audit is inexpensive. The targets for smoking and BMI set by the NSF were achieved by a very large margin before either the index cardiac event or starting CR. PMID- 15284325 TI - Outcome monitoring to facilitate clinical governance; experience from a national programme in the independent sector. AB - BACKGROUND: Since 1998 BUPA has used the Short-Form 36 (SF-36) questionnaire to monitor changes in health status after adult elective surgery. Over 70 independent hospitals across the United Kingdom have collected data on over 100000 patient episodes. SF-36 is one of a number of tools that support clinical governance in the sector. Results are reported confidentially, putting the emphasis on supporting a learning culture. FORMULATION OF PROBLEMS APPARENT AT 3 YEARS: Feedback was sub-optimal: discussions with hospital staff and consultants revealed that the league tables were hard to interpret, and there was uncertainty about the definition of outlier results. The number of patients recruited to the survey was variable across the hospitals. No grouping of low-volume procedures met with agreement. ACTION PLAN FOR YEAR 4: Use 'Shewhart' control charts to distinguish common and special cause variation in results; substitute a condition specific tool in one instance; adoption of an 'alert' process to prompt local audit of unusual results; use of a reminder letter to improve return rate of follow-up questionnaires; and focus programme on a list of 20 common procedures. Discuss these changes with the managerial and clinical leaders of each of BUPA's hospitals. CURRENT POSITION AT YEAR 5 : The use of Shewhart charts has been welcomed by managers and clinicians at BUPA's hospitals. The renewed confidence in the programme has encouraged constructive debate into allowing wider access to previously confidential results. Some changes in clinical practice have occurred. PMID- 15284326 TI - Factors contributing to the cause of a community outbreak of tuberculosis. AB - The diagnosis of pulmonary tuberculosis (TB) in a nursery teacher led to a total of 282 adults and children being screened for TB, and 67 of these contracted the condition. Latent and active factors mitigated against earlier diagnosis of the disease during the multiple contacts by the teacher with the healthcare system over 18 months. A series of barely inter-linked events meant that the system failed the patient and consequently the contacts who contracted the disease. The system errors were widespread and render possible a similar occurrence elsewhere. PMID- 15284327 TI - Environmental and social factors as determinants of respiratory dysfunction in junior schoolchildren in Moscow. AB - BACKGROUND: The process of industrialization of the USSR has left a legacy of widespread and often poorly controlled pollution which is widely believed to have adverse implications for health, in particular for respiratory disease among children. OBJECTIVES: To assess the relationship between area of residence and respiratory function in junior schoolchildren in different districts of Moscow. METHODS: A survey was conducted of 539 children aged 6-12 years who attend school and live in one of three districts of Moscow with varying ambient pollution levels. Spirometry [forced vital capacity (FVC) and forced expiratory volume in 1 s (FEV1)] was assessed at school by trained school health staff. Parents of the children completed a questionnaire asking about respiratory function and factors potentially associated with it, as well as about social and other factors that could influence respiratory development and the health status of their children. RESULTS: There was appreciable difference in the characteristics of the children from the three districts. Children from the lower pollutant districts were generally younger, had higher parental income, and were less frequently exposed to cigarette smoke at home. They were also less likely to report heavy lorry traffic in the streets outside their homes. After adjustment for age, gender and height the FVC was 7.6 per cent (3.6-11.5 per cent) lower in children from the medium pollution district and 9.9 per cent [95 per cent confidence interval (CI) 5.6-14.0 per cent] lower in children from the high pollution district compared with those in the least polluted district (p < 0.001 for trend). These differences were little affected by further adjustment for household income or exposure to household smoking. In contrast, FEV1 showed comparatively little variation across districts. The odds of a forced expiratory ratio (FER) <75 per cent were substantially lower in the high pollution compared with the low pollution district (odds ratio 0.10, 95per cent CI 0.03-0.32 after adjustment for age, gender and height), and there was clear evidence of a trend across pollution categories (p < 0.001). The frequency of reported allergy was also lower in the high pollution district. FVC increased, and the probability of a low FER decreased, with household income. CONCLUSION: Children from areas of high environmental pollution had lower lung capacity but also smaller risk of a low FER compared with those from cleaner areas. The extent to which these differences can be attributed to environmental pollution is unclear without more detailed study. However, socio-economic deprivation, which was associated with pollution, appears to be an important determinant of respiratory function although it was associated with a lower risk of an obstructive pattern of lung function tests. PMID- 15284328 TI - Communicable Disease and Health Protection Quarterly Review: October to December 2003. From the Health Protection Agency, Communicable Disease Surveillance Centre. PMID- 15284329 TI - Mailed questionnaires: length matters. PMID- 15284331 TI - Role of DNA polymerase eta in the bypass of abasic sites in yeast cells. AB - Abasic (AP) sites are major DNA lesions and are highly mutagenic. AP site-induced mutagenesis largely depends on translesion synthesis. We have examined the role of DNA polymerase eta (Poleta) in translesion synthesis of AP sites by replicating a plasmid containing a site-specific AP site in yeast cells. In wild type cells, AP site bypass resulted in preferred C insertion (62%) over A insertion (21%), as well as -1 deletion (3%), and complex event (14%) containing multiple mutations. In cells lacking Poleta (rad30), Rev1, Polzeta (rev3), and both Poleta and Polzeta, translesion synthesis was reduced to 30%, 30%, 15% and 3% of the wild-type level, respectively. C insertion opposite the AP site was reduced in rad30 mutant cells and was abolished in cells lacking Rev1 or Polzeta, but significant A insertion was still detected in these mutant cells. While purified yeast Polalpha effectively inserted an A opposite the AP site in vitro, purified yeast Poldelta was much less effective in A insertion opposite the lesion due to its 3'-->5' proofreading exonuclease activity. Purified yeast Poleta performed extension synthesis from the primer 3' A opposite the lesion. These results show that Poleta is involved in translesion synthesis of AP sites in yeast cells, and suggest that an important role of Poleta is to catalyze extension following A insertion opposite the lesion. Consistent with these conclusions, rad30 mutant cells were sensitive to methyl methanesulfonate (MMS), and rev1 rad30 or rev3 rad30 double mutant cells were synergistically more sensitive to MMS than the respective single mutant strains. PMID- 15284332 TI - Gene trap as a tool for genome annotation and analysis of X chromosome inactivation in human embryonic stem cells. AB - Human embryonic stem (ES) cells were suggested to be an important tool in transplantation medicine. However, they also play a major role in human genetics. Using the gene trap strategy, we have created a bank of clones with insertion mutations in human ES cells. These insertions occurred within known, predicted and unknown genes, and thus assist us in annotating the genes in the human genome. The insertions into the genome occurred in multiple chromosomes with a preference to larger chromosomes. Utilizing a clone where the integration occurred in the X chromosome, we have studied X-chromosome inactivation in human cells. We thus show that in undifferentiated female human ES cells both X chromosomes remain active and upon differentiation one chromosome undergoes inactivation. In the differentiated embryonic cells the inactivation is random, while in the extra-embryonic cells it is non-random. In addition, using a selection methodology, we demonstrate that in a minority of the cells partial inactivation and XIST expression occur even in the undifferentiated cells. We suggest that X chromosome inactivation during human embryogenesis, which coincides with differentiation, may be separated from the differentiation process. The genetic manipulation of human ES cells now opens new ways of analyzing chromosome status and gene expression in humans. PMID- 15284334 TI - 1 alpha,25-dihydroxyvitamin D3 transrepresses retinoic acid transcriptional activity via vitamin D receptor in myeloid cells. AB - Granulocytes and monocytes originate from a common committed progenitor cell. Commitment to either granulocytic or monocytic lineage is triggered by specific extracellular signals involving cytokines or nuclear receptor ligands (all-trans retinoic acid (RA) and 1 alpha,25-dihydroxyvitamin D(3)). Here we show that the stimulatory effect of 1 alpha,25-dihydroxyvitamin D(3) on the production of monocytic colonies (CFU-M) is accompanied by a repression of granulocytic colony (CFU-G) production. We further demonstrate that in bipotent HL-60 myeloid cells as in purified human CD34+ myeloid progenitor cells, the 1 alpha,25 dihydroxyvitamin D(3)-induced monocytic differentiation is concomitant with a direct inhibition of the RA-transcriptional activity. Indeed, a transrepression of the RAR beta RA-target gene promoter via formation of a nuclear complex involving VDR was identified in vitro and in vivo. The fact that binding of RXR RAR on DR3 is not observed suggests that contrary to RA-induced granulocytic differentiation, 1 alpha,25-dihydroxyvitamin D(3)-mediated monocytic differentiation requires positive and negative transcriptional controls both likely mediated by the RXR-VDR heterodimer. These novel findings implicate that 1 alpha,25-dihydroxyvitamin D(3) exerts a dominant negative effect on the RA dependent granulocytic commitment of human bone marrow cells via repression of the RA-target gene promoters. Hence, the transcriptional response to RA and 1 alpha,25-dihydroxyvitamin D(3) in myeloid cells depends on a complex combinatory pattern of interaction among different nuclear receptors with DNA. PMID- 15284333 TI - Detecting single DNA copy number variations in complex genomes using one nanogram of starting DNA and BAC-array CGH. AB - Comparative genomic hybridization to bacterial artificial chromosome (BAC)-arrays (array-CGH) is a highly efficient technique, allowing the simultaneous measurement of genomic DNA copy number at hundreds or thousands of loci, and the reliable detection of local one-copy-level variations. We report a genome-wide amplification method allowing the same measurement sensitivity, using 1 ng of starting genomic DNA, instead of the classical 1 microg usually necessary. Using a discrete series of DNA fragments, we defined the parameters adapted to the most faithful ligation-mediated PCR amplification and the limits of the technique. The optimized protocol allows a 3000-fold DNA amplification, retaining the quantitative characteristics of the initial genome. Validation of the amplification procedure, using DNA from 10 tumour cell lines hybridized to BAC arrays of 1500 spots, showed almost perfectly superimposed ratios for the non amplified and amplified DNAs. Correlation coefficients of 0.96 and 0.99 were observed for regions of low-copy-level variations and all regions, respectively (including in vivo amplified oncogenes). Finally, labelling DNA using two nucleotides bearing the same fluorophore led to a significant increase in reproducibility and to the correct detection of one-copy gain or loss in >90% of the analysed data, even for pseudotriploid tumour genomes. PMID- 15284335 TI - Inhibiting proteasomal proteolysis sustains estrogen receptor-alpha activation. AB - Estrogen receptor-alpha (ER alpha) is a ligand-dependent transcription factor that mediates physiological responses to 17 beta-estradiol (E2). Ligand binding rapidly down-regulates ER alpha levels through proteasomal proteolysis, but the functional impact of receptor degradation on cellular responses to E2 has not been fully established. In this study, we investigated the effect of blocking the ubiquitin-proteasome pathway on ER alpha-mediated transcriptional responses. In HeLa cells transfected with ER alpha, blocking either ubiquitination or proteasomal degradation markedly increased E2-induced expression of an ER responsive reporter. Time course studies further demonstrated that blocking ligand-induced degradation of ER alpha resulted in prolonged stimulation of ER responsive gene transcription. In breast cancer MCF7 cells containing endogenous ER alpha, proteasome inhibition enhanced E2-induced expression of endogenous pS2 and cathepsin D. However, inhibiting the proteasome decreased expression of progesterone receptor (PR), presumably due to the heterogeneity of the PR promoter, which contains multiple regulatory elements. In addition, in endometrial cancer Ishikawa cells overexpressing steroid receptor coactivator 1, 4-hydroxytamoxifen displayed full agonist activity and stimulated ER alpha mediated transcription without inducing receptor degradation. Collectively, these results demonstrate that proteasomal degradation is not essential for ER alpha transcriptional activity and functions to limit E2-induced transcriptional output. The results further indicate that promoter context must be considered when evaluating the relationship between ER alpha transcription and proteasome inhibition. We suggest that the transcription of a gene driven predominantly by an estrogen-responsive element, such as pS2, is a more reliable indicator of ER alpha transcription activity than a gene like PR, which contains a complex promoter requiring cooperation between ER alpha and other transcription factors. PMID- 15284336 TI - Key amino acids located within the transmembrane domains 5 and 7 account for the pharmacological specificity of the human V1b vasopressin receptor. AB - In mammals, the vasopressin V(1b) receptor (V(1b)-R) is known to regulate ACTH secretion and, more recently, stress and anxiety. The characterization of the molecular determinant responsible for its pharmacological selectivity was made possible by the recent discovery of the first V(1b) antagonist, SSR149415. Based upon the structure of the crystallized bovine rhodopsin, we established a three dimensional molecular model of interaction between the human V(1b)-R (hV(1b)-R) and SSR149415. Four amino acids located in distinct transmembrane helices (fourth, fifth, and seventh) were found potentially responsible for the hV(1b)-R selectivity. To validate these assumptions, we selectively replaced the leucine 181, methionine 220, alanine 334, and serine 338 residues of hV(1a)-R by their corresponding amino acids present in the hV(1b)-R (phenylalanine 164, threonine 203, methionine 324, and asparagine 328, respectively). Four mutants, which all exhibited nanomolar affinities for vasopressin and good coupling to phospholipase C pathway, were generated. hV(1a) receptors mutated at position 220 and 334 exhibited striking increase in affinity for SSR149415 both in binding and phospholipase C assays at variance with the hV(1a)-R modified at position 181 or 338. In conclusion, this study provides the first structural features concerning the hV(1b)-R and highlights the role of few specific residues in its pharmacological selectivity. PMID- 15284337 TI - Experimental Peptide Identification Repository (EPIR): an integrated peptide centric platform for validation and mining of tandem mass spectrometry data. AB - LC MS/MS has become an established technology in proteomic studies, and with the maturation of the technology the bottleneck has shifted from data generation to data validation and mining. To address this bottleneck we developed Experimental Peptide Identification Repository (EPIR), which is an integrated software platform for storage, validation, and mining of LC MS/MS-derived peptide evidence. EPIR is a cumulative data repository where precursor ions are linked to peptide assignments and protein associations returned by a search engine (e.g. Mascot, Sequest, or PepSea). Any number of datasets can be parsed into EPIR and subsequently validated and mined using a set of software modules that overlay the database. These include a peptide validation module, a protein grouping module, a generic module for extracting quantitative data, a comparative module, and additional modules for extracting statistical information. In the present study, the utility of EPIR and associated software tools is demonstrated on LC MS/MS data derived from a set of model proteins and complex protein mixtures derived from MCF-7 breast cancer cells. Emphasis is placed on the key strengths of EPIR, including the ability to validate and mine multiple combined datasets, and presentation of protein-level evidence in concise, nonredundant protein groups that are based on shared peptide evidence. PMID- 15284338 TI - Global investigation of p53-induced apoptosis through quantitative proteomic profiling using comparative amino acid-coded tagging. AB - p53-induced apoptosis plays a pivotal role in the suppression of tumorigenesis, and mutations in p53 have been found in more than 50% of human tumors. By comparing the proteome of a human colorectal cancer cell transfected with inducible p53 (DLD-1.p53) with that of the control DLD-1 cell line using amino acid-coded mass tagging (AACT)-assisted mass spectrometry, we have broadly identified proteins that are upregulated at the execution stage of the p53 mediated apoptosis. In cell culturing, the deuterium-labeled (heavy) amino acids were incorporated into the proteome of the induced DLD-1.p53 cells, whereas the DLD-1.vector cells were grown in the unlabeled medium. In high-throughput LC-ESI MS/MS analyses, the AACT-containing peptides were paired with their unlabeled counterparts, and their relative spectral intensities, reflecting the differential protein expression, were quantified. In addition, our novel AACT-MS method utilized a number of different heavy amino acids as internal markers that significantly increased the peptide sequence coverage for both quantitation and identification purposes. As a result, we were able to identify differentially regulated protein isozymes that would be difficult to distinguish by ICAT-MS methods and to obtain a large dataset of the proteins with altered expression in the late stage of p53-induced apoptosis. The regulated proteins we identified are associated with several distinct functional categories: cell cycle arrest and p53 binding, protein chaperoning, plasma membrane dynamics, stress response, antioxidant enzymes, and anaerobic glycolysis. This result suggests that the p53 induced apoptosis involves the systematic activation of multiple pathways that are glycolysis-relevant, energy-dependent, oxidative stress-mediated, and possibly mediated through interorganelle crosstalks. PMID- 15284339 TI - Lesion-induced enhancement of LTP in rat visual cortex is mediated by NMDA receptors containing the NR2B subunit. AB - There is emerging evidence that injury of the cerebral cortex is followed by processes of enhanced neuroplasticity. In the present study, we investigate the functional properties of NMDA receptors (NMDARs) in the surround of focal lesions with recordings of extracellular field potentials (FPs) in acute slices of rat visual cortex at survival times of 2-6 days. FPs were recorded in cortical layer III lateral to the lesion, while long-term potentiation (LTP) was induced by theta-burst stimulation (TBS) in layer IV. The predominantly AMPA receptor mediated FPs displayed a significantly enhanced LTP in the surround of the lesion at distances of 2-3.2 mm. The LTP was completely blocked by the NMDAR antagonist D-AP5. Ifenprodil, an antagonist of NMDARs containing the NR2B subunit, only slightly affected the LTP in slices from sham-operated animals, but significantly reduced the LTP in slices from lesioned rats. We quantitatively analysed the proportion of NMDARs containing the NR2B subunit after lesions by applying ifenprodil to pharmacologically isolated NMDAR-FPs. The NR2B antagonist reduced the NMDAR-FPs significantly more strongly at distances of 2.0-3.2 mm from the border of the lesion. This indicates that the early phase of increased synaptic long-term plasticity in the surround of cortical lesions is accompanied by an up regulation of NMDARs containing the NR2B subunit. PMID- 15284340 TI - Acute nociceptive somatic stimulus sensitizes neurones in the spinal cord to colonic distension in the rat. AB - The common co-existence of fibromyalgia and chronic abdominal pain could be due to sensitization of spinal neurones (SNs), as a result of viscero-somatic convergence. The objective of this study is to explore the influence of acute nociceptive somatic stimulation in the form of acid injections, into the ipsilateral somatic receptive field of neurones responsive to colorectal distension (CRD), and the potential role of ionotropic glutamate receptors on sensitization. Action potentials of CRD-sensitive SNs were recorded extracellularly from the lumbar (L(2)-L(5)) spinal cord. Stimulus-response functions (SRFs) to graded CRD (10-80 mmHg, 30 s) were constructed before and 30 min after ipsilateral injection of low pH (4.0, 100 microl) saline into the somatic receptive fields. In some experiments, cervical (C(1)-C(2)) spinalization was performed to eliminate supraspinal influence. The selective NMDA receptor antagonist CGS 19755 and AMPA receptor antagonist NBQX were injected (25 micromol kg(-1), i.v.) to examine their influence on sensitization. Three types of neurones were characterized as short-latency abrupt (SLA, n = 24), short latency sustained (SLS, n = 12), and long-latency (LL, n = 6) to CRD. Ipsilateral injection of low pH (4.0) in the somatic receptive field, but not the contralateral gastrocnemius (GN) or front leg muscles, sensitized responses of these neurones to CRD. Spinalization had no influence on the development of low pH-induced sensitization. Both CGS 19755 and NBQX significantly attenuated the sensitized response to CRD in intact and spinalized animals. Acute nociceptive somatic stimulus sensitizes CRD-sensitive SNs receiving viscero-somatic convergence. The sensitization occurs at the spinal level and is independent of supraspinal influence. Ionotropic glutamate receptors in the spinal cord are involved in sensitization. PMID- 15284341 TI - Phosphorylated guanidinoacetate partly compensates for the lack of phosphocreatine in skeletal muscle of mice lacking guanidinoacetate methyltransferase. AB - The effects of creatine (Cr) absence in skeletal muscle caused by a deletion of guanidinoacetate methyltransferase (GAMT) were studied in a knockout mouse model by in vivo (31)P magnetic resonance (MR) spectroscopy. (31)P MR spectra of hindleg muscle of GAMT-deficient (GAMT-/-) mice showed no phosphocreatine (PCr) signal and instead showed the signal for phosphorylated guanidinoacetate (PGua), the immediate precursor of Cr, which is not normally present. Tissue pH did not differ between wild-type (WT) and GAMT-/- mice, while relative inorganic phosphate (P(i)) levels were increased in the latter. During ischaemia, PGua was metabolically active in GAMT-/- mice and decreased at a rate comparable to the decrease of PCr in WT mice. However, the recovery rate of PGua in GAMT-/- mice after ischaemia was reduced compared to PCr in WT mice. Saturation transfer measurements revealed no detectable flux from PGua to gamma-ATP, indicating severely reduced enzyme kinetics. Supplementation of Cr resulted in a rapid increase in PCr signal intensity until only this resonance was visible, along with a reduction in relative P(i) values. However, the PGua recovery rate after ischaemia did not change. Our results show that despite the absence of Cr, GAMT-/ mice can cope with mild ischaemic stress by using PGua for high energy phosphoryl transfer. The reduced affinity of creatine kinase (CK) for (P)Gua only becomes apparent during recovery from ischaemia. It is argued that absence of Cr causes the higher relative P(i) concentration also observed in animals lacking muscle CK, indicating an important role of the CK system in P(i) homeostasis. PMID- 15284342 TI - Electroneutral ammonium transport by basolateral rhesus B glycoprotein. AB - The liver and kidney are important tissues for ammonium (NH4+/NH3) metabolism and excretion. The rhesus B glycoprotein (RhBG) is a membrane protein expressed in liver and kidney with similarity to NH4+ transporters found in microorganisms, plants and animals. In the kidney, RhBG is predominantly localized to basolateral membranes of distal tubule epithelia, including connecting tubules and collecting ducts. These epithelia display mainly electroneutral ammonium transport, in contrast to other tubular sites, where net NH4+ transport occurs. In accordance with its localization, human RhBG mediates saturable, electroneutral transport of the ammonium analogue methylammonium when heterologously expressed in Xenopus oocytes. Uptake of methylammonium saturates with a Km = 2.6 mm. Methylammonium uptake is inhibited by ammonium and this inhibition saturates with a Ki approximately 3 mm. Electric current measurements and intracellular pHi determinations suggest that RhBG acts as an electroneutral NH4+ -H+ exchanger. PMID- 15284343 TI - Signalling mechanisms underlying the rapid and additive stimulation of NKCC activity by insulin and hypertonicity in rat L6 skeletal muscle cells. AB - We have investigated the expression and regulation of the Na(+)-K(+)-2Cl(-) cotransporter (NKCC) by insulin and hyperosmotic stress in L6 rat skeletal muscle cells. NKCC was identified by immunoblotting as a 170 kDa protein in L6 myotubes and mediated 54% of K(+) ((86)Rb(+)) influx based on the sensitivity of ion transport to bumetanide, a NKCC inhibitor. The residual (86)Rb(+) influx occurred via the Na(+),K(+)-ATPase and other transporters not sensitive to bumetanide or ouabain. NKCC-mediated (86)Rb(+) influx was enhanced significantly ( approximately 1.6-fold) by acute cell exposure to insulin, but was inhibited significantly by tyrosine kinase inhibitors, wortmannin and rapamycin, consistent with a role for the insulin receptor tyrosine kinase, phosphoinositide 3 (PI3) kinase and mTOR, respectively, in cotransporter activation. In contrast, the hormonal activation of NKCC was unaffected by inhibition of the classical Erk signalling pathway. Subjecting L6 myotubes to an acute hyperosmotic challenge (420 mosmol l(-1)) led to a 40% reduction in cell volume and was accompanied by a rapid stimulation of NKCC activity ( approximately 2-fold). Intracellular volume recovered to normal levels within 60 min, but this regulatory volume increase (RVI) was prevented if bumetanide was present. Unlike insulin, activation of NKCC by hyperosmolarity did not involve PI3-kinase but was suppressed by inhibition of tyrosine kinases and the Erk pathway. While inhibition of tyrosine kinases, using genistein, led to a complete loss in NKCC activation in response to hyperosmotic stress, immunoprecipitation of NKCC revealed that the cotransporter was not regulated directly by tyrosine phosphorylation. Simultaneous exposure of L6 myotubes to insulin and hyperosmotic stress led to an additive increase in NKCC mediated (86)Rb(+) influx, of which, only the insulin-stimulated component was wortmannin-sensitive. Our findings indicate that L6 myotubes express a functional NKCC that is rapidly activated in response to insulin and hyperosmotic shock by distinct intracellular signalling pathways. Furthermore, activation of NKCC in response to hyperosmotic-induced cell shrinkage represents a critical component of the RVI mechanism that allows L6 muscle cells to volume regulate. PMID- 15284344 TI - Nitric oxide synthase inhibition with L-NAME reduces maximal oxygen uptake but not gas exchange threshold during incremental cycle exercise in man. AB - We hypothesized that the effective inhibition of nitric oxide synthase (NOS), achieved via systemic infusion of N(G)-nitro-l-arginine methyl ester (l-NAME), would reduce the gas exchange threshold (GET) and the maximal oxygen uptake (V(.)(O(2)max)) during incremental cycle exercise in man if NO is important in the regulation of muscle vasodilatation. Seven healthy males, aged 18-34 years, volunteered to participate in this ethically approved study. On two occasions, the subjects completed an incremental exercise test to exhaustion on an electrically braked cycle ergometer following the infusion of either l-NAME (4 mg kg(-1) in 50 ml saline) or placebo (50 ml saline, CON). At rest, the infusion of l-NAME resulted in a significant increase in mean arterial pressure (MAP; CON vs. l-NAME, 89 +/- 8 vs. 103 +/- 11 mmHg (mean +/- s.d.; P < 0.05)) and a significant reduction in heart rate (HR; CON vs. l-NAME, 60 +/- 12 vs. 51 +/- 8 beats min( 1); P < 0.01). At submaximal work rates, there was no significant difference in V(.)(O(2)) between the conditions and no difference in the GET (CON vs. l-NAME, 1.94 +/- 0.47 vs. 2.01 +/- 0.41 l min(-1)). However, at higher work rates, differences in V(.)(O(2)) between the conditions became more pronounced such that V(.)(O(2)max) was significantly lower with l-NAME (CON vs. l-NAME, 4.02 +/- 0.41 vs. 3.80 +/- 0.34 l min(-1); P < 0.05). The reduction in V(.)(O(2)max) was associated with a reduction in HR(max) (CON vs. l-NAME, 186 +/- 10 vs. 178 +/- 7 beats min(-1); P < 0.01). These results demonstrate that NOS inhibition with l NAME has no effect on GET but reduces V(.)(O(2)max) during large muscle group exercise in man, presumably by direct or indirect effects on cardiac output and muscle blood flow. PMID- 15284345 TI - The ventilatory responsiveness to CO(2) below eupnoea as a determinant of ventilatory stability in sleep. AB - Sleep unmasks a highly sensitive hypocapnia-induced apnoeic threshold, whereby apnoea is initiated by small transient reductions in arterial CO(2) pressure (P(aCO(2))) below eupnoea and respiratory rhythm is not restored until P(aCO(2)) has risen significantly above eupnoeic levels. We propose that the 'CO(2) reserve' (i.e. the difference in P(aCO(2)) between eupnoea and the apnoeic threshold (AT)), when combined with 'plant gain' (or the ventilatory increase required for a given reduction in P(aCO(2))) and 'controller gain' (ventilatory responsiveness to CO(2) above eupnoea) are the key determinants of breathing instability in sleep. The CO(2) reserve varies inversely with both plant gain and the slope of the ventilatory response to reduced CO(2) below eupnoea; it is highly labile in non-random eye movement (NREM) sleep. With many types of increases or decreases in background ventilatory drive and P(aCO(2)), the slope of the ventilatory response to reduced P(aCO(2)) below eupnoea remains unchanged from control. Thus, the CO(2) reserve varies inversely with plant gain, i.e. it is widened with hyperventilation and narrowed with hypoventilation, regardless of the stimulus and whether it acts primarily at the peripheral or central chemoreceptors. However, there are notable exceptions, such as hypoxia, heart failure, or increased pulmonary vascular pressures, which all increase the slope of the CO(2) response below eupnoea and narrow the CO(2) reserve despite an accompanying hyperventilation and reduced plant gain. Finally, we review growing evidence that chemoreceptor-induced instability in respiratory motor output during sleep contributes significantly to the major clinical problem of cyclical obstructive sleep apnoea. PMID- 15284346 TI - Alpha 5 subunit-containing GABAA receptors affect the dynamic range of mouse hippocampal kainate-induced gamma frequency oscillations in vitro. AB - Though all in vitro models of gamma frequency network oscillations are critically dependent on GABAA receptor-mediated synaptic transmission little is known about the specific role played by different subtypes of GABAA receptor. Strong expression of the alpha5 subunit of the GABAA receptor is restricted to few brain regions, amongst them the hippocampal dendritic layers. Receptors containing this subunit may be expressed on the extrasynaptic membrane of principal cells and can mediate a tonic GABAA conductance. Using hippocampal slices of wild-type (WT) and alpha5-/- mice we investigated the role of alpha5 subunits in the generation of kainate-induced gamma frequency oscillations (20-80 Hz). The change in power of the oscillations evoked in CA3 by increasing network drive (kainate, 50-400 nm) was significantly greater in alpha5-/- than in WT slices. However, the change in frequency of gamma oscillations with increasing network drive seen in WT slices was absent in alpha5-/- slices. Raising the concentration of extracellular GABA by bathing slices in the GABA transaminase inhibitor vigabatrin and blocking uptake with tiagabine reduced the power of gamma oscillations more in WT slices than alpha5-/- slices (43%versus 15%). The data suggest that loss of this GABAA receptor subunit alters the dynamic profile of gamma oscillations to changes in network drive, possibly via actions of GABA at extrasynaptic receptors. PMID- 15284347 TI - Physiological, neurochemical and morphological properties of a subgroup of GABAergic spinal lamina II neurones identified by expression of green fluorescent protein in mice. AB - The processing of sensory, including nociceptive, information in spinal dorsal horn is critically modulated by spinal GABAergic neurones. For example, blockade of spinal GABA(A) receptors leads to pain evoked by normally innocuous tactile stimulation (tactile allodynia) in rats. GABAergic dorsal horn neurones have been classified neurochemically and morphologically, but little is known about their physiological properties. We used a transgenic mouse strain coexpressing enhanced green fluorescent protein (EGFP) and the GABA-synthesizing enzyme GAD67 to investigate the properties of a subgroup of GABAergic neurones. Immunohistochemistry showed that EGFP-expressing neurones accounted for about one third of the GABAergic neurones in lamina II of the spinal dorsal horn. They constituted a neurochemically rather heterogeneous group where 27% of the neurones coexpressed glycine, 23% coexpressed parvalbumin and 14% coexpressed neuronal nitric oxide synthase (nNOS). We found almost no expression of protein kinase Cgamma (PKCgamma) in EGFP-labelled neurones but a high costaining with PKCbetaII (78%). The whole-cell patch-clamp technique was used to intracellularly label and physiologically characterize EGFP- and non-EGFP-expressing lamina II neurones in spinal cord slices. Sixty-two per cent of the EGFP-labelled neurones were islet cells while the morphology of non-EGFP-labelled neurones was more variable. When stimulated by rectangular current injections, EGFP-expressing neurones typically exhibited an initial bursting firing pattern while non-EGFP expressing neurones were either of the gap or the delayed firing type. EGFP expressing neurones received a greater proportion of monosynaptic input from the dorsal root, especially from primary afferent C-fibres. In conclusion, EGFP expression defined a substantial but, with respect to the measured parameters, rather inhomogeneous subgroup of GABAergic neurones in spinal lamina II. These results provide a base to elucidate the functional roles of this subgroup of GABAergic lamina II neurones, e.g. for nociception. PMID- 15284348 TI - Human cerebral venous outflow pathway depends on posture and central venous pressure. AB - Internal jugular veins are the major cerebral venous outflow pathway in supine humans. In upright humans the positioning of these veins above heart level causes them to collapse. An alternative cerebral outflow pathway is the vertebral venous plexus. We set out to determine the effect of posture and central venous pressure (CVP) on the distribution of cerebral outflow over the internal jugular veins and the vertebral plexus, using a mathematical model. Input to the model was a data set of beat-to-beat cerebral blood flow velocity and CVP measurements in 10 healthy subjects, during baseline rest and a Valsalva manoeuvre in the supine and standing position. The model, consisting of 2 jugular veins, each a chain of 10 units containing nonlinear resistances and capacitors, and a vertebral plexus containing a resistance, showed blood flow mainly through the internal jugular veins in the supine position, but mainly through the vertebral plexus in the upright position. A Valsalva manoeuvre while standing completely re-opened the jugular veins. Results of ultrasound imaging of the right internal jugular vein cross-sectional area at the level of the laryngeal prominence in six healthy subjects, before and during a Valsalva manoeuvre in both body positions, correlate highly with model simulation of the jugular cross-sectional area (R(2) = 0.97). The results suggest that the cerebral venous flow distribution depends on posture and CVP: in supine humans the internal jugular veins are the primary pathway. The internal jugular veins are collapsed in the standing position and blood is shunted to an alternative venous pathway, but a marked increase in CVP while standing completely re-opens the jugular veins. PMID- 15284349 TI - Electrophysiological and molecular characterization of the inward rectifier in juxtaglomerular cells from rat kidney. AB - Renin, the key element of the renin-angiotensin-aldosterone system, is mainly produced by and stored in the juxtaglomerular cells in the kidney. These cells are situated in the media of the afferent arteriole close to the vessel pole and can transform into smooth muscle cells and vice versa. In this study, the electrophysiological properties and the molecular identity of the K+ channels responsible for the resting membrane potential (approximately -60 mV) of the juxtaglomerular cells were examined. In order to increase the number of juxtaglomerular cells, afferent arterioles from NaCl-depleted rats were used, and > 90% of the afferent arterioles were renin positive at the distal end of the arteriole. Whole-cell and cell-attached single-channel patch-clamp experiments showed that juxtaglomerular cells are endowed with a strongly inwardly rectifying K+ channel (Kir). The channel was highly sensitive to inhibition by Ba2+ (inhibition constant 37 microM at 0 mV), but relatively insensitive to Cs+ and, with 142 mM K+ in the pipette, had a single-channel conductance of 31.5 pS. Immunocytochemical studies showed the presence of Kir2.1 but no signal for Kir2.2 in the media of the afferent arteriole. In PCR analyses using isolated juxtaglomerular cells, the mRNA for Kir2.1 and Kir2.2 was detected. Collectively, the results show that Kir2.1 is the dominant component of the channel. The current carried by these channels plays a decisive role in setting the membrane potential of juxtaglomerular cells. PMID- 15284350 TI - Selective block of the human 2-P domain potassium channel, TASK-3, and the native leak potassium current, IKSO, by zinc. AB - Background potassium channels control the resting membrane potential of neurones and regulate their excitability. Two-pore-domain potassium (2-PK) channels have been shown to underlie a number of such neuronal background currents. Currents through human TASK-1, TASK-2 and TASK-3 channels expressed in Xenopus oocytes were inhibited by extracellular acidification. For TASK-3, mutation of histidine 98 to aspartate or alanine considerably reduced this effect of pH. Zinc was found to be a selective blocker of TASK-3 with virtually no effect on TASK-1 or TASK-2. Zinc had an IC(50) of 19.8 microM for TASK-3, at +80 mV, with little voltage dependence associated with this inhibition. TASK-3 H98A had a much reduced sensitivity to zinc suggesting this site is important for zinc block. Surprisingly, TASK-1 also has histidine in position 98 but is insensitive to zinc block. TASK-3 and TASK-1 differ at position 70 with glutamate for TASK-3 and lysine for TASK-1. TASK-3 E70K also had a much reduced sensitivity to zinc while the corresponding reverse mutation in TASK-1, K70E, induced zinc sensitivity. A TASK-3-TASK-1 concatamer channel was comparatively zinc insensitive. For TASK-3, it is concluded that positions E70 and H98 are both critical for zinc block. The native cerebellar granule neurone (CGN) leak current, IK(SO), is sensitive to block by zinc, with current reduced to 0.58 of control values in the presence of 100 microM zinc. This suggests that TASK-3 channels underlie a major component of IK(SO). It has recently been suggested that zinc is released from inhibitory synapses onto CGNs. Therefore it is possible that inhibition of IK(SO) in cerebellar granule cells by synaptically released zinc may have important physiological consequences. PMID- 15284352 TI - Direct disease targeting in renal illness: from calcimimetics to cytokines. Proceedings of the 5th Masterclass in Nephrology. 16-17 May 2003, Barcelona, Spain. PMID- 15284353 TI - The clinical consequences of secondary hyperparathyroidism: focus on clinical outcomes. AB - Secondary hyperparathyroidism (SHPT) is a common occurrence in patients with chronic renal failure and is characterized by excessive serum parathyroid hormone (PTH) levels, parathyroid hyperplasia and imbalances in calcium and phosphorus metabolism. PTH acts as a uraemic toxin and it may be responsible for the following long-term consequences: renal osteodystrophy; non-skeletal abnormalities, including severe vascular and heart valve calcification; alterations in cardiovascular structure and function; immune dysfunction; and renal anaemia. The risk of developing SHPT is not the same for all uraemic patients. Black patients appear to have a higher risk of developing SHPT than Caucasian patients, and patients with diabetes have a lower risk than non diabetic patients. Current treatments include dietary phosphate restriction, oral phosphate binders, vitamin D and its analogues, and, in severe cases, parathyroidectomy. These treatments do not provide optimal treatment for many patients, and compounds that directly inhibit PTH secretion may prove a major step forward in the treatment of SHPT. PMID- 15284354 TI - Achieving therapeutic targets in the treatment of secondary hyperparathyroidism. AB - Disturbances in the control of extracellular ionized calcium and phosphorus concentrations, and vitamin D metabolism, in patients with chronic kidney disease (CKD) are associated with prolonged stimulation of the parathyroid glands. This results in increased synthesis and release of parathyroid hormone (PTH) and parathyroid hyperplasia-secondary hyperparathyroidism (SHPT). SHPT is in turn a major driver of the skeletal disturbance that characterizes renal osteodystrophy and is associated with vascular and other soft tissue calcification. Current therapeutic strategies based on vitamin D compounds and calcium-containing phosphate binders are difficult to implement effectively because both agents are associated with substantial, and often dose-limiting, calcaemic actions that prevent the attainment of treatment targets. Calcimimetics are novel agents that increase the sensitivity of calcium-sensing receptors in the parathyroid glands. Consequently, they allow simultaneous reduction of both PTH and extracellular calcium concentrations, thus differing from currently available vitamin D therapies. Reduction of the calcium-phosphorus product (Ca x P) is a consistent feature of calcimimetic therapy and may facilitate the achievement of SHPT treatment targets. PMID- 15284351 TI - PACAP modulation of the colon-inferior mesenteric ganglion reflex in the guinea pig. AB - We investigated the effect of pituitary adenylate cyclase activating peptide (PACAP) on the colon-inferior mesenteric ganglion (IMG) reflex loop in vitro. PACAP27 and PACAP38 applied to the IMG caused a prolonged depolarization and intense generation of fast EPSPs and action potentials in IMG neurones. Activation of PACAP-preferring receptors (PAC1-Rs) with the selective agonist maxadilan or vasoactive intestinal peptide (VIP)/PACAP (VPAC) receptors with VIP produced similar effects whereas prior incubation of the IMG with selective PAC1 R antagonists PACAP6-38 and M65 inhibited the effects of PACAP. Colonic distension evoked a slow EPSP in IMG neurones that was reduced in amplitude by prolonged superfusion of the IMG with either PACAP27, maxidilan, PACAP6-38, M65 or VIP. Activation of IMG neurones by PACAP27 or maxadilan resulted in an inhibition of ongoing spontaneous colonic contractions. PACAP-LI was detected in nerve trunks attached to the IMG and in varicosities surrounding IMG neurones. Cell bodies with PACAP-LI were present in lumbar 2-3 dorsal root ganglia and in colonic myenteric ganglia. Colonic distension evoked release of PACAP peptides in the IMG as measured by radioimmunoassay. Volume reconstructed images showed that a majority of PACAP-LI, VIP-LI and VAChT-LI nerve endings making putative synaptic contact onto IMG neurones and a majority of putative receptor sites containing PAC1-R-LI and nAChR-LI on the neurones were distributed along secondary and tertiary dendrites. These results suggest involvement of a PACAP ergic pathway, operated through PAC1-Rs, in controlling the colon-IMG reflex. PMID- 15284355 TI - The need for better control of secondary hyperparathyroidism. AB - Secondary hyperparathyroidism (SHPT), a frequent complication of chronic kidney disease, develops in response to an imbalance in the serum levels of calcium, phosphorus and vitamin D as a result of altered metabolism. Raised serum levels of parathyroid hormone (PTH) and calcium-phosphorus product have a major effect on morbidity and mortality in dialysis patients. The new Kidney Disease Outcomes Quality Initiative (K/DOQI) guidelines, formulated by the National Kidney Foundation in the USA, propose strict targets for the control of serum levels of PTH, calcium and phosphorus. Meeting these targets will be a challenge for clinicians, because the traditional therapies for SHPT, such as vitamin D sterols and calcium-based phosphate binders, often exacerbate mineral imbalances. Results from a number of recent studies indicate that the majority of haemodialysis patients currently do not meet these new targets. Thus, there is a definite need to improve PTH, calcium and phosphate management of dialysis patients to reduce the incidence of uraemic bone disease and related disturbances of mineral metabolism as well as their unacceptably high cardiovascular morbidity and mortality. PMID- 15284356 TI - Modulation and action of the calcium-sensing receptor. AB - The discovery and cloning of the calcium-sensing receptor (CaR) in 1993 has led to a better understanding of the regulation of calcium homoeostasis. Following activation by extracellular calcium ions, the CaR triggers a cascade of intracellular events. These events result in the release of secondary messengers, which have a number of biological effects, the most important of which is a reduction in parathyroid hormone (PTH) secretion. The way in which calcium acts on the CaR varies depending on the cell type. In the parathyroid gland cell, activation of the CaR by elevated serum levels of calcium leads to a decrease in PTH secretion. In the kidney, CaR activation is thought to have several different actions, leading to enhanced reabsorption of sodium chloride and increased calcium and magnesium excretion in the renal tubules. CaRs are also found in other tissues in the body that are not involved in calcium homoeostasis, suggesting that the CaR has actions that are not associated with calcium homoeostasis. In patients with end-stage renal disease, parathyroid gland hyperplasia is associated with downregulation of the CaR. Discovery of the CaR has allowed the development of a group of drugs called calcimimetics, which mimic or potentiate the actions of extracellular calcium on the CaR. These compounds have considerable potential for the treatment of primary and secondary hyperparathyroidism. PMID- 15284357 TI - Clinical experience with cinacalcet HCl. AB - Secondary hyperparathyroidism (SHPT) is associated with parathyroid gland hyperplasia, increased parathyroid hormone (PTH) production and secretion, disturbed bone and mineral metabolism, soft tissue calcification and an increased risk of death. The condition is an almost universal complication of end-stage renal disease (ESRD) and currently is managed by treatment with phosphate binders and vitamin D compounds, both of which are associated with significant side effects, including hypercalcaemia and hyperphosphataemia. Therapy with calcimimetics is a new approach to the treatment of SHPT. These agents act at the calcium-sensing receptor (CaR), where they increase the sensitivity of the receptor to ionized serum calcium. Activation of the CaR results in a rapid reduction in PTH secretion. The calcimimetic drug cinacalcet HCl currently is undergoing clinical trials in dialysis patients who have uncontrolled SHPT, despite treatment with vitamin D compounds and/or phosphate binders. Clinical trials have confirmed that the drug rapidly reduces plasma PTH, phosphorus and calcium-phosphorus product (Ca x P) levels, and that levels of PTH, phosphorus and Ca x P remain suppressed for up to 3 years. In clinical trials, cinacalcet HCl was a well-tolerated drug; only nausea and vomiting occurred more frequently in patients who took cinacalcet HCl than in those who took placebo, and the occurrence of transient hypocalcaemia was limited to the initial phase of the treatment. Cinacalcet HCl is therefore a potentially highly effective and well tolerated treatment for SHPT in patients with ESRD. PMID- 15284358 TI - Causes and therapy of microinflammation in renal failure. AB - Microinflammation in renal failure has been the subject of numerous studies, but the causes of the inflammatory response in these patients are not clear. There are several potential causes and possible therapies for microinflammation, and they are discussed in this review with regard to uraemia and acidosis, heart failure and volume overload, oxidative stress and iron therapy, and bioincompatibility, especially regarding dialysis membranes. In addition, issues regarding dialysate contamination and access site infection are examined, followed by a discussion of possible drug therapy for microinflammation with angiotensin-converting enzyme inhibitors, angiotensin receptor antagonists, statins, aspirin, and antioxidants, such as vitamin E. PMID- 15284359 TI - Interleukin/cytokine profiles in haemodialysis and in continuous peritoneal dialysis. AB - The uraemic syndrome is a complex condition that results from the retention of "waste" compounds that normally would be excreted into the urine or catabolized by the kidneys. In addition, inflammation has been implicated in symptoms associated with uraemia, including its role in the malnutrition-inflammation atherosclerosis syndrome. Regarding vascular disease, traditional risk factors such as hypertension and gender do not seem to have the same significance in the uraemic population compared with patients without renal failure, and so the possibility has been raised that the uraemic toxins that result in the uraemic syndrome could play a role in this process. In this review, various questions are addressed regarding the involvement of cytokines in uraemia and the effects of dialysis membranes and fluids in patients receiving haemodialysis or peritoneal dialysis on cytokine levels. The effects of non-dialysis-related factors on levels of cytokines, mortality rates and other uraemic disorders are also discussed. It is concluded that cytokines are undoubtedly retained in uraemia, and that the loss of renal excretion almost certainly plays a key role in this process. Many cytokines have a pro-inflammatory role, probably resulting in a number of clinical events that are related to the increased morbidity and mortality of uraemic and haemodialysis patients. Any adjustment of the subtle balance between pro- and anti-inflammation by medical interventions should be conducted carefully because of an enhanced risk of serious infectious episodes. Bioincompatibility of dialysis techniques probably enhances the generation of cytokines as well as other uraemic toxins. PMID- 15284360 TI - Effect of recombinant human erythropoietin on inflammatory status in dialysis patients. AB - Under normal conditions, inflammatory status is regulated by a complex equilibrium between plasma and intracellular mediators. This equilibrium is broken in patients receiving dialysis, which can lead to chronic inflammation causing deleterious consequences on their organs and systems. During recent years, substances that can inhibit the effects of inflammation have been sought. The results of these investigations have produced controversial data on the effects of recombinant human erythropoietin (epoetin) and, in this review, the effects of epoetin on the inflammatory status of dialysis patients are discussed. Aspects discussed include biomarkers of inflammation, and the relationships between epoetin, growth factors, endothelial dysfunction, endothelial fibrinolytic capacity, endothelial damage and oxidative stress. Relationships between epoetin and inflammation in non-uraemic patients are also addressed, as are associations between the malnutrition-inflammation-atherosclerosis syndrome and endothelial dysfunction. It is concluded that although epoetin administration in non-uraemic rats has been shown to have an anti-inflammatory and cytoprotective effect, the mechanisms responsible for regulating inflammation in uraemia are very complex and partially contradictory. The changes in pro thrombotic and pro-atherogenic factors in dialysis patients require further study to evaluate all the factors implicated in the atherogenic process. PMID- 15284361 TI - Anaemia after renal transplantation. AB - Although the presence of anaemia after renal transplantation is well known, specific data on the prevalence and risk factors are scarce. Results from the recent TRansplant European Survey on Anemia Management (TRESAM) survey, conducted in 4263 recipients of a renal transplant from 72 centres in Europe, revealed that 38.6% of patients were anaemic [haemoglobin (Hb) concentrations < or =13 g/dl for male patients and < or =12 g/dl for female patients]. Of these patients, 11.6% had moderate anaemia (Hb concentrations >11 and < or =12 g/dl for male patients and >10 and < or =11 g/dl for female patients), while 8.5% had severe anaemia (Hb concentrations < or =11 g/dl for male patients and < or =10 g/dl for female patients). A strong association existed between Hb concentration and renal graft function. Of the patients with a serum creatinine level >2 mg/dl (which indicates impaired kidney function), 60.1% were anaemic, compared with 29.0% of those with a serum creatinine level < or =2 mg/dl (P<0.01). Other risk factors for anaemia include therapy with angiotensin-converting enzyme inhibitors or angiotensin II receptor antagonists, the use of azathioprine or mycophenolate mofetil, kidneys from older donors and recent infections. Furthermore, only 18.8% of patients with severe anaemia were treated with erythropoietic therapy. The findings from the TRESAM survey are in agreement with the results from another recently published study that included 128 renal transplant patients from two centres in the USA, who were followed for 5 years after transplantation. It was found that 30% of patients were anaemic at some point after transplantation. The prevalence increased with time after transplantation, with 26% of patients being anaemic 5 years' post-transplant. A multivariate logistic regression model identified three risk factors for post-transplant anaemia: serum total CO(2), blood urea nitrogen and creatinine. There is an unexpectedly high incidence of anaemia in patients with a functioning renal transplant: around one-third of these patients are anaemic. Most of the evidence suggests that impaired erythropoietin production by the renal allograft is the most important pathogenic factor of post-transplant anaemia. Whether this high incidence of anaemia may be an additional cardiovascular risk factor in renal transplant patients remains to be proven. However, there does not appear to be any reason why anaemic renal transplant recipients should not be treated like any other patients with renal anaemia. PMID- 15284362 TI - Vascular calcification in patients with end-stage renal disease. AB - Vascular calcification is the most common type of extra-osseous calcification in end-stage renal disease (ESRD), manifesting as both medial and intimal calcification of large arteries. It is highly prevalent, often progressive and is associated with reduced arterial elasticity and increased mortality. Risk factors for calcification in ESRD include age, duration of dialysis, diabetes mellitus, most probably an elevated calcium-phosphorus product (Ca x P) level, the dose of calcium-containing phosphate binders and the induction of the systemic inflammatory response. Uraemic calcification was thought to be a largely physico chemical process facilitated by elevated Ca x P (i.e. "metastatic" calcification). It is now well established, however, that vascular smooth muscle cells actively take up phosphate to form bioapatite. This process is associated with a phenotypic transformation of vascular smooth muscle cells during which they express osteoblast markers. In addition to phosphate, various other factors are likely to increase bioapatite formation, e.g. lipids and inflammatory cytokines. There have also been relatively new insights relating to the role of endogenous inhibitors of calcification [i.e. matrix Gla protein and fetuin-A (alpha(2)-Heremans-Schmid glycoprotein)], in particular the downregulation of fetuin-A in systemic inflammation. Decreased serum fetuin-A has been shown to be associated with a reduced capacity to inhibit calcium phosphate precipitation in vitro and is predictive of mortality in dialysis patients. These new insights into pathogenesis may lead to better prevention and treatment of calcification (e.g. with calcimimetics, anti-cytokines, etc.). However, the only preventive approach to have been established prospectively to date is the replacement of calcium-containing phosphate binders with sevelamer HCl, a non-calcaemic phosphate binder. Yet, it remains unclear whether sevelamer HCl reduces vascular calcification by preventing episodes of hypercalcaemia and/or by reducing low density lipoprotein (LDL)-cholesterol levels. PMID- 15284363 TI - Inflammatory proteins as predictors of cardiovascular disease in patients with end-stage renal disease. AB - Patients with end-stage renal disease (ESRD) are at high risk from potentially devastating cardiovascular sequelae due to the unique clustering of risk factors in these patients. Inflammation is believed to play a key role in the pathogenesis of these cardiovascular lesions. Both pro- and anti-inflammatory cytokines produced from monocytes, and also from adipocytes, have been studied in this regard. Pro-inflammatory cytokines, although cytoprotective acutely, correlate with increased risk of cardiovascular disease (CVD) in chronic situations. Conversely, elevated levels of anti-inflammatory mediators are associated with increased patient survival times. Statistical modelling, calculation of relative risk and cost considerations indicate that determination of serum C-reactive protein levels may be a useful predictor of CVD in ESRD patients. Adipocytes are a rich source of many of the same cytokines produced by monocytes, including interleukin-6, tumour necrosis factor-alpha, as well as adipocyte-specific proteins, leptin and adiponectin (ADPN). ADPN, which is produced in much greater quantities than leptin, is inversely related to body mass index and to insulin resistance, suggesting a possible role in type 2 diabetes. Additionally, ADPN has been shown to modulate the endothelial inflammatory response in vitro. Plasma ADPN levels are an inverse predictor of cardiovascular outcomes among patients with ESRD. Furthermore, ADPN is related to several metabolic risk factors in a manner consistent with the hypothesis that this protein acts as a protective factor for the cardiovascular system. PMID- 15284364 TI - Could anti-inflammatory cytokine therapy improve poor treatment outcomes in dialysis patients? AB - Mortality in dialysis patients is greater than that in the general population across all age groups. The disparity in mortality is greatest among patients aged under 35 years. Chronic kidney disease (CKD) is associated with the malnutrition, inflammation and atherosclerosis (MIA) syndrome, which helps to explain the high mortality rates among patients with CKD. Paradoxically, CKD patients exhibit signs of immune suppression as well as immune system activation. Chronic inflammation and immune system activation are not only integral to the MIA syndrome, but also may underlie resistance to erythropoietin treatment in patients with anaemia. Chronic immune system activation is reflected by abnormally raised T-lymphocyte and monocyte expression of both pro- and anti inflammatory cytokines. Patients who respond well to erythropoietin treatment exhibit fairly normal expression of these cytokines. Patients who persistently fail to respond, however, express abnormally raised levels of the pro inflammatory cytokines tumour necrosis factor-alpha (TNF-alpha) and interferon gamma (IFN-gamma), which are also known to inhibit erythropoiesis. Paradoxically, these patients also express abnormally high levels of the anti-inflammatory cytokines interleukin (IL)-10 and IL-13. Although anti-inflammatory in nature, these cytokines might also affect erythropoiesis. One strategy to overcome the problem of chronic inflammation in anaemic patients with CKD may be treatment with the phosphodiesterase inhibitor, pentoxifylline. Preliminary results suggest that once-daily treatment with 400 mg of pentoxifylline orally not only can reduce T-cell expression of TNF-alpha and IFN-gamma, but can also restore the response to erythropoietin and improve haemoglobin levels. Ongoing studies will investigate further the use of pentoxifylline in erythropoietin resistance. PMID- 15284365 TI - Dietary restriction and immune function. AB - Dietary restriction is beneficial in preventing a multitude of diseases, many of which may involve the immune system in their etiology. Recent reports examining dietary restriction focused on T lymphocytes and macrophages. Dietary restriction delays the onset of T-lymphocyte-dependent autoimmune disease; this may be attributed to improved antioxidant defense mechanisms, blunting shifts in T lymphocyte subset proportions and preventing DNA mutation frequencies. The beneficial effects of dietary restriction were shown in both the CD4 and CD8 T lymphocyte subsets as well as in various immune compartments such as the spleen, mesenteric lymph nodes, peripheral blood, thymus, and salivary glands. In contrast, dietary restriction may have negative effects on macrophage function because recent evidence showed that dietary restriction rendered mice more susceptible to peritonitis and stimulated macrophages produced lower amounts of cytokines. The application of dietary restriction regimens to humans would be difficult; however, understanding the biochemical and molecular targets of dietary restriction in the immune system may lead to the development of new dietary strategies to delay or prevent the onset of aging, cancer, and autoimmune disease. PMID- 15284366 TI - 1,25 dihydroxycholecalciferol-mediated calcium absorption and gene expression are higher in female than in male mice. AB - Recent advances in bone and calcium (Ca) metabolism have relied upon genetically modified mice. However, although human studies have identified gender as an important modulator of Ca metabolism, its effect on Ca metabolism has not been examined in mice. Here we examined basal and vitamin D-regulated Ca absorption (in situ ligated loops) and mRNA levels for the apical membrane calcium channel, TRPV6, and the calcium binding protein, calbindin D(9k) (CaBP) mRNA levels (real time PCR) in duodenum of female and male mice. At 2 mo of age, females fed a 5 g Ca/kg diet had higher Ca absorption (62.3 +/- 4.8 vs. 47 +/- 3.6%) and TRPV6 mRNA levels than males even though plasma 1,25 dihydroxyvitamin D [1,25(OH)(2) D] was not different. In mice fed high (20 g/kg), normal (5 g/kg), or low (0.2 g/kg) Ca diets for 7 d to alter plasma 1,25(OH)(2) D (91 +/- 12, 322 +/- 25, and 587 +/- 43 pmol/L, respectively), the relation between Ca absorption (slope = 0.116 vs. 0.084, P = 0.021) or duodenum TRPV6 mRNA (slope = 0.042 vs. 0.025, P = 0.034) and circulating 1,25(OH)(2) D was steeper in females. After a single 1,25(OH)(2) D injection (200 ng/100 g body weight), peak induction of TRPV6 mRNA was 2-fold greater (at 6 h) and CaBP mRNA was 20% higher in females (at 16 h). Duodenal vitamin D receptor mRNA levels did not differ between genders. Our data indicate that female mice are more sensitive to changes in serum 1,25(OH)(2) D levels than males and that this must be considered when using mice to study calcium and bone biology. PMID- 15284367 TI - Enteral administration of soybean phosphatidylcholine enhances the lymphatic absorption of lycopene, but reduces that of alpha-tocopherol in rats. AB - Dietary phosphatidylcholine (PC) increases the lymphatic absorption of triglyceride (TG). This result suggests that dietary PC might also enhance the absorption of other fat-soluble nutrients. We examined the effects of PC on lycopene and alpha-tocopherol absorption in male rats fitted with a thoracic lymph cannula. The lymphatic output was collected after administration of 1 mL of emulsified test oils containing lycopene and/or alpha-tocopherol in 3 separate experiments. The sodium taurocholate-emulsified test oils contained soybean oil (SO; 113 micromol triglyceride), SO containing soybean PC (SPC; 82.5 micromol SO plus 30.5 micromol purified soybean PC) or SO containing egg PC (EPC; 82.5 micromol SO plus 30.5 micromol purified egg PC) with both lycopene and alpha tocopherol (Expt. 1) or SO, SPC, or EPC with lycopene (Expt. 2) or alpha tocopherol alone (Expt. 3). In rats administered SPC or EPC, the lymphatic outputs of TG and lycopene were higher, and that of alpha-tocopherol was lower compared with rats administered SO (Expt. 1). The absorption rate for lycopene increased from 0.59% (SO group) to 2.16 and 1.28% in the SPC and EPC groups (P < 0.05), respectively, whereas the corresponding rates for tocopherol were 21.5% for the SO, 14.8% for the SPC, and 12.9% for the EPC groups. The increase in lycopene, but not in triglyceride absorption, was higher in the SPC than in the EPC groups. The promotive effects of SPC and EPC were decreased when lycopene alone was added to the test lipids (Expt. 2), and the inhibitory effects of PC were reduced when alpha-tocopherol alone was added to the test lipids (Expt. 3). Dietary PC increased the lymphatic output of lycopene and TG and decreased that of alpha-tocopherol, suggesting that differences exist between lycopene and alpha tocopherol in the absorptive mechanisms. The present results also show that the promotive effects of PC on lycopene absorption are influenced by the type of fatty acids in PC. PMID- 15284368 TI - Soyasaponin I and sapongenol B have limited absorption by Caco-2 intestinal cells and limited bioavailability in women. AB - A human study was conducted to evaluate soyasaponin bioavailability in humans. Eight healthy women ingested a single dose of concentrated soy extract containing 434 micromol of group B soyasaponins, the predominant form of soyasaponins in soybeans. Neither soyasaponins nor their metabolites were detected in a 24-h urine collection. Soyasapogenol B, a major metabolite of group B soyasaponins, was found (36.3 +/- 10.2 micromol) in a 5-d fecal collection but no group B soyasaponins were detected. A human colon cancer Caco-2 cell model was used to evaluate the absorbability of soyasaponins at the mucosal level. The mucosal transfers of soyasaponin I and soyasapogenol B were 0.5-2.9 and 0.2-0.8%, respectively, after 4-h incubation on the Caco-2 monolayer. The apical to basolateral absorptions of soyasaponin I and soyasapogenol B were low with P(app) of 0.9 to 3.6 x 10(-6) and 0.3 to 0.6 x 10(-6) cm/s, respectively. The transport rate and cell uptake of soyasaponin I were saturable and concentration independent. In contrast, soyasapogenol B was taken up by Caco-2 cells in a concentration-dependent manner. Soyasaponin I had no apparent cytotoxic effect on Caco-2 cells at concentrations up to 3 mmol/L, whereas soyasapogenol B at 1 mmol/L or more significantly reduced cell viability. Therefore, ingested soyasaponins have low absorbability in human intestinal cells and seem to be metabolized to soyasapogenol B by human intestinal microorganisms in vivo and excreted in the feces. PMID- 15284369 TI - Isolated soy protein consumption reduces urinary albumin excretion and improves the serum lipid profile in men with type 2 diabetes mellitus and nephropathy. AB - Soy protein was shown to exhibit several beneficial effects on renal function in nondiabetic patients with nephropathy, and to improve serum lipids. This study examined the effects of isolated soy protein consumption on urinary albumin excretion, serum lipids, plasma amino acids, and isoflavones in diabetic patients with nephropathy. Male patients (n = 14) with type 2 diabetes and nephropathy were followed in a crossover design for 7 mo. The study comprised two 8-wk intervention periods, placed between a 4-wk lead-in and two 4-wk washout periods. In the 2 intervention periods, 0.5 g/(kg. d) of the dietary protein was provided as either isolated soy protein (ISP) or casein, in random order. Blood and urine samples were collected at the beginning and end of each period. Data were analyzed by multiple linear regression for a repeated-measure design. ISP consumption led to changes of -9.5% in urinary albumin excretion (P < 0.0001), 0.45 in the total-to-HDL-cholesterol ratio (P < 0.05), -0.20 in the LDL-to-HDL cholesterol ratio (P < 0.05), and +4.3% in HDL cholesterol (P = 0.0040). Plasma arginine concentrations, the arginine-to-lysine ratio, and plasma isoflavone concentrations were higher after ISP consumption (P < 0.05). Urinary albumin excretion was negatively correlated with plasma total isoflavones (rho = -0.441), daidzein (rho = -0.326), and O-desmethylangolesin (rho = -0.389) (P < 0.05). The findings indicate that isolated soy protein consumption improves several markers that may be beneficial for type 2 diabetic patients with nephropathy. PMID- 15284370 TI - The fermentation of different dietary fibers is associated with fecal clostridia levels in men. AB - Only a few reports have compared the fermentation of pectin and cellulose using the hydrogen-breath test, and no studies have examined the relation between the hydrogen breathing pattern and colonic microflora. Using breath-hydrogen measurements, we investigated whether different dietary fibers (DFs) were fermented differently and whether there were individual differences after ingestion of the same DF; we also examined the relation between individual fecal microflora and the fermentation of DF. Results of hydrogen tests in 14 men were compared after they had ingested 20 g of pectin, 20 g of cellulose, or 6 g of lactulose (a DF-like substance). We examined the relation between the breath hydrogen results and the subjects' fecal microflora. We defined significant fermentation (i.e., positive cases) as a continuous rise in hydrogen in the expiratory air of >19 ppm. The subjects were divided into 3 groups according to their hydrogen breath test pattern, i.e., positive for lactulose and pectin (Group LP, n = 4); positive for lactulose alone (Group L, n = 7); and negative for pectin, cellulose, and lactulose (Group N, n = 3). Individual differences were noted in subjects from Group LP and Group L. The detection frequency of lecithinase-negative clostridia was higher in Group LP than in the other groups (P < 0.05), and the detection frequency and the number of lecithinase-positive clostridia were higher in Groups LP and L than in Group N (P < 0.05). These findings suggest that the Clostridium species are associated with hydrogen production. The hydrogen breath test results of DFs depend on both the type of DF and the individual colonic microflora. The amount and constitution of colonic microflora might be predicted by the hydrogen-breath test using different DFs. PMID- 15284371 TI - Lutein bioavailability is higher from lutein-enriched eggs than from supplements and spinach in men. AB - Lutein may be protective against diseases such as age-related macular degeneration (ARMD). At present, data regarding bioavailability of lutein from various sources are insufficient. Healthy men (n = 10) participated in an intervention study with a crossover design. After a 2-wk washout period during which they consumed a low-carotenoid diet, the men were administered 1 of 4 lutein doses (lutein supplement, lutein ester supplement, spinach, and lutein enriched egg) for 9 d. All lutein doses provided 6 mg lutein except for the lutein ester dose, which provided 5.5 mg lutein equivalents. Serum samples were collected from fasting subjects on d -14, 1 (baseline), 2, 3, and 10 and analyzed for changes in lutein concentration. Triacylglycerol-rich lipoproteins (TRL) were separated from postprandial blood samples (0-24 h) after the first lutein dose and analyzed for lutein concentration. Subjects completed all 4 treatments of the study in random order. Results from repeated-measures 1-way ANOVA showed that the baseline and dose-adjusted lutein response in serum was significantly higher after egg consumption than after lutein, lutein ester, and spinach consumption on d 10. There was no significant difference in TRL response. In conclusion, the lutein bioavailability from egg is higher than that from other sources such as lutein, lutein ester supplements, and spinach. The lutein bioavailability from lutein, lutein ester supplements, and spinach did not differ. This finding may have implications for dietary recommendations that may decrease the risk of certain diseases, e.g., ARMD. PMID- 15284372 TI - Meal replacements are as effective as structured weight-loss diets for treating obesity in adults with features of metabolic syndrome. AB - Meal replacements are widely used as a weight-loss strategy; however, their effectiveness outside controlled clinical trial environments is unknown. We compared meal replacements with a structured weight-reduction diet in overweight/obese Australians with raised triglycerides. In a randomized parallel design, 2 groups [meal replacement (MR) and control (C)] of 66 matched subjects underwent a 6000 kJ intervention for 3 mo (stage 1) and a further 3 mo (stage 2). The groups were provided oral and written information. The C group was supplied with shopping vouchers and followed a low fat/energy diet. The MR group was supplied with Slim-Fast trade mark products for 2 meals (1800 kJ) and consumed a low-fat evening meal. Clients were weighed every 2 wk and received structured supervision without professional dietary input, with compliance assessed by 3-d weighed food records. Blood biomarkers were used to assess fruit/vegetable intake and a questionnaire was used to assess attitudes to treatment. Fifty-five subjects completed stage 1 (withdrawals: 7 in the MR group, 4 in the C group) and 42 subjects completed stage 2. Weight loss was 6.0 +/- 4.2 kg (6.3%) for the MR group and 6.6 +/- 3.4 kg (6.9%) for the C group at 3 mo, and 9.0 +/- 6.9 kg (9.4%) for the MR group and 9.2 +/- 5.1 kg (9.3%) for the C group at 6 mo (different over time within but not between treatments). Serum folate and plasma beta-carotene were higher in the MR group. Plasma homocysteine fell in both groups (P < 0.005). Dietary fiber intake was higher in the C group (P < 0.02) and calcium was higher in the MR group (P < 0.001). We concluded meal replacement is equally effective for losing weight compared with conventional but structured weight-loss diets. Dietary compliance and convenience were viewed more favorably by participants who consumed meal replacements than by those in a conventional weight-loss program. PMID- 15284374 TI - Dairy intake is associated with lower body fat and soda intake with greater weight in adolescent girls. AB - Body fat and weight of 9- to 14-y-old girls (n = 323) from Kaiser Permanente were studied in relation to age, ethnicity, and physical activity. Mean age, calcium intake, weight, and iliac skinfold thickness were 11.5 +/- 1.4 y, 736.5 +/- 370.7 mg/d, 44.6 +/- 13.0 kg, and 12.4 +/- 6.1 mm, respectively. Multiple regression with age, ethnicity, height, Tanner breast stage, physical activity, energy, soda, and calcium intake explained 17% of the variation in iliac skinfold thickness. Calcium intake, age, and physical activity were significantly negatively associated with iliac skinfold thickness whereas height, Tanner breast stage, and Pacific Islander ethnicity were significantly positively associated (P < 0.0001, R(2) = 0.165). Substituting total calcium with dairy and nondairy calcium in separate models accounted for 16 and 15% of the variance, respectively (P < 0.0001, both models); 1 mg of total and dairy calcium was significantly associated with 0.0025 mm (P = 0.01) and 0.0026 mm (P = 0.02) lower iliac skinfold thickness. Thus, 1 milk serving was associated with 0.78 mm iliac skinfold thickness. The interaction of Asian ethnicity and dairy intake was significant (P = 0.027). Nondairy calcium was not associated with weight or iliac skinfold thickness. Soda intake was significantly positively associated with weight in both models (P = 0.01, both models). Decreasing soda and increasing dairy consumption among Asians may help maintain body fat and weight during adolescence. PMID- 15284373 TI - Short-term low-protein intake does not increase serum parathyroid hormone concentration in humans. AB - We investigated whether inadequate dietary protein would result in increased serum parathyroid hormone (PTH) concentration, consistent with secondary hyperparathyroidism. Data from 2 controlled feeding studies were utilized. In study 1, 26 healthy women (15 young, 21-46 y, and 11 elderly, 70-81 y) consumed for 12 d each in separate trials 3 levels of protein, 1.00, 0.75, and 0.50 g protein/(kg. d). Blood was drawn from fasting subjects on d 12 of each trial. In study 2, 24 persons (54-80 y) were fed diets with either 1.20 g protein/(kg. d) for 2 wk (HPro, n = 11, 6 men, 5 women) or 1.2 g protein/(kg. d) for 1 wk and then 0.50 g protein/(kg. d) for a 2nd week (IPro, n = 13, 6 men, 7 women). Blood was obtained from fasting subjects after wk 1 and 2. Consistent with altered protein metabolism, urinary total nitrogen excretion and blood urea nitrogen fell progressively with decreasing protein intake in study 1; in study 2, the values decreased from wk 1 to 2 in the IPro group only. Serum intact PTH concentrations did not differ among the 3 protein intakes in study 1, or between the HPro and IPro groups in study 2. These findings do not support the hypothesis that the short-term ingestion of inadequate dietary protein increases serum PTH concentration. PMID- 15284375 TI - Differential response patterns affect food-security prevalence estimates for households with and without children. AB - To produce national prevalence estimates for "food insecurity" and "food insecurity with hunger," the USDA uses a battery of 18 survey items about symptoms of food-related hardship. Ten items refer to adults in the household, while 8 items refer to children in the household and hence are not asked of households without children. To equate food-security status in households with and without children, the USDA uses a statistical model from item response theory, known as the Rasch model. This model requires an assumption that adult referenced items have the same severity calibrations for all households, including households with and without children. However, empirical estimates from the 2000 Current Population Survey showed significantly different severity calibrations for households with and without children. These differences have implications for observable response patterns. Holding constant the number of affirmative responses to adult-referenced items, households with children were more likely to respond that they "worried food would run out," and households without children were more likely to be unable to afford "balanced meals." In light of such differences, the Rasch model cannot be used to equate the food security status of households with and without children. One potential solution would be to estimate household food security by using the same battery of adult referenced survey items for all households. PMID- 15284376 TI - Daily iron alone but not in combination with multimicronutrients increases plasma ferritin concentrations in indonesian infants with inflammation. AB - Iron deficiency is a public health problem in infancy. We assessed the efficacy of iron supplements in infants with inflammation on iron status and subsequent inflammation. This was a prospective, nested, case-control study of 6- to 12-mo old infants participating in the International Research on Infant Supplementation study, Indonesia. Cases (n = 46) were selected on the basis of their inflammation status at baseline, C-reactive protein (>5 mg/L) or alpha-1 acid glycoprotein (>1 g/L); there were 44 controls without inflammation. Infants received 10 mg/d of elemental iron alone or in combination with multimicronutrients, or placebo. Blood samples were collected at baseline and at 6 mo for determinations of plasma ferritin, zinc, copper, retinol, beta-carotene, alpha-tocopherol, and inflammation status. Data on breast-feeding and acute respiratory infections (ARI) were collected daily. At baseline, 33% of infants had iron deficiency, and those with inflammation had lower retinol, beta-carotene, higher concentrations of copper and higher rates of ARI compared with controls. After 6 mo, compared with infants given placebo, ferritin concentration increased significantly in infants administered iron alone independently of inflammation status at baseline or at the end of the study. In those given multimicronutrients with iron, ferritin increased significantly in infants who did not have inflammation at baseline or at the end of the study compared with those given placebo. Consequently, iron alone resolved iron deficiency, whereas multimicronutrients reduced the deterioration of iron stores compared with placebo (chi(2), P < 0.05), without enhancing inflammation. Iron alone is recommended in populations in which iron deficiency is a public health problem despite the presence of inflammation in infants who are still breast-feeding. PMID- 15284377 TI - An adapted version of the U.S. Department of Agriculture Food Insecurity module is a valid tool for assessing household food insecurity in Campinas, Brazil. AB - Until recently, Brazil did not have a national instrument with which to assess household food insecurity (FI). The objectives of this study were as follows: 1) to describe the process of adaptation and validation of the 15-item USDA FI module, and 2) to assess its validity in the city of Campinas. The USDA scale was translated into Portuguese and subsequently tested for content and face validity through content expert and focus groups made up of community members. This was followed by a quantitative validation based on a convenience (n = 125) and a representative (n = 847) sample. Key adaptations involved replacing the term "balanced meal" with "healthy and varied diet," to construct items as questions rather than statements, and to ensure that respondents understood that information would not be used to determine program eligibility. Chronbach's alpha was 0.91 and the scale item response curves were parallel across the 4 household income strata. FI severity level was strongly associated in a dose-response manner (P < 0.001) with income strata and the probability of daily intake of fruits, vegetables, meat/fish, and dairy. These findings were replicated in the 2 independent survey samples. Results indicate that the adapted version of the USDA food insecurity module is valid for the population of Campinas. This validation methodology has now been replicated in urban and/or rural areas of 4 additional states with similar results. Thus, Brazil now has a household food insecurity instrument that can be used to set national goals, to follow progress, and to evaluate its national hunger and poverty eradication programs. PMID- 15284379 TI - Various nondigestible saccharides open a paracellular calcium transport pathway with the induction of intracellular calcium signaling in human intestinal Caco-2 cells. AB - Ingestion of soluble nondigestible saccharides increases calcium absorption, and it is suggested that paracellular calcium transport contributes to this effect. However, cellular mechanisms and the contribution of active transport have not been clarified. This study examined the effects of 4 nondigestible saccharides, difructose anhydride (DFA) III, DFAIV, fructooligosaccharides, and raffinose, on active and passive calcium transport, permeability of paracellular pathways, and intracellular calcium signaling in a human intestinal Caco-2 cell monolayer. Net, active, and passive calcium transport were evaluated using (45)Ca. Transepithelial electrical resistance (TEER) and transport of lucifer yellow were measured as indicators of paracellular passage in differentiated Caco-2 cell monolayers incubated with 0-100 mmol/L of the various saccharides. The changes in intracellular calcium ion concentrations ([Ca(2+)](i)) were measured by fura-2 loading before and after the addition of each saccharide (50 or 100 mmol/L). The addition of 100 mmol/L of each saccharide to the apical medium of the Caco-2 cells enhanced net calcium transport without any changes in active calcium transport. Relative TEER was dose dependently and reversibly decreased by the addition of saccharides, and the decreases in TEER were highly correlated with net calcium transport (P < 0.001). Basolateral application of the saccharides had a slight or no effect on indicators of the paracellular pathway. Each saccharide caused an immediate and dose-dependent rise in [Ca(2+)](i) in the cells. The 4 nondigestible saccharides increased net calcium transport in the cells via the paracellular route through tight junctions. The rise in [Ca(2+)](i) induced by these saccharides may be involved in the opening of tight junctions. PMID- 15284378 TI - Estrogen prevents the reduction in fractional calcium absorption due to energy restriction in mature rats. AB - Weight reduction is a risk factor for bone loss. We previously showed that energy restriction is associated with a decrease in calcium (Ca) absorption and decreased estrogenic activity (EA). We hypothesized that this hypoestrogenic status may be the cause of the decrease in Ca absorption and that estrogen replacement during energy restriction would prevent it. Six-month-old rats were ovariectomized and implanted subcutaneously with 17beta-estradiol (E(2)) pellets to maintain levels within the physiological range. After 3 wk, rats ate ad libitum [control (CTL) group, n = 12] or were 40% energy restricted (EnR group, n = 12) for 10 wk. At the end of this study, rats were divided into 2 groups according to their uterine weight: those with higher EA and those with lower EA. Whereas CTL rats gained approximately 46% weight from baseline, EnR rats maintained their weight throughout the study. Energy restriction was associated with lower Ca absorption (5-d measurement, (45)Ca radioisotope) and Ca balance in lower EA but not higher EA rats. Similarly, Ca absorption was correlated with both serum E(2) (r = 0.68, P < 0.05) and body weight (r = 0.72, P < 0.05) in rats with lower EA but not in those with higher EA. Finally, 24-h corticosterone excretion was higher in EnR than in CTL rats, a response that was blunted in the higher EA rats. Our findings suggest that decreases in estrogen and hyperadrenocorticism with energy restriction play an important role in the regulation of Ca absorption and balance. PMID- 15284380 TI - Ingestion of guar gum hydrolysate, a soluble and fermentable nondigestible saccharide, improves glucose intolerance and prevents hypertriglyceridemia in rats fed fructose. AB - Fructose feeding provides a dietary model of insulin resistance accompanied by hypertriglyceridemia. We examined the effects of guar gum hydrolysate (GGH), a soluble and fermentable nondigestible saccharide with low viscosity, on glucose intolerance and hypertriglyceridemia in rats fed high-fructose diets. Rats were fed either a dextrin-based or a fructose-based diet with or without GGH (75 g/kg) for 30 d. Oral glucose tolerance tests (OGTTs) were performed 0, 14, and 28 d after feeding. High-fructose feeding negatively affected glucose tolerance on d 14 and 28. The addition of GGH to the diets improved glucose intolerance on d 28. Fructose feeding induced hyperinsulinemia after an oral glucose load; this was also improved by GGH on d 28. The glycogen concentration in the gastrocnemius muscles of rats was lowered by dietary fructose, and GGH supplementation abolished this decrease. Triglycerides in the plasma and livers of rats fed fructose diets were elevated, and the increases were ameliorated by supplemental GGH. Regardless of the type of carbohydrate, GGH enlarged the cecum and increased the cecal SCFA pools. In conclusion, supplemental feeding of GGH to rats improved the glucose intolerance and hypertriglyceridemia induced by a high-fructose diet. Possible mediators of these beneficial effects of GGH are the SCFAs produced by microbial fermentation of GGH in the large intestine. PMID- 15284382 TI - Copper deficiency reduces iron absorption and biological half-life in male rats. AB - Dietary copper deficiency (CuD) in rats leads to iron (Fe) deficiency anemia. Is this because CuD reduces Fe absorption? Fe absorption in CuD rats was determined by feeding diets labeled with (59)Fe and using whole-body counting (WBC) to assess the amount retained over time. Two groups, each with 45 male weanling rats, were fed an AIN-93G diet low in Cu (<0.3 mg/kg; CuD) or one containing adequate Cu (5.0 mg/kg; CuA). At intervals over the next 42 d, 5 rats per group were killed and blood was drawn to determine hematocrit, hemoglobin, and other indicators of Fe status. At d 7 and 25, 5 rats per group were fed 1.0 g of their respective diets that had been labeled with (59)Fe. Retained (59)Fe was monitored for 10 d by WBC; then rats were killed and (59)Fe was measured in various organs. Signs of Fe deficiency, such as low hemoglobin, hematocrit, and RBC count, were evident in CuD rats by d 14. At d 7, CuD rats absorbed 90% as much Fe as CuA rats (P > 0.20), but at d 25, CuD rats absorbed only 50% as much as CuA rats (P < 0.001). In the study beginning at d 7, the biological half-life (BHL) of (59)Fe in CuD rats was less (P < 0.02) than that in CuA rats [geometric mean (-SEM, +SEM); 75(62,91) d vs. 175(156,195) d]. In the study beginning at d 25, the BHL was again less (P < 0.02) in the CuD rats than in the CuA rats [33(23,49) d for CuD and 157(148,166) d for CuA]. Apparently, the route of Fe loss in the CuD rats was through the gut. At d 16 and 34, CuD rats lost 4 to 5 times more (P < 0.01) (59)Fe in the feces in a 24-h period than the CuA rats. Also, (59)Fe in the duodenal mucosa of CuD rats was approximately 100% higher (P < 0.01) than in CuA rats. These findings suggest that Fe deficiency anemia in CuD male rats is caused at least in part by reductions in Fe absorption and retention. PMID- 15284381 TI - Piperine enhances the bioavailability of the tea polyphenol (-)-epigallocatechin 3-gallate in mice. AB - (-)-Epigallocatechin-3-gallate (EGCG), from green tea (Camellia sinensis), has demonstrated chemopreventive activity in animal models of carcinogenesis. Previously, we reported the bioavailability of EGCG in rats (1.6%) and mice (26.5%). Here, we report that cotreatment with a second dietary component, piperine (from black pepper), enhanced the bioavailability of EGCG in mice. Intragastric coadministration of 163.8 micromol/kg EGCG and 70.2 micromol/kg piperine to male CF-1 mice increased the plasma C(max) and area under the curve (AUC) by 1.3-fold compared to mice treated with EGCG only. Piperine appeared to increase EGCG bioavailability by inhibiting glucuronidation and gastrointestinal transit. Piperine (100 micromol/L) inhibited EGCG glucuronidation in mouse small intestine (by 40%) but not in hepatic microsomes. Piperine (20 micromol/L) also inhibited production of EGCG-3"-glucuronide in human HT-29 colon adenocarcinoma cells. Small intestinal EGCG levels in CF-1 mice following treatment with EGCG alone had a C(max) = 37.50 +/- 22.50 nmol/g at 60 min that then decreased to 5.14 +/- 1.65 nmol/g at 90 min; however, cotreatment with piperine resulted in a C(max) = 31.60 +/- 15.08 nmol/g at 90 min, and levels were maintained above 20 nmol/g until 180 min. This resulted in a significant increase in the small intestine EGCG AUC (4621.80 +/- 1958.72 vs. 1686.50 +/- 757.07 (nmol/g.min)). EGCG appearance in the colon and the feces of piperine-cotreated mice was slower than in mice treated with EGCG alone. The present study demonstrates the modulation of the EGCG bioavailablity by a second dietary component and illustrates a mechanism for interactions between dietary chemicals. PMID- 15284383 TI - Vitamin A deficiency impairs fetal islet development and causes subsequent glucose intolerance in adult rats. AB - To determine the role of vitamin A in fetal islet development, beta- and alpha cell mass, apoptosis, and alpha- and beta-cell replication were measured in rats using a model of marginal vitamin A deficiency. Female rats before and during pregnancy and their offspring postweaning were fed a diet containing retinol as retinyl palmitate at a low marginal (LM, 0.25 mg/kg diet) or a sufficient (SUFF, 4.0 mg/kg diet) level. Fetal islet size, replication, apoptosis, and offspring glucose tolerance were examined. Both beta-cell area and number per islet were reduced approximately 50% in fetuses from dams fed an LM vitamin A diet compared with those from dams fed the SUFF vitamin A diet. The alpha-cell area and number per fetal islet were not affected by vitamin A deficiency. Apoptosis was not increased. The percentage of newly replicated beta-cells in the LM fetal pancreas was 42% less than that of SUFF offspring, but alpha-cell replication was not affected. To determine whether this decrease in beta-cell area affected adult glucose tolerance and insulin secretion, 65-d-old offspring were subject to glucose tolerance tests. LM rats had a 55% lower plasma insulin level and a 76% higher serum glucose than SUFF rats. The same pattern could be seen in 35-d-old rats. These findings show that vitamin A deficiency decreases beta-cell mass and this reduction can be attributed to a reduced rate of fetal beta-cell replication in LM offspring. This may contribute to impaired glucose tolerance later in adult life. PMID- 15284384 TI - Lactobacillus GG bacteria ameliorate arthritis in Lewis rats. AB - Probiotic bacteria have beneficial effects in infectious and inflammatory diseases, principally in bowel disorders. In the case of chronic progressive autoimmune arthritides, a major goal of treatment is to reduce inflammation. We hypothesized that probiotic bacteria would ameliorate inflammation found in arthritis models. To assess this effect, Lewis rats were injected with 50 microg bovine alpha-tropomyosin (TRM) or complete Freund's adjuvant (CFA) to induce tropomyosin arthritis (TA) or adjuvant arthritis (AA), respectively. In both models, the rats were divided into 6 groups and fed 0.5 mL/d of the following suspensions: 1) heat-killed Lactobacillus GG (LGG) bacteria; 2) live LGG, both 10(11) colony-forming units (cfu)/L; 3) sterilized milk; 4) plain yogurt; 5) yogurt containing 10(11) cfu/L LGG; or 6) sterilized water. In the disease prevention experiments, feeding started 1 wk before or after disease induction. In the therapeutic experiments, feeding was initiated at the onset of clinical arthritis. In all experiments, there were significant interactions between time and treatment (P < 0.001), except for milk, which had no effect in the therapeutic experiment. Histologically, rats fed yogurt containing LGG had a milder inflammation in all experiments (P < 0.05), whereas rats fed plain yogurt exhibited a moderate inflammatory score only in the prevention experiments. Anti TRM antibody titers were not affected by any of the treatments in any of the experiments. Ingestion of live or heat-killed human LGG had a clinically beneficial effect on experimental arthritis. Our observation of the remarkable preventive and curative effect on arthritis using commercial yogurts containing lactobacilli, especially LGG, suggests the need for investigation of these agents in arthritic patients. PMID- 15284385 TI - Biotin deficiency blocks thymocyte maturation, accelerates thymus involution, and decreases nose-rump length in mice. AB - Biotin deficiency in experimental animals causes low body weight as well as several phenomena suggestive of an altered immune system. We reported previously that chronic biotin deficiency in mice decreases body weight and alters the number and proportion of lymphocyte subpopulations in the spleen. To further characterize the effects of biotin deficiency, we studied in detail the maturation of thymocytes and the status of biotin in the thymus, as well as the body length of biotin-deficient mice. Male Balb/cAnN mice were fed for up to 20 wk either standard control diet, a biotin-deficient diet, or a biotin-sufficient diet. At different times, nose-rump length, weight of the thymus, spleen and liver, total number of cells in the spleen and thymus, pyruvate carboxylase (PC) and propionyl CoA carboxylase (PCC) activity in thymus cells, and the proportion of distinct thymocyte subsets were determined. These variables did not differ between mice fed the control and biotin-sufficient diets. In contrast, biotin deficient mice differed from biotin-sufficient mice in all of the analyzed variables. PC and PCC specific activities of thymocytes of mice fed the biotin depleting diet decreased during the first 4 wk by 84.5%. The maturation of thymocytes in biotin-deficient mice was arrested at the double-negative stage. Our results suggest that biotin deficiency in mice causes an accelerated involution of the thymus and decreases nose-rump length, but these effects do not correlate in magnitude or in temporality with the sharp decrease in the activity of the biotin-dependent carboxylases. As such, the possibility that the aforementioned effects are not related directly to the prosthetic function of biotin should be considered. PMID- 15284386 TI - Dietary polyunsaturated fatty acids modulate in vivo, antigen-driven CD4+ T-cell proliferation in mice. AB - Our objective was to determine whether dietary lipids affect in vivo, antigen driven, proliferation of naive CD4(+) T lymphocytes. To accomplish this, we adoptively transferred lymphocytes from T-cell receptor (TCR) transgenic DO11.10 (i.e., donor) mice into syngeneic, nontransgenic BALB/c (i.e., recipient) mice. Before adoptive transfer, recipient mice were fed for 4 wk AIN93G-type diets that differed only in fat source: lard, low in PUFA, fish oil, rich in (n-3) PUFA, and soybean oil, rich in (n-6) PUFA. One week after transfer, recipient mice were immunized with antigen (i.e., ovalbumin), and expansion of CD4(+) T(DO11.10) cells in the spleen and draining lymph nodes (LN) was measured by flow cytometry. Five days postimmunization (p.i.), at the peak of expansion, CD4(+) T(DO11.10) cells in the draining LN and spleen were 5- to 10-fold higher than in unimmunized mice, then quickly declined during the contraction phase (i.e., 7 and 10 d p.i.). Recipients fed the (n-6) PUFA rich diet had approximately 25% greater in vivo expansion of CD4(+) T(DO11.10) cells than lard- and fish oil-fed recipient mice at 5 d p.i. (P < 0.05). However, at 7 and 10 d p.i., CD4(+) T(DO11.10) cells in the draining lymph nodes did not differ between groups, nor in the spleen at 5, 7, and 10 d p.i. In summary, we are the first to demonstrate that dietary PUFAs affect antigen-driven expansion of naive CD4(+) T cells in vivo. Surprisingly, (n 3) PUFA consumption did not reduce CD4(+) T-cell expansion. PMID- 15284387 TI - Abnormal motor function persists following recovery from perinatal copper deficiency in rats. AB - What are the biochemical and behavioral consequences of perinatal copper deficiency? Pregnant Holtzman rats were fed a modified AIN-76A diet low in copper (0.34 mg Cu/kg and 42 mg Fe/kg) starting on gestation d 7. Seven rats received copper in their drinking water (20 mg Cu/L) (+Cu) and 7 drank deionized water ( Cu). Treatments did not affect litter size or pregnancy outcome. Compared with +Cu dams and a sample of +Cu male weanling [postnatal day (P)21] offspring, -Cu rats exhibited signs consistent with copper deficiency. P21 males were switched to a nonpurified copper-adequate diet and sampled biochemically after 3 mo and behaviorally after 3 and 6 mo of repletion (CuR). Compared with controls, CuR rats had lower brain copper and iron levels 3 and 6 mo after repletion; other biochemical differences were not detected. Behavioral assessments after 5 mo of repletion indicated a persistent impairment in motor function of CuR compared with control rats as evaluated by the accelerating rotorod procedure. These results suggest that permanent impairment to motor function can persist after long-term recovery from perinatal copper deficiency. PMID- 15284388 TI - Conjugated linoleic acid isomers and trans fatty acids inhibit fatty acid transport in hepatoma 7288CTC and inguinal fat pads in Buffalo rats. AB - Conjugated linoleic acid (CLA) and some trans fatty acids (FA) decrease tumor growth and alter tumor and host lipid uptake and storage. The goal of this study was to test the hypothesis that the acute inhibitory effects of CLA isomers and trans FAs on FA transport in tumors and white adipose tissue are mediated via an inhibitory G-protein coupled (GPC), FFA receptor (FFAR). Experiments were performed in hepatoma 7288CTC and inguinal fat pads in Buffalo rats during perfusion in situ. CLA isomers and trans FAs (0.03-0.4 mmol/L, in plasma) were added to the arterial blood, and FA uptake or release was measured by arterial minus venous difference. In hepatoma 7288CTC, the CLA isomers, t10,c12-CLA > (+/ )-9-HODE [13-(S)-hydroxyoctadecadienoic acid] > t9,t11-CLA, and the trans FAs, linolelaidic = vaccenic > elaidic, decreased cAMP content and inhibited FA uptake, 13(S)-HODE release, extracellular signal-regulated kinase p44/p42 phosphorylation, and [(3)H]thymidine incorporation. Other CLA isomers, c9,t11 CLA, 13-(S)-HODE, c9,c11-CLA, and c11,t13-CLA, had no effect. In inguinal fat pads, FA transport was inhibited by t10,c12-CLA = linolelaidic acid > trans vaccenic acid, whereas c9,t11-CLA had no effect. In both hepatoma 7288CTC and inguinal fat pad, addition of either pertussis toxin or 8-Br-cAMP to the arterial blood reversed the inhibitions of FA transport. These results support the idea that an inhibitory GPC FFAR reduces cAMP and controls FA transport by CLA isomers and trans FAs. Ligand activity is conferred by the presence of a trans double bond proximal to the carboxyl group. PMID- 15284389 TI - Plasma phytoestrogens are not altered by probiotic consumption in postmenopausal women with and without a history of breast cancer. AB - Soy phytoestrogens were suggested to reduce the risk of a number of diseases including breast cancer. Given that these compounds are metabolized by bacteria, alteration of intestinal bacteria and enzymes may affect phytoestrogen metabolism. We hypothesized that probiotics, when consumed with soy protein, would increase plasma isoflavones, as well as equol producer frequency, in postmenopausal women. We further hypothesized that these effects would differ between women who have had breast cancer and women who have not. To test these hypotheses, 20 breast cancer survivors and 20 controls completed four 6-wk treatments in a randomized, crossover design: supplementation with soy protein (S) (26.6 +/- 4.5 g protein, 44.4 +/- 7.5 mg isoflavones/d); soy + probiotics (S+P) (10(9) colony-forming units Lactobacillus acidophilus DDS+1 and Bifidobacterium longum, 15-30 mg fructooligosaccharide/d); milk protein (M) (26.6 +/- 4.5 g protein/d); and milk + probiotics (M+P). Plasma phytoestrogen concentrations did not differ between controls and survivors, although genistein tended to be lower in survivors at baseline (P = 0.15), and during soy (P = 0.16) and milk protein (P = 0.16) consumption. As expected, soy consumption increased plasma phytoestrogen concentrations (P < 0.0001). Plasma phytoestrogen concentrations and the number of equol producers did not differ between the S and S+P diets. At the same time, plasma equol concentrations as well as urinary equol excretion in 2 subjects were more than 7-fold different between the 2 diets. These results indicate that this particular probiotic supplement does not generally affect plasma isoflavones, although the large differences between plasma and urinary equol in some subjects suggest that equol producer status may be modifiable in some individuals. PMID- 15284390 TI - Dietary isothiocyanates inhibit the growth of human bladder carcinoma cells. AB - Many isothiocyanates (ITCs), some of which are abundant in cruciferous vegetables, have been repeatedly shown to inhibit carcinogenesis in a variety of rodent organs. However, several naturally occurring ITCs also promoted bladder tumorigenesis in rodents, raising the question of whether ITCs behave differently in bladder cells. Alternatively, the observed carcinogenic effects of ITCs may result from prolonged exposure of the bladder epithelium, where the tumors originate, to high concentrations of electrophilic ITCs in the urine. Ingested ITCs are almost exclusively excreted and highly concentrated in the urine as N acetylcysteine conjugates (NAC-ITC). While several NAC-ITCs also are known anticarcinogens, they are unstable and readily dissociate into parent ITCs. In this study, ITCs, including those that have carcinogenic potential in the rodent bladders, induced apoptosis and/or arrested cell-cycle progression in 2 human bladder carcinoma lines (UM-UC-3 and T24) at 7.5-30 micromol/L. Multiple caspases, including caspase-9, -8, and -3, as well as poly(ADP-ribose)polymerase, were cleaved upon ITC exposure. The ITCs blocked cell-cycle progression at the G(2)/M and/or S phases in these cells and downregulated several cell-cycle regulators. However, further increases in ITC concentrations abolished their activities, described above. These findings show that urinary ITC concentrations may need to be maintained at low micromolar concentrations for bladder cancer prevention. PMID- 15284391 TI - Development of a food database of nitrosamines, heterocyclic amines, and polycyclic aromatic hydrocarbons. AB - Some nitrosocompounds that are formed during food preservation, as well as polycyclic aromatic hydrocarbons (PAH) and heterocyclic amines (HA) formed during cooking, may have carcinogenic activity. An accurate assessment of dietary intake of such compounds is difficult, mainly because they are not naturally present in foods, and they are not included in standard food composition tables. Our objective was to develop a food composition database of nitrates, nitrites, nitrosamines, HA, and PAH. We conducted a literature search on the food content of these compounds using the Medline and EMBASE databases. We gathered the following information: 1) Food information: name, cooking methods, preservation methods, cooking doneness, temperature, and time; 2) compound information: type, quantity, value type, analytic method, and sampling methods; and 3) publication information: year, author, and country. We developed a table that includes 207 food items with information concerning the concentration of nitrites, nitrates, and nitrosamines, 297 food items with information about HA concentration and 313 food items with information about PAH. The database is based on 139 references from 23 different countries. It is arranged according to compounds and food groups to facilitate its practical use. The potential limitations are due to the quality of the information we could obtain through Medline and EMBASE databases. This database will allow investigators to quantify dietary exposure to several potential carcinogens, and to analyze their relation to the risk of cancer. PMID- 15284392 TI - The stratum corneum barrier: the final frontier. AB - The stratum corneum (SC) is the differentiated end product of the mammalian epidermis. It is vital to constancy of the milieu interieur (the environment within) because it prevents water loss and the penetration by potentially toxic xenobiotics, damaging radiation, and pathogenic microbes. The intercorneocyte space contains complex nonpolar lipids that constitute the water barrier. The formation of the SC in the process of keratinization is complex providing multiple opportunities for disorders to arise. The final act of keratinization is desquamation and for this to occur the controlled release of single corneocytes is required in which proteases play an important role. Tests and techniques are described that measure the structure and function of the SC. PMID- 15284393 TI - Bacteria in the gut: friends and foes and how to alter the balance. AB - The activities of the bacteria resident in the colon of companion animals can have an impact upon the health of the host. Our understanding of this microbial ecosystem is presently increasing due to the development of DNA-based microbiological tools that allow identification and enumeration of nonculturable microorganisms. These techniques are changing our view of the bacteria that live in the gut, and they are facilitating dietary-intervention approaches to modulate the colonic ecosystem. This is generally achieved by the feeding of either live bacteria (probiotics) or nondigestible oligosaccharides (prebiotics) that selectively feed the indigenous probiotics. Feeding studies with a Lactobacillus acidophilus probiotic have shown positive effects on carriage of Clostridium spp. in canines and on recovery from Campylobacter spp. infection in felines. Immune function was improved in both species. Prebiotic feeding studies with lactosucrose and fructo-oligosaccharides in both cats and dogs have shown positive effects on the microflora balance. Recently synbiotic forms (a probiotic together with a prebiotic) targeted at canines have been developed that show promise as dietary-intervention tools. PMID- 15284394 TI - Body-weight changes during growth in puppies of different breeds. PMID- 15284395 TI - Relationship between electrolyte apparent absorption and fecal quality in adult dogs differing in body size. PMID- 15284396 TI - Maternal diet alpha-linolenic acid during gestation and lactation does not increase docosahexaenoic acid in canine milk. PMID- 15284397 TI - Gastric emptying rate is inversely related to body weight in dog breeds of different sizes. PMID- 15284398 TI - Excess dietary lysine does not cause lysine-arginine antagonism in adult cats. PMID- 15284399 TI - Cats select for adequate methionine but not threonine. PMID- 15284400 TI - Nutritional status of canine and feline patients admitted to a referral veterinary internal medicine service. PMID- 15284401 TI - UV light, temperature, and humidity effects on white hair color in dogs. PMID- 15284402 TI - Canine atopic dermatitis: new targets, new therapies. AB - Atopic dermatitis is a common allergic skin disease of complex etiopathogenesis in both humans and dogs. Immediate-type hypersensitivity to environmental allergens that arises as a result of environmental and genetic factors is a major part of the pathogenesis in most but not all patients. Alterations in epidermal barrier function, priming of cutaneous antigen-presenting cells with IgE, intrinsic keratinocyte defects, and even development of autoimmunity are also factors that contribute to the primary disease. Secondary factors, especially infections with Staphylococcus and yeast organisms, strongly influence the course of this skin disease. The relatively recent understanding of the complexities of atopic dermatitis has resulted in changes in diagnostic and therapeutic strategies for the disease. We now know that the best therapeutic approach is to use combinations of multiple modalities individualized for each patient over the course of his or her lifetime. PMID- 15284403 TI - Diagnosis of adverse reactions to food in dogs: efficacy of a soy-isolate hydrolyzate-based diet. PMID- 15284404 TI - Antioxidant supplementation in horses affected by recurrent airway obstruction. PMID- 15284405 TI - Carbohydrate malabsorption is a feature of feline inflammatory bowel disease but does not increase clinical gastrointestinal signs. PMID- 15284406 TI - Canine and feline diabetes mellitus: nature or nurture? AB - There is evidence for the role of genetic and environmental factors in feline and canine diabetes. Type 2 diabetes is the most common form of diabetes in cats. Evidence for genetic factors in feline diabetes includes the overrepresentation of Burmese cats with diabetes. Environmental risk factors in domestic or Burmese cats include advancing age, obesity, male gender, neutering, drug treatment, physical inactivity, and indoor confinement. High-carbohydrate diets increase blood glucose and insulin levels and may predispose cats to obesity and diabetes. Low-carbohydrate, high-protein diets may help prevent diabetes in cats at risk such as obese cats or lean cats with underlying low insulin sensitivity. Evidence exists for a genetic basis and altered immune response in the pathogenesis of canine diabetes. Seasonal effects on the incidence of diagnosis indicate that there are environmental influences on disease progression. At least 50% of diabetic dogs have type 1 diabetes based on present evidence of immune destruction of beta-cells. Epidemiological factors closely match those of the latent autoimmune diabetes of adults form of human type 1 diabetes. Extensive pancreatic damage, likely from chronic pancreatitis, causes approximately 28% of canine diabetes cases. Environmental factors such as feeding of high-fat diets are potentially associated with pancreatitis and likely play a role in the development of pancreatitis in diabetic dogs. There are no published data showing that overt type 2 diabetes occurs in dogs or that obesity is a risk factor for canine diabetes. Diabetes diagnosed in a bitch during either pregnancy or diestrus is comparable to human gestational diabetes. PMID- 15284407 TI - Use of glucagon stimulation tests to assess beta-cell function in dogs with chronic pancreatitis. PMID- 15284408 TI - High protein intake affects lean body mass but not energy expenditure in nonobese neutered cats. PMID- 15284409 TI - High-protein low-carbohydrate diets enhance weight loss in dogs. PMID- 15284410 TI - Obesity: the integrated roles of environment and genetics. AB - Obesity represents one of the most serious global health issues with approximately 310 million people presently affected. It develops because of a mismatch between energy intake and expenditure that results from behavior (feeding behavior and time spent active) and physiology (resting metabolism and expenditure when active). Both of these traits are affected by environmental and genetic factors. The dramatic increase in the numbers of obese people in Western societies reflects mostly changing environmental factors and is linked to reduced activity and perhaps also increased food intake. However, in all societies and subpopulations, there are both obese and nonobese subjects. These differences are primarily a consequence of genetic factors as is revealed by the high heritability for body mass index. Most researchers agree that energy balance and, hence, body weight are regulated phenomena. There is some disagreement about exactly how this regulation occurs. However, a common model is the "lipostatic" regulation system, whereby our energy stores generate signals that are compared with targets encoded in the brain, and differences between these drive our food intake levels, activity patterns, and resting and active metabolisms. Considerable advances were made in the last decade in understanding the molecular basis of this lipostatic system. Some obese people have high body weight because they have broken lipostats, but these are a rare minority. This suggests that for the majority of obese people, the lipostat is set at an inappropriately high level. When combined with exposure to an environment where there is ready availability of food at low energy costs to obtain it, obesity develops. The evolutionary background to how such a system might have evolved involves the evolution of social behavior, the harnessing of fire, and the development of weapons that effectively freed humans from the risks of predation. The lipostatic model not only explains why some people become obese whereas others do not, but also allows us to understand why energy-controlled diets do not work. Drug-based solutions to the obesity problem that work with the lipostat, rather than against it, are presently under development and will probably be in regular use within 5 10 y. However, several lines of evidence including genetic mapping studies of quantitative trait loci associated with obesity suggest that our present understanding of the regulatory system is still rudimentary. In particular, we know nothing about how the target body weight in the brain is encoded. As our understanding in this field advances, new drug targets are likely to emerge and allow us to treat this crippling disorder. PMID- 15284411 TI - Maternal dietary fatty acids modify canine puppy plasma lipoprotein distributions during the suckling period. PMID- 15284412 TI - Evaluation of corneometry (skin hydration) and transepidermal water-loss measurements in two canine breeds. PMID- 15284413 TI - Urinary isovalthine excretion in adult cats is not gender dependent or increased by oral leucine supplementation. PMID- 15284414 TI - A combination of aloe vera, curcumin, vitamin C, and taurine increases canine fibroblast migration and decreases tritiated water diffusion across canine keratinocytes in vitro. PMID- 15284415 TI - Changes to levels of DNA damage and apoptotic resistance in peripheral blood mononuclear cells and plasma antioxidant potential with age in labrador retriever dogs. PMID- 15284416 TI - A potential nutritional prophylactic for the reduction of feline hairball symptoms. PMID- 15284417 TI - Benefits of bovine colostrum on fecal quality in recently weaned puppies. PMID- 15284418 TI - Dietary sodium promotes increased water intake and urine volume in cats. PMID- 15284419 TI - Immunoglobulin A concentrations in adult dogs vary according to sample type and collection time and method. PMID- 15284420 TI - Application of the comet assay for investigation of oxidative DNA damage in equine peripheral blood mononuclear cells. AB - Oxidative stress occurs when antioxidant defense mechanisms are overwhelmed by free radicals and may lead to DNA damage, which has been implicated in processes such as aging and diseases such as cancer. The two main techniques presently used to quantify DNA damage are measurement of 8-hydroxydeoxyguanosine and the Comet assay (also known as single-cell gel electrophoresis). The aim of this study was to apply the comet assay to equine peripheral blood mononuclear cells (PBMCs) and identify two conditions in which we hypothesized that oxidative DNA damage would be increased in PBMCs: aging and equine recurrent airway obstruction (RAO, a condition similar to human asthma). The images obtained were similar to those previously published for humans, cats, and dogs. The optimum concentration of H(2)O(2) to estimate susceptibility to exogenous damage was 50 microM. Mean intraassay coefficients of variation were 4.7 and 9.7% for endogenous and exogenous tail-DNA quantities, respectively, and 7.3 and 8.3%, respectively, for interassay coefficients. There was no significant difference in either endogenous or exogenous percentages of tail DNA for samples collected from six ponies on three consecutive days. There was no significant difference in endogenous, exogenous, or exogenous (corrected for endogenous) oxidative DNA damage between mature and aged ponies. However, young pony foals had significantly less endogenous DNA damage than mature or aged ponies (P < 0.05). RAO-affected horses without airway inflammation (i.e., in clinical remission) had significantly greater endogenous damage compared with non-RAO-affected control animals (P = 0.009). There was a significant correlation between endogenous percentage of tail DNA in PBMCs and red blood cell hemolysate glutathione concentration (r = 0.720; P < 0.001). In conclusion, the comet assay appears to be suitable for investigating DNA damage in equine PBMCs. PMID- 15284421 TI - Nutrient digestibility of commercial dog foods using mink as a model. PMID- 15284422 TI - Fatty Acid composition in commercial dog foods. PMID- 15284423 TI - Rapid weight loss with a high-protein low-energy diet allows the recovery of ideal body composition and insulin sensitivity in obese dogs. PMID- 15284424 TI - Apparent calcium absorption in growing dogs of two different sizes. PMID- 15284425 TI - Modulation of uncoupling protein 1 and peroxisome proliferator-activated receptor gamma expression in adipose tissue in obese insulin-resistant dogs. PMID- 15284426 TI - Dietary protein source and manufacturing processes affect macronutrient digestibility, fecal consistency, and presence of fecal Clostridium perfringens in adult dogs. PMID- 15284427 TI - Urinary composition of cats is affected by the source of dietary protein. PMID- 15284428 TI - Exocrine pancreatic insufficiency and adverse reaction to food in dogs: a positive response to a high-fat, soy isolate hydrolysate-based diet. PMID- 15284429 TI - Contrast-enhanced MR imaging of the heart: overview of the literature. AB - The use of magnetic resonance (MR) imaging for cardiac diagnosis is expanding, aided by the administration of paramagnetic contrast agents for a growing number of clinical applications. This overview of the literature considers the principles and applications of cardiac MR imaging with an emphasis on the use of contrast media. Clinical applications of contrast material-enhanced MR imaging include the detection and characterization of intracardiac masses, thrombi, myocarditis, and sarcoidosis. Suspected myocardial ischemia and infarction, respectively, are diagnosed by using dynamic first-pass and delayed contrast enhancement. Promising new developments include blood pool contrast media, labeling of myocardial precursor cells, and contrast-enhanced imaging at very high fields. PMID- 15284430 TI - Coronary MR angiography with steady-state free precession: individually adapted breath-hold technique versus free-breathing technique. AB - PURPOSE: To compare image quality and coronary artery stenosis detection with breath-hold (BH) and free-breathing navigator-gated (NAV) coronary magnetic resonance (MR) angiography performed with the same imaging sequence (steady-state free precession) and identical spatial resolution in patients suspected of having coronary artery disease. MATERIALS AND METHODS: Forty consecutive patients suspected of having coronary artery disease underwent steady-state free precession MR imaging of the left or the right coronary artery twice. Correction of breathing motion was performed once with NAV and again with BH. Maximal BH duration and coronary artery rest period were individually determined, and duration of data acquisition was adapted (parallel imaging with different sensitivity encoding factors was used). Quantitative analysis of coronary MR angiography data was performed with multiplanar reformatting software to determine visual score for image quality, vessel sharpness, visible vessel length, and number of visible side branches. Diagnostic accuracy for detection of coronary stenosis of 50% or greater was determined in comparison with results of conventional invasive angiography. The two techniques were compared regarding differences in angiographic parameters with paired Student t testing. chi(2) or Fisher exact testing was used when appropriate. RESULTS: More coronary artery segments were assessable with NAV than with BH MR angiography (254 [79.4%] vs 143 [44.7%] of 320 segments). Overall sensitivity and specificity with NAV were 72% (26 of 36 segments) and 91.7% (200 of 218 segments), versus 63% (12 of 19 segments) and 82.3% (102 of 124 segments) with BH; NAV enabled correct diagnosis in 13% more segments. BH yielded nondiagnostic images in 14 patients, while NAV yielded diagnostic images in all patients. When these 14 patients were excluded, there was a significant increase in visual score for left (3.0 vs 2.4, P <.01) and right (3.3 vs 3.0, P <.05) coronary arteries and no significant difference in vessel sharpness but significant improvement in visible vessel length in left coronary artery (85.9 vs 71.4 mm, P =.003) and number of visible side branches in left (4.9 vs 3.9, P =.04) and right (2.8 vs 2.4, P =.04) coronary arteries on NAV images as compared with BH images. CONCLUSION: Free-breathing NAV was superior to BH coronary MR angiography in terms of image quality and diagnostic accuracy of stenosis detection. PMID- 15284431 TI - Pseudostenosis phenomenon at volume-rendered three-dimensional digital angiography of intracranial arteries: frequency, location, and effect on image evaluation. AB - PURPOSE: To assess the frequency, location, and effect on image interpretation of a pseudostenosis phenomenon at volume-rendered three-dimensional (3D) digital angiography for evaluation of intracranial arteries and to determine the physical characteristics of the phenomenon by using a phantom. MATERIALS AND METHODS: Results of a total of 68 volume-rendered 3D digital angiographic examinations in 56 patients with intracranial aneurysms were retrospectively evaluated in comparison with results of digital subtraction angiography regarding the appearance of a pseudostenosis phenomenon. The phenomenon was analyzed by two radiologists in consensus with regard to frequency, location, percentage stenosis, and angle between the axis of the vessel with pseudostenosis and the axis of rotational angiography. The phenomenon's effect on aneurysm evaluation was also analyzed. For assessing the physical properties of the phenomenon, a phantom study was performed with different lengths of tubing and different angles to the axis of rotational angiography. RESULTS: The pseudostenosis was observed at 23 (34%) of 68 3D digital angiographic examinations and in 53 (5%) of 1161 segments. The percentage stenosis ranged from 3% to 85% (mean, 18% +/- 16.2 [standard deviation]). The arterial segments with pseudostenosis included 15 (25%) of 61 C6 segments, 10 (16%) of 61 M1 segments, and six (10%) of 60 A1 segments. Angles between the two axes ranged from 86 degrees to 93 degrees (mean angle, 90 degrees +/- 1.6). Pseudostenosis affected delineation of the shape and size of two middle cerebral artery aneurysms. At phantom analysis, the phenomenon was most obvious at an angle of 90 degrees and with the longest phantom. CONCLUSION: The pseudostenosis phenomenon on volume-rendered 3D digital angiograms was relatively frequently observed in some segments of the intracranial arteries and affected the delineation of middle cerebral artery aneurysms. This phenomenon was associated with the angle to the axis of rotational angiography and the length of the vessel. PMID- 15284432 TI - Reduced radiation dose helical chest CT: effect on reader evaluation of structures and lung findings. AB - PURPOSE: To assess, by using computer simulation, the effect of the use of reduced computed tomographic (CT) tube current on reader evaluation of structures and lung findings on images obtained at clinically indicated chest CT examinations. MATERIALS AND METHODS: The noise level in the raw scan data of 150 clinically indicated conventional tube current (200-320-mA) chest CT examinations was modified to simulate tube current reduction to 100 and to 40 mA. A total of 450 image sets were thus available. Four radiologists blinded to the tube current used assessed the image sets in random order for 14 structures and lung findings and ranked subjective image quality by using a five-point scale (1 = nondiagnostic, 2 = inferior, 3 = adequate, 4 = good, 5 = excellent). After a 3 week interval, the 150 conventional tube current image sets were rescored so that intraobserver agreement could be assessed. The McNemar statistic was used to determine whether there were more scoring disagreements between interpretations of the conventional and those of the reduced tube current scans or between the two interpretations of the conventional tube current scans. RESULTS: When overall agreement for 14 structures and lung findings was pooled over four observers, significantly more disagreements (P <.05) were seen when scores were compared between conventional and reduced tube current scans than when scores were compared between repeated interpretations of the conventional tube current scans. There was a significant decrease (P <.05) in the subjective image quality of reduced tube current scans compared with the subjective image quality of conventional tube current scans. CONCLUSION: These data indicate that reduced tube current does affect reader evaluation of structures and lung findings and reduces a reader's subjective assessment of image quality. PMID- 15284433 TI - MR procedures: biologic effects, safety, and patient care. AB - The technology used for magnetic resonance (MR) procedures has evolved continuously during the past 20 years, yielding MR systems with stronger static magnetic fields, faster and stronger gradient magnetic fields, and more powerful radiofrequency transmission coils. Most reported cases of MR-related injuries and the few fatalities that have occurred have apparently been the result of failure to follow safety guidelines or of use of inappropriate or outdated information related to the safety aspects of biomedical implants and devices. To prevent accidents in the MR environment, therefore, it is necessary to revise information on biologic effects and safety according to changes that have occurred in MR technology and with regard to current guidelines for biomedical implants and devices. This review provides an overview of and update on MR biologic effects, discusses new or controversial MR safety topics and issues, presents evidence based guidelines to ensure safety for patients and staff, and describes safety information for various implants and devices that have recently undergone evaluation. PMID- 15284434 TI - Improved characterization of liver lesions with liver-phase uptake of liver specific microbubbles: prospective multicenter study. AB - PURPOSE: To evaluate in a prospective multicenter study whether conventional ultrasonographic (US) characterization of liver lesions can be improved by imaging during the liver-specific phase of SH U 508A uptake in the microbubble specific agent detection imaging mode. MATERIALS AND METHODS: One hundred forty two patients with liver lesions underwent conventional gray-scale and color Doppler US and SH U 508A-enhanced US. Two radiologists blindly read digital cine clips and assigned scores for confidence in diagnosis of benignancy or malignancy, diagnosis of specific lesion types, and relative difference in SH U 508A uptake between the lesion and the liver parenchyma (ie, subjective conspicuity score [SCS]). Comparisons were made to see whether the addition of agent detection imaging led to improved diagnostic performance. RESULTS: Receiver operating characteristic analysis revealed improved discrimination of benign and malignant lesions for readers 1 (P =.049) and 2 (P <.001). The number of patients with a correct diagnosis of benignancy or malignancy assigned by readers 1 and 2, respectively, improved from 114 and 113 to 125 and 128 with agent detection imaging (reader 1: P =.027; reader 2: P =.008; McNemar test). Specific diagnoses were made more accurately with agent detection imaging: At McNemar testing, the number of correct lesion type determinations increased from 83 to 92 (P =.022) for reader 1 and from 85 to 99 (P <.001) for reader 2. Both readers assigned high scores for differences in SH U 508A uptake between the liver parenchyma and the lesion for metastases and cholangiocarcinomas and low scores for uptake differences in most of the benign lesions. Hepatocellular carcinomas (HCCs), hemangiomas, and adenomas had more variable uptake differences. Fourteen of 22 hemangiomas were assigned an SCS of less than 50%, and 22 (reader 1) and 15 (reader 2) of 31 HCCs were assigned an SCS of greater than 50%. CONCLUSION: With use of SH U 508A-enhanced agent detection imaging, liver lesion characterization and diagnostic performance are significantly improved. PMID- 15284435 TI - Breast abscess in lactating women: US-guided treatment. AB - PURPOSE: To retrospectively evaluate the use of ultrasonography (US)-guided treatment of breast abscesses in lactating women. MATERIALS AND METHODS: One hundred eight consecutive lactating women who were clinically suspected of having a breast abscess were examined with US. Abscesses depicted at US were treated with US guidance, and the success of US-guided treatment was retrospectively determined. RESULTS: Fifty-six abscesses were identified at US in 43 women; all abscesses were treated with US guidance: 23 with needle aspiration and 33 with catheter drainage. Treatment method was determined according to the size of the abscess. Abscesses that were smaller than 3 cm in maximum diameter were treated with needle aspiration, and abscesses that were 3 cm or larger in maximum diameter were treated with catheter insertion. One patient who was treated with needle aspiration subsequently underwent surgical intervention; all others were successfully treated with US intervention. Catheter placement was well tolerated (mean pain score 2.3 in 22 women by using a subjective pain scale of 0-10). CONCLUSION: US-guided needle aspiration of abscesses smaller than 3 cm and US guided catheter drainage of abscesses 3 cm or larger are successful means of treating breast abscesses. PMID- 15284436 TI - Absolute myocardial perfusion in canines measured by using dual-bolus first-pass MR imaging. AB - PURPOSE: To compare fluorescent microsphere measurements of myocardial blood flow (MBF) with qualitative, semiquantitative, and fully quantitative measurements of first-pass perfusion at magnetic resonance (MR) imaging. MATERIALS AND METHODS: Coronary artery occlusion or intracoronary adenosine infusion was successfully performed in 16 beagles; both procedures were performed simultaneously in one animal. MBF was assessed at microsphere analysis. First-pass myocardial perfusion MR imaging was performed during a dual-bolus administration of gadopentetate dimeglumine (0.0025 mmol/kg followed by 0.10 mmol/kg). The absolute myocardial perfusion at MR imaging was calculated by using Fermi function deconvolution methods. Qualitative, semiquantitative, and absolute myocardial perfusion MR imaging measurements were compared with microsphere MBF measurements by using paired t tests, linear correlation, and Bland-Altman analysis. RESULTS: Fully quantitative (ie, absolute) analysis of MBF at MR imaging correlated with microsphere MBF measurement (r = 0.95, P <.001) across the full range of blood flow rates encountered (from 0 to >5.0 mL/min/g). Similar close correlations were observed in endocardial and epicardial segments (representing approximately 0.85 g of the myocardium). With modest increases in MBF, qualitative measurements plateaued in the hyperemic zones. Semiquantitative measurements did not correlate with MBF as well (r = 0.69-0.89); they plateaued around 3.0 mL/min/g. CONCLUSION: Dual-bolus MR imaging enabled accurate measurement of absolute epicardial and endocardial perfusion across a wide range of blood flow rates (0 to >5.0 mL/min/g). Use of qualitative MR imaging measures such as the contrast enhancement ratio led to substantially underestimated hyperemic blood flow measurements. PMID- 15284437 TI - Acute and subacute intracerebral hemorrhages: comparison of MR imaging at 1.5 and 3.0 T--initial experience. AB - PURPOSE: To assess and describe the appearance of intracerebral hemorrhage (ICH) at 3.0-T magnetic resonance (MR) imaging as compared with the appearance of this lesion type at 1.5-T MR imaging. MATERIALS AND METHODS: Sixteen patients with 21 parenchymal ICHs were examined. ICHs were classified as hyperacute, acute, early subacute, late subacute, or chronic. Patients underwent 1.5- and 3.0-T MR imaging with T2-weighted fast spin-echo, fluid-attenuated inversion-recovery (FLAIR), and T1-weighted spin-echo (1.5-T) and gradient-echo (3.0-T) sequences within 4 hours of each other. The central (ie, core) and peripheral (ie, body) parts of the ICHs were analyzed quantitatively by using contrast-to-noise ratio (CNR) calculations derived from signal intensity (SI) measurements; these values were statistically evaluated by using the Mann-Whitney U test. Two readers qualitatively determined SIs of the cores and bodies of the ICHs, degrees of apparent susceptibility artifacts, and lesion ages. The chi(2) test was used to determine statistically significant differences. RESULTS: With the exception of the bodies of late subacute ICHs at 3.0-T T2-weighted imaging, which had increased positive CNRs and SI scores (P .05). With the exception of minor susceptibility artifacts seen in acute and early subacute ICHs at 3.0-T T1-weighted gradient-echo imaging, no susceptibility artifacts were noticed. The ages of most lesions were identified correctly without significant differences between the two field strengths (P >.05), with the exception of the ages of acute ICHs, which were occasionally misinterpreted as early subacute lesions at 3.0 T. CONCLUSION: At 3.0 T, all parts of acute and early subacute ICHs had significantly increased hypointensity on FLAIR and T2-weighted MR images as compared with the SIs of these lesions at 1.5 T. However, 1.5- and 3.0-T MR images were equivalent in the determination of acute to late subacute ICHs. PMID- 15284438 TI - Multiprotein complex containing succinate dehydrogenase confers mitochondrial ATP sensitive K+ channel activity. AB - The mitochondrial ATP-sensitive K(+) (mitoK(ATP)) channel plays a central role in protection of cardiac and neuronal cells against ischemia and apoptosis, but its molecular structure is unknown. Succinate dehydrogenase (SDH) is inhibited by mitoK(ATP) activators, fueling the contrary view that SDH, rather than mitoK(ATP), is the target of cardioprotective drugs. Here, we report that SDH forms part of mitoK(ATP) functionally and structurally. Four mitochondrial proteins [mitochondrial ATP-binding cassette protein 1 (mABC1), phosphate carrier, adenine nucleotide translocator, and ATP synthase] associate with SDH. A purified IM fraction containing these proteins was reconstituted into proteoliposomes and lipid bilayers and shown to confer mitoK(ATP) channel activity. This channel activity is sensitive not only to mitoK(ATP) activators and blockers but also to SDH inhibitors. These results reconcile the controversy over the basis of ischemic preconditioning by demonstrating that SDH is a component of mitoK(ATP) as part of a macromolecular supercomplex. The findings also provide a tangible clue as to the structural basis of mitoK(ATP) channels. PMID- 15284439 TI - The kinase Grk2 regulates Nedd4/Nedd4-2-dependent control of epithelial Na+ channels. AB - Epithelial Na(+) channels mediate the transport of Na across epithelia in the kidney, gut, and lungs and are required for blood pressure regulation. They are inhibited by ubiquitin protein ligases, such as Nedd4 and Nedd4-2, with loss of this inhibition leading to hypertension. Here, we report that these channels are maintained in the active state by the G protein-coupled receptor kinase, Grk2, which has been previously implicated in the development of essential hypertension. We also show that Grk2 phosphorylates the C terminus of the channel beta subunit and renders the channels insensitive to inhibition by Nedd4-2. This mechanism has not been previously reported to regulate epithelial Na(+) channels and provides a potential explanation for the observed association of Grk2 overactivity with hypertension. Here, we report a G protein-coupled receptor kinase regulating a membrane protein other than a receptor and provide a paradigm for understanding how the interaction between membrane proteins and ubiquitin protein ligases is controlled. PMID- 15284441 TI - Inhibition of apoptosis in acute promyelocytic leukemia cells leads to increases in levels of oxidized protein and LMP2 immunoproteasome. AB - On reaching maturity, animal organs cease to increase in size because of inhibition of cell replication activities. It follows that maintenance of optimal organ function depends on the elimination of oxidatively damaged cells and their replacement with new cells. To examine the effects of oxidative stress and apoptosis on the accumulation of oxidized proteins, we exposed acute promyelocytic leukemia cells to arsenic trioxide (As(2)O(3)) in the presence and absence of a general caspase inhibitor (benzyloxycarbonyl-Val-Ala-Asp fluoromethyl ketone), which is known to inhibit caspase-induced apoptosis. We confirm that treatment of cells with As(2)O(3) induces apoptosis and leads to the accumulation of oxidized proteins. Furthermore, inhibition of caspase activities prevented As(2)O(3)-induced apoptosis and led to a substantial increase in accumulation of oxidized proteins. Moreover, inhibition of caspase activity in the absence of As(2)O(3) led to elevated levels of the LMP2 immunoproteasome protein. We also show that caspase inhibition leads to increases in the levels of oxidized proteins obtained by treatments with hydrogen peroxide plus ferrous iron. Collectively, these results suggest the possibility that an age-related loss in capacity to carry out apoptosis might contribute to the observed accumulation of oxidized proteins during aging and in age-related diseases. PMID- 15284440 TI - Kinetics of regulated protein-protein interactions revealed with firefly luciferase complementation imaging in cells and living animals. AB - Signaling pathways regulating proliferation, differentiation, and apoptosis are commonly mediated through protein-protein interactions as well as reversible phosphorylation of proteins. To facilitate the study of regulated protein-protein interactions in cells and living animals, we optimized firefly luciferase protein fragment complementation by screening incremental truncation libraries of N- and C-terminal fragments of luciferase. Fused to the rapamycin-binding domain (FRB) of the kinase mammalian target of rapamycin and FK506-binding protein 12 (FKBP), respectively, the optimized FRB-N-terminal luciferase fragment (NLuc)/C-terminal luciferase fragment (CLuc)-FKBP luciferase complementation imaging (LCI) pair reconstituted luciferase activity in cells upon single-site binding of rapamycin in an FK506-competitive manner. LCI was used in three independent applications. In mice bearing implants of cells expressing the FRB-NLuc/CLuc-FKBP LCI pair, dose- and time-dependent luciferase activity allowed target-specific pharmacodynamic analysis of rapamycin-induced protein-protein interactions in vivo. In cells expressing a Cdc25C-NLuc/CLuc-14-3-3epsilon LCI pair, drug mediated disruption of cell cycle regulated protein-protein interactions was demonstrated with the protein kinase inhibitor UCN-01 in a phosphoserine dependent manner. When applied to IFN-gamma-dependent activation of Janus kinase/signal transducer and activator of transcription 1 (STAT1), LCI revealed, in the absence of ligand-induced phosphorylation, STAT1 proteins existing in live cells as preformed dimers. Thus, optimized LCI provides a platform for near real time detection and characterization of regulated and small molecule-induced protein-protein interactions in intact cells and living animals and should enable a wide range of novel applications in drug discovery, chemical genetics, and proteomics research. PMID- 15284442 TI - Crystal structure of pyrogallol-phloroglucinol transhydroxylase, an Mo enzyme capable of intermolecular hydroxyl transfer between phenols. AB - The Mo enzyme transhydroxylase from the anaerobic microorganism Pelobacter acidigallici catalyzes the conversion of pyrogallol to phloroglucinol. Such trihydroxybenzenes and their derivatives represent important building blocks of plant polymers. None of the transferred hydroxyl groups originates from water during transhydroxylation; instead a cosubstrate, such as 1,2,3,5 tetrahydroxybenzene, is used in a reaction without apparent electron transfer. Here, we report on the crystal structure of the enzyme in the reduced Mo(IV) state, which we solved by single anomalous-diffraction technique. It represents the largest structure (1,149 amino acid residues per molecule, 12 independent molecules per unit cell), which has been solved so far by single anomalous diffraction technique. Tranhydroxylase is a heterodimer, with the active Mo molybdopterin guanine dinucleotide (MGD)(2) site in the alpha-subunit, and three [4Fe-4S] centers in the beta-subunit. The latter subunit carries a seven stranded, mainly antiparallel beta-barrel domain. We propose a scheme for the transhydroxylation reaction based on 3D structures of complexes of the enzyme with various polyphenols serving either as substrate or inhibitor. PMID- 15284443 TI - MicroRNA profiling reveals distinct signatures in B cell chronic lymphocytic leukemias. AB - Little is known about the expression levels or function of micro-RNAs (miRNAs) in normal and neoplastic cells, although it is becoming clear that miRNAs play important roles in the regulation of gene expression during development [Ambros, V. (2003) Cell 113, 673-676; McManus, M. T. (2003) Semin. Cancer Biol. 13, 253 258]. We now report the genomewide expression profiling of miRNAs in human B cell chronic lymphocytic leukemia (CLL) by using a microarray containing hundreds of human precursor and mature miRNA oligonucleotide probes. This approach allowed us to identify significant differences in miRNome expression between CLL samples and normal CD5+ B cells; data were confirmed by Northern blot analyses and real-time RT-PCR. At least two distinct clusters of CLL samples can be identified that were associated with the presence or absence of Zap-70 expression, a predictor of early disease progression. Two miRNA signatures were associated with the presence or absence of mutations in the expressed Ig variableregion genes or with deletions at 13q14, respectively. These data suggest that miRNA expression patterns have relevance to the biological and clinical behavior of this leukemia. PMID- 15284444 TI - Tubular precipitation and redox gradients on a bubbling template. AB - Tubular structures created by precipitation abound in nature, from chimneys at hydrothermal vents to soda straws in caves. Their formation is controlled by chemical gradients within which precipitation occurs, defining a surface that templates the growing structure. We report a self-organized periodic templating mechanism producing tubular structures electrochemically in iron-ammonium-sulfate solutions; iron oxides precipitate on the surface of bubbles that linger at the tube rim and then detach, leaving behind a ring of material. The acid-base and redox gradients spontaneously generated by diffusion of ammonia from the bubble into solution organize radial compositional layering within the tube wall, a mechanism studied on a larger scale by complex Liesegang patterns of iron oxides formed as ammonia diffuses through a gel containing FeSO(4). When magnetite forms within the wall, a tube may grow curved in an external magnetic field. Connections with free-boundary problems in speleothem formation are emphasized. PMID- 15284445 TI - Does transcatheter closure of patent foramen ovale really "shut the door?" A prospective study with transcranial Doppler. AB - BACKGROUND AND PURPOSE: Transcatheter closure of patent foramen ovale (PFO) is increasingly being performed and monitored with transthoracic or transesophageal echocardiography, whereas contrast-enhanced transcranial Doppler (ce-TCD), which probably represents the most suitable tool to quantify right-to-left shunt (RLS) in the brain vessels, has been systematically overlooked. Our goal is to prospectively assess efficacy and safety of PFO transcatheter closure using ce TCD. METHODS: A total of 140 consecutive patients (mean age, 46+/-13 years; male/female ratio, 63/77) with PFO-related large RLS and no other recognized cause of focal cerebral ischemia underwent transcatheter closure. TCD was done preoperatively and 1 month after the procedure in all patients, after 3 months in 120, after 6 months in 112, and after 1 year in 104 patients. RESULTS: Implantation was successful in all patients. During Valsalva strain, a large shunt was still detectable in 31 of 140 (22%), 15 of 120 (13%), 9 of 112 (8%), and 9 of 104 (9%) patients at the 1-, 3-, 6-, and 12-month visits, respectively. Periprocedural and postprocedural complications included atrial fibrillation in 8% and scintillating scotomata in 6% of patients. During the 1-year follow-up period, only 1 transient ischemic attack was recorded in a patient with paroxysmal atrial fibrillation and complete PFO closure. CONCLUSIONS: Transcatheter PFO closure in patients with cryptogenic stroke and large RLS may be less successful than reported previously. TCD appears the ideal tool to follow up the closure process and to identify early, during follow-up, those patients who will be left with a significant shunt. Atrial fibrillation is more common than believed previously and may underlie the occurrence of further cerebrovascular events despite complete PFO closure. Irritative visual phenomena may occur as a consequence of nickel toxicity. PMID- 15284447 TI - The Willis Lecture 2003: evaluating treatments for stroke patients too slowly: time to get out of second gear. PMID- 15284446 TI - Targets for vascular protection after acute ischemic stroke. AB - BACKGROUND: Vascular damage caused by cerebral ischemia leads to edema, hemorrhage formation, and worsened outcomes in ischemic stroke patients. Therapeutic interventions need to be developed to provide vascular protection. The purpose of this review is to identify the pathophysiologic processes involved in vascular damage after ischemia, which may lead to strategies to provide vascular protection in ischemic stroke patients. SUMMARY OF COMMENT: The pathologic processes caused by vascular injury after an occlusion of a cerebral artery can be separated into acute (hours), subacute (hours to days), and chronic (days to months). Targets for intervention can be identified for all 3 stages. Acutely, superoxide is the predominant mediator, followed by inflammatory mediators and proteases subacutely. In the chronic phase, proapoptotic gene products have been implicated. CONCLUSIONS: Pharmacological agents designed to target specific pathologic and protective processes affecting the vasculature should be used in clinical trials of vascular protection after acute ischemic stroke. PMID- 15284448 TI - Mild to moderate atheromatous disease of the thoracic aorta and new ischemic brain lesions after conventional coronary artery bypass graft surgery. AB - BACKGROUND AND PURPOSE: The presence of new ischemic brain infarcts, detected by diffusion-weighted magnetic resonance imaging (DW-MRI), have been reported in considerable number of patients after cardiac surgery. We sought to determine the role of proximal thoracic aortic atheroma in predicting embolic events and new ischemic brain lesions in patients undergoing conventional coronary revascularization surgery. METHODS: Transesophageal echocardiography and epiaortic scanning was performed to assess the severity of aortic atherosclerosis in the ascending aorta and the aortic arch. Patients were allocated to either low risk group, (intimal thickness < or =2mm), or high-risk group (intimal thickness >2mm). Transcranial Doppler was used to monitor the middle cerebral artery. DW MRI was performed 3-7 days after surgery. The NEECHAM Confusion Scale was used for assessment and monitoring patient consciousness level. RESULTS: Patients in the high-risk group were considerably older; 71+/-6 (n=38) versus 67+/-6 (n=72) years, P=0.004 and were more likely to have impaired left ventricular function. Confusion was present in 6 (16%) patients in the high-risk group and 5 (7%) patients in the low-risk group. Patients in the high-risk group had a three-fold increase in median embolic count, 223.5 versus 70.0, P=0.0003. DW-MRI detected brain lesions were only present in patients from high-risk group, 61.5 versus 0%, P<0.0001. There was significant correlation between the NEECHAM scores and embolic count in the high-risk group; r=0.63, P<0.001. CONCLUSIONS: The findings of this investigation suggest that mild to moderate atheromatous disease of the ascending aorta and the aortic arch (intimal thickness >2mm) is a major contributor to ischemic brain injury after cardiac surgery. PMID- 15284449 TI - Genetic variation in PPARG encoding peroxisome proliferator-activated receptor gamma associated with carotid atherosclerosis. AB - BACKGROUND AND PURPOSE: Peroxisome proliferator-activated receptor gamma is a crucial molecule in atherogenesis because it is associated with metabolic risk factors such as obesity and diabetes and also plays a key role in subcellular metabolism of arterial wall macrophage foam cells. Genetic variation in PPARG has been associated with metabolic and cardiovascular end points. METHODS: We investigated the relationship between 2 common PPARG polymorphisms, namely P12A and c.1431C>T, and carotid atherosclerosis in a sample of 161 Canadian aboriginal people. Dependent variables were carotid intima media thickness (IMT), assessed using B-mode ultrasonography, and total carotid plaque volume (TPV), assessed using 3D ultrasound. RESULTS: Using multivariate analysis, we found that subjects with > or =1 PPARG A12 allele had less carotid IMT than others (0.72+/-0.03 versus 0.80+/-0.02 mm; P=0.0045), with no between-genotype difference in TPV. In contrast, subjects with the PPARG c.1431T allele had greater TPV than others (124+/-18.4 versus 65.1+/-23.7 mm3; P=0.0079), with no between-genotype difference in IMT. CONCLUSIONS: The findings show an association between PPARG genotypes and carotid arterial phenotypes, and further reflect the prevailing view that the PPARG A12 allele protects against deleterious phenotypes. Also, whereas IMT and TPV are somewhat correlated with each other, they might also represent distinct traits with discrete determinants representing different stages of atherogenesis. PMID- 15284450 TI - Intracerebral transplantation of porcine choroid plexus provides structural and functional neuroprotection in a rodent model of stroke. AB - BACKGROUND AND PURPOSE: Choroid plexus (CP) secretes a cocktail of neurotrophic factors. In the present study, CP from neonatal pigs was encapsulated within alginate microcapsules for in vitro and in vivo neuroprotective studies. METHODS: In vitro studies involved serum deprivation of rat embryonic cortical neurons and treatment with a range of concentrations of conditioned media from CP. For in vivo studies, rats received a 1-hour middle cerebral artery occlusion followed by intracranial transplantation of encapsulated or unencapsulated CP, empty capsules, or no transplant. Behavioral testing was conducted on days 1 to 3 after transplantation. Cerebral infarction was analyzed using 2,3,5-triphenyl tetrazolium chloride staining at 3 days after transplantation. RESULTS: Conditioned media from CP produced a significant dose-dependent protection of serum-deprived cortical neurons. Enzyme-linked immunosorbent assay confirmed secretion of GDNF, BDNF, and NGF from CP. Parallel in vivo studies showed that CP transplants improved behavioral performance and decreased the volume of infarction. Both encapsulated and unencapsulated CP transplants were effective; however, more robust benefits accompanied encapsulated transplants. CONCLUSIONS: These data are the first to demonstrate the neuroprotective potential of transplanted CP and raise the intriguing possibility of using these cells as part of the treatment regimen for stroke and other neurological disorders. PMID- 15284453 TI - Crystal structure of Argonaute and its implications for RISC slicer activity. AB - Argonaute proteins and small interfering RNAs (siRNAs) are the known signature components of the RNA interference effector complex RNA-induced silencing complex (RISC). However, the identity of "Slicer," the enzyme that cleaves the messenger RNA (mRNA) as directed by the siRNA, has not been resolved. Here, we report the crystal structure of the Argonaute protein from Pyrococcus furiosus at 2.25 angstrom resolution. The structure reveals a crescent-shaped base made up of the amino-terminal, middle, and PIWI domains. The Piwi Argonaute Zwille (PAZ) domain is held above the base by a "stalk"-like region. The PIWI domain (named for the protein piwi) is similar to ribonuclease H, with a conserved active site aspartate-aspartate-glutamate motif, strongly implicating Argonaute as "Slicer." The architecture of the molecule and the placement of the PAZ and PIWI domains define a groove for substrate binding and suggest a mechanism for siRNA-guided mRNA cleavage. PMID- 15284454 TI - Observation of a one-dimensional Tonks-Girardeau gas. AB - We report the observation of a one-dimensional (1D) Tonks-Girardeau (TG) gas of bosons moving freely in 1D. Although TG gas bosons are strongly interacting, they behave very much like noninteracting fermions. We enter the TG regime with cold rubidium-87 atoms by trapping them with a combination of two light traps. By changing the trap intensities, and hence the atomic interaction strength, the atoms can be made to act either like a Bose-Einstein condensate or like a TG gas. We measure the total 1D energy and the length of the gas. With no free parameters and over a wide range of coupling strengths, our data fit the exact solution for the ground state of a 1D Bose gas. PMID- 15284455 TI - Gefitinib-sensitizing EGFR mutations in lung cancer activate anti-apoptotic pathways. AB - Gefitinib (Iressa, Astra Zeneca Pharmaceuticals) is a tyrosine kinase inhibitor that targets the epidermal growth factor receptor (EGFR) and induces dramatic clinical responses in nonsmall cell lung cancers (NSCLCs) with activating mutations within the EGFR kinase domain. We report that these mutant EGFRs selectively activate Akt and signal transduction and activator of transcription (STAT) signaling pathways, which promote cell survival, but have no effect on extracellular signal-regulated kinase signaling, which induces proliferation. NSCLC cells expressing mutant EGFRs underwent extensive apoptosis after small interfering RNA-mediated knockdown of the mutant EGFR or treatment with pharmacological inhibitors of Akt and STAT signaling and were relatively resistant to apoptosis induced by conventional chemotherapeutic drugs. Thus, mutant EGFRs selectively transduce survival signals on which NSCLCs become dependent; inhibition of those signals by gefitinib may contribute to the drug's efficacy. PMID- 15284456 TI - Argonaute2 is the catalytic engine of mammalian RNAi. AB - Gene silencing through RNA interference (RNAi) is carried out by RISC, the RNA induced silencing complex. RISC contains two signature components, small interfering RNAs (siRNAs) and Argonaute family proteins. Here, we show that the multiple Argonaute proteins present in mammals are both biologically and biochemically distinct, with a single mammalian family member, Argonaute2, being responsible for messenger RNA cleavage activity. This protein is essential for mouse development, and cells lacking Argonaute2 are unable to mount an experimental response to siRNAs. Mutations within a cryptic ribonuclease H domain within Argonaute2, as identified by comparison with the structure of an archeal Argonaute protein, inactivate RISC. Thus, our evidence supports a model in which Argonaute contributes "Slicer" activity to RISC, providing the catalytic engine for RNAi. PMID- 15284457 TI - Evaluation of fetal pulmonary vasculature by power Doppler imaging in congenital diaphragmatic hernia. AB - OBJECTIVE: To evaluate the feasibility of prenatal power Doppler imaging of pulmonary arteries in congenital diaphragmatic hernia and to study its potential to predict outcome. METHODS: A prospective observational study was conducted. Forty-two cases of congenital diaphragmatic hernia (32 left and 10 right) without associated anomalies were analyzed. Qualitative evaluation of pulmonary vasculature was based on power Doppler imaging performed at 26 to 38 weeks. The pulmonary arteries were studied in the lung contralateral to the hernia. Pulmonary Doppler angiography was considered satisfactory when 3 levels of bifurcation defining 3 distinct segments of the pulmonary arteries were imaged and was otherwise considered poor. We also recorded the gestational age at diagnosis, side of the hernia, abdominal circumference below the third percentile, amniotic fluid volume, lung/thoracic area ratio, left/right ventricle ratio, and, in left-sided hernias, stomach position, and we carried out a multivariate analysis to determine the contribution of each factor to predict neonatal mortality. RESULTS: More than 3 divisions of the fetal pulmonary arteries were imaged in 20 cases; 1 or 2 divisions or none were imaged in 22. Neonatal mortality was significantly greater when fewer than 3 divisions of the pulmonary arteries were imaged (18 [82%] of 22) than when 3 divisions could be identified on power Doppler imaging (5 [25%] of 20; P = .0005). However, the lung/thoracic area ratio was the only factor that remained significantly associated with mortality in the multivariate analysis. CONCLUSIONS: An altered pulmonary power Doppler image is associated with neonatal mortality, but estimation of the lung area remains the best predictor of neonatal outcome. PMID- 15284458 TI - The preferred timing of second-trimester obstetric sonography based on maternal body mass index. AB - OBJECTIVE: To determine the preferred timing of sonographic screening of fetal anatomy based on the maternal body mass index (BMI). METHODS: We abstracted the sonographic reports of 2,303 gravidas undergoing routine fetal anatomic screening between 15 and 24 weeks' gestation to determine the completeness of the study. Height and weight information was available on 1444 patients. The maternal BMI (weight [kilograms]/height [square meters]) was categorized as underweight (<19.8), normal weight (19.8-26.0), overweight (26.1-29.0), and obese (>29.0). Completion rates were compared by chi(2) analysis. Multiple logistic regression was used to evaluate for independent predictors of a completed study. RESULTS: Except for underweight women, completion rates for all BMI categories were significantly higher when the sonographic examinations were performed between 18 weeks and 19 weeks 6 days compared with those performed between 15 weeks and 17 weeks 6 days. Body mass index, estimated gestational age, and black race were independent predictors of a completed study. CONCLUSIONS: Except in underweight women, the 18- to 20-week interval appears to be superior to the 15- to 18-week interval when performing sonographic screening of the fetal anatomy. PMID- 15284459 TI - The accreditation of ultrasound practices: impact on compliance with minimum performance guidelines. AB - OBJECTIVE: To determine the effectiveness of the American Institute of Ultrasound in Medicine (AIUM) accreditation program in improving compliance with standards and guidelines for the performance of obstetric and gynecologic ultrasound examinations. METHODS: Scores of case studies of accreditation applications were compared with their respective scores at the time of reaccreditation 3 years later. To account for the element of time, scores of applications that recently completed first-time accreditation were also compared as a control group. RESULTS: Individual obstetric case studies, as well as the average of all obstetric and gynecologic case studies, showed highly significant improvement with the reaccreditation application when compared with the initial accreditation application 3 years earlier (all P <.001). The proportion of practices successfully meeting obstetric and gynecologic AIUM accreditation requirements improved significantly with reaccreditation (obstetric, 57.3% for accreditation compared with 86.6% for reaccreditation; gynecologic, 60% for accreditation compared with 91.9% for reaccreditation; P <.001). Furthermore, reaccreditation scores were significantly higher than scores of recent first-time applications for obstetric case studies as well as scores of the average of obstetric and gynecologic case studies (all P <.05). CONCLUSIONS: Our study confirms that practices that sought and received ultrasound accreditation were able to improve the scores of case studies and compliance with published minimum standards and guidelines for the performance of obstetric and gynecologic ultrasound examinations when reevaluated 3 years after the initial application scores. This improvement should translate into an enhancement of the quality of ultrasound practice. PMID- 15284460 TI - Detection of muscle atrophy on routine sonography of the shoulder. AB - OBJECTIVE: To describe the utility of sonography in visualizing muscle atrophy during routine sonographic examination of the shoulder for evaluation of the rotator cuff tendons. METHODS: A retrospective review of 199 shoulder sonographic examinations performed by 2 musculoskeletal radiologists trained in musculoskeletal sonography with knowledge of the typical sonographic findings of muscle atrophy was performed. Reports were reviewed for the presence of muscle atrophy. If atrophy was present, the reports from those examinations were rereviewed for concomitant rotator cuff abnormalities. RESULTS: Forty-five examinations (23%) showed atrophy in at least 1 muscle on the basis of the criteria of increased echogenicity and decreased bulk. There were a total of 81 individual muscles that showed atrophy, with the following distribution: 16% supraspinatus (n = 13), 31% infraspinatus (n = 25), 36% teres minor (n = 29), 2% subscapularis (n = 2), and 6% biceps brachii (n = 5). In 34 of the 45 examinations with muscle atrophy, there were 57 concomitant full-thickness tendon tears: 64% supraspinatus (n = 29), 38% infraspinatus (n = 17), 7% subscapularis (n = 3), 0% teres minor (n = 0), 16% long head of biceps (n = 7), and 2% deltoid (n = 1). CONCLUSIONS: Although primary sonographic evaluation of the painful shoulder concentrates on the tendons of the rotator cuff, we suggest that examination of the muscles should become a standard component of the comprehensive shoulder sonographic examination, particularly given the potential clinical implications of muscle atrophy on the outcome of rotator cuff surgery. PMID- 15284461 TI - Validation of the depletion kinetic in semiquantitative ultrasonographic cerebral perfusion imaging using 2 different techniques of data acquisition. AB - OBJECTIVE: To validate the potential of ultrasonographic depletion imaging for semiquantitatively visualizing cerebral parenchymal perfusion with contrast burst depletion imaging (CODIM) in comparison with phase inversion harmonic depletion imaging (PIDIM) in healthy volunteers. METHODS: Thirteen healthy adults were examined with both CODIM and PIDIM in accordance with previously described criteria. In addition to the perfusion coefficient, the time to decrease image intensity to 10% above equilibrium intensity from the initial value and the relative error (deviation of measured data from the fitted model) were evaluated to compare the reliability of both techniques in 3 different regions of interest. RESULTS: Perfusion coefficient values did not show significantly differing values in both groups (1.57-1.64 * 10(-2) s(-1) for CODIM and 1.42-1.58 * 10(-2) s(-1) for PIDIM). The relative error was significantly smaller in the PIDIM group (0.38 0.53 for CODIM and 0.18-0.25 for PIDIM; P < .002). CONCLUSIONS: Phase inversion harmonic depletion imaging proved to be more reliable than CODIM because values of the relative error were significantly lower in PIDIM even in this relatively small cohort. This is of interest because the underlying technique, phase inversion harmonic imaging, is more widely available than contrast burst imaging. PMID- 15284462 TI - Evaluation of differences between observers and automatic-manual measurements in calculation of Doppler parameters. AB - OBJECTIVE: We aimed to search for differences between observers and automatic and manual measurements in calculations of Doppler parameters. METHODS: The middle cerebral artery (MCA), central retinal artery, ophthalmic artery (OA), common carotid artery (CCA), vertebral artery (VA), popliteal artery (PA), interlobar renal artery (IRA), and arcuate renal artery (ARA) were evaluated in 20 healthy subjects bilaterally. Peak systolic velocity (PSV), end-diastolic velocity (EDV), time-averaged maximum velocity (TAMAX), resistive index (RI), and pulsatility index (PI) were measured from the same spectrum manually by 3 observers and automatically. Results of 4 measurements were compared by analysis of variance and Pearson tests. RESULTS: The comparison of the 4 measurements revealed significant differences for most parameters except TAMAX of the OA, VA, and ARA and PSV, EDV, and PI of the PA. An automatic calculator yielded lower PSV, RI, and PI values (except the MCA and PA) and higher EDV values compared with manual measurements. The magnitudes of difference were in the range of 1% to 16% for velocities and 4% to 14% for RI and PI. The means of difference were 3.185 cm/s for PSV of the CCA and 0.054 for RI of the IRA. Correlation was high for PSV, EDV, and TAMAX in all arteries (except TAMAX of PA) and relatively low for PI and RI in most of the arteries. CONCLUSIONS: Although our study was performed on healthy subjects, our results showed that, in most cases, readers and the automatic approach disagreed on evaluation of Doppler parameters. This may be important in preventing false diagnoses in cases with Doppler values close to upper limits and may necessitate establishment of new limits for each method. PMID- 15284463 TI - Three-dimensional color-coded duplex sonography for assessment of the vertebral artery origin and vertebral artery stenoses. AB - OBJECTIVE: The aim of the study was to assess the potential of 3-dimensional (3D) color-coded duplex sonography (CDS) for evaluation of the vertebral artery origin and stenoses in this location. METHODS: To compare 2-dimensional (2D) and 3D CDS, both techniques were performed in 25 healthy volunteers and in 18 patients with 21 stenoses of the vertebral artery origin. Stenoses were graded in line with hemodynamic criteria on 2D CDS and according to North American Symptomatic Carotid Endarterectomy Trial criteria on 3D CDS. In 6 patients, digital subtraction angiography (DSA) was performed additionally. Stenoses were graded according to North American Symptomatic Carotid Endarterectomy Trial criteria and compared with 2D and 3D sonographic data. RESULTS: Overall correlation of both sonographic techniques concerning the grading of the stenoses was good (r = 0.69; P < .01). The interobserver correlation for assessment of stenoses by means of 3D CDS was high (r = 0.94; P < .01). Three-dimensional CDS correlated excellently with DSA in 3 of 6 patients but showed only intermediate or no correlation in the remaining 3 patients. In contrast, spatial information on the stenotic morphologic characteristics was always very comparable with the results obtained by DSA. CONCLUSIONS: Three-dimensional CDS represents a valuable tool for assessment of the origin of the vertebral artery, allowing important morphologic information on stenoses in this location. For grading of stenoses of the vertebral artery origin, 3D information should be combined with hemodynamic criteria obtained by spectral Doppler imaging in 2D CDS. Three-dimensional CDS could be a valuable tool before interventional procedures of the proximal vertebral artery, saving time and avoiding iodinated contrast agents. PMID- 15284464 TI - Sonographic follow-up after visceral artery stenting. AB - OBJECTIVE: The aim of this study was to evaluate the sonographic features of stents and the flow parameters of the visceral arteries after stent implantation. METHODS: Since 1996, 34 stenoses of the visceral arteries (2 mesenteric, 4 celiac trunk, and 28 renal arteries) in 28 patients have been treated with metallic stent implantation in the Department of Radiology of Szeged Medical University. All these patients were regularly followed sonographically. For the diagnosis of restenosis, previously published criteria were used. RESULTS: All the mesenteric and celiac stents could be visualized, but none of the renal stents were clearly seen sonographically. The flow parameters could be established in all cases. Sonographic examination revealed 1 occlusion, 2 restenoses, and 1 stent displacement. All these abnormalities were confirmed by other imaging modalities. CONCLUSIONS: Sonography is a useful tool in the follow-up of patients after visceral artery stenting. Despite the fact that none of the renal artery stents were visualized directly, the flow parameters could be evaluated, and the pathologic changes were found. PMID- 15284465 TI - Intraoperative sonography of intracranial arteriovenous malformations: how we do it. AB - OBJECTIVE: We have advanced the application of intraoperative neurosonography by combining gray scale sonographic imaging with pulsed wave Doppler and color flow Doppler imaging to guide and confirm resection of arteriovenous malformations of the brain. We want to share our technique with the imaging community. METHODS: We present a review of our scan technique as it has evolved over the 3 years during which we have been assisting our neurosurgical team. RESULTS AND CONCLUSIONS: Our experience has indicated that a combination of sonographic imaging and color and spectral Doppler imaging improves surgical resection of such lesions. PMID- 15284466 TI - Torsion and beyond: new twists in spectral Doppler evaluation of the scrotum. AB - OBJECTIVE: To illustrate how spectral Doppler waveform analysis plays an adjunctive but very definite role in scrotal sonography. METHODS: The cases illustrate a variety of testicular disorders that were collected at a referral tertiary care center. RESULTS: Normal and a variety of pathologic conditions of the testes are discussed, along with their signature spectral waveforms. CONCLUSIONS: Analysis of the spectral waveform provides important additional information in various scrotal disorders with acute pain. Spectral waveform analysis is critical to diagnosing incomplete torsion when color and power Doppler examinations are indeterminate. PMID- 15284467 TI - Splenogonadal fusion: B-mode and color Doppler sonographic appearances. PMID- 15284468 TI - Iatrogenic pseudoaneurysm resulting in transection of the radial artery. PMID- 15284469 TI - Dural arteriovenous fistula after cerebral sinus thrombosis: a case study of serial venous transcranial color-coded sonography. PMID- 15284470 TI - Conservative management of a uterine arteriovenous malformation diagnosed in pregnancy. PMID- 15284471 TI - Quadruplet pregnancy: two sets of twins, each occupying a horn of a septate (complete) uterus. PMID- 15284472 TI - Prenatal diagnosis of sharp-angled lumbosacral kyphosis with myelomeningocele and spina bifida in a fetus. PMID- 15284473 TI - Congenital epulis: three-dimensional ultrasonographic findings and clinical implications. PMID- 15284474 TI - Contrast-enhanced ultrasonography in the characterization of pancreatic mucinous cystadenoma. PMID- 15284475 TI - History of emergency ultrasound. PMID- 15284476 TI - Echogenic intracardiac foci and choroid plexus cysts. PMID- 15284477 TI - Visualization of a functional KSHV episome-maintenance protein LANA in living cells. AB - The latency-associated nuclear antigen (LANA) of Kaposi's saroma-associated herpesvirus (KSHV) can maintain a plasmid containing the KSHV origin of DNA replication (oriP) as episomes in dividing human cells. Hence, LANA is considered to play crucial roles in persistent KSHV infection in human cells. In this study, we characterized a LANA fusion protein of green fluorescent protein (GFP-LANA). Like the wild-type LANA, GFP-LANA interacted tightly with mitotic chromosomes, and maintained the plasmid selectively with the KSHV oriP for more than three weeks in a human B cell line. Moreover, equivalent amount of GFP-LANA protein was segregated into two daughter cells in living metaphase cells. Our results suggested that the activity of LANA serves the segregation of equivalent amounts of viral genomes tethered with LANA into two daughter progeny cells during cell division. Thus, GFP-LANA is a useful tool for the analyses of the functions and dynamics of LANA in living cells. PMID- 15284478 TI - Molecular analysis of SV-40-CAL, a new slow growing SV-40 strain from the kidney of a caged New World monkey with fatal renal disease. AB - A decline of the Callimico goeldii population in American zoos is presently occurring due to glomerulonephritis of unknown etiology. We hypothesized that this emerging idiopathic fatal renal disease (IFRD) was caused by a virus. We therefore attempted to isolate virus from the kidneys three C. goeldi in Illinois that had IFRD. Along with other viruses, Simian virus 40 (SV-40) strain CAL was isolated. SV-40-CAL is currently the slowest-growing natural isolate of SV-40 in CV-1 cells. Inefficient SV-40-CAL growth in CV-1 cells stems from two features: a suboptimal protoarchetypal regulatory region, and a Large tumor antigen gene sequence like that of SV-40 strain T302, previously considered the slow-growing natural isolate of SV-40. To our knowledge, this is the first documented isolation of SV-40 from a New World monkey outside of a laboratory setting. Though SV-40 is renaltropic, the role of SV-40-CAL in IFRD is uncertain. Transmission of SV-40 to C. goeldii through anthropogenic activity is suspected. PMID- 15284479 TI - Rapid parallel expression in E. Coli and insect cells: analysis of five lef gene products of the Autographa californica multiple nuclear polyhedrosis virus (AcMNPV). AB - A number of strategies are emerging for the high throughput (HTP) expression of recombinant proteins to enable structural and functional study. Here we describe a workable HTP strategy based on parallel protein expression in E. coli and insect cells. Using this system we provide comparative expression data for five proteins derived from the Autographa californica polyhedrosis virus genome that vary in amino acid composition and in molecular weight. Although the proteins are part of a set of factors known to be required for viral late gene expression, the precise function of three of the five, late expression factors (lefs) 6, 7 and 10, is unknown. Rapid expression and characterisation has allowed the determination of their ability to bind DNA and shown a cellular location consistent with their properties. Our data point to the utility of a parallel expression strategy to rapidly obtain workable protein expression levels from many open reading frames (ORFs). PMID- 15284480 TI - Genomic characterization of a slow/low maedi visna virus. AB - The complete genomic sequence of a sheep lentivirus isolate that presents a slow/low phenotype in vitro has been determined. The virus, designated P1OLV, was isolated from lung cells of a naturally infected sheep in Portugal. Three overlapping DNA fragments amplified by PCR, and encompassing the entire viral genome were cloned and sequenced. This isolate has an overall similarity of approximately 80% with the K1514 Maedi Visna virus (MVV) and approximately 70% with the caprine arthritis encephalitis virus (CAEV) Co strain. Phylogenetic analysis based on SU and RT nucleotide sequences grouped P1OLV with previously reported ovine MVV. To determine the virus replication rate, sheep choroid plexus (SCP) and lung cells, macrophages (MO), and goat synovial membrane (GSM) cells were inoculated with either P1OLV or with the lytic North American strain WLC-1. Viral RNA in culture supernatants was measured by one-tube real time quantitative RT-PCR. Significant differences were observed between the replication rates of the two viruses, with WLC-1 growing rapidly and to high levels in all the cells tested, while P1OLV replicated more slowly and to lower levels inducing persistent infections in lung and SCP cells. The U3 region of the LTR of P1OLV lacks the sequence repeats that are present in the LTRs of WLC-1 and MVV prototype K1514 and that contain additional binding sites for the AML(vis) transcriptional factor. To evaluate the contribution of LTR in the virus replication rate in vitro, we measured the basal activity of the promoter from P1OLV and WLC-1 in a luciferase-driven gene expression assay and lower levels of expression were achieved for P1OLV. The genetic and biological properties of P1OLV will be useful for the study of virus transcriptional factors and genes that may be responsible for the slow/low phenotype. PMID- 15284481 TI - Characterization of the baculovirus Bombyx mori nucleopolyhedrovirus gene homologous to the mammalian FGF gene family. AB - We characterized a gene of the baculovirus Bombyx mori nucleopolyhedrovirus (BmNPV) homologous to the mammalian fibroblast growth factor (FGF) family. We termed it vfgf, and examined its transcription and the properties of the gene product (vFGF). RT-PCR analysis showed that vfgf is one of the baculovirus early genes, although there are no consensus sequences of the baculovirus early gene promoters. 5'-RACE analysis revealed that its transcription started at 10 nucleotides upstream of the translation start codon. vFGF has a hydrophobic amino terminus (approximately 16 amino acids), which is a typical signal sequence. As expected, vFGF was efficiently secreted from BmNPV-infected BmN cells. Because possible glycosylation sites are found at positions 44 (Asn) and 171 (Asn), we examined whether BmNPV vFGF is glycosylated or not. Cleavage of recombinant vFGF with PNGase F revealed that BmNPV vFGF was glycosylated. We also found that secretion of vFGF is completely blocked by the treatment of Tunicamycin, which blocks N-linked glycosylation. This is the first report to characterize a virus encoded FGF. PMID- 15284482 TI - Nucleotide sequence and phylogenetic analyses of dengue type 2 virus isolated in the Dominican Republic. AB - Dengue virus infection has been recognized as an important public health problem in the Dominican Republic in the last decade. Complete genomic sequences of three strains of dengue type 2 (DEN-2) virus, DR23/01 and DR31/01 isolated from dengue fever (DF) patients, and DR59/01 isolated from a dengue hemorrhagic fever (DHF) patient, all with primary infection, in the Dominican Republic in 2001, have been established. This achievement constitutes the first genomic characterization of DEN-2 strains from the Dominican Republic. No amino acid differences were observed between the strains isolated from DHF and DF patients. They exhibited extensive homology with the strain from La Martinique, French West Indies. Although phylogenetic analysis was suggestive of their Southeast Asiatic origin, Dominican Republic strains and other Caribbean strains from La Martinique and Jamaica showed 26 amino acid changes that differed from both the Southeast Asia and native American genotypes. PMID- 15284483 TI - Phylogenetic analysis of 5'-UTR and P1 protein of Indian common strain of potato virus Y reveals its possible introduction in India. AB - The 5' untranslated region (UTR) and P1 region of the Indian strain of potato virus Y ordinary strain (PVYO) was cloned and sequenced for the first time. Database searches and multiple sequence alignment showed the highest sequence similarity with the PVYO strains of European origin. Based on the phylogenetic analysis and multiple sequence alignment, the possible evolution of PVYN from PVYO is predicted. PVYO strains from China and India were perhaps introduced into these countries from a similar geographical location. All major PVY strains available in the database can be classified into two major subgroups of North American and European origin. The Chinese and Indian PVYO strains fall within the European union subgroup suggesting a long association since potato was introduced from Europe into these countries by two separate independent events. The possible function of P1 protein in plant virus replication is suggested due to in-silico prediction of nuclear localization signal (NLS) and other phosphorylation regulatory domains at the vicinity of the NLS. PMID- 15284484 TI - Characterization of Tula virus from common voles (microtus arvalis) in Poland: evidence for geographic-specific phylogenetic clustering. AB - Tula virus (TULV), a recently identified arvicolid rodent-borne hantavirus, is harbored by the European common vole (Microtus arvalis) in Central Russia and the Czech and Slovak Republics. We report the isolation and characterization of this hantavirus from M. arvalis captured in Poland, a country where human disease caused by hantaviruses has not been recognized. Of 34 arvicolid rodents (24 Clethrionomys glareolus, 9 M. arvalis, 1 Pitymys sp.) captured in Lodz and Tuszyn, Poland, during June to September 1995, sera from 3 M. arvalis and 3 C. glareolus contained IgG antibodies to Puumala virus (PUUV), as determined by an indirect immunofluorescent antibody assay. Alignment and comparison of the 1852 nucleotide S segment and a 1676-nucleotide region of the G2 glycoprotein-encoding M segment, amplified from lung tissues of two hantavirus-seropositive M. arvalis, revealed 83.9-85.2% and 82.3-83.5% sequence similarity, respectively, with TULV strains from Central Russia and the Czech and Slovak Republics. A > 98% sequence conservation was found at the amino acid level. Phylogenetic analysis indicated that the newly found TULV strains from Poland were closely related to, but distinct from, TULV from elsewhere in Europe. PMID- 15284485 TI - Extended sequence analysis of three Danish potato mop-top virus (PMTV) isolates. AB - The entire nucleotide sequence for the coding regions of a Danish PMTV isolate 54 15 was determined and compared to other known and sequenced isolates of PMTV. Many nucleotide and amino acid changes were found in parts of RNA coding for the triple gene block (TGB) proteins and in the part of the RNA coding for the read through region of the coat protein (CP). These regions for two other isolates, the mild one 54-10 and the severe one 54-19, were sequenced. Only two amino acid changes were found to correlate with the subdivision of isolates according to symptom development into mild and severe subgroups. In addition, the phylogenetic tree was obtained suggesting the closest relationship between isolates 54-15 and 54-10. Although the sequence comparisons indicate a high genetic stability of PMTV populations, a surprising change was found in the newly sequenced isolates- the replacement of the AUG start codon of the fourth gene of the TGB encoding RNA, coding for a cystein-rich protein, by the less efficient GUG start codon. PMID- 15284486 TI - Adenovirus expressing a bioluminescence reporter gene and cMAGI cell assay for the detection of HIV-1. AB - We report a fast, highly sensitive method for detecting and testing drug resistance of M-tropic and T-tropic laboratory and primary HIV-1 isolates. cMAGI cells are infected with an adenovirus vector harboring the luciferase reporter gene controlled by HIV-1 Tat-responsive element, TAR. HIV-1 Tat production by HIV 1 chronically infected cells, or by cMAGI cells as early as two days after being acutely infected with HIV-1, is readily monitored in the presence or absence of antiviral drugs. This method is more sensitive than HIV-1 Tat dependant production of beta-galactosidase in the cMAGI cells. The fast answer, ease and sensitivity as well as the possibility of using this method in high throughput screening, makes it an very attractive tool for phenotypic detection of HIV-1 in clinical samples as well as a sensitive assay for monitoring drug resistant HIV-1 variants. This method can also be used for discovery of novel anti HIV-1 drugs. PMID- 15284487 TI - The reading frame BPLF1 of Epstein-Barr virus: a homologue of herpes simplex virus protein VP16. AB - The open reading frame BPLF1 of Epstein-Barr virus (EBV) shows homology to the Herpes simplex virus 1 (HSV1) protein VP16. This protein is a structural tegument component playing a pivotal role for HSV replication as trans-activator of viral immediate-early genes. An EBV gene with a comparable function has not been described so far. However, computer analysis indicated that BPLF1 may be a tegument protein homologous to VP16. This is the first report on the characterisation of the BPLF1 gene, its transcription, and expression of its gene product in vitro and in vivo. Using RT-PCR and Northern blot assays we demonstrated that the BPLF1 gene belongs to the class of late lytic cycle genes of EBV. Besides a full length transcript of 9.5 kb also a polyadenylated transcript of approximately 3 kb is synthesised. However, no consensus splice sites could be identified. Northern blot experiments using partially overlapping probes and sequencing of a BPLF1-specific cDNA revealed 1,550 nucleotides of the BPLF1 transcript, collinear in sequence with the viral genome from position 64547 to 66097. A recombinant Western blot assay detected BPLF1-specific antibodies in seropositive individuals, in particular in cases with elevated viral replication like infectious mononucleosis, chronic active infection, and nasopharyngeal carcinoma. This demonstrated expression of the BPLF1 protein in vivo. Thus, experimental data and computer analysis strongly support the hypothesis of BPLF1 being a tegument protein of the EBV homologous to VP16 of HSV1 and ORF22 of Varicella zoster virus. PMID- 15284488 TI - Extensive variation of sequence within isolates of Grapevine virus B+. AB - Four regions covering 1247 nucleotides of the RNA genome of 20 isolates of a Vitivirus, Grapevine virus B (GVB), from three countries were analyzed. All the regions in these isolates varied in sequence as compared to the published GVB sequence. Of these, the intergenic region varied the most, with 73.2% nucleotide sequence homology, while ORF4 encoding coat protein varied the least when compared both at nucleotide sequence (80.3% homology) and at amino acid sequence levels (90.6% homology). The variations were scattered along each region length and were higher at the nucleotide level than at the amino acid level, but none resulted in a frame shift or stop codon. These results indicate that GVB may exist as a heterogeneous population, possibly resulting from mixing different strains by grafting practices or by RNA-RNA recombination in the grapevine, the only known natural host of this virus. Although it has been reported that GVB is associated with corky bark disease, no corky bark symptoms were observed in any of the GVB positive grapevine sample collected from Australia. PMID- 15284489 TI - Purification, cloning and functional expression of hydroxyphenylpyruvate reductase involved in rosmarinic acid biosynthesis in cell cultures of Coleus blumei. AB - Hydroxyphenylpyruvate reductase (HPPR) is an enzyme involved in the biosynthesis of rosmarinic acid in Lamiaceae reducing hydroxyphenylpyruvates in dependence of NAD(P)H to the corresponding hydroxyphenyllactates. The HPPR protein was purified from suspension cells of Coleus blumei accumulating high levels of rosmarinic acid by ammonium sulfate precipitation, anion exchange chromatography, hydroxylapatite chromatography, chromatography on 2',5'-ADP-Sepharose 4B and SDS polyacrylamide gel electrophoresis. The protein was tryptically digested and the peptides sequenced. Sequence information was used to isolate a full-length cDNA clone for HPPR (EMBL accession number AJ507733) by RT-PCR, screening of a C. blumei cDNA-library and 5'-RACE-PCR. The open reading frame of the HPPR-cDNA consists of 939 nucleotides encoding a protein of 313 amino acid residues. The sequence showed that HPPR belongs to the family of D-isomer-specific 2 hydroxyacid dehydrogenases. The HPPR-cDNA was heterologously expressed in Escherichia coli and the protein was shown to catalyse the NAD(P)H-dependent reduction of 4-hydroxyphenylpyruvate to 4-hydroxyphenyllactate and 3,4 dihydroxyphenylpyruvate to 3,4-dihydroxyphenyllactate. PMID- 15284490 TI - Rice mutant resources for gene discovery. AB - With the completion of genomic sequencing of rice, rice has been firmly established as a model organism for both basic and applied research. The next challenge is to uncover the functions of genes predicted by sequence analysis. Considering the amount of effort and the diversity of disciplines required for functional analyses, extensive international collaboration is needed for this next goal. The aims of this review are to summarize the current status of rice mutant resources, key tools for functional analysis of genes, and our perspectives on how to accelerate rice gene discovery through collaboration. PMID- 15284491 TI - Isolation and characterization of delta(6)-desaturase, an ELO-like enzyme and delta(5)-desaturase from the liverwort Marchantia polymorpha and production of arachidonic and eicosapentaenoic acids in the methylotrophic yeast Pichia pastoris. AB - The liverwort Marchantia polymorpha contains high proportions of arachidonic and eicosapentaenoic acids. In general, these C20 polyunsaturated fatty acids (PUFA) are synthesized from linoleic and alpha -linolenic acids, respectively, by a series of reactions catalyzed by Delta(6)-desaturase, an ELO-like enzyme involved in Delta(6) elongation and Delta(5)-desaturase. Here we report the isolation and characterization of the cDNAs, MpDES6, MpELO1 and MpDES5, coding for the respective enzymes from M. polymorpha. Co-expression of the MpDES6, MpELO1 and MpDES5 cDNAs resulted in the accumulation of arachidonic and eicosapentaenoic acids in the methylotrophic yeast Pichia pastoris. Interestingly, Delta(6) desaturation by the expression of the MpDES6 cDNA appears to occur both in glycerolipids and the acyl-CoA pool, although other lower-plant Delta(6) desaturases are known to have a strong preference for glycerolipids. PMID- 15284492 TI - Members of a new group of chitinase-like genes are expressed preferentially in cotton cells with secondary walls. AB - Two homologous cotton (Gossypium hirsutum L.) genes, GhCTL1 and GhCTL2, encode members of a new group of chitinase-like proteins (called the GhCTL group) that includes other proteins from two cotton species, Arabidopsis, rice, and pea. Members of the GhCTL group are assigned to family GH19 glycoside hydrolases along with numerous authentic chitinases (http://afmb.cnrs-mrs.fr/CAZY/index.html), but the proteins have novel consensus sequences in two regions that are essential for chitinase activity and that were previously thought to be conserved. Maximum parsimony phylogenetic analyses, as well as Neighbor-Joining distance analyses, of numerous chitinases confirmed that the GhCTL group is distinct. A molecular model of GhCTL2 (based on the three-dimensional structure of a barley chitinase) had changes in the catalytic site that are likely to abolish catalytic activity while retaining potential to bind chitin oligosaccharides. RNA blot analysis showed that members of the GhCTL group had preferential expression during secondary wall deposition in cotton lint fiber. Cotton transformed with a fusion of the GhCTL2 promoter to the beta -d-glucuronidase gene showed preferential reporter gene activity in numerous cells during secondary wall deposition. Together with evidence from other researchers that mutants in an Arabidopsis gene within the GhCTL group are cellulose-deficient with phenotypes indicative of altered primary cell walls, these data suggest that members of the GhCTL group of chitinase-like proteins are essential for cellulose synthesis in primary and secondary cell walls. However, the mechanism by which they act is more likely to involve binding of chitin oligosaccharides than catalysis. PMID- 15284493 TI - Over-expression of an Arabidopsis zinc transporter in hordeum vulgare increases short-term zinc uptake after zinc deprivation and seed zinc content. AB - Increasing the zinc content of cereal grains will be important for improving human nutrition. Improved plant zinc efficiency will lead to increased yields when available zinc is limiting plant growth. The aim of our work was to test how the over-expression of zinc transporters in cereals affects plant growth, seed mineral content, and zinc transport rates. Known zinc transporters from Arabidopsis were over-expressed in Hordeum vulgare cv. Golden Promise by means of a ubiquitin promoter. Multiple transgenic lines were obtained, and the locus number and expression levels were verified. Transgenic lines were tested in long term growth and short-term uptake experiments. Seeds from transgenic lines grown in soil had higher zinc and iron contents than controls. Short-term uptake rates were higher in the transgenic lines after zinc deprivation. Resupply of zinc after a period of deprivation resulted in the rapid decrease in zinc uptake even in transgenic lines in which a zinc transporter gene was constitutively expressed. Similar to processes in yeast and Arabidopsis, we hypothesize that this rapid decrease in zinc transport activity may be caused by the degradation of transporters in response to zinc-sufficient conditions. In the long-term growth experiments, there were no significant differences between transgenic and control lines in leaf zinc content or shoot biomass under zinc-sufficient or deficient conditions. However, root-to-shoot ratios were higher in the transgenic plants grown under low-zinc conditions; this could impact zinc acquisition under field conditions. Increased seed zinc and iron content by over-expression of a zinc transporter provides a new strategy for increasing the micronutrient content of cereals. PMID- 15284494 TI - Evidence that CTR1-mediated ethylene signal transduction in tomato is encoded by a multigene family whose members display distinct regulatory features. AB - Ethylene governs a range of developmental and response processes in plants. In Arabidopsis thaliana, the Raf-like kinase CTR1 acts as a key negative regulator of ethylene responses. While only one gene with CTR1 function apparently exists in Arabidopsis, we have isolated a family of CTR1- like genes in tomato ( Lycopersicon esculentum ). Based on amino acid alignments and phylogenetic analysis, these tomato CTR1- like genes are more similar to Arabidopsis CTR1 than any other sequences in the Arabidopsis genome. Structural analysis reveals considerable conservation in the size and position of the exons between Arabidopsis and tomato CTR1 genomic sequences. Complementation of the Arabidopsis ctr1-8 mutant with each of the tomato CTR genes indicates that they are all capable of functioning as negative regulators of the ethylene pathway. We previously reported that LeCTR1 expression is up-regulated in response to ethylene. Here, quantitative real-time PCR was carried out to detail expression for LeCTR1 and the additional CTR1 -like genes of tomato. Our results indicate that the tomato CTR1 gene family is differentially regulated at the mRNA level by ethylene and during stages of development marked by increased ethylene biosynthesis, including fruit ripening. The possibility of a multi-gene family of CTR1 -like genes in other species besides tomato was examined through mining of EST and genomic sequence databases. PMID- 15284496 TI - Tobacco Nectarin III is a bifunctional enzyme with monodehydroascorbate reductase and carbonic anhydrase activities. AB - Tobacco plants secrete a limited array of proteins (nectarins) into their floral nectar. N-terminal sequencing of the Nectarin II ( NEC2; 35kD) and the Nectarin III ( NEC3; 40kD) proteins revealed that they both share identity with dioscorin, the major soluble protein of yam tubers. These sequences also revealed that NEC2 is a breakdown product of NEC3. Using these N-terminal peptide sequences, degenerate oligonucleotides were designed that permitted the isolation of a partial NEC3 cDNA. This cDNA was then used to probe a nectary specific cDNA library and a full-length NEC3 cDNA clone was isolated. Complete sequence analysis confirmed the identity of NEC3 as a dioscorin-like protein. MALDI-TOF mass spectrometric fingerprinting of tryptic peptides derived from the purified NEC3 confirmed that this protein was encoded by the isolated cDNA. NEC3 was shown to possess both carbonic anhydrase and monodehydroascorbate reductase activities. RT-PCR based expression analyses demonstrated that NEC3 transcript is expressed throughout nectary development as well as in other floral organs. A proposed function in the maintenance of pH and oxidative balance in nectar is discussed. PMID- 15284497 TI - AtToc90, a new GTP-binding component of the Arabidopsis chloroplast protein import machinery. AB - AtToc159 is a GTP-binding chloroplast protein import receptor. In vivo, atToc159 is required for massive accumulation of photosynthetic proteins during chloroplast biogenesis. Yet, in mutants lacking atToc159 photosynthetic proteins still accumulate, but at strongly reduced levels whereas non-photosynthetic proteins are imported normally: This suggests a role for the homologues of atToc159 (atToc132, -120 and -90). Here, we show that atToc90 supports accumulation of photosynthetic proteins in plastids, but is not required for import of several constitutive proteins. Part of atToc90 associates with the chloroplast surface in vivo and with the Toc-complex core components (atToc75 and atToc33) in vitro suggesting a function in chloroplast protein import similar to that of atToc159. As both proteins specifically contribute to the accumulation of photosynthetic proteins in chloroplasts they may be components of the same import pathway. PMID- 15284501 TI - Effect of periradicular methylprednisolone on spontaneous resorption of intervertebral disc herniations. AB - STUDY DESIGN: Prospective comparison of periradicular infiltration with steroid versus saline on the spontaneous resorption of herniated nucleus pulposus in a randomized controlled trial. OBJECTIVES: To evaluate whether periradicular steroid retards the resorption of herniated nucleus pulposus. SUMMARY OF BACKGROUND DATA: Rim enhancement around herniated nucleus pulposus is associated with spontaneous resorption of disc herniations. As rim enhancement consists of a macrophage infiltrate, periradicular steroid could theoretically interfere with the resorption process. METHODS.: Patients with disc herniation-induced sciatica were randomized to receive either periradicular methylprednisolone (in combination with bupivacaine) or saline. Lumbar magnetic resonance imaging (MRI) was performed at baseline, at 2 months, and at 12 months. Disc herniation volume (mm3), coverage of rim enhancement (%), and rim enhancement thickness (mm) were evaluated by a radiologist blinded to the allocation. Operated patients were excluded from the 1-year imaging. Changes in the parameters from baseline to 2 months, and from 2 to 12 months, were evaluated with the Mann-Whitney U test. RESULTS: Change in herniation volume from baseline to 2 months was measurable in 34 patients of both groups, and from 2 to 12 months in 26 patients of the steroid group and 24 patients of the saline group. Significant spontaneous resorption of disc herniations occurred in both groups during the 1-year follow-up. In the subgroup analysis, there tended to be even faster resorption in the steroid group from baseline to 2 months for extrusions, and from 2 months to 12 months for contained herniations. No significant differences were observed in the enhancement parameters (coverage and thickness) between the two treatments. CONCLUSIONS: Periradicular corticosteroid does not have a negative effect on the spontaneous resorption of the herniated nucleus pulposus. PMID- 15284499 TI - Automated SNP detection in expressed sequence tags: statistical considerations and application to maritime pine sequences. AB - We developed an automated pipeline for the detection of single nucleotide polymorphisms (SNPs) in expressed sequence tag (EST) data sets, by combining three DNA sequence analysis programs: Phred, Phrap and PolyBayes. This application requires access to the individual electrophoregram traces. First, a reference set of 65 SNPs was obtained from the sequencing of 30 gametes in 13 maritime pine (Pinus pinaster Ait.) gene fragments (6671 bp), resulting in a frequency of 1 SNP every 102.6 bp. Second, parameters of the three programs were optimized in order to retrieve as many true SNPs, while keeping the rate of false positive as low as possible. Overall, the efficiency of detection of true SNPs was 83.1%. However, this rate varied largely as a function of the rare SNP allele frequency: down to 41% for rare SNP alleles (frequency < 10%), up to 98% for allele frequencies above 10%. Third, the detection method was applied to the 18498 assembled maritime pine (Pinus pinaster Ait.) ESTs, allowing to identify a total of 1400 candidate SNPs, in contigs containing between 4 and 20 sequence reads. These genetic resources, described for the first time in a forest tree species, were made available at http://www.pierroton.inra/genetics/Pinesnps. We also derived an analytical expression for the SNP detection probability as a function of the SNP allele frequency, the number of haploid genomes used to generate the EST sequence database, and the sample size of the contigs considered for SNP detection. The frequency of the SNP allele was shown to be the main factor influencing the probability of SNP detection. PMID- 15284502 TI - Determination of the length of anteromedial screw trajectory by measuring interforaminal distance in the first sacral vertebra. AB - STUDY DESIGN: Measurements of the length of pediculocorporeal screw trajectory were performed on bony sacra. Outer interforaminal distances of the first sacral vertebra were measured on bony sacra and anteroposterior sacral radiographs. OBJECTIVES: To demonstrate the correlation of the length of pediculocorporeal screw trajectory of the first sacral vertebra on bony specimens with outer interforaminal distances on their anteroposterior radiographs. SUMMARY OF BACKGROUND DATA: Posterior fixation of the lumbosacral vertebra has been used in the treatment of unstable lumbar spine disorders. To achieve the strongest biomechanical stability and to avoid injury to anterior sacral structures for anteromedial insertion of the posterior transpedicular screw to the first sacral vertebra (S1), it is crucial to determine the optimum screw length for both unicortical and bicortical screw placement. METHODS: Fifty-one dry bony adult sacra were measured and correlated with anteroposterior lumbosacral radiographs. The length of S1 screw trajectory, which was accepted as between inferolateral border of superior articular facet of S1 and sacral promontory, and the outer interforaminal distances of anterior S1 foramens were measured on bony specimens. The outer interforaminal distances of anterior S1 foramens were also measured on anteroposterior sacral radiographs. RESULTS: The outer interforaminal distances were not statistically different from the length of the S1 screw trajectory on bony specimens. Radiographs of the S1 vertebra have also shown that outer interforaminal distances were not statistically different from the length of the S1 screw trajectory. CONCLUSION: In this study, we described an easy and a reliable method to determine the length of anteromedial screw by measuring outer interforaminal distance of S1 vertebra on anteroposterior radiograph of the sacrum. PMID- 15284498 TI - Sucrose regulates elongation of carrot somatic embryo radicles as a signal molecule. AB - Elongation of carrot somatic embryo radicles was inhibited by sucrose at or above 5% (145 mM). This effect would not be released until the sucrose concentration was lowered again. Morphological and cytological studies as well as determination of ABA content and analysis of the expression mode of a Lea gene, all point to its similarity to natural dormancy and germination of seeds. Use of monosaccharides (glucose and fructose), other disaccharide (maltose), and isomolar concentration of osmotica (mannitol and sorbitol), did not show similar regulatory effect. It is thus clear that the regulatory effect is not a result of simple osmotic stress. Hexokinase inhibitors such as glucosamine and N -acetyl glucosamine did not exert any influence on the regulation-deregulation effects of sucrose. Mannose, which inhibits germination of Arabidopsis seeds, did not prevent carrot somatic embryo radicles from elongating. It is thus inferred that this sucrose-signaling pathway may be independent of hexokinase. As a first step to understand the molecular mechanism of this process, a carrot sucrose transporter gene ( cSUT ) expressed in the embryos and roots specifically was isolated. Studies on transformed yeast mutant with cSUT cDNA identified its sucrose transport activity. Northern hybridization and gel retardation experiment revealed that there is a marked increase in expression of cSUT at the beginning of somatic embryo germination, and this is attributed to regulation on the level of transcription. This suggested the possibility that cSUT has an important role in this sucrose signal regulation system. PMID- 15284503 TI - Comparison of posterolateral lumbar fusion rates of Grafton Putty and OP-1 Putty in an athymic rat model. AB - STUDY DESIGN: Posterolateral lumbar spine fusions in athymic rats. OBJECTIVES: To compare spine fusion rates of two different osteoinductive products. SUMMARY OF BACKGROUND DATA: Many osteoinductive bone graft alternatives are available. Grafton (a demineralized bone matrix [DBM]) and Osteogenic Protein-1 (OP-1, an individual recombinant bone morphogenetic protein) are two such alternatives. The relative efficacy of products from these two classes has not been previously studied. The athymic rat spine fusion model has been validated and demonstrated useful to minimize inflammatory responses to xenogeneic or differentially expressed proteins such as those presented by DBMs of human etiology. METHODS: Single-level intertransverse process fusions were performed in 60 athymic nude rats with 2 cc/kg of Grafton or OP-1 Putty. Half of each study group was killed at 3 weeks and half at 6 weeks. Fusion masses were assessed by radiography, manual palpation, and histology. RESULTS: At 3 weeks, manual palpation revealed a 13% fusion rate with Grafton and a 100% fusion rate with OP-1 (P = 0.0001). At 6 weeks, manual palpation revealed a 39% fusion rate of with Grafton and a 100% fusion rate with OP-1 (P = 0.0007). Similar fusion rates were found by histology at 3 and 6 weeks. Of note, one or two adjacent levels were fused in all of the OP 1 animals and none of the Grafton animals. CONCLUSIONS: Significant differences between the ability of Grafton and OP-1 to induce bone formation in an athymic rat posterolateral lumbar spine fusion model were found. PMID- 15284504 TI - Ultrastructural analysis of metallic debris and tissue reaction around spinal implants in patients with late operative site pain. AB - STUDY DESIGN: Cross-sectional ultrastructural analysis of metallic debris as well as the tissue reaction surrounding spinal implants in patients with late operative site pain. OBJECTIVE: To clarify the cause of late operative site pain by the ultrastructural analysis of the byproducts of metallic corrosion as well as the surrounding soft tissues. SUMMARY OF THE BACKGROUND DATA: Late operative site pain has been identified as the most frequent cause of implant removal. Allergic reaction to metal as well as low-grade infection has been suggested as probable mechanisms. METHODS: Fourteen spinal implants were removed because of late operative site pain. Tissues obtained from different zones surrounding these implants were analyzed by light microscopy, transmission electron microscopy, and scanning electron microscopy along with electron dispersion radiograph analysis. RESULTS: No signs of infection were present. Either one of the 2 types of connective tissue (dense or loose) were found to surround the implants. Macrophage counts were most abundant in zone II (around pedicular screws) when compared to other zones (zone I: around the rods; zone III: around the transverse rod connectors). In contrast, particulate debris was more abundant and larger in size in specimens from zone III. Two types of metallic debris were identified. The rusty appearing particles contained mostly iron (Fe), whereas the black appearing particles were rich in chromium (Cr). CONCLUSION: This study produced useful information regarding the production and distribution of particulate metallic debris around stainless steel spinal implants. PMID- 15284495 TI - Characteristics of the Lotus japonicus gene repertoire deduced from large-scale expressed sequence tag (EST) analysis. AB - To perform a comprehensive analysis of genes expressed in a model legume, Lotus japonicus, a total of 74472 3'-end expressed sequence tags (EST) were generated from cDNA libraries produced from six different organs. Clustering of sequences was performed with an identity criterion of 95% for 50 bases, and a total of 20457 non-redundant sequences, 8503 contigs and 11954 singletons were generated. EST sequence coverage was analyzed by using the annotated L. japonicus genomic sequence and 1093 of the 1889 predicted protein-encoding genes (57.9%) were hit by the EST sequence(s). Gene content was compared to several plant species. Among the 8503 contigs, 471 were identified as sequences conserved only in leguminous species and these included several disease resistance-related genes. This suggested that in legumes, these genes may have evolved specifically to resist pathogen attack. The rate of gene sequence divergence was assessed by comparing similarity level and functional category based on the Gene Ontology (GO) annotation of Arabidopsis genes. This revealed that genes encoding ribosomal proteins, as well as those related to translation, photosynthesis, and cellular structure were more abundantly represented in the highly conserved class, and that genes encoding transcription factors and receptor protein kinases were abundantly represented in the less conserved class. To make the sequence information and the cDNA clones available to the research community, a Web database with useful services was created at http://www.kazusa.or.jp/en/plant/lotus/EST/. PMID- 15284506 TI - Effect of facetectomy on lumbar spinal stability under sagittal plane loadings. AB - STUDY DESIGN: A study using an anatomically accurate finite-element model of a L2 L3 motion segment to investigate the biomechanical effects of graded bilateral and unilateral facetectomies of L3 under flexion and extension loadings. OBJECTIVE: To predict the amount of facetectomy on lumbar motion segment that would cause segmental instability, therefore enhancing the understanding concerning the role of the facet under sagittal loadings. SUMMARY OF BACKGROUND DATA: This study provides a quantitative study on the role of facets in preserving segmental lumbar stability. Previous analytical models lack of three dimensional structural characterization and insufficient element representation for facet joints. METHODS: A validated finite-element L2-L3 model was subjected to sagittal loadings at 7.5 Nm. Effects of ligaments and facets were examined to establish their relative importance on segment response. The effect of iatrogenic changes (graded unilateral and bilateral facetectomy) was then investigated under these loadings to predict the alterations in terms of gross external (angular and coupled) responses, flexibilities, and facet load. RESULTS: This study shows the importance of preserving ligaments to prevent rotational instabilities for motion segment under flexion. The effect of the facetectomy on the motion segment is insignificant under flexion. In extension, unilateral facetectomy and resection on contralateral facet markedly alters the rotational motion and flexibilities as well as coupled motions. Also, unilateral complete facetectomy with resection of less than 100% on contralateral facet generates high facet load. CONCLUSIONS: Clinically, this study suggests that it may be appropriate to incorporate additional stabilization procedure in restoring the spinal strength and stability for surgical intervention of unilateral complete facetectomy and resection on contralateral facet. The exploitation of the finite-element method to simulate clinically related situations permits an improved understanding of lumbar spinal stability to assist in defining clinical expectation for various forms of surgical intervention of the operative procedures. PMID- 15284507 TI - Altered function of lumbar nerve roots in patients with transitional lumbosacral vertebrae. AB - STUDY DESIGN: A retrospective study was conducted on the preoperative neurologic symptoms of patients with lumbar herniated discs. OBJECTIVE: To evaluate the possibility that the muscle innervation pattern and the sensory dermatomes of lumbar nerve roots are altered when a lumbosacral transitional vertebra is present. SUMMARY OF BACKGROUND DATA: In 1962, McCulloch et al suggested with intraoperative recordings that the innervation pattern of the lumbar nerve roots may be altered when a lumbosacral transitional vertebra is present. However, this result was not repeated in the study by Young et al in 1983. METHODS: We retrospectively reviewed the charts of 62 consecutive patients who underwent microdiscectomy for lumbar herniated discs. Lumbarized S1 vertebra was present in 10 of 62 patients (16%). Among these 10 patients, 8 had herniated discs at L5-S1 compressing the S1 nerve root. In the 52 normally configured patients, 22 had herniated discs at L5-S1 compressing the S1 nerve root, and 15 had herniated discs at L4-L5 compressing the L5 nerve root. The preoperative neurologic symptoms caused by the S1 nerve root compression in the patients with lumbarized S1 vertebrae were compared with the symptoms caused by either L5 or S1 nerve root compression in the patients with normal configuration. RESULTS: The distribution of motor weakness caused by the S1 nerve root compression was significantly different between the patients with lumbarized S1 and those with the normal configuration. The motor weakness caused by the S1 root compression in the patients with lumbarized S1 was similar to that of the L5 nerve root compression in the normal configuration. Analysis of sensory symptoms showed similar results. CONCLUSIONS: Our findings suggest that the function of lumbosacral nerve roots is altered in patients with lumbarized S1 so that the S1 nerve root serves the usual function of the L5 nerve root. PMID- 15284509 TI - The flexion-extension profile of lumbar spine in 100 healthy volunteers. AB - STUDY DESIGN: Dynamic lumbar flexion-extension motions were assessed by an electrogoniometer and a videofluoroscopy unit simultaneously. OBJECTIVES: The aims of this study were to assess the motion profile of lumbar spine in different genders and age groups and to assess their differences. SUMMARY OF BACKGROUNDS DATA: The dynamic lumbar flexion-extension motions analysis method has been developed and validated. However, data profile of the spinal motions of healthy volunteers has not been established. METHODS.: A total of 100 healthy volunteers, including 50 men and 50 women, were recruited. They were then divided into four equal groups, following their age ranges of 21 to 30 years, 31 to 40 years, 41 to 50 years, and 51 years and older. Lumbar flexion-extension motion was assessed with an electrogoniometer and videofluoroscopy simultaneously. Radiologic images of the lumbar spine were captured during flexion-extension in 10 degrees intervals. Intervertebral flexion-extension (IVFE) of each vertebral level was calculated. The spinal motion of different genders was compared segment by segment with independent t test. The spinal motion of different age groups was compared with one-way analysis of variance. RESULTS: A linear-liked pattern of the IVFE curves was observed in different genders and age groups. No statistically significant difference in the pattern of motion was found between genders. However, statistically significant difference in the slope of IVFE curves was found at all lumbar levels in subjects whose age was 51 years or older (P < 0.05). CONCLUSIONS: Assessment of motion profile was found to be helpful for the identification of spinal disorders in clinical practice. Because of the normal variation of spinal motion of subjects in different age ranges, interpretation of spinal motion disorders should be careful. Although the sample size in this study was limited, the database generated might be useful to assist the diagnosis of spinal "instability" in the future. PMID- 15284510 TI - Sagittal alignment of the spine and pelvis during growth. AB - STUDY DESIGN: Prospective study of the sagittal plane alignment of the spine and pelvis in the normal pediatric population. OBJECTIVES: To document the sagittal alignment of the spine and pelvis and its change during growth in the normal pediatric population. SUMMARY OF BACKGROUND DATA: Pelvic morphology as well as sagittal alignment of the spine and pelvis in the pediatric population are poorly defined in the literature. METHODS: Five parameters were evaluated on lateral standing radiographs of 180 normal study participants 4 to 18 years of age: thoracic kyphosis, lumbar lordosis, sacral slope, pelvic tilt, and pelvic incidence. Statistical analysis was performed using two-tailed Student t tests and Pearson's coefficients (level of significance = 0.01). RESULTS: The mean thoracic kyphosis, lumbar lordosis, sacral slope, pelvic tilt, and pelvic incidence values were 43.0 degrees, 48.5 degrees, 41.2 degrees, 7.2 degrees and 48.4 degrees, respectively. There was no significant difference between males and females. Thoracic kyphosis, lumbar lordosis, pelvic tilt, and pelvic incidence were found to be weakly correlated with age, while sacral slope remained stable with growth. CONCLUSIONS: Pelvic morphology, as measured by the pelvic incidence angle, tends to increase during childhood and adolescence before stabilizing into adulthood, most likely to maintain an adequate sagittal balance in view of the physiologic and morphologic changes occurring during growth. Pelvic tilt and lumbar lordosis, two position-dependent parameters, also react by increasing with age, most likely to avoid inadequate anterior displacement of the body center of gravity. Sacral slope is achieved with the standing posture and is not further significantly influenced by age. These results are important to establish baseline values for these measurements in the pediatric population, in view of the reported association between pelvic morphology and the development of various spinal disorders such as spondylolisthesis and scoliosis. PMID- 15284511 TI - Predicting persistent neck pain: a 1-year follow-up of a population cohort. AB - STUDY DESIGN: A population cohort study to determine the 1-year persistence of neck pain. OBJECTIVES: The aim of this study was to determine the persistence of neck pain over a 12-month period among the general adult population and to explore socio-demographic, health-related, occupational, physical, and lifestyle factors that might be linked to such persistence. SUMMARY OF BACKGROUND DATA: Musculoskeletal clinicians report that neck patients frequently return to consult for recurring episodes of pain. However, the persistent nature of neck pain has been less researched than other common chronic pain syndromes. METHODS: First, to identify a cohort of current neck pain sufferers, a baseline cross-sectional survey was conducted in a general population of 7,669 adults, 18 to 75 years of age, registered with two primary care practices in South Manchester, UK. The second phase was a follow-up survey, 12 months later, to determine the 1-year persistence of neck pain among those who had reported neck pain at baseline. Persistence of neck pain was compared across groups of responders stratified by potential prognostic factors measured at baseline. "Persistent" neck pain was defined according to shading within the region of the neck on a blank body mannequin. The term "persistent" neck pain could therefore reflect chronic, recurrent, or continuous pain. RESULTS: There were 1,359 neck pain responders in the baseline survey, and these subjects formed the study population for the prospective study. At follow-up, 786 (58%) subjects responded, of whom 48% reported having neck pain lasting for more than 1 day, during the previous month. Significant baseline characteristics, which independently predicted persistent neck pain, were age (odds ratio [OR] = 3.4), being off work at the time of the baseline survey (OR = 1.6), comorbid low back pain (OR = 1.6), and cycling as a regular activity (OR = 2.4). CONCLUSION: Among the general population, neck pain persists at 12 months in around half of those who report neck pain at the start of the period. An increased risk of persistent neck pain was associated with age 45 to 59 years and low back pain, and also with cycling. The link with psychological distress and the absence of a link with occupational factors compares with other previous findings for common musculoskeletal syndromes in the community. PMID- 15284512 TI - Clinical symptoms in lumbar disc herniations and their correlation to the histological composition of the extruded disc material. AB - STUDY DESIGN: Fifty-five consecutive patients undergoing microdiskectomy due to lumbar disc herniation were included in this clinical study over 12 months. OBJECTIVES: To investigate possible correlations between the histologic composition of the herniated disc fragments and pain, disability, clinical signs, and operative findings. SUMMARY OF BACKGROUND DATA: Previous studies have investigated the histologic composition of herniated lumbar disc fragments. Few publications, however, examined correlations with clinical data. METHODS: Before treatment, patients were examined using a standardized clinical protocol; subjective disability and pain were assessed by the Oswestry Disability Questionnaire and the McGill Pain Questionnaire. The herniated disc fragments were examined semiquantitatively for the relative percentages of nucleus pulposus, anulus fibrosus, and cartilaginous endplate. RESULTS: In patients less than 30 years of age, significantly higher percentages of nucleus pulposus were found than in the older group, whereas anulus fibrosus was found in significantly higher percentages in patients > or =30 years. Both higher percentages of cartilage and nucleus pulposus correlated with increased pain intensity values from the McGill Pain Questionnaire. Impaired reflexes before treatment occurred significantly more often in patients with > or =20% of cartilage in the herniated fragments. If nucleus pulposus was <30%, sensory impairment tended to be more severe before treatment. CONCLUSION: The histologic composition of the herniated disc fragments seems to affect pain and clinical symptoms. PMID- 15284513 TI - Predictive factors for neck and shoulder pain: a longitudinal study in young adults. AB - STUDY DESIGN: A longitudinal study. OBJECTIVES: To estimate the prevalence and incidence of neck and shoulder pain in young adults and to identify the associated and predictive factors of neck and shoulder pain based on 7-year follow-up. SUMMARY OF BACKGROUND DATA: Several work-related, psychosocial, and sociodemographic factors have been verified as being related to neck and shoulder pain in adult populations, but far fewer longitudinal studies concerning the topic have been carried out in young populations. METHODS: A random sample of 826 high school students was investigated when they were 15 to 18 years old and again at 22 to 25 years of age. Altogether, 394 (48%) patients participated in both surveys. The outcome variable was weekly neck and shoulder pain during the past 6 months in adulthood, and the explanatory variables included some sociodemographic factors, leisure time activities, self-assessed physical condition, psychosomatic stress symptoms, and symptoms of fatigue and sleep difficulties. RESULTS: In 7 years, the prevalence of weekly neck and shoulder pain increased from 17% to 28%. Among those who were asymptomatic at baseline, 6-month incidence of occasional or weekly neck and shoulder pain was 59% 7 years later. In an adjusted model, psychosomatic symptoms remained an associated factor for prevalent neck and shoulder pain 7 years later for both females and males. In females, neck and shoulder pain in adolescence was associated with prevalent neck and shoulder pain in adulthood, and sports loading dynamically in the upper extremities was an associated factor for a low prevalence of neck and shoulder pain 7 years later. In separate analyses of incident neck and shoulder pain, psychosomatic stress symptoms predicted neck and shoulder pain in adulthood. CONCLUSIONS: In young adults, the incidence of neck and shoulder pain is high, and the associated factors of neck and shoulder pain are already multifactorial in a young population. PMID- 15284514 TI - Health-related quality of life after surgical treatment for lumbar stenosis. AB - STUDY DESIGN: We performed a retrospective study using the Short Form-36 Health Survey questionnaire (SF-36), clinical examination, and neuroradiologic and neurophysiologic measurements. OBJECTIVES: To evaluate patient outcomes, health related quality of life (HRQoL), and clinical and neurophysiologic picture in a follow-up study of surgery for lumbar stenosis (LS). SUMMARY OF BACKGROUND DATA: In LS patients, clinical, neuroradiologic, and neurophysiologic findings were not related with validated measurement of the outcomes that are more relevant to patients such as functional status and symptoms. METHODS: Thirty patients surgically treated for LS were recontacted and evaluated by means of self administered questionnaires (SF-36), clinical examination, and neuroradiologic and neurophysiologic measurements. Preoperative and follow-up clinical and neurophysiologic findings were registered. Relations between patient-oriented data and validated conventional clinical, neuroradiologic, and neurophysiologic measurements were evaluated. RESULTS: A comparison between preoperative and postoperative clinical picture shows an improvement of most parameters tested. A comparison between preoperative and postoperative neurophysiologic picture shows a worsening of most parameters tested. A comparison between the current sample and Italian normative data for the SF-36 shows a worsening of physical aspects of health-related quality of life; conversely, there is an improvement of some mental domains. CONCLUSIONS: Follow-up evaluation of surgical treatment for LS showed a mild impairment of physical aspects of HRQoL, as measured by patient oriented evaluation. Clinical examination findings showed significant improvement. Conversely, neurophysiologic follow-up showed a discordant outcome. We think that, to better assess the surgical indication, further study should be performed focused on natural history and the association between neurophysiologic evolution and patient outcome, etc. PMID- 15284516 TI - The Spanish version of the SRS-22 patient questionnaire for idiopathic scoliosis: transcultural adaptation and reliability analysis. AB - STUDY DESIGN: Validation of the transcultural adaptation of a questionnaire for measuring health-related quality of life. OBJECTIVES: To translate and culturally adapt the SRS-22 questionnaire to Spanish. To determine the metric qualities (internal consistency and test-retest reproducibility) of this questionnaire. SUMMARY OF BACKGROUND DATA: The SRS-22 Patient Questionnaire has proven to be a valid instrument for clinical assessment of patients with idiopathic scoliosis. The widespread use of the SRS-22 in non-English-speaking countries requires its transcultural adaptation. METHODS: Transcultural adaptation of the SRS-22 was carried out according to the International Quality of Life Assessment Project guidelines and included two translations and two back-translations of the material. A committee of experts decided on the final version. The questionnaire was administered to 175 individuals (152 women and 23 men) with idiopathic scoliosis. The mean age of the participants at the time they received the questionnaire was 18.9 years, thoracic curve magnitude was 28.8 degrees, and lumbar curve magnitude was 28.1 degrees. At this time, 85 patients had been treated surgically, 45 had been treated with orthesis, and 45 were under observation. A subgroup of 30 patients completed the questionnaire a second time 1 week later. Internal consistency was determined with Cronbach's alpha coefficient and test-retest reliability with the intraclass correlation coefficient. RESULTS: The overall alpha coefficient of the questionnaire was 0.89. Coefficients for individual domains were as follows: function/activity, 0.67; pain, 0.81; mental health, 0.83; self-image, 0.73; and satisfaction, 0.78. The questionnaire as a whole had an intraclass correlation coefficient of 0.96. Intraclass correlation coefficients for individual domains were as follows: pain, 0.93; function, 0.82; self-image, 0.94; mental health, 0.94; and satisfaction, 0.98. CONCLUSIONS: The Spanish version of the SRS-22 Patient Questionnaire demonstrated adequate internal consistency for the majority of domains and excellent reproducibility. These results suggest that the process of adaptation has produced an instrument that is apparently equivalent to the original and suitable for clinical research. PMID- 15284517 TI - Endoscopic lateral transpsoas approach to the lumbar spine. AB - STUDY DESIGN: A description of a novel surgical approach to the lumbar spine and a prospective evaluation of the early surgical outcomes. OBJECTIVES: Describe the early postoperative results and the operative technique of a new, minimally invasive transpsoas approach for anterior fusion of the lumbar spine that minimizes the risk to large vessels and other critical structures. SUMMARY OF BACKGROUND DATA: Standard anterior endoscopic approaches to the lumbar spine require mobilization of the great vessels and sympathetic plexus. Vascular injury and retrograde ejaculation are complications clearly associated with this approach. A retroperitoneal, transpsoas approach to the lumbar spine may reduce these risks. METHODS: From 1996 to 2002, 21 patients (13 females, 8 males; mean age 50.0 years) underwent an endoscopic, retroperitoneal transpsoas approach for exposure of the lumbar spine. Surgical indications included discogenic pain in 14 patients, spinal instability at a level adjacent to a previous fusion in 3 patients, and progressive degenerative scoliosis in 4 patients. Data were reviewed to document the early postoperative results for this procedure. Illustrations were created to clearly describe this approach. RESULTS: Average operative time for the single level cases was 149 minutes (range 120-170 minutes); blood loss was 150 cc (range 50-650); postoperative hospital stay was 4.1 days. At long-term follow-up, visual analogue scale scores had decreased an average of 5.9. Mean follow-up was 3.1 years (range 2 months-6.0 years). Six patients (30%) experienced paresthesias in the groin/thigh region. Five of these same patients also complained of groin/thigh pain (27%). Two patients had symptoms that lasted longer than 1 month. One patient was converted to a mini open lateral approach. There were no vascular injuries. CONCLUSIONS: Early results show the endoscopic lateral transpsoas approach to the lumbar spine to be a safe, minimally invasive method for anterior fusion of the first through the fourth lumbar vertebrae. Although there is a risk of groin/thigh numbness or pain, and these symptoms are mostly transient. This approach allows for exposure of the lumbar spine without mobilization of the great vessels or sympathetic plexus. PMID- 15284518 TI - The pathology of ligamentum flavum in degenerative lumbar disease. AB - STUDY DESIGN: A pathologic study of the ligamentum flavum in degenerative lumbar disease. OBJECTIVES: To elucidate the clinical significance of each pathologic finding of the ligamentum flavum. SUMMARY OF BACKGROUND DATA: In many reports, researchers observed the ligamentum flavum removed partially during surgery and did not evaluate the whole image of the ligamentum flavum. In addition, there are only a few reports that examined the possible association between various histologic findings and clinical findings. And, thus, there are many unclear points in the clinical significance indicated by each pathologic finding. METHODS: The study participants were 50 patients with degenerative lumbar diseases who underwent surgical decompression with removal of the ligamentum flavum of the affected spinal level. Tissue specimens of the removed ligamentum flavum in cross section were prepared, and changes in the elastic fibers and collagen fibers were evaluated in three grades to evaluate the whole image. In addition, we observed the presence or absence of any focal lesions and statistically analyzed the possible association between these histologic findings and clinical symptoms or image findings. RESULTS: In regard to the association between histologic findings and clinical symptoms or image findings, calcification was observed in significantly older patients, who tended to have low scores in preoperative JOA score, and was frequently observed in patients with cauda equina symptoms. Patients with ossification had a significantly greater % slip, and chondroid cells were frequently observed in patients with spondylolisthesis. CONCLUSION: Various pathologic findings provided important foundations for discussing the pathogenesis of lesions in ligamentum flavum. Calcification was frequently observed in elderly patients and those with cauda equina symptoms, and these patients tended to have severer preoperative symptoms. Chondroid cells were frequently observed in patients with spondylolisthesis, and patients with ossification had a greater % slip, suggesting involvement of mechanical load in ossification of ligaments. The pathologic findings were significantly related to the clinical features, and these findings will be profitable for understanding the pathogenesis of degenerative lumbar disease. PMID- 15284519 TI - Unilateral limited laminectomy as the approach of choice for the removal of thoracolumbar neurofibromas. AB - STUDY DESIGN: The paper reports a minimally invasive approach to the dorsolumbar spine for the removal of neurofibromas. OBJECTIVES: Demonstrating that a limited unilateral approach is the one of choice for this kind of tumors. SUMMARY OF BACKGROUND DATA: Spinal intradural extramedullary tumors are generally removed by single-level or multilevel laminectomy with midline dural incision. Cases of delayed postoperative kyphosis and spinal instability (6%) may be reduced by unilateral microsurgery, causing minimum damage to ligaments and joints. METHODS: Ten patients with dorsolumbar neurofibroma were operated on between June 2000 and June 2002. There were 5 males (all with lumbar) and 5 females (2 with lumbar and 3 with inferior dorsal neurofibromas). One female had 3 lumbar tumors and required two operations. Surgery was performed in the prone position with a unilateral approach, sparing the joint and the ligamentum interspinosum. The dura was opened paramedially and the tumor dissected from the root and removed in one piece when possible. Water-tight dural closure was done with 5-0 or 6-0 stitches. RESULTS: All the patients were mobilized on day 2 and discharged on day 4 or day 5. No complications resulting from the technique were observed. Static and dynamic plain radiograph films showed that none of them had kyphosis and/or instability 6 months postoperatively. Neurologic results were good. CONCLUSIONS: Hospital stay may be reduced and stability may be preserved with an appropriate microsurgical technique. The technique reported in the paper should thus become the one of choice and extended to other spinal intradural extramedullary tumors. PMID- 15284520 TI - Necessity of rib head resection for anterior discectomy in the thoracic spine. AB - STUDY DESIGN: The relation between the rib head and the thoracic disc was investigated anatomically. OBJECTIVES: To clarify the necessity of rib head resection in thoracoscopic discectomy using the anterior approach. SUMMARY OF BACKGROUND DATA: When using the transthoracic anterior approach, the rib head must often be resected. However, there are no reports in which the relation between the rib head and the interspinal disc has been investigated. METHODS: The distance between the inferior margin of the superior vertebral body and the superior margin proximal to the rib (hereafter, "rib index") was measured. RESULTS: The rib index shows negative value in T2-T9 levels, while zero in Tl0 level and positive value T11-T12. The rib index of the fifth to ninth ribs in men was significantly smaller than those in women. CONCLUSIONS: The surgeon should anticipate full removal of the rib head if operating at T9 and only partial resection below that level. PMID- 15284522 TI - Re: Tan M, Wang H, Wang Y, et al: Morphometric evaluation of screw fixation in atlas via posterior arch and lateral mass. Spine. 2003; 28:888-95. PMID- 15284523 TI - Re: Dong Y, Xia Hong M, Jianyi L, et al: Quantitative anatomy of the lateral mass of the atlas. Spine. 2003; 28:860-3. PMID- 15284524 TI - The effect of static torsion on the compressive strength of the spine: an in vitro analysis using a porcine spine model. AB - STUDY DESIGN: Matched porcine cervical spine motion segments were subjected to two main conditions and compared: axial compression and axial compression combined with varying axial torque. OBJECTIVES: To determine the effect of torsion on the acute compressive strength of the spine. SUMMARY OF BACKGROUND DATA: The spine is often subjected to compression together with axial torque as a component of complex loading, yet there is a lack of documentation on its effect on the compressive strength and injury mechanics. METHODS: Matched cohorts of porcine cervical spine (C5-C6) motion segments were compressed to failure at a rate of 3,000 N/s combined with 0 Nm, 5 Nm, 20 Nm, or 30 Nm of axial torque. Three "failure" points were recorded from the stress/strain association: the first "step" (initial microfracture), the initial slope change (yield point or "slow crush" mechanism), and the ultimate failure point (fracture). Furthermore, resultant injuries were documented using planar radiography and visual inspection following dissection of the motion segments. RESULTS: Axial torque affected the failure characteristics during acute compressive loading. The ultimate strength of the motion segments was significantly reduced with increasing static torques. The compressive load at which initial microfracture occurred, indicated by the first "step" in the load-deformation curve, was increased with 5 Nm, 10 Nm, and 20 Nm of applied torsion in comparison to no torque, but this effect was reduced with 30 Nm of torque. The "slow crush" mechanism of failure was not affected by the addition of axial torque. No radiographic gross injuries to the facet joints were observed. Damage appeared to be confined to the endplate and trabecular network of the vertebral body. CONCLUSIONS: Based on this animal model, shown to have similar biomechanical behavior to humans, axial torque appears to significantly reduce the compressive strength of the spine. PMID- 15284525 TI - Clotting parameters and thromboelastography in children with neuromuscular and idiopathic scoliosis undergoing posterior spinal fusion. AB - STUDY DESIGN: An unblinded comparison of laboratory values between patients with idiopathic and neuromuscular scoliosis. OBJECTIVES: To compare standard tests of coagulation and thromboelastography (TEG) parameters between two groups of patients undergoing posterior spinal fusion (PSF). SUMMARY OF BACKGROUND DATA: Children with neuromuscular scoliosis such as cerebral palsy have more intraoperative blood loss than children with idiopathic scoliosis during PSF. Various reasons suggested for this include nutritional deficiencies, altered tissue integrity, hepatic dysfunction, and use of antiepileptic medications that can cause poor hemostasis and altered coagulation. We have observed alterations in coagulation factor levels in patients with cerebral palsy due to spastic quadriplegia with moderate blood volume loss (25% estimated blood volume). METHODS: In a prospective analysis, we compared standard tests of coagulation (prothrombin time [PT], partial thromboplastin time [PTT], platelet count, fibrinogen levels) and TEG at baseline and at a blood loss of 15% estimated blood volume in patients with idiopathic scoliosis and cerebral palsy undergoing PSF. RESULTS: There were no differences between the groups in terms of gender distribution and age. There was a significant difference between the baseline PT and PTT values, although both groups were within laboratory norms. After 15% blood volume loss, there were differences seen in the PT, PTT, maximum amplitude on the TEG, ionized calcium, and serum magnesium levels (P < 0.05). CONCLUSIONS: Children with cerebral palsy undergoing PSF have increased bleeding that starts earlier in the procedure than it does for patients with idiopathic scoliosis undergoing PSF. We found that, even though children with spastic quadriplegia had baseline PT and PTT values within normal limits, they were significantly different when compared with normal patients. PMID- 15284526 TI - Soft tissue neck symptoms following high-energy road traffic accidents. AB - STUDY DESIGN: A single-blinded prospective study was performed into the occurrence and frequency of soft tissue neck symptoms of patients involved in high-energy road traffic accidents. OBJECTIVES: We studied the occurrence of incidental soft tissue neck symptoms in victims of high energy (defined as those severe enough to cause major trauma leading to an injury severity score >16) vehicular collisions causing significant musculoskeletal trauma requiring operative intervention. SUMMARY BACKGROUND DATA: Whiplash is considered to be a soft tissue injury of the neck sustained by occupants of motor vehicles. Use of the term in the past had been restricted to hyperextension injuries following a rear impact, but is commonly now used for all types of impact. No relationship has been found between velocity or force of injury and incidence or outcome of whiplash. The preponderance of whiplash after relatively minor vehicular accidents, the unpredictability of who will develop chronic symptoms and the lack of clinical and radiological evidence of a pathological mechanism suggests that psychosocial variables are important factors in determining the development of persistent neck pain. METHODS: A total of 36 consecutive patients were recruited who had been involved in high-energy road traffic accidents and had chest, musculoskeletal, or abdominal injuries (ISS > 16) requiring admission for treatment, but who had no diagnosed injury of the cervical spine. Patients were asked in a nonspecific or leading manner at the time of admission and again at least 6 to 8 weeks postinjury if they had any neck symptoms, headaches, or paresthesiae. RESULTS: Only 2 of the patients interviewed described any whiplash symptoms. All symptoms were resolved at the time of second interview. CONCLUSIONS: Our study demonstrates a surprisingly low incidence of neck symptoms following high-energy road traffic accidents in which patients sustained unrelated injuries requiring treatment. PMID- 15284527 TI - Neglected major vessel injury after anterior spinal surgery: a case report. AB - STUDY DESIGN: This is a case report. OBJECTIVES: To present a case of neglected major vessel injury after anterior spinal surgery. SUMMARY OF BACKGROUND DATA: Vascular complications during spinal surgery are fortunately few in number. The incidence and management of vascular complications during anterior approaches to the thoracolumbar spine are not well known, and it is likely that most acute or delayed vascular injuries and complications are not reported. METHODS: A case that underwent anterior spinal surgery for T12 burst fracture with an iatrogenic injury to the aorta was presented. RESULTS: Paraparesis (Frankel B) was present before last operation and disappeared completely at 30 months of follow-up. CONCLUSIONS: As with any complication, the best treatment is prevention. Careful and meticulous exposure of the involved anatomic area is very important for prevention. Early recognition with rapid treatment of vascular complications can reduce potential morbidity and mortality. PMID- 15284528 TI - Positional occlusion/stasis of vertebral arteries in a case of cervical rheumatoid arthritis presenting with multiple posterior circulation infarcts: a case report with angiographic demonstration. AB - STUDY DESIGN: A case report of rheumatoid arthritis involving cervical spine, who presented with multiple posterior circulation infarcts, and pertinent literature review are presented. OBJECTIVE: To describe a rare positional occlusion/stasis of vertebral arteries, which resolved following application of cervical traction. The likely mechanism of above findings is proposed. SUMMARY OF BACKGROUND DATA: The mechanism of vertebrobasilar insufficiency with changes in head position in patients with rheumatoid arthritis can result from a number of causes; however, bilateral compromise of vertebral arteries during head extension had been described only once. There is no report of vertebral artery angiogram in any individual following application of traction and relief of compromise in extension. METHODS: A 45-year-old man had rheumatoid arthritis for the last 10 years and presented with symptoms of posterior circulation infarcts. Cervical spine radiographs revealed "mobile" atlantoaxial dislocation and atlantoaxial impaction. Magnetic resonance imaging confirmed odontoid erosions, lateral masses destruction, atlantoaxial dislocation, and atlantoaxial impaction. Angiogram showed occlusion of the left vertebral artery and transient stasis of the right vertebral artery distal to foramen transversarium of C2 vertebra in extension position. The left vertebral artery had narrowing in the same segment in the neutral position. Following traction, repeat angiogram showed no occlusion or narrowing of either vertebral artery in any position. Transoral odontoidectomy and occipitocervical fusion were performed. RESULTS: The patient had no fresh deficits following surgery. CONCLUSIONS: We described a rare case of positional occlusion/stasis of vertebral arteries associated with rheumatoid arthritis, in which angiography following cervical traction showed complete resolution. PMID- 15284530 TI - Association of paraoxonase-1 M55L genotype and alcohol consumption with coronary atherosclerosis: the Helsinki Sudden Death Study. AB - High-density lipoprotein (HDL) level is inversely correlated with coronary heart disease risk. Paraoxonase-1 (PON1) is an HDL-associated anti-atherogenic enzyme. The activity of PON1 is affected by the methionine for leucine substitution at position 55 (M55L) and increased during regular moderate alcohol consumption, consistent with increased HDL cholesterol concentration. We related the PON1 M55L genotypes to the extent of atherosclerosis in left anterior descending coronary artery (LAD) in alcohol abstainers (0-1 g of alcohol/day), moderate consumers (1 36 g of alcohol/day) and drinkers (> 36 g of alcohol/day). The study subjects included an autopsy series of total of 700 middle-aged Finnish men from the Helsinki Sudden Death Study. The LAD was stained for fat and the areas covered with fatty streaks and fibrotic and complicated plaques were measured. Data on coronary artery disease risk factors were obtained from relatives or close friends of the deceased. Compared to the LL homozygotes, carriers of the M55 allele tended to have larger areas of atherosclerotic lesions, the size of which decreased dose-dependently by reported alcohol consumption. Moderate consumers carrying the M55 allele had significantly larger complicated plaques compared to the LL homozygotes drinking as much (P = 0.009). Among the M55 allele carriers, drinkers showed significantly smaller areas of fatty streaks compared to abstainers (P = 0.042) and moderate consumers (P < 0.001) (for the PON1 genotype by alcohol interaction, P = 0.078). Similarly, drinkers with the M55 allele also had statistically significantly smaller areas of complicated lesions than moderate consumers with the M55 allele (P < 0.0001) (for the PON1 genotype by alcohol interaction, P = 0.009). The areas of atherosclerotic lesions in LAD appear to be dependent on the amount of alcohol consumption, especially in men carrying the PON1 M55 allele. PMID- 15284531 TI - Human UGT1A6 pharmacogenetics: identification of a novel SNP, characterization of allele frequencies and functional analysis of recombinant allozymes in human liver tissue and in cultured cells. AB - BACKGROUND: UDP-glucuronosyltransferase (UGT) enzymes catalyze the glucuronidation and typically inactivation of endogenous and exogenous molecules including steroid hormones, bilirubin and many drugs. The UGT1A6 protein is expressed predominantly in liver and metabolizes small phenolic drugs including acetaminophen, salicylates and many beta-blockers. Interindividual variation in the capacity of humans to glucuronidate drugs has been observed. RESULTS: We have identified a novel common single nucleotide polymorphism (SNP) in the human UGT1A6 gene resulting in a Ser7Ala change in encoded amino acid. We have further functionally characterized that polymorphism in the context of two previously reported polymorphisms, Thr181Ala and Arg184Ser. These non-synonymous cSNPs define four common haplotypes. Alleles appear with similar frequencies in Caucasian and African-American populations with distributions adhering to Hardy Weinberg equilibrium. UGT1A6 genotype, rate of substrate glucuronidation and level of immunoreactive UGT1A6 protein was determined. A 25-fold variation in the rate of substrate glucuronidation and an 85-fold variation in level of immunoreactive protein were measured. Liver tissue samples that were homozygous for UGT1A6*2 exhibited a high rate of glucuronidation relative to tissues with other genotypes. Biochemical kinetic studies of recombinant UGT1A6 expressed in HEK293 cells indicated that the UGT1A6*2 allozyme, expressed homozygously, had almost two-fold greater activity toward p-nitrophenol than UGT1A6*1 and when expressed heterozygously (UGT1A6*1/*2) it was associated with low enzyme activity. CONCLUSIONS: These data suggest that common genetic variation in human UGT1A6 confers functionally significant differences in biochemical phenotype as assessed in human tissue and cultured cells expressing recombinant allozymes. This genetic variation might impact clinical efficacy or toxicity of drugs metabolized by UGT1A6. PMID- 15284532 TI - Identification of common polymorphisms in the promoter of the UGT1A9 gene: evidence that UGT1A9 protein and activity levels are strongly genetically controlled in the liver. AB - OBJECTIVES: Polymorphisms in UDP-glucuronosyltransferases (UGTs) can influence detoxifying capacities and have considerable therapeutic implications in addition to influence various (patho)physiological processes. UGT1A9 plays a central role in the metabolism of various classes of therapeutic drugs in addition to carcinogens and steroids. The great interindividual variability of UGT1A9 mediated glucuronidation remains poorly explained, while evidence for its genetic origin exists. METHODS: The proximal UGT1A9 promoter was screened for polymorphisms by sequencing and, the contribution of single nucleotide polymorphisms (SNPs) to the variability of UGT1A9 protein levels and activity was evaluated. RESULTS: We confirmed the presence of the -109 to -98 T10 polymorphism and found ten novel SNPs that generated a diversity of haplotypes in two independent populations. In a panel of 48 human liver microsomes, the UGT1A9 expression varied by 17-fold and was significantly correlated with SNPs -275, 331/-440, -665 and -2152. The base insertion T10 reported to increase reporter gene expression in HepG2 cells [] was not linked to -275 and -2152 SNPs and was not associated with changes in UGT1A9 protein levels. Compared to wild-type individuals, there were statistically significant higher glucuronidating activities in livers with the -275 and -2152 using mycophenolic acid and propofol as UGT1A9 substrates, indicating an extensive glucuronidator phenotype associated with these variants. CONCLUSIONS: This is the first study to demonstrate that naturally occurring sequence variations in the UGT1A9 promoter are informative in predicting the levels of protein and glucuronidating activity, providing a potential mechanism for interindividual variation in UGT1A9-mediated metabolism. PMID- 15284533 TI - Beta2-adrenoceptor Thr164Ile polymorphism is associated with markedly decreased vasodilator and increased vasoconstrictor sensitivity in vivo. AB - BACKGROUND: The uncommon Thr164Ile polymorphism of the beta2-adrenoceptor is associated with profoundly altered responses to agonist in vitro; however its effects on vascular responses in vivo are not known. Altered adrenergic vascular sensitivity may contribute to the decreased survival observed in patients with congestive heart failure carrying the Ile164 allele. METHODS AND RESULTS: We used the linear variable differential transformer dorsal hand vein technique to compare vasodilation in response to the beta-adrenergic receptor agonist, isoproterenol, and vasoconstriction in response to the alpha-adrenergic receptor agonist, phenylephrine, in healthy homozygous (Thr164/Thr164) (n = 21) and heterozygous Thr164/Ile164 (n = 5) women. The dose of isoproterenol required to achieve 50% venodilation (geometric mean; 95% CI) was significantly higher in women with the Ile164 allele (82.5 ng/min; 17.3-394 ng/min) than those without (15.8 ng/min; 11-25 ng/min; P = 0.004). The maximum response to isoproterenol was not different (102 +/- 1% and 102 +/- 3%, respectively, P = 0.9). The dose of phenylephrine needed to induce 50% venoconstriction was significantly lower in women with the Ile164 allele (151 ng/min; 42-543 ng/min) than those without (540 ng/min; 350-835 ng/min; P = 0.02). CONCLUSIONS: The Thr164Ile polymorphism of the beta2-adrenergic receptor is associated with a five-fold reduction in sensitivity to beta2 receptor agonist-mediated vasodilation; vasoconstrictor sensitivity is increased. The overall effect of the Thr164Ile polymorphism is to shift the balance of adrenergic vascular tone toward vasoconstriction. This suggests a mechanistic explanation for the clinical observation of decreased survival in patients with congestive heart failure heterozygous for the Thr164Ile polymorphism. PMID- 15284534 TI - Lipid-lowering response to statins is affected by CYP3A5 polymorphism. AB - Individuals expressing the polymorphic CYP3A5 enzyme might show a more than average efficiency in the metabolism of lovastatin, simvastatin and atorvastatin. We studied whether the expression of CYP3A5 is associated with an impaired lipid lowering response to statins in 69 Caucasian patients. Lovastatin, simvastatin and atorvastatin were significantly less effective in CYP3A5 expressors than in non-expressors. The mean serum total cholesterol concentration at 1 year was 23% higher (P = 0.0014) and the mean serum low-density lipoprotein cholesterol concentration was 24% higher (P = 0.036) in subjects possessing the CYP3A5*1 allele (CYP3A5 expressors, n = 7) than in subjects homozygous for the CYP3A5*3 allele (non-expressors, n = 39). The mean percentage reduction in serum total cholesterol from baseline was significantly smaller in CYP3A5 expressors than in non-expressors (17% versus 31%, P = 0.026). No association between hypolipidemic efficacy and CYP3A5 polymorphism was observed among 23 subjects taking statins that are not dependent on CYP3A5 (fluvastatin, pravastatin). These findings suggest that CYP3A5 may be a genetic determinant of interindividual differences in response to certain statins. PMID- 15284535 TI - Discovery of new potentially defective alleles of human CYP2C9. AB - CYP2C9 is a clinically important enzyme, responsible for the metabolism of numerous clinically important therapeutic drugs. In the present study, we discovered 38 single nucleotide polymorphisms in CYP2C9 by resequencing of genomic DNA from 92 individuals from three different racial groups. Haplotype analysis predicted that there are at least 21 alleles of CYP2C9 in this group of individuals. Six new alleles were identified that contained coding changes: L19I (CYP2C9*7), R150H (CYP2C9*8), H251R (CYP2C9*9), E272G (CYP2C9*10), R335W(CYP2C9*11) and P489S (CYP2C9*12). When expressed in a bacterial cDNA expression system, several alleles exhibited altered catalytic activity. CYP2C9*11 appeared to be a putative poor metabolizer allele, exhibiting a three fold increase in the Km and more than a two-fold decrease in the intrinsic clearance for tolbutamide. Examination of the crystal structure of human CYP2C9 reveals that R335 is located in the turn between the J and J' helices and forms a hydrogen-bonding ion pair with D341 from the J' helix. Abolishing this interaction in CYP2C9*11 individuals could destabilize the secondary structure and alter the substrate affinity. This new putative poor metabolizer (PM) allele was found in Africans. A second potentially PM allele CYP2C9*12 found in a racially unidentified sample also exhibited a modest decrease in the Vmax and the intrinsic clearance for tolbutamide in a recombinant system. Further clinical studies are needed to determine the effect of these new polymorphisms on the metabolism of CYP2C9 substrates. PMID- 15284536 TI - Relative impact of covariates in prescribing warfarin according to CYP2C9 genotype. AB - Patients on warfarin anticoagulant therapy demonstrate wide variation in maintenance dose. Patients possessing variants (*2 and *3) of the cytochrome P450 2C9 gene require reduced maintenance doses compared to those having wild-type alleles (*1). Many other clinical factors have been shown to affect warfarin dose as well. To determine the relative impact of CYP2C9 genotype, age, gender, body surface area, concomitant medication, treatment indication and comorbidity, we conducted a retrospective cohort study in 453 patients managed by the anticoagulation service of a large, horizontally integrated, multispecialty group practice. In this largely Caucasian patient population, the CYP2C9 gene frequencies for *1/*1, *1/*2, *1/*3, *2/*2, *2/*3 and *3/*3 were 65.1%, 19.0%, 12.1%, 1.6%, 1.8% and 0.4%, respectively, approximating Hardy-Weinberg equilibrium. Mean maintenance doses for these genotypes were 36.5, 29.1, 23.5, 28.0, 18.1 and 5.5 mg/week, respectively. In univariate analyses, genotype alone accounted for 19.8% of the variability in maintenance dose. Age, body surface area and male gender accounted for 14.6%, 7.5% and 4.7%, respectively, while cardiac valve replacement as the indication for warfarin accounted for 5.4% of the variability. Collectively, these factors accounted for 33.7% of all dosing variability according to multiple regression. These results will help strengthen the mathematical models that are currently being developed for prospective gene based warfarin dosing. PMID- 15284537 TI - Pharmacogenetic roles of CYP2C19 and CYP2B6 in the metabolism of R- and S mephobarbital in humans. AB - OBJECTIVES AND METHODS: We assessed the relationship between the metabolism of R- and S-mephobarbital (MPB) and genetic polymorphisms of cytochrome P450 (CYP) 2C19 and CYP2B6. Nine homozygous extensive metabolizers (homo-EMs, 2C19*1/2C19*1) of CYP2C19, ten heterozygous EMs (hetero-EMs, 2C19*1/2C19*2, 2C19*1/2C19*3) and eleven poor metabolizers (PMs, 2C19*2/2C19*2, 2C19*3/2C19*3, 2C19*2/2C19*3) recruited from a Japanese population, received an oral 200 mg-dose of racemic MPB. Blood and urine samples were collected, and R-MPB, S-MPB and the metabolites, phenobarbital (PB) and 4'-hydroxy-MPB, were measured. Each subject was also genotyped for CYP2B6 gene. RESULTS: The mean area under the plasma concentration-time curve (AUC) of R-MPB was 92-fold greater in PMs than in homo EMs. R/S ratios for AUC of MPB were much higher in PMs than in EMs (homo- and hetero-). The cumulative urinary excretion of 4'-hydroxy-MPB up to 24 h postdose was 21-fold less in PMs than in homo-EMs. The metabolic ratio of AUCPB/(AUCS-MPB + AUCR-MPB) was higher in PMs than in EMs (homo- and hetero-). In addition, this metabolic ratio was lower in the carriers of CYP2B6*6 compared with that in its non-carriers. CONCLUSIONS: Our results indicate that the 4'-hydroxylation of R MPB is mediated via CYP2C19 and that the rapid 4'-hydroxylation of R-MPB results in a marked difference in the pharmacokinetic profiles between R-MPB and S-MPB in the different CYP2C19 genotypic individuals. In addition, a minor fraction of the interindividual variability in PB formation from MPB may be explainable by the CYP2B6*6 allele. PMID- 15284538 TI - A substrate specific functional polymorphism of human gamma-glutamyl hydrolase alters catalytic activity and methotrexate polyglutamate accumulation in acute lymphoblastic leukaemia cells. AB - We found a significant inverse relationship between gamma-glutamyl hydrolase (GGH) activity and the accumulation of long-chain methotrexate polyglutamates (MTXPG4-7) in non-hyperdiploid B-lineage acute lymphoblastic leukaemia (ALL) cells after uniform treatment with high-dose methotrexate (HDMTX) (1 g/m i.v.). To identify genetic polymorphisms that alter the function of human GGH, we sequenced the GGH exons of genomic DNA from children with ALL, who had a 7.8-fold range of GGH activity in their ALL cells at diagnosis. A single nucleotide polymorphism (452C>T, T127I) was found among patients with low GGH activity, but not found in patients with high GGH activity. Computational modelling indicated that the T127I substitution alters the molecular surface conformation at the catalytic cleft-tail on GGH, which is predicted to alter binding affinity with long chain but not short-chain methotrexate polyglutamates. Enzyme kinetic analysis of heterologously expressed GGH revealed a significantly higher Km (2.7 fold) and lower catalytic efficiency (Vmax/Km reduced 67%) of the T127I variant compared to wild-type GGH using long-chain MTXPG5 as substrate, but not a significant change with short-chain MTXPG2. The 452C>T single nucleotide polymorphism (SNP) was also associated with lower GGH activity in hyperdiploid B lineage and T lineage ALL cells. Caucasians [10.0%; 95% confidence interval (CI) 6.7-13.3%; n = 155] were found to have a significantly higher frequency of the Ile allele than African-Americans (4.4%; 95% CI 1.2-7.5%; n = 80) (P = 0.033). These studies demonstrate a substrate specific functional SNP (452C>T) in the human GGH gene that is associated with lower catalytic activity and higher accumulation of long-chain MTX-PG in leukaemia cells of patients treated with HDMTX. PMID- 15284539 TI - MUC4 and ErbB2 expression in squamous cell carcinoma of the upper aerodigestive tract: correlation with clinical outcomes. AB - OBJECTIVES/HYPOTHESIS: Expression of the membrane mucin MUC4 has been associated with a variety of malignancies, including squamous cell carcinoma of the upper aerodigestive tract. MUC4 modulates cell signaling pathways as an intramembrane ligand of ErbB2. The hypotheses of the study were that MUC4 expression would correlate with ErbB2 expression and that MUC4 expression would correlate with clinical outcomes in squamous cell carcinoma of the upper aerodigestive tract. STUDY DESIGN: Retrospective chart review was combined with immunohistochemical analysis of paraffin-embedded tumor specimens from patients treated with initial definitive surgical resection at an academic tertiary care medical center. METHODS: MUC4 and ErbB2 receptor expression was localized by immunohistochemical studies using archival formalin-fixed and paraffin-embedded tissue. A limited number of fresh-frozen tissues were further analyzed by Western blot. Clinical outcomes and histopathological parameters were determined by retrospective chart review and correlated with immunohistochemical findings. RESULTS: One hundred fifty-four patients were analyzed with a median follow-up of 12 months among 54 patients who died and 49 months among 100 surviving patients. Membrane expression of MUC4 and ErbB2 was seen in 12% and 13% of tumors, respectively. MUC4 expression was not correlated with pathological grade. A significant correlation was found between MUC4 expression and ErbB2 expression. Multivariate survival analyses revealed that patients whose tumors exhibited MUC4 membrane expression had statistically significant improvement in survival and longer time to recurrence compared with patients whose tumors did not express MUC4 as defined by immunohistochemical staining patterns. No correlations between ErbB2 expression and survival or recurrence were observed. CONCLUSION: Patients with tumors that retain MUC4 expression exhibit improved survival and decreased recurrence in squamous cell carcinoma of the upper aerodigestive tract. Correlations between MUC4 expression patterns and ErbB2 expression are also observed, suggesting that MUC4-ErbB2 mediated cell signaling pathways may provide insights into this clinical result. PMID- 15284541 TI - Helical computed tomographic angiography: an excellent screening test for blunt cerebrovascular injury. AB - BACKGROUND: Blunt cerebrovascular injury (BCVI) carries a high morbidity and mortality, especially when diagnosis is delayed. Recent studies have shown that increased recognition of these injuries is achieved with prompt screening, allowing for early treatment and better outcome. Controversy still exists, however, on the best screening test. This study was used to evaluate the role of helical computed tomographic angiography (CTA) of the carotid and vertebral arteries in the early screening of BCVI. METHODS: All patients deemed at risk for BCVI underwent CTA within 24 hours of admission. Patients with a negative CTA test underwent no further radiologic evaluation of the cerebral vasculature. Those patients with positive or equivocal CTA results underwent four-vessel cerebral arteriography as a confirmatory test. Data were collected on the radiologic interpretation of all studies and patient clinical course. RESULTS: Four hundred eighty-six patients fulfilled the criteria for screening and underwent CTA. Nineteen patients were diagnosed with 25 BCVIs during the period of study. There were 7 carotid injuries and 18 vertebral injuries. Eighteen of 19 patients with BCVI were screened with CTA. Seventeen patients were asymptomatic at the time of screening. Results of CTA for BCVI were as follows: sensitivity, 100%; specificity, 94.0%; prevalence (screened patients), 3.7%; positive predictive value, 37.5%; and negative predictive value, 100%. Except for one patient in whom the CTA was clearly misinterpreted by the radiologist, no patient with a negative CTA examination was subsequently found to have a missed injury. CONCLUSION: CTA is an excellent test with which to screen for BCVI. PMID- 15284540 TI - The impact of hypoxia and hyperventilation on outcome after paramedic rapid sequence intubation of severely head-injured patients. AB - BACKGROUND: An increase in mortality has been documented in association with paramedic rapid sequence intubation (RSI) of severely head-injured patients. This analysis explores the impact of hypoxia and hyperventilation on outcome. METHODS: Adult severely head-injured patients (Glasgow Coma Scale score of 3-8) unable to be intubated without neuromuscular blockade underwent paramedic RSI using midazolam and succinylcholine; rocuronium was administered after confirmation of tube position. Standard ventilation parameters were used for most patients; however, one agency instituted use of digital end-tidal carbon dioxide (ETCO2) and oxygen saturation (Spo2) monitoring during the trial. Each patient undergoing digital ETCO2/Spo2 monitoring was matched to three historical nonintubated controls on the basis of age, gender, mechanism, and Abbreviated Injury Scale scores for each of six body regions. Logistic regression was used to explore the impact of oxygen desaturation during laryngoscopy and postintubation hypocapnia and hypoxia on outcome. The relationship between hypocapnia and ventilatory rate was explored using linear regression and univariate analysis. In addition, trial patients and controls were compared with regard to mortality and the incidence of "good outcomes" using an odds ratio analysis. RESULTS: Of the 426 trial patients, a total of 59 had complete ETCO2/Spo2 monitoring data; these were matched to 177 controls. Logistic regression revealed an association between the lowest ETCO2 value and final ETCO2 value and mortality. Matched-controls analysis confirmed an association between hypocapnia and mortality. A statistically significant association between ventilatory rate and ETCO2 value was observed (r = -0.13, p < 0.0001); the median ventilatory rate associated with the lowest recorded ETCO2 value was significantly higher than for all other ETCO2 values (27 mm Hg vs. 19 mm Hg, p < 0.0001). In addition, profound desaturations during RSI and hypoxia after intubation were associated with higher mortality than matched controls. Overall mortality was 41% for trial patients versus 22% for matched controls (odds ratio, 2.51; 95% confidence interval, 1.33-4.72; p = 0.004). CONCLUSIONS: Hyperventilation and severe hypoxia during paramedic RSI are associated with an increase in mortality. PMID- 15284542 TI - Real-time intravascular ultrasound-guided placement of a removable inferior vena cava filter. AB - BACKGROUND: Reports have demonstrated the benefit of prophylactic inferior vena cava filter (IVCF) placement to prevent pulmonary embolism. This series evaluates the potential for the bedside placement of a removable IVCF under "real-time" intravascular ultrasound (IVUS) guidance. METHODS: Twenty trauma patients underwent intensive care unit placement of a removable IVCF with IVUS guidance. All patients had ultrasonography of the femoral veins after placement to rule out postprocedure femoral vein thrombosis and radiographs to identify filter location. RESULTS: Nineteen of 20 IVCFs were placed at approximately the L2 level as verified by radiography. One patient had a large IVC (34 mm) and underwent bilateral common iliac IVCF placement under IVUS. Within 3 weeks of placement, 12 IVCFs were retrieved. Of the remaining eight patients, six had indications for permanent implantation, two had contralateral deep venous thrombosis, and one had ipsilateral deep venous thrombosis. CONCLUSION: Bedside insertion of a removable IVCF with IVUS guidance and its removal are simple, safe, and accurate. PMID- 15284543 TI - Bedside insertion of vena cava filters in the intensive care unit using intravascular ultrasound to locate renal veins. AB - BACKGROUND: Historically, contrast venography has been used to determine renal vein location and assist with vena cava filter placement. This technique, however, exposes the patient to nephrotoxic contrast and radiation. For trauma patients in the intensive care unit (ICU), inferior vena cava filters should ideally be placed without contrast at the bedside to avoid nephrotoxic agents, radiation, and transport of a critically injured patient to the operating room or x-ray department. Previously, the authors have shown that intravascular ultrasound is a safe and accurate method for locating renal veins and assisting with vena cava filter placement. The purpose of this study was to evaluate bedside vena cava filter placement prospectively using only intravascular ultrasound for imaging. METHODS: Between August 2000 and July 2003, 29 patients met trauma service criteria for prophylactic or therapeutic placement of a vena cava filter. The 7 females and 22 males had a mean age of 51.3 years (range, 20 92 years), a mean height of 177 cm (range, 160-218.4 cm), a mean weight of 101.9 kg (range, 59.1-186.4 kg), and a body mass index of 33 (range, 14.7-56.1). Fifteen patients (55.5%) had a body mass index exceeding 30. The mean Injury Severity Score was 25.4 (range, 12-45). Intravascular ultrasound was the sole imaging method, and no contrast or fluoroscopy was used. All procedures were performed in the ICU by trauma surgeons. Data collection was prospective and included demographics, injuries, vena caval anatomy, length of procedure, complications, and follow-up radiographic confirmation of appropriate deployment. RESULTS: The location of the renal veins and vena cava diameter was imaged in all the patients. Three patients were noted to have accessory renal veins, and no patient had thrombus in the vena cava. The inferior vena cava diameter was less than 28 mm in all the patients, thus allowing standard filters to be deployed. Filter deployment was successful for all the patients. Of the 29 patients, 27 had abdominal computed tomography (CT) during their hospital stay. When the location of the renal veins identified by CT was compared with the level of the filter on abdominal x-ray, the filter tip was found to be at or below the level of the most caudal renal vein in 26 of the 27 patients (96.3%). In one patient, the filter tip was purposely placed 2 to 3 cm above an accessory caudal renal vein, but below the main right and left renal veins. The mean procedure time was 37.7 minutes (range, 12-86 minutes). No complications were associated with filter placement. CONCLUSIONS: Intravascular ultrasound is a safe and effective imaging method that may be used for the bedside placement of vena cava filters in the ICU. This technique avoids the use of nephrotoxic intravenous contrast and eliminates the risk of transporting a critically injured patient to the operating room or x-ray department. PMID- 15284544 TI - Current trends in vena caval filtration with the introduction of a retrievable filter at a level I trauma center. AB - OBJECTIVE: The purpose of this study was to document the initial experience, indications, technical success, and complications with an optional vena caval filter at a Level I trauma center. METHODS: The trauma registry and interventional radiology database were reviewed for all venal caval filters placed during a 15-month period. Records were reviewed for age of patient, indication, type of filter, and duration between placement and removal of the filter. RESULTS: One hundred thirty-six filters were placed into 130 patients (55 trauma patients), and the most frequently placed filter was the Gunther Tulip (n = 58, 29 in trauma patients). Forty-five of 1,257 trauma patients received a prophylactic vena cava filter, for a rate of 4%. Twenty-two repositioning (n = 8) or removal procedures (n = 14, 9 in trauma patients) were performed in 15 patients, with a technical success rate of 93%. No minor complications and one major complication occurred. The average duration between placement and removal was 19 days (range, 11-41 days). The mean age of patients selected prospectively for filter removal (29 years; range, 18-71 years) was significantly lower than the mean age (49 years; range, 19-82 years) of trauma, surgical, and intracranial hemorrhage patients selected for placement of prophylactic permanent filters (p < 0.002; 95% confidence interval, 18.0-22.4). CONCLUSION: The Gunther Tulip filter is commonly used at this Level I trauma center as an optional filter that can be left in place as a permanent filter or removed up to 41 days after placement. Without an intervening repositioning procedure, the manufacturer suggests that the Gunther Tulip filter can be safely removed within 14 days of implantation, or it can remain in place as a permanent filter. PMID- 15284545 TI - Futility of resuscitation criteria for the "young" old and the "old" old trauma patient: a national trauma data bank analysis. AB - BACKGROUND: Increasing geriatric trauma is producing disproportionate use of resources. In burn victims, age and burn extent correlate with mortality, yielding the establishment of criteria for futile resuscitation. Such criteria would be useful to trauma patients and their families in making withdrawal-of care decisions while reducing resource use. Our objective, therefore, was to identify injury and physiologic parameters that would indicate a high probability of futile resuscitation among geriatric trauma patients. METHODS: Data pertaining to patients greater than or equal to 65 years of age within the National Trauma Databank from 1994 to 2001 were analyzed. Multivariate logistic regression-with mortality as the outcome variable and head, chest, and/or abdominal injury; base deficit; gender; comorbidities; and admission systolic blood pressure (SBP) as covariates-was performed to develop a stratification scheme providing criteria indicative of a high probability of futile resuscitation. RESULTS: There were 76,304 patients with a mean age of 79.4 years. Head, thoracic, and abdominal injury; age; gender; comorbidities; admission SBP; and base deficit were associated with mortality. Patients with severe chest and/or abdominal injury, moderate to severe head injury, admission SBP less than 90 mm Hg, and significant base deficit had mortalities approaching 100%. Older patients with modest shock and mild to moderate head injury admitted with severe chest and/or abdominal injury had a less than 5% chance of survival. CONCLUSION: Geriatric trauma patients with severe chest and/or abdominal trauma with moderate shock and mild to moderate head injury have an exceedingly low probability of survival. These data support early withdrawal of care in these individuals. PMID- 15284546 TI - Injury patterns among female trauma patients: recognizing intentional injury. AB - BACKGROUND: Intimate partner violence and interpersonal violence are a major source of morbidity and mortality among women. Our objective was to identify patterns of injury consistent with intentional injury in female trauma patients admitted to the hospital. METHODS: Subjects were women aged 16 to 65 years discharged from acute care hospitals in a single year with a primary diagnosis of injury. Data were collected from 14 states across the United States. Analysis was performed using multivariate logistic regression. RESULTS: Women who suffered blunt intentional trauma exhibited very different injury patterns than those hospitalized for motor vehicle collisions or falls. The risk of facial injury with blunt intentional trauma was much higher than for other mechanisms (odds ratio, 4.9; 95% confidence interval, 4.20-5.74). Head injury was also more common in these women (odds ratio, 1.4, 95% confidence interval, 1.15-1.70). CONCLUSION: Physicians can potentially improve identification of cases of intimate partner violence and interpersonal violence by understanding common injuries associated with interpersonal violence. PMID- 15284547 TI - Severity of injury is underestimated in the absence of autopsy verification. AB - BACKGROUND: Trauma disproportionately affects young, productive citizens. To decrease the preventable death rate and morbidity, and to save society some of the estimated 230 million dollars per day revenue loss attributable to these injuries, trauma systems were developed. New York State instituted a population based regional trauma registry to enter all patients meeting appropriate International Classification of Diseases, Ninth Revision diagnostic codes. METHODS: We evaluated the registry records deaths in a single New York State trauma region. We compared the medical records used for registry entry to the autopsy records from the County Medical Examiner's Office to determine accuracy of diagnostic coding. On the basis of autopsy data, the records were then recoded and the extent of the trauma rescored. RESULTS: One hundred thirty-four deaths from 1993 to 1998 were recorded. Twelve records (9%) were accurately entered. One hundred twenty-two records had 452 errors. The mean Injury Severity Score (ISS), based on the medical record face sheet, was 29.93. The revised ISS, based on autopsy review, was 34.44 (p = 0.0108, two-tailed t test). The 95% confidence interval of the difference was 1.05 to 7.96. Z scores were -14.36 before autopsy review and -13.21 after autopsy review (p = 0.4395, not significant). We demonstrated a significantly higher ISS in the patients who died when the autopsy findings were included for coding. This information was not available from the medical record. CONCLUSION: To accurately compare trauma center performance and injury severity, the inclusion of autopsy data is critically important. Present state law does not permit sharing of this information with the trauma centers. When comparing mortality rates of New York State trauma centers, data must be carefully interpreted. PMID- 15284548 TI - Two major hospitals can share level I status in a rural community setting. AB - BACKGROUND: Even though development of Level I trauma centers is thought to improve outcome of injury treatment, the political barriers in communities where two capable hospitals compete for designation can be formidable. This report documents the initial experience of a Level I trauma center developed in a two hospital setting whereby each hospital hosted the trauma center on an alternating annual basis. METHODS: Preliminary efforts began with a community-based report on trauma care to which both hospitals and their associated school of medicine contributed. In addition to confirming need, all parties agreed to develop a single Level I trauma center that would be verified by the appropriate state agency and would receive professional and financial support from all three institutions. RESULTS: The Southern Illinois Trauma Center began to function on July 1, 1999. Prehospital acceptance of the community agreement has resulted in appropriate triage to the designated hospital in 95% of the 1,000 cases seen annually. Integration of trauma care into the surgical residency program has been valuable both for function of the trauma center and as a teaching enhancement for trauma and critical care as reflected by significantly improved American Board of Surgery In-Training Examination scores. The trauma center is governed by a trauma committee with representation from the three institutions. The percentage of trauma transfers has increased over 3 years from 23% to 28%. Four annual institutional site changeovers have now occurred without incident or disruption of service. The trauma center finances are reviewed by the participants on a biannual basis and have been deemed favorable. CONCLUSION: The community medical resources are commonly polarized between two large hospitals but need not prevent centralized trauma center development if preagreed community support can be achieved. Annual site change is not an impediment and could be successfully used in other similar communities, provided they are receptive to the concept of sharing Level I trauma center site designation. PMID- 15284549 TI - Regional air transport of burn patients: a case for telemedicine? AB - BACKGROUND: Air transport of burn patients is plagued by frequent "overtriage." We examined the use of air transport and the feasibility of using alternative methods such as telemedicine to assist in evaluation and treatment of burn patients within our region. METHODS: We reviewed all burn patients transported by air during 2000 to 2001. Each patient was classified as being most appropriate for air, ground, or family transport. In addition, a decision was made regarding whether telemedicine evaluation of the patient before transport could have significantly altered initial treatment decisions. RESULTS: Two hundred twenty five acutely burned patients were transferred from referring hospitals in nine states, at a mean distance of 246 air miles. Mean burn size calculated by burn center physicians was 19.7% total body surface area, whereas referring physicians' mean estimate was 29% total body surface area. In 92 cases, over- or underestimation of burn size by referring physicians of as much as 560% or decisions regarding performance of endotracheal intubation suggested that telemedicine evaluation before transport might have significantly altered transport decisions or care. Air transport charges exceeded hospital charges in 21 cases. CONCLUSION: Frequent discrepancies in burn assessment contribute to overuse of air transport. The ability to evaluate burn patients by telemedicine may have the potential to assist decisions regarding transfer, avoid errors in initial care, and reduce costs. We are currently attempting to develop and test such a system. PMID- 15284550 TI - Prehospital end-tidal carbon dioxide concentration and outcome in major trauma. AB - BACKGROUND: End-tidal carbon dioxide (Petco2) concentration is a marker of the pathophysiologic state because it is a reflection of cardiac output. Petco2 correlates with outcome after prehospital primary cardiac arrest, but association with outcome from prehospital trauma has not been established. METHODS: Between 1998 and 2001, Petco2 was recorded in 191 blunt trauma patients requiring prehospital intubation. Rapid sequence intubation was performed using suxamethonium (1 mg/kg) and etomidate (0.2-0.3 mg/kg). Initial Petco2 after endotracheal intubation (t0) and Petco2 at 20 minutes after endotracheal intubation (t20) were recorded, together with survival to discharge. RESULTS: Median Petco2 at t20 was 4.10 kPa in survivors and 3.50 kPa in nonsurvivors (95% confidence interval of difference between medians, 0.40 to 0.90 kPa; p < 0.0001). Petco2 at t20 was a better predictor of outcome than at t0. CONCLUSION: Only 5% patients with Petco2 < 3.25 kPa survived to discharge. Petco2 at t20 is of value in predicting outcome from major trauma. PMID- 15284551 TI - Diagnostic utility of sublingual PCO2 for detecting hemorrhage in penetrating trauma patients. AB - BACKGROUND: Hemorrhage results in early compromise of splanchnic circulation. Studies have shown that sublingual Pco2 (SLCO2) correlates with gut perfusion. We tested SLCO2's ability to detect hemorrhage. We compared SLCO2 with arterial base deficit (BD) and lactate (LAC). METHODS: This was a prospective study of patients with penetrating torso trauma. SLCO2 was measured at triage. Blood loss was defined as none (group 1), minimal to moderate (<1,500 mL) (group 2), or severe (>/=1,500 mL) (group 3). Data were reported as mean (95% confidence interval) and compared by analysis of variance. Receiver operating characteristic curves compared diagnostic performance between SLCO2, BD, and LAC. RESULTS: One hundred eight patients were enrolled. There was a significant difference (p < 0.001) in SLCO2 between all blood loss groups: group 1, 46.9 mm Hg (44.9-49.0 mm Hg); group 2, 53.5 mm Hg (50.8-56.2 mm Hg); and group 3, 66.0 mm Hg (53.1-78.9 mm Hg). There were no significant (p > 0.05) differences for receiver operating characteristic curves between SLCO2, BD, or LAC. CONCLUSION: SLCO2 differentiated blood loss groups. SLCO2 may be useful in triage of penetrating trauma patients. PMID- 15284552 TI - Apoptogenic effect of fentanyl on freshly isolated peripheral blood lymphocytes. AB - BACKGROUND: Opioids may trigger the apoptotic death of widely ranging cell types, and apoptosis contributes to the immune deficiency of critically ill patients and subjects experiencing surgical trauma. There is evidence that an altered mitochondrial membrane potential constitutes an early and irreversible step in the death-signaling pathway of apoptosis. This study investigated whether fentanyl, a opioid widely used in the management of these patients, may induce apoptosis of T cells by altering their mitochondrial membrane potential. METHODS: Peripheral blood lymphocytes were cultured in the presence of 30 ng fentanyl for 60 (time 1), 90 (time 2), and 120 (time 3) minutes, respectively. The cells then were processed for assessment of mitochondrial membrane potential by means of flow cytometry and confocal scanning microscopy. Furthermore, production of reactive oxygen species, expression of the Fas-Fas L pro-apoptotic pathway, and apoptosis frequency were measured by means of flow cytometry. Control cells were incubated for the same times in the complete culture medium without the drug. RESULTS: Flow cytometry analysis showed a significantly increased rate (p < 0.05) of lymphocytes with disrupted mitochondrial membrane potential after incubation with fentanyl for 90 and 120 minutes, as compared with both control cells and lymphocytes cultured in the presence of fentanyl for 60 minutes. In addition, as early as 60 minutes after exposure to fentanyl, cells displayed a disrupted mitochondrial membrane potential when this was assayed by means of confocal laser scanning. These findings were associated with increased production of reactive oxygen species. The frequency of apoptotic lymphocytes was markedly increased (p < 0.05) after 120 minutes of incubation, as compared with untreated cells and cells exposed to fentanyl for only 60 and 90 minutes. Expression of Fas-FasL was not substantially affected by exposure to fentanyl. CONCLUSIONS: Fentanyl may induce a time-dependent apoptosis of lymphocytes by altering their mitochondrial redox metabolism. PMID- 15284553 TI - Alterations of red blood cell shape in patients with severe trauma. AB - BACKGROUND: Trauma is accompanied by a decrease in red blood cell (RBC) deformability, which may manifest itself earlier than secondary septic complications. The mechanisms of this phenomenon are not clear. The aim of this study is to determine when the alterations of RBC shape appear in trauma patients. METHODS: RBC shape was examined by scanning electron microscopy in 43 patients with multisystem trauma. Blood samples were taken at admission and every 24 hours afterward for 4 to 10 days. The patients were divided into two groups: the survivors and those who died of septic complications. The control group consisted of 10 healthy volunteers. RESULTS: A significant decrease in the percentage of discoid erythrocytes, compared with the volunteers, was observed in both groups of patients at admission (77.6 +/- 11.9 and 66.7 +/- 5.8 vs. 96.0 +/- 2.9%, p < 0.01). The percentage of irreversibly changed RBC (spherostomatocytes, spherocytes) was lower in survivors (12.9 +/- 2.0% vs. 20.3 +/- 9.4%, p < 0.01). This phenomenon remained constant until the end of the study. The survivors only showed a tendency toward the improvement of RBC shape. CONCLUSION: RBC shape alterations appear within the first hours after trauma and persist for at least 7 to 10 days. These changes are more severe in patients with secondary septic complications. PMID- 15284554 TI - Left mediastinal width and mediastinal width ratio are better radiographic criteria than general mediastinal width for predicting blunt aortic injury. AB - BACKGROUND: General mediastinal width, left mediastinal width, and mediastinal width ratio were compared as radiographic predictors of aortic injury. METHODS: A retrospective study investigated the chest radiographs of 51 patients admitted to a level 1 trauma center during a 6-year period for a thorough survey of aortic injury. Mediastinal width (MW >/= 8 cm), left mediastinal width (LMW >/= 6 cm), mediastinal width ratio (MWR >/= 0.60), and a combination of LMW and MWR were compared as predictors of aortic injury. The cutoff points were predetermined by receiver-operator-curve to accommodate 100% sensitivity for each criterion. RESULTS: Of the 51 patients, 21 had aortic injuries and 30 had normal imaging studies. All criteria had 100% negative predictive value. The specificities and positive predictive values, respectively, were 13.3% and 44.7% (MW), 40.0% and 53.8% (LMW), 43.3% and 55.3% (MWR), and 66.7% and 67.7% (combined LMW and MWR). The positive likelihood ratio of aortic injury was 3.00 when LMW was 6 cm or more and MWR was 0.60 or more. CONCLUSIONS: Both an LMW of 6 cm or more and an MWR of 0.60 or more are better radiographic criteria than an MW of 8 cm or more for predicting blunt aortic injury. Trauma patients with positive test results based on the combined LMW and MWR criteria should proceed immediately to aortography or helical computed tomography. PMID- 15284555 TI - Comparison of serum and cerebrospinal fluid protein S-100b levels after severe head injury and their prognostic importance. AB - BACKGROUND: This study aimed to compare serum and cerebrospinal fluid (CSF) S 100b protein levels after a severe head injury. The changes in serum S-100b and CSF S-100b concentrations were investigated as indicators of brain damage for patients suffering from severe head injuries. METHODS: The sample included 48 patients with Glasgow Coma Scale scores of 8 or below who had been admitted to the authors' emergency service soon after their severe head injury occurred. Both blood and CSF samples were taken within 1 to 11 hours after admission, then 24, 48, and 72 hours after the injury. Samples of CSF were taken using a ventricular catheter. The outcome was evaluated 6 to 9 months after hospital discharge using the Glasgow Outcome Scale. RESULTS: The overall mean serum S-100b concentration was 3.5 +/- 6.4 among the patients with unfavorable outcomes and 1.3 +/- 2.5 among those with favorable outcomes. These results were not statistically significant (p > 0.05). The overall mean CSF S-100b concentration was 62.2 +/- 21.8 among the patients with unfavorable outcomes and 21.8 +/- 17.7 among those with favorable outcomes. These results, however, were statistically significant (p < 0.05). CONCLUSION: The results show that CSF S-100b levels clearly are superior to serum S-100b levels for predicting outcome after severe head injury. PMID- 15284556 TI - The applicability of a computer model for predicting head injury incurred during actual motor vehicle collisions. AB - BACKGROUND: Head injury is a significant cause of both morbidity and mortality. Motor vehicle collisions (MVCs) are the most common source of head injury in the United States. No studies have conclusively determined the applicability of computer models for accurate prediction of head injuries sustained in actual MVCs. This study sought to determine the applicability of such models for predicting head injuries sustained by MVC occupants. METHODS: The Crash Injury Research and Engineering Network (CIREN) database was queried for restrained drivers who sustained a head injury. These collisions were modeled using occupant dynamic modeling (MADYMO) software, and head injury scores were generated. The computer-generated head injury scores then were evaluated with respect to the actual head injuries sustained by the occupants to determine the applicability of MADYMO computer modeling for predicting head injury. RESULTS: Five occupants meeting the selection criteria for the study were selected from the CIREN database. The head injury scores generated by MADYMO were lower than expected given the actual injuries sustained. In only one case did the computer analysis predict a head injury of a severity similar to that actually sustained by the occupant. CONCLUSION: Although computer modeling accurately simulates experimental crash tests, it may not be applicable for predicting head injury in actual MVCs. Many complicating factors surrounding actual MVCs make accurate computer modeling difficult. Future modeling efforts should consider variables such as age of the occupant and should account for a wider variety of crash scenarios. PMID- 15284557 TI - External subdural drainage in the treatment of infantile chronic subdural hematoma. AB - BACKGROUND: The management of chronic subdural hematoma (CSDH) in infants remains controversial. The purpose of this study was to analyze the clinical characteristics of CSDH in infancy and evaluate the efficacy of continuous external subdural drainage in the treatment of infantile CSDH. METHODS: We prospectively collected 36 consecutive infants with CSDH, to receive continuous external subdural drainage as the initial management. Medical records were reviewed for comparison of age, gender, cause of injury, clinical presentation, surgical management, and outcome. Diagnosis was made by computed tomography and/or magnetic resonance imaging. RESULTS: There were 20 boys and 16 girls, with ages ranging from 1 to 11 months (average, 5.9 months). The most common cause of CSDH was head injury (44.5%), followed by shaken baby syndrome (36.1%). The most common clinical presentations were seizure, bulging fontanel, and consciousness disturbance. Continuous external subdural drainage was the definite treatment in 34 patients (94.4%). The drains were left in place for no more than 9 days. Only two (5.6%) patients needed permanent subduroperitoneal shunting. No obvious complication was found. At follow-up (17-160 months; mean, 86.6 months), 23 (63.9%) had good recovery, 5 (13.9%) had moderate disability, 3 (8.3%) had severe disability, 4 (11.1%) were in a vegetative state, and 1 (2.8%) died. CONCLUSION: Continuous external subdural drainage was an effective treatment in infantile CSDH, with a low complication rate and good clinical outcome. It might be considered as a strategy before subduroperitoneal shunting in the treatment of CSDH in infants. PMID- 15284558 TI - Functional outcome of nonoperatively managed renal injuries in children. AB - BACKGROUND: This study aimed to define better the functional outcome of nonoperatively managed renal injuries in children. METHODS: All children who had blunt renal trauma managed nonoperatively were reviewed for injury grade, blood urea nitrogen (BUN), creatinine, blood pressure, and percentage of function according to technetium-99m-dimercaptosuccinic acid renal scan after complete healing. RESULTS: Over a 2-year period, 17 children (mean age, 10.4 years) were managed conservatively for their renal injuries. There were two grade 2, two grade 3, nine grade 4, and four grade 5 injuries. Complete healing was documented in all cases within 3 months after injury. Renal scarring and volume loss were evident for all healed high-grade injuries (grades 4 to 5) at follow-up imaging. Technetium-99m-dimercaptosuccinic acid scanning demonstrated a decline in percentage of total renal function corresponding to injury severity (44.7 +/- 8.4% function for grades 2 and 3, 41.8 +/- 9.2% for grade 4 vs 29.5 +/- 7.9% for grade 5). Only two children (22%), however, with grade 4 injury had severe compromise of function (<30%). At the follow-up visit, all the children were asymptomatic and normotensive. None had abnormal BUN or creatinine (mean BUN, 10.5 +/- 5.1 mg/dL; mean creatinine, 0.6 +/- 0.2 mg/dL). CONCLUSIONS: The functional outcome for children with nonoperatively managed kidney injuries is good and correlates with injury grade. Children with grades 2 to 4 injuries managed conservatively retain near normal function. Those with grade 5 injuries have a loss of function attributable to scarring and parenchymal volume loss. Long-term follow-up evaluation of these children may be warranted. PMID- 15284559 TI - Nonmotorized scooters: a source of significant morbidity in children. AB - BACKGROUND: Nonmotorized scooters have increased rapidly in popularity over the past year. However, the morbidity associated with this new type of recreational vehicle is poorly defined. This study examined the pattern of scooter-related injuries sustained in children admitted to a level 1 pediatric trauma center. METHODS: The records of all children admitted to the authors' institution after a scooter-related injury between January 1, 2000 and December 31, 2001 were reviewed. Information regarding patient demographics, mechanism of injury, type of injury sustained, and hospital course was prospectively collected and retrospectively analyzed. RESULTS: During the study period, 27 children were admitted with scooter-related injures. The average age of the patients was 9.1 +/ 1.9 years, and 63% were boys. The average Injury Severity Score (ISS) was 7.9 +/ 6. The most common mechanism of injury was a fall. However, 26% of the patients were involved in a scooter collision with a motor vehicle. Injuries to the head occurred most frequently, followed by extremity injuries. Only 10 of the children (37%) were wearing a helmet at the time of injury. CONCLUSIONS: Scooters are an increasingly popular form of recreational vehicle among children. However, they can result in serious injury, particularly to the head and extremities. The authors recommend that all children riding scooters wear protective equipment and avoid riding in areas that have moving motor vehicles. PMID- 15284560 TI - Role of an extended tertiary survey in detecting missed injuries in children. AB - BACKGROUND: There are limited data on the incidence of delayed diagnosis of injuries in children. We sought to investigate the role of an extended tertiary survey in pediatric trauma patients. METHODS: All children that were admitted to The Children's Hospital at Westmead with an Injury Severity Score (ISS) >/= 9 were included in the study. The trauma fellow performed the tertiary survey the day after admission. This was repeated after extubation in ventilated patients and in head injury patients when they were more mobile and cooperative. RESULTS: Seventy-six patients satisfied the criteria for the study (50 boys and 26 girls). Age ranged from 1 month to 15 years. The median ISS was 14. Sixteen (16%) of the patients had missed injuries, of which skeletal injuries were the most common (10 of 12). Delayed diagnosis of injury occurred most frequently in children involved in motor vehicle injuries. Sixty-six (66%) of the injuries were detected within the first 24 hours. Inadequate assessment and head injury were the most common contributing factors. CONCLUSION: The incidence of missed injury (16%) in our study was comparable to reported figures in the adult literature. There was no correlation between missed injuries and intensive care unit stay or ISS. Head injury often delayed diagnosis and thus ongoing evaluation in this group is recommended. Missed injuries did not result in mortality, but there was significant associated morbidity. A tertiary survey should be part of the evaluation of the pediatric trauma patient. PMID- 15284561 TI - Association of hospital trauma designation with admission patterns of injured children. AB - BACKGROUND: Little is known about the influence of regionalization of trauma care on pediatric trauma care delivery. The purpose of this study was to estimate whether formal adoption of a statewide trauma system was associated with hospital admission patterns of injured children. METHODS: A longitudinal study of children who were residents of Washington State during 1989 to 1999 was conducted. The main outcome measure was hospital admission for trauma. RESULTS: During the 11 year period, there were 24,955 admissions. Admission rates of injured children to pediatric-designated trauma hospitals decreased by 20%, rates at adult-designated hospitals decreased by 60%, and rates at nondesignated hospitals decreased by 66%. Introduction of the trauma system in 1994 was associated with a 12% increase in admission rates to pediatric-designated hospitals, little change (+1%) in admission rates to adult-designated centers, and an 11% decrease in admissions at nondesignated hospitals. CONCLUSION: Trauma designation in Washington was associated with a shift in admissions from nondesignated hospitals to pediatric trauma hospitals. PMID- 15284562 TI - Incidence and patterns of violent and/or traumatic deaths between 1993 and 1999 in the Transkei region of South Africa. AB - BACKGROUND: Incidence and patterns of violent and/or traumatic deaths among 4,525 victims over a 7-year period in the Transkei region of South Africa were investigated. METHOD: Retrospective review and analysis was performed of all medicolegal autopsies (n = 6,181) between January 1993 and December 1999, of which 4,525 were violent or traumatic deaths. RESULTS: During the 7-year period (January 1993-December 1999), violent and/or traumatic deaths in the Transkei region accounted for an average annual rate of 162 per 100,000 of the population. The common causes of deaths per 100,000 of population per year were as follows: motor vehicle collisions, 63; firearm injuries, 43; stab wounds, 32; and blunt trauma, 24. Male subjects outnumbered female subjects by a 3.3:1 ratio. The murder rate in female subjects was 18 per 100,000 population. The murder rate in this area increased from 94 per 100,000 in 1993 to 121 in 1999. Nearly 50% of the violent and/or traumatic deaths occurred in the 21- to 40-year age group. There has been an increase in nontraumatic deaths such as hanging (1.5 times) and poisoning (5 times). CONCLUSION: The average annual incidence of violent and/or traumatic deaths in the Transkei region of South Africa is 162 per 100,000 population. Firearm-related deaths, at 43 per 100,000 of the population per year, have contributed substantially to this high incidence. This is a major cause of concern. PMID- 15284563 TI - Risk adjustment for injured patients using administrative data. AB - BACKGROUND: Risk adjustment methods are needed for population-based studies of injured patients. METHODS: Data were obtained from National Hospital Discharge Surveys, 1996 to 2000. International Classification of Diseases, Ninth Revision, Clinical Modification (ICD-9-CM) diagnoses were used to categorize Abbreviated Injury Scale score, Injury Severity Score, ICD-9-CM Injury Severity Score, injury mechanisms, and comorbidities. Regression models for weighted survey data were constructed from combinations of these classifications, plus age and sex, to predict mortality, length of stay (LOS), or discharge to long-term care (LTC). RESULTS: Increased Abbreviated Injury Scale score, increased Injury Severity Score, or decreased ICD-9-CM Injury Severity Score were similarly associated with mortality, prolonged LOS, or more frequent LTC, as was increased age. Penetrating or burn mechanisms were associated with mortality and longer LOS; penetrating or vehicle mechanisms were associated with less frequent LTC. Different comorbidities affected LOS and LTC. Men had shorter LOS and less frequent LTC than women. CONCLUSION: Hospital outcomes after injury are predictable from age, sex, and standard diagnosis groupings. Anatomic scales gave similar results when adjusted for other factors. PMID- 15284564 TI - Treatment of unstable pelvic fractures: use of a transiliac sacral rod for posterior lesions and an external fixator for anterior lesions. AB - BACKGROUND: This study sought to define the role of transiliac sacral rods used in combination with an external fixator for the management of unstable pelvic fractures. METHODS: This retrospective study evaluated cases in which the surgical strategy was open reduction and internal fixation of posterior lesions with two transiliac sacral rods and closed reduction and external fixation of anterior lesions with an AO external fixator. The data for 65 cases were analyzed. Comprehensive Classification (AO) identified 42 C1 cases, 21 C2, cases and 2 C3 cases. Fractures with iliac bone involvement that impeded the application of an external fixator or transiliac sacral rods were excluded. The follow-up period was 85 months (range, 24-140 months). RESULTS: All the fractures/dislocations healed well. The complications involved 17 cases (26.2%) of persistent posterior pain, 16 cases (24.6%) of irreversible neurologic deficit, 2 cases (3.1%) of posterior wound infection, 3 cases (4.6%) of pin tract infection, and 4 cases (6.2%) of irreversible urologic deficit. The functional results showed that the surgical results were satisfactory in 42 cases (64.6%) and unsatisfactory in 23 cases (35.4%). CONCLUSIONS: For type C pelvic fractures without significant iliac bone involvement, surgical management with posterior transiliac fixation using sacral rods and anterior external fixation yields good radiologic results. The functional results correlated primarily with avoidance of complications and not necessarily with the radiologic results. PMID- 15284565 TI - Rinsing-suction reamer attenuates intramedullary pressure increase and fat intravasation in a sheep model. AB - BACKGROUND: Reamed intramedullary nailing causes an increase of intramedullary pressure. A new rinsing-suction reamer (RSR) can reduce this problem, and it was evaluated in animal experiments in comparison with the AO reamer (AOR) to see its effects on intramedullary pressure and fat intravasation. METHODS: Reamed intramedullary nailing was performed in 14 sheep using the RSR or AOR. The following parameters were evaluated: intramedullary pressure, hemodynamics, blood tests, lung histology, and radiographs of the femur that was operated on. RESULTS: Intramedullary pressure during reaming was significantly (p < 0.001) lower with RSR (9 mm, 34 mm Hg; 9.5 mm, 4 mm Hg; 10 mm, 1 mm Hg) than AOR (9 mm, 750 mm Hg; 9.5 mm, 292 mm Hg; 10 mm, 138 mm Hg). There was a significantly (p < 0.05) higher increase of pulmonary resistance in AOR (from 144 +/- 84 dyne x s x cm to 391 +/- 169 dyne x s x cm) than in RSR (from 137 +/- 51 dyne x s x cm to 258 +/- 105 dyne x s x cm) after nailing and less intravenous fat measured in RSR (0.9; AOR, 2.9; p < 0.05) at all stages of reaming, at nail insertion (RSR, 0.3; AOR, 2.7; p < 0.05), and 30 seconds after nail insertion (RSR, 0.2; AOR, 1.1; p < 0.05) proved by the Gurd test. Pco2 increased (p < 0.05) in AOR (AOR, 36 +/- 5 vs. 40 +/- 7 mm Hg; RSR, 33 +/- 4 vs. 32 +/- 3 mm Hg) and pH dropped significantly (AOR, 7.49 +/- 0.06 vs. 7.45 +/- 0.05; RSR, 7.53 +/- 0.04 vs. 7.54 +/- 0.04; p < 0.05). Semiquantitative histologic analysis proved a significant higher pulmonary fat load in AOR (13.1 +/- 13.4) versus RSR (3.9 +/- 1.5, p = 0.00002). CONCLUSION: Because we found only a minimal increase of the pulmonary arterial pressure as a sign of pulmonary embolism, we conclude that by using the RSR, the systemic side effects caused by intravasation of medullary content during reaming could be reduced as far as possible. PMID- 15284566 TI - High-intensity ultrasound treatment of blunt abdominal solid organ injury: an animal model. AB - BACKGROUND: High-intensity focused ultrasound (HIFU) is effective in producing hemostasis in injuries from organ lacerations and punctures in animals but has not been evaluated in impact injuries. METHOD: High-energy blows were applied to 11 heparinized and anesthetized pigs, resulting in solid organ injury. HIFU was applied to injuries via laparotomy. The animals were closed, administered saline, observed under general anesthesia for 3.6 +/- 0.4 hours, reopened, and inspected, and abdominal free fluid was aspirated. RESULTS: Organ hemostasis was achieved (mean +/- SD) with 15 +/- 6 minutes of HIFU treatment and 54 +/- 3 minutes of operating time, and 18.8 +/- 13.1 mL/kg of blood was recovered from the abdomen. One animal died from an untreated occult injury to a large vein. HIFU-treated sites were hemostatic at relaparotomy, with 8.6 +/- 6.2 mL/kg abdominal serosanguinous fluid recovered. CONCLUSION: HIFU is effective in producing hemostasis by direct treatment of injured parenchyma in blunt trauma. PMID- 15284567 TI - Enterocutaneous fistula and ventral hernia after absorbable mesh prosthesis closure for trauma: the plain truth. AB - HYPOTHESIS: The severity of abdominal injury is the determining factor for the development of enterocutaneous fistula and ventral hernia after absorbable mesh prosthesis closure (AMPC) for trauma. METHODS: We conducted a retrospective analysis of case series that included 140 consecutive trauma patients with AMPC surviving more than 48 hours from October 1, 1989, to March 31, 2000, at a Level I trauma center. The days until abdominal wall reconstruction was used as a measure of exposure of the viscera to the mesh. The abdominal trauma index (ATI) was used as the measure of injury severity. Statistical analysis included t test comparisons, logistic regression analysis, and life-table analysis for hernia development. RESULTS: Enterocutaneous fistula occurred in 10 patients (7.1%). The ATI (mean, 32.5 +/- 23.1) was the only variable independently associated with fistula formation (p = 0.01). The risk of fistula increased by 4% for each 1 unit increase in ATI (95% confidence interval [CI], 1-7%). One hundred seventeen patients (84%) survived to completion of abdominal wall reconstruction over a mean of 18.9 +/- 22.5 days and 3.6 +/- 1.9 operations. The number of days until abdominal wall reconstruction was the only variable independently associated with ventral hernia development (p < 0.001). The likelihood of fascial closure decreased by 26% (95% CI, 16-44%) per day and the risk of ventral hernia increased by 16% (95% CI, 9-23%) per day. The hernia development rate at 4 years (per life table) was 67% for the total, 13% for patients with delayed fascial closure, and 80% for patients requiring other closure techniques. CONCLUSION: Although the severity of abdominal injury is the most important factor for fistula formation, the most important factor for ventral hernia development is the duration of AMPC. Daily interventions, such as mesh tightening, may be necessary to limit ventral hernia in these high-risk patients. PMID- 15284568 TI - Successful treatment of trauma-induced short bowel syndrome with early living related bowel transplantation. AB - BACKGROUND: : Total parenteral nutrition (TPN) is a life-saving therapy for patients with short bowel syndrome. However, TPN is associated with a high incidence of serious complications, poor quality of life, and elevated cost. An attempt was made to avoid TPN-related complications associated with trauma induced short bowel syndrome by using early living related donor bowel transplantation. METHODS: : Three men 27 to 30 years of age with trauma-induced short bowel syndrome received early living related donor bowel transplantation using segmental ileal grafts. RESULTS: : All the donors had an uncomplicated postoperative course. After a mean follow-up period of 40 months, all three recipients were alive and well, and did not require any TPN support. The ileal graft adapted perfectly to support fully the nutritional needs of young, active individuals. CONCLUSIONS: : Early living related donor bowel transplantation is a successful treatment for trauma-induced short bowel syndrome. It is associated with a lower incidence of complications, better quality of life, and lower cost than long-term TPN. PMID- 15284569 TI - A novel use of recombinant factor VIIa in HELLP syndrome associated with spontaneous hepatic rupture and abdominal compartment syndrome. PMID- 15284570 TI - Mitral valve avulsion after a four-wheel all-terrain vehicle crash. PMID- 15284571 TI - Successful resuscitation of a traumatic cardiac arrest victim in hemorrhagic shock with vasopressin: a case report and brief review of the literature. PMID- 15284572 TI - Endovascular repair of a blunt traumatic axillary artery injury presenting with limb-threatening ischemia. PMID- 15284573 TI - Autogenous fascia patch for unresectable and residual arteriovenous malformation within the bone. PMID- 15284574 TI - Retrograde venous bullet embolism: a rare occurrence-case report and literature review. PMID- 15284575 TI - Delayed colonic stricture and obstruction after blunt abdominal trauma: a case report and review of the literature. PMID- 15284576 TI - Pneumomediastinum caused by a lightning strike. PMID- 15284577 TI - Bilateral renal subcapsular hematoma subsequent to seat belt compression. PMID- 15284578 TI - Hyoid bone fracture from a gunshot wound. PMID- 15284581 TI - Xeroderma pigmentosum in four siblings with three different types of malignancies simultaneously in one. AB - Xeroderma pigmentosum (XP) is a rare autosomal recessive disease characterized by marked sensitivity to ultraviolet radiation that leads to the development of multiple skin malignancies. The authors describe four XP siblings in a consanguineous Pakistani family. The first patient was a boy who died at age 2 years. The second and third siblings were girls who died at age 2 and 7 years, respectively. The fourth sibling, the propositus, was a boy diagnosed with XP at age 7 years. He developed three different types of malignancies simultaneously and died at age 13. The authors conclude that it is important to be aware of multiple malignancies of different types in the same patient with XP. PMID- 15284582 TI - Recurrent neuroblastoma with gastric invasion. AB - The authors present an unusual manifestation of neuroblastoma in a young child: upper gastrointestinal bleeding due to erosion of the tumor into the stomach. Included are reviews of gastrointestinal manifestations of neuroblastoma and gastric tumors in children. PMID- 15284583 TI - Successful prophylactic treatment for bleeding in a girl with severe hereditary prothrombin deficiency using a prothrombin complex concentrate (Bebulin VH). AB - The authors describe the evaluation and course of severe hereditary prothrombin deficiency in a 14-year-old girl first diagnosed at age 4 years. Detailed is the evolution of her treatment from episodic fresh-frozen plasma after bleeding events to prophylactic home infusions with the prothrombin complex concentrate Bebulin VH. Pharmacokinetic data on factor II recovery and half-life are presented. The patient has been essentially free of abnormal bleeding while on this prophylactic regimen for 17 months, with no toxicities and with a much improved quality of life. PMID- 15284584 TI - Eukaryotic initiation factor 4E staining as a clinical marker in pediatric neuroblastoma. AB - OBJECTIVES: Neuroblastoma (NBL) has several well-established prognostic factors. Eukaryotic initiation factor 4E (4E) has shown promise as a prognostic marker in other cancers. We hypothesize that a correlation exists between 4E staining and NBL clinical outcome. METHODS: Eleven adrenal NBL patient charts were reviewed for data, including age, stage, MYCN amplification, Shimada classification, and mortality. These patients' surgical specimens were stained with 4E antibody and scored for staining density. 4E expression, quantified by staining density, was compared to clinical data. CONCLUSION: 4E staining significantly correlates with age at diagnosis. The remaining prognostic factors lack statistically significant correlation. Increasing sample size may further establish statistically significant correlations. 4E could become an additional prognostic factor of NBL. PMID- 15284585 TI - Reliability and reproducibility of classification of children as "bleeders" versus "non-bleeders" using a questionnaire for significant mucocutaneous bleeding. AB - The diagnosis of type 1 von Willebrand disease (VWD), the most common inherited bleeding disorder in humans, is greatly dependent on an accurate diagnosis of significant mucocutaneous bleeding. In a previous study, the authors modified the criteria of the International Society on Thrombosis and Haemostasis for significant mucocutaneous bleeding to a format, the Hospital for Sick Children (HSC) criteria, that was more applicable to diagnose significant mucocutaneous bleeding in children. To assess the reliability and reproducibility of classification of subjects as "bleeders" versus "non-bleeders" using a questionnaire for significant mucocutaneous bleeding targeted to children, 39 subjects interviewed for a previous HSC VWD study were reinterviewed for the current study. The original bleeding classification was confirmed in 80% of subjects interviewed for a second time, indicating that this method of classification is reproducible (kappa = 0.65), with a "substantial" agreement among the investigators who reviewed the questionnaire responses (kappa = 0.71). The validity and utility of the HSC questionnaire for primary screening of children with suspected mucocutaneous bleeding disorders merits assessment in further clinical studies. PMID- 15284586 TI - Engraftment syndrome emerges as the main cause of transplant-related mortality in pediatric patients receiving autologous peripheral blood progenitor cell transplantation. AB - The authors examined data from 166 children who received autologous peripheral blood progenitor cell (PBPC) transplantation to ascertain the incidence of early transplant-related mortality (TRM) and the contributing risk factors. Eleven patients (6.6%) (6 boys, 5 girls) died within 180 days following PBPC infusion. The median age was 4 years (range 2-17). The overall probability of TRM was 6.9 +/- 2% at day +180. On univariate analysis, the status of disease at transplantation (complete remission vs. not in complete remission) was identified as the only pretransplant significant predicting factor for TRM (14% of patients who were not in complete remission died within 180 days after PBPC infusion, whereas only 2% of patients in complete remission died) (relative risk [RR] 1.13, 95% confidence interval [CI] 1.01-1.26, P = 0.01). Age, gender, conditioning, and number of CD34+ cells infused were not significantly associated with TRM. In the postinfusion phase, patients who developed multiorgan dysfunction during the neutropenic period, especially when the lung was the first failing organ (RR 16.1, 95% CI 7.16-36.18, P = 0.0001), and those with engraftment syndrome (RR 2.81, 95% CI 1.49-5.24, P = 0.001) had an increased risk for TRM. On multivariate analysis, development of engraftment syndrome was the only significant variable that influenced TRM. In conclusion, the authors found for the first time that engraftment syndrome emerges as the main cause of TRM after autologous PBPC transplantation in children with malignancies. PMID- 15284587 TI - Persistent human parvovirus B19 infection in children under maintenance chemotherapy for acute lymphocytic leukemia. AB - OBJECTIVE: To report on B19 infection management and chemotherapy schedule consequences in five children treated for acute lymphocytic leukemia (ALL). PATIENTS AND METHODS: Between May 2001 and February 2002, five patients between 4 and 12 years of age, receiving maintenance chemotherapy for ALL, presented with symptoms suggesting B19 infection (pallor, fatigue, petechiae and pancytopenia in four patients; generalized rash in two patients; acute hepatitis in one patient). Qualitative polymerase chain reaction (PCR) on peripheral blood was used for diagnosis and follow-up of infection; quantitative PCR was used for viral load measurement. Intravenous nonspecific high-dose immunoglobulin therapy was administered until PCR was negative. RESULTS: Qualitative B19 DNA was found in the peripheral blood of all patients, confirming the infection. Viral load at diagnosis ranged from 10 to 10 particles/mL blood. B19 DNA was detectable in four patients at 45, 21, 40, and 44 weeks, respectively. Chemotherapy was delayed in all patients. No clear benefit of intravenous immunoglobulin was noted. CONCLUSIONS: Infection with B19 is rarely reported in patients with ALL, but it should be suspected when unexplained pancytopenia occurs during chemotherapy. Persistent B19 infection remains a challenge in the management of patients receiving maintenance chemotherapy for ALL, as no specific therapy such as a specific immunoglobulin or vaccine exists. The role of viral load measurement needs to be established in terms of its use in follow-up and evaluation of the therapeutic response. PMID- 15284588 TI - Increased bone marrow angiogenesis in children with severe chronic neutropenia treated with granulocyte colony-stimulating factor. AB - Severe chronic neutropenia (SCN) is a heterogeneous group of conditions characterized by chronically low neutrophil counts and recurrent infections. Granulocyte colony-stimulating factor (G-CSF) is the mainstay of treatment, and evidence exists that G-CSF may promote angiogenesis. To evaluate the effects of G CSF on angiogenesis in children with SCN, the authors assessed microvessel density in bone marrow biopsies from nine children with SCN before starting G-CSF treatment and while receiving G-CSF. In all patients, microvessel density was greater in the on-treatment biopsy. Increased angiogenesis may result from a direct effect of G-CSF on endothelial cells or may be an indirect effect from increased neutrophils. PMID- 15284589 TI - Outcome and prognostic factors of acute lymphoblastic leukemia in children at the National Cancer Institute, Egypt. AB - OBJECTIVE: To improve survival of children with acute lymphoblastic leukemia (ALL) in the authors' institute and to define significant prognostic factors. METHODS: This study included 154 children with newly diagnosed ALL below the age of 18 years during the period August 1, 1998, to December 31, 2000. All patients were treated according to the NCI, Cairo, Egypt, treatment protocol modified from study XIII for high-risk ALL of St. Jude Children's Research Hospital. RESULTS: B cell precursor phenotype was encountered in 73.4% of patients, T-cell in 26.6%. According to NCI/Rome criteria for risk classification, 58.4% of patients were in the high-risk group (90% of T-lineage compared with 47% of B-lineage ALL, P < 0.001). Nine patients (5.8%) died during induction and 10 patients (6.5%) failed to achieve remission. With a median follow-up of 43 months, the 3-year disease free survival and its probability at 5 years were 80 +/- 3.6% and 75.3 +/- 4%, respectively; the 3-year event-free survival and its probability at 5 years were 69 +/- 3.8% and 65.2 +/- 4%, respectively. B-cell precursor ALL had 5-year probabilities of disease-free survival and event-free survival of 80.5 +/- 4% and 71.8 +/- 4.5% compared with 60.2 +/- 8.6% and 46.7 +/- 8% for T-cell, respectively (P < 0.01, log-rank test). Prognostic factors that had a statistically significant unfavorable impact on survival by univariate analysis were age 10 years or more, central nervous system involvement, T-lineage phenotype, high-risk group, DNA index less than 1.16, and slow early response to treatment. By multivariate analysis, central nervous system involvement, high risk group, and slow early response to treatment still had prognostic significance. Risk classification demonstrated prognostic significance for B lineage but not T-lineage ALL. CONCLUSIONS: This treatment protocol was effective in improving ALL survival among patients at the authors' institute compared with previous trials, although the outcome remains lower than that in more industrialized countries. Prognostic factors defined in this study were similar to those identified by other cooperative groups. PMID- 15284590 TI - Probiotics in relapsing and chronic diarrhea. AB - Diarrhea is common in oncology patients; if it becomes chronic and relapsing, it can be debilitating, hinder planned management, and be difficult to treat. The authors describe two patients, one with leukemia who developed recurrent Clostridium difficile colitis and another who developed chronic diarrhea after bone marrow transplantation. In both patients, administration of antibiotics was suspected as the cause. In one patient, relapsing diarrhea resolved after probiotics were given with a 2-day course of metronidazole, and in the other patient, chronic diarrhea resolved after probiotics were given; resolution was maintained after the probiotics were stopped. Probiotics may offer a way to bring about resolution in antibiotic-associated chronic diarrhea. PMID- 15284591 TI - Multimodal treatment of children with unresectable or recurrent desmoid tumors: an 11-year longitudinal observational study. AB - The primary goal of treatment for desmoid tumors is complete surgical resection to achieve negative margins. In adults with unresectable or recurrent lesions, treatment options include noncytotoxic and cytotoxic drugs, but little is known about nonsurgical treatment in children. Between 1992 and 2003 six children (four girls, two boys) with a median age of 2.5 years (range 11 months to 9 years) received multimodal adjuvant therapy for unresectable or recurrent desmoid tumors. Primary treatment consisted of noncytotoxic treatment with tamoxifen (1 mg/kg orally, twice daily) and diclofenac (2 mg/kg rectally, twice daily), whereas two children with life-threatening tumor progression in addition received treatment intensification with weekly vinblastine (6 mg/m intravenously) and methotrexate (30 mg/m intravenously). Of the four children with unresectable tumors, two achieved remarkable tumor shrinkage and two had stable disease, whereas two patients were disease-free for 3.7 and 2.6 years after nonradical resection. Median observation time was 3.1 years (range 1-11 years). Treatment was generally well tolerated; only one patient developed pubertal acceleration after a duration of tamoxifen treatment of 9.3 years. Because of the potential life-threatening situation, the management of children with unresectable or recurrent desmoid tumors requires a multidisciplinary approach. Nonaggressive therapy with tamoxifen and diclofenac may be the first treatment choice in these patients, but in patients with progressive disease, cytotoxic chemotherapy is indicated. Weekly administration of vinblastine and methotrexate seems to be safe and effective in these children. PMID- 15284592 TI - The triad of seizures, hypertension, and neuroblastoma: the first described case. AB - The combination of seizures, hypertensive encephalopathy, and neuroblastoma has not been described before. The authors report one case, which is not only of interest in its own right, but also emphasizes the importance of including blood pressure measurement in the clinical examination of children, especially when hypertension could be the cause of the symptoms. PMID- 15284593 TI - Cryptococcal sepsis diagnosed by bone marrow examination. AB - Disseminated cryptococcal disease is often associated with immunodeficient states. The diagnosis is usually made using standard antigen tests on serum and cerebrospinal fluid in patients with known immunodeficiency. Often, blood and cerebrospinal fluid cultures also yield Cryptococcus neoformans. The authors describe a child whose diagnosis remained elusive until a bone marrow aspiration, performed as part of an evaluation for suspected neoplasm, revealed the offending organism. PMID- 15284594 TI - Thyrotoxicosis after matched unrelated bone marrow transplantation. AB - Acquired hyperthyroidism is most commonly autoimmune in etiology. In the setting of allogeneic bone marrow transplantation (BMT), the use of radiotherapy (total body irradiation) as part of the regimen prior to BMT is known to cause endocrine dysfunction, especially hypopituitarism and hypothyroidism, but hyperthyroidism is rare. The authors report this unusual and late complication in a young boy after BMT for relapsed childhood lymphoblastic leukemia and discuss the possible etiologies. PMID- 15284595 TI - Occurrence of t(8;22)(q24.1;q11.2) involving the MYC locus in a case of pediatric acute lymphoblastic leukemia with a precursor B cell immunophenotype. AB - The authors describe a child with clinical presentation of acute leukemia and an immunophenotype consistent with precursor B-cell acute lymphoblastic leukemia. However, the lymphoblasts had atypical L3 features and the chromosome rearrangement t(8;22)(q24.1;q11.2) involving the MYC locus. The cytogenetic features in this patient were characteristic of mature B/Burkitt leukemia and led to modulation of therapy. This case highlights the need for timely cytogenetic and molecular studies in the diagnosis of acute leukemia. PMID- 15284596 TI - Thrombocytopenia and severe hyperbilirubinemia in the neonatal period secondary to congenital thrombotic thrombocytopenic purpura and ADAMTS13 deficiency. AB - Congenital thrombotic thrombocytopenic purpura (TTP) is an inherited form of TTP due to the deficiency of von Willebrand factor (vWF) cleaving protease ADAMTS13. The authors describe two children with congenital TTP who presented with thrombocytopenia, hemolytic anemia, elevated LDH levels, and schistocytes on peripheral blood smear. In both children, the diagnosis of the disease was delayed despite neonatal histories significant for thrombocytopenia, anemia, and severe hyperbilirubinemia. Severely decreased ADAMTS13 activity (<0.1 U/mL), the absence of an inhibitor to the protease, and partial deficiency found in the parents confirmed the diagnosis of congenital TTP. The authors suggest that congenital TTP should be considered in the differential diagnosis for newborns presenting with severe hyperbilirubinemia, anemia, and thrombocytopenia. PMID- 15284597 TI - 128-channel EEG source imaging in epilepsy: clinical yield and localization precision. AB - The authors evaluated the feasibility, clinical yield, and localization precision of high-resolution EEG source imaging of interictal epileptic activity. A consecutive series of 44 patients with intractable epilepsy of various causes, who underwent a comprehensive presurgical epilepsy evaluation, were subjected to a 128-channel EEG recording. A standardized source imaging procedure constrained to the individual gray matter was applied to the averaged spikes of each patient. In 32 patients, the presurgical workup identified a focal epileptogenic area. The 128-channel EEG source imaging correctly localized this area in 30 of these patients (93.7%). Imprecise localization was explained by simplifications of the recordings and analysis procedure, which was accepted for the benefit of speed and standardization. In a subgroup of 24 patients who underwent operations, the sublobar precision of the 128-channel EEG source imaging was evaluated by calculating the distance of the source maximum to the resected area. This analysis revealed zero distance in 19 cases (79%). The authors conclude that high resolution interictal EEG source imaging is a valuable noninvasive functional neuroimaging technique. The speed, ease, flexibility, and low cost of this technique warrant its use in clinical practice. PMID- 15284598 TI - High resolution spatio-temporal EEG-MEG analysis of rolandic spikes. AB - Using high resolution EEG and MEG and a realistic volume conductor model, the authors investigated spatio-temporal aspects of the sources of spikes in children with benign rolandic epilepsy. A 64-channel EEG and simultaneous 151-channel MEG of interictal spike activity in five children all having general and/or focal seizures were recorded. A spatio-temporal multiple signal classification (MUSIC) analysis was performed on the spike data. Sources having a complex spatio temporal configuration as well as single stationary sources were found. Results for the EEG and MEG were different. In this group of five patients, both high resolution EEG and MEG revealed that in some cases sources well separated in space and time exist, whereas in other cases only single source activity can be resolved. For multiple sources, differences for EEG and MEG in timing and localization of activity suggest that sources are spatio-temporally distributed. Sources can propagate from initial activity in the finger/hand area around the central sulcus down to the mouth/tongue area. PMID- 15284599 TI - Stroboscopic artifact in digital video-EEG. AB - Combined digital video-EEG (DV-EEG) systems eliminate many familiar technical artifacts of older analog recorders; however, new and unanticipated technical issues are becoming evident. In this report, a case is described that identifies one of these technical limitations that could represent a pitfall to accurate data interpretation. An EEG was recorded on an 18-year-old man with history of photically sensitive generalized tonic-clonic seizures, revealing photoparoxysmal responses that appeared to outlast photic stimulation. However, in an attempted video-EEG correlation, the digital video recording showed variable appearance or absence of photic bursts that did not correlate with EEG photic tick marks, initially suggesting desynchronization between video and EEG signals. However, the absence of flashes seen on the video record resulted from stroboscopic artifact caused by mismatch between strobe frequency, video sampling rate, and video display characteristics. Stroboscopic aliasing is a DV-EEG specific artifact that can complicate accurate interpretation of photoparoxysmal responses. PMID- 15284600 TI - Changes in cortical dynamics in the preictal stage of a migraine attack. AB - Neurophysiologic studies suggest that migraineurs without aura have a dysfunction of cortical information processing in the pain-free interval. In this study, the advanced method of nonlinear multielectrode sleep-EEG analysis is used to investigate changes of cortical activity in the preictal time span. Five patients (four women, one man; age range, 29 to 58 years) experiencing migraine without aura participated in the study. The patients spent two blocks in the sleep laboratory. The first block was taken in a headache-free interictal time interval, and the second block when the onset of a migraine attack was most likely. After a nocturnal migraine attack, the patient was asked to mark the maximum of migraine pain in a surface-head scheme. The comparison of preictal and interictal EEGs enabled the authors to obtain a topographical view of changes in cortical dynamics. In each patient map, an area was found that displayed a pronounced focus indicating the region of maximum change in dimensional complexity. It shows a clearly recognizable correspondence with the scalp topography of the later pain perception. These findings indicate an association between cortical status and pain lateralization in the preictal time span. PMID- 15284601 TI - Late blink reflex changes in patients with pure sensory stroke due to geniculo thalamic infarct: a contribution to the long loop theory. AB - Seven patients with a pure sensory stroke due to a geniculo-thalamic infarct underwent blink reflex (BR) and median nerve somatosensory evoked potential studies to explore the mechanism subserving the R2 response. Both ipsilateral and contralateral R2 responses to stimulation of the affected side were significantly delayed in comparison with those obtained with stimulation of the nonaffected side (P < 0.001). Furthermore, in the five patients tested, cortical N20 following median nerve stimulation of the affected side was absent, delayed, or significantly reduced. These findings are consistent with the hypothesis of the transcortical generation of the late component of the BR. BR study appears to be a useful tool to assess long tract function, because changes have also been observed in patients with no demonstrable deficits on sensory examination. PMID- 15284602 TI - Effect of exercise on repetitive nerve stimulation studies: new appraisal of an old technique. AB - Repetitive nerve stimulation (RNS) is a simple and rapid method for evaluation of neuromuscular transmission defects. Although the effect of exercise in conjunction with RNS is well recognized, it has not been standardized in actual patient and control groups. In a prospective study over a period of 1 year, the authors evaluated the effect of exercise in conjunction with RNS in comparison with conventional 3-Hz RNS at rest in the clinical setting. Fifty-four patients who were referred for possible neuromuscular transmission disorders, in addition to 35 healthy control subjects, were studied. Amplitude and area decremental responses with RNS at rest and after 20 seconds of maximal exercise at 1-minute intervals up to 3 minutes were evaluated. The use of RNS with exercise resulted in additional diagnostic yield of up to 36.4% compared with conventional 3-Hz RNS at rest. The standardized use of exercise with RNS is advocated for increasing its diagnostic yield in the neurophysiologic laboratory. PMID- 15284603 TI - Maintaining constant voluntary force in generalized myotonia despite muscle membrane disturbances: insights from a high-density surface EMG study. AB - At medium and high force levels, patients with generalized myotonia (GM) cannot produce normal force because of a peculiar transient paresis. The authors have previously demonstrated that, during transient paresis, there is a disturbed propagation of muscle fiber action potentials along the sarcolemma, resulting in conduction block and paresis. At low force levels, however, these patients can produce a constant force. It is as yet unknown how patients with GM, despite these muscle membrane abnormalities, are able to produce constant force during isometric voluntary low-force contractions. Using high-density surface EMG (SEMG), the authors tested the hypothesis that in patients with GM, muscle membrane function is already disturbed at low force levels despite constant force and that abnormal motor unit recruitment acts as a compensatory mechanism to obtain normal force stability. High-density SEMG was measured on the biceps brachii at 20% of the maximal voluntary contraction (MVC) in seven patients with GM previously shown to have transient paresis at higher force levels. High density SEMG provides spatial and temporal information that is used to analyze propagation disturbances and recruitment strategies. In addition, needle EMG was performed in two patients simultaneously. The SEMG abnormalities included disturbed propagation of motor unit action potentials (MUAPs) over the muscle fiber membrane and abnormal motor unit recruitment patterns, pointing to central adaptation mechanisms. Prolonged periods of low SEMG amplitudes were sometimes present despite constant force, indicating that mechanisms other than motor unit recruitment also contribute to force preservation. During the periods of low SEMG, the density of myotonic discharges, recorded simultaneously with needle EMG, was not increased. Patients with GM can show, despite muscle membrane dysfunction, normal force stability. Abnormal motor unit recruitment as a compensatory mechanism occurs in patients with GM but cannot fully explain constant force. PMID- 15284604 TI - Rhythmic bilateral movement training modulates corticomotor excitability and enhances upper limb motricity poststroke: a pilot study. AB - The recovery of coordinated motor function after stroke onset has been associated with the practice of upper limb movements that required the activation of homologous muscles. This pilot study investigated whether repetitive bimanual coordinated movements enhanced upper limb corticomotor (CM) excitability and motor function poststroke. Patients practiced driving their paretic wrist through passive rhythmical flexion-extension by active flexion-extension of their unaffected wrist using purpose-built manipulanda over a 4-week period. Both preintervention and postintervention motricity was assessed using the upper limb Fugl-Meyer rating scale, and cortical maps of wrist flexor and extensor representations were derived from potentials evoked by transcranial magnetic stimulation. Five of nine subjects improved upper limb motricity in response to this novel active-passive bimanual movement therapy (APBT). Unaffected cortical map volume decreased, especially for a subgroup of five patients who had a postintervention increase in motricity. No change in unaffected map volume was revealed for the four patients who did not improve their postintervention motricity. No consistent shifts in cortical map center of gravity were revealed. These findings suggest that APBT can initiate an improvement in motricity that is accompanied by a balancing of between-hemisphere CM excitability. The findings justify the assessment of the rehabilitative effects of APBT in a homogeneous sample of patients poststroke. PMID- 15284605 TI - MEG in the presurgical investigation of temporal lobe epilepsy. PMID- 15284621 TI - Decision support: helping patients and families to find a balance at the end of life. AB - Terminally ill patients and their families face many decisions at the end of life that can sometimes be overwhelming. Nurses play a key role in providing decision support so that patients and their families can make timely decisions about their health care that reflect their individual needs and circumstances. The Ottawa Decision Support Framework can help nurses to assess patients' decision-making needs, provide tailored decision support and evaluate the effect of their interventions. The theoretical underpinnings of the model and its implications for palliative care clinical practice are discussed. PMID- 15284622 TI - Conferences for researchers--why meet? PMID- 15284623 TI - A phenomenological study of the lived experiences of people with lymphoedema. AB - Lymphoedema occurs in all age groups and is caused by lymphatic insufficiency. It is associated with a wide range of cancer- and non-cancer-related conditions. Researchers have explored the impact of lymphoedema on quality of life but most studies have focused on breast cancer-related lymphoedema or used specific quality of life tools. The study reported here used a phenomenological approach to explore the lived experience of 15 individuals with different types of lymphoedema. Findings highlight the uncertainty surrounding their diagnosis, the difficulties they experienced in accessing appropriate treatment and ways in which they dealt with having lymphoedema. Recommendations are made highlighting the need for increased awareness of lymphoedema and the importance of wider consideration of the emotional and psychosocial dimensions of this chronic condition. PMID- 15284624 TI - Multicomponent psychosocial support for newly diagnosed cancer patients: participants' views. AB - The diagnosis and treatment of cancer carries a heavy psychological burden, particularly in the first year after diagnosis. The purpose of this study was to investigate and report qualitatively on patients who had completed a 10-session programme consisting of education, social interaction, psychological support, exercise and complementary therapies for newly diagnosed cancer patients. A thematic analysis of data obtained from purposefully selected focus groups revealed that peer support was the most highly valued aspect of the service and that a variety of needs were being met. The relaxed atmosphere, sensitivity of staff and complementary therapies were particularly appreciated. The findings also indicated that patients would like the option of diversional activities and a period of structured group therapy in addition to the services offered and that the revision of information-giving procedures would enhance psychological support. Opportunities for staff development are identified. PMID- 15284625 TI - Palliative care stress in a UK community hospital: evaluation of a stress reduction programme. AB - In this study the Nurses Stress Scale and Nurses Coping With Stress Questionnaire were used to investigate work-related stress in 18 nurses providing palliative care in a UK NHS community hospital. These instruments were administered twice before and twice after a stress-reduction programme. In depth qualitative interviews were conducted before and after the programme and a 12-item questionnaire was used to assess whether the nurses found the programme useful. The findings indicated that most nurses did not find their work particularly stressful, and most felt well equipped to cope with palliative care stress. However, the interviews identified a small group of nurses who felt ill equipped to cope and routinely found their work stressful. The principal sources of support for both groups of nurses were family and friends at home rather than colleagues at work, and most felt there was little opportunity to share experiences and feelings with their colleagues. Likewise, few (only 33%) had found an opportunity to practice the relaxation skills they had learnt during the stress-reduction programme. This might explain why there was no evidence of any general improvement in stress and coping scores following the stress-reduction programme. Although the nurses enjoyed the programme and found it helpful, such programmes need to tackle contextual barriers to coping with stress as well as improving the individual coping skills of staff. PMID- 15284626 TI - One palliative care nurse's view of euthanasia: a social movement reflective of a self-serving generation. PMID- 15284627 TI - The matron role is to be developed in the community. PMID- 15284628 TI - Research shows nursing agencies in a positive light. PMID- 15284629 TI - Psychological, sexual and cultural issues for patients with a stoma. AB - Patients undergoing stoma surgery will more readily adapt to their new body image and way of life if they receive professional and voluntary input from the ostomy agencies such as the British Colostomy Association, from the preoperative stage through to rehabilitation and their return to the community. Nursing has moved away from mechanistic, task-oriented care to holistic care and, apart from the physical changes that a stoma will cause, there are other areas to be considered to improve the patient's quality of life after surgery. Patients undergoing stoma surgery experience a profound threat to their sense of physical integrity and self-concept with the change of body image in relation to bodily functions. Sexuality is an integral part of the whole person and is a highly complex phenomenon. Many patients find it difficult to discuss their sexual feelings, especially after a body image change and the nurse should be able to help patients identify and adapt to alterations in sexual self-concept. Cultural background plays an important part in patients' lives, including their beliefs, whether personal or religious, their perceptions of recovery, behaviour and concepts of and attitude towards the disease process. Nurses must combine sound, general information with open, respectful questions to the patient or family and be committed to responding flexibly and constructively. This article describes some of the potential problems that may be faced by patients after stoma surgery and increased knowledge of these areas will allow nurses to improve patient care and satisfaction. PMID- 15284630 TI - Case study: managing and caring for a patient undergoing stoma formation. AB - A 75-year-old man had attended his GP, as he was worried that his altered bowel habits and slight rectal bleeding was more sinister than haemorrhoids. This article describes the subsequent investigations, care and treatment that he received. It highlights how early detection, prompt treatment, appropriate care and management can enable a person to cope with the devastating diagnosis of bowel cancer, surgery and the need for a stoma. This patient was otherwise well which contributed both to the treatment decision and successful outcome. Unfortunately, not all patients have such an experience, and the measures taken to prevent complications are described. PMID- 15284631 TI - Improving the nutritional status of colorectal surgical and stoma patients. AB - This article describes an initiative to improve the quality of nutritional support and information provision to patients undergoing colorectal surgery and stoma formation. It was identified that several aspects of the established hospital routine and catering system overlooked the specific nutritional needs of this group of patients during the postoperative period. This led to the formation of a multidisciplinary strategy group responsible for focusing on the nutritional needs of both those with new stomas and those undergoing colorectal surgery without stoma formation. To ensure that patients had the opportunity to eat suitable foods at regular intervals, a 'colorectal patients' snackbox' was introduced, together with a strategy for individualized nutritional support and advice before admission and on discharge. PMID- 15284632 TI - Sexual counselling by cardiac nurses for patients following an MI. AB - Myocardial infarction (MI) patients often have unanswered questions about resuming sexual activity after this life-threatening event. Coronary care nurses can play an important role in counselling patients in such a sensitive area. However, sexual counselling is an area of nursing practice that is frequently neglected. This article discusses both the patients' and nurses' perceptions of discussing sexual concerns, and explores ways to increase nurses' awareness so that they can offer sexual counselling to patients post-MI. PMID- 15284633 TI - Nursing home manager who failed to provide a safe environment for clients. PMID- 15284634 TI - Clinical skills: assessing and treating shock: a nursing perspective. AB - This article outlines the pathophysiology associated with hypovolaemic, cardiogenic and distributive shock, and discusses how each of these might present clinically in the patient. Nursing assessment of a patient in shock is explored, and the use of tools such as the pulse oximeter is examined. The evidence base for a variety of interprofessional interventions is analysed, including fluid therapies such as blood transfusion, the use of crystalloids and colloids, and drug therapies such as the use of inotropic and vasoactive agents. The nursing role in managing the patient in shock is considered throughout. The importance of recognizing the clinical presentation of shock is highlighted, with an emphasis on understanding the pathophysiology and potential systemic effects. Treatment is discussed and covers: providing optimal oxygen therapy, appropriate patient monitoring and location of care, using effective communication skills, assisting with activities of living, psychological support, and working collaboratively to maximize the overall quality of patient care delivered. PMID- 15284635 TI - Confidentiality of patient information when preparing for death. PMID- 15284636 TI - Preventing cellulitis in older people with persistent lower limb oedema. AB - Persistent lower limb oedema is an underestimated problem in the elderly population. Oedematous tissue predisposes the individual to the development of bacterial infections. Accurate assessment of the underlying aetiology and application of compression hosiery could prevent cellulitis and reduce the risk of recurrent episodes. Awareness of the complications of leg oedema needs to be raised in the general population. Healthcare professions require education to enable the provision of safe, effective treatment to reduce morbidity relating to chronic oedema and cellulitis. The 'National Service Framework for Older People' and the 'British Lymphology Society's Framework for Education' could facilitate the development of national service provision to meet the needs of the population with chronic oedema. PMID- 15284637 TI - Role of the 'ordinary' nurse: concept of qualified caring. PMID- 15284638 TI - Cardiac rehabilitation: goals, interventions and action plans. AB - Cardiac rehabilitation services are now well established in the UK. There is a growing evidence base for cardiac rehabilitation interventions but, despite this, there remains much variation in service models and delivery. The 'National Service Framework for Coronary Heart Disease' sets out goals for cardiac rehabilitation interventions based on lifestyle change; however, many people do not have access to cardiac rehabilitation towards menu-driven approaches, which may suit individual need more than a standardized approach. Nurses play a crucial role in the delivery of cardiac rehabilitation services and hold the necessary skills and expertise so to do. Future challenges to cardiac rehabilitation include developing increasingly patient-focused services across the boundaries of primary, secondary and social care, and increasing patient and public involvement in services. PMID- 15284639 TI - Exploring issues of parental consent to research: a case-study approach. AB - In this article, a novice nurse researcher relates her personal experience of encountering a challenging situation while taking consent from the parents of a critically ill child. This critical incident raises issues for the researcher of ensuring validity of the consent process and ensuring recruitment of an unbiased sample when issues of language and culture could impede communication. The process of reflection allowed the novice researcher to gain greater insight into her responsibilities as a research nurse in relation to recruiting patients and to reflect on how she would integrate this into her future practice, thus enhancing her confidence in her role. PMID- 15284640 TI - Nursing needs to sort out the role of nursing assistants. PMID- 15284641 TI - Hospital-acquired infection rates continue to increase. PMID- 15284642 TI - The importance of regulation of healthcare assistants. PMID- 15284643 TI - Autism: developing a strategy for nursing to prevent discrimination. AB - The final phase of the Disability Discrimination Act (DDA) comes into force in October 2004. The DDA Code of Practice is a document to guide organizations in the implementation of and compliance with the Act. The DDA Code of Practice (2002) makes reference to specific impairments, such as hearing and visual impairments, and there are numerous examples of how 'reasonable adjustments' can be made to enable access to those so disabled. There is recognition that some impairments are hidden, e.g. learning disabilities and diabetes. However, no mention is made of those with autism. The impairments of autism including Asperger syndrome are also 'hidden impairments' as defined by the DDA and this will mean that it will not be immediately obvious as to how the Act will help nurses to amend their practice to enable people with autism full inclusion and access to health care. This article will explore practical implications and outline how nurses can develop their practice to be compliant with the Act and to ensure good 'health care' practice for people diagnosed within the autism spectrum. PMID- 15284644 TI - Exploring the evidence surrounding the debate on MMR and autism. AB - Autism is a poorly understood condition that would appear to be on the increase. There is currently much concern about a possible link between the measles, mumps and rubella (MMR) vaccination and autism which has resulted in a substantial reduction in the uptake of MMR, putting children at risk of three significant childhood diseases. This article looks at the evidence for a link between MMR and autism, finding that while a plausible hypothesis has been constructed, there is not substantive evidence for such a link and that the quality of this evidence is, in many cases, poor. This is due, in part, to the difficulties inherent in pre- and post-licensure vaccination research. These difficulties and the importance of vaccination and parental understanding of risk are discussed. PMID- 15284645 TI - People with learning difficulties who engage in self-injury. AB - This article represents some of the findings of the author's research into the relationship between self-injury and learning disability. It identifies the key theoretical discourses associated with the phenomenon, before elaborating on the main principles of each and identifying resultant intervention strategies. These interventions, psychotropic medication, mechanical restraint, and the behavioural approach are subsequently described. Case-study methodology was employed since this enabled the examination over time of these intervention strategies in the lives of individuals fulfilling the necessary criteria of persistent self-injury and significant communication difficulties. The findings of the research suggest a frequently piecemeal approach to self-injury, with arbitrary selection of behavioural intervention approaches, a continued reliance on powerful medication, and ambivalence concerning the use of mechanical restraint. Nurses were often skilled in working from a behavioural perspective, but were hindered by complex family circumstances and a failure to gain the confidence of direct care staff. There was also a lack of appreciation about the relationship between the individual and his/her self-injury, and recognition of the nature of the intransigence. PMID- 15284646 TI - Caring for people with learning disability using care management. AB - Current Government health policy is moving towards the delivery of services for clients with a learning disability through mainstream primary care services. At present there are difficulties in providing health services that meet the needs of clients. These include lack of resources (financial, physical and human), time and expertise. Yet clients with learning disabilities often have some of the most complex physical and mental health needs and they have a right to expect access to services that are responsive and sensitive to their requirements. In light of the current policy and stated difficulties, all service providers and stakeholders need to work together to review how services are provided and to negotiate shared resources. This article suggests adopting a care management approach, where clients are supported by different providers with a lead practitioner taking responsibility for assessment, planning and review in partnership with the client and his/her carers. PMID- 15284647 TI - Staff nurse whose lack of competence did not improve after further training. PMID- 15284648 TI - Clinical skills: evidence-based nursing care of people with rheumatoid arthritis. AB - Rheumatoid arthritis is a complex inflammatory disease with an unknown cause, uncertain prognosis and no known cure. The physical symptoms of this chronic disease can impact on the patient's psychological state and also affect the family unit and social/economic viability. Nursing patients with such a multifaceted illness is a skilled and complex task and it is imperative that the care provided is optimal, timely and underpinned by patient education. Evidence based nursing care founded on an in-depth knowledge of the disease and its treatments will help to achieve optimal patient outcome. PMID- 15284649 TI - Liability for death: manslaughter, murder and other criminal offences. AB - Case Scenario: Martha Brown by mistake administered twice the dosage of painkiller which was prescribed for Ted Snow. He convulsed and she realized immediately that something was amiss and called the doctor and pharmacist. However, he died before they were able to take remedial action. Since it was only a small mistake (admittedly with horrendous consequences), she is wondering what the consequences in law might be. PMID- 15284650 TI - Changing roles and titles: HCAs now deliver the care. PMID- 15284651 TI - Central venous catheters: choosing the most appropriate access route. AB - Intravenous therapy and care is a complex and intricate area of practice, which is being subsumed into the core role of registered nurses. As more patients become recipients of a vascular access device (VAD), particularly those requiring intermediate to long-term central venous access, it is important to ensure that not only the device but also the most appropriate vein meets their clinical physiological and psychological needs. There is much research and literature on the insertion and care of central venous catheters (CVCs) as well as the detection and treatment of complications. However, apart from a few small studies comparing the performance of devices placed either in the subclavian or jugular veins, there is little to guide doctors or nurses as to which vein is preferable, if secondary complications are to be avoided. This article will describe a number of primary and secondary complications associated with both the subclavian and internal jugular veins and how these can be minimized by selecting the most appropriate vessel. The article concludes with the author's suggestions for correct patient assessment in order that the correct vein is utilized. PMID- 15284652 TI - Chronic obstructive pulmonary disease: importance of diagnosis. AB - Chronic obstructive pulmonary disease is a common, smoking-related respiratory disease. It is responsible for considerable morbidity and mortality and accounts for a large amount of NHS resources. Despite this, it is underdiagnosed, often poorly managed and its impact on patients is underestimated (British Thoracic Society (BTS), 2001). The first step is to make an accurate diagnosis at as early a stage as possible. This article explores the prevalence, impact and presentation of the disease and how it can be identified and diagnosed. PMID- 15284653 TI - Difficulties in developing a nursing research culture in the UK. AB - Despite the obvious advantages to patient care and professional development, nursing research in the UK is relatively poorly developed and research activity in nursing lags behind many of the more established university disciplines. The reasons for this are complex and numerous. The purpose of this article is to explore the most pertinent issues. For example, there is currently a lack of appropriately qualified research-active staff, the 'research culture' in many departments is inadequately developed, dedicated research funding is insufficient and competing demands are commonly placed on nurse academics. In order for research activity to improve significantly many of these problems have to be properly addressed because patient care and professional credibility are at stake. PMID- 15284654 TI - New powers to deal with bad practitioners more easily. PMID- 15284655 TI - We must learn lessons from the Barbara Salisbury case. PMID- 15284656 TI - Clinical skills: nursing considerations in patients with faecal incontinence. AB - Faecal incontinence is a distressing and socially debilitating problem. Nurses are ideally placed to support patients and assist them in improving their quality of life. In order to provide the necessary holistic care the nurse needs to understand the nature of the patient's symptoms and be aware of a variety of management interventions. In this article the author outlines the various causes of faecal incontinence and highlights the importance of a thorough nursing assessment which takes into account the physical, psychological and social aspects of the symptoms. Planned care should be based on a firm knowledge base, but should reflect the needs of the individual. A good nurse-patient interaction facilitates this process and should be valued. PMID- 15284657 TI - Management of urinary incontinence in a patient with multiple sclerosis. AB - Patient care and clinical research looking at urinary incontinence associated with neurological disorders remains under-resourced. With heightened awareness, improved management decisions could be made that would lead to better patient outcomes. With the responsibility that clinical governance conveys, it is vital that an awareness of the risks associated with continence management are identified and minimized. These objectives can be achieved through the formation of a strong evidence base for practice - this will, in turn, allow better planning of care and easier anticipation of potential problems. This article reviews the literature to analyse the care received by a patient with multiple sclerosis and to consider what would have been best practice in this case, proposing alternative treatment options that could have been made available. PMID- 15284658 TI - Toilet training children with learning difficulties: what the literature tells us. AB - Nurses and health visitors are the professional group most likely to be involved in advising and supporting parents of children with disabilities (Bliss and Watson, 1992). Little research has been done into assessing and treating urinary continence difficulties of children with learning difficulties and many questions remain unanswered: what is the extent of the problem; what specific intervention do children with learning difficulties require to attain toileting skills; what expectation can the parent and clinician have that children with learning difficulties can be toilet trained and who is best placed to promote toileting skills? The main findings from the literature will support health and education professionals and carers who are involved in toilet training children with learning difficulties. PMID- 15284659 TI - The case of Barbara Salisbury and the role of the ward sister. PMID- 15284660 TI - Nursing management issues in hip and knee replacement surgery. AB - Total hip replacement (THR) and total knee replacement (TKR) are carried out for the relief of pain in hip or knee joints usually caused by osteoarthritis. Such replacements last for 10-15 years and therefore many nurses will care for patients with a THR/TKR even if that is not the primary reason for the patient seeking care. The different types of THR/TKR and how patients can be prepared for surgery are discussed. The major long-term complications of loosening or dislocation of the components of the THR/TKR and of infection are explored and the presenting symptoms are highlighted. The article is intended to be useful not only for orthopaedic nurses but also for nurses generally. PMID- 15284661 TI - A collaborative care approach to complex diabetic foot ulceration. AB - This article highlights the complex issues that surround the management of diabetic foot ulceration. It describes how the disciplines of podiatry and tissue viability came together to care for a patient who required complex wound management. The importance of collaborative working is highlighted, which has been shown to reduce amputation rates by 50% (Edmonds, 2002). Through exploration of a case study, certain issues emerged--wound infection, wound management, psychosocial factors and teamworking--and these are discussed. The link between diabetic foot ulceration and amputation is explored and the argument put forward that amputation should have been the first choice for this patient bearing in mind that 30% of amputees lose their second leg within 5 years (Geary, 2002). PMID- 15284662 TI - The patient's perspective of the nurses' caring role. PMID- 15284663 TI - An overview of meningitis: signs, symptoms, treatment and support. AB - This article aims to raise awareness of meningitis and its symptoms. It describes some of the practical support that can be offered to sufferers, their families and friends. Meningitis can kill and immediate action can and does save lives. The practice nurse or the nurse working in the accident and emergency (A&E) department may be the first person to be sought for advice; therefore, it is vital that he/she gains an insight and understanding of this disease in order to offer effective advice and take prompt action. A definition of the disease is given, and viral and bacterial meningitis are discussed. Epidemiological data are presented concerning rates of infection and mortality. The signs and symptoms are outlined and the mode of transmission and the treatment needed are also considered. The prognosis and possible complications are described. These include potential problems such as sensorineural deafness, raised intracranial pressure, the need for skin grafting and the amputation of limbs and/or digits. PMID- 15284664 TI - Stress caused to nurses following the death of patients: employer's duty. PMID- 15284665 TI - Nosocomial bloodstream infections in a neonatal intensive care unit. AB - A study was performed to assess the incidence density of, and to identify the risk factors associated with, nosocomial bloodstream infection (BSI) in a neonatal intensive care unit (NICU) in Athens. Twenty-four of 105 patients developed nosocomial BSI (22.9%). The incidence density of BSI was 10.9 per 1000 patient-days. A multivariate model showed that only two factors were significantly and independently responsible for nosocomial BSI: central venous catheter use and umbilical catheter use. Results showed that the incidence density rate was high and the factors that had most influence on the development of nosocomial BSI were associated with the treatment received by neonates during their stay in the NICU. Therefore, surveillance of nosocomial BSI and strategies such as infection control, nursery design and staffing should be implemented to reduce the incidence of these infections. This effort should be multidisciplinary, involving staff who insert and maintain intravascular catheters, and healthcare managers who allocate resources. PMID- 15284667 TI - Community matron gets cautious response. PMID- 15284666 TI - Study raises concerns about school children's health. PMID- 15284668 TI - Older community nurses: perspectives and prospects. AB - In common with the nursing workforces of the developed world, the NHS nursing workforce is ageing. Community nurses, in particular, tend to be older than their acute counterparts and this ageing could have a greater impact on this section of the workforce than on nurses in other sectors. This article reports on a study into the options, decisions and outcomes for nurses over the age of 50 years in the NHS. The study was funded by the Joseph Rowntree Foundation as part of a larger programme of work. Older nurses are a valuable part of the NHS workforce, being a repository of skill and wisdom. While this is generally recognized, and there are policies to support this, there is little recognition or action within the NHS. This study found that older nurses would like to see greater flexibility being shown towards them, in terms of their working hours and conditions of work, without harming their pension prospects. They would like to see return to practice courses and continuing professional development more geared towards their needs. Furthermore, they would like more information about their options. The article includes the experiences of some older nurses working in the community who remain, have left or have returned to the NHS. PMID- 15284669 TI - Fundamental principles of indwelling urinary catheter selection. AB - Catheter selection is a skilled element of continence care, particularly when the catheter is intended to remain in situ for prolonged periods. It is important to choose carefully, referring to catheter length, material, Charriere size and balloon infill volume, any of which may--if not attended to correctly--cause problems. This article gives some advice on catheter selection and outlines what may occur if certain issues are dealt with incorrectly. PMID- 15284670 TI - Multi-compartment medication devices and patient compliance. AB - Multi-compartment medication compliance devices are widely used in primary care. The aim of this review is to reveal whether they are effective in promoting adherence among non-adherent adults living at home. Searches were undertaken using two electronic databases (Medline (1966-2003) and International Pharmaceutical Abstracts (1970-2002)). Only randomized controlled trials (including crossover studies) were included in the review. Participants had to be non-institutionalized adults receiving one or more prescription medicines each day and displaying problems with adherence. Studies had to compare multi compartment medication compliance devices to standard packaging and outcome measures and to include either pill counts, biological assays and/or clinical response. Articles were selected if they described a follow up period of at least three months and demonstrated that over 80% of participants had completed the trial. Two studies were identified that met the criteria, reporting data on a total of 148 patients. The findings from the first study found diabetic patients receiving medication in a compliance device demonstrated better glucose control than patients receiving medication in standard packaging. The second study found compliance devices had no impact on blood pressure control in hypertensive patients. Further research needs to be conducted to assess the effectiveness of multi-compartment medication compliance devices in promoting adherence among non adherent adults living at home. PMID- 15284671 TI - Nurse prescribing: a case for clinical supervision. AB - This article will discuss the implications of nurse prescribing in mainstream primary health care and its impact on the fields of mental health and learning disabilities. Complexities and issues which require serious consideration by those nurses wishing to pursue such a specialist and extended role will be discussed in relation to these practice areas. Titchen's (1998) critical companionship model will be illustrated as an example of one framework for clinical supervision. This is to allow the processes, competencies and contextual issues to be explored by both novice and expert prescribers. The article concludes that for safe, effective and competency-based practice, clinical supervision also assists in mediating the professional and political aspirations in support of supplementary nurse prescribing. PMID- 15284672 TI - The issue of consent and children: who decides? PMID- 15284673 TI - Exploring the significance of place in community nursing. AB - There has been a dramatic shift in emphasis in health care in the UK in recent years towards the community setting. Care for someone in his or her own home takes place in a different context from caring for them in hospital, and requires a different approach. This article highlights a number of issues raised by this change, and suggests that there needs to be a focused effort to understand the impact of these issues on practice. PMID- 15284674 TI - Learning disabilities: lots of room for improvement. PMID- 15284675 TI - Recent progress in heart valve surgery: innovation or evolution? AB - Although heart valve surgery continues to evolve in a dynamic fashion, there is still no optimal solution for all patients. Minimally invasive surgery currently receives considerable attention but its value still needs to be determined. Progress has been made in valve repair, which now allows reconstruction in most patients with mitral valve disease. Reconstruction of the aortic valve is now also possible with results that are now comparable to those of mitral repair. In the future a wider application of repair procedures and further improvements of biologic valves can be anticipated not only to influence long-term results, but also the decision making process for conservative or surgical treatment. PMID- 15284676 TI - Perioperative assessment and management of patients with valvular heart disease undergoing noncardiac surgery. AB - Valvular heart disease in a variety of forms is not uncommon, especially among older patients undergoing noncardiac surgery, and can be associated with increased perioperative cardiac risk. Patients with aortic stenosis are at greatest risk, although other valve lesions also can pose the risk of increased perioperative morbidity. During preoperative evaluation, attention to the presence, nature and severity of valvular heart disease allows appropriate perioperative monitoring and therapy with a goal to minimize the risk of perioperative cardiac morbidity and mortality associated with noncardiac surgery. Appropriate antibiotic prophylaxis reduces the risk of infective endocarditis. Finally, some patients with valvular heart disease and all patients with a mechanical valve prosthesis require long-term anticoagulation, which must be managed during the perioperative period. PMID- 15284677 TI - Aortic valve disease: a practical clinical approach. AB - The diagnosis and treatment of aortic valve disease can be challenging for the clinician. Aortic regurgitation (AR) may be evaluated with Doppler echocardiography or catheterization. Medical therapy consists of vasodilators. Surgery may be appropriate for patients with symptoms, left ventricular dilatation or diminished left ventricle systolic function. Aortic stenosis (AS) can be evaluated with the same modalities as AR. There is evidence that statin therapy may slow the progression of AS. Contrary to conventional wisdom, vasodilators are safe and effective in certain patients. Surgical valve replacement is performed for symptoms and in certain asymptomatic patients. PMID- 15284678 TI - New aspects of infective endocarditis. AB - The current incidence of infective endocarditis (IE) is estimated as 7 cases per 100,000 population per year and continues to increase. The prognosis is significantly influenced by proper diagnosis and adequate therapy. In cases with unconfirmed IE, transesophageal echocardiography is the imaging technique of choice. Culture-negative endocarditis requires either termination of antimicrobial treatment initiated without mircobiological test results and reevaluation of blood samples or serological/molecular biological techniques to identify the causative organism. Antimicrobial therapy should be established only after quantitative sensitivity tests of antibiotics (minimal inhibitory concentrations, MIC) and guided by drug monitoring. In the first 3 weeks after primary manifestation, an index embolism is frequently followed by recurrencies. If vegetations can still be demonstrated by echocardiography after an embolic event, a surgical intervention should seriously be considered. Cerebral embolic events are no contraindication for cardiac surgery, as long as a cerebral bleeding has been excluded by cranial computed tomography immediately preoperatively and the operation is performed before a significant disturbance of the blood-brain barrier (<72 hours) has manifested. A significant prognostic improvement has also been demonstrated for patients with early surgical intervention suffering from myocardial failure due to acute valve incompetence, acute renal failure, mitral kissing vegetations in primary aortic valve IE, and in patients with sepsis persisting for more than 48 hours despite adequate antimicrobial therapy. PMID- 15284679 TI - Genetic risk factors in myocardial infarction at young age. AB - The role of genetic susceptibility to coronary artery disease (CAD) seems to be quite important in young patients. In the last years the attention has been focused on polymorphisms influencing some biological functions (coagulation and fibrinolysis, platelets, vascular function, lipid metabolism, inflammation). The study of prothrombotic polymorphisms has kindled a deep interest. The role of atherosclerosis and thrombosis is different in the different ages. In all the studies we examined, the polymorphism G20210A in the prothrombin gene was associated with an increased risk of acute myocardial infarction (AMI) in young people, especially when other risk factors were present. Contradictory results have been found in the studies on Factor V Leiden: according to many authors the activated protein C resistance (APCR) is associated with an increased risk of AMI only in smokers, above all if women. On the other hand, some polymorphisms of the Factor VII gene seem to be protective. Young AMI could be also caused by a reduction of the fibrinolytic activity, as it was found when the allele 4G in the promoter of plasminogen activator inhibitor (PAI) gene is present. The attention has also been focused on the effects of variations in genes that influence platelet functions. According to a metanalysis of studies published up to 1999, there is no association between the polymorphism PlA1/A2 of the GP IIIa gene and young AMI, whereas there is doubt about the role of the polymorphism in the GP IIb e GP Ib genes. Moreover, it seems to be present an association with the polymorphisms in the thrombopoietin gene (C4830A and A5713G). Also the role of some genes coding for proteins influencing the vascular functions has been valued. Few studies were performed on genetics of the renin-angiotensin aldosterone system and the results are insufficient and contradictory, such as those about the association between the polymorphism G894T in the eNOS gene or the polymorphism C677T in the MTHFR gene and young AMI. Genes coding for proteins involved in the lipid metabolism have been closely examined. Many polymorphisms were discovered in the Apo B gene: the variant C-516T was found to be associated with increased LDL levels, whereas the results about the association between this and other polymorphisms in the same gene (I/D of LAL sequence, PvuII, MspI, Asp4311Ser) and young AMI are discordant. On the other hand, the variant e4 of the ApoE gene was associated with an increased risk of AMI at young age in many works. In the last years, a particular interest has kindled the study of the relationship between inflammation, atherosclerosis and CAD. Even if the studies performed are few, it was found an association between young AMI and polymorphism C-260T in the CD14 gene, between coronarics atherosclerosis and polymorphism A516C in the E Selectin gene or polymorphisms Leu125Val and Ser563Asn in the PECAM1 gene. PMID- 15284680 TI - Plasma adrenomedullin concentrations in patients with renovascular or malignant hypertension. AB - AIM: The aim of the present study was to investigate the behaviour of plasma adrenomedullin (AM), a hypotensive peptide, in patients with malignant (MHT) and renovascular hypertension (RVH), 2 pathologic conditions in which renin angiotensin system (RAS) is activated and to compare them with those in essential hypertensive patients (EHT) and normotensive subjects (NS). METHODS: Three groups of hypertensive patients have been studied: group 1 (4 patients with MHT), group 2 (10 patients with RVH), group 3 (24 patients with EHT) and 21 patients NS were enrolled as controls. In all patients, 10 ml vein blood samples were collected and AM was measured with specific radioimmunoassay. RESULTS: As expected, the plasma renin activity (PRA) levels in the RVH and MHT patients were significantly higher (p<0.0001) respect to NS and EHT. The mean plasma AM (+/-SD) concentrations in EHT (22.5+/-9.1 pg/ml) and RVH (46.8+/-19.4 pg/ml) were significantly (p<0.0001) higher than those in NS (13.7+/-6.1 pg/ml). The plasma AM concentrations were further elevated in MHT patients (107+/-12.3 pg/ml) and were significantly higher (p<0.0001) than those in EHT and RVH patients. In the MHT patients the elevated plasma AM levels, similarly to blood pressure and PRA values, declined after antihypertensive treatment (36.8+/-5.7 pg/ml; p<0.01). CONCLUSION: In conclusion, the findings demonstrated that the plasma AM concentrations were increased in proportion to the severity of arterial hypertension. RAS was activated in patients with MHT and RVH suggesting that activation of this system may contribute to increased in the plasma levels of AM. PMID- 15284681 TI - Left ventricular diastolic impairment during coronary arteriography with a non ionic contrast medium. AB - AIM: The aim of this study was to assess the influence of coronary arteriography with the use of a non-ionic low molecular monomer (iopromide) on left ventricular function. METHODS: Fifty consecutive patients with coronary artery disease (CAD) and normal left ventricular ejection fraction were studied by coronary arteriography for a stable or unstable coronary syndrome by using iopromide. They were divided into 2 groups: group 1, patients with one vessel disease; group 2, patients with multiple vessel disease. A >50% reduction of the lumen diameter by on-line quantitative angiography was considered a significant coronary stenosis. Coronary arteriography was performed by hand injection of 5 ml of iopromide avoiding the use of nitrates during the procedure. Doppler echocardiography monitoring was performed immediately before the coronary arteriography and at the end of the last coronary injection. The following parameter were recorded: E peak velocity (E) (cm/s), A peak velocity (A) (cm/s), E/A ratio, E deceleration time (EDT) (ms), isovolumic relaxation time (IRT) (ms), and left ventricular ejection fraction (EF) (%). RESULTS: No complications were observed during the procedures. A mean amount of 40+/-8 ml of iopromide was used. No significant variation of heart rate and arterial pressure was shown during coronary arteriography. No changes were observed either for E, A, E/A ratio or for left ventricular EF in any group of patients. A significant increase of EDT and IRT in comparison with baseline values was documented only in group 2 (from 140+/-77 to 199+/-44 and from 98+/-33 to 144+/-44, p<0.01), returning to baseline values after 10+/-3 minutes. A positive correlation was observed between EDT and IRT shift from baseline values (r=0.77; p<0.01). CONCLUSION: In conclusion, iopromide temporarily impairs left ventricular diastolic dynamics during selective coronary angiography, but only in patients with multivessel CAD. PMID- 15284682 TI - Contrast enhanced imaging of left internal mammary artery after vasodilation improves the evaluation of the bypass graft. AB - AIM: The aim of the study is to evaluate patency and flow reserve by echocardiography in arterial grafts using the left internal mammary artery (LIMA) to the left anterior descending coronary artery (DA). The main limitations in performing this study routinely are the weakness of the Doppler signal and the exact chest localization of the graft. The purposes of the study were: to verify the feasibility of the echo color Doppler method on LIMA; to verify which between the parasternal or supralavicular view is the better approach to obtain a clear signal; to verify the increase of systolic and diastolic flow velocity of LIMA in basal conditions and after infusion of dipyridamole, and if the visualization of the Doppler signal improves after contrast infusion. METHODS: Twenty patients (all males, mean age 63+/-7.8 years) with previous coronary artery bypass in the last 10 years, and without any significant stenosis in the left mammary artery graft as proved by a recent coronary angiogram (within 6 months), were selected for our study. LIMA was evaluated by two echocardiographic approaches. Patients were studied at rest and after pharmacological infusion of dipyridamole using the protocol of 0.56 mg/kg in 4 minutes. Contrast enhancement was infused in order to improve the Doppler signal using Levovist contrast agent at rest and after vasodilatation. Diastolic and systolic peak flow velocities, their ratio and the diastolic and total velocity time integrals were evaluated. RESULTS: The results showed that using the supraclavicular approach we obtained the visualization of the graft at rest in all patients (100%) and using the parasternal approach in 19 out of 20 (95%) even without contrast injection. At rest, the diastolic and systolic peak flow velocities were 0.417+/-0.133 m/s and 0.368+/-0.1291 m/s; their ratio (diastolic/systolic) was 0.882+/-0.7362. The overall and diastolic velocity time integrals were 0.1571+/-0.0645 m and 0.2232+/- 0.0701 m. After dipyridamole infusion we observed in all patients an increase in diastolic and systolic peak flow velocities as expected by 0.582+/-0.342 m/s (p<0.005) and 0.73+/-0.427 m/s (p<0.005). Contrast injection at rest and after peak dipyridamole infusion showed a better and clearer Doppler signal of the graft allowing an easier evaluation of the velocity curves in all patients. In fact using the association dipyridamole-Levovist the velocity ratio and the total and diastolic velocity time integral values were 1.268+/-0.368 (p<0.05), 0.3492+/ 0.131 m (p<0.05) and 0.2309+/-0.153 m (p<0.05). CONCLUSION: In conclusion, this new echo-color-Doppler approach seems to be valid for the evaluation of the patency rate and flow reserve of the internal mammary artery graft, and helps to better select patients who really need angiography. PMID- 15284683 TI - When should asymptomatic patients with combined severe aortic stenosis and aortic insufficiency undergo valve replacement? A clinical case. AB - Indications to prosthetic aortic valve implantation in patients with aortic stenosis or aortic regurgitation or both stenotic or regurgitant aortic valve, who present without symptoms, are controversial. We present the case of an asymptomatic patient with combined severe aortic stenosis and an equally important insufficiency, undergoing surgery for valve substitution with a bileaflet prosthesis. After surgery he was treated with warfarin according to the doses recommended and underwent follow-up with clinical and echocardiographic exams. Eight months after intervention the patient had an embolic stroke with aphasia and right hemiplegia, despite the therapeutic level of INR. At present, even though he has partly recovered motor function, he reports a noteworthy decline in life quality, because of the persistent speech difficulties. We use this case as the starting point for a discussion of the chance of referring patients affected by aortic valvulopathy to valve substitution, in the absence of symptoms. PMID- 15284684 TI - [Congenital heart diseases in adult age and primary cardiomyopathies at the referral centers]. PMID- 15284685 TI - Knowledge and attitude of clinical students on problem based learning. AB - An intervention study was carried out in Mymensingh Medical College between December 2002 to November 2003 to determine knowledge and attitude of clinical students on problem based learning (PBL) before and after exposure to a PBL course. This is an intervention study. A total of 17 health problems were discussed in the integrated small group tutorials in the departments of Pediatrics and Medicine. Two problems were discussed in a week & each PBL class was lasted for 2 hours. Pretest was done before exposure of the students to the PBL course using self-administered questionnaire and posttest was done immediately after finishing the PBL course. Significance of proportions was calculated by Chi-square (X2) test. PBL course increased knowledge of clinical students on PBL significantly (P < 0.001). Increased number of clinical students also stated that PBL is effective in problem solving (70% before and 97% after the PBL course), and the difference is highly significant (P < 0.001). Increased number of students (about 72% before & 83% after the PBL course) stated that PBL is better than traditional ward teaching (P < 0.01). Majority of our students (about 61% before & 88 % after the PBL course) stated that PBL enhances self directed learning (P < 0.005). A good number of students (about 63% before & 81% after PBL course) recommended PBL to be included in Undergraduate Medical Curriculum (P < 0.05). It can be concluded that exposure of students to PBL course can improve knowledge, attitude & practice of clinical students significantly. Regular exposure (at least twice a year) of clinical students to symposium centered on PBL is recommended PMID- 15284686 TI - Sensitivity of splenic and bone marrow aspirate study for diagnosis of kala-azar. AB - Fifty adult patients of kala-azar were included in this prospective study from Medicine Unit-I of Mymensingh Medical College Hospital. Splenic and bone marrow aspiration were done simultaneously to compare the sensitivity and other related merits and demerits of each procedure. Splenic aspiration appeared to be more sensitive procedure than bone marrow aspiration. Leishman - Donovan (LD) bodies were found in 90 percent and 72 percent of the spleen and bone marrow aspirates respectively. Splenic aspiration was found more acceptable to patients (96%) as it was less painful. Both the procedures were hazardless. There was no major complication except mild pain after splenic aspiration; even a few patients had history of epistaxis. Splenic aspiration was also more acceptable by the physicians because of an easy and reliable diagnostic procedure. PMID- 15284687 TI - Ketosis resistance in under thirty diabetic subjects. AB - Young onset diabetic subjects in tropical developing countries include a group of subjects who exhibits a characteristic ketosis resistance termed as Malnutrition Related Diabetes Mellitus (MRDM) by the WHO Study Group. The mechanism for this resistance to ketosis is still uncertain. To understand this mechanism we have studied the serum responses of glucose, non-esterified fatty acid (NEFA) and triglyceride (TG) to intravenous fat emulsion in newly diagnosed 8 fibrocalculous pancreatic diabetes (FCPD) and 11 low insulin secretory (LIS) subjects under 30 years of age along with 27 age-matched Non Insulin Dependent Diabetes Mellitus (NIDDM) subjects. Overnight fasting subjects were given a 90 min infusion of intralipos 10% (2.5 mg/kg body weight/min) and serum was collected at 0, 60, 90, 120 and 150 min. The fasting NEFA in the 3 groups were almost similar (micromol/l, M +/- SEM: 486 +/- 58, 564 +/- 76 and 559 +/- 34 in FCPD, LIS and NIDDM respectively). Fasting TG also showed a close similarity among 3 groups (mg/dl, M+/-SEM: 117 +/- 11, 110 +/- 22 and 123 +/- 4 in FCPD, LIS and NIDDM respectively). Intravenous fat caused a steady rise of NEFA as well as TG in all groups during the 90 minutes of infusion followed by a gradual fall. No two groups significantly differed regarding NEFA and TG at any time point. Fasting glucose was markedly higher in FCPD (22.9 +/- 2.5, mmol/l, M+/-SEM) and LIS (20.8 +/- 1.6) than NIDDM (11.0 +/- 1.0). In all the 3 groups glucose showed a slow but steady fall. Fasting C-peptide was very low in FCPD (0.42 +/- 0.08, ng/ml, M +/- SEM) and LIS (0.55 +/- 0.09) whereas it was within normal range in NIDDM patients (2.99 +/- 0.24). The results suggest the following: (a) Depleted body fat store do not lead to a decreased supply of NEFA in FCPD and LIS subjects at the fasting state; (b) Increased supply of NEFA in these subjects lead to a normal esterification response as evidenced by a parallel rise of TG; (c) Inspite of markedly low level of the antilipolytic hormone insulin, FCPD and LIS subjects are capable to maintain NEFA and TG responses similar to NIDDM subjects. This may indicate that factor (s) other than substrate and esterification is (are) probably involved in the ketosis resistance of FCPD and LIS subjects; and (d) Although FCPD and LIS differ regarding generalized pancreatic damage (which raises the possibility of involvement of glucagon producing alpha-cells in the FCPD group) the two groups do not differ regarding the ketogenic substrate and esterfication responses. PMID- 15284688 TI - Histopathological types of malignant lesions of esophagus and stomach. AB - Upper part of gastrointestinal tract is common site of malignancy. Histological types of malignancy of esophagus and stomach may vary in different countries and also in different regions in the same countries. To find out the frequencies of different histological malignancies in Mymensingh region of Bangladesh, we analyzed the histopathological reports of 259 biopsy specimens examined in one pathology laboratory in Mymensingh town during the period from 6th January 2002 to 29th April 2004. Endoscopic biopsies were 234 (90.35%) and surgical biopsy specimens were 25 (9.35%). Male patients were 193 (72.52%) and female patients were 66 (25.48%). Specimens of esophagus were 73 (28.19%) and of stomach were 186 (71.81%). Out of 69 (94.53%) adequate samples of esophagus 46 (74.19%) were squamous cell carcinoma and 16 (25.81%) were adenocarcinoma. Adequate samples of stomach were 177 (95.16%) in which malignancies were found in 119 (67.23%). All the malignant tumor of stomach were in adenocarcinoma of which 82 (68.91%) were intestinal type, 17 (14.29%) were diffuse type and 20 (16.81%) mixed type. Intestinal type of gastric adenocarcinoma was found to be significantly associated with male patients (p < 0.005). From the present study it is suggested that in the Mymensingh region the common form of malignant lesion of esophagus may be squamous cell carcinoma followed by adenocarcinoma. Adenocarcinoma is the most common type of gastric malignancy of which intestinal type is more common PMID- 15284689 TI - Placenta previa and it's relation with maternal age, gravidity and cesarean section. AB - The placenta provides the essential connection between the mother and the developing fetus. Placental position were routinely mentioned in an ultrasound report starting from early second trimester to the end of third trimester when asked for pregnancy evaluation. The aim of this study was to see the prevalence of lower segment placenta (placenta previa) and its relations with previous cesarean section delivery, parity and maternal age. The study conducted in Centre for Nuclear Medicine and Ultrasound (CNMU) Mymensingh in a period from January 2001 to December 2002. About 2536 pregnant women (those included in this study) underwent ultrasound examination during pregnancy at third trimester. The prevalence of lower segment placenta was 1.34%. The highest prevalence of placenta previa (2.58%) was seen in 3rd and higher gravida group. Also the highest prevalence were seen 30 yr. and above age group in compare to below 30 yr. age group. No increased prevalence of placenta previa were seen in previous cesarean section (C / S) delivery group (0.65%) in compare to normal delivery group (1.97%). From our study it was seen that development of lower segment placenta has relation with increased number of gravidity and maternal age but no increased prevalence were seen in subjects with previously done cesarean section PMID- 15284690 TI - Attitude of the parents seeking service from the paediatricians. AB - The overuse of antibiotics and other medicines have been standing a dangerous proposition and researchers are coming in fore front analyzing and assessing the aftermath of years of misperceptions and inappropriate usage of drugs prescribed by the pediatricians. Two hundred and two parents from the community as well as from the outpatient department of medical college hospitals were interviewed, to study their attitude while seeking for the health service for their children. Data were collected through a face-to-face interview using a structured questionnaire. One hundred and ninety nine (82.7%) of the parents interviewed were mothers and only 3 (1.5%) were fathers. One hundred and sixty-seven (82.7%) parents were from the urban population and the rest 35 (17.3%) were from rural areas. The mothers mean stay in the educational institution was 11.8 years and that of the father's was 13.1 years. About 34.7% parents had to seek for doctor's advice at least once or twice a month and around 43% had to visit doctor's clinic. Most (66.8%) of the parents were from the family earning tk.6000 to tk.20000 a month, 19.8% and 13.4 % were from the families earning below tk.6000 and above tk.20000 respectively. A vast majority of the parents expressed their preference for advice to medicines. Among the medicines they like antibiotics more than vitamins. Majority of the parents showed their interest to spend more time and to have less medicine. For anorexic children parents preferred advice to medicines. An overwhelming majority (98%) of the parents was interested to get advice and ORS for diarrhea. For ARI also a larger number (56.1%) of parents opted for advice only and no medicine at all but 37.1% kept their option for both. A large majority of the parents were preferring more frequent visit and having fewer medicines than the conditions of more medicines with less frequented visits. PMID- 15284691 TI - Interleukin-6 (IL-6) and tumour necrosis factor-alpha in open and laparoscopic cholecystectomy. AB - Injury to the human body alters normal physiology across several systems and these alterations are proportional to the extent of the injury. Physiological response to minimally invasive surgery appears to be different than those of traditional open surgery. Acute phase protein response appears to be one example. The important cytokines that are known as major mediators of acute phase response are interleukin-6 and TNF-alpha. Thirty patients were studied in which 14 underwent open cholecystectomy and 16 laparoscopic cholecystectomy. Three blood samples were taken from each patient, one pre-operatively and 2 post operatively at 4 and 24 hours. Interleukin-6 and Tumour Necrosis Factor-alpha (TNF-alpha) were raised significantly in post operative blood sample in both groups but the rise was much more in open group than laparoscopic group. This suggest less stress response in laparoscopic group which also showed a direct effect on patient convalescence in terms of less pain, less analgesic requirement and shorter hospital stay PMID- 15284692 TI - Outcome of patients treated with the endoscopic dacryocystorhinostomy. AB - DCR allows an abstracted lacrimal drainage system to be drained into the nasal fossa. Since the development of endonasal endoscopic surgical technique, the endonasal approach presents it self as an alternative choice to the conventional external approach as in the former skin scar can be avoided. We performed DCR of 50 cases in endonasal endoscopic approach in Mymensingh Medical Collage Hospital. The age range of patient was found between 7 to 35 years, average 21 & male female ratio was 1:1.5 with follow up to 2.10 years with high success rate. Associated nasal disease correction septoplasty was done in 7 cases. In each & every case silicon tube was introduced & removed after 3 to 6 months. The only complication was periorbital injury in 5 cases, punctal tear in 2 cases and granuloma formation in 2 cases. Overall success rate was 86%. PMID- 15284693 TI - Effect of fenugreek seeds on the fasting blood glucose level in the streptozotocin induced diabetic rats. AB - In this experiment defatted Trigonella foenumgraecum (fenugreek seeds/methi seeds) has used as the antidiabetogenic herbal medicine. The experiment was carried out in Bangabandhu Sheikh Mujib Medical University and BIRDEM from 1996 to 1998 on a total of 58 Long Evans rats of either sex. They were 50-60 days young rats with average body weight 72-174 gm. Among the total, 10 rats were treated with only vehicle called as non-diabetic control rats, 48 rats were treated with Streptozotocin (STZ) at a dose of 90 mg in 1 ml of citrate buffer solution per kg body weight, among which 20 were diabetics. Ten (1 died, 1 escaped) diabetic rats were again treated with fenugreek called as Fenugreek treated diabetic rats and the rest 10 diabetic rats were called as diabetic control rats. The change in the mean fasting blood glucose (FBG) level in different groups of rat from day 5 from streptozotocin injection were higher in diabetic control group and in fenugreek-treated diabetic group than in non diabetic control group. The FBG level on day 13 the mean in non-diabetic control group was 5.21 mmol/L. In diabetic control group and in fenugreek-treated diabetic group the mean FBG level were 24.33 mmol/L and 9.89 mmol/L respectively. So, from this experiment it may be concluded that fenugreek decreases the FBG level considerably by improving diabetes mellitus. PMID- 15284694 TI - Road traffic accident among motor vehicle drivers in selected high ways. AB - To assess the knowledge of the motor vehicle driver about the causes of road traffic accidents. This cross sectional study was conducted among the 107 motor vehicle drivers of selected Bus & Truck terminal & taxi stands of Dhaka city. The study was carried out during April to June 2002 at the Department of Health Promotion & Health Education of NIPSOM. The study showed about 55.1% learned driving from a friend. Out of 107 respondent 20.6% showed very high speed is one of the reason for road traffic accident.39.3% claimed for poor maintenance of roads. Regarding engine/vehicle defect 46.7% said head light defect. Increased earning is one of the reasons of very fast driving 35.5% respondent opined. These may be the reason of accident proneness of the society along with factor like family pressure, job dissatisfaction etc. Majority of the driver who took part in this study used light vehicle 54.2% and the rest 30.8% were heavy and 15% like medium light vehicle. Most of the respondent 67.3% was working as professional over for 3-6 years. 41.1% for 3 and 26.2% for 6 years. There were significant relationship (p < 0.002) between very fast driving and defective road and also engine defect were highly significant association (p < 0.005) with road traffic accident It is revealed that formal education, driving license through Bangladesh road traffic authority without unnecessary botheration, proper implementation of traffic law, training of drivers and increase level of public awareness through mass communication could reduce the road traffic accident PMID- 15284695 TI - Iron status and infants feeding practice in children with anaemia. AB - A cross sectional study was carried out in the then IPGM&R now (Bangabandhu Sheikh Mujib Medical University, Dhaka, Bangladesh) to detect the pattern of feeding practice among the anemic children who were below 2 years of age and to find out a relationship between feeding practice and iron status. Hemoglobin was measured on finger prick blood samples using Haemoglobinometer. A peripheral blood film, and serum-ferritin was estimated by micro particle enzyme-immonoassay (MEIA). A dietary questionnaire was completed with particular emphasis on the type of feeding, breast and formula and at which age the weaning was introduced. 140 clinically suspected anemic patients were selected randomly for this study. Among them 111 (79.3%) patient had hemoglobin value ranging between 7.2-6.4 gm and their serum-ferritin level at or below 12 ngm/ml. It was observed that female children were predominantly affected and incidentally all patients were malnourished. In this study it was found that babies with an exclusive diet of breast milk were mildly anemic and not deficient in iron. On the contrary infants with mixed feeding habits or prolonged breast-feeding without weaning at all or babies devoid of any breast milk had moderate to severe anaemia and all were deficient in iron. PMID- 15284696 TI - Corneal foreign body--an occupational hazard. AB - An attempt is made to describe the ocular surface injury by foreign body among the people those who are working in welding, grinding, hammering factory. Corneal foreign body which is usually iron in nature is the most common problem among the people those who are working in these factories. Clinical presentation, clinical pictures, and management are discussed. Corneal foreign body is a common cause of ocular morbidity and loss of working hour in the work place. Most of the victims do not use protective glass during work. We conclude that protective glass will useful in reduction of these accidents and alertness of both ophthalmologists and their patients are necessary. PMID- 15284697 TI - Studies on congenital abnormalities and related risk factors. AB - Congenital malformations were studied prospectively covering 11680 consecutive deliveries. The overall incidence of malformations was 2.3%. Musculoskeletal system was the most commonly involved system. The incidence of malformation was higher in still borns, premature, low birth weight babies and those with positive heredo-familial history. It was also higher in babies, born to mothers who were more than 35 years of age and gravida four and above. Consanguinity of marriage, drugs and hormone ingestion during pregnancy, antenatal complication like hydramnios, pre eclamtic toxemia, gestational diabetes was associated with high incidence of congenital malformations. PMID- 15284698 TI - Patch test for the detection of contact allergens. AB - Patch Test was done in 340 suspected allergic contact dermatitis patients having different ages of both sexes during the period of January 2000 to December 2003 in the department of Dermatology and Venerology, Bangabandhu Sheikh Mujib Medical University (BSMMU), Dhaka. Of these 340 cases 145 (42.65%) were male and 195 (57.35%) were female. Among these 340 cases, 250 (73.53%) cases were tested with international standard series allergen (ISS) and 90 (26.47%) cases with cosmetic series allergen (CSS), of which patch test was positive in 125 (50%) cases and 65 (72.22%) cases respectively. The 125 positive cases with ISS consisted of 56% male and 44% female. In 65 CSS positive cases 90.77% were female and 9.23% were male. Highest incidence of positive patch test was found in 61.54% cases in the age group of 30-39 years in ISS and 71.43% cases in the age group of 20-29 yrs in CSS. In ISS--Potassium Dichromate (19.2%), Fragrance mix (17.6%), Nickel Sulphate (15.2%) and PPD (11.2%) were found to be the common allergen and in CSS--Octyl Gallate (27.69%), Getrimide (18.92%), Balsum of Peru (15.38%), Thiomersal (12.31%) were found to be the common allergens. PMID- 15284699 TI - Studies on serum triglyceride level in patients with myocardial infarction. AB - Myocardial Infarction (MI) is the most common form of heart disease and the single most important cause of premature death in the developed and developing world. Unfortunately the incidence of the condition is increasing rapidly in many developing countries like Bangladesh. Effort should therefore be taken to minimize the risk factors of MI. Large scale randomized clinical trials have shown that lowering high triglyceride concentration mainly by drugs reduces the risk of cardiac events like MI. So the present work has been designed to see the serum triglyceride levels in normal healthy subjects, to compare serum triglyceride levels in patients with MI and those of healthy subjects and to evaluate the association of serum triglyceride in Bangladeshi MI patients. The present study was carried out in the Department of Biochemistry, BSMMU in collaboration with Department of Cardiology, BSMMU and NICVD, Dhaka during the period of July 2001 to December 2002. A total of 50 subjects were selected, Group A (30 subjects of control ) and Group B (20 subjects of test ). The mean level of serum triglyceride in control subjects were 117.07 +/- 32.41 mg/d1 and in test subjects were 176.87 +/- 37.15 mg/d1. So the present study showed that serum triglyceride level is significantly higher in patients with MI. From the present study, it is difficult to draw any definite conclusion but suggested that high serum triglyceride concentration is a cause of the incidence of MI. PMID- 15284700 TI - A case report on Klinefelter syndrome. AB - An 18 year old boy presented with small genitalia, failure of eruption of secondary sex hairs, female like voice with eunachoid body habitus, bilateral gynecomastia, infantile external genitalia, small testes and poorly developed musculature. He was diagnosed as a case of 47XXY Klinefelter syndrome on the basis of hormone assay and karyotyping. He has given androgen replacement therapy with the aim to relieve symptoms of androgen deficiency, to reproduce physiological levels of plasma testosterone and to prevent long term consequences of androgen deficiency. PMID- 15284701 TI - A case of molluscum contagiosum. AB - Molluscum contagiosum is a benign contagious disease caused by a poxvirus. The virus proliferates within keratinocytes and forms intracytoplasmic Molluscum bodies. Though it is a common clinical condition, histologically is not yet reported from this region of Mymensingh. We received a skin biopsy specimen in a pathology laboratory for histological examination. The Haematoxylin and Eosin stained sections revealed typical intracytoplasmic Molluscum bodies in keratinocytes. The lesions were in the trunk, which is a common site for Molluscum Contagiosum (MC). As the diagnosis of Molluscum contagiosum is easy by histological examination, every patient suspected to be this disease is recommended to be examined histologically to exclude other similar types of lesions. PMID- 15284702 TI - Post operative diagnosis of osteopetrosis. AB - Osteopetrosis is a rare hereditary disease that was first described by a German Albert Schonberg in 1904. At least five types of osteopetrosis have been described. Among them osteopetrosis congenita (autosomal recessive) and osteopetrosis tarda (autosomal dominant) are most common. Here we are reporting a case of osteopetrosis tarda, who is a female of fifty years of age presented with fracture femur. Preoperatively there was no suspicion of osteopetrosis. She was only suspected while introducing an IM Nail during her fracture reduction. Diagnosis of osteopetrosis was confirmed post operatively by histopathology of medullary cavity of her femur. PMID- 15284703 TI - Prenatal diagnosis of achondroplasia. AB - A 25 years old lady came to Centre for Nuclear Medicine and Ultrasound, Mymensingh for ultrasonographic evaluation of gestational condition. Transabdominal ultrasonogram showed mal development of skeletal system with abnormally shortened limbs. This seems to be a case of achnodroplasia, which was proved subsequently true after termination of pregnancy. PMID- 15284704 TI - A case of brittle bone disease. AB - Brittle bone disease--synonym, osteogenesis imperfecta is a rare genetic disorder of collagen synthesis associated with broad spectrum of musculoskeletal problem, where bones break easily. Recently we got a case of OI, whose name is Babu, 3 days old, full term bay with uneventful home delivery. The baby had multiple fractures in all the extremities with deformities with blue sclera with bilateral inguinal hernia. Other systems were found normal. On 10th day of life he was operated for inguinal hernia with satisfactory postoperative recovery and subsequently he was referred to the orthopedic department for further management. PMID- 15284705 TI - High ear piercing--a dangerous craze. AB - Ear piercing is a primitive tradition among the human being. It reflects the culture of many religions, tribes, and communities, predominately adopted by the females. We reported a sixteen years old girl with painful swelling of both pinnas for last one month following piercing the pinna. She was treated locally by general practitioner without significant improvement. On examination frank abscess were detected in both pinna. Under general anesthesia incision drainage and deep curettage was done. She was treated with ciprofloxacin 750 mg 12 hourly for 2 weeks and recovery was uneventful. After one month she developed unsightly cauliflower ear. With this report we want to sensitize our community regards the risk of transmission of needle prick diseases and deformity of pinna following ear piercing PMID- 15284706 TI - A case report on bezoar. AB - A nine years old girl was admitted into the paediatric surgery word of Dhaka Medical College Hospital (DMCH) with upper abdominal mass and discomfort, was diagnosed radiologically as a case of gastric bezoar which was further confirmed by endoscopy as trichobezoar (hair ball). The girl was undergone laparotomy and trichobezoar was removed from stomach by gastrostomy. After ten days of operation she became well and was discharged from the hospital. PMID- 15284707 TI - Cystic fibrosis in a Bangladeshi child. AB - Cystic fibrosis is one of the common lives limiting inherited diseases in Caucasians population. Recent reports suggest that the diagnosis of cystic fibrosis in this part of the world is missed or delayed due to low index of suspicion. A case of cystic fibrosis is reported here who is a Bangladeshi girl of nine-month-old who presented with the complaints of persistent cough, respiratory distress and failure to thrive. Diagnosis was made on the basis of sweat chloride estimation and mutation analysis, both of which were done, from abroad. She was put on pancreatic enzyme supplementation and nebulized bronchodilators. Cystic fibrosis though rare in Bangladesh its possibility is to be kept in mind in appropriate clinical circumstances PMID- 15284708 TI - Advances in treatment and prevention of hepatorenal syndrome. AB - Hepatorenal syndrome (HRS) remains one of the major therapeutic challenges in clinical medicine today. The pathogenesis is complex, but the final common pathway seems to be that sinusoidal portal hypertension, in the presence of severe hepatic decompensation, leads to splanchnic and systemic vasodilatation and decreased effective arterial blood volume. Renal vasoconstriction increases concomitantly, renal haemodynamics worsens, and renal failure occurs. About 15 years ago it was shown that the renal failure is potentially reversible after liver transplantation. This potential reversibility together with increased understanding of the pathogenesis has led to successful preliminary attempts to reverse HRS nonsurgically with combinations of splanchnic vasoconstrictors and colloid volume expansion, insertion of trans-jugular intrahepatic portovenous shunt radiologically, and improved forms of dialysis. The aim of this chapter is to discuss the advances in the therapy of patients with HRS PMID- 15284709 TI - Neurophysiology in clinical practice. AB - Clinical neurophysiology is concerned with the recording of generation and propagation of electrical potentials of nerve and muscle cells. It can provide important information about brain and neuromuscular disorders. Electro encephalogram (EEG), Evoked potentials (EP), Nerve conduction study (NCS) and electromyogram (EMG) are the four main techniques used in clinical neurophysiology. EEG reflects the electrical activity arising from the cerebral cortex. Evoked Potentials are measurements of classic stimulus response. VEP, SSEP and BSAEP are most useful in diagnosing Multiple Sclerosis, Plexus lesions and 8(th) cranial nerve tumor, CP angle tumor respectively. Nerve conduction studies are the measurement of velocity of nerve impulse. It gives information about the lesion of a myelinated nerve. Electromyogram gives information about the integrity of innervations, condition of neuromuscular junctions and also condition of the muscle fiber itself. PMID- 15284712 TI - Current status of aromatase inhibitors in the management of breast cancer and critique of the NCIC MA-17 trial. AB - BACKGROUND: The third-generation aromatase inhibitors reduce circulating estrogen levels in postmenopausal women and are well tolerated orally for breast cancer. Their role in the management of advanced breast cancer has already been recognized. This article reviews the evidence for their role in the adjuvant treatment of early-stage disease. METHODS: Three large multicenter trials are reviewed. The ATAC trial compared anastrozole and tamoxifen or a combination of the two, for 5 years from the point of diagnosis. The NCIC trial published the results of letrozole compared with placebo after the completion of 5 years of tamoxifen. Most recently, the Intergroup Exemestane Study reported a comparison of 5 years of tamoxifen vs 2 years of tamoxifen followed by 3 years of exemestane. RESULTS: The aromatase inhibitor arm in each of these studies was associated with improved disease-free survival and good tolerability. Because of the three different settings, cross-trial comparisons of the different aromatase inhibitors are impossible, but in each case the novel therapy appears promising. CONCLUSIONS: This review is critical of the early stopping of the NCIC study and recommends more mature follow-up in each case until distant disease-free or overall survival rates can be measured and then correlated with adverse events. The late onset of osteoporotic fractures is a concern that must be addressed before tamoxifen can be abandoned in favor of the aromatase inhibitor in each of the three clinical points: at diagnosis, at midway through a course of tamoxifen, and as an extension to the conventional 5-year period of endocrine therapy. PMID- 15284713 TI - Decreased breast cancer tumor size, stage, and mortality in Rhode Island: an example of a well-screened population. AB - BACKGROUND: Since the 1980s, Rhode Island has achieved one of the highest mammography screening rates in the nation. The objective of this study was to determine the effect of high mammography rates on breast cancer presentation and outcomes. METHODS: Using the Rhode Island Cancer Registry, the incidence of DCIS and invasive cancer, tumor size, stage, rate of BCS and mortality from breast cancer were determined from 1987 to 2001. RESULTS: Over 80% of Rhode Island women report routine mammography. From 1987 to 2001, there were 1,660 cases of DCIS and 11,301 cases of invasive breast cancer. Although the overall incidence of invasive cancer was stable, the median diameter decreased from 2 cm to 1.5 cm with a significant decrease in the incidence of stage III and IV cancers. There was an increase in BCS for women 50 to 64 years of age with stage I and II disease and for women older than 65 years with stage I disease. Disease-specific mortality decreased by 25%. CONCLUSIONS: High mammography rates in Rhode Island are associated with smaller and earlier-stage breast cancers. This largely accounts for the decreased mortality from breast cancer and the increased rate of BCS. PMID- 15284714 TI - The role of sentinel lymph node biopsy in patients with ductal carcinoma in situ or with locally advanced breast cancer receiving neoadjuvant chemotherapy. AB - BACKGROUND: A significant number of patients who are initially diagnosed with pure DCIS will harbor missed or occult invasive disease at their definitive surgery. To provide more accurate staging information and to avoid a second operation, some investigators believe that SLN mapping should be performed in DCIS patients. The role of SLN biopsy after neoadjuvant chemotherapy in patients with advanced breast cancer is controversial. METHODS: A review of the literature was performed to determine the role of SLN biopsy in patients with DCIS or advanced breast cancer receiving neoadjuvant chemotherapy. The success rate of SLN biopsy after neoadjuvant chemotherapy was investigated as well as the percentage of positive SLNs in patients with DCIS. RESULTS: Two consecutive studies revealed metastatic disease to the regional lymph nodes in up to 13% of DCIS patients. In addition, 10% of DCIS patients were upstaged to infiltrating ductal carcinoma at their definitive therapy. The ability of the SLN to predict the status of the remaining non-SLNs after neoadjuvant chemotherapy is unknown. False-negative rates range from 0% to 33%. The success rate for SLN identification for the combined series varies from 84% to 97%. CONCLUSIONS: SLN biopsy is a minimally invasive technique that can be used to evaluate the regional nodal status of DCIS patients. Performing a SLN biopsy during the initial surgical procedure may avoid a second operation in some DCIS patients who are diagnosed with invasive disease at their definitive operation. The success rate of sentinel node identification does not seem to be altered after neoadjuvant therapy. However, the ability of the SLN to predict the pathologic status of the adjacent non-SLNs remains unknown. Therefore, until further prospective randomized trials are conducted, it cannot be assumed that all the regional nodes have the same biologic response to chemotherapy as the SLN. PMID- 15284715 TI - Cancer genetics knowledge and beliefs and receipt of results in Ashkenazi Jewish individuals receiving counseling for BRCA1/2 mutations. AB - BACKGROUND: Genetic counseling for BRCA1 and BRCA2 mutations (mutations associated with increased risk of breast-ovarian cancer) endeavors to communicate information that will help individuals make informed decisions regarding genetic testing. METHODS: This repeated-measures study examined cancer genetics knowledge and beliefs before and after counseling and their relationship to receipt of results for BRCA1/2 mutations in 120 highly educated Ashkenazi Jewish individuals. RESULTS: A repeated-measures analysis examined change in knowledge and beliefs regarding personal behavior, mechanisms of cancer inheritance, meaning of a positive result, practitioner knowledge, frequency of inherited cancer, and meaning of a negative result from pre- to post-counseling with the between subjects variables of education (with/without graduate training) and personal history of breast or ovarian cancer (yes/no), and risk of having a mutation entered as a covariate. Mechanisms of cancer inheritance, meaning of a positive result, and practitioner knowledge increased from pre- to post counseling. Those with graduate training had higher ratings of mechanisms of cancer inheritance ratings and lower ratings of frequency of inherited cancers than those without. Mann-Whitney U tests found those testing had higher ratings in mechanisms of cancer inheritance, specifically in the association of multiple primary cancers with hereditary cancer, than those not testing. CONCLUSIONS: Genetic counseling is helpful in improving overall knowledge of cancer genetics even for highly educated individuals. Particular areas of knowledge improvement should be explored in relation to receipt of results, especially to further elucidate the relationship of knowledge of the association of multiple primary cancers with hereditary cancer to receipt of test results. PMID- 15284716 TI - Enhancing the readability of materials describing genetic risk for breast cancer. AB - BACKGROUND: The number of individuals contemplating genetic testing is increasing, but the current materials and overall subject matter remain complex and not easily understood by many. The goal of this project was to evaluate efforts to revise and increase the readability of an existing information packet describing genetic risk for breast cancer. METHODS: Evaluation was conducted in two stages through two related studies. In Study 1, a focus group of multiethnic breast cancer survivors was assembled to obtain feedback on images included in the revised breast cancer genetics information packet. In Study 2, African American adult students in a literacy program evaluated the revised images (based on the feedback of the focus group in Study 1) and text of the information packet and provided ratings on readability, format, and appearance. RESULTS: Responses from Study 1 participants suggested that some of the images created for the packet needed to be clearer in the concepts they were intended to convey. In Study 2, ratings of adult learners suggested difficulty with word comprehension in spite of the inclusion of definitions and a glossary. The reading level achieved was markedly lower than the college reading level required by the original information packet and other patient-directed cancer genetics materials. CONCLUSIONS: Although efforts to clarify written materials in order to better serve patients with low literacy received generally favorable responses, continued efforts to create more user-friendly patient education materials are warranted. PMID- 15284717 TI - Potential economic effects of volume-outcome relationships in the treatment of three common cancers. PMID- 15284718 TI - Bilateral breast masses. PMID- 15284719 TI - Burnout and balance: how to go the distance in the 21st century. PMID- 15284720 TI - Use of microbial cultures and antibiotics in the prevention of infection associated preterm birth. AB - OBJECTIVE: The purpose of this study was to summarize recent evidence regarding infection-associated preterm birth and to make appropriate recommendations. Antepartum treatment of lower genital tract infection or bacterial colonization has been found to reduce the incidence of preterm birth in the case of asymptomatic bacteriuria and bacterial vaginosis in selected patients but has been proved to be ineffective for vaginal colonization with organisms such as Ureaplasma urealyticum and group B streptococcus. STUDY DESIGN: This is a clinical opinion based on a review of recent data related to 1) the association between lower genital tract infection and preterm birth and 2) antibiotic trials to prevent preterm birth. RESULTS: Antepartum treatment of lower genital tract infection or bacterial colonization has been found to reduce the incidence of preterm birth in the case of asymptomatic bacteriuria and bacterial vaginosis in selected patients, but has been proven to be ineffective for vaginal colonization with organisms such as Ureaplasma urealyticum and group B streptococcus. Large well-designed trials have shown that the routine administration of antibiotics to women with preterm labor and intact membranes is not beneficial; however, antibiotic regimens including macrolides are recommended for preterm premature rupture of the membranes. CONCLUSION: Large well-designed trials have shown that the routine administration of antibiotics to women with preterm labor and intact membranes is not beneficial; however, antibiotic regimens that include macrolides are recommended for preterm premature rupture of the membranes. PMID- 15284721 TI - Clinical feature. Bacterial vaginosis and inflammatory vaginitis. PMID- 15284722 TI - A polymorphism in the promoter region of TNF and bacterial vaginosis: preliminary evidence of gene-environment interaction in the etiology of spontaneous preterm birth. AB - OBJECTIVE: The rarer of 2 alleles of a polymorphism in the promoter of the tumor necrosis factor alpha gene (TNF) has been associated with spontaneous preterm birth following preterm premature rupture of the fetal membranes in some populations. The aim of this study was to assess if the presence of symptomatic bacterial vaginosis amplifies the risk of spontaneous preterm birth in those with a "susceptible" TNF genotype. STUDY DESIGN: A case-control study was performed at our institution. Cases (n=125) were defined as women who delivered before 37 weeks as a result of ruptured membranes or preterm labor, while control subjects (n=250) were defined as women who delivered after 37 weeks. DNA was collected from maternal blood and analyzed for the TNF genotype. Information on symptomatic bacterial vaginosis and other risk factors for preterm birth was obtained by review of the antenatal record. Multiple logistic regression was also used to test the interaction between bacterial vaginosis, the TNF genotype, and preterm birth. RESULTS: Maternal carriers of the rarer allele (TNF-2) were at a significantly increased risk of spontaneous preterm birth [odds ratio (OR) 2.7, 95% CI 1.7-4.5]. The association between TNF-2 and preterm birth was modified by the presence of bacterial vaginosis, such that those with a "susceptible" genotype and bacterial vaginosis had increased odds of preterm birth compared with those who did not (OR 6.1, 95% CI 1.9-21.0). CONCLUSION: This study provides preliminary evidence that an interaction between genetic susceptibilities (ie, TNF-2 carriers) and environmental factors (ie, bacterial vaginosis) is associated with an increased risk of spontaneous preterm birth. PMID- 15284723 TI - Bacterial vaginosis, the inflammatory response and the risk of preterm birth: a role for genetic epidemiology in the prevention of preterm birth. PMID- 15284724 TI - Magnesium sulfate prophylaxis in preeclampsia: Lessons learned from recent trials. AB - In the US, the routine use of magnesium sulfate for seizure prophylaxis in women with preeclampsia is an ingrained obstetric practice. During the past decade, several observational studies and randomized trials have described the use of various regimens of magnesium sulfate to prevent or reduce the rate of seizures and complications in women with preeclampsia. There are only 2 double-blind, placebo-controlled trials evaluating the use of magnesium sulfate in mild preeclampsia. There were no instances of eclampsia among 181 women assigned to placebo, and there were no differences in the percentage of women who progressed to severe preeclampsia (12.5% in magnesium group vs 13.8% in the placebo group, relative risk [RR] 0.90; 95% CI 0.52-1.54). However, the number of women enrolled in these trials is too limited to draw any valid conclusions. There are 4 randomized controlled trials that compare the use of no magnesium sulfate, or a placebo vs magnesium sulfate, to prevent convulsions in patients with severe preeclampsia. The rate of eclampsia was 0.6% among 6343 patients assigned to magnesium sulfate vs 2.0 % among 6330 patients assigned to a placebo or control (RR 0.39; 95% CI 0.28-0.55). However, the reduction in the rate of eclampsia was not associated with a significant benefit in either maternal or perinatal outcome. In addition, there was a higher rate of maternal respiratory depression among those assigned magnesium sulfate (RR 2.06; 95% CI 1.33-3.18). The evidence to date confirms the efficacy of magnesium sulfate in reduction of seizures in women with eclampsia and severe preeclampsia; however, this benefit does not affect overall maternal and perinatal mortality and morbidities. The evidence regarding the benefit-to-risk ratio of magnesium sulfate prophylaxis in mild preeclampsia remains uncertain, and does not justify its routine use for that purpose. PMID- 15284726 TI - The human gynome. AB - This year marks the 50th anniversary of the identification of the 3-dimensional structure of the DNA double helix and the completion of the US Human Genome Project. Now that we have completed the human genome sequence, what have we learned? How will this information benefit humankind? And, what are the implications for our patients in obstetrics and gynecology? Perhaps the biggest surprise is that there are only approximately 30,000 human genes, far fewer than earlier estimated. I propose the term "gynome" to describe that part of the human genome that is unique to women. We have learned that manifestations of diseases and therapeutic response can be gender specific. A major challenge is to define the interplay of the genetic variations of women with variations in their environment and lifestyle. Ultimately, this should lead to improved diagnosis of disease, earlier detection of genetic predispositions to disease, the design of more effective drugs, and gene therapy. PMID- 15284727 TI - Review of the professional medical liability insurance crisis: lessons from Missouri. AB - OBJECTIVE: This study was undertaken to document and generalize the professional liability insurance (PLI) crisis. STUDY DESIGN: Data analysis from physician surveys, Missouri Department of Insurance, and court records. RESULTS: In 2001 2002, many insurers stopped writing new and existing PLI. A survey found 1 in 7 physicians had their PLI terminated and/or application for new insurance denied. Average premiums increased 22% in 2001 and 60% in 2002. Accordingly, 50% of surveyed obstetricians took salary cuts, 18% secured loans, 9% liquidated assets, and 55% significantly limited their clinical services. An adverse court ruling caused insurers to double reserves. Incomplete data led the Missouri Department of Insurance to erroneously conclude a decrease in claim frequency and severity. In contrast, courthouse records and missing data sources revealed increased malpractice filings. PLI premiums continue to rise. CONCLUSION: Many life-saving specialists are being forced out of business. Specialists are less willing to care for emergency and indigent patients for fear of liability exposure. Legislative enactments leading to meaningful tort reform, public support, and judicial restraint must occur to save health care. PMID- 15284729 TI - Evidence supporting a role for blockade of the vascular endothelial growth factor system in the pathophysiology of preeclampsia. Young Investigator Award. AB - OBJECTIVE: Soluble vascular endothelial growth factor receptor 1 (sVEGFR-1), which antagonizes VEGF functions, has been implicated in the pathophysiology of preeclampsia. The purpose of this study was to determine whether preeclampsia is associated with a change in the plasma concentration of sVEGFR-1, and, if so, whether such a change is correlated with the severity of the disease. METHODS: A cross-sectional study was conducted to determine the concentrations of sVEGFR-1 in plasma obtained from normal pregnant women (n=61) and patients with preeclampsia (n=61). Plasma concentrations of sVEGFR-1 were determined by enzyme linked immunoassay. RESULTS: Preeclampsia had a higher median plasma concentration of sVEGFR-1 than normal pregnancy (P <.001). The median plasma concentration of sVEGFR-1 was higher in early-onset (< or =34 weeks) than late onset (>34 weeks) preeclampsia (P=.005), and higher in severe than in mild preeclampsia (P=.002). In normal pregnancy, there was a correlation between plasma concentration of sVEGFR-1 and gestational age (r=0.5; P <.001). In contrast, there was a negative correlation between plasma concentration of sVEGFR 1 and gestational age at the onset of preeclampsia (r=-0.5; P <.001). CONCLUSION: Preeclampsia is associated with an increased plasma sVEGFR-1 concentration. The elevation of sVEGFR-1 concentration is correlated with the severity of the disease. These observations suggest the participation of VEGF and its soluble receptor in the pathophysiology of preeclampsia. PMID- 15284732 TI - Prolonged in utero meconium exposure impairs spatial learning in the adult rat. Central Prize Award. AB - OBJECTIVE: The purpose of this study was to examine the effects of prolonged in utero meconium exposure on adult learning and memory, as measured by the Morris water maze. STUDY DESIGN: Timed pregnant Long-Evans rats were studied. On gestational day 20 (term, 21 days of gestation), laparotomy was performed, and each maternal animal received an injection of clear amniotic fluid or meconium stained amniotic fluid into each gestational sac. The laparotomy incision was closed, and the animals received postoperative monitoring through delivery. On postnatal days 145 to 148, the offspring underwent Morris water maze testing. The mean (+/-SEM) for the latency time was reported for each day's trial and compared between groups. RESULTS: There were significant differences between meconium stained amniotic fluid group and clear amniotic fluid group in the mean time to platform on day 1 (82.7 +/- 1.8 seconds vs 75.9 +/- 3.0 seconds; P=.04), day 2 (60.5 +/- 3.5 seconds vs 47. 8 +/- 4.6 seconds; P=.03), and day 3 (56.5 +/- 4.5 seconds vs 34.7 +/- 4.4 seconds; P=.001). However, there were no differences on days 4 and 5. There were also no differences between recall and response learning trials that were done after a 12-day retention period. CONCLUSION: In the absence of hypoxia or infection, prolonged in utero meconium exposure is associated with a delay of spatial learning in the adult rat. PMID- 15284734 TI - Racial disparity in membrane response to infectious stimuli: a possible explanation for observed differences in the incidence of prematurity. Community Award Paper. AB - OBJECTIVE: This study compares the immune responsiveness of amniochorionic membranes (AC) derived from African American (AA) and white (C) women to an infectious stimulus ex vivo. STUDY DESIGN: AC derived from AA and C women were placed in an organ explant culture for 48 hours and then stimulated with endotoxin. Enzyme-linked immunosorbent assay measured the concentration of matrix metalloproteinase 9 (MMP9), tumor necrosis factor-alpha (TNF-alpha), and soluble TNF receptors (sTNFR1 and sTNFR2) in culture media from stimulated and unstimulated AC. RESULTS: The C group produced 8-fold more TNF-alpha after stimulation than did the AA group. Both soluble receptor (R1 and R2) production increased in the C group and decreased in the AA group after stimulation. Although the C group-derived membranes produced more MMP9 at rest, a 6-fold increase in MMP9 concentration was seen in the AA group-derived membranes after stimulation. No change in MMP9 concentration was seen after stimulation of the C group-derived membranes. CONCLUSION: Although the C group produced more TNF, they also produce higher sTNFRs, which may serve a protective role. The increased MMP9 release by the AA group may be suggestive of the greater risk of premature rupture of membranes in the AA group. PMID- 15284736 TI - A randomized clinical trial of the intrapartum assessment of amniotic fluid volume: amniotic fluid index versus the single deepest pocket technique. AB - OBJECTIVE: The purpose of this study was to determine whether an intrapartum assessment of amniotic fluid identifies a pregnancy that is at risk for an adverse outcome. STUDY DESIGN: Parturients who were admitted for delivery were assigned randomly to have the amniotic fluid assessed either by amniotic fluid index or by the presence of a 2 x 1 pocket. RESULTS: The amniotic fluid index was obtained in 499 pregnancies, and the 2 x 1 technique was performed in 501. Oligohydramnios was diagnosed in 25% of amniotic fluid index pregnancies versus 8% with the use of the 2 x 1 pocket technique (P <.001). Both techniques failed to identify patients who underwent an amnioinfusion for fetal distress (P=.864) or who experienced variable (P=.208) or late decelerations (P=.210) that influenced delivery, fetal distress in labor (P=.220), caesarean delivery for fetal distress (P=.133), and admission to neonatal intensive care unit (P=.686). CONCLUSION: Neither the amniotic fluid index nor the 2 x 1 pocket technique that was undertaken as a fetal admission test identifies a pregnancy that is at risk for an adverse outcome. PMID- 15284738 TI - Automated detection of rare fetal cells in maternal blood: eliminating the false positive XY signals in XX pregnancies. AB - OBJECTIVE: The purpose of this study was to develop a new method to help differentiate XX from XY signals in maternal blood from women carrying XY fetuses. STUDY DESIGN: We have developed a system to scan automatically for cells that bear X and Y fluorescence in situ hybridization signals. These XY target cells are identified by scans at low (x20) magnification, and all identified targets are revisited and verified at high (x100) magnification. The viewer software component of the system displays x20 images of all cells and intracellular fluorescence in situ hybridization signals that are present in each of the 4000 optical fields per slide, along with x100 images of automatically detected target cells. RESULTS: We initially examined 36,000 fields from 18 slides in 12 pregnancies (6 male and 6 female) using our system that is based on fluorescence in situ hybridization with a single probe for the X-chromosome and a single probe for the Y-chromosome and found XY nuclei in all samples, regardless of fetal gender. In the second phase of the study, a refinement of the approach that incorporated 2 independent probes for the Y-chromosome resulted in a false positive rate for detection of XY nuclei in XX cases <0.00005%. CONCLUSION: Our data suggest that this system may allow for excellent "signal to noise" separation, which is required absolutely for fetal cell methods to differentiate aneuploid from normal pregnancies. Quantitation of fetal cells in the maternal circulation and standardization of processes that have been developed for their enrichment are crucial to moving fetal cell assessment from esoteric basic science to applied new technology. PMID- 15284741 TI - Fetal fibronectin and bacterial vaginosis are associated with preterm birth in women who are symptomatic for preterm labor. AB - OBJECTIVE: The purpose of this study was to codify the relationship between bacterial vaginosis/fetal fibronectin and preterm labor/birth. STUDY DESIGN: In this prospective study, 185 women who were symptomatic for preterm labor were assessed for bacterial vaginosis and fetal fibronectin. RESULTS: These women comprised 4 groups: group A (n=23 women; +bacterial vaginosis/+fetal fibronectin); group B (n=31 women; -bacterial vaginosis/+fetal fibronectin); group C (n=47 women; +bacterial vaginosis/-fetal fibronectin); and group D (n=84 women; -bacterial vaginosis/-fetal fibronectin). The time interval from gestational age at testing until delivery was significantly shorter for groups A and B versus groups C and D (P < or =.05 and P <.001, respectively). Similarly, delivery at <32 weeks of gestation was increased in group B (26%) compared with groups A (9%), C (2%), and D (5%; P <.009; odds ratio, 165.90; 95% CI, 30.02, 916.08). CONCLUSION: Women who are symptomatic for preterm labor should be considered for fetal fibronectin and bacterial vaginosis testing. PMID- 15284743 TI - Maternal and perinatal outcomes during expectant management of 239 severe preeclamptic women between 24 and 33 weeks' gestation. AB - OBJECTIVE: This study was undertaken to determine maternal and perinatal outcomes after expectant management of severe preeclampsia between 24 and 33 weeks' gestation. STUDY DESIGN: A prospective observational study of 239 women with severe preeclamptic and undelivered after antenatal steroid prophylaxis was performed. Pregnancy prolongation and maternal and perinatal morbidities were analyzed according to the gestational age at time of expectant management: 24 to 28, 29 to 31, and 32 to 33 weeks. Statistical analysis was performed by Student t test and chi(2) test. RESULTS: The days of pregnancy prolongation were significantly higher among those managed at less than 29 weeks (6) compared with the other groups (4). There were 13 perinatal deaths: 12 in those managed at less than 29 weeks and 1 in those managed at 29 to 31 weeks. Neonatal morbidities were significantly higher among those managed at less than 29 weeks compared with the other groups. There were no instances of maternal death or eclampsia. Maternal morbidities were similar among the groups. CONCLUSION: Expectant management of severe preeclampsia at 24 to 33 weeks in a tertiary care center is associated with good perinatal outcome with a minimal risk for the mother. PMID- 15284746 TI - Prediction of labor in term and preterm pregnancies using non-invasive magnetomyographic recordings of uterine contractions. AB - OBJECTIVE: The purpose of this study was to characterize the noninvasive magnetic field recordings of the uterine electrophysiological activity in patients reporting onset of uterine contractions. STUDY DESIGN: Transabdominal magnetomyographic (MMG) recordings were performed with the use of the SARA system's 151 primary magnetic sensors (CTF Systems Inc, Coquitlam, British Columbia, Canada). Eleven term and 4 preterm patients participated in the study. On all patients, cervical dilation and outcomes were recorded. RESULTS: Of 8 patients having a peak MMG activity exceeding 8 pT (pico Tesla), all but 1 delivered within 48 hours of our recordings. Of the 7 patients with peak activity below 8 pT, 5 failed to deliver within 48 hours of the recordings. This observation reflects an increase in the electrophysiologic activity of the myometrium as labor progresses. CONCLUSION: MMG evaluation provides a new noninvasive method for the prediction of labor. Further studies are in progress to determine the temporal relationship of MMG changes with the onset of labor. PMID- 15284748 TI - Is abnormal labor associated with shoulder dystocia in nulliparous women? AB - OBJECTIVE: This study was undertaken to examine the relationship between labor abnormalities and shoulder dystocia in nulliparous women. STUDY DESIGN: Nulliparous women whose delivery was complicated by shoulder dystocia were studied and compared with a control group selected based on the best possible match for race, labor type (spontaneous or induced), and birth weight. The duration of first and second stage of labor, as well as the rates of labor progress, were calculated and compared between groups. RESULTS: During this 4 year study period, there were 8010 nulliparous singleton deliveries of which 65 (0.8%) were complicated by shoulder dystocia. Compared with controls, there was no difference in the rate of cervical dilation in the active phase of the first stage of labor. In the shoulder dystocia group, more patients had a second stage of labor greater than 2 hours (22% vs 3%; P <.05) and had operative vaginal deliveries (26% vs 1.5%; P <.001). In shoulder dystocia cases with birth weight greater than 4000 g, 33% had a second stage of labor greater than 2 hours. CONCLUSION: In our population, the combination of fetal macrosomia, second stage of labor longer than 2 hours and the use of operative vaginal delivery were associated with shoulder dystocia in nulliparous women. PMID- 15284751 TI - Is there a relationship to dye determined or ultrasound estimated amniotic fluid volume adjusted percentiles and fetal weight adjusted percentiles? AB - OBJECTIVE: This study was undertaken to determine whether there is a correlation between neonatal birth weight and an estimated or dye-determined amniotic fluid volume. STUDY DESIGN: A prospective observational study of women undergoing an amniocentesis and subsequent delivery within 72 hours. Amniotic fluid volume was estimated using the amniotic fluid index (AFI) and single deepest pocket technique and calculated by the dye-dilution technique. RESULTS: One hundred seventy-five women participated in the study. Linear regression modelling showed no significant relationship between birth weight and amniotic fluid volume adjusted for gestational age (P=.062). Similarly, there was no evidence of an association between birth weight and a dye determined amniotic fluid volume (P=.180), fixed cutoffs for the AFI (P=.224), percentiles for the AFI (P=.112), or fixed cutoffs for the single deepest pockets (P=.867). CONCLUSION: Neonatal birth weight is not correlated with a dye-determined or ultrasound estimated amniotic fluid volume. PMID- 15284753 TI - Does advanced ultrasound equipment improve the adequacy of ultrasound visualization of fetal cardiac structures in the obese gravid woman? AB - OBJECTIVE: This study was undertaken to determine the effect of advanced ultrasound equipment on the ability to visualize fetal cardiac structures in obese gravid women. STUDY DESIGN: Singleton pregnancies undergoing initial ultrasound examination between 14 weeks and 23 weeks 6 days were included. Patients were classified by body mass index (BMI) (nonobese [BMI < 30 kg/m(2)] and obese [BMI > or =30 kg/m(2)]). The rate of suboptimal ultrasound visualization (SUV) of the fetal heart (cardiac axis, cardiac position, 4 chamber, and outflow tracts views) was compared between patients examined by standard (HDI 3000) or advanced ultrasound equipment (HDI 5000) (ATL, Philips Medical Systems, Bothell, Wash). RESULTS: Over a 5-year period, 7029 singleton gestations met inclusion criteria; 2498 (35.5%) were clinically obese. There was no difference in gestational age, rate of low amniotic fluid volume, anterior placenta, or vertex fetal presentation between the groups. When the advanced ultrasound equipment was used, SUV of the fetal heart was lower in the nonobese population (20.8% vs 16.4%; P <.001), but not in the obese gravid women (38.1% vs 35.5%; P=.27). However, obese patients who were examined by advanced ultrasound equipment after 18 weeks had less SUV of the outflow tracts (28.5% vs 23.1%, P=.04) but not of the 4-chamber view. CONCLUSION: Despite advanced ultrasound equipment, maternal obesity significantly limits visualization of the fetal heart. However, the advanced ultrasound equipment may somewhat benefit obese gravid women examined after 18 weeks' gestation PMID- 15284756 TI - Expression and regulation of luteinizing hormone/human chorionic gonadotropin receptors in ovarian cancer and its correlation to human chorionic gonadotropin doxorubicin sensitivity. AB - OBJECTIVE: Ovarian cancer cell lines and tissues express gonadotropin receptors. Conjugation of cytostatics to ligands of these receptors may increase the specificity of cytotoxic drugs. STUDY DESIGN: Toxicity of doxorubicin-human chorionic gonadotropin conjugates was determined in 4 ovarian cancer cell lines. Expression and regulation of luteinizing hormone/human chorionic gonadotropin receptors were analyzed before and after treatment with human chorionic gonadotropin, epidermal growth factor, and 8-bromo-cyclic adenosine monophosphate with a nested reverse transcriptase-polymerase chain reaction approach. RESULTS: Toxicity of human chorionic gonadotropin-doxorubicin conjugates was increased compared with unconjugated doxorubicin in OVCAR-3 cells. However, drug conjugates failed to demonstrate increased toxicity in other cell lines, especially after preincubation with human chorionic gonadotropin. All cell lines expressed luteinizing hormone/human chorionic gonadotropin receptors. Receptor expression in OVCAR-3 cells was not effected by human chorionic gonadotropin, endothelial growth factor, or 8-bromo-cyclic adenosine monophosphate treatment. In other cell lines, receptor expression was down-regulated by these agents. CONCLUSION: Cytotoxic activity of doxorubicin was increased specifically by conjugation to human chorionic gonadotropin. However, the regulation of luteinizing hormone/human chorionic gonadotropin receptor expression and other compounds may reduce the drug-uptake of the conjugates. PMID- 15284758 TI - International Classification of Diseases-9th revision coding for preeclampsia: how accurate is it? AB - OBJECTIVE: The purpose of this study was to evaluate the accuracy of the International Classification of Diseases-9th revision codes for preeclampsia and eclampsia. STUDY DESIGN: The University of Illinois Medical Center at Chicago discharge database was used to identify 135 women from 1999 through 2001 whose disease was coded as having preeclampsia or eclampsia. With American College of Obstetrics and Gynecology criteria as the gold standard, the diagnosis that was determined through chart review was compared with the International Classification of Diseases-9th revision code that was present in the discharge database. Patients were classified as true cases if the International Classification of Diseases-9th revision code matched the American College of Obstetricians and Gynecologists diagnosis; the positive predictive value of the code was then calculated. RESULTS: The overall positive predictive value for the complete sample was only 54%, but the positive predictive value for severe preeclampsia was 84.8%, which was high compared with mild preeclampsia (45.3%) and eclampsia (41.7%). Diagnostic (clinician) error was the most common reason for miscoding error. CONCLUSION: The findings suggest that International Classification of Diseases-9th revision codes for preeclampsia/eclampsia vary greatly in their accuracy of diagnosis. Therefore, a review of medical records is required when data are being gathered on the incidence of preeclampsia and eclampsia. PMID- 15284761 TI - Rationing of medical care: Rules of rescue, cost-effectiveness, and the Oregon plan. AB - Doctors who deal with individual patients fail to avoid interventions with minimal expected benefits. This is one reason that the United States spends more on health care services than any of 28 other industrialized nations. Yet, our money has not bought us health; our infant mortality rate ranks 23rd, and our overall life expectancy rate ranks 20th among the 29 nations. Ours is the only nation without a national health system. Our job-based health insurance system has allowed the number of uninsured persons to reach 44 million, which is 18% of the nonelderly population. This article examines the role of such ethical concepts as beneficence, utilitarianism, and justice in the allocation of health care resources. It also examines the innovative Oregon Health Plan and its use of cost-effectiveness analysis for health care allocation that is based on league tables. PMID- 15284762 TI - A follow-up evaluation of sexual misconduct complaints: the Oregon Board of Medical Examiners, 1998 through 2002. AB - OBJECTIVE: The current study presents an analysis of sexual misconduct allegations that were closed from 1998 through 2002 and is compared with allegations from 1991 through 1995 (study 1). One hundred complaints were closed in study 1, which involved 80 licensees; 23.8% of those complaints resulted in reportable board actions. STUDY DESIGN: Retrospective analyses of 47 allegations that were closed and that involved 46 practitioners were evaluated statistically and compared with the previous study. RESULTS: Sexual misconduct was the allegation in 3.1% of all closed complaints, compared with 5.9% in study 1. Of the allegations, 36.2% of the sexual misconduct allegations were for sexual impropriety with no reportable disciplinary outcomes, and 63.8% of the complaints were for sexual violation that resulted in 25 reportable disciplinary actions. Family medicine, psychiatry, and obstetrics/gynecology again reported the highest proportion of total complaints, but psychiatry and obstetrics/gynecology improved both in total complaints and disciplinary actions. Multiple complaints improved significantly in study 2. CONCLUSION: Physician and patient awareness and board actions reduced total complaints of sexual misconduct. Family medicine was an exception, with 12 reportable board actions compared with 4 in study 1. Reportable disciplinary actions involved revocations, suspensions, and surrender of license; the disciplinary actions most often involved probation, education, counseling and/or psychiatric therapy, and practice limitation. Education, the identification of high-risk practitioners, and the appropriate use of deterrence continue to be areas of recommended focus. PMID- 15284763 TI - The insulin resistant subphenotype of polycystic ovary syndrome: clinical parameters and pathogenesis. AB - OBJECTIVE: This study was undertaken to compare clinical and biochemical characteristics of the insulin resistant (IR) and non-IR subphenotypes of polycystic ovary syndrome (PCOS). STUDY DESIGN: Infertile PCOS women were classified as IR (n=32) or non-IR (n=46) on the basis of fasting glucose and insulin levels. The incidence of acanthosis nigricans (AN), hirsutism, and ovulation in response to clomiphene citrate (CC) was compared between the 2 groups, along with serum levels of gonadotropins, and sex steroids. Blood samples from 28 PCOS patients and 8 controls were analyzed by enzymatic immunoassay for autophosphorylated insulin receptor (APIR) and total insulin receptor (TIR) content. RESULTS: Insulin resistance was associated with obesity (odds ratio [OR]=3.5, P <.05), AN (OR=6.0, P <.05), hirsutism (OR=3.1, P <.05), and resistance to CC (OR=5.0, P <.05). Mean levels of LH, LH/FSH ratios, and testosterone were lower in women with IR (11.5 +/- 6.8 mIU/mL, 2.0 +/- 1.0, and 56.6 +/- 29.0 ng/dL, respectively) compared with women without IR (15.0 +/- 13.4 mIU/mL, 2.4 +/- 1.5, and 72.5 +/- 29.8 ng/dL, respectively) (P <.05). Mean APIR/TIR ratios in IR women were lower than in non-IR women (P <.05 at 100 nmol/L of insulin) and controls (P <.01 at 1, 10 and 100 nmol/L insulin). CONCLUSION: Patients with IR are more likely to be obese and have AN, hirsutism, resistance to CC, and lower LH, LH/FSH ratios, and testosterone levels. Furthermore, IR patients appear to have defective autophosphorylation of the insulin receptor, a key element in insulin action, and a possible mechanism for IR in PCOS. PMID- 15284764 TI - Trauma during pregnancy: an analysis of maternal and fetal outcomes in a large population. AB - OBJECTIVE: This study was undertaken to determine the occurrence rates, outcomes, risk factors, and timing of obstetric delivery for trauma sustained during pregnancy. STUDY DESIGN: This is a retrospective cohort study of women hospitalized for trauma in California (1991-1999). International Classification of Disease, ninth revision, Clinical Modification codes, and external causation codes for injury were identified. Maternal and fetal/neonatal outcomes were analyzed for women delivering at the trauma hospitalization (group 1), and women sustaining trauma prenatally (group 2), compared with nontrauma controls. Injury severity scores and injury types were used to stratify risk in relation to outcome. Statistical comparisons are expressed as odds ratios (ORs) with 95% CIs. RESULTS: A total of 10,316 deliveries fulfilling study criteria were identified in 4,833,286 total deliveries. Fractures, dislocations, sprains, and strains were the most common type of injury. Group 1 was associated with the worst outcomes: maternal death OR 69 (95% CI 42-115), fetal death OR 4.7 (95% CI 3.4-6.4), uterine rupture OR 43 (95% CI 19-97), and placental abruption OR 9.2 (95% CI 7.8 11). Group 2 also resulted in increased risks at delivery: placental abruption OR 1.6 (95% CI 1.3-1.9), preterm labor OR 2.7 (95% CI 2.5-2.9), maternal death OR 4.4 (95% CI 1.4-14). As injury severity scores increased, outcomes worsened, yet were statistically nonpredictive. The type of injury most commonly leading to maternal death was internal injury. The risk of fetal, neonatal, and infant death was strongly influenced by gestational age at the time of delivery. CONCLUSION: Women delivering at the trauma hospitalization (group 1) had the worst outcomes, regardless of the severity of the injury. Group 2 women (prenatal injury) had an increased risk of adverse outcomes at delivery, and therefore should be monitored closely during the subsequent course of the pregnancy. This study highlights the need to optimize education in trauma prevention during pregnancy. PMID- 15284765 TI - The diagnosis and reproductive outcome after surgical treatment of the complete septate uterus, duplicated cervix and vaginal septum. AB - OBJECTIVE: This study was undertaken to evaluate the diagnostic management and the reproductive outcome after surgical repair of a rare reproductive malformation. STUDY DESIGN: Sixteen women with a complete septate uterus, double cervix, and a longitudinal vaginal septum were referred for evaluation. Presenting complaints were chiefly pregnancy loss in parous women (n=9) and dyspareunia in nulligravid women (n=7). The combination of hysterosalpingography, ultrasonography, and/or magnetic resonance imaging was used to correctly identify the anomaly in 15 of the 16 cases. Both hysteroscopic (n=11) and transabdominal (n=5) surgical techniques were used to repair the uterine septum. RESULTS: In no case was the correct diagnosis made before referral; the uterus didelphys was the most common misdiagnosis. The preoperative pregnancy loss was 81%. Postoperatively, 12 women conceived for a total of 17 pregnancies; there were 14 term live births or ongoing pregnancies in the third trimester (82%), with a first trimester spontaneous abortion rate of 18%. In 9 women who conceived after hysteroscopic surgery, term live births occurred in 9 of 12 (75%) conceptions. A modified Tompkins metroplasty was performed in 5 women with subsequent term live births or ongoing third trimester pregnancies in 5 of 5 (100%) patients. CONCLUSION: The identification of a duplicated cervix and a vaginal septum is consistent with several uterine malformations, which leads to frequent misdiagnosis and errors in management. Significant pregnancy wastage, obstetric complications, and dyspareunia are common, and surgical treatment is therefore advisable. Making the best choice between hysteroscopic or transabdominal metroplasty depends on the anatomic features of the cervix and the uterine cavity, but optimal patient management requires familiarity with both techniques. PMID- 15284768 TI - The magnetic resonance imaging-based fetal-pelvic index: a pilot study in the community hospital. AB - OBJECTIVE: This study was undertaken to assess feasibility of magnetic resonance imaging (MRI) pelvimetry in conjunction with fetal ultrasonography as a technique in evaluating patients with previous cesarean sections for cephalopelvic disproportion (CPD). STUDY DESIGN: Pregnant patients with one previous cesarean section for CPD who planned a trial of labor after cesarean (TOLAC) were recruited to undergo MRI pelvimetry and fetal ultrasonography at 37 to 38 weeks. Entry criteria included no previous successful vaginal deliveries and no contraindications for vaginal delivery in the ongoing pregnancy. A fetal-pelvic index was calculated for each patient but not disclosed to patients or their physicians. The pregnancies were managed routinely. Analysis after delivery was used to ascertain whether this index would have predicted clinical outcome. RESULTS: There were no difficulties in performing the MRI or ultrasound. Sixteen patients completed their pregnancies. Three patients did not labor. The fetal pelvic index was plotted on a scattergram and compared with the outcome. Three discriminatory zones were identified. Five of 6 patients in the most favorable zone delivered successfully. Two patients in the most unfavorable zone had failed vaginal birth after cesarean section (VBAC) attempts. In the 5 patients in the middle intermediate zone, TOLAC success appeared to depend on fetal presentation and gestational age. CONCLUSION: The use of comparative MRI pelvimetry and fetal ultrasonography is feasible in a community hospital. In this pilot study, it appeared to have potential in enhancing the management of VBAC candidates. This technique may allow sorting of patients before labor into zones that would favor or preclude VBAC attempts. PMID- 15284771 TI - A comparison of orally administered misoprostol to intravenous oxytocin for labor induction in women with favorable cervical examinations. AB - OBJECTIVE: The purpose of this study was to compare orally administered misoprostol with intravenous oxytocin infusion for labor induction in women with favorable cervical examinations (defined as a Bishop score of 6 or more). STUDY DESIGN: One hundred ninety-eight women with indications for labor induction and favorable cervical examinations were assigned randomly to receive oral misoprostol or oxytocin induction. Misoprostol, 100 mg, was administered every 4 hours up to 6 doses, or intravenous oxytocin was administered by standardized protocol. RESULTS: One hundred ten (55.6%) women received misoprostol; 88 (44.4%) received intravenous oxytocin. There was no statistically significant difference in the average interval from start of induction to vaginal delivery, being longer in the misoprostol group (789.4 +/- 510.2 minutes) than in the oxytocin group (654.0 +/- 338.2 minutes, P=.19, log-transformed data). Two women had tachysystole develop in each treatment group. More women in the misoprostol group experienced hyperstimulation (7/110, 6.4%) than in the oxytocin group (0/88, P=.02, Fisher exact test). Nine (8.1%) misoprostol-treated women and 8 (9.1%) oxytocin-treated women underwent cesarean deliveries (P=.82). There was a presumed uterine rupture in a misoprostol-treated multipara women. There were no statistically significant differences in neonatal outcomes between the groups. CONCLUSION: Oral misoprostol offers no benefit over intravenous oxytocin for labor induction in women with favorable cervical examinations. It is associated with a higher likelihood of uterine hyperstimulation and may increase the risk of uterine rupture. PMID- 15284774 TI - Patient satisfaction and disease specific quality of life after uterine artery embolization. AB - OBJECTIVES: This study was undertaken to evaluate changes in fibroid specific symptom severity and health-related quality of life (HRQOL) after uterine artery embolization (UAE) and to consider the impact of these changes on satisfaction with the procedure. STUDY DESIGN: A validated, fibroid specific, symptom, and HRQOL questionnaire was mailed to 80 women who had undergone UAE from 1998 through 2002. Pre- and postprocedure symptom severity and HRQOL scores were obtained. The primary outcome measure was change in fibroid symptoms and HRQOL after UAE. Secondary outcomes included objective measures of patient satisfaction, and the decrease in uterine volume after UAE. RESULTS: Questionnaires were completed by 64 women (80.0%) at a mean of 32.1 months from UAE (range: 57.5-6 months). After UAE, mean uterine volume decreased by 26.3% (95% CI 19.6-33.0), and 17 of 79 women (21.5%) underwent an additional procedure after a mean of 18.6 months. Symptom severity scores decreased by a mean of 35.2% (95% CI 29.3-41.1) and HRQOL scores increased by a mean of 35.7% (95% CI 28.9 42.4). Satisfaction with UAE was correlated with the change in symptom severity and HRQL scores (P <.0001 and P=.0004, respectively) and the decrease in uterine volume after UAE (P=.0196). CONCLUSION: Women who undergo UAE have a significant decrease in symptom severity and increase in HRQOL, associated with high levels of satisfaction with the procedure, even when subsequent therapies are pursued. PMID- 15284776 TI - The role for preimplantation genetic diagnosis in balanced translocation carriers. AB - OBJECTIVE: Preimplantation genetic diagnosis is an established technique that provides an alternative to prenatal diagnosis for patients who are at risk of transmitting a serious genetic disorder to their offspring. Preimplantation genetic diagnosis has been used for couples who have been at risk for having offspring with single gene or X-linked disorders and for screening for common age related aneuploidy and in couples who themselves carry balanced chromosomal rearrangements. The aim of this study was to summarize our experience using preimplantation genetic diagnosis after the identification of a parental balanced translocation, specifically as it relates to the number of embryos that are suitable for transfer after preimplantation genetic diagnosis for a known translocation and aneuploidy screening. STUDY DESIGN: This is a retrospective review of data from a single center that involved 6 couples that initiated the process of preimplantation genetic diagnosis for translocation and aneuploidy screening by fluorescent in situ hybridization. RESULTS: A total of 65 embryos were obtained, of which 56 embryos (86%) were suitable for fluorescent in situ hybridization analysis. After fluorescent in situ hybridization, 1 embryo was diagnosed as normal or balanced (1.7%). Forty-three embryos (76.8%) were unbalanced for the translocation; 8 embryos (14.3%) were aneuploid, and 4 embryos (7.1%) were uninformative. There were no clinical pregnancies. CONCLUSION: In our experience, there are very few embryos that are available for transfer from these patients after translocation and aneuploidy screening because of multiple unbalanced segregation products and a high rate of aneuploidy. Factors that contributed to this may be related to which parent carries the translocation, methods that were used for in vitro fertilization, and advanced maternal age. Although preimplantation genetic diagnosis for translocation carriers theoretically can enhance the pregnancy rate for a couple, there are limitations. This information should be shared with couples who are contemplating preimplantation genetic diagnosis for translocation, and the options of sperm or egg donor should be considered. PMID- 15284779 TI - Comparison of laparoscopic-assisted vaginal hysterectomy with traditional hysterectomy for cost-effectiveness to employers. AB - OBJECTIVE: The purpose of this study was to compare the cost-effectiveness of laparoscopic-assisted vaginal hysterectomy to traditional total abdominal hysterectomy and total vaginal hysterectomy with regard not only to direct hospital costs but also to indirect costs. STUDY DESIGN: This was a combined retrospective cohort study (Canadian Task Force classification II-2) that was conducted in a suburban private practice. The cases of 268 patients who underwent hysterectomies over a 27-month period were analyzed to include clinical outcomes, direct hospital costs, and indirect costs (time to return to normal function, time to return to work, and time away from work required by other family members). RESULTS: For all patients, length of hospital stay and time of return to normal function were shorter for laparoscopic-assisted vaginal hysterectomy than for total abdominal hysterectomy and total vaginal hysterectomy. For working patients, time to return to work and time off for working family members were all significantly shorter after laparoscopic-assisted vaginal hysterectomy when compared with both total abdominal hysterectomy and total vaginal hysterectomy. Operating times were similar for total abdominal hysterectomy and laparoscopic assisted vaginal hysterectomy, and complications were greater for total abdominal hysterectomy. In a comparison of all procedures, direct hospital costs were greatest for laparoscopic-assisted vaginal hysterectomy and least for total vaginal hysterectomy. CONCLUSION: For most patients, laparoscopic-assisted vaginal hysterectomy provides a minimally invasive way to accomplish a hysterectomy with a lower cost to employers (payers) on the basis of lost work hours. PMID- 15284781 TI - Aggressive tocolysis does not prolong pregnancy or reduce neonatal morbidity after preterm premature rupture of the membranes. AB - OBJECTIVE: This study was undertaken to evaluate whether aggressive tocolysis improves pregnancy outcome after preterm premature rupture of the membranes (PPROM). STUDY DESIGN: Retrospective case-control study of patients with PPROM before 34 weeks of gestation, followed by a prospective cohort study with historical controls. The retrospective phase covered 1995 through 1999 when we used tocolysis aggressively. With the use of survival analysis, we compared latency in our cases with 4 published control series in which tocolysis was never used. On the basis of the results, we adopted a new protocol in mid-2000 limiting tocolysis to 48 hours after betamethasone dosing and we conducted a 2-year prospective evaluation of this new protocol. RESULTS: In the retrospective phase, tocolysis was used in 94% of 130 cases and maintained during 84% of 1162 total antenatal patient-days. There was no difference in latency between our cases and the published controls. One or more complications of tocolysis occurred in 18%. In the prospective study, 43% of 63 patients received tocolytics, but these were used at lower doses and were given during only 7% of 770 patient-days. Latency with this very limited tocolytic regimen (median 4.5 days, interquartile range 2.3 to 14.0) was not significantly different than during the last 24 months of aggressive tocolysis (median 3.8 days, 1.8 to 14 days, P=.16) and there were no differences in neonatal morbidity. CONCLUSION: Aggressive tocolysis after PPROM causes significant maternal morbidity, but does not increase latency or decrease neonatal morbidity compared with either very limited tocolysis or no tocolysis at all. PMID- 15284784 TI - A retrospective review of isoimmunized pregnancies managed by middle cerebral artery peak systolic velocity. AB - OBJECTIVE: The purpose of this study was to determine the clinical outcome of isoimmunized pregnancies managed primarily by middle cerebral artery peak systolic velocity. STUDY DESIGN: A retrospective chart review was conducted of isoimmunized pregnancies that underwent ultrasound examinations from January 1, 2001, through May 1, 2003. Ultrasound reports, laboratory tests, and maternal and neonatal charts were reviewed. RESULTS: Women with a clinically significant red blood cell antibody and titer value were included. The study population consisted of 39 women (40 pregnancies, 42 fetuses). Patients with a middle cerebral artery peak systolic velocity of > or =1.5 MoM were offered amniocentesis. Seven pregnancies had an abnormal middle cerebral artery peak systolic velocity. Three of these infants had significant anemia. Six of the 7 pregnancies required an exchange transfusion. None of the 33 pregnancies (35 neonates) with normal middle cerebral artery peak systolic velocity measurements resulted in a neonate with significant anemia or severe hyperbilirubinemia. CONCLUSION: The clinical outcome of these pregnancies supports the use of middle cerebral artery peak systolic velocity measurements in the management of isoimmunized pregnancies. PMID- 15284786 TI - Factors impacting injury documentation after sexual assault: role of examiner experience and gender. AB - OBJECTIVE: This study was undertaken to determine whether physician gender or level of experience is associated with the prevalence of trauma documented in victims after sexual assault. STUDY DESIGN: All female patients 15 years or older reporting to an urban hospital with a complaint of sexual assault between January 1997 and September 1999 underwent a standardized history and physical examination by a second- or third-year resident in obstetrics and gynecology. Data were abstracted and verified. A chi(2) or Fisher exact test was used for categoric analysis. RESULTS: The overall prevalence of genital trauma was 21% in the 662 patients available for analysis. The prevalence of genital trauma documented by second- and third-year residents was 50 of 191 patients (26.2%) and 90 of 471 patients (19.1%), respectively (P=.04), despite similar assault characteristics between the 2 groups. The prevalence of genital trauma documented by male examiners (105/499 [21.0%]) and female examiners (35/160 [21.9%]) did not differ (P=.8). All examiners documented a similar prevalence of body trauma (52%). CONCLUSION: This study supports the hypothesis that the examiner's experience level may influence the prevalence of genital trauma documented after a sexual assault. Genital trauma documented was not associated with examiner gender in this study. PMID- 15284788 TI - Development of a large-scale obstetric quality-improvement program that focused on the nulliparous patient at term. AB - OBJECTIVE: The purpose of this study was to identify an appropriate population and a balanced set of maternal and neonatal measures to drive a hospital network obstetric quality improvement program. STUDY DESIGN: Sutter Health, a large Northern California health care system with>40,000 births annually, served as the site for this project. We chose to focus on the standardized nulliparous patients: term, singleton, and vertex. A multidisciplinary task force evaluated and selected perinatal outcome and process measures. Data from every hospital were collected prospectively electronically and analyzed centrally. RESULTS: Outcome measures that were selected included term, singleton, and vertex rates of 3rd/4th-degree laceration, cesarean birth, 5-minute Apgar score of <7, and patient satisfaction. The process measures included episiotomy, induction (37-41 weeks), and admittance with cervical dilation of > or =3 cm. Data collection completeness improved each quarter; by the end of 2002, the data collection completeness rate had reached 99.7%. Every measure demonstrated a large variation among our hospitals, which indicates opportunities for improvement. CONCLUSION: This balanced set of measures for term, singleton, and vertex patients has been straightforward to collect over a large and diverse hospital system and has engaged all participants successfully. PMID- 15284791 TI - Patient satisfaction after the treatment of vulvovaginal erosive lichen planus with topical clobetasol and tacrolimus: a survey study. AB - OBJECTIVE: The purpose of this study was to compare patient satisfaction with the topical immune system modulator tacrolimus to topical clobetasol during treatment for vulvovaginal erosive lichen planus. STUDY DESIGN: Subjects who had been diagnosed with vulvovaginal erosive lichen planus between June 2000 and May 2001 received a mail survey regarding clinical satisfaction and response to treatment with clobetasol and tacrolimus. Satisfaction was assessed with a 100-mm visual analogue scale (very unsatisfied, 0; very satisfied, 100). Satisfaction was compared with the use of a paired t-test. RESULTS: Nineteen subjects met the inclusion criteria; 17 subjects (89%) returned completed surveys. Sixteen of the 17 women reported clobetasol therapy, and 11 of the 17 subjects acknowledged the use of tacrolimus therapy. All but 1 of the women who received tacrolimus had been treated previously with clobetasol therapy. All subjects reported experiencing sexual pain before their initial examination. After treatment with clobetasol, 2 of 16 women reported pain-free intercourse. Two additional women reported pain-free intercourse after switching to tacrolimus therapy. Ten subjects who had used both treatments rated tacrolimus therapy as significantly more satisfactory than clobetasol therapy (63 vs 38 mm; P=.03). CONCLUSION: The use of topical tacrolimus improves satisfaction and may result in better clinical outcomes than therapy with clobetasol for the treatment of vulvovaginal erosive lichen planus. PMID- 15284794 TI - Potential factors affecting embryo survival and clinical outcome with cryopreserved pronuclear human embryos. AB - OBJECTIVE: This study was undertaken to determine whether the method of fertilization has a significant impact on survival and/or clinical pregnancy rates of cryopreserved human pronuclear (2PN) stage embryos. DESIGN: A retrospective analysis of cryosurvival and clinical pregnancy rates after thawing of 2PN stage embryos from January 2000 through December 2002 in a private Assisted Reproductive Technology (ART) center. MATERIAL AND METHODS: A total of 1408 human 2PN embryos were cryopreserved using a Planer Kryo 10 Series III freezing unit (TS Scientific, Perkasie, Pa) after dehydration/equilibration through Propanediol (Sigma Chemical, St. Louis, Mo) and sucrose. On thawing, embryos were cultured in vitro with P-1 medium with 10% Serum Substitute Supplement (Irvine Scientific, Santa Ana, Calif). Embryo transfer was performed at 40 to 48 hours from time of thaw into a recipient uterus after standard estradiol/progesterone preparation. RESULTS: In 2000, 78% of all frozen 2PN embryos survived and were transferred in 181 cycles producing a delivery rate of 26% per transfer. However, 59% of these cycles were intracytoplasmic sperm injection (ICSI), and the survival of frozen 2PN from these cycles (72%) was lower than the respective survival of frozen 2PN embryos from in vitro fertilization (IVF) (81%; P<.025). Changes to protocols for thawing frozen 2PN embryos were therefore explored and implemented during 2001, resulting in equivalent survival rates of frozen 2PN embryos from IVF and ICSI during 2001 (78% and 80%, respectively) and 2002 (73% and 74%, respectively). Coincidentally, the proportion of all cycles that were performed with ICSI increased (73% in 2001 to 78% in 2002; P<.01) and pregnancy rates after transfer of frozen/thawed 2PN embryos from ICSI increased from 15% in 2000 to 30% in 2002. CONCLUSION: 2PN stage embryo cryosurvival may be negatively affected by ICSI, possibly caused by disruption of the zona pellucida and vitelline membrane before cryopreservation, and/or because ICSI promotes fertilization of some compromised eggs (producing compromised 2PN embryos) that would not have fertilized by conventional IVF. Without close attention to embryo freezing and thawing protocols relative to outcome, lower cryosurvival of unselected ICSI-produced embryos can negatively impact pregnancy outcomes. PMID- 15284796 TI - Symptomatic hypocalcemia after tocolytic therapy with magnesium sulfate and nifedipine. AB - OBJECTIVES: In this study, we presented 2 cases and evaluated the evidence for symptomatic hypocalcemia after treatment with magnesium sulfate alone or combined with use of nifedipine. STUDY DESIGN: Case reports, such as the one that follows, and literature review were used. A 25-year-old gravida presented at 33 weeks' gestation with advanced preterm labor. She received magnesium sulfate followed by nifedipine and experienced bilateral hand contractures 12 hours after discontinuation of magnesium sulfate. Total serum calcium was 5.4 mg/dL. A 35 year-old gravida presented at 26 weeks' gestation with ruptured membranes and received magnesium sulfate until it was discontinued prematurely because of pulmonary edema. Twenty hours later she experienced bilateral hand contractures; total serum calcium was 5.9 m/dL. Symptoms for both patients resolved with calcium gluconate therapy. RESULTS: Hypocalcemia is a well-recognized complication of magnesium sulfate infusion. These are the fifth and sixth symptomatic case reports, as identified by Medline Search. Our first case is the only report in which the subsequent use of nifedipine may have been a factor. Little has been reported on the possible toxicity associated with the combined or sequential use of magnesium sulfate and nifedipine. CONCLUSION: Marked hypocalcemia is clearly associated with magnesium sulfate infusion, is likely dose related, and may appear after discontinuation of magnesium sulfate therapy. Moreover, while the evidence for synergistic toxicity of magnesium sulfate and nifedipine is sparse, caution is advised when these agents are used together. PMID- 15284797 TI - The use of prostaglandin E2 in pregnant patients with asthma. AB - OBJECTIVE: Prostaglandin E(2) is a pharmacologic agent that is used commonly in obstetrics; however, its usage in patients with asthma is unclear. The study objective was to examine pregnant patients with asthma who received prostaglandin E(2). STUDY DESIGN: All pregnancies that were given prostaglandin E(2) suppositories and/or gel were recorded prospectively from January 1989 through December 2000. Those cases with a history of asthma or active asthma were analyzed for any clinical evidence of disease exacerbation after the administration of the agent. Clinical exacerbation was defined as any respiratory complaint that followed drug usage, the initiation of bronchodilator medications by patients currently not on therapy, or an increase in bronchodilator usage by patients with active disease. RESULTS: During the study period, 2513 patients received treatment with the cervical ripening gel, of whom 158 patients had a history of asthma or active asthma. Additionally, 536 patients were administered the 20-mg suppositories, of whom 31 patients had a history of asthma or active asthma. Thus, a total of 189 patients with a history of asthma or active asthma were exposed to prostaglandin E(2), and none of the patients had any evidence of a clinical exacerbation of the disease (0/189 cases; 95% CI, 0- 2%). CONCLUSION: Based on the 95% CI of these data, the maximum risk for the development of a clinical exacerbation of asthma, if exposed to the obstetric forms of prostaglandin E(2), is < or =2%. Although all drug usage in patients with asthma should be monitored carefully, this information would support the usage of prostaglandin E(2), if obstetrically indicated, in pregnant patients with asthma. PMID- 15284799 TI - FISHing for acrocentric associations between chromosomes 14 and 21 in human oogenesis. AB - OBJECTIVE: The purpose of this study was to search for cytologic evidence of robertsonian translocation formation that involves chromosomes 14q and 21q in human oogenesis with the use of dual color fluorescent in situ hybridization with whole chromosome paints. STUDY DESIGN: The oocytes from a chromosomally normal fetus at 23.5 weeks of gestation underwent cohybridization with chromosome specific DNA libraries from chromosomes 14 and 21. The nuclei were scored for the proportion of meiosis I prophase substages and for hybridization efficiency and were evaluated for the presence of hybridization signals that were suggestive of heterologous associations between chromosomes 14q and 21q in zygotene, pachytene, and diplotene. RESULTS: A total of 1769 meiotic nuclei were analyzed. Of 272 informative nuclei at zygotene, pachytene, and diplotene, 1 nucleus at pachytene demonstrated hybridization signals for chromosomes 14 and 21 that could be consistent with a robertsonian translocation. CONCLUSION: A heterologous association between chromosomes 14q and 21q that possibly represent robertsonian translocation formation was observed cytologically with the use of fluorescent in situ hybridization. PMID- 15284801 TI - Human endometriotic xenografts in immunodeficient RAG-2/gamma(c)KO mice. AB - OBJECTIVE: The purpose of this study was to create a novel animal model for studies of endometriosis. STUDY DESIGN: To facilitate the study of the transplantation of endometriosis into immunodeficient RAG-2/gamma(c)KO mice, endometriosis biopsy specimens were collected from 19 women by laparoscopic surgery and grafted subcutaneously into the mice, which were treated subsequently with estradiol and progesterone to create 28-day artificial cycles. The grafts were collected during the first, second, and fourth cycles and were evaluated histologically for evidence of bleeding and immunocytochemically for estrogen receptor and progesterone receptor. RESULTS: Biopsy specimens that contained endometrium-like glands were well accepted (>90% success). These grafts maintained glandular morphologic condition, estrogen receptor, and progesterone receptor; bled after progesterone withdrawal; and formed chocolate cysts. However, biopsy specimens that lacked glands or that consisted of peritoneal adhesions and stroma were accepted poorly <5% success) and failed to show evidence of estrogen receptor, progesterone receptor, or cyclic bleeding. CONCLUSION: Human endometriosis transplanted into RAG-2/gamma(c)KO mice can provide a model for endometriotic bleeding. PMID- 15284808 TI - The DEPA scoring system and its correlation with the healing rate of diabetic foot ulcers. AB - The purpose of this study was to examine the validity of a new scoring system in predicting the outcome of diabetic foot ulcers. The scoring system (DEPA score) includes the depth of the ulcer (D), the extent of bacterial colonization (E), the phase of ulcer healing (P) and the associated underlying etiology (A). The scoring system was validated against the clinical outcome in terms of healing and lower-limb amputations. Eighty-four patients were included in the study: 32 patients had a DEPA score of < or =6, 34 patients had a DEPA score of 7 to 9, and 18 patients had a DEPA score of > or =10. Using the Spearman nonparametric correlation test, DEPA scoring system was accurate in predicting the outcome of management (correlation coefficient, 0.78; 95% confidence interval, 0.68 to 0.86; P <.0001) at a mean follow-up of 20 weeks. The correlation was further validated by using a linear regression model (r = 0.85; slope best-fit value, 0.51; 95% confidence interval, 0.41 to 0.59; P <.0001). All patients with DEPA scores < or =6 had excellent healing, whereas only 15% of those with a score of > or =10 had complete healing in <20 weeks. In conclusion, an increasing DEPA score is associated with increased risk of amputation and poor healing. Furthermore, inclusion of the phase of ulcer healing into the DEPA system increases the accuracy of predicting the outcome of diabetic foot ulcers. PMID- 15284809 TI - Effect of a diode laser on wound healing by using diabetic and nondiabetic mice. AB - The purpose of this study was to evaluate a 980-nm gallium-aluminum-arsenide diode laser for wound healing. Using genetically diabetic and nondiabetic mice, two 6-mm wounds were created on the back of each mouse by using a punch biopsy. The mice were assigned to 1 of 4 subgroups for laser treatment at different fluence and frequency of treatment: 5 W (18 J/cm2) every 2 days, 5 W (18 J/cm2) every 4 days, 10 W (36 J/cm2) every 2 days, and 10 W (36 J/cm2) every 4 days. In addition, control mice were used and the wounds were allowed to heal naturally. Wound healing was evaluated on days 5, 12, and 19 by percentage of wounds healed and percent wound closure. A maximum of 5 mice per subgroup were killed at days 7, 14, and 21, and histology was conducted on the wound sites. For diabetic mice receiving 5 W every 2 days, the percentage of wounds healed after 19 days was 100% versus 40% in the control group. Only 20% of wounds in the 10-W diabetic subgroups achieved healing during the same period. For the subgroups whose wounds did not completely heal, all but the 10 W every 2 days subgroup had average closure of >90%. The 100% closure for the 5 W every 2 days subgroup was significantly greater than the other subgroups. For nondiabetic mice, 100% of the wounds in the 5 W every 4 days and control subgroups were completely healed, whereas 90% of the wounds from the 5 W every 2 days and the 10 W every 4 days subgroups were completely healed. In the latter 2 subgroups, wound closure was 99.4% and 98.8%, respectively. These differences were not significant. The histologic results confirmed these findings. In conclusion, treatment at 18 J/cm2 shows a beneficial effect on wound healing in diabetic mice and does not have a detrimental effect in nondiabetic mice. PMID- 15284810 TI - Vascular perfusion of the long dorsal arm versus chevron osteotomy: a cadaveric injection study. AB - Iatrogenic disruption of the first metatarsal head's blood supply after a distal osteotomy may lead to adverse events such as delayed union or avascular necrosis of the capital fragment. In this cadaveric study, 2 types of distal first metatarsal osteotomies were performed on 7 fresh-frozen specimens (long dorsal arm osteotomy, N = 4; chevron osteotomy, N = 3). For each specimen, the respective osteotomy was created. The dorsalis pedis artery was cannulated at the midfoot level and infiltrated with methylene blue dye until resistance was felt. The nutrient artery to the first metatarsal shaft was then exposed and the capital fragment was dissected free from the remaining soft tissues and visually inspected for the presence and distribution of dye. One of the chevron specimens was excluded because of technical error during the injection portion of the study. In all 4 of the long dorsal arm specimens, dye was observed in the entire dorsal cortical-cancellous wing and the cancellous metatarsal head region. In contrast, dye was completely absent in the capital fragment of the 2 chevron specimens. This cadaveric study showed that the vascular supply to the long dorsal arm osteotomy includes the nutrient artery, whereas the chevron osteotomy does not. This suggests that the long dorsal arm osteotomy may present a less risk of vascular compromise to the capital fragment. PMID- 15284811 TI - Strength analysis of intraosseous wire fixation for avulsion fractures of the fifth metatarsal base. AB - The objective of this study was to gather pilot data of the pullout strength of intraosseous wire for fixation of fifth metatarsal avulsion fractures and to compare intraosseous wire fixation with tension-band wiring. Osteotomies consistent with fifth metatarsal avulsion fractures were created in 5 matched pairs of cadaver limbs. One limb of each pair underwent fixation with intraosseous wiring and the other with tension band wiring. Metatarsals were then loaded to failure, defined as the maximum force achieved before the slope of the load curve moved from a positive to a negative value. Tension-band wiring showed a mean strength of 164.5 +/- 103.7 N compared with a mean strength of 113.7 +/- 46.6 N for intraosseous wiring. No significant difference in strength was shown between methods of fixation. Although the power of this data is small (.143), the data indicate that intraosseous wiring shows similar pullout strength when compared with tension-band wiring for fixation of fifth metatarsal avulsion fractures. PMID- 15284812 TI - Second metatarsophalangeal joint arthrography: a cadaveric correlation study. AB - Arthrography of the second metatarsophalangeal joint is an important diagnostic tool to evaluate the integrity of the plantar plate and to aid in the decision process for surgical intervention. A variety of filling patterns have been identified with lesser metatarsophalangeal joint arthrography and their significance with soft-tissue pathology remains to be completely understood. The purpose of this cadaveric study was to evaluate dye patterns in a series of arthrograms of the second metatarsophalangeal joint and to correlate them with identifiable anatomic lesions or structural variants. Thirty-nine cadaveric specimens (including 28 matched pairs) underwent second metatarsophalangeal joint arthrography with a colored radiopaque dye. Arthrographic findings were observed and recorded. Specimens exhibiting dye extravasation outside of the capsular constraints of the joint were dissected to discover any soft-tissue abnormalities. Twenty-one percent of specimens exhibited abnormal extravasation of dye outside of the joint capsule. A plantar plate tear was identified in 2 of these specimens. Filling of the first intermetatarsophalangeal bursa occurred in 6 specimens. However, because this finding was identified in 2 matched pairs, an anatomic variance is suggested rather than a pathologic entity. This cadaveric study shows that anatomic variances exist concerning the second metatarsophalangeal capsule and that arthrography should be correlated with the clinical scenario. PMID- 15284813 TI - Gait analysis after talonavicular joint fusion: 2 case reports. AB - The purpose of this study was to analyze the alterations in frontal-plane hindfoot motion and in plantar pressure in 2 patients (3 feet) after talonavicular joint fusion. Three-dimensional movement of the hindfoot was recorded by using an electromagnetic motion analysis system. The results indicated that frontal-plane hindfoot motion of the operative foot was substantially reduced during the stance phase of walking. In addition, each patient's frontal-plane hindfoot angle while in the resting standing position was decreased and plantar pressure under the medial longitudinal arch was reduced after surgery. PMID- 15284814 TI - Calcaneal brown tumor with primary hyperparathyroidism caused by parathyroid carcinoma: an atypical localization. AB - Brown tumors are one of the characteristics of primary hyperparathyroidism, although, in some cases, they are noted with secondary hyperparathyroidism as well. The authors present a case of a 50-year-old woman with primary hyperparathyroidism caused by parathyroid carcinoma with an unusual location of a brown tumor in the calcaneus. She first presented with pain and swelling over the heel and ankle, and the diagnosis was suspected by radiographs. Biopsy of the calcaneal lesion confirmed a brown tumor. After the parathyroid lesion was removed surgically, her symptoms were relieved. The calcaneal lesion was treated with immobilization of the foot. PMID- 15284815 TI - Arteriovenous malformation of the foot: a case presentation. AB - Arteriovenous malformations have been reported as rare clinical entities in the foot. When these lesions present, they are most often found in the brain, lungs, and pelvis/thigh of the lower extremity and develop as a result of failed fetal vascular development. Arteriovenous malformations can present with a variety of dermatologic and osseous manifestations. A delayed or missed diagnosis can have severe effects, including chronic ulcerations and osseous manifestations, and lead to amputation. Once identified, a complete evaluation is needed to provide the clinician with a full understanding of the magnitude of the deformity and the potential implications to the limb's vascular status after treatment. A concise review of the literature is provided with a case presentation of an arteriovenous malformation within the foot that was not suspected based on the clinical and imaging evaluation. PMID- 15284816 TI - Excision of a Dermatobia hominis larva from the heel of a South American traveler: a case report. AB - Although foot and ankle specialists are well versed in treating insect bites and foreign bodies, many physicians in the United States are unfamiliar with parasitic organisms that are common in other parts of the world. This article presents a case of a patient inoculated in the posterior heel with the larva of a Dermatobia hominis, or human bot fly. Excision of the larva provided a complete resolution of the patient's symptoms. Although the initial clinical presentation suggested a simple foreign body, the patient's recent travel history to Brazil shows that a thorough history is essential to establishing a complete list of differential diagnoses. PMID- 15284817 TI - Percutaneous tendo-Achilles lengthening with a large-gauge needle: a modification of the Ponseti technique for correction of idiopathic clubfoot. AB - The Ponseti technique has become standard for the treatment of congenital idiopathic clubfoot. Treatment includes serial manipulations and casting, accompanied by percutaneous tenotomy of the Achilles tendon. In this article, the authors describe a modification in the Achilles tenotomy technique by using a large-gauge hypodermic needle in the outpatient setting. PMID- 15284818 TI - Plantar plate repair of the second metatarsophalangeal joint: technique and tips. PMID- 15284819 TI - Verrucous carcinoma. PMID- 15284826 TI - [Cytokine production and dermatophytosis]. AB - The characteristic pathological feature of dermatomycosis is numerous neutrophilic infiltrates within the epidermis. However, the precise mechanism of this infiltration remains unknown. In this study, interleukins 1 beta, 6, and 8, monocyte chemotactic protein-1 (MCP-1), and tumor necrosis factor (TNF)-alpha levels in the medium where keratinocytes were co-cultured with Candida albicans, Malassezia and Trichophyton mentagrophytes, were determined by enzyme-linked immunosorbent assays (ELISAs) in order to estimate the effect of these fungi on the cytokine production from human keratinocytes. The IL-8 level in the supernatants increased with 1 to 14 hours of co-culture in response to live C. albicans, but the other cytokines were undetectable. Furthermore, the mRNA of IL- 8 in keratinocytes was also confirmed to increased. This data suggested that C. albicans directly induce interleukin 8 production from human keratinocytes without activated macrophages. The IL-6, IL-8, and TNF-alpha levels in the culture supernatants increased with 1 to 24 hours of co-culture with keratinocytes and Malassezia species but the MCP-1 level was undetectable. The IL 8 and TNF-alpha levels in the culture supernatants increased with 1 to 24 hours of co-culture with keratinocytes and Trichophyton mentagrophytes but the other cytokine levels were undetectable. The ELISA analysis of cytokine production by human keratinocytes will provide useful information in understanding the pathogenesis of dermatomycosis. PMID- 15284827 TI - [Antimycotics suppress interleukin-4 and interleukin-5 production in anti-CD3 plus anti-CD28-stimulated T cells from patients with atopic dermatitis]. AB - It is reported that antimycotic agents are effective for the treatment of patients with atopic dermatitis (AD). We studied in vitro effects of antimycotics on T helper-1 and T helper-2 cytokine production in anti-CD3 plus anti-CD28 stimulated T cells from AD patients and normal donors. The amounts of interleukin 4 (IL-4) and IL-5 secreted by anti-CD3/CD28-stimulated T cells were higher in AD patients than in normal donors. Azole derivatives, ketoconazole, itraconazole, miconazole and non-azole terbinafine hydrochloride and tolnaftate reduced IL-4 and IL-5 secretion without altering that of IFN-gamma and IL-2 in anti-CD3/CD28 stimulated T cells from both AD patients and normal donors. The azole derivatives were more inhibitory than non-azole antimycotics. These antimycotics reduced the anti-CD3/CD28-induced mRNA expression and promoter activities for IL-4 and IL-5. The cAMP analogue dibutyryl cAMP reversed the inhibitory effects of the antimycotics on IL-4 and IL-5 secretion, mRNA expression, and promoter activities. Anti-CD3/CD28 transiently (< or = 5 min) increased intracellular cAMP in T cells, and the increase was greater in AD patients than in normal donors. The increase of cAMP by anti-CD3/CD28 correlated with IL-4 and IL-5 secretion by anti-CD3/CD28. The transient cAMP increase was suppressed by antimycotics, and azole derivatives were more suppressive than non-azoles. Azole derivatives inhibited the activity of cAMP-synthesizing adenylate cyclase while terbinafine hydrochloride and tolnaftate enhanced the activity of cAMP-hydrolyzing cyclic nucleotide phosphodiesterase in AD and normal T cells. These results suggest that the antimycotics may suppress IL-4 and IL-5 production by reducing cAMP signal, and strengthen the concept of their potential use for the suppression of T helper 2-mediated allergic reactions. PMID- 15284828 TI - [Optimal dosages and cycles of itraconazole pulse therapy for onychomycosis]. AB - OBJECTIVE: To compare the international standard regimen of itraconazole pulse therapy with low daily-dose pulse therapies, which are widely conducted in Japan. DESIGN: Randomized, double-blind, parallel-group comparative study. PATIENTS: 186 patients with a big toenail showing onychomycosis symptoms such as opacity. INTERVENTION: Patients were assigned to Group I (200 mg/d, 3 cycles), Group II (200 mg/d, 6 cycles) or Group III (400 mg/d, 3 cycles). All received itraconazole orally. MAIN OUTCOME MEASURE: Cure or complete response - assessment based on improvement in the opacity ratio and microscopic examination. RESULTS: The clinical response rates (cure plus complete response) at week 24 were Group I: 14.9%; Group II: 25.5%; Group III: 32.7%. At week 48, 17 patients were cured in Group III - up from 3 at week 24. At week 48 the area under the nail plate concentration-time curves (AUC) was: Group I: 6,084 +/- 3,696 ng h/g; Group II: 10,448 +/- 6,980 ng h/g; Group III: 24,189 +/- 15,157 ng h/g. There was no difference among the three groups in the incidence of adverse drug reactions. CONCLUSIONS: The clinical response rates demonstrated that the pulse therapy of 400 mg/d itraconazole for 3 cycles was most effective. The significantly higher Group III AUC (week 48) suggests that, when the same total amount of itraconazole is administered, this drug remains in nail plates longer following pulse therapy with a higher daily dose. It was also suggested that clinical efficacy correlated with the duration of the presence of itraconazole. PMID- 15284829 TI - [Statistical study of dermatomycosis for 30 years (1968-1997) in sendai national hospital]. AB - A statistical study on dermatomycoses for the 30 years from 1968 to 1997 in the dermatologic section of Sendai National Hospital was carried out with the following results. The total number of dermatomycosis patients was 14,259 and accounted for 9.59% of all new outpatients during this period. These cases of dermatomycoses were composed of the following: dermatophytoses 10,656, candidiasis 3,287, malassezia infection 566, sporothrichosis 20, aspergillosis 7, and chromomycosis 1. Annual changes in number of dermatophytoses varied in each clinical type: tinea corporis and tinea cruris had increased by the end of the 1970s, and there after decreased gradually until recent years. Tinea pedis and tinea unguium, on the contrary, increased after the 1980s. Age distribution of all clinical forms of dermatophytoses changed gradually, and its peak of the distribution curve shifted to an older site each year, while the number of younger generation patients decreased. Mycologically Trichophyton (T.) rubrum was mainly isolated from all types of dermatophytoses, and T. mentagrophytes followed. The ratio of these two species (R/M) in tinea pedis was 1.25, and the ratio of T. mentagrophytes was relatively high. Epidermophyton floccosum was continuously isolated, but its frequency in recent years has decreased. Microsporum canis infection increased from the middle period of this research, but after 1990 decreased gradually. The other dermatophytes were found sporadically. In recent years the species isolated have become more simplified. Among candidiasis, infantile candidiasis increased dramatically in the 1970s but soon decreased. Intertrigo type also increased in the same period and then decreased to an intermediate level. Paronychia, onychia and erosio interdigitale were constantly found in small numbers, while malassezia infection remained at a constant level and was found more in male patients. PMID- 15284830 TI - RFLP analysis of the internal transcribed spacer regions of Sporothrix schenckii. AB - Restriction fragment length polymorphism (RFLP) analysis was performed on the internal transcribed spacer regions of 204 Sporothrix schenckii isolates and on one strain each of the related fungi, S. schenckii var. luriei, S. curviconia, S. inflata and Ceratocystis stenoceras. S. schenckii isolates, which have been collected from around the world, have already been typed according to their mitochondrial DNA (mtDNA), and are kept in the Department of Dermatology, Kanazawa Medical University, Japan. Approximately 600 bp of the internal transcribed spacer region 1 (ITS1) of their nuclear ribosomal RNA gene (rDNA), 5.8S rDNA and ITS2 was amplified by PCR. From ITS-RFLP analysis of the PCR products, S. schenckii isolates comprised 4 types, rDNA types I - IV. The rDNA type I - III strains corresponded to the Group A strains (mtDNA types 1 - 3, 11, 14 - 19, 22 and 23), while the rDNA type IV strains corresponded to the Group B strains (mtDNA types 4 - 10, 12, 13, 20 and 21), as previously categorized according to their mtDNA-RFLP. The ITS-RFLP patterns of the above 4 related fungi all differed from those of the 4 rDNA types of S. schenckii. Furthermore, only 22 (3.5%) out of a sequence of about 620 bases of the ITS regions of the rDNA differed among representatives of the mtDNA types 1 - 5, 7, 11, 14 - 19, 22 and 23. This difference in the ITS region is smaller than the 10% difference among isolates when estimated by mtDNA-RFLP. From the phylogenetic tree based on the base sequences, rDNA type I - III strains belong to Group I, while rDNA type IV strains belong to Group II which correspond with Groups A and B based on their mtDNA. The Group I strains are predominant in South America and Africa, while Group II are predominant in Australia and Asia. ITS-RFLP analysis is better than mtDNA-RFLP in allowing faster discrimination and identification, and for its ability to divide the 4 types into groups, which is useful in clinical diagnosis and epidemiological investigations of S. schenckii. PMID- 15284831 TI - [A case of tinea capitis with a couple of nodular lesions possibly resulting from topical application of tacrolimus]. AB - We herein report a case of tinea capitis initially showing a couple of nodular lesions. The patient was a 66-year-old woman who had seen a nearby dermatologist for itching on her head and had been treated with a topical steroid followed by tacrolimus application for one month. Because pseudolymphoma-like erythematous nodules developed at two sites, she visited us. Two weeks after stopping all medication, some slight scaling was found around these nodules. On KOH direct microscopic examination, many filamentous elements around hair shafts were observed. Biopsy of the nodules confirmed the destruction of hair follicles surrounded by granulomatous inflammation histologically. Grocott staining of the same specimen revealed a few short fungal hyphae as well as spores. She was also diagnosed as tinea pedis by direct microscopic examination of her feet. Trichophyton rubrum was isolated from scales of both her head and feet on Sabouraud's dextrose agar at 25 degrees C. Kerion celsi (KC) is usually clinically preceded by a gray patch or black dots. Such a typical course of KC, however, was not observed in our patient. Tacrolimus was thought to have possibly played an important role in modifying tinea capitis. PMID- 15284832 TI - Trends in prenatal diagnosis of critical cardiac defects in an integrated obstetric and pediatric cardiac imaging center. AB - OBJECTIVE: Prenatal diagnosis of critical (requiring neonatal intervention) cardiac defects (critical congenital heart diseases (CCHD)) improves survival, yet detection of such malformations is poor. Our institution changed its practice and integrated a pediatric cardiologist into the perinatal team. The purpose of this study was to evaluate how this change affected the rate of detected congenital heart disease (CHD) and the diagnostic accuracy. STUDY DESIGN: Obstetrical ultrasounds of mothers at high and normal risk for fetal CCHD at a single center between 1991 and 2001 were reviewed. Rate of detected CCHD, positive predictive values and false positives were compared before and after pediatric cardiology integration. RESULTS: Between the first and second time periods, the rate of detected CCHD increased from 6.8/1000 ultrasounds to 12.9/1000 ultrasounds (p=0.007), and positive predictive value increased from 75 to 96%. CONCLUSION: Collaboration with pediatric cardiology can significantly improve the rate of detected CCHD. These findings have significant implications for sonographer education and patient care. PMID- 15284833 TI - Left ventricular and carotid structure in untreated, uncomplicated essential hypertension: results from the Assessment Prognostic Risk Observational Survey (APROS). AB - The impact of hypertension on left ventricular (LV) and vascular structure and the relation of left ventricular hypertrophy (LVH) with vascular changes in untreated essential hypertensives has not been fully explored. This study investigated the prevalence of structural abnormalities of LV and carotid arteries and their determinants in a large population of untreated, uncomplicated essential hypertensive patients. The Assessment of Prognostic Risk Observational Survey was a multicentre (44 centres) prospective study including 1142 untreated hypertensives classified as low or medium cardiovascular risk on the basis of the routine diagnostic work-up recommended by the 1999 World Health Organization/International Society of Hypertension guidelines. All patients underwent ultrasound examinations of the heart and carotid arteries. LVH and carotid structural changes were diagnosed when: (1) LV mass index exceeded 125 g/m(2) in men and 110 g/m(2) in women; (2) there was at least one plaque (focal thickening>1.3 mm) in any segment of either carotid artery or a diffuse common carotid intima-media thickness (IMT) (average of IMT>/=0.8 mm) was present. Overall, 1074 patients (504 women, mean age 48.1+/-11.4 years) completed the study with ultrasonographic examinations of good technical quality. The prevalences of LVH and LV concentric remodelling in the total population were 26.8 and 10.7%, respectively. Eccentric hypertrophy was more prevalent than concentric hypertrophy (15.2 vs 11.6%). One or more carotid plaques or thickening were present in 27.4% of the patients. A stepwise increase in IM thickness occurred from the lowest values in patients with normal cardiac mass and geometry (0.68 mm) to intermediate in those with LV remodelling (0.76 mm) and eccentric LVH (0.81 mm) and to the highest level in patients with concentric LVH (0.87 mm). Patients with LV concentric remodelling and concentric LVH had a significantly greater relative carotid wall thickness than those with normal geometry and eccentric LVH (0.25 and 0.26 vs 0.18 and 0.19, respectively, P<0.01). According to a multivariate analysis age, blood glucose, systolic BP and pulse pressure were the main independent predictors of LVH, while age, systolic BP and total cholesterol were the variables with the greatest impact on IM thickening. To conclude, this study shows that: (1) altered patterns of LV structure and geometry and carotid structural changes occur in a large fraction of patients with untreated essential hypertension; (2) there is a significant association between carotid wall thickening and LVH; (3) the probability of LVH or carotid thickening is significantly greater in elderly, in patients with higher systolic BP and in patients with associated metabolic risk factors. PMID- 15284834 TI - A new atypical antipsychotic: quetiapine-induced sexual dysfunctions. AB - In this paper, we evaluated the new antipsychotic, quetiapine-induced sexual dysfunctions (SDs). The study group consisted of 36 patients with schizophrenia receiving quetiapine. The changes in general sexual functions were assessed by using Arizona Sexual Experience Scale (ASEX) and Udvalg for Kliniske Undersogelser (UKU) Side Effect Rating Scale at baseline and week 4. Also, prolactin (PRL) values were determined at baseline and week 4. There was statistically significant difference with respect to the mean ASEX score at week 4 compared with baseline. The most frequent SD was diminished libido in both male (31.8%) and female subjects (28.6%). No significant correlation was found between ASEX scores and PRL values. The results suggest that SDs are an important problem using even novel antipsychotic, quetiapine. PMID- 15284835 TI - TGF-beta1 neutralizing antibodies decrease the fibrotic effects of ischemic priapism. AB - The objective of this study was to evaluate the possible role of transforming growth factor beta 1 (TGF-beta1) antibodies (ab) for the prevention of fibrotic effects of priapism in a rat model. In total, 30 adult Sprague-Dawley rats were divided into five groups. Priapism with 6 h (group 1), priapism with 6 h+ab (group 2), priapism with 24 h (group 3), priapism with 24 h+ab (group 4) and control (group 5). Priapism was induced with a vacuum erection device and a rubber band was placed at the base of the erect penis. At 1 h after the initiation of priapism, TGF-beta1 antibodies were given intracavernosaly. All rats underwent electrical stimulation of the cavernous nerve after 8 weeks. Intracavernous and systemic blood pressures were measured during the procedure. Rats in group 1 showed significantly higher (intracavernosal pressure (ICP) pressures to cavernous nerve stimulation and had higher ICP/BP ratios when compared to other groups. Similarly, histopathologic examination revealed less fibrosis in group 2, compared with the other groups. Consequently, TGF-beta1 antibodies antagonise the fibrotic effects of TGF-beta1, especially in cases with duration of priapism less than 6 h. PMID- 15284836 TI - Circadian renal rhythms influenced by implanted encapsulated hANP-producing cells in Goldblatt hypertensive rats. AB - Renal excretion in experimental hypertensive rats implanted with encapsulated human atrial natriuretic peptide (hANP)-producing cells is circadian periodic. Chinese hamster ovary (CHO) cells transfected with the plasmid hANP-cDNA were encapsulated in biocompatible polycaprolactone capsules for intraperitoneal implantation into two-kidney, one-clip (2K1C) hypertensive rats. During a 12:12 light-dark cycle, as compared to control CHO cells, the implantation of encapsulated hANP-producing CHO cells was associated with an increase in the net excretion of water, sodium and potassium, and with a reversal of the advanced circadian phases related to renovascular hypertension in 2K1C rats. The increase in blood pressure postimplantation was delayed, and increases in renal blood flow, glomerular filtration rate, sodium output, urinary excretion and urinary cyclic GMP concentrations were also found. Implantation of encapsulated hANP producing cells affects circadian rhythms in kidney excretion functions of 2K1C rats, and may be useful for the treatment of cardiovascular disease. PMID- 15284837 TI - A defective nontransmissible recombinant Sendai virus mediates efficient gene transfer to airway epithelium in vivo. AB - Recombinant Sendai virus (SeV)-mediated gene transfer to differentiated airway epithelial cells has shown to be very efficient, because of its ability to overcome the intra- and extracellular barriers known to limit gene delivery. However, this virus is transmission competent and therefore unlikely to be suitable for use in clinical trials. A nontransmissible, replication-competent recombinant SeV has recently been developed by deleting the envelope Fusion (F) protein gene (SeV/DeltaF). Here we show that SeV/DeltaF is able to mediate beta galactosidase reporter gene transfer to the respiratory tract of mice in vivo, as well as to human nasal epithelial cells in vitro. Further, in an ex vivo model of differentiated airway epithelium, SeV/DeltaF gene transfer was not importantly inhibited by native mucus. When compared to the transmission-competent SeV in vivo, no difference in gene expression was observed at the time of peak expression. The development of an F-defective nontransmissible SeV, which can still efficiently mediate gene transfer to the airway epithelium, represents the first important step towards the use of a cytoplasmic RNA viral vector in clinical trials of gene therapy. PMID- 15284838 TI - Hematopoietic stem cell gene therapy of murine protoporphyria by methylguanine DNA-methyltransferase-mediated in vivo drug selection. AB - Erythropoietic protoporphyria (EPP) is an inherited defect of the ferrochelatase (FECH) gene characterized by the accumulation of toxic protoporphyrin in the liver and bone marrow resulting in severe skin photosensitivity. We previously described successful gene therapy of an animal model of the disease with erythroid-specific lentiviral vectors in the absence of preselection of corrected cells. However, the high-level of gene transfer obtained in mice is not translatable to large animal models and humans if there is no selective advantage for genetically modified hematopoietic stem cells (HSCs) in vivo. We used bicistronic SIN-lentiviral vectors coexpressing EGFP or FECH and the G156A mutated O6-methylguanine-DNA-methyltransferase (MGMT) gene, which allowed efficient in vivo selection of transduced HSCs after O6-benzylguanine and BCNU treatment. We demonstrate for the first time that the correction and in vivo expansion of deficient transduced HSC population can be obtained by this dual gene therapy, resulting in a progressive increase of normal RBCs in EPP mice and a complete correction of skin photosensitivity. Finally, we developed a novel bipromoter SIN-lentiviral vector with a constitutive expression of MGMT gene to allow the selection of HSCs and with an erythroid-specific expression of the FECH therapeutic gene. PMID- 15284839 TI - A clone in the hand. PMID- 15284840 TI - Lack of support for a genetic association of the XBP1 promoter polymorphism with bipolar disorder in probands of European origin. PMID- 15284842 TI - A national DNA bank in The Gambia, West Africa, and genomic research in developing countries. PMID- 15284843 TI - Evidence for lateral gene transfer from salmonids to two Schistosome species. PMID- 15284844 TI - Duplicating SNPs. PMID- 15284845 TI - TGF beta signaling in health and disease. PMID- 15284846 TI - Experiencing VEGF. PMID- 15284847 TI - Complex imprinting. PMID- 15284848 TI - STARTing to recycle. PMID- 15284851 TI - Nonsense-mediated decay approaches the clinic. AB - Nonsense-mediated decay (NMD) eliminates mRNAs containing premature termination codons and thus helps limit the synthesis of abnormal proteins. New results uncover a broader role of NMD as a pathway that also affects the expression of wild-type genes and alternative-splice products. Because the mechanisms by which NMD operates have received much attention, we discuss here the emerging awareness of the impact of NMD on the manifestation of human genetic diseases. We explore how an understanding of NMD accounts for phenotypic differences in diseases caused by premature termination codons. Specifically, we consider how the protective function of NMD sometimes benefits heterozygous carriers and, in contrast, sometimes contributes to a clinical picture of protein deficiency by inhibiting expression of partially functional proteins. Potential 'NMD therapeutics' will therefore need to strike a balance between the general physiological benefits of NMD and its detrimental effects in cases of specific genetic mutations. PMID- 15284852 TI - Imatinib induces a cytogenetic response in blast crisis or interferon failure chronic myeloid leukemia patients with e19a2 BCR-ABL transcripts. PMID- 15284853 TI - In situ leukemic plasmacytoid dendritic cells pattern of chemokine receptors expression and in vitro migratory response. AB - Plasmacytoid dendritic cell (PDC) leukemia/lymphoma is a rare neoplasm presenting cutaneous lesions at the time of diagnosis, followed by dissemination to bone marrow, lymph nodes, and other lymphoid and nonlymphoid organs. Since these leukemic counterparts of human PDC are similar to normal PDC, we studied their chemokine receptor equipment and their migratory capacities. We found both in skin lesions and in invaded lymph nodes an expression by tumor cells of CXCR3, CXCR4, and CCR7, and the concomitant expression by cells in the microenvironment of their respective ligands CXCL9, CXCL12, and CCL19. Moreover, flow cytometry phenotype of leukemic PDC (LPDC) revealed an unexpected expression of CCR6. We show that fresh tumor cells are able to migrate in response to CXCR4, CCR2, CCR5, CCR6, and CCR7 ligands, and the ability of CXCR3 ligands to increase the responsiveness to CXCL12. IL-3- or virus-induced activation of LPDC leads to downregulation of CXCR3 and CXCR4, and upregulation of CCR7, associated with the loss of response to CXCL12, and the acquisition of sensitivity to CCL19. Altogether, these results suggest that the preferential accumulation of LPDC in the skin or lymph nodes could be orchestrated by CXCR3, CXCR4, CCR6, and CCR7 ligands, found in nontumoral structures of invaded organs. PMID- 15284854 TI - Treatment intensity significantly influencing fibrosis in bone marrow independently of the cytogenetic response: meta-analysis of the long-term results from two prospective controlled trials on chronic myeloid leukemia. AB - Bone marrow fibrosis (MF) has been shown to indicate therapy failure in Ph(+) chronic myeloid leukemia (CML). However, the results on the development of MF during interferon-alpha therapy of CML are controversial. The significance of the interferon dose has not been considered as yet. In total, 627 bone marrow biopsies taken prospectively from 200 patients with CML recruited in two studies using different doses of interferon-alpha +/- low-dose cytosine arabinoside were examined for MF before and during therapy. The results showed that the risk of MF depended significantly on the interferon-alpha dose applied (P<0.000005). MF progressed during low-dose therapy (3 x 5 x 10(6) IU/week), but was prevented from progression when applying high dose (5 x 10(6) IU/m(2)/per day). MF disappeared when high-dose interferon-alpha was combined with low-dose cytosine arabinoside (P<0.000005). The risk of death markedly increased when MF occurred or progressed (P<0.0009), independent of all other prognostic factors evaluated including the cytogenetic response. In conclusion, the effectiveness of interferon-alpha on MF depends on the treatment intensity. MF reverses when combining high-dose interferon-alpha with low-dose cytosine arabinoside, but progresses when applying low-dose interferon-alpha. MF appears to be a significant early indicator of ineffective therapy in CML. PMID- 15284855 TI - A novel partner gene CIP29 containing a SAP domain with MLL identified in infantile myelomonocytic leukemia. PMID- 15284856 TI - Hematological defects in the oc/oc mouse, a model of infantile malignant osteopetrosis. AB - Infantile malignant osteopetrosis (IMO) is a rare and lethal disease characterized by an absence of bone resorption due to inactive OCLs. Affected patients display an increased bone mass and hematological defects. The osteopetrotic oc/oc mouse displays a bone phenotype similar to the one observed in IMO patients, and the same gene, Tcirg1, is mutated in this model and in the majority of these patients. Therefore, we explored in oc/oc mice the consequences of the perturbed bone microenvironment on hematopoiesis. We show that the myelomonocytic differentiation is increased, leading to an elevated number of OCLs and dendritic cells. B lymphopoiesis is blocked at the pro-B stage in the bone marrow of oc/oc mouse, leading to a low mature B-cell number. T-cell activation is also affected, with a reduction of IFNgamma secretion by splenic CD4(+) T cells. These alterations are associated with a low IL-7 expression in bone marrow. All these data indicate that the lack of bone resorption in oc/oc mice has important consequences in both myelopoiesis and lymphopoiesis, leading to a form of immunodeficiency. The oc/oc mouse is therefore an appropriate model to understand the hematological defects described in IMO patients, and to derive new therapeutic strategies. PMID- 15284857 TI - Is it now the time to update treatment protocols for lymphomas with new anti virus systems? PMID- 15284858 TI - Transplantation studies in C3-deficient animals reveal a novel role of the third complement component (C3) in engraftment of bone marrow cells. AB - Mice deficient in complement C3 (C3(-/-)) are hematologically normal under steady state conditions, and yet displayed a significant delay in hematopoietic recovery from either irradiation or transplantation of wild-type (WT) hematopoietic stem/progenitor cells (HSPC). Transplantation of histocompatible WT Sca-1(+) cells into C3(-/-) mice resulted in a (i) decrease in day 12 CFU-S, (ii) 5-7-day delay in platelet and leukocyte recovery, and (iii) reduced number of BM CFU-GM progenitors at day 16 after transplantation. Nevertheless, HSPC from C3(-/-) mice engrafted normally into irradiated WT mice, suggesting that there was a defect in the hematopoietic environment of C3(-/-) mice. Since C3(-/-) mice cannot activate/cleave C3, the C3 fragments C3a, C3a(des-Arg), and iC3b were examined for a role in HSPC engraftment. Liquid-phase C3a and C3a(des-Arg) increased CXCR4 incorporation into membrane lipid rafts (thus potentiating HSPC responses to SDF 1 gradients), whereas iC3b was deposited onto irradiated BM cells and functioned to tether CR3(CD11b/CD18)(+)HSPC to damaged stroma. The activity of C3a(des-Arg) suggested that C3aR(+)HSPC also expressed the C5L2 (receptor for C3a and C3a(des Arg)) and this was confirmed. In conclusion, a novel mechanism for HSC engraftment was identified, which involves complement activation and specific C3 fragments that promote conditioning for transplantation and enhance HSPC engraftment. PMID- 15284859 TI - Response to antiviral treatment in hepatitis C virus-associated marginal zone lymphomas. AB - A link between chronic hepatitis C virus (HCV) infection and low-grade B-cell lymphomas has been suggested by epidemiological studies. Marginal zone lymphomas (MZLs) including splenic lymphomas with villous lymphocytes are among the most frequently reported subgroups in the setting of chronic HCV infection. In this study, we examined the effect of antiviral treatment in eight patients with HCV associated MZL. We found that five out of eight patients have responded to interferon alpha and ribavirin. In some cases, hematologic responses were correlated to virologic responses. In addition, we report a case of large granular lymphocyte leukemia occurring in association with MZL and HCV, and responding to interferon and ribavirin. We suggest that there is an etiologic link between HCV and antigen-driven lymphoproliferative disorders. PMID- 15284860 TI - Mutational and expression analysis of the chromosome 12p candidate tumor suppressor genes in pre-B acute lymphoblastic leukemia. AB - Allelic losses on chromosome 12p12-13 are associated with childhood acute lymphoblastic leukemia (ALL) and several solid neoplasias, suggesting the presence of a tumor suppressor locus. The recent construction of a transcription map of this locus has enabled the identification of eight genes, of which five were previously known: ETV6, BCL-G, LRP6, MKP-7, and CDKN1B. The three other candidate genes, LOH12CR1, LOH12CR2, and LOH12CR3, have no known functions. To evaluate whether one (or more) of the candidate genes is the actual target of the 12p12-13 deletions, we examined the genomics and the expression status of these genes in ALL patients. Although we found nine DNA variants in these genes, no inactivating mutations were found in the leukemia cells of patients with 12p hemizygous deletions. Expression analysis revealed that most 12p hemizygously deleted samples also carried a t(12;21) translocation, of which none expressed ETV6 from the nontranslocated allele. Furthermore, we observed one case of t(12;21) without deletion of ETV6, in which the expression of this gene was greatly reduced, indicating a different mechanism of inactivation. None of the other genes showed a significant decrease in expression, suggesting that ETV6 is indeed the target of deletions in ALL patients. PMID- 15284861 TI - The use of housekeeping genes for real-time PCR-based quantification of fusion gene transcripts in acute myeloid leukemia. PMID- 15284862 TI - Myeloid antigen positive acute lymphoblastic leukemia with the Philadelphia translocation and a jumping translocation of 1q in a child. PMID- 15284863 TI - Screening for G-CSF receptor mutations in patients with secondary myeloid or lymphoid transformation of severe congenital neutropenia. A report from the French neutropenia register. PMID- 15284864 TI - Circulating myeloid and lymphoid precursor dendritic cells are clonally involved in myelodysplastic syndromes. AB - Circulating myeloid and lymphoid precursor dendritic cell (pDC) counts were determined in peripheral blood from 22 patients with myelodysplastic syndromes (MDS) by a single-platform flow cytometric protocol. The absolute count of myeloid and lymphoid pDC, as well as their relative number (as proportion of mononuclear cells or total leukocytes) was significantly lower in MDS (n=22) than in healthy controls (n=41). In 11 patients with chromosomal aberrations, purified pDC were examined by interphase fluorescence in situ hybridization. This revealed clonal involvement of myeloid as well as lymphoid pDC in all of them. These data therefore strongly suggest that myeloid and lymphoid pDC share a common precursor. Whether reduced peripheral blood counts of pDC contribute to the immunological abnormalities observed in MDS remains to be investigated. PMID- 15284865 TI - Validation of BIOMED-2 multiplex PCR tubes for detection of TCRB gene rearrangements in T-cell malignancies. AB - The BIOMED-2 Concerted Action BMH4-CT98-3936 on 'Polymerase chain reaction (PCR) based clonality studies for early diagnosis of lymphoproliferative disorders' developed standardized PCR protocols for detection of immunoglobulin (Ig) and T cell receptor (TCR) rearrangements, including TCR beta (TCRB). As no comparable TCRB PCR method pre-existed and only a limited number of samples was tested within the BIOMED-2 study, we initiated this study for further validation of the newly developed TCRB PCR approach by comparing PCR data with previously generated Southern blot (SB) data in a series of 66 immature (ALL) and 36 mature T-cell malignancies. In 91% of cases, concordant PCR and SB results were found. Discrepancies consisted of either failure to detect SB-detected TCRB rearrangements by PCR (6.5%) or detection of an additional non-SB defined rearrangement (2.5%). In 99% of cases (99/100), at least one clonal TCRB rearrangement was detected by PCR in the SB-positive cases. A predominance of complete Vbeta-Jbeta rearrangements was seen in TCRalphabeta(+) T-cell malignancies and CD3-negative T-ALL (100 and 90%, respectively), whereas in TCRgammadelta(+) T-ALL, more incomplete Dbeta-Jbeta TCRB rearrangements were detected (73%). Our results underline the reliability of this new TCRB PCR method and its strategic applicability in clonality diagnostics of lymphoproliferative disorders and MRD studies. PMID- 15284866 TI - Application of comparative genomics to narrow-leafed lupin (Lupinus angustifolius L.) using sequence information from soybean and Arabidopsis. AB - The completion of genome-sequencing initiatives for model plants and EST databases for major crop species provides a large resource for gaining fundamental knowledge of complex gene interactions and the functional significance of proteins. There are increasingly numerous opportunities to transfer this information to other plant species with uncharacterized genomes and make advances in genome analysis, gene expression, and predicted protein function. In this study, we have used DNA sequences from soybean and Arabidopsis to determine the feasibility of applying comparative genomics to narrow-leafed lupin. We have used transcribed sequences from soybean and showed that a high proportion cross hybridize to lupin DNA, identifying similar genes and providing landmarks for estimating the degree of chromosomal synteny between species. To further investigate comparative relationships in this study, a detailed analysis of three lupin genes and comparison of orthologs from soybean and Arabidopsis shows that, in some cases, gene structure and expression are highly conserved and their proteins may have similar function. In other cases, genes show variation in expression profiles indicating alternative functions across species. The advantages and limitation of using soybean and Arabidopsis sequences for comparative genomics in lupins are discussed. PMID- 15284867 TI - Assessment of genetic variability of haploids extracted from tetraploid (2n = 4x = 48) Solanum tuberosum. AB - The objectives of this study were to assess the genetic variability of haploids (2n = 2x = 24) extracted from tetraploid Solanum tuberosum through 4x x 2x crosses with Solanum phureja. Molecular and phenotypic analyses were performed to fingerprint the genotypes used and to evaluate their potential use in breeding programs. AFLP analysis revealed the presence of specific bands derived from the tetraploid seed parent S. phureja, as well as ex novo originated bands. On average, 210 bands were visualized per genotype, 149 (70%) of which were common to both parental genotypes. The percentage of S. tuberosum specific bands ranged from 25.1% to 18.6%, with an average of 22%. The fraction of genome coming from S. phureja ranged from 1.9% to 6.5%, with an average value of 4%. The percentage of ex novo bands varied from 1.9% to 9.0%. The presence of S. phureja DNA is very interesting because it indicated that S. phureja pollinator is involved in the mechanism of haploid formation. The characterization for resistance to Erwinia carotovora subsp. carotovora and potato virus X (PVX) provided evidence that haploids may express traits that are lacking in the tetraploids they come from, which can be useful for both genetic studies and breeding purposes. It is noteworthy that genotypes combining resistance to both diseases and good pollen stainability were identified. Other possible breeding implications owing to the presence of S. phureja genome in the haploids analyzed are discussed. PMID- 15284868 TI - Gene expression profiling of the bovine gastrointestinal tract. AB - Basal gene expression levels across the bovine gastrointestinal tract (GI) were examined in an attempt to formulate genetic explanations for the differences in function that are known or thought to exist between the various regions. Gene expression along the tract was studied through the random sequencing of a total of 16 412 clones from seven tissue-specific cDNA libraries spanning its length. The expressed sequence tags (ESTs) within each library were clustered to reduce clone redundancy and obtain longer consensus sequences. BLASTN and BLASTX searches against the NCBI human RefSeq databases were used to find putative matches for the bovine sequences and gene ontology assignments were made. Notable similarities and differences in gene expression were observed among the various compartments of the GI tract of the bovine. Many of the prominent transcripts have yet to be reliably identified and the prominence of others may be worthy of further examination. This collection of ESTs represents an important resource for the future construction of a GI tract specific microarray for further gene expression studies. PMID- 15284869 TI - Isolation of a family of resistance gene analogue sequences of the nucleotide binding site (NBS) type from Lens species. AB - Most known plant disease-resistance genes (R genes) include in their encoded products domains such as a nucleotide-binding site (NBS) or leucine-rich repeats (LRRs). Sequences with unknown function, but encoding these conserved domains, have been defined as resistance gene analogues (RGAs). The conserved motifs within plant NBS domains make it possible to use degenerate primers and PCR to isolate RGAs. We used degenerate primers deduced from conserved motifs in the NBS domain of NBS-LRR resistance proteins to amplify genomic sequences from Lens species. Fragments from approximately 500-850 bp were obtained. The nucleotide sequence analysis of these fragments revealed 32 different RGA sequences in Lens species with a high similarity (up to 91%) to RGAs from other plants. The predicted amino acid sequences showed that lentil sequences contain all the conserved motifs (P-loop, kinase-2, kinase-3a, GLPL, and MHD) present in the majority of other known plant NBS-LRR resistance genes. Phylogenetic analyses grouped the Lens NBS sequences with the Toll and interleukin-1 receptor (TIR) subclass of NBS-LRR genes, as well as with RGA sequences isolated from other legume species. Using inverse PCR on one putative RGA of lentil, we were able to amplify the flanking regions of this sequence, which contained features found in R proteins. PMID- 15284870 TI - Conversion of AFLP markers associated with FHB resistance in wheat into STS markers with an extension-AFLP method. AB - Amplified fragment length polymorphism (AFLP) has proven a powerful tool for tagging genes or quantitative trait loci (QTLs) of interest in plants. However, conversion of AFLP markers into sequence-tagged site (STS) markers is technically challenging in wheat owing to the complicated nature of its genome. In this study, we developed an "extension-AFLP" method to convert AFLP markers associated with Fusarium head blight (FHB) resistance into STS markers. When an AFLP marker of interest was detected with an EcoRI+3-MseI+4-selective primer combination, the PCR product was used as a template for an additional selective amplification with four primer pairs, in which one additional selective base (either A, C, G, or T) was added to the 3' end of one of the two primers. The extended primer pair that produced the targeted band was further extended by adding each of the four selective nucleotide bases for the next round of selective amplification. Extension selective amplification was performed until the target bands became clear enough for subsequent cloning and sequencing. By using the extension-AFLP method, we successfully converted two AFLP markers located on chromosome 3BS and associated with FHB resistance into STS markers. Our results indicated that the extension-AFLP method is an efficient approach for converting AFLP markers into STS markers in wheat. The developed STS markers might be used for marker-assisted selection (MAS) for FHB resistance in wheat breeding programs. PMID- 15284871 TI - Comparative analysis of a Brassica BAC clone containing several major aliphatic glucosinolate genes with its corresponding Arabidopsis sequence. AB - We compared the sequence of a 101-kb-long bacterial artificial chromosome (BAC) clone (B21H13) from Brassica oleracea with its homologous region in Arabidopsis thaliana. This clone contains a gene family involved in the synthesis of aliphatic glucosinolates. The A. thaliana homologs for this gene family are located on chromosome IV and correspond to three 2-oxoglutarate-dependent dioxygenase (AOP) genes. We found that B21H13 harbors 23 genes, whereas the equivalent region in Arabidopsis contains 37 genes. All 23 common genes have the same order and orientation in both Brassica and Arabidopsis. The 16 missing genes in the broccoli BAC clone were arranged in two major blocks of 5 and 7 contiguous genes, two singletons, and a twosome. The 118 exons comprising these 23 genes have high conservation between the two species. The arrangement of the AOP gene family in A. thaliana is as follows: AOP3 (GS-OHP) - AOP2 (GS-ALK) - pseudogene - AOP1. In contrast, in B. oleracea (broccoli and collard), two of the genes are duplicated and the third, AOP3, is missing. The remaining genes are arranged as follows: Bo-AOP2.1 (BoGSL-ALKa) - pseudogene - AOP2.2 (BoGSL-ALKb) - AOP1.1 - AOP1.2. When the survey was expanded to other Brassica accessions, we found variation in copy number and sequence for the Brassica AOP2 homologs. This study confirms that extensive rearrangements have taken place during the evolution of the Brassicacea at both gene and chromosomal levels. PMID- 15284872 TI - Regeneration of somatic hybrids in relation to the nuclear and cytoplasmic genomes of wheat and Setaria italica. AB - Somatic hybridization via PEG (Polyethylene 6000)-mediated protoplast fusion was achieved between two different wheat culture lines (Triticum aestivum L., "Jinan"177, T1 and T2) and Setaria italica (L.) P. Beauv. The T1 recipient originated from non-regenerable long-term cell suspensions, while T2 was derived from embryogenic calli with a high regeneration capacity. Donor protoplasts were obtained from embryogenic calli of S. italica (S) (with low regeneration capacity) irradiated with different doses of ultraviolet light. Twenty-three putative hybrid cell lines were produced in fusion combinations with the donor protoplasts treated with UV light for 30 s (combination I) and 1 min (combination II), but only one (from combination II) differentiated into green plants. Three cell lines from combination I and five cell lines from combination II possessed the nuclear genomes of T1, T2, and S. italica as revealed by cytological, isozyme, RAPD, and 5S rDNA spacer sequence analyses. Genomic in situ hybridization (GISH) analysis showed that most hybrid cell lines had 22-36 wheat chromosomes, 0-2 S. italica chromosomes, and 1-6 wheat - S. italica recombinant chromosomes, whereas the regenerable cell line had 44-56 wheat chromosomes and 3 6 recombinant chromosomes, but no intact S. italica chromosomes. RFLP analysis of organellar DNA revealed that mitochondrial and chloroplast DNA of both parents coexisted in all hybrid cell lines and recombined in most hybrid cell lines. These results indicate that the regeneration of hybrid plants involves not only the integration of S. italica nuclear and organellar DNA, but also the genome complementation of T1 and T2. PMID- 15284873 TI - AFLP analysis of Cynodon dactylon (L.) Pers. var. dactylon genetic variation. AB - Cynodon dactylon (L.) Pers. var. dactylon (common bermudagrass) is geographically widely distributed between about lat 45 degrees N and lat 45 degrees S, penetrating to about lat 53 degrees N in Europe. The extensive variation of morphological and adaptive characteristics of the taxon is substantially documented, but information is lacking on DNA molecular variation in geographically disparate forms. Accordingly, this study was conducted to assess molecular genetic variation and genetic relatedness among 28 C. dactylon var. dactylon accessions originating from 11 countries on 4 continents (Africa, Asia, Australia, and Europe). A fluorescence-labeled amplified fragment length polymorphism (AFLP) DNA profiling method was used to detect the genetic diversity and relatedness. On the basis of 443 polymorphic AFLP fragments from 8 primer combinations, the accessions were grouped into clusters and subclusters associating with their geographic origins. Genetic similarity coefficients (SC) for the 28 accessions ranged from 0.53 to 0.98. Accessions originating from Africa, Australia, Asia, and Europe formed major groupings as indicated by cluster and principal coordinate analysis. Accessions from Australia and Asia, though separately clustered, were relatively closely related and most distantly related to accessions of European origin. African accessions formed two distant clusters and had the greatest variation in genetic relatedness relative to accessions from other geographic regions. Sampling the full extent of genetic variation in C. dactylon var. dactylon would require extensive germplasm collection in the major geographic regions of its distributional range. PMID- 15284874 TI - QTL detection for rice grain quality traits using an interspecific backcross population derived from cultivated Asian (O. sativa L.) and African (O. glaberrima S.) rice. AB - An interspecific advanced backcross population derived from a cross between Oryza sativa "V20A" (a popular male-sterile line used in Chinese rice hybrids) and Oryza glaberrima (accession IRGC No. 103544 from Mali) was used to identify quantitative trait loci (QTL) associated with grain quality and grain morphology. A total of 308 BC3F1 hybrid families were evaluated for 16 grain-related traits under field conditions in Changsha, China, and the same families were evaluated for RFLP and SSR marker segregation at Cornell University (Ithaca, N.Y.). Eleven QTL associated with seven traits were detected in six chromosomal regions, with the favorable allele coming from O. glaberrima at eight loci. Favorable O. glaberrima alleles were associated with improvements in grain shape and appearance, resulting in an increase in kernel length, transgressive variation for thinner grains, and increased length to width ratio. Oryza glaberrima alleles at other loci were associated with potential improvements in crude protein content and brown rice yield. These results suggested that genes from O. glaberrima may be useful in improving specific grain quality characteristics in high-yielding O. sativa hybrid cultivars. PMID- 15284875 TI - Colocation between a gene encoding the bZip factor SPA and an eQTL for a high molecular-weight glutenin subunit in wheat (Triticum aestivum). AB - The quality of wheat grain is largely determined by the quantity and composition of storage proteins (prolamins) and depends on mechanisms underlying the regulation of expression of prolamin genes. The endosperm-specific wheat basic region leucine zipper (bZIP) factor storage protein activator (SPA) is a positive regulator that binds to the promoter of a prolamin gene. The aim of this study was to map SPA (the gene encoding bZIP factor SPA) and genomic regions associated with quantitative variations of storage protein fractions using F7 recombinant inbred lines (RILs) derived from a cross between Triticum aestivum "Renan" and T. aestivum "Recital". SPA was mapped through RFLP using a cDNA probe and a specific single nucleotide polymorphism (SNP) marker. Storage protein fractions in the parents and RILs were quantified using capillary electrophoresis. Quantitative trait loci (QTLs) for protein were detected and mapped on six chromosome regions. One QTL, located on the long arm of chromosome 1B, explained 70% of the variation in quantity of the x subunit of Glu-B1. Genetic mapping suggested that SPA is located on chromosome arm 1L and is also present in the confidence interval of the corresponding QTL for Glu-B1x on 1BL, suggesting that SPA might be a candidate gene for this QTL. PMID- 15284876 TI - Evolution of duplicated growth hormone genes in autotetraploid salmonid fishes. AB - A defining character of the piscine family Salmonidae is autotetraploidy resulting from a genome-doubling event some 25-100 million years ago. Initially, duplicated genes may have undergone concerted evolution and tetrasomic inheritance. Homeologous chromosomes eventually diverged and the resulting reduction in recombination and gene conversion between paralogous genes allowed the re-establishment of disomic inheritance. Among extant salmonine fishes (e.g. salmon, trout, char) the growth hormone (GH) gene is generally represented by two functional paralogs, GH1 and GH2. Sequence analyses of salmonid GH genes from species of subfamilies Coregoninae (whitefish, ciscos) and Salmoninae were used to examine the evolutionary history of the duplicated GH genes. Two divergent GH gene paralogs were also identified in Coregoninae, but they were not assignable to the GH1 and GH2 categories. The average sequence divergence between the coregonine GH genes was more than twofold lower than the corresponding divergence between the salmonine GH1 and GH2. Phylogenetic analysis of the coregonine GH paralogs did not resolve their relationship to the salmonine paralogs. These findings suggest that disomic inheritance of two GH genes was established by different mechanisms in these two subfamilies. PMID- 15284877 TI - Cytogenetic characterization and fae1 gene variation in progenies from asymmetric somatic hybrids between Brassica napus and Crambe abyssinica. AB - Sexual progenies of asymmetric somatic hybrids between Brassica napus and Crambe abyssinica were analyzed with respect to chromosomal behavior, fae1 gene introgression, fertility, and fatty-acid composition of the seed. Among 24 progeny plants investigated, 11 plants had 38 chromosomes and were characterized by the occurrence of normal meiosis with 19 bivalents. The other 13 plants had more than 38 chromosomes, constituting a complete chromosomal set from B. napus plus different numbers of additional chromosomes from C. abyssinica. The chromosomes of B. napus and C. abyssinica origin could be clearly discriminated by genomic in situ hybridization (GISH) in mitotic and meiotic cells. Furthermore, meiotic GISH enabled identification of intergenomic chromatin bridges and of asynchrony between the B. napus and C. abyssinca meiotic cycles. Lagging, bridging and late disjunction of univalents derived from C. abyssinica were observed. Analysis of cleaved amplified polymorphic sequence (CAPS) markers derived from the fae1 gene showed novel patterns different from the B. napus recipient in some hybrid offspring. Most of the progeny plants had a high pollen fertility and seed set, and some contained significantly greater amounts of seed erucic acid than the B. napus parent. This study demonstrates that a part of the C. abyssinica genome can be transferred into B. napus via asymmetric hybridization and maintained in sexual progenies of the hybrids. Furthermore, it confirms that UV irradiation improves the fertility of the hybrid and of its sexual progeny via chromosomal elimination and facilitates the introgression of exotic genetic material into crop species. PMID- 15284878 TI - Molecular evolution of homologous gene sequences in germline-limited and somatic chromosomes of Acricotopus. AB - The origin of germline-limited chromosomes (Ks) as descendants of somatic chromosomes (Ss) and their structural evolution was recently elucidated in the chironomid Acricotopus. The Ks consist of large S-homologous sections and of heterochromatic segments containing germline-specific, highly repetitive DNA sequences. Less is known about the molecular evolution and features of the sequences in the S-homologous K sections. More information about this was received by comparing homologous gene sequences of Ks and Ss. Genes for 5.8S, 18S, 28S, and 5S ribosomal RNA were choosen for the comparison and therefore isolated first by PCR from somatic DNA of Acricotopus and sequenced. Specific K DNA was collected by microdissection of monopolar moving K complements from differential gonial mitoses and was then amplified by degenerate oligonucleotide primer (DOP)-PCR. With the sequence data of the somatic rDNAs, the homologous 5.8S and 5S rDNA sequences were isolated by PCR from the DOP-PCR sequence pool of the Ks. In addition, a number of K DOP-PCR sequences were directly cloned and analysed. One K clone contained a section of a putative N-acetyltransferase gene. Compared with its homolog from the Ss, the sequence exhibited few nucleotide substitutions (99.2% sequence identity). The same was true for the 5.8S and 5S sequences from Ss and Ks (97.5%-100% identity). This supports the idea that the S homologous K sequences may be conserved and do not evolve independently from their somatic homologs. Possible mechanisms effecting such conservation of S derived sequences in the Ks are discussed. PMID- 15284879 TI - Karyotype characterization of the lake sturgeon, Acipenser fulvescens (Rafinesque 1817) by chromosome banding and fluorescent in situ hybridization. AB - A karyotype analysis using several staining techniques was carried out on the North American lake sturgeon, Acipenser fulvescens. The chromosome number was found to be 2n = 262 +/- 6. A representative karyotype of 264 chromosomes was composed of 134 meta- and submetacentrics, 70 telo- and acrocentrics, and 60 microchromosomes. The constitutive heterochromatin, revealed by C banding, was localized in various positions on several chromosomes, including microchromosomes. The signals of fluorescent in situ hybridization (FISH) with a HindIII satellite DNA probe were visible as centromeric heterochromatin blocks on 48 chromosomes. The telomeric repeat (TTAGGG)n detected by FISH was localized at both ends of all chromosomes and two chromosomes were entirely marked. Fluorescent staining with GC-specific chromomycin A3 showed recognizable fluorescent regions, whereas a more uniform base composition was revealed by the AT-specific 4',6-diamidino-2-phenylindole (DAPI). After silver staining, the active nucleolar organizer regions (NORs) were detected on 12 chromosomes. FISH with the 5S probe showed four signals on four small chromosomes. Our data suggest that A. fulvescens is a tetraploid species. PMID- 15284880 TI - A direct repeat sequence associated with the centromeric retrotransposons in wheat. AB - A high-density BAC filter of Triticum monococcum was screened for the presence of a centromeric retrotransposon using the integrase region as a probe. Southern hybridization to the BAC digests using total genomic DNA probes of Triticum monococcum, Triticum aestivum, and Hordeum vulgare detected differentially hybridizing restriction fragments between wheat and barley. The fragments that hybridized to genomic DNA of wheat but not to that of barley were subcloned. Fluorescence in situ hybridization (FISH) analysis indicated that the clone pHind258 hybridized strongly to centromeric regions in wheat and rye and weakly to those in barley. The sequence of pHind258 was homologous to integrase and long terminal repeats of centromeric Ty3-gypsy retrotransposons of cereal species. Additionally, pHind258 has a pair of 192-bp direct repeats. FISH analysis indicated that the 192-bp repeat probe hybridized to centromeres of wheat and rye but not to those of barley. We found differential FISH signal intensities among wheat chromosomes using the 192-bp probe. In general, the A-genome chromosomes possess strong FISH signals, the B-genome chromosomes possess moderate signals, and the D-genome chromosomes possess weak signals. This was consistent with the estimated copy numbers of the 192-bp repeats in the ancestral species of hexaploid wheat. PMID- 15284881 TI - Telomeric and interstitial telomeric-like DNA sequences in Orthoptera genomes. AB - A (TTAGG)n-specific telomeric DNA probe was hybridized to 11 orthopteroid insect genomes by fluorescence in situ hybridization. Nine different genera, mainly distributed within two evolutionary branches with male chromosome numbers 2n = 23 and 2n = 17 were included in the analysis. Telomere sequences yielded positive signals in every telomere and there was a considerable number of interstitial telomeric-like sequences, mainly located at the distal end of some, but not all, subterminal chromosome regions. One of the species, Pyrgomorpha conica, showed massive hybridization signals associated with constitutive heterochromatin. The results are discussed along two lines: (i) the chromosomal evolutionary trends within this group of insects and (ii) the putative role that ITs may play in a genome when they are considered telomere-derived, but not telomere-functional, DNA sequences. PMID- 15284882 TI - Segmental duplications within the Glycine max genome revealed by fluorescence in situ hybridization of bacterial artificial chromosomes. AB - Soybean (Glycine max L. Merr.) is presumed to be an ancient polyploid based on chromosome number and multiple RFLP fragments in genetic mapping. Direct cytogenetic observation of duplicated regions within the soybean genome has not heretofore been reported. Employing fluorescence in situ hybridization (FISH) of genetically anchored bacterial artificial chromosomes (BACs) in soybean, we were able to observe that the distal ends of molecular linkage group E had duplicated regions on linkage groups A2 and B2. Further, using fiber-FISH, it was possible to measure the molecular size and organization of one of the duplicated regions. As FISH did not require repetitive DNA for blocking fluorescence signals, we assume that the 200-kb genome region is relatively low in repetitive sequences. This observation, along with the observation that the BACs are located in distal euchromatin regions, has implications for genome structure/evolution and the approach used to sequence the soybean genome. PMID- 15284883 TI - Cloning and characterization of a transposable-like repeat in the heterochromatin of the darkling beetle Misolampus goudoti. AB - A long repeat unit of the PstI family in Misolampus goudoti (Coleoptera, Tenebrionodae) is characterized in this work. The 30 sequenced units have small differences in length (consensus 1169 bp), but very similar nucleotide composition (mean 61.1% A+T). PstI repeats contain a 36-bp-long inverted repeat at both the 5' and 3' ends, with a fully conserved 16-bp-long motif similar to those found in class II transposable elements. However, the transposable-like PstI repeats seems to be defective, since they do not encode for any protein related with transposition. Interestingly, energetically stable hairpins resembled the structure of a miniature interspersed transposable element, suggesting that the PstI satellite DNA family in M. goudoti may have originated from an ancestral active transposable element as also described in Drosophila guanche. The presence of transposable-like structure along with the non-detection of gene conversion or unequal crossing-over events suggest that transposition could be one of the putative molecular mechanisms involved in the strong amplification and (or) homogenization of these repeats. A putative transposition of PstI repeats allowing their genomic mobility also could explain why this satellite is widely distributed to all heterochromatic regions, telomeres, pericentromeric regions, and on the Y chromosome, whereas satellites of other tenebrionids lacking transposable-like structures are restricted only to pericentromeric regions. PMID- 15284884 TI - [The genome of alpha-proteobacteria : complexity, reduction, diversity and fluidity]. AB - The alpha-proteobacteria displayed diverse and often unconventional life-styles. In particular, they keep close relationships with the eucaryotic cell. Their genomic organization is often atypical. Indeed, complex genomes, with two or more chromosomes that could be linear and sometimes associated with plasmids larger than one megabase, have been described. Moreover, polymorphism in genome size and topology as well as in replicon number was observed among very related bacteria, even in a same species. Alpha-proteobacteria provide a good model to study the reductive evolution, the role and origin of multiple chromosomes, and the genomic fluidity. The amount of new data harvested in the last decade should lead us to better understand emergence of bacterial life-styles and to build the conceptual basis to improve the definition of the bacterial species. PMID- 15284885 TI - Population patterns and antimicrobial resistance of Aeromonas in urban playa lakes. AB - Bacteria belonging to the genus Aeromonas are indigenous to aquatic environments. Once regarded as unimportant human pathogens, reports of opportunistic infections caused by these organisms have appeared increasingly in the medical literature. To estimate the potential for human infection by Aeromonas where limited water resources are being used intensively, we studied the spatial and temporal variation and incidence of antimicrobial resistance among environmental isolates of Aeromonas from two urban playa lakes in Lubbock, Texas. Aeromonas population densities varied seasonally, with the highest densities occurring from mid-April to late October. The greatest range of densities was 100-fold, from 2.50 to 255.17 colony-forming units per 0.1 mL of water sample. Densities also varied with water depth, although the variation did not display a consistent pattern. One hundred fifty-one Aeromonas isolates were divided into 10 species or subspecies groups by using the BIOLOG identification system. Nine isolates displayed resistance to co-trimoxazole, tetracycline, and cefuroxime, and none was resistant to more than one of these antimicrobial agents. In summary, the results of this study showed that the densities of Aeromonas peak in the late spring and again in late summer, times when human activity around the playa lakes is also high. Thus, we infer that human exposure to these potential pathogens varies seasonally. Compared to other published studies, the incidence of antimicrobial-resistant Aeromonas is relatively low in urban playa lakes in Lubbock, Texas. Nevertheless, resistant organisms were detected. PMID- 15284886 TI - Assessment of intra-species diversity among strains of Acinetobacter baumannii isolated from sites contaminated with petroleum hydrocarbons. AB - A total of 96 crude oil-degrading bacterial strains were isolated from 5 geographically diverse sites in India that were contaminated with different types of petroleum hydrocarbons. The strains were identified by sequencing the genes that encode for 16S rRNA. Out of the 96 isolates, 25 strains were identified as Acinetobacter baumannii and selected for the study. All of the selected strains could degrade the total petroleum hydrocarbon fractions of crude oil. These 25 strains were biochemically profiled and grouped into 8 phenovars on the basis of multivariate analysis of their substrate utilization profiles. PCR-based DNA fingerprinting was performed using intergenic repetitive DNA sequences, which divided the selected 25 strains into 7 specific genomic clusters. tRNA intergenic spacer length polymorphism was performed to determine the intra-species relatedness among these 25 strains. It delineated the strains into 8 genomic groups. The present study detected specific variants among the A. baumannii strains with differential degradation capacities for different fractions of crude oil. This could play a significant role in in situ bioremediation. The study also revealed the impact of environmental factors that cause intra-species diversity within the selected strains of A. baumannii. PMID- 15284887 TI - The fate of a genetically modified Pseudomonas strain and its transgene during the composting of poultry manure. AB - The fate of the genetically modified (GM) Pseudomonas chlororaphis strain 3732 RN L11 and its transgene (lacZ insert) during composting of chicken manure was studied using plate count and nested polymerase chain reaction (PCR) methods. The detection sensitivity of the nested PCR method was 165 copies of the modified gene per gram of moist compost or soil. Compost microcosms consisted of a 100-g mixture of chicken manure and peat, whereas soil microcosms were 100-g samples of sandy clay loam. Each microcosm was inoculated with 4 x 1010 CFU of P. chlororaphis RN-L11. In controlled temperature studies, neither P. chlororaphis RN-L11 nor its transgene could be detected in compost microcosms after incubation temperature was elevated to 45 degrees C or above for one or more days. In contrast, in the compost microcosms incubated at 23 degrees C, the target organism was not detected by the plate count method after 6 days, but its transgene was detectable for at least 45 days. In compost bins, the target organism was not recovered from compost microcosms or soil microcosms at different levels in the bins for 29 days. However, the transgene was detected in 8 of the 9 soil microcosms and in only 1 of the 9 compost microcosms. The compost microcosm in which transgene was detected was at the lower level of the bin where temperatures remained below 45 degrees C. The findings indicated that composting of organic wastes could be used to reduce or degrade heat sensitive GM microorganisms and their transgenes. PMID- 15284888 TI - Use of the surface proteins GapC and Mig of Streptococcus dysgalactiae as potential protective antigens against bovine mastitis. AB - Streptococcus dysgalactiae is a significant pathogen associated with bovine mastitis in lactating and nonlactating dairy cows, causing a severe inflammatory response of the mammary gland, which results in major economic losses to the dairy industry. Two proteins from S. dysgalactiae strain SDG8 were tested for their protective capacity against a homologous bacterial challenge in a dry cow model. The first was a bovine plasmin receptor protein (GapC), which shares 99.4% sequence identity to the plasmin-binding Plr protein of group A streptococci. The second protein product was Mig, a alpha2-M-, IgG-, and IgA-binding protein present on the cell surface of SDG8. We investigated the efficacy of immunization with purified recombinant forms of GapC and Mig by measuring the number of somatic cells and assessing the presence of the challenge strain in mammary secretions following challenge. In this model, we found that, although the number of quarters containing SDG8 was significantly reduced in the GapC- but not in the Mig-immunized animals, the somatic cell counts from teat secretions were significantly decreased in both the GapC and Mig vaccinates. PMID- 15284889 TI - Genetic analysis of Canadian isolates of C:2a:P1.2,5 and B:2a:P1.2,5 Neisseria meningitidis strains belonging to the hypervirulent clone of ET-15. AB - Isolates of the hypervirulent Neisseria meningitidis clone ET-15 found to express the serogroup B antigen were investigated and compared with representative members of serogroup B and C isolates. Clonal-clustering methods clearly grouped the B:ET15 isolates with C:ET15 isolates, indicating the only major difference between the two groups was in the capsule expressed. The organization of the cps operon from the B:ET15 isolates was found to be consistent with typical serogroup B isolates and differed from serogroup C isolates only in the sialyl transferase gene present. This suggests that these strains arose via recombination of the sialyl transferase gene. Specific points of recombination could not be identified, however, the majority (64%) of the B:ET15 isolates contained a copy of pseudo-IS1106 downstream of the cps operon indicating the potential for a common ancestral origin. The combination of pulsed-field gel electrophoresis (PFGE) and sequence analysis of targeted regions of the cps operon permitted the differentiation of most B:ET15 isolates indicating that they likely arose from separate genetic events and do not represent the emergence and spread of a new clone. However, two isolates that appeared identical by all methods employed were temporally and geographically related although no epidemiological evidence is available indicating a link between these strains. PMID- 15284890 TI - Evaluation of the in vitro and in vivo dimorphism of Sporothrix schenckii, Blastomyces dermatitidis, and Paracoccidioides brasiliensis isolates after preservation in mineral oil. AB - Morphological differentiation has commanded attention for its putative impact on the pathogenesis of invasive fungal infections. We evaluated in vitro and in vivo the dimorphism from mycelial to yeast-phase of Sporothrix schenckii, Blastomyces dermatitidis and Paracoccidioides brasiliensis isolates, two strains for each species, preserved in mineral oil. S. schenckii strains showed typical micromorphology at 25 degrees C but one strain was unable to complete the dimorphic process in vitro. After in vivo passage through mice the strains had the ability to turn into yeast-like cells and to form colonies on brain-heart infusion medium at 36 degrees C. B. dermatitidis strains grew as dirty white to brownish membranous colonies at 25 degrees C and their micromorphology showed thin filaments with single hyaline conidia. At 36 degrees C the colonies did not differ from those grown at 25 degrees C, but produced a transitional micromorphology. P. brasiliensis strains grew as cream-colored cerebriform colonies at 25 degrees C showing a transitional morphology. B. dermatitidis and P. brasiliensis strains did not turn into yeast-like cells in vivo. The present results demonstrate that B. dermatitidis and P. brasiliensis strains were unable to complete the dimorphic process even after in vivo passage, in contrast to the S. schenckii strain. PMID- 15284891 TI - Enhancement of the antifungal activity of Bacillus subtilis F29-3 by the chitinase encoded by Bacillus circulans chiA gene. AB - Bacillus subtilis F29-3 is an antagonistic bacterium against a wide range of fungal species. In order to determine the effect of chitinase on the antifungal activity of B. subtilis F29-3, a 2.4-kb DNA fragment containing the chiA gene of Bacillus circulans WL-12 was ligated into a shuttle vector pHY300PLK and transformed into B. subtilis F29-3. A bioassay conducted on the culture supernatant showed that, in comparison to the B. subtilis control strain, B. subtilis F29-3 expressing the chiA gene exhibited a greater inhibition of spore germination of Botrytis elliptica, indicating that chitinase could enhance the antifungal function conferred by B. subtilis F29-3. PMID- 15284892 TI - Pyrimidine nucleotide synthesis in Pseudomonas citronellolis. AB - Pyrimidine biosynthesis was active in Pseudomonas citronellolis ATCC 13674 and appeared to be regulated by pyrimidines. When wild-type cells were grown on succinate in the presence of uracil, the de novo enzyme activities were depressed while only four enzyme activities were depressed in the glucose-grown cells. On either carbon source, orotic acid-grown cells had diminished aspartate transcarbamoylase, dihydroorotase or OMP decarboxylase activity. Pyrimidine limitation of glucose-grown pyrimidine auxotrophic cells resulted in de novo enzyme activities, except for transcarbamoyolase activity, that were elevated by more than 5-fold compared to their activities in uracil-grown cells. Since pyrimidine limitation of succinate-grown mutant cells produced less enzyme derepression, catabolite repression appeared to be a factor. At the level of enzyme activity, aspartate transcarbamoylase activity in P. citronellolis was strongly inhibited by all effectors tested. Compared to the regulation of pyrimidine biosynthesis in taxonomically-related species, pyrimidine biosynthesis in P. citronellolis appeared more highly regulated. PMID- 15284893 TI - Molecular basis of transcriptional silencing in budding yeast. AB - Transcriptional silencing is a phenomenon in which the transcription of genes by RNA polymerase II or III is repressed, dependent on the chromosomal location of a gene. Transcriptional silencing normally occurs in highly condensed heterochromatin regions of the genome, suggesting that heterochromatin might repress transcription by restricting the ability of sequence-specific gene activator proteins to access their DNA target sites. However, recent studies show that heterochromatin structure is inherently dynamic, and that sequence-specific regulatory proteins are able to bind to their target sites in heterochromatin. The molecular basis of transcriptional silencing is plainly more complicated than simple steric exclusion. New ideas and experiments are needed. PMID- 15284894 TI - The FACT chromatin modulator: genetic and structure/function relationships. AB - The chromatin configuration of DNA inhibits access by enzymes such as RNA polymerase II. This inhibition is alleviated by FACT, a conserved transcription elongation factor that has been found to reconfigure nucleosomes to allow transit along the DNA by RNA polymerase II, thus facilitating transcription. FACT also reorganizes nucleosomes after the passage of RNA polymerase II, as indicated by the effects of certain FACT mutations. The larger of the two subunits of FACT is Spt16/Cdc68, while the smaller is termed SSRP1 (vertebrates) or Pob3 (budding yeast). The HMG-box domain at the C terminus of SSRP1 is absent from Pob3; the function of this domain for yeast FACT is supplied by the small HMG-box protein Nhp6. In yeast, this "detachable" HMG domain is a general chromatin component, unlike FACT, which is found only in transcribed regions and associated with RNA polymerase II. The several domains of the larger FACT subunit are also likely to have different functions. Genetic studies suggest that FACT mediates nucleosome reorganization along several pathways, and reinforce the notion that protein unfolding and (or) refolding is involved in FACT activity for transcription. PMID- 15284895 TI - Synthetic zinc finger peptides: old and novel applications. AB - In the last decade, the efforts in clarifying the interaction between zinc finger proteins and DNA targets strongly stimulated the creativity of scientists in the field of protein engineering. In particular, the versatility and the modularity of zinc finger (ZF) motives make these domains optimal building blocks for generating artificial zinc finger peptides (ZFPs). ZFPs can act as transcription modulators potentially able to control the expression of any desired gene, when fused to an appropriate effector domain. Artificial ZFPs open the possibility to re-program the expression of specific genes at will and can represent a powerful tool in basic science, biotechnology and gene therapy. In this review we will focus on old, novel and possible future applications of artificial ZFPs. PMID- 15284896 TI - Nucleoplasmin: a nuclear chaperone. AB - In this article, we briefly review the structural and functional information currently available on nucleoplasmin. Special emphasis is placed on the discussion of the molecular mechanism involved in the sperm chromatin remodelling activity of this protein. A model is proposed based on current crystallographic data, recent biophysical and functional studies, as well as in the previously available information. PMID- 15284897 TI - Histone modifications and DNA double-strand break repair. AB - The roles of different histone modifications have been explored extensively in a number of nuclear processes, particularly in transcriptional regulation. Only recently has the role of histone modification in signaling or facilitating DNA repair begun to be elucidated. DNA broken along both strands in the same region, a double-strand break, is damaged in the most severe way possible and can be the most difficult type of damage to repair accurately. To successfully repair the double-strand break, the cell must gain access to the damaged ends of the DNA and recruit repair factors, and in the case of homologous recombination repair, the cell must also find, colocalize, and gain access to a suitable homologous sequence. In the repair of a double-strand break, the cell must also choose between homologous and non-homologous pathways of repair. Here, we will briefly review the mechanisms of double-strand-break repair, and discuss the known roles of histone modifications in signaling and repairing double-strand breaks. PMID- 15284898 TI - Activation domains of gene-specific transcription factors: are histones among their targets? AB - Activation domains of promoter-specific transcription factors are critical entities involved in recruitment of multiple protein complexes to gene promoters. The activation domains often retain functionality when transferred between very diverse eukaryotic phyla, yet the amino acid sequences of activation domains do not bear any specific consensus or secondary structure. Activation domains function in the context of chromatin structure and are critical for chromatin remodeling, which is associated with transcription initiation. The mechanisms of direct and indirect recruitment of chromatin-remodeling and histone-modifying complexes, including mechanisms involving direct interactions between activation domains and histones, are discussed. PMID- 15284899 TI - Gene regulation by Sp1 and Sp3. AB - The Sp family of transcription factors is united by a particular combination of three conserved Cys2His2 zinc fingers that form the sequence-specific DNA-binding domain. Within the Sp family of transcription factors, Sp1 and Sp3 are ubiquitously expressed in mammalian cells. They can bind and act through GC boxes to regulate gene expression of multiple target genes. Although Sp1 and Sp3 have similar structures and high homology in their DNA binding domains, in vitro and in vivo studies reveal that these transcription factors have strikingly different functions. Sp1 and Sp3 are able to enhance or repress promoter activity. Regulation of the transcriptional activity of Sp1 and Sp3 occurs largely at the post-translational level. In this review, we focus on the roles of Sp1 and Sp3 in the regulation of gene expression. PMID- 15284900 TI - The long hand of the small RNAs reaches into several levels of gene regulation. AB - Small RNA molecules such as siRNAs and miRNAs represent a new class of molecules that have been implicated in a wide range of diverse gene silencing phenomena. It is now becoming clear that these two similar molecules share several common features in both their biogenesis and their mechanism of action. Thus, the siRNA and miRNA pathways may have evolved from a common ancestral mechanism that has diverged to play important roles in developmental regulation, genomic organisation, and cellular defence against foreign nucleic acids. PMID- 15284901 TI - Functional diversity of ISWI complexes. AB - The yeast SWI/SNF ATP-dependent chromatin remodeling complex was first identified and characterized over 10 years ago (F. Winston and M. Carlson. 1992. Trends Genet. 8: 387-391.) Since then, the number of distinct ATP-dependent chromatin remodeling complexes and the variety of roles they play in nuclear processes have become dizzying (J.A. Martens and F. Winston. 2003. Curr. Opin. Genet. Dev. 13: 136-142; A. Vacquero et al. 2003. Sci. Aging Knowledge Environ. 2003: RE4)--and that does not even include the companion suite of histone modifying enzymes, which exhibit a comparable diversity in both number of complexes and variety of functions (M.J. Carrozza et al. 2003. Trends Genet. 19: 321-329; W. Fischle et al. 2003. Curr. Opin. Cell Biol. 15: 172-183; M. Iizuka and M.M. Smith. 2003. Curr. Opin. Genet. Dev. 13: 1529-1539). This vast complexity is hardly surprising, given that all nuclear processes that involve DNA--transcription, replication, repair, recombination, sister chromatid cohesion, etc.--must all occur in the context of chromatin. The SWI/SNF-related ATP-dependent remodelers are divided into a number of subfamilies, all related by the SWI2/SNF2 ATPase at their catalytic core. In nearly every species where researchers have looked for them, one or more members of each subfamily have been identified. Even the budding yeast, with its comparatively small genome, contains eight different chromatin remodelers in five different subfamilies. This review will focus on just one subfamily, the Imitation Switch (ISWI) family, which is proving to be one of the most diverse groups of chromatin remodelers in both form and function. PMID- 15284902 TI - New twists on H2A.Z: a histone variant with a controversial structural and functional past. AB - Integration of histone variants into chromatin organization allows for functional specification of chromatin regions. Recent functional analyses of H2A.Z have ascribed to it a multiplicity of complex and often opposing roles in developmental and homeostatic regulation. However, although the manner in which this essential histone variant is able to mediate its effects is not entirely well understood, current work has sought to investigate its mode of action. It is becoming increasingly clear that H2A.Z does not necessarily act independently, but rather, in conjunction with trans-acting factors to elicit chromatin changes. The nature of these structural changes has remained somewhat controversial but nevertheless exemplifies the seemingly multifaceted nature of H2A.Z. PMID- 15284903 TI - Telomeres, telomerase, and apoptosis. AB - Telomeres are specialized high-order chromatin structures that cap the ends of eukaryotic chromosomes. In vertebrates, telomeric DNA is composed of repetitions of the TTAGGG hexanucleotide, is bound to a set of specific proteins, and is elongated by the reverse transcriptase enzyme telomerase. Telomerase activity is promptly detected in cells with an indefinite replicative potential, such as cancer cells, while is almost undetectable in normal cells, which are characterized by a limited life span. Mounting evidence indicates that the maintenance of telomere integrity and telomerase protect cells from apoptosis. Disruption of the telomere capping function and (or) telomerase inhibition elicit an apoptotic response in cancer cells, while restoration of telomerase activity in somatic cells confers resistance to apoptosis. The possible mechanisms linking telomeres, telomerase and apoptosis are discussed in this review, together with the impact of this field in anticancer research. PMID- 15284904 TI - S100 proteins and their influence on pro-survival pathways in cancer. AB - The S100 gene family is composed of at least 20 members that share a common structure defined in part by the Ca2+ binding EF-hand motif. These genes which are expressed in a discriminate fashion in specific cells and tissues, have been described to have either an intracellular or extracellular function, or both. S100 proteins are implicated in the immune response, differentiation, cytoskeleton dynamics, enzyme activity, Ca2+ homeostasis and growth. A potential role for S100 proteins in neoplasia stems from these activities and from the observation that several S100 proteins have altered levels of expression in different stages and types of cancer. While the precise role and importance of S100 proteins in the development and promotion of cancer is poorly understood, it appears that the binding of Ca2+ is essential for exposing amino acid residues that are important in forming protein-protein interactions with effector molecules. The identity of some of these effector molecules has also now begun to emerge, and with this the elucidation of the signaling pathways that are modulated by these proteins. Some of these interactions are consistent with the diverse functions noted above. Others suggest that, many S100s may also promote cancer progression through specific roles in cell survival and apoptosis pathways. This review summarizes these findings and their implications. PMID- 15284906 TI - Nanomedicine -- a tremendous research opportunity for analytical chemists. PMID- 15284907 TI - Ultrasound: promoting electroanalysis in difficult real world media. AB - This article outlines the recent progress in the field of sonoelectroanalysis with strong emphasis on 'real world media' analysis. General principles of stripping analysis and the effects of ultrasound are explained. A section on 'Electroanalysis in extreme media' presents several examples of applications including detection of copper in beer, manganese in tea, lead and cadmium in saliva. The benefits of diamond electrodes are described and in the final section 'Metal ion detection in blood' several novel approaches based on the use of power ultrasound and based on bismuth electrodes are discussed. Specifically it is shown that the combination of ultrasound with classical stripping voltammetry permits quantifiable measurements in media hitherto impossible to study using conventional methods. PMID- 15284908 TI - Elizabeth a. H. Hall, University of Cambridge. PMID- 15284909 TI - Three-dimensional solution NMR spectroscopy of complex structures and mixtures. AB - This paper reviews the non-biological applications of three dimensional NMR (3D NMR) spectroscopy methodologies for studying chemical structures in polymer science, dendrimer research, organometallic chemistry, organosilicon chemistry, and mixtures of small organic molecules. Four methodologies for solving chemical structure problems are described, where the appropriate method is determined by the presence or absence of a third X nucleus (in addition to (1)H and (13)C) with suitable NMR properties. PMID- 15284910 TI - Low-pressure gradient micro-ion chromatography with ultra-short monolithic anion exchange column. AB - A 1.0 x 0.4 cm silica based monolithic anion exchanger has been produced and evaluated for use within a miniature gradient suppressed ion chromatography system based upon low-pressure micro-scale peristaltic pumps. PMID- 15284911 TI - Standards in the extraction phase, a new approach to calibration of microextraction processes. AB - A standard, preloaded onto the extraction phase prior to the extraction step, partially desorbs to the sample matrix during sampling. The amount lost can be used as a means of calibration. PMID- 15284912 TI - Development and assessment of a miniaturised centrifugal chromatograph for reversed-phase separations in micro-channels. AB - This paper describes the micro-fabrication and preliminary assessment of a miniature polydimethylsiloxane (PDMS) device for performing rapid, parallel liquid phase chromatographic separations driven by centrifugal force in microchannels. Device components include a main separating channel, into which a high performance liquid chromatography (HPLC) particulate stationary phase was packed under pressure by application of centrifugal force, in addition to solvent and sample reservoirs. Also described are methods for sealing such devices based upon partial polymerisation of PDMS. The mobile phase flow rate through a typical device was measured and several important chromatographic parameters determined from a test separation. An expression describing mobile phase flow through packed channels was also developed, based upon work on liquid flow in open micro channels. Good agreement between predicted and measured flow rates were observed. Some predictions for potential uses of such devices and possibilities for further miniaturisation are discussed. PMID- 15284913 TI - HPLC simultaneous analysis of thiols and disulfides: on-line reduction and indirect fluorescence detection without derivatization. AB - A liquid chromatography method with indirect fluorescence detection has been developed for simultaneous detection of cysteine, cystine, homocysteine, homocystine, glutathione and glutathione disulfide. After separation in their native forms, a post-column solution of tris(2-carboxyethyl)phosphine (TCEP) at 32 mM, pH 8 is added to reduce the disulfides on-line to the corresponding thiol. The effluent is then merged with a second post-column solution of the highly fluorescent complex Cd(HQS)(2)(2-). The cadmium is complexed by the eluting thiols, effectively quenching the fluorescence. Optimization of the separation, the on-line reduction and the indirect fluorescence detection are discussed. Detection limits from 0.3-4.3 microM (0.04 to 2.6 ppm) are achieved for the six analytes in a 20 min separation. PMID- 15284914 TI - Extraction of arsenic species from spiked soils and standard reference materials. AB - The abilities of various extractants to recover four arsenic species [As(iii), As(v), dimethylarsinic acid (DMA), and monomethylarsonic acid (MMA)] from soils spiked with 20 micro g g(-1) As were investigated. The extractants were water, buffer solutions (citrate and ammonium dihydrogen phosphate), acidic solutions (phosphoric acid and acetic acid), a basic solution (sodium hydroxide) and household chemicals (vinegar and Coca Cola). Gentle shaking at room temperature with each extractant for 24 h gave different recoveries for the different arsenic species. With 0.1 M NaOH solution 46% As(iii), 53% DMA, 100% MMA and 84% As(v) were recovered. A rapid extraction procedure using a sonicator probe has been developed to obtain higher extraction efficiencies. Extracts of arsenic-spiked soil, SRM 2711 Montana soil and SRM 2709 San Joaquin soil were analyzed by HPLC ICP-MS. In the SRM water extracts, DMA and MMA were identified in addition to inorganic arsenic. The solution detection limits (3s) were 0.1, 0.12, 0.13 and 0.15 ng mL(-1) for As(iii), DMA, MMA and As(v), respectively for HPLC-ICP-MS. PMID- 15284915 TI - Analysis of corticosteroids by high performance liquid chromatography electrospray mass spectrometry. AB - A high performance liquid chromatography electrospray mass spectrometry (HPLC-ESI MS) method, for the detection of corticosteroids in cosmetics has been developed. A water-acetonitrile linear gradient on a C-18 reversed-phase column was found to be suitable in separating triamcinolone and its main derivatives, which greatly differ in lipophilicity. Detection was performed in negative electrospray ionisation mode. Good correlation between peaks areas and solutions concentration was found in the range 0.05-10.0 micro g ml(-1) and the detection limits resulted in the range of 20-45 pg injected. The method was successfully applied to the analysis of real samples of shampoo. PMID- 15284916 TI - LC-ESI-MS-MS method for the analysis of tetrabromobisphenol A in sediment and sewage sludge. AB - Tetrabromobisphenol A (4,4'-isopropylidenebis(2,6-dibromophenol), TBBPA) is the most widely used brominated flame retardant in the world. Due to its low water solubility TBBPA released in aquatic ecosystems ultimately accumulates in sediments, but the lack of data on its environmental level and temporal trend in sediment cores precludes establishing if the input of TBBPA is an on-going environmental problem. We developed an analytical method involving HPLC-ESI-MS-MS (ion trap) with detection of the negative pseudo-molecular ion of TBBPA and its fragmentation pattern. Recovery of TBBPA from spiked marine sediment (both lyophilized and wet) and dehydrated sewage sludge was better than 95%. The current detection limit of TBBPA is 60 pg injected and the linearity of the response is at least three orders of magnitude, ranging from 7 ng ml(-1) to 7000 ng ml(-1). The method was also applied to the analysis of urban sewage sludge where TBBPA was detected at a concentration of 300 ng g(-1)(dry weight). With an analysis time of less than 20 min, this method is adequate for a rapid re assessment of archived sediment samples avoiding cumbersome derivatization procedures. PMID- 15284917 TI - Disequilibrium effects in metal speciation by capillary electrophoresis inductively coupled plasma mass spectrometry (CE-ICP-MS); theory, simulations and experiments. AB - A theoretical-experimental approach to evaluate disequilibrium effects in capillary electrophoresis inductively coupled plasma mass spectrometry (CE-ICP MS) is presented. Electrophoresis requires metal ligand (ML) complexes to be stable on the time scale of separation and detection. By expressing ML complex stability in terms of half-life during a CE separation, an evaluation of separation artifacts can be made. Kinetically slow metals like Cr, Al or Fe form complexes that are stable on the time scale of electrophoretic separations. Kinetically fast metals, like Pb, Hg, Cu, Cd and REE, however tend to form labile complexes which unless complexed by strong chelators will dissociate during CE separations. A reactive transport simulation model of CE separations involving ML complexes allows a more detailed prediction of disequilibrium bias and identifies kinetically limited from mobility-limited types of dissociation. Complementary experimental results are given for kinetic and equilibrium binding experiments of Sm with humic acid. The equilibrium logK for Sm-Leonardite humic acid (HA) binding at pH 7 and 0.01 mol L(-1) ionic strength was determined to be 13.04. Kinetic rates of formation and dissociation for SmHA were 5.9 10(8) and 5.3 10( 5) mol s(-1). PMID- 15284918 TI - Electrospray ionization mass spectrometry fingerprinting of propolis. AB - Crude ethanolic extracts of propolis, a natural resin, have been directly analysed using electrospray ionization mass (ESI-MS) and tandem mass spectrometry (ESI-MS/MS) in the negative ion mode. European, North American and African samples have been analyzed, but emphasis has been given to Brazilian propolis which displays diverse and region-dependent chemical composition. ESI-MS provides characteristic fingerprint mass spectra, with propolis samples being divided into well-defined groups directly related to their geographical origins. Chemometric multivariate analysis statistically demonstrates the reliability of the ESI-MS fingerprinting method for propolis. On-line ESI-MS/MS tandem mass spectrometry of characteristic [M - H](-) ion markers provides an additional dimension of fingerprinting selectivity, while structurally characterizing the ESI-MS marker components of propolis. By comparison with standards, eight such markers have been identified: para-coumaric acid, 3-methoxy-4-hydroxycinnamaldehyde, 2,2 dimethyl-6-carboxyethenyl-2H-1-benzopyran, 3-prenyl-4-hydroxycinnamic acid, chrysin, pinocembrin, 3,5-diprenyl-4-hydroxycinnamic acid and dicaffeoylquinic acid. The negative mode ESI-MS fingerprinting method is capable of discerning distinct composition patterns to typify, to screen the sample origin and to reveal characteristic details of the more polar and acidic chemical components of propolis samples from different regions of the world. PMID- 15284919 TI - Poly(decyl methacrylate)-based fluorescent PEBBLE swarm nanosensors for measuring dissolved oxygen in biosamples. AB - 150-250 nm Poly(decyl methacrylate)(PDMA) fluorescent ratiometric nanosensors for dissolved oxygen have been developed. Platinum octaethylporphine ketone (PtOEPK), the oxygen-sensitive dye, and octaethylporphyrin (OEP), the oxygen-insensitive dye, have been incorporated into PDMA nanoparticles to make the sensors ratiometric. Based on the corresponding Stern-Volmer plot, these nanosensors exhibit almost complete linearity over the whole range of dissolved molecular oxygen from 0 to 42.5 ppm (deoxygenated to pure oxygen-bubbled water). The overall quenching response is up to 97.5%, the best so far for all dissolved oxygen optical sensors. These PEBBLE nanosensors also show very good reversibility and stability to leaching and photobleaching, as well as very short response times and no perturbation by proteins. In human plasma they demonstrate a robust oxygen sensing capability, little affected by light scattering and autofluorescence. Potential applications include intracellular oxygen imaging and microresolved pressure profiles in biological and other heterogenous environments. PMID- 15284920 TI - Use of sodium dodecyl sulfate as an antifouling and homogenizing agent in the direct determination of heavy metals by anodic stripping voltammetry. AB - The effectiveness of the anionic surfactant sodium dodecyl sulfate (SDS) for suppressing adsorption interferences in anodic stripping voltammetry (ASV) was investigated. The samples included fruit juices, wine, beer, milk powder and waste water, and the analytes were cadmium, lead and copper. In most of the samples, the ASV signals were severely depressed or even absent due to electrode fouling, but addition of SDS in concentrations up to 10 g L(-1) proved effective in restoring the ASV response. By using SDS as an interference suppressor, the content of lead and copper in a milk powder reference material was determined, and the results were in agreement with the certified values. In this determination, which could not have been performed without SDS, the surfactant also served as a homogenizing agent, preventing separation of the sample components. The effect of SDS was explained by the interaction of the surfactant with the electrode surface and with the constituents of the sample matrix. PMID- 15284921 TI - Electrocatalytic detection of thiols using an edge plane pyrolytic graphite electrode. AB - The first example of using an edge plane pyrolytic graphite electrode in electroanalysis is reported as the determination of homocysteine, N acetylcysteine, cysteine and glutathione is studied. The response of the electrode in the direct oxidation of thiol moieties is explored and found to be electrocatalytic producing a reduction in the overpotential while having enhanced signal-to-noise characteristics compared to glassy carbon and basal plane pyrolytic graphite electrodes. The effectiveness of the methodology is examined in the determination of cysteine species in a growth tissue media that contains a high number of common biological interferences. The advantageous properties of this electrode for thiol determination lie in its excellent catalytic activity, sensitivity and simplicity. PMID- 15284922 TI - Chemical polymerization of m-phenylenediamine, in the presence of glucose oxidase, produces an enzyme-retaining electrooxidisable polymer used to produce a biosensor for amperometric detection of glucose from brain dialysate. AB - A new procedure involving chemical polymerization of a monomer of m phenylenediamine (m-ppd) containing glucose oxidase (GOx) and subsequent electro synthesis of the functional GOx containing polymer onto platinum needle electrodes (PTNE) was used for the amperometric analysis of glucose concentration in brain dialysates. Monomer solutions of o-phenylenediamine (o-ppd) and m-ppd were polymerized by low concentrations of glutaraldehyde (GA) and precipitated from solution. The 1,3 position of the amines on the benzene was amenable to stable polymerization by GA but polymerization of o-ppd (1,2 position) by GA was unstable and degraded. Polymerization of m-ppd appears to proceed by dehydration synthesis. GA induced polymerization of m-ppd polymer in the presence of GOx produced a polymer with strongly bound, functional GOx. This GOx-m-ppd polymer formed a stable matrix that was effectively employed in flow injection analysis (FIA) of glucose. If maintained under O(2) free atmosphere after chemical polymerization, the GOx-m-ppd polymer retained the ability to be electropolymerized. PTNE coated with GOx-m-ppd polymer by repeated dip/amperometry produced stable, sensitive amperometric glucose sensors with good interference exclusion properties and long shelf life. Scanning EM demonstrated that amperometry modified the structure of the GOx-m-ppd on the PTNE surface. GOx m-ppd PTNE glucose sensors and bare PTNE were placed in a radial flow cell and FIA was employed for the simultaneous measurement of glucose and ascorbic acid, respectively, from dialysates of brain tissue. PMID- 15284923 TI - Robustness of the extraction step when parallel factor analysis (PARAFAC) is used to quantify sulfonamides in kidney by high performance liquid chromatography diode array detection (HPLC-DAD). AB - The robustness of a multiresidue method has been analysed for the extraction and quantification of sulfamethoxypyridazine, sulfamethoxazole and sulfadimethoxine in porcine kidney by HPLC-DAD through a Plackett-Burman design. Two experimental responses were examined, the mean recovery from three replicates (accuracy) and their standard deviation (precision). Three factors were tested: the volume of phosphoric acid (pH) added in the extraction step, the time used for passing the sample through the solid-phase extraction cartridge (flow rate) and methanol volume to elute the analytes from the cartridge. Due to the non-specificity of the chromatograms (unknown matrix interferences coelute with each sulfonamide) the PARAFAC model was employed to evaluate the concentration recovered in the experiments of the Plackett-Burman design as well as to identify the spectra of the substances according to the criteria set in the European Decision 2002/657/EC for the analysis of residues. The extraction step was concluded to be robust to the recovery and the standard deviation of all three analytes. PMID- 15284924 TI - Isotope selective analysis of CO(2) with tunable diode laser (TDL) spectroscopy in the NIR. AB - The performance of a home-built tunable diode laser (TDL) spectrometer, aimed at multi-line detection of carbon dioxide, has been evaluated and optimized. In the regime of the (30(0)1)(III) <-- (000) band of (12)CO(2) around 1.6 microm, the dominating isotope species (12)CO(2), (13)CO(2), and (12)C(18)O(16)O were detected simultaneously without interference by water vapor. Detection limits in the range of few ppmv were obtained for each species utilizing wavelength modulation (WM) spectroscopy with balanced detection in a long-path absorption cell set-up. High sensitivity in conjunction with high precision -- typically +/ 1 (per thousand) and +/-6 (per thousand) for 3% and 0.7% of CO(2), respectively - renders this experimental approach a promising analytical concept for isotope ratio determination of carbon dioxide in soil and breath gas. For a moderate (12)CO(2) line, the pressure dependence of the line profile was characterized in detail, to account for pressure effects on sensitive measurements. PMID- 15284925 TI - Design and characterization of a directly polarized radioluminescent light source. AB - The properties of several directly polarized radioluminescent light (RL) sources are described. The RL sources are composed of a [small beta]-particle-emitting (90)Sr radioisotope coupled to polarization-oriented light-emitting plastic scintillators. The polarization of the scintillator molecules was achieved through a mechanical stress applied to the polymer matrix. The emission stability of the RL sources is described. It was found that the degree of polarization of light emitted by the sources varied from zero to 26%, depending on the matrix and scintillating material. The RL source with the highest degree of polarization was constructed from polystyrene and anthracene. PMID- 15284926 TI - Calcium selective test strip for water and milk. AB - We have developed a selective and reversible test strip based on an ion-exchange mechanism to determine calcium. The optical test strip contains a polymeric film of plasticised PVC that contains all of the reagents necessary to produce a response to calcium, among them the new ionophore, 4,13-bis[(N adamantylcarbamoyl)propionyl]-1,7,10,16-tetraoxa-4,13-diazacyclooctadecane. The measurement of the absorbance at 655 nm in a standard photometer makes it possible to determine calcium activities. The composition of the membrane and reaction conditions have been adjusted to obtain adequate selectivity. The test strip responded linearly to calcium between 0.050 and 135 mM in activities. The reproducibility intermembrane at a medium level of the range was 6.2%, as RSD, of log a(Mg(2+)), and 3.4% as RSD intramembrane. The procedure was applied to the determination of calcium ion in different types of waters (tap, well, spring and mineral) and milks (whole, skimmed, skimmed with calcium added, special types) validating the results against a reference procedure. PMID- 15284928 TI - A ton of Clinics. PMID- 15284927 TI - Anxious patients in the medical setting. PMID- 15284929 TI - Biostatistics 301. Repeated measurement analysis. PMID- 15284930 TI - Assessment of maternal anxiety levels before and after amniocentesis. AB - INTRODUCTION: To assess anxiety levels in mothers before and after undergoing amniocentesis. The secondary aim was to see how counselling by nurse-counsellors affected maternal anxiety levels. METHODS: A prospective study was carried out from February 2000 to August 2000 at the Kandang Kerbau Women's and Children's Hospital in Singapore. We used standard statistical analysis and Spielberger's state-trait anxiety inventory (STAI), that consisted of 40 items, to assess anxiety levels. Anxiety levels were assessed at different stages: before and after counselling; before amniocentesis and after amniocentesis; when results were disclosed; and after the routine 20-week screening ultrasound scan was acknowledged four to six weeks later. English-speaking women were recruited for the study as the STAI questionnaire has only been validated for an English speaking population. 195 at-risk mothers (advanced maternal age, abnormal nuchal translucency on ultrasound scan, previous abnormal baby and high-risk maternal serum screening results) and patients requesting for amniocentesis between 15 to 20 weeks gestation were recruited. RESULTS: 156 mothers agreed to amniocentesis. 38 mothers declined amniocentesis. S-anxiety levels declined significantly after counselling by trained nurse-counsellors in all mothers counselled. S-anxiety levels were highest and significantly higher compared to all other times just prior to amniocentesis despite counselling. Anxiety levels were the lowest and significantly lower compared to all other times at the last assessment stage. CONCLUSION: High level of anxiety prior to amniocentesis despite counselling is understandable due to the invasive nature of the procedure. There is no long lasting post-procedural anxiety to the mother. PMID- 15284931 TI - Does maternal serum screening for Down syndrome induce anxiety in younger mothers? AB - INTRODUCTION: To assess anxiety levels in mothers with low-risk pregnancies before and after offering routine serum screening. METHODS: A prospective study was carried out at the Kandang Kerbau Women's and Children's Hospital in Singapore from February 2000 to August 2000. We used standard statistical analysis and Spielberger's state-trait anxiety inventory (STAI) which consists of 40 items to assess anxiety. Anxiety levels were assessed at several stages: before serum screening counselling, after counselling but before serum screening, before the routine 20-week obstetrical screening ultrasound scan, and after ultrasound scan results were acknowledged four to six weeks later. As the STAI questionnaire has only been validated for an English-speaking population, only English-speaking women were recruited for the study. The subjects included 111 women between 15 to 20 weeks gestation that were randomly selected (without any risk factors) for serum screening counselling. RESULTS: Anxiety levels did not decline significantly after counselling by a trained nurse-counsellor. They were highest prior to counselling and were significantly higher compared to all other times in which anxiety was assessed. Anxiety levels were lowest after the serum screening and routine 20-week screening ultrasound scan results were acknowledged. They were also significantly lower compared to all other times in which anxiety was assessed. CONCLUSION: Anxiety before serum screening was not abnormally high and routine serum screening offered by trained nurse counsellors did not significantly increase maternal anxiety in mothers with low risk pregnancies. PMID- 15284932 TI - The role of computed tomography in clinically-suspected but equivocal acute appendicitis. AB - INTRODUCTION: To study the role of contrast-enhanced computed tomography (CT) of the abdomen and pelvis in the evaluation of patients with clinically-suspected but equivocal acute appendicitis. METHODS: The medical records of 206 consecutive patients who had CT of the abdomen and pelvis for equivocal signs and symptoms of acute appendicitis were reviewed. 7 mm collimated axial sections from the diaphragm to the iliac crest and 5mm collimated sections of the pelvis with intravenous and oral contrast were obtained. The criteria used to diagnose acute appendicitis were: (a) a thickened appendix of more than 7 mm or (b) inflammatory changes in the periappendiceal fat. The CT findings were correlated with the histological diagnosis at appendectomy. If the CT findings were negative for acute appendicitis and surgery not performed, the results were correlated with other corroborating diagnostic investigations or clinical follow-up. RESULTS: A total of 206 patients were scanned, of which 39 were excluded due to lack of any follow-up. Of the final 167 that were studied, there were 36 true positives, 127 true negatives, 4 false negatives and no false positives, resulting in a sensitivity of 93.9 percent, specificity of 100 percent and accuracy of 98.5 percent. CONCLUSION: We have found CT to be a safe, reliable and accurate modality in the diagnosis of acute appendicitis in patients with equivocal presentation. PMID- 15284933 TI - Cardiac and electrocardiographical manifestations of acute organophosphate poisoning. AB - INTRODUCTION: To study the extent, frequency and pathogenesis of the cardiac and electrocardiographical manifestations of acute organophosphate poisoning. METHODS: 37 adult patients admitted over a three-year period with a diagnosis of acute organophosphate or carbamate poisoning were studied prospectively. The clinical features and electrocardiographical finding were recorded. RESULTS: Cardiac complications developed in 23 patients (62.2 percent). These were: non cardiogenic pulmonary oedema in eight cases (21.6 percent), electrocardiographical abnormalities including prolonged Q-Tc interval in 14 cases (37.8 percent), ST-T changes in 11 cases (29.7 percent), and conduction defects in two cases (5.4 percent). Sinus tachycardia occurred in 15 patients (40.5 percent) and sinus bradycardia in seven patients (18.9 percent). Hypertension developed in five patients (13.5 percent) and hypotension in four patients (10.8 percent). Five patients (13.5 percent) needed respiratory support because of respiratory depression of which two patients developed intermediate syndrome. Out of 14 patients with prolonged Q-Tc interval, only one had polymorphic ventricular tachycardia of the torsade de pointes type. Two patients died from non-cardiogenic pulmonary oedema and one from ventricular fibrillation, giving a hospital mortality of 8.1 percent. CONCLUSION: Cardiac complications usually occur during the first hour after exposure. Hypoxemia, electrolyte derangements and acidosis are major predisposing factors for the development of these complications. Intensive supportive treatment, meticulous respiratory care and administration of atropine in adequate doses vary early in the course of the illness will reduce the mortality. PMID- 15284934 TI - Endobronchial tuberculosis simulating bronchial asthma. AB - Pulmonary tuberculosis is still a major health problem worldwide, but the principles of diagnosis and treatment are well-established. Endobronchial tuberculosis (EBTB) is known to complicate pulmonary tuberculosis and its importance lies in the potential for bronchostenosis. In the absence of parenchymal disease, EBTB is less well-recognised and can lead to difficulties in diagnosis. We report a 26-year-old woman who presented with symptoms of cough, shortness of breath and wheezing simulating bronchial asthma. Although the chest radiograph did not show any lung infiltrate, a bronchoscopy was carried out. The findings, suspicious of malignancy, were actually due to EBTB, which was confirmed on histology by special stains and on culture for Mycobacterium tuberculosis. The patient subsequently developed bronchostenosis, a well described complication. PMID- 15284935 TI - Epithelioid schwannoma of the vestibular nerve. AB - Epithelioid schwannomas are rarely encountered intracranially, with only four cases involving the eighth nerve reported in the literature. Histological behaviour ranging from benign to aggressive has been described. We report a 45 year-old woman who presented with right-sided tinnitus and hearing impairment. Magnetic resonance imaging revealed a tumour in the right cerebellopontine angle with intracanalicular extension. The patient underwent retromastoid craniectomy with near-total tumour excision. Microscopical examination confirmed the diagnosis of epithelioid schwannoma of the vestibular nerve. Intraoperative findings of sharp circumscription, bland histological appearance, low proliferative activity, coupled with the indolent clinical course, point to the quiescent nature of the lesion in this case. PMID- 15284936 TI - Clinics in diagnostic imaging (100). Migrated pharyngeal fish bone. AB - A 71-year-old man presented with a suspected swallowed fish bone. The lateral radiograph of the neck showed a curvilinear radio-opaque density in the swollen pre-vertebral soft tissues. The diagnosis of a migrated fish bone was confirmed on computed tomography and during subsequent surgery. The patient made a good recovery. As calcified normal structures, particularly the laryngeal cartilages, can mimic abnormal radio-opaque foreign bodies, it is important to be able to recognise the normal calcified structures seen on the neck radiograph. A sound knowledge of radiological anatomy is required in order to avoid unnecessary investigation and to provide prompt and appropriate management. PMID- 15284937 TI - Air and blood occupational exposure limits for lead and representing results. PMID- 15284940 TI - [Reliability survey and standardization of the narcissism inventory based on a healthy sample]. AB - The German "Narzissmusinventar" (Narcissism Inventory, Deneke and Hilgenstock 1989) was used in many clinical studies, but until now there was only very little data about healthy controls. Therefore, this study will survey data from a bigger healthy sample, examine the influence of sex and age and study the retest reliability. Data of 273 healthy controls was gathered in a dentist practice and 100 of the participants filled out the questionnaire for a second time after one month. Except for the scale "smallness self", all scales revealed fairly good reliability values (retest and internal consistency). Also the item-total correlations and the connection between the items and the scales were satisfying. There were significant differences between men and women in 9 of the 18 scales. The group of young people (ages 15 to 20) differed significantly from the older age groups in 13 scales. These young probands described themselves as more anxious and more driven by emotions. Therefore separate evaluations for this group are suggested. Overall, the good psychometric properties of the Narcissism Inventory have been verified. PMID- 15284941 TI - [Improving medical education by enhancing behavioural and social science content of medical school curricular (not only in the USA)]. PMID- 15284942 TI - [Cardiological and psychosocial status of patients with malignant ventricular arrhythmias before implantation of a cardioverter defibrillator. First results from the German Austrian ICD Multicenter Study (GAIMS)]. AB - In a prospective multicenter study of coping, subjective well-being and objective course of the disease we recruited patients with life-threatening cardiac arrhythmias awaiting the implantation of a cardioverter defibrillator. All patients received a semistructured interview and a number of well validated self assessment questionnaires. In addition, detailed cardiological findings were documented. The present paper describes the study rationale and design as well as the main study hypotheses. In addition, we present representativity data for the inclusion sample and cross-sectional psychometric findings obtained before implantation of the device. The study sample consists of 286 patients with severe ventricular arrhythmias and is almost representative for all ICD recipients in the participating centers and ICD recipients in general. Despite their severe physical impairment, patients only showed moderate levels of psychological abnormalities. Only patients with severe heart failure or a history of repeated resuscitations showed elevated rates of anxiety or depression. However, there were relevant associations among the self-rating scales: Patients with abnormal anxiety or depression scores reported significantly elevated levels of physical complaints and depressive coping. They also showed low social support and an impaired quality of life. These cross-sectional findings add to the international literature on coping and well-being of patients with malignant cardiac arrhythmias. On the background of earlier research findings and clinical experience our results show high plausibility. Prospective changes over time in the different dimensions of psychosocial adjustment and their prognostic power for future quality of life and arrhythmic events will be reported separately. PMID- 15284943 TI - [Coherence in case conceptualizations--concept and evaluation of a computer supported training program]. AB - In several domains of professional expertise it has been found that experts' action is based on a deep understanding of the problems they act on, and also the outcome of psychotherapies depends on the quality of the view which therapists develop of their patients. Unfortunately there are signs that the quality of case conceptualizations is suboptimal in many instances, even after special (traditional) training. This was reason for developing an innovative program for the training of partial abilities required to develop differentiated case conceptualizations. In the program specifically the graphical representation of the relations a therapist sees is supported by a computer program. The program gives fast feedback on several formal aspects of the case conceptualization, among others its coherence, and supports its further elaboration. An evaluation of the program in a controlled trial with a total of 34 participating therapists showed a good acceptance and significant effects in several criteria for the quality of the representation of the analyzed cases. PMID- 15284944 TI - [Attachment security and quality of life in atopic dermatitis]. AB - OBJECTIVE: Attachment style is an important variable of influence on general and disease specific quality of life. Insecure attachment may go along with increased autonomic vulnerability and with compromised social support and coping resources. Patients with atopic dermatitis often report interactions of the status and severity of their disease with psychic wellbeing. It was the purpose of the present study to clarify the influence of attachment style and social support on the quality of life in probands with atopic dermatitis. We hypothesized that securely attached test persons with severe skin affection feel less constrained in their quality of life, because they may "buffer" strains from the disease more successfully due to better social support deriving from more satisfying relationships. METHOD: We carried through a cross-sectional study on 124 probands with atopic dermatitis of varying severity, assessed with self report (Neurodermitis Severity Index, NSI) and in part by clinical dermatological ratings (NSI/SCORAD). Attachment style was captured with the Relationship Scales Questionnaire (RSQ), social support with the F-SOZU (K-22), and quality of life with the Skindex-29. RESULTS: We found slightly more insecurely attached individuals in the study sample than it was to be expected from representational healthy samples. Securely attached participants have no lower severity of their disease, but show a higher amount of social support. Irrespective of severity, securely attached persons feel less limited in their quality of life compared to insecurely attached participants. In their symptom based quality of life both feel equally limited. Severity predicts quality of life: contrary to our expectation of a buffering effect of attachment security this association is closer in securely attached probands. CONCLUSION: Suffering from atopic dermatitis may go along with more attachment insecurity. The unexpected stronger association between severity and quality of life in securely attached individuals can be interpreted as a more direct reaction to their skin condition in the absence of other encumbering factors (relationship problems etc.). PMID- 15284945 TI - [Acute otitis media in children: an evidence-based practice guideline]. AB - Acute Otitis media is one of the most common acute respiratory infections managed in primary care and the most common infection among in children. Diagnostic criteria, however, do not always correspond to scientific evidence. They often differ depending on individual preferences and competences. Treatment, also, is controversial. In Germany, most children attending their pediatrician or primary care physician will be prescribed antibiotics. Evidence from several randomized studies and systematic reviews suggests that routine usage of antibiotics provides only modest benefit. The benefit of prescribing antibiotics should not only be balanced against the increased likelihood of side effects such as diarrhoea but also against the potential to contribute to longterm antibiotic resistance. PMID- 15284946 TI - [Concerning specific language impairment: intelligibility in expressive language]. AB - BACKGROUND: Even in the age of two years toddlers with Expressive Language Impairment (SLI-E) differ regarding their phonetic inventories compared with an age-matched group developing normally. METHOD AND PATIENTS: PCC-R scores of 19 children with expressive language disorder, aged between four and six years, are compared with age and sex matched controls with normal language acquisition. RESULTS: There are significant differences between PCC-R scores, children with expressive language disorder show lower scores than children with normal language acquisition. Age, sex and nonverbal intelligence do not influence performance measured with PCC-R. Analysis regarding early, middle and late consonants shows, that expressive language disordered children show bigger problems in aquiring late than early and middle consonants. PMID- 15284947 TI - A comparison of efficacy between two natural exogenous surfactant preparations in premature infants with respiratory distress syndrome. AB - The mortality and various morbidity rates have been substantially reduced by means of exogenous surfactant replacement, the cornerstone in the treatment of respiratory distress syndrome (RDS) in premature infants. The objective of this study is to compare two natural surfactant preparations (Alveofact(R), Survanta(R)) in terms of effectiveness and side-effects. A total of 50 infants with RDS were given surfactant due to RDS were taken into the scope of this study. Survanta(R) and Alveofact(R) were administered to randomized infants with RDS and the results obtained during clinical observations were compared. Second hour mean FiO (2), MAP and a/APO (2) values showed changes in favour of Alveofact(R) (n = 25) group compared to the Survanta(R) (n = 25) group (p < 0.05 for each parameter). However, this difference disappeared in the 6 (th) hour. No statistical difference was established between the two groups with regard to sideeffects (pneumothorax, sepsis, intraventricular hemorrhage, bronchopulmonary dysplasia), duration of mechanical ventilation in survivors, duration of hospitalization in survivors and mortality before the 28 (th) day. It was concluded that results obtained with different surfactant preparations having dissimilar compositions were not different in terms of final impacts and side effects. PMID- 15284948 TI - [Multilocular erythema migrans in borreliosis]. AB - BACKGROUND: Borreliosis is the most common vector transmitted disease in childhood. Although the disease manifests with an erythema migrans in 80 % of the patients, multilocular skin manifestations are only observed in 2-18 % of these. Differential diagnoses of erythema migrans include erysipelas, persistent insect bite reaction, and fixed drug eruption, in particular when the clinical history does not reveal a tick bite. PATIENT: We report on a 5-year-old boy showing nine erythemas with central pallor on his face, trunk, arms and legs. He recalled a tick bite 3 weeks before. RESULTS: Serological studies revealed an acute infection with Borrelia burgdorferi. After antibiotic treatment with orally administered amoxicillin skin manifestations resolved within three days. During a follow-up period of six months the patient revealed no signs of persistent borreliosis. CONCLUSION: Multilocular erythema migrans is a possible manifestation of borreliosis and is classified as disseminated early infection which is frequently associated with systemic reactions, including malaise, arthritis, carditis, headache and even meningeal signs. Treatment is based on antibiotics, which should preferably be given intravenously in case of systemic signs. PMID- 15284949 TI - Yield of bronchial forceps biopsies in addition to nasal brushing for ciliary function analyses in children. AB - BACKGROUND: The diagnosis of primary ciliary dyskinesia (PCD) is unlikely, if ciliary beat frequency (CBF) is normal. The aim of this study was to test the diagnostic value of an additional bronchial biopsy in cases where nasal CBF are abnormal. PATIENTS, METHODS: In a paediatric bronchitis population nasal brush biopsies and bronchial forceps biopsies were taken. In both samples we measured CBF and compared results to nasal CBF of infants and children without respiratory disease. RESULTS: Patients with bronchitis (n = 31; 0.3 to 14.6 years; 10 girls) had a normal CBF in their nasal biopsies in 68 %, and in bronchial biopsies in 48 %, compared to the reference group (n = 72; 0.5 to 17.5 years; 23 girls). One patient had an abnormal nasal, but a normal bronchial ciliary activity. When cilia were beating at both sites (n = 14), nasal CBF agreed well with bronchial CBF (mean difference -0.78 Hz, 95 % confidence interval -1.81 Hz to 0.25 Hz). CONCLUSIONS: By adding the investigation of bronchial mucosa to the measurement of nasal CBF the diagnostic yield to exclude PCD was only improved from 68 % to 71 %. Consequently, if nasal ciliary activity is abnormal in infants and children with bronchitis, we do not recommend additional bronchoscopy to obtain another biopsy. PMID- 15284950 TI - Long-term outcome after neonatal parenchymatous brain lesions. AB - BACKGROUND: Although considered of high prognostic impact, knowledge on the long term outcome after neonatal parenchymatous brain lesions (PBL) is limited. PATIENTS: 29 children with either unilateral (n = 19) or bilateral (n = 10) hemorrhagic/ischemic PBL. METHODS: The patients were reinvestigated at 9 9/12 +/- 3 4/12 years of age, using a standardized clinical investigation, the Beery Buktenica Scales of Visuomotor Integration (VMI) and the Bruininks-Oseretzky Test of Motor Proficiency (BOT). The parents were questioned by means of a standardized questionnaire and the Child Behavior Checklist (CBCL). RESULTS: 90 % of the children showed cerebral palsy (including 12 with hemi- and 8 with tetraplegia). Only 11 % showed normal results on BOT and 39 % on VMI testing. 50 % were bed wetters. Six had required ventriculoperitoneal shunting and 11 were on long-term antiepileptic therapy. Herewith bilateral versus unilateral lesions and low 5-minute APGAR scores were associated with poorer outcome (Cox model and Kaplan-Meier analysis). During follow-up the impact of different disabilities changed. Despite the high rate of cerebral palsy, 71 % learned to walk unaided and 86 % to communicate with words. The last patient learned to walk at 7 years of age. Only one showed poor seizure control. No severe shunt-related complications occurred after 5 years of age. Social, cognitive and behavioral problems increased with age. Only 34 % could attend mainstream schools or kindergartens, and only 50 % displayed normal behavior according to CBCL data, with attention deficiency and social problems being the most important domains. In consequence, nearly all children required 24-hour supervision. CONCLUSIONS: After birth, organic problems such as delayed motor development, epilepsy and ventriculoperitoneal shunting are of major importance for children with PBL. Although delayed, basic skills such as verbal communication are achieved by the majority of patients. In later childhood and adolescence, social, behavioral and cognitive problems increase. In the future, vast resources will be required to provide adequate education and carers as substitutes for elderly parents. PMID- 15284951 TI - Are sequence family variants useful for identifying deletions in the human Y chromosome? PMID- 15284953 TI - Problematic use of Greenberg's linguistic classification of the Americas in studies of Native American genetic variation. PMID- 15284954 TI - The phylogeography of mitochondrial DNA haplogroup L3g in Africa and the Atlantic slave trade. PMID- 15284956 TI - Beckwith-Wiedemann syndrome and IVF: a case-control study. PMID- 15284957 TI - [A randomised open trial comparing monotherapy with topiramate versus carbamazepine in the treatment of paediatric patients with recently diagnosed epilepsy]. AB - INTRODUCTION: In some studies in children, topiramate showed efficacy. AIM: To evaluate efficacy, tolerability and safety of topiramate in monotherapy in newly diagnosed epilepsy vs carbamazepine in children. PATIENTS AND METHODS: In a multicentre, open-label, comparative and randomized study patients with partial epilepsy, were randomized to received topiramate or carbamazepine treatment. Patients with degenerative disease were excluded. Data were analysed by SPSS statistical program v. 11.0, and non parametric test. Comparisons between groups were made with chi square test and t Student's test. RESULTS: In total were included 88 patients, 33 in group 1 (topiramate), 32 group 2 (carbamazepine), 23 were drop-outs because adverse events and lost in follow-up (13 in group 1 y 10 group 2). In both groups were observed good efficacy, in month 6 and 9 of follow up, the average of seizures in group 1 were better than group 2 (p = 0.01, t Student's test). The percentage of free seizure patients was greater in group 1 than group 2 (statistical significance p = 0.02 chi square test). The adverse events were similar in both groups and mild, somnolence 9%, weight loss 6% in group 1 and somnolence 19%, dizziness 3% and seizure discontrol 3% in group 2. CONCLUSIONS: Good efficacy in both groups, and topiramate in good treatment choice in newly diagnosed epilepsy in children because it's the efficacy and tolerability in comparison with the gold standard carbamazepine. PMID- 15284958 TI - [Neuropsychological tests abbreviated and adapted to Spanish speakers: review of previous findings and validity studies for the discrimination of patients with anterior vs. posterior lesions]. AB - INTRODUCTION: There is a lack of neuropsychological tests for Spanish-speakers validated by the cerebral location of the injury. Then the antecedents of the present instrument are described. AIMS: To study the validity of the battery for the differentiation of healthy subjects versus patients and, in patients, anterior versus posterior telencephalic lesions. PATIENTS AND METHODS: 37 patients with frontal lesions, 37 with posterior lesions and 148 healthy subjects were studied. The three groups were matched by sex, age, years of study, manual preference, laterality of the injury, type of lesion and time since onset of condition. RESULTS: The discriminant analysis reached a percentage of correctly predicted cases of 86% for posterior lesions, 85% for anterior lesions, and 99% for healthy subjects. The analysis of the items of the initial interview, only with the group of patients, resulted in a correct prediction of the observations of 90% or over for both lesions. In general, tests which best discriminated anterior from posterior lesions were the ones that worst discriminated anterior lesion from healthy subjects with exception of an easy paired-associated word learning test which turned out to be valid for the discrimination of the three groups. CONCLUSION: Present results demonstrated that this battery of abbreviated tests adapted to Spanish speakers was valid for the discrimination of the three studied groups. Methodological approaches based on objective evaluations and multiple comparisons rather than on clinical appreciation are proposed. PMID- 15284959 TI - [Time evolution of TNF-alpha, VCAM-1, IL-4, IL-10, neopterin and CD-30 in patients treated with interferon]. AB - INTRODUCTION: Interferon (IFN) diminishes the outbreaks of multiple sclerosis (MS) and slows down its progression. Follow-up of patients is performed using clinical and resonance imaging parameters, and no biological markers are available that allow us to determine its efficiency. AIMS: 1. To discover the effects of IFN on the serum levels of TNF-alpha, IL-4, IL-10, VCAM-1, neopterin and CD-30 in patients with MS; 2. To determine how these modifications evolve over time; 3. To find out the clinical value of its determination in isolation. PATIENTS AND METHODS: We studied 19 patients with MS who were clinically stable and undergoing IFN therapy. Samples were obtained every 3 months over a 2.5 year period and always immediately before injecting the drug. The ELISA method was used to determine interleukins. RESULTS: Serum levels of neopterin, CD-30 and VCAM-1 were not modified, TNF-alpha levels oscillated regardless of the clinical status of the patient and IL-4 and IL-10 had a significant serum peak at 9-12 months after beginning treatment. CONCLUSIONS: The existence of a significant IL 4 and IL-10 peak between 6 and 12 months of therapy indicates that IFN reaches its possible immunomodulatory effect after several months and, therefore, a poor initial clinical response must not be a reason for discontinuing medication. The specific determination of the serum levels of IL is not useful in following up patients treated with IFN. PMID- 15284960 TI - [Analysis of the aetiologies of headaches in a paediatric emergency service]. AB - INTRODUCTION: Headaches are a frequent cause of children's visits to different health care services but the epidemiological profile varies widely according to whether the sample is taken from among visits to hospital clinics, neuropaediatric units or emergency services. AIMS: The aim of this study was to analyse the aetiologies of the cases of headache attended in a paediatric emergency service. PATIENTS AND METHODS: We conducted a retrospective analysis of patient records with headache as the main reason for visiting over a six month period at the Paediatric Emergency Department at the Centro Hospitalario Pereira Rossell. Aetiologies were classified according to International Headache Society (IHS) criteria and the information was completed by telephone if the medical records contained insufficient data. RESULTS: The records of 185 children were studied, which represented 0.58% of the total number of visits. The most common aetiologies were non-cranial infections (43.1%), migraines (14.6%), tension-type headaches (9.2%), traumatic injury (8.1%) and sinusitis (5.4%). Serious causes constituted 4.9% of the total, with a predominance of tumours and infections of the central nervous system. The low percentage of viral meningitis in our series (1%) leads us to suspect an underdiagnosis of this disorder, given the scant number of cases in which the cerebrospinal fluid was examined. CONCLUSIONS: Our study confirmed the existence of a wide and varied range of aetiologies causing headaches, with a clear predominance of extracranial disorders and a low percentage of serious intracranial causes. Non-classifiable headaches made up 10.8% of the sample. Brain imaging studies were performed in 9.7% of cases, which is a considerably lower percentage than that of other series and demonstrates a rational use of this resource. PMID- 15284961 TI - [A link between congenital malformations of the central nervous system and epilepsy in paediatric patients in Mexico]. AB - INTRODUCTION: Since epilepsy is one of the most frequent causes of visits in Paediatric Neurology, attention must be given to one of the causes linked to it, namely congenital malformation, which is the second most common cause of epilepsy. To this end, different forms of congenital defects related to epilepsy in Paediatric medicine can be identified according to their neurodevelopment. AIMS: The purpose of this study was to determine the different congenital malformations associated to epilepsy in Paediatrics. PATIENTS AND METHODS: We took a sample consisting of 116 patients diagnosed as suffering from epilepsy associated with congenital malformations of the central nervous system, following an evaluation of imaging studies, magnetic resonance and computerised tomography brain scans; subjects were then grouped according to the normal embryonic chronological development of the human being. RESULTS: From the total number of cases, a selection was made according to age, where the predominant group was found in those below one year of age and in the group of school-age children, and migration disorders, where the main malformation included was lissencephaly; the other group was made up of proliferation disorders. Similarly, the associated types of epilepsy were the most common childhood epileptic syndromes, West and Lennox-Gastaut syndrome. The types of epileptic seizures that were found were partial seizures. CONCLUSIONS: The study outlined above shows congenital malformations of the central nervous system to be the main cause associated to epilepsy and the most sensitive neuroimaging study currently available is magnetic resonance. For this reason we suggest the use of this procedure in cases in which no apparent cause can be found so that this nosological entity can be defined to a greater degree of precision. Despite its multifactorial causation, being below 25 years of age and above 35 at the time of pregnancy is considered to constitute a higher potential risk, while no geographic location was found to predominate. PMID- 15284962 TI - [Stroke as the first manifestation of temporal arteritis: three case reports and a review of its pathogenesis and treatment]. PMID- 15284963 TI - [Neurological complications arising from dengue virus infection]. AB - INTRODUCTION: Dengue is the most common of the arbovirosis that humans can suffer from. The frequency with which the central nervous system (CNS) is affected by this viral infection remains unknown, although isolated cases with neurological complications have been reported in Asia and South America. In Rio de Janeiro, Brazil, dengue virus infection has become an important public health concern. CASE REPORTS: The authors describe two cases of immune-mediated CNS involvement following classic infection by the dengue virus: one involving post-infectious disseminated acute encephalitis and the other consisting of Guillain-Barre syndrome. In both cases dengue was diagnosed using the ELISA technique, and other viral aetiologies in the cerebrospinal fluid (CSF) were excluded. A 10-year-old female, following a bout of classic dengue, presented symptoms of a diminished level of consciousness, spastic tetraparesis, cerebellar syndrome and frontal symptoms. A resonance brain scan showed areas of hypersignal in T2 sequences in the cerebral peduncle, lentiform nuclei and internal capsule on both sides of the brain, which suggested post-infectious encephalitis. The second patient, a 14 year-old male, presented an areflexive flaccid ascending tetraparesis that suggested acute polyradiculoneuritis, following a bout of classic dengue. CSF albuminocytologic dissociation was also observed. This patient's electroneuromyogram recording showed a polyradiculoneuropathy of a primarily demyelinating nature with an associated axonal component. CONCLUSIONS: The immunological mechanisms involved in the pathophysiology of this type of neurological complications after suffering from dengue may be part of the physiological response to the viral infection. PMID- 15284964 TI - [Cerebellopontine angle lipoma: a case report]. AB - INTRODUCTION: Intracranial lipomas are infrequent tumours and especially so when they are situated in the cerebellopontine angle (CPA), when they only constitute 0.05% of the total number. CASE REPORT: We present the case of a CPA lipoma diagnosed in a 38-year-old female with hypoacusis as the presenting symptom and in which magnetic resonance (MR) was the complementary test that proved most useful as a diagnostic aid. There was a hyperintense image in T1 in the right side CPA that did not take up contrast and which was interpreted as a fatty tumour. CONCLUSIONS: We discuss the clinical manifestations of CPA lipomas, the means that can be used to diagnose them (mainly focused on the value of MRI) and the procedure to be followed in dealing with these tumours according to the experience obtained from the relatively scarce number of cases reported to date. We underline the idea that these lesions are currently considered to be embryonic development disorders rather than actual neoplasias. PMID- 15284965 TI - [Hypotonia and lethargy: initial manifestations of a new case of galactosemia]. AB - INTRODUCTION: Galactosemia is a metabolic disease that is transmitted by autosomal recessive inheritance in which there is an enzymatic deficit that prevents the metabolism of galactose. Three enzymes could be involved, but the lack of galactose-1-phosphate uridyltransferase (GALT) is the most frequent. Incidence is two cases per 100,000 newborn infants. As a consequence of this enzymatic deficit, on ingesting milk the newborn infant will present a progressive neurological deterioration, cataracts and digestive tract and kidney disorders. An early diagnosis is essential so that galactose can be withdrawn from the diet as soon as possible, which in the newborn infant means discontinuing mother's milk and feeding with galactose-free milk. CASE REPORT: We report the case of a newborn female, the daughter of consanguineous parents (second cousins) from the gypsy ethnic group, who was diagnosed as suffering from galactosemia with a total GALT deficit. The patient was given normal milk for the first 10 days of her life and presented hypotonia, lethargy, jaundice, hepatomegaly, refusal to eat, low weight gain and a urinary infection caused by gram negative bacteria. Following diagnosis, galactose was withdrawn from the diet (she was given soy milk) and the physical exploration became progressively more normal. CONCLUSIONS: This is an extremely unusual pathology, but the patient's outcome is largely dependent on an early diagnosis and treatment. PMID- 15284966 TI - [The role of protein p53 in neurodegenerative processes throughout the 25 years of its history]. AB - INTRODUCTION: In this review we will study the role of protein p53 in neurodegenerative processes and conduct a detailed analysis of the mechanisms responsible for regulating its levels and biological activity. We analyse the neuropathologies in which this protein is involved, such as Alzheimer's and Parkinson's diseases and amyotrophic lateral sclerosis, and we will also examine its regulation by second messengers such as the reactive species of oxygen and calcium, showing the signalling paths involved in the apoptotic processes. DEVELOPMENT: The year 2004 sees the 25th anniversary of the discovery of protein p53. At first p53 was wrongly attributed with an oncogenic function due to its capacity to bind to the T antigen of the virus SV40 in transformed cells. Nevertheless, it was not until 1989 that it was attributed with its true physiological function as a tumour-suppressing protein. This milestone constitutes a turning point in the short life of this protein. Protein p53 plays a fundamental role in the mechanisms the cell uses to respond to damage or mutation in the genome. There is, therefore, a correlation between deletions or mutations in the p53 gene and the development of some kinds of cancer; additionally, increases in the protein levels of its native form have been reported in pathologies where apoptotic processes are high. CONCLUSIONS: Protein p53 plays an essential role in the mechanisms by which the cell responds to damage or mutation in the genome. It can activate two signalling mechanisms that lead either to stopping the cell cycle or to the death of the cell due to apoptosis if the cell cannot repair the damage to the genome. There is a correlation between its deletions and mutations and the development of cancer, and increases in its native form have been described in pathologies where apoptotic processes are high, as is the case of some neurodegenerative diseases. PMID- 15284967 TI - [Epilepsy beginning in the neonatal period and early infancy]. AB - INTRODUCTION: In this study we analyse several epileptic syndromes that begin in the neonatal period or early infancy, up to two years of age, and we also define their clinical, neurophysiological and progressive aspects as well as their differential diagnoses. Both neonatal and febrile convulsions are excluded. DEVELOPMENT: The different syndromes are classified according to the predominant type of seizures, which is the one that identifies them, although it is not the only type of seizure presented. In line with this reasoning, the syndromes were divided into four main groups: 1. Epileptic spasms: infantile spasms (West's syndrome), periodic spasms as described by Gobbi and bouts of epileptic seizures without hypsarrhythmia. 2. Tonic seizures: Lennox-Gastaut syndrome. 3. Myoclonias: benign myoclonic epilepsy in infancy, Dravet's severe myoclonic epilepsy, myoclonic-astatic epilepsy and a myoclonic state in non-progressive encephalopathies; and 4. Partial seizures: non-idiopathic location-related epilepsies, malign epilepsy with migratory partial seizures and benign infantile familial and non-familial seizures. CONCLUSIONS: Thus, it will be possible to establish a plan of studies, differential diagnoses and a rational therapeutic approach depending on the clinical manifestations, while at the same even enabling us to distinguish between the idiopathic, cryptogenic and symptomatic forms. PMID- 15284969 TI - [Pituitary apoplexy as the presenting symptom of a recurrent craniopharyngioma]. PMID- 15284970 TI - [Tachyarrhythmia as the first manifestation in a classic Rett syndrome]. PMID- 15284972 TI - Mini-incision for total hip arthroplasty: a prospective, controlled investigation with 5-year follow-up evaluation. AB - A group of 42 primary total hip arthroplasties performed through an abridged surgical incision (group 1) was prospectively compared to a cohort of 42 primary total hip arthroplasties performed through a standard surgical incision (group 2). The length of the incision was 8.8 +/- 1.5 cm for group 1 and 23.0 +/- 2.1 cm for group 2. The groups were not significantly different with respect to age, height, preoperative Harris Hip scores (HHS), estimated blood loss, or length of hospital stay (P>.05). Group 1 patients had a lower body mass index than group 2 patients (P<.01). Length of surgery was slightly less for group I (P =.02). A 0% incidence was found of infection, nerve palsy, component malposition, and aseptic loosening in both groups. No dislocations occurred in group 1, and one dislocation occurred in group 2. Patients in group 1 have expressed considerable enthusiasm regarding the cosmetic appearance of the surgical incisions, and their postoperative HHS are slightly higher than those of group 2 (P =.042). Total hip arthroplasty can be performed safely and effectively through an abridged surgical incision, but this investigation confirms no dramatic clinical benefit other than cosmetic appeal. PMID- 15284973 TI - Cementless titanium tapered-wedge femoral stem: 10- to 15-year follow-up. AB - This investigation is an ongoing clinical and radiographic analysis of a titanium tapered-wedge femoral component with a proximal plasma-spray porous coating. Integral femoral stems (Biomet, Warsaw, IN) were implanted in 200 hips in 186 patients. Nineteen patients died before 10-year follow-up, and 50 patients were lost to follow-up. The mean follow-up of the remaining 129 hips was 11.6 years. Harris Hip Scores improved from 58 to 93. Thigh pain was 2.3%. Radiographic analysis revealed adaptive distal remodeling in zones 2, 3, 5, 6, and 13, with no evidence of osteolysis below the level of the calcar and the greater trochanteric region. Only 2 femoral stems were revised: 1 with suspected fibrous fixation at 7 years postoperatively and another with a broken trunion at 10 years postoperatively. A tapered titanium femoral stem with circumferential plasma spray porous coating provides excellent long-term fixation, durable clinical outcome, and protects against osteolysis below the level of the calcar and greater trochanter. PMID- 15284974 TI - Balancing severe valgus deformity in total knee arthroplasty using a lateral cruciform retinacular release. AB - We report the technique and results of a cruciform lateral release performed on 35 consecutive knees having > or =15 degrees of valgus with minimum 2-year follow up. The posterior cruciate ligament (PCL) was preserved in all knees. Preoperative valgus averaged 17 degrees, and range of motion averaged 10 degrees to 107 degrees. Postoperative valgus averaged 4.8 degrees, and average postoperative range of motion was 2 degrees to 110 degrees. The PCL was partially released in 5 knees, and further lateral release of the lateral collateral ligament and popliteus were required in 3 knees. Stable flexion and extension gaps were achieved in all cases, and stability was maintained at follow-up. This lateral cruciform retinacular release provides a simple surgical technique for most valgus deformities of the knee and allows for stable ligamentous balancing. PMID- 15284975 TI - Soft-tissue tension total knee arthroplasty. AB - It is far from clear how best to define the proper strength of soft-tissue tensioning in total knee arthroplasty (TKA). We attached a torque driver to the Monogram balancer/tensor device and measured soft-tissue tension in full extension and 90 degrees flexion during TKA. In our surgical procedure, when we felt proper soft-tissue tension was being applied, the mean distraction force was noted to be 126N in extension and 121N in flexion. There was no significant correlation between soft-tissue tension and the postoperative flexion angle finally achieved. To the best of our knowledge, this is the first study to assess the actual distraction forces in relation to soft-tissue tension in TKA. Further study may reveal the most appropriate forces to achieve proper soft-tissue tension in the wide variety of circumstances presenting at knee arthroplasty. PMID- 15284976 TI - Revision and salvage patellar arthroplasty using a porous tantalum implant. AB - The purpose of this study was to assess the clinical results of a new surgical technique and novel porous tantalum implant for augmentation or arthroplasty of the patella for 11 patients who underwent revision total knee arthroplasty. Preoperative average knee function and pain scores were 24 and 20, respectively, and average range of motion (ROM) was 62 degrees. The low knee scores reflect the immobility, trauma, and/or pain associated with the patients' presenting conditions. At the most recent follow-up (average, 32 months), the average knee function and pain scores were 69 and 53, respectively, and the average ROM was 103 degrees. Radiographically, all implants were stable, and patient satisfaction has been excellent. These results indicate that this surgical technique and porous tantalum prosthesis can substantially improve function and reduce pain for patients with severe patellar bone loss and other complicating factors. PMID- 15284977 TI - Cementless revision hip arthroplasty using strut allografts and primary cementless proximal porous-coated prosthesis. AB - This study evaluated the role of a primary cementless proximal porous-coated prosthesis in revision arthroplasty of the hip with substantial bone loss. It included 14 men and 22 women with a mean age at the time of the index revision of 56.8 years (range, 39 to 68 years). They were followed up for a minimum of 5 years (mean 6.5 years; range, 5 to 8 years). The Harris Hip score improved from a mean of 40 points (range, 22 to 55 points) before revision surgery to a mean of 85 points (range, 51 to 93 points) at the latest review. One femoral stem (3%) was revised for aseptic loosening. Two stems (5.5%) had a varus shift, and these were interpreted as radiologic loosening. All allografts were predictably united to the host femur. The medium-term result of this technique in selected cases was excellent, and the clinical outcome was very satisfactory. PMID- 15284978 TI - The value of hip aspiration versus tissue biopsy in diagnosing infection before exchange hip arthroplasty surgery. AB - The exclusion of infection is the most important factor in determining treatment options in a failed hip arthroplasty. Preoperative biopsy in the form of aspiration or tissue biopsy can be used to diagnose infection. Preoperative aspiration and tissue drill biopsy were performed in 273 consecutive patients under general anesthesia 3 months before exchange arthroplasty surgery. The results of the preoperative aspiration and tissue biopsy cultures were compared with the definitive intraoperative tissue cultures obtained at surgery. Seventy one of 273 (26%) hips were infected. Overall accuracy of aspiration was 90.1% and tissue biopsy 87.9%. The sensitivity and specificity was 80% and 94% for aspiration and 83% and 90% for tissue biopsy. Positive predictive value and negative predictive values were 81.4% and 93.1% for aspiration and 73.8% and 93.8% for tissue biopsy. The more invasive tissue drill biopsy offers no advantage over aspiration in terms of bacterial accuracy and results in more false positives. PMID- 15284979 TI - Medial screws and cement: a possible mechanical augmentation in total knee arthroplasty. AB - The purpose of this study was to determine whether screws placed beneath the medial tibial plateau in cemented total knee arthroplasty helps prevent collapse of the medial tibia. A previous study found that the AGC all-polyethylene tibial component had a 14% rate of collapse of the medial subchondral region in the first postoperative year. Of 536 implanted AGC all-polyethylene tibial components, 20 had screws inserted beneath the medial tibial plateau. No AGC all polyethylene tibial components with screws failed because of aseptic loosening or collapse of the medial tibial plateau. The study included 125 cemented metal backed total knee arthroplasties with screws inserted beneath the medial tibial plateau. We also found 2 cases of collapse of the medial tibial plateau and 1 case of collapse on the lateral side. No revisions were performed. The placement of screws beneath the medial tibial plateau to fill large defects is an excellent precaution against collapse of the medial tibia. PMID- 15284980 TI - Sagittal plane kinematics of a mobile-bearing unicompartmental knee arthroplasty at 10 years: a comparative in vivo fluoroscopic analysis. AB - This study compares in vivo sagittal plane kinematics of the Oxford mobile bearing unicompartmental knee arthroplasty (UKA) at 1 and 10 years' postsurgery (10 knees) with a fixed-bearing total knee arthroplasty (TKA) (5 knees) and the normal knee (5 knees), using dynamic fluoroscopic measurement of the patellar tendon angle. The Oxford UKA preserved normal changes in patellar tendon angle with flexion, and this was maintained at 10 years. In contrast, an abnormal pattern was seen with the TKA. The results suggest that a normal pattern of sagittal plane knee kinematics exists following Oxford medial UKA and imply that anterior cruciate ligament function is maintained in the long term. PMID- 15284981 TI - The initial stability of an exeter femoral stem after impaction bone grafting in combination with segmental defect reconstruction. AB - Bone impaction grafting of the femur is associated with more complications when segmental defects are present. The effect of segmental defect repair on initial stem stability was studied in an in vitro study with fresh-frozen goat femora. A standardized medial segmental defect was reconstructed using a cortical strut or a metal mesh. As controls, we used intact femora and femora with a nonreconstructed defect. In all 4 groups, impacted bone grafting was performed in combination with a cemented Exeter stem. Each group contained 5 femora. Reconstructions were dynamically loaded up to 1,500 N. Migration was measured with Roentgen stereophotogrammetric analysis. All cases with a nonreconstructed segmental defect failed into excessive varus rotation. None of the femora with a reconstructed defect failed. Cortical struts and metal meshes were equally effective in creating a stable stem construction (varus rotation, 2.89 +/- 2.27 and 2.27 +/- 0.57, respectively). Reconstructions with a metal mesh were more reproducible, although the obtained stability was significantly lower (P<.01) when compared with impaction grafting in an intact femur (varus rotation, 0.58 +/ 0.36). PMID- 15284982 TI - Cement microcracks in thin-mantle regions after in vitro fatigue loading. AB - An in vitro study of cemented femoral hip components was conducted to determine if microcracks in the cement mantle would preferentially form in thin-mantle regions as a result of cyclic fatigue loading via stair-climbing. Overall, there was not an increased amount of microcracks in thin-mantle (<2 mm) regions (number found/number expected = 0.59, P<.03). However, through cracks that extended between the stem to the bone were more prevalent in thin-mantle regions (number found/number expected = 2.93, P<.03). Although cracks form throughout the cement mantle and appear to grow at the same rate, thin-mantle regions are most likely to have through cracks after fatigue loading. This is consistent with results from at-autopsy studies of well-fixed femoral components and supports the general guideline that thin-mantle regions should be avoided in the cementing of the femoral stem. PMID- 15284983 TI - Distal rotational alignment of the Chinese femur and its relevance in total knee arthroplasty. AB - The rotational alignments of the distal femur in southern Chinese subjects were investigated. Chinese femurs are significantly more externally rotated. This is new evidence to suggest a racial difference in the distal femoral geometry. During total knee arthroplasty, a common recommendation is to allow 4 degrees of external rotation. The clinical significance of our study is that this need to be modified to 6 degrees for Chinese women and 5 degrees for Chinese men. This racial difference can be incorporated into a new concept of "a mountain and a molehill." This concept is presented in this report. PMID- 15284984 TI - Three-dimensional lower extremity alignment assessment system: application to evaluation of component position after total knee arthroplasty. AB - A 3-dimensional (3D) lower extremity alignment assessment system based on biplanar computed radiography, a camera calibration procedure, and 3D image fitting techniques was developed. The goal of this study was to examine the errors associated with applying this 3D technique to determine component alignment after total knee arthroplasty (TKA). Tibial and femoral component alignment after TKA (varus--valgus, flexion-extension, and internal--external angulations) in 4 patients was determined directly with reconstructions from computed tomography (CT) scans and indirectly with the proposed bone and component image fitting techniques. Mean differences between alignment values from the CT and image fitting techniques ranged from 0.18 degrees to 0.78 degrees, and maximum differences ranged from 0.30 degrees to 0.90 degrees. The proposed 3D method can be reliably used to assess component alignment after TKA. PMID- 15284985 TI - Accuracy of haptic assessment of patellar symmetry in total knee arthroplasty. AB - Resection resulting in asymmetrically thick patellar arthroplasty in knee arthroplasty may lead to increased compression forces, wear, fracture, or loss of quadriceps power. Assessing the cut patella between the thumb and forefinger (haptic assessment) represents a convenient way to recognize asymmetry. In 2 test series, 8 orthopedic surgeons evaluated 24 precut solid foam patellae of varying asymmetric thickness by feeling the patella between thumb and forefinger, without visualization. Of 384 responses, in 73.2% asymmetry was underestimated, 10.4% assessments were exact, and in 16.4% asymmetry was overestimated. Specifically, 35.9% were correct within 1 mm, 60.7% within 2 mm, 81% within 3 mm, and 91.4% within 4 mm. The thickest half of the patella was correctly identified in 90.6% of responses. Haptic assessment of patellar symmetry is a useful technique in knee arthroplasty. PMID- 15284986 TI - Lack of mental-status changes in patients undergoing total knee arthroplasty. AB - According to the literature, a significant number of patients present with mental status changes following total joint arthroplasty. It has been postulated that mental-status changes would be increased in patients undergoing bilateral joint arthroplasty. To assess the mental status of patients undergoing total knee arthroplasty (TKA), a validated test was performed in a prospective fashion. Thirty patients underwent unilateral TKA, and 30 underwent sequential simultaneous bilateral TKA. The Folstein Mini Mental Status examination was performed before and after surgery. No significant changes were noted in this patient population after surgery. Furthermore, there were no significant differences between the unilateral and the bilateral groups. PMID- 15284987 TI - Massive wear of an incompatible metal-on-metal articulation in total hip arthroplasty. AB - We report on a case of massive wear because of an incompatible metal-on-metal combination. In a 62-year-old man, a cobalt-chromium (CoCr) inlay and a stainless steel head were paired by accident. Because of persistent pain, revision surgery was performed 7 months later. Histologic analysis of the surrounding tissue revealed massive metallosis. The wear volume was increased by a factor of 18 for the head and 2 for the cup compared with normal metal-on-metal articulation. The serum concentrations of chromium and cobalt were increased by a factor of 20 and 4 over levels of a healthy population, respectively. Incompatible metal-on-metal combinations should be revised immediately. In case of delayed diagnosis, no metal-on-metal articulation should be implanted because of the high volume of metal in the human body. PMID- 15284988 TI - Fixation using acetabular reconstruction cage and cancellous allografts for intraoperative acetabular fractures associated with cementless acetabular component insertion. AB - Displaced acetabular fractures that occur during insertion of cementless acetabular component are a rare but very serious complication. We report two cases that were treated successfully with the utilization of an acetabular reconstruction cage and morsellized allografts. PMID- 15284989 TI - Late dissociation of an alumina-on-alumina bearing modular acetabular component. AB - Alumina-on-alumina bearings for total hip arthroplasty have been introduced as a promising alternative to reduce wear debris and to increase the life expectancy of the prosthesis. We report a case of a late dissociation of an alumina-on alumina bearing modular acetabular component, which occurred 2 years, 8 months after surgery. Detailed analysis of the retrieved prosthesis suggested that the cause of the failure may be strong rotational torque developed by a roughening of the bearing's alumina surface caused by edge loading (microseparation). The strong friction torque in articulation may explain the higher reported rate of mechanical loosening of the acetabular component in alumina-on-alumina bearing total hip prostheses. PMID- 15284990 TI - Polyethylene post failure in posterior stabilized total knee arthroplasty. AB - Posterior-stabilized (PS) prostheses have been used extensively in total knee arthroplasty (TKA), with excellent long-term results. The key feature of these prostheses is the femoral cam and tibial post mechanism that limits posterior displacement and produces femoral rollback. Although articular-surface polyethylene wear of the tibial component has not been a significant clinical problem, tibial post wear has been reported. In distinction to chronic post wear, little information exists about catastrophic post failure. We present the case of a 56-year-old woman who presented 63 months after TKA with a PS prosthesis with acute fracture of the polyethylene post. The evaluation and treatment of this patient, including the previously unreported use of computed tomography arthrography to diagnose this rare problem, is reviewed. PMID- 15284991 TI - Fatigue failure of the femoral component of a cementless total hip arthroplasty. AB - This case report describes a fracture of the neck of a femoral component of a Furlong (JRI Limited, London) hydroxyapatite-coated cementless total hip arthroplasty. PMID- 15284992 TI - Dislocation of polyethylene liner in uncemented Harris Galante II acetabulum: report of 6 cases. AB - We report the dislocation of 6 liners out of the metal backing shells of Harris Galante II cementless total hip prostheses implanted between 1993 and 1996. The time between primary implantation and failure varied from 46 to 95 months. The dislocations were probably the result of suboptimal mechanical fixation of the polyethylene liner in the shell compounded by suboptimal positioning of the acetabular component. In 5 cases, both the metal backing and the liner were revised because of broken tines. In one case, only the liner was exchanged. PMID- 15284993 TI - Fifty-one-year survival of a Judet polymethylmethacrylate hip prosthesis. AB - Revision arthroplasty of a Judet polymethylmethacrylate (PMMA) hip hemiprosthesis was performed 51 years after implantation. This makes it the first implant to have survived for longer than 50 years. No signs of aggressive osteolysis were found on histologic analysis, which confirms good tissue tolerance for PMMA. Judet's hip prosthesis did not follow the concept of low-friction arthroplasty; nevertheless, it represents an important step in the development of the hip prosthesis currently used. PMID- 15284994 TI - Correct limb alignment after a knee arthroplasty. PMID- 15284996 TI - Reconstruction of the iliofemoral ligament with an artificial ligament for recurrent anterior dislocation of total hip arthroplasty. PMID- 15285000 TI - [Glomerulonephritis and renal sclerosis: new therapeutic proposals (review)]. AB - The treatment of immuno-mediated glomerulonephritides is presently based upon a limited series of drugs. Albeit evidence-based medicine relies solely upon controlled trials, there is a need to follow new perspectives with an open mind, since they may lead to tomorrow's therapy. Several original and innovative approaches to treat inflammatory glomerular diseases have been recently reported, including drugs designed to limit the effect of pro-inflammatory and pro sclerotic cytokines (recombinant monoclonal antibodies, receptor antagonists, gene therapy providing viral transfection of genes, antisense oligonucleotides, aptamers, inhibition of transcription factors, active immunization). Moreover, newer options are being proposed, as in the case of enhancing natural anti inflammatory cytokines or intracellular signalling limiting inflammation. Some of these proposals, which are briefly reviewed in this article, are likely to enter soon clinical investigation and to become in the next future standard treatment for glomerular diseases. PMID- 15285001 TI - [Autoimmune and lymphoproliferative HCV-correlated manifestations: example of mixed cryoglobulinaemia (review)]. AB - Mixed cryoglobulinaemia (MC) is a systemic vasculitis involving small vessels (arterioles, capillaries, venules). The histological hallmark of the disease is the leukocytoclastic vasculitis secondary to the vascular deposition of circulating immune-complexes (CIC), mainly cryoglobulins and complement. The immune-mediated vasculitic lesions are responsible for different MC clinical features, including cutaneous and visceral organ involvement. Hepatitis C virus (HCV) represents the triggering factor in the large majority of MC patients (>90%). Moreover, several epidemiological, clinico-pathological and laboratory investigations suggested a possible role for HCV in a wide spectrum of immuno lymphoproliferative disorders; namely, porphyria cutanea tarda, diabetes, polyarthritis, lung fibrosis, poly-dermatomyositis, thyroiditis, thyroid cancer, B-cell non-Hodgkin's lymphomas (B-NHL), etc. Renal involvement with or without MC syndrome can be observed in HCV-infected individuals. There is great geographical etherogeneity in the prevalence of HCV-related disorders. This epidemiological observation suggests a multifactorial and multistep process in the pathogenesis of these conditions, involving other unknown genetic and/or environmental factors. HCV lymphotropism may explain the mono-oligoclonal B-lymphocyte expansion observed in HCV-infected individuals, particularly in MC patients. The 'benign' lymphoproliferative disorder, classified as monotypic lymphoproliferative disorders of undetermined significance (MLDUS), may be responsible for the wide production of CIC, including cryoglobulins, rheumatoid factor and different organ and non-organ specific autoantibodies. The consequence is the appearance of various HCV-related autoimmune diseases, including MC syndrome. This latter may be complicated by B-NHL in 10% of the cases; moreover, HCV infection has been confirmed in a significant percentage of 'idiopathic B NHL. For a correct therapeutic approach to cryoglobulinaemic vasculitis, as well as to other HCV-related disorders, we should deal with concomitant, conflicting conditions: HCV infection, autoimmune and lymphoproliferative alterations. In this scenario, we can treat the diseases at three different levels by means of etiologic, pathogenetic and/or symptomatic therapies. The eradication of HCV by combined interferon and ribavirin therapy can be achieved in only a minority of cases. On the contrary, severe complications such as glomerulonephritis, sensory motor neuropathy or diffuse vasculitis can be effectively treated by a combination of corticosteroids, plasma exchange and cyclophosphamide. More recently, a pathogenetic treatment with rituximab, a monoclonal chimeric antibody that binds to the B-cell surface antigen CD20 with selective B-cell blockade, was proposed in patients with HCV-related MC syndrome. PMID- 15285002 TI - [Disturbances of mineral metabolism and vascular calcifications in dialysis patients (review)]. AB - Vascular calcifications are more frequent in dialysis patients than in the general population or in patients with cardiovascular disease and normal renal function. The reasons for this high incidence are multiple. They include traditional factors such as hypertension, diabetes, dyslipidaemia, and specific factors such as sodium overload, hyperomocysteinaemia, chronic inflammation, oxidative stress as well as disturbance of mineral metabolism. Specifically, hyperphosphataemia and the elevated calcium (Ca) x phosphate product have been associated with an increased risk for development of vascular calcification and death. Even though a causal relationship between the use of Ca- containing phosphate binders and the development of vascular calcifications has not been documented, treatment with Ca salts can induce hypercalcaemia, increased Ca x phosphate product, and Ca overload. A net intestinal Ca absorption of 180-500 mg has been documented in uraemic patients after a meal containing 1200 mg of Ca. Thus, treatment with Ca salts may induce Ca overload when a patient is dialyszed against a high dialysate Ca (> 1.5 mmol/L) solution, which is known to determine a positive dialysis balance. On the contrary, an overall negative Ca balance can result from the use of a low Ca dialysate (1.25 mmol/L) when the patients do not receive Ca supplements or vitamin D metabolites. Maintaining a normal Ca and phosphate balance remains one of the primary goals in the management of dialysis patients. Control of hyperphopshataemia should be obtained using either Ca and aluminium- free phosphate binders, such as sevelamer, or Ca salts, while avoiding a daily oral elemental Ca intake > 1.5 g. PMID- 15285003 TI - [Are convective treatments equivalent to the traditional ones? The Hemo Study and beyond (review)]. AB - Dialysis treatments have allowed 'terminal patients' to live for years and years. However, life expectancy and quality are still consistently reduced in renal dialysis patients. Consequently, all efforts to device alternative treatments to the conventional ones are highly justified. Recently, the Hemo Study showed that neither the use of high flux membranes, nor the increase of the dialysis dose above the conventional, were capable to reduce significantly patient's mortality and morbidity, although 8% reduction of the risk of death was seen in patients treated with high flux vs. patients treated with low-flux dialysis. A relevant question is if convective treatments may offer an overprotection from morbidity and mortality, in comparison with low flux and high flux treatments. Data from the Registro Lombardo di Nefrologia e Trapianto published in 2000 showed a trend toward a better survival (RR= 90) and a significantly better protection from tunnel carpal syndrome (RR= 0.58; p= 0.03) in patients treated with convective treatments (hemofiltration and/or hemodiafiltration) vs. patients treated with diffusive dialysis. Except than a better cardiovascular stability observed on hemofiltration and an higher beta2-microglobuline clearance given by online hemofiltration and hemodiafiltration, evident clinical benefits of convective treatments, over the conventional high flux treatments, are not yet clearly demonstrated. Notwithstanding that, online convective treatments, that are performed with high flux compatible membranes and high technology machines, producing high quality water, offer at the moment the best bases for the improvement of clinical results of dialysis, especially in some category of patients. PMID- 15285004 TI - [When the history of nephrology changed that of medicine]. AB - Medicine owes much to nephrology. Indeed many of the practical and doctrinal acquisitions, through nephrology have derived their first intuitions, explanations and applications which have become epochal conquests of scientific progress. This article is a historical reconstruction of six of the milestones which have marked the medical and scientific human progress: Galeno, the ligature of the ureters and the birth of experimental medicine; uroscopy and the introduction of laboratory exams; the synthesis of urea in the laboratory and the beginnings of biothecnology; the kidney and the introduction of systematic parenteral antibiotic therapy; the kidney and the first artificial organs; the kidney and the start of the transplantation era. PMID- 15285005 TI - [Italian study on the treatment of anaemia in chronic dialysis patients switched over to less frequent doses of darbepoetin from human recombinant erythropoietin (rHuEPO)]. AB - BACKGROUND: Darbepoetin alpha is a novel erythropoiesis stimulating protein with unique properties as compared to recombinant human erythropoietin (rHuEPO), including a three-fold longer elimination half-life that allows for less frequent dosing. This study was aimed at testing the efficacy and safety of darbepoetin alpha in a large number of chronic dialysis patients switched from rHuEPO. METHODS: Nine hundred and fifty dialysis patients in stable treatment with rHuEPO were switched to darbepoetin alpha. Patients receiving rHuEPO 2 or 3 times weekly were switched to once weekly darbepoetin alpha and those receiving rHuEPO once weekly were switched to once every other week darbepoetin alpha. Patients received darbepoetin alpha by the same route of administration (SC or IV) as the one used for rHuEPO. The unit doses of darbepoetin alpha (10-150 microg) were titrated to maintain haemoglobin concentration within -1.0 and +1.5 g/dL of the individual mean baseline haemoglobin levels and between 10 and 13 g/dL for 24 weeks. RESULTS: The mean change in haemoglobin from baseline to the evaluation period (weeks 21-24) was statistically but not clinically significant [-0.10 g/dL (95% CI: -0.18, -0.02]. In general, the geometric mean weekly dose of study drug from screening/baseline to evaluation period remained substantially unmodified [(from 26.10 micro g/wk to 25.90 microg/wk; percentage change -0.40% (95% CI: 3.78, 3.10)]. Overall, darbepoetin alpha was well tolerated. CONCLUSIONS: The treatment of anaemia of a large dialysis patient population with unit dosing of darbepoetin alpha is effective and safe in maintaining target haemoglobin concentration at reduced dose frequency. PMID- 15285006 TI - [Quality guide in nephrology and dialysis]. AB - The Quality Guide, a strategic tool for any nephrologist wishing to initiate a quality policy in his own renal unit, describes to the organisation of total quality management. This article gives a short description of a Quality Guide in Nephrology and Dialysis. According to ISO 9004:2000 Norms, the Quality Guide can be subdivided into four sections: Managerial Accountability, Resource Management; Implementation of Services; Analysis, Evaluation and Continuous Quality Improvement. The quality cycle begins by identifying the customers, who added to the resources constitute the input that will be transformed by the service into output (results). By measuring customer satisfaction the results will be evaluated and analyzed to allow continuous improvement of the service. In conclusion, the drafting of the Guide by all staff members involved has a twofold value: it organizes the quality management within the renal unit and certifies the quality level guaranteed to the customer. PMID- 15285007 TI - [To delay may be wise]. AB - We report a case of acute renal failure, quickly evolved, in which the coexistence of parenchimal nephropaty and renal mass, have induced not a common diagnostic and therapeutic approach, finalized to optimize the interventional nephrology procedures, with the use of various imaging procedures. It is followed a multidisciplinar therapeutic approach, with the employment of dialysis, steroid therapy and surgical treatment. PMID- 15285008 TI - [The activity of the Soci SIN Mailing List (ML-SIN): an update]. AB - The Soci SIN Mailing List (ML-SIN) is the automatic on-line moderated discussion group for nephrology professionals of the Italian Society of Nephrology, which regularly operates since October 2000. The new publishing initiative of the Giornale Italiano di Nefrologia reserves to the ML-SIN a special survey in which the most discussed topics will be summarised and commented. This paper presents an update of the last six months (October 2003 - march 2004) of ML-SIN activity. The number of ML-SIN members has increased from 525 to 603 (+15%), thus confirming an increasing interest in the ML-SIN as discussion instrument and reliable source of professional advice for the Italian nephrological community. The number of messages received and accepted was 1024, for the most part (49%) related to general or organizational topics (i.e., regulatory information, meeting announcements and management issues) and only 51% to scientific debate. Some of the topics are discussed in detail, e.g. the proposal for a retrospective study on cholesterol crystal embolism and the analysis of dialysis treatment reimbursement rates by the Health System Financial Organization. PMID- 15285009 TI - [Nutcracker syndrome: a difficult case of recurrent gross hematuria]. AB - BACKGROUND: The Nutcracker Syndrome (NS) is an uncommon clinical condition caused by compression and entrapment of the renal vein (LRV) as it passes through the angle between the superior mesenteric artery (SMA) and the aorta (meso-aortic angle). Intermittent macrohematuria, left peripelvic and gonadal vein varices and aspecific abdominal pain may be common manifestations of this syndrome. CASE REPORT: A 15-year-old white boy developed recurrent macrohaematuria in June 2002. He had a history of upper respiratory infection prior the first episode of gross hematuria followed by 4 other episodes of macrohematuria 'sine causa'. Blood pressure and renal function were normal. Routine laboratory tests showed only an increase in serum LDH levels (901 IU/L) with negative Coombs' test, both direct and indirect, and absence of schistocytes in the blood smear. Renal ultrasonography showed normal kidneys while an intravenous pyelography showed a 'minus' in the right ureter. For this reason, a cystoscopy and a retrograde pielography were performed but resulted normal. A renal biopsy was carried out because of the presence of one episode of post-infective macrohaematuria, but light microscopy and immunofluorescence examinations were found to be normal. Renal ultrasonography and Color Doppler ultrasonography (CD-USG) oriented our diagnosis towards NS. An abdominal computerised tomography (CT) scan confirmed that the LRV was compressed between the aorta and the SMA. CONCLUSIONS: NS cannot be diagnosed with routine diagnostic methods. Endoscopic and radiological methods may provide some clues for the presence of NS, while CD-USG may be considered to be the first level non-invasive method for diagnosis. The sensitivity and specificity of this test for diagnosing NS is reported as being 78% and 94%, respectively. The best treatment available for this syndrome is still being debated. PMID- 15285010 TI - Drug therapy for ulcerative colitis. AB - Ulcerative colitis (UC) is an inflammatory destructive disease of the large intestine occurred usually in the rectum and lower part of the colon as well as the entire colon. Drug therapy is not the only choice for UC treatment and medical management should be as a comprehensive whole. Azulfidine, Asacol, Pentasa, Dipentum, and Rowasa all contain 5-aminosalicylic acid (5-ASA), which is the topical anti-inflammatory ingredient. Pentasa is more commonly used in treating Crohn's ileitis because Pentasa capsules release more 5-ASA into the small intestine than Asacol tablets. Pentasa can also be used for treating mild to moderate UC. Rowasa enemas are safe and effective in treating ulcerative proctitis and proctosigmoiditis. The sulfa-free 5-ASA agents (Asacol, Pentasa, Dipentum and Rowasa) have fewer side effects than sulfa-containing Azulfidine. In UC patients with moderate to severe disease and in patients who failed to respond to 5-ASA compounds, systemic (oral) corticosteroids should be used. Systemic corticosteroids (prednisone, prednisolone, cortisone, etc.) are potent and fast acting drugs for treating UC, Crohn's ileitis and ileocolitis. Systemic corticosteroids are not effective in maintaining remission in patients with UC. Serious side effects can result from prolonged corticosteroid treatment. To minimize side effects, corticosteroids should be gradually reduced as soon as the disease remission is achieved. In patients with corticosteroid-dependent or unresponsive to corticosteroid treatment, surgery or immunomodulator is considered. Immunomodulators used for treating severe UC include azathioprine/6 MP, methotrexate, and cyclosporine. Integrated traditional Chinese and Western medicine is safe and effective in maintaining remission in patients with UC. PMID- 15285011 TI - Construction and selection of subtracted cDNA library of mouse hepatocarcinoma cell lines with different lymphatic metastasis potential. AB - AIM: In order to elucidate the molecular mechanism of lymphatic metastasis of hepatocarcinoma, we detected the difference of gene expression between mouse hepatocarcinoma cell lines Hca-F and Hca-P with different lymphatic metastasis potential. METHODS: cDNA of Hca-F cells was used as a tester and cDNA of Hca-P cells was used as a driver. cDNAs highly expressed in Hca-F cells were isolated by the suppression subtractive hybridization (SSH) method. The isolated cDNA was cloned into T/A cloning vector. The ligation products were transformed into DH5alpha competent cells. Individual clones were randomly selected and used for PCR amplification. Vector DNA from positive clones was isolated for sequencing. RESULTS: There were 800 positive clones in amplified subtracted cDNA library. Random analysis of 160 clones with PCR showed that 95% of the clones contained 100-700 bp inserts. Analysis of 20 sequenced cDNA clones randomly picked from the SSH library revealed 4 known genes (mouse heat shock protein 84 ku, DNA helicase, ribosomal protein S13, ethanol induced 6 gene) and 3 expressed sequence tags (ESTs). Four cDNAs showed no homology and presumably represent novel genes. CONCLUSION: A subtracted cDNA library of differentially expressed genes in mouse heptocarcinoma cell lines with different lymphatic metastasis potential was successfully constructed with SSH and T/A cloning techniques. The library is efficient and lays a solid foundation for searching new lymphatic metastasis related genes. The expression of mouse heat shock protein gene, DNA helicase and other 4 novel gene may be different between mouse heptocarcinoma cell lines with different lymphatic metastasis potential. PMID- 15285012 TI - COX-2 expression and tumor angiogenesis in colorectal cancer. AB - AIM: Cyclooxygenase-2 (COX-2) is one of the rate-limiting enzymes in metabolism of arachidonic acid, and COX-2 inhibitors demonstrate preventive effects on cancer, especially on colorectal cancer. The underlying mechanism remains unclear. The aim of this study was to illustrate the relationship between angiogenesis and COX-2 in carcinogenesis of colorectal cancer. METHODS: One hundred and seventy patients with colorectal cancer were enrolled in our study from January 1993 to September 2001 in School of Oncology, Peking University. COX 2 and VEGF expression were detected with the immunohistochemistry (IHC) technique. IHC assays were carried out with the aid of tissue microarray (TMA) procedure. Specimens from 35 of these patients were examined with reverse transcriptase PCR (RT-PCR). RESULTS: COX-2 and VEGF expressions were stronger in colorectal cancer than those in the corresponding normal tissues, at both protein and mRNA levels. One hundred patients were eligible for analysis after IHC assay of COX-2 and VEGF. The positive rate of VEGF was much higher in COX-2 positive group (47/85) than in COX-2 negative group (chi (2) = 4.181, P = 0.041). The result was further verified by the result of RT-PCR (chi (2) = 8.517, P = 0.003). Correlation coefficient was 0.409 after Spearman correlation analysis (P = 0.015). CONCLUSION: COX-2 may be involved in the course of tumor angiogenesis of colorectal cancer and acts through VEGF. PMID- 15285013 TI - New serum biomarkers for detection of HBV-induced liver cirrhosis using SELDI protein chip technology. AB - AIM: To find new serum biomarkers for liver cirrhosis (LC) in chronic carriers of hepatitis B virus (HBV). METHODS: Surface enhanced laser desorption/ionization time-of-flight (SELDI-TOF) mass spectrometry was used to discover biomarkers for differentiating HBV induced LC from non-cirrhotic cohorts. A training population of 25 patients with HBV-induced LC, 20 patients with HCC, and 25 closely age matched healthy men, was studied. RESULTS: Two biomarkers with M(r) 7 772 and 3 933 were detected in sera of non-cirrhotic cohorts, but not in patients with HBV induced LC. A sensitivity of 80% for all LC patients, a specificity of 81.8% for all non-cirrhotic cohorts and a positive predictive value of 75% for the study population were obtained. CONCLUSION: These two serum biomarkers for HBV-induced LC might be used for diagnosis and assessment of disease progression. PMID- 15285014 TI - Association of interleukin-12 p40 gene 3'-untranslated region polymorphism and outcome of HCV infection. AB - AIM: To investigate the effect of interleukin-12 p40 gene (IL12B) 3'-untranslated region polymorphism on the outcome of HCV infection. METHODS: A total of 133 patients who had been infected with HCV for 12-25 (18.2+/-3.8) years, were enrolled in this study. Liver biochemical tests were performed with an automated analyzer and HCV RNA was detected by fluorogenic quantitative polymerase chain reaction. B-mode ultrasound was used for liver examination. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was used for the detection of IL12B (1188A/C) polymorphism. RESULTS: Self-limited infection was associated with AC genotype (OR = 3.48; P = 0.001) and persistent infection was associated with AA genotype (OR = 0.34; P = 0.014) at site 1188 of IL12B. In patients with persistent HCV infection, no significant differences were found regarding the age, gender, duration of infection and biochemical characteristics (P>0.05). According to B-mode ultrasound imaging and clinical diagnosis, patients with persistent infection were divided into groups based on the severity of infection. No significant differences were found in the frequency of IL-12 genotype (1188A/C) between different groups (P>0.05). CONCLUSION: The polymorphism of IL12B (1188A/C) appears to have some influence on the outcome of HCV infection. PMID- 15285015 TI - Helicobacter pylori enhances tumor necrosis factor-related apoptosis-inducing ligand-mediated apoptosis in human gastric epithelial cells. AB - AIM: To investigate the relations between tumor necrosis factor-related apoptosis inducing ligand (TRAIL) and Helicobacter pylori (H pylori) infection in apoptosis of gastric epithelial cells and to assess the expression of TRAIL on the surface of infiltrating T-cells in H pylori-infected gastric mucosa. METHODS: Human gastric epithelial cell lines and primary gastric epithelial cells were co cultured with H pylori in vitro, then recombinant TRAIL proteins were added to the culture. Apoptosis of gastric epithelial cells was determined by a specific ELISA for cell death. Infiltrating lymphocytes were isolated from H pylori infected gastric mucosa, and expression of TRAIL in T cells was analyzed by flow cytometry. RESULTS: The apoptosis of gastric epithelial cell lines and primary human gastric epithelial cells was mildly increased by interaction with either TRAIL or H pylori alone. Interestingly, the apoptotic indices were markedly elevated when gastric epithelial cells were incubated with both TRAIL and H pylori (Control vs TRAIL and H pylori: 0.51+/-0.06 vs 2.29+/-0.27, P = 0.018). A soluble TRAIL receptor (DR4-Fc) could specifically block the TRAIL-mediated apoptosis. Further studies demonstrated that infiltrating T-cells in gastric mucosa expressed TRAIL on their surfaces, and the induction of TRAIL sensitivity by H pylori was dependent upon direct cell contact of viable bacteria, but not CagA and VacA of H pylori. CONCLUSION: H pylori can sensitize human gastric epithelial cells and enhance susceptibility to TRAIL-mediated apoptosis. Modulation of host cell sensitivity to apoptosis by bacterial interaction adds a new dimension to the immunopathogenesis of H pylori infection. PMID- 15285016 TI - Antigen epitope of Helicobacter pylori vacuolating cytotoxin A. AB - AIM: To construct and select antigen epitopes of vacuolating cytotoxin A (VacA) for nontoxic VacA vaccine against Helicobacter pylori (H pylori ) infection. METHODS: Eleven VacA epitopes were predicted according to VacA antigenic bioinformatics. Three candidates of VacA epitope were constructed through different combined epitopes. The candidate was linked with E. coli heat-labile enterotoxin B (LTB) by a linker of 7 amino acids, and cloned into plasmid pQE-60 in which fusion LTB-VacA epitope was efficiently expressed. To test the antigencity of the candidate, 6 BALB/c mice were treated with the fusion LTB-VacA epitope through intraperitoneal injection. To explore the ability of inhibiting the toxicity of VacA,cantiserum against the candidate was used to counteract VacA that induced HeLa cells to produce cell vacuoles in vitro. RESULTS: Serum IgG against the candidate was induced in the BALB/c mice. In vitro, the three antisera against the candidate efficiently counteracted the toxicity of VacA, and decreased the number of cell vacuoles by 14.17%, 20.20% and 30.41% respectively. CONCLUSION: Two of the three candidates, LZ-VacA1and LZ-VacA2, can be used to further study the mechanism of vacuolating toxicity of VacA, and to construct nontoxic VacA vaccine against H pylori infection. PMID- 15285017 TI - 4-hydroxy-2, 3-nonenal activates activator protein-1 and mitogen-activated protein kinases in rat pancreatic stellate cells. AB - AIM: Activated pancreatic stellate cells (PSCs) are implicated in the pathogenesis of pancreatic inflammation and fibrosis, where oxidative stress is thought to play a key role. 4-hydroxy-2,3-nonenal (HNE) is generated endogenously during the process of lipid peroxidation, and has been accepted as a mediator of oxidative stress. The aim of this study was to clarify the effects of HNE on the activation of signal transduction pathways and cellular functions in PSCs. METHODS: PSCs were isolated from the pancreas of male Wistar rats after perfusion with collagenase P, and used in their culture-activated, myofibroblast-like phenotype unless otherwise stated. PSCs were treated with physiologically relevant and non-cytotoxic concentrations (up to 5 micromol/L) of HNE. Activation of transcription factors was examined by electrophoretic mobility shift assay and luciferase assay. Activation of mitogen-activated protein (MAP) kinases was assessed by Western blotting using anti-phosphospecific antibodies. Cell proliferation was assessed by measuring the incorporation of 5-bromo-2' deoxyuridine. Production of type I collagen and monocyte chemoattractant protein 1 was determined by enzyme-linked immunosorbent assay. The effect of HNE on the transformation of freshly isolated PSCs in culture was also assessed. RESULTS: HNE activated activator protein-1, but not nuclear factor kappaB. In addition, HNE activated three classes of MAP kinases: extracellular signal-regulated kinase, c-Jun N-terminal kinase, and p38 MAP kinase. HNE increased type I collagen production through the activation of p38 MAP kinase and c-Jun N-terminal kinase. HNE did not alter the proliferation, or monocyte chemoattractant protein 1 production. HNE did not initiate the transformation of freshly isolated PSCs to myofibroblast-like phenotype. CONCLUSION: Specific activation of these signal transduction pathways and altered cell functions such as collagen production by HNE may play a role in the pathogenesis of pancreatic disorders. PMID- 15285018 TI - Structure prediction and activity analysis of human heme oxygenase-1 and its mutant. AB - AIM: To predict wild human heme oxygenase-1 (whHO-1) and hHO-1 His25Ala mutant (delta hHO-1) structures, to clone and express them and analyze their activities. METHODS: Swiss-PdbViewer and Antheprot 5.0 were used for the prediction of structure diversity and physical-chemical changes between wild and mutant hHO-1. hHO-1 His25Ala mutant cDNA was constructed by site-directed mutagenesis in two plasmids of E. coli DH5alpha. Expression products were purified by ammonium sulphate precipitation and Q-Sepharose Fast Flow column chromatography, and their activities were measured. RESULTS: rHO-1 had the structure of a helical fold with the heme sandwiched between heme-heme oxygenase-1 helices. Bond angle, dihedral angle and chemical bond in the active pocket changed after Ala25 was replaced by His25, but Ala25 was still contacting the surface and the electrostatic potential of the active pocket was negative. The mutated enzyme kept binding activity to heme. Two vectors pBHO-1 and pBHO-1(M) were constructed and expressed. Ammonium sulphate precipitation and column chromatography yielded 3.6-fold and 30-fold higher purities of whHO-1, respectively. The activity of delta hHO-1 was reduced 91.21% after mutation compared with whHO-1. CONCLUSION: Proximal His25 ligand is crucial for normal hHO-1 catalytic activity. delta hHO-1 is deactivated by mutation but keeps the same binding site as whHO-1. delta hHO-1 might be a potential inhibitor of whHO-1 for preventing neonatal hyperbilirubinemia. PMID- 15285019 TI - Effect of SNPs in protein kinase C zeta gene on gene expression in the reporter gene detection system. AB - AIM: To investigated the effects of the SNPs (rs411021, rs436045, rs427811, rs385039 and rs809912) on gene expression and further identify the susceptibility genes of type 2 diabetes. METHODS: Ten allele fragments (49 bp each) were synthesized according to the 5 SNPs mentioned above. These fragments were cloned into luciferase reporter gene vector and then transfected into HepG2 cells. The activity of the luciferase was assayed. Effects of the SNPs on RNA splicing were analyzed by bioinformatics. RESULTS: rs427811T allele and rs809912G allele enhanced the activity of the reporter gene expression. None of the 5 SNPs affected RNA splicing. CONCLUSION: SNPs in protein kinase Cz (PKCZ) gene probably play a role in the susceptibility to type 2 diabetes by affecting the expression level of the relevant genes. PMID- 15285020 TI - Prolongation of liver allograft survival by dendritic cells modified with NF kappaB decoy oligodeoxynucleotides. AB - AIM: To induce the tolerance of rat liver allograft by dendritic cells (DCs) modified with NF-kappaB decoy oligodeoxynucleotides (ODNs). METHODS: Bone marrow (BM)-derived DCs from SD rats were propagated in the presence of GM-CSF or GM CSF+IL-4 to obtain immature DCs or mature DCs. GM-CSF+IL-4-propagated DCs were treated with double-strand NF-kappaB decoy ODNs containing two NF-kappaB binding sites or scrambled ODNs to ascertain whether NF-kappaB decoy ODNs might prevent DC maturation. GM-CSF-propagated DCs, GM-CSF+NF-kappaB decoy ODNs or scrambled ODNs-propagated DCs were treated with LPS for 18 h to determine whether NF-kappaB decoy ODNs could prevent LPS-induced IL-12 production in DCs. NF-kappaB binding activities, costimulatory molecule (CD40, CD80, CD86) surface expression, IL-12 protein expression and allostimulatory capacity of DCs were measured with electrophoretic mobility shift assay (EMSA), flow cytometry, Western blotting, and mixed lymphocyte reaction (MLR), respectively. GM-CSF-propagated DCs, GM CSF+IL-4 -propagated DCs, and GM-CSF+NF-kappaB decoy ODNs or scrambled ODNs propagated DCs were injected intravenously into recipient LEW rats 7 d prior to liver transplantation and immediately after liver transplantation. Histological grading of liver graft rejection was determined 7 d after liver transplantation. Expression of IL-2, IL-4 and IFN-gamma mRNA in liver graft and in recipient spleen was analyzed by semiquantitative RT-PCR. Apoptosis of liver allograft infiltrating cells was measured with TUNEL staining. RESULTS: GM-CSF-propagated DCs, GM-CSF+NF-kappaB decoy ODNs-propagated DCs and GM-CSF+ scrambled ODNs propagated DCs exhibited features of immature DCs, with similar low level of costimulatory molecule (CD40, CD80, CD86) surface expression, absence of NF kappaB activation, and few allocostimulatory activities. GM-CSF+IL-4-propagated DCs displayed features of mature DCs, with high levels of costimulatory molecule (CD40, CD80, CD86) surface expression, marked NF-kappaB activation, and significant allocostimulatory activity. NF-kappaB decoy ODNs completely abrogated IL-4-induced DC maturation and allocostimulatory activity as well as LPS-induced NF-kappaB activation and IL-12 protein expression in DCs. GM-CSF+NF-kappaB decoy ODNs-propagated DCs promoted apoptosis of liver allograft-infiltrating cells within portal areas, and significantly decreased the expression of IL-2 and IFN gamma mRNA but markedly elevated IL-4 mRNA expression both in liver allograft and in recipient spleen, and consequently suppressed liver allograft rejection, and promoted liver allograft survival. CONCLUSION: NF-kappaB decoy ODNs-modified DCs can prolong liver allograft survival by promoting apoptosis of graft-infiltrating cells within portal areas as well as down-regulating IL-2 and IFN-gamma mRNA and up-regulating IL-4 mRNA expression both in liver graft and in recipient spleen. PMID- 15285021 TI - Grey scale enhancement of rabbit liver and kidney by intravenous injection of a new lipid-coated ultrasound contrast agent. AB - AIM: To assess the grey scale enhancement of a new lipid-coated ultrasound contrast agent in solid abdominal organs as liver and kidney. METHODS: Size distribution and concentration of the lipid-coated contrast microbubbles were analyzed by a Coulter counter. Two-dimensional (2D) second harmonic imaging of the hepatic parenchyma, the inferior vena cava and the right kidney of the rabbits were acquired before and after contrast agent injection. Images were further quantified by histogram in Adobe Photoshop 6.0. Time-intensity curves of hepatic parenchyma, inferior vena cava and renal cortex were generated from the original grey scale. RESULTS: The 2D images of hepatic parenchyma and cortex of the kidney were greatly enhanced after injection and the peak time could last more than 50 min. CONCLUSION: This new lipid ultrasound contrast agent could significantly enhance the grey scale imaging of the hepatic parenchyma and the renal cortex for more than 50 min. PMID- 15285022 TI - Intestinal barrier damage caused by trauma and lipopolysaccharide. AB - AIM: To investigate the intestinal barrier function damage induced by trauma and infection in rats. METHODS: Experimental models of surgical trauma and infection were established in rats. Adult Sprague-Dawley rats were divided into 4 groups: control group (n = 8), EN group (n = 10), PN group (n = 9) and Sep group (n = 8). The rats in PN and Sep groups were made into PN models that received isonitrogenous, isocaloric and isovolumic TPN solution during the 7-d period. Rats in EN and Sep groups received laparotomy and cervical catheterization on day 1 and received lipopolysaccharide injection intraperitoneally on d 7. On the 7(th) day all the animals were gavaged with lactulose and mannitol to test the intestinal permeability. Twenty-four hours later samples were collected and examined. RESULTS: The inflammatory responses became gradually aggravated from EN group to Sep group. The mucosal structure of small intestine was markedly impaired in PN and Sep groups. There was a low response in IgA level in Sep group when compared with that of EN group. Lipopolysaccharide injection also increased the nitric oxide levels in the plasma of the rats. The intestinal permeability and bacterial translocation increased significantly in Sep group compared with that of control group. CONCLUSION: One wk of parenteral nutrition causes an atrophy of the intestinal mucosa and results in a moderate inflammatory reaction in the rats. Endotoxemia aggravates the inflammatory responses that caused by laparotomy plus TPN, increases the production of nitric oxide in the body, and damages the intestinal barrier function. PMID- 15285023 TI - DA-9601 for erosive gastritis: results of a double-blind placebo-controlled phase III clinical trial. AB - AIM: To determine the efficacy and safety of DA-9601 on erosive gastritis versus cetraxate as a standard drug by gastrointestinal endoscopy. METHODS: Five hundred and twelve patients with erosive gastritis were divided into three groups. The groups received 180 mg or 360 mg of DA-9601, or 600 mg of cetraxate (Neuer) t.i.d. for 2 wk, respectively. Endoscopic observations were performed before and 2 wk after the treatment, and the cure and improvement rates were investigated. RESULTS: Of the 512 intention-to-treat (ITT) population, 457 patients comprised the per protocol (PP) analysis. Endoscopic cure rate was significantly higher in the DA-9601 group than in the cetraxate group in both the PP (56%, 58% vs 36%; DA 9601 180 mg, 360 mg and cetraxate, respectively) and ITT (52%, 51% vs 35%) populations. Two DA-9601 groups (180 and 360 mg) had significantly higher endoscopic improvement rates than the cetraxate group in both the PP (67%, 65% vs 46%) and ITT (63%, 58% vs 45%) populations. The percentage of symptom relief over the 2 wk was found not significantly different between groups. During the study, both DA-9601 and cetraxate produced no treatment-associated adverse events. CONCLUSION: From these results, it appears that DA-9601 has excellent efficacy on erosive gastritis. This study also confirms the safety profile of DA-9601. PMID- 15285024 TI - Impaired gallbladder motility and delayed orocecal transit contribute to pigment gallstone and biliary sludge formation in beta-thalassemia major adults. AB - AIM: Gallbladder and gastrointestinal motility defects exist in gallstones patients and to a lesser extent in pigment gallstone patients. To investigated the role of gallbladder and gastrointestinal motility disorders in pigment gallstone formation in beta-thalassemia major. METHODS: Twenty-three patients with beta-thalassemia major (16 females; age range 18-37 years) and 70 controls (47 females, age range 18-40 years) were studied for gallbladder and gastric emptying (functional ultrasonography), orocecal transit (OCTT, H(2)-breath test), autonomic dysfunction (sweat-spot, cardiorespiratory reflex tests), bowel habits, gastrointestinal symptoms and quality of life (all with questionnaires). Gallbladder content (ultrasonography) was examined before and during 8-12 mo follow-up. RESULTS: Gallstones and/or biliary sludge were found in 13 (56%) patients. beta-thalassemia major patients had increased fasting (38.0+/-4.8 mL vs 20.3+/-0.7 mL, P = 0.0001) and residual (7.9+/-1.3 mL vs 5.1+/-0.3 mL, P = 0.002) volume and slightly slower emptying (24.9+/-1.7 min vs 20.1+/-0.7 min, P = 0.04) of the gallbladder, together with longer OCTT (132.2+/-7.8 min vs 99.7+/-2.3 min, P = 0.00003) than controls. No differences in gastric emptying and bowel habits were found. Also, patients had higher dyspepsia (score: 6.7+/-1.2 vs 4.9+/-0.2, P = 0.027), greater appetite (P = 0.000004) and lower health perception (P = 0.00002) than controls. Autonomic dysfunction was diagnosed in 52% of patients (positive tests: 76.2% and 66.7% for parasympathetic and sympathetic involvement, respectively). Patients developing sludge during follow-up (38%, 2 with prior stones) had increased fasting and residual gallbladder volume. CONCLUSION: Adult beta-thalassemia major patients have gallbladder dysmotility associated with delayed small intestinal transit and autonomic dysfunction. These abnormalities apparently contribute together with haemolytic hyperbilirubinemia to the pathogenesis of pigment gallstones/sludge in beta-thalassemia major. PMID- 15285025 TI - Clinical features of human intestinal capillariasis in Taiwan. AB - Human intestinal capillariasis is a rare parasitosis that was first recognized in the Philippines in the 1960 s. Parasitosis is a life threatening disease and has been reported from Thailand, Japan, South of Taiwan (Kaoh-Siung), Korea, Iran, Egypt, Italy and Spain. Its clinical symptoms are characterized by chronic diarrhea, abdominal pain, borborygmus, marked weight loss, protein and electrolyte loss and cachexia. Capillariasis may be fatal if early treatment is not given. We reported 14 cases living in rural areas of Taiwan. Three cases had histories of travelling to Thailand. They might have been infected in Thailand while stayed there. Two cases had the diet of raw freshwater fish before. Three cases received emergency laparotomy due to peritonitis and two cases were found of enteritis cystica profunda. According to the route of transmission, freshwater and brackish-water fish may act as the intermediate host of the parasite. The most simple and convenient method of diagnosing capillariasis is stool examination. Two cases were diagnosed by histology. Mebendazole or albendezole 200 mg orally twice a day for 20-30 d is the treatment of choice. All the patients were cured, and relapses were not observed within 12 mo. PMID- 15285026 TI - Accuracy of indocyanine green pulse spectrophotometry clearance test for liver function prediction in transplanted patients. AB - AIM: To investigate whether the non-invasive real-time Indocyanine green (ICG) clearance is a sensitive index of liver viability in patients before, during, and after liver transplantation. METHODS: Thirteen patients were studied, two before, three during, and eight following liver transplantation, with two patients suffering acute rejection. The conventional invasive ICG clearance test and ICG pulse spectrophotometry non-invasive real-time ICG clearance test were performed simultaneously. Using linear regression analysis we tested the correlation between these two methods. The transplantation condition of these patients and serum total bilirubin (T. Bil), alanine aminotransferase (ALT), and platelet count were also evaluated. RESULTS: The correlation between these two methods was excellent (r(2)=0.977). CONCLUSION: ICG pulse spectrophotometry clearance is a quick, non-invasive, and reliable liver function test in transplantation patients. PMID- 15285027 TI - Prevalence of subclinical hepatic encephalopathy in cirrhotic patients in China. AB - AIM: Subclinical hepatic encephalopathy (SHE) is a common complication of liver diseases. The aim of this study was to find out the normal value of psychometric test and to investigate the prevalence of SHE in Chinese patients with stabilized hepatic cirrhosis. METHODS: Four hundred and nine consecutive cirrhotic patients without overt clinical encephalopathy were screened for SHE by using number connection test part A (NCT-A) and symbol digit test (SDT). SHE was defined as presence of at least one abnormal psychometric test. The age-corrected normal values were defined as the mean+/-2 times standard deviation (2SD), and developed in 356 healthy persons as normal controls. Four hundred and sixteen patients with chronic viral hepatitis were tested as negative controls to assess the diagnostic validity of this test battery. RESULTS: There was no significant difference in NCT scores and SDT quotients between healthy controls and chronic hepatitis group (P>0.05). In all age subgroups, the NCT and SDT measurements of cirrhotic patients differed significantly from those of the controls (P<0.05). When mean+/ 2SD of SDT and NCT measurements from healthy control group was set as the normal range, 119 cirrhotic patients (29.1%) were found to have abnormal NCT-A and SDT tests, 53 (13.0%) were abnormal only in SDT and 36 (8.8%) only in NCT-A. Taken together, SHE was diagnosed in 208 (50.9%) cirrhotic patients by this test battery. The prevalence of SHE increased from 39.9% and 55.2% in Child-Pugh's grade A and B groups to 71.8% in Child-Pugh's grade C group (P<0.05). After the adjustment of age and residential areas required from the tests, no correlation was found in the rate of SHE and causes of cirrhosis, education level and smoking habit. CONCLUSION: Psychometric tests are simple and reliable indicators for screening SHE among Chinese cirrhotic patients. By using a NCT and SDT battery, SHE could be found in 50.9% of cirrhotic patients without overt clinical encephalopathy. The prevalence of SHE is significantly correlated with the severity of liver functions. PMID- 15285028 TI - Prevalence of a newly identified SEN virus in China. AB - AIM: To establish nested-PCR methods for the detection of SENV-D and SENV-H and to investigate the epidemiology of SEN virus in China. METHODS: According to published gene sequences, primers from the conserved region were designed. Then, 135 samples from healthy voluntary blood donors and 242 samples from patients with various forms of liver disease were detected by nested-PCR of SENV-D/H. Some PCR products were cloned and sequenced. RESULTS: By sequencing, the specificity of genotype-specific PCR was confirmed. SENV-D/H DNA was detected in 31% of the blood donors, which was higher than those in America and Italy (2%), and in Japan and Taiwan (15-20%). The prevalence of SENV-D/H viremia was significantly higher in patients with hepatitis B and hepatitis C than in blood donors (59-85% vs 31%, P<0.05). The prevalence among patients with non-A-E hepatitis was significantly higher than among blood donors (68% vs 31%, P<0.01), and equivalent to that among patients with hepatitis B and C. CONCLUSION: Nested-PCR with genotype-specific primers could serve as a useful SENV screening assay. SENV has the same transmission modes as HBV and HCV. The high prevalence in patients with non-A-E hepatitis may attribute to the transmission modes, and SENV may not serve as the causative agents. PMID- 15285029 TI - E-cadherin and calretinin as immunocytochemical markers to differentiate malignant from benign serous effusions. AB - AIM: To investigate the expressions of E-cadherin and calretinin in exfoliated cells of serous effusions and evaluate their values in distinguishing malignant effusions from benign ones. METHODS: Fresh serous effusion specimens were centrifuged and exfoliated cells were collected. Cells were then processed with a standardized procedure, including paraformaldehyde fixation, BSA-PBS solution washing and smears preparation. E-cadherin and calretinin were detected by immunocytochemistry (ICC). RESULTS: In the exfoliated cells of serous effusions, most of carcinoma cells only expressed E-cadherin, and most of mesothelial cells only expressed calretinin, and benign cells (lymphocytes and granulocytes) did not express either of them. For E-cadherin, 85.7% (30/35) of malignant effusions and 8.1% (3/37) of benign fluids were ICC-positive (P<0.001). The sensitivity of E-cadherin ICC in the diagnosis of malignant effusions was 85.7%, specificity 91.9%, and diagnostic rate 88.9%. For calretinin, 94.6% (35/37) of benign effusions and 11.4% (4/35) of malignant effusions were ICC-positive (P<0.001). The sensitivity of calretinin ICC in the diagnosis of benign effusions was 94.6%, specificity 88.6%, and diagnostic rate 91.7%. For diagnosis of benign and malignant effusions by combining E-cadherin ICC and calretinin ICC, the specificities were up to 100% and 97.1%, respectively. CONCLUSION: E-cadherin ICC and calretinin ICC are sensitive and specific in differential diagnosis of benign and malignant serous effusion specimens and specificities are evidently improved when both markers are combined. PMID- 15285030 TI - Assessment of correlation between serum titers of hepatitis C virus and severity of liver disease. AB - AIM: The significance of hepatitis C virus (HCV) serum titers has been examined in several clinical situations. There is much evidence that patients with a lower viral load have better response rates to anti-viral therapy compared to those with higher levels. Moreover, a direct association has been observed between serum titers of HCV and transmission rates of the virus. The aim of the present study was to determine if there was any correlation between HCV viral load and the severity of liver disease. METHODS: Fifty patients with HCV infection were included in the study. These comprised of 34 subjects with a history of alcohol use and 16 non-alcoholics. Quantitative serum HCV RNA assay was carried out using the branched DNA (bDNA) technique. Linear regression analysis was performed between serum viral titers and liver tests. In addition, for the purpose of comparison, the subjects were divided into two groups: those with low viral titers (< or = 50 genome mEq/mL) and high titers (> 50 mEq/mL). RESULTS: All subjects were men, with a mean +/- SD age of 47 +/- 7.8 years. The mean HCV RNA level in the blood was 76.3 x 10(5) +/- 109.1 genome equivalents/mL. There was no correlation between HCV RNA levels and age of the patients (r = 0.181), and the history or amount (g/d) of alcohol consumption (r = 0.07). Furthermore, no correlation was observed between serum HCV RNA levels and the severity of liver disease as judged by the values of serum albumin (r = 0.175), bilirubin (r = 0.217), ALT (r = 0.06) and AST (r = 0.004) levels. Similarly, no significant difference was observed between patients with low viral titers and high titers with respect to any of the parameters. CONCLUSION: Our results indicate that the severity of liver disease is independent of serum levels of hepatitis C virus. These findings are important since they have a direct impact on the current debate regarding the role of direct cytopathic effect of hepatitis C virus versus immune-mediated injury in the pathogenesis of HCV-related liver damage. PMID- 15285031 TI - Hypertrophied anal papillae and fibrous anal polyps, should they be removed during anal fissure surgery? AB - AIM: Hypertrophied anal papillae and fibrous anal polyps are not given due importance in the proctology practice. They are mostly ignored being considered as normal structures. The present study was aimed to demonstrate that hypertrophied anal papillae and fibrous anal polyps could cause symptoms to the patients and that they should be removed in treatment of patients with chronic fissure in anus. METHODS: Two groups of patients were studied. A hundred patients were studied in group A in which the associated fibrous polyp or papillae were removed by radio frequency surgical device after a lateral subcutaneous sphincterotomy for relieving the sphincter spasm. Another group of a hundred patients who also had papillae or fibrous polyps, were treated by lateral sphincterotomy alone. They were followed up for one year. RESULTS: Eighty-nine percent patients from group A expressed their satisfaction with the treatment in comparison to only 64% from group B who underwent sphincterotomy alone with the papillae or anal polyps left untreated. Group A patients showed a marked reduction with regard to pain and irritation during defecation (P = 0.0011), pricking or foreign body sensation in the anus (P = 0.0006) and pruritus or wetness around the anal verge (P = 0.0008). CONCLUSION: Hypertrophied anal papillae and fibrous anal polyps should be removed during treatment of chronic anal fissure. This would add to effectiveness and completeness of the procedure. PMID- 15285032 TI - Lack of evidence for leukocyte maternal microchimerism in primary biliary cirrhosis. AB - AIM: It is reasonable to assume that microchimerism could also be involved in the induction of primary biliary cirrhosis (PBC). However, previous reports investigated only fetus-microchimerism in women patients. Maternal microchimerism has not been investigated until now. The current study aimed to clear either maternal microchimerism was involved in the pathogenesis of PBC or not. METHODS: We used fluorescence in situ hybridization on paraffin-embedded tissue (We called "Tissue-FISH".) to determine whether maternal cells infiltrated in male patients who were diagnosed as having PBC. Tissue-FISH was performed by using both X and Y specific probes on the biopsy liver sample of 3 male PBC patients. RESULTS: Infiltrating lymphocytes demonstrated both X and Y signals in all 3 male patients. CONCLUSION: Maternal microchimerism dose not play a significant role in PBC. PBC may not relate to fetus and maternal microchimerism. PMID- 15285033 TI - Gastrointestinal stromal tumor: Computed tomographic features. AB - AIM: Gastrointestinal stromal tumor (GIST) is a rare type of cancer. Computed tomography (CT) is an imaging modality of choice for diagnosing GIST. The aim of this retrospective study was to review the CT imaging features of 17 GIST patients. METHODS: From 1995 to 2003, there were 47 patients with pathologically proven GISTs at our hospital. Of these, 17 patients underwent preoperative CT. We collected and analyzed these CT images. The CT imaging features included tumor diameter, number and location, tumor margin, location of metastasis, hounsfield units of tumor and effect of contrasts. In addition, we also recorded the surgical findings, including complications, tumor size and location for comparative analysis. RESULTS: The results showed that 12 (70%) tumors were located in the stomach and five (30%) were located in the jejunum mesentery. GISTs were extraluminal in 12 (70%) patients. The tumor margins of 13 (76%) tumors were well defined and irregular in four (24%). The effect of contrast enhancement on GIST CT imaging was homogenous enhancement in 13 (76%) and heterogeneous enhancement in four (24%). The hounsfield units (HU) were 30.41 +/ 5.01 for precontrast images and postcontrast hounsfield units were 51.80 +/- 9.24. CONCLUSION: The stomach was the commonest site of GIST occurrence among our patients. The CT features of GIST were well-defined tumor margins, homogenous enhancement on postcontrast CT images. PMID- 15285034 TI - Effect of parenteral and enteral nutrition combined with octreotide on pancreatic exocrine secretion of patients with pancreatic fistula. AB - AIM: To evaluate the effect of parenteral and enteral nutrition combined with octreotide on pancreatic exocrine secretion of the patients with pancreatic fistula. METHODS: Pancreatic juice, drained directly from the pancreatic fistula, was collected, and the volume, protein, amylase, HCO(3)(-), K(+), Na(+) and Cl(-) were determined on d 1, 4 and 7 before and after 7-d treatment with octreotide, respectively. RESULTS: No differences in exocrine pancreatic secretion were observed during the enteral and parenteral nutrition period (t = 2.03, P > 0.05); there were significant decreases in pancreatic juice secretion volume, protein, amylase, HCO(3)(-), K(+), Na(+) and Cl(-) after parenteral and enteral nutrition combined with octreotide compared with octreotide pretreatment (t = 4.14, P < 0.05). CONCLUSION: There is no stimulatory effect on the pancreatic secretion by intrajejunal nutrition and parenteral nutrition. Octreotide is effective on the reduction of pancreatic fistula output. PMID- 15285035 TI - Risk factors for alcoholic liver disease in China. AB - AIM: To examine the association of daily alcohol intake, types of alcoholic beverage consumed, drinking patterns and obesity with alcoholic liver disease in China. METHODS: By random cluster sampling and a 3-year follow-up study, 1 270 alcohol drinkers were recruited from different occupations in the urban and suburban areas of Xi'an City. They were examined by specialists and inquired for information on: Medical history and family medical history, alcohol intake, types of alcoholic beverage consumed, drinking patterns by detailed dietary questionnaires. Routine blood tests and ultrasonography were done. RESULTS: Multivariate analysis showed that: (1) The risk threshold for developing alcoholic liver disease was ingestion of more than 20 g alcohol per day, keeping on drinking for over 5 years in men. The highest OR was at the daily alcohol consumption > or =160 g, the occurrence rate of ALD amounted to 18.7% (P<0.01). No ALD occurred when ingestion of alcohol was less than 20 g per day. (2) 87.9% of all drank only at mealtimes. The cumulative risk of developing ALD was significantly higher in those individuals who regularly drank alcohol without food than in those who drank only at mealtimes, especially for those who regularly drank hard liquors only and multiple drinks (P<0.05). (3) The alcohol consumption in those with BMI > or =25 was lower than in those with BMI<25, but the risk increased to 11.5%, significantly higher than that of general population, 6.5% (P<0.01). (4) Abstinence and weight reduction could benefit the liver function recovery. CONCLUSION: In the Chinese population the ethanol risk threshold for developing ALD is 20 g per day, and this risk increases with increased daily intake. Drinking 20 g of ethanol per day and for less than 5 years are safe from ALD. Drinking alcohol outside mealtimes and drinking hard liquors only and multiple different alcohol beverages both increase the risk of developing ALD. Obesity also increases the risk. Abstinence and weight reduction will directly affect the prognosis of ALD. Doctor's strong advice might influence the prognosis indirectly. PMID- 15285036 TI - Effect of insulin on hyperkalemia during anhepatic stage of liver transplantation. AB - AIM: To investigate the effectiveness of insulin on decreasing serum potassium concentration during anhepatic stage of orthotopic liver transplantation. METHODS: Sixteen patients with serum potassium concentrations greater than 4.0 mmol/L at the onset of anhepatic stage were randomized into two groups. The patients in control group (n = 8) received no treatment, while those in treatment group (n = 8) received an intravenous bolus injection of regular insulin (20 U) 10 min into the anhepatic stage, followed by a glucose infusion (500 mL 50 g/L dextrose) over 15 min. RESULTS: In control group, potassium concentration underwent no changes whereas in treatment group, it decreased from 4.8+/-0.48 mmol/L to 4.19+/-0.55 mmol/L (mean+/-SD) within 15 min and to 3.62+/-0.45 mmol/L 60 min after the therapy. The potassium concentration was lower in treatment group than in control group within 30 min of treatment (3.94+/-0.57 vs 4.47+/ 0.42 mmol/L, respectively; P<0.05), and increased similarly 30 s after graft reperfusion in both groups of patients, but remained lower in treatment group (5.81+/-1.78 vs 7.44+/-1.75 mmol/L, respectively; P<0.05). The potassium concentration returned to pre-reperfusion levels within 5 min after graft reperfusion. CONCLUSION: In patients undergoing orthotopic liver transplantation, the administration of insulin rapidly decreases serum potassium concentration even in the absence of the liver, suggesting an important contribution by extrahepatic tissues in insulin-stimulated uptake of potassium. PMID- 15285037 TI - Receptor binding characteristics and cytotoxicity of insulin-methotrexate. AB - AIM: To characterize the receptor binding affinity and cytotoxicity of insulin methotrexate (MTX) for the potential utilization of insulin as carriers for carcinoma target drugs. METHODS: MTX was covalently linked to insulin. Insulin MTX conjugate was purified by Sephadex G-25 column and analyzed by high performance liquid chromatography. Hepatocellular carcinoma cell membrane fractions were isolated by sucrose density gradient centrifugation. Competitive displacement of (125)I-insulin with insulin and insulin-MTX binding to insulin receptors were carried out. Cytoreductive effect of insulin-MTX on human hepatoma BEL7402 cells and human hepatocyte cell line HL7702 was evaluated using the MTT assay. RESULTS: Insulin-MTX competed as effectively as insulin with (125)I insulin for insulin receptors. The values of Kd for insulin-MTX and insulin were 93.82+/-19.32 nmol/L and 5.01+/-1.24 nmol/L, respectively. The value of Kd for insulin-MTX was significantly increased in comparison with insulin (t=7.2532, n=4, P<0.005). Insulin-MTX inhibited the growth of human hepatoma cells (BEL7402) almost as potently as MTX. The inhibitory effect reached a peak on the 5 th day when the growth of cells was inhibited by 79% at a concentration of 5.0 microg/mL insulin-MTX. Treatment with 5.0 microg/mL of MTX and 5.0 microg/mL of insulin-MTX merely resulted in inhibition of HL7702 cells by 31.5% and 7.8% on the 5 th day. CONCLUSION: Insulin-MTX specifically recognizes insulin receptors and inhibits the growth of BEL7402 cells. These results suggest that insulin can be used as a carrier in receptor mediated carcinoma-targeting therapy. PMID- 15285038 TI - Analysis of multiple factors of postsurgical gastroparesis syndrome after pancreaticoduodenectomy and cryotherapy for pancreatic cancer. AB - AIM: To explore the etiology, pathogenesis, diagnosis, and treatment of postsurgical gastroparesis syndrome (PGS) after pancreatic cancer cryotherapy (PCC) or pancreatico-duodenectomy (PD), and to analyze the correlation between the multiple factors and PGS caused by the operations. METHODS: Clinical data of 210 patients undergoing PD and 46 undergoing PCC were analyzed retrospectively. RESULTS: There were 31 (67%, 31/46) patients suffering PGS in PCC group, including 29 with pancreatic head and uncinate tumors and 2 with pancreatic body and tail tumors. Ten patients (4.8%, 10/210) developed PGS in PD group, which had a significantly lower incidence of PGS than PCC group (chi= 145, P<0.001). In PCC group, 9 patients with PGS were managed with non-operative treatment (drugs, diet, nasogastric suction, etc.), and one received reoperation at the 16th day, but the symptoms were not relieved. In PD group, all the patients with PGS were managed with non-operative treatment. The PGS in patients undergoing PCC had close association with PCC, tumor location, but not with age, gender, obstructive jaundice, hypoproteinemia, preoperative gastric outlet obstruction and the type and number of gastric biliary tract operations. The mechanisms of PGS caused by PD were similar to those of PGS following gastrectomy. The damage to interstitial cells of Cajal might play a role in the pathogenesis of PGS after PCC, for which multiple factors were possibly responsible, including ischemic and neural injury to the antropyloric muscle and the duodenum after freezing of the pancreatico duodenal regions or reduced circulating levels of motilin. CONCLUSION: PGS after PCC or PD is induced by multiple factors and the exact mechanisms, which might differ between these two operations, remain unknown. Radiography of the upper gastrointestinal tract and gastroscopy are main diagnostic modalities for PGS. Non-operative treatments are effective for PGS, and reoperation should be avoided in patients with PGS caused by PCC. PMID- 15285039 TI - Simultaneous detection of HBV and HCV by multiplex PCR normalization. AB - AIM: To design and establish a method of multiplex PCR normalization for simultaneously detecting HBV and HCV. METHODS: Two pairs of primers with a 20 bp joint sequence were used to amplify the target genes of HBV and HCV by two rounds of amplification. After the two rounds of amplification all the products had the joint sequence. Then the joint sequence was used as primers to finish the last amplification. Finally multiplex PCR was normalized to a single PCR system to eliminate multiplex factor interference. Four kinds of nucleic acid extraction methods were compared and screened. A multiplex PCR normalization method was established and optimized by orthogonal design of 6 key factors. Then twenty serum samples were detected to evaluate the validity and authenticity of this method. RESULTS: The sensitivity, specificity, diagnostic index and efficiency were 83.3%, 70%, 153.3% and 72.2%, respectively for both HBsAg and anti-HCV positive patients, and were 78.6%, 80%, 258.6% and 79.2%, respectively for HBsAg positive patients, and were 75%, 90%, 165% and 83.3%, respectively for anti-HCV positive patients. CONCLUSION: The multiplex PCR normalization method shows a broad prospect in simultaneous amplification of multiple genes of different sources. It is practical, correct and authentic, and can be used to prevent and control HBV and HCV. PMID- 15285040 TI - Role of endoscopic miniprobe ultrasonography in diagnosis of submucosal tumor of large intestine. AB - AIM: To evaluate the role of miniprobe ultrasonography under colonoscope in the diagnosis of submucosal tumor of the large intestine, and to determine its imaging characteristics. METHODS: Thirty-five patients with submucosal tumors of the large intestine underwent miniprobe ultrasonography under colonoscope. The diagnostic results of miniprobe ultrasonography were compared with pathological findings of specimens by biopsy and surgical resection. RESULTS: Lipomas were visualized as hyperechoic homogeneous masses located in the submucosa with a distinct border. Leiomyomas were visualized as hypoechoic homogeneous mass originated from the muscularis propria. Leiomyosarcomas were shown with inhomogeneous echo and irregular border. Carcinoids were presented as submucosal hypoechoic masses with homogenous echo and distinct border. Lymphangiomas were shown as submocosal hypoechoic masses with cystic septal structures. Malignant lymphomas displayed as hypoechoic masses from mucosa to muscularis propria, while pneumatosis cystoids intestinalis originated from submucosa with a special sonic shadow. One large leiomyoma was misdiagnosed as leiomyosarcoma. CONCLUSION: Endoscopic miniprobe ultrasonography can provide precise information about the size, layer of origin, border of submucosal tumor of the large intestine and has a high accuracy in the diagnosis of submucosal tumor of the large intestine. Pre operative miniprobe ultrasonography under colonoscope may play an important role in the choice of therapy for submucosal tumor of the large intestine. PMID- 15285041 TI - Gastrointestinal autonomic nerve tumors: a surgical point of view. AB - AIM: Gastrointestinal autonomic nerve tumors are uncommon stromal tumors of the intestinal tract. Their histological appearance is similar to that of other gastrointestinal stromal tumors. We report two cases and performed an analysis of the literature by comparing our findings with the available case reports in the medical literature. METHODS: Two patients were admitted with abdominal tumor masses. One occurred in the stomach with large multiple liver metastases and the second originated in Meckel's diverticulum. The latter site has never been reported previously. Both patients underwent surgery. In one patient gastrectomy, right liver resection and colon transversum resection were performed to achieve aggressive tumor debulking. In the other patient the tumor bearing diverticulum was removed. RESULTS: Postoperative recovery of both patients was uneventful. Histological examination, immunohistochemical analysis and electron microscopy revealed the diagnosis of a gastrointestinal autonomic nerve tumor. The patient with the tumor in Meckel's diverticulum died 6 mo after surgery because of pneumonia. The patient with liver metastases have been alive 13 years after initial tumor diagnosis and 7 years after surgery with no evidence of tumor progression. In light of our results, we performed a thorough comparison with available literature reports. CONCLUSION: Radical surgical resection of gastrointestinal autonomic nerve tumors seems to be the only available curative approach to date, and long term survival is possible even in large metastasized tumors. PMID- 15285042 TI - Granular cell tumor of colon: report of a case and review of literature. AB - Granular cell tumor (GCT) is uncommon in the colon and rectum. Here we report a case of GCT in the transverse colon. A 48-year-old male patient underwent a screening colonoscopy. A yellowish sessile lesion, about 4 mm in diameter, was found in the transverse colon. An endoscopic snare resection was performed without complication. Histological examination revealed the tumor consisted of plump neoplastic cells with abundant granular eosinophilic cytoplasm containing acidophilic periodic acid Schiff-positive, diastase-resistant granules. Immunohistochemical analysis showed the tumor cells expressed S-100 protein and neuron-specific enolase. Thus, the resected tumor was diagnosed as a GCT. Since GCTs are usually benign, endoscopic resection constitutes an easy and safe treatment. Colonoscopists should consider the possibility of GCT in the differential diagnosis of submucosal tumors of the colon. PMID- 15285043 TI - A case of leptospirosis simulating colon cancer with liver metastases. AB - We report a case of a 61-year-old man who presented with fatigue, abdominal pain and hepatomegaly. Computed tomography (CT) of the abdomen showed hepatomegaly and multiple hepatic lesions highly suggestive of metastatic diseases. Due to the endoscopic finding of colon ulcer, colon cancer with liver metastases was suspected. Biochemically a slight increase of transaminases, alkaline phosphatase and gammaglutamyl transpeptidase were present; alpha-fetoprotein, carcinoembryogenic antigen and carbohydrate 19-9 antigen serum levels were normal. Laboratory and instrumental investigations, including colon and liver biopsies revealed no signs of malignancy. In the light of spontaneous improvement of symptoms and CT findings, his personal history was reevaluated revealing direct contact with pigs and their tissues. Diagnosis of leptospirosis was considered and confirmed by detection of an elevated titer of antibodies to leptospira. After two mo, biochemical data, CT and colonoscopy were totally normal. PMID- 15285045 TI - Dysphagia lusorium in elderly: a case report. AB - AIM: Late onset of dysphagia due to vascular abnormalities is a rare condition. We aimed to present a case of right subclavian artery abnormalities caused dysphagia in the elderly. METHODS: A 68-year-old female was admitted with dysphagia seven months ago. Upper endoscopic procedures and routine examinations could not demonstrate any etiology. Multislice computed thorax tomography was performed for probable extra-esophagial lesions. RESULTS: Multislice computed thorax tomography showed right subclavian artery abnormality and esophagial compression with this aberrant artery. CONCLUSION: Causes of dysphagia in the elderly are commonly malignancies, strictures and/or motility disorders. If routine examinations and endoscopic procedures fail to show any etiology, rare vascular abnormalities can be considered in such patients. Multislice computed tomography is a usefull choice in such conditions. PMID- 15285044 TI - Acute esophageal necrosis and liver pathology, a rare combination. AB - Acute esophageal necrosis (AEN) or "black esophagus" is a clinical condition found at endoscopy. It is a rare entity the exact etiology of which remains unknown. We describe a case of 'black esophagus', first of its kind, in the setting of liver cirrhosis and hepatic encephalopathy. PMID- 15285047 TI - Editorial. The late professor Gen Matsumoto. AB - At the opening of this special issue dedicated to the work of the late Prof. Gen Matsumoto, I would like to look back at Gen Matsumoto's research life and to personally share with the readers of this journal his dream of creating a real brain-like computer. Gen Matsumoto had been planning to create a brain-like computer for thirty years. I have been most fortunate in being able to personally see progress toward Gen Matsumoto's ultimate goal, and have been inspired by his persistence, perseverance, and full belief that one day a brain-like computer would be operative. Gen Matsumoto left a definite impression on me so much, so that I considered him my reference of life rather than just my research partner. PMID- 15285046 TI - Minute gastric carcinoid tumor with regional lymph node metastasis: a case report and review of literature. AB - We have encountered an unusual case of gastric carcinoid tumor. Gastroscopic examination of this 32-year-old male patient showed a smooth protrusion at the greater curvature of the gastric body with a central depression, identified by subsequent biopsy as carcinoma. The patient had a normal serum gastrin level and was negative for anti-parietal cell antibody. Histological examination of the resected gastric tissues showed that the tumor was a carcinoid, 0.3 cm x 0.3 cm in size with only one regional lymph node metastasis. We reviewed the pathogenesis, clinical presentation, diagnosis and treatment of gastric carcinoids and raise the possibility of being a lymph vessel-related metastasis even for a minute carcinoid tumor. Sentinel lymph node biopsy is recommended for surgery of minute carcinoid tumors. PMID- 15285048 TI - Prologue--The pioneering work of the late professor Gen Matsumoto. AB - The late Professor Gen Matsumoto not only accomplished pioneering work of neuroscience such as nerve excitation as a dissipative structure in far-from equilibrium systems but also inspired many students and young researchers by his sincere and attractive personality. PMID- 15285049 TI - On the conduction velocity of nonmyelinated nerve fibers. AB - Nerve impulse conduction in nonmyelinated nerve fibers is analyzed by considering this process as a direct consequence of the coexistence of two structurally distinct regions, active and resting. Assuming that the active (i.e. swollen) region of the fiber is in direct contact with the resting (i.e. shrunken) region, a simple procedure for deriving the conduction velocity equation is described. The physico-chemical significance of the quantities in this velocity equation is briefly discussed. PMID- 15285050 TI - The brain-computer: origin of the idea and progress in its realization. AB - The Brain-Computer is a physical analogue of a real organism which uses both a brain-inspired memory-based architecture and an output-driven learning algorithm. This system can be realized by creating a scaled-down model car that learns how to drive by heuristically connecting image processing with behavior control. This study proves that learning efficiency progresses rapidly when the acquired behaviors are prioritized. We develop a small real-world device that moves about purposefully in an artificial environment. The robot uses imaging information acquired through its random actions to make a mental map. This map, then, provides the cognitive structure for acquiring necessary information for autonomous behavior. PMID- 15285051 TI - Output-driven operation and memory-based architecture principles embedded in a real-world device. AB - Two principles of neurocomputational design are implemented into an autonomous real-world device, such as a helicopter. The helicopter has a motivational component towards emitting motor responses in a manner similar to a fledging bird. We expect these two principles, together with an understanding of integrative brain activity and memory in which functionally and selectively distributed neural networks operate in vivo will eventually lead to the embodiment of cognition in a brain-like computer as an engineering counterpart of a real brain. PMID- 15285052 TI - Synchronization of neural oscillations as a possible mechanism underlying episodic memory: a study of theta rhythm in the hippocampus. AB - Rat hippocampal cells exhibit characteristic phase-dependent firings when oscillation in frequency range around 8Hz is present. Based on the hypothesis that theta phase coding is generated by synchronization of neural activities, an autoassociative network model of the hippocampus and the entorhinal cortex was analyzed to explore mechanisms underlying episodic memory. Phase coding in theta rhythm enables instantaneous acquisition of experienced events including temporal and spatial contents. Further comparison with electrophysiological data from both rodents and primates suggests a possible role of theta oscillations in memory encoding and online processing of episodic events. PMID- 15285053 TI - Chaotic itinerancy as a mechanism of irregular changes between synchronization and desynchronization in a neural network. AB - We investigate the dynamic character of a network of electrotonically coupled cells consisting of class I point neurons, in terms of a finite dimensional dynamical system. We classify a subclass of class I point neurons, called class I* point neurons. Based on this classification, we use a reduced Hindmarsh-Rose (H-R) model, which consists of two dynamical variables, to construct a network model consisting of electrotonically coupled H-R neurons. Although biologically simple, the system is sufficient to extract the essence of the complex dynamics, which the system may yield under certain physiological conditions. The network model produces a transitory behavior as well as a periodic motion and spatio temporal chaos. The transitory dynamics that the network model exhibits is shown numerically to be chaotic itinerancy. The transitions appear between various metachronal waves and all-synchronization states. The network model shows that this transitory dynamics can be viewed as a chaotic switch between synchronized and desynchronized states. Despite the use of spatially discrete point neurons as basic elements of the network, the overall dynamics exhibits scale-free activity including various scales of spatio-temporal patterns. PMID- 15285054 TI - Functional relevance of 'excitatory' gaba actions in cortical interneurons: a dynamical systems approach. AB - The non-classical, but frequently reported behavior of GABA(A) receptor-mediated excitation in mature CNS has long been regarded as a puzzle. We theorize that the function of cortical GABAergic interneurons, which might constitute a subsystem of brain's GABA interneurons, is their ability of switching from inhibitory action to excitatory action depending on the level of spatio-temporal activity in progress. From the perspective of a dynamical systems approach, such "excitatory" GABAergic responses may serve to temporarily invoke attractor-like memories when extensively activated by, for example, top-down signals as category information or attention, while an ongoing background state of GABA changes its action to inhibition, returning the dynamical nature of the memory structure back to attractor ruins. PMID- 15285055 TI - Chaotic spiking in the Hodgkin-Huxley nerve model with slow inactivation of the sodium current. AB - The Hodgkin-Huxley (HH) equations with a modification in which the inactivation process (h variable) of sodium channels is slightly slowed down are investigated. It is shown that this slight modification changes the HH dynamics to a completely different one, with chaotic spiking and very long interspike intervals appearing in a generic manner, although the initiation mechanism of repetitive firing is a simple Hopf bifurcation. PMID- 15285056 TI - Scaling laws for myelinated axons derived from an electrotonic core-conductor model. AB - A macroscopic cable equation, which describes the passive linear ("electrotonic") response of a myelinated axon, was previously derived from a segmented cable equation using Keller's two-space homogenization method [Basser, PJ, Med. and Biol. Comput., 1993, Vol. 31, pp. S87-S92]. Here we use the space and length constants of this averaged cable equation to predict classical scaling laws that govern relationships among the inner and outer diameters of the axon's myelin sheath and the distance separating adjacent nodes of Ranvier. These laws are derived by maximizing the characteristic speed of an electrical disturbance along the axon, i.e., the ratio of the characteristic length and the characteristic time constants of the macroscopic cable, subject to the constraint that the nodal width is constant. Using this result, it is also possible to show that all myelinated axons are equally fault tolerant. No free parameters were used in this analysis; all variables and physical constants used in these calculations were taken from published experimental data. PMID- 15285057 TI - [A prospective study about an imaging quality of low dosage of contrast media with single-detector CT]. AB - PURPOSE: The steadily elevating cost of the contrast-enhanced CT has been a problem in last decade. One approach to curtail the cost is to reduce the amount of contrast media (CM). The purpose of this study was to examine the possibility of reducing the volume of CM in single-detector CT (SDCT) without compromising diagnostic ability. MATERIALS AND METHODS: One hundred ml of Iohexol 300 and 75 ml of Ioversol 320 were compared in a prospective CT study about their imaging quality. One hundred patients were randomly divided into two parallel groups (A and B). The former agent was used for group A and the latter for B. CT attenuations of the aorta, central vein, spleen and kidney were measured and each imaging quality was reviewed by three radiologists. RESULTS: CT attenuations and the imaging quality were significantly superior (p<0.05) in A group, however, comparing in the condition under 50 kg patients, they showed no significant differences and the quality was tolerable for diagnosis in both group. CONCLUSION: This demonstrates the difficulty to perform the high-quality CT with a reduced CM in SDCT. However, this does not negate the diagnostic ability of low dose of CM, but reflects the importance of determining the acceptable lowest doze of CM for diagnosis. PMID- 15285059 TI - [Patient information. Gastroesophageal reflux disease]. PMID- 15285058 TI - Recurrent isolation of an uncommon yeast, Candida pararugosa, from a sarcoma patient. AB - A yeast was repeatedly isolated from the saliva of a sarcoma patient. A relatively uncommon species, Candida maris, was identified based on the API 20C profile. The yeast species most frequently obtained from the patient's mother and from clinic staff was Candida albicans. A comparison of the yeast obtained from the patient with the type strain of C. maris strongly suggested that the former was not representative of C. maris. Analysis of partial ribosomal DNA sequences of the patient strain and from the type strain of C. maris showed that the two are phylogenetically not closely related. The patient strain was very close to Candida pararugosa, a relatively uncommon asporogenous yeast. DNA reassociation studies among C. pararugosa and patient isolates showed that they were conspecific. We could not determine the source of the yeast infection. This case will alert hospital staff to be aware of the possibility of unexpected environmental microorganisms as causes of infections, colonizations and persistent environmental contamination events in immunocompromised patients. PMID- 15285060 TI - Medicare program; interest calculation. Final Rule. AB - This final rule changes the way we calculate interest on Medicare overpayments and underpayments to providers, suppliers, health maintenance organizations, competitive medical plans, and health care prepayment plans to be more reflective of current business practices. This change reduces the amount of interest assessed on overpayments and underpayments and simplifies the way the interest is calculated. This change in the way we calculate interest also applies to Medicare Secondary Payer debt. PMID- 15285061 TI - New Pennsylvania law requires error reporting for learning purposes. PMID- 15285062 TI - Study establishes potential utility of biomarkers in ovarian cancer diagnostics. PMID- 15285063 TI - Study explores case for routine prenatal and pre-pregnancy thyroid screening. PMID- 15285064 TI - New treatment for thyroid ablation as effective as traditional therapy without side-effects of hypothyroidism. PMID- 15285065 TI - The Registry of the International Society for Heart and Lung Transplantation: twenty-first official adult heart transplant report--2004. PMID- 15285066 TI - The Registry of the International Society for Heart and Lung Transplantation: twenty-first official adult lung and heart-lung transplant report--2004. PMID- 15285068 TI - Recent references. PMID- 15285067 TI - The changing face of heart and lung transplantation: Presidential Address, 2003 Annual Meeting of the International Society for Heart and Lung Transplantation. PMID- 15285069 TI - Recent advances in alpha1-adrenoceptor pharmacology. PMID- 15285070 TI - WHO releases new report on global problem of oral diseases. PMID- 15285071 TI - Primary care intervention reduces suicidal ideation in older patients with depression. PMID- 15285072 TI - On the Szaszian argument. AB - The Szaszian argument claims that psychiatry is a rhetorical enterprise aimed at providing justification for involuntary treatment. Such treatment, the argument holds, is just when provided to those suffering from demonstrable brain lesions, but it is unjust in cases of "mental illness" because such "illnesses" lack objective histopathology and are therefore fictional. It is here argued that this distinction is irrelevant to the morality or immorality of involuntary treatment, since such treatment inevitably rests on a subjective determination of competency or dangerousness, which is not rendered substantially more objective by the criterion of histopathology. The Szaszian argument subscribes to a naturalistic fallacy in this regard, which leads it to inconsistencies in its philosophy of mind. PMID- 15285073 TI - Impaired regulation of blood flow as a risk for inadequate blood flow to the optic nerve under challenge. PMID- 15285074 TI - Informed consent and defenses to a lack of informed consent. PMID- 15285075 TI - Trauma in reproductive-age women. PMID- 15285076 TI - Endarterectomy vs. Angioplasty in Patients with Symptomatic Severe Carotid Stenosis (EVA-3S) Trial. AB - Endarterectomy versus Angioplasty in Patients with Symptomatic Severe Carotid Stenosis (EVA-3S) is a French multicenter, non-inferiority randomized trial with national research organisation funding. In brief, patients are eligible if they have experienced a carotid TIA or non-disabling stroke within 4 months before randomisation and if they have an atherosclerotic stenosis of the region of the ipsilateral carotid bifurcation of 60% or more, as determined by the NASCET method, that investigators believe is suitable for both carotid surgery and angio plasty. Carotid surgery is performed using standard operative techniques. Carotid angioplasty consists of pri-mary stenting with cerebral protection. The primary end-points are: (a) any stroke or death within 30 days of the procedure and (b) any stroke or death within 30 days of the procedure plus ipsilateral stroke. To join the study, a centre must form a multidisciplinary team, including avascular neurologist, a vascular surgeon and an interventionalist. Operator experience must be substantiated through documentation of a sufficient number of cases performed. PMID- 15285077 TI - [Sepsis in the newborn]. PMID- 15285078 TI - Tobacco smoke and involuntary smoking. PMID- 15285079 TI - [Timing of the oral feeding after total laryngectomy]. PMID- 15285080 TI - [Central auditory assessment and its influencing factors]. PMID- 15285081 TI - In memoriam--Mandel E. Cohen, MD (March 8, 1907 - November 19, 2000). PMID- 15285082 TI - Zernike expansion of separable functions of cartesian coordinates. AB - A Zernike expansion over a circle is given for an arbitrary function of a single linear spatial coordinate. The example of a half-plane mask (Hilbert filter) is considered. The expansion can also be applied to cylindrical aberrations over a circular pupil. A product of two such series can thus be used to expand an arbitrary separable function of two Cartesian coordinates. PMID- 15285083 TI - Characterization of single and double fiber-coupled diffusing spheres. AB - A fiber-optic sensor, consisting of an optical fiber with and without a 1.59-mm diameter spherical ceramic tip, inserted into a 19-mm-diameter Spectralon sphere has been characterized. This sensor is evaluated as a large-area omnidirectional sensor. An optical transport measurement system that rotates the sphere about two axes has been designed. The system measured the UV transport efficiency at 351 nm of light impinging on the sphere with an f-6 cone angle. When a bare fiber was placed at the center of the target sphere, the detection sensitivity was biased in the forward direction. The peak of the sensitivity of the inverse integrating sphere shifted from front to back as the bare fiber was withdrawn from the sphere and the numerical aperture of the fiber viewed more of the scattering volume. The response function with respect to the angle of incidence of the dual sphere was much more uniform than that obtained with a bare fiber in the center of the sphere. PMID- 15285084 TI - Color and shape measurement with a three-color photometric stereo system. AB - The purpose of our research has been to develop a system for measurement of microtopography based on the photometric stereo principle. In a previous system, in which we used two illumination directions facing each other, we had difficulties in detecting topography variations perpendicular to the illumination direction. With the new measurement system we avoid this problem by use of three colored and two white illumination modules. The integration problem that occurred when the gradient was known in more than one direction was solved by use of weight functions in the spatial-frequency domain. The results show that true three-dimensional surface height functions can be obtained with the new method. This is a significant improvement from the two-source system, in which we could make only estimates of profiles perpendicular to the illumination direction. To quantitatively evaluate the new measurement system, the results were compared with measurements of a mechanical stylus instrument. Comparisons between mechanical and optical measurements show a coefficient of determination r2 between 0.71 and 0.81 for the new system and r2 between 0.57 and 0.88 for the two source system. PMID- 15285085 TI - High-accuracy measurement of the optical transmittance of optical bulk materials at deep-ultraviolet wavelengths. AB - A measurement setup that is capable of measuring the internal transmittance of fused-silica prisms at 193 nm with a precision better than 0.01%/cm (3sigma) is presented. Its application to materials and wavelengths other than those that were chosen here for demonstration is straightforward. A lack of any standards makes it impossible to determine the absolute accuracy (also called measurement uncertainty) experimentally; however, calculations indicate that it is almost within the same margin as the precision. PMID- 15285086 TI - High-performance nonscanning Fourier-transform spectrometer that uses a Wollaston prism array. AB - A high-performance nonscanning Fourier-transform spectrometer is reported that is composed mainly of a Wollaston prism array and a two-dimensional photodetector array. It is a substantial improvement over existing Wollaston prism based nonscanning Fourier-transform spectrometers because it offers finer spectral resolution and smaller size. Such spectrometers will find important applications in remote chemical and biological sensing, environmental monitoring, medical diagnosis, etc. Experimental results are consistent with theoretical analyses. PMID- 15285087 TI - Simple phase-shifting method in a wedge-plate lateral-shearing interferometer. AB - A simple phase-shifting method in a wedge-plate lateral shearing interferometer is described. Simply moving the wedge plate in an in-plane parallel direction gives the amount of phase shift required for phase-shifting interferometry because the thickness of a wedge plate is not constant and varies along the wedge direction. This method requires only one additional linear translator to move the wedge plate. The required moving distance for a phase shift of the wave front with this method is of the order of a millimeter, whereas the typical moving distance for another method that uses a piezoelectric transducer is of the order of a wavelength. This method yields better precision in controlling the moving distance than do the other methods. PMID- 15285088 TI - Spectroscopic studies of autofluorescence substances existing in human tissue: influences of lactic acid and porphyrins. AB - The influence of lactic acid or porphyrins on the optical properties of tissue fluorophores is investigated by autofluorescence (AF) spectroscopy measurement with a GaN-based ultraviolet laser diode along with Fourier-transform IR (FTIR) spectroscopy measurement. As the lactic-acid concentration becomes dense, the AF peak intensity from elastin and desmosine solutions become wholly weak. A similar reduction in the AF intensity is observed for nicotinamide adenine dinucleotide (NADH) solutions. FTIR analysis indicates that the lactic acid causes the conformational change in elastin and the oxidation of NADH, which can be related to changes in the AF properties. The peak intensity of the tissue fluorophores also becomes weak when porphyrins are added, although the conformational change in each tissue fluorophore is not confirmed from FTIR analysis. Judging from the change in the scattering-light intensity of the excitation source, the observed change mainly originates from the absorption of the excitation source by porphyrins. PMID- 15285089 TI - Dimensionless parameters for the design of optical traps and laser guidance systems. AB - Optical traps are routinely used for the manipulation of neutral particles. However, optical trap design is limited by the lack of an accurate theory. The generalized Lorenz-Mie theory (GLMT) solves the scattering problem for arbitrary particle size and predicts radial forces accurately. Here we show that the GLMT predicts the observed radial and axial forces in a variety of optical manipulators. We also present a dimensionless parameter beta for the prediction of axial forces. Coupled with our correlation for radial escape forces, we now have a set of two simple correlations for the practical design of radiation-force based systems. PMID- 15285090 TI - Liquid-crystal bandpass filter based on the optical rotatory dispersion effect. AB - A bandpass spectral filter and the test measurement results are presented and described. The filter is developed and constructed by use of a ferroelectric liquid-crystal polarization rotator in combination with dispersive quartz optical rotators, and its wavelength selection is mechanically tunable and electrically switchable. PMID- 15285091 TI - Shift-tolerance property of fully phase encryption that uses a joint transform correlator. AB - We technically investigate the robustness of an image encryption technique that uses a virtual phase image and a joint transform correlator (JTC) in the frequency domain. An encrypted image is obtained by the Fourier transform of the product of a virtual phase image, which camouflages the original image, and a random phase image. The resulting image is then decrypted by use of a decrypting key made from the proposed phase assignment rule in order to enhance the level of security. We demonstrate that the encrypted image generated by the proposed JTC based decryption technique is robust to data loss and image shift. PMID- 15285092 TI - Comment on "Holographic characteristics of a 1-mm-thick photopolymer to be used in holographic memories.". AB - We believe there is an error in the calculation of the M/# in a previous paper [Appl. Opt. 42, 7008 (2003)]. From the data provided, we calculate an M/# of 3.8 rather than the reported value of 38 for the 1-mm sample tested. PMID- 15285093 TI - Shrinkage control in a photopolymerizable hybrid solgel material for holographic recording. AB - We report the correction of the shrinkage observed during UV postrecording curing in a holographic solgel material that was recently achieved by the use of various chemical formulations for the composition of the hybrid supporting matrix. We found that a chemical modification of the matrix noticeably attenuates the shrinkage (from 1.3% to 0.4% of the material's initial thickness with the inclusion of just 20% tetramethylorthosilicate), providing a material with improved stability for permanent data storage applications. The holographic properties of samples with different binders are also reported. In addition, a theoretical study has revealed the way by which to compensate for angular deviation in the Bragg condition during UV postrecording by tailoring the binder shrinkage (s), the maximum refractive-index modulation capability of the photosensitive mixture (deltan), or both. PMID- 15285094 TI - Relevance of mask-roughness-induced printed line-edge roughness in recent and future extreme-ultraviolet lithography tests. AB - The control of line-edge roughness (LER) of features printed in photoresist poses significant challenges to next-generation lithography techniques such as extreme ultraviolet (EUV) lithography. Achieving adequately low LER levels requires accurate resist characterization as well as the ability to separate resist effects from other potential contributors to LER. One potentially significant contributor to LER arises from roughness on the mask coupling to speckle in the aerial image and consequently to LER in the printed image. Here I numerically study mask surface roughness and phase roughness to resist LER coupling both as a function of illumination coherence and defocus. Moreover, the potential consequences of this mask effect for recent EUV lithography experiments is studied through direct comparison with experimental through-focus printing data collected at a variety of coherence settings. Finally, the effect that mask roughness will play in upcoming 0.3-numerical-aperture resist testing is considered. PMID- 15285095 TI - Behavior of GeSbTeBi phase-change optical recording media under subnanosecond pulsed laser irradiation. AB - We investigated the variations in reflectivity during the phase transition between amorphous and crystalline states of a Bi-doped GeTe-Sb2Te3 pseudobinary compound film with subnanosecond laser pulses, using a pump-and-probe technique. We also used a two-laser static tester to estimate the onset time of crystallization under 2.0-micros pulse excitation. Experimental results indicate that the formation of a melt-quenched amorphous mark is completed in approximately 1 ns, but that crystalline mark formation on an as-deposited amorphous region requires several hundred nanoseconds. Simple arguments based on heat diffusion are used to explain the time scale of amorphization and the threshold for creation of a burned-out hole in the phase-change film. PMID- 15285097 TI - SIMBAD: a field radiometer for satellite ocean-color validation. AB - A hand-held radiometer, called SIMBAD, has been designed and built specifically for evaluating satellite-derived ocean color. It provides information on the basic ocean-color variables, namely aerosol optical thickness and marine reflectance, in five spectral bands centered at 443, 490, 560, 670, and 870 nm. Aerosol optical thickness is obtained by viewing the Sun disk and measuring the direct atmospheric transmittance. Marine reflectance is obtained by viewing the ocean surface and measuring the upwelling radiance through a vertical polarizer in a geometry that minimizes glitter and reflected sky radiation, i.e., at 45 degrees from nadir (near the Brewster angle) and at 135 degrees in azimuth from the Sun's principal plane. Relative inaccuracy on marine reflectance, established theoretically, is approximately 6% at 443 and 490 nm, 8% at 560 nm, and 23% at 670 nm for case 1 waters containing 0.1 mg m(-3) of chlorophyll a. Measurements by SIMBAD and other instruments during the Second Aerosol Characterization Experiment, the Aerosols-99 Experiment, and the California Cooperative Oceanic Fisheries Investigations cruises agree within uncertainties. The radiometer is compact, light, and easy to operate at sea. The measurement protocol is simple, allowing en route measurements from ships of opportunity (research vessels and merchant ships) traveling the world's oceans. PMID- 15285096 TI - Reducing variability that is due to secondary pigments in the retrieval of chlorophyll a concentration from marine reflectance: a case study in the western equatorial Pacific Ocean. AB - A neural network is developed to retrieve chlorophyll a concentration from marine reflectance by use of the five visible spectral bands of the Sea-viewing Wide Field-of-view Sensor (SeaWiFS). The network, dedicated to the western equatorial Pacific Ocean, is calibrated with synthetic data that vary in terms of atmospheric content, solar zenith angle, and secondary pigments. Pigment variability is based on in situ data collected in the study region and is introduced through nonlinear modeling of phytoplankton absorption as a function of chlorophyll a, b, and c and photosynthetic and photoprotectant carotenoids. Tests performed on simulated yet realistic data show that chlorophyll a retrievals are substantially improved by use of the neural network instead of classical algorithms, which are sensitive to spectrally uncorrelated effects. The methodology is general, i.e., is applicable to regions other than the western equatorial Pacific Ocean. PMID- 15285098 TI - Atmospheric optical turbulence over land in middle east coastal environments: prediction modeling and measurements. AB - Beam intensity scintillations, characterized by a refractive-index structure parameter and caused by variations of macrometeorological features of the coastal atmosphere such as air temperature, wind speed and direction, and relative humidity, are examined theoretically and experimentally. In our theoretical analysis we present two well-known models considered separately for over-water and over-land atmospheric optical communication or imaging channels. By means of comparison with our experiments carried out in midland coastal environments in southern and northern Israel, we show the limitations of the models to predict the refractive-index structure Cn2 parameter for both daytime and nighttime turbulent atmospheres in different coastal zone meteorological conditions. We also present an extension of an existing model with two different practical applications that, as is shown experimentally, can be a good predictor of Cn2 for optical atmospheric paths over midland coastal zones. PMID- 15285099 TI - Phase retrieval from subdivision of the focal plane with a lenslet array. AB - A phase retrieval algorithm derived from subdivision of the complex field at the focal plane is proposed. This subdivision is achieved with a lenslet array at the focal plane in a manner similar to the pyramid wave-front sensor. The phase retrieval algorithm significantly improves the wave-front estimate that can be attained as a linear combination of the aperture images. This phase retrieval algorithm also avoids the twin-image stagnation problem inherent in phase retrieval and phase retrieval in conjunction with the Shack-Hartmann sensor. PMID- 15285100 TI - Spatial structure of dye-doped polymer nanoparticle laser media. AB - The spatial structure of dye-doped polymer-nanoparticle gain media, reported to generate spatially homogeneous single-transverse-mode laser beams, has been examined by electron microscopy in the nanometer scale. It is found that the distribution of the silica nanoparticles in the laser dye-doped polymer is fairly uniform. There is some aggregation of silica particles into loose nanoclusters. However, the nanocluster dimensions are smaller than those necessary to satisfy the conditions for internal interference in the visible spectrum. This explains the absence of macroscopic spatial inhomogeneities in the emission laser beams. PMID- 15285101 TI - Mode properties produced by a corner-cube cavity. AB - The laser mode properties of a corner-cube resonator, in which the corner cube is the key element and a flat mirror is used as the output mirror, are analyzed by a numerical simulating method. Examples of numerical calculations are given to illustrate mode propagation through an optical system. The simulated results agree with the experimental ones. PMID- 15285102 TI - Properties of a phase-conjugate etalon mirror and its application to laser resonator spatial-mode control. AB - The concept of a phase-conjugate etalon mirror consisting of one flat and one aspheric surface is introduced. This new element can be used as an end mirror of a conventional resonator to promote spatial-mode selection and mode shaping. A phase-conjugate etalon designed for the fundamental Gaussian mode is experimentally implemented and tested with a single-mode He-Ne laser. PMID- 15285103 TI - Dispersion compensation by two vertically coupled asymmetric ridge waveguides. AB - Dispersion compensation was demonstrated with a highly dispersive waveguide consisting of two vertically coupled InGaAsP/InP asymmetric ridge waveguides with a large contrast between their refractive indices. A 3.3-ps positively chirped pulse was compressed-at a compression ratio of 62%--by the dispersion compensation produced by the dispersive waveguide. Moreover, experimental and theoretical investigations of dispersion compensation in this waveguide showed that the supermodes of the two coupled asymmetric waveguides play an important role in the observed pulse compression. PMID- 15285104 TI - Tunable dual-wavelength optical short-pulse generation by use of a fiber Bragg grating and a tunable optical filter in a self-seeding scheme. AB - A simple self-seeding scheme is developed to generate tunable dual-wavelength optical short pulses in a flexible manner and with an increased wavelength-tuning range. The wavelength selection and tuning are achieved by simultaneous use of a fiber Bragg grating and a tunable optical filter. The side-mode suppression ratio of the output pulses is better than 30 dB over a wavelength-tuning range of 33.8 nm. The system is compact and convenient for dual-wavelength tuning. PMID- 15285105 TI - Casein gelation under simultaneous action of transglutaminase and glucono-delta lactone. AB - Casein solutions (5% w/v) were treated with microbial transglutaminase (MTG) and glucono-delta-lactone (GDL) under varying conditions in order to obtain gels. Storage modulus (G') and gelation time of the gels were measured by oscillation rheometry, while protein cross-linking was determined by gel permeation chromatography. The addition of only GDL to milk resulted in very weak gels, while MTG on its own was not able to create gel networks. Simultaneous action of both ingredients led to gels, the firmness of which was linearly related to the added amount of MTG, but passed through a maximum with rising GDL concentrations. Using chromatographical analysis, increasing G' values were interrelated with the formation of MTG-induced oligomers. The gelation time was directly proportional to the GDL concentration but not influenced by the addition of MTG within the studied range of concentration. PMID- 15285106 TI - Influence of cooking and microwave heating on microstructure and mechanical properties of transgenic potatoes. AB - The transgenic potato clones of cultivar Irga with improved resistance to a necrotic strain of potato virus Y (PVY(N)) were subjected to heat treatment in order to determine their technological quality. The technological quality was determined on the basis of differences between mechanical properties of unmodified potato and transgenic clones during cooking and microwave heating. The compression test was applied in order to evaluate the mechanical resistance of raw, cooked and microwave-treated potatoes. Compression resistance was expressed by fracture stress F (kPa), fracture strain D (mm/mm), and Young modulus E (kPa). The differences in microstructure of potato tubers (unmodified and modified) were investigated using scanning electron microscopy (SEM). The observed differentiation in the mechanical properties of heat-treated potatoes was less connected with genetic modification but most of all with a kind of the process used. The heat processes caused a distinct decrease in mechanical resistance in all the examined tubers. However, the process of microwave heating resulted in more significant changes in mechanical properties of tubers than cooking. Deformation of parenchyma cells during cooking was directly connected with starch, gelatinisation and gel formation. Microwave heating affected significantly cellular water evaporation which resulted in intercellular failure, collapsing of cells, and limitation of starch gelatinisation. PMID- 15285107 TI - Purification and physicochemical characterization of ovine beta-lactoglobulin and alpha-lactalbumin. AB - Ovine whey proteins were fractionated and studied by using different analytical techniques. Anion-exchange chromatography and reversed-phase high-performance liquid chromatography (HPLC) showed the presence of two fractions of beta lactoglobulin but only one of alpha-lactalbumin. Gel permeation and sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis allowed the calculation of the apparent molecular mass of each component, while HPLC coupled to electrospray ionisation-mass spectrometry (ESI-MS) technique, giving the exact molecular masses, demonstrated the presence of two variants A and B of ovine beta lactoglobulin. Amino acid compositions of the two variants of beta-lactoglobulin differed only in their His and Tyr contents. Circular dichroism spectroscopy profiles showed pH conformation changes of each component. The thermograms of the different whey protein components showed a higher heat resistance of beta lactoglobulin A compared to beta-lactoglobulin B at pH 2, and indicated high instability of ovine alpha-lactalbumin at this pH. PMID- 15285108 TI - Removal of phenolic compounds in pomegranate juices using ultrafiltration and laccase-ultrafiltration combinations. AB - Phenolic compounds of fruit juices are responsible for haze and sediment formation as well as for color, bitterness and astringency. The influence of ultrafiltration (UF) and laccase-UF combination was investigated on phenolic contents of pomegranate juices and on filtration output. Laccase-treated and then ultrafiltered pomegranate juices have shown a rapid increase in their color, when compared to only ultrafiltered (control) samples. Kinetic parameters of laccase were also determined. During the oxidation period, the changes occurring in pomegranate juices were estimated from phenolic contents, color and anthocyanin measurements. Results have shown that laccase oxidation produced a significant decrease in phenolic content of pomegranate juices while juice color the increased. However, in recent literatures, the possibility to remove polyphenols in apple juices was reported. We decided in this study that laccase treatment can not be applied due to the loss of natural red color and unwanted dark brownish color formation in pomegranate juice. PMID- 15285109 TI - Inhibitory effect of pollen and propolis extracts. AB - Bee pollen and propolis were collected from Apis mellifera colonies in five regions of Turkey. The antifungal properties of methanol extracts of pollen and propolis (2% and 5% concentrations) were determined on Alternaria alternata and Fusarium oxysporium f. sp. melonis. The least active concentration towards the tested fungi was 2% concentration of both extracts. The inhibitory effect of all propolis extracts on growth of F. oxysporium and A. alternata were generally higher when compared with pollen extracts. The growth of A. alternata and F. oxysporium were not affected at both concentrations of pollens. However, F. oxysporium against propolis extracts was more sensitive than A. alternata (P < 0.01). None of the pollen extracts tested completely inhibited mycelial growth of fungi used in our experiment. Percent inhibition of both pollen concentrations against A. alternata and F. oxysporium was lower than 50%. However, both concentrations of Alanya and Beysehir propolis extracts were 100% effective on mycelial growth of F. oxysporium until the 7th day of incubation (P < 0.01). 2% Alanya and Beysehir pollen extracts completely stimulated mycelial growth of F. oxysporium on the 7th day of incubation. Both concentrations of propolis extract showed more than 50% inhibition against E. oxysporium. It is suggested that high concentrations ofpropolis extract could be used as an antifungal agent against tested fungi. PMID- 15285110 TI - Effect of gamma-radiation and microwave heating of wheat grain on some starch properties in irradiated grain as well as in grain of the next generation crops. AB - Grains of the Polish winter wheat variety Begra were subjected to gamma-radiation (grain harvested in 1996) within the dose range of 0.05-10 kGy and microwave heating (grain harvested in 1997) from 28 degrees C to 98 degrees C. Later the grains were divided into two parts, the first was used for direct analyses after treatment. The second part was sown on the experimental fields. The obtained crop was described as the first generation and divided into two parts. One part was destined to determination of starch properties and the second part was sown in order to obtain the second generation crop. The same pattern was conducted in order to achieve the third generation crop. gamma-Irradiation directly applied on the wheat grain reduced statistically significant falling number values and gelatinisation enthalpy (deltaH) of the grain treated by 5 and 10 kGy. Calculated linear regression correlation coefficient between the falling number values and the gelatinisation enthalpy was equal to 0.94 (p < or = 0.001) and showed that these two starch characteristics are well correlated in the case of directly irradiated wheat grain. The falling number values, peak temperatures (Tp) and gelatinisation enthalpy (deltaH), in three generations of wheat grain crop studied, did not show any statistically significant differences as a result of indirect effect of gamma-irradiation. Microwave direct heating of wheat grain to 98 degrees C caused a statistically significant increase in the falling number value and decrease in starch gelatinisation power expressed by the enthalpy of gelatinisation (deltaH). The statistically significant changes in the falling number values, slight changes in the peak temperatures (Tp) and enthalpy of gelatinisation (deltaH) were found in all three generation crops as an indirect effect of microwave heating. PMID- 15285111 TI - Electrochemical detection of ascorbic acid (vitamin C) using a glassy carbon electrode. AB - A method for the detection of ascorbic acid (AA) was developed using square wave anodic stripping voltammetric (SWASV) analysis with a glassy carbon electrode (GCE). The results indicated a sensitive oxidation peak current of AA on the GCE. A linear curve was obtained within a concentration range of 1-130.0 microg/L with a preconcentration time of 270 s. The relative standard deviation of 30.0 microg/L observed was 0.1016% (n = 12) under optimum conditions. The low detection limit (S/N) was pegged at 0.30 microg/L. Results showed that the method developed can be used to assay biological and pharmaceutical samples, and food samples, as well as other materials requiring AA analyses. PMID- 15285112 TI - Change in glyceride composition of olive pomace oil during enzymatic esterification. AB - Enzymatic esterification of free fatty acids of olive pomace oil with glycerol was investigated. The esterification reaction was carried out in the absence and presence of glycerol (glycerol to free fatty acids (FFA) molar ratio of 1/3 and 1/6). In the absence of glycerol, the FFA concentration decreased from 32 to 21% while the triglyceride concentration increased from 33 to 40% after of 8 h of reaction time. The most significant decrease in FFAs and increase in triglycerides was observed at the limiting concentration of glycerol (glycerol to FFA molar ratio of 1/3). The FFA concentration decreased to 2.5% and the triglyceride concentration increased up to 78%. The change in both FFA and triglyceride concentrations was found to be statistically significant (P < 0.05). PMID- 15285113 TI - A study on the lipid fraction of Adriatic sardine filets (Sardina pilchardus). AB - Sardine (Sardina pilchardus Walb.) is an important Mediterranean commercial fish species. In this study, the lipids of sardine filets, fished in the Adriatic Sea at different times, were examined. In function of their total lipid (TL) content, sardine filet samples were grouped into lean (TL < 4%) and fat (TL > 4%). It was demonstrated that the differences of TL were exclusively due to a seasonal cyclical increase of neutral lipids. In fact, during moderate-hot months, sardines accumulated reserve fat that was metabolised during the winter months. The fatty acid composition was similar in both sardine sample groups and the fatty acid profile was equally distributed among saturated fatty acids, on average 38.3%, 31.2% monounsaturated, and 30.4% polyunsaturated. The polyunsaturated fatty acid n3 (PUFA-n3) represented on average 20.9%, always higher than PUFA-n6. C20:5n3 eicosapentaenoic acid, (EPA) and C22:6n3 docosahexaenoic acid (DHA) were the must abundant PUFA-n3. Under nutritional aspects, the lipids of 100 g of fat-sardines provided PUFA-n3 quantities, in particular EPA and DHA, significantly higher than human daily requirements. In lean-sardines, the PUFA-n3 input drastically decreased and was estimated that EPA and DHA inputs in 100 g of sardines covered around 17% and 50% of daily requirements, respectively. Finally, cholesterol was equal to 93 mg/100 g of sardines, ranging from 67 to 131 and it did not increase in relation to the total lipid content. In conclusion, this study has highlighted that, under nutritional aspects, regarding EPA and DHA inputs, it is preferable to consume sardines with at least 4% total lipids. PMID- 15285114 TI - Chemical, technological, and nutritional characteristics of two lines of "farro" (Triticum turgidum ssp. dicoccum). AB - In recent years, the renewed interest for foods with a natural image has increased the demand for dry pasta produced from "hulled" wheat such as the Triticum turgidum ssp. dicoccum, also known as "farro". In order to contribute to the general knowledge, two lines of farro were considered in this study. To have a comparison, an old cultivar of Triticum turgidum ssp. durum (Senatore Cappelli) in addition to a commercial semolina were also examined. All semolina samples were used to produce pasta samples. Results showed some differences among pasta samples that seem to be due not to the presence of specific protein subunits but especially to the quantitative ratio between the different subunits. Results also reconfirmed the role played by the drying technology that is able to affect the sensory characteristics of pasta products. PMID- 15285115 TI - Metabolic activities of Lactobacillus spp. strains isolated from kefir. AB - A total of 21 strains of Lactobacillus species were isolated from Turkish kefir samples, in order to select the most suitable strains according to their metabolic activities including probiotic properties. As a result of the identification tests, 21 Lactobacillus isolates were identified as L. acidophilus (4%), L. helveticus (9%), L. brevis (9%), L. bulgaricus (14%), L. plantarum (14%), L. casei (19%) and L. lactis (28%). The amount of produced lactic acid, hydrogen peroxide, proteolytic activity, and acetaldehyde productions of Lactobacillus spp. were determined. Different amounts of lactic acid were produced by strains studies; however, lactic acid levels were 1.7-11.4 mg/mL. All strains produced hydrogen peroxide. L. bulgaricus Z14L strain showed no proteolytic activity, L. casei Z6L strain produced the maximum amount (0.16 mg/mL) of proteolytic activity. Acetaldehyde concentration produced in Lactobacillus strains ranged between 0.88-3.52 microg/mL. PMID- 15285116 TI - Composition and in vitro digestibility of raw versus cooked white- and colour flowered peas. AB - Ten pea cultivars (four white-flowered, Pisum sativum ssp. hortense, and six colour-flowered, Pisum sativum ssp. arvense) grown in Latvia were analyzed and tested in in vitro experiments, as raw and cooked seeds. The colour-flowered (CF) had a greater proportion of hulls and a higher acid detergent fibre (ADF) content than white-flowered (WF) pea seeds (10.7 vs. 8.2% and 92.2 vs. 84.5 g/kg dry matter (DM), respectively). Three out of six CF varieties had a significantly greater amount of protein bound to neutral detergent fibre (NDF) than WF peas. The tannin content was higher in CF than in WF peas (8.46 vs. 0.37 g/kg DM). In vitro protein and amino acid digestibility was about 8% higher in WF than in CF varieties. Cooking decreased the tannin content in CF peas (8.46 vs. 5.51 g/kg DM) but had no effect on in vitro protein digestibility. Heat treatment reduced significantly trypsin inhibitor activity and amount of protein bound to NDF in CF and WF varieties (from 6.50 to 0.52 and from 6.54 to 0.46 trypsin inhibitor units (TIU)/mg DM; from 1.250 to 0.831 and 0.761 to 0.209 g N/100 g NDF, respectively). However, the protein bound to NDF content in pea DM increased in CF and decreased in WF varieties (from 1.525 to 2.145 and from 0.913 to 0.502 g N/kg DM, respectively). Cooking resulted in an increased NDF content over two times in both CF and WF pea seeds (from 122 to 259 and from 120 to 262 g/kg DM, respectively). The results suggest that colour-flowered pea may be considered as an interesting dietary alternative to white-flowered pea since cooking removes trypsin inhibitor activity (TIA), decreases tannins, and increases dietary fibre contents. PMID- 15285117 TI - Preparation and functional properties of extracts from bee bread. AB - Three extracts, namely hot-water fraction (HWF), water-soluble fraction (WSF), and ethanol-soluble fraction (ESF), were prepared from fresh bee bread imported from Lithuania. The protein and total phenolic contents of these samples were very high. Among them, WSF at 100% concentration showed the highest antioxidative ability and scavenging ability. On the other hand, ESF at 10% concentration possessed the highest ability against 1,1-diphenyl-2-picrylhydrazyl (DPPH) and hydroxyl radicals. Bee bread will apply more and more as health food and medicine due to its functional properties such as antioxidative ability and scavenging activities of reactive oxygen species. PMID- 15285118 TI - Antioxidant and angiotension-converting enzyme inhibition capacities of various parts of Benincasa hispida (wax gourd). AB - Vegetables and fruits have been shown to be good sources of antioxidants. Benincasa hispida (wax gourd) has been used in traditional Chinese medicine to treat hypertension and inflammation. The aims of this study were to investigate the abilities of antioxidation and inhibition of angiotensin-converting enzyme (ACE) activity of wax gourd pulp, core, seed, and peel prepared by different extraction methods. The fresh weights required to reach 50% inhibition of linoleic acid oxidation were higher in fresh extracts, compared to other extraction methods. Fresh weights required to reach 50% inhibition were the lowest in seed. The seed had the lowest Cu2+ -induced low-density lipoprotein (LDL) oxidation percentage and inhibition level of ACE activity among all parts. The higher antioxidant capacity of the seed may result from the higher total phenolics contents and superoxide dismutase activity. The abilities of antioxidation and ACE activity inhibition may provide protective effects against cardiovascular diseases and cancers. PMID- 15285119 TI - Effect of gamma-irradiation on aflatoxin B1 production by Aspergillus flavus and chemical composition of three crop seeds. AB - The effect of gamma-irradiation on aflatoxin B1 production by Aspergillus flavus, and the chemical composition of some different crop seeds were investigated. A. flavus infected seeds behaved differently according to their principal constituents. A. flavus caused an increase in protein and decrease in lipids and carbohydrate contents of wheat, soyabean and fababean seeds. Growth of A. flavus and production of aflatoxin B1 was inhibited at a dose level of 5 kGy. A. flavus utilizes carbohydrates of seeds for its growth and aflatoxin production. Crops were arranged, in descending order, according to aflatoxin produced in seeds as wheat > soyabean > fababean. There were no changes in chemical constituents of irradiated seeds, such as protein, lipids, and carbohydrates. PMID- 15285120 TI - Causal judgment from contingency information: a systematic test of the pCI rule. AB - Contingency information is information about the occurrence or nonoccurrence of an effect when a possible cause is present or absent. Under the evidential evaluation model, instances of contingency information are transformed into evidence and causal judgment is based on the proportion of relevant instances evaluated as confirmatory for the candidate cause. In this article, two experiments are reported that were designed to test systematic manipulations of the proportion of confirming instances in relation to other variables: the proportion of instances on which the candidate cause is present, the proportion of instances in which the effect occurs when the cause is present, and the objective contingency. Results showed that both unweighted and weighted versions of the proportion-of-confirmatory-instances rule successfully predicted the main features of the results, with the weighted version proving more successful. Other models, including the power PC theory, failed to predict the results. PMID- 15285121 TI - Counterfactual thinking: the temporal order effect. AB - People often think about how things might have happened differently. Their counterfactual thoughts tend to mentally undo the most recent event in an independent sequence. Consider a game in which two players must each pick the same color card, both red or both black. The first picks black and the second picks red and so they lose. People think, "If only the second player had picked black." Our study tested the idea that the ways in which the players could have won provide counterfactual alternatives to the facts. In three experiments, the same set of facts (both players picked black cards), and the same winning conditions (to win in this new game they must pick different color cards) were presented, but the description of the winning conditions varied (e.g., "if one or the other but not both picks a red card" vs. "if one or the other but not both picks a black card"). The results showed that the temporal order effect can be produced or reversed by different descriptions. The descriptions make accessible different elements of the winning possibilities. A theory of the mental representations and cognitive processes underlying counterfactual thinking in the temporal order effect is described. PMID- 15285122 TI - Measures of similarity in models of categorization. AB - This paper concerns the use of similarities based on geometric distance in models of categorization. Two problematic implications of such similarities are outlined. First, in a comparison between two stimuli, geometric distance implies that matching features are not taken into account. Second, missing features are assumed not to exist. Only nonmatching features enter into calculations of similarity. A new model is constructed that is based on the ALCOVE model (Kruschke, 1992), but it uses a feature-matching similarity measure (see, e.g., Tversky, 1977) rather than a geometric one. It is an on-line model in the sense that both dimensions and exemplars are constructed during the categorization process. The model accounts better than ALCOVE does for data with missing features (Experiments 1 and 2) and at least as well as ALCOVE for a data set without missing features (Nosofsky, Kruschke, & McKinley, 1992). This suggests that, at least for some stimulus materials, similarity in categorization is more akin to a feature-matching procedure than to geometric distance calculation. PMID- 15285123 TI - False prototype enhancement effects in dot pattern categorization. AB - Results from the classic dot pattern distortion paradigm have sometimes yielded prototype enhancement effects that could not be accounted for by exemplar models of categorization. However, in these experiments the status of the prototype was confounded with certain stimulus-specific properties as well as with the frequency of presentation of the prototype during testing. In two mock-subliminal experiments, participants made categorization judgments to patterns that were generated as prototypes, low-level distortions, or high-level distortions. The participants rated the prototypes as being more likely to be members of a category, although no patterns were presented during training, and there was no objective category structure. In two other experiments, greater prototype enhancement effects were observed when the prototype and low-level distortions were presented with greater frequency during transfer. These results suggest that classic prototype enhancement effects may not be due to the abstraction of a prototype at time of original learning, but rather to other factors not formalized in extant models. PMID- 15285124 TI - Spatial updating of virtual displays during self- and display rotation. AB - In four experiments, we examined observers' ability to locate objects in virtual displays while rotating to new perspectives. In Experiment 1, participants updated the locations of previously seen landmarks in a display while rotating themselves to new views (viewer task) or while rotating the display itself (display task). Updating was faster and more accurate in the viewer task than in the display task. In Experiment 2, we compared updating performance during active and passive self-rotation. Participants rotated themselves in a swivel chair (active task) or were rotated in the chair by the experimenter (passive task). A minimal advantage was found for the active task. In the final experiments, we tested similar manipulations with an asymmetrical display. In Experiment 3, updating during the viewer task was again superior to updating during the display task. In Experiment 4, we found no difference in updating between active and passive self-movement. These results are discussed in terms of differences in sources of extraretinal information available in each movement condition. PMID- 15285125 TI - Spatial memory and perspective taking. AB - Giving directions or describing an environment often requires assuming perspectives other than one's own. We employed a spatial perspective-taking task to investigate how describing familiar versus novel perspectives affects subsequent memory. One participant (the director) viewed a display of objects from a single perspective and described the display to another participant (the matcher) from a perspective that varied by 0 degrees, 45 degrees, 90 degrees, 135 degrees, or 180 degrees from the viewing perspective. Following the description, we assessed the director's memory for the display, using judgments of relative direction, scene recognition, and map drawing. The participants imagined and recognized familiar views faster and/or more accurately than novel views. Moreover, different tasks showed different degrees of facilitation for the visually perceived and described views, suggesting multiple representations for different aspects of spatial memory. These findings emphasize the importance of understanding distinctions among spatial experiences and underscore differences in the tasks used to probe spatial memory. PMID- 15285126 TI - Cultural life scripts structure recall from autobiographical memory. AB - Three classes of evidence demonstrate the existence of life scripts, or culturally shared representations of the timing of major transitional life events. First, a reanalysis of earlier studies on age norms shows an increase in the number of transitional events between the ages of 15 and 30 years, and these events are associated with narrower age ranges and more positive emotion than events outside this period. Second, 1,485 Danes estimated how old hypothetical centenarians were when they had been happiest, saddest, most afraid, most in love, and had their most important and most traumatic experiences. Only the number of positive events showed an increase between the ages of 15 and 30 years. Third, undergraduates generated seven important events that were likely to occur in the life of a newborn. Pleasantness and whether events were expected to occur between the ages of 15 and 30 years predicted how frequently events were recorded. Life scripts provide an alternative explanation of the reminiscence bump. Emphasis is on culture, not individuals. PMID- 15285127 TI - Autobiographical memories for the September 11th attacks: reconstructive errors and emotional impairment of memory. AB - College students were asked about their personal memories from September 11, 2001. Consistency in reported features over a 2-month period increased as the delay between the initial test and 9/11 increased. Central features (e.g., Where were you?) were reported with greater consistency than were peripheral features (What were you wearing?) but also contained a larger proportion of reconstructive errors. In addition, highly emotional participants demonstrated poor prospective memory and relatively inconsistent memory for peripheral details, when compared with less emotional participants. Highly emotional participants were also more likely to increase the specificity of their responses over time but did not exhibit greater consistency for central details than did less emotional participants. The results demonstrated reconstructive processes in the memory for a highly consequential and emotional event and emotional impairment of memory processing of incidental details. PMID- 15285128 TI - Conditions affecting the revelation effect for autobiographical memory. AB - In four experiments involving 184 participants, people rated their confidence that particular events had happened in their childhood (e.g., "Broke a window playing ball"). If participants had to unscramble a key word in a phrase just before rating it (e.g., "Broke a nwidwo [window] playing ball"), confidence ratings increased-the revelation effect. However, the pattern of revelation effects depended on the particular way in which participants processed key words (e.g., visualizing vs. counting vowels in the word window) approximately 10 min prior to rating life events that contained those words. Prior exposure to key words never in itself directly affected confidence ratings. These results demonstrate that one can manipulate the revelation effect by altering the processing that participants perform on words prior to unscrambling them. These results also pose difficulties for many accounts of the revelation effect. The major puzzle posed by our present findings is that unscrambling key words increases confidence that an event has happened in childhood, whereas prior exposure to these words does not. PMID- 15285129 TI - False memories for compound words: role of working memory. AB - Subjects studied pairs of compound words; pair members were presented simultaneously (one above the other) for 2 sec or sequentially (one immediately following the other) for 1 sec each, and 6-sec interstimulus intervals separated the end of presentation of one pair and the start of that of another. A subsequent recognition test included within-pair and between-pair conjunction foils (recombinations of stimulus parts from the same study pair and from separate pairs, respectively). Previous experiments using faces as stimuli have demonstrated that when faces are presented simultaneously there are many more false alarms to within-pair than to between-pair conjunction items, and when faces are presented sequentially there is an equal number of false alarms in those two conditions. However, Experiment 1 showed that for compound word stimuli there were equally high false alarm rates toboth types of foils in both study conditions relative to completely new test items. Experiment 2 showed that when rehearsal of compound words was prevented, the pattern of conjunction errors was very similar to the one typically obtained for faces. In Experiment 3, subjects falsely recalled conjunctions of within-pair compound words but not conjunctions of between-pair words in the simultaneous-study condition; no conjunctions were recalled in the sequential-study condition. The results support the idea that working memory processing is necessary for binding stimulus parts together in episodic memory. PMID- 15285130 TI - Relations between emotion, memory, and attention: evidence from taboo stroop, lexical decision, and immediate memory tasks. AB - This article reports five experiments demonstrating theoretically coherent effects of emotion on memory and attention. Experiments 1-3 demonstrated three taboo Stroop effects that occur when people name the color of taboo words. One effect is longer color-naming times for taboo than for neutral words, an effect that diminishes with word repetition. The second effect is superior recall of taboo words in surprise memory tests following color naming. The third effect is better recognition memory for colors consistently associated with taboo words rather than with neutral words. None of these effects was due to retrieval factors, attentional disengagement processes, response inhibition, or strategic attention shifts. Experiments 4 and 5 demonstrated that taboo words impair immediate recall of the preceding and succeeding words in rapidly presented lists but do not impair lexical decision times. We argue that taboo words trigger specific emotional reactions that facilitate the binding of taboo word meaning to salient contextual aspects, such as occurrence in a task and font color in taboo Stroop tasks. PMID- 15285131 TI - On the control of visual word recognition: changing routes versus changing deadlines. AB - Two different accounts have been proposed to explain the fact that (1) an effect of word frequency is present when readers of transparent orthographies read only words aloud and (2) the effect of word frequency is eliminated when subjects name words and nonwords mixed together in a single block. In the route-shifting account, subjects shift from using a lexical route that can read only words to using a nonlexical route that can read both words and nonwords via the use of sublexical spelling-sound correspondences (hence, no word frequency effect). The essence of the second, time criterion account is that the elimination of the word frequency effect is determined by the speed with which the nonwords are processed, because subjects attempt to homogenize the point in time at which they release an articulation. These two different accounts are pitted against each other in a series of naming experiments utilizing the transparent Turkish orthography. A word frequency effect persists even when words and nonwords are mixed together, provided that nonword sets are matched so as to be named as quickly as the high-frequency words and as slowly as the low-frequency words, respectively. This result is argued to be consistent with the time criterion account, but not with the unadorned route-shifting account. PMID- 15285132 TI - Redintegration and lexicality effects in children: do they depend upon the demands of the memory task? AB - The effect of long-term knowledge upon performance in short-term memory tasks was examined for children from 5 to 10 years of age. The emergence of a lexicality effect, in which familiar words were recalled more accurately than unfamiliar words, was found to depend upon the nature of the memory task. Lexicality effects were interpreted as reflecting the use of redintegration, or reconstruction processes, in short-term memory. Redintegration increased with age for tasks requiring spoken item recall and decreased with age when position information but not naming was required. In a second experiment, redintegration was found in a recognition task when some of the foils rhymed with the target. Older children were able to profit from a rhyming foil, whereas younger children were confused by it, suggesting that the older children make use of sublexical phonological information in reconstructing the target. It was proposed that redintegrative processes in their mature form support the reconstruction of detailed phonological knowledge of words. PMID- 15285133 TI - Readers' sensitivity to linguistic cues in narratives: how salience influences anaphor resolution. AB - Despite the general assumption that anaphoric inferences are necessary inferences, Levine, Guzman, and Klin (2000) concluded that the probability of resolving noun phrase anaphors depends both on the degree of accessibility in memory of the antecedent concepts and the extent to which resolution is necessary to create a coherent discourse representation. Four experiments are presented in which the factors that influence readers' standard of coherence are investigated. We examine the hypothesis that readers are more likely to resolve anaphors that are perceived as salient; salience was manipulated both with a syntactic focusing structure (wh- clefts) and with the addition of prenominal adjectival modifiers. The results of a probe recognition time task provide support for the hypothesis that a variety of linguistic cues serve as mental processing instructions (Givon, 1992), which instruct readers as to how much attention to devote to processing. PMID- 15285134 TI - Plant host associations of Penthaleus species and Halotydeus destructor (Acari: Penthaleidae) and implications for integrated pest management. AB - Integrated pest management programs seek to minimise reliance on pesticides and provide effective long-term control of pests. Cultural control strategies, such as crop rotations, trap and border crops, and weed management, require a thorough understanding of pest host associations. This paper examines the effects of different plant hosts on the persistence and reproduction of blue oat mites, Penthaleus spp., and the redlegged earth mite, Halotydeus destructor (Tucker), which are major agricultural pests in southern Australia. Field and shade-house experiments were conducted testing several crop and plant types. All species survived and reproduced from one mite season to the next when confined to pasture. Canola and a common weed, 'bristly ox-tongue', were suitable hosts for H. destructor and Penthaleus falcatus (Qin and Halliday), whereas Penthaleus sp. x and Penthaleus major (Duges) failed to persist on these plants. A mixture of wheat and oats sustained P. sp. x and H. destructor, but not P. falcatus or P. major. Lentils were generally a poor host plant for all mite species. These findings show that earth mite species differ in their ability to persist on different plant types, highlighting the importance of distinguishing mite species before implementing control strategies. Results are discussed with respect to cultural control options for the management of these winter pests. PMID- 15285135 TI - Below-ground plant parts emit herbivore-induced volatiles: olfactory responses of a predatory mite to tulip bulbs infested by rust mites. AB - Although odour-mediated interactions among plants, spider mites and predatory mites have been extensively studied above-ground, belowground studies are in their infancy. In this paper, we investigate whether feeding by rust mites (Aceria tulipae) cause tulip bulbs to produce odours that attract predatory mites (Neoseiulus cucumeris). Since our aim was to demonstrate such odours and not their relevance under soil conditions, the experiments were carried out using a classic Y-tube olfactometer in which the predators moved on a Y-shaped wire in open air. We found that food-deprived female predators can discriminate between odours from infested bulbs and odours from uninfested bulbs or artificially wounded bulbs. No significant difference in attractiveness to predators was found between clean bulbs and bulbs either wounded 30 min or 3 h before the experiment. These results indicate that it may not be simply the wounding of the bulbs, but rather the feeding by rust mites, which causes the bulb to release odours that attract N. cucumeris. Since bulbs are belowground plant structures, the olfactometer results demonstrate the potential for odour-mediated interactions in the soil. However, their importance in the actual soil medium remains to be demonstrated. PMID- 15285136 TI - The relationship between dietary specialism and availability of food and water on cannibalistic interactions among predatory mites in protected crops. AB - The interplay between dietary specialism, the tolerance of food and water stress and level of cannibalism is likely to be important in determining the outcome of biological control using inundative releases of multiple natural enemies, such as phytoseiid mites in protected crops. The dietary specialist, Phytoseiulus persimilis, with a short immature development time (4-5 days) when plentiful food was available had a low ability to survive without food (5 days), even with access to water. The dietary generalists, Neoseiulus californicus, N. cucumeris and lphiseius degenerans, had longer immature development times (by up to 2 days) than P. persimilis. Survival ability differed amongst the generalist species when they were starved but provided with constant access to water. Both N. californicus and N. cucumeris survived the longest (8-10 days) and I. degenerans survived the shortest period (4 days). No negative intra-specific interaction between immatures was observed with P. persimilis when food was available and in the absence of food this species tended to starve rather than act cannibalistically. Both N. californicus and N. cucumeris showed a low degree of cannibalism between immatures, either when food was available, or when starved but given access to water. Even when food was available survival of I. degenerans fell by 30% in 4 days and remained at 60-70% for 3 further days; survival continued to decline rapidly when they were starved but provided with water. This indicates that immatures of I. degenerans could either feed on dead conspecifics or that they were capable of a degree of cannibalism. Adult females of P. persimilis did not feed on conspecific eggs even when deprived of food but provided with water. Adult female N. californicus and N. cucumeris did feed on conspecific eggs but at a low level (<1 egg per day), which occurred only after 48 h starvation. Although egg cannibalism occurred more consistently with adult female I. degenerans than with other mite species it was at a low level (<1 egg per day). If the tendency to cannibalism, not just of eggs but with more susceptible life stages such as larvae, is reduced when water is available freely this could be important in determining the interactions that occur under natural conditions. PMID- 15285137 TI - The predatory mite Typhlodromus pyri (Acari: Phytoseiidae) causes feeding scars on leaves and fruits of apple. AB - Typhlodromus pyri Scheuten (Acari: Phytoseiidae) is the most important predator of Panonychus ulmi (Koch) (Acari: Tetranychidae) in orchards and vineyards. It was recently found that adult T. pyri females cause microscopic scars on apple leaves. The present laboratory experiments were carried out to confirm the production of scars on apple leaves and to assess if females cause scars on fruits as well. Scar production on apple leaves and/or fruits was investigated under various nutritional conditions: no food, pollen of Scots pine (Pinus sylvsestris L.) only, nymphs of P. ulmi only, and pollen + prey. Both on leaves and fruits, either offered alone or in combination, feeding scars were produced under all nutritional conditions, but mostly in the 'no food' treatment. The predators consumed significantly more P. ulmi nymphs when offered alone than when offered in combination with pollen. T. pyri laid eggs under all nutritional conditions, but mostly in the 'pollen + prey' treatment and least when no food was offered. T. pyri females caused scars on both leaves and fruits when offered simultaneously, but more on leaves than on fruits. The scars were also bigger on leaves than on fruits in all experiments. T. pyri survived and reproduced on plant material in the absence of other food sources. Whether the scars produced on leaves and fruits harm the quality of fruits or the yield of apple cannot be concluded from the present experiments. PMID- 15285138 TI - Effects of NaCl-stressed citrus plants on life-history parameters of Tetranychus urticae (Acari: Tetranychidae). AB - Tetranychus urticae is an important pest of citrus, especially lemon and mandarin, under Mediterranean climate. Factors leading to this problem are poorly understood, but saline stress is suspected to contribute to spider mite outbreaks. In this study, the effect of NaCl concentration in nutritive solutions used to water potted young mandarin trees on population growth of T. urticae reared on leaf discs obtained from these plants was investigated. Although the differences observed between treated (5, 10, 30 and 60 mM NaCl) and control groups were in most cases not significant, when all biological parameters calculated were combined to obtain Ro, T and r(m), remarkable differences appeared, and a concentration-dependent effect was detected. Although high salt concentrations negatively affected T. urticae, at the lowest concentration tested the r(m) value was significantly higher than at the water control and this may contribute to the observed field explosions of T. urticae. PMID- 15285139 TI - Tactic responses of the parasitic mite, Psoroptes ovis, to light and temperature. AB - The astigmatid mite, Psoroptes ovis (Hering) (Acari: Psoroptidae), is an obligate, non-burrowing ectoparasite of vertebrates, of particular economic importance in domestic sheep flocks where it causes clinical psoroptic mange. To help understand the behaviour which facilitates transmission via the environment, the responses of P. ovis derived from rabbits (syn. Psoroptes cuniculi) to temperature and light were examined in the laboratory. On a vertical surface of uniform temperature, the presence and direction of illumination had a significant effect on the distance and direction moved by the mites. In darkness or with illumination from both above and below, the mites moved relatively little, but this movement was upwards. In contrast, with illumination from above only, mites moved downwards. When the direction of the illumination was reversed so that it came from below only, the mites moved upwards. On a vertical surface with a temperature gradient, in darkness or with illumination from both above and below, the mites moved up or down towards the area of highest temperature, depending on whether this was above or below, respectively. However, the movement of the mites in response to the temperature gradient was strongly displaced up or down by the presence of unidirectional illumination from above or below, respectively. The results indicate that the movement of these mites is strongly directed towards areas of high temperature but away from higher light intensity. These behaviours might be expected to maintain the position of the mites on a host animal and help them locate the skin surface of a new host when displaced into the environment. PMID- 15285140 TI - Rapid quantitative assay for acaricidal effects on Sarcoptes scabiei var. suis and Otodectes cynotis. AB - Brimer et al. (Vet. Parasitol. 51: 123-135, 1993 and 59: 249-255, 1995) developed a migration assay for acaricidal effect of acetylcholinesterase inhibitors and macrocyclic lactones utilising Sarcoptes scabiei var. suis mites. In contrast to many others, this assay is fully quantitative but quite time-consuming. The aim of the present investigation was to modify this assay to become faster and simpler. As a result accurate determinations can now be obtained within 6h, as opposed to 24h. Furthermore it was demonstrated that also Otodectes cynotis mites can be used with only minor modifications of the procedures. The cholinesterase inhibitor diazinon and the formamide amitraz were used as acaricides. Thus, the mite migration assay now has been proven useful for acaricidal compounds belonging to three chemical groups with different modes of action, namely organophosphorous cholinesterase inhibitors, macrocyclic lactones acting on the glutamanergic/GABAegic motoneurons, and formamide inhibitors of the octopamine systems of arthropods. PMID- 15285142 TI - Phenology of Bryobia cristata (Acari, Prostigmata) in hayfields in northern Iceland. PMID- 15285141 TI - Neotrombicula autumnalis (Acari, Trombiculidae) as a vector for Borrelia burgdorferi sensu lato? AB - Larvae of the trombiculid mite Neotrombicula autumnalis were collected at 18 sites in and around Bonn, Germany, to be screened for infection with Borrelia burgdorferi s.l. by means of PCR. Questing larvae numbering 1380 were derived from the vegetation and 634 feeding ones were removed from 100 trapped micromammals including voles, mice, shrews and hedgehogs. In a laboratory infection experiment, a further 305 host-seeking larvae from the field were transferred onto Borrelia-positive mice and gerbils, and examined for spirochete infection at various intervals after repletion. In three cases borrelial DNA could be amplified from the mites: (1) from a larva feeding on a wild-caught greater white-toothed shrew (Crocidura russula), (2) from a pool of four larvae feeding on a B. garinii-positive laboratory mouse, and (3) from a nymph that had fed on a B. afzelii-positive laboratory gerbil as a larva. In the first case, borrelial species determination by DNA hybridization of the PCR product was only possible with a B. burgdorferi complex-specific probe but not with a species specific one. In the second case, probing showed the same borrelial genospecies (B. garinii) as the laboratory host had been infected with. In the latter case, however, DNA hybridization demonstrated B. valaisiana while the laboratory host had been infected with B. afzelii. Subsequent DNA sequencing confirmed much higher similarity of the PCR product to B. valaisiana than to B. afzelii indicating an infection of the mite prior to feeding on the laboratory host. The negligible percentage of positive mites found in this study suggests that either the uptake of borrelial cells by feeding trombiculids is an extremely rare event or that ingested spirochetes are rapidly digested. On the other hand, the results imply a possible transstadial and transovarial transmission of borreliae once they are established in their trombiculid host. However, unless the transmission of borreliae to a given host is demonstrated, a final statement on the vector competence of trombiculid mites is not possible. PMID- 15285143 TI - Amblyomma cajennense ticks induce immediate hypersensitivity in horses and donkeys. AB - Since host immune reaction to ticks interferes with tick-borne pathogen transmission, it is important to recognize naturally occurring tick-host immune relationships to better understand the epidemiology of such infectious diseases. Amblyomma cajennense is an important tick-borne disease vector in the Neotropical region and horses maintain it in domestic environments. In the present work intradermal testing of A. cajennense tick exposed horses and donkeys using crude tick antigens was used to evaluate the type of hypersensitivity induced by infestations. Animals sensitized by A. cajennense infestation displayed an immediate hypersensitivity reaction at the antigen inoculation site. Foals sensitized with experimental infestations and field sensitized horses presented the most intense reactions (40% of ear thickness increase). Field sensitized donkeys presented less intense reaction reaching no more than 22% of mean thickness increase. Control horses (non-sensitized) had the least intense reaction, with a peak of no more than 12% of increase. The presence of a prominent immediate hypersensitivity in equids sensitized experimentally or by field infestations indicates that A. cajennense ticks induce in this host an immune response that is associated with IgE production and which is known to be inappropriate against intracellular pathogens. Differences observed between horses and donkeys are discussed. PMID- 15285144 TI - A reassessment of argasid tick salivary gland ultrastructure from an immuno cytochemical perspective. AB - Previous morphological and histochemical studies of argasid tick salivary glands indicated that they were less complex than ixodid salivary glands, with only three granular cell types. The present study shows that there exist at least four different granular cell types in the salivary glands of the argasid tick Ornithodoros savignyi, based on immuno-localization of the anti-hemostatic factors, apyrase and savignygrin. Both anti-hemostatic factors were localized to dense core granule type 'a' and to granule type 'b', that shares a similar homogenous morphology with non-labeled granule type 'd'. Furthermore, the major tick salivary gland proteins (TSGPs), previously implicated in granule biogenesis, were localized to all the granular cell types. This indicates that granular cell types with different morphologies can express the same proteins, while cell types that show similar morphologies may not express the same proteins. Argasid tick salivary glands seem to be more complex than previously thought and might not be amenable to morphological classification alone. Alternative classification methodologies that rely on physical expression patterns of the salivary gland proteome might be more reliable as markers for a specific granular cell type. PMID- 15285145 TI - Regeneration of Haller's sensory organ in two species of hard ticks of the genus Haemaphysalis (Acari: Ixodidae). AB - A study of Haller's organ regeneration in nymphs and adults of Haemaphysalis turturis and parthenogenetic females of H. longicornis, from which the forelegs had been amputated during the previous instar, revealed structural changes in regenerated organs. The adult regenerates reestablished atavistic structural features, while the nymphal regenerates retained larval features, which is typical of regenerates of two other genera examined previously (Ixodes and Hyalomma). Data on regeneration of Haller's sensory organ testify to an ancient character of the genus, standing closely to the base of the phylogenetic tree of hard ticks. PMID- 15285146 TI - Cross-sectional survey of ticks (Acari: Ixodidae) in sheep from an area of the southern Italian Apennines. AB - A cross-sectional survey of ticks was conducted on 197 ovine farms with animals pasturing in an area (3971 km2) of the southern Italian Apennines. The farms were selected to be uniformly distributed throughout the study area using Geographical Information System (GIS). Ticks were collected from 309 (31.4%) out of the 985 sheep sampled, belonging to 92 (46.7%) out of the 197 farms included in the study. The following tick species were found (farm prevalence): Dermacentor marginatus (37.6%), Haemaphysalis punctata (29.4%), H. sulcata (2.5%), H. parva (2.0%). H. inermis (0.5%), Ixodes gibbosus (2.0%), I. ricinus (0.5%), Rhipicephalus sanguineus group (1.0%), and R. bursa (0.5%). A point distribution map (PDM) was drawn by GIS in order to display the distribution of each tick genus in the study area. The general trends of the PDM show that Dermacentor marginatus and Haemaphysalis spp. were widely and homogeneously spread throughout the study area, whereas Rhipicephalus spp. and Ixodes spp. were present only in a few concentrated zones of the study area in accordance to their biological and ecological characteristics. PMID- 15285147 TI - First record of immature stages of Amblyomma tigrinum (Acari: Ixodidae) on wild birds in Chile. AB - For the first time, larvae and nymphs of Amblyomma tigrinum ticks were found on three species of wild birds (Zenaida auriculata, Callipepla californica and Nothoprocta perdicaria) in Chile. A significant higher number of A. tigrinum was found on fledglings of Z. auriculata and N. perdicaria than on adults of these species of birds. A significant higher number was also observed on N. perdicaria living in non-irrigated areas as compared with irrigated areas. Infestation levels were 6.5, 6.3 and 10.2 ticks for Z. auriculata, C. californica and N. perdicaria, respectively. Our results suggest that birds are important in the maintenance of the life cycle of A. tigrinum ticks in the area. PMID- 15285148 TI - [Embryonic stem cell therapy in experimental stroke: host-dependent malignant transformation]. AB - INTRODUCTION: Therapeutic application of embryonic stem cells in neurodegenerative disorders like stroke is widely investigated in preclinical animal models. AIM: The authors studied the therapeutic potential of murine embryonic stem cells in two rodent models of stroke. METHODS: Undifferentiated and predifferentiated stem cells were implanted into the non-ischemic hemisphere of mice and rats following focal brain ischemia. The brains were analysed by immunohistochemistry and histology. The in vitro differentiation of the cells was checked by immunocytochemistry and Western-blot. RESULTS: After xenotransplantation in rats undifferentiated cells migrated along the corpus callosum towards the ischemic injury. Later stem cells differentiated into neurons in the border zone of the lesion. In the homologous mouse brain, the same murine embryonic stem cells did not migrate, but produced highly malignant teratocarcinomas at the site of implantation, independent of whether they were predifferentiated in vitro to neural progenitor cells. These experiments demonstrated a hitherto unrecognized adverse outcome after xenotransplantation and homologous transplantation of embryonic stem cells. CONCLUSION: This observation raises serious concerns about safety provisions when the therapeutic potential of human embryonic stem cells is tested in preclinical animal models. The clinical trials are based on the positive outcome of the xenologous experiments. PMID- 15285149 TI - [The role of essential metal ions in the human organism and their oral supplementation to the human body in deficiency states]. AB - The role of essential nutrient metal ions (Mg, Fe, Cu, Zn, Mn and Co) often deficient in our foodstuffs, although vitally essential in the function of the human organism as well as the different reasons for these deficiencies both in foods and in the human body have been studied. The most frequent nutritional disease is iron deficient anaemia. Inorganic salts, artificial synthetic monomer organic metal complexes of high stability or organic polymer complexes of high molecular mass are unsatisfactory for supplementation to the human body, owing to poor absorption, low availability and/or harmful side effects. In contrast, we have recently found that mixed metal complexes of oligo/polygalacturonic acids with medium molecular weight prepared from natural pectin of plant origin are efficient for oral supplementation. Sufficient absorption of essential metal ions from metal oligo/polygalacturonate mixed complexes with polynuclear innersphere structure is due to the high ionselectivity and medium stability values. Metal oligo/polygalacturonate mixed complexes contain all deficient essential metal ions in adequate amounts and ratios for higher bioavailability of metal ions and optimal vital function. Therefore, by oral administration of these complexes, metal ion homeostasis and optimal interactions with vitamins and hormones can be ensured. Prelatent or latent macroelement Mg deficiency can often be observed among clinical or ambulance patients. Latent or manifest mesoelement iron deficiency is the most common, however, the occurrence of microelement copper, zinc, manganese and cobalt latent deficiencies is not seldom either. Supplementation studies utilizing essential metal oligo/polygalacturonate complexes led to satisfactory outcome without harmful side effects. PMID- 15285151 TI - [Application of robots in the upper limb physiotherapy of patients with hemiparesis]. AB - OBJECTIVE: Authors describe the robotic physiotherapy system developed in the scope of Reharob project belonging to the 5th Framework Programme of the European Union. The system is able to execute robot mediated physiotherapy of the shoulder and the elbow of patients with spastic hemiparesis. Due to the fact, that spasticity is velocity dependent, it is important to execute the exercises with a relatively slow and constant velocity. In such cases a robot can support the work of the physiotherapist first of all when delivering exercises with high repetition number. The objective of the first clinical trial was to gain experiences with the system, to prove its functionality and security. METHOD: 4 healthy subjects and 8 patients with spastic hemiparesis were given 30 minutes long robot mediated physiotherapy on 20 consecutive days. RESULTS: During 240 therapeutic events the robots executed the exercises according to the programme established by the physiotherapist, without any significant mechanical problem or dangerous situation. CONCLUSIONS: The clinical trial confirmed the functionality of the Reharob Therapeutic System. The development of the system will be continued according to the experiences gained during the trial. PMID- 15285150 TI - [Mitochondrial DNA deletions in newborn brain samples]. AB - INTRODUCTION: Mitochondrial DNA deletion affecting 4977 base pairs (mtDNA4977) thought to be the most common somatic mutation in man was analysed in samples taken from various parts of the brains at autopsy in order to analyse the supposition whether this mitochondrial damage may play a role in the causation of neurological dysfunction in childhood. METHODS: DNA was isolated from the samples of 15 newborns and 8 adults taken during autopsy. mtDNA4977 deletion was determined by polymerase chain reaction. RESULTS: mtDNA4977 could be demonstrated not only in adults but also in every newborn sample. Estimation of the amount of mtDNA4977 indicated that the level of mtDNA4977 was smaller in the newborn samples than in the elderly's. CONCLUSION: Results suggest that mtDNA4977, contrary to the generally accepted opinion stating that it is acquired during life span, may already be present in the beginning of life. However, the possibility can not be excluded that mutations in the extreme sensitive mtDNA against oxidative damage might be generated by perinatal hypoxia and intensive care. Such a causative role of mtDNA mutations may be an important additional factor in explaining the pathomechanism of cerebral palsy and mental retardation frequently observed in surviving children. PMID- 15285152 TI - [Liver resection for a rare parasitic infection--visceral larva migrans syndrome]. AB - The authors describe a case of an 38-year-old woman suffering from a parasitical infection which is rare in Hungary. It was diagnosed in connection with a surgical liver segment resection. Visceral larval migrans is an infection caused by migration of the roundworm Toxocara larvae to organs and tissues. The authors describe the pathophysiology of the disease, clinical symptoms, diagnostic and therapeutic possibilities. PMID- 15285153 TI - [Up-to-date management of glaucoma]. PMID- 15285154 TI - [Terminology for vertebral deformities]. PMID- 15285155 TI - [The importance of infant nutrition for the development of type 1 diabetes]. PMID- 15285156 TI - [Lung cancer--should the PET be used?]. PMID- 15285157 TI - [Lung cancer--should positron-emission tomography be used?]. PMID- 15285158 TI - [Gluten and type 1 diabetes mellitus]. PMID- 15285159 TI - [The clinical examination of hip joints in adults]. PMID- 15285160 TI - [Surgical treatment of extreme obesity. Presentation of a Cochrane analysis]. PMID- 15285161 TI - [Dying patients at home and in hospital]. PMID- 15285162 TI - [Adalimumab in the treatment of rheumatoid arthritis]. PMID- 15285163 TI - [Cost-effectiveness of regionalization of esophageal resections in Denmark]. PMID- 15285164 TI - [Esophageal resection in an accelerated postoperative recovery regimen]. PMID- 15285165 TI - [Densities of the tick (Ixodes ricinus) and coexistence of the Louping ill virus and tick borne encephalitis on the island of Bornholm]. PMID- 15285166 TI - [Iodinated contrast media and nephropathy. Estimated creatinine clearance for better assessment of renal function and dosage]. PMID- 15285167 TI - [Decline in AIDS and death rates in the EuroSIDA study. An observational study]. PMID- 15285168 TI - [Allergic bronchopulmonary aspergillosis with fatal outcome]. PMID- 15285169 TI - [Neonatal meningitis and sepsis with hemolytic group A streptococci]. PMID- 15285170 TI - ["An extraordinary initiative"]. PMID- 15285171 TI - [The "money back" concept is a new dimension in a rational pharmacotherapy]. PMID- 15285172 TI - [The sugar report and lipogenesis 2]. PMID- 15285173 TI - [Death attest four years late]. PMID- 15285174 TI - Th1 and Th2 subsets in orthopaedic surgery by single-cell cytokine analysis. AB - Significant immunosuppression can occur following allogeneic blood transfusion or surgery. Cytokine stimulation controls immune responses and determines their type and intensity. Infusion of autologous or allogeneic blood provides elements, including cytokines, which may result in transfusion-associated immunomodulation. This study investigates to what extent autologous/cell salvage transfusions affect levels of intracellular cytokines interferon-gamma and interleukin-4, and if this indicates a shift in the T-helper 1/T-helper 2 cell ratio using a novel method of detecting intracellular cytokines, the Magnetic Activated Cell Sorter Cytokine Secretion Assay (MACS Assay). Comparisons were made between patients receiving autologous blood or no blood transfusion, for pre- and post-operation levels of interferon-gamma and interleukin-4. Interferon-gamma producing T-helper 1 cells decreased post-operatively. Concomitantly, interleukin-4 producing T helper 2 cells increase. These results demonstrate a measurable shift from T helper 1 to T-helper 2 cells post-operatively. Secondly, the study showed surgery alone instigates the same level of immunomodulation as autologous/cell salvage blood transfusion in combination with surgery. PMID- 15285175 TI - Markers of bone resorption--measurement in serum, plasma or urine? AB - OBJECTIVE: We compared an established urine marker (total deoxypyridinoline (DPD), related to creatinine (cr)) with a plasma marker (CTX) to evaluate whether the methods are equally suited to detect increased bone resorption in women after the menopause and to see whether both markers show normal bone resorption in postmenopausal women on hormone replacement therapy (HRT). METHODS: DPD in first morning void urines was measured by an automated HPLC system. CTX ("beta CrossLaps") was measured on the automated electrochemiluminescence analyzer E170 (Roche Diagnostics, Germany). For CTX, EDTA plasma and serum samples taken from fasting patients in the morning between 08.00 and 08.30 were analyzed. 49 women were premenopausal, 43 women were postmenopausal without HRT, and 13 women were postmenopausal on HRT (mostly on oral medication) for more than six months. RESULTS AND DISCUSSION: Inter-assay coefficients of variation (CVs) at three different concentrations were 6.0%, 6.7% and 7.2% for the assay of DPD and 2.58%, 1.83% and 1.99% for CTX, respectively. CTX was more stable in EDTA plasma than in serum. 21/43 (49%) of postmenopausal women without HRT showed increased DPD/cr ratios and 19/43 (44%) showed elevated CTX concentrations. Regarding postmenopausal women on HRT, DPD/cr ratios were elevated in 3/13 women, whereas plasma CTX showed levels within the premenopausal range in all 13 women. It is discussed that in some cases a lower muscle mass as a result of increasing age or as a result of oral HRT might increase the urinary DPD/cr ratio by lowered excretion of cr. This effect would raise the number of cases with an elevated DPD/cr ratio after the menopause out of proportion. CONCLUSION: CTX is determined with very high precision on the E170. CTX, if measured in EDTA plasma samples from fasting patients in the morning, seems to indicate bone resorption in women on HRT correctly as normal. The DPD/cr ratio in urine of women on HRT is increased in some cases above normal, presumably by lowered excretion of cr. According to our results, plasma (or serum) markers of bone resorption seem to be preferable over cr-related urine markers. PMID- 15285176 TI - Clinical value of a new fecal elastase test for detection of chronic pancreatitis. AB - INTRODUCTION: Several pancreatic function tests exist, but their role in the diagnosis of chronic pancreatitis is controversial. Here we analyzed the clinical value of a newly available fecal elastase assay. MATERIALS AND METHODS: Fecal samples from 212 patients treated in our hospital from January to September 2002 were taken. Chronic pancreatitis was assumed when ductal alterations were present in ERCP. The severity of disease was assessed according to the Cambridge classification. Elastase (test from ScheBo, Germany) and chymotrypsin (Roche Diagnostics) were measured. As a new parameter an elastase ELISA from BIOSERV (Rostock, Germany) was employed. Specificity, sensitivity, and accuracy of each test as well as the ROC curves were calculated. RESULTS: In 45 patients (21.2%) chronic pancreatitis was diagnosed. The sensitivities of elastase from ScheBo, elastase from BIOSERV and chymotrypsin were 68.9%, 77.8%, and 57.8%, respectively. The corresponding specificities were 77.2%, 76.0%, and 52.7%. When a cut-off for the elastase tests of 100 U/ml was used the sensitivities (57.8%) and the specificities (89.2%) for both elastase tests were similar. The areas under the ROC curves for the ScheBo elastase, BIOSERV elastase, and chymotrypsin were 0.805, 0.840, and 0.628, respectively. The higher AUC of the BIOSERV test was maintained when patients with mild-moderate chronic pancreatitis (Cambridge I II) were analyzed separately. CONCLUSION: The new elastase assay could probably replace the older test and was also much better than chymotrypsin. PMID- 15285177 TI - Ultra-rapid detection of Chlamydia trachomatis by real-time PCR in the LightCycler using SYBR green technology or 5'-nuclease probes. AB - Chlamydia trachomatis infections are among the most common sexually transmitted diseases and of great epidemiological importance world-wide. Identification of this pathogen can be difficult, and it is highly desirable to have a rapid and accurate nucleic acid based detection method. Several commercial PCR test systems are available (e.g. CobasAmplicor, Roche, Mannheim, Germany) but they require post-amplification detection by hybridization resulting in extended work-up time and possible cross-contamination. The objective of our study was to develop a routine diagnostic method for the sensitive, specific and rapid detection of C. trachomatis. The obvious choice is real-time PCR without any post-amplification procedures. The dye SYBR Green I (intercalating in dsDNA) provides a simple and fast real-time PCR in the LightCycler. Specific primer design combined with melting curve analysis allows a reliable and sensitive identification of C. trachomatis. In addition, a new commercial real-time PCR system (RealArt C. trachomatis LC PCR Reagents, artus, Hamburg, Germany) was evaluated, that combines sequence-specific primers and fluorescence-labelled (FRET) 5'-nuclease probes. An internal control integrated in this system detects false negative results and erroneous PCR conditions. All results were compared with the corresponding data from an analysis using the CobasAmplicor system (Roche). (Clin PMID- 15285178 TI - Evaluation of a new fully automated one-step C-peptide chemiluminescence assay (LIAISON C-Peptid). AB - The determination of C-peptide, a 31 amino acid fragment of proinsulin which is a byproduct of insulin formation, is used as a marker for insulin secretion. Clinically, the determinations are performed to detect autonomous insulinoma, factitious hypoglycemia, and in general to assess the function of beta-cells in patients with diabetes mellitus. The analysis is frequently performed by radioimmunoassays (RIA), which have several disadvantages, for instance the use of radioactivity and time and resource requirements. We performed an evaluation of a new fully automated chemiluminescence assay (LIAISON C-Peptid, Byk-Sangtec) at two clinical sites, in Germany and Italy, with regard to imprecision and clinical relevance of the obtained data, and the correlation with a standard RIA method and another chemiluminescence test. The new assay showed a good correlation with the RIA (r = 0.950) and the chemiluminescence assay (r = 0.967). The intra-assay variability and inter-assay variability was 3.5% and 8.7% in Germany, and 2.4% and 9.6% in Italy. The clinical evaluation of samples derived from 19 oral glucose tolerance tests, 13 insulin suppression tests, and 2 insulin secretion stimulation tests revealed a clinical specificity of 100%, i.e. all cases resulted in the same clinical diagnosis with all tests. With regard to the practical performance of the assays, the new chemiluminescence test, as a single step fully automated method, offered the advantage of being a non-radioactive, less complex and much faster method than the RIA and also had timely advantages over the comparative chemiluminescence test. In general, the new LIAISON chemiluminescence assay compared favorably with the RIA and comparative chemiluminescence test and offers an attractive alternative for C-peptide analysis. PMID- 15285179 TI - Evaluation of analytical performance of Super G1, Super G2+ and Biosen 5030 glucose amperometric analyzers. AB - Three glucose mono-analyzers Super G1, Super G2+ and Biosen 5030 were evaluated. The within run imprecision for the Super G1 analyzer was 0.4% to 5.6%, for the Super G2+ from 3.7% to 7.9% and for the Biosen 5030 from 1.04% to 2.97%. Between run imprecision was in the range of 4.6% to 8.6% for the Super G1 and Super G2+ analyzers and ranged from 1.68% to 2.78% for the Biosen 5030 analyzer, respectively. Analytical bias was 0.0% to 4.6% for the Super G1 and Super G2+ analyzers and 0.64% to 8.43% for the Biosen 5030, respectively. The bias of 8.43% was observed at near normal glucose concentrations. All glucose analyzers studied showed a perfect linearity of response versus blood glucose concentrations in the range from 1.4 to 16 mmol/l. The calculated differences between glucose concentrations obtained by the Super G1 and Super G2+ analyzers amounted to 0.015, 0.06 and 0.21 mmol/l at glucose concentrations of 2.5, 5 and 10 mmol/l, respectively, and differences between the Super G2+ and Biosen 5030 amounted to 0.27, 0.07 and 0.33 mmol/l, respectively. The Bland Altmann difference analysis yielded mean difference values amounting to -0.024 mmol/l; 0.036 mmol/l and 0.049 mmol/l for theSuper G1, G2+ and Biosen 5030, respectively. The carried evaluation has shown that the analytical performance of all three electrochemical glucose analyzers studied approximates the reference laboratory method. Besides of some outlying results this type of glucose meters can be reliably used as laboratory back-up analyzers and as glucose analyzers employed in various satellite locations. PMID- 15285180 TI - Diagnostic value of tests for Toxoplasma gondii-specific antibodies in patients undergoing bone marrow transplantation. AB - The value of Toxoplasma gondii-specific antibodies for the diagnosis of reactivated toxoplasmosis in immunocompromised patients is controversely discussed. The present study was performed to investigate the value of testing for antibodies against Toxoplasma gondii in patients undergoing bone marrow transplantation (BMT). Of 75 patients enrolled in the study, 53 (70.7%) were seropositive before BMT. Of these, 7 (13.2%) developed parasitemia after BMT as detected by polymerase chain reaction (PCR); three of these developed clinical symptoms compatible with reactivated toxoplasmosis that resulted in fatal toxoplasmosis in two cases. We did not detect specific antibody patterns or changes in specific antibody titers that could have predicted the development of parasitemia. Notably, 18 (81.8%) of 22 patients without specific IgG before BMT showed anti-T. gondii IgG after BMT, but none of them developed parasitemia. The results of the present study indicate that blood from both the donor and the recipient should be tested before BMT. Seronegative patients should be tested regularly after BMT to detect primary infections. Routine serological monitoring of patients seropositive before BMT does not contribute to the diagnosis of reactivated disease following BMT. In contrast, blood of seropositive patients should be screened by polymerase chain reaction (PCR) for Toxoplasma-specific DNA for the diagnosis of reactivation of infection with T. gondii. PMID- 15285181 TI - Evaluation of the hemoglobin A1c-analyzer TOSOH HLC-723 G7. AB - The TOSOH HLC-723 G7 is a compact analyzer designed for the measurement of HbA1c under routine laboratory conditions. The analyzer has an automatic blood tube supply and positive sample identification. Samples are transported automatically via racks in a continuous-load mode, cap piercing is optional. Tests devoted to the assessment of reproducibility and accuracy of analytical results indicated that over a test period of 17 days, the intra-assay variation (CV) was 1.79%, and the inter-assay variation 2.60%, respectively. A comparison with the predecessor model G5 showed a very good correlation (r = 0.997, y = 1.0041x - 0.00172; n = 149). The presence of high triglyceride, bilirubin or urea concentrations in patient samples did not influence the analytical precision. The labile HbA1c fraction (L-A1c) is clearly separated during chromatography and thus does not compromise HbA1c analysis. With a protocol of 1.2 minutes, the TOSOH G7 is a very fast analyzer, designed for laboratories with a high throughput of samples. PMID- 15285182 TI - Decision limit for troponin I on ADVIA:Centaur and evaluation of the analytical precision at low concentrations. AB - Recent guidelines have defined that the decisional cut-off of cardiac troponin (cTn) would recognize "the normal" subjects, and in theory they do not have "measurable" values of cTn. The 99th percentile of this population is chosen as cut-off but using an assay with an analytical CV < 10% at this value. Objectives of this study were to set the decisional limits for cTroponin I (cTnI) measured by the ADVIA:Centaur, and to evaluate the analytical precision around these limits. 120 normal plasma samples were tested for cTnI levels. The 99th percentile defined the decisional level, according to the recent guidelines. The precision was estimated by 10 replicates for 21 samples. The 99th percentile was 0.17 microg/L, with an analytical CV <10%. Since this analytical method achieves the recommended analytical precision, the cTnI decision level for myocardial damage by the ADVIA:Centaur is 0.17 microg/L. PMID- 15285183 TI - Oxidative stress in hemodialysis--postdialytic changes. AB - BACKGROUND: There is evidence for production of free oxygen radicals during hemodialysis. Hemodialysis is an intervention that is intermittent and is usually undertaken once in two or three days. It is known that the free oxygen radicals are short lived. Hence, it is necessary to know how long the effects of this oxidative stress are seen in the postdialytic period and whether they are carried over to the next dialysis. Review of the literature showed that there is no information in this area. Hence, this study was undertaken in order to learn whether oxidative stress due to a dialysis session is carried over to next dialysis session or not. METHODS: The effects were studied after four different types of membrane and dialysate--Polysulphone-Bicarbonate (PB), Polysulphone Acetate (PA), Cuprophan-Acetate (CA) and Cuprophan-Bicarbonate (CB). Two consecutive dialysis sessions were studied to know the effect of re-use of the membrane. For each dialysis session, blood samples were collected at 0 (immediately prior to dialysis or preHD), 4 (immediate postdialysis), 6, 12, 24 and 48 hours (start of next session). Lipid peroxides, SOD and GP were determined in erythrocytes. Vitamins A and E and lipid peroxides were estimated in plasma. RESULTS: In the postdialytic phase there was an increase in plasma lipid peroxide levels. Plasma vitamin E levels increased significantly in all groups after first use dialysis, whereas the increase found after re-use dialysis was not statistically significant. Erythrocyte lipid peroxide levels showed a significant decrease. No significant changes were observed in the plasma vitamin A, erythrocyte SOD and GP levels. There was no significant change in any of the parameters between preHD and either 48-hour or 96-hour samples in all groups studied. CONCLUSIONS: Our results show that there is no carry-over of oxidative stress produced by dialysis to the next session regardless of the type of dialysis. PMID- 15285184 TI - The use of biochips opens up new dimensions in diagnostic practice. AB - The past decades have reflected a continuous increase in healthcare costs that is dramatic compared to growth rates of the national economy. Not only this, the legislative demands made on medical services and medical devices have become far more complex. All this makes a strong impact on the field of laboratory diagnostics. Although scientific and technical possibilities have greatly improved, they have done little to reduce costs. PMID- 15285185 TI - The norovirus on the march: triggers of acute diseases of the stomach and intestine. AB - Gastroenteritis (diseases of the stomach and intestine) is widespread all over the world and one of the most frequent causes of infant mortality in the Third World. In industrial countries, gastroenteritis occurs second to respiratory diseases in children, rarely, however, with any complications. PMID- 15285186 TI - Antibodies to Chlamydia trachomatis heat shock protein 60 (cHSP60) and Chlamydia trachomatis major outer membrane protein (MOMP) in women with different tubal status. PMID- 15285187 TI - Antibodies to Chlamydia trachomatis heat shock protein (cHSP60) and major outer membrane protein (MOMP) in men with impaired fertility. PMID- 15285188 TI - The extended Chlamydia trachomatis diagnosis in patients with pelvic inflammatory disease--a better approach for the diagnosis of upper genital tract infections. PMID- 15285189 TI - Rights of patients in the European context, ten years and after. PMID- 15285190 TI - Patients' rights and patients' participation. Individual and collective involvement: partnership and participation in health law. PMID- 15285191 TI - The right to health care. PMID- 15285192 TI - Medical confidentiality--quo vadis? PMID- 15285193 TI - Communicable diseases and human rights. PMID- 15285194 TI - Wrongful birth and wrongful life actions. PMID- 15285195 TI - Medical research involving human beings: some reflections on the main principles of the international regulatory instruments. PMID- 15285196 TI - Genetics, tissue- and databases. PMID- 15285197 TI - Health legislation in Eastern European countries: the Baltic States. PMID- 15285198 TI - Genetic databases: assessing the benefits and the impact on human and patient rights--a WHO report. PMID- 15285199 TI - International Declaration on Human Genetic Data. PMID- 15285200 TI - CJEC 2004/1, case C-322/00, Deutscher Apothekerverband ev/0800 Doc Morris NV. PMID- 15285201 TI - CJEC 2004/2, case T-326/99, Olivieri/Commisssion. PMID- 15285202 TI - ECHR 2004/1 case of Arcila Henao v. The Netherlands, 24 June 2003, no. 13669/03 (second section). PMID- 15285203 TI - ECHR 2004/2 case of Sentges v. The Netherlands, 8 July 2003, no.27677/02 (second section). PMID- 15285204 TI - ECHR 2004/3 case of Tkacik v. Slovakia, 14 October 2003, no. 42472/98 (fourth section). PMID- 15285205 TI - ECHR 2004/4 case of I.P. v. Poland, 14 October 2003, no. 77831/01 (fourth section). PMID- 15285206 TI - ECHR 2004/5 case of Rakevich v. Russia, 28 October 2003, no. 58973/00 (second section). PMID- 15285207 TI - Rewarding results: Improving the quality of treatment for people with alcohol and drug problems. AB - Substance use disorders are the nation's number one health problem, and lie at the root of many public safety and workplace issues. Improving quality of treatment is as important as improving access to treatment. Leadership for improvement must come from many sources: Congress, SAMHSA, state legislatures, state and local treatment agencies, criminal justice, welfare and other public agencies, employers and managed care organizations, providers, and community leaders. We hope that our report helps leaders see ways to improve treatment quality. Our recommendations can be summed up in a single phrase: reward results. We recognize that there are many avenues for treatment quality improvement, including training, credentialing, best practice dissemination, work force development, facility licensing standards, improvement and implementation of new models for treatment of dual diagnosis patients. We believe, however, that rewarding results is essential to motivating action for improvement. We also believe that if providers receive rewards for improved results, they will creatively open new avenues for improvement--a focus on results gives greater freedom than more detailed mandates for change. Finally, we believe that rewards for result may lead to a restructured treatment system with greater stability and correspondingly greater capacity to improve. While we have placed central emphasis on the role of institutional buyers and managers of care, we believe that the voices of patients and families must be heard. People who have progressed to the stage of recovery, and their families, often have essential insight into what did and did not work for them--their personal stories are frequently compelling and persuasive. We also believe that providers of treatment for substance use disorders are profoundly committed to serving their patients, and often have great understanding of what works. Wise managers will listen very carefully and systematically to the voices of consumers, their families, and their providers. We have advocated that buyers reward results as a central strategy for improving quality. We believe that this is the best long-term strategy, not only to effect quality improvement, but also to end stigmatization and increase resources. We wish, in closing, to reemphasize our shared beliefs that adequate resources are essential, that treating persons with substance use disorders is always emotionally challenging and often profoundly frustrating and alienating, and that those on the front line deserve our unequivocal support and our profound gratitude for their service. Legislators should adequately fund public treatment and mandate parity in insurance reimbursement for treatment of substance use disorders. PMID- 15285208 TI - Outcomes from a therapeutic community for homeless addicted mothers and their children. AB - A women's therapeutic community (TC) designed to prevent homelessness was evaluated using a quasi-experimental process. Propensity analysis selected comparable experimental (E) and comparison (C) participants. Significant improvements were found for the E group at the domain level, both in "psychological" dysfunction on symptoms (e.g., depression), and in "health," including ratings of health and adherence to medication regimens. No significant difference was found at the domain level for "parenting" or "housing stabilization," but specific outcomes did differ. For example, a greater number of children resided with the E group mothers who also assumed financial responsibility for more of their children. PMID- 15285209 TI - Substance use and five-year survival in Washington State mental hospitals. AB - This report combines five-year survival and cause of death in individuals discharged from Washington State mental hospitals with (1) mental illness only, (2) co-occurring mental illness and substance use disorder, or (3) substance use disorder only. Five-year survival was similar in the three groups, although after adjusting for age, individuals with co-occurring disorders or substance use disorder were almost 50% more likely to die than those with mental illness only. Persons with these conditions need treatment to prevent premature death and medical conditions directly related to substance use disorder. PMID- 15285210 TI - Harm reduction: a historic parallel and parable. PMID- 15285211 TI - Self-help groups in Hong Kong. PMID- 15285212 TI - The Howard Gilman Foundation Lecture. Where have we come from and where are we going? Valve management past, present and future. AB - Leonardo da Vinci's anatomical drawings of quadricuspid, tricuspid, and bicuspid aortic valves underscored the hydraulic superiority of a three leaflet valve with cuspal equality. William Harvey demonstrated that venous valves were designed for unidirectional flow and to prevent reflux from the heart, observations that served as the basis of his immortal de Mortu Cordis. Joseph Rouanet of Paris proposed that heart sounds originated from the closing movements of cardiac valves. The Cardiodynamics of Mitral Insufficiency by Wiggers and Feil was followed three decades later by Paul Wood's An Appreciation of Mitral Stenosis. The Bland/Sweet operation indirectly addressed mitral stenosis by means of a venous shunt. Sir Henry Souttar's early digital repair of mitral stenosis was later reintroduced independently by Harken and Bailey; Doyen, Sellers, and Brock employed surgical valvotomy for pulmonary stenosis, and Bailey employed surgical valvotomy for aortic stenosis. Management of abnormal cardiac valves includes repair (reconstruction), replacement with mechanical or biologic prostheses, and interventional catheterization. The first mechanical valve was inserted extracardiac by Hufnagel into the descending thoracic aorta of patients with severe aortic regurgitation. The Starr caged ball mechanical prosthesis was designed for intracardiac replacement of an abnormal cardiac valve. The peripheral flow ball valve was followed by hydraulically superior and less thrombogenic central flow monoleaflet or bileaflet mechanical valves, and by homograft and heterograft bioprosthetic valves. Improved methods of preparing exogenous bioprostheses and innovative techniques of aortic valve reconstruction are evolving. Cardiac catheterization as a therapeutic intervention is routinely applied to stenotic mitral, aortic and pulmonary valves, and transcatheter replacement of an abnormal pulmonary valve is now a reality. PMID- 15285213 TI - The epidemiology of valvular heart diseases: the problem is growing. PMID- 15285214 TI - Heart failure in aortic regurgitation: the role of primary fibrosis and its cellular and molecular pathophysiology. PMID- 15285215 TI - Is prophylactic beta-adrenergic blockade appropriate in mitral regurgitation: impact of cellular pathophysiology. PMID- 15285216 TI - Valve surgery in the asymptomatic patient with aortic regurgitation: current indications and the effect of change rates in objective measures. PMID- 15285217 TI - Valve repair versus replacement when aortic regurgitation is caused by aortic root aneurysms: relative advantages and disadvantages and the impact of decision on surgical indications. PMID- 15285218 TI - Assessment of myocardial damage in regurgitant valvular disease. PMID- 15285219 TI - Cholesterol-lowering studies for aortic stenosis. PMID- 15285221 TI - Natural history of mitral stenosis and echocardiographic criteria and pitfalls in selecting patients for balloon valvuloplasty. AB - In summary, MS is a progressive disease characterized by a long latent period between the initial attack of acute rheumatic fever and the development of symptoms. For patients with MS who require mechanical relief of obstruction PMV is the preferred treatment and gives results comparable to surgical commissurotomy. Two-dimensional echocardiographic assessment of mitral valve morphology is the most important predictor of outcome. An echocardiographic score <8 predicts good immediate and long-term results. In patients undergoing PMV, TEE is useful for detecting left atrial and left atrial appendage thrombi, guiding transseptal puncture, assessing results and detecting complications. PMID- 15285220 TI - Perspectives on diseases of the thoracic aorta. PMID- 15285222 TI - Surgical treatment of degenerative mitral regurgitation: should we approach differently patients with flail leaflets of simple mitral valve prolapse? PMID- 15285223 TI - Ventricular arrhythmias in mitral regurgitation: frequency, clinical and prognostic importance, management before and after mitral valve surgery. PMID- 15285224 TI - The case for bioprosthetic mitral valve replacement in patients aged 60-70. PMID- 15285225 TI - Endocardial cushion defects: embryology,anatomy and pathophysiology. PMID- 15285226 TI - Surgery for atrioventricular septal defects. PMID- 15285227 TI - When should tricuspid valve replacement/repair accompany mitral valve surgery? PMID- 15285228 TI - Strategies for management of postcardiotomy cardiogenic shock following valvular heart surgery. PMID- 15285229 TI - Neurological dysfunction after coronary artery bypass surgery: facts vs. fiction. PMID- 15285230 TI - Robotic valve surgery: how does the future look? PMID- 15285231 TI - Selection of techniques for combined valve surgery and coronary artery bypass grafting: the impact of combined procedures involving the aortic or mitral valve. PMID- 15285232 TI - Aortic valve and non-ischemic mitral valve surgery in patients undergoing coronary artery bypass grafting. PMID- 15285233 TI - Echocardiographic doppler evaluation of prosthetic valve function and dysfunction. PMID- 15285234 TI - Massive rectal bleeding from a Dieulafoy's ulcer of the rectum. AB - Massive per rectal bleeding caused by a Dieulafoy's ulcer located within the rectum is extremely rare. We herein report such a case occurring in a 76-year-old male patient with a history of chronic renal failure, who presented with massive fresh bleeding in the rectum. He was diagnosed during an endoscopic inspection and was promptly treated by clipping at the same time. Although this is a rare entity, Dieulafoy's ulcer should therefore be taken into consideration in the differential diagnosis of patients presenting with massive lower gastrointestinal bleeding. PMID- 15285235 TI - Gunshot liver trauma with disruption of the right hepatic duct managed by surgery, radiology, and endoscopy: a case report. AB - During the process of treating a complicated gunshot wound of an upper limb, chest, abdomen, and spine, there appeared--sometime after the initial treatment- a necrosis of the right hepatic duct in the bullet path within the liver. Although laparotomy was the life-saving operation during the first and second period, the final diagnosis and solution were based on bypassing the defect, i.e., a combination of a percutaneous and endoscopic approach. The efficiency of this method was also proved by an examination carried out 1 year after the end of the treatment. PMID- 15285236 TI - Primary torsion of the greater omentum. AB - Primary torsion of the greater omentum is an uncommon cause of acute abdominal pain, often mimicking other acute abdominal conditions. There has been little in the English literature concerning diagnostic imaging of the torsion of the greater omentum because it is not usually diagnosed until surgically operated on for acute abdomen that has been interpreted as appendicitis. Resection of the infracted segment is the treatment of choice, offering rapid recovery and reducing the possibility of adhesion formation. We report a case of torsion of the greater omentum that was diagnosed correctly with preoperative computed tomography and discuss the therapeutic implications of this entity. PMID- 15285237 TI - Combined endoscopic approach for the management of a difficult recto-sigmoid anastomotic stricture. AB - A 48-year-old woman underwent an emergency laparotomy for a perforated diverticular abscess. At operation, a peritoneal lavage was carried out, but no colonic resection was undertaken. Subsequently, she developed recurrent sepsis and underwent a second laparotomy. The patient was referred to our institution for a definitive left hemicolectomy and diverting loop colostomy. Before closing the colostomy, a contrast enema revealed obstruction of the lumen at the anastomotic site. This was refractory to conventional colonoscopic dilatation. A simultaneous endoscope was passed through the distal loop of the colostomy in addition to a conventional approach. A defect was created that allowed passage of a guide-wire and balloon dilator. One week later, the anastomosis remained patent, and the colostomy closed. The patient remains well, with normal bowel function. This novel combined endoscopic approach for dealing with colonic stenoses avoids the higher morbidity and mortality associated with an open surgical procedure. PMID- 15285238 TI - Symptomatic fibroid uterus in a 15-year-old girl. AB - A 15 year-old gravida 1, abortus 1 black girl presented with chief complaints of menorrhagia, severe dysmenorrhea, and progressively worsening abdominal pain, which was caused by a very large uterine leiomyoma. The symptoms began 6 months earlier, shortly after a 10-week spontaneous abortion at age 14. A solitary 25-cm uterine leiomyoma was removed uneventfully with an abdominal laparotomy. In the English literature of the past 50 years, this case represents the sixth, and we believe, the largest, documented uterine fibroid among teenagers, which required corrective surgery. PMID- 15285239 TI - Report of 100 open inguinal hernia repairs using a 2-cm incision: a novel technique. AB - We report a novel open technique for the repair of inguinal hernias using a 2-cm incision. This allows bilateral cases to be performed safely as day case procedures, where laparoscopic repair is not possible. A 2-cm incision is made over the deep inguinal ring. Blunt dissection distracts the cord and hernial sac, which can be dissected and reduced. A stapler is used to secure mesh through an incision too small to allow satisfactory suturing. This is a fast, safe, and easy to-learn technique that produces a good repair with minimal bruising and quick wound healing. It is a useful substitute to laparoscopic totally extraperitoneal procedure (TEP) repair where this is not possible. PMID- 15285240 TI - Indigenous cost-effective peritoneo-venous shunt for refractory ascites. AB - About 5% of patients with chronic liver disease develop massive refractory ascites. These patients cease to respond to diuretic therapy and may develop prerenal azotemia. There is a small but definite role for the peritoneo-venous shunt in these patients. In our study of 36 patients, managed with locally made, single-valved peritoneo-venous shunts (GSAIMS shunts), shunt failure and complication rates were assessed postoperatively. There is a definite improvement in quality of life with this cost-effective locally made shunt if patients are selected carefully. Long-term follow-up of these patients is not possible because most of these patients succumb to advanced liver disease. PMID- 15285241 TI - Omental wrapping of skeletonized major vessels after pancreaticoduodenectomy. AB - Occasionally, a pancreatic leakage results in a poor outcome. Even today, its rate of occurrence ranges from 2 to 13%. The purpose of this study is to describe the omental-wrapping technique and to evaluate its clinical utility for preventing serious complications following pancreatic leakage. The clinical records of 54 patients who underwent this technique were reviewed for their short term outcome. After completing all reconstruction, an omental flap was spread out behind the pancreaticojejunostomy, allowing complete isolation of the skeletonized major vessels from the pancreatic anastomosis. There was no operative death or intra-abdominal bleeding. Pancreatic leakage occurred in 5 patients, all associated with bacterial infection. However, drains placed in the retroperitoneal space, which was isolated from the pancreatic anastomosis by the flap, were aseptic in all patients. The pancreatic leakage was closed with no additional serious conditions related to the leak. Our studies demonstrate that the omental wrapping technique is one of the useful surgical options for preventing serious conditions following a pancreatic leakage. PMID- 15285242 TI - Strategy for the treatment of unresectable hepatoblastoma: neoadjuvant chemotherapy followed by delayed primary operation or liver transplantation. AB - We present our experience in using neoadjuvant regional and systemic chemotherapy together with surgical resection as a strategy for the treatment of unresectable hepatoblastoma. Neoadjuvant chemotherapy was given prior to surgical treatment in six children with unresectable hepatoblastoma. Furthermore, the neoadjuvant chemotherapy was intensified according to response to the initial treatment. Surgical resection was performed when the tumor was judged to be resectable. The adjuvant chemotherapy was given after delayed primary operation. Five of six children receiving neoadjuvant chemotherapy responded to the treatment and subsequently received delayed primary operation or living donor liver transplantation. All five children who had successful surgery have completed treatment and show no evidence of disease to date (27-115 months after surgery). It is concluded that neoadjuvant chemotherapy given as a combination of regional and systemic chemotherapy was effective for tumor reduction in cases with early stage or stage III disease. Also, to increase the cure rate of children with localized disease that was still unresectable after chemotherapy, living donor liver transplantation, which offers some advantage in timing of transplant compared with cadaveric liver transplantation, seems to be a possible procedure. PMID- 15285243 TI - Microsatellite instability and survival rate in the solid or nonsolid types of poorly differentiated colorectal adenocarcinoma. AB - Poorly differentiated colorectal adenocarcinoma accounts for 5-10% of all cases of colorectal cancer, and its poor prognosis is known, but there are many unclear points regarding its biological features. Because the histology of colorectal cancer is roughly classified into two patterns (severe and mild interstitial infiltration), in this study, we focused on this point and investigated clinicopathological findings and microsatellite instability (MSI) in the two interstitial infiltration patterns of poorly differentiated colorectal adenocarcinoma. We encountered 502 patients with colorectal cancer, and 30 patients (6%) who were diagnosed with the dominant histologic type of poorly differentiated colorectal adenocarcinoma were selected as subjects. The lesion was not resectable in three patients, and thus, the clinicopathology was unknown in these patients. In the 27 patients who underwent resection of poorly differentiated colorectal adenocarcinoma, the cancers were histologically classified based on the interstitial infiltration patterns into solid and non solid types, and the clinicopathology and survival rates were investigated. DNA was extracted from the primary lesions, and normal tissue and MSI were investigated using five microsatellite markers. Histopathologically, 20 cancers were classified as the solid type and 7 as the nonsolid type. In the clinicopathological findings in the nonsolid and solid types, lymph node metastasis was observed in 100% and 60%; liver metastasis in 57% and 20%, and peritoneal dissemination in 29% and 5%, respectively, showing that the positive rates were higher in the nonsolid type. The 5-year survival rate was also poor in the nonsolid type. MSI was observed in 50% of the solid type and 20% of the nonsolid type, showing a high frequency in the solid type. Poorly differentiated colorectal adenocarcinoma was classified based on the infiltration pattern into the nonsolid and solid types. In the nonsolid type, the cancer was advanced in many cases, and the frequency of MSI was lower than that of the solid type. Therefore, there were differences in clinico-pathological features and biological malignancy between the nonsolid and solid types, suggesting different carcinogenesis mechanisms for the two types. PMID- 15285244 TI - Tensionless Spigelian herniorrhaphy using a bilayered prosthetic patch: historical, anatomical, diagnostic, and operative perspectives. AB - Spigelian hernias, which represent <2% of all hernias of the vellum abdominis (abdominal wall) anterior, can be a diagnostic challenge for clinicians. Noninvasive imaging techniques, including ultrasonography (US) and computerized axial tomography (CAT), substantially complement clinical inferences based on interrogation and physical examination. Successful definitive care mandates comprehension of the regional, topographical, and visceral anatomy in axial, coronal, and transverse planes. Reported herein is the successful use of a bilayered prosthetic patch, advantageous because of its unimodular and biplanar configuration, to perform a tensionless herniorrhaphy. PMID- 15285245 TI - Handgrip strength and endurance as a predictor of postoperative morbidity in surgical patients: can it serve as a simple bedside test? AB - Postoperative morbidity is related to preoperative nutritional status. Current methods of assessing this are cumbersome, lack sufficient accuracy to completely separate all those at risk from others, and require a laboratory backup. This study was done to evaluate handgrip manometry, a simple bedside test as an indicator of preoperative nutritional status to predict risk of postoperative complications. Normal values for maximal grip strength and endurance times were established in 496 controls. Similar values were recorded in 100 patients undergoing elective surgery preoperatively. Postoperative variables studied included complication rate and hospital stay. The efficacy of four parameters, serum albumin, clinical nutritional score based on subjective global assessment, maximal grip strength (MGS), and grip endurance time, in predicting complications were studied. It was found that age and presence of co-morbidity increased risk of complications. Of the four predictive parameters studied, it was found that MGS is better than serum albumin. Clinical scoring was superior to MGS in predicting risk of complications. MGS is a simple bedside test, which can be easily performed. It can be used as a complimentary test to clinical scoring in identifying patients at risk of complications after surgery. Patients with abnormal MGS require urgent preoperative correction to reduce the risk of complications. PMID- 15285246 TI - Multi-modality tissue-mimicking phantom for thermal therapy. AB - A tissue-mimicking phantom material has been developed for use with thermal therapy devices and techniques. This material has magnetic resonance properties (primarily T2) which change drastically upon thermal coagulation, enabling its use for device characterization and treatment verification using simple T2 weighted imaging techniques. The coagulation temperature of the phantom can be changed from 50-60 degrees C by adjusting the pH from 4.3 to 4.7. The energy absorption properties can be adjusted to match the acoustical and optical properties of tissues. T2 relaxation measurements are provided as a function of temperature, along with T2-weighted MR images to illustrate the visualization of heating patterns. A complete recipe for fabricating phantoms is provided. PMID- 15285247 TI - Electromagnetic fields in the human body due to switched transverse gradient coils in MRI. AB - Magnetic resonance imaging scans impose large gradient magnetic fields on the patient. Modern imaging techniques require this magnetic field to be switched rapidly for good resolution. However, it is believed that this can also lead to the unwanted side effect of peripheral nerve stimulation, which proves to be a limiting factor to the advancement of MRI technology. This paper establishes an analytical model for the fields produced within an MRI scanner by transverse gradient coils of known current density. Expressions are obtained for the magnetic induction vector and the electric field vector, as well as for the surface charge and current densities that are induced on the patient's body. The expressions obtained are general enough to allow the study of any combination of gradient coils whose behaviour can be approximated by Fourier series. For a realistic example coil current density and switching function, it is found that spikes of surface charge density are induced on the patient's body as the gradient field is switched, as well as loops of surface current density that mimic the coil current density. For a 10 mT m(-1) gradient field with a rise time of 100 micros, the magnitude of the radial electric field at the body is found to be 10.3 V m(-1). It is also found that there is a finite limit to radial electric field strength as rise time approaches zero. PMID- 15285248 TI - Neural attractor network for application in visual field data classification. AB - The purpose was to introduce a novel method for computer-based classification of visual field data derived from perimetric examination, that may act as a 'counsellor', providing an independent 'second opinion' to the diagnosing physician. The classification system consists of a Hopfield-type neural attractor network that obtains its input data from perimetric examination results. An iterative relaxation process determines the states of the neurons dynamically. Therefore, even 'noisy' perimetric output, e.g., early stages of a disease, may eventually be classified correctly according to the predefined idealized visual field defect (scotoma) patterns, stored as attractors of the network, that are found with diseases of the eye, optic nerve and the central nervous system. Preliminary tests of the classification system on real visual field data derived from perimetric examinations have shown a classification success of over 80%. Some of the main advantages of the Hopfield-attractor-network-based approach over feed-forward type neural networks are: (1) network architecture is defined by the classification problem; (2) no training is required to determine the neural coupling strengths; (3) assignment of an auto-diagnosis confidence level is possible by means of an overlap parameter and the Hamming distance. In conclusion, the novel method for computer-based classification of visual field data, presented here, furnishes a valuable first overview and an independent 'second opinion' in judging perimetric examination results, pointing towards a final diagnosis by a physician. It should not be considered a substitute for the diagnosing physician. Thanks to the worldwide accessibility of the Internet, the classification system offers a promising perspective towards modern computer assisted diagnosis in both medicine and tele-medicine, for example and in particular, with respect to non-ophthalmic clinics or in communities where perimetric expertise is not readily available. PMID- 15285249 TI - A phenomenological model for the relative biological effectiveness in therapeutic proton beams. AB - To study the effects of a variable relative biological effectiveness (RBE) in inverse treatment planning for proton therapy, fast methods for three-dimensional RBE calculations are required. We therefore propose a simple phenomenological model for the RBE in therapeutic proton beams. It describes the RBE as a function of the dose, the linear energy transfer (LET) and tissue specific parameters. Published experimental results for the dependence of the parameters alpha and beta from the linear-quadratic model on the dose averaged LET were evaluated. Using a linear function for alpha(LET) in the relevant LET region below 30 keV per micrometre and a constant beta, a simple formula for the RBE could be derived. The new model was able to reproduce the basic dependences of RBE on dose and LET, and the RBE values agreed well with experimental results. The model was also applied to spread-out Bragg peaks (SOBP), where the main effects of a variable RBE are an increase of the RBE along the SOBP plateau, and a shift in depth of the distal falloff. The new method allows fast RBE estimations and has therefore potential applications in iterative treatment planning for proton therapy. PMID- 15285250 TI - Effect on tumour control of time interval between surgery and postoperative radiotherapy: an empirical approach using Monte Carlo simulation. AB - In this work, a procedure, based on Monte Carlo techniques, to analyse the effect on the tumour control probability of the time interval between surgery and postoperative radiotherapy is presented. The approach includes the tumour growth as well as the survival of tumour cells undergoing fractionated radiotherapy. Both processes are described in terms of the binomial distribution. We have considered two different growth models, exponential and Gompertz, the parameters of which have been fixed to reproduce the clinical outcome corresponding to a retrospective study for patients with head and neck squamous cell carcinomas. In the cases analysed, we have not found significant differences between the results obtained for both growth models. The mean doubling times found for residual clonogens after surgery are less than 40 days. The rate of decrease in local control is around 0.09% per day of delay between surgery and radiotherapy and the corresponding time factor is about 0.11 Gy per day. PMID- 15285251 TI - A new analytical model for Varian enhanced dynamic wedge factors. AB - Dynamic and physical (hard) wedges are used in 3D conformal radiotherapy in order to improve dose distribution in patients. Unlike wedge factors for physical wedges that depend on wedge material and thickness, wedge factors for Varian dynamic wedges depend on the relationship between the position of the moving jaw and the number of delivered monitor units. In this study, we describe a new analytical model for dynamic wedge factors. We also review the existing analytical models and compare calculated and measured wedge factors. The comparison is performed for different wedge angles, symmetric and asymmetric fields and two different photon energies. The obtained results indicate that the new dynamic wedge model provides the best overall agreement (within 1%) with the measured wedge factors. PMID- 15285252 TI - Degeneracy, frequency response and filtering in IMRT optimization. AB - This paper attempts to provide an answer to some questions that remain either poorly understood, or not well documented in the literature, on basic issues related to intensity modulated radiation therapy (IMRT). The questions examined are: the relationship between degeneracy and frequency response of optimizations, effects of initial beamlet fluence assignment and stopping point, what does filtering of an optimized beamlet map actually do and how could image analysis help to obtain better optimizations? Two target functions are studied, a quadratic cost function and the log likelihood function of the dynamically penalized likelihood (DPL) algorithm. The algorithms used are the conjugate gradient, the stochastic adaptive simulated annealing and the DPL. One simple phantom is used to show the development of the analysis tools used and two clinical cases of medium and large dose matrix size (a meningioma and a prostate) are studied in detail. The conclusions reached are that the high number of iterations that is needed to avoid degeneracy is not warranted in clinical practice, as the quality of the optimizations, as judged by the DVHs and dose distributions obtained, does not improve significantly after a certain point. It is also shown that the optimum initial beamlet fluence assignment for analytical iterative algorithms is a uniform distribution, but such an assignment does not help a stochastic method of optimization. Stopping points for the studied algorithms are discussed and the deterioration of DVH characteristics with filtering is shown to be partially recoverable by the use of space-variant filtering techniques. PMID- 15285253 TI - Characterization of dual layer phoswich detector performance for small animal PET using Monte Carlo simulation. AB - A positron emission tomograph dedicated to small animal imaging should have high spatial resolution and sensitivity, and dual layer scintillators have been developed for this purpose. In this study, simulations were performed to optimize the order and the length of each crystal of a dual layer phoswich detector, and to evaluate the possibility of measuring signals from each layer of the phoswich detector. A simulation tool GATE was used to estimate the sensitivity and resolution of a small PET scanner. The proposed scanner is based on dual layer phoswich detector modules arranged in a ring of 10 cm diameter. Each module is composed of 8 x 8 arrays of phoswich detectors consisting of LSO and LuYAP with a 2 mm x 2 mm sensitive area coupled to a Hamamatsu R7600-00-M64 PSPMT. The length of the front layer of the phoswich detector varied from 0 to 10 mm at 1 mm intervals, and the total length (LSO + LuYAP) was fixed at 20 mm. The order of the crystal layers of the phoswich detector was also changed. Radial resolutions were kept below 3.4 mm and 3.7 mm over 8 cm FOV, and sensitivities were 7.4% and 8.0% for LSO 5 mm-LuYAP 15 mm, and LuYAP 6 mm-LSO 14 mm phoswich detectors, respectively. Whereas, high and uniform resolutions were achieved by using the LSO front layer, higher sensitivities were obtained by changing the crystal order. The feasibilities for applying crystal identification methods to phoswich detectors consisting of LSO and LuYAP were investigated using simulation and experimentally derived measurements of the light outputs from each layer of the phoswich detector. In this study, the optimal order and lengths of the dual layer phoswich detector were derived in order to achieve high sensitivity and high and uniform radial resolution. PMID- 15285254 TI - Non-invasive determination of the irradiation dose in fingers using low-frequency EPR. AB - Several reports in the literature have described the effects of radiation in workers who exposed their fingers to intense radioactive sources. The radiation injuries occurring after local exposure to a high dose (20 to 100 Gy) could lead to the need for amputation. Follow-up of victims needs to be more rational with a precise knowledge of the irradiated area that risks tissue degradation and necrosis. It has been described previously that X-band electron paramagnetic resonance (EPR) spectroscopy could be used to assess the dose in irradiated amputated fingers. Here, we propose the use of low-frequency EPR spectroscopy to evaluate non-invasively the absorbed dose. Low-frequency microwaves are indeed less absorbed by water and penetrate more deeply into living material (approximately 10 mm in tissues using 1 GHz spectrometers). This work presents preliminary results obtained with baboon and human fingers compared with human dry phalanxes placed inside a surface-coil resonator. The EPR signal increased linearly with the dose. The ratio of the slopes of the dry bone to whole finger linear regression lines was around 5. The detection limit achievable with the present spectrometer and resonator is around 60 Gy, which is well within the range of accidentally exposed fingers. It is likely that the detection limit could be improved in the future, thanks to further technical spectrometer and resonator developments as well as to appropriate spectrum deconvolution into native and dosimetric signals. PMID- 15285255 TI - The most likely path of an energetic charged particle through a uniform medium. AB - Presented is a calculation of the most likely path for a charged particle traversing a uniform medium and suffering multiple-Coulomb scattering when the entrance and exit positions and angles are known. The effects of ionization energy loss are included and the results are verified using Monte Carlo simulation. The application to proton computed tomography is discussed. PMID- 15285256 TI - Exact and approximate algorithms for helical cone-beam CT. AB - This paper concerns image reconstruction for helical x-ray transmission tomography (CT) with multi-row detectors. We introduce two approximate cone-beam (CB) filtered-backprojection (FBP) algorithms of the Feldkamp type, obtained by extending to three dimensions (3D) two recently proposed exact FBP algorithms for 2D fan-beam reconstruction. The new algorithms are similar to the standard Feldkamp-type FBP for helical CT. In particular, they can reconstruct each transaxial slice from data acquired along an arbitrary segment of helix, thereby efficiently exploiting the available data. In contrast to the standard Feldkamp type algorithm, however, the redundancy weight is applied after filtering, allowing a more efficient numerical implementation. To partially alleviate the CB artefacts, which increase with increasing values of the helical pitch, a frequency-mixing method is proposed. This method reconstructs the high frequency components of the image using the longest possible segment of helix, whereas the low frequencies are reconstructed using a minimal, short-scan, segment of helix to minimize CB artefacts. The performance of the algorithms is illustrated using simulated data. PMID- 15285257 TI - Quality assurance of a helical tomotherapy machine. AB - Helical tomotherapy has been developed at the University of Wisconsin, and 'Hi Art II' clinical machines are now commercially manufactured. At the core of each machine lies a ring-gantry-mounted short linear accelerator which generates x rays that are collimated into a fan beam of intensity-modulated radiation by a binary multileaf, the modulation being variable with gantry angle. Patients are treated lying on a couch which is translated continuously through the bore of the machine as the gantry rotates. Highly conformal dose-distributions can be delivered using this technique, which is the therapy equivalent of spiral computed tomography. The approach requires synchrony of gantry rotation, couch translation, accelerator pulsing and the opening and closing of the leaves of the binary multileaf collimator used to modulate the radiation beam. In the course of clinically implementing helical tomotherapy, we have developed a quality assurance (QA) system for our machine. The system is analogous to that recommended for conventional clinical linear accelerator QA by AAPM Task Group 40 but contains some novel components, reflecting differences between the Hi-Art devices and conventional clinical accelerators. Here the design and dosimetric characteristics of Hi-Art machines are summarized and the QA system is set out along with experimental details of its implementation. Connections between this machine-based QA work, pre-treatment patient-specific delivery QA and fraction-by fraction dose verification are discussed. PMID- 15285258 TI - Evaluation of the adjoint equation based algorithm for elasticity imaging. AB - Recently a new adjoint equation based iterative method was proposed for evaluating the spatial distribution of the elastic modulus of tissue based on the knowledge of its displacement field under a deformation. In this method the original problem was reformulated as a minimization problem, and a gradient-based optimization algorithm was used to solve it. Significant computational savings were realized by utilizing the solution of the adjoint elasticity equations in calculating the gradient. In this paper, we examine the performance of this method with regard to measures which we believe will impact its eventual clinical use. In particular, we evaluate its abilities to (1) resolve geometrically the complex regions of elevated stiffness; (2) to handle noise levels inherent in typical instrumentation; and (3) to generate three-dimensional elasticity images. For our tests we utilize both synthetic and experimental displacement data, and consider both qualitative and quantitative measures of performance. We conclude that the method is robust and accurate, and a good candidate for clinical application because of its computational speed and efficiency. PMID- 15285259 TI - A second-order finite element algorithm for solving the three-dimensional EEG forward problem. AB - A finite element algorithm has been developed to solve the electroencephalogram (EEG) forward problem. A new computationally efficient approach to calculate the stiffness matrix of second-order tetrahedral elements has been developed for second-order tetrahedral finite element models. The present algorithm has been evaluated by means of computer simulations, by comparing with analytic solutions in a multi-spheres concentric head model. The developed finite element method (FEM) algorithm has also been applied to address questions of interest in the EEG forward problem. The present simulation study indicates that the second-order FEM provides substantially enhanced numerical accuracy and computational efficiency, as compared with the first-order FEM for comparable numbers of tetrahedral elements. The anisotropic conductivity distribution of the head tissue can be taken into account in the present FEM algorithm. The effects of dipole eccentricity, size of finite elements and local mesh refinement on solution accuracy are also addressed in the present simulation study. PMID- 15285260 TI - Physical mechanisms of gas and perfluoron retinopexy and sub-retinal fluid displacement. AB - Injection of gas into the eye, followed by face-down positioning, is a common protocol for the reseating of the retina in posterior and superior retinal tears and breaks. The physical mechanism by which injected gas helps reattach retinal flaps is often ascribed to the 'buoyancy' force of the injected gas bubble. The various forces at play in this system (surface tension and buoyancy) were calculated and compared. The results are extended to the case in which the retina is intact (pneumatic displacement of blood) and to the use of intraocular perfluoron (n-perfluorooctane). We show that buoyancy forces are applicable only for gas or n-perfluorooctane bubbles that are smaller than the detached retina and that do not invade underneath the retina. For larger bubbles, as is normally used in reattachment protocols, we show that it is the interfacial tension that reattaches the retina. The range of angles within which patients can position, and still maintain a gas-vitreous interface along a tear is calculated as a function of the volume of injected gas and size of the tear. The maximum retinal flap size that can be reattached using surface tension forces is also estimated. PMID- 15285261 TI - An evaluation of safety guidelines to restrict exposure to stray radiofrequency radiation from short-wave diathermy units. AB - Short-wave diathermy (SWD), a form of radiofrequency radiation used therapeutically by physiotherapists, may be applied in continuous (CSWD) or pulsed (PSWD) mode using either capacitive or inductive methods. Stray radiation emitted by these units may exceed exposure guidelines close to the equipment. Discrepant guidelines exist on a safe distance from an operating unit for operators and other personnel. Stray electric (E-field) and magnetic (H-field) field strengths from 10 SWD units in six departments were examined using a PMM 8053 meter and two isotropic probes (EP-330, HP-032). A 5 l saline phantom completed the patient circuit. Measurements were recorded in eight directions between 0.5 m and 2 m at hip and eye levels while the units operated at maximum output and data compared to current guidelines. Results found stray fields from capacitive CSWD fell below operator limits at 2 m (E-field 4.8-39.8 V/m; H-field 0.015-0.072 A/m) and at 1 m for inductive CSWD (E-field 0-36 V/m; H-field 0.01 0.065 A/m). Capacitive PSWD fields fell below the limits at 1.5 m (E-field 1.2 19.9 V/m; H-field 0.002-0.045 A/m) and at 1m for inductive PSWD (E-field 0.74.0 V/m; H-field 0.009-0.03 A/m). An extra 0.5 m was required before fields fell below the guidelines for other personnel. These results demonstrate, under a worst case scenario, emissions from SWD exceed the guidelines for operators at distances currently recommended as safe. Future guidelines should include recommendations for personnel other than physiotherapists. PMID- 15285262 TI - Photon transport parameters of diffusive media with highly anisotropic scattering. AB - The dependence of the photon transport parameters on the optical characteristics of diffusive media such as biological tissue with strongly forward biased scattering is examined with respect to the influence of the large angle scattering component and higher moments of the phase function. The latter are particularly significant for the temporal evolution of the angular intensity. The P3 approximation gives clear physical insight into the influence of boundaries on the radiative flux and is applied here as an analytic method of evaluating certain phase functions reported in the literature, while higher order P(N) approximations are used to calculate accurate time-dependent angular intensity distributions of the scattered light. PMID- 15285263 TI - A design study for a 'spiral staircase' ionization chamber for the quality control of electron beams. AB - In order to verify that the energies of electron beams used for external beam therapy remain constant, IPEM 81 recommends a constancy check based on the ratio of ionization chamber measurements at two depths along the central axis. Such measurements for a range of electron energies can be a time consuming process. The purpose of this study was to design a device that would use several ion chambers simultaneously to measure electron depth dose curves, and hence the electron energy. A design was developed for a device consisting of ten independent ionization chambers, shaped and arranged in a solid phantom like the steps of a spiral staircase, the axis of the staircase being coincident with the axis of the electron beam. Measurements were carried out to test the design of individual chambers and to optimize the radius of the spiral and both the depth intervals and the lateral spacing between adjacent chambers. For ranges of electron energy from 6-12 MeV and 12-20 MeV the radii of the spirals needed were found to be 36.5 mm and 30.9 mm, the angular separations between edges of the chambers were 52 degrees and 30 degrees and chamber depths were found to be 10, 15, 20, 25, 30, 35, 40, 45, 50, 55 mm and 20, 40, 45, 50, 55, 60, 65, 70, 75, 80 mm, respectively. PMID- 15285264 TI - Effects of temperature variation on MOSFET dosimetry. AB - This note investigates temperature effects on dosimetry using a metal oxide semiconductor field effect transistor (MOSFET) for radiotherapy x-ray treatment. This was performed by analysing the dose response and threshold voltage outputs for MOSFET dosimeters as a function of ambient temperature. Results have shown that the clinical semiconductor dosimetry system (CSDS) MOSFET provides stable dose measurements with temperatures varying from 15 degrees C up to 40 degrees C. Thus standard irradiations performed at room temperature can be directly compared to in vivo dose assessments performed at near body temperature without a temperature correction function. The MOSFET dosimeter threshold voltage varies with temperature and this level is dependent on the dose history of the MOSFET dosimeter. However, the variation can be accounted for in the measurement method. For accurate dosimetry, the detector should be placed for approximately 60 s on a patient to allow thermal equilibrium before measurements are taken with the final reading performed whilst still attached to the patient or conversely left for approximately 120 s after removal from the patient if initial readout was measured at room temperature to allow temperature equilibrium to be established. PMID- 15285265 TI - Surface dose extrapolation measurements with radiographic film. AB - Assessment of surface dose delivered from radiotherapy x-ray beams for optimal results should be performed both inside and outside the prescribed treatment fields. An extrapolation technique can be used with radiographic film to perform surface dose assessment for open field high energy x-ray beams. This can produce an accurate two-dimensional map of surface dose if required. Results have shown that the surface percentage dose can be estimated within +/-3% of parallel plate ionization chamber results with radiographic film using a series of film layers to produce an extrapolated result. Extrapolated percentage dose assessment for 10 cm, 20 cm and 30 cm square fields was estimated to be 15% +/- 2%, 29% +/- 3% and 38% +/- 3% at the central axis and relatively uniform across the treatment field. The corresponding parallel plate ionization chamber measurements are 16%, 27% and 37%, respectively. Surface doses are also measured outside the treatment field which are mainly due to scattered electron contamination. To achieve this result, film calibration curves must be irradiated to similar x-ray field sizes as the experimental film to minimize quantitative variations in film optical density caused by varying x-ray spectrum with field size. PMID- 15285266 TI - Uniaxial and biaxial irradiation protocols for microbeam radiation therapy. AB - Synchrotron-generated x-ray beams for microbeam radiation therapy (MRT) are fixed in space, so three-dimensional treatment planning would require that a patient be secured to, and moved in a gantry between exposures. Two protocols for such movements are proposed: one for uniaxial opposing-fields cross-planar MRT, the other for biaxial orthogonal-fields co-planar MRT. PMID- 15285267 TI - Nutritional therapy: an important component of integrative medicine. PMID- 15285268 TI - Retroactive prayer: a preposterous hypothesis. PMID- 15285269 TI - Yoga research and Richard Freeman. PMID- 15285270 TI - A brief evidence-based review of two gastrointestinal illnesses: irritable bowel and leaky gut syndrome. PMID- 15285271 TI - Food as healer, food as slayer. PMID- 15285272 TI - An overview of the pathophysiology of neurodegenerative disorders. PMID- 15285273 TI - An analytic review of studies on measuring effects of external QI in China. AB - Scientists have long been interested in measuring external qi (EQ or wai qi) during qigong healing, and have produced a large body of literature over the past 20 years. This paper reviews the major research on measuring EQ in China and tries to help other researchers to get a picture on what has been done so as to eliminate the simple replication of already verified results. Starting with the historical background of EQ studies in China, this paper analytically reviews the major studies of EQ effects from five different categories of detectors: 1) physical signal detectors; 2) chemical dynamics methods; 3) detectors using biological materials; 4) detectors using life sensors; and 5) detectors using the human body. The focus is on the pros and cons of each detector. These studies documented some important correlates of EQ process or qi healing, which cannot be explained by psychological effect or the known biological processes. Even though the extant literature suggests that intent plays a critical role in the effect or characteristics of EQ we know little about its role in EQ effect and its relationship with qigong healing from these experiments. These studies have confirmed the existence of measurable EQ effects from various perspectives; however, none has really revealed the primary nature of EQ or how EQ healing works. Given the fact that qigong therapy is based on the dialectic view of two interdependent spheres, while modern science and medicine is based on the reductionist view of one material world, it is recommended that future studies should use more biological or life-sensor detectors to increase our understanding of the healing potentials of qigong, instead of stay at the level of verification of signals. New methodologies, new theories, and new perspectives are urgently needed for further understanding what qigong is and how EQ healing works. PMID- 15285274 TI - Dangshen (Codonopsis pilosula) and Bai guo (Gingko biloba) enhance learning and memory. AB - CONTEXT: An exploration of the usefulness of several common Chinese herbs used for Acquisition and Retention singly or in combination is required. OBJECTIVE: To test (1) whether Gingko biloba (Bai guo) in combination with Codonopsis pilosula (Dangshen) or Gingko biloba (GB) alone could enhance memory acquisition and retention of normal human subjects, better than a placebo and (2) to investigate whether the overall health status can be affected by these supplements. DESIGN: A double blind randomized placebo controlled trial design was used to determine the efficacy of these two products compared to placebo. PARTICIPANTS AND SETTING: Sixty participants, aged 21- 60 years, who were either students or faculty of the Southern California University of Health Sciences. INTERVENTION: Each combination capsule was made up of 75mg of Codonopsis pilosula total glycosides and 40mg GB extract. Each GB capsule consisted of 40mg of Gingko biloba as an aqueous extract and. The placebo pill was similar in shape and color to that of the other two capsules. All the participants regardless of the group were instructed to take one pill twice a day with food. OUTCOME MEASURES: The primary outcome measure was a computerized, standardized acquisition and retention test provided by Digital Acumen, Inc. The secondary outcome measures were the SF-12 and the Medical Symptom Questionnaire ( 1997 healthcomm International, Inc. And Immuno Laboratories, Inc.) RESULTS: Mean age was 28 years; almost two thirds of the sample were males. The study adopted repeated measure analysis using data from three measurement points for between group analysis. GB group was compared to placebo, and CPG/GB group was also compared to placebo. These individual comparisons were made to determine whether the active products produced better results than chance. The between groups findings indicate that both products are better than placebo at producing improvements in acquisition and retention and overall health status. CONCLUSION: In our participant population, the combination product seems to be better than GB alone in improving the cognitive function and overall health. PMID- 15285275 TI - Symptom relief and adherence in the rotary diversified diet, a treatment for environmental illness. AB - CONTEXT: The rotary diversified diet, which involves food elimination and rotation of remaining allowed foods, is commonly used in the management of environmental illness. No studies have considered patient adherence while evaluating the effectiveness of the diet in controlling symptoms. OBJECTIVE: The study examined the severity of patients' perceived symptoms and dietary adherence during treatment with a rotary diversified diet. DESIGN: A prospective and exploratory study using purposive sampling and the following data collection methods: personal interviews, symptom severity questionnaires, and food records to assess dietary adherence. SETTING: Private clinic of a Toronto, Ontario physician specializing in environmental medicine. PATIENTS OR OTHER PARTICIPANTS: Twenty-five female residents of Toronto, Ontario (aged 25-67 years) diagnosed with environmental illness. INTERVENTION: Patients were treated with a rotary diversified diet for 16 weeks. MAIN OUTCOME MEASURES: Symptom severity and dietary adherence were assessed after 4, 10, and 16 weeks of treatment. Adherence was assessed by comparing food records to the diet prescription. RESULTS: At 16 weeks, patients reported a 50% decline in symptom severity for 5 of the 6 symptom categories assessed and for all categories combined. Those with closer elimination and rotation adherence reported a greater decline in gastrointestinal symptoms at 4 and 10 weeks of treatment, respectively. Improvement in total symptom severity was associated with closer rotation adherence at 10 weeks. Patients experienced difficulties in adhering to the diet. CONCLUSIONS: Results suggest that the diet, if followed, is beneficial, especially in improving gastrointestinal symptoms. Further evaluation of its effectiveness is limited by its complexity and the nature of environmental illness. Because the diet is difficult to follow over time, patients require extensive nutritional counseling and support. PMID- 15285276 TI - A pilot study of mind-body changes in adults with asthma who practice mental imagery. AB - CONTEXT: Despite the growing number of studies of imagery and the use of complementary and alternative modalities as treatments for asthma, research on mental imagery in adults with asthma is practically, nonexistent. The purpose of this feasibility study was to lay groundwork for a larger follow-up clinical trial. OBJECTIVE: To determine whether pulmonary function, asthma symptoms, quality of life, depression, anxiety, and power differ over time in adults with asthma who do and do not practice mental imagery (MI). (Power is the ability to make aware choices with the intention of freely involving oneself in creating desired change.) DESIGN: Randomized controlled study using univariate repeated measures analysis of variance (ANOVA) and replacement through block design. SETTING: Lenox Hill Hospital, an affiliate of New York University Medical School, New York, NY. SUBJECTS: Sixty-eight adults with symptomatic asthma, after 4 weeks of baseline data collection and analysis, met requirements for this randomized controlled study. Thirty-three completed pulmonary function as well as self report tests at 4 time points over 17 weeks. The 16 experimental participants also completed the 4-session imagery protocol. INTERVENTION: Individual imagery instruction (week 1) and follow-up (weeks 4, 9, 15). Participants were given 7 imagery exercises to select from and practice 3 times a day for a total of 15 minutes. MAIN OUTCOME MEASURES: 1) Spirometry (FEV1); 2) medication use; 3) Asthma Quality of Life Questionnaire; 4) Beck Depression Inventory; 5) Spielberger Anxiety Scales (A-State and A-Trait); 6) Barrett Power as Knowing Participation in Change Tool, Version II; 7) Epstein Balloon Test of Ability to Image. RESULTS: There was little evidence of statistical change in this feasibility study; yet, valuable lessons were learned. Paired t-tests indicated there was a significant difference in the total power scores in the imagery group, and in the expected direction (two-tailed, t-statistic = -2.3, P = 0.035) and the choices sub-scale (two-tailed, tstatistic = -2.93, P = 0.01) of the power instrument from weeks one to 16 of the study. Eight of 17 (47%) participants in the MI group reduced or discontinued their medications. Three of 16 (19%) participants in the control group reduced their medications; none discontinued. Chi-square indicated differences between groups (X2 = 4.66, P = 0.05). Persons who reduced or discontinued their medications showed neither an increase in pulmonary function prior to medication discontinuation, nor a fall in these parameters following discontinuation. CONCLUSIONS: Findings related to major outcome measures must be viewed with caution due to the small sample size resulting from attrition related to labor intensiveness and, therefore, low statistical power. However, the study did provide significant data to plan a larger scale study of the use of mental imagery with adult asthmatics. The study also demonstrated that imagery is inexpensive, safe and, with training, can be used as an adjunct therapy by patients themselves. Its efficacy needs additional exploration. Further research for adults with asthma who practice imagery is important, as current treatments are not entirely efficacious. Lessons learned in this study may facilitate improvement in research designs. PMID- 15285277 TI - The effect of osteopathic manipulative treatment on immune response to the influenza vaccine in nursing homes residents: a pilot study. PMID- 15285279 TI - Mark Hyman, MD practicing medicine for the future. PMID- 15285278 TI - Cochrane for CAM providers: evidence for action. PMID- 15285280 TI - Welfare implications of new and existing breeding technologies. PMID- 15285281 TI - The interferon-gamma field trial: background, principles and progress. AB - Since November 2002, the State Veterinary Service and the Veterinary Laboratories Agency have been conducting a field trial to assess the interferon-gamma test as an ancillary parallel test to the tuberculin skin test in herds with confirmed tuberculosis breakdowns. Here, Martin Vordermeier, Tony Goodchild, Richard Clifton-Hadley and Ricardo de la Rua describe the background to the trial, discuss the principle of the test and provide an update of progress so far PMID- 15285282 TI - Disposal and disease rates in British dairy herds between April 1998 and March 2002. AB - Data derived over four years from 434 dairy herds in 1998/99 to 244 in 2001/02 revealed average disposal rates of 22.6 per cent per year, half of which were for poor fertility, mastitis and lameness. The quartile of herds with the lowest disposal rates sold an average of 11.5 per cent annually and the quartile with the highest rates sold 35.5 per cent. Average annual disease rates over the four years were as follows: for assisted calving 7.8 per cent, for digestive disease 1.2 per cent, for ketosis 0.5 per cent, for hypomagnesaemia 0.5 per cent, for hypocalcaemia 5.0 per cent and for injuries 0.8 per cent. The incidence of mastitis increased from 36.0 to 43.3 per cent of cows per year. The incidence of lameness decreased from 23.3 per cent in 1998/99 to 20.7 per cent in 2000/01 but increased to 21.9 per cent in 2001/02. Data received from the same 219 farms during the first three years showed no effective differences from the full set of data for each of the three years. The lowest annual incidences of mastitis and lameness on individual farms were below 7 per cent and 2.5 per cent, respectively. In general, housing cows in cubicles was associated with a greater risk of lameness, and housing them in straw yards with a greater risk of mastitis. However, some of the lowest rates of lameness were recorded in cubicle housed cows and some of the lowest rates of mastitis were recorded in cows housed in yards. Larger herds were not associated, in general, with higher rates of mastitis. PMID- 15285283 TI - Mortality due to fox predation in free-range poultry flocks in Britain. AB - Information derived from questionnaires sent to producers of free-range eggs, chickens, turkeys and geese was used to assess the extent of fox predation in terms of the density of the fox population and farm management factors. The mean reported bird mortality was less than 2 per cent for all the producers, but there were marked differences between them. Egg producers reported losing many more birds to foxes than other types of producer (up to 1000 birds in a laying cycle). On average, egg and goose producers lost the highest proportions of their total flocks (0.5 per cent). The extent of predation was not associated either with large-scale estimations of the density of the fox population or with variations in the farms' habitat. Chicken predation was not linked to differences in types of housing or fencing. However, there was a positive association between losses due to other causes and chicken predation. The results suggest that changes in farm management would be the most cost-effective means of reducing fox predation, rather than greater fox control. PMID- 15285284 TI - Retinal dysplasia in wild otters (Lutra lutra). AB - Eyes from 88 otters found dead in south-west England between 1990 and 2000, were collected as part of a larger pathological study. Histopathological examination of 131 eyes revealed dysplastic changes such as rosetting and folding in the retinas of 26 of the otters. In the eyes of 42 of the otters there were postmortem and fixation-induced retinal detachment which complicated the differentiation of dysplastic from normal retina, but 11 eyes had folds which probably indicated a dysplastic pathology. The eyes of 18 of the otters had inflammatory or autolytic changes which precluded a definitive evaluation of their dysplastic status. Liver samples from 55 of the otters were analysed for a range of polychlorinated hydrocarbons and for vitamin A. The otters with dysplastic retinas had significantly lower concentrations of vitamin A and higher concentrations of dieldrin than the otters with normal retinas. PMID- 15285285 TI - Association between Mycoplasma hyopneumoniae at different respiratory sites and presence of histopathological lung lesions. PMID- 15285286 TI - Pulmonary chondroma in a dog. PMID- 15285287 TI - Tularaemia in a captive golden-headed lion tamarin (Leontopithecus chrysomelas) in Switzerland. PMID- 15285288 TI - Potentially novel segmental polioencephalomyelitis in weaner pigs. PMID- 15285289 TI - Acetonaemia and displaced abomasum. PMID- 15285290 TI - New IT system to support TB and brucellosis testing. PMID- 15285291 TI - Slaughter without prestunning. PMID- 15285292 TI - Diagnostic and therapeutic dilemmas in vulvar cancer. AB - Invasive vulvar cancer is rare. The etiology of vulvar cancer is incompletely understood. Human papilloma virus is known to be the major causal factor. The keystep in the diagnosis is punch biopsy with attention to unifocal or multifocal lesions and involvement of adjacent regions. The therapeutic procedure is based mostly on histomorphologic parameters e.g. unifocal/multifocal lesion, localisation, tumor typing/grading, depth of invasion; the goal of these morphologic and morphometric parameters is to reduce radicality in order to avoid postoperative morbidity without jeopardising the chances of cure. This individualization considers treatment options in both the vulvar and the groin. PMID- 15285293 TI - Pseudomyxoma peritonei usually originates from the appendix: a review of the evidence. AB - Pseudomyxoma peritonei (PMP) is a rare condition, said to be more common in females during the fourth or fifth decade of life with an incidence believed to be in the region of one per million per year. Although PMP has been reported as originating from many intra-abdominal organs, in the majority of cases an ovarian or appendix cystadenoma or cystadenocarcinoma has been implicated as the primary site. Our experience suggests that most cases arise from the appendix. We have reviewed the clinical and scientific evidence. In the four largest reported series of 393 patients, 181 (46%) were males. Immunohistochemistry techniques in women with both appendical and ovarian tumours favour an appendiceal primary in most cases. The distinction between "benign" adenomucinosis and mucinous adenocarcinoma is important in both treatment and prognosis. Experience suggests that there may well be a spectrum of disease and possibly an "adenoma carcinoma sequence". PMID- 15285294 TI - Carcinoma in situ and early breast carcinoma survey of the Portuguese Senology Society on treatment in Portugal and its evolution between 1985 and 2000. AB - By means of a questionnaire sent to Portuguese hospitals which diagnose and treat most female patients with breast cancer, it was intended to assess the situation regarding the treatment of carcinoma in situ and early breast cancer (T1 or T2, N0 or N1), as well as their evolution between 1985 and 2000. The hospital participation rate was 65% and a sample of 865 patients was collected, distributed by the years 1985, 1990, 1995 and 2000. It was observed that, in terms of surgery, there was an increase in conservative surgery, which was over 40% in 2000, as well as an increase in the average of excised axillary lymph nodes. Progress in the surgical approach was similar both in cancer centres and in large and university hospitals, when compared with the other surveyed hospitals. Also, no differences between these two hospital groups in disease-free survival and overall survival were found. Postoperative radiotherapy was employed in more than 90% of the patients submitted to conservative surgery and adjuvant chemotherapy was used in 39% of all the patients, while tamoxifen as adjuvant treatment was used in 58% of the patients. PMID- 15285295 TI - Establishment and characterization of two cell lines (HEC-155, HEC-180) derived from uterine papillary serous adenocarcinoma. AB - Uterine papillary serous adenocarcinoma (UPSC) is an uncommon histologic subtype of endometrial cancer that characteristically behaves aggressively with a poor prognosis. We established two novel cell lines derived from UPSC designated HEC 155 and HEC-180. Both cell lines have been growing steadily in monolayer cultures for over ten years. Overexpression of p53, Ki67 and p27 was detected in both cell lines by immunohistochemistry. Using a DNA sequencing technique, a point mutation of p53 was detected in exon 8, codon 286 in HEC-155 and in exon 6, codon 195 in HEC-180. These newly established cell lines should be useful for investigating the characteristics of UPSC. PMID- 15285296 TI - Flow cytometric analysis of DNA ploidy and S-phase fraction of Stage IIIB cervical carcinoma. AB - OBJECTIVES: To evaluate the role of flow cytometry-measured DNA ploidy and S phase fraction as survival prognostic indicators in women with FIGO Stage IIIB squamous cell carcinoma of cervix. METHODS: We retrospectively reviewed the medical and pathological records of women with Stage IIIB squamous cell cervical carcinoma treated between 1993 and 1996. Flow cytometric analysis of DNA ploidy and S-phase fraction was performed by the modified Hedley technique using paraffin-embedded tissue. Survival was calculated using the Kaplan-Meier life table analysis. RESULTS: Of the 75 cases, 66 were analyzable. Diploid tumors were found in 73%. The mean S-phase fraction was 14% (SD = 5.4). The overall 5-year survival rate was 60%. The survival of patients with aneuploidy tumors was significantly worse than that of the diploid tumors (p = 0.001). The survival of the patients who had S-phase fraction > 12% was significantly worse than those who had S-phase fraction < or =12% (p = 0.04). CONCLUSIONS: In this homogeneous study population, we found that aneuploidy and S-phase fraction >12% correlated with poor survival. Identifying this poor prognostic group would be of benefit in considering additional treatment for a better outcome. PMID- 15285297 TI - Clinical review of 63 cases of sex cord stromal tumors. AB - PURPOSE OF INVESTIGATION: A retrospective analysis of 63 cases of sex cord stromal tumors treated in a 22-year period to evaluate the prognostic impact of different clinical parameters. METHODS: Sixty-three cases of sex cord stromal tumors were studied. These neoplasms are characteristically detected at an early stage and may recur locally years after the initial diagnosis. The most frequent cell type was adult granulosa cell tumor (75%); a total of 37 patients (62%) had Stage IA lesions. RESULTS: The cornerstone of treatment is surgery. Conservative surgical treatment was performed in 11 out of 47 cases (23%) of early stage tumor and in one of 13 patients affected by advanced neoplasm. Five of these 12 patients became pregnant after the treatment. Endometrial hyperplasia and uterine adenocarcinoma were diagnosed in 26.5% and 8.8% of the cases, respectively. Twenty-one patients (35%) received adjuvant therapy: 20 chemotherapy and one chemo-radiation treatment. Eight patients (13%) either progressed or recurred. All the recurrent patients but one had been treated with adjuvant chemotherapy (VAC and/or PVB). Overall survival by stage was 88.2% for Stage I and 30% for Stage III-IV. CONCLUSION: Tumor stage is the most important clinical parameter of prognostic relevance. Tumor size and laterality significantly affected prognosis in terms of overall survival; survival rate did not seem to be affected either by the age of the patients or by the modality of surgical treatment. PMID- 15285298 TI - Laparoscopic treatment of endometrial cancer: five-year recurrence and survival rates. AB - PURPOSE: To evaluate the 5-year recurrence and survival of patients with clinical Stage I endometrial cancer treated by the laparoscopic approach. METHODS: Retrospective review of 56 patients with clinical Stage I endometrial cancer treated laparoscopically. The mean follow-up was 6.4 (4.8-9.6) years. The International Federation of Obstetricians and Gynecologists (FIGO) surgical staging was: I, 45 (80.4%); II, three (5.4%); III, six (10.7%); and IV, two (3.6%). RESULTS: For patients with surgical Stage I (n = 45), the 5-year recurrence rate was 4.9% and the 5-year cause-specific survival was 94.7%. Factors univariately associated with survival were grade (p = .017), depth of myometrial invasion (p = .018), node metastasis (p = .013), and surgical stage according to FIGO (p = .097). CONCLUSION: The laparoscopic approach provided 5 year survival and recurrence rates similar to those previously attained by laparotomy in our institution. PMID- 15285300 TI - Does improving communication and information for women increase attendance at colposcopy in an inner city clinic? A randomised controlled trial. AB - PURPOSE: To establish whether information leaflets and appointment reminders improve attendance for diagnostic colposcopy. DESIGN & SETTING: Randomised controlled trial in an inner city colposcopy clinic. PARTICIPANTS: 500 women newly referred to the colposcopy clinic with abnormal cervical screening smear results were randomly assigned to an intervention or control group using computer generated numbers. INTERVENTION: 233 women referred for colpoposcopy were sent a comprehensive information leaflet with their appointment details and additionally were sent reminder letters regarding their appointment 7-10 days prior to their appointment date. CONTROL: 267 women were sent the standard basic information prior to their appointment. MAIN OUTCOME MEASURE: Attendance and default rates for diagnostic colposcopy. RESULTS: Default in the intervention arm was 42 out of 233 (18%) compared with 93 out of 267 (35%) in the control arm. CONCLUSION: Improved communication and information in the form of a detailed leaflet and a reminder letter for women with a recently abnormal smear result increased attendance for initial colposcopy assessment. PMID- 15285299 TI - Clinical value of the ultrasound Doppler index in determination of ovarian tumor malignancy. AB - PURPOSE OF INVESTIGATION: Clinical evaluation of the newly created Doppler index (DS) in preoperative early differentiation of ovarian tumor malignancy. PATIENTS AND METHODS: The study involved data concerning 464 ovarian tumor examinations of women treated between 1994 and 2002. One hundred and sixty-five woman had malignant ovarian tumors, and the other 299 tumors were benign on histopathology. Menopausal status of the studied women was also considered. Created ROC curves compare the diagnostic value of the suggested index. The number of vessels, their location and arrangement, shape of velocity waves as well as presence of the protodiastolic notch in the arterial vessels of the tumor were analyzed. Each of the mentioned features could acquire a point-value of 0 or 1. RESULTS: A cut-off point for the suggested Doppler index was determined at the level of 4 for the following prognostic values: sensitivity, specificity and accuracy which amounted to 86.7%; 93.3% and 90.9%, respectively. In the group of 101 postmenopausal women accuracy was higher--94.1%, specificity and positive predictive value (PPV) was estimated as 100%. CONCLUSIONS: The suggested Doppler index enables a precise prognosis of ovarian tumor malignancy and as a half-amount analysis helps in the decision making of the most effective therapeutic treatment. PMID- 15285301 TI - Gemcitabine in heavily pretreated patients with recurrent ovarian, peritoneal and fallopian tube carcinoma. AB - PURPOSE OF INVESTIGATION: To report the experience of a single institution in the south of Israel with gemcitabine in heavily pretreated patients with platinum resistant recurrent ovarian, peritoneal and fallopian tube carcinoma. METHODS: The hospital records of 21 patients with ovarian, peritoneal and fallopian tube carcinoma who had salvage chemotherapy with gemcitabine between October 1998 and November 2003 were retrospectively reviewed. Gemcitabine, 1000 mg/m2, was given on days 1, 8, and 15 of every 28 days. Dose intensity and relative dose intensity of gemcitabine were calculated. Response was determined using clinical evaluation, radiological reports and CA-125 level. Toxicity was graded using the National Cancer Institute (NCI) criteria. RESULTS: The median relative dose intensity of gemcitabine received by the patients was 0.91, with 17 (81%) patients receiving more than 80% of the planned standard dose intensity. Two (9.5%) patients had complete response of disease lasting for ten and 33 months, respectively, eight (38.1%) had stable disease and 11 (52.4%) had progressive disease. Three (14.3%) patients had CA-125 complete response, five (23.8%) had CA 125 partial response, six (28.5%) had CA-125 stable levels and seven (33.3%) had CA-125 progressive levels. Toxicity was mainly hematological with grade 3-4 toxicity as follows: leukopenia--two (9.5%) patients, neutropenia--four (19%), thrombocytopenia--three (14.3%) and anemia--one (4.7%). CONCLUSION: Gemcitabine has some activity and low and well tolerated toxicity in heavily pretreated patients with platinum-resistant recurrent ovarian, peritoneal and fallopian tube carcinoma. PMID- 15285302 TI - Laparoscopic coagulation of the uterine blood supply in laparoscopic-assisted vaginal hysterectomy is associated with less blood loss. AB - BACKGROUND: Does laparoscopic coagulation of the uterine blood supply decrease blood loss compared with transvaginal ligature of the uterine vessels? METHODS: Intra- and postoperative data of 446 patients undergoing laparoscopic-assisted vaginal hysterectomy at the Department of Gynecology, University of Jena, between 1998 and 2001 were analysed. In 213 patients the uterine blood supply was transected laparoscopically at the origin of the uterine vessels (LAVH type II) and in 233 patients (LAVH type I) transvaginally. RESULTS: Patients in both groups were comparable with respect to median age, Quetelet index, and parity. The drop of hemoglobin between the preoperative day and postoperative day 3 was 0.8 mmol/l or 0.6 mmol/l for LAVH type I without or with BSO vs 0.3 mmol/l or 0.4 mmol/l for LAVH type II without or with BSO (p = 0.001), respectively. Median operative time was similar for both techniques: LAVH type I 136 min or with BSO 128 min vs LAVH type II 126 min or with BSO 131 min. The weight of the removed uteri was significantly lower in LAVH type I vs type II (220 vs 270 grams), but similar when LAVH was combined with BSO (160 vs 178 grams). The rate of intraoperative complications was 2.2% vs 0.9% between LAVH type I or II (n.s.), but 9% vs 3.3% for overall postoperative complications (p = 0.01). CONCLUSIONS: Laparoscopic coagulation of the uterine blood supply at the origin of uterine vessels is a safe technique which minimizes blood loss in LAVH. In patients with a low preoperative hemoglobin value this technique is indicated. PMID- 15285303 TI - Estrogen receptor beta (ERbeta) expression in breast carcinomas is not correlated with estrogen receptor alpha (ERalpha) and prognosis: the Greek experience. AB - Estrogen receptors (ER) are members of the nuclear receptor superfamily of ligand activated transcription factors and mediate the effects of estrogen on target tissues. ERalpha was the first estrogen receptor to be characterized, and ERbeta was identified ten years later. The role of ERbeta in breast cancer pathobiology is largely unknown because specific antibodies have not been available until recently. The purpose of this study was to explore the expression of ERbeta in breast neoplasms and to correlate it with ERalpha and prognosis. ERa and ERbeta expression was monitored immunohistochemically in 59 breast carcinomas. We found no correlation between ERalpha and ERbeta expression, between ERbeta expression and the known prognostic indicators such as tumor size, grade or lymph node status, or between ERbeta expression and survival. Our findings contribute to the better understanding of the role of ERbeta in breast cancer. PMID- 15285304 TI - Reporting of "LSIL with ASC-H" on cervicovaginal smears: is it a valid category to predict cases with HSIL follow-up? AB - Recently it has been shown that there is a 15-30% risk of associated cervical intraepithelial neoplasia (CIN 2-3 or greater) for a low-grade squamous intraepithelial lesion (LSIL) diagnosis. We tried to define a subgroup of "LSIL with atypical squamous cells of undetermined significance. High-grade squamous intraepithelial lesion (LASC-H)" in cervicovaginal screening which may aid in predicting the cases associated with high risk cannot be ruled out. In the years between 2001 and 2003 a total of 21,342 cervicovaginal smears were evaluated. The smears with pure LSIL and LASC-H diagnosis which had histologic follow-up were selected. The cases with diagnosis of LASC-H contained numerous typical cells of LSIL and only a few cells with features suggesting high-grade squamous intraepithelial lesion (HSIL). Eight (61%) of 13 cases with a diagnosis of LASC-H but three (11%) of 27 cases with a diagnosis of pure LSIL resulted in CIN 2-3 histology (p < 0.05). Diagnosis of LASC-H may be a valid diagnostic category in distinguishing patients with LSIL that would have HSIL in follow-up. PMID- 15285305 TI - Expression of beta-human chorionic gonadotropin in ovarian cancer tissue. AB - The purpose of the present investigation was to determinate expression of human chorionic gonadotropin gene in ovarian cancer tissue. The study included 15 patients with epithelial ovarian carcinoma. The expression of mRNA hCGbeta was determinated by the RT PCR method and the distribution of the hormone in study tissue was analyzed immunohistochemicaly. In all 15 study specimens of the ovarian carcinoma tissue the active hCGbeta gene was found, whereas noncancerous tissue demonstrated lack of the hormone expression. Thus, the study clearly shows that the expression of hCGbeta is the feature of ovarian cancer tissue. PMID- 15285306 TI - The presence of hereditary BRCA1 gene mutations in women with familial breast or ovarian cancer and the frequency of occurrence of these tumours in their relatives. AB - In 48 women with familial breast cancer as well as in 22 women with familial ovarian cancer, the presence of pathogenic mutations in BRCA1 gene were found in 35.4% and 54.6% of patients, respectively. From the patients with mutations we created two groups: the CaM--probands with breast cancer and CaOv--probands with ovarian cancer. The probands with breast cancer were younger by a mean of five years than the probands with ovarian cancer (p = 0.048). METHODS: The PCR-SSCP procedure was used to find mutations in the BRCA1 gene. Fragments suspected of mutation were subjected to nucleotide sequencing. RESULTS: In the CaM group, which consisted of 17 women with breast cancer, the following mutations in the BRCA1 gene were detected: 5382insC, T300G, 3819del5 and IVS20+60ins12. The probands of the CaM group and their relatives developed a total of 49 breast and ovarian cancers. Among all these tumours the breast cancers of the probands made up 34.7%, the breast cancers of proband relatives made up 57.1% and the ovarian cancers of probands and their relatives made up only 8.2%. The CaOv group consisted of 12 probands with ovarian cancers in whom we detected only two kinds of mutations: 5382insC and 185delAG. The probands of the CaOv group and their relatives developed a total of 38 ovarian and breast cancers. Among all these tumours the ovarian cancers of the probands made up 31.6%, the ovarian cancers of their relatives made up 34.2% and the breast cancers of the relatives 34.2% of tumours. In the probands with breast or ovarian cancer the predominant mutation was 5382insC--in the BRCA1 gene detected in 76.5%, and 91.7%, respectively. Despite the predominant presence of the same mutation in probands from both groups the ratio of the number of breast cancers to the number of ovarian cancers in their relatives differed significantly (p = 0.0003). CONCLUSION: This data shows that the presence of the 5382insC mutation in the BRCA1 gene is not always associated with the development of ovarian cancer. It is very likely that the development of ovarian cancer requires some additional factor, which was common among the familial ovarian cancer patients, and was almost inexistent among the familial breast cancer patients. On the other hand, the development of ovarian cancer at a later age than breast cancer in probands suggests that some factors exist which slow down the development of ovarian cancer or which accelerate the development of breast cancer. PMID- 15285307 TI - A second Pap smear during colposcopy: is it really worth it? AB - The aim of this study was to determine whether the clinical value of a second Pap smear during colposcopy outweighed its cost-effectiveness and reliability parameters. We studied retrospectively 569 cases focusing on A) The initial Pap smear, B) The smear performed during colposcopy, C) The colposcopic findings, and D) The histopathological reports of the cases where biopsy sampling was performed. In 380 patients (67%), the second Pap smear corresponded to the first one. In 13% of the patients, the cytological lesions were worse (particularly in 2% of the patients staging increased from HPV-associated reactive cellular changes to CIN II, or from CIN I to CIN III), and in 20% slighter than the initial. In 79% of the cases revealing more serious lesions in the second smear, the histological result of the biopsy corresponded to that of the initial smear. Conclusively, only 2% seem to benefit from a second repeat Pap smear during referral colposcopy. PMID- 15285308 TI - The effect of recombinant GM-CSF on IL-6 and TNF-alpha levels in epithelial ovarian cancer patients who received paclitaxel and cisplatinum: preliminary results. AB - OBJECTIVE: We investigated the effects of GM-CSF factor on IL-6 and TNF-alpha levels prior to paclitaxel-cisplatinum combination chemotherapy for advanced epithelial ovarian cancer. MATERIALS AND METHODS: Twenty-three consecutive patients with FIGO (International Federation of Gynecology and Obstetrics) Stage III-IV epithelial ovarian cancer were enrolled in the study. Following cytoreductive surgery patients received 175 mg/m2 paclitaxel and 75 mg/m2 cisplatinum on the same day. These 23 patients also received RhuGM-CSF five days before at a dose of 5 microg/kg/day by subcutaneous injection for three days. IL 6 and TNF-alpha levels were measured before and 24 hours later following the last dose of RhuGM-CSF. RESULTS: White blood cell counts on the 10th day of the cycle were lower than preGM-CSF white blood cell counts and the difference was statistically significant (p = 0.003). Platelet levels on the 10th day of the chemotherapy cycle were lower than pre GM-CSF levels, however were not statistically significant (p = 0.097). Post GM-CSF TNF-alpha and IL-6 levels were higher than pre GM-CSF levels. This difference was statistically significant for TNF-alpha (p = 0.002) however for IL-6 a statistically significant difference was not detected (p = 0.55). GM-CSF does not significantly effect IL-6 levels in contrast to TNF-alpha. CONCLUSION: Clinical implications of increased levels of TNF-alpha are unclear and for a precise determination further studies are needed. PMID- 15285309 TI - Immunohistochemical profile of intravenous leiomyomatosis. AB - To determine the immunohistochemical staining profile of intravenous leiomyomatosis (IVL), we analysed six IVLs and 12 ordinary leiomyomas (LM) for immunoreactivity with a panel of 11 antibodies. All IVLs and LMs reacted with antibodies to alpha-smooth muscle actin (alphasm), h caldesmon, vimentin and progesterone receptor (PR). Five of six IVLs and all LMs reacted with desmin. All IVLs were negative for CD-10. Only one LM exhibited focal CD-10 positivity. Three of six IVLs and nine of 12 LMs showed estrogen receptor expression. All IVLs and LMs showed immunnegativity with MIB-1 and inhibin. There were not any significant differences between immunoreactivity patterns of IVL and LM for asm, desmin, h caldesmon, CD-10, MIB-1 and PR. We conclude that, although they appear to be useful markers in differentiating IVL from ESS and LMS, a larger study also including ESS and LMS would be necessary to confirm their validity. PMID- 15285310 TI - K-ras gene point mutations and p21ras immunostaining in human ovarian tumors. AB - It is well recognized that genetic alterations within oncogenes, tumor suppressor genes, DNA mismatch repair and excision repair genes contribute to tumorigenesis within the human ovary. This study was undertaken to screen for the existence of K-ras gene point mutations in paraffin-embedded slides randomly selected from benign and malignant ovarian tumors applying the PCR-RFLP technique. Expression of p21ras was also assessed in 30 primary ovarian adenocarcinomas immunohistochemically. K-ras codon 12 point mutations occurred in nine of 40 (22.5%) cases. They were not identified in two benign mucinous cystadenomas, but in one out of two (50%) mucinous tumors of LMP (low malignant potential), in five out of 30 (17%) ovarian adenocarcinomas, and in one case of adenocarcinoma metastatic to the ovary. K-ras activation was also detected in one out of four (25%) sex cord-stromal cell tumors (folliculoma), and in one dysgerminoma. None of these tumors exhibited K-ras codon 13 point mutations. Gene alterations were more frequently found in mucinous than in non-mucinous (30% vs 10%) tumors, although the difference did not reach significance (p > 0.05). The frequency of K ras point mutations was correlated neither with clinical nor with pathological variables of cancer. Cytoplasmic p21ras was expressed in all adenocarcinomas negative for K-ras point mutations, whereas one of five (20%) K-ras-positive tumors exhibited lack of immunoreactivity. In conclusion, these findings confirm the role of K-ras activation in mucinous ovarian tumors. p21ras expression is not necessarily associated with K-ras gene alterations in human ovarian adenocarcinomas. PMID- 15285311 TI - Stromomyomas of the uterus-- importance of total circumferential evaluation of the margin. AB - Four stromomyomas were extensively dissected to represent the entire circumference of the uterus on sequential histologic sections. In all cases the smooth muscle component was extensive, and irregular interdigitation of stromal neoplasia with a smooth muscle component made evaluation of the margin difficult. It was impossible to determine where the smooth muscle component of the neoplasm ended and where peritumoral normal myometrium began. This makes the detection of vascular invasion more important. At the end of a thorough evaluation of sections, large vessel invasion was found on the circumference of three stromomyomas in a particular foci of the margin. Extensive circumferential evaluation of the margin has been evaluated as a promising procedure to allow effective distinction of stromomyomas with focal angioinvasion. PMID- 15285312 TI - Mammographic changes during postmenopausal hormonal replacement therapy with tibolone. AB - PURPOSE OF INVESTIGATION: To evaluate mammographic changes in postmenopausal women receiving hormonal replacement therapy (HRT) with tibolone. METHODS: 83 postmenopausal women aged 45-62 received the same dose of tibolone for a period of six months to five years without interruption. The women were examined mammographically every six to 12 months during the observation period. RESULTS: There was a low incidence of breast tenderness. Also, we did not observe any significantly increased mammographic density or neoplasmatic disease of the breast. As far as dysplastic changes are concerned, no remarkable aggravation in the mammographic picture was noted. CONCLUSION: The new synthetic steroid tibolone, in contrast to conventional HRT, rarely causes breast pain. At least short-period tibolone therapy (less than 5 years) has good effects on climacteric disorders and does not cause breast changes (dysplasia or cancer). Our study is on-going. PMID- 15285313 TI - Fibroadenomas of the breast: is there any association with breast cancer? AB - PURPOSE: The experience of our Breast Unit in the diagnosis and treatment of fibroadenomas is presented in this retrospective study, focusing specifically in cancer development within these common benign tumors of young age. MATERIAL METHODS: 310 women with histologically verified breast fibroadenomas who had surgical management in our Unit over the last 14 years were included in the study. RESULTS: Most of them (n: 255) presented with palpable lesions and had triple assessment preoperatively, while the remaining 55 had nonpalpable lesions and underwent needle-wire localization biopsies. Nine cases had cancer development, usually in situ, within the fibroadenomas (2.9%). CONCLUSION: Coexistence of fibroadenomas and breast cancer is relatively rare, but it should not be ignored by breast surgeons, and patients should be properly informed. PMID- 15285314 TI - Postoperative patient-controlled analgesia with intravenous tramadol, intravenous fentanyl, epidural tramadol and epidural ropivacaine+fentanyl combination. AB - PURPOSE: The aim of this study was to compare the effects of IV tramadol, IV fentanyl, epidural tramadol, and an epidural ropivacaine+fentanyl combination in patient-controlled analgesia (PCA) after lower abdominal surgery. METHODS: Eighty adult patients undergoing lower abdominal surgery were randomly allocated to one of four groups to receive analgesics with PCA pumps. Patients in group I received IV tramadol, group II patients IV fentanyl, group III patients epidural tramadol, and group IV patients an epidural infusion of 0.125% ropivacaine + 2 microg ml( 1) fentanyl combination. Analgesic effectiveness and side-effects were assessed at 1, 2, 3, 4, 5, 6, 8, 12, 16, 20, and 24 hours after surgery. RESULTS: Adequate analgesia was achieved in all groups. The analgesia was highest in group IV (p < 0.05), and lowest in group III patients (p < 0.05). Eleven patients (55%) in group I and eight patients (40%) in group II suffered from nausea/vomiting. CONCLUSION: Although adequate pain relief was achieved with all regimens that were used in the study, intravenous tramadol and intravenous fentanyl are associated with a high incidence of nausea and vomiting. PMID- 15285315 TI - The value of epithelial membrane antigen overexpression in hyperplastic and malignant endometrium and its relationship with steroid hormone receptor expression. AB - The aim of this study was to evaluate the value of epithelial membrane antigen overexpression (EMA OE) in benign, hyperplastic and neoplastic endometrium and to analyze its association with estrogen and progesterone receptors (ER, PR) immunohistochemistry, tumor grade and myometrial invasion in patients with endometrial carcinoma (EC). The OE of EMA was analysed immunohistochemically in nine patients with benign endometrium (BE), in 18 patients with atypical complex endometrial hyperplasia (ACH) and in 29 patients with EC. EMA OE was present in 13 of 29 patients (44.8%) with EC, in two of 18 patients (11.1 %) with ACH, and in none of nine patients with BE (p < 0.05). EMA OE of endometrial carcinoma was statistically correlated with the International Federation of Gynecology and Obstetrics (FIGO) grade (G1 vs G2 and G3, p < 0.05) and depth of myometrial invasion (< 1/2 vs > 1/2, p < 0.05). EMA OE was significantly associated with PR negativity (p < 0.001). However it did not show any association with ER immunohistochemistry (p = 0.14). PR immunohistochemistry had significant correlations with FIGO grade (p < 0.001) and depth of myometrial invasion (p < 0.05) but ER loss showed a nearly significant association only with advanced FIGO grade (p = 0.054). In conclusion, EMA shows increased expression as the lesion progresses to malignancy and can also aid discrimination between hyperplastic and neoplastic states. The correlation of imunohistochemical findings with tumor grade and myometrial invasion could help in predicting behavior of the tumor and planning treatment in patients with endometrial carcinoma. PMID- 15285316 TI - Adenosarcoma of the ovary. A case report. AB - Adenosarcoma of the ovary is a rare condition. We report a case of a 32-year-old patient that has been treated in our Department. The diagnosis of ovarian adenosarcoma was carried out after laparoscopy with removal of an ovarian endometriotic cyst. Laparoscopic homolateral ovariectomy was then performed and conservative treatment was decided on considering the young age, low stage and low grade of the disease. Second-look laparoscopy, clinical evaluation and ultrasound were performed for four years of follow-up. No recurrence has been detected. Conservative treatment should be proposed in fertile age with low-grade ovarian adenosarcoma, but a strict follow-up is always necessary. PMID- 15285317 TI - A case of rhabdomyosarcoma of the vagina in an elderly woman. AB - Most rhabdomyosarcomas of the vagina (RMSV) occur in infants and children up to six years old. RMSV in elderly patients is extremely rare. We report a case of a 70-year-old woman with RMSV. She had received surgery for uterine endometrial cancer one year before and a vaginal polypoid tumor was noted during routine follow-up vaginal examination. She was referred to our department for radiation therapy following partial tumorectomy of the lesion. She was given three sessions of intra-vaginal radiation therapy, once a week with 6 Gy at 7.5 mm below the vaginal surface and external irradiation of 50 Gy to the pelvis. However, paraaortal lymph node metastasis developed during initial radiation therapy. Furthermore, multiple bone metastases appeared at the completion of the radiation therapy. Six months after initial treatment the patient died from progression of the disease. Autopsy demonstrated small residual tumor at the primary site as well as multiple systemic metastases. PMID- 15285318 TI - Massive ovarian edema: a case report. AB - Massive ovarian edema is considered a non-neoplastic lesion characterized by a tumor-like enlargement of one or occasionally both ovaries secondary to an accumulation of edema fluid within the stroma. It is an uncommon entity that usually leads to oophorectomy as an unnecessary treatment in children, adolescents and young women. A diagnostic wedge resection with subsequent frozen section is essential. A definitive diagnosis of massive ovarian edema can not be made on preoperative imaging. In this article a case of massive ovarian edema in a 15-year-old female thought to be a solid neoplasm in the preoperative period is described. PMID- 15285319 TI - Primary ovarian leiomyosarcoma. Proliferation rate and survival. AB - OBJECTIVE: A case of Stage IIA primary ovarian leiomyosarcoma (LMS) with an unfavorable outcome 24 months after total abdominal hysterectomy with bilateral salpingo-oophorectomy, despite chemotherapy treatment, is described. Eighteen months from surgery the patient showed peritoneal spreading with ascites, liver and lung metastases. The present study was aimed to compare tumor growth fraction with cell density, lesion stage and clinical course. METHODS: The surgical specimens were evaluated by histological, histochemical, and immunocytochemical methods. Under microscopy, mitotic index (MI) was estimated, as a ratio of mitotic figures per 1000 tumor cells. Immunohistology was also carried out to reveal some intermediate-type filamentous proteins, as histogenetic markers, and the MIB1 monoclonal antibody was used to assess the percent of MIB1-positive nuclei (MIB1 labeling index). RESULTS: The histologic findings and immunohistology of the assayed intermediate filamentous proteins substantiated a diagnosis of LMS, with associated coagulation necrosis and not rare mitotic figures. A dual tumor component was observed, representing both the pleomorphic and myxoid LMS-variants. On the basis of the quantitative findings, a MI of 10.9 and a MIB1-LI of 23.1 were calculated, on average. CONCLUSIONS: The proliferation indices in the described variant of ovarian LMS, denote a fast growing malignancy. They agree with the tumor stage at operation and the subsequent fatal outcome. PMID- 15285320 TI - Pelvic lipomatosis complicating ovarian cyst removal: a case report. AB - Pelvic lipomatosis consists of an abdominal capsulated mass containing lipidic tissue, generally with remarkable dimensions, responsible for urinary tract disturbances. Here we describe the first case to our knowledge of accidental intraoperative diagnosis associated to an ovarian cyst in absence of objective symptoms and signs. PMID- 15285321 TI - Aggressive angiomyxoma of the vagina: a case report and review of the literature. AB - Aggressive angiomyxoma (AA) is a rare mesenchymal tumor of the lower pelvis and genital region, characterized by local infiltration and frequent, even multiple recurrences. In the present paper a case of a small-sized AA of the vagina, in a 55-year-old woman is reported. We describe the histological appearance and the immunohistochemical phenotype of this tumor and discuss its differential diagnosis from other mesenchymal lesions occurring in the pelvic and genital region. Furthermore, we attempt to enlighten the possible mechanisms that govern the pathogenesis and the biological behavior of this "mysterious" neoplasm. PMID- 15285322 TI - Primary fallopian tube cancer--a ten year review. Clinicopathological study of 12 cases. AB - Primary fallopian tube cancer is the rarest of all gynecologic cancers, presenting as benign pelvic disease or more often as ovarian cancer and almost all cases are diagnosed at operation or autopsy. Primary adenocarcinoma is the most common histological type of primary tube cancer which has traditionally been managed and treated in the same manner as epithelial ovarian cancer. However, unlike ovarian cancer, fallopian tube cancer is not routinely suspected and treatment may be delayed and also seems to have a worst prognosis than ovarian cancer. We present a retrospective study involving 12 patients with primary fallopian tube cancer treated in our department. The clinicopathologic characteristics and treatment are reviewed. PMID- 15285323 TI - Total laparoscopic radical hysterectomy (type III) and pelvic lymphadenectomy. AB - A case of Stage IIA, G2 carcinoma of the cervix treated by total laparoscopic radical hysterectomy and pelvic lymphadenectomy is reported. To our knowledge, a total laparoscopic radical hysterectomy with laparoscopic lymphadenectomy has not been previously described in Italy. PMID- 15285324 TI - Sclerosing stromal tumor of the ovary associated with Meigs' syndrome: a case report. AB - Sclerosing stromal tumors of the ovary are distinct, but rare benign neoplasms. These tumors appear solid and are very vascular giving the impression of malignant tumors. They occur mostly in young women. Morphologically they have distinct characteristics which differentiate them from other stromal tumors. Benign ovarian tumors associated with Meigs' syndrome are rare. In this article a case of ovarian sclerosing stromal tumor associated with Meigs' syndrome in a 17 year-old women is described and the differential diagnosis is also discussed. PMID- 15285325 TI - Expression of estrogen receptors alpha and beta in two uterine mesenchymal tumors after prolonged tamoxifen therapy. Report of two cases. AB - INTRODUCTION: Tamoxifen therapy is associated with an increased risk of endometrial carcinoma, and possibly uterine sarcomas. Little is known about hormone receptor expression in mesenchymal tumors of the uterus after tamoxifen therapy. CASES: The cases of two patients with uterine mesenchymal tumors after prolonged tamoxifen therapy due to breast cancer are presented. The expression of estrogen receptors alpha (ERalpha) and beta (ERbeta) and progesterone receptors (PR) was studied immunohistochemically in both cases. Both tumors were negative for ERalpha and positive for ERbeta. In the first case the tumor was negative for PR, while in the second only 20% of nuclei were PR-positive. CONCLUSIONS: Consistent with previous studies, uterine mesenchymal tumors after tamoxifen therapy do not express ERalpha. The results of the present report provide for the first time evidence that tamoxifen might exert a stimulatory effect on the uterus, at least during tumor progression, through ERbeta but not through ERalpha. PMID- 15285330 TI - Solubilization and solid-state characterization of a poorly soluble 5-alpha reductase inhibitor. AB - GI197111X is a 5-alpha reductase inhibitor for the treatment of androgenetic alopecia. Equilibrium solubilities of GI197111X were determined in multiple solvents or cosolvents. A polymorph screen was conducted using suspension equilibration and solution recrystallization methods. Single crystals were grown from pyridine/water and crystal structure was determined using a Bruker SMART diffractometer. Crystal structure data were imported into Cerius2 to provide visualization of the crystal structure and calculation of the simulated X-ray powder diffraction (XRPD) pattern. The solubility of GI19711IX was low at 25 degrees C in all vehicles suitable for animal and human dosing. The solubility of 6.4 mg/mL in Capmul MCM made it the only choice for a soft gel dosage form for phase I/II. Solution recrystallization and suspension equilibration of GI197111X have produced only one crystal form. Crystal structure data: orthorhombic P2(1) 2(1) 2(1); a= 10.8960(6) A, b=11.5683(6) A, c=20.9019(11) A; unit cell volume 2634.65(24) A3; Z=4; calculated density= 1.248 g/cc. The molecule has seven chiral centers, and single-crystal analysis eliminated all possible stereo isomers except the expected conformation or its enantiomer. Hydrogen bonds occur from both carbonyl oxygens to an H-N group. Simulated vacuum-based crystal morphology (habit) calculated using the Bravais-Friedel-Donnay-Harker, Growth Morphology, and Hartman-Perdok modules in Cerius2 was a close match to the morphology observed by light microscopy. PMID- 15285336 TI - The effect of citric acid added to hydroxypropyl methylcellulose (HPMC) matrix tablets on the release profile of vinpocetine. AB - Vinpocetine is a pH-dependent experimental drug with a short half-life. The sustained-release matrix tablets of vinpocetine were prepared by direct compression using hydroxypropyl methylcellulose (HPMC) and different amounts of citric acid to set up a system bringing about gradual release of this drug. In order to investigate the influence of citric acid and the pH value of medium on the drug release from HPMC matrix tablets, an in vitro release test was carried out in either phosphate buffer pH 6.8 [0.5% sodium dodecyl sulfate (SDS)] for 12 hr or in 0.1 N HCl (0.5% SDS) (0-2 hr) and phosphate buffer pH 6.8 (0.5% SDS) (2 12 hr). Dissolution curves were described by the Peppas equation: M(t)/M(inf)=ktn, and the influence of citric acid on the dissolution mechanism was estimated according to the regression parameter-n and k values. The addition of citric acid and the pH value of medium could notably influence the dissolution behavior and mechanism of drug-release from matrices. Increasing the amounts of citric acid produced an increase in drug release rate, which showed a good linear relationship between contents of citric acid and drug accumulate release (%) in phosphate buffer pH 6.8 (0.5% SDS) (r>0.99). Moreover, a higher drug release rate could be found in 0.1 N HCl (0.5% SDS) than that in phosphate buffer pH 6.8 (0.5% SDS) during the first two hours when the content of citric acid added to matrices was lower than 45 mg/tab., but no significant difference could be found when the content of citric acid was above that value. Increasing amounts of citric acid produced decreasing values of n and increasing values of k, in a linear relationship, which indicated there was a trend favoring the mechanism of diffusion with the addition of increasing quantities of citric acid. PMID- 15285329 TI - Effect of formulation composition on the properties of controlled release tablets prepared by roller compaction. AB - This study discusses the effect of formulation composition on the physical characteristics and drug release behavior of controlled-release formulations made by roller compaction. The authors used mixture experimental design to study the effect of formulation components using diclofenac sodium as the model drug substance and varying relative amounts of microcrystalline cellulose (Avicel), hydroxypropyl methylcellulose (HPMC), and glyceryl behenate (Compritol). Dissolution studies revealed very little variability in drug release. The t70 values for the 13 formulations were found to vary between 260 and 550 min. A reduced cubic model was found to best fit the t70 data and gave an adjusted r square of 0.9406. Each of the linear terms, the interaction terms between Compritol and Avicel and between all three of the tested factors were found to be significant. The longest release times were observed for formulations having higher concentrations of HPMC or Compritol. Tablets with higher concentrations of Avicel showed reduced ability to retard the release of the drug from the tablet matrix. Crushing strength showed systematic dependence on the formulation factors and could be modeled using a reduced quadratic model. The crushing strength values were highest at high concentrations of Avicel, while tablets with a high level of Compritol showed the lowest values. A predicted optimum formulation was derived by a numerical, multiresponse optimization technique. The validity of the model for predicting physical attributes of the product was also verified by experiment. The observed responses from the calculated optimum formulation were in very close agreement with values predicted by the model. The utility of a mixture experimental design for selecting formulation components of a roller compacted product was demonstrated. These simple statistical tools can allow a formulator to rationally select levels of various components in a formulation, improve the quality of products, and develop more robust processes. PMID- 15285328 TI - Transdermal delivery of ketorolac tromethamine: effects of vehicles and penetration enhancers. AB - The effects of vehicles and penetration enhancers on the in vitro permeation of ketorolac tromethamine (KT) across excised hairless mouse skins were investigated. Among pure vehicles examined, propylene glycol monolaurate (PGML) showed the highest permeation flux, which was 94.3 +/- 17.3 microg/cm2/h. Even though propylene glycol monocaprylate (PGMC) alone did not show high permeation rate, the skin permeability of KT was markedly increased by the addition of diethylene glycol monoethyl ether (DGME); the enhancement factors were 19.0 and 17.1 at 20% and 40% of DGME, respectively. When DGME was added to PGML, the permeation fluxes were almost two times at 20-60% of DGME compared to PGML alone. In the cosolvent system consisting of propylene glycol (PG)-oleyl alcohol, the permeation rate increased as the ratio of PG increased. In the study to investigate the effect of drug concentration on the permeation rate of KT, the permeation rates increased as the drug concentration increased in all vehicles used, and the dramatic increase in permeation rate was obtained when the drug concentration was higher than its solubility. For the effects of fatty acids on the permeation of KT, five fatty acids were added to PG at concentrations of 1%-, 3%-, 5%- and 10%- caprylic acid, capric acid, lauric acid, oleic acid, and linoleic acid. The enhancing effects of fatty acids were different, depending on the concentration as well as the sort of fatty acids. The highest enhancing effect was attained with 10% caprylic acid in PG; the permeation flux was 113.6 +/- 17.5 microg/cm2/h. The lag time of KT was reduced as the concentration of fatty acids increased except for caprylic acid. PMID- 15285332 TI - In situ gelling pectin formulations for oral sustained delivery of paracetamol. AB - The purpose of this study was to evaluate the potential of a pectin formulation with in situ gelling properties for the oral sustained delivery of paracetamol (acetaminophen). The formulations consisted of dilute aqueous solutions (1% to 2% w/v) of low methoxy pectin containing calcium ions in complexed form, which on release in the acidic environment of the stomach caused gelation of the pectin. In vitro studies demonstrated diffusion-controlled release of paracetamol from the gels over a period of 6 h. A bioavailability of approximately 96% of that of a paracetamol solution could be achieved from gels containing an identical dose of drug formed in situ in the stomachs of rats, with appreciably lower peak plasma levels and a sustained release of drug over a period of at least 6 h. PMID- 15285335 TI - In vitro-in vivo characterization of release modifying agents for parenteral sustained-release ketorolac formulation. AB - One of the prerequisites for a parenteral preparation is that the excipients incorporated are biocompatible and biodegradable. In the present study hydrophilic and hydrophobic excipients were investigated for developing an intramuscular sustained-release formulation of ketorolac. Kollidon 17 PF, Peceol (glyceryl monooleate), and castor oil were chosen as the potential release retarding agents, each with a distinct mechanism of action. They were evaluated by in vitro drug-release profiles and in vivo pharmacodynamic and pharmacokinetic study in mice. Cumulative drug release was determined for standard and test formulations in modified Franz diffusion cell. Pharmacodynamic parameter, T = 70% response of peak analgesic response, was used to compare the performance of test formulations. Based on pharmacodynamic/pharmacokinetic correlation in the animal studies, Css(max) and Css(min) of 51.39 and 30.0 microg/mL, respectively, were determined and considered as performance markers for pharmacokinetic evaluation of test formulations. The study suggested that the sustained-release capability of glyceryl monooleate was maximum followed by that of castor oil and Kollidon 17 PF, when compared to conventional ketorolac tromethamine formulation. It was inferred that water soluble excipient, though, showed release retarding property in vitro but could not maintain it in the in vivo environment. Glyceryl monooloeate-based formulation produced the most favorable drug blood concentration vs. time profile. PMID- 15285331 TI - Comparison of the physical and chemical stability of niclosamide crystal forms in aqueous versus nonaqueous suspensions. AB - In an effort to produce physically stable and pharmaceutically acceptable suspensions of niclosamide, this study reports the differences in physical and chemical stability of aqueous vs. nonaqueous suspensions of a niclosamide anhydrate, two monohydrates HA and HB, a 1:1 niclosamide N,N-dimethylformamide solvate, a 1:1 niclosamide dimethyl sulfoxide solvate, a 1:1 niclosamide methanol solvate, and a 2:1 niclosamide tetraethylene glycol hemisolvate. Evaluation of aqueous and nonaqueous suspensions showed that in aqueous suspensions anhydrous, and solvated niclosamide crystal forms were transformed to a monohydrate, HA, which was reasonably stable but which did eventually transform to the most stable monohydrate HB. The order in which these crystal forms transformed to monohydrate HB were: Anhydrate > N,N-dimethylformamide > dimethyl sulfoxide > methanol > tetraethylene glycol > monohydrate HA. In a nonaqueous propylene glycol vehicle, the transformation to the monohydrous forms was not observed and on desolvation the solvated crystals transformed to the anhydrous form. In all cases, immediately upon desolvation or dehydration, the crystal structures of the desolvated materials were similar to that of the solvated materials. However, the isomorphic structures, formed after desolvation, were unstable and rehydrated or resolvated when exposed to the solvent or converted to the anhydrous form in a dry environment. The crystal forms remained chemically stable in both aqueous and nonaqueous suspensions for the length of the study. PMID- 15285327 TI - Acute renal failure after anterior pelvic exenteration: a case report and review of the literature. AB - This is a report of a case of advanced cervical carcinoma in a 34-year-old woman treated with anterior pelvic exenteration at the Department of Gynecology of the Medical University in Gdansk. Despite annual gynecological check-ups, the patient presented with profuse bleeding from the genital tract. IVa cervical carcinoma according to the International Federation of Gynecology and Obstetrics (FIGO) staging was diagnosed. A vesicovaginal fistula was confirmed. In the postoperative period acute renal failure occurred. Twenty-four days after the operation when normal renal parameters had been restored, the patient was transferred to the Department of Radiotherapy for supplementary treatment. Pelvic exenteration offers the last chance for some women with gynecological malignancy and can provide a good chance of long-term survival in carefully selected patients with gynecological cancer. PMID- 15285337 TI - In vitro and ex vivo permeation studies of chlorpheniramine maleate gels prepared by carbomer derivatives. AB - The antihistaminic chlorpheniramine maleate (CPM) is used for symptomatic relief of hypersensitivity reactions and in pruritic skin disorders. At present, the drug is marketed in tablet, capsule, syrup, cream, and injectable dosage forms. Chlorpheniramine maleate has some side effects when taken orally. Due to its first pass effect, only 25%-45% of the orally administered dose reaches the blood circulation. To bypass these disadvantages, we aimed to investigate percutaneous absorption of CPM from gel formulations prepared with different carbomer derivatives (Carbopol 934, 940, 941, 2984, 980, and 981; main differences are related to presence of a comonomer and cross-link density). Cellulose membrane was used as the diffusion barrier for all the formulations' drug-release studies. The release of active substance from carbopol derivatives, which have the least cross-linking density (Carbopol 941 and 981) was found to be numerically higher than the others. The formulation (F8; 1% Carbopol 941) that exhibited the maximum drug release through the cellulose membrane was further studied for drug release by using polyurethane membrane, excised rat skin, and human skin. The penetration of the active substance through different diffusion barriers was found to be statistically different (p<0.05) when compared. Of all the different diffusion barriers, rat skin gave the closest results to human skin. Thus topical application of CPM in the carbomer gel may be of potential use for local activity. The type and concentration of carbomers can affect drug release. The synthetic membranes are useful in assessments of formulations in quality assurance but they do not give definite indication of how a formulation will behave when it is used on skin. PMID- 15285334 TI - Evaluation of selected polysaccharide excipients in buccoadhesive tablets for sustained release of nicotine. AB - Some naturally occurring biocompatible materials were evaluated as mucoadhesive controlled release excipients for buccal drug delivery. A range of tablets were prepared containing 0-50% w/w xanthan gum, karaya gum, guar gum, and glycol chitosan and were tested for swelling, drug release, and mucoadhesion. Guar gum was a poor mucoadhesive and lacked sufficient physical integrity for buccal delivery. Karaya gum demonstrated superior adhesion to guar gum and was able to provide zero-order drug release, but concentrations greater than 50% w/w may be required to provide suitable sustained release. Xanthan gum showed strong adhesion to the mucosal membrane and the 50% w/w formulation produced zero-order drug release over 4 hours, about the normal time interval between daily meals. Glycol chitosan produced the strongest adhesion, but concentrations greater than 50% w/w are required to produce a nonerodible matrix that can control drug release for over 4 hours. Swelling properties of the tablets were found to be a valuable indicator of the ability of the material to produce sustained release. Swelling studies also gave an indication of the adhesion values of the gum material where adhesion was solely dependent upon penetration of the polymer chains into the mucus layer. PMID- 15285333 TI - In vitro and in vivo evaluation of glibenclamide in solid dispersion systems. AB - The purpose of this work is to improve the dissolution and bioavailability characteristics of glibenclamide as compared to Daonil tablets (Hoechst). Solid dispersions of glibenclamide in Gelucire 44/14 (Formula 1) and in polyethylene glycol 6000 (PEG 6000) (Formula 2) were prepared by fusion method. In vitro dissolution studies showed that the dispersing systems containing glibenclamide and Gelucire 44/ 14 or PEG 6000 gave faster dissolution rates than the reference product Daonil. The in vivo bioavailability study was assessed in six healthy male volunteers in crossover design with a 1-week washout period. Both formulas were found to be significantly different from Daonil with regard to the extent of absorption as indicated by the area under serum concentration-time curve. Both formulas are not significantly different from Daonil with respect to time of peak plasma concentration Tmax. It is concluded from this pilot study that the ranking of the in vitro dissolution is similar to the ranking of in vivo availability. The ranking of the three preparations in term of dissolution rate and extent of absorption is as follows: Formula 2>Formula 1 >Daonil. PMID- 15285338 TI - Effects of oily drug rheology on content uniformity in granules obtained by wet granulation with a high-shear mixer. AB - The purpose of the present work was to elucidate the effects of viscosity on the content uniformity of an oily drug in granules obtained by wet granulation with a high-shear mixer. For this purpose, we used d-alpha-tocopheryl acetate diluted with a medium chain fatty acid triglyceride having viscosities in the range from 26.0 to 726.0 mPas. It was found that independent of viscosity, nuclei rich in the oily drug were formed in the process of mixing with powder and that those nuclei prevented uniform distribution of the drug throughout the granules. To achieve content uniformity, it is necessary for the nuclei formed before granulation to be fragmented and for the oily drug to be distributed uniformly throughout granules. Tensile strength of the nuclei was attributed to the viscosity of the oily drug, according to a model for tensile strength of a granule under dynamic conditions. When viscosity of the oily drug increased, tensile strength of the nuclei increased and the extent of the drug demixing in granules was large and constant independent of granulation time. On the other hand, when viscosity of the oily drug decreased, tensile strength of the nuclei decreased. The extent of the drug demixing was small with lower viscosity but increased with a prolonged granulation time. In the case of the oily drug, we found that a decrease in its viscosity led to the improvement of the content uniformity in granules. The viscosity of the oily drug significantly affects its content uniformity in granules by a high-shear mixer granulation. PMID- 15285340 TI - An explanation for the physical instability of a marketed fixed dose combination (FDC) formulation containing isoniazid and ethambutol and proposed solutions. AB - An investigation was carried out to explore the possible reason for the physical instability of a marketed strip packaged anti-TB fixed dose combination (FDC) tablet containing 300 mg of isoniazid (H) and 800 mg of ethambutol hydrochloride (E). The instability was in the form of distribution of white powder inside the strip pockets. High-performance liquid chromatography (HPLC) and liquid chromatography-mass spectrometry (LC-MS-MS) studies confirmed that both H and E were present in the powder. The same was also confirmed through Fourier-transform infrared (FTIR) spectroscopy, which also indicated absence of interaction between the two drugs. No sublimation of the drugs was observed up to 110 degrees C, indicating that the observed instability was not due to this reason. Subsequently, attention was paid to the possibility of moisture gain by the tablets through defective packaging (which was established) due to hygroscopicity of E. To understand the phenomenon further, pure drugs and their mixtures were stored under accelerated conditions of temperature and humidity [40 degrees C/75% relative humidity (RH)] and both increase in weight and physical changes were recorded periodically. The mixtures gained moisture at a higher rate than pure E and those with higher content of E became liquid, which on withdrawal from the chambers, became crystallized. The drug mixture containing H:E at a ratio of 30:70 w/w, which was similar to the ratio of the drugs in the tablets (27:73 w/w), crystallized fastest, indicating formation of a rapid crystallizing saturated system at this ratio of the drugs. It is postulated that the problem of instability arises because of the formation of a saturated layer of drugs upon moisture gain through the defective packaging material and drying of this layer with time. The study suggests that barrier packaging free from defects and alternatively (or in combination) film coating of the tablets with water resistant polymers are essential for this formulation. PMID- 15285326 TI - Sclerosing stromal cell tumor of the ovary in pregnancy: a case report. AB - A rare case of benign ovarian stromal cell tumor during pregnancy is presented. Because of a rapidly growing solid ovarian mass, 6 x 7 cm in diameter, a 21-year old woman at 14 weeks of gestation was explored via laparotomy. Histopathological diagnosis was sclerosing stromal tumor of the ovary. She had no complaint of menstrual irregularities before pregnancy and there was no clinical or hormonal evidence of active androgenic hormone secretion. Immunohistochemical staining showed positive vimentin, smooth-muscle actin and desmin reactions. Sclerosing stromal tumor is a very rare condition in pregnancy and our case is only the eighth case detected during pregnancy according to the literature. PMID- 15285339 TI - Preparation, evaluation, and NMR characterization of vinpocetine microemulsion for transdermal delivery. AB - A novel microemulsion was prepared to increase the solubility and the in vitro transdermal delivery of poorly water-soluble vinpocetine. The correlation between the transdermal permeation rate and structural characteristics of vinpocetine microemulsion was investigated by pulsed field gradient nuclear magnetic resonance (PFG-NMR). For the microemulsions, oleic acid was chosen as oil phase, PEG-8 glyceryl caprylate/caprate (Labrasol) as surfactant (S), purified diethylene glycol monoethyl ether (Transcutol P) as cosurfactant (CoS), and the double-distilled water as water phase. Pseudo-ternary phase diagrams were constructed to obtain the concentration range of each component for the microemulsion formation. The effects of various oils and different weight ratios of surfactant to cosurfactant (S/CoS) on the solubility and permeation rate of vinpocetine were investigated. Self-diffusion coefficients were determined by PFG NMR in order to investigate the influence of microemulsion composition with the equal drug concentration on their transdermal delivery. Finally, the microemulsion containing 1% vinpocetine was optimized with 4% oleic acid, 20.5% Labrasol, 20.5% Transcutol P, and 55% double-distilled water (w/w), in which drug solubility was about 3160-fold higher compared to that in water and the apparent permeation rate across the excised rat skin was 36.4 +/- 2.1 microg/cm2/h. The physicochemical properties of the optimized microemulsion were examined for the pH, viscosity, refractive index, conductivity, and particle size distribution. The microemulsion was stable after storing more than 12 months at 25 degrees C. The irritation study showed that the optimized microemulsion was a nonirritant transdermal delivery system. PMID- 15285341 TI - Enhancement effect of p-menthane-3,8-diol on in vitro permeation of antipyrine and indomethacin through Yucatan micropig skin. AB - The enhancing effect of p-Menthane-3,8-diol (MDO) on skin permeation of antipyrine (ANP) and indomethacin (IM) through Yucatan micropig skin in vitro was compared with 1-menthol. p-Menthane-3,8-diol is a metabolite of 1-menthol and has little odor. It is easy to combine the vehicle because of lower lipophilicity than 1-menthol. All formulations contained 40% (v/v) ethanol. The permeation of ANP increased with MDO about three times that without enhancer by increasing ANP concentration in the skin. However, the MDO effect was about a quarter that of 1 menthol. The permeation of IM with MDO was about 15 times that with no enhancer and it was almost the same as that with 1-menthol. The lag time of permeation was not significantly changed by MDO, which was not so in the case of 1-menthol. Skin concentration of IM increased about 11 times and six times with MDO and 1 menthol, respectively. MDO and 1-menthol partitioned to the skin relatively high concentrations, 5.9 and 2.5 mg/ cm3, respectively. The solubility of IM in the skin was improved by MDO, and consequently, the permeation of IM was enhanced. PMID- 15285344 TI - Persistent organic pollutants in air and vegetation from the Canadian Rocky Mountains. AB - The exchange of chlorinated organic pollutants between air and vegetation in cold, mountain environments was investigated through the extraction of coniferous vegetation and high-volume air samples collected from the Canadian Rocky Mountains during the summers of 1999 and 2000. Concentrations of several compounds in vegetation increased as temperatures decreased, whereas atmospheric concentrations were not related to temperature. Daily cycling of these compounds between air and vegetation as a result of diurnal temperature changes was not observed. Compared with concentrations in vegetation from the Canadian Rocky Mountains, plant samples from the western valley in British Columbia (Canada) showed higher pollutant levels. Chemical partitioning between vegetation and air was not correlated with temperature, indicating that air contamination is governed by long-range transport and not by local revolatilization events. Based on these observations, we show that both deposition at higher altitudes and long range atmospheric transport influence chemical accumulation in vegetation from the Canadian Rocky Mountains. PMID- 15285343 TI - Sorption of steroid estrogens to soils and sediments. AB - Steroid estrogens at sub-micrograms per liter levels are frequently detected in surface water, and increasingly cause public concern of their potential impacts on ecosystems and human health. Assessing the environmental fate and risks of steroid estrogens requires accurate characterization of various physicochemical and biological processes involving these chemicals in aquatic systems. This paper reports sorption of three estrogens, 17beta-estradiol (estradiol), estrone, and 17alpha-ethinyl estradiol (EE2), by seven soil and sediment samples at both equilibrium and rate-limiting conditions. The results indicated that attainment of sorption equilibrium needs about 2 d when aqueous estrogen concentrations (C(t)s) are 25 to 50% of their solubility limits (S(W)S), but equilibrium requires 10 to 14 d when the Ct is 20 times lower than the S(W). The measured sorption isotherms are all nonlinear, with the Freundlich model parameter n ranging from 0.475 to 0.893. The observed isotherm nonlinearity correlates to a gradual increase of single-point organic carbon-normalized sorption distribution coefficient (capacity) (K(OC)) as the equilibrium estrogen concentration (Ce) decreases. At Ce = 0.5S(W), all three estrogens have log K(OC) values of 3.14 to 3.49, whereas at Ce = 0.02S(W), the log K(OC) values for estrone, EE2, and estradiol are within ranges of 3.40 to 3.81, 3.45 to 3.85, and 3.71 to 4.12, respectively. This study suggests that, when at sub-micrograms per liter levels, these estrogenic chemicals may exhibit even slower rates and greater capacities of sorption by soils and sediments. PMID- 15285346 TI - Toxicity testing of 16 priority polycyclic aromatic hydrocarbons using Lumistox. AB - Hazard assessment of industrial sites contaminated with coal tar and its products usually focuses on selected pollutants such as the 16 polycyclic aromatic hydrocarbons (PAHs) prioritized by the U.S. Environmental Protection Agency (U.S. EPA). The aim of this study was to investigate to which extent these 16 PAHs contribute to the Vibrio fischeri bioluminescence inhibition measured by the acute Lumistox luminescent bacteria test. Five of the 16 PAHs-naphthalene (NAP), acenaphthylene (ACY), acenaphthene (ACE), fluorene (FLU), and phenanthrene (PHE) revealed inhibiting effects when measuring saturated aqueous solutions of these compounds. However, in elutriates of PAH-contaminated soils, the amount of leached PAHs was very low, and the 16 PAHs did not considerably contribute to the observed bioluminescence inhibition. Nevertheless, bioluminescence inhibition was higher for elutriates with increased PAH concentration indicating the presence of other toxicants that co-occur with the 16 PAHs. No evidence was observed for increased bioluminescence inhibition due to synergistic effects among PAHs as calculated on the basis of toxic units for an aqueous solution containing all 16 priority PAHs. Data suggest that the U.S. EPA PAHs play only a minor role in causing acute toxicity to V. fischeri when exposed to aqueous elutriates of PAH contaminated soils. PMID- 15285345 TI - Influence of hydroxypropyl-beta-cyclodextrin on the extraction and biodegradation of phenanthrene in soil. AB - A study was conducted to investigate the effect of hydroxypropyl-beta cyclodextrin (HPCD) on the aging and biodegradation of phenanthrene (PHE) in soil. Soil was spiked with PHE at 25 mgPHE/kgSOIL and HPCD at a range of concentrations from 0 to 3.5 gHPCD/kgSOIL and aged for 1, 84, and 322 d. At each time point, a variety of analyses were performed to assess the loss and aging of the PHE in the soil. Methods included determination of total PHE remaining, dichloromethane (DCM) and butan-1-ol (BuOH) extractions, and determination of PHE extractable by an aqueous HPCD shake extraction. Mineralization assays were also carried out to assess the availability of the PHE to a PHE-degrading bacterial inoculum. It was found that the presence of HPCD in the soils increased PHE loss from the aged soil systems, particularly at the higher application rates. Dichloromethane and BuOH extractabilities were reduced with aging and increasing HPCD concentration, as was the amount of PHE that was extractable using an aqueous HPCD shake extraction or that was available for mineralization. The DCM and BuOH extraction yielded similar results, and both greatly overestimated the availability of the PHE to the degraders, whereas the HPCD extraction results were very similar to that of PHE biodegradation. This study indicates that cyclodextrins have potential for use as alternatives to surfactants in enhancing the desorption/solubilization and degradation of recalcitrant organic contaminants in soil. PMID- 15285342 TI - Extrusion granulation and high shear granulation of different grades of lactose and highly dosed drugs: a comparative study. AB - Formulations containing different lactose grades, paracetamol, and cimetidine were granulated by extrusion granulation and high shear granulation. Granules were evaluated for yield, friability, and compressibility. Tablets were prepared from those granules and evaluated for tensile strength, friability, disintegration time, and dissolution. The different lactose grades had an important effect on the extrusion granulation process. Particle size and morphology affected powder feeding and power consumption, but had only a minor influence on the granule and tablet properties obtained by extrusion granulation. In contrast, the lactose grades had a major influence on the granule properties obtained by high shear granulation. Addition of polyvinylpyrrolidone (PVP) was required to process pure paracetamol and cimetidine by high shear granulation, whereas it was feasible to granulate these drugs without PVP by extrusion granulation. Granules prepared by extrusion granulation exhibited a higher yield and a lower friability than those produced by high shear granulation. Paracetamol and cimetidine tablets compressed from granules prepared by extrusion granulation showed a higher tensile strength, lower friability, and lower disintegration time than those prepared from granules produced by high shear granulation. Paracetamol tablets obtained via extrusion granulation exhibited faster dissolution than those obtained via high shear granulation. For all lactose grades studied, extrusion granulation resulted in superior granule and tablet properties in comparison with those obtained by high shear granulation. These results indicate that extrusion granulation is more efficient than high shear granulation. PMID- 15285350 TI - Anaerobic transformation of compounds of technical toxaphene. 2. Fate of compounds lacking geminal chlorine atoms. AB - The major toxaphene metabolites in sediment and soils (2-exo,3-endo,6-exo,8,9,10 hexachlorobornane [B6-923] and 2-endo,3-exo,5-endo,6-exo,8,9,10 heptachlorobornane [B7-1001]) were incubated with the isolated gram-negative bacterium Dehalospirillum multivorans. Within 14 d, biotransformation of B7-1001 was nearly quantitative, resulting in two penta- and six hexachlorobornanes, as well as one unsaturated hexachloro compound of technical toxaphene. The major transformation product (approximately 50% of all metabolites) was identified as 2 exo,3-endo,5-exo,8,9,10-hexachlorobornane (B6-903). Abiotic dehydrochlorination of B7-1001 with methanolic KOH resulted in the formation and subsequent identification of the lone unsaturated compound as 2,5-endo,6-exo,8,9,10 hexachloroborn-2-ene. Thus, dehydrochlorination was found to be a minor process of the anaerobic transformation of toxaphene. Biotransformation of 70% of amended B6-923 within 14 d demonstrated that reductive dechlorination was not exclusively associated with geminal Cl atoms, as previously suggested. Three pentachlorobornanes were identified as transformation products, one of which was identical with a transformation product of B7-1001. This commonality unequivocally proves this metabolite to be 2-exo,3-endo,8,9,10 pentachlorobornane. Fifteen previously unknown metabolites of B6-923, B7-1001, and other toxaphene compounds identified in this study were detected in sediment from Lake Ontario (Canada), underscoring the importance of microbial toxaphene transformation in natural, aquatic environments. PMID- 15285351 TI - Determination of degradation products of alkylphenol polyethoxylates in municipal wastewaters and rivers in Tokyo, Japan. AB - An analytical method for the simultaneous determination of nonylphenoxy acetic acid (NP1EC), nonylphenol monoethoxy acetic acid (NP2EC), nonylphenol (NP), octylphenol (OP), and nonylphenol monoethoxylate (NP1EO) was developed. The method was applied to environmental samples to demonstrate the distribution and behavior of nonylphenol polyethoxylates and their degradation intermediates in aquatic environments in Tokyo, Japan. In sewage treatment plants, more than 85% of NP, OP, and NP1EO were removed, whereas NP1EC and NP2EC were generated during the treatment. Concentrations of NP1EC and NP2EC in secondary effluents (1.9-2.9 microg/L) were higher than those of NP and NPIEO (0.12-0.63 microg/L). In river waters. NPIEC and NP2EC concentrations (0.11-2.8 microg/L) were higher than NP and NP1EO concentrations (<0.015-3.4 microg/L). with some exceptions. In surface sediments, neither NPIEC nor NP2EC was detectable (<0.01 microg/g dry wt) whereas NP and NPIEO were detected significantly (0.03-1.82 microg/g dry wt and 0.04-0.46 microg/g dry wt, respectively). PMID- 15285352 TI - Derivation of a chronic site-specific water quality standard for selenium in the Great Salt Lake, Utah, USA. AB - The purpose of this study was to develop a site-specific water quality standard for selenium in the Great Salt Lake, Utah, USA. The study examined the bioavailability and toxicity of selenium, as selenate, to biota resident to the Great Salt Lake and the potential for dietary selenium exposure to aquatic dependent birds that might consume resident biota. Because of its high salinity, the lake has limited biological diversity with bacteria, algae, diatoms, brine shrimp, and brine flies being the only organisms present in the main (hypersaline) portions of the lake. To evaluate their sensitivity to selenium, a series of acute and chronic toxicity studies were conducted on brine shrimp (Artemia franiciscana), brine fly (Ephydra cinerea), and a hypersaline alga (Dunaliella viridis). The resulting acute and chronic toxicity data indicated that resident species are more selenium tolerant than many freshwater species. Because sulfate is known to reduce selenate bioavailability, this selenium tolerance is thought to result in part from the lake's high ambient sulfate concentrations (>5,800 mg/L). The acute and chronic test results were compared to selenium concentrations expected to occur in a mining effluent discharge located at the south end of the lake. Based on these comparisons, no appreciable risks to resident aquatic biota were projected. Field and laboratory data collected on selenium bioaccumulation in brine shrimp demonstrated a linear relationship between water and tissue selenium concentrations. Applying a dietary selenium threshold of 5 mg/kg dry weight for aquatic birds to this relationship resulted in an estimate of 27 microg/L Se in water as a safe concentration for this exposure pathway and an appropriate chronic site-specific water quality standard. Consequently, protection of aquatic birds represents the driving factor in determining a site-specific water quality standard for selenium. PMID- 15285353 TI - Glycosidation of chlorophenols by Lemna minor. AB - Metabolic fate of xenobiotics in plant tissues has an important role in the ultimate fate of these compounds in natural and engineered systems. Chlorophenols are an important class of xenobiotics used in a variety of biocides and have been shown to be resistant to microbial degradation. Three chlorophenyl glycosides were extracted from tissues of Lemna minor exposed to 2,4-dichlorophenol (DCP). The products were identified as 2,4-dichlorophenyl-beta-D-glucopyranoside (DCPG), 2,4-dichlorophenyl-beta-D-(6-O-malonyl)-glucopyranoside (DCPMG) and 2,4 dichlorophenyl-beta-D-glucopyranosyl-(6 --> 1)-beta-D-apiofuranoside (DCPAG). Identification was based on reverse phase retention (C18), electrospray mass spectra collected in negative and positive mode (ESI-NEG and ESI-POS, respectively), and nuclear magnetic resonance (NMR) spectra comparisons to reference materials synthesized in the laboratory. Liquid chromatography-mass spectrometry (LC-MS) analysis of plants exposed to 2,4,5-trichlorophenol (TCP) formed analogous compounds: 2,4,5-trichlorophenyl-beta-D-glucopyranoside (TCPG), 2,4,5-trichlorophenyl-beta-D-(6-O-malonyl)-glucopyranoside (TCPMG) and 2,4,5 trichlorophenyl-beta-D-glucopyranosyl-(6 --> 1)-beta-D-apiofuranoside (TCPAG). Enzyme catalyzed hydrolysis with beta-glucosidase was ineffective in releasing the beta-glucosides with chemical modifications at C6. Presence of these glucoconjugates confirmed that L. minor was capable of xenobiotic uptake and transformation. Identification of these products suggested that chlorophenols were incorporated into vacuoles and cell walls of L. minor. PMID- 15285354 TI - Behavioral response of young rainbow trout (Oncorhynchus mykiss) to forest fire retardant chemicals in the laboratory. AB - Fire-retardant chemicals often are applied in relatively pristine and environmentally sensitive areas that are potentially inhabited by endangered or threatened aquatic species. Avoidance of contaminants is an adaptive behavior that may reduce exposure to harmful conditions. We evaluated the avoidance responses of rainbow trout (Oncorhynchus mykiss) to concentrations of fire retardant chemicals and alternate constituent formulations ranging from 0.65 to 26 mg/L. Countercurrent avoidance chambers were used in a flow-through design with receiving water at each end and a drain at the center to create a distinct boundary between treatment water and reference water. Rainbow trout consistently avoided water treated with retardants at all concentrations tested. The magnitude of the avoidance response did not appear to follow a concentration-response relationship, but rather was an all-or-none response. PMID- 15285348 TI - Combined effects of humic acids and salinity on solid-phase microextraction of DDT and chlorpyrifos, an estimator of their bioavailability. AB - The unbound portion of dissolved hydrophobic environmental contaminants generally is presumed to be the most bioavailable to aquatic organisms. The effects of differing concentrations of Aldrich humic acid (HA) and salinity on the freely dissolved fraction of chlorpyrifos (CPF) and 4,4'-dichlorodiphenyltrichloroethane (DDT) in water were assessed through their relative uptake by solid-phase microextraction (SPME). This extraction method has been recently suggested as a valuable biomimetic tool. Increasing salinity (0-20 parts per thousand [parts per thousand]) alone had no effect on the uptake of DDT by the SPME fiber, but generally enhanced the SPME uptake of the more water soluble CPE Solid-phase microextraction uptake of DDT was decreased at an HA concentration of 10 mg/L, but 100 mg/L was required to decrease CPF uptake. Binding of CPF and DDT by HA was greatly reduced by the presence of salt at 5 to 20 parts per thousand. The extent of the HA and salinity effects appeared to be pesticide-dependent. These factors may ultimately impact the contaminant's environmental fate, transport, and bioavailability, for example, in estuarine situations. PMID- 15285349 TI - Effect of humic acids on toxicity of DDT and chlorpyrifos to freshwater and estuarine invertebrates. AB - The effects of dissolved humic acids (HAs) on the acute toxicities of the organophosphate pesticide chlorpyrifos and the organochlorine pesticide 4,4' dichlorodiphenyltrichloroethane (DDT) were assessed by using freshwater crustaceans (Ceriodaphnia dubia) and saltwater crustaceans (Americamysis bahia). The effects of filtered Aldrich HAs (10-100 mg/L) on organism mortality were determined. Humic acids had no effect on mortality of A. bahia for either pesticide at a salinity of 20 parts per thousand, but greatly reduced the mortality of C. dubia for both pesticides in freshwater (0 parts per thousand). In the latter case, the effect was proportional to the HA concentration. The difference in toxicity mitigation as a function of salinity is believed to be due to conformational changes in the HA molecules, which impact pesticide-HA binding, rather than to organismal effects. PMID- 15285347 TI - Chemometric modeling of main contamination sources in surface waters of Portugal. AB - Various chemometric data analysis methods, such as principal components analysis, multivariate curve resolution-alternating least squares, parallel factor analysis, and Tucker3, are proposed and compared for the resolution and modeling of main contamination sources in a large environmental data array obtained in an exhaustive environmental monitoring program that examined the quality of surface waters of Portugal. The study covered the analysis of 19 priority semivolatile organic compounds (SVOCs) frequently found in a total number of 644 surface water samples, including 46 different sites from throughout Portugal and corresponding to a period of 14 months, from April 1999 to May 2000. Main contamination sources of the analyzed SVOCs were identified and interpreted according to their chemical composition and according to their resolved geographical and temporal distribution profiles. PMID- 15285358 TI - Mechanisms of cadmium toxicity in terrestrial pulmonates: programmed cell death and metallothionein overload. AB - A sublethal dose of cadmium (Cd2+) administered via the diet during short-term exposure over 10 d induced programmed cell death in the hepatopancreas of the terrestrial pulmonate snail Helix pomatia. Condensed cell residues were predominantly phagocytosed by calcium cells, suggesting a specific function of these epithelial cells in metal detoxification or in clearing the organ of cellular debris from cell death. The considerable cell loss recorded by histological analysis was accompanied by enhanced cell proliferation. Intoxication with Cd was further associated with the pronounced abundance of residual bodies, predominantly recorded in excretory cells, and with pathological changes in the endoplasmic reticulum. During long-term Cd exposure, mortality increased with increasing Cd concentrations in the diet, as demonstrated by feeding experiments in the laboratory. Lethal effects of Cd appeared to be correlated with Cd overloading of the Cd-specific metallothionein isoform (Cd MT), isolated and characterized previously from the animal's hepatopancreas. Stoichiometric analysis shows that the capacity of Cd-MT to bind six molar equivalents of Cd corresponds to a tissue Cd concentration of approximately 4 micromol/g dry weight. At this tissue concentration, all high-affinity metal binding sites of Cd-MT are occupied by Cd2+. Cadmium exposure beyond this level gives rise to progressive destabilization of Cd-MT cluster structure in vitro, resulting in increasing proportions of weakly bound, or even unbound, Cd2+ ions. Our results suggest that in vivo, the observed overburdening of Cd-MT with Cd2+ reduces the viability of affected animals. PMID- 15285355 TI - Trivalent chromium alters gene expression in the mummichog (Fundulus heteroclitus). AB - Mummichogs (Fundulus heteroclitus) were used as a model fish species to study the effects of trivalent chromium exposure. To ascertain chromium's effects, we examined altered gene expression by differential display between fish exposed in the laboratory and fish collected from a chromium-impacted estuarine site. Twenty differentially expressed genes were found from either laboratory-exposed fish or in fish collected from the field site. Database sequence searches indicated that several of these genes are highly homologous to known sequences, including a fatty acid-binding protein (FABP), cytochrome P4502N2 (CYP2N2), and a precursor to the translation initiation factor eIF2B. Verification of the differentially expressed genes by real-time polymerase chain reaction (PCR) revealed that the fatty acid-binding protein was repressed to a 3.6-times greater extent (3.6-fold) in the field-site animals as compared to a reference site, eIF2B was repressed 2 fold, and an expressed sequence tag (EST) termed A31 was induced 2.6-fold. In the laboratory-exposed animals, A31 was also induced between 2- and 4-fold. However, in contrast to the field-site fish, FABP was upregulated in the chromium-exposed animals. We hope to be able to use A31 as a biomarker for ascertaining the impacts of chromium exposure on fish. PMID- 15285356 TI - Interlaboratory comparison of a reduced volume marine sediment toxicity test method using the amphipod Ampelisca abdita. AB - The U.S. Environmental Protection Agency has standardized methods for performing acute marine amphipod sediment toxicity tests. A test design reducing sediment volume from 200 to 50 ml and overlying water from 600 to 150 ml was recently proposed. An interlaboratory comparison was conducted to evaluate the precision of this reduced sediment volume toxicity test method using the marine amphipod Ampelisca abdita. A negative control and three sediment samples of varying degrees of toxicity ranging from low to high were tested by six laboratories. Complete agreement was reached in rank of relative toxicity for all samples tested by five out of six laboratories. Test acceptability for control survival was achieved by all laboratories, and 69% agreement in classification of the sediments as toxic or nontoxic was documented. Coefficients of variation in all test samples were similar to those reported in other interlaboratory studies using marine amphipods. Results of this study indicate that the reduced sediment volume test using A. abdita is a reliable and precise measure of acute toxicity in marine sediment samples. PMID- 15285359 TI - A comparison of the fish bioconcentration factors for brominated flame retardants with their nonbrominated analogues. AB - Flame retardants (FR) play a significant role in reducing the flammability of many consumer products. On a volume basis, approximately 25% of the FRs in use today utilize bromine as the active flame-retarding moiety. Their applications are those requiring high FR performance or in resins needing an FR that is active during the gas phase. Laboratory fish bioconcentration factors (BCFs) for 11 brominated FRs (BFRs) or their components were compared with those for their nonbrominated analogues. Bioconcentration, defined here as a BCF of greater than 1,000, was not observed in those brominated molecules examined with molecular weights of 700 or greater. These included the decabromodiphenyl oxide and octabromodiphenyl oxide commercial products, ethane 1,2-bis(pentabromophenyl), ethylene bis-tetrabromophthalimide, and decabromobiphenyl. Tetrabromobisphenol A, with a molecular weight of less than 700, also did not bioconcentrate. This likely relates to the ease with which it is metabolized and eliminated. Within the BFR class of polybrominated diphenyl oxides/ethers, the BCFs for those congeners with molecular weights of between approximately 450 and 700 varied with the number and position of the bromine atoms. The BCF of hexabromocyclododecane appeared to be related to its cyclododecane substructure, not to its bromine content. Bioconcentration was not a characteristic feature of the BFRs examined here. PMID- 15285360 TI - Effects of exposure to a combination of zinc- and lead-spiked sediments on mouthpart development and growth in Chironomus tentans. AB - Exposures to either zinc or lead in contaminated sediments have been shown to induce characteristic deformities in larval chironomids. This study examined the effects of exposure to lead and zinc in combination on Chironomus tentans larvae. Proportions of mouthpart deformities in populations of larvae reared in sediments containing nominal combinations of lead and zinc were tested for additive, synergistic, and antagonistic interactions using logistic regression. Metal body burdens, body size measurements, and survival were used to evaluate toxicity and developmental impacts. Results demonstrate zinc and lead mixtures produce fewer deformities than the individual metal, so their interaction may be characterized as antagonistic. However, exposure to the metal mixtures also caused delayed development and failure to hatch. The apparent decline in deformities may be an artifact of higher mortalities or developmental effects. This research provides better understanding of some of the problems and considerations for use of chironomid population deformity proportions in bioassessments for sediment metal contamination. PMID- 15285357 TI - Tumor prevalence and biomarkers of exposure and response in brown bullhead (Ameiurus nebulosus) from the Anacostia River, Washington, DC and Tuckahoe River, Maryland, USA. AB - We evaluated liver and skin tumor prevalence and biomarkers of exposure and response in brown bullhead (Ameiurus nebulosus) from three locations in the Anacostia River (Washington, DC, USA), a Chesapeake Bay region of concern. The Tuckahoe River (Maryland, USA) served as a reference. Each river was sampled in fall 2000 and spring 2001. In the Anacostia, prevalence of liver tumors was 50 to 68%, and prevalence of skin tumors was 13 to 23% in large (> or = 260 mm, age > or = 3 years) bullheads. Liver and skin tumor prevalence was 10 to 17% and 0%, respectively, in small (150-225 mm, age 1-2 years) bullheads. Tuckahoe bullhead liver tumor prevalence was 0 to 3% (large) and 0% (small); none had skin tumors. Biliary polynuclear aromatic hydrocarbon (PAH)-like fluorescent metabolites and liver DNA adduct concentrations were elevated in large and small Anacostia bullheads. Mean adduct concentrations were 16 to 28 times higher than those in Tuckahoe fish. Chromatograms revealed a diagonal radioactive zone, indicating polycyclic aromatic compound (PAC)-DNA adducts. The biomarker data and the 10 to 17% liver tumor prevalence at ages 1 to 2 suggest that these year classes are likely to have a high prevalence as they reach age 3 and older. This study provides the strongest evidence to date of the role of PAHs in tumor development in Anacostia bullheads. PMID- 15285364 TI - Mechanistic approaches for evaluating the toxicity of reactive organochlorines and epoxides in green algae. AB - Reactive electrophilic chemicals, such as reactive organochlorine compounds or epoxides, react specifically with a broad spectrum of nucleophilic biomolecules, including proteins and DNA. Conventional toxicity tests for algae, involving the observation of growth inhibition, i.e., the inhibition of cell multiplication, after several days, yield unreliable information for risk assessment because reactive compounds hydrolyze to different extents during the exposure period. The diversity of their modes of toxic action further complicates effect assessment and calls for methods yielding additional information on the mechanisms of toxicity. One of the primary targets of reactive chemicals in cells is the tripeptide glutathione (GSH), which is important for detoxification but can also be regarded as a toxicity sensor because changes in glutathione levels indicate stress. A vital system for algae is the photosynthetic system, which is indirectly affected by reactive chemicals. The test systems developed in this study for the assessment of reactive toxicity toward algae were therefore based not only on nonspecific toxicity indicators like growth inhibition but also on indicators for disturbance of photosynthesis (inhibition of photosystem II quantum yield) and glutathione metabolism. The application of the developed test systems on Scenedesmus vacuolatus after short-term exposure of 2 h showed that these tests can be used as fast screening tests for algal toxicity and in mode-of action-based test batteries. PMID- 15285361 TI - Trace metal availability and effects on benthic community structure in floodplain lakes. AB - Effects of contaminants on communities are difficult to assess and poorly understood. We analyzed in situ effects of trace metals and common environmental variables on benthic macroinvertebrate communities in floodplain lakes. Alternative measures of trace metal availability were evaluated, including total metals, metals normalized on organic carbon (OC) or clay, simultaneously extracted metals (SEM), combinations of SEM and acid-volatile sulfide (AVS), and metals accumulated by detritivore invertebrates (Oligochaeta). Accumulated metal concentrations correlated positively with sediment trace metals and negatively with surface water dissolved OC. Sixty-eight percent of the variation in benthic community composition was explained by a combination of 11 environmental variables, including sediment, water, and morphological characteristics with trace metals. Metals explained 2 to 6% of the community composition when SEM-AVS or individual SEM concentrations were regarded. In contrast, total, normalized, and accumulated metals were not significantly linked to community composition. We conclude that examination of SEM or SEM - AVS concentrations is useful for risk assessment of trace metals on the community level. PMID- 15285362 TI - Silver uptake by a marine diatom and its transfer to the coastal copepod Acartia spinicauda. AB - Silver (Ag) is an important metal contaminant in many coastal waters and often is accompanied by high nutrient concentrations in the effluent outfall. The biological uptake of Ag by the marine diatom Thalassiosira pseudonana at various levels of nitrate, ammonium, and phosphate was examined under different growth conditions of the cells. The biological uptake of Ag increased significantly with increasing concentrations of nitrate, ammonium, and phosphate, presumably because of the increase in algal growth rate or increasing sulfur-containing ligands available for Ag transport. The calculated Ag uptake rate constants increased by 3 to 16 times with increasing nitrate and ammonium concentration from 5.88 microM to 176 microM. The assimilation efficiency (AE) of Ag by the coastal copepod Acartia spinicauda was quantified under different ecological and chemical conditions of the diatom prey. The Ag AEs were in the range of 3 to 23%, and increased with decreasing diatom food concentrations or when the diatoms were grown at a lower nutrient level. Significant correlations were demonstrated between the AE and the distribution of Ag in the diatom's cytoplasm, the Ag concentration factor in the diatoms, Ag retention in the particles during the feeding period, and the Ag gut passage time in the copepods. Desorption within the copepod's gut appears to play a critical role in Ag assimilation and partially accounts for the variability of Ag AEs under different food and geochemical conditions. Our study highlights that several geochemical and physiological processes all significantly affect Ag trophic transfer in marine copepods. Given the dependence of Ag transfer on its concentration in ingested particles and food concentration, dietary uptake of Ag is probably variable in natural environments. The influence of eutrophication on Ag trophic transfer is dependent on the degree to which each kinetic parameter is affected by nutrient enrichments. PMID- 15285363 TI - Acute and chronic toxicity of nickel to a cladoceran (Ceriodaphnia dubia) and an amphipod (Hyalella azteca). AB - This study evaluated acute and chronic nickel (Ni) toxicity to Ceriodaphnia dubia and Hyalella azteca with the objective of generating information for the development of a biotic ligand model for Ni. Testing with C. dubia was used to evaluate the effect of ambient hardness on Ni toxicity, whereas the larger H. azteca was used to derive lethal body burden information for Ni toxicity. As was expected, acute C. dubia median lethal concentrations (LC50s) for Ni increased with increasing water hardness. The 48-h LC50s were 81, 148, 261, and 400 microg/L at hardnesses of 50, 113, 161, and 253 mg/L (as CaCO3), respectively. Ceriodaphnia dubia was found to be significantly more sensitive in chronic exposures than other species tested (including other daphnids such as Daphnia magna); chronic toxicity was less dependent on hardness than was acute toxicity. Chronic 20% effective concentrations (EC20s) were estimated at <3.8, 4.7, 4.0, and 6.9 microg/L at hardnesses of 50, 113, 161, and 253 mg/L, respectively. Testing with H. azteca resulted in a 96-h LC50 of 3,045 microg/L and a 14-d EC20 of 61 microg/L at a hardness of 98 mg/L (as CaCO3). Survival was more sensitive than was growth in the chronic study with H. azteca. The 20% lethal accumulation effect level based on measured Ni body burdens was 247 nmol/g wet weight. PMID- 15285365 TI - Gender and spatial patterns in metal concentrations in brown anoles (Anolis sagrei) in southern Florida, USA. AB - Comparatively little is known about heavy metal levels in reptiles, particularly for lizards. Yet lizards often are common predators that could serve as bioindicators of contamination on a small spatial scale. This study examined the differences in metal concentrations of adult brown anoles (Anolis sagrei) for arsenic, cadmium, chromium, lead, manganese, mercury, and selenium in six locations in southern Florida, USA, from Lake Okeechobee (Pahokee) south to Key West. We predicted that levels of contaminants would be higher in the industrialized ports (Port Everglades, Port of Miami) and at a landfill (in Key West) compared to a campground, tropical garden, and marine laboratory. Males were significantly larger than females, as expected. Although there were significant locational differences in metal concentrations, they did not fit a consistent pattern, either among sites or between reference and study sites. However, the lowest levels were generally found at Everglades Marina and Campground (except for selenium) and at Fairchild Tropical Garden (except for manganese). For females, size was significantly correlated with cadmium, lead, and mercury levels, while for males, there were significant correlations between length and arsenic, chromium, and manganese concentrations. Where there were gender differences in metal levels, females had significantly higher levels than males even though they were smaller. We attribute these gender differences to differences in diet due to microhabitat differences in foraging locations. Females spend more time feeding near the ground and males spend more time feeding on tree trunks and branches where they consume more flying insects. This is the first study that examines concentrations of metals in lizards in the United States, and suggests that nonnative lizards may be useful in toxicological studies. PMID- 15285367 TI - Organochlorine contaminants in sea turtles: correlations between whole blood and fat. AB - Monitoring toxic organochlorine (OC) compounds is an important aspect in wildlife studies, especially in protected species such as sea turtles. The goal of this study was to determine whether blood OC concentrations can predict those in adipose tissue of sea turtles. Blood offers many benefits for monitoring OCs. It can be collected nondestructively from live turtles and can be sampled repeatedly for continuous monitoring. Organochlorine concentrations in blood may better represent the exposure levels of target tissues, but blood concentrations may fluctuate more than those in fatty tissues following recent dietary exposure or lipid mobilization. Paired fat and blood samples were collected from 44 live, juvenile loggerhead sea turtles and 10 juvenile Kemp's ridley sea turtle carcasses. Organochlorines were analyzed using gas chromatography with electron capture detection and mass spectrometry. Lipid-normalized OC concentrations measured in the blood significantly correlated to levels found in the fat samples of both species. This result suggests that sea turtle blood is a suitable alternative to fatty tissues for measuring OCs because blood concentrations reasonably represent those observed in the paired fat samples. However, blood OC concentrations calculated on a wet-mass basis were significantly and inversely correlated to lipid content in the fat samples. Therefore, caution should be used when monitoring spatial or temporal trends, as OC levels may increase in the blood following mobilization of fat stores, such as during long migrations, breeding, or disease events. PMID- 15285370 TI - Biomonitoring of contaminants in birds from two trophic levels in the North Pacific. AB - The presence and accumulation of persistent contaminants at high latitudes from long-range transport is an important environmental issue. Atmospheric transport has been identified as the source of pollutants in several arctic ecosystems and has the potential to severely impact high-latitude populations. Elevated levels of contaminants in Aleutian Island avifauna have been documented, but the great distance from potential industrial sources and the region's complex military history have confounded identification of contaminant origins. We sampled bird species across the natural longitudinal transect of the Aleutian Archipelago to test three contaminant source hypotheses. We detected patterns in some polychlorinated biphenyl congeners and mercury that indicate abandoned military installations as likely local point sources. Carbon isotopes were distinct among island groups, enabling us to rule out transfer through migratory prey species as a contaminant source. The long-range transport hypothesis was supported by significant west-to-east declines in contaminant concentrations for most detected organochlorines and some trace metals. Although relatively low at present, concentrations may increase in Aleutian fauna as Asian industrialization increases and emitted contaminants are atmospherically transported into the region, necessitating continued monitoring in this unique ecosystem. PMID- 15285368 TI - Effect of carbon dioxide on uranium bioaccumulation in the freshwater clam Corbicula fluminea. AB - This paper presents the results of a study examining the impact of CO2 variations in water on uranium bioaccumulation in the bivalve Corbicula fluminea. The objectives were to evaluate the effect of CO2 on bivalve behavior (valve activity and ventilation rate) that are related to bioaccumulation and on the bioavailability of uranium carbonate complexes to the bivalve. It was demonstrated that at a total inorganic carbon concentration of Cco2 = 276 micromol/L, the daily valve opening duration and ventilation rate are significantly (p < 0.05) lower than those obtained at 27.6 micromol/L (-28 and 47%, respectively). For both Cco2 values, exposure to uranium at 0.25 micromol/L had no impact on valve activity; however, ventilation decreased significantly compared to the reference condition, down to the same lower level for the two Cco2 conditions. Consequently, the quantity of uranium passing through the bivalve was identical for both Cco2 conditions. Thus, bivalve ventilatory and valve activity could not explain increased bioaccumulation in the gills and mantle measured under the low-Cco2 condition. Consequently, we suggest that the quantity of carbonate bound to the U fraction must be less bioavailable than other U species such as the free-ion UO(2)2+, which is in accordance with the biotic ligand model. PMID- 15285366 TI - Use of vegetative furrows to mitigate copper loads and soil loss in runoff from polyethylene (plastic) mulch vegetable production systems. AB - The transport of runoff with high copper concentrations and sediment loads into adjacent surface waters can have adverse effects on nontarget organisms as a result of increased turbidity and degraded water quality. Runoff from vegetable production utilizing polyethylene mulch can contain up to 35% of applied copper, a widely used fungicide/bactericide that has adverse effects on aquatic organisms. Copper is primarily transported in runoff with suspended particulates; therefore, implementation of management practices that minimize soil erosion will reduce copper loads. Replacing bare-soil furrows with furrows planted in rye (Secale cereale) significantly improved the sustainability of vegetable production with polyethylene mulch and reduced the potential environmental impact of this management practice. Vegetative furrows decreased runoff volume by >40% and soil erosion by >80%. Copper loads with runoff were reduced by 72% in 2001, primarily as a result of reduced soil erosion since more than 88% of the total copper loads were transported in runoff with suspended soil particulates. Tomato yields in both years were similar between the polyethylene mulch plots containing either bare-soil or vegetative furrows. Replacing bare-soil furrows with vegetative furrows greatly reduces the effects of sediments and agrochemicals on sensitive ecosystems while maintaining crop yields. PMID- 15285374 TI - Increased kidney, liver, and testicular cell death after chronic exposure to 17alpha-ethinylestradiol in medaka (Oryzias latipes). AB - Sublethal effects observed in fish exposed to environmental estrogens may be mediated via stimulation of cell death. To investigate whether cell death is induced in fish after chronic exposure to estrogenic chemicals, Japanese medaka (Oryzias latipes) were exposed from hatch until sexual maturity to 10 ng/L 17alpha-ethinylestradiol (EE2) or acetone solvent (control). Cell death was evaluated in blinded histological sections of whole medaka using terminal dideoxynucleotidyl-mediated dUTP nick end-labeling (TUNEL), which labels nuclei of cells containing apoptotic or necrotic (fragmented) DNA. The major impact of EE2 exposure in both male and female medaka was to significantly increase the number of TUNEL-positive hepatocytes and kidney tubule cells compared to control. Cell morphology was consistent with apoptosis in the liver and cloudy swelling or necrosis in the tubule cells. The number of TUNEL-positive interstitial (hematopoietic) and glomerular cells was significantly greater in the kidneys of EE2-exposed male, but not female, medaka. The EE2 exposure also significantly increased the number of TUNEL-positive testicular cells in medaka compared to corresponding controls, namely Leydig cells, Sertoli cells, spermatocytes, and spermatids. In medaka with gonadal intersex, areas of fibrosis and areas containing female gonadal cells were relatively unstained with TUNEL. No effect of EE2 exposure on the number of TUNEL-positive ovarian somatic cells or on the rate of female ovarian follicle atresia was found. These results suggest that chronic exposure to EE2 in medaka is hepatotoxic and nephrotoxic in both sexes, whereas gonadal toxicity is specific to males. PMID- 15285373 TI - Alterations to gonadal development and reproductive success in Japanese medaka (Oryzias latipes) exposed to 17alpha-ethinylestradiol. AB - The Japanese medaka (Oryzias latipes) was used as an in vivo model to evaluate the effects of exposure to the synthetic estrogen 17alpha-ethinylestradiol (EE2) on reproductive behavior and reproductive success in fish. Exposures to EE2 began within 2 to 5 d posthatch and continued until medaka were sexually mature, between four and six months of age. Among male and female medaka exposed to EE2 at nominal concentrations of 0.2 and 2 ng/L, mating behavior and reproductive success were normal in reproductive trials. However, reproductive behavior (i.e., copulations) was suppressed in the treatment with 10 ng/L EE2. Among 19 males exposed to 10 ng/L EE2 and placed with unexposed females in reproductive trials, 16 males did not copulate and reproductive success was very low. None of the females exposed to 10 ng/L of EE2 participated in reproductive behavior with unexposed males. The reproductive trials in combination with a histological survey indicated that male fish with gonadal intersex (i.e., testis-ova) were still capable of reproductive behavior and could fertilize eggs. Even though females exposed to 10 ng/L EE2 had poor reproductive success, their ovaries showed normal development and oogenesis. These data are relevant to observations of intersex in feral fish populations. Although intersex in male fish may be an indicator of exposure to estrogenic compounds, it appears that the presence of oocytes in testicular tissue may not directly impact the reproductive capability of the male fish. However, it is clear that concentrations of endocrine disrupting chemicals (EDCs) that cause gonadal intersex are sufficient to reduce reproductive performance. PMID- 15285369 TI - Surveys of plasma vitellogenin and intersex in male flounder (Platichthys flesus) as measures of endocrine disruption by estrogenic contamination in United Kingdom estuaries: temporal trends, 1996 to 2001. AB - Plasma vitellogenin (VTG) concentrations and the presence of the ovo-testis (intersex) condition have been recorded in male flounder (Platichthys flesus) captured from several United Kingdom (UK) estuaries since 1996 as part of the endocrine disruption in the Marine Environment (EDMAR) project and earlier programs. It has been confirmed that plasma VTG concentrations in male flounder have remained elevated in several UK estuaries (e.g., Tees, Mersey, and Tyne) throughout the period covered by this study. However, the time-series data indicate that plasma VTG, a measure of environmental estrogen contamination, has decreased in fish captured from several estuaries, especially those of the Tyne and Mersey. Shorter time-series data sets from the Forth and Clyde estuaries also suggest a decrease in estrogen contamination at these sites. Trends associated with specific point sources of estrogenic contamination show site-specific patterns. For instance, plasma VTG levels in male flounder captured near the Howdon sewage treatment outfall (Tyne) have shown a steady decline to near baseline levels in 2001, while the plasma of male fish captured at a site adjacent to the Dabholm Gut discharge in the Tees estuary have shown little evidence of a sustained decline. The occurrence of the intersex condition was detected at a low but consistent prevalence through the study period, with the majority of cases recorded in fish captured from the Tyne and Mersey estuaries. The data set does not allow conclusions to be drawn about any temporal trends associated with this condition. The significance of the findings and possible mitigating influences are discussed in terms of the impacts on wild fish and the role of effluent treatment in reducing these. PMID- 15285376 TI - [A clinical evaluation of the practical reliability in video-assisted thoracic surgery for right primary lung cancer]. AB - Recently, lobectomy by video-assisted thoracic surgery (VATS lobectomy: VL) has been widely applied to peripheral lung cancer because of its less invasive approach compared to standard thoracotomy (ST). However, the appropriate approach in VL still remains to be solved. The aim of this study was to evaluate the practical reliability of our technical devices in VL for right primary lung cancer. For the VATS procedures, a mini-thoracotomy measuring about 6-7 cm was made in the fourth or fifth intercostal space (ICS) under the auscultatory triangle without rib resection. Two access holes 12 mm in size were also made in the fourth ICS at the anterior axillary line and in the seventh ICS at the posterior axillary line, respectively. These access holes were used for insertion of thoracoscope, endoscopic stapler or retracting instrument according to operative procedure. After stapling of the vessels and bronchus, the resected pulmonary lobe was removed from the thorax using a plastic retrieval bag. The present study showed the technical feasibility of this unique thoracoscopic approach in the standard lobectomy with systematic nodal dissection for right lung cancer. PMID- 15285380 TI - [A technique for off-pump ablation of atrial fibrillation on beating heart using the epicardial radiofrequency approach]. AB - Recently ablation surgery for atrial fibrillation, so-called maze procedure, has become popular. However, this procedure usually needs cardiopulmonary bypass. Here we describe 3 patients for whom the simple, new procedure in which epicardial radiofrequency ablation is sucessfully done without cardiopulmonary bypass prior to concomitant cardiac operation. Our modified maze procedure consists of isolation of pulmonary veins using the FLEX 7 radiofrequency ablation device (COBRA). Following the surgery, all patients quickly attained regular sinus rhythm. We believe our procedure would be especially useful for patients for whom concomitant cardiac procedure can be done on beating heart. PMID- 15285381 TI - [Pulmonary thromboendarterectomy for an elderly patient]. AB - We describe a case of successful pulmonary thromboendarterectomy for an elderly patient. A 75-year-old female suffered from severe dyspnea in spite of administration of pulmonary vascular dilators and anticoagulants. Her pulmonary vascular resistance was 545 dyn x sec x cm(-5) and, her pulmonary angiogram and perfusion scan revealed that almost all branches of her right pulmonary artery were occluded. Pulmonary thromboendarterectomy was performed under cardiopulmonary bypass. A videoscope was employed to see inside of the pulmonary artery. Flexible aspirator chip was used to peed abnormal intima easily. A non invasive intracranial oxygen saturation monitor was used for secure circulatory arrest. The duration of cardiopulmonary bypass was 101 minutes and minimum body temperature was 24.5 degrees C. Her symptoms improved markedly after operation, and she went back to the ordinary life. Pulmonary thromboendarterectomy was useful even for elderly patients under some modifications of the operative procedures. PMID- 15285379 TI - [Staged total cavopulmonary connection following Starnes operation]. AB - A three-year-old boy with Ebstein's anomaly, who had received Starnes operation in the neonate state, successfully underwent staged total cavopulmonary connection. Despite of hypoplasticity of the pulmonary artery, the postoperative cardiac catheterization showed no elevation of pulmonary artery pressure and good cardiac performance. Although he suffered from supraventricular tachycardia in the early postoperative period, subsequent course was uneventful. PMID- 15285382 TI - [Reduction of preparatory bank blood requirements in adult cardiac surgery]. AB - The importance of blood conservation to minimize homologous blood use in cardiac surgery is well-accepted. In addition, it is financially important to minimize unnecessary and avoidable bank blood prepared, because once the blood products were taken into a hospital, it could not be returned to the blood bank in Japan. We tried to safely reduce the amount of bank blood products requested in 185 consecutive adult cardiac operations for 18 months. First 6 months' observation revealed that only 2 patients received blood transfusion among 34 patients whose products of weight (kg) and hemoglobin (g/dl) were more than 700. We called the product "a transfusion index", and the amounts of blood prepared were newly set as follows: a transfusion index less than 500; 6 units or more of cross-matched red cell concentrate, 500-700; 2 to 4 units of cross-matched red cell concentrate, more than 700; no cross-matched bank blood. In next 6 months, the amounts of cross-matched blood reduced from 4.8 units to 3.9 units per patient. Last 6 months, we prepared no blood products for the patients whose transfusion indexes were more than 700. The amounts of prepared red cell concentrate reduced to 2.1 units per patient, however, additional blood requirements during the operations did not show significant increase. A transfusion index depends on the patient's weight and preoperative hemoglobin is a simple and useful indicator to anticipate blood requirements. PMID- 15285378 TI - [Squamous cell carcinoma caused by dysplasia on the bullous wall]. AB - We experienced a squamous cell lung carcinoma caused by dysplasia around the bullous wall after right upper lobectomy by tuberculosis. A case is 70 years old male who was resected right upper lobe caused by tuberculosis 50 years ago. He was pointed out an abnormal shadow on the chest X-ray in March 2002. There were bullous change in right lung field on the chest computed tomography (CT). There was appeared a tumor contiguous to the bullous wall. A part of bullous wall surrounding the tumor was thickened bronchio-alveolar lavage gave proof of squamous cell carcinoma. Right basal segmentectomy and subcarinal lymph node dissection was done, because of severe adhesion a right middle lobe. Radiation therapy at the mediastinum is performed, because of positive subcarinal lymph nodes. It is rare case of squamous cell carcinoma caused by dysplasia, we reported. PMID- 15285371 TI - A method for predicting bioavailability of rare earth elements in soils to maize. AB - A single-extraction procedure using low-molecular-weight organic acids (LMWOAs) as extractant and the first and second steps of a three-step extraction procedure recommended by the European Community Bureau of Reference (BCR; now European Community Standards, Measurement and Testing Programme, Brussels, Belgium) were performed to extract the light rare earth elements (LREEs) La, Ce, Pr, and Nd from wet rhizosphere soil. The extracted soil solutions were successively filtered through membranes with a pore size of less than 0.45 microm and a molecular weight cutoff of less than 1 kDa, which were termed colloidal and truly dissolved fractions, respectively. Apoplastically and symplastically bound LREEs in maize roots were experimentally distinguished by ultrasound-assisted desorption with 1 mM CaCl2 solution at 0 degrees C in ice-cooled water bath. When the LMWOAs extraction method was used, a good correlation was obtained between LREEs in soil colloidal and truly dissolved fractions and LREEs bound to apoplasm and symplasm of maize root. Both apoplastically and symplastically bound LREEs are the result of bioavailability. However, a poor correlation was obtained between LREEs in fractions water soluble, exchangeable and carbonate bound (B1) and Fe-Mn oxide bound (B2) of the BCR method and LREEs in apoplasm and symplasm and in intact roots. Hence, the LMWOAs extraction method is recommended for measuring the bioavailability of LREEs in soils. PMID- 15285372 TI - Fish full life-cycle testing for androgen methyltestosterone on medaka (Oryzias latipes). AB - Abstract-We studied the chronic effects of methyltestosterone (MT) on reproductive status of medaka (Oryzias latipes) over two generations under continuous exposure to verify the applicability of the fish full life-cycle test (FFLC) for this androgen with this species. The exposure of parental (F0) medaka to MT was begun on embryos within 12 h postfertilization and continued for up to 101 d; assessment endpoints included embryological development, hatching, posthatch survival, growth, sexual differentiation, reproduction, and hepatic vitellogenin (VTG) levels under flow-through exposure to MT at each mean measured concentration of 0.35, 1.09, 3.29, 9.98, and 27.75 ng/L. Eggs (F1) spawned from the F0 fish at 98, 99, and 100 d posthatch were examined for hatchability, survival after hatching, growth, sexual differentiation, and hepatic VTG level until 60 d posthatch. In the FFLC with medaka, MT induced masculinization of both secondary sex characteristics and gonads. We observed that all F0 fish in the 27.75-ng/L treatment group showed male secondary sex characteristics in which no fish with ovary could be discerned. Several fish with ovaries in F0 and F1 generations treated with 9.98 ng/L showed male secondary sex characteristics. We also observed swollen abdomens in the F0 and F1 female fish in the 9.98-ng/L treatment group. These swollen abdomens were induced by enlarged ovaries and were accompanied with declined fecundity and fertility in the F0 generation. These results indicate that MT reduces the reproductive potential of medaka and that the FFLC with this species is applicable to the evaluation of androgens. PMID- 15285375 TI - The spatial extent of contaminants and the landscape scale: an analysis of the wildlife, conservation biology, and population modeling literature. AB - Many contaminant releases to the terrestrial environment are of small areal extent. Thus, rather than evaluating the ecological impact on species in the immediate vicinity of the release, it may be more ecologically meaningful to determine if population impacts occur at the landscape level. In order to do this, the cumulative impact of all releases in the landscape under consideration must be evaluated. If the release sites are viewed as localized areas that are no longer available for use by ecological receptors (i.e., no longer part of the habitat), this can be thought of as a form of habitat fragmentation. Habitat fragmentation is typically viewed as the loss of large areas of habitat within a landscape, leaving small isolated patches of intact habitat within a hostile matrix. Small-scale contaminant releases, on the other hand, result in small uninhabitable areas within a primarily intact habitat. With this consideration in mind, we analyzed the wildlife and conservation biology literature to determine if information on habitat size requirements such as home-range or critical patch size could inform us about the potential for impact at the landscape level from release sites based on the size of the release alone. We determined that evaluating the impact of release size had to be conducted within a contextual basis (considering the existing state of the landscape). Therefore, we also reviewed the population modeling literature to determine if models could be developed to further evaluate the impact of the spatial extent of chemical releases on the landscape. We identified individual-based models linked to geographic information systems to have the greatest potential in investigating the role of release size with respect to population impacts at the landscape level. PMID- 15285377 TI - [Management of remaining coronary artery dissection after the replacement of the ascending aorta in acute type A aortic dissection]. AB - The authors report a case study of a 54-year-old male admitted to our hospital with severe chest pain and ST depression in II, III and aVf lead on the electrocardiogram. The chest X-ray showed an enlarged superior mediastinum. An enhanced computed tomography (CT) was performed and confirmed the diagnosis of acute type A aortic dissection. The patient underwent emergency surgical repair with the replacement of the ascending aorta. The patient recovered without complication until the fifteenth postoperative day, when another severe chest pain appeared. Emergency coronary angiography revealed a remaining dissection in both the left anterior descending artery (LAD) and the left circumflex artery (LCx). Implantation of Elite stents to the LAD and the LCx was performed. The patient recovered uneventfully after this operation. Remaining coronary artery dissection after the replacement of the ascending aorta is very rare. In this case coronary intervention with Elite stents was effective. PMID- 15285383 TI - [Surgical treatment of thymic carcinomas]. AB - Five cases of surgically treated thymic carcinoma are reported. The patients (4 men and a woman) ranged in age from 46 to 76 years old with a mean of 64.6. Four patients were asymptomatic and an abnormal shadow on X-ray films was noted. One remaining patient suffered from hoarseness. One patient had stage II disease and the others had stage III. Surgical tumor resection was performed in all cases. Only 1 patient among the 5 underwent a successful complete resection. Histological examinations of the resected specimens revealed squamous cell carcinoma of thymus. Four specimens were poorly differentiated and 1 is moderately differentiated carcinoma. All patients received radiation therapy post operatively. Three patients are alive without any recurrence 6, 8 and 109 months after the surgery. Thymic carcinomas are frequently invasive or metastatic at the time of diagnosis. But poorly differentiated group, in squamous cell carcinoma, mucoepidermoid carcinoma and besaloid carcinoma, are characterized by a low incidence of local recurrence and distant metastasis. They also have a good sensitivity for the radiation. Therefore complete surgical resection combined with postoperative radiation therapy should be a choice in treating thymic carcinomas. We considered that complete resection and postoperative radiation therapy is a curative therapy for thymic carcinomas. PMID- 15285385 TI - [Aortic dissection (DeBakey type I) associated with pseudoaneurysm 21 years after aortic valve replacement; report of a case]. AB - We report a case of 63-year-old male who suffered from aortic dissection associated with pseudoaneurysm 21 years after aortic valve replacement. Preoperative computed tomography (CT) scan and aortography revealed DeBakey type I dissecting aneurysm with maximum diameter of 70 mm on ascending aorta and calcified pseudoaneurysm at previous aortotomy. He underwent modified Bentall procedure preserving previous mechanical valve and total aortic arch replacement with elephant trunk technique. Postoperative course was uneventful. PMID- 15285384 TI - [A surgical repair of ruptured atherosclerotic coronary artery aneurysm; report of a case]. AB - We reported a successful surgery for the huge ruptured coronary aneurysm resulting in cardiac shock. The rupture of atherosclerotic coronary aneurysm is extremely rare except congenital cardiac disease and as a coronary artery pulmonary artery fistula or a coronary arterio-venous fistula. Excision of the aneurysm, closure of the coronary ostium and coronary artery bypass using saphenous vein graft was performed. The post operative course was uneventful. The histological findings revealed atherosclerotic coronary aneurysm. PMID- 15285386 TI - [Stickler syndrome with rapidly progressive mitral valve regurgitation: report of a case]. AB - A 36-year-old man with Stickler syndrome who underwent mitral valve replacement for rapidly progressive mitral regurgitation due to prolapse of an anterior leaflet. Stickler syndrome is a relatively rare condition caused by a defective collagen gene and characterized by high myopia, sensorineural-hearing deficit and flattened facial features. The prevalence of mitral valve prolapse in Stickler syndrome is reported to be much higher than in the general population because of its connective tissue dysplasia. Mitral regurgitation rapidly and refractorily progresses. Prompt surgical treatment should be recommended to mitral regurgitation in Stickler syndrome, and mitral valve replacement with a prosthetic valve is a better selection than mitral valve plasty for tissue fragility. PMID- 15285387 TI - [Re-coronary artery bypass grafting necessitated by flow insufficiency of right internal thoracic artery; report of a case]. AB - A 75-year-old woman required re-coronary artery bypass grafting due to flow insufficiency of the right internal thoracic artery (RITA). After the first operation, coronary angiography revealed a narrowed RITA. Myocardial ischemia occurred when the native coronary artery was occluded by the catheter. A stringlike internal thoracic artery is probably unable to increase blood supply immediately in response to the demand created by acute obstruction of the recipient native coronary artery. PMID- 15285388 TI - [Aortic regurgitation due to nonpenetrating trauma of the chest; report of a case]. AB - A 64-year-old man was admitted to our hospital with multiple traumas, caused by motorcycle accident. Six months later, he revisited our hospital, because he was found to have aortic regurgitation and subsequent congestive heart failure. Therefore, the aortic valve was excised and replaced with 23 mm CarboMedics prosthesis. His postoperative course was uneventful. Aortic regurgitation due to nonpenetrating trauma of the chest is rare. PMID- 15285389 TI - [Multiple myxoma in the left ventricle and the left atrium]. AB - Left ventricular myxoma is particulary rare. Our case is a 77-year-old female. Transesophageal echocardiography showed a giant tumor in the left atrium. An urgent operation was performed. A giant mass was excised en bloc via a transinteratrial septal approach. Histopathologically it was myxoma. As a transthoracic echocardiography at 1-year postoperation showed a tumor in the left ventricle. A mass was excised en bloc via a vertical approach. Histopathologically it was diagnosed again as myxoma. We looked at the earliest transesophageal echocardiogram again, and found the small tumor on the same area under the posterior mitral leaflet. At the diagnose of cardiac tumor, possibility of multiple formation should be always considered. PMID- 15285390 TI - [Idiopathic hemothorax associated with shock during transporting in an ambulance; report of a case]. AB - A 37-year-old man was admitted to our hospital. The patient noted sudden right back pain after coughing before 1 hour. Loss of consciousness was occurred in an ambulance. Chest X-P revealed whole fluid in the right chest. Enhanced chest computed tomography (CT) revealed extravasation of contrast media into the pleural cavity from the right chest wall. Thoracentesis was performed to relieve dyspnea and 2,000 ml of blood was removed. Then hemoglobin count was dropped to 3.8 g/dl. At thoracotomy whole blood was sucked about 3,900 ml. Bleeding point was found at third intercostal vein. The vein was knotted and sutured by prolene thread. The bleeding lesion was no inflammation and no string like tissue. We report a case of idiopathic hemothorax and enhanced chest CT was useful for diagnosis of bleeding lesion of pleural cavity. PMID- 15285391 TI - [Thoracoscopically diagnosed multicentric Castleman disease; report of a case]. AB - A 57-year-old female was admitted because of chest and back pain. Computed tomography (CT) revealed that many well-marginated lymph nodes were located in mediastinum and abdominal para-aortic area, especially in the right lower mediastinum. These lymph nodes were enhanced at contrast material-enhanced CT. We performed thoracoscopic surgery. The histopathologic diagnosis was multicentric Castleman disease (MCD). MCD should be considered in the differential diagnosis of multiple lymph nodes swelling with hyper globulinemia. Thoracoscopic surgery is the useful method to resect the lymph nodes and diagnose MCD. PMID- 15285392 TI - [Congenital bronchial atresia treated with video-assisted thoracoscopic surgery; report of a case]. AB - We report an adult case of congenital lobar emphysema due to bronchial atresia. A 24-year-old man was referred to our department because of cough and fever. A chest roentgenogram on admission showed multiple cystic shadows in the left lower lung field. Chest computed tomography (CT) demonstrated multiple cysts with neveau and scattered infiltration in the left lingual segment and lower lobe. Surgical treatment was scheduled because of no improvement of the chest lesions. The interlobar fissure was not found between the upper and the lower lobes, but between the upper and the lingual divisions. Additionally, the lingual bronchus was not bifurcated from the upper bronchus, but from the lower bronchus. As inflammatory changes were extended to the lingual division and the lower lobe, a left lingual segmentectomy and a lower lobectomy with video-assisted thoracoscopic surgery were performed. His postoperative course was uneventful and he was discharged at the seventh day after surgery. PMID- 15285393 TI - [Schwannoma of the recurrent laryngeal nerve; report of a case]. AB - A 57-year-old woman was admitted to our hospital because of an abnormal shadow on her chest X-ray film and a palpable left neck mass. She had a mild cough. Computed tomography (CT) scans and magnetic resonance imaging (MRI) of the chest demonstrated a well-circumscribed, huge mass (approximately 14 cm in diameter) in the left anterior mediastinum. On June 12th, 2000, the mass was resected. During surgery, the tumor seemed to arise from the left recurrent laryngeal nerve. The pathological examination revealed the tumor as a schwannoma. Recurrent laryngeal nerve schwannoma is extremely rare. PMID- 15285394 TI - [Solitary papilloma of the lung; report of a case]. AB - A 66-year-old woman admitted our hospital due to an abnormal shadow in the right lung field on a routine chest X-ray film. Preoperative diagnosis was not made, we didn't have correct diagnosis of benign or malignancy by intraoperative frozen section specimen, so segmentectomy without lymph nodes dissection was performed. The histopathological diagnosis was glandular papilloma. A solitary papilloma in the lung field is rare, it is considered to be difficult to differentiate malignancy or benign clinically. PMID- 15285395 TI - Developments in environmental auditing by supreme audit institutions. AB - At the end of the 1980s, Supreme Audit Institutions (SAIs) became aware of their responsibility towards the environment and environmental policy. In this article, the development of environmental auditing by SAIs during the last 10 years is presented, as well as the current state of the art. The description is based on the results of three questionnaire surveys held in 1994, 1997, and 2000 by the INTOSAI Working Group in Environmental Auditing. In most countries, the government has stipulated some form of environmental policy, and the SAI has a mandate to carry out regularity and/or performance audits. The activities of SAIs have developed substantially since 1993. Nowadays, environmental auditing is a substantial and regular part of the audit work of more than half of the SAIs. Environmental problems are often transboundary in nature. SAIs can contribute to international environmental cooperation by auditing the compliance of their national government with international environmental obligations and commitments. The INTOSAI Working Group on environmental auditing wants to enhance this type of audit and has provided guidelines for the audit process and the selection of international agreements. Moreover, cooperation between SAIs is a good method to exchange experiences and to learn from each other. PMID- 15285396 TI - Application of decision analysis to forest road deactivation in unstable terrain. AB - Resource managers require objective methodologies to optimize decisions related to forest road deactivation and other aspects of road management, especially in steep terrain, where road-related slope failures inflict extensive environmental damage. Decision analysis represents a systematic framework that clearly identifies real options and critical decision points. This framework links current decisions with expected future outcomes and provides advantages such as a common currency to systematically explore the liability consequences of limited budget expenditures to road deactivation and other road-related activities. Furthermore, the decision framework prevents the analysis from becoming hopelessly entangled by the vast number of possibilities generated by the alternative occurrences, magnitudes, and consequences of landslide/debris flow events and provides the information required for the first step of an adaptive management process. Here, a structured analysis of potential environmental risks for a road deactivation project in coastal British Columbia, Canada is presented. The application of decision analysis generates a ranking of the expected benefits of proposed deactivation activities on various road sections. The ranking distinguishes between road sections that offer high expected benefit from those that offer moderate to low expected benefit. Seventeen of 171, 100-m road segments accounted for 18% of the cumulative cost and 98% of the cumulative expected net benefits from road deactivation. Furthermore, the cost of deactivating a section of road is related to the expected benefit from such deactivation, thus providing the basis for more effective resource allocation and budgeting decisions. PMID- 15285397 TI - Current and future strategies for water and wastewater management of Istanbul City. AB - Istanbul has experienced rapid increases in population to more than 12 million people, which has created infrastructure problems of water supply and wastewater treatment and disposal. In this article, the achievements and approaches of the Istanbul Water and Sewerage Administration (ISKI) to solve the water shortage problem and to improve services are summarized. Istanbul had a very severe water shortage problem in 1994 because of ignorance of the implementation of the needed projects. After reviewing the reasons and causes of the problem, new priority criteria adopted after 1994 are given. Following the implementation of the projects determined according to the aforementioned criteria, water supplied has exceeded the water demand. The added capacity is equal to one to three times of the capacity built up to 1994 for water treatment, service reservoirs, pumping stations, transmission lines, and the water distribution network; water quality has been improved the meet local and international potable water standards. Unaccounted for water has been reduced from 60% to 27%. The percentage of treated wastewater has been increased from 10% to 90% in 8 years, resulting in drastic improvements and rehabilitation of the Golden Horn and coastal water quality. Through improved customer services, complaints were reduced from 33% in 1994 to 0.3%. Some of the main criteria and the approaches behind this success are summarized. PMID- 15285398 TI - Conflict and impacts of divers and anglers in a marine park. AB - The New South Wales State Government (Australia) gazetted the Jervis Bay Marine Park (JBMP) in 1998. During the preparation of the draft zoning plan in 2000, societal data on two conflicting park user groups--recreational scuba divers and fishers (anglers)--was collected. While conflict resolution was a plan priority, other factors, such as cumulative environmental impacts of users and protection for the critically endangered grey nurse shark (Carcharias taurus), further complicated planning. Both scuba diving and angling are primary summer activities and are disproportionately concentrated around the headlands of the bay. Furthermore, shore based game-fishing was concentrated on the northern headland, where the conflict was centered. However, when the exact locations of divers and anglers were determined, there was a partial partitioning of the available space, with only a small contested overlap. To resolve conflict and maximize positive environmental outcomes, a sanctuary zone and noanchoring zone option in the draft zoning plan was sought to formalize this partition. The human dimension data proved valuable in guiding environmental management in this politically volatile situation. A baseline study conducted 11 years previously was also used to gain a limited perspective on change in user numbers. Comparison between study periods indicated dive numbers had remained similar, while the number of dive charter trips was significantly less. The numbers of anglers, for the four months compared, had doubled and tripled. The actual data used to inform management is presented and the limitations of this "best available data" approach are discussed. PMID- 15285399 TI - A multidisciplinary decision support system for forest fire crisis management. AB - A wildland fire is a serious threat for forest ecosystems in Southern Europe affecting severely and irreversibly regions of significant ecological value as well as human communities. To support decision makers during large-scale forest fire incidents, a multidisciplinary system has been developed that provides rational and quantitative information based on the site-specific circumstances and the possible consequences. The system's architecture consists of several distinct supplementary modules of near real-time satellite monitoring and fire forecast using an integrated framework of satellite Remote Sensing, GIS, and RDBMS technologies equipped with interactive communication capabilities. The system may handle multiple fire ignitions and support decisions regarding dispatching of utilities, equipment, and personnel that would appropriately attack the fire front. The operational system was developed for the region of Penteli Mountain in Attika, Greece, one of the mountain areas in the country most hit by fires. Starting from a real fire incident in August 2000, a scenario is presented to illustrate the effectiveness of the proposed approach. PMID- 15285400 TI - Land evaluation for maize based on fuzzy set and interpolation. AB - The objective of this article is to apply fuzzy set and interpolation techniques for land suitability evaluation for maize in Northern Ghana. Land suitability indices were computed at point observations using the Semantic Import (SI) model, whereas spatial interpolation was carried out by block kriging. Interpolated land suitability shows a high correlation (R2 = 0.87) with observed maize yield at the village level. This indicates that land suitability is closely related to maize yield in the study area. Membership functions were further used to assess the degree of limitation of land characteristics to maize. Sixty percent of the data has membership functions ranging from 0.23 for ECEC to 1.00 for drainage. ECEC, organic C, and clay are the major constraints to maize yield. The use of the fuzzy technique is helpful for land suitability evaluation, especially in applications in which subtle differences in soil quality are of a major interest. Furthermore, the use of kriging that exploits spatial variability of data is useful in producing continuous land suitability maps and in estimating uncertainties associated with predicted land suitability indices. PMID- 15285401 TI - Influence of motivations for seeking ISO 14001 certification on perceptions of EMS effectiveness in China. AB - This study examines the motivations of mainland Chinese facilities in seeking ISO 14001 certification and examines the linkages between these motivations and self reports of the effectiveness of major environmental management system (EMS) components. In a sample of 128 facilities in Beijing, Shanghai, and Guangzhou, the main drivers for certification were reported to be to ensure regulatory compliance, to enhance the firm's reputation, and to improve environmental performance, in that order. Although motivation to achieve cost reductions were least emphasized, a broad range of motivations appears to be considered in the decision to seek certification to ISO 14001. Regression models linking these motivations to the EMS components suggests that internal motivations have an influence on most EMS components. One interesting exception to this, however, is that no significant relationship was observed between internal motivations and the promulgation of environmental objectives and targets. The relationships associated with external motivations for certification (i.e., those in response to customer and other stakeholder pressures) and EMS components, on the other hand, are weaker and tend to occur earlier in the process cycle. No significant relationships were found between motivations to reduce costs and perceptions of the effectiveness of EMS components. Overall, these findings suggest that ISO 14001, as currently being implemented in mainland China, may have a modestly useful role when used in combination with other policy mechanisms to move the Chinese economy toward more sustainable practices. It is asserted that the ISO standard could provide even greater benefits if Chinese registrars were more proactive in developing EMS in conjunction with even more rigorous third-party audits. PMID- 15285402 TI - Sustainable mining in the European Union: the legislative aspect. AB - This paper is a review of the community legislation of the European Union ("acquis communautaire") with regard to the mineral extractive industry. It highlights the existing inconsistencies of the acquis, which require correction. Historically, the mining industry has received privileged treatment within the European Community. The treaties declare the promotion of a policy of using natural resources prudently and rationally to avoid their unconsidered exhaustion. However, mining is excluded from the scope of major environmental directives or reserves a certain freedom for interpretation. This has led to an increasing number of related cases at the European Court of Justice. The regulatory tools of the environmental acquis are rather diverse in controlling the emission sources, the pollution pathways, and the impacted receptor media through administrative measures or assigning environmental quality targets. A combined approach is needed for controlling the environmental impacts of the extractive industries. The amendment of the Seveso II Directive, the elaboration of a reference document on best available techniques and the preparation of a widescope directive on mining waste management might provide a frame for the elimination of the above discrepancies. In addition, a coherent European Community policy based on the balanced consideration of economic, environmental, and social aspects could ensure a sustainable development of the mining industry. PMID- 15285403 TI - Local cultural knowledge and water resource management: the Wind River Indian Reservation. AB - Ecology and culture comprise interacting components of landscapes. Understanding the integrative nature of the landscape is essential to establish methods for sustainable management. This research takes as a unifying theme the idea that ecological and cultural issues can be incorporated through management. As a first step in developing integrative management strategies, information must be collected that compares and contrasts ecological and cultural issues to identify their areas of intersection. Specifically how can local cultural knowledge enable water resource management that reflects cultural and ecological values? This research examines Native American cultural knowledge for setting water resource management priorities in the Wind River Indian Reservation in central Wyoming. A cross-cultural approach is adopted to assess the relationship between indigenous cultural knowledge and Euro-American perspectives through a comparative examination of the Wind River Water Code and Wyoming Water Law. This research indicates that cultural perspectives provide a rich arena in which to examine management issues. Understanding and identifying cultural practices may be an important first step in collaborative resource management between different cultural groups to prevent conflict and lengthy resolution in court. PMID- 15285404 TI - Attitudes toward wilderness study areas: a survey of six southeastern Utah counties. AB - Southeastern Utah is a region of world-renowned red-rock sandstone formations, large tracts of federal public land, rural communities centered on agriculture and extractive industries, and is often at the epicenter of environmental protection efforts in the western United States. Environmental groups have proposed formal Wilderness designations for much of the region's public land- proposals that have been actively fought by rural community leaders who do not want large areas "locked-up" from traditional livelihood and recreational uses. The debate over wilderness designation in the region has been characterized in the media as one that is particularly contentious and polarizing. A survey of southeastern Utah residents was conducted in order to better understand this conflict. The survey focused on attitudes toward wilderness designation and management. We found that residents of southeastern Utah have negative attitudes towards the designation and management of Wilderness Study Areas. We propose that these attitudes should be carefully considered and engaged in future policy and management decisions. We suggest that negative opinions expressed by residents of southeastern Utah are not directed primarily at the concept of environmental protection but rather at the strong perception that these programs and initiatives have been carried out in a heavy-handed manner and dominated by outside influences that have overwhelmed local "voices." PMID- 15285405 TI - [Rifampicin use for potency stabilization in Rif(r) mutants of Streptomyces recifensis subsp. lyticus, an organism producing lytic enzymes]. AB - Rif(r) mutants 1P-92 and 2P-15 were isolated as a result of selection of Streptomyces recifensis subsp. lyticus, an organism producing lytic enzymes. The effect of rifampicin on the biosynthetic potency of the mutants was studied. When added to the medium for cultivation of Rif(r) mutants 1P-92 and 2P-15 in the optimal concentrations (7.5 and 10.0 mcg/ml respectively), the antibiotic showed stabilizing effect on their potency in successive subcultures and recovered the initial potency of the old laboratory strains. Preliminary cultivation of strain 2P-15 after its storage for 6 years at a temperature of -20 degrees C made it possible to increase the efficiency of the initial potency recovery in the analytical selection. PMID- 15285406 TI - [Determination of biological activity of hydrogel preparations based on ketoconazole and clotrimazole]. AB - Biological activity of original hydrogel preparations based on ketoconazole and clotrimazole was estimated biologically with the 3-dose variant of the agar diffusion method. The optimal concentrations of the active substances in the hydrogels were the following: 2% of ketoconazole and 1% for clotrimazole. PMID- 15285407 TI - [Paraendocervical route for metronidazole administration in complex treatment of chronic Trichomonas endocervicitis]. AB - The data on paraendocervical administration of metronidazole in the treatment of patients with relapsing Trichomonas endocervicitis are presented. Metronidazole was administered as 0.5% solution in a dose of 0.04 g once a day for 8-10 days in complex traditional therapy including oral use of metronidazole and immunocorrigating and local treatment. It was shown that paraendocervical administration of the protistocidal agent provided earlier regression of the urogenital symptoms and 2.8 times lower frequency of the relapses. PMID- 15285408 TI - [Impact of special treatment methods on life quality and lifespan of patients with widespread forms of ovarian cancer]. AB - The experience of the treatment of 138 patients with widespread forms of ovarian cancer within the period of 1998 to 2002 is described. The efficacies of the palliative treatment methods in improvement of the life quality and increase of the lifespan of the patients were compared. The results showed that combination of the chemotherapy with the subsequent citoreductive surgical interventions was advantageous. Such a treatment provided not only increase of the average lifespan of the patients from 13 to 23 months but also improvement of the life quality in 77.8% of the cases. PMID- 15285409 TI - [Clarithromycin: specific features of antimicrobial spectrum and clinical use]. PMID- 15285410 TI - [Linezolid (Zivox), a new highly efficient antibiotic for therapy of Gram positive infections]. PMID- 15285411 TI - [Glycopeptides and therapy of infections caused by multiresistant Gram-positive organisms]. PMID- 15285413 TI - [Transcranial magnetic stimulation (TMS): physiology, psychology, brain mapping and clinical applications]. AB - Non-invasive Transcranial Magnetic Stimulation (TMS) can painlessly induce conductive current to excite cortex. This changes physiological processing in the brain. Varying the parameters of TMS, differential physiologic effects, psychological impacts can be examined. Modulation of sensory perception, suppression or facilitation of cognitive capacity and behavioral performance depicts differential modulatory operation in the brain. TMS could be applied in the research of neurological disease, psychiatric disorders, and pharmacological investigation. Stereotaxic and frameless TMS can improve the anatomy-guided TMS and use for navigation in brain surgery. Finally, the principles derived from acupuncture mechanisms can be integrated into the frequency modulation effects on the brain, shared by TMS. PMID- 15285414 TI - [Factors influencing axon regeneration after CNS damage]. AB - In contrast to peripheral nerves, damaged axons in the mature mammalian brain and spinal cord rarely regenerate, and always lead to permanent loss of function. There are many reasons that inhibit the axon regeneration in the central nerve system, including the formation of glial scar, the deficiency of neurotrophic factors and the expression of inhibitory molecules. Here several factors, their structures, distributions, contributions and possible mechanisms are briefly reviewed. PMID- 15285415 TI - [C-reactive protein and atherosclerosis]. AB - Human C-reactive protein (CRP) is an acute phase protein which arises rapidly and tremendously in serum when the subject is exposed to infection and tissue injury. CRP can effectively opsonize by both activating the complement classical pathway and potentiating the phagocytosis of phagocytes, thus clearing the invading pathogens and tissue cells undergoing injury, necrosis and apoptosis and performing an important protective role in the innate immune system. It has been over 70 years since CRP was first discovered. Traditional wisdom holds that CRP is a non-specific marker of inflammation. However, accumulating evidence during the last decade has elucidated that CRP, plays a direct role in the inflammation and cardiovascular disease such as atherosclerosis and can be recognized as the most powerful risk factor and predicator of cardiovascular disease, hence at present receiving attention worldwidely. PMID- 15285416 TI - [Recent advances of protein phosphorylation in proteome]. AB - With the advent of post-genomic era, it is an important task to study the expression, modification and interaction of the total proteins in cells, tissue and organs for proteomics in biological organism. The protein phosphorylation is one of the most popular mechanisms in signal transduction and enzyme modulation. It is estimated that about 2% of the total genes encoded 500 kinases and 100 phosphatases in human genome. As the key point of expression modulation in prokaryotic and eukaryotic cells, the protein phosphorylation and dephosphorylation may help to reveal the status of the life system at the molecular level. The phosphorylation in proteome remains challenging for functional genomics. In the present review, some new advances such as identification and characterization of protein phosphorylation in proteome are summarized. PMID- 15285417 TI - [Application of RNAi technology to knockdown gene expression in vivo in mammalians]. AB - Post-transcriptional gene silencing (PTGS) initiated by dsRNA, which result in specific degradation of homologous mRNA, is called RNAi. The discovery of RNAi greatly intrigued researchers, and was followed by a flood of papers that described the phenomenon and mechanism of RNAi. More excitingly, RNAi has recently been developed into a new tool, showing promising role in reverse genetics, gene therapy and anti-viral infection. This review provides the progress of the application of RNAi technology in mammalian animals. PMID- 15285418 TI - [Discovery of susceptibility gene for coronary heart disease]. AB - Coronary heart disease (CHD) is a leading cause of death for adults in many countries. It is multifactorial in origin with both genetic and environmental factors contributing to its development. But the molecular basis of CHD pathogenesis remains obscure, which represents a major obstacle to the identification of CHD susceptibility genes. Knowledge of genetic factors contributing to the pathogenesis of CHD will undoubtedly advance efforts to identify CHD susceptibility genes. The benefits promised are great, but progress to gene identification in CHD has been rather disappointing so far. This review aims to discuss the current and future approaches to gene discovery in CHD. And these approaches can be applied equally to other multifactorial traits in the cardiovascular arena--stroke, peripheral arterial occlusive disease, hypertension, heart failure and beyond. PMID- 15285419 TI - [Progress in Notch signaling]. AB - Notch signaling plays a vital role in cell fate decisions in both invertebrate and vertebrate development. Signals exchanged between neighboring cells through the Notch receptor amplify and consolidate molecular differences, which eventually dictate cell fates. In this review, the details of notch signaling will be discussed, with a focus on Lin/Notch repeats (CSL) independent modes. PMID- 15285420 TI - [The role of myeloid differential protein-2 in innate immunity]. AB - As typical PRRs (pattern-recognition receptors), TLRs (Toll-like receptors) play an important role during innate immunity recognition. MD-2 (myeloid differential protein-2) may contain distinct functional domains that can separately and simultaneously bind TLRs (TLR4 or TLR2) and TLR ligands, such as lipopolysaccharide (LPS). The special structure of MD-2 may result in its three main functions: (1) An association with TLR4 that amplifies TLR4 responsiveness to ligands, especially LPS. (2) Enabling TLR2-mediated responses to LPS and enhancing TLR2-mediated responses to bacteria and their cell wall components. (3) Increasing the expression of TLR2 and TLR4 and possibly influencing the correct intracellular distribution of TLR4. Importantly, while MD-2 regulation of TLR expression and distribution is well established, determining whether the interaction is direct or not will require further study. Thus, MD-2 is not only an assistant molecule of TLR4 but is also a key regulatory molecule in innate immunity, and may play important roles during infection, inflammation, immune responses and many other pathologic and physiologic processes. PMID- 15285421 TI - [Nongenomic mechanism of rapid effects of steroids on immunocytes]. PMID- 15285422 TI - [Reviews on relationship between nitric oxide and olfactory recognition memory]. PMID- 15285423 TI - [Recent advance of studies on telomerase function in stem cells]. PMID- 15285424 TI - [Upper cervical cord: integrator of somatic and visceral afferent inputs]. PMID- 15285425 TI - [Tetrahydrobiopterin and endothelial dysfunction]. PMID- 15285426 TI - [The anti-angiogenesis effect of plasminogen kringle 5]. PMID- 15285427 TI - [Development and application of NF-kappaB decoy oligodeoxynucleiotide]. PMID- 15285428 TI - [The surface charge theory and influences of sialic acid on the gating of sodium and potassium channels]. PMID- 15285429 TI - [Roles of hydrogen sulphide played in central nervous system]. PMID- 15285430 TI - [The relationship between calcineurin and learning and memory]. PMID- 15285431 TI - [The research advance of nuclear receptor Nurr1]. PMID- 15285432 TI - [The inflammation in Alzheimer's disease]. PMID- 15285433 TI - [Progress on gonadotropin-releasing hormone cell models]. PMID- 15285435 TI - The scientific basis of ethics. PMID- 15285434 TI - 'Why do my patients leave me?'. PMID- 15285436 TI - Doctors and the medical aid industry. PMID- 15285437 TI - Infant feeding and prevention of mother-to-child HIV transmission. PMID- 15285438 TI - Auto-antibody testing in obstetric patients. PMID- 15285439 TI - Pressure chamber explosion--Southern African Underwater and Hyperbaric Medical Association Statement. PMID- 15285440 TI - Psychopathology and coping in recently diagnosed HIV/AIDS patients--the role of gender. PMID- 15285441 TI - Tariff relativity. PMID- 15285442 TI - Compensation for injury from medical treatment. PMID- 15285443 TI - Back to the future for dispensing. PMID- 15285444 TI - AIDS pandemic--still time to prepare for the orphan tide. PMID- 15285445 TI - 'Emergency Mac' retires--well almost. PMID- 15285446 TI - The angel in 'scrubs' together we realise well-being. PMID- 15285447 TI - Board of Health Funders of Southern Africa conference, May 2004. PMID- 15285449 TI - Sexual risk behaviours are influenced by knowing someone with HIV/AIDS. PMID- 15285448 TI - The clinical relevance of fasting serum insulin levels in obese subjects. PMID- 15285450 TI - Worms wanted, dead or alive. PMID- 15285451 TI - Medication prescription for depression in children and adolescents. PMID- 15285452 TI - Identifying acute HIV infection--a major new public health challenge. PMID- 15285453 TI - Lower limb amputation--still a challenging procedure. PMID- 15285454 TI - Incidence of acute lymphoblastic leukaemia in white and coloured children in the Western Cape. AB - Objectives. To record the age-specific incidence rate (ASIR) for diagnosed acute lymphoblastic leukaemia (ALL) in coloured and white children aged 0 - 12 years in the Western Cape (WC). DESIGN: A retrospective population-based study using the 1991 population census to calculate the mean annual childhood population and the ASIR for ALL in the 0 - 4, 5 - 9 and 10 - 12-year age groups in rural and Cape Town metropolitan areas for the period 1983 - 1999. Odds ratios were calculated using EpiInfo 2000. SETTING: Registry records of the paediatric cancer units at Tygerberg and Red Cross War Memorial Children's hospitals where all children with ALL in the WC were initially treated. SUBJECTS: All white and coloured children aged 0 - 12 years diagnosed as having ALL from 1983 - 1999. OUTCOME MEASURES: The ASIR by age and ethnic group in rural and metropolitan patients in the WC. RESULTS: The estimated annual childhood population in 1991 was 709 151 with 80.4% coloured and 19.6% white children, of whom 60% were resident in the Cape Town metropolitan area and 40% in the rural area of the WC. Of 246 children with ALL diagnosed in the period 1983 - 1999, 144 were male and 102 female. The ASIR in coloured children aged 0 - 4 years was 17.1/10(6) in the rural and 30.5/10(6) in the metropolitan area, compared with 55.7/10(6) and 56.2/10(6) respectively in white children. In the 5 - 9-year age group the ASIR in coloured children was 10.0/10(6) in the rural and 16.6/10(6) in the metropolitan area compared with 27.6/10(6) and 26.7/10(6) respectively in white children. The 10 - 12-year age group had comparable incidence rates in both populations and geographical areas. Only one case occurred within a 20 km radius of the Koeberg nuclear reactor. CONCLUSIONS: White children have an ASIR for ALL comparable to rates of diagnosis in the USA, while only half as many coloured children aged 0 - 9 years were diagnosed in both the rural and metropolitan areas. This contrast may indicate significant underdiagnosis of ALL in coloured children over the period in question. The change in health policy since 1994, which has improved access to primary health care, may improve the rate of diagnosis among coloured and black children. PMID- 15285456 TI - Profile of children with head injuries treated at the trauma unit of Red Cross War Memorial Children's Hospital, 1991 - 2001. AB - OBJECTIVE: To describe the profile of childhood head injury patients treated in a trauma unit. DESIGN: A retrospective record-based study. SETTING: The trauma unit of the Red Cross War Memorial Children's Hospital. SUBJECTS: Children (under 13 years of age) presenting with head injuries between January 1991 and December 2001. RESULTS: Of the almost 94 000 records, more than one-third were children presenting with head injuries. Fifty-nine per cent were boys, with more than half the sample under 5 years of age. The majority of children presented with superficial lacerations and abrasions, mostly affecting the scalp and skull. Injuries were mainly caused by falls from a variety of heights, and traffic related injuries. Almost two-thirds of traffic-related injuries involved children as pedestrians being struck by a motor vehicle. More than 60% of injuries occurred in or around the child's own home. CONCLUSIONS: Head injuries in children are a significant cause of morbidity. Prevention, especially in the home and on the streets, needs urgent attention. PMID- 15285455 TI - Marital status and risk of HIV infection in South Africa. AB - OBJECTIVE: Available evidence on the relationship between marital status and HIV is contradictory. The objective of this study was to determine HIV prevalence among married people and to identify potential risk factors for HIV infection related to marital status in South Africa. METHODS: A multistage probability sample involving 6 090 male and female respondents, aged 15 years or older was selected. The sample was representative of the South African population by age, race, province and type of living area, e.g. urban formal, urban informal, etc. Oral fluid specimens were collected to determine HIV status. A detailed questionnaire eliciting information on socio-demographic, sex behaviour and biomedical factors was administered through face-to-face interviews from May to September 2002. RESULTS: HIV prevalence among married people was 10.5% compared with 15.7% among unmarried people (p-value < 0.001). The risk of HIV infection did not differ significantly between married and unmarried people (odds ratio (OR) = 0.85, 95% confidence interval (CI): 0.71 - 1.02) when age, sex, socio economic status, race, type of locality, and diagnosis of a sexually transmitted infection (STI) were included in the logistical regression model. However, the risk of HIV infection remained significantly high among unmarried compared with married people when only sex behaviour factors were controlled for in the model (OR 0.55; 95% CI: 0.47 - 0.66). CONCLUSIONS: The relationship between marital status and HIV is complex. The risk depends on various demographic factors and sex behaviour practices. Increased prevention strategies that take socio-cultural context into account are needed for married people. PMID- 15285457 TI - Estimation of the diagnostic accuracy of organ electrodermal diagnostics. AB - OBJECTIVE: To estimate the diagnostic accuracy and the scope of utilisation of a new bio-electronic method of organ diagnostics. DESIGN: Double-blind comparative study of the diagnostic results obtained using organ electrodermal diagnostics (OED), with clinical diagnosis as the criterion standard. SETTING: Department of Surgery, Helen Joseph Hospital, Johannesburg. PATIENTS: Two hundred pre-selected inpatients of mean age 38 years (standard deviation 9 years) with suspected pathology of one (or more) of the following organs: oesophagus, stomach, gallbladder, pancreas, colon, kidneys, urinary bladder and prostate. In total, 714 of the abovementioned internal organs were selected for statistical consideration. MAIN OUTCOME MEASURES: The degree of rectification of the measuring current once the resistance 'breakthrough effect' has been induced in the skin, as well as the difference in impedance measured at organ projection areas (OPAs) (skin zones corresponding to particular internal organs). RESULTS: In total, 630 true OED results were obtained from the 714 subjects considered, with a detection rate of 88.2% (95% confidence interval (CI): 85.6 - 90.5%). Established OED sensitivity was 89.5% (CI: 85.2 - 92.8%) and OED specificity equalled 87.5% (CI: 84.0 - 90.4%). The predictive value for positive OED results was 81.7% (CI: 76.9 - 85.9%) and for negative OED results 93.0% (CI 90.1 - 95.2%). Healthy organs usually produced the OED result 'healthy' or 'within normal limits', while subacute pathology displayed 'subcute' and acute pathology 'acute'. The OED results were not affected by either the type or the aetiology of disease, i.e. OED estimated the actual extent of pathological process activity within particular organs but did not directly explain the cause of pathology. CONCLUSIONS: So-called OPAs do exist on the skin surface. Pathology of a particular organ causes a related OPA to rectify electrical currents once the resistance 'breakthrough effect' has been induced in the skin. Pathology of an internal organ also increases the impedance of the corresponding OPA. The degree of rectification or difference in impedance is proportional to the extent of the pathological process within this organ. OED which utilises the abovementioned electrical phenomena of the skin, is a reliable bio-electronic method of non invasive medical diagnostics, with high rates of sensitivity, specificity and predictive values. OED may be used to detect diseased organs and estimate the extent of pathological process activity. PMID- 15285458 TI - C F L Leipoldt (1880 - 1947)--journalist doctor. PMID- 15285459 TI - [Current views on allergic diseases]. AB - Allergic diseases are an increasing health problem in the industrialised and developed countries especially in children and young adult persons. They are considered diseases of modern civilisation. The reported cumulative prevalence of allergic diseases in childhood of 25-30% includes allergic rhinoconjunctivitis, asthma and dermatitis. The reasons for this increasing prevalence are unknown. The main risk factors are genetic predisposition, allergen exposure, environmental pollutants, decreased stimulation of immune system during the critical period of development and lifestyle. Allergic diseases must be treated as common health disorder. They can express themselves in any age groups and in many different organs. IgE antibody is the main connection between involved organs. Specific IgE is still being identified using the 100 years old skin testing method and quantitative immunoenzymatic method in serum. In spite of the permanent improvement of both methods, neither skin reactivity to allergens nor measurable specific IgE necessarily mean a clinically manifested disease. The interpretation of these findings is still in the clinician's domain. Allergic diseases rarely have a fatal outcome, but have a long duration. They need a complex treatment and are a substantial individual and public socio-economic burden. Studies of factors influencing the ontogeny and maturation of the immune system in the early human development as well as studies of interaction between environment and genetic predisposition will provide a new insight in the aetiology of allergic diseases. This rewiev presents curent views on the epidemiology, pathogenesis, diagnosis, therapy and prevention of allergic diseases. PMID- 15285461 TI - Quality assurance and standardisation of patch tests for contact allergens skin testing. AB - The misbranding of raw materials used in the manufacturing of patch tests and the counterfeiting of the final product of these test materials have gained increasing importance in the last decade with respect to the production process in the pharmaceutical industry. To guarantee the performance and safety of a semi solid pharmaceutical testing form, it is important to make oneself acquainted with the different quality standards and official requirements demanded by the national and international regulations. Moreover it will be necessary to get an idea about the major factors which influence the quality of patch tests and therefore finally the quality of the medical allergic diagnosis. PMID- 15285460 TI - [ARIA--one airway, one disease: what links our research to the concept?]. AB - ARIA (Allergic Rhinitis and its Impact on Asthma) is a public health initiative of the World Health Organization (WHO) based on the "one airway, one disease" concept. The trigger for the initiative was a growing evidence (from epidemiology, anatomy, pathophysiology, clinical diagnostics, therapy) of a pandemic increase in the prevalence of allergic disorders of the respiratory tract (allergic rhinitis and asthma), of the deficiency of primary and secondary preventive measures, and of incomplete therapeutic control over these conditions using current management strategies. Evidence gathered so far suggest a strong link between allergic rhinitis and asthma (anatomical and histological features, aetiology, similar pathophysiological mechanisms, related therapeutic results). Therefore, ARIA as a public health initiative aims at uniting the management of these, till now separated disorders. This review discusses the evidence on which the "one airway, one disease" concept and ARIA are based, including a contribution made by scientists from the Institute of Medical Research and Occupational Health, Zagreb, Croatia. PMID- 15285462 TI - Hypersensitivity to pollen allergens on the Adriatic coast. AB - This paper describes a study of air concentrations of pollens and a calendar of pollination around the town of Split on the Croatian Adriatic in 1994. High pollen concentrations of Parietaria officinalis dominated during the year (up to 20% from April to June) followed by the pollens of Pistacia lentiscus, Olea europaea, Pinus halepensis, Juniperus oxycedrus, Acacia baileyana, Artemisia vulgaris, Ambrosia elatior and Cistus monspeliensis. In 1994-95, skin prick tests using commercially available standard inhalation allergens and specially prepared pollen allergens were performed on 3,500 patients with allergic respiratory symptoms. About 30% were allergic to standard pollen allergens (mixed grass pollen, mixed tree pollen, Parietaria officinalis and Pittosporum tobira). Hypersensitivity to more than one allergen was found in 45% of patients, whereas 15% did not react to any of the standard allergens. Additional testing with newly prepared individual allergens (P. lentiscus, O. europaea, P. halepensis, A. baileyana, C. monspeliensis, A. vulgaris, A. elatior) revealed hypersensitivity in a number of patients, but 36% showed no reaction. This finding suggests that further studies of this kind are needed for additional identification, isolation, and characterisation of pollen allergens that are present in the Adriatic coast. PMID- 15285463 TI - [Ragweed hypersensitivity in Osijek-Baranja region]. AB - Hypersensitivity of the respiratory system to pollens depends on the dominant kind of pollen grains in the air. It manifests as seasonal allergic rhinoconjunctivitis and asthma. The appearance and the severity linearly correlate with the pollen concentration in the air. In the last few years an increase in the number of people hypersensitive to pollen has been observed in the Osijek-Baranja region. It correlates with the expansion of ragweed and the increase of its air pollen concentrations. Since 2001, pollen and spores are being monitored in Osijek. This project has been initiated by the Association for the Prevention of Allergic Diseases and has been carried out in cooperation with the Regional Office of Public Health. Measurements are done using the Burkard volumetric apparatus according to the predetermined standard and are presented in pollen/m3 of air. Ragweed pollen is found in the air from the beginning of August until the end of September, with the maximum contentrations from the end of August until the middle of September. PMID- 15285464 TI - [Frequencies of airborne moulds in Zagreb]. AB - Airborne fungi are sometimes associated with several respiratory diseases and allergies. This paper describes a study of qualitative and quantitative variations in the occurrence of airborne moulds in Zagreb area on three locations: centre of the city (C), Pharmaceutical Botanical Garden "Fran Kusan" (BG) and the mountain Medvednica (M) during autumn, winter, spring and summer 2002-03. Lower concentrations of airborne moulds were found in all three locations in autumn (up to 76.88 CFU/m3) and winter (31.46 CFU/m3), with significantly higher levels in C and BG than in M (P<0.001). In spring and summer, these concentrations were much higher in all sampling sites and were significantly higher in C (160.00 CFU/m3) and BG (134.00 CFU/m3) in spring than in M (90.07 CFU/m3) (P<0.001). In summer, significantly higher concentration was found in C (237.5 CFU/m3) than in BG (186.50 CFU/m3) (P<0.01), while concentrations in C and M (216.70 CFU/m3) were similar. Airspora belonging to 29 fungal genera were identified, and allergologicaly significant moulds, Cladosporium (up to 79.5%) and Alternaria (up to 59.4%) dominated in all sampling sites. Penicillium, Fusarium and Aspergillus were also constant fungal entities (43.0-70.5%), but in much lower concentrations than Cladosporium and Alternaria. Airsporas of Cladosporium and Alternaria were more frequent in spring and summer in all locations, with significantly higher concentrations in C and BG (P<0.05). The risk from allergies increases with higher airspora concentrations in spring and summer due to an increase in Cladosporium and Alternaria. PMID- 15285465 TI - [Identification of Langerhans cells in dermatology]. AB - This paper describes our own findings on the role of Langerhans' cells in dermatology and discusses literature data on their detection in seven different dermatoses. The skin is an integral part of immune system. During the past 30 years, increasing evidence has been accumulated that the skin contains cellular elements which are needed for the initiation and expression of immune response. Langerhans' cells (LCs) are dendritic cells originating in the bone marrow. They reside mainly within stratified squamous epithelia and constitute approximately 2 4% of epithelial cells. LCs are epidermal antigen presenting cells which play a crucial role in allergic contact hypersensitivity, viral diseases, graft versus host disease and elimination of neo-plastic cell clones. They express antigens conjugated with major histocompatibility complex (MHC) class II positive molecules on their surfaces for presentation to T-helper lymphocytes. LCs cannot be identified in routinely prepared histologic testing but can be visualised at the light microscope level by histochemical and immunologic techniques. Appropriate methods for the detection of Langerhans' cells in dermatology (also shown by our own experience) are histoenzymatic methods of adenosintriphosphatase (ATP-ase), acid phosphatase (AP), alpha-naphthylacetatesterase (ANAE and peroxidase-antiperoxidase immunohistochemistry method with polyclonal S-100 protein antibody (PAP). LCs are the only cells in normal skin with ATP-ase activity. Histoenzymatic methods used in patients with atopic dermatitis, vitiligo, mycosis fungoides, Behcet's disease, lichen ruber planus, psoriasis vulgaris, irritant dermatitis and allergic contact dermatitis demonstrated LSs in epidermis and dermis. ANAE and AP showed concordance and were suitable histochemical markers for LC distribution and macrophages in the dermis in mycosis fungoides, atopic dermatitis, psoriasis vulgaris, irritant chronic dermatitis and Bechet's disease. Our experience of the human skin showed a strong activity of calcium-activated adenosine triphosphatase in LCs. LCs in the guinea pig skin can be demonstrated by Mg++ and Ca++ activated adenosine triphosphatase, but a stronger activity of Ca++ activated adenosine triphosphatase in LCs after irritation. Ca++ ATP-ase as an indicator of energy-dependent pump is the reflection of intracellular calcium level, which is a significant factor for regulating the growth and metabolism of the cells. LCs are found as target cells during the efferent phase of contact allergic reaction. Immunohistochemical methods, define the role of LCs in dermatology more precisely and allow complete immunologic recognition within the epidermis. PMID- 15285466 TI - Endotoxin measurement in house dust using the end-point Limulus amoebocyte lysate method. AB - Endotoxin is a lipopolysaccharide, a part of gram-negative bacteria cell membrane commonly present in general and many occupational environments. This paper describes sample preparation and endotoxin measurement in 16 samples of house dust from urban homes (Zagreb, Croatia) using end-point chromogenic Limulus amoebocyte lysate (LAL) bioassay. House dust was collected on cellulose filters by vacuuming bedroom and living room floors, and was kept frozen until assayed. Samples were extracted from filters with a 0.05% solution of Tween-20 in endotoxin-free water. Serial dilutions of samples were measured in duplicates. The linearity of the standard curve was satisfying (r=0.983), as well as the recovery (92 and 110%) and repeatability (coefficient of variation from 0 to 8.5%). The endotoxin levels found in the house dust samples ranged from 4.8 to 200 EU/mg, with the arithmetic mean of 49.5 EU/mg (standard error of the mean of 12.1 EU/mg), and were in the range of house dust endotoxin values obtained by other authors. PMID- 15285467 TI - [Exposure and allergy to dust mites in general and working environment in Croatia]. AB - This paper gives a review of the most important impacts of exposure to dust mites in general and working environment on human health. The Institute for Medical Research and Occupational Health in Zagreb, Croatia, has been investigating the frequency and exposure levels of allergies to pyroglyphid and non-pyroglyphid mites in Croatia for the last 10 years. Investigations were performed in general urban and rural population from the inland and coastal Croatia, and several industrial inland populations occupationally exposed to organic dusts. Mite species and levels of pyroglyphid mites allergens (Der p 1, Der f 1) were established in house dust samples taken from the floors of bedrooms and living rooms and in several industrial dust samples. The frequency of allergies to pyroglyphid mites in general urban population of inland Croatia is about 20%, with significant general indoor exposure to these mites (median value for Der p 1: 0.85 microg/g of dust). General adult population of the coastal region had a significantly higher exposure to pyroglyphid mites (median value for Der p 1: 4.5 microg/g of dust), yet showed a significantly lower frequency of allergies to these mites (about 5%). New studies are necessary to investigate possible genetic and environmental factors involved in the mechanisms which protect coastal population from the development of mite allergy. Acarological and statistical analyses have shown that the high frequency of sensitisation to non-pyroglyphid mites found in the general and working populations of the inland region is not related to environmental exposure to these mites, but to the cross-reactivity between pyroglyphid and non-pyroglyphid mites and to false positive skin reactions in prick testing, particularly to T. putrescentiae. So far, results do not indicate that pyroglyphid mites are occupational allergens in paper recycling, fish-processing and tobacco-processing. PMID- 15285468 TI - Allergy to endoprostheses. AB - This paper describes a study of hypersensitivity and tissue reaction to loosened metal implants in 40 patients who underwent repeated hip arthroplasty. Metal sensitivity was tested using a standard cutaneous patch test. Nine patients were positive for chromium, nickel, cobalt, metal rust, endoprosthesis scrapings or combinations of these allergens. Patients with positive or negative patch test did not differ in terms of their age, sex, primary diagnosis, number of endoprosthesis revisions, length of implant function, presence of other metal parts around the implant, circulating immunocomplexes, and histological appearance of the tissue around the implant. In conclusion, stainless steel endoprostheses are safe for revised hip arthroplasty, and hypersensitivity to metals probably does not play a significant role in the loosening of endoprosthesis. PMID- 15285469 TI - Exercise and allergic diseases. AB - The aim of this study was to compare exercise-induced bronchial reaction between healthy control subjects and subjects with allergic rhinitis (AR) and allergic asthma (AA). It included 16 controls, 16 subjects with AR and 19 subjects with AA. A skin prick test, pulmonary function test, histamine challenge test and exercise challenge test (ECT) were performed in all subjects. Bronchial reaction to exercise was expressed as the fall index FEV1 (%), AUC(0-30) (min x %), and fall index FEF(25-75) (%). After ECT, subjects with AA had a significantly greater bronchial reaction to exercise than subjects with AR and controls (respective fall index FEV1 8.4, 2.9, and 2.4%, P=0.0083; AUC(0-30) 127.7, 29.6, and 33.1 min x %, P=0.025; and fall index FEF(25-75) 14.6, 0.06, and 1.9%, P<0.001). No difference was found between subjects with AR and controls. In conclusion, ECT induced a significantly greater bronchial reaction in patients with AA and bronchial hyperreactivity to histamine than in patients with AR and bronchial normoreactivity to histamine and controls. This difference was not found between subjects with AR and controls. PMID- 15285470 TI - [Stress and allergy]. AB - Stress is one of the components in the complex interaction of environmental, genetic, physiological, psychological, behavioural and social factors that can influence the body's ability to remain healthy or become healthy, to resist or overcome a disease. Stress can alter neuroendocrine and immune mechanisms of health and disease through various psychosocial processes. In addition, it can affect health through the impact on health-impairing behaviours and on compliance with medical regimens. At the same time, the relationship between stress and health is not unidirectional but bi-directional. Current views on the relation between stress and allergy vary from the denial of any relationship that could fundamentally help in allergy treatment to the widespread opinion that psychological stress can exacerbate some skin symptoms and precipitate asthma. The role of stress in the genesis, incidence and symptomatology of allergy still remains a controversial issue since the mechanisms of that relationship are not well understood. Starting from the biopsychosocial model of disease, we introduced the Social Readjustment Rating Scale which measures stressful life events, and the WHOQOL-BREF which measures subjective quality of life, into an extensive multidisciplinary study of immunotoxic effects of indoor bioaerosols and lifestyle. This paper describes the characteristics of those two questionnaires and discusses the relationship between stress and various domains of the quality of life. The Social Readjustment Rating Scale proved to be a reliable predictor for quality of life in the domains of physical health and environment. Future analyses will examine the role of stress and subjective quality of life in allergy. PMID- 15285471 TI - Organic aerosols and the development of allergic disorders. AB - The aim of this study was to investigate skin reactivity to organic dust extracts and total serum IgE and their relation to the prevalence of respiratory symptoms and ventilatory capacity in workers occupationally exposed to organic aerosols. It included workers employed in processing coffee, tea, dried fruits, spices, animal food, soy, hemp, cotton, swine farmers, and control groups of workers non exposed to organic dust. All underwent a skin prick test (SPT) with water extracts of organic dust 1:10 w/v, Dermatophagoides pteronyssinus, mixed moulds, bacteria, histamin solution (1 mg/ml) and buffer solution. SPT was considered positive if the diameter of the observed wheal was 3 mm greater than that of buffer solution. The total IgE was measured by the PRIST method (Pharmacia Diagnostics AB, Upsala) and the values > 125 kU/L were considered increased. Data on respiratory symptoms were collected by standardized questionnaire. Ventilatory capacity was measured by recording MEFV curve. Airborne industrial dust were measured as total and respirable fraction. The exposed workers had a greater prevalence of positive SPT to organic dust extracts, except in soy processing. Increased IgE was found in workers processing coffee, tea, hemp, cotton and animal food, compared to non-exposed workers (P<0.05). Workers with positive SPT had a significantly higher total IgE. As there was no correlation between acute and chronic changes in ventilatory function, positive SPT, and level of total IgE, our findings could not predict objective respiratory impairment. PMID- 15285473 TI - Effective patient grievance policy can be vital tool for improvement. AB - Even if it were not required, quality managers would implement patient grievance policy. Hurt, frightened patients may be more likely to complain. Know the difference between a complaint and a grievance. PMID- 15285472 TI - Allergic diseases in relationship with environmental factors in a population of school children in Zagreb, Croatia. AB - Most scientists believe that increasing number of people with allergic diseases may be connected with some aspects of the "Western lifestyle". This paper discusses data obtained from questionnaires originally designed by the International Study of Asthma and Allergies in Childhood (ISAAC) Steering Committee concerning exposure to different environmental factors. The study included 1047 children. Allergic and non-allergic groups showed statistically significant differences in the attendance of kindergarten, vaccination against pertussis, pertussis infection, and parasite infestation. Exposure to tobacco smoke during pregnancy and exposure to dampness and moulds also entailed a risk for allergy. We speculate that changing some conditions, such as use of carpets and use of feather pillows, were connected with the expression of allergic diseases. Some correlations were consistent with earlier observations of other authors, while others differed and need further confirmation on a larger sample. PMID- 15285474 TI - Second time's the charm for open-visiting effort. AB - First attempt at instituting open visiting lacked adequate planning and defined processes. Including, not excluding, family from the intensive care unit is a growing trend. The two-month pilot ultimately led to adoption of a new policy. PMID- 15285475 TI - National model begun at regional level. AB - Improvements were seen both in self-care efforts and clinical numbers. The model has been copied by other states and quality improvement organizations. The program is based on evidence of how adults learn best. PMID- 15285476 TI - Collaborative starts 2nd phase of connectivity push. AB - The Robert Wood Johnson Foundation gives financial boost to next phase of work. Incremental road map needed to prioritize actions and foster innovation. Demonstration project is planned to test products in real-world settings. PMID- 15285477 TI - Quality gap found between award winners and others. PMID- 15285478 TI - The critical path to medical innovation. AB - In the United States,the Food and Drug Administration (FDA) and the National Institutes of Health have noted a significant decrease in the number of innovative products reaching patients, in spite of a rapidly expanding basis of fundamental scientific knowledge. A recent FDA report proposes some solutions to this problem. PMID- 15285479 TI - Brand: a company's most valuable asset. AB - Brands have the potential to outlive all other forms of intellectual property. Awareness of the issues will maximise their value. PMID- 15285480 TI - Assessing breathing-system filters. AB - An assessment of 104 different breathing-system filters has been published by the United Kingdom's Medicines and Healthcare Products Regulatory Agency. Filtration performance was measured by determining the penetration of sodium chloride particles. Wide variations in performance were found. This rapid, easy assessment technique could be used to test each filter during manufacture. PMID- 15285481 TI - The important role of material and chemical characterisation in device evaluation. AB - Parts 18 and 19 of ISO 10993, Biological Evaluation of Medical Devices, currently in draft, are receiving even more emphasis as they become an integral part of the biological safety evaluation of biomaterials and medical devices. An important step in this process is the characterisation of the material and identification of chemicals that can migrate or extract from the polymer components. This article discusses which tests will meet the requirements. PMID- 15285482 TI - Incorporating microporous membrane into medical devices. AB - A range of factors needs to be considered when choosing a microporous membrane for a medical device application. In addition to the composition and performance characteristics of the membrane, engineers should also evaluate potential suppliers for their quality systems, support capabilities, general financial strength and commitment to manufacturing the product. This article provides a checklist of considerations. PMID- 15285483 TI - Micro- and nanotechnology fabrication processes for metals. AB - Many medical products are taking advantage of the developments in micro-and nanotechnology. Fabrication processes for metal products are reviewed here. Those featured include stents, surgical blades and needles, wound dressings and fluorescent labels for diagnostic testing. PMID- 15285484 TI - Novel approaches to innovation in medical device manufacture. AB - Innovation is an imperative for the industry. This article explores new ways for companies to fill deficiencies in their innovation capabilities and to overcome the problems posed by globalisation. PMID- 15285485 TI - The essential element of ethics. AB - Companies must operate in a cost-effective manner in an increasingly competitive business environment. At the same time, their products must meet patient and user needs, be safe, perform as labelled and meet regulatory requirements. This article discusses three codes of practice developed to assist companies in balancing these sometimes opposing forces. PMID- 15285486 TI - EU and US clinical investigation adverse event reporting. AB - Revised international standards on clinical investigations were recently published. ISO 14155 consists of two parts under the general title Clinical Investigation of Medical Devices for Human Subjects. This article outlines the requirements of the revised standard and contrasts these with United States reporting requirements. PMID- 15285487 TI - Latest medical applications of polypropylene. AB - PP volumes for use in medical applications increase every year for some obvious reasons. The polymer can be processed by practically all techniques and sterilisation and transparency processes, and its properties are continuously improved. Furthermore, it can replace many other materials, and is attractive from a cost and versatility perspective. PMID- 15285488 TI - Systems for research and development in medical ultrasound imaging. AB - The system requirements of engineers conducting research and development into new ultrasonic methods for medical applications are reported. The design criteria for these systems are explained together with a review of what is currently available. PMID- 15285489 TI - Germany: financing innovation is the future challenge. AB - Changes are required to encourage innovation in the industry. Some ways forward are described. PMID- 15285490 TI - Opting for clean manufacturing. PMID- 15285491 TI - Genetic variation in body weight and egg production in an experimental line selected long term for increased egg production, a commercial dam line, and reciprocal crosses between lines. AB - A line (E) of turkeys selected long term (40 generations) for increased egg production was reciprocally crossed with a commercial dam line (BD). The BD line was larger and had better conformation than the E line, but the E line laid more eggs than the BD line. Heterosis was negative and significant for BW at 8, 16, and 20 wk of age (both sexes) and at 50% production (females only). Values for heterosis ranged from -3.1 to -5.5%. Negative heterosis was observed for breast width at 16 wk of age (10.8 and 3.0% for males and females, respectively). Positive heterosis values of 9.2, 10.2, and 11.3% were observed for egg production when based on 84, 120, and 180 d, respectively. Heterosis (-8.2%) was significant for rate of response to stimulatory lighting of 14 h light per day (days from stimulatory lighting to production of first egg). Based on data for a 180-d production period, heterosis was observed in average clutch length (12.6%) and rate of lay [number of eggs laid: (180 - total days broody)] (8.2%) but not in total days lost from broodiness or the effective length of the laying period (180 minus days lost in periods of 5 or more consecutive days at the end of the laying period). The present results along with those in the literature suggest that more heterosis in egg production traits might be expected in crosses of lines exhibiting extreme differences in BW and body conformation. The negative heterosis in BW in the BD and E reciprocal crosses reduces the commercial importance of such a cross to improve egg production. PMID- 15285492 TI - Inheritance of breast muscle morphology in turkeys at sixteen weeks of age. AB - The inheritance of morphology of the pectoralis major muscle in turkeys at 16 wk of age was studied in a randombred control line (RBC2), a subline (F) of RBC2 selected long term for increased 16 wk BW only, and F1 and F2 crosses of the F and RBC2 lines. Samples of pectoralis major muscle were obtained from 10 males and 10 females of each genetic group in a manner to avoid muscle contraction. After being fixed and cross sectioned, the muscle samples were stained with hematoxylin and eosin to view muscle morphology. The stained sections were analyzed for muscle fiber width, number of fibers in a 136-microm2 area, and extracellular matrix perimysial (PW) and endomysial (EW) width in areas of sections in which accurate measurements could be made. Because muscle damage was evident in some sections and, therefore, morphological measurements might not have provided a complete overview of muscle morphology, sections of the F2 crosses were subjectively rated by 4 people. The ratings ranged from 1 (little extracellular matrix and indistinct muscle fibers) to 5 (large extracellular space and distinct muscle fibers). Ratings of 2 to 4 were intermediate to these extremes. Creatine kinase concentrations of blood samples taken immediately prior to collecting muscle tissue were obtained and correlated with muscle section ratings within genetic group and sex. The F and RBC2 lines differed in PW and EW but not in individual fiber measurements. In the F1 generation, heterosis was 10.4% (P < or = 0.01), 19.7% (P < or = 0.05), -25.2% (P < or = 0.01), and -34.3% (P < or = 0.01), respectively, for fiber width, number of muscle fibers, PW, and EW. The F2 crosses differed only in EW based on measurements of sections in which accurate measurements could be made. However, based on subjective ratings of the muscle sections, possible maternal inheritance was suggested, as the orthogonal contrast was significant (P < or = 0.01) for crosses with F dams as F1 parents vs. those with RBC2 dams as F1 parents, confirming a previous study. The correlation coefficient between creatine kinase concentration and muscle section ratings was -0.282 (P < or = 0.01) after adjustment for line and sex effects. PMID- 15285493 TI - A comparison of growth and development patterns in diverse genotypes of broilers. 1. Male broiler growth. AB - Selection for breast muscle yield and BW in commercial broilers has resulted in genotypes far different from broilers processed in the past. When comparative studies with commercial lines are conducted, it is often difficult to differentiate between carcass effects resulting from direct genetic selection vs. correlated effects that partially reflect genetic changes in BW. The objective of the present experiment was to compare growth and development characteristics of male broilers from commercial lines exhibiting similar rates of BW gain based on a percentage of 8-wk BW but exhibiting different carcass traits. Male broilers from 2 commercial genotypes exhibiting increased breast muscle yield (A and B) were compared with broilers from a commercial line that was "unimproved" with respect to conformation (C). All birds were fed a commercial-type broiler starter diet throughout the study and were processed at 8 wk of age for carcass comparisons. No differences were observed among lines for breast-free BW or weight of the abdominal fat pad. The absolute and relative weights of pectoralis major and pectoralis minor breast muscles were heavier in lines A and B than C (P < 0.001). Length (C > A > B; P < 0.001), width (B > C >A; P < 0.001), and depth (A = B > C; P < 0.001) of the pectoralis major were different between lines. Drum and thigh weights were heavier in lines A and B than C (P < 0.001). There were genotype differences in tibia length (C > A = B, P < 0.001) and femur length (C > A = B, P < 0.001). The comparison of line C with selected lines A and B, suggested that line C may provide a useful model for studying carcass development between commercial genotypes with similar growth patterns. PMID- 15285494 TI - A comparison of growth and development patterns in diverse genotypes of broilers. 2. Pullet growth. AB - Genetic selection within commercial broiler lines continues to generate improvements in BW, feed conversion, and breast meat yield. The objective of the current experiment was to compare carcass and reproductive characteristics of broiler breeder pullets from 2 dam lines that produce heavy broilers with increased breast yield (A and B) with pullets from a commercial line that does not have the extremes in breast yield (C). Restricted-fed BW were similar at all ages of the experiment in the 3 genotypes. All pullets were photostimulated at 23 wk of age, and carcass and reproductive tract measurements were made at 27 wk of age. There were no differences in BW among the lines at 27 wk of age, but the weights of the pectoralis major and minor breast muscles were heavier in lines A and B compared with line C (P < 0.001). The weight of the abdominal fad pad, however, was heavier in line C (P < 0.04). There was no difference in total drum weight or total thigh weight among lines A, B, or C. Tibia length and tibia width were similar in lines A and C, and the measurements were larger in these lines than in line B (P < 0.02). Femur length was longer (P < 0.001) in line C than in lines A and B, whereas femur width was greater in line C than in line A (P < 0.001) but similar to line B (P < 0.001). Oviduct weight was greater in line C compared with lines A and B (P < 0.004), but there were no differences in total ovarian weight, follicle number, or follicle weight. The information gathered in the present experiment suggested that line C may prove to be useful for reproductive comparisons with commercial lines exhibiting significant differences in carcass traits. PMID- 15285495 TI - Genetic and phenotypic correlations between feather pecking behavior, stress response, immune response, and egg quality traits in laying hens. AB - The objective of the current study was to estimate genetic and phenotypic correlations among feather pecking (FP) behavior and stress response, immune response, and egg quality parameters. These traits have been measured in an F2 cross, coming from a cross between a high and a low FP line of laying hens. Heritabilities (h2) of stress response (32 wk), primary immune response to keyhole limpet hemocyanin (KLH) (36 wk) and Mycobacterium butyricum (39 wk), and egg quality parameters (35, 44, and 50 wk of age) were calculated. The h2 was 0.05 +/- 0.05 (SE) for stress response, 0.15 +/- 0.07 for antibody response to KLH, and 0.08 +/- 0.06 for antibody response to M. butyricum. The h2 for egg quality traits were in the range of 0.12 to 0.30. Significant phenotypic correlations were found between gentle FP in adult hens and stress response, egg weight at 44 and 50 wk, and egg deformation at 50 wk. Significant additive genetic correlations were found between severe FP in adult hens and antibody response to KLH (0.79 +/- 0.35), and between ground pecking in adult hens and egg deformation at 50 wk (0.63 +/- 0.26), and between ground pecking and eggshell strength at 35, 44, and 50 wk of age (-0.86 +/- 0.29, -0.81 +/- 0.20, -0.76 +/- 0.24, respectively). PMID- 15285496 TI - Modeling of parameters affecting phytate phosphorus bioavailability in growing birds. AB - The current study was undertaken to establish a population from an unselected random-mating chicken population for the development of a model to predict factors that affect phytate P utilization in growing birds. A population was established from a mating of 40 male and 200 female chickens from the Athens Canadian randombred population. At 4 wk of age, birds were housed in individual metabolic cages and fed a diet containing 1.06% Ca, 0.35% total P, and 0.03% available P. After 3 d of acclimatization, feed consumption (FC) was measured and excreta produced in 3 consecutive d were collected. Individual 4-wk BW, BW gain (BWG), phytate P intake (PPI), inorganic P intake (IPI), Ca intake (CaI), N intake (NI), and energy intake (EI) during the 3 d excreta collection period were also measured. Feed conversion ratios (FCR) and relative growth rate were calculated. Phytate P bioavailability (PPB), Ca bioavailability (CaB), and N bioavailability (NB) were estimated from the disappearance of the nutrients during the passage of feed through the gastrointestinal tract. Energy bioavailability (EB) was measured by bomb calorimetry as the difference in the gross energy of the feed and the energy of the excreta. The major factors affecting PPB were CaB and EB for both sexes. In the males, BW contributed significantly to PPB. However, in the females, NB also contributed significantly to PPB. Faster growing birds tended to have a reduced retention time of feed compared with slow growing birds, and as a result utilized phytate P less. Birds that are able to utilize phytate P better are putatively able to release P for energy utilization. Therefore, the birds that were able to utilize phytate P better were also better energy utilizers. PMID- 15285497 TI - The influence of selection for increased body weight and sex on pectoralis major muscle weight during the embryonic and posthatch periods. AB - Skeletal muscle development and growth results from a complex series of highly organized processes. To address how myogenesis was influenced by selection for increased BW and by sex, both sexes from a turkey line (F) selected only for increased 16-wk BW and its genetic control line (RBC2) were used. Pectoralis major muscle was isolated and weighed from 15 individuals of each sex of the F and RBC2 lines at 14, 16, 18, 20, and 24 d of embryonic development and at 1, 8, 12, and 16 wk of age posthatch. The F line had significantly heavier p. major muscle weights than the RBC2 line beginning at 16 d of embryonic development, and the magnitude of the line differences generally increased with age through 16 wk posthatch The p. major muscle was consistently heavier in males than in females, but the differences between sexes were significant only at 16, 18, and 24 d of embryonic development and at 8 wk posthatch. There was no significant interaction between line and sex for weight of the p. major muscle at any age. The results indicated that selection for increased 16-wk BW in the F line altered growth of the p. major muscle by 16 d of embryonic development and changes were similar for both sexes. PMID- 15285498 TI - Microflora ecology of the chicken intestine using 16S ribosomal DNA primers. AB - The microflora in the gastrointestinal tract of broiler chickens influences digestion, health, and wellbeing. Analysis of chicken gut microflora has been mainly by culture-based methods. Studies using these techniques have been useful for identification and analysis of specific groups of bacteria, however, the use of enrichment medium precludes even relative quantitation of bacterial species. Recent advances in ribosomal DNA-based molecular techniques make it possible to identify different bacterial populations in environmental samples without cultivation. In this study, the intestinal microflora was examined using 16S ribosomal DNA (rDNA) targeted probes from bacterial DNA isolated from intestinal and cecal contents of chickens at 4, 14, and 25 d of age. The ribosomal gene sequence was amplified using PCR with universal primers to determine total bacterial DNA and specific primers directed at 6 bacterial species: Lactobacillus, Bifidobacterium, Salmonella, Campylobacter, Escherichia coli, and Clostridium. The use of universal primers extends these methods to allow determination of relative proportions of different bacterial species. The results indicated that in young chicks the major species present in the small intestines and ceca was Lactobacilli, with a Bifidobacteria population becoming more dominant in the ceca at older age. Clostridium was detected in some segments of the small intestine in young chicks. In older chickens, Salmonella, Campylobacter, and E. coli species were found in the ceca. This study has demonstrated the use of molecular techniques for determining relative proportions of bacterial species and monitoring pathogens in the chick gastrointestinal tract. PMID- 15285499 TI - Competitive exclusion of a glycopeptide-resistant Enterococcus faecium in the presence of vancomycin but not equivalent concentrations of tylosin or gentamicin. AB - The effect of subtherapeutic concentrations of antibiotics (10.0 and 40.0 microg/mL of vancomycin, gentamicin, and tylosin) on the efficacy of a mixed anaerobe culture of chicken microflora (CCF) was studied in a continuous-flow fermentation system. Efficacy of CCF posttreatment was assessed by challenge with glycopeptide-resistant Enterococcus faecium (GRE) at 6.0 log10 cfu/mL. Bacterial enumeration of endogenous CCF isolates, volatile fatty acid (VFA) analysis, and challenge with GRE indicated that CCF efficacy was affected by all antibiotic treatments. Although CCF treated with 10.0 microg/mL of vancomycin eliminated GRE13 at a rate of 0.61 log10 cfu/ mL per day, it was unable to eliminate E. coli, a gram-negative challenge organism. All other antibiotic treatments allowed GRE persistence at approximately 2.0 to 6.5 log10 cfu/mL. All antibiotic-treated cultures had decreased concentrations of acetic and propionic acids. Our data suggest that low concentrations of antimicrobials may adversely affect the microbial ecology of gut microflora with respect to its ability to exclude exogenous bacteria. Moreover, gentamicin had an adverse effect on the inhibitory stringency of CCF even though it showed little anti-anaerobic activity against CCF strict anaerobes in pure culture. Verification of the results in live animals will be necessary to determine if antimicrobial treatment could compromise the effectiveness of normal microflora to serve as a natural host defense against infection. PMID- 15285500 TI - Evaluation of culture media for detecting airborne Salmonella enteritidis collected with an electrostatic sampling device from the environment of experimentally infected laying hens. AB - Detection of Salmonella enteritidis in the environment of commercial laying hens is critical for reducing the production of contaminated eggs by infected flocks. In the present study, an inexpensive and portable electrostatic air sampling device was used to collect S. enteritidis in rooms containing experimentally infected laying hens. After hens were orally inoculated with a phage type 13a S. enteritidis strain and housed in individual cages, air samples were collected 3 times each week with electrostatic devices onto plates of 6 types of culture media (brilliant green agar, modified lysine iron agar, modified semisolid Rappaport-Vassiliadis agar, Rambach agar, XLD agar, and XLT4 agar). Air sampling plates were incubated at 37 degrees C, examined visually for presumptive identification of typical S. enteritidis colonies and then subjected to confirmatory enrichment culturing. Air samples (collected using all 6 culture media) were positive for S. enteritidis for 3 wk postinoculation. Because visual determination of the presence or absence of typical S. enteritidis colonies on air sampling plates was not consistently confirmed by enrichment culturing, the postenrichment results were used for comparing sampling strategies. The frequency of positive air sampling results using brilliant green agar (66.7% overall) was significantly greater than was obtained using most other media. A combination of several plating media (brilliant green agar, modified lysine iron agar, and XLT4 agar) allowed detection of airborne S. enteritidis at an overall frequency of 83.3% over the 3 wk of sampling. When used with appropriate culture media, electrostatic collection of airborne S. enteritidis can provide a sensitive alternative to traditional methods for detecting this pathogen in the environment of laying flocks. PMID- 15285501 TI - Comparison of four Salmonella isolation techniques in four different inoculated matrices. AB - The poultry industry is now operating under increased regulatory pressure following the introduction of the pathogen reduction and hazard analysis critical control point (HACCP) rule in 1996. This new operation scheme has greatly increased the need for on-farm food safety risk management of foodborne bacteria, such as Salmonella. Information needed to make informed food safety risk management decisions must be obtained from accurate risk assessments, which rely on the sensitivity of the isolation techniques used to identify Salmonella in the production environment. Therefore, better characterization of the Salmonella isolation and identification techniques is warranted. One new technique, immunomagnetic separation (IMS), may offer a benefit to the poultry industry, as it has been shown to be efficacious in the isolation of Salmonella from various sample matrices, including some poultry products. In this work, we compared the isolation ability of 4 Salmonella-specific protocols: IMS, tetrathionate (TT) broth, Rappaport-Vassiliadis R10 (RV) broth, and a secondary enrichment (TR) procedure. All 4 methods were compared in 4 different spiked sample matrices: Butterfield's, poultry litter, broiler crops, and carcass rinses. IMS was able to detect Salmonella at 3.66, 2.09, 3.06, and 3.97 log10 cfu/mL in Butterfield's, poultry litter, carcass rinse, and broiler crop matrices, respectively. For the broiler litter and Butterfield's solution, there were no (P > 0.05) differences among the 4 isolation protocols. However, in the carcass rinse and crop samples, there were no differences among the isolation of Salmonella using RV, TR, or TT, but all 3 were (P < or = 0.05) more successful at recovering Salmonella than the IMS method. PMID- 15285502 TI - A comprehensive screen for chicken proteins that interact with proteins unique to virulent strains of Marek's disease virus. AB - Genetic resistance to Marek's disease (MD) has been proposed as a method to augment current vaccinal control of MD. Although it is possible to identify QTL and candidate genes that are associated with MD resistance, it is necessary to integrate functional screens with linkage analysis to confirm the identity of true MD resistance genes. To help achieve this objective, a comprehensive 2 hybrid screen was conducted using genes unique to virulent Marek's disease virus (MDV) strains. Potential MDV-host protein interactions were tested by an in vitro binding assay to confirm the initial two-hybrid results. As a result, 7 new MDV chicken protein interactions were identified and included the chicken proteins MHC class II beta (BLB) and invariant (Ii) chain (CD74), growth-related translationally controlled tumor protein (TPT1), complement component Clq-binding protein (C1QBP), retinoblastoma-binding protein 4 (RBBP4), and alpha-enolase (ENO1). Mapping of the encoding chicken genes suggests that BLB, the gene for MHC class II beta chain, is a positional candidate gene. In addition, the known functions of the chicken proteins suggest mechanisms that MDV might use to evade the chicken immune system and alter host gene regulation. Taken together, our results indicate that integrated genomic methods provide a powerful strategy to gain insights on complex biological processes and yield a manageable number of genes and pathways for further characterization. PMID- 15285503 TI - Effects of mushroom and herb polysaccharides on cellular and humoral immune responses of Eimeria tenella-infected chickens. AB - We investigated the effects of polysaccharide extracts from 2 mushrooms, Lentinus edodes (LenE) and Tremella fuciformis (TreE), and an herb, Astragalus membranaceus (AstE), on cellular and humoral immune responses of Eimeria tenella infected chickens. A total of 150 broiler chicks were assigned to 5 treatment groups: 3 groups were infected with E. tenella and fed with extracts (LenE, TreE, and AstE), and 2 control groups were infected with or without E. tenella. The 3 extracts were given at the level of 1 g/kg of the diet from 8 to 14 d of age. Specific systemic and cecum mucosal antibody production, proliferation of splenocytes, and peripheral T and B lymphocytes were measured during the 3 wk following Eimeria infection. A significantly higher production of specific IgA, IgM (at d 14 and 21 postinfection), and IgG (at d 21 postinfection) were detected in the Eimeria-infected groups fed with the extracts than in the infected group not fed with the extracts. Of the 3 extracts, TreE stimulated a slightly higher production of specific IgM (P = 0.052), and a significantly higher IgG production at 21 d postinfection. The cecal antibody production showed a similar trend to that of serum antibodies. The overall mean levels of cecal-specific IgA and IgG of the groups fed with extracts were significantly higher at 14 and 21 d postinfection compared with the group not fed with extracts. Of the 3 extracts, the AstE-fed group showed the highest IgG production at d 7 postinfection. Both TreE- and LenE-fed groups had significantly higher IgM and IgG levels compared with the AstE group at d 21 postinfection. The extract-fed groups also showed a significantly higher antigen-specific proliferation of splenocytes at 14 and 21 d postinfection compared with the group not fed with the extracts. The overall mean of erythrocyte rosette-forming cells (ERFC %) (at d 14 and 21) and erythrocyte antibody-complement cells (EAC %) (at d 14) of the groups fed with the extracts was significantly higher compared with the group not fed the extracts. It is concluded from this study that supplementation with mushroom and herb extracts resulted in enhancement of both cellular and humoral immune responses in E. tenella-infected chickens. PMID- 15285504 TI - Differential effects of lipopolysaccharide and lipoteichoic acid on the primary antibody response to keyhole limpet hemocyanin of chickens selected for high or low antibody responses to sheep red blood cells. AB - Various bacterial components are potent activators of the innate immune system and probably (in)directly determine subsequent specific immune responses. Therefore, effects of i.v. administered Salmonella enteriditis-derived lipopolysaccharide (LPS) and Staphylococcus aureus-derived lipoteichoic acid (LTA), respectively, on the primary antibody (Ab) response to keyhole limpet hemocyanin (KLH) were studied in cocks from 2 lines divergently selected for high (H line) and low (L line) Ab responses to SRBC. The Ab responses to KLH were significantly affected by a line-by-treatment-by-time interaction. Significantly higher Ab titers to KLH, S. aureus LTA, and S. enteriditis LPS were found in H line birds than in the L line birds. Ab titers to KLH were enhanced if the chickens were intravenously pretreated 24 h earlier with LTA but decreased if the chickens were intravenously pretreated 24 h earlier with LPS. Ab responses to S. enteriditis LPS were significantly enhanced when birds were immunized with KLH or pretreated with S. aureus LTA. Ab responses to S. aureus LTA were also significantly enhanced when birds were immunized with KLH or pretreated with LTA and subsequently immunized with KLH. Our findings suggest that LTA and LPS have immunomodulatory features in chickens, albeit in opposite directions. In addition, KLH acted in an immunomodulatory role too. Possible mechanisms underlying our observations and the role of LTA and LPS in polarization of the specific immune response in chickens are discussed. PMID- 15285505 TI - Utilization of various carbohydrate sources as affected by age in the chick. AB - In 3 experiments, New Hampshire x Columbian male chicks were fed carbohydrate soybean meal (SBM) or casein diets from 0 to 21 d of age, and MEn was determined at 0 to 2, 3 to 4, 7, 14, and 21 d of age. Carbohydrate sources evaluated in experiment 1 were dextrose (D-glucose), conventional cornstarch, dextrinized cornstarch, corn-syrup solids, pregelatinized unmodified cornstarch, pregelatinized tapioca starch, tapioca dextrin, high-amylose starch, and polycose (mixed glucose polymers). Carbohydrate sources evaluated in experiments 2 and 3 were conventional corn, waxy corn, high-oil corn, corn flour, rice flour, dextrose, and sucrose. In experiment 1, chicks fed the dextrose diet had the highest weight gains, and the chicks fed high-amylose starch and pregelatinized unmodified cornstarch diets had the lowest weight gains. The MEn values varied among carbohydrate sources with MEn being highest for the dextrose diet and lowest for the high amylose starch diet. In experiment 2, chicks fed waxy corn, high-oil corn, or dextrose-SBM diets had (P < 0.05) higher growth rates than chicks fed conventional corn, corn flour, or rice flour. The MEn values increased with age for all diets except the dextrose-SBM, which was consistently high at all ages. In experiment 3, the dextrose diets (SBM or casein) yielded higher growth performance and MEn values than the sucrose-diets, and the differences were greater at younger ages. The MEn values were also much higher for the casein than the SBM diets, and MEn of the SBM diets increased with increasing age. The results of this study indicate that MEn, varies among carbohydrate sources and increases with age for most carbohydrate-SBM diets. PMID- 15285506 TI - Effects of dietary mannan oligosaccharide, bacitracin methylene disalicylate, or both on the live performance and intestinal microbiology of turkeys. AB - Hybrid male turkeys were fed to 18 wk of age in a completely randomized design with 10 replicate pens (18 birds each) per treatment to compare growth promoters. Four dietary treatments were used: negative control (CON), bacitracin methylene disalicyate (BMD) at 55 mg/kg to 6 wk and 27.5 mg/kg thereafter, mannan oligosaccharide (MOS) at 0.1% to 6 wk and 0.05% thereafter, and BMD and MOS at concentrations listed above. There were 3 toms/m2 (3.59 ft2/tom) on fresh pine shavings inoculated with used litter. A 6-phase feeding program was used, with crumbles the first 3 wk and pellets thereafter. At wk 6 and 18, one bird per pen was killed, and the large intestinal microbial populations, after being frozen, were enumerated (i.e., bifidobacteria, Clostridium perfringens, coliforms, enterococci, Escherichia coli, lactobacilli, and total anerobes). Body weights at wk 18 were as follows: CON, 11.87 kg; BMD, 12.46 kg; MOS: 12.56 kg; and BMD + MOS, 12.79 kg. The BMD and MOS turkeys were (P < 0.05) heavier than CON birds, and those fed the combination were significantly heavier than all other treatments. At wk 18, BMD + MOS feed conversion ratio of 2.66 was significantly lower than CON at 3.00 with BMD and MOS being intermediate (2.83 and 2.79, respectively). Mortality was not affected by treatment. The BMD and MOS each reduced large intestinal concentrations of Clostridium perfringens (log transformed data analysis) at wk 6 but not at wk 18. The BMD or MOS each improved turkey performance, and when used together, exhibited further beneficial effects. PMID- 15285507 TI - Fatty acid content in chicken thigh and breast as affected by dietary polyunsaturation level. AB - One hundred ninety-two female broiler chickens were randomly distributed into 16 experimental treatments as a result of the combination of 4 levels of dietary polyunsaturated fatty acids (PUFA) (15, 34, 45, and 61 g/kg) and 4 levels of supplementation with alphatocopheryl acetate (alpha-TA) (0, 100, 200, and 400 mg/kg), to determine the modification of the amount and type of fatty acids (FA) deposited in raw and cooked chicken tissues. At 44 d, quantified FA of thighs and breasts were not affected by dietary supplementation with alpha-TA. Total FA content of breast was less than 15% of the total FA content of thigh. However, increasing the PUFA content of the diet by 46 g, from 15 to 61 g/kg, decreased total FA of thigh 17%, but did not affect FA content in breast meat. Monounsaturated fatty acid (MUFA) and saturated fatty acid (SFA) content of thigh (y) decreased linearly as the inclusion of dietary PUFA (x) increased (MUFA: y = 89.34 - 0.92x, R2 = 0.70; SFA: y = 53.81 - 0.43x, R2 = 0.57), whereas the relationship between PUFA content of feed (x) and thighs (y) was exponential (y = 92.03 92.03e(-00155x), R2 = 0.75). A similar response was observed in breast, with less variation and more incorporation of PUFA than thigh. Cooking of thigh meat led to a reduction in total FA content that affected SFA, MUFA, and PUFA in a similar proportion. PMID- 15285508 TI - Predicting effective caloric value of nonnutritive factors: I. Pellet quality and II. Prediction of consequential formulation dead zones. AB - Two experiments were conducted with male broilers to 1) establish a methodology for predicting effective caloric value (ECV), defined as dietary caloric density (CD) necessary for broilers to achieve specific BW and feed conversion ratio (FCR) combinations under standardized conditions and 2) quantify the ECV attributable to pellet quality (PQ), defined as the pellet to pellet fines ratio in the feeder. In experiment 1, chicks were reared to 56 d on diets varying in CD. Dietary caloric densities examined ranged from 2,650 to 3,250 kcal of MEn/kg. Pen BW, feed intake, and FCR were measured at 21, 42, and 56 d. On 42 and 56 d, carcass traits were measured. Increasing CD significantly enhanced BW, energy consumption, and FCR. Feed intake remained similar across the upper 3 CD treatments to 42 d. By d 56, feed consumption tended to decline as CD increased. Increasing CD beyond 3,066 kcal of MEn/kg diet did not increase lean tissue accretion, while fat deposition rose disproportionately. Experiment 1 results enabled development of equations whereby CD, hence ECV, might be predicted using BW and FCR. In experiment 2, 38-d-old broilers were used to evaluate PQ effects on growth, feed intake, FCR, and behavior in a 7-d FCR assay. The BW gain and FCR were significantly enhanced by pelleting and were positively correlated with PQ. Feed intake was not affected by PQ. The experiment 1 model was validated for experiment 2, as it closely estimated the CD for diets of similar PQ used in experiment 1. Results suggest pelleting contributes 187 kcal/kg of diet at 100% PQ and that the ECV declines curvilinearly as PQ falls. Birds were observed eating less and resting more as PQ increased, suggesting that ECV of pelleting is mediated by energy expenditure for activity. These studies provide a method for estimating ECV of nonnutritive factors that impact BW, FCR, or both. Further, the application reveals potential for creation of formulation "dead zones" whereby dietary changes to enhance CD may be offset due to reduced ECV. PMID- 15285509 TI - Towards complete dephosphorylation and total conversion of phytates in poultry feeds. AB - The rate of phytate P removal from feed (level of dephosphorylation, DL) and the extent to which the molecule of phytic acid is deprived of phosphate moieties (conversion degree, CD) were studied in vitro and in a feeding trial with broilers fed corn-soybean diets. In the in vitro model, phytase A asymptotically increased DL and CD. Phytase B influenced DL only at low dosages of phytase A [0 or 250 phytase activity units (FTU)/kg], but it enhanced CD irrespective of phytase A activity. In the feeding trial, 3-phytase A and 6-phytase A (at 750 FTU/kg) exerted similar effects on broiler performance and similarly influenced bone mineralization, P retention, and Ca retention. Phytase B [6,400 acid phosphatase activity units (ACPU)/kg] enhanced feed intake, BW gain (BWG), toe ash, and P retention but not the retention of Ca. Myo-inositol fed at 0.1% significantly increased BWG, but it reduced P retention. Under conditions of a higher CD (excess of phytase B), 3-phytase A was more effective in enhancing performance than 6-phytase A, but it reduced Ca retention. Lower phytase B activities (0 to 3,200 ACPU/kg) with added 6-phytase A were more necessary for optimal growth of chickens than for enhanced P and Ca retention (4,800 to 6,400 ACPU/kg). The efficacy of both forms of phytase A and phytase B depended on the Ca level in feed. There is enough evidence to conclude that myo-inositol phosphates resulting from simultaneous action of 3-phytase A and phytase B affect bird physiology differently than intermediates accumulated by the action of 6 phytase A and phytase B. PMID- 15285510 TI - Phytase, citric acid, and 1alpha-hydroxycholecalciferol improve phytate phosphorus utilization in chicks fed a corn-soybean meal diet. AB - Previous research from our laboratory has shown that phytase, citric acid, and 1alpha-hydroxycholecalciferol [1alpha-(OH) D3] individually improve phytate P use in young chicks fed a P-deficient corn-soybean meal (C-SBM) diet. The current study was conducted to evaluate combinations of these additives on phytate P utilization. In 3 chick experiments, male crossbred chicks (New Hampshire x Columbian) were fed experimental diets from 8 to 21 d of age. The C-SBM basal diet used in all assays contained no supplemental P and was calculated to provide 23% CP, 0.13% nonphytate P (0.39% total P), 0.62% Ca, 25 mg of cholecalciferol/kg, and 3,260 kcal of TME/kg. In all 3 experiments, factorial arrangements (2 x 2 or 2 x 2 x 2) were used to evaluate 2 levels of phytase (0 and 300 units/kg), citric acid (0 and 3 or 4%), and 1alpha(OH) D3 (0 and 5, 10, or 15 microg/kg). Phytase, citric acid, and 1alpha-(OH) D3 each increased weight gain and tibia ash in all 3 experiments. There were some significant interactions among the compounds, but these were not consistent across experiments. Using standard curve methodology for tibia ash data, it was estimated that 0.03, 0.02, and 0.04% P were released by 3% citric acid, 300 units of phytase/kg, and 5 microg 1alpha-(OH) D3/kg, respectively, and that the combination of all 3 compounds resulted in the release of 0.13% P. Our results indicate that all 3 compounds increased phytate P use, and that their effects were generally additive, with some possible synergism between citric acid and 1alpha-(OH) D3. PMID- 15285511 TI - Proliferative and steroidogenic effects of follicle-stimulating hormone on cultured chick embryo testis cells. AB - The present study evaluated the follicle-stimulating hormone (FSH) effect on cell proliferation and steroid production by chick embryo testis. Dissociated cells from 18-d-old embryos were cultured on polycarbonate membranes in defined media. In some experiments, cells were further separated by a metrizamide gradient, and 5 cellular subpopulations were recovered and cultured. [3H]thymidine was added to the culture media. When necessary, 17beta-estradiol, human FSH (hFSH), recombinant human FSH (rhFSH), or human chorionic gonadotropin (hCG) was added to the medium at the beginning of the culture. The total number of cells and the incorporation of [3H]thymidine increased when hFSH or rhFSH was added. No changes were produced by the addition of hCG or 17beta-estradiol. The dose-response curve to hFSH resulted in an ED50 of 0.25 IU/mL. The stimulatory effect of hFSH on total number of cells and on [3H]thymidine incorporation was significant at 36 h of culture and was maintained up to 60 h. Testosterone production increased with the addition of FSH or rhFSH, meanwhile estradiol production was below the limit of detection of RIA. The hFSH proliferative effect measured as [3H]thymidine incorporation was observed only in the F3, F4, and F5 fractions of the density gradient. Present results show that hFSH and rhFSH, but not hCG or estradiol, stimulate testis cell proliferation in a time- and dose-dependent manner. The combination of [3H]thymidine incorporation and testosterone production in fractions obtained from the metrizamide density gradients suggests that the cell fractions of the chick embryo testis show a differential response to FSH. PMID- 15285512 TI - Effect of a single short-term reduction in photoperiod on photorefractoriness in turkey hens. AB - In a prior study, we reported that a high proportion of hens in a winter-laying flock became relatively photorefractory (rPR) early in the reproductive cyand that successive short-term reductions in photoperiod in such hens each initially depressed egg production but then caused a rebound in rate of lay to briefly exceed that of hens that did not exhibit rPR. The present study was conducted to assess rPR in a summer-laying flock and to determine whether a single short-term reduction in day length early in the reproductive cycle might enhance egg production and delay the onset of absolute photorefractoriness (aPR). Control hens received a photoperiod of 16L:8D throughout the experiment. Experimental hens were photostimulated with 16L:8D, received a reduced (but still stimulatory) photoperiod of 11.5L:12.5D for 2 wk beginning 8 wk after photostimulation, and then were returned to 16L:8D for the remainder of the 23-wk test period. Results showed that a single 2-wk reduction in day length shortly after the hens reached peak egg production did not significantly reduce overall flock egg production, but it also did not improve late-season egg production or retard the onset or incidence of aPR. The incidence of rPR was substantially less in this study than we had observed with a winter-laying flock (32.9 vs. 67.1%), but similar proportions of treated hens exhibited the most severe rPR response (a brief but complete cessation of egg production) in both studies (21.1 vs. 24.0%), and all treated hens that subsequently became aPR had shown this severe rPR response to the test photoperiod. We concluded that a core proportion of hens (approximately one-fifth) exhibited a strong rPR response when presented with a reduced photoperiod early in the reproductive cycle, regardless of season of the year, and that such hens were more likely to subsequently exhibit poor egg production or become aPR than flockmates that did not exhibit rPR. Therefore, some indication of the incidence of rPR early in the lay period may have a predictive value for the overall egg production of the flock. PMID- 15285513 TI - Relationship between egg productivity and insulin-like growth factor-I genotypes in Korean native Ogol chickens. AB - Endocrine factors, such as steroid hormones and growth factors, regulate egg productivity in terms of the quantity of egg production, egg weight, sexual maturity, and the number of small yellow follicles (SYF). Insulin-like growth factors (IGF) are involved in the regulation of ovulation rate and ovarian follicular development in chickens, and a relationship between IGF-I genotype and egg weight has been reported. However, the effect of IGF on egg productivity in Korean Native Ogol chickens (KNOC) has been little studied. Therefore, this study was conducted to identify the relationship among endocrine factors (IGF-I, IGF II, estradiol, and progesterone), IGF-I genotypes, and egg productivity. Frequencies of IGF-I genotypes (AA, AB, BB) were 17.3, 26.9, and 55.8%, respectively, within a population. When compared with the IGF-I genotypes, the AB genotype had the highest serum levels of estradiol and progesterone at 40 and 30 wk of age, respectively; the highest IGF-II concentration in F1 follicles at 60 wk; and was positively associated with the number of SYF at 60 wk. The results showed that the A allele was associated with a higher IGF-II expression in the follicles and stimulated the development of follicles, indicating a positive association of the A allele with egg production and the number of SYF. Therefore, these results suggest that there is a possibility of IGF-I genotypes acting as a genetic marker for egg productivity of KNOC. PMID- 15285514 TI - Localization of aquaporins in the sperm storage tubules in the turkey oviduct. AB - Oviductal sperm storage tubules (SST), located at the uterovaginal junction, are the primary site of sperm storage in turkeys. Sperm reside within these storage sites and may be released via a dynamic interaction between sperm mobility and a fluid current generated by the SST epithelial cells. In this study, aquaporins 2, 3, and 9 (proteins that form water channels in the plasmalemma of a variety of cells) were immunocytochemically localized within the apical aspect of the epithelial cells that form the SST. These observations support the contention that the SST epithelial cells are capable of water exchange that may interact with sperm residing within the SST. PMID- 15285515 TI - Recovery of Campylobacter from broiler feces during extended storage of transport cages. AB - Feces deposited in transport cages by a Campylobacter-positive flock can cause the spread of Campylobacter to subsequent flocks placed in the same cages. This experiment was designed to determine the effect of extended cage storage on the viability of Campylobacter in feces deposited on the cage floor during commercial transport and holding. After 4 h of feed (but not water) withdrawal, Campylobacter-positive broilers were caught by commercial catching crews, placed into 3 new commercial cages and transported with the rest of the flock to the holding area at a commercial processing facility. Broilers were allowed to remain in the cages for 8 h before being unloaded by facility personnel. After removal of the broilers, empty cages were held under a shed and sampled at 7 intervals for the presence of viable Campylobacter. Cages were sampled by removing all the feces out of a different randomly assigned compartment in each cage at 0.5, 2, 4, 6, 8, 24, and 48 h after unloading. No decrease in Campylobacter numbers was noted through 8 h of storage. After 24 h in both replications, Campylobacter was detected in 2 of 3 compartments by direct plating and detected in the third by enrichment only. After 48 h, Campylobacter was detected in one replication by enrichment only, and was not detected in the second replication at all. Storing soiled transport cages for 48 h between uses results in lower numbers of Campylobacter in feces, but may not eliminate Campylobacter entirely. Due to cage cost and space requirements, routine cage storage between uses would not be practical. PMID- 15285516 TI - Thermal inactivation of Salmonella and Listeria monocytogenes in ground chicken thigh/leg meat and skin. AB - Thermal inactivation D and z values of Salmonella and Listeria monocytogenes were obtained for chicken thigh and leg meat and skin. The D values of Salmonella at 55 to 70 degrees C were 43.33 to 0.07 min in the meat and 43.76 to 0.09 min in the skin. The D values of L. monocytogenes at 55 to 70 degrees C were 38.94 to 0.04 min in the meat and 34.05 to 0.05 min in the skin. The z value of Salmonella was 5.34 degrees C in the meat and 5.56 degrees C in the skin. The z value of L. monocytogenes was 5.08 degrees C in the meat and 5.27 degrees C in the skin. For Salmonella or L. monocytogenes, the z value of the meat was not different from that of skin. However, the z value of Salmonella in meat or skin was different from that of L. monocytogenes in meat or skin. The z value of Salmonella or L. monocytogenes in chicken thigh and leg meat was different from that in the skin. The results from this study are useful for predicting process lethality of Salmonella and L. monocytogenes in products that contain chicken thigh and leg meat or skin. PMID- 15285517 TI - Quantification of bioaerosols in automated chicken egg production plants. AB - The quantity and composition of bioaerosols in a typical automated chicken egg layer management system (LMS) with a controlled internal climate (B) and without (A) were compared. The LMS-A used a fecal matter disposal system featuring a central opening in the floor through which the matter automatically dropped to an open-air lower level; the LMS-B used a conveyer belt below each hen battery set, which removed the fecal matter frequently. Bioaerosols were collected by impaction on agar. Humidity, wind velocity, temperature, and dust particle concentration were also analyzed at several locations in the LMS. The average bioaerosol concentrations (total viable aerobic bacteria) associated with the inside of LMS-A reached X = 1.1 x 10(5) cfu/m3 with counts in LMS-B being X = 9.2 x 10(4) cfu/m3. In both systems, the bacterial counts were significantly higher on the inside of the LMS than the outside. The LMS-A showed yeast counts of X = 6.7 x 10(1) cfu/m3 with none detectable in LMS-B. Total culturable mold counts were X = 7.0 x 10(2) cfu/m3, with significantly higher presumptive Salmonella spp. counts (X = 6.6 x 10(1) cfu/m3) inside both LMS when compared with the outside. Escherichia coli and total culturable gram-negative counts were significantly higher in LMS-B at concentrations of X = 3.6 x 10(1) cfu/m3. These counts were significantly higher compared with the outside environment. We concluded that the live birds were the major source of bioaerosols in both LMS, with the fecal matter disposal systems attributing to the difference in bioaerosol composition. Modifications to the operation protocols of both LMS to limit the contamination of eggs by bioaerosols are suggested. PMID- 15285518 TI - Effects of carcass washers on Campylobacter contamination in large broiler processing plants. AB - Campylobacter, a major foodborne pathogen found in poultry products, remains a serious problem facing poultry processors. Campylobacter research has primarily focused on detection methods, prevalence, and detection on carcasses; limited research has been conducted on intervention. The aim of this study was to assess the effectiveness of carcass washing systems in 4 large broiler-processing plants in removing Campylobacter species. Washing systems evaluated included combinations of inside/outside carcass washers and homemade cabinet washers. Processing aids evaluated were trisodium phosphate (TSP) and acidified sodium chlorite (ASC). The washer systems consisted of 1 to 3 carcass washers and used from 2.16 to 9.73 L of water per carcass. The washer systems used chlorinated water with 25 to 35 ppm of total chlorine. These washer systems on average reduced Campylobacter populations by log 0.5 cfu/mL from log 4.8 cfu/mL to log 4.3 cfu/mL. Washer systems with TSP or ASC reduced Campylobacter populations on average by an additional log 1.03 to log 1.26, respectively. Total average reductions in Campylobacter populations across the washer system and chill tank were log 0.76 cfu/mL. Washer systems that included antimicrobial systems had total average reductions in Campylobacter populations of log 1.53 cfu/mL. These results suggest that carcass washer systems consisting of multiple washers provide minimal reductions in Campylobacter populations found on poultry in processing plants. A more effective treatment of reducing Campylobacter populations is ASC or TSP treatment; however, these reductions, although significant, will not eliminate the organism from raw poultry. PMID- 15285519 TI - Lipid oxidation in frozen, mechanically deboned turkey meat as affected by packaging parameters and storage conditions. AB - Mechanically deboned turkey meat (MDTM) was stored in different packaging materials (film produced with natural antioxidant (alpha-tocopherol) or synthetic antioxidant) at -20 degrees C for 12 mo in a vacuum, modified atmosphere, or air. One-half of the samples were thawed at 4 degrees C for 24 h after 1 mo of storage and then refrozen. Oxidative rancidity was evaluated during storage by measuring the development of 2-thiobarbituric acid reactive substances (TBARS), and hexanal, a volatile oxidation product. Vacuum- and modified atmosphere-packaged samples had lower TBARS values and hexanal content than air-packaged samples with corresponding treatments. Hexanal content and TBARS values increased with storage time, and the highest levels were obtained after 6 mo of storage. The largest increase was obtained with presence of oxygen. Mechanically deboned turkey meat stored in packages where a natural antioxidant (alpha-tocopherol) was used in production of one of the PE layers, had, in almost every instance, the lowest TBARS values and hexanal content when stored in vacuum or modified atmosphere. However, this difference was not statistically significant. Neither TBARS values nor hexanal content showed dependency of the temperature profile (frozen or frozen/thawed/refrozen) during storage. PMID- 15285520 TI - Diverticulitis: when and how to operate? AB - Diverticular disease, and particularly diverticulitis, has increasing incidence in industrialised countries. Diverticular disease can be classified as symptomatic uncomplicated disease, recurrent symptomatic disease, and complicated disease. Conservative or medical management is usually indicated for acute uncomplicated diverticulitis. Indications for surgery include recurrent attacks and complications of the disease. Surgical treatment options have changed considerably over the years along with the inventions of new diagnostic tools and new surgical therapeutic approaches. Indications and timing for surgery of diverticular disease are determined mainly by the stage of the disease. In addition to this major factor, the individual risk factors of the patient along with the course of the disease after conservative or operative therapy do play a big role in decision-making and treatment of this disease. In this context, the purpose of this article is to review the surgical treatment of diverticulitis with regard to indications, timeliness of operative intervention, operative options and techniques, and special circumstances. PMID- 15285521 TI - Stool test for Helicobacter pylori. PMID- 15285522 TI - Predicting outcome in severe ulcerative colitis. PMID- 15285523 TI - Diagnosis of Helicobacter pylori infection in dyspeptic patients by stool antigen detection usefulness of a new monoclonal enzyme immunoassay test. AB - BACKGROUND: Helicobacter pylori antigens can be measured in human stools with an enzyme immunoassay, which may prove to be a valuable non-invasive diagnostic tool. Aim. To evaluate the usefulness of a new monoclonal enzyme immunoassay for detecting H. pylori antigens in dyspeptic patients' faeces (FemtoLab H. pylori, Connex, Martinsried, Germany). PATIENTS: H. pylori infection was determined in 75 patients (49 men, 26 women, mean age 52 + 16.5) for histology and rapid urease test. METHODS: H. pylori status was established by concordance of the reference tests. FemtoLab H. pylori was measured in triplicate. In addition, two determinations of a polyclonal faecal antigen test (HpSA, Platinum Premier HpSA, Meridian Diagnostic Inc., Cincinnati, USA) were also performed. Sensitivity, specificity, positive and negative predictive values were calculated. Concordance between different measurements was estimated by Kappa statistics. RESULTS: The sensitivity of the FemtoLab H. pylori immunoassay ranged from 98 to 100% and its specificity was 76%. Positive and negative predictive values were 91 and 94-100%, respectively. Concordance coefficients ranged from 0.81 to 0.92. Corresponding HpSA values were 69, 86, 92 and 53%, respectively. Concordance coefficient was 0.61. CONCLUSIONS: FemtoLab H. pylori is a very sensitive, specific, highly reproducible and easy-to-perform tool for diagnosis of H. pylori infection. PMID- 15285524 TI - Helicobacter pylori impairs iron absorption in infected individuals. AB - BACKGROUND: Infection with Helicobacter pylori is recognised as a major risk factor for chronic gastritis, peptic ulcer disease and gastric cancer. The association between H. pylori infection and iron deficiency anaemia has been established. Multiple mechanisms have been advocated to explain the relationship between H. pylori and iron status and their association might reduce iron deposit. AIM: Aim of this study was to investigate whether H. pylori infection affects iron absorption. METHODS: The study was designed on a prospective basis. Fifty-five subjects underwent upper gastrointestinal endoscopy and biopsy to investigate the presence of H. pylori and, when this was positive, also search of serum anti-CagA was performed. Tests included an oral iron absorption test with the administration of 1 mg/kg of Fe2+. Iron levels were measured before and 2 h after iron administration (delta iron). H. pylori-positive subjects were administered antibiotic therapy for 1 week and, 2 months later, the oral iron absorption test was repeated and urea-breath test was first performed. RESULTS: H. pylori-positive subjects had lower serum level of ferritin and lower delta iron compared to H. pylori-negative subjects. That difference is significant in anaemic women and is independent of the presence of serum anti-CagA antibodies. After H. pylori eradication iron absorption test was similar to those of non infected subjects. CONCLUSION: H. pylori infection impairs iron uptake. That mechanism, together with others, may contribute to the depletion of iron in infected patients. PMID- 15285525 TI - Prognosis of severe attacks in ulcerative colitis: effect of intensive medical treatment. AB - BACKGROUND: Severe attacks of ulcerative colitis have a high risk of colectomy. AIMS: To evaluate the effects of standard medical management and to identify the clinical and laboratory variables capable of predicting the clinical outcome. MATERIALS AND METHODS: Prospective study monitoring the clinical and laboratory variables in 67 patients with severe colitis. Therapy consisted of prednisone, cyclosporin if no response, and azathioprine for maintenance. End-points were colectomy or remission. Logistic regression analysis was applied for statistical evaluation. RESULTS: Fourteen (20%) patients required colectomy, 34 (50%) patients achieved remission with steroids, 25 (37%) patients received cyclosporin, 19 (76%) with benefit. Increased body temperature, pulse rate, sedimentation rate and C-reactive protein levels on admission were significantly associated with colectomy. Sedimentation rate greater than 75 mm/h and body temperature exceeding 38 degrees C at admission had 4.6- and 8.8-fold increased risk of colectomy. Less than 40% reduction in the bowel movements within 5 days predicted no response to steroids. Azathioprine maintained remission in 70% of the patients. CONCLUSIONS: Elevated sedimentation rate and fever at day 1 best predict colectomy in severe colitis. Less than 40% reduction in the bowel movements at day 5 predicts no response to steroids. Cyclosporin has a high rate of success in acute attacks and azathioprine in maintaining remission. PMID- 15285526 TI - Diagnostic value of faecal calprotectin in paediatric gastroenterology clinical practice. AB - BACKGROUND: Faecal calprotectin (FC) is a new marker of intestinal inflammation. Data on FC in paediatric gastroenterology clinical practice are still scarce. AIMS: To assess FC values in different paediatric gastrointestinal diseases comparing them with those obtained in healthy children. PATIENTS: Two hundred and eighty-one children (age range 13-216 months) consecutively referred for gastrointestinal symptoms. Seventy-six healthy controls (age range 13-209 months). The exclusion criteria in healthy children were the following: any known underlying chronic disease or a history of abdominal pain, diarrhoea, acute respiratory tract infection, intake of non-steroidal anti-inflammatory drugs, gastric acidity inhibitors, antibiotics, drugs influencing gut motility, and menstrual or nasal bleeding in the last 3 weeks. METHODS: Stool samples stored, prepared and analyzed by an ELISA assay. RESULTS: In healthy children the median FC value was 28.0 microg/g (15-57 interquartile range) with a 95th percentile value of 95.3 microg/g. An increase in FC concentration was observed in all diseases characterized by gastrointestinal mucosa inflammation, and the active inflammatory bowel disease patients showed the higher FC values. All children affected by functional bowel disorders or by non-inflammatory diseases showed normal values. We calculated an optimized FC cut off value of 102.9266 microg/g (revealed by the receiver operating characteristic curve) to distinguish patients with active organic/inflammatory disorders from healthy subjects and from patients with functional bowel disorders. CONCLUSIONS: Calprotectin is a sensitive, but not disease specific, marker to easily detect inflammation throughout the whole gastrointestinal tract. It may help in identifying an organic disease characterized by intestinal mucosa inflammation and in the differential diagnosis of functional bowel disorders. PMID- 15285527 TI - C4BQ0: a genetic marker of familial HCV-related liver cirrhosis. AB - BACKGROUND AND METHODS: Host may have a role in the evolution of chronic HCV liver disease. We performed two cross-sectional prospective studies to evaluate the prevalence of cirrhosis in first degree relatives of patients with cirrhosis and the role of two major histocompatibility complex class III alleles BF and C4 versus HCV as risk factors for familial clustering. FINDINGS: Ninety-three (18.6%) of 500 patients with cirrhosis had at least one cirrhotic first degree relative as compared to 13 (2.6%) of 500 controls, (OR 7.38; CI 4.21-12.9). C4BQ0 was significantly more frequent in the 93 cirrhotic patients than in 93 cirrhotic controls without familiarity (Hardy-Weinberg equilibrium: chi2 5.76, P = 0.016) and in 20 families with versus 20 without aggregation of HCV related cirrhosis (29.2% versus 11.3%, P = 0.001); the association C4BQ0-HCV was found almost only in cirrhotic patients with a family history of liver cirrhosis. CONCLUSIONS: Our studies support the value of C4BQ0 as a risk indicator of familial HCV related cirrhosis. PMID- 15285528 TI - A tumour vaccine of fixed tumour fragments in a controlled-release vehicle with cytokines for therapy of hepatoma in mice. AB - BACKGROUND: Cytokines can be strong potentiators for a tumour vaccine, but they have very short life in vivo when administered as a solution. AIMS: To evaluate the slow release of interleukin 2 from a cytokine-vehicle in vitro and in vivo and to evaluate the anti-tumour activity of a new tumour vaccine in vivo. METHODS: The tumour vaccine was composed of formalin-fixed Hepa 1-6 hepatoma tissue fragments, tuberculin and a lipid based vehicle containing granulocyte macrophage colony-stimulating factor and interleukin 2. The quantity of interleukin 2 release from the cytokine-vehicle in vitro and in vivo was determined by a proliferation assay with CTLL-2 cell line. Hepa 1-6 hepatoma model system with C57BL/6J mice was used to examine protective and therapeutic anti-tumour effect of the vaccine. RESULTS: Release of interleukin 2 from the cytokine-vehicle lasted 5 days in vitro and 3 days in vivo. The vaccine protected 67% of mice from a Hepa 1-6 cell challenge and had a therapeutic effect by prolonging the life span of mice bearing established Hepa 1-6 tumours of 5 mm in diameter. Of the treated mice, 20% became completely tumour-free. CONCLUSIONS: Formalin-fixed tumour fragments and cytokines in controlled-release vehicle are useful in the rational design of tumour vaccines. PMID- 15285529 TI - Osteitis fibrosa cystica, coeliac disease and Turner syndrome. A case report. AB - Today, osteitis fibrosa cystica is seldom present in primary hyperparathyroidism while it is mainly observed in uraemic osteodystrophy. We describe the case of a 54-year-old woman who was found to have huge bone cysts due to osteitis fibrosa cystica in the long bones. A parathyroid adenoma was identified and removed. Coeliac disease and Turner syndrome were diagnosed. Metabolic bone disease due to secondary hyperparathyroidism is common in coeliac disease; however, osteitis fibrosa cystica has not yet been described. PMID- 15285530 TI - Coeliac disease associated with Sjogren's syndrome, renal tubular acidosis, primary biliary cirrhosis and autoimmune hyperthyroidism. AB - Although coeliac disease may occur in patients affected by another immune mediated disorder, its coexistence with multiple autoimmune diseases is not frequently described. We report here the case of a 45-year-old woman referred to our centre because of diarrhoea and weight loss, who had already received a diagnosis of primary biliary cirrhosis, Sjogren's syndrome and renal tubular acidosis. Following the development of diarrhoea we established the diagnosis of coeliac disease, based on the presence of anti-endomysium antibodies and a compatible duodenal biopsy. Despite gluten withdrawal she went on to develop an autoimmune hyperthyroidism. The patient tested positive for HLA DRB1*03 and DQB1*02. The association is unlikely to be casual and may be explained by autoimmune mechanisms, genetic susceptibility and favouring environmental factors commonly shared by the diseases of our patient. PMID- 15285531 TI - The global village of celiac disease. AB - In the last years our knowledge on epidemiology of celiac disease has increased: there is a wide spectrum of its clinical presentation (classical, atypical, silent and latent forms of celiac disease), and of its pathological mucosal intestinal features, which range from early and mild pictures to severe villous atrophy (Marsh stages). In addition, a strong genetic component, associated with the susceptibility to the disease (HLA and non HLA genes), has been found. This knowledge, together with the availability of new high sensitive and specific serological tests (antigliadin, antiendomysium and antitransglutaminase antibodies), has led us to the realization that celiac disease is the most common food intolerance in the world, involving genetically predisposed individuals consuming gluten-containing cereals in their diet. So, today it is well known that celiac disease is a common disorder not only in Europe but also in populations of European ancestry (North and South Americas, Australia), in North Africa, in the Middle East and in South Asia, where until a few years ago it was historically considered extremely rare. Therefore, celiac disease is spread worldwide as in a common "Global Village", and greater levels of awareness and attention on gluten intolerance are needed, both in the Old and in the New World. PMID- 15285532 TI - A case of death after insertion of an intragastric balloon for treatment of morbid obesity. PMID- 15285533 TI - Role of surgery in ovarian cancer: an update. AB - Rupture of an ovarian malignant tumor should be avoided at the time of surgery for an early ovarian cancer. Laparoscopic removal of ovarian cysts should be restricted to patients with preoperative evidence that the cyst is benign. Degree of differentiation is the most important independent prognostic factor in stage I disease and should be used in decisions on therapy in clinical practice and the future FIGO-classification of Stage I. In early ovarian cancer staging adequacy and tumor grade were the only 2 statistical significant prognostic factors for survival in the multivariate analysis of the EORTC ACTION-trial. According to the present data there is no scientific basis to rely only on adjuvant chemotherapy or on optimal staging procedure in medium and high risk stage I ovarian cancer. Primary debulking surgery by a gynecologic oncologist remains the standard of care in advanced ovarian cancer. Optimal debulking surgery should be defined as no residual tumor load. Interval debulking is defined as an operation performed after a short course of induction chemotherapy, usually 2 or 3 cycles. Based on the randomized EORTC-GCG trial, interval debulking by an experienced surgeon improves survival in some patients who did not undergo optimal primary debulking surgery. Based on the GOG 152 data, interval debulking surgery does not seem to be indicated in patients who underwent primarily a maximal surgical effort by a gynecological oncologist. Open laparoscopy is probably the most valuable tool for evaluating the operability primarily or at the time of interval debulking surgery. In retrospective analyses neoadjuvant chemotherapy followed by interval debulking surgery does not seem to worsen prognosis compared to primary debulking surgery followed by chemotherapy. However, we will have to wait for the results of the EORTC-GCG/NCI Canada randomized trial to know whether neoadjuvant chemotherapy followed by interval debulking surgery is as good as primary debulking surgery in some or all stage IIIc and IV patients. The most suitable candidates for secondary debulking surgery are those who had an initial complete response to chemotherapy, a long treatment-free interval (e.g. more than 12 months), and resectable disease (without diffuse carcinomatosis). PMID- 15285534 TI - Heparin-induced thrombocytopenia and vascular surgery. AB - Heparin-induced thrombocytopenia (HIT) is of special interest to vascular surgeons as heparin is the predominant anticoagulant used before, during, and after vascular surgery. Further, the prothrombotic nature of this antibody mediated disorder leads to a high frequency of limb ischemia due to large arterial occlusion by platelet-rich ("white") clots or because of extensive venous thrombosis involving large veins and small venules. This latter syndrome has been associated with coumarin anticoagulation of HIT-associated deep-vein thrombosis (coumarin-induced venous limb gangrene). Non-heparin anticoagulants, such as the direct thrombin inhibitors (lepirudin, argatroban), may be needed for intraoperative management of a patient with suspected acute HIT who requires vascular surgery. The transience of HIT antibodies provides a rationale for intraoperative use of heparin in a patient who has recovered from HIT and in whom HIT antibodies are no longer detectable. PMID- 15285535 TI - Concerns on clinical application of composite tissue allotransplantation. AB - Composite tissue allograft has become a clinical reality: hands, vascularized femoral diaphyses, abdominal walls, a larynx have all been transplanted throughout the world. Conventional immunosuppressive protocol has shown to be sufficient and effective. Rejection has been prevented in most cases and when it did occur it was successfully reversed. Skin has been confirmed as the principal target of acute and chronic rejection. There has been no mortality or early graft losses and, particularly in hand transplantation, the survival graft rate is 91% with a follow-up period ranging from 6 months to 61 months. The side effects of immunosuppression are limited and include primarily transient hyperglycemia, an increase in creatinine values and some opportunistic infections (i.e. cytomegalovirus infection). Nerve regeneration and cortical reorganization have been demonstrated in hand transplantation. Functional results have been encouraging particularly for hand and larynx transplantation. Appropriate indications and patient selection, based particularly on patient motivation and compliance, are essential requirements for composite tissue allograft success. PMID- 15285536 TI - The nomenclature of the veins of the lower limbs, based on their planar anatomy and fascial relationships. PMID- 15285537 TI - Do we need a better classification than CEAP? AB - The CEAP classification, C Clinical, E Etiology, A Anatomy and P Pathophysiology, corresponds to the four main headings of chronic venous disease. Each heading is composed of clearly defined subheadings. This classification is therefore complete and well structured and, as a result of this innovation, far superior to previous classifications. On the other hand, this classification is complex and difficult to use for many clinicians. It also lacks a whole series of important items, such as vascular history, corona phlebectatica, widely used in Europe, and a varicose veins score. More complete and more rigorous studies could be conducted if these items were included under the "C" heading. A number of improvements have been proposed over recent years. They are designed to simplify the CEAP, without introducing any structural changes. The scientific justification for these simple modifications would be an improvement of the coherence. However, these proposals must be validated before being presented to the American Venous Forum. One of the most recent proposals is the development of computer software which would considerably facilitate the use of this classification. Further studies are necessary to demonstrate the value of these modifications. PMID- 15285538 TI - The role of postoperative neovascularisation in recurrence of varicose veins: from historical background to today's evidence. AB - Recurrence of varicose veins at the sapheno-femoral or sapheno-popliteal junction can not always be explained by technical inadequacy of the original intervention. Its development has also been attributed to neovascularisation in the granulation tissue around the ligated saphenous stump. Already in the 19th century, surgeons had noticed that a new vein channel could be formed after ligation or extirpation of a piece of a vein, which could be responsible for recurrence after surgery. After the introduction of the 'high ligation' (crossectomy) in the groin, this theory became less popular. The majority of authors in the 20th century claimed that recurrence was due to the development of incompetence in pre-existing collaterals, which had not been adequately ligated by the previous surgeon. During the period 1960-80's, Dr Glass (Belfast) was one of the first surgeons to focus again on recurrence of varicose veins through regrowth of veins or 'neovascularisation' in interesting series of clinical and experimental work. Neovascularisation seemed to play a crucial role in recurrence after correctly performed sapheno-femoral ligation. With the introduction of duplex scanning, postoperative observation of the phenomena happening at the ligated sapheno femoral or sapheno-popliteal stump has nowadays led to a better understanding of the causes of varicose vein recurrence. Neovascularisation at the ligated stump seems to play a key role in recurrence, which explains why ongoing research efforts are mainly directed at mitigating recurrent reflux related to neovascularisation. PMID- 15285539 TI - Heart transplantation: how to select patients? What are the alternatives? AB - The gap between the number of potential recipients of a cardiac graft and the availability of donor hearts is still growing. A proper selection of heart transplant candidates is mandatory to ensure that patients in critical need, who are likely to benefit from this procedure, are identified. The work-up of a patient with terminal chronic heart failure includes a comprehensive cardiac and systemic evaluation. Critical in the decision is the exclusion of irreversible pulmonary hypertension. Furthermore, underlying diseases that could compromise short- and long-term prognosis after transplantation should be carefully assessed. The mortality on the waiting list and the growing pool of patients with chronic heart failure that is excluded from transplantation has stimulated the search for alternative treatment modalities. Besides a pharmacologic approach, the last few years have witnessed a tremendous evolution in so-called mechanical devices, designed to improve both morbidity and mortality of these patients. Although several of these devices have only just entered the clinical phase, internal cardioverter defibrillators, left ventricular assist devices and biventricular pacemakers can no longer be viewed as experimental tools. PMID- 15285540 TI - Heart transplantation: outcome. PMID- 15285541 TI - Cerebral protection during percutaneous carotid intervention: wich device should be used ? AB - Embolic complications remain the major and unpredictable clinical event during carotid angioplasty and stenting. Cerebral protection devices could play an important role in the prevention of such emboli. Protection devices such as occlusion balloons, filters and reversed flow devices are currently undergoing clinical evaluation and appear to be promising in reducing the incidence of embolic events. This article provides an overview of the three different types of embolic protection devices. PMID- 15285542 TI - Hirschsprung's disease in adults: the Duhamel procedure. AB - Hirschprung's disease is a rare condition in the adult. We present three patients with Hirschsprung's disease who successfully underwent the modified Duhamel procedure. The diagnosis was suspected on the basis of the typical medical history of lifelong constipation. The diagnosis was confirmed by anorectal manometric studies and/or full thickness rectal biopsy. The relief of the marked preoperative abdominal distention was complete and the bowel movements were active and smooth. Two out of three had perfect faecal control and continence. The Duhamel-Martin procedure seems a safe and reliable technique in the treatment of Hirschsprung's disease. PMID- 15285543 TI - Mini-invasive treatment of complications following laparoscopic cholecystectomy. AB - BACKGROUND: Even though laparoscopic cholecystectomy (LC) has become the customary method for treating gallstones, some incidents and complications appear rather more frequently than with the open technique. Several aspects of these complications and their treatment possibilities are analysed. MATERIALS AND METHODS: Over the last 9 years 9542 laparoscopic cholecystectomies have been performed, of which 13.9% were carried out for acute cholecystitis, 38.4% in obese patients and 7.6% in patients aged > 65 years. RESULTS: The main postoperative complications were bile leakage and choleperitoneum (54 cases), haemorrhage (15 cases), subhepatic abscess (10 cases) and retained bile duct stones (11 cases). Classic re-interventions were practiced in 28.8% of cases with complications. Mini-invasive techniques were used in 42.2% of cases with complications: laparoscopic re-interventions (15 cases) for choleperitoneum, haemoperitoneum and subhepatic abscess and endoscopic sphincterotomy (22 cases) for prolonged bile leak on subhepatic drain and for early diagnosed remnant lithiasis of the common bile duct. All cases healed. Another 26 patients were treated conservatively. DISCUSSION: With a precise diagnosis and a good indication, the mini-invasive treatment of complications was completed with good results. 16 laparoscopic re-operations and 22 endoscopic sphyncterotomies were performed (for the treatment of bile leakage and remnant gallbladder stones). PMID- 15285544 TI - Use of radiofrequency in the treatment of minor anal pathology. AB - BACKGROUND: Conventional approaches dealing with chronic anal fissures focus on relieving the anal spasm by manipulation of the internal sphincter. The concomitant pathologies like sentinel piles, anal papillae, or haemorrhoids, however, are often ignored for unknown reasons. MATERIAL AND METHODS: This study describes radiofrequency procedures to remove the above named associated pathologies after performing anal sphincterotomy. A separate, blinded and prospective study between conventional and radiofrequency excision of sentinel pile is described. The measured parameters included the procedure time, intraoperative blood loss, postoperative pain and wound healing time. RESULTS: This combined technique was found useful in treating anal fissures as well as the associated pathologies. The comparative study showed that the intraoperative bleeding was less (p < 0.0001) and wound healing was faster (p < 0.0001) in the radiofrequency group. CONCLUSION: Radiofrequency can be used as an alternative in treating pathologies like the sentinel tags, anal papillae, post fissure fistula or non prolapsing haemorrhoids found associated with anal fissures. PMID- 15285545 TI - Hepatoblastoma in children. AB - Hepatoblastoma is the most common primary liver tumour in children. Complete surgical removal is the treatment of choice for cure; however, in most cases the tumour is unresectable because of its extensive hepatic involvement. Nineteen pediatric cases (11 boys, 8 girls) with ages ranging from three months to 17 years were referred for management to our clinic from 1982 until 2000. All but three suffered from abdominal distention. The other frequent complaints were abdominal mass, anorexia, fatigue, abdominal pain and fever. Physical examination revealed enlarged liver in all patients. In addition to laboratory studies, they were pre-operatively examined by ultrasonography and, in recent cases, computed tomography was also used. Serum alpha-fetoprotein levels were found to be elevated in all patients. In thirteen cases, hepatic resections (10 lobectomies, 2 trisegmentectomies, 1 segmentectomy) were performed. In six children only liver biopsies could be done because of the huge tumour size. However, in three of them the tumours were excised at the second laparotomy, but only one patient survived. All of the patients - except two who were lost in the early postoperative period received chemotherapy whether the tumour was excised or biopsied. In this series the mortality rate was found to be very high (91%) in the 1980s, and more reasonable (50%) in the 1990s, with an overall mortality rate of 73 per cent. This result might be explained with late referral and advanced stage at diagnosis. In addition, we speculate that a combination of improved chemotherapy and technical advances in anesthesia and hepatic resection caused the obvious differences in the survival rates between the two periods. PMID- 15285546 TI - Aneurysm of the splenic vein. AB - Aneurysms of the portal venous system are increasingly reported in the past five years. Congenital weakness of the venous wall, trauma, pancreatitis and portal hypertension are possible etiologies. Surgical intervention is indicated in case of symptomatic aneurysms with or without progressive expansion of the aneurysm diameter. The treatment of asymptomatic splenic vein aneurysms remains debated. We report the case of an asymptomatic and uncomplicated splenic vein aneurysm for which a conservative approach was advocated with regular follow-up by means of Doppler ultrasonography. After six years of follow-up the aneurysm diameter has not changed and no complications were observed. PMID- 15285547 TI - Surgical repair for aortic valve fibro-elastoma: a case report. AB - Valvular cardiac tumours are rare but account for a high prevalence of stroke or sudden death. We report a case of an incidental finding on echocardiographic screening of a 65-year-old man. He was admitted to the emergency department for an episode of sinus bradycardia due to B-blocker overdose. Diagnosis of fibro elastoma was rapidly suspected and the patient underwent surgery in order to prevent embolization. Epidemiology, diagnosis and treatment are discussed. PMID- 15285548 TI - The latissimus dorsi flap for reconstruction of a paralysed deltoid. AB - A case with complete paralysis of the deltoid muscle with irreperable axillary nerve lesion was treated with a functional muscle transfer. The latissimus dorsi muscle was preparated as an island flap, turned over and sutured to the trapezius at one side and to the original deltoid insertion on the humerus at the other side. Function and cosmesis were excellent. PMID- 15285549 TI - The retroperitoneoscopic repair of a lumbar hernia of Petit. Case report and review of literature. AB - In this paper, we comment on a patient who consulted us because of his "lower backpain" together with the appearance of a small swelling at the left side. Anamnesis and clinical examination were suggestive and further simple diagnostic methods confirmed the exceptional diagnosis of a "lumbar hernia of Petit". We describe the retroperitoneoscopic approach of this hernia, its reduction and the fixation of a polypropylene mesh at the surrounding structures with a Tacker. This approach provided a good postoperative comfort, a shorter hospital stay and an early recovery of autonomy and activity. Furthermore, we give a review of the literature concerning lumbar hernias and the evolution of the different reconstruction techniques. PMID- 15285550 TI - Muir-Torre syndrome presenting with ileus: a case report. AB - The Muir-Torre syndrome is characterized by cutaneous neoplasms and visceral malignancies. At least one sebaceous adenoma, epithelioma or carcinoma and at least one internal malignancy are required to make a reliable diagnosis. According to medical literature only two cases of Muir-Torre syndrome with jejunal carcinoma have been reported to date and there is no reported case with intestinal obstruction. Here, we report an unusual case of jejunal carcinoma presenting with ileus. PMID- 15285551 TI - Peripancreatic tuberculous lymphadenitis mimicking carcinoma: report of a case. AB - Peripancreatic tuberculous lymphadenitis is a rare clinical entity and it usually raises serious diagnostic problems. We report a case of a solitary abdominal tuberculoma. A 45-year old woman was admitted to hospital with obstructive jaundice. An exploratory laparotomy was performed. A conglomerated mass, penetrating into the pancreas was found. Since exact diagnosis could not be obtained by peroperative frozen sections, standard Whipple procedure, segmental portal vein resection and reconstruction with autogenous saphenous vein were performed. Histopathological examination of the resected specimen revealed tuberculous lymphadenitis. The patient was given an anti-tuberculous treatment and a good response was noted. Abdominal tuberculoma is often mistaken for a malignant neoplasm and a high grade of suspicion is neccessary in order to make the exact diagnosis and optimal medical treatment of this entity. PMID- 15285552 TI - Malignant transformation of a benign enchondroma of the hand to secondary chondrosarcoma with isolated pulmonary metastasis. AB - Malignant transformation of solitary enchondromas of the hand to secondary chondrosarcomas is extremely rare. We report a case of a recurrent chondromatous tumor of the hand that initially presented with the typical histology of a cellular enchondroma of the small tubular bones but with clinical and radiological signs of malignancy. After development of a single pulmonary metastasis of a chondromyxoid tumor a malignant transformation of the primary enchondroma of the hand must be assumed. PMID- 15285553 TI - Late appearance and slow progression of a breast carcinoma metastasis of the index metacarpal. AB - A case of metastasis into the index metacarpal is described. It appeared 11 years after mastectomy and progressed slowly. The patient has already survived this metastasis for 4 years. PMID- 15285554 TI - Gall-bladder agenesis presenting with obstructive jaundice and elevated CA 19-9. AB - We report the case of an 81-year-old man with agenesis of the gall-bladder that presented with choledocholithiasis, obstructive jaundice, and very high CA 19-9 serum level (2765 U/ml). On ultrasound and CT scan, the gallbladder was not visualised and it was assumed shrunken and filled with gall-stones. After repeated unsuccessful endoscopic retrograde cholangiopancreatography, the patient was operated on for common bile duct (CBD) stones. At laparotomy the gall-bladder was not identified but a 3 cm long gall-stone was removed from the CBD. After decompression of the CBD all symptoms disappeared and the CA 19-9 returned to normal. We believe that this is the first report in the literature of gall bladder agenesis presenting with high serum level of CA 19-9. PMID- 15285555 TI - Non-functioning malignant neuro-endocrine tumour of the pancreas: a case report with a review of the literature. PMID- 15285556 TI - Pancreatic Castleman's tumour: an unusual case. AB - We present an uncommon case of hyaline vascular type Castleman's disease mimicking a pancreatic tumour. A 56-year-old woman with constitutional symptoms was investigated. Pre-operative interventions failed to produce a definitive diagnosis. Surgical excision was performed and the tumour was diagnosed to be the hyaline vascular type of Castleman's disease histopathologically. Pancreatic Castleman's disease should remain a consideration in the differential diagnosis of a pancreatic mass. PMID- 15285557 TI - Choice of protection devices in carotid angioplasty. Two case reports. AB - Patient selection is of the utmost importance with all endovascular procedures. It is equally important to select the appropriate protection device during carotid angioplasty/stenting (CAS). CAS in two patients was unsuccessful due to the chosen protection device. Occlusion of the external carotid artery while deploying a Parodi Anti-Embolic-System Device (PAEC - ArteriA) resulted in cerebral ischaemia. Conversion to the Angioguard System (Cordis) made carotid angioplasty possible, but was complicated by thrombosis of the filter. In the second case it was impossible to negotiate tortuous vessels with the Angioguard system. Our experience illustrates that both devices have limitations. Choosing the wrong device may have serious consequences. PMID- 15285558 TI - Renal cell carcinoma. PMID- 15285559 TI - Epidemiology of renal cell carcinoma. AB - The increasing incidence of RCC in most populations may in part be due to increasing numbers of incidentally detected cancers with new imaging methods. Further, the increase is not only limited to small local tumours but also includes more advanced tumours, which may to some part explain the still high mortality rates. The variation in incidence between populations may have several other explanations. Traditionally the starting point has included thoughts of environmental exposures, which so far have only in part explained the causes of RCC, by means of cigarette smoking and obesity, which may account for approximately 40% of cases in high-risk countries (Table 2). Further, the genetic variations may be of importance as a cause of the difference between populations. Continued research in RCC is needed with the knowledge that nearly 50% of patients die within 5 years after diagnosis. The further search for environmental exposures should take in account the knowledge that RCC consists of different types with specific genetic molecular characteristics. These genetic alterations have in some cases been suggested to be associated with specific exposures. Furthermore, there might exist a modulating effect of genetic polymorphisms among metabolic activation and detoxification enzymes. Hence, a further understanding of the genetic and molecular processes involved in RCC will hopefully give us a better knowledge how to analyse and interpret exposure associations that have importance for both initiation and progression of RCC. PMID- 15285560 TI - Natural and clinical course of renal cell carcinoma--better prospect for the patients. AB - The many improvements in diagnostic and therapeutic tools over the last half century have changed the prospects for patients with renal cell carcinoma (RCC). Higher survival rates and less morbidity have been achieved. The disease is, however, still unpredictable and represents many unsolved problems. In this paper we focus on determinants of the natural and clinical course of RCC and the changes in regard to this over the last decades. PMID- 15285561 TI - Inherited forms of renal cell carcinoma. AB - It is estimated that up to 2% of renal cell cancer (RCC) clusters in families. Several forms of hereditary RCC have been characterized with specific clinical, histopathological, and genetic features. The most common of these is von Hippel Lindau (VHL) disease caused by mutations in the VHL gene and predisposing to clear cell RCC. Predisposition to papillary RCC is present in hereditary leiomyomatosis and renal cell cancer (HLRCC) and hereditary papillary renal cell carcinoma (HPRC). Identification of the genetic defects causing these diseases has enlightened the molecular pathogenesis of RCC, and moreover, provided means to improve patient management. Genetic testing enables early diagnosis of the disease, after which individuals at-risk can be guided to regular surveillance. Screening facilitates detection of presymptomatic early tumors broadening treatment options and potentially improving prognosis. Thus, identification of individuals with inherited cancer susceptibility is important as special management of these patients improves disease outcome. The purpose of this review is to provide clues for identification and management of hereditary renal cancer patients in clinical practice. PMID- 15285562 TI - Progression of malignancy in clear cell renal cell carcinomas. AB - Much progress has recently been obtained in the classification and characterization of RCC by using cytogenetic, gene microarray and proteomic techniques. Pivotal for the understanding of the progression of malignancy of clear cell renal cell carcinomas are findings connecting its biology to inactivation of the von Hippel-Lindau tumour suppressor gene product (VHLp), found in most CC-RCCs. Disruption of VHLp function appears to be involved in altered cell cycle control, resistance to hypoxia, hyperangiogenesis and changes in the organization of cytoskeletal and extracellular matrix proteins in RCC. These changes are reflected in the overexpression of the vascular endothelial growth factor (VEGF) and the subunits of hypoxia-inducible factor (HIF), and other angiogenetic and metastasis-promoting factors. Other changes related to progression of malignancy in RCC are the upregulation of proinflammatory cytokines and changes in cell adhesion proteins. PMID- 15285563 TI - Prognostic factors in renal cell carcinoma. PMID- 15285564 TI - Nephron-sparing surgery--strategies for partial nephrectomy in renal cell carcinoma. AB - The use of partial nephrectomy for renal cell carcinoma has continuously changed in the clinical practice. Previously it was mostly used in imperative cases, in patients with a solitary kidney or in patients with a risk of renal failure. An increased number of incidentally detected renal cell carcinomas are diagnosed due to the advances of the radiological methods. These tumours tend to be smaller and generally with a lower stage. The reported excellent results of partial nephrectomy have promoted the use of nephron-sparing surgery also in patients with a normal contralateral kidney and tumours smaller than 4-5 cm. The technical outcome is excellent with a low operative morbidity and a good oncologic control. Therefore partial nephrectomy has become a standard technique in the treatment of properly selected patients. Laparoscopy with its reduced postoperative pain and shorter rehabilitation time, has encouraged the interest in minimally invasive nephron sparing surgical techniques. Although low, the risk of local tumour recurrence and surgical complications are higher after nephron-sparing surgery compared with radical nephrectomy. Furthermore, long-term renal function remains adequate in most patients with a normally functioning contralateral kidney also after radical nephrectomy. Albeit these facts, there is convincing evidence justifying nephron-sparing surgery to be used routinely for patients with a small renal cell carcinoma and a normal functioning contralateral kidney. PMID- 15285565 TI - Laparoscopic versus open nephrectomy for renal cell carcinoma? AB - Laparoscopic radical nephrectomy has become a well-standardized and reproducible, but technically demanding procedure. It is rapidly replacing the traditional open technique in radical nephrectomy with T1-2 tumours. Open operation will mainly be reserved for T3 tumours. Nephron-sparing surgery will play a major role in small (<4 cm) peripheral tumours. Open technique is still the standard for NSS, but with the refined techniques, laparoscopy may be soon coming. PMID- 15285566 TI - Minimally invasive treatments in renal cell carcinoma. PMID- 15285567 TI - Caval involvement of renal cell carcinoma. PMID- 15285568 TI - Surgery for metastases of renal cell carcinoma. AB - The role of surgery for RCC in the era of emerging effective systemic therapy (usually immunotherapy) is not yet defined except for solitary metastasis. The retrospective analysis of patients subjected to aggressive surgical management after systemic therapy reinforces the need to find better therapeutic modalities in order to achieve complete eradication of metastatic disease. In the meantime, however, we propose these guidelines. First, we would encourage aggressive surgical resection of the clinically solitary metastasis, whether synchronous or metachronous. Continue to follow those patients indefinitely, because relapse is quite likely, but do not give adjuvant systemic therapy unless on protocol. Second, limited metastases in only one organ may behave similarly to a solitary metastasis, and if the metastases are in a site amenable to surgical resection, e.g., lung, initial surgery might be reasonable. Systemic therapy for these patients is highly recommended and need not necessarily wait for recurrence. Third, for patients with multiple metastases, initial systemic therapy followed then by resection of any residual disease in selected patients seems to be supported by the experience at several medical centers. Apparently prolonged survival times have been observed after systemic therapy followed by surgery in highly selected patients, despite finding viable cancer in the overwhelming majority of specimens. One must be mindful of the morbidity of an attempt to remove all known disease, however, and try to weigh this against potential benefit. Only a prospective, randomized trial could ever confirm the value of an aggressive surgical approach to metastatic RCC. In the meantime, however, metastasectomy offers, at the very least, the opportunity to confirm the histologic response to systemic therapy, render some patients disease-free, and possibly promote long-term survival in selected patients. PMID- 15285569 TI - Systemic therapy in metastatic renal cell carcinoma. AB - During the last two decades considerable advances have been made in the understanding of the biology of RCC. Although the best therapeutic options for patients with metastatic RCC have not been defined, it is apparent that use of immunomodulating cytokines like interferon-alpha and interleukin-2 either alone or combined with chemotherapeutic agents provides the best available results in routine clinical practice. Numerous studies have confirmed that objective tumour responses are seen only in a small fraction of patients (averagely in 15-20%). In spite of a lot of evidence that these treatments prolong survival, expectations of only 5-10% long-term survivals with complete and durable regression of tumours are realistic. Recently, some new promising investigational approaches have been reported. These may already in near future further improve overall treatment results. PMID- 15285570 TI - Vaccination immunotherapy--an update. AB - Renal cell carcinoma is often regarded as an "immunogenic tumor". Organ-confined tumors are best treated by operative removal. Adjuvant strategies, however, may improve the outcome after operative therapy. Recently, a phase-III trial using an autologous renal tumor cell vaccine was able to demonstrate a reduction of the risk of progression in patients after radical nephrectomy for renal cell carcinoma larger than 2.5 cm. These results were achieved with minimal side effects. Patients with metastases have a poor prognosis. Thirty years ago autologous tumor cell vaccination resulted in remissions in a small number of patients. Almost all vaccination reports focus on patients with metastatic renal cell carcinoma. These reports differ considerably in their modes of preparation, stimulation, application route and intervals and other relevant parameters. More important, clinical response is limited in most studies. For metastatic renal cell carcinoma none of the various vaccination approaches are being currently investigated in phase-III trials. Ongoing efforts focus on development of more powerful vaccines. This review summarizes vaccination approaches for renal cell carcinomas published in the past 4 years. PMID- 15285571 TI - TT virus infection in acute and chronic liver diseases and in patients regularly receiving blood products in Belgium. AB - BACKGROUND: TT viruses are single-stranded DNA viruses, suggested to be involved in non A-E hepatitis. We studied the prevalence of TTV infection in acute or chronic hepatitis in Belgium in comparison with that in blood donors and in patients regularly receiving blood products. METHODS: TTV-DNA was detected by PCR using the primer set of Takahashi et al (1998) or a nested-PCR specific for genotype-2, because it had been reported that this subtype might be more pathogenic (Tagger et al. 1999). RESULTS: TTV-DNA was present in 49% of 128 patients with chronic hepatitis C, in 54% of 54 with chronic hepatitis B and in 54% of 24 with acute liver failure. This prevalence is similar to the 47% in 127 patients with clotting disorders, or the 64% in 103 undergoing chronic haemodialysis, but lower than the 29.7% found in 340 healthy blood donors. Significant differences in clinical or biochemical characteristics between TTV- positive or TTV-negative patients could not be substantiated. The genotype-2 subgroup comprised 3.9%, but they also did not differ from non genotype-2 patients. CONCLUSIONS: The prevalence of TTV infection was higher in patients than in healthy blood donors. Its clinical significance remains questionable since clinical and biochemical characteristics were not different between TTV positive and TTV negative patients. The higher prevalence of TTV in patients might be related to parenteral transmission, but the relatively high prevalence in healthy blood donors points to an additional presumably faeco-oral infection. The presence of TTV in animals suggests that infection might also originate from food. Long term follow-up will have to define whether co-infection with TTV eventually alters the natural history of chronic hepatitis. PMID- 15285572 TI - Ethical perspectives about organ allocation for transplantation. AB - The ethical questions suggested by the allocation of organs for transplantation can be assembled around the notion of justice. It is a domain of health care in which there is a chronic lack of resources, but for patients whose life is at stake, sometimes at short term. It has similarities with the triage, imposed in some conditions of emergency medicine, at war or during civil catastrophes. The question will be approached from the points of view of the main stakeholders. PMID- 15285573 TI - History of pediatric liver transplantation in Europe. PMID- 15285574 TI - The paediatric liver transplantation program at the Universite catholique de Louvain. AB - The Paediatric Liver Transplant Program at Saint-Luc University Clinics constitutes a substantial single centre experience, including 667 transplantations performed between March 1984 and April 2003, and the history of this program reflects the tremendous progress in this field since twenty years. Liver transplantation in children constitutes a considerable undertaking and its results depend on multiple, intermingled risk factors. An analysis of the respective impact of several surgical and immunological parameters on patient/graft outcome and allograft rejection after paediatric liver transplantation showed a significant learning curve effect as well as the respective impact of pre-transplant diagnosis on survival and of primary immunosuppression on the rejection incidence. The introduction of living related liver transplantation in 1993 not only permitted to provide access to liver replacement in as many as 74% more candidate recipients, but also resulted in better graft survival and reduced retransplantation rate. The results of a recent pilot study suggest that steroid avoidance is not harmful, and could even be beneficial for paediatric liver recipients, particularly regarding growth, and that combining tacrolimus with basiliximab (anti-CD25 chimeric monoclonal antibody) for steroid substitution appears to constitute a safe alternative in this context. The long-term issues represent the main future challenges in the field, including the possibility of a full rehabilitation through immunosuppression withdrawal and tolerance induction, the development of adolescence transplant medicine, and the risk of early atherogenesis in the adulthood. PMID- 15285576 TI - The psychological challenge of paediatric organ transplantation: gift and incorporation. AB - The specific feature of organ transplantation is the confrontation of the patient with a body part not being his own and coming from somebody else. Thus the psychological challenge of transplantation will be the gift and incorporation of the graft. The given organ confronts the patients with mental manifestations related to emotions of grieving and guilt, most constantly directed to the donor himself. It can also cause a fantastical imagination related to the idea of having to some extent inherited the character of the donor. This very special gift relationship has to be questioned, as it can be interpreted in terms of a tyranny of the dept. A dept the recipient will never be able to reimburse. In this context the contribution of sociological knowledge is determining. It makes it possible to reconsider the problem, and to discover that a gift relationship offers transplant patients much vaster possibilities than a materialistic conception of the gift, based on the dept, would do. PMID- 15285575 TI - Genetic cholestatic liver diseases: the example of progressive familial intrahepatic cholestasis and related disorders. PMID- 15285577 TI - Adult liver transplantation at UCL: update 2002. AB - The authors present the results of a single centre study of 587 liver transplants performed in 522 adults during the period 1984-2002. Results have improved significantly over time due to better pre-, peri- and post-transplant care. One, five, ten and fifteen year actuarial survivals for the whole patient group are 81.2; 69.8; 58.9 and 51.2%. The high incidence of de novo tumors (12.3%), of cardiovascular diseases (7.5%) and of end-stage renal function (3.6%) should be further incentives to tailor the immunosuppression to the individual patient and to direct the attention of the transplant physician to the long-term quality of life of the liver recipient. PMID- 15285578 TI - Viral hepatitis B, C and D. PMID- 15285579 TI - Treatment of hepatocellular carcinoma at the dawn of the third millennium: liver transplantation and its alternatives. AB - Hepatocellular carcinoma is one of the most frequent tumors worldwide, and its frequency is increasing. The management of hepatocellular carcinoma has changed in recent years, this because screening allows to discover tumors at an earlier stage, and because of effective treatments are available, such as liver transplantation, liver resection, percutaneous ablation and transarterial chemoembolization. Each one of these treatments has its own advantages and drawbacks, and range of application according to the stage of the tumor and of the underlying liver disease. This review summarizes the recent progress in the management of HCC and the practice in our unit. PMID- 15285580 TI - Metastatic follicular dendritic cell sarcoma of the stomach: a case report and review of the literature. AB - Follicular dendritic cell (FDC) sarcomas are rare tumours, typically seen in lymph nodes. However, in about one third of the reported cases, a FDC sarcoma presents as an extranodal mass. Involvement of the gastrointestinal tract is extremely rare, and only 3 cases have been described to date. We report on a 40 year-old female patient with a follicular dendritic cell sarcoma located in the stomach and the presence of a metastasis in the liver at the time of diagnosis. Severe asthenia, nausea, back pain and loss of weight were the presenting symptoms. A CT scan of the abdomen and an upper gastrointestinal endoscopy revealed a tumour mass in the stomach. The diagnosis of a FDC sarcoma was made on histological and immunohistochemical findings. We report the second case of a FDC sarcoma presenting in the stomach. Due to its rarity, a FDC sarcoma seldom enters the differential diagnosis of spindle cells neoplasms of the gastrointestinal tract. Complete surgical resection is the treatment of choice for FDC sarcoma. PMID- 15285581 TI - Perianal Paget's disease: case report and review of the literature. AB - Cutaneous Paget's disease (PD) is a rare entity, predominantly involving the breasts. Anal involvement is rather exceptional, and can be associated with underlying malignancies, among which prostate and rectal adenocarcinoma. We report the case of a 71-year-old man suffering from a long history of anal itching, associated with an erythematosquamous lesion of the right buttock extending up to the anus. The diagnosis of perianal PD (PAPD) was confirmed by histopathological demonstration of Paget's cells from a biopsy performed after ineffective topical treatment. Radiological and further clinical inspections allowed us to exclude any synchronous malignancy. A first-step surgery consisted in coelioscopic diverting sigmoid colostomy, along with multiple perianal, anal and rectal biopsies revealing an anal canal involvement. Coelioscopic abdominoperineal surgery and a wide cutaneous excision were then performed. Histopathological analysis revealed positive posterior margin, requiring further excision. No adjuvant therapy was prescribed, and to this day, after a one-year and a half follow-up, the patient remains disease-free. Our case report and review of PAPD stress that appropriate management is required to improve the poor prognosis of this rare affection. PMID- 15285582 TI - Idiopathic eosinophilic oesophagitis: atypical presentation of a rare disease. AB - A 72 year-old man presented severe dysphagia and weight loss of recent onset. Repeated oesophageal endoscopy and biopsies with macroforceps were normal. Oesophageal manometry disclosed features compatible with achalasia. Oesophageal EUS endoscopy localized an infiltrating process between muscular layers of the oesophageal wall and CT scan delimited a circular thickening in the inferior part of the oesophagus. Because of severe clinical presentation mimicking a possible oesophageal neoplasm like a lymphoma, partial oesophagectomy was performed and revealed eosinophilic oesophagitis. This unusual presentation emphasizes that idiopathic eosinophilic oesophagitis must be proposed in the differential diagnosis of dysphagia, even in old patient without apparent oesophageal lesion at endoscopy. PMID- 15285583 TI - MR imaging findings in a patient with hepatic veno-occlusive disease. AB - We report the MRI findings in a 31-year-old woman with veno-occlusive disease. MRI demonstrated patent hepatic veins and patchy signal enhancement of the liver after gadolinium chelate injection. This enhancement was compatible with sinusoidal congestion. The diagnosis of veno-occlusive disease was confirmed by histological examination of liver biopsy. The diagnosis of veno-occlusive disease should be evoked when patchy liver enhancement suggestive of sinusoidal congestion is observed in the absence of hepatic vein thrombosis and congestive heart failure. PMID- 15285584 TI - [Thoracoscopic thymectomy]. AB - BACKGROUND/PURPOSE: The use of videothoracoscopy in thoracic pediatric pathology has been progressively accepted in different diagnostic and therapeutic procedures along last decade. The aim of this work is to analyze our initial experience in thymectomy through this approach. PATIENTS AND METHODS: We have used the thoracoscopic approach in the last two cases of thymus pathology with surgical indication. Case 1: 9 year-old patient recently diagnosed on myasthenia gravis and several hospital admissions because of clinical worsening. Case 2: 9 year-old patient with a 7x8x3.5 cm. cervico-mediastinal tumour. FINAL DIAGNOSIS: Multilocular cystic thymoma. In both cases we used right approach in lateral decubitus, and the harmonic scalpel. RESULTS: Mean operating time was 125 minutes. No procedure-related complications. They were discharged on the 6th and 4th postoperative day. After 6 and 7 month follow-up, no incidences have been found, and case 1 has shown a sympthomatic improvement and a decrease in drug dosage. CONCLUSIONS: Thoracoscopy is a good alternative in thymus approach. Its cosmetic and recovery advantages upon transcervical and transsternal are obvious. Despite our very initial experience, we believe that this approach at least equals classic ones in the ability to resect the whole thymus. PMID- 15285585 TI - [Contrast-enhanced sonourethrography versus conventional miction cystourethrography in the assessment of the urethra: preliminary study]. AB - BACKGROUND AND AIMS: Voiding cystourethrography (VCUG) is the first choice imaging modality for assessing the urethra, but this technique exposes patients to ionizing radiation. Cystosonography with echocontrast (CS) has proved to be a reliable technique to detect and grade vesicoureteral reflux (VUR) without exposing patients to ionizing radiation, but its capacity to adequately study the urethra has yet to be demonstrate in large series of patients. The aim of this study is to demonstrate the reliability of contrast-enhanced CS for assessing the urethra by comparing the results with those of the VCUG. MATERIAL AND METHODS: 108 patients were studied with ultrasound (US) using a galactose-based contrast agent. This exam was always followed by VCUG. Basal and voiding urethral US studies were performed with. Patients were studied in supine decubitus position. Girls were examined by longitudinal translabial approach, with the probe (a 7.5 MHz liner array transducer) positioned longitudinally at the introitus, to evaluate the bladder neck and urethra. In boys the transducer was initially placed longitudinally in the escrotum at ventral root of the penis to assess the bladder neck and proximal bulbar urethra, and then displaced distally toward the penile urethra. On basal study the echogenic urethral mucosa and the collapsed sonolucent urethral lumen were indentified and measured when distended. The patients were asked to void with the probe in place. During voiding attention was focused on elasticity and distention of urethral walls, as well as in the caliber of the entire urethra. Patients unable to void during either CS or VCUG were excluded. RESULTS: The bladder neck and the entire urethra were well demonstrate with CS. All females and 43 males showed a normal urethra both in CS and VCUG. Four patients were dignoses of posterior urethral valves (PUV) with CS and confirmed at VCUG, one patient had anterior urethral valves and 5 showed urethral stenosis at both techniques. Three patients with a vesicosphincteric dysinergia, 14 with resected PUV and one with a resected syringocele were adequately evaluated. Twelve girls showed vaginal reflux. The information provided by CS was equivalent to the VCUG in all patients but two with a syringocele (only seen on VCUG). CONCLUSIONS: CS is a reliable imaging modality sufficiently sensitive and specific to study the urethra, adding dynamic information to VCUG and can be used as a complement to VCUG. PMID- 15285586 TI - [Rectosigmoidectomy and end to end coloanal anastomosis with mechanical stapler for treatement of Hirschsprung disease]. AB - The experience of the Pediatric Surgical Service of the Materno Infantil University Hospital from Malaga on surgical treatment of the Hirschsprung disease by means of a modified technique of Swenson and Rehbein operations is presented. MATERIAL AND METHODS: Between 1992 and 2001 25 patients were operated of a rectosigmoidectomy. Of them, 23 were diagnosed of Hirschsprung's disease, one suffered a rectal angiodisplasia and the other one presented with a rectal stenosis secondary to a previous rectosigmoidectomy. In all of them a transabdominal rectosigmoidectomy with coloanal end to end anastomosis by means of a circular intraluminal stapler was performed. In 10 of them (group A), a modified Rehbein operation with intraabdominal anastomosis was performed. In the remaining 15 patients (group B), a modified Swenson operation with exteriorization of the aganglionic colon through the anus and extrabdominal anastomosis was performed. RESULTS: The postoperative course was evaluated by measuring the postoperative fasting time and the first spontaneous deposition. The medium hospital stay was of 9 days, nevertheless in 16 patients (64%) was lower than 7 days. The postoperative complications are presented. It consists in 1 anastomotic leakage (4%), postoperative enterocolitis 1 case (4%) and transient anastomotic stenosis in 4 patients (16%). All of them were treated with conservative treatment except one case of stenosis which needed a sphincterotomy. CONCLUSIONS: The rectosigmoidectomy and coloanal end to end anastomosis with endoluminal stapler is a safe and easy to do technique to treat the Hirschsprung's disease allowing a deep rectal resection which is very difficult to achieve by manual suture. The anastomosis is located in and extraperitoneal position, with a minimum risk of peritoneal involvement in case of anastomotic leakage. The patients presented a fast recovery, a minimum of complications and good functional result. PMID- 15285587 TI - [Indication of appendectomy in the recurrent abdominal pain]. AB - INTRODUCTION: The frequency of the recurrent abdominal pain without demonstrable pathology is high and in occasions it takes to the surgeon to practice an appendectomy. In most of the cases the pain disappears. This performance although subjective, it doesn't base on objective data, what takes us to carry out this work. MATERIAL AND METHOD: The study is carried out in 47 patients with recurrent abdominal pain and subjected to appendectomy, 17 were excluded by diverse causes. The vermiform appendixes were studied by morphological and immunohistochemical methods, using the monoclonal antibodies, valuing morphologic sing of a chronic inflammation of the appendix. RESULTS: 100% of the samples studied presented microscopically chronic inflammatory changes microscopically. We observe fibrose in 100%, lympho-plasmocytic inflammatory infiltrate in 60% and a lymphocytes T prevalence has more than enough lymphocytes B. CONCLUSIONS: Our results support the existence of a clinical-pathological condition that can be defined as chronic appendicitis and therefore it justifies the appendectomy in recurrent abdominal pain when another disease has been excluded. PMID- 15285589 TI - [New Modern Magpi: meatal advancement and glanuloplasty clinical course]. AB - INTRODUCTION: Meatal advancement and glanuloplasty (MAGPI) described by Duckett, has been the most accepted technique for distal hypospadias repair along the last 20 years. Only 50% of the distal variants are amenable to the Magpi; last years several modifications has been reported in order to reach better cosmetic results and to make it available for the most of the distal hypospadias. We report the first 20 cases with some modifications of Magpi. MATERIALS AND METHODS: The basic steps of the technique are: we remove a triangular segment of glanular tissue distal to the meatus. Dissection of the dorsal and lateral urethral sides is made, accomplishing the urethral advancement without any tension. Strips of glandular epithelium are excised on each side, and glans tissue is sutured above the ventral urethral wall. RESULTS: There were 11 coronal, and 9 glanular hypospadias (3 with megameatus). 8 cases showed a slight incurvation that disappeared after releasing any cutaneous chordee. The posoperative follow-up was 35 weeks (R=7-48 weeks). Cosmetic and functional results were excellent, showing a natural circumcized penis. There wasn't any complications as meatal retraction, stenosis or incurvation. There was only a transitional hematoma and a minimal fistula that closed expontanely. CONCLUSIONS: New Modern Magpi adds minimal variations to the original technique, avoiding the cosmetic limitations derived from borderline indications. New Modern Magpi is amenable to near all of glanular hypospadias and most of coronal hypospadias. PMID- 15285588 TI - [Diagnosis and surgical treatment for male pseudohermaphroditism in a multidisciplinary unit of intersexual conditions]. AB - The birth of a child with ambiguous genitalia represents a very stressing situation for the family, and afterwards has great social and psychological repercussion for the patient itself. Until now, the sex assignation is being done according to the phenotype. Now, with the molecular diagnosis of the genes that play a role in the sexual development, the assignation must also take into account the prognosis of response to androgens. The aim of this work is to review the 40 male pseudohermaphroditism cases controlled in our hospital and the genetic molecular diagnosis done in 19 cases, thus obtaining the certainty diagnosis. In 15 patients the mutations were located in the AR gene (androgen receptor). In 2 cases the mutation affected the SRD5A2 gene (deficiency of 5a reductase) and in the other 2 cases it affected the HSD17BIII gene (deficiency of 17-ketoreductase). If the mutations affect the AR gene they must be assigned to the feminine sex, because of the impossibility of virilisation at puberty (lack of response to androgens). If the mutations are located in the other 2 genes they can be assigned to the masculine sex, since in puberty, they will present good response to androgens and will virilize. The molecular diagnosis offers us also the possibility to establish a prenatal diagnosis and to offer genetic advice. PMID- 15285590 TI - [Puberal gynaecomastia. Review of nine cases]. AB - Gynaecomastia is more frequent during the adolescence. In this paper, a retrospective study of nine cases diagnosed and treated on a period of six years is presented. The reports of nine patients are reviewed. The mean age at diagnosis was 10.4 years (range 9-12). In all cases a bilateral mammary hypertrophy was detected, symmetrical in 6 cases (67%) and asymmetric in 3 (33%). After a mean follow-up period of 24 months, surgical treatment was indicated: a subtotal subcutaneous mastectomy was performed in all nine cases. Only two patients developed early complications: one wound infection and one haematoma. The long-term plastic results were satisfactory after a mean follow-up postoperative period of 11 months. The general management of this kind of pathologic process is presented. In the authors experience the elective surgical procedure is the subtotal subcutaneous mastectomy as simple procedure with a very short morbidity and excellent plastic results. PMID- 15285591 TI - [Transanal endorectal pull-through alone as treatment of Hirschsprung's disease]. AB - The aim of this work is to present our experience in the treatment of Hirschsprung's disease (HD) with the technique described by De la Torre. MATERIALS AND METHODS: Seven children diagnosed with recto-sigmoid aganglionism have been treated with this surgical technique, to which a few modifications have been done. RESULTS: There were no intra- nor early postoperative complications. Surgical time ranged 150 to 240 minutes (average 198). All children began oral feedings 2 or 3 days postoperatively (average 2.4). Hospital stay averaged 5.2 days. Follow-up ranges from 6 months to 3 years (average 16 months). Two late complications were seen--one anastomotic stricture and one constipation--and successfully treated as out patients. CONCLUSIONS: The transanal only approach carries a rapid recovery. Family satisfaction is high because of the lack of scars. We believe this is the treatment of choice when confronting rectosigmoid aganglionism. PMID- 15285592 TI - [Congenital neuroblastomas]. AB - BACKGROUND: We consider congenital neuroblastomas (CN) those detected in pregnancy or at the very first hours of life. Due to perinatal sonography, its incidence has increased in the last years. We present herein our experience in the treatment of this condition and we try to find out any different clinical pattern from those neuroblastomas diagnosed later in life. METHODS: We review the CN treated in our hospital from 1990 to 2003, analyzing diagnosis, localization, tumor staging, N-myc amplification, treatment and evolution. RESULTS: Among the 107 neural tumors managed during this period (89 neuroblastomas, 18 ganglioneuromas), 8 were congenital neuroblastomas (7 girls, 1 boy). Two patients had prenatal diagnosis and 6 tumours were detected in routine exploration or casual findings upon neonatal examination. Six were abdominal, 1 thoracoabdominal and 1 abdominopelvic with dumbbell invasion. Three tumours were classified like stage 1, 1 stage 2, 1 stage 3, 1 stage 4 and 2 stage 4S. Although most of them had unfavorable histology, we didn't find N-myc amplification in any tumor. All patients were operated upon, with preoperative chemotherapy in 2 of them. Resection was complete in 7 out of the 8 tumors. The patient who presented neurological symptoms at birth recovered neither motility nor bladder function after resection. All of them survive after 60+/-53 months of follow-up. CONCLUSIONS: The outcome in this group of neuroblastomas is better that expected, probably because of its abdominal location. On the contrary in dumbbell neuroblastomas, neurological damage at birth seems to be irreversible. PMID- 15285593 TI - [Diagnosis and management of necrotizing tracheobronchitis]. AB - OBJECTIVE: Necrotizing tracheobronchitis has been described as a complication of mechanical ventilation of newborns with respiratory failure. Neonates with necrotizing tracheobronchitis present a diverse clinical spectrum from asymptomatic disease to severe airway obstruction that causes 45% of mortality. The objective of our study is to analyze our experience in the management of these patients. MATERIAL AND METHODS: In the last three years we have treated eight patients with necrotizing tracheobronchitis The mean age was 0.84 +/- 0.95 months, gestational age of 37.43 +/- 2.3 weeks. The mean weight was 3.07 +/- 1.04 kg. Five patients had a congenital heat disease (62.5%) and three have a respiratory failure (37.5%). We have analyzed the contributing factors, symptoms, diagnosis, treatment and results. RESULTS: All patients presented episodes of shock with treatment of drugs. Five patients have conventional ventilation (62.5%) while three have high frequency oscillatory ventilation (37.5%). There were not significant differences in the ventilator parameters of both groups (PMA, PIP, PEEP). Three patients were supported by ECMO when they developed necrotizing tracheobronchitis. The treatment was bronchoscopic removal of necrotic tissue. There was not any complication after the procedure. A patient suffered a stenosis in the left main bronchus. Three patients have died during follow-up for different causes. The time of pursuit is of 10.33 +/- 7.61 months. CONCLUSIONS: Necrotizing tracheobronchitis may be increasing in the Neonatal Intensive Care Units, due to a bigger survival of patient with serious respiratory failure and shock. Hypotension and shock seems to be a major contributing factor in the development of this lesion. Bronchoscopy is necessary for treatment and survival of the patients. PMID- 15285594 TI - [Paraneoplasic syndrome in inflammatory pseudotumor of the lung]. AB - BACKGROUND: Inflammatory pseudotumor (IPT) is the most frequent pulmonary mass in childhood. It is histologically benign but locally aggressive. Atelectasis and recurrent airway infections are the most frequent presenting findings. We present two children in whom first clinical signs were paraneoplasic syndromes. MATERIAL AND METHODS: Retrospective study of two cases of IPT treated in our clinic from 1998 to 2002. Age, clinical presentation, preoperative diagnosis, treatment, histological diagnosis and postoperative outcome were reviewed from clinical chart. RESULTS: Case 1: 7 year old male with incidental diagnosis of superior right lobe IPT in routine study because of diabetes. The mass collapsed superior and median lobar arteries and compressed superior cava vein. Right pneumonectomy was necesary to complete removal of the mass. From immediate postoperatory the child became normoglycemic and is free of insulin. Case 2: 11 year old male with rheumatologic clinic consisting in hypertrophic osteoarthropathy, arthralgy and knees liquid lasting for two years. X-ray examination showed mediastinic mass. He underwent complete removal of a pulmonary mass. Few months after the operation the rheumatologic-like symptoms were resolved. CONCLUSIONS: IPT can present with paraneoplasic syndromes, although physiopathology is not well understood. In children with recurrent respiratory infections one diagnosis to take in mind is IPT, even more when clinic is associated with symptoms compatible with paraneoplasic syndrome. PMID- 15285596 TI - Informal reasoning: theory and method. PMID- 15285595 TI - [Jejunal diaphragm. Laparoscopic treatment in newborns]. AB - Intestinal obstructions in newborns secondary to atresias or membranes have been treated until the current moment through laparotomy and resection with anastomosis. Recently, it has been reported the use of minimally invasive techniques to correct this congenital anomaly. We present a case of jejunal diaphragm treated in our Service. Diagnosis and treatment by mean of endoscopic techniques was achieved in the fourth day of life. Operative time was 180 minutes. Postoperative course was uneventful. Results in terms of function and cosmetic were excellent. Laparoscopic management in selected cases of intestinal atresia has proven to be safe and effective and represents an alternative to neonatal open surgery. PMID- 15285597 TI - A Bayesian approach to the argument from ignorance. AB - In this paper, we re-examine a classic informal reasoning fallacy, the so-called argumentam ad ignorantiam. We argue that the structure of some versions of this argument parallels examples of inductive reasoning that are widely viewed as unproblematic. Viewed probabilistically, these versions of the argument from ignorance constitute a legitimate form of reasoning; the textbook examples are inductive arguments that are not unsound but simply weak, due to the nature of the premises and conclusions involved. In an experiment, we demonstrated some of the variables affecting the strength of the argument, and conclude with some general considerations towards an empirical theory of argument strength. PMID- 15285598 TI - Theory and data interactions of the scientific mind: evidence from the molecular and the cognitive laboratory. AB - A number of researchers and scholars have stressed the importance of disconfirmation in the quest for the development of scientific knowledge (e.g., Popper, 1959). Paradoxically, studies examining human reasoning in the laboratory have typically found that people display a confirmation bias in that they are more likely to seek out and attend to data consistent rather than data inconsistent with their initial theory (Wason, 1968). We examine the strategies that scientists and students use to evaluate data that are either consistent or inconsistent with their expectations. First, we present findings from scientists reasoning "live" in their laboratory meetings. We show that scientists often show an initial reluctance to consider inconsistent data as "real." However, this initial reluctance is often overcome with repeated observations of the inconsistent data such that they modify their theories to account for the new data. We further examine these issues in a controlled scientific causal thinking simulation specifically developed to examine the reasoning strategies we observed in the natural scientific environment. Like the scientists, we found that participants in our simulation initially displayed a propensity to discount data inconsistent with a theory provided. However, with repeated observations of the inconsistent data, the students, like the scientists, began to see the once anomalous data as "real" and the initial bias to discount that data was significantly diminished. PMID- 15285599 TI - Conditions for the acceptance of deontic conditionals. AB - Recent psychological research has investigated how people assess the probability of an indicative conditional. Most people give the conditional probability of q given p as the probability of if p then q. Asking about the probability of an indicative conditional, one is in effect asking about its acceptability. But on what basis are deontic conditionals judged to be acceptable or unacceptable? Using a decision theoretic analysis, we argue that a deontic conditional, of the form if p then must q or if p then may q, will be judged acceptable to the extent that the p & q possibility is preferred to the p & not-q possibility. Two experiments are reported in which this prediction was upheld. There was also evidence that the pragmatic suitability of permission rules is partly determined by evaluations of the not-p & q possibility. Implications of these results for theories of deontic reasoning are discussed. PMID- 15285600 TI - Promise is debt, threat another matter: the effect of credibility on the interpretation of conditional promises and threats. AB - Evidence from reasoning tasks shows that promises and threats both tend to receive biconditional interpretations. They also both display high speaker control. On the face of it, the only difference seems to be the positive or negative signing of the consequent. In a promise, the speaker tries to persuade the hearer to do something by holding out the prospect of a particular reward; in a threat, the speaker tries to refrain the hearer from doing something by holding out the prospect of a particular punishment. This paper investigates the respects in which conditional promises and threats differ further by means of an inference task. The credibility of the consequent was manipulated in order to examine whether the acceptability ratings of inferences based on promises and on threats would be equally affected. The results of the inference task and an analysis of the reasons people give for their answers suggest that the credibility of promises is less affected by the use of excessive consequents than the credibility of threats. In other words, promise remains debt, whereas threat is another matter. PMID- 15285601 TI - Counterfactual and prefactual conditionals. AB - We consider reasoning about prefactual possibilities in the future, for example, "if I were to win the lottery next year I would buy a yacht" and counterfactual possibilities, for example, "if I had won the lottery last year, I would have bought a yacht." People may reason about indicative conditionals, for example, "if I won the lottery I bought a yacht" by keeping in mind a few true possibilities, for example, "I won the lottery and I bought a yacht." They understand counterfactuals by keeping in mind two possibilities, the conjecture, "I won the lottery and I bought a yacht" and the presupposed facts, "I did not win the lottery and I did not buy a yacht." We report the results of three experiments on prefactuals that examine what people judge them to imply, the possibilities they judge to be consistent with them, and the inferences they judge to follow from them. The results show that reasoners keep a single possibility in mind to understand a prefactual. PMID- 15285602 TI - The story of some: everyday pragmatic inference by children and adults. AB - The statement, some elephants have trunks, is logically true but pragmatically infelicitous. Whilst some is logically consistent with all, it is often pragmatically interpreted as precluding all. In Experiments 1 and 2, we show that with pragmatically impoverished materials, sensitivity to the pragmatic implicature associated with some is apparent earlier in development than has previously been found. Amongst 8-year-old children, we observed much greater sensitivity to the implicature in pragmatically enriched contexts. Finally, in Experiment 3, we found that amongst adults, logical responses to infelicitous some statements take longer to produce than do logical responses to felicitous some statements, and that working memory capacity predicts the tendency to give logical responses to the former kind of statement. These results suggest that some adults develop the ability to inhibit a pragmatic response in favour of a logical answer. We discuss the implications of these findings for theories of pragmatic inference. PMID- 15285603 TI - Reasoning, integration, inference alteration, and text comprehension. AB - This paper reports how the study of formal logical reasoning provides insight into more everyday types of reasoning, such as that involved in language comprehension. Both of these types of cognition are thought to involve the use of mental models, and so it is reasonable to think that the cognitive operations needed for formal logical reasoning would be involved in everyday reasoning as well. We focused on three aspects of formal reasoning: (a) the integration of information into a common mental model, (b) the drawing of inferences, and (c) the coordination of alternative possibilities. We were able to show that the integration and inference components were related to narrative comprehension processes, but the coordination of alternative models was not. Thus, there is evidence for some overlap in the mental processes used in formal and everyday reasoning. This further justifies the study of formal logical reasoning as a window into certain types of everyday reasoning. PMID- 15285604 TI - Cultivating expertise in informal reasoning. AB - People generally develop some degree of competence in general informal reasoning and argument skills, but how do they go beyond this to attain higher expertise? Ericsson has proposed that high-level expertise in a variety of domains is cultivated through a specific type of practice, referred to as "deliberate practice." Applying this framework yields the empirical hypothesis that high level expertise in informal reasoning is the outcome of extensive, deliberate practice. This paper reports results from two studies evaluating the hypothesis. University student participants completed 12 weeks of deliberate practice in informal reasoning. Quantity of practice was recorded by computer, and additionally assessed via self-report. The hypothesis was supported: Students in both studies showed a large improvement, and practice, as measured by computer, was related to amount of improvement in informal reasoning. These findings support adopting a deliberate practice approach when attempting to teach or learn expertise in informal reasoning. PMID- 15285606 TI - [Polymorphisms of CYP1B1 and COMT in breast and endometrial cancer]. AB - CYP1B1 and COMT code for the key enzymes of catecholestrogen biosynthesis and metabolism, and their polymorphisms determine a variation of enzymic activities. RFLP analysis was used to study the allele and genotype frequency distributions of CYP1B1 polymorphisms Arg48Gly, Ala119Ser, and Val432Leu and COMT polymorphism Val158Met in 210 breast cancer patients, 138 endometrial cancer patients, and 152 healthy women. The COMT polymorphism showed no significant association with breast or endometrial cancer. For the first time, such association was observed for the CYP1B1 polymorphisms. CYP1B1 allele C (Arg48), which codes for the enzyme more active in estradiol 4-hydroxylation, was associated with higher risk of breast (OR = 3.22, CI 2.34-4.43, p = 0.000) and endometrial (OR = 2.43, CI 1.72 3.44, p = 0.000) cancer. Similar data were obtained for CYP1B1 allele G (Ala119): OR = 2.18, CI 1.58-3.01, p = 0.000 in breast cancer and OR = 2.52, CI 1.78-3.56, p = 0.000 in endometrial cancer. Risk of endometrial, but not breast, cancer was significantly higher in carriers of CYP1B1 genotype Val432/Val. This was explained by stronger estrogen dependence and, consequently, higher estrogen reactivity of the endometrium as compared with the mammary gland. PMID- 15285605 TI - [Functions of p21Waf1 in norm and in stress]. AB - Cyclin-dependent kinase inhibitor p2(Waf1/Cip1/Sdi1/CAP20) plays the key part in cell cycle arrest at the G1/S checkpoint in response to DNA damage, and is involved in the assembly of active cyclin-kinase complexes, in particular, cyclin D-Cdk4/6. Recent studies extended the range of known p21Waf1 functions. In addition to the cell-cycle control, p21Waf1 participates in important cell processes such as differentiation, senescence, and apoptosis. A balance of p21Waf1 functional activity seems to shift depending on the cell state (senescence, exposure to stress, expression of viral oncogenes). This is due to direct or indirect interaction with various modulators or to modification (phosphorylation, partial proteolysis) of p21Waf1. The review considers the structure of p21Waf1, its posttranslational modification, interactions with various cell or viral proteins, and their effects on the p21Waf1 function and the cell. PMID- 15285607 TI - [Modulation of serine/threonine protein kinase activity in chloramphenicol resistant mutants of Streptomyces avermitilis]. AB - A mutation to chloramphenicol resistance (Cmlr) stimulates production of macrolide avermectin in Streptomyces avermitilis; production starts in early stationary growth. By labeling in vivo, the Cmlr mutation was shown to stimulate phosphorylation of Ser and Thr in several proteins in the same growth phase. Autophosphorylation of active protein kinases (PK) was analyzed in gel after one- or two-dimensional PAGE for the original S. avermitilis strain ATCC 31272, its Cmlr mutant, and a Cmls revertant. An increase in in vivo phosphorylation was associated with an increase in autophosphorylation of Ser/Thr-PK 41K, 45K, 52K, 62K, and 85K and complete suppression of autophosphorylation of PK 66K. Comparison of the PK molecular weights and pI with the parameters deduced for putative PK encoded by S. avermitilis genes identified the 41K, 45K, 52K, 62K, and 85K PK as pkn 24, pkn 32, pkn 13, pkn 12, and pkn 5, respectively. Prenylamine lactate, a modulator of calmodulin-dependent processes, substantially reduced the avermectin production, impaired the Cml resistance, and selectively inhibited Ca2+-dependent PK 85K in the Cmlr mutant. It was assumed that PK 85K is involved in regulating the avermectin production. PMID- 15285608 TI - [Polymorphism of the serotonin 2A receptor gene (5HTR2A) and personality traits]. AB - The interindividual variation of temperament features (such as anxiety, neuroticism, harm avoidance) is determined, in particular, by allele polymorphism of genes involved in serotonin metabolism and has earlier been associated with the insertion/deletion polymorphism of the serotonin transporter gene. Polymorphic alleles of the serotonin 2A receptor gene (5HTR2A) were tested for association with personality traits assessed with several tests. The T102C and A1438G polymorphisms were associated with a variation in emotionality, activity, and sociability, which are integral characteristics of temperament. With each polymorphism, differences were significant only between heterozygotes and homozygotes. Carriers of T102C genotype A1/A2 displayed a lower level of anxiety related traits, a higher score on scale Hypomania, and a lower score on scale Social Introversion and were assumed to have higher activity and sociability. Carriers of A1428G genotype A/G differed from homozygotes G/G in having a lower level of social introversion and a lower score on scale No close friends, which testified to higher sociability of heterozygotes. Thus, the polymorphic alleles of SHTR2A proved to be associated with personality traits in mentally healthy people. PMID- 15285609 TI - [Cytoplasmic male sterility-associated structural variation of the mitochondrial genome regions containing rps3 and orf215 in sugar beet Beta vulgaris L]. AB - Several 5'-degenerate primers were selected by computer analysis and used for mtDNA typing in sugar beet cultivars with cytoplasms of the S (typical for cytoplasmic male sterility) or N (normal) type. A number of N- or S-specific markers were found to correspond to transcribed mitochondrial genes. One was from the orf215 region of the N-type mtDNA. A physical map of the corresponding region was constructed for the S-type mtDNA, and a substantial difference observed for the two genome types. One N-specific marker proved to contain a rearranged rps3 region and a truncated atp9 copy. With the known nucleotide sequence of this marker, three-primer PCR was designed and showed that both variants of the rps3 region simultaneously take place in the mtDNA pool, the new one occurring in a substochiometric proportion. PMID- 15285610 TI - [Editing of the mitochondrial cox3 transcript may yield a new site of protein protein interactions in wild cereal Elymus sibiricus L]. AB - With PCR, RT-PCR, and direct sequencing, complete nucleotide sequences were established for the Elymus sibiricus mitochondrial cytochrome c oxidase subunit 3 gene (cox3) and its cDNA. The cox3 transcript was shown to have 12 editing sites with changes affecting the amino acid sequence of the protein product. The editing of the primary cox3 transcript was found to change the position of a site of protein-protein interactions. The results demonstrate again the important role of mRNA editing in posttranscriptional regulation of the expression of plant mitochondrial genes. PMID- 15285611 TI - [Comparative restriction enzyme analysis of the genome in variola virus strains from the Russian collection]. AB - Comparative RFLP analysis was for the first time performed for 21 variola virus (VARV) strains of the Russian collection with 20 amplicons covering the total VARV genome. The amplicons were synthesized in the long polymerase chain reaction. A database useful as a reference for identifying VARV strains was generated. VARV strains isolated in different geographical regions were compared and proved to vary mostly in variable genome regions. Each of the dendrograms constructed included three clusters of African, Asian, and VARV-alastrim isolates. The VARV-alastrim isolates differed to the greatest extent from the other strains. VARV strains isolated during an ecdemic variola burst in Moscow (1960) grouped with Asian isolates. Polymorphism of VARV strains was for the first time observed for a single variola burst with a few affected patients. PMID- 15285612 TI - [The replication system of plasmids from Bacillus subtilis environmental isolates]. AB - Restriction enzyme analysis, cloning, and sequencing showed that large (more than 90-kb) plasmids isolated from different Bacillus subtilis strains are identical in structure of the region ensuring stable inheritance of plasmid replicons and are widespread in Belarussian environmental strains of B. subtilis. PMID- 15285613 TI - [Similarity of mutation spectra of the mitochondrial DNA hypervariable segment 1 in Homo and Pan species]. AB - The mutation spectrum of mtDNA hypervariable segment 1 (HVS1) was compared for east chimpanzee Pan troglodytes schweigfurthi and human. The two HVS1 had much the same nucleotide composition, and their mutation spectra were similar in major characteristics (substantial prevalence of transitions over transversions, pyrimidine transitions over purine ones, and C --> T over T --> C). DNA strand displacement (dislocation) during replication was identified as a major mechanism of context-dependent mutagenesis in human and chimpanzee mtDNAs. Nucleotide positions with mutations fitting the model of dislocation mutagenesis accounted for 21% of all variable positions in the chimpanzee HVS1. Variable motifs proved to be similar in the chimpanzee and human HVS1. Comparison of the Neanderthal and modern human HVS1 nucleotide sequences showed that most variable nucleotides are in DNA sites allowing context-dependent mutagenesis. PMID- 15285614 TI - [Analysis of chromosome translocations involving MML by hybridization with an oligonucleotide microarray]. AB - Identification of chromosome rearrangements is of importance for exact diagnosis, risk assessment, and therapy in blood malignancies. A new method was proposed for rapid and accurate identification of leukemia forms caused by chromosome rearrangements involving MLL (11q23). The method combines reverse transcription multiplex PCR and hybridization with an oligonucleotide microarray. The microarray was designed to detect the five most common MLL rearrangements: t(4;11) MLL/AF4, t(9;11) MLL/AF9, t(11;19) MLL/ELL, t(11;19) MLL/ENL, and dup(11) MLL/MLL. With clinical specimens, the method was shown to efficiently identify the chromosome translocations in leukemia patients. PMID- 15285615 TI - [The association of NAT2 polymorphisms with sporadic breast cancer]. PMID- 15285616 TI - [Phylogenetic analysis of alpha-galactosidases of the GH27 family]. AB - Amino acid sequence analysis of alpha-galactosidases and other proteins of glycoside hydrolase family 27 (GH27) allowed isolation of three major subfamilies, 27a-27c. Unique isomalto-dextranase of Arthrobacter globiformis clustered separately. Eukaryotic proteins formed five clusters on a phylogenetic tree of the family. Bacterial GH27 proteins, which are relatively few, did not form stable clusters. A monophyletic origin of the GH27 family was demonstrated with the use of related proteins of the GH36 family. The structure of the active center and evolution of alpha-galactosidases are discussed. PMID- 15285617 TI - [Expression of the mouse serotonin receptor (5HT1c) cDNA in cultured insect cells]. AB - Baculovirus-mediated cloning and expression of the mouse serotonin receptor (5HT1c) cDNA in insect cells was proposed to obtain an alternative to an oocyte based system, which is commonly employed in electrophysiological studies of ionic channels. A recombinant bacmid was constructed, and the 5HT1c cDNA transferred into the AcNPV genome to yield a recombinant baculovirus. Infected inset Sf9 cells produced recombinant 5HT1c. PMID- 15285618 TI - [Identification of signal sequences determining the specific nucleolar localization of fibrillarin in HeLa cells]. AB - Fibrillarin is one of the major nucleolar proteins and is involved in pre-rRNA maturation. Its three main regions are a glycine and arginine-rich (GAR) domain, an RNA-binding domain, and an alpha-helical region, which presumably has a methyltransferase activity. Yet the roles of these regions in nucleolus-specific localization of fibrillarin are still unclear. To elucidate this issue, a series of plasmids was constructed to express human fibrillarin mutants fused with the green fluorescent protein. Localization of the chimeric proteins was studied in interphase and mitotic HeLa cells after single transfection with the plasmids. Deletion or a mutation of any domain proved to alter the specific fibrillarin location coinciding with sites of pre-rRNA synthesis. The GAR domain and the first spacer together were sufficient for fibrillarin migration into the nucleolus. Fibrillarin mutants located within the interphase nucleolus did not differ in mitotic location from the wild-type fibrillarin. PMID- 15285619 TI - [Protein environment of the sense codon of the template in the A site of the human ribosome as inferred from crosslinking to oligoribonucleotide derivatives]. AB - The protein environment of each nucleotide of the template codon located in the A site of the human ribosome was studied with UUCUCAA and UUUGUU derivatives containing a Phe codon (UUC and UUU, respectively) and a perfluoroarylazido group at U4, U5, or U6. The analogs were positioned in the ribosome with the use of tRNA(Phe), which is cognate to the UUC or UUU codon and directs it to the P site, bringing a modified codon in the A site with a modified nucleotide occupying position +4, +5, or +6 relative to the first nucleotide of the P-site codon. On irradiation of ribosome complexes with tRNA(Phe) and mRNA analogs with mild UV light, the analogs crosslinked predominantly to the 40S subunit, modifying the proteins to a greater extent than the rRNA. The 18S rRNA nucleotides crosslinking to the analogs were identified previously. Of the small-subunit proteins, S3 and S15 were the major targets of modification in all cases. The former was modified both in ternary complexes and in the absence of tRNA, and the latter, only in ternary complexes. The extent of crosslinking of mRNA analogs to S15 decreased when the modified nucleotide was shifted from position +4 to position +6. The results were collated with the data on ribosomal proteins located at the decoding site of the 70S ribosome, and conclusion was made that the protein environment of the A-site codon strikingly differs between bacterial and eukaryotic ribosomes. PMID- 15285620 TI - [The first nucleotide of the codon located in the P site of the 80S ribosome is close to G1702 of the 18S rRNA as revealed by crosslinking to a pUUUGUU derivative containing a perfluorophenylazido group at the guanine]. AB - Modification of the 18S rRNA with a pUUUGUU derivative carrying a perfluorophenylazido group at N7 of G was studied in the complex with the human 80S ribosome and Val-tRNA(Val), which directs modified GUU to the P site. Reverse transcription reported modification of invariant G1702 of the 18S rRNA. On evidence of the results and the earlier data on affinity modification of the human ribosome with tetra- or heptaribonucleotide derivatives carrying an alkylating group at the 3' end, the template was assumed to make a bend between the A- and P-site codons, which brings both codons closer to G1702 of the 18S rRNA. PMID- 15285621 TI - [The effect of Clp proteins on DnaK-dependent refolding of bacterial luciferases]. AB - A study was made of the refolding of bacterial luciferases of Vibrio fischeri, V. harveyi, Photobacterium phosphoreum, and Photorhabdus luminescens. By reaction rate, luciferases were divided into two groups. The reaction rate constants of fast luciferases of V. fischeri and Ph. phosphoreum were about tenfold higher than those of slow luciferases of Ph. luminescens and V. harveyi. The order of increasing luciferase thermostability was Ph. phosphoreum, V. fischeri, V. harveyi, and Ph. luminescens. The refolding of thermoinactivated luciferases completely depended on the active DnaK-DnaJ-GrpE chaperone system. Thermolabile fast luciferases of V. fischeri and Ph. phosphoreum showed highly efficient rapid refolding. Slower and less efficient refolding was characteristic of thermostable slow luciferases of V. harveyi and Ph. luminescens. Chaperones of the Clp family were tested for effect on the efficiency of DnaK-dependent refolding of bacterial luciferases in Escherichia coli cells. The rate and extent of refolding were considerably lower in the clpB mutant than in wild-type cells. In E. coli cells with mutant clpA, clpP, of clpX showed a substantially lower luciferase refolding after heat shock. PMID- 15285622 TI - [Survival strategy of photosynthetic organisms. 1. Variability of the extent of light-harvesting pigment aggregation as a structural factor optimizing the function of oligomeric photosynthetic antenna. Model calculations]. AB - In accordance with our concept of rigorous optimization of photosynthetic machinery by a functional criterion, this series of papers continues purposeful search in natural photosynthetic units (PSU) for the basic principles of their organization that we predicted theoretically for optimal model light-harvesting systems. This approach allowed us to determine the basic principles for the organization of a PSU of any fixed size. This series of papers deals with the problem of structural optimization of light-harvesting antenna of variable size controlled in vivo by the light intensity during the growth of organisms, which accentuates the problem of antenna structure optimization because optimization requirements become more stringent as the PSU increases in size. In this work, using mathematical modeling for the functioning of natural PSUs, we have shown that the aggregation of pigments of model light-harvesting antenna, being one of universal optimizing factors, furthermore allows controlling the antenna efficiency if the extent of pigment aggregation is a variable parameter. In this case, the efficiency of antenna increases with the size of the elementary antenna aggregate, thus ensuring the high efficiency of the PSU irrespective of its size; i.e., variation in the extent of pigment aggregation controlled by the size of light-harvesting antenna is biologically expedient. PMID- 15285623 TI - [Survival strategy of photosynthetic organisms. 2. Experimental proof of the size variability of the unit building block of light-harvesting oligomeric antenna]. AB - The present series of papers is part of an integrated research program to understand the effective functional strategy of native light-harvesting molecular antennae in photosynthetic organisms. This work tackles the problem of the structural optimization of light-harvesting antennae of variable size. In vivo, the size responds to the illumination intensity, thus implying more sophisticated optimization strategies, since larger antenna size demands finer structural tuning. Earlier modeling experiments showed that the aggregation of the antenna pigments, apart from being itself a universal structural factor of functional antenna optimization with any (!) spatial lattice of light-harvesting molecules, determines the antenna performance provided that the degree of aggregation varies: the larger the unit building block, the higher the efficacy of the whole structure. It means that altering the degree of pigment aggregation in response to the antenna size is biologically expedient. In the case of the oligomeric chlorosomal antenna of green bacteria, the strategy of variable antenna structural optimization in response to the illumination intensity was demonstrated to take place in vivo and facilitate high antenna performance regardless of its size, thus allowing bacteria to survive in diverse illumination conditions. PMID- 15285624 TI - [Studies of alpha- and betaL-crystallin complex formation in solution at 60 degrees C]. AB - Studies of molecular mechanisms of chaperone-like activity of alpha-crystallin became an active field of research over last years. However, fine interactions between alpha-crystallin and the damaged protein and their complex organization remain largely uncovered. Complexation between alpha- and betaL-crystallins was studied with thermal denaturation of betaL-crystallin at 60 degrees C using small angle X-ray scattering (SAXS), light scattering, gel-permeation chromatography and electrophoresis. A mixed solution of alpha- and betaL-crystallins in concentrations about 10 mg/ml incubated at 60 degrees C was found to contain their soluble complexes with mean radius of gyration approximately 14 nm, mean molecular weight approximately 4000 kDA and maximal size approximately 40 nm. In pure betaL-crystallin solution, complexes were not observed at 60 degrees C. In SAXS studies, transitions in the alpha-crystallin quaternary structure at 60 degrees C were shown to occur and result in a double increase of the molecular weight. It suggests that during the temperature-induced denaturation of betaL crystallin it binds with modified alpha-crystallin or, alternatively, alpha-betaL crystallin complexation and alpha-crystallin modifications are concurrent. Estimates of the alpha-betaL-crystallin dimensions and relative contents of alpha and betaL-crystallins in the complex suggest that several alpha-crystallin molecules are involved in complex formation. PMID- 15285625 TI - [Binding of unnatural alpha,beta-oligocytidylates with DNA duplexes]. AB - Binding of short fluorescently labeled AT-containing DNA duplexes with modified oligocytidylates is studied. The latter are modified to contain unnatural alpha anomers along with natural beta-nucleotides; the nucleotide composition is selected according to putative pattern of unconventional triplex formation between duplex and oligomer bases. Nondenaturing gel electrophoresis is used to study complexation of fluorescent duplexes with cytidyl oligomers and oligocytidylate self-association at low temperatures. A DNA duplex of random AT composition is shown to bind with an excess of the corresponding oligocytidylate in 0.1 M Tris-HCl in the presence of Mg2+. Binding is observed at neutral pH values, while more basic pH (8.0) prevents complexation of the AT duplex and oligocytidylate. Contrary to oligonucleotides of irregular composition, a regular dA30:dT30 duplex does not bind with the dC strand. It is also shown that alternating self-complementary duplex d(AT)16 and oligocytidylate d(CbetaCalpha)15 do not form complexes, and poly-dC self-associates are formed instead. The effect of 2'-O-methylation of the third strand on complex formation and self-association is also analyzed. The results suggest that a modified oligocytidylate binds with a random-composition duplex, albeit with lower efficiency. PMID- 15285627 TI - [Terrorism and mental health (problem's scale, population tolerance, management of care)]. AB - The consequences of terrorist threat and terrorist acts for mental health of the individual, groups of individuals and community in general are analyzed. Mental disorders emerging in the victims of terrorism is described. The problem of terrorist threats use as a psychic weapon is discussed. Tolerance of population to terrorism can be divided into two types--psychophysiological and socio psychological. The ways for elevating tolerability to terrorist threat and terrorist acts are suggested. Help in the centers of terrorist act must be of the complex character, being provided by different specialists including psychologists and psychiatrists. The importance of state structures and community support in this work is emphasized. PMID- 15285626 TI - [RNA-binding properties of proteins of the beet yellows closterovirus]. AB - Recombinant p64, p65, p24, p22, p21 of the beet yellows closterovirus and pcp, hel, mtr, and pol fragments encoded by the replication genes of the virus were purified and tested for RNA binding. North-Western blotting revealed the RNA binding activity for p64 and hel a 21-kDa fragment of the helicase domain with conserved motifs V and VI. Gel retardation assay confirmed hel binding with a randomized RNA probe in vitro, and a cooperative RNA-hel interaction was assumed on evidence of the binding pattern. The RNA-hel complexes proved to be stable at a high ionic strength. PMID- 15285628 TI - [The peculiarities of panic disorders determined by gender dimorphism]. AB - In contrast to panic disorder (PD) epidemiology, gender-related etiological and clinical PD peculiarities are not studied well so far. Seventy-five PD patients (27 male, 48 female) with and without agoraphobia diagnosed according to ICD-10 criteria have been studied. No differences were found by gender for the key clinical signs: interrelations in parent family, age at debut, panic attacks frequency, combination with agoraphobia, social maladaptation and response to antidepressant therapy. Along with common features, we revealed gender differences in the clinical picture. In women, neuroendocrinal and motivation disorders preceded PD debut, panic attacks comprised more symptoms, with autonomic symptoms in the attack being subjectively considered as the key ones, autonomic function was more pronounced in between-attack period and depressive symptoms predominated in the clinical picture. Men considered psychic anxious symptoms (fear of death, forcing a patient to seek an outside assistance) as more prominent, and PD in men was more often combined with hypochondria and alcohol abuse. PMID- 15285629 TI - [The age peculiarities of nonpsychotic forms of psychogenic disorders caused by everyday stressors]. AB - Three groups of patients were examined in the period of 1994-1999: adolescent (aged 15-18 years), middle-aged (25-59 years) and elderly (60 years and older) with non-psychotic mental disorders developing under the influence of everyday stressors. The highest tolerability to stressors was detected in the middle-aged group. The dominating stressors in the groups were: social situations in the adolescences; interfamily conflicts--in the middle-aged patients and a presence of disease--in the elderly. Stressogenic affective disorders occurred most often in all the groups; anxious-phobic states prevailed in adolescents; neurotic pictures with hysteri-form symptoms, converse type--in the middle age and anxious phobic disorders with primary psychoorganic appearances--in the elderly. Risk factors for psychogenic disorders are: male sex, "organic predisposition", accentuation of the personality, "destructive" types of family bringing up in the adolescences; female sex, signs of endogenic diathesis and reactive lability for the middle-aged patients and female sex, aging factor and related changes of personality as well as somatic diseases in the elderly. PMID- 15285630 TI - [Risk and prevention of atrial fibrillation of non-valvular origin]. AB - Atrial fibrillation (AF) is an independent risk factor for ischemic stroke. In patients with AF, cardioembolias present about 10% of ischemic strokes. Transesophageal echocardiography is an ideal instrument for diagnostics of intracardiac thrombi. An aim of the study was to find the high risk markers for stroke in patients with AF of non-valvular origin. The patients have been divided into 2 groups with and without stroke in anamnesis. To search for stroke dependence of clinical and echocardiographic high risk markers, the data were analyzed using Poly Analyst Power statistical package. In the group of the patients with stroke in the anamnesis, echocardiographic markers for high risk of thromboembolia occurred significantly more frequently. Thrombi in the left atrial or its appendage were registered in 12.5% patients without stroke in anamnesis and in 31% of those, who survived stroke. The independent risk factors for stroke were age, AF duration, left ventricular ejection fraction, diabetes mellitus and arterial hypertension. PMID- 15285631 TI - [Influence of long-term quetiapine (Seroquel) and haloperidol therapy on cognitive deficit in patients with paranoid schizophrenia]. AB - In of 67 patients with ICD-10 diagnosis of paranoid schizophrenia compared influence of atypical neuroleptic Seroquel (31 patients) and typical neuroleptic haloperidol (36 patients) on cognitive disturbances. The latter were evaluated using 8 neuropsychological tests and clinical symptoms were measured by the PANSS. There were 39 patients with the first episode and 28 with a chronic disease course. A mean therapeutic dosage for Seroquel was 316.21 mg/day, for haloperidol--12.34 mg/day, treatment duration was 3-12 months. The effect was evaluated 1, 3, 6 and 12 months after the beginning of the treatment. Compared to haloperidol, Seroquel proved to be more effective for all the indices studied, in particular for executive functions and verbal productivity that correlated with negative symptoms reduction measured with the PANSS. Cognitive functions related to positive symptoms (attention, verbal memory) improved in both groups. Seroquel positively affected motor functions. The study confirmed the earlier reports on possibility of neurocognitive deficit correction by atypical antipsychotic drugs in the first episode as well as in chronic patients with schizophrenia. PMID- 15285632 TI - [Use of cholinomimetics in the treatment of endogenous autism in children]. AB - Twenty children with endogenous autism of mild and moderate severity (30-44.5 scores according to the CARS), aged 3-8 years, were treated with choline alfoscerate (CA), 400 mg/day, during 8 weeks in the presence of maintenance therapy with neuroleptics (17 cases). Positive therapeutic effect was observed in 89% of the patients: significant improvement--in 61% and minimal efficacy--in 28%. Statistically significant positive changes in the patient's state were observed in the general improvement of behavior (p<0.001), development of social and communicative skills, as well as self-service, reduction of marked speech disturbances (p<0.001) and motor sphere (p<0.001), enhancement of learning activity and productivity (p<0.05). Good tolerability to the therapy, without patient's state worsening was registered. Some patients exhibited strengthening of affective lability in the first weeks of the treatment which attenuated to the 4th week as the CA dosages decreased to 400 mg every other day. CA may be recommended for combined therapy with neuroleptics as an effective and safe medicine for the treatment of cognitive and behavioral disorders in patients with children's autism. PMID- 15285633 TI - [Dynamics of spatial synchronization of brain biopotentials in intensive attention during hypnotic state]. AB - Twelve psychiatrically normal women aged 21-38 years were examined in awakened and hypnotic state using an EEG method with evaluation of spatial synchronization of brain biopotentials (SSBB). Suggesting of high intensive attention to the subjects in hypnotic state leads to substantial reorganization of SSBB manifesting as SSBB increase between both occipital regions, right temporal and other brain cortex areas. Dynamics of brain SSBB in intensive attention in awakened and hypnotic state is of opposite character that is likely to be the result of temporary switching off of the functions of the frontal brain cortex areas, providing the conscious control and regulation of the current activity. PMID- 15285634 TI - [Epilepsy in East Siberia]. PMID- 15285635 TI - [Prognostic value of some spectral EEG indices in patients in acute period of brain supratentorial hemorrhagic stroke]. PMID- 15285636 TI - [Intrabrain hemorrhage in hemophilia]. PMID- 15285637 TI - [Children's aphasias and Landau-Kleffner syndrome in the light of brain plasticity]. PMID- 15285639 TI - [On the activity of dissertation boards on "nervous diseases" specialty, in 1998 2002]. PMID- 15285638 TI - [The features of atypical neuroleptic amisulpride action]. PMID- 15285640 TI - Construction of a novel Mo/Cu/S cluster with a closed double-cubane-like polyhedron and a chain polymer of W/Cu/S clusters. AB - Reaction of [MoOS(3)](2)(-) and [WS(4)](2)(-) with Cudtp (dtp = diethyl dithiophosphate) gave rise to the clusters [Bu(4)N](2)[(MoOS(3))(4)Cu(12)(dtp)(6)], 1, and [Et(4)N][(WS(4)Cu(4))(dtp)(3)], 2, respectively. In cluster 1, the dtp- ligands act as both monodentate and bidentate ligands that bridge between Cu atoms and link together a closed double cubane-like [Mo(2)O(2)S(6)Cu(6)](2+) core and two incomplete cubane-like [MoOS(3)Cu(3)]+ units. In cluster 2, the [WS(4)Cu(4)](2+) fragments were connected via bidentate and doubly bridging dtp- bridges to give a chain polymeric anion. Cluster 1 is the first example of a Mo/Cu/S cluster that contains a closed double-cubane-like structure. Compound 2 is also rare and the first W/Cu/S polymer with dtp- linkages. PMID- 15285641 TI - Control of oxygen atom chirality and chelate ring conformation by protected/free sugar hydroxyl groups in glucose-pendant dipicolylamine-copper(II) complexes. AB - A pair of copper(II) complexes 1 and 2 exhibit an enantiomeric chiral center at the oxygen atom that coordinates to the metal center. The configurations of the oxygen atom chirality and the chelate ring conformation are simply controlled by protected/free hydroxyl groups of the sugar moiety, yielding mirror image CD spectra. In this system, repulsive and attractive forces are used to regulate chirality on the copper-coordinated oxygen atom both in the solid state and in solution. PMID- 15285642 TI - Structural characterization and luminescence behavior of a silver(I) 1D polymeric chain constructed via a Bridge with unusual 4,5-diazospirobifluorene and perchlorate. AB - A new polymeric silver complex, [Ag(2)(L(2))(ClO(4))(2)] (L = 4,5 diazospirobifluorene), has been synthesized and shown to exhibit interesting luminescence properties in a single crystal. Structural analysis reveals a one dimensional chain, which contains a [Ag(2)(L(2))](2+) dimer bridged with ClO(4)( ). The Ag...Ag distances are 2.776(1) and 4.575(1) A incorporated by two L ligands and by a ClO(4)(-) bridge, respectively. PMID- 15285643 TI - [NH4]2Mn3(H2O)4[Mo(CN)7]2.4H2O: tuning dimensionality and ferrimagnetic ordering temperature by cation substitution. AB - [NH(4)](2)Mn(3)(H(2)O)(4)[Mo(CN)(7)](2).4H(2)O (1) has been synthesized by slow diffusion of aqueous solutions containing K(4)[Mo(CN)(7)].2H(2)O, [Mn(H(2)O)(6)](NO(3))(2), and (NH(4))NO(3). Compound 1 crystallizes in the monoclinic C2/c space group. The basic motif of the three-dimensional structure consists of a Mo1-Mn1 gridlike sheet parallel to the bc plane. Two of these sheets are connected through CN-Mn2-NC linkages to form a bilayer reminiscent of the K(2)Mn(3)(H(2)O)(6)[Mo(CN)(7)](2).6H(2)O (2) two-dimensional structure. In 1, [NH(4)](+) cations allow these bilayers to be connected through direct Mo1-CN-Mn1 bridges to form a three-dimensional network, whereas in 2, they are isolated by (H(2)O)K(+) cations. As shown by the magnetic measurements, this increase of dimensionality by counterion substitution induces an enhancement of the ferrimagnetic critical temperature from 39 K in 2 to 53 K in 1. PMID- 15285644 TI - A novel mu(4)-oxo bridged copper tetrahedron derived by self-assembly: first example of double helical bis(tridentate) coordination of a hexadentate amine phenol ligand. AB - In methanol, the reaction of Cu(ClO(4))(2).6H(2)O and the hexadentate amine phenol ligand (H(2)bahped) in the presence of triethylamine affords a tetranuclear copper(II) complex having the formula [Cu(4)(mu(4) O)(bahped)(2)](ClO(4))(2). The X-ray structure of this complex shows a tetrahedral central [Cu(II)(4)(mu(4)-O)]unit coordinated to two hexadentate bridging (via the central ethylenediamine part) ligands. The compound is the first example of a mu(4)-oxo tetranuclear copper(II) complex without any bridging ligand along the six tetrahedral edges. Variable-temperature magnetic data clearly show an S(t) = 0 spin ground state for antiferromagnetic interactions between four (2)B(2) copper(II) ions in a dimer of dimers. PMID- 15285645 TI - A Mn12 single-molecule magnet [Mn12O12(OAc)12(dpp)4] (dppH = diphenyl phosphate) with no coordinating water molecules. AB - The preparation and physical characterization are reported for a novel single molecule magnet [Mn(12)O(12)(OAc)(12)(dpp)(4)] (dppH = diphenyl phosphate) with no coordinating water molecules. The crystal structure analysis reveals that there are four five-coordinate Mn(III) ions with Mn.H approaches. Addition of water in CD(2)Cl(2) solution was monitored by (1)H NMR, which showed that H(2)O could coordinate to a vacant site of a five-coordinate Mn(III) ion in solution. The measurements and analyses of magnetization hysteresis and ac magnetic susceptibility indicate that the title complex is a single-molecule magnet with a quantum tunneling behavior, whose ground state was tentatively assigned to S = 10 with g = 1.78 and D = -0.60 K. PMID- 15285647 TI - Synthesis of novel amorphous boron carbonitride ceramics from the borazine derivative copolymer via hydroboration. AB - A preceramic polymer for boron carbonitride was synthesized for the first time by the hydroboration of borazine derivatives, B-triethynyl, N-trimethyl borazine and borazine, without a catalyst. A homogeneous, amorphous boron carbonitride ceramic was prepared by the thermolysis of a hydroborated copolymer in an argon atmosphere. PMID- 15285646 TI - Comparison of cysteine and penicillamine ligands in a Co(II) maquette. AB - l-Penicillamine (Pen) has been investigated as a ligand for metalloprotein design by examining the binding of Co(II) to the sequence NH(2) KL(Pen)EGG.(Pen)IG(Pen)GA(Pen).GGW-CONH(2). For comparison, we have studied Co(II) binding to the analogous sequence with Cys ligands, the ferredoxin maquette ligand IGA that was originally designed to bind a [4Fe-4S] cluster. The Co(II) affinity and UV-vis spectroscopic properties of IGA indicate formation of a pseudotetrahedral tetrathiolate ligated Co(II). In contrast, IGA-Pen showed formation of a pseudotetrahedral complex with Co(II) bound by three Pen ligands and an exogenous H(2)O. EXAFS data on both Co(II) complexes confirms not only the proposed primary coordination spheres but also shows six Co(II)-C(beta) methyl group distances in Co(II)-IGA-Pen. These results demonstrate that ligand sterics in simple peptides can be designed to provide asymmetric coordination spheres such as those commonly observed in natural metalloproteins. PMID- 15285648 TI - Reaction of AgN3 with SOCl2: evidence for the formation of thionyl azide, SO(N3)2. AB - Pure thionyl azide SO(N(3))(2), which is the only gaseous reaction product, has been generated in a vacuum by the heterogeneous reaction of SOCl(2) vapor with AgN(3) at room temperature at a SOCl(2) vapor pressure of 1 x 10(-3) Torr. Evidence for the formation of SO(N(3))(2) is given by on line photoelectron spectroscopy (PES) combined with outer valence Green's function (OVGF) calculations with the 6-311++G(2df) basis set. The good agreement between the PES experiment and the OVGF calculation shows that SO(N(3))(2) has C(1) symmetry. The first ionization energy of SO(N(3))(2) is 10.18 eV. PMID- 15285649 TI - Coordination chemistry of diselenophosphinate complexes: the X-ray single-crystal structures of [K(Se2PPh2)(THF)2]2 and [In(Se2PPh2)3].L (L = THF, PhMe). AB - Reaction of potassium diphenylphosphide with elemental selenium is shown to give [K(Se(2)PPh(2))(THF)(2)](2) 1, which further reacts with InCl(3) to yield [In(Se(2)PPh(2))(3)] 2. Crystallization of 2 from either THF or PhMe gave 2.THF or 2.PhMe, respectively, both of which form loosely linked dimers in the solid state via Se...Se intermolecular van der Waals interactions. Decomposition of 2 has been studied by TGA. PMID- 15285650 TI - Kinetics evidence for a complex between peroxynitrous acid and titanium(IV). AB - The reaction between TiO(2+) and ONOOH in 0.9 M H(2)SO(4) provides evidence for direct formation, previously unobserved, of a HOONO-metal complex. The reaction proceeds via formation of an end-on complex (k = 3.0 x 10(2) M(-1) s(-1)) that rearranges to form a side-on complex (k approximately equal to 20 s(-1)). With ONOOH in excess, this rearrangement proceeds more slowly (k approximately equal to 0.1 s(-1)), probably because multiple hydrogen oxoperoxonitrate molecules form end-on complexes with oxotitanium(IV) and hinder rearrangement to the side-on complex. The absorption spectrum of the final product is that of TiO(2)(2+). Presumably, during the rearrangement or later, NO+ is lost. PMID- 15285651 TI - Incommensurate nature of the multilayered molecular ferromagnetic metals based on bis(ethylenedithio)tetrathiafulvalene and bimetallic oxalate complexes. AB - The salt [ET]x[MnRh(ox)(3)].CH(2)Cl(2) (x = 2.526(1)) has been obtained and characterized. This paramagnetic metal is essentially isostructural to the ferromagnetic metal [ET]y[MnCr(ox)(3)].CH(2)Cl(2) (y approximately equal to 3) and provides a definite answer on the origin of the structural disorder present on such systems. As in the ferromagnetic analogue, this material shows high electrical conductivity at room temperature (13 S.cm(-1)) and metallic behavior. PMID- 15285652 TI - Grid-type two-dimensional magnetic multinuclear metal complex: strands of ([CuII(mu-4,4'-bpy)]2+)n cross-linked by octacyanotungstate(V) ions. AB - Reaction of the preorganized strands of ([Cu(II)(mu-4,4'-bpy)](2+))n (4,4'-bpy = 4,4'-bipyridine) with [W(V)(CN)(8)](3)(-) leads to a novel cyano-bridged Cu(II)(3)W(V)(2) complex [Cu(mu-4,4'-bpy)(DMF)(2)][Cu(mu-4,4' bpy)(DMF)](2)[W(V)(CN)(8)](2).2DMF. 2H(2)O 1. The structure of 1 consists of the expected 2-dimensional grid-type network which is built of infinite ([Cu(II)(mu 4,4'-bpy)](2+))n chains cross-linked by octacyanotungstate units. The Cu(II)-NC W(V)-CN-Cu(II) linkage exhibits the topology of a 3,2-chain. The skeleton of the layer is additionally stabilized by a hydrogen bond network formed by terminal cyano ligands of the [W(CN)(8)](3-) moiety and water molecules. The distance between the adjacent Cu(3)(II)W(2)(V) chains within the layer is 11.12 A along the a axis. The layers are connected by H-bonds of NCN-NDMF-NCN linkages into 3-D supramolecular architecture. The magnetic properties correspond to a dominant ferromagnetic coupling within the Cu(II)(3)W(V)(2) pentamer units (J = +35(4) cm( 1)) and much weaker effective AF interunit coupling which include both intra- and inter-3,2-chain interactions between pentamers (J' = -0.05(1) cm(-1)). PMID- 15285653 TI - Unprecedented migration of a methyl group in 2-(2',6'-dimethylphenylazo)-4 methylphenol mediated by ruthenium. AB - An unprecedented chemical transformation of 2-(2',6'-dimethylphenylazo)-4 methylphenol has been observed upon its reaction with [Ru(PPh(3))(2)(CO)(2)Cl(2)] whereby the methyl group at the 2' position migrates to the 4' or 6' position. PMID- 15285654 TI - Chiral induction in a ribose-decorated metallostar through intrinsic and interionic diastereomeric interactions. AB - A ribose-functionalized bpy ligand has been prepared and shown to give modest diastereomeric excesses of Lambda-[FeL(3)](2+) complexes; interconversion of Delta and Lambda cations is relatively fast, and in CHCl(3), the favored complexes with Delta- or Lambda-TRISPHAT counterions are homochiral, (Delta(+)Delta(-)) or (Lambda(+)Lambda(-)). In the case of the Delta-TRISPHAT salt, a single diastereomer is observed (de > or = 96%). PMID- 15285655 TI - Imine hydrolysis and role of a rhodium(I)-imine-amine complex in homogeneous H2 hydrogenation of the imine and a rare example of inequivalent NH2 protons. AB - A Rh-catalyzed, homogeneous hydrogenation of the imine, PhCH(2)N=CHPh, is shown to involve a Rh-imine-amine species that subsequently activates H(2), the amine (benzylamine) being formed via a Rh-catalyzed hydrolysis of the imine by adventitious water. The imine-amine complex, cis-(Rh[P(p tolyl)(3)](2)(PhCH(2)N=CHPh)(NH(2)CH(2)Ph))PF(6) (2b), is structurally characterized, and the solution (1)H NMR data reveal inequivalent NH(2) protons. PMID- 15285656 TI - Synthesis, structural, spectroscopic, and electrochemical characterization of high oxidation state diruthenium complexes containing four identical unsymmetrical bridging ligands. AB - Six Ru2(6+) derivatives of the form Ru2(L)4(C[triple bond]CC6H5)(2), where L = 2 Fap, 2,3-F(2)ap, 2,4-F(2)ap, 2,5-F(2)ap, 3,4-F(2)ap, or 2,4,6-F(3)ap, are synthesized and characterized as to their electrochemical, spectroscopic, and/or structural properties. These compounds are synthesized from a reaction between LiC[triple bond]CC6H5 and Ru2(L)4Cl. Two of the investigated complexes exist in a (4,0) isomeric form while four adopt a (3,1) geometric conformation. These two series of geometric isomers are compared with previously characterized (4,0) Ru2(ap)4(C[triple bond]CC6H5)(2), (4,0) Ru2(F5ap)4(C[triple bond]CC6H5)(2), and (3,1) Ru2(F5ap)4(C[triple bond]CC6H5)(2). The overall data on the nine compounds thus provide an opportunity to systematically examine how the electrochemical and structural properties of these Ru2(6+) complexes vary with respect to isomer type and electronic properties of the bridging ligands. PMID- 15285657 TI - New compounds from tellurocyanide rhenium cluster anions and 3d-transition metal cations coordinated with ethylenediamine. AB - The compounds [Ni(en)(3)](2)[Re(6)Te(8)(CN)(6)].10H(2)O (1), [Ni(NH(3))(4)(en)](2)[Re(6)Te(8)(CN)(6)].2H(2)O (2), [Ni(NH(3))(2)(en)(2)][(Ni(en)(2))(3)(Re(4)Te(4)(CN)(12))(2)].38H(2)O (3), [Co(NH(3))(2)(en)(2)](2)[(Co(en)(2))Re(6)Te(8)(CN)(6)]Cl(2).H(2)O (4),and [(Zn(H(2)O)(en)(2))(Zn(en)(2))Re(6)Te(8)(CN)(6)].3H(2)O (5) (en = ethylenediamine) have been synthesized and characterized. Compounds 1, 4, and 5 have been synthesized by the diffusion of an aqueous (for 1 and 5) or an ammonia (for 4) solution of Cs(4)[Re(6)Te(8)(CN)(6)].2H(2)O into a glycerol solution of NiCl(2).6H(2)O (for 1), CoCl(2).6H(2)O (for 4), or ZnCl(2) (for 5). Compounds 2 and 3 have been synthesized by the reaction of an aqueous solution of Cs(4)[Re(6)Te(8)(CN)(6)].2H(2)O (for 2) or K(4)[Re(4)Te(4)(CN)(12)].5H(2)O (for 3) with an ammonia solution of Ni(en)(2)Cl(2). Compounds 1 and 2 are ionic whereas compounds 4 and 5 are one-dimensional polymers. Compound 3, a two dimensional polymer, possesses hexagonal shaped channels of approximate diameter 10-12 A. Because the framework of compound 3 is robust, it is an attractive host for guest molecules of appropriate size and shape. The potential "guest" volume is about 37% of the unit cell volume. PMID- 15285658 TI - Two series of novel rare earth complexes with dicyanamide [Ln(dca)2(phen)2(H2O)3][dca].(phen), (Ln = Pr, Gd, and Sm) and [Ln(dca)3(2,2' bipy)2(H2O)]n, (Ln = Gd, Sm, and La): syntheses, crystal structures, and magnetic properties. AB - Two series of novel complexes, [Ln(dca)(2)(Phen)(2)(H(2)O)(3)](dca).(phen) (Ln = Pr (1), Gd (2), and Sm (3), dca = N(CN)(-), phen = 1,10-phenanthroline) and [Ln(dca)(3)(2,2'-bipy)(2)(H(2)O)](n), (Ln = Gd (4), Sm (5), and La (6), 2,2'-bipy = 2,2'-bipydine), have been synthesized and structurally characterized by X-ray crystallography. The crystal structures of the first series (1-3) are isomorphous and consist of discrete [Ln(dca)(2)(Phen)(2)(H(2)O)(3)]+ cations, dca anions, and lattice phen molecules; whereas the structures of the second series (4-6) are characterized by infinite chains [Ln(dca)(3)(2,2'-bipy)(2)(H(2)O)](n). The Ln(III) atoms in all complexes are nine-coordinated and form a distorted tricapped trigonal prism environment. The three-dimensional frameworks of 1-6 are constructed by intermolecular hydrogen bond interactions. Variable-temperature magnetic susceptibility measurements for complexes 1, 2, 4, and 5 indicate a Curie-Weiss paramagnetic behavior over 5-300 K. PMID- 15285659 TI - Mechanism of abstraction reactions of dimetallenes (R2X=XR2; X = C, Si, Ge, Sn, Pb) with halocarbons: a theoretical study. AB - Potential energy surfaces for the abstraction reactions of dimetallenes with halocarbons have been studied using density functional theory (B3LYP). Five dimetallene species, (SiH(3))(2)X=X(SiH(3))(2), where X = C, Si, Ge, Sn, and Pb, have been chosen in this work as model reactants. The present theoretical investigations suggest that the relative dimetallenic reactivity increases in the order C=C << Si=Si < Ge=Ge < Sn=Sn < Pb=Pb. That is to say, for halocarbon abstractions there is a very clear trend toward lower activation barriers and more exothermic reactions on going from C to Pb. Moreover, for a given dimetallene, the overall barrier heights are determined to be in the order CF(4) > CCl(4) > CBr(4) > CI(4). That is, the heavier the halogen atom (Y), the more facile its abstraction from CY(4). Halogen abstraction is always predicted to be much faster than the abstraction of a CY(3) group irrespective of the dimetallene or halocarbon involved. Our model conclusions are consistent with some available experimental findings. Furthermore, both a configuration mixing model based on the work of Pross and Shaik and bonding dissociation energies can be used to rationalize the computational results. These results allow a number of predictions to be made. PMID- 15285660 TI - Theoretical analysis of the Jahn-Teller distortions in tetrathiolato iron(II) complexes. AB - Crystallographic studies of [Fe(SR)(4)](2-) (R is an alkyl or aryl residue) have shown that the Fe(II)S(4) cores of these complexes have (pseudo) D2d symmetry. Here we analyze the possibility that these structures result from a Jahn-Teller (JT) distortion that arises from the e(3z(2) - r(2), x(2) - y(2)) orbital ground state of Fe(II) in T(d)symmetry. Special attention is paid to the influence of the second-nearest neighbors of Fe, which lowers the symmetry and reduces the full JT effect to a smaller, pseudo JT effect (PJT). To estimate the size of the PJT distortion, we have determined the vibronic parameters and orbital state energies for a number of [Fe(SR)(4)](2-) models using density functional theory (DFT). Subsequently, this information is used for evaluating the adiabatic potential surfaces in the space of the JT-active coordinates of the FeS(4) moiety. The surfaces reveal that the JT effect of Fe(II) is completely quenched by the tetrathiolate coordination. PMID- 15285661 TI - Theoretical analysis of the three-dimensional structure of tetrathiolato iron complexes. AB - The three-dimensional structures of a number of [M(SR)(4)](n-) complexes, where M is a 3d transition metal and R is an alkyl or aryl group, have been analyzed using density functional theory (DFT). Special attention is paid to the Fe(II)/Fe(III) mimics of rubredoxin. The Fe(II) model complex [Fe(SCH(3))(4)](2-) has an equilibrium conformation with D2d symmetry. The DFT energy has been decomposed into contributions for ligand-ligand and metal-ligand interactions. The latter contribution is analyzed with the angular overlap model (AOM) and constitutes the dominant stereospecific interaction in the Fe(II) complex. The sulfur lone-pair electrons exert anisotropic pi interactions on the 3d(6) shell of Fe(II), which are controlled by the torsion angles, omega(i), for the rotations of the S(i)-C(beta) bonds around the Fe-S(i) axes. In contrast, the pi interactions acting on the high-spin 3d(5) shell of Fe(III) are isotropic. As a consequence, the stereochemistry of the Fe(III) complexes is determined by the Coulomb repulsions between the ligands and has S(4) symmetry. The electrostatic repulsions between the lone pairs of the sulfurs are an essential component of the ligand-ligand interaction. The lone-pair repulsions distort the 90 degree angle SFeS' angles (delta + delta(t)) and give rise to a correlation between delta and omega, which is confirmed by crystallographic data. Both the Fe(II) and Fe(III) complexes exhibit structural bistability due to the presence of low-lying equilibrium conformations with S(4) symmetry in which the complex can be trapped by the crystalline host. PMID- 15285662 TI - Nonexponential relaxation of the metastable state of the spin-crossover system [Fe(L)2](ClO4)2.H2O [L = 2,6-Bis(pyrazol-1-ylmethyl)pyridine]. AB - The relaxation of the metastable state of the spin-crossover compound [Fe(L)(2)](ClO(4))(2).H(2)O, with L = 2,6-bis(pyrazol-1-ylmethyl)pyridine, populated by the LIESST (light induced excited spin state trapping) effect, has been investigated by magnetic measurements. The time dependence of the relaxation curve at several temperatures, starting from different initial states, is in the shape of stretched exponentials, and the thermal variation of the photostationary state under constant photoexcitation is progressive and reversible. These features are satisfactorily modeled by considering noninteracting two-level systems with a distribution of activation energies. A suggested origin for the distribution is the conformational flexibility of the nonplanar heterocyclic ligands. The effect of the intensity distribution during the LIESST process is also accounted for in a simple way. PMID- 15285663 TI - N-methylmonothiocarbamatopentamminecobalt(III): restricted C-N bond rotation and the acid-catalyzed O- to S-bonded rearrangement. AB - High-resolution (1)H and (13)C NMR studies on the linkage isomers [(NH(3))(5)CoOC(S)NHCH(3)](2+) and [(NH(3))(5)CoSC(O)NHCH(3)](2+) reveal that the O-bonded form exists as a 5:1 mixture of Z and E isomers arising from restricted rotation about the C-N bond. Similarly, restricted rotation is observed (at 20 degrees C) for the S-bonded isomer (Z/E ca. 18:1), but not for the isoelectronic carbamate ion [(NH(3))(5)CoOC(O)NHCH(3)](2+), nor for the unsubstituted carbamato complex [(NH(3))(5)CoOC(O)NH(2)](2+). An analysis of the variable-temperature NMR data for the O-bonded carbamato and urea complexes has provided quantitative data on the rotational barriers, and these ions involve much faster C-N bond rotations than the thiocarbamato complexes. The acid-catalyzed reaction of [(NH(3))(5)CoOC(S)NHCH(3)](2+) is confirmed, but there is much less parallel hydrolysis (ca. 2%) than previously reported (40 +/- 10%) for 0.1 M HClO(4). In 1 M HClO(4), [(NH(3))(5)CoSC(O)NHCH(3)](2+) and [(NH(3))(5)CoOH(2)](3+) are formed in parallel as an 83:17 mixture. The kinetic data suggest that the protonated form is at least 20-fold more reactive than the free ion and that the linkage isomerization and hydrolysis pathways are both acid-catalyzed, the latter clearly more so than the rearrangement. PMID- 15285664 TI - 4-cyanoimidazolate: a new pseudo-cyanide? AB - Because of its exobidentate nature, pK(a), and electronic properties, 4 cyanoimidazole has been examined as a ligand and as a pseudo-cyanide. The ligand reacts readily as an anion with both cobalt and nickel ions in solution to form coordination polymers. The magnetic susceptibility and thermal stability of these materials are reported. 4-Cyanoimidazolate forms a hexakis complex with nickel, to form the first observed hexakis imidazolate nickelate complex. PMID- 15285665 TI - Ytterbium and samarium bis(diphosphanylamides): syntheses and structures of lanthanide complexes having two [(Ph2P)2N]- ligands in the coordination sphere. AB - Bis(diphosphanylamide) complexes of the lanthanides have been synthesized. Two approaches to obtain these compounds are shown. Reaction of YbCl3 with a slight excess of [K(THF)n][N(PPh2)2] gives [((Ph2P)2N)2 YbCl(THF)2], which can be further reacted with K(C5Me5) to give the corresponding pentamethylcyclopentadienyl complex [((Ph2P)2N)2Yb(C5Me5)]. In a second approach to bis(diphosphanylamide) complexes of the lanthanides, Na(C(5)H(5)) was treated with SmCl3 to generate [(C5H5)SmCl2(THF)3] in situ. Further reaction with 2 equiv of [K(THF)n][N(PPh2)2] gave the desired complex [((Ph2P)2N)2Sm(C5H5)(THF)]. PMID- 15285666 TI - Resonance Raman detection of the Fe2+-C-N modes in heme-copper oxidases: a probe of the active site. AB - Resonance Raman spectroscopy has been employed to investigate the reduced cyano complexes of cytochrome aa(3) from bovine heart and Rhodobacter sphaeroides and of cytochrome bo(3) from E. coli. In the aa(3)-type oxidases, the frequency of the Fe-CN stretching mode is located at 468 cm(-1), and the bending Fe-C-N vibration, at 500 cm(-1). The fully reduced cytochrome bo(3)-CN complex gives rise to a stretching vibration at 468 cm(-1), a bending vibration at 491 cm(-1), and a stretching C-N vibration at 2037 cm(-1). The observed differences between aa(3) and bo(3) oxidases in the frequencies of the Fe-C-N group suggest a quantitative difference in the structure of the His-heme a(3)(2+)/Cu(B)(1+) and His-heme o(3)(2+)/Cu(B)(1+) binuclear pockets upon CN- binding. PMID- 15285667 TI - Diruthenium complexes [((acac)2RuIII)2(mu-OC2H5)2], [((acac)(2RuIII)2(mu L)](ClO4)2, and [((bpy)(2RuII)2(mu-L)](ClO4)4 [L = (NC5H4)2-N-C6H4-N-(NC5H4)2, acac = acetylacetonate, and bpy = 2,2'-bipyridine]. synthesis, structure, magnetic, spectral, and photophysical aspects. AB - Paramagnetic diruthenium(III) complexes (acac)(2)Ru(III)(mu OC(2)H(5))(2)Ru(III)(acac)(2) (6) and [(acac)(2)Ru(III)(mu L)Ru(III)(acac)(2)](ClO(4))(2), [7](ClO(4))(2), were obtained via the reaction of binucleating bridging ligand, N,N,N',N'-tetra(2-pyridyl)-1,4-phenylenediamine [(NC(5)H(4))(2)-N-C(6)H(4)-N-(NC(5)H(4))(2), L] with the monomeric metal precursor unit (acac)(2)Ru(II)(CH(3)CN)(2) in ethanol under aerobic conditions. However, the reaction of L with the metal fragment Ru(II)(bpy)(2)(EtOH)(2)(2+) resulted in the corresponding [(bpy)(2)Ru(II) (mu-L) Ru(II)(bpy)(2)](ClO(4))(4), [8](ClO(4))(4). Crystal structures of L and 6 show that, in each case, the asymmetric unit consists of two independent half-molecules. The Ru-Ru distances in the two crystallographically independent molecules (F and G) of 6 are found to be 2.6448(8) and 2.6515(8) A, respectively. Variable-temperature magnetic studies suggest that the ruthenium(III) centers in 6 and [7](ClO(4))(2) are very weakly antiferromagnetically coupled, having J = -0.45 and -0.63 cm(-)(1), respectively. The g value calculated for 6 by using the van Vleck equation turned out to be only 1.11, whereas for [7](ClO(4))(2), the g value is 2.4, as expected for paramagnetic Ru(III) complexes. The paramagnetic complexes 6 and [7](2+) exhibit rhombic EPR spectra at 77 K in CHCl(3) (g(1) = 2.420, g(2) = 2.192, g(3) = 1.710 for 6 and g(1) = 2.385, g(2) = 2.177, g(3) = 1.753 for [7](2+)). This indicates that 6 must have an intermolecular magnetic interaction, in fact, an antiferromagnetic interaction, along at least one of the crystal axes. This conclusion was supported by ZINDO/1-level calculations. The complexes 6, [7](2+), and [8](4+) display closely spaced Ru(III)/Ru(II) couples with 70, 110, and 80 mV separations in potentials between the successive couples, respectively, implying weak intermetallic electrochemical coupling in their mixed-valent states. The electrochemical stability of the Ru(II) state follows the order: [7](2+) < 6 < [8](4+). The bipyridine derivative [8](4+) exhibits a strong luminescence [quantum yield (phi) = 0.18] at 600 nm in EtOH/MeOH (4:1) glass (at 77 K), with an estimated excited-state lifetime of approximately 10 micros. PMID- 15285668 TI - Chiral bisphosphinite metalloligands derived from a P-chiral secondary phosphine oxide. AB - Interaction of PdCl(2)(MeCN)(2) with 2 equiv of (S(P))-(t)BuPhP(O)H (1H) followed by treatment with Et(3)N gave [Pd((1)(2)H)](2)(micro-Cl)(2) (2). Reaction of 2 with Na[S(2)CNEt(2)] or K[N(PPh(2)S)(2)] afforded Pd[(1)(2)H](S(2)CNEt(2)) (3) or Pd[(1)(2)H)[N(PPh(2)S)(2)] (4), respectively. Treatment of 3 with V(O)(acac)(2) (acac = acetylacetonate) and CuSO(4) in the presence of Et(3)N afforded bimetallic complexes V(O)[Pd(1)(2)(S(2)CNEt(2))](2) (5) or Cu[Pd(1)(2)(S(2)CNEt(2))](2) (6), respectively. X-ray crystallography established the S(P) configuration for the phosphinous acid ligands in 3 and 6, indicating that 1H binds to Pd(II) with retention of configuration at phosphorus. The geometry around Cu in 6 is approximately square planar with the average Cu-O distance of 1.915(3) A. Treatment of 2 with HBF(4) gave the BF(2)-capped compound [Pd((1)(2)BF(2))](2)(micro-Cl)(2) (7). The solid-state structure of 7 containing a PdP(2)O(2)B metallacycle has been determined. Chloride abstraction of 7 with AgBF(4) in acetone/water afforded the aqua compound [Pd((1)(2)BF(2))(H(2)O)(2)][BF(4)] (8) that reacted with [NH(4)](2)[WS(4)] to give [Pd((1)(2)BF(2))(2)](2)[micro-WS(4)] (9). The average Pd-S and W-S distances in 9 are 2.385(3) and 2.189(3) A, respectively. Treatment of [(eta(6)-p cymene)RuCl(2)](2) with 1H afforded the phosphinous acid adduct (eta(6)-p cymene)RuCl(2)(1H) (10). Reduction of [CpRuCl(2)](x)() (Cp = eta(5)-C(5)Me(5)) with Zn followed by treatment with 1H resulted in the formation of the Zn(II) phosphinate complex [(CpRu(eta(6)-C(6)H(5)))(t)BuPO(2))](2)(ZnCl(2))(2) (11) that contains a Zn(2)O(4)P(2) eight-membered ring. PMID- 15285669 TI - Cobalt half-sandwich, sandwich, and mixed sandwich complexes with soft tripodal ligands. AB - Reaction of sodium hydrotris(methimazolyl)borate (NaTm(Me)) with cobalt halides leads to the formation of paramagnetic pseudotetrahedral [Co(Tm(Me))X] (X = Cl, Br, I), of which the bromide has been crystallographically characterized. Mass spectrometry reveals the presence of higher molecular weight fragments [Co(Tm(Me))(2)](+) and [Co(2)(Tm(Me))(2)X](+) in solution. Aerial oxidation in donor solvents (e.g. MeCN) leads to formation of the [Co(Tm(Me))(2)](+) cation, which has been crystallographically characterized as the BF(4)(-), ClO(4)(-), Br( ), and I(-), salts. Attempts to prepare the mixed sandwich complex, [Co(Cp)(Tm(Me))](+), resulted in ligand decomposition to yield [Co(mtH)(3)I]I (mtH = 1-methylimidazole-2-thione), but with the more electron donating methylcyclopentadienyl (Cp(Me)) ligand, [Co(Cp(Me))(Tm(Me))]I was isolated and characterized. Electrochemical measurements reveal that the cobalt(III) Tm(Me) complexes are consistently more difficult to reduce than their Tp and Cp congeners. PMID- 15285670 TI - Coordination of 9-ethylguanine to the mixed-ligand compound alpha [Ru(azpy)(bpy)Cl2] (azpy = 2-phenylazopyridine and bpy = 2,2'-bipyridine). An unprecedented ligand positional shift, correlated to the cytotoxicity of this type of [RuL2Cl2] (with L = azpy or bpy) complex. AB - The striking difference in cytotoxic activity between the inactive cis [Ru(bpy)(2)Cl(2)] and the recently reported highly cytotoxic alpha [Ru(azpy)(2)Cl(2)] (alpha indicating the isomer in which the coordinating Cl atoms, pyridine nitrogens, and azo nitrogens are in mutual cis, trans, cis orientation) encouraged the synthesis of the mixed-ligand compound cis [Ru(azpy)(bpy)Cl(2)]. The synthesis and characterization of the only occurring isomer, i.e., alpha-[Ru(azpy)(bpy)Cl(2)], 1 (alpha denoting the isomer in which the Cl ligands are cis related to each other and the pyridine ring of azpy is trans to the pyridine ring of bpy), are described. The solid-state structure of 1 has been determined by X-ray structure analysis. The IC(50) values obtained for several human tumor cell lines have indicated that compound 1 shows mostly a low to moderate cytotoxicity. The binding of the DNA model base 9-ethylguanine (9 EtGua) to the hydrolyzed species of 1 has been studied and compared to DNA model base binding studies of cis-[Ru(bpy)(2)Cl(2)] and alpha-[Ru(azpy)(2)Cl(2)]. The completely hydrolyzed species of 1, i.e., alpha-[Ru(azpy)(bpy)(H(2)O)(2)](2+), has been reacted with 9-EtGua in water at room temperature for 24 h. This resulted in the monofunctional binding of only one 9-EtGua, coordinated via the N7 atom. The product has been isolated as alpha-[Ru(azpy)(bpy)(9 EtGua)(H(2)O)](PF(6))(2), 2, and characterized by 2D NOESY NMR spectroscopy. The NOE data show that the 9-EtGua coordinates (under these conditions) at the position trans to the azo nitrogen atom. Surprisingly, time-dependent (1)H NMR data of the 9-EtGua adduct 2 in acetone-d(6) show an unprecedented positional shift of the 9-EtGua from the position trans to the azo nitrogen to the position trans to the bpy nitrogen atom, resulting in the adduct alpha'-[Ru(azpy)(bpy)(9 EtGua)(H(2)O)](PF(6))(2) (alpha' indicating 9-EtGua is trans to the bpy nitrogen). This positional isomerization of 9-EtGua is correlated to the cytotoxicity of 1 in comparison to both the cytotoxicity and 9-EtGua coordination of cis-[Ru(bpy)(2)Cl(2)], alpha-[Ru(azpy)(2)Cl(2)], and beta-[Ru(azpy)(2)Cl(2)]. This positional isomerization process is unprecedented in model base metal chemistry and could be of considerable biological significance. PMID- 15285671 TI - Density functional study of ammonia activation by late first-row transition metal cations. AB - Density functional theory (DFT) in its B3LYP implementation is used to investigate the reaction of ammonia with the late (Co(+), Ni(+), and Cu(+)) first row transition metal cations in both high- and low-spin states. The potential energy surfaces (PES's) leading to three different exit channels are closely examined. The binding energies for the reaction products are calculated and compared with the corresponding experimental values. A comparison with our earlier works covering the reactivity of the Sc-Fe series of cations is made in order to underline similarities and differences of the reaction mechanisms as well as to establish trends along the row. PMID- 15285672 TI - Coordination networks through the dimensions: from discrete clusters to 1D, 2D, and 3D silver(I) coordination polymers with rigid aliphatic amino ligands. AB - The use of a ligand directed strategy in the assembly of discrete clusters, 1D chains, 2D layers, and 3D networks using aliphatic N-donor ligands has been investigated. The ligands are a family of amines with rigid backbones [cis,cis 1,3,5-triaminocyclohexane (cis-tach), cis,trans-1,3,5-triaminocyclohexane (trans tach), cis-1,3-diaminocyclohexane (cis-dach), and cis-3,5-diaminopiperidine (cis dapi)], and their complexation with Ag(I) salts results in a diverse set of architectures with the following compositions: [Ag3(cis-tach)2]F3.4CH(3)OH.0.5H2O (1), [Ag3(cis-tach)2]F3.6H2O (2), ([Ag(cis-dach)]ClO4)n (3), ([Ag(cis-tach)]NO3)n (4), ([Ag(trans-tach)]PF6)n(5), and ([Ag(cis-dapi)]CF3SO3)n (6). Structural analysis shows that compounds 1 and 2 are discrete M(3)L(2) cage-type clusters with varying solvent molecule content. Short Ag...Ag contacts (3.021(8) A) are observed to dimerize discrete units in compound 2. Compound 3 is a 1D zigzag chain formed by coordination to the two primary amines of cis-dach, whereas the tridentate ligands in compounds 4 and 5 (cis-tach and trans-tach, respectively) are able to form tubular architectures by virtue of their ability to "wrap" round the channel walls. An infinite 2D coordination network is demonstrated by compound 6, in which the three coplanar amino donors of cis-dapi coordinate to the trigonal planar Ag(I) ions to form a layered structure of 6(3) topology. These are compared with a previously reported 3D structure, ([Ag(trans tach)]NO3)n (7), that belongs to this family of architectures. PMID- 15285673 TI - Analysis of low oxidation state transition metal clusters by laser desorption/ionization time-of-flight mass spectrometry. AB - A variety of homonuclear and heteronuclear transition metal carbonyl clusters have been analyzed by ultraviolet laser desorption/ionization time-of-flight mass spectrometry. The spectra were recorded in negative and positive ion modes, using both linear and reflective techniques. A range of different clusters based on different nuclearities, geometries, and ligand types, which include hydrides, phosphines, nitriles, and cyclopentadienyl ligands and naked main group atoms, were studied. These experiments have allowed us to construct a detailed picture of the technique for the analysis of transition metal carbonyl clusters and their derivatives. In general, extensive reactions are observed, cluster aggregation reactions in particular, and from a comparison of the spectra obtained, some mechanistic inferences concerning the aggregation processes have been drawn. PMID- 15285674 TI - Investigation of the electronic and structural properties of potassium hexaboride, KB6, by transport, magnetic susceptibility, EPR, and NMR measurements, temperature-dependent crystal structure determination, and electronic band structure calculations. AB - The electronic and structural properties of potassium hexaboride, KB(6), were examined by transport, magnetic susceptibility, EPR, and NMR measurements, temperature-dependent crystal structure determination, and electronic band structure calculations. The valence bands of KB(6) are partially empty, but the electrical resistivity of KB(6) reveals that it is not a normal metal. The magnetic susceptibility as well as EPR and NMR measurements show the presence of localized electrons in KB(6). The EPR spectra of KB(6) have two peaks, a broad ( approximately 320 G) and a narrow (less than approximately 27 G) line width, and the temperature-dependence of the magnetic susceptibility of KB(6) exhibits a strong hysteresis below 70 K. The temperature-dependent crystal structure determination of KB(6) shows the occurrence of an unusual variation in the unit cell parameter hence supporting that the hysteresis of the magnetic susceptibility is a bulk phenomenon. The line width DeltaH(pp) of the broad EPR signal is independent of temperature and EPR frequency. This finding indicates that the line broadening results from the dipole-dipole interaction, and the spins responsible for the broad EPR peak has the average distance of approximately 1.0 nm. To explain these apparently puzzling properties, we examined a probable mechanism of electron localization in KB(6) and its implications. PMID- 15285675 TI - Synthesis, structure, and bonding of BaAuTl3 and BaAuIn3: stabilization of BaAl4 type examples of the heavier triels through gold substitution. AB - The title compounds have been synthesized by high temperature means and characterized by X-ray structural analysis, physical property measurements, and electronic structure calculations. The compounds crystallize in the three dimensional tetragonal structure of BaAl(4), I4/mmm, Z = 2 (a = 4.8107(4), 4.8604(2) A, and c = 11.980(2), 12.180(2) A for BaAuIn(3) and BaAuTl(3), respectively). Gold randomly substitutes for 50% of the In or Tl in the apical (4e) positions in the network, generating apical-apical atom distances of 2.77 and 2.70 A, respectively, values that are comparable to the single bond metallic radii sum for Au plus In, and 0.08 A less than that for Au plus Tl. Relativistic effects appear to be important for both of the latter elements. The shrinkage in distances and increase in bond strengths evidently stabilize BaAuTl(3) relative to the distorted BaTl(4) with a presumably oversized triel lattice. EHTB band calculations indicate that the two compounds are electron-deficient relative to optimal Au-Tr and Au-Au bonding and metallic, the latter in agreement with measured properties of BaAuTl(3). PMID- 15285676 TI - Excited-state dynamics of fac-[ReI(L)(CO)3(phen)]+ and fac-[ReI(L)(CO)3(5-NO2 phen)]+ (L = imidazole, 4-ethylpyridine; phen = 1,10-phenanthroline) complexes. AB - The nature and dynamics of the lowest excited states of fac [Re(I)(L)(CO)(3)(phen)](+) and fac-[Re(I)(L)(CO)(3)(5-NO(2)-phen)](+) [L = Cl(-), 4-ethyl-pyridine (4-Etpy), imidazole (imH); phen = 1,10-phenanthroline] have been investigated by picosecond visible and IR transient absorption spectroscopy in aqueous (L = imH), acetonitrile (L = 4-Etpy, imH), and MeOH (L = imH) solutions. The phen complexes have long-lived Re(I) --> phen (3)MLCT excited states, characterized by CO stretching frequencies that are upshifted relative to their ground-state values and by widely split IR bands due to the out-of-phase A'(2) and A"nu(CO) vibrations. The lowest excited states of the 5-NO(2)-phen complexes also have (3)MLCT character; the larger upward nu(CO) shifts accord with much more extensive charge transfer from the Re(I)(CO)(3) unit to 5-NO(2)-phen in these states. Transient visible absorption spectra indicate that the excited electron is delocalized over the 5-NO(2)-phen ligand, which acquires radical anionic character. Similarly, involvement of the -NO(2) group in the Franck Condon MLCT transition is manifested by the presence of an enhanced nu(NO(2)) band in the preresonance Raman spectrum of [Re(I)(4-Etpy)(CO)(3)(5-NO(2) phen)](+). The Re(I) --> 5-NO(2)-phen (3)MLCT excited states are very short lived: 7.6, 170, and 43 ps for L = Cl(-), 4-Etpy, and imH, respectively, in CH(3)CN solutions. The (3)MLCT excited state of [Re(I)(imH)(CO)(3)(5-NO(2) phen)](+) is even shorter-lived in MeOH (15 ps) and H(2)O (1.3 ps). In addition to (3)MLCT, excitation of [Re(I)(imH)(CO)(3)(5-NO(2)-phen)](+) populates a (3)LLCT (imH --> 5-NO(2)-phen) excited state. Most of the (3)LLCT population decays to the ground state (time constants of 19 (H(2)O), 50 (MeOH), and 72 ps (CH(3)CN)); in a small fraction, however, deprotonation of the imH.+ ligand occurs, producing a long-lived species, [Re(I)(im.)(CO)(3)(5-NO(2)-phen).-]+. PMID- 15285677 TI - Application of a universal force field to mixed Fe/Mo-S/Se cubane and heterocubane clusters. 1. Substitution of sulfur by selenium in the series [Fe4X4(YCH3)4]2-; X = S/Se and Y = S/Se. AB - A series of Fe-S and Fe-Se cubane clusters containing all four combinations of the general formula [Fe(4)X(4)(Y-CH(3))(4)](2)(-) (X = S/Se, Y = S/Se) is investigated with FTIR and Raman spectroscopy. The terminally selenolate coordinated clusters (Y = Se) are prepared by a new synthetic route. All four cluster compounds are structurally characterized by X-ray single-crystal structure determination. Infrared and Raman spectra of all compounds are presented and interpreted with normal coordinate analysis. The corresponding force fields are based on that developed for the Fe(4)S(4)-benzyl cluster (Czernuszewicz, R. S.; Macor, K. A.; Johnson, M. K.; Gewirth, A.; Spiro, T. G. J. Am.Chem. Soc. 1987, 109, 7178-7187). An empirical procedure is presented to convert Fe-S into Fe-Se force constants. Only minor changes in force constants are found upon S --> Se exchange, reflecting the similarity of the Fe-S and Fe-Se bonds. The drastic frequency shifts in the metal-ligand region observed upon substitution of sulfur by selenium are, therefore, primarily due to the corresponding mass changes. PMID- 15285678 TI - Application of a universal force field to mixed Fe/Mo-S/Se cubane and heterocubane clusters. 2. Substitution of iron by molybdenum in Fe4(S/Se)4 clusters with terminal halide and thiolate ligands. AB - Infrared and Raman spectra of Fe(4)(S/Se)(4) clusters with terminal halide ligands and MoFe(3)S(4) clusters with terminal thiolate and halide ligands are presented and interpreted on the basis of the force fields determined in the accompanying paper. The Raman spectra of halide coordinated Fe(4)(S/Se)(4) clusters are characterized by the fact that vibrations of the terminal ligands appear with little or vanishing intensity. Infrared and Raman spectra of MoFe(3)S(4) clusters with terminal thiolates are correlated to those of corresponding Fe(4)S(4) systems, which were investigated in part 1 of this study and interpreted with normal coordinate analysis. Band assignments are checked by employing MoFeS(4) clusters with terminal halide ligands. Spectra of these systems are in turn compared to those of their Fe(4)S(4) counterparts, i.e., Fe-S cubane clusters with chloro, bromo, and iodo ligands. A consistent interpretation of all spectra is presented. General implications of these results are discussed. PMID- 15285679 TI - Acidic aqueous decomposition of thiocyanogen. AB - The aqueous reaction of acidic Cl2 with excess SCN- rapidly generates a UV absorbing intermediate identified as an equilibrium mixture of thiocyanogen, (SCN)2, and trithiocyanate, (SCN)3(-). The decomposition of this mixture can be described as 3(SCN)2 + 4H2O --> 5HSCN + H2SO4 + HCN. Under our conditions the decomposition is sufficiently slow that its kinetics can be studied using standard stopped-flow methodology. Over the pH range 0-2 the decomposition rate law is -d[(SCN)2]/dt = (3/2)[k(disp)K(hyd)2[(SCN)2]2/([SCN-]2[H+]2 + K(SCN)3-[SCN ]3[H+]2 + K(hyd)[SCN-][H+])] with K(SCN)3(-) = 0.43 +/- 0.29 M(-1), K(hyd) = (5.66 +/- 0.77) x 10(-4) M2, and k(disp) = (6.86 +/- 0.95) x 10(4) M(-1) s(-1) at 25 degrees C and micro = 1 M. The K(SCN)3(-) and K(hyd) terms are significant enhancements relative to one of the rate laws conventionally cited. In the proposed mechanism, K(SCN)3(-) refers to the formation of (SCN)3(-) by association of SCN- with (SCN)2, K(hyd) refers to the hydrolysis of (SCN)2 to form HOSCN, and k(disp) is the rate constant for the bimolecular irreversible disproportionation of HOSCN, which leads ultimately to SO4(2-) and HCN. Ab initio calculations support the values of K(SCN)3(-) and K(hyd) reported herein. The high value for k(disp) indicates that HOSCN is a short-lived transient, while the magnitude of K(hyd) provides information on its thermodynamic stability. These results bear on the physiological role of enzymes that catalyze the oxidation of SCN- such as salivary peroxidase and myeloperoxidase. PMID- 15285680 TI - Electronic structures of five-coordinate iron(III) porphyrin complexes with highly ruffled porphyrin ring. AB - The spin states of the iron(III) complexes with a highly ruffled porphyrin ring, [Fe(TEtPrP)X] where X = F-, Cl-, Br-, I-, and ClO4(-), have been examined by 1H NMR, 13C NMR, EPR, and Mossbauer spectroscopy. While the F-, Cl-, and Br- complexes adopt a high-spin (S = 5/2) state, the I- complex exhibits an admixed intermediate-spin (S = 5/2, 3/2) state in CD2Cl2 solution. The I- complex shows, however, a quite pure high-spin state in toluene solution as well as in the solid. The results contrast those of highly saddled [Fe(OETPP)X] where the I- complex exhibits an essentially pure intermediate-spin state both in solution and in the solid. In contrast to the halide-ligated complexes, the ClO4(-) complex shows a quite pure intermediate-spin state. The 13C NMR spectra of [Fe(TEtPrP)ClO4] are characterized by the downfield and upfield shifts of the meso and pyrrole-alpha carbon signals, respectively: delta(meso) = +342 and delta(alpha-py) = -287 ppm at 298 K. The data indicate that the meso carbon atoms of [Fe(TEtPrP)ClO4] have considerable amounts of positive spin, which in turn indicate that the iron has an unpaired electron in the d(xy) orbital; the unpaired electron in the d(xy) orbital is delocalized to the meso positions due to the iron(d(xy))-porphyrin(a(2u)) interaction. Similar results have been obtained in analogous [Fe(TiPrP)X] though the intermediate-spin character of [Fe(TiPrP)X] is much larger than that of the corresponding [Fe(TEtPrP)X]. On the basis of these results, we have concluded that the highly ruffled intermediate spin complexes such as [Fe(TEtPrP)ClO4] and [Fe(TiPrP)ClO4] adopt a novel (d(xz), d(yz))3(d(xy))1(d(z)(2)1 electron configuration; the electron configuration of the intermediate-spin complexes reported previously is believed to be (d(xy))2(d(xz)), d(yz))2(d(z)(2))1. PMID- 15285681 TI - Probing the limits of the Zintl concept: structure and bonding in rare-earth and alkaline-earth zinc-antimonides Yb9Zn4+xSb9 and Ca9Zn4.5Sb9. AB - A new transition metal Zintl phase, Yb(9)Zn(4+x)Sb(9), was prepared by high temperature flux syntheses as large single crystals, or by direct fusion of the corresponding elements in polycrystalline form. Its crystal structure was determined by single-crystal X-ray diffraction. Its Ca-counterpart, hitherto known as Ca(9)Zn(4)Sb(9), and the presence of nonstoichiometry in it were also studied. Yb(9)Zn(4+x)Sb(9) was found to exist in a narrow homogeneity range, as suggested from the crystallographic data at 90(3) K (orthorhombic, space group Pbam (No. 55), Z = 2): (1) a = 21.677(2) A, b = 12.3223(10) A, c = 4.5259(4) A, R1 = 3.09%, wR2 = 7.18% for Yb(9)Zn(4.23(2))Sb(9); (2) a = 21.706(2) A, b = 12.3381(13) A, c = 4.5297(5) A, R1 = 2.98%, wR2 = 5.63% for Yb(9)Zn(4.380(12))Sb(9); and (3) a = 21.700(2) A, b = 12.3400(9) A, c = 4.5339(4) A, R1 = 2.75%, wR2 = 5.65% for Yb(9)Zn(4.384(14))Sb(9). The isostructural Ca(9)Zn(4.478(8))Sb(9) has unit cell parameters a = 21.830(2) A, b = 12.4476(9) A, and c = 4.5414(3) A (R1 = 3.33%, wR2 = 5.83%). The structure type in which these compounds crystallize is related to the Ca(9)Mn(4)Bi(9) type, and can be considered an interstitially stabilized variant. Formal electron count suggests that the Yb or Ca cations are in the +2 oxidation state. This is supported by the virtually temperature-independent magnetization for Yb(9)Zn(4.5)Sb(9). Electrical resistivity data show that Yb(9)Zn(4.5)Sb(9) and Ca(9)Zn(4.5)Sb(9) are poor metals with room-temperature resistivity of 10.2 and 19.6 mOmega.cm, respectively. PMID- 15285682 TI - Synthesis and characterization of a new family of bi-, tri-, tetra-, and pentanuclear ferric complexes. AB - Nine members of a new family of polynuclear ferric complexes have been synthesized and characterized. The reaction of Fe(O(2)CMe)(2) with polydentate Schiff base proligands (H(2)L) derived from salicylidene-2-ethanolamine, followed in some cases by reaction with carboxylic acids, has afforded new complexes of general formulas [Fe(2)(pic)(2)(L)(2)] (where pic(-) is the anion of 2-picolinic acid), [Fe(3)(O(2)CMe)(3)(L)(3)], [Fe(4)(OR)(2)(O(2)CMe)(2)(L)(4)], and [Fe(5)O(OH)(O(2)CR)(4)(L)(4)]. The tri-, tetra-, and pentanuclear complexes all possess unusual structures and novel core topologies. Mossbauer spectroscopy confirms the presence of high-spin ferric centers in the tri- and pentanuclear complexes. Variable-temperature magnetic measurements suggest spin ground states of S = 0, 1/2, 0, and 5/2 for the bi-, tri-, tetra-, and pentanuclear complexes, respectively. Fits of the magnetic susceptibility data have provided the magnitude of the exclusively antiferromagnetic exchange interactions. In addition, an easy-axis-type magnetic anisotropy has been observed for the pentanuclear complexes, with D values of approximately -0.4 cm(-)(1) determined from modeling the low-temperature magnetization data. A low-temperature micro SQUID study of one of the pentanuclear complexes reveals magnetization hysteresis at nonzero field. This is attributed to an anisotropy-induced energy barrier to magnetization reversal that is of molecular origin. Finally, an inelastic neutron scattering study of one of the trinuclear complexes has revealed that the magnetic behavior arises from two distinct species. PMID- 15285683 TI - Electronic factors affecting second-order NLO properties: case study of four different push-pull bis-dithiolene nickel complexes. AB - The paper presents a detailed experimental and theoretical study of the four mixed nickel-bisdithiolene complexes [Ni(Pr(i)(2)pipdt)(dmit)] (1b, Pr(i)(2)pipdt = 1,4-diisopropyl-piperazine-3,2-dithione; dmit = 1,3-dithiolo-2-tione-4,5 dithiolato), [Ni(R(2)pipdt)(mnt)] (2b", R = 2-ethylhexyl; mnt = maleonitriledithiolato), [Ni(Pr(i)(2)timdt)(dmit)] (3b, Pr(i)(2)timdt = 1,3 diisopropyl-imidazoline-2,4,5-trithione), and [Ni(Pr(i)(2)timdt)(mnt)] (4b), and their models. All the complexes, with common (C(2)S(2))Ni(C(2)S(2)) core and two different terminal groups, are uncharged and square-planar coordinated. Previous measurements of the first molecular hyperpolarizability indicated that some of the species are potential NLO chromophores due to the pi-delocalized character of two frontier levels (HOMO and LUMO) which is asymmetrically perturbed by the combination of one push (R(2)pipdt, R(2)timdt) with one pull ligand (dmit and mnt). The X-ray structure of complex 1b is presented and its geometry is compared with those available in the literature for the four types of complexes under study. The results of electrochemical and spectroscopic measurements (oxidation and reduction potentials, IR, dipole moment, molecular absorptivities, etc.) indicate rather different responses between the pairs of complexes 1-2 and 3-4. Hence, DFT calculations on the model compounds 1a-4a, where hydrogen atoms replace the alkyl groups of R(2)pipdt and R(2)timdt, have been carried out to correlate geometries and electronic structures. Moreover, the first molecular hyperpolarizabilities have been calculated and their components have been analyzed with the simplest two-level approximation. The derived picture highlights the different roles of the two push and pull ligands, but also the peculiar perturbation of the pi-electron density induced by the terminal CS(3) grouping of the ligand dmit. PMID- 15285684 TI - Electrochemical determination of triple helices: electrocatalytic oxidation of guanine in an intramolecular triplex. AB - Electrocatalytic oxidation of the oligonucleotide 5'- GAA GAG GTT TTT CCT CTT CTT TTT CTT CTC C (TS) by Ru(bpy)(3)(2+) was studied by cyclic voltammetry. This oligonucleotide forms either an intramolecular triplex, hairpin, or single strand, depending on the pH (Plum, G. E.; Breslauer, K. J. J. Mol. Biol. 1995, 248, 679-695). In the triplex form, the guanine doublet in TS is buried inside the folded structure, and as such is less susceptible to oxidation by electrogenerated Ru(bpy)(3)(3+). Digital simulations of the catalytic voltammograms gave a rate constant of 3.5 +/- 0.2 x 10(2) M(-1) s(-1) for oxidation of the triplex form, while oxidation of the duplex and single-stranded forms occurred with much faster rate constants of (3.5-9.1) x 10(4) M(-1) s(-1). Experiments using a truncated form of TS that lacked the third strand of the triplex were consistent with these measurements. The Ru(bpy)(3)(3+) complex was also generated by photolyzing Ru(bpy)(3)(2+) in the presence of Fe(CN)(6)(3-). This reaction produced strand scission following piperidine treatment, which was visualized using high-resolution gel electrophoresis. These experiments showed decreased reactivity for the triplex form, and also gave an unusual reversal of a common selectivity for the 5'-G of GG doublets generally seen in B-form DNA. This reversal was ascribed to strain caused by the location of the GG doublet adjacent to the hairpin loop. PMID- 15285685 TI - Organic cross-linked electropolymers as supported oxidation catalysts: poly((tetrakis(9,9'-spirobifluorenyl)porphyrin)manganese) films. AB - Anodic oxidation of free base and manganese complexes of tetraspirobifluorenylporphyrins leads to the coating of the working electrode by insoluble electroactive poly(9,9'-spirobifluorene-free and manganese porphyrin) films which electrochemical behavior and physicochemical properties are described. After removal from the electrode, the manganese-complexed polymers were evaluated as catalysts for the oxidation of alkenes by iodobenzene diacetate or iodosylbenzene. The results show that the reactions proceeded very efficiently at room temperature with good yields. The electrosynthesized polymer catalysts can be recycled by simple filtration and reused even up to the eighth cycle without loss of activity and selectivity. These results represent an important improvement over those previously described for manganese-porphyrin-catalyzed epoxidation reactions. PMID- 15285686 TI - Synthesis, crystal structure, and luminescent properties of a binuclear gallium complex with mixed ligands. AB - By introducing tridentate Schiff base ligands, a binuclear gallium complex with mixed ligands, bis(salicylidene-o-aminophenolato)-bis(8-quinolinolato)-bis gallium(III) [Ga(2)(saph)(2)q(2)], has been synthesized and structurally characterized by single-crystal X-ray crystallography. Crystal data for C(44)H(30)Ga(2)N(4)O(6) are as follows: space group, triclinic, P; a = 11.357(3) A, b = 12.945(3) A, c = 12.947(3) A, alpha = 103.461(15) degrees, beta = 100.070(7) degrees, gamma = 96.107(18) degrees, Z = 2. This complex was identified as a dimeric complex of hexacoordinated gallium with strong intermolecular and intramolecular pi-pi stacking interactions between the pyridyl/pyridyl rings. The thermal analysis showed that Ga(2)(saph)(2)q(2) can readily form a stable amorphous glass with a high glass transition temperature (T(g) = 204 degrees C), which is 27 degrees C higher than that of tris(8 hydroxyquinolinolate)aluminum (Alq(3)). In addition, a high photoluminescence efficiency (phi(PL)) of 0.318 in DMF has been demonstrated, although the central gallium atom can result in heavy-atom quenching. Organic light-emitting diodes (OLEDs) based on this complex displayed a turn-on voltage as low as 2.5 V and a high efficiency. Even at a low doping concentration of 1%, the doped Ga(2)(saph)(2)q(2) devices with 4-(dicyanomethylene)-2-tert-butyl-6-(1,1,7,7 tetramethyljulolidyl-9-enyl)-4H-pyran (DCJTB) as the dopant exhibited excellent red emission centered at 628 nm with improved durability, compared with the case of Alq(3) as the host. These distinguishing properties of Ga(2)(saph)(2)q(2) make it a good candidate as a novel electron-transporting and emitting material for OLEDs. PMID- 15285687 TI - Emitting-state displacements in ligand-centered vibrational modes in the trans [OsO2(NCS)4]2- complex determined from near-infrared luminescence spectroscopy. AB - Low-temperature luminescence spectra from three salts of the trans [OsO(2)(NCS)(4)](2-) complex exhibit highly resolved vibronic structure in both metal-ligand high-frequency O=Os=O (885 cm(-1)) and lower-frequency Os-N(CS) (255 cm(-1)) symmetric stretching modes as well as in a ligand-centered CS stretching mode (858 cm(-1)). Band maxima range from 10000 to 12000 cm(-1), and spectra contain irregular frequency intervals that correspond to transitions from more than one origin and phonon sidebands. Experimental band shapes are distinctly different for all three compounds and are calculated assuming harmonic potential energy surfaces for both the ground and emitting states. Normal-coordinate offsets along all displaced vibrational modes are determined and compared for the three compounds. The analyses reveal emitting-state displacement of high frequency ligand-centered (CS) and metal-ligand (O=Os=O) symmetric stretching modes, leading to observed high-frequency intervals (855-880 cm(-1)) that do not match any frequencies determined from ground-state Raman spectra. The values for the high-frequency normal-coordinate offsets, DeltaQ(O=Os=O) and DeltaQ(CS), were found to be on the order of 0.06 A. Offsets along the 255 cm(-1) Os-N mode varied noticeably between the three compounds and were largest for the compound with the largest value of DeltaQ(CS). PMID- 15285688 TI - Ruthenium(II) dipyridoquinoxaline-norbornene: synthesis, properties, crystal structure, and use as a ROMP monomer. AB - The synthesis, X-ray structure, and electrochemical and photophysical characterization of [Ru(phen)(2)dpq-n][PF(6)](2) (phen = phenanthroline, dpq-n = dipyridoquinoxaline-norbornene) are described. This complex contains a Ru(phen)(3)(2+) moiety in close conjugation with a norbornene unit and is the first example of a Ru(II) diimine complex capable of undergoing ring-opening metathesis polymerization. Luminescence studies of this complex showed an increase in quantum efficiency in polar solvents and in water. Preliminary ring opening metathesis polymerization studies, carried out at low monomer-to initiator ratio, showed the formation of an oligomeric mixture composed mainly of the dimer of this complex. This dimer exhibits photophysical and redox properties similar to those of the monomer, indicating that the Ru(phen)(3)(2+) moiety remains intact during the polymerization. PMID- 15285689 TI - Selective dehydrogenation of amines with respect to coordination geometry: different oxidation products of tricyano[bis(2-pyridylmethyl)amine]ferrate(II) between mer- and fac-isomers. AB - Both the fac- and mer-isomers of tricyano[bis(2-pyridylmethyl)amine = 2 DPA]ferrate(II) were isolated and their spectroscopic properties compared. The fac-isomer was converted to the corresponding iron(III) complex under acidic conditions. Oxidation of the ferrate(II) complexes with ammonium peroxodisulfate yielded the mer-isomer of the dehydrogenated ferrate(II) compound, but only the metal-oxidized iron(III) complex for the fac-isomer was produced under neutral or basic conditions. Electrochemical measurements confirmed this difference in the oxidation behavior, in which the nature of the coordination governs the ease of oxidative dehydrogenation. PMID- 15285690 TI - Mixed valence aspects of diruthenium complexes [((L)ClRu)2(mu-tppz)]n+ incorporating 2-(2-pyridyl)azoles (L) as ancillary functions and 2,3,5,6 tetrakis(2-pyridyl)pyrazine (Tppz) as bis-tridentate bridging ligand. AB - Tppz [2,3,5,6-tetrakis(2-pyridyl)pyrazine]-bridged complexes [((L)ClRu)(2)(mu tppz)]n+ with structurally similar but electronically different ancillary ligands, 2-(2-pyridyl)azoles (L), were synthesized as diruthenium(II) species. Cyclic voltammetry, EPR of paramagnetic states, and UV-vis-NIR spectroelectrochemistry show that the first two reduction processes occur at the tppz bridge and that oxidation involves mainly the metal centers. The mixed valent intermediates from one-electron oxidation exhibit moderate comproportionation constants 10(4) < K(c) < 10(5) but appear to be valence averaged according to the Hush criterion. Redox potentials, EPR, and UV-vis-NIR results show the effect of increasing donor strength of the ancillary ligands along the sequence L(1) < L(2) < L(4) << L(3), L(1) = 2-(2-pyridyl)benzoxazole, L(2) = 2-(2-pyridyl)benzthiazole, L(3) = 2-(2-pyridyl)benzimidazolate, L(4) = 1 methyl-2-(2-pyridyl)-1H-benzimidazole. Whereas the mixed valent complexes with L(1) and L(2) remain EPR silent at 4 K, the analogues with L(4) and L(3) exhibit typical ruthenium(III) EPR signals, albeit with some noticeable ligand contribution in the case of the L(3)-containing complex. Intervalence charge transfer (IVCT) bands were found in the visible spectrum for the complex with L(3) but in the near-infrared range (at ca. 1500 nm) for the other systems. PMID- 15285691 TI - New terpyridine-containing macrocycle for the assembly of dimeric Zn(II) and Cu(II) complexes coupled by bridging hydroxide anions and pi-stacking interactions. AB - The synthesis of the new terpyridine-containing macrocycle 2,5,8,11,14 pentaaza[15](6,6' ')cyclo(2,2':6',2' ')terpyridinophane (L) is reported. The ligand contains a pentaamine chain linking the 6,6' ' positions of a terpyridine unit. A potentiometric, (1)H NMR, UV-vis spectrophotometric and fluorescence emission study on the acid-base properties of L in aqueous solutions shows that the first four protonation steps occur on the polyamine chain, whereas the terpyridine nitrogens are involved in proton binding only at strongly acidic pH values. L can form both mono- and dinuclear Cu(II), Zn(II), Cd(II), and Pb(II) complexes in aqueous solution. The crystal structures of the Zn(II) and Cd(II) complexes ([ZnLH](2)(micro-OH))(ClO(4))(5) (6) and ([CdLH](2)(micro Br))(ClO(4))(5).4H(2)O (7) show that two mononuclear [MLH](3+) units are coupled by a bridging anion (OH(-) in 6 and Br(-) in 7) and pi-stacking interactions between the terpyridine moieties. A potentiometric and spectrophotometric study shows that in the case of Cu(II) and Zn(II) the dimeric assemblies are also formed in aqueous solution containing the ligand and the metals in a 1:1 molar ratio. Protonation of the complexes or the addition of a second metal ion leads to the disruption of the dimers due to the increased electrostatic repulsions between the two monomeric units. PMID- 15285692 TI - Comparative structural studies of iodide complexes of uranium(III) and lanthanide(III) with hexadentate tetrapodal neutral N-donor ligands. AB - The syntheses, the solution structures, and the crystal structures of the two new tetrapodal N-donor ligands N,N,N',N'-tetrakis(2-pyrazylmethyl)-1,3 trimethylenediamine (tpztn), 1, and N,N,N',N'-tetrakis(2-pyrazylmethyl)-trans-1,2 cyclohexanediamine (tpzcn), 2, are described. Two different geometric isomers of the cation [La(tpztn)I(2)](+) were isolated in which the ligand adopts two different conformations leading to strong differences in the metal-ligand bond distances. The crystal structure of isostructural complexes of La, U, Ce, and Nd were determined by X-ray diffraction studies for the ligands tpztn and tpzcn. In both series of complexes the two methylpyrazyl arms and the diamine spacer (trimethylene or cyclohexane) around each aliphatic nitrogen adopt the same helical configuration. The complexes crystallize as a racemic mixture of Lambda,Lambda and Delta,Delta enantiomers with distorted square antiprism geometries. In these complexes the M-N(pyrazine) distances show a decrease from La to Ce and from La to Nd which corresponds well to the decrease in ionic radius as expected in a purely ionic bonding model. Conversely the mean value of the U N(pyrazine) distances is shorter (0.043(3) A for tpztn and 0.054(11) A for tpzcn) than the mean value of the La-N(pyrazine) distances. These differences are significantly larger than the decrease expected from the variation of the ionic radii and can be interpreted in terms of a stronger M-N interaction for U(III). Previously reported extraction studies have shown that while the tripod tris[(2 pyrazyl)methyl]amine (tpza) containing three pyrazyl nitrogens extracts An(III) preferentially to Ln(III), tpztn and tpzcn display no selectivity despite the presence of four pyrazyl groups connected to a different spacer. The structural studies described here show that despite the lack of selectivity observed in the extraction conditions, the arrangement of pyrazyl nitrogens in the tetrapodal architectures of tpztn and tpzcn allows for metal-ligand interaction similar to that observed for tpza. PMID- 15285693 TI - Photochemistry of the dinitrosyl iron complex [S5Fe(NO)2]- leading to reversible formation of [S5Fe(mu-S)2FeS5]2-: spectroscopic characterization of species relevant to the nitric oxide modification and repair of [2Fe-2S] ferredoxins. AB - The reaction of [PPN][Fe(CO)(3)(NO)] and S(8) in a 1:1 molar ratio in THF proceeded to give the dinitrosyl iron complex [PPN][S(5)Fe(NO)(2)] (1) and the known [PPN](2)[S(5)Fe(mu-S)(2)FeS(5)] (2). EPR signals of g values g(z) = 2.0148, g(x) = 2.0270, and g(y) = 2.0485 at 77 K confirmed the existence of the unpaired electron in compound 1. The temperature-dependent magnetic moment of complex 1 indicates that the ground state is one unpaired electron with (S(t), S(L)) = ((1)/(2), 1) at very low temperature (S(t) is the total spin quantum number of the system; S(L) is the sum of the spin quantum numbers of two NO ligands). The O K-edge absorptions of complex 1 and [(NO)Fe(S(2)CNEt(2))(2)] at 532.1 and 532.5 eV are assigned to the transition of 1s --> pi(NO) and 1s --> pi(NO(+)), respectively. For the electronic structure of the [Fe(NO)(2)] core, DFT calculations, magnetic susceptibility measurement, EPR, and Fe K-/L-edge XAS spectroscopy of complex 1 lead to a description of [Fe(1+)(.NO)(2)](9). [2Fe-2S] cluster 2 treated with nitric oxide in THF shows that cluster 2 is transformed into the dinitrosyl iron complex 1 identified by IR, UV-vis, and X-ray diffraction analysis. The reaction may be reversed by the photolysis of the THF solution of 1 in the presence of the NO-accepting reagent [(C(4)H(8)O)Fe(S,S C(6)H(4))(2)](-) to reform 2. This result demonstrates a successful biomimetic reaction cycle of the degradation and reassembly of [2Fe-2S] cluster [S(5)Fe(mu S)(2)FeS(5)](2)(-) relevant to the repair of nitric oxide-modified [2Fe-2S] ferredoxin by cysteine desulfurase and l-cysteine in vitro. PMID- 15285694 TI - Structural consequences of the prohibition of hydrogen bonding in copper guanidine systems. AB - The synthesis and structure of copper(I) complexes supported by N-substituted bicyclic guanidines is described. The N-methyl-substituted bicyclic guanidine 1,3,4,6,7,8-hexahydro-1-methyl-2H-pyrimido[1,2-a]pyrimidine (hppMe) reacted with copper(I) chloride to afford the ion-pair [Cu(hppMe)(2)][CuCl(2)] (1), a rare example of a compound containing an unsupported Cu...Cu interaction. The analogous reaction with CuI, however, afforded the molecular mu,mu-dihalo-bridged dimer [CuI(hppMe)](2) (2). Inclusion of a trimethylsilyl substituent at nitrogen provided a sufficiently sterically encumbered environment to support a two coordinate copper center in CuCl(hppSiMe(3)) (3). Compounds 1-3 have been fully characterized, including single-crystal X-ray diffraction studies. PMID- 15285695 TI - Carnitine palmitoyltransferase-I, a new target for the treatment of heart failure: perspectives on a shift in myocardial metabolism as a therapeutic intervention. AB - Although the heart is capable of extracting energy from different types of substrates such as fatty acids and carbohydrates, fatty acids are the preferred fuel under physiological conditions. In view of the presence of diverse defects in myocardial metabolism in the failing heart, changes in metabolism of glucose and fatty acids are considered as viable targets for therapeutic modification in the treatment of heart failure. One of these changes involves the carnitine palmitoyltransferase (CPT) enzymes, which are required for the transfer of long chain fatty acids into the mitochondrial matrix for oxidation. Since CPT inhibitors have been shown to prevent the undesirable effects induced by mechanical overload, e.g. cardiac hypertrophy and heart failure, it was considered of interest to examine whether the inhibition of CPT enzymes represents a novel approach for the treatment of heart disease. A shift from fatty acid metabolism to glucose metabolism due to CPT-I inhibition has been reported to exert beneficial effects in both cardiac hypertrophy and heart failure. Since the inhibition of fatty acid oxidation is effective in controlling abnormalities in diabetes mellitus, CPT-I inhibitors may also prove useful in the treatment of diabetic cardiomyopathy. Accordingly, it is suggested that CPT-I may be a potential target for drug development for the therapy of heart disease in general and heart failure in particular. PMID- 15285696 TI - The role of paraoxonase 1 activity in cardiovascular disease: potential for therapeutic intervention. AB - The antioxidant activity of high density lipoprotein (HDL) is largely due to the paraoxonase (PON) 1 located on it. Experiments with transgenic PON1 knockout mice indicate the potential for PON1 to protect against atherogenesis. This effect of HDL in decreasing low density lipoprotein (LDL) lipid peroxidation is maintained for longer than that of antioxidant vitamins and could therefore be more protective. Several important advances in the field of PON research have occurred recently, not least the discovery that two other members of the PON gene family PON2 and PON3 - may also have important antioxidant properties. Significant advances have been made in understanding the basic biochemical function of PON1 and the discovery of possible modulators of its activity. Case-control studies of PON1 activity and coronary heart disease (CHD) have shown a clear association between CHD and low serum PON1 activity. This relationship has been further strengthened by the publication of the first prospective study showing low serum PON1 activity to be an independent predictor of new CHD events. Furthermore, decreased CHD risk has been revealed by meta-analysis to be associated with the polymorphisms of PON1, which are most active in lipid peroxide hydrolysis. Although this is likely to be an underestimate of the true contribution of PON1 to CHD (because these polymorphisms explain only a small component of the variation in PON1 activity), it is important because genetic influences are unlikely to be confounded by other factors linked with both CHD and diminished PON1 activity. PON1 is being extensively researched and it is hoped that therapeutic approaches will emerge to increase its activity. Clinical trials of these, if successful, will not only provide a novel means of preventing atherosclerosis, but also provide a more satisfactory means of testing the oxidant hypothesis of atherosclerosis than antioxidant vitamin supplementation has proved to be. PMID- 15285697 TI - Cost considerations in selecting coronary artery revascularization therapy in the elderly. AB - This article presents some of the cost factors involved in selecting coronary artery revascularization therapy in an elderly patient. With the percentage of gross national product allocated to healthcare continuing to rise in the US, resource allocation has become an issue. Percutaneous coronary intervention continues to be a viable option for many patients, with lower initial costs. However, long-term angina-free results often require further interventions or eventual surgery. Once coronary artery revascularization therapy is selected, it is worthwhile to evaluate the cost considerations inherent to various techniques. Off-pump coronary artery bypass graft surgery has seen a resurgence, with improved technology and lower hospital costs than on-pump bypass surgery. Numerous factors contributing to cost in coronary surgery have been studied and several are documented here, including the potential benefits of early extubation and the use of standardized optimal care pathways. A wide range of hospital-level cost variation has been noted, and standardization issues remain. With the advent of advanced computer-assisted robotic techniques, a push toward totally endoscopic bypass surgery has begun, with the eventual hope of reducing hospital stays to a minimum while maximizing outcomes, thus reducing intensive care unit and stepdown care times, which contribute a great deal toward overall cost. At the present time, these techniques add a significant premium to hospital charges, outweighing any potential length-of-stay benefits from a cost standpoint. As our elderly population continues to grow, use of healthcare resource dollars will continue to be heavily scrutinized. Although the clinical outcome remains the ultimate benchmark, cost containment and optimization of resources will take on a larger role in the future. PMID- 15285699 TI - Genetic polymorphisms in cytochrome P450 enzymes: effect on efficacy and tolerability of HMG-CoA reductase inhibitors. AB - Adverse drug reactions are common; they are responsible for a number of debilitating side effects and are a significant cause of death following drug therapy. It is now clear that a significant proportion of these adverse drug reactions, as well as therapeutic failures, are caused by genetic polymorphism, genetically based interindividual differences in drug absorption, disposition, metabolism, or excretion. HMG-CoA reductase inhibitors are generally very well tolerated and easy to administer with good patient acceptance. There are only two uncommon but potentially serious adverse effects related to HMG-CoA reductase inhibitor therapy: hepatotoxicity and myopathy. The occurrence of lethal rhabdomyolysis in patients treated with cerivastatin has prompted concern on the part of physicians and patients regarding the tolerability of HMG-CoA reductase inhibitors. Apart from pravastatin and rosuvastatin, HMG-CoA reductase inhibitors are metabolized by the phase I cytochrome P450 (CYP) superfamily of drug metabolizing enzymes. The best-characterized pharmacogenetic polymorphisms are those within this enzyme family. One of these enzymes, CYP2D6, plays an important role in the metabolism of simvastatin. It has been shown that the cholesterol lowering effect as well as the efficacy and tolerability of simvastatin is influenced by CYP2D6 genetic polymorphism. Because the different HMG-CoA reductase inhibitors differ, with respect to the degree of metabolism by the different CYP enzymes, genotyping may help to select the appropriate HMG-CoA reductase inhibitor and the optimal dosage during the start of the treatment and will allow for more efficient individual therapy. A detailed knowledge of the genetic basis of individual drug response is potentially of major clinical and economic importance. PMID- 15285698 TI - Dyslipidemia in visceral obesity: mechanisms, implications, and therapy. AB - Visceral obesity is frequently associated with high plasma triglycerides and low plasma high density lipoprotein-cholesterol (HDL-C), and with high plasma concentrations of apolipoprotein B (apoB)-containing lipoproteins. Atherogenic dyslipidemia in these patients may be caused by a combination of overproduction of very low density lipoprotein (VLDL) apoB-100, decreased catabolism of apoB containing particles, and increased catabolism of HDL-apoA-I particles. These abnormalities may be consequent on a global metabolic effect of insulin resistance. Weight reduction, increased physical activity, and moderate alcohol intake are first-line therapies to improve lipid abnormalities in visceral obesity. These lifestyle changes can effectively reduce plasma triglycerides and low density lipoprotein-cholesterol (LDL-C), and raise HDL-C. Kinetic studies show that in visceral obesity, weight loss reduces VLDL-apoB secretion and reciprocally upregulates LDL-apoB catabolism, probably owing to reduced visceral fat mass, enhanced insulin sensitivity and decreased hepatic lipogenesis. Adjunctive pharmacologic treatments, such as HMG-CoA reductase inhibitors, fibric acid derivatives, niacin (nicotinic acid), or fish oils, may often be required to further correct the dyslipidemia. Therapeutic improvements in lipid and lipoprotein profiles in visceral obesity can be achieved by several mechanisms of action, including decreased secretion and increased catabolism of apoB, as well as increased secretion and decreased catabolism of apoA-I. Clinical trials have provided evidence supporting the use of HMG-CoA reductase inhibitors and fibric acid derivatives to treat dyslipidemia in patients with visceral obesity, insulin resistance and type 2 diabetes mellitus. Since drug monotherapy may not adequately optimize dyslipoproteinemia, dual pharmacotherapy may be required, such as HMG-CoA reductase inhibitor/fibric acid derivative, HMG-CoA reductase inhibitor/niacin and HMG-CoA reductase inhibitor/fish oils combinations. Newer therapies, such as cholesterol absorption inhibitors, cholesteryl ester transfer protein antagonists and insulin sensitizers, could also be employed alone or in combination with other agents to optimize treatment. The basis for a multiple approach to correcting dyslipoproteinemia in visceral obesity and the metabolic syndrome relies on understanding the mechanisms of action of the individual therapeutic components. PMID- 15285700 TI - The paclitaxel (TAXUS)-eluting stent: a review of its use in the management of de novo coronary artery lesions. AB - The TAXUSExpress stent contains paclitaxel 1 microg/mm(2). On deployment, paclitaxel is slowly released into the intimal tissue of the coronary artery to prevent cell proliferation and neointimal hyperplasia. When deployed in patients with previously untreated coronary artery lesions, the paclitaxel-eluting stent (PES) effectively reduces the need for revascularization without increasing the risk of in-stent thrombosis. While long-term outcomes data and comparative efficacy and cost-benefit trials versus other drug-eluting stents are required, the PES appears to be an attractive alternative for the management of de novo coronary artery lesions. PHARMACOLOGIC PROPERTIES: The PES comprises a stainless steel stent coated with a non-erodible biocompatible polyolefin matrix containing paclitaxel 1 microg/mm(2). Paclitaxel dose dependently inhibits vascular smooth muscle cell proliferation at therapeutic concentrations as a result of binding to and stabilizing cellular microtubules. This prevents the cascade of events associated with obstructive in-stent neointimal hyperplasia. Paclitaxel was released in a controlled manner from the stent coating in in vitro studies. The higher release rate in the first 2 days after implantation (to reduce response to implantation injury) slows over the next 8-10 days. The drug is rapidly taken up by intimal cells with minimal dispersion in the plasma; it was not detected systemically after stent deployment in clinical trials. Paclitaxel is extensively bound to proteins (88-98%), and is principally metabolized in the liver, undergoing biliary clearance after systemic administration. THERAPEUTIC EFFICACY: The efficacy of the PES was compared with that of a bare-metal stent (BMS) in a number of randomized, double-blind, multicenter trials in patients with de novo coronary artery lesions. The TAXUS I and II trials used the NIR stent, while the pivotal TAXUS IV trial used the Express stent. The primary endpoints of the well designed TAXUS II and IV trials indicated superiority for the PES over the BMS. Twice as many patients receiving the BMS required target vessel revascularization at 9 months postprocedure and, 6 months following the procedure, the in-stent neointimal volume in the PES system was only one-third of that in the BMS. The incidence of cumulative major adverse cardiac events (MACE; cardiac death, myocardial infarction [MI], or target vessel revascularization [TVR]) was also significantly lower in PES than BMS recipients at 9 and 12 months postprocedure. The incidence of cardiac death and MI was low and similar between treatment groups; however, TVR was significantly reduced by the PES versus the BMS. Other secondary endpoints, such as target lesion revascularization, luminal diameter stenosis, minimal luminal diameter, and serial intravascular ultrasound measurements from one or both trials supported these results. Preliminary analysis of the subgroup of patients with diabetes mellitus in the TAXUS IV trial suggested that the PES was also effective in diabetic patients who receive oral medications. The group receiving insulin was too small to draw meaningful conclusions. TOLERABILITY: Because of the small paclitaxel dosages and the mainly local uptake, systemic adverse events associated with the PES are considered unlikely. The incidences of cardiac death and MI were very low and similar in both groups. Local events such as in-stent aneurysms, incomplete stent apposition, or in-stent thrombosis occurred at a similar rate in PES and BMS recipients. There has been no evidence of late thrombosis in PES recipients followed for 2 years. The rate of late luminal loss in the 5mm of vessel proximal and distal to the stent edges was significantly lower in PES than BMS systems. PHARMACOECONOMIC CONSIDERATIONS: Initial deployment costs associated with the PES are likely to be offset by savings in repeat procedures, according to a cost effectiveness analysis in the UK. PMID- 15285701 TI - AIDS vaccines. PMID- 15285703 TI - The roles of nonhuman primates in the preclinical evaluation of candidate AIDS vaccines. AB - Preclinical studies in nonhuman primates (NHP) play key roles in AIDS vaccine development efforts. In addition to their traditional utilization to gauge vaccine safety and immunogenicity, NHP models are currently employed to an unprecedented extent and in unprecedented ways in contemporary basic and applied vaccine development efforts. Current studies employ NHP models to probe fundamental mechanisms of primate immune system regulation, to investigate pathogenic mechanisms of AIDS, and to optimize immunization strategies involving novel vaccine vectors. The use of experimental challenges of immunized NHPs with either simian immunodeficiency virus or chimeric simian/human immunodeficiency virus to generate preclinical vaccine efficacy data has emerged as an important criterion for facilitating entry of a given vaccine candidate into early phase clinical evaluation in humans. However, for studies of the biology of AIDS virus transmission, AIDS virus disease pathogenesis and AIDS virus vaccine efficacy that are predicated on experimental viral challenge to be most valuable, additional efforts need to be devoted to generating challenge models that more closely recapitulate HIV-1 infection in humans. Towards this end, improved communication between clinical and preclinical investigators, to promote a bidirectional flow of information focusing on individual research needs and shared goals should enable the NHP models to most effectively expedite progress toward the development of a safe and effective AIDS vaccine. PMID- 15285704 TI - Neutralizing antibody responses to HIV: role in protective immunity and challenges for vaccine design. AB - AIDS continues to be a major health problem throughout the world with a high degree of mortality and morbidity. Therefore, there is an urgent need for an effective anti-HIV vaccine. Although the correlates of protective immunity against infection by HIV remain unidentified, recent studies have demonstrated that both humoral and cellular responses are required for controlling viral replication. Vaccine efforts should therefore aim at developing broad and potent humoral as well as cellular responses. Anti-HIV T-cell responses can be generated both in animals and humans by several vaccine modalities. In contrast, broadly neutralizing antibody responses against HIV have not been elicited by any strategy tested in the clinic thus far. The presence of such responses has the potential to prevent the establishment of infection. If not, the presence of neutralizing antibodies may significantly reduce the number of cells that become infected, therefore reducing the inoculum, which may delay viral spread and allow for a better control of viral replication in the infected host. Finally, cytotoxic T-lymphocytes may facilitate the clearance of virally infected cells. One of the biggest challenges in HIV vaccine development is to design a HIV envelope immunogen that can induce protective neutralizing antibodies effective against the diverse HIV-1 strains that characterize the global pandemic. The focus of this article is to review the importance of antibodies and the strategies that are currently being used for inducing such antibodies. PMID- 15285705 TI - Requirement of diverse T-helper responses elicited by HIV vaccines: induction of highly targeted humoral and CTL responses. AB - With the continued spread of the HIV/AIDS epidemic at alarming proportions there is a sense of urgency for an effective prophylactic HIV vaccine. However, in addition to the social, geopolitical and public health problems, the scientific challenges often seem insurmountable. Empirical approaches to develop an HIV/AIDS vaccine have been unsuccessful and this, coupled with the recent failure of the first Phase III clinical trials, calls for a strong rational approach based on a deeper scientific understanding of the correlates of immunity observed in both preclinical and clinical settings. While the field has been polarized between those who have been proponents of vaccines that induce strong cytotoxic T-cell responses, and those who advocate inducing neutralizing antibody responses, we have maintained middle ground. Based on our early preclinical observations in rigorous nonhuman primate vaccine efficacy studies, we have focused on vaccine strategies that induce potent T-helper immune responses capable of driving both cytotoxic, as well as broad highly effective neutralizing antibodies. The critical issue remains in the selection of the specific vaccine antigens. To date, our approach has been to utilize multiple structural as well as regulatory HIV antigens containing highly conserved epitopes. The current challenge faced is to design novel antigens based on mimicking envelope structures capable of inducing broad neutralizing antibodies. Our aim is to combine these with immunization strategies capable of eliciting potent cellular as well as humoral immune responses with the ultimate goal of providing mucosal barriers to HIV entry. PMID- 15285706 TI - Mucosal AIDS vaccines: current status and future directions. AB - Natural transmission of HIV occurs through mucosal surfaces. New information in immunology, virology and vaccinology has emerged regarding strategies for development of new mucosal vaccines against HIV. The intestinal mucosa represents a major site of HIV replication and amplification, and the initial site of CD4+ T cell depletion. Local mucosal CD8+ cytotoxic T-lymphocytes (CTLs) and mucosal antibody can control AIDS virus replication within local tissues prior to systemic dissemination and can be more effective than a systemic immune response. Mucosal HIV vaccine delivery should be considered among the most effective immunization routes in the induction of mucosal antibody and CD8+ CTLs and protection against mucosal infection. New mucosal vaccine strategies, such as prime-boost, using a new generation of mucosal adjuvants, a synergistic combination of cytokines, chemokines, costimulatory molecules, CpG oligodeoxynucleotides, and targeting lymph nodes which drain mucosal sites, show promise to improve the efficacy of mucosal vaccines. PMID- 15285707 TI - Poxvirus-based vaccine candidates for HIV: two decades of experience with special emphasis on canarypox vectors. AB - Poxvirus vectors have emerged as important vectors for licensed veterinary vaccines and candidate vaccines for humans. Vaccinia, highly-attenuated vaccinia strains and avipoxviruses have been assessed extensively in preclinical models, as well as in humans, to determine their immunogenicity and protective efficacy against HIV. The attenuated vaccinia strains and avipoxviruses have been shown to be safe and able to carry HIV genes and express their proteins to induce both antibodies and cellular immune responses. Preclinical studies show protection against HIV challenge. When using a live attenuated vector system, one must be cognizant of the potential for immune dampening because of vector-specific immunity. In this regard, avipoxviruses, such as canarypox, appear free of the inhibitory effects of vector immunity and repeated use. Unlike vaccinia-based vectors derived from classical vaccine strains, NYVAC and modified vaccinia Ankara may be less susceptible to this effect. In the coming 5 to 10 years, we will certainly know whether this class of vaccine candidates, either alone or in a prime-boost format with other vectors or proteins, will contribute to HIV disease management either from a preventive or therapeutic perspective. Additional Phase I and II studies, as well as human efficacy trials will provide new information. Furthermore, it is hoped that this body of data will contribute to a better understanding of the relevance of specific immunogenicity end points to protection and the predictive value of available animal models in HIV vaccine development. PMID- 15285708 TI - MVA as a vector for vaccines against HIV-1. AB - A vaccine against HIV Type 1 (HIV-1) is urgently needed. Modified vaccinia virus Ankara is an attenuated smallpox vaccine which can be adapted to express HIV-1 antigens. In this review, we discuss the features which make modified vaccinia virus Ankara an attractive vector for genetic vaccines and have put it, together with several other recombinant viral vectors, at the forefront of HIV-1 vaccine development. Many candidate vaccines including those vectored by modified vaccinia virus Ankara are now entering human trials, the results of which will become available in the coming years. PMID- 15285709 TI - ALVAC-HIV vaccines: clinical trial experience focusing on progress in vaccine development. AB - A decade of clinical trial experience with the preventive use of ALVAC-HIV (Aventis Pasteur) has revealed important information on the safety and immunogenicity of HIV vaccines in general. The ability to induce mucosal immune responses and the feasibility of assessing those responses with a systemically delivered HIV vaccine was recently shown with ALVAC-HIV. A critical advance for the HIV vaccine field was the reported direct relationship between cellular immune responses to ALVAC-HIV vaccine in human leukocyte antigen (HLA) B27 and HLA B57 carriers, alleles linked with a favorable HIV prognosis in the setting of natural infection. Other advances include the use of ALVAC-HIV as a heterologous viral boost to enhance cellular responses to an adenovirus-HIV vaccine. All of these observations have important implications for the future of HIV vaccines and need to be considered in the design of the next generation of products. PMID- 15285710 TI - A call for replicating vector prime-protein boost strategies in HIV vaccine design. AB - A key challenge to HIV vaccine development is the integration of HIV proviral DNA into the host genome upon infection. Therefore, an optimal vaccine should block infection within hours of viral exposure, providing 'sterilizing immunity' at mucosal sites and in blood via potent, broadly reactive antibody to the HIV envelope glycoprotein. This is difficult due to the envelope's conformational complexity and sequence diversity. Antibodies that do not completely prevent infection nevertheless could reduce the viral infectious burden, allowing strong cellular immunity to control viremia, delay disease progression and prevent viral transmission, while also providing help for T- and B-cell responses. Rapidly responsive, potent, persistent immunity might best be achieved using prime-boost strategies incorporating a replicating vector and an optimally designed envelope subunit. PMID- 15285711 TI - Progress towards the use of Listeria monocytogenes as a live bacterial vaccine vector for the delivery of HIV antigens. AB - Listeria monocytogenes is a facultative intracellular bacterium that enters the cell by phagocytosis after which it colonizes the cytosol of the host cell. It is thus a potent vaccine vector for the presentation of passenger antigens to the major histocompatability complex class II and class I pathways of antigen processing and presentation. This article shall review the progress made in developing this unusual bacterium as a vaccine vector. In mouse models, recombinant Listeria carrying a number of different antigens have been shown to provide protective immunity against infectious organisms and therapeutic immunity directed towards tumor-associated antigens. Listeria has been engineered to express a number of HIV/SIV antigens. Measurements of immune responses using these recombinant strains in the mouse, after oral and parenteral immunization, and in the rhesus macaque after oral immunization indicate that strong cell mediated immunity can be induced against these antigens. This review also discusses safety issues associated with live bacterial vaccine vectors and problems to be overcome in developing Listeria as a HIV vaccine for human use. PMID- 15285712 TI - Approaches for the design and evaluation of HIV-1 DNA vaccines. AB - Although it is not clear what arm of the immune response correlates with protection from HIV-1 infection or disease, a robust broad cellular and humoral immune response will likely be needed to control this infection. Accordingly, it is crucial to characterize which HIV-1 gene products are potential targets to elicit these responses. DNA vaccination has been shown to be effective for induction of both humoral and cellular immune responses in animal models. Most DNA vaccine strategies studied to date have been based on targeting structural HIV-1 proteins, but others have focused on the regulatory/accessory HIV-1 proteins as an approach to induce immune responses able to recognize early infected cells. It has also become clear that HIV-DNA vaccine efficacy in humans requires improvement. Combinations of HIV-1 genes, improvement of the DNA vector itself, or addition of genetic adjuvants (cytokines or costimulatory molecules) as part of the DNA vaccine itself, have been evaluated by several groups as approaches for enhancing DNA vaccine-induced immune responses. Encouraging results have been obtained in primate models, supporting that these strategies should be further evaluated in humans, for either prophylaxis or immune therapy of HIV-1. PMID- 15285713 TI - Immunogen sequence: the fourth tier of AIDS vaccine design. AB - While worldwide efforts to develop an effective HIV-1 vaccine are underway, the virus continues to spread, particularly in developing countries where the delivery of antiviral therapies presents formidable challenges. Vaccine research has largely focused on three general aspects: vectors, adjuvants, and immunization schedules. Our group favor the use of computational methods to design potential immunogens that capture the genetic and biological features of circulating viruses. These methods allow researchers to predict, in silico, the presence of potential glycosylation sites, humoral immune responses, and epitope coverage. This review shall compare three computational approaches for immunogen design: the consensus sequence, which has at each site the modal nucleotide or amino acid residue across a sequence alignment; the most recent common ancestor, the sequence estimated at the basal node of the clades seen in the HIV-1 phylogeny; and the center of tree method, which minimizes the evolutionary distance to all sequences in the data set. PMID- 15285714 TI - Centralized immunogens as a vaccine strategy to overcome HIV-1 diversity. AB - Genetic variation of HIV-1 represents a major obstacle for AIDS vaccine development. With the amino acid sequence divergence as high as 30% in envelopes between different subtypes among HIV-1 group M viruses, it is unlikely that cross subtype protection will occur equally well among all subtypes. Computer programs have been used to generate 'centralized' HIV gene sequences: consensus, ancestor or center of the tree. These sequences can decrease the genetic distances between the 'centralized' and wild-type gene immunogens to half of those between any wild type immuongens to each other. Recent studies demonstrated that an artificial group M consensus env gene is equidistant from any subtype and recombinants. It is biologically functional and preserves antigenicity similar to contemporary Env proteins. Most importantly, the group M consensus Env immunogen can elicit both T and B-cell responses to wild-type HIV-1 isolates. PMID- 15285715 TI - Evaluating therapeutic vaccines in patients infected with HIV. AB - Long-term survival of HIV infection can mean decades of treatment for a patient, with major side effects and costs that limit their efficacy and accessibility. Although antiretroviral therapy remains the only standard of care, alternative therapeutic strategies must be found to ensure efficient and safe clinical management of the disease in the long term. Therapeutic immunization against HIV might be a significant approach to enhancing immune control of the virus and limiting disease progression and thus the requirement for medication. Several anti-HIV vaccines are currently being evaluated in attempts to prolong periods of treatment interruption in HIV patients. The design and end-points of clinical trials, and the clinical settings in which these new strategies should be evaluated and will be of benefit, have yet to be defined and are the focus of this review. PMID- 15285716 TI - Therapeutic vaccination against HIV. AB - The challenge for an immunotherapeutic vaccine is to increase antiviral responses in an increasingly immunocompromised host and to provide immunity to epitopes that have been neglected by the infected host. Therapeutic vaccination with structural and regulatory genes and proteins of HIV are reviewed. The most promising clinical results consist of short-term improvement in survival without antiretroviral therapy. Together with antiviral therapy, it is reported that immunization has provided a prolonged time to virological failure. It is clear, however, that additional help will be needed from adjuvants and/or modulators that activate natural killer and T-cells, or other immune molecules. Vaccine therapy should start early, while adequate reservoirs of appropriate T-helper and memory cells are available and still inducible. PMID- 15285717 TI - APC-targeted immunization for the treatment of HIV-1. AB - Therapeutic immunization may be thought of as an adjunct to highly active antiretroviral therapy to prime the immune system and possibly correct for immunological defects. Most therapeutic vaccine strategies currently under investigation aim to increase HIV-specific cellular responses. This may be most successfully accomplished by utilizing professional antigen-presenting cells. Autologous dendritic cells may be isolated, cultured, loaded with antigen and re injected into the subject (ex vivo) or antigen may be directly delivered in situ to Langerhans cells or dermal dendritic cells, which are located respectively at the epidermal and dermal layer of the skin. Once Langerhans cells or dermal dendritic cells have incorporated the antigen, they are expected to mature and migrate to the lymph node to present antigen and stimulate naive T-cells. Exciting results have been obtained in nonhuman primates with both ex vivo and topical antigen-presenting cell-based therapeutic immunization. PMID- 15285718 TI - Alternative translation initiation generates a novel isoform of insulin-degrading enzyme targeted to mitochondria. AB - IDE (insulin-degrading enzyme) is a widely expressed zinc-metallopeptidase that has been shown to regulate both cerebral amyloid beta-peptide and plasma insulin levels in vivo. Genetic linkage and allelic association have been reported between the IDE gene locus and both late-onset Alzheimer's disease and Type II diabetes mellitus, suggesting that altered IDE function may contribute to some cases of these highly prevalent disorders. Despite the potentially great importance of this peptidase to health and disease, many fundamental aspects of IDE biology remain unresolved. Here we identify a previously undescribed mitochondrial isoform of IDE generated by translation at an in-frame initiation codon 123 nucleotides upstream of the canonical translation start site, which results in the addition of a 41-amino-acid N-terminal mitochondrial targeting sequence. Fusion of this sequence to the N-terminus of green fluorescent protein directed this normally cytosolic protein to mitochondria, and full-length IDE constructs containing this sequence were also directed to mitochondria, as revealed by immuno-electron microscopy. Endogenous IDE protein was detected in purified mitochondria, where it was protected from digestion by trypsin and migrated at a size consistent with the predicted removal of the N-terminal targeting sequence upon transport into the mitochondrion. Functionally, we provide evidence that IDE can degrade cleaved mitochondrial targeting sequences. Our results identify new mechanisms regulating the subcellular localization of IDE and suggest previously unrecognized roles for IDE within mitochondria. PMID- 15285719 TI - NALP1 is a transcriptional target for cAMP-response-element-binding protein (CREB) in myeloid leukaemia cells. AB - NALP1 (also called DEFCAP, NAC, CARD7) has been shown to play a central role in the activation of inflammatory caspases and processing of pro-IL1b (pro interleukin-1b). Previous studies showed that NALP1 is highly expressed in peripheral blood mononuclear cells. In the present study, we report that expression of NALP1 is absent from CD34+ haematopoietic blast cells, and its levels are upregulated upon differentiation of CD34+ cells into granulocytes and to a lesser extent into monocytes. In peripheral blood cells, the highest levels of NALP1 were observed in CD3+ (T-lymphocytes), CD15+ (granulocytes) and CD14+ (monocytes) cell populations. Notably, the expression of NALP1 was significantly increased in the bone marrow blast cell population of some patients with acute leukaemia, but not among tissue samples from thyroid and renal cancer. A search for consensus sites within the NALP1 promoter revealed a sequence for CREB (cAMP response-element-binding protein) that was required for transcriptional activity. Moreover, treatment of TF1 myeloid leukaemia cells with protein kinase C and protein kinase A activators induced CREB phosphorylation and upregulated the mRNA and protein levels of NALP1. Conversely, ectopic expression of a dominant negative form of CREB in TF1 cells blocked the transcriptional activity of the NALP1 promoter and significantly reduced the expression of NALP1. Thus NALP1 is transcriptionally regulated by CREB in myeloid cells, a mechanism that may contribute to modulate the response of these cells to pro-inflammatory stimuli. PMID- 15285720 TI - Guanidinium chloride denaturation of the dimeric Bacillus licheniformis BlaI repressor highlights an independent domain unfolding pathway. AB - The Bacillus licheniformis 749/I BlaI repressor is a prokaryotic regulator that, in the absence of a beta-lactam antibiotic, prevents the transcription of the blaP gene, which encodes the BlaP beta-lactamase. The BlaI repressor is composed of two structural domains. The 82-residue NTD (N-terminal domain) is a DNA binding domain, and the CTD (C-terminal domain) containing the next 46 residues is a dimerization domain. Recent studies have shown the existence of the monomeric, dimeric and tetrameric forms of BlaI in solution. In the present study, we analyse the equilibrium unfolding of BlaI in the presence of GdmCl (guanidinium chloride) using different techniques: intrinsic and ANS (8 anilinonaphthalene-l-sulphonic acid) fluorescence, far- and near-UV CD spectroscopy, cross-linking, analytical ultracentrifugation, size exclusion chromatography and NMR spectroscopy. In addition, the intact NTD and CTD were purified after proteolysis of BlaI by papain, and their unfolding by GdmCl was also studied. GdmCl-induced equilibrium unfolding was shown to be fully reversible for BlaI and for the two isolated fragments. The results demonstrate that the NTD and CTD of BlaI fold/unfold independently in a four-step process, with no significant co-operative interactions between them. During the first step, the unfolding of the BlaI CTD occurs, followed in the second step by the formation of an 'ANS-bound' intermediate state. Cross-linking and analytical ultracentrifugation experiments suggest that the dissociation of the dimer into two partially unfolded monomers takes place in the third step. Finally, the unfolding of the BlaI NTD occurs at a GdmCl concentration of approx. 4 M. In summary, it is shown that the BlaI CTD is structured, more flexible and less stable than the NTD upon GdmCl denaturation. These results contribute to the characterization of the BlaI dimerization domain (i.e. CTD) involved in the induction process. PMID- 15285722 TI - The European Blood Directive: a new era of blood regulation has begun. PMID- 15285721 TI - Proteomic characterization of two snake venoms: Naja naja atra and Agkistrodon halys. AB - Snake venom is a complex mixture of proteins and peptides, and a number of studies have described the biological properties of several venomous proteins. Nevertheless, a complete proteomic profile of venom from any of the many species of snake is not available. Proteomics now makes it possible to globally identify proteins from a complex mixture. To assess the venom proteomic profiles from Naja naja atra and Agkistrodon halys, snakes common to southern China, we used a combination strategy, which included the following four different approaches: (i) shotgun digestion plus HPLC with ion-trap tandem MS, (ii) one-dimensional SDS/PAGE plus HPLC with tandem MS, (iii) gel filtration plus HPLC with tandem MS and (iv) gel filtration and 2DE (two-dimensional gel electrophoresis) plus MALDI TOF (matrix-assisted laser desorption ionization-time-of-flight) MS. In the present paper, we report the novel identification of 124 and 74 proteins and peptides in cobra and viper venom respectively. Functional analysis based upon toxin categories reveals that, as expected, cobra venom has a high abundance of cardio- and neurotoxins, whereas viper venom contains a significant amount of haemotoxins and metalloproteinases. Although approx. 80% of gel spots from 2DE displayed high-quality MALDI-TOF-MS spectra, only 50% of these spots were confirmed to be venom proteins, which is more than likely to be a result of incomplete protein databases. Interestingly, these data suggest that post translational modification may be a significant characteristic of venomous proteins. PMID- 15285723 TI - Using patient-identifiable data for epidemiological research. AB - The use of patient-identifiable data in epidemiological research is subject to increasingly complex regulation. This article reports the experience of a research team in setting up the Epidemiology and Survival of Transfusion Recipients (EASTR) study in which patient-identifiable information was needed in order to link data from two sources for analysis and obtain long-term survival patterns of transfusion recipients. The process of establishing the study involved obtaining separate ethical, research and development and data protection approval, including application to the newly formed Patient Information Advisory Group, set up under Section 60 of the Health and Social Care Act, 2001. We describe the high cost in administrative procedures and time now necessary to gain statutory approval before such a study can begin, which has been the result of recent legislation. Issues arising from our experience are discussed. PMID- 15285724 TI - A simple automatized audit system for following and managing practices of platelet and plasma transfusions in a neonatal intensive care unit. AB - During neonatal intensive care, blood components are often used in clinical situations where both their efficacy and safety lack solid justification. A practical system to continuously analyse actual transfusion practices is a prerequisite for improvements of quality in transfusion therapy. We hypothesized that such a system would reveal inappropriate variations in clinical decision making and offer a means for staff education and quality improvement and assurance. The study consisted of three 120-152-day periods (P I, P II and P III) between January 2000 and October 2001 and involved 543 new patient admissions (141 patients with birth weight < 1501 g) and 6227 days of patient care at a single tertiary level NICU. P I was a control with no intervention, P II was after technically introducing the computer system and, the last period, P III was after presenting and discussing the results of P I and P II at a staff meeting. Upon an order of platelet or fresh frozen plasma (FFP) unit from the blood bank, a computer-based audit system compared the last platelet count or prothrombin time [expressed as percentage of normal clotting activity, prothrombin time (PT %)] to predefined criteria. In the case of exceeding the preset thresholds, the system required additional information and recorded the pretransfusion laboratory values for later analysis. Thirty-two per cent of platelet transfusions were given with pretransfusion platelet count >49 x 10(9) L(-1), and 60% of these transfusions (19% of all platelet transfusions) could not be clinically justified in retrospective chart review. There was no significant change in this practice from P II to P III. FFP transfusions were given with significantly different pretransfusion PT-% values during P II and P III. The proportions of FFP transfusions with pretransfusion PT-% > 49% were 7.8% and 0.9% during P II and P III, respectively (P < 0.0001). In chart review, none of the FFP transfusions with pretransfusion PT-% > 49% could be justified by clinical grounds. Inappropriate transfusions of both platelets and plasma remain a significant challenge for quality assurance of neonatal intensive care. Automated recording of pretransfusion platelet count and prothrombin time reliably identified the poorly justified transfusions and thus offered a practical resource-saving tool for quality assurance of transfusion in the NICU. A significant shift towards more appropriate use of plasma was demonstrated after implementation of the audit system. PMID- 15285725 TI - Therapeutic efficacy of pooled buffy-coat platelet components prepared and stored with a platelet additive solution. AB - Despite the introduction of platelet additive solutions for the preparation of pooled platelet components, only a few studies of limited scope have evaluated the clinical efficacy of platelets stored in these solutions. The current report presents an analysis of data to evaluate the response to the transfusion of pooled buffy-coat components suspended in storage solution with reduced (35%) plasma content in comparison with 100% plasma products. During the euroSPRITE clinical trial of platelet components treated with a pathogen inactivation process, control treatment group platelet components were prepared in 100% allogeneic donor plasma (plasma control) or in platelet additive solution (T-Sol) mixed with plasma (T-Sol control). Control group thrombocytopenic patients received either plasma control or T-Sol control platelet components. One-hour and 24-h platelet count increments (CIs) and corrected count increments (CCIs) were analysed for these two types of preparation. In addition, haemostatic assessments were conducted for each transfusion. One-hour and 24-h mean platelet CIs and post transfusion haemostatic scores were not significantly different for patients receiving platelet components suspended in 100% plasma and T-Sol plasma mixtures. Pooled buffy-coat platelet components prepared in reduced plasma content mixtures provided therapeutic platelet CIs with effective haemostasis. PMID- 15285726 TI - Plasma quality after whole-blood filtration depends on storage temperature and filter type. AB - The study evaluated the quality of plasma obtained after whole-blood filtration with four different polyester filters and one polyurethane filter. The activities of coagulation factors and proteinase inhibitors were not or only negligibly affected by filtration, in all experiments. Filtration did not increase markers of clotting and fibrinolysis. Only a strong neutrophil and complement activation was observed, which depended on the type of filter and whole-blood storage conditions. However, as neutrophil elastase-specific degradation products did not increase and the complement-derived anaphylatoxin C3a was found in its inactivated form, C3a-desArg, these filtration-dependent changes apparently have little impact on the therapeutic quality of whole-blood-filtered fresh frozen plasma for transfusion. PMID- 15285727 TI - Soluble urokinase plasminogen activator receptor in blood transfusion components. AB - Post-transfusion infectious complications associated with allogeneic blood components may depend on storage time and may be related to extracellular accumulation of bioactive substances during storage. The glycoprotein, soluble urokinase plasminogen activator receptor (suPAR), which is located in specific granules of neutrophils, plays a role in inflammation and remodelling of the extracellular matrix. Using enzyme-linked immunosorbent assay, suPAR was determined in serum, plasma and blood cell lysates. In addition, suPAR was measured in whole blood (WB), buffy-coat-depleted saline-adenine-glucose-mannitol (SAGM) blood, platelet-rich plasma (PRP) and buffy-coat-derived platelet (BCP) pools with and without pre-storage leucofiltration, and in non-filtered WB, SAGM blood and platelet concentrates prepared using apheresis (APC) at different time points during storage. Mean suPAR concentration was significantly higher in cell lysates, compared to that in a corresponding serum (P = 0.007) and in plasma samples (P = 0.004). Mean suPAR levels in WB, BCP and SAGM were significantly reduced using leucofiltration (WB: 3.4 versus 2.0 ng mL(-1); BCP: 1.6 versus 1.1 ng mL(-1); SAGM: 2.8 versus 0.19 ng mL(-1)), whereas no difference was observed in PRP. In non-filtered WB, SAGM and APC, extracellular suPAR accumulated significantly in a storage-time-dependent manner (WB: P < 0.01; SAGM: P < 0.001; APC: P < 0.001). The present study demonstrates that cell lysates contain significantly more suPAR, compared to both serum and plasma. This can be explained by the release of suPAR from intracellular granules during cell lysis. The amount of suPAR is significantly increased during storage of blood transfusion components, an accumulation that is reduced using pre-storage leucofiltration. PMID- 15285728 TI - SERF: a new antigen in the Cromer blood group system. AB - The Cromer blood group system consists of eight high incidence and three low incidence antigens carried on decay-accelerating factor (DAF). This report describes the identification and characterization of a new Cromer high incidence antigen, named SERF. Sequence analyses of DNA from a Thai female whose serum contained the alloantibody to a high incidence antigen in the Cromer blood group system (anti-SERF) and from her two children were performed. Reverse transcriptase-polymerase chain reaction (RT-PCR) and sequence analysis on cDNA from the proband and PCR-restriction fragment length polymorphism analysis on DNA from Thais were also performed. To map the epitope, DAF deletion mutants were tested by immunoblotting with anti-SERF. Sequence analysis revealed a substitution of 647C>T in exon 5 DAF in the proband. The proband's two children and two of 100 Thais were heterozygotes 647C/T. Analysis using DAF deletion mutants revealed the antigenic determinant to be within short consensus repeat 3 (SCR3), which is encoded by exon 5. This study describes a novel high incidence antigen (SERF) in the Cromer blood group system characterized by the amino acid proline at position 182 in SCR3 of DAF. The SERF-negative proband has a substitution mutation that predicts for leucine at this position. SERF has been provisionally assigned the International Society of Blood Transfusion number 021.012 (CROM 12). PMID- 15285729 TI - The first probable case of platelet transfusion-transmitted visceral leishmaniasis. AB - Visceral leishmaniasis (VL; kala-azar), a life-threatening infection of the mononuclear phagocytic system, is transmitted by the bite of infected sand flies. Though peripheral parasitaemia is documented for Leishmania spp. causing VL, reports of transfusion-transmitted infections are rare. A case of probable platelet transfusion-acquired VL is reported from India and issues related to transfusion safety in endemic areas are discussed. PMID- 15285730 TI - The integration of data logging and transport technology to provide waste-free emergency transfusion support to distant hospital facilities. PMID- 15285731 TI - Recurrent paroxysmal cold haemoglobinuria. PMID- 15285734 TI - Some international aspects of nursing. PMID- 15285733 TI - Lessons from Denmark on caring for the elderly. PMID- 15285735 TI - Global leaders team up for better world health. PMID- 15285739 TI - Nurse migration: let's tackle the real issues. PMID- 15285740 TI - International collaboration to address common problems in health care: processes, practicalities and power. AB - This article reports on an international collaborative study into the management of chronic leg ulcers by nurses working in the community in Kronoberg, Sweden and the East Riding and Hull, UK. The management of leg ulcers like many other services provided by nurses working in the community is one that is growing out of community need, often without matching resources. This article reports on some of the processes that were involved and some of the practical considerations that were faced and addressed in formulating and conducting an international collaborative study. The article also explores the 'power' international collaboration brings to the research enterprise. PMID- 15285741 TI - Nursing the dying: essential elements in the care of terminally ill patients. AB - AIM: To verify those aspects of care that nurses view as important when assisting patients beyond therapeutic possibilities and who are not under intensive care. OBJECTIVES: (1) To find out how nurses cope with daily confrontation with the death and suffering of dying patients, (2) To identify whether nurses feel it is important to have communication skills in order to assist the terminally ill patient, (3) To estimate nurses' degree of work satisfaction, and (4) To explore the humane aspects of nursing assistance to the dying. METHOD: Data were collected in January and February of 2002 by means of individual semistructured interviews with 14 nurses from the unit of haematology at a general hospital in the city of Sao Paulo, Brazil. Interviews were recorded, transcribed, and further analysed according to the qualitative method proposed by Bardin (1977). RESULTS: We found that Brazilian nurses caring for dying patients should be receiving psychological and emotional support. Results also highlighted different individual approaches in the endeavour to communicate with terminally ill patients, as well as the avoidance patterns developed by some nurses. The latter appeared to be as a result of personal difficulties in coping with the reality of human suffering and death. Finally, there is a need for better preparation in communication skills for nurses caring for terminally ill patients. CONCLUSION: Although the number of interviewed nurses in our study was small, the results corroborated the findings of other studies on the subject. PMID- 15285742 TI - A review of the literature on how important water is to the world's elderly population. AB - AIM: This article reviews the literature on how important water is to the world's elderly population. BACKGROUND: Water is a finite resource, so we must preserve the water that we have. Physiological aspects and what water requirements our bodies maintain sum up this essential nutrient for life. Dehydration is a concern in the elderly. CONCLUSIONS: Five strategies related to water intake can promote health: (1) assess for symptoms which may indicate dehydration, (2) encourage ingestion of fluids and foods to maintain an optimal fluid level, (3) be alert to physical and clinical conditions affecting hydration in the elderly, (4) consider environmental factors which may affect body fluids, electrolytes and acid-base balance, and (5) encourage methods to increase fluid consumption. PMID- 15285743 TI - Symptom management for HIV-positive persons in Norway. AB - PURPOSE: To record symptoms experienced by people with HIV/AIDS, and describe useful self-care strategies and how symptoms impact on daily life. METHOD: A questionnaire was used with self-reported answers and descriptions of how symptoms impact on daily activities and suggestions for useful self-care strategies. Four hundred and twenty-two (n = 422) HIV-positive persons from seven sites in the USA and one site in Norway (n = 20) answered the questionnaire. RESULTS: In this article subjective symptom descriptions from the Norwegian sample are presented along with self-care strategies and their effectiveness. Findings revealed the Norwegian sample to be a little older, maybe less anxious and depressed, than participants in the larger study. This difference might be explained by the structure of the national social and health care system in Norway, where treatments, medications and social services are available to all citizens without cost. Respondents described a number of related symptoms as well as their subjective explanations of what triggered the symptoms. Most of the self care strategies were learnt by trial and error. RECOMMENDATIONS: Community health providers, nurses and physicians should become more knowledgeable about the symptoms that infected persons struggle with, as well as the impact these symptoms have on routine daily self-care activities and a person's quality of life. There is need for more research about helpful self-care strategies to assist HIV-positive persons to manage their symptoms and to disseminate these to community health providers and support groups for HIV infected persons. PMID- 15285744 TI - From east to west: Nepalese women's experiences. AB - BACKGROUND: Nepal is a small mountainous South Asian country located between the nations of India and China. Forty-two per cent of the 22 million Nepalese people live in poverty. As a result, immigration to a developed country is the dream of many but available to few. Some immigrants from Nepal have arrived in Australia in recent years entering the 'Skill' stream of eligibility categories. Nepalese immigrants to Australia are predominantly young married couples with professional education qualifications. AIM: To generate knowledge of the childbirth and early experiences of Nepalese women in their mother country and in Australia. The aspect presented here is the immigration experiences of Nepalese women to Australia. METHOD: An ethnographic, grounded theory approach was used to observe and analyse the experiences of 11 Nepalese participants. FINDINGS: Analysis of data suggests that Nepalese female immigrants with the ability to comprehend and speak English and a level of education and skill required by Australia can successfully negotiate the change of culture and adapt to their new society. Major benefits of immigration for the women were the opportunities to work, become independent and to share in decision making for their family. CONCLUSION AND IMPLICATIONS FOR NURSING PRACTICE: Severance from the Nepalese joint family, a male dominant hierarchical society, and a new way of life allow a Nepalese woman to become an individual rather than a member of a collective. This study has produced transcultural information from the perspective of the educated professional Nepalese female immigrant that will assist in the provision of midwifery and nursing care. PMID- 15285745 TI - Evaluation of training of trainers courses for Oromia (Ethiopia) health professional schools' tutors. AB - BACKGROUND: In Ethiopia training tutors in the health system are part of a major health programme. A previous study disclosed that tutors felt they lacked sufficient teaching skills. AIM: The Teaching Methodology Course (TMC) described here was designed to correct these deficiencies. The aim of the study was firstly to evaluate the usefulness of questionnaires in terms of TMC quality control and secondly to evaluate whether the subcourses differed in terms of self-progress evaluation and course evaluation. CONTENT: The TMC consists of eight subcourses and is evaluated according to a quality circle described previously. METHODS: Two questionnaires were used as part of the TMC quality control, one to assess tutors' self-progress and the other to examine tutors' opinion about the course instructors' pre-defined teaching skills. RESULTS: In the questionnaires a distinction was made between 'good' and 'bad' subcourses. Moreover, the quantitative evaluation of subcourses was in accord with the tutors' written qualitative comments. CONCLUSION: The questionnaires can be used for TMC quality control in order to improve at least some subcourses. PMID- 15285746 TI - Clinical characteristics of normotensive renal transplant recipients with microalbuminuria and effects of angiotensin II type I receptor antagonist on urinary albumin excretion. AB - AIM: Microalbuminuria is typically observed in renal transplant recipients with systemic hypertension. The effects of angiotensin II type 1 receptor antagonist (losartan) on the hypertensive recipients have been evaluated. However, the clinical background of normotensive recipients with microalbuminuria and the effect of losartan administration in those subjects have not been clarified. One of the two purposes for the present study was to investigate the clinical characteristics of normotensive recipients with microalbuminuria. The other was to evaluate the effect of losartan on urinary excretion of albumin in these patients. METHODS: The clinical data and the change of the single kidney glomerular filtration rate (GFR) for the graft by radionuclide study were assessed in 13 normotensive recipients with microalbuminuria. These were compared with the data of 13 normotensive patients without microalbuminuria. The 13 recipients with microalbuminuria were treated with losartan for one year and urine excretion of albumin, N-acetyl-beta-D-glucosaminidase (NAG) and serum creatinine (S-Cr) levels were measured. RESULTS: The GFR of the grafts from donors to recipients significantly increased (30.9 to 55.2 mL/min) in microalbuminuric recipients, but did not significantly increase in the non microalbuminuric recipients. Decreases of the urinary excretion rate of albumin (351 +/- 261 at baseline to 158 +/- 14 mg/gCr at 12 months), NAG (13 +/- 5 to 10 +/- 3 IU/gCr) and S-Cr (1.7 +/- 0.6 to 1.5 +/- 0.4 mg/DL) were observed in the microalbuminuric recipients with losartan administration. CONCLUSIONS: The present study suggests that an increased single kidney GFR of the graft from the donor in situ to the recipient might be a cause of microalbuminuria in normotensive recipients. The one-year effects of losartan were observed in terms of the decrease in urinary excretion of albumin, NAG and S-Cr levels. PMID- 15285747 TI - Urodynamic effects and safety of modified intravesical oxybutynin chloride in patients with neurogenic detrusor overactivity: 3 years experience. AB - BACKGROUND: Intravesical oxybutynin chloride with hydroxypropylcellulose (HPC) (modified intravesical oxybutynin) has been reported to be effective for treatment of overactive bladder. We reported the short-term effects of modified intravesical oxybutynin previously. In the present article, we detail the results of a 3-year follow-up study of patients from our previous analysis and report the efficacy and side-effects of modified intravesical oxybutynin. METHODS: Modified intravesical oxybutynin (5 mg/10 mL, twice a day) was applied for more than 3 years to six neurogenic overactive detrusor patients (three men and three women, average age 53.3 years) who were not satisfied with oral anticholinergic agents or the other therapy. A cystometogram (CMG) was performed before, 1 week after and 3 years after the start of modified intravesical oxybutynin treatment. We evaluated the patient's satisfaction of this treatment after 4 weeks and again after 3 years. We compared the patients' answers before and after the therapy (excellent, good, fair, unchanged and worse). We also monitored systemic and topical side-effects in these patients during this period. RESULTS: CMG studies showed that two of six patients no longer exhibited uninhibited contraction 1 week after the treatment and that the cystocapacity of patients before, 1 week after and 3 years after the initial modified intravesical oxybutynin was 129.7 +/ 19.4, 283.5 +/- 40.4 and 286.8 +/- 38.1 mL, respectively. For the evaluation of patients' satisfaction with this treatment, four patients considered the therapy excellent and one patient described it as good after both 4 weeks and after 3 years. Two patients dropped out of the study; one developed left ureteral cancer (2.25 years) and the other developed ileus (1.5 years). Dry mouth and acute cystitis were observed in both patients. CONCLUSION: Modified intravesical oxybutynin is an effective and relatively safe option of therapy for overactive bladder patients. However, this therapy requires careful observation for emergent side-effects. PMID- 15285748 TI - Comparative study of novel endoluminal ultrasonography and conventional transurethral ultrasonography in staging of bladder cancer. AB - BACKGROUND: Recent advances in ultrasonic techniques have improved the image quality and diagnostic accuracy for staging of bladder cancer. The aim of this study was to assess the feasibility and usefulness of endoluminal ultrasonography (ELUS) in staging of bladder cancer, and to compare them with those of conventional transurethral ultrasonography (TUUS). METHODS: From 2000 to 2002, 19 patients with bladder cancer were evaluated by ELUS and TUUS before transurethral resection or biopsy. Clinical staging using ELUS, TUUS, computed tomography (CT) and magnetic resonance imaging (MRI) was compared with the results of pathological staging. RESULTS: In 16 of 19 patients, both ELUS and TUUS were able to diagnose tumor stage. In the remaining three patients, both methods were unable to evaluate stage of tumor. In two of these patients, this inability to evaluate tumor state was caused by a difficulty in depicting the tumor base in rectangular scanning. In the remaining patient, the inability to evaluate tumor stage was caused by a difficulty in recognizing the normal muscularis because of edema around the tumor base. Both diagnostic accuracies of ELUS and TUUS were 84%, which were superior to those of CT (44%) and MRI (82%). CONCLUSIONS: Endoluminal ultrasonography and TUUS were equally useful for staging diagnosis of bladder cancer. Because the ELUS probe is very small in diameter and can be manipulated under direct vision, it is superior to the TUUS in safety and in fine visualization. However, the main limitations of ELUS include an inability to evaluate the depth of invasion of large tumors and an inability to visualize the tumor base in the position of the bladder neck. PMID- 15285749 TI - Extracorporeal magnetic innervation treatment for urinary incontinence. AB - BACKGROUND: Extracorporeal magnetic innervation (ExMI) is a new technology used for pelvic muscle strengthening for the treatment of stress urinary incontinence. We explored whether this new technology is effective for patients with urge incontinence, as well as those with stress urinary incontinence. METHODS: We studied 20 patients with urge incontinence and 17 patients with stress urinary incontinence. The Neocontrol system (Neotonus Inc., Marietta, GA) was used. Treatment sessions were for 20 min, twice a week for 8 weeks. Evaluations were performed by bladder diaries, one-hour pad weight testing, quality-of-life surveys and urodynamic studies. RESULTS: Of the urge incontinence cases, five patients were cured (25.0%), 12 patients improved (60.0%) and three patients did not show any improvement (15.0%). Leak episodes per day reduced from 5.6 times to 1.9 times at 8 weeks (P < 0.05). Eight patients with urge incontinence recurred within 24 weeks after the last treatment (47.1%). Of the stress incontinence cases, nine patients were cured (52.9%), seven patients improved (41.1%) and one patient did not show any improvement (6%). In one-hour pad weight testing, the mean pad weight reduced from 7.9 g to 1.9 g at 8 weeks (P < 0.05). Three patients returned to the baseline values within 24 weeks after the last treatment (17.6%). No side-effects were experienced by any of the patients. CONCLUSION: Although the results for urge incontinence were less effective than for stress urinary incontinence, ExMI therapy offers a new option for urge incontinence as well as stress urinary incontinence. PMID- 15285750 TI - Effects of acupuncture for chronic pelvic pain syndrome with intrapelvic venous congestion: preliminary results. AB - BACKGROUND: The present study was designed to reveal the usefulness of acupuncture for chronic pelvic pain syndrome with intrapelvic venous congestion as evaluated by symptom scores, transrectal ultrasonography (TRUS) and magnetic resonance (MR) venography. METHODS: Ten male patients suffering from non inflammatory chronic pelvic pain syndrome (NIH category IIIB) with intrapelvic venous congestion were treated using acupuncture. Eight patients had previously received pharmacotherapy, which was unsuccessful. Acupuncture was performed using disposable stainless steel needles, which were inserted into the bilateral BL-33 points and rotated manually for 10 min. The treatment was repeated every week for 5 weeks without other therapeutic maneuvers. Results from TRUS and MR venography, as well as clinical symptoms based on the NIH chronic prostatitis symptom index (NIH-CPSI) and the international prostate symptom score (IPSS), were compared before and after the treatment. RESULTS: No side-effects were recognized throughout the treatment period. The average pain and QOL scores of the NIH-CPSI 1 week after the 5th acupuncture treatment decreased significantly (P < 0.05 and P < 0.01, respectively) compared with the baseline. The maximum width of the sonolucent zone 1 week after the 5th treatment also decreased significantly (P < 0.01, compared with the baseline). Intrapelvic venous congestion demonstrated by MR venography was significantly improved in four patients. CONCLUSION: This study provided novel information concerning the therapeutic effects of acupuncture on non-inflammatory chronic pelvic pain syndrome. PMID- 15285751 TI - Transrectal ultrasound-guided transperineal 14-core systematic biopsy detects apico-anterior cancer foci of T1c prostate cancer. AB - AIM: The optimal biopsy strategy for prostate cancer detection, especially in men with isolated prostate-specific antigen (PSA) elevation, remains to be defined. We evaluated diagnostic yield and safety of transrectal ultrasound (TRUS)-guided transperineal systematic 14-core biopsy and compared the spatial distribution of cancer foci detected with this technique in men with and without abnormality on digital rectal examination (DRE). METHODS: In a prospective study, 289 men aged between 50 and 87 years (median age, 70 years) underwent TRUS-guided transperineal systematic 14-core prostate biopsy because of elevated PSA and/or abnormal DRE findings. Using the fan technique, 12 cores from the peripheral zone and two cores from the transition zone were obtained systematically. To characterize the spatial distribution of cancer positive cores, site-specific overall and unique cancer detection rates were compared between stage T1c and T2 cancers. RESULTS: Prostate cancer was detected in 105 of the 289 patients (36%). Major complications requiring prolonged hospital stay or re-hospitalization during a 4-week postbiopsy period were rare (1.4%). Sixty-seven stage T1c cancers were identified. These cancers were associated with significantly lower PSA and a smaller number of cancer positive cores when compared with stage T2 cancers (n= 38). The overall cancer detection rate was highest at the anterior peripheral zone and the posterior peripheral zone in stage T1c and stage T2 cancers, respectively. The unique cancer detection rate at the anterior peripheral zone was significantly higher in stage T1c cancers than in stage T2 cancers. Therefore, when the prostate is extensively biopsied using the transperineal approach, cancer positive cores are characteristically distributed anteriorly in stage T1c cancers and posteriorly in stage T2 cancers. CONCLUSIONS: TRUS-guided transperineal systematic 14-core biopsy showed an apico-anterior distribution of cancer foci in stage T1c prostate cancers. PMID- 15285752 TI - Health related quality of life in Japanese men after radical prostatectomy or radiation therapy for localized prostate cancer. AB - BACKGROUND: We performed a retrospective survey of general and disease specific health-related quality of life (HRQOL) after radical prostatectomy (RP) and external beam radiotherapy (XRT) in Japanese men. METHODS: A total of 186 patients underwent RP and 78 underwent XRT for clinically localized prostate cancer between 2000 and 2002. We measured the general and disease specific HRQOL with the MOS 36-Item Health Survey and the University of California, Los Angeles Prostate Cancer Index, respectively. Each treatment group was further divided into four subgroups according to the time scale. RESULTS: Patients from the RP group were significantly younger than those from the XRT group. The tumor characteristics differed significantly in their distributions among the treatment groups. Patients undergoing XRT had low scores in most of the general measures of HRQOL just after treatment, but after 6 months there were no differences between the treatment groups, except for the physical domains. The RP group was associated with worse urinary function, whereas the XRT group had worse bowel function and bother during the first 6 months after treatment. Thereafter, however, urinary and bowel domain did not differ between the groups. Both groups reported poor sexual function, although the RP group scored lower sexual bother. CONCLUSION: The patients who underwent RP had significantly worse urinary and better bowel function than those treated with XRT. Both treatment groups had decrements in sexual function throughout the post-treatment period; careful attention should be paid to this side-effect in preoperative counselling, especially in younger patients, regardless of the primary treatments. PMID- 15285753 TI - Basic study on velocity-flow urodynamics using Doppler sonography: simultaneous detection of cavitation and Doppler signals in an artificial urethral model. AB - BACKGROUND: We have developed velocity-flow urodynamics using Doppler sonography based on the hypothesis that microbubbles formed in the urethra are responsible for Doppler signals. In order to confirm this hypothesis derived from Bernoulli's principle, we investigated the simultaneous detection of cavitation noise and Doppler signals in an experimental system. METHODS: An experimental circuit was built in which a stenosis was created using a glass or silicon tube with tap water used as the sample fluid. Doppler signals, pressure before and after the stenosis, flow rate, flow velocity and cavitation noise were measured. Direct detection of cavitation with a high-speed charged-coupled device (CCD) camera was conducted in the glass tube. The relationship between cross-sectional area and flow velocity in terms of the detection of Doppler signals was analyzed in the silicon tube study. RESULTS: In the glass tube study, a high-speed CCD camera clearly detected masses of microbubbles associated with cavitation. The range of flow rates creating cavitation completely corresponded with those producing Doppler signals detected by ultrasonography. A similar correlation was observed in the silicon tube study, which showed that a low flow velocity of 41.5 cm/sec through a stenosis with a cross-sectional area of 20 mm(2) created Doppler signals at a flow rate of 8.3 mL/sec. CONCLUSION: The results of the present study confirmed that microbubbles created in flowing urine are responsible for Doppler signals. Measurement of velocity-flow urodynamics has great potential to become a non-invasive and reliable alternative to conventional pressure- flow urodynamic studies. PMID- 15285754 TI - Resection of pulmonary metastases following chemotherapy for high stage testicular tumors. AB - BACKGROUND: Our objective was to analyze retrospectively our experience with 19 patients who had metastatic germ cell testicular tumor and had undergone resection of pulmonary metastases following chemotherapy. We wished to determine the necessity of thoracic surgery on these patients. METHODS: Of 103 patients in need of postchemotherapeutic surgery for metastatic germ cell testicular tumors, 19 patients (mean age 31) underwent surgery for thoracic masses following cis platin based chemotherapy. Resection of pulmonary metastases was performed on patients with normal tumor markers after chemotherapy, who did not achieve complete radiological remission. Histopathological findings, correlation with the pathology of abdominal surgery and probable prognostic factors for disease-free and overall survivals were evaluated. RESULTS: Disease-free and overall survival rates were 14/19 (73%) and 16/19 (84%), respectively, within a median follow-up time of 30 months (15-212 months). Patients with and without viable tumor cells in their thoracic histopathological specimen had 40% and 85% disease-free survival rates, respectively (P < 0.05). Eight patients had both abdominal and thoracic postchemotherapy surgery. Only two (25%) of these patients had the same histopathological features at both sites. CONCLUSIONS: All patients with residual thoracic masses must be considered candidates for surgery, because there are no predictive factors to determine the thoracic pathology without surgery. With the resection of the pulmonary metastases only, surgery can be performed without significant morbidity and is essential to select patients for further chemotherapy, to remove all visible masses and to provide histopathological confirmation. Patients with viable tumor cells in the thoracic surgical specimen have a poor prognosis. PMID- 15285755 TI - Risk factors in past histories and familial episodes related to development of testicular germ cell tumor. AB - BACKGROUND: A retrospective study was conducted to examine the host factors of 240 testicular germ cell tumor patients. This study was performed to address a new theory proposed by Skakkebaek called testicular dysgenesis syndrome which claims that cryptorchism, hypospadias, poor semen quality and testicular germ cell tumors are symptoms of an underlying testicular dysgenesis in uterus. METHODS: The past health histories and familial episodes of 240 testicular germ cell tumor patients were examined. The past health histories included cryptorchism, hypospadias, infertility, atrophic testis and inguinal hernia. RESULTS: Of the 240 patients, 13 (5.4%) had a history of cryptorchism or orchidopexy. Two (0.8%) showed existence of hypospadias or had experienced urethroplasty. Among 129 married couples, 104 (80.6%) couples were fertile. Three (1.3%) patients developed testicular tumors after they were diagnosed as infertile or came to the hospital with the complaints of infertility. Four (1.7%) had contralateral atrophic testis. 19 (7.9%) had experienced inguinal herniorrhaphy before age 15. Three (1.3%) had testicular germ cell tumor patients among their family or relatives. CONCLUSIONS: The testicular germ cell tumor patients showed a considerable incidence of complications such as cryptorchism, hypospadias and incomplete closure of processus vaginalis. Cryptorchism, perinatal factors and familial factors could be risks for developing testicular germ cell tumors. PMID- 15285756 TI - Antitumor effect of simultaneous transfer of interleukin-12 and interleukin-18 genes and its mechanism in a mouse bladder cancer model. AB - BACKGROUND: The objectives of this study were to evaluate the antitumor effects of the simultaneous introduction of interleukin 12 (IL-12) and IL-18 genes into a mouse bladder cancer cell line (MBT2). We intended to compare these with those of either gene alone and to investigate the mechanism of the effects induced by the transfer of IL-12 and/or IL-18 genes in this model system. METHODS: We transfected the IL-12 and/or IL-18 genes into MBT2 cells by the liposome-mediated gene transfer method. We confirmed the secretion of IL-12 and/or IL-18 by enzyme linked immunosorbent assay. Parental (MBT2/P), IL-12-transfected (MBT2/IL-12), IL 18-transfected (MBT2/IL-18) or both IL-12- and IL-18-transfected (MBT2/Both) cells were subcutaneously or intravenously injected into syngeneic C3H mice. To analyze the mechanism of tumor rejection, these clones were subcutaneously injected into naive nude mice and those depleted with natural killer (NK) cells by antibody. RESULTS: MBT2/IL-12, MBT2/IL-18 and MBT2/Both were completely rejected when they were injected subcutaneously or intravenously into syngeneic mice. However, MBT2/IL-12, but not MBT2/IL-18, could grow in nude mice. Moreover, the antitumor effect of MBT2/IL-18 was partially abrogated when injected into nude mice of which NK cells were depleted by antibody treatment. MBT2/Both was completely rejected in both nude mice with and without NK cells. CONCLUSION: The results of the present study indicate that T cells and NK cells seem to play important roles in the antitumor effects by the secretion of IL-12 and IL-18, respectively, and MBT2/Both possesses both mechanisms. PMID- 15285757 TI - Cystic hamartoma of the renal pelvis. AB - Cystic hamartoma of the renal pelvis is a rare benign tumor in the same category as mixed epithelial and stromal tumors. We present a 33-year-old woman with a solid and cystic intrarenal tumor extending into the renal pelvis. She underwent radical nephrectomy and ureterectomy under the diagnosis of renal tumor or renal pelvic tumor. Histopathologically, the tumor was composed of a biphasic proliferation of epithelial and mesenchymal elements. We believe the present case is best classified as a cystic hamartoma of the renal pelvis in the category of mixed epithelial and stromal tumors because of the coexistence of hamartomatous lesions, such as the proliferation of adipose cells and well to poorly differentiated fibromuscular lesions. PMID- 15285758 TI - BK virus subtype 1 infection associated with tubulointerstitial nephritis in a renal allograft recipient. AB - The BK polyomavirus (BKV) infects most of the human population, but clinically relevant infections are usually limited to individuals who are in an immunosuppressed state. The significance of BKV infection was investigated in a 50-year-old man who underwent cadaveric kidney transplantation and was treated with tacrolimus, mycophenolate mofetil and prednisolone. By staining renal biopsy specimens with a monoclonal antibody against BK large T antigen, we were able to observe the relationship between the appearance of the BKV antigen and the extent of immunosuppression in this patient. We also determined that BKV belonged to genotype I by analysis of viral DNA from the patient's urine. PMID- 15285759 TI - Massive bilateral perirenal hematoma following urinary catheterization for urinary obstruction. AB - Urinary tract obstruction is a common problem associated with many complications. Decompression of an enlarged bladder has been associated with several complications, mainly vesicular bleeding. We report a case of a 42-year-old male patient who developed bilateral renal subcapsular hematomas secondary to relief of an extremely enlarged bladder. PMID- 15285760 TI - Intracranial aspergillosis in a non-immunocompromised patient treated for muscle invasive bladder cancer. AB - We report a case of intracerebral Aspergillosis in a patient undergoing radical cystectomy for the treatment of muscle-invasive bladder cancer who did not reveal any deterioration of the immune system. Aspergillus fumigatus is an ubiquitously present, airborne fungus that tends to infect the upper respiratory tract. However, the latter was not observed in the patient presented. Complications in the form of an involvement of the central nervous system are very rarely recognized as a result of an Aspergillus infection and primarily occur in patients who are not immunologically competent. To our knowledge, we present the first case of intracerebral invasive Aspergillosis in an otherwise healthy patient diagnosed with an urological malignancy. PMID- 15285761 TI - An unusual complication of Burch colposuspension. AB - Burch colposuspension remains one of the successful operations for genuine stress incontinence. We report a patient who developed an intravesical foreign body granuloma post-Burch colposuspension. Any patient developing unexplained lower urinary tract symptoms following bladder or pelvic surgery for incontinence must be evaluated endoscopically in order to exclude this complication. PMID- 15285762 TI - Primary extragonadal germ cell tumor of the prostate in a young man. AB - We report on a case of malignant prostatic teratoma treated with radical cystoprostatectomy and cisplatin-based chemotherapy because of intraoperative tumor rupture. During chemotherapy the patient suffered acute myocardial infarction and was treated with percutanous coronarangiography and stenting. PMID- 15285763 TI - Small cell carcinoma arising from the proximal urethra. AB - Primary neuroendocrine carcinomas of the lower urinary tract are distinctly rare, locally aggressive neoplasms with a high rate of metastasis. We present a case of primary small cell carcinoma of the urethra occurring in a 64-year-old man. The clinical, histological and immunohistochemical features of urethral small cell carcinoma are highlighted with respect to the differential diagnosis of neuroendocrine and other urethral tumors. The possible histogenesis of urethral small cell carcinoma, reported at this location in only a small number of cases, is briefly discussed. We favor an origin from pluripotent epithelial stem cells as one of the possible histogenic pathways. PMID- 15285764 TI - Successful treatment for squamous cell carcinoma of the female urethra with combined radio- and chemotherapy. AB - We report on two cases of women with locally advanced squamous cell carcinoma of the urethra. Patient 1 also displayed regional lymph node metastasis. Treatment comprised combined radiotherapy to 60 Gy and chemotherapy with 5-fluorouracil and cisplatin. Complete response was obtained in both patients, including the inguinal lymph nodes of Patient 1. Patient 1 experienced recurrent inguinal lymph node metastasis on the contralateral side at 42 months after initial treatment, and the same treatment was performed followed by surgical excision. Both patients remain alive with no evidence of disease, at 12 months after recurrence in Patient 1, and at 27 months after treatment in Patient 2. PMID- 15285765 TI - Relapse of brucellosis simulating testis tumor. AB - We report on a case of a 32-year-old man referred for evaluation of a painless left testicular mass suggesting a testicular tumor. Previous history was uneventful except for a 3-year history of systemic brucellosis without epididymo orchitis. Radical inguinal orchidectomy was performed. Clinical and histopathological findings indicated a brucellar abscess of the left testis. Even in the absence of systemic symptoms, the possible relapse of brucellosis as an abscess formation in the testis should be considered as a rare cause of testicular mass in patients who live in endemic regions. PMID- 15285766 TI - Xanthogranulomatous orchitis with scrotal fistulas. AB - Xanthogranulomatous orchitis is an extremely rare inflammatory change of testis which is difficult to distinguish from testicular tumor. We report on a 21-year old man who presented with left testicular swelling and pyogenic discharge from the scrotum. Testicular tumor markers were normal. Scrotal ultrasonography showed a testicular tumor in the left testis. Because of severe adhesion between the scrotum and intrascrotal structures, radical orchiectomy combined with hemiscrotectomy was performed to exclude possible malignancy. Histopathological findings showed xanthogranulomatous orchitis. PMID- 15285767 TI - Extramammary Paget's disease of the vulva subclinically extending to the bladder neck: correct staging obtained with endoscopic urethral biopsy. AB - We report a 65-year-old woman with extramammary Paget's disease which recurred on the skin flap. We performed a mapping-biopsy of the urethra by ureteroscopy to select the appropriate treatment. This method is presumed to be useful for determining the margin, deciding the correct staging for extramammary Paget's disease and for making a judgement on appropriate therapy. PMID- 15285768 TI - TRIPSTAR: prioritizing oral triptan treatment attributes in migraine management. AB - Migraine can be associated with severe pain and is often very disabling. Optimal treatment should provide rapid and sustained, complete pain relief, be well tolerated and restore normal function. The seven commercially available triptans show differences in performance on individual treatment attributes. The TRIPSTAR multiattribute decision model compares the profiles of the oral triptans, using efficacy and tolerability data weighted for importance, to identify if measurable differences are clinically relevant. Application of the TRIPSTAR model was demonstrated at the Migraine Trust International Symposium 2002, where delegates collectively prioritized treatment attributes according to the needs of a specific patient case history. The TRIPSTAR model identified the preferred triptans for this patient. These three triptans, almotriptan 12.5 mg, eletriptan 80 mg and rizatriptan 10 mg, standout in a triptan meta-analysis, three TRIPSTAR surveys and in a demonstration of the TRIPSTAR model at a symposium in the USA. Taken together the findings suggest that some differences amongst triptans may be relevant in clinical practice. PMID- 15285769 TI - Efficacy of levetiracetam in pharmacoresistant continuous spikes and waves during slow sleep. AB - OBJECTIVE: To evaluate the efficacy of levetiracetam (LEV) in continuous spikes and waves during slow sleep (CSWS). Despite first description dates back to 1971, no agreement exists about CSWS treatment. The condition is rare and controlled clinical trials are very difficult to perform, so the reports about efficacy of different drugs are anecdotal. PATIENTS AND METHODS: We introduced LEV in three children affected by symptomatic focal epilepsy and pharmacoresistant CSWS and evaluated clinical, neuropsychological and electroencephalographic outcome. RESULTS: Two cases responded completely, one case showed only a mild reduction of spikes and waves during slow sleep. CONCLUSION: Even if our report is anecdotal, LEV expands the spectrum of antiepileptic drugs that can be used for the treatment of CSWS. LEV efficacy should be confirmed in larger series. PMID- 15285770 TI - Bilateral mesial temporal lobe epilepsy: comparison of scalp EEG and hippocampal MRI-T2 relaxometry. AB - OBJECTIVE: Bilateral hippocampal abnormality is frequent in mesial temporal lobe sclerosis and might affect outcome in epilepsy surgery. The objective of this study was to compare the lateralization of interictal and ictal scalp EEG with MRI T2 relaxometry. MATERIAL AND METHODS: Forty-nine consecutive patients with intractable mesial temporal lobe epilepsy (MTLE) were studied with scalp EEG/video monitoring and MRI T2 relaxometry. RESULTS: Bilateral prolongation of hippocampal T2 time was significantly associated with following bitemporal scalp EEG changes: (i) in ictal EEG left and right temporal EEG seizure onsets in different seizures, or, after regionalized EEG onset, evolution of an independent ictal EEG over the contralateral temporal lobe (left and right temporal asynchronous frequencies or lateralization switch; P = 0.002); (ii) in interictal EEG both left and right temporal interictal slowing (P = 0.007). Bitemporal T2 changes were not, however, associated with bitemporal interictal epileptiform discharges (IED). Lateralization of bilateral asymmetric or unilateral abnormal T2 findings were associated with initial regionalization of the ictal EEG in all but one patient (P < 0.005), with lateralization of IED in all patients (P < 0.005), and with scalp EEG slowing in 28 (82,4%) of 34 patients (P = 0.007). CONCLUSION: Our data suggest that EEG seizure propagation is more closely related to hippocampal T2 abnormalities than IED. Interictal and ictal scalp EEG, including the recognition of ictal propagation patterns, and MRI T2 relaxometry can help to identify patients with bitemporal damage in MTLE. Further studies are needed to estimate the impact of bilateral EEG and MRI abnormal findings on the surgical outcome. PMID- 15285771 TI - CSF outflow resistance as predictor of shunt function. A long-term study. AB - OBJECTIVES: Little is known about the long-term impact of a CSF shunt on the human CSF hydrodynamic system. In patients with communicating hydrocephalus, patency of the shunt system is not regularly assessed. In order to reveal postoperative changes in the CSF hydrodynamic system, we prospectively investigated the features of the system in shunted patients with idiopathic adult hydrocephalus syndrome (IAHS) over a 3-year period. MATERIAL AND METHODS: Thirty two patients with IAHS were studied at baseline and at 3, 9, 18 and 36 months postoperatively. All patients were operated on with a Hakim standard valve system and a ventriculo-peritoneal approach. At each visit, the patients were investigated with computed tomography/magnetic resonance imaging, video recording of gait and a lumbar constant pressure infusion method. Six brand-new Hakim valves were investigated in a bench test, and these results were compared with the in vivo results. RESULTS: After shunt insertion, the CSF outflow resistance was significantly decreased (13.6 vs 3.8 mmHg/ml/min). The mean outflow resistance of the six in vitro tested valves corresponded to the postoperative values. The variation in resistance in the functioning shunts at different postoperative investigations was negligible. The mean intracranial pressure in the supine position was 13.8 mmHg at the baseline and 14.3, 14.5, 14.8 and 15.7 mmHg at the follow-up visits, respectively. Postoperatively, the CSF pressure after sitting for 10 min (i.e. 'siphoning effect') decreased significantly (mean decrease -5.3, -5.4, -4.7 and -5.3 mmHg at each visit, respectively). Shunt related complications occurred in seven patients (underdrainage four, overdrainage three). Despite a functioning shunt, eight patients never improved and another nine patients first improved but later deteriorated. CONCLUSIONS: The CSF outflow resistance is much decreased postoperatively and does not alter over time in patients with functioning shunts. We consider CSF outflow resistance to be a reliable indicator of shunt function and of fundamental importance to distinguish a dysfunctioning shunt from an aggravation of the primary condition in patients with communicating hydrocephalus. The unaltered intracranial pressure together with the in vitro model results, suggests that the intra-abdominal pressure might be a major determinant of the postoperative intracranial pressure. PMID- 15285772 TI - Internuclear ophthalmoplegia: causes and long-term follow-up in 65 patients. AB - OBJECTIVES: To analyze the causes and long-term clinical outcome of internuclear ophthalmoplegia (INO) in a consecutive series of 65 patients. PATIENTS AND METHODS: From a clinical registry of a neuroophthalmological department, patients with diagnosis of INO were retrospectively identified. Patients were classified into three groups: unilateral INO, bilateral INO, and one-and-a-half syndrome. RESULTS AND CONCLUSIONS: Thirty-three men (50.8%) and 32 women (49.2%), with a mean age of 38.4 years were included in the study. INO was unilateral in 36 patients (55.4%), bilateral in 22 (33.8%) and one-and-a-half syndrome in seven (10.8%). The most common causes were vascular (36.9%), multiple sclerosis (32.3%), and infectious diseases (13.8%). Resolution of INO was documented in 32 patients (49.2%): 15 patients showed INO resolution during the first 3 months and 17 patients in 3-9 months. INO persisted in 33 patients (50.8%) even after a follow-up longer than 12 months. PMID- 15285773 TI - Repeated vs single physical maneuver in benign paroxysmal positional vertigo. AB - OBJECTIVES: To evaluate the effectiveness and possible side effects of a single session of repeated particle repositioning maneuver (PRM) to treat posterior canal benign paroxysmal positional vertigo (BPPV) and the usefulness of post treatment restrictions. MATERIALS AND METHODS: A total of 125 consecutive patients with idiopathic BPPV participated in the study. Fifty patients received a single session of repeated PRM only (group I). Results were compared with those of 50 patients with BPPV who received a single PRM (group IIb), and 25 patients who received a single PRM followed by the use of a neck collar and keeping the head upright for 48 h (group IIa). RESULTS: Forty-six patients (92%) of group I, 40 patients (80%) of group IIb, and 21 patients (84%) of group IIa were completely free of signs and symptoms when re-examined 1 week after treatment. Transient nausea and disequilibrium following treatment were reported equally in all subgroups and well tolerated. Nearly all patients of group IIa considered the post-treatment restrictions very inconvenient. CONCLUSIONS: A single session of repeated physical procedure seems to be clinically superior to one single maneuver and well tolerated. Additional post-treatment measurements are inconvenient and should be abandoned. PMID- 15285774 TI - Surgery for unruptured intracranial aneurysms in a low-volume neurosurgical unit. AB - OBJECTIVES: To evaluate outcomes after surgical treatment for unruptured intracranial aneurysms in a low-volume neurosurgical unit. MATERIAL AND METHODS: Consecutive patients operated during the years 1988-98 at the Department of Neurosurgery, University Hospital of North Norway, were studied retrospectively. Procedure-related complications were registered. The modified Rankin Scale and the Glasgow Outcome Scale, Extended version (GOS-E), were used for assessment of outcome. RESULTS: Thirty-six aneurysms were repaired in 32 patients during 34 surgical procedures. Surgery-related central nervous system complications occurred in eight (25%) patients. The complication rate was 35% for the first 17 procedures, and 12% for the last 17. The latter group of procedures were performed during a period of 3 years, whereas the former group was spread over a period of 8 years. A favourable outcome (GOS-E score 6-8) was reached in 27 (84%) patients. CONCLUSION: Decision-making in patients with unruptured intracranial aneurysms must be based on knowledge about institution-specific complication rates, preferably followed on a prospective basis. PMID- 15285775 TI - Src family kinase-inhibitor PP2 reduces focal ischemic brain injury. AB - OBJECTIVES: To investigate the neuroprotective potential of the Src family kinase (SFK) inhibitor 4-amino-5-(4-chlorophenyl)-7-(t-butyl) pyrazolo(3,4-d)pyrimidine (PP2) in transient focal cerebral ischemia in the rat. MATERIAL AND METHODS: Sprague-Dawley rats were exposed to transient (90 min) middle cerebral artery occlusion (MCAO) and evaluated after 1 day of survival. PP2 (1.5 mg/kg i.p.) or vehicle was given 30 min after MCAO. The lesions were examined with magnetic resonance imaging (MRI), tri-phenyl tetrazolium chloride (TTC) staining and the functional outcome was determined using neurological scoring according to Bederson et al. RESULTS: PP2-treated rats showed approximately 50% reduction of infarct size on T2-weighted MRI and in TTC staining compared with controls (P < 0.05). Moreover, the neurological score was better in the PP2 group than controls (P < 0.05). CONCLUSION: PP2 is a potential neuroprotective agent in cerebral ischemia-reperfusion. The interference of PP2 with SFKs and/or other pathways remains to be elucidated. PMID- 15285776 TI - Factors of importance for weight loss in elderly patients with Parkinson's disease. AB - OBJECTIVE: Weight loss is reported frequently in patients with Parkinson's disease (PD). The objective of this study was to find the underlying factors of this phenomenon. PARTICIPANTS AND METHODS: Twenty-six L-dopa-treated patients with PD and 26 age- and sex-matched healthy controls were assessed twice within a 1-year interval. Body weight, body fat mass, resting energy expenditure, physical activity, energy intake, thyroid hormones and cognitive function were investigated. RESULTS: Nineteen (73%) of the PD patients lost body weight, although energy intake and the time for rest increased. Weight loss was most marked in patients with more severe PD symptoms and in whom cognitive function had decreased. Multiple regression analyses showed that determinants for weight loss were female gender, age and low physical activity. CONCLUSION: Weight loss was common in PD patients, in spite of the increased energy intake and was most obvious in patients with increased PD symptoms and decreased cognitive function. PMID- 15285777 TI - Levodopa effect on [18F]fluorodopa influx to brain: normal volunteers and patients with Parkinson's disease. AB - OBJECTIVES: Levodopa is the immediate precursor of dopamine and the substrate for DOPA decarboxylase, an enzyme subject to regulation in living brain. To test whether this regulation changes in disease, we used Positron Emission Tomography (PET) with parametric mapping to measure the effect of levodopa on the net clearance of [(18)F]fluorodopa to brain (K, ml/g/min). METHODS: Five patients with early Parkinson's disease with pause of medication for 3 days and six age matched healthy volunteers were studied in a baseline condition and after levodopa challenge. RESULTS: Levodopa (200 mg as Sinemet) increased the magnitude of the net clearance K in the left and right putamen of the healthy volunteers by 11% relative to the baseline condition. In contrast, resumption of medication with levodopa did not significantly alter the magnitude of K in putamen of the Parkinson's disease patients. Compartmental analysis was used to probe the physiological basis of the activation of K: levodopa treatment increased by 15% the apparent distribution volume of [(18)F]fluorodopa in cerebellum (, ml/g) of both patients and control subjects, without significantly altering the unidirectional blood-brain clearance (, ml/g/min) or the relative activity of DOPA decarboxylase (, min(-1)) in putamen. CONCLUSION: We conclude that levodopa treatment increases the distribution volume of [(18)F]fluorodopa in brain, increasing its availability for utilization in dopamine terminals. We speculate that levodopa act as a direct beta-adrenergic agonist at receptors regulating the permeability of the blood-brain barrier to levodopa. However, the PET analytical method was without sufficient power to detect the consequent increase in magnitude of K in brain of only five Parkinson's disease subjects. PMID- 15285778 TI - Dejerine-Sottas' neuropathy caused by the missense mutation PMP22 Ser72Leu. AB - OBJECTIVE: To describe a patient with the Dejerine-Sottas' syndrome due to a de novo Ser72Leu amino acid substitution in the PMP22 protein and summarize the phenotype associated with this frequent mutation. CASE REPORT: The proband has a medical history of early onset, severe, and progressive demyelinating neuropathy, accompanied by mild ptosis and limitations of eye movements. Ulnar nerve motor conduction velocities were extremely reduced (2.6 and 2.2 m/s), and the sural nerve biopsy showed onion bulbs and thinly myelinated axons. Duplication of chromosome 17p11.2 was ruled out, and the Ser72Leu substitution was found upon sequencing the PMP22 gene. CONCLUSION: The Ser72Leu substitution is being confirmed as the most frequent point mutation in the PMP22 gene. This 'hot spot' should be considered in the strategy of looking for point mutations in the hereditary demyelinating neuropathies. PMID- 15285779 TI - Embolization of indirect carotid-cavernous sinus fistulas using the superior ophthalmic vein approach. AB - OBJECTIVES: In indirect carotid-cavernous sinus fistulas (CCF), abnormal connections exist between tiny dural branches of the external and/or internal carotid system and the cavernous sinus. Usually this kind of fistula occurs spontaneously and is characterized by a low shunt volume. Alternative vascular approaches for embolization are required when standard interventional neuroradiological access via arterial or transfemoral venous routes is not feasible. PATIENTS AND METHODS: Two symptomatic patients with indirect CCFs are described. Transarterial and transfemoral venous approach was unsuccessful or resulted in incomplete occlusion of the CCF. Therefore, the superior ophthalmic vein (SOV) was surgically exposed and retrograde catheterized to allow the delivery of platinum coils to the fistula point via a microcatheter. RESULTS: Complete fistula obliteration was accompanied by recovery of the clinical symptoms. CONCLUSION: The surgical SOV approach might be sufficient when standard neuroradiological procedures do not succeed. The technique is safe and effective when performed by an interdisciplinary team. PMID- 15285780 TI - Stress echocardiography in heart failure. AB - Echocardiography has the ability to noninvasively explore hemodynamic variables during pharmacologic or exercise stress test in patients with heart failure. In this review, we detail some important potential applications of stress echocardiography in patients with heart failure. In patients with coronary artery disease and chronic LV dysfunction, dobutamine stress echocardiography is able to distinguish between viable and fibrotic tissue to make adequate clinical decisions. Exercise testing, in combination with echocardiographic monitoring, is a method of obtaining accurate information in the assessment of functional capacity and prognosis. Functional mitral regurgitation is a common finding in patients with dilated and ischaemic cardiomyopathy and stress echocardiography in the form of exercise or pharmacologic protocols can be useful to evaluate the behaviour of mitral regurgitation. It is clinical useful to search the presence of contractile reserve in non ischemic dilated cardiomyopathy such as to screen or monitor the presence of latent myocardial dysfunction in patients who had exposure to cardiotoxic agents. Moreover, in patients with suspected diastolic heart failure and normal systolic function, exercise echocardiography could be able to demonstrate the existence of such dysfunction and determine that it is sufficient to limit exercise tolerance. Finally, in the aortic stenosis dobutamine echocardiography can distinguish severe from non-severe stenosis in patients with low transvalvular gradients and depressed left ventricular function. PMID- 15285781 TI - A simulation model of African Anopheles ecology and population dynamics for the analysis of malaria transmission. AB - BACKGROUND: Malaria is one of the oldest and deadliest infectious diseases in humans. Many mathematical models of malaria have been developed during the past century, and applied to potential interventions. However, malaria remains uncontrolled and is increasing in many areas, as are vector and parasite resistance to insecticides and drugs. METHODS: This study presents a simulation model of African malaria vectors. This individual-based model incorporates current knowledge of the mechanisms underlying Anopheles population dynamics and their relations to the environment. One of its main strengths is that it is based on both biological and environmental variables. RESULTS: The model made it possible to structure existing knowledge, assembled in a comprehensive review of the literature, and also pointed out important aspects of basic Anopheles biology about which knowledge is lacking. One simulation showed several patterns similar to those seen in the field, and made it possible to examine different analyses and hypotheses for these patterns; sensitivity analyses on temperature, moisture, predation and preliminary investigations of nutrient competition were also conducted. CONCLUSIONS: Although based on some mathematical formulae and parameters, this new tool has been developed in order to be as explicit as possible, transparent in use, close to reality and amenable to direct use by field workers. It allows a better understanding of the mechanisms underlying Anopheles population dynamics in general and also a better understanding of the dynamics in specific local geographic environments. It points out many important areas for new investigations that will be critical to effective, efficient, sustainable interventions. PMID- 15285782 TI - Adenoid cystic carcinoma of the parotid metastasizing to liver: case report. AB - BACKGROUND: Adenoid cystic carcinoma is a rare malignant parotid tumor. Metastasis can occur even a decade or more after initial treatment of the primary. CASE PRESENTATION: We report a 60 year old female patient who presented with adenoid cystic carcinoma of the parotid gland. She underwent a total conservative parotidectomy followed by adjuvant radiotherapy. While on follow up, patient developed multiple liver metastases which manifested three years later. Patient lived for another two years before she died of her disease. CONCLUSIONS: Although distant metastases of adenoid cystic carcinoma develop frequently, isolated metastasis to liver is unusual. Even after manifestation of distant metastasis, patients can be expected to live for a number of years. Palliative chemotherapy can be considered in symptomatic cases while the usefulness of metastasectomy is controversial. PMID- 15285783 TI - Chronic pain self-management for older adults: a randomized controlled trial [ISRCTN11899548]. AB - BACKGROUND: Chronic pain is a common and frequently disabling problem in older adults. Clinical guidelines emphasize the need to use multimodal therapies to manage persistent pain in this population. Pain self-management training is a multimodal therapy that has been found to be effective in young to middle-aged adult samples. This training includes education about pain as well as instruction and practice in several management techniques, including relaxation, physical exercise, modification of negative thoughts, and goal setting. Few studies have examined the effectiveness of this therapy in older adult samples. METHODS/DESIGN: This is a randomized, controlled trial to assess the effectiveness of a pain self-management training group intervention, as compared with an education-only control condition. Participants are recruited from retirement communities in the Pacific Northwest of the United States and must be 65 years or older and experience persistent, noncancer pain that limits their activities. The primary outcome is physical disability, as measured by the Roland Morris Disability Questionnaire. Secondary outcomes are depression (Geriatric Depression Scale), pain intensity (Brief Pain Inventory), and pain-related interference with activities (Brief Pain Inventory). Randomization occurs by facility to minimize cross-contamination between groups. The target sample size is 273 enrolled, which assuming a 20% attrition rate at 12 months, will provide us with 84% power to detect a moderate effect size of.50 for the primary outcome. DISCUSSION: Few studies have investigated the effects of multimodal pain self management training among older adults. This randomized controlled trial is designed to assess the efficacy of a pain self-management program that incorporates physical and psychosocial pain coping skills among adults in the mid old to old-old range. PMID- 15285785 TI - Assessment of genetic diversity in Trigonella foenum-graecum and Trigonella caerulea using ISSR and RAPD markers. AB - BACKGROUND: Various species of genus Trigonella are important from medical and culinary aspect. Among these, Trigonella foenum-graecum is commonly grown as a vegetable. This anti-diabetic herb can lower blood glucose and cholesterol levels. Another species, Trigonella caerulea is used as food in the form of young seedlings. This herb is also used in cheese making. However, little is known about the genetic variation present in these species. In this report we describe the use of ISSR and RAPD markers to study genetic diversity in both, Trigonella foenum-graecum and Trigonella caerulea. RESULTS: Seventeen accessions of Trigonella foenum-graecum and nine accessions of Trigonella caerulea representing various countries were analyzed using ISSR and RAPD markers. Genetic diversity parameters (average number of alleles per polymorphic locus, percent polymorphism, average heterozygosity and marker index) were calculated for ISSR, RAPD and ISSR+RAPD approaches in both the species. Dendrograms were constructed using UPGMA algorithm based on the similarity index values for both Trigonella foenum-graecum and Trigonella caerulea. The UPGMA analysis showed that plants from different geographical regions were distributed in different groups in both the species. In Trigonella foenum-graecum accessions from Pakistan and Afghanistan were grouped together in one cluster but accessions from India and Nepal were grouped together in another cluster. However, in both the species accessions from Turkey did not group together and fell in different clusters. CONCLUSIONS: Based on genetic similarity indices, higher diversity was observed in Trigonella caerulea as compared to Trigonella foenum-graecum. The genetic similarity matrices generated by ISSR and RAPD markers in both species were highly correlated (r = 0.78 at p = 0.001 for Trigonella foenum-graecum and r = 0.98 at p = 0.001 for Trigonella caerulea) indicating congruence between these two systems. Implications of these observations in the analysis of genetic diversity and in supporting the possible Center of Origin and/or Diversity for Trigonella are discussed. PMID- 15285784 TI - Maternal microchimerism in the livers of patients with biliary atresia. AB - BACKGROUND: Biliary atresia (BA) is a neonatal cholestatic disease of unknown etiology. It is the leading cause of liver transplantation in children. Many similarities exist between BA and graft versus host disease suggesting engraftment of maternal cells during gestation could result in immune responses that lead to BA. The aim of this study was to determine the presence and extent of maternal microchimerism (MM) in the livers of infants with BA. METHODS: Using fluorescent in situ hybridization (FISH), 11 male BA & 4 male neonatal hepatitis (NH) livers, which served as controls, were analyzed for X and Y-chromosomes. To further investigate MM in BA, 3 patients with BA, and their mothers, were HLA typed. Using immunohistochemical stains, the BA livers were examined for MM. Four additional BA livers underwent analysis by polymerase chain reaction (PCR) for evidence of MM. RESULTS: By FISH, 8 BA and 2 NH livers were interpretable. Seven of eight BA specimens showed evidence of MM. The number of maternal cells ranged from 2-4 maternal cells per biopsy slide. Neither NH specimen showed evidence of MM. In addition, immunohistochemical stains confirmed evidence of MM. Using PCR, a range of 1-142 copies of maternal DNA per 25,000 copies of patients DNA was found. CONCLUSIONS: Maternal microchimerism is present in the livers of patients with BA and may contribute to the pathogenesis of BA. PMID- 15285786 TI - Slowly developing depression of N-methyl-D-aspartate receptor mediated responses in young rat hippocampi. AB - BACKGROUND: Activation of N-methyl-D-aspartate (NMDA) type glutamate receptors is essential in triggering various forms of synaptic plasticity. A critical issue is to what extent such plasticity involves persistent changes of glutamate receptor subtypes and many prior studies have suggested a main role for alpha-amino-3 hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptors in mediating the effect. Our previous work in hippocampal slices revealed that, under pharmacological unblocking of NMDA receptors, both AMPA and NMDA receptor mediated responses undergo a slowly developing depression. In the present study we have further addressed this phenomenon, focusing on the contribution via NMDA receptors. Pharmacologically isolated NMDA receptor mediated excitatory postsynaptic potentials (EPSPs) were recorded for two independent synaptic pathways in CA1 area using perfusion with low Mg2+ (0.1 mM) to unblock NMDA receptors. RESULTS: Following unblocking of NMDA receptors, there was a gradual decline of NMDA receptor mediated EPSPs for 2-3 hours towards a stable level of ca. 60-70 % of the maximal size. If such an experimental session was repeated twice in the same pathway with a period of NMDA receptor blockade in between, the depression attained in the first session was still evident in the second one and no further decay occurred. The persistency of the depression was also validated by comparison between pathways. It was found that the responses of a control pathway, unstimulated in the first session of receptor unblocking, behaved as novel responses when tested in association with the depressed pathway under the second session. In similar experiments, but with AP5 present during the first session, there was no subsequent difference between NMDA EPSPs. CONCLUSIONS: Our findings show that merely evoking NMDA receptor mediated responses results in a depression which is input specific, induced via NMDA receptor activation, and is maintained for several hours through periods of receptor blockade. The similarity to key features of long-term depression and long-term potentiation suggests a possible relation to these phenomena. Additionally, a short term potentiation and decay (<5 min) were observed during sudden start of NMDA receptor activation supporting the idea that NMDA receptor mediated responses are highly plastic. PMID- 15285788 TI - Significant receptor affinities of metabolites and a degradation product of mometasone furoate. AB - Mometasone furoate (MF) is a highly potent glucocorticoid used topically to treat inflammation in the lung, nose and on the skin. However, so far no information has been published on the human glucocorticoid receptor activity of the metabolites or degradation products of MF. We have now determined the relative receptor binding affinities of the known metabolite 6beta-OH MF and the degradation product 9,11-epoxy MF to understand their possible contribution to undesirable systemic side effects. In competition experiments with human lung glucocorticoid receptors we have determined the relative receptor affinities (RRA) of these substances with reference to dexamethasone (RRA = 100). We have discovered that 6beta-OH MF and 9,11-epoxy MF display RRAs of 206 +/- 15 and 220 +/- 22, respectively. This level of activity is similar to that of the clinically used inhaled corticosteroid flunisolide (RRA 180 +/- 11). Furthermore we observed that 9,11-epoxy MF is a chemically reactive metabolite. In recovery experiments with human plasma and lung tissue we found a time dependent decrease in extractability of the compound. Hence, we provide data that might contribute to the understanding of the pharmacokinetics as well as the clinical effects of MF. PMID- 15285789 TI - Stem cells and repair of lung injuries. AB - Fueled by the promise of regenerative medicine, currently there is unprecedented interest in stem cells. Furthermore, there have been revolutionary, but somewhat controversial, advances in our understanding of stem cell biology. Stem cells likely play key roles in the repair of diverse lung injuries. However, due to very low rates of cellular proliferation in vivo in the normal steady state, cellular and architectural complexity of the respiratory tract, and the lack of an intensive research effort, lung stem cells remain poorly understood compared to those in other major organ systems. In the present review, we concisely explore the conceptual framework of stem cell biology and recent advances pertinent to the lungs. We illustrate lung diseases in which manipulation of stem cells may be physiologically significant and highlight the challenges facing stem cell-related therapy in the lung. PMID- 15285787 TI - C-type lectin-like domains in Fugu rubripes. AB - BACKGROUND: Members of the C-type lectin domain (CTLD) superfamily are metazoan proteins functionally important in glycoprotein metabolism, mechanisms of multicellular integration and immunity. Three genome-level studies on human, C. elegans and D. melanogaster reported previously demonstrated almost complete divergence among invertebrate and mammalian families of CTLD-containing proteins (CTLDcps). RESULTS: We have performed an analysis of CTLD family composition in Fugu rubripes using the draft genome sequence. The results show that all but two groups of CTLDcps identified in mammals are also found in fish, and that most of the groups have the same members as in mammals. We failed to detect representatives for CTLD groups V (NK cell receptors) and VII (lithostathine), while the DC-SIGN subgroup of group II is overrepresented in Fugu. Several new CTLD-containing genes, highly conserved between Fugu and human, were discovered using the Fugu genome sequence as a reference, including a CSPG family member and an SCP-domain-containing soluble protein. A distinct group of soluble dual-CTLD proteins has been identified, which may be the first reported CTLDcp group shared by invertebrates and vertebrates. We show that CTLDcp-encoding genes are selectively duplicated in Fugu, in a manner that suggests an ancient large-scale duplication event. We have verified 32 gene structures and predicted 63 new ones, and make our annotations available through a distributed annotation system (DAS) server http://anz.anu.edu.au:8080/Fugu_rubripes/ and their sequences as additional files with this paper. CONCLUSIONS: The vertebrate CTLDcp family was essentially formed early in vertebrate evolution and is completely different from the invertebrate families. Comparison of fish and mammalian genomes revealed three groups of CTLDcps and several new members of the known groups, which are highly conserved between fish and mammals, but were not identified in the study using only mammalian genomes. Despite limitations of the draft sequence, the Fugu rubripes genome is a powerful instrument for gene discovery and vertebrate evolutionary analysis. The composition of the CTLDcp superfamily in fish and mammals suggests that large-scale duplication events played an important role in the evolution of vertebrates. PMID- 15285790 TI - The expanding role of Tax in transcription. AB - The viral transactivator of HTLV-I, Tax, has long been shown to target the earliest steps of transcription by forming quaternary complexes with sequence specific transcription factors and histone-modifying enzymes in the LTR of HTLV I. However, a new study suggests that Tax preferentially transactivates the 21-bp repeats through CREB1 and not other bZIP proteins. The additional transactivation of Tax-responsive promoters subsequent to initiation is also presented. This result highlights a potentially novel role of Tax following TBP recruitment (i.e. initiation) and may expand the mechanism of Tax transactivation in promoter clearance and transcriptional elongation. PMID- 15285792 TI - Improved methods for the mathematically controlled comparison of biochemical systems. AB - The method of mathematically controlled comparison provides a structured approach for the comparison of alternative biochemical pathways with respect to selected functional effectiveness measures. Under this approach, alternative implementations of a biochemical pathway are modeled mathematically, forced to be equivalent through the application of selected constraints, and compared with respect to selected functional effectiveness measures. While the method has been applied successfully in a variety of studies, we offer recommendations for improvements to the method that (1) relax requirements for definition of constraints sufficient to remove all degrees of freedom in forming the equivalent alternative, (2) facilitate generalization of the results thus avoiding the need to condition those findings on the selected constraints, and (3) provide additional insights into the effect of selected constraints on the functional effectiveness measures. We present improvements to the method and related statistical models, apply the method to a previously conducted comparison of network regulation in the immune system, and compare our results to those previously reported. PMID- 15285791 TI - Specific TATAA and bZIP requirements suggest that HTLV-I Tax has transcriptional activity subsequent to the assembly of an initiation complex. AB - BACKGROUND: Human T-cell leukemia virus type I (HTLV-I) Tax protein is a transcriptional regulator of viral and cellular genes. In this study we have examined in detail the determinants for Tax-mediated transcriptional activation. RESULTS: Whereas previously the LTR enhancer elements were thought to be the sole Tax-targets, herein, we find that the core HTLV-I TATAA motif also provides specific responsiveness not seen with either the SV40 or the E1b TATAA boxes. When enhancer elements which can mediate Tax-responsiveness were compared, the authentic HTLV-I 21-bp repeats were found to be the most effective. Related bZIP factors such as CREB, ATF4, c-Jun and LZIP are often thought to recognize the 21 bp repeats equivalently. However, amongst bZIP factors, we found that CREB, by far, is preferred by Tax for activation. When LTR transcription was reconstituted by substituting either kappaB or serum response elements in place of the 21-bp repeats, Tax activated these surrogate motifs using surfaces which are different from that utilized for CREB interaction. Finally, we employed artificial recruitment of TATA-binding protein to the HTLV-I promoter in "bypass" experiments to show for the first time that Tax has transcriptional activity subsequent to the assembly of an initiation complex at the promoter. CONCLUSIONS: Optimal activation of the HTLV-I LTR by Tax specifically requires the core HTLV-I TATAA promoter, CREB and the 21-bp repeats. In addition, we also provide the first evidence for transcriptional activity of Tax after the recruitment of TATA binding protein to the promoter. PMID- 15285793 TI - Inhibition of microglial inflammatory responses by norepinephrine: effects on nitric oxide and interleukin-1beta production. AB - BACKGROUND: Under pathological conditions, microglia produce proinflammatory mediators which contribute to neurologic damage, and whose levels can be modulated by endogenous factors including neurotransmitters such as norepinephrine (NE). We investigated the ability of NE to suppress microglial activation, in particular its effects on induction and activity of the inducible form of nitric oxide synthase (NOS2) and the possible role that IL-1beta plays in that response. METHODS: Rat cortical microglia were stimulated with bacterial lipopolysaccharide (LPS) to induce NOS2 expression (assessed by nitrite and nitrate accumulation, NO production, and NOS2 mRNA levels) and IL-1beta release (assessed by ELISA). Effects of NE were examined by co-incubating cells with different concentrations of NE, adrenergic receptor agonists and antagonists, cAMP analogs, and protein kinase (PK) A and adenylate cyclase (AC) inhibitors. Effects on the NFkappaB:IkappaB pathway were examined by using selective a NFkappaB inhibitor and measuring IkappaBalpha protein levels by western blots. A role for IL-1beta in NOS2 induction was tested by examining effects of caspase-1 inhibitors and using caspase-1 deficient cells. RESULTS: LPS caused a time dependent increase in NOS2 mRNA levels and NO production; which was blocked by a selective NFkappaB inhibitor. NE dose-dependently reduced NOS2 expression and NO generation, via activation of beta2-adrenergic receptors (beta2-ARs), and reduced loss of inhibitory IkBalpha protein. NE effects were replicated by dibutyryl cyclic AMP. However, co-incubation with either PKA or AC inhibitors did not reverse suppressive effects of NE, but instead reduced nitrite production. A role for IL-1beta was suggested since NE potently blocked microglial IL-1beta production. However, incubation with a caspase-1 inhibitor, which reduced IL 1beta levels, had no effect on NO production; incubation with IL-receptor antagonist had biphasic effects on nitrite production; and NE inhibited nitrite production in caspase-1 deficient microglia. CONCLUSIONS: NE reduces microglial NOS2 expression and IL-1beta production, however IL-1beta does not play a critical role in NOS2 induction nor in mediating NE suppressive effects. Changes in magnitude or kinetics of cAMP may modulate NOS2 induction as well as suppression by NE. These results suggest that dysregulation of the central cathecolaminergic system may contribute to detrimental inflammatory responses and brain damage in neurological disease or trauma. PMID- 15285794 TI - Antioxidant protection from HIV-1 gp120-induced neuroglial toxicity. AB - BACKGROUND: The pathogenesis of HIV-1 glycoprotein 120 (gp120) associated neuroglial toxicity remains unresolved, but oxidative injury has been widely implicated as a contributing factor. In previous studies, exposure of primary human central nervous system tissue cultures to gp120 led to a simplification of neuronal dendritic elements as well as astrocytic hypertrophy and hyperplasia; neuropathological features of HIV-1-associated dementia. Gp120 and proinflammatory cytokines upregulate inducible nitric oxide synthase (iNOS), an important source of nitric oxide (NO) and nitrosative stress. Because ascorbate scavenges reactive nitrogen and oxygen species, we studied the effect of ascorbate supplementation on iNOS expression as well as the neuronal and glial structural changes associated with gp120 exposure. METHODS: Human CNS cultures were derived from 16-18 week gestation post-mortem fetal brain. Cultures were incubated with 400 microM ascorbate-2-O-phosphate (Asc-p) or vehicle for 18 hours then exposed to 1 nM gp120 for 24 hours. The expression of iNOS and neuronal (MAP2) and astrocytic (GFAP) structural proteins was examined by immunohistochemistry and immunofluorescence using confocal scanning laser microscopy (CSLM). RESULTS: Following gp120 exposure iNOS was markedly upregulated from undetectable levels at baseline. Double label CSLM studies revealed astrocytes to be the prime source of iNOS with rare neurons expressing iNOS. This upregulation was attenuated by the preincubation with Asc-p, which raised the intracellular concentration of ascorbate. Astrocytic hypertrophy and neuronal injury caused by gp120 were also prevented by preincubation with ascorbate. CONCLUSIONS: Ascorbate supplementation prevents the deleterious upregulation of iNOS and associated neuronal and astrocytic protein expression and structural changes caused by gp120 in human brain cell cultures. PMID- 15285795 TI - Human CNS cultures exposed to HIV-1 gp120 reproduce dendritic injuries of HIV-1 associated dementia. AB - HIV-1-associated dementia remains a common subacute to chronic central nervous system degeneration in adult and pediatric HIV-1 infected populations. A number of viral and host factors have been implicated including the HIV-1 120 kDa envelope glycoprotein (gp120). In human post-mortem studies using confocal scanning laser microscopy for microtubule-associated protein 2 and synaptophysin, neuronal dendritic pathology correlated with dementia. In the present study, primary human CNS cultures exposed to HIV-1 gp120 at 4 weeks in vitro suffered gliosis and dendritic damage analogous to that described in association with HIV 1-associated dementia. PMID- 15285796 TI - Selective COX-2 inhibition prevents progressive dopamine neuron degeneration in a rat model of Parkinson's disease. AB - Several lines of evidence point to a significant role of neuroinflammation in Parkinson's disease (PD) and other neurodegenerative disorders. In the present study we examined the protective effect of celecoxib, a selective inhibitor of the inducible form of cyclooxygenase (COX-2), on dopamine (DA) cell loss in a rat model of PD. We used the intrastriatal administration of 6-hydroxydopamine (6 OHDA) that induces a retrograde neuronal damage and death, which progresses over weeks. Animals were randomized to receive celecoxib (20 mg/kg/day) or vehicle starting 1 hour before the intrastriatal administration of 6-OHDA. Evaluation was performed in vivo using micro PET and selective radiotracers for DA terminals and microglia. Post mortem analysis included stereological quantification of tyrosine hydroxylase, astrocytes and microglia. 12 days after the 6-OHDA lesion there were no differences in DA cell or fiber loss between groups, although the microglial cell density and activation was markedly reduced in animals receiving celecoxib (p < 0.01). COX-2 inhibition did not reduce the typical astroglial response in the striatum at any stage. Between 12 and 21 days, there was a significant progression of DA cell loss in the vehicle group (from 40 to 65%) that was prevented by celecoxib. Therefore, inhibition of COX-2 by celecoxib appears to be able, either directly or through inhibition of microglia activation to prevent or slow down DA cell degeneration. PMID- 15285797 TI - PPARgamma, neuroinflammation, and disease. AB - BACKGROUND: Peroxisome proliferator-activated receptors (PPARs) are a class of nuclear transcription factors that are activated by fatty acids and their derivatives. One of these, PPARgamma, regulates responsiveness to insulin in adipose cells, and PPARgamma-activating drugs such as pioglitazone are used in the treatment of type 2 diabetes. PPARgamma acts in myeloid-lineage cells, including T-cells and macrophages, to suppress their activation and their elaboration of inflammatory molecules. PPARgamma activation also suppresses the activated phenotype in microglia, suggesting that PPARgamma-activating drugs may be of benefit in chronic neuroinflammatory diseases. Some, but not all, nonsteroidal anti-inflammatory agents (indomethacin and ibuprofen in particular) also have activating effects on PPARgamma. DISCUSSION AND CONCLUSIONS: These observations suggest on the one hand a role for PPARgamma-activating drugs in the treatment of chronic neuroinflammatory diseases-as shown for a patient with secondary progressive multiple sclerosis by Pershadsingh et al. in this issue of the Journal of Neuroinflammation-and suggest on the other hand a possible explanation for confusing and contradictory results in trials of nonsteroidal anti-inflammatory agents in Alzheimer's disease. PMID- 15285798 TI - Ginkgolide B inhibits the neurotoxicity of prions or amyloid-beta1-42. AB - BACKGROUND: Neuronal loss in Alzheimer's or prion diseases is preceded by the accumulation of fibrillar aggregates of toxic proteins (amyloid-beta1-42 or the prion protein). Since some epidemiological studies have demonstrated that the EGb 761 extract, from the leaves of the Ginkgo biloba tree, has a beneficial effect on Alzheimer's disease, the effect of some of the major components of the EGb 761 extract on neuronal responses to amyloid-beta1-42, or to a synthetic miniprion (sPrP106), were investigated. METHODS: Components of the EGb 761 extract were tested in 2 models of neurodegeneration. SH-SY5Y neuroblastoma cells were pre treated with ginkgolides A or B, quercetin or myricetin, and incubated with amyloid-beta1-42, sPrP106, or other neurotoxins. After 24 hours neuronal survival and the production of prostaglandin E2 that is closely associated with neuronal death was measured. In primary cortical neurons apoptosis (caspase-3) in response to amyloid-beta1-42 or sPrP106 was measured, and in co-cultures the effects of the ginkgolides on the killing of amyloid-beta1-42 or sPrP106 damaged neurons by microglia was tested. RESULTS: Neurons treated with ginkgolides A or B were resistant to amyloid-beta1-42 or sPrP106. Ginkgolide-treated cells were also resistant to platelet activating factor or arachidonic acid, but remained susceptible to hydrogen peroxide or staurosporine. The ginkgolides reduced the production of prostaglandin E2 in response to amyloid-beta1-42 or sPrP106. In primary cortical neurons, the ginkgolides reduced caspase-3 responses to amyloid beta1-42 or sPrP106, and in co-culture studies the ginkgolides reduced the killing of amyloid-beta1-42 or sPrP106 damaged neurons by microglia. CONCLUSION: Nanomolar concentrations of the ginkgolides protect neurons against the otherwise toxic effects of amyloid-beta1-42 or sPrP106. The ginkgolides also prevented the neurotoxicity of platelet activating factor and reduced the production of prostaglandin E2 in response to platelet activating factor, amyloid-beta1-42 or sPrP106. These results are compatible with prior reports that ginkgolides inhibit platelet-activating factor, and that platelet-activating factor antagonists block the toxicity of amyloid-beta1-42 or sPrP106. The results presented here suggest that platelet-activating factor antagonists such as the ginkgolides may be relevant treatments for prion or Alzheimer's diseases. PMID- 15285799 TI - Effect of pioglitazone treatment in a patient with secondary multiple sclerosis. AB - BACKGROUND: Ligands of the peroxisome proliferator-activated receptor-gamma (PPARgamma) induce apoptosis in activated T-lymphocytes and exert anti inflammatory effects in glial cells. Preclinical studies have shown that the thiazolidinedione pioglitazone, an FDA-approved PPARgamma agonist used to treat type 2 diabetes, delays the onset and reduces the severity of clinical symptoms in experimental autoimmune encephalomyelitis, an animal model of multiple sclerosis (MS). We therefore tested the safety and therapeutic potential of oral pioglitazone in a patient with secondary MS. CASE PRESENTATION: The rationale and risks of taking pioglitazone were carefully explained to the patient, consent was obtained, and treatment was initiated at 15 mg per day p.o. and then increased by 15 mg biweekly to 45 mg per day p.o. for the duration of the treatment. Safety was assessed by measurements of metabolic profiles, blood pressure, and edema; effects on clinical symptoms were assessed by measurement of cognition, motor function and strength, and MRI. Within 4 weeks the patient exhibited increased appetite, cognition and attention span. After 12 months treatment, body weight increased from 27.3 to 35.9 kg (32%) and maintained throughout the duration of the study. Upper extremity strength and coordination improved, and increased fine coordination was noted unilaterally after 8 months and bilaterally after 15 months. After 8 months therapy, the patient demonstrated improvement in orientation, short-term memory, and attention span. MRIs carried out after 10 and 18 months of treatment showed no perceptible change in overall brain atrophy, extent of demyelination, or in Gd-enhancement. After 3.0 years on pioglitazone, the patient continues to be clinically stable, with no adverse events. CONCLUSIONS: In a patient with secondary progressive MS, daily treatment with 45 mg p.o. pioglitazone for 3 years induced apparent clinical improvement without adverse events. Pioglitazone should therefore be considered for further testing of therapeutic potential in MS patients. PMID- 15285800 TI - Induction of serine racemase expression and D-serine release from microglia by amyloid beta-peptide. AB - BACKGROUND: Roles for excitotoxicity and inflammation in Alzheimer's disease have been hypothesized. Proinflammatory stimuli, including amyloid beta-peptide (Abeta), elicit a release of glutamate from microglia. We tested the possibility that a coagonist at the NMDA class of glutamate receptors, D-serine, could respond similarly. METHODS: Cultured microglial cells were exposed to Abeta. The culture medium was assayed for levels of D-serine by HPLC and for effects on calcium and survival on primary cultures of rat hippocampal neurons. Microglial cell lysates were examined for the levels of mRNA and protein for serine racemase, the enzyme that forms D-serine from L-serine. The racemase mRNA was also assayed in Alzheimer hippocampus and age-matched controls. A microglial cell line was transfected with a luciferase reporter construct driven by the putative regulatory region of human serine racemase. RESULTS: Conditioned medium from Abeta-treated microglia contained elevated levels of D-serine. Bioassays of hippocampal neurons with the microglia-conditioned medium indicated that Abeta elevated a NMDA receptor agonist that was sensitive to an antagonist of the D serine/glycine site (5,7-dicholorokynurenic acid; DCKA) and to enzymatic degradation of D-amino acids by D-amino acid oxidase (DAAOx). In the microglia, Abeta elevated steady-state levels of dimeric serine racemase, the apparent active form of the enzyme. Promoter-reporter and mRNA analyses suggest that serine racemase is transcriptionally induced by Abeta. Finally, the levels of serine racemase mRNA were elevated in Alzheimer's disease hippocampus, relative to age-matched controls. CONCLUSIONS: These data suggest that Abeta could contribute to neurodegeneration through stimulating microglia to release cooperative excitatory amino acids, including D-serine. PMID- 15285801 TI - Microglia and neuroinflammation: a pathological perspective. AB - Microglia make up the innate immune system of the central nervous system and are key cellular mediators of neuroinflammatory processes. Their role in central nervous system diseases, including infections, is discussed in terms of a participation in both acute and chronic neuroinflammatory responses. Specific reference is made also to their involvement in Alzheimer's disease where microglial cell activation is thought to be critically important in the neurodegenerative process. PMID- 15285802 TI - Tumor necrosis factor-mediated inhibition of interleukin-18 in the brain: a clinical and experimental study in head-injured patients and in a murine model of closed head injury. AB - Tumor necrosis factor (TNF) and interleukin-(IL)-18 are important mediators of neuroinflammation after closed head injury (CHI). Both mediators have been previously found to be significantly elevated in the intracranial compartment after brain injury, both in patients as well as in experimental model systems. However, the interrelation and regulation of these crucial cytokines within the injured brain has not yet been investigated. The present study was designed to assess a potential regulation of intracranial IL-18 levels by TNF based on a clinical study in head-injured patients and an experimental model in mice. In the first part, we investigated the interrelationship between the daily TNF and IL-18 cerebrospinal fluid levels in 10 patients with severe CHI for up to 14 days after trauma. In the second part of the study, the potential TNF-dependent regulation of intracerebral IL-18 levels was further characterized in an experimental set-up in mice: (1) in a standardized model of CHI in TNF/lymphotoxin-alpha gene deficient mice and wild-type (WT) littermates, and (2) by intracerebro ventricular injection of mouse recombinant TNF in WT C57BL/6 mice. The results demonstrate an inverse correlation of intrathecal TNF and IL-18 levels in head injured patients and a TNF-dependent inhibition of IL-18 after intracerebral injection in mice. These findings imply a potential new anti-inflammatory mechanism of TNF by attenuation of IL-18, thus confirming the proposed "dual" function of this cytokine in the pathophysiology of traumatic brain injury. PMID- 15285803 TI - Chronic brain inflammation leads to a decline in hippocampal NMDA-R1 receptors. AB - BACKGROUND: Neuroinflammation plays a prominent role in the progression of Alzheimer's disease and may be responsible for degeneration in vulnerable regions such as the hippocampus. Neuroinflammation is associated with elevated levels of extracellular glutamate and potentially an enhanced stimulation of glutamate N methyl-D-aspartate receptors. This suggests that neurons that express these glutamate receptors might be at increased risk of degeneration in the presence of chronic neuroinflammation. METHODS: We have characterized a novel model of chronic brain inflammation using a slow infusion of lipopolysaccharide into the 4th ventricle of rats. This model reproduces many of the behavioral, electrophysiological, neurochemical and neuropathological changes associated with Alzheimer's disease. RESULTS: The current study demonstrated that chronic neuroinflammation is associated with the loss of N-methyl-D-aspartate receptors, as determined both qualitatively by immunohistochemistry and quantitatively by in vitro binding studies using [3H]MK-801, within the hippocampus and entorhinal cortex. CONCLUSION: The gradual loss of function of this critical receptor within the temporal lobe region may contribute to some of the cognitive deficits observed in patients with Alzheimer's disease. PMID- 15285804 TI - Effect of anti-inflammatory agents on transforming growth factor beta over expressing mouse brains: a model revised. AB - BACKGROUND: The over-expression of transforming growth factor beta-1(TGF-beta1) has been reported to cause hydrocephalus, glia activation, and vascular amyloidbeta (Abeta) deposition in mouse brains. Since these phenomena partially mimic the cerebral amyloid angiopathy (CAA) concomitant to Alzheimer's disease, the findings in TGF-beta1 over-expressing mice prompted the hypothesis that CAA could be caused or enhanced by the abnormal production of TGF-beta1. This idea was in accordance with the view that chronic inflammation contributes to Alzheimer's disease, and drew attention to the therapeutic potential of anti inflammatory drugs for the treatment of Abeta-elicited CAA. We thus studied the effect of anti-inflammatory drug administration in TGF-beta1-induced pathology. METHODS: Two-month-old TGF-beta1 mice and littermate controls were orally administered pioglitazone, a peroxisome proliferator-activated receptor-gamma agonist, or ibuprofen, a non steroidal anti-inflammatory agent, for two months. Glia activation was assessed by immunohistochemistry and western blot analysis; Abeta precursor protein (APP) by western blot analysis; Abeta deposition by immunohistochemistry, thioflavin-S staining and ELISA; and hydrocephalus by measurements of ventricle size on autoradiographies of brain sections. Results are expressed as means +/- SD. Data comparisons were carried with the Student's T test when two groups were compared, or ANOVA analysis when more than three groups were analyzed. RESULTS: Animals displayed glia activation, hydrocephalus and a robust thioflavin-S-positive vascular deposition. Unexpectedly, these deposits contained no Abeta or serum amyloid P component, a common constituent of amyloid deposits. The thioflavin-S-positive material thus remains to be identified. Pioglitazone decreased glia activation and basal levels of Abeta42- with no change in APP contents - while it increased hydrocephalus, and had no effect on the thioflavin-S deposits. Ibuprofen mimicked the reduction of glia activation caused by pioglitazone and the lack of effect on the thioflavin-S-labeled deposits. CONCLUSIONS: i) TGF-beta1 over-expressing mice may not be an appropriate model of Abeta-elicited CAA; and ii) pioglitazone has paradoxical effects on TGF-beta1-induced pathology suggesting that anti-inflammatory therapy may reduce the damage resulting from active glia, but not from vascular alterations or hydrocephalus. Identification of the thioflavin-S-positive material will facilitate the full appraisal of the clinical implication of the effects of anti-inflammatory drugs, and provide a more thorough understanding of TGF-beta1 actions in brain. PMID- 15285806 TI - Welcome to the Journal of Neuroinflammation! AB - Welcome to the Journal of Neuroinflammation, an open-access, peer-reviewed, online journal that focuses on innate immunological responses of the central nervous system, involving microglia, astrocytes, cytokines, chemokines, and related molecular processes. 'Neuroinflammation' is an encapsulization of the idea that microglial and astrocytic responses and actions in the central nervous system have a fundamentally inflammation-like character, and that these responses are central to the pathogenesis and progression of a wide variety of neurological disorders. This concept has its roots in the discoveries of inflammatory cytokines and proteins in the plaques of Alzheimer disease, and these ideas have been extended to other neurodegenerative diseases, to ischemic/toxic diseases, to tumor biology and even to normal brain development. The Journal of Neuroinflammation, published by BioMed Central, will bring together work focusing on microglia, astrocytes, cytokines, chemokines, and related molecular processes in the central nervous system. All articles published in the Journal of Neuroinflammation will be immediately listed in PubMed, and access to published articles will be universal and free through the internet. PMID- 15285807 TI - Evaluation of in vivo labelled dendritic cell migration in cancer patients. AB - BACKGROUND: Dendritic Cell (DC) vaccination is a very promising therapeutic strategy in cancer patients. The immunizing ability of DC is critically influenced by their migration activity to lymphatic tissues, where they have the task of priming naive T-cells. In the present study in vivo DC migration was investigated within the context of a clinical trial of antitumor vaccination. In particular, we compared the migration activity of mature Dendritic Cells (mDC) with that of immature Dendritic Cells (iDC) and also assessed intradermal versus subcutaneous administration. METHODS: DC were labelled with 99mTc-HMPAO or 111In Oxine, and the presence of labelled DC in regional lymph nodes was evaluated at pre-set times up to a maximum of 72 h after inoculation. Determinations were carried out in 8 patients (7 melanoma and 1 renal cell carcinoma). RESULTS: It was verified that intradermal administration resulted in about a threefold higher migration to lymph nodes than subcutaneous administration, while mDC showed, on average, a six-to eightfold higher migration than iDC. The first DC were detected in lymph nodes 20-60 min after inoculation and the maximum concentration was reached after 48-72 h. CONCLUSIONS: These data obtained in vivo provide preliminary basic information on DC with respect to their antitumor immunization activity. Further research is needed to optimize the therapeutic potential of vaccination with DC. PMID- 15285805 TI - Modelling neuroinflammatory phenotypes in vivo. AB - Inflammation of the central nervous system is an important but poorly understood part of neurological disease. After acute brain injury or infection there is a complex inflammatory response that involves activation of microglia and astrocytes and increased production of cytokines, chemokines, acute phase proteins, and complement factors. Antibodies and T lymphocytes may be involved in the response as well. In neurodegenerative disease, where injury is more subtle but consistent, the inflammatory response is continuous. The purpose of this prolonged response is unclear, but it is likely that some of its components are beneficial and others are harmful. Animal models of neurological disease can be used to dissect the specific role of individual mediators of the inflammatory response and assess their potential benefit. To illustrate this approach, we discuss how mutant mice expressing different levels of the cytokine transforming growth factor beta-1 (TGF-beta1), a major modulator of inflammation, produce important neuroinflammatory phenotypes. We then demonstrate how crosses of TGF beta1 mutant mice with mouse models of Alzheimer's disease (AD) produced important new information on the role of inflammation in AD and on the expression of different neuropathological phenotypes that characterize this disease. PMID- 15285808 TI - Recurrence of granulosa cell tumour after thirty years with small bowel obstruction. AB - Granulosa cell tumours of the ovary are rare, comprising around 3% of ovarian tumours. These tumours have preponderance for local spread and extremely late recurrence. Although previous cases of recurrence have been described, it is extremely unusual for these tumours to recur after thirty years. We describe a case of recurrence of granulosa cell tumour after 30 years, presenting as small bowel obstruction. The patient had not been followed up after the original surgery, and on histological analysis, recurrence of the original tumour was confirmed. This case report emphasises the necessity for lifelong follow-up of patients who have had these tumours excised, and also the unusual way in which these tumours can recur. PMID- 15285809 TI - Capsular contraction following immediate reconstructive surgery for breast cancer - An association with methylene blue dye. AB - Capsular contraction following implantation of breast prostheses occurs in 2-33% of patients undergoing breast augmentation. This condition can be debilitating for patients, and often requires revisional surgery. The aetiology of capsular contraction is unclear, but may be due to infection, haematoma or foreign body type reactions.Methylene blue dye is a substance known to cause localised tissue inflammation, and is often used during breast cancer surgery to allow identification of the sentinel lymph node. We report a case of Baker Grade 4 capsular contraction necessitating revisional surgery, occurring in a patient who underwent immediate breast reconstruction during surgery for breast cancer. Methylene blue dye was used to locate the sentinel nodes during the original surgery, and was found to have heavily discoloured the prosthesis at subsequent revisional surgery. Capsular contraction may have been caused in part by a localised tissue reaction initiated by, or involving the dye. PMID- 15285810 TI - There is no correlation between c-Myc mRNA expression and telomerase activity in human breast cancer. AB - BACKGROUND: Telomerase is a ribonucleoprotein enzyme that synthesises telomeres after cell division and maintains chromosomal length and stability thus leading to cellular immortalisation. The hTERT (human telomerase reverse transcriptase) subunit seems to be the rate-limiting determinant of telomerase and knowledge of factors controlling hTERT transcription may be useful in therapeutic strategies. The hTERT promoter contains binding sites for c-Myc and there is experimental and in vitro evidence that c-Myc may increase hTERT expression. MATERIALS AND METHODS: RNA was extracted from 18 breast carcinomas and c-Myc mRNA expression was estimated by quantitative reverse transcriptase-PCR (RT-PCR) with Taqman methodology. These tumours had already been analysed for ER and PgR status using ligand-binding assays and had had their DNA ploidy and S-phase fractions measured by flow cytometry. Telomerase activity had already been determined by using a modified telomeric repeat and amplification protocol (TRAP) assay. RESULTS: Telomerase activity ranged from 0 to 246 units of Total Protein Generated (TPG), where one unit of TPG was equal to 600 molecules of telomerase substrate primers extended by at least three telomeric repeats. Median levels of TPG were 60 and mean levels 81. There was no significant correlation between levels of c-Myc mRNA expression, telomerase activity, S phase fraction or PgR. There was a significant negative correlation with ER status. CONCLUSION: Although the hTERT promoter contains potential binding sites for c-Myc oncoprotein, we have found no correlation between c-Myc mRNA levels and telomerase activity. PMID- 15285811 TI - Welcome To International Seminars In Surgical Oncology. AB - This editorial marks the launch of a new online journal for surgical and medical oncology. The internet has produced a profound shift in the way in which clinicians, researchers and patients seek, interpret and utilise medical information and research. The launch of our journal comes at a time when these changes are in a rapid phase of development and consolidation. PMID- 15285812 TI - Sad, blue, or depressed days, health behaviors and health-related quality of life, Behavioral Risk Factor Surveillance System, 1995-2000. AB - BACKGROUND: Mood disorders are a major public health problem in the United States as well as globally. Less information exists however, about the health burden resulting from subsyndromal levels of depressive symptomatology, such as feeling sad, blue or depressed, among the general U.S. population. METHODS: As part of an optional Quality of Life survey module added to the U.S. Behavioral Risk Factor Surveillance System, between 1995-2000 a total of 166,564 BRFSS respondents answered the question, "During the past 30 days, for about how many days have you felt sad, blue, or depressed?" Means and 95% confidence intervals for sad, blue, depressed days (SBDD) and other health-related quality of life (HRQOL) measures were calculated using SUDAAN to account for the BRFSS's complex sample survey design. RESULTS: Respondents reported a mean of 3.0 (95% CI = 2.9-3.1) SBDD in the previous 30 days. Women (M = 3.5, 95% CI = 3.4-3.6) reported a higher number of SBDD than did men (M = 2.4, 95% CI = 2.2-2.5). Young adults aged 18-24 years reported the highest number of SBDD, whereas older adults aged 60-84 reported the fewest number. The gap in mean SBDD between men and women decreased with increasing age. SBDD was associated with an increased prevalence of behaviors risky to health, extremes of body mass index, less access to health care, and worse self-rated health status. Mean SBDD increased with progressively higher levels of physically unhealthy days, mentally unhealthy days, unhealthy days, activity limitation days, anxiety days, pain days, and sleepless days. CONCLUSION: Use of this measure of sad, blue or depressed days along with other valid mental health measures and community indicators can help to assess the burden of mental distress among the U.S. population, identify subgroups with unmet mental health needs, inform the development of targeted interventions, and monitor changes in population levels of mental distress over time. PMID- 15285814 TI - Efficacy of adjuvant capecitabine compared with bolus 5-fluorouracil/leucovorin regimen in dukes C colon cancer: results from the X-ACT trial. PMID- 15285815 TI - Bevacizumab improves the efficacy of 5-fluorouracil/leucovorin in patients with advanced colorectal cancer. PMID- 15285816 TI - Radiofrequency ablation of primary and metastatic hepatic malignancies. AB - Radiofrequency interstitial tissue ablation is a local ablative therapy in which tumors are destroyed in situ by thermal coagulation and protein denaturation through frictional heating produced by tissue ionic agitation from high-frequency alternating current. This technology can be used to destroy primary and metastatic hepatic lesions generally considered nonresectable or nonoperable, thus providing patients with these tumors, who have few treatment options, a relatively safe and effective alternative with the potential for improved chance of survival. Knowledge of the broad spectrum of potential complications associated with radiofrequency ablation (RFA) is essential for prevention, early detection, and proper management. Combining RFA with other modalities such as surgical resection or hepatic artery infusional chemotherapy is feasible, has increased the pool of operable patients, and may improve treatment efficacy and clinical outcome in properly selected patients. The approach to perform RFA percutaneously, laparoscopically, or during laparatomy should take into consideration tumor characteristics, imaging and technical limitations, and the role of other treatment modalities. Therefore, patients considered for RFA should be evaluated within the context of a multidisciplinary approach to insure proper patient selection and coordination of adjunct therapy. PMID- 15285817 TI - Retrospective study of resection of pulmonary metastases in patients with advanced colorectal cancer: the development of a preoperative chemotherapy strategy. AB - Considerable data are available to support the resection of hepatic metastases in patients with colorectal cancer, but there are relatively few studies on the role of pulmonary metastectomy. The small number of such studies is mainly noncontemporaneous and predates the use of preoperative neoadjuvant chemotherapy. A retrospective analysis of 31 patients with pulmonary metastases from colorectal cancer treated with surgery and perioperative chemotherapy between 1995 and 2003 was performed. Twenty patients (65%) proceeded directly to surgery and 5 of these received postoperative chemotherapy. Eleven patients (35%) received preoperative chemotherapy, which consisted of a fluoropyrimidine in combination with oxaliplatin or mitomycin-C, except for 1 patient who received single agent irinotecan. Nine of 11 patients (82%) had a partial response and 2 patients (18%) had stable disease. A total of 39 thoracic surgeries (6 bilateral and 1 incomplete) were performed. There were no postoperative deaths. Four of 20 patients (20%) who had initial surgery had postoperative complications, compared with 18% of the preoperative chemotherapy group. Overall 3- and 5-year survival rates after the first thoracic surgery were 65.2% (95% CI, 35.1%-83.9%) and 26.1% (95% CI, 4.3%-56.2%), respectively. Based on the limited data from this study, disease-free interval, number of pulmonary metastases, previous resection of hepatic metastases, prethoracotomy carcinoembryonic antigen levels, and preoperative chemotherapy were not found to be significant prognostic factors for survival. Therefore, surgical resection of lung metastases is associated with low morbidity and mortality and results in long-term survival for 20%-30% of patients. Moreover, preoperative chemotherapy produced a high response rate, with no patients experiencing disease progression before surgery. PMID- 15285818 TI - Development of rationally designed, target-based agents for the treatment of advanced colorectal cancer. AB - Although there have been several recent additions to the conventional armamentarium used to treat patients with advanced colorectal cancer, principally as a result of the development of selective and nonselective pharmacologic agents and antibodies, the general outcome of patients with advanced disease is still poor. However, a greater understanding of cancer biology, as well as major advances in biotechnology, is beginning to identify and characterize molecular aberrations that are common in patients with colorectal cancer. These advances have resulted in the development of a wide range of rationally designed, target based anticancer therapeutic agents, which, by virtue of their selectivity, would be expected to produce less nonspecific toxicity and therefore higher therapeutic indices compared with nonspecific cytotoxic agents. This review will discuss several novel targets and therapeutic agents, particularly those designed to interrupt aberrant signal transduction and apoptotic processes. It will also emphasize the complexity of these systems and the need to incorporate novel clinical development paradigms based on a thorough scientific understanding of these targets. PMID- 15285819 TI - Endorectal ultrasound in the preoperative evaluation of rectal cancer. AB - For rectal cancer, the decisions about neoadjuvant therapy, radical resection, or local excision depend on accurate preoperative staging. Multiple modalities are available to stage rectal cancer, including digital rectal examination, computed tomography, magnetic resonance imaging (MRI), and endorectal ultrasound (ERUS). Digital rectal examination accuracy varies from 58% to 88% for depth of penetration. Computed tomography accuracy varies from 53% to 94% for depth of penetration and from 54% to 70% for lymph node metastases. Magnetic resonance imaging accuracy varies from 66% to 92% for depth of penetration and from 60% to 90% for lymph node metastases. Endorectal ultrasound varies from 62% to 92% for depth of penetration and from 64% to 88% for lymph node metastases. In all radiologic modalities, overstaging and understaging occurs. Endorectal ultrasound has the advantage of being portable and often office-based, requiring only minimal preparation and is well tolerated by the patient. Although MRI with the use of an endorectal coil may have a slightly higher accuracy for detecting lymph nodes, ERUS has been shown to be the most accurate method for the determination of the depth of wall penetration, and is comparable for lymph node metastases. Interpretation varies with operator experience. Three-dimensional (3D) ERUS may further improve staging accuracy. Endorectal ultrasound is an accurate method to preoperatively stage rectal cancers. Although operator-dependent, it can be readily performed at the time of patient evaluation with minimal preparation or patient discomfort. We are prospectively evaluating modifications to the current staging system and the use of 3D ERUS. PMID- 15285824 TI - A survey of staff attitudes and comparative managerial and non-managerial views in a clinical directorate. AB - Widespread implementation of clinical directorates (CDs) has displaced traditional structures of hospitals over the past 20 years. Responses range from support for involving clinicians in organizational processes along with the associated managerial benefits, to criticism that foreshadows potential negative effects and warns that CDs will not of themselves resolve embedded health sector problems. There is limited empirical evidence about the transition and the views of staff toward CDs. To investigate staff attitudes, a questionnaire was developed and administered in a survey of 107 staff in a large hospital that had introduced CDs three years previously. Attitudes were assessed in terms of their intensity, polarity, uncertainty and positivism toward CDs. Managers and other staff held similar attitudes on 66% of questionnaire items. Significant differences were found in the remaining one-third of items. Managers were positive about CDs, whereas non-managers' approval was limited and muted. Managers' attitudes were more intense, less uncertain and less polarized than were non-managers'. They differed primarily in the areas of working relations and power. Over recent years, CDs seem to have become institutionalized and investigations into their operation have declined. Our results suggest that taking the benefits of CDs for granted is premature. PMID- 15285825 TI - Doctors as managers: investors and reluctants in a dual role. AB - Government reform of the NHS in the UK has sought to increase the involvement of doctors (clinicians) in hospital management. Using frameworks from the psychological contract and organisational misbehaviour literatures, this paper examines the processes involved when clinicians assume management roles. This literature seeks to explain breaches to expectations regarding prior agreements with management and subsequent actions of 'getting even' as a result of breaches to the employment relationship. A qualitative methodology using interviews was undertaken, which identified two distinct groups of clinician-manager. Investors actively pursued a management opportunity as an alternative to clinical medicine, whilst reluctants tended to assume a management role to protect particular specialities from outside influence or from those they thought would be inappropriate clinician-managers. Investors and reluctants often had very little prior experience of management and managers and had problems reconciling their dual clinician-management role. Poor relationships with hospital managers who often had no understanding of their dual responsibilities led to tensions and conflict, which questions continued developments in this important area of UK health policy. Suggestions for improving this process are outlined. PMID- 15285826 TI - Change management of mergers: the impact on NHS staff and their psychological contracts. AB - The NHS has experienced a significant amount of organisational change and restructuring, which has included numerous mergers and de-mergers, since the Labour party came to power in the UK in 1997. However, to date there has been little in the way of evaluation of such changes, particularly the impact of organisational restructuring on the staff involved. This paper examines the human aspect of a merger, and subsequent de-merger, within a primary care trust (PCT) in the North of England, using a focus group methodology. The findings demonstrate that leadership and management styles have a significant impact on staff experiencing such changes. In addition, the psychological contract can be damaged due to the impact of several factors, inducing exit or intention to leave. Employees experienced a constant cycle of change with little time for stabilisation or adjustment, leading to negativity and lowered motivation at times. PMID- 15285827 TI - Healthcare and the information revolution: re-configuring the healthcare service encounter. AB - Utilisation of internet-derived information by patients within the healthcare service encounter is increasing. Encompassing both the use of unidirectional information sites as well as bidirectional computer-mediated communities, this is manifest in the growth of consumerist expectations on the part of patients. Based on interviews with patients, professionals and internet site managers, this paper examines the role of the internet as a source of patient information and support, and in particular the effect on the relationship between 'informed' consumers and professionals involved in the delivery of healthcare services. The core challenge for informed consumers is to develop frameworks that facilitate robust dialogue, exchange of information and emotional support to complement their rising authority. The parallel challenge is for the established medical profession to recognize the consequences of this evolving dialogue and develop approaches to service delivery that effectively engage with consumers on the basis of this increasing authority. PMID- 15285828 TI - Using routinely recorded ethnicity: analysis of waiting times for elective admissions by ethnic group. AB - OBJECTIVE: To assess whether patients from non-white ethnic groups wait longer than white patients for elective in-patient admissions at St Mary's Hospital in London. METHODS: Patients who came off the waiting list for an elective inpatient admission between 1 April 2000 and 31 March 2001 were selected. A multivariable log linear model was developed to assess geometric mean waiting times for Black, Asian, Other and Missing ethnic groups compared to the White group, adjusted for age, sex, urgency and distance. RESULTS: Caution is needed in interpreting results, as a large number of patients had no usable ethnic code. There was no strong evidence that waiting times for ethnic groups were systematically different than for the White group. However, there was some evidence that white patients waited longer for a coronary arteriography than patients in other ethnic groups. This was partially explained by age, sex, clinical urgency and residential distance from St Mary's. CONCLUSIONS: The large proportion of patients with no usable ethnic code, lack of robust methods for case-mix adjustment and multiple ethnic categories makes analysis methodologically difficult. Regular and informative analysis of ethnic coded data is a necessary step in improving the accuracy and completeness of coding. PMID- 15285829 TI - Moderation of cognition-intention and cognition-behaviour relations: a meta analysis of properties of variables from the theory of planned behaviour. AB - Meta-analysis was used to quantify the moderating effects of seven properties of cognitions-accessibility, temporal stability, direct experience, involvement, certainty, ambivalence and affective-cognitive consistency-on cognition-intention and cognition-behaviour relations. Literature searches revealed 44 studies that could be included in the review. Findings showed that all of the properties, except involvement, moderated attitude-behaviour consistency. Similarly, all relevant moderators improved the consistency between intentions and behaviour. Temporal stability moderated PBC-behaviour relations, certainty moderated subjective norm-intention relations, and ambivalence, certainty, and involvement all moderated attitude-intention relations. Overall, temporal stability appeared to be the strongest moderator of cognition-behaviour relations. PMID- 15285830 TI - Citizenship in practice. AB - The idea of citizenship dates back to classical antiquity. It was originally concerned to address legitimacy of occupancy in the public sphere. Our empirical study contributes to the project of developing a social psychology of the citizen by focusing on the dynamics of such membership, specifically rights and identities. The authors briefly describe a number of existing psychological models of the citizen. Drawing on the main theoretical principles of discursive psychology, rather than asking, 'who is the citizen?' in terms of mental states, we suggest a shift in focus to the more social question, 'how do people claim citizenship and to what ends?'. We present an analysis of private letters of complaint that formed part of a larger mixed data set used in a recent research programme centred on disputes over Britain's newer travellers' rights of settlement. Specifically our analysis demonstrates how some of the letter writers generate a basis for claims-making by making relevant a citizenship/ governance alignment of identities. We also demonstrate how the entitlements associated with the category citizen are built up and action-oriented rather than flowing from the (unproblematic) assumption of citizenship. Finally we discuss how citizenship can be used for the purposes of inclusion and exclusion. PMID- 15285831 TI - Affective and cognitive control of persons and behaviours. AB - Three studies assessed the relative contribution of affect and cognition to determining behavioural intentions for a variety of behaviours using both between participants and within-participants analyses. The between-participants analyses showed that affect tends to make more of a contribution than does cognition for more behaviours. However, the within-participants analyses indicated that there are strong individual differences among people. Some people are more under affective control, across behaviours, whereas other people are more under cognitive control. The most interesting finding was that, despite the potential independence of between-participants and within-participants analyses (Mischela, 1990), between-participants analyses on subsamples created from the within participants analyses showed significant dependence. The predictive validity of affect vs. cognition depended upon whether participants were affectively or cognitively controlled. PMID- 15285832 TI - Making sense of 'barebacking': gay men's narratives, unsafe sex and the 'resistance habitus'. AB - Why do some gay men continue to engage in unsafe sexual practices despite the known and widely publicized risks of HIV infection? Dominant models of health promotion have been criticized for reducing this complex psychosocial issue to a question of instrumental technique - 'use a condom every time'. Accordingly, there has recently been an attempt to develop a deeper understanding of unsafe sexual practices by paying attention to the 'subjective meaning' manifest in gay men's own accounts of their behaviour. However, this paper argues that such studies have failed to develop an adequate understanding of the historical nature of such behaviours and the way in which they are embedded in a 'cultural psyche' that the individual gay man may not even be consciously aware of. Accordingly, using published autobiographical and fictional narratives written by gay men over three different periods of gay history (pre-AIDS, during AIDS and 'post'-AIDS), this paper aims to show that contemporary 'barebacking' behaviour may constitute one manifestation of a 'resistance' or 'transgressional' 'habitus' that has remained a consistent feature of gay men's individual and social psyche since the early days of gay liberation. The paper discusses the potentially health-damaging implications of this 'habitus' and the possibility and desirability of facilitating change. PMID- 15285833 TI - Psychoanalysis as a resource for understanding emotional ruptures in the text: the case of defensive masculinities. AB - Recent theory and research on men and masculinities within feminist and critical social psychologies has largely drawn upon social constructionism and discourse analysis. This work has been useful in extending our understanding of contemporary discourses drawn upon by men to construct masculine identities and/or to construct 'subordinated others' such as women and gay men. But it has been pointed out that discourse analytic work does not adequately account for emotional or experiential dimensions to (masculine) identities. To address this problem, several writers have turned to versions of psychoanalytic theory as this perspective is directly concerned with emotional life. Psychoanalysis has been reworked so that concepts traditionally read as intra-psychic essences (e. g. anxiety, desire, defence) are re-interpreted as interpersonal and contextual. Informed by this work, I argue that a psychoanalytic, particularly Kleinian, reading of focus group discussions with heterosexual men can help illuminate aspects of the contemporary reproduction of masculinities. I use data collected from a 'men and masculinities' project and focus primarily on emotive talk which 'others' gay men and women. Concepts such as 'projection' are used to connect the men's constructions of others with shared anxieties about masculinities. The implications and advantages of pursuing psychoanalytic accounts of (masculine) subjectivities within social psychology are then discussed. PMID- 15285834 TI - Procrastination and counterfactual thinking: avoiding what might have been. AB - The possible negative consequences of counterfactuals were explored in the current study by examining the relationship between counterfactual direction and trait procrastination, a self-defeating behavioural style. Eighty participants generated counterfactuals in response to two experimental anxiety inductions. Trait procrastination was overall related to avoiding thoughts about how things could have been better (making more downward and relatively fewer upward counterfactuals) in response to the two anxiety-provoking scenarios, suggesting the involvement of a self-enhancement motive (mood repair). Evidence for the involvement of this self-motive in procrastinating behaviour also emerged, as procrastination was more related to making more downward counterfactuals for a delay-specific anxiety scenario than for a general anxiety scenario. The pattern of results supports the proposal that downward counterfactuals may be associated with negative behavioural styles such as procrastination and implicates self enhancement motives in this relationship. The behavioural and motivational consequences of downward counterfactuals are discussed and possible connections between downward counterfactuals and other self-defeating behaviours are presented. PMID- 15285835 TI - Why are men more likely to support group-based dominance than women? The mediating role of gender identification. AB - Arguing from a sociobiological perspective, Sidanius and Pratto (1999) have shown that the male/female difference in social dominance orientation (SDO) is largely invariant across cultural, situational and contextual boundaries. The main objective of this study was to test the validity of Social Dominance Theory (SDT) by contrasting it with a model derived from Social Identity Theory (SIT). More specifically, while SIT predicts that gender identification mediates the effect of gender on SDO, SDT predicts the reverse. According to SDT, the degree to which men and women endorse status legitimizing ideology should determine to what extent they identify with their gender group. Using structural equation modelling, the results provide strong support for the SIT model and no support for SDT predictions. Implications of these results for social dominance theory and its sociobiologically based invariance hypothesis are discussed. PMID- 15285836 TI - Prejudice on the stage: self-monitoring and the public expression of group attitudes. AB - According to self-monitoring theory (Snyder, 1987), high self-monitors tailor their self-presentation for the sake of desired public appearances, whereas low self-monitors are relatively unlikely to practice such situationally guided impression management strategies. It was therefore predicted that, when asked to publicly express their attitudes regarding a social group, high self-monitors would modify their expressive behaviour in a direction consistent with the attitudes attributed to their audience. Conversely, low self-monitors would be unaffected by their audience's attitude towards this group. A study was conducted to test this hypothesis: participants, whose level of self-monitoring and prejudice towards homosexuals had been previously measured, were asked to report their thoughts regarding this group in an open-ended manner. They anticipated discussing these thoughts with an audience perceived as either prejudiced or tolerant, or they expected that their responses would remain private. In line with predictions, high self-monitors expressed more prejudice when the audience was perceived as prejudiced than tolerant, whereas low self-monitors were not affected by the audience's attitude. PMID- 15285837 TI - Comparative evaluation of the powder properties and compression behaviour of a new cellulose-based direct compression excipient and Avicel PH-102. AB - This study compares the compression behaviour of a new cellulose-based tableting excipient, hereinafter referred to as UICEL-A/102, and Avicel PH-102, a commercial direct compression excipient commonly referred to as microcrystalline cellulose (MCC). UICEL-A/102 shows the cellulose II lattice, while Avicel PH-102 belongs to the cellulose I polymorphic form. The median particle diameters of UICEL-A/102 and Avicel PH-102 fractions used in the study were 107 and 97 microm, respectively. Compared with Avicel PH-102, UICEL-A/102 was more dense; the relative poured and tapped densities were: 0.277 and 0.327 (vs 0.195 and 0.248 for Avicel PH-102), respectively. The true density, rhotrue, of the two materials was comparable ( approximately 1.56 g cm(-3)). The slopes of the in-die and out of-die Heckel curves for Avicel PH-102 were steeper than for UICEL-A/102. The relative density versus applied pressure plot was in good agreement with the modified Heckel equation. The out-of-die and in-die minimal pressure susceptibility (chipmin) values calculated were 3.36 x 10(-3) and 8.09 x 10(-3) MPa(-1) for UICEL-A/102 and 8.00 x 10(-3) and 16.12 x 10(-3) MPa(-1) for Avicel PH-102, respectively. The elastic recovery profiles showed UICEL-A/102 to be more elastic than Avicel PH-102. In conclusion, UICEL-A/102 and Avicel PH-102 differ in their compression behaviour under pressure. The different polymorphic forms could provide a possible explanation. PMID- 15285838 TI - A comparison of topical formulations for the prevention of human schistosomiasis. AB - Recently, a dimeticone formulation has been shown to be effective at preventing Schistosoma cercariae infecting skin, while DEET (N,N-diethyl-m-toluamide), a highly effective insecticide, has been shown to have activity against cercariae. Seven formulations, 3 containing DEET, were prepared and applied to excised human skin in Franz cells for 1 h. Schistosoma cercariae were applied for 30 min at 1 and 24 h, and the number that penetrated the skin calculated (n = 9). DEET could not be incorporated into the dimeticone formulation, yet it remained the most effective at preventing cercarial penetration, both 1 and 24 h after application. The ointments that contained DEET did prevent penetration but their mode of action was due to the toxicity of DEET against the cercariae. The persistence of the protection afforded by the dimeticone formulation after washing suggests that the formulation may be interacting with the stratum corneum to prevent cercarial recognition of skin. PMID- 15285840 TI - Identification of P-glycoprotein substrates and inhibitors among psychoactive compounds--implications for pharmacokinetics of selected substrates. AB - The pharmacokinetics of antipsychotic drugs has become an integral part in understanding their pharmacodynamic activity and clinical effects. In addition to metabolism aspects, carrier-mediated transport, particularly secretion by ABC transporters, has been discussed as potentially relevant for this group of therapeutics. In this study, the psychoactive compounds perphenazine, flupentixol, domperidone, desmethyl clozapine, haloperidol, fluphenazine, fluvoxamine, olanzapine, levomepromazine, perazine, desmethyl perazine, clozapine, quetiapine and amisulpride were characterized in terms of P glycoprotein (P-gp) affinity and transport. Experimental methods involved a radioligand displacement assay with [3H]talinolol as radioligand and transport- as well as transport inhibition--studies of the P-gp substrate [3H]talinolol across Caco-2 cell monolayers. In addition, the physicochemical descriptors log P and deltalog P were determined to test potential correlations between transporter affinity and lipophilicity parameters. All of the tested antipsychotics showed affinity to P-gp albeit their IC50 values (concentration of competitor that displaced 50% of the bound radioligand) differed by a factor exceeding 1000, when compared using the transport inhibition assay. From the group of P-gp substrates, amisulpride and fluphenazine were selected for in-vivo drug-drug interaction studies in rats to demonstrate the in-vivo relevance of the in-vitro findings. Compounds were administered by intraperitoneal injection either alone or in combination with 50 mg kg(-1) ciclosporin. The concentration versus time profiles for both drugs were followed in serum as well as in brain tissues. Significant differences between the treatments with the antipsychotic alone versus the combination of antipsychotic with ciclosporin were found for amisulpride. The distribution of amisulpride to the brain was increased and systemic serum levels were likewise increased indicating decreased systemic clearance for the combination regimen. For fluphenazine, systemic levels with and without co administration of ciclosporin were comparable while higher brain-to-serum concentration ratios were found after co-administration of ciclosporin. The findings are explained on the basis of the limited contribution of P-gp-mediated transport to the elimination of fluphenazine and to a direct effect with respect to its distribution into the brain. PMID- 15285839 TI - Ethanol inhibits in-vitro metabolism of nifedipine, triazolam and testosterone in human liver microsomes. AB - Although extended exposure to ethanol induces CYP3A metabolism in-vivo, the acute effects of ethanol on CYP3A metabolism have not been fully evaluated in-vitro. We assessed the effect of ethanol on CYP3A-mediated biotransformation using human liver microsomes in-vitro with three prototypic CYP3A-mediated reactions: nifedipine to oxidized nifedipine, triazolam to its 1-hydroxy (1-OH TRZ) and 4 hydroxy (4-OH TRZ) metabolites, and testosterone to 6beta-hydroxytestosterone (6beta-OH TST). Ethanol inhibited metabolism of nifedipine (oxidized nifedipine IC50 3 mg dL(-1), where the IC50 value is the inhibitor concentration corresponding to a 50% reduction in metabolite formation velocity), triazolam (1 OH TRZ IC50 1.1 mg dL(-1), 4-OH TRZ IC50 2.7 mg dL(-1)) and testosterone (6beta OH TST IC50 2.4 mg dL(-1)). The inhibitory potency of ethanol was similar for the three substrates representing the three hypothetical CYP3A substrate categories. The IC50 values obtained were lower than clinically relevant blood alcohol concentrations. In conclusion, ethanol is an inhibitor of human CYP3A metabolism and may contribute to clinically important interactions. PMID- 15285841 TI - Comparative assessment of ocular tissue distribution of drug-related radioactivity after chronic oral administration of 14C-levofloxacin and 14C chloroquine in pigmented rats. AB - Fluoroquinolones have been reported to have a high affinity for melanin. The ocular tissue distribution and accumulation of radioactivity was compared after repeated oral administration of 14C-levofloxacin and 14C-chloroquine at daily doses of 20 mg (0.054 mmol) kg(-1) and 28 mg (0.054 mmol) kg(-1), respectively, in pigmented rats for 84 days. The mean serum level at 24 h following each dose of 14C-levofloxacin was almost constant in the range of 0.33-0.45 nmol equiv mL( 1) after the 14th dose and thereafter. The melanin-containing ocular tissues, such as iris ciliary body and stratum pigment chorioides sclera, showed a much higher concentration of radioactivity than other non-pigmented ocular tissues. The respective concentration in iris ciliary body and stratum pigment chorioides sclera after the 1st dose was 126.47 and 74.91 nmol equiv g(-1), and gradually increased with increasing dose number, reaching 1261.81 and 447.45 nmol equiv g( 1) after the 84th dose, which was ca. 10 and 6 times higher, respectively, than after the 1st dose. The mean serum level following each dose of 14C-chloroquine was almost constant in the range 0.51-0.87 nmol equiv mL(-1) after the 7th dose and thereafter. The respective concentration in iris ciliary body and stratum pigment chorioides sclera after the 1st dose was 572.10 and 709.41 nmol equiv g( 1), and gradually increased with increasing dose number, reaching 33 317.92 and 12 322.90 nmol equiv g(-1) after the 84th dose, which was ca. 58 and 17 times higher, respectively, than after the 1st dose. The concentration in aqueous humour, cornea, lens, vitreous body and retina after the 84th dose was 1.84, 6.33, 0.48, 5.60 and 11.42 nmol equiv g(-1) for 14C-levofloxacin and 18.84, 264.99, 27.26, 158.43 and 1020.89 nmol equiv g(-1) for 14C-chloroquine (ca. 10, 42, 57, 28 and 89 times higher, respectively, than for 14C-levofloxacin). Especially, the concentration in the retina was markedly higher after 14C chloroquine administration than after 14C-levofloxacin administration. The concentration and the extent of accumulation of radioactivity not only in melanin containing ocular tissues but also in other non-pigmented ocular tissues, such as retina, after chronic oral administration of 14C-levofloxacin once daily for 84 days were much lower than those after multiple dosing with 14C-chloroquine under the same conditions. These results indicate that levofloxacin would have a much lower risk for ocular toxicity than chloroquine after chronic dosing. PMID- 15285842 TI - Concentration dependency of modulatory effect of amlodipine on P-glycoprotein efflux activity of doxorubicin--a comparison with tamoxifen. AB - Modulators of P-glycoprotein (P-gp) can enhance or limit the permeability of a number of therapeutic agents that are considered substrates of this efflux pump protein. The modulatory effect of amlodipine (4-dihydropyridine calcium antagonist) on P-gp efflux activity has not been fully elucidated. We have studied the concentration dependency of its modulatory effect and compared it qualitatively with tamoxifen (a non-esteroid anti-estrogen). The investigation was conducted on transmembrane efflux of doxorubicin at a fixed concentration of 5 microM across a Caco-2 monolayer in the presence of various concentrations of amlodipine or tamoxifen. The maximum flux of doxorubicin from basolateral to apical (ba) occurred at 4.5 microM amlodipine and at 0.02 microM tamoxifen. At higher concentrations, the apical to basolateral (ab) flux and the net flux of doxorubicin (ba - ab) declined steadily in a concentration-dependent manner. We analysed the observed net flux data by fitting different mathematical models to the data. A composite sigmoidal Emax/Imax (stimulatory/inhibitory) model was found to be the most appropriate to define the system. The observed and calculated parameters supported the modulatory role of both compounds and clearly indicated that the stimulation and inhibition of transmembrane efflux occurred simultaneously in the presence of amlodipine or tamoxifen. It was concluded that amlodipine, similar to tamoxifen, modulated the transporter-dependent transmembrane flux of the P-gp substrate in a concentration-dependent manner. PMID- 15285843 TI - Lack of congruence between cysteine dioxygenase activity and S-carboxymethyl-L cysteine S-oxidation activity in rat cytosol. AB - The identity of the enzyme(s) responsible for the S-oxidation of the mucoactive drug S-carboxymethyl-L-cysteine (SCMC) is unknown but the protein(s) are a susceptibility factor for a number of chronic degenerative diseases. The structural similarities between the amino acid L-cysteine and SCMC have raised the possibility that cysteine dioxygenase (CDO) may be responsible for this biotransformation reaction. Both CDO and SCMC S-oxygenase were found to require Fe2+ for enzymatic activity, and both enzyme activities were inhibited by Fe2+ and Fe3+ chelators. However, sulphydryl group modification of the enzymes resulted in the activation of the S-oxidation of SCMC but inhibition of the S oxidation of L-cysteine. When the two enzyme activities were quantified in 20 female hepatic cytosolic fractions no linear correlation in the production of their respective metabolites was seen. The results of this investigation indicate that CDO is not responsible for the S-oxidation of SCMC in the rat. PMID- 15285844 TI - Inhibition of P-glycoprotein function by tea catechins in KB-C2 cells. AB - We studied the effects of tea catechins, (-)-epicatechin (EC), (-) epigallocatechin (EGC), (-)-epicatechin gallate (ECG), and (-)-epigallocatechin gallate (EGCG) on the P-glycoprotein (P-gp) function in multidrug-resistant P-gp over-expressing KB-C2 cells. EC did not have any effects on cellular accumulation of P-gp substrates, rhodamine-123 and daunorubicin, but the other catechins increased the accumulation in the order of EGC < ECG < EGCG. The effects of EGCG were larger than those of verapamil and quercetin. Since these catechins inhibited the efflux of P-gp substrates, the elevation of substrate accumulation seemed to be induced by the inhibition of the efflux transporter. The results showed that the inhibitory effects of the catechins did not depend on their total hydrophobicity, but significantly depended on their chemical structure. The presence of the galloyl moiety on the C-ring markedly increased the n-octanol/PBS partition coefficients of the catechins and their activity on P-gp. On the other hand, the presence of the trihydric pyrogallol group as the B-ring decreased the partition coefficients but increased the activity on P-gp, compared with the action of the corresponding catechins with a dihydric catechol B-ring. PMID- 15285845 TI - Inhibition of in-vitro simvastatin metabolism in rat liver microsomes by bergamottin, a component of grapefruit juice. AB - Grapefruit juice can modify the pharmacokinetic parameters of many drugs, in particular simvastatin, an orally active cholesterol-lowering agent. The exact components in grapefruit juice responsible for drug interactions are not perfectly known. However, it seems that bergamottin, a furocoumarin derivative, is one of the main active components within grapefruit juice. The objective of this paper was to quantify and to characterize in-vitro the inhibitory effect of bergamottin on simvastatin metabolism by using rat and human liver microsomes. In rat liver microsomes, the incubation conditions (+/-NADPH) of bergamottin were found to influence its inhibiting capacity. In co-incubation with simvastatin, the Ki value (the equilibrium dissociation constant for the enzyme-inhibitor complex) was higher (Ki = 174 +/- 36 microM) than in pre-incubation (Ki = 45 +/- 6 microM and 4 +/- 2 microM, without and with NADPH, respectively). It thus seems that the pre-incubation of bergamottin (in particular with NADPH) increases its inhibiting capacity on simvastatin metabolism. Bergamottin metabolism study in rat liver microsomes showed the formation of two metabolites that were CYP-450 dependent. In contrast, in human liver microsomes, the incubation conditions of bergamottin did not influence its inhibiting capacity of simvastatin metabolism (Ki = 34 +/- 5 microM, Ki = 22 +/- 5 microM, Ki = 27 +/- 11 microM in coincubation and pre-incubation without and with NADPH, respectively). In rat and man, bergamottin was found to be a mixed-type inhibitor of simvastatin hepatic metabolism. However, in rat, bergamottin was partially a mechanism-based inhibitor by involvement of either bergamottin alone or one of its metabolites. The results highlight the importance of validating in-vitro models to help verify the suitability of the in-vitro model for predicting the nature and degree of metabolic drug interactions. PMID- 15285846 TI - Lack of effect of proteinase-activated receptor-2 (PAR-2) deletion on the pathophysiological changes produced by lipopolysaccharide in the mouse: comparison with dexamethasone. AB - This study tested the hypothesis that activation of proteinase-activated receptor 2 (PAR-2) contributes towards the pathophysiology of lipopolysaccharide (LPS) induced shock in the mouse. The effects of LPS on plasma glucose, biochemical markers of hepatic, renal and pancreatic exocrine function and lung content of myeloperoxidase (MPO) were examined in homozygous PAR-2 knockout mice (PAR-2 -/-) and genetically equivalent, homozygous PAR-2 +/+ mice. The effect of LPS was also examined in normal mice receiving dexamethasone (10 mg kg(-1), i. p.) or saline as a positive control. At six hours after intraperitoneal injection, LPS (40 mg kg(-1)) produced an increase in rectal temperature, hypoglycaemia and elevations in serum concentrations of alanine aminotransferase (ALT), creatinine and lipase, as well as an increase in lung MPO content. Dexamethasone treatment reduced LPS induced hypoglycaemia and elevation of serum ALT concentrations but did not modify elevations in serum creatinine and lipase concentrations or the increase in lung MPO content. The changes in serum concentrations of glucose, ALT, creatinine and lipase produced by LPS in PAR-2 -/- mice were not different from those seen in wild-type or PAR-2 +/+ mice. These data suggest that activation of PAR-2 may not play a pivotal role in LPS-induced multi-organ dysfunction. PMID- 15285847 TI - Calcium channel antagonists attenuate cross-sensitization to the rewarding and/or locomotor effects of nicotine, morphine and MK-801. AB - The present study focused on the evaluation of behavioural cross-sensitization, particularly in locomotor activities and conditioned rewarding effects, between nicotine and morphine, cocaine, amphetamine or MK-801. Nicotine (0.5 mg kg(-1)) experienced mice manifested an enhanced locomotor response to morphine (5 mg kg( 1)) or MK-801 (0.3 mg kg(-1)). No cross-sensitization was observed between nicotine and amphetamine (2 mg kg(-1)) or cocaine (15 mg kg(-1)). Additionally, the L-type voltage-dependent calcium-channel antagonists, nimodipine and verapamil, but not diltiazem, at a dose of 20 mg kg(-1) injected before morphine or MK-801 challenge, blocked the expression of this cross-sensitization. In the second test, an enhancement of morphine place conditioning in rats pre-exposed to nicotine (0.5 mg kg(-1), injected daily for 5 days) was demonstrated. After two conditioning sessions, morphine (5 mg kg(-1)) induced a clear place preference only in animals that had previously received nicotine injections. The administration of nimodipine (10 and 20 mg kg(-1)), verapamil (10 and 20 mg kg( 1)) and diltiazem (10 and 20 mg kg(-1)) prior to nicotine dose-dependently prevented this sensitization to the rewarding effect of morphine produced by prior injections of nicotine. These findings support the hypothesis that similar neural calcium-dependent mechanisms are involved in the appetitive effects of nicotine and morphine and in the sensitized locomotor stimulant effects of nicotine and morphine or MK-801. PMID- 15285848 TI - Solid-phase synthesis and evaluation of libraries of substituted 4,5 dihydropyridazinones as vasodilator agents. AB - The solid-phase parallel preparation of a library of 4,5-dihydropyridazin-3(2H) one derivatives substituted at position 6 with piperazinylmethyl or tetrahydroquinolinylmethyl groups and analogues (3) is reported. Polymer supported gamma-keto-delta-aminoesters prepared from Wang resin reacted with hydrazine or methylhydrazine to afford pyridazinones in good yields after a cyclization cleavage approach. We have evaluated these novel analogues and several compounds of other series (1, 2) for their vasorelaxant effect. Among the products tested, 3l and 3d proved to be efficacious and potent relaxant agents of the isolated rat aorta. Inhibitors of phosphodiesterase (PDE3), responsible for the breakdown of cyclic AMP in the vascular smooth muscle, are currently developed for cardiac heart failure because of their inotropic effect and coronary vasodilatation. We had expected that the vasodilatation induced by 3l, as efficient as reference PDE3 inhibitors, milrinone or CI-930, to be due to PDE3 inhibition. However 3l and 3d exhibited a low inhibitory effect against PDE3 isoenzyme activity. These compounds induced a significant vasorelaxation, which could be of therapeutic interest even if their mechanism of action remains to be determined. PMID- 15285849 TI - Inhibition of human cytochromes P450 by components of Ginkgo biloba. AB - The extraction, isolation and characterization of 29 natural products contained in Ginkgo biloba have been described, which we have now tested for their in-vitro capacity to inhibit the five major human cytochrome P450 (CYP) isoforms in human liver microsomes. Weak or negligible inhibitory activity was found for the terpene trilactones (ginkgolides A, B, C and J, and bilobalide), and the flavonol glycosides. However 50% inhibitory activity (IC50) was found at concentrations less than 10 microg L(-1) for the flavonol aglycones (kaempferol, quercetin, apigenin, myricetin, tamarixetin) with CYP1A2 and CYP3A. Quercetin, the biflavone amentoflavone, sesamin, as well as (Z,Z)-4,4'-(1,4-pentadiene-1,5-diyl)diphenol and 3-nonadec-8-enyl-benzene-1,2-diol, were also inhibitors of CYP2C9. The IC50 of amentoflavone for CYP2C9 was 0.019 microg mL(-1) (0.035 microM). Thus, the principal components of Ginkgo biloba preparations in clinical use (terpene trilactones and flavonol glycosides) do not significantly inhibit these human CYPs in-vitro. However, flavonol aglycones, the biflavonol amentoflavone and several other non-glycosidic constituents are significant in-vitro inhibitors of CYP. The clinical importance of these potential inhibitors will depend on their amounts in ginkgo preparations sold to the public, and the extent to which their bioavailability allows them to reach the CYP enzymes in-situ. PMID- 15285850 TI - Parameters of drug antagonism: re-examination of two modes of functional competitive drug antagonism on intraocular muscles. AB - There are two distinct kinetic functional pharmacological procedures by which the equilibrium affinity constant, KB, of a competitive reversible blocker is obtained. The classical method on an organ system requires the study of the parallel displacement of the agonist concentration-response curve in the presence of the blocker. In the second method, the agonist-evoked functional mechanical response is reduced to half by the blocker IC50 (the concentration required for 50% inhibition). In relation to these parameters the role of the ionization constant pKa and liposolubility log Pc or log D of blockers was examined. On the ciliary muscle from human eye, IC50/KB ratios for (+/-)-atropine, its quaternary analogue (+/-)-methylatropine, (-)-scopolamine, (+/-)-cyclopentolate, (-) tropicamide, (+/-)-oxybutynin and pirenzepine were 15, 23, 4.4, 2.6, 1.66, 1.46 and 1.71, respectively. The ratios on the iris sphincter were comparable with those of ciliary muscle. When compared with large proportions of ionized molecules with water soluble properties of (+/-)-atropine and (+/-) methylatropine, relatively high amounts of un-ionized and/or with greater partitioning of all other blockers in the lipoid barrier correlated well to low IC50/KB ratios, as predicted by the classical theory of competitive drug antagonism. It was hypothesized that due to receptor biophase access, the reduction of the mechanical response of the agonist by the highly ionized water soluble antagonist at IC50 represented time-distorted "pseudoequilibrium" estimation, where a higher concentration of the blocker was needed. On the other cholinergic effectors, like that of rat anococcygeus muscle or frog rectus abdominus muscle, IC50/KB ratios of respective blockers atropine or (+) tubocurarine and hexamethonium were close to 1. Thus physicochemical properties, which affect the distribution coefficient log D and the tissue morphology (where asymmetric distribution of receptors may occur), appeared to be a critical factor in the analysis of the affinity parameters of the competitive reversible blocker. On the intraocular muscles, two functional pharmacological procedures for obtaining KB and IC50 values were not kinetically equivalent. PMID- 15285851 TI - Effect of lansoprazole and rabeprazole on tacrolimus pharmacokinetics in healthy volunteers with CYP2C19 mutations. AB - The aim of this study was to investigate the effects of the proton pump inhibitors (PPIs), lansoprazole and rabeprazole, on tacrolimus pharmacokinetics in healthy volunteers with mutations in the cytochrome P450 (CYP) 2C19 gene (CYP2C19). An open-label crossover study was performed with 19 healthy subjects. Tacrolimus (2 mg) was administered orally with and without lansoprazole (30 mg per day for 4 days) or rabeprazole (10 mg per day for 4 days). Blood concentrations of tacrolimus were determined before and 1, 2, 4 and 8 h after dosing. Genotyping for CYP2C19 was conducted by a polymerase chain reaction restriction fragment length polymorphism method. Coadministration of lansoprazole significantly decreased the oral tacrolimus clearance, resulting in an increase in the area under the blood concentration-time curve (AUC0-8) (control vs with lansoprazole: 29.7 +/- 3.5 vs 44.1 +/- 5.0 ng h mL(-1), P < 0.05). Large individual variation was observed in the effects of lansoprazole on tacrolimus AUC0-8 owing to CYP2C19 genotype status. The percent change for tacrolimus AUC0-8 in subjects with and without CYP2C19 mutant alleles was 81% and 29%, respectively. Coadministration of rabeprazole also increased the mean AUC0-8 of tacrolimus, but the difference was not statistically significant. These observations suggest that drug interaction between tacrolimus and lansoprazole occurs in subjects with higher lansoprazole blood concentrations corresponding to CYP2C19 genetic status. In contrast, rabeprazole has minimal effect on tacrolimus pharmacokinetics regardless of CYP2C19 genotype status. PMID- 15285852 TI - Reversal of P-gp mediated multidrug resistance in-vitro and in-vivo by FG020318. AB - Overexpression of P-glycoprotein (P-gp) by tumours results in multidrug resistance (MDR) to structurally and functionally unrelated chemotherapeutic drugs. Combined therapy with MDR-related cytotoxins and MDR modulators is a promising strategy to overcome clinical MDR. This study was performed to explore the MDR reversal activity of a novel compound 2-[4-(2-pyridin-2-yl-vinyl) phenyl] 4,5-bis-(4-N,N-diethylaminophenyl)-1(H)-imidazole (FG020318) in-vitro and in vivo. Tetrazolium (MTT) assay was used to evaluate the ability of FG020318 to reverse drug resistance in two P-gp-expressing tumour cell lines, KBv200 and MCF 7/adr. Intracellular doxorubicin accumulation was determined by fluorescence spectrophotometry in MCF-7/adr cell line. The effect of FG020318 on P-gp function was demonstrated by rhodamine 123 (Rh123) accumulation in KBv200 cells. KBv200 cell xenograft models were established to study the in-vivo effect of FG020318 on reversing MDR. FG020318 was not cytotoxic by itself against P-gp expressing KBv200 cells and MCF-7/adr cells and their parental drug-sensitive KB cells and MCF-7 cells. FG020318 could significantly increase the sensitivity of MDR cells to antitumour drugs including doxorubicin and vincristine in MCF-7/adr cells and KBv200 cells, respectively. It was much stronger than the positive control verapamil in reversal of MDR. FG020318 also increased the intracellular accumulation of doxorubicin in a concentration-dependent manner in MCF-7/adr cells, but did not affect the accumulation of doxorubicin in drug-sensitive MCF-7 cells. The Rh123 accumulation in resistant KBv200 cells was also increased by the addition of FG020318, but Rh123 accumulation was not affected by FG020318 in drug sensitive KB cells. FG020318 potentiated the antitumour activity of vincristine to KBv200 xenografts and was an efficacious modulator in-vivo. Our results suggested that FG020318 was a highly potent, efficacious MDR modulator not only in-vitro but also in-vivo. The reversal of drug resistance by FG020318 was probably related to the increased anticancer drug accumulation and its inhibition of P-gp function of MDR tumour cells. PMID- 15285853 TI - Neck dissection: historical perspective. PMID- 15285854 TI - Ear, nose and throat manifestations of ectodermal dysplasia. AB - This review of ectodermal dyplasias (ED) presents the particular syndromes that might present to the otolaryngologist for management and discusses the ear, nose and throat manifestations of the condition. PMID- 15285855 TI - Training opportunities for specialist registrars post-calmanization: audit of trainees' exposure to repair of the tympanic membrane. AB - Specialist training at the registrar grade in the United Kingdom has undergone significant change in the recent past. One of the effects has been a reduction in the length of time spent in training. This and the application of the European working time directive have the potential to reduce trainees' surgical exposure. The proportion of tympanic membrane procedures performed by reconstructive otolaryngology registrars in the Grampian University hospitals was audited to monitor the impact of these changes. Case notes of all patients who underwent myringoplasty or tympanoplasty between July 1998 and June 1999 were analysed retrospectively. Details of the surgeons' grade were recorded. The proportion of myringoplasties performed by registrars as determined by the Royal College of Surgeons of England National Comparative Audit survey, carried out in 1995 before the widespread implementation of recommended changes in otolaryngology registrar training, was set as the gold standard. In the period July 1998-1999 registrars had performed fewer myringoplasties than the standard, 17 per cent versus 34.2 per cent respectively (p = 0.035). A strategy to increase registrar exposure to myringoplasty surgery was then adopted by the department and the proportion of myringoplasties performed by registrars re-audited prospectively. The proportion of myringoplasties undertaken by registrars increased in the period January 2001 to July 2001 compared to July 1998-1999, 53 per cent versus 17 per cent respectively (p < 0.0007). Changes in working practice can address shortfalls in registrars' exposure to surgical procedures. PMID- 15285856 TI - Noise exposure in orthopaedic practice: potential health risk. AB - Noise exposure is one of the major causes of permanent hearing loss in society. Exposure of health service staff to intense levels of noise in the workplace is a potential risk for the development of temporary and permanent hearing loss. In this prospective study, 18 members of the orthopaedic staff underwent hearing assessment by pure tone audiometry and speech discrimination prior to noise exposure at the workplace and immediately following cessation of work. The number of hours of exposure and noise levels in the workplace was also analysed. Only minimal temporary sensorineural threshold shifts were detected post-noise exposure. There was no change in speech discrimination scores and no individuals complained of tinnitus. The number of hours of exposure ranged from 1.5 to 8.5 hours (mean 5.2 hours). Recorded sound levels for instruments ranged from 119.6 dB at source to 73.1 decibels at 3 metres. Although high sound levels are recorded in the orthopaedic operating theatre, the intermittent nature exposure to the intense noise may protect staff against hearing loss, speech discrimination difficulties and tinnitus. PMID- 15285857 TI - Complications following adult cochlear implantation: experience in Manchester. AB - Cochlear implantation is regarded as a safe and effective treatment for the profoundly deaf. However, a proportion of patients suffer complications after implant surgery. This paper examines the complications encountered in 240 adult cochlear implant operations performed in Manchester between June 1988 and June 2002. Minor complications were defined as those that either settled spontaneously or with conservative management. The total number of minor complications was 61 (25.4 per cent of cases). Non-auditory stimulation, which resolved with implant reprogramming, was present in 53 cases (22.1 per cent). Major complications were defined as those requiring further surgery, explantation or causing a significant medical problem, and occurred in 15 patients (6.25 per cent). These included implant extrusion, implant sepsis, electrode migration, flap-related problems, and persistent non-auditory stimulation. Nine of the 15 patients suffering a major complication required explantation. There were no post-operative deaths, cases of meningitis, nor persistent facial palsies in the series. PMID- 15285858 TI - Waveform reliability with different recording electrode placement in facial electroneuronography. AB - Electroneuronography (ENoG) has become a useful test for estimating the degree of facial nerve degeneration and predicting the prognosis in patients with facial nerve palsy. Test results may be influenced by several factors, including the electrode positions, skin resistance, stimulus magnitude, and possible artifacts. Regarding recording electrode positions, different groups have used two different locations, the nasolabial fold and nasal ala. The authors compared the waveforms recorded from these two locations in ENoG recordings to obtain the optimal waveform. Twenty healthy volunteers and 25 patients with unilateral facial nerve palsy were included in this study. Recordings were carried out with the recording electrode placed on the nasolabial fold, followed by placement on the nasal ala after 10 minutes. The following parameters were assessed: (1) the supramaximal threshold, (2) amplitude and shape of the waveform, (3) interside difference, and (4) test-retest variability. There was no significant difference in the amplitude of the waveform, interside difference, and test-retest variability between the two groups. However, when the electrode was placed on the nasal ala, the threshold was significantly lower, an ideal biphasic configuration was present in almost all cases (97.5 per cent) of normal volunteers and it was easier to identify the waveform. Placement of the recording electrode on the nasal ala would be the preferred method. PMID- 15285859 TI - Endoscopic dacryocystorhinostomy in cases of dacryocystitis due to atrophic rhinitis. AB - Atrophic rhinitis is a chronic inflammatory disease of the nose, which is more common in India. Chronic dacryocystitis is its rare complication. The authors found four cases of chronic dacryocystitis from March 2002 to October 2003 due to atrophic rhinitis. It was diagnosed clinically by the regurgitation test and lacrimal syringing. These cases were treated conservatively for a period of six weeks to make the nasal mucosa healthier and were then subjected to endoscopic dacryocystorhinostomy (end-DCR) under local anaesthesia. The procedure was found to be more difficult due to bleeding and the healing time was prolonged as compared to other cases of end-DCR. After one to one and half years of follow-up the primary success rate was 75 per cent but after revision surgery in one case, all cases were successful. Hence it was concluded that atrophic rhinitis is no more a contraindication for end-DCR. However, meticulous initial preparation and post-operative follow-up is necessary to improve the result. PMID- 15285860 TI - Laryngeal ultrasound to assess vocal fold paralysis in children. AB - The purpose of this study was to assess the practicality and validity of laryngeal ultrasound to establish vocal fold movement in children with suspected vocal fold palsy. Fifty-five consecutive patients (age range three days to 12 years) with suspected vocal fold palsy underwent both laryngoscopy and laryngeal ultrasound. Ultrasonographic findings correlated with endoscopic findings in 81.2 per cent of cases. This, however, rose to a concordance rate of 89.5 per cent in patients aged over 12 months. Laryngeal ultrasound is well-tolerated, safe and non-invasive and the authors feel that it is a useful adjunct to endoscopy in the diagnosis of vocal fold palsy. PMID- 15285861 TI - Optimizing the assessment of quality of life after laryngeal cancer treatment. AB - Reports of the impact of larynx cancer treatment modality on quality of life are conflicting, in part due to varying study methodology. The aims of this study were to (1) provide preliminary comparisons of quality of life following radiotherapy or combination therapy; (2) evaluate a number of measures of quality of life and thereby (3) inform future prospective studies. Thirty-six laryngeal cancer patients, 24 following radiotherapy, 12 following radiotherapy and laryngectomy completed the Functional Assessment of Cancer Therapy (FACT) - General/Head and Neck subscale; Nottingham Health Profile (NHP); and the Hospital Anxiety and Depression scale (HAD), three to 12 months post-treatment. Results showed trends towards a less good quality of life in the combined therapy group over a wide range of outcomes, significant for the disease specific FACT head and neck subscale, NHP emotion (p = 0.04) and isolation (p = 0.027). To the authors' knowledge, however, this is the first demonstration of greater impact of laryngeal cancer on quality of life in younger subjects, who had lower scores among others on emotional wellbeing (p = 0.015) and anxiety (p = 0.035). Younger patients thus appear more likely to need more intensive support through treatment. Many of the physical and psychosocial domains derived from the three tools used were highly correlated. In other words, given the known high morbidity of the disease and its treatment, the selection of tools for head and neck quality of life assessment may be much less important than their universal application. PMID- 15285862 TI - Use of steroids in the treatment of peritonsillar abscess. AB - Peritonsillar abscess is the most common deep infection of the head and neck that occurs in adults; the treatment of the disease remains controversial. A prospective study using a single high dose steroid treatment for peritonsillar abscess, was undertaken in 62 patients to determine the treatment's effectiveness in relieving symptoms such as fever, throat pain, dysphagia and trismus. All patients were randomly assigned to two groups: 28 patients received intravenous antibiotic therapy and a single dose placebo and 34 patients were treated with single use of high dose steroid in addition to intravenous antibiotic. Patients were hospitalized after needle aspiration and therefore their clinical courses and responses to therapy could be rigorously assessed. Comparison of clinical outcomes with respect to hours hospitalized, throat pain, fever, trismus were assessed between the two groups. Clinical outcomes revealed a statistically significant difference between the two groups (p < 0.01), indicating that single use of high dose steroid prior to antibiotic therapy is more effective than the use of an antibiotic alone. These results suggest that single intravenous use of steroid in addition to antibiotic therapy is an excellent choice for the management of peritonsillar abscess. PMID- 15285863 TI - Surgical treatment of squamous cell carcinoma of the hypopharynx: analysis of treatment results, failure patterns, and prognostic factors. AB - Patients with squamous cell carcinoma of the hypopharynx (HPSCC) are often seen in advanced stages and have a poor prognosis. The authors analysed 104 patients who had HPSCC and underwent surgery as the primary treatment between 1986 and 1995 in their institute. Of the 104 patients, 83 patients (80 per cent) had advanced T(3) or T(4) staged and 64 patients (62 per cent) had cervical metastasis. Thirteen patients (13 per cent) had conservation surgery with laryngeal preservation and 69 patients (66 per cent) received post-operative radiotherapy. The five-year overall and disease-specific survival was 47 per cent and 62 per cent, respectively. Recurrence occurred in 38 patients (37 per cent), including 12 (12 per cent) with local, 22 (21 per cent) with regional, and 12 (12 per cent) with distant recurrence. Sixteen patients (15 per cent) had recurrence at multiple sites. The site and size of the primary tumour, neck biopsy before surgery, early post-operative complications, and pathological nodal stage were significant prognostic factors of disease-specific survival (DSS) in univariate analysis. Neck biopsy before surgery and site of primary tumour were significant factors in multivariate analysis. In conclusion, surgical treatment for the HPSCC patients has achieved good local-regional control and survival. Bilateral neck dissection for the tumour across the midline and avoiding neck biopsy before surgery may reduce regional recurrence. PMID- 15285864 TI - Sudden hearing loss in intralabyrinthine haemorrhage in a child. AB - This paper reports an unusual case in which aseptic meningitis presented with sudden sensorineural hearing loss (SSNHL) associated with intralabyrinthine haemorrhage (ILH). A seven-year-old girl presented with sudden right-sided hearing loss with dizziness. She did not have a previous history of bleeding disorders. This child was assessed using audiograms and magnetic resonance imaging (MRI). The patient's hearing loss was irreversible. Steroid therapy was not effective. SSNHL associated with ILH can be one of the negative prognostic factors in children. PMID- 15285865 TI - Antrochoanal polyp and obstructive sleep apnoea in children. AB - Antrochoanal polyps were first documented in the 18th century. They represent one of the most common types of nasal polyps in children without cystic fibrosis. Only a few reports on children who had a history of snoring due to an antrochoanal polyp and only two cases where the antrochoanal polyp caused documented obstructive sleep apnoea (OSA) have been published so far. This report adds a third case of paediatric OSA induced by an antrochoanal polyp in a 12-year old boy. After endonasal endoscopically-controlled polypectomy and a recurrence, transoral osteoplastic antrotomy in combination with endoscopic endonasal polypectomy eliminated the antrochoanal polyp and OSA was resolved. The authors have reviewed essential historical aspects about children suffering from snoring and/or OSA caused by an antrochoanal polyp. PMID- 15285866 TI - Corticosteroids and peritonsillar abscess formation in infectious mononucleosis. AB - Peritonsillar abscess formation is an uncommon complication of infectious mononucleosis (IM). Early case reports implicated corticosteroids in the development of such abscesses, however, subsequent studies suggested that these drugs do not promote the formation of abscesses at several sites outside the central nervous system. It has recently been demonstrated that zwitterionic polysaccharides, in bacterial capsules, form complexes with CD4(+) T lymphocytes leading to abscess formation. A patient is presented who developed peritonsillar abscess a few days after initiation of corticosteroid therapy for IM; the medical literature was reviewed in respect of this subject. It appears that the occurrence of these abscesses in IM is not strongly linked to corticosteroid treatment. The authors, therefore, recommend that steroids should not be withheld from patients with severe IM on the basis that they may precipitate the development of peritonsillar abscess. PMID- 15285867 TI - Tracheal stenting: a better method of dealing with airway obstruction due to thyroid malignancies than tracheostomy. AB - Tracheostomy remains the primary method of treatment of acute airway obstruction due to malignant invasion and compression of the trachea. However, the development of tracheal stents has provided an alternative effective treatment modality. This case report and literature review highlights the benefits of intra luminal stenting, including resolution of distressing obstructive symptoms, and subsequent improvement in quality of life. PMID- 15285868 TI - Retropharyngeal space swelling secondary to minor blunt head and neck trauma. AB - Retropharyngeal space swelling is a rare occurrence following minor head and neck trauma. Upper airway obstruction is a potentially life-threatening sequela. The authors present a case of retropharyngeal space haematoma following minor blunt head and neck trauma. Management was conservative with gradual spontaneous resolution of the haematoma. The literature is reviewed and the management and treatment principles presented. PMID- 15285869 TI - Subcutaneous emphysema of the neck in infancy: under-recognized presentation of child abuse. AB - Two cases of subcutaneous emphysema of the neck as a result of abuse in infancy are presented to add to the variety of findings associated with the maltreatment of infants. Pharyngeal perforations as a result of abuse are rare. These cases are presented as a reminder to the attending clinician that although most cases of subcutaneous emphysema may resolve without any complications a detailed examination for the cause should be performed and the history carefully verified. PMID- 15285870 TI - Spontaneous cervical haematoma resulting from intrathoracic pathology. AB - Spontaneous cervical haematomas are rare occurrences for which a wide variety of aetiologies have been reported. Although the pathogenesis of this condition can be quite diverse, all known cases have emanated from a cervical source. The authors report the first case of a spontaneous cervical haematoma arising from an intrathoracic source. A case review and Medline search from 1962-2003 was carried out. No prior experience with intrathoracic sources for cervical haematomas has previously been reported. Transcervical haematoma evacuation confirmed the intrathoracic origin and a median sternotomy was required to gain control of this haemorrhage. Thoracic sources should be considered in the differential diagnosis of a spontaneous cervical haematoma. Preparations for a combined procedure with a thoracic surgeon can be predicted pre-operatively based on imaging studies. PMID- 15285871 TI - Hypercellularity of the mastoid as a cause of spontaneous pneumocephalus. AB - In this paper two cases are reported in which spontaneous entry of air into the head appears to have occurred through a hypercellular mastoid air cell system. In both these cases forceful sneezing and nose blowing were considered contributory factors. They underwent surgical repair of the bony defects which, combined with less vigorous nose blowing, has affected a successful repair. The aetiology of pneumocephalus is discussed and a review of the pertinent literature is also presented. PMID- 15285872 TI - Management of lateral sinus thrombosis: update and literature review. J Laryngol Otol 2003;117:932-9. PMID- 15285873 TI - Gold standard or wrong standard? PMID- 15285874 TI - Rationale for sentinel node biopsy to stage N0 head and neck squamous-cell carcinoma. AB - Sentinel node biopsy is an evolving staging tool in the management of patients with squamous-cell carcinoma of the head and neck. This tool provides a more detailed understanding of the lymphatic pathways within the head and neck and the role of these pathways regarding tumor spreading. By targeting nodes that are most likely to harbor disease, a more detailed pathological evaluation of sentinel nodes is possible-thus improving staging by the identification of micrometastases. The rationale behind the use of sentinel node biopsy to stage the N0 neck are discussed within this paper. PMID- 15285875 TI - Low-dose-rate irradiation by 131I versus high-dose-rate external-beam irradiation in the rat pancreatic tumor cell line CA20948. AB - AIM: The rat pancreatic CA20948 tumor cell line is widely used in receptor targeted preclinical studies because many different peptide receptors are expressed on the cell membrane. The response of the tumor cells to peptide radionuclide therapy, however, is dependent on the cell line's radiosensitivity. Therefore, we measured the radiosensitivity of the CA20948 tumor cells by using clonogenic survival assays after high-energy external-beam radiotherapy (XRT) in vitro. It can, however, be expected that results of high-dose-rate XRT are not representative for those after low-dose-rate radionuclide therapy (RT), such as peptide-receptor radionuclide therapy. Therefore, we compared clonogenic survival in vitro in CA20948 tumor cells after increasing doses of XRT or RT, the latter using (131)I. METHODS: Survival of CA20948 cells was investigated using a clonogenic survival assay after RT by incubation with increasing amounts of (131)I, leading to doses of 1-10 Gy after 12 days of incubation (maximum dose rate, 0.92 mGy/min), or with doses of 1-10 Gy using an X-ray machine (dose rate, 0.66 Gy/min). Colonies were scored after a 12-day-incubation period. Also, the doubling time of this cell line was calculated. RESULTS: We observed a dose dependent reduction in tumor-cell survival, which, at low doses, was similar for XRT and RT. For high-dose-rate XRT, the quadratic over linear component ratio (alpha/beta) for CA20948 was 8.3 Gy, whereas that ratio for low-dose-rate RT was calculated to be 86.5 Gy. The calculated doubling time of CA20948 cells was 22 hours. CONCLUSIONS: Despite the huge differences in dose rate, RT tumor cell killing effects were approximately as effective as those of XRT at doses of 1 and 2 Gy, the latter being the common daily dose given in fractionated external-beam therapies. At higher doses, RT was less effective than XRT. PMID- 15285876 TI - Dosimetry model for radioactivity localized to intestinal mucosa. AB - BACKGROUND: This paper provides a new model for calculating radiation-absorbed doses to the full thickness of the small and large intestinal walls, and to the mucosal layers. The model was used to estimate the intestinal radiation doses from yttrium-90-labeled-DOTA-biotin binding to NR-LU-10-streptavidin in patients. METHODS: We selected model parameters from published data and observations, and used the model to calculate energy-absorbed fractions using the EGS4 radiation transport code. We determined the cumulated (90)Y activity in the small and large intestines of patients from gamma camera images, and calculated absorbed doses to the mucosal layer and to the whole intestinal wall. RESULTS: The mean absorbed dose to the wall of the small intestine was 16.2 mGy/MBq (60 cGy/mCi) administered from (90)Y localized in the mucosa, and 70 mGy/MBq (260 cGy/mCi) to the mucosal layer within the wall. Doses to the large intestinal wall and to the mucosa of the large intestine were lower than those for the small intestine by a factor of about 2.5. These doses are greater by factors of about 5 to 6 than those that would have been calculated using the standard MIRD models that assume the intestinal activity is in the bowel contents. CONCLUSIONS: The specific uptake of radiopharmaceuticals in mucosal tissues may lead to dose-related intestinal toxicities. Tissue dosimetry at the sub-organ level is useful for a better understanding of intestinal tract radiotoxicity and associated dose response relationships. PMID- 15285877 TI - A theoretical radiobiological assessment of the influence of radionuclide half life on tumor response in targeted radiotherapy when a constant kidney toxicity is maintained. AB - The potential of targeted radionuclide therapy may be limited if the antibody affinity to the tumor is relatively low and if significant normal tissue damage occurs while the tumor is sterilized. One way to increase the efficiency of the antibody-radionuclide complex might be to use knowledge of the radiobiological processes to select a near-optimal radionuclide half-life. In this paper, the role of physical half-life in targeted radiotherapy optimization is investigated using the linear quadratic (LQ) radiobiological model in conjunction with a range of radiobiological parameters relevant to the tumor. Five radionuclides ((211)At, (90)Y, (131)I, (86)Rb, and (114m)In) were selected, providing a half-life range from 0.3-49.5 days. The dose-limiting organ was assumed to be the kidney, with a simple fractional link between the initial (extrapolated) dose-rate to the tumor and the initial dose-rate to the kidney. The results suggest that short-lived radionuclides (half-life in the range of 1-10 days) have an advantage over medium and long-lived radionuclides. Furthermore, for very rapid tumor uptake (uptake half-time of a few hours), very short-lived radionuclides (half-life of less than 1 day) could be efficiently employed. Ultimately, however, treatment outcome (in terms of tumor cell kill) is limited by the antibody affinity to the tumor. PMID- 15285878 TI - Engineered fusion hybrid vaccine of IL-18 gene-modified tumor cells and dendritic cells induces enhanced antitumor immunity. AB - Dendritic cell (DC)-tumor fusion hybrid vaccines that facilitate antigen presentation represent a novel powerful strategy in cancer immunotherapy. In our study, we investigated the antitumor immunity derived from the vaccination of fusion hybrids between engineered J558/IL-18 myeloma cells secreting Th1 cytokine IL-18 and DCs. DC/J558/IL-18 could secret a higher level of IL-18 than DCs, efficiently expressed J558 tumor antigen P1A, and enhanced ability of allogeneic T cell stimulation when compared to J558/IL-18. Our data showed that the immunization of BALB/c mice with DC/J558/IL-18 hybrids induced the most potent protective immunity against 1 x 10(6) cells with a J558 tumor challenge, compared to those immunized with the mixture of DCs and J558/IL-18, J558/IL-18, or J558. Furthermore, the immunization of mice with engineered DC/J558/IL-18 hybrids elicited stronger NK activity and J558 tumor-specific cytotoxic T lymphocyte (CTL) responses in vitro. In addition, DC/J558/IL-18 tumor cells into syngeneic mice induced a Th1 dominant immune response to J558 and resulted in tumor regression, which indicated that the antitumor effect mediated by DC/J558/IL-18 appeared to be dependent on TH1 cytokine production. These results demonstrate that the engineered fusion hybrid vaccines that combine Th1 gene-modified tumor with DCs may be an attractive strategy for cancer immunotherapy. PMID- 15285879 TI - Predictive impact of retinoid X receptor-alpha-expression in renal-cell carcinoma. AB - PURPOSE: Retinoid receptors are nuclear transcription factors that mediate the effects of retinoids on gene expression. In our study, we analyzed the expression of retinoid X receptor-alpha (RXR-alpha) and its prognostic value in renal-cell carcinoma (RCC) patients exhibiting stage IV disease upon first diagnosis or thereafter. MATERIALS AND METHODS: Detection of RXR-alpha was performed on tumor specimens from 63 patients with primary RCC, using immunohistochemical techniques. For our evaluation of the immunostaining results, we developed a new cell-counting system based on the subcellular distribution of immunoreactivity. The impact of the subcellular distribution of RXR-alpha on the prognosis of patients with RCC was analyzed statistically among other clinicopathologic factors. The primary end point was survival. RESULTS: In 34 RCC samples (54.0%), RXR-alpha was detected predominantly in the nucleus, while 25 RCC specimens (39.7%) displayed an aberrant subcellular distribution pattern, with a predominantly cytoplasmic staining with nuclear sparing in 15 specimens (23.8%), and a combined nuclear and cytoplasmic staining in 10 specimens (15.9%). Very faint to undetectable immunoreactivity was noted in 4 cases (6.3%) of RCC. Univariate Kaplan-Meier analysis showed that patients with a predominantly nuclear localization of RXR-alpha had a significantly prolonged survival after primary tumor diagnosis, when compared to patients with a predominantly aberrant subcellular distribution profile (p < 0.01). Furthermore, multivariate analysis revealed that an aberrant subcellular distribution of RXR-alpha in RCC was an independent predictor of poor survival (p < 0.01). CONCLUSIONS: Our study indicated that the subcellular intratumoral distribution pattern of RXR-alpha could independently predict the survival of RCC patients. However, the exact mechanisms underlying the aberrant compartmentalization and the functions of RXR alpha in RCC remain to be determined. PMID- 15285880 TI - Effect of simultaneous administration of Avemar and cytostatic drugs on viability of cell cultures, growth of experimental tumors, and survival tumor-bearing mice. AB - Avemar (Biromedicina Co., Budapest, Hungary), a wheat germ preparation with immunomodulant and antimetastatic activity, was applied simultaneously with cytostatic drugs of different modes of action, in vitro and in vivo, in order to find out whether this simultaneous administration exerts an antagonistic or a synergistic effect on the viability of cell cultures, tumor growth, and survival of animals, inoculated with a transplantable mouse tumor (3LL-HH). In vitro, Avemar did not influence the effect on the viability of MCF-7, HepG2, or Vero cells, exerted by Dacarbazine, 5-fluorouracyl, or Adriblastina. In vivo, Avemar, combined with Endoxan, Navelbine, and doxorubicin, did not prevent the tumor growth inhibitory effect of the cytostatic drugs. The combination of Avemar with the cytostatic drugs did not increase the toxicity of the cytostatic compounds, and did not exert any toxic effect. Avemar may be administered together with cytostatic drugs, without the risk of increasing toxicity or decreasing antiproliferative activity. PMID- 15285881 TI - Repeated cycles with 72-hour continuous infusion interleukin-2 in kidney cancer and melanoma. AB - While high-dose bolus inpatient interleukin-2 is generally given on 8-week cycles, continuous infusion interleukin-2 could potentially allow for more rapidly repeated cycles. Fourteen (14) patients with an Eastern Cooperative Oncology Group (ECOG) performance status (PS) of 0 or 1, having either kidney cancer (6) or melanoma (8), have been treated with continuous infusion (CIV) interleukin-2 (IL-2) 18 MIU/m(2)/24 hours for 72 hours. Cycles were repeated every 3 weeks up to 4 cycles, then every 3-4 weeks for 2 cycles, then every 6-8 weeks, until progression or intolerable toxicity. All patients received famotidine 20 mg intravenously (i.v.) twice per day during the 72-hour infusions. Patient characteristics included a median ECOG performance status of 1; median age = 63 (range: 25-79); most common metastatic sites: lung (9), bone (5), lymph nodes (5), and the liver (3). No patients with metastatic kidney cancer underwent a nephrectomy prior to interleukin-2. Median number of cycles received = 5 (1-9). No patients required Intensive Care Unit (ICU) admission. There have been no treatment-related deaths. Most common toxicities have been rigors, fever, nausea/emesis, and the reversible elevation of creatinine. One complete response and three partial responses (67% response rate; 95% confidence interval: 30%-90%) have been seen in kidney cancer, and two partial responses (25% response rate; 95% confidence interval: 7%-60%) have occurred in melanoma. Median survival has not been reached at >9+ months. Responding sites include the liver, bone, lung, lymph node and subcutaneous sites. Inpatient 72-hour continuous infusion interleukin-2 at this dose and schedule is well tolerated by patients with an ECOG performance status of 0 or 1 and has activity in kidney cancer and melanoma. PMID- 15285882 TI - Introduction to kidney dose-response for radionuclide therapy. PMID- 15285883 TI - Clinical aspects of radiation nephropathy. AB - Small radiolabeled molecules are finding increasing clinical use for targeted radionuclide therapy. With the administration of radiolabeled small molecules, the bone marrow is not necessarily the first organ to show radiation toxicity. Rapid excretion of radioactivity through the urinary tract and the retention of radiolabeled small-protein molecules in the kidneys may expose the kidneys to radiation sufficient enough to cause toxicity--and in clinical trials, radiation toxicity of the urinary tract has become clinically relevant. The cells of the kidneys are slowly repairing cells; thus, the radiation toxicity may not be manifest for several months. The clinical and pathological features associated with radiation nephropathy, and issues particular to radiation nephropathy following targeted radionuclide therapy, are described here. PMID- 15285884 TI - Use of the linear-quadratic radiobiological model for quantifying kidney response in targeted radiotherapy. AB - This paper reviews the generalized application of the linear quadratic (LQ) model of radiobiological effect to targeted radiotherapy. Special attention is given to formulations for normal tissue responses and these are applied, in particular, to the kidney. Because it is derived from self-consistent bio-physical principles, the LQ model currently remains the standard formalism for assessing biological responses for the whole range of radiotherapy treatments. A central feature of the model is the derivation of biologically effective doses (BEDs), which may be used to quantify the impact of a treatment on both tumors and normal tissues. BEDs are routinely derived for conventional external-beam treatments. The likely limits of targeted radiotherapy may, thus, be assessed by comparing the expected normal-tissue BEDs for such treatments with those known to be just tolerable in conventional therapy. The main parameters required in the model are defined, and data are provided which demonstrate the tentative link between targeted radiotherapy doses and those used in conventional radiotherapy. The extension of the LQ method to targeted radiotherapy involves using parameters for which the numerical values may not be accurately known at present. This places a restriction on the overall predictive accuracy of the model and the necessary caveats are, therefore, outlined. PMID- 15285885 TI - Nonuniform absorbed dose distribution in the kidney: the influence of organ architecture. AB - The development of novel, systemically administered radionuclide therapies (such as radioimmunotherapy) relies on the ability to predict dose-limiting toxicity to normal tissue. Where the kidney is the principal route of excretion of the radionuclide preparation and/or breakdown of products, nephrotoxicity may be the dose-limiting factor. Until recently, conventional (MIRD) dosimetry assumed the distribution in the kidney to be uniform. A new MIRD phantom of the kidney models it as a set of uniform suborgans. In the work described here, the assumption of uniformity of distribution and of heterogeneity of dose rate (and, thus, absorbed dose) was tested in the mouse model. In this paper, we examine the nonuniformity of distribution and the subsequent dose rate for 4 antibody preparations (IgG (150 kD), F(ab)'(2) (100 kD), Fab (50 kD) and sFv (27 kD)) labeled with 4 radionuclides ((125)I, (131)I, (186)Re, and (90)Y) of interest in radioimmunotherapy (RIT). The kidney was considered as a whole and as two suborgans (cortex and medulla), and the nonuniformity of the dose-rate distribution was measured by a correlation of modeled dose-rate distribution with the dose-rate distribution obtained for an equivalent uniform radionuclide distribution. The heterogeneity of distribution, the inter- and intra-suborgan, was seen to increase as the molecular weight of the antibodies decreased. The assumption of uniform activity distribution for the whole kidney gives a poor estimation of the distribution of the dose rate. In the cortex, the longer-range emitters smooth out the effect of heterogeneous distribution and, in mice, an assumption of uniform cortex self-dose distribution may be sufficient for simple calculations. It is unclear how much this smoothing would be relevant in the human kidney. PMID- 15285886 TI - Relevance of external beam dose-response relationships to kidney toxicity associated with radionuclide therapy. AB - The importance of the kidney as a dose-limiting organ is likely to increase as smaller molecular vectors and radiometals become more commonly used in targeted radionuclide therapy. Data derived from kidney irradiation by external-beam therapy (XRT) indicate that the kidney is radiosensitive. The features of radiation nephropathy seen post-treatment appear similar between local XRT, total body irradiation (TBI), and radionuclide therapy. For uniform kidney irradiation, tolerance doses appear to be approximately 15-17 Gy in 2 Gy fractions for local XRT and probably less than this (approximately 12 Gy in 2 Gy fractions) when radiation is delivered systemically as TBI in the context of bone marrow transplant protocols. Animal studies indicate that the linear quadratic (LQ) model with an alpha/beta parameter of 1.5-3 Gy seems to adequately describe the XRT fractionation sensitivity of kidney for doses per fraction down to approximately 1 Gy, but may underestimate the effectiveness of fraction sizes less than this. Animal studies have also clarified the dose-dependency of the time to expression of radiation nephropathy and have indicated that radiation nephropathy may be alleviated by pharmacological means. PMID- 15285887 TI - Summary and perspectives on kidney dose-response to radionuclide therapy. PMID- 15285889 TI - Extreme value distribution based gene selection criteria for discriminant microarray data analysis using logistic regression. AB - One important issue commonly encountered in the analysis of microarray data is to decide which and how many genes should be selected for further studies. For discriminant microarray data analyses based on statistical models, such as the logistic regression models, gene selection can be accomplished by a comparison of the maximum likelihood of the model given the real data, L(D|M), and the expected maximum likelihood of the model given an ensemble of surrogate data with randomly permuted label, L(D(0)|M). Typically, the computational burden for obtaining L(D(0)M) is immense, often exceeding the limits of available computing resources by orders of magnitude. Here, we propose an approach that circumvents such heavy computations by mapping the simulation problem to an extreme-value problem. We present the derivation of an asymptotic distribution of the extreme-value as well as its mean, median, and variance. Using this distribution, we propose two gene selection criteria, and we apply them to two microarray datasets and three classification tasks for illustration. PMID- 15285890 TI - Joint classifier and feature optimization for comprehensive cancer diagnosis using gene expression data. AB - Recent research has demonstrated quite convincingly that accurate cancer diagnosis can be achieved by constructing classifiers that are designed to compare the gene expression profile of a tissue of unknown cancer status to a database of stored expression profiles from tissues of known cancer status. This paper introduces the JCFO, a novel algorithm that uses a sparse Bayesian approach to jointly identify both the optimal nonlinear classifier for diagnosis and the optimal set of genes on which to base that diagnosis. We show that the diagnostic classification accuracy of the proposed algorithm is superior to a number of current state-of-the-art methods in a full leave-one-out cross-validation study of five widely used benchmark datasets. In addition to its superior classification accuracy, the algorithm is designed to automatically identify a small subset of genes (typically around twenty in our experiments) that are capable of providing complete discriminatory information for diagnosis. Focusing attention on a small subset of genes is useful not only because it produces a classifier with good generalization capacity, but also because this set of genes may provide insights into the mechanisms responsible for the disease itself. A number of the genes identified by the JCFO in our experiments are already in use as clinical markers for cancer diagnosis; some of the remaining genes may be excellent candidates for further clinical investigation. If it is possible to identify a small set of genes that is indeed capable of providing complete discrimination, inexpensive diagnostic assays might be widely deployable in clinical settings. PMID- 15285891 TI - Physical network models. AB - We develop a new framework for inferring models of transcriptional regulation. The models, which we call physical network models, are annotated molecular interaction graphs. The attributes in the model correspond to verifiable properties of the underlying biological system such as the existence of protein protein and protein-DNA interactions, the directionality of signal transduction in protein-protein interactions, as well as signs of the immediate effects of these interactions. Possible configurations of these variables are constrained by the available data sources. Some of the data sources, such as factor-binding data, involve measurements that are directly tied to the variables in the model. Other sources, such as gene knock-outs, are functional in nature and provide only indirect evidence about the variables. We associate each observed knock-out effect in the deletion mutant data with a set of causal paths (molecular cascades) that could in principle explain the effect, resulting in aggregate constraints about the physical variables in the model. The most likely settings of all the variables, specifying the most likely graph annotations, are found by a recursive application of the max-product algorithm. By testing our approach on datasets related to the pheromone response pathway in S. cerevisiae, we demonstrate that the resulting model is consistent with previous studies about the pathway. Moreover, we successfully predict gene knock-out effects with a high degree of accuracy in a cross-validation setting. When applying this approach genome-wide, we extract submodels consistent with previous studies. The approach can be readily extended to other data sources or to facilitate automated experimental design. PMID- 15285892 TI - Optimizing exact genetic linkage computations. AB - Genetic linkage analysis is a challenging application which requires Bayesian networks consisting of thousands of vertices. Consequently, computing the probability of data, which is needed for learning linkage parameters, using exact computation procedures calls for an extremely efficient implementation that carefully optimizes the order of conditioning and summation operations. In this paper, we present the use of stochastic greedy algorithms for optimizing this order. Our algorithm has been incorporated into the newest version of SUPERLINK, which is a fast genetic linkage program for exact likelihood computations in general pedigrees. We demonstrate an order of magnitude improvement in run times of likelihood computations using our new optimization algorithm and hence enlarge the class of problems that can be handled effectively by exact computations. PMID- 15285893 TI - A polynomial-time nuclear vector replacement algorithm for automated NMR resonance assignments. AB - High-throughput NMR structural biology can play an important role in structural genomics. We report an automated procedure for high-throughput NMR resonance assignment for a protein of known structure, or of a homologous structure. These assignments are a prerequisite for probing protein-protein interactions, protein ligand binding, and dynamics by NMR. Assignments are also the starting point for structure determination and refinement. A new algorithm, called Nuclear Vector Replacement (NVR) is introduced to compute assignments that optimally correlate experimentally measured NH residual dipolar couplings (RDCs) to a given a priori whole-protein 3D structural model. The algorithm requires only uniform( 15)N labeling of the protein and processes unassigned H(N)-(15)N HSQC spectra, H(N) (15)N RDCs, and sparse H(N)-H(N) NOE's (d(NN)s), all of which can be acquired in a fraction of the time needed to record the traditional suite of experiments used to perform resonance assignments. NVR runs in minutes and efficiently assigns the (H(N),(15)N) backbone resonances as well as the d(NN)s of the 3D (15)N-NOESY spectrum, in O(n(3)) time. The algorithm is demonstrated on NMR data from a 76 residue protein, human ubiquitin, matched to four structures, including one mutant (homolog), determined either by x-ray crystallography or by different NMR experiments (without RDCs). NVR achieves an assignment accuracy of 92-100%. We further demonstrate the feasibility of our algorithm for different and larger proteins, using NMR data for hen lysozyme (129 residues, 97-100% accuracy) and streptococcal protein G (56 residues, 100% accuracy), matched to a variety of 3D structural models. Finally, we extend NVR to a second application, 3D structural homology detection, and demonstrate that NVR is able to identify structural homologies between proteins with remote amino acid sequences using a database of structural models. PMID- 15285894 TI - Approximation of protein structure for fast similarity measures. AB - The structural comparison of two proteins comes up in many applications in structural biology where it is often necessary to find similarities in very large conformation sets. This work describes techniques to achieve significant speedup in the computation of structural similarity between two given conformations, at the expense of introducing a small error in the similarity measure. Furthermore, the proposed computational scheme allows for a tradeoff between speedup and error. This scheme exploits the fact that the Calpha representation of a protein conformation contains redundant information, due to the chain topology and limited compactness of proteins. This redundancy can be reduced by approximating subchains of a protein by their centers of mass, resulting in a smaller number of points to describe a conformation. A Haar wavelet analysis of random chains and proteins is used to justify this approximated representation. Similarity measures computed with this representation are highly correlated to the measures computed with the original Calpha representation. Therefore, they can be used in applications where small similarity errors can be tolerated or as fast filters in applications that require exact measures. Computational tests have been conducted on two applications, nearest neighbor search and automatic structural classification. PMID- 15285895 TI - Methods in comparative genomics: genome correspondence, gene identification and regulatory motif discovery. AB - In Kellis et al. (2003), we reported the genome sequences of S. paradoxus, S. mikatae, and S. bayanus and compared these three yeast species to their close relative, S. cerevisiae. Genomewide comparative analysis allowed the identification of functionally important sequences, both coding and noncoding. In this companion paper we describe the mathematical and algorithmic results underpinning the analysis of these genomes. (1) We present methods for the automatic determination of genome correspondence. The algorithms enabled the automatic identification of orthologs for more than 90% of genes and intergenic regions across the four species despite the large number of duplicated genes in the yeast genome. The remaining ambiguities in the gene correspondence revealed recent gene family expansions in regions of rapid genomic change. (2) We present methods for the identification of protein-coding genes based on their patterns of nucleotide conservation across related species. We observed the pressure to conserve the reading frame of functional proteins and developed a test for gene identification with high sensitivity and specificity. We used this test to revisit the genome of S. cerevisiae, reducing the overall gene count by 500 genes (10% of previously annotated genes) and refining the gene structure of hundreds of genes. (3) We present novel methods for the systematic de novo identification of regulatory motifs. The methods do not rely on previous knowledge of gene function and in that way differ from the current literature on computational motif discovery. Based on genomewide conservation patterns of known motifs, we developed three conservation criteria that we used to discover novel motifs. We used an enumeration approach to select strongly conserved motif cores, which we extended and collapsed into a small number of candidate regulatory motifs. These include most previously known regulatory motifs as well as several noteworthy novel motifs. The majority of discovered motifs are enriched in functionally related genes, allowing us to infer a candidate function for novel motifs. Our results demonstrate the power of comparative genomics to further our understanding of any species. Our methods are validated by the extensive experimental knowledge in yeast and will be invaluable in the study of complex genomes like that of the human. PMID- 15285896 TI - Multilevel modeling and inference of transcription regulation. AB - The understanding of transcription regulation is a major goal of today's biology. The challenge is to utilize diverse high-throughput data in order to infer mechanistic models of transcription control. We propose a new model which integrates transcription factor-gene affinity, protein abundance, and gene expression profiles. The model provides a detailed, yet computationally tractable description of the relations between transcription factors, their binding sites at gene promoters, and the combinatorial regulation of transcription. At the core, our model manipulates dose-affinity-response functions that associate transcription factor concentrations and transcription factor-DNA affinities to determine the rate of transcription factor-DNA reactions. We study computational problems that arise in optimizing such models and develop polynomial algorithms for certain problems. We show how to assess missing values (notably protein abundance) and describe a novel framework to infer models from currently available data. On budding yeast carbohydrate metabolism data, our results demonstrate the sensitivity and specificity of the approach. They also suggest new active binding sites and a regulation model for the transcription program of the galactose system. PMID- 15285897 TI - Maximum entropy modeling of short sequence motifs with applications to RNA splicing signals. AB - We propose a framework for modeling sequence motifs based on the maximum entropy principle (MEP). We recommend approximating short sequence motif distributions with the maximum entropy distribution (MED) consistent with low-order marginal constraints estimated from available data, which may include dependencies between nonadjacent as well as adjacent positions. Many maximum entropy models (MEMs) are specified by simply changing the set of constraints. Such models can be utilized to discriminate between signals and decoys. Classification performance using different MEMs gives insight into the relative importance of dependencies between different positions. We apply our framework to large datasets of RNA splicing signals. Our best models out-perform previous probabilistic models in the discrimination of human 5' (donor) and 3' (acceptor) splice sites from decoys. Finally, we discuss mechanistically motivated ways of comparing models. PMID- 15285898 TI - Score functions for determining regional conservation in two-species local alignments. AB - We construct several score functions for use in locating unusually conserved regions in a genomewide search of aligned DNA from two species. We test these functions on regions of the human genome aligned to the mouse genome. These score functions are derived from properties of neutrally evolving sites on the mouse and human genome and can be adjusted to the local background rate of conservation. The aim of these functions is to try to identify regions of the human genome that are conserved by evolutionary selection because they have an important function, rather than by chance. We use them to get a very rough estimate of the amount of DNA in the human genome that is under selection. PMID- 15285899 TI - Combining phylogenetic and hidden Markov models in biosequence analysis. AB - A few models have appeared in recent years that consider not only the way substitutions occur through evolutionary history at each site of a genome, but also the way the process changes from one site to the next. These models combine phylogenetic models of molecular evolution, which apply to individual sites, and hidden Markov models, which allow for changes from site to site. Besides improving the realism of ordinary phylogenetic models, they are potentially very powerful tools for inference and prediction--for example, for gene finding or prediction of secondary structure. In this paper, we review progress on combined phylogenetic and hidden Markov models and present some extensions to previous work. Our main result is a simple and efficient method for accommodating higher order states in the HMM, which allows for context-dependent models of substitution--that is, models that consider the effects of neighboring bases on the pattern of substitution. We present experimental results indicating that higher-order states, autocorrelated rates, and multiple functional categories all lead to significant improvements in the fit of a combined phylogenetic and hidden Markov model, with the effect of higher-order states being particularly pronounced. PMID- 15285901 TI - Efficient extraction of mapping rules of atoms from enzymatic reaction data. AB - Many computational problems and methods have been proposed for analysis of biological pathways. Among them, this paper focuses on extraction of mapping rules of atoms from enzymatic reaction data, which is useful for drug design, simulation of tracer experiments, and consistency checking of pathway databases. Most of existing methods for this problem are based on maximal common subgraph algorithms. In this paper, we propose a novel approach based on graph partition and graph isomorphism. We show that this problem is NP-hard in general, but can be solved in polynomial time for wide classes of enzymatic reactions. We also present an O(n(1.5)) time algorithm for a special but fundamental class of reactions, where n is the maximum size of compounds appearing in a reaction. We develop practical polynomial-time algorithms in which the Morgan algorithm is used for computing the normal form of a graph, where it is known that the Morgan algorithm works correctly for most chemical structures. Computational experiments are performed for these practical algorithms using the chemical reaction data stored in the KEGG/LIGAND database. The results of computational experiments suggest that practical algorithms are useful in many cases. PMID- 15285900 TI - Scalable heuristics for design of DNA probe arrays. AB - Design of DNA arrays for very large-scale immobilized polymer synthesis (VLSIPS) (Fodor et al., 1991) seeks to minimize effects of unintended illumination during mask exposure steps. Hannenhalli et al. (2002) formulate this requirement as the Border Minimization Problem and give an algorithm for placement of probes at array sites under the assumption that the array synthesis is synchronous; i.e., nucleotides are synthesized in a periodic sequence (ACGT)(k) and every probe grows by exactly one nucleotide with every group of four masks. Drawing on the analogy with VLSI placement, in this paper we describe and experimentally validate the engineering of several scalable, high-quality placement heuristics for both synchronous and asynchronous DNA array design. We give empirical results on both randomly generated and industry test cases confirming the scalability and improved solution quality enjoyed by our methods. In general, our techniques improve on state-of-the-art industrial results by over 4% and surpass academically published results by up to 35%. Finally, we give lower bounds that offer insights into the amount of available further improvements. PMID- 15285902 TI - An integrated probabilistic model for functional prediction of proteins. AB - We develop an integrated probabilistic model to combine protein physical interactions, genetic interactions, highly correlated gene expression networks, protein complex data, and domain structures of individual proteins to predict protein functions. The model is an extension of our previous model for protein function prediction based on Markovian random field theory. The model is flexible in that other protein pairwise relationship information and features of individual proteins can be easily incorporated. Two features distinguish the integrated approach from other available methods for protein function prediction. One is that the integrated approach uses all available sources of information with different weights for different sources of data. It is a global approach that takes the whole network into consideration. The second feature is that the posterior probability that a protein has the function of interest is assigned. The posterior probability indicates how confident we are about assigning the function to the protein. We apply our integrated approach to predict functions of yeast proteins based upon MIPS protein function classifications and upon the interaction networks based on MIPS physical and genetic interactions, gene expression profiles, tandem affinity purification (TAP) protein complex data, and protein domain information. We study the recall and precision of the integrated approach using different sources of information by the leave-one-out approach. In contrast to using MIPS physical interactions only, the integrated approach combining all of the information increases the recall from 57% to 87% when the precision is set at 57%-an increase of 30%. PMID- 15285903 TI - Towards optimally multiplexed applications of universal arrays. AB - We study a design and optimization problem that occurs, for example, when single nucleotide polymorphisms (SNPs) are to be genotyped using a universal DNA tag array. The problem of optimizing the universal array to avoid disruptive cross hybridization between universal components of the system was addressed in previous work. Cross-hybridization can, however, also occur assay specifically, due to unwanted complementarity involving assay-specific components. Here we examine the problem of identifying the most economic experimental configuration of the assay-specific components that avoids cross-hybridization. Our formalization translates this problem into the problem of covering the vertices of one side of a bipartite graph by a minimum number of balanced subgraphs of maximum degree 1. We show that the general problem is NP-complete. However, in the real biological setting, the vertices that need to be covered have degrees bounded by d. We exploit this restriction and develop an O(d)-approximation algorithm for the problem. We also give an O(d)-approximation for a variant of the problem in which the covering subgraphs are required to be vertex disjoint. In addition, we propose a stochastic model for the input data and use it to prove a lower bound on the cover size. We complement our theoretical analysis by implementing two heuristic approaches and testing their performance on synthetic data as well as on simulated SNP data. PMID- 15285904 TI - Model-based inference of haplotype block variation. AB - The haplotype block structure of SNP variation in human DNA has been demonstrated by several recent studies. The presence of haplotype blocks can be used to dramatically increase the statistical power of genetic mapping. Several criteria have already been proposed for identifying these blocks, all of which require haplotypes as input. We propose a comprehensive statistical model of haplotype block variation and show how the parameters of this model can be learned from haplotypes and/or unphased genotype data. Using real-world SNP data, we demonstrate that our approach can be used to resolve genotypes into their constituent haplotypes with greater accuracy than previously known methods. PMID- 15285905 TI - Haplotype reconstruction from SNP alignment. AB - In this paper, we describe a method for statistical reconstruction of haplotypes from a set of aligned SNP fragments. We consider the case of a pair of homologous human chromosomes, one from the mother and the other from the father. After fragment assembly, we wish to reconstruct the two haplotypes of the parents. Given a set of potential SNP sites inferred from the assembly alignment, we wish to divide the fragment set into two subsets, each of which represents one chromosome. Our method is based on a statistical model of sequencing errors, compositional information, and haplotype memberships. We calculate probabilities of different haplotypes conditional on the alignment. Due to computational complexity, we first determine phases for neighboring SNPs. Then we connect them and construct haplotype segments. Also, we compute the accuracy or confidence of the reconstructed haplotypes. We discuss other issues, such as alternative methods, parameter estimation, computational efficiency, and relaxation of assumptions. PMID- 15285906 TI - An interview with James Wilbur, Ph.D. General Manager, Life Sciences, Meso Scale Discovery. AB - James L. Wilbur, Ph.D. received a Bachelor's degree from the University of California, San Diego and a Ph.D. in Chemistry from Stanford University. After completing an NIH Postdoctoral Fellowship with Professor George M. Whitesides in the Department of Chemistry at Harvard University, he joined IGEN International, Inc., where he held a variety of positions in Research and Development. During that time, he was part of the team that developed the core technology and products for Meso Scale Discovery. He assumed his current position in 2001 when Meso Scale Discovery launched the products discussed here. PMID- 15285907 TI - Functional assay of voltage-gated sodium channels using membrane potential sensitive dyes. AB - The discovery of novel therapeutic agents that act on voltage-gated sodium channels requires the establishment of high-capacity screening assays that can reliably measure the activity of these proteins. Fluorescence resonance energy transfer (FRET) technology using membrane potential-sensitive dyes has been shown to provide a readout of voltage-gated sodium channel activity in stably transfected cell lines. Due to the inherent rapid inactivation of sodium channels, these assays require the presence of a channel activator to prolong channel opening. Because sodium channel activators and test compounds may share related binding sites on the protein, the assay protocol is critical for the proper identification of channel inhibitors. In this study, high throughput, functional assays for the voltage-gated sodium channels, hNa(V)1.5 and hNa(V)1.7, are described. In these assays, channels stably expressed in HEK cells are preincubated with test compound in physiological medium and then exposed to a sodium channel activator that slows channel inactivation. Sodium ion movement through open channels causes membrane depolarization that can be measured with a FRET dye membrane potential-sensing system, providing a large and reproducible signal. Unlike previous assays, the signal obtained in the agonist initiation assay is sensitive to all sodium channel modulators that were tested and can be used in high throughput mode, as well as in support of Medicinal Chemistry efforts for lead optimization. PMID- 15285908 TI - Characterization of CHO cells stably expressing a G alpha 16/z chimera for high throughput screening of GPCRs. AB - G protein-coupled receptors (GPCRs) are important therapeutic targets for drug discovery. The identification and characterization of new ligands ideally requires the use of high throughput assays that are applicable to all GPCR subtypes. To circumvent the problem of different GPCRs coupling to distinct intracellular second messenger pathways, we describe a new method that uses the chimeric Galpha protein 16z25 to facilitate this process. Stably expressed in Chinese hamster ovary cells, 16z25 allows G(i/o)- and G(s)-coupled receptors to mobilize intracellular Ca(2+) upon agonist stimulation. We have generated nine cell lines each stably expressing 16z25 and a GPCR. All cell lines respond to appropriate agonist stimulation in fluorometric imaging plate reader (FLIPR) assays with robust and potent Ca(2+) mobilization. Several of these lines have been pharmacologically characterized using agonists and antagonists. We also demonstrate that the coexpression of GPCR and 16z25 does not interfere with the receptors' ability to activate endogenous signaling pathways. The ability of 16z25 to functionally mediate the agonist stimulation of a broad spectrum of GPCRs indicates that the use of cell lines stably coexpressing this chimera and GPCRs will simplify the drug screening process and aid in the deorphanization of new receptors. PMID- 15285909 TI - Development of a sensitive and HTS-compatible reporter gene assay for functional analysis of human adenosine A2a receptors in CHO-K1 cells. AB - Adenosine A2a receptor, a member of the G protein-coupled receptor superfamily, has been demonstrated to be an important pharmacological target. It couples to stimulatory G protein and activates adenylate cyclase upon agonist stimulation. Here we attempted to stably transfect Chinese hamster ovary (CHO-K1) cells, which lack any known subtypes of adenosine receptors, with recombinant human adenosine A2a receptors (hA2aR). Rapid down-regulation of hA2aR in a clonal cell line, CHOA2a-2, was observed over a short period of time in culture. This is consistent with other groups' findings of low expression and poor G protein coupling of this receptor in several cell systems. To facilitate pharmacological profiling for hA2aR ligand, we introduced a cyclic AMP response element (CRE)-linked beta galactosidase reporter gene into CHOA2a-2 cells to generate a stable cell line, CHOA2a-2CREbetagal#26. Robust cyclic AMP signal amplification was obtained using a colorimetric assay measuring beta-galactosidase activity. The EC(50) of 5'-N ethylcarboxamidoadenosine (NECA), a potent A2a agonist, for inducing beta galactosidase activity was 23.3 +/- 3.5 nM, similar to 22.7 +/- 3.9 nM, which was the NECA EC(50) in the direct measurement of cyclic AMP of CHOA2a-2 cells in early culture. Subsequently we validated this assay for high throughput screening for hA2aR agonists. The Z' factor for robotic assay performance was 0.79 +/- 0.03, the ratio of signal/noise was 157 +/- 36, and the ratio of signal/background was 10.6 +/- 1.2, demonstrating that this assay is well suitable for quality high throughput screening. High throughput screening of Johnson & Johnson libraries uncovered a couple of distinct series of nonadenosine small molecules, in addition to adenosine analogues, as potential hA2aR agonists with EC(50) values of 2-6 microM. Preliminary characterization of those compounds was presented. PMID- 15285910 TI - WGA-coated yttrium oxide beads enable an imaging-based adenosine 2a receptor binding scintillation proximity assay suitable for high throughput screening. AB - Adenosine receptors belong to the superfamily of G protein-coupled receptors and are involved in a variety of physiologic functions. Traditionally, binding assays to detect adenosine 2a (A2a) antagonists and agonists have used filtration methods that are cumbersome to run and not amenable to HTS. We developed scintillation proximity assays (SPA trade mark ) utilizing HEK293 RBHA2AM cell membranes, either wheat germ agglutinin (WGA)-coated yttrium silicate (YSi) or red-shifted yttrium oxide (YO) beads and the A2a-selective radioligand [(3)H]SCH 58261. Both beads gave windows (total binding/nonspecific binding) of >5 and K(d) values of 2-3 nM for the radioligand, in agreement with results obtained by filtration. In contrast, WGA-polyvinyltoluene as well as other bead types had windows of <3 and significant radioligand binding to the uncoated beads. A 384 well WGA-YO bead SPA was optimized utilizing a LEADseeker imaging system and an automated trituration process for dispensing the dense yttrium-based beads. Signals were stable after 4 h, and Z' values were 0.7-0.8. The LEADseeker imaging assay tolerated 2% dimethyl sulfoxide and generated IC(50) values of 3-5 nM for the A2a antagonist CGS 15943, comparable to that obtained by the filtration method. A number of adenosine and xanthine analogues were identified as hits in the Library of Pharmacologically Active Compounds (LOPAC). This imaging-based A2a SPA enables HTS and is a major improvement over the filtration method. PMID- 15285911 TI - A homogeneous scintillation proximity format for monitoring the activity of recombinant human long-chain-fatty-acyl-CoA synthetase 5. AB - Fatty acyl coenzyme A (CoA) synthetases are a group of enzymes responsible for the activation of fatty acids through ligated high-energy CoA thioester bonds. Ultimately these fatty acyl-CoA conjugates are routed toward either anabolic or catabolic pathways. Long-chain-fatty-acid-CoA ligase 5 (LACS 5) utilizes a wide range of saturated fatty acids with a substrate preference for C16-C18 unsaturated fatty acids. This enzyme represents a new class of potential drug targets, and, hence, our efforts were focused upon developing a robust assay for utilization in a high throughput screen. Toward that end, we describe a radiometric homogeneous measurement of the enzymatic reaction by employing ionic capture of the reaction product onto YSi scintillation proximity assay (SPA) beads. We present kinetic and inhibition data for LACS 5 using this SPA format. Our results show that the assay method is both robust and well suited for this class of lipid-metabolizing enzymes. PMID- 15285912 TI - Mechanized syringe homogenization of human and animal tissues. AB - Tissue homogenization is a prerequisite to any fractionation schedule. A plethora of hands-on methods are available to homogenize tissues. Here we report a mechanized method for homogenizing animal and human tissues rapidly and easily. The Bio-Mixer 1200 (manufactured by Innovative Products, Inc., Oklahoma City, OK) utilizes the back-and-forth movement of two motor-driven disposable syringes, connected to each other through a three-way stopcock, to homogenize animal or human tissue. Using this method, we were able to homogenize human or mouse tissues (brain, liver, heart, and salivary glands) in 5 min. From sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis and a matrix-assisted laser desorption/ionization time-of-flight mass spectrometric enzyme assay for prolidase, we have found that the homogenates obtained were as good or even better than that obtained used a manual glass-on-Teflon (DuPont, Wilmington, DE) homogenization protocol (all-glass tube and Teflon pestle). Use of the Bio-Mixer 1200 to homogenize animal or human tissue precludes the need to stay in the cold room as is the case with the other hands-on homogenization methods available, in addition to freeing up time for other experiments. PMID- 15285913 TI - Patent prosecution in structural proteomics. AB - This paper presents a brief overview of intellectual property rights and the various areas in proteomics to which intellectual property rights may be applicable. Technology transfer, including licensing and business agreements, are not covered in this paper. Instead, issues and complications related to national and overseas patent prosecution in this relatively new field will be discussed. PMID- 15285921 TI - Cyclooxygenase-2 and gastrointestinal cancer. AB - The cyclooxygenase (COX) enzymes (COX-1 and COX-2) are key enzymes of prostaglandin (PG) biosynthesis. Nonselective non-steroidal anti-inflammatory drugs (NSAIDs) inhibit the enzymatic activity of both COX-1 and COX-2. Selective COX-2 inhibitors have been developed that appear to have 50% less gastrointestinal toxicity than traditional nonselective NSAIDs. Experimental evidence suggests that the COX pathway is involved in tumor promotion. Evidence to support this comes from both clinical and laboratory findings suggesting that chronic NSAID use reduces the relative risk for developing colorectal cancer (CRC). Although the precise mechanism or mechanisms by which these drugs affect tumor progression is not completely understood, it is likely that part of their anti-tumor effect is due to inhibition of the COX- 2 enzyme. COX-2 levels are increased in CRC as well as in several other solid malignancies. COX-2-derived bioactive lipid products promote tumor-associated n eovascularization, inhibit cell death, and stimulate cell proliferation and motility. Additionally, treatment with COX-2-selective inhibitors reduces polyp burden in animal models of intestinal neoplasia and in humans with familial adenomatous polyposis (FAP). Ongoing human clinical trails are under way to test the efficacy of COX-2 selective inhibitors in a number of human cancers. PMID- 15285922 TI - Progression rates of colorectal cancer in high-risk individuals. PMID- 15285923 TI - Iodine versus palladium for prostate brachytherapy: the controversy continues. PMID- 15285924 TI - Treatment margins and prostate brachytherapy: "size matters," but how much and where? PMID- 15285925 TI - Progression rates of colorectal cancer by Dukes' stage in a high-risk group: analysis of selective colorectal cancer screening. AB - PURPOSE: The progression rates of colorectal cancer by Dukes' stage in a high risk group were estimated and applied to evaluate the efficacy of different screening regimens. PATIENTS AND METHODS: Of 6303 high-risk subjects invited to a colorectal cancer screening project with colonoscopy, 39 screen-detected cases and 16 postscreening cases were diagnosed with information available on Dukes' stage. A five-state Markov process was applied to estimate parameters pertaining to the disease natural history of colorectal cancer by Dukes' stage. RESULTS: The estimates of the mean sojourn time in years were 3.10 for preclinical Dukes' A and B and 1.92 for preclinical Dukes' stages C and D. The predicted reductions of Dukes' stages C and D achieved by annual, biennial, 3-yearly, and 6-yearly screening regimens against the control group were 60%, 49%, 40%, and 25%, respectively. These, in turn, yield the corresponding predicted mortality reductions of 39%, 33%, 28%, and 18%. CONCLUSIONS: These findings suggest that to achieve a 30% mortality reduction, as observed in annual fecal occult blood testing, a prudent interscreening interval with colonoscopy for this high-risk group should not be longer than 3 years. PMID- 15285926 TI - Iodine 125 versus palladium 103 implants for prostate cancer: clinical outcomes and complications. AB - PURPOSE: The purpose of this study was to evaluate the clinical outcomes and compare complication rates for patients with prostate cancer treated with iodine 125 ((125)I) and palladium 103 ((103)Pd) prostate brachytherapy at a single institution. PATIENTS AND METHODS: Between 1992 and 2002, 272 patients with prostate cancer were treated with ultrasound-guided transperineal implantation incorporating (125)I (107 patients) or (103)Pd (165 patients). Three months of hormonal therapy was incorporated into the treatment program in 33% of the patients in both groups. Nineteen percent of those treated with (125)I were treated with a combination of implantation plus external-beam radiation therapy. Only 6% of the group receiving (103)Pd implants were treated with such a combination. For those treated with (125)I implantation alone, the minimum tumor dose was 145 Gy. The minimum tumor dose for those treated with (103)Pd alone was 125 Gy. Those treated with a combination of external-beam radiation therapy and (125)I received 45 Gy via 1.8-Gy fractions followed by implantation with a minimum tumor dose of 110 Gy. For those treated with external-beam radiation therapy and (103)Pd, the doses were 45 Gy via 1.8-Gy fractions followed by implantation with minimum tumor dose of 98 Gy. Outcomes were evaluated based on radionuclide used, T stage, Gleason score, prostate-specific antigen, and prognostic group. Complications were also evaluated for each radionuclide. The mean follow-up for the (125)I group was 55 months, and the range was 12-108 months. The mean follow-up for the (103)Pd group was 44 months, and the range was 12-72 months. RESULTS: The 5-year biochemical disease-free survival rates for those in the favorable group (clinical stage T1c or T2, prostate-specific antigen level <10, Gleason score <7) were 92% for the (125)I group and 92% for the patients treated with (103)Pd. The 5-year disease-free survival rates for those in the intermediate and poor prognostic groups, which were combined, was 72% and 74%, respectively, for (125)I and (103)Pd. There was no statistically significant difference for either modality for any treatment group tested. In those treated with implantation alone, patients treated with (125)I had higher complication rates than those treated with (103)Pd (15% vs 4%). (125)I-treated patients had a grade 2 complication rate of 8% and a grade 3-4 complication rate of 7%, compared with 3% and 1%, respectively, for the (103)Pd-treated patients. CONCLUSION: Despite the different management recommendations that evolved during the study period, the clinical outcome for patients treated with either radionuclide were similar with respect to biochemical disease-free survival. Although specific dosimetric comparisons are not valid given differences in imaging over the study course, the complication rate appears to be somewhat higher for (125)I, which is consistent with a radiobiologic model. PMID- 15285927 TI - Treatment margins predict biochemical outcomes after prostate brachytherapy. AB - PURPOSE: Due to the theoretical role of treatment margins (TMs) in cancer, we have correlated biochemical outcomes with post-implant TMs in patients treated with brachytherapy for early stage prostate cancer. METHODS: From November 1998 through September 2003, 492 of a planned total of 600 patients with 1997 AJC clinical stage T1c-T2a prostatic carcinoma (Gleason score 5 or 6, PSA 4 to 10 ng/mL) have been randomized to implantation with (125)I (144 Gy, TG-43) versus (103)Pd (125 Gy, NIST-99). This preliminary analysis included only the first 122 analyzable patients, while accrual to the trial finishes. Isotope implantation was performed by standard techniques, using a modified peripheral loading pattern. Axial CT images at 3 mm intervals were acquired within four hours postoperatively for post-implant dosimetry. The contoured images and sources were entered into Varian Variseed system 7.1 (Charlottesville, VA). After completion of standard dosimetric calculations, the 100% prescription dose TMs were measured and tabulated around the prostate periphery at the 0.0, 1.0, 2.0 and 3.0 cm planes, going distal from the bladder-prostate interface. Measurements were limited to the transverse planes. Freedom from biochemical failure was defined as a serum PSA < or = 0.5 ng/mL at last follow-up. Patients were censored at last follow-up if their serum PSA was still decreasing. Patients whose serum PSA nadired at a value >0.5 ng/mL were scored as failures at the time at which their PSA nadired. The follow-up period for non-failing patients ranged from 2.1-5.0 years (median: 3.3 years). RESULTS: The average 100% prescription dose treatment margin (for individual patients) ranged from -5.0 to 8.7 mm, with an overall average of 2.6 mm (+/-3.1). In univariate analysis, the D(90) was the best predictor of biochemical control for (125)I, while the average TM was the best predictor for (103)Pd. Similarly, in multivariate analysis using the D(90), V(100), and average TM as the independent variables and biochemical control as the dependent variable, the D(90) was most closely related to biochemical control for (125)I patients, while average TM was most closely related for (103)Pd patients. In separate analysis of TM by site, the anterior TMs were the best predictors of biochemical outcomes. CONCLUSION: V(100), D(90), and TMs all appear to have a bearing on biochemical freedom from relapse after prostate brachytherapy. Efforts to better identify and test geographic dosimetric parameters are theoretically appealing, and supported by the clinical data summarized here. PMID- 15285928 TI - The impact of prostate volume and neoadjuvant androgen-deprivation therapy on urinary function following prostate brachytherapy. AB - PURPOSE: The purpose of this article is to evaluate the impact of prostate size and the magnitude of cytoreduction after neoadjuvant androgen-deprivation therapy (ADT) on catheter dependency, urinary symptomatology, and need for postbrachytherapy surgical intervention. MATERIALS AND METHODS: From February 1998 to August 2002, 186 consecutive patients under went monotherapeutic brachytherapy (no supplemental external-beam radiotherapy or ADT), and 101 consecutive patients received < or = 6 months of ADT (a luteinizing hormone releasing hormone agonist and an anti-androgen) in conjunction with brachytherapy without supplemental external-beam radiotherapy for clinical Tlc-T2b (2002 American Joint Committee on Cancer) prostate cancer. ADT was initiated approximately 3 months before brachytherapy. The median follow-up was 38.6 months. An alpha-blocker was initiated before implantation and continued at least until the International Prostate Symptom Score (IPSS) returned to baseline levels. Evaluated parameters included patient age, pretreatment prostate-specific antigen, Gleason score, clinical T stage, preimplantation IPSS, ultrasound volume, hormonal status, isotope, D(90), V(100/150/200), and urethral dose (average and maximum). RESULTS: Patients receiving neoadjuvant ADT were statistically older, presented with higher preimplantation IPSS scores, and larger prostate volumes. Patients receiving ADT were likelier to require a urinary catheter for the first 3 days after implantation; however, by day 4, no statistical difference in catheter dependency could be discerned between the two cohorts. Hormonal status did not predict for postbrachytherapy surgical intervention. IPSS returned to baseline at a mean of 1.8 and 1.7 months in hormone-naive and ADT patients, respectively. In multivariate Cox regression analysis, the preimplantation IPSS and the maximum postimplantation IPSS predicted for IPSS normalization overall and in both cohorts. Ultrasound prostate volume did not predict for IPSS normalization, catheter dependency, or need for postimplantation surgical intervention. CONCLUSION: Although patients receiving ADT were likelier to require a urinary catheter for the first three days after implantation, hormonal manipulation did not affect IPSS normalization, prolonged catheter dependency, or need for postbrachytherapy surgical intervention in these patients treated with brachytherapy without supplemental external-beam radiotherapy. PMID- 15285929 TI - Eicosapentaenoic acid induces Fas-mediated apoptosis through a p53-dependent pathway in hepatoma cells. AB - Eicosapentaenoic acid (EPA) has been demonstrated to induce apoptosis and cell cycle arrest in various cancer cell lines in vitro. In this study, we investigated the anti-tumor effects of EPA on hepatoma cell lines and the mechanisms responsible for induced cell death. Three hepatoma cell lines tested had different p53 status: HepG2 with a wild-type p53; Hep3B, of which the endogenous p53 was deleted; and Huh7 with its p53 mutated. MTT assay showed reduced viability of HepG2 cells after exposure to EPA, and the cytotoxicity of EPA was time and dose dependent. However, EPA had no effect on the viability and cell death in the two other hepatoma cell lines containing dysfunctional p53. DNA fragmentation analysis and TUNEL (terminal deoxynucleotidyl transferase [TdT] mediated deoxyuridine diphosphate [dUTP] nick end labeling) staining showed a typical pattern of DNA laddering and DNA breaks staining, respectively, in wild type p53-containing HepG2 cells after EPA treatment. We also observed that EPA induced transient nuclear accumulation of P53 protein that subsequently up regulated the expression of Fas messenger RNA and protein in HepG2 cells. In contrast, these findings were not observed in Hep3B and Huh7 cells exposed to EPA. Most notably, EPA-induced apoptosis in HepG2 cells could be reduced almost completely by treatment with FasL antisense oligonucleotides. We conclude that EPA inhibits the growth of HepG2 cells and mediates its effect, at least in part, via the Fas-mediated apoptosis. It appears that the effects of EPA on hepatoma cells are determined by the status of p53 and that wild-type p53 is a prerequisite for the anticancer effect of EPA. PMID- 15285955 TI - Acute and delayed posttraumatic stress disorders: a history and some issues. PMID- 15285956 TI - Gender differences in the prescribing of antipsychotic drugs. AB - OBJECTIVE: The aim of this article is to offer recommendations and rationale for gender-specific antipsychotic treatment. METHOD: The author summarizes reviews of recent literature in psychiatric clinical trials, pharmacology, drug safety, toxicology, obstetrics and gynecology, and pediatrics. RESULTS: The pharmacokinetics and pharmacodynamics of antipsychotic drugs differ in women and men and are influenced by gender-specific factors such as body build, diet, smoking, concurrent medication, exercise, substance use, and hormonal transitions. In general, and for some drugs in particular, women require lower doses in order to stay well. Because preliminary drug testing is not done in pregnant women, the issue of effective dosing during pregnancy is unstudied, and safety for fetuses and nursing infants may not become evident until a drug is widely used. Specific adverse effects on issues crucial to women (e.g., parenting) have not been well studied, but some side effects, such as weight gain, passivity, hypotension, and hyperprolactinemia, are reported to be particularly problematic for women. Some serious side effects are more often seen among women than among men. CONCLUSIONS: Optimal maintenance regimens of antipsychotics for women and men are not the same. PMID- 15285957 TI - Physical health monitoring of patients with schizophrenia. AB - OBJECTIVE: Schizophrenia is associated with several chronic physical illnesses and a shorter life expectancy, compared with life expectancy in the general population. One approach to improving the health of patients with schizophrenia is to improve the monitoring of physical health that occurs in psychiatric settings. The authors discuss a consensus panel's recommendations for improving the physical health monitoring of patients with schizophrenia who are treated in outpatient settings. METHOD: A consensus meeting including psychiatric and other medical experts assembled on October 17-18, 2002, to evaluate the existing literature and to develop recommendations for physical health monitoring of patients with schizophrenia. Conference participants reviewed the literature in the following areas: 1) weight gain and obesity; 2) diabetes; 3) hyperlipidemia; 4) prolongation of the QT interval on the ECG; 5) prolactin elevation and related sexual side effects; 6) extrapyramidal side effects, akathisia, and tardive dyskinesia; 7) cataracts; and 8) myocarditis. Experts for each topic area formulated monitoring recommendations that were discussed by all of the participants until a consensus was reached. RESULTS: Consensus recommendations included regular monitoring of body mass index, plasma glucose level, lipid profiles, and signs of prolactin elevation or sexual dysfunction. Information from monitoring should guide the selection of antipsychotic agents. Specific recommendations were made for cardiac monitoring of patients who receive medications associated with QT interval prolongation, including thioridazine, mesoridazine, and ziprasidone, and for monitoring for signs of myocarditis in patients treated with clozapine. Patients who receive both first- and second generation antipsychotic medications should be examined for extrapyramidal symptoms and tardive dyskinesia. Patients with schizophrenia should receive regular visual examinations. CONCLUSIONS: The conference participants recommended that mental health care providers perform physical health monitoring that typically occurs in primary care settings for their patients who do not receive physical health monitoring in those settings. This change in usual practice is recommended on the basis of the conference participants' belief that this additional monitoring will result in the earlier detection of common, serious risk factors that could, without detection and intervention, contribute to impaired health of patients with schizophrenia. PMID- 15285958 TI - Refining personality disorder diagnosis: integrating science and practice. AB - OBJECTIVE: Personality disorder researchers are currently evaluating a range of potential solutions to problems with the DSM-IV diagnostic categories. This article proposes changes to the diagnostic categories and criteria based on empirical findings from a national sample of patients with personality disorder diagnoses. METHOD: The Shedler-Westen Assessment Procedure (SWAP-200) is a personality assessment tool designed to capture the richness and complexity of clinical personality descriptions while providing reliable and quantifiable data. A national sample of experienced psychiatrists and psychologists used the SWAP 200 to describe either their conceptions (prototypes) of personality disorders (N=267) or current patients with personality disorder diagnoses (N=530). RESULTS: Clinicians" conceptions of personality disorders and their descriptions of actual patients overlapped with the DSM descriptions but also differed in systematic ways. Their descriptions were clinically richer than the DSM descriptions and placed greater emphasis on patients" mental life or inner experience. The study identifies potential diagnostic criteria that may be more defining of personality syndromes than some of the current DSM criteria. CONCLUSIONS: Diagnostic criterion sets should be expanded to better address the multiple domains of functioning inherent in the concept of personality and should more explicitly address patients' mental life or inner experience. The authors offer recommendations for revision of the diagnostic categories and criteria and also propose a prototype matching approach to personality disorder diagnosis that may overcome limitations inherent in the current diagnostic system. PMID- 15285959 TI - Structure of the human prefrontal cortex. PMID- 15285961 TI - Acute stress disorder, posttraumatic stress disorder, and depression in disaster or rescue workers. AB - OBJECTIVE: The events of Sept. 11, 2001, highlighted the importance of understanding the effects of trauma on disaster workers. To better plan for the health care of disaster workers, this study examined acute stress disorder, posttraumatic stress disorder (PTSD), early dissociative symptoms, depression, and health care utilization in disaster workers. METHOD: Exposed disaster workers (N=207) and unexposed comparison subjects (N=421) were examined at 2, 7, and 13 months after an airplane crash. RESULTS: Exposed disaster workers had significantly higher rates of acute stress disorder, PTSD at 13 months, depression at 7 months, and depression at 13 months than comparison subjects. Those who were younger and single were more likely to develop acute stress disorder. Exposed disaster workers with acute stress disorder were 3.93 times more likely to be depressed at 7 months. Those with high exposure and previous disaster experience or who had acute stress disorder were more likely to develop PTSD. Similarly, those who were depressed at 7 months were 9.5 times more likely to have PTSD. Those who were depressed at 13 months were 7.96 times more likely to also meet PTSD criteria. More exposed disaster workers than comparison subjects obtained medical care for emotional problems at 2, 7, and 13 months. Overall, 40.5% of exposed disaster workers versus 20.4% of comparison subjects had acute stress disorder, depression at 13 months, or PTSD. CONCLUSIONS: Exposed disaster workers are at increased risk of acute stress disorder, depression, or PTSD and seek care for emotional problems at an increased rate. PMID- 15285962 TI - Use of psychotropic medications before and after Sept. 11, 2001. AB - OBJECTIVE: This study examined patterns of psychotropic medication use after the Sept. 11, 2001, terrorist attacks. METHOD: It drew from two large pharmacy data sets, one providing nationally representative aggregate projections for all U.S. prescriptions (156.9 million claims for psychotropic medications during the study period) and a second from the nation's largest pharmacy benefit management organization (36.4 million enrollees per month), 4.1% of whom had a prescription for a psychotropic medication during the study period. Analyses examined use of antidepressant, antipsychotic, anxiolytic, and hypnotic medications in the 12 weeks before and after Sept. 11, 2001, compared with the same weeks during 2000. RESULTS: Nationally and in Washington, D.C., there was no evidence of an increase in overall prescriptions, new prescriptions, or daily doses for psychotropic medications. In New York City, there was an increase in the proportion of existing users with psychotropic dose increases in the weeks after the attacks (16.9% in 2001 versus 13.6% in 2000) but no significant increase in the rate of new psychotropic prescriptions. CONCLUSIONS: For most of the nation, the distress associated with the terrorist attacks was not accompanied by a commensurate increase in the use of psychotropic medications. In New York City, there was a statistically significant but modest increase in the proportion of individuals with dose increases in their psychotropic medications. PMID- 15285963 TI - The psychological impact of terrorism: an epidemiologic study of posttraumatic stress disorder and associated factors in victims of the 1995-1996 bombings in France. AB - OBJECTIVE: A wave of bombings struck France in 1995 and 1996, killing 12 people and injuring more than 200. The authors conducted follow-up evaluations with the victims in 1998 to determine the prevalence of and factors associated with posttraumatic stress disorder (PTSD). METHOD: Victims directly exposed to the bombings (N=228) were recruited into a retrospective, cross-sectional study. Computer-assisted telephone interviews were conducted to evaluate PTSD, per DSM IV criteria, and to assess health status before the attack, initial injury severity and perceived threat at the time of attack, and psychological symptoms, cosmetic impairment, hearing problems, and health service use at the time of the follow-up evaluation. Factors associated with PTSD were investigated with univariate logistic regression followed by multiple logistic regression analyses. RESULTS: A total of 196 respondents (86%) participated in the study. Of these, 19% had severe initial physical injuries (hospitalization exceeding 1 week). Problems reported at the follow-up evaluation included attack-related hearing problems (51%), cosmetic impairment (33%), and PTSD (31%) (95% confidence interval=24.5%-37.5%). Results of logistic regression analyses indicated that the risk of PTSD was significantly higher among women (odds ratio=2.54), participants age 35-54 (odds ratio=2.83), and those who had severe initial injuries (odds ratio=2.79) or cosmetic impairment (odds ratio=2.74) or who perceived substantial threat during the attack (odds ratio=3.99). CONCLUSIONS: The high prevalence of PTSD 2.6 years on average after a terrorist attack emphasizes the need for improved health services to address the intermediate and long-term consequences of terrorism. PMID- 15285964 TI - Posttraumatic stress disorder and depression following trauma: understanding comorbidity. AB - OBJECTIVE: Posttraumatic stress disorder (PTSD) and major depression occur frequently following traumatic exposure, both as separate disorders and concurrently. This raises the question of whether PTSD and depression are separate disorders in the aftermath of trauma or part of a single general traumatic stress construct. This study aimed to explore the relationships among PTSD, depression, and comorbid PTSD/depression following traumatic injury. METHOD: A group of 363 injury survivors was assessed just prior to discharge from hospital and 3 and 12 months postinjury. Canonical correlations were used to examine the relationship between PTSD and depression symptom severity and a set of predictor variables. Multinomial logistic regression was used to identify whether the diagnostic categories of PTSD, depression, and comorbid PTSD/depression were associated with different groups of predictors. RESULTS: The majority of psychopathology in the aftermath of trauma was best conceptualized as a general traumatic stress factor, suggesting that when PTSD and depression occur together, they reflect a shared vulnerability with similar predictive variables. However, there was also evidence that in a minority of cases at 3 months, depression occurs independently from PTSD and was predicted by a different combination of variables. CONCLUSIONS: While PTSD and comorbid PTSD/depression are indistinguishable, the findings support the existence of depression as a separate construct in the acute, but not the chronic, aftermath of trauma. PMID- 15285965 TI - The ACTH response to dexamethasone in PTSD. AB - OBJECTIVE: Enhanced negative feedback and reduced adrenal output are two different models that have been put forth to explain the paradoxical observations of increased release of corticotropin-releasing factor in the face of low cortisol levels in posttraumatic stress disorder (PTSD). To discriminate between these models, the authors measured levels of adrenocorticopic hormone (ACTH) and cortisol at baseline and in response to dexamethasone in medically healthy subjects with and without PTSD. Under conditions of enhanced negative feedback inhibition, ACTH levels would not be altered relative to cortisol levels, but the ACTH response to dexamethasone would be augmented, in concert with the enhanced cortisol response to dexamethasone. In contrast, under conditions of reduced adrenal output, ACTH levels would be expected to be higher at baseline relative to cortisol levels, but the ACTH response to dexamethasone would be unchanged in PTSD relative to healthy comparison subjects. METHOD: The ACTH and cortisol responses to 0.50 mg of dexamethasone were assessed in 19 subjects (15 men and four women) with PTSD and 19 subjects (14 men and five women) without psychiatric disorder. RESULTS: The ACTH-to-cortisol ratio did not differ between groups before or after dexamethasone, but the subjects with PTSD showed greater suppression of ACTH (as well as cortisol) in response to dexamethasone. CONCLUSIONS: The data support the hypothesis of enhanced cortisol negative feedback inhibition of ACTH secretion at the level of the pituitary in PTSD. Pituitary glucocorticoid receptor binding, rather than low adrenal output, is implicated as a likely mechanism for this effect. PMID- 15285966 TI - Impact of sleep deprivation and subsequent recovery sleep on cortisol in unmedicated depressed patients. AB - OBJECTIVE: One night of sleep deprivation induces a transient improvement in about 60% of depressed patients. Since depression is associated with abnormalities of the hypothalamic-pituitary-adrenal (HPA) axis, the authors measured cortisol secretion before, during, and after therapeutic sleep deprivation for 1 night. METHOD: Fifteen unmedicated depressed inpatients participated in a combined polysomnographic and endocrine study. Blood was sampled at 30-minute intervals during 3 consecutive nights before, during, and after sleep deprivation. Saliva samples were collected at 30-minute intervals during the daytime before and after the sleep deprivation night. RESULTS: During the night of sleep deprivation, cortisol levels were significantly higher than at baseline. During the daytime, cortisol levels during the first half of the day were higher than at baseline in the patients who responded to sleep deprivation but not in the nonresponders. During recovery sleep, cortisol secretion returned to baseline values. CONCLUSIONS: This study demonstrated a significant stimulatory effect of 1 night of sleep deprivation on the HPA axis in unmedicated depressed patients. The results suggest that the short-term effects of antidepressant treatments on the HPA axis may differ from their long-term effects. A higher cortisol level after sleep deprivation might transiently improve negative feedback to the hypothalamus or interact with other neurotransmitter systems, thus mediating or contributing to the clinical response. The fast return to baseline values coincides with the short clinical effect. PMID- 15285967 TI - Traumatic grief among adolescents exposed to a peer's suicide. AB - OBJECTIVE: The phenomenology of grief among children and adolescents is not well studied. A syndrome of traumatic grief, distinct from depression and anxiety, has been described among bereaved adults. The purpose of this study was to describe the symptoms and course of traumatic grief among adolescents exposed to a peer's suicide and to examine the relationship between traumatic grief and depression and posttraumatic stress disorder (PTSD) in this population. METHOD: A total of 146 friends and acquaintances of 26 suicide victims were included in this study. Subjects were interviewed at 6, 12-18, and 36 months after the peer's suicide. A subgroup was also interviewed 6 years afterward. The Texas Revised Inventory of Grief was administered at 6, 12-18, and 36 months; the Inventory of Complicated Grief was administered at the 6-year assessment. RESULTS: Principal component analysis of the Texas Revised Inventory of Grief resulted in two factors: one assessing a traumatic grief reaction and another assessing a milder or even normal grief reaction. The occurrence of traumatic grief was found to be independent from that of depression and PTSD. Traumatic grief at 6 months predicted the onset or course of depression and PTSD at subsequent assessments. CONCLUSIONS: Similar to adults, adolescents experience a traumatic grief reaction after exposure to a peer's suicide. Clinicians should be alerted to the occurrence of traumatic grief reactions among adolescents and the need to assess these reactions and address them in their treatment approaches. PMID- 15285968 TI - Implications of childhood trauma for depressed women: an analysis of pathways from childhood sexual abuse to deliberate self-harm and revictimization. AB - OBJECTIVE: Data from depressed women with and without a history of childhood sexual abuse were used to characterize clinical features that distinguished the two groups and to examine relationships of childhood sexual abuse to lifetime deliberate self-harm and recent interpersonal violence. METHOD: One hundred twenty-five women with depressive disorders were interviewed and completed self report questionnaires. Path analysis was used to examine relationships of several childhood and personality variables with deliberate self-harm in adulthood and recent interpersonal violence. RESULTS: Women with a childhood sexual abuse history reported more childhood physical abuse, childhood emotional abuse, and parental conflict in the home, compared to women without a childhood sexual abuse history. The two groups were similar in severity of depression, but the women with a childhood sexual abuse history were more likely to have attempted suicide and/or engaged in deliberate self-harm. The women with a history of childhood sexual abuse also became depressed earlier in life, were more likely to have panic disorder, and were more likely to report a recent assault. Path analysis confirmed the contributory role of childhood sexual abuse to deliberate self-harm and the significance of childhood physical abuse for recent interpersonal violence. CONCLUSIONS: Childhood sexual abuse is an important risk factor to identify in women with depression. Depressed women with a childhood sexual abuse history constitute a subgroup of patients who may require tailored interventions to combat both depression recurrence and harmful and self-defeating coping strategies. PMID- 15285969 TI - Promising treatments for women with comorbid PTSD and substance use disorders. AB - OBJECTIVE: The authors' goal was to compare the efficacy of a manualized cognitive behavior therapy that addresses both posttraumatic stress disorder (PTSD) and substance abuse (seeking safety) with a manualized cognitive behavior therapy that addresses only substance abuse (relapse prevention) and with standard community care for the treatment of comorbid posttraumatic stress disorder (PTSD) and substance use disorder. METHOD: One hundred seven women from an urban, low-income population who had comorbid PTSD and substance use disorder were randomly assigned to receive the two kinds of cognitive behavior therapy or received standard community treatment. Participants were recruited from both community and clinical populations and evaluated with structured clinical instruments. Forty-one women received seeking safety therapy, 34 received relapse prevention therapy, and 32 received standard community care. RESULTS: At the end of 3 months of treatment, participants in both cognitive behavior therapy conditions had significant reductions in substance use, PTSD, and psychiatric symptoms, but community care participants worsened over time. Both groups receiving cognitive behavior therapy sustained greater improvement in substance use and PTSD symptoms at 6-month and 9-month follow-ups than subjects in the community care group. CONCLUSIONS: Seeking safety and relapse prevention are efficacious short-term treatments for low-income urban women with PTSD, substance use disorder, and other psychiatric symptoms. PMID- 15285970 TI - Prospective study of clinical predictors of suicidal acts after a major depressive episode in patients with major depressive disorder or bipolar disorder. AB - OBJECTIVE: The authors investigated the predictive potential of a stress diathesis model for suicidal behavior based on correlates of past suicidal acts. In this model, suicidal acts are precipitated by stressors such as life events or a major depressive episode in the setting of a propensity for acting on suicidal urges. This diathesis is expressed as the tendency to develop more pessimism in response to a stressor and/or the presence of aggressive/impulsive traits. The predictive potential of the diathesis was tested by determining whether clinical correlates of past suicidal behavior predict suicidal acts during a 2-year follow up of patients with a major depressive episode. METHOD: Patients with DSM-III-R major depressive disorder or bipolar disorder (N=308) were assessed at presentation for treatment of a major depressive episode. Potential predictors of suicidal acts in the 2 years after study enrollment were identified on the basis of an association with previous suicidal behavior and were tested by using Cox proportional hazards regression analysis. In addition, pessimism and aggression/impulsivity factors were generated, and their predictive ability was tested by using Cox proportional hazards regression analysis. RESULTS: The three most powerful predictors of future suicidal acts were a history of suicide attempt, subjective rating of the severity of depression, and cigarette smoking, each of which had an additive effect on future risk. The pessimism and aggression/impulsivity factors both predicted suicidal acts, and each factor showed an additive effect. CONCLUSIONS: In addition to obtaining a history of suicidal behavior, clinicians may find it useful to assess patients' current level of pessimism, aggressive/impulsive traits, and comorbidity with substance use disorders, including nicotine-related disorders, to help identify patients at risk for suicidal behavior after major depression. Interventions such as aggressive pharmacotherapeutic prophylaxis to prevent relapse or recurrence of depressive symptoms may protect such at-risk individuals from future suicidal behavior. PMID- 15285971 TI - Factors contributing to therapists' distress after the suicide of a patient. AB - OBJECTIVE: Factors contributing to therapists' severe distress after the suicide of a patient were investigated. METHOD: Therapists for 34 patients who died by suicide completed a semistructured questionnaire about their reactions, wrote case narratives, and participated in a workshop. RESULTS: Thirteen of the 34 therapists were severely distressed. Four factors were identified as sources of severe distress: failure to hospitalize an imminently suicidal patient who then died, a treatment decision the therapist felt contributed to the suicide, negative reactions from the therapist's institution, and fear of a lawsuit by the patient's relatives. Although one emotion was sometimes dominant in the therapist's response to the suicide, severely distressed therapists, compared to others, reported a significantly larger number of intense emotional states. CONCLUSIONS: Over one-third of therapists who experienced a patient's suicide were found to suffer severe distress, pointing to the need for further study of the long-term effects of patient suicide on professional practice. PMID- 15285972 TI - Correlates of 1-year prospective outcome in bipolar disorder: results from the Stanley Foundation Bipolar Network. AB - OBJECTIVE: The purpose of the study was to examine potential correlates of outcome in patients treated for bipolar disorder. METHOD: During a 1-year period, 258 patients with DSM-IV bipolar disorder or schizoaffective disorder were rated with the prospective NIMH-Life Chart Method, which characterizes each day in terms of the severity of manic and depressive symptoms on the basis of patients' mood-related impairment in their usual educational, social, or occupational roles. Mean ratings for the severity of mania, depression, and overall bipolar illness and the number of manic, depressive, and overall illness episodes were calculated. Potential risk factors were assessed at the start of the study, and multivariate linear regression analysis was used to determine the correlates of the six 1-year outcome measures. RESULTS: Three of the six outcome measures were largely independent of each other and were used in the analysis. The mean rating for severity of mania was associated with comorbid substance abuse, history of more than 10 prior manic episodes, and poor occupational functioning at study entry. The mean rating for severity of depression was associated with a history of more than 10 prior depressive episodes and poor occupational functioning at study entry. The total number of overall illness episodes was associated with a positive family history of drug abuse, a history of prior rapid cycling, and poor occupational functioning. In addition, the mean rating for severity of mania and the total number of overall illness episodes were both initially associated with a history of childhood abuse, but these relationships were lost with the addition of other illness variables to the analysis. CONCLUSIONS: Clinicians who treat patients with bipolar disorder should consider a family history of drug abuse, a history of childhood abuse, prior course of illness, comorbid substance abuse, and occupational functioning in determining prognosis and setting goals for further treatment. PMID- 15285973 TI - Improving access to geriatric mental health services: a randomized trial comparing treatment engagement with integrated versus enhanced referral care for depression, anxiety, and at-risk alcohol use. AB - OBJECTIVE: The authors sought to determine whether integrated mental health services or enhanced referral to specialty mental health clinics results in greater engagement in mental health/substance abuse services by older primary care patients. METHOD: This multisite randomized trial included 10 sites consisting of primary care and specialty mental health/substance abuse clinics. Primary care patients 65 years old or older (N=24,930) were screened. The final study group consisted of 2,022 patients (mean age=73.5 years; 26% female; 48% ethnic minority) with depression (N=1,390), anxiety (N=70), at-risk alcohol use (N=414), or dual diagnosis (N=148) who were randomly assigned to integrated care (mental health and substance abuse providers co-located in primary care; N=999) or enhanced referral to specialty mental health/substance abuse clinics (i.e., facilitated scheduling, transportation, payment; N=1,023). RESULTS: Seventy-one percent of patients engaged in treatment in the integrated model compared with 49% in the enhanced referral model. Integrated care was associated with more mental health and substance abuse visits per patient (mean=3.04) relative to enhanced referral (mean=1.91). Overall, greater engagement was predicted by integrated care and higher mental distress. For depression, greater engagement was predicted by integrated care and more severe depression. For at-risk alcohol users, greater engagement was predicted by integrated care and more severe problem drinking. For all conditions, greater engagement was associated with closer proximity of mental health/substance abuse services to primary care. CONCLUSIONS: Older primary care patients are more likely to accept collaborative mental health treatment within primary care than in mental health/substance abuse clinics. These results suggest that integrated service arrangements improve access to mental health and substance abuse services for older adults who underuse these services. PMID- 15285974 TI - Residential versus community treatment of personality disorders: a comparative study of three treatment programs. AB - OBJECTIVE: The aim of this study was to compare the effectiveness of three treatment models for personality disorder: 1) a long-term psychoanalytically oriented residential specialist program, 2) a phased "step-down" specialist psychosocial program that included a briefer residential stay and an outpatient component, and 3) a general community psychiatric model. METHOD: One hundred forty-three patients with a personality disorder diagnosis were allocated according to geographical criteria to the three treatment conditions. Outcome was prospectively evaluated at 6, 12, and 24 months through the use of a standardized battery of instruments that included measures of general symptom severity, social adaptation, assessment of mental health functioning, frequency of self-harm and suicide attempts, and rates and duration of hospital readmissions. RESULTS: By 24 months, patients in the step-down condition showed significant improvements on all measures. Patients in the long-term residential model showed significant improvements in symptom severity, social adaptation, and global functioning, while no changes were achieved in self-harm, attempted suicide, and readmission rates. Patients in the general psychiatric group showed no improvement on any variables except self-harm and hospital readmissions. CONCLUSIONS: The results of this study suggest that for personality disorders, a specialist step-down program is more effective than both long-term residential treatment and general psychiatric treatment in the community. Replication is needed that includes a random allocation of patients to conditions to ensure that geographical factors did not account for the observed differences. PMID- 15285975 TI - Treatment outcomes in depression: comparison of remote treatment through telepsychiatry to in-person treatment. AB - OBJECTIVE: Telepsychiatry is an increasingly common method of providing psychiatric care, but randomized trials of telepsychiatric treatment compared to in-person treatment have not been done. The primary objective of this study was to compare treatment outcomes of patients with depressive disorders treated remotely by means of telepsychiatry to outcomes of depressed patients treated in person. Secondary objectives were to determine if patients' rates of adherence to and satisfaction with treatment were as high with telepsychiatric as with in person treatment and to compare costs of telepsychiatric treatment to costs of in person treatment. METHOD: In this randomized, controlled trial, 119 depressed veterans referred for outpatient treatment were randomly assigned to either remote treatment by means of telepsychiatry or in-person treatment. Psychiatric treatment lasted 6 months and consisted of psychotropic medication, psychoeducation, and brief supportive counseling. Patients' treatment outcomes, satisfaction, and adherence and the costs of treatment were compared between the two conditions. RESULTS: Hamilton Depression Rating Scale and Beck Depression Inventory scores improved over the treatment period and did not differ between treatment groups. The two groups were equally adherent to appointments and medication treatment. No between-group differences in dropout rates or patients' ratings of satisfaction with treatment were found. Telepsychiatry was more expensive per treatment session, but this difference disappeared if the costs of psychiatrists' travel to remote clinics more than 22 miles away from the medical center were considered. Telepsychiatry did not increase the overall health care resource consumption of the patients during the study period. CONCLUSIONS: Remote treatment of depression by means of telepsychiatry and in-person treatment of depression have comparable outcomes and equivalent levels of patient adherence, patient satisfaction, and health care cost. PMID- 15285976 TI - Academic performance of psychiatrists compared to other specialists before, during, and after medical school. AB - OBJECTIVE: This study was designed to compare psychiatrists with other physicians on measures of academic performance before, during, and after medical school. METHOD: More than three decades of data for graduates of Jefferson Medical College (N=5,701) were analyzed. Those who pursued psychiatry were compared to physicians in seven other specialties on 18 performance measures. Analysis of covariance was used to control for gender effect. RESULTS: Compared to other physicians, psychiatrists scored higher on measures of verbal ability and general information before medical school and on evaluations of knowledge and skills in behavioral sciences during medical school, but they scored lower on United States Medical Licensing Examinations step 3. CONCLUSIONS: The results generally confirmed the authors' expectations about psychiatrists' academic performance. More attention should be paid to the general medical education of psychiatrists. PMID- 15285977 TI - Psychopathology after rape. AB - OBJECTIVE: This study evaluated the psychopathological consequences of a single rape occurring in adult women. METHOD: The psychiatric symptoms reported by 40 women who were victims of rape during the previous 9 months as decided by a court of law were compared with the symptoms of 32 women who underwent severe, nonsexual, life-threatening events (car accidents, physical attacks, or robberies). None of the raped women had experienced previous sexual abuse during childhood or adolescence. RESULTS: The raped women showed a significantly greater prevalence of posttraumatic stress disorder, as well as sexual, eating, and mood disorders. CONCLUSIONS: These findings indicate that the psychopathological consequences of a rape could be specific and may warrant particular attention. PMID- 15285978 TI - Differential response to placebo among patients with social phobia, panic disorder, and obsessive-compulsive disorder. AB - OBJECTIVE: Placebo effects in treatment of three anxiety disorders were compared. METHOD: Treatment response and patients' treatment expectancy were examined by using data from 70 patients with obsessive-compulsive disorder, social phobia, or panic disorder who received placebo in three randomized, controlled trials comparing cognitive behavior therapy, medication, and their combination to placebo. RESULTS: Patients with obsessive-compulsive disorder were less likely to respond to placebo than patients with generalized social phobia or panic disorder. Differential expectancy did not account for these findings. CONCLUSIONS: Further examination of the placebo effect across the anxiety disorders may elucidate maintenance mechanisms of these disorders and have implications for development of more effective treatments. PMID- 15285979 TI - Low-dose cortisol for symptoms of posttraumatic stress disorder. AB - OBJECTIVE: Because elevated cortisol levels inhibit memory retrieval in healthy human subjects, the present study investigated whether cortisol administration might also reduce excessive retrieval of traumatic memories and related symptoms in patients with chronic posttraumatic stress disorder (PTSD). METHOD: During a 3 month observation period, low-dose cortisol (10 mg/day) was administered orally for 1 month to three patients with chronic PTSD in a double-blind, placebo controlled, crossover design. RESULTS: In each patient investigated, there was a significant treatment effect, with cortisol-related reductions of at least 38% in one of the daily rated symptoms of traumatic memories, as assessed by self administered rating scales. In accordance, Clinician-Administered PTSD Scale ratings assessed after each month showed cortisol-related improvements for reexperiencing symptoms and, additionally, in one patient for avoidance symptoms. CONCLUSIONS: The results of this pilot study indicate that low-dose cortisol treatment reduces the cardinal symptoms of PTSD. PMID- 15285980 TI - Sustained remission of schizophrenia among community-dwelling older outpatients. AB - OBJECTIVE: The frequency and nature of sustained remission of schizophrenia are controversial. METHOD: The authors assessed the prevalence of sustained remission among 155 middle-aged and elderly outpatients living independently. They compared patients with sustained remission to symptomatic schizophrenia patients and normal comparison subjects using standardized psychopathological, cognitive, and functional measures. RESULTS: Eight percent of the older schizophrenia patients living independently met criteria for sustained remission. Their level of psychopathology was similar to that in normal subjects and lower than that in symptomatic patients. On cognition, quality of well-being, and everyday functioning, the group with sustained remission was intermediate between the normal and symptomatic groups and differed significantly from the normal subjects. CONCLUSIONS: Sustained remission can occur even in older patients with very chronic illness, but its prevalence is lower than that in several published reports. Remission may reflect a return to premorbid functioning, consistent with neurodevelopmental hypotheses of schizophrenia. PMID- 15285981 TI - Awareness of disorder and suicide risk in the treatment of schizophrenia: results of the international suicide prevention trial. AB - OBJECTIVE: Schizophrenia is characterized by high suicide risk and low awareness of disorder. Although awareness has benefits for medication compliance and clinical outcome, it is unclear how it may relate to suicide risk in this population. METHOD: This multicenter investigation assessed awareness and suicide related behavior in 980 patients with schizophrenia or schizoaffective disorder. Patients were followed over 2 years and assessed by blinded raters for suicide related events. RESULTS: Awareness of psychiatric condition at baseline was associated with increased risk of suicide events over the follow-up. This effect was mediated by depression and hopelessness levels. By contrast, changes in awareness associated with treatment decreased the risk of suicide. CONCLUSIONS: Although some patients may become depressed after acknowledging the clinical handicaps of their disorder, treatment-related changes in awareness are generally associated with a positive outcome relative to suicide risk. The complex interactions and mediation effects of these clinical variables require careful monitoring. PMID- 15285982 TI - Associated alterations of striatal dopamine D2/D3 receptor and transporter binding in drug-naive patients with schizophrenia: a dual-isotope SPECT study. AB - OBJECTIVE: In vivo imaging studies have revealed deregulated presynaptic or postsynaptic function of the midbrain dopaminergic system in patients with schizophrenia. To further delineate the neuropathological involvement of the presynaptic and postsynaptic dopamine neurons in schizophrenia, the authors examined brain D(2)-family receptor and dopamine transporter binding simultaneously in patients with drug-naive schizophrenia using the dual-isotope single photon emission computed tomography (SPECT) imaging technique. METHOD: Eleven patients with schizophrenia and 12 healthy comparison subjects were recruited. Striatal dopamine D(2)/D(3) receptor was measured with SPECT and [(123)I]iodobenzamide ([(123)I]IBZM), while dopamine transporter was measured with SPECT and [(99m)Tc]TRODAT-1. RESULTS: Striatal D(2)/D(3) receptor and dopamine transporter binding were unaltered in these drug-naive patients with schizophrenia. Nonetheless, D(2)/D(3) receptor binding measures were positively correlated with dopamine transporter binding measures in these patients but not in the comparison subjects. CONCLUSIONS: The associated presynaptic and postsynaptic disturbances of midbrain dopamine neurons could be clinically relevant in drug-naive patients with schizophrenia. PMID- 15285983 TI - C-reactive protein and serotonin syndrome. PMID- 15285984 TI - Hypophosphatemia in panic disorder. PMID- 15285985 TI - Clozapine treatment of dimenhydrinate abuse. PMID- 15285986 TI - Suicide methods from the internet. PMID- 15285987 TI - Elevated binding of D8/17-specific monoclonal antibody to B lymphocytes in Tic disorder patients. PMID- 15285988 TI - Clozapine-induced acute interstitial nephritis. PMID- 15285989 TI - Depression and the decision to abort. PMID- 15285990 TI - The relevance of epigenomics to psychiatry. PMID- 15285991 TI - Ziprasidone and mania. PMID- 15285992 TI - Illness severity and depression in multiple sclerosis. PMID- 15285993 TI - Serotonin 1A receptors in memory function. PMID- 15285994 TI - Mortality and poststroke depression. PMID- 15285995 TI - Mortality and poststroke depression. PMID- 15285996 TI - Terror management theory. PMID- 15285997 TI - Violence against women. PMID- 15285998 TI - Sample selection in Finnish adoptive family study. PMID- 15285999 TI - C-type natriuretic peptide attenuates bleomycin-induced pulmonary fibrosis in mice. AB - C-type natriuretic peptide (CNP) has been shown to play an important role in the regulation of vascular tone and remodeling. However, the physiological role of CNP in the lung remains unknown. Accordingly, we investigated whether CNP infusion attenuates bleomycin (BLM)-induced pulmonary fibrosis in mice. After intratracheal injection of BLM or saline, mice were randomized to receive continuous infusion of CNP or vehicle for 14 days. CNP infusion significantly reduced the total number of cells and the numbers of macrophages, neutrophils, and lymphocytes in bronchoalveolar lavage fluid. Interestingly, CNP markedly reduced bronchoalveolar lavage fluid IL-1beta levels. Immunohistochemical analysis demonstrated that CNP significantly inhibited infiltration of macrophages into the alveolar and interstitial regions. CNP infusion significantly attenuated BLM-induced pulmonary fibrosis, as indicated by significant decreases in Ashcroft score and lung hydroxyproline content. CNP markedly decreased the number of Ki-67-positive cells in fibrotic lesions of the lung, suggesting antiproliferative effects of CNP on pulmonary fibrosis. Kaplan Meier survival curves demonstrated that BLM mice treated with CNP had a significantly higher survival rate than those given vehicle. These results suggest that continuous infusion of CNP attenuates BLM-induced pulmonary fibrosis and improves survival in BLM mice, at least in part by inhibition of pulmonary inflammation and cell proliferation. PMID- 15286000 TI - Upregulation of erythropoietin receptor during postnatal and postpneumonectomy lung growth. AB - Circulating erythropoietin (EPO) stimulates erythrocytosis, whereas organ specific local EPO receptor (EPOR) expression has been linked to angiogenesis, tissue growth, and development. On the basis of the observation of concurrent enhancement of lung growth and erythrocyte production during exposure to chronic hypoxia, we hypothesized that a paracrine EPO system is involved in mediating lung growth. We analyzed EPOR protein expression in normal dog lung tissue during postnatal maturation and during compensatory lung growth after right pneumonectomy (PNX). Membrane-bound EPOR was significantly more abundant in the immature lung compared with mature lung and in the remaining lung 3 wk after PNX compared with matched sham controls. COOH-terminal cytosolic EPOR peptides, which were even more abundant than membrane-bound EPOR, were also upregulated in immature lung but differentially processed after PNX. Apoptosis was enhanced during both types of lung growth in direct relationship to cellular proliferation and EPOR expression. We conclude that both developmental and compensatory lung growth involve paracrine EPO signaling with parallel upregulation but differential processing of EPOR. PMID- 15286001 TI - Fibrogenic cytokine levels in bronchoalveolar lavage aspirates 15 years after exposure to sulfur mustard. AB - Over 100,000 Iranian war veterans suffer from chronic effects of mustard gas exposure. Sulfur mustard was used by Iraq during the Iraqi-imposed war on Iran (between 1980 and 1988). The major complaints of these patients are mild interstitial fibrosis and bronchiolitis. We aimed to determine the state of fibrosis progression and assessed transforming growth factor (TGF)-beta1 levels in pulmonary samples and in bronchoalveolar lavage (BAL) aspirates. A total of 126 war veterans confirmed for lung disease were assessed and compared with three control groups: 1) 64 veterans not exposed to chemical agents, 2) 12 idiopathic pulmonary fibrosis civilian patients, and 3) 33 normal persons. BAL was performed via a flexible fiber-optic bronchoscope and the standard manual method. Total protein was measured by Bradford assay, and samples were corrected with regard to coefficients. Samples were concentrated 15-fold by lyophilization and resolubilization. Samples were double-checked using an ELISA test kit. The Mann Whitney test was used for the data analysis using commercial software. We detected that significant differences between TGF-beta1 levels between the case group and control group 1 (P = 0.001) and control group 3 (P = 0.003). No significant differences were found between the case group and control group 2 (P = 0.57). Inflammation and fibrotic processes in lung tissue of patients exposed to sulfur mustard may be progressive so IFN-gamma may be a useful drug to these patients' treatment. PMID- 15286002 TI - BMPR-II heterozygous mice have mild pulmonary hypertension and an impaired pulmonary vascular remodeling response to prolonged hypoxia. AB - Heterozygous mutations of the bone morphogenetic protein type II receptor (BMPR II) gene have been identified in patients with primary pulmonary hypertension. The mechanisms by which these mutations contribute to the pathogenesis of primary pulmonary hypertension are not fully elucidated. To assess the impact of a heterozygous mutation of the BMPR-II gene on the pulmonary vasculature, we studied mice carrying a mutant BMPR-II allele lacking exons 4 and 5 (BMPR-II(+/-) mice). BMPR-II(+/-) mice had increased mean pulmonary arterial pressure and pulmonary vascular resistance compared with their wild-type littermates. Histological analyses revealed that the wall thickness of muscularized pulmonary arteries (<100 mum in diameter) and the number of alveolar-capillary units were greater in BMPR-II(+/-) than in wild-type mice. Breathing 11% oxygen for 3 wk increased mean pulmonary arterial pressure, pulmonary vascular resistance, and hemoglobin concentration to similar levels in BMPR-II(+/-) and wild-type mice, but the degree of muscularization of small pulmonary arteries and formation of alveolar-capillary units were reduced in BMPR-II(+/-) mice. Our results suggest that, in mice, mutation of one copy of the BMPR-II gene causes pulmonary hypertension but impairs the ability of the pulmonary vasculature to remodel in response to prolonged hypoxic breathing. PMID- 15286003 TI - Barrier dysfunction and RhoA activation are blunted by homocysteine and adenosine in pulmonary endothelium. AB - RhoA GTPases modulate endothelial permeability. We have previously shown that adenosine and homocysteine enhance basal barrier function in pulmonary artery endothelial cells by a mechanism involving diminution of RhoA carboxyl methylation and activity. In the current study, we investigated the effects of adenosine and homocysteine on endothelial monolayer permeability in cultured monolayers. Adenosine and homocysteine significantly attenuated thrombin-induced endothelial barrier dysfunction and intercellular gap formation. We found significantly diminished RhoA associated with the membrane subcellular fraction in endothelial cells pretreated with adenosine and homocysteine, compared with vehicle-treated endothelial cells. Additionally, adenosine and homocysteine significantly blunted RhoA activation following thrombin exposure. Incubation with adenosine and homocysteine also enhanced in vitro interactions between RhoA and RhoGDI, as well as subcellular translocation of p190RhoGAP to the cytosol. These data demonstrate that elevated intracellular concentrations of homocysteine and adenosine enhance endothelial barrier function in cultured endothelial cells isolated from the main pulmonary artery and lung microvasculature, suggesting a potentially protective effect against pulmonary edema in response to lung injury. We speculate that homocysteine and adenosine modulate the level of endothelial barrier dysfunction through modulation of RhoA posttranslational processing resulting in diminished GTPase activity through altered interactions with modulators of RhoA activation. PMID- 15286004 TI - Superoxide dismutase moderates basal and induced bacterial adherence and interleukin-8 expression in airway epithelial cells. AB - Bacterial infection of the tracheobronchial tree is a frequent, serious complication in patients receiving treatment with oxygen and mechanical ventilation, resulting in increased morbidity and mortality. Using human airway epithelial cell culture models, we examined the effect of hyperoxia on bacterial adherence and the expression of interleukin-8 (IL-8), an important mediator involved in the inflammatory process. A 24-h exposure to 95% O(2) increased Pseudomonas aeruginosa (PA) adherence 57% in A549 cells (P < 0.01) and 115% in 16HBE cells (P < 0.01) but had little effect on Staphylococcus aureus (SA) adherence. Exposure to hyperoxia, followed by a 1-h incubation with SA, further enhanced PA adherence (P < 0.01), suggesting that hyperoxia and SA colonization may enhance the susceptibility of lung epithelial cells to gram-negative infections. IL-8 expression was also increased in cells exposed to both hyperoxia and PA. Stable or transient overexpression of manganese superoxide dismutase reduced both basal and stimulated levels of PA adherence and IL-8 levels in response to exposure to either hyperoxia or PA. These data indicate that hyperoxia increases susceptibility to infection and that the pathways are mediated by reactive oxygen species. Therapeutic intervention strategies designed to prevent accumulation of intracellular reactive oxygen species may reduce opportunistic pulmonary infections. PMID- 15286005 TI - Association between baseline radiographic damage and improvement in physical function after treatment of patients with rheumatoid arthritis. AB - OBJECTIVES: To identify factors associated with poor physical function in rheumatoid arthritis and to assess whether baseline joint damage has an impact on improvement in physical function during infliximab treatment. METHODS: 428 patients with active rheumatoid arthritis despite methotrexate treatment received methotrexate alone or with infliximab (3 mg/kg or 10 mg/kg every four or eight weeks) for 54 weeks (the ATTRACT trial). Data on clinical outcomes and physical function (assessed by the health assessment questionnaire (HAQ)) were collected. Structural damage was assessed using the van der Heijde modification of the Sharp score. Odds ratios (OR) for factors associated with severe functional disability (HAQ > or =2.0) at baseline were estimated using multiple logistic regression analyses, and baseline factors related to the change in physical function after treatment at week 54 were determined. RESULTS: Baseline radiographic scores were correlated with baseline HAQ scores. After adjustment for demographic characteristics in the logistic regression model, baseline disease activity scores, radiological joint damage, fatigue, and morning stiffness were found to be associated with severe functional disability (HAQ >2.0), with OR values of 2.00 (1.53 to 2.63), 1.82 (1.15 to 2.87), 1.19 (1.05 to 1.34), and 1.07 (1.01 to 1.13), respectively. In multiple linear regression analysis, physical disability, joint damage, and fatigue at baseline were correlated with less improvement in physical function after treatment. Infliximab treatment was associated with greater improvement in physical function. CONCLUSIONS: Greater joint damage at baseline was associated with poorer physical function at baseline and less improvement in physical function after treatment, underlining the importance of early intervention to slow the progression of joint destruction. PMID- 15286006 TI - Deciding on progression of joint damage in paired films of individual patients: smallest detectable difference or change. AB - Progression of radiological joint damage is usually based on the simultaneous assessment of a series of films from an individual patient ("paired", with or without known sequence). In this setting the degree of progression that can be reliably detected above the measurement error is best determined by the smallest detectable change, and overestimated by the traditionally calculated smallest detectable difference. This knowledge is important for calculation of the proportion of patients showing radiographic progression in clinical trials. PMID- 15286007 TI - Effect of treatment of rheumatoid arthritis with infliximab on IFN gamma, IL4, T bet, and GATA-3 expression: link with improvement of systemic inflammation and disease activity. AB - OBJECTIVE: To study interferon gamma (IFN gamma) production and the expression of T-bet and GATA-3, the transcription factors associated with Th1 and Th2, in peripheral blood mononuclear cells (PBMC) from patients with rheumatoid arthritis before and during infliximab treatment, so as to distinguish between a disease specific and a disease activity dependent defect. METHODS: Rheumatoid PBMC were obtained at weeks 0 and 6 of infliximab treatment and cultured for seven days with or without interleukin (IL)12 or the combination of IL12 and IL18. IFN gamma concentrations in supernatants were determined by ELISA. mRNA expression of IFN gamma, IL4, T-bet, and GATA-3 was determined by real time RT-PCR in whole blood at weeks 0 and 22. RESULTS: A reduction in spontaneous IFN gamma production and in the response to Th1 inducing cytokines occurred in rheumatoid PBMC. Reduction of systemic inflammation with infliximab treatment increased IFN gamma production in response to IL12 or IL12+IL18. The IFN gamma/IL4 expression ratio of rheumatoid blood before treatment was lower than in healthy controls but was increased by infliximab treatment. T-bet expression or T-bet/GATA-3 ratio of rheumatoid blood was less than in controls. The T-bet/GATA-3 ratio was not influenced by infliximab treatment. CONCLUSIONS: Regulation of T-bet and GATA-3 or IFN gamma and IL4 expression appeared different. The IFN gamma/IL4 ratio might express the blood Th1/Th2 balance better than the T-bet/GATA-3 ratio. Reduced IFN gamma production by rheumatoid PBMC and levels of IFN gamma and IL4 mRNA expression in blood were linked to disease improvement, indicating an association between this systemic Th1 feature and disease activity. PMID- 15286008 TI - Is rheumatoid arthritis disappearing? AB - During the past decades a number of studies have examined the incidence of rheumatoid arthritis (RA) in different geographical settings and at different times. Some studies from the 1970s and 1980s reported a higher incidence of RA than seen during recent years, where reported incidence numbers seems to have flattened out at a lower level. Besides a real time dependent decline of RA incidence, changing methodology in classification may be an equally important explanation. Today we may assume that annually 25-50 people from a population of 100,000 will develop typical RA. PMID- 15286009 TI - Anti-C1q antibodies in nephritis: correlation between titres and renal disease activity and positive predictive value in systemic lupus erythematosus. AB - OBJECTIVE: To investigate antibodies to complement 1q (anti-C1q) and investigate the correlation between anti-C1q titres and renal disease in systemic lupus erythematosus (SLE). METHODS: 151 SLE patients were studied. In patients with biopsy proven lupus nephritis (n = 77), activity of renal disease was categorised according to the BILAG renal score. Sera were tested for anti-C1q by enzyme immunoassay. Serum samples were randomly selected from 83 SLE patients who had no history of renal disease, and the positive and negative predictive value of the antibodies was studied. RESULTS: Patients with active lupus nephritis (BILAG A or B) had a higher prevalence of anti-C1q than those with no renal disease (74% v 32%; relative risk (RR) = 2.3 (95% confidence interval, 1.6 to 3.3)) (p<0.0001). There was no significant difference in anti-C1q prevalence between SLE without nephritis and SLE with non-active nephritis (BILAG C or D) (32% v 53%, p = 0.06) or between active and non-active nephritis (74% v 53%, p = 0.06). Patients with nephritis had higher anti-C1q levels than those without nephritis (36.0 U/ml (range 4.9 to 401.0) v 7.3 U/ml (4.9 to 401.0)) (p<0.001). Anti-C1q were found in 33 of 83 patients (39%) without history of renal disease. Nine of the 33 patients with anti-C1q developed lupus nephritis. The median renal disease-free interval was nine months. One patient with positive anti-C1q was diagnosed as having hypocomplementaemic urticarial vasculitis syndrome during follow up. CONCLUSIONS: Anti-C1q in SLE are associated with renal involvement. Monitoring anti-C1q and their titres in SLE patients could be important for predicting renal flares. PMID- 15286010 TI - Floral traits and pollination systems in the Caatinga, a Brazilian tropical dry forest. AB - BACKGROUND AND AIMS: Pollination is a critical stage in plant reproduction and thus in the maintenance and evolution of species and communities. The Caatinga is the fourth largest ecosystem in Brazil, but despite its great extent and its importance few studies providing ecological information are available, with a notable lack of work focusing on pollination biology. Here, general data are presented regarding the frequency of pollination systems within Caatinga communities, with the aim of characterizing patterns related to floral attributes in order to make possible comparisons with data for plant communities in other tropical areas, and to test ideas about the utility of syndromes. This paper also intends to provide a reference point for further studies on pollination ecology in this threatened ecosystem. METHODS: The floral traits and the pollination systems of 147 species were analysed in three areas of Caatinga vegetation in northeastern Brazil, and compared with world-wide studies focusing on the same subject. For each species, floral attributes were recorded as form, size, colour, rewards and pollination units. The species were grouped into 12 guilds according to the main pollinator vector. Analyses of the frequencies of the floral traits and pollination systems were undertaken. KEY RESULTS: Nectar and pollen were the most common floral resources and insect pollination was the most frequent, occurring in 69.9 % of the studied species. Of the entomophilous species, 61.7 % were considered to be melittophilous (43.1 % of the total). Vertebrate pollination occurred in 28.1 % of the species (ornithophily in 15.0 % and chiropterophily in 13.1 %), and anemophily was recorded in only 2.0 %. CONCLUSIONS: The results indicated that the pollination systems in Caatinga, despite climatic restrictions, are diversified, with a low percentage of generalist flowers, and similar to other tropical dry and wet forest communities, including those with high rainfall levels. PMID- 15286011 TI - Pod set related to photosynthetic rate and endogenous ABA in soybeans subjected to different water regimes and exogenous ABA and BA at early reproductive stages. AB - BACKGROUND AND AIMS: The physiological reasons for reduced pod set in soybean (Glycine max) caused by drought during anthesis are not established. The objective of this study was to investigate the involvement of photosynthetic rate (A), pod endogenous abscisic acid (ABA) and exogenously applied ABA and 6 benzylaminopurine (BA) in regulating pod set in soybean during drought. METHODS: Two pot experiments were done in a controlled-environment glasshouse. In expt I, soybeans were either well-watered (WW) or droughted by withholding water from 4 d before to 4 d after anthesis (DAA). In expt II, soybeans were drought-stressed (DS) from -4 to 4 DAA. From -2 to 4 DAA, some of the WW and DS plants were treated with 0.1 mm ABA or 1 mm BA. KEY RESULTS: Drought stress decreased A, but increased pod ABA concentration ([ABA]). Pod set decreased only when A had decreased by 40 %, and pod [ABA] had increased 1.5-fold. Beyond the thresholds, pod set correlated positively with A and negatively with pod [ABA]. Exogenously applied ABA decreased A and pod set in WW plants, whilst it increased A and pod set in DS plants; exogenous BA had opposite effects. In these plants, pod set correlated linearly with A. CONCLUSIONS: Drought-induced decrease in A is significant in inducing pod abortion, probably as a consequence of carbohydrate deprivation. The effects of ABA and BA on pod set may be partially due to their effects on photosynthate supply. PMID- 15286012 TI - Characterization of anisocotylous leaf formation in Streptocarpus wendlandii (Gesneriaceae): significance of plant growth regulators. AB - BACKGROUND AND AIMS: Unifoliate species of Gesneriaceae are unique, as they bear only one leaf throughout their life history. The development of this leaf (termed a macrocotyledon) derived from one of two cotyledons is intriguing. The other cotyledon does not develop further and is termed a microcotyledon. This process of unequal cotyledon development is termed anisocotyly. In this study the process of macrocotyeldon formation was studied and the effects of plant hormones on the macrocotyledon development were investigated. METHODS: Streptocarpus wendlandii was chosen as the main subject material, as it was found to be suitable for experimental studies in laboratory conditions. Morphological analyses were carried out with light and scanning electron microscopy. Plant hormones were applied exogenously. KEY RESULTS: The macrocotyledon of S. wendlandii is produced through cell division activity in the basal meristem of the enlarging cotyledon. The newly developed region in the macrocotyledon displayed distinct morphological changes, including the formation of long, needle-shaped trichomes. The newly formed region was surrounded by lateral veins. No such change was observed in the microcotyledon. Furthermore, it was shown that development of anisocotyly is suppressed by the application of cytokinin, resulting in the formation of two nearly equal-sized cotyledons. Both cotyledons displayed macrocotyledon characteristics. This observation in S. wendlandii was confirmed using Monophyllaea glabra, another unifoliate species in the same family. CONCLUSIONS: It is proposed that developmental changes of the macrocotyledon have characteristics of a developmental phase-change, and cytokinins may be involved in its formation. These results are discussed in the light of current knowledge of phase-change transitions in plant vegetative development. PMID- 15286013 TI - Morphological features and inheritance of Foliaceous Stipules of primary leaves in cowpea (Vigna unguiculata). AB - BACKGROUND AND AIMS: The presence of connate foliaceous stipules of primary leaves and their inheritance in cowpea (Vigna unguiculata) genotype EC394736 is reported for the first time. METHODS: The development of foliaceous stipules (FS) and their persistence were examined throughout the growth and developmental stages of the plants of the genotype EC394736. The shape, size, colour, texture and other parameters were examined in the field during the period 15-50 d after sowing. The area of FS was measured using image analysis software. The inheritance of FS was studied by making a cross between the genotype EC394763 with rudimentary stipules (RS) and the genotype EC394736, which has connate foliaceous stipules of primary leaves. The presence or absence of FS in plants of the F1, F2 and F3 generations was recorded. KEY RESULTS: The stipules developed along with the primary leaves in the genotype EC394736. One stipule of each primary leaf fused with the adjacent stipule of the other primary leaf forming a foliaceous structure. These stipules persisted on the plants for >50 d, even after the primary leaves had withered off. The F1 plants showed an absence of FS indicating the rudimentary stipules to be dominant over foliaceous stipules. The F2 segregation into 15 (RS) : 1 (FS) indicated that duplicate recessive genes controlled the presence of the FS. This was confirmed from the segregation pattern in the F3 generation. CONCLUSIONS: The presence of FS is a unique feature in cowpea genotype EC394736 and duplicate recessive genes govern it. The FS can be used as a morphological marker for identification of cowpea varieties. PMID- 15286014 TI - Model-based estimation of relative risks and other epidemiologic measures in studies of common outcomes and in case-control studies. AB - Some recent articles have discussed biased methods for estimating risk ratios from adjusted odds ratios when the outcome is common, and the problem of setting confidence limits for risk ratios. These articles have overlooked the extensive literature on valid estimation of risks, risk ratios, and risk differences from logistic and other models, including methods that remain valid when the outcome is common, and methods for risk and rate estimation from case-control studies. The present article describes how most of these methods can be subsumed under a general formulation that also encompasses traditional standardization methods and methods for projecting the impact of partially successful interventions. Approximate variance formulas for the resulting estimates allow interval estimation; these intervals can be closely approximated by rapid simulation procedures that require only standard software functions. PMID- 15286015 TI - Poliovirus vaccination during pregnancy, maternal seroconversion to simian virus 40, and risk of childhood cancer. AB - Before 1963, poliovirus vaccine produced in the United States was contaminated with simian virus 40 (SV40), which causes cancer in animals. To examine whether early-life SV40 infection can cause human cancer, the authors studied 54,796 children enrolled in the US-based Collaborative Perinatal Project (CPP) in 1959 1966, 52 of whom developed cancer by their eighth birthday. Those children whose mothers had received pre-1963 poliovirus vaccine during pregnancy (22.5% of the children) had an increased incidence of neural tumors (hazard ratio = 2.6, 95% confidence interval: 1.0, 6.7; 18 cases) and hematologic malignancies (hazard ratio = 2.8, 95% confidence interval: 1.2, 6.4; 22 cases). For 50 CPP children with cancer and 200 CPP control children, the authors tested paired maternal serum samples from pregnancy for SV40 antibodies using a virus-like particle enzyme immunoassay and a plaque neutralization assay. Overall, mothers exhibited infrequent, low-level SV40 antibody reactivity, and only six case mothers seroconverted by either assay. Using the two SV40 assays, maternal SV40 seroconversion during pregnancy was not consistently related to children's case/control status or mothers' receipt of pre-1963 vaccine. The authors conclude that an increased cancer risk in CPP children whose mothers received pre-1963 poliovirus vaccine was unlikely to have been due to SV40 infection transmitted from mothers to their children. PMID- 15286016 TI - Case-control study of cancer among US Army veterans exposed to simian virus 40 contaminated adenovirus vaccine. AB - Simian virus 40 (SV40) was an accidental contaminant of vaccines produced in monkey kidney tissue cultures in the 1950s and early 1960s, including a parenteral adenovirus vaccine given to several hundred thousand US military recruits. Detection of SV40 DNA in tumor tissues by some laboratories suggests that SV40 contributes to human cancers. To determine if entry into US Army service during periods of administration of SV40-contaminated adenovirus vaccine was associated with an increased risk of cancer, the authors conducted a case control study of cancer occurring in male Army veterans who entered service in 1959-1961. Cases of brain tumors (n = 181), mesothelioma (n = 10), and non Hodgkin's lymphoma (n = 220) were identified through a Veterans Administration hospital discharge database, as were colon cancer and lung cancer controls (n = 221). Exposure to adenovirus vaccine was assigned on the basis of known periods of adenovirus vaccine administration and dates of Army entry obtained for cancer cases and controls. The odds ratios associated with exposure to SV40-contaminated adenovirus vaccine were 0.81 (95% confidence interval (CI): 0.52, 1.24) for brain tumors, 1.41 (95% CI: 0.39, 5.15) for mesothelioma, and 0.97 (95% CI: 0.65, 1.44) for non-Hodgkin's lymphoma. These findings do not support a role for SV40 in the development of these cancers. PMID- 15286017 TI - Blood transfusion and risk of non-Hodgkin's lymphoma in Connecticut women. AB - The incidence and mortality rates of non-Hodgkin's lymphoma have been increasing worldwide. Allogeneic blood transfusion has been suggested as a risk factor for non-Hodgkin's lymphoma, but the results from epidemiologic studies have been inconsistent. Data from a population-based case-control study of Connecticut women were analyzed to evaluate this relation. A total of 601 histologically confirmed, non-Hodgkin's lymphoma incident cases identified between 1996 and 2000 and 717 randomly selected controls were included in this study. Allogeneic blood transfusion was not associated with the increased risk of non-Hodgkin's lymphoma overall (odds ratio = 1.0, 95% confidence interval: 0.7, 1.3) or by subtype of the disease. The risk also did not vary by number of allogeneic blood transfusions, age at first transfusion, or time since first transfusion. When the reason for blood transfusion was considered, an increased risk of non-Hodgkin's lymphoma was found only for allogeneic blood transfusion for reason of anemia. In summary, the authors' findings do not support the hypothesis that allogeneic blood transfusion increases the risk of non-Hodgkin's lymphoma. PMID- 15286018 TI - Methods of calculating deaths attributable to obesity. AB - Previously reported estimates of deaths attributable to obesity in the United States have been based on a method that only partially adjusts for confounding and does not allow for effect modification. In this study, the authors investigated the possible magnitude and direction of bias in estimating deaths attributable to obesity when such a method is used. Hypothetical examples are based on 1991 US population data and published relative risks. Incomplete adjustment for confounding of the obesity-mortality relation by age and sex led to a 17% overestimation of deaths due to obesity. Additional bias resulted from slight differences between the derivation cohort and the target population. For example, a difference of three percentage points in the proportion of people 80 years of age or older led to a 42% overestimation of deaths due to obesity. In addition, these estimates appear to be sensitive to minor differences in relative risks between a derivation cohort and the target population. A difference of 0.20 in relative risks almost doubled the number of deaths (97% overestimation). Estimates of deaths attributable to obesity can be biased if confounding and effect modification are not properly taken into account or if the relative risks are not estimated accurately. PMID- 15286019 TI - Dietary supplement use by US adults: data from the National Health and Nutrition Examination Survey, 1999-2000. AB - Data from the 1999-2000 National Health and Nutrition Examination Survey, a nationally representative, cross-sectional survey of US health and nutrition, were analyzed to assess prevalence of dietary supplement use overall and in relation to lifestyle and demographic characteristics. Fifty-two percent of adults reported taking a dietary supplement in the past month; 35% took a multivitamin/multimineral. Vitamin C, vitamin E, B-complex vitamins, calcium, and calcium-containing antacids were taken by more than 5% of adults. In bivariate analyses, female gender, older age, more education, non-Hispanic White race/ethnicity, any physical activity, normal/underweight, more frequent wine or distilled spirit consumption, former smoking, and excellent/very good self reported health were associated with greater use of any supplement and of multivitamin/multiminerals; in multivariable comparisons, the latter three characteristics were not associated with supplement use. Most supplements were taken daily and for at least 2 years. Forty-seven percent of adult supplement users took just one supplement; 55% of women and 63% of adults aged >or=60 years took more than one. These findings suggest that, to minimize possible spurious associations, epidemiologic studies of diet, demography, or lifestyle and health take dietary supplement use into account because of 1) supplements' large contribution to nutrient intake and 2) differential use of supplements by demographic and lifestyle characteristics. PMID- 15286020 TI - Relation between maternal recreational physical activity and plasma lipids in early pregnancy. AB - The authors examined the relation between recreational physical activity and plasma lipid concentrations in early pregnancy. Between 1996 and 2000, 925 normotensive, nondiabetic pregnant women in Washington State were interviewed at approximately 13 weeks' gestation regarding type, frequency, and duration of physical activity during the previous 7 days. Plasma triglyceride, total cholesterol, and high density lipoprotein cholesterol concentrations were measured in contemporaneous blood samples. After adjustment, mean triglyceride concentration was 12.7 mg/dl lower in women performing any physical activity versus none (95% confidence interval (CI): -22.7, -2.6). Mean triglyceride concentration was lower in women in the highest tertiles of time performing physical activity (-23.6 mg/dl, 95% CI: -34.9, -12.2), energy expenditure (-23.6 mg/dl, 95% CI: -35.1, -12.2), and peak intensity (-18.1 mg/dl, 95% CI: -29.5, 6.8) versus inactive women. Reductions in mean total cholesterol were also observed for women with the highest levels of time performing physical activity, energy expenditure, and peak intensity. Linear relations were observed across levels of physical activity measures for triglyceride and total cholesterol. No association was found between physical activity and high density lipoprotein cholesterol. These data suggest that habitual physical activity may attenuate pregnancy-associated dyslipidemia. PMID- 15286021 TI - Risk factors for proximal humerus fracture. AB - This case-control study of proximal humerus fracture included 448 incident female and male cases and 2,023 controls aged 45 years or older identified in five Northern California Kaiser Permanente Medical Centers in 1996-2001. Data were collected by using an interviewer-administered questionnaire. Some factors related to low bone mass, including number of fractures since age 45 years and low dietary calcium intake, were associated with increased risks of fracture, and factors thought to protect against bone loss, such as menopausal hormone therapy and calcium carbonate tablet use, were associated with reduced risks. Fall related risk factors included previous falls, diabetes mellitus, and difficulty walking in dim light. Possible fall-related risk factors suggested for the first time in this study were seizure medication use (adjusted odds ratio (OR) = 2.80, 95% confidence interval (CI): 1.45, 5.42), depression (OR = 1.34, 95% CI: 0.98, 1.84), almost always using a hearing aid (OR = 1.92, 95% CI: 1.12, 3.31 vs. never prescribed), and left-handedness (OR = 2.36, 95% CI: 1.51, 3.68 vs. right handedness). Difficulty with activities of daily living and lack of physical activity tended to be associated with increased risk. Prevention of falls among frail, osteoporotic persons would likely reduce the frequency of proximal humerus fracture. PMID- 15286022 TI - Folate intake and risk of Parkinson's disease. AB - In clinical studies, individuals with Parkinson's disease have had higher concentrations of plasma homocysteine than did controls, and experimental evidence suggests that folate deficiency or focal administration of homocysteine sensitizes dopaminergic neurons to the neurotoxicity of 1-methyl-4-phenyl-1,2,3,6 tetrahydropyridine. The authors thus prospectively investigated whether higher intake of folate, vitamin B(6), or vitamin B(12) was related to a lower risk of Parkinson's disease in the Health Professionals Follow-up Study (1986-2000) and the Nurses' Health Study (1980-1998). They documented Parkinson's disease diagnoses in 248 men and 167 women during the follow-up. Folate intake was not associated with the risk of Parkinson's disease; the relative risks for the highest compared with the lowest quintiles were 1.0 (95% confidence interval: 0.7, 1.5) in men and 1.3 (95% confidence interval: 0.8, 2.3) in women. Neither did they find significant associations in analyses stratified by age, smoking, alcohol consumption, or lactose intake. Intake of vitamin B(6) or vitamin B(12) also was not related to the risk of Parkinson's disease. The current study does not support the hypothesis that higher intake of folate or related B vitamins lowers the risk of Parkinson's disease. PMID- 15286023 TI - Excess stroke in Mexican Americans compared with non-Hispanic Whites: the Brain Attack Surveillance in Corpus Christi Project. AB - Mexican Americans are the largest subgroup of Hispanics, the largest minority population in the United States. Stroke is the leading cause of disability and third leading cause of death. The authors compared stroke incidence among Mexican Americans and non-Hispanic Whites in a population-based study. Stroke cases were ascertained in Nueces County, Texas, utilizing concomitant active and passive surveillance. Cases were validated on the basis of source documentation by board certified neurologists masked to subjects' ethnicity. From January 2000 to December 2002, 2,350 cerebrovascular events occurred. Of the completed strokes, 53% were in Mexican Americans. The crude cumulative incidence was 168/10,000 in Mexican Americans and 136/10,000 in non-Hispanic Whites. Mexican Americans had a higher cumulative incidence for ischemic stroke (ages 45-59 years: risk ratio = 2.04, 95% confidence interval: 1.55, 2.69; ages 60-74 years: risk ratio = 1.58, 95% confidence interval: 1.31, 1.91; ages >or=75 years: risk ratio = 1.12, 95% confidence interval: 0.94, 1.32). Intracerebral hemorrhage was more common in Mexican Americans (age-adjusted risk ratio = 1.63, 95% confidence interval: 1.24, 2.16). The subarachnoid hemorrhage age-adjusted risk ratio was 1.57 (95% confidence interval: 0.86, 2.89). Mexican Americans experience a substantially greater ischemic stroke and intracerebral hemorrhage incidence compared with non Hispanic Whites. As the Mexican-American population grows and ages, measures to target this population for stroke prevention are critical. PMID- 15286024 TI - Monte Carlo sensitivity analysis and Bayesian analysis of smoking as an unmeasured confounder in a study of silica and lung cancer. AB - Conventional confidence intervals reflect uncertainty due to random error but omit uncertainty due to biases, such as confounding, selection bias, and measurement error. Such uncertainty can be quantified, especially if the investigator has some idea of the amount of such bias. A traditional sensitivity analysis produces one or more point estimates for the exposure effect hypothetically adjusted for bias, but it does not provide a range of effect measures given the likely range of bias. Here the authors used Monte Carlo sensitivity analysis and Bayesian bias analysis to provide such a range, using data from a US silica-lung cancer study in which results were potentially confounded by smoking. After positing a distribution for the smoking habits of workers and referents, a distribution of rate ratios for the effect of smoking on lung cancer, and a model for the bias parameter, the authors derived a distribution for the silica-lung cancer rate ratios hypothetically adjusted for smoking. The original standardized mortality ratio for the silica-lung cancer relation was 1.60 (95% confidence interval: 1.31, 1.93). Monte Carlo sensitivity analysis, adjusting for possible confounding by smoking, led to an adjusted standardized mortality ratio of 1.43 (95% Monte Carlo limits: 1.15, 1.78). Bayesian results were similar (95% posterior limits: 1.13, 1.84). The authors believe that these types of analyses, which make explicit and quantify sources of uncertainty, should be more widely adopted by epidemiologists. PMID- 15286025 TI - Influence of study population on the identification of risk factors for sexually transmitted diseases using a case-control design: the example of gonorrhea. AB - The population prevalence of many sexually transmitted diseases (STDs) is low. Thus, most epidemiologic studies of STDs are conducted among STD clinic populations to maximize efficiency. However, STD clinic patients have unique sociobehavioral characteristics. To examine the potential effect of study population on identification of risk factors, the authors compared 1) STD clinic patients with a random digit dialing telephone sample, 2) general population cases with random digit dialing controls, and 3) STD clinic cases with STD clinic controls (Seattle, Washington, 1992-1995). Risk factors for gonorrhea identified among STD clinic patients formed a subset of those identified in the general population. In both populations, risk decreased with age (odds ratio for the general population (OR(GP)) = 0.4, 95% confidence interval (CI): 0.22, 0.59; odds ratio for the clinic population (OR(clinic)) = 0.5, 95% CI: 0.30, 0.81) and was increased among Blacks (OR(GP) = 15.5, 95% CI: 4.93, 49.0; OR(clinic) = 10.5, 95% CI: 4.51, 24.68) and persons whose partner had been jailed (OR(GP) = 5.4, 95% CI: 2.07, 13.9; OR(clinic )= 3.1, 95% CI: 1.32, 7.30). Additional factors associated with gonorrhea in the general population included secondary education (OR = 0.3, 95% CI: 0.11, 0.70), anal intercourse (OR = 10.5, 95% CI: 2.01, 54.7, STD history (OR = 5.9, 95% CI: 1.76, 19.5), meeting partners in structured settings (OR = 0.2, 95% CI: 0.09, 0.50), no condom use (OR = 3.2, 95% CI: 1.30, 7.89), and divorce (OR = 3.6, 95% CI: 1.07, 11.9). Risk factors identified in STD clinics will probably be confirmed in a general population sample, despite overcontrolling for shared behaviors; however, factors associated with both disease and STD clinic attendance may be missed. PMID- 15286026 TI - Re: "Four-year review of the use of race and ethnicity in epidemiologic and public health research". PMID- 15286027 TI - Re: "Relation of education and occupation-based socioeconomic status to incident Alzheimer's disease". PMID- 15286028 TI - Mitochondrial dysfunction in mouse oocytes results in preimplantation embryo arrest in vitro. AB - Oocyte mitochondrial dysfunction has been proposed as a cause of high levels of developmental retardation and arrest that occur in human preimplantation embryos generated using assisted reproductive technology in the treatment of some causes of female infertility. To investigate this, a model of mitochondrial dysfunction was developed in mouse oocytes using a method of photosensitization of the mitochondrion-specific dye, rhodamine-123. After in vitro fertilization, dye loaded and photosensitized oocytes showed developmental arrest in proportion to irradiation time. Morphological and metabolic assessments of zygotes indicated an increase in mitochondrial permeability that subsequently resulted in apoptotic degeneration. Development was partially restored by inhibition of mitochondrial permeability transition pore formation by oocyte pretreatment with cyclosporin A. Oocyte mitochondria are therefore physiological regulators of early embryo development and potential sites of pathological insult that may perturb oocyte and subsequent preimplantation embryo viability. These findings have important implications for the treatment of clinically infertile women using assisted reproductive technologies. PMID- 15286029 TI - Efficient generation of transgenic mice with intact yeast artificial chromosomes by intracytoplasmic sperm injection. AB - The production of animals with large transgenes is an increasingly valuable tool in biotechnology and for genetic studies, including the characterization and manipulation of large genes and polygenic traits. In the present study, we describe an intracytoplasmic sperm injection (ICSI) method for the stable incorporation and phenotypic expression of large yeast artificial chromosomes (YAC) constructs of submegabase and megabase magnitude. By coinjecting spermatozoa and YACs into metaphase II oocytes, we were able to produce founders exhibiting germline transmission of an intact and functional transgene of 250 kilobases, carrying the mouse tyrosinase locus, used here as a reporter gene to rescue the albinism of recipient mice. More than 35% transgenesis was obtained for this YAC transgene. When compared with the pronuclear microinjection standard method, the efficiency of the ICSI-mediated YAC transfer system was significantly greater. In summary, we describe, for the first time, stable incorporation in the host genome and correct phenotypic expression of large DNA constructs mediated by ICSI. PMID- 15286030 TI - Characterization of a novel postacrosomal perinuclear theca-specific protein, CYPT1. AB - The perinuclear theca (PT) is a unique cytoskeletal structure that surrounds the nucleus of the sperm. The posterior acrosome segment of the PT (postacrosomal PT) is thought to play roles in shaping the nucleus during differentiation of the spermatid and in activating the oocyte during fertilization. We isolated a cDNA clone that encoded a novel haploid germ cell-specific cysteine-rich perinuclear theca protein, CYPT1. The transcripts were expressed exclusively in testicular germ cells after meiotic division. Sequence analysis revealed that CYPT1 comprised 168 amino acids and that the N-terminal was rich in basic amino acids, including cysteine clusters. Immunohistochemical and biochemical analyses localized CYPT1 to the postacrosomal PT of elongated spermatids and mature sperm. The cypt1 had three paralogs that were expressed in adult testis. A comparison of genomic structure suggested that two of the three cypt1 paralogs were generated by gene triplication on the X chromosome, while one paralog was retrotransposed to an autosome. Interestingly, the 5'-flanking regions of these genes were highly homologous with the promoter region of the spermatid-specific gene Zfy-2. CYPT1 and the proteins of the paralogous genes constitute a novel, basic cysteine-rich sperm protein family that may contribute to the function of the postacrosomal PT during nuclear shaping. PMID- 15286031 TI - Isolation of genes associated with developmentally competent bovine oocytes and quantitation of their levels during development. AB - We have performed suppressive subtraction hybridization (SSH) of populations of developmentally competent and incompetent bovine oocytes from large (> or =5-mm) and small (< or =2-mm) follicles to isolate messenger RNA associated with the attainment of developmental competency. RNA was amplified in a linear fashion and then subjected to the SSH procedure to produce a library enriched for genes associated with competency. One thousand clones of this library were subjected to a differential screening approach to identify 31 potentially upregulated isolates. Sequencing revealed these to represent 21 genes. To rigorously identify the degree of upregulation and reproducibility thereof, we examined the expression of these genes in three separate pools of developmentally competent and incompetent oocytes by quantitative real-time PCR. Results indicated that upregulation varied from zero to threefold, showing that accurate quantification is essential for the interpretation of such differential screening experiments. Furthermore, it appears that the molecular causes for poor developmental capacity may be highly complex and be reliant on many small changes. We further characterized a selection of these novel and known maternally expressed genes for their absolute expression levels during maturation in the presence or absence of an inhibitor of transcription and during preattachment development. Last, the effect of nuclear transfer on the levels of these genes was assayed. Nuclear transfer was found to differentially affect transcript levels of genes expressed after embryonic genome activation but did not prevent the degradation of maternal transcripts or result in activation of maternal genes that are silent at blastocyst stages. PMID- 15286032 TI - Isolation of nascent messenger RNA from mouse preimplantation embryos. AB - The expression of the zygotic genome starts at the late one-cell stage in mouse embryos, and its regulation changes dynamically until the late two-cell stage. To understand this process, it is important to accumulate the profiles of the genes transcribed at any given instant at each stage of development. However, because large amounts of maternal mRNA accumulate in embryos to sustain early development, it is difficult to determine the profile of newly synthesized mRNA just after gene activation. To overcome this difficulty, we established a novel method of isolating nascent mRNA from the large pool of preexisting mRNA. Briefly, the procedure was as follows. Embryos were electrically permeabilized and loaded with 5-bromouridine-5'-triphosphate (BrUTP). Nascent mRNA with incorporated BrU was isolated by immunoprecipitation with an antibody recognizing BrU. The cDNA was synthesized from the isolated mRNA, and its abundance was evaluated using semiquantitative real-time PCR. Using this method, we examined the amounts of newly synthesized eIF-1A, MuERV-L, and cyclin-A2 transcripts in two-cell mouse embryos and compared them with the quantities of these transcripts present in the total mRNA pool. The amount of each transcript in the nascent mRNA fraction and in the total mRNA pool changed differently over time, demonstrating that this method can be used to obtain profiles of genes transcribed during development. PMID- 15286034 TI - Progesterone regulates granulosa cell viability through a protein kinase G dependent mechanism that may involve 14-3-3sigma. AB - Progesterone (P4) inhibits granulosa cell and spontaneously immortalized granulosa cell (SIGC) apoptosis by regulating membrane-initiated events. However, the nature of the signal transduction pathway that is induced by these membrane initiated events has not been defined. To gain insights into the P4-regulated signal transduction pathway, mouse granulosa cells and SIGCs were cultured with 8 br-cGMP and P4. In culture, 8-br-cGMP mimicked P4's antiapoptotic actions. Because cGMP activates protein kinase G (PKG), the effect of PKG antagonists on P4-regulated SIGC viability was assessed. P4's antiapoptotic action was attenuated by the PKG inhibitors, Rp-8-pCPT-cGMP, KT5823, the PKG-1alpha-specific inhibitor, DT-3, and a dominant negative PKG-1alpha. Further, the type I isoform of PKG was shown to be expressed by SIGCs and activated by P4. P4's antiapoptotic action was not affected by the PKA inhibitor, KT5720. Collectively, these findings indicate that P4 maintains SIGC viability by activating PKG-1alpha. PKG 1alpha-GFP was shown to localize predominantly to the cytoplasm of SIGCs. To identify potential cytoplasmic targets of PKG-1alpha, SIGCs were cultured for 5 h with P4 in the presence or absence of DT-3. Cell lysates were prepared and subjected to two-dimensional electrophoresis. The resulting gels were sequentially stained with ProQ-Diamond Gel Stain and Coomassie Blue to reveal phosphorylated proteins. The two-dimensional gels revealed one major protein, the phosphorylation status of which was abrogated by DT-3. Mass spectrometric analysis identified this protein as 14-3-3sigma, with 14-3-3sigma being phosphorylated on tyrosine 19, serine 28, serine 69, serine 74, threonine 90, threonine 98, and serine 116. Finally, difopein, a specific 14-3-3 inhibitor, was shown to induce apoptosis even in the presence of serum. These data suggest that 1) P4 regulates the phosphorylation status of 14-3-3sigma through a PKG-dependent pathway and 2) 14-3-3sigma plays a central and essential role in maintaining the viability of SIGCs. PMID- 15286033 TI - Regulation of matrix metalloproteinase-19 messenger RNA expression in the rat ovary. AB - Matrix metalloproteinases (MMPs) are instrumental in the constant tissue remodeling in the ovary. An induction of MMP-19 mRNA in periovulatory follicles has been reported in mouse ovaries. However, little is known about MMP-19 expression during the follicular and luteal periods or about the ovarian regulation of MMP-19 mRNA expression. We examined the expression pattern of MMP 19 mRNA during various reproductive phases and the periovulatory regulation of MMP-19 mRNA in the rat ovary. In gonadotropin-primed, immature rat ovaries, levels of MMP-19 mRNA transiently increased during both follicular growth and ovulation. The MMP-19 mRNA was localized to the theca-interstitial layer of growing follicles and to the granulosa and theca-interstitial layers of periovulatory follicles. A similar expression pattern of MMP-19 mRNA in periovulatory follicles was observed in ovaries from naturally cycling adult rats. Accumulation of MMP-19 mRNA was detected in regressing corpus luteum. The regulation of MMP-19 mRNA expression during the periovulatory period was investigated via in vivo studies and through in vitro culture studies on follicular cells. The hCG-induction of MMP-19 mRNA was mimicked by treating granulosa cells, but not theca-interstitial cells, from preovulatory follicles with LH or activators of the protein kinase (PK) A or PKC pathways. Cycloheximide blocked the LH- or forskolin-induced MMP-19 mRNA expression, demonstrating the requirement for new protein synthesis. In contrast, blocking activation of the progesterone receptor or prostaglandin synthesis had no effect on the increase in MMP-19 mRNA expression. In conclusion, the induction of MMP-19 mRNA suggests an important role of this proteinase during follicular growth, ovulation, and luteal regression. PMID- 15286035 TI - A mixture of the "antiandrogens" linuron and butyl benzyl phthalate alters sexual differentiation of the male rat in a cumulative fashion. AB - Prenatal exposure to environmental chemicals that interfere with the androgen signaling pathway can cause permanent adverse effects on reproductive development in male rats. The objectives of this study were to 1) determine whether a documented antiandrogen butyl benzyl phthalate (BBP) and/or linuron (an androgen receptor antagonist) would decrease fetal testosterone (T) production, 2) describe reproductive developmental effects of linuron and BBP in the male, 3) examine the potential cumulative effects of linuron and BBP, and 4) investigate whether treatment-induced changes to neonatal anogenital distance (AGD) and juvenile areola number were predictive of adult reproductive alterations. Pregnant rats were treated with either corn oil, 75 mg/kg/day of linuron, 500 mg/kg/day of BBP, or a combination of 75 mg/kg/day linuron and 500 mg/kg/day BBP from gestational Day 14 to 18. A cohort of fetuses was removed to assess male testicular T and progesterone production, testicular T concentrations, and whole body T concentrations. Male offspring from the remaining litters were assessed for AGD and number of areolae and then examined for alterations as young adults. Prenatal exposure to either linuron or BBP or BBP + linuron decreased T production and caused alterations to androgen-organized tissues in a dose additive manner. Furthermore, treatment-related changes to neonatal AGD and infant areolae significantly correlated with adult AGD, nipple retention, reproductive malformations, and reproductive organ and tissue weights. In general, consideration of the dose-response curves for the antiandrogenic effects suggests that these responses were dose additive rather than synergistic responses. Taken together, these data provide additional evidence of cumulative effects of antiandrogen mixtures on male reproductive development. PMID- 15286036 TI - Angiogenesis and morphometry of bovine placentas in late gestation from embryos produced in vivo or in vitro. AB - The objective of this study was to determine the effects of in vitro embryo production on angiogenesis and morphometry of the bovine placenta during late gestation. Blastocysts produced in vivo were recovered from superovulated Holstein cows. Blastocysts produced in vitro were obtained after culture of in vitro-matured and -fertilized Holstein oocytes. Single blastocysts from each production system were transferred into heifers. Fetuses and placentas were recovered on Day 222 of gestation (in vivo, n=12; in vitro, n=12). Cotyledonary and caruncular tissues were obtained for quantification of vascular endothelial growth factor (VEGF) and peroxisome proliferator-activated receptor-gamma (PPARgamma) mRNA and protein. Tissue sections of placentomes were prepared for morphometric analysis. Fetuses and placentas were heavier from embryos produced in vitro than from embryos produced in vivo. More placentas from embryos produced in vitro had an excessive volume of placental fluid. There was no effect of treatment on the expression of mRNA for VEGF and PPARgamma in either cotyledonary or caruncular tissues. The expression of VEGF protein in cotyledons and caruncles as well as the expression of PPARgamma protein in cotyledons were not different between the in vitro and in vivo groups. However, caruncles from the in vitro group had increased expression of PPARgamma protein. The total surface area of endometrium was greater for the in vitro group compared with controls. In contrast, the percentage placentome surface area was decreased in the in vitro group. Fetal villi and binucleate cell volume densities were decreased in placentomes from embryos produced in vitro. The proportional tissue volume of blood vessels in the maternal caruncles was increased in the in vitro group. Furthermore, the ratios of blood vessel volume density-to-placentome surface area were increased in the in vitro group. In conclusion, these findings are consistent with the concept that compensatory mechanisms exist in the vascular beds of placentas from bovine embryos produced in vitro. PMID- 15286037 TI - Mesenteric arterial relaxation to calcitonin gene-related peptide is increased during pregnancy and by sex steroid hormones. AB - The present study investigated whether pregnancy and circulatory ovarian hormones increase the sensitivity of the mesenteric artery to calcitonin gene-related peptide (CGRP)-induced relaxation and possible mechanisms involved in this process. Mesenteric arteries from young adult male rats or female rats (during estrous cycle, after ovariectomy, at Day 20 of gestation, or Postpartum Day 2) were isolated, and the responsiveness of the vessels to CGRP was examined with a small vessel myograph. The CGRP (10(-10) to 10(-7) M) produced a concentration dependent relaxation of norepinephrine-induced contractions in mesenteric arteries of all groups. Arterial relaxation sensitivity to CGRP was significantly (P < 0.05) greater in female rats compared with male rats. Pregnancy increased the sensitivity to CGRP significantly (P < 0.05) compared to ovariectomized and Postpartum Day 2 rats. In pregnant rats, CGRP-receptor antagonist, CGRP(8-37), inhibited the relaxation responses produced by CGRP. The CGRP-induced relaxation was not affected by N(G)-nitro-l-arginine methyl ester (nitric oxide inhibitor, 10(-4) M) but was significantly (P < 0.05) attenuated by an inhibitor of guanylate cyclase (1H-[1 , 2 , 4 ]oxadizaolo[4 , 3 -a]quinoxalin-1-one, 10(-5) M). Relaxation responses of CGRP on mesenteric arteries were blocked (P < 0.05) by a cAMP-dependent protein kinase A inhibitor, Rp-cAMPs (10(-5) M). The CGRP induced vasorelaxation was significantly (P < 0.05) attenuated by calcium dependent (tetraethylammonium, 10(-3) M), but not ATP-sensitive (glybenclamide, 10(-5) M), potassium channel blocker. Therefore, the results of the present study suggest that mesenteric vascular sensitivity to CGRP is higher during pregnancy and that cAMP, cGMP, and calcium-dependent potassium channels appear to be involved. Therefore, we propose that CGRP-mediated vasodilation may be important to maintain vascular adaptations during pregnancy. PMID- 15286038 TI - Prolactin mediates photoperiodic immune enhancement: effects of administration of exogenous prolactin on circulating concentrations, receptor expression, and immune function in steers. AB - Changes in photoperiod can significantly impact the physiology of many species. For example, we have observed an improvement in cellular immune function in cattle on short-day photoperiod (SDPP) relative to long-day photoperiod (LDPP). In addition, prolactin (PRL) and PRL receptor (PRL-R) are affected by photoperiod management. Our hypothesis is that the inverse relationship observed between PRL and PRL-R mRNA expression during photoperiod treatment alters the sensitivity of the animal to PRL, thereby affecting the changes in their cellular immune function. The objective of this study was to determine the effects of exogenous PRL on photoperiodic-mediated immune responses. Eight Holstein steers received each of four treatments: LDPP (16L:8D), SDPP (8L:D), SDom (SDPP plus PRL via osmotic minipump for 10 days), and SDinj (SDPP plus PRL via 3x daily injections for 10 days). Steers on SDPP had decreased PRL relative to the other treatments. Expression of PRL-R mRNA was increased in SDPP animals relative to LDPP, SDom, and SDinj. Prior to PRL treatment, SDPP animals had greater lymphocyte proliferation and neutrophil chemotaxis relative to LDPP animals. Following PRL treatment, cellular immune function of SDom and SDinj animals was reduced to the level of LDPP animals. Addition of PRL to the in vitro lymphocyte proliferation did not alter response of LDPP animals but increased proliferation of lymphocytes from SDPP animals. The results of these experiments suggest that an animal's responsiveness to PRL correlate to changes in cellular immune function that occur with photoperiod manipulation. PMID- 15286039 TI - Parathyroid hormone-related protein in preeclampsia: a linkage between maternal and fetal failures. AB - Preeclampsia is a disorder associated with pregnancy that affects both the mother and the fetus. Typical features of the disease are maternal hypertension, proteinuria, and edema as well as fetal growth retardation. Although the etiological details are still being debated, a consensus exists that the starting point is deficient placentation in the first half of pregnancy. The crucial early steps are reduced trophoblast invasiveness and enhanced apoptotic death. In the present review, we demonstrate that parathyroid hormone-related protein is involved not only in the maternal and fetal failures but also in the etiological aspects of the disease. We hypothesize that reduced local production of the peptide is a major causative event. PMID- 15286040 TI - Covalent transfer of heavy chains of inter-alpha-trypsin inhibitor family proteins to hyaluronan in in vivo and in vitro expanded porcine oocyte-cumulus complexes. AB - Previous studies have shown that the heavy chains (HCs) of serum-derived inter alpha-trypsin inhibitor (IalphaI) molecules become covalently linked to hyaluronan (HA) during in vivo mouse cumulus expansion and significantly contribute to cumulus matrix organization. Experiments with mice suggest that the incorporation of such proteins in cumulus matrix appears to be rather complex, involving LH/hCG-induced changes in blood-follicle barrier and functional cooperation between cumulus cells, granulosa cells, and oocyte within the follicle. We demonstrate here that HC-HA covalent complexes are formed during in vivo porcine cumulus expansion as well. Western blot analysis with IalphaI antibody revealed that follicular fluids from medium-sized follicles and those from large follicles unstimulated with hCG contain high levels of all forms of IalphaI family members present in pig serum. The same amount of HCs were covalently transferred from IalphaI molecules to HA when pig oocyte-cumulus complexes (OCCs) were stimulated in vitro with FSH in the presence of pig serum or follicular fluid from unstimulated or hCG-stimulated follicles. In addition, HC-HA coupling activity was stimulated in cumulus cells by FSH treatment also in the absence of oocyte. Collectively, these results indicate that IalphaI molecules can freely cross the blood follicle barrier and that follicular fluid collected at any stage of folliculogenesis can be successfully used instead of serum for improving OCC maturation. Finally, pig cumulus cells show an autonomous ability to promote the incorporation of IalphaI HCs in the cumulus matrix. PMID- 15286041 TI - Primordial germ cell migration in the rat: preliminary evidence for a role of galactosyltransferase. AB - The precise cellular mechanism of primordial germ cell (PGC) migration remains unknown. Cell surface galactosyltransferase (GalTase) is known to play unique roles in the process of locomotion of many migratory cells. With an objective to seek evidence for possible involvement of GalTase in the migratory process of PGC, we evaluated germ cell migration in the rat following experimental modulation of embryonic GalTase activity. Pregnant rats were laparotomized under anesthesia on Day 10 of pregnancy. While embryos of one uterine horn received lysozyme (100 microg/fetus), those of the other received alpha-lactalbumin (LA; 100 microg/fetus), N-acetylglucosamine (GlcNAc; 250 nmole/fetus), uridine 5' monophosphate (UMP; 2.5 micromole/fetus), uridine diphosphate-galactose (UDP-gal; 250 nmole/fetus), or a combination of 250 nmole of UDP-gal and 2.5 micromole of UMP/fetus. Between gestation Days 12 and 14, embryos were dissected out and processed for histochemical localization of PGC on the basis of binding of Dolichos biflorus agglutinin on the surface glycoconjugate of the germ cells. The number of PGC in each embryo was counted. There was a daywise increase in the number of PGC in all groups. As compared with lysozyme-exposed controls, the numbers of PGCs at the day-specific sites on all days of examination were significantly lower in the LA- as well as GlcNAc-exposed groups. UMP or UDP-gal individually exerted little or no influence, while the total PGC count rose significantly over the respective control values under simultaneous exposure to UMP and UDP-gal. The present findings suggest a likely catalytic role of GalTase in the process of germ cell migration. PMID- 15286042 TI - Localization of the chaperone proteins GRP78 and HSP60 on the luminal surface of bovine oviduct epithelial cells and their association with spermatozoa. AB - Upon their transit through the female genital tract, bovine spermatozoa bind to oviduct epithelial cells, where they are maintained alive for long periods of time until fertilization. Although carbohydrate components of the oviduct epithelial cell membrane are involved in these sperm/oviduct interactions, no protein candidate has been identified to play this role. To identify the oviduct factors involved in their survival, sperm cells were preincubated for 30 min with apical membranes isolated from oviduct epithelial cells, washed extensively, and further incubated for up to 12 h in the absence of apical membranes. During this incubation, sperm viability, motility, and acrosomal integrity were improved compared with cells preincubated in the absence of apical membranes. This suggests that, during the 30-min preincubation with apical membrane extracts, either an oviductal factor triggered intracellular events resulting in positive effects on spermatozoa or that such a factor strongly attached to sperm cells to promote a positive action. Similarly, spermatozoa were incubated with apical membranes isolated from oviduct epithelial cells labeled with [35S]-methionine and, upon extensive washes, proteins were separated by two-dimensional (2-D) gel electrophoresis to identify the factors suspected to have beneficial effects on spermatozoa. The six major proteins, according to their signal intensity on the autoradiographic film, were extracted from a 2-D gel of oviduct epithelial cell proteins run in parallel and processed for N-terminal sequencing of the first 15 amino acids. Of these, one was identical to heat shock protein 60 (HSP60) and one to the glucose-regulated protein 78 (GRP78). Their identities and association with spermatozoa were confirmed using an antibody directed against these proteins. This paper reports the localization of both GRP78 and HSP60 on the luminal/apical surface of oviduct epithelial cells, their binding to spermatozoa, and the presence of endogenous HSP60 in the sperm midpiece. PMID- 15286043 TI - Cryopreservation and thawing is associated with varying extent of activation of apoptotic machinery in subsets of ejaculated human spermatozoa. AB - We investigated the impact of cryopreservation and thawing on levels of caspases 3, -8, and -9 activity, intact mitochondrial membrane potential (Deltapsim), and DNA fragmentation in human spermatozoa. Eleven pools of cryopreserved and eight pools of fresh semen samples were examined. Mature and immature fractions were separated on a two-layer density gradient (47% and 90%) and further subdivided based on the externalization of phosphatidylserine and its binding to annexin V labeled superparamagnetic microbeads (ANMB). Levels of activated caspases were assessed using fluorescein-labeled inhibitors of caspases (FLICA), Deltapsim using a lipophilic cationic dye, and DNA fragmentation by the terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) assay. Cryopreservation was significantly associated with activation of caspases-3, -8, and -9, as well as disruption of the mitochondrial membrane potential but no significant changes were observed in DNA fragmentation. In mature sperm, caspase activation was only detected in the ANMB+ fraction, whereas in immature sperm, both ANMB+ and ANMB- fractions showed activated caspase levels. In ANMB+ immature sperm, apoptosis seemed to be triggered by a surface ligand-receptor mechanism as well as by disruption of mitochondria, whereas in ANMB- immature sperm, apoptosis was induced by activation of caspase-9 following loss of intact Deltapsim. These results demonstrate that selection of annexin V-negative mature spermatozoa might be of clinical relevance for fertility preservation, as this sperm fraction shows no activated apoptosis during the cryopreservation process. PMID- 15286044 TI - Time course of air hunger mirrors the biphasic ventilatory response to hypoxia. AB - Determining response dynamics of hypoxic air hunger may provide information of use in clinical practice and will improve understanding of basic dyspnea mechanisms. It is hypothesized that air hunger arises from projection of reflex brain stem ventilatory drive ("corollary discharge") to forebrain centers. If perceptual response dynamics are unmodified by events between brain stem and cortical awareness, this hypothesis predicts that air hunger will exactly track ventilatory response. Thus, during sustained hypoxia, initial increase in air hunger would be followed by a progressive decline reflecting biphasic reflex ventilatory drive. To test this prediction, we applied a sharp-onset 20-min step of normocapnic hypoxia and compared dynamic response characteristics of air hunger with that of ventilation in 10 healthy subjects. Air hunger was measured during mechanical ventilation (minute ventilation = 9 +/- 1.4 l/min; end-tidal Pco(2) = 37 +/- 2 Torr; end-tidal Po(2) = 45 +/- 7 Torr); ventilatory response was measured during separate free-breathing trials in the same subjects. Discomfort caused by "urge to breathe" was rated every 30 s on a visual analog scale. Both ventilatory and air hunger responses were modeled as delayed double exponentials corresponding to a simple linear first-order response but with a separate first-order adaptation. These models provided adequate fits to both ventilatory and air hunger data (r(2) = 0.88 and 0.66). Mean time constant and time-to-peak response for the average perceptual response (0.36 min(-1) and 3.3 min, respectively) closely matched corresponding values for the average ventilatory response (0.39 min(-1) and 3.1 min). Air hunger response to sustained hypoxia tracked ventilatory drive with a delay of approximately 30 s. Our data provide further support for the corollary discharge hypothesis for air hunger. PMID- 15286045 TI - Cardiac remodeling and functional adaptations consecutive to altitude training in rats: implications for sea level aerobic performance. AB - This study questioned the effect of living and training at moderate altitude on cardiac morphological and functional adaptations and tested the incidences of potential specific adaptations compared with aerobic sea level training on maximal left ventricular performance. Sea level-native rats were randomly assigned to N (living in normoxia), NT (living and training 5 days/wk for 5 wk in normoxia), CH (living in hypoxia, 2,800 m), and CHT (living and training 5 days/wk for 5 wk in hypoxia, 2,800 m) groups. Cardiac adaptations were evaluated throughout the study period by Doppler echocardiography. Maximal stroke volume (LV(SVmax)) was measured during volume overloading before and after the study period. Finally, at the end of the study period, passive pressure-volume relationships on isolated heart and cardiac weighing were obtained. Altitude training resulted in a specific left ventricular (LV) remodeling compared with NT, characterized by an increase in wall thicknesses without any alteration in internal dimensions. These morphological adaptations associated with hypoxia induced alterations in pulmonary outflow and preload conditions led to a decrease in LV filling and subsequently no improvement in LV performance during resting physiological conditions in CHT compared with NT. Such a lack of improvement was confirmed during volume overloading that simulated maximal effort (LV(SVmax) pretest: NT = 0.58 +/- 0.05, CHT = 0.57 +/- 0.08 ml; posttest: NT = 0.72 +/- 0.06, CHT = 0.58 +/- 0.07 ml; NT vs. CHT in posttest session, P < 0.05). Maximal aerobic velocities increased to the same extent in NT and CHT rats despite marked polycythemia in the latter. The lack of LV(SVmax) improvement resulting from altitude training-induced cardiac morphological and functional adaptations could be responsible for this phenomenon. PMID- 15286046 TI - Lingual, splanchnic, and systemic hemodynamic and carbon dioxide tension changes during endotoxic shock and resuscitation. AB - Sublingual and intestinal mucosal blood flow and Pco(2) were studied in a canine model of endotoxin-induced circulatory shock and resuscitation. Sublingual Pco(2) (Ps(CO(2))) was measured by using a novel fluorescent optrode-based technique and compared with lingual measurements obtained by using a Stowe-Severinghaus electrode [lingual Pco(2) (Pl(CO(2)))]. Endotoxin caused parallel changes in cardiac output, and in portal, intestinal mucosal, and sublingual blood flow (Q(s)). Different blood flow patterns were observed during resuscitation: intestinal mucosal blood flow returned to near baseline levels postfluid resuscitation and decreased by 21% after vasopressor resuscitation, whereas Q(s) rose to twice that of the preshock level and was maintained throughout the resuscitation period. Electrochemical and fluorescent Pco(2) measurements showed similar changes throughout the experiments. The shock-induced increases in Ps(CO(2)) and Pl(CO(2)) were nearly reversed after fluid resuscitation, despite persistent systemic arterial hypotension. Vasopressor administration induced a rebound of Ps(CO(2)) and Pl(CO(2)) to shock levels, despite higher cardiac output and Q(s), possibly due to blood flow redistribution and shunting. Changes in Pl(CO(2)) and Ps(CO(2)) paralleled gastric and intestinal Pco(2) changes during shock but not during resuscitation. We found that the lingual, splanchnic, and systemic circulations follow a similar pattern of blood flow variations in response to endotoxin shock, although discrepancies were observed during resuscitation. Restoration of systemic, splanchnic, and lingual perfusion can be accompanied by persistent tissue hypercarbia, mainly lingual and intestinal, more so when a vasopressor agent is used to normalize systemic hemodynamic variables. PMID- 15286047 TI - Regulation of fuel metabolism by preexercise muscle glycogen content and exercise intensity. AB - To date, the results of studies that have examined the effects of altering preexercise muscle glycogen content and exercise intensity on endogenous carbohydrate oxidation are equivocal. Differences in the training status of subjects between investigations may, in part, explain these inconsistent findings. Accordingly, we determined the relative effects of exercise intensity and carbohydrate availability on patterns of fuel utilization in the same subjects who performed a random order of four 60-min rides, two at 45% and two at 70% of peak O(2) uptake (Vo(2 peak)), after exercise-diet intervention to manipulate muscle glycogen content. Preexercise muscle glycogen content was 596 +/- 43 and 202 +/- 21 mmol/kg dry mass (P < 0.001) for high-glycogen (HG) and low glycogen (LG) conditions, respectively. Respiratory exchange ratio was higher for HG than LG during exercise at both 45% (0.85 +/- 0.01 vs. 0.74 +/- 0.01; P < 0.001) and 70% (0.90 +/- 0.01 vs. 0.79 +/- 0.01; P < 0.001) of Vo(2 peak). The contribution of whole body muscle glycogen oxidation to energy expenditure differed between LG and HG for exercise at both 45% (5 +/- 2 vs. 45 +/- 5%; P < 0.001) and 70% (25 +/- 3 vs. 60 +/- 3%; P < 0.001) of Vo(2 peak). Yet, despite marked differences in preexercise muscle glycogen content and its subsequent utilization, rates of plasma glucose disappearance were similar under all conditions. We conclude that, in moderately trained individuals, muscle glycogen availability (low vs. high) does not influence rates of plasma glucose disposal during either low- or moderate-intensity exercise. PMID- 15286048 TI - Fluctuations in plantar flexion force are reduced after prolonged tendon vibration. AB - The purpose of the study was to examine the effect of prolonged vibration on the force fluctuations during a force-matching task performed at low-force levels. Fourteen young healthy men performed a submaximal force-matching task of isometric plantar flexion before and after Achilles tendon vibration (n = 8, vibration subjects) or lying without vibration (n = 6, control subjects) for 30 min. The target forces were 2.5-10% of the previbration maximal voluntary contraction force. The standard deviation of force decreased by a mean of 29 +/- 20% across target forces after vibration, whereas it did not decrease significantly in control subjects (-5 +/- 12%). This change was significantly greater compared with control subjects (P < 0.01 for both). Power spectral density of the force was predominantly composed of signals of low-frequency bandwidth (200 microM), CMPF appears to also inhibit the enzymatic metabolism of Ery. In contrast, IS did not significantly inhibit the hepatocyte uptake of Ery, even at the highest concentration (800 microM) tested, but reduced metabolite generation (p < 0.001). The other uremic toxins, HA, IA, IG, and GSA, did not affect either hepatic uptake or microsomal metabolism of Ery. CMPF, IS, and HA were shown not to inhibit differential P-glycoprotein transport of Ery in cellular systems. Our results suggest that CMPF can directly inhibit the uptake of Ery by inhibiting Oatp2, whereas IS is more likely to inhibit the enzymatic metabolism of Ery. PMID- 15286057 TI - Pharmaco-epidemiology: what do (and don't) we know about utilisation and impact of psychotropic medications in real-life conditions? PMID- 15286056 TI - Efficacy of topical non-steroidal anti-inflammatory drugs in the treatment of osteoarthritis: meta-analysis of randomised controlled trials. AB - OBJECTIVE: To assess the efficacy of topical non-steroidal anti-inflammatory drugs (NSAIDs) in the treatment of osteoarthritis. DATA SOURCES: Medline, Embase, Scientific Citation Index, CINAHL, Cochrane Library, and abstracts from conferences. REVIEW METHODS: Inclusion criterion was randomised controlled trials comparing topical NSAIDs with placebo or oral NSAIDs in osteoarthritis. Effect size was calculated for pain, function, and stiffness. Rate ratio was calculated for dichotomous data such as clinical response rate and adverse event rate. Number needed to treat to obtain the clinical response was estimated. Quality of trial was assessed, and sensitivity analyses were undertaken. RESULTS: Topical NSAIDs were superior to placebo in relieving pain due to osteoarthritis only in the first two weeks of treatment. Effect sizes for weeks 1 and 2 were 0.41 (95% confidence interval, 0.16 to 0.66) and 0.40 (0.15 to 0.65), respectively. No benefit was observed over placebo in weeks 3 and 4. A similar pattern was observed for function, stiffness, and clinical response rate ratio and number needed to treat. Topical NSAIDs were inferior to oral NSAIDs in the first week of treatment and associated with more local side effects such as rash, itch, or burning (rate ratio 5.29, 1.14 to 24.51). CONCLUSION: Randomised controlled trials of short duration only (less than four weeks) have assessed the efficacy of topical NSAIDs in osteoarthritis. After two weeks there was no evidence of efficacy superior to placebo. No trial data support the long term use of topical NSAIDs in osteoarthritis. PMID- 15286058 TI - There is only one functional somatic syndrome. PMID- 15286059 TI - Impact of stressful life events, familial loading and their interaction on the onset of mood disorders: study in a high-risk cohort of adolescent offspring of parents with bipolar disorder. AB - BACKGROUND: Stressful life events are established as risk factors for the onset of mood disorders, but few studies have investigated their impact on the development of mood disorders in adolescents. AIMS: To study the effect of life events on the development of mood disorders in the offspring of parents with bipolar disorder, with respect to the possibility of a decay effect and modification by familial loading. METHOD: In a high-risk cohort of 140 Dutch adolescent offspring of parents with bipolar disorder, we assessed life events, current and past DSM-IV diagnoses and familial loading. To explore their interaction and impact on mood disorder onset, we constructed four different models and used a multivariate survival analysis with time-dependent covariates. RESULTS: The relationship between life events and mood disorder was described optimally with a model in which the effects of life events gradually decayed by 25% per year. The effect of life event load was not significantly stronger in the case of high familial loading. CONCLUSIONS: Independent of familial loading, life events increase the liability to mood disorders in children of patients with bipolar disorder but the effects slowly diminish with time. PMID- 15286060 TI - Vascular disease/risk and late-life depression in a Korean community population. AB - BACKGROUND: Associations between vascular risk factors and late-life depression are controversial. AIMS: To investigate the association between measures of vascular disease/risk and depression and confounding and effect modification by APOE genotype and cognitive function. METHOD: In a Korean community population aged 65+ (n=732), diagnosis of depression (Geriatric Mental State Schedule) and information on vascular status, disability, APOE genotype and cognitive function were obtained. RESULTS: Previous stroke and lower high-density lipoprotein cholesterol level (but neither hypertension nor diabetes) were significantly associated with depression (independently of disability and cognitive function). These associations were stronger in participants with borderline cognitive impairment, although not to a significant extent. CONCLUSIONS: Except for previous stroke and an atherogenic lipid profile, associations between depression and other common risk factors for cerebrovascular disease were not evident. PMID- 15286061 TI - Premorbid adjustment in first-episode non-affective psychosis: distinct patterns of pre-onset course. AB - BACKGROUND: Knowledge about premorbid development in psychosis can shed light upon theories about aetiology and schizophrenic heterogeneity, and form a basis for early detection initiatives. AIMS: To identify and validate patterns of premorbid functioning in first-episode psychosis. METHOD: The Premorbid Adjustment Scale was used to examine 335 patients. RESULTS: Social and academic function constituted fairly independent dimensions. Cluster analysis identified groups varying both in level and course. Patients with a stable social course compared with a deteriorating one had a shorter duration of untreated psychosis, were older, had more friends and less negative symptoms. Good childhood academic function correlated with more education, more meaningful activities and better working memory. Patients with a stable academic course were older at admission. CONCLUSIONS: Patterns of premorbid development suggest both neurodevelopmental and neuroregressive pathways to illness. PMID- 15286062 TI - War-related psychological stressors and risk of psychological disorders in Australian veterans of the 1991 Gulf War. AB - BACKGROUND: Questions remain about the long-term health impacts of the 1991 Gulf War on its veterans. AIMS: To measure psychological disorders in Australian Gulf War veterans and a military comparison group and to explore any association with exposure to Gulf War-related psychological stressors. METHOD: Prevalences of DSM IV psychological disorders were measured using the Composite International Diagnostic Interview. Gulf War-related psychological stressors were measured using a service experience questionnaire. RESULTS: A total of 31% of male Gulf War veterans and 21% of the comparison group met criteria for a DSM-IV disorder first present in the post-Gulf War period. The veterans were at greater risk of developing post-Gulf War anxiety disorders including post-traumatic stress disorder, affective disorders and substance use disorders. The prevalence of such disorders remained elevated a decade after deployment. The findings can be explained partly as a 'war-deployment effect'. There was a strong dose-response relationship between psychological disorders and number of reported Gulf War related psychological stressors. CONCLUSIONS: Service in the 1991 Gulf War is associated with increased risk of psychological disorders and these are related to stressful experiences. PMID- 15286063 TI - Psychological impact of severe acute respiratory syndrome on health workers in a tertiary hospital. AB - BACKGROUND: The sudden emergence of severe acute respiratory syndrome (SARS) caused international anxiety owing to its highly contagious and pandemic transmission. Health workers are vulnerable and are at high risk of infection. AIMS: To assess SARS-related stress and its immediate psychological impact and responses among health workers. METHOD: Health workers in a tertiary hospital affected by SARS were invited to complete a questionnaire designed to evaluate exposure experience, psychological impact and psychiatric morbidity. The risk and rates of psychiatric morbidity were estimated for exposure experience. RESULTS: Altogether, 1257 health workers successfully completed the survey. In the initial phase of the outbreak, when the infection was spreading rapidly, feelings of extreme vulnerability, uncertainty and threat to life were perceived, dominated by somatic and cognitive symptoms of anxiety. During the 'repair' phase, when the infection was being brought under control, depression and avoidance were evident. The estimated prevalence of psychiatric morbidity measured by the Chinese Health Questionnaire was about 75%. CONCLUSIONS: The outbreak of SARS could be regarded as an acute episode of a bio-disaster, leading to a significantly high rate of psychiatric morbidity. PMID- 15286064 TI - Childhood experience and health care use in adulthood: nested case--control study. AB - BACKGROUND: Few studies have considered the role of childhood experiences in adult health care use. AIMS: To examine the hypotheses that individuals frequently attending primary care report childhood adversities and illness exposures more commonly than the remainder of patients and that any association is independent of adult psychiatric disorder. METHOD: A nested case-control study was carried out in a single general practice in Manchester, UK. Fifty frequent attenders (randomly selected from adult patients in the top decile of consultation frequency) and fifty normal attenders (randomly selected from the remainder of adult patients) underwent a structured psychiatric interview and a detailed, semi-structured interview of childhood experience. RESULTS: There was a strong association between frequent attendance and childhood experiences. Multivariate analysis suggested that reported childhood illness exposures and reports of childhood adversity were each associated independently with adult consultation behaviour, even after adjustment for adult psychiatric disorder. CONCLUSIONS: Interventions for high users of health care may need to address childhood experiences of illness and childhood adversities, as well as adult psychiatric disorder. PMID- 15286065 TI - Behavioural interventions in the rehabilitation of acute v. chronic non-organic (conversion/factitious) motor disorders. AB - BACKGROUND: Repeated case series have documented the effectiveness of multidisciplinary in-patient behavioural treatment for conversion disorders. However, in the absence of controlled research, treatment success could be attributed to providing patients with a face-saving opportunity to get better. AIMS: The present study contrasts two behavioural treatments to elucidate the factors underlying successful in-patient rehabilitation of this population. METHOD: Thirty-nine patients underwent a standard behavioural programme. Using a crossover design, patients who did not improve underwent a strategic-behavioural treatment in which they and their families were told that full recovery constituted proof of an organic aetiology whereas failure to recover was definitive proof of a psychiatric aetiology. RESULTS: Chart review indicated that the standard behavioural treatment was effective for 8/9 'acute' patients but only for 1/28 'chronic' patients. Of the 21 patients with chronic motor disorder who then under went the strategic-behavioural intervention, 13 were symptom-free at discharge. CONCLUSIONS: The strategic intervention was superior to standard behavioural treatment for patients with chronic motor disorder. Treatment components previously deemed critical for the effectiveness of behavioural treatment may be unnecessary. PMID- 15286066 TI - Prolactin levels in antipsychotic treatment of patients with schizophrenia carrying the DRD2*A1 allele. AB - BACKGROUND: Hyperprolactinaemia induced by D(2) dopamine receptor antagonist antipsychotic medication can result in significant health problems. AIMS: To examine the role of DRD2 polymorphism on prolactin levels in patients treated with antipsychotic medication. METHOD: Antipsychotic drugs with different degrees of D(2) receptor binding were given to 144 patients with schizophrenia. Serum prolactin levels were obtained and Taq1A DRD2 alleles were determined. RESULTS: Prolactin levels increased across medication groups reflecting increasingly tight D(2) receptor binding (clozapine, olanzapine, typical antipsychotics and risperidone). In the combined medication group, patients with the DRD2(*)A1allele had 40% higher prolactin levels than patients without this allele. In patients treated with clozapine (the loosest D(2) receptor binding agent), patients with the DRD2(*)A1allele had prolactin levels twice those of patients without this allele. CONCLUSIONS: Patients with the DRD2A1 allele receiving antipsychotic medications had higher prolactin levels and were overrepresented among those with hyperprolactinaemia, suggesting greater functional D(2) receptor binding in this group. PMID- 15286067 TI - Testing for diabetes in hospitalised patients prescribed antipsychotic drugs. AB - BACKGROUND: Studies using computer databases suggest that atypical antipsychotic agents are more likely to be associated with diabetes than are conventional drugs. AIMS: To discover the extent of testing for diabetes mellitus in hospital in-patients prescribed antipsychotics. METHOD: Prescription charts were screened to identify patients prescribed antipsychotics. Case notes were then searched for evidence of testing for diabetes. RESULTS: In all, 606 patients were prescribed antipsychotics, of whom 250 (41.3%) had evidence of prior testing for diabetes. Patients prescribed atypicals were 40% more likely to have been tested than those prescribed conventional drugs (RR =1.4,95% CI1.1-1.9). Adjusted odds ratios v. conventional antipsychotics for conventional antipsychotics for testing were significantly higher for clozapine (OR=4.64,95% CI 2.42-8.90), olanzapine (OR=1.85,95% CI1.04-3.30) and antipsychotic polypharmacy (OR=2.96,95% CI1.59 5.52). CONCLUSIONS: Testing for diabetes was undertaken in less than half of the patients studied. Testing was more common in those receiving atypical antipsychotics. Apparent differences in claimed causal association of the use of some antipsychotics with diabetes may in part reflect different rates of testing. PMID- 15286068 TI - Effect of a medication management training package for nurses on clinical outcomes for patients with schizophrenia: cluster randomised controlled trial. AB - BACKGROUND: Non-compliance attenuates the efficacy of treatments for physical and mental disorders. AIMS: To assess the effectiveness of a medication management training package for community mental health nurses (CMHNs) in improving compliance and clinical outcomes in patients with schizophrenia. METHOD: Pragmatic randomised controlled trial. Sixty CMHNs in geographical clusters were assigned randomly to medication management training or treatment as usual. Each CMHN identified two patients on their case-load who were assessed at baseline and again after 6 months by a research worker. The primary efficacy outcome of interest was psychopathology, measured using the Positive and Negative Syndrome Scale (PANSS). RESULTS: Nurses who had received medication management training produced a significantly greater reduction in patients'overall psychopathology compared with treatment as usual at the end of the 6-month study period (change in PANSS total scores: medication management -16.62, treatment as usual 1.17; difference -17.79; 95% CI -24.12 to -11.45; P<0.001). CONCLUSIONS: Medication management training for CMHNs is effective in improving clinical outcomes in patients with schizophrenia. PMID- 15286069 TI - Systematic assessments of need and care planning in severe mental illness: cluster randomised controlled trial. AB - BACKGROUND: Surveys have shown high levels of unmet need in representative samples of people with severe mental illness. Introducing standardised needs assessment into the care planning process might reduce these needs and improve outcome. AIMS: To determine whether feedback from a standardised assessment of need enhances the effectiveness of care planning and whether exposing care coordinators to feedback on some patients improves their care of other patients. METHOD: A single-blind, cluster randomised trial involving a within-cluster individually randomised arm: patients' needs were evaluated using the Cardinal Needs Schedule and the findings were fed back to their care coordinators under the care programme approach. A total of 304 patients were recruited from 72 care coordinators and 242 patients (79.6%) were reassessed at 12 months. RESULTS: The only significant effect of the intervention was on patient satisfaction. Patients cluster-randomised to receive feedback were more satisfied than controls, but patients individually randomised to receive feedback were not. CONCLUSIONS: Standardised needs assessment did not substantially enhance care planning in this trial. However, giving care coordinators some experience of feedback from a standardised assessment of need could improve satisfaction. PMID- 15286070 TI - Social cognition and face processing in schizophrenia. AB - Summary Studies of face processing have begun to elucidate the brain regions involved in social cognition, which include frontal and temporal regions known to be reduced in volume in schizophrenia. In this case-control study participants with schizophrenia (n=20) showed marked deficits in their ability to interpret social cues from faces, and those experiencing positive symptoms were impaired in recognising even basic facial emotions. PMID- 15286071 TI - Value of early intervention in psychosis. PMID- 15286072 TI - Value of early intervention in psychosis. PMID- 15286073 TI - Testosterone and psychosis. PMID- 15286074 TI - Neurosurgery for mental disorder. PMID- 15286075 TI - Stigma and somatisation. PMID- 15286076 TI - Disability and post-traumatic stress. PMID- 15286077 TI - In defence of complainants. PMID- 15286078 TI - GHB and date rape. PMID- 15286079 TI - Creative debate misses the point. PMID- 15286080 TI - Comedians: fun and dysfunctionality. PMID- 15286081 TI - Dok-6, a Novel p62 Dok family member, promotes Ret-mediated neurite outgrowth. AB - Activation of Ret, the receptor-tyrosine kinase for the glial cell line-derived neurotrophic factor (GDNF) family ligands (GFLs), results in the recruitment and assembly of adaptor protein complexes that function to transduce signals downstream of the receptor. Here we identify Dok-6, a novel member of the Dok-4/5 subclass of the p62 Dok family of intracellular adaptor molecules, and characterize its interaction with Ret. Expression analysis reveals that Dok-6 is highly expressed in the developing central nervous system and is co-expressed with Ret in several locations, including sympathetic, sensory, and parasympathetic ganglia, as well as in the ureteric buds of the developing kidneys. Pull-down assays using the Dok-6 phosphotyrosine binding (PTB) domain and GDNF-activated Ret indicate that Dok-6 binds to the phosphorylated Ret Tyr(1062) residue. Moreover, ligand activation of Ret resulted in phosphorylation of tyrosine residue(s) located within the unique C terminus of Dok-6 predominantly through a Src-dependent mechanism, indicating that Dok-6 is a substrate of the Ret-Src signaling pathway. Interestingly, expression of Dok-6 potentiated GDNF-induced neurite outgrowth in GDNF family receptor alpha1 (GFRalpha1)-expressing Neuro2A cells that was dependent upon the C-terminal residues of Dok-6. Taken together, these data identify Dok-6 as a novel Dok-4/5 related adaptor molecule that may function in vivo to transduce signals that regulate Ret-mediated processes such as axonal projection. PMID- 15286082 TI - Identification of distinct gamma-secretase complexes with different APH-1 variants. AB - The gamma-secretase complex catalyzes the final intramembraneous cleavage of the beta-amyloid precursor protein, liberating the neurotoxic amyloid beta-peptide implicated in Alzheimer's disease. Apart from the catalytic subunit presenilin (PS), three additional subunits, nicastrin, APH-1, and PEN-2, have been identified. In mammals, two PS homologues, PS1 and PS2, which are part of distinct gamma-secretase complexes, exist. Likewise, two APH-1 homologues, APH-1a and APH-1b, have been identified. Furthermore, two APH-1a splice forms, APH-1aS and APH-1aL, have been reported. Here we show that both APH-1a splice forms and APH-1b are expressed in peripheral and neuronal cells. APH-1aS, APH-1aL, and APH 1b form separate, proteolytically active gamma-secretase complexes containing either one of the two PSs. Deficiency of APH-1a caused a decrease in nicastrin, PS, and PEN-2 levels and an increase in the levels of APH-1b, whereas deficiency of APH-1b did not affect the levels of APH-1a or the other complex components. Consistent with this finding, we found that deficiency of APH-1a was associated with reduced gamma-secretase activity, whereas deficiency of APH-1b was not. Thus, APH-1b gamma-secretase complexes may fulfill redundant functions. Taken together, our results suggest that, dependent on the tissue expression of the individual subunits, six distinct gamma-secretase complexes composed of the known subunits can exist in human cells. PMID- 15286083 TI - Structural and thermodynamic evidence for a stabilizing role of Nop5p in S adenosyl-L-methionine binding to fibrillarin. AB - In Archaea, fibrillarin and Nop5p form the core complex of box C/D small ribonucleoprotein particles, which are responsible for site-specific 2'-hydroxyl methylation of ribosomal and transfer RNAs. Fibrillarin has a conserved methyltransferase fold and employs S-adenosyl-l-methionine (AdoMet) as the cofactor in methyl transfer reactions. Comparison between recently determined crystal structures of free fibrillarin and fibrillarin-Nop5p-AdoMet tertiary complex revealed large conformational differences at the cofactor-binding site in fibrillarin. To identify the structural elements responsible for these large conformational differences, we refined a crystal structure of Archaeoglobus fulgidus fibrillarin-Nop5p binary complex at 3.5 A. This structure exhibited a pre-formed backbone geometry at the cofactor binding site similar to that when the cofactor is bound, suggesting that binding of Nop5p alone to fibrillarin is sufficient to stabilize the AdoMet-binding pocket. Calorimetry studies of cofactor binding to fibrillarin alone and to fibrillarin-Nop5p binary complex provided further support for this role of Nop5p. Mutagenesis and thermodynamic data showed that a cation-pi bridge formed between Tyr-89 of fibrillarin and Arg 169 of Nop5p, although dispensable for in vitro methylation activity, could partially account for the enhanced binding of cofactor to fibrillarin by Nop5p. Finally, assessment of cofactor-binding thermodynamics and catalytic activities of enzyme mutants identified three additional fibrillarin residues (Thr-70, Glu 88, and Asp-133) to be important for cofactor binding and for catalysis. PMID- 15286084 TI - The Epstein-Barr virus polymerase accessory factor BMRF1 adopts a ring-shaped structure as visualized by electron microscopy. AB - Epstein-Barr virus (EBV) encodes a set of core replication factors used during lytic infection in human cells that parallels the factors used in many other systems. These include a DNA polymerase and its accessory factor, a helicase/primase, and a single strand binding protein. The EBV polymerase accessory factor has been identified as the product of the BMRF1 gene and has been shown by functional assays to increase the activity and processivity of the polymerase. Unlike other members of this class of factors, BMRF1 is also a transcription factor regulating certain EBV genes. Although several polymerase accessory factors, including eukaryotic proliferating cell nuclear antigen, Escherichia coli beta protein, and T4 gene 45 protein have been shown to form oligomeric rings termed sliding clamps, nothing is known about the oligomeric state of BMRF1 or whether it forms a ring. In this work, BMRF1 was purified directly from human cells infected with an adenovirus vector expressing the BMRF1 gene product. The protein was purified to near homogeneity, and examination by negative staining electron microscopy revealed large, flat, ring-shaped molecules with a diameter of 15.5 +/- 0.8 nm and a distinct 5.3-nm diameter hole in the center. The size of these rings is consistent with an oligomer of 6 monomers, nearly twice as large as the trimeric proliferating cell nuclear antigen ring. Unlike the herpes simplex virus UL42 homologue, BMRF1 was found to self-associate in solution. These findings extend the theme of polymerase accessory factors adopting ring-shaped structures and provide an example in which the ring is significantly larger than any previously described sliding clamp. PMID- 15286085 TI - Novel regulation of cystic fibrosis transmembrane conductance regulator (CFTR) channel gating by external chloride. AB - The cystic fibrosis transmembrane conductance regulator (CFTR) is vital for Cl(-) and HCO(3)(-) transport in many epithelia. As the HCO(3)(-) concentration in epithelial secretions varies and can reach as high as 140 mm, the lumen-facing domains of CFTR are exposed to large reciprocal variations in Cl(-) and HCO(3)(-) levels. We have investigated whether changes in the extracellular anionic environment affects the activity of CFTR using the patch clamp technique. In fast whole cell current recordings, the replacement of 100 mm external Cl(-) ((Cl(o)( ))) with HCO(3)(-), Br(-), NO(3)(-), or aspartate(-) inhibited inward CFTR current (Cl(-) efflux) by approximately 50% in a reversible manner. Lowering Cl(o)(-) alone by iso-osmotic replacement with mannitol also reduced Cl(-) efflux to a similar extent. The maximal inhibition of CFTR current was approximately 70%. Raising cytosolic calcium shifted the Cl(-) dose-inhibition curve to the left but did not alter the maximal current inhibition observed. In contrast, a reduction in the internal [Cl(-)] neither inhibited CFTR nor altered the block caused by reduced Cl(o)(-). Single channel recordings from outside-out patches showed that lowering Cl(o)(-) markedly reduced channel open probability with little effect on unitary conductance. Together, these results indicate that alterations in Cl(o)(-) alone and not the Cl(-)/HCO(3)(-) ratio regulate the gating of CFTR. Physiologically, our data have implications for current models of epithelial HCO(3)(-) secretion and for the control of pH at epithelial cell surfaces. PMID- 15286086 TI - Pharmacological and clinical profile of moexipril: a concise review. AB - Angiotensin-converting enzyme (ACE) inhibitors are effective and safe antihypertensive drugs, with the exception of the rare occasion of angioedema. These drugs have demonstrated additional cardiovascular protective effects to their blood pressure lowering, and their combination with the diuretic hydrochlorothiazide potentiates their antihypertensive effectiveness. Moexipril is a long-acting ACE inhibitor suitable for once-daily administration, and like some ACE inhibitors, moexipril is a prodrug and needs to be hydrolyzed in the liver into its active carboxylic metabolite, moexiprilat, to become effective. Moexipril alone and in combination with low-dose hydrochlorothiazide has been shown in clinical trials to be effective in lowering blood pressure and be well tolerated and safe given in single daily doses. In this review, the pharmacological profile of this drug and its clinical usefulness are discussed. PMID- 15286087 TI - Physiological, pharmacokinetic, and pharmacodynamic changes in space. AB - Medications have been taken since the first Mercury flight in 1967 and, since then, have been used for several indications such as space motion sickness, sleeplessness, headache, nausea, vomiting, back pain, and congestion. As the duration of space missions get longer, it is even more likely that astronauts will encounter some of the acute illnesses that are frequently seen on Earth. Microgravity environment induces several physiological changes in the human body. These changes include cardiovascular degeneration, bone decalcification, decreased plasma volume, blood flow, lymphocyte and eosinophil levels, altered hormonal and electrolyte levels, muscle atrophy, decreased blood cell mass, increased immunoglobulin A and M levels, and a decrease in the amount of microsomal P-450 and the activity of some of its dependent enzymes. These changes may be expected to have severe implications on the pharmacokinetic and pharmacodynamic properties of drug substances. PMID- 15286088 TI - Influence of genetic variants in UGT1A1 and UGT1A9 on the in vivo glucuronidation of SN-38. AB - The uridine diphosphate glucuronosyltransferase (UGT) 1A1 and 1A9 isoforms are involved in the phase II biotransformation of the irinotecan metabolite, SN-38. Recently, several variants in the UGT1A1 and UGT1A9 genes have been described with altered functionality in vitro. The aim of this study was to evaluate the functional consequence of the UGT1A1(TA)(7)TAA (UGT1A1(*)28), UGT1A9 766G>A (D256N; UGT1A9(*)5), and UGT1A9 98T>C (M33T; UGT1A9(*)3) variants in Caucasian patients treated with irinotecan. Pharmacokinetic studies were performed after the first course of irinotecan in 47 males and 47 females. The mean (SD) area under the curves (AUCs) of irinotecan and SN-38 were 20,348 +/- 6466 ng x h/mL and 629 +/- 370 ng x h/mL, respectively, which is in line with earlier findings. For UGT1A9(*)5,novariant alleles were observed, whereas for UGT1A9(*)3, 1 patient with the variant allele was found (allele frequency, 0.633%). The distribution of the UGT1A1(*)28 variant showed 44 wild-type patients (Wt), 37 heterozygotes (Het), and 5 homozygotes (Var). The median AUC ratio of SN-38G to SN-38 was significantly reduced in carriers of the variant UGT1A1(*)28 allele (7.00 [Wt] vs. 6.26 [Het] vs. 2.51 [Var]; p =.022). It is concluded that UGT1A9 functional variants are rare in Caucasians and likely to be clinically insignificant in irinotecan regimens. Screening for the UGT1A1(*)28 polymorphism may identify patients with altered SN-38 pharmacokinetics. PMID- 15286089 TI - Lack of attenuation in the antitumor effect of tamoxifen by chronic CYP isoform inhibition. AB - Tamoxifen is a selective estrogen receptor modulator used in estrogen receptor positive breast cancer. Tamoxifen is metabolized to an extremely potent antiestrogen by cytochrome P450 (CYP) 2D6, 2C9, and 3A isoforms. The selective serotonin reuptake inhibitors (SSRIs) are potent inhibitors of these CYPs. Since the prevalence of depression in breast cancer patients is nearly triple that of the general population, it is likely that a subgroup of breast cancer patients will receive long-term treatment with both an SSRI and tamoxifen. A case control design was used to investigate the possibility that a resultant decrease in production of the 4-hydroxy metabolite from chronic inhibition results in the attenuation of the antitumor effect of tamoxifen. Twenty-eight patients without recurrences of breast cancer (controls) were matched to an equal number of cases (recurrences) by cancer stage and year of diagnosis. Data were analyzed on all chronic medication exposure (> 3 months) in both cases and controls classified as to their status as CYP 2D6, 2C9, and 3A inhibitors, substrates, or inducers. No significant difference was found for CYP inhibitor or substrate exposure between cases and controls. Indeed, controls showed a slightly greater exposure to inhibitors of the relevant CYP isoforms compared to cases. These results suggested a trend toward the null hypothesis. It is unlikely that the effect of chronic exposure to potent CYP isoform inhibitors affects the antitumor effect of tamoxifen and its 4-hydroxy metabolite, supporting the safety of the continued practice of concomitant SSRI administration to breast cancer patients with depression. PMID- 15286090 TI - Pharmacokinetics of meloxicam in patients with juvenile rheumatoid arthritis. AB - The pharmacokinetics of a meloxicam suspension were studied in 18 children with juvenile rheumatoid arthritis. Children received a single 0.25-mg/kg dose up to a maximum of 15 mg. Pharmacokinetic parameters after the first dose were calculated by noncompartmental methods. Geometric mean (percent coefficient of variation for geometric mean [gCV]) C(max), AUC(0- infinity ), apparent clearance, apparent volume of distribution, and elimination half-life values were 1.24 microg/mL (47% gCV), 25.6 microg x h/mL (81% gCV), 0.17 mL/min/kg (83% gCV), 0.19 L/kg (63% gCV), and 13.4 hours (54% gCV) in the younger group and 1.89 microg/mL (25% gCV), 35.8 microg x h/mL (21% gCV), 0.12 mL/min/kg (23% gCV), 0.13 L/kg (22% gCV), and 12.7 hours (21% gCV) for the older group, respectively. Area under the curve, volume of distribution, and clearance tended to be higher in the younger group, whereas elimination half-lives were similar. A post hoc comparison to pharmacokinetic data in adults revealed no relevant differences. Thus, a common body weight-normalized dose is considered appropriate for children older than 2 years. PMID- 15286091 TI - Pharmacokinetics of gemtuzumab ozogamicin as a single-agent treatment of pediatric patients with refractory or relapsed acute myeloid leukemia. AB - Gemtuzumab ozogamicin is currently approved to treat CD33-positive acute myeloid leukemia (AML) in first relapse in patients older than age 60 years. The objective of this study was to characterize the pharmacokinetics of gemtuzumab ozogamicin in pediatric patients with relapsed or refractory AML. The study population comprised 29 subjects younger than age 18 with AML in first relapse. Dosages of 6, 7.5, and 9 mg/m(2) were administered during the study. Pharmacokinetic parameters were determined following each dose for hP67.6, total calicheamicin derivatives, and unconjugated calicheamicin derivatives. hP67.6 pharmacokinetic parameters had a consistent and statistically significant change between the first and second doses. Increases in AUC and decreases in both CL and V(ss) from the first dose to the second dose were consistent with those of the adult population. Changes between dose periods for total calicheamicin derivatives and unconjugated calicheamicin derivatives were consistent with those of hP67.6. Changes in pharmacokinetic parameters between dose periods are attributed to saturation of CD33 binding sites and diminished clearance resulting from a lower peripheral blast burden and antigen. Children receiving 9 mg/m(2) had the following hP67.6 pharmacokinetic parameters: C(max), 3.47+/-1.04 mg/L; AUC, 136 +/- 107 mg x h/L; CL, 0.12 +/- 0.15 L/h/m(2); V(ss), 6.5 +/- 5.5 L; and t(1/2), 64 +/- 44 h after their first dose. Mean pharmacokinetic values are similar to values reported in adults. Individual children demonstrated large intersubject variability, similar to adults. The pharmacokinetics of gemtuzumab ozogamicin in pediatric patients closely follow the profile and variability of adult patients. PMID- 15286092 TI - Effects of nicotine on the number and activity of circulating endothelial progenitor cells. AB - Recently, some studies have shown that nicotine increased neovascularization, which involves endothelial progenitor cells (EPCs). The effects of nicotine on EPCs are still unclear at present. Therefore, the authors investigated whether nicotine had influences on EPC number and activity. The EPCs were stimulated with nicotine (to make a series of final concentrations: 10(-12) mol/L, 10(-10) mol/L, 10(-8) mol/L, 10(-6) mol/L, 10(-4) mol/L) or vehicle control for the respective time points(12, 18, 24, 32, and 48 hours). The EPCs were characterized as adherent cells double positive for DiLDL uptake and lectin binding by direct fluorescent staining under a laser-scanning confocal microscope. They were further documented by demonstrating the expression of KDR, VEGFR-2, and AC133 with flow cytometry. The EPC proliferation, migration, and in vitro vasculogenesis activity were assayed with the 3-(4, 5-dimethylthiazol-2-yl)-2, 5 diphenyltetrazolium bromide assay; the modified Boyden chamber assay; and the in vitro vasculogenesis kit, respectively. The EPC adhesion assay was performed by replating those on fibronectin-coated dishes and then counting the adherent cells. As a result, nicotine dose dependently increased the EPC number and the proliferative, migratory, adhesive, and in vitro vasculogenesis capacity at nicotine concentrations of 10(-12) to 10(-8) mol/L. The peak effects on EPCs were observed at concentrations of nicotine 10(-8) mol/L, similar to those in the blood of smokers. In addition, nicotine (10(-8) mol/L) time dependently increased the EPC number and activity. However, cytotoxicity was seen at higher nicotine concentrations (> 10(-6) mol/L). In conclusion, nicotine had complex effects on EPCs: nicotine might induce the augmentation of EPCs with enhanced functional activity at relatively low concentrations. However, cytotoxicity was seen at higher nicotine concentrations. PMID- 15286094 TI - Pharmacokinetics of M100240 and MDL 100,173, a dual angiotensin-converting enzyme/neutral endopeptidase inhibitor, in healthy young and elderly volunteers. AB - M100240 is an acetate thioester of MDL 100,173-a dual angiotensin-converting enzyme (ACE)/neutral endopeptidase (NEP) inhibitor-in phase II development. The pharmacokinetics of M100240 and MDL 100,173 were compared in young and elderly subjects. Pharmacokinetic data were obtained from 12 young (ages 18-45 years, 10 male, 2 female) and 12 elderly (ages 65-85 years, 7 male, 5 female) healthy subjects in a parallel-group, open-label study. Following an overnight fast, subjects received a single 25-mg oral dose of M100240. Serial plasma concentrations of M100240 and MDL 100,173 were determined using a validated liquid chromatography/tandem mass spectrometry (LC/MS/MS) method, and pharmacokinetic parameters were calculated with noncompartmental methods. Single dose treatment with M100240 was well tolerated in both groups of subjects, with no clinically significant changes in vital signs, ECG recordings, or laboratory safety parameters. M100240 was rapidly absorbed and converted to MDL 100,173, with M100240 concentrations no longer detectable at 3 to 4 hours postdose in both groups. The pharmacokinetics of the pharmacologically active MDL 100,173 were similar for both groups. Although maximum concentrations of M100240 were generally higher in elderly versus young subjects (C(max) 0.48 ng/mL vs. 0.17 ng/mL), systemic availability of M100240 was quite low and variable with plasma, and this apparent difference in parent drug exposure is unlikely to have important clinical implications. No age-related differences in the pharmacokinetic parameters of MDL 100,173 (C(max) 8.16 vs. 9.62 ng/mL, t(max) 1.25 vs. 1.5 h, AUC((0-last)) 81.6 vs. 72.2 ng x h/mL) were observed between young and elderly subjects, respectively. In conclusion, there are no age-related differences in the pharmacokinetics of MDL 100,173 between young and elderly subjects. PMID- 15286093 TI - Central effects of fexofenadine and cetirizine: measurement of psychomotor performance, subjective sleepiness, and brain histamine H1-receptor occupancy using 11C-doxepin positron emission tomography. AB - Histamine H1-receptor (H1R) antagonists, or antihistamines, often induce sedative side effects when used for the treatment of allergic disorders. This study compared the sedative profiles of the second-generation antihistamines, fexofenadine and cetirizine, using 3 different criteria: subjective sleepiness evaluated by the Stanford Sleepiness Scale, objective psychomotor tests (simple and choice reaction time tests and visual discrimination tests at 4 different exposure durations), and measurement of histamine H1-receptor occupancy (H1RO) in the brain. Subjective sleepiness and psychomotor performance were measured in 20 healthy Japanese volunteers at baseline and 90 min after administration of fexofenadine 120 mg or cetirizine 20 mg in a double-blind, placebo-controlled crossover study. Hydroxyzine 30 mg was included as a positive control. H1RO was measured using positron emission tomography (PET) with (11)C-doxepin in 12 of the 20 subjects, and a further 11 volunteers were recruited to act as controls. In psychomotor tests, fexofenadine was not significantly different from placebo and significantly less impairing than cetirizine on some tasks, as well as significantly less impairing than hydroxyzine on all tasks. For subjective sleepiness, fexofenadine was not significantly different from placebo, whereas cetirizine showed a trend toward increased sleepiness compared with fexofenadine and placebo. H1RO was negligible with fexofenadine (-0.1%) but moderately high with cetirizine (26.0%). In conclusion, fexofenadine 120 mg is distinguishable from cetirizine 20 mg, as assessed by H1RO and psychomotor testing. PMID- 15286095 TI - Safety, tolerability, and pharmacokinetic profile of BIA 2-093, a novel putative antiepileptic, in a rising multiple-dose study in young healthy humans. AB - This was a double-blind, randomized, placebo-controlled study to investigate rising oral doses of BIA 2-093 (S-(-)-10-acetoxy-10,11-dihydro-5H dibenz/b,f/azepine-5-carboxamide), a putative new antiepileptic drug. Within each of 4 dosage groups of 8 healthy male adult subjects, 2 subjects were randomized to receive placebo, and the remaining 6 subjects were randomized to receive BIA 2 093 (200 mg bid, 400 mg qd, 800 mg qd, and 1200 mg qd) for 8 days. Concentrations of BIA 2-093 in plasma or urine were generally not measurable. Median maximum plasma concentrations of the major metabolite (licarbazepine, (+/-)-10,11-dihydro 10-hydroxy-5H-dibenz/b,f/azepine-5-carboxamide) were attained (t(max)) at 2 to 3 h postdose; thereafter, plasma concentrations declined with a mean apparent terminal half-life of 9 to 13 h following repeated dosing. The extent of systemic exposure to licarbazepine increased in an approximately dose-proportional manner following single and repeated administration. Licarbazepine accumulated in plasma following repeated administration of BIA 2-093; the mean extent of accumulation (R(O), calculated from AUC(0-tau) (day 8)/AUC(0-tau) (day 1)) was 3.0 after repeated, twice-daily dosing and 1.4 to 1.7 after once-daily dosing. Steady-state plasma licarbazepine concentrations were attained at 4 to 5 days of once- or twice-daily dosing, consistent with an effective half-life on the order of 20 to 24 h. The mean renal clearance of licarbazepine from plasma was approximately 20 to 30 mL/min, which is low compared with the glomerular filtration rate. The total amount of licarbazepine recovered in urine was approximately 20% within 12 h postdose and 40% within 24 h postdose. All adverse events were mild in severity, except for 1 case of somnolence of moderate severity, which occurred in a subject receiving 1200 mg BIA 2-093. The incidence of adverse events was similar between all treatment groups, including placebo. There were no serious adverse events. In conclusion, BIA 2-093 was well tolerated and appeared to be rapidly and extensively metabolized to licarbazepine following single and repeated administration to healthy young subjects. PMID- 15286096 TI - Renal interaction between itraconazole and cimetidine. AB - Renal drug interactions can result from competitive inhibition between drugs that undergo extensive renal tubular secretion by transporters such as P-glycoprotein (P-gp). The purpose of this study was to evaluate the effect of itraconazole, a known P-gp inhibitor, on the renal tubular secretion of cimetidine in healthy volunteers who received intravenous cimetidine alone and following 3 days of oral itraconazole (400 mg/day) administration. Glomerular filtration rate (GFR) was measured continuously during each study visit using iothalamate clearance. Iothalamate, cimetidine, and itraconazole concentrations in plasma and urine were determined using high-performance liquid chromatography/ultraviolet (HPLC/UV) methods. Renal tubular secretion (CL(sec)) of cimetidine was calculated as the difference between renal clearance (CL(r)) and GFR (CL(ioth)) on days 1 and 5. Cimetidine pharmacokinetic estimates were obtained for total clearance (CL(T)), volume of distribution (Vd), elimination rate constant (K(el)), area under the plasma concentration-time curve (AUC(0-240 min)), and average plasma concentration (Cp(ave)) before and after itraconazole administration. Plasma itraconazole concentrations following oral dosing ranged from 0.41 to 0.92 microg/mL. The cimetidine AUC(0-240 min) increased by 25% (p < 0.01) following itraconazole administration. The GFR and Vd remained unchanged, but significant reductions in CL(T) (655 vs. 486 mL/min, p < 0.001) and CL(sec) (410 vs. 311 mL/min, p = 0.001) were observed. The increased systemic exposure of cimetidine during coadministration with itraconazole was likely due to inhibition of P-gp mediated renal tubular secretion. Further evaluation of renal P-gp-modulating drugs such as itraconazole that may alter the renal excretion of coadministered drugs is warranted. PMID- 15286097 TI - No pharmacokinetic or pharmacodynamic interaction between atorvastatin and the oral direct thrombin inhibitor ximelagatran. AB - In this randomized, 2-way crossover study, the potential for interaction was investigated between atorvastatin and ximelagatran, an oral direct thrombin inhibitor. Healthy female and male volunteers (n = 16) received atorvastatin 40 mg as a single oral dose and, in a separate study period, ximelagatran 36 mg twice daily for 5 days plus a 40-mg oral dose of atorvastatin on the morning of day 4. In the 15 subjects completing the study, no pharmacokinetic interaction was detected between atorvastatin and ximelagatran for all parameters investigated, including melagatran (the active form of ximelagatran) area under the plasma concentration versus time curve (AUC) and maximum plasma concentration, atorvastatin acid AUC, and AUC of active 3-hydroxy-3-methyl glutaryl-coenzyme-A (HMG-CoA) reductase inhibitors. Atorvastatin did not alter the melagatran-induced prolongation of the activated partial thromboplastin time, and both drugs were well tolerated when administered in combination. In conclusion, no pharmacokinetic or pharmacodynamic interaction between atorvastatin and ximelagatran was observed in this study. PMID- 15286098 TI - No pharmacokinetic or pharmacodynamic interaction between digoxin and the oral direct thrombin inhibitor ximelagatran in healthy volunteers. AB - The interaction potential of digoxin and ximelagatran, an oral direct thrombin inhibitor being developed for the prevention and treatment of thromboembolic disease, was investigated in this randomized, double-blind, 2-way crossover study. On 2 separate occasions, healthy female and male volunteers (n = 16) received ximelagatran 36 mg or placebo twice daily for 8 days separated by a 4- to 14-day washout period. All volunteers received a single oral dose of digoxin 0.5 mg on day 4 of both study periods. No interaction between ximelagatran and digoxin was detected in the pharmaco-kinetic parameters (using a 90% confidence interval [CI] of least squares mean estimate ratios), including melagatran (the active form of ximelagatran) AUC(tau) and C(max) and digoxin AUC(t) and C(max). Digoxin did not alter the melagatran-induced prolongation of the activated partial thromboplastin time, and both drugs were well tolerated when administered in combination. In conclusion, no pharmacokinetic or pharmacodynamic interaction between digoxin and ximelagatran was observed in this study. PMID- 15286099 TI - Ethics education in dental school: continuing the conversation. PMID- 15286100 TI - Dentistry for persons with special needs: how should it be recognized? PMID- 15286101 TI - A call for increased education in genetics for dental health professionals. PMID- 15286102 TI - Status of genetics education in U.S. dental schools. AB - Genomics research is rapidly increasing our understanding of the genetic basis of normal and abnormal growth, development, and disease. Genetic information and technologies are also being applied to develop new diagnostic and treatment strategies. Many diseases with dental, oral, and craniofacial manifestations have a genetic basis. Effective clinical application of genomics to oral medicine will depend on the education of health care professionals, the general public, and policymakers. Dentists must understand genetics to provide accurate information to patients and be able to discuss benefits and limitations of the biological, clinical, and ethical issues related to genomic-based health care. Genetics education in dental schools will significantly impact the integration of genetics into oral medicine. Fifty-three U.S. dental schools completed a survey in 2001 to assess the status of genetics curricula in dental schools in the United States. Ninety-four percent of schools did not require genetics education for entry to dental school, and a formal genetics course was conducted in only eight of the fifty-three schools (15 percent). The genetics education currently offered to undergraduate dental students is not standardized, and the content varies considerably among schools. These findings suggest more emphasis on genetics education is needed in U.S. dental schools. PMID- 15286103 TI - Need for genetics education in U.S. dental and dental hygiene programs. AB - The two major afflictions of the oral cavity are dental caries and gingival/periodontal disease. While microorganisms have long been acknowledged as important etiologic factors, the most recent research data demonstrate that both of these pathologic conditions have a strong hereditary base, i.e., even in the presence of putative pathogenic microorganisms, if the host individual is not genetically susceptible, ensuing disease will be mild or even nonexistent. In the face of this evidence for heritability of the two major oral diseases, we evaluated what educational experiences in genetics were provided to students in U.S. dental schools and dental hygiene programs in 2003-04. Our survey of fifty four dental schools revealed that only one requires a formal genetics course before admission, and only six incorporate a required genetics course within the dental curriculum. Of the 264 dental hygiene programs surveyed, none require a formal genetics course as a prerequisite for admission, and none require a formal genetics course within their curricula. The enormous successes, and future promise, of the Human Genome Project suggest that genetics will soon dominate the future of medicine and dentistry, in prediction of diseases, disease diagnosis, and, eventually, therapy for genetically based disorders. It is therefore incumbent upon dental and dental hygiene education programs to provide genetics education for tomorrow's practitioners. PMID- 15286104 TI - A snapshot of the U.S. postdoctoral pediatric dentistry faculty workforce, 2002. AB - This study characterizes the faculty shortage in U.S. postdoctoral pediatric dentistry (PD) education. The objectives of the study were to determine: 1) the changes in PD faculty numbers and teaching loads between 1995 and 2002 for postdoctoral PD education, 2) current faculty age and training, and 3) distribution of faculty by age. A questionnaire was sent in 2002 to fifty-four programs, of which forty-six responded (85 percent). Dental school and residency mean class sizes increased in the seven-year study period from 82.8 to 91.8 and from 6.0 to 8.5, respectively. Full- and part-time mean faculty positions increased as did vacancies, the latter growing from 15 to 38.9 and changing during the period from 5 to 10.8 percent of available positions. About one-third of programs used general dentists to teach PD, while programs using foreign trained educators grew from 4 to 13 percent. Twenty-nine percent of full-time and 27 percent of part-time faculty are fifty-five years or older, and young entry level faculty, age twenty-five to twenty-nine, represent only 2 percent and 5 percent of full- and part-time faculty respectively. Faculty vacancies have increased along with numbers of students and residents, and the largest segment of PD faculty is within a decade of retirement age. PMID- 15286105 TI - Improving performance on the endodontic section of the Florida Dental Licensure Examination. AB - In an attempt to improve performance of University of Florida College of Dentistry (UFCD) graduates on the endodontic section of the Florida Dental Licensure Examination, a retrospective analysis was conducted for classes graduating between 1996 and 2003 to assess potential relationships between passing and failing performance and three factors with potential impact on "first attempt" pass rates. The three factors were clinical endodontic experience, performance on the senior mock board examination, and dialogue with representatives of the licensure examination, which resulted in modification of the endodontic section of the licensure exam. Using ANOVA, we found no differences in performance on the endodontic section of the senior mock board exam between graduates who passed the endodontic section of the dental licensure exam and those who failed this section. Furthermore, no differences were found in the mean number of clinical endodontic experiences (number of teeth treated) between graduates who passed the endodontic section of the licensure exam and those who failed. However, in 2003 following dialogue between representatives of the Florida Board of Dentistry and endodontic faculty from the two dental schools in Florida, a significant difference in senior mock board endodontic scores (p>0.05) and a significant difference in performance on the endodontic section of the licensure exam scores (p>0.005) was observed for the 2003 graduates when compared to the 2002 graduates. The mean mock board scores and the mean state board endodontic section scores were higher for the 2003 graduates. In addition, the UFCD failure rate on the endodontic section of the state board exam dropped from 34 percent in 2002 to 6 percent in 2003. The primary factors believed responsible for these improvements were a direct result of dialogue between dental school faculty and state board representatives. They include a greater appreciation by the UFCD faculty for the performance criteria used by the Board of Dentistry to evaluate procedures and a change by the board in the tooth selection criteria for the endodontic experience. The options in tooth-type used in the board exams increased from a two-rooted maxillary premolar to any anterior or premolar tooth. In conclusion, this report supports the positive benefits from ongoing discussions between dental school faculty and representatives of the state licensure board. PMID- 15286106 TI - Enamel matrix derivative for periodontal tissue regeneration in treatment of intrabony defects: a Cochrane systematic review. AB - We reviewed the literature on the efficacy of enamel matrix derivative (EMD) in comparison with open flap debridement, guided tissue regeneration (GTR), and bone grafting for the treatment of intrabony defects. We searched four major electronic databases for randomized controlled trials (RCTs) with at least one year of follow-up. Several journals were handsearched with no language restrictions. Outcome measures were: tooth loss, changes in probing attachment levels (PAL), pocket depths (PPD), gingival recessions (REC), marginal bone levels on intraoral radiographs, and postoperative infections. Screening of eligible RCTs, assessment of the methodological quality, and data extraction were conducted in duplicate. No difference in tooth loss was observed. A meta-analysis (eight trials) showed that EMD-treated sites displayed statistically significant PAL improvements (mean difference 1.3 mm) and PPD reduction (1 mm) when compared to flap surgery. When EMD was compared to GTR (six trials), GTR showed a statistically significant reduction of PPD (0.6 mm) and increase of REC (0.5 mm). No difference in postoperative infections was observed. No trials compared EMD with bone grafts alone. EMD is able to significantly improve PAL levels and PPD reduction when compared to flap surgery; however, there is no evidence that more teeth could be saved. There was no evidence of important differences between EMD and GTR. PMID- 15286107 TI - Academy: strengthening the educational mission in academic health centers. AB - A dental school in the modern academic health center often finds that its traditional mission of educating dentists has become more complex as a result of the increased emphasis placed on research and patient care. Although health care education clearly benefits from a rich environment including research and patient care, faculty often find themselves conflicted about their roles in the complex mix of activities in the health center. In order to support faculty in their educational activities, several schools have formed organizational structures with the specific purpose of providing financial and developmental assistance. This article describes a model for an organizational structure within the Harvard School of Dental Medicine designed to promote educational excellence and then describes two academies dedicated to promoting teaching quality at Harvard Medical School and at the University of California at San Francisco. PMID- 15286108 TI - Predoctoral dental school curriculum for catastrophe preparedness. AB - Preparing for catastrophic events, both human-made and natural, is in the national interest and has become a priority since catastrophic events in Oklahoma City, Washington, DC, and New York City. Dentists are a large source of non physician health manpower that could contribute to the public welfare during catastrophic events that require additional public health human resources. Dentists, by virtue of their education, understand biomedical concepts and have patient care skills that can be directly applied during a catastrophic event. Dentists also can provide training for other types of health care workers and can supervise these individuals. In this article, we propose that dentistry can make a significant contribution as part of a national response before, during, and after a catastrophic event or at the time of a public health emergency. We describe the potential collaboration among a dental school, city and state health departments, law enforcement, the military, and others to develop a curriculum in catastrophe preparedness. Then we describe one dental school's effort to build a catastrophe preparedness curriculum for our students. The competencies, goals and objectives, and sources of content for this catastrophe preparedness curriculum are described as well as suggestions for sequencing instruction. PMID- 15286109 TI - Student learning preferences and teaching implications. AB - One of the most serious challenges that dental educators face today is improving the level of student satisfaction with the curriculum and learning environment. To determine whether a particular teaching method might enhance student satisfaction with the learning process, a learning preference survey linked to sensory modalities was given to students in the four classes of the Temple University School of Dentistry. New Zealand educator Neil Fleming developed the survey called VARK (an acronym for Visual, Aural, Read/Write, and Kinesthetic) in 1998. The purpose of this study was to measure the distribution of learning preference mean scores of the dental students and note any significant differences among classes, gender, and a sample population determined using 31,243 participants on the VARK website. Results clearly demonstrate that the dominant preference distributions for the two populations (dental student and sample population) are different. In particular, the proportions of learners who selected visual or kinesthetic are significantly different for the two populations, while the proportions of learners who selected aural or read/write are not significantly different. Dental students prefer visual learning at a higher percentage and kinesthetic learning at a lower percentage than the sample population measured in the VARK website. Inter-class differences varied, and gender differences were not significant. The distribution of dental student scores shows a preference for instructors who use strong visual presentations and facilitate note-taking during lectures. Dental educators should be aware of these differences in order to explore opportunities for making the educational experience more productive and enjoyable. PMID- 15286110 TI - Equivalence study of a dental anatomy computer-assisted learning program. AB - Tooth Morphology is a computer-assisted learning program designed to teach the anatomy of the adult dentition. The purpose of this study was to test whether Tooth Morphology could teach dental anatomy to first-year dental students as well as the traditional lecture. A randomized controlled trial was performed with forty-five first-year dental students. The students were randomly assigned to either the Tooth Morphology group (n=23), which used the computer-assisted learning program and did not attend lecture, or the lecture group (n=22), which attended the traditional lecture and did not use Tooth Morphology. The Tooth Morphology group had a final exam average of 90.0 (standard deviation=5.2), and the lecture group had a final exam average of 90.9 (sd=5.3). Analysis showed that the two groups' scores were statistically equivalent (p<0.05), with a priori equivalence bounds around the difference between the groups set at +/-5 points. It was concluded that Tooth Morphology taught the anatomy of the adult dentition as well as traditional lecture, as measured by exams. Based on the results of this study and student feedback, Tooth Morphology, in combination with interactive class meetings, has replaced the traditional dental anatomy lectures. PMID- 15286111 TI - A comparison of retreatment decisions among general dental practitioners and endodontists. AB - This study compared the difference in decision making regarding retreatment of endodontically treated teeth by general dental practitioners and endodontists. Thirty radiographs of endodontically treated teeth taken from undergraduate records with their respective case descriptions were submitted to fifteen endodontists and fifteen general dental practitioners. Seven treatment alternatives were given as choices; reasons for retreatment, if chosen, were also requested and presented as choices. The results showed statistically different decisions among these two groups regarding retreatment cases. More endodontists opted for retreatment of cases, while higher percentages of general dentists decided to observe, not treat or extract. To prevent misdiagnosis and eventually mistreatment, endodontic decision making should be taught. Currently, there are no specific guidelines for management of failed root canal retreatment. It is suggested that guidelines generated by evidence-based dentistry may produce less variation in clinical decision making. PMID- 15286112 TI - Applicants to U.S. dental schools: an analysis of the 2002 entering class. AB - In 2002, there were 7,537 applicants to all entering dental school classes in the United States. This represents a 1.7 percent increase over the number of applicants in 2001. Between the peak of applicants in 1997 (at 9,829) and 2001, the number declined 25.0 percent. (This is similar to the percent decline that occurred in medical school applicants since their peak in 1996, at 46,968.) Dental schools reported 4,372 first-time, first-year enrollees in 2002. This is an increase of 105 first-time, first-year enrollees over the number reported in 2001. With the 1.7 percent increase in applicants and the 2.5 percent increase in first-time, first-year enrollees over last year, 58 percent of the dental school applicants were enrolled in 2002. This is up very slightly from 57.6 percent in 2001. Since 1989 when dental school enrollment once again began to increase, the number of first-time, first-year enrollees has increased 17.7 percent. (Total first-year enrollment, which includes first-time enrollees and repeat students, has increased 11.8 percent since 1989.) The number of applicants per first-time, first-year position was 1.72 in 2002. It was 2.31 in 1997. (The most recent low was 1.34 in 1989.) The average GPA and DAT scores of first-time, first-year enrollees in 2002 were essentially unchanged from what they were in 2001. Women were 43.7 percent of the applicants and 42.7 percent of first-time, first-year enrollees in 2002, slight increases from what they were in 2001. Underrepresented minorities comprised 12.8 percent of the applicants and 11.4 percent of the first time, first-year enrollees in 2002. These percentages are little changed from those reported in 2001. PMID- 15286114 TI - The classic caries clinical trial: constraints and opportunities. AB - The history of clinical trials would include events in 1747 on board the Salisbury, a British Navy vessel at sea with 12 seamen critically ill with scurvy. Involving these 12 sailors in a study, an officer on board by the name of Lind evaluated six potential treatments for scurvy, and rapidly reached the conclusion that daily consumption of citrus fruits returned the men fit for duty in approximately six days (Bull, 1959). The concept of experimental randomization was first developed by Sir R.A. Fisher (1925, 1926), and the method was introduced to medical research via a study of tuberculosis treatment by Amberson and co-workers (1931), who randomized 24 TB patients into two groups, one to receive the experimental therapy, the other serving as the control. Amberson et al. also incorporated the concept of blinding into their study. Sir Austin Bradford Hill codified and built on the principles of scientific experimentation developed by Fisher, and introduced the use of random numbers in the allocation of patients in the British Medical Research Council (1948) study of the effect of streptomycin in the treatment of tuberculosis (Daniels and Hill, 1952; Hill, 1952). The first applications of clinical trial methodology for testing interventions on dental, oral, and maxillofacial diseases and conditions are more difficult to determine. For dental caries prevention, however, Chilton and Fertig (1958) and Slack and Martin (1964) were certainly among the early caries clinical trial pioneers. As clinical trials have come into the mainstream of clinical research in medicine and dentistry, a great deal of developmental work has focused on their methodological enhancement. The most successful of these efforts have come from fruitful, ongoing collaborations among clinician investigators, biostatisticians, data management specialists, biomedical ethicists, and others with an academic interest in clinical trial design and utilization. During the past 25 years, the emergence of systematic reviews and the evidence-based medicine (EBM) movement have also contributed significantly to the increasing reliance on randomized clinical trial outcomes for the advancement of better clinical practice (Richards et al., 1997; Straus and Sackett, 1998; www.cochrane.org/cochrane/ccbroch.htm#BDL, 2002). PMID- 15286115 TI - A report on the NIH Consensus Development Conference on Diagnosis and Management of Dental Caries Throughout Life. PMID- 15286116 TI - New developments in medical clinical trials. AB - This paper reviews several new developments and long-standing good practices for conducting clinical trials. Discussion starts with the need for clear statements of study objectives, proceeds to clarify target and sample population, and elaborates on primary vs. secondary variables with the need for alpha adjustment in the presence of multiple outcomes. Here we also review the issue of surrogate endpoints. Study design issues--including blinding, randomization, and multicenter studies--come next. Then we discuss the current trend of the replacement of placebo-controlled trials by active controlled non-inferiority trials, the increasing use of Independent Data Monitoring Committees, the prominence of analysis on Intention-to-Treat samples, and the importance of imputation of missing data. We close with a brief discussion of the unit of analysis, the role of newer statistical analysis methods, safety issues, subset analysis, and, most importantly, clinical significance. PMID- 15286117 TI - The future of web-based clinical research in dentistry. AB - Clinical research is a growing industry in dentistry, requiring more efficient methods for research conduct. The Internet provides an opportunity for investigators to expand, enhance, and potentially redefine the way they conduct clinical research. A prime area for improved research is in clinical trials processes, due to their current high cost and inefficiencies. The Web offers the advantages of both centralization of information and coordination of multiple clinical trials processes. The result is improvements for the investigator sites, the sponsor, and subject safety. Observational studies also provide the opportunity for investigators to collect greater amounts of information in a more realistic dental environment. The Internet offers investigators the opportunity to collect outcomes information from sources worldwide, and more efficiently, accurately, and quickly, providing new valuable research answers. The tremendous explosion of genetic and gene therapy research will substantially increase the need for research methods that can manage the vast amount of complex new data, depending on the Internet to manage these research needs. The Internet holds promise for improving dental clinical research through its gains in efficiency, accuracy, and safety. PMID- 15286118 TI - Overview of the impact of changing global patterns of dental caries experience on caries clinical trials. PMID- 15286119 TI - What constitutes dental caries? Histopathology of carious enamel and dentin related to the action of cariogenic biofilms. AB - Substantial pH fluctuations within the biofilm on the tooth surface are a ubiquitous and natural phenomenon, taking place at any time during the day and night. The result may be recordable in the dental tissues at only a chemical and/or ultrastructural level (subclinical level). Alternatively, a net loss of mineral leading to dissolution of dental hard tissues may result in a caries lesion that can be seen clinically. Thus, the appearance of the lesion may vary from an initial loss of mineral, seen only in the very surface layers at the ultrastructural level, to total tooth destruction. Regular removal of the biofilm, preferably with a toothpaste containing fluoride, delays or even arrests lesion progression. This can occur at any stage of lesion progression, because it is the biofilm at the tooth or cavity surface that drives the caries process. Active enamel lesions involve surface erosion and subsurface porosity. Inactive or arrested lesions have an abraded surface, but subsurface mineral loss remains, and a true subsurface remineralization is rarely achievable, because the surface zone acts as a diffusion barrier. The dentin reacts to the stimulus in the biofilm by tubular sclerosis and reactionary dentin. PMID- 15286120 TI - The continuum of dental caries--evidence for a dynamic disease process. AB - The eventual outcome of dental caries is determined by the dynamic balance between pathological factors that lead to demineralization and protective factors that lead to remineralization. Pathological factors include acidogenic bacteria, inhibition of salivary function, and frequency of ingestion of fermentable carbohydrates. Protective factors include salivary flow, numerous salivary components, antibacterials (both natural and applied), fluoride from extrinsic sources, and selected dietary components. Intervention in the caries process can occur at any stage, either naturally or by the insertion of some procedure or treatment. Dental caries covers the continuum from the first atomic level of demineralization, through the initial enamel or root lesion, through dentinal involvement, to eventual cavitation. The dynamic balance between demineralization and remineralization determines the end result. The disease is reversible, if detected early enough. Since demineralization can be quantified at early stages, before frank cavitation, intervention methods can be tested by short-term clinical trials. PMID- 15286121 TI - Modern concepts of caries measurement. AB - Following the consideration of several recent systematic and other reviews, there is a growing professional and scientific consensus that caries measurement methodology in caries clinical trials (CCT) should be updated to reflect progress made elsewhere in cariology. In this paper, therefore, "modern" means accepted in contemporary dental research and dental practice on the basis of sound research evidence--not necessarily new or requiring the use of new technology. Caries measurement should be seen in the context of the objectives of modern clinical caries management and the continuum of disease states, ranging from sub-surface carious changes through to more advanced lesions. Measurement concepts can be applied to at least three levels: the tooth surface, the individual, or the group/population. All are relevant to CCTs. Modern clinical caries management can be seen as comprised of seven discrete but linked steps (Steps 2, 3, and 4 are directly concerned with measurement.): (1) 'Caries detection' represents a yes/no decision as to whether caries is present; (2) lesion measurement assesses defined stages of the caries process, taking into account the histopatholgical morphology and appearance of different sizes and types of lesion and the diagnostic threshold(s) being used; (3) lesion monitoring by repeated measures at a series of examinations is used when lesions are less advanced than the stage judged to require operative intervention (A comparison of serial measurements permits the efficacy of preventive care aiming either to arrest or to reverse the lesion to be assessed.); (4) caries activity measures would be very valuable, but are relatively poorly developed and tested at present; (5) diagnosis, prognosis, and clinical decision-making are the important human processes in which all the information obtained from steps 1 to 4 is synthesised; (6) interventions/treatments, both preventive and operative, are now routinely used for caries management; and (7) outcome of caries control/management assesses caries management by examining evidence on the long-term outcomes. A challenge for the future is to define a range of optimal caries measurement methods--in use or in development in recent trials, in clinical practice, and/or in caries epidemiology--that will best contribute to more efficient, modern caries clinical trials. PMID- 15286122 TI - The challenges of validating diagnostic methods and selecting appropriate gold standards. AB - Caries diagnostic methods are usually methods for caries lesion detection and measurement. Caries lesions occur on a continuous scale of tissue damage, from subclinical surface changes to macroscopic cavities reaching the pulp. Any change of a lesion on this continuous scale offers the opportunity for the diagnosis of disease activity or remission. Research aimed at remineralizing agents may focus on lesions that are amenable to remineralization, and select a method that will measure small changes in early lesions. General caries management strategies depend on detecting all stages of lesion development, and methods covering early to late stages are preferred. This paper addresses some methodological issues in validating caries diagnostic methods. The available gold standards for caries lesions are discussed, with their suitability in different applications, and their "validity" as far as it is known or can be inferred. The gold standards are compared as far as their measurement of lesion parameters and reproducibility is concerned. Tentative conclusions are formulated, and recommendations for future research are given. PMID- 15286123 TI - The challenges of validating diagnostic methods relative to a conventional two year caries clinical trial. AB - This paper is directed to the question, "What are the appropriate validation criteria for the use of a new clinical trial methodology as a replacement for a conventional two- to three-year caries study?" It is important to recognize that the objective of a two- to three-year randomized, controlled caries trial is to test a precisely framed hypothesis, regarding an experimental product's efficacy relative to a control product. The external validity of conventional two- to three-year caries clinical studies in determining the efficacy and safety of anti caries products is well-accepted. However, caries clinical trials are not without limitations and have increasingly been viewed as inefficient with respect to measuring the disease process in a holistic manner. The endpoint of a caries lesion with loss of enamel integrity (cavitation) focuses on one end of the caries progression continuum at the expense of early caries initiation and progression. Several early caries detection methods have been developed that correlate with mineral loss of the tooth surface. These diagnostics differ from conventional visual-tactile and radiographic methods in that they are capable of detecting early non-cavitated lesions, and this can generate continuous data. As diagnostic methods become accepted, they will lead to study designs that diverge from the conventional two- to three-year caries studies. Modification of the existing two- to three-year conventional caries design for assessment of product effectiveness, whether by the introduction of a new diagnostic method or by modification of the overall clinical design, must result in a clinical design that is able to differentiate known treatments on the basis of caries prevention efficacy. Given that the fluoride dose response has been characterized in the literature, this should form the basis of any validation package for new methodologies. In conclusion, a minimum expectation for acceptance as a replacement to conventional testing should be that the method or design can differentiate products of known efficacy from one another and that the efficacy relationship observed in a two- to three-year conventional study can be observed with the new method or design. PMID- 15286124 TI - Visual and visuo-tactile detection of dental caries. AB - The objective of this review is to describe and discuss the content validity of a sample of caries detection criteria reported in the literature between January 1, 1966, and May 1, 2000. Using filters to locate randomized or controlled clinical trials on dental caries, fluorides, sealants, and "restorative" care, I identified a total of 171 documents from MEDLINE and the Cochrane Collaboration's Oral Health Group (CC-OHG) special register. These articles met the following inclusion criteria: (1) Data had been collected from samples of patients or populations; and (2) dental caries was assessed clinically, and criteria were either published or described in the paper. From the selected articles, evidence tables were prepared describing each caries detection criterion. Analysis of the content validity of the criteria systems was based on evaluation of the disease process, exclusion of non-caries lesions, subjectivity, use of explorers, and drying of teeth prior to examination. This review included 29 unique criteria systems. Of those, 13 originated from the UK, 3 from the USA, 4 from Denmark, and others from the World Health Organization (WHO), Sweden, Switzerland, Norway, Netherlands, and Canada. Thirteen of the criteria systems either measured active and inactive early and cavitated lesions or defined separate criteria for smooth and occlusal tooth surfaces. Nine systems measured early as well as cavitated stages of the caries process, and 7 measured cavitation only. Eleven of the criteria systems provided explicit descriptions of the disease process measured or information on how to exclude non-caries from caries lesions. The use of explorers and drying and cleaning of teeth varied widely among the criteria. The majority of the newly developed criteria systems originated from Europe. In conclusion, this review of the content validity of the 29 criteria systems found substantial variability in disease processes measured, inclusion and exclusion criteria, and examination conditions. PMID- 15286125 TI - Improving clinical visual detection--potential for caries clinical trials. AB - It has been the norm in caries clinical trials to measure caries increment by several different caries-recording systems, including the crude DMF-S/T index. However, there is a reserved attitude as to whether to subdivide the non cavitated lesions and use arrested lesions in clinical trials. This has been due to the belief that it is not possible to achieve reliable data of the early stages of the disease (Radike, 1972). However, recently, Ekstrand et al.(1997, 1998) showed that it was possible: (1) to differentiate between different stages of non-cavitated occlusal lesions, (2) to differentiate between active and inactive occlusal lesions, and (3) to predict the depth of the lesion. In at least 4 other clinical studies, the reproducibility of recording initial active lesions, cavitated active lesions, and arrested lesions was found to be adequate (Carvalho et al., 1989; Nyvad et al, 1999; Ekstrand et al., 2000; Machiulskiene et al., 2001). Since caries today is a more slowly developing disease in many countries in the world, this will result in prolongation of the duration of the clinical trial, which will increase the costs. As indicated above, there is now sufficient evidence that caries can be clinically diagnosed accurately and reliably in earlier stages as well as in an arrested stage. If such stages of caries are used as outcome variables in caries clinical trials, they may have a positive influence on the trials' duration and costs. PMID- 15286126 TI - Bitewing and digital bitewing radiography for detection of caries lesions. AB - When radiography is applied in the clinic for caries detection, the recommended technique is bitewing projection (Grondahl, 1994). This technique was introduced by Raper (1925) and has continued with only minor alterations. The aims of this report are to provide evidence for (1) optimal bitewing recording for individual examinations and clinical trials, (2) advantages and disadvantages of digital receptors for bitewing examinations, (3) the diagnostic outcomes and limitations of bitewing radiography, and (4) computer-automated detection of caries. PMID- 15286127 TI - Electrical measurements for use in caries clinical trials. AB - This paper reviews the use of electrical measurements of caries, particularly in relation to caries clinical trials. Electrical measurements change as tooth tissue porosity alters in the caries process, but several other variables also have a significant effect on these electrical measurements and hence upon their diagnostic validity. Available electrical-method data, in the context of clinical trials, relate to the use of the Electronic Caries Monitor (ECM), which measures "bulk" resistance. The device is presently limited in scope to occlusal surfaces, and only limited ECM data from clinical trials are available. In the context of clinical trials, more work is needed to determine the potential role of electrical measurements. Such research will need to focus both on an understanding of those electrical parameters which are most valuable in identifying changes and stages in the caries process in individual teeth and also on identifying the extent of the effects of the variables affecting these measurements. PMID- 15286128 TI - DIAGNOdent: an optical method for caries detection. AB - The onset of caries is characterized by demineralization of dental hard tissues. Optimal fluoridation with respective oral hygiene habits and diet may stop the progression of a lesion and even allow for its remineralization. The aim of modern dentistry must be a preventive approach rather than invasive repair of the disease. This is possible only with early detection and respective preventive measures. Some of today's diagnostic tools are not sensitive enough to detect this early onset of destruction. Tools based on fluorescence could have the possibility to overcome this problem. This overview will focus on today's knowledge of one possible tool, the DIAGNOdent. PMID- 15286129 TI - Optical methods--quantitative light fluorescence. AB - Considerable research during the past two decades has focused upon the development of new technologies for the detection of dental caries. Of these technologies, the method that has been most extensively studied is based upon the indirectly assessed changes in the fluorescence of enamel associated with the loss of mineral. The purpose of this presentation was to review the available information regarding the use of this technology, commonly known as quantitative light fluorescence, for caries detection, particularly early caries detection, and the potential for the routine use of this technology in clinical caries trials. This technology is unique in its ability to measure small changes in the mineral content of enamel lesions quantitatively. The results of recent small scale clinical trials have indicated that the impact of caries-preventive measures can be determined within a six-month period. With current hardware and software refinements and the results of long-term clinical validation studies that are in progress, it may be that this technology will be the future method of choice for caries clinical trials. PMID- 15286130 TI - A review of potential new diagnostic modalities for caries lesions. AB - This paper aims to present a simple overview of potential new diagnostic methods for dental caries. There are several novel methods of caries detection (with potential application to diagnosis) which have been proposed in the last few years, in addition to those that are gaining some commercial exposure and clinical acceptance. For the most part, these methods have been demonstrated in laboratories and are generally many years away from routine clinical application. They include multi-photon imaging, infrared thermography and infrared fluorescence, optical coherence tomography, ultrasound, and terahertz imaging. PMID- 15286131 TI - Efficiency issues among statistical methods for demonstrating efficacy of caries prevention. AB - Although repeated tooth-surface-specific information is commonly collected during a longitudinal caries clinical trial, traditional methods often make limited use of the repeated measures. Newer methods of analysis, such as methods based on time-to-event and methods for longitudinal or clustered data, have the potential to increase the efficiency of the statistical analysis. We compare a range of analytical methods from the traditional analysis based only on the number of caries onsets to newer methods that incorporate time at risk and surface-specific information, such as Poisson regression methods for clustered data, with respect to the efficiency of treatment comparisons. Under most circumstances, the greatest gain in efficiency associated with time-to-event methods will be due to the ability of subjects to contribute caries onsets to the analysis until they are lost from the study. Incorporating the number of surfaces at risk, the surface time at risk, and surface-specific characteristics will typically produce only a modest gain in efficiency. PMID- 15286132 TI - Using survival methodologies in demonstrating caries efficacy. AB - Exploiting recent advances in statistical methods, particularly for correlated intra-subject data, could increase the efficiency of caries clinical trials. Methods of analysis using the tooth surface as the unit should be investigated. Whole-mouth measures such as the DMFS increment ignore the variation in the number of surfaces at risk between subjects and within a subject over time. The use of "survival time" for each surface as the outcome measure--i.e., the time from the start of the trial to when a surface is recorded as decayed or filled- is proposed. Data from caries clinical trials could be described as clustered survival data, where clustering of tooth surfaces exists such that survival times within the same cluster or subject are correlated. Advances in the analysis of clustered survival data, such as the use of marginal models with robust variance estimators, have recently been exploited in the analysis of caries clinical trials. The analysis produced results similar to those achieved by conventional DMFS-based analysis. The results using survival analysis are easily interpreted for example, the median survival time of tooth surfaces in female subjects using a toothpaste with a higher level of fluoride (1500 ppm F) is 1.07 times the median survival time of surfaces in female subjects using toothpaste with less fluoride (1000 ppm F). Further research is required to investigate if survival analysis is a more sensitive method of analysis, i.e., whether causative factors can be identified with fewer subjects than with the conventional method of analysis. PMID- 15286133 TI - Analysis of clinical trials involving non-cavitated caries lesions. AB - Treatments to halt or reverse the progression of non-cavitated caries lesions are of increasing interest. Diagnostic technologies under development offer potential for the assessment of gradual progression and regression of such lesions. Many therapies directed at correcting demineralization-remineralization imbalance should, in principle, protect enamel similarly across lesion severities from initiation to near cavitation. If this is so, and if acceptable reproducibility and predictive validity can be demonstrated for a diagnostic of acceptable cost, then clinical trials of agents to prevent cavitation can become more efficient by the use of outcome indices that reflect, in addition to cavitation, the expansion and regression of non-cavitated lesions. However, to achieve such a benefit will require data analyses that fully exploit ordinal or continuous-scale outcome measures. We consider comparison of such measures of lesion status between treatment groups, with most attention to ordinal categorical data. Interim data from a clinical trial in Lithuanian children are used for illustration. PMID- 15286134 TI - Statistical issues for combining multiple caries diagnostics for demonstrating caries efficacy. AB - Caries efficacy in clinical trials has been based primarily on visual examinations supplemented by Fiber Optic Transillumination (FOTI) and radiography, with the assessments combined at the surface level to classify each surface as to its caries status. Newer caries diagnostics techniques measure the caries process in a quantitative manner and so thus yield continuous rather than ordinal results. The objective of this study was to examine various methods for the analysis of multiple outcomes in clinical trials and to compare their usefulness for the analysis of caries trials. Four global tests (rank sum, ordinary least squares, general least squares, and generalized estimating equations) and two caries indices (based on average and maximum values of the methods) were evaluated with the use of one-year follow-up data from 1063 children in a recent caries trial. A new hybrid method was also developed and evaluated. All of the methods performed well when the diagnostic measures showed product differences in caries in the same direction. Ease of use, interpretability, and distributional assumptions must be considered before a consensus method for analysis of multiple diagnostic measures in caries trials can be determined. PMID- 15286135 TI - Current issues in clinical equivalence trials. AB - A clinical trial designed to show that an experimental treatment E is similar to a control treatment S in a specified direction is a one-sided equivalence or similarity trial--in the terminology of the International Conference on Harmonisation, a non-inferiority trial (ICH, 1998). We design such a study to show that E is not worse than S (often an accepted or standard treatment) by as much as a pre-specified margin theta0. The quantity theta0 can be either a difference or ratio of an appropriate outcome in individuals treated with E and S. A critical issue is whether one can conclude from a non-inferiority trial that E is effective. Closely related is an appropriate choice of theta0, which should be substantially less than the estimated effect of S if available from previous studies; theta0 should also be acceptable to clinicians, either because of advantages of E or because a difference or ratio less than theta0 is considered unimportant clinically. Another possible approach for showing that E is effective is to estimate its effect compared with placebo from historical data. If previous studies that consistently show an effect of S are not available, alternative study designs should be considered. Findings of superiority or non-inferiority of E, when the study was planned to show the other, are possible and may be supportable. A finding that E is at the same time statistically significantly worse than S and "non-inferior" to S should not be a problem, if the criterion theta0 is appropriate and this possibility was considered in the protocol. Various sorts of non-adherence may make treatments appear similar, even if they are not. In particular, random non-adherence of study participants to the assigned treatment regimen may cause an intention-to-treat analysis to give a misleading result of similarity. Thus, maintaining a high degree of adherence to protocol is especially important in an equivalence or non-inferiority trial. Interim analysis does not present statistical problems in these trials; early stopping may not be wise in many cases, however, because strong interim evidence for non-inferiority may actually be an indicator that E is superior to S. PMID- 15286136 TI - The role of risk factors in the identification of appropriate subjects for caries clinical trials: design considerations. AB - In seeking new and more effective therapies to delay or prevent caries development, investigators must design clinical trials focused on high-risk populations with a predictable incidence of caries over a limited period of time. In children and adolescents, the strongest predictors of caries incidence appear to be baseline levels of caries activity (present caries, e.g., dmfs, DMFT, caries lesions in first molars). Other predictors of caries risk typically include oral hygiene level, counts of cariogenic micro-organisms in plaque and saliva, fluoride history, sucrose intake, and parent's socio-economic level. This paper will briefly review existing literature to address the most useful and relevant prognostic factors for predicting future caries onset. The relative merits of identifying high-risk subjects based on these factors, either singly or in combination, will be explored in terms of statistical efficiency. Particular attention will focus on the advantages of covariate adjustment in the context of survival-based methods for the analysis of caries data. Further, with the advent of more sophisticated diagnostic procedures (e.g., quantitative light fluorescence) to screen and monitor study subjects for caries activity, there is the potential for earlier states of lesion initiation and progression (or regression) to be detected, with, therefore, improved experimental sensitivity to treatment effects. The validity of risk assessment and outcome measurement on the basis of these new diagnostic tools vs. more conventional methods will be examined. PMID- 15286137 TI - Methods for pooling results from multi-center studies. AB - A multi-center study is one conducted simultaneously in several participating centers following an agreed protocol, where the randomization has been carried out independently within each center. The main consideration for pooling the data from the individual centers is the choice between a weighted analysis, which weights centers relative to the number of patients in them, or an unweighted analysis as the primary statistical method. The unweighted analysis is used to investigate whether there was an interaction between the centers and study groups. Another issue is whether a fixed- or random-effects model should be used. There is unresolved controversy among statisticians about whether to use a weighted (type II) or unweighted analysis (type III), since there are advantages and disadvantages to the use of either method. The weighted analysis provides the most powerful test of the treatment contrast if there is no interaction between treatment and center. If there is an interaction, the unweighted analysis leads to unbiased estimates. Although, from an estimation and hypothesis testing standpoint, there is no need to balance the number of patients between the sites, it is sensible to avoid major imbalances among the study sites. There is agreement among statisticians that a fixed-effects model should be used. PMID- 15286138 TI - Potential modern alternative designs for caries clinical trials (CCTs) and how these can be validated against the conventional model. AB - The main reasons that industry runs caries clinical trials (CCTs) are to provide proof of efficacy and to collect in vivo safety data on new products. In recent years, predominantly due to declining caries levels and the use of positive controls, the cost of performing these CCTs has escalated. It is now reaching the stage where it is becoming commercially prohibitive to conduct new studies. This is likely to stifle innovation of new anticaries products, and we now need new, more discriminatory, faster, and less expensive study designs. There are many ways in which the design of CCTs may be changed, such as improving diagnostic efficiency, improving data handling/statistical modeling, and using high-risk populations. However, it is paramount that the overriding principle behind CCT design validation must be that the results/conclusions from any new design are in line with those shown previously by 'conventional' CCTs, to ensure the maintenance of standards for both efficacy and safety. It is suggested that the validation of any new trial design must involve comparisons with regimens previously shown in conventional CCTs to have different anticaries efficacies. For example, since several clinical trials have shown convincing evidence for a monotonic dose response for fluoride at least up to levels of 2500 ppm F, one could choose two products, differing solely in their fluoride level. One aim for this workshop is to identify and agree on validation principles for new clinical trial designs. This will facilitate general international acceptance of novel smaller/faster CCTs designs both now and in the future. We recognize that any new design must not compromise the standard of proof of either efficacy or safety. In addition, any principles will need to take account of current understanding of the caries process, while recognizing the need for change to match future developments in cariology. Finally, the mechanism of action of the test product must be considered, in assessments of the acceptability of novel designs, if this differs markedly from the regimens used to validate the design. PMID- 15286139 TI - International Consensus Workshop on Caries Clinical Trials (ICW-CCT)--final consensus statements: agreeing where the evidence leads. PMID- 15286140 TI - Dealing with the genotype x environment interaction via a modelling approach: a comparison of QTLs of maize leaf length or width with QTLs of model parameters. AB - Quantitative genetics of adaptive traits is made difficult by the genotypexenvironment interaction. A classical assumption is that QTLs identified in both stressed and control conditions correspond to constitutive traits whereas those identified only in stressed treatments are stress-specific and correspond to adaptive traits. This hypothesis was tested by comparing, in the same set of experiments, two ways of analysing the genetic variability of the responses of maize leaf growth to water deficit. One QTL detection was based on raw phenotypic traits (length and width of leaf 6) of 100 recombinant inbred lines (RILs) in four experiments with either well-watered or stressing conditions in the field or in the greenhouse. Another detection followed a method proposed recently which consists of analysing intrinsic responses of the same RILs to environmental conditions, determined jointly over several experiments. QTLs of three responses were considered: (i) leaf elongation rate per unit thermal time in the absence of stress, (ii) its response to evaporative demand in well-watered plants, and (iii) its response to soil water status in the absence of evaporative demand. The QTL of leaf length differed between experiments, but colocalized in seven cases out of 13 with QTLs of the intrinsic leaf elongation rate, even in experiments with stressing conditions. No colocalization was found between QTLs of leaf length under water deficit and QTLs of responses to air or soil water status. By contrast, QTLs of leaf width colocalized in all experiments, regardless of environmental conditions. The classical method of identifying the QTL of constitutive versus adaptive traits therefore did not apply to the experiments presented here. It is suggested that identification of the QTL of parameters of response curves provides a promising alternative for dealing with the genetic variability of adaptive traits. PMID- 15286141 TI - Regulation of calcium signalling and gene expression by glutathione. AB - The glutathione redox couple is an information-rich redox buffer that interacts with numerous cellular components. To explore the role of glutathione in redox signalling, leaf contents were increased either chemically, by feeding reduced glutathione (GSH), or genetically, by over-expressing the first enzyme of the GSH biosynthetic pathway, gamma-glutamylcysteine synthetase (gamma-ECS). Leaf discs were also fed glutathione disulphide (GSSG), leading to increases in both GSH and GSSG. The effects of increases in GSH were compared with non-specific changes in leaf thiol status induced by feeding dithiothreitol (DTT) or the monothiol beta mercaptoethanol (beta-ME). Photosynthesis measurements showed that none of the feeding treatments greatly disrupted leaf physiology. Transgenic plants expressing aequorin were used to analyse calcium signatures during the feeding treatments. Calcium release occurred soon after the onset of GSH or GSSG feeding, but was unaffected by DTT or beta-ME. Pathogenesis-related protein 1 (PR-1) was induced both in the gamma-ECS overexpressors and by feeding GSH, but not GSSG. Feeding DTT also induced PR-1. Key transcripts encoding antioxidative enzymes were much less affected, although glutathione synthetase was suppressed by feeding thiols or GSSG. It is concluded that modulation of glutathione contents transmits information through diverse signalling mechanisms, including (i) the establishment of an appropriate redox potential for thiol/disulphide exchange and (ii) the release of calcium to the cytosol. PMID- 15286142 TI - Control of sulphate assimilation and glutathione synthesis: interaction with N and C metabolism. AB - Sulphate assimilation is an essential pathway being a source of reduced sulphur for various cellular processes and for the synthesis of glutathione, a major factor in plant stress defence. Many reports have shown that sulphate assimilation is well co-ordinated with the assimilation of nitrate and carbon. It has long been known that, during nitrate deficiency, sulphate assimilation is reduced and that the capacity to reduce nitrate is diminished in plants starved for sulphate. Only recently, however, was it shown that adenosine 5' phosphosulphate reductase (APR), the key enzyme of sulphate assimilation, is regulated by carbohydrates. In plants treated with sucrose or glucose APR was induced, whereas the activity was strongly reduced in plants grown in CO(2)-free air. The availability of cysteine is a crucial factor in glutathione synthesis, but an adequate supply of glutamate and glycine are also important. The molecular mechanisms for the co-ordination of S, N, and C assimilation are not known. O acetylserine, a precursor of cysteine, was proposed to be the signal regulating sulphate assimilation, but most probably is not the outgoing signal to N and C metabolism. cDNA arrays revealed the induction of genes involved in auxin synthesis upon S-starvation, pointing to a possible role of phytohormones. Clearly, despite significant progress in understanding the regulation of sulphate assimilation and glutathione synthesis, their co-ordination with N and C metabolism achieved, and several potential signal molecules identified, present knowledge is still far from being sufficient. PMID- 15286143 TI - Irrigation scheduling: advantages and pitfalls of plant-based methods. AB - This paper reviews the various methods available for irrigation scheduling, contrasting traditional water-balance and soil moisture-based approaches with those based on sensing of the plant response to water deficits. The main plant based methods for irrigation scheduling, including those based on direct or indirect measurement of plant water status and those based on plant physiological responses to drought, are outlined and evaluated. Specific plant-based methods include the use of dendrometry, fruit gauges, and other tissue water content sensors, while measurements of growth, sap flow, and stomatal conductance are also outlined. Recent advances, especially in the use of infrared thermometry and thermography for the study of stomatal conductance changes, are highlighted. The relative suitabilities of different approaches for specific crop and climatic situations are discussed, with the aim of indicating the strengths and weaknesses of different approaches, and highlighting their suitability over different spatial and temporal scales. The potential of soil- and plant-based systems for automated irrigation control using various scheduling techniques is also discussed. PMID- 15286144 TI - myo-Inositol and sucrose concentrations affect the accumulation of raffinose family oligosaccharides in seeds. AB - Raffinose family oligosaccharides (RFOs) fulfil multiple functions in plants. In seeds, they possibly protect cellular structures during desiccation and constitute carbon reserves for early germination. Their biosynthesis proceeds by the transfer of galactose units from galactinol to sucrose. Galactinol synthase (GolS), which mediates the synthesis of galactinol from myo-inositol and UDP galactose, has been proposed to be the key enzyme of the pathway. However, no significant relationship was detected between the extractable GolS activity and the amount of RFOs in seeds from seven pea (Pisum sativum L.) genotypes selected for high variation in RFO content. Instead, a highly significant correlation was found between the levels of myo-inositol and RFOs. Moderately strong relationships were also found between sucrose and RFO content as well as between myo-inositol and galactinol. Further evidence for a key role of myo-inositol for the synthesis of galactinol was obtained by feeding exogenous myo-inositol to intact pea seeds and by the analysis of four barley (Hordeum vulgare L.) low phytic acid mutants. In seeds of three of these mutants, the reduced demand for myo-inositol for the synthesis of phytic acid (myo-inositol 1,2,3,4,5,6 hexakisphosphate) was associated with an increased level in myo-inositol. The mutants seeds also contained more galactinol than wild-type seeds. The results suggest that the extent of RFO accumulation is controlled by the levels of the initial substrates, myo-inositol and sucrose, rather than by GolS activity alone. PMID- 15286145 TI - The gene geranylgeranyl reductase of peach (Prunus persica [L.] Batsch) is regulated during leaf development and responds differentially to distinct stress factors. AB - Plant geranylgeranyl hydrogenase (CHL P) reduces free geranylgeranyl diphosphate to phytil diphosphate, which provides the side chain to chlorophylls, tocopherols, and plastoquinones. In peach, the single copy gene (PpCHL P) encodes a deduced product of 51.68 kDa, which harbours a transit peptide for cytoplasm-to chloroplast transport and a nicotinamide binding domain. The PpCHL P message was abundant in chlorophyll-containing tissues and flower organs, but barely detected in the roots and mesocarp of ripening fruits, suggesting that transcription was related to plastid types and maturation. The message was not revealed in shoot apical meristems, but spread thoroughly in leaf cells during the early stages and was located mainly in the palisade of mature leaves, which exhibited higher transcript levels than young ones. Hence, the transcription of PpCHL P was likely to be regulated during leaf development. Gene expression was monitored in leaves responding to natural dark, cold, wounding, stress by imposed darkening, and during the curl disease. Transcription was stimulated by light, but repressed by dark and cold stress. In darkened leaves, the PpCHL P message was augmented concomitantly with that of CATALASE. In wounded leaves, the message decreased, but recovered rapidly, whereas in curled leaves, a reduction in gene expression was related to leaf damage intensity. However, transcript signals increased locally both in cells mechanically wounded by a needle and in those naturally injured by the pathogenic fungus Taphrina deformans. These data suggest that PpCHL P expression was regulated by photosynthetic activity and was possibly involved in the defence response. PMID- 15286146 TI - Accumulation of menaquinones with incompletely reduced side chains and loss of alpha-tocopherol in rice mutants with alternations in the chlorophyll moiety. AB - The rice mutants M249 and M134 accumulate chlorophyllides a and b which are esterified with incompletely reduced alcohols such as geranylgeraniol, dihydrogeranylgeraniol, and tetrahydrogeranylgeraniol. Quantities of alpha tocopherol, phylloquinone, and menaquinones in leaves of these mutants were determined by high performance liquid chromatography (HPLC) with a fluorescence detector after post-column chemical reduction to convert quinones to fluorescent quinols. Methylnaphthoquinones, varying in the reduction state of the side chain (menaquinones), were detected in leaf segments of the rice mutants on HPLC analyses with both high selectivity and sensitivity to plant quinones. Mutant M249 preferentially accumulated menaquinone, which contains tetrahydrogeranylgeraniol as its side chain. However, mutant M134 exhibited preferential accumulation of menaquinone with a geranylgeraniol side chain. In both mutants, the accumulation patterns of menaquinones with different prenyl side chains were similar to those of chlorophyll with the corresponding prenyl side chains. The content of P700, the photosystem I primary electron donor, in the wild type was greater than that of either mutant, on both a chlorophyll and a fresh weight basis. However, the ratios of total methylnaphthoquinones to P700 were similar in both the wild type and the mutants. Since no comparative large differences in photosynthetic activity exist between the wild type and the mutants, these results suggest that the hydrogenation of the methylnaphthoquinone side chain to phytol is not an essential requirement for it to function as an electron acceptor in photosystem I. On the other hand, alpha-tocopherol was detected in fully developed leaves of the wild type, but not in those of the mutants. Accumulation of menaquinones and the loss of alpha-tocopherol in mutant leaves suggest that the reduction of chlorophyll-geranylgeraniol to phytol and that of geranylgeranyl pyrophosphate to phytyl pyrophosphate are catalysed by the same enzyme. PMID- 15286147 TI - Grain yields with limited water. AB - Plant reproduction is sensitive to water deficits, especially during the early phases when development may cease irreversibly even though the parent remains alive. Grain numbers decrease because of several developmental changes, especially ovary abortion in maize (Zea mays L.) or pollen sterility in small grains. In maize, the water deficits inhibit photosynthesis, and the decrease in photosynthate flux to the developing organs appears to trigger abortion. Abscisic acid also increases in the parent and may play a role, perhaps by inhibiting photosynthesis through stomatal closure. Recent work indicates that invertase activity is inhibited and starch is diminished in the ovaries or affected pollen. Also, sucrose fed to the stems rescues many of the ovaries otherwise destined to abort. The feeding restores some of the ovary starch and invertase activity. These studies implicate invertase as a limiting enzyme step for grain yields during a water deficit, and transcript profiling with microarrays has identified genes that are up- or down-regulated during water deficit-induced abortion in maize. However, profiling studies to date have not reported changes in invertase or starch synthesizing genes in water-deficient ovaries, perhaps because there were too few sampling times. The ovary rescue with sucrose feeding indicates either that the changes identified in the profiling are of no consequence for inhibiting ovary development or that gene expression reverts to control levels when the sugar stream recovers. Careful documentation of tissue- and developmentally specific gene expression are needed to resolve these issues and link metabolic changes to the decreased sugar flux affecting the reproductive organs. PMID- 15286148 TI - The promoter of the Arabidopsis nuclear gene COX5b-1, encoding subunit 5b of the mitochondrial cytochrome c oxidase, directs tissue-specific expression by a combination of positive and negative regulatory elements. AB - In the present work, the promoter of the Arabidopsis thaliana nuclear gene COX5b 1, encoding subunit 5b of the mitochondrial cytochrome c oxidase, has been analysed. For this purpose, plants, stably transformed with different promoter fragments fused to the beta-glucuronidase reporter gene, have been obtained. Histochemical staining indicated that the COX5b-1 promoter directs expression in meristems and in vascular tissues of cotyledons, roots, and hypocotyls, as well as in anthers and pollen and the central leaf vein. Quantitative measurements in extracts prepared from different organs suggested that expression is higher in roots. The analysis of progressive upstream deletions of the promoter suggested the presence of negative regulatory elements, preferentially active in leaves, between nucleotides -609 and -387 from the translation start site. A further deletion down to nucleotide -195 completely abolished expression. The inclusion of sucrose or the cytokinin 6-benzylaminopurine in the culture medium induced COX5b-1 promoter-dependent beta-glucuronidase expression. This induction was observed with all constructs that produced beta-glucuronidase activity. Putative regulatory elements involved in the regulation of other genes were detected in the promoter fragment required for expression. A detailed analysis of these elements will help to elucidate the molecular mechanisms that participate in the expression of this and, possibly, other components of the cytochrome c-dependent respiratory pathway. PMID- 15286149 TI - Physico-chemical characteristics of cell walls from Arabidopsis thaliana microcalli showing different adhesion strengths. AB - Changes in the composition and structure of cell walls and extracellular polysaccharides (ECP) were studied during the growth of suspension-cultured Arabidopsis thaliana microcalli. Three growth phases, namely the cell division phase, the cell expansion phase, and the stationary phase, were distinguished and associated with a decreasing cell cluster adhesion strength. Degradation of the homogalacturonan pectic backbone and of linear pectic side chains (1,4)-beta-D galactan were observed concomitantly with the cell expansion and stationary phases and the decrease in cell adhesion. Also, in the stationary phase, branched (1,5)-alpha-L-arabinans were linearized. The AGP content of the culture medium increased while it decreased in the cell wall during cell growth and as cell adhesion decreased. These data suggest that, in addition to homogalacturonan, pectic side chains and AGP are involved in plant cell development and particularly in cell-cell attachment. PMID- 15286150 TI - Cell wall metabolism during maturation, ripening and senescence of peach fruit. AB - Cell wall changes were examined in fruit of a melting flesh peach (Prunus persica L.) allowed to ripen on the tree. Three phases to softening were noted, the first of which began prior to the completion of flesh colour change and an increase in ethylene evolution. Softening in young mature fruit, prior to ripening, was associated with a depolymerization of matrix glycans both loosely and tightly attached to cellulose and a loss of Gal from all cell wall fractions. After the initiation of ripening, but before the melting stage, softening was associated with continuing, progressive depolymerization of matrix glycans. A massive loss of Ara from the loosely bound matrix glycan fraction was observed, probably from side chains of glucuronoarabinoxylan, pectin, or possibly arabinogalactan protein firmly bound into the wall and solubilized in this extract. An increase in the solubilization of polyuronides also occurred during this period, when softening was already well advanced. The extensive softening of the melting period was marked by substantial depolymerization of both loosely and tightly bound matrix glycans, including a loss of Ara from the latter, an increase in matrix glycan extractability, and a dramatic depolymerization of chelator-soluble polyuronides which continued during senescence. Depolymerization of chelator-soluble polyuronides thus occurred substantially after the increase in their solubilization. Ripening-related increases were observed in the activities of exo and endo-polygalacturonase (EC 3.2.1.67; EC 3.2.1.15), pectin methylesterase (EC 3.1.1.11), endo-1,4-beta-glucanase (EC 3.2.1.4), endo-1,4-beta-mannanase (EC 3.2.1.78), alpha-arabinosidase (EC 3.2.1.55), and beta-galactosidase (EC 3.2.1.23), but the timing and extent of the increases differed between enzymes and was not necessarily related to ethylene evolution. Fruit softening in peach is a continuous process and correlated closely with the depolymerization of matrix glycans, which proceeded throughout development. However, numerous other cell wall changes also took place, such as the deglycosylation of particular polymers and the solubilization and depolymerization of chelator-soluble polyuronides, but these were transient and occurred only at specific phases of the softening process. Fruit softening and other textural changes in peach appear to have a number of stages, each involving a different set of cell wall modifications. PMID- 15286151 TI - Maternal apo E genotype is a modifier of the Smith-Lemli-Opitz syndrome. AB - BACKGROUND: Smith-Lemli-Opitz syndrome (MIM 270400) is an autosomal recessive malformation and mental retardation syndrome that ranges in clinical severity from minimal dysmorphism and mild mental retardation to severe congenital anomalies and intrauterine death. Smith-Lemli-Opitz syndrome is caused by mutations in the Delta7 sterol-reductase gene (DHCR7; EC 1.3.1.21), which impair endogenous cholesterol biosynthesis and make the growing embryo dependent on exogenous (maternal) sources of cholesterol. We have investigated whether apolipoprotein E, a major component of the cholesterol transport system in human beings, is a modifier of the clinical severity of Smith-Lemli-Opitz syndrome. METHOD: Common apo E, DHCR7, and LDLR genotypes were determined in 137 biochemically characterised patients with Smith-Lemli-Opitz syndrome and 59 of their parents. RESULTS: There was a significant correlation between patients' clinical severity scores and maternal apo E genotypes (p = 0.028) but not between severity scores and patients' or paternal apo E genotypes. In line with their effects on serum cholesterol levels, the maternal apo epsilon2 genotypes were associated with a severe Smith-Lemli-Opitz syndrome phenotype, whereas apo E genotypes without the epsilon2 allele were associated with a milder phenotype. The correlation of maternal apo E genotype with disease severity persisted after stratification for DHCR7 genotype. There was no association of Smith-Lemli-Opitz syndrome severity with LDLR gene variation. CONCLUSIONS: These results suggest that the efficiency of cholesterol transport from the mother to the embryo is affected by the maternal apo E genotype and extend the role of apo E and its disease associations to modulation of embryonic development and malformations. PMID- 15286152 TI - Positive association of the DIO2 (deiodinase type 2) gene with mental retardation in the iodine-deficient areas of China. AB - BACKGROUND: Iodine deficiency is the commonest cause of preventable mental retardation (MR) worldwide. However, in iodine-deficient areas not everyone is affected and familial aggregation is common. This suggests that genetic factors may also contribute. Thyroid hormone (TH) plays an important role in fetal and early postnatal brain development. The pro-hormone T4 (3,3',5,5' triiodothyronine) is converted in the brain to its active form, T3, or its inactive metabolite, reverse T3, mainly by the action of deiodinase type 2 (DIO2). METHODS: To investigate the potential genetic contribution of the DIO2 gene, we performed a case-control association study using three common SNPs in the gene (rs225014, rs225012, and rs225010) that were in strong linkage disequilibrium with each other. RESULTS: Single marker analysis showed a positive association of MR with rs225012 and rs225010. Particularly with rs255012 [corrected], CC [corrected] genotype frequency was significantly higher in MR cases than in controls (chi squared [corrected] = 9.18, p = 0.00246). When we compared the distributions of common haplotypes, we also found significant differences between mental retardation and controls in the haplotype combination of rs225012 and rs225010 (chi2 = 15.04, df 2, global p = 0.000549). This association remained significant after Bonferroni correction (p = 0.0016470). CONCLUSION: We conclude that allelic variation in the DIO2 gene may affect the amount of T3 available and in an iodine-deficient environment may partly determine overall risk of MR. PMID- 15286153 TI - Mutations of ESPN cause autosomal recessive deafness and vestibular dysfunction. AB - We mapped a human deafness locus DFNB36 to chromosome 1p36.3 in two consanguineous families segregating recessively inherited deafness and vestibular areflexia. This phenotype co-segregates with either of two frameshift mutations, 1988delAGAG and 2469delGTCA, in ESPN, which encodes a calcium-insensitive actin bundling protein called espin. A recessive mutation of ESPN is known to cause hearing loss and vestibular dysfunction in the jerker mouse. Our results establish espin as an essential protein for hearing and vestibular function in humans. The abnormal vestibular phenotype associated with ESPN mutations will be a useful clinical marker for refining the differential diagnosis of non-syndromic deafness. PMID- 15286154 TI - A pleiomorphic GH pituitary adenoma from a Carney complex patient displays universal allelic loss at the protein kinase A regulatory subunit 1A (PRKARIA) locus. AB - Carney complex (CNC) is a familial multiple endocrine neoplasia syndrome associated with GH-producing pituitary tumours and transmitted as an autosomal dominant trait. Mutations of the PRKAR1A gene are responsible for approximately half the known CNC cases but have never found in sporadic pituitary tumours. Pituitary tissue was obtained from an acromegalic CNC patient heterozygote for a common (PRKARIA)i-inactivating mutation. Both immunohistochemistry and electron microscopy showed a highly pleiomorphic pituitary adenoma. The cell culture population appeared morphologically heterogeneous and remained so after more than 30 passages. The mixture was comprised of cells strongly immunostained for GH, spindle-shaped myofibroblast-like cells, and cuboid cells with large axonal projections (negative for GH). The population appeared to have both epithelial and mesenchymal cells. Both at baseline and at passage 30, cytogenetic analysis indicated the presence of normal 46, XY diploid karyotype, whereas losses of the PRKARIA(i) locus were demonstrated in more than 98% of the cells by fluorescent in situ hybridisation, supporting this gene's involvement in pituitary tumorigenesis. Allelic loss may have occurred in a single precursor cell type that differentiated and clonally expanded into several phenotypes. Epithelial-to mesenchymal transition may also occur in CNC-associated pleiomorphic pituitary adenomas. PMID- 15286155 TI - Genomic imprinting of PPP1R9A encoding neurabin I in skeletal muscle and extra embryonic tissues. PMID- 15286156 TI - Homozygous missense mutation in the lamin A/C gene causes autosomal recessive Hutchinson-Gilford progeria syndrome. PMID- 15286157 TI - Molecular analysis of the mitochondrial 12S rRNA and tRNASer(UCN) genes in paediatric subjects with non-syndromic hearing loss. PMID- 15286158 TI - A novel mutation in the ATP1A2 gene causes alternating hemiplegia of childhood. PMID- 15286159 TI - Polymorphisms in the mannose binding lectin gene affect the cystic fibrosis pulmonary phenotype. PMID- 15286160 TI - A novel neurodegenerative disease characterised by posterior column ataxia and pyramidal tract involvement maps to chromosome 8p12-8q12.1. PMID- 15286161 TI - A new method for autozygosity mapping using single nucleotide polymorphisms (SNPs) and EXCLUDEAR. PMID- 15286163 TI - Ratio of female to male offspring of women tested for BRCA1 and BRCA2 mutations. PMID- 15286162 TI - Novel mutations in the gene SALL4 provide further evidence for acro-renal-ocular and Okihiro syndromes being allelic entities, and extend the phenotypic spectrum. PMID- 15286164 TI - The congenital myasthenic syndrome mutation RAPSN N88K derives from an ancient Indo-European founder. PMID- 15286165 TI - Autosomal recessive erythropoietic protoporphyria in the United Kingdom: prevalence and relationship to liver disease. PMID- 15286166 TI - A novel mutation in the Connexin 46 gene causes autosomal dominant congenital cataract with incomplete penetrance. PMID- 15286167 TI - Clinical features of type 2 Stickler syndrome. PMID- 15286168 TI - Expression analysis of an FGFR2 IIIc 5' splice site mutation (1084+3A->G). PMID- 15286170 TI - All the lonely people: by Caveman. PMID- 15286169 TI - Recurrent 17 bp duplication in PITX3 is primarily associated with posterior polar cataract (CPP4). PMID- 15286171 TI - ATP-dependent nucleosome remodelling: factors and functions. PMID- 15286172 TI - Interview with Caroline Damsky. PMID- 15286173 TI - MAP kinases as structural adaptors and enzymatic activators in transcription complexes. AB - Mitogen-activated protein kinase (MAPK) pathways regulate eukaryotic gene expression in response to extracellular stimuli. MAPKs and their downstream kinases phosphorylate transcription factors, co-regulators and chromatin proteins to initiate transcriptional changes. However, the spatial context in which the MAPKs operate in transcription complexes is poorly understood. Recent findings in budding yeast show that MAPKs can form integral components of transcription complexes and have novel structural functions in addition to phosphorylating local substrates. Hog1p MAPK is stably recruited to target promoters by specific transcription factors in response to osmotic stress, and acts as both a structural adaptor and enzymatic activator driving the assembly and activation of the transcription complex. We review the evidence that suggests a similar bifunctional role for MAPKs in mammalian transcription complexes. PMID- 15286174 TI - Aminoacyl-tRNA synthetase complexes: beyond translation. AB - Although aminoacyl-tRNA synthetases (ARSs) are housekeeping enzymes essential for protein synthesis, they can play non-catalytic roles in diverse biological processes. Some ARSs are capable of forming complexes with each other and additional proteins. This characteristic is most pronounced in mammals, which produce a macromolecular complex comprising nine different ARSs and three additional factors: p43, p38 and p18. We have been aware of the existence of this complex for a long time, but its structure and function have not been well understood. The only apparent distinction between the complex-forming ARSs and those that do not form complexes is their ability to interact with the three non enzymatic factors. These factors are required not only for the catalytic activity and stability of the associated ARSs, such as isoleucyl-, methionyl-, and arginyl tRNA synthetase, but also for diverse signal transduction pathways. They may thus have joined the ARS community to coordinate protein synthesis with other biological processes. PMID- 15286175 TI - Inner envelope protein 32 is imported into chloroplasts by a novel pathway. AB - The 32 kDa chloroplast inner envelope protein (IEP32) is imported into the organelle in the absence of a cleavable N-terminal pre-sequence. The ten N terminal amino acids form an essential portion of this targeting information as deduced from deletion mutants. Recognition and translocation of IEP32 is not catalysed by the general chloroplast outer envelope translocon subunits Toc159, Toc75III and Toc34, because IEP32 import is neither inhibited by proteolytic removal of Toc34 and Toc159 nor by inhibition of the Toc75 import channel by CuCl(2) or spermine. Import of IEP32 only requires ATP concentrations of below 20 microM indicating that stromal chaperones are not involved in the process, but that IEP32 might be directly inserted from the intermembrane space into the inner envelope by a so far unidentified pathway. IEP32 may require the assistance of Tic22, an intermembrane space translocon subunit for import as indicated by the presence of a chemical crosslinked product between both polypeptides. PMID- 15286176 TI - Human Dlg protein binds to the envelope glycoproteins of human T-cell leukemia virus type 1 and regulates envelope mediated cell-cell fusion in T lymphocytes. AB - Human homologue of the Drosophila Dlg tumor suppressor (hDlg) is a widely expressed scaffold protein implicated in the organization of multi-protein complexes at cell adhesion sites such as the neuronal synapse. hDlg contains three PDZ domains that mediate its binding to the consensus motifs present at the C-termini of various cell surface proteins, thus inducing their clustering and/or stabilization at the plasma membrane. Using a yeast two-hybrid screen, we identified hDlg as a cellular binding partner of a viral membrane integral protein, the envelope glycoprotein (Env) of human T-cell leukemia virus type 1 (HTLV-1). HTLV-1 is a human retrovirus that infects CD4+ T lymphocytes and is preferentially transmitted via direct contacts between infected and target cells, through a structure referred to as the virological synapse. Here, we demonstrate that hDlg interacts with a classical PDZ domain-binding motif present at the C terminus of the cytoplasmic domain of HTLV-1 Env and conserved in the related HTLV-2 virus. We further document that, in HTLV-1 infected primary T cells, hDlg and Env are concentrated in restricted areas of the plasma membrane, enriched in molecules involved in T-cell contacts. The presence of Gag proteins responsible for viral assembly and budding in these areas indicated that they constitute platforms for viral assembly and transmission. Finally, a mutant virus unable to bind hDlg exhibited a decreased ability to trigger Env mediated cell fusion between T lymphocytes. We thus propose that hDlg stabilizes HTLV-1 envelope glycoproteins at the virological synapse formed between infected and target cells, hence assisting the cell-to-cell transmission of the virus. PMID- 15286177 TI - Peroxisome elongation and constriction but not fission can occur independently of dynamin-like protein 1. AB - The mammalian dynamin-like protein DLP1 belongs to the dynamin family of large GTPases, which have been implicated in tubulation and fission events of cellular membranes. We have previously shown that the expression of a dominant-negative DLP1 mutant deficient in GTP hydrolysis (K38A) inhibited peroxisomal division in mammalian cells. In this study, we conducted RNA interference experiments to 'knock down' the expression of DLP1 in COS-7 cells stably expressing a GFP construct bearing the C-terminal peroxisomal targeting signal 1. The peroxisomes in DLP1-silenced cells were highly elongated with a segmented morphology. Ultrastructural and quantitative studies confirmed that the tubular peroxisomes induced by DLP1-silencing retained the ability to constrict their membranes but were not able to divide into spherical organelles. Co-transfection of DLP1 siRNA with Pex11pbeta, a peroxisomal membrane protein involved in peroxisome proliferation, induced further elongation and network formation of the peroxisomal compartment. Time-lapse microscopy of living cells silenced for DLP1 revealed that the elongated peroxisomes moved in a microtubule-dependent manner and emanated tubular projections. DLP1-silencing in COS-7 cells also resulted in a pronounced elongation of mitochondria, and in more dispersed, elongated Golgi structures, whereas morphological changes of the rER, lysosomes and the cytoskeleton were not detected. These observations clearly demonstrate that DLP1 acts on multiple membranous organelles. They further indicate that peroxisomal elongation, constriction and fission require distinct sets of proteins, and that the dynamin-like protein DLP1 functions primarily in the latter process. PMID- 15286178 TI - Lateral diffusion of Toll-like receptors reveals that they are transiently confined within lipid rafts on the plasma membrane. AB - The innate immune system utilises pattern recognition receptors in order to recognise microbial conserved molecular patterns. The family of Toll-like receptors (TLRs) has been shown to act as the main pattern recognition receptors for the innate immune system. Using biochemical as well as fluorescence imaging techniques, TLR2 and TLR4 were found to be recruited within microdomains upon stimulation by bacterial products. Furthermore their lateral diffusion in the cell membrane as determined by fluorescence recovery after photobleaching revealed that upon stimulation by bacterial products TLRs encounter barriers to their lateral movement, thus supporting the notion that specialised domains on the plasma membrane facilitate the innate recognition. PMID- 15286179 TI - The roles of MAPK cascades in synaptic plasticity and memory in Aplysia: facilitatory effects and inhibitory constraints. AB - Synaptic plasticity is thought to contribute to memory formation. Serotonin induced facilitation of sensory-motor (SN-MN) synapses in Aplysia is an extensively studied cellular analog of memory for sensitization. Serotonin, a modulatory neurotransmitter, is released in the CNS during sensitization training, and induces three temporally and mechanistically distinct phases of SN MN synaptic facilitation. The role of protein kinase A and protein kinase C in SN MN synaptic facilitation is well documented. Recently, it has become clear that mitogen-activated protein kinase (MAPK) cascades also play a critical role in SN MN plasticity. Here, we summarize the roles of MAPK cascades in synaptic plasticity and memory for sensitization in Aplysia. PMID- 15286180 TI - Protein synthesis inhibition blocks consolidation of an acrobatic motor skill. AB - To investigate whether motor skill learning depends on de novo protein synthesis, adult rats were trained in an acrobatic locomotor task (accelerating rotarod) for 7 d. Animals were systemically injected with cycloheximide (CHX, 0.5 mg/kg, i.p.) 1 h before sessions 1 and 2 or sessions 2 and 3. Control rats received vehicle injections before sessions 1, 2, and 3. Although CHX did not affect improvement of performance within session 1, between-session improvement was impaired. In overtrained animals, comparable injections of CHX had no effect on rotarod performance. These findings suggest that consolidation of motor skills requires protein synthesis. PMID- 15286181 TI - Providing explicit information disrupts implicit motor learning after basal ganglia stroke. AB - Despite their purported neuroanatomic and functional isolation, empirical evidence suggests that sometimes conscious explicit processes can influence implicit motor skill learning. Our goal was to determine if the provision of explicit information affected implicit motor-sequence learning after damage to the basal ganglia. Individuals with stroke affecting the basal ganglia (BG) and healthy controls (HC) practiced a continuous implicit motor-sequencing task; half were provided with explicit information (EI) and half were not (No-EI). The focus of brain damage for both BG groups was in the putamen. All of the EI participants were at least explicitly aware of the repeating sequence. Across three days of practice, explicit information had a differential effect on the groups. Explicit information disrupted acquisition performance in participants with basal ganglia stroke but not healthy controls. By retention (day 4), a dissociation was apparent--explicit information hindered implicit learning in participants with basal ganglia lesions but aided healthy controls. It appears that after basal ganglia stroke explicit information is less helpful in the development of the motor plan than is discovering a motor solution using the implicit system alone. This may be due to the increased demand placed on working memory by explicit information. Thus, basal ganglia integrity may be a crucial factor in determining the efficacy of explicit information for implicit motor-sequence learning. PMID- 15286182 TI - Calcium signaling in mitral cell dendrites of olfactory bulbs of neonatal rats and mice during olfactory nerve Stimulation and beta-adrenoceptor activation. AB - Synapses formed by the olfactory nerve (ON) provide the source of excitatory synaptic input onto mitral cells (MC) in the olfactory bulb. These synapses, which relay odor-specific inputs, are confined to the distally tufted single primary dendrites of MCs, the first stage of central olfactory processing. beta adrenergic modulation of electrical and chemical signaling at these synapses may be involved in early odor preference learning. To investigate this possibility, we combined electrophysiological recordings with calcium imaging in olfactory bulb slices prepared from neonatal rats and mice. Activation of ON-MC synapses induced postsynaptic potentials, which were associated with large postsynaptic calcium transients. Neither electrical nor calcium responses were affected by beta-adrenergic agonists or antagonist. Immunocytochemical analysis of MCs and their tufted dendrites revealed clear immunoreactivity with antibodies against alpha1A (Cav2.1, P/Q-type) and alpha1B (Cav2.2, N-type), but not against alpha1C (Cav1.2, L-type) or alpha1D (Cav1.3, L-type) calcium channel subunits. Moreover, nimodipine, a blocker of L-type calcium channels, had no effect on either electrical or calcium signaling at ON-MC synapses. In contrast to previous evidence, we concluded that in neonatal rats and mice (P5-P8), mitral cells do not express significant amounts of L-type calcium channels, the calcium channel type that is often targeted by beta-adrenergic modulation. The absence of beta adrenergic modulation on either electrical or calcium signaling at ON-MC synapses of neonatal rats and mice excludes the involvement of this mechanism in early odor preference learning. PMID- 15286183 TI - Preserved fronto-striatal plasticity and enhanced procedural learning in a transgenic mouse model of Alzheimer's disease overexpressing mutant hAPPswe. AB - Mutations in the amyloid precursor protein (APP) gene inducing abnormal processing and deposition of beta-amyloid protein in the brain have been implicated in the pathogenesis of Alzheimer's disease (AD). Although Tg2576 mice with the Swedish mutation (hAPPswe) exhibit age-related Abeta-plaque formation in brain regions like the hippocampus, the amygdala, and the cortex, these mice show a rather specific deficit in hippocampal-dependent learning and memory tasks. In view of recent findings showing that neural systems subserving different forms of learning are not simply independent but that depressing or enhancing one system affects learning in another system, we decided to investigate fronto-striatal synaptic plasticity and related procedural learning in these mutants. Fronto striatal long-term depression (LTD) induced by tetanic stimulation of the cortico striatal input was similar in Tg2576 and wild-type control mice. Behavioral data, however, pointed to an enhancement of procedural learning in the mutants that showed robust motor-based learning in the cross maze and higher active avoidance scores. Thus, in this mouse model of AD, an intact striatal function associated with an impaired hippocampal function seems to provide neural conditions favorable to procedural learning. Our results suggest that focusing on preserved or enhanced forms of learning in AD patients might be of interest to describe the functional reorganization of the brain when one memory system is selectively compromised by neurological disease. PMID- 15286184 TI - Contributions of striatal subregions to place and response learning. AB - The involvement of different subregions of the striatum in place and response learning was examined using a T-maze. Rats were given NMDA lesions of the dorsolateral striatum (DLS), anterior dorsomedial striatum (ADMS), posterior dorsomedial striatum (PDMS), or sham surgery. They were then trained to retrieve food from the west arm of the maze, starting from the south arm, by turning left at the choice point. After 7 d of training, with four trials a day, a probe test was given in which the starting arm is inserted as the north arm, at the opposite side of the maze. A left turn would indicate a "response" strategy; a right turn, a "place" strategy. The rats were then trained for 7 more days, followed by a second probe test. Unlike rats in the other groups, most of the rats in the PDMS group turned left, using the response strategy on both probe tests. These results suggest that the PDMS plays a role in spatially guided behavior. PMID- 15286185 TI - Overexpectation: response loss during sustained stimulus compounding in the rabbit nictitating membrane preparation. AB - Rabbits were given reinforced training of the nictitating membrane (NM) response using separate conditioned stimuli (CSs), which were a tone, light, and/or tactile vibration. Then, two CSs were compounded and given further pairings with the unconditioned stimulus (US). Evidence of both overexpectation and summation effects appeared. That is, responding to the individual CSs declined despite their continued pairing with the US on compound trials (overexpectation), and responding on the compound trials was greater than responding to the individual CSs (summation). The response loss appeared regardless of the testing regime, that is, whether the test presentations of the individual CSs were themselves reinforced (Experiment 2), not reinforced (Experiment 1), or deferred until the end of compound training (Experiment 2). The results are discussed with respect to the roles of excitatory versus inhibitory processes, elemental versus configural processes, and the possible roles of cerebellar and hippocampal pathways. PMID- 15286186 TI - The surgeon's job: how should we assess the trainee? PMID- 15286187 TI - Healing in survivors of torture. PMID- 15286188 TI - New approaches to the diagnosis of psychopathy and personality disorder. PMID- 15286189 TI - Cognitive dysfunction in homeless adults: a systematic review. AB - Homelessness is associated with multiple adversities that might impact upon brain function. We performed a review of published work to assess evidence of cognitive dysfunction among adults who are homeless. Despite liberal inclusion criteria only seventeen publications were identified, these describing eighteen samples mainly from the USA. Although the total number of individuals studied is small (about 3300) and the samples are heterogeneous, most studies indicate a considerable burden of cognitive dysfunction among homeless people. Such dysfunction might be expected to impact upon their ability to reintegrate into society, thereby undermining policies of inclusiveness. In clinical practice, assessment of homeless adults should include their cognitive state. PMID- 15286190 TI - Doctors' confusion over ratios and percentages in drug solutions: the case for standard labelling. AB - The different ways of expressing concentrations of drugs in solution, as ratios or percentages or mass per unit volume, are a potential cause of confusion that may contribute to dose errors. To assess doctors' understanding of what they signify, all active subscribers to doctors.net.uk, an online community exclusively for UK doctors, were invited to complete a brief web-based multiple choice questionnaire that explored their familiarity with solutions of adrenaline (expressed as a ratio), lidocaine (expressed as a percentage) and atropine (expressed in mg per mL), and their ability to calculate the correct volume to administer in clinical scenarios relevant to all specialties. 2974 (24.6%) replied. The mean score achieved was 4.80 out of 6 (SD 1.38). Only 85.2% and 65.8% correctly identified the mass of drug in the adrenaline and lidocaine solutions, respectively, whilst 93.1% identified the correct concentration of atropine. More would have administered the correct volume of adrenaline and lidocaine in clinical scenarios (89.4% and 81.0%, respectively) but only 65.5% identified the correct volume of atropine. The labelling of drug solutions as ratios or percentages is antiquated and confusing. Labelling should be standardized to mass per unit volume. PMID- 15286191 TI - Mapping rehabilitation resources for head injury. AB - Several reports have pointed to the unevenness in the UK of services for rehabilitation after head injury. A study was conducted in the Eastern Region of England to define the key stages in recovery and rehabilitation, by an iterative process of questionnaire, interview and consensus conference. Findings were translated into a draft set of maps showing current availability of services which were revised after feedback. Working groups then developed a set of definitions and classification codes for each stage of rehabilitation which were likewise disseminated for feedback. The maps were then redrafted to correspond with the definitions together with a flowchart of potential head injury rehabilitation services. The definitions were piloted at a regional neurosurgery unit and a rehabilitation hospital. Core services for neurorehabilitation region wide were found to be variable and uncoordinated with fragmented and inequitable allocation of resources. The definitions and mapping system that emerged from this study should facilitate the design of care pathways for patients and identify gaps in the services. PMID- 15286192 TI - Mesothelioma and a high potassium. PMID- 15286193 TI - Return of a normal functioning spleen after traumatic splenectomy. PMID- 15286194 TI - Pulmonary embolism caused by Candida albicans. PMID- 15286195 TI - An epigastric swelling. PMID- 15286196 TI - Penicillium marneffei: a pathogen on our doorstep? PMID- 15286197 TI - Channelling the Emperor: what really killed Napoleon? AB - Arsenic was present in Napoleon's hair before he arrived on Saint Helena and the findings at necropsy are consistent only with the diagnosis of ulcerating, regionally invasive, gastric carcinoma. The question of whether Napoleon died of, or merely with, arsenic poisoning is illuminated by developments in the treatment of promyelocytic leukaemia. Arsenic trioxide induces remission in many, but treatment can be complicated by QT prolongation, torsades de pointes and sudden death. At clinically relevant concentrations, arsenic blocks both I(Kr) and I(ks) channels and, at the same time, activates I(K-ATP) channels. The balance of these forces is easily disrupted, and QT prolongation is worsened by hypokalaemia. Napoleon was chronically treated with tartar emetic for gastrointestinal symptoms, and the day before he died he was given a huge dose of calomel (mercurous chloride) as a purgative. Both treatments would have caused potassium wastage. In addition, the Emperor was being treated with a decoction containing 'bark'-presumably 'Jesuit's bark'. The quinine in Jesuit's bark is another cause of QT prolongation. It is likely that the immediate cause of the Emperor's death was torsades de pointes, brought on by chronic exposure to arsenic and a medication error. PMID- 15286198 TI - French leave. PMID- 15286199 TI - Effectiveness of cycle helmets and the ethics of legislation. PMID- 15286201 TI - Osteophytes and throat symptoms. PMID- 15286202 TI - Art about hospitals. PMID- 15286203 TI - Rapid-access ultrasonography for testicular lesions. PMID- 15286204 TI - Addison and Bright at the St Alban's Club. PMID- 15286206 TI - Long-term administration of Delta9-tetrahydrocannabinol desensitizes CB1-, adenosine A1-, and GABAB-mediated inhibition of adenylyl cyclase in mouse cerebellum. AB - Cannabinoid CB(1) receptors in the cerebellum mediate the inhibitory effects of Delta(9)-tetrahydrocannabinol (THC) on motor coordination. Intracellular effects of CB(1) receptors include inhibition of adenylyl cyclase via activation of G(i/o) proteins. There is evidence for the convergence of other neuronal receptors, such as adenosine A(1) and GABA(B), with the cannabinoid system on this signaling pathway to influence motor function. Previous studies have shown that brain CB(1) receptors are desensitized and down-regulated by long-term THC treatment, but few studies have examined the effects of long-term THC treatment on downstream effector activity in brain. Therefore, these studies examined the relationship between CB(1), adenosine A(1), and GABA(B) receptors in cerebella of mice undergoing prolonged treatment with vehicle or THC at the level of G protein activation and adenylyl cyclase inhibition. In control cerebella, CB(1) receptors produced less than additive inhibition of adenylyl cyclase with GABA(B) and A(1) receptors, indicating that these receptors are localized on overlapping populations of cells. Long-term THC treatment produced CB(1) receptor down regulation and desensitization of both cannabinoid agonist-stimulated G protein activation and inhibition of forskolin-stimulated adenylyl cyclase. However, G protein activation by GABA(B) or A(1) receptors was unaffected. It is noteworthy that heterologous attenuation of GABA(B) and A(1) receptor-mediated inhibition of adenylyl cyclase was observed, even though absolute levels of basal and forskolin or G(s)-stimulated activity were unchanged. These results indicate that long term THC administration produces a disruption of inhibitory receptor control of cerebellar adenylyl cyclase and suggest a potential mechanism of cross-tolerance to the motor incoordinating effects of cannabinoid, GABA(B), and A(1) agonists. PMID- 15286207 TI - Molecular determinants of frequency dependence and Ca2+ potentiation of verapamil block in the pore region of Cav1.2. AB - Verapamil block of Ca(v)1.2 is frequency-dependent and potentiated by Ca(2+). We examined the molecular determinants of these characteristics using mutations that effect Ca(2+) interactions with Ca(v)1.2. Mutant and wild-type Ca(v)1.2 channels were transiently expressed in tsA 201 cells with beta(1b) and alpha(2)delta subunits. The four conserved glutamates that compose the Ca(2+) selectivity filter in Ca(v)1.2 were mutated to Gln (E363Q, E709Q, E1118Q, E1419Q) and the adjacent conserved threonine in each domain was mutated to Ala (T361A, T707A, T1116A, T1417A). The L-type-specific residues in the domain III pore region (F1117G) and the C-terminal tail (I1627A) were also mutated and assayed for block by verapamil using whole-cell voltage-clamp recordings in 10 mM Ba(2+) or 10 mM Ca(2+). In Ba(2+), none of the pore-region mutations reduced the fraction of current blocked by 30 microM verapamil at 0.05 Hz stimulation. However, all of the pore-region mutations abolished Ca(2+) potentiation of verapamil block at 0.05 Hz. The T1116A, F1117G, E1118Q, and E1419Q mutations all significantly reduced frequency-dependent verapamil block (1-Hz stimulation) in both Ba(2+) and Ca(2+). The I1627A mutation, which disrupts Ca(2+)-dependent inactivation, increased the fraction of closed channels blocked by 30 microM verapamil in Ba(2+) but did not affect frequency-dependent block in Ba(2+) or Ca(2+). Our data suggest that the pore region of domain III may contribute to a high affinity verapamil binding site accessed during 1-Hz stimulation and that Ca(2+) binding to multiple sites may be required for potentiation of verapamil block of closed channels. PMID- 15286208 TI - Specific inhibition of nuclear factor-kappaB-dependent inflammatory responses by cell type-specific mechanisms upon A2A adenosine receptor gene transfer. AB - Adenosine is a potent inhibitor of inflammatory processes, and the A(2A) adenosine receptor (A(2A)AR) plays a key nonredundant role as a suppresser of inflammatory responses in vivo. In this study, we demonstrate that increasing A(2A)AR gene expression suppressed multiple inflammatory responses in both human umbilical vein endothelial cells (HUVECs) and rat C6 glioma cells in vitro. In particular, the induction of the adhesion molecule E-selectin by either tumor necrosis factor alpha (TNFalpha) or Escherichia coli lipopolysaccharide (LPS) was reduced by more than 70% in HUVECs, whereas inducible nitric-oxide synthase (iNOS) induction was abolished in C6 cells after exposure to interferon-gamma in combination with LPS and TNFalpha, suggesting that the receptor inhibited a common step in the induction of each of these pro-inflammatory genes. Consistent with this hypothesis, A(2A)AR expression inhibited the activation of NF-kappaB, a key transcription factor whose proper function was essential for optimal iNOS and E-selectin induction. However, although NF-kappaB binding to target DNA was severely compromised in both cell types, the mechanisms by which this occurred were distinct. In C6 cells, A(2A)AR expression blocked IkappaBalpha degradation by inhibiting stimulus-induced phosphorylation, whereas in HUVECs, A(2A)AR expression inhibited NF-kappaB translocation to the nucleus independently of any effect on IkappaBalpha degradation. Together, these observations suggest that A(2A)AR-mediated inhibition NF-kappaB activation is a critical aspect of its anti inflammatory signaling properties and that the molecular basis of this inhibition varies in a cell type-specific manner. PMID- 15286209 TI - Regulation of cyclin-dependent kinase 5 and calcium/calmodulin-dependent protein kinase II by phosphatidylinositol-linked dopamine receptor in rat brain. AB - A brain dopamine receptor that modulates phosphatidylinositol (PI) metabolism via the activation of phospholipase Cbeta (PLCbeta) has been described previously. The present study aims to define the downstream signaling cascade initiated by the PI-linked dopamine receptor. Incubation of rat brain frontal cortical slices with 6-chloro-7,8-dihydroxy-3-methyl-1-(3-methylphenyl)-2,3,4,5-tetrahydro-1H-3 benzazepine (SKF83959), a recently identified selective agonist of the PI-linked D1-like dopamine receptor, elicited transient time- and dose-dependent stimulations of cyclin-dependent kinase 5 (cdk5) and calcium/calmodulin-dependent protein kinase II (CaMK II) activities. The stimulation of these kinases is blocked by 20 microM R-(+)-7-chloro-8-hydroxy-3-methyl-1-phenyl-2,3,4,5 tetrahydro-1H-3-benzazepine (SCH23390) or the PLCbeta antagonist 1-[6-[[17beta methoxyestra-1,3,5(10)-trien-17-yl]amino]hexyl]-1H-pyrrole-2,5-dione (U-73122) and is attenuated by the protein kinase inhibitor calphostin C or by the intracellular calcium chelator BAPTA, indicating that SKF83959 stimulates cdk5 and CaMK II activities via a PI-linked D1-like dopamine receptor, and PLCbeta and is dependent on protein kinase C and calcium. Although cdk5 and CaMK II are physically associated in native brain tissue, no change in this association was observed in response to SKF83959 stimulation or to the inhibition of either cdk5 by roscovitine or of CaMK by 2-[N-(2-hydroxyethyl)]-N-(4 methoxybenzenesulfonyl)]amino-N-(4-chlorocinnamyl)-N-methylbenzylamine) (KN93), suggesting that SKF83959-mediated stimulation of cdk5 or CaMK II is independent of the other kinase and that the association of the two kinases is not modulated by change of kinase activity. Moreover, we found that cdk5 phosphorylates dopamine and cAMP-regulated phosphoprotein at Thr75, whereas CaMK II is responsible for the activation of cAMP response element-binding protein in response to SKF83959 stimulation. The present data provide the first insight into the signaling mechanism for the PI-linked dopamine receptor. This information, in turn, may help in exploring the functional consequences of stimulation of this brain receptor. PMID- 15286210 TI - Mapping of maurotoxin binding sites on hKv1.2, hKv1.3, and hIKCa1 channels. AB - Maurotoxin (MTX) is a potent blocker of human voltage-activated Kv1.2 and intermediate-conductance calcium-activated potassium channels, hIKCa1. Because its blocking affinity on both channels is similar, although the pore region of these channels show only few conserved amino acids, we aimed to characterize the binding sites of MTX in these channels. Investigating the pH(o) dependence of MTX block on current through hKv1.2 channels, we concluded that the block is less pH(o) - sensitive than for hIKCa1 channels. Using mutant cycle analysis and computer docking, we tried to identify the amino acids through which MTX binds to hKv1.2 and hIKCa1 channels. We report that MTX interacts with hKv1.2 mainly through six strong interactions. Lys(23) from MTX protrudes into the channel pore interacting with the GYGD motif, whereas Tyr(32) and Lys(7) interact with Val(381), Asp(363), and Glu(355), stabilizing the toxin onto the channel pore. Because only Val(381), Asp(363), and the GYGD motif are conserved in hIKCa1 channels, and the replacement of His(399) from hKv1.3 channels with a threonine makes this channel MTX-sensitive, we concluded that MTX binds to all three channels through the same amino acids. Glu(355), although important, is not essential in MTX recognition. A negatively charged amino acid in this position could better stabilize the toxin-channel interaction and could explain the pH(o) sensitivity of MTX block on current through hIKCa1 versus hKv1.2 channels. PMID- 15286211 TI - Effect of folic acid on homocysteine-induced trophoblast apoptosis. AB - In trophoblast cells exposed to homocysteine (Hcy) we observed cellular apoptosis and the inhibition of trophoblast functions. Because folate and Hcy, linked in the same metabolic pathway, are inversely related, we investigated the role of folic acid in reversing the Hcy effect in human placenta. In primary trophoblast cells we examined the cytosolic release of cytochrome c, both M30 and terminal deoxynucleotidyl transferase-mediated dUDP nick-end labelling (TUNEL) and DNA laddering. Hcy (20 micromol/l) treatment resulted in cytochrome c release from mitochondria to the cytosol, and an increased number of M30-positive trophoblast cells and TUNEL positive nuclei. Furthermore, DNA cleavage in agarose gel and the determination of histone-associated DNA fragments have been investigated. Homocysteine induced DNA fragmentation and significantly reduced hCG secretion. The addition of folic acid (20 nmol/l) resulted in inhibition of the effects of Hcy on human trophoblast. These results suggest a protective role of folic acid in the prevention of trophoblast apoptosis linked to Hcy. PMID- 15286212 TI - Comparative proteomes of the proliferating C(2)C(12) myoblasts and fully differentiated myotubes reveal the complexity of the skeletal muscle differentiation program. AB - When cultured in low serum-containing growth medium, the mouse C(2)C(12) cells exit cell cycle and undergo a well-defined program of differentiation that culminates in the formation of myosin heavy chain-positive bona fide multinucleated muscle cells. To gain an understanding into this process, we compared total, membrane- and nuclear-enriched proteins, and phospho-proteins from the proliferating C(2)C(12) cells and the fully differentiated myotubes by the combined methods of two-dimensional PAGE, quantitative PDQuest image analysis, and MS. Quantification of more than 2,000 proteins from C(2)C(12) myoblasts and myotubes revealed that a vast majority of the abundant proteins appear to be relegated to the essential, housekeeping and structural functions, and their steady state levels remain relatively constant. In contrast, 75 proteins were highly regulated during the phenotypic conversion of rapidly dividing C(2)C(12) myoblasts into fully differentiated, multi-nucleated, post mitotic myotubes. We found that differential accumulation of 26 phospho-proteins also occurred during conversion of C(2)C(12) myoblasts into myotubes. We identified the differentially expressed proteins by MALDI-TOF-MS and LC-ESI quadrupole ion trap MS/MS. We demonstrate that more than 100 proteins, some shown to be associated with muscle differentiation for the first time, that regulate inter- and intracellular signaling, cell shape, proliferation, apoptosis, and gene expression impinge on the mechanism of skeletal muscle differentiation. PMID- 15286213 TI - Epidemiology of childhood tuberculosis in the United States, 1993-2001: the need for continued vigilance. AB - OBJECTIVE: To describe trends and highlight epidemiologic and clinical characteristics of childhood tuberculosis (TB) in the United States. METHODS: All verified TB cases reported to the national TB surveillance system from 1993 to 2001 were included. A child was defined as a person younger than 15 years. RESULTS: A total of 11,480 childhood TB cases were reported. Case rates (TB cases/100,000 population) in all children declined from 2.9 (n = 1663) in 1993 to 1.5 (n = 931) in 2001. Among children, those who were younger than 5 years had the highest rate. California, Texas, and New York accounted for 48% of all childhood TB cases. In 2001, TB case rates were higher for foreign-born (12.2) than US-born children (1.1). Hispanic and non-Hispanic black children accounted for nearly three quarters of all cases. Twenty-four percent of children with TB were foreign-born children, with the largest number originating from Mexico (39.8%), the Philippines (8.6%), and Vietnam (5.7%). Most children had evidence of pulmonary TB disease (78.9%). Among culture-positive cases without previous TB, drug resistance to at least isoniazid was 7.3% and to isoniazid and rifampin was 1.6%. In 1999, 82.9% of children received directly observed therapy for at least part of their treatment and 94.8% completed treatment. CONCLUSIONS: Although the overall TB case number among children is declining in the United States, certain groups of children (eg, younger children, racial and ethnic minorities, foreign-born) are at higher risk for TB. As the United States moves toward the elimination of TB, future efforts should endeavor to prevent all cases of childhood TB. PMID- 15286214 TI - Developing community-specific recommendations for first-line treatment of acute otitis media: is high-dose amoxicillin necessary? AB - OBJECTIVES: National recommendations are to use high-dose amoxicillin (80-90 mg/kg per day) to treat uncomplicated acute otitis media (AOM) in children who are at high risk for infection with nonsusceptible Streptococcus pneumoniae (NSSP). However, high-dose treatment may not be necessary if the local prevalence of NSSP is low. The objective of this study was to estimate the local prevalence of NSSP in children with acute upper respiratory illnesses and to develop community-specific recommendations for first-line empiric treatment of AOM. METHODS: We conducted a cross-sectional prevalence study in the offices of 7 community pediatricians in St Louis, Missouri. S pneumoniae was isolated from nasopharyngeal swabs collected from children who were younger than 7 years and had AOM, nonspecific upper respiratory infection, cough, acute sinusitis, or pharyngitis. Children were excluded from the study when they had received an antibiotic in the previous 4-week period. Parents and providers completed a brief questionnaire to assess risk factors for carriage of NSSP. On the basis of National Clinical Chemistry Laboratory Standards, isolates with a penicillin minimum inhibitory concentration > or =0.12 microg/mL were considered to be nonsusceptible to penicillin (NSSP), and isolates with a penicillin minimum inhibitory concentration >2 microg/mL were categorized as nonsusceptible to standard-dose amoxicillin (35-45 mg/kg per day; NSSP-A). RESULTS: S pneumoniae was isolated from the nasopharynx of 85 (40%) of 212 study patients (95% confidence interval [CI]: 33%-47%); 41 (48%) of 85 isolates were NSSP (95% CI: 37%-59%), and 6 (7%) were NSSP-A (95% CI: 1.5%-13%). Among the 212 study patients, the prevalence of NSSP was 19% (95% CI: 14%-25%), and the prevalence of NSSP-A was 3% (95% CI: 0.6%-5%). Carriage of NSSP was increased in child care attendees compared with nonattendees (29% vs 14%; odds ratio: 2.6; 95% CI: 1.3 5.2). CONCLUSIONS: In our community, although the prevalence of NSSP among isolates of S pneumoniae identified from the nasopharynx of symptomatic children is high (48%), the probability of NSSP-A infection among symptomatic children is <5%. Our data support a recommendation to treat most children who have uncomplicated AOM with standard-dose amoxicillin. Children who attend child care or have recently received an antibiotic may require treatment with high-dose amoxicillin. Other communities may benefit from a similar assessment of the prevalence of NSSP and NSSP-A. PMID- 15286215 TI - Marginal increase in cost and excess length of stay associated with nosocomial bloodstream infections in surviving very low birth weight infants. AB - OBJECTIVE: Nosocomial bloodstream infections (NBIs) are associated with serious morbidity and prolonged length of stay (LOS) in very low birth weight (VLBW) infants. However, the marginal costs and excess LOS associated with these infections have never been measured in different birth weight (BW) categories after adjustment for many of the potentially confounding demographic variables, comorbidities, and treatments. The objective of this study was to measure the marginal cost and excess LOS caused by NBIs in surviving VLBW infants in different BW categories. METHODS: This retrospective study examined data previously collected as part of the Neonatal Intensive Care Quality Improvement Collaborative 2000 and the Vermont Oxford Network clinical outcomes database. Univariate analyses and multiple regression were used to examine the effect of NBIs on hospital costs and LOS. Seventeen neonatal intensive care units that participated in the Neonatal Intensive Care Quality Improvement Collaborative 2000 submitted both clinical and financial data on their VLBW infants who were born from January 1, 1998, to December 31, 1999. This study included data from both university and community hospitals. RESULTS: NBIs occurred in 19.7% of 2809 patients included in this study. NBI was associated with significantly increased treatment costs for infants with BW 751 to 1500 g. The marginal costs of NBIs, as estimated by multiple regression, varied from 5875 dollars for VLBW infants with a BW of 401 to 750 g to 12,80 dollars for those with BW of 751 to 1000 g. LOS was significantly increased in all BW categories. The excess LOS estimated by multiple regression varied from 4 days in VLBW infants with a BW of 1001 to 1251 g to 7 days in those with a BW of 751 to 1000 g. CONCLUSIONS: NBIs are associated with increased hospital treatment costs and LOS but by varying amounts depending on the BW. Preventing a single NBI could reduce the treatment costs of a VLBW infant by at least several thousand dollars. These savings are a greater percentage of the total treatment costs in VLBW infants with BW 1001 to 1500 g than in smaller infants. PMID- 15286216 TI - Triage of the pediatric patient in the emergency department: are we all in agreement? AB - OBJECTIVE: To compare triage categorization as a measure of perceived patient acuity on presentation to the emergency department by pediatric emergency medicine (PEM) attending physicians, nurses, and pediatric residents with their general emergency medicine (GEM) counterparts. METHODS: A questionnaire that contained 12 pediatric triage scenarios was sent to all PEM attending physicians, triage-trained nurses, and pediatric residents and their GEM counterparts at a large urban hospital with separate pediatric and general emergency departments. Participants were asked to use a 3-tier triage system (emergent, urgent, nonurgent) to assign a triage level for each patient scenario. RESULTS: The response rate was 99%. The kappa level of agreement was highest (.39) among the PEM physicians. Significantly more GEM attending physicians triaged the following scenarios at a higher acuity level as compared with PEM attending physicians with a trend toward emergent triage: simple febrile seizure, 50% (95% confidence interval [CI]: 30%-70%) versus 7.7% (95% CI: 1%-34%); 18-month-old with fever and bumps on lips, 21% (95% CI: 9%-43%) versus 0% (95% CI: 0%-23%); and 15-month-old well-appearing child with high fever, 50% (95% CI: 30%-70%) versus 7.7% (95% CI: 1%-34%). Significant differences were found between GEM and PEM triage-trained nurses only in the 15-month-old high fever scenario and between GEM and pediatric residents in the 15-month-old high fever scenario, the 18-month-old with fever and bumps on lips scenario, and a fever/limp scenario. CONCLUSIONS: The level of agreement of triage assignment within each group was only fair. GEM participants and PEM participants agreed on most scenarios. However, GEM participants were more likely to triage children with certain febrile illnesses at higher acuity levels as compared with their PEM counterparts. PMID- 15286217 TI - Initial hypoglycemia and neonatal brain injury in term infants with severe fetal acidemia. AB - OBJECTIVE: To determine the potential contribution of initial hypoglycemia to the development of neonatal brain injury in term infants with severe fetal acidemia. METHODS: A retrospective chart review was conducted of 185 term infants who were admitted to the neonatal intensive care unit between January 1993 and December 2002 with an umbilical arterial pH <7.00. Short-term neurologic outcome measures include death as a consequence of severe encephalopathy and evidence of moderate to severe encephalopathy with or without seizures. Hypoglycemia was defined as an initial blood glucose < or =40 mg/dL. RESULTS: Forty-one (22%) infants developed an abnormal neurologic outcome, including 14 (34%) with severe hypoxic ischemic encephalopathy who died, 24 (59%) with moderate to severe hypoxic ischemic encephalopathy, and 3 (7%) with seizures. Twenty-seven (14.5%) of the 185 infants had an initial blood sugar < or =40 mg/dL. Fifteen (56%) of 27 infants with a blood sugar < or =40 mg/dL versus 26 (16%) of 158 infants with a blood sugar >40 mg/dL had an abnormal neurologic outcome (odds ratio [OR]: 6.3; 95% confidence interval [CI]: 2.6-15.3). Infants with abnormal outcomes and a blood sugar < or =40 mg/dL versus >40 mg/dL had a higher pH (6.86 +/- 0.07 vs 6.75 +/- 0.09), a lesser base deficit (-19 +/- 4 vs -23.8 +/- 4 mEq/L), and lower mean arterial blood pressure (34 +/- 10 vs 45 +/- 14 mm Hg), respectively. There was no difference between groups in the proportion of infants who required cardiopulmonary resuscitation (7 [46%] vs 15 [57%]) and those with a 5-minute Apgar score <5 (11 [73%] vs 22 [85%]). By multivariate logistic analysis, 4 variables were significantly associated with abnormal outcome: initial blood glucose < or =40 mg/dL versus >40 mg/dL (OR: 18.5; 95% CI: 3.1-111.9), cord arterial pH < or =6.90 versus >6.90 (OR: 9.8; 95% CI: 2.1-44.7), a 5-minute Apgar score < or =5 versus >5 (OR: 6.4; 95% CI: 1.7-24.5), and the requirement for intubation with or without cardiopulmonary resuscitation versus neither (OR: 4.7; 95% CI: 1.2-17.9). CONCLUSION: Initial hypoglycemia is an important risk factor for perinatal brain injury, particularly in depressed term infants who require resuscitation and have severe fetal acidemia. It remains unclear, however, whether earlier detection of hypoglycemia, such as in the delivery room, in this population could modify subsequent neurologic outcome. PMID- 15286218 TI - Brain volumes in adult survivors of very low birth weight: a sibling-controlled study. AB - OBJECTIVES: To establish whether adults who were born very low birth weight (VLBW) show altered volumes of certain brain structures. METHODS: Unmatched case control study was conducted of 33 individuals from a cohort of VLBW (<1500 g) infants who were born between 1966 and 1977 and 18 of their normal birth weight siblings. Whole brain, gray matter, ventricular, corpus callosum, and hippocampal volumes were measured on structural magnetic resonance imaging scans. RESULTS: VLBW individuals had a 46% increase in total ventricular volume and a 17% reduction in posterior corpus callosum volume. No differences in whole brain, gray matter, or hippocampal volumes were observed. CONCLUSION: Specific differences exist in the volumes of certain brain structures in adults who were born VLBW compared with their normal birth weight siblings. PMID- 15286219 TI - Clinical outcomes of near-term infants. AB - OBJECTIVE: To test the hypothesis that near-term infants have more medical problems after birth than full-term infants and that hospital stays might be prolonged and costs increased. METHODS: Electronic medical record database sorting was conducted of 7474 neonatal records and subset analyses of near-term (n = 120) and full-term (n = 125) neonatal records. Cost information was accessed. Length of hospital stay, Apgar scores, clinical diagnoses (temperature instability, jaundice, hypoglycemia, suspicion of sepsis, apnea and bradycardia, respiratory distress), treatment with an intravenous infusion, delay in discharge to home, and hospital costs were assessed. RESULTS: Data from 90 near-term and 95 full-term infants were analyzed. Median length of stay was similar for near-term and full-term infants, but wide variations in hospital stay were documented for near-term infants after both vaginal and cesarean deliveries. Near-term and full term infants had comparable 1- and 5-minute Apgar scores. Nearly all clinical outcomes analyzed differed significantly between near-term and full-term neonates: temperature instability, hypoglycemia, respiratory distress, and jaundice. Near-term infants were evaluated for possible sepsis more frequently than full-term infants (36.7% vs 12.6%; odds ratio: 3.97) and more often received intravenous infusions. Cost analysis revealed a relative increase in total costs for near-term infants of 2.93 (mean) and 1.39 (median), resulting in a cost difference of 2630 dollars (mean) and 429 dollars (median) per near-term infant. CONCLUSIONS: Near-term infants had significantly more medical problems and increased hospital costs compared with contemporaneous full-term infants. Near term infants may represent an unrecognized at-risk neonatal population. PMID- 15286220 TI - Can an alternative umbilical arterial catheter solution and flush regimen decrease iatrogenic hemolysis while enhancing nutrition? A double-blind, randomized, clinical trial comparing an isotonic amino acid with a hypotonic salt infusion. AB - OBJECTIVE: In the process of sampling blood through an umbilical arterial catheter (UAC), infant blood comes into stagnant contact with infusion solution in the "waste syringe" before being reinfused. We have previously demonstrated in vitro that this process is associated with less hemolysis of red blood cells (RBCs) with use of an isotonic solution compared with a hypotonic 0.25 normal saline (NS) solution. The objective of this study was to compare the in vivo effect on hemolysis of 2 UAC infusion/flush regimens (an isotonic regimen vs a hypotonic regimen) and to assess the early nutritional benefit of an amino acid solution as the isotonic UAC infusion solution. METHODS: Infants who had a birth weight of < or =1.5 kg and were expected to have a UAC for > or =3 days were enrolled within 24 hours of life into this prospective, double-blind, randomized, clinical trial of 2 UAC infusion solution/flush regimens. Power analysis demonstrated that 40 infants were needed to determine differences in hemolysis quantified by plasma-free hemoglobin (PFH) level. Nutrition from glucose was evaluated by measurement of daily dextrose calories. C-peptide was measured to evaluate endogenous insulin production. Adverse events and protein tolerance were tracked. RESULTS: Twenty-two infants (mean gestational age: 27 weeks; 945 g birth weight) were enrolled in each group, for an average of 4.2 days (range: 2.5-8 days). There were no group differences in demographics. PFH levels were lower for infants who received isotonic amino acid (IAA) in comparison with 0.25 NS (33 +/- 14 mg/dL vs 62 +/- 27 mg/dL, respectively). C-peptide was higher in those who received IAA, as were nonprotein calories received on days 4 to 6 of the study (51 +/- 11 kcal/kg/day vs 44 +/- 12 kcal/kg/day, IAA vs 0.25 NS, respectively). CONCLUSIONS: Lower PFH levels in IAA versus 0.25 NS group were consistent with our hypothesis of decreased hemolysis with an isotonic infusion/flush regimen. IAA use may also allow greater early glucose nutrition, as indicated by the higher level of endogenous insulin production and improved glucose tolerance. IAA seems to be a superior UAC solution to 0.25 NS in that it is associated with less hemolysis and improved nutrition. PMID- 15286221 TI - A national assessment of children with special health care needs: prevalence of special needs and use of health care services among children in the military health system. AB - OBJECTIVE: Children are frequently perceived to be healthy, low-risk individuals with a majority of clinical services devoted to health maintenance and preventive clinical services. However, a subset of children have unique needs that require specialized care to achieve optimal health outcomes. The purpose of this research was to use survey tools that have been developed to identify children with special health care needs (CSHCN) to measure prevalence and resource needs of these children in the military health system (MHS). METHODS: The US Department of Defense manages the MHS, which is one of the largest integrated health care systems in the world and provides care to almost 2,000000 children. We incorporated the CSHCN survey screener and assessment questions into the annual health care survey of beneficiaries who are eligible for benefits within the MHS. In addition, we used claims information available from inpatient and outpatient services. We used parent reports from the survey to estimate the prevalence of CSHCN. Incorporating claims data and restricting our analyses to those who were enrolled continuously in a military health maintenance organization (TRICARE Prime), we described utilization of different types of health care resources and compared CSHCN with their healthy counterparts. Finally, we examined alternative types of special needs and performed regression analyses to identify the major determinants of health needs and resource utilization to guide system management and policy development. RESULTS: CSHCN compose 23% of the TRICARE Prime enrollees who are younger than 18 years and whose parents responded to the survey. The needs of a majority of these children consist of prescription medications and services targeting medical, mental health, and educational needs. CSHCN experience 5 times as many admissions and 10 times as many days in hospitals compared with children without special needs. CSHCN are responsible for nearly half of outpatient visits for enrolled children and more than three quarters of inpatient days. Service utilization varies dramatically by type of special need and other demographic variables. CONCLUSION: CSHCN represent a major challenge to organized systems of care and our society. Because they represent a group of children who are particularly at risk with potential for improved health outcomes, efforts to improve quality, coordinate care, and optimize efficiency should focus on this target population. PMID- 15286222 TI - Insurance for children with special health care needs: patterns of coverage and burden on families to provide adequate insurance. AB - OBJECTIVE: To update national estimates of insurance coverage for children with special health care needs (CSHCN) to reflect better the current economic and policy environment and to examine the burden on families and adequacy of coverage. METHODS: I analyzed data on children who were aged 0 to 17 and included in the sample child files of the 2000 and 2001 National Health Interview Survey (NHIS). CSHCN were identified using a noncategorical approach. Various measures of insurance coverage type, premium contributions, unmet need for care, and out of-pocket spending were compared for CSHCN and children without special needs across all incomes and stratified by poverty status. RESULTS: Compared with other children, CSHCN had higher rates of public insurance (29.8% vs 18.5%), lower rates of private insurance (62.5% vs 69.1%), and a smaller percentage without insurance (8.1% vs 11.5%). More than 13% of low-income CSHCN were uninsured. Most (78.1%) families of CSHCN contributed to private insurance premiums. Family premium contributions for employer-sponsored insurance plans averaged 2058 dollars, or 4.4% of income; premiums for private nongroup insurance were higher (3593 dollars) and consumed a larger percentage of income (6.6%). For children with insurance, rates of unmet need for specific services were relatively low, suggesting that insurance coverage was adequate. However, almost 20% of low income CSHCN experienced some form of unmet need and of out-of-pocket spending was significantly higher for families with CSHCN compared with those without CSHCN. CONCLUSIONS: CSHCN are more likely to have insurance coverage, but among low-income CSHCN, lack of insurance remains a problem. In addition, the burden on families of CSHCN to provide insurance is greater, yet coverage purchased is not always adequate to meet the needs of many children and places addition burdens on families to pay directly for care. PMID- 15286223 TI - Bias in reported neurodevelopmental outcomes among extremely low birth weight survivors. AB - OBJECTIVES: The purpose of this study was to investigate possible bias in the evaluation of neurodevelopment and somatic growth at 18 to 22 months' postmenstrual age among extremely low birth weight (ELBW) survivors (401-1000 g at birth). METHODS: Data from a cohort of 1483 ELBW infant survivors who were born January 1993 through December 1994 and cared for at centers in the Neonatal Research Network of the National Institute of Child Health and Human Development were examined retrospectively. Children who were compliant with an 18- to 22 month follow-up visit, who visited but were not measured, or who made no visit were compared regarding 4 outcomes: 1) Bayley Scales of Infant Development, 2nd edition, Mental Developmental Index (MDI) <70 and 2) Psychomotor Developmental Index (PDI) <70, 3) presence or absence of cerebral palsy, and 4) weight <10th percentile for age. Logistic regression models were used to predict likelihood of these outcomes for children with no follow-up evaluation, and predicted probability distributions were compared across the groups. RESULTS: Compared with children who were lost to follow-up, those who were compliant with follow-up were more likely to have been 1 of a multiple birth, to have received postnatal glucocorticoids, and to have had chronic lung disease. These factors were significantly associated with MDI and PDI <70 in the compliant group. Chronic lung disease was associated with increased risk of cerebral palsy (CP). MDI and PDI scores <70 were found in 37% and 29% of children who were evaluated at follow up, respectively. Prediction models revealed that 34% and 26% of infants in the no-visit group would have had MDI and PDI scores <70. Compliant children tended to have greater incidence of MDI <70 compared with those predicted in the no visit group but not PDI <70. CP was identified in 17% of the compliant group and predicted for 18% of the no-visit group. Predicted probabilities of having CP were marginally higher among the no-visit infants compared with those who were compliant with follow-up. There were no statistically significant somatic growth differences among the compliant, visit but not measured, and no-visit groups. CONCLUSION: ELBW infant survivors who weighed 401 to 1000 g at birth and who are compliant with follow-up evaluations may have worse Bayley Scales of Infant Development, 2nd edition, MDI scores than infants with no visit. Thus, follow-up studies based on infants who are compliant with follow-up care may lead to an overestimation of adverse outcomes in ELBW survivors. PMID- 15286224 TI - What follows newborn screening? An evaluation of a residential education program for parents of infants with newly diagnosed cystic fibrosis. AB - OBJECTIVE: The diagnosis of a severe life-limiting condition, such as cystic fibrosis (CF), is generally followed by assessment and treatment of the child and education and counseling for parents. The introduction of newborn screening for CF provides an opportunity for standardized assessment and education. The aim of this study was to evaluate a 5-day residential assessment and education program for parents of infants who receive a diagnosis of CF after newborn screening. METHODS: Eligible parents had a 6- to 30-month-old infant with CF diagnosed by newborn screening. Parents were interviewed by telephone using a structured questionnaire that addressed 3 main themes: 1) initial communication of the diagnosis of CF, 2) the perceived value of the 5-day assessment and education program, and 3) the perceived advantages and disadvantages of the residential component (Care-By-Parent unit) of the program. RESULTS: Fifteen of 17 eligible families took part in the 5-day assessment and education program, 12 of whom used the residential Care-By-Parent unit. At the end of the program, parents believed that they had the knowledge and skills required to manage their child's CF at home. One hundred percent endorsed the timing of the assessment and education program immediately after the child's diagnosis and would recommend it to other families in the same situation. Perceived advantages of the residential program were not having to travel (89%), being able to concentrate on CF (50%), and the benefit of a "home base" at the hospital (39%). Twenty-two percent reported that financial costs related to participation (paternal time off work) were a disadvantage, 17% reported additional strain on family members caring for siblings, and 17% mentioned lack of comfort within the unit. CONCLUSIONS: This time-intensive residential program was evaluated positively by parents of children with newly diagnosed CF. It provides a model for education programs after the diagnosis of CF by newborn screening, as well as for other pediatric conditions that require intensive parent education. PMID- 15286225 TI - Cost-effectiveness of inhaled nitric oxide for the management of persistent pulmonary hypertension of the newborn. AB - OBJECTIVE: Inhaled nitric oxide (iNO) is a selective pulmonary vasodilator that has become part of the standard management for persistent pulmonary hypertension of the newborn (PPHN). This treatment modality, like many in neonatology, has not been well studied using quantitative economic techniques. The objective of this study was to evaluate the economic impact of adding iNO to the treatment protocol of PPHN for term infants from birth to the time of discharge from their initial hospitalization. METHODS: We used decision analysis modeling from a societal perspective to obtain an incremental cost-effectiveness ratio. Outcome probabilities were taken from the medical literature and a cohort of 123 infants who were treated with PPHN at The Children's Hospital of Philadelphia between 1991 and 2002. Costs were estimated from daily resources used by these infants in 2001 dollars. Survival and quality-adjusted life years were used as effectiveness measures. One-way, threshold, and probabilistic sensitivity analyses were performed to assess the robustness of the base-case estimate. RESULTS: The addition of iNO to the treatment regimen of PPHN increased the cost of treating an infant by an average of 1141 dollars, primarily from an increased number of mechanical ventilation days. Use of iNO led to 3.4% more lives saved and a 6% increase in the average utility gained per infant. The incremental cost effectiveness ratio was 33,234 dollars per life saved and 19,022 dollars per quality-adjusted life year gained. The model was robust to changes in outcome probabilities, cost, and utility variables. Only 3.6% of the trials using probabilistic sensitivity analysis found iNO to be more expensive with a worse outcome than conventional therapy alone, whereas 35.7% of the trials found iNO to be cheaper and more effective than conventional treatment alone. CONCLUSIONS: iNO is cost-effective but not cost-saving in treating infants with PPHN from a societal perspective. There are critical time points during an infant's hospitalization that could improve the efficiency and consequently the cost of care for this patient population. PMID- 15286226 TI - Neonatal loss of motor function in human spina bifida aperta. AB - OBJECTIVE: In neonates with spina bifida aperta (SBA), leg movements innervated by spinal segments located caudal to the meningomyelocele are transiently present. This study in neonates with SBA aimed to determine whether the presence of leg movements indicates functional integrity of neuronal innervation and whether these leg movements disappear as a result of dysfunction of upper motor neurons (axons originating cranial to the meningomyelocele) and/or of lower motor neurons (located caudal to the meningomyelocele). METHODS: Leg movements were investigated in neonates with SBA at postnatal day 1 (n = 18) and day 7 (n = 10). Upper and lower motor neuron dysfunction was assessed by neurologic examination (n = 18; disinhibition or inhibition of reflexes, respectively) and by electromyography (n = 12; absence or presence of denervation potentials, respectively). RESULTS: Movements, related to spinal segments caudal to the meningomyelocele, were present in all neonates at postnatal day 1. At day 1, leg movements were associated with signs of both upper (10 of 18) and lower (17 of 18) motor neuron dysfunction caudal to the meningomyelocele. In 7 of 10 neonates restudied after the first postnatal week, leg movements had disappeared. The absence of leg movements coincided with loss of relevant reflexes, which had been present at day 1, indicating progression of lower motor neuron dysfunction. CONCLUSIONS: We conclude that the presence of neonatal leg movements does not indicate integrity of functional lower motor neuron innervation by spinal segments caudal to the meningomyelocele. Present observations could explain why fetal surgery at the level of the meningomyelocele does not prevent loss of leg movements. PMID- 15286227 TI - Normal annual increase of bone mineral density during two years in patients with cystic fibrosis. AB - OBJECTIVES: To determine prospectively for 2 years the change in bone mineral density (BMD) in patients with cystic fibrosis (CF) and to correlate clinical data and routine biochemical parameters of bone metabolism and infection with BMD. METHODS: Fifty-four patients with CF, aged 6 to 33 years, were included. BMD was measured using dual-energy x-ray absorptiometry in lumbar spine (LS) and femoral neck (FN). Anthropometric data and biochemical markers of bone metabolism and infection were measured. The number of intravenous antibiotic courses per year (ivAC) and pulmonary function were assessed. RESULTS: The patients had normal anthropometric data and normal growth, but 36% and 33% of the patients had BMD z score <-1 standard deviation in LS and in FN, respectively. Nevertheless, BMD increased at a normal rate during the 2 years and was correlated to weight and lung function. Intact parathyroid hormone was positively correlated with the increase of BMD in both LS and FN during childhood. Blood sedimentation rate, serum concentration of immunoglobulin G, and ivAC were negatively correlated with BMD in FN. Patients with 2 more severe CF transmembrane conductance regulator mutations had significantly lower BMD in FN than other genetic combinations. CONCLUSION: The study suggests that low BMD in CF is multifactorial and depends on infection and nutritional parameters. Differences in BMD of LS and FN suggested higher susceptibility to infection in FN at all ages. Longitudinal studies starting early before bacterial colonization would be valuable to determine the relative role of infection in the development of BMD in CF. PMID- 15286228 TI - Molecular epidemiology of childhood mitochondrial encephalomyopathies in a Finnish population: sequence analysis of entire mtDNA of 17 children reveals heteroplasmic mutations in tRNAArg, tRNAGlu, and tRNALeu(UUR) genes. AB - OBJECTIVES: Many heteroplasmic point mutations in tRNA genes of mitochondrial DNA (mtDNA) have been associated with human diseases. We recently reported on a prospective 7-year study in which we enrolled 116 consecutive children with undefined encephalomyopathy. Seventeen of them were found to have both a defect in the mitochondrial respiratory chain and abnormal ultrastructure of muscle mitochondria, suggesting a clinically probable mitochondrial encephalopathy. METHODS: We determined the frequency of mtDNA mutations in these 17 children by analyzing the entire sequence of mtDNA by conformation-sensitive gel electrophoresis and sequencing. RESULTS: Three heteroplasmic tRNA mutations that were considered to be pathogenic were detected. Two of the mutations were novel transitions, 10438A>G in the tRNA(Arg) gene and 14696A>G in the tRNA(Glu) gene, whereas the third one was 3243A>G, the common MELAS mutation. The mutant load was very high in the blood and skeletal muscle of the patients and markedly lower in the blood of asymptomatic maternal relatives. The 10438A>G mutation changes the nucleotide flanking the anticodon, whereas 14696A>G changes a nucleotide in the stem of the pseudouridine loop, creating a novel base pair and reducing the wobble. CONCLUSIONS: Our results emphasize that the analysis of the entire sequence of mtDNA is worthwhile in the diagnostic evaluation of patients with clinically probable mitochondrial encephalomyopathy. The frequency of pathogenic mtDNA mutations was found to be 18% among children with biochemically and histologically defined mitochondrial disease, suggesting that the likelihood of nuclear DNA mutations in such a group is several times higher than that of mtDNA mutations. PMID- 15286229 TI - Terminal 22q deletion syndrome: a newly recognized cause of speech and language disability in the autism spectrum. AB - OBJECTIVE: Cryptic subtelomeric chromosome rearrangements account for 6% to 10% of idiopathic mental retardation. As cytogenetic and molecular techniques have become more sophisticated, the number of genetic syndromes attributed to these microdeletions has increased. To date, 64 patients have been described in the literature with a more recently recognized microdeletion syndrome, del 22q13.3. The purpose of this study is to present 11 new cases of this recently described syndrome to delineate further the phenotype and to alert the clinician to another genetic condition that should be considered in the differential diagnosis of early hypotonia, delayed speech acquisition, and autistic behavior. METHODS: Eleven patients were evaluated in 3 academic institutions. Clinical features and results of cytogenetic testing were recorded and tabulated. Reasons for referral for genetic evaluation included developmental delay, severe expressive speech and language delay, and dysmorphic features. RESULTS: Age of presentation ranged from 5 months to 46 years. There were 10 female patients and 1 male patient. All of the patients exhibited delayed motor development, some degree of hypotonia, and severe expressive speech and language delay. Dysmorphic facial features included epicanthal folds, large cupped ears, underdeveloped philtrum, loss of cupid's bow, and full supraorbital ridges. Six patients exhibited autistic-like behaviors. Microscopically visible chromosome deletions were observed in 6 patients. In the remainder, the deletion was detected with the use of fluorescence in situ hybridization. CONCLUSIONS: Hypotonia and developmental delay are nonspecific findings observed in many malformation and genetic syndromes. However, in association with severe speech and language delay and autistic-like behavior, this phenotype may be a significant indication to consider the 22q13 deletion syndrome as a potential cause. PMID- 15286230 TI - Complications associated with image-guided gastrostomy and gastrojejunostomy tubes in children. AB - OBJECTIVE: To evaluate the complications associated with the image-guided insertion of gastrostomy (G) and gastrojejunostomy (GJ) tubes in children, performed by the retrograde percutaneous route. METHODS: A convenience sample of 208 charts of 840 patients recorded as having G and/or GJ tubes placed by the interventional radiology service in a 4-year period (1995-1999) at the Hospital for Sick Children in Toronto, Canada, were selected for review. Complications were categorized as major (including subcutaneous abscess, peritonitis, septicemia, gastrointestinal bleeding, and death) or minor. RESULTS: In total, 253 tubes (208 G tubes, 41 GJ tubes, 4 G and GJ tubes) were placed in the 208 patients reviewed. The median age at the time of insertion was 15 months (range: 7 days-18 years). The most common diagnostic category was neurologic disease (47%). The main indications for tube insertion were recorded as failure to thrive (57%) and risk of aspiration (47%). Major complications were seen in 5% of patients. Peritonitis was noted in 3%, and there was 1 death related to tube insertion (0.4%). Minor complications were found in 73% of patients, including tube dislodgement (37%), tube leakage (25%), and G-tube site skin infection (25%). GJ tubes had a higher rate than G tubes of obstruction, migration, dislodgement, leakage, and intussusception. Site infection, gastroesophageal reflux, and bleeding from the site were seen less frequently in patients with GJ tubes compared with G tubes. CONCLUSION: G and GJ tubes placed by the image guided retrograde percutaneous method are associated with a wide range of complications. The majority of these are minor and are predominantly related to tube maintenance, but major complications, including death, do occur. PMID- 15286231 TI - Gastrointestinal involvement in chronic granulomatous disease. AB - OBJECTIVE: Chronic granulomatous disease (CGD) is a rare disorder of phagocyte oxidative metabolism. In addition to infectious complications, granulomatous lesions often involve hollow viscera, especially the gastrointestinal (GI) tract. The objective of this study was to evaluate the clinical presentation, prevalence, and consequences of GI involvement in patients with CGD. METHODS: The medical records of 140 patients with CGD (67% X-linked) followed at the National Institutes of Health were reviewed and abstracted for GI manifestations. All available GI pathology was reviewed. RESULTS: GI involvement was recorded in 46 (32.8%) of 140 patients with CGD, 89% of whom had X-linked inheritance. The median age at the time of initial GI manifestations was 5 years (range: 0.8-30 years); 70% of the affected patients presented with GI involvement in the first decade of life. Abdominal pain was the most frequent symptom (100%), and hypoalbuminemia was the most frequent sign (70%). Prednisone controlled symptoms and signs in the majority of affected patients, but relapse of symptoms occurred in 71%. GI involvement had no effect on mortality and was unassociated with interferon-gamma use. CONCLUSION: GI involvement is a common and recurring problem in CGD, especially in those with X-linked inheritance. Currently, there is no clear evidence for an infectious cause. The frequency of GI involvement is unaffected by the use of interferon-gamma and does not affect mortality. GI involvement should be sought in patients who have CGD with abdominal pain, growth delay, or hypoalbuminemia. PMID- 15286232 TI - Inhaled corticosteroids and the risk of fractures in children and adolescents. AB - OBJECTIVE: To determine whether children or adolescents who are exposed to inhaled corticosteroids (ie, beclomethasone, budesonide, fluticasone) are at a higher risk of having bone fractures compared with nonexposed individuals. METHODS: We performed a population-based nested case-control analysis using data from the United Kingdom-based General Practice Research Database. Within a base population of 273,456 individuals aged 5 to 79 years, we identified by International Classification of Diseases codes children or adolescents who were aged 5 to 17 years with a fracture diagnosis and up to 6 control subjects per case matched to cases on age, gender, general practice attended, calendar time, and years of history in the GPRD. We compared use of inhaled steroids before the index date between fracture cases and control patients. RESULTS: We identified 3744 cases and 21,757 matched control subjects aged 5 to 17 years. Current exposure to inhaled steroids did not reveal a substantially altered fracture risk compared with nonusers, even in individuals with current longer term exposure (ie, > or =20 prescriptions; adjusted odds ratio 1.15; 95% confidence interval: 0.89-1.48). In individuals with current or previous exposure to oral steroids, the adjusted odds ratio for current long-term inhaled steroid use compared with nonusers was 1.21 (95% confidence interval: 0.99-1.49). CONCLUSIONS: Exposure to inhaled steroids does not materially increase the fracture risk in children or adolescents compared with nonexposed individuals. PMID- 15286233 TI - 2003 Job Lewis Smith award acceptance address: achieving the unfinished agenda through public-private partnerships. PMID- 15286234 TI - The brave new bipartisan world of health care reform: how will low-income families with children with special health care needs fare? PMID- 15286235 TI - The Early Treatment for Retinopathy of Prematurity study: better outcomes, changing strategy. PMID- 15286236 TI - Can we restore aspects of the in utero environment in premature infants to prevent disease? PMID- 15286237 TI - Journeys from childhood to midlife: risk, resilience, and recovery. PMID- 15286238 TI - Pathology research into sudden infant death syndrome: where do we go from here? PMID- 15286239 TI - Closing the gap between guidelines and practice: ensuring safe and healthy beginnings. PMID- 15286240 TI - Evaluation and treatment of the human immunodeficiency virus-1--exposed infant. AB - In developed countries, care and treatment are available for pregnant women and infants that can decrease the rate of perinatal human immunodeficiency virus type 1 (HIV-1) infection to 2% or less. The pediatrician has a key role in prevention of mother-to-child transmission of HIV-1 by identifying HIV-exposed infants whose mothers' HIV infection was not diagnosed before delivery, prescribing antiretroviral prophylaxis for these infants to decrease the risk of acquiring HIV-1 infection, and promoting avoidance of HIV-1 transmission through human milk. In addition, the pediatrician can provide care for HIV-exposed infants by monitoring them for early determination of HIV-1 infection status and for possible short- and long-term toxicities of antiretroviral exposure, providing chemoprophylaxis for Pneumocystis pneumonia, and supporting families living with HIV-1 infection by providing counseling to parents or caregivers. PMID- 15286242 TI - Otitis media meta-analysis. PMID- 15286243 TI - The cost-benefit threshold for low birth weight infants. PMID- 15286244 TI - The review process fails to require appropriate statistical analysis of a group randomized trial. PMID- 15286245 TI - Television viewing and attention deficits in children. PMID- 15286246 TI - Addressing disparity in treatment received. PMID- 15286247 TI - Comparison of alternative diagnostic approaches for managing appendicitis in children: the effect of disease prevalence and spectrum. PMID- 15286248 TI - Italian attention-deficit/hyperactivity disorder registry. PMID- 15286249 TI - Diagnostic accuracy in pediatric appendicitis. PMID- 15286250 TI - Implementation of newborn screening for cystic fibrosis varies widely between states. PMID- 15286251 TI - Identification, evaluation, and management of obesity in an academic primary care center. AB - BACKGROUND: The rapidly increasing prevalence of obesity among children is one of the most challenging dilemmas facing pediatricians today. While the medical community struggles to develop effective strategies for the treatment of this epidemic, timely identification of obesity by pediatric health care providers remains the crucial initial step in the management of obesity. OBJECTIVE: Direct assessment of pediatric clinicians' performance in identifying and managing obesity in clinical practice has not been conducted to date. The objective of this study was to determine rates of identification of obesity by pediatric residents, nurse practitioners, and faculty members in an academic primary care setting and to describe the actions taken by these providers in their evaluation and management of obesity. DESIGN: A retrospective medical record review of all health supervision visits for children 3 months to 16 years of age, examined between December 1, 2001, and February 28, 2002, was performed. For children <5 years of age, a weight >120% of the 50th percentile of weight-for-height was defined as obese. For children > or =5 years of age, a body mass index of >95th percentile for age and gender was defined as obese. SETTING: A large, primary care practice located in a tertiary-care, academic, pediatric hospital, which serves a predominantly urban, minority (70% African American), Medical Assistance insured (90%) population. PARTICIPANTS: Primary care providers, including pediatric residents, nurse practitioners, and faculty physicians. RESULTS: Of the 2515 visits reviewed, a total of 244 patients met the study definition of obesity, yielding an estimated prevalence of obesity visits of 9.7% among health supervision visits for children 3 months to 16 years of age. This prevalence of obesity visits cannot be used to estimate the population prevalence of obesity, given the skewed distribution of visits toward very young children. For all children who met the study definition of obesity, providers documented obesity in their assessments in only 53% of the reviewed visits (129 visits). Although the majority of charts (69%) contained an adequate dietary history, only 15% included a description of the child's activity level or television viewing. Obesity was noted in the physical examination in 39% of cases. For children for whom obesity was identified as a problem by their clinicians (129 patients), 81% of charts contained an adequate dietary history, whereas 27% contained a description of the child's activity level or television viewing. Obesity was noted in the physical examination in 64% of cases. Most children identified as obese by their providers received some management specific to their obesity, including education, screening, and specialist referral. Dietary changes were recommended for 71%, increased activity for 33%, and limitation of television viewing for only 5%. Eighty-three percent of providers recommended close follow-up monitoring. Other recommendations included referral to a dietitian (22%), screening laboratory studies (13%), a food diary (9%), endocrine referral (5%), or preventive cardiology referral (3%). Provider identification of obesity was affected by the age of the patient and by the degree of obesity. Obesity identification was lowest among preschool children (31%) and highest among adolescent patients (76%). Providers evaluating older and heavier children were also more likely to obtain activity histories, note obesity in the physical examination, recommend changes in activity, refer the patient to a nutritionist, obtain screening laboratory studies, and recommend close follow-up monitoring. Identification of obesity and other outcome variables were not significantly influenced by the level of provider training or patient gender. CONCLUSIONS: Although the prevalence of childhood obesity has now reached epidemic proportions, it was under-recognized and under-treated by pediatric primary care providers in our study. Providers identified obesity as a problem for only one-half of the obese children examined for health supervision. The lowest rates of obesity identification occurred among children <5 years of age and those with milder degrees of obesity. Identification did not improve with additional years of pediatric training. Even for the subset of children identified as obese by their providers, evaluation and treatment often were not consistent with current recommendations. For example, more attention was given to the role of diet, compared with activity, in the evaluation of obesity. In particular, only a small number of providers (5%) recommended a decrease in television viewing to their obese patients, despite evidence linking television viewing and pediatric obesity. This finding is of concern, because obesity is known to be a multifactorial disease that responds only to significant changes in both dietary and activity behaviors. Only 13% of providers requested laboratory studies as part of their recommendations. The American Academy of Pediatrics currently recommends obtaining a lipid profile, total cholesterol level, and screening test for type 2 diabetes mellitus as part of the evaluation of obesity. The majority of clinicians who requested laboratory studies included thyroid function tests, which are not recommended by the American Academy of Pediatrics because of the very low likelihood of hypothyroidism as a cause of obesity. Although the extent of evaluation and management for children who were recognized as obese did not meet current guidelines, it was far better than that for patients who were not identified as obese by their providers. This demonstrates the importance of timely identification as the crucial initial step in the management of obesity. The results of this study are disheartening, especially as evidence mounts regarding the importance of early intervention in preventing the medical and psychosocial sequelae of obesity, as well as the persistence of obesity into adulthood. This study highlights the need for increased awareness and identification of obesity in the primary care setting, especially among younger children and those with mild obesity. PMID- 15286252 TI - Clinical outcomes and secondary diagnoses for infants born with hypoplastic left heart syndrome. AB - OBJECTIVE: To explore clinical outcomes and secondary diagnoses present at discharge for infants born with hypoplastic left heart syndrome (HLHS), from a national perspective. METHODS: We examined hospitalizations for infants < or =30 days of age who were born with HLHS, using hospital discharge data from the 1997 Kids Inpatient Database. To explore treatment choices, clinical outcomes, and resource use, we used International Classification of Diseases, 9th Revision, Clinical Modification diagnostic and procedure codes to classify discharges according to type of surgical intervention versus no surgical intervention. To investigate outcomes in more detail, we identified secondary diagnoses noted at discharge, using International Classification of Diseases, 9th Revision, Clinical Modification codes, and stratified results according to type of surgical intervention. RESULTS: Of a total of 550 patients with HLHS, 234 underwent the Norwood procedure, 17 underwent orthotopic heart transplantation, and 106 died in the hospital with no reported surgical intervention. Although we found no demographic variables to be significantly associated with the type of treatment received, discharged patients who died without surgical intervention were significantly more likely to have received care in hospitals identified as small (odds ratio [OR]: 1.5; 95% confidence interval [CI]: 1.03-3.1) or not children's hospitals (OR: 2.02; 95% CI: 1.13-3.6). Secondary diagnoses of cardiac arrest (OR: 2.0; 95% CI: 1.1-3.4) and seizures (OR: 2.6; 95% CI: 1.2-5.5) occurred more frequently in orthotopic heart transplantation cases than in Norwood procedure cases. CONCLUSIONS: These data from a national perspective reflect outcomes of infants with HLHS during a time when rates of initial survival after surgical intervention were considered to be improved. These findings may be useful to clinicians when they are considering and recommending initial medical and surgical strategies currently being proposed for the treatment of HLHS. PMID- 15286253 TI - A modified screening tool for autism (Checklist for Autism in Toddlers [CHAT-23]) for Chinese children. AB - BACKGROUND: There is a recent trend of a worldwide increase in the incidence of autistic spectrum disorder. Early identification and intervention have proved to be beneficial. The original version of the Checklist for Autism in Toddlers (CHAT) was a simple screening tool for identification of autistic children at 18 months of age in the United Kingdom. Children with an absence of joint attention (including protodeclarative pointing and gaze monitoring) and pretend play at 18 months were at high risk of autism. Section A of the CHAT was a self-administered questionnaire for parents, with 9 yes/no questions addressing the following areas of child development: rough and tumble play, social interest, motor development, social play, pretend play, protoimperative pointing (pointing to ask for something), protodeclarative pointing, functional play, and showing. Section B of the CHAT consisted of 5 items, which were recorded with observation of the children by general practitioners or health visitors. The 5 items addressed the child's eye contact, ability to follow a point (gaze monitoring), pretend (pretend play), produce a point (protodeclarative pointing), and make a tower of blocks. A 6-year follow-up study of >16,000 children screened with the CHAT at 18 months in the United Kingdom showed a sensitivity of only 0.40 and a specificity of 0.98, with a positive predictive value (PPV) of 0.26. Rescreening using the same instrument at 19 months for those who failed the 18-month screening yielded a higher PPV of 0.75. Therefore, children were likely to have autism if they failed the CHAT at 18 months and failed again at 19 months. It was estimated that consistent failure in 3 key questions (ie, protodeclarative pointing, gaze monitoring, and pretend play) at 18 months indicated an 83.3% risk of having autism. Because of the poor sensitivity of the original CHAT for autism, a Modified Checklist for Autism in Toddlers (M-CHAT), consisting of 23 questions, with 9 questions from the original CHAT and an additional 14 questions addressing core symptoms present among young autistic children, was designed in the United States. The original observational part (ie, section B) was omitted. The M-CHAT was designed as a simple, self-administered, parental questionnaire for use during regular pediatric visits. The more questions children failed, the higher their risk of having autism. Two criteria were used to measure the sensitivity and specificity of M-CHAT. Criterion 1 used any 3 of the 23 questions, and criterion 2 used 2 of the 6 best questions that could be used to discriminate autism from other groups. The sensitivity and specificity for criterion 1 were 0.97 and 0.95 and those for criterion 2 were 0.95 and 0.99, respectively. M-CHAT had a better sensitivity than the original CHAT, because children up to 24 months of age were screened, with the aim of identifying those who might regress between 18 and 24 months. The 6 best questions of the M-CHAT addressed areas of social relatedness (interest in other children and imitation), joint attention (protodeclarative pointing and gaze monitoring), bringing objects to show parents, and responses to calling. Joint attention was addressed in the original CHAT, whereas the other areas were addressed only in the M-CHAT. To date, there has been no study of the application of either the original CHAT or the M-CHAT for Chinese populations. OBJECTIVES: CHAT-23 is a new checklist translated into Chinese, combining the M-CHAT (23 questions) with graded scores and section B (observational section) of the CHAT. We aimed to determine whether CHAT-23 could discriminate autism at mental ages of 18 to 24 months for Chinese children and to determine the best combination of questions to identify autism. METHODS: A cross sectional cohort study was performed with 212 children with mental ages of 18 to 24 months. The children were categorized into 2 groups, ie, group 1 (N = 87) (autistic disorder: N = 53; pervasive developmental disorder: N = 33) and group 2 (N = 125) (nonautistic). The checklist included self-ad25) (nonautistic). The checklist included self-administered questionnaires with 23 questions (part A) and direct observations of 5 items by trained investigators (part B). We performed discriminant function analysis to We found that 7 key questions, addressing areas of joint attention, pretend play, social relatedness, and social referencing, were identified as discriminative for autism. For part A, failing any 2 of 7 key questions, ie, question 13 (does your child imitate you? [eg, you make a face; will your child imitate it?]), question 5 (does your child ever pretend, for example, to talk on the phone or take care of dolls, or pretend other things?), question 7 (does your child ever use his/her index finger to point, to indicate interest in something?), question 23 (does your child look at your face to check your reaction when faced with something unfamiliar?), question 9 (does your child ever bring objects over to you [parent] to show you something?), question 15 (if you point at a toy across the room, does your child look at it?), and question 2 (does your child take an interest in other children?), yielded sensitivity of 0.931 and specificity of 0.768. Failing any 6 of all 23 questions produced sensitivity of 0.839 and specificity of 0.848. For part B, failing any 2 of 4 items produced sensitivity of 0.736, specificity of 0.912, and PPV of 0.853. The 4 observational items were as follows: item B1: during the appointment, has the child made eye contact with you? item B2: does the child look across to see what you are pointing at? item B3: does the child pretend to pour out tea, drink it, etc?; item B4: does the child point with his/her index finger at the light? CONCLUSION: We found that integrating the screening questions of the M-CHAT (from the United States) and observational section B of the original CHAT (from the United Kingdom) yielded high sensitivity and specificity in discriminating autism at 18 to 24 months of age for our Chinese cohort. This new screening instrument (CHAT-23) is simple to administer. We found that a 2-stage screening program for autism can offer a cost-effective method for early detection of autism at 18 to 24 months. For CHAT-23, use of both the parental questionnaire and direct observation and use of the criterion of failing any 2 of 7 key questions yielded the highest sensitivity but a relatively lower specificity, whereas use of part B yielded the highest specificity but a lower sensitivity. We recommend identifying the possible positive cases with part A (parental questionnaire) and then proceeding to part B (observation) with trained assessors. The proposed algorithm for screening for autism is as follows. 1) The parents or chief caretakers complete a 23-item questionnaire when their children are 18 to 24 months of age. 2) The parents mail, fax, or hand this 23 item questionnaire to the local child health agency. 3) Clerical staff members check for and score failure, with the criteria of failing any 2 of 7 key questions or failing any 6 of 23 questions; if either criterion is met, then the staff members highlight the medical records of the suspicious cases. 4) Trained child health care professionals observe the children who failed any 2 of 7 key questions or any 6 of 23 questions. These identified patients are observed for 5 minutes for part B of the CHAT-23. 5) Any child who fails any 2 of 4 items requires direct referral to a comprehensive autism evaluation team, for early diagnostic evaluation and early intervention. The high sensitivity and specificity of the criteria observed in our study suggested that CHAT-23 might be used to differentiate children with autism. Additional international collaboration with the use of the CHAT, M-CHAT, and CHAT-23 could provide more prospective epidemiologic data, to establish whether there is a genuine increase in the worldwide incidence of autism. PMID- 15286254 TI - Reduced bone density among children with severe hemophilia. AB - OBJECTIVE: Children with severe hemophilia are at risk for reduced bone mineral density (BMD) because of reduced weight-bearing exercise and hepatitis C infection. Reduced bone density in childhood is a risk factor for osteoporosis in later life. STUDY DESIGN: We performed a cross-sectional survey of bone density among 19 children with severe hemophilia, at the Royal Children's Hospital. Results were correlated with findings of blinded objective evaluations of the joints of the lower limb and with hepatitis C status. RESULTS: The mean lumbar bone mineral apparent density for patients was reduced (0.102 g/cm3), compared with that for control subjects (0.113 g/cm3). The mean areal BMD z score was 0.92, which was significantly reduced, compared with that for control subjects. The difference in bone density was independent of body size. There was a statistically significant relationship between the lumbar BMD z scores and the maximal single joint evaluation scores, but there was no difference based on hepatitis C status. CONCLUSIONS: Our results suggest that children with severe hemophilia have reduced BMD. Patients at risk are those with signs of hemophilic arthropathy. Because osteoporosis may complicate the future treatment of patients with hemophilia, screening of young patients for reduced bone density is recommended. PMID- 15286255 TI - The health of primary caregivers of children with cerebral palsy: how does it compare with that of other Canadian caregivers? AB - BACKGROUND: Caring for any child involves considerable resources, but the demands for these resources are often increased when caring for a child with a disability. These demands have implications for the psychologic and physical health of the caregiver (CG). Although a number of recent trends in health care stress the importance of studying and promoting the health of CGs of children with disabilities, the literature in this area exhibits 2 major weaknesses, ie, most studies draw conclusions from relatively small, potentially biased, clinic based samples and the majority of work has focused on the psychologic health of CGs, whereas little research has been undertaken to study their physical well being. The goal of this study was to compare the physical and psychologic health of CGs of children with cerebral palsy (CP) with that of the general population of CGs. METHODS: Data on the physical and psychologic health of 468 primary CGs of children with CP, drawn from 18 of 19 publicly funded children's rehabilitation centers in Ontario, Canada, were collected with a self-completed questionnaire and a face-to-face interview. Identical items and scales had been administered previously to nationally representative samples of the Canadian population in 2 large-scale Canadian surveys, ie, the National Population Health Survey (NPHS) and the National Longitudinal Study of Children and Youth (NLSCY). Subsamples of those data, restricted to adult residents of the province of Ontario who were parents, allowed a comparison of our sample of CGs of children with CP with parent samples from both the NLSCY (n = 2414) and the NPHS (n = 5549). OUTCOME MEASURES: Demographic variables included CG age, gender, education, income, and work-related variables. Psychologic health and support variables included social support, family functioning, frequency of contacts, distress, and emotional and cognitive problems. Physical health variables included the number and variety of chronic conditions, vision, hearing, and mobility problems, and experience of pain. RESULTS: CGs of children with CP had lower incomes than did the general population of CGs (proportion with income over 60,000 dollars: CG: 40.9%; NLSCY: 51.4%), despite the absence of any important differences in education between the 2 samples. Results showed that CGs of children with CP were less likely to report working for pay (CG: 66%; NLSCY: 81.2%), less likely to be engaged in full-time work (CG: 67.5%; NLSCY: 73.2%), and more likely to list caring for their families as their main activity (CG: 37.2%; NLSCY: 28.4%). Measures of support showed no difference in reported social support (CG: mean score: 14.5; SD: 3.4; NLSCY: mean score: 14.3; SD: 2.7) or family functioning (CG: mean score: 8.6; SD: 5.6; NLSCY: mean score: 9.0; SD: 4.9) between the 2 samples, although the CG sample did report a statistically greater number of support contacts (CG: mean score: 4.5; SD: 0.7; NPHS: mean score: 4.2; SD: 0.9). Measures of psychologic health showed greater reported distress (CG: mean score: 4.7; SD: 4.4; NPHS: mean score: 2.2; SD: 2.7), chronicity of distress (CG: mean score: 5.5; SD: 1.4; NPHS: mean score: 5.2; SD: 1.1), emotional problems (CG: 25.3% indicating problems; NPHS: 13.7%), and cognitive problems (CG: 38.8%; NPHS: 14.3%) among CGs of children with CP. They also reported a greater likelihood of a variety of physical problems, including back problems (CG: 35.5% reporting the condition; SE: 2.2%; NLSCY: 12.2%; SE: 0.7%), migraine headaches (CG: 24.2%; SE: 2.0%; NLSCY: 11.2%; SE: 0.7%), stomach/intestinal ulcers (CG: 8.4%; SE: 1.3%; NLSCY: 1.7%; SE: 0.3%), asthma (CG: 15.8%; SE: 1.7%; NLSCY: 6.3%; SE: 0.5%), arthritis/rheumatism (CG: 17.3%; SE: 1.8%; NLSCY: 7.3%; SE: 0.5%), and experience of pain (CG: 28.8%; SE: 2.1%; NPHS: 11.0%; SE: 0.5), as well as a greater overall number of chronic physical conditions (CG: 24.1% reporting no chronic conditions; NLSCY: 55.2%). CONCLUSIONS: Although many families cope well despite the added challenges of caring for a child with a disability, our findings suggest that the demands of their children's disabilities can explain differences in the health status of parents and that parents of children with CP are more likely to have a variety of physical and psychologic health problems. Many of these findings are consistent with a stress process model, in which stress from caregiving can directly or indirectly affect a variety of measures of health, although some of the findings (asthma and arthritis) seem to strain this hypothesis. Alternate interpretations of these findings include the possibility that parents who are in regular contact with the health care system may have more opportunities to discuss and receive attention for their own health concerns than do comparison adults or that the greater number of health issues reported by CGs is related to the nature of our study, perhaps leading these parents to focus on their health and well-being in more depth than is usually feasible in a population survey. CGs of children with CP also had lower incomes, despite the absence of any important differences in education. The findings are consistent with the idea that the financial burden of caring for a child with a disability results in part from a reduced availability of these parents to work for pay. IMPLICATIONS FOR SERVICE PROVIDERS: Physicians and other health care professionals should be aware of the important relationship between child disability and CG health. Family-centered policies and services that explicitly consider CG health are likely to benefit the well-being of both CGs and their families. Future work should address the extent to which the family centeredness of services, as experienced by CGs, is associated with better health outcomes for parents and their families. PMID- 15286256 TI - Decreased arousals among healthy infants after short-term sleep deprivation. AB - OBJECTIVE: Sleep deprivation is a risk factor for sudden infant death syndrome (SIDS). Recent changes in normal life routines were more common among SIDS victims, compared with control infants. Sleep deprivation can result from handling conditions or from sleep fragmentation attributable to respiratory or digestive conditions, fever, or airway obstructions during sleep. Compared with matched control infants, future SIDS victims exhibited fewer complete arousals by the end of the night, when most SIDS cases occur. Arousal from sleep could be an important defense against potentially dangerous situations during sleep. Because the arousal thresholds of healthy infants were increased significantly under conditions known to favor SIDS, we evaluated the effects of a brief period of sleep deprivation on sleep and arousal characteristics of healthy infants. DESIGN: Fourteen healthy infants, with a median age of 8 weeks (range: 6-18 weeks), underwent polygraphic recording during a morning nap and an afternoon nap, in a sleep laboratory. The infants were sleep-deprived for 2 hours before being allowed to fall asleep. Sleep deprivation was achieved by keeping the infants awake, with playing, handling, and mild tactile or auditory stimulations, for as long as possible beyond their habitual bedtimes. To avoid any confounding effect attributable to differences in sleep tendencies throughout the day, sleep deprivation was induced before either the morning nap or the afternoon nap. Seven infants were sleep-deprived before the morning nap and 7 before the afternoon nap. The sleep and arousal characteristics of each infant were compared for the non-sleep-deprived condition (normal condition) and the sleep-deprived condition. During each nap, the infants were exposed, during rapid eye movement (REM) sleep, to white noise of increasing intensity, from 50 dB(A) to 100 dB(A), to determine their arousal thresholds. Arousal thresholds were defined on the basis of the lowest auditory stimuli needed to induce arousal. After the induced arousal, the infants were allowed to return to sleep to complete their naps. RESULTS: Sleep deprivation lasted a median of 120 minutes (range: 90-272 min). Most sleep characteristics were similar for the normal and sleep-deprived conditions, including sleep efficiency, time awake, percentages of REM sleep and non-REM sleep, frequency and duration of central apnea and of periodic breathing, duration of obstructive apnea, mean heart rate and variability, and mean breathing rates during REM sleep and non-REM sleep. After sleep deprivation, the duration of the naps increased, whereas there were decreases in the latency of REM sleep and in the density of body movements. More-intense auditory stimuli were needed for arousal when the infants were sleep-deprived, compared with normal nap sleep. Sleep deprivation was associated with a significant increase in the frequency of obstructive sleep apnea episodes, especially during REM sleep. No significant differences were noted when the effects of morning and afternoon sleep deprivation were compared. No correlation was found between the duration of sleep deprivation and either the frequency of obstructive apnea or the changes in arousal thresholds, although the infants who were more sleep-deprived exhibited tendencies toward higher auditory arousal thresholds and shorter REM sleep latencies, compared with less sleep-deprived infants. There were tendencies for a negative correlation between the auditory arousal thresholds and REM sleep latencies and for a positive correlation between the auditory arousal thresholds and the frequencies of obstructive apnea during REM sleep. CONCLUSIONS: Short term sleep deprivation among infants is associated with the development of obstructive sleep apnea and significant increases in arousal thresholds. As already reported, sleep deprivation may induce effects on respiratory control mechanisms, leading to impairment of ventilatory and arousal responses to chemical stimulation and decreases in genioglossal electromyographic activity during REM sleep. These changes in respiratory control mechanisms could contribute to the development of obstructive apnea. The relationship between the development of obstructive apnea and increases in arousal thresholds remains to be evaluated. Adult subjects with obstructive sleep apnea exhibited both sleep fragmentation and increases in arousal thresholds. Conversely, sleep deprivation increased the frequency and severity of obstructive sleep apnea. In this study, the increases in arousal thresholds and the development of obstructive apnea seemed to result from the preceding sleep deprivation. The depressed arousals that follow sleep deprivation have been attributed to central mechanisms, rather than decreases in peripheral sensory organ function. Such mechanisms could include disturbances within the reticular formation of the brainstem, which integrates specific facilitory inputs, such as ascending pathways from auditory receptors, and inhibitory inputs from the cortex. It remains to be determined whether the combination of upper airway obstruction and depressed arousability from sleep contributes to the increased risk of sudden death reported for sleep deprived infants. PMID- 15286257 TI - Body mass index, waist circumference, and clustering of cardiovascular disease risk factors in a biracial sample of children and adolescents. AB - OBJECTIVE: To derive optimal body mass index (BMI) and waist circumference thresholds for children and adolescents, to predict risk factor clustering. DESIGN: Cross-sectional receiver operating characteristic curve analysis. SETTING: The Bogalusa Heart Study, a community-based study of cardiovascular disease risk factors in early life. PARTICIPANTS: A total of 2597 black and white children and adolescents, 5 to 18 years of age, who were examined between 1992 and 1994. MAIN OUTCOME MEASURES: The presence or absence of > or =3 age-adjusted risk factors (low high-density lipoprotein cholesterol level, high low-density lipoprotein cholesterol level, high triglyceride level, high glucose level, high insulin level, and high blood pressure) was predicted from age-adjusted BMI and waist circumference values. RESULTS: The areas under the receiver operating characteristic curves were significantly different from 0.5 for both BMI and waist circumference for all gender/race groups, ranging from 0.73 to 0.82. The optimal BMI thresholds were at the 53rd and 50th percentiles for white and black male subjects, respectively, and at the 57th and 51st percentiles for white and black female subjects, respectively. Similarly, the optimal waist circumference thresholds were at the 56th and 50th percentiles for white and black male subjects, respectively, and at the 57th and 52nd percentiles for white and black female subjects, respectively. The sensitivity and specificity at the thresholds were similar for all gender/race groups, ranging from 67% to 75%. CONCLUSIONS: The use of BMI and waist circumference for the prediction of risk factor clustering among children and adolescents has significant clinical utility. In this sample, race and gender differences in the optimal thresholds were minimal. PMID- 15286258 TI - Pediatric stroke among Hong Kong Chinese subjects. AB - BACKGROUND: The incidence of pediatric stroke was estimated to be 2.5 to 2.7 cases per 100,000 children per year in North America and 13 cases per 100,000 children per year in France. Stroke is among the top 10 causes of death among children in the United States, with the highest incidence in the first 1 year of life. The annual mortality rate was 0.34 deaths per 100,000 person-years, with an average of 244 deaths per year. Interethnic differences have been demonstrated to be important in pediatric stroke. However, most population-based studies on pediatric stroke were from Europe or North America, and there was a lack of data on the incidence of stroke among Chinese or Asian children. Whether the etiologic patterns and risk factors for death and morbidity among Chinese children with stroke were similar to those described for other ethnic groups was unknown. OBJECTIVES: To calculate the incidence of stroke among Chinese children in Hong Kong and to examine the clinical spectrum, causes, patterns, risk factors, and outcomes of pediatric stroke among Chinese subjects. METHODS: The population of Hong Kong was 6.7 million in 2001, and >98% of our population is Chinese in origin. In Hong Kong, public hospitals under the Hospital Authority provide >95% of the hospital service for the region. We identified children (>1 month to <15 years of age) who were admitted and given a discharge diagnosis of stroke from the Clinical Data Analysis and Reporting System, which is a centralized computerized database for all public hospitals. The discharge coding of stroke used codes from the International Classification of Diseases, 9th Revision, Clinical Modification. Only first admissions during the study period were included. We excluded any subsequent admissions by using multiple demographic characteristics of the patients. The incidence of pediatric stroke was estimated as the number of first hospitalizations divided by the person-years at risk. Since 1991, we had been collecting a database on pediatric stroke (ages of 1 month to 16 years) from a single center (the university-affiliated pediatric unit). The clinical presentation, causes, risk factors, and outcomes for those in the Hong Kong Children's Stroke Registry with follow-up data for > or =2 years were analyzed. Data on outcomes, in terms of survival and neurologic deficits, were studied. For survivors, neurologic deficits were defined as short-term if they resolved within 3 months and long-term if they persisted for >3 months. The severity of deficits was defined as mild when function was minimally affected and the patient remained independent in activities of daily living, moderate when the patient required supervision or partial assistance in activities of daily living or when the deficit caused delay in developmental milestones, and severe when the patient required total or near-total care in activities of daily living. Potential risk factors for death and poor neurologic outcomes, including gender, age at the time of stroke, clinical presentation, causes, and neuroimaging findings, were analyzed. RESULTS: Using projections from census data in 2001, the number of children <15 years of age in Hong Kong from 1998 to 2001 was estimated to be 1,104100 to 1,158800, resulting in 4,545300 person-years. During the same period, 94 children with discharge coding of stroke were identified. Therefore, the estimated incidence of pediatric stroke between 1998 and 2001 was 2.1 cases per 100,000 children-years. The average number of new cases treated annually was 4.5 (0-15 cases/year). Fifty children (28 boys and 22 girls; male/female ratio: 1.27:1) were identified in the 11-year period. The mean age at presentation was 5.6 +/- 4.9 years. Thirty-six strokes (72%) were ischemic and 14 (28%) were hemorrhagic. Despite evaluation for possible underlying causes, 12% (6 cases) remained idiopathic. Eighteen patients with ischemic strokes had cerebral thrombosis, whereas 15 had cerebral embolism. We did not observe any case of sinovenous thrombosis. The 36 cases of ischemic stroke were subtyped according to vascular territories. Eleven cases had infarction involving the middle cerebral artery territory; 2 were limited to the cortical region, 3 were limited to subcortical structures such as the basal ganglia or internal capsule or both, and 6 had complete middle cerebral artery involvement, with cortical and subcortical stroke. Involvement of the anterior cerebral artery occurred in 2 cases, with involvement of cerebellar/basilar artery territories in another 2 cases. The remaining 15 cases had multiple sites of infarction. Three patients experienced secondary hemorrhagic transformation after the initial thrombotic event. Of the 14 patients with hemorrhagic strokes, only 1 had subarachnoid hemorrhage. All others had intracerebral bleeding, at single (N = 9) or multiple (N = 4) loci. Important causes included complications related to congenital heart diseases (N = 15, 30%), vascular diseases (N = 13, 26%), and hematologic diseases (N = 14, 28%). Six cases had no determined causes. One case involved mitochondrial encephalopathy with lactic acidosis and stroke-like episodes and constituted the only case with a metabolic cause. For the 7 patients for whom prothrombotic screening was performed, findings were negative. Seizures (52%) and hemiplegia (34%) were the most common presenting features. Other presenting clinical features included headaches (22%), decreased consciousness (30%), visual field defects (12%), dysphasia (10%), and lethargy (8%). Only 1 patient, with moyamoya disease, had a family history of stroke. The median follow-up time was 8.7 years (range: 2-12.4 years). Nine patients (18%) died, 5 with ischemic stroke and 4 with hemorrhagic stroke. Among the 5 cases of death with ischemic stroke, 3 involved hemorrhagic transformation before death. Seven patients (77%) died within 31 days (range: 2-31 days), whereas the other 2 died 6 months and 2.5 years after the episode. Recurrence occurred in 5 cases (10%). Long-term neurologic deficits occurred among 41% of survivors, including mental retardation (N = 11), epilepsy (N = 7), and hemiplegia (N = 10). The functional deficits were classified as severe in 7 cases, moderate in 3 cases, and mild in 7 cases, for patients with long-term neurologic deficits. Decreased levels of consciousness, hematologic causes, and hemorrhagic transformation (applicable only in ischemic stroke) were significant risk factors associated with high mortality rates. For the 41 patients who survived, the only significant risk factor for long-term neurologic deficits was seizures at the initial presentation. Other factors, such as gender, age, other clinical features, stroke type, vascular territory, other causes, and recurrence of stroke, were all insignificant for both death and long term deficits. The 3 risk factors identified for death were analyzed in multivariate logistic regression analyses, with adjustment for the confounding variables, and only decreased levels of consciousness remained significant (odds ratio = 15.6). CONCLUSIONS: The incidence of stroke among Chinese children was slightly lower than that in Europe or North America. The etiologic pattern was different in our cohort, and there was no sickle cell anemia, thrombophilia, or sinovenous thrombosis. Despite these differences, however, mortality and long term neurologic deficit rates were similar. PMID- 15286259 TI - Hyperbilirubinemia among African American, glucose-6-phosphate dehydrogenase deficient neonates. AB - BACKGROUND: Although glucose-6-phosphate dehydrogenase (G-6-PD) deficiency is prevalent in African Americans, their risk of associated neonatal hyperbilirubinemia has not been prospectively studied. OBJECTIVE: To compare hemolysis and the risk of hyperbilirubinemia among African American, G-6-PD deficient neonates (study group) and G-6-PD-normal control subjects. METHODS: Consecutive, healthy, term and near-term, male neonates born to African American mothers comprised the patient cohort. G-6-PD testing was performed with umbilical cord blood samples. Routine management included measurement of the end tidal carbon monoxide level corrected for ambient carbon monoxide level (ETCOc) within 4 hours after delivery (assessment of hemolysis), > or =1 predischarge bilirubin determination, and additional bilirubin testing as clinically indicated. Indications for phototherapy were identical for study patients and control subjects. Neonates were monitored for the first 1 week of life. ETCOc results, the incidence of hyperbilirubinemia (defined as a transcutaneous or plasma total bilirubin concentration of > or =95th percentile for the hour of life), and the need for phototherapy were compared between the G-6-PD-deficient and G-6-PD normal groups. RESULTS: Five hundred male patients were enrolled, of whom 64 (12.8%) were G-6-PD-deficient. ETCOc values (median and interquartile range) were higher among G-6-PD-deficient neonates than among control neonates (2.4 ppm [2.0 2.9 ppm] vs 2.1 ppm [1.7-2.5 ppm]). More G-6-PD-deficient neonates developed hyperbilirubinemia than did control subjects (14 of 64, 21.9%, vs 29 of 436, 6.7%; relative risk: 3.27; 95% confidence interval: 1.83-5.86), whereas 13 (20.3%) met the criteria for phototherapy, compared with 25 control subjects (5.7%) (relative risk: 3.53; 95% confidence interval: 1.91-6.56). No cases of kernicterus were observed. CONCLUSIONS: Within the African American neonatal population, there is a subgroup of G-6-PD-deficient infants with elevated rates of hemolysis, a higher incidence of hyperbilirubinemia, and a greater requirement for phototherapy, compared with G-6-PD-normal control subjects. These newborns should be monitored vigilantly for the development of hyperbilirubinemia. PMID- 15286260 TI - Dating violence and associated sexual risk and pregnancy among adolescent girls in the United States. AB - OBJECTIVES: To assess the annual prevalence of physical violence from dating partners among a representative sample of sexually experienced adolescent girls attending US public and private high schools, as well as sexual risk behaviors and pregnancy among this population. DESIGN, SETTING, PARTICIPANTS, AND MEASURES: Female students (9th through 12th grade) (N = 6864) participating in the 2001 National Youth Risk Behavior Survey completed measures of physical dating violence during the previous year, as well as assessments of health risk behaviors. Annual rates of physical dating violence were estimated for sexually experienced (n = 3085) and inexperienced girls. Multiple logistic regression models were constructed to assess whether physical dating violence in the previous year was associated with sexual health risks and pregnancy, after controlling for effects of potentially confounding demographic features and risk behaviors. RESULTS: Slightly less than 1 of 5 sexually experienced US adolescent girls (17.7%) reported being intentionally physically hurt by a date in the previous year, and approximately 1 of 25 girls (3.7%) who reported no sexual experience reported such violence. Dating violence among sexually experienced adolescent girls was related to increased risks for both sexual risk behaviors (eg, recent multiple sexual partners: odds ratio: 2.0; 95% confidence interval: 1.3-3.1) and pregnancy (odds ratio: 1.8; 95% confidence interval: 1.3-2.4). CONCLUSIONS: Dating violence is prevalent among US adolescent girls, especially those reporting having had sexual intercourse. Adolescent girls intentionally hurt by a date in the previous year are more likely to experience sexual health risks, including those increasing vulnerability to human immunodeficiency virus infection and other sexually transmitted infections, and to have been pregnant. Dating violence should be integrated into sexual health and pregnancy prevention programs, and greater efforts to identify girls experiencing dating violence are needed among those providing care related to adolescent sexual and reproductive health. PMID- 15286261 TI - Association between patterns of maternal substance use and infant birth weight, length, and head circumference. AB - OBJECTIVE: To determine the effects of patterns of drug use during term pregnancy on infant growth parameters at birth. METHODS: Histories of cocaine, opiate, alcohol, tobacco, and marijuana use during the 3-month period before pregnancy and the 3 trimesters of pregnancy were recorded at the infants' 1-month visit. Patterns of use were categorized as consistently high, moderate, or low/none or increasing/decreasing, and effects on growth parameters were analyzed in multivariate linear regression analyses, with adjustment for clinical site, maternal age, prepregnancy weight, multidrug use, and socioeconomic status. RESULTS: A total of 241 cocaine-exposed women and 410 non-cocaine-exposed women participated in the study. In the cocaine-exposed group, 75% used alcohol, 90% used tobacco, and 53% used marijuana; in the non-cocaine-exposed group, 57% used alcohol, 34% used tobacco, and 19% used marijuana. Birth weight, birth length, and head circumference were significantly greater among infants born to women who used no drugs, compared with women with any cocaine, opiate, alcohol, tobacco, or marijuana use, and were greater among infants born to cocaine nonusers, compared with cocaine users. With adjustment for confounders, birth weight was significantly affected by cocaine (deficit of 250 g with consistently low pattern) and tobacco (deficits of 232 g with consistently high pattern, 173 g with consistently moderate pattern, 153 g with decreasing pattern, and 103 g with consistently low pattern). Head size was affected by cocaine (deficit of 0.98 cm with consistently moderate pattern) and tobacco (deficits of 0.72 cm with consistently high pattern and 0.89 cm with consistently moderate pattern). Birth length was affected by tobacco use only (deficits of 0.82 cm with consistently high pattern and 0.98 cm with decreasing use). CONCLUSION: Patterns of tobacco use during pregnancy affect birth weight, length, and head circumference, whereas cocaine affects birth weight and head size, when adjustments are made for confounders, including multidrug use. PMID- 15286262 TI - Natural history of lipid abnormalities and fat redistribution among human immunodeficiency virus-infected children receiving long-term, protease inhibitor containing, highly active antiretroviral therapy regimens. AB - OBJECTIVE: To characterize the type and frequency of biochemical lipid abnormalities and physical changes in body composition associated with the use of protease inhibitor (PI)-containing antiretroviral therapy among human immunodeficiency virus-infected children treated for up to 6 years. METHODS: A retrospective study of human immunodeficiency virus-infected pediatric patients enrolled in research protocols between August 1995 and December 2001 was performed. All patients who had received a PI for > or =2 years as part of their investigational antiretroviral treatment regimens during the study period were eligible. Of the 110 patients identified as having received PI therapy, 94 met the study criteria. RESULTS: Of the 94 patients evaluated, 9 patients (10%) developed fat redistribution as well as dyslipidemia, 49 patients (52%) developed dyslipidemia without associated physical changes, and 36 patients (38%) exhibited no elevation of lipid levels or physical signs of fat redistribution. For all 9 patients with fat redistribution, the onset of the physical changes was closely associated with changes during pubertal development. Fat redistribution was also associated with lower viral loads and higher, more sustained levels of dyslipidemia. The onset of dyslipidemia and fat redistribution peaked between 10 and 15 years of age. CONCLUSION: Among pediatric patients receiving PI therapy, there seems to be an age range in which children are at greater risk of developing hypercholesterolemia and subsequent fat redistribution, suggesting that unidentified physiologic changes associated with puberty may predispose pediatric patients treated with PI therapy to developing lipodystrophy. PMID- 15286263 TI - Polymorphism of tumor necrosis factor-alpha and risk and severity of bronchopulmonary dysplasia among very low birth weight infants. AB - BACKGROUND: Preterm infants with bronchopulmonary dysplasia (BPD) exhibit prolonged elevation of inflammatory indices in their tracheal aspirates. Tumor necrosis factor-alpha (TNF-alpha) is a central mediator of the inflammatory response. The adenine-containing alleles of TNF-alpha-308 and lymphotoxin alpha+250 have been associated with increased levels of TNF-alpha, whereas the adenine allele of TNF-alpha-238 produces lower levels of TNF-alpha after stimulation. High levels of TNF-alpha may promote chronic inflammation by overwhelming counter-regulatory mechanisms and may lead to the development of BPD. Low levels of TNF-alpha may decrease the risk and/or severity of BPD. OBJECTIVE: To determine whether alleles of TNF-alpha play a role in the susceptibility and/or severity of BPD among very low birth weight infants. METHODS: Infants with birth weights of < or =1250 g were included. Genotypic analyses (polymerase chain reaction-restriction fragment length polymorphism assays) were performed with DNA extracted from whole-blood samples. RESULTS: Infants who developed BPD (fraction of inspired oxygen at postconceptional age of 36 weeks of >0.21, n = 51) had a younger gestational age (mean +/- SD: 27 +/- 4 vs 29 +/- 2 weeks) and lower birth weight (853 +/- 184 vs 997 +/- 193 g) than did infants without BPD (n = 69). The genotypic distributions of lymphotoxin alpha+250 and TNF-alpha-308 were comparable among the groups of infants. However, the AA and GA TNF-alpha-238 genotypes were much less likely to occur among infants with BPD than among infants without BPD. The adenine allele of TNF-alpha 238 was absent among infants with severe BPD and occurred significantly less often among infants with moderate or severe BPD, compared with infants with mild BPD. The number of adenine alleles of TNF-alpha-238 was correlated inversely with the severity of BPD (r = -.341). CONCLUSION: The adenine allele of TNF-alpha-238 may reduce the risk and severity of BPD. PMID- 15286264 TI - Procalcitonin: a marker of severity of acute pyelonephritis among children. AB - OBJECTIVE: Febrile urinary tract infection (UTI) is a common problem among children. The diagnosis and management of acute pyelonephritis is a challenge, particularly during infancy. The distinction between acute pyelonephritis and UTI without renal involvement is very important, because renal infection may cause parenchymal scarring and thus requires more aggressive investigation and follow up monitoring. However, this distinction is not easy among children, because common clinical findings and laboratory parameters are nonspecific, especially among young children. In an attempt to differentiate acute pyelonephritis from febrile UTI without renal lesions in a group of 100 children, we measured serum levels of procalcitonin (PCT), a new marker of infection. The objective of the study was to determine the accuracy of PCT measurements, compared with C-reactive protein (CRP) measurements, in diagnosing acute renal involvement during febrile UTI and in predicting subsequent scars, as assessed with 99mTc-dimercaptosuccinic acid (DMSA) scintigraphy. DESIGN: Serum CRP levels, erythrocyte sedimentation rates, leukocyte counts, and PCT levels were measured for 100 children, 1 month to 13 years of age, admitted for suspected febrile UTI (first episode). Renal parenchymal involvement was evaluated with DMSA scintigraphy within 5 days after admission. The DMSA study was repeated 6 months later if the initial results were abnormal. RESULTS: The mean PCT level was significantly higher in acute pyelonephritis than in UTI without renal lesions (4.48 +/- 5.84 ng/mL vs 0.44 +/- 0.30 ng/mL). In these 2 groups, the mean CRP levels were 106 +/- 68.8 mg/L and 36.4 +/- 26 mg/L, mean erythrocyte sedimentation rates were 79.1 +/- 33 mm/hour and 58.5 +/- 33 mm/hour, and leukocyte counts were 18 492 +/- 6839 cells/mm3 and 16 741 +/- 5302 cells/mm3, respectively. For the prediction of acute pyelonephritis, the sensitivity and specificity of PCT measurements were 83.3% and 93.6%, respectively; CRP measurements had a sensitivity of 94.4% but a specificity of only 31.9%. Positive and negative predictive values for prediction of renal involvement with PCT measurements were 93.7% and 83% and those with CRP measurements were 61.4% and 83.3%, respectively. When inflammatory markers were correlated with the severity of the renal lesions, as assessed with DMSA scintigraphy, a highly significant correlation with both PCT and CRP levels was found. However, when the 2 parameters were correlated with renal scarring in follow-up scans, a significant positive association was found only for PCT levels. CONCLUSIONS: Serum PCT levels may be a sensitive and specific measure for early diagnosis of acute pyelonephritis and determination of the severity of renal parenchymal involvement. Therefore, this measurement could be useful for the treatment of children with febrile UTIs, allowing prediction of patients at risk of permanent parenchymal renal lesions. PMID- 15286265 TI - Acute myocardial infarction in a child: possible pathogenic role of patent foramen ovale associated with heritable thrombophilia. AB - We report an 8-year-old girl who presented with clinical features of an acute myocardial infarction. The angiographic appearance of the coronary arteries was normal. A thrombophilic state caused by a homozygote genotype for the prothrombin G20210A mutation was detected, and a patent foramen ovale (PFO) with right-to left shunting after Valsalva maneuver was demonstrated by transesophageal contrast echocardiography. No other embolic source was identified. We suggest that paradoxical embolization through a PFO resulted in a myocardial infarction in this young patient with hereditary thrombophilia. We closed the patient's PFO with a 25-mm PFO occluder. She was anticoagulated with warfarin for 6 months. After 6 months, a contrast echocardiogram showed no evidence of residual atrial shunt. There has been no evident recurrent paradoxical embolization. PMID- 15286266 TI - Neonatal genital herpes simplex virus type 1 infection after Jewish ritual circumcision: modern medicine and religious tradition. AB - OBJECTIVE: Genital neonatal herpes simplex virus type 1 (HSV-1) infection was observed in a series of neonates after traditional Jewish ritual circumcision. The objective of this study was to describe neonate genital HSV-1 infection after ritual circumcision and investigate the association between genital HSV-1 after circumcision and the practice of the traditional circumcision. METHODS: Eight neonates with genital HSV-1 infection after ritual circumcision were identified. RESULTS: The average interval from circumcision to clinical manifestations was 7.25 +/- 2.5 days. In all cases, the traditional circumciser (the mohel) had performed the ancient custom of orally suctioning the blood after cutting the foreskin (oral metzitzah), which is currently practiced by only a minority of mohels. Six infants received intravenous acyclovir therapy. Four infants had recurrent episodes of genital HSV infection, and 1 developed HSV encephalitis with neurologic sequelae. All four mohels tested for HSV antibodies were seropositive. CONCLUSION: Ritual Jewish circumcision that includes metzitzah with direct oral-genital contact carries a serious risk for transmission of HSV from mohels to neonates, which can be complicated by protracted or severe infection. Oral metzitzah after ritual circumcision may be hazardous to the neonate. PMID- 15286267 TI - Pertussis pneumonia, hypoxemia, hyperleukocytosis, and pulmonary hypertension: improvement in oxygenation after a double volume exchange transfusion. AB - A 3-month-old infant of 33 weeks' gestation was hospitalized with pneumonia caused by Bordetella pertussis. Respiratory insufficiency worsened, and on hospital day 3, there was severe pulmonary dysfunction (arterial oxygen pressure/fraction of inspired oxygen ratio: 120), extreme leukocytosis (white blood cell count 104,000/mm3), and severe pulmonary hypertension as assessed by 2 dimensional echocardiogram. A double volume exchange transfusion was performed to reduce the leukocyte mass. Oxygenation began to improve during the exchange and continued to improve over the ensuing 31 hours (arterial oxygen pressure/fraction of inspired oxygen ratio: 280). The white blood cell count fell dramatically after the exchange, and the rate of rise was slower after exchange therapy compared with preexchange. PMID- 15286268 TI - Intrauterine baclofen exposure: a multidisciplinary approach. AB - Maternal use of the antispasmodic baclofen during pregnancy is an uncommon clinical scenario and leads to uncertainty regarding neonatal risks. We present a team-based, peripartum management plan designed for safe monitoring and minimizing the risk of neonatal withdrawal after unusual drug exposure. Incorporating the expertise of neonatology, nursing, pharmacy, neurology, and the lactation service, as well as parental input, this consensus approach was implemented in a case of maternal oral baclofen use with a successful outcome for the infant and family. PMID- 15286269 TI - All that is vesicular is not herpes: incontinentia pigmenti masquerading as herpes simplex virus in a newborn. AB - Incontinentia pigmenti is a multisystem genodermatosis characterized by cutaneous, neurologic, ophthalmologic, and dental abnormalities. The skin lesions associated with the disease progress through 4 stages, the first being erythematous vesicles linearly distributed along the lines of Blaschko. We report a case of an infant who had incontinentia pigmenti and presented with 2 crops of vesicles and was initially thought to have herpes simplex virus. PMID- 15286270 TI - Varicella zoster virus meningitis in a previously immunized child. AB - We are reporting a previously well 5-year-old child with varicella-zoster meningitis who had a history of a previous immunization against varicella. This child also developed a transient sensorineural hearing loss. The child was treated with acyclovir and made a full recovery. PMID- 15286272 TI - Physical abuse of children. PMID- 15286273 TI - Consultation with the specialist: encopresis: assessment and management. PMID- 15286274 TI - Chloramphenicol: a review. PMID- 15286275 TI - Index of suspicion. PMID- 15286276 TI - Question from the clinician: head circumference. PMID- 15286277 TI - The fourth report on the diagnosis, evaluation, and treatment of high blood pressure in children and adolescents. PMID- 15286278 TI - Synopsis book: best articles relevent to pediatric allergy and immunology. PMID- 15286279 TI - Tapasin enhances MHC class I peptide presentation according to peptide half-life. AB - Understanding how peptides are selected for presentation by MHC class I is crucial to vaccination strategies based on cytotoxic T lymphocyte priming. We have studied this selection of the MHC class I peptide repertoire in terms of the presentation of a series of individual peptides with a wide range of binding to MHC class I. This series was expressed as minigenes, and the presentation of each peptide variant was determined with the same MHC class I peptide-specific antibody. In wild-type cells, the hierarchy of presentation followed peptide half life. This hierarchy broke down in cells lacking tapasin but not in cells lacking calreticulin or in cells lacking transporter associated with antigen processing associated ERp57. We demonstrate a key role for tapasin in shaping the MHC class I peptide repertoire, as enhancement of presentation in the presence of tapasin correlated with peptide half-life. PMID- 15286280 TI - Peroxynitrite inhibition of Coxsackievirus infection by prevention of viral RNA entry. AB - Although peroxynitrite is harmful to the host, the beneficial effects of peroxynitrite are less well understood. We explored the role of peroxynitrite in the host immune response to Coxsackievirus infection. Peroxynitrite inhibits viral replication in vitro, in part by inhibiting viral RNA entry into the host cell. Nitrotyrosine, a marker for peroxynitrite production, is colocalized with viral antigens in the hearts of infected mice but not control mice. Nitrotyrosine coprecipitates with the viral polypeptide VP1 as well. Guanidinoethyl disulfide, a scavenger of peroxynitrite, blocks peroxynitrite inhibition of viral replication in vitro and permits an increase in viral replication in vivo. These data suggest that peroxynitrite is an endogenous effector of the immune response to viruses. PMID- 15286282 TI - Competition among fishermen and fish causes the collapse of Barents Sea capelin. AB - The vast majority of the world's fisheries are typically managed within a single species perspective, ignoring the dynamic feedback mechanisms generated by the ecological web of which they are a part. Here we show that the dynamics of the Barents Sea capelin (Mallotus villosus), the world's largest stock of this species, is strongly influenced by both within-system ecological feedback mechanisms and the impact of harvesting. Both overexploitation and predation by herring (Clupea harengus) can cause the population to collapse, whereas predation by cod (Gadus morhua) is demonstrated a delay in the stock's recovery after a collapse. Such collapses, which have occurred twice in 20 years, affect the entire Barents Sea ecosystem, a region that for ages has provided food for all of Europe. PMID- 15286281 TI - Kinetic profiles of p300 occupancy in vivo predict common features of promoter structure and coactivator recruitment. AB - Understanding the language encrypted in the gene regulatory regions of the human genome is a challenging goal for the genomic era. Although customary extrapolations from steady-state mRNA levels have been effective, deciphering these regulatory codes will require additional empirical data sets that more closely reflect the dynamic progression of molecular events responsible for inducible transcription. We describe an approach using chromatin immunoprecipitation to profile the kinetic occupancy of the transcriptional coactivator and histone acetyltransferase p300 at numerous mitogen-induced genes in activated T cells. Comparison of these profiles reveals a class of promoters that share common patterns of inducible expression, p300 recruitment, dependence on selective p300 domains, and sensitivity to histone deacetylase inhibitors. Remarkably, this class also shares an evolutionarily conserved promoter composition and structure that accurately predicts additional human genes with similar functional attributes. This "reverse genomic" approach will have broad application for the genome-wide classification of promoter structure and function. PMID- 15286283 TI - Use of sequence duplication to engineer a ligand-triggered, long-distance molecular switch in T4 lysozyme. AB - We have designed a molecular switch in a T4 lysozyme construct that controls a large-scale translation of a duplicated helix. As shown by crystal structures of the construct with the switch on and off, the conformational change is triggered by the binding of a ligand (guanidinium ion) to a site that in the wild-type protein was occupied by the guanidino head group of an Arg. In the design template, a duplicated helix is flanked by two loop regions of different stabilities. In the "on" state, the N-terminal loop is weakly structured, whereas the C-terminal loop has a well defined conformation that is stabilized by means of nonbonded interactions with the Arg head group. The truncation of the Arg to Ala destabilizes this loop and switches the protein to the "off" state, in which the duplicated helix is translocated approximately 20 A. Guanidinium binding restores the key interactions, restabilizes the C-terminal loop, and restores the "on" state. Thus, the presence of an external ligand, which is unrelated to the catalytic activity of the enzyme, triggers the inserted helix to translate 20 A away from the binding site. The results illustrate a proposed mechanism for protein evolution in which sequence duplication followed by point mutation can lead to the establishment of new function. PMID- 15286284 TI - Resolution of organelle docking and fusion kinetics in a cell-free assay. AB - In vitro assays of compartment mixing have been key tools in the biochemical dissection of organelle docking and fusion. Many such assays measure compartment mixing through the enzymatic modification of reporter proteins. Homotypic fusion of yeast vacuoles is measured with a coupled assay of proteolytic maturation of pro-alkaline phosphatase (pro-ALP). A kinetic lag is observed between the end of docking, marked by the acquisition of resistance to anti-SNARE reagents, and ALP maturation. We therefore asked whether the time taken for pro-ALP maturation adds a kinetic lag to the measured fusion signal. Prb1p promotes ALP maturation; overproduction of Prb1p accelerates ALP activation in detergent lysates but does not alter the measured kinetics of docking or fusion. Thus, the lag between docking and ALP activation reflects a lag between docking and fusion. Many vacuoles in the population undergo multiple rounds of fusion; methods are presented for distinguishing the first round of fusion from ongoing rounds of fusion. A simple kinetic model distinguishes between two rates, the rate of fusion and the rate at which fusion competence is lost, and allows estimation of the number of rounds of fusion completed. PMID- 15286285 TI - Slow wave sleep in crayfish. AB - Clear evidence of sleep in invertebrates is still meager. Defined as a distinct state of reduced activity, arousability, attention, and initiative, it is well established in mammals, birds, reptiles, and teleosts. It is commonly defined by additional electroencephalographic criteria that are only well established in mammals and to some extent in birds. Sleep states similar to those in mammals, except for electrical criteria, seem to occur in some invertebrates, based on behavior and some physiological observations. Currently the most compelling evidence for sleep in invertebrates (evidence that meets most standard criteria for sleep) has been obtained in the fruit fly Drosophila melanogaster. However, in mammals, sleep is also characterized by a brain state different from that at rest but awake. The electrophysiological slow wave criterion for this state is not seen in Drosophila or in honey bees. Here, we show that, in crayfish, a behavioral state with elevated threshold for vibratory stimulation is accompanied by a distinctive form of slow wave electrical activity of the brain, quite different from that during waking rest. Therefore, crayfish can attain a sleep state comparable to that of mammals. PMID- 15286286 TI - Phycobilisome diffusion is required for light-state transitions in cyanobacteria. AB - Phycobilisomes are the major accessory light-harvesting complexes of cyanobacteria and red algae. Studies using fluorescence recovery after photobleaching on cyanobacteria in vivo have shown that the phycobilisomes are mobile complexes that rapidly diffuse on the thylakoid membrane surface. By contrast, the PSII core complexes are completely immobile. This indicates that the association of phycobilisomes with reaction centers must be transient and unstable. Here, we show that when cells of the cyanobacterium Synechococcus sp. PCC7942 are immersed in buffers of high osmotic strength, the diffusion coefficient for the phycobilisomes is greatly decreased. This suggests that the interaction between phycobilisomes and reaction centers becomes much less transient under these conditions. We discuss the possible reasons for this. State transitions are a rapid physiological adaptation mechanism that regulates the way in which absorbed light energy is distributed between PSI and PSII. Immersing cells in high osmotic strength buffers inhibits state transitions by locking cells into whichever state they were in prior to addition of the buffer. The effect on state transitions is induced at the same buffer concentrations as the effect on phycobilisome diffusion. This implies that phycobilisome diffusion is required for state transitions. The main physiological role for phycobilisome mobility may be to allow such flexibility in light harvesting. PMID- 15286287 TI - Characterization of the Arabidopsis lysine-rich arabinogalactan-protein AtAGP17 mutant (rat1) that results in a decreased efficiency of agrobacterium transformation. AB - Arabinogalactan-proteins (AGPs) are a family of complex proteoglycans widely distributed in plants. The Arabidopsis rat1 mutant, previously characterized as resistant to Agrobacterium tumefaciens root transformation, is due to a mutation in the gene for the Lys-rich AGP, AtAGP17. We show that the phenotype of rat1 correlates with down-regulation of AGP17 in the root as a result of a T-DNA insertion into the promoter of AGP17. Complementation of rat1 plants by a floral dip method with either the wild-type AGP17 gene or cDNA can restore the plant to a wild-type phenotype in several independent transformants. Based on changes in PR1 gene expression and a decrease in free salicylic acid levels upon Agrobacterium infection, we suggest mechanisms by which AGP17 allows Agrobacterium rapidly to reduce the systemic acquired resistance response during the infection process. PMID- 15286288 TI - Understanding in vivo benzenoid metabolism in petunia petal tissue. AB - In vivo stable isotope labeling and computer-assisted metabolic flux analysis were used to investigate the metabolic pathways in petunia (Petunia hybrida) cv Mitchell leading from Phe to benzenoid compounds, a process that requires the shortening of the side chain by a C(2) unit. Deuterium-labeled Phe ((2)H(5)-Phe) was supplied to excised petunia petals. The intracellular pools of benzenoid/phenylpropanoid-related compounds (intermediates and end products) as well as volatile end products within the floral bouquet were analyzed for pool sizes and labeling kinetics by gas chromatography-mass spectrometry and liquid chromatography-mass spectrometry. Modeling of the benzenoid network revealed that both the CoA-dependent, beta-oxidative and CoA-independent, non-beta-oxidative pathways contribute to the formation of benzenoid compounds in petunia flowers. The flux through the CoA-independent, non-beta-oxidative pathway with benzaldehyde as a key intermediate was estimated to be about 2 times higher than the flux through the CoA-dependent, beta-oxidative pathway. Modeling of (2)H(5) Phe labeling data predicted that in addition to benzaldehyde, benzylbenzoate is an intermediate between l-Phe and benzoic acid. Benzylbenzoate is the result of benzoylation of benzyl alcohol, for which activity was detected in petunia petals. A cDNA encoding a benzoyl-CoA:benzyl alcohol/phenylethanol benzoyltransferase was isolated from petunia cv Mitchell using a functional genomic approach. Biochemical characterization of a purified recombinant benzoyl CoA:benzyl alcohol/phenylethanol benzoyltransferase protein showed that it can produce benzylbenzoate and phenylethyl benzoate, both present in petunia corollas, with similar catalytic efficiencies. PMID- 15286290 TI - Rapid regulation of the methylerythritol 4-phosphate pathway during isoprene synthesis. AB - More volatile organic carbon is lost from plants as isoprene than any other molecule. This flux of carbon to the atmosphere affects atmospheric chemistry and can serve as a substrate for ozone production in polluted air. Isoprene synthesis may help leaves cope with heatflecks and active oxygen species. Isoprene synthase, an enzyme related to monoterpene synthases, converts dimethylallyl diphosphate derived from the methylerythritol 4-phosphate pathway to isoprene. We used dideuterated deoxyxylulose (DOX-d(2)) to study the regulation of the isoprene biosynthetic pathway. Exogenous DOX-d(2) displaced endogenous sources of carbon for isoprene synthesis without increasing the overall rate of isoprene synthesis. However, at higher concentrations, DOX-d(2) completely suppressed isoprene synthesis from endogenous sources and increased the overall rate of isoprene synthesis. We interpret these results to indicate strong feedback control of deoxyxylulose-5-phosphate synthase. We related the emission of labeled isoprene to the concentration of labeled dimethylallyl diphosphate in order to estimate the in situ K(m) of isoprene synthase. The results confirm that isoprene synthase has a K(m) 10- to 100-fold higher for its allylic diphosphate substrate than related monoterpene synthases for geranyl diphosphate. PMID- 15286291 TI - The spatially variable inhibition by water deficit of maize root growth correlates with altered profiles of proton flux and cell wall pH. AB - Growth of elongating primary roots of maize (Zea mays) seedlings was approximately 50% inhibited after 48 h in aerated nutrient solution under water deficit induced by polyethylene glycol 6000 at -0.5 MPa water potential. Proton flux along the root elongation zone was assayed by high resolution analyses of images of acid diffusion around roots contacted for 5 min with pH indicator gel. Profiles of root segmental elongation correlated qualitatively and quantitatively (r(2) = 0.74) with proton flux along the surface of the elongation zone from water-deficit and control treatments. Proton flux and segmental elongation in roots under water deficit were remarkably well maintained in the region 0 to 3 mm behind the root tip and were inhibited from 3 to 10 mm behind the tip. Associated changes in apoplastic pH inside epidermal cell walls were measured in three defined regions along the root elongation zone by confocal laser scanning microscopy using a ratiometric method. Finally, external acidification of roots was shown to specifically induce a partial reversal of growth inhibition by water deficit in the central region of the elongation zone. These new findings, plus evidence in the literature concerning increases induced by acid pH in wall extensibility parameters, lead us to propose that the apparently adaptive maintenance of growth 0 to 3 mm behind the tip in maize primary roots under water deficit and the associated inhibition of growth further behind the tip are related to spatially variable changes in proton pumping into expanding cell walls. PMID- 15286289 TI - Brassinosteroid deficiency due to truncated steroid 5alpha-reductase causes dwarfism in the lk mutant of pea. AB - The endogenous brassinosteroids in the dwarf mutant lk of pea (Pisum sativum) were quantified by gas chromatography-selected ion monitoring. The levels of castasterone, 6-deoxocastasterone, and 6-deoxotyphasterol in lk shoots were reduced 4-, 70-, and 6-fold, respectively, compared with those of the wild type. The fact that the application of brassinolide restored the growth of the mutant indicated that the dwarf mutant lk is brassinosteroid deficient. Gas chromatography-selected ion monitoring analysis of the endogenous sterols in lk shoots revealed that the levels of campestanol and sitostanol were reduced 160- and 10-fold, respectively, compared with those of wild-type plants. These data, along with metabolic studies, showed that the lk mutant has a defect in the conversion of campest-4-en-3-one to 5alpha-campestan-3-one, which is a key hydrogenation step in the synthesis of campestanol from campesterol. This defect is the same as that found in the Arabidopsis det2 mutant and the Ipomoea nil kbt mutant. The pea gene homologous to the DET2 gene, PsDET2, was cloned, and it was found that the lk mutation would result in a putative truncated PsDET2 protein. Thus it was concluded that the short stature of the lk mutant is due to a defect in the steroidal 5alpha-reductase gene. This defect was also observed in the callus induced from the lk mutant. Biosynthetic pathways involved in the conversion of campesterol to campestanol are discussed in detail. PMID- 15286292 TI - The role of the C4 pathway in carbon accumulation and fixation in a marine diatom. AB - The role of a C(4) pathway in photosynthetic carbon fixation by marine diatoms is presently debated. Previous labeling studies have shown the transfer of photosynthetically fixed carbon through a C(4) pathway and recent genomic data provide evidence for the existence of key enzymes involved in C(4) metabolism. Nonetheless, the importance of the C(4) pathway in photosynthesis has been questioned and this pathway is seen as redundant to the known CO(2) concentrating mechanism of diatoms. Here we show that the inhibition of phosphoenolpyruvate carboxylase (PEPCase) by 3,3-dichloro-2-dihydroxyphosphinoylmethyl-2-propenoate resulted in a more than 90% decrease in whole cell photosynthesis in Thalassiosira weissflogii cells acclimated to low CO(2) (10 microm), but had little effect on photosynthesis in the C(3) marine Chlorophyte, Chlamydomonas sp. In 3,3-dichloro-2-dihydroxyphosphinoylmethyl-2-propenoate-treated T. weissflogii cells, elevated CO(2) (150 microm) or low O(2) (80-180 microm) restored photosynthesis to the control rate linking PEPCase inhibition with CO(2) supply in this diatom. In C(4) organic carbon-inorganic carbon competition experiments, the (12)C-labeled C(4) products of PEPCase, oxaloacetic acid and its reduced form malic acid suppressed the fixation of (14)C-labeled inorganic carbon by 40% to 50%, but had no effect on O(2) evolution in photosynthesizing diatoms. Oxaloacetic acid-dependent O(2) evolution in T. weissflogii was twice as high in cells acclimated to 10 microm rather than 22 microm CO(2), indicating that the use of C(4) compounds for photosynthesis is regulated over the range of CO(2) concentrations observed in marine surface waters. Short-term (14)C uptake (silicone oil centrifugation) and CO(2) release (membrane inlet mass spectrometry) experiments that employed a protein denaturing cell extraction solution containing the PEPCKase inhibitor mercaptopicolinic acid revealed that much of the carbon taken up by diatoms during photosynthesis is stored as organic carbon before being fixed in the Calvin cycle, as expected if the C(4) pathway functions as a CO(2) concentrating mechanism. Together these results demonstrate that the C(4) pathway is important in carbon accumulation and photosynthetic carbon fixation in diatoms at low (atmospheric) CO(2). PMID- 15286293 TI - Phosphorylation of transitory starch is increased during degradation. AB - The starch excess phenotype of Arabidopsis mutants defective in the starch phosphorylating enzyme glucan, water dikinase (EC 2.7.9.4) indicates that phosphorylation of starch is required for its degradation. However, the underlying mechanism has not yet been elucidated. In this study, two in vivo systems have been established that allow the analysis of phosphorylation of transitory starch during both biosynthesis in the light and degradation in darkness. First, a photoautotrophic culture of the unicellular green alga Chlamydomonas reinhardtii was used to monitor the incorporation of exogenously supplied (32)P orthophosphate into starch. Illuminated cells incorporated (32)P into starch with a constant rate during 2 h. By contrast, starch phosphorylation in darkened cells exceeded that in illuminated cells within the first 30 min, but subsequently phosphate incorporation declined. Pulse-chase experiments performed with (32)P/(31)P orthophosphate revealed a high turnover of the starch-bound phosphate esters in darkened cells but no detectable turnover in illuminated cells. Secondly, leaf starch granules were isolated from potato (Solanum tuberosum) plants grown under controlled conditions and glucan chains from the outer granule layer were released by isoamylase. Phosphorylated chains were purified and analyzed using high performance anion-exchange chromatography and matrix-assisted laser desorption/ionization mass spectrometry. Glucans released from the surface of starch granules that had been isolated from darkened leaves possessed a considerably higher degree of phosphorylation than those prepared from leaves harvested during the light period. Thus, in the unicellular alga as well as in potato leaves, net starch degradation is accompanied with an increased phosphorylation of starch. PMID- 15286294 TI - Metabolic discrimination of Catharanthus roseus leaves infected by phytoplasma using 1H-NMR spectroscopy and multivariate data analysis. AB - A comprehensive metabolomic profiling of Catharanthus roseus L. G. Don infected by 10 types of phytoplasmas was carried out using one-dimensional and two dimensional NMR spectroscopy followed by principal component analysis (PCA), an unsupervised clustering method requiring no knowledge of the data set and used to reduce the dimensionality of multivariate data while preserving most of the variance within it. With a combination of these techniques, we were able to identify those metabolites that were present in different levels in phytoplasma infected C. roseus leaves than in healthy ones. The infection by phytoplasma in C. roseus leaves causes an increase of metabolites related to the biosynthetic pathways of phenylpropanoids or terpenoid indole alkaloids: chlorogenic acid, loganic acid, secologanin, and vindoline. Furthermore, higher abundance of Glc, Glu, polyphenols, succinic acid, and Suc were detected in the phytoplasma infected leaves. The PCA of the (1)H-NMR signals of healthy and phytoplasma infected C. roseus leaves shows that these metabolites are major discriminating factors to characterize the phytoplasma-infected C. roseus leaves from healthy ones. Based on the NMR and PCA analysis, it might be suggested that the biosynthetic pathway of terpenoid indole alkaloids, together with that of phenylpropanoids, is stimulated by the infection of phytoplasma. PMID- 15286295 TI - Roles for Class III HD-Zip and KANADI genes in Arabidopsis root development. AB - Meristems within the plant body differ in their structure and the patterns and identities of organs they produce. Despite these differences, it is becoming apparent that shoot and root apical and vascular meristems share significant gene expression patterns. Class III HD-Zip genes are required for the formation of a functional shoot apical meristem. In addition, Class III HD-Zip and KANADI genes function in patterning lateral organs and vascular bundles produced from the shoot apical and vascular meristems, respectively. We utilize both gain- and loss of-function mutants and gene expression patterns to analyze the function of Class III HD-Zip and KANADI genes in Arabidopsis roots. Here we show that both Class III HD-Zip and KANADI genes play roles in the ontogeny of lateral roots and suggest that Class III HD-Zip gene activity is required for meristematic activity in the pericycle analogous to its requirement in the shoot apical meristem. PMID- 15286296 TI - 13C labeling reveals chloroplastic and extrachloroplastic pools of dimethylallyl pyrophosphate and their contribution to isoprene formation. AB - Isoprene emitted from plants is made in chloroplasts from dimethylallyl pyrophosphate (DMAPP). Leaves of Populus nigra and Phragmites australis exposed to (13)CO(2) for 15 min emitted isoprene that was about 90% (13)C, but DMAPP isolated from those leaves was only 28% and 36% (13)C, respectively. The labeled DMAPP is likely to represent chloroplastic DMAPP contributing to isoprene emission. A substantial (13)C labeling was also found in both emission and DMAPP pool of low-emitting, young leaves of Phragmites. This confirms that low emission of young leaves is not caused by absence of chloroplastic DMAPP but rather by enzyme characteristics. A very low (13)C labeling was found in the DMAPP pool and in the residual isoprene emission of leaves previously fed with fosmidomycin to inhibit isoprene formation. This shows that fosmidomycin is a very effective inhibitor of the chloroplastic biosynthetic pathway of isoprene synthesis, that the residual isoprene is formed from extra-chloroplastic sources, and that chloroplastic and extrachloroplastic pathways are not cross-linked, at least following inhibition of the chloroplastic pathway. Refixation of unlabeled respiratory CO(2) in the light may explain incomplete labeling of isoprene emission, as we found a good association between these two parameters. PMID- 15286297 TI - A dominant mutation in the pea PHYA gene confers enhanced responses to light and impairs the light-dependent degradation of phytochrome A. AB - Phytochrome A (phyA) is an important photoreceptor controlling many processes throughout the plant life cycle. It is unique within the phytochrome family for its ability to mediate photomorphogenic responses to continuous far-red light and for the strong photocontrol of its transcript level and protein stability. Here we describe a dominant mutant of garden pea (Pisum sativum) that displays dramatically enhanced responses to light, early photoperiod-independent flowering, and impaired photodestruction of phyA. The mutant carries a single base substitution in the PHYA gene that is genetically inseparable from the mutant phenotype. This substitution is predicted to direct the replacement of a conserved Ala in an N-terminal region of PHYA that is highly divergent between phyA and other phytochromes. This result identifies a region of the phyA photoreceptor molecule that may play an important role in its fate after photoconversion. PMID- 15286298 TI - Why use remote guidance to steer catheters and guide wires? PMID- 15286302 TI - Chronic pulmonary embolism: combining MR angiography with functional assessment. PMID- 15286303 TI - Building a team for change in an academic radiology department. PMID- 15286305 TI - Rectal cancer: review with emphasis on MR imaging. AB - One concern after rectal cancer surgery is the high local recurrence rate. Randomized trials have shown that the best local control rate for rectal cancer patients as a group is achieved after a short course of radiation therapy followed by optimal surgery. It is debatable, however, whether all patients with rectal cancer should undergo preoperative radiation therapy. Preoperative identification of those most likely to benefit from neoadjuvant therapy is important. Therefore, the challenge for preoperative imaging in rectal cancer is to determine subgroups of patients with different risks for recurrence: those with superficial tumors, who can be treated with surgery alone; those with operable tumors and a wide circumferential resection margin, who can be treated with a short course of radiation therapy followed by total mesorectal excision; and those with advanced cancer and a close or involved resection margin, who require a long course of radiation therapy, with or without chemotherapy, and extensive surgery. So far, there is no consensus on the role of diagnostic imaging (endorectal ultrasonography, computed tomography, and magnetic resonance [MR] imaging) in the care of patients with primary rectal cancer. Preoperative staging has long relied on digital examination alone, which indicates that it has been difficult to achieve accuracy levels high enough for clinical decision making with preoperative imaging. In this review, the relevance of preoperative imaging in staging the local extent of primary rectal cancer will be discussed. Research on various imaging modalities, with an emphasis on MR, will be discussed under four main headings that address the most relevant aspects of local spread of rectal tumors: T stage, circumferential resection margin, locally advanced rectal cancer, and N stage. PMID- 15286306 TI - Comparison of Z-axis automatic tube current modulation technique with fixed tube current CT scanning of abdomen and pelvis. AB - PURPOSE: To compare image quality, diagnostic acceptability, and radiation exposure associated with 16-section multi-detector row computed tomographic (CT) examinations of abdomen and pelvis performed with z-axis modulation technique of automatic tube current modulation and with manual selection of fixed tube current. MATERIALS AND METHODS: Sixty-two consecutive subjects (mean age, 60 years; age range, 19-84 years; male-to-female ratio, 35:27) underwent follow-up CT of abdomen and pelvis with use of a 16-section multi-detector row scanner and z-axis modulation technique (10.5-12.0-HU noise index, 10-380 mA). Scanning parameters included 140 kVp, 0.5-1.0-second gantry rotation time, 0.938:1 beam pitch, and 5-mm reconstructed section thickness. For each subject, images obtained with z-axis modulation were compared with previous images obtained with fixed tube current (200-300 mA) and with other parameters identical. Images were compared for noise and diagnostic acceptability by two subspecialty radiologists using a five-point scale (1, unacceptable; 3, acceptable; 5, excellent) at five levels: upper liver at diaphragm, porta hepatis, right kidney hilum, iliac crest, and upper margin of acetabulum. Tube current and gantry rotation time used for acquisitions at these levels were recorded. Data were analyzed with parametric and nonparametric statistical tests. RESULTS: Although no significant differences were found (P =.34), images acquired with z-axis modulation at the levels of the upper liver (diaphragm) and acetabulum had a higher noise and lower diagnostic quality, compared with images acquired with fixed tube current. Compared with fixed tube current, z-axis modulation resulted in tube current-time product reduction in 54 (87%) of 62 examinations (mean reduction, 71.2 mAs) and increase in eight (13%) (mean increase, 17.0 mAs). CONCLUSION: Compared with manually selected fixed tube current, z-axis automatic tube current modulation resulted in reduced tube current-time product and similar image noise and diagnostic acceptability at CT of abdomen and pelvis. PMID- 15286307 TI - Laparoscopic splenectomy: multi-detector row CT for preoperative evaluation. AB - PURPOSE: To prospectively evaluate multi-detector row spiral computed tomography (CT) for determination of splenic volume, splenic vascular anatomy, and presence of accessory spleens and parenchymal lesions in patients who were undergoing laparoscopic splenectomy. MATERIALS AND METHODS: Twenty-two patients who were candidates for laparoscopic splenectomy underwent multiphasic multi-detector row CT. Two observers evaluated splenic volume with two hand-tracing editing modalities. Variability between the two observers was calculated with a reliability coefficient (Cronbach alpha). A linear regression equation for each modality was generated to identify the correlation between the two observers. Multi-detector row CT angiography was evaluated for assessment of splenic vascular anatomy. Presence and number of both accessory spleens and parenchymal lesions were recorded. RESULTS: Mean splenic volume was 1,050 and 1,046 mL, respectively, for observers A and B by using each-section editing (technique 1) and 1,067 and 1,068 mL for observers A and B by using distanced editing (technique 2). For each editing modality, alpha reliability coefficient was higher than 0.99. Both techniques 1 and 2 were very highly predictive of specimen weight and had R2 values of greater than 0.99 (P <.001). CT angiograms correctly showed polar arteries in all cases and the presence of the arteria pancreatica magna in one case. Multi-detector row CT demonstrated the presence, number, and size of all accessory spleens and of focal parenchymal lesions. CONCLUSION: Multi detector row CT volumetric and anatomic evaluation provided accurate and reproducible information. PMID- 15286308 TI - Case 77. PMID- 15286309 TI - Case 73: Nasolacrimal duct mucocele. PMID- 15286310 TI - Fetal anomalies: comparison of MR imaging and US for diagnosis. AB - PURPOSE: To compare prenatal ultrasonography (US) and magnetic resonance (MR) imaging for the diagnosis of fetal anomalies. MATERIALS AND METHODS: Images of 27 fetuses (28 diagnostic cases) with anomalies diagnosed at US were evaluated; in these fetuses, prenatal MR imaging was performed within 15 days of US. Prenatal US and MR imaging findings were compared with postnatal diagnoses. Postnatal evaluation included US, MR imaging, autopsy, surgery, voiding cystourethrography, computed tomography, angiography, and physical examination. RESULTS: In seven diagnostic cases, US and MR imaging findings were in complete agreement with postnatal diagnoses. MR imaging correctly provided additional information to the US-determined diagnosis in another seven and correctly changed the US diagnosis in three. The MR imaging-determined diagnosis was incorrect and the US diagnosis was correct in four cases. In seven cases, the diagnoses at both US and MR imaging were incorrect when correlated with the postnatal outcome. MR imaging was most valuable in the assessment of anomalies of the central nervous system. CONCLUSION: MR imaging may have a place as an adjunct to US in evaluation of fetal anomalies, particularly those involving the central nervous system. PMID- 15286311 TI - Enhancing research in academic radiology departments: recommendations of the 2003 Consensus Conference. AB - Opportunities for funded radiologic research are greater than ever, and the amount of federal funding coming to academic radiology departments is increasing. Even so, many medical school-based radiology departments have little or no research funding. Accordingly, a consensus panel was convened to discuss ways to enhance research productivity and broaden the base of research strength in as many academic radiology departments as possible. The consensus panel included radiologists who have leadership roles in some of the best-funded research departments, radiologists who direct other funded research programs, and radiologists with related expertise. The goals of the consensus panel were to identify the attributes associated with successful research programs and to develop an action plan for radiology research based on these characteristics. PMID- 15286312 TI - Radiation from "extra" images acquired with abdominal and/or pelvic CT: effect of automatic tube current modulation. AB - PURPOSE: To retrospectively determine the number and usefulness of images acquired beyond the intended anatomic area of interest with abdominal and/or pelvic computed tomography (CT) and to assess the effect of automatic tube current modulation (ATCM) on associated radiation. MATERIALS AND METHODS: Superior and inferior levels at routine abdominal and/or pelvic CT were defined as the dome of the diaphragm and the inferior margin of the pubic symphysis, respectively. Records of 106 consecutive examinations (male-to-female ratio, 45:61; age range, 21-86 years) performed from June 1 to June 30, 2003, were reviewed to determine the number of "extra" images. Sixty-two abdominal and/or pelvic CT examinations performed concurrently with chest or thigh CT or for trauma were not included in the 106. Abdominal and/or pelvic CT was performed with either ATCM (n = 44) or manual selection of tube current (n = 62). CT parameters recorded for each extra image included tube current, peak kilovoltage, and gantry rotation time. Mean and median tube current-time products were calculated for extra images. Extra images were analyzed for pathologic findings. Statistical analysis was performed with the Student t test. RESULTS: Extra images were acquired above the dome of the diaphragm in 103 (97%) of 106 examinations and below the pubic symphysis in 100 (94%) of 106. A total of 1,280 extra images were acquired in 106 examinations (mean, 12 images per examination). Nineteen additional findings were observed on extra images. With ATCM, mean tube current time product was 74.5 and 120.6 mAs for extra images acquired above the diaphragm and below the pubic symphysis, respectively; with manual selection, mean tube current-time products were 167.5 and 168.3 mAs (P <.05). CONCLUSION: Most extra images acquired at abdominal and/or pelvic CT contributed no additional information. With ATCM, the radiation dose was reduced by a mean of 56% (median, 72%) for extra images above the diaphragm and 29% (median, 36%) for images below the pubic symphysis, compared with dose levels with manual selection. PMID- 15286313 TI - Implementation of an international teleradiology staffing model. AB - Although teleradiology is presently being used extensively in the United States for both overseas subspecialty consultations and overnight coverage of imaging services at domestic medical centers-there has been limited investigation of its potential to help provide staffing support to U.S. medical centers from offshore locations. In this review, the authors describe an empirical assessment of the clinical feasibility and applicability of body computed tomographic (CT) image cases that originated at a U.S. university hospital being interpreted nearly contemporaneously by a staff radiologist in India. During a 3-month period, nonemergent CT cases obtained at a tertiary care institution (Yale-New Haven Hospital) were transmitted daily to a satellite reading facility in Bangalore, India. The cases were interpreted at the satellite reading facility by a faculty member radiologist who maintained hospital privileges and academic appointment at the parent institution in the United States. CT imaging reports were transcribed and uploaded directly to the parent institution's radiology information system. Technical problems temporarily prevented the transfer of image cases twice during the study period. Overall, the project results demonstrated the feasibility and reliability of an international teleradiology staffing model. PMID- 15286314 TI - Characterization of focal liver lesions with contrast-specific US modes and a sulfur hexafluoride-filled microbubble contrast agent: diagnostic performance and confidence. AB - PURPOSE: To assess whether characterization of solid focal liver lesions could be improved by using ultrasonographic (US) contrast-specific modes after sulfur hexafluoride-filled microbubble contrast agent injection, as compared with lesion characterization achieved with preliminary baseline US. MATERIALS AND METHODS: Four hundred fifty-two solid focal hepatic lesions that were considered indeterminate at baseline gray-scale and color Doppler US were examined after microbubble contrast agent injection performed by using low-acoustic-power contrast-specific modes during the arterial (10-40 seconds after injection), portal venous (50-90 seconds after injection), and late (100-300 seconds after injection) phases. Two readers independently and retrospectively reviewed baseline and contrast material-enhanced US scans and classified each depicted lesion as malignant or benign according to standard diagnostic criteria. Sensitivity, specificity, accuracy, and positive and negative predictive values and areas under the receiver operating characteristic curve (Az) were calculated by considering histologic analysis (317 patients) or contrast-enhanced helical computed tomography followed by serial US 3-6 months apart (135 patients) as the reference standards. RESULTS: Different contrast enhancement patterns were observed according to lesion characteristics. During the late phase, benign lesions were predominantly hyper- or isoechoic relative to the adjacent liver parenchyma, whereas malignant lesions were predominantly hypoechoic. Review of the contrast-enhanced US scans after baseline image review yielded significantly improved diagnostic performance (P <.05). Overall diagnostic accuracy was 49% before versus 85% after review of the contrast-enhanced scan for reader 1 and 51% before versus 88% after review of the contrast-enhanced scan for reader 2. Diagnostic confidence-that is, the Az-was 0.820 before versus 0.968 after review of the contrast-enhanced scan for reader 1 and 0.831 before versus 0.978 after review of the contrast-enhanced scan for reader 2. CONCLUSION: The use of contrast-specific modes with a sulfur hexafluoride contrast agent led to improved characterization of solid focal liver lesions. PMID- 15286315 TI - Quantification of internal carotid artery stenosis with duplex US: comparative analysis of different flow velocity criteria. AB - PURPOSE: To compare 13 previously published sets of duplex ultrasonographic (US) criteria with the US criteria used at the authors' institution in terms of agreement with carotid artery angiographic results. MATERIALS AND METHODS: The authors studied 1,006 carotid arteries in 503 patients at duplex US and angiography. The degree of stenosis was determined by using duplex flow US velocities and applying 13 previously published sets of criteria and the criteria used at the authors' institution. Two independent observers evaluated the angiograms according to North American Symptomatic Carotid Endarterectomy Trial (NASCET) criteria. kappa statistics, sensitivities, specificities, positive predictive values (PPVs), negative predictive values (NPVs), and generalized linear mixed regression models were used to assess agreement between duplex US and angiographic findings. RESULTS: Stenoses of 0%-29%, 30%-49%, 50%-69%, 70% 99%, and 100% could be differentiated with 73% overall agreement between duplex US and angiographic findings according to flow velocity criteria (kappa = 0.57; 95% confidence interval [CI]: 0.54, 0.60); however, with duplex US, the angiographic degree of stenosis tended to be overestimated. In the differentiation of stenoses of less than 70%, only 45% agreement (kappa = 0.26; 95% CI: 0.23, 0.29) was observed, whereas in the differentiation of high-grade (> or =70%) stenoses, 96% agreement was observed (kappa = 0.85; 95% CI: 0.83, 0.87). The PPV and NPV for the identification of 70%-99% angiographic stenosis were 69% and 98%, respectively, with use of the most sensitive duplex US criteria. CONCLUSION: Duplex US is an excellent examination to screen for high-grade carotid artery stenosis; however, it tends to lead to an overestimation of the degree of stenosis. Exclusion of 70%-99% angiographic stenosis can be achieved with a sensitivity of up to 98%. PMID- 15286316 TI - Supraglottic carcinoma treated with curative radiation therapy: identification of prognostic groups with MR imaging. AB - PURPOSE: To retrospectively assess the prognostic meaning of tumor characteristics depicted on pretreatment magnetic resonance (MR) images for local outcome in supraglottic squamous cell carcinoma treated with definitive radiation therapy. MATERIALS AND METHODS: Pretreatment MR images acquired in 84 patients with supraglottic carcinoma treated with curative radiation therapy were reviewed for tumor involvement of laryngeal sites including glottis, subglottis, pre epiglottic space, laryngeal cartilages, and hypopharynx, and for extralaryngeal extension. The volume of each tumor was estimated, and mean tumor volume was calculated for the group of tumors in each T staging category. RESULTS: Results of univariate analysis showed MR imaging-determined primary tumor volume (P =.03), involvement of pre-epiglottic space (P =.008), abnormal signal intensity in thyroid cartilage (P =.04), and extralaryngeal extension beyond thyroid and/or cricoid cartilage (P =.02) to be significant predictors of local control rate. Results of multivariate analysis with the Cox regression model confirmed statistical significance for invasion of pre-epiglottic space (P =.004) and for abnormal signal intensities in thyroid cartilage adjacent to the anterior commissure (P =.04) and in cricoid cartilage (P =.01). Five-year local control rates were calculated from the regression coefficients of three independent MR imaging prognostic factors, and three prognostic groups were identified on the basis of these control rates. The 5-year local control rate in the high-risk group was 35%, significantly lower than the rates in the intermediate- and low risk groups (60% and 89%, respectively; P =.002). CONCLUSION: MR imaging determined pre-epiglottic space involvement and abnormal signal intensities in the thyroid cartilage adjacent to the anterior commissure and/or the cricoid cartilage are strong predictors of local outcome in supraglottic carcinoma treated with definitive radiation therapy. PMID- 15286317 TI - The Mount Fuji sign. PMID- 15286318 TI - Radiofrequency ablation: in vivo comparison of four commercially available devices in pig livers. AB - PURPOSE: To compare in vivo coagulation necrosis obtained with four radiofrequency (RF) ablation devices, to determine shape and reproducibility of induced coagulation by means of three-dimensional measurements of the ablation zone, and to achieve representations of the coagulated areas in three-dimensional spaces. MATERIALS AND METHODS: Four commercially available RF devices (perfusion, internally cooled cluster, and nine- and 12-tine expandable electrodes) that represent the most widely used systems on the market were tested. Sixteen in vivo ablation procedures were performed in porcine livers (four ablations for each RF system). After macroscopic and histopathologic analyses of 3-mm-thick liver sections, morphometric and volumetric findings in the central zone of white coagulation necrosis were assessed. Coagulation volume, diameter, length, and shape were determined digitally. After analysis of variance, measurements with each system were tested with the Tukey post hoc test. RESULTS: Mean coagulation volumes were 31.5 cm3 +/- 15.8 (SD) for the perfusion electrode, 20.5 cm3 +/- 2.6 for the cluster electrode, 16.2 cm3 +/- 7.3 for the 12-tine electrode, and 9.8 cm3 +/- 3.2 for the nine-tine electrode (P <.05, perfusion vs nine-tine electrode). No significant differences were observed regarding the mean short axis perpendicular to the needle shaft: 2.30 cm +/- 0.94, 3.04 cm +/- 0.26, 3.44 cm +/- 0.21, and 2.70 cm +/- 0.76, respectively. Variation coefficients were 0.50, 0.13, 0.45, and 0.33, respectively. CONCLUSION: Larger coagulation volumes were obtained with the perfusion and internally cooled cluster devices. More spherical volumes of ablation were achieved with the 12-tine and cluster electrodes. The former proved superior with regard to the short axis perpendicular to the needle shaft. The cluster and nine-tine electrode produced better reproducibility, which is suggestive of improved predictability of the extent of coagulation with these systems. PMID- 15286319 TI - Long-term cardiovascular morbidity, mortality, and reintervention after endovascular treatment in patients with iliac artery disease: The Dutch Iliac Stent Trial Study. AB - PURPOSE: To compare long-term cardiovascular morbidity and mortality and their determinants in a population initially treated with one of two endovascular treatment strategies for stenosis or short occlusion of an iliac artery. MATERIALS AND METHODS: A total of 279 symptomatic patients with stenosis or short (< or =5-cm) occlusion of the iliac arteries were randomly assigned to undergo either primary stent placement or primary angioplasty followed by selective stent placement (in case of a residual mean pressure gradient greater than 10 mm Hg at the treated site). Follow-up data for all 279 patients were provided by the general practitioners and referring clinicians. Events of interest were arterial interventions, reinterventions in the iliac arteries, cardiovascular events (myocardial infarction, stroke, or extracranial bleeding), and death. Regression analysis was performed to identify predictors of reintervention and of cardiovascular morbidity and mortality. RESULTS: The mean follow-up period was 5.6 years +/- 1.3 (+/- standard deviation). There were no significant differences between primary stent placement and primary angioplasty treatment groups in regard to number of reinterventions in the treated iliac arteries (33 [18%] of 187 segments and 33 [20%] of 169 segments, respectively) or in the ipsilateral legs (45 [25%] of 181 legs and 50 [30%] of 164 legs, respectively). The risk of other cardiovascular events in primary stent placement and primary angioplasty groups was 13% (18 of 143) and 11% (15 of 136), and the risk of death was 15% (21 of 143 patients) and 16% (22 of 136 patients), respectively. Sex, presence of critical ischemia, and length of stenosis were predictors of whether a patient would require iliac reintervention. Myocardial infarction, stroke, and vascular death were predicted on the basis of a patient's creatinine level and walking distance as tested at the time of inclusion. CONCLUSION: No difference was found in the number of reinterventions between the two treatment groups 5 years after treatment. Patients with iliac artery disease are at high risk of cardiovascular morbidity and mortality. PMID- 15286320 TI - Percutaneous intentional extraluminal recanalization in patients with chronic critical limb ischemia. AB - PURPOSE: To review percutaneous intentional extraluminal recanalization (PIER) for treatment of patients who are poor candidates for infrainguinal arterial bypass surgery (IABS) and have arterial occlusions and chronic critical limb ischemia (CCLI). MATERIALS AND METHODS: Patients with CCLI who were poor candidates for IABS were candidates for PIER. PIER was performed to create continuous arterial flow to the foot for limb salvage. PIER was attempted in 40 patients (22 men, 18 women; median age, 69 years; age range, 44-87 years). Of these patients, 24 (60%) had diabetes, 17 (42%) had renal disease, and 26 (66%) had coronary artery disease. Wound healing was evaluated at follow-up. Kaplan Meier curves were constructed to evaluate limb salvage, survival, and amputation free survival. RESULTS: Fifty procedures were attempted in 44 limbs. Tissue loss was present in 40 (91%) limbs, and rest pain was present in four (9%); technical success occurred in 38 (86%). Thirty-seven (84%) of 44 limbs treated with PIER involved tibial vessels (tibial vessels only, n = 15; tibial and superior femoral artery [SFA] and/or popliteal vessels, n = 22). Sixty-six infrainguinal arterial vessel segments (SFA, n = 29; tibial, n = 37) in 38 limbs (1.7 segments per limb) were successfully treated with PIER. Thirty-five (95%) of 37 tibial occlusions and 24 (83%) of 29 SFA and/or popliteal occlusions were longer than 10 cm. Median run-off scores were 5.3 (range, 3-8) and 6.6 (range, 3-9) for patients with tibial occlusions and SFA and/or popliteal occlusions, respectively, as scored with modified Rutherford weighting of run-off arteries. Median follow-up was 7.8 months (range, 1-24 months). Twelve months after PIER, Kaplan-Meier analysis showed limb salvage rate was 66%, survival rate was 71%, and amputation-free survival rate was 48% in these patients. The 30-day mortality rate was 2.5%. Major complications occurred in four (10%) patients, and minor complications occurred in an additional four (10%). CONCLUSION: PIER is a useful percutaneous technique for limb salvage in patients with CCLI. PMID- 15286321 TI - Dysfunctional autogenous hemodialysis fistulas: outcomes after angioplasty--are there clinical predictors of patency? AB - PURPOSE: To determine the primary and secondary patency rates for fistulas treated with angioplasty, as well as clinical predictors of fistula patency after angioplasty. MATERIALS AND METHODS: The authors reviewed their institutional experience with autogenous fistulas from June 1997 to June 2002. A total of 104 men and 36 women were treated. Mean age +/- standard deviation of patient cohort was 62.4 years +/- 15.6. Patient age and sex, age of fistula at initial intervention, presence of diabetes, side and location of fistula, location of stenosis, and number of venous stenoses dilated were examined. Patency after angioplasty was estimated by using the Kaplan-Meier method, and predictors of patency were examined by using a Cox proportional hazards model. RESULTS: One hundred fifty-one dysfunctional fistulas (94 radiocephalic and 57 brachiocephalic) were treated with angioplasty initially. Clinical success rate was 98.0% (297 of 303 interventions). At 3, 6, and 12 months, respectively, primary patency rates +/- standard errors of the estimate were 73% +/- 6, 51% +/- 7, and 39% +/- 7 for brachiocephalic fistulas and 85% +/- 4, 75% +/- 5, and 62% +/- 5 for radiocephalic fistulas; secondary patency rates were 96% +/- 2.4, 89% +/- 4, and 85% +/- 5 for brachiocephalic fistulas and 91% +/- 3, 88% +/- 3, and 86% +/- 4 for radiocephalic fistulas. For all time points, there was a significant difference in primary (P =.004) but not secondary (P =.45) patency between radiocephalic and brachiocephalic fistulas. Stenosis was most prevalent within 3 cm of the arteriovenous anastomosis in 74 (64%) of the 116 dysfunctional radiocephalic fistulas and at the cephalic arch in 22 (30%) of the 74 dysfunctional brachiocephalic fistulas. The clinical variables examined did not influence outcome. Complications occurred in seven (2.3%) of 303 interventions. CONCLUSION: Patency after angioplasty in dysfunctional autogenous hemodialysis fistulas exceeds that observed in hemodialysis grafts. None of the clinical or anatomic variables examined affected patency outcome. PMID- 15286322 TI - Primary patency of femoropopliteal arteries treated with nitinol versus stainless steel self-expanding stents: propensity score-adjusted analysis. AB - PURPOSE: To evaluate, in a propensity score-adjusted analysis, the intermediate term primary patency rates associated with nitinol versus stainless steel self expanding stent placement for treatment of atherosclerotic lesions in femoropopliteal arteries. MATERIALS AND METHODS: The authors analyzed the clinical and imaging data of 175 consecutive patients with peripheral artery disease and either intermittent claudication (n = 150) or critical limb ischemia (n = 25) who underwent femoropopliteal artery implantation of nitinol (n = 104) or stainless steel (n = 123) stents in a nonrandomized setting. The stents were placed owing to either significant residual stenosis (ie, >30% lumen diameter reduction) or flow-limiting dissection after initial balloon angioplasty of the femoropopliteal artery. Patients were followed up for a median period of 9 months (mean, 13 months; range, 6-66 months) for the detection of a first in-stent restenosis, defined as a greater than 50% lumen diameter reduction that was seen at color-coded duplex ultrasonography and confirmed at angiography. RESULTS: Cumulative patency rates at 6, 12, and 24 months were 85%, 75%, and 69%, respectively, after nitinol stent placement versus 78%, 54%, and 34%, respectively, after stainless steel stent placement (P =.008, log-rank test). There were no statistically significant differences in associated patency among the three different nitinol stents used (P =.72, log-rank test). Multivariate Cox proportional hazard analysis, in which the effect of propensity to receive a nitinol stent was considered, revealed a significantly reduced risk of restenosis with the nitinol stents compared with the risk of restenosis with the stainless steel stents (adjusted hazard ratio, 0.44; 95% confidence interval: 0.22, 0.85; P =.014). CONCLUSION: Nitinol stents are associated with significantly improved primary patency rates in femoropopliteal arteries compared with stainless steel stents. Randomized controlled trials are needed to confirm these results. PMID- 15286323 TI - 131I therapeutic efficacy is not influenced by stunning after diagnostic whole body scanning. AB - PURPOSE: To determine if stunning can be seen with a 185-MBq (5-mCi) dose of iodine 131 (131I) at diagnostic whole-body scanning and, if stunning is seen, determine if there is any 131I therapeutic efficacy. MATERIALS AND METHODS: A retrospective review of findings involving 166 patients who underwent thyroidectomy for differentiated thyroid carcinoma was performed. Diagnostic 131I scans were compared with postablation scans for evidence of stunning. Stunning was defined when the diagnostic scan showed activity that was subsequently decreased on the postablation scan. The sample population was divided into two groups: group NS, patients with no stunning, and group S, patients with stunning. Patients were considered successfully treated if no functioning thyroid tissue and/or metastases were seen on follow-up diagnostic scans. Fisher exact and Student t tests were used to evaluate the statistical significance of therapy success rates, clinical characteristics, and scanning parameters between the two groups. RESULTS: Group NS included 135 (81.3%) of 166 patients, with 36 (26.7%) of 135 lost to follow-up. Group S included 31 (18.7%) of 166 patients, with eight (26%) of 31 patients lost to follow-up. There was no significant difference (P =.61) in treatment success rates between group NS (87 of 99, 88%) and group S (21 of 23, 91%). The treatment success rates for thyroid remnants were 87% (48 of 55) for group NS and 91% (10 of 11) for group S (P =.63). Treatment success rates for metastases (mostly lymph nodes) were 89% (39 of 44) for group NS and 83% (10 of 12) for group S (P =.55). CONCLUSION: Thyroid stunning can occur with 185 MBq of 131I in diagnostic imaging. However, data did not show any effect of stunning on the efficacy of 131I therapy for differentiated thyroid carcinoma. PMID- 15286324 TI - CT features of lung disease in patients with systemic sclerosis: comparison with idiopathic pulmonary fibrosis and nonspecific interstitial pneumonia. AB - PURPOSE: To evaluate computed tomographic (CT) patterns of lung disease in patients with systemic sclerosis (SSc) and compare them with CT appearance in patients with biopsy-proved idiopathic pulmonary fibrosis (IPF) and idiopathic nonspecific interstitial pneumonia (NSIP). MATERIALS AND METHODS: The CT features of consecutive patients with SSc (n = 225; male patients, 44; female patients, 181; median age, 47 years; age range, 16-78 years), IPF (n = 40; men, 26; women, 14; median age, 54.5 years; age range, 36-77 years) and NSIP (n = 27; men, 18; women, nine; median age, 53 years; age range, 32-68 years) were quantified separately by two observers. The extent of interstitial lung disease, ground glass opacification, emphysema, and the coarseness of a reticular pattern were quantified. Group comparisons were made nonparametrically with the Wilcoxon rank sum test. Differences in CT features were identified with multiple logistic regression analysis. RESULTS: The coarseness of fibrosis was similar in patients with SSc and idiopathic NSIP but strikingly different between patients with SSc (median coarseness score, 5.5; range, 0.0-13.3) and IPF (median coarseness score, 8.8; range, 2.5-15.0) (P <.001). The proportion of ground-glass opacification at CT was similar in patients with SSc and idiopathic NSIP but differed significantly between patients with SSc (median proportion, 49.9%; range, 0.0% 100.0%) and IPF (median proportion, 23.5%; range, 0.0%-97.2%) (P <.001). At logistic regression analysis, there were no differences in the CT features between patients with SSc and those with NSIP after controlling for age, disease extent, and the percentage predicted forced vital capacity and carbon monoxide diffusing capacity. CONCLUSION: Interstitial lung disease in patients with SSc is less extensive, less coarse, and characterized by a greater proportion of ground glass opacification than that in patients with IPF. The CT features of lung disease in patients with SSc closely resemble those in patients with idiopathic NSIP. PMID- 15286325 TI - Characterization of bone and soft-tissue tumors with in vivo 1H MR spectroscopy: initial results. AB - PURPOSE: To determine if in vivo detection of choline by using hydrogen 1 (1H) magnetic resonance (MR) spectroscopy with dynamic contrast material-enhanced MR imaging can help differentiate between benign and malignant musculoskeletal tumors. MATERIALS AND METHODS: MR imaging was performed in 36 consecutive patients with bone and soft-tissue tumors larger than 1.5 cm in diameter. Examinations were performed at 1.5 T with a surface coil appropriate for the location of the lesions. Single-voxel 1H MR spectroscopy was performed by using a point-resolved spectroscopic sequence with echo times of 40, 135, and 270 msec. The volume of interest within lesions was positioned on the areas of early enhancement (<8 seconds after arterial enhancement) according to the findings of dynamic contrast-enhanced MR imaging with subtraction. The criterion for determining whether choline was present in a lesion was a clearly identifiable peak at 3.2 ppm in at least two of the three spectra acquired at echo times. MR spectroscopic results and histopathologic findings were determined in blinded fashion and compared with kappa statistics. P <.001 was considered to indicate a significant difference. RESULTS: Choline was detected in 18 of 19 patients with malignant tumors and in three of 17 patients with benign lesions. The three benign lesions included one perineurioma, one giant cell tumor, and one abscess. Choline was not detected in 14 patients with benign lesions nor in one patient with a densely ossifying low-grade parosteal osteosarcoma. In vivo 1H MR spectroscopy characterized bone and soft-tissue tumors, resulting in a sensitivity of 95%, specificity of 82%, and accuracy of 89% (P <.001). CONCLUSION: Choline can be reliably detected in large malignant bone and soft tissue tumors by using a multiecho point-resolved spectroscopic protocol. 1H MR spectroscopy can help differentiate malignant from benign musculoskeletal tumors by revealing the presence or absence of water-soluble choline metabolites. PMID- 15286326 TI - Acute pulmonary embolism: thin-collimation spiral CT versus planar ventilation perfusion scintigraphy. PMID- 15286327 TI - The golden hour: how to spend your time and money in trauma radiology. PMID- 15286328 TI - Comments on T2 measurements of prostate tissue. PMID- 15286329 TI - Standardization of terms and reporting criteria for image-guided tumor ablation. PMID- 15286330 TI - Here come the Olympics. PMID- 15286331 TI - Theoretical physics. Hawking slays his own paradox, but colleagues are wary. PMID- 15286332 TI - U.S. science budget. Caught in a squeeze between tax cuts and military spending. PMID- 15286333 TI - Biomedicine. An end to the prion debate? Don't count on it. PMID- 15286335 TI - Environment. States sue over global warming. PMID- 15286334 TI - Archaeology: Wisconsin dig seeks to confirm pre-Clovis Americans. PMID- 15286336 TI - Kennewick Man. Court battle ends, bones still off-limits. PMID- 15286338 TI - National labs. Los Alamos suspends 19 employees. PMID- 15286337 TI - U.S. science policy. Congressmen clash on politics and scientific advisory committees. PMID- 15286339 TI - Virology. Tiptoeing around Pandora's Box. PMID- 15286340 TI - Neuroscience. Crime, culpability, and the adolescent brain. PMID- 15286341 TI - Neuroscience. Adolescence: akin to mental retardation? PMID- 15286342 TI - Solar physics. Solar physicists expose the roots of the sun's unrest. PMID- 15286343 TI - NIOSH and the CDC reorganization. PMID- 15286344 TI - NIOSH's work on firefighting. PMID- 15286345 TI - International students at Vanderbilt. PMID- 15286346 TI - Foreign-born physician-scientists. PMID- 15286347 TI - Asian brains: U.S. v. Europe. PMID- 15286348 TI - Steps to end the obesity epidemic. PMID- 15286349 TI - The virtues of teaching. PMID- 15286350 TI - Climate. The Bush administration's approach to climate change. PMID- 15286352 TI - Ecology. Herbivores rule. PMID- 15286351 TI - Ecology. Ecogenomics benefits community ecology. PMID- 15286353 TI - Geology. A new period for the geologic time scale. PMID- 15286354 TI - Planetary science. A unique chunk of the Moon. PMID- 15286355 TI - Materials science. Watching the nanograins roll. PMID- 15286356 TI - The pathophysiology of mitochondrial cell death. AB - In the mitochondrial pathway of apoptosis, caspase activation is closely linked to mitochondrial outer membrane permeabilization (MOMP). Numerous pro-apoptotic signal-transducing molecules and pathological stimuli converge on mitochondria to induce MOMP. The local regulation and execution of MOMP involve proteins from the Bcl-2 family, mitochondrial lipids, proteins that regulate bioenergetic metabolite flux, and putative components of the permeability transition pore. MOMP is lethal because it results in the release of caspase-activating molecules and caspase-independent death effectors, metabolic failure in the mitochondria, or both. Drugs designed to suppress excessive MOMP may avoid pathological cell death, and the therapeutic induction of MOMP may restore apoptosis in cancer cells in which it is disabled. The general rules governing the pathophysiology of MOMP and controversial issues regarding its regulation are discussed. PMID- 15286358 TI - A race to the starting line. PMID- 15286359 TI - Mighty mice: inspiration for rogue athletes? PMID- 15286360 TI - Do pool sharks swim faster? PMID- 15286361 TI - Peering under the hood of Africa's runners. PMID- 15286362 TI - An everlasting gender gap? PMID- 15286363 TI - Graceful, beautiful, and perilous. PMID- 15286364 TI - Engineering peak performance. PMID- 15286365 TI - Long gone or gone wrong? PMID- 15286366 TI - APOBEC-mediated editing of viral RNA. AB - Retroviral DNA can be subjected to cytosine-to-uracil editing through the action of members of the APOBEC family of cytidine deaminases. Here we demonstrate that APOBEC-mediated cytidine deamination of human immunodeficiency virus (HIV) virion RNA can also occur. We speculate that the natural substrates of the APOBEC enzymes may extend to RNA viruses that do not replicate through DNA intermediates. Thus, cytosine-to-uracil editing may contribute to the sequence diversification of many viruses. PMID- 15286367 TI - Engineered interface of magnetic oxides. AB - Interface-selective probing of magnetism is a key issue for the design and realization of spin-electronic junction devices. Here, magnetization-induced second-harmonic generation was used to probe the local magnetic properties at the interface of the perovskite ferromagnet La(0.6)Sr(0.4)MnO3 with nonmagnetic insulating layers, as used in spin-tunnel junctions. We show that by grading the doping profile on an atomic scale at the interface, robust ferromagnetism can be realized around room temperature. The results should lead to improvements in the performance of spin-tunnel junctions. PMID- 15286368 TI - Grain boundary-mediated plasticity in nanocrystalline nickel. AB - The plastic behavior of crystalline materials is mainly controlled by the nucleation and motion of lattice dislocations. We report in situ dynamic transmission electron microscope observations of nanocrystalline nickel films with an average grain size of about 10 nanometers, which show that grain boundary mediated processes have become a prominent deformation mode. Additionally, trapped lattice dislocations are observed in individual grains following deformation. This change in the deformation mode arises from the grain size dependent competition between the deformation controlled by nucleation and motion of dislocations and the deformation controlled by diffusion-assisted grain boundary processes. PMID- 15286369 TI - Pinpointing the source of a lunar meteorite: implications for the evolution of the Moon. AB - The lunar meteorite Sayh al Uhaymir 169 consists of an impact melt breccia extremely enriched with potassium, rare earth elements, and phosphorus [thorium, 32.7 parts per million (ppm); uranium, 8.6 ppm; potassium oxide, 0.54 weight percent], and adherent regolith. The isotope systematics of the meteorite record four lunar impact events at 3909 +/- 13 million years ago (Ma), approximately 2800 Ma, approximately 200 Ma, and <0.34 Ma, and collision with Earth sometime after 9.7 +/- 1.3 thousand years ago. With these data, we can link the impact melt breccia to Imbrium and pinpoint the source region of the meteorite to the Lalande impact crater. PMID- 15286370 TI - Foundering lithosphere imaged beneath the southern Sierra Nevada, California, USA. AB - Seismic tomography reveals garnet-rich crust and mantle lithosphere descending into the upper mantle beneath the southeastern Sierra Nevada. The descending lithosphere consists of two layers: an iron-rich eclogite above a magnesium-rich garnet peridotite. These results place descending eclogite above and east of high P wave speed material previously imaged beneath the southern Great Valley, suggesting a previously unsuspected coherence in the lithospheric removal process. PMID- 15286371 TI - Herbivores promote habitat specialization by trees in Amazonian forests. AB - In an edaphically heterogeneous area in the Peruvian Amazon, clay soils and nutrient-poor white sands each harbor distinctive plant communities. To determine whether a trade-off between growth and antiherbivore defense enforces habitat specialization on these two soil types, we conducted a reciprocal transplant study of seedlings of 20 species from six genera of phylogenetically independent pairs of edaphic specialist trees and manipulated the presence of herbivores. Clay specialist species grew significantly faster than white-sand specialists in both soil types when protected from herbivores. However, when unprotected, white sand specialists dominated in white-sand forests and clay specialists dominated in clay forests. Therefore, habitat specialization in this system results from an interaction of herbivore pressure with soil type. PMID- 15286372 TI - Osedax: bone-eating marine worms with dwarf males. AB - We describe a new genus, Osedax, and two new species of annelids with females that consume the bones of dead whales via ramifying roots. Molecular and morphological evidence revealed that Osedax belongs to the Siboglinidae, which includes pogonophoran and vestimentiferan worms from deep-sea vents, seeps, and anoxic basins. Osedax has skewed sex ratios with numerous dwarf (paedomorphic) males that live in the tubes of females. DNA sequences reveal that the two Osedax species diverged about 42 million years ago and currently maintain large populations ranging from 10(5) to 10(6) adult females. PMID- 15286373 TI - The complete genome sequence of Propionibacterium acnes, a commensal of human skin. AB - Propionibacterium acnes is a major inhabitant of adult human skin, where it resides within sebaceous follicles, usually as a harmless commensal although it has been implicated in acne vulgaris formation. The entire genome sequence of this Gram-positive bacterium encodes 2333 putative genes and revealed numerous gene products involved in degrading host molecules, including sialidases, neuraminidases, endoglycoceramidases, lipases, and pore-forming factors. Surface associated and other immunogenic factors have been identified, which might be involved in triggering acne inflammation and other P. acnes-associated diseases. PMID- 15286374 TI - Synthetic mammalian prions. AB - Recombinant mouse prion protein (recMoPrP) produced in Escherichia coli was polymerized into amyloid fibrils that represent a subset of beta sheet-rich structures. Fibrils consisting of recMoPrP(89-230) were inoculated intracerebrally into transgenic (Tg) mice expressing MoPrP(89-231). The mice developed neurologic dysfunction between 380 and 660 days after inoculation. Brain extracts showed protease-resistant PrP by Western blotting; these extracts transmitted disease to wild-type FVB mice and Tg mice overexpressing PrP, with incubation times of 150 and 90 days, respectively. Neuropathological findings suggest that a novel prion strain was created. Our results provide compelling evidence that prions are infectious proteins. PMID- 15286375 TI - KIF1A alternately uses two loops to bind microtubules. AB - The motor protein kinesin moves along microtubules, driven by adenosine triphosphate (ATP) hydrolysis. However, it remains unclear how kinesin converts the chemical energy into mechanical movement. We report crystal structures of monomeric kinesin KIF1A with three transition-state analogs: adenylyl imidodiphosphate (AMP-PNP), adenosine diphosphate (ADP)-vanadate, and ADP-AlFx (aluminofluoride complexes). These structures, together with known structures of the ADP-bound state and the adenylyl-(beta,gamma-methylene) diphosphate (AMP-PCP) bound state, show that kinesin uses two microtubule-binding loops in an alternating manner to change its interaction with microtubules during the ATP hydrolysis cycle; loop L11 is extended in the AMP-PNP structure, whereas loop L12 is extended in the ADP structure. ADP-vanadate displays an intermediate structure in which a conformational change in two switch regions causes both loops to be raised from the microtubule, thus actively detaching kinesin. PMID- 15286377 TI - Pathogenesis and spectrum of autoimmunity. AB - The immune system specifically recognizes and eliminates foreign antigens and thus protects the integrity of the host. During maturation of the immune system, tolerance mechanisms develop that prevent or inhibit potentially harmful reactivities to self-antigens. Autoreactive B and T cells that are generated during immune responses are eliminated by apoptosis in the thymus, lymph nodes, or peripheral circulation or are actively suppressed by regulatory T cells. However, autoreactive cells may survive because of failure of apoptosis or molecular mimicry, that is, presentation and recognition of cryptic epitopes of self-antigens or aberrant lymphokine production. Development of immune responses and tolerance is determined by an interplay of genetic and environmental factors. Autoimmunity is a result of the breakdown of one or more of the mechanisms of immune tolerance. PMID- 15286378 TI - Mapping the systematic lupus erythematosus susceptibility genes. AB - Systemic lupus erythematosus (SLE) is a prototype systemic, autoimmune inflammatory disease that can involve virtually any organ or tissue type. The disease has a strong familial tendency but, like most human illness, has a complex pattern of inheritance that is consistent with multiple susceptibility genes as well as environmental risk factors. Association studies have been performed, especially for the major histocompatibility complex on chromosome 6 and for various complement components. Several large familial studies have begun to report results for genetic linkage. Linkage has been established for many genetic intervals. SLE is a complex clinical illness, and investigation of the genetics of the illness based on clinical manifestations revealed linkages not found without consideration of the phenotype of the disease. PMID- 15286379 TI - T-cell signaling abnormalities in human systemic lupus erythematosus. AB - Abnormal expression of key signaling molecules and defective functions of T lymphocytes play a significant role in the pathogenesis of systemic lupus erythematosus (SLE). T-cell receptor (TCR)/CD3-mediated stimulation of SLE T cells shows increased protein tyrosine phosphorylation of cellular proteins, with faster kinetics, heightened calcium response, and decreased interleukin (IL)-2 production. The molecular mechanism of T-cell signaling abnormalities in SLE T cells is complex and cannot be explained fully by the current theories of T-cell signaling. Current research on lymphocyte signaling abnormalities in SLE has been directed toward investigating various factors that contribute to abnormal tyrosine phosphorylation, intracellular calcium response, and cytokine production. Latest developments suggest multiple components, including altered receptor structure, supramolecular assembly, modulation of membrane clustering, aberrant cellular distribution, and precompartmentalization with lipid rafts invariably contributing to abnormal T-cell signaling in SLE T cells. The methods and protocols described here pertaining to T-cell signaling abnormalities in SLE T cells are very much optimized in many ways, and they were derived by the combined tasks and continuous efforts of many researchers in the laboratory over a long period. These simplified protocols can be readily applied to study T-cell signaling abnormalities in SLE to identify the genetic, molecular, and biochemical factors contributing to aberrant immune cell function and unravel the pathophysiology of SLE. PMID- 15286380 TI - TCR zeta-chain abnormalities in human systemic lupus erythematosus. AB - A growing number of studies have revealed that the expression of many genes is abnormal in T lymphocytes of patients with systemic lupus erythematosus (SLE). Although aberrant expression of signaling molecules may arise intrinsically or in response to the environment, these abnormalities play a significant role in the pathogenesis of this autoimmune disease. Modern research on lymphocyte signaling abnormalities in SLE has been directed toward identifying defective expression of various signaling molecules, understanding the molecular basis of the deficiency, and dissecting the T-cell signaling abnormalities that result from abnormal gene expression. The developments suggest that interplay of abnormal transcriptional factor, aberrant messenger RNA processing/editing, ubiquitination, proteolysis, oxidative stress, and changes in chromatin structure invariably contribute to the abnormal expression of numerous signaling molecules in SLE T cells. The contribution of each of these mechanisms in the abnormal expression of signaling molecules in SLE T cells is not known. In addition to abnormalities in gene expression, multiple factors, including altered cellular distribution of the protein, rewiring of the receptor, modulation of membrane clustering, and lipid raft distribution of signaling molecules and defective signal-silencing mechanisms play a key role in delivering the anomalous T-cell receptor/CD3 mediated intracellular calcium response in SLE T cells. The optimized methods and protocols described here pertaining to TCR zeta-chain expression and related T cell signaling abnormalities can be very well applied to other molecules aberrantly expressed in SLE T cells. PMID- 15286381 TI - Protein kinase A and signal transduction in T lymphocytes: biochemical and molecular methods. AB - Abnormal T-cell effector functions in systemic lupus erythematosus (SLE) are present and may be associated with disease immunopathogenesis. Our work has led to the characterization of a signaling defect, involving protein kinase A (PKA), leading to abnormal T-cell effector functions in SLE. PKA is a component of the adenylyl cyclase/cyclic adenosine monophosphate/PKA (AC/cAMP/PKA) pathway, a principal signal transduction system in T cells. The aim of this chapter is to provide a comprehensive, technical, step-by-step approach to studying PKA function in T cells. The methods detailed here are (a) chromatographic fractionation of PKA-I and PKA-II isozymes and PKA phosphotransferase activity in purified T cell populations, (b) Western immunoblotting to identify the presence of regulatory (R)-subunit proteins of PKA, and (c) isolation of RNA, and quantification of PKA R subunit-specific transcripts by competitive polymerase chain reaction. Although our emphasis in the chapter is T cells, these methods may be useful for investigation of signaling via PKA in other cell types as well. PMID- 15286382 TI - Apoptosis and mitochondrial dysfunction in lymphocytes of patients with systemic lupus erythematosus. AB - Systemic lupus erythematosus (SLE) is characterized by abnormal activation and cell death signaling within the immune system. Activation, proliferation, or death of cells of the immune system are dependent on controlled reactive oxygen intermediate (ROI) production and ATP synthesis in mitochondria. The mitochondrial transmembrane potential (Delta(Psi)m) reflects the energy stored in the electrochemical gradient across the inner mitochondrial membrane, which in turn is used by F0F1-ATPase to convert adenosine 5'-diphosphate to ATP during oxidative phosphorylation. Mitochondrial hyperpolarization and transient ATP depletion represent early and reversible steps in T-cell activation and apoptosis. By contrast, T lymphocytes of patients with SLE exhibit elevated Delta(Psi)m, that is, persistent mitochondrial hyperpolarization, cytoplasmic alkalinization, increased ROI production, as well as diminished levels of intracellular glutathione and ATP. Oxidative stress affects signaling through the T-cell receptor as well as the activity of redox-sensitive caspases. ATP depletion may be responsible for diminished activation-induced apoptosis and sensitize lupus T cells to necrosis. Mitochondrial dysfunction is identified as a key mechanism in the pathogenesis of SLE. PMID- 15286383 TI - Methods for inducing apoptosis. AB - Apoptotic cells are sources of tolerogenic material during tissue homeostasis; abnormalities in apoptosis or in the clearance of apoptotic material generate a novel source of antigens against which an autoimmune response may be initiated. In our laboratory, we study the biochemistry and cell biology of systemic autoimmune disease autoantigens during different forms of cell death. Several different methods for inducing apoptosis, and for assaying the induction of this cellular process, are routinely performed. This chapter describes methods for inducing apoptosis via ultraviolet B irradiation, small molecule drug treatments, death receptor ligation, and exposure to granule components of cytotoxic lymphocytes. Assays to confirm the induction of apoptosis by quantifying changes in mitochondrial membrane potential, phosphatidylserine membrane localization, DNA content, and autoantigen cleavage are also detailed. PMID- 15286384 TI - Measurement of cytokines in autoimmune disease. AB - Systemic autoimmune diseases are characterized by extensive alterations in immune system function, with cytokines and autoantibodies contributing to impaired immunoregulation and tissue damage. Characterization of the expression and function of cytokines is important for elucidation of pathogenic mechanisms and for identification of therapeutic targets and strategies. We reviewed the utility of assays that reflect individual variability in cytokine gene sequence, expression of messenger ribonucleic acid (mRNA) or protein, as well as measurement of expression of target genes regulated by cytokines. Real-time reverse transcriptase polymerase chain reaction and intracellular staining for cytokine expression are two sensitive and quantitative approaches for analysis of cytokine mRNA and protein, respectively. Detailed methods are provided for these assays. PMID- 15286385 TI - Evaluation of autoimmunity to transaldolase in multiple sclerosis. AB - Transaldolase is a target of autoimmunity mediated by T cells and antibody (Ab) in patients with multiple sclerosis. Functional T-cell assays, T- and B-cell epitope mapping, and detection of transaldolase-specific antibodies in patients with multiple sclerosis are described. Recombinant transaldolase was produced in a prokaryotic expression vector for use in Western blot analysis of sera of these patients. Overlapping transaldolase peptides 15 amino acids (aa) long were synthesized onto cellulose membranes to map immunodominant B-cell epitopes. Amino acid sequence homologies between viral peptides and immunodominant B-cell epitopes of transaldolase were identified using a computer-based algorithm. Direct assessment of molecular mimicry between transaldolase B-cell epitopes and related viral peptides is also shown. T-cell epitopes are mapped in a T-cell proliferation assay using multiple sclerosis patient and control donor cells. Autoantigen-specific T cells are identified by MHC-peptide tetramer staining using flow cytometry analysis. PMID- 15286386 TI - Animal models for autoimmune myocarditis and autoimmune thyroiditis. AB - This chapter describes four murine models of autoimmune diseases: two related to autoimmune myocarditis and two related to autoimmune thyroiditis. The first model, Coxsackie virus B3 (CB3)-induced myocarditis, results in the development of acute myocarditis in susceptible as well as resistant mouse strains, whereas chronic myocarditis develops only in genetically susceptible mice. CB3-induced myocarditis closely resembles the course of human myocarditis, which is believed to be initiated by viral infection. Mouse cardiac myosin heavy chain has been identified as the major antigen associated with the late chronic phase of viral myocarditis. The second model is cardiac myosin-induced experimental autoimmune myocarditis (EAM) and, in a modification, cardiac alpha-myosin heavy chain peptide-induced myocarditis. In the EAM model, cardiac myosin or the relevant peptide in Freund's complete adjuvant (FCA) is injected subcutaneously into mice. The immune response, the histological changes, and the genetic susceptibility seen in EAM are similar to those of CB3-induced myocarditis. The third model is experimental autoimmune thyroiditis (EAT). EAT can be induced in genetically susceptible strains of mice by immunization with mouse thyroglobulin in FCA or lipopolysaccharide. Mice susceptible to EAT have the H-2A(k), H-2A(s), or H-2A(q) alleles. We describe here a standard technique for the induction of EAT; it was developed in our laboratory and is widely used as a model for studying Hashimoto's thyroiditis. The fourth model presented in this chapter is that of spontaneous autoimmune thyroiditis in NOD.H2h4 mice. These mice express the H 2A(k) allele on an NOD genetic background and develop spontaneous thyroiditis, which is exacerbated with dietary iodine. PMID- 15286387 TI - Animal models of insulin-dependent diabetes. AB - Animal models have contributed enormously to study in the field of type 1 diabetes. Perhaps the most intensively studied model is the nonobese diabetic (NOD) mouse, which develops an autoimmune-mediated spontaneous diabetes associated with the development of insulin autoantibodies and insulitis. Accurate measurement of antiislet autoantibodies by radioassay and detection of antigen specific T cells using major histocompatibility complex tetramers are possible. Various strategies have been developed in preventing diabetes in animal models; a peptide-induced model of type 1 diabetes has been described. Finally, the development of peptide vaccines is hampered by the risk of anaphylaxis in both mouse and humans. In this chapter, methods and strategies to measure antiinsulin autoantibodies, to detect antigen-specific T cells by tetramer analysis, and to prevent diabetes using peptide vaccines are discussed. Along with these topics, a protocol of peptide-induced diabetes and peptide vaccine-induced anaphylaxis are described, serving as a reminder of the potential dangers that could exist in human trials. In summary, animal models have become necessary in the study of type 1 diabetes and provide researchers important tools to conduct studies that could not otherwise be performed in humans. PMID- 15286388 TI - Generation, maintenance, and adoptive transfer of diabetogenic T-cell lines/clones from the nonobese diabetic mouse. AB - The ability to generate, maintain, and use cloned lines of T cells reactive for self-antigens has opened up a new avenue of investigation for researchers. These T-cell clones allow the rapid induction of tissue-specific autoimmunity with the intent of dissecting the contribution of the different cell types involved. T cells from the diabetes-prone nonobese diabetic mouse are proving to be a vital asset for understanding the T-cell-mediated pathogenesis that leads to overt beta cell destruction. T-cell clone adoptive transfer protocols have been developed for use in immunodeficient strains, thus reducing the complexity of mechanism of disease initiation. Furthermore, these T-cell clones have been used to derive T cell receptor transgenic (TCR-Tg) animals carrying only self-reactive T cells. The use of these TCR-Tg animals to study pathogenesis has also evolved from the ability to generate, maintain, and use T-cell cloned lines. This chapter focuses on primary culture for the generation of T-cell lines and clones, their long-term maintenance, and their use in disease transfer for studying the pathogenesis of end-organ autoimmunity. PMID- 15286389 TI - Experimental use of murine lupus models. AB - Murine models of systemic lupus erythematosus provide fertile research systems for the pathogenesis and therapy of systemic autoimmune disease. Their phenotypes span the broad range of clinical manifestations of human lupus and consist of both spontaneous and experimentally induced disease in both inbred and targeted mutant animals. This chapter contrasts the clinical characteristics of these various models, providing an outline for the use and analysis of these in vivo autoimmune systems. PMID- 15286390 TI - The anti-DNA knock-in model of systemic autoimmunity induced by the chronic graft vs-host reaction. AB - The injection of spleen cells from bm12 mice into C57BL/6 recipients induces a chronic graft-vs-host reaction characterized by systemic autoimmunity, including anti-double-stranded DNA (anti-dsDNA) autoantibodies and immune complex-type proliferative glomerulonephritis. If the B6 recipient mice express an anti-DNA Vh site-directed transgene, the repertoire is skewed even more toward the anti-DNA response. Over a period of several weeks, high titers of serum anti-DNA antibodies appear and the mice develop renal damage. This permits the examination of the role of somatic immunoglobulin genetics and B-cell tolerance in a model of systemic lupus erythematosus. PMID- 15286391 TI - Murine models of lupus induced by hypomethylated T cells. AB - CD4+ T-cell DNA hypomethylation may contribute to the development of drug-induced and idiopathic human lupus. Inhibiting DNA methylation in mature CD4+ T cells causes autoreactivity specific to the major histocompatibility complex in vitro. The lupus-inducing drugs hydralazine and procainamide also inhibit T-cell DNA methylation and induce autoreactivity, and T cells from patients with active lupus have hypomethylated DNA and a similarly autoreactive T-cell subset. Further, T cells treated with DNA methylation inhibitors demethylate the same sequences that demethylate in T cells from patients with active lupus. The pathological significance of the autoreactivity induced by inhibiting T-cell DNA methylation has been tested by treating murine T cells in vitro with drugs that modify DNA methylation, then injecting the cells into syngeneic female mice. Mice receiving CD4+ T cells demethylated by a variety of agents, including procainamide and hydralazine, develop a lupuslike disease. This chapter describes the protocols for inducing autoreactivity in murine T cells in vitro and using the cells to induce autoimmunity in vivo. PMID- 15286392 TI - The mouse model of collagen-induced arthritis. AB - Collagen-induced arthritis (CIA) is an experimental autoimmune disease that can be elicited in susceptible strains of rodents (rat and mouse) and nonhuman primates by immunization with type II collagen (CII), the major constituent protein of articular cartilage. Following immunization, these animals develop an autoimmune polyarthritis that shares several clinical and histological features with rheumatoid arthritis. Susceptibility to CIA in rodents is linked to the class II molecules of the major histocompatibility complex (MHC), and the immune response to CII is characterized by both the stimulation of collagen-specific T cells and the production of high titers of antibody specific for both the immunogen (heterologous CII) and the autoantigen (mouse CII). Histologically, murine CIA is characterized by an intense synovitis that corresponds precisely with the clinical onset of arthritis. Because of the pathological similarities between CIA and rheumatoid arthritis, the CIA model has been the subject of extensive investigation. Here, we describe the specifics for establishing the murine model of CIA, including specific requirements for the handling and preparation of the CII antigen, procedures for immunization, selection of susceptible mouse strains for study, and procedures for the evaluation and quantitation of the autoimmune arthritis. PMID- 15286394 TI - Mouse models of multiple sclerosis: experimental autoimmune encephalomyelitis and Theiler's virus-induced demyelinating disease. AB - Experimental autoimmune encephalomyelitis (EAE) and Theiler's murine encephalitis virus-induced demyelinating disease (TMEV-IDD) are two clinically relevant murine models of multiple sclerosis (MS). Like MS, both are characterized by mononuclear cell infiltrate into the central nervous system and demyelination. EAE is induced by either the administration of protein or peptide in adjuvant or by the adoptive transfer of encephalitogenic T-cell blasts into naive recipients. The relative merits of each of these protocols are compared. Depending on the type of question asked, different mouse strains and peptides are used. Different disease courses are observed with different strains and different peptides in active EAE. These variations are addressed, and grading of mice in EAE is discussed. In addition to EAE induction, useful references for other disease indicators, such as delayed type hypersensitivity, in vitro proliferation, and immunohistochemistry, are provided. TMEV-IDD is a useful model for understanding the potential viral etiology of MS. This chapter provides detailed information on the preparation of viral stocks and subsequent intracerebral infection of mice. In addition, virus plaque assay and disease assessment are discussed. Recombinant TMEV strains have been created for the study of molecular mimicry; these strains incorporate 30 various amino acid myelin epitopes within the leader region of TMEV. PMID- 15286393 TI - Proteoglycan aggrecan-induced arthritis: a murine autoimmune model of rheumatoid arthritis. AB - This chapter describes the major principals, methods, and immunization protocols for the induction of a systemic autoimmune arthritis in genetically susceptible murine strains. The model is called proteoglycan-induced arthritis (PGIA) because the antigenic/arthritogenic material is isolated from cartilage. This autoimmune systemic disease is induced by intraperitoneal immunization of either BALB/c or certain C3H colonies with cartilage proteoglycan, an abundant component in articular cartilage. The chapter presents (a) methodological details on how to purify cartilage proteoglycan aggrecan by cesium chloride gradient centrifugation; (b) substitution of this highly purified antigenic/arthritogenic material with a crude cartilage extract obtained from knee joint cartilages removed during joint replacement surgery; and (c) substitution of human cartilage proteoglycan with pig, dog, sheep, or bovine cartilage proteoglycans for arthritis induction. The cartilage proteoglycan aggrecan requires partial deglycosylation, and necessary materials, methods, and protocols are described. In addition, basic methods for measuring antigen-specific T-cell-dependent immune responses, antibody production, serum cytokine levels, and alternative solutions for adoptive transfers are also described. PMID- 15286395 TI - Experimental autoimmune encephalomyelitis. AB - Experimental autoimmune encephalomyelitis (EAE) is an inflammatory demyleinating disease of the central nervous system that is induced in laboratory animals by the generation of an immune response against myelin epitopes. It has been used as a prototype of Th1-driven, organ-specific autoimmunity and as a model for the human disease multiple sclerosis. This chapter describes two classic protocols for EAE induction (active immunization and adoptive transfer). Subheading 3.3. describes methods for rating clinical disease in symptomatic animals. Subheading 3.4. includes instructions for the isolation of mononuclear cells from the inflamed spinal cords from mice with EAE. PMID- 15286396 TI - Animal models of scleroderma. AB - Although no single animal model of systemic sclerosis (SSc) faithfully reproduces all features of the human disease, certain animal models that display some of the features of SSc are potentially useful as they may be helpful in gaining a better understanding of the pathogenesis of SSc as well as developing novel therapeutic interventions. This chapter gives the detailed description of the two most useful animal models of SSc: bleomycin-induced skin fibrosis and the sclerodermatous graft-vs-host disease in mice. It provides the methodology of the induction as well as the repertoire of the different approaches that can be used to investigate the skin fibrosis in these models, including histopathology, immunohistochemistry, dermal thickness, hydroxyproline content of the skin, and analysis of dermal cells by flow cytometry. PMID- 15286397 TI - Rodent models of experimental autoimmune uveitis. AB - The model of experimental autoimmune uveitis (EAU) in mice and in rats is described. EAU targets immunologically privileged retinal antigens and serves as a model of autoimmune uveitis in humans as well as a model for autoimmunity in a more general sense. EAU is a well-characterized, robust, and reproducible model that is easily followed and quantitated. It is inducible with synthetic peptides derived from retinal autoantigens in commonly available strains of rats and mice. The ability to induce EAU in various gene-manipulated, including HLA-transgenic, mouse strains makes the EAU model suitable for the study of basic mechanisms as well as in clinically relevant interventions. PMID- 15286398 TI - Autoimmune depigmentation following sensitization to melanoma antigens. AB - Generating an antitumor immune response can be thought of as eliciting an immune response to cells derived from self-tissue. As such, tumor immunity may result in autoimmunity. Melanoma patients undergoing immunotherapy often develop a form of autoimmune depigmentation referred to as vitiligo, in which T cells with antigenic specificity for pigmentation antigens destroy normal melanocytes. The models described in this chapter can be used to study immunity to melanoma antigens. These models employ a well-characterized pigmentation antigen relevant to melanoma and a common transplantable murine melanoma cell line. As more sophisticated approaches to cancer therapy are developed, models such as these may be key in understanding how immunity to self-antigens can be manipulated to elicit tumor immunity. PMID- 15286399 TI - Involvement of Leishmania donovani major surface glycoprotein gp63 in promastigote multiplication. AB - The major surface glycoprotein gp63 of the kinetoplastid protozoal parasite Leishmania is implicated as a ligand mediating uptake of the parasite into, and survival within, the host macrophage. By expressing gp63 antisense RNA from an episomal vector in L. donovani promastigotes, gp63-deficient transfectants were obtained. Reduction of the gp63 level resulted in increased generation times, altered cell morphology, accumulation of cells in the G2/M phase of the cell cycle, and increased numbers of binucleate cells with one or two kinetoplasts. Growth was stimulated, and the number of binucleate cells reduced, by addition to the culture of a bacterially expressed fusion protein containing the fibronectin like SRYD motif and the zinc-binding (metalloprotease) domain of gp63. These observations support an additional role of gp63 in promastigote multiplication; the fibronectin-like properties of gp63 may be important in this process PMID- 15286400 TI - Dibutyryl c-AMP as an inducer of sporidia formation: biochemical and antigenic changes during morphological differentiation of Karnal bunt (Tilletia indica) pathogen in axenic culture. AB - Effect of dibutyryl adenosine 3',5'-cyclic monophosphate (dbc-AMP), an analogue of c-AMP, was investigated on growth and morphological differentiation of Tilletia indica. Exponential growth was observed up to 21 days in both presence and absence of dbc-AMP; however, increasing concentration of dbc-AMP was deleterious to mycelial growth in liquid culture. A slow increase of mycelial biomass up to 21 days and decline at 30 days in the presence of 2.5 mM dbc-AMP was observed, therefore, this concentration was chosen in subsequent investigations. The inhibitory influence of dbc-AMP was further substantiated by decrease in soluble protein. The fungus on exposure to dbc-AMP experienced morphological differentiation from vegetative mycelial phase to sporogenous mycelial phase, and was induced to produce filiform sporidia. Use of quantitative ELISA further suggested that sporidia formation took more than 21 days in the presence of dbc-AMP. Variations of proteins during different stages of T. indica grown in the presence and absence of dbc-AMP suggested the expression of stage specific proteins or differential expression of proteins induced by dbc-AMP. The changes in expression of cell surface antigens as evidenced from decrease and increase binding of anti-mycelial and anti-sporidial antibodies in dbc-AMP treated culture by ELISA was further interpreted on the basis of morphological differentiation from mycelial to sporidial phase PMID- 15286401 TI - Identification of a hypothetical membrane protein interactor of ribosomal phosphoprotein P0. AB - The ribosomal phosphoprotein P0 of the human malarial parasite Plasmodium falciparum (PfP0) has been identified as a protective surface protein. In Drosophila, P0 protein functions in the nucleus. The ribosomal function of P0 is mediated at the stalk of the large ribosomal subunit at the GTPase centre, where the elongation factor eEF2 binds. The multiple roles of the P0 protein presumably occur through interactions with other proteins. To identify such interacting protein domains, a yeast two-hybrid screen was carried out. Out of a set of sixty clones isolated, twelve clones that interacted strongly with both PfP0 and the Saccharomyces cerevisiae P0 (ScP0) protein were analysed. These belonged to three broad classes: namely (i) ribosomal proteins; (ii) proteins involved in nucleotide binding; and (iii) hypothetical integral membrane proteins. One of the strongest interactors (clone 67B) mapped to the gene YFL034W which codes for a hypothetical integral membrane protein, and is conserved amongst several eukaryotic organisms. The insert of clone 67B was expressed as a recombinant protein, and immunoprecipitaion (IP) reaction with anti-P0 antibodies pulled down this protein along with PfP0 as well as ScP0 protein. Using deletion constructions, the domain of ScP0, which interacted with clone 67B, was mapped to 60-148 amino acids. It is envisaged that the surface localization of P0 protein may be mediated through interactions with putative YFL034W-like proteins in P. falciparum. PMID- 15286402 TI - Studies on middle silkgland proteins of cocoon colour sex-limited silkworm (Bombyx mori L.) using two-dimensional polyacrylamide gel electrophoresis. AB - Qualitative and quantitative differences in proteins expressed in the middle silkglands of male and female silkworm larvae that differ in silk colour were investigated by high resolution two-dimensional polyacrylamide gel electrophoresis (2-D PAGE), followed by computer assisted image analysis. About 1000 protein spots were resolved in both the sexes and most proteins were shown to be distributed in the area from 15 kDa to 70 kDa and pH 4-8. It was found that some proteins displayed higher expression in yellow cocoon, while two proteins were only expressed in female silkworm silkgland tissue through the comparison and analysis by two-D software. These proteins especially existed in female silkworm middle silkgland tissue of yellow cocoon. Furthermore, these proteins might be involved in the expression of cocoon colour phenotype PMID- 15286403 TI - Up-regulation of mu-opioid receptors in the spinal cord of morphine-tolerant rats. AB - Though morphine remains the most powerful drug for treating pain, its effectiveness is limited by the development of tolerance and dependence. The mechanism underlying development of tolerance to morphine is still poorly understood. One of the factors could be an alteration in the number of micro receptors within specific parts of the nervous system. However, reports on changes in the micro-opioid receptor density in the spinal cord after chronic morphine administration are conflicting. Most of the studies have used subcutaneously implanted morphine pellets to produce tolerance. However, it does not simulate clinical conditions, where it is more common to administer morphine at intervals, either by injections or orally. In the present study, rats were made tolerant to morphine by injecting increasing doses of morphine (10-50 mg/kg, subcutaneously) for five days. In vitro tissue autoradiography for localization of micro-receptor in the spinal cord was done using [3H]-DAMGO. As compared to the spinal cord of control rats, the spinal cord of tolerant rats showed an 18.8% increase or up-regulation in the density of micro-receptors in the superficial layers of the dorsal horn. This up-regulation of micro-receptors after morphine tolerance suggests that a fraction of the receptors have been rendered desensitized, which in turn could lead to tolerance PMID- 15286404 TI - Analysis of human chorionic gonadotropin-monoclonal antibody interaction in BIAcore. AB - Kinetic studies of macromolecular ligant-ligate interaction have generated ample interest since the advent of plasmon resonance based instruments like BIAcore. Most of the studies reported in literature assume a simple 1:1 Langmuir binding and complete reversibility of the system. However we observed that in a high affinity antigen-antibody system [human chlorionic gonadotropin-monoclonal antibody (hCG-mAb)] dissociation in insignificant and the sensogram data cannot be used to measure the equilibrium and kinetic parameters. At low concentrations of mAb the complete sensogram could be fitted to a single exponential. Interestingly we found that at higher mAb concentrations, the binding data did not conform to a sample biomolecular model. Instead, the data fitted a two-step model, which may be because of surface heterogeneity of affinity sites. In this paper, we report on the global fit of the sensograms. We have developed a method by which a single two-minute sensogram can be used in high affinity systems to measure the association rate constant of the reaction and the functional capacity of the ligand (hCG) immobilized on the chip. We provide a rational explanation for the discrepancies generally observed in most of the BIAcore sensograms. PMID- 15286405 TI - An optimized gossypol high-performance liquid chromatography assay and its application in evaluation of different gland genotypes of cotton. AB - A comparative study on gossypol content of various genetic types of pigment glands of cotton varieties was conducted through an optimized high-performance liquid chromatography (HPLC) on a C18 column (4.6 mm x 250 mm, 5 microm particle) with methanol-0.5% acetic acid aqueous solution, 90 : 10 (v/v), as mobile phase, at a flow rate of 0.8 ml/min and UV detection at 254 nm. The method was shown to be highly reproducible, with precision [as relative standard deviation (RSD)] and accuracy [as relative mean error (RME)] < 10%, both intra-day and inter-day. Absolute recoveries were > 94%. The results revealed major differences among the different gland varieties or species of cotton, including the special and ordinary glandless and glanded Gossypium hirsutum, G. barbadense, and displayed the precious resources of different glands of extraordinary cotton. PMID- 15286406 TI - Protective effects of sodium orthovanadate in diabetic reticulocytes and ageing red blood cells of Wistar rats. AB - The reticulocytes and the ageing red blood cells (RBCs) namely young (Y), middle aged (M) and old RBCs (O) of female Wistar rats from different groups such as control animals (C), controls treated with vanadate (C + V), alloxan-induced diabetic (D), diabetic-treated with insulin (D + I) and vanadate (D + V), were fractionated on a percoll/BSA gradient. The following enzymes were measured - hexokinase (HK), glutathione peroxidase (GSH-Px), glutathione reductase (GSSG-R), glutathione-s-transferase (GST), alanine aminotransferase (AlaAT), aspartate aminotransferase (AsAT) and arginase in the hemolysates of all the RBCs fractions. Decreases in the activity of HK and AsAT by about 70%, arginase and GSH-Px by 30% in old RBCs were observed in comparison to reticulocytes of control animals. Increases in the activity of GSSG-R by 86%, AlaAT by more than 400% and GST by 70% were observed in old RBCs in comparison to reticulocytes of control animals. Alloxan diabetic animals showed a further decrease in the activities of HK in Y RBCs by 37%, M RBCs by 39% and O RBCs by 32%, GSH-Px activity in Y RBCs by 13%, M RBCs by 20% and O RBCs by 33% and GST activity in Y RBCs by 14%, M RBCs by 42% and O RBCs by 60% in comparison to their corresponding cells of control animals. An increase in the activity of all the enzymes studied was also observed in reticulocytes of diabetic animals in comparison to reticulocytes of controls. The GSSG-R activity was found to be increased in Y RBCs by 49%, M RBCs by 67% and O RBCs by 64% as compared to the corresponding age-matched cells of control animals. The activity of arginase also decreased in Y RBCs by about10%, M RBCs by 20% and O RBCs by 30% in comparison to the age-matched cells of control animals. A decrease in the activity of AsAT in Y and M RBCs by 30%, and O RBCs by 25% was observed in diabetic animals in comparison to the age-matched cells of control animals. The activity of AlaAT was found to be decreased by more than 10% in Y and M RBCs and 25% in O RBCs of diabetic animals in comparison to the age-matched cells of control animals. Insulin administration to diabetic animals reversed the altered enzyme activity to control values. Vanadate treatment also reversed the enzyme levels except for that of GST in old cells. PMID- 15286408 TI - Golgi analysis of tangential neurons in the lobula plate of Drosophila melanogaster. AB - The lobula plate (LP), which is the third order optic neuropil of flies, houses wide-field neurons which are exquisitely sensitive to motion. Among Diptera, motion-sensitive neurons of larger flies have been studied at the anatomical and physiological levels. However, the neurons of Drosophila lobula plate are relatively less explored. As Drosophila permits a genetic analysis of neural functions, we have analysed the organization of lobula plate of Drosophila melanogaster. Neurons belonging to eight anatomical classes have been observed in the present study. Three neurons of the horizontal system (HS) have been visualized. The HS north (HSN) neuron, occupying the dorsal lobula plate is stunted in its geometry compared to that of larger flies. Associated with the HS neurons, thinner horizontal elements known as h-cells have also been visualized in the present study. Five of the six known neurons of the vertical system (VS) have been visualized. Three additional neurons in the proximal LP comparable in anatomy to VS system have been stained. We have termed them as additional VS (AVS)-like neurons. Three thinner tangential cells that are comparable to VS neurons, which are elements of twin vertical system (tvs); and two cells with wide dendritic fields comparable to CH neurons of Diptera have been also observed. Neurons comparable to VS cells but with 'tufted' dendrites have been stained. The HSN and VS1-VS2 neurons are dorsally stunted. This is possibly due to the shape of the compound eye of Drosophila which is reduced in the fronto dorsal region as compared to larger flies. PMID- 15286407 TI - Effects of sodium-orthovanadate and Trigonella foenum-graecum seeds on hepatic and renal lipogenic enzymes and lipid profile during alloxan diabetes. AB - Sodium-orthovanadate (SOV) and seed powder of Trigonella foenum graecum Linn. (common name: fenugreek, family: Fabaceae) (TSP) besides being potential hypoglycemic agents have also been shown to ameliorate altered lipid metabolism during diabetes. This study evaluates the short-term effect of oral administration of SOV and TSP separately and in concert (for 21 days) on total lipid profile and lipogenic enzymes in tissues of alloxan diabetic rats. Diabetic rats showed 4-fold increase in blood glucose. The level of total lipids, triglycerides and total cholesterol in blood serum increased significantly during diabetes. During diabetes the level of total lipids increased significantly (P < 0.001) in liver and in kidney by 48% and 55%, respectively, compared to control. Triglycerides level increased by 32% (P < 0.01) in liver and by 51% (P < 0.005) in kidney, respectively, compared to control. Total cholesterol level also increased significantly in both liver and kidney (P < 0.01 and P < 0.001, respectively). The activities of NADP-linked enzymes; namely glucose-6-phosphate dehydrogenase (G6PDH), malic enzyme (ME), isocitrate dehydrogenase (ICDH), and the activities of lipogenic enzymes namely ATP-citrate lyase (ATP-CL) and fatty acid synthase (FAS) were decreased significantly in liver and increased in kidney during diabetes as compared to control. SOV and TSP administration to diabetic animals prevented the development of hyperglycemia and alteration in lipid profile in plasma and tissues and maintained it near normal. Maximum prevention was observed in the combined treatment with lower dose of SOV (0.2%) after 21 days. We are presenting for the first time effectiveness of combined treatment of SOV and TSP in amelioration of altered lipid metabolism during experimental type I diabetes. PMID- 15286409 TI - Factors influencing offspring traits in the oviparous multi-clutched lizard, Calotes versicolor (Agamidae). AB - The determinants of offspring size and number in the tropical oviparous multi clutched lizard, Calotes versicolor, were examined using both univariate and multivariate (path) analyses. In C. versicolor maternal snout-vent length (SVL) and body condition influence clutch mass and clutch size but have no significant influence on offspring size. The positive effect of maternal SVL and body condition on offspring number is counterbalanced by a negative effect of breeding time on egg mass. In fact, breeding time directly influences the offspring body mass and condition through variation in the egg mass. There is a trade-off between offspring mass and condition with offspring number, and breeding time influences both. Offspring hatched from the eggs of early (May-June) or mid (July August) breeding periods invariably show lower mass and condition than those hatched from the eggs of late breeding season (September-October). Yet, there is no variation in offspring SVL among early, mid and late clutches. Thus, in C. versicolor offspring SVL is optimized while body mass and condition are not optimized. PMID- 15286410 TI - Adaptive response and split-dose effect of radiation on the survival of mice. AB - Although the importance of radiation-induced adaptive response has been recognized in human health, risk assessment and clinical application, the phenomenon has not been understood well in terms of survival of animals. To examine this aspect Swiss albino mice were irradiated with different doses (2-10 Gy) at 0.015 Gy/s dose rate and observed on a regular basis for 30 days. Since almost 50% lethality was seen with 8 Gy, it was selected as the challenging dose for further studies. Irradiation of mice with conditioning doses (0.25 or 0.5 Gy) and subsequent exposure to 8 Gy caused significant increase in the survival of mice compared to irradiated control. The splitting of challenging dose did not influence the efficiency of conditioning doses (0.25 Gy and 0.5 Gy) to induce an adaptive response. However conditioning doses given in fractions (0.25 Gy + 0.25 Gy) or (0.5 Gy + 0.5 Gy) were able to modulate the response of challenging dose of 8 Gy. These results clearly showed the occurrence of adaptive response in terms of survival of animals. The conditioning dose given in small fractions seemed to be more effective. The findings have been discussed from a mechanistic point of view. The possible biological implications, potential medical benefits, uncertainties and controversies related to adaptive response have also been addressed PMID- 15286411 TI - On the dependence of speciation rates on species abundance and characteristic population size. AB - The question of the potential importance for speciation of large/small population sizes remains open. We compare speciation rates in twelve major taxonomic groups that differ by twenty orders of magnitude in characteristic species abundance (global population number). It is observed that the twenty orders of magnitude's difference in species abundances scales to less than two orders of magnitude's difference in speciation rates. As far as species abundance largely determines the rate of generation of intraspecific endogenous genetic variation, the result obtained suggests that the latter rate is not a limiting factor for speciation. Furthermore, the observed approximate constancy of speciation rates in different taxa cannot be accounted for by assuming a neutral or nearly neutral molecular clock in subdivided populations. Neutral fixation is only relevant in sufficiently small populations with 4N(e)v < 1, which appears an unrealistic condition for many taxa of the smaller organisms. Further research is clearly needed to reveal the mechanisms that could equate the evolutionary pace in taxa with dramatically different population sizes PMID- 15286412 TI - A novel potassium deficiency-induced stimulon in Anabaena torulosa. AB - Potassium deficiency enhanced the synthesis of fifteen proteins in the nitrogen fixing cyanobacterium Anabaena torulosa and of nine proteins in Escherichia coli. These were termed potassium deficiency-induced proteins or PDPs and constitute hitherto unknown potassium deficiency-induced stimulons. Potassium deficiency also enhanced the synthesis of certain osmotic stress-induced proteins. Addition of K+ repressed the synthesis of a majority of the osmotic stress-induced proteins and of PDPs in these bacteria. These proteins contrast with the dinitrogenase reductase of A. torulosa and the glycine betaine-binding protein of E. coli, both of which were osmo-induced to a higher level in potassium supplemented conditions. The data demonstrate the occurrence of novel potassium deficiency-induced stimulons and a wider role of K+ in regulation of gene expression and stress responses in bacteria PMID- 15286413 TI - KCTCCA, a peptide-based facilitator for bioelectrochemistry. AB - Electrochemical and scanning tunnelling microscopy (STM) studies have been carried out to investigate the suitability of the hexapeptide KCTCCA as a facilitator for bioelectrochemistry. The stable, quasi-reversible electrochemical response of cytochrome b562 on a KCTCCA modified gold electrode and the high degree of surface coverage of KCTCCA on gold (111), as observed by STM, indicate applicability of the molecule as an electrochemical facilitator PMID- 15286414 TI - Microbial acetate oxidation in horizontal rotating tubular bioreactor. AB - The aim of this work was to investigate the possibility of conducting a continuous aerobic bioprocess in a horizontal rotating tubular bioreactor (HRTB). Aerobic oxidation of acetate by the action of a mixed microbial culture was chosen as a model process. The microbial culture was not only grown in a suspension but also in the form of a biofilm on the interior surface of HRTB. Efficiency of the bioprocess was monitored by determination of the acetate concentration and chemical oxygen demand (COD). While acetate inlet concentration and feeding rate influenced efficiency of acetate oxidation, the bioreactor rotation speed did not influence the bioprocess dynamics significantly. Gradients of acetate concentration and pH along HRTB were more pronounced at lower feeding rates. Volumetric load of acetate was proved to be the most significant parameter. High volumetric loads (above 2 g acetate l-1 h-1) gave poor acetate oxidation efficiency (only 17 to 50%). When the volumetric load was in the range of 0.60-1.75 g acetate l-1 h-1, acetate oxidation efficiency was 50-75%. At lower volumetric loads (0.14-0.58 g acetate l-1 h-1), complete acetate consumption was achieved. On the basis of the obtained results, it can be concluded that HRTB is suitable for conducting aerobic continuous bioprocesses. PMID- 15286415 TI - Effects of anisotonicity on pentose-phosphate pathway, oxidized glutathione release and t-butylhydroperoxide-induced oxidative stress in the perfused liver of air-breathing catfish, Clarias batrachus. AB - Both hypotonic exposure (185 mOsmol/l) and infusion of glutamine plus glycine (2 mmol/l each) along with the isotonic medium caused a significant increase of 14CO2 production from [1-14C]glucose by 110 and 70%, respectively, from the basal level of 18.4 +/- 1.2 nmol/g liver/min from the perfused liver of Clarias batrachus. Conversely, hypertonic exposure (345 mOsmol/l) caused significant decrease of 14CO2 production from [1-14C]glucose by 34%. 14CO2 production from [6 14C]glucose was largely unaffected by anisotonicity. The steady-state release of oxidized glutathione (GSSG) into bile was 1.18 +/- 0.09 nmol/g liver/min, which was reduced significantly by 36% and 34%, respectively, during hypotonic exposure and amino acid-induced cell swelling, and increased by 34% during hypertonic exposure. The effects of anisotonicity on 14CO2 production from [1-14C]glucose and biliary GSSG release were also observed in the presence of t butylhydroperoxide (50 mmol/l). The oxidative stress-induced cell injury, caused due to infusion of t-butylhydroperoxide, was measured as the amount of lactate dehydrogenase (LDH) leakage into the effluent from the perfused liver; this was found to be affected by anisotonicity. Hypotonic exposure caused significant decrease of LDH release and hypertonic exposure caused significant increase of LDH release from the perfused liver. The data suggest that hypotonically-induced as well as amino acid-induced cell swelling stimulates flux through the pentose phosphate pathway and decreases loss of GSSG under condition of mild oxidative stress; hypotonically swollen cells are less prone to hydroperoxide-induced LDH release than hypertonically shrunken cells, thus suggesting that cell swelling may exert beneficial effects during early stages of oxidative cell injury probably due to swelling-induced alterations in hepatic metabolism. PMID- 15286416 TI - Acetylcholinesterase in central vocal control nuclei of the zebra finch (Taeniopygia guttata). AB - The distribution of acetylcholinesterase (AChE) in the central vocal control nuclei of the zebra finch was studied using enzyme histochemistry. AChE fibres and cells are intensely labelled in the forebrain nucleus area X, strongly labelled in high vocal centre (HVC) perikarya, and moderately to lightly labelled in the somata and neuropil of vocal control nuclei robust nucleus of arcopallium (RA), medial magnocellular nucleus of the anterior nidopallium (MMAN) and lateral magnocellular nucleus of the anterior nidopallium (LMAN). The identified sites of cholinergic and/or cholinoceptive neurons are similar to the cholinergic presence in vocal control regions of other songbirds such as the song sparrow, starling and another genus of the zebra finch (Poephila guttata), and to a certain extent in parallel vocal control regions in vocalizing birds such as the budgerigar. AChE presence in the vocal control system suggests innervation by either afferent projecting cholinergic systems and/or local circuit cholinergic neurons. Co occurrence with choline acetyltransferase (ChAT) indicates efferent cholinergic projections. The cholinergic presence in parts of the zebra finch vocal control system, such as the area X, that is also intricately wired with parts of the basal ganglia, the descending fibre tracts and brain stem nuclei could underlie this circuitry's involvement in sensory processing and motor control of song PMID- 15286417 TI - Foraging behaviour in tadpoles of the bronze frog Rana temporalis: experimental evidence for the ideal free distribution. AB - The ability of bronze frog Rana temporalis tadpoles (pure or mixed parental lines) to assess the profitability of food habitats and distribute themselves accordingly was tested experimentally using a rectangular choice tank with a non- continuous input design. Food (boiled spinach) was placed at two opposite ends of the choice tank in a desired ratio (1 : 1, 1 : 2 or 1 : 4) to create habitat A and B. The tadpoles in Gosner stage 28-33, pre-starved for 24 h, were introduced in an open ended mesh cylinder placed in the center of the choice tank, held for 4 min (for acclimation) and then released to allow free movement and habitat selection. The number of tadpoles foraging at each habitat was recorded at 10, 15, 20, 25 and 30 min time intervals. The actual suitability, Si (the food available in a habitat after colonization of tadpoles) of each habitat was obtained from the equation Si = Bi - fi (di) where Bi is basic suitability (amount of food provided at each habitat before release of tadpoles), fi is the rate of depletion of food (lowering effect) with introduction of each tadpole, and di is the density of tadpoles in habitat i. The expected number of tadpoles at each habitat was derived from the actual suitability. With no food in the choice tank, movement of the tadpoles in the test arena was random indicating no bias towards any end of the choice tank or the procedure. In tests with a 1 : 1 food ratio, the observed ratio of tadpoles (11.71 : 12.28) was comparable with the expected 12 : 12 ratio. The observed number of tadpoles in the habitats with a 1 : 2 food ratio was 8.71 : 15.29 and 7.87 : 16.13 for pure and mixed parental lines respectively. In both cases, the observed ratios were close to the expected values (7 : 17). Likewise, in experiments with a 1 : 4 food ratio, the observed number of tadpoles in the two habitats (10.78 : 37.22) did not differ significantly from the expected ratio of 7 : 41. In all tests, the number of R. temporalis tadpoles matched ideally with habitat profitability (undermatching index K approximate, equals 1). The study shows that tadpoles of the bronze frog exhibit an ideal free distribution while foraging regardless of whether they are siblings or non-siblings in a group, which correlates well with their group living strategy in nature PMID- 15286418 TI - Severity, affect, family and environment (SAFE) approach to evaluate chronic pelvic pain in adolescent girls. AB - BACKGROUND: Pelvic pain is common in adolescent girls in day-to-day practice. Severity, Affect, Family and Environment (SAFE) is a recent interview strategy to approach these patients and their families. AIM: 1. To find the prevalence of pelvic pain in adolescent girls. 2. To find out the feasibility and acceptability of "SAFE" approach in evaluating chronic pelvic pain in adolescent girls. SETTINGS & DESIGN: 200 adolescent girls aged 13-23 years were selected at random from school and colleges nearby hospital campus. METHOD: A questionnaire method was adopted. Adolescent girls selected were divided into two groups, group I (with pain) and group II (without pain). STATISTICAL ANALYSIS: It was performed using chi-square and Fischer tests with significance of p value being taken at 0.05. RESULTS: Out of 200 adolescent girls selected for interview, prevalence of pelvic pain was found to be 37.0%. Dysmenorrhoea was also found to have significant relationship with pelvic pain. Depression and anxiety, school absences and loss of weight were found to have significant association with pain. Pelvic pain was found to be more in adolescent girls with large family size (> 4 members), single parent, both parents working, ongoing marital problems at home and was less reported when there was good parent-child communication about sex and when the teenager was well prepared for menarche by the parents and the association with the above mentioned factors was found to be significant. CONCLUSION: "SAFE" approach contributes in identifying health problems in adolescent girls with chronic pelvic pain. PMID- 15286419 TI - Spectrum of anti-neutrophil cytoplasmic antibodies in patients with pulmonary tuberculosis overlaps with that of Wegener's granulomatosis. AB - BACKGROUND AND OBJECTIVES: Mycobacterial infections are known to induce the development of autoantibodies and a few of these antibodies are also known to be diagnostic markers for some other diseases and it is uncertain whether these autoantibodies play a role in the pathogenesis of autoimmune disorders. This study was undertaken to determine the prevalence of autoantibodies like anti neutrophil cytoplasmic antibodies (ANCA), anti-nuclear antibodies (ANA), anti double stranded antibodies (anti-dsDNA) and anti-histone antibodies (AHA)in pulmonary Tuberculosis. MATERIALS & METHODS: Seventy consecutive pulmonary TB patients, 30 patients of interstitial lung disease and 100 normal individuals were studied. ANCA and ANA were detected by indirect immunofluorescence test (IIF). Anti-dsDNA and AHA were tested by ELISA. RESULTS: ANCA was detected in 30% cases, and of these 52.4% showed perinuclear pattern (p-ANCA), 38.1% cytoplasmic (c-ANCA) and 9.5% showed an "atypical" pattern. ANCA specificities by ELISA revealed that, 47.6% had anti-Myeloperoxidase (anti-MPO), 28.6% had anti Proteinase3 (anti-PR3) and 19.1% had anti-Lactoferrin (anti-LF) antibodies. ANA and AHA were present in 24.3% and 21.4% cases respectively whereas anti-ds DNA antibodies were absent. Normal controls showed 4% and 2% positivity for ANA and ANCA whereas disease control group of ILD showed 7% of ANA and ANCA posititivy. CONCLUSION: The presence of autoantibodies in TB patients could have a multifactorial etiology. Clinically relevant is the presence of anti-PR3 antibodies. This finding along with pulmonary and renal manifestations could lead to a false diagnosis of Wegener's granulomatosis or vice versa because these autoantibodies may be present in both diseases. PMID- 15286420 TI - A school-based intervention to teach 3-4 grades children about healthy heart; the Persian Gulf healthy heart project. AB - BACKGROUND: Cardiovascular health promotion in children has the potential to reduce the risk of atherosclerosis in both the individual child and the population at large. It thus seems eminently reasonable to initiate healthful lifestyle training in childhood to promote improved cardiovascular health in adult life. AIMS: To test the hypothesis that a year long, classroom-based education for the third and fourth graders could change their knowledge scores about healthy heart. SETTINGS AND DESIGN: A randomized, controlled trial in elementary schools of Bushehr/Iran. METHODS AND MATERIALS: A total of 14 elementary schools, categorized by socioeconomic types and male and female setting were selected and randomized into control or intervention groups. Subjects were 1128 third and fourth graders, aged 9 to 10 years (49.1% boys and 50.9% girls). Over a course of 8 weeks, health educators and sport teachers of the elementary schools presented two hours sessions per week on heart function, nutrition, and exercise for healthy heart and living tobacco free for the intervention group. The education program was based on HeartPower! Program, an American Heart Association program. STATISTICAL ANALYSIS: Mann-Whitney U test and Wilcoxon matched-pairs signed rank test and Bonferroni correction for the two pair wise comparisons were used. RESULTS: Total heart knowledge at posttest was 25% correct higher in the intervention than in the control group (p< 0.001). Difference in means of total healthy heart knowledge scores between control and intervention group increased from 1.43 points in baseline to 4.02 points in posttest (p< 0.001). CONCLUSION: It can be concluded that the classroom-based cardiovascular health promotion had a significant effect on the heart healthy knowledge. Therefore, schools provide an excellent setting for introducing comprehensive healthy heart education and promotion of cardiovascular health to the general population. PMID- 15286421 TI - Short tandem repeat technology has diverse applications: individual identification, phylogenetic reconstruction and chimerism based post haematopoietic stem cell transplantation graft monitoring. AB - BACKGROUND: Short Tandem Repeat (STR) loci are widely considered to be effective for variety of applications including forensic applications, phylogenetic reconstruction and chimerism based post Haematopoietic Stem Cell Transplantation (HSCT) graft monitoring. For each application, specific sets of STR loci are used. AIMS: In the present study, we have attempted to use same set of STR loci for varied purposes based on their efficacy and informativity. SETTINGS AND DESIGN: Population and patient based study. MATERIALS AND METHODS: We have analyzed 5 STR loci--vWA, Tho1, FES, F13 and TPOX in 1000 North Indians. All five markers were also analyzed for chimerism based graft monitoring after HSCT in 42 HLA matched pair of patient-donor to predict the outcome of transplantation. STATISTICAL ANALYSIS: The analysis was done for Hardy Weinberg equilibrium (HWE), Heterozygosity, Polymorphism information content (PIC) and Power of Exclusion and Phylogenetic assessment. RESULTS AND CONCLUSIONS: High allelic variability in term of Heterozygosity (0.68-0.76), PIC (0.66-0.74) and high Power of exclusion (0.28-0.38) indicating high forensic utility. The ensuing PC plots finely resolved three basal clusters corresponding to three geo-ethnic groups of African, Orientals, and Caucasians. In post HSCT chimerism analysis, it was found that together these markers were informative in 38 pairs (98%) and were able to predict the chimerism status successfully. There is a possibility that these STR loci along with forensic and phylogenetic importance, can predict the outcome of HSCT successfully. PMID- 15286422 TI - Is today's male population really less fertile? Declining semen quality--a global phenomenon? PMID- 15286423 TI - Seroprevalence of HIV, HBV, HCV and syphilis in blood donors. PMID- 15286424 TI - Pattern of use of tobacco based products in urban area of Chandigarh. PMID- 15286425 TI - Megaloblastic anemia--Part II. PMID- 15286426 TI - Involvement of TRPM7 in cell growth as a spontaneously activated Ca2+ entry pathway in human retinoblastoma cells. AB - We investigated the possible involvement of the melastatin family protein TRPM7 in Ca(2+)-mediated proliferative control of human retinoblastoma (RB) cells. The growth of RB cell was facilitated by elevating the extracellular Ca(2+) concentration with a parallel increase in the magnitude of spontaneous Ca(2+) influx. Under nystatin-perforated voltage-clamp, RB cells exhibited an outward rectifying, spontaneous cation current (I(spont)) having Ca(2+)/Mg(2+)-inhibited but -permeating properties. Various cation channel blockers inhibiting I(spont) (Gd(3+), La(3+), LOE908, 2-APB) suppressed the spontaneous Ca(2+) influx and decelerated the growth of RB cells with similar efficacies. Excision of the RB cell membrane (inside-out) into MgATP-free solution induced a 70pS single channel activity, which was effectively inhibited by millimolar concentrations of Mg(2+) or MgATP. RT-PCR and immunocytochemical experiments revealed the expression of TRPM7 mRNA and protein in RB cells, and heterologous expression of TRPM7 in HEK293 cells reproduced the key features of I(spont). In contrast, elimination of this protein from RB cells by siRNA silencing markedly reduced I(spont) density and the magnitude of spontaneous Ca(2+) influx, which was paralleled by decreased TRPM7 immunoreactivity, decelerated cell proliferation, and retarded G(1)/S cell cycle progression. These results suggest a significant regulatory role of TRPM7 for RB cell proliferation as a spontaneously activated Ca(2+) influx pathway. PMID- 15286427 TI - Uridine 5'-triphosphate stimulates alveolar fluid clearance in the isolated rat lungs. AB - Uridine 5'-triphosphate (UTP) increases chloride secretion followed by fluid movement into the proximal airspaces. However, little is known about whether UTP affects fluid movement in the distal airspaces. We studied the effect of UTP on basal and stimulated alveolar fluid clearance in the isolated rat lungs. Isosmotic 5% albumin solution was instilled into the alveolar spaces of isolated rat lungs, which were then inflated with 100% oxygen at an airway pressure of 7 cmH(2)O. Alveolar fluid clearance was measured by the progressive increase in albumin concentrations over 1 h. Although UTP (10(-9) - 10(-6) M) did not increase alveolar fluid clearance, UTP (10(-5) - 10(-3) M) and isoproterenol (10( 5) M), a beta-adrenergic agonist, increased alveolar fluid clearance by 40% and 120% of the basal values, respectively. A combined treatment of UTP (10(-4) M, 10(-3) M) and isoproterenol increased alveolar fluid clearance by 280% of the basal value. The effects of UTP in the presence and absence of isoproterenol were abolished by blockers of a P2 purinoceptor and chloride channels. These results indicate that UTP stimulates alveolar fluid clearance in the distal airspaces of rat lungs. PMID- 15286428 TI - Histamine H1 receptor down-regulation mediated by M3 muscarinic acetylcholine receptor subtype. AB - Heterologous down-regulation of histamine H(1) receptor (H1R) mediated by muscarinic acetylcholine receptor subtype was investigated using five kinds of Chinese hamster ovary (CHO) cells stably co-expressing the human H1R and one of the five (M(1) - M(5)) muscarinic acetylcholine receptors, CHO-H1/M1, CHO-H1/M2, CHO-H1/M3, CHO-H1/M4, and CHO-H1/M5 cells. Among the CHO-H1/M1, CHO-H1/M3, and CHO-H1/M5 cells, carbachol treatment of the CHO-H1/M3 cells time-dependently led to remarkable down-regulation of the H1R to 60% of the control level. In contrast, stimulation of CHO-H1/M1 cells by carbachol induced negligible effect on the down-regulation. Stimulation of CHO-H1/M5 cells by carbachol induced significant but only small H1R down-regulation. M(2) and M(4) muscarinic receptors showed negligible effect on the down-regulation. H1R-mediated accumulation of inositol phosphates in CHO-H1/M3 cells with long-term expose to carbachol was decreased to 60% compared with non-treated cells. Heterologous phosphorylation of H1R was induced by the stimulation of each muscarinic receptor. H1R was phosphorylated by about twofold from the basal level through five subtypes of muscarinic receptor. The M(3) muscarinic receptor-mediated phosphorylation of H1R was reversed by the inhibition of protein kinase C. In the present study we demonstrated that the M(3) muscarinic acetylcholine receptor mediated remarkable down-regulation of the H1R with decreased receptor signaling. PMID- 15286429 TI - Apafant, a potent platelet-activating factor antagonist, blocks eosinophil activation and is effective in the chronic phase of experimental allergic conjunctivitis in guinea pigs. AB - Platelet-activating factor (PAF) plays important roles in allergic reactions. In particular, there are many concerns about PAF, eosinophils, and the chronicity of allergic diseases. The purpose of the present studies is to elucidate the role of PAF in eosinophil activation at conjunctiva and to confirm the efficacy of Apafant (a potent PAF antagonist) ophthalmic solution in chronic experimental allergic conjunctivitis. Guinea pigs were actively immunized and allergic conjunctivitis was induced by repetitive instillation of 2.5% ovalbumin. PAF solution was topically applied and eosinophil activation was assessed by measuring the eosinophil peroxidase (EPO) activity in the tear fluid. Itch scratching episodes and clinical symptoms scores were evaluated in the repetitive challenge conjunctivitis. From the instillation of PAF solution into guinea pig eyes, which were in a state of chronic allergic conjunctivitis, a significant increase in EPO activity was observed, and this increase was inhibited by pre treatment with Apafant. In the repetitive challenge model, the animals treated with Apafant ophthalmic solution showed a significant reduction of clinical symptoms and the itch-scratch response in both the first and the second challenges. PAF has an activity, that induces mediator release from eosinophils in the conjunctival tissues and may be involved in the chronic phase of allergic conjunctivitis. PMID- 15286430 TI - Vanadate activates Rho A translocation in association with contracting effects in ileal longitudinal smooth muscle of guinea pig. AB - We characterized the effects of vanadate, an inhibitor of tyrosine phosphatase, on the tension, the level of myosin light chain (MLC) phosphorylation, and Rho A activation in intact ileal longitudinal smooth muscle of the guinea pig to study the role of tyrosine phosphorylation in contraction signaling. Vanadate exerted a sustained contraction with a slow onset of tension development, in a concentration-dependent manner. The contractile effects of vanadate were accompanied by increases in the level of MLC phosphorylation. The tyrosine kinase inhibitor genistein; the MLC kinase inhibitor 1-(5-chloronaphthalene-1-sulfonyl) 1H-hexahydro-1,4-diazepine hydrochloride (ML-9); and the Rho kinase inhibitor (+) (R)-trans-4-(1-aminoethyl)-N-(4-pyridyl) cyclohexanecarboxamide dihydrochloride, monohydrate (Y-27632) inhibited the vanadate-induced contraction and MLC phosphorylation. Vanadate caused Rho A translocation from the cytosol to the membrane fraction, which was inhibited by genistein, but not by ML-9 and Y-27632. These data indicate that vanadate induces Rho A activation probably via protein tyrosine phosphorylation and the subsequent contraction through increases in the level of MLC phosphorylation. PMID- 15286431 TI - The regulation of human beta-defensin 2 by the ETS transcription factor MEF (myeloid Elf-1-like factor) is enhanced by promyelocytic leukemia protein. AB - The human beta-defensin 2 (HBD 2) is a potent anti-bacterial peptide with a wide spectrum of activity. MEF (myeloid elf-1-like factor) or Elf4, a member of the ETS transcription factor family, has been shown to up-regulate the basal expression of the HBD 2 gene in epithelial cells. The mammalian cell nucleus is organized into distinct nuclear domains, one of which is the PML (promyelocytic leukemia) nuclear body involved in the regulation of transcription. Here, we show that PML stimulated MEF transcriptional activity, resulting in the up-regulation of endogenous HBD 2 expression. PMID- 15286432 TI - Characterization of the trafficking pathway of cystic fibrosis transmembrane conductance regulator in baby hamster kidney cells. AB - To examine the unknown trafficking pathway of the cystic fibrosis transmembrane conductance regulator (CFTR) from the endoplasmic reticulum (ER), we utilized baby hamster kidney cells stably expressing CFTR fused with green fluorescent protein. CFTR trafficking from the ER was visualized and analyzed by immunocytochemical analyses. Here we show that CFTR was exported from the ER to the cis-Golgi and early endosome, suggesting that CFTR transport in the early secretory pathway may utilize a non-conventional pathway. This CFTR trafficking pathway may be a target for pharmacological modulation that selectively stimulates CFTR transport. PMID- 15286433 TI - Polymorphisms in interleukin-1 beta and Interleukin-1 receptor antagonist genes are associated with kidney failure in Korean patients with type 2 diabetes mellitus. AB - BACKGROUND/AIM: Cytokines play an important role in the pathogenesis of kidney diseases. The aim of the study was to investigate the impact of interleukin (IL) 1 cluster genes on diabetic nephropathy in Korean patients with type 2 diabetes mellitus (DM). METHODS: We investigated -511 C/T polymorphism of IL-1 beta and tandem repeat polymorphism in intron 2 of IL-1 receptor antagonist in type 2 DM patients with end-stage kidney failure as compared with patients without nephropathy. RESULTS: The IL1B2 allele was found more frequently in patients with kidney failure than in controls (57.4 vs. 46.1%, p < 0.05). An excessive homozygous carriage of IL1B2 was found in patients with kidney failure when compared with controls (30.5 vs. 18.3%, p < 0.05). The allelic frequency of IL1RN*2 was also higher in cases than in controls without nephropathy (8.4 vs. 2.8 %, p < 0.05). The carriage rate of IL1RN*2 was significantly associated with an increased risk of kidney failure (15.8 vs. 5.6%; OR 3.19, 95% CI 1.24-8.17). The risk of kidney failure was highest in those carrying both IL1RN*2 and IL1B2 (OR 3.90, 95% CI 1.34-11.40). CONCLUSION: IL1B2 and IL1RN*2 genotypes of the IL-1 cluster genes are associated with diabetic nephropathy in Korean patients with type 2 DM. PMID- 15286434 TI - Stress and acute biliary pancreatitis. PMID- 15286435 TI - Influence of stress in acute pancreatitis and correlation with stress-induced gastric ulcer. AB - BACKGROUND AND AIMS: In the general adaptation syndrome, gastric lesions are the first manifestation of stress. We hypothesized that acute pancreatitis (AP), an inflammatory acute disease, will be exacerbated if unchained following stress. Visceral hypersensitivity will be enhanced due to catecholaminergic discharges leading to an over-induction of the intrapancreatic cholinergic tone with increased response of the pancreocyte to cholecystokinin (CCK). Our aim was to investigate the influence of stress before AP on the later AP, and the effect of AP on underlying diseases such as gastric ulceration. METHODS: The model of stress induced by restraint was followed by the bilio-pancreatic duct outlet exclusion closed duodenal loops model. The effect of autonomous arc reflex (AAR) interruption by anesthetics after stress but before AP was assessed. The participation of the vagal and sympathetic pathways and involvement of CCK-A receptors were considered. The degree of severity was evaluated using biochemical and histopathological analyses. RESULTS: Induction of AP after stress was more severe than in its absence. Acinar and fat necrosis, hemorrhage and neutrophil infiltrate foci were evenly distributed, being significantly greater in size and number after stress. Gastric ulceration evolved to ulcer, hemorrhage and gastric necrosis after AP triggering. Serum amylase, lipase, C-reactive protein, IL-6, IL 10 and plasmatic hsp72 as well as pancreatic and lung myeloperoxidase were significantly elevated in AP after stress while pancreatic amylase and lipase were significantly reduced. AAR blockage ameliorated AP after stress. CONCLUSIONS: Stress aggravates pancreatic pathology while AP deteriorates gastric pathology, and anesthetic treatment was beneficial for both. Restraint in other animal models can be useful to study the influence of stress in the evolution of other diseases. PMID- 15286436 TI - Gonadal hormones humour the brain. AB - The relationship between the brain and the endocrine system is now seen to extend far beyond the regulation of somatic hormone production by the hypothalamus and pituitary: the brain itself can be considered both as an endocrine organ, producing hormones that act both within and outside the central nervous system, and as a target for hormones. The current extent of this concept with respect to the gonadal hormones was explored at a recent meeting ('Hormones and the Brain', Third Endocrinology Colloquium of the Fondation Ipsen, Paris, December 8, 2003). The discussion, reviewed in this article, ranged from intracellular signalling pathways and intercellular networks regulating hormone production and action in the central nervous system to hormone involvement in the generation of sexual behaviour and in development, plasticity, neuroprotection and repair. The hormonal contribution to psychiatric and neurodegenerative illnesses was also examined. The picture presented is complex, with layers of controls and with hormones that have diverse actions at different sites in the central nervous system. This richness of actions and functions is providing some interesting leads for developing new therapeutics. PMID- 15286437 TI - Blockade of endothelin receptors attenuates end-organ damage in homozygous hypertensive ren-2 transgenic rats. AB - BACKGROUND/AIMS: A growing body of evidence suggests that the interplay between the endothelin (ET) and the renin-angiotensin systems (RAS) plays an important role in the development of the malignant phase of hypertension. The present study was performed to evaluate the role of an interaction between ET and RAS in the development of hypertension and hypertension-associated end-organ damage in homozygous male transgenic rats harboring the mouse Ren-2 renin gene (TGRs) under conditions of normal-salt (NS, 0.45% NaCl) and high-salt (HS, 2% NaCl) intake. METHODS: Twenty-eight-day-old homozygous male TGRs and age-matched transgene negative male normotensive Hannover Sprague-Dawley (HanSD) rats were randomly assigned to groups with NS or HS intake. Nonselective ET(A/B) receptor blockade was achieved with bosentan (100 mg/kg/day). Systolic blood pressure (BP) was measured in conscious animals by tail plethysmography. Rats were placed into metabolic cages to determine proteinuria and clearance of endogenous creatinine. At the end of the experiment the final arterial BP was measured directly in anesthetized rats. Kidneys were taken for morphological examination. RESULTS: All male HanSD fed either the NS or HS diet exhibited a 100% survival rate until 180 days of age (end of experiment). The survival rate in untreated homozygous male TGRs fed the NS diet was 41%, which was markedly improved by treatment with bosentan to 88%. The HS diet reduced the survival rate in homozygous male TGRs to 10%. The survival rate in homozygous male TGRs on the HS diet was significantly improved by bosentan to 69%. Treatment with bosentan did not influence either the course of hypertension or the final levels of BP in any of the experimental groups of HanSD rats or TGRs. Although the ET-1 content in the renal cortex did not differ between HanSD rats and TGRs, ET-1 in the left heart ventricle of TGRs fed the HS diet was significantly higher compared with all other groups. Administration of bosentan to homozygous male TGRs fed either the NS or HS diet markedly reduced proteinuria, glomerulosclerosis and attenuated the development of cardiac hypertrophy compared with untreated TGR. CONCLUSIONS: Our data show that nonselective ET(A/B) receptor blockade markedly improves the survival rate and ameliorates end-organ damage in homozygous male TGRs without significantly lowering BP. PMID- 15286438 TI - Eosinophils accumulate in the gastric mucosa of food-allergic mice. PMID- 15286439 TI - Proteomic analysis of putative latex allergens. AB - BACKGROUND: Extensive analysis of allergenic proteins is generally time-consuming and labor-intensive. Accordingly, a rapid and easy procedure for allergen identification is required. As sequence information on proteins and genes is accumulated in databases, it is becoming easier to identify a candidate protein using proteomic strategies, i.e. two-dimensional gel electrophoresis, site specific fragmentation, mass spectrometry and then database search. In this study, we evaluated the usefulness of a proteomic strategy for identifying putative allergens through its application to latex proteins. METHODS: Latex proteins were separated with two-dimensional gel electrophoresis, and putative allergens were visualized by IgE immunoblotting using pooled serum from latex sensitive patients. The IgE-interactive proteins were cut out from the negatively stained two-dimensional gel and subjected to in-gel digestion by trypsin. Then the resulting peptides were analyzed with mass spectrometry. Based on the mass spectrometric data we obtained, the allergen candidates were assigned by a database search. RESULTS: Five previously reported allergens and five new allergen candidates were identified with the proteomic approach without isolating the individual proteins. Less than 1 mg of crude latex protein was sufficient for the entire protocol. Because plural proteins can be processed in parallel, analysis of about 50 IgE-interactive proteins was accomplished within 1 week. CONCLUSIONS: Analysis of putative allergens with proteomic strategies (allergenomics) is a promising avenue for rapid and exhaustive research. The high resolving power of two-dimensional gel electrophoresis is superior to conventional gel electrophoresis. Moreover, the notable sensitivity and speed of mass spectrometry have pronounced advantages over the N-terminal sequencing that has generally been used for protein identification. PMID- 15286440 TI - Thrombin affects eosinophil migration via protease-activated receptor-1. AB - BACKGROUND: Protease-activated receptors (PARs) are a unique class of G-protein coupled receptors, which are activated by proteolytic cleavage of the amino terminus of the receptor itself. Although expression of the PAR1, which is typically activated by thrombin, on human eosinophils has been demonstrated, no effect of thrombin on eosinophil function has been shown yet. Thus we investigated whether thrombin affects eosinophil migration in vitro. METHODS: Eosinophils were obtained from venous blood of healthy donors. Cell migration was studied by micropore filter assays. Involvement of PARs in thrombin-dependent migration was tested functionally using selective agonist peptides for PARs and a cleavage blocking PAR1 antibody. RESULTS: Thrombin significantly stimulated eosinophil chemotaxis in a dose-dependent manner. This effect was mimicked by the PAR1 but not the PAR2 agonist and was reversed by the cleavage blocking PAR1 antibody. Checkerboard experiments indicated that eosinophil migration depends on the presence of thrombin in a concentration gradient. CONCLUSIONS: Data suggest that activation of PAR1 by thrombin stimulates directed migration of human eosinophils and thereby may affect eosinophils in tissue and allergic inflammation. PMID- 15286441 TI - Isodon japonicus decreases immediate-type allergic reaction and tumor necrosis factor-alpha production. AB - BACKGROUND: The immediate-type allergic reaction is involved in many allergic diseases such as asthma, allergic rhinitis and sinusitis. The discovery of drugs for the treatment of immediate-type allergic disease is a very important subject in human health. Isodon japonicus Hara (Labiatae) (IJAE) has been used for centuries as a traditional medicine in Korea and is known to have an anti inflammatory effect. However, its specific mechanism of action is still unknown. In this report, we investigated the effect of IJAE on the immediate-type allergic reaction and studied its possible mechanisms of action, focusing on the mast cell mediated allergic reaction. METHODS: IJAE extracts were anally administered to mice for high and fast absorption. Compound 48/80-induced mortality and compound 48/80- or immunoglobulin E (IgE)-induced histamine release were measured to evaluate the antiallergic effects of IJAE. The effect of IJAE on the model of local allergic reaction in vivo, passive cutaneous anaphylaxis (PCA), was investigated. The production of tumor necrosis factor-alpha (TNF-alpha) was measured by Western blotting. RESULTS: IJAE inhibited compound 48/80-induced systemic reactions and plasma histamine release in mice. IJAE decreased the PCA reaction activated by anti-dinitrophenyl (DNP) IgE antibody. IJAE dose dependently reduced histamine release from rat peritoneal mast cells activated by compound 48/80 or anti-DNP IgE. Furthermore, IJAE decreased the production of TNF alpha in phorbol 12-myristate 13-acetate plus calcium ionophore A23187-stimulated human mast cells. CONCLUSION: Our findings provide evidence that IJAE inhibits mast cell-derived immediate-type allergic reactions, and also demonstrate the involvement of TNF-alpha in these effects. We propose the clinical use of IJAE in mast cell-mediated immediate-type allergic diseases. PMID- 15286442 TI - Intratracheal instillation of cytoplasmic granules from Phleum pratense pollen induces IgE- and cell-mediated responses in the Brown Norway rat. AB - BACKGROUND: Release of cytoplasmic granules from grass pollen upon contact with water is thought to be an important source of airborne allergens. OBJECTIVES: To investigate the humoral and cellular responses to intratracheal instillation of Phleum pratense (timothy grass) pollen cytoplasmic granules (PCG) in the Brown Norway rat. METHODS: PCG were purified from timothy grass pollen by filtration through 5-microm-mesh filters. Rats were sensitized (day 0) and challenged (day 21) intratracheally with purified PCG suspended in saline (6 x 10(6) PCG/rat). Rats were then challenged 4 weeks later (1.5 x 10(6) PCG/rat). Blood samples, bronchial lymph nodes and lungs were collected from the rats 4 days after the second challenge. PCG-specific IgE and IgG1 levels and specificity were determined by ELISA and Western blotting. Pollen, pollen extract and PCG-induced proliferation of lymph node cells were monitored by [(3)H]-thymidine incorporation in a lymph node assay. Histopathological examination was carried out on the lungs. RESULTS: Specific IgE and IgG1 were present in the sera. Cultured lymph node cells proliferated in the presence of pollen, pollen extract and PCG. Western blots showed that all major pollen allergens are recognized by IgE and IgG1 from PCG-treated rats. Histopathological examination revealed features of a mild allergic reaction. CONCLUSIONS: In our rat model of allergy, purified timothy grass PCG instillation induced specific antibodies and lymph node cell responses, comparable to those obtained with intact pollen. PMID- 15286443 TI - Sputum levels of reduced glutathione increase 24 hours after allergen challenge in isolated early, but not dual asthmatic responders. AB - BACKGROUND: The pathogenesis of late asthmatic reactions after allergen challenge in contrast to isolated early responses is incompletely understood. Recently, the antioxidant glutathione and endogenous nitrosothiols were shown to protect against bronchoconstriction. We compared reduced (GSH) and oxidized glutathione (GSSG) and nitrosothiols in induced sputum following allergen challenge in mild asthmatics with isolated early (EAR) and dual early and late (LAR) asthmatic responses. METHODS: Exhaled nitric oxide, sputum cells and sputum supernatant concentrations of GSH, GSSG and nitrosothiols were quantified 2-5 days prior to and 24 h after allergen challenge in 24 mild asthmatics (12 EAR, 12 LAR, only beta-agonists prn). RESULTS: There were no differences at baseline between EAR and LAR asthmatics for any of the parameters (p > 0.1, all comparisons). Mean +/- SD fall in forced expiratory volume in 1 s, expressed as the percentage decrease compared to the baseline value, between 3 and 8 h after allergen challenge was 1 +/- 5% in the group of patients without LAR vs. 24.9 +/- 8.7% in the group of patients with LAR (p < 0.001). Sputum eosinophils increased in both groups (p < 0.05, both comparisons), whereas neutrophils only increased in LAR subjects (p = 0.06 vs. EAR). In contrast, GSH was significantly increased 24 h after challenge only in EAR asthmatics [geometric mean with 95% confidence intervals: before: 3.3 microM (1.25-7.9 microM), after: 5.9 microM (2.7-12.9 microM), p = 0.05; mean difference vs. LAR subjects: 6 microM (0.1-12 microM), p = 0.048], and the proportion of GSSG was positively correlated with postallergen eosinophils in all patients (rho = 0.4, p = 0.05). There was no change in nitrosothiols after 24 h in either EAR or LAR subjects (p > 0.23, all comparisons). CONCLUSIONS: GSH increases 24 h after allergen challenge in isolated early responders. These data suggest that different adoptive responses to allergen may result in different physiologic phenotypes. Further studies on the role of glutathione in allergen induced bronchoconstriction are clearly warranted. PMID- 15286444 TI - A case of peanut cross-allergy to lupine flour in a hot dog bread. AB - BACKGROUND: In a case monitored by the Norwegian National Register for Severe Allergic Reactions to Food, a patient with peanut allergy experienced an allergic reaction after eating a particular brand of hot dog bread. The aim of this study was to identify the eliciting allergen. METHODS: Extracts from the hot dog bread and reference material from peanut, lupine and lupine-fortified food products were analysed by immunochemical methods with patient serum and a new polyclonal anti-lupine antibody. RESULTS: Evidence could be provided that the hot dog bread contained proteins from lupine but not from peanut. CONCLUSION: Crossed peanut lupine allergy can have clinical significance. A peanut-allergic patient reacted against hidden lupine protein in a hot dog bread. Presented with our results, the producer confirmed the use of lupine flour and changed the ingredient list. PMID- 15286445 TI - Evaluation of recombinant and native timothy pollen (rPhl p 1, 2, 5, 6, 7, 11, 12 and nPhl p 4)- specific IgG4 antibodies induced by subcutaneous immunotherapy with timothy pollen extract in allergic patients. AB - BACKGROUND: Allergen immunotherapy is a widely accepted treatment for IgE mediated allergies. The evaluation of immunotherapy-induced IgG4 antibodies based on allergen extract is questionable because the amount of allergen-extract specific IgG4 to individual disease-eliciting allergens cannot be determined using crude allergen extracts. In this study, we examined the specific IgE and IgG4 serum binding profiles to individual Phleum pratense allergens in grass pollen-sensitive patients who had received grass-pollen-specific immunotherapy (SIT). METHODS: The study included 33 patients from North-West Italy. All suffered from seasonal rhinoconjunctivitis and/or asthma. A modified "cluster" regimen of injections of a standardized aluminium-adsorbed P.pratense extract, with once-weekly visits and 10 injections for 5 weeks followed by 3 weeks of maintenance injections was instituted. Patients' sera were analyzed for specific IgE and IgG4 reactivity to individual P. pratense allergens (recombinant Phl p 1, Phl p 2, Phl p 5, Phl p 6, Phl p 7, Phl p 11, Phl p12 and native Phl p 4) and natural P. pratense extract using the Pharmacia CAP system. RESULTS: IgE reactivities to new allergen components were not detected by CAP in treated patients after 15 weeks and a cumulative dose of approximately 65 microg of the major allergen Phl p 5. Patients lacking specific IgE reactivity towards individual allergens at the start of SIT did not produce significant levels of specific serum IgG4 to serum IgE-negative allergens. On the other hand, an increase in specific IgG4 only to allergens to which patients were previously sensitized was observed. Significant increases in specific IgG4 levels to rPhl p 1 (p < 0.05), 2 (p < (0.01), 5 (p < 0.0001), 6 (p < 0.0001), 7 (p < 0.05), 11 (p < 0.05) and nPhl p 4 (p < 0.01) were observed after P. pratense extract immunotherapy. No significant rPhl p 12-specific IgG4 antibody increase was documented after treatment. CONCLUSION: These findings suggest that Phl p 12 was underrepresented in the extract used, as indicated by the low specific IgG4 response induced by this grass-pollen-specific vaccine. Thus, the simple detection of specific serum IgG4 antibodies a few weeks after the start of SIT could represent a valuable tool to estimate the presence of relevant allergens in a given immunotherapeutic allergen extract. PMID- 15286446 TI - Soluble interleukin-5 receptor alpha is increased in acute exacerbation of chronic obstructive pulmonary disease. AB - BACKGROUND: During chronic obstructive pulmonary disease (COPD) exacerbations (AE COPD), an influx of eosinophils into the bronchial mucosa has been described. Eosinophilic cationic protein (ECP) and soluble interleukin-5 receptor alpha (sIL5Ralpha) are secreted by eosinophils and increased in eosinophilic airway diseases. METHODS: We studied ECP and sIL5Ralpha expression in patients with COPD compared to healthy controls and smokers and investigated a possible association to viral exacerbations of COPD. Expression of sIL5Ralpha in serum was analyzed by ELISA and ECP by the Uni-Cap system. Induced sputum from patients with COPD was analyzed for six different respiratory viruses by nested PCR. RESULTS: ECP and sIL5Ralpha were significantly elevated in AE-COPD subjects (n = 54) compared to healthy controls (n = 11, p = 0.018). Furthermore, there was a significant increase in sIL5Ralpha, but not in ECP, in 30 patients with virus-associated AE COPD compared to smokers without COPD (n = 16) and healthy controls. The increase in FEV(1) after resolution of the AE-COPD correlated with the decrease in sIL5Ralpha (r = 0.269, p = 0.034). CONCLUSIONS: sIL5Ralpha is increased in AE COPD and not affected by smoking like ECP. sIL5Ralpha is increased in patients with virus-associated AE-COPD compared to smokers and controls. Concentrations of sIL5Ralpha mirror changes in the clinical status and lung function. These data support the involvement of eosinophils in acute exacerbations of COPD. PMID- 15286447 TI - The childhood component of the ICP model is appropriate for growth analysis of short Israeli children. AB - OBJECTIVE: To determine whether the childhood component of the infancy-childhood puberty (ICP) model is appropriate for growth analysis of short Israeli children. SUBJECTS AND METHODS: From 204 short, prepubertal children, 2-16 years of age, 1,516 height measurements were analyzed. For each child's measurements, a best fitted line based on C equation of ICP has been drawn and the distribution of measurement points around that line was calculated. RESULTS: Ninety percent of the measurements were at a distance of no more than +/- 2 cm from the best-fitted ICP line. CONCLUSION: The C component of ICP model can be used as a growth analysis tool for shorter than average, prepubertal, Israeli children, older than 2 years of age. PMID- 15286448 TI - Lost and found testes: the importance of the hCG stimulation test and other testicular markers to confirm a surgical declaration of anorchia. AB - BACKGROUND: In patients with impalpable testes,laparoscopy or open surgery is considered conclusive in establishing the absence of testicular tissue. METHODS: Retrospective chart review. RESULTS: Over a 22-year period, 4 out of 82 patients with a diagnosis of bilateral anorchia by laparoscopy or laparotomy had persistent testicular tissue suggested by endocrine evaluations. The clue to the presence of testicular tissue was: (1) a pubertal rise in plasma testosterone (2 patients); (2) the presence of possible Mullerian structures and of a detectable plasma anti-Mullerian hormone (1 patient), and (3) the fact that one of the gonads had not been seen at surgery (1 patient who still had a testosterone response to hCG postoperatively). Testes were localized by venography (3 patients) and laparotomy (1 patient). CONCLUSION: A surgical diagnosis of bilateral anorchia needs to be confirmed by hCG stimulation, gonadotropin levels, or other markers of testicular function. PMID- 15286449 TI - Expression of leptin receptor isoforms and effects of leptin on the proliferation and hormonal secretion in human pituitary adenomas. AB - OBJECTIVE: To pursue whether leptin regulates anterior pituitary cells, we studied the ex vivo expression of several isoforms of the leptin receptor (OB-R) as well as the in vitro effects of leptin administration in human pituitary adenomas. METHODS: OB-R mRNA expression and in vitro response to leptin were studied in 39 pituitary macroadenomas. RESULTS: All 4 OB-R subtypes were expressed in most adenomas. The expression was significantly more pronounced in GH-secreting adenomas as compared to non-functioning tumor cells (p < 0.05). Leptin administration in vitro did not significantly influence cell proliferation or the secretion of GH, FSH, LH or alpha-subunit. CONCLUSIONS: (1) Several isoforms of the OB-R, including the signal transducing full-length receptor, are expressed in most human pituitary adenomas. (2) This expression ex vivo is not associated with significant effects of leptin in vitro. PMID- 15286450 TI - Magnetic resonance spectroscopy features of uterine leiomyomas. AB - OBJECTIVE: The purpose of this study is to investigate the in vivo magnetic resonance spectroscopy features of uterine leiomyomas using long echo time and to characterize the spectral patterns of these lesions. METHODS: We calculated metabolites in 15 patients with uterine leiomyomas and myometrium of 20 healthy control subjects using single-voxel proton MR spectroscopy (point resolved spectroscopy technique, TE:136 ms). Voxels were placed at the center of the uterine leiomyomas. The peak areas of creatine, choline, lipid and lactate were determined. The MR spectroscopy results of uterine leiomyomas were compared with the spectroscopy results obtained from the myometrium of healthy control subjects. RESULTS: The characteristically obtained signal was choline, which was detected not only in 14 of the 15 leiomyomas (93.3%) but also in 18 of the 20 myometrium of control subjects (90%). The lipid signals were determined in 9 of 15 patients with uterine leiomyomas (60%) and 8 of 20 control subjects (40%). The lactate signal was obtained from six of 15 patients with leiomyomas (40%) but only two of myometrium (10%). The creatine signal was obtained from 4 of 15 patients with leiomyomas (26.6%) and 5 of 20 myometrium (25%). Among the tested parameters only lactate peak was statistically significant (p < 0.05). CONCLUSION: Proton MR spectroscopic imaging may be helpful for the investigation of the underlying pathophysiology of uterine leiomyomas. The presence of lactate and lipid signals in the spectrum may be a useful indicator of metabolic pathway of uterine leiomyomas. PMID- 15286451 TI - Predictive factors for noncompliance with follow-up among women treated for cervical intraepithelial neoplasia. AB - The aim of this study was to explore the magnitude and causes of noncompliance with follow-up after cervical intraepithelial neoplasia (CIN) treatment. Women who were treated by large loop excision of the transformation zone (LLETZ) for CIN between January 1994 and June 1996 were enrolled. Women who kept all their appointments were grouped as compliant. To the noncompliant group belonged those women who missed at least two follow-up appointments during the first year of follow-up. Seventy-four of 141 women (52.5%) who were treated for CIN were noncompliant. Fifty women from each group were then evaluated. Demographic, social and contraceptive parameters were found to be similar between the groups. The noncompliant group had a nonsignificant trend towards having had only elementary education, undergoing surgery in the past, additional diseases, taking chronic medications and having diseases in the family. We conclude that noncompliance with follow-up after treatment of CIN by LLETZ may be much higher than expected. Noncompliant patients do not have any significantly defined characteristics. PMID- 15286452 TI - A preclinical phase in vascular dementia: cognitive impairment three years before diagnosis. AB - Alzheimer's disease (AD) and vascular dementia (VaD) patients exhibit similar patterns of deficits in many cognitive tasks in the early clinical stages. Considering that preclinical cognitive deficits are well documented in AD, the purpose of the present study was to investigate if such deficits are also present in VaD. The cognitive outcome measure was the Mini-Mental State Examination (MMSE). The sample was taken from a population-based study and consisted of 699 persons who were nondemented at baseline, but out of whom 35 persons were diagnosed with VaD and 170 with AD at a 3-year follow-up. Both the incident VaD and AD cases exhibited baseline deficits on the total score of the MMSE and three of the subscales: orientation to time, orientation to place, and delayed memory. Further, both dementia groups exhibited precipitous decline on most MMSE subscales during the 3-year follow-up period. Logistic regression analyses showed that all subscales that revealed deficits at baseline predicted dementia status at follow-up. Delayed memory was the best predictor in both preclinical VaD and preclinical AD. Thus, these results demonstrate preclinical cognitive deficits in VaD in a measure of global cognitive functioning, which closely resemble those observed in AD. This observation suggests that circulatory disturbance is associated with cognitive problems several years before the actual VaD diagnosis. PMID- 15286453 TI - Impact of different diagnostic criteria on prognosis of delirium: a prospective study. AB - A 2-year follow-up study was performed to compare the prognosis of delirium defined according to 4 different diagnostic classifications (DSM-III, DSM-III-R, DSM-IV and ICD-10 clinical criteria) among 425 elderly geriatric hospital patients and nursing home residents. The proportion of delirium varied from 24.9% (DSM-IV) to 10.1% (ICD-10). The prognoses were similar particularly according to all DSM classifications: 31.3-36.3% of the delirious patients died within 1 year and 57.8-62.5% within 2 years. The number of subjects diagnosed as delirious according to the ICD-10 was small, and their prognosis did not differ significantly from the others either. The DSM-IV has simplified the criteria of delirium. It identifies new, acutely ill and relatively nondependent subjects as delirious who share the poor prognosis of patients diagnosed with the previous criteria. PMID- 15286454 TI - SREBP-1a polymorphism influences the risk of Alzheimer's disease in carriers of the ApoE4 allele. AB - Sterol regulatory element-binding proteins (SREBPs) are transcription factors involved in cholesterol and fatty acid synthesis. Recently, a polymorphism in the 5'-region of the SREBP-1a gene has been described to be correlated with alterations in the plasma levels of cholesterol. Consequently the relationship between this SREBP-1a gene polymorphism and Alzheimer's disease (AD) alone and in combination with the apolipoprotein E (ApoE) 4 allele was evaluated. No association between SREBP-1a polymorphism alone and AD could be seen. However, in the group of healthy ApoE4 allele carriers, the number of homozygote SREBP-1a DeltaG allele carriers was significantly higher than in AD patients. Cerebrospinal fluid levels of cholesterol were lower in AD patients who were carriers of the SREBP-1a DeltaG allele, and the ratio of 24S-hydroxycholesterol to cholesterol was increased in these probands. Our data suggest a reduced risk of AD in carriers of an ApoE4 allele who are additionally homozygous for the SREBP-1a DeltaG allele, which is possibly due to the influence of SREBP-1a polymorphism on brain cholesterol metabolism. This is the first report on a genetic factor which prevents the deleterious effect of the ApoE4 allele and thus reduces the risk of AD. PMID- 15286455 TI - The scoring scheme of the informant questionnaire on cognitive decline in the elderly needs revision: results of rasch analysis. AB - The purpose of this study was to examine the optimal scoring scheme (category use), unidimensionality, item fit, and redundancy of the Informant Questionnaire on Cognitive Decline in the Elderly (IQCODE) in a cohort of Hong Kong Chinese stroke survivors. At 3 months after the index stroke, a research assistant administered the IQCODE to relatives of 284 Chinese patients with acute stroke who were consecutively admitted to a general hospital. A psychiatrist, who was blinded to the IQCODE scores, interviewed all 284 patients and made DSM-IV diagnosis of dementia, which served as the benchmark for judging the performance of IQCODE in screening dementia. The results suggest that the optimal IQCODE scoring scheme has 2 rather than the original 5 categories. Although the IQCODE was unidimensional overall, there was evidence of item redundancy, thus indicating that a shortened version is desirable. PMID- 15286456 TI - Genetic association of CYP46 and risk for Alzheimer's disease. AB - An increasing number of studies suggest that cholesterol plays an important role in regulating beta-amyloid (Abeta) metabolism in Alzheimer's disease (AD). One of the most important mechanisms for the elimination of excess brain cholesterol is its conversion into the 24S-hydroxycholesterol catalyzed by cholesterol 24S hydroxylase (CYP46). Preliminary evidence indicates that an intron 2 CYP46 T/C gene polymorphismis associated with increased brain Abeta load and higher risk of AD. A case-control study utilizing a clinically well-defined group of 321 sporadic AD patients and 315 control subjects was performed to test this association. Our results indicate that the intron 2 CYP46 C/C genotype may predispose to AD, and this association is independent of the apolipoprotein E genotype. PMID- 15286457 TI - Polymorphisms of the macrophage inhibitory factor and C-reactive protein genes in subjects with Alzheimer's dementia. AB - Neuroinflammation is a central feature of Alzheimer's disease (AD). C-reactive protein (CRP) is a key molecule of the acute phase of inflammation that has been localized in the two characteristic lesions of AD brain, senile plaque and neurofibrillary tangles. On the other hand, the macrophage migration inhibitory factor (MIF) is a cytokine with multiple biological activities, including the ability to act as potent amyloid beta (A-beta)-binding protein. Two common polymorphisms have been recently detected in the genes encoding for CRP and MIF and have been associated with significant modifications of plasma levels and activity of the corresponding proteins. Following these observations, we hypothesized that CRP and MIF gene polymorphisms might contribute to the development and progression of neurodegenerative disorders and evaluated their association with AD. CRP and MIF gene polymorphisms were examined by polymerase chain reaction and restriction enzyme analysis in 116 Italian subjects affected by probable AD and 184 age- and sex-matched controls. We did not find a statistically significant difference in the distribution of CRP and MIF genotypes and alleles between AD subjects and controls. Although these data need further confirmation, they indicate that CRP and MIF gene polymorphisms are not associated with AD. PMID- 15286458 TI - Plasma antioxidant status, immunoglobulin g oxidation and lipid peroxidation in demented patients: relevance to Alzheimer disease and vascular dementia. AB - A large body of evidence supports a role of oxidative stress in Alzheimer disease (AD) and in cerebrovascular disease. A vascular component might be critical in the pathophysiology of AD, but there is a substantial lack of data regarding the simultaneous behavior of peripheral antioxidants and biomarkers of oxidative stress in AD and vascular dementia (VaD). Sixty-three AD patients, 23 VaD patients and 55 controls were included in the study. We measured plasma levels of water-soluble (vitamin C and uric acid) and lipophilic (vitamin E, vitamin A, carotenoids including lutein, zeaxanthin, beta-cryptoxanthin, lycopene, alpha- and beta-carotene) antioxidant micronutrients as well as levels of biomarkers of lipid peroxidation [malondialdehyde (MDA)] and of protein oxidation [immunoglobulin G (IgG) levels of protein carbonyls and dityrosine] in patients and controls. With the exception of beta-carotene, all antioxidants were lower in demented patients as compared to controls. Furthermore, AD patients showed a significantly higher IgG dityrosine content as compared to controls. AD and VaD patients showed similar plasma levels of plasma antioxidants and MDA as well as a similar IgG content of protein carbonyls and dityrosine. We conclude that, independent of its nature-vascular or degenerative-dementia is associated with the depletion of a large spectrum of antioxidant micronutrients and with increased protein oxidative modification. This might be relevant to the pathophysiology of dementing disorders, particularly in light of the recently suggested importance of the vascular component in AD development. PMID- 15286459 TI - Late-onset depression with mild cognitive deficits: electrophysiological evidences for a preclinical dementia syndrome. AB - Mild cognitive impairment (MCI) is present in up to 60% of patients with late onset depression and constitutes a major diagnostic problem in geriatric psychiatry. Searching for sensitive markers for the detection of early brain changes suggestive of dementia, we compared this depressive risk population with mildly to moderately demented patients and cognitively unimpaired depressed patients using EEG power and coherence. We found a considerable similarity between Alzheimer's disease patients and cognitively impaired depressed patients regarding the cognitive profile and EEG pattern. Changes in EEG power and coherence at frontotemporal leads in depressive patients with MCI thereby allowed discrimination from cognitively unimpaired patients with a sensitivity of 88% and a specifity of 81%. PMID- 15286460 TI - Pathogenic factors in vascular dementia and Alzheimer's disease. Multiple actions of heparin that probably are beneficial. AB - The following areas are discussed in this review: atherogenesis; cerebrovascular factors; hypoperfusion; beta-amyloid production; beta-amyloid fibril formation; beta-sheets; metal cations; reactive oxygen species/free radicals; chronic inflammatory factors; endogenous plasma heparin; lipoprotein lipase; polyamines; protein kinase C; casein kinases; phospholipase A2; serine proteases; myeloperoxidase; cyclooxygenase 2; cysteine proteases; caspases; proprotein convertases; aspartic proteases; cyclin proteinases; thrombin; tau hyperphosphorylation; advanced glycosylation end products; activator protein 1; calcium; apolipoprotein E epsilon4; histamine; blood-brain barrier; glutamate; transglutaminase; insulin-like growth factor 1. PMID- 15286461 TI - Comparative effects of two phototherapy delivery systems on cerebral blood flow velocity in term neonates. AB - BACKGROUND: Phototherapy is an effective and generally safe method of treating neonatal hyperbilirubinemia. However, there is some concern that it may affect cerebral blood flow velocity (CBFV) in newborns with fragile cerebral vasculature. OBJECTIVES: To measure and compare the effects of two different phototherapy units, delivering similar irradiance, on CBFVs. METHODS: Doppler flow velocities were measured in term infants under fluorescent overhead and fluorescent BiliBed phototherapy units, respectively, at baseline, 4 and 24 h of therapy. RESULTS: Peak systolic CBFV increased during treatment in infants treated under overhead phototherapy (n = 18) but not in those treated in BiliBeds (n = 12). CONCLUSIONS: Different phototherapy delivery modalities can have differential effects on CBFV in term neonates. PMID- 15286462 TI - Effects of mild vitamin a deficiency on lung maturation in newborn rats: a morphometric and morphologic study. AB - OBJECTIVE: To evaluate the effects of a 60% vitamin A deficiency (VAD) on the two postnatal stages of lung development: alveolarization and microvascular maturation. Lungs from deficient rats were compared to age-matched controls. STUDY DESIGN: Starting at 3 weeks before mating, female rats were maintained under a diet lacking vitamin A. Due to the slow depletion of the vitamin A liver stores the pregnant rats carried to term and delivered pups under mild VAD conditions. Mothers and offspring were then kept under the same diet what resulted in a mean reduction of vitamin A plasma concentration of about 60% vs. controls during the whole experimental period. Pups were sacrificed on days 4, 10 and 21 and their lungs fixed and analyzed by means of a combined morphologic and morphometric investigation at light and electron microscopic levels. RESULTS: During the whole experiment, body weights of VAD animals were lower than controls with a significant decrease on day 10. On days 4, 10 and 21 the pulmonary structure was in a comparable gross morphologic state in both groups. Despite this morphologic normality, quantitative alterations in some functional parameters could be detected. On day 4, lung volume and the volume and surface area of air spaces were decreased, while the arithmetic mean barrier thickness and type 2 pneumocyte volume were increased in the VAD group. On day 21, some changes were again manifest mainly consisting in an augmentation of the vascularization and a decrease in interstitial volume in deficient animals. CONCLUSIONS: Mild VAD causes no gross disturbances in the postnatal phases of lung development in rats. However, a body weight-related transient retardation of lung maturation was detectable in the first postnatal week. At 3 weeks, the VAD lungs showed a more mature vascular system substantiated by an increase in volume of both capillary volume and the large non-parenchymal vessels. In view of these quantitative alterations, we suspect that mild VAD deregulates the normal phases of body and lung growth, but does not induce serious functional impairments. PMID- 15286463 TI - Differences in testis cancer survival by race and ethnicity: a population-based study, 1973-1999 (United States). AB - OBJECTIVE: We examined the relationship between race/ethnicity and testis cancer survival in a population-based setting. METHODS: We analyzed 16,086 cases of primary testis cancer diagnosed during 1973-1999 and reported to 12 cancer registries participating in the National Cancer Institute's Surveillance, Epidemiology, and End Results Program. We compared testis cancer-specific survival between patients from different racial/ethnic groups by use of the hazard ratio (HR) and 95% confidence intervals (CI) calculated from Cox proportional hazards models, adjusting for stage, histology, and period of diagnosis. RESULTS: Relative to non-Hispanic whites, a greater proportion of African American, Native American, Hawaiian, and Hispanic patients were diagnosed at late stages. There were 886 deaths among 16,086 testis cancer patients and overall 5-year survival was 95%. After adjustment for stage, histology, and period of diagnosis, the risk of dying from testis cancer was increased among African Americans (HR = 2.3; CI: 1.6-3.2), Native Americans (HR = 2.1; CI: 1.1 3.9), Filipinos (HR = 3.6; CI: 1.3-9.5), Hawaiians (HR = 2.4; CI: 1.4-4.1), and Hispanics (HR = 1.4; CI: 1.1-1.8), compared to non-Hispanic whites. CONCLUSION: These findings are consistent with previous reports of race/ethnic disparities in stage at diagnosis and survival in testis cancer patients as well as other cancer patients. Further research is needed to understand the reasons underlying these disparities. PMID- 15286464 TI - Occupational exposure to vinyl chloride, acrylonitrile and styrene and lung cancer risk (europe). AB - Several industry-based cohort studies have addressed the risk of lung cancer following exposure to vinyl chloride, acrylonitrile and styrene, with inconsistent results and usually without smoking adjustment. These exposures are addressed here in a large case-control study with full adjustment for smoking. Almost 6000 subjects were included in a case-control study conducted in seven European countries. For each job they held, local experts assessed the exposure to a number of occupational agents, including vinyl chloride, acrylonitrile and styrene, on the basis of detailed occupational questionnaires. Information on tobacco consumption and other risk factors was also collected. The odds ratio (OR) for ever exposure to vinyl chloride was 1.05 (95% confidence interval, CI: 0.68-1.62) and a modest, non-significant increase in the risk of lung cancer was found in the highest exposed subgroup. The OR for ever exposure to acrylonitrile was 2.20 (95% CI: 1.11-4.36) with a positive dose-response relationship between estimated cumulative exposure and lung cancer risk. No association between exposure to styrene and lung cancer risk was found. In conclusion, we cannot exclude a weak association between occupational exposure to vinyl chloride and lung cancer risk. Exposure to acrylonitrile was associated in our study with risk of lung cancer. Exposure to styrene does not seem to increase lung cancer risk. PMID- 15286465 TI - Prevalence of whole-body skin self-examination in a population at high risk for skin cancer (Australia). AB - OBJECTIVE: Whole-body skin self-examination (SSE) with presentation of suspicious lesions to a physician may improve early detection of melanoma. The aim of this study was to establish the prevalence and determinants of SSE in a high-risk population in preparation for a community-based randomised controlled trial of screening for melanoma. METHODS: A telephone survey reached 3110 residents older than 30 years (overall response rate of 66.9%) randomly selected from 18 regional communities in Queensland, Australia. RESULTS: Overall, 804 (25.9%) participants reported whole-body SSE within the past 12 months and 1055 (33.9%) within the past three years. Whole-body SSE was associated in multivariate logistic regression analysis with younger age (< 50 years); higher education; having received either a whole-body skin examination, recommendation or instruction on SSE by a primary care physicial; giving skin checks a high priority; concern about skin cancer and a personal history of skin cancer. CONCLUSION: Overall, the prevalence of SSE in the present study is among the highest yet observed in Australia, with about one-third of the adult population reporting whole-body SSE in the past three years. People over 50 years, who are at relatively higher risk for skin cancer, currently perform SSE less frequently than younger people. PMID- 15286466 TI - Incidence of transitional cell carcinoma of the bladder and arsenic exposure in New Hampshire. AB - OBJECTIVE: Arsenic is a known bladder carcinogen and populations exposed to high arsenic levels in their water supply have reported elevated bladder cancer mortality and incidence rates. To examine the effects of lower levels of arsenic exposure on bladder cancer incidence, we conducted a case-control study in New Hampshire, USA where levels above 10 micro/l are commonly found in private wells. METHODS: We studied 383 cases of transitional cell carcinoma of the bladder cancer, newly diagnosed between July 1, 1994 and June 30, 1998 and 641 general population controls. Individual exposure to arsenic was determined in toenail clippings using instrumental neutron activation analysis. RESULTS: Among smokers, an elevated odds ratio (OR) for bladder cancer was observed for the uppermost category of arsenic (OR: 2.17, 95% CI: 0.92-5.11 for greater than 0.330 mcg/g compared to less than 0.06 micro/g). Among never smokers, there was no association between arsenic and bladder cancer risk. CONCLUSIONS: These, and other data, suggest that ingestion of low to moderate arsenic levels may affect bladder cancer incidence, and that cigarette smoking may act as a co-carcinogen. PMID- 15286467 TI - Physical activity, body size, and estrogen metabolism in women. AB - OBJECTIVES: Physical activity has demonstrable effects on estrogen levels in pre- and postmenopausal women. Increased oxidation of estrone to 2-hydroxyestrone (2HE) relative to 16alpha-hydroxyestrone (16HE) has been hypothesized to reduce breast cancer risk, but little is known about the effect of physical activity and body size in relation to the ratio of 2HE and 16HE in women. We examined these relationships in cross-sectional analyses of 157 North American and Chinese women. METHODS: Physical activity was assessed using validated questionnaires. Adiposity was assessed as body mass index (BMI, kg/m2) and by anthropometric methods (% body fat). Estrone metabolites, 2HE and 16HE, were determined from urine via ELISA. RESULTS: Regression analyses on the 2HE/16HE ratio revealed an interaction between leisure-time physical activities and adiposity in both North American and Chinese women (p < or = 0.05). Women reporting low levels of leisure time physical activity who had higher BMI levels had 2HE/16HE ratios that were lower than their lean counterparts. In contrast, women with higher BMI levels that were physically active maintained higher 2HE/16HE ratios. CONCLUSION: These findings suggest that physical activity participation has the potential to modify the adverse effect of increased adiposity on estrogen metabolism in North American and Chinese women. PMID- 15286468 TI - Green tea consumption and subsequent risk of gastric cancer by subsite: the JPHC Study. AB - OBJECTIVE: To investigate the relationship between green tea consumption and subsequent risk of gastric cancer at different anatomical subsites in a population-based prospective study. METHODS: The Japan Public Health Center-based prospective study (JPHC Study) was established in 1990 for Cohort I and in 1993 for Cohort II. Among 72,943 subjects (34,832 men and 38,111 women), 892 gastric cancer cases (665 men and 227 women) were identified from 1990 to 2001. RESULTS: While no association between green tea consumption and gastric cancer was observed among men, a decreased risk of gastric cancer was observed among women after adjustment for potential confounding factors. This result was more remarkable when only the tumors in the distal portion were analyzed; for that subsite, the relative risk was 0.51 (95% confidence interval 0.30-0.86) in the highest category of green tea consumption (5 or more cups per day versus less than 1 cup per day) (p for trend = 0.01). The null association for upper-third gastric cancer was consistent for both sexes. CONCLUSIONS: An inverse association between green tea consumption and distal gastric cancer was observed among women. More prospective studies with detailed information are needed to confirm the role of green tea in the occurrence of gastric cancer. PMID- 15286469 TI - Folate intake, MTHFR C677T polymorphism, alcohol consumption, and risk for sporadic colorectal adenoma (United States). AB - OBJECTIVE: The purpose of this study was to investigate whether folate intake is associated with risk for incident sporadic colorectal adenoma, and whether the association differs according to methylenetetrahydrofolate reductase (MTHFR) genotypes or is modified by intakes of alcohol or other micronutrients in the folate metabolism pathway. METHODS: The authors analyzed data from a colonoscopy based case-control study (n = 177 cases, 228 controls) conducted in North Carolina between 1995 and 1997. RESULTS: The multivariate-adjusted odds ratio (OR) comparing the highest to lowest tertile of total folate intake was 0.61 (95% confidence interval [CI] 0.35-1.05); for MTHFR C677T polymorphism CT and TT genotypes relative to the CC genotype they were, respectively, 1.09 (CI: 0.71 1.66) and 0.68 (CI: 0.29-1.61); and for heavy drinkers (> 3 drinks/week) compared to non-drinkers it was 1.67 (CI: 1.00-2.81). The multivariate-adjusted ORs comparing the highest to lowest tertile of total folate intake according to those with the MTHFR CC, CT, and TT genotypes, were, respectively, 0.65 (CI: 0.30 1.39), 0.57 (CI: 0.23-1.44), and 0.22 (CI: 0.02-3.19). For those in the lowest tertile of folate intake who drank more than three drinks a week compared to those who were in the highest tertile of folate intake and did not drink alcohol the OR was 6.54 (CI: 1.96-21.80). There was no substantial evidence for interactions of folate with intakes of methionine, vitamins B2, B6, or B12. CONCLUSIONS: These data are consistent with hypotheses and previous findings that higher folate intake may reduce risk for colorectal neoplasms, perhaps especially among those who consume more alcohol. PMID- 15286470 TI - Better knowledge translation for effective cancer control: a priority for action. AB - Increasing cancer rates are a world wide problem. Efforts towards controlling cancer are most effectively implemented through national cancer control programs. The literature has emphasized prevention and screening as main starting points; by applying what we know a substantial amount of cancer could be prevented. As well, in the areas of access to care, treatment and palliation, there are also many gains to be made. However, despite advances in fundamental and applied research across the cancer continuum, there continue to be delays between research discovery and application. Translation of research knowledge has focused on means traditionally part of the research process such as publication in journals. While knowledge may be disseminated via these methods, they appear to have little impact on implementation of new approaches in practice or policy. Research in the area of knowledge translation identifies important elements and strategies most effective in the translation of research findings. Adding a knowledge translation component to national cancer control programs can help ensure that even small efforts directed at cancer control can have maximum impact. PMID- 15286471 TI - Occupational exposure to carbon black and risk of cancer. AB - OBJECTIVES: To investigate cancer risk in dockyard workers exposed to carbon black. METHODS: Cancer incidence was ascertained among 2101 longshoremen employed at the dockyard of Genova, Italy They were categorized a priori as exposed to low, moderate, and high level of carbon black dust. Incidence rates for the male population of the City of Genova were used to compute standardized incidence ratios (SIR). RESULTS: A positive exposure-response relation with carbon black exposure was detected only for bladder cancer (SIR = 204, 95%CI = 112-343, in highly exposed workers). Increased incidence of pleural mesothelioma (SIR = 751, 95%CI = 302-1547) and melanoma (SIR = 288, 95%CI = 125-2168) were detected. CONCLUSION: Exposure to carbon black experienced by dockyard workers was associated with a two-fold increased risk of bladder cancer. PMID- 15286472 TI - Stem cells and prenatal origin of breast cancer. AB - The hypothesis that in utero exposure to pregnancy hormones, notably estrogens, is related to the occurrence of breast cancer in the offspring has been examined in a number of epidemiological and experimental studies. Many studies have provided direct or indirect evidence that supports the hypothesis of an intrauterine component in the origin of breast cancer. Human studies to examine the underlying biological mechanisms, however, have been limited. We review the likely role of stem cells in hormone-mediated carcinogenic process, particularly as intermediate steps between in utero exposure to hormones and breast cancer. We summarize also studies related to the assumptions of the hypothesis concerning in utero exposure. We propose the use of stem cell potential as a measurable variable of the 'fertile soil', a term that has been used to characterize the consequences of fetal exposure to intrauterine environment. We conclude by outlining a feasible population-based study that measures stem cell potential to explore mechanisms mediating the relation between in utero exposure to pregnancy hormones and breast cancer risk in the offspring. PMID- 15286485 TI - Assessing pain in patients with advanced dementia. PMID- 15286486 TI - Closing the case on the keep-vein-open rate. PMID- 15286490 TI - Inserting an indwelling urinary catheter in a female patient. PMID- 15286491 TI - Promoting cardiac rehabilitation. PMID- 15286492 TI - How do I care for a patient with alcohol withdrawal syndrome? PMID- 15286500 TI - Clostridium perfringens. PMID- 15286511 TI - Seizures 101. PMID- 15286512 TI - "Eating our young" isn't practiced here. PMID- 15286513 TI - The changing face of AIDS. PMID- 15286514 TI - Understanding pleural effusion. PMID- 15286517 TI - Managing venous and neuropathic ulcers. PMID- 15286518 TI - Tips on implementing safety devices. PMID- 15286522 TI - Epistaxis. PMID- 15286525 TI - Expanding the scope of practice for cardiac rehabilitation: managing patients with implantable cardioverter defibrillators. PMID- 15286526 TI - Women's Initiative for Nonsmoking-VII: evaluation of health service utilization and costs among women smokers with cardiovascular disease. AB - PURPOSE: The Women's Initiative for Nonsmoking (WINS), a randomized clinical trial of a smoking cessation intervention for women with cardiovascular disease, permitted an assessment of the types and costs of health services women used during the 30 months after their hospitalization with cardiovascular disease. METHODS: A prospective design nested within WINS was used for this study. A structured telephone interview guide included questions about medical services and 15 categories of prevention services, including cardiac rehabilitation at 6, 12, 24, and 30 months. Costs were estimated from state and national databases. RESULTS: The 277 women studied had a mean age of 60.7 +/- 10 years. They had smoked approximately 40 +/- 11.4 years. More than 50% of the women had one or more risk factors for cardiovascular disease. During the first 6 months after the index hospitalization, 94% had a physician visit, 39% had an emergency-room visit, and 36% had a hospital admission. Prevention services used were home healthcare by nurse or home health aide (26%), a cardiac rehabilitation program, including Multifit and Heart Smart (19%), and physical therapy (14%). Usage decreased over the 30 months. For the women who used any service, the mean total monthly cost per woman was 913 dollars +/- 1204 dollars. CONCLUSIONS: This is the first report on health service use by women smokers with cardiovascular disease. Data collection using a telephone interview guide proved feasible for evaluating health service use. The greatest costs resulted from hospital admissions and physician and emergency-room visits. Considering the high prevalence of risk factors in this cohort, secondary prevention services were severely underutilized. By increasing referrals to such services, physicians and nurses might influence women to reduce their risk for subsequent cardiovascular disease. PMID- 15286527 TI - Acceptability of a low-fat vegan diet compares favorably to a step II diet in a randomized, controlled trial. AB - PURPOSE: This study aimed to assess the acceptability of a low-fat vegan diet, as compared with a more typical fat-modified diet, among overweight and obese adults. METHODS: Through newspaper advertisements, 64 overweight, postmenopausal women were recruited, 59 of whom completed the study. The participants were assigned randomly to a low-fat vegan diet or, for comparison, to a National Cholesterol Education Program Step II (NCEP) diet. At baseline and 14 weeks later, dietary intake, dietary restraint, disinhibition, and hunger, as well as the acceptability and perceived benefits and adverse effects of each diet were assessed. RESULTS: Dietary restraint increased in the NCEP group (P <.001), indicating a greater subjective sense of constraint with regard to diet requirements, but was unchanged in the vegan group. Disinhibition and hunger scores fell in each group (P <.001 and P <.01, respectively). The acceptability of both diets was high, although the vegan group participants rated their diet as less easy to prepare than their usual diets (P <.05) and the NCEP participants foresaw continuation of their assigned diet to be more difficult than continuation of their baseline diets (P <.05). There were no between-group differences on any acceptability measures. CONCLUSIONS: The acceptability of a low-fat vegan diet is high and not demonstrably different from that of a more moderate low-fat diet among well-educated, postmenopausal women in a research environment. PMID- 15286528 TI - Effects of cardiac rehabilitation on the recovery outcomes of older adults after coronary artery bypass surgery. AB - PURPOSE: This study aimed to examine differences in lower extremity function as well as perception of physical and mental function between adults 70 years of age or older who participated in a phase 2 cardiac rehabilitation program (CRP) (n = 32) and those who did not participate in a CRP (n = 33) after coronary artery bypass surgery (CABS). METHODS: In this two-group longitudinal comparative study, recovery outcomes measured at baseline (6 weeks) and 6 months after CABS were compared between older adults who participated and those who did not participate in a CRP. RESULTS: In study groups that were equivalent before the CRP, analysis of covariance (controlling for baseline scores) showed that 6 months after hospital discharge, those who participated in a CRP had greater lower extremity strength (F = 3.9; P =.04), greater ankle range of motion (F = 4.2; P =.02), better dynamic balance (F = 8.2; P =.003), better static balance (F = 3.3; P =.04), better gait (F = 4.7; P =.02), and perceptions of better physical function (F = 14.8; P =.00). The results remained the same when control was used for the effects of social support, self-efficacy, depression, comorbidity, cardiac functional status, and gender for all the variables except static balance. No difference related to perception of mental function was found between the study groups (F =.10; P =. 74). CONCLUSIONS: Participation in a CRP by older individuals improves lower extremity function (an important dimension in preventing disability) and perception of physical function. Cardiac rehabilitation programs can be used to optimize the recovery outcomes of older individuals after CABS. PMID- 15286529 TI - Strengthening the evidence base to support the use of cardiac rehabilitation with older individuals. PMID- 15286530 TI - Gender alters the high-density lipoprotein cholesterol response to cardiac rehabilitation. AB - PURPOSE: A reduced level of high-density lipoprotein cholesterol (HDL-C) is a powerful independent risk factor for the development and progression of coronary heart disease. This study assessed the effects of cardiac rehabilitation exercise training on HDL-C and other lipid subfractions, giving close attention to the role of gender and baseline values. METHODS: The study population consisted of 340 patients with coronary heart disease who enrolled in outpatient cardiac rehabilitation and completed 36 sessions of exercise over a 12-week period. With the National Cholesterol Education Panel ATP III guidelines used to create categories of HDL-C, patients were stratified at baseline into four subgroups: (1) males with high HDL-C, (2) males with low HDL-C, (3) women with high HDL-C, and (4) women with low HDL-C. RESULTS: Overall, women experienced a significantly greater improvement in HDL-C after exercise training than men (14% vs 7.1%; P <.0001). Among the patients with a high HDL-C at baseline, the women increased HDL-C by 8.4%, whereas there was no change (0.9%) in the men (P <.001 between groups). Additionally, the women with low HDL-C experienced a significantly greater improvement than the men (15.3% vs 11.5%, P <.03). CONCLUSIONS: The study results demonstrate that women experience a greater improvement in HDL-C with cardiac rehabilitation than men despite similar changes in fitness and body composition. Women, regardless of baseline HDL-C, demonstrated improvements in HDL-C, whereas only men with low HDL-C experienced an increase in HDL-C. These results describe a differing impact of cardiac rehabilitation on changes in HDL-C based on gender. Clinicians should consider the impact of gender when assessing an individual's risk factor goals and therapeutic options. PMID- 15286532 TI - Gender differences in lipids and lipoproteins after cardiac rehabilitation. PMID- 15286533 TI - Randomized trial of an individualized coronary risk factor intervention in patients from rural communities undergoing percutaneous coronary revascularization. PMID- 15286534 TI - Use of the 6-minute walk test for women with diastolic heart failure. PMID- 15286535 TI - Effect of exercise intensity on postexercise hypotension. PMID- 15286536 TI - Reductions in functional balance, coordination, and mobility measures among patients with stable chronic obstructive pulmonary disease. AB - PURPOSE: To compare measures of balance, coordination, and mobility between patients with chronic obstructive pulmonary disease (COPD) and healthy control subjects, and to determine whether differences in these measures are associated with measures of disease severity. METHODS: The subjects were divided into three groups: 15 patients with COPD who required the use of supplemental oxygen (WO), 15 patients with COPD who did not require the use of supplemental oxygen (NO), and 21 healthy control subjects (CO). The subjects performed spirometry and several measures of balance, coordination, and mobility including the Community Balance and Mobility Scale, the timed up and go test, the fast-gait speed test, posturography, and both a finger-to-nose test and a toe-tapping coordination test. Significance was set at an alpha less than 0.05. RESULTS: When control was used for age, significant differences were found between the WO group and the CO group for the finger-to-nose test, and for both the sway index and peak sway index for the eyes open, moving-platform test. Differences were found among all three groups for the Community Balance and Mobility Scale overall score. The scores for the WO group were significantly worse than for the NO group on the timed up and go and the fast-gait speed tests. Moderate correlation was found among all of the measures, demonstrating significant differences in forced expiratory volume in 1 second (FEV1), peak expiratory flow, and forced-expiratory volume. When controls were used for both age and FEV1, between-group differences disappeared. CONCLUSIONS: Patients with COPD exhibit deficiencies in functional balance, coordination, and mobility tasks associated with disease severity or differences in activity levels, but not in the requirement for supplemental oxygen. PMID- 15286537 TI - Early lactate clearance is associated with improved outcome in severe sepsis and septic shock. AB - OBJECTIVE: Serial lactate concentrations can be used to examine disease severity in the intensive care unit. This study examines the clinical utility of the lactate clearance before intensive care unit admission (during the most proximal period of disease presentation) as an indicator of outcome in severe sepsis and septic shock. We hypothesize that a high lactate clearance in 6 hrs is associated with decreased mortality rate. DESIGN: Prospective observational study. SETTING: An urban emergency department and intensive care unit over a 1-yr period. PATIENTS: A convenience cohort of patients with severe sepsis or septic shock. INTERVENTIONS: Therapy was initiated in the emergency department and continued in the intensive care unit, including central venous and arterial catheterization, antibiotics, fluid resuscitation, mechanical ventilation, vasopressors, and inotropes when appropriate. MEASUREMENTS AND MAIN RESULTS: Vital signs, laboratory values, and Acute Physiology and Chronic Health Evaluation (APACHE) II score were obtained at hour 0 (emergency department presentation), hour 6, and over the first 72 hrs of hospitalization. Therapy given in the emergency department and intensive care unit was recorded. Lactate clearance was defined as the percent decrease in lactate from emergency department presentation to hour 6. Logistic regression analysis was performed to determine independent variables associated with mortality. One hundred and eleven patients were enrolled with mean age 64.9 +/- 16.7 yrs, emergency department length of stay 6.3 +/- 3.2 hrs, and overall in-hospital mortality rate 42.3%. Baseline APACHE II score was 20.2 +/- 6.8 and lactate 6.9 +/- 4.6 mmol/L. Survivors compared with nonsurvivors had a lactate clearance of 38.1 +/- 34.6 vs. 12.0 +/- 51.6%, respectively (p =.005). Multivariate logistic regression analysis of statistically significant univariate variables showed lactate clearance to have a significant inverse relationship with mortality (p =.04). There was an approximately 11% decrease likelihood of mortality for each 10% increase in lactate clearance. Patients with a lactate clearance> or =10%, relative to patients with a lactate clearance <10%, had a greater decrease in APACHE II score over the 72-hr study period and a lower 60 day mortality rate (p =.007). CONCLUSIONS: Lactate clearance early in the hospital course may indicate a resolution of global tissue hypoxia and is associated with decreased mortality rate. Patients with higher lactate clearance after 6 hrs of emergency department intervention have improved outcome compared with those with lower lactate clearance. PMID- 15286538 TI - Utility of B-type natriuretic peptide for the evaluation of intensive care unit shock. AB - OBJECTIVES: Among patients with congestive heart failure, B-type natriuretic peptide measurement is useful to estimate filling pressures and to prognosticate adverse outcome. However, among critically ill intensive care unit patients with shock, the utility of B-type natriuretic peptide to assess cardiac hemodynamics or prognosis has not been explored. DESIGN: Clinical investigation. SETTING: Hospital. PATIENTS: Forty-nine patients with shock and indication for pulmonary artery catheterization. INTERVENTIONS: Analysis for B-type natriuretic peptide was performed on blood obtained at the time of catheter placement. MEASUREMENTS AND MAIN RESULTS: Correlations between B-type natriuretic peptide and pulmonary artery occlusion pressure as well as cardiac index were calculated using Spearman analysis. Mortality at the time of study completion was correlated with B-type natriuretic peptide values and Acute Physiology and Chronic Health Evaluation II scores, and logistic regression identified independent predictors of mortality. A wide range of B-type natriuretic peptide concentrations was seen in intensive care unit patients (<5 to >5000 pg/mL); only eight patients (16%) had normal B type natriuretic peptide concentrations. Log-transformed B-type natriuretic peptide concentrations did not correlate with interpatient cardiac index or pulmonary artery occlusion pressure (all p = not significant); however, a B-type natriuretic peptide <350 pg/mL had a negative predictive value of 95% for the diagnosis of cardiogenic shock. Median B-type natriuretic peptide concentrations were higher in those who died than those who survived (943 pg/mL vs. 378 pg/mL, p <.001). In multivariable analysis, a B-type natriuretic peptide concentration in the highest log-quartile was the strongest predictor of mortality (odds ratio = 4.50, 95% confidence interval = 1.87-99.0, p <.001). CONCLUSION: B-type natriuretic peptide concentrations are frequently elevated among critically ill patients in the intensive care unit and cannot be used as a surrogate for pulmonary artery catheterization. B-type natriuretic peptide concentrations in intensive care unit shock may provide powerful information for use in mortality prediction. PMID- 15286539 TI - Factors associated with nurse assessment of the quality of dying and death in the intensive care unit. AB - OBJECTIVE: To determine the feasibility of using nurse ratings of quality of dying and death to assess quality of end-of-life care in the intensive care unit and to determine factors associated with nurse assessment of the quality of dying and death for patients dying in the intensive care unit. DESIGN: Prospective cohort study. SETTING: Hospital intensive care unit. PATIENTS: 178 patients who died in an intensive care unit during a 10-month period at one hospital. INTERVENTIONS: Nurses completed a 14-item questionnaire measuring the quality of dying and death in the intensive care unit (QODD); standardized chart reviews were also completed. MEASUREMENTS AND MAIN RESULTS: Five variables were found to be associated with QODD scores. Higher (better) scores were significantly associated with having someone present at the time of death (p <.001), having life support withdrawn (p =.006), having an acute diagnosis such as intracranial hemorrhage or trauma (p =.007), not having cardiopulmonary resuscitation in the last 8 hrs of life (p <.001), and being cared for by the neurosurgery or neurology services (p =.002). Patient age, chronic disease, and Glasgow Coma Scale scores were not associated with the 14-item QODD. Using multivariate analyses, we identified three variables as independent predictors of the QODD score: a) not having cardiopulmonary resuscitation performed in the last 8 hrs of life; b) having someone present at the moment of death; and c) being cared for by neurosurgery or neurology services. CONCLUSIONS: Intensive care unit nurse assessment of quality of dying and death is a feasible method for obtaining quality ratings. Based on nurse assessments, this study provides evidence of some potential targets for interventions to improve the quality of dying for some patients: having someone present at the moment of death and not having cardiopulmonary resuscitation in the last 8 hrs of life. If nurse-assessed quality of dying is to be a useful tool for measuring and improving quality of end-of-life care, it is important to understand the factors associated with nurse ratings. PMID- 15286540 TI - Survival of critically ill patients hospitalized in and out of intensive care units under paucity of intensive care unit beds. AB - OBJECTIVE: The demand for intensive care beds far exceeds their availability in many European countries. Consequently, many critically ill patients occupy hospital beds outside intensive care units, throughout the hospital. The outcome of patients who fit intensive care unit admission criteria but are hospitalized in regular wards needs to be assessed for policy implications. The object was to screen entire hospital patient populations for critically ill patients and compare their 30-day survival in and out of the intensive care unit. DESIGN: Screening teams visited every hospital ward on four selected days in five acute care Israeli hospitals. The teams listed all patients fitting a priori developed study criteria. One-month data for each patient were abstracted from the medical records. SETTING: Five acute care Israeli hospitals. PATIENTS: All patients fitting a priori developed study criteria. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Survival in and out of the intensive care unit was compared for screened patients from the day a patient first met study criteria. Cox multivariate models were constructed to adjust survival comparisons for various confounding factors. The effect of intensive care unit vs. other departments was estimated separately for the first 3 days after deterioration and for the remaining follow-up time. Results showed that 5.5% of adult hospitalized patients were critically ill (736 of 13,415). Of these, 27% were admitted to intensive care units, 24% to specialized care units, and 49% to regular departments. Admission to an intensive care unit was associated with better survival during the first 3 days of deterioration, after we adjusted for age and severity of illness (p =.018). There was no additional survival advantage for intensive care unit patients (p =.9) during the remaining follow-up time. CONCLUSIONS: The early survival advantage in the intensive care unit suggests a window of critical opportunity for these patients. Under economic constraints and dearth of intensive care unit beds, increasing the turnover of patients in the intensive care unit, thus exposing more needy patients to the early benefit of treatment in the intensive care unit, may be advantageous. PMID- 15286541 TI - Open randomized phase II trial of an extracorporeal endotoxin adsorber in suspected Gram-negative sepsis. AB - OBJECTIVE: An initial phase II trial to investigate the safety and therapeutic effect of the endotoxin adsorber system EN 500 in septic patients suffering from presumed Gram-negative infection. DESIGN: Open, controlled, prospective, randomized, multiple-center, parallel-group clinical trial. SETTING: Intensive care units of 31 university-affiliated and community hospitals in Europe. PATIENTS: One hundred forty-five patients with a clinical diagnosis of severe sepsis or septic shock due to suspected Gram-negative infection. INTERVENTIONS: Patients were randomized to receive either standard therapy alone for sepsis (n = 76) or standard therapy plus extracorporeal endotoxin adsorption (n = 67) daily for the first 4 days following study entry. MEASUREMENTS AND MAIN RESULTS: The primary end point was the proportion of responders (defined as a decrease in Acute Physiology and Chronic Health Evaluation II score by > or =4 points from study entry to day 4). Secondary outcomes were the Sequential Organ Failure Assessment score and its components, length of intensive care unit stay, survival rate, and safety of the adsorber treatment. Patient characteristics at entry were well balanced between the two treatment groups, except for a higher Sequential Organ Failure Assessment score in the adsorber group. On all-subjects-treated analysis, 65% of the adsorber group were responders vs. 57% for the standard (p =.389). A planned interim analysis restricted further enrollment to patients with peritonitis, in whom a slightly higher proportion of responders was observed with the adsorber treatment (69%) vs. standard treatment (54%, p =.159). There were no differences in survival, but adsorption treatment in peritonitis patients was associated with trends toward a reduction in length of intensive care unit stay and a more rapid decline in plasma endotoxin concentrations. There was a significantly greater reduction in platelet count with the adsorber; however, this did not require extra treatment. CONCLUSIONS: The endotoxin adsorber system did not result in a significantly improved primary end point in patients with presumed Gram-negative sepsis. In patients with peritonitis, the adsorber treatment likewise did not result in significantly improved Acute Physiology and Chronic Health Evaluation II scores. There were no clinically important side effects. These results provide encouragement for further study of adsorber treatment in patients with high likelihood of Gram-negative sepsis (e.g., peritonitis). PMID- 15286542 TI - Diuretics and mortality in acute renal failure. AB - OBJECTIVE: According to recent research, diuretics may increase mortality in acute renal failure patients. The administration of diuretics in such patients has been discouraged. Our objective was to determine the impact of diuretics on the mortality rate of critically ill patients with acute renal failure. DESIGN: Prospective, multiple-center, multinational epidemiologic study. SETTING: Intensive care units from 54 centers and 23 countries. PATIENTS: Patients were 1,743 consecutive patients who either were treated with renal replacement therapy or fulfilled predefined criteria for acute renal failure. INTERVENTIONS: Three distinct multivariate models were developed to assess the relationship between diuretic use and subsequent mortality: a) a propensity score adjusted multivariate model containing terms previously identified to be important predictors of outcome; b) a new propensity score adjusted multivariate model; and c) a multivariate model developed using standard methods, compensating for collinearity. MEASUREMENTS AND MAIN RESULTS: Approximately 70% of patients were treated with diuretics at study inclusion. Mean age was 68 and mean Simplified Acute Physiology Score II was 47. Severe sepsis/septic shock (43.8%), major surgery (39.1), low cardiac output (29.7), and hypovolemia (28.2%) were the most common conditions associated with the development of acute renal failure. Furosemide was the most common diuretic used (98.3%). Combination therapy was used in 98 patients only. In all three models, diuretic use was not associated with a significantly increased risk of mortality. CONCLUSIONS: Diuretics are commonly prescribed in critically ill patients with acute renal failure, and their use is not associated with higher mortality. There is full equipoise for a randomized controlled trial of diuretics in critically ill patients with renal dysfunction. PMID- 15286543 TI - Effect of once-daily dosing vs. multiple daily dosing of tobramycin on enzyme markers of nephrotoxicity. AB - OBJECTIVE: To determine the incidence of nephrotoxicity of once-daily dosing (ODD) and multiple daily dosing (MDD) regimens of tobramycin in critically ill patients. DESIGN: Randomized, prospective clinical trial. SETTING: : Adult intensive care units at two university hospitals. PATIENTS: Fifty-eight critically ill patients with a suspected or documented aerobic Gram-negative infection. INTERVENTIONS: Patients were randomized to receive tobramycin by ODD (7 mg/kg) or MDD. Baseline urine aliquots and 24-hr urine collections were collected on days 3, 7, and 11 during therapy and on days 3, 7, and 11 following discontinuation of therapy for measurement of alanine aminopeptidase (AAP), N acetyl-beta-d-glucosaminidase (NAG), and creatinine. MEASUREMENTS AND MAIN RESULTS: Fifty-four patients were evaluable (ODD n = 25; MDD n = 29). The groups were similar with regard to demographic and clinical variables. The tobramycin dose was higher in the ODD group compared with the MDD group (425 +/- 122.5 mg vs. 312.8 +/- 116.6 mg, p <.001). Patients in the MDD group received a mean of 3.89 +/- 1.14 mg.kg(-1)day(-1) at intervals of 11.92 +/- 3.12 hrs. In the ODD group, patients had a higher measured creatinine clearance at the end of therapy compared with MDD group (70 +/- 18.6 vs. 64.8 +/- 17.5 mL/min, p =.047). Fewer patients in the ODD group developed nephrotoxicity than the MDD group (5 vs. 12, p =.142). Although there were increases in urinary enzymes in both treatment groups (AAP, 8.7 +/- 2.9 vs. 5.2 +/- 2.1 units/24 hrs, p <.01 MDD vs. ODD; NAG, 14.7 +/- 4.9 vs. 6.8 +/- 3.1, p <.01 MDD vs. ODD), the increases in the ODD group were significantly lower than in the MDD group. CONCLUSIONS: : The ODD tobramycin regimen appeared to be less nephrotoxic than the MDD regimen despite significantly higher doses. Tobramycin administered by ODD may be the preferred dosing method in selected critically ill medical patients to reduce the incidence and extent of renal damage. PMID- 15286544 TI - Unidentified acids of strong prognostic significance in severe malaria. AB - OBJECTIVE: To calculate, using the Stewart approach to acid-base disorders, the strong anion gap as an estimate for the contribution of unmeasured plasma anions other than lactate to the metabolic acidosis that characterizes severe falciparum malaria and to assess its relative prognostic significance. DESIGN: Cohort study. SETTING: The intensive care unit of an infectious diseases hospital in southern Vietnam. PATIENTS: Consecutive adult patients (n = 268) with severe falciparum malaria. INTERVENTIONS: The intervention was clinical management in a dedicated unit. We measured baseline venous lactate, electrolytes, biochemical variables, admission arterial blood pH, and gas tensions for calculation of the strong anion gap. MEASUREMENTS AND MAIN RESULTS: The mean (95% confidence interval) admission strong anion gap was 11.1 (10.4-11.9) mEq/L, compared with lactate (geometric mean, 95% confidence interval) at 2.9 (2.7-3.2) mmol/L. Strong anion gap had a high predictive value for mortality (area under the receiver operating characteristic curve 0.73 (95% confidence interval, 0.65-0.82), which was independent of plasma lactate and creatinine concentrations. Renal failure and hepatic dysfunction were both associated with, but were not the sole determinants of, high levels of strong anion gap. CONCLUSIONS: In severe malaria, unidentified anions other than lactate are the most important contributors to metabolic acidosis, a major cause of death. The strong anion gap is a powerful prognostic indicator in patients with severe malaria. PMID- 15286545 TI - A prospective, randomized, study comparing early percutaneous dilational tracheotomy to prolonged translaryngeal intubation (delayed tracheotomy) in critically ill medical patients. AB - OBJECTIVE: The timing of tracheotomy in patients requiring mechanical ventilation is unknown. The effects of early percutaneous dilational tracheotomy compared with delayed tracheotomy in critically ill medical patients needing prolonged mechanical ventilation were assessed. DESIGN: Prospective, randomized study. SETTING: Medical intensive care units. PATIENTS: One hundred and twenty patients projected to need ventilation >14 days. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Patients were prospectively randomized to either early percutaneous tracheotomy within 48 hrs or delayed tracheotomy at days 14-16. Time in the intensive care unit and on mechanical ventilation and the cumulative frequency of pneumonia, mortality, and accidental extubation were documented. The airway was assessed for oral, labial, laryngeal, and tracheal damage. Early group showed significantly less mortality (31.7% vs. 61.7%), pneumonia (5% vs. 25%), and accidental extubations compared with the prolonged translaryngeal group (0 vs. 6). The early tracheotomy group spent less time in the intensive care unit (4.8 +/- 1.4 vs. 16.2 +/- 3.8 days) and on mechanical ventilation (7.6 +/- 2.0 vs. 17.4 +/- 5.3 days). There was also significantly more damage to mouth and larynx in the prolonged translaryngeal intubation group. CONCLUSIONS: This study demonstrates that the benefits of early tracheotomy outweigh the risks of prolonged translaryngeal intubation. It gives credence to the practice of subjecting this group of critically ill medical patients to early tracheotomy rather than delayed tracheotomy. PMID- 15286546 TI - Neutrophil elastase inhibition in acute lung injury: results of the STRIVE study. AB - OBJECTIVE: Neutrophil elastase is believed to be an important mediator of acute lung injury. Sivelestat (ONO-5046, Elaspol) is a small molecular weight inhibitor of neutrophil elastase. The primary objectives of this study were to determine whether sivelestat would reduce 28-day all-cause mortality or increase the number of ventilator-free days (days alive and free from mechanical ventilation from day 1 to day 28) compared with placebo in mechanically ventilated patients with acute lung injury. DESIGN: Multiple-center, double-blind, placebo-controlled trial administering a continuous infusion of sivelestat at a dose of 0.16 mg.kg(-1)hr( 1). SETTING: One hundred and five institutions in the United States, Canada, Belgium, Spain, Australia, and New Zealand. PATIENTS: A total of 492 mechanically ventilated patients with acute lung injury. INTERVENTIONS: Patients were randomized in a 1:1 fashion to sivelestat or placebo. Study drug was administered as a continuous infusion for the duration of mechanical ventilation plus 24 hrs for a maximum of 14 days. All patients were managed using low tidal volume mechanical ventilation. MEASUREMENTS AND MAIN RESULTS: The study was stopped prematurely at the recommendation of an external Data and Safety Monitoring Board, which noted a negative trend in long-term mortality rate. Final analysis revealed no effect of sivelestat on the primary end points of ventilator-free days (day 1-day 28) or 28-day all-cause mortality. There were 64 deaths in each treatment group within the 28-day study period, and the mean number of ventilator free days was 11.4 and 11.9 in the sivelestat and placebo treatment groups, respectively (p =.536). There was no evidence of effect on measures of pulmonary function, including Pao2/Fio2, static lung compliance, and time to meeting weaning criteria. There was no difference in adverse events or serious adverse events between treatment groups. A comparison of the Kaplan-Meier 180-day survival curves showed no difference between treatment groups (p =.102), but there was an increase in 180-day all-cause mortality in the sivelestat treatment group compared with the placebo group (p =.006). CONCLUSIONS: Intravenous sivelestat had no effect on 28-day all-cause mortality or ventilator-free days in a heterogeneous acute lung injury patient population managed with low tidal volume mechanical ventilation. PMID- 15286547 TI - Randomized, controlled trial of immediate versus delayed goal-directed ultrasound to identify the cause of nontraumatic hypotension in emergency department patients. AB - OBJECTIVE: We examined a physician-performed, goal-directed ultrasound protocol for the emergency department management of nontraumatic, symptomatic, undifferentiated hypotension. DESIGN: Randomized, controlled trial of immediate vs. delayed ultrasound. SETTING: Urban, tertiary emergency department, census >100,000. PATIENTS: Nontrauma emergency department patients, aged >17 yrs, and initial emergency department vital signs consistent with shock (systolic blood pressure <100 mm Hg or shock index >1.0), and agreement of two independent observers for at least one sign and symptom of inadequate tissue perfusion. INTERVENTIONS: Group 1 (immediate ultrasound) received standard care plus goal directed ultrasound at time 0. Group 2 (delayed ultrasound) received standard care for 15 mins and goal-directed ultrasound with standard care between 15 and 30 mins after time 0. MEASUREMENTS AND MAIN RESULTS: Outcomes included the number of viable physician diagnoses at 15 mins and the rank of their likelihood of occurrence at both 15 and 30 mins. One hundred eighty-four patients were included. Group 1 (n = 88) had a smaller median number of viable diagnoses at 15 mins (median = 4) than did group 2 (n = 96, median = 9, Mann-Whitney U test, p <.0001). Physicians indicated the correct final diagnosis as most likely among their viable diagnosis list at 15 mins in 80% (95% confidence interval, 70-87%) of group 1 subjects vs. 50% (95% confidence interval, 40-60%) in group 2, difference of 30% (95% confidence interval, 16-42%). CONCLUSIONS: Incorporation of a goal-directed ultrasound protocol in the evaluation of nontraumatic, symptomatic, undifferentiated hypotension in adult patients results in fewer viable diagnostic etiologies and a more accurate physician impression of final diagnosis. PMID- 15286548 TI - Relationship of continuous infusion lorazepam to serum propylene glycol concentration in critically ill adults. AB - OBJECTIVES: The primary objective was to evaluate the relationship between high dose lorazepam and serum propylene glycol concentrations. Secondary objectives were a) to document the occurrence of propylene glycol accumulation associated with continuous high-dose lorazepam infusion; b) to assess the relationship between lorazepam dose, serum propylene glycol concentrations, and propylene glycol accumulation; and c) to assess the relationship between the osmol gap and serum propylene glycol concentrations. DESIGN: Prospective, observational study. SETTING: Tertiary care, medical intensive care unit. PATIENTS: Nine critically ill adults receiving high-dose lorazepam (> or =10 mg/hr) infusion. INTERVENTIONS: Cumulative lorazepam dose (mg/kg) and the rate of infusion (mg.kg( 1).hr(-1)) were monitored from initiation of lorazepam infusion until 24 hrs after discontinuation of the high-dose lorazepam infusion. Serum osmolarity was collected at 48 hrs into the high-dose lorazepam infusion and daily thereafter. Serum propylene glycol concentrations were drawn at 48 hrs into the high-dose lorazepam infusion, and the presence of propylene glycol accumulation, as evidenced by a high anion gap (> or =15 mmol/L) metabolic acidosis with elevated osmol gap (> or =10 mOsm/L), was assessed at that time. MEASUREMENTS AND MAIN RESULTS: The mean cumulative high-dose lorazepam received and mean high-dose lorazepam infusion rate were 8.1 mg/kg (range, 5.1-11.7) and 0.16 mg.kg(-1).hr ( 1)(range, 0.11-0.22), respectively. A significant correlation between high-dose lorazepam infusion rate and serum propylene glycol concentrations was observed (r =.557, p =.021). Osmol gap was the strongest predictor of serum propylene glycol concentrations (r =.804, p =.001). Propylene glycol accumulation was observed in six of nine patients at 48 hrs. No significant correlation between duration of lorazepam infusion and serum propylene glycol concentrations was observed (p =.637). CONCLUSION: Propylene glycol accumulation, as reflected by a hyperosmolar anion gap metabolic acidosis, was observed in critically ill adults receiving continuous high-dose lorazepam infusion for > or =48 hrs. Study findings suggest that in critically ill adults with normal renal function, serum propylene glycol concentrations may be predicted by the high-dose lorazepam infusion rate and osmol gap. PMID- 15286549 TI - Expression and secretion of procalcitonin and calcitonin gene-related peptide by adherent monocytes and by macrophage-activated adipocytes. AB - OBJECTIVE: To explore the roles of peripheral blood mononuclear cells (PBMCs) and PBMC-derived macrophages in sepsis-related increased procalcitonin and calcitonin gene-related peptide (CGRP) I production. DESIGN: Prospective, in vitro primary human cell culture study and human tissue samples gene expression analysis. SETTING: University hospital research laboratories. PATIENTS: Cells from healthy donors and septic patients. INTERVENTIONS: PBMCs were obtained from healthy donors. Isolation of pure monocyte cultures was performed by magnetic depletion of nonmonocyte cells from PBMCs. Adipose tissue biopsies and circulating leukocytes were collected from septic patients. Expressions of calcitonin messenger RNA and CGRP I messenger RNA were analyzed using reverse transcriptase polymerase chain reaction and quantitative real-time polymerase chain reaction. Supernatant procalcitonin and CGRP protein content were determined by ultrasensitive chemiluminometric and radioimmunoassays, respectively. MEASUREMENTS AND MAIN RESULTS: PBMCs expressed and secreted procalcitonin and CGRP within 3-5 hrs after adherence to endothelial cells or plastic surfaces. This induction was transient, as it was not detectable after 18 hrs. No calcitonin or CGRP I messenger RNA was observed in leukocytes obtained from septic patients with markedly increased serum procalcitonin concentrations. Stimulation with cytokines, endotoxin, or Escherichia coli did not induce expression of calcitonin and CGRP I messenger RNA in PBMC-derived macrophages. However, inflammatory factors released from activated macrophages induced a marked expression of procalcitonin and CGRP in co-cultured human adipocytes. CONCLUSIONS: The adhesion-induced, transient expression and secretion of procalcitonin and CGRP in vitro may play an important role during monocyte adhesion and migration in vivo. PBMC-derived macrophages may contribute to the marked increase in circulating procalcitonin by recruiting parenchymal cells within the infected tissue, as exemplified with adipocytes. PMID- 15286550 TI - A potential role of hyperbaric oxygen exposure through intestinal nuclear factor kappaB. AB - OBJECTIVE: Recent studies have demonstrated the therapeutic effectiveness and pharmacologic mechanisms of hyperbaric oxygen therapy (HBOT) in the treatment of a systemic shock state. To elucidate the in vivo role of HBOT during sepsis, we evaluated the effects of HBOT on intestinal mucosal injury and bacterial translocation after lipopolysaccharide challenge. DESIGN: Experimental study. SETTING: First Department of Surgery and Division of Emergency Care, Kagoshima University School of Medicine, Kagoshima, Japan. SUBJECTS: : Male rats were treated with lipopolysaccharide by an intraperitoneal route or with lipopolysaccharide and HBOT. INTERVENTIONS: The survival rate, small intestinal tissue damage, and bacterial translocation in the HBOT-treated group were compared with those in the untreated group. Moreover, plasma tumor necrosis factor-alpha and nitrite/nitrate concentrations, inducible nitric oxide synthase and myeloperoxidase activities, and nuclear factor-kappaB in ileal mucosa were investigated. HBOT was initiated 3 hrs after lipopolysaccharide challenge and administered as 100% oxygen, at 2.53 x 10 kPa (2.5 atm absolute), for 60 mins. MEASUREMENTS AND MAIN RESULTS: When a sublethal dose of lipopolysaccharide (24 mg/kg) was given, the survival rate was much better in the HBOT-treated group (75%) than in the untreated group (33%). HBOT given 3 hrs after lipopolysaccharide injection (10 mg/kg) also lessened the histologic tissue damage of the terminal ileum and the incidence and magnitude of bacterial translocation to mesenteric lymph nodes at 24 hrs after the lipopolysaccharide injection. Moreover, HBOT was able to reduce mucosal inducible nitric oxide synthase and myeloperoxidase activities and plasma nitrite/nitrate concentrations but not serum tumor necrosis factor-alpha concentrations. Immunohistochemical examination revealed that HBOT specifically modified the mucosal nuclear factor kappaB activation within 4-6 hrs after the injection. CONCLUSIONS: HBOT performed 3 hrs after lipopolysaccharide challenge alleviates intestinal barrier dysfunction and improves survival rates. Herein, we propose one possible mechanism for these beneficial effects: HBOT can modify the nuclear factor-kappaB activation in the intestinal mucosa and attenuate the sequential nitric oxide overproduction and myeloperoxidase activation. Consequently, bacterial translocation could be potentially decreased. We believe that the present study should lead to an improved understanding of HBOT's potential role in sepsis. PMID- 15286551 TI - Mechanisms of postburn intestinal barrier dysfunction in the rat: roles of epithelial cell renewal, E-cadherin, and neutrophil extravasation. AB - OBJECTIVE: Our group has previously shown that the intestinal epithelium exhibits increased postburn barrier permeability and bacterial translocation associated with deranged neutrophil activity. The purpose of this investigation is to explore possible underlying intestinal structural mechanisms, leading to those functional changes with emphasis on (1) neutrophil influx and extravasation in the intestinal lamina propria 1-3 days after burn and (2) enterocyte proliferation, migration, apoptosis, and E-cadherin junctional epithelium levels 3 days after burn. DESIGN: Freshly isolated ileum specimens were quick frozen, then cut by a cryostat into 30-micron-thick sections. Sections from day 1 postburn rats were immunostained with (1) anti-granulocyte or anti-elastase antibodies to assess neutrophil influx or (2) combined anti-granulocyte and anti von Willebrand factor double immunolabeling to compare levels of neutrophil extravasation. Sections from day 3 postburn rats were immunostained with (1) bromodeoxyuridine immunohistochemistry 1, 3, 6, or 18 hrs after bromodeoxyuridine injection to assess enterocyte proliferation and migration, (2) cytokeratin-18 M30-immunohistochemistry to compare levels of enterocyte apoptosis, and (3) E cadherin immunohistochemistry to compare junctional E-cadherin integrity. Ileal myeloperoxidase activity and bacterial translocation of Enterococcus faecalis were assessed biochemically and by E. faecalis-specific bacterial cultures, respectively, in day 3 postburn rats. SETTING: : Research laboratories in a medical center and an academic institution. SUBJECTS: Male Sprague-Dawley rats given sham treatment or treatment as a burn model with full-thickness skin scald over 30% total body surface area. CONCLUSIONS: We report (1) increased levels of neutrophil influx and extravasation in villi lamina propriae, including elastase positive cells (postburn day 1), (2) heightened levels of intestinal myeloperoxidase activity (postburn day 3), (3) decreased levels of epithelial cell proliferation, migration, and E-cadherin (postburn day 3), and (4) increased enterocyte apoptosis and E. faecalis bacterial translocation (postburn day 3). Based on these structural and functional abnormalities, we propose a mechanism for burn injury-related intestinal barrier dysfunction that includes increased trans- and para-cellular leakage caused by impaired enterocyte renewal and decreased junctional E-cadherin levels subsequent to increased neutrophil influx and extravasation within the villus lamina propria microenvironment. PMID- 15286552 TI - Escherichia coli pneumonia enhances granulopoiesis and the mobilization of myeloid progenitor cells into the systemic circulation. AB - OBJECTIVE: The process by which hematopoietic tissues respond to a pulmonary infection remains poorly understood. This study investigated the potential role of lung-derived granulopoietic cytokines in facilitating this response. DESIGN: Laboratory investigation. SETTING: University laboratory. SUBJECTS: Male Balb/c mice. INTERVENTIONS: Mice were challenged with intratracheal Escherichia coli or granulocyte colony-stimulating factor (G-CSF). Bone marrow cells were isolated from normal mice and treated in vitro with G-CSF. MEASUREMENTS AND MAIN RESULTS: Bronchoalveolar lavage fluid concentrations of G-CSF, macrophage inflammatory protein-2, and keratinocyte-derived chemokine were elevated 3 and 6 hrs after intratracheal E. coli. The increases in intrapulmonary G-CSF and keratinocyte derived chemokine were associated with increases of their concentrations in the plasma. The numbers of granulocyte-macrophage colony forming units in bone marrow, spleen, and blood were increased 48 hrs after intratracheal E. coli or G CSF. In addition, plasma G-CSF and the number of progenitor cells (lin-ckit+Sca 1(-)) in the blood were increased at 30 mins and 48 hrs, respectively, following intratracheal G-CSF. Signal transducer and activator of transcription-3 in bone marrow cells was activated following intratracheal E. coli or G-CSF in addition to activation by in vitro G-CSF stimulation. CONCLUSIONS: During pulmonary infection, locally produced cytokines enter the circulation and may play an important role in initiating a granulopoietic response. PMID- 15286553 TI - Delayed administration of human inter-alpha inhibitor proteins reduces mortality in sepsis. AB - OBJECTIVE: We have recently shown that administration of human inter-alpha inhibitor proteins (IalphaIp) very early after the onset of sepsis maintains cardiovascular stability and reduced mortality. However, it remains unknown whether injection of IalphaIp at later time points of sepsis has any beneficial effects. We therefore hypothesized that IalphaIp and its active component bikunin are reduced in sepsis and that the delayed administration of IalphaIp also improves survival rate. DESIGN: : Prospective, controlled, and randomized animal study. SETTING: A research institute laboratory. SUBJECTS: : Male adult Sprague Dawley rats. INTERVENTIONS: Rats were subjected either to polymicrobial sepsis by cecal ligation and puncture (CLP) or to sham operation followed by the administration of normal saline solution (i.e., fluid resuscitation). MEASUREMENTS AND MAIN RESULTS: : Bikunin gene expression in the liver was measured by reverse transcription polymerase chain reaction. Plasma concentrations of IalphaIp were determined by Western blot at 5 and 20 hrs after CLP. IalphaIp clearance was assessed by injecting radioactive IalphaIp at 12 hrs post-CLP, and the half-life was determined. In addition, IalphaIp (30 mg/kg of body weight) or vehicle was administered at 1, 5, or 10 hrs (single treatment) or at both 10 and 20 hrs (double treatment) post-CLP. The necrotic cecum was excised at 20 hrs post-CLP, and 10-day survival was recorded. The results indicate that bikunin gene expression decreased significantly at 20 hrs post-CLP. Moreover, IalphaIp concentrations decreased significantly at 5 and 20 hrs post-CLP, and its half-life increased from 5.6 +/- 0.3 hrs to 11.8 +/- 2.7 hrs (p <.05), suggesting down-regulation of IalphaIp in sepsis despite the decreased clearance. Administration of IalphaIp at 1 hr post-CLP improved the survival rate from 50% to 92% (p <.05), whereas there was no significant improvement when IalphaIp was administrated at 5 or 10 hrs post-CLP. However, double injection of IalphaIp at 10 and 20 hrs post-CLP (i.e., severe sepsis) increased the survival rate from 44% to 81% (p <.05). CONCLUSION: Since delayed but repeated administration of human IalphaIp improves survival after CLP, this compound appears to be a useful agent for the treatment of severe sepsis. PMID- 15286554 TI - Tumor necrosis factor-alpha is associated with early postresuscitation myocardial dysfunction. AB - OBJECTIVE: Left ventricular dysfunction after successful cardiopulmonary resuscitation contributes to early death following resuscitation. The stress induced proinflammatory cytokines, particularly tumor necrosis factor-alpha and interleukin-1beta, are known to depress myocardial function. We hypothesized that tumor necrosis factor-alpha and interleukin-1beta, synthesized and released in response to the stress of global ischemia accompanying cardiac arrest, play a role in development of postresuscitation left ventricular dysfunction. METHODS: Hemodynamic variables, tumor necrosis factor-alpha , interleukin-1beta, interleukin-6 (enzyme-linked immunosorbent assay method), and ionized calcium were measured in ten anesthetized swine before and after 7 mins of cardiac arrest and during the early postresuscitation period (60-90 mins). RESULTS: Tumor necrosis factor-alpha increased three-fold within 15 mins of restoration of circulation and remained elevated throughout the observation period. A significant negative correlation was observed between tumor necrosis factor-alpha and left ventricular systolic change in pressure over time (r = -.54, p <.001). Interleukin-1beta was undetectable before and after resuscitation, and interleukin-6 was detectable in only two animals after resuscitation. Although a significant decline in ionized calcium was observed and correlated with left ventricular systolic change in pressure over time, an independent role for ionized calcium in postresuscitation left ventricular dysfunction was not demonstrated. CONCLUSION: Tumor necrosis factor-alpha increases during the early postresuscitation period and may play a role in postresuscitation myocardial dysfunction. PMID- 15286555 TI - Role of Toll-like receptor 4 on pancreatic and pulmonary injury in a mice model of acute pancreatitis associated with endotoxemia. AB - OBJECTIVE: Infection of pancreatic necrosis is a severe complication of acute pancreatitis. Because Toll-like receptor 4 (TLR4) has been identified as a receptor necessary to transduct the signal of bacteria-derived lipopolysaccharide into cells, we investigated the role of TLR4 on pancreatic and pulmonary injury in acute pancreatitis and acute pancreatitis associated with endotoxemia in wild type and TLR4-deficient mice. DESIGN: Laboratory investigation. SETTING: University laboratory. SUBJECTS: Heterozygous TLR4 mice. INTERVENTIONS: Mice were injected intraperitoneally with a supramaximal dose of cerulein each hour for 10 hrs. To mimic infection, additional groups of mice were injected with lipopolysaccharide in the presence or absence of cerulein injections. MEASUREMENTS AND MAIN RESULTS: The severity of acute pancreatitis was assessed by serum amylase activity, pancreatic edema, acinar cell necrosis, and pancreas myeloperoxidase activity. Lung injury was quantitated by lung microvascular permeability and lung myeloperoxidase activity. Injections of cerulein induced an edematous pancreatitis that was of similar severity in wild-type and TLR4 deficient mice. Lipopolysaccharide alone had no toxic effect on pancreas and lungs and did not worsen the pancreatic injury induced by cerulein in wild-type and TRL4-deficient mice. In contrast, lipopolysaccharide worsened pancreatitis associated lung injury, and the deficiency in TLR4 fully prevented this aggravation. CONCLUSIONS: TLR4 may not play a role in the pancreatitis-associated lung injury but participates in the pulmonary injury mediated by endotoxemia. PMID- 15286556 TI - Time-dependent mitochondrial-mediated programmed neuronal cell death prolongs survival in sepsis. AB - OBJECTIVE: To investigate whether apoptosis is a possible mechanism of brain dysfunction occurring in septic syndrome. DESIGN: Experimental prospective study. SETTING: Laboratory of Surgical Research at the University of Athens. SUBJECTS: Male pathogen-free Wistar rats. INTERVENTIONS: Rats (n = 112) were subjected to sepsis by cecal ligation and puncture. Sham-operated animals (n = 40) underwent the same procedure but without ligation or puncture. Septic animals were either randomly divided (n = 62) in six groups and studied at 6, 12, 24, 36, 48, and 60 hrs after the operation or monitored (n = 50) for 48 hrs as a survival study group. Sham-operated animals were killed at 6, 12, 24, 36, 48, and 60 hrs after the procedure. Brain and cecum were then removed and postfixed in paraffin sections. Apoptosis was evaluated by light microscopy in hematoxylin and eosin stained specimens and by transmission electron microscopy. In paraffin-embedded sections, immunostaining for bax, bcl-2, cytochrome c, and caspase-8 was done. MEASUREMENTS AND MAIN RESULTS: In septic rats, increased apoptosis was detected in neurons of the CA1 region of the hippocampus, in choroid plexus, and in Purkinje cells of the cerebellum. Bax immunopositivity was found decreased after the septic insult (p =.03). Bax immunoreactivity was altered as the septic syndrome evolved; it was up-regulated in the early stages (6-12 hrs) and progressively decreased in the late phases (p =.001). Cytochrome c presented a similar regional pattern of expression and was found to be the sole gene marker carrying an independent prognostic role (p =.03). Both bcl-2 and caspase-8 expression remained at constant levels at all times evaluated. CONCLUSIONS: There is evidence that more neurons undergo apoptosis during sepsis than in normal brain tissue in certain sites where the blood-brain barrier is compromised. In this phenomenon, mitochondrial gene regulators such as bax and products such as cytochrome c seem to play important regulating and prognostic roles, respectively. PMID- 15286557 TI - Fluid overload before continuous hemofiltration and survival in critically ill children: a retrospective analysis. AB - OBJECTIVE: Continuous venovenous hemofiltration (CVVH) is used for renal replacement and fluid management in critically ill children. A previous small study suggested that survival was associated with less percent fluid overload (%FO) in the intensive care unit (ICU) before hemofiltration. We reviewed our experience with a large series of pediatric CVVH patients to evaluate factors associated with outcome. DESIGN: Retrospective chart review. SETTING: Tertiary children's hospital. PATIENTS: CVVH pediatric ICU patients from November 1997 to January 2003. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: %FO was defined as total fluid input minus output (up to 7 days before CVVH for both hospital stay and ICU stay) divided by body weight. One hundred thirteen patients received CVVH; 69 survived (61%). Multiple organ dysfunction syndrome (MODS) was present in 103 patients; 59 survived (57%). Median patient age was 9.6 yrs (25th, 75th percentile: 2.5, 14.3). Median %FO was significantly lower in survivors vs. nonsurvivors for all patients (7.8% [2.0, 16.7] vs. 15.1% [4.9, 25.9]; p =.02] and in patients with > or =3-organ MODS (9.2% [5.1,16.7] vs. 15.5% [8.3, 28.6]; p =.01). The Pediatric Risk of Mortality Score III at CVVH initiation also was associated with survival in these groups, but by multivariate analysis, %FO was independently associated with survival in patients with > or =3-organ MODS (p =.01). CONCLUSIONS: Survival in critically ill children receiving CVVH in this large series was higher than in previous reports. CVVH survival may be associated with less %FO in patients with > or =3-organ MODS. Prospective studies are necessary to determine whether earlier use of CVVH to control fluid overload in critically ill children can improve survival. PMID- 15286558 TI - Use of recombinant tissue plasminogen activator in children with meningococcal purpura fulminans: a retrospective study. AB - OBJECTIVE: Meningococcal disease causes septic shock with associated disseminated intravascular coagulation and hemorrhagic skin necrosis. In severe cases, widespread vascular thrombosis leads to gangrene of limbs and digits and contributes to morbidity and mortality. Uncontrolled case reports have suggested that thrombolytic therapy may prevent some complications, and the use of tissue plasminogen activator (t-PA) has been widespread. Our aim was to summarize the clinical outcome and adverse effects where systemic t-PA has been used to treat children with fulminant meningococcemia. DESIGN: International, multiple-center, retrospective, observational case note study between January 1992 and June 2000. SETTING: Twenty-four different hospitals in seven European countries and Australia. PATIENTS: A total of 62 consecutive infants and children with severe meningococcal sepsis in whom t-PA was used for the treatment of predicted amputations and/or refractory shock (40 to treat severe ischemia, 12 to treat shock, and ten to treat both). INTERVENTIONS: t-PA was administered with a median dose of 0.3 mg.kg(-1).hr(-1) (range, 0.008-1.13) and a median duration of 9 hrs (range, 1.2-83). MAIN RESULTS: Twenty-nine of 62 patients died (47%; 95% confidence interval, 28-65). Seventeen of 33 survivors had amputations (11 below knee/elbow or greater loss; six less severe). In 12 of 50 patients to whom t-PA was given for imminent amputation, no amputations were observed. Five developed intracerebral hemorrhages (five of 62, 8%; 95% confidence interval, 0.5-16). Of these five, three died, one developed a persistent hemiparesis, and one recovered completely. CONCLUSIONS: The high incidence of intracerebral hemorrhage in our study raises concerns about the safety of t-PA in children with fulminant meningococcemia. However, due to the absence of a control group in such a severe subset of patients, whether t-PA is beneficial or harmful cannot be answered from the unrestricted use of the drug that is described in this report. Our experience highlights the need to avoid strategies that use experimental drugs in an uncontrolled fashion and to participate in multiple-center trials, which are inevitably required to study rare diseases. PMID- 15286559 TI - Challenges in end-of-life care in the ICU: statement of the 5th International Consensus Conference in Critical Care: Brussels, Belgium, April 2003: executive summary. AB - OBJECTIVE: The purpose of the conference was to provide clinical practice guidance in end-of-life care in the ICU via answers to previously identified questions relating to variability in practice, inadequate predictive models for death, elusive knowledge of patient preferences, poor communication between staff and surrogates, insufficient or absent training of healthcare providers, the use of imprecise and insensitive terminology and incomplete documentation in the medical record. PARTICIPANTS: Presenters and jury were selected by the sponsoring organizations (American Thoracic Society, European Respiratory Society, European Society of Intensive Care Medicine, Society of Critical Care Medicine, Societe de Reanimation de Langue Francaise). Presenters were experts on the question they addressed. Jury members were general intensivists without special expertise in the areas considered. Experts presented in an open session to jurors and other healthcare professionals. EVIDENCE: Experts prepared review papers on their specific topics in advance of the conference for the jury's reference in developing the consensus statement. CONSENSUS PROCESS: Jurors heard experts' presentations over 2 days and asked questions of the experts during the open sessions. Jury deliberation with access to the review papers occurred for 2 days following the conference. A writing committee drafted the consensus statement for review by the entire jury. The 5 sponsoring organizations reviewed the document and suggested revisions to be incorporated into the final statement. CONCLUSIONS: Strong recommendations for research to improve end-of-life care were made. The jury advocates a shared approach to end-of-life decision-making involving the caregiver team and patient surrogates. Respect for patient autonomy and the intention to honor decisions to decline unwanted treatments should be conveyed to the family. The process is one of negotiation, and the outcome will be determined by the personalities and beliefs of the participants. Ultimately, it is the attending physician's responsibility, as leader of the team, to decide on the reasonableness of the planned action. If a conflict cannot be resolved, an ethics consultation may be helpful. The patient must be assured of a pain-free death. The jury subscribes to the moral and legal principles that prohibit administering treatments specifically designed to hasten death. The patient must be given sufficient analgesia to alleviate pain and distress; if such analgesia hastens death, this "double-effect" should not detract from the primary aim to ensure comfort. PMID- 15286560 TI - Lac-time? PMID- 15286561 TI - Evolution of B-type natriuretic peptide in evaluation of intensive care unit shock. PMID- 15286562 TI - Nurse-assessed tool for evaluating death in the intensive care unit. PMID- 15286563 TI - The survival benefit of intensive care. PMID- 15286564 TI - Extracorporeal endotoxin adsorption in septic patients--does it work? PMID- 15286565 TI - Acute renal failure and diuretics: propensity, equipoise, and the need for a clinical trial. PMID- 15286566 TI - Unidentified acids in severe malaria: lessons for critical care. PMID- 15286567 TI - Early tracheostomy--has its time arrived? PMID- 15286568 TI - Sonographic assessment of the hypotensive patient: is this Jones a winner? PMID- 15286569 TI - Medication vehicle injury--using the proper restraint? PMID- 15286570 TI - Procalcitonin mode of action: new pieces in a complex puzzle. PMID- 15286571 TI - Multiple converging mechanisms for postburn intestinal barrier dysfunction. PMID- 15286572 TI - Programmed neuronal death in sepsis: caught in a crossfire or a planned sacrifice? PMID- 15286573 TI - Fluid overload in multiple organ dysfunction syndrome: a prediction of survival. PMID- 15286574 TI - Recombinant tissue plasminogen activator in children with meningococcal purpura fulminans--role uncertain. PMID- 15286575 TI - Guidelines on critical care services and personnel--meeting the manpower needs. PMID- 15286577 TI - Pacemakers and implantable cardioverter defibrillators. PMID- 15286580 TI - [The SOR program of guidelines for clinical practice. Palliative chemotherapy as first line treatment of metastatic colorectal cancer, 2003 update]. PMID- 15286592 TI - Type 2 diabetes in youth. PMID- 15286594 TI - Designer disasters, a legacy to leave behind. PMID- 15286595 TI - Lessons from North Carolina. PMID- 15286596 TI - Physician preparedness for bioterrorism recognition and response: a Utah-based needs assessment. AB - The medical community accepts that they have a need to be prepared to recognize and respond to bioterrorism events. A needs assessment conducted among physicians throughout the state of Utah provided insights into their perceived training needs and preferred methods of learning. Physicians have many competing demands on their time and tailored educational offering can increase the intended audience's acceptance and learning. PMID- 15286597 TI - Planning for hospital emergency mass-casualty decontamination by the US Department of Veterans Affairs. PMID- 15286598 TI - Triage of a febrile patient with a rash: a comparison of chickenpox, monkeypox, and smallpox. AB - The immediate and correct recognition of an infectious exanthema can be aided with a focused history and minor assessment. False alarms can consume health care resources and create unnecessary anxiety. Clinicians can use specific references to not only help with educating staff, but to ensure a more accurate diagnosis and trigger notification of appropriate infectious disease protocols. The authors recommend that al emergency departments have a process in place to immediately isolate suspicious cases until a more thorough medial workup can be performed. PMID- 15286599 TI - Large-scale urban disaster drill involving an explosion: lessons learned by an academic medical center. AB - Disaster drills are an effective way to test a hospital's preparedness for real life disasters, but an extensive amount of coordination and time is necessary to host a successful drill with a large number of victims. The lessons learned in this drill include a number of planning, education, orientation, and follow-up issues. It is not realistic to believe that a drill can be perfectly planned and practiced; therefore each drill provides another opportunity to improve on past experience. PMID- 15286610 TI - Simply secure. PMID- 15286611 TI - Perseverance pays off. PMID- 15286612 TI - Paybacks from way back: memorable moves by funny people. PMID- 15286613 TI - The ins and outs of outsourcing. PMID- 15286614 TI - The Heimlich in near-drownings. PMID- 15286617 TI - Articulating knee injuries: placing proper emphasis on the recognition & stabilization of severely dislocated knees. PMID- 15286618 TI - EMS armor: PPE is more than BSI protection. PMID- 15286619 TI - HIV: test results in minutes: reviewing the prehospital occupational risk of acquiring HIV & the availability & value of rapid post-exposure testing. PMID- 15286620 TI - Exertional hyponatremia: the other heat-related emergency. PMID- 15286621 TI - Road safety vehicle simulator--just like the real thing! PMID- 15286622 TI - Kevlar for your feet: magnum's new uniform boot coddles & protects. PMID- 15286623 TI - Wireless broadband lives! PMID- 15286625 TI - Beer over brawn. PMID- 15286624 TI - Surprise! surprise! surprise!... PMID- 15286626 TI - Politics & Turf wars may continue to cost lives. PMID- 15286627 TI - Boldness, but with realism. PMID- 15286628 TI - Expose your clinical thinking. PMID- 15286629 TI - IPO plans sour on both sides of the pond. PMID- 15286630 TI - Patent drop reveals pressures on industry. PMID- 15286631 TI - Japanese firms broaden investment focus to early biotech. PMID- 15286632 TI - India aims to become the main bioinformatics hub. PMID- 15286634 TI - Biotech holds its own in Q2. PMID- 15286635 TI - Fast track to MS drug. PMID- 15286637 TI - Negative fallout from public sentiment in Japan. PMID- 15286639 TI - Debunking technophobes. PMID- 15286640 TI - Bioethic$ Inc. PMID- 15286641 TI - Biotech IPOs--flop or pop? PMID- 15286643 TI - Redesigner drugs. AB - Drug development is a risky business, and the final product can have serious, sometimes deadly flaws. But by focusing on fixing those flaws, companies are catapulting themselves to profitability. PMID- 15286644 TI - Zooming in and out with quantum dots. PMID- 15286645 TI - Tagging all genes. PMID- 15286646 TI - Editing the message from A to I. PMID- 15286647 TI - The need for speed in stochastic simulation. PMID- 15286651 TI - Australia experiments with 'experimental use' exemption. AB - Lawmakers are asking whether Australian researchers need an express 'experimental use' defense against patent infringement. PMID- 15286655 TI - Where did the BLOSUM62 alignment score matrix come from? AB - Many sequence alignment programs use the BLOSUM62 score matrix to score pairs of aligned residues. Where did BLOSUM62 come from? PMID- 15286656 TI - Should software hold data hostage? PMID- 15286657 TI - Ligand selectivity and competition between enzymes in silico. AB - In a cell, there are many possibilities for cross interactions between enzymes and small molecules, arising from the similarities in the structures of the metabolites and the flexibility in binding of protein active sites. Despite this promiscuity, the cognate partners must be able to recognize each other in vivo, for the cell to function efficiently. This study examines the basis of this selectivity in recognition using standard docking calculations and finds significant improvement when proteins and ligands are cross-docked. We find that cognate molecules rarely form the most stable complexes and that specificity may be driven either by recognition of the substrate by the enzyme or the recognition of the enzyme by the substrate. Despite limitations of the in silico methods, especially the scoring functions, these calculations highlight the need to consider cross reactions in the cell and suggest that localization and compartmentalization must be important factors in the evolution of complex cells. However, the inherent promiscuity of these interactions can also benefit an organism, by facilitating the evolution of new functions from old ones. The results also suggest that high-throughput screening should involve not just a panel of small molecules, but also a panel of proteins to test for cross reactivity. PMID- 15286659 TI - Structural basis for inhibition of the replication licensing factor Cdt1 by geminin. AB - To maintain chromosome stability in eukaryotic cells, replication origins must be licensed by loading mini-chromosome maintenance (MCM2-7) complexes once and only once per cell cycle. This licensing control is achieved through the activities of geminin and cyclin-dependent kinases. Geminin binds tightly to Cdt1, an essential component of the replication licensing system, and prevents the inappropriate reinitiation of replication on an already fired origin. The inhibitory effect of geminin is thought to prevent the interaction between Cdt1 and the MCM helicase. Here we describe the crystal structure of the mouse geminin-Cdt1 complex using tGeminin (residues 79-157, truncated geminin) and tCdt1 (residues 172-368, truncated Cdt1). The amino-terminal region of a coiled-coil dimer of tGeminin interacts with both N-terminal and carboxy-terminal parts of tCdt1. The primary interface relies on the steric complementarity between the tGeminin dimer and the hydrophobic face of the two short N-terminal helices of tCdt1 and, in particular, Pro 181, Ala 182, Tyr 183, Phe 186 and Leu 189. The crystal structure, in conjunction with our biochemical data, indicates that the N-terminal region of tGeminin might be required to anchor tCdt1, and the C-terminal region of tGeminin prevents access of the MCM complex to tCdt1 through steric hindrance. PMID- 15286660 TI - CD10 is expressed in a subset of chromophobe renal cell carcinomas. AB - CD10 has been considered a useful marker in the diagnosis of renal carcinomas, because of its expression in clear cell and papillary renal cell carcinomas and its absence in chromophobe renal cell carcinomas. On the other hand, chromophobe renal cell carcinoma expresses parvalbumin, which is absent in clear cell and papillary renal cell carcinomas. To further address the relevance of these markers, we studied the expression of CD10 and parvalbumin in 42 samples of chromophobe renal cell carcinoma (seven of which had aggressive features, including invasion beyond the renal capsule, renal vein invasion, metastases, or sarcomatoid transformation), 75 clear cell renal cell carcinomas (eight metastatic) and 51 papillary renal cell carcinomas (two metastatic). CD10 was found in 100% of clear cell renal cell carcinomas, 63% of papillary renal cell carcinomas and in all metastatic cases of both types. At variance with previous studies, we found CD10 expression in from 30 to 90% of the neoplastic cells, in 11 of 42 (26%) chromophobe renal cell carcinomas. The CD10-positive cases included five of the seven (71%) chromophobe renal cell carcinoma with aggressive features. Statistical analysis showed significant association of CD10-positive tumors with clinicopathologic aggressiveness (P=0.003) and mitotic figures (P=0.04). Parvalbumin was strongly expressed in all primary and metastatic chromophobe renal cell carcinomas. Western blot analysis was utilized to confirm the expression of both CD10 and parvalbumin in chromophobe renal cell carcinomas. PMID- 15286661 TI - Localization of heparanase in esophageal cancer cells: respective roles in prognosis and differentiation. AB - In this study, we examined the distribution of heparanase protein in 75 esophageal squamous cell carcinomas by immunohistochemistry and analyzed the relationship between heparanase expression and clinicopathological characteristics. In situ hybridization showed that the mRNA expression pattern of heparanase was similar to that of the protein, suggesting that increased expression of the heparanase protein at the invasive front was caused by an increase of heparanase mRNA in tumor cells. Heparanase expression correlated significantly with depth of tumor invasion, lymph node metastasis, tumor node metastasis (TNM) stage and lymphatic invasion. Overexpression of heparanase in esophageal cancers was also associated with poor survival. In addition to its localization in the cytoplasm and cell membrane, heparanase was also identified in the nuclei of normal epithelial and tumor cells by immunohistochemistry. Furthermore, nuclear heparanase was detected in nuclear extract of cancer cell lines by Western blot and immunohistochemistry. Examination of the role of nuclear heparanase in cell proliferation and differentiation by double immunostaining for proliferating cell nuclear antigen (PCNA) and cytokeratin 10 (CK10) showed significant relationship between nuclear heparanase expression and differentiation (heparanase vs CK10), but not for proliferative state of esophageal cancer cells (heparanase vs PCNA). Our results suggest that cytoplasmic heparanase appears to be a useful prognostic marker in patients with esophageal cancer and that nuclear heparanase protein may play a role in differentiation. Inhibition of heparanase activity may be effective in the control of esophageal tumor invasion and metastasis. PMID- 15286662 TI - AL-amyloidosis and light-chain deposition disease light chains induce divergent phenotypic transformations of human mesangial cells. AB - Human mesangial cells (HMCs) are injured by either excessive amounts or abnormal light chains (LCs), or a combination of both in patients with plasma cell dyscrasias. Consequently, these HMCs undergo phenotypic transformations. HMCs were incubated with eight different light-chains (LCs) for 96 h. These cells, in addition to 51 patient samples from patients with AL-amyloidosis (AL-Am), light chain deposition disease (LCDD), myeloma cast nephropathy (MCN) and controls were analyzed by immunohistochemistry for CD68, muscle-specific actin (MSA), smooth muscle actin (SMA), CD14, and Ham56 protein expressions. All samples were also studied using electron microscopy. Greater staining (four- and three-fold) expressions of CD68 and Ham56, respectively, were observed in the HMCs incubated with AL-Am-LCs compared to those with LCDD-LCs and control. SMA expression levels were five-fold higher in LCDD-LC-treated cells compared to the other categories of LC-treated and control cells. Similar results were obtained in the renal specimens, however, CD68 levels were 12-fold higher in the AL-Am cases compared to the LCDD cases, respectively. Conversely, MSA and SMA levels were three fold higher in the LCDD cases than in the AL-Am ones. No CD14 expression was noted in any of the samples and CD-34 staining of HMCs treated with the various LCs only showed rare positive cells. Dynamic real-time studies to visualize the rough endoplasmic reticulum (RER) and lysosomal compartments in HMCs incubated with LCDD and AL-Am-LCs showed striking expansion of each of the above-mentioned compartments, respectively. This indicates the presence of more RER in the LCDD LC-treated HMCs and a striking increase in lysosomes noticeable in the AL-Am-LC treated cells. Data obtained in this study highlighted that HMCs incubated with LCDD-LCs undergo a myofibroblastic phenotypic transformation, while AL-Am-LCs induce a macrophage-like phenotype in these cells. PMID- 15286663 TI - Cone- and rod-mediated multifocal electroretinogram in early age-related maculopathy. AB - PURPOSE: To investigate the cone- and rod-mediated multifocal electroretinograms (mfERG) in early age-related maculopathy (early ARM). METHODS AND SUBJECTS: We investigated the cone- and rod-mediated mfERG in 17 eyes of 17 subjects with early ARM and 16 eyes of 16 age-matched control subjects with normal fundi. All subjects had a visual acuity of 6/12 or better. We divided the ARM subjects into two groups based on drusen size and retinal pigment epithelium abnormalities-a less advanced (ARM1) and a more advanced (ARM2) group. The mfERG data were compared to templates derived from the control group. We analysed the mfERG results for the central and peripheral fields (CP method) and the superior and inferior fields (SI method). RESULTS: While the mean cone results showed no statistically significant difference between the groups, the rods showed significantly delayed responses in the ARM1 group for the CP and the SI methods, but not in the ARM2 group, although there was a trend of longer latencies compared to the control group. CONCLUSION: Our results show a functional impairment of the rods in early ARM subjects. As there is histopathological evidence showing earlier rod than cone impairment in early ARM, following the rod function with the mfERG might be helpful in diagnosis or for monitoring the progression of early ARM. PMID- 15286664 TI - Calcification of Aqua-Sense intraocular lenses. PMID- 15286665 TI - Recurrent endophthalmitis caused by Burkholderia cepacia. PMID- 15286666 TI - Bilateral, multiple choroidal effusions after vomiting. PMID- 15286667 TI - Inflammatory markers and retinopathy in pregnancies complicated with type I diabetes. AB - AIM: The relation of maternal cytokine levels to retinopathy progression during diabetic pregnancy is a less studied subject. Therefore, we investigated levels of systemic proinflammatory markers, C-reactive peptide (CRP), interleukin-6 (IL 6) and circulating vascular cell adhesion molecule-1 (VCAM-1) during pregnancy and postpartum in relation to the progression of diabetic retinopathy (DR). METHODS: A prospective follow-up study of 39 pregnant women with Type I diabetes and eight nondiabetic pregnant women was performed. DR was graded from fundus photographs. Plasma levels of systemic proinflammatory markers were measured by immunofluorometric assay (CRP) and by enzyme-linked immunosorbent assay (IL-6 and VCAM-1) in the first, second (diabetics only), third trimester of pregnancy, and 3 and 6 months postpartum (diabetics only). RESULTS: Our diabetic women had good glycaemic control (HbA1c 6.9 +/- 0.8). The levels of IL-6, VCAM-1, and CRP did not differ between diabetic and nondiabetic women throughout pregnancy and postpartum (repeated measures ANOVA between the groups). An association between CRP and progression of retinopathy was observed in diabetic women (P = 0.037). Additional evidence of inter-relationship could be revealed as CRP was higher in those diabetic women with worse glycaemic control (HbA1c) (P = 0.038). CONCLUSIONS: During pregnancy and postpartum, levels of proinflammatory factors (IL-6, CRP, VCAM-1) seem to be generally similar in Type I diabetic women compared to nondiabetic controls. However, CRP levels were higher in those diabetic women with progression of retinopathy and in those with worse glycaemic control. PMID- 15286668 TI - An unusual appearance of limbal conjunctival follicles in a patient on brimonidine and dorzolamide. PMID- 15286669 TI - Blinking and superficial punctate keratopathy in patients with diabetes mellitus. AB - AIM: To evaluate blinking patterns in patients with diabetes mellitus and whether blinking contributes to the formation of superficial punctate keratopathy in diabetic patients. METHODS: We examined 163 patients with type II diabetes mellitus and 76 without diabetes. Blinks were recorded, analysed using six parameters, and compared between patients with and without diabetes. Multivariate regression analysis was performed to assess the influence of other ocular factors, such as status of tear lipid layer, tear breakup time, corneal sensitivity, the result of cotton thread test, or blinking rate related to superficial punctate keratopathy. RESULTS: In patients with diabetes, the average mean and maximum interblinking times were longer, the average coefficient of variation of interblinking time was higher, and the average blinking rates were lower than those in patients without diabetes. Multivariate regression analysis revealed that the status of tear lipid layer and tear breakup time were significantly relevant to superficial punctate keratopathy (P < 0.01). CONCLUSION: Interblinking time was longer in diabetic patients, resulting in a decreased blinking rate. The prevalence of superficial punctate keratopathy cannot be predicted from blinking patterns in patients with diabetes. PMID- 15286670 TI - Hemiretinal vein occlusion associated with pseudotumour orbit: an observational case report. PMID- 15286671 TI - Risk management strategies following analysis of cataract negligence claims. AB - INTRODUCTION: Clinical governance and risk management is very important in today's clinical practice. Cataract surgery is one of the most common procedures performed in the NHS, with around 200,000 operations per year. In order to help minimise the frequency of negligence claims, we performed a collaborative study to analyse claims relating to cataract surgery, dealt with by the defence organisations of England, Scotland, Wales, and Northern Ireland. MATERIALS AND METHODS: All claims dealt with by the Medical Defence Union, the Medical Protection Society, and the Medical and Dental Defence Union of Scotland from January 1990 to December 1999, were analysed by three ophthalmologists with at least 5 years' speciality experience. Recurrent themes were identified and claims were grouped by major causative factor. The findings were discussed by a panel compromising the authors in conjunction with the defence unions and risk management strategies were designed. RESULTS: There were 96 claims within the 10- year period analysed. Of these, the largest group (52) pertained to claims that related to accepted complications of cataract surgery. The remainder comprised two groups: 'Medical Errors' (anaesthetic, surgeon, and biometry) and 'Other Claims' comprising subjective complaints, pain and poor visual outcome. A total of 16 claims had been settled by May 2002, 45 are ongoing and 35 have closed without settlement. CONCLUSIONS: The majority of claims pertained to well recognised complications of cataract surgery. If these risks are adequately explained to the patient before surgery and if the care provided reaches a standard acceptable to a responsible body of professional opinion, all such claims should be defensible. Good visual outcome does not protect against litigation. PMID- 15286672 TI - Amniotic membrane transplantation in acute chemical burns. AB - PURPOSE: To evaluate the outcome of fresh amniotic membrane transplantation (AMT) for ocular surface reconstruction in acute chemical burns. METHODS: A prospective study of 15 consecutive eyes with acute chemical burns was performed. In all, 10 eyes had lime burns and five eyes had acid burns. There were three eyes of grade II, four eyes of grade III and eight eyes of grade IV burns. AMT was performed within 3 weeks of injury. RESULTS: Patients were followed up for 10.14 +/- 4.41 months. All patients had immediate relief of pain postoperatively. Of 15 eyes, nine (60%) showed epithelialization within 1-4 weeks (15.33 +/- 9.91 days). The final visual acuity improved in 10 of 15 eyes (66.66%). Eyes with burns of grade II and III showed more visual improvement than those with grade IV burns. None of the eyes showed perforation. Symblepharon was seen in nine of 15 eyes (60%). Of 15 eyes, 12 (80%) experienced limbal stem cell deficiency and showed superficial corneal vascularization. CONCLUSIONS: Amniotic membrane transplantation with fresh amniotic membrane increases patient comfort and reduces inflammation. In mild burns, AMT alone restores corneal and conjunctival surfaces. In moderate to severe burns, it probably reduces conjunctival scarring sequelae, but does not prevent the sequelae of limbal stem cell deficiency that requires further limbal stem cell transplantation. In the acute stage, amniotic membrane transplantation probably has a protective role against the progressive melting and perforation. PMID- 15286673 TI - Correlation of lipid layer thickness measurements with fluorescein tear film break-up and Schirmer's test. PMID- 15286674 TI - En bloc excision of intraocular epithelial cystic downgrowth using syngeneic auricular cartilage. AB - AIM: To describe the surgical management of epithelial cystic downgrowth by en bloc excision and use of syngeneic auricular cartilage in two children. STUDY DESIGN: Retrospective interventional case series. METHOD: The charts of two patients treated for epithelial cystic downgrowth with en bloc excision and auricular cartilage transplant were reviewed. Details of ocular history, preoperative and postoperative visual acuity, intraocular pressure, ocular examination findings, surgical procedure and subsequent management were noted. RESULTS: Two children aged 4 and 6 years, with epithelial cysts who underwent en bloc excision were identified. The cysts had developed following penetrating eye injury. Surgery involved en bloc resection of the cyst and associated tissue, and replacement of the excised corneoscleral tissue with syngeneic auricular cartilage. One patient additionally required synechiolysis, discission of a secondary cataract and anterior vitrectomy. In both cases, the epithelial tissue was successfully removed and the auricular cartilage transplant was well-apposed. Visual acuity remained at the preoperative level in the first patient due to amblyopia; in the second patient visual acuity improved to 6/7.5 with mild astigmatic correction. CONCLUSION: En bloc excision provides the most definitive surgical treatment of cystic epithelial downgrowth. Auricular cartilage may be used for sclerokeratoplasty when donor cornea or sclera is unavailable. PMID- 15286675 TI - Choroidal neovascularization secondary to choroidal osteoma: successful treatment with photodynamic therapy. PMID- 15286676 TI - Photodynamic therapy of choroidal haemangioma associated with Sturge-Weber syndrome. PMID- 15286677 TI - Extranasal nasal glioma. PMID- 15286678 TI - Inter-device reproducibility of the scanning laser polarimeter with variable cornea compensation. AB - OBJECTIVE: To evaluate the interdevice reproducibility of retinal nerve fibre layer (RNFL) thickness measurements obtained with the commercially available GDx VCC, a scanning laser polarimeter with variable (individualized) corneal compensation. METHODS: A prospective instrument validation study in which 13 GDx VCC devices were tested. One eye each, from three normal subjects were used to test each of the devices, on the same day, by an experienced operator. Variability and reproducibility for each of five GDx parameters were calculated. RESULTS: For each of five tested GDx parameters, the coefficient of variation and 95% confidence interval range (microm), for the 13 devices, respectively, were: TSNIT avg: 5.1%, 3.84 microm; Superior avg: 5.3%, 4.82 microm; Inferior avg: 6.1%, 5.50 microm; TSNIT standard deviation: 8.6%, 2.92 microm; and nerve fibre indicator (NFI): N/A, 5.69. Item reliability (Cronbach's alpha) for the five GDx parameters are: TSNIT-Avg: 0.97, Sup-Avg: 1.00, Inf-Avg: 0.84, TSNIT-SD: 0.99, NFI: 0.99. CONCLUSIONS: With the commercially available GDx-VCC, our results indicate that RNFL measurements appear reproducible across devices. PMID- 15286680 TI - Phacoemulsification and intraocular lens implantation following pars plana vitrectomy: a prospective study. PMID- 15286681 TI - Oncolytic viral therapies. AB - Molecular research has vastly advanced our understanding of the mechanism of cancer growth and spread. Targeted approaches utilizing molecular science have yielded provocative results in the treatment of cancer. Oncolytic viruses genetically programmed to replicate within cancer cells and directly induce toxic effect via cell lysis or apoptosis are currently being explored in the clinic. Safety has been confirmed and despite variable efficacy results several dramatic responses have been observed with some oncolytic viruses. This review summarizes results of clinical trials with oncolytic viruses in cancer. PMID- 15286682 TI - Mycoplasma fermentans inhibits tumor necrosis factor alpha-induced apoptosis in the human myelomonocytic U937 cell line. AB - Mycoplasma fermentans (M. fermentans) was shown to be involved in the alteration of several eukaryotic cell functions (i.e. cytokine production, gene expression), and was suggested as a causative agent in arthritic diseases involving impaired apoptosis. We investigated whether M. fermentans has a pathogenic potential by affecting tumor necrosis factor (TNF)alpha-induced apoptosis in the human myelomonocytic U937 cell line. A significant reduction in the TNFalpha-induced apoptosis (approximately 60%) was demonstrated upon either infection with live M. fermentans or by stimulation with non-live M. fermentans. To investigate the mechanism of M. fermentans antiapoptotic effect, the reduction of mitochondrial transmembrane potential (DeltaPsim) and the protease activity of caspase-8 were measured. In the infected cells, the reduction of DeltaPsim was inhibited (approximately 75%), and an approximately 60% reduction of caspase-8 activity was measured. In conclusion, M. fermentans significantly inhibits TNFalpha-induced apoptosis in U937 cells, and its effect is upstream of the mitochondria and upstream of caspase-8. PMID- 15286684 TI - Heat stress induces different forms of cell death in sea anemones and their endosymbiotic algae depending on temperature and duration. AB - Bleaching of reef building corals and other symbiotic cnidarians due to the loss of their dinoflagellate algal symbionts (=zooxanthellae), and/or their photosynthetic pigments, is a common sign of environmental stress. Mass bleaching events are becoming an increasingly important cause of mortality and reef degradation on a global scale, linked by many to global climate change. However, the cellular mechanisms of stress-induced bleaching remain largely unresolved. In this study, the frequency of apoptosis-like and necrosis-like cell death was determined in the symbiotic sea anemone Aiptasia sp. using criteria that had previously been validated for this symbiosis as indicators of programmed cell death (PCD) and necrosis. Results indicate that PCD and necrosis occur simultaneously in both host tissues and zooxanthellae subject to environmentally relevant doses of heat stress. Frequency of PCD in the anemone endoderm increased within minutes of treatment. Peak rates of apoptosis-like cell death in the host were coincident with the timing of loss of zooxanthellae during bleaching. The proportion of apoptosis-like host cells subsequently declined while cell necrosis increased. In the zooxanthellae, both apoptosis-like and necrosis-like activity increased throughout the duration of the experiment (6 days), dependent on temperature dose. A stress-mediated PCD pathway is an important part of the thermal stress response in the sea anemone symbiosis and this study suggests that PCD may play different roles in different components of the symbiosis during bleaching. PMID- 15286683 TI - Expression of apoptosis inhibitor protein Mcl1 linked to neuroprotection in CNS neurons. AB - Mcl1 is a Bcl2-related antiapoptotic protein originally isolated from human myeloid leukemia cells. Unlike Bcl2, expression has not been reported in CNS neurons. We isolated Mcl1 in a direct screen for candidate modifier genes of neuronal vulnerability by differential display of mRNAs upregulated following prolonged seizures in two mouse strains with contrasting levels of hippocampal cell death. Mcl1 is widely expressed in neurons, and transcription is rapidly induced in both strains. In resistant C57Bl/6J mice, Mcl1 protein levels remain persistently elevated in hippocampal pyramidal neurons after seizures, but fall rapidly in C3H/HeJ hippocampus, coinciding with extensive neuronal apoptosis. DNA damage and caspase-mediated cell death were strikingly increased in Mcl1 deficient mice when compared to +/+ littermates after similar seizures. We identify Mcl1 as a neuronal gene responsive to excitotoxic insult in the brain, and link relative levels of Mcl1 expression to inherited differences in neuronal thresholds for apoptosis. PMID- 15286685 TI - Thiotepa and fractionated TBI conditioning prior to allogeneic stem cell transplantation for advanced hematologic malignancies: a phase II single institution trial. AB - Relapse of hematologic malignancies after allogeneic stem cell transplantation remains a common problem, in particular for patients who have advanced disease at the time of transplantation. Thiotepa has excellent antileukemic and immunosuppressive activity, and could therefore be a useful drug in the conditioning regimen for patients with advanced hematologic neoplasms. We retrospectively analyzed toxicity, engraftment and survival data of 41 patients who received a conditioning regimen of thiotepa (600 mg/m2) and hyperfractionated TBI (10 Gy) prior to matched related (n = 25) or matched unrelated (n = 16) allogeneic stem cell transplantation. The mean age at transplantation was 37.8 years (range 20-59), all but five patients had advanced hematologic malignancies at the time of transplantation. GVHD prophylaxis was with standard cyclosporine and methotrexate. Engraftment was excellent, but the regimen was associated with a high incidence of grade III renal (41%) and hepatic (15%) toxicity, and high transplant-related mortality (44% at day +90). The 3-year event-free survival was 13% and overall survival 14%. We conclude that this regimen requires modification to reduce toxicity. PMID- 15286686 TI - Graft-versus-lymphoma effect in refractory cutaneous T-cell lymphoma after reduced-intensity HLA-matched sibling allogeneic stem cell transplantation. AB - Cutaneous T-cell lymphomas (CTCL) are rare diseases that, in their advanced stages or in transformation, have a poor prognosis. Autologous stem cell transplantation (Au-SCT) after high-dose therapy has yielded disappointing results. Allogeneic transplantation (allo-SCT) provides the potential advantage of an immune-mediated graft-versus-lymphoma (GVL) effect. Reduced-intensity allo SCT potentially offers a GVL effect, but with diminished toxicity related to the induction regimen; however, published experience with this approach in CTCL is limited. We report a series of three patients (age 35-49) with advanced, refractory (n=2) or transformed (n=1) CTCL who underwent reduced-intensity allo SCT in the context of active disease. All three survived the peri-transplant period and, despite later having disease relapse, all exhibited evidence of a GVL effect. Relapses of the disease were in the context of immune suppression for graft-versus-host disease (GVHD), and when immune suppression was reduced, responses were regained. A comparison is made of these results to those in a review of the published literature to date. We conclude that while a GVL can be achieved for CTCL with reduced-intensity allogeneic transplantation, the clinical benefits are short lived and novel approaches are required to obtain sustained remissions. PMID- 15286688 TI - Successful autologous stem cell transplantation in Ph-positive ALL patient with refractory invasive pulmonary aspergillosis. PMID- 15286687 TI - Mycophenolate mofetil and cyclosporine for graft-versus-host disease prophylaxis following reduced intensity conditioning allogeneic stem cell transplantation. AB - The use of reduced intensity conditioning (RIC) regimens for allogeneic stem cell transplantation (allo-SCT) can result in a significant decrease in early procedure-related toxicity in patients not eligible for standard myeloablative regimens. However, acute graft-versus-host disease (aGVHD) remains a matter of concern after RIC allo-SCT, and its incidence might be expected to be higher in elderly and high-risk patients. This report investigated mycophenolate mofetil (MMF) and cyclosporin A (CsA) combination (n=14) in comparison to CsA alone (n=20) for GVHD prophylaxis in cancer patients aged over 50 years (27 haematological malignancies and seven solid tumours) receiving an HLA-identical sibling antithymocyte-globulin (ATG)-based RIC allo-SCT. Baseline demographic characteristics and risk factors for aGVHD were comparable between both groups. Although MMF administration was not associated with any significant toxicity, the cumulative incidence of any form of GVHD was comparable between both groups (cumulative incidence of grade II-IV aGVHD, 50% (95% CI, 28-72%) for CsA alone, as compared to 64% (95% CI, 39-89%) to CsA and MMF, P=NS), suggesting that adjunction of MMF to CsA is feasible, but does not translate towards a significant reduction of aGVHD, at least in the context ATG-based RIC allo-SCT. PMID- 15286689 TI - Infectious complications and outcomes after allogeneic hematopoietic stem cell transplantation in Korea. AB - We reviewed 242 allogeneic hematopoietic stem cell transplantation (HSCT) recipients retrospectively over a 2-year period (January 1998-December 1999) in order to analyze the characteristics and assess the outcomes of infectious complications in patients after HSCT in Korea. Bacteria were the major pathogens before engraftment, and viral and fungal infections predominated during the post engraftment period. Varicella zoster virus was the most common viral pathogen after engraftment. Cytomegalovirus disease occurred mainly in the late-recovery phase. The frequency of mold infection was higher than that of yeast. There was a relatively high incidence of tuberculosis (3.0%) and Pneumocystis carinii pneumonia (6.5%). One case of death by measles confirmed by autopsy was also noted. Overall, cumulative mortality was 43% (104/242), and 59.6% of these deaths (62/104) were infection-related. Allogeneic HSCT recipients from unrelated donors were prone to infectious complication and higher mortality than those from matched sibling (17/39 (43.6%) vs 45/203 (22.2%), respectively; P<0.01; odd ratio 2.5; 95% confidence interval 1.2-5.1). As infection was the main post-HSCT complication in our data, more attention should be given to the management of infections in HSCT recipients. PMID- 15286690 TI - The thymus is typically small at 1 year after autologous or allogeneic T-cell replete hematopoietic cell transplantation into adults. PMID- 15286691 TI - Single agent dexamethasone for pre-stem cell transplant induction therapy for multiple myeloma. AB - Given the survival advantage, high-dose therapy (HDT) remains the standard of care for patients with multiple myeloma eligible for the procedure. For those undergoing HDT, initial therapy aimed at reducing tumor burden is given prior to stem cell harvest. Various regimens, mostly variations of VAD (vincristine, doxorubicin, dexamethasone), are used for induction therapy. We retrospectively evaluated if single agent dexamethasone would be an effective induction therapy, given that it is the most active drug in these combinations. A total of 35 patients who received induction therapy with dexamethasone alone were compared to a similar group of 72 patients who received VAD as the initial therapy. We found a 63% response rate with dexamethasone compared to 74% with VAD (P=0.25). Including minimal responses, the overall response rate for Dex and VAD was 74 and 86%, respectively (P=0.13). The overall and complete response rates to transplant, respectively, were 97 and 26% for the dexamethasone group and 100 and 39% for the VAD group; P=0.33 and 0.18. No significant differences were observed in the progression-free and overall survival at 1 year post transplant. Single agent dexamethasone appears to be an effective alternative to VAD for induction therapy prior to HDT in myeloma. PMID- 15286692 TI - Adoption of long-term cultures to evaluate the cryoprotective potential of trehalose for freezing hematopoietic stem cells. AB - Dimethylsulfoxide (DMSO), which is widely used as a cryoprotectant for hematopoietic stem cells (HSC), has considerable toxicity for both the thawed cells and the patient. The aim of this study was to evaluate the cryoprotective potential of trehalose in comparison to DMSO for human HSC. Human bone marrow (BM) and peripheral blood stem cells (PBSC) of volunteer donors were cryopreserved in the presence of different concentrations of trehalose with and without insulin, as well as with DMSO 10%. After thawing to 37 degrees C colony forming unit (CFU) assays were performed. Long-term marrow-cultures (LTMC) were established and used for the detection of long-term culture-initiating cells (LTCIC). The total amount of CFUs detected was 104+/-134 (mean+/-s.d.) in DMSO preserved cells and 179+/-34 in trehalose-protected cells. For LTMC the best feeder layer proved to be fresh human BM and the most useful concentration of trehalose was 0.5 M. Using these culture conditions we could detect after 5 weeks LTMC a total of 172+/-28 CFUs for trehalose-protected cells and 170+/-52 for DMSO preserved cells. In conclusion, trehalose exerts a similar cryoprotective potential for hematopoietic progenitor and stem cells like DMSO and could possibly replace DMSO at least in part as cryoprotectant in the setting of hematopoietic cell transplantation. PMID- 15286693 TI - Frequent severe liver iron overload after stem cell transplantation and its possible association with invasive aspergillosis. AB - Iron overload is associated with free radical generation and tissue damage. Our main objective was to ascertain the frequency and severity of iron overload in a group of 59 patients who died after conventional-intensity autologous (n=24) or allogeneic (n=35) haematopoietic stem cell transplantation (HSCT). A second objective was to investigate associations between liver-iron concentration and causes of transplant-related mortality. The median age was 41 years (range, 19 66), 41 were males and 18 females. In total, 26 patients had acute leukaemia or MDS, 10 CML, 17 lymphoma, four myeloma and two aplastic anaemia. The median hepatic iron concentration (HIC) was 138 micromol/g dry weight (7.7 mg/g; range 31-631 micromol/g). In total, 4/32 (12%) patients with HIC <150 micromol/g and 10/27 (37%) with hepatic iron > or =150 micromol/g showed invasive aspergillosis at autopsy (P=0.035). This was significant in multivariate analysis (RR 9.0; 95% CI 1.6-50.3, P=0.012). In conclusion, severe iron overload is frequent in patients who die following HSCT and is associated with invasive aspergillosis. PMID- 15286694 TI - High-dose treosulfan in patients with relapsed or refractory high-grade lymphoma receiving tandem autologous blood stem cell transplantation. AB - This phase I/II study evaluated high-dose treosulfan in patients with high-grade lymphoma. In all, 21 patients (median age 51, 25-60 years) with primary refractory disease (n=3) or early (n=11) or late (n=7) relapse received DexaBEAM and one course etoposide for cytoreduction and PBPC mobilization. Subsequently, 16 patients received 30 g/m2 treosulfan and 140 mg/m2 melphalan, followed by autologous transplantation. Nine patients received a 2nd high-dose treatment (HDT) with 30 g/m2 treosulfan and 750 mg/m2 thiotepa. Recovery time to >1/nl leukocytes and >25/nl thrombocytes was 8.9 (range 8-11) and 11.9 (8-16) days after 1st and 9.6 (7-13) and 13 (9-19) days after 2nd HDT. Reversible grade 3 or 4 nonhematologic toxicities included mucositis (n=7), infection (n=7) and one episode of re-entry tachycardia. Two treatment-related deaths occurred after 2nd HDT. Since three dose-limiting toxicities occurred among nine patients receiving tandem HDT, 30 g/m2 of treosulfan was considered MTD in this setting. Patients with late compared to early relapse or refractory disease had a higher probability of CR (6/7 vs 3/14 patients, P=0.017) and overall survival (71 vs 21%, P<0.05, 24-49 months follow-up). In conclusion, high-dose treosulfan as major therapy component induces sustained complete remissions in relapsed high grade lymphoma with acceptable toxicity. PMID- 15286695 TI - Bone metabolism in patients more than five years after bone marrow transplantation. AB - We investigated the bone metabolism of 22 patients (median age 38 years) over 6 years after allogeneic bone marrow transplantation (BMT). Biplanar roentgenograms of the thoracic and lumbar spine were used to diagnose vertebral deformities caused by fractures. The actual bone mineral density (BMD) of the lumbar spine and the femoral neck were measured. Laboratory tests included calcium, phosphate, parathyroid hormone, a marker of bone resorption (beta-crosslaps, CTX), markers of bone formation (osteocalcin, bone-specific alkaline phosphatase), osteoprotegerin (OPG)--antagonist of the osteoclast differentiation factor RANKL, and sex hormone status. One patient had a vertebral fracture. Seven patients (28%) had osteopenia in the lumbar spine while 12 patients (48%) had osteopenia in the femoral neck. Bone resorption was increased in nine patients (43%) and bone formation was increased in four patients (20%). BMT recipients had significantly increased serum levels of OPG (P=0.029). Three women (75%) and four men (25%) were hypogonadal. The data showed that BMD is reduced and bone metabolism is still disturbed more than 6 years after BMT. The RANKL/osteoprotegerin system appears to play an important role in the pathophysiology of late post transplantation osteoporosis. PMID- 15286696 TI - Adoptive immunotherapy with donor lymphocyte infusions and interleukin-2 after high-dose therapy and autologous stem cell rescue for multiple myeloma. AB - In an attempt to induce a graft-versus-myeloma effect, we administered donor lymphocyte infusions (DLI) after high-dose therapy with autologous stem cell transplant rescue to seven patients with refractory or relapsed multiple myeloma. High-dose therapy consisted of melphalan, idarubicin and etoposide (days -9 to 6) followed by autologous stem cell infusion on day 0. DLI (five of seven donors with two or three HLA antigens mismatched) were administered on days +1, +5 and +10 along with IL-2 (from day +1 through +12). Six of the seven patients developed acute graft-versus-host disease (GVHD), which resolved spontaneously, coincidentally with autologous hematopoietic reconstitution. One patient failed to engraft and received a second autologous graft. One patient died from complications of a pulmonary hemorrhage after experiencing GVHD. With a minimum follow-up of 38 months, five patients remain without disease progression in complete remission or with minimal residual disease. In this setting, DLI/IL-2 is biologically active resulting in GVHD. A graft-versus-myeloma effect is suggested by the improved outcome of our small cohort of high-risk patients. The use of partially mismatched related donors makes this approach potentially available to nearly all patients. PMID- 15286697 TI - Fatal pulmonary fibrosis associated with BCNU: the relative role of platelet derived growth factor-B, insulin-like growth factor I, transforming growth factor beta1 and cyclooxygenase-2. AB - Pulmonary fibrosis is a severe complication associated with bis-chloronitrosourea (BCNU) therapy. However, the pathogenetic mechanism has never been well investigated. We report here a 26-year-old female with diffuse large B-cell lymphoma who died of severe pulmonary fibrosis 81 days after the administration of high-dose BCNU (600 mg/m2). Thoracoscopic wedge resection of left upper lung performed 10 days before patient's death showed severe pulmonary fibrosis with prominent hyperplasia of alveolar macrophages and type II pneumocytes. We further used immunohistochemistry (IHC) to examine the relative role of platelet-derived growth factor-B (PDGF-B), insulin-like growth factor I (IGF-I), transforming growth factor-beta1 (TGF-beta1) and cyclooxygenase-2 (COX-2) in the pathogenesis of BCNU-related pulmonary fibrosis. Strong expressions of PDGF-B and IGF-1 on alveolar macrophages and type II pneumocytes were clearly demonstrated, but in contrast, the expressions of TGF-beta1 and COX-2 were almost undetectable. In conclusion, pulmonary fibrosis can develop early and progress rapidly after the administration of high-dose BCNU. The markedly increased expression of fibrogenic factors PDGF-B and IGF-1 on hyperplastic alveolar macrophages and hyperplastic type II pneumocytes may play an important role in the fibrogenesis of this disease. These novel findings may offer specific therapeutic targets in the treatment of BCNU-associated pulmonary fibrosis. PMID- 15286698 TI - G-CSF or not G-CSF? That is the question. PMID- 15286699 TI - Rust and corrosion in hematopoietic stem cell transplantation: the problem of iron and oxidative stress. AB - Iron overload is a common acute and long-term event associated with autologous and allogeneic hematopoietic stem cell transplantation (HSCT). In a state of iron excess, free iron becomes available to catalyze the conversion of reactive oxygen species (ROS) intermediates such as superoxide anion (O2*-) and hydrogen peroxide (H2O2) to highly toxic free radicals such as hydroxyl radical (OH*). ROS may help to promote chronic liver disease, sinusoidal obstruction syndrome, idiopathic pneumonia syndrome and bacterial, fungal and other opportunistic infections. Phlebotomy has been effectively and safely used to deplete excess iron stores post-HSCT in thalassemic and other iron-overloaded patients. Intracellular iron levels may also be decreased through pharmacologic chelating agents, while antioxidants such as N-acetylcysteine, glutamine (glutathione precursor) and captopril have been shown to replenish glutathione redox potential and scavenge free radicals. A better understanding of the mechanisms involved in the iron generated pro-oxidant state associated with HSCT will likely lead to reduced toxicity and improved patient outcomes. PMID- 15286700 TI - Prolactin induces c-Myc expression and cell survival through activation of Src/Akt pathway in lymphoid cells. AB - Stimulation of resting W53 cells (lymphoid murine cells expressing prolactin (PRL) receptor) by PRL induced expression of growth-related immediate-early genes (IEG), and proliferation through activation of the Src kinases. Since IEG are essential for cell cycle progression, we have studied how PRL controls expression of c-Myc mRNA and c-Fos. Stimulation of W53 cell proliferation by PRL required activation of MAPK, as the Mek1/2 inhibitor PD184352 eliminated Erk1/2 stimulation, cell proliferation, and expression of c-Fos mRNA. In contrast, PD184352 did not alter PRL activation of c-Myc mRNA expression or stimulation of p70S6K, Akt, and the Jak2/Stat5 pathway. Activation of the PI3K by PRL was necessary for the expression of c-MycmRNA and W53 cell proliferation, as the PI3K inhibitor LY294002 abolished them. However, it did not modify PRL stimulation of c-Fos mRNA expression or activation of Erk1/2 and Stat5. Furthermore, rapamycin, an inhibitor of mTOR and consequently of p70S6K, did not alter PRL stimulation of c-Myc and c-Fos mRNA expression and it had a very minor inhibitory effect on PRL stimulation of W53 cell proliferation. In addition, rapamycin did not affect PRL stimulation of Akt or Stat5. However, it reinforced PRL activation of Erk1/2. Overexpression of a constitutively activated Akt (myristoylated Akt) in W53 cells overcame the inhibitory effect of LY294002 on c-Myc expression, as well as cell death upon PRL deprivation. Consistently, inducible expression of Akt-CAAX Box in W53 cells caused inhibition of c-Myc expression. PRL stimulation of W53 cells resulted in Akt translocation to the nucleus, phosphorylation of FKHRL1 transcription factor, and its nuclear exclusion. In contrast, induced expression of Akt-CAAX Box caused inhibition of FKHRL1 phosphorylation. Furthermore, transient expression of nonphosphorylatable FKHRL1-A3 mutant impaired PRL-induced activation of the c-Myc promoter. Akt activation also resulted in phosphorylation and inhibition of glycogen synthetase kinase 3 (GSK3), which in turn promoted c Myc stability. Consistently, treatment of W53 with selective inhibitors of GSK3 such as SB415286 and lithium salts resulted in increased levels of c-Myc. Also, overexpression of c-Myc in W53 cells overcame the decrease in cell proliferation induced by LY294002. These findings defined a PRL-signalling cascade in W53 cells, involving Src kinases/PI3K/Akt/FKHRL1-GSK3, that mediates stimulation of c Myc expression. PMID- 15286701 TI - Antitumor agent parthenolide reverses resistance of breast cancer cells to tumor necrosis factor-related apoptosis-inducing ligand through sustained activation of c-Jun N-terminal kinase. AB - The antitumor activity of the sesquiterpene lactone parthenolide, an active ingredient of medicinal plants, is believed to be due to the inhibition of DNA binding of transcription factors NF-kappaB and STAT-3, reduction in MAP kinase activity and the generation of reactive oxygen. In this report, we show that parthenolide activates c-Jun N-terminal kinase (JNK), which is independent of inhibition of NF-kappaB DNA binding and generation of reactive oxygen species. Parthenolide reversed resistance of breast cancer cells to tumor necrosis factor related apoptosis-inducing ligand (TRAIL)-induced apoptosis. Cancer cells treated with a combination of TRAIL and parthenolide underwent massive typical apoptosis and atypical apoptosis involving the loss of plasma membrane integrity. JNK activity is necessary for the parthenolide-induced sensitization to TRAIL because a dominant-negative JNK or the JNK inhibitor SP600125 reduced TRAIL plus parthenolide-induced apoptosis. Parthenolide induced phosphorylation of Bid and increased TRAIL-dependent cleavage of Bid without affecting caspase 8 activities. Cytochrome c but not Smac/DIABLO was released from the mitochondria in cells treated with parthenolide alone. Parthenolide through JNK increased the TRAIL mediated degradation of the antiapoptotic protein X-linked inhibitor of apoptosis (XIAP). Enhanced XIAP cleavage correlated with increased and prolonged caspase 3 activity and PARP cleavage, suggesting that the sensitization to TRAIL involves 'feed forward' activation of caspase 3. These results identify a new antitumor activity of parthenolide, which can be exploited to reverse resistance of cancer cells to TRAIL, particularly those with elevated XIAP levels. PMID- 15286702 TI - Regulation of Snail transcription during epithelial to mesenchymal transition of tumor cells. AB - Expression of Snail transcriptional factor is a determinant in the acquisition of a mesenchymal phenotype by epithelial tumor cells. However, the regulation of the transcription of this gene is still unknown. We describe here the characterization of a human SNAIL promoter that contains the initiation of transcription and regulates the expression of this gene in tumor cells. This promoter was activated in cell lines in response to agents that induce Snail transcription and the mesenchymal phenotype, as addition of the phorbol ester PMA or overexpression of integrin-linked kinase (ILK) or oncogenes such as Ha-ras or v-Akt. Although other regions of the promoter were required for a complete stimulation by Akt or ILK, a minimal fragment (-78/+59) was sufficient to maintain the mesenchymal specificity. Activity of this minimal promoter and SNAIL RNA levels were dependent on ERK signaling pathway. NFkappaB/p65 also stimulated SNAIL transcription through a region located immediately upstream the minimal promoter, between -194 and -78. These results indicate that Snail transcription is driven by signaling pathways known to induce epithelial to mesenchymal transition, reinforcing the role of Snail in this process. PMID- 15286703 TI - Differential immunization identifies PHB1/PHB2 as blood-borne tumor antigens. AB - Early diagnosis of cancer is crucial for successful treatment. Noninvasive assays for detecting tumor-derived antigens in serum and other bodily fluids have the potential to screen healthy individuals for hitherto undetected cancers. Very few such assays have been successfully developed, in part because identifying potential target antigens remains a challenge. To identify new blood-borne tumor antigens for the purpose of establishing such assays, we have developed a novel technique called differential immunization. Using this method, we have identified PHB1 and PHB2, proteins thought to function as mitochondrial chaperones and transcriptional regulators, as antigens released from colorectal tumors in vivo. Serum from colorectal patients contains significantly higher levels of these antigens compared to serum from healthy volunteers. These data demonstrate that differential immunization is an effective new method for identifying tumor derived antigens in serum. PMID- 15286704 TI - Effects of novel inhibitors of poly(ADP-ribose) polymerase-1 and the DNA dependent protein kinase on enzyme activities and DNA repair. AB - DNA-dependent protein kinase (DNA-PK) and poly (ADP-ribose) polymerase-1 (PARP-1) participate in nonhomologous end joining and base excision repair, respectively, and are key determinants of radio- and chemo-resistance. Both PARP-1 and DNA-PK have been identified as therapeutic targets for anticancer drug development. Here we investigate the effects of specific inhibitors on enzyme activities and DNA double-strand break (DSB) repair. The enzyme activities were investigated using purified enzymes and in permeabilized cells. Inhibition, or loss of activity, was compared using potent inhibitors of DNA-PK (NU7026) and PARP-1 (AG14361), and cell lines proficient or deficient for DNA-PK or PARP-1. Inactive DNA-PK suppressed the activity of PARP-1 and vice versa. This was not the consequence of simple substrate competition, since DNA ends were provided in excess. The inhibitory effect of DNA-PK on PARP activity was confirmed in permeabilized cells. Both inhibitors prevented ionizing radiation-induced DSB repair, but only AG14361 prevented single-strand break repair. An increase in DSB levels caused by inhibition of PARP-1 was shown to be caused by a decrease in DSB repair, and not by the formation of additional DSBs. These data point to combined inhibition of PARP-1 and DNA-PK as a powerful strategy for tumor radiosensitization. PMID- 15286705 TI - Implication of BRG1 and cdk9 in the STAT3-mediated activation of the p21waf1 gene. AB - STAT transcription factors (signal transducers and activators of transcription) are cytoplasmic proteins that induce gene activation in response to cytokine receptor stimulation. Following tyrosine phosphorylation, STAT proteins translocate into the nucleus and activate specific target genes. We have previously reported that STAT3 activates the expression of the p21waf1 gene through its association with the NcoA/SRC1a and CBP coactivators. In this study, we explore the role of BRG1, a component of the SWI/SNF chromatin-remodeling complex, and the role of cdk9, a component of the elongation factor P-TEFb, in the STAT3-mediated expression of p21waf1. We found using pull-down experiments and co-immunoprecipitation assays that both proteins associate with STAT3. Chromatin immunoprecipitation (ChIP) experiments indicate that STAT3 DNA binding results in histone H3 acetylation and BRG1 recruitment. Using Southern blot analysis, we found that the loading of BRG1 is followed by an increased accessibility of the proximal p21waf1 promoter and by the association of RNA polymerase II. As a next step, STAT3 then recruits the cdk9 kinase to phosphorylate the carboxy-terminal domain of the RNA polymerase at serine 2. Accordingly, the elongating form of the polymerase can be detected by ChIP experiments on the coding region of the gene, probably initiating mRNA synthesis. Therefore, STAT3 not only promotes the initiation of transcription but also regulates chromatin remodeling and transcription elongation through its interaction with BRG1 and cdk9. PMID- 15286706 TI - Regulated proteolysis of the IFNaR2 subunit of the interferon-alpha receptor. AB - The type I interferons (IFNs) bind surface receptors, induce JAK kinases and activate STAT transcription factors to stimulate the transcription of genes downstream of IFN-stimulated response elements (ISREs). In this study, we demonstrate that IFNaR2, a subunit of the type I IFN receptor, is proteolytically cleaved in a regulated manner. Immunoblotting shows that multi-step cleavage occurs in response to phorbol ester (PMA) and IFN-alpha, generating both a transmembrane 'stub' and the intracellular domain (ICD), similar to Notch proteolysis. Isolated membrane fractions process IFNaR2 to release the ICD. A chimeric receptor construct is utilized to show that cleavage requires the presenilins and occurs in response to epidermal growth factor and protein kinase C-delta overexpression, as well as PMA and type I IFNs. Fluorescence microscopy demonstrates that a green fluorescent protein-ICD fusion localizes predominantly to the nucleus. A fusion between the ICD and the Gal4 DNA-binding domain represses transcription, in a histone deacetylase-dependent manner, of a Gal4 upstream activating sequence-regulated reporter, while overexpression of the ICD alone represses transcription of a reporter linked to an ISRE. Proteolytic cleavage events may facilitate receptor turnover or, more likely, function as a mechanism for signaling similar to that employed by Notch and the Alzheimer's precursor protein. PMID- 15286707 TI - Crosstalk of the mitotic spindle assembly checkpoint with p53 to prevent polyploidy. AB - Treatment of cells with microtubule inhibitors results in activation of the mitotic spindle assembly checkpoint, leading to mitotic arrest before anaphase. Upon prolonged treatment, however, cells can adapt and exit mitosis aberrantly, resulting in the occurrence of tetraploid cells in G1. Those cells subsequently arrest in postmitotic G1 due to the activation of a p53-dependent G1 checkpoint. Failure of the G1 checkpoint leads to endoreduplication and further polyploidization. Using HCT116 and isogenic p53-deficient or spindle checkpoint compromised derivatives, we show here that not only p53 but also a functional spindle assembly checkpoint is required for postmitotic G1 checkpoint function. During transient mitotic arrest, p53 stabilization and activation is triggered by a pathway independent of ATM/ATR, Chk1 and Chk2. We further show that a prolonged spindle checkpoint-mediated mitotic arrest is required for proper postmitotic G1 checkpoint function. In addition, we demonstrate that polyploid cells are inhibited to re-enter mitosis by an additional checkpoint acting in G2. Thus, during a normal cell cycle, polyploidization and subsequent aneuploidization is prevented by the function of the mitotic spindle checkpoint, a p53-dependent G1 checkpoint and an additional G2 checkpoint. PMID- 15286708 TI - Host-derived plasminogen activator inhibitor-1 (PAI-1) concentration is critical for in vivo tumoral angiogenesis and growth. AB - Plasminogen activator inhibitor type 1 (PAI-1) plays a key role in tumor progression and is believed to control proteolytic activity and cell migration during angiogenesis. We report here that host PAI-1, at physiological concentration, promotes in vivo tumor invasion and angiogenesis. In sharp contrast, inhibition of tumor vascularization was observed when PAI-1 was produced at supraphysiologic levels, either by host cells (transgenic mice overexpressing PAI-1) or by tumor cells (after transfection with murine PAI-1 cDNA). This study provides for the first time in vivo evidence for a dose dependent effect of PAI-1 on tumor angiogenesis. Of great interest is the finding that PAI-1 produced by tumor cells, even at high concentration, did not overcome the absence of PAI-1 in the host, emphasizing the importance of the cellular source of PAI-1. PMID- 15286709 TI - Stability of nucleolar versus non-nucleolar forms of human p14(ARF). AB - Fusion proteins containing the amino-terminal domain of human p14(ARF) linked to green fluorescent protein are able to bind MDM2 and stabilize p53 without localization in the nucleolus. However, these fusion proteins are inherently unstable, with half-lives considerably shorter than either authentic ARF or chimaeras containing the entire coding domain, both of which are predominantly nucleolar. We present evidence that the unstable fusion proteins are significantly stabilized if redirected to the nucleolus by addition of a basic motif based on the nuclear localization signal of SV40 T-antigen. Moreover, the stability of these proteins can be enhanced by modulating the functions of MDM2 and p53. These data are consistent with a model in which ARF interacts with MDM2 in the nucleoplasm but is consequently subject to proteasomal degradation. Nucleolar localization may serve to store or stabilize ARF. PMID- 15286710 TI - Analysis of the transforming and growth suppressive activities of the PAX3-FKHR oncoprotein. AB - The 2;13 chromosomal translocation occurs in most cases of the cancer alveolar rhabdomyosarcoma (ARMS), and juxtaposes the genes encoding the PAX3 and FKHR transcription factors. The resulting chimeric protein PAX3-FKHR is a potent transcriptional activator, and is hypothesized to function as a dominant acting oncogene. To investigate its biological function, PAX3-FKHR was transduced into three immortalized murine cell lines in either a constitutive or inducible manner. These cells only tolerate expression of low PAX3-FKHR levels, which is sufficient for transformation in NIH3T3 cells. In contrast, higher PAX3-FKHR levels, which are comparable to the endogenous level expressed in ARMS cells, result in growth suppression. To determine as to which PAX3 functional domains are needed for growth suppression and transformation, inactivating mutations were introduced into the paired box and homeodomain of PAX3-FKHR. In these experiments, the homeodomain is necessary for transformation, but not growth suppression; whereas the paired box is not required for transformation but mediates growth suppression. In summary, our findings demonstrate that the transforming and growth suppressive activities of PAX3-FKHR are dominant at different activity levels and are mediated by distinct functional domains. These findings are consistent with the hypothesis that distinct expression pathways are operative in these opposing phenotypic end points. PMID- 15286711 TI - P53 in cytoplasm may enhance the accuracy of DNA synthesis by human immunodeficiency virus type 1 reverse transcriptase. AB - The tumor suppressor protein p53 displays 3' --> 5' exonuclease activity and can provide a proofreading function for DNA polymerases. Reverse transcriptase (RT) of human immunodeficiency virus (HIV)-1 is responsible for the conversion of the viral genomic ssRNA into the proviral DNA in the cytoplasm. The relatively low fidelity of HIV-1 RT was implicated as a dominant factor contributing to the genetic variability of the virus. The lack of intrinsic 3' --> 5' exonuclease activity, the formation of 3'-mispaired DNA and the subsequent extension of this DNA were shown to be determinants for the low fidelity of HIV-1 RT. It was of interest to analyse whether the cytoplasmic proteins may affect the accuracy of DNA synthesis by RT. We investigated the fidelity of DNA synthesis by HIV-1 RT with and without exonucleolytic proofreading provided by cytoplasmic fraction of LCC2 cells expressing high level of wild-type functional p53. Two basic features related to fidelity of DNA synthesis were studied: the misinsertion and mispair extension. The misincorporation of noncomplementary deoxynucleotides into nascent DNA and subsequent mispair extension by HIV-1 RT were substantially decreased in the presence of cytoplasmic fraction of LCC2 cells with both RNA/DNA and DNA/DNA template-primers with the same target sequence. The mispair extension frequencies obtained with the HIV-1 RT in the presence of cytoplasmic fraction of LCC2 cells were significantly lower (about 2.8-15-fold) than those detected with the purified enzyme. In addition, the productive interaction between polymerization (by HIV-1 RT) and exonuclease (by p53 in cytoplasm) activities was observed; p53 preferentially hydrolyses mispaired 3'-termini, permitting subsequent extension of the correctly paired 3'-terminus by HIV-1 RT. The data suggest that p53 in cytoplasm may affect the accuracy of DNA replication and the mutation spectra of HIV-1 RT by acting as an external proofreader. Furthermore, the decrease in error prone DNA synthesis with RT in the presence of external exonuclease, provided by cytoplasmic p53, may partially account for lower mutation rate of HIV-1 observed in vivo. PMID- 15286712 TI - Comparable transforming capacities and differential gene expression patterns of variant FUS/CHOP fusion transcripts derived from soft tissue liposarcomas. AB - The chromosomal translocation t(12;16)(q13;p11) is a common genetic alteration in myxoid and round-cell liposarcomas. It results in transcription of various chimeric FUS/CHOP fusion transcripts that encode different oncogenic proteins. Recent reports suggest that these may have different neoplastic transformation activities. To audit this hypothesis, we transfected expression plasmids for the two major variant FUS/CHOP transcripts I and II in NIH 3T3 cells and determined the number of outgrowing foci as well as their growth potential in soft agar. In addition, we compared tumour growth in nude mice upon subcutaneous injection of the respective transfectants. No significant differences in transformation assays in vitro and in vivo were observed, suggesting that both variant transcripts confer comparable transforming activities. The histopathological picture of tumours derived from both cell populations resembles high-grade spindle cell sarcomas. This suggests that both FUS/CHOP variants cause similar patterns of differential gene expression. This hypothesis was confirmed by mRNA-expression profiles of the respective cell clones. Strong overexpression of the pentaxin related gene (PTX), the osteoblast-specific factor 2 (osf-2), the basic Kruppel like factor (bklf), the leucoprotease inhibitor, and the cyclophilin B were observed in both types of FUS/CHOP-transfected cell clones. Taken together, our data suggest that different FUS/CHOP variants cause transformation of mesenchymal cells via the same pathways with comparable efficacy. PMID- 15286713 TI - Apoptosis-inducing factor determines the chemoresistance of non-small-cell lung carcinomas. AB - Non-small-cell lung carcinomas (NSCLCs) are resistant to the induction of apoptosis by conventional anticancer treatment. However, NSCLC cell lines are sensitive to the action of the broad protein kinase inhibitor, staurosporine (STS). In the NSCLC cell line U1810, STS induced the mitochondrial release of apoptosis-inducing factor (AIF) and cytochrome c (Cyt c) followed by activation of caspases, nuclear condensation, DNA fragmentation and finally cell death. Although preincubation of U1810 cells with the broad-spectrum caspase inhibitor z VAD.fmk delayed the occurrence of nuclear apoptosis induced by STS, it did not impede mitochondrial alterations (such as the release of Cyt c and AIF) and cell death to occur. Moreover, the microinjection of neither Cyt c nor recombinant active caspase-3 into the cytoplasm promoted nuclear apoptosis-related changes in U1810 cells. Evaluation of the role of the caspase-independent factor AIF in STS mediated death revealed that, upon immunodepletion of AIF, cytosols from STS treated U1810 lost their capacity to induce nuclear condensation when incubated with isolated nuclei. In addition, microinjection of an anti-AIF antibody prevented AIF from translocating to the nuclei of STS-treated U1810 cells and reduced STS-induced cell death. Finally, although the transfection-enforced overexpression of AIF was not sufficient to induce cell death, it did enhance STS mediated cell killing. Altogether, these results indicate that activation of caspases is not sufficient to kill U1810 cells and rather suggests an important role for the AIF-mediated mitochondrial-mediated death pathway. PMID- 15286714 TI - Parity-induced mammary epithelial cells facilitate tumorigenesis in MMTV-neu transgenic mice. AB - Using a Cre-lox-based genetic labeling technique, we have recently discovered a parity-induced mammary epithelial subtype that is abundant in nonlactating and nonpregnant, parous females. These mammary epithelial cells serve as alveolar progenitors in subsequent pregnancies, and transplantation studies revealed that they possess features of multipotent progenitors such as self-renewal and the capability to contribute to ductal and alveolar morphogenesis. Here, we report that these cells are the cellular targets for transformation in MMTV-neu transgenic mice that exhibit accelerated mammary tumorigenesis in multiparous animals. The selective ablation of this epithelial subtype reduces the onset of tumorigenesis in multiparous MMTV-neu transgenics. There is, however, experimental evidence to suggest that parity-induced mammary epithelial cells may not be the only cellular targets in other MMTV-promoter-based transgenic strains. In particular, the heterogeneous MMTV-wnt1 lesions predominantly express the ductal differentiation marker Nkcc1 that is absent in MMTV-neu-derived tumors. Our observations support the idea that tumors originate from distinctly different epithelial subtypes in selected MMTV-promoter-driven cancer models and that diverse oncogenes might exert discrete effects on particular mammary epithelial subtypes. PMID- 15286715 TI - The farnesyl transferase inhibitor R115777 (Zarnestra) synergistically enhances growth inhibition and apoptosis induced on epidermoid cancer cells by Zoledronic acid (Zometa) and Pamidronate. AB - Pamidronate (PAM) and zoledronic acid (ZOL) are aminobisphosphonates (BPs) able to affect the isoprenylation of intracellular small G proteins. We have investigated the antitumor activity of BPs and R115777 farnesyl transferase inhibitor (FTI) against epidermoid cancer cells. In human epidermoid head and neck KB and lung H1355 cancer cells, 48 h exposure to PAM and ZOL induced growth inhibition (IC(50) 25 and 10 microM, respectively) and apoptosis and abolished the proliferative and antiapoptotic stimuli induced by epidermal growth factor (EGF). In these experimental conditions, ZOL induced apoptosis through the activation of caspase 3 and a clear fragmentation of PARP was also demonstrated. A strong decrease of basal ras activity and an antagonism on its stimulation by EGF was recorded in the tumor cells exposed to BPs. These effects were paralleled by impaired activation of the survival enzymes extracellular signal regulated kinase 1 and 2 (Erk-1/2) and Akt that were not restored by EGF. Conversely, farnesol induced a recovery of ras activity and antagonized the proapoptotic effects induced by BPs. The combined treatment with BPs and R115777 resulted in a strong synergism both in growth inhibition and apoptosis in KB and H1355 cells. The synergistic activity between the drugs allowed BPs to produce tumor cell growth inhibition and apoptosis at in vivo achievable concentrations (0.1 micromolar for both drugs). Moreover, the combination was highly effective in the inhibition of ras, Erk and Akt activity, while farnesol again antagonized these effects. In conclusion, the combination of BPs and FTI leads to enhanced antitumor activity at clinically achievable drug concentrations that resides in the inhibition of farnesylation-dependent survival pathways and warrants further studies for clinical translation. PMID- 15286716 TI - Characterization of a conserved aphidicolin-sensitive common fragile site at human 4q22 and mouse 6C1: possible association with an inherited disease and cancer. AB - Fragile sites are classified as common or rare depending on their occurrence in the populations. While rare sites are mainly associated with inherited diseases, common sites have been involved in somatic rearrangements found in the chromosomes of cancer cells. Here we study a mouse locus containing the ionotropic glutamate receptor delta 2 (grid2) gene in which spontaneous chromosome rearrangements occur frequently, giving rise to mutant animals in inbred populations. We identify and clone common fragile sites overlapping the mouse grid2 gene and its human ortholog GRID2, lying respectively at bands 6C1 and 4q22 in a 7-Mb-long region of synteny. These results show a third example of orthologous common sites conserved at the molecular level, and reveal an unexpected link between an inherited disease and an aphidicolin-sensitive region. Recurrent deletions of subregions of band 4q22 have been previously described in human hepatocellular carcinomas. This 15-Mb-long region appears precisely centered on the site described here, which strongly suggests that it also plays a specific role in hepatic carcinogenesis. PMID- 15286717 TI - Inhibitory effects of cyclosporin A on calcium mobilization-dependent interleukin 8 expression and invasive potential of human glioblastoma U251MG cells. AB - Interleukin (IL)-8 produced from glioblastoma is suggested to contribute to its own proliferation and progression. Since various external stimuli have been shown to increase intracellular Ca(2+) in glioma cells, we investigated Ca(2+) mobilization-dependent IL-8 expression and effect of cyclosporin A (CsA), an inhibitor of calcineurin (Cn), on the expression and invasive potential of human glioblastoma U251MG cells. Combined treatment with Ca(2+)-ionophore and phorbol myristate-acetate (A23187/PMA) increased IL-8 mRNA and protein levels. This increase was suppressed by CsA and by another Cn inhibitor FK506. Luciferase reporter gene assay and electrophoretic mobility shift assay revealed that activation of p65-containing nuclear factor-kappaB was essential for A23187/PMA dependent activation of IL-8 promoter. CsA suppressed the promoter activity by attenuating IkappaB-alpha degradation. U251MG cells expressed IL-8 receptors CXCR 1 and -2, and Matrigel invasion assay revealed that CsA attenuated A23187/PMA dependent stimulation of invasive potential, probably by inhibiting IL-8 production. In addition, IL-8-dependent proliferation was also suppressed by CsA. Taken together, these results demonstrate the novel inhibitory effects of CsA on glioblastoma cell functions, suggesting CsA as a potential therapeutic adjuvant for glioma treatment. PMID- 15286718 TI - Protein patterns and proteins that identify subtypes of glioblastoma multiforme. AB - Glioblastoma multiforme (GBM) has been subdivided into two types based on clinical and genetic findings: primary tumors, which arise de novo, and secondary tumors, which progress from lower grade gliomas to GBMs. To analyse this dichotomy at the protein level, we employed selective tissue microdissection to obtain pure populations of tumor cells, which we studied using two-dimensional protein gel electrophoresis (2-DGE) and protein sequencing of select target proteins. Protein patterns were analysed in a blinded manner from the clinical and genetic data. 2-DGE clearly identified two distinct populations of tumors. 2 DGE was reproducible and reliable, as multiple samples analysed from the same patient gave identical results. In addition, we isolated and sequenced 11 proteins that were uniquely expressed in either the primary or the secondary GBMs, but not both. We demonstrate that specific proteomic patterns can be reproducibly identified by two-dimensional gel electrophoresis from limited numbers of selectively procured, microdissected tumor cells and that two patterns of GBMs, primary versus secondary, previously distinguished by clinical and genetic differences, can be recognized at the protein level. Proteins that are expressed distinctively may have important implications for the diagnosis, prognosis, and treatment of patients with GBM. PMID- 15286719 TI - Transcriptional activation of c-myc proto-oncogene by WT1 protein. AB - The Wilms' tumor 1 gene (WT1) plays an essential role in urogenital development and malignancy. Through DNA binding, WT1 can either enhance or repress transcription depending on the context of the DNA-binding sites or the cell type in which it is expressed. WT1 is overexpressed in a variety of human cancers, including leukemia and breast cancer; in these diseases, the expression of WT1 is associated with a poor prognosis. To determine how WT1 affects c-myc expression in the context of breast cancer cells, we have examined the ability of both endogenous and exogenous WT1 proteins in breast cancer cells to bind to the c-myc promoter in vivo. Using c-myc-promoter-driven luciferase constructs, we found that different forms of WT1 could enhance the expression of the reporter. Unlike other studies where WT1 is reported to be a negative regulator of c-myc, we found that both the - and + KTS forms of WT1 could act to enhance c-myc expression, depending on the cell type. The WT1-binding site near the second major transcription start site of the c-myc promoter was confirmed to be involved in upregulation of human c-myc by WT1. Finally, we demonstrated that overexpression of WT1 induced a significant increase in the abundance of endogenous c-myc protein in breast cancer cells, consistent with the upregulation of c-myc transcription following WT1 induction. These observations strongly argue that in the case of breast cancer WT1 is functioning as an oncogene in part by stimulating the expression of c-myc. PMID- 15286720 TI - Mechanochemical coupling of two substeps in a single myosin V motor. AB - Myosin V is a double-headed processive molecular motor that moves along an actin filament by taking 36-nm steps. Using optical trapping nanometry with high spatiotemporal resolution, we discovered that there are two possible pathways for the 36-nm steps, one with 12- and 24-nm substeps, in this order, and the other without substeps. Based on the analyses of effects of ATP, ADP and 2,3 butanedione 2-monoxime (a reagent shown here to slow ADP release from actomyosin V) on the dwell time and the occurrence frequency of the main and the intermediate states, we propose that the 12-nm substep occurs after ATP binding to the bound trailing head and the 24-nm substep results from a mechanical step following the isomerization of an actomyosin-ADP state on the bound leading head. When the isomerization precedes the 12-nm substep, the 36-nm step occurs without substeps. PMID- 15286721 TI - The PM2 virion has a novel organization with an internal membrane and pentameric receptor binding spikes. AB - Biological membranes are notoriously resistant to structural analysis. Excellent candidates to tackle this problem in situ are membrane-containing viruses where the membrane is constrained by an icosahedral capsid. Cryo-EM and image reconstruction of bacteriophage PM2 revealed a membrane bilayer following the internal surface of the capsid. The viral genome closely interacts with the inner leaflet. The capsid, at a resolution of 8.4 A, reveals 200 trimeric capsomers with a pseudo T = 21 dextro organization. Pentameric receptor-binding spikes protrude from the surface. It is evident from the structure that the PM2 membrane has at least two important roles in the life cycle. First, it acts as a scaffold to nucleate capsid assembly. Second, after host recognition, it fuses with the host outer membrane to promote genome entry. The structure also sheds light on how the viral supercoiled circular double-stranded DNA genome might be packaged and released. PMID- 15286722 TI - An antibiotic factory caught in action. AB - The synthesis of aromatic polyketides, such as actinorhodin, tetracycline and doxorubicin, begins with the formation of a polyketide chain. In type II polyketide synthases (PKSs), chains are polymerized by the heterodimeric ketosynthase-chain length factor (KS-CLF). Here we present the 2.0-A structure of the actinorhodin KS-CLF, which shows polyketides being elongated inside an amphipathic tunnel approximately 17 A in length at the heterodimer interface. The structure resolves many of the questions about the roles of KS and CLF. Although CLF regulates chain length, it does not have an active site; KS must catalyze both chain initiation and elongation. We provide evidence that the first cyclization of the polyketide occurs within the KS-CLF tunnel. The mechanistic details of this central PKS polymerase could guide biosynthetic chemists in designing new pharmaceuticals and polymers. PMID- 15286723 TI - A novel tunnel in mycobacterial type III polyketide synthase reveals the structural basis for generating diverse metabolites. AB - The superfamily of plant and bacterial type III polyketide synthases (PKSs) produces diverse metabolites with distinct biological functions. PKS18, a type III PKS from Mycobacterium tuberculosis, displays an unusual broad specificity for aliphatic long-chain acyl-coenzyme A (acyl-CoA) starter units (C(6)-C(20)) to produce tri- and tetraketide pyrones. The crystal structure of PKS18 reveals a 20 A substrate binding tunnel, hitherto unidentified in this superfamily of enzymes. This remarkable tunnel extends from the active site to the surface of the protein and is primarily generated by subtle changes of backbone dihedral angles in the core of the protein. Mutagenic studies combined with structure determination provide molecular insights into the structural elements that contribute to the chain length specificity of the enzyme. This first bacterial type III PKS structure underlines a fascinating example of the way in which subtle changes in protein architecture can generate metabolite diversity in nature. PMID- 15286724 TI - Myosin VI walks hand-over-hand along actin. AB - Myosin VI is a molecular motor that can walk processively on actin filaments with a 36-nm step size. The walking mechanism of myosin VI is controversial because it takes very large steps without an apparent lever arm of required length. Therefore, myosin VI is argued to be the first exception to the widely established lever arm theory. It is therefore critical to directly demonstrate whether this motor walks hand-over-hand along actin despite its short lever arm. Here, we follow the displacement of a single myosin VI head during the stepping process. A single head is displaced 72 nm during stepping, whereas the center of mass previously has been shown to move 36 nm. The most likely explanation for this result is a hand-over-hand walking mechanism. We hypothesize the existence of a flexible element that would allow the motor to bridge the observed 72-nm distance. PMID- 15286726 TI - Helping the CD8(+) T-cell response. PMID- 15286725 TI - Fibrotic disease and the T(H)1/T(H)2 paradigm. PMID- 15286727 TI - Transmembrane adaptor proteins: organizers of immunoreceptor signalling. PMID- 15286728 TI - Matrix metalloproteinases as modulators of inflammation and innate immunity. PMID- 15286729 TI - HIV and SIV CTL escape: implications for vaccine design. PMID- 15286730 TI - Inflammation and necrosis promote tumour growth. PMID- 15286731 TI - Seventeen-colour flow cytometry: unravelling the immune system. PMID- 15286732 TI - A new oral anticoagulant: the 50-year challenge. PMID- 15286733 TI - Fragment-based lead discovery. PMID- 15286734 TI - Drug repositioning: identifying and developing new uses for existing drugs. PMID- 15286735 TI - Recombinant biologics for treatment of bleeding disorders. PMID- 15286736 TI - Lipid metabolic enzymes: emerging drug targets for the treatment of obesity. PMID- 15286737 TI - Can the pharmaceutical industry reduce attrition rates? PMID- 15286738 TI - Overweight, obesity and cancer: epidemiological evidence and proposed mechanisms. PMID- 15286739 TI - Pathways of apoptotic and non-apoptotic death in tumour cells. PMID- 15286740 TI - Biologically active sphingolipids in cancer pathogenesis and treatment. PMID- 15286741 TI - X-chromosome genetics and human cancer. PMID- 15286742 TI - Chemical-induced DNA damage and human cancer risk. PMID- 15286743 TI - Integrated global profiling of cancer. PMID- 15286744 TI - MALT lymphoma: from morphology to molecules. PMID- 15286745 TI - Making an impact? PMID- 15286746 TI - Public attitudes to nanotechnology in Europe and the United States. PMID- 15286747 TI - Metastable high-pressure phases of low-Z compounds: creation of a new chemistry or a prompt for old principles? AB - Recent experiments demonstrate that high pressure is a powerful tool for the synthesis of new unusual inorganic polymers consisting of low-Z elements. However, experience within organic chemistry, for example, polyethylene, provides evidence that polymeric phases with high thermal stability can be potentially synthesised by conventional chemical techniques without applying high pressure. PMID- 15286749 TI - Electrochemistry: arrays of polymer nanowires. PMID- 15286750 TI - Nanoceramic composites: tough to test. PMID- 15286751 TI - Organic electronics: supra solutions. PMID- 15286752 TI - Polymer composites: swell properties and swift processing. PMID- 15286753 TI - Material witness: faux food to the rescue? PMID- 15286754 TI - Nanostructuring of metals by severe plastic deformation for advanced properties. AB - Despite rosy prospects, the use of nanostructured metals and alloys as advanced structural and functional materials has remained controversial until recently. Only in recent years has a breakthrough been outlined in this area, associated both with development of new routes for the fabrication of bulk nanostructured materials and with investigation of the fundamental mechanisms that lead to the new properties of these materials. Although a deep understanding of these mechanisms is still a topic of basic research, pilot commercial products for medicine and microdevices are coming within reach of the market. This progress article discusses new concepts and principles of using severe plastic deformation (SPD) to fabricate bulk nanostructured metals with advanced properties. Special emphasis is laid on the relationship between microstructural features and properties, as well as the first applications of SPD-produced nanomaterials. PMID- 15286755 TI - Carbon nanotube filters. AB - Over the past decade of nanotube research, a variety of organized nanotube architectures have been fabricated using chemical vapour deposition. The idea of using nanotube structures in separation technology has been proposed, but building macroscopic structures that have controlled geometric shapes, density and dimensions for specific applications still remains a challenge. Here we report the fabrication of freestanding monolithic uniform macroscopic hollow cylinders having radially aligned carbon nanotube walls, with diameters and lengths up to several centimetres. These cylindrical membranes are used as filters to demonstrate their utility in two important settings: the elimination of multiple components of heavy hydrocarbons from petroleum-a crucial step in post-distillation of crude oil-with a single-step filtering process, and the filtration of bacterial contaminants such as Escherichia coli or the nanometre sized poliovirus ( approximately 25 nm) from water. These macro filters can be cleaned for repeated filtration through ultrasonication and autoclaving. The exceptional thermal and mechanical stability of nanotubes, and the high surface area, ease and cost-effective fabrication of the nanotube membranes may allow them to compete with ceramic- and polymer-based separation membranes used commercially. PMID- 15286756 TI - Subwavelength-scale tailoring of surface phonon polaritons by focused ion-beam implantation. AB - Recent advances in optical nanotechnologies by controlling surface plasmon polaritons in metallic nanostructures demonstrate high potential for subwavelength-scale waveguiding of light, data storage, microscopy or biophotonics. Surprisingly, surface phonon polaritons-infrared counterparts to surface plasmon polaritons-have not been widely explored for nanophotonic applications. As they rely on the infrared or terahertz excitation of lattice vibrations in polar crystals they offer totally different material classes for nanophotonic applications, such as semiconductors and insulators. In an initial step towards nanoscale surface phonon photonics we show evidence that the local properties of surface phonon polaritons can be tailored at a subwavelength-scale by focused ion-beam modification of the crystal structure, even without significant alteration of the surface topography. Such single-step-fabricated, monolithic structures could be used for controlling electromagnetic energy transport by surface phonon polaritons in miniaturized integrated devices operating at infrared or terahertz frequencies. We verify the polaritonic properties of an ion-beam-patterned SiC surface by infrared near-field microscopy. The near-field images also demonstrate nanometre-scale resolved infrared mapping of crystal quality useful in semiconductor processing or crystal growth. PMID- 15286757 TI - Science, not politics. PMID- 15286758 TI - Asian epidemic, abstinence top AIDS meeting agenda. PMID- 15286759 TI - Research with premature infant livers touches a nerve. PMID- 15286760 TI - London calling for British medical research institute. PMID- 15286761 TI - Disputed expenses at heart of Moncada's Spanish spat. PMID- 15286763 TI - Experts call for smarter approach to targeted therapies. PMID- 15286764 TI - Ovarian transplants restore fertility in cancer patients. PMID- 15286766 TI - Profile: Irving Weissman. PMID- 15286767 TI - The virtuous virus. PMID- 15286768 TI - Urgently needed: a filter for the HIV-1 vaccine pipeline. AB - Research groups worldwide are trying to make immunogens intended to elicit neutralizing antibody responses as part of a prophylactic vaccine to counter the spread of HIV-1. The relative merits of different designs can only be gauged properly through comparative studies, and particularly by evaluating human or animal antisera under identical, or comparable, conditions. Hence there is a need for assay standardization and for the creation of a centralized testing facility that could distribute consensus protocols and reagents. PMID- 15286770 TI - RNAi quashes polyQ. PMID- 15286772 TI - Breast cancer progression with a Twist. PMID- 15286773 TI - Redistricting the retroviral restriction factors. PMID- 15286774 TI - Staphylococcal protein A inflames the lungs. PMID- 15286775 TI - Gene delivery goes global. PMID- 15286776 TI - Type 1 diabetes: focus on prevention. PMID- 15286778 TI - Treating cancer's kinase 'addiction'. PMID- 15286780 TI - Cancer genes and the pathways they control. AB - The revolution in cancer research can be summed up in a single sentence: cancer is, in essence, a genetic disease. In the last decade, many important genes responsible for the genesis of various cancers have been discovered, their mutations precisely identified, and the pathways through which they act characterized. The purposes of this review are to highlight examples of progress in these areas, indicate where knowledge is scarce and point out fertile grounds for future investigation. PMID- 15286781 TI - Regulatory T cells and mechanisms of immune system control. AB - The immune system evolved to protect the host against the attack of foreign, potentially pathogenic, microorganisms. It does so by recognizing antigens expressed by those microorganisms and mounting an immune response against all cells expressing them, with the ultimate aim of their elimination. Various mechanisms have been reported to control and regulate the immune system to prevent or minimize reactivity to self-antigens or an overexuberant response to a pathogen, both of which can result in damage to the host. Deletion of autoreactive cells during T- and B-cell development allows the immune system to be tolerant of most self-antigens. Peripheral tolerance to self was suggested several years ago to result from the induction of anergy in peripheral self reactive lymphocytes. More recently, however, it has become clear that avoidance of damage to the host is also achieved by active suppression mediated by regulatory T (T(reg)) cell populations. We discuss here the varied mechanisms used by T(reg) cells to suppress the immune system. PMID- 15286782 TI - Correlates of immune protection in HIV-1 infection: what we know, what we don't know, what we should know. AB - The field of vaccinology began in ignorance of how protection was instilled in vaccine recipients. Today, a greater knowledge of immunology allows us to better understand what is being stimulated by various vaccines that leads to their protective effects: that is, their correlates of protection. Here we describe what is known about the correlates of protection for existing vaccines against a range of different viral diseases and discuss the correlates of protection against disease during natural infection with HIV-1. We will also discuss why it is important to design phase 3 clinical trials of HIV vaccines to determine the correlates of protection for each individual vaccine. PMID- 15286785 TI - Donor APCs are required for maximal GVHD but not for GVL. AB - Graft-versus-host disease (GVHD) is a major source of morbidity in allogenic stem cell transplantation. We previously showed that recipient antigen-presenting cells (APCs) are required for CD8-dependent GVHD in a mouse model across only minor histocompatibility antigens (minor H antigens). However, these studies did not address the function of donor-derived APCs after GVHD is initiated. Here we show that GVHD develops in recipients of donor major histocompatibility complex class I-deficient (MHC I(-)) bone marrow. Thus, after initial priming, CD8 cells caused GVHD without a further requirement for hematopoietic APCs, indicating that host APCs are necessary and sufficient for GHVD. Nonetheless, GVHD was less severe in recipients of MHC I(-) bone marrow. Therefore, once initiated, GVHD is intensified by donor-derived cells, most probably donor APCs cross-priming alloreactive CD8 cells. Nevertheless, donor APCs were not required for CD8 mediated graft-versus-leukemia (GVL) against a mouse model of chronic-phase chronic myelogenous leukemia. These studies identify donor APCs as a new target for treating GVHD, which may preserve GVL. PMID- 15286784 TI - Reticulon family members modulate BACE1 activity and amyloid-beta peptide generation. AB - Inhibiting the activity of the beta-amyloid converting enzyme 1 (BACE1) or reducing levels of BACE1 in vivo decreases the production of amyloid-beta. The reticulon family of proteins has four members, RTN1, RTN2, RTN3 and RTN4 (also known as Nogo), the last of which is well known for its role in inhibiting neuritic outgrowth after injury. Here we show that reticulon family members are binding partners of BACE1. In brain, BACE1 mainly colocalizes with RTN3 in neurons, whereas RTN4 is more enriched in oligodendrocytes. An increase in the expression of any reticulon protein substantially reduces the production of Abeta. Conversely, lowering the expression of RTN3 by RNA interference increases the secretion of Abeta, suggesting that reticulon proteins are negative modulators of BACE1 in cells. Our data support a mechanism by which reticulon proteins block access of BACE1 to amyloid precursor protein and reduce the cleavage of this protein. Thus, changes in the expression of reticulon proteins in the human brain are likely to affect cellular amyloid-beta and the formation of amyloid plaques. PMID- 15286786 TI - Evidence for a fruit fly hemangioblast and similarities between lymph-gland hematopoiesis in fruit fly and mammal aorta-gonadal-mesonephros mesoderm. AB - The Drosophila melanogaster lymph gland is a hematopoietic organ and, together with prospective vascular cells (cardioblasts) and excretory cells (pericardial nephrocytes), arises from the cardiogenic mesoderm. Clonal analysis provided evidence for a hemangioblast that can give rise to two daughter cells: one that differentiates into heart or aorta and another that differentiates into blood. In addition, the GATA factor gene pannier (pnr) and the homeobox gene tinman (tin), which are controlled by the convergence of Decapentaplegic (Dpp), fibroblast growth factor (FGF), Wingless (Wg) and Notch signaling, are required for the development of all cardiogenic mesoderm, including the lymph gland. Here we show that an essential genetic switch that differentiates between the blood or nephrocyte and vascular lineages involves the Notch pathway. Further specification occurs through specific expression of the GATA factor Serpent (Srp) in the lymph-gland primordium. Our findings suggest that there is a close parallel between the molecular mechanisms functioning in the D. melanogaster cardiogenic mesoderm and those functioning in the mammalian aorta-gonadal mesonephros mesoderm. PMID- 15286787 TI - Mutations in SLC6A19, encoding B0AT1, cause Hartnup disorder. AB - Hartnup disorder, an autosomal recessive defect named after an English family described in 1956 (ref. 1), results from impaired transport of neutral amino acids across epithelial cells in renal proximal tubules and intestinal mucosa. Symptoms include transient manifestations of pellagra (rashes), cerebellar ataxia and psychosis. Using homozygosity mapping in the original family in whom Hartnup disorder was discovered, we confirmed that the critical region for one causative gene was located on chromosome 5p15 (ref. 3). This region is homologous to the area of mouse chromosome 13 that encodes the sodium-dependent amino acid transporter B(0)AT1 (ref. 4). We isolated the human homolog of B(0)AT1, called SLC6A19, and determined its size and molecular organization. We then identified mutations in SLC6A19 in members of the original family in whom Hartnup disorder was discovered and of three Japanese families. The protein product of SLC6A19, the Hartnup transporter, is expressed primarily in intestine and renal proximal tubule and functions as a neutral amino acid transporter. PMID- 15286788 TI - Hartnup disorder is caused by mutations in the gene encoding the neutral amino acid transporter SLC6A19. AB - Hartnup disorder (OMIM 234500) is an autosomal recessive abnormality of renal and gastrointestinal neutral amino acid transport noted for its clinical variability. We localized a gene causing Hartnup disorder to chromosome 5p15.33 and cloned a new gene, SLC6A19, in this region. SLC6A19 is a sodium-dependent and chloride independent neutral amino acid transporter, expressed predominately in kidney and intestine, with properties of system B(0). We identified six mutations in SLC6A19 that cosegregated with disease in the predicted recessive manner, with most affected individuals being compound heterozygotes. The disease-causing mutations that we tested reduced neutral amino acid transport function in vitro. Population frequencies for the most common mutated SLC6A19 alleles are 0.007 for 517G --> A and 0.001 for 718C --> T. Our findings indicate that SLC6A19 is the long-sought gene that is mutated in Hartnup disorder; its identification provides the opportunity to examine the inconsistent multisystemic features of this disorder. PMID- 15286789 TI - Detection of large-scale variation in the human genome. AB - We identified 255 loci across the human genome that contain genomic imbalances among unrelated individuals. Twenty-four variants are present in > 10% of the individuals that we examined. Half of these regions overlap with genes, and many coincide with segmental duplications or gaps in the human genome assembly. This previously unappreciated heterogeneity may underlie certain human phenotypic variation and susceptibility to disease and argues for a more dynamic human genome structure. PMID- 15286790 TI - Temporal plasticity in the primary auditory cortex induced by operant perceptual learning. AB - Processing of rapidly successive acoustic stimuli can be markedly improved by sensory training. To investigate the cortical mechanisms underlying such temporal plasticity, we trained rats in a 'sound maze' in which navigation using only auditory cues led to a target location paired with food reward. In this task, the repetition rate of noise pulses increased as the distance between the rat and target location decreased. After training in the sound maze, neurons in the primary auditory cortex (A1) showed greater responses to high-rate noise pulses and stronger phase-locking of responses to the stimuli; they also showed shorter post-stimulation suppression and stronger rebound activation. These improved temporal dynamics transferred to trains of pure-tone pips. Control animals that received identical sound stimulation but were given free access to food showed the same results as naive rats. We conclude that this auditory perceptual learning results in improvements in temporal processing, which may be mediated by enhanced cortical response dynamics. PMID- 15286791 TI - Neural fate of ignored stimuli: dissociable effects of perceptual and working memory load. AB - Observers commonly experience functional blindness to unattended visual events, and this problem has fuelled an intense debate concerning the fate of unattended visual information in neural processing. Here we used functional magnetic resonance imaging (fMRI) to demonstrate that the type of task that a human subject engages in determines the way in which ignored visual background stimuli are processed in parahippocampal cortex. Increasing the perceptual difficulty of a foveal target task attenuated processing of task-irrelevant background scenes, whereas increasing the number of objects held in working memory did not have this effect. These dissociable effects of perceptual and working memory load clarify how task-irrelevant, unattended stimuli are processed in category-selective areas in human ventral visual cortex. PMID- 15286792 TI - The sequential activity of the GTPases Rap1B and Cdc42 determines neuronal polarity. AB - The establishment of a polarized morphology is an essential step in the differentiation of neurons with a single axon and multiple dendrites. In cultured rat hippocampal neurons, one of several initially indistinguishable neurites is selected to become the axon. Both phosphatidylinositol 3,4,5-trisphosphate and the evolutionarily conserved Par complex (comprising Par3, Par6 and an atypical PKC (aPKC) such as PKClambda or PKCzeta) are involved in axon specification. However, the initial signals that establish cellular asymmetry and the pathways that subsequently translate it into structural changes remain to be elucidated. Here we show that localization of the GTPase Rap1B to the tip of a single neurite is a decisive step in determining which neurite becomes the axon. Using GTPase mutants and RNA interference, we found that Rap1B is necessary and sufficient to initiate the development of axons upstream of Cdc42 and the Par complex. PMID- 15286793 TI - Adding muscle to neuromuscular degeneration investigation. PMID- 15286794 TI - Umbilical cord blood cells and brain stroke injury: bringing in fresh blood to address an old problem. AB - Degeneration of brain tissue following stroke leads to functional impairment with limited brain self-repair. New evidence suggests that delivery of circulating CD34(+) human umbilical cord blood cells can produce functional recovery in an animal stroke model with concurrent angiogenesis and neurogenesis leading to some restoration of cortical tissue. While some alternative interpretations of this data are offered herein, the study provides encouraging evidence of functional recovery from stroke in an animal model using stem cell therapy. PMID- 15286795 TI - Unlocking the secrets of the pancreatic beta cell: man and mouse provide the key. AB - Failure of the pancreas to secrete sufficient insulin results in type 2 diabetes, but the pathogenesis of pancreatic beta cell dysfunction is still poorly understood. New insights into beta cell failure come from defining the genes involved in rare genetic subtypes of diabetes and creating appropriate animal models. A new mouse model of transient neonatal diabetes mellitus emphasizes that both the number of beta cells and their function are critical for insulin secretion and may be regulated by imprinted genes. PMID- 15286796 TI - IL-5 links adaptive and natural immunity in reducing atherosclerotic disease. AB - Oxidized LDL induces changes in several facets of the immune system, although the relationships between these facets and their contributions to atherogenesis have yet to be fully elucidated. A report in this issue of the JCI provides a novel demonstration of the adaptive immune system influencing the production of natural antibodies. The results demonstrate that injection of malondialdehyde-modified LDL promotes a Th2 response that in turn increases the titers of the natural antibody T15/EO6, which recognizes the oxidized phospholipid POVPC. Atherosclerotic lesion size in LDL receptor-deficient mice is reduced as a consequence of the increase in natural antibody titers, and IL-5 is identified as the link between the adaptive and natural immune systems. PMID- 15286797 TI - Pulmonary fibrosis: thinking outside of the lung. AB - Pulmonary fibrosis is a devastating condition that leads to progressive lung destruction and scarring. Previous mechanistic research has focused on the local fibroproliferative process in the lung. However, emerging evidence suggests that circulating cells of hematopoietic origin play a crucial role in the pathogenesis of this disease. PMID- 15286798 TI - Obstruction of extrahepatic bile ducts by lymphocytes is regulated by IFN-gamma in experimental biliary atresia. AB - The etiology and pathogenesis of bile duct obstruction in children with biliary atresia are largely unknown. We have previously reported that, despite phenotypic heterogeneity, genomic signatures of livers from patients display a proinflammatory phenotype. Here, we address the hypothesis that production of IFN gamma is a key pathogenic mechanism of disease using a mouse model of rotavirus induced biliary atresia. We found that rotavirus infection of neonatal mice has a unique tropism to bile duct cells, and it triggers a hepatobiliary inflammation by IFN-gamma-producing CD4(+) and CD8(+) lymphocytes. The inflammation is tissue specific, resulting in progressive jaundice, growth failure, and greater than 90% mortality due to obstruction of extrahepatic bile ducts. In this model, the genetic loss of IFN-gamma did not alter the onset of jaundice, but it remarkably suppressed the tissue-specific targeting of T lymphocytes and completely prevented the inflammatory and fibrosing obstruction of extrahepatic bile ducts. As a consequence, jaundice resolved, and long-term survival improved to greater than 80%. Notably, administration of recombinant IFN-gamma led to recurrence of bile duct obstruction following rotavirus infection of IFN-gamma-deficient mice. Thus, IFN-gamma-driven obstruction of bile ducts is a key pathogenic mechanism of disease and may constitute a therapeutic target to block disease progression in patients with biliary atresia. PMID- 15286799 TI - Administration of CD34+ cells after stroke enhances neurogenesis via angiogenesis in a mouse model. AB - Thrombo-occlusive cerebrovascular disease resulting in stroke and permanent neuronal loss is an important cause of morbidity and mortality. Because of the unique properties of cerebral vasculature and the limited reparative capability of neuronal tissue, it has been difficult to devise effective neuroprotective therapies in cerebral ischemia. Our results demonstrate that systemic administration of human cord blood-derived CD34(+) cells to immunocompromised mice subjected to stroke 48 hours earlier induces neovascularization in the ischemic zone and provides a favorable environment for neuronal regeneration. Endogenous neurogenesis, suppressed by an antiangiogenic agent, is accelerated as a result of enhanced migration of neuronal progenitor cells to the damaged area, followed by their maturation and functional recovery. Our data suggest an essential role for CD34(+) cells in promoting directly or indirectly an environment conducive to neovascularization of ischemic brain so that neuronal regeneration can proceed. PMID- 15286800 TI - Impaired glucose homeostasis in transgenic mice expressing the human transient neonatal diabetes mellitus locus, TNDM. AB - Transient neonatal diabetes mellitus (TNDM) is a rare inherited diabetic syndrome apparent in the first weeks of life and again during early adulthood. The relative contributions of reduced islet beta cell number and impaired beta cell function to the observed hypoinsulinemia are unclear. The inheritance pattern of this imprinted disorder implicates overexpression of one or both genes within the TNDM locus: ZAC, which encodes a proapoptotic zinc finger protein, and HYMAI, which encodes an untranslated mRNA. To investigate the consequences for pancreatic function, we have developed a high-copy transgenic mouse line, TNDM29, carrying the human TNDM locus. TNDM29 neonates display hyperglycemia, and older adults, impaired glucose tolerance. Neonatal hyperglycemia occurs only on paternal transmission, analogous to paternal dependence of TNDM in humans. Embryonic pancreata of TNDM29 mice showed reductions in expression of endocrine differentiation factors and numbers of insulin-staining structures. By contrast, beta cell mass was normal or elevated at all postnatal stages, whereas pancreatic insulin content in neonates and peak serum insulin levels after glucose infusion in adults were reduced. Expression of human ZAC and HYMAI in these transgenic mice thus recapitulates key features of TNDM and implicates impaired development of the endocrine pancreas and beta cell function in disease pathogenesis. PMID- 15286801 TI - Central and peripheral actions of somatostatin on the growth hormone-IGF-I axis. AB - Somatostatin (SRIF) analogs provide safe and effective therapy for acromegaly. In a proportion of patients, however, SRIF analogs may lead to discordant growth hormone (GH) and IGF-I suppression, which suggests a more complex mechanism than attributable to inhibition of GH release alone. To elucidate whether SRIF acts peripherally on the GH-IGF-I axis, we showed that rat hepatocytes express somatostatin receptor subtypes-2 and -3 and that IGF-I mRNA and protein levels were suppressed in a dose-dependent manner by administration of octreotide. The inhibitory effect of SRIF was not apparent without added GH and in the presence of GH was specific for IGF-I induction and did not inhibit GH-induced c-myc or extracellular signal regulated kinase (ERK) phosphorylation. Pertussis toxin treatment of hepatocytes incubated with GH and SRIF, or with GH and octreotide, abrogated the inhibitory effect on GH-induced IGF-I, which confirms the requirement for the inhibitory G-protein. Treatment with SRIF and GH increased protein tyrosine phosphatase (PTP) activity and inhibited signal transducer and activator of transcription-5b (STAT5b) phosphorylation and nuclear localization. Octreotide also inhibited GH-stimulated IGF-I protein content of ex vivo-perfused rat livers. The results demonstrate that SRIF acts both centrally and peripherally to control the GH-IGF-I axis, providing a mechanistic explanation for SRIF analog action in treating patients with GH-secreting pituitary adenomas. PMID- 15286802 TI - Regulation of hypothalamic prohormone convertases 1 and 2 and effects on processing of prothyrotropin-releasing hormone. AB - Regulation of energy balance by leptin involves regulation of several neuropeptides, including thyrotropin-releasing hormone (TRH). Synthesized from a larger inactive precursor, its maturation requires proteolytic cleavage by prohormone convertases 1 and 2 (PC1 and PC2). Since this maturation in response to leptin requires prohormone processing, we hypothesized that leptin might regulate hypothalamic PC1 and PC2 expression, ultimately leading to coordinated processing of prohormones into mature peptides. Using hypothalamic neurons, we found that leptin stimulated PC1 and PC2 mRNA and protein expression and also increased PC1 and PC2 promoter activities in transfected 293T cells. Starvation of rats, leading to low serum leptin levels, decreased PC1 and PC2 gene and protein expression in the paraventricular nucleus (PVN) of the hypothalamus. Exogenous administration of leptin to fasted animals restored PC1 levels in the median eminence (ME) and the PVN to approximately the level found in fed control animals. Consistent with this regulation of PCs in the PVN, concentrations of TRH in the PVN and ME were substantially reduced in the fasted animals relative to the fed animals, and leptin reversed this decrease. Further analysis showed that proteolytic cleavage of pro-thyrotropin-releasing hormone (proTRH) at known PC cleavage sites was reduced by fasting and increased in animals given leptin. Combined, these findings suggest that leptin-dependent stimulation of hypothalamic TRH expression involves both activation of trh transcription and stimulation of PC1 and PC2 expression, which lead to enhanced processing of proTRH into mature TRH. PMID- 15286803 TI - Cancer cachexia is regulated by selective targeting of skeletal muscle gene products. AB - Cachexia is a syndrome characterized by wasting of skeletal muscle and contributes to nearly one-third of all cancer deaths. Cytokines and tumor factors mediate wasting by suppressing muscle gene products, but exactly which products are targeted by these cachectic factors is not well understood. Because of their functional relevance to muscle architecture, such targets are presumed to represent myofibrillar proteins, but whether these proteins are regulated in a general or a selective manner is also unclear. Here we demonstrate, using in vitro and in vivo models of muscle wasting, that cachectic factors are remarkably selective in targeting myosin heavy chain. In myotubes and mouse muscles, TNF alpha plus IFN-gamma strongly reduced myosin expression through an RNA-dependent mechanism. Likewise, colon-26 tumors in mice caused the selective reduction of this myofibrillar protein, and this reduction correlated with wasting. Under these conditions, however, loss of myosin was associated with the ubiquitin dependent proteasome pathway, which suggests that mechanisms used to regulate the expression of muscle proteins may be cachectic factor specific. These results shed new light on cancer cachexia by revealing that wasting does not result from a general downregulation of muscle proteins but rather is highly selective as to which proteins are targeted during the wasting state. PMID- 15286804 TI - Novel mode of action of c-kit tyrosine kinase inhibitors leading to NK cell dependent antitumor effects. AB - Mutant isoforms of the KIT or PDGF receptors expressed by gastrointestinal stromal tumors (GISTs) are considered the therapeutic targets for STI571 (imatinib mesylate; Gleevec), a specific inhibitor of these tyrosine kinase receptors. Case reports of clinical efficacy of Gleevec in GISTs lacking the typical receptor mutations prompted a search for an alternate mode of action. Here we show that Gleevec can act on host DCs to promote NK cell activation. DC mediated NK cell activation was triggered in vitro and in vivo by treatment of DCs with Gleevec as well as by a loss-of-function mutation of KIT. Therefore, tumors that are refractory to the antiproliferative effects of Gleevec in vitro responded to Gleevec in vivo in an NK cell-dependent manner. Longitudinal studies of Gleevec-treated GIST patients revealed a therapy-induced increase in IFN-gamma production by NK cells, correlating with an enhanced antitumor response. These data point to a novel mode of antitumor action for Gleevec. PMID- 15286805 TI - Expanded B cell population blocks regulatory T cells and exacerbates ileitis in a murine model of Crohn disease. AB - SAMP1/YitFc mice develop discontinuous, transmural inflammatory lesions in the terminal ileum, similar to what is found in human Crohn disease. Compared with the mesenteric lymph nodes (MLNs) of AKR control mice, SAMP1/YitFc MLNs contain a 4.3-fold expansion in total B cell number and a 2.5-fold increased percentage of CD4(+) T cells expressing the alpha(E)beta(7) integrin. Although alpha(E)beta(7)(+)CD4(+) T cells possess a regulatory phenotype (CD25(+), L selectin(lo), and CD45RB(lo)), express IL-10, and suppress effector T cell proliferation in vitro, they cannot prevent ileitis development in SCID mice adoptively transferred with effector CD4(+) T cells, although the CD4(+)CD25(+) subset, which overlaps with the alpha(E)beta(7)(+)CD4(+) subset, prevents colitis. The alpha(E)beta(7)(+)CD4(+) T cells express high levels of ICOS, a costimulatory molecule that augments B cell function, suggesting their involvement in the increase in B cells, IgA(+) cells, and soluble IgA found within the MLNs and ileum of SAMP1/YitFc mice. MLN B cell numbers correlate with ileitis severity in SAMP1/YitFc mice, and cotransfer of SAMP1/YitFc MLN B cells along with CD4(+) T cells increases ileitis severity in SCID mice compared with transfer of CD4(+) T cells alone. SAMP1/YitFc B cells prevent alpha(E)beta(7)(+)CD4(+) T cells from suppressing effector T cell proliferation. We conclude that SAMP1/YitFc MLN B cells contribute to the development of SAMP1/YitFc ileitis. PMID- 15286806 TI - The anaphylatoxin C3a downregulates the Th2 response to epicutaneously introduced antigen. AB - Mechanical injury to the skin results in activation of the complement component C3 and release of the anaphylatoxin C3a. C3a binds to a seven-transmembrane G protein-coupled receptor, C3aR. We used C3aR(-/-) mice to examine the role of C3a in a mouse model of allergic inflammation induced by epicutaneous sensitization with OVA. C3aR(-/-) mice exhibited an exaggerated Th2 response to epicutaneous but not to intraperitoneal sensitization with OVA, as evidenced by significantly elevated levels of serum OVA-specific IgG1 and significantly increased secretion of the Th2 cytokines IL-4, IL-5, and IL-10 by antigen-stimulated splenocytes. Presentation of OVA peptide by C3aR(-/-) APCs caused significantly more IL-4 and IL-5 secretion by T cells from OVA-T cell receptor (OVA-TCR) transgenic mice compared with presentation by WT APCs. C3a inhibited the ability of splenocytes, but not of highly purified T cells, to secrete Th2 cytokines in response to TCR ligation. This inhibition was mediated by IL-12 secreted by APCs in response to C3a. These results suggest that C3a-C3aR interactions inhibit the ability of APCs to drive Th2 cell differentiation in response to epicutaneously introduced antigen and may have important implications for allergic skin diseases. PMID- 15286807 TI - Integrin engagement regulates monocyte differentiation through the forkhead transcription factor Foxp1. AB - The precise signals responsible for differentiation of blood-borne monocytes into tissue macrophages are incompletely defined. "Outside-in" signaling by integrins has been implicated in modulation of gene expression that affects cellular differentiation. Herein, using differential display PCR, we have cloned an 85-kDa forkhead transcription factor (termed Mac-1-regulated forkhead [MFH] and found subsequently to be identical to Foxp1) that is downregulated in beta(2)-integrin Mac-1-clustered compared with Mac-1-nonclustered monocytic THP-1 cells. MFH/Foxp1 is expressed in untreated HL60 cells, and its expression was markedly reduced during phorbol ester-induced monocyte differentiation, but not retinoic acid induced granulocyte differentiation. Overexpression of MFH/Foxp1 markedly attenuated phorbol ester-induced expression of c-fms, which encodes the M-CSF receptor and is obligatory for macrophage differentiation. This was accompanied by decreased CD11b expression, cell adhesiveness, and phagocytosis. Using electromobility shift and reporter assays, we have established that MFH/Foxp1 binds to previously uncharacterized sites within the c-fms promoter and functions as a transcriptional repressor. Deficiency of Mac-1 is associated with altered regulation of MFH/Foxp1 and monocyte maturation in vivo. Taken together, these observations suggest that Mac-1 engagement orchestrates monocyte-differentiation signals by regulating the expression of the forkhead transcription repressor MFH/Foxp1. This represents a new pathway for integrin-dependent modulation of gene expression and control of cellular differentiation. PMID- 15286808 TI - Bone marrow-derived immune cells regulate vascular disease through a p27(Kip1) dependent mechanism. AB - The cyclin-dependent kinase inhibitors are key regulators of cell cycle progression. Although implicated in carcinogenesis, they inhibit the proliferation of a variety of normal cell types, and their role in diverse human diseases is not fully understood. Here, we report that p27(Kip1) plays a major role in cardiovascular disease through its effects on the proliferation of bone marrow-derived (BM-derived) immune cells that migrate into vascular lesions. Lesion formation after mechanical arterial injury was markedly increased in mice with homozygous deletion of p27(Kip1), characterized by prominent vascular infiltration by immune and inflammatory cells. Vascular occlusion was substantially increased when BM-derived cells from p27(-/-) mice repopulated vascular lesions induced by mechanical injury in p27(+/+) recipients, in contrast to p27(+/+) BM donors. To determine the contribution of immune cells to vascular injury, transplantation was performed with BM derived from RAG(-/-) and RAG(+/+) mice. RAG(+/+) BM markedly exacerbated vascular proliferative lesions compared with what was found in RAG(-/-) donors. Taken together, these findings suggest that vascular repair and regeneration is regulated by the proliferation of BM derived hematopoietic and nonhematopoietic cells through a p27(Kip1)-dependent mechanism and that immune cells largely mediate these effects. PMID- 15286809 TI - IL-5 links adaptive and natural immunity specific for epitopes of oxidized LDL and protects from atherosclerosis. AB - During atherogenesis, LDL is oxidized, generating various oxidation-specific neoepitopes, such as malondialdehyde-modified (MDA-modified) LDL (MDA-LDL) or the phosphorylcholine (PC) headgroup of oxidized phospholipids (OxPLs). These epitopes are recognized by both adaptive T cell-dependent (TD) and innate T cell independent type 2 (TI-2) immune responses. We previously showed that immunization of mice with MDA-LDL induces a TD response and atheroprotection. In addition, a PC-based immunization strategy that leads to a TI-2 expansion of innate B-1 cells and secretion of T15/EO6 clonotype natural IgM antibodies, which bind the PC of OxPLs within oxidized LDL (OxLDL), also reduces atherogenesis. T15/EO6 antibodies inhibit OxLDL uptake by macrophages. We now report that immunization with MDA-LDL, which does not contain OxPL, unexpectedly led to the expansion of T15/EO6 antibodies. MDA-LDL immunization caused a preferential expansion of MDA-LDL-specific Th2 cells that prominently secreted IL-5. In turn, IL-5 provided noncognate stimulation to innate B-1 cells, leading to increased secretion of T15/EO6 IgM. Using a bone marrow transplant model, we also demonstrated that IL-5 deficiency led to decreased titers of T15/EO6 and accelerated atherosclerosis. Thus, IL-5 links adaptive and natural immunity specific to epitopes of OxLDL and protects from atherosclerosis, in part by stimulating the expansion of atheroprotective natural IgM specific for OxLDL. PMID- 15286811 TI - The influence of the human genome on chronic viral hepatitis outcome. AB - The mechanisms that determine viral clearance or viral persistence in chronic viral hepatitis have yet to be identified. Recent advances in molecular genetics have permitted the detection of variations in immune response, often associated with polymorphism in the human genome. Differences in host susceptibility to infectious disease and disease severity cannot be attributed solely to the virulence of microbial agents. Several recent advances concerning the influence of human genes in chronic viral hepatitis B and C are discussed in this article: a) the associations between human leukocyte antigen polymorphism and viral hepatic disease susceptibility or resistance; b) protective alleles influencing hepatitis B virus (HBV) and hepatitis C virus (HCV) evolution; c) prejudicial alleles influencing HBV and HCV; d) candidate genes associated with HBV and HCV evolution; d) other genetic factors that may contribute to chronic hepatitis C evolution (genes influencing hepatic stellate cells, TGF-beta 1 and TNF-alpha production, hepatic iron deposits and angiotensin II production, among others). Recent discoveries regarding genetic associations with chronic viral hepatitis may provide clues to understanding the development of end-stage complications such as cirrhosis or hepatocellular carcinoma. In the near future, analysis of the human genome will allow the elucidation of both the natural course of viral hepatitis and its response to therapy. PMID- 15286810 TI - Circulating fibrocytes traffic to the lungs in response to CXCL12 and mediate fibrosis. AB - Previous reports have identified a circulating pool of CD45(+) collagen I(+) CXCR4(+) (CD45(+)Col I(+)CXCR4(+)) cells, termed fibrocytes, that traffic to areas of fibrosis. No studies have demonstrated that these cells actually contribute to fibrosis, however. Pulmonary fibrosis was originally thought to be mediated solely by resident lung fibroblasts. Here we show that a population of human CD45(+)Col I(+)CXCR4(+) circulating fibrocytes migrates in response to CXCL12 and traffics to the lungs in a murine model of bleomycin-induced pulmonary fibrosis. Next, we demonstrated that murine CD45(+)Col I(+)CXCR4(+) fibrocytes also traffic to the lungs in response to a bleomycin challenge. Maximal intrapulmonary recruitment of CD45(+)Col I(+)CXCR4(+) fibrocytes directly correlated with increased collagen deposition in the lungs. Treatment of bleomycin-exposed animals with specific neutralizing anti-CXCL12 Ab's inhibited intrapulmonary recruitment of CD45(+)Col I(+)CXCR4(+) circulating fibrocytes and attenuated lung fibrosis. Thus, our results demonstrate, we believe for the first time, that circulating fibrocytes contribute to the pathogenesis of pulmonary fibrosis. PMID- 15286812 TI - Paracoccidioidomycosis in children: clinical presentation, follow-up and outcome. AB - From February, 1981 to May, 2001, 63 children under 15 y old (ages 2 - 15 y, median = 8 y, mean +/- 1 SD = 8 +/- 3 y) presenting 70 episodes of Paracoccidioidomycosis were admitted. The main clinical manifestations and laboratory features observed upon admission were: lymph node enlargement (87.1%), fever (75.7%), weakness (48.6%), pallor (41.4%), hepatomegaly (40%), splenomegaly (35.7%), anemia (90%), hypergammaglobulinemia (88.5%), eosinophilia (75.5%) and hypoalbuminemia (72.5%). Moderate to severe malnutrition was detected in 35.7% of the episodes (Gomez's criterion). Radiographic and technetium studies showed bone lesions in 20 of the episodes, most of them being multiple lytic lesions, involving both long (70%) and plain bones (30%). First line treatment consisted of an association of sulfametoxazole-trimethoprin, which was used, exclusively, in 50 episodes. Follow-up of hemoglobin levels, number of eosinophils in the peripheral blood, albumin and gammaglobulin serum levels revealed significant sequential improvement one and six months after hospital admission, being quite useful to evaluate treatment effectiveness. Six patients died (9.3%) and four developed sequelae (6.3%). In conclusion, the juvenile and disseminated forms can be observed in about 70% of the episodes of PCM occurring in children younger than 15 y old, most of them presenting with a febrile lymphoproliferative syndrome associated to anemia, eosinophilia and hypergammaglobulinemia. PMID- 15286814 TI - Airborne fungi isolated from Fortaleza city, State of Ceara, Brazil. AB - Airbone fungi are considered important causes of allergic rhinitis and allergic asthma. The knowledge of these fungi in a city or region is important for the ecological diagnosis and specific treatment of allergic manifestations induced by inhalation of fungal allergens. The airborne fungi of Fortaleza, State of Ceara, Brazil, were studied during a one year period. Five hundred and twenty Petri dishes with Sabouraud dextrose agar medium were exposed at ten different locations in the city. The dishes exposed yielded one thousand and five hundred and twenty one colonies of twenty four genera. The most predominants were: Aspergillus (44.7%), Penicillium (13.3%), Curvularia (9.8%), Cladosporium (6.8%), Mycelia sterilia (6.0%), Fusarium (3.5%), Rhizopus (3.1%), Drechslera (2.6%), Alternaria (2.4%) and Absidia (2.2%). The results shown that Aspergillus, Penicillium, Mycelia sterilia, Fusarium and Alternaria were found during all months in the year. Absidia was more frequent during the dry season. Anemophilous fungi and the high concentration of spores in the air are important because may result in an increased number of people with allergic respiratory disease. PMID- 15286816 TI - Human herpesvirus-7 as a cause of exanthematous ilnesses in Belem, Para, Brazil. AB - We screened sera from 370 patients suffering from exanthematous illnesses in Belem, North Brazil, for the presence of human herpesvirus-7 (HHV-7) IgM and IgG antibodies. Samples were obtained from January 1996 to December 2002 and were processed by a HHV-7-specific indirect immunofluorescence assay (IFA). HHV-7 specific IgM and/or IgG antibodies were found in 190 (51.4%) of these patients, with similar prevalence rates (IgM+ and IgG+ subgroups taken together) for female and male subjects: 52.5% and 50.3%, respectively. Serological status as defined by IgG was identified in 135 (36.5%) patients. In 55 (14.9%) of the patients HHV 7 IgM antibodies were detected. HHV-7 IgM- and- IgG antibody rates were similar (p > 0.05) when male and female subjects are compared: 14.4% versus 15.3% and 38.1% versus 35.0%, respectively. Statistically significant difference (p = 0.003) was noted when HHV-7-IgM-positive female and male patients aged 5-8 months are compared. Prevalence rates ranging from 4.6% (female, 5-8 months of age) to 93.3% (female, > 10 years of age) and 12.2% (male, 5-8 months) to 80.0% (male, 8 10 years of age) were noted in the IgG- positive subgroups. A subgroup (n = 131) of patients with IgM or IgG HHV-7 antibodies were examined for the presence of DNA using a polymerase chain reaction/nested PCR. Recent/active HHV-7 infection occurred at a rate of 11.0% (6/55) among patients whose samples presented IgM+ specific antibodies. In a subgroup (n = 76) of patients with high HHV-7-IgG antibody levels (titre > 1:160) DNA could not be detected in sera examined by PCR/nested PCR. Of the six recent/active infections, four subjects with less than 1 year and two with 3 and 6 years of age, presented typical exanthem subitum (E.S), as defined by higher fever (> 38.0 degrees C) with duration of 24 to 72 hours, followed by a maculopapular skin rash. Our results underscore the need for searching HHV-7 infection in patients with exanthematous diseases, particularly those presenting with typical E.S. HHV-7 appears therefore to emerge as a newly recognized pathogen of exanthem in our region. PMID- 15286818 TI - Molecular analysis of the dengue virus type 1 and 2 in Brazil based on sequences of the genomic envelope-nonstructural protein 1 junction region. AB - The genomic sequences of the Envelope-Non-Structural protein 1 junction region (E/NS1) of 84 DEN-1 and 22 DEN-2 isolates from Brazil were determined. Most of these strains were isolated in the period from 1995 to 2001 in endemic and regions of recent dengue transmission in Sao Paulo State. Sequence data for DEN-1 and DEN-2 utilized in phylogenetic and split decomposition analyses also include sequences deposited in GenBank from different regions of Brazil and of the world. Phylogenetic analyses were done using both maximum likelihood and Bayesian approaches. Results for both DEN-1 and DEN-2 data are ambiguous, and support for most tree bipartitions are generally poor, suggesting that E/NS1 region does not contain enough information for recovering phylogenetic relationships among DEN-1 and DEN-2 sequences used in this study. The network graph generated in the split decomposition analysis of DEN-1 does not show evidence of grouping sequences according to country, region and clades. While the network for DEN-2 also shows ambiguities among DEN-2 sequences, it suggests that Brazilian sequences may belong to distinct subtypes of genotype III. PMID- 15286819 TI - Echinococcosis in southern Brazil: efforts toward implementation of a control program in Santana do Livramento, Rio Grande do Sul. AB - This investigation aimed to design a strategy for echinococcosis control in Santana do Livramento county, an endemic area in state of Rio Grande do Sul (Brazil). Fecal samples from 65 dogs were obtained from urban, suburban and rural areas. Purging with Arecoline Bromhidrate (AB) was done to visualize Echinococcus granulosus, and Enzyme Linked Immunosorbent Assay (ELISA) was performed to detect parasite coproantigen. Samples were obtained at the beginning and at the end of treatment with Praziquantel. A third fecal sampling was also done in rural areas four months after the end of treatment. Each dog was treated immediately after the first purging and every 30 days for eight months. In urban and suburban areas no infected dogs were found. In rural areas, first evaluation showed 11.36% and 27.69% of infected dogs by AB and ELISA, respectively. No infected dogs were diagnosed in the second evaluation and in the third evaluation 36.84% and 47.37% infected dogs were identified by AB and ELISA, respectively. Medication program to combat dog infection resulted in successful interruption of parasite transmission, but the project failed to create awareness of the need for dog prophylaxis among rural populations as well as to establish a permanent control program in this municipality. PMID- 15286820 TI - Comparative study of the fecundity and fertility of Biomphalaria glabrata (Say, 1818) and Biomphalaria straminea (Dunker, 1848) in a laboratory through self fertilization and cross-fertilization. AB - The aim of this study was to compare the fecundity and fertility of B. glabrata and B. straminea by cross- and self-fertilization. To attain this objective, laboratory-raised strains of B. glabrata and B. straminea were used. The former originated from natural breeding grounds in the municipality Paulista, state of Pernambuco, Brazil. The latter originated from irrigation ditches in the municipality of Petrolandia, in the same state. Snail populations of B. glabrata and B. straminea were maintained for 240 days in laboratory. Their fecundity was evaluated by noting the number of egg-masses, eggs and eggs per mass. Their fertility was evaluated by the number of viable eggs and the hatching rate. B. straminea was markedly more fecund than B. glabrata through cross- and self fertilization, namely: greater egg-mass; higher egg production and more eggs per mass. Regarding fertility, there seemed to be no preferential period for occlusion to occur or a trend in the rhythm of producing viable eggs. PMID- 15286822 TI - A simple and cheaper in house varicella zoster virus antibody indirect ELISA. AB - We have developed a cheaper an simple in house indirect ELISA that uses the live attenuated VZV vaccine as a coating antigen. The alternative ELISA had an agreement of 94% when compared with a commercial VZV ELISA kit. Moreover, our ELISA proved to be more reliable than the kit when assessing true negative samples. By adding a standard serum, we were able to produce results in international units per millilitre. Also, the addition of an extra step with 8M urea allowed the assessment of VZV IgG avidity without excessive costs. The cost per sample to test VZV IgG was 2.7 times cheaper with our ELISA, allowing the testing of many samples without the burden of production of VZV antigen in the laboratory. PMID- 15286823 TI - Salmonella enterica subsp houtenae serogroup O:16 in a HIV positive patient: case report. AB - We described a case of salmonellosis in a 33-year old HIV-infected patient. The patient presented oral and esophageal candidiasis, intense epigastric and retrosternal pain. During the physical examination he was hypochloraemic, acyanotic, hypohydrated, anicteric and afebrile. Admittance laboratory tests indicated: red cells 3.6 millions/mm(3); hemoglobin, 10.1 g/dL; leukocyte count, 3,000/mm(3), with 1% of eosinophils, 14% of non-segmented and 53% of segmented neutrophils and 31% of lymphocytes. The blood culture was positive for Salmonella enterica subsp houtenae serogroup O:16. This is probably the first human report of bacteremia due to Salmonella enterica subsp houtenae in Brazil associated to HIV-infected patient. PMID- 15286824 TI - Carrion's disease (Bartonellosis bacilliformis) confirmed by histopathology in the High Forest of Peru. AB - Bartonellosis or Carrion's disease is endemic in some regions of Peru, classically found in the inter-Andean valleys located between 500 and 3200 meters above sea level. We report the case of a 43 year-old male patient, farmer, who was born in the Pichanaki district (Chanchamayo, Junin), located in the High Forest of Peru. He presented with disseminated, raised, erythematous cutaneous lesions, some of which bled. The distribution of these lesions included the nasal mucosa and penile region. Additionally subcutaneous nodules were distributed over the trunk and extremities. Hematologic exams showed a moderate anemia. Serologic studies for HIV and Treponema pallidum were negative. The histopathologic results of two biopsies were compatible with Peruvian wart. Oral treatment with ciprofloxacin (500 mg BID) was begun. Over 10 days, the patient showed clinical improvement. This is the first report of a confirmed case of bartonellosis in the eruptive phase originating from the Peruvian High Forest, showing the geographical expansion of the Carrion's disease. PMID- 15286825 TI - Azithromycin in the treatment of mucosal leishmaniasis. AB - This report describes three elderly patients with mucosal form of American tegumentary leishmaniasis associated with chronic cardiopathy. Due to the known toxicity of classical drugs with activity against Leishmania sp., the patients received three oral courses of azithromycin therapy in single 500 mg daily dose during ten days, every other month. All lesions healed after the third series. One of the patients relapsed and a new series of azithromycin was prescribed. Azithromycin may be an alternative drug for the treatment of leishmaniasis in special situations due to its optimal mucosal and intraphagocyte concentration, single daily posology, high tolerance and oral administration. The mechanism of this drug on Leishmania sp. is unknown at present. PMID- 15286827 TI - Dental and facial characteristics of patients with juvenile idiopathic arthritis. AB - OBJECTIVE: It has been shown that the temporomandibular joint is frequently affected by juvenile idiopathic arthritis, and this degenerative disease, which may occur during facial growth, results in severe mandibular dysfunction. However, there are no studies that correlate oral health (tooth decay and gingival diseases) and temporomandibular joint dysfunction in patients with juvenile idiopathic arthritis. The aim of this study is to evaluate the oral and facial characteristics of the patients with juvenile idiopathic arthritis treated in a large teaching hospital. METHOD: Thirty-six patients with juvenile idiopathic arthritis (26 female and 10 male) underwent a systematic clinical evaluation of their dental, oral, and facial structures (DMFT index, plaque and gingival bleeding index, dental relationship, facial profile, and Helkimo's index). The control group was composed of 13 healthy children. RESULTS: The mean age of the patients with juvenile idiopathic arthritis was 10.8 years; convex facial profile was present in 12 juvenile idiopathic arthritis patients, and class II molar relation was present in 12 (P =.032). The indexes of plaque and gingival bleeding were significant in juvenile idiopathic arthritis patients with a higher number of superior limbs joints involved (P =.055). Anterior open bite (5) and temporomandibular joint noise (8) were present in the juvenile idiopathic arthritis group. Of the group in this sample, 94% (P =.017) had temporomandibular joint dysfunction, 80% had decreased mandibular opening (P = 0.0002), and mandibular mobility was severely impaired in 33% (P =.015). CONCLUSION: This study confirms that patients with juvenile idiopathic arthritis a) have a high incidence of mandibular dysfunction that can be attributed to the direct effect of the disease in the temporomandibular joint and b) have a higher incidence of gingival disease that can be considered a secondary effect of juvenile idiopathic arthritis on oral health. PMID- 15286828 TI - Effect of clarithromycin on the cell profile of bronchoalveolar lavage fluid in mice with neutrophil-predominant lung disease. AB - OBJECTIVE: Macrolide antibiotics have anti-inflammatory properties in lung diseases. The aim of this study was to investigate the effect of clarithromycin in pulmonary cellular inflammatory response in mice. METHOD: Eight adult Swiss mice were studied. All animals received an intranasal challenge (80 micro L) with dead Pseudomonas aeruginosa (1.0 x 10(12) CFU/mL). Bronchoalveolar lavage was performed 2 days later, with total cell count and differential cell analysis. The study group (n = 4) received clarithromycin treatment (50 mg/kg/day, intraperitoneal) for 5 days. Treatment was initiated 2 days before intranasal challenge. RESULTS: There was no significant difference in total cell count between the groups (mean: 2.0 x 10(6) and 1.3 x 10(6), respectively). In both groups, there was a predominance of neutrophils. However, the study group had a higher percentage of lymphocytes in the bronchoalveolar lavage than the control group (median of 19% vs 2.5%, P =.029). CONCLUSION: Clarithromycin alters the cytological pattern of bronchoalveolar lavage of Swiss mice with neutrophil pulmonary inflammation, significantly increasing the percentage of lymphocytes. PMID- 15286829 TI - Different doses of exogenous surfactant for treatment of meconium aspiration syndrome in newborn rabbits. AB - OBJECTIVE: To evaluate the effects of 2 different doses of exogenous surfactant on pulmonary mechanics and on the regularity of pulmonary parenchyma inflation in newborn rabbits. METHOD: Newborn rabbits were submitted to tracheostomy and randomized into 4 study groups: the Control group did not receive any material inside the trachea; the MEC group was instilled with meconium, without surfactant treatment; the S100 and S200 groups were instilled with meconium and were treated with 100 and 200 mg/kg of exogenous surfactant (produced by Instituto Butantan) respectively. Animals from the 4 groups were mechanically ventilated during a 25 minute period. Dynamic compliance, ventilatory pressure, tidal volume, and maximum lung volume (P-V curve) were evaluated. Histological analysis was conducted using the mean linear intercept (Lm), and the lung tissue distortion index (SDI) was derived from the standard deviation of the means of the Lm. One way analysis of variance was used with a = 0.05. RESULTS: After 25 minutes of ventilation, dynamic compliance (mL/cm H2O.kg) was 0.87 +/- 0.07 (Control); 0.49 +/- 0.04 (MEC*); 0.67 +/- 0.06 (S100); and 0.67 +/- 0.08 (S200), and ventilatory pressure (cm H2O) was 9.0 +/- 0.9 (Control); 16.5 +/- 1.7 (MEC*); 12.4 +/- 1.1 (S100); and 12.1 +/- 1.5 (S200). Both treated groups had lower Lm values and more homogeneity in the lung parenchyma compared to the MEC group: SDI = 7.5 +/- 1.9 (Control); 11.3 +/- 2.5 (MEC*), 5.8 +/- 1.9 (S100); and 6.7 +/- 1.7 (S200) (*P < 0.05 versus all the other groups). CONCLUSIONS: Animals treated with surfactant showed significant improvement in pulmonary mechanics and more regularity of the lung parenchyma in comparison to untreated animals. There was no difference in results after treatment with either of the doses used. PMID- 15286831 TI - Nonsmall cell lung cancer: evaluation of 737 consecutive patients in a single institution. AB - OBJECTIVE: To analyze surgical and pathological parameters and outcome and prognostic factors of patients with nonsmall cell lung cancer (NSCLC) who were admitted to a single institution, as well as to correlate these findings to the current staging system. METHOD: Seven hundred and thirty seven patients were diagnosed with NSCLC and admitted to Hospital do Cancer A. C. Camargo from 1990 to 2000. All patients were included in a continuous prospective database, and their data was analyzed. Following staging, a multidisciplinary team decision on adequate management was established. Variables included in this analysis were age, gender, histology, Karnofsky index, weight loss, clinical stage, surgical stage, chemotherapy, radiotherapy, and survival rates. RESULTS: 75.5% of patients were males. The distribution of histologic type was squamous cell carcinoma 51.8%, adenocarcinoma 43.1%, and undifferentiated large cell carcinoma 5.1%. Most patients (73%) presented significant weight loss and a Karnofsky index of 80%. Clinical staging was IA 3.8%, IB 9.2%, IIA 1.4%, IIB 8.1%, IIIA 20.9%, IIIB 22.4%, IV 30.9%. Complete tumor resection was performed in 24.6% of all patients. Surgical stage distribution was IA 25.3%, IB 1.4%, IIB 17.1%, IIIA 16.1%, IIIB 20.3%, IV 11.5%. Chemotherapy and radiotherapy were considered therapeutic options in 43% and 72%, respectively. The overall 5-year survival rate of nonsmall cell lung cancer patients in our study was 28%. Median survival was 18.9 months. CONCLUSIONS: Patients with NSCLC who were admitted to our institution presented with histopathologic and clinical characteristics that were similar to previously published series in cancer hospitals. The best prognosis was associated with complete tumor resection with lymph node dissection, which is only achievable in earlier clinical stages. PMID- 15286830 TI - Systemic hypertension, diabetes mellitus, and dyslipidemia in relation to body mass index: evaluation of a Brazilian population. AB - OBJECTIVE: To determine the prevalence of systemic hypertension, diabetes mellitus, hypercholesterolemia, and hypertriglyceridemia in a Brazilian population in relation to body mass index. METHOD: Retrospective evaluation of 1213 adults (mean age: 45.2 +/- 12.8; 80.6% females) divided into groups according to body mass index [normal (18.5 - 24.4 kg/m2); overweight (25 - 29.9 kg/m2); grade 1 obesity (30 - 34.9 kg/m2); grade 2 obesity (35 - 39.9 kg/m2), and grade 3 obesity (> or = 40 kg/m2)]. The prevalence of hypertension, diabetes mellitus, hypercholesterolemia, and hypertriglyceridemia were analyzed in each group. The severity of cardiovascular risk was determined. High-risk patients were considered those reporting 2 or more of the following factors: systemic hypertension, HDL < or = 35 mg/dL, total cholesterol > or = 240 mg/dL, triglycerides > or = 200 mg/dL when HDL < or = 35 mg/dL, and glycemia > or = 126 mg/dL. Moderate-risk patients were those reporting 2 or more of the following factors: systemic hypertension, HDL < or = 45, triglycerides > or = 200 mg/dL, and total cholesterol > or = 200 mg/dL. RESULTS: The prevalence of systemic hypertension, diabetes mellitus, hypertriglyceridemia, and low HDL-cholesterol levels increased along with weight, but the prevalence of hypercholesterolemia did not. The odds ratio adjusted for gender and age, according to grade of obesity compared with patients with normal weight were respectively 5.9, 8.6, and 14.8 for systemic hypertension, 3.8, 5.8, and 9.2 for diabetes mellitus and 1.2, 1.3, and 2.6 for hypertriglyceridemia. We also verified that body mass index was positively related to cardiovascular high risk (P < .001) CONCLUSION: In our population, cardiovascular risk increased along with body mass index. PMID- 15286832 TI - Adnexal torsion following gonadotropin-releasing hormone analog therapy: a case report. AB - Adnexal torsion may occur in girls and adolescents. Often it is associated with ovarian diseases resulting in ovarian enlargement. Adnexal torsion may involve the ovary, fallopian tube or both, and the main symptom is acute pelvic pain. An 8-year-old girl complaining of acute pelvic and abdominal pain, who was previously diagnosed with precocious puberty and who received treatment with a GnRH analog, is reported. Ultrasound demonstrated a normal-sized uterus and bilaterally enlarged ovaries with multiple internal cysts. At laparotomy, we found a complete torsion in the right adnexa. The histological examination revealed massive edema associated with multiple antral follicles and reduction of the follicular reserve. PMID- 15286833 TI - Concomitant involvement of the small intestine and the distal esophagus in an infant with massive necrotizing enterocolitis. AB - Necrotizing enterocolitis is a disease of the newborn that may involve the small intestine and/or the colon, and the stomach. To our knowledge, massive necrosis of the small intestine with concomitant involvement of the esophagus has never been reported. A case of a 6-month-old boy with necrotizing enterocolitis and pan necrosis of the small intestine, cecum, and the lower third of the esophagus is presented. After 70 days of treatment, intestinal transit was established by an anastomosis between the first centimeter of jejunum and the ascending colon. Finally, esophageal transit was established by a total gastric transposition with cervical esophagogastric anastomosis. The patient was maintained under total parenteral nutrition, and after 19 months he developed fulminant hepatic failure due to parenteral nutrition; he then underwent combined liver and small bowel transplantation. After 2 months, the patient died due to undefined neurologic complications, probably related to infection or immunosuppressive therapy. PMID- 15286834 TI - Conjunctival keratoacanthoma. AB - Keratoacanthoma generally occurs on the skin; it is rarely found in the conjunctiva. A case of a 34-year-old woman with a rapidly growing conjunctival mass is reported. The tumor was excised with a safety margin to exclude squamous cell carcinoma. Histopathologically it was crateriform and consistent with atypical keratoacanthoma. There has been no recurrence in 2 years of follow-up. Conjunctival keratoacanthoma is rare; differential diagnosis of conventional squamous cell carcinoma and keratoacanthoma can be difficult. We recommend complete surgical excision and careful follow-up of crateriform squamous proliferations. PMID- 15286835 TI - Involvement of C4 allotypes in the pathogenesis of human diseases. AB - The complement system is an important humoral defense mechanism that plays a relevant role against microbial agents, inflammatory response control, and immunocomplex clearance. Classical complement pathway activation is antibody dependent. The C4 component participates in the initial step of activation, and C4 expression is determined by 2 pairs of allotypes: C4A and C4B. Deficiencies in C4 allotypes have been associated with several diseases. The aim of the present review is evaluate the reported data in the literature regarding specific C4A and C4B deficiencies and characterize their clinical relevance. We searched the MEDLINE and LILACS databases. Papers referring to total C4 deficiency without allotype evaluation and case reports of primary C4 deficiency were not included. Deficiencies in C4 allotypes have been associated with Mycobacterium leprae infection, erythema nodosum, systemic sclerosis with anti-topoisomerase I antibodies, intermediate congenital adrenal hyperplasia with DR5 genotype, diabetes mellitus type 1 with DR3,4 genotype, and diabetes mellitus with antibodies against islet cells. C4 allotype deficiency is also related to C4B deficiency and autoimmune-associated diseases, such as systemic lupus erythematosus, or diseases with an autoimmune component, such as autism. Some reports associate C4A with thyroiditis after delivery as well as limited and systemic sclerosis without anti-topoisomerase I antibodies. However, the studies with C4A and C4B have been concentrated in isolated populations, and some of the studies could not be reproduced by other authors. PMID- 15286836 TI - Proton magnetic resonance spectroscopy of the frontal lobe in schizophrenics: a critical review of the methodology. AB - Schizophrenic patients undergoing proton magnetic resonance spectroscopy show alterations in N-acetyl aspartate levels in several brain regions, indicating neuronal dysfunction. The present review focuses on the main proton magnetic resonance spectroscopy studies in the frontal lobe of schizophrenics. A MEDLINE search, from 1991 to March 2004, was carried out using the key-words spectroscopy and schizophrenia and proton and frontal. In addition, articles cited in the reference list of the studies obtained through MEDLINE were included. As a result, 27 articles were selected. The results were inconsistent, 19 papers reporting changes in the N-acetyl aspartate levels, while 8 reported no change. Methodological analysis led to the conclusion that the discrepancy may be due the following factors: (i) number of participants; (ii) variation in the clinical and demographic characteristics of the groups; (iii) little standardization of the acquisition parameters of spectroscopy. Overall, studies that fulfill strict methodological criteria show N-acetyl aspartate decrease in the frontal lobe of male schizophrenics. PMID- 15286837 TI - Homeopathy: do not condemn what we do not understand. PMID- 15286840 TI - [Adverse events -- or is it normal?]. PMID- 15286839 TI - [The importance of pharmaceutical medicine]. PMID- 15286841 TI - [Incidental finding of hepatic nodule. What is its importance?]. PMID- 15286842 TI - [Second-hour hemo-sedimentation velocity: What is its usefulness]. PMID- 15286844 TI - [Diagnosis of hepatic encephalopathy]. PMID- 15286843 TI - [What is the importance of video-assisted hysteroscopy in the climacteric?]. PMID- 15286845 TI - [Which are the alterations in the fetal heart rate during the intrapartum of greater prediction of neonatal acidemia?]. PMID- 15286846 TI - [Infectious endocarditis]. PMID- 15286853 TI - [Milk of calcium]. PMID- 15286854 TI - [Cryptococcal neoformans meningitis as a cause of prolonged fever in a patient with AIDS ]. PMID- 15286855 TI - [Clinical and anatomopathologic diagnosis: discordances]. PMID- 15286856 TI - [Airborne fungi]. PMID- 15286857 TI - [Evaluation of scientific papers]. PMID- 15286858 TI - [Muscular energetic metabolism study in athletes using 31P-MRS]. AB - BACKGROUND: The aim of this study was to characterize the muscular reservoirs of phosphorilated energetic components of athletes using 31P-MRS. METHODS: The sample was formed by 14 elite athletes from the Center for High Sportive Performance (CAR, Sant Cugat del Valles, Spain). The pattern of the phosphorilated metabolites was measured from the muscle vastus medialis by 31P MRS. Oral supplementation of 20 g of Creatine monohydrate was given during 14 days. Two groups of athletes were formed according to their physical characteristics (weight, height, body mass index, maximum O2 uptake). The first group received a placebo (maltodextrine), while the second group received a diet of creatine supplement. The exercise was performed inside the resonance tunnel with a frequency of 60 RPM with both legs. RESULTS: The results showed that significant decrease occurred in phosphocreatine (PCr), inorganic phosphate (Pi) and intracellular pH after supplementation. CONCLUSION: It was concluded that the exercise performed by the long distance runners recruited in this study, detected by 31P-MRS, reduced the consumption of PCr during exercise owing to creatine supplementation diet. PMID- 15286859 TI - [Acquisition of skills in medical ethics in learning-teaching small groups. Comparing problem-based learning with the traditional model]. AB - BACKGROUND: Aiming to evaluate the acquisition of skills on Medical Ethics among medical students from Marilia Medical School, some of them from the small group learning-teaching method, others from traditional teaching method. METHODS: A prospective analytical study was done based on the application of questionnaires about general themes on Ethics, at two different times. RESULTS: There weren't significant differences on the skills' acquisition between the two methods. Students from late graduation years showed a significantly better performance than those from early years. The themes that presented worse results were medical secret, legal responsible consent, patient autonomy, medical prescription, medical handbook and corporative feeling in the presence of medical mistake. CONCLUSION: The most important difference between the groups was not the pedagogical pattern but the exposition time to the theme. PBL gives the chance to distribute the theme in different situations accelerating the acquisition of knowledge in Medical Ethics. It was realized that a revitalization on Medical Ethics teaching is necessary at our institution, aiming a better integration with the socio-economical situation in our country. PMID- 15286860 TI - [Spirometric values of obese and non-obese subjects on orthostatic, sitting and supine positions]. AB - BACKGROUND: It is possible that obesity could lead to pulmonary restriction with decreasing lung volumes. However, controversies about this restriction and its etiology still exist. Thus, the purpose of this report was to evaluate the effects of body weigh excess on spirometry, on three different body positions, evaluated by Body Mass Index (BMI), percentage of fatness and the ratio of abdominal girth to hip breadth (AG/HB). METHODS: Forty-six sedentary volunteers, with ages between 20 and 40 years, were studied and distributed on five groups, based on BMI. Skin fold thickness and ratio of abdominal girth to hip breadth (AG/HB) of the volunteers were measured. FVC, FEV1 and ratio of FEV1 to FVC were measured on three different body positions--sitting, supine and orthostatic positions. RESULTS: Comparing the values measured and predicted between the groups, no difference was detected. Comparing body positions, the supine position shows lower values than sitting and orthostatic positions (p<0.05). Associations between CVF, VEF1 e VEF1/CVF values and BMI, percentage of fatness and ratio of AG/HB were not found. CONCLUSIONS: Spirometric values from obese people are into normality range and decrease on the supine position. PMID- 15286861 TI - [Ophthalmological findings in HIV infected patients in the post-HAART (Highly Active Anti-retroviral Therapy) era, compared to the pre-HAART era]. AB - BACKGROUND: To determine the frequency of the ophthalmological findings in Human Immunodeficience Virus (HIV) infected patients in the post-HAART (Highly Active Anti-retroviral Therapy) era and to compare it to data from the Federal University of Sao Paulo in the pre-HAART era10,11. METHODS: Charts from 200 consecutive patients examined from May 2000 to February 2001 were reviewed. RESULTS: From the total of 200 patients, 84 (42%) presented ophthalmological findings related to HIV infection in the first ocular examination: 36 (18%) with Cytomegalovirus retinitis (9 active and 27 inactive), 22 (11%) with ocular toxoplasmosis, (9 active and 13 inactive), 15 (7.5%) with retinal detachment (10 secondary to CMV retinitis, 3 to ocular toxoplasmosis and 2 not determined), 8 (4%) with cataract (5 secondary to CMV retinitis, 1 to ocular toxoplasmosis and 2 not determined), 8 (4%) with cotton wool spots, 6 (3%) with acute retinal necrosis (4 active and 2 inactive) and 3 (1.5%) with intersticial keratitis. All patients with acute retinal necrosis showed T CD4 cells under 100 cells/microL. CONCLUSION: The authors identified in the post-HAART era, rise on number of patients with normal ophthalmologic exam and decreased number of cases of CMV retinitis and ocular toxoplasmosis when compared to the pre-HAART era. PMID- 15286862 TI - [Baseline clinical features of 483 children and adolescents with primary vesicoureteral reflux: a retrospective study]. AB - OBJECTIVES: To evaluate the clinical/radiological features of patients with primary vesicoureteral reflux (VUR) admitted to a single institution from 1969 to 1999. METHODS: At admission, after the institution of chemoprophylaxis, patients were investigated by CUM, DMSA, and US. The children were managed with periodical clinical and laboratory evaluations. Analyses were performed with the data obtained at admission. RESULTS: A total of 483 patients were enrolled in the protocol. There was a predominance of females (70%) and caucasian race (70%). Mean age at VUR diagnosis was 26 months and 92.5% of the patients had urinary infection before admission. A total of 710 refluxing units were analyzed. The distribution of reflux grade was: grade I (49; 7%); II (254; 36%); III (190; 26%); IV (161; 23%) e V (56; 8%). Approximately one half of the units analyzed were from patients presenting renal damage at admission. The distribution of the severity of renal damage was as follows: mild (36%), moderate (34%), and severe (30%). There was a significant risk of severe renal damage for the males (OR = 1.74, 95% CI = 1.2 - 2.5, p = 0.002). CONCLUSION: There was a predominance of VUR among females and most presented a mild or moderate degree. However, there was a high percentage of renal damage at admission and a tendency to greater morbidity for the males. PMID- 15286863 TI - [Hypertension in women: study in mothers of students from Jaboatao dos Guararapes, Pernambuco, Brazil]. AB - OBJECTIVE: The purpose of this research was to study the prevalence of hypertension in apparently healthy women community with a view preparing a program for primary prevention of hypertension. METHODS: The population survey involved a representative sample of mothers of 126.800 students enrolled at schools of Jaboatao dos Guararapes, Pernambuco, Brasil. The population size (1.273 students) was selected from a 5% +/- 2% expected frequency of the hypertension in children with a confidence level of 99.9%. This study comprised 21 schools and a random sample of 986 mothers of 1.601 students). Two blood pressure measurements separated by 2 minutes were carried out in the schools on two subsequent visits. The corporal mass index was determined in 893 mothers. In a group of 671 mothers, a midstream urine specimen was collected and the dip - stick was used to assess the presence of urinary abnormalities. RESULTS: The prevalence of the hypertension was 27.7% and it was significantly higher (p<0.01) in overweight (28.5%) and obese women (57%) than normal weight women (14%) and underweight women (12%). There was significant difference regarding the presence of glucosuria between the hypertensives and the normotensives women (3% and 0%, respectively). In a group of 62 hypertensives women with followup at 3 to 12 month interval, the blood pressure has been controlled in 13 (21%) and remains below the initial hypertensive stage in 27 (43.5%). CONCLUSIONS: The prevalence of hypertension (27,7%) do agree with the data in the literature concerning hypertension in women (26%), showing a higher tendency in overweight and obese women. We are continuing the followup of the hypertensive women as well as developing a community program for primary prevention of hypertension in the target population. PMID- 15286864 TI - [Puberty and growth in children and adolescents with neuro fibromatosis type 1]. AB - BACKGROUND: This study evaluates anthropometric and pubertal development parameters in children and adolescents with sporadic and familial occurrence of neurofibromatosis type 1 (NF-1). METHODS: The study involved 23 patients (15 boys, 8 girls) with sporadic (n=12) and familial NF-1(7 affected mothers and 4 fathers), with average age of 12 years (range 5-20 years). All patients answered to a standard questionnaire including family and personal data and, following, they underwent systematized physical examination, with special attention to anthropometric data, dermatologic and pubertal stage according to Tanner's criteria evaluation. RESULTS: Short stature was detected in a 17 year-old patient, 148.5cm height, with score Z of -4.16 and being the even outside of the family conduit. Three male patients had head circumference above of + 2DP from on average for the age. It was not found no case of precocious or delayed puberty. CONCLUSIONS: In our study, do not we find significant alterations regarding the stature and pubertal development. PMID- 15286865 TI - [Identification of insulinomas by endoscopic ultrasonography]. AB - BACKGROUND: The aim of this study is to compare EUS and the others diagnostics tests in the correct localization of insulinomas. METHODS: We prospectively investigated 30 patients with endoscopic ultrasound with a clinical diagnosis of insulinomas prior to surgical exploration. They were submitted to abdominal ultrasonography, spiral computed tomography and four patients were submitted to magnetic ressonance before EUS. Surgery was the gold standard for tumor localization. RESULTS: Twenty-six tumors were benign (86.6%) and four were malign (13.4%). The median size tumors detected by EUS was 1.5 cm. The overall sensitivity of EUS in identifying insulinomas was 86.6% compared to 33% for CT, 40% to MRI and 90.9% to IUS. In 12 patients we were able to perform EUS-guided fine needle aspiration. Insulinoma was diagnosed in ten cytological specimens (83.3%). Tumors located in the head and body of the pancreas were seen by EUS in all patients, respectively but those located in the tail were diagnosed only in 55.5% of the cases. CONCLUSIONS: EUS has a high sensibility in the identification and localization of pancreatic insulinomas and should replace traditional methods of image when clinical suspicion is high. PMID- 15286866 TI - [Obesity prevalence among oldest-old and its association with risk factors and cardiovascular morbidity]. AB - OBJECTIVE: To evaluate the obesity prevalence and its association with risk factors and cardiovascular morbidity in the oldest old (>80 years old) residing at the municipality of Veranopolis - RS, Brazil. METHODS: 196 elderly participated in the study (69 male and 127 female), 91% of the population aged >80 until June, 1996. For obesity evaluation and classification, we used the body mass index (BMI) and the World Health Organization (WHO) and National Health and Nutrition Examination Survey (NHANES III) criteria. The cardiovascular risk factors investigated were sex, age, systemic hypertension, hypercholesterolemia, hypertriglyceridemia, diabetes mellitus (DM) and smoking. For cardiovascular morbidities, we considered acute myocardial infarction, intermittent claudication and stroke. Waist-hip ratio (W/H), regular consumption of alcoholic beverages and physical activity were investigated too. RESULTS: The obesity prevalence was 23.3% according to WHO (without difference between sex, p=0.124) and 45.6% according to NHANES III criteria (significantly higher in female, p=0.05). Obesity associations with risk factors were sex-dependent (the obese females presented higher levels of systolic blood pressure and glucose, lower levels of HDL-c, and higher systemic hypertension and DM frequencies; while the obese males presented higher levels of diastolic blood pressure, total cholesterol, LDL-c and higher hypercholesterolemia frequency). W/H and triglycerides, as well as hypertriglyceridemia frequency, were higher in obese people. CONCLUSIONS: The obesity prevalence was high among the long-living elderly, and its association with cardiovascular risk factors was sex-dependent. As regards morbidities, we did not observe differences between obese and non-obese people. PMID- 15286867 TI - [Clinical diagnosis and anatomic-pathologic diagnosis: disagreements]. AB - Many pathologic entities in the clinical practice generate disagreements regarding its identification, not only by its likeness with other lesions but also by its semantics. BACKGROUND: The goal of this work is to clarify which disagreements are more frequent in the clinical practice, supply new knowledges to facilitate the identification of the larger pathologies controversies and to enlarge its differentials diagnosis. METHODS: We revised 1825 reports of referring biopsies to the period of 1992 up to 1999, belonging to the Laboratory of Pathological Anatomy of Unoeste, being excluded 439 lauds that did not introduce diagnostic hypothesis, or that had as signals hypothesis and clinical indications or "for clearing". We confronted the clinical hypothesis with the diagnosis Anatomic-pathological, obtaining itself 444 (32.05%) of discordant cases. RESULTS: Note that the biggest disagreements were between Hansen's disease clinical diagnosis, which in 65.7% cases were chronicles Unspecific Dermatitises, between Sebaceous Cyst, which in 80% cases were cyst of Epidermical Inclusion and Incomplete Abortion, which in 68.2% cases were complete Abortion. CONCLUSION: We conclude that an adequate concepts definition discerning an anamnese and narrow correlation of the clinical characteristics of the lesions promote a minor disagreements number among clinical diagnosis and Anatomic-pathological. PMID- 15286868 TI - [Clinical and epidemiologic evaluation of pressure ulcers in patients at the Hospital Sao Paulo]. AB - BACKGROUND: To determine the profile of patients at the Hospital Sao Paulo with pressure ulcers (PU). METHODS: A prospective study was carried out. Seventy-eight patients with pressure ulcers were evaluated between May 1st and 31st of 2002. The questionnaire consisted of demographic and clinical data of the population, PU classification and Braden Scale. RESULTS: Concerning 78 patients with PU: 66.7% were over 61 years old (average: 64). The average period of hospitalization was 33 days. 68% (53) developed ulcers while hospitalized, of which 43.7% (34) were pre-ulcers. The most frequent causes of hospitalization were neurological diseases (29.5%) and cancer (29.5%). Regarding the classification of the pressure ulcers, all stages were found on the sacrum, being 24.4% (19) pre-ulcer, 38.5% (30) stage II, 11.5%(9) stage III and 12.8%(10) at stage IV. According to the Braden Scale, half of the patients hospitalized showed severe risk of PU development, while 20.5% (16) showed moderate risk and 19.3% (15) showed low risk. CONCLUSIONS: The study showed a high risk of PU development, indicating the importance of having knowledge about the main characteristics of the hospitalized patients who may develop pressure ulcers, and, thus, preventing them. PMID- 15286869 TI - [Predictors of choledocholithiasis in patients sustaining gallstones]. AB - BACKGROUND: The aim of this study was to identify clinical, biochemical and ultrasonographic predictors of choledocholithiasis in patients sustaining gallstones assessed by cholangiography. METHODS: In a prospective study, 148 patients were analyzed regarding clinical, biochemical and ultrasonographic data. All patients underwent cholangiography, either preoperative endoscopic or during cholecystectomy. Each variable was compared between the ones who sustained lithiasis in the biliary tree and the others, in order to find out the predictors of choledocholithiasis. Sensibility, specificity, positive predictive value, negative predictive value (NPV) and accuracy were calculated. Spearman correlation, Odds ratio and logistic regression were employed for the statistical analysis, considering p<0.05 as significant. RESULTS: The variables that showed statistical significance were: presence of jaundice, elevated blood serum levels of alkaline phosphatase, g glutamyltransferase (gamma GT), aspartate aminotransferase, alanine aminotransferase, total bilirrubin, and biliary tract dilatation or choledocholithiasis in the ultrasound. The logistic regression presented an equation capable of predicting the probability of choledocholithiasis based in the variables: jaundice, presence of choledocholithiasis in the ultrasound, and blood levels of gamma GT. The best option to exclude the presence of choledocholithiasis was gamma GT, as it held the higher NPV. Every patient with choledocholithiasis in this sample sustained at least one of the preoperative criteria analyzed. CONCLUSIONS: Jaundice and choledocholithiasis at the ultrasound were the best predictors of choledocholithiasis; as well as gamma GT was the most reliable factor to exclude this diagnosis. PMID- 15286870 TI - [Racemic bupivacaine, levobupi vacaine and ropivacaine in regional anesthesia for ophthalmology -- a comparative study]. AB - BACKGROUND AND OBJECTIVE: Racemic bupivacaine, used in peribulbar anesthesia owing to its high potential to promote motor blockade, presents a smaller safety margin for cardiotoxicity in relation to ropivacaine and levobupivacaine. The objective of this study was to compare the degree of motor blockade and alteration of intraocular pressure (IOP) produced by racemic bupivacaine, levobupivacaine and ropivacaine in peribulbar block. METHOD: Ninety seven patients, ASA physical status I and II, submitted to peribulbar anesthesia, were randomly allocated into three groups: group A-(n=16) receiving racemic bupivacaine 0.75% with epinephrine 1:200.000; group B -(n=16) levobupivacaine 0.75% with epinephrine 1:200.000; group C -(n=15) ropivacaine 0.75%. A single inferior injection peribulbar anesthesia was performed with 7 ml of the anesthetic solution plus 280 UI of hyaluronidase. The IOP and the degree of motor blockade were registered five minutes before injection and 1,2,3,4,5 and 10 minutes after it. The motor blockade was evaluated according to Nicoll's scale. For statistical analysis, Wilcoxon's test, simple frequency analysis, and Student t test were used. p<0.05 was considered significant. RESULTS: There were no significant differences between groups with respect to the degree of motor blockade. The IOP variation between the groups was not clinically significant. CONCLUSIONS: Considering the advanced age of most of these patients and the high concentrations of local anesthetics used in peribulbar blockade, the use of ropivacaine and levobupivacaine produces motor blockade as effective as racemic bupivacaine while minimising risks for cardiotoxicity. PMID- 15286871 TI - [Validation of the Brazilian version of Alcohol, Smoking and Substance Involvement Screening Test (ASSIST)]. AB - BACKGROUND: Considering the increasing levels of psychoactive substance use in Brazil and around the world, it is necessary the development of an instrument to early detection, that could be considered valid, reliable and useful at primary health care settings. In order to do that, an international team of researchers, supported by the World Health Organization (WHO) developed the Alcohol Smoking and Substance Screening Test (ASSIST). The objective of the present study was to evaluate the psychometric properties of the Brazilian version of ASSIST, as well as its reliability and concurrent validity in the detection of psychoactive substance use and related problems. METHODS: The ASSIST and three validated diagnostic instruments (Mini-Plus, AUDIT, RTQ) were applied to 99 patients of primary/secondary health care services and to 48 patients in treatment for alcohol and drug dependence in specialized services, at Sao Paulo and Curitiba cities. RESULTS: The ASSIST' scores to alcohol showed good correlation with AUDIT' scores. The ASSIST presented high sensitivity and specificity in the detection of alcohol, cannabis and cocaine abuse/dependence, considering the MINI Plus diagnostic the "gold-standard". The reliability of the instrument was good (Cronbach's alpha of 0.80 to alcohol, 0.79 to cannabis and 0.81 to cocaine). CONCLUSIONS: The psychometric properties of the Brazilian version of the ASSIST seem to be satisfactory, recommending its use in patients of primary/secondary health care services. PMID- 15286872 TI - [Proteinuria in hypertensive syndrome of pregnancy: maternal and perinatal outcome]. AB - AIM: The purpose of this study was to determine the role of proteinuria on pregnancy outcome in hypertensive syndrome with singleon pregnancies. METHODS: Transversal study with retrospective data of 334 pregnancies complicated by hypertensive syndromes who were delivered in the Department of Obstetrics of UNIFESP/EPM from January 1, 1999 to December 31, 2002. RESULTS: The patients were divided into four groups: (I) without proteinuria (n-203); (II) with proteinuria of 0.3 to 1.0g (n-39); (III) 1.0 to 2.0g (n-45); and (IV) 2.0g or more. Without proteinuria there was one case of placental abruption. The presence of proteinuria predicted adverse maternal outcome with increase of complications proportional to his elevation; among them, HELLP syndrome was the most frequent with 30.5% (40/131) followed by eclampsia with 3.8% (5/131), DPP 3.1% (4/131) and renal insufficiency with 0.7% (1/131). It was confirmed one maternal death in that group, when Maternal Mortality of 763/100.000nv was added up. As to the perinatal effects there was not increase of adverse effects without proteinuria. In the presence of proteinuria and its levels was observed the worst perinatal outcome with the elevation of the following indicatives: increase prematurely (62.2% vs 11.5%), newborn with weight < 2500g (6.5% vs 1.5%), newborn with Apgar < 7 in the 5th minute (30.4% vs 3.5%), concepts with growing restriction of intrauterine (41.9% vs 6.5%), newborn interned in the neonatal undid, (59.8% vs 15.5%) stillborn (14.4% vs 1.4%), neonatal deaths (6.1% vs 0.98%). The Perinatal Mortality was greater with proteinuria (175 vs 19,7) and, when = 2.0g (297.8 vs 19.6). CONCLUSIONS: The presence of proteinuria in the hipertensives syndromes during gestation and the elevation of their levels increase the risks of maternal complications, especially HELLP syndromes and eclampsia. Besides, it was observed a significative incidence of premature birth, newborn with Apgar < 7, weight < 2500g, IUGR, stillborn and neonatal deaths. PMID- 15286873 TI - [Acute myocardial infarction -- acute coronary syndrome with ST-segment elevation]. AB - Cardiovascular diseases continue to be the first cause of death in Brazil -- responsible for almost 32% of all deaths. In addition, they are the third major cause of admission in the country. Among them, acute myocardial infarction is still one of the major causes of morbidity and mortality. Despite of the last decade's therapeutic advances, acute myocardial infarction still shows remarkable rates of mortality, and great part of the patients do not receive the adequate treatment. The opening of the Coronary Care Units and the introduction of reperfusion treatment with fibrinolytics or primary angioplasty were fundamental to reduce mortality and complications related to myocardial infarction. Important beneficial effects to the current treatment include less ventricular dysfunction and better control of ventricular arrhythmias. The need of early reperfusion is crucial for the good prognosis after a myocardial infarction. The objective of this review is to emphasize the modern basic concepts of the pathophysiology, diagnosis and treatment of acute myocardial infarction, according to national and international guidelines. PMID- 15286874 TI - [Evidence based clinical practice. Part III -- Critical appraisal of clinical research]. AB - Evidence based health care begins with a clinical question and the search on data bases to retrieve the relevant information, that was the issue of two preceding articles of this series. At present it will be discussed how to critically appraise the medical literature using the clinical epidemiological methodology. Clinical research aims to develop diagnostic and therapeutic procedures measuring association and causality between the exposure and outcome. In this case the exposures are signs, symptoms, laboratorial or image exam, and therapy intervention. It is a mistake to take surrogate end-points instead of clinical outcomes. The main types of clinical study design are diagnostic, prognostic, therapeutic and harm/etiology. Experimental, physiologic and animal studies are useful for the medical undergraduate education, but do not contribute with clinical decisions. The study designs are classified according with the presence of a control group, patient's follow-up, and therapy interventions. The evidence hierarchy was done by the previous characteristics and the presence of systematic bias. Systematic reviews are stronger than the primary observational studies and are on the top when they revised randomized clinical trial. Since 1998 the proportion of evidence based practice guidelines was increasing compared with systematic reviews or other types of practice guidelines, although the former still are in a few numbers. The article critical appraisal must answer the clinical question, and need to have consistent study design and bias under control. In conclusion we ought to offer methodological actualization to interested physicians and put the information already critically assessed on evidence-based practice guidelines. PMID- 15286876 TI - Are anti-interferon antibodies the cause of failure in: chronic HCV hepatitis treatment? AB - A follow-up study was made of 94 chronic hepatitis C patients at a hepatitis clinic in Brazil, after interferon alpha (IFN-alpha) therapy, to determine the influence of anti-interferon antibodies on treatment outcome. Patients diagnosed as having chronic hepatitis C, confirmed by PCR (HCV RNA) and liver biopsy, were treated with interferon alpha 2a or 2b for at least six months, and were followed up for 24 weeks after termination of treatment in order to assess biochemical, virological and clinical pathology responses. Only 6% of the 94 patients developed anti-IFN antibodies, 70% presented a biochemical response and 23% maintained a sustained virological response. Clinical evaluation revealed that in only 2 patients was there progression of fibrosis; the necro-inflammatory score indicated that 72% maintained the same activity, 12% had worsening necro inflammatory activity, and the remaining 16% had decreased activity. There was no significant correlation of demographic and laboratory variables with levels of anti-interferon antibodies. Similarly, biochemical and virological responses were not influenced by anti-interferon antibodies. Multivariate analysis by logistic regression revealed that clinical pathological parameters, staging and necro inflammatory activity did not influence the response to the virus. PMID- 15286877 TI - Accidents with biological material among undergraduate nursing students in a public Brazilian university. AB - During their academic activities, undergraduate nursing students are exposed to contamination by bloodborne pathogens, as well as by others found in body fluids, among which are the Human Immunodeficiency (HIV), Hepatitis B and C viruses. We developed a profile of victimized students, characterizing accidents with biological material occurring among undergraduate nursing students at a public university in Sao Paulo State, Brazil. We identified the main causes and evaluated the conduct adopted by students and their reactions and thoughts concerning the accidents. Seventy-two accidents were identified, of which 17% involved potentially contaminated biological material. Needles were the predominant cause of accidents. The most frequently involved topographic areas were the fingers. Only five students reported the accidents and sought medical care. Among these, two students were advised to begin prophylactic treatment against HIV infection by means of antiretroviral drugs. It was found that the risk of accidents is underestimated and that strategies such as formal teaching and continual training are necessary in order to make students aware of biosafety measures. PMID- 15286878 TI - SENTRY antimicrobial surveillance program report: Latin American and Brazilian results for 1997 through 2001. AB - The alarming emergence and spread of antimicrobial resistance among common bacteria threatens the effectiveness of therapy for many infections. Surveillance of antimicrobial resistance is essential to identify the major problems and guide adequate control measures. Several resistance surveillance programs have been implemented in North America and Europe in the last decade; however, very few programs have assessed antimicrobial resistance in Latin American countries. The SENTRY Antimicrobial Surveillance Program was initiated in 1997 and represents the most comprehensive surveillance program in place at the present time worldwide. The SENTRY Program collects consecutive isolates from clinically documented infections in more than 80 medical centers worldwide (10 in Latin America). The isolates are collected according to the type of infection (objectives) and susceptibility tested in a central microbiology laboratory by reference broth microdilution methods according to NCCLS guidelines. The Program also incorporated molecular typing (ribotyping and PFGE) and resistance mechanism analysis of selected isolates. In this report we present a very broad analysis of the data generated by testing almost 20,000 bacterial isolates against more than 30 antimicrobial agents. The susceptibility results (MIC(50), MIC(90) and % susceptible) are presented in 11 tables according to the organism and site of infection. The data from Brazil, as well as the data from isolates collected in 2001, are analyzed separately. This report allows the evaluation of the activities numerous antimicrobial agents against clinical isolates collected in Latin American countries. PMID- 15286879 TI - Phagocytosis and killing of epidemic methicillin-resistant Staphylococcus aureus by human neutrophils and monocytes. AB - Staphylococcus aureus is a pathogen that has been associated with nosocomial infections since the preantibiotic era. Since the introduction of antibiotics in medical practice in the 1940 s, methicillin-resistant S. aureus (MRSA) strains have been emerging in various parts of the world. In view of the important role of the phagocytic system in the defense against this bacteria, we decided to study phagocytosis by neutrophils and monocytes of an epidemic MRSA strain in Sao Paulo, Brazil, in comparison with methicillin-sensitive strains. Complement system opsonins are fundamental for efficient ingestion of the resistant and sensitive strains by both types of phagocytes. We found no association of the opsonic requirement of the MRSA strain with the multiresistance phenotype. On the other hand, the MRSA strain was found to be more resistant to the effector mechanisms of neutrophils than both sensitive strains when opsonized with fresh serum, despite the phagocytosis results. This fact suggests that the intracellular killing of S. aureus is an additional parameter of bacterial virulence, but new approaches must be implemented to study the interactions of this MRSA strain with phagocytes in order to investigate the possible factors involved in its behavior in response to neutrophil effector mechanisms. PMID- 15286880 TI - Gatifloxacin in the treatment of community-acquired pneumonias: a comparative trial of ceftriaxone, with or without macrolides, in hospitalized adult patients with mild to moderately severe pneumonia. AB - Community-acquired pneumonia is very common, but some of the cases do require hospitalization for treatment, particularly when older patients and/or co morbidities are involved; both "typical" and "atypical" respiratory pathogens take part etiologically, and there is increasing concern about the emergence of resistance. There is interest in therapeutic options that can: a) comprehend such a spectrum of bacteria and resistance; b) allow parenteral to oral sequential treatment. We made a multicenter, prospective and randomized trial to compare the "standard" treatment of ceftriaxone IV alone or in combination with erythromycin IV, followed by clarithromycin PO (ceftriaxone treatment arm), with gatifloxacin IV, followed by oral administration (gatifloxacin treatment arm). The need for hospitalization was based on clinical criteria as judged by the investigators. Standardized criteria for diagnosis and follow-up were employed. Fifty-six patients were enrolled, with 48% over 65 years old, and there were frequent co morbidities. Of these, 51 were clinically evaluable, 26 in the gatifloxacin and 25 in the ceftriaxone arm, with comparable success rates, 92% and 88%, respectively, even when major prognostic factors were considered. There were no serious adverse events or significant laboratory value changes attributable to the study drugs. Gatifloxacin as monotherapy (initially IV then orally until completion of treatment) was shown to be effective and safe, comparable to ceftriaxone IV alone or in combination with a macrolide (initially IV then orally until completion of treatment), in empirical therapy for community-acquired pneumonias, for patients that, at the physician s discretion, require initial treatment as inpatients. PMID- 15286881 TI - Effect of citrinin and in association with aflatoxin B(1) on the infectivity and proliferation of Toxoplasma gondii in vitro. AB - Macrophages exposed to 10 mug/mL citrinin (CTR) or 0.01 mug CTR mixed with 0.04 mug aflatoxin B1 (AFB1) for a period of 2 h at 37 masculine C, were infected with 10(6) Toxoplasma gondii tachyzoites/muL. The parasites were treated with mycotoxins (2 h at 37 masculine C) before being added to the macrophage culture. The number of tachyzoites was quantified 2, 24, 48, 72 and 96 h after infection. During the first 2 hours, 59% infectivity was observed in the control. After exposure to CTR or the mixture of toxins (CTR-AFB1), macrophages were infected with 77.5% and 75% of the inoculated tachyzoites, respectively. Similarly, 72.3% of the cells were infected when cultured together with previously treated parasites. The treatment with CTR-AFB1 gave rise to 2.9 times more tachyzoites than the control at 72 h. An increased number of parasites was recovered from macrophages exposed to CTR after 96 h, and to CTR-AFB1 after 72 h of culture; The number of tachyzoites recovered from the supernatant was 1.94 and 2.06 times higher, respectively, than in the control (5 x 10(5) +/- 0.054 /mL). PMID- 15286882 TI - Klebsiella pneumoniae with multiple antimicrobial resistance. AB - A Klebsiella pneumoniae strain was isolated from the urine of a patient at one of the centers participating in the 2001 edition of the MYSTIC program in Brazil. The initial phenotypic findings of the isolated K. pneumoniae presented an unusual MIC of 8 microg/mL to meropenem, 2 microg/mL to imipenem, elevated MICs to broad spectrum cephalosporins (ceftazidime/cefotaxime/cefepime MIC > 256 microg/mL), aminoglycosides (gentamycin > 256 microg/mL and tobramycin = 48 microg/mL), piperacillin/tazobactam (MIC > 256 microg/mL) and susceptibility to ciprofloxacin (MIC = 0.25 microg/mL). The strain also tested positive for ESBL production with double-disk and E-test methodologies. More detailed investigation revealed that the strain produced a SHV-4 type enzyme and also lacked a 36 kDa outer membrane porin. PMID- 15286883 TI - Tuberculosis of the cystic duct lymph node. AB - Tuberculosis of the cystic duct lymph node associated with cholelithiasis is rare. We report a case of a 40 year-old woman with this pathology. She presented with anorexia, biliary colic, postprandial fullness and fever. Imaging studies revealed cholelithiasis and several visible portal lymph nodes. Cholecystectomy was performed and histopathological examination showed tuberculosis of the cystic duct lymph node without affecting the gallbladder. The presence of gallstones and lymphadenopathy in computed tomography, associated with persistent fever and symptoms that resemble cholecystitis, should cause suspicion of tuberculosis. However, diagnosis is usually achieved by microscopic appearance of caseating granulomas and isolation of Mycobacterium tuberculosis. The treatment in this case consisted of cholecystectomy and antitubercular chemotherapy. PMID- 15286884 TI - Meningoencephalitis and new onset of seizures in a patient with normal brain CT and multiple lesions on MRI. AB - Toxoplasmic encephalitis is the most common cerebral mass lesion in patients with AIDS. The definitive diagnosis requires direct demonstration of the tachyzoite form of Toxoplasma gondii in cerebral tissue. The presumptive diagnosis is based on serology, clinical and radiological features, and on response to anti Toxoplasma therapy. Typically, patients have a subacute presentation of focal neurological signs, with multiple lesions in computed tomography (CT) or magnetic resonance imaging (MRI). However, the neurological and CT scan spectrum is broad. We report a case of toxoplasmic encephalitis in a heterosexual man without prior history of HIV infection. He was admitted with four days of headache, confusion, and new onset of seizures. His brain CT disclosed no alterations and MRI revealed multiple lesions. Empirical specific anti-Toxoplasma therapy was initiated and the patient experienced excellent clinical and radiological improvement. His HIV tests were positive and the CD4+ cell count was 74 cells/ml (8.5 %). On follow up, three months later, the general state of the patient was good, without neurological sequelae and with a normal MRI. We concluded that toxoplasmic encephalitis should be considered in the differential diagnosis of meningoencephalitis in sexually active individuals, including cases without prior history or suspicion of HIV infection, and no abnormalities on CT scan. PMID- 15286885 TI - [The role of nitric oxide in trauma and infection]. AB - Nitric oxide (NO) is a very important molecule for homeostasis. It is involved in several pathological conditions ranging from hypertension to septic shock. It is synthesized from L-arginine, which is catalyzed by nitric oxide synthase (NOS). Both constitutional and inducible NOS are involved in NO synthesis. While NO produced by constitutional NOS is required for normal physiologic processes, excessive production by inducible NOS results in injury and tissue damage. Induced NO may be either protective or damaging in acute inflammatory conditions. As a result of pluripotent activities, NO presents as a paradoxical phenomenon in almost all conditions in which confusing aspects arise concerning the pathophysiology. This article reviews the role of NO in trauma and infections. PMID- 15286886 TI - [The beneficial effect of L-carnitine in rat renal ischemia-reperfusion injury]. AB - BACKGROUND: We examined the effect of L-carnitine on experimental renal ischemia reperfusion (I-R) injury. METHODS: Seventy-two Wistar albino rats were divided into five groups, four of which underwent right nephrectomy and left renal I-R for 60 min (groups 2 and 4) and 90 min (groups 3 and 5). The sham operated group (n=8) was left as controls. Intraperitoneal carnitine (200 mg/kg) was given to groups 4 and 5 three hours before operation. For histopathologic studies, half of the rats in the study groups (n=8) and the remaining rats were sacrificed 15 min and seven days after reperfusion, respectively. Blood urea nitrogen (BUN) and serum creatinine levels were also measured at 15 min and on Day 3 and Day 7. RESULTS: Concerning the 90 min I-R groups, survival was improved in carnitine treated rats (7/8 vs 5/8). Of untreated groups, the 90 min I-R group exhibited significantly higher BUN (p=0.0023) and serum creatinine (p=0.0086) levels on Day 3. Although BUN and serum creatinine levels did not show significant differences between carnitine-treated and untreated rats in the two 60 min I-R groups throughout the study period, carnitine-treated rats in the 90 min I-R group exhibited significantly improved BUN (p=0.0063) and serum creatinine (p=0.0013) levels on Day 3. The severity of histopathologic changes was lower in all carnitine-treated rats 15 min after reperfusion. On Day 7, the mean histopathologic score was significantly lower in carnitine-treated rats in the 90 min I-R group than that of untreated rats exposed to I-R injury of same duration (p=0.0078). CONCLUSION: Our results suggest that L-carnitine has beneficial effects on renal function, histopathologic changes, and survival in renal ischemia-reperfusion injury. PMID- 15286888 TI - The value of 5-hydroxyindolacetic acid levels in spot urine in the diagnosis of acute appendicitis. AB - BACKGROUND: We investigated the value of 5-hydroxyindolacetic acid (5-HIAA) levels in spot urine in the diagnosis of acute appendicitis. METHODS: Forty-three patients (11 females, 32 males; mean age 26.3 years; range 16 to 73 years) who were admitted to the emergency room with acute abdominal pain and suspected acute appendicitis were evaluated by means of physical examination, leukocyte counts, urine analysis, measurements of 5-HIAA in spot urine, abdominal x-rays, and sonography. Of these, 25 patients underwent appendectomy after a clinical diagnosis of acute appendicitis. Spot urine analyses were made in all the patients within the first hour of admission with the use of spectrophotometric and colorimetric methods; 5-HIAA levels were read at 540 nm. RESULTS: At laparotomy, 22 patients (88%; 3 females, 19 males; mean age 26.18 years; range 15 to 38 years) were found to have acute appendicitis (11 phlegmonous, 11 gangrenous), whereas three patients (12%; 2 females, 1 male) had negative findings. No significant differences were found between patients with confirmed acute appendicitis, patients without appendectomy, and those with negative laparotomy with respect to the mean leukocyte counts and 5-HIAA levels (p>0.05). The sensitivity, specificity, positive predictive value, negative predictive value, and accuracy of 5-HIAA for the diagnosis of acute appendicitis were calculated as 22%, 93%, 71%, 62%, and 63%, respectively. CONCLUSION: Measurement of 5-HIAA levels in spot urine is not helpful in the diagnosis of acute appendicitis. PMID- 15286887 TI - Interhospital transport of pediatric patients requiring emergent care: current status in Turkey. AB - BACKGROUND: This study was designed to evaluate the current situation of interhospital transport of pediatric patients requiring emergent care. METHODS: Using a clinical prospective and multicenter design, 1,666 interhospital transports of pediatric patients were evaluated in 18 centers. Non-emergency transports and newborn transports were not included, so 854 transports were eligible for evaluation. Data were collected by means of a comprehensive form filled by a physician at the receiving hospital. RESULTS: The physicians who gave the decisions for the transports were pediatricians in 60%, general physicians in 15.4%, and residents in 6%, while no identification existed in 159 transports (18.6%). The receiving hospitals were not notified prior to the transport in 79.3%. Pretransport information about the patients were adequate in 26.1% and inadequate in 31.8%; no information was available in 42.1%. Ambulances were used in 64.4% of the transports, of which only 16.2% was fully equipped. Unqualified or inexperienced personnel were in charge in 42.8% of the transports. In 26.3% of the transports, the patients arrived at the receiving hospital in an agonized state. CONCLUSION: It appears that there are no established guidelines for the emergency transport of pediatric children in Turkey. PMID- 15286889 TI - [Factors affecting morbidity and mortality in mechanical intestinal obstruction]. AB - BACKGROUND: We evaluated the effects of factors related to both the patient and the treatment on morbidity and mortality of patients with mechanical intestinal obstruction. METHODS: A total of 152 patients (83 males, 69 females; mean age 55.05 years; range 15 to 90 years) who underwent surgery for intestinal obstruction were retrospectively evaluated. The effect of several factors on morbidity and mortality during hospitalization (mean 8.6 days; range 1 to 105 days) was assessed, including age, sex, associated diseases, the presence of intestinal necrosis, previous abdominal operations, the time of the operation, and the level of obstruction. Statistical evaluations were made with the use of a multiple logistic regression analysis. RESULTS: Complications were encountered in 52 patients (34.2%), the most common being systemic complications (15.8%) followed by wound site infections (11.2%). During hospitalization, mortality occurred in six patients (4%), all of whom had at least one associated disease. Logistic regression analysis showed that age beyond 75 years (p<0.01), male gender (p<0.01), associated disease (p<0.05), non-viable strangulation (p<0.001), previous operations (p<0.05) and malignancies (p<0.05) were significant independent factors affecting the complication rate. On the other hand, only non viable strangulation (p<0.05) had significant independent effect on mortality. CONCLUSION: Patient-related factors should be taken into consideration in the evaluation and treatment of mechanical intestinal obstruction. PMID- 15286890 TI - [Is interval appendectomy necessary after conservative treatment of appendiceal masses?]. AB - BACKGROUND: This prospective study was conducted to investigate whether interval appendectomy was necessary after successful conservative treatment of appendiceal masses. METHODS: Thirty-seven patients with a diagnosis of appendiceal mass by physical examination and ultrasonography were initially treated conservatively with broad-spectrum antibiotics, anti-inflammatory drugs, and, if required, intravenous fluid treatment. Interval appendectomy was ruled out in 28 patients who responded well to conservative treatment, three of whom were then lost to follow-up. The remaining 25 patients (9 females, 16 males; mean age 25 years; range 17 to 54 years) were monitored for recurrent appendicitis and other causes of appendiceal mass. The mean follow-up period was 35 months (range 6 to 66 months). RESULTS: The mean duration of abdominal symptoms was nine days (range 3 to 20 days). The mean length of hospital stay was 14 days (range 10 to 21 days) in patients who responded to conservative treatment. Recurrent appendicitis developed in three patients (12%; 2 males, 1 female). Two patients who presented with acute appendicitis within six months after discharge and one patient who developed chronic abdominal right lower quadrant pain unresponsive to medical treatment a year after discharge underwent appendectomy. No other complications were seen with conservative treatment. CONCLUSION: We do not recommend routine interval appendectomy in patients who benefit from conservative treatment for an appendiceal mass unless recurrent appendicitis develops. PMID- 15286891 TI - Our experience with telemedicine in traumatology and orthopedics. AB - BACKGROUND: Telemedicine widely takes root in all branches of modern medicine including traumatology and orthopedics. The main goal of this work was to present our experience with asynchronic teleconsultation in daily clinical practice, in particular in the treatment of polytrauma patients. METHODS: Throughout 2000 and 2003, we carried out 144 teleconsultations for 92 men and 52 women (age range three months to 80 years). Of these, we were the inquiring party in 51 cases, the consulting one in 88 cases, and the mediator in five cases. Time passed till the completion of consultations ranged from 12 to 24 hours. RESULTS: The number of consultants was one, two, three, and more in 99, 22, 3, and 15 teleconsultations, respectively. The most common questions (n=128) were those of treatment tactics. In the majority of cases, the consultant approved of the diagnosis suggested by the inquirer and formulated or corrected the scheme of the treatment. The majority of teleconsultations were concerned with various problems of traumatology (n=83) and orthopedics (n=31). For each clinical case, we received a mean of 2.6 replies (range 1 to 8). The effectiveness of the suggested treatment methods accounted for approximately 80% in final decision making. Teleconsultations provided considerable benefits in the treatment of polytrauma patients, including decreases in in-hospital treatment necessities (16%), in complication rates (9.2%) and their severity, the relative risk of developing complications (10%), and in the need for re-hospitalization (0.4%). CONCLUSION: In view of our experience, we recommend that asynchronic consultations on the basis of the Internet-technology be more commonly used in the treatment of polytrauma patients. PMID- 15286892 TI - [An evaluation of the relationship between blood glucose level and prognosis in acute phase of head injuries]. AB - BACKGROUND: We evaluated the relationship between blood glucose level and Glasgow Coma Scale (GCS) and prognosis in acute phase of head injuries. METHODS: The study included 62 patients (26 females, 36 males; mean age 61 years; range 4 to 76 years) who were admitted within the first three hours following isolated head injuries. Initial GCS scores were determined and blood glucose levels were measured. Cranial computed tomography (CT) findings were classified as cerebral edema, fracture, contusion, hemorrhage, and multiple pathologies. Relationships were sought between GCS scores, outcome, CT findings, and blood glucose levels. RESULTS: Cranial CT findings were cerebral edema in 15 patients (24.2%), hemorrhage in nine patients (14.5%), fractures in four patients (6.5%), contusion in two patients (3.2%), and multiple injuries in 32 patients (51.6%). The mean GCS score was 8 (range 3 to 13), which was 8 or lower in 37 patients (59.7%), and above 8 in 25 patients (40.3%). The mean blood glucose level was 219 mg/dl (range 136 to 397 mg/dl) in the study group. It was 293 mg/dl in those with a GCS score of 8 or lower, of which 18 patients died. Significant inverse relationships were found between (i) blood glucose level and GCS scores (p<0.01), and (ii) GCS scores and mortality (p<0.01). Blood glucose levels were significantly correlated with mortality (p<0.01). Computed tomography findings were not correlated with mortality and blood glucose levels. CONCLUSION: High blood glucose levels in acute phase of head injuries may be a sign for poor prognosis. PMID- 15286893 TI - [Successful results with the use of a new reconstruction material for soft tissue defects of the lower extremity: neurocutaneous flaps]. AB - BACKGROUND: We evaluated the use of distally based neurocutaneous flaps in the reconstruction of soft tissue defects in the lower extremity caused by trauma or other reasons. METHODS: Sixty-six patients (49 males, 17 females; mean age 42 years; range 16 to 75 years) underwent reconstruction of lower extremity soft tissue defects with the use of neurocutaneous flaps. The defects were in the external (n=11) and internal malleolus (n=10), Achilles tendon (n=6), heel (n=11), foot (n=7), distal (n=9) and middle (n=7) third of the anterior tibia, the mid-plantar region (n=3), and the popliteal region (n=2). Sural and saphenous neurocutaneous flaps were used in 34 and 32 patients, respectively. The mean flap area was 135 square centimetres (range 54 to 286). The mean follow-up period was 32 months (range 11 to 58 months). RESULTS: Of sixty-six patients treated with neurocutaneous flaps, complete survival was achieved in 57 flaps (86.4%). Partial necrosis was encountered in six patients (9%). Total flap loss occurred in three patients (4.6%) who had diabetes, hypertension, and electrical burns, respectively. The most common complication during the postoperative period was venous congestion seen in 14 patients (21%), the majority of whom had diabetes (n=5), hypertension (n=3), and electrical burns (n=3), predisposing them to microcirculation disorders. This complication was successfully managed by leech therapy in 78.6%. CONCLUSION: Neurocutaneous flaps offer maximum reconstruction capacity with minimum donor site morbidity, making them a successful alternative in the treatment of soft tissue defects of the lower extremity in recent years. PMID- 15286894 TI - [Urgent partial sternotomy for the treatment of iatrogenic vascular injury to the thoracic outlet: a report of two cases]. AB - Two patients underwent surgery for iatrogenic vascular injuries at the thoracic outlet. Injuries occurred during mediastinoscopy in one patient (male, aged 60 years) and removal of a Portovac catheter in the other (female, aged 49 years). We performed partial sternotomy in both cases to expose the proximal of a. carotis communis and vena cava superior, respectively. After distal and proximal control of the vessels, the injured vessel was primarily sutured in the first patient, and the tip of the catheter was removed through a venotomy incision in the latter. No postoperative complications occurred in both patients. Partial sternotomy incision provides an appropriate and quick access to the injured vessels and surrounding structures at the thoracic outlet. PMID- 15286895 TI - [A case of superior sagittal sinus thrombosis after closed head injury]. AB - Superior sagittal sinus thrombosis (SSST) is a rare entity, most often arising from infections, dehydration, and hematologic disorders. Development of this condition secondary to trauma is extremely rare. In this report, a 13-year-old boy who developed SSST following a closed head injury is presented. Imaging studies showed SSST caused by a depressed skull fracture. Neurologic examination of the patient was normal other than bilateral papillary stasis. He was treated with antiedematous and anticonvulsant drugs. Magnetic resonance venography obtained eight months after the diagnosis showed unoccluded superior sagittal sinus, neurologic examination findings were normal, as well. PMID- 15286896 TI - [Radial artery pellet embolism: a case report]. AB - A fifty-four-year-old male patient presented to our emergency department with a gunshot injury in his arm, caused by pellets. Surgical exploration showed injury to the brachial artery, which was then repaired with a saphenous vein graft. Conservative treatment was planned for associated nerve injuries. After arterial flow began, two pellet-like solid bodies were palpated in the radial artery trace at the wrist level and the pellets were removed from the lumen of the artery. Ten days after surgery, the patient had profuse bleeding in the arm. On exploration, partial necrosis was detected in the saphenous vein graft and primary repair was performed. However, on the 19th day, the bleeding recurred and increased necrosis and rupture of the artery, 2-3 cm in size, were detected. Arterial repair was repeated with another saphenous vein graft from the other limb. The patient returned to normal daily activities nine months after the injury, with slight cold intolerance. Clinical examination and Doppler studies did not show any signs of vascular deficiency. Arterial or venous pellet embolism should be included in the evaluation of patients with gunshot injuries. PMID- 15286898 TI - [Are we Spanish ophthalmologists good scientists? (Wisen up. [you jerk]... or they'll eat us for breakfast!)]. PMID- 15286899 TI - [The use of Xalacom and deterioration in cases of vitiligo. Is there a causal relationship?]. PMID- 15286900 TI - [Changes in axial length/corneal radius ratio (AL/CR) according to refractive state of the eye. Relationship with ocular components]. AB - PURPOSE: Determination of the role of the axial length/corneal radius ratio (AL/CR) in the refractive state and investigation of its relationship with the ocular optical components: AL, CR, anterior chamber depth (ACD), crystalline lens thickness (CT) and vitreous chamber depth (VCD). METHODS: The RE (right eye) of 193 University students 22.27 (SD 3.24) years, with different refractive errors (spherical equivalent range: +3.00 D to -11.00 D), being divided into: emmetropes, hyperopes and myopes (low, moderate and high). The ACE, the CT, the VCD and the AL were measured by ultrasonography (unidimensional echography); and the mean CR by videokeratoscopy. RESULTS: The value of AL/CR obtained was 2.98 (SD 0.69) for emmetropes, 2.89 (SD 0.87) for hyperopes, 3.01 (SD 0.07) in low myopias, 3.10 (SD 0.11) in moderate myopias and 3.23 (SD 0.12) in high myopias. The AL/CR ratio showed a higher correlation with the refractive error. Besides, all the refractive groups were observed to have lower CE values as the AL/CR increased. This tendency is statistically significant in hyperopes, emmetropes and low myopes; and is not in moderate and high myopias. All the study groups could be observed to have a positive and statistically significant correlation between AL/CR and ACD. CONCLUSION: The ratio AL/CR is the most important parameter and the best predictor of the refractive state of the human eye. It provides important information on how best to determine the degree of emmetropization given by the crystalline, decreasing its power and the ACD in concordance with the LA. A value for the ratio AL/CR above 3.00 could be considered as a risk factor for the development of myopia in emmetropic eyes. PMID- 15286901 TI - [Transcanalicular dacryocystorhinostomy technique using diode laser]. AB - PURPOSE: To describe the surgical technique and to evaluate the clinical results of carrying out transcanalicular dacryocystorhinostomies using a diode laser (DCR TC), including the advantages and limitations of this technique. METHODS: 43 DCR TC were studied and they were analysed using a prospective, interventional, non randomized and non comparative study. We used local and topical anaesthesia in patients with a clinical history of epiphora or dacryocystitis for nasolacrimal obstruction. A diode laser was used to effect a vaporization of the lacrimal sac, osteotomy and vaporization with coagulation of nasal mucosa. The mean duration of surgery was 14 minutes (range 7 to 29 minutes). In all cases, and during a two month period, bicanalicular intubation was carried out using a silicone tube and prolene filament. Follow-up was between 4 to 38 months. The degree of epiphora was evaluated using the Munk scale and lacrimal permeability was evaluated using Jones I and II tests by direct videoendoscopic control in all cases. RESULTS: 43 DCR-TC were realized, 39 cases are asymptomatic (90.7%). 2 of them (4.65%) had epiphora (degree 2 on the Munk scale) and permeable tract. One case (2.32%) presented lower canaliculi obstruction. One patient showed total osteotomy closing. CONCLUSIONS: Transcanalicular dacryocystorhinostomy carried out using a diode laser is a useful method because it does not cause cutaneous scarring, it hardly produces pain and bleeding, it respects the lacrimal pump, it needs less surgical time, it can be carried out in an out-patient surgery and it generates minimal intra and postsurgical morbility. PMID- 15286902 TI - [Corneal graft data collection: evaluation of results 1995-2000]. AB - PURPOSE: To examine the collection of data from the corneal grafts performed between 1995 and 2000, thus providing a long term follow-up. METHODS: Records of corneal grafts were made according to the following design and divided into three basic groups: 1a) donor data: age, storage time, endothelial density, ABO group; 1b) recipient data: age, ABO group, sight threatening pathology, diagnosis, visual acuity, corneal oedema, inflammation, vascularisation; 2) surgery data: surgery type, graft size, suture, intraocular lens, vitrectomy, complications; 3) data at first month and at each year follow-up: visual acuity, astigmatism, endothelial density, graft rejection and complications. A retrospective study of data collection percentage was performed. RESULTS: Although 100% of data from donors and recipients was collected, some data such as ABO group was not available in all cases. First indication was keratoconus and bullous keratopathy. Mean visual acuity at first year was 0.31, collected in 60% and endothelial density, 2,223 cls/mm2, available in 26.6 percent. CONCLUSIONS: A good basic data collection improves the long-term follow-up for corneal grafts. Further analysis in the future of this data will be helpful for clinical audit. PMID- 15286903 TI - [Granulocyte-apheresis in ocular Behcet's disease resistant to medical treatment]. AB - OBJECTIVE: To assess the feasibility of granulocyte apheresis (GCAP) in patients with Ocular Behcet's Disease (BD) resistant to immunosuppressive therapy (prednisone, cyclosporin, azathyoprine or mycophenolate mofetil). METHODS: Prospective observational study of five cases, carried out in a university centre. Four patients were resistant to medical treatment and one refused immunosuppressive drugs. The intervention procedure consisted of an extracorporeal GCAP using a column filled with cellulose acetate beads (Adacolumn, JIMRO, Takasaki, Japan). All patients received underwent a schedule of therapy of five sessions, once a week for five consecutive weeks. Visual acuity (Snellen lines), the degree of intraocular inflammation and doses of immunosuppressive therapy were measured and observed every week. RESULTS: Visual acuity improved in the five cases. Intraocular inflammation was measured and observed in every case, relapses were avoided, and treatment with prednisone was reduced by more than half of the initial dose (average reduction 52.7%, SD 14). CONCLUSIONS: GCAP has been shown to be safe and effective as a new therapy in five cases of Ocular Behcet's Disease refractory to medical treatment. Further research is needed in order to confirm the promising results of these initial investigations. PMID- 15286904 TI - [Glaucoma drainage device obstruction]. AB - PURPOSE: To determine the pathophysiology and to discuss possible solutions for tube obstruction since this is a frequent complication in glaucoma drainage device surgery. MATERIAL AND METHODS: 81 eyes with a glaucoma drainage device (GDD) fitted by our Department during the last twelve years (1991-2002) were retrospectively reviewed and eleven cases of tube obstruction were detected. The treatments included Nd-Yag laser, anterior vitrectomy, blood clot removal and intracamerular rTPA. RESULTS: The obstruction was proximal in ten cases and distal in one. Of the former, the obstruction was caused by a blood clot in six cases, by vitreous in three cases and by a fibrinous membrane in one case. The nature of the distal obstruction was not determined. One case of vitreous obstruction occurred after a posterior capsulotomy with Nd-Yag laser two years after positioning the GDD. Complete success was achieved in five cases, and partial success in three where antiglaucoma therapy had to be maintained postoperatively. Complete failure was found in the other three cases. CONCLUSIONS: The material which most frequently caused the tube obstruction was blood and although in some cases this may resolve spontaneously, clot removal with forceps or rTPA intracameral injection may be helpful. In vitreous obstruction, Nd-Yag laser may temporariril clear the tube, but extensive anterior vitrectomy is required to maintain long-term permeability. We suggest carrying out a small posterior capsulotomy in pseudophakic patients with a GDD. PMID- 15286905 TI - [Adventitial sheathotomy in branch retinal vein occlusion with non ischemic macular edema]. AB - PURPOSE: To determine whether surgical decompression of branch retinal vein occlusion at the arterio-venous (A/V) crossing is effective as a treatment for associated non ischemic macular edema. METHOD: A total of 13 consecutive patients with non ischemic macular edema secondary to branch retinal vein occlusion were treated in our clinic with A/V decompression: pars plana vitrectomy and an arteriovenous adventitial sheathotomy. An internal limiting membrane (ILM) maculorhexis was additionally performed in five patients. We assessed visual acuity, fundus biomicroscopy, and fluorescein angio-graphy preoperatively and after surgery. RESULTS: The surgical procedure and postoperative period were uneventful. Postoperative visual acuities improved in 12 patients (92%) at twelve months. Functional results in patients with internal limiting membrane (ILM) maculorhexis were significantly improved and in a shorter time span. CONCLUSIONS: Our results with surgical decompression of branch retinal vein occlusions describe the therapeutical effect of surgical A/V decompression in treatment of macular non ischemic macular edema secondary to branch retinal vein occlusion, and the additional performance of internal limiting membrane maculorhexis may improve visual prognosis. Whether sheathotomy is beneficial in the treatment of non ischemic macular edema has to be investigated further. PMID- 15286906 TI - [Amaurosis after lachrymo-nasal stent surgery]. AB - CLINICAL CASE: A 74 year old woman with lachrymo nasal stent introduced three days before referring periocular pain and loss of visual acuity in left eye. With aggressive treatment the inflammatory illness improved but not the visual acuity (no light perception). The lachrymal stent was removed and, later, the lachrymal sack. DISCUSSION: Orbital cellulitis is an inflammation of fat and intraorbital structures that needs aggressive treatment in order to prevent serious complications. PMID- 15286907 TI - [Acute and bilateral uveitis secondary to moxifloxacin]. AB - CASE REPORT: We report a case of a 77-year-old woman with an anterior acute bilateral uveitis and pigmentary dispersion after treatment of a neumococic pneumonia with moxifloxacin. Her response to treatment was good but she presented as a result of treatment abundance of pigment over all the structures of the anterior segment. DISCUSSION: Many drugs can produce uveitis. After Med-line revision, we can not find any previously reported case of anterior Uveitis associated with fourth generation quinolones. PMID- 15286908 TI - [Myths regarding the origin of eyeglasses (II)]. PMID- 15286910 TI - Kidney mass and relative medullary thickness of rodents in relation to habitat, body size, and phylogeny. AB - We tested the hypotheses that relative medullary thickness (RMT) and kidney mass are positively related to habitat aridity in rodents, after controlling for correlations with body mass. Body mass, mass-corrected kidney mass, mass corrected RMT, mass-corrected maximum urine concentration, and habitat (scored on a semiquantitative scale of 1-4 to indicate increasing aridity) all showed statistically significant phylogenetic signal. Body mass varied significantly among habitats, with the main difference being that aquatic species are larger than those from other habitats. Mass-corrected RMT and urine concentration showed a significant positive correlation (N=38; conventional r=0.649, phylogenetically independent contrasts [IC] r=0.685), thus validating RMT as a comparative index of urine concentrating ability. RMT scaled with body mass to an exponent significantly less than 0 (N=141 species; conventional allometric slope=-0.145 [95% confidence interval (CI)=-0.172, -0.117], IC allometric slope=-0.132 [95% CI=-0.180, -0.083]). Kidney mass scaled to an exponent significantly less than unity (N=104 species; conventional slope=0.809 [95% CI=0.751, 0.868], IC slope=0.773 [95% CI=0.676, 0.871]). Both conventional and phylogenetic analysis indicated that RMT varied among habitats, with rodents from arid areas having the largest values of RMT. A phylogenetic analysis indicated that mass-corrected kidney mass was positively related to habitat aridity. PMID- 15286911 TI - Differences in isolated mitochondria are insufficient to account for respiratory depression during diapause in artemia franciscana embryos. AB - In response to cues signifying the approach of winter, adult Artemia franciscana produce encysted embryos that enter diapause. We show that respiration rates of diapause embryos collected from the field (Great Salt Lake, Utah) are reduced up to 92% compared with postdiapause embryos when measured under conditions of normoxia and full hydration. However, mitochondria isolated from diapause embryos exhibit rates of state 3 and state 4 respiration on pyruvate that are equivalent to those from postdiapause embryos with active metabolism; a reduction in these rates (15%-27%) is measured with succinate for two of three collection years. Respiratory control ratios for diapause mitochondria are comparable to or higher than those from postdiapause embryos. The P : O flux ratios are statistically identical. Our calculations suggest that respiration of intact, postdiapause embryos is operating close to the state 3 oxygen fluxes measured for isolated mitochondria. Cytochrome c oxidase (COX) activity is 53% lower in diapause mitochondria during one collection year; the minimal impact of this COX reduction on mitochondrial respiration appears to be due to the 31% excess COX capacity in A. franciscana mitochondria. Transmission electron micrographs of embryos reveal mitochondria that are well differentiated and structurally similar in both states. As inferred from the similar amounts of mitochondrial protein extractable, tissue contents of mitochondria in diapause and postdiapause embryos are equivalent. Thus, metabolic depression during diapause cannot be fully explained by altered properties of isolated mitochondria. Rather, mechanisms for active inhibition or substrate limitation of mitochondrial metabolism in vivo may be operative. PMID- 15286912 TI - Oxygen uptake and local Po2 profiles in submerged larvae of phaeoxantha klugii (Coleoptera: Cicindelidae), as well as their metabolic rate in air. AB - We studied whether oxygen uptake from the surrounding water might enhance survival in submerged third instar larvae of Phaeoxantha klugii, a tiger beetle from the central Amazonian floodplains. Local oxygen partial pressures (Po(2)) were measured with microcoaxial needle electrodes close to larvae submerged in initially air-saturated still water. The Po(2) profiles showed that the larvae exploit oxygen from the aquatic medium. Metabolism in the air of more or less resting larvae was determined by measuring the rate of CO(2) production (sV dot co2) with an infrared gas analyzer at 29 degrees C. The sV dot co2 was around 1.8 mu L g(-1) min(-1), equivalent to an oxygen consumption rate (sV dot o2) of 1.8 2.6 mu L g(-1) min(-1). Oxygen consumption (V dot o2) of individually submerged larvae measured in closed respiration chambers at 19-10.3 kPa Po(2) (initially air saturated, 29 degrees C) ranged between 0.05 and 0.2 mu L min(-1) and was not correlated with body mass. The sV dot o2 ranged between 0.1 and 0.4 mu L min(-1), that is, 4%-22% of the metabolic rate measured in air. Mean V dot o2 decreased with declining Po(2); however, some individuals showed contrary patterns. V dot o2 was additionally measured in dormant larvae, in larvae submerged for 1-2 d in open water or for 30-49 d within sediment, as well as in larvae exposed to anoxia before the measurements. The range of V dot o2 was similar in all groups, indicating that the larvae exploit oxygen from the water whenever available. Similar V dot o2 across the whole range of body mass investigated (0.31-0.76 g) suggests that oxygen uptake occurs by spiracular uptake. Assuming that larvae survive for some time at rates comparable to depressed metabolic rates reported for other insect species, it can be concluded that oxygen uptake from water can sustain aerobic metabolism even under quite severe hypoxia. It might therefore play an important role for survival during inundation periods. PMID- 15286913 TI - Strategies for surviving high concentrations of environmental ammonia in the swamp eel Monopterus albus. AB - The swamp eel Monopterus albus lives in muddy ponds, swamps, canals, and rice fields in the tropics. It encounters high concentrations of environmental ammonia (HEA) during dry seasons or during agricultural fertilization in rice fields. This study aimed at determining the tolerance of M. albus to environmental ammonia and at elucidating the strategies that it adopts to defend against ammonia toxicity in HEA. In the laboratory, M. albus exhibited very high environmental ammonia tolerance; the 48-, 72-, and 96-h median lethal concentrations of total ammonia at pH 7.0 and 28 degrees C were 209.9, 198.7, and 193.2 mM, respectively. It was apparently incapable of actively excreting ammonia against a concentration gradient. In addition, it did not detoxify ammonia to urea, the excretion of which would lead to a loss of nitrogen and carbon, during ammonia loading. The high tolerance of M. albus to HEA was attributable partially to its exceptionally high tolerance to ammonia at the cellular and subcellular levels. During the 144 h of exposure to 75 mM NH(4)Cl at pH 7.0, the ammonia contents in the muscle, liver, brain, and gut of M. albus reached 11.49, 15.18, 6.48, and 7.51 mu mol g(-1), respectively. Such a capability allowed the accumulation of high concentrations of ammonia in the plasma (3.54 mu mol mL(-1)) of M. albus exposed to HEA, which would reduce the net influx of exogenous ammonia. Subsequent to the buildup of internal ammonia levels, M. albus detoxified ammonia produced endogenously to glutamine. The glutamine contents in the muscle and liver reached 10.84 and 17.06 mu mol g(-1), respectively, after 144 h of exposure to HEA, which happened to be the highest known for fish. Unlike urea, the storage of glutamine in the muscle during ammonia loading allowed its usage for anabolic purposes when the adverse environmental condition subsides. Glutamine synthetase activity increased significantly in the liver and gut (2.8- and 1.5-fold, respectively) of specimens exposed to HEA for 144 h. These results suggest that the liver was the main site of ammonia detoxification and the gut was more than a digestive/absorptive organ in M. albus. Monopterus albus did not undergo a reduction in amino acid catabolism during the first 24 h of ammonia exposure. However, assuming a total inhibition of excretion of endogenous ammonia, there was a deficit of -312 mu mol N between the reduction in nitrogenous excretion (3,360 mu mol N) and the retention of nitrogen (3,048 mu mol N) after 72 h of aerial exposure. The deficit became much greater after 144 h, reaching a value of -3,243 mu mol N. These results suggest that endogenous ammonia production in M. albus was suppressed in order to prevent the newly established internal steady state concentration of ammonia from rising to an intolerable level after an extended period of exposure to HEA. PMID- 15286915 TI - Freeze resistance in rainbow smelt (Osmerus mordax): seasonal pattern of glycerol and antifreeze protein levels and liver enzyme activity associated with glycerol production. AB - Rainbow smelt (Osmerus mordax) inhabit inshore waters along the North American Atlantic coast. During the winter, these waters are frequently ice covered and can reach temperatures as low as -1.9 degrees C. To prevent freezing, smelt accumulate high levels of glycerol, which lower the freezing point via colligative means, and antifreeze proteins (AFP). The up-regulation of the antifreeze response (both glycerol and AFP) occurs in early fall, when water temperatures are 5 degrees -6 degrees C. The accumulation of glycerol appears to be the main mechanism of freeze resistance in smelt because it contributes more to the lowering of the body's freezing point than the activity of the AFP (0.5 degrees C vs. 0.25 degrees C for glycerol and AFP, respectively) at a water temperature of -1.5 degrees C. Moreover, AFP in smelt appears to be a safeguard mechanism to prevent freezing when glycerol levels are low. Significant increases in activities of the liver enzymes glycerol 3-phosphate dehydrogenase (GPDH), alanine aminotransferase (AlaAT), and phosphoenolpyruvate carboxykinase (PEPCK) during the initiation of glycerol production and significant correlations between enzyme activities and plasma glycerol levels suggest that these enzymes are closely associated with the synthesis and maintenance of elevated glycerol levels for use as an antifreeze. These findings add further support to the concept that carbon for glycerol is derived from amino acids. PMID- 15286914 TI - Cl- uptake mechanism in freshwater-adapted tilapia (Oreochromis mossambicus). AB - In this study, the correlation between Cl(-) influx in freshwater tilapia and various transporters or enzymes, the Cl(-)/HCO(3)(-) exchanger, Na(+),K(+) ATPase, V-type H(+)-ATPase, and carbonic anhydrase were examined. The inhibitors 2x10(-4) M ouabain (a Na(+),K(+)-ATPase inhibitor), 10(-5) M NEM (a V-type H(+) ATPase inhibitor), 10(-2) M ACTZ (acetazolamide, a carbonic anhydrase inhibitor), and 6x10(-4) M DIDS (a Cl(-)/HCO(3)(-) exchanger inhibitor) caused 40%, 60%-80%, 40%-60%, and 40%-60% reduction in Cl(-) influx of freshwater tilapia, respectively. The inhibitor 2x10(-4) M ouabain also caused 50%-65% inhibition in gill Na(+),K(+)-ATPase activity. Western blot results showed that protein levels of gill Na(+),K(+)-ATPase, V-type H(+)-ATPase, and carbonic anhydrase in tilapia acclimated in low-Cl(-) freshwater were significantly higher than those acclimated to high-Cl(-) freshwater. Based on these data, we conclude that Na(+),K(+)-ATPase, V-H(+)-ATPase, the Cl(-)/HCO(3)(-) exchanger, and carbonic anhydrase may be involved in the active Cl(-) uptake mechanism in gills of freshwater-adapted tilapia. PMID- 15286916 TI - Biotic and abiotic factors affect the nest environment of embryonic leatherback turtles, Dermochelys coriacea. AB - Clutches of leatherback turtles, Dermochelys coriacea, have lower hatching success than those of other sea turtles, but causes of high embryonic mortality are unknown. We measured characteristics of clutches along with spatial and temporal changes in PO(2) and temperature during incubation to determine the extent to which they affected the developmental environment of leatherback embryos. Minimum PO(2) in nests decreased as both the total number and mass of metabolizing embryos increased. Increases in both the number and mass of metabolizing embryos caused an increase in maximum nest temperature. However, neither PO(2) nor temperature was correlated with hatching success. Our measurements of relatively high nest PO(2) (lowest 17.1 kPa or 16.9% O(2)) indicate that hypoxia apparently does not cause the low hatching success of leatherback clutches. Oxygen partial pressure increased and temperature decreased from the center toward the periphery of leatherback nests. We inferred from these measurements that positions of eggs within nests vary in quality and potentially affect overall developmental success of entire clutches. The large metabolic mass of leatherback clutches and limits to gas flux imposed by the sand create a situation in which leatherback embryos collectively affect their own environment. PMID- 15286918 TI - Maternal effects on offspring locomotion: influence of density and corticosterone elevation in the lizard Lacerta vivipara. AB - Offspring phenotype can be affected by maternal history before and during gestation. Offspring sensitivity to maternal conditions is believed to have evolved to favor preadaptation of offspring to environmental factors they are likely to encounter. Because the locomotor capacity of an individual is likely to have important fitness consequences, we examined the role of long-term and short term prenatal conditions on offspring's locomotor performance in the lizard Lacerta vivipara. To examine long-term prenatal effects, we manipulated the density of two populations, leaving two additional populations as unmanipulated. We then collected pregnant females within these four populations (Cevennes, Massif Central, France) and kept them in the laboratory until parturition. To examine short-term prenatal effects, we manipulated the corticosterone level of half the females within each population. We took two different measurements of offspring locomotion: sprint speed and endurance. As already documented, sprint speed was positively correlated with offspring body size. Although population density significantly affected female fecundity, neither the density manipulation nor the population of origin influenced offspring phenotype. Corticosterone administered during gestation decreased juvenile sprint speed but did not affect juvenile endurance. Furthermore, we observed that the motivation to run was influenced by maternal hormonal treatment. Juveniles born from corticosterone treated mothers needed more stimuli than those born from control mothers. We conclude, therefore, that the action of corticosterone on sprint speed could be more behavioral than physiological. Offspring phenotype as measured by endurance and sprint speed appeared partly under maternal control. PMID- 15286917 TI - Aerobic performance of wild-derived house mice does not change with cold exposure or intestinal parasite infection. AB - Aerobic performance is affected by numerous endogenous and exogenous factors. We investigated the effects of ambient temperature and parasite infection on resting metabolism and maximal exercise-induced oxygen consumption in wild-derived house mice (Mus musculus). We also collected preliminary data for effects of lactation on these measures of aerobic performance. Mice were experimentally infected with a naturally occurring intestinal nematode (Heligmosomoides polygyrus) and then exposed to cold temperatures for 10 d or allowed to mate and reproduce. Wild derived house mice did not change their resting metabolism with H. polygyrus infection or cold exposure, which is in stark contrast to similar studies with laboratory mice. Preliminary data also showed no effect of lactation on aerobic performance. Similarly, maximal exercise-induced oxygen consumption and hematocrit and hemoglobin were unaffected by all experimental treatments. We conclude that resting metabolism, maximal oxygen consumption, and hematology of wild-derived house mice are unaffected by exogenous (temperature) and endogenous (H. polygyrus) demands and, therefore, wild-derived mice respond to these demands without incurring potential costs associated with changes in aerobic performance. PMID- 15286919 TI - Ontogenetic variation in digestion by the herbivorous lizard Ctenosaura pectinata. AB - I tested the hypothesis that an animal with an ontogenetic diet shift must have different digestive efficiencies for foods that correspond to its diet shift, so that nutrient and energy extraction are maximized. The iguanine lizard Ctenosaura pectinata undergoes an ontogenetic diet shift from eating insects as a juvenile to plants as an adult. When fed six different pure foods from the natural diets of different age classes, C. pectinata assimilated nutrients and energy differently depending on food type and age class. Extraction of energy and nutrients in insect larvae was maximized by juvenile lizards. Calcium, phosphorus, and energy were readily assimilated from flowers and fruit by immature and adult lizards. Magnesium levels were highest in leaves and were extracted by immature and adult lizards, but xenobiotic effects of one plant leaf (Croton suberosus), eaten by adults, killed juvenile lizards. Although juvenile C. pectinata ate some flowers (Senna wislizenii) naturally, they were less efficient at digesting cell walls from these plant parts than were older lizards. Ontogenetic changes in ctenosaur digestive physiology were not the result of a trade-off involving ecological costs of different foods; rather, each age class preferred a diet that maximized its physiological benefit. PMID- 15286920 TI - Diet quality and food limitation affect the dynamics of body composition and digestive organs in a migratory songbird (Zonotrichia albicollis). AB - Migrating songbirds interrupt their feeding to fly between stopover sites that may vary appreciably in diet quality. We studied the effects of fasting and food restriction on body composition and digestive organs in a migratory songbird and how these effects interacted with diet quality to influence the rate of recovery of nutrient reserves. Food limitation caused white-throated sparrows to reduce both lean and fat reserves, with about 20% of the decline in lean mass represented by a decline in stomach, small intestine, and liver. During refeeding on diets similar in nutrient composition to either grain or fruit, food-limited grain-fed birds ate 40% more than did control birds, and they regained body mass, with on average 60% of the increase in body mass composed of lean mass including digestive organs. In contrast, food-limited fruit-fed birds did not eat more than did control birds and did not regain body mass, suggesting that a digestive constraint limited their food intake. The interacting effects of food limitation and diet quality on the dynamics of body composition and digestive organs in sparrows suggest that the adequacy of the diet at stopover sites can directly influence the rate of recovery of body reserves in migrating songbirds and hence the pace of their migration. PMID- 15286921 TI - Differential accumulation and pigmenting ability of dietary carotenoids in colorful finches. AB - Many animals develop bright red, orange, or yellow carotenoid pigmentation that they use to attract mates. Colorful carotenoid pigments are acquired from the diet and are either directly incorporated as integumentary colorants or metabolized into other forms before deposition. Because animals often obtain several different carotenoids from plant and animal food sources, it is possible that these pigments are accumulated at different levels in the body and may play unique roles in shaping the ultimate color expression of individuals. We studied patterns of carotenoid accumulation and integumentary pigmentation in two colorful finch species--the American goldfinch (Carduelis tristis) and the zebra finch (Taeniopygia guttata). Both species acquire two main hydroxycarotenoids, lutein and zeaxanthin, from their seed diet but transform these into a series of metabolites that are used as colorful pigments in the plumage (goldfinches only) and beak (both species). We conducted a series of carotenoid-supplementation experiments to investigate the relative extent to which lutein and zeaxanthin are accumulated in blood and increase carotenoid coloration in feathers and bare parts. First, we supplemented the diets of both species with either lutein or zeaxanthin and measured plasma pigment status, feather carotenoid concentration (goldfinches only), and integumentary color. Zeaxanthin-supplemented males grew more colorful feathers and beaks than lutein-supplemented males, and in goldfinches incorporated a different ratio of carotenoids in feathers (favoring the accumulation of canary xanthophyll B). We also fed goldfinches different concentrations of a standard lutein-zeaxanthin mix and found that at physiologically normal and high concentrations, birds circulated proportionally more zeaxanthin over lutein than occurred in the diet. Collectively, these results demonstrate that zeaxanthin is preferentially accumulated in the body and serves as a more potent substrate for pigmentation than lutein in these finches. PMID- 15286922 TI - Physiological ecology of Mediterranean blue tits (Parus caeruleus L.): effects of ectoparasites (Protocalliphora spp.) and food abundance on metabolic capacity of nestlings. AB - The consequences of nest ectoparasites, such as Protocalliphora larvae, on nestling birds have been the subject of numerous studies. Despite observed reductions in mass and hematocrit of chicks from parasitized nests, no studies have found any effect of Protocalliphora on nestling survival, suggesting that fitness consequences of Protocalliphora are either weak or occur after fledging. From experiments on the metabolic performance of chicks, we found that parasitized chicks suffer from reduced thermogenic and metabolic capacities as a result of decreased mass and hematocrit. Hence, Protocalliphora may potentially affect nestling survival after fledging, when energetically costly activities such as flight and moult are undertaken. Previous studies have demonstrated an increase in parental feeding rate to compensate for the detrimental consequences of parasite infestation. We tested whether parasite effects on nestling aerobic capacity were dependent on food availability during the feeding period. Measures of caterpillar densities and experimental manipulations of parasite loads allowed us to investigate relationships among host, parasite, and environment. A positive relationship between chick aerobic and thermogenic performances and caterpillar density suggests that negative effects of parasitism may be offset by increased food availability. This study provides the first measurement of the effects of an ectoparasite on metabolic competence in wild birds and documentation of the effect of food availability on ectoparasite virulence using a quantitative measure of food abundance. PMID- 15286923 TI - The allometry of avian basal metabolic rate: good predictions need good data. AB - Basal metabolic rate (BMR) is often predicted by allometric interpolation, but such predictions are critically dependent on the quality of the data used to derive allometric equations relating BMR to body mass (Mb). An examination of the metabolic rates used to produce conventional and phylogenetically independent allometries for avian BMR in a recent analysis revealed that only 67 of 248 data unambiguously met the criteria for BMR and had sample sizes with n>/=3. The metabolic rates that represented BMR were significantly lower than those that did not meet the criteria for BMR or were measured under unspecified conditions. Moreover, our conventional allometric estimates of BMR (W; logBMR= 1.461+0.669logMb) using a more constrained data set that met the conditions that define BMR and had n>/=3 were 10%-12% lower than those obtained in the earlier analysis. The inclusion of data that do not represent BMR results in the overestimation of predicted BMR and can potentially lead to incorrect conclusions concerning metabolic adaptation. Our analyses using a data set that included only BMR with n>/=3 were consistent with the conclusion that BMR does not differ between passerine and nonpasserine birds after taking phylogeny into account. With an increased focus on data mining and synthetic analyses, our study suggests that a thorough knowledge of how data sets are generated and the underlying constraints on their interpretation is a necessary prerequisite for such exercises. PMID- 15286924 TI - Measuring fat mass in small birds by dual-energy x-ray absorptiometry. PMID- 15286925 TI - Bone alkaline phosphatase as a sensitive indicator of skeletal development in birds: a study of the great tit nestlings. PMID- 15286926 TI - Twelve-lead electrocardiogram: the advantages of an orderly frontal lead display including lead -aVR. AB - BACKGROUND: It is possible that efforts in ECG review by both young experienced clinicians are currently discouraged-and risk to be completely dismissed-by the conventional (ie, disorderly) display of the frontal plane leads, with lead aVR at -150 degrees. METHODS: We reviewed studies on the usefulness of leads aVR and aVR as well as on the history of the frontal leads in electrocardiography. RESULTS: Lead aVR and particularly, lead -aVR, provide useful information when systematically analyzed. In addition, if lead -aVR is examined in its anatomically logical sequence, ie, aVL, I, -aVR, II, aVF, and III, the frontal plane of the 12-lead ECG is more easily understood. This "panoramic" or "orderly" display is in common use in countries such as Sweden, but it is rarely seen in the United States. CONCLUSIONS: ECG interpretation would be enhanced by displaying the limb leads in an orderly arrangement that starts with lead aVL and ends with lead III, and many ECG changes would be ideally displayed by a lead aVR at 30 degrees. PMID- 15286927 TI - Refinement and interobserver agreement for the electrocardiographic Sclarovsky Birnbaum Ischemia Grading System. AB - BACKGROUND: Electrocardiogram-derived grades of ischemia at the time of patient presentation with acute myocardial infarction have proved useful in predicting the salvageability by reperfusion therapy, final infarct size, severity of left ventricular dysfunction, and short- and long-term prognosis. SUBJECTS AND METHODS: The Sclarovsky-Birnbaum Ischemia Grading System based on the relation between the acute appearances of the T wave, the ST segment, and the QRS complex was considered as a means of enhanced ECG analysis in this group of patients. The evaluation of a training population (n = 46) resulted in refinement of the published description of the Sclarovsky-Birnbaum Ischemia Grading System, and a test population (n = 50) was utilized for investigating the interobserver agreement among 5 observers in determining the grade of ischemia. RESULTS: The agreement among the observers applying the "refined" Sclarovsky-Birnbaum Ischemia Grading System was 0.89. Complete agreement was found for the ECGs of 80% of the patients, and the most common reason for disagreement was the application of the terminal T-negativity criterion. CONCLUSIONS: The refined Sclarovsky-Birnbaum Ischemia Grading System can be performed manually with low interobserver variability. It has potential for support of the acute myocardial infarction triage decision as an electrocardiographic method for evaluating the level of ischemic protection at the time of either pre-hospital or emergency-department presentation. PMID- 15286928 TI - Reduced high-frequency QRS components in patients with ischemic heart disease compared to normal subjects. AB - Analysis of high-frequency QRS components (HF-QRS) might provide an additional method when diagnosing various heart diseases, for example ischemic heart disease (IHD). This study compares HF-QRS in normal subjects to those in patients with IHD, and also analyzes HF-QRS considering gender and age. A total of 63 normal subjects and 64 patients with IHD were included. Signal-averaged electrocardiograms (ECGs) from the 12 standard leads were analyzed in the frequency interval of 150-250 Hz. The results showed that the summed 12 lead HF QRS in patients with IHD were significantly lower than in normal subjects (mean summed HF-QRS was 33.5 microV in the IHD group, 43.7 microV in normal individuals, P <.0005). HF-QRS were not statistically associated with gender or age (P =.820 and P =.573, respectively). However, the inter-individual variation of HF-QRS was large in both groups which probably limits the clinical usefulness of the method. PMID- 15286929 TI - Repeatability of heart rate variability measures. AB - Due to the sparse data on the repeatability of short and ultra-short term heart rate variability (HRV) measures, we measured the repeatability of common HRV measures derived from 10-second, 2-minute, and 6-minute recordings in 63 healthy men and women, aged 45-64, in Chapel Hill, North Carolina. Three 10-second and 2 six-minute heart rate recordings were obtained during each of 2 visits, separated by 1 to 2 weeks. We partitioned the measurement error into components and computed intraclass correlation coefficients using nested, random effects models. Repeatability improved with the length of recording: intraclass correlation coefficients were greater than 0.7 for 6-minute measures and 2-minute time domain measures and greater than 0.5 for 2-minute frequency domain measures. Repeatability of measures from 10-second records was lower, but improved considerably when the mean from 2 or 3 records was used. Correlations between the same measures from different length recordings were quite high. Our findings support the use of records of at least 5 minutes in length in epidemiological studies, in accordance with previous guidelines. Researchers using 10-second records should consider taking the mean of several recordings, when possible, or using statistical methods to correct for measurement error. PMID- 15286930 TI - Complexity of the dynamic QT variability and RR variability in patients with acute anterior wall myocardial infarction: a novel technique using a non-linear method. AB - QT and RR intervals' fluctuations are implicated in the development of malignant arrhythmias. Recent research has quantified repolarization lability using stochastic and linear methods. However, QT-RR intervals are nonlinearly coupled. To this end, QT and RR intervals were extracted from twenty four patients with acute myocardial infarction (AMI) and 13 controls, and a measure of local dimensional complexity (pointwise correlation dimension, PD2) was calculated. PD2 of QT intervals was significantly higher for the patients than for controls (4.83 +/- 0.81 versus 3.40 +/- 0.76, P =.0001), and vice versa for RR intervals (2.51 +/- 0.62 versus 2.91 +/- 0.42, P =.028). The RR and QT measures of complexity were highly correlated only among controls (r = 0.769, P =.0021). Our results support the presence of autonomic abnormalities during infarction and might complement existing tools for assessment of increased risk for sudden death after AMI. PMID- 15286931 TI - Effect of premature ventricular beats on manual and automatic repolarization measurements. AB - OBJECTIVE: To compare QT interval and QT dispersion in ventricular ectopic beats with measurements from the preceding and the immediately following sinus beats, and investigate differences between manual and automatic measurements. PATIENTS: Eleven chronic uremic patients. MAIN OUTCOME MEASURES: ECGs were recorded during hemodialysis treatment and 12-lead sections containing five consecutive beats were extracted, each containing four sinus beats and one centrally-positioned premature ventricular beat. QT measurements were performed both manually and with a computer-automated technique. RESULTS: T wave amplitude was greater in the ectopic beats compared to the sinus beats (0.61 +/- 0.18 vs. 0.23 +/- 0.06 mV, P <.001). The ectopic beats had a greater QT than the sinus beats when measured manually (415 +/- 35 ms vs. 386 +/- 28 ms, P <.001), or automatically (375 +/- 30 vs. 366 +/- 27 ms, P<.01). The sinus beats following the ectopics had a greater QT than the preceding sinus beats (400 +/- 27 vs. 386 +/- 28 ms, P<.001, manual; 382 +/- 24 vs. 366 +/- 27 ms, P<.001, automatic). Differences in QT dispersion were seen only between the ectopic and sinus beats (91 +/- 31 vs. 58 +/- 27 ms, P <.001, manual; 68 +/- 33 vs. 49 +/- 35 ms, P <.001, automatic). CONCLUSIONS: Manual measurement resulted in greater QT values than automatic measurement. Both techniques identified differences between sinus and ectopic beats. The ventricular ectopic beats resulted in an increase in the QT of the immediately following sinus beats. These results confirm the need to interpret QT measurements with care in the presence of ectopic beats. PMID- 15286932 TI - Transmural dispersion of repolarization and ventricular tachyarrhythmias. AB - BACKGROUND: Myocardial transmural dispersion of repolarization (TDR) has been associated with reentrant arrhythmias in animal studies but a clinical association has not yet to been demonstrated. The present study examines the relationship between TDR and ventricular tachyarrhythmias in human subjects. METHODS: This study consisted of 65 patients with non-sustained ventricular tachycardia, sustained ventricular tachycardia, ventricular fibrillation or unexplained syncope with organic heart disease. The control group included 65 patients with paroxysmal supraventricular tachycardia. The 12 ECG was recorded at a recording rate of 100 mm/sec. The interval from the peak to the end of the T wave in the precordial (ECG), referred to as TpTe was assumed to be representative of TDR. RESULTS: Patients were divided into three groups based on the ability to induce VT at the time of electrophysiologic study: VT inducible group (n=37), VT non-inducible group (n=25) and control group (n=65). V4 TpTe/ radical RR was significantly prolonged in the VT inducible group, as compared to the VT non-inducible group (n=25) and the control group (118.9 +/- 26.1 vs. 103.9 +/- 25.7, 104.1 +/- 22.6 ms, P<.05). Patients who develop VT spontaneously (n=13) during a mean follow-up period of 25 months, displayed significantly prolonged V3 TpTe/ radical RR, compared to patients who did not develop VT spontaneously or the control group (132.5 +/- 37.4 vs. 109.8 +/- 26.3, 107.1 +/- 24.1 ms, P <.05). CONCLUSION: Prolonged TDR is associated with inducibility as well as spontaneous development of VT in higher risk patients. TDR may be a useful index for predicting ventricular tachyarrhythmias. PMID- 15286933 TI - Right lateral decubitus position reduces QT dispersion in patients with chronic heart failure. AB - A greater QT dispersion in patients with chronic heart failure (CHF) appears to be a non-invasive marker of susceptibility to malignant ventricular arrhythmias. We evaluated whether QT dispersion in CHF patients is modified by the patients' recumbent position. In 12 CHF patients, and age and sex-matched 12 normal subjects, a single 12-lead surface ECG was recorded in each postural position [left lateral decubitus position (L), supine position (S), and right lateral decubitus position (R)]. In normal subjects, the QT dispersion was comparable in the three recumbent positions [L: 47+/-15 (SD) ms, S: 40+/-9 ms, R: 38+/-14 ms, P=NS]. In contrast, in CHF patients, QT dispersion was significantly shorter in R than those in L and S (L: 93+/-42 ms*, S: 81+/-29 ms*, R: 63+/-24 ms, *P <.05 vs. R). In conclusion, reclining in R reduces the prolonged QT dispersion in CHF patients. PMID- 15286934 TI - Different features of ventricular arrhythmias and the RR-interval dynamics in atrial fibrillation related to the patient's clinical characteristics: an analysis using RR-interval plotting. AB - The clinical features of ventricular arrhythmia and RR-interval dynamics in AF patients remain unresolved. We successively plotted points on an X-Y plain as (X, Y) = (RRn, RRn + 1) from the consecutive RR-intervals of Holter ECGs. Eighty of 175 AF-patients were thus diagnosed to have ventricular arrhythmia based on the different plotting patterns between ventricular premature contractions (VPCs) and aberrations. Different characteristics of the RR-interval dynamics before VPCs were observed such as fixed or variable coupling, and a regular or irregular RR interval sequence. Malignant arrhythmias occurred more frequently in AF-patients with variable coupling VPCs and/or an irregular RR-interval sequence before VPCs than in those with the fixed coupling VPCs and/or the regular RR-interval sequence before VPCs. The RR-interval plotting method enabled us to distinguish different types of VPCs which were related to the clinical characteristics of the AF-patients. PMID- 15286935 TI - Electrocardiographic signs of right ventricular overload in patients who underwent pulmonary embolism event(s). Are they useful in diagnosis of chronic thromboembolic pulmonary hypertension? AB - Diagnosis of chronic thromboembolic pulmonary hypertension (CTEPH) remains a major clinical problem. An attempt was made to learn whether electrocardiography has the potential to alleviate that problem. Sensitivity, specificity, negative and positive predictive value as well as a likelihood ratio of electrocardiogram (ECG) signs of right ventricular overload (RVO) were determined in 56 patients with chronic pulmonary embolism and a mean pulmonary artery pressure (mPAP) ranging from 15 to 80 mm Hg. CPE was recognized as the only disease in 44 patients (Group A) whereas the remaining 12 CPE patients suffered from concomitant cardiac and pulmonary diseases (Group B). Thirty three (59%) of the patients were diagnosed with CTEPH (mPAP exceeding 30 mm Hg). Twelve ECG signs of RVO were identified in the entire population of patients. At least a single ECG sign of RVO was found in 72% of the patients and 7 ECG signs were found exclusively in CTEPH patients. Negative T wave in precordial V1-V5 leads, negative T wave in II, III, aVF, pulmonary P wave and right axis deviation >90% occurred with the highest incidence that was determined to be 43%, 32%, 30%, and 30% respectively. These ECG signs of RVO had positive predictive value of those signs ranged from 80 to 100% in Group A and 25 to 75% in Group B, whereas their negative predictive value ranged from 44 to 76% and from 66.5 to 87.5% respectively. It is concluded that ECG signs of RVO appear to have the potential to aid in diagnosing CTEPH in the patients who underwent acute embolic event in the past. PMID- 15286936 TI - Arrhythmias of a sudden traumatic death. AB - A 73-year-old man was treated because of his paroxysmal palpitations became persistent. At the time of hospital admission atrial flutter was found. Antiarrhythmic drug treatment was unsuccessful therefore electrical cardioversion was indicated which restored his sinus rhythm. After one year of uneventful follow up Holter monitoring was indicated again. When the machine was hooked up sinus rhythm was observed. After 53 minutes a tram knocked down the patient, and he died immediately. During autopsy brain laceration, multiple cranial fractions, mediastinal hemorrhage were found. The Holter recording time was 66 minutes. Before the accident sinus rhythm was recorded. At the time of accident an electrical noise was found, followed by long sinus arrest, atrial fibrillation, nodal escape rhythm, sinus bradycardia, ventricular flutter, tachycardia, fibrillation and "dying heart" rhythm. According to our knowledge this is the first Holter monitoring observation during a sudden traumatic death. PMID- 15286937 TI - Giant R wave, convex ST-segment elevation, and negative T wave during exercise treadmill test. AB - The giant R wave syndrome is characterized by giant R wave accompanied by widening of the QRS complex, marked ST segment elevation, QRS axis deviation, and the formation of monophasic QRS-ST complex with obliteration of S wave in leads facing the ischemic zone. This report describes a 65-year-old-man with variant angina who had a transient giant R wave syndrome during an exercise treadmill test. Initially, at peak exercise, there was a convex ST segment elevation ending in a negative T wave in the same (inferior) leads which showed giant R waves. Later, in the recovery period and coinciding with an amelioration of myocardial ischemia, there was a less marked increase of R wave amplitude associated with concave ST segment elevation and positive T wave in the inferolateral leads. Subsequently, a ST segment depression in the inferolateral leads preceded the ECG normalization. The patient had also a concave ST segment elevation and positive T wave in inferolateral leads during a spontaneous episode of variant angina at rest. An emergency coronary arteriography showed a dominant right coronary artery with an 80% and a 75% diameter stenosis of the middle and distal segment, respectively; the other arteries and left ventriculogram were normal. The underlying mechanisms of the different shapes of ST segment elevation and T waveform in the setting of acute transmural myocardial ischemia are discussed. PMID- 15286938 TI - Retrospective diagnosis of prolonged QT interval and Torsades de Pointes made by analysis of ICD electrograms. AB - We describe a case where a patient received multiple shocks from his implantable cardioverter defibrillator (ICD) for pause dependent polymorphic ventricular tachycardia. The stored far-field electrograms of the ICD were used to make the diagnosis of prolonged QT interval and Torsades de Pointes retrospectively. PMID- 15286939 TI - Rate-dependent QRS prolongation during exercise testing associated with hyperkalemia. AB - The present case showed gradual increase of QRS duration from 100 ms up to 180 msec during an ergometer exercise test along with the heart rate increase. After exercise, QRS duration shortened and normalized. Laboratory test showed hyperkalemia (K = 8.0 mEq/l). T1 myocardial scintigraphy revealed exercise induced transient ischemia in posterolateral region of left ventricle. Coronary angiography showed significant stenosis in the distal portion of left circumflex coronary artery. The increase of QRS duration was possibly due to the combination of hyperkalemia and the effect of mexiletine. The rate dependent blocking effect on sodium channel of mexiletine might be intensified under hyperkalemia. PMID- 15286940 TI - E. B. Wilson's study of cleavage in the egg of the mollusc Dentalium. PMID- 15286941 TI - Ontogenic and ecological control of metamorphosis onset in a carapid fish, Carapus homei: experimental evidence from vertebra and otolith comparisons. AB - In Carapus homei, reef colonisation is associated with a penetration inside a sea cucumber followed by heavy transformations during which the length of the fish is reduced by 60%. By comparing vertebral axis to otolith ontogenetic changes, this study aimed (i) to specify the events linked to metamorphosis, and (ii) to establish to what extent these fish have the ability to delay it. Different larvae of C. homei were caught when settling on the reef and kept in different experimental conditions for at least 7 days and up to 21 days: darkness or natural light conditions, presence of sea cucumber or not, and food deprivation or not. Whatever the nutritional condition, a period of darkness seems sufficient to initiate metamorphosis. Twenty-one days in natural light conditions delayed metamorphosis, whereas the whole metamorphosis process is the fastest (15 days) for larvae living in sea cucumbers. Whether the metamorphosis was initiated or not, otoliths were modified with the formation of a transition zone, whose structure varied depending on the experimental conditions. At day 21, larvae maintained in darkness had an otolith transition zone with more increments (around 80), albeit wider than those (more or less 21) of individuals kept under natural lighting. These differences in otolith growth could indicate an increased incorporation rate of released metabolites by metamorphosing larvae. However, the presence of a transition zone in delayed-metamorphosis larvae suggests that these otolith changes record the endogenously-induced onset of metamorphosis, whereas body transformations seem to be modulated by the environmental conditions of settlement. PMID- 15286942 TI - Effects of temperature and moisture on embryonic diapause of the veiled chameleon (Chamaeleo calyptratus). AB - The development of lizard embryos is typically initiated at fertilization and continues until birth or hatching. In contrast, embryonic development of some chameleons is arrested at the gastrula stage, and embryos remain at this stage for months after the eggs are laid. Our research tested the hypothesis that increased temperature, moisture, or both, are associated with the resumption of development by diapausing embryos of Chamaeleo calyptratus, the veiled chameleon. After 40 days of incubation at 25 degrees C in a relatively dry substrate, eggs were subjected to: 1) no change in temperature or moisture, 2) no change in temperature but change from a dry to a wet substrate, 3) change to a warmer temperature but no change in substrate moisture, or 4) an increase in both temperature and substrate moisture. Overall, embryos initiated development after 50-60 days to 80 or more days of incubation. Neither substrate moisture nor water uptake by eggs was related to the interval when development resumed. In contrast, development was initiated about 10 days earlier for eggs in the high temperature treatment compared to eggs in the low temperature treatment. Our results suggest that neither water availability nor water uptake by eggs affect the length of diapause but that an increase in ambient temperature initiates development of diapausing embryos of C. calyptratus. PMID- 15286943 TI - The effect of impaired thyroid function during development on the mechanical properties of avian bone. AB - Thyroid hormones show fluctuating levels during the post-hatching development of birds. In this paper we report the results of the first mechanical tests to quantify the effect of hypothyroidism, during post-natal development, on the skeletal properties of a precocial bird, the barnacle goose, as determined by microhardness testing. The effect of hypothyroidism is tissue-specific; bone from the femora of birds is not significantly affected by induced hypothyroidism, however, there is a strong positive relationship between the levels of circulating thyroid hormones and the mechanical properties of bone from humeri. In the barnacle goose the development of the wing skeleton and musculature depends on an increase in circulating thyroid hormones and our analysis shows that, in its absence, the mechanical competence of the bone mineral itself is reduced in addition to the decreased bone length and muscle development previously reported in the literature. PMID- 15286944 TI - The effects of opercular linkage disruption on prey-capture kinematics in the teleost fish Sarotherodon melanotheron. AB - The kinematics of prey capture in blackchin tilapia (Sarotherodon melanotheron) subjected to three experimental treatments (control, anesthetization, and opercular linkage disruption) were analyzed using high-speed video to explore the role of the opercular four-bar linkage in depressing the lower jaw in teleost fishes. A series of two-way mixed model analyses of variance (random effects=fish; fixed effects=treatment) revealed that maximum gape, lower jaw angle, gape cycle, and time to lower jaw depression differed among treatments. Tukey post-hoc comparisons revealed that the opercular linkage disruption treatment differed from the control and anesthetization treatments, suggesting that severing the opercular linkage affected the ability of fish to depress the lower jaw. We hypothesize that although the opercular four-bar linkage system may not be the only linkage mechanism involved in depressing the lower jaw, it plays a very important role in opening the mouth during feeding in teleost fishes. PMID- 15286945 TI - Immunolocalization of Na+/K+-ATPase, carbonic anhydrase II, and vacuolar H+ ATPase in the gills of freshwater adult lampreys, Geotria australis. AB - As adults, anadromous lampreys migrate from seawater into freshwater rivers, where they require branchial ion (NaCl) absorption for osmoregulation. In teleosts and elasmobranchs, pharmological, immunohistochemical, and molecular data support roles for Na+/K+-ATPase (NPPase), carbonic anhydrase II (CAII), and vacuolar H+-ATPase (V-ATPase) in two different models of branchial ion absorption. To our knowledge, these transport-related proteins have not been studied in adult freshwater lampreys, and therefore it is not known if they are expressed, or have similar functions, in lampreys. The purpose of this study was to localize NPPase, CAII, and V-ATPase in the gills of adult freshwater lampreys and determine if any of these transport-related proteins are expressed in the same cells. Heterologous antibodies were used to localize the three proteins in gill tissue from pouched lamprey (Geotria australis). Immunoreactivity (IR) for all three proteins occurred between, and at the base of, lamellae in cells that match previous descriptions of mitochondrion-rich-cells (MRCs). NPPase-IR was always on the basolateral side of cells that did not stain for CAII or V-ATPase. In contrast, CAII-IR was always on the apical side of cells that also contained diffuse V-ATPase-IR. Therefore, we have identified two types of MRC in adult freshwater lamprey gills based on immunohistochemical staining for three transport proteins. A model of ion transport, based on our results, is proposed for adult freshwater lampreys. PMID- 15286946 TI - Contribution of the submentalis muscle to feeding mechanics in the leopard frog, Rana pipiens. AB - This study investigated the functional contributions of the submentalis muscle to the coordination of feeding behavior in the leopard frog, Rana pipiens. Additionally, the anatomical origins of the motor neurons innervating this muscle are identified and described. The m. submentalis is a small muscle connecting the distal mandibular tips. Depending upon the anuran species studied, this muscle contributes to mandibular bending and the degree to which the tongue is protracted, or has little or no role in feeding biomechanics. High-speed videography was used to quantify feeding attempts before versus after bilateral denervation of the m. submentalis. Additionally, the terminal branch of the trigeminal nerve prior to innervating the m. submentalis was retrogradely labeled to identify the origins of motor neurons innervating the muscle. For the kinematic analyses, denervation of the submentalis resulted in significant increases in the time to maximum tongue protrusion, and the duration of tongue protrusion. Neither mandibular bending, nor tongue length variables differed significantly between normal conditions and deafferented conditions. However, when unsuccessful feeding attempts were quantified following the denervation, failed attempts were nearly always due to the tongue not reaching the prey. None of the unsuccessful feedings prior to denervation were due to inadequate tongue protrusion. Anatomical data show a much larger rostral-caudal distribution of the trigeminal motor neurons than previously described for anurans. These data suggest a larger role for the submentalis muscle in Rana than in previously studied anurans with long protrusible tongues, and suggests a feedback mechanism from the trigeminal nerve to the nerves coordinating tongue protraction and retraction. PMID- 15286947 TI - Photoreceptor topography in the duplex retina of the paddlefish (Polyodon spathula). AB - Retinal whole-mount preparations from the eyes of the North American paddlefish, Polyodon spathula, were examined with a combination of bright field and differential interference contrast microscopy. The entire retina was mapped and population counts of rod and cone photoreceptors were made at regular intervals throughout the retina. The retina is dominated by rods, but a significant percentage (ca. 38%) of the photoreceptors are cones. Mean cone packing density for the entire retina is 6,402+/-1,216 cones/mm2. There is a small (16%) but statistically significant difference between cone packing density in the dorsal retina (6,674+/-1,168 cones/mm2) and the ventral retina (5,745+/-1,076 cones/mm2). There is no region of unusually high cone concentration that might be construed as a fovea or a visual streak. Mean rod packing density for the entire retina is 10,271+/-1,205 rods/mm2. Except in the far periphery, where rods are less numerous, the density of rods is fairly uniform throughout the retina. The data are discussed with regard to paddlefish habitat and behavior. PMID- 15286948 TI - Baseline and stress-induced glucocorticoids during reproduction in the variable flying fox, Pteropus hypomelanus (Chiroptera: Pteropodidae). AB - Baseline and stress-responsive glucocorticoid (GC) levels were assessed during early pregnancy, late pregnancy, and lactation in female variable flying foxes (Pteropus hypomelanus) and in males over the same time period. Animals were maintained in a breeding colony in captivity. High levels of both cortisol and corticosterone were detected, with total plasma GC levels being among the highest documented in vertebrates (up to 3000 ng/ml in individual animals, with cortisol being the primary GC, accounting for approximately 78% of total GCs), and significantly greater in males than in females. Plasma levels of cortisol and corticosterone showed nearly identical profiles within each sex, with the exception of females in late pregnancy, in which corticosterone, but not cortisol, increased significantly. Baseline levels of plasma cortisol were highest in September (when pups were between 1 and 2 months of age) in both sexes, which may be related to the approaching onset of the mating period. There was a continuum in the magnitude of the response to stress (handling and sampling) over time in females, with the greatest stress response in early pregnancy, a dampened response during late pregnancy, and no significant stress response during lactation. Surprisingly, males failed to exhibit elevated GCs after this stress, but did have significant stress-induced hyperglycemia and suppression of plasma testosterone levels. This may be due to their high (perhaps maximal) baseline levels, which suggests that being in a breeding group was chronically stressful for males. PMID- 15286949 TI - cDNA cloning and mRNA expression of a FTZ-F1 homologue from the pituitary of the orange-spotted grouper, epinephelus coioides. AB - A FTZ-F1 homologue was cloned from the pituitary cDNA library of the orange spotted grouper. The full-length cDNA of the orange-spotted grouper FTZ-F1 spanned 1735 bp including a poly (A) tail. The open reading frame encodes a protein of 468 amino acids. Sequence analysis indicated that it had a structure typical of the orphan nuclear receptor superfamily, and the FTZ-F1 box, a characteristic of the FTZ-F1 family. Phylogenetic analysis indicated that the orange-spotted grouper FTZ-F1 was closely related to medaka FTZ-F1 and did not belong to either the SF-1/Ad4BP group or the LRH-1/FTF group. Virtual Northern Blot detected a major transcript of about 1.7 kb and a minor transcript of 2.2 kb of FTZ-F1 in the orange-spotted grouper pituitary gland. The expression of FTZ-F1 homologue gene in different tissues and during embryonic development of the orange-spotted grouper was determined using one-step RT-PCR coupled with Southern blot analysis. In addition to the pituitary gland, the orange-spotted grouper FTZ F1 was also expressed in the hypothalamus, forebrain, heart, liver, kidney, and ovary. The stronger signal from the gel image indicated that the expression level of FTZ-F1 homologue gene was higher in the ovary of stage 3 than stage 2. During embryonic development, mRNA for the orange-spotted grouper FTZ-F1 homologue was present in newly fertilized eggs, but disappeared in embryos at 50 min post fertilization. The orange-spotted grouper FTZ-F1 homologue mRNA reappeared in embryos at 1.5 hr post fertilization. Its expression level was increased from late blastula to neurula stages. Taken together, results of the current study suggest that the orange-spotted grouper FTZ-F1 homologue exhibits characteristics indicative of both the LRH-1/FTF- and the SF-1/Ad4BP-like genes, and may also play important roles in the hypothalamus-pituitary-gonadal axis, cholesterol metabolism, and embryogenesis. PMID- 15286951 TI - Using information technology to improve surgical safety. PMID- 15286952 TI - Angiogenesis and neoplastic transformation of Barrett's epithelium. PMID- 15286953 TI - Postoperative mortality following oesophagectomy and problems in reporting its rate. AB - BACKGROUND: There have been three previous reviews of the world literature describing postoperative mortality rate following oesophagectomy. The first documented rates in the first half of the last century, the second the period 1960-1979 and the third the interval 1980-1988. The aim of this review was to document the rate for the period 1990-2000. METHODS: Reports were sourced through PubMed and/or Medline listings. RESULTS: The number of papers included in the review was 312, involving 70,756 patients. The overall mortality rate was 6.7 per cent. The 30-day mortality rate was 4.9 per cent and the in-hospital mortality rate 8.8 per cent. Survival rates, where reported, were 62.7 per cent at 1 year and 27.9 per cent at 5 years. CONCLUSION: Operative mortality rates following oesophagectomy have continued to fall. However, the true rate is almost certainly higher than that reported here, for a variety of reasons. The 1-year survival of patients was only reported in about a quarter of the papers. It may be a more meaningful figure than postoperative mortality rate. PMID- 15286954 TI - Systematic review and meta-analysis of controlled trials assessing spinal cord stimulation for inoperable critical leg ischaemia. AB - BACKGROUND: Spinal cord stimulation (SCS) may have a place in the treatment of patients with inoperable chronic critical leg ischaemia. METHODS: A systematic review and meta-analysis was performed of all controlled studies comparing SCS in addition to any form of conservative treatment for inoperable chronic critical leg ischaemia. Main endpoints were limb salvage, pain relief and clinical situation. Systematic methodological appraisal and data extraction were performed by independent reviewers. RESULTS: Of the 18 reports found, nine trials, comprising 444 patients, matched the selection criteria. After pooling, limb salvage at 12 months appeared significantly greater in the SCS group (risk difference (RD) -0.13 (95 per cent confidence interval (c.i.) -0.04 to -0.22)). Significant pain relief occurred in both treatment groups, but patients who received SCS required significantly less analgesia and reached Fontaine stage 2 more often than those who did not have SCS (RD 0.33 (95 per cent c.i. 0.19 to 0.47)). Complications of SCS were problems of implantation (8.2 per cent), changes in stimulation requiring reintervention (14.8 per cent) and infection (2.9 per cent). CONCLUSION: The addition of SCS to standard conservative treatment improves limb salvage, ischaemic pain and the general clinical situation in patients with inoperable chronic critical leg ischaemia. These benefits should be weighed against the cost and the (minor) complications associated with the technique. PMID- 15286955 TI - A review of clinical guidelines. AB - BACKGROUND: Clinical guidelines are increasingly used in patient management but few clinicians are familiar with their origin or appropriate application. METHODS: A Medline search using the terms 'clinical guidelines' and 'practice guidelines' was conducted. Additional references were sourced by manual searching from the bibliographies of articles located. RESULTS AND CONCLUSION: Clinical guidelines originated in the USA in the early 1980s, initially as a cost containment exercise. Significant improvements in the process and outcomes of care have been demonstrated following their introduction, although the extent of improvement varies considerably. The principles for the development of guidelines are well established but many published guidelines fall short of these basic quality criteria. Guidelines are only one aspect of improving quality and should be used within a wider framework of promoting clinical effectiveness. Understanding their limitations as well as their potential benefits should enable clinicians to have a clearer view of their place in everyday practice. PMID- 15286956 TI - Low molecular weight heparin for the prevention of venous thromboembolism after abdominal surgery. AB - BACKGROUND: Abdominal surgery carries a significant risk of venous thrombosis, a risk further increased in patients with cancer. METHODS: Embase and Pubmed searches between 1980 and 2003, using the key words 'heparin,' 'surgery,' 'abdominal or rectal or colorectal or rectum or colon' and 'clinical trial', were conducted to identify studies of thromboprophylaxis in patients having abdominal surgery. RESULTS: A total of 16 comparative studies were identified. These show that low molecular weight heparin (LMWH) is as effective as unfractionated heparin (UFH) in reducing venous thromboembolism after abdominal surgery and, at appropriate doses, can reduce bleeding complications. In very high-risk patients, a higher dose of LMWH may offer increased efficacy without increasing bleeding risk. Extending the standard 7-10-day period of prophylaxis may benefit certain high-risk groups; recent data show a significant benefit of 4-week enoxaparin thromboprophylaxis compared with a standard regimen, at no cost to safety. CONCLUSION: Patients undergoing abdominal surgery should be stratified according to thromboembolism risk and managed accordingly. LMWH is a recommended alternative to UFH in moderate- or high-risk patients. In patients with cancer, high doses of LMWH may offer increased efficacy without increasing the bleeding risk and an extended 4-week period of prophylaxis appears beneficial. PMID- 15286957 TI - Randomized clinical trial of laparoscopic versus open fundoplication for gastro oesophageal reflux disease. AB - BACKGROUND: The aim of this study was to compare laparoscopic and open Nissen fundoplication for gastro-oesophageal reflux disease in a randomized clinical trial. METHODS: Ninety-nine patients were randomized to either laparoscopic (52) or open (47) Nissen fundoplication. Patients with oesophageal dysmotility, those requiring a concurrent abdominal procedure and those who had undergone previous antireflux surgery were excluded. Independent assessment of dysphagia, heartburn and patients' satisfaction 1, 3, 6 and 12 months after surgery was performed using multiple standardized clinical grading systems. Objective measurement of oesophageal acid exposure and lower oesophageal sphincter pressure before and after surgery, and endoscopic assessment of postoperative anatomy, were performed. RESULTS: Operating time was longer in the laparoscopic group (median 82 versus 46 min). Postoperative pain, analgesic requirement, time to solid food intake, hospital stay and recovery time were reduced in the laparoscopic group. Perioperative outcomes, postoperative dysphagia, relief of heartburn and overall satisfaction were equally good at all follow-up intervals. Reduction in oesophageal acid exposure, increase in lower oesophageal sphincter tone and improvement in endoscopic appearances were the same for the two groups. CONCLUSION: The laparoscopic approach to Nissen fundoplication improved early postoperative recovery, with an equally good outcome up to 12 months. PMID- 15286958 TI - Randomized clinical trial of the effects of preoperative and postoperative oral nutritional supplements on clinical course and cost of care. AB - BACKGROUND: Postoperative oral nutritional supplementation has been shown to be of clinical benefit. This study examined the clinical effects and cost of administration of oral supplements both before and after surgery. METHODS: This was a randomized clinical trial conducted in three centres. Patients undergoing lower gastrointestinal tract surgery were randomized to one of four groups: group CC received no nutritional supplements, group SS took supplements both before and after surgery, group CS received postoperative supplements only, and group SC were given supplements only before surgery. Preoperative supplements were given from the time it was decided to operate to 1 day before surgery. Postoperative supplements were started when the patient was able to take free fluids and continued for 4 weeks after discharge from hospital. Data collected included weight change, complications, length of stay, nutritional intake, anthropometrics, quality of life and detailed costings covering all aspects of care. RESULTS: Some 179 patients were randomized, of whom 27 were withdrawn and 152 analysed (CC 44, SS 32, CS 35, SC 41). Dietary intake was similar in all four groups throughout the study. Mean energy intake from preoperative supplements was 536 and 542 kcal/day in the SS and SC groups respectively; that 2 weeks after discharge from hospital was 274 and 361 kcal/day in the SS and CS groups respectively. There was significantly less postoperative weight loss in the SS group than in the CC and CS groups (P < 0.050), and significantly fewer minor complications in the SS and CS groups than the CC group (P < 0.050). There were no differences in the rate of major complications, anthropometrics and quality of life. Mean overall costs were greatest in the CC group, although differences between groups were not significant. CONCLUSION: Perioperative oral nutritional supplementation started before hospital admission for lower gastrointestinal tract surgery significantly diminished the degree of weight loss and incidence of minor complications, and was cost-effective. PMID- 15286959 TI - D-dimer testing in patients with suspected acute thromboembolic occlusion of the superior mesenteric artery. AB - BACKGROUND: There is no accurate non-invasive method available for the diagnosis of acute thromboembolic occlusion of the superior mesenteric artery (SMA). The aim of this study was to assess the diagnostic properties of the fibrinolytic marker D-dimer. METHODS: From September 2000 to April 2003 consecutive patients aged over 50 years admitted to hospital with acute abdominal pain were studied. Patients with possible acute SMA occlusion at presentation had blood samples taken within 24 h of the onset of the pain for analysis of D-dimer, plasma fibrinogen, activated partial thromboplastin time, prothrombin time and antithrombin. The value of D-dimer testing to diagnose SMA occlusion was assessed by means of likelihood ratios. RESULTS: Nine of 101 patients included had acute SMA occlusion. The median D-dimer concentration was 1.6 (range 0.4-5.6) mg/l, which was higher than that in 25 patients with inflammatory disease (P = 0.007) or in 14 patients with intestinal obstruction (P = 0.005). The combination of a D dimer level greater than 1.5 mg/l, atrial fibrillation and female sex resulted in a likelihood ratio for acute SMA occlusion of 17.5, whereas no patient with a D dimer concentration of 0.3 mg/l or less had acute SMA occlusion. CONCLUSION: D dimer testing may be useful for the exclusion of patients with suspected acute SMA occlusion. PMID- 15286960 TI - Oesophagocardioplasty for residual dysphagia following multiple pneumatic dilatations for achalasia. PMID- 15286961 TI - Effect of surveillance of Barrett's oesophagus on the clinical outcome of oesophageal cancer. AB - BACKGROUND: Surveillance programmes for Barrett's oesophagus have been implemented in an effort to detect oesophageal adenocarcinoma at an earlier and potentially curable stage. The aim of this study was to examine the impact of endoscopic surveillance on the clinical outcome of patients with adenocarcinoma complicating Barrett's oesophagus. METHOD: Consecutive patients who underwent oesophageal resection for high-grade dysplasia or adenocarcinoma arising from Barrett's oesophagus were studied retrospectively. The pathological stage and survival of patients identified as part of a surveillance programme were compared with those of patients presenting with symptomatic adenocarcinoma. RESULTS: Seventeen patients in the surveillance group and 74 in the non-surveillance group underwent oesophagectomy. Disease detected in the surveillance programme was at a significantly earlier stage: 13 of 17 versus 11 of 74 stage 0 or I, three versus 26 stage II, and one versus 37 stage III or IV (P < 0.001). Lymphatic metastases were seen in three of 17 patients in the surveillance group and 42 of 74 who were not under surveillance (P = 0.004). Three-year survival was 80 and 31 per cent respectively (P = 0.008). CONCLUSION: Patients with surveillance-detected adenocarcinoma of the oesophagus are diagnosed at an earlier stage and have a better prognosis than those who present with symptomatic tumours. PMID- 15286962 TI - Value of positron emission tomography in the diagnosis of recurrent oesophageal carcinoma. AB - BACKGROUND: Positron emission tomography (PET) with [18F]fluorodeoxyglucose (FDG) might be useful for staging oesophageal squamous cell carcinoma (SCC). FDG-PET may be more accurate than computed tomography (CT) in diagnosing lymph node metastasis. This retrospective study compared the ability of FDG-PET and CT to diagnose recurrent oesophageal carcinoma. METHODS: Fifty-five patients with thoracic oesophageal SCC who had undergone radical oesophagectomy were studied. The accuracy of FDG-PET and CT in detecting recurrence during follow-up was calculated using data from the first images generated by either modality that suggested the presence of recurrent disease. Lesions deemed to be equivocal on these scans were considered as positive for recurrence. RESULTS: Twenty-seven of the 55 patients had recurrent disease in a total of 37 organs. Locoregional recurrence was observed in 19 patients (35 per cent). Distant recurrent disease occurred in 15 patients (27 per cent) in 18 organs. Six patients had recurrence in the liver, four in the lung, six in bone and two in distant lymph nodes. FDG PET showed 96 per cent sensitivity, 68 per cent specificity and 82 per cent accuracy in demonstrating recurrent disease. The corresponding values for CT were 89, 79 and 84 per cent. The sensitivity of FDG-PET was higher than that of CT in detecting locoregional recurrence, but its specificity was lower because of FDG uptake in the gastric tube and thoracic lymph nodes. In distant organs the sensitivity of PET in detecting lung metastasis was lower than that of CT, but its sensitivity for bone metastasis was higher. CONCLUSION: FDG-PET has a larger field than CT. Combined PET-CT would appear to be an appropriate modality for the detection of recurrent oesophageal cancer. PMID- 15286963 TI - Ineffective oesophageal motility does not affect the clinical outcome of open Nissen fundoplication. AB - BACKGROUND: Nissen fundoplication is considered the 'gold standard' in antireflux surgery but some surgeons employ a different surgical strategy when gastro oesophageal reflux disease (GORD) is associated with motor disorders of the oesophageal body. METHODS: Ninety-three patients undergoing surgery for GORD were divided into two groups: 52 patients (group 1) had normal oesophageal body motility and 41 (group 2) had ineffective oesophageal motility (IOM). All patients had a short Nissen fundoplication via a laparotomy. The median follow-up was 5 years in group 1 and 6.5 years in group 2. RESULTS: The clinical outcome was satisfactory in more than 90 per cent of the patients in both groups. Only one of ten patients with IOM and dysphagia before operation still had dysphagia after surgery. One patient in each group developed postoperative dysphagia. Six of 52 patients with normal motility and eight of 41 with IOM had persistent pathological acid reflux after surgery. Significant increases in contractile wave pressure and a decrease in the percentage of non-propagated waves were found in group 2 after fundoplication. CONCLUSION: Patients with IOM did not have an increased rate of dysphagia after total fundoplication compared with those with normal motility, but they did have a higher rate of recurrence of endoscopic and pH-proven reflux. PMID- 15286964 TI - Prospective study of routine contrast radiology after total gastrectomy. AB - BACKGROUND: The practice of routine contrast radiology before recommencing oral nutrition after total gastrectomy is not evidence based. The aim of this prospective study was to evaluate the clinical role and timing of this investigation. METHODS: Seventy-six consecutive patients underwent total gastrectomy with a stapled oesophagojejunal anastomosis. A contrast swallow using non-ionic contrast and barium was performed routinely 5 and 9 days after surgery. The surgeon was blinded to the result of the first of these examinations. Patients with clinical evidence of a leak underwent contrast radiology and upper gastrointestinal videoendoscopy. RESULTS: Eight patients (11 per cent) developed a clinical leak from the oesophagojejunal anastomosis, seven before the first scheduled contrast swallow. Contrast radiology identified a leak in four of six patients. Endoscopy detected a leak in both patients with a false-negative swallow and in two patients who were not fit to undergo contrast radiology. Routine contrast radiology identified a subclinical leak in a further five patients (7 per cent), none of whom developed clinical signs. Four of seven in hospital deaths were associated with an anastomotic leak. CONCLUSION: There is no role for routine contrast swallow after total gastrectomy with a stapled oesophagojejunal anastomosis, but patients with clinical suspicion of leakage should undergo urgent contrast radiology, plus endoscopy if the contrast examination is normal. PMID- 15286965 TI - Major resection for chronic pancreatitis in patients with vascular involvement is associated with increased postoperative mortality. AB - BACKGROUND: The aim was to evaluate the outcome of major resection for chronic pancreatitis in patients with and without vascular involvement. METHODS: Of 250 patients with severe chronic pancreatitis referred between 1996 and 2003, 112 underwent pancreatic resection. The outcome of 17 patients (15.2 per cent) who had major vascular involvement was compared with that of patients without vascular involvement. RESULTS: The 95 patients without vascular involvement had resections comprising Beger's operation (39 patients), Kausch-Whipple pancreatoduodenectomy (28), total pancreatectomy (25) and left pancreatectomy (three). Twenty-five major vessels were involved in the remaining 17 patients. One or more major veins were occluded and/or compressed producing generalized or segmental portal hypertension, and three patients also had major arterial involvement. Surgery in these patients comprised Beger's operation (eight), total pancreatectomy (five), Kausch-Whipple pancreatoduodenectomy (two) and left pancreatectomy (two). Perioperative mortality rates were significantly different between the groups (two of 95 versus three of 17 respectively; P = 0.024). There were similar and significant improvements in long-term outcomes in both groups. CONCLUSION: Resection for severe chronic pancreatitis in patients with vascular complications is hazardous and is associated with an increased mortality rate. Vascular assessment should be included in the routine follow-up of patients with chronic pancreatitis, to enable early identification of those likely to develop vascular involvement and prompt surgical intervention. PMID- 15286966 TI - Urinary trypsinogen activation peptide as a marker of severe acute pancreatitis. AB - BACKGROUND: Trypsinogen activation peptide (TAP) may be an early marker of severe pancreatitis. Previous studies have included all patients with organ failure in the group with severe pancreatitis, although patients with transient organ failure may have a good prognosis. The aim of this study was to determine the value of urinary TAP estimation for prediction of severity of acute pancreatitis, and to validate use of several markers of prediction of severity against a new, stringent definition of severity. METHODS: Patients with acute pancreatitis were recruited within 24 h of onset of symptoms. Urine and blood samples were collected for 24 h, and Acute Physiology And Chronic Health Evaluation (APACHE) II (24 h), Ranson (48 h) and Glasgow (48 h) scores were calculated. Severe acute pancreatitis was defined by the presence of a local complication or the presence of organ failure for more than 48 h. RESULTS: Urinary TAP levels were significantly greater in patients with severe pancreatitis than in those with mild disease during the first 36 h of admission. The highest of three estimations of TAP in the first 24 h was as effective as APACHE II at 24 h in predicting severity. At 24 h after admission, urinary TAP was better than C-reactive protein (CRP) in predicting severity. The combination of TAP and CRP at 24 h allowed identification of high- and low-risk groups. The new definition of severity excluded 24 of 190 patients with transient organ failure; none of these patients died. CONCLUSION: Use of TAP improved early prediction of the severity of acute pancreatitis. Organ failure that resolves within 48 h does not signify a severe attack of acute pancreatitis. PMID- 15286967 TI - Induction of adhesion molecule expression in liver ischaemia-reperfusion injury is associated with impaired hepatic parenchymal microcirculation. AB - BACKGROUND: Activated neutrophils may be important mediators in liver ischaemia reperfusion injury (I/R). Adhesion of leucocytes to the endothelial cell surface is a result of activation of cell adhesion molecules. The aim of this study was to investigate the effect of I/R on the hepatic microcirculation (HM) and intercellular adhesion molecule (ICAM) 1 expression. METHODS: Four groups of six Sprague-Dawley rats underwent laparotomy for liver exposure. Group 1 acted as controls, and groups 2-4 underwent partial liver ischaemia for 30, 45 and 60 min respectively followed by reperfusion for 60 min. Flow in the HM was measured by laser Doppler flowmetry. Liver biopsies were taken at the end of the reperfusion period. ICAM-1 expression was assessed by immunohistochemistry (graded 0-3). RESULTS: Mean flow in the HM was significantly reduced with I/R (mean(s.e.m.) red cell flux 140(21), 52(3) and 43(2) with 30, 45 and 60 min ischaemia compared with control 230(17); all P < 0.001). ICAM-1 expression was significantly induced (mean(s.e.m.) 1.30(0.21), 2.50(0.22) and 2.80(0.17) with 30, 45 and 60 min ischaemia versus control 0.50(0.22); all P < 0.001). CONCLUSION: I/R produced a significant upregulation of ICAM-1 expression which correlated with impaired flow in the HM. PMID- 15286968 TI - Prognostic significance of both surgical and pathological assessment of curative resection for rectal cancer. AB - BACKGROUND: Information on whether surgery has been 'curative' is essential for prediction of prognosis and for selection of patients for adjuvant treatment. The aim of this study was to analyse the prognostic value of surgeons' and pathologists' assessments of tumour clearance in patients with primary rectal cancer who underwent resection. METHODS: A total of 1550 patients were studied prospectively. Data were collected from reports made by surgeons and pathologists on whether tumour clearance had been 'complete', 'uncertain' or 'incomplete'. The predictive value in relation to outcome after at least 5 years of follow-up was assessed. RESULTS: In patients assessed as having a complete surgical clearance, tumour recurrence developed in 33.3 per cent. For patients with an uncertain or incomplete clearance the recurrence rate was 59.5 and 61 per cent respectively (P < 0.001). The relative risk of recurrence was twice as high when the surgeon and pathologist disagreed than when they both agreed on the complete clearance. Survival in patients with a complete, uncertain or incomplete surgical clearance was 55.3, 23.0 and 10 per cent respectively (P = 0.050). CONCLUSION: Assessments of tumour clearance were of strong prognostic value in relation to outcome. When the surgeon or pathologist was uncertain, or there was disagreement about the completeness of clearance, the risk of recurrence was similar to that among patients in whom an incomplete resection had been performed. PMID- 15286969 TI - Mortality and morbidity of planned relaparotomy versus relaparotomy on demand for secondary peritonitis. AB - BACKGROUND: Planned relaparotomy (PR) and relaparotomy on demand (ROD) are both frequently used in the treatment of secondary peritonitis. The aim of this study was to evaluate the mortality, morbidity and long-term outcome associated with PR compared with ROD in patients with secondary peritonitis admitted to a university hospital. METHOD: This retrospective study included 278 consecutive patients who underwent emergency laparotomy for secondary peritonitis between January 1994 and January 2000. Outcome was analysed based on the decision made by the surgeon during the first operation to perform either ROD (197 patients) or PR (81). RESULTS: The Acute Physiology And Chronic Health Evaluation II score was comparable in ROD and PR groups (10.8 versus 11.7; P = 0.222). The in-hospital mortality rate was significantly lower with ROD than PR (21.8 versus 36 per cent; P = 0.016). Two-year survival(s.e.) was 65.8(3.4) per cent in the ROD group and 55.5(5.5) per cent in the PR group (P = 0.031). CONCLUSION: The in-hospital and long-term survival rates were higher in patients with secondary peritonitis treated by ROD than in those with disease of comparable severity treated by PR. Choice of treatment strategy was an independent predictor of survival. PMID- 15286970 TI - Detection of circulating oesophageal squamous cancer cells in peripheral blood and its impact on prognosis. AB - BACKGROUND: Many studies have attempted to detect cancer cells using the reverse transcription-polymerase chain reaction (RT-PCR) for specific mRNAs. None has examined the correlation between the presence of circulating oesophageal cancer cells in peripheral blood and long-term outcome. METHODS: Blood samples were obtained on admission, and before, during and after operation from 70 patients with squamous oesophageal cancer who had complete clinicopathological records and who underwent curative oesophagectomy between June 1997 and June 2000. RT-PCR for mRNA encoding squamous cell carcinoma antigen (SCCA mRNA) was used to detect oesophageal cancer cells in peripheral blood. RESULTS: Twenty-three patients (33 per cent) were positive for SCCA mRNA on admission and 17 of these patients developed recurrent disease. SCCA mRNA on admission correlated with the depth of tumour invasion (P < 0.001) and with venous invasion (P < 0.001). Eleven of 24 patients with a positive intraoperative result were positive for SCCA mRNA only during operation, of whom seven also developed recurrence. CONCLUSION: RT-PCR for SCCA mRNA can detect oesophageal cancer cells in peripheral blood. The presence of such cells in blood samples obtained on admission or during operation is a useful predictor of outcome in patients with oesophageal squamous cell carcinoma. PMID- 15286971 TI - Laparoscopically assisted distal gastrectomy for early gastric cancer in the elderly. AB - BACKGROUND: Open gastrectomy is associated with increased morbidity and a longer hospital stay than laparoscopically assisted gastrectomy. The aim of this study was to clarify the value of laparoscopically assisted distal gastrectomy (LDG) in the elderly, in whom co-morbid disease is generally more common. METHODS: Forty five elderly patients (aged 70 years or more) and 57 younger patients who underwent LDG, and 28 elderly patients who underwent open distal gastrectomy (ODG) for early gastric cancer between January 1994 and April 2003 were studied. Demographics and postoperative outcomes were compared. RESULTS: : Co-morbidity was more common in elderly patients than in younger patients who underwent LDG (25 of 45 versus 16 of 57; P = 0.004). The postoperative complication rate, time to solid diet and postoperative hospital stay were similar in these two groups. Elderly patients who underwent LDG had a significantly reduced medical complication rate (two of 45 versus six of 28; P = 0.023), time to first flatus (3.7 versus 4.2 days; P = 0.042), time to solid diet (4.6 versus 5.5 days; P = 0.011) and postoperative hospital stay (16.3 versus 23.9 days; P = 0.011) than elderly patients who had ODG. CONCLUSION: LDG offers particular advantages to elderly patients with early gastric cancer, including rapid return of gastrointestinal function, fewer complications and a shorter hospital stay. PMID- 15286972 TI - Factors influencing survival after resection of pulmonary metastases from colorectal cancer. AB - BACKGROUND: Selection of patients for resection of lung metastases from colorectal cancer is problematic. The aim of this study was to evaluate clinically relevant prognostic factors and to define a subgroup of patients who would most benefit from such surgery. PATIENTS: Seventy-five patients (median age 58 (range 33-82) years) with pulmonary metastases from colorectal cancer underwent 104 R0 lung resections. Median follow-up was 33 (range 4-116) months. Patients who had no evidence of recurrent extrathoracic disease, no more than three metastases on either side, lobectomy as the maximal surgical procedure, and adequate cardiorespiratory function were eligible for surgery. Univariate and multivariate Cox regression, and classification and regression tree subgroup analyses were performed. RESULTS: Overall median survival was 33 months, with 3- and 5-year survival rates of 47 and 27 per cent respectively. Size of metastases (relative risk (RR) 2.6) and extent of resection (RR 0.4) were identified as independent prognostic factors. Primary tumour stage was significant in univariate analysis. Subgroup analysis defined two statistically relevant prognostic groups: patients with a maximum metastasis size of 3.75 cm or less with a disease-free interval of more than 10 months and patients with larger metastases and a shorter disease-free interval. Median survival and 5-year survival were 45 months and 39 per cent in the former group, and 24 months and less than 11 per cent in the latter. CONCLUSION: Subgroup analysis provided criteria for the selection of patients for R0 resection of lung metastases from colorectal cancer and differentiated between those at high or low risk of early tumour progression; the latter patients would benefit most from surgery. PMID- 15286973 TI - Critical evaluation of the different staging systems for hepatocellular carcinoma (Br J Surg 2004; 91: 400-408). PMID- 15286974 TI - Drawbacks of endoscopic thoracic sympathectomy (Br J Surg 2004; 91: 264-269). PMID- 15286975 TI - Long-term outcome following loose-seton technique for external sphincter preservation in complex anal fistula (Br J Surg 2004; 91: 476-480). PMID- 15286976 TI - Chronic anal fissure (Br J Surg 2004; 91: 270-279). PMID- 15286980 TI - Molecular understanding of oxygen-tension and patient-variability effects on ex vivo expanded T cells. AB - Immunotherapy with ex vivo cultured T cells depends on a large supply of biologically active cells. Understanding the effects of culture parameters is essential for improving the proliferation and efficacy of the expanded cells. Low oxygen tension (5% pO(2)) was previously reported to improve T-cell expansion and alter cellular phenotypic characteristics compared to T cells cultured at 20% pO(2). Here we report the use of DNA-array based transcriptional analysis coupled with protein-level analysis to provide molecular insights into pO(2) and patient variability effects on expanded primary human T cells. Analysis of seven blood samples showed that reduced pO(2) results in higher expression of genes important in lymphocyte biology, immune function, and cell-cycle progression. 20% pO(2) resulted in higher expression of genes involved in stress response, cell death, and cellular repair. Expression of granzyme A (gzmA) was found to be significantly regulated by oxygen tension with cells at 5% pO(2) having greater gzmA expression than at 20% pO(2). Protein-level analysis of gzmA was consistent with transcriptional analysis. Granzyme K (gzmK) was coexpressed with gzmA, whereas Granzyme B (gzmB) expression was found to precede the expression of both gzmA and gzmK in 15-day cultures. Temporal gene expression patterns for seven blood samples demonstrate that most genes are expressed by all patient samples in similar temporal patterns. However, several patient-specific gene clusters were identified, and one cluster was found to correlate well with cell proliferation and may potentially be used to predict patient-specific T-cell expansion. PMID- 15286981 TI - Mechanisms of biosorption of different heavy metals by brown marine macroalgae. AB - The biosorption mechanisms of different heavy metallic cations (Cd, Ni, Pb) to active chemical groups on the cell wall matrix of the nonliving brown marine macroalga, Sargassum vulgaris in its natural form, were examined by the following instrumental and chemical techniques: Fourier-transform infrared (FTIR) analysis, X-ray photoelectron spectroscopy (XPS), scanning electron microscopy (SEM), and extraction of alginic acid and sulfated polysaccharides, which act as metal binding moieties present in cell wall. From the different techniques used and the known chemical composition of the algal cell wall, it was observed that biosorption of the metallic cations to the algal cell wall component was a surface process. The binding capacities of the different metal cations were between 1 and 1.2 mmol metal/g on a dry weight basis. The main chemical groups involved in the metallic cation biosorption were apparently carboxyl, amino, sulfhydryl, and sulfonate. These groups were part of the algal cell wall structural polymers, namely, polysaccharides (alginic acid, sulfated polysaccharides), proteins, and peptidoglycans. The main cadmium cation sequestration mechanism by the algal biomass was apparently chelation, while the nickel cation sequestration mechanism was mainly ion exchange. Lead cations exhibit higher affinity to the algal biomass, and their binding mechanism included a combination of ion exchange, chelation, and reduction reactions, accompanied by metallic lead precipitation on the cell wall matrix. During the ion exchange process, calcium, magnesium, hydrogen cations, and probably other cations (sodium and potassium) in the algal cell wall matrix were replaced by the tested heavy metals. PMID- 15286982 TI - Increased production of Bacillus keratinase by chromosomal integration of multiple copies of the kerA gene. AB - To increase the production of keratinase, stable strains of Bacillus licheniformis carrying multiple keratinase gene copies in the chromosome were developed. Integrative vectors carrying kerA with or without P43-promoter were constructed and subcloned into B. licheniformis T399D and Bacillus subtilis DB104. In T399D, multiple copies of kerA integration into the chromosome were identified and determined by Southern blot. The optimal integration of kerA was found in the range of 3-5 copies. Higher integration of gene copies (>5) caused reduced processing and secretion of the extracellular keratinase. In DB104, kerA was cloned in the plasmid, not integrated into the chromosome. The strong constitutive promoter P43 not only increased the keratinase production in plasmid based expression in DB104 but also improved the enzyme yield of the integrants of T399D. New strains were able to enhance cell growth and enzyme yield at higher concentrations of medium substrate. When they were grown in either soy or feather medium, the keratinase activity was stable and improved by about 4-6 times. PMID- 15286983 TI - Substrate replenishment extends protein synthesis with an in vitro translation system designed to mimic the cytoplasm. AB - Cytoplasmic mimicry has recently led to the development of a novel method for cell-free protein synthesis called the "Cytomim" system. In vitro translation with this new system produced more than a 5-fold yield increase of chloramphenicol acetyl transferase (CAT) relative to a conventional method using pyruvate as an energy substrate. Factors responsible for activating enhanced protein yields, and causes leading to protein synthesis termination have been assessed in this new system. Enhanced yields were caused by the combination of three changes: growing the extract source cells on 2x YTPG media versus 2x YT, replacing polyethylene glycol with spermidine and putrescine, and reducing the magnesium concentration from conventional levels. Cessation of protein synthesis was primarily caused by depletion of cysteine, serine, CTP, and UTP. Substrate replenishment of consumed amino acids, CTP, and UTP extended the duration of protein synthesis to 24 h in fed-batch operation and produced 1.2 mg/mL of CAT. By also adding more T7 RNA polymerase and plasmid DNA, yields were further improved to 1.4 mg/mL of CAT. These results underscore the critical role that nucleotides play in the combined transcription-translation reaction and highlight the importance of understanding metabolic processes influencing substrate depletion. PMID- 15286984 TI - Continuous glucose monitoring and control in a rotating wall perfused bioreactor. AB - A glucose control system consisting of a single in-line glucose sensor, concentrated glucose solution, and computer hardware and software were developed. The system was applied to continuously control glucose concentrations of a perfusion medium in a rotating wall perfused vessel (RWPV) bioreactor culturing BHK-21 cells. The custom-made glucose sensor was based on a hydrogen peroxide electrode. The sensor continuously and accurately measured the glucose concentration of GTSF-2 medium in the RWPV bioreactor during cell culture. Three sets of two-point calibrations were applied to the glucose sensor during the 55 day cell culture. The system first controlled the glucose concentration in perfusing medium between 4.2 and 5.6 mM for 36 days and then at different glucose levels for 19 days. A stock solution with a high glucose concentration (266 mM) was used as the glucose injection solution. The standard error of prediction (SEP) for glucose measurement by the sensor, compared to measurement by the Beckman glucose analyzer, was +/-0.4 mM for 55 days. PMID- 15286985 TI - BioLogic gates enable logical transcription control in mammalian cells. AB - The architecture of gene regulatory networks is reminiscent of electronic circuits. Modular building blocks that respond in a logical way to one or several inputs are connected to perform a variety of complex tasks. Gene circuit engineers have pioneered the construction of artificial gene regulatory networks with the intention to pave the way for the construction of therapeutic gene circuits for next-generation gene therapy approaches. However, due to the lack of a critical amount of eukaryotic cell-compatible gene regulation systems, the field has so far been limited to prokaryotes. Recent development of several mammalian cell-compatible expression control systems laid the foundations for the assembly of transcription control modules that can respond to several inputs. Herein, three approaches to evoke combinatorial transcription control have been followed: (i) construction of artificial promoters with up to three operator sites for regulatory proteins, and (ii) parallel and (iii) serial linking of two gene regulation systems. We have combined tetracycline-, streptogramin-, macrolide-, and butyrolactone transcription control systems to engineer BioLogic gates of the NOT IF-, AND-, NOT IF IF-, NAND-, OR-, NOR-, and INVERTER-type in mammalian cells, which are able to respond to up to three different small molecule inputs. BioLogic gates enable logical transcriptional control in mammalian cells and, in combination with modern transduction technologies, could serve as versatile tools for regulated gene expression and as building blocks for complex artificial gene regulatory networks for applications in gene therapy, tissue engineering, and biotechnology. PMID- 15286986 TI - Growth-rate recovery of Escherichia coli cultures carrying a multicopy plasmid, by engineering of the pentose-phosphate pathway. AB - Expression of plasmid-encoded genes in bacteria is the most common strategy for the production of specific proteins in biotechnological processes. However, the synthesis of plasmid-encoded proteins and plasmid-DNA replication often places a metabolic load (metabolic burden) into the cell's biochemical capacities that usually reduces the growth rate of the producing culture (Glick BR. Biotechnol Adv 1995;13:247-261). This metabolic burden may be related to a limited capacity of the cell to supply the extra demand of building blocks and energy required to replicate plasmid DNA and express foreign multicopy genes. Some of these required blocks are intermediaries of the pentose phosphate (PP) pathway, e.g., ribose-5 phosphate, erythrose-4-phosphate. Due to the important impact of metabolic burden on biotechnological processes, several groups have worked on developing strategies to overcome this problem, like reduction of plasmid copy number (Seo JH, Bailey JE. Biotechnol Bioeng 1985;27:1668-1674; Jones KL, Kim S, Keasling JD. Metab Eng 2000;3:328-338), chromosomal insertion of the gene which product is desired, or changing the plasmid-coded antibiotic resistance gene (Hong Y, Pasternak JJ, Glick BR. Can J Microbiol 1995;41:624-628). However, few efforts have been attempted to overcome the reduction of growth rate due to protein over expression, by modifying central metabolic pathways (Chou C-H, Bennett GN, San KY. Biotechnol Bioeng 1994;44:952-960). We constructed a high-copy number plasmid carrying the gene for glucose-6-phosphate dehydrogenase, zwf, under the control of an inducible trc promoter (pTRzwf04 plasmid). By transforming a wild-type strain and inducing with IPTG, it was possible to recover growth-rate from 0.46 h(-1) (uninduced) to 0.64 h(-1) (induced). The same transformation in an Escherichia coli zwf(-), allows a growth-rate recovery from 0.43 h(-1) (uninduced) to 0.62 h(-1) (induced). We also studied this effect as part of a laboratory-scale biotechnology process: production of a recombinant insulin peptide by co-transforming E. coli JM101 strain with pTRzwf07, a low-copy-number plasmid that carries the same inducible construction as pTRzwf04, and with the pTEXP-MMRPI vector that carries a TrpLE-proinsulin hybrid gene. In this system, production of TrpLE-proinsulin strongly reduces growth rate; however, overexpression of zwf gene recovers with a growth rate from 0.1 h(-1) in the TrpLE-proinsulin induced strain, to 0.37 h(-1) when both zwf and TrpLE-proinsulin genes were induced. In this paper, we show that the engineering of the pentose phosphate pathway by modulation of the zwf gene expression level partially overcomes the possible bottleneck for the supply of building blocks and reducing power synthesized through the PP pathway, that are required for plasmid replication and plasmid-encoded protein expression. PMID- 15286987 TI - Potato flour viscosity improvement is associated with the expression of a wheat LMW-glutenin gene. AB - It has been previously shown that expression of a high-molecular-weight glutenin (HMW-GS) in transgenic wheat seeds resulted in the improvement of flour functional properties. In this study, potato flour viscosity was improved through a specific expression of a low-molecular-weight glutenin (LMW-GS-MB1) gene in tuber. The resulting construct was introduced into potato leaf explants (Solanum tuberosum cv Kennebec) through Agrobacterium tumefaciens-mediated gene transfer. Southern and Northern analysis of transgenic potato confirmed that the integration of LMW-GS-MB1 in genomic DNA was stable and its mRNA was abundant in transgenic line 16 tubers. Western blot analysis of line 16 extract shows a LMW GS subunit accumulation in tuber. To demonstrate the capacity of transgenic lines to produce tubers with improved flour functional properties, transgenic lines 9 and 16 exhibiting, respectively, moderate and high expression of LMW-GS-MB1 mRNA and nontransgenic plants were transferred to field plots. The mean viscosity value of flour obtained from the field-grown tubers of transgenic line 16 exhibited a 3-fold increase in viscosity at 23 degrees C when compared to flour from nontransgenic tubers. PMID- 15286988 TI - Empirical modeling of batch fermentation kinetics for poly(glutamic acid) production and other microbial biopolymers. AB - An empirical kinetic model is proposed for the batch production of poly(glutamic acid) from Bacillus subtilis IFO 3335. In addition, the proposed model was used to fit the kinetic data of poly(glutamic acid) production from other bacterial strains using different media, as well as kinetic data from different strains for the production of the exocellular biopolymers dextran, hyaluronic acid, xanthan, alginate, and the endocellular biopolymer polyhydroxybutyrate. The empirical model treats the biopolymer as a component of the biomass and fits the experimental biomass data using a sigmoidal relationship that includes the maximum specific growth rate, mu(max), and the substrate saturation parameter, K(S). An empirical parameter, the relative coefficient (r), quantifies, in relative terms, the degree of nongrowth-associated biopolymer formation. PMID- 15286989 TI - Metabolic engineering and protein directed evolution increase the yield of L phenylalanine synthesized from glucose in Escherichia coli. AB - L-phenylalanine (L-Phe) is an aromatic amino acid with diverse commercial applications. Technologies for industrial microbial synthesis of L-Phe using glucose as a starting raw material currently achieve a relatively low conversion yield (Y(Phe/Glc)). The purpose of this work was to study the effect of PTS (phosphotransferase transport system) inactivation and overexpression of different versions of feedback inhibition resistant chorismate mutase-prephenate dehydratase (CM-PDT) on the yield (Y(Phe/Glc)) and productivity of L-Phe synthesized from glucose. The E. coli JM101 strain and its mutant derivative PB12 (PTS(-)Glc(+) phenotype) were used as hosts. PB12 has an inactive PTS, but is capable of transporting and phosphorylating glucose by using an alternative system constituted by galactose permease (GalP) and glucokinase activities (Glk). JM101 and PB12 were transformed with three plasmids, harboring genes that encode for a feedback inhibition resistant DAHP synthase (aroG(fbr)), transketolase (tktA) and either a truncated CM-PDT (pheA(fbr)) or its derived evolved genes (pheA(ev1) or pheA(ev2)). Resting-cells experiments with these engineered strains showed that JM101 and PB12 strains expressing either pheA(ev1) or pheA(ev2) genes produced l-Phe from glucose with Y(Phe/Glc) of 0.21 and 0.33 g/g, corresponding to 38 and 60% of the maximum theoretical yield (0.55 g/g), respectively. In addition, in both engineered strains the reached q(Phe) high levels of 40 mg/g dcw.h. The metabolic engineering strategy followed in this work, including a strain with an inactive PTS, resulted in a positive impact over the Y(Phe/Glc), enhancing it nearly 57% compared with its PTS(+) counterpart. This is the first report wherein PTS inactivation was a successful strategy to improve the Y(Phe/Glc). PMID- 15286990 TI - A novel convenient procedure for extractive work-up of whole-cell biotransformations using de-emulsifying hydrolases. AB - Extractive work-up of whole-cell biotransformations generally suffers from the formation of stable gels and slimes upon addition of the organic solvent to the cell suspension and the cell-free solution, respectively. This problem has been overcome by enzymatic lysis of emulsifying agents present in the medium through addition of hydrolases. Of these agents, proteases have exhibited the most powerful de-emulsifying activity. Enzyme treatment of cell-free culture media of Saccharomyces cerevisiae with pronase E drastically reduced phase separation time (t(p)) from 1 week to 30 min without significantly affecting product integrity. Yeast cell suspensions were de-emulsified best with protease N-01, where phase separation was complete after 1 h. As was exemplified with cell-free culture media of Lactobacillus kefir, wherein addition of pronase E or protease N-01 reduced t(p) from 1 week to 2 h each, this practical, ready-to-use method is appropriate for both fungal and bacterial biocatalysts. PMID- 15286991 TI - Serum-free large-scale transient transfection of CHO cells. AB - To date, methods for large-scale transient gene expression (TGE) in cultivated mammalian cells have focused on two transfection vehicles: polyethylenimine (PEI) and calcium phosphate (CaPi). Both have been shown to result in high transfection efficiencies at scales beyond 10 L. Unfortunately, both approaches yield higher levels of recombinant protein (r-protein) in the presence of serum than in its absence. Since serum is a major cost factor and an obstacle to protein purification, our goal was to develop a large-scale TGE process for Chinese hamster ovary (CHO) cells in the absence of serum. CHO-DG44 cells were cultivated and transfected in a chemically defined medium using linear 25 kDa PEI as a transfection vehicle. Parameters that were optimized included the DNA amount, the DNA-to-PEI ratio, the timing and solution conditions for complex formation, the transfection medium, and the cell density at the time of transfection. The highest levels of r-protein expression were observed when cultures at a density of 2.0 x 10(6) cells/ml were transfected with 2.5 microg/ml DNA in RPMI 1640 medium containing 25 mM HEPES at pH 7.1. The transfection complex was formed at a DNA:PEI ratio of 1:2 (w/w) in 150 mM NaCl with a 10-min incubation at room temperature prior to addition to the culture. The procedure was scaled up for a 20-L bioreactor, yielding expression levels of 10 PMID- 15286992 TI - Multienzyme mevalonate pathway bioreactor. AB - The five-carbon metabolic intermediate isopentenyl diphosphate constitutes the basic building block for the biosynthesis of all isoprenoids in all forms of life. Two distinct pathways lead from amphibolic intermediates to isopentenyl diphosphate. The Gram-positive cocci and certain other pathogenic bacteria employ exclusively the mevalonate pathway, a set of six enzyme-catalyzed reactions that convert 3 mol of acetyl-CoA to 1 mol each of carbon dioxide and isopentenyl diphosphate. The survival of the Gram-positive cocci requires a fully functional set of mevalonate pathway enzymes. These enzymes therefore represent potential targets of inhibitors that might be employed as antibiotics directed against multidrug-resistant strains of certain bacterial pathogens. A rapid throughput, bioreactor-based assay to assess the effects of potential inhibitors on several enzymes simultaneously should prove useful for the survey of candidate inhibitors. To approach this goal, and as a proof of concept, we employed enzymes from the Gram-positive pathogen Enterococcus faecalis. Purified recombinant enzymes that catalyze the first three reactions of the mevalonate pathway were immobilized in two kinds of continuous flow enzyme bioreactors: a classical hollow fiber bioreactor and an immobilized plug flow bioreactor that exploited a novel method of enzyme immobilization. Both bioreactor types employed recombinant acetoacetyl-CoA thiolase, HMG-CoA synthase, and HMG-CoA reductase from E. faecalis to convert acetyl-CoA to mevalonate, the central intermediate of the mevalonate pathway. Reactor performance was monitored continuously by spectrophotometric measurement of the concentration of NADPH in the reactor effluent. Additional potential applications of an Ni(++) affinity support bioreactor include using recombinant enzymes from extremophiles for biosynthetic applications. Finally, linking a Ni(++) affinity support bioreactor to an HPLC mass spectrometer would provide an experimental and pedagogical tool for study of metabolite flux and pool sizes of intermediates to model regulation in intact cells. PMID- 15286993 TI - Study of dye decolorization in an immobilized laccase enzyme-reactor using online spectroscopy. AB - Decolorization of textile dyes by a laccase from Trametes modesta immobilized on gamma-aluminum oxide pellets was studied. An enzyme reactor was equipped with various UV/Vis spectroscopic sensors allowing the continuous online monitoring of the decolorization reactions. Decolorization of the dye solutions was followed via an immersion transmission probe. Adsorption processes were observed using diffuse reflectance measurements of the solid carrier material. Generally, immobilization of the laccase does not seem to sterically affect dye decolorization. A range of commercial textile dyes was screened for decolorization and it was found that the application of this enzymatic remediation system is not limited to a certain structural group of dyes. Anthrachinonic dyes (Lanaset Blue 2R, Terasil Pink 2GLA), some azo dyes, Indigo Carmine, and the triphenylmethane dye Crystal Violet were efficiently decolorized. However, the laccase displayed pronounced substrate specificities when a range of structurally related model azodyes was subjected to the biotransformation. Azodyes containing hydroxy groups in ortho or para position relative to the azo bond were preferentially oxidized. The reactor performance was studied more closely using Indigo Carmine. PMID- 15286994 TI - Transposon vectors for gene-trap insertional mutagenesis in vertebrates. AB - The function of most vertebrate genes remains unknown or uncertain. Insertional mutagenesis offers one approach to identify and understand the function of these genes. Transposons have been used successfully in lower organisms and plants for insertional mutagenesis, but until activation of the Sleeping Beauty (SB) transposon system, there was no indication of active DNA-based transposons in vertebrates. Investigator-driven insertional mutagenesis in vertebrates has relied on retroviral insertions or selection of low-frequency integration of naked DNA in ES cell lines. We have combined the highly active SB transposon with gene-trapping technology to demonstrate that transposon traps can be used for insertional mutagenesis screens in vertebrates. In our studies about one-fourth of the trap insertions appear to be in transcriptional units, a rate that is commensurate with random integration. We show that gene-traps coupled to a fluorescent protein reporter gene can be used to detect insertions into genes active in specific cells of living zebrafish embryos, supporting use of our transposon traps for high-throughput functional genomic screens in vertebrates. PMID- 15286995 TI - Floxed allele for conditional inactivation of the voltage-gated sodium channel Scn8a (NaV1.6). AB - The sodium channel gene Scn8a encodes the channel NaV1.6, which is widely distributed in the central and peripheral nervous system. NaV1.6 is the major channel at the nodes of Ranvier in myelinated axons. Mutant alleles of mouse Scn8a result in neurological disorders including ataxia, tremor, paralysis, and dystonia. We generated a floxed allele of Scn8a by inserting loxP sites around the first coding exon. The initial targeted allele containing the neo-cassette was a severe hypomorph. In vivo deletion of the neo-cassette by Flp recombinase produced a floxed allele that generates normal expression of NaV1.6 protein. Ubiquitous deletion of the floxed exon by Cre recombinase in ZP3-Cre transgenic mice produced the Scn8a(del) allele. The null phenotype of Scn8a(del) homozygotes confirms the in vivo inactivation of Scn8a. Conditional inactivation of the floxed allele will make it possible to circumvent the lethality that results from complete loss of Scn8a in order to investigate the physiologic role of NaV1.6 in subpopulations of neurons. PMID- 15286996 TI - Refining GAL4-driven transgene expression in Drosophila with a GAL80 enhancer trap. AB - We constructed an enhancer-trap element, P[GAL80], that encodes the yeast GAL80 repressor to refine expression of transgenes driven by the binary GAL4/UAS system. GAL80 blocks GAL4 activity by binding to its transcriptional activation domain. We screened GAL80 enhancer-traps for repression of GAL4-induced green fluorescent protein (GFP) in the intact larval nervous system. We selected one line that repressed GFP in a large set of cholinergic neurons. This line was used to refine GFP expression from a set of over 200 neurons to a subset of 20 neurons in a preselected GAL4 line. Expression of tetanus neurotoxin, a potent blocker of neurotransmitter release, in these 20 neurons reproduced an aberrant larval turning behavior previously assigned to the parental set of 200 neurons. Our results suggest that targeted GAL80 expression could become a useful means of spatially refining transgene expression in Drosophila. PMID- 15286997 TI - Control of bract formation in Drosophila: poxn, kek1, and the EGF-R pathway. AB - In Drosophila, the sensory organs are formed by cells that derive from a precursor cell through a fixed lineage. One exception to this rule is the bract cell that accompanies some of the adult bristles. The bract cell is derived from the surrounding epidermis and is induced by the bristle cells. On the adult tibia, bracts are associated with all mechanosensory bristles, but not with chemosensory bristles. The differences between chemosensory and mechanosensory lineages are controlled by the selector gene pox-neuro (poxn). Here we show that poxn is also involved in suppressing bract formation near the chemosensory bristles. We have identified the gene kek1, described as an inhibitor of the EGF R signaling pathway, in a screen for poxn downstream genes. We show that kek1 can suppress bract formation and can interfere with other steps of sensory development, including SMC determination and shaft differentiation. PMID- 15286998 TI - Efficient DNA cassette exchange in mouse embryonic stem cells by staggered positive-negative selection. AB - Recombinase-mediated cassette exchange (RMCE), when applied to mouse embryonic stem (ES) cells, promises to increase the ease with which genetic alterations can be introduced into targeted genomic loci in the mouse. However, existing selection strategies for identifying ES cells in which replacement DNA cassettes from a carrier plasmid have been exchanged correctly into a defined locus are suboptimal. Here, we report the generation in mouse ES cells of a loxed cassette acceptor (LCA) allele within the glucokinase (gk) gene locus. Using the gkLCA as a test allele, we developed a staggered positive-negative selection strategy that facilitates efficient identification of ES cell clones in which a DNA replacement cassette from a carrier plasmid has been exchanged correctly into the gkLCA allele. This selection strategy, by facilitating more efficient production of ES cell clones with various replacement DNA cassettes, should accelerate targeted repetitive introduction of gene modifications into the mouse. PMID- 15286999 TI - New class of Son-of-sevenless (Sos) alleles highlights the complexities of Sos function. AB - The guanine nucleotide exchange factor (GEF) Son-of-sevenless (Sos) encodes a complex multidomain protein best known for its role in activating the small GTPase RAS in response to receptor tyrosine kinase (RTK) stimulation. Much less well understood is SOS's role in modulating RAC activity via a separate GEF domain. In the course of a genetic modifier screen designed to investigate the complexities of RTK/RAS signal transduction, a complementation group of 11 alleles was isolated and mapped to the Sos locus. Molecular characterization of these alleles indicates that they specifically affect individual domains of the protein. One of these alleles, SosM98, which contains a single amino acid substitution in the RacGEF motif, functions as a dominant negative in vivo to downregulate RTK signaling. These alleles provide new tools for future investigations of SOS-mediated activation of both RAS and RAC and how these dual roles are coordinated and coregulated during development. PMID- 15287000 TI - Transcription cofactor Vgl-2 is required for skeletal muscle differentiation. AB - TEF-1 transcription factors regulate gene expression in skeletal muscle but are not muscle-specific. Instead, TEF-1 factors rely on the muscle-specific cofactor Vestigial-like 2 (Vgl-2), a protein related to Drosophila vestigial. Previously, we showed that Vgl-2 promotes skeletal muscle differentiation and activates muscle-specific promoters. However, the mechanism whereby Vgl-2 regulates TEF-1 factors and the requirement for Vgl-2 for muscle-specific gene expression were not known. In Drosophila, vestigial alters DNA binding specificity of the TEF-1 homolog scalloped to drive wing and flight muscle-specific gene expression. Here, gel mobility shift assays show that Vgl-2 differentially affects DNA binding of different TEF-1 factors. Using an antisense morpholino, we blocked the expression of Vgl-2 and a muscle-specific gene in the myogenic C2C12 cell line and in chick embryos by electroporation. These results demonstrate that Vgl-2 is required for muscle gene expression, in part by switching DNA binding of TEF-1 factors during muscle differentiation. PMID- 15287001 TI - Localization of swallow-Green Fluorescent Protein in Drosophila oogenesis and implications for the role of swallow in RNA localization. AB - The localization of a hybrid protein composed of swallow and Green Fluorescent Protein (GFP) during Drosophila oogenesis is reported. I constructed a hybrid gene with GFP inserted into an internal position of swallow. This gene was integrated into the Drosophila genome and provides full swallow+ function, as assayed by the complete rescue of strong swallow mutants. Swallow-GFP is localized at all points along the oocyte cortex from vitellogenic stages of oogenesis through the end of oogenesis. Higher concentrations of swallow-GFP are present at the anterior oocyte cortex than at the lateral and posterior oocyte cortices at Stages 10 and 11, when bicoid and htsN4 mRNA transport from nurse cells and localization in the oocyte are most active. At Stage 9 and at Stages 12 14 swallow-GFP is equally distributed at the anterior, lateral, and posterior oocyte cortices. The position of swallow-GFP in vitellogenic stages is identical to the position of endogenous swallow protein determined by indirect immunofluorescence using an anti-swallow antibody. At the oocyte cortex, swallow GFP is present in particulate structures that lie within or just internal to the dense cortical actin meshwork. These particles show little or no movement, suggesting that they are attached to or embedded in the oocyte cortex. These observations are most easily interpreted in the context of mRNA anchoring or microtubule organizing functions for the swallow protein. PMID- 15287002 TI - Hypervolemic hypertension in mice with systemic inactivation of the (floxed) guanylyl cyclase-A gene by alphaMHC-Cre-mediated recombination. AB - To dissect the tissue-specific functions of atrial natriuretic peptide (ANP), we recently introduced loxP sites into the murine gene for its receptor, guanylyl cyclase-A (GC-A), by homologous recombination (tri-lox GC-A). For either smooth muscle or cardiomyocyte-restricted deletion of GC-A, floxed GC-A mice were mated to transgenic mice expressing Cre-recombinase under the control of the smooth muscle SM22 or the cardiac alphaMHC promoter. As shown in these studies, Cre mediated recombination of the floxed GC-A gene fully inactivated GC-A function in a cell-restricted manner. In the present study we show that alphaMHC-Cre, but not SM22-Cre, with high frequency generates genomic recombinations of the floxed GC-A gene segments which were transmitted to the germline. Alleles with partial or complete deletions were readily recovered from the next generation, after segregation of the Cre-transgene. We took advantage of this strategy to generate a new mouse line with global, systemic deletion of GC-A. Doppler echocardiographic and physiological studies in these mice demonstrate for the first time the tremendous impact of ANP/GC-A dysfunction on chronic blood volume homeostasis. PMID- 15287003 TI - Chemiluminescence detection of Escherichia coli in fresh produce obtained from different sources. AB - A chemiluminescence-based assay is developed for the rapid detection of Escherichia coli in fresh produce. The assay was based on the reaction of beta galactosidase enzyme from E. coli with a phenylgalactosidase-substituted dioxetane substrate. Light emitted from the reaction was measured in a luminometer and data correlated with counts of E. coli enumerated on sorbitol MacConkey agar plates. A strain of E. coli O157:H7 was used to inoculate samples of fresh produce to differentiate the inoculum from the natural E. coli potentially present on the produce. Fresh market samples were tested for generic E. coli and E. coli O157:H7. Significant differences in light emission were found in samples with high initial E. coli counts when market samples were compared to respective heat-treated samples. The assay was able to detect E. coli in all produce tested, particularly at higher contamination or inoculation levels. The sensitivity of the assay ranged between 10(2)-10(5) CFU within 30 min. The chemiluminescence assay provides a simple and rapid method for detection of viable E. coli, an important step towards enhancing food safety. PMID- 15287004 TI - Scavenger effect of flavonols on HOCl-induced luminol chemiluminescence. AB - Hypochlorous acid (HOCl), the main product of the myeloperoxidase system, is a strong oxidant and a potent chlorinating agent, which can damage host tissues. In the present work, the scavenger effect of three aglycone flavonols (myricetin, quercetin and kaempferol) and of the natural glycoside flavonol, rutin, was studied towards HOCl using luminol-dependent chemiluminescence (CL). At 1 micro mol/L fi nal concentration, rutin was the most powerful scavenger of HOCl with an inhibitory luminol oxidation of 91.4% +/- 3.2%. Quercetin, kaempferol and myricetin inhibited the luminol-dependent CL at the same concentration only by 75.9% +/- 3.4%, 57.7% +/- 5.3% and 43.3% +/- 3.5%, respectively. With increasing concentration of these flavonols, a dose-dependent inhibition of luminol CL was observed. In order to prove to what extent flavonols scavenge HOCl, their concentrations that gave 50% inhibition of luminescence (IC50) were compared to IC50 values of the sulphur-containing compounds N-acetyl cysteine (NAC) and taurine. The scavenging activities of compounds tested decrease in the order: rutin > NAC > quercetin > kaempferol > taurine. The present study revealed that rutin was the most effective scavenger agent. PMID- 15287005 TI - The emitting species formed by the oxidation of hydrazides with hypohalites or N halosuccinimides. AB - The chemiluminescence accompanying the oxidation of salicylic hydrazide (2 hydroxybenzoic acid hydrazide) with hypochlorite, hypobromite, N chlorosuccinimide, N-bromosuccinimide or hydrogen peroxide with cobalt(II) matched the photoluminescence emission of salicylic acid. In a related reaction, the oxidation of a mixture of isoniazid and ammonia, a synergistic effect was observed. The chemiluminescence spectrum for this reaction matches that accompanying the oxidation of the hydrazide, rather than the oxidation of ammonia. These results were used to assess mechanisms proposed by previous authors. PMID- 15287006 TI - A rapid screening method for the detection of viable spores in powder using bioluminescence. AB - A rapid diagnosis of a biological threat in a powder sample is important for fi rst responders who have to make decisions on-site. The present culture-based method does not provide timely results, which is a critical barrier for a quick response when a suspicious powder sample is found. The ATP bioluminescence method, combined with a heat shock, was investigated to determine the presence of spores in powder. The results show that only spore-containing powder samples provided a dramatic increase in the bioluminescence signal after the heat shock, which induces germination of the spores. Various conditions were tested to fi nd the most effective and rapid germination procedure. Elevated temperatures (37 degrees C and 50 degrees C) were more effective in germination than room temperature. At 50 degrees C, a double-strength germinant was more effective in germination than the regular strength. The 37 degrees C/15 min procedure induced the germination of spores most effectively, while a 50 degrees C/2 min procedure provided reasonably high signals, so it could make the entire procedure even faster (< 5 min). The detection limit of the bioluminescence method is < 100 spores. PMID- 15287007 TI - Multi-channel electrochemiluminescence of luminol at a copper electrode. AB - Multi-channel electrochemiluminescence (ECL) of luminol at a copper electrode has been studied under conventional cyclic voltammetric (CV) conditions. Compared with the ECL of luminol at other electrodes, three ECL peaks were observed at 0.30, -0.24 and -0.65 V (vs. SCE), respectively, which was also imaged by a CCD camera. The effects of potential scan direction, anodic reverse potential, the presence of N2 and O2 of the solution, the pH of the solution, the NaNO3 concentration and the potential scan rate were examined. The effect of n alkanethiol self-assembled monolayers on copper electrodes and 20 L-amino acids, dopamine, adrenaline and noradrenaline on the ECL of luminol were also investigated. The emission spectra of various ECL peaks at different potentials demonstrated that all ECL peaks were related to the luminol reaction. The results show that the oxygen dissolved in solution and copper oxide covered on the surface of the electrode play an important role in the luminol ECL process at a copper electrode. It has been proposed that three ECL channels of luminol at a copper electrode resulted from the reactions of luminol or luminol radical electrooxidized by luminol with various electrogenerated oxygen-containing species, such as O2, OOH- and copper oxides at different potentials. PMID- 15287008 TI - Potential of the luminol reaction in the sensitive detection of pesticide residues by flow injection analysis. AB - This study presents the first analytical application of the luminol chemiluminescence (CL) reaction for the sensitive detection of carbamate residues. Some experiments have been carried out to check the influence of the presence of traces of a N-methylcarbamate (carbaryl) on the CL emission produced from the oxidation of luminol using different oxidants, showing a significant enhancing effect on the CL emission when the oxidation of luminol is produced by potassium permanganate in alkaline medium, this enhancement being proportional to the carbaryl concentration. This fact has permitted the establishment of a sensitive chemiluminescence flow-injection (CL-FIA) method for the direct determination of carbaryl. The optimization of instrumental and chemical variables influencing the CL response has been carried out by applying experimental designs. Under the optimal conditions, the CL intensity was linear for a carbaryl concentration over the range 5-100 ng/mL with a detection limit of 4.9 ng/mL. This luminol-KMnO4-based FIA-CL system in basic medium shows an easy, fast and cheap alternative detection mode for the analysis of carbaryl residues in environmental water samples. PMID- 15287009 TI - Discarding multidrug resistance inducers, the possible role of a biosensing reporter in antimicrobial discovery. AB - The aim of the present work was to determine whether the luminescence-based reporter plasmid pQacLux could be applied to drug discovery in order to discard compounds with defined properties. Non-pathogenic Staphylococcus aureus RN4220 cells bearing pQacLux were incubated with different concentrations of a disinfectant of common use in hospitals. The in vivo light emission response of the plasmid to the given stimuli was then quantified and compared to a negative control for the construction of dose-response curves. The selected disinfectant provided a convenient model for the activity of quaternary ammonium compounds. In spite of the use of a raw model solution, the system revealed high levels of sensitivity. According to the results obtained, pQacLux could be conveniently used in the first steps of drug development in order to discard all possible multidrug resistance inducers. PMID- 15287010 TI - Luminescence literature. Green fluorescent protein (GFP). PMID- 15287013 TI - 3-D morphological characterization of the liver parenchyma by atomic force microscopy and by scanning electron microscopy. AB - A comparative study of atomic force microscopy (AFM) and scanning electron microscopy (SEM) imaging of the healthy human liver parenchyma was carried out to determine the similarities and the differences. In this study, we compared the fine hepatic structures as observed by SEM and AFM. Although AFM revealed such typical hepatic structures as bile canaliculi and hepatocytes, it also showed the location of the nucleus and chromatin granules in rough relief structure, which was not visible by SEM. By contrast, SEM visualized other structures, such as microvilli, the central vein, and collagenous fibers, none of which was visualized by AFM. For better orientation and confirmation of most of the structures imaged by SEM and AFM, Congo Red-stained specimens were also examined. Amyloid deposits in the Disse's spaces were shown especially clearly in these images. The differences between the SEM and AFM images reflected the characteristics of the detection systems and methods used for sample preparation. Our results reveal that more detailed information on hepatic morphology is obtained by exploiting the advantages of both SEM and AFM. PMID- 15287014 TI - Glycogen autophagy. AB - Glycogen autophagy, which includes the sequestration and degradation of cell glycogen in the autophagic vacuoles, is a selective process under conditions of demand for the massive hepatic production of glucose, as in the postnatal period. It represents a link between autophagy and glycogen metabolism. The formation of autophagic vacuoles in the hepatocytes of newborn animals is spatially and biochemically related to the degradation of cell glycogen. Many molecular elements and signaling pathways including the cyclic AMP/cyclic AMP-dependent protein kinase and the phosphoinositides/TOR pathways are implicated in the control of this process. These two pathways may converge on the same target to regulate glycogen autophagy. PMID- 15287015 TI - Differences in X-ray absorption due to cadmium treatment in Saponaria officinalis leaves. AB - A method for detecting cadmium uptake in leaves of Saponaria officinalis doped with a solution of cadmium acetate is described. The technique based on the exposure of dried leaves to X-rays of a wavelength close to that of the metal K edge could be useful for phytoremediation studies as it could reveal the bioaccumulation in plants due to the treatment either in vivo or in vitro with heavy metals. X-ray microradiography measurements are in agreement with those from peroxidase enzyme assay utilized to follow the oxidative damage induced by heavy metals. At present, as we will see in this report, microradiography has still poorer sensitivity in comparison with enzyme assay, but it has the advantage of being faster, not destructive, and usable even at very high doping levels, where the enzyme assay technique results are fully saturated. Further analysis of the optical density values could lead to a quantitative measurement of the heavy metal in the sample. Thus, the technology developed in this article could be useful for tracing the intake in phytoremediation studies. PMID- 15287016 TI - Expression of brain natriuretic peptide in the rat heart studies during heart growth and in relation to sympathectomy. AB - Brain natriuretic peptide (BNP) might be of importance during heart development and is described to be increasingly expressed in congestive heart failure and to affect the progress of this condition. However, details in the normal expression of BNP are still unclear in various parts of the adult and growing heart, including the conduction system. In this study, we investigated the expression of BNP in relation to that of atrial natriuretic peptide (ANP) in the growing as well as in the adult rat heart. The effects of chemical sympathectomy in adult rats were also examined. Contrary to previous BNP immunohistochemical studies, the BNP antiserum was preabsorbed with an excess of ANP before staining to abolish the crossreactivity with ANP. There was a pronounced BNP immunoreaction in the auricles, the trabeculated ventricular walls, and the peripheral parts of the conduction system at 0-1 days postnatally. The degree of immunoreaction gradually decreased with increasing age. A similar developmental pattern was seen concerning ANP expression, but the magnitude of the latter clearly exceeded that for BNP. Immunoreaction for BNP was never detected in the atrioventricular (AV) node and AV bundle at any stage. In contrast to the situation for ANP previously observed, no obvious changes in BNP immunoreaction patterns were observed in response to sympathectomy. This is the first study to thoroughly demonstrate the expression of BNP in the various regions of the rat heart during growth and in the normal and sympathectomized adult stage. The observations are related to possible functions of natriuretic peptides in the growing and adult heart. PMID- 15287017 TI - Confocal laser scanning microscope study of cytokeratin immunofluorescence differences between villous and extravillous trophoblast: cytokeratin downregulation in pre-eclampsia. AB - Pre-eclampsia is a disease characterized by failures in interstitial implantation. One product of the implantation process is the basal plate; a structure whose complexity makes it hard to fully appreciate the pathological changes in significant diseases of pregnancy. This article describes our use of CLSM immunofluorescence to examine the cytokeratin composition of the cells of trophoblastic origin in the term placental basal plate. Large differences in the content of the structural polymeric protein were compared using analysis of digital images. We show that greater pancytokeratin immunofluorescence is observed in extravillous cytotrophoblast cells as compared with villous trophoblast. There is a >30-fold difference in the mean area percent of the most intensely immunofluorescent pixels in the tissue containing these cells. This is a very high, statistically significant difference as defined by the Wilcoxon Signed Ranks Test Asym. Sig. (two-tailed): P < 0.001. The most invasive population of cells of the trophoblast lineage (the extravillous trophoblast) exhibits a significant reduction in cytokeratin immunofluorescence when comparisons of healthy and pre-eclamptic pregnancies are made. This ratio was 2.4:1. It was tested using the Mann-Whitney U-test. From healthy to pre-eclamptic the reduction was from mean rank 83.42((healthy)) to 51.13((pre-eclamptic)). The difference was very highly statistically significant (n = 53 + 75 = 128; U = 984.500; Z = -4.852; P < 0.001). There was also less cytokeratin-related immunofluorescence in villous trophoblast when healthy villi were compared with pre-eclamptic villi. The observed alterations in trophoblastic cytoskeletal components are expected to damage the anchorage and motility of cells. The extravillous trophoblast is known to be necessary for implantation. This leads to a cellular hypothesis of the failure of implantation resulting in reduced depth of uterine invasion and failure to adapt the spiral arterioles for low-pressure perfusion of the intervillus space, two well-known features of pre-eclampsia. The reduction in cytokeratin-related immunofluorescence in the villus trophoblast seen on comparing healthy term placentae with those from pre-eclamptics implies that the trophoblast is a weaker epithelial layer in the hypertensive pregnancy. This could account for the rise in deported trophoblast associated with pre eclampsia. Deported trophoblast has been invoked as the systemic messenger that leads to generalized maternal hypertension seen in this condition. PMID- 15287018 TI - Healthy and pre-eclamptic placental basal plate lining cells: quantitative comparisons based on confocal laser scanning microscopy. AB - Immunocytochemical confocal laser scanning microscope images of the monolayer of cells lining the intervillus space at the basal plate of term placentae were analysed using stereology. Immunoreactively-distinct regions of this mosaic layer were measured. In basal plate from healthy pregnancies, trophoblast epithelium occupied 18.91% of the surface area and endothelium 60.81%. In pre-eclampsia the equivalent areas were 15.57% and 67.63%. Acellular fibrinoid covers the remaining area and this component decreases in area in pre-eclampsia. The statistically significant increase in the cellular endothelial compartment may be relevant to the hypertensive pathology of pre-eclampsia as endothelial signalling plays a major role in regulation of blood pressure. PMID- 15287019 TI - Glycoconjugates of the urodele amphibian testis shown by lectin cytochemical methods. AB - Lectin histochemistry is a useful method that allows the in situ identification of the terminal sugar moieties of the carbohydrates that form the glycoconjugates. Moreover, when it is combined with chemical or enzymatic deglycosylation pretreatments, lectin histochemistry can be employed to determine if carbohydrates are linked to the protein core by means of an N- or O-glycosidic linkage or, indeed, to partially sequence the sugar chains. One of the most interesting model organs for the study of spermatogenesis is the amphibian urodele testis. However, this organ has not been very widely investigated with lectin histochemical research. In the last few years, we have carried out a research project to identify and locate glycoconjugates in the testis of the urodele Pleurodeles waltl, the Spanish newt, as a first approach to identify possible carbohydrates with key roles in spermatogenesis. Our findings reveal some glycan chains located in a fusome-like structure in early (diploid) germ cells, oligosaccharides with terminal GalNAc in the acrosome, the occurrence of glycan modifications in the acrosomal contents during spermiogenesis, and changes in glycan composition of follicle and interstitial cells during the spermatogenetic cycle. Furthermore, the similar labeling pattern of follicle and duct cells supports the hypothesis for a common origin of both cell types. PMID- 15287020 TI - Ultrastructural study of apoptotic U937 [corrected] cells treated with different pro-apoptotic substances. AB - Human monocytic leukemia U937 cells readily undergo apoptosis when exposed to various stimuli, including inhibition of protein synthesis, oxidative stress, antitumoral agents, etc. The sequential, step-by-step morphological changes in U937 cells that occur during the apoptotic program are largely determined by the activation of a specific class of proteases, the caspases. The action of these proteases were followed at the ultrastructural level. From our observations 1) no unique morphological feature exists during apoptosis, even in the same cell type; 2) the extent of the morphological modifications are inducer- and dose-dependent; 3) double or triple treatments amplify the morphological modifications with a single inducer, but not the rate of apoptosis; and 4) in the case of double treatment the second inducer has to have a cytoplasmatic target because damage to the cytoplasm occurs before nuclear modifications become visible. These data should facilitate a more objective evaluation of apoptosis in conditions where antiproliferative drugs, like antiblastic or immunosuppressive molecules, are used to monitor the efficiency of treatment. PMID- 15287021 TI - Analysis of somatic APC mutations in rare extracolonic tumors of patients with familial adenomatous polyposis coli. AB - Patients with familial adenomatous polyposis coli (FAP) carry heterozygous mutations of the APC gene. At a young age, these patients develop multiple colorectal adenomas that consistently display a second somatic mutation in the remaining APC wild-type allele. Inactivation of APC leads to impaired degradation of beta-catenin, thereby promoting continuous cell-cycle progression. The role of APC inactivation in rare extracolonic tumors of FAP patients has not been characterized sufficiently. Among tissue specimen from 174 patients with known APC germ-line mutations, we identified 8 tumors infrequently seen in FAP. To investigate the pathogenic role of APC pathway deregulation in these lesions, they were analyzed for second-hit somatic mutations in the mutational cluster region of the APC gene. Immunohistochemistry was performed to compare the expression pattern of beta-catenin to the mutational status of the APC gene. Exon 3 of the beta-catenin gene (CTNNB1) was analyzed for activating mutations to investigate alternative mechanisms of elevated beta-catenin concentration. Although CTNNB1 mutations were not observed, second somatic APC mutations were found in 4 of the 8 tumors: a uterine adenocarcinoma, a hepatocellular adenoma, an adrenocortical adenoma, and an epidermal cyst. These tumors showed an elevated concentration of beta-catenin. No APC mutations were seen in focal nodular hyperplasia of the liver, angiofibrolipoma, and seborrheic wart. This is the first study reporting second somatic APC mutations in FAP-associated uterine adenocarcinoma and epidermal cysts. Furthermore, our data strengthen a role for impaired APC function in the pathogenesis of adrenal and hepatic neoplasms in FAP patients. PMID- 15287022 TI - Mutational activation of the MAP3K8 protooncogene in lung cancer. AB - The MAP3K8 protooncogene (Cot/Tpl-2) activates the MAP kinase, SAP kinase, and NF kappaB signaling pathways. MAP3K8 mutations occur in the rat homologue, but activating mutations have yet to be identified in primary human tumors. We have identified MAP3K8 as a transforming gene from a human lung adenocarcinoma and characterized a 3' end mutation in the cDNA. In addition, we confirmed that the mutation occurs in the original lung tumor, and we screened a series of lung cancer cell lines to determine whether the MAP3K8 mutation is a common occurrence in lung tumorigenesis. The oncogene was isolated and identified with the NIH3T3 nude mouse tumorigenicity assay and cDNA library screening. The gene was analyzed by polymerase chain reaction (PCR), single-strand conformational polymorphism (SSCP), and 3'RACE for mutations. The mutation was localized to MAP3K8 exon 8 and confirmed in the primary tumor DNA. Both wild-type and mutant MAP3K8 cDNAs transformed NIH3T3 cells, but the transforming activity of the mutant was much greater than that of the wild type. PCR-SSCP screening of cell line cDNAs identified one silent polymorphism in cell line SK-LU-1. Although we were unable to find additional activating mutations, these data support a role for MAP3K8 activity in cellular transformation, but suggest that mutational activation of the gene is a rare event in lung cancer. PMID- 15287023 TI - Different mechanisms of chromosome 16 loss of heterozygosity in well- versus poorly differentiated ductal breast cancer. AB - Loss of heterozygosity (LOH) at the long arm of chromosome 16 is a frequent genetic alteration in breast cancer. It can occur by physical loss of part of or the entire chromosomal arm, resulting in a decrease in copy number or loss followed by mitotic recombination. Comparative genomic hybridization (CGH) demonstrated that well-differentiated breast tumors showed significantly more physical loss of 16q than did poorly differentiated ones and that this difference was already discernable in the preinvasive stage. However, polymorphic markers detected no difference in the frequency of 16q LOH between invasive tumors of different histological grade. Here, by combining data on LOH (n=52), fluorescence in situ hybridization (n=18) with chromosome 16-specific probes, and CGH (n=34), we show a preference in well-differentiated grade I tumors for physical loss of chromosome arm 16q, whereas in poorly differentiated grade III tumors LOH is accompanied by mitotic recombination. This clarifies the discrepancies observed between CGH and LOH for 16q in breast cancer. These different somatic genetic mechanisms may reflect the presence of multiple tumor suppressor genes that are the target of LOH at chromosome arm 16q. PMID- 15287024 TI - Promoter methylation of the PTEN gene is a common molecular change in breast cancer. AB - About 25-50% of women with Cowden disease, a syndrome associated with germ-line mutations of the PTEN gene (at 10q23), develop breast cancer (BC), but PTEN mutations have been found in only 5% of sporadic BCs. However, 29-48% of BCs display loss of heterozygosity in 10q23, and about 40% of BCs show a decrease or absence of PTEN protein levels at the time of diagnosis. Promoter hypermethylation has been identified as an alternative mechanism of tumor suppressor gene inactivation, but its importance in PTEN silencing in sporadic BC is unknown. We investigated PTEN promoter hypermethylation in 90 sporadic BCs and its correlations with 11 molecular and pathologic parameters, including mRNA levels of PTEN. The study, a methylation-specific PCR assay, was carried out with methylated specific primers designed in a region with scarce homology with the psiPTEN pseudogene. Expression was analyzed by real-time PCR. We found that the PTEN promoter was hypermethylated in 43 BCs (48%). PTEN hypermethylation was associated with ERBB2 overexpression, larger size, and higher histologic grade (P=0.012, 0.03, and 0.009, respectively). We concluded that PTEN promoter hypermethylation is a common event in sporadic BC, correlating with other well established prognostic factors of this malignancy. Additionally, PTEN mRNA expression was lower in tumors with aberrant methylation. PMID- 15287025 TI - Alternative mechanisms of gene amplification in human cancers. AB - Gene amplification is a common phenomenon in cancer. Cytogenetic analyses have indicated that breakage-fusion-bridge (BFB) cycles drive intrachromosomal amplification of some oncogenes in a head-to-head manner in human cancers. However, the complex structures of an amplified sequence found in cancers are not always explained by the BFB model. At the 17q21 locus, which is not linked to common fragile sites, we discovered a recombination hot spot harboring amplicon repeats in tandem in a head-to-tail orientation, with the interamplicon junctions in each cancer cell being homogeneous. These findings clearly show the presence of alternative mechanisms other than BFB cycles in oncogene amplification. PMID- 15287026 TI - Spontaneously immortalized human T lymphocytes develop gain of chromosomal region 2p13-24 as an early and common genetic event. AB - To gain further insight into the molecular events responsible for the extended life span and immortalization of human lymphoid cells, we analyzed a series of spontaneously immortalized, IL2-dependent human T-cell lines using molecular cytogenetic techniques. Two of the cell lines were derived from normal spleen and three from patients with Nijmegen breakage syndrome (NBS), a recessive disorder characterized by a high incidence of lymphoid malignancies. Here we show that spontaneous immortalization of the five T-cell lines was associated with the acquisition of copy number gains involving chromosomal region 2p13-24 as common early alterations. In addition, we found an amplification of 8q21-24 after prolonged propagations in all three NBS-derived cell lines as well as early development of near-tetraploidy in two of these lines. Gains involving the short arm of chromosome 2 recently were found in several lymphoid malignancies. Therefore, the cell lines described here can be used for identification and characterization of genes involved in the pathogenesis of lymphoid neoplasms and would also provide a useful tool for better understanding the mechanisms responsible for cell immortalization. PMID- 15287028 TI - Divergent patterns of telomere maintenance mechanisms among human sarcomas: sharply contrasting prevalence of the alternative lengthening of telomeres mechanism in Ewing's sarcomas and osteosarcomas. AB - Two types of telomere maintenance mechanisms (TMMs) have been described in human tumors: telomerase activation and alternative lengthening of telomeres (ALT). Although the vast majority of epithelial tumors rely on telomerase activation, many mesenchymal tumors rely on ALT for telomere maintenance, but within this tumor group, the TMMs used by translocation-associated sarcomas have not been systematically studied. We studied telomere lengths and telomerase expression and activity in 30 uncultured tumor samples and in 10 cell lines of Ewing's sarcoma, a prototypical translocation-associated sarcoma, and compared the data to an identical analysis of 60 osteosarcomas, the most common type of sarcoma lacking a specific translocation. Telomerase activity was demonstrated in 21 Ewing's sarcoma tumor samples (70%) and in 9 of 10 Ewing's sarcoma cell lines. Evidence of ALT, indicated by the presence of long and heterogeneous telomeres, was observed only in the cell line without telomerase activity and in none of the 30 Ewing's sarcoma tumor samples. The 9 Ewing's sarcoma patients whose tumors lacked detectable telomerase activity did not differ significantly from the remaining patients in age, stage, EWSR1-FLI1 fusion type, prevalence of TP53 point mutations, or overall survival. The low prevalence of ALT in Ewing's sarcoma contrasted sharply with our data on TMMs in 60 osteosarcomas, which showed ALT in 38 of 60 cases (P<0.0001). The present study, together with emerging published data on other sarcoma types, suggests that a predominance of telomerase activation in the absence of ALT may characterize sarcomas with specific chromosomal translocations (such as Ewing's sarcoma), whereas a high prevalence of ALT appears typical of sarcomas with nonspecific complex karyotypes (such as osteosarcoma). PMID- 15287027 TI - Loss of chromosome arm 18q with tumor progression in head and neck squamous cancer. AB - Loss of 18q was analyzed in 21 sets of head and neck squamous cell carcinoma (HNSCC) cell lines derived from primary and secondary tumors in the same patients. Only 3 of the 21 cell line pairs had no loss of 18q. In the remaining 18 sets, loss of heterozygosity (LOH) affecting 18q was found in either the primary or the secondary lines or both. In every case but one, the same chromosome was affected in both the primary and secondary cell lines. In 8 sets, the 18q loss occurred in the primary tumor and remained stable through the subsequent tumor progression. The primary and secondary lines differed in 18q loss in 10 of 18 (56%) cases with 18q LOH. In 3 of the 10 pairs that differed, 18q LOH was found in only the primary line, indicating that the loss developed after the metastatic or recurrent tumor population had diverged from the primary tumor population. In the other 7 pairs, 18q LOH developed or progressed with tumor recurrence or metastasis. Of these, 3 of 7 had 18q LOH in only the secondary lines, and 4 of 7 had 18q LOH in both the primary and secondary lines, but the extent of LOH was greater in the secondary lines than in the primary lines, indicating that additional rearrangements of the same chromosome occurred with progression. These cases showed that interstitial loss often progresses to consolidated loss in vivo. However, in vitro, the cell lines from the primary tumors with interstitial loss maintain those chromosomes over long-term culture. LOH on 18q in cell lines from previously untreated primary tumors was significantly associated with advanced tumor stage (P=0.0242) and decreased survival (P=0.0453). The findings are consistent with the concept that 18q LOH is an event associated with tumor progression and suggest that inactivation and loss of one or more genes on 18q contributes to aggressive tumor behavior. PMID- 15287029 TI - Mapping of a translocation breakpoint in a Peutz-Jeghers hamartoma to the putative PJS locus at 19q13.4 and mutation analysis of candidate genes in polyp and STK11-negative PJS cases. AB - Germ-line mutations in the serine-threonine kinase gene STK11 (LKB1) cause Peutz Jeghers syndrome (PJS), a rare autosomal dominantly inherited disease, characterized by hamartomatous polyposis and mucocutaneous pigmentation. STK11 mutations only account for about half of PJS cases, and a second disease locus has been proposed at chromosome segment 19q13.4 on the basis of genetic linkage analysis in one family. We identified a t(11;19)(q13;q13.4) in a PJS polyp arising from the small bowel in a female infant age 6 days. Because the breakpoint in 19q13.4 may disrupt the putative PJS disease gene mapping to this region, we mapped the breakpoint and analyzed DNA from the case and a series of STK11-negative PJS cases. Using two-color interphase fluorescence in situ hybridization, the breakpoint region was refined to a 0.5-Mb region within 19q13.4. Eight candidate genes mapping to the breakpoint region--U2AF2, EPN1, NALP4, NALP11, NALP5, ZNF444, PTPRH, and KIAA1811--were screened for mutations in germ-line and polyp DNA from the case and from 15 PJS cases that did not harbor germ-line STK11 mutations. No pathogenic mutations in the candidate genes were identified. This report provides further evidence of the existence of a second PJS disease locus at 19q13.4 and excludes involvement of eight candidate genes. PMID- 15287030 TI - Genetic and epigenetic screening for gene alterations of the chromatin-remodeling factor, SMARCA4/BRG1, in lung tumors. AB - The SMARCA4/BRG1 gene product is a component of the SWI-SNF chromatin-remodeling complex and regulates gene expression by disrupting histone-DNA contacts in an ATP-dependent manner. Inactivating mutations of the SMARCA4 gene, on chromosome arm 19p, are present in several human cancer cell lines, including cell lines derived from lung cancers. Interestingly, loss of heterozygosity (LOH) at 19p and absence of the SMARCA4 protein have been reported in lung tumors. To evaluate further the possible contribution of SMARCA4 gene inactivation to lung carcinogenesis, we performed a complete analysis of the SMARCA4 gene to search for (a) point mutations in all 35 coding exons, including an existing splicing variant and the intron-exon boundaries, and (b) abrogation of gene expression through promoter hypermethylation by using the methylation-specific polymerase chain reaction (MSP) assay. We selected genomic DNA from 20 lung primary tumors with LOH on 19p for the screening of point mutations and 10 lung cancer cell lines and 52 lung primary tumors for the MSP analysis. Through our mutational screening, we identified an in-frame and germ-line insertion of 24 bp in exon 4 whose biological relevance is unknown. This variant was not detected in the germ line of the 62 additional individuals analyzed, indicating it is not a common polymorphism. Moreover, two missense alterations were identified in the tumors of 2 patients, a somatic Gly1160Arg mutation and a Ser1176Cys mutation. Neither was present in the germ line of the 51 additional lung cancer individuals tested. Because these mutations lead to substitution of highly conserved amino acids, they may affect the ATPase function of the protein. Finally, no promoter hypermethylation was observed in any lung primary tumor or cancer cell line, indicating that this is not a major mechanism for SMARCA4 inactivation during lung carcinogenesis. In conclusion, our data revealed that somatic point mutations of the SMARCA4 gene are present in a small subset of lung tumors, although mutations affecting the ATPase domain may be a hot-spot for SMARCA4 gene inactivation. We cannot rule out that other mechanisms, such as complete or partial deletions of the SMARCA4 gene, are contributing to the loss of the SMARCA4 protein in lung cancer. PMID- 15287031 TI - Level of MYC overexpression in pediatric Burkitt's lymphoma is strongly dependent on genomic breakpoint location within the MYC locus. AB - Increased transcriptional activity of the MYC gene is a characteristic feature of Burkitt's lymphoma. Aberrant MYC expression is caused by (1) chromosomal translocation to one of the loci carrying an immunoglobulin gene, (2) mutation within the translocated allele, (3) loss of the block to transcription elongation, or (4) promoter shift. To investigate the influence of breakpoint locations within the MYC gene on MYC transcript levels, we determined both the precise genomic MYC/IGH breakpoints and the amount of MYC mRNA in 25 samples of pediatric Burkitt's lymphoma with translocation t(8;14)(q24;q32). Patients with breakpoints that were 5' from MYC exon 1 had significantly lower expression of MYC than did patients who had a breakpoint within exon 1 or intron 1 (P < 0.05 and 0.005, respectively). The highest mRNA level of MYC (1,006 copies per 100 copies ABL1) was detected in patients with loss of the first exon and transcription initiation from a cryptic P3 promoter within the first intron of the MYC gene. In contrast, there was no obvious correlation between breakpoint locations within the IgH locus and the amount of MYC mRNA. PMID- 15287032 TI - Functional outcomes of transoral laser surgery of supraglottic carcinoma compared with a transcervical approach. AB - BACKGROUND: Several functional advantages have been described for the transoral laser supraglottic laryngectomy as compared with open techniques. However, comparative studies have been rarely performed. METHODS: Functional results in 26 patients treated with a transoral approach were retrospectively compared with those of a comparable series of 26 patients who underwent a transcervical approach. RESULTS: The only significant differences found between the two groups were a lesser number of temporary tracheotomies and a shorter time of removal of the nasogastric tube in the laser group. No significant differences were found in the incidence of postoperative complications, hospital stay, and swallowing capacity. CONCLUSIONS: The rate of functional problems after transoral laser surgery did not greatly decrease compared with the rate after the conventional procedure. The mayor advantage of the transoral approach was the lower incidence of temporary tracheotomies. PMID- 15287033 TI - The National Cancer Data Base report on squamous cell carcinoma of the base of tongue. AB - BACKGROUND: This study provides the largest contemporary overview of presentation, care, and outcome for base of tongue squamous cell carcinoma (SCC). METHODS: We extracted 16,188 cases from the National Cancer Data Base (NCDB). Chi square analyses were performed on selected cross-tabulations. Observed and disease-specific survival were used to analyze outcome. RESULTS: Three-quarters had advanced-stage (III-IV) disease. Radiation therapy alone (24.5%) and combined with surgery (26.9%) were the most common treatments. Five-year observed and disease-specific survival rates were 27.8% and 40.3%, respectively. Poorer survival was significantly associated with older age, low income, and advanced stage disease. For early-stage disease, surgery with or without irradiation had higher survival than irradiation alone. For advanced-stage disease, surgery with irradiation had the highest survival. CONCLUSIONS: Survival rates were low for base of tongue SCC, with most deaths occurring within the first 2 years. Income, stage, and age were significant prognostic factors. In this nonrandomized series, surgery with radiation therapy offered patients with advanced-stage disease the best survival. PMID- 15287034 TI - Pediatric submandibular triangle masses: a fifteen-year experience. AB - BACKGROUND: The purpose of this study was to evaluate the surgical results of pediatric submandibular triangle masses, with specific attention to neoplastic processes. METHODS: We retrospectively reviewed the medical records of 105 patients aged 6 months to 21 years who underwent surgery in the submandibular triangle at a major pediatric tertiary care hospital from 1987 to 2001. RESULTS: One hundred five patients who underwent surgery in the submandibular triangle were included in the study. Twenty patients had neoplastic processes, six of which were of primary salivary origin (two mucoepidermoid carcinomas and four pleomorphic adenomas). Twenty-four patients underwent excision of inflamed or infected lymph nodes, and 23 patients underwent excision of inflamed or infected submandibular glands. Thirty-eight patients were included who underwent surgery for sialorrhea or to gain access for another surgical procedure. Complications included tumor recurrence, transient and permanent marginal mandibular nerve weakness, ranula, postoperative fluid collection, and cellulitis. Duration of follow-up ranged from no follow-up to 11 years. CONCLUSION: Surgical excision of submandibular triangle masses is uncommon. We present our experience with these lesions, with a discussion of diagnosis, surgical indications, and surgical complications. PMID- 15287035 TI - Salivary gland carcinoma: independent prognostic factors for locoregional control, distant metastases, and overall survival: results of the Dutch head and neck oncology cooperative group. AB - BACKGROUND: We analyzed the records of patients with malignant salivary gland tumors, as diagnosed in centers of the Dutch Head and Neck Oncology Cooperative Group, in search of independent prognostic factors for locoregional control, distant metastases, and overall survival. METHODS: In 565 patients, we analyzed general results and looked for the potential prognostic variables of age, sex, delay, clinical and pathologic T and N stage, site (332 parotid, 76 submandibular, 129 oral cavity, 28 pharynx/larynx), pain, facial weakness, clinical and pathologic skin involvement, histologic type (WHO 1972 classification), treatment, resection margins, spill, perineural and vascular invasion, number of neck nodes, and extranodal disease. The median follow-up period was 74 months; it was 99 months for patients who were alive on the last follow-up. RESULTS: The rates of local control, regional control, distant metastasis-free and overall survival after 10 years were, respectively, 78%, 87%, 67%, and 50%. In multivariable analysis, local control was predicted by clinical T-stage, bone invasion, site, resection margin, and treatment. Regional control depended on N stage, facial nerve paralysis, and treatment. The relative risk with surgery alone, compared to surgery plus postoperative radiotherapy, was 9.7 for local recurrence and 2.3 for regional recurrence. Distant metastases were independently correlated with T and N stage, sex, perineural invasion, histologic type, and clinical skin involvement. Overall survival depended on age, sex, T and pN stage, site, skin and bone invasion. CONCLUSIONS: Several prognostic factors for locoregional control, distant metastases, and overall survival were found. Postoperative radiotherapy was found to improve locoregional control. PMID- 15287036 TI - High-resolution image cytometry on smears of normal oral mucosa: a possible approach for the early detection of laryngopharyngeal cancers. AB - BACKGROUND: The purpose of this study was to investigate the possibility of identifying laryngopharyngeal cancers by nuclear chromatin texture feature analysis of cell nuclei from mucosal scrapings obtained from clinically and cytologically noncancerous areas of the soft palate in patients with cancer. METHODS: The collective consisted of 68 controls and 77 cases of laryngopharyngeal carcinomas. After Feulgen staining, 3000 cell nuclei were automatically measured using a high-resolution image analyser (CytoSavant Oncometrics, Vancouver, BC, Canada). Texture features were extracted for calculation of a discriminant function, which allows the two groups to be distinguished. RESULTS: Two parameters allowed the two populations to be distinguished. The classifier reached an overall performance of 72.7% sensitivity, 82.4% specificity, a positive predictive value of 80.5%, a negative predictive value of 75.1%, and an area under the receiver operating characteristics (ROC) curve of 0.7754. CONCLUSION: Our work shows that subtle changes in the chromatin distribution in cell nuclei from ostensibly normal cells in the vicinity of carcinomas are demonstrable in the oral cavity of patients suffering from laryngopharyngeal cancers. It may be possible to develop this method into a valuable clinical tool to reduce the high rate of delayed diagnosis of oral and laryngopharyngeal cancers. PMID- 15287037 TI - Supraglottic hemipharyngolaryngectomy for the treatment of T1 and T2 carcinomas of laryngeal margin and piriform sinus. AB - BACKGROUND: We evaluated the functional and oncologic results of supraglottic hemipharyngolaryngectomy as treatment for T1 and T2 lateral laryngeal margin and piriform sinus carcinomas. METHODS: Eighty-seven patients underwent this surgical treatment. The disease was classified T1 in 14 of these cases (16.1%) and T2 in 73 cases (83.9%). The nodal status indicated 39 cases of N0 (44.8%), 18 cases of N1 (20.7%), 28 cases of N2 (32.2%), and two cases of N3 (2.2%). With regard to the N0 cases, 15 (38.4%) were positive at the histologic examination. Within the N+ group, 52.1% involved capsular rupture. RESULTS: Two patients died of complications during the postoperative period. The mean duration of nasogastric tube feeding was 20 days. Six patients (7.27%) had feeding resumption problems. All patients were decanulated after a mean period of 16 days. All patients underwent postoperative radiation therapy, except two with T1N0N- disease and three who had previously undergone this treatment. The 5-year actuarial survival rate was 60.3% (T1, 83.3%; T2, 49.9%). The rates of local and regional recurrence, second primary cancer, and metastasis were 19.5%, 24.1%, and 28.1%, respectively. The infringement of the pharyngoepiglottic fold was significantly correlated with locoregional recurrence. The survival rate was significantly correlated with the nodal status and extracapsular spread. CONCLUSIONS: Initial staged cancers of the laryngeal margin and piriform sinus can be successfully managed with conservative surgery called supraglottic hemipharyngolaryngectomy combined with nodal neck dissection. Postoperative radiation therapy is still recommended in most cases because of the high recurrence potential and prevalence of secondary regional cancers. This combined treatment seems to be a suitable therapeutic choice in the treatment of patients with T1 and T2 carcinomas of the laryngeal margins and piriform sinus. PMID- 15287038 TI - Effects of p53 or p27 overexpression on cyclooxygenase-2 gene expression in head and neck squamous cell carcinoma cell lines. AB - BACKGROUND: Although cyclooxygenase-2 (COX-2) has been suggested to play an important role in carcinogenesis, the effects of tumor suppressors on COX-2 gene expression and the combined antitumor effects of tumor suppressors and COX-2 inhibitors have rarely been investigated. METHODS: The effects of p53 or p27 gene transfer on COX-2 expression by adenoviral vector and the combined effects of p53 or p27 gene transfer and COX-2 inhibitor exposure on the proliferation of cancer cells were investigated in head and neck squamous cell carcinoma (HNSCC) cell lines. RESULTS: Overexpression of p53 markedly downregulated the transcription of COX-2, but the overexpression of p27 did not affect COX-2 levels in HNSCC cell lines. The combined antitumor effects of p53 or p27 gene transfer and of a COX-2 inhibitor (NS 398) were mainly at least additive in terms of the inhibition of cell proliferation and cell cycle arrest and additive in terms of apoptotic induction. CONCLUSIONS: These results suggest that the overexpression of p53 could exert antitumor effects, at least in part, through the suppression of COX-2 gene expression, whereas growth suppression by the overexpression of p27 probably occurs by mechanisms other than the downregulation of COX-2 expression. In addition, the administration of COX-2 inhibitors, as an adjunct to p53 or p27 gene therapy, could offer a new strategy of cancer treatment and prevention. PMID- 15287039 TI - Parathyroid carcinoma: a 22-year experience. AB - PURPOSE: Because parathyroid carcinoma is rare, clear consensus is not available regarding the optimal management of patients with this condition. Treatment strategies generally derive from clinical and anecdotal experiences. We report our experience with this entity. METHODS: We included all patients with parathyroid carcinoma seen at The University of Texas M. D. Anderson Cancer Center since January 1, 1980. The medical records and pathology specimens were reviewed and verified in all cases. RESULTS: Since 1980, 27 patients (16 men and 11 women) registered at M. D. Anderson Cancer Center with parathyroid carcinoma and a minimum follow-up of 2 years. The age at initial diagnosis (mean +/- SD) was 46.7 +/- 15.3 years. All patients were seen with hypercalcemia (mean calcium, 13.4 +/- 1.5 mg/dL). Eighteen patients had locally invasive disease, eight had localized disease, and one had distant metastasis. Parathyroid cancer was treated with complete surgical excision with curative intent in 18 patients. In the other nine patients, who had clinical and/or radiographic evidence of soft tissue extension, the tumor was treated by comprehensive "en bloc" soft tissue resection. Of six patients who received adjuvant radiotherapy after initial surgery, only one had a local relapse. In contrast, of 20 patients who did not receive adjuvant radiotherapy, 10 had a local relapse, excluding the one patient who had distant metastases. The 5-year survival was 85%, and the 10-year survival was 77%. Five patients died of parathyroid carcinoma; all deaths were hypercalcemia related. CONCLUSIONS: Parathyroid carcinoma can be an indolent disease with morbidity and mortality related to hypercalcemia. Adjuvant radiotherapy may improve local control and limit the occurrence of local relapse. A comprehensive multidisciplinary approach with surgery, radiation therapy, and medical treatment for hypercalcemia is needed to optimize patient outcome. PMID- 15287040 TI - Cutaneous metastatic squamous cell carcinoma to the parotid gland: analysis and outcome. AB - BACKGROUND: Our aim was to review the presentation, treatment, and outcome of patients with metastatic cutaneous squamous cell carcinoma involving the parotid gland at a tertiary referral center. METHODS: We performed a retrospective chart review of the cancer registry at the Princess Margaret Hospital, Toronto, from 1970 to 2001. All patients had a previously untreated metastatic cutaneous head and neck squamous cell carcinoma involving the parotid gland. A minimal follow-up of 1 year was mandatory for inclusion in the study. RESULTS: Fifty-six white patients (43 men and 13 women), with a median age of 76 years (range, 49-97 years), were eligible for inclusion. The disease in all patients was retrospectively staged according to a new system. Twenty patients had P1 disease, 14 had P2, and 22 had P3. Therapy included surgery and adjuvant external beam radiation in 37 patients, single-modality external beam radiation in 12, and surgery alone in seven patients. The overall recurrence rate was 29%. The disease specific survival was significantly worse in patients treated with external beam radiation alone (p <.05). Tumor size >6 cm (p <.01) and the presence of facial nerve involvement (p <.01) were poor prognostic factors. CONCLUSIONS: Metastatic cutaneous squamous cell carcinoma to the parotid gland is an aggressive neoplasm that requires combination therapy. The presence of a lesion in excess of 6 cm or with facial nerve involvement is associated with a poor prognosis. PMID- 15287041 TI - Combined evaluation of expression of telomerase, survivin, and anti-apoptotic Bcl 2 family members in relation to loss of differentiation and apoptosis in human head and neck cancers. AB - BACKGROUND: Head and neck squamous cell carcinoma (HNSCC) is one of the most common cancers, and it accounts for 5% of all adult cancers worldwide. Loss of growth control and a marked resistance to apoptosis are considered major mechanisms driving tumor progression. Little is known about the distribution of inhibitors of apoptosis in HNSCC or how they correlate with other biomarkers of malignancy, such as telomerase, an enzyme that plays a critical role in cellular immortalization. The objective of this study was to assess the protein expression of anti-apoptotic members of Bcl-2 family and survivin and correlate them with telomerase activity. METHODS: We compared anti-apoptotic protein expression in tumor tissue sections of 50 HNSCC patients and 19 histopathologically normal tissues by immunohistochemistry and Western blotting. Apoptotic index was studied by TUNEL assay. Telomerase activity was determined by polymerase chain reaction (PCR)-enzyme-linked immunosorbent assay (ELISA). RESULTS: Bcl-2, Bcl-XL, and survivin were found to be significantly upregulated in tumor tissues as compared with the normal tissue. Protein expression of Bcl-2 and survivin was significantly associated with the loss of differentiation in tumors and that of Bcl-XL with nodal metastasis. Telomerase activity was found to be upregulated in tumors as compared with the normal tissue (p <.001) and was found to be significantly associated with the loss of differentiation in tumors. CONCLUSIONS: Mechanisms that promote both cell proliferation (telomerase activity) and cell survival (expression of inhibitors of apoptosis protein [IAPs]) appear to be activated in HNSCC. This is the first study of its kind to look into the correlation of the apoptotic pathway and proliferation promoting enzyme activity simultaneously in relation to loss of apoptosis and differentiation in HNSCCs. Telomerase activity in these tumors was found to be correlated with Bcl-2, Bcl XL, and survivin overexpression and with reduced apoptosis, thereby suggesting that novel strategies can be developed to control cancer cell growth by co targeting telomerase and apoptotic pathways. PMID- 15287042 TI - Cavernous carotid aneurysm presenting with epistaxis. AB - BACKGROUND: Carotid artery aneurysms are a rare cause of epistaxis. The most common presentation for nontraumatic cavernous internal carotid artery aneurysms is mass effect, with only 3% presenting with hemorrhage. We present a case of epistaxis caused by a nontraumatic cavernous internal carotid artery aneurysm. METHODS: A 73-year-old white woman was seen with a 1-month history of recurrent right-sided epistaxis. The patient had essential hypertension and a family history of intracranial aneurysm. A complete otolaryngologic, neurologic, and ophthalmologic examinations were normal. Contrast-enhanced CT of the paranasal sinuses revealed a trilobed aneurysm of the cavernous segment of the right internal carotid artery. Coil embolization of the cavernous aneurysm and right internal artery was performed. RESULTS: The patient has had no further episodes of epistaxis and has remained neurologically intact. CONCLUSION: Carotid artery aneurysms must be considered in the differential diagnosis of profuse epistaxis. PMID- 15287043 TI - The successful use of maggots in necrotizing fasciitis of the neck: a case report. AB - BACKGROUND: The use of maggots to digest necrotic tissue as a form of wound debridement has a long history in medicine. Necrotizing fasciitis of the neck has a high mortality rate despite aggressive surgical and medical intervention. The use of maggots in this disease has been reported only once before. METHODS: We report the case of a 73-year-old woman, who underwent neck dissection and had necrotizing fasciitis of the neck develop shortly after. After initial surgical wound debridement, we used maggots as a biosurgical method for further debridement. A net containing 100 maggots (Biobag; BioMonde, Germany) was used. RESULTS: Daily wound dressing showed rapid improvement of the wound; 4 days after beginning treatment, the wound was free of necroses. CONCLUSION: In this case, we could avoid repeated surgical wound debridement with the use of sterile maggots. The frequently rapid progression of necrotizing fasciitis could be well controlled. PMID- 15287045 TI - Fetal lung lesions: a spectrum of disease. New classification based on pathogenesis, two-dimensional and color Doppler ultrasound. PMID- 15287046 TI - Newer imaging modalities in the prenatal diagnosis of skeletal dysplasias. PMID- 15287047 TI - Fetoscopic tracheal occlusion (FETO) for severe congenital diaphragmatic hernia: evolution of a technique and preliminary results. AB - BACKGROUND: Congenital diaphragmatic hernia (CDH) with liver herniation and a lung area to head circumference ratio (LHR) < 1 [corrected] is associated with a high rate of neonatal death due to pulmonary hypoplasia. METHODS: We report the development of a minimally invasive and reversible fetoscopic tracheal occlusion (FETO) with a balloon, carried out in 21 consecutive fetuses with severe CDH. RESULTS: Endotracheal placement of the balloon was successfully performed in all 21 cases and the mean duration of the operation was 20 (range, 5-54) min. The median gestation at FETO was 26 (range, 25-33) weeks. There were no maternal complications such as hemorrhage, placental abruption or pulmonary edema. In 11 (52.4%) patients there was postoperative prelabor amniorrhexis, which occurred within 2 weeks in five patients and after 2 weeks in six patients. Ultrasound scans after FETO demonstrated an increase in the echogenicity of the lungs within 48 h and improvement in the LHR from a median 0.7 (range, 0.4-0.9) before FETO to 1.8 (range, 1.1-2.9) within 2 weeks following surgery. The median gestation at delivery was 34 (range, 27-38) weeks and in 17 (77.3%) patients delivery occurred after 32 weeks. Nine babies died in the neonatal period due to complications from pulmonary hypoplasia. Surgical repair of the diaphragmatic hernia was carried out in 12 babies and in all but one the defect was extensive and required the insertion of a patch. Ten of these babies survived, and at the time of writing were aged 6-25 (median, 18) months and were developing normally. Survival was 30% in the first group of 10 fetuses and 63.6% in the second group of 11 fetuses. The total number of cases was too small for definite conclusions to be drawn as to the causes of this apparent improvement in survival. Nevertheless, improved survival coincided with a shift in the timing of FETO from the third to the second trimester, the administration of epidural rather than general anesthesia, reduced incidence of postoperative amniorrhexis and a change in the policy on the timing of removal of the balloon from the intrapartum to the prenatal period. During the same period of study there were 17 cases examined in the participating centers that met the criteria for FETO but which declined prenatal therapy. In all cases there was isolated left-sided CDH with liver in the thorax and LHR of 0.4-0.9 (mean, 0.7). In five cases the parents elected to terminate the pregnancy. In the 12 cases with expectant management all babies were liveborn but 11 died in the neonatal period due to pulmonary hypoplasia and only one (8.3%) survived. CONCLUSION: Severe CDH can be successfully treated with FETO, which is minimally invasive and may improve postnatal survival. PMID- 15287049 TI - Prenatal diagnosis of fetal skeletal dysplasias by combining two-dimensional and three-dimensional ultrasound and intrauterine three-dimensional helical computer tomography. AB - OBJECTIVE: To evaluate the contribution of new imaging techniques in the prenatal diagnosis of skeletal dysplasia. METHODS: Between May and October 2003, a prospective study was conducted in a single referral center. Three-dimensional ultrasound (3D-US) and three-dimensional helical computer tomography (3D-HCT) were performed after two-dimensional ultrasound (2D-US) in six cases of skeletal dysplasia. Diagnostic accuracy and detailed findings with each of the three techniques were compared with postnatal radiological findings. RESULTS: There were three cases of achondroplasia, two cases of osteogenesis imperfecta type II and one case of chondrodysplasia punctata. Termination of pregnancy was performed in five cases and one fetus with osteogenesis imperfecta type II was delivered at term by Cesarean section. 2D-US made the correct diagnosis in four cases. 3D-US and 3D-HCT achieved an accurate diagnosis in all six cases. 3D-HCT and 3D-US identified significantly more abnormalities than did 2D-US (3D-HCT: 94.3% (33/35); 3D-US: 77.1% (27/35); 2D-US: 51.4% (18/35); P < 0.01). The diagnosis was made between 27 and 36 weeks' gestation in all cases. The advantage of 3D-HCT over 3D-US was the possibility of imaging the entire fetus. CONCLUSION: 3D-US and 3D-HCT seem to be useful complementary methods to 2D-US, and may improve accuracy of the prenatal diagnosis of skeletal disorders. These new imaging technologies may have a role in the prenatal multidisciplinary approach to the diagnosis of skeletal dysplasias. PMID- 15287048 TI - Fetal lung dysplasia: clinical outcome based on a new classification system. AB - OBJECTIVE: To evaluate the clinical application of a new classification system of fetal lung anomalies. METHODS: Forty fetal diagnoses of lung lesions were analyzed according to our proposed classification system in which each lung component is considered using two-dimensional ultrasound and color and power Doppler technology. Medical files, natural history and neonatal follow-up were recorded. RESULTS: Type I dysplasia: Four cases of agenesis of the lung were diagnosed, three with right lung agenesis and one with left lung agenesis. Three of the four patients elected to undergo termination of pregnancy (TOP). The surviving fetus was diagnosed with scimitar syndrome and postnatal embolization of the aberrant vessel was performed. Type II dysplasia: One case of normal lung with abnormal systemic feeding artery was diagnosed with normal neonatal outcome. Type III dysplasia: Abnormal lung with abnormal vascularity was found in 14 cases, presenting in most cases as echogenic lung masses. Seven were supradiaphragmatic, six subdiaphragmatic and one case was of undetermined position. All 14 fetuses showed an aberrant systemic artery emerging from the aorta. Abnormal venous drainage could be identified in only five (36%) of the fetuses: three had prominent azygos vein, one showed drainage to the inferior vena cava and one had multiple intrapulmonary veins forming a huge arteriovenous (A-V) shunt. Two cases in this group underwent TOP, the case with A-V shunt following development of hydrops, and one on maternal request. The remaining 12 fetuses (86%) survived and were alive and well at the time of writing; only one of them needed immediate postnatal embolization of the bilateral aberrant feeding arteries. Type IV dysplasia: Abnormal lung with no vascular abnormality was diagnosed in 20 fetuses. In this group there was one case of intrauterine fetal death, two patients underwent TOP, one complicated with hydrops and one on maternal request. The survival rate in this group was 85%. Only two cases needed immediate surgical repair. Type V miscellaneous dysplasia: One fetus demonstrated echogenic lung with split notochord syndrome and survived. CONCLUSIONS: Congenital bronchopulmonary and related vascular anomalies can be categorized using the new classification system. This new approach enabled prenatal evaluation of each lung component and facilitated cogent management of the fetus with congenital lung dysplasia. PMID- 15287050 TI - Foramen ovale changes in growth-restricted fetuses. AB - OBJECTIVE: In animal experiments hypoxemia induces an increased shunting through the fetal foramen ovale (FO). Based on the hypothesis that the FO is expanded to permit more flow, the aim of this study was to determine the size of the FO in growth-restricted human fetuses. METHODS: Thirty-one women with singleton pregnancies complicated with growth-restriction (< 5th percentile) were examined at 24-39 weeks of gestation. The diameter between the FO valve and the atrial septum was determined during maximum excursion in a horizontal transverse section of the fetal heart, and the transverse diameter of the right atrium (RA) was noted. The pulsatility index (PI) was determined in the umbilical artery (UA) and absent or reversed end-diastolic flow velocity was noted. The measurements were compared with a reference population using Z-scores. RESULTS: In comparison with normally grown fetuses, the growth-restricted fetuses had a normal RA size (P = 0.08) but a smaller FO (P = 0.002), particularly when expressed as a relative size by the ratio FO/RA (P < 0.0001). This effect on the FO and FO/RA was seen mainly at < 32 weeks of gestation (P = 0.003 and P < 0.0001, respectively), and was not significant later in pregnancy. There was a tendency towards a negative relationship between relative size of the FO (FO/RA) and progressive placental compromise (overall P < 0.0001). CONCLUSIONS: Growth-restricted fetuses maintain a normally grown heart (expressed by the RA diameter) but a reduced FO diameter. The effect is seen before 32 weeks and tends to be more marked in fetuses with pronounced hemodynamic compromise of the placenta. This supports the theory that FO shunting is impaired in severely premature fetuses with placental compromise. PMID- 15287051 TI - Fetal cerebral venous Doppler velocimetry in normal and high-risk pregnancy. AB - OBJECTIVE: In previous pilot studies, fetal vein of Galen (GV) blood velocity has been shown to be non-pulsatile in normal pregnancies. A pulsating pattern in high risk pregnancies has been related to adverse outcome of pregnancy. The aim of this study was to establish reference ranges for fetal cerebral venous blood flow and compare them to the recordings in high-risk pregnancies in terms of predicting adverse perinatal outcome. METHODS: The GV, straight sinus (SS) and transverse sinus (TS) were located by color Doppler ultrasound in 189 normal pregnancies between 23 and 43 weeks of gestation. Recordings were also made in 102 pregnancies complicated by pregnancy-induced hypertension and/or intrauterine growth restriction. The following parameters were measured: peak systolic velocity, minimum diastolic velocity, time-averaged maximum velocity, pulsatility index for veins (PIV) and preload index (PLI). GV pulsations were noted. In high risk pregnancies, Doppler measurements were correlated to pregnancy outcome, including emergency operative intervention and/or neonatal distress. Umbilical vein and umbilical, uterine and middle cerebral artery blood velocities were also recorded at the same time. RESULTS: In normal pregnancy, pulsating venous blood velocity was observed in GV in 8% of cases, in SS in 79% of cases and in TS in 100% of cases. GV and SS maximum velocity increased with gestational age and TS PIV showed linear decreasing values and TS-PLI showed increasing values with gestational age. In high-risk pregnancies, pulsating blood velocity in the GV was found in 59 (58%) cases and was related to adverse outcome of pregnancy including mortality. Abnormal values for TS-PIV and PLI and SS maximum velocity were found in nine, six and five cases, respectively and were only related to perinatal mortality. GV pulsations were more frequent than umbilical venous pulsations. CONCLUSIONS: Of the fetal cerebral veins studied, the presence of pulsations in the GV seems to be the best predictor of adverse outcome of high-risk pregnancy. Pulsations in the GV are more frequent than in the umbilical vein and might therefore appear earlier during worsening fetal condition, and thus be of potential value for fetal surveillance in high-risk pregnancies. PMID- 15287052 TI - Neurological outcome of children who were treated for fetal tachycardia complicated by hydrops. AB - OBJECTIVE: Fetal tachycardia is a condition associated with congestive heart failure and development of fetal hydrops, which may result in neurological morbidity and mortality. The aim of this study was to investigate the long-term outcome of hydropic fetuses. METHODS: This was a retrospective study on cognitive and neurological functioning of 11 infants, aged 6 months to 12 years, who experienced fetal tachycardia complicated by hydrops. RESULTS: Seven fetuses had supraventricular tachycardia (SVT), three had atrial flutter (AF) and one had ventricular tachycardia (VT). Nine fetuses converted to sinus rhythm within a mean time of 8.2 days of presentation; resolution of hydrops was achieved in six of these patients in a mean time of 8.8 days. Mean gestational age (GA) at birth was 35 + 4 weeks. Neonatal cranial ultrasound was normal in seven infants and all but one of these were normal at follow-up: one infant who initially had no abnormalities developed multiple cerebral lesions as a result of a malignant long QT syndrome (LQTS) and died at the age of 2 years. Three infants had periventricular echogenicity (PVE) on neonatal cranial ultrasound, associated with a pseudocyst in one infant. The remaining infant showed a parenchymal hemorrhage of antenatal onset, seen as a porencephalic cyst at birth. One of these infants was normal at follow-up, one died 2 days after birth and two infants had neurological abnormalities at follow-up, consisting of mild hemiplegia with normal cognitive function in one, and a cognitive developmental delay in the other. CONCLUSIONS: In this study, neurological outcome was good in eight out of 11 infants. Initiation of therapy should not be withheld or delayed on the assumption of poor neurological outcome. PMID- 15287053 TI - Prevalence of neurological damage in monochorionic twins with selective intrauterine growth restriction and intermittent absent or reversed end-diastolic umbilical artery flow. AB - OBJECTIVE: To assess the incidence of parenchymal lesions on early and late neonatal brain scans and its association with the presence or absence of intermittent absent or reversed end-diastolic umbilical artery flow velocity (A/REDV) in monochorionic twins complicated by selective intrauterine growth restriction (IUGR), as compared to dichorionic twins and monochorionic twins without selective IUGR. METHODS: This was a prospective cohort study involving 42 monochorionic twins diagnosed with selective IUGR and managed expectantly. The presence or absence of intermittent A/REDV was recorded in all cases. This study group was compared to dichorionic twins (n = 29) and monochorionic twins without selective IUGR (n = 32) delivered at 26-34 weeks during the study period. All infants underwent an early neonatal brain scan (at or before the fourth day of postnatal life) and at least one follow-up scan during the first 28 days of postnatal life. Perinatal outcome and the incidence of neurological damage were compared between the study groups. RESULTS: The incidence of intrauterine fetal death (IUD) and periventricular leukomalacia was significantly increased in monochorionic twins complicated with selective IUGR, as compared with the other study groups. Intermittent A/REDV was observed in 22/42 (52.4%) twin pairs, and was always present in the growth-restricted twin. The incidence of IUD (overall 9/44 (20.5%) vs. 0/40, P < 0.001; smaller twin 6/22 (27.3%) vs. 0/20, P < 0.05) and parenchymal brain damage (overall 7/35 (20.0%) vs. 2/40 (5.0%), P = 0.07; larger twin 7/19 (36.8%) vs. 1/20 (5.0%), P < 0.05) was significantly higher in pregnancies with intermittent A/REDV than in those without intermittent A/REDV. Brain damage usually occurred in the larger twin, irrespective of whether the smaller twin was liveborn or not. CONCLUSIONS: The presence of intermittent A/REDV in monochorionic twins with selective IUGR identifies a subgroup with an elevated risk of intrauterine demise of the smaller twin and neurological damage in the larger twin; this latter finding is not restricted to cases with IUD of the cotwin. PMID- 15287054 TI - Ultrasound assessment of venous blood flow before and after laser therapy: approach to understanding the pathophysiology of twin-twin transfusion syndrome. AB - OBJECTIVE: To estimate direction and volume of blood exchange between the donor twin and recipient twin by ultrasound assessment of blood flow in the umbilical vein before and after selective laser photocoagulation of communicating vessels (SLPCV) for twin-twin transfusion syndrome (TTTS). METHODS: Forty-one TTTS patients underwent Doppler examination of the umbilical vein before and 24 h after SLPCV. The diameter and mean time-averaged velocity of the umbilical vein were estimated. Total umbilical venous flow (TUVF) was calculated as follows: TUVF (mL/min) = mean time-averaged velocity (cm/s) x mean cross-sectional area (cm2) x 60 (s). RESULTS: TUVF was significantly higher in the recipient (111.2 mL/min) than in the donor twin (44.8 mL/min) before SLPCV (P < 0.0001). However, TUVF was no different between the recipient and the donor twin after SLPCV (93.1 vs. 70.7 mL/min, recipient and donor twin, respectively, P = 0.11). The donor twin's TUVF increased after surgery (P < 0.0001), while the recipient twin's TUVF decreased (P = 0.041). The median postoperative increase in the donor twin's TUVF of 25.9 mL/min had a corresponding decrease of TUVF in the recipient twin of 18.1 mL/min (P = 0.27). CONCLUSIONS: Our data suggest that untreated TTTS is characterized by excessive umbilical venous blood flow in the recipient twin relative to the donor twin. Laser surgery results in concordant changes in umbilical venous flow in opposite directions between the donor and recipient twins, eliminating the initial imbalance. Our results lend support to the fundamental hypothesis of unbalanced blood flow exchange (net flow from donor to recipient) between monochorionic twins as the cause for TTTS and that laser surgery eliminates the pathophysiological cause. PMID- 15287055 TI - Disappearance of enlarged nuchal translucency before 14 weeks' gestation: relationship with chromosomal abnormalities and pregnancy outcome. AB - OBJECTIVE: The aim of this study was to investigate the natural course of enlarged nuchal translucency (NT) and to determine if its disappearance before 14 weeks' gestation is a favorable prognostic sign in relation to fetal karyotype and pregnancy outcome. METHODS: A total of 147 women with increased NT (> 95th centile) at first measurement were included in this study. A second measurement was performed in all cases, at an interval of at least 2 days. Both measurements were taken between 10 + 3 and 14 + 0 weeks. All women underwent chorionic villus sampling or amniocentesis for subsequent karyotyping. In those women with a normal karyotype, a fetal anomaly scan was performed at 20 weeks' gestation. Pregnancy outcome was recorded in all cases. The finding of persistent or disappearing NT enlargement was analyzed in relation to fetal karyotype and pregnancy outcome. RESULTS: Of the 147 paired measurements, NT remained enlarged at the second measurement in 121 (82%) cases. An abnormal karyotype was found in 35% of these cases. In 26 (18%) fetuses the NT measurement was found to be below the 95th percentile at the second measurement and in only two of them an abnormal karyotype was found (8%). In the 103 chromosomally normal fetuses an adverse outcome (i.e. fetal loss or structural defects) was recorded in 22 fetuses with persistent enlargement (28%) and in four fetuses with disappearing enlargement (17%). CONCLUSIONS: Disappearance of an enlarged NT before 14 weeks' gestation is not a rare phenomenon and seems to be a favorable prognostic sign with respect to fetal karyotype. Overall, no significant difference in pregnancy outcome was found between chromosomally normal fetuses with persisting or disappearing NT enlargement. PMID- 15287056 TI - Accuracy of rectal endoscopic ultrasonography and magnetic resonance imaging in the diagnosis of rectal involvement for patients presenting with deeply infiltrating endometriosis. AB - OBJECTIVE: To compare the accuracy of rectal endoscopic ultrasonography (REU) and magnetic resonance imaging (MRI) for predicting rectal wall involvement in patients presenting histologically proven deeply infiltrating endometriosis (DIE). METHODS: This was a retrospective study of a continuous series of 81 patients presenting histologically proven DIE who underwent preoperative investigations using both REU and MRI. The sonographer and the radiologist, who were unaware of the clinical findings and patient history, but knew that DIE was suspected, were asked whether there was involvement of the digestive wall. RESULTS: Rectal DIE was confirmed histologically in 34 of the 81 (42%) patients. For the diagnosis of rectal involvement, sensitivity, specificity and positive and negative predictive value for REU were 97.1%, 89.4%, 86.8% and 97.7% and for MRI they were 76.5%, 97.9%, 96.3% and 85.2%. CONCLUSION: The sensitivity and negative predictive value of REU were higher than those of MRI suggesting that REU performs better than MRI in the diagnosis of rectal involvement for patients presenting with DIE. Prospective studies with a large number of patients are needed in order to validate these preliminary results. PMID- 15287057 TI - Diagnostic accuracy of transvaginal sonography for deep pelvic endometriosis. AB - OBJECTIVE: To determine the accuracy of transvaginal sonography (TVS) for the diagnosis of deep pelvic endometriosis. METHODS: In a prospective study, 142 women with clinical signs of endometriosis underwent TVS followed by surgical and histopathological investigations. The presence and extent of endometriosis involving the uterosacral ligaments, vagina, rectovaginal septum, intestines, bladder and ovaries shown by TVS were compared with surgical and histological findings. The sensitivity, specificity, predictive values and accuracy of TVS for predicting deep pelvic endometriosis were assessed. RESULTS: Ovarian and deep pelvic endometriosis were found by surgery and histology in respectively 83 (58.5%) and 79 (55.6%) of the 142 patients. The sensitivity, specificity, and positive and negative predictive values of TVS for the diagnosis of deep pelvic endometriosis were 78.5%, 95.2%, 95.4% and 77.9%, respectively. The sensitivity and specificity of TVS for endometriotic involvement of the uterosacral ligaments, vagina, rectovaginal septum and intestines were 70.6% and 95.9%, 29.4% and 100%, 28.6% and 99.3%, and 87.2% and 96.8%, respectively. The sensitivity and specificity of TVS for bladder involvement were 71.4% and 100%, respectively. CONCLUSION: TVS accurately diagnoses intestinal and bladder endometriosis, but is less accurate for uterosacral, vaginal and rectovaginal septum involvement. PMID- 15287058 TI - Determination of bladder neck position by intraoperative introital ultrasound in colposuspension: outcome at 6-month follow-up. AB - OBJECTIVE: To determine whether open colposuspension modified by intraoperative ultrasound to prevent overcorrection is a safe and effective procedure. METHODS: Ninety women operated on for urodynamically proven genuine stress urinary incontinence underwent intraoperative introital ultrasound in a prospective observational clinical study. The positions of the bladder neck and proximal urethra were assessed by determining the parameters height (H), distance (D) and the urethrovesical angle (beta) perioperatively and for up to 6 months postoperatively. Colposuspension of the bladder neck was performed with a vertical height correction, DeltaH (resting H(intraop) - resting H(preop)) of 1 to 10 mm. Bladder neck positions were determined on an individual basis by introital ultrasound before, during and after surgery. RESULTS: Surgical elevation of the bladder neck (median height correction, DeltaH 4 mm) resulted in a median intraoperative elevation of 9 mm (6 months: 8 mm). All postoperative measurements showed a significant reduction of the median linear movement of the bladder neck during straining (P < 0.0001). Anti-incontinence surgery resulted in a significant reduction of funneling and hypermobility 6 months after surgery (P < 0.0001). At 6-month follow-up, 94% (85/90) of the women were continent. Evaluation immediately after surgery showed voiding difficulties and urge symptoms in 9% (8/90) of the patients each and de novo urge incontinence in 1% (1/90). CONCLUSIONS: Intraoperative introital ultrasound can help to optimize the colposuspension procedure. Ultrasonographic measurement of height H allows for objectively assessing the surgical procedure and can reduce postoperative complications by preventing excessive correction. PMID- 15287059 TI - Tissue Doppler gated (TDOG) dynamic three-dimensional ultrasound imaging of the fetal heart. AB - Dynamic three-dimensional (3D) ultrasound imaging of the fetal heart is difficult due to the absence of an electrocardiogram (ECG) signal for synchronization between loops. In this study we introduce tissue Doppler gating (TDOG), a technique in which tissue Doppler data are used to calculate a gating signal. We have applied this cardiac gating method to dynamic 3D reconstructions of the heart of eight fetuses aged 20-24 weeks. The gating signal was derived from the amplitude and frequency contents of the tissue Doppler signal. We used this signal as a replacement for ECG in a 3D-volume reconstruction and visualization, utilizing techniques established in ECG-gated 3D echocardiography. The reliability of the TDOG signal for fetal cardiac cycle detection was experimentally investigated. Simultaneous recordings of tissue Doppler of the heart and continuous wave (CW) spectral Doppler of the umbilical artery (UA) were performed using two independent ultrasound systems, and the TDOG signal from one system was compared to the Doppler spectrum data from the other system. Each recording consisted of a two-dimensional (2D) sector scan, transabdominally and slowly tilted by the operator, covering the fetal heart over approximately 40 cardiac cycles. The total angle of the sweep was estimated by recording a separate loop through the center of the heart, in the elevation direction of the sweep.3D reconstruction and visualization were performed with the EchoPAC-3D software (GE Medical Systems). The 3D data were visualized by showing simultaneous cineloops of three 2D slices, as well as by volume projections running in cineloop. Synchronization of B-mode cineloops with the TDOG signal proved to be sufficiently accurate for reconstruction of high-quality dynamic 3D data. We show one example of a B-mode recording with a frame rate of 96 frames/s over 20 seconds. The reconstruction consists of 31 volumes, each with 49 tilted frames. With the fetal heart positioned 5-8 cm from the transducer, the sampling distances were approximately 0.15 mm in the beam direction, 0.33 degrees approximately 0.37 mm azimuth and 0.45 degrees approximately 0.51 mm elevation. From this single dataset we were able to generate a complete set of classical 2D views (such as four-chamber, three-vessel and short-axis views as well as those of the ascending aorta, aortic and ductal arches and inferior and superior venae cavae) with high image quality adequate for clinical use. PMID- 15287060 TI - Prenatal diagnosis of hemifacial microsomia and ipsilateral cerebellar hypoplasia in a fetus with oculoauriculovertebral spectrum. AB - Oculoauriculovertebral spectrum, or Goldenhar syndrome, is characterized by varying degrees of prevalently unilateral underdevelopment of craniofacial structures (orbit, ear, mandible) and spinal anomalies. We report the prenatal ultrasonographic diagnosis made at 24 weeks' gestation in a family with a negative history. The prenatal diagnosis was suspected due to the presence of marked hemifacial microsomia and moderate ipsilateral cerebellar hemisphere hypoplasia in the absence of facial clefting. PMID- 15287061 TI - Quantitative three-dimensional power Doppler ultrasound predicts the outcome of placental chorioangioma. AB - The relationship of large and vascularized chorioangiomas to adverse pregnancy outcome is well recognized. We present a patient with a large placental tumor and signs of impending fetal cardiac failure. The angioarchitecture of the tumor depicted by three-dimensional (3D) power Doppler ultrasound enabled us to accurately diagnose a placental chorioangioma. During the follow-up period, quantitative flow data obtained using 3D power Doppler indicated altered hemodynamics in the tumor and concomitant improvement in the condition of the fetus, enabling us to manage the mother conservatively. Spontaneous delivery occurred at 38 weeks without any complications. This report demonstrates the potential value of 3D power Doppler in prenatal diagnosis and monitoring of pregnancies complicated by large, vascularized chorioangioma. PMID- 15287062 TI - Prenatal sonographic diagnosis of cardiac hemangioma with postnatal spontaneous regression. PMID- 15287063 TI - First-trimester diagnosis of conjoined twins after in-vitro fertilization-embryo transfer (IVF-ET) at blastocyst stage. PMID- 15287064 TI - Some more about fetal intrapericardial teratoma. PMID- 15287065 TI - Three-dimensional sonographic diagnosis of vasa previa. PMID- 15287066 TI - Methods to study the phytochemistry and bioactivity of essential oils. AB - Many essential oils are extracted, analysed and their main components are identified, characterised and then published without any biological testing whatsoever. Their useful biological activities can remain unknown for years. Yet, the search for these activities often increases our knowledge of the potential use of oils in therapeutics. Therefore, there is a real need for a simple, reliable and reproducible methods to study the bioactivity of essential oils and their constituents which can detect a broad spectrum of action or specific pharmacological activities in aromatic plants. These methods can then be employed by natural product chemists, pharmacologists and biologists to conduct their scientific research and to valorize natural products. Standardisation of some of these methods is therefore desirable to permit more comprehensive evaluation of plant oils, and greater comparability of the results obtained by different investigators. PMID- 15287067 TI - Stereospecific effects of catechin isomers on insulin induced lipogenesis in 3T3 L1 cells. AB - We studied the influence of (+)- and (-)-catechin contained in green tea on insulin induced lipogenesis in 3T3-L1 cells. In 14 days of culture with insulin, the intracellular triglyceride concentration and the activities of glycerophosphate dehydrogenase, a marker of adipose conversion, increased. The addition of 0.02 mg/ml (+)-isomer stimulated the accumulation of triglyceride induced by insulin, but the addition of the same concentration of (-)-isomer inhibited lipogenesis. The activities of glycerophosphate dehydrogenase were changed by (+)- and (-)-catechin in the same direction as the corresponding changes in triglyceride accumulation. These data suggest a biological significance of catechins, with opposite effects on lipid metabolism depending on the isomer. PMID- 15287068 TI - The in vitro antioxidant and antimicrobial activities of the essential oil and methanol extracts of Achillea biebersteini Afan. (Asteraceae). AB - The essential oil and methanol extracts from A. biebersteinii Afan. (Asteraceae) were evaluated for their antimicrobial and antioxidant activities in vitro. The oil showed stronger antimicrobial activity than the extracts. Their antioxidant features were also evaluated using diphenyl-1-picrylhydrazyl (DPPH), inhibition of superoxide and hydroxyl radicals and inhibition of the lipid peroxidation assays. Particularly, polar subfraction of the methanol extract showed antioxidant activity. The GC-MS analysis of the oil has resulted in the identification of 23 components; piperitone, eucalyptol, camphor, chrysanthenone and borneol were the main components. Antimicrobial activity tests carried out with the fractions of the oil showed that the activity was mainly observed in those containing eucalyptol and camphor, in particular, followed by borneol and piperitone. PMID- 15287069 TI - Inhibitory effects of leaf extracts of Stachytarpheta jamaicensis (Verbenaceae) on the respiratory burst of rat macrophages. AB - The anti-oxidant effects of ethyl acetate (EAcE) and n-hexane extracts (nHE) of dried leaves of Stachytarpheta jamaicensis Vahl. (Verbenaceae) on the reactive oxygen species (ROS) generating during the respiratory burst of rat peritoneal macrophages were investigated. Only EAcE, at concentrations between 0.4 and 40 microg/ml, inhibited the extracellular release of oxygen radicals by resident peritoneal macrophages stimulated with phorbol-12-myristate 13-acetate (PMA). At concentrations above 40 microg/ml, EAcE inhibited the production of nitric oxide (NO) in macrophages stimulated in vivo with sodium thioglycollate then in vitro with lipopolysaccharide (LPS) and gamma-interferon (IFN-gamma). nHE extracts at concentrations between 0.4 and 40 microg/ml did not scavenge (O-)2 generated enzymatically by hypoxanthine/xanthine oxidase (HX/XO) system, but EAcE at the same concentrations showed potent (O-)2-scavenging activity. At 40 microg/ml, EAcE also inhibited XO activity. These results suggest that the EAcE extract of S. jamaicensis may be a potential pharmaceutical value in treatment of immunopathological diseases related to oxidative stress. PMID- 15287070 TI - Antifungal constituents of Clytostoma ramentaceum and Mansoa hirsuta. AB - Ethanol extracts of Clytostoma ramentaceum Bur. & K. Schum and Mansoa hirsuta DC. (Bignoniaceae) inhibited the growth of standardized cultures of Aspergillus niger and Fusarium oxysporum, at concentrations of 400 microg and 500 microg, in bioautographic assays. The activity-guided fractionation of C. ramentaceum extract afforded ursolic acid and 2-(3',4'-dihydroxyphenyl) ethanol, both active against the test fungi (100 microg). These compounds are reported for the first time in C. ramentaceum and were not detected in M. hirsuta extract, according to HPLC analysis. The bioguided study of M. hirsuta resulted in five active fractions (100 to 200 microg), whose GC-MS analysis allowed us to identify 11 compounds, mostly alkanols and alkanodiols, that may be regarded as the antifungal constituents of M. hirsuta. PMID- 15287071 TI - Biological screening of Nigella damascena for antimicrobial and molluscicidal activities. AB - The essential oil, various extracts at different polarity, fractions, and pure compounds obtained from Nigella damascena plants and seeds were screened for biological activity. Antimicrobial tests showed the essential oil to be active only against Gram positive bacteria; among the extracts, the BuOH was active against Pseudomonas aeruginosa and Staphylococcus aureus. Molluscicidal activity was absent. PMID- 15287072 TI - Lepidium peruvianum chacon restores homeostasis impaired by restraint stress. AB - Lepidium peruvianum root has been traditionally utilized by native Peruvians, since before the time of the Incas, for both nutritional and putative medicinal purposes as an adaptogen and also to enhance fertility in humans and animals. The present research was conducted to evaluate the anti-stress activity of the methanolic extract of Lepidium peruvianum. The drug is capable of attenuating or even eliminating variations in homeostasis produced by stress since it reduces or abolishes stress-induced ulcers, elevated corticosterone levels, the reduction of glucose and the increase in the weight of adrenal glands produced by stress. It also eliminates the decrease in free fatty-acids (FFA) in plasma produced by stress and we obtain a positive result in the forced-swimming test. Thus, it did not appear to affect restraint stress-induced immunosuppression. PMID- 15287073 TI - Structural criteria for depigmenting mechanism of arbutin. AB - Arbutin, hydroquinone-O-beta-D-glucopyranoside (1) was found to inhibit the oxidation of l-tyrosine (monophenolase activity) catalyzed by mushroom tyrosinase. However, arbutin itself was oxidized as a monophenol substrate at an extremely slow rate, and this oxidation was accelerated as soon as catalytic amounts (0.01 mM) of l-3,4-dihydroxyphenylalanine (L-DOPA) became available as a cofactor. The result observed was supported by monitoring oxygen consumption. The depigmenting mechanism of arbutin previously reported is supportable if a cofactor is not available in the melanocytes. The combination with L-ascorbic acid is a useful application, particularly when oxygen is limited. PMID- 15287074 TI - Antiviral activity of the red marine alga Ceramium rubrum. AB - An extract from the red marine alga Ceramium rubrum (Huds.) Ag. from the Bulgarian Black Sea seacoast considerably inhibited the reproduction of influenza viruses type A and B in vitro and in ovo. The virus-induced cytopathogenic effect (CPE), infectious virus yields and the production of hemagglutinin were all reduced at non-toxic concentrations of the extract. The virus-inhibitory effect was selective, dose-related and strain-specific; selectivity indices ranged 9.5 68.3. The inhibition affected adsorption as well as the intracellular stages of viral replication. The extract inhibited also the reproduction of herpes simplex virus (HSV) type 1 and type 2 in cell cultures. The preparation exhibited a strong HSV-inactivating activity. PMID- 15287075 TI - Evaluation of the gastric antiulcerogenic effects of Portulaca oleracea L. extracts in mice. AB - Portulaca oleracea is commonly used in Iranian folk medicine. The aqueous and ethanolic extracts were studied in mice for their ability to inhibit gastric lesions induced by HCl or absolute ethanol. In addition, their effects on gastric acid secretion were measured. Both extracts showed a dose-dependent reduction in severity of ulcers. The highest dose of extracts exerted similar activity to sucralfate. The oral and intraperitoneal administration of extracts reduced the gastric acidity in pylorus-ligated mice. These results suggest that Portulaca oleracea has gastroprotective action and validates its use in folk medicine for gastrointestinal diseases. PMID- 15287076 TI - Attenuating effect of a traditional korean formulation, Paeng-Jo-Yeon-Nyeon-Baek Ja-In-Hwan (PJBH), on hydrogen peroxide-induced injury in PC12 cells. AB - The Paeng-Jo-Yeon-Nyeon-Baek-Ja-In-Hwan (PJBH) prescription is a dried decoctum consisting of a mixture of 18 medicinal herbs that include Semen Biotae, Fructus Torilis seu cnidii, Fructus Rubi, Herba Dendrobii, Radix Morindae officinalis, Cortex Eucommiae, Radix Aspragi, Radix Polygalae, Radix Dipsaci, Ramulus Cinnamomi, Rhizoma Acori graminei, Rhizoma Alismatis, Rhizoma Dioscoreae, Radix Ginseng, Radix Rehmanniae preparata, Fructus corni, Fructus Schisandrae and Herba Cistanches. The effect of PJBH extracts on H2O2-induced toxicity in the rat pheochromocytoma line PC12 was examined by measurements of cell lesion, level of lipid peroxidation and antioxidant enzyme activities, since free radicals are involved in neurodegeneration in Alzheimer's disease (AD). After a 30 min exposure of the cells to H2O2 (150 microM), a marked decrease in cell survival, activities of glutathione peroxidase and catalase as well as an increased production of malondialdehyde (MDA) were found. Pretreatment of the cells with PJBH (0.5-10 microg/ml) prior to H2O2 exposure significantly elevated cell survival, antioxidant enzyme activities and resulted in a decrease in the level of MDA. The effects of the PJBH on hydrogen peroxide-induced injury in PC12 cells were also examined. PJBH had a remarkable elevating effect on catalase and GSH-Px activities as well as cell survival, suggesting that cytoprotective effects of the PJBH are involved in stimulation against intermediate concentrations of H2O2 induced PC12 cell injury. The above-mentioned neuroprotective effects were also compared with the effect of tacrine. The results suggest that PJBH has potential for use as a novel neuronal therapeutic agent. PMID- 15287077 TI - Antifungal activity of actinomycetes from Cuban soils. AB - This study reports on the capacity of actinomycete strains isolated from Cuban soils to produce antifungal agents. The antimicrobial activities were determined by susceptibility disk assay methods. We isolated 563 different actinomycetes and 286 produced compounds with antifungal activity. Our screening method indicated the presence of many possible polyene macrolide antibiotics and the important antifungal activity in the soils rich in minerals. PMID- 15287078 TI - Antibacterial activity of the Chinese traditional medicine, Zi Hua Di Ding. AB - The antibacterial activity of extracts from species of plants used in the Chinese medicine, Zi Hua Di Ding (Viola yedoensis, V. prionantha, Corydalis bungeana and Gueldenstaedtia verna), was tested against Bacillus subtilis and Pseudomonas syringae using a bioautographic assay. The petroleum ether and ethyl acetate extracts of all four species of plants showed activity against both species of bacteria, whereas the methanol and aqueous methanol extracts were inactive. Three fractions from the petroleum ether extracts of V. yedoensis, V. prionantha and C. bungeana showed activity against B. subtilis at 6.25 microg/mL. Preliminary analysis of these active fractions indicates that they contain long chain carboxylic acids. PMID- 15287079 TI - Quantile estimation following non-parametric phase I clinical trials with ordinal response. AB - A non-parametric multi-dimensional isotonic regression estimator is developed for use in estimating a set of target quantiles from an ordinal toxicity scale. We compare this estimator to the standard parametric maximum likelihood estimator from a proportional odds model for extremely small data sets. A motivating example is from phase I oncology clinical trials, where various non-parametric designs have been proposed that lead to very small data sets, often with ordinal toxicity response data. Our comparison of estimators is performed in conjunction with three of these non-parametric sequential designs for ordinal response data, two from the literature and a new design based on a random walk rule. We also compare with a non-parametric design for binary response trials, by keeping track of ordinal data for estimation purposes, but dichotomizing the data in the design phase. We find that a multidimensional isotonic regression-based estimator far exceeds the others in terms of accuracy and efficiency. A rule by Simon et al. (J. Natl. Cancer Inst. 1997; 89:1138-1147) yields particularly efficient estimators, more so than the random walk rule, but has higher numbers of dose limiting toxicity. A small data set from a leukemia clinical trial is analysed using our multidimensional isotonic regression-based estimator. PMID- 15287080 TI - A general statistical principle for changing a design any time during the course of a trial. AB - A general method is presented that allows the researcher to change statistical design elements such as the residual sample size during the course of an experiment, to include an interim analysis for early stopping when no formal rule for early stopping was foreseen, to increase or reduce the number of planned interim analyses, to change time points and the type I error spending function for the further design of interim analyses, or to change the test statistic, the outcome measure, etc. At the time of a pre-planned interim analysis for early stopping or at any time of an interim look without spending part of the type I error level the method offers the option to completely redesign the remaining part of the trial, without affecting the type I error level. The method is described in the usual Brownian motion model and extended to the general context of statistical decision functions. It is based on the conditional rejection probability of a decision variable. PMID- 15287081 TI - A new approach to modelling interactions between treatment and continuous covariates in clinical trials by using fractional polynomials. AB - We consider modelling interaction between a categoric covariate T and a continuous covariate Z in a regression model. Here T represents the two treatment arms in a parallel-group clinical trial and Z is a prognostic factor which may influence response to treatment (known as a predictive factor). Generalization to more than two treatments is straightforward. The usual approach to analysis is to categorize Z into groups according to cutpoint(s) and to analyse the interaction in a model with main effects and multiplicative terms. The cutpoint approach raises several well-known and difficult issues for the analyst. We propose an alternative approach based on fractional polynomial (FP) modelling of Z in all patients and at each level of T. Other prognostic variables can also be incorporated by first constructing a multivariable adjustment model which may contain binary covariates and FP transformations of continuous covariates other than Z. The main step involves FP modelling of Z and testing equality of regression coefficients between treatment groups in an interaction model adjusted for other covariates. Extensive experience suggests that a two-term fractional polynomial (FP2) function may describe the effect of a prognostic factor on a survival outcome quite well. In a controlled trial, this FP2 function describes the prognostic effect averaged over the treatment groups. We refit this function in each treatment group to see if there are substantial differences between groups. Allowing different parameter values for the chosen FP2 function is flexible enough to detect such differences. Within the same algorithm we can also deal with the conceptually different cases of a predefined hypothesis of interaction or searching for interactions. We demonstrate the ability of the approach to detect and display treatment/covariate interactions in two examples from controlled trials in cancer. PMID- 15287082 TI - A measurement error model with a Poisson distributed surrogate. AB - We study a linear model in which one of the covariates is measured with error. The surrogate for this covariate is the event count in unit time. We model the event count by a Poisson distribution, the rate of which is the unobserved true covariate. We show that ignoring the measurement error leads to inconsistent estimators of the regression coefficients and propose a set of unbiased estimating equations to correct the bias. The method is computationally simple and does not require using supplemental data as is often the case in other measurement error analyses. No distributional assumption is made for the unobserved covariate. The proposed method is illustrated with an example from the Wisconsin Sleep Cohort Study. PMID- 15287083 TI - On sample size for sensitivity and specificity in prospective diagnostic accuracy studies. AB - The design of a study of disease screening tests may be based on hypothesis tests for the sensitivity and specificity of the tests. The case-control study requires knowledge of the disease status of patients at the time of enrollment. This may not be possible in a prospective setting, when the gold standard is obtained subsequent to the initial screening and the number of diseased individuals is random and can not be fixed by design. Several ad hoc procedures for determining the total sample size are commonly used by practitioners, for example, the prevalence inflation method. The properties of these methods are not well understood. We develop a formal method for sample size and power calculations based on the unconditional power properties of the test statistics. The approach provides novel insights into the behaviour of the commonly used methods. We find that the ad hoc prevalence inflation method may serve as a useful approximation to our rigorous framework for sample size determination in the prospective set up. The design of a large population-based study of mammography for breast cancer screening illustrates the key issues. PMID- 15287084 TI - A sample size computation method for non-linear mixed effects models with applications to pharmacokinetics models. AB - We propose a simple method to compute sample size for an arbitrary test hypothesis in population pharmacokinetics (PK) studies analysed with non-linear mixed effects models. Sample size procedures exist for linear mixed effects model, and have been recently extended by Rochon using the generalized estimating equation of Liang and Zeger. Thus, full model based inference in sample size computation has been possible. The method we propose extends the approach using a first-order linearization of the non-linear mixed effects model and use of the Wald chi(2) test statistic. The proposed method is general. It allows an arbitrary non-linear model as well as arbitrary distribution of random effects characterizing both inter- and intra-individual variability of the mixed effects model. To illustrate possible uses of the method we present tables of minimum sample sizes, in particular, with an illustration of the effect of sampling design on sample size. We demonstrate how (D-)optimal or frequent sampling requires fewer subjects in comparison to a sparse sampling design. We also present results from Monte Carlo simulations showing that the computed sample size can produce the desired power. The proposed method greatly reduces computing times compared with simulation-based methods of estimating sample sizes for population PK studies. PMID- 15287085 TI - Validation and updating of predictive logistic regression models: a study on sample size and shrinkage. AB - A logistic regression model may be used to provide predictions of outcome for individual patients at another centre than where the model was developed. When empirical data are available from this centre, the validity of predictions can be assessed by comparing observed outcomes and predicted probabilities. Subsequently, the model may be updated to improve predictions for future patients. As an example, we analysed 30-day mortality after acute myocardial infarction in a large data set (GUSTO-I, n = 40 830). We validated and updated a previously published model from another study (TIMI-II, n = 3339) in validation samples ranging from small (200 patients, 14 deaths) to large (10,000 patients, 700 deaths). Updated models were tested on independent patients. Updating methods included re-calibration (re-estimation of the intercept or slope of the linear predictor) and more structural model revisions (re-estimation of some or all regression coefficients, model extension with more predictors). We applied heuristic shrinkage approaches in the model revision methods, such that regression coefficients were shrunken towards their re-calibrated values. Parsimonious updating methods were found preferable to more extensive model revisions, which should only be attempted with relatively large validation samples in combination with shrinkage. PMID- 15287086 TI - A simple significance test for quantile regression. AB - Where OLS regression seeks to model the mean of a random variable as a function of observed variables, quantile regression seeks to model the quantiles of a random variable as functions of observed variables. Tests for the dependence of the quantiles of a random variable upon observed variables have only been developed through the use of computer resampling or based on asymptotic approximations resting on distributional assumptions. We propose an exceedingly simple but heretofore undocumented likelihood ratio test within a logistic regression framework to test the dependence of a quantile of a random variable upon observed variables. Simulated data sets are used to illustrate the rationale, ease, and utility of the hypothesis test. Simulations have been performed over a variety of situations to estimate the type I error rates and statistical power of the procedure. Results from this procedure are compared to (1) previously proposed asymptotic tests for quantile regression and (2) bootstrap techniques commonly used for quantile regression inference. Results show that this less computationally intense method has appropriate type I error control, which is not true for all competing procedures, comparable power to both previously proposed asymptotic tests and bootstrap techniques, and greater computational ease. We illustrate the approach using data from 779 adolescent boys age 12-18 from the Third National Health and Nutrition Examination Survey (NHANES III) to test hypotheses regarding age, ethnicity, and their interaction upon quantiles of waist circumference. PMID- 15287087 TI - Hierarchical modelling of small area and hospital variation in short-term prognosis after acute myocardial infarction. A longitudinal study of 35- to 74 year-old men in Denmark between 1978 and 1997. AB - Models for analysis of trends in hospital and small area variation in case fatality after acute myocardial infarction are presented. The data are from administrative registries in Denmark. Hierarchical modelling in a logistic regression with a Bayesian approach is used. Model selection is undertaken using the deviance and the Bayesian information criteria. There is a modest trend for hospital variation in case-fatality rates that coincides with the introduction of new treatment strategies. This hospital variation is considerably larger than the variation at the area level. There is no trend for variation of the case-fatality rates at the area level. Unstructured random effects slightly outperform spatially correlated random effects at the area level. Somewhat high correlations over time within hospitals and within areas were detected for the case-fatality rates. Heavy-tailed distributions (T-distributions) could be an alternative for the random effect distribution in data from administrative registries and compete in the model selection with the normal distribution in this study. PMID- 15287089 TI - Anti-tumor effects and lack of side effects in mice of an immunotoxin directed against human and mouse prostate-specific membrane antigen. AB - BACKGROUND: Prostate-specific membrane antigen (PSMA) is a transmembrane protein that is largely restricted to prostatic epithelial cells in humans and is strongly upregulated on prostatic carcinoma cells. It is also expressed on the endothelium of tumor vasculature in humans, but not on the vasculature of normal tissues. Expression of low levels of PSMA has also been found on non-vascular cells in several normal tissues, most prominently on the brain and kidney in humans. PSMA is an excellent candidate for targeting prostate cancer or targeting tumor vasculature of various solid tumors. The high potential clinical benefit of these agents has prompted the search for an animal model in which to assess the efficacy and safety of anti-PSMA monoclonal antibody (mAb)-based therapies. METHODS: A rat monoclonal antibody, E6 that recognizes both mouse and human PSMA was generated using conventional hybridoma techniques. The antibody was characterized by enzyme-linked immunosorbent assay (ELISA), Western blot, and immunohistochemistry. An immunotoxin composed of E6, antibody and deglycosylated ricin A-chain (dgA) was prepared chemically. The anti-tumor effects of the immunotoxin were determined in vitro and in mice bearing subcutaneous LnCaP human prostate tumors, which express PSMA on the tumor cell surface. RESULTS: E6 recognizes the extracellular domain of both human and mouse PSMA in ELISA, immunoblot and by immunohistochemistry. E6 strongly stained the vascular endothelium of tumors from humans but not from mice. E6 stained proximal tubules in mouse and human kidneys, and neurons in the mouse and human hippocampus but, unlike the human, did not detectably stain epithelial cells in mouse prostate or small intestine. An E6-dgA immunoconjugate strongly inhibited the growth of LnCaP tumor xenografts without causing apparent toxicity to the mice. Histological observation indicated that the anti-tumor effects were mediated through direct cytotoxic effects on the tumor cells. CONCLUSIONS: We have generated and characterized a rat mAb (E6) that reacts specifically with both human and mouse PSMA and have demonstrated that an immunotoxin constructed from E6 is safe and effective against human prostatic carcinoma cells growing subcutaneously in nude mice. PMID- 15287090 TI - Anti-prostate specific membrane antigen designer T cells for prostate cancer therapy. AB - BACKGROUND: Designer T cells are T lymphocytes engineered toward specific antibody-type membrane antigens through chimeric immunoglobulin-T-cell receptor (IgTCR) genes that have been used for adoptive cellular immunotherapy. We have extended this approach to prostate specific membrane antigen (PSMA) as a means to attack prostate cancer. METHODS: A chimeric anti-PSMA IgTCR gene was constructed based on an anti-PSMA monoclonal antibody, 3D8. Both T-cell lines and primary cultured human T lymphocytes were transduced with the chimeric anti-PSMA IgTCR construct and were analyzed for IgTCR expression, specific activation by PSMA, cytotoxicity against PSMA-expressing tumor cells in vitro, and retardation of tumor growth in an animal model. RESULTS: The IgTCR was incorporated into the TCR CD3 complex and formed a functional chimeric complex. The IgTCR-modified T cells were specifically activated through the chimeric receptor with PSMA as measured by IL-2 production and increased CD25 expression and specifically lysed the PSMA expressing prostate cancer cells in vitro as well as retarded tumor growth in an animal model. CONCLUSIONS: The anti-PSMA designer T cells exhibit an antibody type specificity that can recognize PSMA expressing tumor cells in a MHC independent fashion, resulting in T-cell activation, target cell lysis in vitro and inhibition of tumor growth in vivo. PMID- 15287091 TI - Transgenic mouse with human mutant p53 expression in the prostate epithelium. AB - BACKGROUND: Apoptosis is disrupted in prostate tumor cells, conferring a survival advantage. p53 is a nuclear protein believed to regulate cancer progression, in part by inducing apoptosis. To test this possibility in future studies, the objective of the present study was to generate a transgenic mouse model expressing mutant p53 in the prostate (PR). METHODS: Transgene incorporation was tested using Southern analysis. Expression of mutant p53 protein was examined using immunofluorescence microscopy. Apoptosis in the PR was evaluated using the Tunnel method. RESULTS: A construct, consisting of the rat probasin promoter and a mutant human p53 fragment, was prepared and used to generate transgenic mice. rPB-mutant p53 transgene incorporation, as well as nuclear accumulation of mutant human p53 protein, was demonstrated. Prostatic intraepithelial neoplasia (PIN) III and IV were found in PR of 52-week old transgenic mice, whereas no pathological changes were found in the other organs examined. PR ability to undergo apoptosis following castration was reduced in rPB-mutant p53 mice as compared to non transgenic littermates. CONCLUSIONS: Transgenic rPB-mutant p53 mice accumulate mutant p53 protein in PR, resulting in neoplastic lesions and reduced apoptotic potential in the PR. Breeding rPB-mutant p53 mice with mice expressing an oncogene in their PR will be useful in examining interactions of multiple genes that result in progression of slow growing prostate tumors expressing oncogenes alone to metastatic cancer. PMID- 15287092 TI - Ionizing radiation enhances the therapeutic potential of TRAIL in prostate cancer in vitro and in vivo: Intracellular mechanisms. AB - BACKGROUND: We assessed the influence of sequential treatment of ionizing radiation followed by tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) on intracellular mechanisms of apoptosis of prostate tumor cells in vitro and in vivo. METHODS: Prostate normal and cancer cells were exposed to irradiation and TRAIL. Four- to 6-week-old athymic nude mice were injected s.c. with PC-3 tumor cells. Tumor bearing mice were exposed to irradiation and TRAIL, either alone or in combination (TRAIL after 24 hr of irradiation), and tumor growth, apoptosis, and survival of mice were examined. Expressions of death receptors, Bcl-2 family members, and caspase were measured by Western blotting, ELISA, and ribonuclease protection assay; tumor cellularity was assessed by H&E staining; inhibition of p53 was performed by RNA interference (RNAi) technology, and apoptosis was measured by annexin V/propidium iodide staining, and terminal deoxynucleotidyltransferase-mediated nick end labeling assay. RESULTS: Irradiation significantly augmented TRAIL-induced apoptosis in prostate cancer cells through upregulation of DR5, Bax, and Bak, and induction of caspase activation. Dominant negative FADD and p53 siRNA inhibited the synergistic interaction between irradiation and TRAIL. The pretreatment of cells with irradiation followed by TRAIL significantly enhanced more apoptosis than single agent alone or concurrent treatment. Furthermore, irradiation sensitized TRAIL resistant LNCaP cells to undergo apoptosis. The sequential treatment of xenografted mice with irradiation followed by TRAIL-induced apoptosis through activation of caspase-3, induction of Bax and Bak, and inhibition of Bcl-2, and completely eradicated the established tumors with enhanced survival of nude mice. CONCLUSION: The sequential treatment with irradiation followed by TRAIL can be used as a viable option to enhance the therapeutic potential of TRAIL in prostate cancer. PMID- 15287093 TI - Adenovirus-mediated expression of a soluble fibroblast growth factor receptor inhibits in vitro growth of prostate DU145 cells. AB - BACKGROUND: Fibroblast growth factor (FGF) family plays a key role in prostate cancer. The soluble FGF receptor (sFGFR) has been studied with regards to inhibiting cancer growth and was shown to have a dominant negative effect on cellular signaling and function. Using replication deficient adenovirus-mediated gene transfer, we tested if sFGFR expression may have a suppressive effect on in vitro growth of prostate cancer cells. METHODS: Western analysis was used to verify expression of sFGFR1 and to examine the effect of sFGFR1 on MAP kinase phosphorylation. The effect on proliferation and invasiveness of DU145 cells was examined using the WST-1 and Matrigel Invasion assay, respectively. RESULTS: Activation of MAP kinase (pERK1 and 2) by exogenous FGF1, 2, and 7 was suppressed to baseline levels by sFGFR, which was not seen with EGF. Proliferation and invasion of DU145 cells were significantly suppressed by sFGFR. CONCLUSIONS: A replication deficient adenoviral vector system reproducibly expresses sFGFR in prostate cells. Suppression of in vitro growth in DU145 cells by sFGFR provides the basis of a novel therapeutic approach. PMID- 15287094 TI - Chronic inflammation in benign prostate hyperplasia is associated with focal upregulation of cyclooxygenase-2, Bcl-2, and cell proliferation in the glandular epithelium. AB - BACKGROUND: Chronic inflammation has been suggested to be linked to the development and progression of prostate cancer. An inflammatory microenvironment may support the development of malignancy by upregulation of proinflammatory cytokines and cyclooxygenase-2 (COX-2). Recent studies have suggested that COX-2 is upregulated in cancer and in proliferative inflammatory atrophy (PIA) of the prostate. METHODS: Immunohistochemistry (IHC) was used to investigate the expression of COX-2 in prostate epithelium. The relationships between COX-2 expression and inflammatory cells, proliferation (proliferating cell nuclear antigen (PCNA) and Ki-67), and apoptosis (Bcl-2) were studied in 45 BPH samples. RESULTS: COX-2 expression was detected in prostate luminal epithelial cells in all 45 samples studied. The overall percentage of COX-2 positive glands was 7.5%, distributed as 0.2% positive glands in normal prostate tissue, 25.7% in postatrophic hyperplasia (PAH), and 11.9% in simple atrophy (SA). The highest proliferation index was found in COX-2 positive stained epithelium. COX-2 expression was associated with Bcl-2 immunostaining in atrophic lesions (P < 0.0001). T lymphocytes and macrophages were the predominant inflammatory cells related to the COX-2 expression in prostate epithelium. CONCLUSIONS: The data demonstrate that T lymphocytes and macrophages appeared to play an important role in the induction of COX-2 expression in prostate epithelium, which was associated with increased cell proliferation and possibly, due to overexpression of Bcl-2, apoptotic resistance. This suggests that pro-inflammatory cytokines released by adjacent inflammatory cells may induce COX-2 in the epithelial cells in prostate atrophic lesions. In addition, COX-2 expressing cells may be involved in the pathogenesis of prostate cancer. PMID- 15287095 TI - Effect of permixon on human prostate cell growth: lack of apoptotic action. AB - BACKGROUND: Permixon, a phytotherapeutic agent derived from the saw palmetto or Serenoa repens plant, is a lipid/sterol extract that is believed to interfere with 5alpha-reductase activity, thus inhibiting prostate growth. In this study, we investigated the magnitude and specificity of the effect of Permixon on cell proliferation and apoptosis in human prostate cancer cells. METHODS: The effect of Permixon was examined in androgen-independent PC-3 prostate cancer cells, androgen-sensitive LNCaP prostate cancer cells, and MCF-7 breast cancer cells in vitro. Cell growth, apoptosis induction, and cell proliferation was studied after exposure to Permixon at two concentrations (10 and 100 microg/ml). Cell proliferation and cell cycle progression were determined after 24 hr on the basis of (3)[H]-thymidine incorporation assay and flowcytometric analysis, respectively. Apoptosis induction was evaluated in treated and untreated cultures using the Hoescht staining and caspase-3 activation. RESULTS: Exposure of prostate and breast cancer cells to a high dose of Permixon (100 microg/ml) resulted in a significant decrease in the rate of cell growth; an effect that was not time-dependent and was not associated with cell cycle arrest. Permixon treatment (at either high or low dose) had no effect on apoptosis induction in prostate cancer cell lines (P > 0.6). Furthermore, in vitro Permixon was a weak inhibitor of 5alpha-reductase activity type 2 in prostatic homogenates. CONCLUSIONS: The results indicate the ability of Permixon to affect prostate cancer cell growth without inducing apoptosis or cell cycle arrest. This effect was not prostate-specific and was only manifested at high concentrations of Permixon. Furthermore our findings indicate that Permixon is weak inhibitor of 5alpha-reductase compared to finasteride. This study challenges previous evidence on the anti-growth effect of Permixon in the prostate and its ability to inhibit 5alpha-reductase activity, while questioning apoptosis as a mechanism of action of this phytotherapeutic against prostate growth, a concept that may have therapeutic significance. PMID- 15287096 TI - Surgical margin and Gleason score as predictors of postoperative recurrence in prostate cancer with or without chromosome 8p allelic imbalance. AB - BACKGROUND: Identification of prostate cancer patients at risk for postoperative disease recurrence is an important clinical issue. Existing pathological markers can predict disease recurrence only to a certain extent, and there is a need for more accurate predictors. METHODS: Using "counting alleles," a novel experimental method, we determined allelic status of chromosome 8p in 107 prostatectomy specimens. Statistical analyses examined the association between pathologic predictors (Gleason score, stage, surgical margin, etc.) and cancer recurrence in patients with and without 8p allelic imbalance (8p AI). RESULTS: 8p AI cancers were more likely to recur in the presence of a positive surgical margin, whereas recurrence of 8p retaining tumors was associated with the Gleason score, but not with the surgical margin. CONCLUSIONS: Our findings suggest that chromosome 8p allelic status affects the predictive value of "traditional" markers of prostate cancer recurrence. If confirmed by larger studies, these results may have important clinical implications. PMID- 15287097 TI - Nitroprusside stimulates mitochondrial aconitase gene expression through the cyclic adenosine 3',5'-monosphosphate signal transduction pathway in human prostate carcinoma cells. AB - BACKGROUND: Mitochondrial aconitase (mACON), an iron-requiring enzyme, is a major target of nitric oxide (NO) in cells, which causes the oxidant-mediated disruption of the [4Fe-4S] prosthetic group of the enzyme. In this study, the effect of NO on mACON enzymatic activity and gene expression were investigated. METHODS: Three NO generators, sodium nitroprusside (SNP), S-nitoso-N acetylpenicillamine (SNAP), and 3-morpholinosydnonimine (SIN) were used to determine the regulation of mACON enzymatic activity by NO. The effect of SNP on mACON, which modulates citrate secretion and cellular bioenergetics in PC-3 cells, was investigated by determining the effect of SNP on mACON gene expression using Western blot and transient gene expression assays. RESULTS: SNP upregulated mACON enzymatic activity and gene expression in PC-3 cells. However, treating cells with other NO generators, SNAP and SIN, resulted in decreased mACON enzymatic activity. The addition of ascorbic acid to the SNP treatment resulted in a decrease in mACON enzymatic activity and gene expression. Our results showed that both SNP and dibutyryl-cAMP increased the mACON promoter activity 2-fold while the effect was blocked by adding H-89. Mutation of the cAMP response element (CRE) to the AGAGCT abolished the activating effects of SNP and dibutyryl cAMP on mACON promoter activity. CONCLUSIONS: These results establish the function of nitroprusside as a signaling molecule for mACON gene expression through the cAMP signal transduction pathway in human prostatic carcinoma cells. PMID- 15287099 TI - The integration of Darwinism and evolutionary morphology: Alexej Nikolajevich Sewertzoff (1866-1936) and the developmental basis of evolutionary change. AB - The growth of evolutionary morphology in the late 19th and early 20th centuries was inspired by the work of Carl Gegenbaur (1826-1903) and his protege and friend Ernst Haeckel (1834-1919). However, neither of them succeeded in creating and applying a strictly Darwinian (selectionist) methodology. This task was left to the next generation of evolutionary morphologists. In this paper we present a relatively unknown researcher, Alexej Nikolajevich Sewertzoff (1866-1936) who made important contributions towards a synthesis of Darwinism and evolutionary morphology. PMID- 15287100 TI - When did theropods become feathered?--evidence for pre-Archaeopteryx feathery appendages. AB - Filamentous impressions associated with locomotive theropod tracks in the Lower Jurassic Turners Falls Formation of western Massachusetts, U.S.A. represent the oldest evidence of feathered dinosaurs. Feather impressions are preserved with sitting traces which bear integumentary structures along the outlines of the pre pubic and ischiadic impressions. Extant palaeognathous down feathers provide a valuable comparative model for these filamentous integumental structures and for similar structures described in Chinese theropods from younger deposits. The described morphologies are congruent with Stage II of Prum ('99) and support that plumulaceous morphologies evolved before the origin of the rhachis and the planar vane. Appearance of feathery appendages in theropods may be linked to evolution of higher metabolic rates, improved locomotory abilities, and/or distinct behavior(s) and visual communication. Development of feathery integument might have also played a crucial role in the competitiveness and successful radiation of maniraptoriform theropods and their actively flying descendants in the Jurassic. PMID- 15287101 TI - Dermo-epidermal interactions in reptilian scales: speculations on the evolution of scales, feathers, and hairs. AB - The dermal influence on the epidermis during scale formation in reptiles is poorly known. Cells of the superficial dermis are not homogeneously distributed beneath the epidermis, but are instead connected to specific areas of the epidermis. Dermal cells are joined temporarily or cyclically through the basement membrane, with the reactive region of the epidermis forming specific regions of dermo-epidermal interactions. In these regions morphoregulatory molecules may be exchanged between the dermis and the connected epidermis. Possible changes in the localization of these regions in the skin may result in the production of different appendages, in accordance with the genetic makeup of the epidermis in each species. Regions of dermo-epidermal interactions seem to move their position during development. A hypothesis on the development and evolution of scales, hairs, and feathers from sarcopterigian fish to amniotes is presented, based on the different localization and extension of regions of dermo-epidermal interactions in the skin. It is hypothesized that, during phylogenesis, possible variations in the localization and extension of these regions, from the large scales of basic amniotes to those of sauropsid amniotes, may have originated scales with hard (beta)-keratin. In extant reptiles, extended regions of dermo epidermal interaction form most of the expanse of outer scale surface. It is hypothesized that the reduction of large regions of dermo-epidermal interactions into small areas in the skin were the origin of dermal condensations. In mammals, small regions of dermo-epidermal interactions have invaginated, forming the dermal papilla with the associated hair matrix epidermis. In birds, small regions of dermo-epidermal interactions have reduced the original scale surface of archosaurian scales, forming the dermal papilla. The latter has invaginated in association with the collar epidermis from which feathers were produced. PMID- 15287102 TI - Preliminary observations on the spawning conditions of the European amphioxus (Branchiostoma lanceolatum) in captivity. AB - Members of the subphylum Cephalochordata, which include the genus Branchiostoma (i.e. amphioxus), represent the closest living invertebrate relatives of the vertebrates. To date, developmental studies have been carried out on three amphioxus species (the European Branchiostoma lanceolatum, the East Asian B. belcheri, and Floridian-Caribbean B. floridae). In most instances, adult animals have been collected from the field during their ripe season and allowed (or stimulated) to spawn in the laboratory. In any given year, dates of laboratory pawning have been limited by two factors. First, natural populations of these three most studied species of amphioxus are ripe, at most, for only a couple of months each year and, second, even when apparently ripe, animals spawn only at unpredictable intervals of every several days. This limited supply of living material hinders the development of amphioxus as a model system because this limitation makes it more difficult to work out protocols for new laboratory techniques. Therefore we are developing laboratory methods for increasing the number of amphioxus spawning dates per year. The present study found that a Mediterranean population of B. lanceolatum living near the Franco-Spanish border spawned naturally at the end of May and again at the end of June in 2003. Re feeding experiments in the laboratory demonstrated that the gonads emptied at the end of May refilled with gametes by the end of June. We also found that animals with large gonads (both, obtained from the field and kept and fed at the laboratory during several weeks) could be induced to spawn in the laboratory out of phase with the field population if they were temperature shocked (spawning occurred 36 hours after a sustained increase in water temperature from 19 degrees C to 25 degrees C). PMID- 15287103 TI - Molecular evolution in the yeast transcriptional regulation network. AB - We analyze the structure of the yeast transcriptional regulation network, as revealed by chromatin immunoprecipitation experiments, and characterize the molecular evolution of both its transcriptional regulators and their target (regulated) genes. We test the hypothesis that highly connected genes are more important to the function of gene networks. Three lines of evidence-the rate of molecular evolution of network genes, the rate at which network genes undergo gene duplication, and the effects of synthetic null mutation in network genes provide no strong support for this hypothesis. In addition, we ask how network genes diverge in their transcriptional regulation after duplication. Both loss (subfunctionalization) and gain (neofunctionalization) of transcription factor binding play a role in this divergence, which is often rapid. On the one hand, gene duplicates experience a net loss in the number of transcription factors binding to them, indicating the importance of losing transcription factor binding sites after gene duplication. On the other hand, the number of transcription factors that bind to highly diverged duplicates is significantly greater than would be expected if loss of binding played the only role in the divergence of duplicate genes. PMID- 15287104 TI - The clinical-skills examination. PMID- 15287105 TI - The clinical-skills examination. PMID- 15287106 TI - The clinical-skills examination. PMID- 15287107 TI - The clinical-skills examination. PMID- 15287108 TI - The clinical-skills examination. PMID- 15287109 TI - AIDS threat growing throughout Europe. PMID- 15287111 TI - Current awareness in NMR in biomedicine. PMID- 15287110 TI - Highlights from the American Heart Association Annual Scientific Sessions 2003: November 9 to 12, 2003. PMID- 15287112 TI - Earth science: role of fO2 on fluid saturation in oceanic basalt. AB - Assessing the conditions under which magmas become fluid-saturated has important bearings on the geochemical modelling of magmas because volatile exsolution may profoundly alter the behaviour of certain trace elements that are strongly partitioned in the coexisting fluid. Saal et al. report primitive melt inclusions from dredged oceanic basalts of the Siqueiros transform fault, from which they derive volatile abundances of the depleted mantle, based on the demonstration that magmas are not fluid-saturated at their eruption depth and so preserve the mantle signature in terms of their volatile contents. However, in their analysis, Saal et al. consider only fluid-melt equilibria, and do not take into account the homogeneous equilibria between fluid species, which, as we show here, may lead to a significant underestimation of the pressure depth of fluid saturation. PMID- 15287113 TI - Where involuntary commitment, civil liberties, and the right to mental health care collide: an overview of California's mental illness system. PMID- 15287114 TI - Rethinking protections for human subjects. PMID- 15287115 TI - Study finds research consent forms difficult to comprehend. PMID- 15287116 TI - Bush appoints new advisory committee on human subjects. PMID- 15287117 TI - The mythical threat of genetic determinism. PMID- 15287118 TI - New accrediting body forms for human-subjects research. PMID- 15287119 TI - After heated debate, U.S. House votes again to ban cloning. PMID- 15287120 TI - Penn anthropologist fights subpoenas for field notes in medical case. PMID- 15287121 TI - Cloning debate moves to the states: with an impasse in Congress, legislatures are now facing bills on research. PMID- 15287122 TI - Nature takes only tiny steps but still surpasses our reckoning. PMID- 15287123 TI - Banking on DNA: Estonia's genetic database promises medical advances--maybe. PMID- 15287124 TI - Choosing eugenics: how far will nations go to eliminate a genetic disease? PMID- 15287125 TI - The misuse of race in medical diagnosis. PMID- 15287126 TI - When should a scholar's notes be confidential? An anthropologist involved in a medical lawsuit says she'll go to jail rather than turn hers over. PMID- 15287127 TI - Defining embryo death would permit important research. PMID- 15287128 TI - Senators grill NIH director over restrictions on stem-cell research. PMID- 15287129 TI - Transsexual 'subjects' complain about professor's research methods. PMID- 15287130 TI - Medical-research ethics under the microscope: schools try to plot the fine line between commercial links and conflicts of interest. PMID- 15287131 TI - Crossing the line? Medical research on brain-dead people raises ethical questions. PMID- 15287132 TI - California governor signs bills promoting stem-cell research. PMID- 15287133 TI - Advisory committee; Pediatric Advisory Committee; establishment. Final Rule. AB - The Food and Drug Administration (FDA) is announcing the establishment of a Pediatric Advisory Committee in the Office of the Commissioner. Elsewhere in this issue of the Federal Register, FDA is publishing a document requesting nominations for the membership on this committee. This document adds the Pediatric Advisory Committee to the agency's list of standing advisory committees in 21 CFR 14.100. PMID- 15287134 TI - Did a university let a sex researcher go too far? A psychology professor returned to his lab, only to draw complaints for a second time. PMID- 15287135 TI - NIH begins review of studies that were questioned at a congressional hearing. PMID- 15287136 TI - Lawmakers examine the possibility of favoritism in a $40-million NIH contract with Harvard. PMID- 15287137 TI - Bill to ban patents on human organisms nears congressional approval. PMID- 15287138 TI - Northwestern U. psychologist accused of having sex with research subject. PMID- 15287139 TI - When good institutions behave badly. PMID- 15287140 TI - Strong medicine for doctors: 'Just say no' to gifts from drug reps, a Columbia U. physician urges his colleagues. PMID- 15287141 TI - Korean investigators harvest first stem cells from a cloned human embryo. PMID- 15287142 TI - Ecstasy agonistes: a retracted study on a controversial substance raises questions about the reliability of government-sponsored research on drugs. PMID- 15287143 TI - Bush cuts 2 dissenters from federal bioethics advisory council. PMID- 15287144 TI - 2 scientists on bioethics council say its reports favor Bush ideology. PMID- 15287145 TI - Jury clears cancer center of failing to tell participants of clinical-trial risks. PMID- 15287146 TI - A dangerous surplus of sons? Two political scientists warn that Asia's lopsided sex ratios threaten world peace. PMID- 15287147 TI - Inconceivable? Deducting the costs of fertility treatment. AB - This Article considers whether infertile taxpayers can deduct their fertility treatment costs as medical expenses under Internal Revenue Code section 213 and whether they should be able to deduct them. Internal Revenue Code section 213 defines medical expenses as "amounts paid-for the diagnosis, cure, mitigation, treatment, or prevention of disease, or for the purpose of affecting any structure or function of the body." This definition is interpreted by reference to a baseline of normal biological functioning, which includes reproductive functioning. Most people conceive and bear children without having to incur expenses for fertility treatment. Expenses incurred to approximate the baseline of normal reproductive health are deductible, even if the taxpayer winds up better off, with a child, after the fertility treatment. The medical profession recognizes that infertility is a disease or condition. Infertility is a loss, just as a broken leg is a loss. Fertility treatment costs are thus medical expenses under section 213. In addition, given the existence of the medical expense deduction, taxpayers should be able to deduct the cost of fertility treatments, including IVF, egg donor, and surrogate procedures, under either an "ability-to-pay" or consequentialist normative approach. Reproduction is extremely important to most people. In addition, allowing taxpayers to deduct the costs of fertility treatment will encourage infertile taxpayers to elect the most effective treatment option and reduce the rate of risky multifetal pregnancies. This Article concludes that fertility treatment costs are deductible as medical expenses under current law and should be deductible as medical expenses. PMID- 15287148 TI - A new kind of bioethics: eschewing the academic mainstream, Bush panel focuses on technology's dangers. PMID- 15287149 TI - HIV and women: when words speak louder than actions. PMID- 15287150 TI - Eicosanomics: targeted lipidomics of eicosanoids in biological systems. AB - Recent advancements in mass spectrometry, especially the development of electrospray tandem mass spectrometry (ESI/LC/MS2) and matrix-assisted laser desorption time-of-flight mass spectrometry (MALDI/TOF), have greatly facilitated analysis of complex biomolecules. It has now become possible to profile, in relatively short periods of time, large multicomponent groups of compounds biosynthesized by biological systems. The efficiency and accuracy of analysis have led to the development of new concepts of mass spectrometric profiling, mapping, and imaging. Profiling of proteins in biological material (proteomics) has become a widely accepted strategy for identification of mechanisms involved in the biochemistry of disease processes, and has become a novel tool for unraveling new drug targets. Evolution of proteomics has relied on ESI/LC/MS2 and MALDI/TOF, techniques that are also useful in the novel area of quantitative proteomics. PMID- 15287151 TI - cagA+ Helicobacter pylori induces greater levels of prostaglandin E2 than cagA- strains. AB - cagA+ Helicobacter pylori (HP) infection is associated with an increased risk of distal gastric cancer. Previous studies investigating the effect of HP infection on prostaglandin E2 (PGE2) levels have not differentiated between cagA+ and cagA- strains and consequently have produced contradictory results. The aim was to investigate the effect of cagA+ strains on PGE2 and enhance the understanding of the mechanisms leading to gastric diseases. Hundred patients without peptic ulcers and not on medication were recruited (one later excluded) from endoscopy clinics: six biopsies were obtained from each patient. PGE2, colonization density and histology were determined. In addition, HP status was assessed by histology, CLOtest and culture with cagA+ being determined by PCR. Sixty-nine patients were HP- and 30 HP+ (10 cagA+, 18 cagA-, 2 undetermined). In age and sex-matched patients, PGE2 was significantly greater (P = 0.04) in HP+ (37.2 +/- 1.2 pg/mg per 20 min) than in HP- (22.6 +/- 1.2). In patients without atrophy, those infected with cagA+ had significantly higher (P = 0.03) PGE2 levels (53 +/- 1.1) than HP- patients (22.6 +/- 1.1) and greater levels (P = 0.29) than cagA- patients (35 +/- 1.3). In conclusion, the increased levels of PGE2 in the presence of cagA+ infection could be an important factor by which cagA+ strains enhance the gastric mucus layer protective functions leading to established colonization, gastritis and increased risk of gastric cancer. However, further evaluation with a large-scale multi-centre study is required to substantiate this hypothesis. PMID- 15287153 TI - Detrimental effects of prostaglandin F2alpha on preimplantation bovine embryos. AB - Two studies were performed to determine effects of prostaglandin F2alpha (PGF2alpha) on continued development of pre-compacted (in vitro-produced) and compacted (in vivo-derived) bovine embryos. In Experiment 1, pre-compacted embryos were placed in KSOM media supplemented with polyvinyl alcohol (0.3%) and assigned to the following treatments: (1) control; (2) PGF-1 (1 ng/mL PGF2alpha); (3) PGF-10 (10 ng/mL PGF2alpha); (4) PGF-100 (l00 ng/mL PGF2alpha); or (5) PGE-5 (5 ng/mL PGE2). Following 4 days of incubation in assigned treatments, continued development of pre-compacted embryos to blastocysts was reduced by addition of PGF2alpha in culture medium (P = 0.002). Development did not differ between control and PGE2 treatments (P > 0.10). In Experiment 2, compacted morula' s were placed in KSOM-PVA supplemented media and assigned to one of four treatments: (1) control; (2) PGF-0.1 (0.1 ng/mL PGF2alpha); (3) PGF-1 (1 ng/mL PGF2alpha); and (4) PGF-10 (10 ng/mL PGF2alpha). After 24h in culture, embryos were washed and placed in KSOM-BSA (0.5%) without PGF2alpha for an additional 48 h until assessment for development. Continued development of compacted morula to blastocyst was not affected by addition of PGF2alpha to the culture medium (P > 0.10). However, hatching rates of embryos cultured with PGF2alpha were lower (P = 0.05). In conclusion, it is suggested that PGF2alpha has a direct negative effect on continued embryonic development of pre-compacted and compacted bovine embryos. PMID- 15287152 TI - Thiazolidinediones increase arachidonic acid release and subsequent prostanoid production in a peroxisome proliferator-activated receptor gamma-independent manner. AB - Thiazolidinedione, peroxisome proliferator-activated receptor gamma (PPARgamma) agonist, has been used as an anti-diabetic drug and as an useful tool to elucidate multiple PPARgamma functions by in vitro and in vivo studies. We investigated the effects of thiazolidinediones on prostanoid production in lipopolysaccharide-stimulated cells. The high concentrations (>10 microM) of rosiglitazone and pioglitazone significantly increased lipopolysaccharide stimulated prostanoid production such as thromboxane A2 and prostaglandin E2. However, PPARgamma antagonist could not inhibit them. In PPARgamma-deficient cells, thiazolidinediones increased prostaglandin E2 production. Thiazolidinediones increased arachidonic acid (AA) release from the cell membrane by not stimulating AA releasing process involving several phospholipase A2s but inhibiting AA reuptaking process. The expression of cyclooxygenase-1 and cyclooxygenase-2 were not affected by thiazolidinediones. In this study, we demonstrated that high concentrations of TZDs increased AA release by the inhibition of AA reuptaking process, leading to subsequent increase in the prostanoid production in a PPARgamma-independent manner. This mechanism provides useful information for the elucidation of multiple PPARgamma functions and diabetic drug therapy. PMID- 15287154 TI - Alterations in embryo development in progestogen-supplemented cows administered prostaglandin F2alpha. AB - The objective was to examine effects of elevated prostaglandin F2alpha (PGF) on embryo development in cows supplemented with exogenous progestogen. Cows were artificially inseminated at estrus (Day 0) and a synthetic progestogen supplemented in the feed from Days 3 to 8. Cows were allotted randomly to receive either 15 mg PGF (TRT) or saline (CON) at 06:00, 14:00 and 22:00 h from Days 5 to 8. Blood samples were collected at 06:00 and 22:00 h from Days 5 to 8 for determination of progesterone and 13,14-dihydro-15-keto-PGF2alpha (PGFM). Single embryos were recovered on Day 8, assigned a quality score, and stage of development recorded. Progesterone was lower from Days 5 to 8 in TRT versus CON cows (P = 0.0001). Concentrations of PGFM from Days 5 to 8 were elevated in TRT compared to CON cows (P = 0.0001). Embryo quality was reduced in TRT cows compared to CON cows (P = 0.059). Percentage of embryos considered transferable was decreased by administration of PGF (P = 0.003). Sixty-four percent of TRT embryos were retarded in development at Day 8, whereas 80% of CON embryos had developed to expanded blastocysts (P = 0.003). In conclusion, treatment of progestogen-supplemented cows with PGF reduced quality and retarded development of embryos. Decreased fertility in conditions causing elevated concentrations of PGF may result from altered embryo development and quality. PMID- 15287155 TI - Exogenous prostaglandin E2 inhibits TPA induced matrix metalloproteinase-9 production in MCF-7 cells. AB - Elevated levels of prostaglandin E2 (PGE2) have been reported in many high metastatic human breast cancers, but no relationship between exogenous PGE2 activity, expression of matrix metalloproteinases (MMPs) and metastasis in human tumor cells has been reported. The poorly invasive human breast cancer cell line MCF-7 was cultured for 24h in the presence of both phorbol ester 12-O tetradecanoylphorbol-13-acetate (TPA, 50 nM) and PGE2 (1 microM) and the activity of MMP-9, one of the MMPs involved in metastasis, was measured, in growth medium by gelatin substrate zymography. TPA induced a strong production of MMP-9 while exogenous PGE2 had no effect on the basal MMP-9 level, but inhibited the TPA induced enzyme expression and matrigel invasiveness. We showed that MCF-7 cells expressed EP2, EP3 and EP4 receptors for PGE2 and that its action was probably mediated by EP4 receptor and adenylyl cyclase activation while cAMP dependent PKA was not involved in the process of inhibition of MMP-9 production. These findings suggest a possible inhibitory role for exogenous PGE2 in the metastatic process development. PMID- 15287156 TI - Do calcium-mediated cellular signalling pathways, prostaglandin E2 (PGE2), estrogen or progesterone receptor antagonists, or bacterial endotoxins affect bovine placental function in vitro? AB - The major objective of this experiment was to determine whether the bovine placenta could be stimulated to secrete progesterone, since the bovine placenta secretes little progesterone when the corpus luteum is functional. Secondly, we wanted to determine whether reported abortifacients or progesterone or estrogen receptor antagonists affected bovine placental prostaglandin secretion. The ovine placenta secretes half of the circulating progesterone at day 90 of pregnancy and PGE2 appears to regulate ovine placental progesterone secretion. Calcium has been reported to regulate placental progesterone secretion in cattle. Diced 186-245 day placental slice explants from six Brahman and six Angus cows were incubated in vitro at 39.5 degrees C under 95% air: 5% CO2 at pH 7.2 in 5 ml of M-199 for 1 h in the absence of treatments and for 4 and 8 h in the presence of treatments. Treatments were: vehicle; R24571; compound 48/80; IP3; PGE2; CaCl2; cyclosporin A; lipopolysaccharide (endotoxin) from Salmonella abortus equi., enteriditis, and typhimurium; monensin; ionomycin; arachidonic acid; mimosine; palmitic acid; progesterone, androstenedione; estradiol-17beta; A23187; RU-486; or MER-25. Jugular and uterine venous plasma and culture media were analyzed for progesterone, PGE2 and PGF2alpha by radioimmunoassay (RIA). Plasma hormone data were analyzed by a One-Way Analysis of Variance (ANOVA). Hormone data in culture media were analyzed for breed and treatment effects by a Factorial Design (2 breeds, 2-range of days, 21 treatments) for ANOVA (2 x 2 x 21). Since hormone data secreted by placental tissue in vitro did not differ (P > or = 0.05) by breed or range of days of pregnancy, data were pooled and analyzed by a One-Way ANOVA. Concentrations of PGE2 in uterine venous blood were two-fold greater (P < or = 0.05) in Angus than Brahman cows. PGE2 and PGF2alpha in vehicle controls increased from 4 to 8h (P < or = 0.05), but not progesterone (P > or = 0.05) Progesterone in culture media treated with RU-486 increased (P < or = 0.05) at 4 and 8 h compared to vehicle controls and was not affected by other treatments (P > or = 0.05). Concentrations of PGE2 in media at 4 and 8 h were lower (P < or = 0.05) when compared to controls except treatment with PGE2 at 4 and 8h and RU-486 at 8h (P > or = 0.05). PGF2alpha was increased (P < or = 0.05) by RU-486 at 8h and no other treatment affected PGF2alpha at 4 or 8 h (P < or = 0.05). In conclusion, modulators of cellular calcium signalling pathways given alone do not affect bovine placental progesterone secretion at the days studied and progesterone receptor-mediated events appear to suppress placental progesterone, PGF2alpha, and PGE2 secretion in cattle. In addition, PGE2 does not appear to regulate bovine placental progesterone secretion when the corpus luteum is functional and bacterial endotoxin does not appear to affect bovine placental secretion of PGF2alpha or PGE2. PMID- 15287157 TI - What can animal models tell us about human sexual response? AB - In all species, sexual behavior is directed by a complex interplay between steroid hormone actions in the brain that give rise to sexual arousability and experience with sexual reward that gives rise to expectations of competent sexual activity, including sexual arousal, desire, and performance. Sexual experience allows animals to form instrumental and Pavlovian associations that predict sexual outcome and thereby directs the strength of sexual responding. Although the study of animal sexual behavior by neuroendocrinologists has traditionally been concerned with mechanisms of copulatory responding, more recent use of conditioning and preference paradigms, and a focus on environmental circumstances and experience, has revealed behaviors and processes that resemble human sexual responses. In this paper, we review behavioral paradigms used with rodents and other species that are analogous or homologous to human sexual arousal, desire, reward, and inhibition. The extent to which these behavioral paradigms offer predictive validity and practicality as preclinical tools and models is discussed. Identification of common neurochemical and neuroanatomical substrates of sexual responding between animals and humans suggests that the evolution of sexual behavior has been highly conserved and indicates that animal models of human sexual response can be used successfully as preclinical tools. PMID- 15287158 TI - The menopause and sexual functioning: a review of the population-based studies. AB - Sexual problems are among the most frequently presented health concerns of women attending menopause clinics. We examine rigorous observational studies of the menopausal transition to determine whether there are changes in sexual functioning associated with the menopausal transition and the relative roles of aging and hormonal factors. We detail the methodological limitations of menopause research. We then review studies documenting the effects of aging on women's sexual functioning prior to reviewing studies that document both aging and menopausal status. These latter studies are divided into both cross-sectional and longitudinal studies. In summary, there is an age-related decline in sexual functioning but an added incremental decline associated with the menopausal transition. There have been relatively few studies that have been prospective, population-based, utilised a validated measure of sexual functioning, and carried out concurrent hormonal sampling. The Melbourne Women's Midlife Health Project is a prospective, observational study of a community-based sample of Australian born women aged 45-55 at baseline. There were eight annual assessments using a self report questionnaire based on the McCoy Female Sexuality Questionnaire and blood sampling for hormone levels. From early to late menopausal transition, the percentage of women with scores indicating sexual dysfunction rose from 42% to 88%. Decreasing scores correlated with decreasing estradiol but not with androgens. By the postmenopausal phase there was a significant decline in sexual arousal and interest, frequency of sexual activities, and the Total Score. There was a significant increase in vaginal dryness and dyspareunia and women's reports of their partner's problems in sexual performance. Women with low scores of sexual functioning were more likely to be distressed on the Female Sexual Distress Scale. In conclusion, there is a dramatic decline in female sexual functioning with the natural menopausal transition. PMID- 15287159 TI - The effect of hysterectomy on sexual functioning. AB - The effect of hysterectomy on sexual function is an issue of debate. There are reasons to believe that removal of the uterus can have adverse effects on female sexual functioning by disrupting the anatomical relations in the pelvis. In this article, we review the literature on the impact of hysterectomy (without oophorectomy and for benign conditions) on the sexual functioning of premenopausal women. There is evidence that women for whom there is a clinical indication for hysterectomy are often experiencing a decreased quality of life. After successful treatment of dysfunctional uterine bleeding, either by hysterectomy or uterus-saving alternatives, the majority of women report experiencing improved sexual functioning. Nonetheless, the research on the effect of hysterectomy on female sexual functioning is not conclusive. Prehysterectomy sexual functioning and psychosocial state are significant predictors for posthysterectomy sexual dysfunction and depression. A minority of women report developing sexual dysfunctions as a result of hysterectomy. The nature and extent of these dysfunctions have not been adequately investigated. Many investigations in this area are flawed by methodological imperfections. For example, qualitative changes in sexual functioning and changes in the physiology of sexual function often were not adequately addressed. In the future, researchers should include both objective measures of physiological functioning and use standardized and validated self-report questionnaires. A critical attitude towards the indications of hysterectomy remains mandatory. PMID- 15287160 TI - Beyond the male condom: the evolution of gender-specific HIV interventions for women. AB - As the number of HIV infections in women has increased, there has been a concomitant recognition that prevention efforts to reduce sexual transmission must address the gendered context in which risk behavior occurs. This paper provides a longitudinal perspective on the emergence of the HIV epidemic in U.S. women and the parallel development of interventions to reduce risk. In the first portion of this paper, we briefly discuss the growth of the epidemic among women and how public health responses reflected the early discourse about infected women. We also address methods of protection available to women, and the emerging recognition of the importance of gender relations. In the second half of this paper, we show how gender-specificity in prevention efforts has evolved, using a framework developed by Geeta Gupta (2001) and relying on published reviews of the intervention literature in the past 10 years. Finally, we discuss in detail several recent examples. We conclude with a discussion of future directions. PMID- 15287161 TI - What ever happened to ritualized homosexuality? Modern sexual subjects in Melanesia and elsewhere. AB - In this paper, I examine the legacy of ritualized homosexuality as a behavioral practice and as an analytic category of research in Melanesia since the early 1980s. A case study of striking change among the Gebusi of Papua New Guinea suggests that ritualized homosexuality and insemination of boys have become behaviorally vestigial or moribund and that characterizing sexual practices in these terms has been difficult to begin with (as the original proponent of these terms has himself suggested). Historical change in Melanesia reveals linkage between the contemporary construction of heterosexual norms and desires for locally modern development and progress. A larger issue is how researchers of sexuality may unwittingly accept Western ideologies of sexual choice and freedom while positing historical and non-Western practices as culturally bound rather than being open to individual exploration and interpersonal diversity. PMID- 15287162 TI - Group sex, sex change, and parasitic males: sexual strategies among the fishes and their neurobiological correlates. AB - Sexual selection, in the form of intrasexual competition and mate choice, has driven the evolution of a variety of sexual phenotypes amongst the vertebrates (Andersson, 1994). As a result, vertebrate species utilize many different approaches to acquire fertilizations. Humans and other primates show a wide range of sexual behaviors, but this range is dwarfed by the remarkable variation seen in advanced fishes. The goals of this review are (a) to acquaint the reader with the tremendous sexual diversity exhibited by fishes, (b) to demonstrate how this diversity provides unique opportunities to examine the neurobiological correlates of vertebrate sexual strategies, and (c) to highlight the parallels between the neuroendocrine correlates of the sexual strategies of fish with the mechanisms underlying sexual phenotypes in other vertebrates, showing the utility of fish studies for understanding sexual variation in general. PMID- 15287163 TI - Regression strategies for analyzing the study and pharmacological treatment of sexual response: ANOVA and beyond. AB - As the study of human sexual response becomes increasingly complex to meet the demands of this interdisciplinary field, the need for appropriate analytical tools has grown as well. In this review, we attempt to familiarize sexologists with the wide range of applications of regression analysis to the study of human sexual response. To this end, a typical experiment in human sexual response is described, and the basic regression model for the experiment is provided. Then, depending on the specific question under investigation, the nature of the study variables, and the design of the study, a number of different regression models are suggested, with each describing a specific variation from the basic regression model. Sample outputs are included to enable comparisons across several different models. With this information, researchers may develop a better understanding of the use and flexibility of the regression model in handling the kinds of problems often encountered in the experimental study of human sexual response. PMID- 15287164 TI - Remote sensing as a tool for mapping mosquito breeding habitats and associated health risk to assist control efforts and development plans: a case study in Wadi El Natroun, Egypt. AB - Limited mosquito ground surveys were combined with remote sensing and GIS technologies to identify mosquito breeding habitats in Natroun lakes area and to delineate associated health risks. Mosquito larval surveys were carried out in a small area to characterize positive breeding habitats and determine their geographic coordinates. Mosquitoes (Anopheles multicolor, Culex antennatus and Cx. theileri) were found breeding in water-flooded habitats with dense vegetation cover spatially associated to existing lakes. Chemical analysis indicated that mosquito breeding water was found to be polluted by several sources including agriculture, industrial and domestic sources. This information served as a training set to characterize the spectral signature of mosquitogenic (mosquito producing) habitats using reflectance data of the Thematic Mapper (TM) sensor aboard Landsat 5 satellite. Following characterization of the spectral signature, satellite data were used to predict, potential mosquito breeding patches over the whole study area. Field surveys were then carried out to assess the accuracy of predicted habitats and those surveys have indicated that all checked sites were positive for mosquito larvae demonstrating an accuracy of 100%. Based on an average adult mosquito flight range of 2 km, GIS was used to create buffer zones around breeding habitats describing areas at risk from mosquito nuisance and disease transmission. The obtained results could thus provide a new basis for directing the control of mosquito vectors as they provide health authorities with precise maps of mosquito breeding habitats in a timely manner. Moreover, the generated map delineating risk areas could be used by project developers to either re-site the project or invest in mosquito control activities in order to avoid health risks and ensure sustainability of their development. The approach adopted in this investigation demonstrated the practical and successful application of remote sensing and GIS in assisting health and development decision making. PMID- 15287165 TI - Non-histological assessment of liver fibrosis in HCV infection. AB - This study found a correlation between some serum markers [AST/ALT ratio, level of matrix metalloproteinase 9 (MMP9), level of viraemia and HCV serotype] and severity of liver fibrosis in HCV-infected patients. The study included 72 human cases referred to the Early Cancer Detection Unit, for liver biopsy assessment. The severity of liver fibrosis was staged using the METAVIR scoring system into 4 stages. The level of viraemia did not differ significantly in the different stages of liver fibrosis. Also, the type of HCV had no effect on the severity of liver fibrosis. However, the transaminases ratio differed significantly in the different fibrosis stages (P < 0.01). This serum test has a relatively high sensitivity and specificity (92.6% and 94.3%, respectively) in diagnosing severe fibrosis and cirrhosis. The level of MMP9 was, however, inversely correlated with the fibrosis stages and was found to have an 88.9% sensitivity and an 88.6% specificity when diagnosing severe fibrosis and cirrhosis. Although, the sensitivity of these serum markers did not reach 100%, yet their use can reduce the number of liver biopsies when diagnosing and treating HCV-infected patients. PMID- 15287166 TI - Investigation of induced biochemical and histopathological parameters of acetonitril extract of Jatropha carcus in albino rats. AB - Toxicological and histopathological investigations were carried on the acetonitril extract from J. carcus in comparison to praziquantel, the known antischistosomal drug. On a constant weight dose bases (single dose of 50 mg/Kg body weight injected orally to albino rats), the acetonitril extract from J. carcus showed mild toxicological parameters (AST p < 0.001, ALT & creatinine, non significant), biochemical parameters (total lipids, cholesterol, triglycerides, HDL, LDL, proteins, albumins globulins, ascorbic acid and bilirubin, non significant) and histo-pathological profile (slight alterations in liver, kidney and spleen), in comparison to control. However, these side-effects were very little as compared to the severe side-effects caused by Praziquantel. PMID- 15287167 TI - The protective activity of serum and fractionated serum from rats against Babesia divergens. AB - The rat adapted strain of bovine Babesia, B. divergens was used as a model to investigate the mechanisms of immunity to this parasite. The participation of humoral factors in acquired immunity to B. divergens was investigated in splenectomised rats. Antibodies to B. divergens were detected by IFA test in sera collected during infection and at different times after recovery. The protective activity of the same serum was tested in vivo by passive transfer and compared with the antibody levels as measured by IFA test. The role of antibody in protection was confirmed after fractionating IgM and IgG from immune and hyper immune sera. The protective activity of sera collected immediately after recovery was mainly due to IgM antibodies. The protective activity of sera collected 3-4 days after recovery and of hyper-immune sera was mainly due to IgG antibodies. In hyper-immune serum IgM antibodies were partially protective. PMID- 15287168 TI - Efficacy of Myrrh in the treatment of schistosomiasis (haematobium and mansoni) in Ezbet El-Bakly, Tamyia Center, El-Fayoum Governorate, Egypt. AB - Schistosomiasis remains a public health problem in Egypt, despite the continuous control effort. Most of the anti-schistosomal drugs have deleterious side effects or low efficacy. This necessitates a search for new safe and effective drug. A field survey was done in Ezbet El-Bakly (Tamyia Center) El-Fayoum Governorate to determine the magnitude of schistosomiasis haematobium and mansoni and to evaluate the efficacy of Mirazid (the oleo-resin extract from Myrrh of Commiphora molmol tree, family: Burseraceae) in the treatment of both types of schistosomiasis. Among the 1019 individuals parasitologically examined, the prevalence of S. haematobium and S. mansoni were 4.2% and 2.4% respectively and the geometric mean egg count (GMEC) were 33.2 eggs/10 ml urine and 113.3 eggs/gram stools. Most of the patients with haematobiasis and mansoniasis were <15 years (56.4% & 53.8%), males (56.4% & 53.8%) & illiterates (46.2% & 46.2%). All cases were treated by Myrrh (Mirazid) as two capsules (600 mg) on an empty stomach an hour before breakfast for six consecutive days and were followed up clinically and parasitologically by urine analysis by the sedimentation and nucleopore techniques and by hatching test and by stool analysis by sedimentation and Kato-Katz techniques and by hatching test. The parasitological cure rate after three months was 97.4% and 96.2% for S. haematobium and S. mansoni cases with the marvelous clinical cure without any side-effects. Patients not completely responded to a single course of treatment showed marked reduction of egg intensity. It is concluded that Mirazid proved to be safe and very effective in treatment of S. haematobium and S. mansoni infections under field conditions. PMID- 15287170 TI - Prevalence of dipterous flies associated with human and animal diseases in Al Obour and 6th October wholesale markets, Egypt. AB - The present study is a survey to identify the dipterous flies associated with human and animal diseases in the two wholesale markets: Al-Obour and 6th October. The results indicated that 20,824 flies belonging to 9 families, 24 genera and 31 species where trapped during the period of investigation (January-December, 2003). Musca domestica were the most abundant species in the two markets. Statistical analysis showed that species of the families: Calliphoridae, Milichiidae, Muscidae, Piophilidae, Otitidae, Sarcophagidae and Sphaeroceridae were significantly higher in Al-Obour than 6th October, due to spread of garbage and decaying fish. PMID- 15287169 TI - Molecular characterization of Cryptosporidium parvum isolates obtained from humans. AB - Fifty stool specimens collected from severe diarrheic patients attending Misr University Hospital, were examined microscopically for protozoan parasites mainly, Cryptosporidium parvum. Stool examination revealed 22 cases with C. parvum, 8 with E. histolytica, 14 with G. intestinalis and six were parasite free. The results were compared with the established nested PCR assay to detect DNA directly from stool specimens. After the extraction of DNA from stool, a 402 bp fragment of C. parvum DNA was amplified with two 26-mer outer primers. The amplified products, 194-bp DNA fragment, were used for a second run. This study indicated that the used primers are specific for DNA of C. parvum. The PCR detected a total of 28 positives; six of these cases were negative by AF stool examination, which eventually confirmed to be positive by several successive examinations of the stool and/or duodenal aspiration. Microscopy exhibited 78.5% sensitivity and 100% specificity compared to 100% specificity and sensitivity with PCR. Consequently, PCR is more sensitive and easier to interpret but required more hands-on time to perform and is more expensive than microscopy. However, PCR batch analysis reduces the cost considerably. PMID- 15287171 TI - Genotyping of human giardiasis in relation to anti-Giardia secretory IgA and mucosal histopathology. AB - Comparative study between the prevalence of pathological grading and Giardia genotypes revealed that, in patients infected with Giardia group I and II, out of patients having Giardia genotype I the prevalence of grade 0 was 13.16%, grade I was 21.05%, grade II was 47.37%, grade III was 13.16% and grade IV was 2.26% in comparison to 0%, 30.77%, 46.15%, 7.69% and 15.38% in genotype II (13 patients) and 10%, 40%, 20%, 20% and 10% in group III (10 patients) also in relation to 25%, 43.75%, 18.75%, 6.25% and 6.25% in mixed genotype infections group (16 patients) and 25%, 25%, 35.71%, 10.71% and 3.57% in undetermined infection group (28 patients) for grade 0, I, II, III & IV pathology respectively. There was no statistically significant difference regarding the prevalence of pathological grading in different Giardia genotypes in Gs I & II (P > 0.05). The mean OD of anti-Giardia secretory IgA in relation to Giardia genotypes in patients infected with Giardia Gs I & II was significantly different in the mean OD values of anti Giardia secretory IgA in patients with different Giardia genotypes which were 1.091 +/- 0.377, 1.079 +/- 0.474, 1.524 +/- 0.503, 1.292 +/- 0.472 & 1.004 +/- 0.31 groups of genotype I, II, III, mixed genotypes infection and undetermined infection group respecttively (P > 0.05), being more increased in patients infected with Giardia genotype III and in mixed genotype infection. PMID- 15287172 TI - DNA ploidy and S-phase fraction in the patients of chronic HCV and hepatocellular carcinomas. AB - Four hundred blue Fulgen-stained nuclei were measured from each lesion by using DNA image cytometry. The histopathological and cytopathological observations revealed that (52 cases, 69.3%) had a variable degrees of chronic hepatitis, (12cases 16 %) were emerging into cirrhosis, while (11 cases 14.7%) represented different grades of HCC. Most of the cases with minimal or mild chronic hepatitis were female, while most of male had moderate or severe chronic hepatitis, cirrhosis and HCC. DNA image analysis data gave the support of to the histological observations. All of chronic hepatitis C and cirrhotic cases showed normal diploid and/or tetraploid histograms, although increasing S-phase fraction s' values of the highly diseased chronic hepatitis and cirrhotic cases. Hepatocellular carcinomas and one cirrhotic case only revealed aneuploidy (diploid and tetraploid), while one case of poorly differentiated HCC revealed multiploid histogram. So, histopathological severity in cases of progressive chronic hepatitis seems to be associated with the age and sex of Egyptian society. Also, demonstrates the potential usefulness of image cytometry for the evaluation of the different histopathological problems. PMID- 15287173 TI - Bancroftian filariasis: spatial patterns, environmental correlates and landscape predictors of disease risk. AB - Lymphatic filariasis has been identified as the second leading cause of permanent and long-term disability. This article is an attempt to discuss the disease spatial context in light of current interest in GIS and satellite remote sensing. Field validation of outputs obtained through the application of these technologies in the Nile Delta, Egypt is also summarized. PMID- 15287174 TI - Human Monieziasis expansa: the first Egyptian parastic zoonosis. AB - This paper reports the first Egyptian zoonotic infection with the common tapeworm of sheep, Moniezia expansa. Perhaps this is the first human monieziasis expansa in the sheep raising countries worldwide. Diagnosis was based on recovery of the characteristic eggs from the stool of a fifteen-years-old shepherd. A single dose of Niclosamide one gram proved effective. PMID- 15287175 TI - Toxoplasmosis in pregnant women in Sanliurfa, Southeastern Anatolia City, Turkey. AB - Blood samples from 1149 pregnant women (age 26.9 +/- 5.3 years; mean 20 weeks gestation) on their first prenatal visit to Obstetrics and Gynecology Department and analyzed for Toxoplasma-specific immunoglobulin (IgM) and immunoglobulin (IgG) by ELISA. 694 (60.4%) of the women were IgG-positive, indicating previous maternal infection, while 35 (3%) were IgM-positive, indicating recent infection, and 420 (38.9%) were sero-negative. The high sero-positive rate may be linked to traditional raw meat consumption. Treatment with spiramycin was effective in preventing congenital toxoplasmosis in children of sero-positive mothers who consented to treatment. PMID- 15287176 TI - Induced changes in biochemical parameters of the molluscan tissues non-infected using two potent plants molluscicides. AB - Effect of Capparis spinosa (C. spinosa) and Acacia arabica (A. arabica) dry powder as plant molluscicide on some glycolytic and gluconeogenic enzymes on snail tissues, was investigated. Lactate debydrogenase (LDH), Pyruvate Kinase (PK), Hexokinase (HK), phosphofructokinase (PFK), glucose phosphate isomerase (GPI) as important glycolytic enzymes, were markedly manipulated by both plants when measured one day and one week post-treatment. On the other hand glucose-6 phosphatase (G-6Pase), fructose 1.6 diphosphatase (FDpase), phosphoenol pyruvate carboxykinase (PEPCK) as gluconeogenic enzymes were significantly affected by the moluscicidal plants. In addition, some other parameters as glycogen, glucose, total protein, 5-nucleotidase alpha-hydroxybutyrate dehydrogenase (HBDH) and succinate dehydrogenase (SDH) as kreb's cycle enzyme were tested. As conclusion, LC25 and LC50 concentrations of C. spinosa and A. arabica might render B. alexandrina physiologically unsuitable for S. mansoni infection. PMID- 15287177 TI - Avidity of immunoglobulin G antibody response to the different antigenic fractions of soluble Schistosoma mansoni adult worm antigen preparation (SWAP) using avidity immunoblotting assay. AB - The detection of IgG antibodies against the different SWAP antigenic fractions plus the determination of their avidity in avidity immunoblotting assay using 6 M urea wash, presents a novel alternative for the characterization of optimal antigenic markers for acute and chronic phases of Schistosoma mansoni infection and for vaccine development. Human serum samples from 25 acute schistosomiasis patients, 20 chronic cases and 15 normal healthy controls were analysed by IgG avidity enzyme linked immunosorbent assay (ELISA) and IgG avidity immunoblotting assay. Using avidity ELISA, a pronounced overlap of avidity index values was observed between acute and chronic infections with a range of uncertainty (0.86 1) which was encountered in both groups. Using avidity immunoblotting assay, antigenic bands at >116, 84, 48, 40 & 34 KDa were exclusive for the acute phase. From these bands, 34 KDa was recognized mostly by low-avidity antibodies and showed a high sensitivity (96%) and specificity (100%) making it an optimal marker for the acute phase. 40 KDa band was recognized mostly by high-avidity antibodies even during acute infection. Bands of 80, 70, 42, 36, 30 & 26 KDa were exclusive for the chronic phase. Only 70 KDa band was recognized by high-avidity antibodies yet, with low sensitivity (35%), that limits its use as an optimal marker for the chronic infection. Meanwhile, 70 and 40 KDa bands, recognized by high-avidity antibodies, are considered as potential vaccine antigen candidates. PMID- 15287178 TI - Evaluation of semi-quantitative PCR and IgG & IgM ELISA in diagnosis of toxoplasmosis in females with miscarriage. AB - One hundred female (age 20-42 yrs) patients were classified into group I: 40 patients presented with abortion in the first trimester; group II: 33 patients with abortion in the second trimester and group III: 27 patients with intrauterine fetal death (IUFD). The positive percentages of semi-quantitative PCR and both IgG & IgM ELISA were 38% and 35% respectively. Ten (26.3%) cases out of 38 were positive for toxoplasmosis by both PCR and ELISA-IgG, while 5 (13.2%) cases out of 38 were positive by both PCR and ELISA-IgM, whereas 16 (42.1%) cases out of 38 PCR positive cases were positive by both ELISA IgG & IgM. Sensitivity and specificity of both ELISA IgG and IgM were 81.57% & 93.54% respectively. False negative by ELISA were found in 7 cases out of 38 positive toxoplasmosis cases detected by semi-quantitative PCR. Three cases out of the 7 cases with false negative by ELISA were detected with a trophozoite copy load of 10(1) trophozoite /mL in the blood sample by semi-quantitative PCR. So, the semi quantitative PCR detected low levels of parasite DNA recommending its usefulness especially in the early stages of the disease when low amount of antibodies can't be detected by serological method or even by the conventional PCR. PMID- 15287179 TI - Echis carinatus (Serpentes: Viperidae) as a new host for Caryospora maxima (Apicomplexa: Eimeriidae) in Saudi Arabia. AB - Caryospora maxima is redescribed from the intestinal contents of the viperid snake, Echis carinatus collected from Gazan area in the southern region of Saudi Arabia. This report represents a new host and geographic location for the parasite. Sporulated oocysts of this coccidian are spherical to subspherical, 42.8 x 41.2 (40.3-45.9 x 39.6-43.8) microm, with smooth brownish-yellow bilayered wall, 1.9 (1.5-2.3) microm. thick. Micropyle and oocyst residuum are absent. Some oocysts had a small polar granule. Sporocysts are broadly ellipsoid, 22.1 x 16.8 (21.0-23.3 x 15.5-17.4) microm., with a prominent stieda and substieda bodies. Sporocyst residuum is present consisting of many granules in compact mass. Sporozoites are banana-shaped, each with two refractile globules. Oocyst measurements were similar to those reported from Psammophis schokari in Jordan. Except for the presence of a spherical polar granule, the oocyst morphology was identical to the original description of C. maxima. PMID- 15287180 TI - Host-parasite relationships between Schistosoma mansoni and Echinostoma liei and their intermediate host Biomphalaria alexandrina using RAPD-PCR analysis. AB - Biomphalaria alexandrina are known to be intermediate hosts for both Schistosoma mansoni and Echinostoma liei. RAPD-PCR assay offers a new approach to host parasite relationships. This was performed by investigating the genetic variation and compatibility among S. mansoni, E. liei and their intermediate host B. alexandrina with special emphasis on variations occurring in snails infected with S. mansoni and/or E. liei. Six primers were screened for DNA analysis and gave total patterns from 28 to 37 reproducible bands for each species. All specimens analyzed by the RAPD-PCR gave interpretable electrophoretic banding patterns that were polymorphic and compatible in the amplified products of these primers within each species. PMID- 15287181 TI - Detection of trichomoniasis in vaginal specimens by both conventional and modern molecular tools. AB - Out of 23 symptomatic cases 21 specimens were positive for T. vaginalis by one or more methods. 21 were positive by PCR (91.3%) and 17 (72.9%) by culture, 14 (60.8%) by Acridine orange (AO) stain and 13 (56.5%) by wet mount microscopy. The PCR detected all the positive cases and no PCR negative cases proved to be positive by any other technique. So, it is the test of choice for diagnosis of trichomoniasis. The modified Diamond's media proved to be highly sensitive (80.95%) and the results of the culture were significantly associated with those of PCR. The results of both wet mount and AO stain were insignificantly associated. The wet mount although widely used, easy, rapid and inexpensive yet, it has low sensitivity. So, negative cases should be repeated by either culture or PCR. PMID- 15287182 TI - Diagnostic potentials of copro-antigen detection based ELISA, compared to microscopy in intestinal amoebiasis. AB - Ninety three patients clinically presumed to have intestinal amoebiasis were chosen and stool samples were collected from all of them. Stool samples were subjected to microscopic examination and Entamoeba copro-antigens detection using Entamoeba and Entamoeba II tests. Out of 93 clinically positive samples, 51 (54.8%) were found positive by microscopy, while 49 (52.7%) were detected by Entamoeba test as having antigens specific for E. histolytica/E. dispar (88.24% sensitivity). Among 49 specimens, 4 were demonstrated as microscopy negative (90.48% specificity). Entamoeba II test demonstrated 16 specimens having antigens specific for the pathogenic E. histolytica among 49 positive by Entamoeba test, while 33 were detected as positive for nonpathogenic E. dispar copro-antigens. Copro-antigen assay using ELISA has shown to be more sensitive and specific than microscopy in different-tiation between pathogenic and nonpathogenic Entamoeba species. Extensive use of this technique allowed for revising the epidemiology of the true pathogenic E. histolytica and obviate the need for unnecessary chemotherapy with its costs and risk of side effects. PMID- 15287183 TI - Selective identification of the pathogenic E. histolytica in fresh stool samples using polymerase chain reaction (PCR). AB - Stool samples from 93 individuals clinically presumed to have intestinal amoebiasis and subjected to microscopic examination and DNA extraction. The PCR amplification was performed using two sets of primers that differentiate between pathogenic and nonpathogenic Entamoeba DNA. Of 93 clinically positive cases, 51 (54.8%) were positive by microscopy, while 53 (56.9%) were detected by PCR as having DNA specific for either E. histolytica / E. dispar. A specificity of 85.71% and a sensitivity of 92.15% were with PCR compared to microscopy. Among 53 PCR positive specimens, three different DNA sequences were demonstrated: 8 specimens had DNA sequences specific of E. histolytica, 31 with DNA specific for E. dispar and 14 specimens have mixed DNA sequences for E. histolytica and E. dispar. PCR is a sensitive and a specific tool. PCR application is better the epidemiology in endemic areas through keeping indefinite DNA records for prospective and retrospective studies. PMID- 15287184 TI - Potential diagnosis of Giardia lamblia infection through specific antibody detection in saliva. AB - Thirty six patients presenting with gastrointestinal symptoms were studied for the presence of specific anti-Giardia lamblia salivary IgA antibodies. Stool samples were examined for parasites especially G. lamblia by direct smear. Duodenal aspirate was examined for the parasite. Saliva samples were collected from each patient and examined by ELISA technique for the specific anti-Giardia lamblia IgA antibodies. 94.4% of positive cases for G. laimblia by stool analysis had positive anti-Giardia salivary IgA antibodies. 33.3% of stool negative cases were positive for anti-Giardia salivary IgA antibodies. All Giardia negative cases by duodenal aspirate examination were negative for anti-Giardia salivary IgA antibodies. Detection of antiGiardia salivary IgA antibodies was an excellent tool for screening G. lamblia in patients with long standing symptoms of more than one month duration. PMID- 15287185 TI - Fasciola gigantica: immunization of rabbits with proteins isolated from coproantigen. AB - Two fractions were isolated from coproantigen by ion-exchange chromatography in which DEAE cellulose was utilized. Both fractions and crude antigen were characterized by SDS-polyacrylamide gel electrophoresis which revealed 13 bands of molecular weight ranged from 205-31 in crude coproantigen. While fraction I resolved into six bands of molecular weight 198, 178, 148, 111, 101 & 45. Fraction II showed seven bands of 191 KDa, 178KDa, 166KDa, 118KDa, 98.5KDa, 72KDa & 32KDa. Fraction 1I was higher immunoreactivity than fraction by ELISA. Three immunoreactive bands of 191KDa, 118KDa & 98.5KDa were identified in fraction II using immunoblot assay. Five bands of 178KDa, 148KDa, 111KDa, 101KDa & 45KDa were detected in fraction I. Immunization of rabbits twice with fraction II in Freund's adjuvant with two weeks interval followed by challenge with F. gigantica metacercariae resulted in 66.6% protection from infection. The protection was assessed by detect-ion of hepatic damage, worm recoveries and antibody response. High level of IgG response in vaccinated rabbits than control infected ones occurred and being responsible for the recorded protection. PMID- 15287186 TI - Feeding ecology and food composition of the black carp Mylopharyngodon piceus and the grass carp Ctenopharyngodon idella inhabiting the fish pond of Al-Abbassa fish hatchery with emphasis given to vector snails. AB - Stomach contents of M. piceus and C. idella collected from earthen fish ponds at Al-Abbassa fish hatchery during different seasons were examined. Food items were analyzed by three methods; the numerical (N%) percentage of point assessment (P%) and frequency of occurrence (F%). A comparison of the different methods emphasized the importance of snails as the major food resources in the diet of M. piceus during all seasons. During winter, stomachs of the most fish samples collected from fish ponds were empty or had traces of food. The index of relative importance (I.R.I) indicated that snails were the major food category in the diet of M. piceus during summer, autumn and spring represented by 7331, 9696 and 11670 respectively. Plants were the main food item in the diet of C. idella during summer (4604), meanwhile they came 3rd in relative importance in the diet during autumn and spring. Artificial fish food came as the 2nd food item in the diet of M. piceus during summer (1398.9) and autumn (5896.3) but it completely absent in spring. It is considered the main food item in diet of C. idella during autumn (10384.3) and spring (6293.5) and being 3rd in summer (1395.5). The results showed that snails reached highest level in the diet of M. piceus during spring (about 80% of all diet, decreased during summer (about 70%) and reached lowest proportion in autumn (50%). Plants exhibited marked seasonal variation in abundance with maximum level during summer (about 41% of all diet decreased during spring (27%) and autumn (22%). Artificial fish food was the highest in the diet of M. piceus during autumn (39%), decreased in summer (22%) until disappeared during spring. It reached highest level in diet of C. idella during autumn (49%), decreased in spring (38%) and reached lowest proportion during summer (21 %). PMID- 15287187 TI - Influence of Capparis spinosa and Acacia arabica on certain biochemical haemolymph parameters of Biomphalaria alexandrina. AB - The work investigated the molluscicidal potency of dried Capparis spinosa and Acacia arabica leaves on selected biochemical parameters of Bionimphalaria alexandrina, in order to render them, physiologically, unsuitable for S. mansoni infection or at least disturb the life-cycle of the parasite within its respective snail host. The effect of the two plants on lactate dehydrogenase (LDH), 5'-nucleotidase, acid phosphatase (AP), aspartate and alanine aminotransferases (AST & ALT), alkaline phosphatase (ALP) and glucose content were studied. This work was extended to evaluate the effect of these two plants on protein profile as well as total protein (TP) content of snail's in haemolymph after 24 hours and one week of snails plants feeding. The study revealed that both plants induced marked alteration in all the measured parameters, where LC50 of C. spinosa after fed one week showed the most potent effect. PMID- 15287188 TI - Detection of genetic variabiltiy in nonhuman isolates of Fasciola hepatica and Fasciola gigantica by the RAPD-PCR technique. AB - The present study shows the molecular characterization of Fasciola gigantica and F. hepatica isolates collected from cows and sheep, using the random amplified polymorphic DNA fragments-polymerase chain reaction (RAPDs-PCR) technique. Optimal standardization of amplification conditions and thermocyclation were made, using genetic markers. The methodology used compared the genetic pattern between the two species (inter-species) and inside each species (intra-species) between cow and sheep and the amplification fragments were between 135 and 741 base pairs of marker. The results showed genetic variations (polymorphisms) of Fasciola gigantica and F. hepa-tica with amplification fragment based on a 500 400 base pair (bp). Inside each species, there were genetic variations in bovine and ovine and the amplification fragments were between 600 and 400 base pairs (bp). This assay is useful for both individual diagnosis and epidemiological surveys in endemic regions. PMID- 15287189 TI - Evaluation of seven assays detecting serum immunoglobulin classes and subclasses and salivary and faecal secretory IgA against Fasciola excretory/secretory (ES) antigens in diagnosing fascioliasis. AB - Seven assays detecting serum IgM, IgG, IgG1, IgG4, IgA and salivary and fecal excretory IgA against Fasciola excretory/secretory (ES) antigens were evaluated in diagnosing fascioliasis, for cross reactivity with Schistosoma mansoni sera and for evaluation of cure of Fasciola infection after treatment. Assays detecting sera IgM, IgG1, IgG4 and IgA against Fasciola ES antigens showed 100% specificity and sensitivity. Assays detecting IgM and IgG showed 98% and 96% sensitivity and 100% and 94.6% specificity respectively. Assays detecting salivary and faecal IgA showed 92% & 96% sensitivity and 100% & 100% specificity respectively. Assays detecting IgM and IgG4 were the best in evaluation of cure and assays detecting IgG4 & IgA showed the lowest cross-reactivity with sera from S. mansoni infected patients. So, assays detecting serum IgA, IgG1 & IgG4 against Fasciola ES antigens were highly sensitive and specific for diagnosis of fascioliasis and assays detecting salivary and faecal IgA were promising and of great help in diagnosis of fascioliasis especially in epidemiologic studies. PMID- 15287190 TI - Prevalence of asymptomatic bacteriuria among Egyptian elder persons living in the rural areas. AB - This study detected the prevalence of asymptomatic bacteriuria among elder Egyptians living in the rural areas Serrey village (Qaulyobia G.). Complete history taking, clinical examination, urine analysis and culture, blood glucose level and serum creatinine were performed for 86 elders. The asymptomatic bactiruria was 24.4% and the most common organism was Klebsiella followed by Staphylococcus and the least was E. coli. Regular urine culture is a must for the elders in rural areas. PMID- 15287191 TI - Myrrh (Commiphora molmol) in treatment of human and sheep dicrocoeliasis dendriticum in Saudi Arabia. AB - Dicrocoeliasis dendriticum is now imposing itself as an animal and zoonotic helminthic disease in many Arabian countries. Myrrh extract of Commiphora molmol (Mirazid) successfully and safely treated clinically and parasitologically proven 18 human dicrocoeliasis dendriticum patients. The dose was 2 capsules (300 mg each) given on an empty stomach an hour before the breakfast for six successive days. Cure (100%) was achieved clinically and by stool analysis for two months follow up. Besides, fifteen sheep naturally infected with Dicrocoelium dendriticum as proven parasitologically were successfully and safely treated with 2 capsules (300 mg each) on an empty stomach an hour before breakfast for four successive days. Cure (100%) was successfully achieved by stool analysis for seven days and macroscopically for detection of any adult worm after being slaughtered. The total dose required to treat infected sheep (2400 mg) was less than that required for human treatment (3600 mg). PMID- 15287192 TI - Role of cytochrome P4502E1 activation in proximal tubular cell injury induced by hydrogen peroxide. AB - BACKGROUND: There is now good evidence to suggest that cytochrome P450 (CYP450) may act as an iron-donating catalyst for the production of hydroxyl ion (OH*), which contributes to proximal tubular cell injury. However, it remains unclear which isoform of CYP450 is involved in this process. Cytochrome P4502E1 (CYP2E1) is a highly labile isoform which is not only involved in free radical generation, but has also been shown to be a source of iron in cisplatin-induced renal injury. This study investigates the role of CYP2E1 in the proximal tubular cell injury induced by hydrogen peroxide (H2O2). METHODS: Porcine proximal tubular cells (LLC PK1) were incubated with H2O2 (1 mM) for 4 h in the presence or absence of 0.1 mM of two CYP2E1 inhibitors; diallyl sulfide (DAS), or disulfiram (DSF), desferrioxamine (DFO) (0.1-0.4 mM), or catalase (CT) (78, 150, 300 U/mL). Cell death was determined by measuring LDH release. CYP2E1 activity was determined by p-nitrophenol hydroxylation after 2 h incubation with H2O2. RESULTS: Exposure of LLC-PKI to H2O2 significantly increased cell death. CT, DFO, DAS and DSF significantly reduced H2O2-mediated cell death. Incubation with H2O2 increased CYP2EI activation in time- and dose-dependent manner, which was significantly reduced by CT, DFO, DAS and DSF. CONCLUSION: We propose that CYP2E1 activation occurs possibly due to OH* and contributes to H2O2-mediated LLC-PK1 cell necrosis by acting as a source of iron and perpetuating the generation of OH* via the Fenton reaction. Inhibition of CYP2E1 may be a novel approach for the prevention of tubular injury caused by oxidative stress. PMID- 15287193 TI - Electrophysiological abnormalities in upper extremities after brachiocephalic A-V fistulas construction in predialysis patients. AB - BACKGROUND: Peripheral neuropathy is considered a common complication in patients suffering from advanced chronic kidney disease (CKD). Superimposed peripheral multiple neuropathies may complicate arteriovenous (A-V) fistulas construction. AIM: To evaluate, prospectively, the influence of brachiocephalic A-V fistulas construction on the peripheral nerves of the same extremity and to characterize the patients at risk for developing ischemic and neurological complications. PATIENTS AND METHODS: Twenty patients suffering from advanced CKD were enrolled in the study: 10 diabetic and 10 non-diabetic patients. All patients underwent electrophysiological evaluation one week before, 3 weeks and 3 months after surgery. Median, ulnar and radial nerves were studied. RESULTS: In non-diabetic patients MNCV was normal before and after surgery, but were significantly lower and reduced progressively and significantly after surgery in diabetic patients (p< or =0.02). In both non-diabetic and diabetic patients SNCV was reduced, but were significantly lower in diabetic patients before and after surgery (p< or =0.03). In diabetic patients it reduced progressively and significantly after surgery (p<0.01). Thirty percent of patients developed local edema and significant decrease of CMAP of median nerve three weeks after surgery (p=0.02) with complete resolution at three months. CONCLUSION: Diabetic uremic patients are at increased risk to develop disabling neurological complications after the construction of A-V fistulas. Diabetes was the only predictive risk factor for developing these complications. Prevention requires careful preoperative electrophysiological evaluation and postoperative follow-up. PMID- 15287194 TI - The influence of G-protein beta3-subunit gene and endothelial nitric oxide synthase gene in exon 7 polymorphisms on progression of autosomal dominant polycystic kidney disease. AB - BACKGROUND: A significant phenotypical variability is observed in autosomal dominant polycystic kidney disease (ADPKD). The variability cannot be fully explained by the genetic heterogeneity of the disease. We examined the influence of G-protein beta3-subunit C825T polymorphism and endothelial nitric oxide synthase Glu298Asp polymorphism on the progression of ADPKD towards end stage renal failure (ESRF). METHODS: 306 ADPKD patients (pts) were analyzed; 261 pts (136 males, 125 females) with ESRF, with subgroup of 73 pts (44 males, 29 females) with ESRF before 45 years (rapid progressors), 46 pts (20 males, 26 females) with ESRF later than in 63 years (slow progressors) and 45 ADPKD pts (17 males, 28 females) in mean age 51 years with serum creatinine under 110 micromol/L (slow progressors) and 100 genetically unrelated healthy Czech subjects. DNA samples from collected blood were genotyped for G-protein beta3 subunit C825T genotype in exon 10 and for endothelial nitric oxide synthase Glu298Asp genotype in exon 7. RESULTS: The G-protein beta3-subunit C825T genotype exhibited no significant differences among the groups of slow progressors (6.6% (6/91) TT, 54.9% (50/91) CT, 38.8% (35/91) CC), rapid progressors (9.6% (7/73) TT, 46.6% (34/73) CT, 43.8% (32/73) CC), ADPKD group with ESRF between 40-63 years (9.2% (13/142) TT, 50% (71/142) CT, 40.8% (58/142) CC) and control group (12% TT, 44% CT, 44% CC). When comparing the ages of ESRF of all patients with ESRF, we did not find significant differences in the ages: males TT--51.7+/-8.8 years, CT--51.9+/-10.3 years, CC--49.7+/-10.2 years and females TT--56+/-9.9 years, CT--53.2+/-8.5 years, CC--53.9+/-8.7 years. The endothelial nitric oxide synthase Glu298Asp and Asp29Asp genotypes were significantly more frequent in rapid progressors (9.6% (7/73) Asp/Asp, 39.7% (29/73) Asp/Glu, 50.7% (37/73) Glu/Glu) and in ADPKD group with ESRF between 40-63 years (11.3% (16/142) Asp/Asp, 41.5% (59/142) Asp/Glu, 47.2% (67/142) Glu/Glu) in comparison with slow progressors (8.8% (8/91) Asp/Asp, 24.2% (22/91) Asp/Glu, 67.0% (61/91) Glu/Glu) and with control group (8% Asp/Asp, 32% Asp/Glu, 60% Glu/Glu) (Chi-square test, p<0.05). Comparing the ages of ESRF of all patients with ESRF, we did not find significant differences in the ages in males with Asp/Asp--54.9+/-10.4 years, Asp/Glu--50.2+/-9.4 years, Glu/Glu--51.0+/-10.4 years. We found out in homozygous Asp/Asp females significantly earlier onset of ESRF (49.2+/-5.6 years) in comparison with heterozygous females (53.3+/-7.2 years) and with Glu/Glu homozygous females (54.8+/-9.7 years) (t-test, p<0.05). CONCLUSION: We excluded the significance of G-protein beta3-subunit C825T polymorphism on the progression of ADPKD. We established the negative prognostic value of the carriers of Asp variant of eNOS polymorphism. Finding of new modifiers could have in future clinical consequences for ADPKD patients. PMID- 15287195 TI - Is circulating phospholipase A2 removed by large-pore continuous venovenous hemodiafiltration in septic acute renal failure? AB - Group II A phospholipase A2 (PLA2) produces many inflammatory lipid mediators, and the elevation in the level during sepsis has been correlated positively with the decrease in the arterial blood pressure. We studied the effect of large-pore continuous venovenous hemodiafiltration (LP-CVVHDF) on the plasma PLA2 concentration and the clearance mechanism during septic acute renal failure. The subjects were 10 consecutive patients with septic acute renal failure receiving CVVHDF. Simultaneous samples of arterial, and filter inlet and outlet blood, and ultradiafiltrate were collected before starting CVVHDF (0 hr), and 4 hr, 12 hr and 24 hr after starting CVVHDF. PLA2 activity was measured in plasma and ultradiafiltrate. We eluted PLA2 bound to hemofilter from patient and the classification of PLA2 type of eluting solution and ultradiafiltrate was done using Western blot analysis. Plasma clearance (mL/min) was 28.1+/-7.6 at 4 hr, 23.2+/-8.9 at 12hr and 17.5+/-8.0 at 24 hr. Plasma clearance at 4 hr was higher than that at either 12 hr or 24 hr. Plasma clearance mainly consisted of adsorption by LP-CVVHDF. The changes in arterial plasma PLA2 activity were not statistically significant. One mg/mL of heparin eluted PLA2 bound to the large pore hemofilter. The PLA2 in eluting solution and in ultradiafiltrate were identified as an approximately 70 kD band in Western blot analysis using anti human secretory II A-PLA2 monoclonal antibody. The results show that circulating PLA2 can be removed by adsorption with LP-CVVHDF to some extent and that plasma PLA2 activity is not significantly decreased. Because PLA2 clearance with LP CVVHDF is estimated as <1% of total body PLA2 clearance, LP-CVVHDF could not be a clinically efficient therapy to remove the circulating PLA2. PMID- 15287196 TI - Salubrious effect of vitamin E supplementation on renal stone forming risk factors in urogenital tuberculosis patients. AB - The incidence of renal calculi has been evaluated to be 25% in urogenital tuberculosis patients. The stone could be caused due to the host, the pathogenic organism, or possibly by the treatment. Studies were carried out to find out the efficacy of vitamin E supplementation in reducing the risk of stone formation in renal tuberculosis patients. The study constituted four groups, Group I with 30 normal volunteers, the second group comprised of 36 renal tuberculosis patients (GuTb) a day before treatment. Third group comprised of 24 patients with regular anti tuberculosis drug regimen for sixty days. In the fourth group, 12 patients were treated with anti tuberculosis drug regimen along with supplementation of antioxidant vitamin E (200 mg/day) for sixty days. Hyperuricosuria and hypercalciuria were observed in group II and group III patients, along with increased excretion of oxalate and creatinine, accompanied by decreased excretion of inhibitors such as citrate and glycosaminoglycans (GAGs). Renal damage was evident with increased leakage of Lactate dehydrogenase (LDH), Alkaline phosphatase (ALP) and gamma-Glutamyl transferase (gamma-GT) in renal tuberculosis patients. From the results of the above study, it is obvious that increased urinary oxalate levels leads to cellular damage in GuTb patients, which is a prerequisite for crystal retention as revealed by the elevated urinary marker enzymes. Antioxidant therapy prevents membrane injury thereby reducing the risk of stone formation. Hence vitamin E supplementation has a salubrious effect in preventing stone forming tendency with routine anti tuberculosis drug regimen. PMID- 15287197 TI - Effects of altered volume loading on left ventricular hemodynamics and diastolic filling during hemodialysis. AB - BACKGROUND: Changes in the circulating volume associated with hemodialysis (HD) resulted in alternations of left ventricular (LV) filling. However, previous studies offered conflicting findings. This study thus evaluated the impact of HD on LV diastolic filling indices and hemodynamics. MATERIALS AND METHODS: Forty patients with end-stage renal disease were studied by Doppler echocardiography immediately before and after HD. The cardiac size, volume and mass were determined by M-mode and two-dimensional echocardiography. LV diastolic filling parameters and hemodynamics were assessed from mitral inflow using Doppler echocardiography. RESULTS: Left atrial and LV dimension, LV volume, and LV mass decreased significantly after HD (p<0.001). Cardiac output declined from 5.74+/ 1.37 to 4.98+/-1.27 L/min (p<0.001), whereas, the ejection fraction remained unchanged. HD elicited marked changes in the early diastolic E (95.1+/-20.5 to 70.3+/-18.2 cm/s, p<0.001) and late atrial filling A velocities (104.3+/-20.9 to 88.9+/-16.9 cm/s, p<0.001). In addition, correction of the deceleration time of E and isovolumic relaxation time prolonged significantly (p=0.011 and p<0.001, respectively). CONCLUSIONS: Findings in this study indicate that HD altering the loading condition significantly influenced the LV diastolic function and hemodynamics. Moreover, Doppler echocardiography provides an effective means of assessing the effects on LV diastolic filling and hemodynamics during HD. PMID- 15287198 TI - Electrolyte mass balance during CVVH: lactate vs. bicarbonate-buffered replacement fluids. AB - OBJECTIVE: To compare the effect of lactate vs. bicarbonate-buffered replacement fluids on electrolyte mass balance during isovolemic continuous veno-venous hemofiltration (CVVH). DESIGN: Randomized controlled study with double cross over. SETTING: Intensive care unit of a tertiary university hospital. PATIENTS AND PARTICIPANTS: Eight patients with acute renal failure (ARF). INTERVENTIONS: Isovolemic CVVH (2L/hr of replacement fluid) was performed in random order with either bicarbonate or lactate-buffered replacement fluid delivered pre-filter. MEASUREMENTS AND RESULTS: Sodium, potassium, chloride, magnesium, and phosphate, were measured in each sample. There was a mass gain of sodium, which was similar under both conditions (bicarbonate: 23.3+/-4.9 mmol/hr, lactate: 22.7+/-3.5 mmol/hr). Mass chloride gains occurred with bicarbonate-buffered replacement fluid only (12.8+/-5.3 mmol/hr), while there was an overall net loss of chloride with lactate fluids (-2.5+/-5.2 mmol/hr), resulting in a significant difference in chloride mass balance (p<0.0001). Magnesium mass balance was negative with bicarbonate buffer only (-0.6+/-0.2 mmol/hr) and also differed significantly from that obtained with lactate fluids (-0.1+/-0.2 mmol/hr, p<0.0001). Phosphate losses (bicarbonate: -1.7+/-0.7 mmol/hr, lactate: -1.7+/-0.5 mmol/hr) were equivalent with both buffers. Potassium mass balance was neutral. CONCLUSIONS: Mass balance during isovolemic CVVH is significantly affected by the type of replacement fluid administered prefilter. Isovolemic CVVH is not isonatremic and the use of bicarbonate-buffered fluid results in a significant accumulation of chloride and a loss of magnesium. PMID- 15287199 TI - Can a different priming process of the dialyzer affect dialysis adequacy in chronic hemodialysis patients? AB - In this study, we investigated whether a different priming process of the dialyzer could affect the dialysis adequacy in chronic hemodialysis (HD) patients. 20 HD patients (M/F:12/8) with a median age of 40 (20-74) were included in this study. All the patients were clinically stable and were on bicarbonate based hemodialysis program 3 times in a week. During the study period of 6 months, we tried to keep the vascular accesses, types and surfaces of the membranes and also the blood and dialysate flow rates almost the same for all patients. For the first 3 months of the study we performed our routine priming process by flushing 1 L of saline from the bloodline without any dialysate passing through the dialyzer. For the next 3 months, we carried out a different priming process. While we passed 1 L of saline through the blood compartment of the dialyzer, we also started the dialysate pump to get a flow rate of 500 mL/min for 30 minutes. After a 3 month period of different priming process, significant increases were observed in Kt/V (1.19+/-0.14 to 1.35+/-0.14, p=0.000), URR (%) (62.3+/-1.1 to 66.9+/-1.25, p=0.000) and nPCR (1.09+/-0.04 to 1.25+/-0.04, p=0.002) parameters. Our findings show that a priming process of the dialyzer by passing both saline and dialysate from the dialyzer for half an hour before starting every dialysis session can improve dialysis adequacy parameters. We suggest that this procedure, by increasing dialysis adequacy, can provide great clinical benefits. PMID- 15287200 TI - Bone mineral density in patients on maintenance hemodialysis and effect of chronic hepatitis C virus infection. AB - OBJECTIVE: To determine the prevalence of osteopenia and osteoporosis in HD patients at our center; to investigate whether HCV infection affects BMD in hemodialysis patients; to test for correlations between bone mineral density (BMD) and clinical and laboratory parameters in this population. SUBJECTS AND METHODS: The study involved 76 end-stage renal disease patients. Forty-three (56.6%) patients were tested negative for anti-HCV antibodies and HCV-RNA. Thirty three (43.4%) of them had positivity of anti-HCV antibodies and permanent or intermittent HCV-RNA positivity at least for two years. Mean HD duration was 86.4 months. Patients completed a standard questionnaire that listed age, sex, occupation, education level; cause of renal failure, smoking history, dialysis duration, and sports activities engaged in during life, and pathologic bone fractures. The women answered additional items about age at menarche, number of pregnancies and menopausal status. Each subject underwent a baseline physical examination, including measurement of body weight and height for calculation of body mass index. The results of laboratory tests that had been done at monthly visits in the previous year were retrospectively evaluated, and mean levels for the year were used for correlation testing. Bone mineral density was measured in the spine, femoral neck and forearm. Relationships between BMD values and chronic HCV infection, laboratory results and clinical parameters were analyzed. RESULTS: In the 43 patients who were negative for anti-HCV antibodies and HCV-RNA, spine BMD testing showed osteopenia in 16 (37.2%) cases and osteoporosis in 7 (16.3%) cases. The corresponding values for the neck of the femur were 14 (32.6%) and 6 (14.0%), and for the forearm were 19 (44.2%) and 15 (34.9%). In the 33 anti-HCV antibodies and HCV-RNA positive patients; spine BMD testing showed osteopenia in 10 (30.3%) cases and osteoporosis in 7 (21.2%) cases. The corresponding values for the neck of the femur were 17 (51.5%) and 4 (12.1%), and for the forearm were 4 (12.1%) and 25 (75.8%). Bone mineral density decreased as dialysis duration increased (p<0.05). There was no statistical difference between BMD measurements of chronic HCV infection positive and negative group. CONCLUSION: However the mean BMD values for all three sites in the 76 HD patients were low HCV infection may not be a risk factor for low BMD in this population. PMID- 15287201 TI - Hemorrhagic stroke in chronic dialysis patients. AB - AIMS: This study was designed to retrospectively investigate the clinical profiles, disease course and management of hemorrhagic stroke in chronic dialysis patients. We emphasized on the factors affecting the prognosis. PATIENTS AND METHODS: We retrospectively studied (January 1991-June 1999) the chronic dialysis patients who were admitted to our facility with a diagnosis of acute hemorrhagic stroke. The medical results were reviewed in detail and the clinical characteristics, laboratory data and management records of each individual were collected for analysis. RESULTS: There were 16 patients analyzed in total, 9 males and 7 females. The average age was 59.4+/-13.3 years old. Before admission, 14 patients received chronic hemodialysis (HD) and two patients peritoneal dialysis (PD). The co-morbidities included hypertension (16/16), Diabetes Mellitus (DM) (9/16), previous cerebrovascular accidents (9/16) and hyperlipidemia (5/16). The locations of cerebral hemorrhage (CH) were: the putamen (6/16), brain stem (3/16), thalamus (3/16) and others (4/16). Among the 14 HD patients, 8 remained on HD after onset of CH, while 6 switched to PD. Those who received PD before their development of CH continued to perform PD. The overall mortality was 44% (7/16). One of the 8 patients who continued on HD died (mortality 12.5%). Among the 8 patients who received PD, 6 died (mortality 75%). Two patients who underwent surgical intervention also passed away. The major cause of death was neurological deterioration. The interval between the onset of CH and death was short (15+/-13 days, range 2-39 days). CONCLUSION: The overall prognosis of CH in the chronic dialysis population is poor. Patients with lower hemoglobin levels upon presentation and those performing PD after CH may have even worse prognosis. PMID- 15287202 TI - Therapeutic approach of patients with IgA nephropathy. AB - Immunoglobulin A nephropathy (IgAN) is the most commonly encountered primary glomerulonephritis and it usually follows an indolent clinical course. However, hypertensive patients with proteinuria and renal insufficiency at presentation and patients with severe histological involvement are at high risk to develop end stage renal failure. There is no consensus for the treatment of patients with IgA nephropathy. In general, patients with normal renal function, mild proteinuria (3 g/24 h) and in progressive disease despite treatment with ACE inhibitors. Fish oil might be an alternative to corticosteroids in cases with renal insufficiency and chronic histological lesions. Combinations of corticosteroids and cytotoxic drugs are saved for patients with IgA nephropathy and a rapidly progressive course. PMID- 15287203 TI - The red blood cell deformability in patients suffering from end stage renal failure on hemodialysis or continuous ambulatory peritoneal dialysis. AB - Anemia is the main problem for patients suffering from end stage renal disease (ESRD). This study aimed to determine whether the index of rigidity (IR), that shows red blood cells (RBCs) deformability and the possible IR disturbances can provide an explanation about the cause of anemia, in patients undergoing maintenance hemodialysis (HD) or on peritoneal dialysis. The IR was determined in 39 hemodialyzed patients, who were already in dialysis for a period of time ranging from 16 to 120 months (mean+/-SD=41.8 +/-24.1) (Group A). Furthermore, the IR was measured in 32 patients on continuous ambulatory peritoneal dialysis (CAPD), who were in CAPD for a period of time ranging from 6 to 60 months (mean+/ SD = 10.7+/-9.9) (Group B). Finally, the IR was determined in 17 normal individuals (group C). The RBCs IR was measured twice in group A (before and after the end of a hemodialysis session) and once in groups B and C. The IR was determined by hemorrheometry (method of filtration), using special equipment. In group A the IR was increased in comparison to the control group (C) (17.9+/-6.2 vs. 10.2+/-1.8, p<0.0001). This increase was even higher in the measurement at the end of the hemodialysis session (paired t-test, p < 0.0001). The RBCs IR in CAPD patients was significantly lower than that of HD patients (12+/-3.8 vs. 17.9+/-6.2, p<0.0001) and was not statistically different from the control group (12+/-3.8 vs. 10.2+/-1.8, p=0.068). It is concluded from the study that: 1) in HD patients occur disturbances in the deformability of the RBCs, that are worsened by the hemodialysis session; 2) the index of rigidity of RBCs is significantly higher in the HD patients than in CAPD patients; 3) in patients on CAPD, the disturbance of deformability of the RBCs was less in comparison to the control group, which however does not reach the statistically significant levels. PMID- 15287204 TI - Other glomerular pathologies in three patients with diabetes mellitus. AB - Diabetic nephropathy is the common cause of end stage renal disease in diabetes mellitus. But other glomerular pathologies have been also described in diabetic patients. We described 3 cases with diabetes mellitus and other glomerular diseases. Case I: A 59-year-old male patient with type 2 diabetes mellitus for 4 years was evaluated for generalized edema. Physical examination showed pretibial edema and no diabetic retinopathy. The cause of nephrotic syndrome was membranoproliferative glomerulonephritis. Conservative therapy could not control the severe proteinuria and renal dysfunction. With corticosteroid and cyclophosphamide therapy partial remission was obtained. Case II: A 46-year-old diabetic woman was evaluated for severe proteinuria. Diabetic retinopathy was not found on her funduscopic examination. Mesangioproliferative glomerulonephritis was found on renal biopsy. Proteinuria did not regress with conservative therapy and corticosteroid and cyclophosphamide. Case III: A 48-year-old male patient with diabetes mellitus type 2 for 2 years was admitted to the hospital because of nephrotic syndrome and weakness. At another hospital his diagnosis with biopsy showed minimal change disease. He was treated with corticosteroid since 3 months. His renal biopsy was reevaluated and found amyloid deposition but not diabetic nephropathy or minimal change disease. In diabetic patients, nondiabetic nephropathy is not uncommon and it was reported as common as about 30%. In addition to therapy for diabetes mellitus these patients can need specific therapy. PMID- 15287205 TI - Ethambutol-induced optic neuritis in patients with end stage renal disease on hemodialysis: two case reports and literature review. AB - Ethambutol, a synthetic bacteriostatic agent, is a first line agent against Mycobacterium tuberculosis. Although optic neuritis is the most serious adverse effect of ethambutol, most cases in the literature are reversible. Renal failure prolongs the half-life of ethambutol and increases the risk of ethambutol-induced optic neuritis. We present two patients with end stage renal disease (ESRD), who were on maintenance dialysis and suffering ethambutol-induced optic neuritis. The first woman had been suffering ESRD on hemodialysis for 2 years. After tuberculosis was diagnosed, she was prescribed three-combined anti-tuberculosis medications, including ethambutol 800 mg/day. Bilateral blurred vision suddenly occurred 4 months after the start of treatment, and she became totally blind despite discontinuing ethambutol. The second woman had been on hemodialysis for 5 months. Tuberculosis was diagnosed by lung biopsy. After 3 weeks of three combined anti-tuberculosis medications including ethambutol (1,200 mg/day), reduced visual acuity and color vision defects occurred. One year after the discontinuation of ethambutol, visual acuity remained little improved. Physicians should be aware of ethambutol-induced optic neuritis and ethambutol should be used cautiously in patients with renal failure. PMID- 15287206 TI - Renal sodium handling study in an atypical case of Bartter's syndrome associated with mitochondriopathy and sensorineural blindness. AB - Bartter's syndrome is a disorder that has been linked to mutations in one of three ion transporter proteins: NKCC2 (type I), ROMK (type II) and CCLNKB (type III), which affects a final common pathway that participates in ion transport by thick ascending limb cells. We present an atypical case of mitochondriopathy combined with tubule functional disturbances compatible with Bartter's syndrome and definitive sensorineural blindness. Our patient had a peculiar clinical presentation with signs of salt and volume depletion, low blood pressure and secondary hyperaldosteronism, associated with hypokalemic metabolic alkalosis, hypocalcemia and severe hypomagnesemia, uncommon in genetic forms of Bartter's syndrome. The enhanced absolute and fractional sodium excretion in our patient compared to volunteers was accompanied by increased post-proximal sodium rejection, suggesting a striking ion transport dysfunction in these nephron segments. These findings lead to the Bartter's syndrome diagnosis, accompanied by a suppose mitochondrial tick ascending loop of Henle epithelium dysfunction that may reflect the high energy supplied by mitochondria electron transport chain, required for this nephron segment to maintain normal ion transport. PMID- 15287207 TI - Hemodynamic correction and early detection of tubulointerstitial fibrosis prevent disease progression in chronic kidney disease. PMID- 15287208 TI - More the same than different: worldwide drug policy issues. AB - Pharmaceutical policies in Europe were described in the previous issue of the journal. These policies were described in 5 areas: regulation and access; pricing, price control, and cost; publicly administered drug insurance; patient advocacy; and research ethics. While the underlying policy issues are strikingly similar, each country has developed its own set of solutions to address them. This article provides insight into the Canadian drug policy environment by comparing and contrasting it with the European experience. PMID- 15287209 TI - Comparative drug policies--heresy: learning from other countries. PMID- 15287210 TI - Differential pricing of pharmaceuticals in the internet age. AB - The Internet provides healthcare consumers with more information about available prices and provides pharmaceutical manufacturers with more information about consumers' willingness to pay. The former effect tends to undermine price differences while the latter tends to support them. We believe that the former effect will dominate and that the Internet will undermine differential pricing of pharmaceuticals. This should be a concern for manufacturers and policy makers, because differential pricing of pharmaceuticals can increase access for the poor and increase incentives for innovation. We suggest strategic responses for manufacturers and policy makers. PMID- 15287211 TI - Estimating emergency service treatment bed needs. AB - Estimating the required number of emergency service treatment beds must be sensitive to utilization patterns and strategic operational assumptions. This article describes key issues and illustrates techniques for the analysis of arrival and service times. Seasonal arrival patterns, time of day of arrivals, and common statistical distributions for length of stay are discussed. Alternative modeling approaches to estimate future bed needs are described, including visits/year per treatment space, simple queuing modeling, and detailed computer simulation. Sample estimates of treatment rooms needs are provided for typical arrival rates and lengths of stay. A generalized regression model based on the simulation trials is suggested for cases that fall outside of the illustrated simulation case studies. PMID- 15287212 TI - Universal design concepts in the emergency department. AB - Universal patient bedrooms have been a design focus in healthcare settings for over a decade. The challenges among designers and healthcare organizations include the definition of universal, the application of the concept to more than simply patient bedrooms, and importantly, the long-term efficacy of the concept. This article addresses each of those challenges, first, by offering a range of definitions and then by testing the application of those definitions within an emergency department and offering case studies indicating initial successes. PMID- 15287213 TI - Adapting to change: the advantage of modular clinic design. AB - Architects have applied modular design concepts for years. Projects benefiting most are those where a few room types are repeated often. Ambulatory care facilities are excellent candidates because this project type typically consists of a large number of rooms that can be standardized. The new Kaiser Permanente Santa Clara Medical Office Building illustrates the benefits and flexibility of modular design. PMID- 15287214 TI - Physician and staff turnover in community primary care practice. AB - The effect of a rapidly changing healthcare system on personnel turnover in community family practices has not been analyzed. We describe physician and staff turnover and examine its association with practice characteristics and patient outcomes. A cross-sectional evaluation of length of employment of 150 physicians and 762 staff in 77 community family practices in northeast Ohio was conducted. Research nurses collected data using practice genograms, key informant interviews, staff lists, practice environment checklists, medical record reviews, and patient questionnaires. The association of physician and staff turnover with practice characteristics, patient satisfaction, and preventive service data was tested. During a 2-year period, practices averaged a 53% turnover rate of staff. The mean length of duration of work at the current practice location was 9.1 years for physicians and 4.1 years for staff. Longevity varied by position, with a mean of 3.4 years for business employees, 4.0 years for clinical employees, and 7.8 years for office managers. Network-affiliated practices experienced higher turnover than did independent practices. Physician longevity was associated with a practice focus on managing chronic illness, keeping on schedule, and responding to insurers' requests. No association was found between turnover and patient satisfaction or preventive service delivery rates. Personnel turnover is pervasive in community primary care practices and is associated with employee role, practice network affiliation, and practice focus. The potentially disruptive effect of personnel turnover on practice functioning, finances, and longitudinal relationships with patients deserves further study despite the reassuring lack of association with patient satisfaction and preventive service delivery rates. PMID- 15287215 TI - Comparing the health status of VA and non-VA ambulatory patients: the veterans' health and medical outcomes studies. AB - The purpose of this study was to compare health status and disease profiles of ambulatory patients in specific Veterans Administration (VA) and civilian healthcare settings. A random sample of 2425 male veterans seeking care at 4 Boston-area VA outpatient clinics, who took part in the Veterans Health Study (VHS) in 1993-1995, were compared to 1318 male patients seeking civilian outpatient care in 3 major metropolitan areas covered in the Medical Outcomes Study (MOS) in 1986. The MOS sampled patients who had 1 of 5 conditions- hypertension, noninsulin-dependent diabetes, recent myocardial infarction, congestive heart failure, or depression. These 2 samples were age adjusted and compared in terms of the SF-36 Health Status/Quality of Life measures, and a list of 100 clinical variables (diagnostic, symptom, and medical event reports) collected with comparable instruments by a trained clinical observer. Individual odds ratios (VHS to MOS) were calculated for each measure and clinical variables. SF-36 measures of patient health in the VHS were lower than those in the MOS by more than one half of a standard deviation (SD) on 4 of 8 scales, by more than one quarter of a SD on the other 4, by 58% of a SD on the physical health summary scale, and by 37% of a SD on the mental health summary scale (P < .0001 in all cases). The median odds ratio was 2.2 among the SF-36 scales and 1.9 among clinical variables. Outpatients in the 4 VA clinics had more than twice the illness burden than did patients in the MOS. Current economic condition and service-connected disability explain most, if not all, of the differences. The differences were clinically and socially meaningful and would be consistent with substantially higher expected healthcare use. PMID- 15287216 TI - Improving the response choices on the veterans SF-36 health survey role functioning scales: results from the Veterans Health Study. AB - Role functioning and its limitations due to one's health is an important aspect of health-related quality of life (HRQoL). The Medical Outcomes Study (MOS) SF-36 includes 2 role functioning scales: role limitations due to physical health problems (RP) or emotional problems (RE). Although they capture important concepts of HRQoL, these 2 scales have some limitations in their measurement properties. Using dichotomized sets of response choices, the scales are limited in their distributional properties (eg, higher standard deviation than other SF 36 scales) and ability to discriminate between clinically relevant groups. In this study, we ascertain the improvements to these 2 scales using 5-point ordinal response choices for each of the scale items. Two thousand one hundred sixty-two patients from the Veterans Health Study (VHS), an observational study of health outcomes in patients receiving ambulatory care, completed a health status questionnaire and a medical history. The health questionnaire included (1) the MOS SF-36, in which the RP and RE items used dichotomized yes/no responses; and (2) a set of modified RP and RE items that used 5-response choices for each of the items, ranging from "no, none of the time" to "yes, all of the time." We compared the original and modified RP and RE scales using internal consistency reliability and factor analysis. We tested item convergent and discriminant validity using multitrait scaling, and scale discriminant validity using ordinary least squares regression. Results indicate that the modifications to the original RP and RE scales accomplish important gains in the distributional properties of the scales. The floor and ceiling effects of the 2 scales have been reduced and the reliability of the RP scale has increased (0.87-0.95). Factor analysis and multitrait scaling tests indicate that the modified items have the same interpretation as the original items. Tests of discriminant validity indicate that the modified RP and RE scales have greater explanatory power for measures of disease burden, depression, and disease severity. The modified SF-36 role scales are clearly superior to the original versions. The modifications have increased the explained variability, suggesting greater explanatory power and more information obtained by the role functioning measures. The modified RP and RE are capturing a wider spectrum of disease severity, in part due to the lowering of the floor and raising of the ceiling of the scales. Additional work needs to test these improvements in other populations and to expand the analysis to track the responsiveness of the modified scales to clinically and socially important changes over time. PMID- 15287217 TI - Comorbidity assessments based on patient report: results from the Veterans Health Study. AB - The objective of the study was to develop a self-reported measure of patients' comorbid illnesses that could be readily administered in ambulatory care settings and that would improve assessment of their health-related quality of life and utilization of health services. Data were analyzed from the Veterans Health Study, an observational study of health outcomes in patients receiving Veterans Administration (VA) ambulatory care. Patients who received ambulatory care services in 4 VA outpatient clinics in the greater Boston area between August 1993 and March 1996 were eligible for inclusion. Among the 4137 patients recruited, 2425 participated in the Veterans Health Study, representing a response rate of 59%. Participants were mailed a health-related quality of life questionnaire, the Medical Outcomes Study Short Form Health Survey (SF-36). They were also scheduled for an in-person interview at which time they completed a medical history questionnaire. We developed a comorbidity index (CI) that included 30 self-reported medical conditions (physical CI) and 6 self-reported mental conditions (mental CI). The physical CI and the mental CI were significantly associated with all SF-36 scales and explained 24% and 36%, respectively, of the variance in the physical component summary and the mental component summary of the SF-36. Both indexes were also significant predictors of future outpatient visits and mortality. The CI is an independent predictor of health status, outpatient visits, and mortality. Its use appears to be a practical approach to case-mix adjustment to account for differences in comorbid illnesses in observational studies of the quality of healthcare. It can be administered to large patient populations at relatively low cost. This method may be particularly valuable for clinicians and researchers interested in population based studies, case-mix adjustment, and clinical trials. PMID- 15287218 TI - Juvenile injustice. PMID- 15287219 TI - VA pursues bar-code quality. PMID- 15287220 TI - Enforcement outdoes education at eliminating unsafe abbreviations. PMID- 15287221 TI - Poison control centers' basic funding should be federal, IOM says. PMID- 15287222 TI - Developers aim to bring laboratory tests to the point of care. PMID- 15287223 TI - Nonadministration of injectable antineoplastic drugs in a French hospital. PMID- 15287224 TI - Availability of pralidoxime and implications of inadequate stocking. PMID- 15287225 TI - Experience with nesiritide at a community hospital. PMID- 15287226 TI - Gauging patients' interest in an arthritis Web site. PMID- 15287227 TI - Pharmacists in health-assessment program. PMID- 15287228 TI - Pharmacist-managed pain clinic at a Veterans Affairs Medical Center. PMID- 15287229 TI - New JCAHO standards: challenges and opportunities. PMID- 15287230 TI - New JCAHO medication management standards for 2004. PMID- 15287231 TI - JCAHO's Shared Visions-New Pathways: the new hospital survey and accreditation process for 2004. PMID- 15287232 TI - Atazanavir: a new protease inhibitor to treat HIV infection. AB - PURPOSE: The pharmacology, pharmacokinetics, and clinical trials of and drug interactions and formulary considerations associated with atazanavir, the newest protease inhibitor (PI) to treat human immunodeficiency virus (HIV) infection, are evaluated. SUMMARY: Clinical and pharmacokinetic trials were identified through a MEDLINE search. In addition, all available literature citations and meeting abstracts were obtained from the drug's manufacturer. All articles identified from the data sources were evaluated, and all information deemed relevant was included in this review. Data on atazanavir for the treatment of HIV infection are limited to several phase II and III trials, one of which is still ongoing. Atazanavir has shown efficacy comparable with other PIs and the nonnucleoside reverse-transcriptase inhibitor efavirenz in reducing HIV RNA levels, increasing CD4+ lymphocyte counts, and increasing the percentage of patients achieving clinically undetectable HIV RNA levels when given as the sole PI in treatment-naive patients, in combination with saquinavir in treatment experienced patients, and with ritonavir-boosting regimens in highly treatment experienced patients. Treatment-naive patients receiving atazanavir commonly develop a protease enzyme mutation on codon 50, which decreases HIV's susceptibility to atazanavir but may increase the susceptibility of the virus to other PIs. When atazanavir is given to patients with preexisting PI-related mutations, the virus's susceptibility to atazanavir is greatly reduced. The occurrence of lipid abnormalities, which has been a major concern with previous PIs, has not been shown to be troublesome in patients receiving atazanavir. CONCLUSION: Atazanavir may be used alone as a first-line PI, with saquinavir in treatment-experienced patients, or in combination with a ritonavir-boosting regimen in highly treatment-experienced patients as part of a salvage regimen. PMID- 15287233 TI - Restricting patients' medication supply to one month: saving or wasting money? AB - PURPOSE: A state Medicaid program's pharmacy expenditures associated with dispensing one- and three-month supplies of drugs were examined. METHODS: We simulated the effect of a policy change from a maximum of a 100-day supply of prescription medication to one where only a 34-day supply was allowed. All North Carolina prescription claims from Medicaid enrollees who filled a prescription for at least one of six medication categories during fiscal years 1999 and 2000 were included. The six categories were angiotensin-converting-enzyme inhibitors, antiulcers, antipsychotics, nonsteroidal antiinflammatory drugs, selective serotonin-reuptake inhibitors, and sulfonylureas. The dollar value of the medication wasted, the amount of medication wastage diverted after a change to a shorter prescription length, and the total costs incurred by the increases in prescription refills were calculated. RESULTS: For each therapeutic category, 255,000-783,000 prescription drug claims were analyzed. No valid drug claims were excluded for any reason. Although 5-14% of total drug wastage, attributed to switches of drug therapy, could be saved by dispensing a 34-day supply, this saving could not make up for a larger increase in dispensing costs, as consumers would fill prescriptions more often. In addition, reducing the amount of drug dispensed each time may be costly to consumers through increased transportation and other expenses. CONCLUSION: Simulated calculation showed that the cost of drug therapy to North Carolina's Medicaid program would probably increase if 34 day rather than 100-day supplies of medications are dispensed to patients. PMID- 15287234 TI - Reported medication errors associated with methotrexate. AB - PURPOSE: Medication errors reported to FDA as adverse events in which methotrexate was identified as a possible contributor were studied. METHODS: All adverse-event reports submitted to FDA between November 1997 and December 2001 indicating potential medication errors involving methotrexate were analyzed to determine the indication for use, the type of error, and the point in the medication-use process where the error occurred. RESULTS: A total of 106 cases of reported medication errors associated with methotrexate were identified, including errors resuiting in 25 deaths (24%) and 48 other serious outcomes (45%). The most common types of errors involved confusion about the once-weekly dosage schedule (30%) and other dosage errors (22%). The most frequently involved indication for use was rheumatoid arthritis (42%). Of the errors, 39 (37%) were attributable to the prescriber, 21 (20%) to the patient, 20 (19%) to dispensing, and 18 (17%) to administration by a health care professional. CONCLUSION: A review of medication errors involving methotrexate revealed that errors occurred during all phases of use, often resulted from confusion about dosage, and often caused death or other serious adverse effects. PMID- 15287235 TI - Effects of a pharmacist-initiated educational intervention on patient knowledge about the appropriate use of antibiotics. PMID- 15287236 TI - Benefits of forming pharmacy technician teams. PMID- 15287238 TI - Effect on amphotericin B lipid complex use of a clinical decision support system for computerized prescriber order entry. PMID- 15287237 TI - Therapeutic interchange involving replacement of rofecoxib or celecoxib with valdecoxib. PMID- 15287239 TI - "Her body dissected. PMID- 15287240 TI - Victimization: a newly recognized outcome of prematurity. AB - Victimization by peers affects 10 to 20% of school children under the age of 12 years. Physical, verbal, and psychological victimization (being pushed, hit, called names, teased, being the target of rumours, theft, extortion) is associated with short- and long-term adjustment problems, such as peer rejection, social withdrawal, low self-esteem, anxiety, loneliness, and depression, as well as academic problems and school drop-out. Research on populations of school children (primary and secondary) has associated victimization with personal risk factors (the victim's characteristics and behaviour) and interpersonal risk factors (social relationships between peers). Studies on the social adjustment of preterm children at school age show that, even in the absence of a major motor or cognitive disability, this population has several personal risk factors associated with victimization. The objective of this study was to compare the level of victimization experienced by a group of 96 seven-year-old children born extremely preterm (EP, < 29 weeks of gestation; 49 females) against that experienced by a group of 63 term children (34 females) matched for age and sex, maternal level of education, and family socioeconomic status. The children born EP had a mean gestational age of 27.3 weeks (SD 1.2) and a mean birthweight of 1001.1g (SD 223) and normal birthweight children had a mean gestational age of 39.5 weeks (SD 1.5) and a mean birthweight of 3468.7g (SD 431). Physical and verbal victimization were assessed in a school setting by peers with individual sociometric interviews (Modified Peer Nomination Inventory). After controlling for physical growth (height and weight) at the age of 7 years, the data indicate two independent effects: males were more victimized than females, and children born preterm experienced more verbal victimization by their peers than their term classmates, even when participants with a visible motor, intellectual, or sensory disability were excluded. Several hypotheses are presented to account for the higher incidence of verbal victimization of preterm children. PMID- 15287241 TI - Long-term prognosis of cerebral venous sinus thrombosis in childhood. AB - Cerebral venous sinus thrombosis (CVST) is a rare but potentially serious disorder in children. There is no literature on the long-term neuropsychological and emotional sequelae and implications for quality of life. We studied 17 children who had CVST after the neonatal period, aged between 1 month and 16 years at the time of CVST (mean age at CVST was 6 years, median 4 years 8 months). Five children died during follow-up. The cause of death was related to CVST in one child. Twelve children participated in a clinical follow-up assessment. Mean follow-up was 2 years 8 months. One child had physical sequelae with impairment of skilled movement. All children had average or high intelligence scores. Two children with CVST due to an uncomplicated mastoiditis had mild cognitive deficits: one child had difficulty with written language; the other had diminished cognitive efficiency with concentration and attention problems associated with decreased psychosocial functioning. Decreased physical well-being was reported in three of 12 children. We conclude that children who had survived CVST had a fair prognosis. Most had normal cognitive and physical development, although mild cognitive deficits or decreased physical and psychosocial well-being can occur. PMID- 15287242 TI - Is exposure to cocaine or cigarette smoke during pregnancy associated with infant visual abnormalities? AB - The aim of this study was to assess the association between cocaine or cigarette smoke exposure in utero and visual outcome. A total of 153 healthy infants (89 males, 64 females; gestational age range 34 to 42 weeks) were prospectively enrolled in a masked, race-matched study. Quantitative analyses of urine and meconium were used to document exposure to cigarette smoke and cocaine. Infants with exposure to other illicit drugs, excepting marijuana, were excluded. At 6 weeks of age, grating acuity and visual system abnormalities (VSA; eyelid oedema, gaze abnormalities, and visual inattention) of 96 infants from the original study sample were assessed with the Teller acuity card procedure and a detailed neurological examination. Neither cocaine nor cigarette smoke exposure was associated with acuity or VSA. However, VSAs were associated with abnormal neurological examination, independent of drug exposure and other risk factors (odds ratio 7.9; 95% confidence interval 2.0 to 31.5;p=0.004). This unexpected finding could prove a helpful clinical marker for the infant at risk for neurological abnormalities. PMID- 15287243 TI - Vernier acuity is selectively affected in infants and children with cortical visual impairment. AB - Cortical visual impairment (CVI) refers to bilateral impairment of vision that is usually due to damage occurring perinatally in the visual cortex and/or optic radiations. The most common cause of this damage is hypoxia, and other causes include encephalitis, meningitis, and trauma. Relatively little research has been done to quantify visual abilities in children with CVI. In the present study, we used an electrophysiological technique (visual evoked potentials) to measure two aspects of spatial vision in 35 infants and children with CVI (15 females, 20 males; mean age 3 years 6 months, SD 3 years 5 months; age range 4 months to 16 years). We measured each child's grating acuity (resolution for detecting high contrast stripe patterns) and vernier acuity (resolution for localizing pattern elements). Performance on grating acuity and vernier acuity in individuals with CVI was compared with that of age-matched individuals with normal vision, and it was found that vernier acuity was relatively lower than grating acuity in children with CVI. Results support the theory that vernier acuity is cortically mediated, and suggest that vernier acuity is a more sensitive measure than grating acuity for quantifying vision deficits in patients with CVI. PMID- 15287244 TI - Development of displacement of centre of mass during independent walking in children. AB - The aims of this study were to assess the characteristics of three-dimensional displacement of the centre of mass of the body (CMb) during walking in healthy children and to compare it with those of young adults. Twenty-one children (11 males, 10 females; age range 1 to 9 years) were recruited from the nursery and school attached to the Universite catholique de Louvain, Brussels, Belgium; and three young adults (one male two females; mean age 26 years 4 months) were recruited from the Rehabilitation and Physical Medicine Unit of the same university. Displacement of CMb was assessed at different walking velocities in the children and adults by two successive mathematical integrations of ground reaction forces, measured by a large strain-gauge force platform. Displacement of CMb was controlled for leg length of the participant to eliminate the scaling effect that is dependent on growth. Results showed that vertical and lateral amplitudes of the CMb when controlled for leg length were greater for children before 4 years of age and that the forward amplitude when controlled for leg length was greater for children before 7 years of age. We conclude that the development of mature human CMb displacement during independent walking is a gradual neural process, evolving until the age of 7 years. PMID- 15287245 TI - Effectiveness of selective muscle-release surgery for children with cerebral palsy: longitudinal and stratified analysis. AB - The purpose of this study was to determine the effectiveness of muscle-release surgery for children with cerebral palsy (CP) using longitudinal and stratified analysis. Twenty-five children with CP (15 females, 10 males; age range 4 to 16 years; mean age 8 years 2 months, SD 3 years 2 months) were selected from five treatment centres in Japan. Twenty-two children had spastic diplegia, two had spastic quadriplegia, and one had athetospastic quadriplegia. Motor function for each child was assessed using the Gross Motor Function Measure (GMFM). Assessment was conducted on eight separate occasions: 1 month and 1 week before surgery, and 1, 2, 4, 6, 9, and 12 months after surgery. Participants' motor function before surgery was classified using the Gross Motor Function Classification System (GMFCS). Six children were classified at level I, three at level II, six at level III, and 10 at level IV. A significant difference was found after surgery in the GMFCS levels III and IV groups (p<0.05). Improvement in GMFM scores between 1 week before surgery and 12 months after surgery were 1, 5, 8.5, and 8.5 for GMFCS levels I to IV respectively. Results indicate that this treatment is advantageous for improving motor function in children within GMFCS levels III and IV. PMID- 15287246 TI - Care provider assessment of intrathecal baclofen in children. AB - Intrathecal baclofen is used increasingly to manage severe spasticity in children. Before implanting the baclofen pump, care providers typically ask how it will benefit their child. The purpose of this study was to describe the perceptions of care providers about intrathecal baclofen for management of spasticity in 80 individuals (52 males, 28 females). The individuals were younger than 22 years at the time of implantation (mean age 11 years, SD 5 years; range 3 to 21 years). Participants had been implanted with the pump for a minimum of one year at the time of evaluation. The most common diagnoses were quadriplegic and diplegic cerebral palsy and traumatic brain injury. Most participants were at level IV and V on the Gross Motor Function Classification System. After pump implantation most participants had tone reduction on the Ashworth scale of 1 to 1.9 in the lower extremities and 0 to 0.9 in the upper extremities. Lower extremity range of motion was maintained in 43 of 51 individuals (84%) and lost in 8 participants (16%). Complications requiring surgery occurred in 63 of a larger group of 152 patients (incidence per patient-year of follow-up was 0.19). Thirty-one of the 80 children had orthopedic procedures after pump placement. Only one of these was unexpected and none had rapid progression of scoliosis. Most treatment goals were achieved. Goals most commonly chosen (decreased pain, prevention of worsening of deformity, and improved ease of care) were improved in 91%, 91%, and 88% of participants respectively. Ninety-five per cent of care providers agreed that they would have this procedure performed again (81% strongly agreed, 14% slightly agreed). All care providers reported improvement in scores on the Caregiver Questionnaire. This information has been helpful to families considering intrathecal baclofen therapy. PMID- 15287247 TI - Social skills of children with neurofibromatosis type 1. AB - Children with neurofibromatosis type 1 (NF1) have difficulties in forming friendships and are often rejected by their peers. Factors that contribute to these negative social outcomes are poorly understood. This study investigated the social skills of children with NF1 and the influence of comorbid conditions, such as attention-deficit-hyperactivity disorder (ADHD) and specific learning dfficulties. We assessed and analyzed data from 79 children with NF1 (42 males, 37 females; mean age 11 years 6 months, SD 2 years 4 months) and 46 unaffected siblings (19 males, 27 females; mean age 12 years 1 month, SD 2 years 6 months; age range 8 to 16 years). Social skills were measured with the Social Skills Rating System. Children with NF1 had significantly poorer social outcomes than their unaffected siblings, and significantly poorer social skills in comparison with normative data. The presence of ADHD (in 39% of children with NF1) was the major risk factor for poor social functioning. Children with NF1 and ADHD had the poorest social skills and social outcomes when compared with children with NF1 only or children with NF1 and learning difficulties. These findings dispel the previous assumption that NF1 alone is associated with poor social functioning and has major implications for the development of effective interventions. PMID- 15287248 TI - Neurodevelopmental pattern of succinic semialdehyde dehydrogenase deficiency (gamma-hydroxybutyric aciduria). AB - Succinic semialdehyde dehydrogenase (SSADH deficiency) (MIM 271980) is a defect in gamma-aminobutyric acid catabolism, resulting in the accumulation of gamma hydroxybutyric acid (GHB) and causing neurological and cognitive disorders of varying severity. The non-specific nature and the difficulties in detection of urinary GHB explain why this disorder is largely underdiagnosed. Of 350 patients identified worldwide, to date only six adults with SSADH deficiency have been reported in the literature. Here we describe two additional cases in brothers up to ages 26 and 28 years. This retrospective report sheds light on the clinical features of SSADH deficiency in relation to the physiopathological involvement of GHB, and tries to identify the specific neurodevelopmental pattern of this learning disability.* Features of this are: early impaired psychomotor development with hypotonia and disturbances in motor coordination; impaired development of language, mainly due to poor auditory perception; and seizures and psychotic features in late adolescence or adulthood. Moreover, narcolepsy-like symptoms could be a consistent feature of the disease. PMID- 15287249 TI - Is treatment with growth hormone effective in children with cerebral palsy? AB - Children with cerebral palsy (CP) often have poor linear growth during childhood, resulting in a diminished final adult height. Here we report a female with CP and short stature but without growth hormone (GH) deficiency who exhibited increased growth during treatment with GH. We also report two other children with CP who were treated with GH: one female with a history of leukemia, and a male with Klinefelter syndrome. These two children were both found to be GH-deficient by insulin provocative GH testing and responded to treatment with increased growth rate. Growth improved to a greater extent in the two children with apparent GH deficiency. In summary, it is felt that GH therapy might be beneficial for children with CP and warrants further investigation. PMID- 15287250 TI - The Human Tissue Bill: the death of pathology? PMID- 15287251 TI - Energy cost of walking in children with cerebral palsy: relation to the gross motor function classification system. PMID- 15287252 TI - 'Congenital peripheral neuropathy presenting as apnoea and respiratory insufficiency: spinal muscular atrophy with respiratory distress type 1 (SMARD1)'. PMID- 15287253 TI - [Biological characteristics and ecological factors determining mammalian radioresistance in the Rodentia order]. AB - Both the results of our investigations and published data on mammalian radioresistance, biological characteristics and ecological factors which modify it are generalized in this paper. The described diversity among large taxa in radioresistance is based on the data obtained on the rodents from the Old and the New Worlds (totally 51 samples from 22 genera of four families--Sciurudae, Muridae, Cricetidae, Heteromyidae). It was found that radioresistance substantially depended on which family the species belonged. The main biological characteristics and ecological factors (body size, nutrition type, biotopical restriction) were revealed; in sum they determined 40% of radioresistance of small mammals. PMID- 15287254 TI - [Polysystemic assessment of the state of sanogenesis in workers employed in nuclear fuel plants. The analysis of metabolism regulation processes]. AB - Using the method of laser correlation spectroscopy of biological fluids (blood serum, urine, oropharyngeal washout fluid) we studied the types of metabolic shifts in workers employed in nuclear fuel complex plant. In was found that the incidence of catabolic shifts considerably increased in workers with higher level of occupational exposure. In individuals contacting with open radiation sources we found the contribution of anabolic immunomodifying shifts with predominance of autoimmune sensibilization. A risk group for blood diseases was identified. PMID- 15287255 TI - [Monitoring of tundra vole (Microtus oeconomus Pall.) population inhabiting areas with increased radiation background]. AB - The data of complex examinations of morphophysiological state of Tundra vole (Microtus oeconomus Pall.), the lipid peroxidation (LPO) processes and the energy exchange in their organs, the cytogenetic analysis of marrow cells and the process of multiplication in populations inhabiting areas with increased natural radiation background more 50 years are generalized. The data obtained support the existence of qualitative differences of Tundra vole subpopulations inhabiting control and radioactive nuclide contaminating areas. They allow to suppose that the process of adaptation to chronic low intensity radiation effect of Tundra vole in natural conditions involves the transition of the LPO and another regulatory systems to different level of function and is directed on population survival and homeostasis maintenance in changing radioecological conditions. PMID- 15287256 TI - [Polysystemic assessment of the state of sanogenesis in workers employed in nuclear fuel plants. The analysis of functional state of cardiovascular, respiratory, and psychomotor systems]. AB - The state of the cardiovascular, respiratory, and psychomotor systems of workers employed in nuclear fuel plant was evaluated using a computer-assisted laboratory complex. It was found that neuroendocrine regulation of the peripheral circulation and muscular activity responsible for fine motions are most vulnerable to industrial hazardous factors. PMID- 15287257 TI - [The cytogenetic damage, radiosensitivity and adaptive response in children living in the different districts of Moscow]. AB - The spontaneous level of blood lymphocytes with micronuclei (MN), the sensitivity to 1.0 Gy irradiation and adaptive response (AR) after adaptive irradiation with a dose of 0.05 Gy 5 hr later have been studied in children population living in different districts of Moscow. It was shown that spontaneous frequency of cells with MN, the sensitivity to 1.0 Gy acute irradiation and the AR manifestation have significant differences in samples taken from children living in different districts. The individual variability is significant also. In each group of children the individuals with the enhanced radiosensitivity after adaptive irradiation have been observed. In conformance with the data of radioecological inspection the radiation situation in different Moscow districts is quite safe on overage but in some districts the spontaneous level of lymphocytes with MN, and radiosensitivity after 0.05 Gy irradiation were enhanced, the AR was not found. PMID- 15287258 TI - [Radioecological environment in the territory of Yakutia]. AB - The present radioecological environment in the territory of Yakutia is analyzed. The general description of the natural gamma-background and the sources of pollution of permafrost landscaped with natural and artificial radionuclides is represented. It was revealed that the radiation environment was not good in some regions of Yakutia. PMID- 15287259 TI - [A new approach to assessment of biological consequences of exposure to low-dose radiation]. AB - A high sensitivity of characteristics of the lipid metabolism in erythrocytes to exposure to low doses of gamma- and X-rays was found; the changes in lipid peroxidation (LPO) of blood components were persistent, which substantially influenced the development of biological consequences of low doses of radiation. The effect of low doses of radiation on the interrelation between the LPO intensity in blood plasma and the lymphocyte DNA structural integrity or the cell free DNA content in animal blood plasma was found. Different sensitivity of the examined parameters (neither normalization nor linear dependence on a dose rate was found) implies the transition of the LPO regulatory system to another level of functioning due to scale changes of interrelations between the examined parameters under influence of low doses of radiation. These findings make possible to assess biological consequences of radiation factor for animal groups by the scale changes of interrelations between the examined characteristics in blood plasma. PMID- 15287260 TI - [Molecular epidemiology of the late radiation effects]. AB - The scientific bases, problems, methods and prospects for the development of a new scientific direction "Molecular epidemiology of late consequences of ionizing radiation influence on the man" are reviewed. It is marked, that for two decades on the basis of achievements in the field of molecular biology, biochemistry, genetics and genomics it has arisen and is developing the important direction in infectious and non-infectious epidemiology--molecular epidemiology. Molecular epidemiology is a new section of epidemiology on border with molecular biology and genetics, integrating efforts of epidemiologists, molecular biologists, genetics and researchers from many other areas of knowledge in a direction of an estimation of individual risk of development of chronic diseases at practically healthy people, development of recommendations on their preventive maintenance in cohorts of risk and, hence, in all population by studying by molecular methods of an etiology of illnesses, mechanisms of a susceptibility to them, and also biological mechanisms and frequency of their development in human cohorts. Special development in molecular epidemiology was noticed for the methods named biomarkers, subdivided on 4 categories: 1) markers of an internal dose; 2) markers of an effective dose; 3) markers of preclinical biological effects; 4) markers of a susceptibility. Use of biomarkers in molecular-epidemiological researches appeared especially productive in oncology. As radiogenic cancers are the main remote consequence of exposure to ionizing radiations, approaches and achievements of molecular epidemiology of nonradiation cancers are perspective at becoming of molecular epidemiology of radiation effects, with an object to solve problems concerning mechanisms and features of radiation carcinogenesis, risk assessment, distribution and preventive maintenance of radiogenic cancers in cohorts of professionals of the various branches having contact to sources of ionizing radiation, and the population, exposed to radiation medically. PMID- 15287261 TI - [Role of NO-mediated mechanisms in post-irradiation changes of neurohormonal regulation of the heart function]. AB - Investigations on isolated Langendorff-perfused rat heart have shown that after the impact of ionizing radiation in 1 Gy dose the myocardial contractility and relaxation are decreased. The inhibition of NO synthesis attenuates chrono- and inotropic responses of the heart and reaction of coronary vessels to norepinephrine (NE), increases negative inotropic and vasoconstrictor effects of carbamylcholine (CCh). In the irradiated animals the reaction of isolated heart to NE and CCh is decreased. The post-irradiation changes in adreno- and cholinoreactivity caused not only by changes in receptor sensitivity and density, but also by impairment of NO-mediated regulation of coronary flow and biomechanical heart function. PMID- 15287262 TI - [Hormonal regulation of the function of the myocyte intracellular membranes under radiation influence]. AB - It was shown that the experimental hypofunction of thyroid, induced by thyroidectomy and strong gamma-irradiation with a dose of 1 Gy lead to disturbance of function and structure of membranes of sarcoplasmic reticulum of myocites in rats. Introduction in vivo of L-thyroxine rose functional capacity of membranes with insignificaut changes in their lipid bilayer. PMID- 15287263 TI - [The radioadaptation condition at various stages of transition process, induced by gamma-irradiation in bacteria Escherichia coli JM 101]. AB - The dynamics of transition process caused by gamma-irradiation in bacterial Escherichia coli JM 101 culture was investigated. The bacteria radiostability at different phases of this process was defined. The received results testified to phase gamma-irradiation doze dependent character of bacteria population reaction. It was shown, that the increase of bacteria cell radioresistance occured at the postradiation restoration stage, whereas at the inhibition stage the radioresistance decreased. PMID- 15287264 TI - [Genotoxic studies in populations occupationally exposed to low doses of physical and chemical agents in Republic Croatia]. AB - In Croata Republic personnel occupationally exposed to radiation is examined with a method of chromosomal aberrations every 5 years, by the law. A population exposed to chemical factors is examined with a method of sister chromatid exchange as well as with micronucleus test. In the paper it is discussed a significance of various methods of examination of low doses of clastogenic and aneugenic factors on human genome. PMID- 15287265 TI - [Radioprotective capacity of indralin in reducing radiation injury to salivary glands]. AB - Radioprotective capacity of radioprotector indraline (alpha-1(B)-adrenoagonist) was studied by its effect on early displays of a local radiation injury to salivary glands in white rats after X-ray irradiation of an animal head with a dose of 18.7 Gy. Indraline was found to be capable to reduce a radiation injury to parotid glands registered by reduction of gland mass on 6th day after irradiation. In experiments on rats, radioprotective efficiency of indraline (100 mg/kg) in term of DRF is close to 1.5. PMID- 15287266 TI - [Regularities of changes in 137Cs content in milk in the long term after the Chernobyl nuclear reactor accident]. AB - Regularities of changes in 137Cs content in cattle milk in the long term after the Chernobyl accident have been analyzed. Contamination levels of haylands and pastures, soil properties, specific features of agricultural production and time after the fallout play a crucial role in 137Cs concentration changes in animal products. Trends have been studied that reflect the influence of these factors and their significance assessed. The half-life periods of 137Cs decay in milk vary over the period of 1994 to 2000 between 7.1 and 14.8 years and approach similar periods calculated for the long term after global radioactive fallout from nuclear weapons tests. PMID- 15287267 TI - [Accumulation of 90Sr in the bone tissue of the northern mole vole, living in the head part of the East Ural Radioactive Trace]. AB - Deposition of 90Sr in a bone tissue of the northern mole-vole (Ellobius talpinus Pallas, 1770) living in the head part of the East Ural Radioactive Trace (EURT) (density of pollution by 90Sr 37 MBq/m2 - 1000 Ci/km2) was investigated. Features of the rodent are digging (underground) way of life, family organization of settlements and weak migratory activity. Authentic (p < 0.01) interfamily distinctions in concentration 90Sr are revealed on the background of absence of sexual and are features of accumulation the radionuclides inside separate families, thus individual parameters differ in 7 times. It is established, that the level of accumilation of 90Sr is comparable with that in other species of small mammals in this territory. Age inversion of accumulation of 90Sr earlier found out in other species of mammals on the EURT is confirmed. PMID- 15287268 TI - [Modelling of radiocaesium soil-plant transfer]. AB - Existing methods of predicting of radiocaesium transfer from soil to plants was critcally reviewed. The analysis of radiocaesium behavior in the system "soil solid phase/pore solution/plant" was carried out. Equations for calculation of radiocaesium uptake by plants as a function of soil properties were obtained and tested using the reported experimental data. Key soil parameters which natural variability and estimation difficulty are the main sources of prediction uncertainty were identified. PMID- 15287269 TI - [Radioactive contamination of aquatic organisms of the Yenisei River in the area affected by the activity of the Mining-and-Chemical Combine]. AB - The study was done to investigate the content of manmade radionuclides in aquatic organisms of the Yenisei River near the Mining-and-Chemical Combine (MCC) and to estimate the exposure dose rates to organisms from various sources. The results of the investigation and calculations suggest that the main source of radioactive contamination of aquatic organisms is the coolant of the third MCC reactor, which is still being released into the Yenisei. Gamma-spectrometric analysis revealed 23 manmade radionuclides in the biomass of aquatic plants. The aquatic animal Phylolimnogammarus viridis and diatoms also contain manmade radionuclides. Among aquatic organisms, the highest dose rate is received by aquatic plants (up to 39 microGy/day). For most aquatic organisms under study, the dose received from the technogenic irradiation is an order of magnitude higher than the dose received from natural irradiation. The water moss (Fontinalis antipyretica) features the highest capacity to accumulate manmade radionuclides; hence, it accumulates the largest technogenic exposure dose among the study aquatic organisms. PMID- 15287271 TI - Three year health outcomes among older women at risk of elder abuse: women's health Australia. AB - BACKGROUND: Older women are at increasing risk of various forms of familial violence, yet detection is poor and very little is known of the long-term health effects of this psychosocial problem. The effectiveness of the 'Vulnerability to Abuse' Screening Scale (VASS) in predicting three year health outcomes was investigated among women enrolled in the Australian Longitudinal Study on Women's Health, now known as Women's Health Australia. METHODS: The sample comprised a cohort of 10,421 women aged 73-78 who completed the 1996 and 1999 postal surveys (attrition rate 19.5%). The Time 2 sample had a small bias towards lower risk for elder abuse at Time 1 and better health on SF-36 and self-rated health. The VASS is a 12-item self-report measure with 4 factors: vulnerability, coercion, dependence and dejection. RESULTS: Overall, physical health (PCS) declined while mental health (MCS) increased over the three year period. Decline in physical health was predicted by only the dejection factor, but not by factors which seem to more directly measure abuse. The predictive validity of the VASS for three year mental health outcomes was given partial support. Three of the four VASS factors (dejection, vulnerability, and coercion) predicted decline in mental health at the univariate level, however, after adjusting for confounders, only one VASS factor (dejection) independently predicted decline in mental health. CONCLUSIONS: While the VASS shows some promise as a marker of health risk in older women, only the dejection factor proved consistently predictive of declining health status. Further research is needed to determine longer term predictive validity of the scale and to gain a clearer picture of how abusive experiences impact on older women's health. PMID- 15287270 TI - Outcomes research in cancer clinical trial cooperative groups: the RTOG model. AB - BACKGROUND: The Radiation Therapy Oncology Group (RTOG), a National Cancer Institute sponsored cancer clinical trials research cooperative, has recently formed an Outcomes Committee to assess a comprehensive array of clinical trial endpoints and factors impacting the net effect of therapy. METHODS: To study outcomes in a consistent, comprehensive and coordinated manner, the RTOG Outcomes Committee developed a model to assess clinical, humanistic, and economic outcomes important in clinical trials. RESULTS: This paper reviews how the RTOG incorporates outcomes research into cancer clinical trials, and demonstrates utilization of the RTOG Outcomes Model to test hypotheses related to non-small cell lung cancer (NSCLC). In this example, the clinical component of the model indicates that the addition of chemotherapy to radiotherapy (RT) improves survival but increases the risk of toxicity. The humanistic component indicates that esophagitis is the symptom impacting quality of life the greatest and may outweigh the benefits in elderly (> or =70 years) patients. The economic component of the model indicates that accounting for quality-adjusted survival, concurrent chemoRT for the treatment of NSCLC is within the range of economically acceptable recommendations. CONCLUSION: The RTOG Outcomes Model guides a comprehensive program of research that systematically measures a triad of endpoints considered important to clinical trials research. PMID- 15287272 TI - Functional ability, social support, and depression in rheumatoid arthritis. AB - OBJECTIVE: First, to investigate the patterns of functional ability, depressive feelings, and social support in early stage rheumatoid arthritis (RA) patients. Second, to demonstrate the stress buffering effect of social support. Social support is thought to reduce the impact of chronic stress on psychological well being; for patients without social support the impact of functional ability on depressive feelings will be stronger. METHODS: In 4 waves with an intervening period of 1 year, longitudinal data was collected of 264 Dutch RA patients, of which 65% was female. At T1, the mean age of these patients was 53 years, while their mean disease duration was 22 months. In an interview at the patients' homes, data was collected on functional ability, social support en psychological well-being. The buffering effect of social support was examined by testing the significance of the (computed) stressor by social support interaction term in a regression analysis on depressive feelings. RESULTS: Although large differences between subjects existed, the mean scores on functional ability, social support, and depressive feelings barely changed from year to year. Patients who deteriorated in functional ability during one year had the best chances to improve next year, and visa versa. Furthermore, the stress by support interaction terms had no significant effect on depressive feelings in a regression analysis. CONCLUSIONS: This study demonstrated clearly the fluctuating pattern of RA in the first years after onset. The patients' level of depressive feelings was linearly related to the level of functional ability. Like many other studies, also this study could not provide evidence for the stress buffering effect of social support. PMID- 15287273 TI - Quality of life in old people with and without cancer. AB - The aim was to investigate the influence of age and gender on quality of life (QoL), complaints, and the presence and nature of self-reported diseases in persons aged 75 and older with cancer (n = 150), compared to a matched group without cancer (n = 138). A second aim was to investigate factors associated with poor QoL in people aged 75 and older. QoL was measured with Short Form (SF-12) and European Organisation for Research and Treatment of Cancer Quality of Life Questionnaire (EORTC QLQ-30). The study showed that the cancer group had lower (poorer) scores in different domains of QoL, more complaints, and more self reported diseases than the group without cancer. In both groups, oldest old persons had more complaints than the youngest old. The cancer group had significantly more complaints than the noncancer group. In the youngest old, the cancer group had significantly more complaints than the comparison group. Women with cancer reported the poorest QoL compared to men with cancer and women without cancer. Receiving help for daily living from others and degree of complaints were associated with poor QoL for both the physical and mental component scores (PCS, MCS) of the SF-12. Thus, caregivers should be aware that the most vulnerable cancer patients are women, and that the complaints by cancer patients have implications for QoL especially among the youngest old. PMID- 15287274 TI - Quality of life of primary caregivers of elderly with cerebrovascular disease or diabetes hospitalized for acute care: assessment of well-being and functioning using the SF-36 health questionnaire. AB - OBJECTIVE: To use the Short Form 36 (SF-36) to determine the extent to which health-related quality of life (HRQOL) is decreased among primary caregivers of patients with cerebrovascular accident (CVA) or diabetes mellitus (DM) compared to normal Taiwanese population and to identify the determinants of this decrease. METHODS: Data from a cross-sectional survey of 187 primary caregivers who had responsibility for inpatients with a medically verified diagnosis of CVA or DM were compared to those of randomly selected residents. Data were collected by face-to-face interviews with trained interviewers. RESULTS: The age- and gender adjusted mean differences of caregivers on primarily mental scales of SF-36 were significantly negative compared to general population, as much as a 3-12 points reduction on this 100-point scale. While the age- and gender-adjusted mean differences on physical functioning and bodily pain scales were significantly positive, as much as a 3-6 points increase. Effects of caregiving on the perceived social life of the caregiver and disability of inpatients in eating and getting in/out of bed were associated with the SF-36 Physical Component Scale (PCS) score while female gender, type of caregiver, care conflicts, degree of care demand of daily living, and effects of caregivering on perceived social life of caregivers were negatively associated with the SF-36 Mental Component Scale (MCS) score. CONCLUSIONS: Primary caregivers of CVA or DM hospitalized elderly have poorer mental but better physical well-being than the population norm. Both caregiver and inpatient factors contribute to caregivers' HRQOL. PMID- 15287275 TI - The relationship between quality of life and psychiatric impairment for a Taiwanese community post-earthquake. AB - This purpose of this study was to investigate the relationship between quality of life and psychiatric impairment in a Taiwanese community located near the epicenter of the 1999 earthquake, as assessed four to six months after the natural catastrophe. Trained assistants interviewed the 4223 respondents using the disaster-related psychological screening test (DRPST), an instrument specifically designed and validated by senior psychiatrists for assessment of psychiatric impairment after natural catastrophe. Additionally, the 36-Item Short Form Health Survey (SF-36) was used to evaluate quality of life. The collected results were analyzed using Windows SPSS 10.0 software. Psychiatric impairment rated moderate to severe was assessed for 1448 (34.3%) of the responding residents. The 4223 respondents were divided into 4 psychiatric-impairment groups based on DPRST score: healthy (n = 952); mild impairment (n = 1823); moderate impairment (n = 1126); and, severe impairment (n = 322). The four groups were compared for a number of salient factors, including gender, age, current marital status and psychiatric-impairment score, to determine impact on quality of life. Respondents assessed as psychiatrically impaired tended to be older, female, divorced/widowed, and less educated, and they were more likely to have experienced major familial financial loss as an immediate consequence of the earthquake. Further, the greater the severity of the psychiatric impairment, the lower the scores for quality of life, for both the physical and mental aspects of this important general indicator. PMID- 15287276 TI - The development and validation of a screening instrument to identify hospitalized medical patients in need of early functional rehabilitation assessment. AB - The purpose of this study was to develop a scale that identifies hospitalized patients in need of physical therapy (PT) and/or occupational therapy (OT) assessments. Preliminary scale items were tested for reliability among 52 patients and remaining items were then administered to 299 patients and items that were associated with the concept of 'need for an assessment' on multivariate analyses were selected as final scale items. The concept of need was based on the clinical judgment of physical and occupational therapists. Receiver operator characteristic (ROC) curves were constructed to determine a cut-off score and the predictive ability of this score in determining length of stay and utilization of services was evaluated among 200 patients. The final scale contains two components. The PT component addresses ambulation, falls, breathing, and activities of daily living (ADL). The OT component addresses swallowing, ADL and instrumental activities of daily living (IADL). The area under the ROC curves of the PT and OT components were 0.71 and 0.72, respectively. Both components predicted length of stay and utilization (p < 0.05). In summary, this scale provides a mechanism for targeting patients for early PT and OT assessment and provides a basis for testing the effectiveness of early PT and OT interventions. PMID- 15287277 TI - Longitudinal changes in health status using the chronic respiratory disease questionnaire and pulmonary function in patients with stable chronic obstructive pulmonary disease. AB - Long-term changes in health status have been less evaluated in patients with chronic obstructive pulmonary disease (COPD), in comparison to the changes in forced expiratory volume in 1 s (FEV1). Accordingly, we examined the clinical course of health status as well as pulmonary function in COPD patients, and investigated the relationship between the change in health status and the change in pulmonary function in a 3-year longitudinal study involving 224 patients with COPD. Health status using the Chronic Respiratory Disease Questionnaire (CRQ) and pulmonary function were measured at baseline and every six months over three years. We used the random effects model for the slopes to estimate the longitudinal changes. A total of 147 patients completed the 3-year study. The dyspnoea, fatigue, and emotional function domains of the CRQ declined slowly but significantly over 3 years (p = 0.001, 0.003, and 0.004, respectively) with a mean decline rate of 0.08/year. This means that it would take about 6 years to reach the minimal important change of 0.5 on the CRQ. The mean decline in post bronchodilator FEV1 was 60 ml/year. None of the changes in any of the domains of the CRQ were significantly correlated with the changes in pulmonary function. We have found that, in comparison to the decline in pulmonary function, health status evaluated by the CRQ declined significantly but very slowly in three of four domains over three years in patients with COPD. Furthermore, we have demonstrated that there was no significant relationship between the change in health status and the change in pulmonary function. PMID- 15287278 TI - Assessing the quality of life of adults with chronic respiratory diseases in routine primary care: construction and first validation of the 10-Item Respiratory Illness Questionnaire-monitoring 10 (RIQ-MON10). AB - BACKGROUND: As doctors' judgements about the burden of a disease often differ from patients' own assessments a manageable method to incorporate the latter into routine care might support patient-centered decision-making. For this purpose we shortened the 55-Item Quality of Life for Respiratory Illness Questionnaire (QoL RIQ). METHODS: Secondary analyses of the data of 3 controlled studies (n = 328, 502 and 555). PROCEDURES: inter-item correlations, scale distributions, Cronbach's alpha and factor analysis. Dyspnoea, forced expiratory volume in 1 s (FEV1), COOP/WONCA charts, the Medical Research Council-ECCS symptoms questionnaire and the MOS-SF 36 served as criteria to test validity and responsiveness. RESULTS: Item-reduction resulted in a 10-item short form (alpha's 0.87-0.90), consisting of 2 5-item factors: (1) physical and emotional complaints and (2) physical and social limitations. The correlations of the short form with dyspnoea (r from 0.57 to 0.60), the generic health status instruments (r from 0.39 to 0.59) and lung function (r from 0.10 to 0.15) fulfilled the criteria. FURTHER RESULTS: a clinical relevant score difference (> 0.5) between upper and lower quartiles of the convergent instruments, an intraclass correlation between repeated scores in a stable group of 0.82 and a standardised response mean of 0.86 in an improved group of patients. CONCLUSIONS: The short form (RIQ-MON10) maintained the psychometric properties of the original instrument and is promising for assessing quality of life (QoL) during routine primary care visits. PMID- 15287279 TI - Differences in health-related quality of life and treatment preferences among black and white patients with end-stage renal disease. AB - BACKGROUND: Relatively little is known about racial differences in health-related quality of life (HRQL) among patients receiving dialysis for end-stage renal disease (ESRD) or how such differences may relate to preferences for renal transplantation. METHODS: We surveyed 1392 patients, ages 18-54 approximately 10 months after they initiated dialysis in 4 regions of the United States. The HRQL measures analyzed were overall health, emotional health, physical activity, energy level, social activity, and effect of ESRD on daily life. We also examined whether the measures of HRQL were associated with patients' preferences for renal transplantation by race. RESULTS: After adjustment for socioeconomic and clinical characteristics, Black women and men reported better overall health than White women and men, respectively. Black women reported higher energy levels than White women, and Black men reported less negative effects of ESRD on daily life compared to White men. Black men with high levels of physical activity were less likely to be certain about preferring a transplant than White men with similar levels of physical activity. CONCLUSIONS: Black patients receiving dialysis reported better HRQL than White patients, even after controlling for potential confounders. Racial differences in preferences for renal transplantation among men may be associated with their levels of physical activity. PMID- 15287280 TI - Development of a pain and discomfort module for use with the WHOQOL-100. AB - Clinicians and researchers have become increasingly interested in the impact of chronic pain (CP) on quality of life (QoL). This report describes the qualitative stages of developing a pain and discomfort module for persons with CP to be used with the UK World Health Organisation generic measure of quality of life (WHOQOL). The aims were to investigate patients' perceptions of CP and its effect on QoL, and to generate items to be used in the development of a module appended to the UK WHOQOL-100. At the first stage (study 1), six focus groups of patients were invited to discuss how living with pain and discomfort affected QoL. At the second stage (study 2), an international web survey was conducted with English speaking respondents. Ten new facets of QoL were identified by the focus groups: flare-ups, pain relief, anger and frustration, vulnerability/fear/worry, uncertainty, loss/loneliness/feeling alone, positive strategies, communication, guilt and burdening others, and relationship with health care providers. The web survey confirmed and validated these new facets. Although the WHOQOL-100 is a reliable and valid measure of QoL for use in CP, this study shows that additional areas must be assessed when measuring the impact of CP on QoL. PMID- 15287281 TI - Glycemia and the quality of well-being in patients with diabetes. AB - OBJECTIVES: To investigate the cross-sectional relationships among self-reported frequencies of symptomatic hyperglycemia and hypoglycemia, HbA1c, and symptoms in the Quality of Well-Being Self-Administered (QWB-SA), and to examine the associations among these measures of glycemia and health-utility scores. METHODS: The study group included 1522 patients with diabetes who attended University of Michigan Health System clinics. Published studies were reviewed to identify symptoms in the QWB-SA that might be associated with measures of glycemia. Linear regression analyses were performed to evaluate the strength of the associations among the frequency of self-reported measures of glycemia, QWB-SA symptoms, and QWB-SA-derived health-utility scores. RESULTS: Frequency of hyperglycemic symptoms was associated with 3% of the variance in the QWB-SA-derived health utility score in type-1 diabetes and with 5% of the variance in type-2 diabetes. Frequency of hypoglycemic symptoms was not associated with the QWB-SA-derived health-utility score in type-1 diabetes but was associated with 1% of the variance in type-2 diabetes. HbAlc levels were not significantly associated with QWB-SA-derived health-utility scores. After controlling for age, gender, and complications, frequency of hyperglycemic symptoms was significantly associated with QWB-SA-derived health-utility scores in type-1 and type-2 diabetes. CONCLUSIONS: Reported frequency of hyperglycemic symptoms is associated with symptoms included in the QWB-SA and with QWB-SA-derived health-utility scores. The QWB-SA may be an appropriate measure to assess the health burden of hyperglycemia. PMID- 15287282 TI - Design of an individualised measure of the impact of macular disease on quality of life (the MacDQoL). AB - The aim of this study was to design an individualised questionnaire to measure the impact of macular disease (MD) on quality of life (QoL). Principles underlying the Schedule for the Evaluation of Individual Quality of Life (SEIQoL) interview method and the Audit of Diabetes-Dependent Quality of Life (ADDQoL) diabetes-specific questionnaire influenced the Macular Disease-Dependent Quality of Life (MacDQoL) design. The MacDQoL specifies domains of QoL that were selected using focus group methodology and refined following a postal pilot study of members of the UK Macular Disease Society (MDS). Respondents rated the impact of MD on each domain and the importance of each domain to their QoL. Mean domain scores from 69 respondents indicated that MD had a negative impact on all the domains of QoL investigated in the measure. There was preliminary evidence of good internal consistency reliability (Cronbach's alpha = 0.93, n = 37). Respondents who were registered partially sighted (P/S) or blind reported poorer QoL than those who were not registered (Kruskal-Wallis: chi2 = 14.03, n = 62, p < 0.001). This evidence suggests that the measure will be sensitive to subgroup differences. The instrument has been further refined following the pilot study. The MacDQoL is being used in clinical trials and psychometric evaluation of the measure will be carried out using trial data. The measure is available for clinical use and has been linguistically validated in 15 other languages. PMID- 15287283 TI - The current status of psychotherapy. AB - Psychotherapy has a long history but its practice has always been strewn with controversy. In this review, the current status of psychotherapy is examined by setting its development in historical perspective. While previous practice was often based on the pronouncements of "masters", current approaches are almost always embedded in both rigorous theoretical formulations and frequently also in empirically derived data on efficacy. A fundamental understanding about the mechanisms of action of psychotherapy is a promising new development that is emanating from modern techniques of neurosciences and neuroimaging. Whether such understanding will lead to a renaissance in the clinical utility of psychotherapy is still early to say. However, there is little doubt that the provision of a holistic care for patients with psychological and mental disorders in particular, and most physical conditions in general, should be informed by an appreciation of the bi-directional nature of the relationship between the mind and the body and should therefore include the provision of appropriate psychotherapeutic interventions. PMID- 15287284 TI - Hepatitis B virus in Nigerians with lichen planus. AB - BACKGROUND: [corrected] Lichen planus had been reported as one of the cutaneous manifestations of Hepatitis B virus (HBV) in the literature. The prevalence of HBV among Nigerians with lichen planus has not been documented in the literature despite the high prevalence of HBV in the community, and the reports of a possible relationship between lichen planus and HBV from this centre and from other regions. The aim of this study was to determine the prevalence of HBsAg amongst Nigerians with lichen planus. METHOD: Sixty Nigerians with lichen planus (LP group) and 30 patients with other dermatoses not reportedly associated with HBV (control group A) and 30 apparently normal subjects (control group B) were screened for the presence of HBsAg by second generation ELISA. RESULTS: Nine (15%) of the 60 LP group, 2 (6.2%) of the 30 control group A and 2 (6.2%) of the 30 control group B were HBsAg seropositive. CONCLUSION: This study found a higher prevalence of HBsAg in patients with lichen planus when compared with patients with other cutaneous dermatoses and apparently normal individuals. Although a causal relationship between HBV has not been established from this study, this report reiterates the importance of screening patients with lichen planus for the presence of HBV and instituting therapy in those found positive. PMID- 15287285 TI - Attitudes of patients towards voluntary human immunodeficiency virus counselling and testing in two Nigerian tertiary hospitals. AB - BACKGROUND: Despite new scientific evidence establishing the benefits of counselling and testing as key elements in human immunodeficiency virus/acquired immune deficiency syndrome (HIV/AIDS) prevention strategy, inappropriate HIV screening without consent and counselling is frequent in Africa. Presumed high rejection rate of HIV test has been adduced to this practice. OBJECTIVE: To determine the acceptability of HIV conselling and testing among Nigerians. METHODS: Consecutive sixty indigenous Nigerians aged 35.10+/-11.31 years with male: female ratio of 2:1, and made of clients with clinically suspected AIDS (20), diseases unrelated to AIDS (15), dermatological problems (10), sexually transmitted diseases (9), and asymptomatic persons (6) were studied. RESULTS: Fifty-three (88.3%) subjects gave informed consent to HIV screening. Of the 53 consenters, 32 (60.4%) were seropositive while 21 (39.6%) were seronegative. Five clients (9.4%) (1seropositive + 4 seronegative consenters) did not turn up for their results, and 2 (3.8%) seronegative consenters did not want to know their serostatus. The reactions to disclosure of seropositive results included grief 9 (28.1 %), indifference 8 (25 %), surprise 5 (15.6%), family concern 5 (15.6%), denial 3 (9.4%) and suicidal ideation 2 (6.3%). Thirteen (40.6%) seropositive clients showed willingness to disclosure of their serostatus to family members including the father (58%), senior brother (23%), wife (11%) and others (8%). Direct cost of screening was N400.00 (U$3.10) per client. An average of 18 minutes per client was spent on counselling. CONCLUSIONS: This study demonstrates the feasibility of VCT in Nigerian hospitals. PMID- 15287286 TI - Radiological changes and complications associated with nasal polyposis. AB - BACKGROUND: Nasal polyps are pedunculated portion of oedematous mucosa of the nose or paranasal sinuses. Simple mucous or benign nasal polyps are an early recognizable clinical entity. METHODOLOGY: A 5-year (1998--2002) study of 63 nasal polyposis patients seen at the department of Otorhinolaryngology of University College Hospital Ibadan, Nigeria with radiological changes and complications was done. RESULTS: Out of these 63 patients, 55 (87 %) had associated radiological changes of which 34 (62 %) were males and 21(38%) females with M:F of 1.6:1 and with an average age of 34 years. The duration of symptoms ranged from 2 months-14 years. The radiological changes were mainly seen in the maxillary sinus (40:33%), nasal cavity (37:30%) and ethmoidal air cells (21:17%), Sinus opacifications (90-100%) constituted the most common features seen followed by nasal cavity loss of radio-translucency (97 %). 50(79 %) had complications with sinusitis (98 %) being the most common complication seen. CONCLUSION: The nasal polyposis has been found in this study to cause the radiological changes mainly in the maxillary sinus. The plain radiographs of the paranasal sinuses demonstrated by at least the occipito-frontal, occipito-mental and lateral views would show the extent of the disease in the nose and paranasal sinuses to certain good extent. PMID- 15287287 TI - Enhanced wound contraction in fresh wounds dressed with honey in Wistar rats (Rattus Novergicus). AB - BACKGROUND: Due to reports that honey accelerates wound healing, an investigation on its role in wound contraction in fresh wounds inflicted on wistar rats was carried out. METHOD: Twenty adult male wistar rats had 2cm by 2cm square wound inflicted on their right dorsolateral trunk. They were divided into two groups. The experimental group had their wounds dressed with honey while the control group had normal saline dressing. Wound dressing was done every five days and measurements taken at each dressing. Wound morphology was also assessed. RESULTS: Dressing with honey significantly enhanced percentage wound contraction on day 10 with value of 79.20+/-2.94 compared to control value of 53.50+/-4.32. p=0.0. The mean wound measurement on day 10 reduced significantly in honey group, 1.15+/ 0.18 compared to control group 2.38+/-0.28. p=0.002. However, there was no significant difference in fibroblast count per high power field in honey group 68.0+/-2.59 compared to control 90.2+/-17.40, p=0.242. Honey dressing increased mean blood vessel count per high power field, 18.8+/-3.77 albeit non significantly when compared to control value of 13.4+/-2.44, p=0.264. Also honey dressing caused increased granulation tissue formation in wounds dressed with honey compared to control group. CONCLUSION: Our study suggests that honey dressing enhances wound contraction in fresh wounds which is one of the key features of wound healing. PMID- 15287288 TI - A cephalometric study of antero-posterior skeletal jaw relationship in Nigerian Hausa-Fulani children. AB - OBJECTIVE: To determine the antero-posterior skeletal jaw relationship in Hausa Fulani children in Nigeria. SETTING: This study was carried out in 1998 at the Maxillo-facial unit of the Ahmadu Bello University Teaching Hospital, Kaduna, Nigeria. MATERIALS AND METHODS: 100 subjects aged 11-13 years of Hausa-Fulani ancestry with no previous orthodontic treatment were selected for the study. Lateral cephalometric radiographs were traced and analysed to produce values for SNA, SNB and ANB. RESULTS: The mean SNA was 82.4 degrees, mean SNB 803 degrees and mean ANB 2.1degrees The normal range of ANB values was 0.5 degrees-4 degrees CONCLUSION: The values obtained differ from those of other population groups and should be used as guidelines in the orthodontic treatment of the ethnic group studied. PMID- 15287289 TI - Eclampsia and abnormal QTc. AB - BACKGROUND: Abnormalities of calcium and magnesium metabolism are known risk factors for QT interval prolongation and have been reported in eclampsia. OBJECTIVE: To determine if eclampsia is associated with QT abnormalities METHODS: In a prospective study, the QT interval corrected for heart rate (QTc), serum calcium, magnesium and potassium were compared among 30 intrapartum eclamptics and 30 age, parity and gestational age-matched women with uncomplicated pregnancy RESULTS: Primigravidae made up 93.3 % of patients. Mean age was 19.5+/-4.2 years Blood pressure was significantly higher among patients than controls (163.0+/ 34.7mmHg versus 120.4+/-18.6mmHg systolic, p<0.05) and (104.7+/-15.2mmHg versus 79.6+/-10.7mmHg, p<0.05). Heart rate ranged from 76 to 163 beats per minute (bpm) and 65 to 112bpm among patients and controls respectively. The corresponding QTc were 390-572 and 390-460 respectively. Compared to the controls, patients had significantly higher mean heart rate (121.1+/-24.9bpm versus 89.3+/-12.9bpm, p<0.05) and higher QTc (470.4+/-42.4 versus 436.3+/-17.7; p<0.05). Compared to the controls, the eclamptics also had higher frequency of sinus tachycardia (90% versus, 13.3%) Odd Radio (OR) =29.57; 95% Confidence Interval (CI) = 5,87-174.08, p=0.00), abnormal QTc (46.7% versus 6.6%, OR = 9.2; 95% CI =1.61-68.48, p=0.01) and T-axis deviation (26.6% versus 3.3%, OR=10.55, 95% CI=1.2-244.3, p=0.03) Serum calcium level was significantly lower among patients than controls (2.0+/ 0.4mmol/L versus 2.3+/-0.2mmol/L, p<0.05). The eclamptics had higher frequency of hypocalcaemia than the controls (40 % versus 6.6%; OR = 14, 95% Cl = 1.58-316.9, p=0.01). Serum magnesium, potassium, urea and creatinine levels were similar between the two groups. CONCLUSION: Eclampsia is associated with abnormal QTc that may be related to maternal hypocalcaemia. Correction of electrolyte abnormalities and cautions about agents that have potential for QT prolongation are vital in the care of eclamptics. PMID- 15287290 TI - Day case transurethral prostatectomy in Nigeria. AB - BACKGROUND: Caudal block regional anaesthesia has been used over the years for out-patient procedures, and in transurethral resection of the prostate gland in Nigeria since 1995. In a preliminary study involving 10 selected patients undergoing TURP, spontaneous voiding resumed on the operative day, and their discharge on the same day did not in any way lead to any adverse events. This larger series further confirms the safety of transurethral resection of the prostate gland as a day case procedure. METHOD: One hundred and eighty patients with obstructing benign prostatic enlargement on urethral catheter drainage with prostate glands weighing 60g or less on ultrasound assessment, were subjected to transurethral resection of the prostate gland (TURP) as day-cases under caudal block regional anesthesia using 2% xylocaine with 1 in 80,000 adrenaline. Hemostasis was secured until effluent of the irrigation fluid from the bladder was totally free of any visible trace of blood. A catheter was not inserted postoperatively. The patients were discharged on the same day after they had satisfactorily voided. RESULTS: These patients resumed spontaneous voiding postoperatively before discharge on the operative day. Their discharge on the same day did not in any way lead to any adverse events. CONCLUSIONS: With a better understanding of the anatomy of the innervations and blood supply of the prostate gland, and proper patient selection, this larger series has confirmed that day-case TURP without postoperative catheterization can now be safely added to the list of day case procedures. PMID- 15287291 TI - A review of genitourinary cancers at the Korle-Bu teaching hospital Accra, Ghana. AB - OBJECTIVE: To determine the frequency and pattern of genitourinary malignancies seen at the Korle-Bu Teaching Hospital, Accra, Ghana MATERIALS AND METHODS: A retrospective review of genitourinary malignancies seen at the Korle-Bu Teaching Hospital from 1980 to 1990 was undertaken. Data was obtained from the operating theatre register, histopathology reports, and patient case notes. Information retrieved included age and sex of patients, organ involved and laterality where appropriate and tumour type. RESULTS: 548 genitourinary malignancies were seen, of which 479 (87.4%) were in males and 69 (12.6%) in females. Adults comprised 93.4% and children 6.6%. The organ-specific distribution was as follows-prostate 349 (63.7%), bladder 117 (21.3%), kidney 57 (10.4%), testis 13 (2.4%), penis 10 (1.8%) and one each of the ureter and urethra. The kidney tumours comprised nephroblatoma (56.1%), adenocarcinoma (35.1%) with the rest being of urothelial origin. Of the bladder tumours, 50.4 % and 44.8 % were transitional cell and squamous cell carcinoma respectively. Virtually all the prostatic cancers (99%) were adenocarcinomas. Of the testicular tumours 8(61.5%) were of germ cell and 5 (38.5%) non-germ cell origin. The penile cancers were all squamous. The ureteric and urethral tumours were due to transitional cell and squamous cell carcinomas respectively. CONCLUSION: Prostatic carcinoma was the predominant genitourinary tumour, accounting for nearly two-thirds of cases, followed by the bladder and the kidney. Other tumours were relatively uncommon. Transitional cell carcinoma of the bladder was seen a little more commonly than the squamous type. PMID- 15287292 TI - Nasopharyngeal cancer at the University College Hospital Ibadan Cancer Registry: an update. AB - BACKGROUND: Only two reports on nasopharyngeal cancer (NPC) are available from this large centre and both covered the years 1961--1966 and 1966--1980. OBJECTIVE: This study describes the update of nasopharyngeal cancer at the Ibadan Cancer Registry from 1981 to 2000. METHOD: This is a retrospective review of all histologically confirmed cases of nasopharyngeal cancer accumulated in the Ibadan Cancer Registry from 1981--2000. RESULTS: Two hundred and twenty three cases were analysed consisting of 156 (70%) males and 67 (30%) females with a male: female of 2.3:1. There is a steady increase in the incidence of nasopharyngeal cancer over the last two decades. Overall, the mean age was 41.1years (age range 10 to 81 years). The females had a mean age of 36.1years (age range 11 to 80 years) and males 43.2 years (age range 10 to 81 years). The peak age group of incidence for the females was 20-29 and 50-59 for the males with an almost equal incidence in the preceding three (3) decades. The ratio of Regaud to Schmincke type cancer reversed with increasing age amongst the females with the Schmincke type more common in the first decade but this was not reproduced in the males. CONCLUSION: There is a steady increase in the incidence of nasopharyngeal cancer over the last two decades. The two main histological types showed differential variation between the sexes suggesting a possible biological effect in the manifestation of this disease. PMID- 15287293 TI - Early results of transurethral vaporisation of prostate in Nigeria. AB - OBJECTIVE: to determine the effectiveness of transurethral electrovaporisation of prostate for the relief of obstructive prostatic disease in Nigeria. METHODS: Consecutive patients presenting with obstructive prostatic disease with prostate size of less than 40gm were treated by transurethral electrovaporisation of prostate using a 5mm rollerball electrode. RESULTS: A total of 27 patients were entered into the study, 7(26%) had clinically malignant prostates; 2 patients (7%) had immediate post operative retention requiring resection of residual prostate. No patients exhibited the clinical features of TUR syndrome, all patients were discharged by the 4th post operative day and required no blood transfusions. CONCLUSIONS: Transurethral electrovaporisation is effective for treatment of obstructive prostatic disease, benign or malignant. using the 5mm rollerball instead of the recommended vaportrode still confers benefit over TURP. TEVP confers all the benefits of TURP with less mobidity. PMID- 15287294 TI - Characteristics of type 2 diabetics presenting with end stage renal disease at the Jos University Teaching Hospital, Nigeria. AB - BACKGROUND: Diabetes mellitus is an increasingly common cause of end-stage renal disease (ESRD) in Nigeria. We describe the clinical characteristics of Nigerian diabetics presenting with ESRD, as data obtained would provide baseline information for management policy formulation. METHODS: Twenty-one diabetics (16 males and 5 females) with ESRD seen in the Nephrology Unit of the Jos University Teaching Hospital were studied. Both clinical and laboratory parameters were assessed. RESULTS: The mean age of the patients was 55.5+/-9.8 years with a mean duration of diabetes being 7.7+/-8.2 years. Retinopathy, hypertension and peripheral vascular disease were present in 75.5%, 71.4% and 57.1% of the patients respectively. The mean fasting blood glucose was 6.0+/-2.7 mmol/L. Hypertriglyceridemia was the most common dyslipidemia seen in 38.1% of the patients, followed by reduced high- density lipoprotein (HDL) cholesterol in 33.3% and hypercholesterolemia in 23.8%. Common electrocardiographic abnormalities included myocardial ischemia, left atrial hypertrophy and left ventricular hypertrophy. CONCLUSION: The care of these patients should take into consideration the control of hypertension and dyslipidemia as cardiovascular event is common in them. PMID- 15287295 TI - Chronic dehydration and symptomatic upper urinary tract stones in young adults in Ibadan, Nigeria. AB - INTRODUCTION: Upper urinary tract stones are relatively uncommon in Nigeria and they are most often seen in men in their 4th and 5th decades. There is however no recent report on this disease from our locality. This retrospective study was done to evaluate the pattern of presentation of upper tract urolithiasis in our institution. PATIENTS AND METHODS: We reviewed all cases of renal stones referred to a single Consultant in a Teaching Hospital in Southwestern Nigeria over a two year period. RESULTS: Twenty cases of renal calculi presented within the study period. All patients presented with sudden onset of loin or back pain, and the diagnosis was confirmed radiologically. The mean age of our patients was 27 years (age range 13-38 years), and the male to female ratio was 1.5: 1. Nineteen patients (95%) reported poor fluid intake (<1.5L/day), and 17/20 (85%) frequently total fasted totally (no water or food intake) for religious reasons. Serum calcium was normal in 15 of 16 patients (94%) and only marginally raised in the remaining patient. All patients were treated with a high fluid intake (3L/day) and analgesics, and 16 of the patients (80%) passed their stones spontaneously. The other four are currently pain free, one of whom is awaiting surgery. CONCLUSION: This (uncommon) occurrence of upper tract urolithiasis in young adults in Ibadan may be related to chronic dehydration exacerbated by religious fasting. Further studies are required to explore this relationship, PMID- 15287296 TI - Association between HIV/AIDS and malignancies in a Nigerian tertiary institution. AB - BACKGROUND: Following the outbreak of the Human Immunodeficiency Virus (HIV) infection in 1981, there has been a widespread increase in the incidence of many malignancies including non-Hodgkin's lymphoma, cervical carcinoma and Kaposi's sarcoma. The scarcity of reports linking HIV infection with malignancies in Nigeria necessitated this study. We prospectively screened one hundred patients with various forms of malignancies seen at the University College Hospital Ibadan, Nigeria between October 2001 and June 2002 for HIV infection by the Enzyme Linked Immunosorbent Assay (ELISA) method and confirmed with the Western Blot method. RESULTS: Six of the patients were found to be seropositive for HIV antibodies. There were 41 males (41%) and 59 females (59%) with age ranging from 7 months to 80 years and a median of 46 years. The HIV seropositive patients were between 29 and 35 years of age. Two patients with non-Hodgkin's lymphoma (NHL) and one patient each with carcinoma of the cervix, Kaposi's sarcoma. Hodgkin's lymphoma and carcinoma of the breast were HIV seropositive. All the p values were greater than 0.05. CONCLUSION: The seroprevalence of HIV infection in patients with malignancies in this study was 6%. Despite the HIV/AIDS epidemic, there is yet no significant statistical relationship between HIV infection and malignancies in this environment. Larger, preferably multicenter studies need to be carried out to ascertain the relationship between HIV infection and malignancies in Nigeria. PMID- 15287297 TI - Normal values of medial and lateral canthal distances in 3 to 18 year-old Nigerians. AB - BACKGROUND: This study was carried out on male and female Nigerians whose age ranged from 3 to 18 years in order to provide a database of canthal measurements for a predominantly black population and compare them with Caucasians. STUDY DESIGN: All the healthy pupils and students were randomly selected. The ages of the children, adolescents and young adults were approximated to their nearest birthdays. Two different researchers measured each parameter and the mean values were recorded. SETTING: Nursery, primary, secondary schools and the University of Benin, in Benin City, Nigeria. RESULTS: Four hundred and sixty eight males (53.4%) and 408 (46.6%) females making up a total of 876 subjects were studied. The mean values for medial canthal distance for male Nigerians are slightly higher than those established for male Caucasians and these differences are significant (p>0.05). There was no significant difference in the lateral canthal distance between the two groups (p<0.05). Nigerian and Caucasian females have significant difference (p>0.05) in mean values for medial canthal distances but not in lateral canthal distance (p<0.05). The difference in these distances between Nigerian males and females are not significant. Weight and lateral canthal distance showed a covariance of 6.980 while age and lateral canthal distance, age and medial canthal distance showed a covariance of 2.970 and 1.140 respectively. There was no correlation between age, height, weight and the distances measured. CONCLUSION: Medial canthal distances between male and female Nigerians compared to males and female Caucasians show significant variations but not in the lateral canthal distances. PMID- 15287298 TI - Pattern of chronic liver disease in Omani children--a clinicopathological review. AB - OBJECTIVE: To determine the pattern of chronic liver disease in Omani children. STUDY DESIGN: Seventy six children [43M : 33F] aged 4 days to 10 years, referred to the Paediatric Gastroenterology clinic of the Sultan Qaboos University Hospital, Muscat, Oman, between 1995--2000 for evaluation of liver disease were studied. Liver biopsies were performed in all and tissues obtained processed and examined for histological lesions. RESULT: The main histological diagnoses were neonatal hepatitis (22) biliary atresia (9) biliary hypoplasia (7), cirrhosis (7) and congenital hepatic fibrosis (5). Hepatomegaly with or without jaundice was the indication for liver biopsy in the majority of patients studied. CONCLUSION: The study has provided background information on the occurrence of specific liver diseases in Omani children. Neonatal hepatitis syndrome was the most common diagnosis before the age of 2 years. PMID- 15287299 TI - Computed tomography and childhood seizure disorder in Ibadan. AB - BACKGROUND: Computed Tomography (CT) is an important tool for neuroimaging, it offers an opportunity to investigate structural lesions as a cause of seizures with little morbidity. This study is designed to evaluate it's applicability in children with epileptic seizures. METHOD: It is a descriptive study of the CT scans of the 103 consecutive children who were referred to the CT suite of the University College Hospital on account of seizure disorders over a 5 year period (1997--2001). RESULTS: Only 103 (4.6%) of the subjects who had cranial scans done in five years were children with seizures disorders. The CT scans were abnormal in 53 (51.5 %). Hydrocephalus was the most common finding in 14 (13.6%). Cerebral atrophy and infarct were reported in 10.6% and 8.7% respectively. The outlined cranial fractures found in 6.8% were all depressed. A high incidence (74.4%) of abnormal scans was reported in the children with partial seizures. Thirty-three (62.3%) of the abnormal scans were amenable to surgery. The presence of neurologic deficit increased the yield of abnormal CT features. CONCLUSION: CT scans are of extreme value in the screening and definitive evaluation of seizures in children. It is advocated for excluding treatable conditions and monitoring progression of the disorder. PMID- 15287300 TI - A case of neuropsychiatric lupus with myelopathy successfully treated with corticosteroids. AB - This report describes a 16-year old female patient who presented with acute paresis in both lower limbs, acute urinary retention, blurred vision and arthritis. The patient was diagnosed as having systemic lupus erythematosus with myelitis and bilateral abducent nerves palsy. The administration of steroids resulted in marked improvement in her neurological symptoms. PMID- 15287302 TI - Maxillary haemangiopericytoma: a case report. AB - BACKGROUND: Haemangiopericytoma is a very rare slow-growing vascular tumour with a variable malignant potential, constituting less than 1% of all neoplasms. It may arise from any blood vessel and in any organ of the body. Primary haemangiopericytoma of bone is even rarer, constituting about 0.1% of bone tumours. The tumour is extremely rare in Africans and particularly in the head and neck region. STUDY DESIGN: We describe the case of a 66-year old Nigerian with haemangiopericytoma of the maxilla, who presented with a recurrent but painless jaw mass. RESULTS: Surgical resection of this tumour is potentially bedevilled with the risk of torrential haemorrhage and high rate of recurrence. This risk may be substantially reduced by wide surgical resection with a careful microscopical examination of the resection margins and the institution of adjuvant radiotherapy in incompletely resected tumours. Chemotherapy has no known role in the management of haemangiopericytoma. Postoperative radiation therapy appears to be effective against tumour recurrence. CONCLUSION: Even then, long term follow-up is essential in all cases. To our knowledge, this is the first report of this entity in an African. PMID- 15287301 TI - Mature sacrococcygeal teratoma: a case report and literature review. AB - BACKGROUND: Sacrococcygeal teratomas are derived from embryonic germ cell layers. They present mostly in infancy and are extremely rare in adults; with an associated risk of malignancy. Modern imaging technique may be helpful to delineate the extent of the mass but surgical excision is generally indicated at the time of detection. PATIENTS AND METHODS: A case report is presented with a review of literature utilising MEDLINE, Microsoft Net and Yahoo search engines. RESULTS: A three day old female baby presented with a mature sacrococcygeal teratoma containing well-developed limb buds. She had surgical excision and primary repair with good results. A two-year follow up utilising serial serum alpha-fetoprotein assay and CT Scan revealed no evidence of tumour recurrence. CONCLUSION: Sacrococcygeal teratoma is a rare tumour that may be benign or malignant. Complete excision is the primary therapy and is adequate if the tumour is benign. Chemotherapy and radiotherapy are however indicated in malignant cases and in recurrence after previous excision. PMID- 15287303 TI - Gastrodiscoides hominis infection in a Nigerian-case report. AB - Gastrodiscoides hominis is a large fluke of pig and human and constitutes an important parasite of human in Assam, Indian, the Philippines and Southeast Asia. This parasite has not been reported in Nigeria and possibly other parts of Africa. This is a case report of a seven year old Nigerian child who presented with features of malnutrition and anaemia and was found to have Gastrodiscoides hominis and Ascaris lumbricoides. Following clearance of the worms there was tremendous improvement of the health status of the child. The detailed epidemiology of this parasite still remains to be studied in this environment. PMID- 15287304 TI - Recurrent aphthous ulcers today: a review of the growing knowledge. AB - Recurrent aphthous ulcers represent a very common but poorly understood mucosal disorder. They occur in men and women of all ages, races and geographic regions. It is estimated that at least 1 in 5 individuals has at least once been afflicted with aphthous ulcers. The condition is classified as minor, major, and herpetiform on the basis of ulcer size and number. Attacks may be precipitated by local trauma, stress, food intake, drugs, hormonal changes and vitamin and trace element deficiencies. Local and systemic conditions, and genetic, immunological and microbial factors all may play a role in the pathogenesis of recurrent aphthous ulceration (RAU). However, to date, no principal cause has been discovered. Since the aetiology is unknown, diagnosis is entirely based on history and clinical criteria and no laboratory procedures exist to confirm the diagnosis. Although RAU may be a marker of an underlying systemic illness such as coeliac disease, or may present as one of the features of Behcet's disease, in most cases no additional body systems are affected, and patients remain otherwise fit and well. Different aetiologies and mechanisms might be operative in the aetiopathogenesis of aphthous ulceration, but pain, recurrence, self-limitation of the condition, and destruction of the epithelium seem to be the ultimate outcomes. There is no curative therapy to prevent the recurrence of ulcers, and all available treatment modalities can only reduce the frequency or severity of the lesions. PMID- 15287305 TI - Microvascular autologous submandibular gland transfer in severe cases of keratoconjunctivitis sicca. AB - The objective is to evaluate the technique of microvascular autologous submandibular gland transfer for the treatment of severe keratoconjunctivitis sicca. From August 1999 to April 2002, 38 patients with severe keratoconjunctivitis sicca were treated by autologous submandibular gland transfer to the temporal region of the skull. The related vessels were anastomosed to the superficial temporal artery and vein. When the vein was too small, venous bridging was applied. Prior to cutting off the gland, the facial artery was preserved and infused with heparin in normal saline after the gland had been freed to allow inspection of the blood oozing from the three veins. This would be helpful in the selection of a relevant vein for anastomosis. Wharton's duct was transplanted to the upper lateral conjunctiva fornix, and the gland was left denervated. Postoperative scintigraphy with Tc99m pertechnetate, follow-up studies, and management of complications were performed. The transplantations were successful in 33 cases, their symptoms of xerophthalmia disappeared. The discomfort resulting from bright light and wind was also relieved. These patients could stop applying artificial tears. In five patients the transplanted glands did not survive. Epiphora occurred in eight cases. They were successfully treated by reducing the size of the graft. Obliteration of Wharton's duct took place in two cases and was treated by the reconstruction of the duct or duct orifice. Microvascular autologous submandibular gland transfer is a lasting and effective solution for severe cases of keratoconjunctivitis sicca. PMID- 15287306 TI - Are there any complications with bioabsorbable fixation devices? A 10 year review in orthognathic surgery. AB - Bioabsorbable fixation devices have been used in our departments between November 1991 and November 2001 in orthognathic surgery. The aim of this retrospective study was to assess all complications experienced during this time period, when we have operated 163 patients who have undergone 329 orthognathic osteotomies fixated with bioresorbable devices. No postoperative intermaxillary fixation was used. Light guiding elastics were used for 5 to 7 weeks. Patients' acceptance was generally excellent and very few complications occurred during this follow-up of 10 years. The complications were minor and did not affect the end results of the operations. Minor complications occurred in 14 patients (8.6%). Only one patient (0.6%) had a postoperative infection with elevated infection parameters. The other minor complications consisted mainly of dehiscence of the wound and plate exposure together with granulation tissue in the operation field. The rest of the complications occurred in the beginning of our study, when large screw heads on top of the bone irritated the patient and had to be removed. Insufficient fixation resulted in open bite in three patients (1.8%) in the beginning of the trial use of new devices, which no longer are used. Based on our experience, bioresorbable devices are safe to be used in orthognathic procedures. However, there is a learning curve, as there is with all new methods introduced. PMID- 15287307 TI - Le Fort I miniplate osteosynthesis: a randomized, prospective study comparing resorbable PLLA/PGA with titanium. AB - The aim of this study was to compare the use of resorbable osteosynthesis material (LactoSorb) with titanium osteosynthesis for the fixation of Le Fort I osteotomies with respect to long-term stability and morbidity. To achieve exact cephalometric measures, five tantalum micro implants were inserted in the maxilla during surgery. A total of 60 patients undergoing a non-segmented Le Fort I osteotomy were randomized to one of the treatments and were followed for 1 year postoperatively. For the osteotomies fixated with LactoSorb, the lateral cephalometric analysis demonstrated a statistically significant difference in the vertical position of the maxilla after 6 weeks as the position became more superior compared with the postoperative situation (mean change of 0.6 mm). In the titanium group no significant changes in position were observed. There were no statistically significant changes in the position of the maxilla from 6 weeks to 12 months in either of the treatment groups. The changes in maxillary postion were not clinically noticeable in either of the treatment groups, and all treatments were completed with satisfactory results. There were two cases of infection and wound dehiscence in the LactoSorb group, whereas titanium osteosynthesis was more often palpable after 6-12 months and required surgical removal in three cases. PMID- 15287308 TI - Shoulder complaints after nerve sparing neck dissections. AB - The purpose of the study was to analyse the prevalence of shoulder complaints after nerve sparing neck dissection at least 1 year after surgery, and to analyse the influence of radiation therapy on shoulder complaints. Patients were interviewed for shoulder complaints, and patients filled out the shoulder disability questionnaire to evaluate shoulder disability in daily activities. In total 137 patients; 51 after modified radical neck dissection (MRND), 21 after postero-lateral neck dissection (PLND), and 65 after supraomohyoid neck dissection (SOHND) were analysed. After MRND 33.3% of the patients experienced shoulder complaints, after PLND 66.7%, and after SOHND 20% of the patients experienced shoulder complaints. Type of neck dissection was significantly (P < 0.001) related to shoulder complaints. Outcome on the shoulder disability questionnaire also showed a significant (P < 0.01) difference in outcome for type of neck dissection. The prevalence of shoulder complaints after SOHND are low, and reduce disability in daily activities. Radiation therapy does not have a significant effect on shoulder complaints and disability. PMID- 15287309 TI - 10-year follow-up of onlay bone grafts and implants in severely resorbed maxillae. AB - Thirty patients with extremely resorbed maxillae had reconstructive bone grafts from the ala iliaca and endosseous implants in a one-stage procedure. The first ten patients constituted a development group and the following 20 patients a routine group. The marginal bone level and implants success rate was evaluated in a prospective long-term follow-up for a minimum of 10 years (10-13 years). Clinical and radiographic examinations were performed at 6 months and then annually up to 5 years. The final examinations were performed at the 10-year follow-up. The bridges were removed at every clinical examination. Marginal bone loss was seen up to the 3-year examination, where it averaged 4.6 mm in the routine group. Between the 3- and 10-year follow-up no significant change was registered. The initial bone loss was probably due to the design of the 3.6 mm conical unthreaded marginal part of the implant. The implant success rate was 83.1% in the routine group. Failures mostly occurred during the first 2 years (14 out of 20). A substantial amount of bone can be gained in patients with extremely resorbed maxillae, when treated with bone graft according to the procedure described in this study. PMID- 15287310 TI - Peripheral odontogenic tumours--differential diagnosis in gingival lesions. AB - Peripheral odontogenic tumours (POT) are rare benign focal overgrowths of the oral soft tissue, usually occurring in the gingiva. Between 1996-2000, 6 out of 406 excised gingival lesions were diagnosed as POT (1.5%). Tumours included peripheral odontogenic fibroma (2 patients), peripheral calcifying odontogenic cyst (2 patients), peripheral ameloblastoma (1 patient), and peripheral calcifying epithelial odontogenic tumour (1 patient). Review of the literature reveals that peripheral odontogenic fibroma and peripheral ameloblastoma were the most common POT. The purpose of this article was to analyse the clinical data of these tumours according to the presented cases and the literature review, to elucidate typical features of each tumour type and enhance easy identification. PMID- 15287311 TI - Preoperative intravenous tramadol versus ketorolac for preventing postoperative pain after third molar surgery. AB - The objective of this study was to compare the analgesic efficacy of a single dose of preoperative intravenous tramadol versus ketorolac in preventing pain after third molar surgery. Sixty-four patients undergoing elective third molar surgery were randomly assigned into one of the two groups (32 in each group): Group I received tramadol 50 mg, and Group 2 received ketorolac 30 mg intravenously preoperatively before the surgery. After injection of the study drugs, a standard intravenous sedation technique was administered and the impacted third molars were removed under local anaesthetic. The difference in postoperative pain was assessed by four primary end-points: pain intensity as measured by a 100-mm visual analogue scale hourly for 12 h, median time to rescue analgesic, postoperative acetaminophen consumption, and patient's global assessment. Throughout the 12-h investigation period, patients reported significantly lower pain intensity scores in the ketorolac versus tramadol group (P = 0.05, Mann-Whitney U-test). Patients also reported significantly longer median time to rescue analgesic (9.0 h versus 7.0 h, P = 0.007, log rank test), lesser postoperative acetaminophen consumption (P = 0.02, Mann-Whitney U-test) and better global assessment (P = 0.01, chi2 test) for the ketorolac versus tramadol group. Preoperative intravenous ketorolac 30 mg is more effective than tramadol 50 mg in the prevention of postoperative dental pain. PMID- 15287312 TI - Oestrogen replacement therapy promotes bone healing around dental implants in osteoporotic rats. AB - The aim of this study was to investigate the effect of oestrogen replacement therapy on bone healing around titanium implants in osteoporotic rats. Sixty 32 week-old female SD rats were used in this study. Ovariectomies were performed in 40 rats, and other 20 rats had sham operation. Eighty-four days after surgery, osteoporotic changes in proximal tibiae were seen in four ovariectomized rats when compared with two sham-operated rats. Then pure titanium implants were placed in the bilateral proximal metaphyses of the tibiae of the remaining animals. Oestrogen replacement therapy was administrated in 18 ovariectomized rats after implantation. Nine rats from each group (ovariectomized, oestrogen treated and sham-operated) were killed at 28 and 84 days after implantation surgery respectively, and the tibiae specimens were harvested and examined. Both at 28 and 84 days after implantation surgery, most bone histomorphometric indices in the oestrogen-treated group were significantly increased compared with those in the ovariectomized group (P < 0.05 or P < 0.01). Although the oestrogen treated group showed lower trabecular bone volume at 28 days after implant surgery and lower mineralization rate at both the two time points than the sham operated group, there were no significant differences in other bone histomorphometric indices between the oestrogen-treated group and the sham operated group both at 24 and 84 days after implantation. The results of this study suggest that oestrogen replacement therapy may promote bone healing around titanium implants under osteoporotic state, and therefore it seemed to be beneficial to long-term success of dental implants in clinical postmenopausal patients. PMID- 15287313 TI - The effect of alendronate on resorption of the alveolar bone following tooth extraction. AB - Maintenance of alveolar bone width and height following tooth loss is essential with regard to the restoration of missing teeth with endosseous dental implants or prosthodontics approaches. A various amount of alveolar ridge resorption is likely to occur after tooth extraction at the buccal and lingual alveolar bone plates. Bisphosphonates, alendronate, is well known for its potent inhibition of osteoclast-mediated bone resorption. The objective of this study was to examine the inhibitory effect of alendronate on alveolar bone resorption following tooth extraction in rats. Male Wistar Albino rats were divided into three groups: baseline group, saline-treated group and alendronate-treated group. The saline treated group was administered with daily saline solution for 2 and 4 weeks respectively while the alendronate-treated group was given a daily amount of 0.25 mg/kg alendronate subcutaneously for the same periods. The level of urinary calcium, creatinine, and serum calcium, alkaline phosphatase and phosphate were measured. Serum alkaline phosphatase level was measured as a marker of osteoblastic activity. Histopathological sections of 4 microm thickness were obtained from the right first mandibular molar region in a bucco-lingual direction. The number of osteoclasts, osteoblasts, and haversian canals, the number and size of resorptive lacunae, and osteoid formation were evaluated histopathologically. The mean thickness of buccal and lingual alveolar bone was measured. In the alendronate-treated group, both buccal and lingual alveolar bone volume reduction was significantly less than the saline treated group. Significant reduction in serum and urinary calcium levels and the number of osteoclasts revealed the pronounced suppression of bone resorption in the alendronate-treated group. PMID- 15287314 TI - Experimental study of mechanical analysis in mandibular lengthening. Application of strain gauge measurement. AB - In distraction osteogenesis, the assessment of new bone formation and the decision of when to remove the distraction device are very important from clinical viewpoint. The purpose of this study was to measure bone stiffness using a strain gauge during and after mandibular lengthening, and to compare the results with those from radiographic and histological examinations. Twelve adult mongrel dogs served as the experimental subjects. An external distraction device was connected to the mandible, and distraction was started 7 days after operation and continued for 10 consecutive days. The animals underwent strain gauge measurement and radiographic examination of the mandible at the completion of distraction and every 2 weeks until the end of the consolidation period. The animals were then sacrificed at 2, 4, 6, 8, 10, and 12 weeks after completion of the distraction for histologic examinations. There was correlation between the amount of decrease in strain, increase in ratio of radiopacity, and newly formed bone in the distracted area. PMID- 15287315 TI - Arteriovenous fistula after temporomandibular joint arthroscopy successfully treated with embolization. AB - Temporomandibular joint arthoscopy is a minimal invasive surgical procedure commonly used to effectively treat some internal derangement of the TMJ. However, this method is not free of complications. Arteriovenous fistula (AVF) is a lesion that communicates the high flow arterial system and the low flow venous network. We describe a new case of preauricular traumatic AVF successfully treated with external carotid embolization, along with a review of the medical literature. PMID- 15287316 TI - Ultrasound guided removal of an air gun pellet from the temporal fossa: a technical note. PMID- 15287317 TI - Small cell carcinoma metastatic to the mandible. Report of a case. AB - Small cell carcinoma either primary or metastatic is an uncommon malignancy in the oral and perioral tissues. We are reporting such a case. PMID- 15287318 TI - Osteoma of the zygomatic arch--report of a case. AB - An osteoma of the zygomatic arch in a 55-year-old woman is reported. Surgical resection through a preauricular incision was easily performed. The lesion was very large, but pedicled. PMID- 15287319 TI - Rhino-sinusitis related to endosseous implants extending into the nasal cavity. A case report. AB - Rhino-sinusitis may develop as a result of an altered airflow in the nasal cavity causing irritation of the nasal mucosa. A patient is presented who developed recurrent rhino-sinusitis complaints following placement of endosseous implants in the maxilla. Inspection of the nasal floor revealed that two implants had perforated the floor of the nasal cavity. The part of the implants protruding in the nasal cavity was surgically resected via an endonasal approach whereupon the rhino-sinusitis complaints disappeared. PMID- 15287320 TI - Medication use and misuse. PMID- 15287321 TI - New technology for medication adherence: electronically managed medication dispensing system. PMID- 15287322 TI - Perceived health promotion practice by older women: use of herbal products. AB - Women age 65 and older appear to use herbal products as well as multiple prescribed and non-prescribed drugs as part of their health-promoting practice. The purpose of this study was to identify changes in herbal-product use in a sample of older women during an 18-month period and to explore differences in locus of control and perceived health competence between herbal-product users and non-users. A longitudinal and descriptive study design was used. Instruments used to guide the two sets of structured interviews included the Multidimensional Health Locus of Control (MHLC), the Perceived Health Competence Scale (PHCS), and a questionnaire about herbal-product use. The prevalence of herbal-product usage and the number of products used remained consistent between the initial and follow-up interviews, with 43% of the participants using an average of 2.6 herbal products. Neither MHLC nor PHCS scores differed between herbal-product users and non-users. Mean scores of women in the study indicated higher internal than external locus of control. PMID- 15287323 TI - HIV and older adults: clinical outcomes in the era of HAART. AB - As the human immunodeficiency virus (HIV) epidemic enters its third decade, nurses are caring for increasing numbers of older adults with HIV who are on complicated medication regimens or highly active antiretroviral therapy (HAART). Although HAART has revolutionized HIV and acquired immunodeficiency syndrome (AIDS) care, little is known about how older adults respond to the new therapies. A review of the medical records of 19 older (> or = 50 years) and 18 younger (< 40 years) adults initiated on their first HAART regimen revealed both older and younger adults had similar positive clinical outcomes. Nurses need to individualize their care to patients of all ages rather than develop specific clinical guidelines for older adults with HIV. PMID- 15287324 TI - Multicultural medication adherence: a comparative study. AB - The purpose of this study was to evaluate the effect of several interventions on improving medication adherence among White, Black, and Hispanic older women. A total of 109 women older than age 65 who were participating in a clinical osteoporosis trial were recruited for this 12-month study examining medication adherence. After baseline medication adherence was assessed, participants underwent standardized teaching. Participants were contacted monthly by telephone and were seen in a clinic setting every 3 months. All participants used a pillbox for 6 months, and the minority women used an electronic monitoring bottle for 6 months. Adherence was highest in White women. Black women showed significant improvement in adherence at 9 and 12 months, and Hispanic women demonstrated a significant increase in adherence at 12 months. The use of electronic monitors had a positive effect on adherence for the minority women. PMID- 15287325 TI - Medication non-adherence among older adults: a review of strategies and interventions for improvement. AB - Medication non-adherence among older adults is a prevalent and costly problem; approximately one half have problems following their prescribed medication regimen, and more than 10% of hospital admissions are the result of medication non-adherence. In this literature review, medication non-adherence is defined and described among adults age 50 and older. Factors associated with medication non adherence are presented, interventions to improve medication non-adherence are discussed, and methods for assessing medication non-adherence are reviewed. In addition, nursing assessment and intervention to improve medication non-adherence are described. PMID- 15287326 TI - Warfarin therapy in older adults: managing treatment in the primary care setting. AB - Oral anticoagulant therapy with war farin is commonly used to prevent thromboembolic events in patients at risk. The degree of anticoagulation is variable among individuals and is influenced by many factors; therefore, patients must be monitored frequently to assess for potential adverse effects related to treatment. Individuals older than age 65 are at particular risk for thromboembolic events as well as anticoag ulant-related complications. Because of these factors, elderly individuals pose a unique challenge in maintain ing anticoagulant control. The purpose of this article is to revisit the role of warfarin therapy for elderly individ uals in the primary care setting and to provide nurse practitioners with the information necessary to prescribe and monitor this medication appropriately. This article provides indications for warfarin therapy and also identifies potential barriers to effective management with specific implications for the older population. PMID- 15287327 TI - Greater rigor for corporate compliance: amended sentencing guidelines tighten program requirements. PMID- 15287328 TI - Coder education: will demand, will deliver. PMID- 15287329 TI - Four options for implementing SNOMED. PMID- 15287330 TI - Verbal orders compliance and how we achieved it. PMID- 15287331 TI - An updated toolkit for security strategies. PMID- 15287332 TI - The EHR and the CCR: compatible or competitive? PMID- 15287333 TI - Kick starting the security risk analysis. PMID- 15287334 TI - A strategic approach to requesting capital. PMID- 15287335 TI - ICD-10 preparation checklist: part 2. PMID- 15287336 TI - Understanding new Medicare coverage determinations. PMID- 15287337 TI - A framework for human functioning--the ICF in Australia. PMID- 15287338 TI - Consultant contract: the view of the Hospital Consultants and Specialists Association. PMID- 15287339 TI - Consultant contract: the view of the British Medical Association. PMID- 15287340 TI - Unusual presentations of tuberculosis: let the clinician beware. PMID- 15287341 TI - New aspects of inner ear research. AB - At birth, one in 850 babies are profoundly deaf, and hearing loss affects more than 50% of all people over 60 years of age. While hearing loss caused by disease of the external and/or middle ear is treatable, hearing loss as a result of damage to and loss of hair cells and/or auditory neurons can only be alleviated using prosthetic devices such as hearings aids or cochlear implants. PMID- 15287342 TI - Antenatal diagnosis of fetal heart disease. AB - Antenatal diagnosis of congenital heart disease is most commonly made at the routine 20-week anomaly scan. Not all abnormalities can be detected by prenatal ultrasound but detection can be improved by obtaining outlet views and by the use of colour Doppler. This article provides an overview of the uses and limitations of fetal echocardiography. PMID- 15287343 TI - Increasing the use of orphan drugs in clinical practice. AB - Orphan drugs allow people with rare disease to access well-tolerated, safely formulated and effective drugs. Over the next few years, orphan drugs will probably be increasingly adopted into everyday clinical practice, partly because of genomic stratification. PMID- 15287344 TI - Population screening for lung cancer. AB - The feasibility of diagnosing small stage 1 lung cancers using low-dose chest computed tomography in asymptomatic at-risk individuals has been demonstrated in multiple studies. However, it has yet to be proved that the introduction of a chest computed tomography screening programme would do more good than harm at an acceptable cost. PMID- 15287345 TI - Cruciate ligament reconstruction. AB - Injuries to the cruciate ligaments of the knee can be disabling. Advances in treatment over recent years have made their early diagnosis imperative. Surgical reconstruction is not appropriate for all. Once surgical candidates have been identified a number of reconstructive options exist. PMID- 15287346 TI - Persistent pulmonary hypertension of the newborn. PMID- 15287347 TI - Latex allergy and otorhinolaryngological surgery. AB - Latex allergy in the otorhinolaryngology patient requires forward planning and coordination of management plans between the surgical, nursing and anaesthetic departments in order to ensure a safe outcome. PMID- 15287348 TI - Intravenous urography and imaging of the urinary tract. AB - This article briefly reviews the commonly used radiological investigations of the renal tract. The intravenous urogram (IVU) is a widely available imaging technique and one of the first-line investigations, yet is often poorly understood by junior medical teams. This article therefore discusses the IVU in detail, and reviews the common pathologies seen using this technique. PMID- 15287349 TI - Medical students' views about Modernising Medical Careers: a questionnaire survey of London students. AB - A survey of London medical students asked for their views of the changes to postgraduate medical education starting in August 2005. The majority had clear ideas about their career plans and did not want their career held back by the introduction of an extra year. They overwhelmingly preferred to start their early training in or reasonably near to where they had undertaken their medical studies. PMID- 15287350 TI - Chest wall tuberculosis involving the second rib in a young Ethiopian woman. PMID- 15287351 TI - Tuberculous meningitis in the developed world: a lurking menace. PMID- 15287352 TI - Tuberculous osteomyelitis: chasing the elusive tubercle. PMID- 15287353 TI - Training to recognize trainees in difficulty. PMID- 15287354 TI - Resistant ascites: a high price to pay. PMID- 15287355 TI - Should ketamine be used as a regular analgesic for patients with chronic pain? PMID- 15287356 TI - [Epidemiology and prognostic aspects of ankylosing spondylitis]. AB - The spondyloarthritides (SpA) comprise ankylosing spondylitis (AS), psoriatic SpA (PsSpA), reactive SpA (ReSpA), arthritis associated with chronic inflammatory bowel disease (SpAIBD) and undifferentiated SpA (uSpA). There are characteristic clinical features of SpA: inflammatory back pain (IBP), asymmetric peripheral arthritis, enthesitis, anterior uveitis, positive family history and others. The SpA, mainly AS, are strongly associated with HLA B27. AS is the most frequent and potentially most severe subtype, next to PsSpA. The prevalence of all SpA is rather high and not much different from rheumatoid arthritis (RA) and AS patients carry a burden of disease similar to RA patients. The prognosis of AS has not been extensively studied but some factors have been identified. There is a clear role for imaging modalities in the diagnosis of AS. Changes in the sacroiliac joint as detected by radiography still constitute the basis for the diagnosis of AS (New York criteria 1984). A diagnosis of sacroiliitis as made by magnetic resonance imaging (MRI) provides more objective evidence to a diagnosis of IBP arguing in favour of SpA which is defined on the basis of the ESSG criteria 1991 mainly on a clinical basis. Radiographic spinal changes such as syndesmophytes are important for the staging and outcome of AS. MR based assessment of spinal changes in are now being increasingly used to assess disease activity of AS patients. The presence of spinal radiographic changes at time of presentation was found to be the best predictor of further deterioration using the score modified SASSS' in a recent study. Other clinical features such as hip arthritis, early onset of disease, dactylitis, oligoarthritis, limitation of spinal mobility and poor efficacy of nonsteroidal antiinflammatory drugs were found to also have negative prognostic value. PMID- 15287357 TI - [Magnetic resonance tomography of sacroiliitis: anatomy, histological pathology, MR-morphology, and grading]. AB - The diagnosis of spondyloarthropathy is based on radiography of the sacroiliac joints, beside the patient's history and clinical examination. In cases where the clinical examination and radiography yield discrepant findings, contrast-enhanced magnetic resonance imaging (MRI) is a sensitive modality for the diagnosis of early sacroiliitis. Knowledge of the morphologic anatomy of the sacroiliac joints and of their abnormal micro- and macroanatomy in sacroiliitis and enthesitis are helpful for interpreting MR images. Arthritis of the sacroiliac joints is characterized by subchondral sclerosis, erosions, transarticular bone bridges, accumulation of periarticular fat, juxta-articular osteitis, synovtis, capsulitis, and enthesitis. The major histologic finding in active sacroiliitis is the presence of proliferative, pannus-like connective tissue destroying cartilage and bone. This tissue contains fibroblasts and fibrocytes as well as T cells and macrophages with a shift of the CD4/CD8 ratio toward the CD4 T helper cell population. The well-established grading of MRI findings by means of a chronicity and activity index, which are determined quantitatively from dynamic MR images, is supplemented by an alternative, semi-quantitative grading of activity. The following grades were defined for the short tau inversion recovery (STIR) sequence or the T1-weighted, fatsaturated spin-echo sequence for each quadrant (iliac anterior, iliac posterior, sacral anterior, sacral posterior): 0: no signal increase, 1: local increase in the joint cavity or within erosions, 2: small areas of increased juxta-articular signal, 3: moderate sized areas of increased juxta-articular signal, 4: large areas of increased juxta-articular signal. Values of the 4 quadrants are summed to an activity score (range 0-16). The new grading system is proposed to facilitate the examination and shorten image interpretation time. PMID- 15287358 TI - [Technique and radiation dose of conventional X-rays and computed tomography of the sacroiliac joint]. AB - Anterior-posterior (a.p.) or posterior-anterior X-rays of the sacroiliac joint, sometimes supplemented by a transverse view, have been the method of choice for diagnosis of patients suspected of having sacroiliitis. The sensitivity and specificity of conventional X-rays are relatively low, which can delay the diagnosis of sacroiliitis. Computed tomography (CT) is superior to conventional X rays for diagnosis of sacroiliitis, but does emit a relatively higher dose of radiation. For this reason, particularly for females, CT should be optimized by employing semi-coronal planes which require a lower radiation dose than axial planes. CT in a semi-coronal plane causes minimal radiation to the ovaries, and the effective radiation dose for women might even be lower than with conventional AP X-rays. Therefore, for suspected sacroiliitis in young women, CT in the semi coronal plane is the preferred imaging method with respect to diagnostics and radiation protection when magnetic resonance imaging (MRI) is not available. Male gonads can be protected from radiation doses in conventional X-rays, and CT as the primary imaging method can only be justified in these cases because of its better diagnostic capabilities. Due to the lack of inherent risk factors, MRI is superior to CT for diagnostics since it provides images of inflammatory signs in addition to joint destruction. Thus, when available, MRI should be given preference for diagnosis of sacroiliitis. PMID- 15287359 TI - [Grading sacroiliitis with emphasis on MRT-imaging]. AB - Cross-sectional imaging techniques play a decisive role in identification, localization, and characterization of alterations in the sacroiliac joint during the early stage of seronegative spondylarthropathy (SpA). Although several studies showed that the diagnostic capabilities of MRI and CT are superior to those of conventional radiography, they have not yet become established and accepted as methods for evaluating the grade of ankylosing spondylitis (AS) in contrast to conventional radiography. The lack of acceptance for MRI and/or CT methods for evaluating and grading changes in the sacroiliac joint makes it difficult to include the results of these procedures in classifying the grade of SpA. Moreover, grading the changes in the sacroilac joint in SpA with a method more sensitive than conventional radiography will be of prime importance in assessing treatment, e.g., the efficacy of new biological therapeutic agents directed against the tumor necrosis factor-alpha (TNF-alpha). An overview of the available grading methods is provided and MRI and CT techniques are presented. PMID- 15287360 TI - [Imaging of bacterial infections of the sacroiliac joint]. AB - Infection of the sacroiliac joint can be pyogenic or granulomatous and is usually unilateral. There are a number of predisposing conditions including drug abuse and intra articular steroid injection, but in 44% of cases, no definite predisposing factors can be identified. Considerable delay between presentation and diagnosis is recognized. The clinical picture may be non-specific and variable, and clinical suspicion may be low due to the relatively low incidence of the condition. This is compounded by difficulties in clinical examination of the SUs. The diagnosis is based on a history suggestive of infection, clinical or radiographic localization to the SUs, and a positive blood culture or joint aspirate. The pathology of pyogenic sacroiliitis is reviewed with respect to the anatomy of the SU, and the differential diagnoses considered. The imaging findings, and relative merits of all the modalities are discussed with particular consideration given to changes over the course of the disease. Imaging strategies are evaluated and proposed. As the commonest presenting symptom is low back pain, consideration should be given to the addition of a STIR sequence covering the SUs on all routine lumbar spine MR examinations. MR imaging is the most sensitive and specific imaging modality, while CT-guided arthrocentesis improves diagnostic confidence. Tc99MDP blood pool imaging mirrors the clinical features of resolution, and scintigraphy may be the best method to monitor response to treatment. Targeted antibiotic therapy usually leads to a full recovery. A high incidence of clinical suspicion, with MR imaging at an early stage are the essential prerequisites to an accurate diagnosis of bacterial sacroiliitis. PMID- 15287361 TI - [Minimal invasive stabilization of osteoporotic vertebral compression fractures. Methods and preinterventional diagnostics]. AB - PURPOSE: Minimal invasive stabilizations represent a new alternative for the treatment of osteoporotic compression fractures. Vertebroplasty and balloon kyphoplasty are two methods to enhance the strength of osteoporotic vertebral bodies by the means of cement application. MATERIAL AND METHODS: Vertebroplasty is the older and technically easier method. The balloon kyphoplasty is the newer and more expensive method which does not only improve pain but also restores the sagittal profile of the spine. RESULTS: By balloon kyphoplasty the height of 101 fractured vertebral bodies could be increased up to 90% and the wedge decreased from 12 to 7 degrees. Pain was reduced from 7.2 to 2.5 points. The Oswestry disability index decreased from 60 to 26 points. This effect persisted over a period of two years. Cement leakage occurred in only 2% of vertebral bodies. Fractures of adjacent vertebral bodies were found in 11%. CONCLUSION: Good preinterventional diagnostics and intraoperative imaging are necessary to make the balloon kyphoplasty a successful application. PMID- 15287362 TI - [Prolegomena of a system of ideas for medical radiology]. PMID- 15287363 TI - [Visual defects and nystagmus]. PMID- 15287364 TI - [What benefit has an MRT-screening]. PMID- 15287365 TI - [Contrast agents in MRT. Substance, effects, pharmacology and validity]. AB - The use of intravenous contrast agents in magnetic resonance imaging (MRI) has become well established clinical practice. Contrast agents provide additional information in many applications. Gadolinium chelates constitute the largest group of MR contrast agents and are considered to be safe. Different groups of contrast agents are established for clinical application: low concentrated gadolinium chelates, high concentrated gadolinium chelates, superparamagnetic iron oxide particles and hepatobiliary contrast agents. The review discusses the clinical applications and the safety issues involved with administration of intravenous contrast agents in MR imaging. Several approaches of intravascular or blood pool agents are also presented. PMID- 15287366 TI - Corneal epithelial cells and stromal keratocytes efficently produce CC chemokine ligand 20 (CCL20) and attract cells expressing its receptor CCR6 in mouse herpetic stromal keratitis. AB - PURPOSE: CC chemokine-ligand 20 (CCL20) is known to be selectively expressed by surface-lining mucosal epithelial cells and skin epidermal keratinocytes and to attract cells such as immature dendritic cells and effector T cells via CCR6. This study evaluated the ability of corneal epithelial cells and stromal keratocytes to produce CCL20 in vitro and in vivo. METHODS: Human corneal epithelial cells (HCE) and corneal keratocytes (HCK) were treated without or with various cytokines and expression of CCL20 mRNA and secreion of its protein were evaluated by RT-PCR and ELISA. Induction of CCL20 mRNA in HCE and HCK was also examined upon in vitro infection with HSV-1. Using a mouse model of herpetic stromal keratitis (HSK), induction of CCL20 expression and accumulation of cells expressing CCR6 were evaluated by RT-PCR and immunohistochemistry. RESULTS: Not only corneal epithelial cells but also stromal keratocytes efficiently expressed CCL20 mRNA and protein upon stimulation with IL-1beta and TNF-alpha. In vitro infection with HSV-1 also induced CCL20 mRNA in both types of cells. In a mouse herpetic stromal keratitis model, prominent accumulation of CCL20 and CCR6 mRNA was revealed in HSV-1-infected corneas. Furthermore, immunohistochemistry demonstrated production of CCL20 by corneal epithelial cells as well as stromal keratocytes and stromal infiltration of DEC205+ dendritic cells, CD4+ T cells and CD8+ T cells. Double staining revealed that CCR6-expressing cells were mostly MHC class II+ dendritic cells. CONCLUSIONS: Not only epithelial cells but also stromal keratocytes are efficient producers of CCL20 in the cornea and recruit CCR6-expressing cells such as dendritic cells into inflamed cornea. PMID- 15287367 TI - The in vitro and in vivo proliferative capacity of serum-free cultivated human conjunctival epithelial cells. AB - PURPOSE: To investigate the in vitro and in vivo proliferative capacity of human conjunctival epithelial cells cultured in serum-free media, and to compare this with current methods that utilize serum-containing media and 3T3 feeder layers. METHODS: Human conjunctival epithelial cells were cultivated in serum-free media alone, serum-free media with a 3T3 feeder layer, and serum-containing media with a 3T3 feeder layer. The areas of outgrowth, colony-forming efficiencies and number of population doublings were compared. The in vivo proliferative potential was assessed by analyzing the number of cells generated by the implantation of cultured cells into athymic mice. Cultured cells were evaluated for the expression of cytokeratins K3, K4, K12, K19, as well as the gel-forming goblet cell mucin, MUC5AC. RESULTS: Cells cultivated in serum-free media, serum-free media and feeder cells, and serum-containing media and feeder cells achieved colony-forming efficiencies of 14.5 +/- 4.1%, 10.1 +/- 3.1%, and 20.4 +/- 6.7%, respectively, and number of population doublings of 24.8 +/- 4.3, 14.8 +/- 3.6, and 30.0 +/- 5.0, respectively. Nine-day old athymic mice conjunctival cysts derived from serum-free cultures comprised 1.29 x 10(6) +/- 0.46 x 10(6) cells, while cysts derived from serum-containing cultures comprised 1.30 x 10(6) +/- 0.53 x 10(6) cells. The degree of epithelial stratification was similar in both conditions. Serum-free cultivated conjunctival cells retained their in vivo characteristics and expressed K4, K19 and MUC5AC. The presence of MUC5AC mRNA in these cells was confirmed by RT-PCR. CONCLUSIONS: Conjunctival epithelial cells propagated in serum-free media demonstrated a similar in vivo proliferative capability, as compared to serum-containing media with 3T3 feeder cells. This has important clinical implications, as the serum-free ex vivo expansion of cells for clinical transplantation overcomes the problems associated with the use of animal serum and cells. PMID- 15287368 TI - A novel porcine dry eye model system (pDEM) with simulated lacrimation/blinking system: preliminary findings on system variability and effect of corneal drying. AB - PURPOSE: To investigate inter- and intra-system variations and the effect of corneal drying using a recently developed pDEM. METHODS: pDEM was used to simulate "normal" and "dry eye" conditions using two "lacrimation-blink" intervals (20 s and 60 s). Corneas were examined/graded with sodium fluorescein before and after the experiment. At the end of each experiment, corneas were assessed by trypan blue exclusion technique. Two duplicated pDEM systems were set up and tested to investigate reproducibility. RESULTS: There was no significant difference in the results produced by the two pDEM systems. In the eyes under "normal" condition, there was no significant increase in the fluorescein grading. However, in eyes under "dry eye" condition, fluorescein staining increased and the number of non-viable cells in the central cornea increased. CONCLUSIONS: This novel pDEM system provides a useful assessment tool for the study of causative factors and new treatment strategies for dry eye syndrome. PMID- 15287369 TI - Alternative culture conditions for isolation and expansion of retinal progenitor cells. AB - PURPOSE: To investigate different in vitro model systems for retinal progenitor cell (RPC) isolation and expansion. METHODS: RPCs were isolated from embryonic day (E) 17 Long Evans rat retinas. Three different culture media: (1) modified serum free defined media (2) serum-containing media and (3) embryonic stem cell (ES)-conditioned media were used for RPC isolation and long term expansion. Expression of various cellular markers and cell morphologies were compared among the three culture systems at different passages by immunostaining and confocal microscopy. RESULTS: All three culture systems could maintain RPCs as nestin positive cells (78-87%) after long-term in vitro expansion. However, RPCs appeared to proliferate faster in the serum-free culture system. The ES conditioned media provided the best RPC survival. Cells appeared smaller at early passages compared with later passages. This morphology change occurred at P9-P10 in the serum-free medium, and at P5-P6 in the other two culture systems. CONCLUSIONS: The serum-free medium may be superior for preventing RPC differentiation during expansion. PMID- 15287370 TI - Quantitative comparison of fluoroquinolone therapies of experimental gram negative bacterial keratitis. AB - PURPOSE: To determine the effectiveness of topically applied fluoroquinolones for experimental Pseudomonas or Serratia keratitis. METHODS: Bacteria were injected intrastromally (10(3) colony forming units [CFU]). From 16 to 22 hours post infection (PI), a single topical drop of moxifloxacin (Vigamox, 0.545%), levofloxacin (Quixin, 0.5%), ofloxacin (Ocuflox, 0.3%) or ciprofloxacin (Ciloxan, 0.3%) was applied every 30 minutes. At 23 hours PI, corneas were cultured quantitatively. RESULTS: For Pseudomonas keratitis, untreated eyes contained 7 log CFU/cornea and antibiotic-treated eyes demonstrated a > or = 5-log reduction in CFU/cornea (p < or = 0.0001). Moxifloxacin, levofloxacin, or ciprofloxacin therapies were not significantly different from each other (p > or = 0.67). For Serratia keratitis, untreated eyes contained 7 logCFU/cornea whereas treated eyes had a > or = 2-log reduction (p < or = 0.0001). Moxifloxacin therapy proved most effective (p < or = 0.001). CONCLUSIONS: Overall, moxifloxacin was the most effective of the four fluoroquinolones in reducing CFU/cornea in the rabbit model of gram-negative keratitis. PMID- 15287371 TI - S-crystallin and arginine kinase bind F-actin in light- and dark-adapted octopus retinas. AB - PURPOSE: Rhabdomere microvilli dramatically reorganize in conditions of light and dark. This reorganization involves remodeling of the microvillus actin cytoskeleton. We are using the rhabdomeric retina of Octopus bimaculoides to identify actin-binding proteins that may be involved in this remodeling. METHODS: Octopus were light-/dark-adapted, retinas separated into dorsal and ventral halves, and homogenized. Actin-binding proteins were recognized using F-actin overlay blot assays and selected proteins from the overlays were identified using N-terminal sequencing methods or mass spectroscopy. Protein concentrations were quantified and compared by statistical analysis. RESULTS: Total protein gels of light-/dark-adapted, ventral/dorsal halves were almost identical except for a protein band at 26 kD. The relative amount of this protein in the dark was almost double that found in the light. The levels of other proteins did not vary significantly between the light and dark. F-actin overlays also showed matching patterns of actin-binding proteins except for the 26 kD protein. Although the 26 kD protein from light-adapted retinas transferred to the blotting membranes, it did not bind F-actin while the 26 kD protein on overlays from dark-adapted retinas always demonstrated F-actin binding. Besides the 26 kD protein, other proteins at 200 kD, 80 kD, 40 kD appeared on the overlays. These proteins and the 26 kD protein were sequenced and identified as hemocynanin, transitional ER ATPase, arginine kinase and S-crystallin, respectively. CONCLUSIONS: The amount of S-crystallin present in the octopus retina is significantly greater in dark adapted retinas and it binds to F-actin. In the light, the level of S-crystallin is greatly reduced and there is no apparent F-actin binding. No other studies, to our knowledge, show that S-crystallin binds to the actin cytoskeleton or that its expression is regulated by light. Arginine kinase may provide energy for cytoskeletal remodeling as it may in other neural tissues. PMID- 15287372 TI - Influence of exercise induced hyperlactatemia on retinal blood flow during normo- and hyperglycemia. AB - PURPOSE: Short term hyperglycemia has previously been shown to induce a blood flow increase in the retina. The mechanism behind this effect is poorly understood. We set out to investigate whether exercise-induced hyperlactatemia may alter the response of retinal blood flow to hyperglycemia. METHODS: We performed a randomized, controlled two-way cross over study comprising 12 healthy subjects, performed a 6-minutes period of dynamic exercise during an euglcaemic or hyperglycaemic insulin clamp. Retinal blood flow was assessed by combined vessel size measurement with the Zeiss retinal vessel analyzer and measurement of red blood cell velocities using bi-directional laser Doppler velocimetry. Retinal and systemic hemodynamic parameters were measured before, immediately after and 10 and 20 minutes after isometric exercise. RESULTS: On the euglycemic study day retinal blood flow increased after dynamic exercise. The maximum increase in retinal blood flow was observed 10 minutes after the end of exercise when lactate plasma concentration peaked. Hyperglycemia increased retinal blood flow under basal conditions, but had no incremental effect during exercise induced hyperlactatemia. CONCLUSIONS: Our results indicate that both lactate and glucose induce an increase in retinal blood flow in healthy humans. This may indicate a common pathway between glucose and lactate induced blood flow changes in the human retina. PMID- 15287373 TI - Inhibition of experimental choroidal neovascularization in rats by an alpha(v) integrin antagonist. AB - PURPOSE: Integrin alpha(v)beta3 is predominantly expressed on endothelial cells in choroidal neovascularization (CNV). We evaluated the efficacy of cyclic RGD (Arg-Gly-Asp) peptide, an alpha(v)-integrin antagonist, in a rat model of laser induced CNV METHODS: We evaluated the effect of cyclic RGD on the adhesion and cell viability of bovine choroidal endothelial cells (BCECs) by cell counting and the trypan blue dye exclusion test. CNV was induced by laser photocoagulation in female Long Evans rats (day 0), followed by intravitreal administration of one dose of cyclic RGD of 200 (n = 9), 100 (n = 10), 50 (n = 4), or 0 microg (n = 9) on days 9 and 11. We assessed the area of CNV by fluorescein angiography and the thickness microscopically on histologic sections. Neovascular vessels were detected by an antibody against factor VIII. RESULTS: Cyclic RGD (0.02 to 200 microg/ml) inhibited adhesion of BCECs in a dose-dependent manner without changing the cell viability (p < 0.01). In eyes treated with two injections of 200 or 100 microg of cyclic RGD peptide, the development of CNV was significantly (p < 0.01) inhibited in the area of leakage on fluorescein angiography. Histologically, the CNV membrane was observed beneath the retina and the factor VIII-positive cells and red blood cells were involved. The thickness of the lesions was significantly (p < 0.01) reduced in eyes that received 200 or 100 microg of RGD. CONCLUSIONS: Cyclic RGD effectively inhibited CNV progression in a rat model of laser-induced CNV, suggesting that this alpha(v)-integrin antagonist may be beneficial in the treatment of CNV. PMID- 15287374 TI - Arachidonic acid metabolites and peroxide-induced inhibition of [3H]D-aspartate release from bovine isolated retinae. AB - We have previously shown that hydrogen peroxide (H2O2) can inhibit K+ depolarization-evoked [3H]D-aspartate release from bovine isolated retinae. In the present study, we investigated the role of arachidonic acid metabolites in the inhibitory response elicited by H2O2 in the bovine retinae. Furthermore, we examined the direct effect of H2O2 on the production of prostaglandins and isoprostanes in this tissue. Isolated bovine retinae were prepared for studies of [3H]D-aspartate release using the Superfusion Method. Release of [3H]D-aspartate was elicited by Krebs solution containing an iso-osmotic concentration of KCl (50 mM). A direct action of H2O2 on prostaglandin E2 (PGE2) and 8-isoprostane F2alpha (8-iso-PGF2alpha) was also measured by enzyme-linked immunosorbant assay (ELISA). The cyclooxygenase inhibitor, flurbiprofen (3 microM), or the thromboxane receptor antagonist, SQ 29548 (10 microM) had no significant (p > 0.05) effect on K+-evoked [3H]D-aspartate release. On the other hand, both flurbiprofen (3 microM) and SQ 29548 (10 microM) blocked the inhibition of K+-evoked [3H]D aspartate induced by H2O2 (30 microM). In concentrations up to 100 microM, H2O2 caused an increase in PGE2 and 8-iso-PGF2alpha over basal levels. For instance, H2O2 (100 microM) increased PGE2 and 8-iso-PGF2alpha over basal levels by 348 +/- 41% and 185 +/- 26 (n = 4), respectively. We conclude that the peroxide-mediated inhibition of [3H]D-aspartate may involve the production of prostaglandins and isoprostanes in the bovine isolated retinae. PMID- 15287375 TI - Invasive pneumococcal disease in Greenland. AB - INTRODUCTION: The Inuit of Alaska and Canada have a higher incidence of invasive pneumococcal disease than non-inuit. The purpose of the present study was to evaluate incidence and outcome of these infections in Greenland with special reference to pneumococcal serotypes. STUDY DESIGN: Retrospective study of invasive pneumococcal infections in Greenland in the period 1996-2002. METHODS: Cases were defined as patients with positive cultures of Streptococcus pneumoniae from blood and/or CSF samples received at the microbiological laboratory of Dronning Ingrids Hospital Nuuk. Isolates were sent to Statens Serum Institut in Copenhagen for serotyping. Medical charts were reviewed. RESULTS: Fifty one cases were identified. The incidence among Inuit was 54 and among non-inuit 17 per 100,000 per year. Twenty one patients were in the age group 35-49 years and 20 in the age group 50-64 years. Twenty patients had meningitis (incidence 6 per 100,000/year). Seventeen patients died (33%). The most common serotypes were 1 (6 cases) and 12F (8 cases). The mortality rate was significantly higher among patients with 12F than among others (p < 0.01). No patients with serotype 1 died. CONCLUSION: As in Canada and Alaska the incidence of invasive pneumococcal disease, especially meningitis, is high among the Inuit in Greenland. Young and middle aged adults were most frequently affected. Pneumococcal serotype seems to be an important determinant for the outcome of invasive pneumococcal disease. PMID- 15287376 TI - Comparative study of community-acquired pneumonia caused by Streptococcus pneumoniae, Legionella pneumophila or Chlamydia pneumoniae. AB - The objective of this study was to compare epidemiological data and clinical presentation of community-acquired pneumonia (CAP) caused by Streptococcus pneumoniae, Legionella pneumophila or Chlamydia pneumoniae. From May 1994 to February 1996, 157 patients with S. pneumoniae (n = 68), L. pneumophila (n = 48) and C. pneumoniae (n = 41) pneumonia with definitive diagnosis, were prospectively studied. The following comparisons showed differences at a level of at least p < 0.05. Patients with S. pneumoniae pneumonia had more frequently underlying diseases (HIV infection and neoplasm) and those with C. pneumoniae pneumonia were older and had a higher frequency of chronic obstructive pulmonary disease (COPD), while L. pneumophila pneumonia prevailed in patients without comorbidity, but with alcohol intake. Presentation with cough and expectoration were significantly more frequent in patients with S. pneumoniae or C. pneumoniae pneumonia, while headache, diarrhoea and no response to betalactam antibiotics prevailed in L. pneumophila pneumonia. However, duration of symptoms > or = 7 d was more frequent in C. pneumoniae pneumonia. Patients with CAP caused by L. pneumophila presented hyponatraemia and an increase in CK more frequently, while AST elevation prevailed in L. pneumophila and C. pneumoniae pneumonia. In conclusion, some risk factors and clinical characteristics of patients with CAP may help to broaden empirical therapy against atypical pathogens until rapid diagnostic tests are available. PMID- 15287377 TI - Epidemiological and clinical characteristics of bacteraemia caused by Aeromonas spp. as compared with Escherichia coli and Pseudomonas aeruginosa. AB - We reviewed 75 episodes of Aeromonas spp. bacteraemia observed at our institution in 1979-2002, with special reference to episodes occurring in elderly patients (> or = 65 y) and to nosocomial episodes. In addition, we compared monomicrobial bacteraemic episodes caused by Aeromonas spp. (n = 54) with those caused by Escherichia coli (n = 108) and Pseudomonas aeruginosa (n = 108), to assess differences in clinical presentation and outcome. The 75 episodes of Aeromonas spp. bacteraemia occurred mainly in males (72%), suffering from chronic liver disease (36%) or neoplasm (33%). They had an abdominal origin in 52% of cases, were recorded as primary bacteraemia in 40%, and showed a 30-d case fatality rate of 36%. 22 episodes (29%) were nosocomial, 36 (48%) occurred in elderly patients and 21 (28%) were polymicrobial infections. In comparison with Aeromonas spp., E. coli bacteraemia was more often associated with less severe underlying conditions, a community-acquired infection (74%), and a urinary tract (52%) or abdominal (27%) origin and had a 30-d case fatality rate of 24%. P. aeruginosa bacteraemia occurred mainly in patients with severe underlying conditions, was associated with nosocomial infection (69%), and had a 30-d case fatality rate of 43%. In conclusion, Aeromonas spp. bacteraemia is a serious infection that should be considered in patients with chronic liver disease or neoplasm. It may occur in the elderly and as a nosocomial infection, and differs in clinical findings from bacteraemia due to other common pathogens. PMID- 15287379 TI - Tularaemia in Europe: an epidemiological overview. AB - Tularaemia exists endemically in most European countries. In some areas, such as Finland and Sweden, outbreaks comprising hundreds of cases are recorded at least once a decade. In other areas, outbreaks of such a magnitude occur only occasionally, except in times of war. Between outbreaks, the natural reservoir of the causative agent, Francisella tularensis, is unknown. The organism replicates intracellularly in protozoans. An association of tularaemia to natural water may be of significance in locating the reservoir. Epidemiological work has to date been slow, but is now facilitated by the development of new molecular methods. Due to a variation in numbers of short sequence-tandem repeats in the bacterial genome, individual strains of F. tularensis can today be distinguished. PMID- 15287378 TI - PCR identification and automated ribotyping of Pseudomonas aeruginosa clinical isolates from intensive care patients. AB - Nosocomial isolates of Pseudomonas aeruginosa exhibit high rates of resistance to antibiotics, and are often multidrug resistant. P. aeruginosa clinical isolates (n = 56) were obtained from ICU patients in a hospital in Pakistan over a 3-y period. Antimicrobial susceptibility of the 56 P. aeruginosa clinical isolates was investigated using 7 antibiotics and the resistance rates were as follows: aztreonam (68% resistant), ceftazidime (67%), imipenem (66%), ofloxacin (59%), amikacin (56%), gentamicin (44%), and piperacillin-tazobactam (27%) (p < 0.01). In addition, 55% of the P. aeruginosa clinical isolates were resistant to 4 or more antibiotics. Imipenem-resistant strains were frequently associated with ceftazidime, ofloxacin, aztreonam, and more strikingly, amikacin resistance (p < 0.05). PCR (using P. aeruginosa-specific primers VIC1 + VIC2 and P1 + P2, respectively) was highly specific and sensitive, and was positive for all 56 P. aeruginosa isolates tested. Automated ribotyping was used to investigate the clonal diversity of the 56 P. aeruginosa isolates. Automated ribotyping indicated that the clinical isolates were clonally related and could be clustered into 4 major ribogroups based on their similarity index, with ribogroup II being the dominant one. The P. aeruginosa isolates in ribogroup II were correlated with their antibiotic resistance pattern and, interestingly, there seemed to be a gradual acquisition of multiple antibiotic resistance associated with the isolates within this group over time. The ribotyping data, together with the antibiotic resistance profile, provide valuable molecular epidemiology information for the control of hospital-acquired P. aeruginosa infections. PMID- 15287380 TI - Identification of Seoul hantavirus in Rattus norvegicus in Indonesia. AB - The first genetic evidence for the presence of Seoul hantavirus (SEOV) in Indonesia is presented. Partial M segment sequence was recovered from the lung tissue of Rattus norvegicus trapped in central Jakarta. The sequence belongs to SEOV genotype and is most closely related to the strain B-1 from Japan. PMID- 15287381 TI - Demonstration of neutralizing mucosal IgA response to intranasal HIV-1 env DNA vaccines with or without the V3 glycosylation site. AB - HIV-1 env based DNA vaccines are generally found to be poor B-cell immunogens. We examined the role of an N-glycan located in the V3 loop of HIV-1 (N306) that is known to modulate the immunogenicity of gp120. Here we describe intranasal immunizations with env (HIV-1 BRU) based genetic immunogens in combination with subcutaneous boosts of recombinant gp160 (rgp160) in mice. Immunization with DNA alone resulted in detectable IgA responses to rgp160 in both faeces and bronchoalveolar lavage (BAL) fluid, but the additional boosting increased the faecal IgA titres only. Protein boosting was required for induction of faecal IgA antibodies capable of neutralizing a homologous laboratory strain and a subtype B primary isolate. The B-cell response towards V3 loop peptides was not only directed against the homologous subtype B but also against the subtype F. In contrast to our previous observations on IgG, there were no differences in anti gp160 IgA titres elicited by the N-glycan mutant and the wild-type immunogen. These results indicate that intranasal administration of plasmids containing env in combination with a subcutaneous boost proved to be an effective way of eliciting neutralizing mucosal IgA against HIV-1. PMID- 15287382 TI - Use of inflammatory markers for early detection of bacteraemia in patients with febrile neutropenia. AB - The aim of the study was to evaluate the ability of procalcitonin, C-reactive protein, serum amyloid A, interleukin-6 and interleukin-8 to predict bacteraemia during the 2 first d of fever in neutropenic patients. A total of 94 febrile neutropenic episodes in 60 patients were studied. Plasma samples were analysed at 10-h intervals from the onset of fever. Clinical events were categorized into 4 groups: 1) bacteraemia caused by other agents than coagulase-negative staphylococci (non-CNS bacteraemia) (n = 21), 2) coagulase-negative staphylococci bacteraemia (n = 15), 3) microbiologically or clinically documented infection without bacteraemia (n = 26) and 4) fever of unknown origin (n = 32). In non-CNS bacteraemia all markers, except for serum amyloid A, showed significantly higher levels compared to patients with fever of unknown origin (p < 0.05). For non-CNS bacteraemia the highest negative predictive value was found for procalcitonin (94%), followed by interleukin-6 (89%), C-reactive protein (88%) and interleukin 8 (87%). Procalcitonin, with a cut-off level of 1.4 ng/ml during 10-20 h after fever onset, showed the highest positive predictive value (67%) for a non-CNS bacteraemia. In conclusion, the value of the analysed markers to predict a non CNS bacteraemia in neutropenic patients was limited due to low sensitivity and positive predictive value. However, procalcitonin, interleukin-6, C-reactive protein, and interleukin-8 could give useful information for the clinician in excluding a non-CNS bacteraemia. PMID- 15287383 TI - A multicenter, open-label clinical study of micafungin (FK463) in the treatment of deep-seated mycosis in Japan. AB - The efficacy and safety of micafungin (FK463), which is a new lipopeptide antifungal agent of the echinocandin class and is active against both Aspergillus and Candida species, were investigated in patients with deep-seated mycosis in this study. 70 patients were treated with micafungin 12.5-150 mg/d intravenously for up to 56 d. The overall clinical response rates were 60% (6/10) in invasive pulmonary aspergillosis, 67% (6/9) in chronic necrotizing pulmonary aspergillosis, 55% (12/22) in pulmonary aspergilloma, 100% (6/6) in candidemia, and 71% (5/7) in esophageal candidiasis. The response rates for patients with prior antifungal treatment which was considered ineffective or toxic, were similar to rates for patients without prior treatment. Mycological eradication was observed in patients infected with Aspergillus fumigatus, Aspergillus flavus, Aspergillus terreus, Aspergillus niger, Candida albicans, Candida glabrata, or Candida krusei. Adverse events related to micafungin were reported in 21 patients (30%), and there was no dose-related occurrence of any adverse event. It is concluded that treatment with micafungin as monotherapy seems to be effective and safe in patients with deep-seated mycosis. PMID- 15287384 TI - Post-operative meningitis caused by drug-resistant Streptococcus pneumoniae: two case reports. AB - We report 2 patients with post-operative meningitis caused by drug-resistant Streptococcus pneumoniae (DRSP), following correction of frontoethmoidal encephalomeningocele in 1 patient and adenotonsillectomy in the other. Both patients responded well to vancomycin plus cefotaxime. DRSP may be colonized in the upper respiratory tract and causes serious infections after surgical operation. PMID- 15287385 TI - Brain abscess due to Nocardia otitidiscaviarum: report of a case and review. AB - We present a case of brain abscess caused by Nocardia otitidiscaviarum in an immunocompromized 44-y-old male. Only 7 other cases of N. otitidiscaviarum brain abscess or involvement were found in the literature. The mortality was 75% despite treatment among cases reviewed. There is a lack of therapeutic guidelines regarding brain abscesses due to Nocardia. PMID- 15287386 TI - Endobronchial actinomycosis secondary to a tooth aspiration. AB - We report a middle aged smoker with recurrent pneumonia caused by endobronchial actinomycosis secondary to a tooth aspiration. Unlike previously reported cases, our patient was not chronically debilitated. The case suggests that a follow-up bronchoscopy is beneficial after the initiation of antibiotic therapy for endobronchial actinomycosis. PMID- 15287387 TI - Bilateral facial palsy in a case of leptospirosis. AB - When returning from a 5-month trip to China, a 21-y-old Dutch male developed clinical signs, symptoms, and an antibody response compatible with leptospirosis. On d 15 of disease, he also developed facial palsy with a bilateral Bell's phenomenon. Facial palsy is a rare finding in leptospirosis, and if a causal relation exists, the delay of onset in the present case would suggest vasculitis rather than a direct neurotoxic effect. PMID- 15287388 TI - Fulminant endogene gas gangrene in a previously healthy male. AB - Spontaneous or non-traumatic gas gangrene is a rare condition. The present report refers to a previously healthy 57-y-old male who developed gas gangrene in the left lumbar region, left flank, left scapular, inguinal and suprapubic regions. Despite surgical, intensive care treatment, and antibiotic therapy, the patient died 32 h after the onset of the first symptoms. PMID- 15287389 TI - Severe metabolic acidosis and renal failure in an HIV-1 patient receiving tenofovir. AB - A 55-y-old male developed severe metabolic acidosis and renal failure during tenofovir therapy. Increased tenofovir exposure due to low body weight and chronic stable renal insufficiency could have enhanced the risk of nephrotoxicity, and creatinine clearance should be estimated before initiation of tenofovir. PMID- 15287390 TI - Painful neuropathy vasculitis in 2 patients with long-standing human immunodeficiency virus-1 infection. AB - We describe 2 most unusual cases of distal symmetrical painful polyneuropathy in patients with long-standing HIV-1 infection well controlled by HAART. Sural nerve biopsies revealed vasculitis in both cases and steroid therapy led to resolution of symptoms not influenced by analgesics and anti-inflammatory drugs. These unusual cases outline the importance of nerve biopsies in order to reach a diagnosis. PMID- 15287391 TI - Extrarespiratory Chlamydophila pneumoniae infection. AB - A series of 3 patients with prolonged fever due to Chlamydophila pneumoniae is reported. Laboratory data revealed hepatitis and various haemogram abnormalities. The main features of acute extrarespiratory infection caused by C. pneumoniae are discussed. PMID- 15287392 TI - Successful long-term application of highly purified natural interferon-alpha (multiferon) after preceding interferon approaches in a chronic hepatitis C patient with thrombocytopenia. AB - Treatment approaches with recombinant IFN-alpha2b and natural IFN-beta in a patient with chronic hepatitis C (genotype 1b) and cirrhosis had, in both cases, to be terminated prematurely due to breakthrough phenomena and thrombo leukocytopenia up to WHO grade 3. After the patient was switched to highly purified natural IFN-alpha (Multiferon) the thrombocyte and leukocyte counts increased significantly, and sustained complete biochemical and virological response could be achieved. PMID- 15287393 TI - Stenotrophomonas maltophilia bacteremia. PMID- 15287394 TI - Management of onychomycosis: examining the role of monotherapy and dual, triple, or quadruple therapies. AB - The high prevalence of onychomycosis warrants effective lasting treatment. Currently available monotherapeutic options in the United States include surgical or chemical nail avulsion/debridement, a topical antifungal nail lacquer, and systemic antifungal agents. Failure to respond to therapy and relapse rates of approximately 25% to 50% both point to the need for a shift in the approach to treating this chronic disease. In vitro data indicate synergistic and additive effects when combining certain antifungal agents, eg, ciclopirox and terbinafine. Clinical reports suggest that combining topical and oral antifungal agents (eg, ciclopirox nail lacquer and oral terbinafine), administered for a shortened duration compared with the standard regimen, may yield cure rates as good as, if not better than, the indicated oral monotherapy regimen. Drug penetration to different parts of the nail unit and complimentary modes of action may contribute to the success of combination therapy. PMID- 15287395 TI - Onychomycosis: review of recurrence rates, poor prognostic factors, and strategies to prevent disease recurrence. AB - Treatment of onychomycosis is associated with substantial disease reappearance rates. Identification of factors associated with therapeutic failure may help develop strategies to prevent recurrence of onychomycosis. Aspects of a patient's health and lifestyle, local factors involving the nail, therapeutic options, and environmental conditions are associated with poor therapeutic response. Strategies to reduce recurrence of disease involve the reduction of both relapse (delayed failure) and reinfection. The topical antifungal agent ciclopirox nail lacquer, may be a consideration for prophylaxis of this chronic disease. PMID- 15287396 TI - Onychomycosis: management and treatment. Proceedings of a clinical roundtable. PMID- 15287397 TI - Treatment of aggressive fibromatosis: a multidisciplinary approach. PMID- 15287398 TI - Autologous bone marrow injection in the management of simple bone cysts in children. AB - The majority of simple bone cysts (SBC's) is not symptomatic and remains undiagnosed or is discovered fortuitously. A number of simple bone cysts are only diagnosed after a pathological fracture which occurs as a presenting symptom. Fractures are managed either conservatively or surgically, based on criteria such as the age of the child and the type and localisation of the fracture. The risk for fracture can be evaluated radiographically. In the absence of a fracture risk, plain radiographic follow-up is sufficient. In case of a high fracture risk, percutaneous aspiration and injection of bone marrow may be performed. The result of this treatment in 21 simple bone cysts with a high risk for fracture is reported. Slow regression of the cyst and progressive healing were obtained in 15 cases (71.4%) whereas no response was noted in 3 cases (14.3%) and recurrence in another 3 (14.3%), after a mean follow-up of 37.1 months. Guidelines are proposed for the follow-up and management of SBC. PMID- 15287399 TI - Shoulder hemiarthroplasty in the management of humeral head fractures. AB - The results of hemiarthroplasty for shoulder fracture were evaluated in 26 patients, 20 women and 6 men with a mean age of 64.7 +/- 8.2 years. The follow-up period was 2 to 7 years. Cofield prostheses were used for the first 10 patients and subsequently 9 Global and 7 Aequalis prostheses were implanted, all cemented. The clinical outcome was assessed using the Constant-Murley scale. The mean score, at the last follow-up, was 70.4 +/- 16.4 (39-96). Mean forward elevation of the arm was 150 degrees (300-175 degrees), mean abduction was 145 degrees (30 degrees -170 degrees), and mean external rotation was 30 degrees (10 degrees-45 degrees). In most of the cases internal rotation corresponded with a position of the dorsum of the hand at the L3 vertebrae. The patients in our series achieved their optimal clinical result within the first 6 months after operation. Shoulder hemiarthroplasty is a worthwhile procedure, giving predictable results provided the patients have been carefully selected, the individual anatomy of the shoulder is restored and an aggressive rehabilitation program is implemented during the first six months after surgery. PMID- 15287400 TI - Grammont's reverse shoulder prosthesis for rotator cuff arthropathy. A retrospective study of 32 cases. AB - The authors have retrospectively studied a series of 32 reverse shoulder prostheses implanted in 30 patients by the same surgeon between 1992 and 2000. The mean age was 71 years; 26 patients were female, 4 were male. Thirteen patients (14 prostheses) were clinically and radiologically examined at follow up; 9 patients (10 prostheses) were questioned by telephone and their radiological records were studied. With a mean follow-up of 31 months, 92 % (22/24) were entirely satisfied with the operation. The mean Constant score for the 14 shoulders which were clinically evaluated at follow-up was 60/100. There was one failure, related with glenoid loosening. These findings are in line with the good short-term results previously reported with this prosthesis. Glenoid notching is a well-known problem with the reversed prosthesis; it may lead to implant failure. We noted such an image in 50 % of our patients. In several cases however, the radiological finding was more suggestive of osteophytic formation than of real bony erosion, an observation that has not been reported before. The image remained stable over time and did not lead to glenoid loosening within the time limits of the study. Nevertheless, notching remains a concern with respect to the long-term survival of these implants which should therefore, in our opinion, be used only in elderly patients. Whether improved technique or design modification can address this issue still has to be established. PMID- 15287401 TI - The Sauve-Kapandji procedure for posttraumatic disorders of the distal radioulnar joint. AB - The authors report their experience with the Sauve-Kapandji procedure for the management of posttraumatic disorders of the distal radioulnar joint in 20 patients. The mean age was 39 years (range, 19 to 62 years), the mean duration of follow-up was 76 months (range, 60-97 months) and the mean time interval between initial injury and the Sauve-Kapandji procedure was 24 months (range, 6-120 months). Postoperatively all patients experienced relief of pain. Rotation of the forearm increased to near normal values. The patients scored an average of 77 points on the Modified Mayo Wrist Score (range, 65-95 points). Three patients had an excellent result, six had a good result, seven had a fair result and one had a poor result. There were no major complications. Fifteen of seventeen employed patients had returned to work. Eighteen of nineteen patients were very satisfied or satisfied by the result of surgery. The procedure performed satisfactorily in addressing posttraumatic problems of the distal radioulnar joint, but must still be considered a salvage procedure. PMID- 15287402 TI - The use of primary total hip arthroplasty in university hospitals of the European Union. AB - Current practice in primary total hip replacement was investigated by postal survey in 125 university hospitals of the European Union (EU). Most hospitals (78.4%) use a hip register and implant cemented as well as uncemented stems (72.0%) and cups (68.8%). In Scandinavian & Anglo-Saxon countries, 42.9% of the departments implant cemented stems in all their patients, and 16.7% implant cemented cups in all their patients. In these countries, modern cementing techniques are commonly used and therapeutic choices are strongly influenced by hip registers. In Southern Europe, cemented cups have been abandoned in 31.1% and modern cementing techniques are less common. Benelux & Germanic countries have a practice in between. Three cemented (Exeter, Charnley, Lubinus) and three uncemented stems (Zweymuller, ABG, Bi-contact) represent 41.9% and 25.3% of stem types in use. Most departments (70.4%) have adopted alternative bearings. Ceramic ceramic and metal-metal are both used in almost half of the hospitals. Metal polyethylene has been abandoned in 15.2%. These trends are taught to new generations of surgeons in the EU and could become common practice in a near future. PMID- 15287403 TI - Prophylaxis for heterotopic ossification after primary total hip arthroplasty. A cohort study between indomethacin and meloxicam. AB - The authors have conducted a prospective cohort study of the efficacy of a 7 days administration of Indomethacin (n = 89) versus Meloxicam (n = 92), in the prophylaxis of heterotopic ossification (HO) in primary total hip arthroplasty. To assess the interobserver variability of the Brooker classification, all radiographs were evaluated by three investigators. In the Indomethacin group 25 patients developed HO (grade I: 22, grade III: 2 and grade IV: 1). In the Meloxicam group 34 developed HO (grade I: 30, grade II: 1 and grade III: 3). We were not able to show any difference between Indomethacin and Meloxicam in preventing heterotopic ossification after primary hip arthroplasty. We found a high interobserver variability in the grading system according to Brooker, in particular for the higher grades(grade II, III and IV). PMID- 15287404 TI - The potential benefit of thermal shrinkage for lax anterior cruciate ligaments. AB - Radiofrequency thermal shrinkage of the anterior cruciate ligament (ACL) injured knee is a relatively recent treatment. The purpose of this study was to retrospectively analyse the results in a cohort of 32 patients with a minimum follow-up of 12 months (range 12-37 months). Cases were acute (1) or chronic (31), involving either a native or reconstructed ligament. Native ACL injuries were: lax but intact (12 + 1 acute), partial tears with remaining intact femoral attachment (4) or completely torn and attached to the posterior cruciate ligament (PCL) (6). Reconstructions were: Hamstrings (3), Bone-Patella Tendon-Bone (5) or allograft (1). We also recorded concomitant pathology, function status, Lysholm and Tegner scores and KT2000 values. The procedure was successful in limiting instability overall in 72% (23/32), and in particular 44% of reconstructed ligaments (4/9). There was no correlation of results to the treatment of co existent pathology. The clinical scores remained high in successful cases although sporting function did reduce by follow-up. We conclude that despite these results the technique is still of value in appropriately selected, counselled and rehabilitated individuals. It is a safe technique that may avoid the necessity to proceed to reconstruction and can be considered as a planned procedure or as an adjunct to other therapy for instability at the time of arthroscopy for other pathology. PMID- 15287405 TI - Do thigh tourniquets contribute to the formation of intra-operative venous emboli? AB - The authors undertook a randomised prospective study to investigate the contribution of thigh tourniquets to the formation of intra-operative venous emboli during lower limb surgery. Patients were randomised to have a thigh tourniquet or no tourniquet and transoesophageal echocardiography was used to detect embolic signals in the right heart during and after knee arthroscopy. Three physicians blinded to patient demographics and tourniquet status separately assessed videotapes of the echocardiograms for evidence of emboli. Of the 32 patients randomised, 18 underwent knee arthroscopy with and 14 without tourniquet. Emboli were seen in 72% (95% CI 55 to 84) of patients, in 14 patients with tourniquet and in 9 patients without tourniquet. There was an estimated 13% greater incidence of emboli in the tourniquet group compared to the non tourniquet group, a difference which was not statistically significant (Fisher's Exact Test, p = 0.45). No patients suffered symptoms or signs attributable to a pulmonary embolus. PMID- 15287406 TI - Arthroscopic debridement of the osteoarthritic knee under local anaesthesia. AB - This prospective study compared the efficacy of arthroscopic debridement in osteoarthritic knees under local, general or peridural anaesthesia. Between 1997 and 2001, 201 arthroscopic debridements were performed in 197 patients (173 partial meniscectomies, 192 articular trimmings, 119 microfractures, 201 lavage procedures) in 197 patients. Patients were treated under local (Group "L", n = 67), general (Group "G", n = 65) or peridural anesthesia ( Group "P", n = 65). No tourniquet was used. The follow-up ranged from 24 to 72 months (mean: 32 months). No major complication was noted. Results were assessed according to the scale of Baumgaertner et al independently from the type of anaesthesia used (p = 0.71). Results were excellent in 85 cases (L: 30, G: 27, E: 28), good in 75 (L: 25, G: 24, E: 26), fair in 27 (L: 9, G: 8, E: 10), poor in 14 (L: 7, G: 4, E: 3). Arthroscopic debridement of the osteoarthritic knee under local anaesthesia appears as an efficient, simple, safe, painless and cost-effective method of treatment. PMID- 15287407 TI - Surgical repair of chronic rupture of the distal end of the biceps brachii. A modified anterior surgical repair technique. AB - The authors have used a modified surgical technique for repair of the distal end of the biceps brachii in three patients who presented with chronic rupture, all more than 6 weeks old. All patients were males; two lesions were on the dominant right side and one was on the non-dominant left side. An anterior incision was made over the cubital fossa, a hole was drilled over the radial tuberosity and a simple pull-through technique with an Ethibond suture was used to attach the tendon to an endobutton over the posterior surface of the radius. All patients returned to their employment and preinjury activity levels by six months. There were no postoperative complications and clinically all repairs remained intact. The patients regained their normal range of movements in 3 months; all but one regained the endurance and strength of their bicep in 6 months as assessed by Cybex testing. Surgical repair of the distal end of the biceps using the technique reported has given excellent results in these three patients. PMID- 15287408 TI - Atraumatic retrosternal dislocation of the clavicle. AB - Atraumatic retrosternal dislocation of the clavicle is an exceedingly rare event and three out of four previously reported cases lack any radiological evidence. We report the case of a 30-year-old male patient who presented an atraumatic retrosternal dislocation of the clavicle without a history of previous injury and underlying pathology. The diagnosis was delayed and established by a CT scan ten days after and initial presentation of the symptoms. A successful, stable, closed reduction under general anaesthesia was performed ten days after the initial presentation, having a cardiothoracic surgeon immediately available. There was no recurrence and the patient remains asymptomatic 18 months later. PMID- 15287409 TI - Isolated complex dislocation of the metacarpophalangeal joint of the third finger. AB - Two unusual cases of isolated closed complex dislocation of the metacarpophalangeal joint of the third finger are presented. The single most important element preventing reduction was interposition of the volar plate between the proximal end of the phalanx and the head of the metacarpal, but the deep transverse ligament was also intimately involved in the entrapment mechanism. Such dislocations require open reduction as in the two cases presented, and we found the dorsal approach to be simple and effective. PMID- 15287410 TI - Mycobacterium marinum causing tenosynovitis. 'Fish tank finger'. AB - Mycobacterium marinum is an unusual atypical mycobacterium with low pathogenicity for humans in comparison with Mycobacterium tuberculosis. Among the non tuberculous mycobacterial pathogens, Mycobacterium marinum is the most common pathogen to cause skin infections. Mycobacterium marinum infection causes chronic cutaneous lesions and in some cases deeper infections such as tenosynovitis, septic arthritis and rarely osteomyelitis. We report the case of a male patient presenting with tenosynovitis of the distal upper extremity secondary to Mycobacterium marinum infection. PMID- 15287411 TI - MRI abnormalities of the ischiopubic synchondrosis in children: a case report. AB - The authors report the case of a five and a half year-old-boy with symptomatic MRI abnormalities of the ischiopubic synchondrosis. The concept of "ischiopubic osteochondritis" is reviewed in the light of modern imaging, and the importance of its differentiation from different pathological entities such as osteomyelitis, tumour, fracture or other pathologic entities is recalled. PMID- 15287412 TI - Osteomyelitis of the accessory navicular bone in the foot. A case report. AB - The accessory navicular is one of the most symptomatic bones of the foot. Osteonecrosis and fracture of this bone have been previously described. We report a case of osteomyelitis of an accessory navicular bone in a young girl, to make treating clinicians aware of this rare possibility. PMID- 15287413 TI - Septic arthritis of a lumbar facet joint case report and review of the literature. AB - Septic arthritis of a lumbar facet joint (SALFJ) is a very rare condition. It has mostly been described in adults. Only one other paediatric case has been reported. We present a case of septic arthritis of the left L5-S1 lumbar facet joint, associated with epiduritis and paraspinal abscess, in an 8-year-old boy. Plain radiographs and Technetium bone scan were negative. The diagnosis was made by blood cultures, which isolated staphylococcus epidermidis, and by MRI. The child was treated successfully with antibiotics only. PMID- 15287414 TI - Parenting, family contexts, and personality characteristics in youngsters with VCFS. AB - Parenting, family contexts, and personality characteristics in youngsters with VCFS: The personality profiles for 48 youths with Velo-Cardio-Facial syndrome (VCFS) were described using the California Child Q-Set (CCQ). Associations between personality characteristics and parenting (i.e., Control and Warmth vs. Anger) and family contexts (i.e., Experienced Family Stress, Marital Conflict and Parental Consistency) were investigated. Personality characteristics were found to be related to parenting (in particular, Parental Warmth vs. Anger) but not to family contexts. Parents who reported more Parental Warmth (and less Anger) in interactions had children with higher Agreeableness, Conscientiousness and Emotional Stability and with lower Irritability and Dependency. Parental Control was positively related to children's Dependency and negatively to children's Conscientiousness. Compared to fathers, mothers exerted more Control. Differences in parenting and family contexts were related to the mode of inheritance but not to IQ, age, gender, and cardiac defects. Families in which a familial deletion occurred reported higher levels of Marital Conflict and lower Warmth in the parent-child interactions. PMID- 15287415 TI - A review of 35 cases of asymmetric crying facies. AB - A review of 35 cases of asymmetric crying facies: Congenital asymmetric crying facies (ACF) is caused by congenital hypoplasia or agenesis of the depressor anguli oris muscle (DAOM) on one side of the mouth. It is well known that this anomaly is frequently associated with cardiovascular, head and neck, musculoskeletal, respiratory, gastrointestinal, central nervous system, and genitourinary anomalies. In this article we report 35 ACF patients (28 children and 7 adults) and found additional abnormalities in 16 of them (i.e. 45%). The abnormalities were cerebral and cerebellar atrophy, mega-cisterna magna, mental motor retardation, convulsions, corpus callosum dysgenesis, cranial bone defect, dermoid cyst, spina bifida occulta, hypertelorism, micrognatia, retrognatia, hemangioma on the lower lip, short frenulum, cleft palate, low-set ears, preauricular tag, mild facial hypoplasia, sternal cleft, congenital heart defect, renal hypoplasia, vesicoureteral reflux, hypertrophic osteoarthropathy, congenital joint contractures, congenital hip dislocation, polydactyly, and umbilical and inguinal hernia. Besides these, one infant was born to a diabetic mother, and had atrial septal defect and the four other children had 4p deletion, Klinefelter syndrome, isolated CD4 deficiency and Treacher-Collins like facial appearance, respectively Although many of these abnormalities were reported in association with ACF, cerebellar atrophy, sternal cleft, cranial bone defect, infant of diabetic mother, 4p deletion, Klinefelter syndrome, isolated CD4 deficiency and Treacher-Collins like facial appearance were not previously published. PMID- 15287416 TI - Sensorineural deafness in two infants: a novel feature in the 22q distal duplication syndrome. Cardinal signs in trisomies 22 subtypes. AB - Sensorineural deafness in two infants: a novel feature in the 22q distal duplication syndrome. cardinal signs in trisomies 22 subtypes: Distal trisomy 22 has been described in more than 15 individuals. The features are severe mental and growth retardation, failure to thrive, congenital hypotonia, hydrocephalus, microcephaly, cleft palate, epicanthic folds, low-set ears, broad prominent nasal bridge, long philtrum, micrognathia, finger-like thumbs, cryptorchidism. We describe a girl deceased at the age of 12 years and an 11 year old boy, both with a duplication of distal 22q due to a parental pericentric inversion (22) (p13q12). Their phenotypes are compatible with distal trisomy of chromosome 22. However, they did not present cleft palate, but the survival of both patients permitted us to discover sensorineural deafness not previously reported in this chromosomal duplication. PMID- 15287417 TI - Familial cylindromatosis. AB - Familial cylindromatosis: we report a daughter with turban tumor and her mother with cylindromatosis. The dermal eccrine cylindroma arose as small, solitary lesions on the head of the mother when she was 28 years old. The following years other tumors became apparent. She was operated on several times. The first lesions appeared on the frontal part of the scalp of the daughter when she was 23 years old. Other tumors grew on the scalp. Histopathological examination of the excised tumors showed the same lesions in both the mother and the daughter: dermal eccrine cylindromata. Family history showed that the daughter's maternal aunt had a few tumors. Dermal eccrine cylindroma should be differentiated from malignant syndromes such as basal naevoid carcinoma or metastases and from neurofibromata. The gene of familial cylindromatosis was localised to chromosome 16q12-q13 and it was proposed that this gene is a tumor supressor gene. PMID- 15287418 TI - Maternal uniparental disomy 16 and genetic counseling: new case and survey of published cases. AB - Uniparental disomy (UPD) is the occurrence of both homologous chromosomes from one parent. Maternal UPD(16) is the most often reported UPD other than UPD(15); almost all cases are associated with confined placental mosaicism (CPM). Most of maternal UPD(16) cases are characterised by intrauterine growth retardation (IUGR) and different congenital malformations. Maternal UPD(16) has therefore been suspected to have clinical effects: however, the lack of uniqueness and specificity of the birth defects observed suggests that the phenotype may be related in parts to placental insufficiency. We report on a new case of maternal UPD(16) associated with low level trisomy 16 mosaicism in placenta and fetus. IUGR was noticed at 19 gestational weeks and the fetus died intrauterine. Apart from different craniofacial dysmorphisms she showed anal atresia. While IUGR is probably associated with trisomy 16 mosaicism, anal atresia is more characteristic for maternal UPD( 16). Considering the features in our patient as well as those in maternal UPD (16) cases from the literature, indications for UPD (16) testing can be defined: They include trisomy 16 mosaicism, IUGR and congenital anomalies (anal atresia, congenital heart defects). However, there is an overlap of clinical signs in mosaic trisomy 16 cases mosaic for maternal UPD(16) as opposed to those mosaic for biparental disomy 16. The management of trisomy 16 pregnancies should not differ from those in which maternal UPD(16) is confirmed. Therefore, a prenatal testing for UPD(16) is not useful, but it should be offered postnatally. The molecular genetic proof of maternal UPD(16) excludes an increased recurrence risk for the family for further pregnancies. PMID- 15287419 TI - Mesomelic form of chondrodysplasia and congenital glaucoma associated with de novo translocation (13;18)(q14;q23). AB - Mesomelic form of chondrodysplasia and congenital glaucoma associated with de novo translocation (13;18)(q14:q23): Mesomelic dysplasias are characterized by limb shortening most prominent of the middle segment of the extremities (forearm and lower leg). In addition to several syndromic forms a few patients with sporadic or familial forms and without precise nosological classification have been reported so far. In this report we present a young female with disproportionate mesomelic dwarfism, dysmorphic facial features, bilateral glaucoma, patent ductus arteriosus, low and hoarse voice, and generalized muscular hypotonia. At the age of 2.5 years mental development is normal. High resolution G-banded chromosome studies revealed a de novo reciprocal translocation with karyotype 46,XX t (13;18)(q14;q23). The concurrence of this de novo autosomal translocation with this distinct phenotype supports the hypothesis that disruption of (a) gene(s) at the translocation breakpoints causes this unusual, apparently new form of skeletal chondrodysplasia. PMID- 15287420 TI - Screening of Y chromosome microdeletion which contains AZF regions in 71 Turkish azoospermic men. AB - Screening of Y chromosome microdeletion which contains AZF regions in 71 turkish azoospermic men: In 71 Turkish men Y chromosome microdeletions have been studied before intracytoplasmic sperm injection (ICSI). DNA samples were amplified with 18 STS primers of the azoospermia factor (AZF) region on the Y chromosome by using multiplex polymerase chain reaction (PCR). Microdeletions were detected in 4 azoospermic men (5.6 %); one with a deletion in the AZFb region, while the 3 others had a large deletion extending over multiple chromosomal regions (AZFb+c+d and AZFa+b+c+d). In the patients with microdeletion, no spermatogenetic activity could be detected in testis biopsies. This result confirms the idea that Y chromosome microdeletion analysis is important in investigating the possibility of finding sperm in testicular sperm extraction (TESE). Therefore, we point out the importance of genetic testing and counselling regarding Y chromosome microdeletion for couples requesting ICSI. PMID- 15287421 TI - Fetal alcohol syndrome in association with Rett syndrome. AB - Fetal alcohol syndrome in association with RETT syndrome: We report on a girl with neonatal dystrophy, microcephaly, heart defect, and the characteristic features of alcohol embryopathy. Later, she developed distinctive features of RETT syndrome including loss of early acquired developmental skills and presented typical symptoms of RETT syndrome as reduction of communication skills, reduction of hand function, hyperventilation, and grinding of teeth. Molecular analysis of the MECP2 gene revealed the c.808T>C (R270X) mutation located in the nuclear localisation signal sequence of the gene. Our report highlights the importance of considering the diagnosis of RETT syndrome even in patients who are already suffering from a defined disease. PMID- 15287422 TI - Hereditary gingival fibromatosis and sensorineural hearing loss in a 42-year-old man with Jones syndrome. AB - Hereditary gingival fibromatosis and sensorineural hearing loss in a 42-year-old man with Jones syndrome: Gingival fibromatosis is a rare disease, which can be seen as an isolated condition or associated with some uncommon syndromes. This case report describes the evaluation and treatment of a 42-year-old male patient with hereditary gingival fibromatosis, sensorineural hearing loss, undescended testis and maxillary odontogenic cyst (Jones Syndrome). Six years follow up of the index patient after the surgery revealed no recurrence of the gingival fibromatosis. This report also describes the anamnestic data of the patient's family that showed progressive deafness and gingival enlargement in three generations. PMID- 15287423 TI - Neonatal Marfan syndrome caused by an exon 25 mutation of the fibrillin-1 gene. AB - Neonatal Marfan syndrome caused by an exon 25 mutation of the Fibrillin-1 gene: We describe a male infant with severe arachnodactyly, hypermobility of the fingers, flexion contractures of elbows, wrists, hips, and knees, microretrognathia, crumpled ears, rockerbottom feet, loose redundant skin, and lens dislocations. Cardiac valve insufficiency and aortic dilatation resulted in cardiac failure, decompensated with digitalisation and death occurred at the age of 4 months. This case represents the severe end of the clinical spectrum of Marfan syndrome, namely neonatal Marfan syndrome. Molecular diagnostic analyses confirmed a de novo exon 25 mutation in the FBN1 gene. PMID- 15287424 TI - Pectus excavatum and situs inversus totalis: a new combination or a coincidence? PMID- 15287425 TI - Cleidocranial dysplasia with new additional findings. PMID- 15287426 TI - Micropenis in a newborn with acrocallosal syndrome. PMID- 15287427 TI - Primary body stalk anomaly in a first trimester fetus. PMID- 15287428 TI - The nuts and bolts of homeopathy. PMID- 15287429 TI - Homeopathy and dental caries: implications for dental practice and veterinary research. PMID- 15287430 TI - Repertory and likelihood ratio: time for structural changes. AB - If the likelihood ratio (LR) method is introduced, the repertory will gradually change as more symptoms are assessed. It will also change the use of the repertory: the most important medicines of each symptom rubric can be identified and relied on, even in large rubrics. This is also a good opportunity to correct structural shortcomings of the repertory, for instance, entries should be based on systematic analysis of materia medica instead of casual observations. PMID- 15287431 TI - How do homeopaths make decisions? An exploratory study of inter-rater reliability and intuition in the decision making process. AB - The validity of clinical decision making in homeopathy is largely unexplored and little is understood about the process or its reliability. This exploratory study investigated, in the context of a questionnaire based re-proving of Belladonna 30c, the extent to which decisions are based on clinical facts or intuition and how reliable decisions are. Three experienced, independent homeopathic clinicians/proving researchers rated the symptom diaries of the 206 subjects taking part. They reported their proving decision (ie positive proving response, no proving response or undecided) based on the total symptom profiles and rated (on a scale of 0-10) their use of clinical facts or intuition. Keynote symptoms and overall confidence scores were also reported. The level of agreement between raters was generally poor (weighted kappa 0.349-0.064). All raters used both facts and intuition. The rater's reliance on the facts was significantly associated with classifying those subjects who had no proving response [rater 1, P<0.001; rater 2, P<0.001]. Raters used significantly higher intuition scores when classifying a prover [rater 2, P= 0.001; rater 3, P= 0.012]. Issues regarding the education and practice of homeopathy are discussed. PMID- 15287432 TI - Homeopathically prepared dilution of Rana catesbeiana thyroid glands modifies its rate of metamorphosis. AB - One strand of research on the scientific basis of homeopathy is based on inversion effects of dilutions and the biophysical properties of information transfer. A model developed by Endler, was the basis for the study of the influence of high-diluted solution (1:1026 part by weight) of thyroid glands on the rate of metamorphosis of the frog Rana catesbeiana from the no legged to four legged stage. The glands were obtained from tadpoles and prepared according by (dilution and succussion). Similar pure hydroalcoholic solution (unsuccussed) was used as control. In order to identify significant differences in the frequencies of four-legged tadpoles, in homeopathic and control group, we used a chi-square goodness-of-fit test (P<0.01) and the cumulative risk for metamorphosis by Cox's Proportional Hazards model (P<0.05). The number of animals that reached the four legged stage is generally smaller in the treated group, than in the hydroalcoholic control group. It was postulated that thyroid hormones transmitted information' specific to the molecules used to prepare the solution, even though the molarity was beyond Avogadro's number. PMID- 15287433 TI - The effect of fluorine and homeopathic medicines in rats fed cariogenic diet. AB - Although some sectors of dentistry have benefited from technological advances, dental caries is still a major problem. Prevention and treatment of dental caries by fluorine is considered a major advance in public health. Nevertheless fluorosis, caused by ingestion of excessive amounts of fluorine during the period of teeth formation, is of great concern. In accordance with the homeopathic doctrine, minimum doses of fluorine and other substances could prevent and/or treat caries. In this experiment, we compared the preventive action of fluorine and evaluated the effect of homeopathic medicines on the teeth of rats fed a cariogenic diet. None of the groups included in this study developed caries. However, microscopy revealed the presence of precipitate and/or deposit in the groups treated with homeopathic medicines. This phenomenon might be due to deposit in the dental surface or precipitation of bacterial plaque or calcium salts. It was not possible to identify the composition of the deposit/precipitate due for technical reasons. In one of the groups treated with homeopathic medicines fur loss was observed in 40% of animals. These reactions might be caused due to the action of the homeopathic medicines. PMID- 15287434 TI - Permanent physico-chemical properties of extremely diluted aqueous solutions of homeopathic medicines. AB - The purpose of this study was to obtain information about the influence of successive dilutions and succussions on the water structure. 'Extremely diluted solutions' (EDS) are solutions obtained through the iteration of two processes: dilution in stages of 1:100 and succussion, typically used in homeopathic medicine. The iteration is repeated until extreme dilutions are reached, so that the chemical composition of the solution is identical to that of the solvent. Nine different preparations, were studied from the 3cH to 30cH (Hahnemannian Centesimal Dilution). Four of those were without the active principle (potentized water). Two different active principles were used: Arsenicum sulphuratum rubrum (ASR), As4S4, 2,4-dichlorophenoxyacetic acid (2,4D). The solvents were: a solution of sodium bicarbonate and of silicic acid at 5 x 10(-5) M (mol/l) each, and solutions of sodium bicarbonate 5 x 10(-5), 7.5 x 10(-5) and 10 x 10(-5) M (mol/l) in double-distilled water. The containers were Pyrex glass to avoid the release of alkaline oxide and silica from the walls. Conductivity measurements of the solutions were carried out as a function of the age of the potencies. We found increases of electrical conductivity compared to untreated solvent. Successive dilution and succussion can permanently alter the physico-chemical properties of the aqueous solvent. But we also detected changes in physio chemical parameters with time. This has not previously been reported. The modification of the solvent could provide an important support to the validity of homeopathic medicine, that employs 'medicines without molecules'. The nature of the phenomena here described remains still unexplained, nevertheless some significant experimental results were obtained. PMID- 15287435 TI - The structure of the Organon. AB - The Organon of Medicine is the seminal text of Homeopathy. However, its grammar and structure make it obtuse and remote to both new students and veterans. We propose a demarcation of the Organon into sections, exposing the didactic structure of the Organon, and display this demarcation in concise graphic form. It is hoped that this representation will improve accessibility and understanding of the Organon for readers at all levels. PMID- 15287436 TI - Patient-practitioner-remedy (PPR) entanglement. Part 6. Miasms revisited: non linear quantum theory as a model for the homeopathic process. AB - The possibility that non-linear quantum theory could be used to model PPR entanglement is discussed in relation to the treatment of miasms. In this model, miasms are imagined as disease entities behaving like solitary waves, or 'solitons' which, when trapped in a therapeutic state space, requiring equally soliton-like (miasmatic or high potency) remedies to effectively 'annihilate' them. PMID- 15287437 TI - A landmark for basic research in homeopathy. PMID- 15287438 TI - The imaginer and the imagined. AB - A transference of the imaginer and the imagined, arising from largely unconscious fantasies of the way parent and child interact to construct a view of reality, is present in all analyses. For narcissistic patients, primitive fantasies of the imaginer and imagined form an enduring organization, and the enactment of these fantasies in transference and countertransference distorts the way analyst and patient construct meaning. Clinical material demonstrates the deepening that occurs when these fantasies are interpreted. PMID- 15287439 TI - The analyst's fantasy of the ideal patient. AB - Using detailed clinical vignettes, the author illustrates how the analyst's fantasy of the ideal patient can be used to advance an analysis at the same time as it fuels mutual resistances. The author suggests that all analysts carry with them a fantasy of the ideal patient that varies from analyst to analyst and from school to school. Such fantasies are often related to images of an ideal free associative process. They are for the most part descriptively unconscious, becoming conscious only when prompted by the clinical moment. As such, they are part of a countertransference, broadly defined, that is responsive to both the analyst's and the patient's conflictual life. PMID- 15287440 TI - The psychoanalyst as individual: self-analysis and gradients of functioning. AB - The authors discuss the position of the analyst as an individual and the idea that his mental functioning can be seen as a meaningful element of the analytic field. The first part of the article shows the importance of the analyst's self analysis, with particular attention to periods when the analyst is facing a difficult time, self-analysis in supervision, and the exploration of transgenerational influences. The authors go on to discuss the many gradients of the analyst's mental functioning, and these are mirrored in the patient's text, an indication of attunement. PMID- 15287441 TI - One form of self-analysis. AB - Beginning with Freud, psychoanalysts have discovered media through which they may achieve a self-analytic experience (for example, by use of dreams, fantasies, reveries, memories, and even visual images). Each of these media is a kind of "fiction" created by the analyst that provides an imaginative space where he or she may gain access to unconscious life. The author demonstrates how a generative self-analytic experience may be accomplished through the medium of psychoanalytic writing: a fictional autobiographical form of writing through which a self analytic experience is created that has much in common with the analytic experience created by the analyst and analysand. PMID- 15287442 TI - Haunted by parents--a literary example of change meaning loss: Edna St. Vincent Millay. AB - The author uses the life and personal history of the poet Edna St. Vincent Millay, as revealed through her diaries and letters and in biographies, to illustrate a particular type of psychoanalytic patient. Such patients are resistant to change, particularly when it involves letting go of the internalization of early parental figures. Although some of these patients fail to achieve successful analytic outcomes, Millay is an example of someone with similar circumstances who nevertheless made significant creative contributions. PMID- 15287443 TI - Introduction to Jean-Jacques Blevis's "remains to be transmitted: Primo Levi's traumatic dream". PMID- 15287444 TI - Remains to be transmitted: Primo Levi's traumatic dream. AB - Drawing on the writings of Primo Levi and the psychoanalysis of Jacques Lacan, the author attempts to conceive psychic trauma as a coalescence of traumas, since this is perhaps the only way to prevent a subject from being forced back into identification with the catastrophic event, whatever that may have been. A recurrent dream of Primo Levi's suggests to the author the way that traumas may have coalesced within Levi. The hope would be to restore the entire significance of what remains from that traumatic event to the speech (parole) of the Other, to the speech of every human, even the most helpless, bruised, or destroyed among us. PMID- 15287445 TI - Three psychoanalytic sessions. PMID- 15287446 TI - Endoscopy-assisted craniosynostosis. PMID- 15287447 TI - Endoscopy-assisted wide-vertex craniectomy, barrel stave osteotomies, and postoperative helmet molding therapy in the management of sagittal suture craniosynostosis. AB - OBJECT: Endoscopic techniques were introduced 7 years ago for the surgical management of patients with sagittal synostosis. In this study of 139 patients with sagittal synostosis, the authors assessed the efficacy, safety, complications, and outcomes after performing endoscopy-assisted wide-vertex craniectomies with bitemporal and biparietal barrel stave osteotomies. METHODS: The sample population consisted of a total of 99 boys and 40 girls who ranged in age from 0.4 to 9.2 months (mean 3.6 months). Two small incisions were made near the lambda and vertex. Using endoscopic visualization, wide-vertex craniectomies with bilateral temporal and parietal barrel stave osteotomies were performed. Postoperative treatment included custom-made surlyn cranial orthotic devices for cranial reshaping and maintenance. The mean craniectomy width was 5.4 cm and the length was 10 cm. The overall blood transfusion rate was 9% (two intraoperative and 12 postoperative transfusions). The mean estimated blood loss was 29 ml (range 5-150 ml). The mean preoperative hematocrit was 32%, whereas the postoperative level was 27%. One hundred thirty-two patients were discharged the morning following surgery. The majority of patients did not experience facial swelling, and none suffered postoperative fevers. Anthropometric cephalic index measurements indicated that excellent results were obtained in 87% of the patients (cephalic index > 75); good results in 8.7% (cephalic index 70-75); and poor results in 4.3% (cephalic index > 70). There were no cases of intraoperative death, infection, hemorrhage, or venous sinus injury. CONCLUSIONS: Analysis of the results indicates that use of the aforedescribed procedure in the early treatment of infants with sagittal synostosis provides excellent outcomes and that the morbidity rate is lower than that associated with traditional cranial vault reconstruction. Detailed anthropometric and radiographic analyses demonstrated that with adequate helmet therapy in our patients normocephaly was achieved and maintained without the need for secondary operations. PMID- 15287448 TI - Advances in the endoscopic management of suprasellar arachnoid cysts in children. AB - OBJECT: Suprasellar arachnoid cysts present unique management problems. The authors retrospectively reviewed six cases, in which endoscopic ventriculocystocisternotomy was performed, to identify specific neuroimaging features that aid both the accurate diagnosis of this entity and the postoperative assessment of fenestration patency. METHODS: Six consecutive children underwent treatment for suprasellar arachnoid cysts. Consistent radiographic features in all cases were identified. Through a single entry site, endoscopic fenestration was performed at both the apical and basal cyst membranes. Outcome was assessed using clinical examination, quantitative changes in cyst size, and triplanar magnetic resonance (MR) imaging with flow-sensitive (long TR) sequences. In every case, the suprasellar cysts displayed three diagnostic MR imaging features: 1) vertical displacement of the optic chiasm/tracts; 2) upward deflection of the rostral mesencephalon and mammillary bodies; and 3) effacement of the ventral pons. Two patients initially underwent placement of a ventriculoperitoneal shunt before the cysts were recognized, but MR images obtained after shunt placement revealed the cysts. In a mean follow-up period of 26.2 months, all patients improved clinically. Postoperative imaging revealed a mean cyst volume decrease of 52.7% and a return to more normal suprasellar and prepontine anatomy. Flow-sensitive MR imaging confirmed pulsation artifact at all 12 fenestration sites. There was no surgery-related death and no additional cerebrospinal fluid diversion procedure was required. CONCLUSIONS: To aid in the accurate diagnosis of prepontine arachnoid cysts, the authors identified several pathognomonic features on sagittal MR images: vertical deflection of the optic chiasm and mammillary bodies, as well as pontine effacement. Dual endoscopic fenestration into the intraventricular compartment and basal cistern is safe, and it effectively provides symptomatic relief by decreasing the cyst size. Triplanar flow-sensitive MR imaging sequences can confirm fenestration patency without the need for cine-mode MR imaging. PMID- 15287449 TI - Endoscopic third ventriculostomy in children younger than 1 year of age. AB - OBJECT: The authors analyzed data obtained in 36 patients younger than 1 year of age who were treated with endoscopic third ventriculostomy (ETV) for obstructive hydrocephalus at their institution. METHODS: There were 17 boys and 19 girls who ranged in age from 3 days to 11 months (mean 4.7 months). The causes of the hydrocephalus were Chiari Type II malformation (11 cases), aqueductal stenosis (11 cases), and other (14 cases). The success rate was 64% (p < 0.05, confidence interval 0.48-0.8) and there were four complications, mainly meningitis. The follow-up period ranged from 22 to 69 months (mean 47.4 months). CONCLUSIONS: Based on the 64% success rate in children younger than 1 year of age, ETV should be the treatment of choice for obstructive hydrocephalus in this age group, although larger studies involving specific causes of hydrocephalus are needed. PMID- 15287450 TI - Neuroendoscopic findings in patients with intracranial germinomas correlating with diabetes insipidus. AB - OBJECT: Intracranial germinomas commonly occur in the pineal region, the floor of the third ventricle (hypothalamus), or both, and they are often associated with diabetes insipidus (DI). The authors conducted a study to correlate preoperative DI with the endoscopic and magnetic resonance (MR) imaging evidence of tumor on the third ventricle floor. METHODS: The authors reviewed hospital records, office charts, and MR imaging studies obtained in patients in whom a biopsy sampling procedure was performed with or without endoscopic third ventriculostomy (ETV) at Children's Hospital, Birmingham, Alabama between May 1998 and July 2002. Ten patients with the pathological diagnosis of pure germinoma were identified. Preoperative MR imaging findings and presenting symptoms were correlated with intraoperative neuroendoscopic findings. Seven patients presented with symptomatic hydrocephalus and underwent concomitant ETV. Six patients presented with DI and MR imaging evidence of involvement of the third ventricle floor. Two patients presented with DI and no initial MR imaging evidence of neoplastic involvement of the third ventricle floor; in both there was endoscopic evidence of neoplastic involvement of the floor of the third ventricle. In two children without DI, neither MR imaging nor endoscopic evidence of involvement of the third ventricle floor was observed. CONCLUSIONS: In the authors' experience with intracranial germinoma, endoscopic tumor biopsy sampling, and ETV provide an effective, safe, and minimally invasive means of obtaining diagnostic tissue and treating any concomitant hydrocephalus. The authors found that preoperative DI is an absolute predictor of neoplastic involvement of the hypothalamus regardless of MR imaging findings. Therefore, in the setting of DI and intracranial germinoma without neuroimaging evidence of hypothalamic involvement, germinomatous involvement of the hypothalamus should be assumed present, if not confirmed endoscopically at the time of biopsy sampling or third ventriculostomy, when devising adjuvant treatment plans for such patients. PMID- 15287451 TI - Initial endoscopic management of pineal region tumors and associated hydrocephalus: clinical series and literature review. AB - OBJECT: The authors report their experience in six patients with pineal tumors and associated hydrocephalus who underwent an endoscopic biopsy procedure and third ventriculocisternostomy (ETVC) in a single sitting. METHODS: The ETVC was successfully performed without complication in all patients; however, a ventriculoperitoneal shunt was eventually required in four. Histological diagnosis was successfully established in four patients. The authors also reviewed the literature to assess reports involving ETVC and tumor biopsy sampling in patients with pineal tumors and hydrocephalus. A total of 54 cases, including those in this study, have been reported. Fifteen percent of the patients eventually required placement of a ventricular shunt. The transient complication rate was 15% with no death. A positive tissue diagnosis was established in 89% of the cases overall. CONCLUSIONS: The authors conclude that the endoscopic management of patients with pineal region masses and hydrocephalus may be a preferred initial strategy. PMID- 15287452 TI - Predictors of death in pediatric patients requiring cerebrospinal fluid shunts. AB - OBJECT: Despite improved therapeutic strategies and better diagnostic techniques in the management of pediatric hydrocephalus there continues to be a significant mortality rate associated with cerebrospinal fluid (CSF) shunts. The goal of this study was to determine the long-term outcome and predictors of death in these patients. METHODS: Data were collected in all patients requiring a CSF shunt presenting to a single tertiary care pediatric institution during a 10-year period. Patients with neoplasms were excluded because their deaths were predominantly related to the tumor. Descriptive statistics were obtained on the patient characteristics, surgical features, and shunt characteristics. The time and cause of death were determined. Kaplan-Meier survival estimates were used to determine overall survival of patients. Univariate analysis was performed using the log-rank test. Multivariate analysis included use of Cox regression model to determine the significance of age (at the time of initial shunt insertion), the number of shunt-related failures and infections, and whether the shunts were complex or multiple in nature in predicting death. Hazard ratios, 95% confidence intervals and probability values were calculated. Of 907 patients, 124 died. The most common causes were myelomeningocele (191 cases), intraventricular hemorrhage (114 cases), and tumor (190 cases) with 7.9, 3.5, and 32.6% dying, respectively, during the study period. Restricting all analyses to cases without neoplasms, the incidence of shunt-related failures was 58.1% in patients who died and 55.3% in those who survived, with an incidence of shunt-related infection of 19.4% in the former and 18.5% in the latter. The overall mortality rates in all patients at 1, 5, and 10 years were 4.5, 8.9, and 12.4%, respectively, from time of initial shunt insertion to death or last follow-up visit. The infection rate per procedure (that is, following the first shunt insertion) was 10.9% (78 of 717 cases). Evaluation of predictors of death revealed a statistically significant effect of infection with a hazard ratio of 1.66 (p = 0.04). CONCLUSIONS: The mortality rate in shunt-treated pediatric patients with hydrocephalus remains high, dependent on the underlying reason for CSF shunt insertion and the subsequent development of infection of the shunt apparatus. PMID- 15287453 TI - Pathophysiological changes in cerebrovascular distensibility in patients undergoing chronic shunt therapy. AB - OBJECT: Patients undergoing long-term shunt therapy following shunt malfunction often present with acute neurological deterioration, high intracranial pressure (ICP), and yet small or slit ventricles. It is believed that low brain compliance prevents ventricle enlargement in such cases. To elucidate the underlying pathophysiology, the authors estimated compliance as a function of cerebrovascular distensibility in 45 patients undergoing chronic shunt therapy. METHODS: The ICP and pressure-volume index (PVI) were measured at end-tidal CO2 of 30 mm Hg (PVI30) and 40 mm Hg (PVI40). The ventricle volume was dichotomized as slit/small/normal or dilated based on the frontooccipital horn ratio. In 18 patients PVI30 was normal (18.4 +/- 4 ml), whereas in 27 patients it was significantly elevated (45.5 +/- 14 ml). Clinical symptoms or ventricle size at presentation did not correlate with the PVI30. The ICP and PVI at end-tidal CO2 of 40 mm Hg were significantly higher than those at end-tidal CO2 of 30 mm Hg (p < 0.001 and < 0.02, respectively) suggesting an increased cerebrovascular distensibility. CONCLUSIONS: The authors did not observe a low compliance in patients undergoing chronic shunt therapy who, at shunt malfunction, presented with a slit/small/normal ventricle; however, analysis of the findings strongly indicated that an increased cerebrovascular distensibility was present in these patients. This may explain the high ICP and acute clinical deterioration following shunt malfunction in such cases. PMID- 15287454 TI - A pilot trial comparing cerebral perfusion pressure-targeted therapy to intracranial pressure-targeted therapy in children with severe traumatic brain injury. AB - OBJECT: The authors sought to compare cerebral perfusion pressure (CPP)- with intracranial pressure (ICP)-targeted therapy in children with severe traumatic brain injury (TBI). METHODS: A randomized controlled trial was developed to assess CPP and ICP therapies in 17 children (range 15 months-15 years of age) with poststabilization Glasgow Coma Scale (GCS) scores of less than or equal to 8 who were admitted to a pediatric intensive care unit at a Level I trauma center. Goals in the ICP group were to maintain ICP lower than 20 mm Hg and CPP higher than 50 mm Hg. In the CPP group, goals were to maintain CPP higher than 70 mm Hg for patients at least 2 years old and higher than 60 mm Hg for patients younger than 2 years of age. The study outcomes were death or functional outcome at 1 year postinjury. The median GCS scores in the CPP group (12 patients) and the ICP group (five patients) were 6 and 7, respectively. In the CPP group, two patients died, one was lost to follow up, four were unimpaired, and five had mild impairment. In the ICP group, all patients survived; one was lost to follow up, two had mild impairment, and two had hemiparesis and moderate impairment. There were four unimpaired survivors in the CPP arm compared with none in the ICP arm (p = 0.08). CONCLUSIONS: The CPP method appears to be safe, although this feasibility study does not establish that the CPP therapy is superior to ICP therapy. PMID- 15287455 TI - Persistent syringomyelia following pediatric Chiari I decompression: radiological and surgical findings. AB - OBJECT: The incidence of syringomyelia in patients with Chiari I malformation is reported to be between 20 and 75%. In the series reported herein 10.6% of patients with both clinical entities have continued to have a significant syrinx following their first decompressive procedure. All but one patient had resolution of their syrinx after a second posterior fossa operation. The authors have analyzed this smaller group of patients to look for possible radiological or surgical findings that may aid in predicting which patients are less likely to respond to the initial first decompressive procedure. METHODS: The authors retrospectively reviewed radiological and operative data in eight patients who continued to have syringomyelia following a decompressive procedure. Seven (88%) of these patients had complete resolution of their syrinx following a second operation. At repeated operation, obstruction at the foramen of Magendie was seen in six patients. In one patient in whom the dura was not opened during first operation, the second operation revealed an arachnoid veil that occluded the foramen of Magendie. CONCLUSIONS: No single radiological measurement was found to aid in the prediction of which patients would not respond to the first decompressive procedure. Furthermore, no operative finding was extraordinarily unique to any single patient. All but one patient in whom confirmation of a patent foramen of Magendie was made at repeated operation-that is, lysing of arachnoid veils, stent placement, unilateral tonsillar coagulation-had resolution of their syringomyelia. Surgical reexploration should be considered in cases of persistent syringomyelia. PMID- 15287456 TI - Arachnoid veils and the Chiari I malformation. AB - OBJECT: The literature contains scant data regarding variations in anatomy at the level of the foramen of Magendie in patients with Chiari I malformation and syringomyelia. METHODS: Based on their operative experience and hospital data, the authors detailed the incidence of arachnoid veils found in juxtaposition to the foramen of Magendie in patients with hindbrain herniation. Additionally, radiological studies were retrospectively reviewed in cases in which such an anomaly was noted intraoperatively. Of 140 patients with Chiari I malformation who underwent decompressive surgery, an associated syrinx was demonstrated in 80 (57%). The foramen of Magendie was obstructed by an arachnoid veil in 10 (12.5%) of these patients; once the lesion was punctured, the cerebrospinal fluid drained freely from this median aperture. On retrospective review of imaging studies, none of these anomalous structures was evident. In all patients with an arachnoid veil and syringomyelia resolution of syringomyelia was revealed on postoperative imaging. CONCLUSIONS: In the absence of a clear pathophysiology of syrinx production, the authors would recommend that patients with syringomyelia and Chiari I malformation undergo duraplasty so that, if present, these veils can be fenestrated. PMID- 15287457 TI - Feasibility and advisability of resections of thalamic tumors in pediatric patients. AB - OBJECT: The author conducted a retrospective review of 19 cases in which he resected thalamic tumors between 1986 and 2001. METHODS: The median age of the children was 8 years. Five different routes were used to resect the tumors; two tumors were resected via two approaches. The extent of resection was evaluated by postoperative imaging. Gross-total resections were performed in six cases and resections of greater than 90% of the tumor were conducted in 10 others; 90% or more of the tumor was resected in 84% of the cases. Seven tumors were low-grade gliomas and 12 were high grade. There was one postoperative death, and two children suffered permanent morbidity. Five of seven children with low-grade tumors are alive 2 to 12 years postoperatively; three of 11 children with high grade tumors are alive at 2, 3, and 16 years, respectively. CONCLUSIONS: Treatment of pediatric thalamic tumors must be individualized, with consideration given to the tumor's location, enhancement, and margins as well as the published data about the correlation between extent of resection and prognosis. PMID- 15287458 TI - Intracranial midline dermoid and epidermoid cysts in children. AB - OBJECT: Dermoid and epidermoid cysts are rare space-occupying lesions of the central nervous system. Although characterized by a slow growth rate, they are often associated with serious complications. Surgery is the only effective treatment, and radical resection of the entire cyst, whenever possible, generally succeeds in achieving a cure. Authors of large series have only occasionally reported on pediatric cases, and these reports often lack a specific analysis of those cerebral midline lesions. METHODS: The authors report on the treatment of 19 patients (16 with intracranial intradural dermoid and three with epidermoid cysts located along the cerebral midline). All patients underwent surgery at the Catholic University Medical School in Rome. The patients ranged in age from 3 months to 16 years. Nine cysts were located in the posterior cranial fossa, six in the frontobasal subarachnoid spaces, two in the third ventricle, and two in the quadrigeminal plate cistern. In the cases presenting with dermal sinus tracts, attempts at resecting the dermoid cyst and the associated dermal sinus were made in a single stage to achieve an en bloc removal. In the cases without dermal sinus tracts, and in the three with epidermoid cysts, a standard craniotomy was performed to reach the lesion. There were no surgery-related deaths, and the morbidity rate was low. Total and subtotal resections were achieved in 15 and four cases, respectively; however, regrowth of the residual tumor was observed only in two of them. After repeated resection, tumor progression has not been observed. At a mean follow-up period of 92.7 months, excellent clinical results were achieved in 18 patients. CONCLUSIONS: Because surgery is the only effective treatment modality for these lesions, radical resection should be performed in all cases to avoid tumor recurrence; however, because the cyst capsule can adhere firmly to vital structures and attempts at its radical removal can be dangerous, subtotal resection may be a wise option in selected cases. PMID- 15287459 TI - Krev1 interaction trapped-1/cerebral cavernous malformation-1 protein expression during early angiogenesis. AB - OBJECT: Molecular genetic studies of cerebral cavernous malformation (CCM) have identified three loci, CCM1-3, that can lead to CCM when mutated. Examination of the CCM1 locus established KRIT1 (Krev1 Interaction Trapped genre 1) as the CCM1 gene. Despite the identification of KRIT1 as the gene mutated in CCM1, little has been learned regarding its function. The authors recently demonstrated specific KRIT1 expression in endothelial cells. Based on this result and the fact that the CCM phenotype features defects in microvasculature, we hypothesized that KRIT1 may take an active part in normal angiogenesis. METHODS: In this study, the authors investigated the spatial and temporal expression of KRIT1 during normal vessel development and maturation by examining KRIT1 protein in both in vitro and in vivo angiogenic systems with the use of postconfluent endothelial cell cultures along with placental tissues from different developmental stages. CONCLUSIONS: The results demonstrate that KRIT1 is expressed during capillary like tube formation in the early stages of angiogenesis in vitro. Histological examination of placental tissue, a well-established in vivo model of angiogenesis, shows KRIT1 expression in active angiogenic and vasculogenic areas of the immature placental villi. As the placenta matures, KRIT1 expression is restricted to microvascular and small arterial endothelial cells with little or no expression seen in the intima of large vessels. It can therefore be concluded that KRIT1 is expressed during early angiogenesis by endothelial cells and may play a key role in vessel formation and/or development. PMID- 15287460 TI - Radiation-induced atypical meningioma with rapid growth in a 13-year-old girl. Case report. AB - This case illustrates the potential growth rate of an atypical meningioma in a pediatric patient, emphasizes one of the potential risks after therapeutic radiation, and underscores the importance of clinical evaluation and follow up of the symptomatic patient after tumor resection and radiation therapy. The authors report a case of a radiation-induced atypical meningioma of the olfactory groove in a 13-year-old girl who received 36 Gy of radiation to the craniospinal axis and 72 Gy to the primary site of a primitive neuroectodermal epithelial tumor of the left parietooccipital lobe when she was 4 years of age. This tumor was not present on routine magnetic resonance imaging performed 13 months prior to the discovery of this lesion or on computerized tomography scanning obtained 6 months prior to the discovery of this tumor. At the time of its discovery, the tumor was 5 x 5 x 4 cm. This tumor was resected and the patient's symptoms improved. This case illustrates the importance of continued close follow up after cranial irradiation in the pediatric population. PMID- 15287461 TI - Transient local response and persistent tumor control in a child with recurrent malignant glioma: treatment with combination therapy including dendritic cell therapy. Case report. AB - Treatment of malignant glioma is difficult and discouraging. Even after resection and maximal adjuvant therapy, the prognosis remains poor. The authors sought a novel form of treatment, such as stimulating the patient's own immune response against the tumor, and developed a protocol of tumor vaccination in which autologous dendritic cells (DCs) were used in patients with recurrent malignant glioma. A 4-year-old girl was treated by means of biopsy sampling and radiotherapy for a rolandic low-grade glioma. Ten years later, a Grade III recurrence was discovered and treated with subtotal resection, interstitial radiation, six courses of oral temozolomide, and 12 courses of oral VP 16. At the end of the chemotherapy cycle, a new rapidly growing recurrence was diagnosed. A macroscopically complete resection was performed. Afterward, the girl was vaccinated with autologous DCs that had been pulsed ex vivo with the homogenate of the resection specimen. She received six vaccines in total. The efficacy of immunization was checked by a positive delayed-type hypersensitivity skin reaction after the second injection. After the fifth vaccine, a transient contrast enhancement without mass effect was visualized on magnetic resonance imaging. Simultaneously, positron emission tomography imaging revealed a transient increase of metabolic activity around the resection cavity, but the metabolic uptake ratio remained below 1.8. The patient's disease is still in complete remission 24 months after the last surgery. She is clinically well with minor and stable left hemiparesis. This case report illustrates the potential of vaccination with DCs loaded with crude tumor homogenate as adjuvant therapy to induce prolonged tumor control of malignant glioma and the objective noninvasively monitored immune response against infiltrating tumor cells. PMID- 15287462 TI - Subependymal giant cell astrocytoma with cranial and spinal metastases in a patient with tuberous sclerosis. Case report. AB - This 17-year-old male patient with tuberous sclerosis developed increased headaches and lethargy. Magnetic resonance imaging of the brain revealed increased ventricle size and increased size of a subependymal giant cell astrocytoma at the foramen of Monro, as well as spinal cord metastases of giant cell tumors. Decompressive surgery of the foramen of Monro lesion resulted in temporary resolution of the hydrocephalus. Increased Ki-67 labeling of tumor as well as rare spinal enhancement both possibly indicated malignant features for this entity. PMID- 15287463 TI - Neurocutaneous melanosis associated with Dandy-Walker malformation and a meningohydroencephalocele. Case report. AB - Neurocutaneous melanosis and Dandy-Walker malformation are both forms of rare congenital neurodysplasia. Interestingly, 8 to 10% of patients with neurocutaneous melanosis also harbor an associated Dandy-Walker malformation, indicating that these developmental abnormalities share a common origin. The authors describe a case of neurocutaneous melanosis associated with Dandy-Walker malformation and an occipital meningohydroencephalocele with a giant melanotic nevus. Multiple congenital liver masses were also observed in the infant. The occipital nevus was totally excised, and ventriculoperitoneal and cyst-peritoneal shunts were created to prevent subsequent hydrocephalus. Findings in this case support the possibility that excessive melanocytes hinder normal mesenchymal development, causing Dandy-Walker malformation and an occipital meningocele. PMID- 15287464 TI - Endoscopic disconnection for hypothalamic hamartoma with intractable seizure. Report of four cases. AB - Although intractable epilepsy associated with hypothalamic hamartoma (HH) can be controlled by microsurgical resection of the lesion, excision of deep-seated lesions is often associated with morbidity and mortality. Endoscopic disconnection is less invasive and seems to be well suited for this indication. The authors discuss the role of endoscopic-assisted surgery in the management of HH-induced seizures. Four patients with HH-related intractable gelastic seizure underwent endoscopic disconnection surgery. Postoperatively, all patients exhibited improvement. Two patients became seizure free immediately after endoscopic disconnection surgery, one patient with a widespread seizure focus involving the motor strip continued to experience rare complex partial seizures but gelastic seizures ceased, and one experienced a reduced frequency of seizures but persistence of some generalized seizures. Three patients suffered postoperative disconnection-like syndrome, which continued 3 to 7 days and spontaneously disappeared. The authors advocate the endoscopic disconnection surgery as a safe and effective treatment for HH-related epilepsy by blocking the spread of epileptic discharges from the lesion. PMID- 15287465 TI - Further characterization of traumatic subdural collections of infancy. Report of five cases. AB - Bilateral convexity and interhemispheric subdural hematomas are common neuroimaging patterns seen in infants who have sustained nonaccidental head injuries (NAHIs). These collections often appear aschronic or acute-on-chronic on computerized tomography (CT) studies. To determine the nature of these extraaxial fluid collections and their relationship to cerebrospinal fluid (CSF) dynamics, the authors studied five infants with suspected NAHI in whom symptomatic bilateral mixed- or low-density subdural collections were revealed on imaging studies; the patients underwent burr hole evacuation of the hematoma and external drainage. Once decompression was achieved, radiotracer was injected into the lumbar subarachnoid space, and the subdural drainage system was monitored for appearance of the isotope. In all five cases, the radiotracer moved rapidly from the lumbar subarachnoid space into the convexity subdural space and then into the external drainage system. This indicated the possibility that some of these mixed density subdural collections were acute blood mixed with CSF rather than acute-on chronic collections arising from rebleeding subdural membranes. The authors propose that, during infancy, tears in the loosely adherent arachnoid envelope at the main arachnoid granulation site along the superior sagittal sinus may result in a considerable amount of CSF mixing with acute blood in the subdural space, creating a hematohygroma. PMID- 15287466 TI - Craniopagus: second Brisbane case. Case report. AB - Craniopagus is a rare and intriguing condition with an incidence of one in 2.5 million births. The chance of a neurosurgeon seeing a case in a working lifetime is unlikely. The chances of two cases from the same community within 12 months are remote in the extreme. The authors present a second case of craniopagus born and separated in Brisbane, Australia, in 2001 and discuss the intricacies of surgical separation and the lessons learned. PMID- 15287467 TI - Juvenile pilocytic astrocytoma presenting with subarachnoid hemorrhage. Case report and review of the literature. AB - The authors report a case of a 13-year-old boy with juvenile pilocytic astrocytoma (JPA) presenting with subarachnoid hemorrhage (SAH). The patient experienced sudden onset of headache, vomiting, and loss of consciousness. Cranial computerized tomography scanning revealed blood within basal cisterns and the third ventricle. Angiography demonstrated normal cerebral vasculature and upward displacement of the bilateral A, segments of the anterior cerebral artery. Magnetic resonance (MR) imaging revealed a chiasmatic/hypothalamic mass with evidence of hemorrhage. The mass was surgically decompressed. Histopathological examination showed evidence of JPA. In all cases of SAH in which there is blood around the third ventricle and a raised A1 segment on angiography, MR imaging should be performed. The presence of a normal sella turcica, as well as indistinct margins between the tumor and the opticochiasmatic apparatus should raise suspicion about the lesion. PMID- 15287468 TI - Bone growth causing ventriculoperitoneal shunt malfunction in a patient with osteopetrosis. Case report. AB - Osteopetrosis is an inherited skeletal condition of defective osteoclastic resorption of bone resulting in increased bone density. Osseous changes occur most severely at the base of the skull. Important clinical symptoms include cranial nerve palsies due to uni- or bilateral obliteration of cranial nerve foramina including deafness, facial paralysis, and optic nerve compression. Thickening of the skull and progressive diminution of the cranial capacity may lead to elevation of intracranial pressure, papilledema, and hydrocephalus. The authors present an unusual case of a patient with osteopetrosis in whom a ventriculoperitoneal shunt became obstructed at its exit from the skull by the developing bone mass. To the best of their knowledge, this has not been described. PMID- 15287469 TI - Vertebral osteoid osteoma associated with paravertebral soft-tissue changes on magnetic resonance imaging. Report of two cases. AB - Soft-tissue changes associated with osteoid osteoma have been described in the digits of the hands and feet as well as the long bones. Only six cases in which such changes occurred in the spine have been reported. Magnetic resonance (MR) imaging facilitates the determination of such changes. Establishing a diagnosis, however, is especially difficult in spinal osteoid osteoma when using MR imaging. Therefore, osteoid osteoma-related soft-tissue changes demonstrated on MR imaging raise the question of malignancy and may lead to unnecessary long-term treatment or biopsy sampling. The authors report two cases of spinal osteoid osteoma in which paravertebral soft-tissue changes were observed on MR imaging to mimic malignant soft-tissue tumors. PMID- 15287470 TI - Cranial index of symmetry: an objective semiautomated measure of plagiocephaly. Technical note. AB - OBJECT: The prevalence of deformational, or positional, plagiocephaly has increased during the last decade. Treatments vary among centers, ranging from expectant management to orthotic helmet therapy to craniofacial remodeling. This management variability is partially due to a lack of objective methods with which to measure the severity of plagiocephaly, as well as procedures that are not cumbersome or require radiographic studies. A simple and objective method of determining the degree of cranial deformation has been developed. METHODS: A headband placed around the head was marked with two adjustable points--one denoting the nasion and the other, the inion. A digital camera was used to image the head from a vertex view. The shape of the headband and the area of each hemisphere were then determined by analyzing the image on a personal computer in a semiautomated fashion. A cranial index of symmetry (CIS) was calculated and, by definition, equaled 100% for a perfectly symmetrical head. In this preliminary study, the authors studied eight children referred for evaluation of their plagiocephaly and eight infants referred for noncraniofacial entities. In the unaffected infants the mean CIS was 96.3 +/- 1.3% (+/- standard deviation). In children with clinical evidence of plagiocephaly, however, the CIS was 81.9 +/- 3.4% (p < 0.001). Although the CIS in healthy children was never less than 95%, that in all infants with plagiocephaly was below 90%. CONCLUSIONS: Although preliminary, this objective nonradiographic measurement of cranial symmetry appears to allow grading of the severity of positional plagiocephaly. The aforementioned methodology may potentially be used as an unbiased means of comparing different treatment modalities in longitudinal studies. PMID- 15287471 TI - Moyamoya. PMID- 15287472 TI - [Community-associated methicillin-resistant Staphylococcus aureus: current status and molecular epidemiological perspective]. AB - MRSA has been a major causative agent of nosocomial infection. However, recently MRSA has become increasingly isolated from community-associated infections. We summarized here up to date information about community-associated MRSA (C-MRSA) infections and characteristics of C-MRSA strains based on molecular analysis. By using the SCCmec typing, strong evidence was provided for the independent derivation of healthcare-associated MRSA and C-MRSA clones. PMID- 15287473 TI - [Influenza C virus isolated in Hiroshima Prefecture during the 1999/2000 winter season--a clinical and epidemiological study]. AB - Influenza C virus (Inf. C) is one of pathogens of human respiratory tract infection and prevalent throughout the world at an early stage in life. However, Inf. C has been isolated only accidentally and there have been few reports on its clinical and epidemiological features. From November 1999 to March 2000, Inf. C was isolated from clinical specimens (throat swabs) of 4 pediataric patients with respiratory tract illness at Hiroshima Prefectural Hospital and was isolated in 4 peditaric patients at the other medical institutions in Hiroshima prefecture. There were no differences in clinical features including duration of illness, duration of fever, maximum body temperature between 4 patients with Inf. C infection and patients with influenza A (H1N1 and H3N2) and influenza B infection from 1992 to 2000. We investigated geographical distribution of patients with inf. C infection and analyzed for antigenic characteristics with a set of monoclonal antibodies against hemagglutinin-esterase glycoproteins. The data suggested that at least two antigenically different Inf. C prevalented in a region during winter from 1999 to 2000. PMID- 15287474 TI - [Isolation of influenza A H1N2 virus from a returning traveller at Nagoya International Airport]. AB - A reassortant influenza A H1N2 virus was isolated from a returning traveller arriving at Nagoya International Airport, Japan from Indonesia in May, 2002. A Hemagglutination inhibition test revealed that the virus was similar to a vaccine strain of A/NewCaledonia/20/99. A phylogenetic analysis demonstrated that the virus forms a cluster with other influenza A H1N2 viruses isolated in other countries. The reassortment event was theoretically assumed to have occurred between the 1999/2000 and 2000/2001 influenza seasons. Neither A H1N2 nor A H3N1 virus was detected from 256 isolates of AH1 or 177 of AH3 influenza viruses isolated in Aichi Prefecture, Japan between the 1999/2000 and 2001/2002 influenza seasons. This finding suggests the importance of influenza surveillance at an airport quarantine office to detect promptly a novel influenza virus penetrating to Japan. PMID- 15287475 TI - [Antimicrobial resistance in Streptococcus pneumoniae and Haemophilus influenzae isolated from nasopharynx in children]. AB - The aims of this study are to investigate the antimicrobial susceptibility of bacteria isolated from children and clarify the risk factors for the carriage of the resistant strains. We examined the minimum inhibitory concentrations (MICs) of antimicrobial agents against 949 strains of Streptococcus pneumoniae (S. pneumoniae) and 791 strains of Haemophilus influenzae (H. influenzae) isolated at our department between September, 2001 and May, 2003. Of those, 226 S. pneumoniae strains and 115 H. influenzae strains were analysed for the resistance genes. Also we retrospectively reviewed the profiles of 1,359 patients with either S. pneumoniae, H. influenzae, or both in nasopharynx. From the view point of MICs, PSSP strains were 185 (19%), PISP strains were 443 (47%), and PRSP strains were 321 (34%) in 949 S. pneumoniae strains, and BLNAS strains were 545 (69%), low BLNAR strains were 104 (13%), BLNAR strains were 81 (11%), and BLPAR strains were 61 (8%) in 791 H. influenzae strains. The results of gene analysis showed that all resistant strains by MICs such as PISP, PRSP, BLNAR, and BLPAR had resistant genes and that 55% of and 21% of susceptible strains of S. pneumoniae (PSSP) and H. influenzae (BLNAS), respectively, had resistant genes. From the investigation for profiles of 1,359 patients, age less than 3 years old, day nursery, and use of antimicrobial agents in last 3 month, seemed to be the risk factors for carriage of resistant strains. To prevent the resistant bacteria from disseminating we should re-consider how to use the antimicobial agents and nurse the young children. PMID- 15287476 TI - [Efficacy of azithromycin as the empiric therapy in children with community acquired pneumonia who were isolated macrolide resistant Streptococcus pneumoniae from nasopharynx]. AB - Identification of pathogens in childhood community-acquired pneumonia (CAP) is not easy. However, it is believed that nasopharyngeal colonization of pathogenic bacteria leads to childhood CAP, so the etiology is inferred by the isolates obtained from nasopharynx of children with CAP. Among the pathogens of childhood CAP, Streptococcus pneumoniae (SP) is the most important agent and macrolides resistant SP (MRSP) is increasingly reported. We investigated the characterization of the mechanism of macrolide resistance in isolates of MRSP by the presence of the ermB gene or the mefA gene and clindamycin (CLDM) resistance. In addition, we also assessed the efficacy of azithromycin (AZM) in children with CAP who were isolated MRSP from nasopharynx. During a 6 month period between January and June in 2002, children with CAP who were treated with a 3 day regimen of AZM and isolated SP from nasopharynx were enrolled. Clinical outcome was based on assessment of fever on the fourth day of treatment. MIC measurements were obtained by broth microdilution and interpreted according to NCCLS criteria. 53 patients were enrolled and MRSP were isolated in 41 children. Of 41 MRSP isolates, 25 isolates were identified CLDM resistance. The AZM MIC90 of CLDM resistant MRSP isolates was 128 microg/ml. On the other hand, that of CLDM sensitive MRSP isolates was 8 microg/ml. However, AZM was effective in 20 children isolated CLDM resistant MRSP and 15 out of 16 children isolated CLDM sensitive MRSP. On this background, despite high rates of MRSP in Japan, AZM continues to be clinically effective for the treatment of childhood CAP. PMID- 15287477 TI - [Mycoplasma pneumoniae infection in hospitalized children with acute pneumonia under the Mycoplasma epidemic]. AB - Since October 2000, Mycoplasma pneumonia has been a recurring epidemic in Japan. To become clear the importance of Mycoplasma pneumoniae infection in children, we investigated cross-sectionally M. pneumoniae infection by serology in the hospitalized children age under seven years with acute pneumonia retrospectively reviewing pediatric patients of the four studies about lower respiratory tract infection which we had been treated during 2001 to 2003. Firstly, we found M. pneumoniae infection in 75 patients (33.8%) among a total of 222 patients with asthma exacerbation and acute pneumonia in 2001. Second, we had evaluated a total of 46 hospitalized children with acute pneumonia for M. pneumoniae infection in November 2002 and 18 patients (39.1%) were found. Thirdly, we found M. pneumoniae infection in 8 patients (34.8%) among 23 patients with respiratory syncitial virus and acute pneumonia age under two years during October 2002 to April 2003. Fourthly, we found M. pneumoniae infection in 19 patients (35.8%) among 53 patients with asthma exacerbation and acute pneumonia during January from June in 2003. Even only among the patients age under two years M. pneumoniae infection was found to be 24.3% (16/70), 27.8% (5/8), 34.8% (8/23) and 33.3% (7/21), respectively. These findings demonstrate that M. pneumoniae is common pathogen of acute pneumonia even in infants and young children under Mycoplasma epidemic. Not only typical bacteria and but also M. pneumoniae should be considered as important pathogens in the treatment of acute pneumonia in infants and young children under Mycoplasma epidemic. PMID- 15287478 TI - [Profiles of the expression of phospholipase A2 family mRNAs by macrophages infected with Mycobacterium tuberculosis]. AB - Previously, we found that collaboration of reactive nitrogen intermediates (RNI) and free fatty acids (FFA) plays crucial roles in the expression of macrophage (Mphi) antimicrobial activity against Mycobacterium tuberculosis (MTB). Phospholipase A2 (PLA2) families hydrolyze phospholipids and release FFA from their sn-2 position. In the present study, we examined profiles of the mRNA expression of PLA2 families by Mphis stimulated with MTB by using RT-PCR method, and the following results were obtained. First, the expression of type IV cytosolic PLA2 (cPLA2), which is highly specific to arachidnic acid moiety, was significantly up-regulated by MTB stimulation of Mphis. Second, the expression of type IIa secretory PLA2 (sPLA2) was not observed for Mphis with or without MTB stimulation. Third, the profile of mRNA expression of type V sPLA2 was nearly the same as that of type IV cPLA2 for Mphis before and after MTB stimulation. These findings suggest that type IV cPLA2 and type V sPLA2 both play important roles in the FFA-mediated Mphi Cap antimicrobial activity against MTB organisms. PMID- 15287479 TI - [Analysis of the resistance gene in Streptococcus pneumoniae]. AB - Resistance genes were determinded for 81 strains of Streptococcus pneumoniae isolated from Ehime University hospital, during 2002 and 2003 by various clinical material. In penicillin-binding proteins of mutation, there were 74 strains; pbp2x mutation 23 strains (28.4%), pbp2b mutation one strain (1.2%), pbp1a + pbp2x mutations 5 strains (6.2%), pbp2x + pbp2b mutations 18 strains (22.2%) and all mutations 27 strains (33.3%). As for the result of macrolide resistance genes, there were 67 strains; mefA gene 20 strains (24.7%), ermB gene 46 strains (56.8%) and both gene one strain (1.2%). In the analysis of gyrA gene and parC gene, 3 strains (3.7%) had both gene mutations, and 26 strains (32.1%) had only parC gene mutation. There was more of an increase than before in isolates, two or more mutation strains with PBPs gene, ermB gene holding strains and the levofloxacin resistance strain. These results suggest that the gyrA gene or parC gene mutation strains hold PBPs gene mutation and macrolide resistance genes in a high rate, and there will be more drug resistance in the future. PMID- 15287480 TI - [A case of pulmonary coccidioidomycosis presented with bilateral infiltrative opacities and eosinophilia]. AB - A 53-year-old male was admitted to our hospital complaining of high fever with chillness, cough and dyspnea after traveling to Arizona in the United States. The chest X-ray films taken on admission showed consolidation in the right middle lung field and bilateral nodular shadows. The laboratory data revealed an increase in white blood cell counts with eosinophilia, and a rise in erythrocyte sediment rate and serum C-reactive protein. The biopsied lung specimen by video assisted thoracoscopic surgery showed granulomatous inflammation consisting of eosinophils and giant cells. In addition, typical spherules filled with endopores were detected in the specimen. The diagnosis of primary pulmonary coccidioidomycosis was made. After the treatment of a three months' regimen with itraconazole at the daily dosage of 200 mg, the patient's symptoms, laboratory data and radiological findings markedly improved. PMID- 15287481 TI - [Evaluation of a new standard beta-glucan CSBG in the measurement of beta-glucan in plasma by alkaline treatment, chromogenic automated kinetic assay]. PMID- 15287482 TI - [Strategy for omental transplantation in moyamoya disease]. AB - Intracranial omental transplantation is sometimes indicated for treatment of ischemia in the territory both of the anterior cerebral artery and of the posterior cerebral artery in certain cases with moyamoya disease. The surgical process for omental transplantation is thought to be complicated and time consuming. For this reason some technical improvements for intracranial omental transplantation are presented in this report. 1) Technique for harvest of omentum. 2) A gastroepiploic artery is anastomosed to a scalp artery in an end-to side fashion, or in an end-to-end fashion. 3) A gastroepiploic vein is anastomosed to a cortical vein in an end-to-side fashion. PMID- 15287483 TI - [Nocardial brain abscess: surgery and postoperative antibiotic therapy]. AB - Nocardial infections in an immunocompromised host have been increasingly reported. Nocardial brain abscess, the most common presentation of nocardiosis in the central nervous system, is associated with a high mortality rate because of its delayed diagnosis and its unresponsiveness to the usual antibiotic therapy. We report four patients who experienced a long-term cure of nocardial brain abscess due to treatment by a combination of surgery and postoperative antibiotic therapy; 1 man and 3 women, ages ranging from 43 to 67 years old. Two patients were associated with systemic lupus erythematosus and two with autoimmune hemolytic anemia. Patients underwent surgical aspiration and drainage of brain abscess. Nocardia was identified from the aspirated specimen and postoperative antibiotic therapy for 5-6 weeks was performed using effective antibiotic agents; sulfamethoxazole/trimethoprim (ST), imipenem/cilastatin and minocycline (MINO) in Case 1, ST and MINO in Case 2, erythromycin in Case 3, and panipenem/betamipron and cefotaxime in Case 4. Case 3 and Case 4 with multilobulated brain abscess underwent total excision of the brain abscess. All patients showed successful cure of nocardial brain abscess with no recurrence for the period of 1-8 years. The combination of surgery and postoperative antibiotic therapy provides a good prognosis for nocardial brain abscess. PMID- 15287484 TI - [A guide to initial management of minor head injury]. AB - We reviewed the records of 1,335 minor head injury patients with initial Glasgow Coma Scale (GCS) scores of 15 treated by our neurosurgery service between January 1998 and December 2000. Skull X-ray was performed in 945 patients (71%), and Computed tomography (CT) was performed in 590 patients (44%). Skull fracture was shown radiographically in 24 patients (2.5%), and abnormalities on the initial CT were seen in 29 patients (4.9%). The most frequent intracranial lesion on CT was acute epidural hematoma with skull fracture. Significantly more intracranial lesions were found in those with a fracture than in those without by chi2 analysis. Post-traumatic vomiting was significantly associated with radiographical abnormalities, but headache and nausea did not increase the risk of skull fracture and intracranial lesions on the CT. Patients required neurosurgical intervention in 4 cases, and all of those were acute epidural hematoma with skull fracture. In this study, the first thing we should do for asymptomatic minor head injury patients with a GCS score of 15 is to investigate the presence of a skull fracture by skull X-ray. Head trauma patients with a skull fracture and post-traumatic vomiting should undergo CT to facilitate detection of intracranial lesions, even when there are no abnormal neurological signs. PMID- 15287485 TI - [Ion-beam irradiated ePTFE for the therapy of intracranial aneurysms]. AB - Intracranial aneurysms are frequently treated with either microsurgical clipping or endovascular coiling. However, as so called broad-neck aneurysms are not suitable for these treatment options, a wrapping technique using muslin gauze, muscle piece, ePTFE is applied for such cases. The material for aneurysmal wrapping demands both stable adherence and no reactive inflammatory response such as inert artificial wall. Authors have developed a new improved ePTFE by ion-beam irradiation technique that is biologically inert and able to adhere firmly to surrounding tissue. Based on the last studies, Ar+ ion at an energy of 150 keV with a fluence of 5 x 10(14) ions/cm2 was chosen to irradiate ePTFE. A cell adhesion test and direct implantation of ion-beam irradiated ePTFE as wrapping material to rabbit common carotid arteries (CCA) were examined. It was demonstrated that the surface of ion-beam irradiated ePTFE exhibits remarkably greater adhesion and promotes cell proliferation on the surface more effectively than that of non-irradiated ePTFE. The carotid artery well-wrapped by ion-beam irradiated ePTFE strongly adhered to the mural wall and induced little inflammatory reaction. The results of this investigation indicate that application of this technology would offer the best means for aneurysm wrapping. PMID- 15287486 TI - [Adult type tethered cord syndrome with chronic attackwise pain in the bilateral feet]. AB - The authors report a case of chronic attackwise pain in the bilateral feet for five years due to tethered cord syndrome. Despite extensive examinations, this condition had been overlooked. The patient is a 21-year-old man. He had suffered attackwise pain resembling sticking a thumbtack in the soles of his feet, since he was 16 years old. The pain appeared several times a day and continued for 30 seconds to 30 minutes for 5 years. Physical examination revealed hammer toes and high-arched feet. The fingers and knee joints showed hyperextension. The neurological findings showed weakness of toe extension, hyporeflexia of deep tendon reflexes in the leg. Mild hypesthesia was seen in the bilateral soles. Myelography showed sacral dural ectasia. Magnetic resonance images showed dorsal displacement of the conus medullaris, the filum terminale and the cauda equina. A computed tomographic scan after myelography also showed a dorsally located thick filum terminale (the diameter is 2 mm). Surgery disclosed thick and tight filum terminale directly under the dura mater. Its flexibility was diminished. Abnormal lesions such as lipoma, spinal dysraphysm, diastematomyelia, myelomeningocele were not observed. After the untethering operation, the pain attacks decreased dramatically. The condition of the present case is adult onset tethered cord Group 2 described by Yamada. When unusual pain is manifested, we always have to keep this syndrome in mind. PMID- 15287487 TI - [A case of convulsion during selective intra-arterial infusion of fasudil hydrochloride for treatment of vasospasm following subarachnoid hemorrhage]. AB - The authors report a case of convulsion during intra-arterial selective infusion of fasudil hydrochloride (FSD) for treatment of vasospasm following subarachnoid hemorrhage (SAH). A 47-year-old man (Hunt and Kosnik grade I) presented with sudden onset of headache and was diagnosed with SAH on CT, and admitted to our hospital. Digital subtraction angiography (DSA) performed on admission revealed an anterior communicating artery aneurysm. Neck clipping of the aneurysm was performed on the same day and no neurological deficits were noted postoperatively. Motor aphasia appeared on day 11 after the operation, and emergency DSA revealed vasospasm of the left middle cerebral artery and its branches. Emergency percutaneous transluminal angioplasty was performed with successful dilation of the left M1 artery, and 25 milligrams of FSD was injected into the left M1 artery selectively. During this injection, right hemifacial convulsion appeared, and three minutes later disappeared. No treatment was needed for the seizure. Additional injection of 30 milligrams of FSD into the left internal carotid artery resulted in vasodilatation of the left M1 artery and its branches, improvement of their blood flow on angiography, and recovery from motor aphasia. The patient was discharged 1 month later with no neurological deficits. Intra-arterial selective infusion of FSD plays an important role in treatment for vasospasm following SAH, however, we must be aware of risks of complications such as convulsion. PMID- 15287488 TI - [A surgically treated case with a ruptured bacterial aneurysm of the middle cerebral arterial bifurcation following occlusion]. AB - A 56-year-old woman with aortic regurgitation (AR) developd a high fever on April 25th, 2003, followed by the sudden onset of left hemiparesis and dysarthria on May 10th, 2003. MRI and MRA showed cerebral infarction due to occlusion of the right proximal portion of the middle cerebral artery. Streptococcus was isolated from arterial blood culture at the time of admission and cardiac examination such as echocardiography revealed active infective endocarditis. Cerebral angiography on the 31st day after the onset of symptoms demonstrated a fusiform-shaped aneurysm at the occluded M2 portion of the middle cerebral artery. Despite administration of antibiotics, a small subcortical hematoma was observed in the right temporal lobe surrounding the aneurysm on the 35th day. The direct surgery of aneurysmal trapping and resection was subsequently performed to prevent rebleeding. The sylvian fissure and perianeurysmal area were strongly adherent to granulation tissue and blood clot. After exposing the aneurysm, the dilated portion of the vessel was successfully trapped and resected. Other than residual left hemiparesis, the postoperative course was uneventful. Histological examination confirmed bacterial aneurysm due to bacterial embolization originating from infective endocarditis (IE). We report a rare case having a ruptured bacterial aneurysm of the middle cerebral arterial bifurcation requiring surgery following occlusion due to bacterial embolization after sepsis and meningitis due to infective endocarditis. PMID- 15287489 TI - [A multicentric glioma presenting different pathological appearances: a case report]. AB - We report a multicentric glioma case which revealed different pathological appearances. A 45-year-old male had been admitted to our hospital complaining of an attack of transient sudden aphasia. On magnetic resonance imaging (MRI), T1 weighted images revealed a low intensity and T2-weighted images demonstrated a homogeneous high intensity abnormal mass in the frontal lobe, which was not enhanced with gadolinium. Removal of the tumor was performed through a right frontal transcortical approach in March, 2002. Histological diagnosis was gemistocytic astrocytoma. The patient's condition was uneventful and postoperative MRI revealed a marked decrease in the volume of the tumor. A total of 54 Gy radiation to the brain in the locality was performed. Four months after the initial surgery, the patient suffered from incomplete right hemiparesis. MRI showed a left parietal abnormal mass which had a ring formation enhancement after gadolinium administration. This Neuro-radiological examination demonstrated complete independence from the initial right frontal tumor. A second surgery which was concerned with cyst aspiration was carried out on August 10, 2002. During the next month, a third operation for partial removal of a left parietal abnormal mass was performed. Histological diagnosis was anaplastic astrocytoma. The right frontal and left parietal tumors revealed neither continuous relation suggesting intracerebral invasion, nor dissemination through the subarachnoid space nor intracerebral metastasis. Our case was diagnosed as multicentric glioma with different pathological appearances, of which only 9 cases have been reported previously. PMID- 15287490 TI - [Idiopathic spinal cord herniation which extended remarkably up- and downward from dural defect: case report]. AB - A case of idiopathic spinal cord herniation which extended remarkably up- and downward from a dural defect is described. A 53-year-old woman presented with numbness and pain of the right lower limb. Magnetic resonance (MR) imaging revealed ventral displacement of the spinal cord and dilation of the dorsal subarachnoid space at T4-5. CT Myelography showed ventral deviation of the spinal cord at the T4/5 level. A laminectomy of T3-T5 was performed, and the herniated spinal cord was untethered and wrapped by Goretex membrane. Postoperative MR image revealed normal location of the spinal cord. Among many cases of spinal cord herniations, this one is considered to be a rare case of idiopathic spinal cord herniation which showed marked protrusion up- and downward from a dural defect. PMID- 15287491 TI - [Large kissing aneurysms of the middle cerebral artery: a case report- classification of kissing aneurysms]. AB - We report a rare case of kissing aneurysms located at the middle cerebral artery. A 69-year-old man had a severe subarachnoid hemorrhage associated with intracerebral hematoma (Hunt and Hess grade 5, WFNS grade V). Angiography revealed two large-sized aneurysms of the middle cerebral artery, and these aneurysms were seen as contacting each other. Both aneurysms were adherent, with fibrous tissue at each dome site. Neck clipping was performed. The difficulty of neck clipping with kissing aneurysms is dependent upon the relationship between the two aneurysmal necks. We classify kissing aneurysms into two groups based on the location of the aneurysmal neck (Type 1: each aneurysmal neck is located on the same parent artery. Type 2: each aneurysmal neck is located on different parent arteries.). In Type 1, preoperative diagnosis of kissing aneurysms is difficult and premature rupture during the application of a clip occurs frequently. Therefore, careful and meticulous dissection between the aneurysms is especially required. On the other hand, with Type 2 cases, large aneurysms (> 15 mm) are seen much more frequently than in cases of Type 1. Our classification of kissing aneurysms is useful to assess the difficulty of neck clipping for these aneurysms. PMID- 15287492 TI - [Risk management and medical safety in neurosurgical field--from the standpoint of characteristics of neurosurgery]. PMID- 15287493 TI - [Thalamic deep brain stimulation therapy: a guide to stereotactic brain operations]. PMID- 15287494 TI - [Clinico-pathology of skull tumor: osteoma, osteoblastoma, osteosarcoma]. PMID- 15287495 TI - [SREBP controls cholesterol homeostasis]. PMID- 15287496 TI - [Regulation of cholesterol homeostasis by nuclear receptors]. PMID- 15287497 TI - [Activation of "fat burning sensor" peroxisome proliferator-activated receptor delta induces fatty acid beta-oxidation in skeletal muscle and attenuates metabolic syndrome]. PMID- 15287498 TI - [ABC proteins involved in lipid homeostasis]. PMID- 15287499 TI - [ABCA1-mediated generation of HDL and its non-transcriptional regulation]. PMID- 15287500 TI - [Tissue-specific expression of ABC transporters involved in lipid transport]. PMID- 15287502 TI - [Lipid transfer proteins]. PMID- 15287501 TI - [Transporters for bile lipids]. PMID- 15287503 TI - [The molecular machinery CERT for intracellular trafficking of ceramide]. PMID- 15287504 TI - [Intracellular cholesterol transport by NPC1/HE1]. PMID- 15287505 TI - [Lipid droplet as an independent organelle]. PMID- 15287506 TI - [Etiology and pathogenesis of Parkinson's disease: from mitochondrial dysfunctions to familial Parkinson's disease]. AB - Parkinson's disease (PD) is the second most common neurodegenerative disease after Alzheimer's disease. It is urgently needed to elucidate the cause of the disease and to establish neuroprotective treatment. We have been working on the etiology and pathogenesis of PD for many years and we found selective loss of mitochondrial complex I and the alpha-ketoglutarate dehydrogenase complex in the nigral neurons of patients with PD. Our observation firmly established mitochondrial defects in PD. Mitochondrial respiratory failure induces oxidative damage in neurons, and we found increase in hydroxynonenal and 8-oxo deoxyguanine, indices of oxidative damage, in the nigral neurons of PD. These abnormalities can trigger apoptotic cell death. The primary events which induce mitochondrial failure and oxidative damage are not known, however, it has been postulated that the interaction of genetic risk factors and environmental factors would initiate the degenerative process. Based on this assumption, we conducted genetic association studies by the candidate gene methods. We found that polymorphic mutations of superoxide dismutase-2 and 24-kDa subunit of mitochondrial complex I were associated increased risk of developing Parkinson's disease. While we were doing this genetic association study, we found a family, in which parkinsonian phenotype completely segregated with a polymorphic mutation of the superoxide dismutase-2 gene. In this family, 4 out of 6 siblings were affected with early onset parkinsonism and the parents were apparently normal. Thus the mode of inheritance appeared to be autosomal recessive and this type is now called as AR-JP or Park2. We confirmed the linkage of this type of familial Parkinson's disease to the superoxide dismutase loci that is located in the telomeric region of chromosome 6 by the linkage analysis using microsatellite markers in this region. Then we found another family, in which an affected patient showed lack of one of the microsatellite markers (D6S315), which we were using in the linkage analysis. This observation prompted us to initiate the molecular cloning of the disease gene utilizing D6S315 as the initial probe. The molecular cloning was done with the collaboration with Professor Nobuyoshi Shimizu of Keio University. We identified a novel gene and confirmed that mutations of this novel gene were found only in the patients with autosomal recessive Parkinson's disease. The novel gene was named parkin. We conducted mutational analysis on more than 700 families with Parkinson's disease. We also established a method to detect compound heterozygotes of parkin mutations. Mutinous of the parkin gene were found in approximately 50% of autosomal recessive families. Many kinds of exonic deletions and point mutations were found. This type of familial Parkinson's disease had been considered to be unique among Japanese, but since we started mutational analysis of the parkin gene, we confirmed the world wide distribution of parkin gene mutations. Then we analyzed functions of parkin protein with the collaboration with Dr. Keiji Tanaka of Tokyo Metropolitan Institute of Medical Sciences. We found that parkin protein was a ubiquitin-protein ligase of the ubiquitin system. Now we are working on the candidate substrates of parkin protein as a ubiquitin ligase. We found that CDCrel-1, a synaptic vesicle protein, was a candidate substrate of parkin protein. In addition, we found two additional candidate proteins, i.e., alpha synuclein 22 and PAEL receptor, with the collaboration of Professor Denis Selkoe of Harvard Medical School and Dr. Ryosuke Takahashi of RIKEN, respectively. Accumulation of PAEL receptor in the endoplasmic reticulum causes endoplasmic reticulum stress and apoptotic cell death. We found evidence to indicate accumulation of PAEL receptor and the presence of endoplasmic reticulum stress in a patient with AR-JP (Park2). Thus our studies firmly established that a genetic defect of an enzyme in the ubiquitin-proteasome system induces selective nigral neuronal death. We indicated the important role of the ubiquitin-proteasome system in neurodegeneration in general. In many other neurodegenerative disorders, such as Alzheimer's disease, Huntington's disease, Machado-Joseph disease, dentatorubral-pallidoluysian atrophy, and ALS, ubiquitinated proteins are accumulated in neurons. Thus protein handling in the ubiquitin-proteasome system appears to be affected in these neurodegenerative disorders despite the difference in the primary defects. Our studies also suggest many potential approaches for the discovery of neuroprotective treatment for not only Parkinson's disease but also other neurodegenerative disorders. PMID- 15287507 TI - [Comparison between MRI and 3D-SSP in olivopontocerebellar atrophy and cortical cerebellar atrophy]. AB - We compared images of three-dimensional stereotactic surface projections (3D-SSP) of SPECT with MRI images in spinocerebellar degeneration patients (13 olivopontocerebellar atrophy (OPCA) and 7 cortical cerebellar atrophy (CCA)). We analyzed a brain blood flow pattern with an image of statistics by 123I-IMP SPECT. In OPCA patients, a blood flow reduction was more remarkable in 3D-SSP than a degree of cerebellar atrophy in MRI. In patients with CCA, the cerebellum showed little blood flow reduction in 3D-SSP despite of apparent atrophy in MRI. Simultaneous examination both MRI and 3D-SSP might be useful for differential diagnosis of spinocerebellar degenerations. PMID- 15287508 TI - [Respiratory system elastance and resistance measured by proportional assist ventilation in patients with respiratory muscle weakness]. AB - OBJECTIVE: Non-invasive ventilatory therapy has prolonged survival of myopathy patients with hypoventilation. Efficacy of non-invasive ventilation depends on both elastance and resistance of the respiratory system. Although these parameters are important in the prescription of respiratory management, conventional respiratory function test does not show the appropriate answer in patients with severe respiratory muscle weakness. In muscular dystrophy, muscle tends to be shortened due to its fibrosis, when muscle becomes atrophic and weak; fibrosis of respiratory muscle tissues presumably causes high thoracic elastance. We evaluated the total respiratory system elastance and resistance during proportional assist ventilation (PAV) in myopathy patients. METHODS: In PAV with 100% assist, using BiPAP Vision ventilator, airway pressure exceeds 20 cmH2O or tidal volume exceeds 1.5 liter (run-away phenomenon) when the volume assist or the flow assist is higher than the individual elastance or the resistance, respectively. Twenty myopathy patients with ventilatory failure and 7 healthy controls were evaluated, including 7 patients with Duchenne muscular dystrophy (DMD), 2 patients with congenital myopathy (CM), 1 patient with limb-girdle muscular dystrophy (LG), 6 patients with myotonic dystrophy (MyD) and 4 patients with acid maltase deficiency (AMD). Seventeen patients used a nasal mask and 3 patients had a tracheostomy tube. Fifteen patients used a pressure-preset ventilator, and 3 patients used a volume-preset ventilator. RESULTS: In all patients with DMD, CM and LG, respiratory system elastance was higher than 20 (cmH2O/L) and than in all patients with AMD and MyD except 1 MyD patient. Follow up measurement after half a or one year showed increase of respiratory system elastance in 2 DMD patients and 1 CM patient, but almost no change in 3 AMD patients. The elastance measured during PAV was consistent with the clinical impression of muscle shortening. One exceptional MyD patient showed extremely high elastance (more than 58 cmH2O/L), which reflected the fixed thoracic spine and increase of abdominal visceral fat. Resistance was normal in all patients except a LG patient with pulmonary aspergillosis and a history of pulmonary tuberculosis who showed 14 (cmH2O/L/s). In a CM patient who developed emphysema, resistance increased from 5 to 12 (cmH2O/L/s) in a year, although forced expiratory volume 1.0% (FEV1.0/FVC) remained normal. Respiratory system resistance measurement was useful to detect a lung disease, because obstructive disorder is underestimated with FEV1.0/FVC when vital capacity is low. CONCLUSION: The respiratory system elastance and resistance measured during PAV are useful parameters in evaluation of mechanical features of the lung, thorax and airway. It is recommended to keep both parameters normal in patients who may require ventilatory assist due to progression of respiratory muscle weakness. PMID- 15287509 TI - [A case of CADASIL (cerebral autosomal dominant arteriopathy with subcortical infarcts and lekoencephalopathy) with Notch 3 (Arg169Cys) mutation and typical granular osmiophilic materials in peripheral small arteries]. AB - We report a 64-year-old Japanese woman with recurrent ischemic strokes and progressive dementia without any cardiovascular risk factors. Her first stroke was at 45 years old, and she has a family history of ischemic strokes compatible with an autosomal dominant trait. Marked leukoaraiosis and multiple lacunar infarcts were shown on brain MR images, and no atherosclerotic changes were observed in her extra- and intra-cranial arteries by cervical arterial echography and intracranial MR angiography. Excluded other inherited or metabolic diseases causing leukodystrophy by examination of her blood samples, her disease was diagnosed as CADASIL (cerebral autosomal dominant arteriopathy with subcortical infarcts and lekoencephalopathy). We demonstrated granular osmiophilic materials (GOM) on the wall of small arteries from a biopsied peripheral nerve tissue specimen and detected a mutation Arg169Cys of Notch 3 gene. Many CADASIL patients have been reported and over 28 kinds of mutations of the Notch 3 were identified in western countries, while few CADASIL patients have been reported in Japanese people. Among them, eleven CADASIL families have been reported and only five mutations (Arg133Cys, Cys174Phe, Arg213Lys, Arg90Cys and Arg141Cys) have been determined so far. The mutation of Notch 3 in our patient was determined as Arg169Cys, and this is the first report on a Japanese patient with CADASIL due to this mutation. PMID- 15287510 TI - [A case with severe respiratory muscle weakness due to chronic myositis associated with PBC]. AB - We report a 37-year-old woman with slowly developing muscular weakness for 2 years following insidious stiffness of calf muscle. Serum CK was elevated up to 4,207 IU/l. She presented sleepiness, weakness of proximal and truncal muscles and systemic muscular atrophy. While she had not experienced dyspnea, her arterial blood gas analysis revealed extreme hypoxia and hypercapnea due to weakness of respiratory muscles. Echocardiogram showed thinness and hypokinesis of left ventricular wall, and arrhythmia was pointed out by holter ECG. Needle elctromyogram of the proximal muscles exhibited polyphasic units with low amplitude. Muscle biopsy showed degeneration and necrosis of muscle fibers as well as regeneration. Mild infiltration of inflammatory cells was shown. Serological examination showed positive antimitochondrial M2 antibody, especially specific for primary biliary chirrhosis (PBC). She was diagnosed as chronic myositis associated with PBC. Four cases of idiopathic myositis with severe weakness of respiratory muscle, associated with PBC had been reported. These cases and our present case share the similar feature in respect of insidious or chronic course and resistance to therapy. In our present patient, respiration had been supported by BiPAP and she has been successfully improving slowly by oral steroid following 4 courses of methylprednisolone pulse therapy. PMID- 15287511 TI - [Palpable orbital subcutaneous masses in chronic inflammatory demyelinating polyneuropathy. MRI and neurophysiological study of multiple peripheral nerve swelling]. AB - We report a 46-year-old woman with chronic inflammatory demyelinating polyneuropathy (CIDP) in whom swelling of the first branch of the trigeminal nerves in the bilateral orbits were observed as subcutaneous masses in the upper eyelids. The disease developed when the patient was 33 years old, and weakness of the four limbs, double vision, unilateral hypoglossal neuroparalysis, and unilateral facial paralysis frequently occurred during the course of the illness. On nerve conduction studies, conduction block was detected in the motor nerves. Steroid therapy and immunoglobulin treatment improved the symptoms. At 43 years old, subcutaneous phymas were noted in the bilateral upper eyelids, and fat suppressed MRI detected the phymas localized along the upper region of the superior straight muscle in the bilateral orbits, and the muscles were slightly compressed downward. The masses branched in the orbits, and were diagnosed as nerve swelling of the supraorbital nerve, the first branch of the trigeminal nerve. Fat-suppressed MRI also identified nerve swelling of the extracranial maxillary and mandibular nerves. However, the patient had no subjective sensory disturbance in the trigeminal nerve region. Blink reflex did not induce R1 and R2 exhibited low amplitude and delayed latency. MRI confirmed asymmetric nerve swelling in the regions of the bilateral median nerves with motor nerve conduction block. Lumbar MRI detected nerve swelling in the peripheral nerves distant from the lumbar ganglion. MRI detected no nerve swelling in the arachnoid space such as the cervical cord, thoracic cord, or cauda equina. Aggravation of CIDP was treated with steroids and immunoglobulin. Diplopia occasionally appeared, but was not consistent with aggravation of CIDP. The bilateral supraorbital nerves remained unchanged for three years on MRI. PMID- 15287512 TI - [A patient with mitochondrial encephalomyopathy presenting gynecomastia with elevation of serum estriol level]. AB - We report a 23-year-old man with mitochondrial encephalomyopathy. At 21 years of age, he noted speech distubance. Since his dysarthria did not improve thereafter, he was admitted to our hospital. On admission, he showed mild gynecomastia. Neurological examination revealed mild decrease in performance IQ in WAIS-R, mild scanning speech, mild left hearing disturbance, mild to moderate muscle weakness in proximal four extremities, mild bilateral limb ataxia, and mild to moderate truncal ataxia. While, no brisk deep tendon reflex, pathological reflex, aberrant muscle tonus, sensory disturbance, retinopathy, myoclonus or autonomic disorder was found. Serum levels of lactate (23.2 mg/dl, normal<18.7) and pyruvate (1.23 mg/dl, normal<0.94) were elevated, and serum lactate levels were markedly elevated (118.1 mg/dl) after 15-minute exercise (15 Watts/minute). CSF levels of lactate (31.2 mg/dl, normal<12.5) and pyruvate (1.48 mg/dl, normal<0.75) were also elevated. Head MRI showed mild cerebral and cerebellar atrophy, but 1H-MRS showed no lactate peak. Moreover, muscle biopsy from left biceps muscle showed lots of ragged-red fibers, and he was thus diagnosed as having mitochondrial encephalomyopathy. However, nt3243 mutation of mitochondria DNA was not present. Next, we confirmed gynecomastia by mammography, and checked serum levels of estrogens. Mildly decreased estradiol (19.9 pg/ml, normal, 20-59), normal estrone (24.0 pg/ml, normal<30.0) and mildly increased estriol (6.03 pg/ml, normal<5.0) were found. While, the serum levels of cortisol, dehydroepiandrosterone-sulfate (DHEA-S), androstenedione, testosterone, luteinizing hormone (LH) and follicle stimulating hormone (FSH) were all within normal limits. Since the steroid hormone synthesis system and hypothalamus-pituitary system seem to be normal, 16alpha-hydroxylase that converts estradiol to estriol may be upregulated. While, aromatase (P-450arom) is well known to convert androgens to estrogens. In addition, 16alpha-hydroxylase and P-450arom convert DHEA-S to estriol. Since it is recently reported that P-450arom is considerably expressed in muscle tissues as well as fat tissues and that muscle tissue may be a major organ to produce estrogens in men and postmenopausal women, estriol production may be increased in the present patient's muscle. Although hypogonadism due to hypothalamus-pituitary disorders was sometimes reported, there have been no reports that suggest an increased estrogen production in skeletal muscles in mitochondrial encephalomyopathies. Recently, estrogen has been known to protect muscle fibers from oxidative damages due to exercise. Thus, it is of potential that estrogens increased locally in muscle tissues of the patients with mitochondrial encephalomyopathies protect muscle fibers from oxidative damage due to mitochondrial dysfunction. PMID- 15287513 TI - [Neurosyphilis presenting the left total ophthalmoplegia: a case report]. AB - We report a 73-year-old woman with meningitis-type neurosyphilis presenting the main symptom of the left total ophthalmoplegia. Three months after the appearance of the deviation of the eyeball to the inside and ptosis of the left eyelid, the left eyelid was completely closed. On admission, about four months after the appearance of neurosyphilis, she showed paralysis of the left oculomotor nerve, trochlearis nerve and abducens nerve, and the right mydriasis and absent light reflex. She was diagnosed as meningovascular neurosyphilis because syphilitic antibodies reactions in both serum and cerebrospinal fluid were highly positive. We treated her with intravenous infusion drop of penicillin G (eighteen-million units/day) for ten days, and those symptoms mentioned above other than light reflex were completely recovered. Bilateral internal carotid arteries situated close each other at the supraclinoid portion. The internal carotid arteries were not enhanced on Gd-MRI and the stenosis of the arteries were not detected on MRA. However, we suppose that the inflammation of meninges at that portion spreads to the bilateral internal carotid arteries, and that the III, IV and VI nerves close to the left internal carotid artery were damaged. There have been no reports of meningovascular neurosyphilis with the manifestation of unilateral ophthalmoplegia. In the patients of meningovascular neurosyphilis, however, various cranial nerve palsies can be appeared. Therefore we suggest that neurosyphilis should always be taken into consideration as differential diagnosis of cranial nerve palsies. PMID- 15287514 TI - [Eosinophilic fasciitis associated with Borrelia afzelii infection]. AB - A 58-year-old woman suffered from stiffness, swelling, rubor and muscle pain in the extremities one month after she climbed a mountain in Kyushu, an island in southern Japan. On admission, mild proximal weakness was present in the extremities. Her range of motion in the extremities was limited due to firm skin and subcutaneous stiffness which was similar to scleroderma. She showed peripheral blood eosinophilia and hypergammaglobulinemia as well as a high erythrocyte sedimentation rate. An IgM antibody against Borrelia afzelii was positive. MRI of the skeletal muscles demonstrated enhancing fascia around the biceps brachii muscle, and a biopsy specimen revealed perivascular infiltration of mononuclear cells within the hypertrophic fascia. Eosinophilic infiltration was absent. We treated the patient with prednisolone, doxycycline and amoxicillin, which alleviated the symptoms. This is the first report to show that Borrelia afzelii was involved in eosinophilic fasciitis. PMID- 15287515 TI - [Revision of guidelines for diagnosis and treatment of chronic idiopathic thrombocytopenic purpura]. PMID- 15287516 TI - [Assessment of intergroup practice guideline for management of childhood ITP]. PMID- 15287517 TI - [Clinical practice guidelines for malignant lymphomas]. PMID- 15287518 TI - [Guidelines for ensuring the safety of blood transfusion]. PMID- 15287519 TI - [Principles and guidelines for the management of infectious diseases]. PMID- 15287521 TI - [Ethical issues in the development of regenerative medicine: research on human embryonic and adult stem cells]. PMID- 15287520 TI - [Human vascular lesions and thrombosis]. PMID- 15287522 TI - [Hematologic malignancies and the 1p36 chromosomal translocations]. PMID- 15287523 TI - [Thalidomide treatment of patients with refractory myeloma in the institutes participating in the Japan Myeloma Study Group]. AB - Thalidomide was used in 73 patients with refractory myeloma in 15 of 45 institutes participating in the Japan Myeloma Study Group. The mean age and male/female ratio were 63.8 years and 0.92 (35/38), respectively. Thirty-four patients (47%) were treated with only thalidomide, 27 patients (37%) were treated with thalidomide and steroids, and 12 (16%) were treated with thalidomide and chemotherapy. The mean initial, maximum, and maintenances dose of thalidomide were 111.0, 204.8, and 163.0 mg/day, respectively. Almost all of the patients were maintained on low-dose thalidomide between 100-200 mg/day. Complete, near complete and partial response was obtained in 31 patients (42.5%). The progression-free and overall survivals after thalidomide therapy were 9.8 and 21.3 months, respectively. The most common adverse effects were gastrointestinal disturbance, peripheral neuropathy, psychological signs, and skin eruption. In contrast to reports from Europe and America, no deep vein thrombosis was observed in this study. On the other hand, leukopenia was relatively frequently observed, and might be recognized as a serious adverse effect in myeloma patients. In conclusion, low-dose thalidomide is a useful and safe tool for the treatment of refractory myeloma. PMID- 15287524 TI - [Successful treatment with antiviral agents for human herpesvirus type 6 encephalitis following reduced intensity stem cell transplantation in a patient with myelodysplastic syndrome]. AB - We report here a patient who suffered from PCR-confirmed human herpesvirus type 6 (HHV-6) encephalitis following reduced intensity stem cell transplantation (RIST) from her HLA-matched sibling donor. A 66-year-old woman with MDS-RA underwent RIST from her HLA-matched brother. Engraftment was favorable and grade 2 GVHD (skin and intestine) was observed with good response to 60 mg of prednisolone. On day 162, she developed fever, headache, diplopia, disorientation and abnormal neurological findings including cervical stiffness and nystagmus. An analysis of cerebrospinal fluid (CSF) revealed 80 cells/microl, a glucose level of 50 mg/dl and a protein level of 97 mg/dl on day 162. Although computed tomography (CT) of the brain and electroencephalography (EEG) were nonspecific, HHV-6 was detected in the CSF using polymerase chain reaction (PCR) techniques and the patient was diagnosed as having encephalitis due to local reactivation of HHV-6. Administration of ganciclovir (GCV) and acyclovir (ACV) were started from day 162. Treatment with antiviral agents was effective, with total resolution of her symptoms and the DNA of this virus disappeared from the CSF after 23 days of treatment. This case shows that HHV-6 infection has to be considered in patients with neurological symptoms following stem cell transplantation, and suggests the necessity of PCR for HHV-6 virus from the CSF. PMID- 15287525 TI - [Registration of hematological disorders by the Kyushu Hematology Organization for Treatment (K-HOT) Study Group]. AB - The Kyushu Hematology Organization for Treatment (K-HOT) Study Group was organized in 1999 to study hematological disorders diagnosed in the participating institutions in the Kyushu district. We registered all new patients with hematological disorders and from February 2000 to the end of 2003, a total of 2908 patients had been registered. They include non-Hodgkin's lymphoma in 803 patients, leukemia in 556, multiple myeloma (MM) in 276, myelodysplastic syndrome in 273, and adult T-cell leukemia/lymphoma (ATL) in 269 followed in a decreasing order by idiopathic thrombocytopenic purpura, aplastic anemia, and other benign hematological disorders and myeloproliferative disorders. The annual incidence of MM is estimated to be much higher than that previously reported. It is also confirmed that ATL is still one of the frequently encountered lymphoid malignancies in the Kyushu district. PMID- 15287526 TI - [Brugada syndrome occurring after autologous peripheral blood stem cell transplantation for acute myeloid leukemia]. AB - A 74-year-old male was found to be suffering from the Brugada syndrome after undergoing high dose chemotherapy followed by autologous peripheral blood stem cell transplantation (ABSCT) for acute myeloid leukemia. A specific ECG pattern of right bundle-branch block and a coved-type ST-segment elevation in leads V1 through V3, which is compatible with the Brugada syndrome, was unmasked by febrile neutropenia on the 8th day after ABSCT. He experienced syncope on the 11th day due to ventricular tachycardia, which was immediately improved with the administration of intravenous lidocaine. The Brugada syndorome should be considered in febrile patients if they have a episode of syncope or ECG change. PMID- 15287527 TI - Mimics of brain tumor on neuroimaging: part II. AB - Brain tumor is a distinct pathological entity that differs from other diseases, including cerebrovascular, demyelinating, inflammatory, infectious, and various miscellaneous diseases. Insidious onset and gradual progression of signs and symptoms are common in patients with brain tumors, whereas the onset of cerebrovascular diseases is usually acute or sudden. Patients with demyelinating, inflammatory, or infectious diseases show subacute onset. Differentiation of brain tumors from other disorders is usually possible from the clinically and radiologically characteristic features. However, in some diseases other than brain tumors, an atypical clinical course and/or radiological findings may suggest or simulate those of brain tumors. The diagnosis of brain tumor is confirmed histopathologically, and appropriate therapies are given to the patient based on the histopathological type and grade of the tumor. In order to obtain a specimen for histopathological examination, surgical intervention is required. Other diseases are usually diagnosed clinically and radiologically. Invasive procedures should be avoided in making a diagnosis. Therefore, differentiation of brain tumors from other diseases is a critical issue for neuroimaging. Detailed inspection of images is necessary, and characteristic findings, and additional imaging methods, such as diffusion-weighted imaging, are often helpful for the differential diagnosis. We assess the imaging findings of diseases simulating brain tumors and review the literature. PMID- 15287528 TI - Clinical use of a new mechanical detachable coil system for percutaneous intravenous embolization of cavernous sinus dural arteriovenous fistulas. AB - PURPOSE: The purpose of this study was to evaluate the usefulness of a new mechanical detachable coil system (Detach-18/-11) for percutaneous transvenous embolization (TVE) of dural arteriovenous fistulas (AVFs) involving the cavernous sinus. MATERIALS AND METHODS: Five patients with dural AVF involving the cavernous sinus were treated by TVE with the use of the Detach-18/-11 system alone. All procedures were analyzed with regard to the processes of introduction, delivery, and/or retrieval of the Detach-18/-11 system, and angiographical and clinical outcome. RESULTS: A total of 70 detachable coils (37 spiral-type coils and 33 J-type coils) were used. Two coils were easily retrieved after introduction. The remaining 68 coils were easily delivered within 30 seconds. Neither premature detachment nor coil migration was observed. Of the five dural AVFs, three were completely occluded and two were nearly completely occluded immediately after embolization. Follow-up angiography showed complete occlusion in all cases. Clinical symptoms had disappeared within one month, and no recurrent symptoms were observed during follow-up (from 5 to 36 months). CONCLUSION: Our results support the safety and reliability of the Detach-18/-11 system for TVE of dural AVF. The availability of various types of this coil system allows sufficient packing of the involved sinus. PMID- 15287529 TI - Clinical evaluation of choline measurement by proton MR spectroscopy in patients with malignant tumors. AB - PURPOSE: To examine whether choline measurement by proton magnetic resonance (MR) spectroscopy in patients with malignant tumors is clinically meaningful in addition to routine MR imaging. MATERIALS AND METHODS: MR spectroscopy and MR imaging were performed in 27 consecutive patients with suspected malignant disease. Malignancy was assessed based on total choline compound levels using proton MR spectroscopy, and the results were compared with MR imaging findings. RESULTS: The sensitivity of MR imaging (84%, 21/25) was not significantly different from that of MR spectroscopy (88%, 22/25) among the 25 actual malignant diseases. Both MR imaging and MR spectroscopy produced two false-negative results. In one case, MR spectroscopy produced a false-negative result, whereas MR imaging produced a true-positive result. In two cases of benign breast disease, MR imaging produced false-positive results. MR spectroscopy produced one true-negative result and one false-positive result. CONCLUSION: Although choline measurement by MR spectroscopy is a useful tool in the evaluation of malignant disease, it should be reserved for patients with suspected malignant disease that cannot be detected by MR imaging, such as those with non-palpable prostate tumor with elevated sPSA levels. PMID- 15287530 TI - Transcatheter arterial embolization for advanced tumor thrombus with marked arterioportal or arteriovenous shunt complicating hepatocellular carcinoma. AB - PURPOSE: To evaluate the efficacy and safety of transcatheter arterial embolization with gelatin sponge immersed in an anti-cancer agent (GIA-TAE) alone or combined with radiation therapy, in hepatocellular carcinoma with portal vein tumor thrombus (PVTT) or hepatic vein tumor thrombus (HVTT) complicated by marked arterioportal or arteriovenous shunts. MATERIALS AND METHODS: GIA-TAE was performed in 15 patients with PVTT and arterioportal shunts to the main portal trunk or first-order branch, and five with HVTT and arteriovenous shunts, adding radiation therapy in suitable cases. Primary efficacy, hemodynamic changes, Child Pugh score, and survival rates were evaluated. RESULTS: GIA-TAE with or without radiation therapy was effective for tumor thrombus in 11 patients with PVTT and in four with HVTT. Treatment was effective for the main tumor in eight patients with PVTT and three with HVTT. Shunts disappeared in seven of 13 patients available for follow-up. Child-Pugh scores before and after the treatment were not significantly different. Median survival times of PVTT and HVTT groups were 8.7 and 12.2 months, respectively. One-year survival rates for both groups were about 15.6% and 50.0%, respectively. CONCLUSION: TAE with GIA alone or combined with radiation therapy is effective and safe for severe arteriovenous shunts with PVTT or HVTT, and also favors patient survival. PMID- 15287531 TI - A single institutional subset analysis of the WJLCG study comparing concurrent and sequential chemoradiotherapy for stage III non-small-cell lung cancer. AB - PURPOSE: To supplement findings of the West Japan Lung Cancer Group (WJLCG) study, treatment outcomes in our institution were reviewed from the perspective of radiation oncology. MATERIALS AND METHODS: Chemotherapy consisted of cisplatin (80 mg/m2 on days 1 and 29), vindesine (3 mg/m2 on days 1, 8, 29, and 36), and mitomycin (8 mg/m2 on days 1 and 29). In the concurrent arm, radiation therapy began on day 2 with a dose of 56 Gy in 28 fractions over 6.8 weeks, with an interval of 10 days at 28 Gy. In the sequential arm, radiation therapy began on day 50 with a dose of 56 Gy in 28 fractions over 5.6 weeks, without an interval. RESULTS: Twenty-four patients in the concurrent arm and 25 patients in the sequential arm in our institution were eligible for the WJLCG study. In the concurrent arm, three patients could not receive the full dose of radiation therapy and 12 patients required interruption of radiation therapy for more than 4 days. The median survival time among per-protocol patients and in those with interruption or with incomplete radiation therapy was 28.9 months and 14.1 months, respectively (p = 0.02). In the sequential arm, one patient could not receive the full dose of radiation therapy and none of the patients required such interruption. Local relapse and distant metastases as the first site of relapse occurred in 12 (11 in-field, 1 marginal) and five patients, respectively, in the concurrent arm, and in eight (7 in-field, 1 marginal) and 11 patients, respectively, in the sequential arm. CONCLUSION: In the concurrent regimen, noncompletion or interruption of radiation therapy was frequent, and the prognosis of such patients was poor. PMID- 15287532 TI - Manual aspiration thrombectomy with a standard PTCA guiding catheter for treatment of acute massive pulmonary thromboembolism. AB - PURPOSE: To evaluate the efficacy and safety of percutaneous manual aspiration thrombectomy for the treatment of acute massive pulmonary thromboembolism with hemodynamic impairment. MATERIALS AND METHODS: Over a period of 6 years and 9 months, 15 patients with hemodynamic impairment (4 men, 11 women; aged 27-79 years) were treated by manual clot aspiration with a standard, large-lumen percutaneous transluminal coronary angioplasty (PTCA) guiding catheter. RESULTS: After treatment, angiography demonstrated improvement of pulmonary perfusion in all patients (mean Miller score: before treatment 18.9, after treatment 12.1; P < 0.01). Mean pulmonary arterial pressure decreased from 29.6 to 22.5 mmHg (P < 0.01). The mean treatment time was 114.2 min. All of the patients survived and their clinical status improved. No patient had any significant complication. CONCLUSION: Percutaneous manual aspiration thrombectomy with a standard 8 Fr PTCA guiding catheter achieved rapid, safe improvement of the hemodynamic situation in cases of acute massive pulmonary thromboembolism, with low cost both in terms of time and money. PMID- 15287533 TI - Experimental study of the rat for the assessment of barium coating on the gastric mucosa: efficacy of ranitidine and acetylcysteine administration. AB - PURPOSE: In order to improve the preparation method for barium examination of the stomach by ranitidine and acetylcysteine use, the effect on the rat gastric mucosa caused by the administration of ranitidine and acetylcysteine was studied. MATERIALS AND METHODS: Rat stomach that had been treated with ranitidine or acetylcysteine at different intervals and examined in vivo was excised and coated with a barium suspension. A radiograph was subsequently taken and evaluated in regard to the removal of gastric mucus and imaging of the areae gastricae (AG). The removal of mucus was assessed by six blind observers. The imaging of AG was estimated as a percentage of the imaged AG area per total gastric corpus. RESULTS: No change was seen on the radiograph with ranitidine preparation, while the mucus was distinctly removed and AG well-imaged in the group studied 15 minutes after the peroral administration of acetylcysteine. CONCLUSION: Proper preparation for barium study of the stomach should involve treatment with a mucolytic agent about 15 minutes before the examination. H2-blockers must be used supplementally in the short term. PMID- 15287534 TI - Respiratory symptoms in rheumatoid arthritis: relation between high resolution CT findings and functional impairment. AB - PURPOSE: The objectives of this study were to analyze the high resolution computed tomography (HRCT) findings in rheumatoid arthritis (RA) patients with respiratory symptoms and to evaluate the relation between the extent of HRCT findings and functional impairment as assessed by spirometry. MATERIALS AND METHODS: HRCT examination of the thorax and pulmonary function tests (PFTs) were performed in 34 RA patients with respiratory symptoms. Patients with smoking history or with emphysema evident on HRCT were excluded from the study. CT findings were assessed for the presence and pattern of abnormalities. Extent was scored based on the number of pulmonary segments involved. PFTs included forced expiratory flows (FEFs) and forced vital capacity (FVC). RESULTS: Bronchial wall thickening was detected in 29 of 34 RA patients (85%), small nodules in 24 patients (71%), and bronchial dilatation in 21 patients (62%). The extent of bronchial wall thickening correlated with FEF25-75, FEF75, and FEF50 (p<0.0001, respectively) (Spearman's rank correlation). Extent of small nodules correlated with FEF25-75, FEF50, and FEF25 (p<0.01, respectively). Stepwise regression analysis showed independent correlations of bronchial wall thickening with decreases in FEF25-75 and FEF75 (p<0.0001, both). Bronchial dilatation was also independently associated with a decrease in FVC (p<0.05). CONCLUSION: The most common HRCT findings in RA patients with respiratory symptoms are bronchial wall thickening and small nodules, and the extent of these findings correlates significantly with functional impairment. PMID- 15287535 TI - Evaluation of major aortopulmonary collateral arteries (MAPCAs) using three dimensional CT angiography: two case reports. AB - Pulmonary atresia (PA) and ventricular septal defect (VSD) are usually associated with major aortopulmonary collateral arteries (MAPCAs). Preoperative evaluation of MAPCAs is essential for effective planning of unifocalization. Multidetector row computed tomography (MDCT) with three-dimensional volume rendering (3D-VR) demonstrates MAPCAs as well as conventional angiography. We report two cases of PA and VSD associated with MAPCAs examined pre- and postoperatively by means of angiography and MDCT with the 3D-VR technique. MDCT with the 3D-VR technique had the potential to elucidate all MAPCAs demonstrated by invasive angiography. This technique is useful with preoperative conventional angiography as a standard examination for the planning of staged surgery. PMID- 15287536 TI - Carbon monoxide poisoning: two cases of assessment by magnetization transfer ratios and 1H-MRS for brain damage. AB - We describe two patients with carbon monoxide (CO) poisoning. One developed diseased hypoxic encephalopathy, and the other recovered completely. In the patient with progressive hypoxic encephalopathy, the magnetization transfer ratios (MTRs) had already decreased during the lucid period, while the patient with complete recovery showed no reduction of MTRs during this period. 1H-MRS in the two patients revealed no definite abnormality during this lucid period. The MTR maps suggested that irreversible change had already occurred during the lucid period, and 1H-MRS was useful to determine the duration of treatment. The combination of MTRs and 1H-MRS may help to manage patients with CO poisoning. PMID- 15287537 TI - Intrahepatic portosystemic venous shunt: occurrence in a child patient without liver cirrhosis. AB - There are multiple reports of intrahepatic portosystemic venous shunt (PSVS) cases in adult patients. We report the case of a 4-year-old child with PSVS and pulmonary arteriovenous malformation (PAVM). Abdominal sonography and computed tomography (CT) revealed the presence of PSVS. T1-weighted magnetic resonance imaging (MRI) demonstrated multiple intracranial hyperintense lesions, mainly in the globus pallidus, which suggested portosystemic encephalopathy. Tc-99m labeled microsphere study showed diffusely increased uptake in the thyroid and kidneys. The scan suggested the existence of PAVM. Pulmonary angiography was performed in order to evaluate pulmonary hypertension, and pulmonary venous pressure was slightly elevated. Contrast echocardiography suggested the presence of an intrapulmonary arteriovenous malformation with significant right-to-left shunt, as evidenced by rapid filling of the left atrium with dissolved bubbles. In this case, contrast echocardiography was helpful in diagnosing the patient's PAVM. In conclusion, we present the case of PSVS with PAVM in childhood. The incidence of PSVS is low, and data from the literature remain limited. However, further investigation is required to clarify the possible correlation between PSVS and PAVM. PMID- 15287538 TI - Adenoid cystic carcinoma with hyperostosis after stereotactic radiosurgery. AB - Adenoid cystic carcinoma (ACC) of the head and neck is a relatively rare tumor characterized by slow evolution, recurrences, and protracted clinical course. We performed stereotactic radiosurgery in order to palliate symptoms for intraorbital recurrence of ACC that had been treated with total resection of the left parotid gland, radical neck dissection, and conventionally fractionated radiotherapy. We experienced interesting hyperostosis in the area irradiated by the stereotactic radiosurgery. PMID- 15287539 TI - Postoperative mental disorder following prolonged oral surgery. AB - A prolonged period of oral surgery is a potential risk factor of postoperative mental disorders although no such report has been published to date. We retrospectively studied perioperative features in 36 patients who underwent prolonged (10 hours or more) of oral surgery. Patients were categorized as pre delirium (Pre-D) when they manifested 1 or 2 symptoms and delirium (D) when they showed more than 2 symptoms, according to the modified International Classification of Diseases, 10th edition. Of the 36 patients who returned to a normal mental state without drug therapy, 13 were classified as D and 14 were Pre D. A number of patients had moderate complications preoperatively, and massive hemorrhaging occurred during the operation in some Pre-D and D patients. Age was greater in D (62.0 +/- 9.9 years) than in Pre-D (56.0 +/- 13.8 years) patients. Propofol-based general anesthesia was most commonly employed. The time prior to appearance of pre-delirium was significantly shorter in D (30.0 +/- 16.7 hours) than in Pre-D (55.0 +/- 35.0 hours) group patients. Our results indicate that, in general, patients predisposed to postoperative mental disorders have moderate complications preoperatively, are generally older than 50-years-old, receive propofol-based general anesthesia and/or experience a massive hemorrhage during the operation. PMID- 15287540 TI - Prevention of trabecular bone loss in the mandible of ovariectomized rats. AB - The effect of therapeutic agents on trabecular bone loss in the mandible was investigated in ovariectomized rats. Eighty-seven Wistar SPF female rats were ovariectomized (OVX) or given a sham operation (Sham), and maintained on a diet containing 0.1% calcium. Four weeks later, groups of OVX rats were treated with estriol (E3), calcitonin (CT), etidronate, or 2-carboxyethylgermanium sesquioxide (Ge-132). The Basal group was maintained on a diet containing 1.0% calcium, and the OVX and sham groups on a diet containing 0.1% calcium. The trabecular bone mineral density (BMD) and trabecular bone mineral content (BMC) in 11 mandibular slices from 0.5 mm at the mesial margin of the first molar to 0.5 mm at the distal margin of the third molar, were measured using peripheral Quantitative Computed Tomography (pQCT). The BMD in the OVX group was lower than that in the Sham group, and decreased BMC was observed only in the molar region. BMD and BMC were increased in the etidronate-treated group, but only BMC was increased in the CT group. E3 treatment increased BMD and BMC; significant increases were also observed beneath the molar. Ge-132 treatment increased both BMD and BMC, especially the latter. PMID- 15287541 TI - Proliferative activity in oral salivary gland tumors: the role of PCNA and AgNOR assessed by a double staining technique. AB - The aim of this study was to evaluate the role of PCNA and AgNOR in the assessment of salivary gland tumor proliferation using a double staining technique. Ten cases of pleomorphic adenoma (PA) and seventeen cases of adenoid cystic carcinoma (ACC) were examined. Numeric and morphometric parameters of AgNOR were evaluated and compared in PCNA-positive and PCNA-negative nuclei. There were statistically significant differences in AgNOR numbers, perimeters and contour indices between PCNA-positive and -negative nuclei in the PA samples. The ACC samples demonstrated significant differences only in the AgNOR areas. Our results show that in salivary gland tumors there is not always a relationship between proliferative activity evaluated by AgNOR numeric and morphometric parameters and PCNA immunostaining. PMID- 15287542 TI - Monitoring of dnaK gene expression in Porphyromonas gingivalis by oxygen stress using DNA microarray. AB - Porphyromonas gingivalis, a Gram-negative anaerobe associated with adult periodontitis, expresses numerous potential virulence factors. dnaK, a member of the heat shock protein family, functions as a molecular chaperone and plays a role in microbial pathogenicity. However, little is known regarding its gene expression caused by oxygen stress in P. gingivalis. In the present study, a custom-made DNA microarray was designed and used to monitor dnaK gene expression in P. gingivalis caused by oxygen stress. The results demonstrated that dnaK mRNA was up-regulated in a short time, and the DNA microarray results were confirmed by real-time polymerase chain reaction analysis. These findings suggest that oxygen stress stimulates gene expression of dnaK and may have a relationship to the aerotolerance activity of this organism as well as its expression of pathogenesis. PMID- 15287543 TI - Oral colonization of Candida species in perinatally HIV-infected children in northern Thailand. AB - Human immunodeficiency virus (HIV) infection remains a serious problem in northern Thailand. A high prevalence of perinatally HIV-infected children with oral candidiasis has been observed in the region. The objective of this study was to determine oral colonization of Candida spp. in children with perinatal HIV infection. Samples were collected by oral rinse or oral swab from 40 HIV-infected children and from 15 HIV-negative children as a control group. Yeasts recovered in culture were identified and quantified. The mean ages of HIV-infected children and HIV-negative children were 5.5 years (SD = 3.5) and 2.9 years (SD = 2.0) respectively. Eighteen HIV-infected children (45%) had clinical symptoms of oral candidiasis while none of the HIV-negative children had any such symptoms. By culture technique, yeasts were isolated from 28/40 (70%) of the HIV-infected children and 6/15 (40%) of the HIV-negative children. C. albicans was the most common species recovered from HIV-infected and HIV-negative children. Statistically, HIV infection was significantly associated with Candida spp. detection (P-value = 0.04). In contrast, the association between HIV infection and asymptomatic oral carriage of Candida spp. was not significant (P-value = 0.74). These findings demonstrate that oral colonization of Candida spp. is prevalent in HIV-infected children and suggest that prevention and treatment of oral candidiasis is needed for these children. PMID- 15287544 TI - Influence of different vehicles on the pH of calcium hydroxide pastes. AB - The main known benefit of calcium hydroxide as an intracanal medicament lies in the bactericidal effect conferred by its pH. The objective of this work was to determine the influence of the vehicle on the pH of calcium hydroxide pastes after usage in patients and in vitro. The incisor root canals of 180 patients were instrumented and filled with calcium hydroxide pastes containing distilled water, chlorhexidine, propylene glycol, anesthetic solution, camphorated p monochlorophenol and camphorated p-monochlorophenol-propylene glycol. The pH of the paste in the patients' root canals was measured at 7, 14 and 21 days. Similarly, pH was measured in vitro up to 21 days. The pH of all the pastes remained constant throughout the time periods assessed. The calcium hydroxide water combination showed significantly higher pH values than the other pastes in clinical use. Comparative analysis showed that the pH values of the anesthetic solution, camphorated p-monochlorophenol and camphorated p-monochlorophenol propylene glycol were significantly higher in vitro. The type of vehicle was shown to influence the final pH of the pastes. However, the alkalinity of all pastes was maintained over time under the experimental conditions. PMID- 15287545 TI - Masticatory efficiency before and after surgery in oral cancer patients: comparative study of glossectomy, marginal mandibulectomy and segmental mandibulectomy. AB - This study evaluated the effect of oral cancer surgery on masticatory efficiency. Masticatory efficiency was measured using the ATP absorption method. Eating ability was measured using a questionnaire. Two groups were employed as controls: The "normal occlusion group" consisted of subjects who had a complete set of natural maxillary teeth opposed to mandibular teeth, and the "unilateral occlusion group" consisted of subjects who had lost their molar and premolar teeth on one side of the mandible as a result of caries or periodontal diseases. Three treatment groups, each of 6 patients, were studied: a glossectomy group, a marginal mandibulectomy group and a segmental mandibulectomy group. There were no differences in masticatory efficiency between two control groups. Masticatory efficiencies of the three oral cancer treatment groups were lower than in the unilateral occlusion group, even 12 months after surgery. Masticatory efficiency of the glossectomy group was significantly higher 12 months after surgery compared with pre-surgery. Masticatory and eating abilities of the marginal mandibulectomy group and the segmental mandibulectomy were reduced at 3 and 6 months after surgery. The masticatory efficiency 12 months after surgery was higher in the marginal mandibulectomy group than the segmental mandibulectomy group, although the difference was not statistically significant. The self assessed eating ability 12 months after surgery was significantly higher in the marginal mandibulectomy group than the segmental mandibulectomy group. These results suggest that discontinuation of the mandible may lead patients to eat only foods that do not require a substantial amount of chewing. Hence, the quality of life of patients in the marginal mandibulectomy group was considered to be better than that in the segmental mandibulectomy group. PMID- 15287546 TI - Age-related changes in IGF-1 expression in submandibular glands of senescence accelerated mice. AB - Saliva is known to play important roles in such functions as swallowing, mastication, speech, and taste. Furthermore, salivary glands synthesize and secrete a number of growth factors involved in cell/tissue homeostasis. It has been demonstrated that IGF-1, which is structurally analogous to insulin, has been shown to be expressed in mouse submandibular glands, and that IGF-1 stimulates DNA synthesis, amino acid uptake, protein synthesis, and glucose transport in various cells. Diminished function of the salivary glands is thought to lead to increased dental caries and periodontal diseases, which are commonly associated with aging. However, very little is known regarding the effects of age on IGF-1 expression in submandibular glands. The senescence-accelerated mouse (SAM), an experimental murine model of accelerated aging, has been extensively used to examine the mechanisms responsible for aging. In the present study, IGF-1 production and mRNA levels in the submandibular glands of SAM-P1 mice were examined. IGF-1 levels were determined by radioimmunoassay and IGF-1 mRNA levels by semi-quantitative RT-PCR. We found that IGF-1 protein levels in homogenates and IGF-1 mRNA levels decreased with age in SAMP1 mice. These findings suggest that IGF-1 synthesis in submandibular glands decreases with aging, and this may result in lower levels of cellular proliferation, regeneration and wound healing in aged oral tissues. PMID- 15287547 TI - Characteristics and willingness of patients to pay for regular dental check-ups in Japan. AB - The purpose of this survey was to investigate the relationship between demographic characteristics and willingness of patients to pay for regular dental check-ups in Japan. Questionnaires were distributed at private dental offices and collected via postage-paid return envelopes addressed to the center of the study groups. Questions focused on demographics and willingness to pay for regular check-ups. Five thousand one hundred thirty-two questionnaires were collected (response rate 56.8%). The 3 groups most likely to have regular dental check-ups were found to be the under 20s, 50 to 59 year olds and civil servants. Of these groups, civil servants were found to be the most likely of all to have regular check-ups. More females than males were represented in the sample. More than 60% of the patients responded that they would be willing to pay for regular check-ups if the cost were less than 2,000 yen (about 20 dollars). However, no statistically significant differences were observed in relation to household income. The results suggested that participation in regular dental check-ups might be related to gender and age, but not to household income. PMID- 15287548 TI - Mechanical evaluation of effect of grape seed proanthocyanidins extract on debilitated mandibles in rats. AB - Grape seed proanthocyanidins extract (GSPE), whose principal ingredient is proanthocyanidins, shows many activities such as cholesterol lowering effects, antioxidant effects, anti-tumor effects, cardioprotective effects, and protection against ultraviolet rays. However, reports of the effects of GSPE on bone are rare. We performed a mechanical analysis of the effect of GSPE on the interior structure of rat mandibular bone in the growth period, using three-dimensional peripheral quantitative computed tomography (pQCT). A low-calcium/high-calcium diet with supplementary GSPE was compared to a low-calcium/high-calcium diet in rats with debilitated mandibular bones. The group who received added GSPE showed a significant increase in cortical bone density, cross-sectional area, and trabecular bone mineral content (p<0.05). A significant increase was also seen in the results of a non-invasive stress strain index (SSI) (p<0.01) in the added GSPE. Our findings suggest that GSPE can increase bone quality and bone strength of rat mandibles in the growth period. PMID- 15287549 TI - Abutment forms and restorative materials in adhesive prosthesis: a finite element analysis. AB - This study evaluated experimental abutment forms utilizing adhesion for clinical treatment by the three-dimensional finite element method. Three experimental abutment forms with no axial wall were evaluated: Form 1 was the occlusal surface reduced by 1.5 mm, Forms 2 and 3 were the crown cut perpendicular to the tooth axis 2 mm or 4 mm from the central groove. The restorations were made of 3 types of materials: composite resin, porcelain, and a gold-based alloy. Restorations were bonded to the abutments with an adhesive resin. A vertical load of 500 N was applied to the center of the inner incline of the buccal cusp. The stresses in the adhesives were largest in Form 1 with composite resin and in Form 3. It was indicated that the stresses were greatly affected by the form of the abutment and the restorative materials. PMID- 15287550 TI - High temperature characteristics and solidification microstructures of dental metallic materials. Part II. ADAS Type 3 gold alloy. AB - Previously, high temperature properties of the silver-palladium-copper-gold alloy were investigated. In this study, the thermal expansion percentage and coefficient, and high temperature strengths of ADAS Type 3 gold alloy were investigated up to the liquidus temperature. Furthermore, microstructural and compositional changes in the solid/liquid dual phase were studied. The following conclusions were obtained. (1) The solidus point of the Type 3 gold alloy was 899.3+/-11.7 degrees C, and the liquidus point was 962.3+/-2.4 degrees C. (2) The thermal expansion percentage at the solidus point was 1.636+/-0.046%, while it was 4.853+/-0.213% for the liquidus point. The thermal expansion percentage of the melt was 3.217+/-0.257%. (3) The melt expansion was observed even under the measuring pressure of 373.75 HPa, which was quite different from the fact that the melt expansion disappeared at the pressure of 20.87 HPa for the silver palladium-copper-gold alloy. (4) The morphology of solid phase in the solid/liquid dual zone of this alloy was quite different from those observed with the silver-palladium-copper-gold alloy. PMID- 15287551 TI - Bond durability of resin cements to Au-Pd-Ag alloy under cyclic impact load. AB - The bond durability of resin cements to a 12% Au-Pd-Ag alloy was studied through cyclic impact tests with different loads. A piece of casting alloy was bonded to a cast block with two types of resin cements, Super Bond C&B and Bistite II. A shear load was applied onto a small piece of alloy until debonding of the specimen, using different weights of plungers, 130 g, 230 g, 330 g and 430 g. The specimen bonded with Super Bond exhibited a higher resistance than that with Bistite II. The fracture modes of the debonded cements were completely different from each other. That is, Bistite II showed a bulk fracture of cement by the crack penetrating through the cement layer. On the other hand, Super Bond showed damages limited to the surface and no bulk fracture. The mode of fracture was dependent not on the loading weight but the types of resin cements used. PMID- 15287552 TI - Relationship between plasticizer content and tensile bond strength of soft denture liners to a denture base resin. AB - The purpose of this study was to determine the influence of plasticizer content on the tensile bond strength of heat-cured acrylic soft denture liners to a denture base resin. Differences among materials were significant, except for 100 wt% Dibutyl Sebacate (DBS) and 80 wt% DBS of tensile bond strength. The bond strength of all materials to the denture base increased with an increase in thermal cycles significantly except for 40 wt% DBS. The tensile bond strength of soft denture liners to the denture base resin significantly decreased with an increase of plasticizer contents. Differences were found among the difference plasticizer contents in failure types between the denture base resin and soft denture liners. The results suggest that the tensile bond strengths of heat-cured acrylic soft denture liners to the denture base resin were lower with an increase in plasticizer content. PMID- 15287553 TI - Adhesion of 4-META/MMA-TBB resin to collagen-depleted dentin--effect of conditioner with ascorbic acid/ferric chloride. AB - The purpose of this study was to evaluate ascorbic acid (AS) and ferric chloride (FE) for bonding 4-META/MMA-TBB resin to dentin that had been treated with NaClO. An experimental dentin conditioner consisting of 10%AS and 5%Fe (10AS-5FE) and three controls (10AS-0FE, 0AS-5FE, and 0AS-0FE) were prepared. Ascorcic acid neutralizes NaClO. The flattened dentin surfaces were modified sequentially with phosphoric acid etchant, NaClO agent, and the experimental conditioner, then each surface was bonded to a stainless steel rod with 4-META/MMA-TBB resin. The Super Bond C&B (10-3/SB) system was also used. 24-hour tensile bond strengths were determined. The bonding system using 10AS-5FE conditioner showed significantly high bond strength compared to 10AS-0FE, 0AS-5FE, and 0AS-0FE. No significant differences were observed between 10AS-5FE and 10-3/SB. Microphotographs suggested that no hybrid layer formed in the 10AS-5FE group. Although the use of phosphoric acid and NaClO resulted in decreased bond strength between 4-META/MMA TBB resin and dentin, additional conditioning with ascorbic acid and ferric chloride improved the bond strength. PMID- 15287554 TI - Cytotoxicities of a 4-META/MMA-TBBO resin against human pulp fibroblasts. AB - This study was designed to investigate the cytotoxicity of MMA (methyl methacrylate) and 4-META (4-methacryloyloxyethoxycarbonylphthalic anhydride)/MMA with or without TBBO (tri-n-butylborane partially oxide), an initiator and TBBO alone against human pulp fibroblasts (HPF). Cell viabilities were measured by MTT assay. The cytotoxicity of 4-META/MMA-TBBO was comparable with that of MMA-TBBO. TBBO showed higher cytotoxicity than 4-META/MMA. The cytotoxicity induction of a 4-META/MMA-TBBO resin may be preferably associated with TBBO. PMID- 15287555 TI - Corrosion behavior and microstructures of experimental Ti-Au alloys. AB - Anodic polarization was performed in 0.9% NaCl and 1% lactic acid solutions to characterize the relationship between the corrosion behavior and microstructures of cast Ti-Au (5-40%) alloys. An abrupt increase in the current density occurred at approximately 0.6 V vs. SCE for the 30% and 40% Au alloys in the 0.9% NaCl solution. The microstructures after corrosion testing indicated that this breakdown may have been caused by the preferential dissolution of the Ti3Au. However, the potential for preferential dissolution was higher than the breakdown potential of stainless steel or Co-Cr alloy, which meant that the corrosion resistance of the Ti-Au alloys was superior. In 1% lactic acid solution, the corrosion resistance of the Ti-Au alloys was excellent, with no breakdown at any composition. In the present test solutions, the Ti-Au alloys up to 20% Au had good corrosion resistance comparable to that for pure titanium. PMID- 15287556 TI - Bond strength of permanent cements in cementing cast to crown different core build-up materials. AB - The purpose of this laboratory investigation was to evaluate the bond strength of permanent cement (Duo-cement Kit, Meron, Durelon) to commonly used core build-up materials (President, Dyract AP, Ionofil, Vitremer). Sixty specimens (five of each product) were fabricated as a canine core build- up. Full crown castings were made to fit each core specimen. Full crown castings were cemented to core samples and stored at 37 degrees C and 100% humidity for 10 days. After storage, the bond strength was measured with a Haunsfield tensometer in tensile mode at a crosshead speed of 5 mm/min. Statistical evaluation was performed with univariate analysis of variance (P<0.001). The cement types affected the bond strength of full crown castings to core materials (F: 14.80; P<0.001). The interaction between the cement and core materials was significant (F: 3.69; P<0.01). According to the Duncan's test it was found that the values of Duo-cement were statistically different from the other cements. PMID- 15287557 TI - 3D shape measurement of dental casts using medical X-ray CT. AB - Three-dimensional (3D) digitizing and computerization of dental casts is a trend in dentistry especially for orthodontics to substitute stone casts. Generally used laser scanners have a blind side in the measurement of undercuts. As alternative equipment that can digitize regardless of the undercut, the potential of recent multi-slice medical CT was examined. In 3D shape reconstruction, the CT window level affects the size of the object. It was examined, and a CT window level of 800 was found to be suitable. However, the size became slightly smaller than the real object. Then, a correction ratio of 1.002, 1.015 and 1.013 on the X , Y- and Z-axis was given, and error within 0.08% was accomplished. The measurement and 3D imaging of dental casts was completed within 10 min. The reproducibility of the complicated morphology of dental casts was slightly inferior to that of the latest laser scanners, but the accuracy and operationality regardless of the undercut is noteworthy for clinical application. PMID- 15287558 TI - Development of casting investment preventing blackening of noble metal alloys part 3. Effect of reducing agent addition on the strength and expansion of the investments. AB - Different reducing agents (B, Al, Si and Ti) were individually added to two gypsum-bonded investments to prepare investments preventing surface blackening of some noble cast alloys. The effect of different additive contents on green-body and burnout compressive strength, setting and thermal expansion of the investments were evaluated. The strength and expansion of the investments were changed by the additives. The compressive strength of Al-, Si- and Ti-added investments decreased with the increase of additive contents. The burnout strength of B-added investments significantly increased while green-body strength remained unchanged. The setting expansion of the B-added investments increased while those of the Al-, Si- and Ti-added investments decreased with the increase of additive contents. The thermal expansion of the Si- and Ti-added investments decreased, and that of the Al- and B-added investments remained unchanged. Further study is necessary to evaluate the effects of these additives on the accuracy of dental castings. PMID- 15287559 TI - Effects of repeated baking on the mechanical and physical properties of metal ceramic systems. AB - This study evaluates effects of repeated baking processes on the mechanical and physical properties of single and triple applications of opaque, body and enamel porcelains fused to three different metal substrates (precious metal, semi precious metal and non-precious metal). The vintage halo porcelain system was employed and fused to metals. Fused samples were subjected to three-point bend tests to evaluate bend strength and modulus of elasticity. It was found that, by increasing repeated baking cycles, (1) body and enamel porcelains increased bend strengths but opaque porcelain did not show any changes, (2) all triple-layered porcelains fired to metals increased bend strengths, and (3) all three porcelains and metal substrates did not exhibit changes in thermal expansion percentage. It was concluded that repeating baking procedures up to 10 cycles did not exhibit any adverse effects on the final properties of porcelain-fired to metals, rather it was noticed that mechanical strengths increased by increasing cycles. PMID- 15287560 TI - Effect of adding spherical silica filler on physico-mechanical properties of resin modified glass-ionomer cement. AB - This study investigated the effects of spherical silica fillers on the physical and mechanical properties of resin-modified glass-ionomer cement (RMGIC). Specimens were fabricated by mixing untreated (UF) or silanized (SF) spherical silica filler into the powder of a commercially prepared RMGIC. The original RMGIC and a preparation containing 20 wt% spherical silica filler were also examined with regard to their fractured surface and fluoride release. The fillers increased the compressive strength remarkably: up to 17% in the case of SF and 9% in the case of UF. Both UF and SF increased the flexural strength by up to 17%. The addition of SF increased the DTS up to 38%, but UF decreased the DTS. The addition of SF improved the workability and the mechanical properties of the RMGIC. PMID- 15287561 TI - Optical and color stabilities of paint-on resins for shade modification of restorative resins. AB - The purpose of this study was to examine the optical and color stabilities of the paint-on resin used for shade modification of restorative resins. Three shades of paint-on resin and two crown and bridge resins were used. The light transmittance characteristics of the materials during accelerated aging tests such as water immersion, toothbrush abrasion, ultraviolet (UV) light irradiation, and staining tests were measured. Discolorations of materials resulting from tests were also determined. There were no significant effects of water immersion, toothbrush abrasion and UV light irradiation on the light transmittance and visible color change of paint-on resins, whereas the staining tests significantly decreased the light transmittance and increased color change of the translucent shades of materials. Our results indicate that the paint-on resins exhibit stable optical properties and color appearance, which are at least as good as the crown and bridge resins. PMID- 15287562 TI - Polymerization characteristics of ethyl methacrylate-based resin initiated by TBB. AB - Polymerization characteristics of 4 types of ethyl methacrylate (EMA)-based resin, composed of EMA and EMA/methyl methacrylate (MMA) copolymers and initiated by tributylborane (TBB), were studied from the aspects of long term changes of residual monomer (RM) and molecular weight (Mw). The resins were polymerized at room temperature for 30 min, stored at 37 degrees C for 24 hr, 1 and 4 weeks and then analyzed by high performance liquid chromatography and size exclusion chromatography to determine RM and Mw. RM decreased significantly from 4.47 11.61% after 30 min to 0.94-7.73%, 0.81-7.81%, and 0.78-7.10% after 24 hr, 1 and 4 weeks, respectively, demonstrating a tendency to decrease with time for each resin. Mw showed a tendency to decrease from 427 x 10(3)-551 x 10(3) after 30 min to 354 x 10(3)-530 x 10(3) after 4 weeks. TBB-initiated EMA resin had polymerization characteristics similar to those for TBB-initiated MMA resin in the temporal changes in RM and Mw during postpolymerization. PMID- 15287563 TI - Fatigue strengths of particulate filler composites reinforced with fibers. AB - The aim of this study was to evaluate the dynamic fatigue strengths at 10(5) cycles and the strains of particulate filler composite resins with and without reinforcing fibers. An UHMWPE (Ribbond), a polyaromatic polyamide fiber (Fibreflex), and three glass fibers (GlasSpan, FibreKor, Vectris Frame) were used to reinforce the particulate filler composite resins. The fatigue properties were measured in three-point bending mode using a servohydraulic universal testing machine at a frequency of 5 Hz, until failure occurred or 10(5) cycles had been completed. The fatigue strengths at 10(5) cycles were determined by the staircase method. The fractured aspects of specimens were evaluated by an optical and scanning electron microscope. The fatigue strengths of particulate filler composite resins were 49-57 MPa, and those of fiber-reinforced were 90-209 MPa. Unidirectional glass fibers showed higher reinforcing effects on the fatigue strengths of composite resins. The strain of UHMWPE-reinforced composite was largest. PMID- 15287564 TI - Effects of heat and pH in silanation process on flexural properties and hydrolytic durabilities of composite resin after hot water storage. AB - The objectives of this study were to evaluate the effect of heat application after silanation and the pH effect of silanated solution on flexural properties and hydrolytic durability of experimental composites. A spherical-shaped filler was silanated with 0.66 mass% of gamma-MPTS in 70 vol% ethanol solution. For the heating effect, the filler was heated after silanation for 3 hours at 50 degrees C, 80 degrees C and 110 degrees C. For the effect of pH, ethanol solution with HCl solution of pH 1, 2 and 4 were used. All fillers were mixed to make photopolymerized experimental composite resins. All specimens after immersion in 37 degrees water for 24 hours were additionally immersed in Soxhlet's extractor for another 1, 7 and 28 days. A three-point bending test was performed in 37 degrees C water. The heat application of 50 degrees C and 80 degrees C increased flexural strength while heat of 110 degrees C and pH showed no effects on flexural properties at p<0.05. PMID- 15287565 TI - Application of alumina coping to porcelain laminate veneered crown: part 1 masking ability for discolored teeth. AB - A densely sintered high-purity alumina has been successfully utilized as a coping for all-ceramic crown. In order to apply the alumina coping to the porcelain laminate veneer restoration for the discolored teeth, the present study evaluated its masking ability when it was thinner than the proposed thickness for the crown. Colorimetric examination was performed on white and black backgrounds for the 0.7 mm thick porcelain laminate veneer with 0.40 or 0.25 mm thick alumina coping and 0.7 mm thick porcelain without coping. With the presence of the coping, the porcelain appeared significantly lighter. Judging from the calculated color differences delta E and the literature, it was suggested that the masking ability of the alumina coping would be sufficient with a 0.25 mm thickness for the porcelain laminate veneer for heavily discolored teeth. PMID- 15287566 TI - Evidence that HSP70 gene expression may be useful for assessing the cytocompatibility of dental biomaterials. AB - In the current studies, we examined the possibility of using HSP70 gene regulation as a cytocompatibility test for dental biomaterials. For this reason, we assessed the effects of three metal salts, HgCl2, CuSO4 and NiCl2 on HSP70 gene expression in HeLa S3 cells using real-time Taqman quantitative PCR. Incubation of the cells for 4 h in medium containing HgCl2 (20 or 40 microM), CuSO4 (157, 313, 625 or 1250 microM) or NiCl2 (5000 and 10000 microM) significantly induced HSP70 mRNA. The real-time Taqman quantitative PCR was able to detect HSP70 mRNA induction at 4-fold lower concentrations of HgCl2 and 8-fold lower concentrations of CuSO4 than the Neutral Red cell viability assay. These results indicate that real-time Taqman quantitative PCR, in combination with the monitoring of cell viability, may be a valuable tool for distinguishing between specific HSP70 mRNA induction and cytocompatibility of metals in dental biomaterials. PMID- 15287567 TI - Durability of tungsten carbide burs for the fabrication of titanium crowns using dental CAD/CAM. AB - The purpose of this study was to evaluate the durability of tungsten carbide burs for the fabrication of titanium crowns using two dental CAD/CAM systems (DECSY, Digital Process, Japan and Cadim, Advance, Japan). A tungsten carbide bur in each system was examined and used without fracture to fabricate 51 titanium crowns. For both systems tiny chips were found on the bur blade at the 11th machining. These chips gradually enlarged as the number of machining times increased. At the first machining no significant difference in the average surface roughness was found on the crown between the two systems (1.6 microm for DECSY and 1.2 microm for Cadim). The cutting grooves became dull and the average surface roughness increased as the number of machining times increased. It is concluded that the tungsten carbide burs for both systems can be used to fabricate up to 51 titanium crowns. PMID- 15287568 TI - In vitro corrosion characteristics of commercially available orthodontic wires. AB - The corrosion characteristics of orthodontic alloy wires were investigated both in as-received and grinded conditions in 0.9% NaCl solution by atomic absorption spectrophotometry and potentiodynamic polarization measurements. The amount of each metal ion released from most alloys was larger for the grinded wires than for the as-received wires (p<0.01). The fact that the beta-Ti alloy wire (Ti-Mo Zr) does not contain allergenic metals such as Ni, Co, and Cr, and the finding that resistance to both general and localized corrosion is the highest among the six wires investigated suggest that this wire is the most biocompatible orthodontic wire. Since a small amount of Ni, Cr or Co ions were released from Ni Ti, Co-Cr and stainless steel wires, special attention should be paid during their clinical use for patients with allergic tendencies. PMID- 15287569 TI - Mechanical properties and grindability of experimental Ti-Au alloys. AB - Experimental Ti-Au alloys (5, 10, 20 and 40 mass% Au) were made. Mechanical properties and grindability of the castings of the Ti-Au alloys were examined. As the concentration of gold increased to 20%, the yield strength and the tensile strength of the Ti-Au alloys became higher without markedly deteriorating their ductility. This higher strength can be explained by the solid-solution strengthening of the a titanium. The Ti-40%Au alloy became brittle because the intermetallic compound Ti3Au precipitated intensively near the grain boundaries. There was no significant difference in the grinding rate and grinding ratio among all the Ti-Au alloys and the pure titanium at any speed. PMID- 15287570 TI - Mold filling and microhardness of 1% Fe titanium alloys. AB - We examined the mold filling capacity and microhardness of two industrial 1% Fe titanium alloys: Super-TIX800 (Nippon Steel Corp.) (Fe: 0.910%, O: 0.370%, N: 0.005%) and Super-TIX800N (Nippon Steel Corp.) (Fe: 0.960%, O: 0.300%, N: 0.041%). Two wedge-shaped acrylic patterns (with 30 degrees or 15 degrees angles) were prepared. Each alloy was cast in a centrifugal casting machine. Mold filling was evaluated as the missing length between the tip of the casting and the theoretical tip. Vickers hardness of the edge of the castings was also determined. For both angles tested, there were no significant differences (p>0.05) in mold filling among these alloys and the control (CP Ti). The results of testing the microhardness near the cast surfaces indicated that the hardened reaction layers on these alloys were thinner at the edge compared to CP Ti. PMID- 15287571 TI - Dimensional changes of ring-shaped pattern. AB - Ring shaped wax patterns, having the same outside diameter and different inside diameters, were invested with a gypsum-bonded cristobalite investment. The wax pattern was eliminated in an electric furnace at 120 degrees C. A fusible alloy with a melting point of 47 degrees C was cast at room temperature. The dimensional deviations between the fusible alloy casting and the wax pattern were calculated using the inside diameter, ring width and outside diameter. On the other series, a gold alloy casting of the same size was fabricated in the usual manner of the dental precise casting procedure, and the dimension was compared with that of the wax pattern. In the comparison of 2 types of patterns, dimensional change by setting expansion was different. Dimensional change of the small inside diameter specimen differed at 3 portions measured, but that of the large inside diameter specimen was comparable at 3 portions. Concerning the resultant gold alloy casting, dimensional change at the outside diameter differed from each other, but those at ring width and inside diameter were comparable to each other. The difference in the inside diameter influenced dimensional change by setting expansion as well as that of the resultant casting. PMID- 15287572 TI - Toothbrush abrasion of paint-on resins for shade modification and crown resins: effect of water absorption. AB - In order to investigate the clinical application of paint-on resins, the effect of water absorption on toothbrush abrasion and light transmittance of ten crown resins including three paint-on resins was examined. Water absorption into each material ranged from 0.29 to 0.89 mg/cm2 after storage in distilled-water for 6 weeks and their hardnesses decreased by 3.5-22.3%. Maximum surface roughness (Rmax) of the materials stored in distilled water for 6 weeks increased with an increasing number of toothbrush abrasion cycles and ranged from 1.9 to 10.5 microm after 100,000 cycles. Also, Maximum depth and weight loss as an indicator of the amount of each material lost by abrasion showed similar behaviors similar to Rmax. These results indicated that the abrasion resistance of paint-on resins was located in the middle among all materials examined. PMID- 15287573 TI - Gypsum-bonded investment and dental precision casting (III) Composition of investment for the quick casting technique. AB - A simultaneous differential thermal analysis and thermogravimetry method previously established was used to estimate the composition of gypsum-bonded investment marketed for the quick casting technique. Three commercial investments of this type were heated to 700 degrees C at 10 degrees C/min and the hemihydrate content was estimated by the mass decrease reached at 300 degrees C after subtracting the mass decrease at 100 degrees C as moisture content. The hemihydrate contents were between 25% and 30%, which appears to be the range also chosen for the conventional gypsum-bonded investment of cristobalite type over 70 years by the industry. However, the new type of investment contained both cristobalite and quartz. The small sample size is a disadvantage of the present method but this can be overcome by more frequent use of the method by investigators. PMID- 15287574 TI - Comparative study of water sorption and solubility of soft lining materials in the different solutions. AB - In this study, two acrylic based materials and three silicone rubber soft lining materials were investigated to determine the percentage of absorption and solubility in artificial saliva, distilled water, and denture cleanser. In addition, the effect of denture cleanser on surface properties of soft lining materials was also evaluated. For sorption and solubility testing, 75 discs (50 mm x 0.5 mm) were prepared and divided into 5 groups with 15 samples in each group. The specimens were stored in different solutions, and tested after 1, 4, and 16 weeks. Analysis of variance was used to find the significant differences between the materials at all time intervals. The acrylic resin soft lining materials had higher solubility (3.432% Visco-gel in artificial saliva) and absorption (3.349% Visco-gel in distilled water) than Molloplast-B after 16 weeks of aging. The greatest hardness and color change were shown in the acrylic soft lining materials. PMID- 15287575 TI - [The compatibility of ronidazol in racing pigeons in a controlled clinical study]. AB - Ronidazol is often used in racing pigeons for the treatment of Trichomonas infections and diseases. Therefore, in this study, the compatibility of the drug was examined by oral application over 7 days. For this purpose a randomized blind study was performed using four different groups (control group, 10 mg = therapy group, 20 mg = double-dose-group and 40 mg = high-dose-group) of pigeons (Columba livia f. domestica) with 6 male and 6 female birds each. All birds were clinically healthy and between 6 and 12 weeks of age. The application of ronidazol at a dose of 10 mg/racing pigeon did show no side-effect within the duration of the study, e.g. no influence could be seen on clinical, haematological, blood-chemical and pathological parameters. Low-to middle-grade clinical alterations of the gastro-intestinal tract occurred in the high-dose group at day 6 and 7 of the application of the drug. Therefore a fourfold overdosing of ronidazol should be avoided. PMID- 15287576 TI - [Establishment of an immunological labelling of pigs using synthetic peptides]. AB - The objective of this study was to develop a suitable combination consisting of synthetic peptides, carrier protein and adjuvant for immunological labelling of pigs. Specific antibody titres were evaluated by ELISA technique. From 9 peptides 4 were excluded from following investigations showing cross reactivity or low immunogenic effects, sufficient anti-peptide titres were achieved by 5 peptides. Labelling control is possible after 7 days at the earliest and can be used for the whole fattening period after single immunization using an effective carrier adjuvant-combination. Mixing of peptides in one labelling dose had no negative influence on titres against single peptides. Different combinations of 4 carriers (keyhole limpet haemocyanin (KLH), palmitic-acid-3-cystein-acid (Pam3Cys), palmitic acid (Pam) and dextran) and 4 adjuvants (Montanide IMS 1313, Montanide ISA 25, Quil A and Diluvac forte) were tested. Optimal labelling could be seen by combination of 50 nmol peptide, KLH as carrier and Montanide IMS 1313 as adjuvant. Generally after booster injection titres were higher, however, a booster dose was not necessary using most effective adjuvants. A less immunogenic, but cost effective alternative for short time labelling (7 weeks) was a peptide-Pam3Cys-conjugate (75 nmol) combined with Quil A as an adjuvant (2 mg/ml). Immunological labelling of pigs is recommended as a good method for tracing back the origin of animals and meat products. It may be also used for vaccine labelling to prove vaccination of pigs. PMID- 15287577 TI - [Results of parasitological examinations of faecal samples from horses, ruminants, pigs, dogs, cats, hedgehogs and rabbits between 1998 and 2002]. AB - The results of coproscopical examinations in horses, ruminants, pigs, dogs, cats, hedgehogs and rabbits between 1998 and 2002 are presented. In 4399 samples from horses 37.4% stages of strongylids, 1.4% anoplocephalids, 1.3% Strongyloides westeri, 0.9% Parascaris equorum, 0.04% Oxyuris equi, 0.04% Eimeria sp. and 0.04% Fasciola hepatica were found. In 998 samples of cattle 22.1% stages of strongylids, 11.2% of Eimeria spp., 3.5% of cryptosporidium, 2.9% of Moniezia spp., 1.3% of Trichuris spp., 0.7% of Dictyocaulus sp., 0.6% of Fasciola hepatica, 0.6% of Strongyloides sp., 0.5% of Nematodirus spp. and 0.4% of Capillaria sp. could be detected. In 524 samples of sheep 60.7% eggs of strongylids, 43.1% oozysts of Eimeria spp., 11.1% stages of Nematodirus spp., 9.5% of Moniezia spp., 7.8% of Trichuris spp., 6.7% of Strongyloides sp., 1.7% of Fasciola hepatica, 1% of Capillaria spp., 0.4% of protostrongylidae, 0.2% of Skrjabinema sp. and 0.2% of Dictyocaulus sp. were found. 33.9% of the 118 samples of goats that were examined were positive for oocysts of Eimeria spp., 30.5% for eggs of strongylids, 6.8% for Nematodirus spp., 4.2% for Trichuris spp., 3.4% for Moniezia spp., 0.8 for protostrongylids and 0.8% for Strongyloides sp. 5.7% of 1427 samples of pigs contained stages of strongylids, 1.5% of Ascaris suum, 0.4% of Isospora, 0.3% of Eimeria spp., 0.3% of Trichuris sp., 0.1% of Giardia sp., 0.1% of cryptosproidium as well as 0.1% of metastrongylids. In 1281 of the samples of dogs 2.3% Giardia sp., 2.3% Isospora sp., 2.2% Toxocara canis, 1.4% ancylostomids, 0.8% taeniids, 0.6% larvae of Crenosoma sp., 0.2% Capillaria sp, 0.2% Trichuris vulpis and 0.2% Hammondia-like oocysts were found. In 441 samples of cats 10.7% stages of Isospora sp., 3.9% eggs of Toxocara cati, 1.6% of ancylostomids, 1.4% of taeniids, 1.1% of Giardia sp., 0.7% of Toxoplasma-like oocysts, 0.7% of Aelurostrongylus abstrusus, 0.5% of Toxascaris leonina and 0.2% of Capillaria spp. were found. Furthermore 0.2% of the samples contained proglottids of Mesocestoides and 0.2% stages of Dipylidium sp. Eggs of Capillaria sp. were found in 33% of the 106 samples of hedgehogs, larvae of Crenosoma striatum in 27.4%, oocysts of Isospora sp. in 5.7% of the cases. In 232 samples of rabbits 56.9% oocysts of Eimeria sp., 4.8% stages of Passalurus ambiguus, 1.3% of strongylids, 0.9% of Strongyloides sp., 0.4% of trematodes were found. PMID- 15287578 TI - [Disinfectant tests at 20 and 10 degrees C to determine the virucidal activity against circoviruses]. AB - To evaluate virucidal activity against porcine circovirus type 2 (PCV2), four disinfectants were tested under laboratory conditions. As basis to perform the testing the "Guidelines for testing chemical disinfectants" of the German Veterinary Association (DVG-guidelines) were applied. For simulation of field conditions, the tests were carried out in virus carrier tests, at 20 and 10 degrees C, and under protein load (40% foetal calf serum (FCS) in virus suspension). For disinfection of PCV2 at 20 degrees C an exposure time of 120 min in 2% Disinfectant 1 (20% glutaraldehyde, 12% 2-propenal, polymer with formaldehyde) or Disinfectant 2 (55% formic acid, 7% glyoxylic acid) was necessary. 1% of Disinfectant 3 (Component 1: Potassium peroxomonosulphate. Component 2: Active detergents) disinfected PCV2 on carriers within 180 min. After a reaction time of 120 min with 1% and 60 min with 2% Disinfectant 4 (21% glutaraldehyde, 17% formaldehyde) there could not be detected any virus. Reduction on effectivity through temperature reduction to 10 degrees C were more significant for aldehyde containing preparations Disinfectant 1 and Disinfectant 4 than for Disinfectant 2 and Disinfectant 3. These losses on effectivity could be corrected through extension of exposure time or increase of concentration. PMID- 15287579 TI - [Intersexuality in dogs: causes and genetics]. AB - Failures in the establishment of chromosomal, gonadal and phenotypic sex can cause intersexuality in dogs. Thus, diagnosis of chimaerism, mosaicism, sex reversal syndrome, and male or female pseudohermaphroditism in intersex individuals has to be based on the inspection of the chromosomes, gonads and the phenotypic appearance of the reproductive organs. In a study over two years, seven dogs of different breeds suspected to be intersexes were cytogenetically investigated. A sry-negative XX-sex reversal syndrome was diagnosed in a Jack Russel Terrier. In a mixbred dog a persistent Mullerian duct syndrome (PMDS) was found and a Border Terrier Dog showed an XX/XY chromosomal chimaerism. In further four dogs of different breeds, a female constitution of sex chromosomes was seen. As a sign of intersexuality each of these dog showed an enlarged clitoris. A differentiation between XX-sex reversal syndrome and female pseudohermaphroditism was not possible because there was no information on the internal genital tract and gonads available. PMID- 15287580 TI - [Prevalence of anti-Yersinia antibodies in European brown hares in North-Rhine Westphalia, Germany]. AB - Yersinia (Y.) pseudotuberculosis infections may lead to significant lethality in European brown hare (Lepus europaeus, Pallas) populations especially during the cold and wet seasons. In recent decades, also Y. enterocolitica was isolated from hares found dead. Consequently, a Western-blot technique proved to be valuable for the detection of antibodies against all pathogenic Yersinia isolates was applied to monitor the prevalence of antibodies in hare populations in North Rhine Westphalia, Germany. A total of 89.6% of the 230 animals tested was seropositive. Further investigations should be performed to elucidate the role of subclinical yersiniosis in the decline of European brown hare populations in Germany. PMID- 15287581 TI - Proteomics approach reveals novel proteins on the surface of malaria-infected erythrocytes. AB - Proteins on the surface of parasite-infected erythrocytes (PIESPs) have been one of the major focuses of malaria research due to their role in pathogenesis and their potential as targets for immunity and drug intervention. Despite intense scrutiny, only a few surface proteins have been identified and characterized. We report the identification of two novel surface proteins from Plasmodium falciparum-infected erythrocytes. Surface proteins were fractionated through biotin-streptavidin interaction and analyzed by shotgun proteomics. From a list of 36 candidates, two were selected for further characterization. The surface location of both proteins was confirmed by confocal microscopy using specific antibodies. PIESP1 and PIESP2 are unlikely to be associated with knobs, the protrusions on the parasite-infected erythrocyte (PIE) surface. In contrast to other known PIESPs, such as PfEMP1 and Rifin, these novel proteins are encoded by single copy genes, highly conserved across Plasmodium ssp., making them good targets for interventions with a broad specificity to various P. falciparum isolates. PMID- 15287582 TI - Type I signal peptidase from Leishmania is a target of the immune response in human cutaneous and visceral leishmaniasis. AB - The gene encoding type I signal peptidase (Lmjsp) has been cloned from Leishmania major. Lmjsp encodes a protein of 180 amino residues with a predicted molecular mass of 20.5 kDa. Comparison of the protein sequence with those of known type I signal peptidases indicates homology in five conserved domains A-E which are known to be important, or essential, for catalytic activity. Southern blot hybridisation analysis indicates that there is a single copy of the Lmjsp gene. A recombinant SPase protein and a synthetic peptide of the L. major signal peptidase were used to examine the presence of specific antibodies in sera from either recovered or active individuals of both cutaneous and visceral leishmaniasis. This evaluation demonstrated that sera from cutaneous and visceral forms of leishmaniasis are highly reactive to both the recombinant and synthetic signal peptidase antigens. Therefore, the Leishmania signal peptidase, albeit localised intracellularly, is a significant target of the Leishmania specific immune response and highlights its potential use for serodiagnosis of cutaneous and visceral leishmaniasis. PMID- 15287583 TI - Functional properties of Schistosoma mansoni single-stranded DNA-binding protein SmPUR-alpha. Description of the interaction between SmPUR-alpha and SMYB1. AB - PUR-alpha is a highly conserved protein in eukaryotes belonging to the family of single-stranded DNA-binding proteins. Because PUR-alpha is a multifunctional protein that participates in several regulatory events at the level of gene transcription, it became relevant to investigate the structural features of Schistosoma mansoni PUR-alpha (SmPUR-alpha) that could be correlated to its mode of action. Using deletion constructs on a dot blot assay we mapped the domains of GST-SmPUR-alpha fusion protein involved in the interactions with DNA and RNA. Individually, the N-terminal amino acid residues 1-26 and the C-terminal residues 196-276 of GST-SmPUR-alpha which did not contain nucleic acid-binding domains, did not bind ssDNA or RNA. In contrast, domains encompassing the N-terminal and Class I and C-terminal plus Class I exhibited the highest binding affinity. Seemingly, the latter (GST-SmPUR-alpha 174-276) played a major role in nucleic acid interaction as judged by affinity alone. Other combinations of the deletion constructs displayed either intermediary or no binding affinity to the DNA or RNA probes. Gel shift competition assay showed that GST-SmPUR-alpha bound to ssDNA with higher affinity than to RNA. Because SmPUR-alpha contains two putative phosphorylation sites the protein was tested as a substrate to casein kinase II. GST-SmPUR-alpha could be phosphorylated in vitro by casein kinase II at both, the N- and C-terminus of the protein. The multifunctional nature of SmPUR-alpha was demonstrated by experiments measuring the physical interaction between SmPUR alpha and the transcription factor SMYB1. This was determined in vivo (yeast two hybrid) and in vitro (GST-pull down). Furthermore, we showed that SmPUR-alpha and SMYB1 acted synergistically to bind preferentially to pyrimidine-rich sequences. PMID- 15287584 TI - Signal transduction through the Gal-GalNAc lectin of Entamoeba histolytica involves a spectrin-like protein. AB - Capping followed by uroid formation in Entamoeba histolytica has been implicated in resistance against the host immune response during development of amoebiasis. The amebic actomyosin cytoskeleton is essential for such a process. A protein from the spectrin family co-localizes with the Gal-GalNAc lectin during capping of this surface protein complex. Co-localization is not observed when capping of the Gal-GalNAc lectin is specifically inhibited by production of the carboxyl terminal region of its heavy chain that includes the lectin cytoplasmic tail. A peptide encompassing the lectin last 77 amino acids fused to glutathione-S transferase interacts in vitro with purified spectrin. The spectrin-binding site was narrowed down to a stretch of 21 amino acids within the lectin cytoplasmic domain. This is the first report identifying an amino acid sequence involved in the interaction between the Gal-GalNAc lectin and cytoskeletal spectrin. PMID- 15287585 TI - The Trypanosoma brucei reference strain TREU927/4 contains T. brucei rhodesiense specific SRA sequences, but displays a distinct phenotype of relative resistance to human serum. AB - The Trypanosoma brucei reference strain TREU927/4 exhibits some resistance to lysis by normal human serum (NHS), but this resistance is never complete even after selection. The genome of this strain contains a minimum of eight sequences related to the T. brucei rhodesiense-specific serum resistance-associated gene (SRA), which encodes a truncated variant surface glycoprotein (VSG) conferring full resistance to lysis by NHS. We selected two sequences showing the highest similarity to SRA and also preceded by a region ("cotransposed region") present immediately upstream from SRA in the VSG expression site termed R-ES, where SRA is expressed in T. brucei rhodesiense. Whereas one of these sequences appears to be a pseudogene, the other, which is contained within a cluster of VSG basic copies (BCs), encodes a VSG truncated in the C-terminal domain. In the latter gene, an inserted region encoding surface-exposed loops similar to those of the BoTat 1.20 VSG interrupts the full SRA sequence. Therefore, this gene was termed SRA-BC, for the putative VSG basic copy from which SRA was derived. Neither this gene nor other SRA-like sequences appeared to be responsible for the relative resistance of TREU927/4 to NHS, since (i) transfection of SRA-BC in T. brucei brucei did not confer increased resistance; (ii) SRA transcripts could not be detected in this strain, even when focusing the search on the limited SRA sequence necessary to confer resistance and when using strain-specific SRA probes on RNA from cells selected in the presence of NHS. PMID- 15287586 TI - Trichinella spiralis secretes a homologue of prosaposin. AB - Infective larvae and adult stage Trichinella spiralis secrete a protein homologous to prosaposin, the precursor of sphingolipid activator proteins (saposins) A-D originally defined in vertebrates. The protein contains four saposin domains, with the six cysteine residues which form the three intramolecular disulphide bonds in close register in each case. It differs substantially from vertebrate prosaposins in the N-terminal prodomain, the region separating saposins A and B, and completely lacks the C-terminal domain which has been demonstrated to be essential for lysosomal targetting in these organisms. The protein is secreted in unprocessed form with an estimated mass of 56 kDa, and contains a single N-linked glycan which is bound by the monoclonal antibody NIM M1, characteristic of the TSL-1 antigens which are capped by tyvelose (3,6 dideoxy-D-arabinohexose). Immuno-electron microscopy localised the protein to membrane-bound vesicles and more complex multi-lamellar organelles in diverse tissues including the hypodermis, intestine and stichosomes, although it was absent from the dense-core secretory granules typical of the latter. Possible functions of a secreted prosaposin are discussed. PMID- 15287587 TI - Population dynamics of Wolbachia bacterial endosymbionts in Brugia malayi. AB - The human filarial nematode Brugia malayi contains an endosymbiotic bacterium, Wolbachia. We used real-time quantitative polymerase chain reaction (QPCR) and microscopy to investigate the population dynamics of the bacterium-nematode association. Two Wolbachia (wsp and ftsZ) and one nematode (gst) genes were amplified from all life-cycle stages of B. malayi and results expressed as gene copies per worm and as Wolbachia/nematode ratios. Since the genes were single copy and there was one genome per Wolbachia, the gene copy numbers were equivalent to the numbers of bacteria. These were similar in microfilariae and the mosquito-borne larval stages (L2 and L3), with the lowest ratios of Wolbachia/nematode DNA. However, within 7 days of infection of the mammalian host, bacteria had increased 600-fold and the bacteria/worm ratio was the highest of all life-cycle stages. The rapid multiplication continued throughout L4 development, so that the major period of bacterial population growth occurred within 4 weeks of infection of the definitive host. Microscopy confirmed that there were few bacteria in mosquito-derived L3 but many, in large groups, in L4 collected 9 and 21 days after infection. In adult male worms up to 15 months of age, the bacterial populations were maintained, whilst in females, bacteria numbers increased as the worms matured and as the ovary and embryonic larval stages became infected. These results support the hypothesis that the bacteria are essential for larval development in the mammalian host and for the long-term survival of adult worms. PMID- 15287588 TI - Purification and molecular cloning of a major allergen from Anisakis simplex. AB - A heat-stable allergen with a molecular weight of 21 k was purified from larvae of the nematode Anisakis simplex by gel filtration, anion-exchange FPLC and reverse-phase HPLC. When analyzed by immunoblotting and ELISA, seven of eight patient sera reacted to the 21 k allergen, demonstrating that this protein is a major allergen of A. simplex. A full-length cDNA encoding the 21 k allergen was cloned by a combination of 3'RACE and screening of an expression library with DIG labeled DNA probes. The precursor of the 21 k allergen was judged to be composed of a signal peptide (23 residues) and a mature protein (171 residues). As compared to the N-terminal amino acid sequence (up to the 17th residue) of Ani s 1 previously identified as the major allergen, the 21 k allergen has only one replacement, suggesting that the 21 k allergen belongs to the same protein family of Ani s 1. Although the 21 k allergen was found to have 30-40% sequence identity with Kunitz-type trypsin inhibitor domain containing hypothetical proteins of Caenorhabditis elegans, it lacked inhibitory activity against trypsin. The 21 k allergen was successfully expressed in Escherichia coli as a GST-fusion protein showing reactivity with IgE in patient sera. PMID- 15287589 TI - Genetic and metabolic analysis of folate salvage in the human malaria parasite Plasmodium falciparum. AB - Antifolate drugs that target the biosynthesis and processing of essential folate cofactors are widely used for treatment of chloroquine-resistant falciparum malaria. Salvage of pre-formed folate can strongly compromise the efficacy of these drugs in vitro and the availability of folate from the human host in natural infections also influences therapeutic outcomes. To investigate how different parasite lines respond to the presence of exogenous folate, we measured the effect of the latter on the susceptibility of parasites to sulfa-drug blockage of folate biosynthesis, utilising the parents and 22 progeny of the HB3 Dd2 genetic cross of Plasmodium falciparum, together with selected unrelated lines. Complete linkage of the folate utilisation phenotype was observed to a DNA sequence of 48.6 kb lying between nucleotide positions 738,489 and 787,058 of chromosome 4 and encompassing the dihydrofolate reductase-thymidylate synthase (dhfr-ts) gene locus. Examination of the putative ORFs on this fragment upstream (3) and downstream (4) of dhfr-ts revealed no plausible candidate genes for folate processing. Similarly, a marked heterogeneity in the 5'-UTR regions of Dd2 and HB3, manifest as a directly repeated 256 bp sequence in the former, also did not correlate with the folate utilisation phenotype nor apparently influence levels of dhfr-ts transcripts or protein products. By contrast, the nature of the coding sequence of the dhfr domain appeared to play a direct role, with the single mutant (S108N) HB3-type utilising folic acid much less efficiently than other allelic variants. We also compared the processing of exogenous folic acid, folinic acid and p-aminobenzoic acid (pABA) in metabolic labelling studies of HB3 and Dd2. These support the view that DHFR is likely to have a low-level folate reductase activity as well as its normal function of reducing dihydrofolate to tetrahydrofolate, and that a significant hurdle in the utilisation of exogenous folic acid is the initial reduction of fully oxidised folic acid to dihydrofolate, an activity that the single mutant enzyme found in HB3 is postulated to perform particularly poorly. This would mirror earlier studies indicating that the DHFR activity of HB3 is also compromised relative to other variants. PMID- 15287590 TI - Sequences required for the flagellar targeting of an integral membrane protein. AB - Previous studies have established that the ISO1 glucose transporter of Leishmania enriettii resides primarily in the flagellar membrane, whereas the ISO2 glucose transporter is located in the pellicular plasma membrane surrounding the cell body. This pronounced difference in subcellular targeting is conferred by the NH2 terminal domain of the transporters, since this is the only region of the two permeases that differs in sequence. Analysis of the 130 residue NH2-terminal domain of ISO1 using multiple terminal deletion mutants and various internal deletion mutants established that a sequence located between amino acids 84 and 100 of this domain is required for flagellar trafficking. In addition, chimeras between ISO1 and ISO2 indicated that the region between residues 110 and 118 of ISO1 is also required for flagellar targeting. These results imply that flagellar targeting information for this integral membrane protein does not constitute a simple linear sequence of amino acids but is at least bipartite in structure. PMID- 15287591 TI - Recombinant expression and enzymatic subsite characterization of plasmepsin 4 from the four Plasmodium species infecting man. AB - Plasmepsin 4 from Plasmodium falciparum and orthologs from Plasmodium malariae, Plasmodium ovale and Plasmodium vivax have been expressed in recombinant form, and properties of the active site of each enzyme characterized by kinetic analysis. A panel of chromogenic peptide substrates systematically substituted at the P3, P2, P2' and P3' positions was used to estimate enzyme/ligand interactions in the corresponding enzyme subsites based upon kinetic data. The kinetic parameters kcat, Km and kcat/Km were measured to identify optimal substrates for each enzyme and also sequences that were readily cleaved by the plasmepsins but poorly by host aspartic peptidases. Computer generated models were utilized to compare enzyme structures and interpret kinetic results. The orthologous plasmepsins share highly similar subsite specificities. In the S3 and S2 subsites, the plasmepsin 4 orthologs all preferred hydrophobic amino acid residues, Phe or Ile, but rejected charged residues such as Lys or Asp. In S2' and S3' subsites, these plasmepsins tolerated both hydrophobic and hydrophilic residues. Subsite specificities of the plasmepsin 4 family of orthologs are similar to those of human cathepsins D and E, except in S3' where the plasmepsins accept substrates containing Ser significantly better than either of these human aspartic proteases. Peptidomimetic methyleneamino reduced-peptide inhibitors, which have inhibition constants in the picomolar range, were prepared for each plasmepsin 4 ortholog based upon substrate preferences. A peptidomimetic inhibitor designed for plasmepsin 4 from P. falciparum having Ser in P3' had the lowest Ki of the series of inhibitors prepared, but did not significantly improve the selectivity of the inhibitor for plasmepsin 4 versus human cathepsin D. PMID- 15287592 TI - The role of Steinernema feltiae body-surface lipids in host-parasite immunological interactions. AB - Interactions between entomopathogenic nematodes (Steinernema feltiae) and insect host (Galleria mellonella) immune system were investigated. We focused on the immunosuppressive properties of the parasite cuticle and on its interaction with hemolymph humoral components. Effects of parasite cuticle against host proPO system enzymatic cascade were evaluated a short time after infection. The presence of parasite cuticles decreased both normal and LPS-elicited proPO system activity, suggesting that S. feltiae body surface plays a key role in the early parasitation phase, probably interfering with host proPO activation pathways. The data obtained showed that cuticle lipidic compounds are able to interact with host humoral components, removing them from the hemolymph. The depletion of these molecules, arbitrarily named host-interacting proteins (HIPs), seems to be responsible of the drastic decrease in proPO system activity. Moreover, hemolymph HIPs showed LPS-binding properties and parasite cuticle cross-reacted with anti LPS antibodies. Finally, we also assessed the involvement of parasite body surface on immunoevasion strategies of S. feltiae against host cell-mediated encapsulation processes. We conclude that S. feltiae body surface is responsible for short-term immunosuppression and immunoevasion processes; since it is able to sequester host hemolymph compounds involved in proPO system activation and this process could be responsible for a molecular disguise strategy against cellular encapsulation. PMID- 15287593 TI - Eimeria tenella sporozoites and merozoites differentially express glycosylphosphatidylinositol-anchored variant surface proteins. AB - Little is known about glycosylphosphatidylinositol (GPI)-linked surface proteins in the coccidian parasite Eimeria tenella. Examination of 28,550 EST sequences from the sporozoite and second merozoite developmental stages of the parasite led to the identification of 37 potential GPI-linked variant surface proteins, termed EtSAGs. Analysis of the complete nucleotide sequences of 23 EtSAG genes separated them into two multi-gene families. All the predicted EtSAG proteins (which vary in length from 228 to 271 residues) have an N-terminal hydrophobic signal peptide, a C-terminal hydrophobic GPI signal-anchor peptide and an extracellular domain organised around six cysteine residues, the positions of which are conserved within each family. Using specific antibodies against a small number of recombinant-expressed EtSAGs, the surface localisation and GPI-anchorage of members of both families was confirmed experimentally. Expression of EtSAGs is differentially regulated between the oocyst/sporozoite and second generation merozoite stages, with only one expressed specifically in the sporozoite, a small number expressed in both stages and the majority expressed specifically in the second generation merozoite. Preliminary data support a model in which multiple variant surface antigens are co-expressed on individual parasites, rather than a model of antigenic switching. The biological role(s) of EtSAGs and the effect(s) that expression of a complex repertoire of variant surface antigens by the second generation merozoite has on host adapted immunity are unknown. PMID- 15287594 TI - Substrate specificity of the Leishmania donovani myo-inositol transporter: critical role of inositol C-2, C-3 and C-5 hydroxyl groups. AB - Inositol is an essential precursor for the formation of glycosyl phosphatidylinositol (GPI)-anchors found in the majority of surface molecules in trypanosomatids, in addition to its requirement for phoshatidylinositol signal transduction pathways. In Leishmania donovani, high-affinity inositol transport is catalyzed by the active myo-inositol/H+ transporter MIT, which is driven by a proton gradient across the parasite membrane. We have characterized the substrate specificity and pharmacology of L. donovani MIT in vitro and in promastigote cultures. High substrate specificity of myo-inositol transport was shown in competition studies with 14 different monosaccharides and MIT function was unaffected by the structurally similar pentose sugars or hexoses. L-Fucose and D xylose, both inhibitors of the Na+-dependent inositol transport system in the human host, did not affect MIT transport function in the parasite. Competition studies with eight different inositol isomers revealed that proton bonds between the C-2, C-3 and C-5 hydroxyl groups of myo-inositol and the transporter protein played a critical role for substrate recognition, and the C-3 hydroxyl oxygen appears to act as an electron donor to form an H-bond with a positive charge of the MIT permease. The cytotoxic inositol analogue 3-fluoro-myo-inositol was recognized by MIT with similar affinity as myo-inositol and showed an IC50 value of 42 +/- 8 microM in L. donovani cultures. Finally, substrate affinities of MIT revealed apparent Km values of 84 +/- 8 microM for myo-inositol and 5.4 +/- 0.9 nM for H+, equal pH 8.27 + 0.08, suggesting that the L. donovani myo-inositol/H+ symporter is fully activated at physiological pH in the sandfly midgut or macrophage phagolysosome. We conclude that Leishmania MIT constitutes an attractive target for delivery of cytotoxic inositol analogues and differs significantly from the sodium-coupled myo-inositol transport system of the human host. PMID- 15287595 TI - Plasmodium falciparum genes differentially expressed during merozoite morphogenesis. PMID- 15287596 TI - Purification and identification of a fatty acyl-CoA synthetase from Trypanosoma brucei. PMID- 15287597 TI - Differential gene expression and the effects of Biomphalaria glabrata embryonic (Bge) cell factors during larval Schistosoma mansoni development. PMID- 15287598 TI - Expansion of wild type allele rather than back mutation in pfcrt explains the recent recovery of chloroquine sensitivity of Plasmodium falciparum in Malawi. PMID- 15287599 TI - Regulatory pathways in ion homeostasis involving calcineurin and a calcium transporting ATPase are different between yeast and schistosomes. PMID- 15287600 TI - Management of splenic abscess in a critically ill patient. AB - Because of the increased number of immunocompromised patients within the general population, the incidence of splenic abscesses has increased over the last decade. This cohort of immune-deficient patients with splenic abscesses engenders a distinct evolution in the pathogenesis and microbiology of the disease process. Moreover, the morbidity and mortality rates for splenic abscesses are increased in this unique population. Clinically, these patients do not have a characteristic presentation. Diagnostically, computed tomography of the abdomen is the test of choice. Antibiotics and splenectomy remain the standard of care in most clinical settings. However, percutaneous drainage is reported with solitary and unilocular abscesses and in poor operative candidates. An unusual case of a patient with a splenic abscess awaiting heart transplantation is presented. This patient was successfully treated with percutaneous drainage and antibiotics. The literature regarding the presentation, diagnosis, pathogenesis, and treatment of splenic abscesses is reviewed as well. PMID- 15287601 TI - New tools for laparoscopic division of the pancreas: a comparative animal study. AB - We tested the hypothesis that the pancreas can be safely divided laparoscopically using non-suture devices. Twelve pigs were randomized into 4 groups: 1) laparoscopic distal pancreatectomy (LDP) using an ultrasonic scalpel; 2) LDP using an ultrasonic scalpel with pancreatic stump suture reinforcement; 3) LDP using a 35-mm laparoscopic linear vascular stapler; 4) LDP using a prototype 35 mm radio-frequency laparoscopic linear vascular stapler. There were no serious complications related to distal pancreatectomy. All groups gained weight by postoperative day (POD) 14. Serum amylase, glucose, electrolytes and total bilirubin levels were measured preoperatively and on POD 1, 3, 7, and 14, and peripancreatic peritoneal fluid amylase levels were measured on POD 7 and 14; all remained normal in all groups. Fewer adhesions to the pancreatic stump were found in the ultrasonic scalpel groups as compared with the stapler groups. Ultrasonic dissection may be the superior means oflaparoscopic transection of the pancreas. PMID- 15287602 TI - Is mini-laparoscopic appendectomy feasible for children. AB - Mini-laparoscopic appendectomy (mini-LA) can be performed safely and efficiently. It is the first treatment choice for patients with acute, uncomplicated appendicitis in our hospital. To evaluate the feasibility of mini-LA for simple and ruptured appendicitis in children, we retrospectively compared the outcomes between mini-LA and open appendectomy for appendicitis in children. From October 1998 to August 2000, the medical records of 585 patients with appendicitis were retrospectively reviewed. Children were defined as patients younger than 15 years of age. The percentage of mini-LA, operation time, time to first flatus passage, duration of hospital stay, and demand for intra-muscular pethidine injection were compared between the mini-LA and open appendectomy. The complications among simple and ruptured appendicitis and the cost of mini-LA and open appendectomy were also analyzed. The operation was performed with one infra-umbilical 10-mm incision, and pneumoperitoneum was established at 12 to 15 mm Hg. A 2-mm laparoscope was inserted via the supra-pubic port, and another 2-mm working port was set-up between the other two ports. Statistical testing using the Whitney Mann U test and Fisher exact test was performed as appropriate. Of the 585 patients, there were 100 children. The youngest patient was 4 years of age and only 7 patients were younger than 5 years. Among children, 18% had a perforated appendix. Mini-LA accounted for 83% of appendectomies in the pediatric group, but it increased yearly (from 41.7% in 1998 to 92.5% in 2000). The operation time of mini-LA and open appendectomy were 57.32 minutes and 49.12 minutes. There was significant improvement in mini-LA from 1998 to 1999. Flatus passage, hospital stay, and pethidine use all favored the mini-LA. For pediatric appendicitis involving a ruptured appendix, postoperative ileus and length of hospital stay were significantly shortened in the mini-LA group. The postoperative complication was not significantly different between mini-LA and open appendicitis. Mini-LA can be safely performed in pediatric patients and it provides early postoperative recovery and short hospital stay. Even for a ruptured appendix, the mini-LA can be the treatment of choice in a well-equipped hospital with well-trained surgeons. PMID- 15287603 TI - Preliminary report of multi degrees of freedom forceps for endoscopic surgery. AB - Laparoscopic suturing requires more complex techniques than conventional open surgery, because of the limited degrees of freedom of endoscopic devices. The prototype multi degrees of freedom forceps was developed with a concept of a flexibility that frees us from the restriction on suturing during endoscopic surgery. It was designed for a needle holder. We calculated the movement of the forceps' distal joint at the time of horizontal suturing. The learning curve was also investigated for ten surgeons. The device was clinically applied in several surgeries. We could perform suturing freely using this prototype forceps at any point and in any directions. The learning curve had its peak within five training sessions, and may be mastered during about ten training sessions. All clinical applications were successfully achieved. With the increased degree of freedom for forceps, the operability for endoscopic surgery was improved. PMID- 15287604 TI - Chemical pleurodesis to improve patients' quality of life in the management of malignant pleural effusions: the 15 year experience of the National Cancer Institute of Milan. AB - We analyzed chemical pleurodesis role in recurrent neoplastic pleural effusions management, performed by two different techniques: VATS and minimal lateral thoracotomy. We compared the results obtained using the two different procedures, and we also evaluated the two sclerosing agents used (talc and alcohol). From January 1987 to March 2002, we treated 565 patients with malignant pleural effusion: 355 (63%) by VATS and 210 (37%) through mini-thoracotomy all of them underwent chemical pleurodesis: 442 (78%) by means of talc and 123 (22%) by alcohol. Chemical pleurodesis therapeutic success was globally obtained in 436 patients (77%). Dealing with surgical approaches, VATS reduced operating time (33 versus 44 minutes: P < 0.001), mean drainage time (3 versus 5 days: P < 0.001), complications (2% versus 7%: P = 0.006) and mean postoperative course (5 versus 7 days: P < 0.001). Therapeutic success of VATS-treated patients was 81% versus 65% of those undergoing thoracotomy (P < 0.001). We obtained a significantly lower relapse rate in the patients of all the two groups treated with talc versus alcohol (12% versus 35% in VATS group and 25% versus 59% in thoracotomic group). Our data indicate that chemical pleurodesis represents a good palliative treatment of neoplastic pleural effusion. Talc was superior to alcohol as sclerosant agent regardless of the surgical procedure. Comparing the two techniques, VATS should be preferred to minimal thoracotomy. We can suggest talc pleurodesis by VATS as the choice treatment in case of recurrent pleural effusions. PMID- 15287605 TI - Laparoscopic Roux-en-Y choledochojejunostomy for benign biliary disease. AB - Roux-en-Y choledochojejunostomy (RYCJ) is frequently used for biliary bypass surgery. However, reports on laparoscopic RYCJ are scarce. Between February 1997 and February 2002, laparoscopic RYCJ was performed in 6 patients with a recurrent common bile duct (CBD) stone or a benign biliary stricture. The first procedure involved the laparoscopic preparation of the CBD. The Roux limb prepared with an endo-GIA was brought up near the proximal CBD. An end-to-side choledochojejunostomy was then performed using an intracorporeal suture. Jejunojejunostomy was performed using an endo-GIA. The mean operating time was 358.3 minutes. A postoperative complication occurred in 1 patient who had an episode of melena, which resolved spontaneously. All patients were free of symptoms during the follow-up period (27.5 months). Laparoscopic RYCJ may be a useful option, especially in the treatment of benign biliary disease. PMID- 15287606 TI - Middle colic artery thrombosis as a result of retained intraperitoneal gallstone after laparoscopic cholecystectomy. AB - Spilled gallstones during laparoscopic cholecystectomy are associated with a number of complications such as infection, abscess, inflammation, adhesions, cutaneous sinuses, small bowel obstruction, incarcerated hernia, and generalized septicemia. We report a case of a patient with middle colic vessel erosion and thrombosis secondary to a retained gallstone following laparoscopic cholecystectomy 11 years ago. At operation, the patient was found to have a necrotic transverse colon with a large 2 cm gallstone compressing her middle colic vessels. An extended right hemicolectomy was performed with a primary anastomosis. The importance of stone retrieval during laparoscopic cholecystectomy is emphasized. PMID- 15287607 TI - Thoracoscopic enucleation of a giant submucosal tumor of the esophagus. AB - Since the introduction of thoracoscopy in the surgical field, many thoracic interventions have been considered feasible via thoracoscopic route. The authors reported a case of thoracoscopic enucleation of a giant esophageal submucosal tumor (8.5 cm in diameter) situated along the left side of the midesophagus. Histopathologic evaluation revealed a gastrointestinal stromal tumor (GIST). Postoperative period was uneventful and the patient was discharged on the eighth postoperative day. Given the well-known advantages of minimally invasive surgery, we assume that the removal of esophageal submucosal tumors can first be attempted by thoracoscopic approach, even if the tumor is of a big size. In cases of histopathologically unknown tumors preoperatively, definitive examination of the complete specimen provides the basis for further therapeutic decisions. PMID- 15287608 TI - Laparoscopic hand-assisted surgery for hydatid cysts of the liver. PMID- 15287609 TI - Laparoscopic treatment of ileocecal intussusception caused by primary ileal lymphoma. AB - Intussusception seldom occurs naturally in adults, but is frequently found due to an underlying disease. We present the case of a 46-year-old man with the chief complaint of repeated abdominal cramping pain, especially in the right lower quadrant, and diarrhea of 1 year's duration. Abdominal sonography and computed tomography showed ileocecal intussusception, and colonoscopy found one protruding tumor at the cecum. Biopsy of the tumor revealed malignant lymphoma cells. Due to persistent symptoms, fear of intestine obstruction caused by further intussusception, and the possibility of mesenteric vascular compression, the patient underwent laparoscopic right hemicolectomy before systemic chemotherapy. The symptoms were relieved successfully after surgery. We emphasize that the majority of adults with intussusception may have an underlying malignancy. The most important rule in treatment is avoidance of tumor emboli spread during manipulation. Therefore, a trial of reduction of the intussuscepted intestine should be prohibited. PMID- 15287610 TI - Laparoscopic repair of bilateral morgagni hernia. AB - Morgagni hernias are rare and comprise approximately 3% of the congenital diaphragmatic hernias that are observed in 1 in 4800 live births. A 50-year-old female patient who complained of nonspecific chest pain radiating to the right arm for approximately 1 year presented. Chest x-ray revealed a right-sided paracardiac mass diagnosed Morgagni hernia by computerized tomography of the thorax. Laparoscopic exploration showed that the retrosternal hernia was actually a bilateral one. After reducing the contents of the hernial sacs, the defects were closed as a single defect with hernia stapler starting from each end and an appropriate sized polyprolene mesh was closed over the repair site. The patient did not have any symptoms of recurrence after 3 months. Recent advances in video endoscopic surgery made it possible to perform repairs of these hernias less traumatic and more comfortable to the patients than the conventional transabdominal or transthoracic methods. PMID- 15287611 TI - Traumatic abdominal wall hernia: same-admission laparoscopic repair. AB - Traumatic abdominal wall hernias are uncommon. They are traditionally treated with open surgery. We report a case repaired using laparoscopic technique and prosthetic reinforcement. PMID- 15287612 TI - Re: Appendix retrieval after laparoscopic appendectomy: a safe and inexpensive technique. PMID- 15287613 TI - Re: Thoracoscopic talc pleurodesis for recurrent, symptomatic pleural effusion following cardiac operations. Surg Lap Endosc Percut Tech. PMID- 15287614 TI - Ambiguous choices. PMID- 15287615 TI - EMRs save time, increase productivity. PMID- 15287616 TI - Body bequest programs in Minnesota. PMID- 15287617 TI - University seeks private funding for embryonic stem cell research. PMID- 15287618 TI - Thoughtful Trekkie. Medical practice, faith, philosophy, and a bit of sci-fi inform the thinking of Mayo Clinic hematologist and ethicist C. Christopher Hook, M.D. PMID- 15287619 TI - Doing the right thing. Hospitals ethics committees help clinicians, families, and facilities wrestle with tough questions. PMID- 15287620 TI - Daily decisions. Even as new biotechnologies expand the horizons of bioethics, focus needs to be placed on the ethics of everyday practice. PMID- 15287621 TI - Patient trust. Trust remains fundamental to the ethical practice of medicine even in the age of the patient as consumer. PMID- 15287622 TI - Ethics of organ distribution in lung transplantation. PMID- 15287624 TI - We don't play politics with science. PMID- 15287623 TI - Why address medical futility now? New guidelines aim to resolve the inevitable differences of opinion that occur when health care providers or family members deem that further medical care is futile. PMID- 15287625 TI - A perspective on homelessness, ethics, and medical care. AB - People who are homeless are known to have high rates of morbidity and mortality. Yet their attitudes, values, and desires regarding end-of-life care have not been studied. This article reports on the findings from interviews with groups of homeless individuals in the Twin Cities performed by University of Minnesota researchers. It explores the implications of homelessness on two commonly discussed ethical principles-autonomy and justice. PMID- 15287626 TI - [Evolution of anastomotic techniques in oesophageal surgery: experience at the Milan University Department of Surgery]. AB - Despite the fact that the incidence and severity of postoperative complications after oesophagectomy have substantially decreased over the past two decades, anastomotic leakage is still a potentially catastrophic event. In this article, the experience of a single surgical unit is analysed. Over the period from 1992 to 2003, 435 oesophagectomies with oesophagogastroplasty were performed at the Milan University Department of Surgery. The overall mortality rate was 1.6%. The incidence of anastomotic leakage was 8.5% (6.5% for intrathoracic anastomoses and 14% for cervical anastomoses), and the mortality rate due to leakage was 13.5%. The authors discuss the factors associated with anastomotic leakage by comparing their personal experience with data from the international literature. PMID- 15287627 TI - Current indications for laparoscopic adrenalectomy in the era of minimally invasive surgery. AB - The aim of this study was to clarify the current indications for laparoscopic adrenalectomy, reviewing both our own experience and the literature data. Since January 2000, 22 patients have undergone adrenalectomy in our department: 17 (77.3%) with the laparoscopic approach and 5 (22.7%) with the traditional one. The indications for laparoscopy were: 6 Cushing's adenomas, 4 aldosterone producing adenomas, 4 non-functional adenomas, 2 pituitary-dependent bilateral adrenocortical hyperplasias and 1 metachronous adrenal metastasis. The conversion rate to laparotomy was 11.7%. The indications for the open approach were: tumours greater than 7 cm and previous abdominal surgery. The mean size of laparoscopic specimens was smaller than those removed by the open procedure (3.9 cm versus 6.7 cm). The mean postoperative hospital stay in the laparoscopic group was 4.9 days as compared to 10.2 days in the open group. Morbidity was encountered in 2/17 laparoscopically treated patients (11.7%) and in 2/5 patients in the open group. In our early experience, laparoscopic adrenalectomy has been the procedure of choice for removing unilateral or bilateral tumours measuring less than 7 cm in diameter. Nevertheless, apart from diameter cut-off, on the basis of evidence from the literature, an invasive carcinoma is currently considered the only absolute contraindication to laparoscopy. PMID- 15287628 TI - [Robotic fundoplication for gastro-oesophageal reflux disease]. AB - Presented as a possible "second" revolution in general surgery after the introduction of laparoscopy during the last few years, the robotic approach to mini-invasive surgery has not yet witnessed wide, large-scale diffusion among general surgeons and is still considered an "experimental approach". In general surgery, the laparoscopic treatment of gastrooesophageal reflux is the second most frequently performed robot-assisted procedure after cholecystectomy. A review of the literature and an analysis of the costs may allow a preliminary evaluation of the pros and cons of robotic fundoplication, which may then be applicable to other general surgery procedures. Eleven articles report 91 cases of robotic fundoplication (75 Nissen, 9 Thal, 7 Toupet). To date, there is no evidence of benefit in terms of duration of surgery, rate of complications and hospital stay. Moreover, robotic fundoplication is more expensive than the traditional laparoscopic approach (the additional cost per procedure due to robotics is 1,882.97 euros). Only further technological upgrades and advances will make the use of robotics competitive in general surgery. The development of multi-functional instruments and of tactile feedback at the console, enlargement of the three-dimensional laparoscopic view and specific "team" training will enable the use of robotic surgery to be extended to increasingly difficult procedures and to non-specialised environments. PMID- 15287629 TI - [Uncommon abdominal sites of hydatid disease. Our experience with the surgical treatment of 15 cases]. AB - The aim of this retrospective study is to report on a series of 15 patients with abdominal hydatid disease in uncommon sites submitted to surgery in our unit over the period 1974-2003. Eight women and 7 men (mean age: 48.4 years) were included in the study. The hydatid cysts were located in the peritoneum in 8 patients, in the spleen in 5, in the kidney in 1 and in the retroperitoneum in 1. In 4 cases the peritoneal cysts were solitary, while 4 patients in this subgroup presented multiple cysts and 2 had concomitant liver hydatidosis. The splenic cysts were solitary in 2 cases, associated with a hepatic cyst in 2 and with a lung cyst in 1. The renal and retroperitoneal cysts were both solitary. The diagnosis was made at operation in 3 cases, while in 12 patients it was made by serological tests, ultrasonography and/or CT. All patients were operated on: we performed a total cystectomy in 7 patients with peritoneal cysts and in the patient with a retroperitoneal location, splenectomy in the 5 splenic cysts and a partial cystectomy with external drainage of the residual cavity in 1 peritoneal cyst and in the renal location. The postoperative course was regular with no mortality and no major morbidity in 14 patients, while 1 patient submitted to splenectomy developed a subphrenic abscess that required surgical drainage. Two recurrences occurred in patients with peritoneal cysts 71 and 20 months, respectively, after the first operation and these were managed by total cystectomy. The diagnosis of uncommon abdominal sites of hydatid disease is more accurate today because of the new imaging techniques, which are often able to show specific radiological signs of hydatid disease. The treatment of choice is surgical and complete removal of the cyst is the gold standard, but its feasibility is related to the location of the cyst. PMID- 15287630 TI - The role of intraoperative angiography in arterial thromboembolectomy for non traumatic acute upper limb ischaemia. AB - The aim of the study was to investigate the influence of intraoperative arteriography on the outcome of upper limb acute arterial thromboembolectomy. In a retrospective study, 63 thromboembolectomies were analysed in 59 patients with acute ischaemia of the upper limbs (51 embolectomies and 12 thrombectomies). In 19 interventions, intraoperative angiography was performed either routinely or because of difficulty in passing the Fogarty catheter and/or absence of backflow. No angiography was performed in 44 interventions because the surgeons were dubious as to the benefits of the procedure. In 6/19 cases (32%) intraoperative angiography led to an extension of the intervention, with 1 intraoperative transluminal angioplasty, 2 patchplasties, 4 distal thromboembolectomies and 2 thromboendarterectomies. In patients with embolic occlusion, the adoption of routine intraoperative angiography resulted in a significantly lower re-occlusion rate at 6 months in comparison with patients who were not submitted to angiography (P<0.05). Also in patients with thrombotic occlusion, the adoption of angiography resulted in a lower re-occlusion rate at 6 months compared to patients not submitted to angiography (P<0.05). There were no amputations at 1 month. We recommend intraoperative arteriography as a routine procedure due to its positive influence on the outcome of thromboembolectomy for acute upper limb ischaemia. PMID- 15287631 TI - [Liver resection after downstaging with neoadjuvant chemotherapy for "unresectable" colorectal metastases]. AB - Hepatic resection is the only treatment that offers a chance of long-term survival in patients with metastases from colorectal cancer. Nevertheless, a curative resection can be performed in only 10-20 per cent of patients: multiple bilobar metastases or "unresectable" disease are the greatest obstacles to surgical radicality. Techniques such as preoperative portal embolisation, preoperative portal ligation, two-stage hepatectomy, and neoadjuvant chemotherapy, have extended the possibility of liver surgery to patients with advanced metastatic colorectal cancer. The outcomes of two patients treated successfully with neoadjuvant chemotherapy (one case with FOL-F-OX, and one with FOL-F-IRI) followed by liver resection were analyzed. In both patients neoadjuvant chemotherapy enabled a curative liver resection to be performed without significant complications. In some patients, neoadjuvant chemotherapy permits the "downsizing" of metastatic disease to such an extent that a surgical approach proves feasible. This advance can dramatically improve the prognosis of patients with multiple or unresectable liver metastases from colorectal cancer. PMID- 15287632 TI - Multiple pulmonary nodules: a decisional algorithm. AB - The present international system for staging lung cancer classifies multiple pulmonary nodules present in one pulmonary lobe or in distinct ipsilateral or controlateral pulmonary lobes as stage IIIB and IV, respectively. In our opinion this approach does not guarantee the patient the best choice of therapy. Moreover, some studies support the choice of surgery for patients with multiple pulmonary nodules without mediastinal lymph node involvement. Introducing PET may be of considerable help in making an informed choice. In this work we propose a diagnostic/therapeutic decisional algorithm for patients with multiple pulmonary nodules. PMID- 15287633 TI - [Retroperitoneal approach in the treatment of supra- and infrarenal aortic aneurysms]. AB - The retroperitoneal approach for the treatment of thoracoabdominal type IV and infrarenal aortic aneurysms is an accepted alternative to thoraco phrenolaparotomy. The purpose of this retrospective study was to report our experience and results in terms of respiratory and renal complications. From January 1997 to December 2003, 48 patients (36 with thoracoabdominal type IV and 12 with infrarenal aortic aneurysms) were treated by a retroperitoneal extrapleural approach in intercostal space X or XI. We performed 40 aorto-aortic and 8 aorto-basilar reconstructions. The perioperative mortality was 2%. Postoperative respiratory insufficiency was 8%, and postoperative renal insufficiency 12%. Permanent dialysis was necessary in 4% of cases. The survival rates were 98%, 89.4% and 58.7 at 1, 5 and 7 years, respectively. Retroperitoneal extrapleural access with a partial phrenotomy results in a significantly reduced incidence of postoperative respiratory complications. PMID- 15287634 TI - [Thyroid surgery: total and partial resection. Analysis of complications and a review of the literature]. AB - The authors review the recent international literature relating to approximately 36,800 cases of thyroid surgery, analysing the complications associated with total thyroidectomy versus partial resection of the thyroid, with a view to drawing up lines of conduct in terms of indications for surgery of benign disease and suggestions on surgical technique to reduce complications such as recurrent laryngeal nerve injuries (transient and definitive), hypocalcaemia (transient and definitive) and superior laryngeal nerve injuries. PMID- 15287635 TI - [Gastrin-secreting neuroendocrine carcinoma: two cases identified and treated surgically with a radio-guided technique]. AB - The authors present two cases of gastrin-producing carcinoma demonstrated by scintigraphy with In-111 pentetreotide. The possible malignant evolution of such tumours and the good results that can be achieved with surgical therapy suggest the usefulness of a radio-guided intraoperative strategy to locate the lesions and minimise invasive procedures. This approach enabled us to detect a lymph-node metastasis concealed in the adipose tissue in the first case and to exclude any other gastrin-producing tumour localisations in the second. We recommend that the radio-guided strategy should be included among the guidelines for the treatment of gastrinomas and believe that the procedure can also be used in other functioning endocrine tumours. PMID- 15287636 TI - [Identification of gallbladder pedicle anatomy during laparoscopic cholecystectomy]. AB - Laparoscopic cholecystectomy is widely accepted nowadays as the gold standard in the treatment of cholelithiasis. This new technique was initially associated with a significant increase in morbidity, and in particular in iatrogenic biliary injuries and arterial haemorrhages, perhaps due to a lack of knowledge of the "laparoscopic anatomy" of the gallbladder pedicle. In this technique the anatomical structures are viewed on a two-dimensional video monitor, and the dissection is performed with long instruments without manual sensitivity. Therefore, the laparoscopic surgeon has to deal with new anatomical views and must be aware of the possible arterial and biliary variants. In this review we describe our technique of laparoscopic cholecystectomy, with particular reference to manoeuvres useful for identifying the various anatomical structures at the gallbladder hilum. In our opinion, it is mandatory to avoid cutting any duct if its identity has yet to be established. For this reason, we pay great attention to the anatomical dissection of Calot's triangle, in order to accurately identify the cystic duct and the cystic artery and any other vascular or biliary structures. Routine intraoperative cholangiography may be useful for identifying the biliary anatomy. When in doubt, the surgeon should not hesitate to convert the procedure to open surgery. PMID- 15287637 TI - Surgical infections after laparoscopic cholecystectomy: ceftriaxone vs ceftazidime antibiotic prophylaxis. A prospective study. AB - The incidence of surgical infections after laparoscopic cholecystectomy is reported to be <2%, because of the minimal trauma due to this approach. We report the results of a prospective study of antibiotic prophylaxis in laparoscopic cholecystectomy, comparing ceftriaxone vs ceftazidime. From Jan 1 to Dec 31 2002 a consecutive series of 242 cholecystectomies were performed, consisting in 18 open cholecystectomies and 224 laparoscopic cholecystectomies, 7 of which (3.1%) were converted to open cholecystectomies for technical and/or anatomical reasons. One hundred and eleven patients received 1 g i.v. ceftazidime 1 h before surgery, and 105 patients 1 g i.v. ceftriaxone on an alternate basis. Thirty-nine patients (17.4%) with acute cholecystitis received at least one booster dose at the end of the operation; 30 out of 39 were given further therapy for 2-3 days, i.e. 1 g i.v. bid. Twenty-two patients treated elsewhere with ceftriaxone or ceftazidime before surgery were transferred to another prophylactic regimen. The final diagnosis in the laparoscopic cholecystectomy group was 219 bile stones, 3 adenomas, and 2 occult carcinomas. We had 4 complications (1.8% of 217 laparoscopic cholecystectomies), 2 of which were minor (infection of the umbilical access by S. aureus) and 2 major (1 biliary fistula [accessory duct] and 1 acute pancreatitis), both treated conservatively. Positive bile cultures (27 cases) were unrelated to the clinical course. The incidence of infections after laparoscopic cholecystectomy in our prospective series was <2%. Ceftriaxone is confirmed as the gold standard in biliary tract surgery, but ceftazidime was equivalent (no statistical difference between the two antibiotics, P=0.59 NS). Ultra-short prophylaxis is enough in most cases, except in cholecystitis. We found no correlation between positive bile cultures and surgical infections after laparoscopic cholecystectomy. The umbilicus was the preferred site of infection in obese patients after the laparoscopic procedure. Major complications are usually related to technical pitfalls. PMID- 15287638 TI - [Antibiotic prophylaxis in laparotomic and laparoscopic cholecystectomy]. AB - Antibiotic prophylaxis in biliary surgery, when correctly used, has led to the minimisation of postoperative infections. Conventional cholecystectomy, and particularly laparoscopic cholecystectomy give rise to a very complicated issue concerning the use of antimicrobial prophylaxis, especially in relation to low risk patients. The authors describe their experience with the use of short-term prophylaxis in biliary surgery based on a hundred consecutive laparoscopic cholecystectomies. In addition, the literature on this topic strengthens the authors' conviction that antimicrobial prophylaxis may be indicated in all surgical cholecystectomy procedures, also in view of the difficult management of postoperative infection risk factors. PMID- 15287639 TI - Introduction of laparoscopic appendectomy: a retrospective comparison with the open technique. AB - Laparoscopic appendectomy (LA) is gaining favour as an alternative to the traditional laparotomic technique (OA) despite the persistence of controversy regarding indications, morbidity, hospital stay, costs and surgical time. We present a retrospective analysis of our first three years of experience with the procedure. During this period we performed 235 appendectomies (102 laparoscopic and 133 laparotomic). The conversion rate was 9.7%, due to severe peritonitis, high-grade inflammation and an unfavorable position of the appendix; we found a significantly higher percentage of difficulty due to these factors in the laparotomic procedures. Operating time was similar in the two groups. The rate of associated pathology was higher (22.5% vs 6%) after laparoscopy, but conversion to laparotomy was never necessary for treatment. Early morbidity was limited to 2 patients who underwent laparoscopic appendectomy (1 re-operation for a micro abscess and 1 conservatively treated haemorrhage), while wound infections (13.5% vs 1.9%) and incisional hernias (0% vs 2.3%) were more frequent in the open procedures. Hospital stay was slightly less in the laparoscopic group (4.0 vs 4.7 days). In our initial experience, laparoscopic appendectomy has shown significant advantages in terms of intraoperative diagnosis of associated diseases and diminished morbidity. We advocate a laparoscopic approach to appendicular disease, reserving conversion to laparotomy for selected cases after exploration. PMID- 15287640 TI - One-day surgery in a series of 150 breast cancer patients: efficacy and cost benefit analysis. AB - Financing health-care expenditure has become increasingly more difficult. Considering the high frequency of breast cancer, which affects one million women in the world each year, the reductions of medical expenditure for the treatment of this disease is highly desirable within the limits of medical efficiency and safety. One hundred and fifty patients with carcinoma of the breast underwent surgery in our department with one-day hospitalization. Patients were discharged with the drainage tube still in place and later treated in the out-patient setting, for dressing and checking the wound, and removing the stitches and drainage tube. Four cases of seroma were registered, all resolved by aspiration of the fluid in a single visit, 1 case of haematoma and 1 case of infection. Patients who underwent this short-stay treatment were amply satisfied. Our experience demonstrates that this type of treatment is both safe and effective. Moreover, it provides considerable benefits in terms of national health-care costs as well as being psychologically better for the patients. PMID- 15287641 TI - [Prevention of lymphoedema secondary to the treatment of breast cancer: a case report and proposal for a prevention protocol]. AB - Secondary lymphoedema of the upper limbs is a fairly frequent complication of breast cancer treatment. It is related to dissection of the axillary lymph nodes, and manifests itself in the form of clinically important lymphostasis, particularly when the dissection is combined with radiotherapy. Despite the fact that the surgical treatment of mammary cancer has become more conservative and, at the same time, radiotheraphy (when necessary) now proves less aggressive and more efficacious, secondary lymphoedema is still reported with incidence rates ranging from 5 to 25%, with an increase of up to 35% and more, when dissection of the axillary lymph nodes is followed by radiotherapy. The aim of this study was to highlight the essential importance of an early diagnosis of secondary lymphoedema, above all in relation to the prevention of this pathology. We report the case of a patient who, at the same time as the axillary lymphnode dissection, underwent a microsurgical operation consisting in the construction of lymphatic venous shunts in the arm as a preventive measure, because lymphoscintigraphy of the upper limbs, carried out in advance, had revealed a predisposition to the development of lymphedema. PMID- 15287642 TI - Surgical treatment of wounds of the lower leg and foot with loss of substance. AB - In the surgical treatment of wounds of the distal regions of the leg with loss of substance, particular anatomical conditions and the frequent coexistence of previous diseases or of diseases connected to the trauma make it hard to choose the most appropriate surgical strategy and require a multidisciplinary approach. Timing and treatment modalities must be decided by different operators in order to ensure a lower risk of post-surgical complications and disabling outcomes. The need for plastic and reconstructive treatment is growing as a result both of the improvement in reconstructive techniques and of the increased incidence of major injuries. The authors report their own experience with the treatment of 21 patients (14 males and 7 females). Partial-thickness skin grafts were performed in 11 cases (7 lower-third leg injuries and 4 foot wounds). Fasciocutaneous flaps were used in 8 patients (5 lower-third leg injuries and 3 wounds to the distal third of the foot), while microsurgical flaps were used in 2 cases of exposed injuries to the foot. The authors report the results obtained and the healing times in relation to the severity of the injuries and the complications observed. Patient treatment modalities are discussed and the results analysed. PMID- 15287643 TI - [Pancreaticojejunostomy by simple introduction after pancreaticoduodenectomy]. AB - The method which most surgeons still prefer in the treatment of the pancreatic stump after pancreaticoduodenectomy is pancreaticojejunostomy. In this article, we describe our preliminary experience with a fast, effective method, consisting in an end-to-end pancreaticojejunostomy by simple introduction, in 11 cases operated on without morbidity or mortality. From 1998 to 2002, 11 patients with pancreatic head or distal bile duct neoplasms underwent pancreaticoduodenectomy. After removal of the specimen, the residual pancreatic stump was prepared towards the left for about two centimetres, mobilizing the posterior surface from the porto-meseraic axis. A single layer of interrupted suture, consisting only in two posterior stitches, was enough in all cases; each stitch was done taking the stump full-thickness at about one centimetre from the transection margin (so as to introduce a corresponding portion of parenchyma into the jejunal lumen), and from the superior and inferior margin, respectively, of the pancreas. On the intestinal side, the stitches were passed full-thickness from the inner surface to the outside, 6 to 7 millimetres from the transection margin. After introducing the stump completely into the intestinal lumen, three anterior stitches were always done and knotted between the pancreatic capsule and the jejunum. All the anastomoses proved to be perfectly sealed. PMID- 15287644 TI - Insertion of prolonged venous access device: a comparison between surgical cutdown and percutaneous techniques. AB - The use of totally implantable prolonged venous access devices (TIPVAD) in chemotherapy for oncological patients is now consolidated practice, whereas the choice between surgical cutdown and the percutaneous technique is still a controversial matter. The aim of this study was to retrospectively evaluate the validity and safety of the surgical approach by comparison with percutaneous techniques. Over a period of 17 months, 106 patients (mainly oncological cases) underwent surgical cutdown for TIPVAD placement in the cephalic vein. During a mean follow-up of 8 months (range 1-21), we evaluated the surgical and management complications and compared them with reported results obtained with the percutaneous technique. We observed a lower incidence of pneumothorax, 2 cases of malfunction due to kinking, and no catheter fractures, while management complications were similar to the findings in the literature. In expert hands, the surgical approach is a fast, safe technique with a lower rate of intraoperative complications than the percutaneous approach and less discomfort for the patient. Adequate training of medical and paramedical staff is the most important factor in making TIPVAD reliable and safe in the long term. PMID- 15287645 TI - [Personal treatment of a case of gastrointestinal autonomic nerve tumour of the second duodenal segment]. AB - Gastrointestinal autonomic nerve tumours are an uncommon form of gastrointestinal stromal tumours. Since both gastrointestinal stromal tumours and gastrointestinal autonomic nerve tumours are potentially malignant, radical surgical excision is always required. We report a case of a gastrointestinal nerve tumour measuring about 5 cm in diameter and arising from the medial wall of the second portion of the duodenum about 1.5 cm below the papilla of Vater. Because of this rare location a very invasive procedure (duodenocephalo-pancreatectomy) might have been required for tumour resection. We avoided this operation and implemented an alternative solution. Endoscopic ultrasonography was very helpful for this purpose, revealing that the tumour was confined within the duodenal wall and separated from the papilla. We employed a non-conventional surgical technique consisting in a duodenal resection comprising the tumour and a direct TT anastomosis of the duodenal stumps. Two technical devices were of fundamental importance for carrying out this procedure: (i) Ligasure which made dissection between the pancreatic head and duodenal wall a safe manoeuvre with little bleeding; and (ii) Valtrac, which allowed us to perform a large anastomosis without any tension on the duodenal stumps. Intraoperative endoscopy was also important. No anastomotic leakage occurred. At follow-up at 12 months the patient is in good health and CT scan and endoscopic ultrasonography have shown no recurrence of disease. PMID- 15287646 TI - Bouveret's syndrome: a case report. AB - Duodenal obstruction due to a gallstone from a cholecystoduodenal fistula (Bouveret's syndrome) is a rare complication of gallstone disease. Patients present gastric outlet occlusion with vomiting, abdominal distension and a state of dehydration. Plain film of the abdomen, ultrasonography and CT scans reveal pneumobilia and the obstructing gallstone. Endoscopy is essential for diagnosis and therapy, with a view to the possibility to relieving the occlusion. Endoscopy, however, cannot be used to treat the fistula and is often unsuccessful because of the very large size of the stone. Surgical therapy can be effective both for relieving the occlusion and for fistula treatment. The authors report a case of Bouveret's syndrome successfully treated surgically in an otherwise healthy patient. In critically ill patients, the primary goal of therapy is relief of the occlusion. PMID- 15287647 TI - [Adrenal incidentaloma: a case of subclinical Cushing's syndrome]. AB - The authors describe a case of adrenal incidentaloma that was the cause of subclinical Cushing's syndrome and take the opportunity to weigh up some of the clinical, diagnostic and therapeutic aspects. Besides the particular expression of the symptoms which were difficult to interpret before reaching a diagnosis, the authors describe the diagnostic work-up adopted, aimed at precisely identifying the type of tumour and the surgical procedure implemented laparoscopically, the outstanding validity of which is confirmed compared to traditional adrenalectomy techniques. PMID- 15287648 TI - Epithelioid haemangioendothelioma of the parotid salivary gland: a case report. AB - Haemangioendothelioma is a vascular tumour characterised by the proliferation of endothelial cells with an epithelioid appearance. The behaviour of this neoplasm is intermediate between haemangioma and angiosarcoma. It may be localised in a wide range of sites, with a preference for soft and bone tissue. It is only rarely localised in the head and neck and even more rarely in the salivary glands. We describe a case of haemangioendothelioma in a 28-year-old man that originated in the retroneural region of the parotid gland, compressing the gland tissue and posteriorly infiltrating the muscular plane. A total parotidectomy surgical operation including the removal of lymph nodes in the region was performed followed by radiotherapy. An immunohistochemical investigation, carried out using the oxidase-antiperoxidase method, indicated that the neoplastic elements of the marker of the endothelial cells CD34 were positive for vimentine and for muscle-specific actin, showing a moderate proliferative action of the cellular elements with MIB-1 positivity estimated at around 6%. The peculiarity of the case we describe resides in the rarity of the haemangioendothelioma localisation in the parotid gland. PMID- 15287649 TI - [Abdominal cystic lymphangioma: a case report]. AB - Intra-abdominal cystic lymphangioma is a rare dysembryogenetic tumour. Although laparoscopic treatment may be feasible and has been reported in the literature, the extension of the mass to the retroperitoneum can make a minimally invasive surgical approach difficult or impossible. We describe the case of a patient with lymphangioma with retroperitoneal extension who underwent successful surgical therapy. PMID- 15287650 TI - [Iatrogenic hemothorax and unknown spontaneous rupture of the oesophagus, double dramatic disease]. AB - Iatrogenic haemothorax is a dramatic event and generally lethal if not treated appropriately and rapidly. Any thoracic co-morbidity increases the risk of death. Spontaneous rupture of the oesophagus is an equally lethal illness if not treated. We report a case of left haemothorax after a thoracic drain for spontaneous pneumothorax with ipsilateral effusion in a 77-year old male. The patient was operated on 6 hours after admission to hospital. We found a laceration of the left common carotid and an unsuspected rupture of the supradiaphragmatic oesophagus. Repair of the lesions in a single session led to no further complications. The patient was discharged in good condition. We know of only one case in the literature with Boerhaave's syndrome not treated surgically, whereas all the other cases had a negative outcome if surgery was not performed promptly. The non-specific symptoms in our case delayed the correct diagnosis of the spontaneous rupture of the oesophagus. The mortality rate is 31% in the literature even when there is an early diagnosis with well performed surgical reapair. A rapid decision as to the best surgical tactics and sending these patients to referral centres specialising in oesophageal disease are the keys to achieving good results. PMID- 15287651 TI - Supercritical fluid chromatography. PMID- 15287652 TI - Determination of diazepam in cream biscuits by liquid chromatography. AB - An analytical procedure was developed for the detection and quantitation of diazepam in cream biscuits, which were used to commit crime. The method involves the extraction of diazepam with ethanol at room temperature, and the extract is filtered, evaporated to dryness, and redissolved in the mobile phase, methanol acetonitrile-tetrahydrofuran-water (15 + 55 + 4 + 26, v/v). The separation is achieved on a C18 reversed-phase column with the mobile phase and diode array detection (lambda(max)) at 230 nm. Medazepam is used as the internal standard is for quantification. The calibration plot for the determination of diazepam is based on linear regression analysis (y = 0.6687x + 0.0372; r2 = 0.995). The limit of detection for diazepam in the biscuit samples was estimated as 600 ng/mL. The limit of quantitation for diazepam was estimated as 1.75 microg/mL. The diazepam detected per piece of biscuit was found to be in the range of 0.27-0.45 mg. Pure diazepam was added to biscuit samples at 3 levels (100 and 500 microg/g, and 1 mg/g), and the recoveries were found to be 95%. The mean retention time of diazepam was 2.7 min and that of medazepam (IS) was 4 min. The relative standard deviations of the diazepam level in the biscuit samples were estimated to be 0.4% for retention time and 1.02% for peak area in intraday analysis, whereas the corresponding values were and 0.61 and 2.34% in interday analysis. The method is rapid and reliable for qualitative and quantitative analysis of cream biscuits laced with diazepam, and it can be used by law enforcement laboratories for routine analysis. PMID- 15287653 TI - Colorimetric assay of propranolol tablets by derivatization: novel application of diazotized 4-amino-3,5-dinitrobenzoic acid (ADBA). AB - A novel colorimetric assay of propranolol tablets has been developed. The assay is based on chemical derivatization (aromatic ring derivatization technique) using diazotized 4-amino-3,5-dinitrobenzoic acid (ADBA) as the chromogenic derivatizing reagent and resultant formation of azo dyes. Optimization studies established an optimal reaction time of 5 min at 30 degrees C after mixing on a Vortex mixer the drug/reagent mixture for 10 s. A new absorption maximum (lambda(max)) was found at 470 nm, which was selected as the analytical wavelength. The assays were linear over 1-8 microg/mL propranolol, and the reaction occurred by a 1:1 reagent/drug stoichiometric ratio. The developed method has a low limit of detection of 0.76 microg/mL and is reproducible. It has been applied successfully to the assay of propranolol tablets and is of equivalent accuracy (p > 0.05) with the official (British Pharmacopoeia) ultraviolet spectrophotometric method. The new method has the main advantage of using more affordable instrumentation and could be applied to the in-process quality control of propranolol tablets. PMID- 15287654 TI - Relationships between LC retention, octanol-water partition coefficient, and fungistatic properties of 2-(2,4-dihydroxyphenyl)benzothiazoles. AB - The retention behavior of newly synthesized compounds with antimycotic activity from the 2-(2,4-dihydroxyphenyl)benzothiazole group by high-performance liquid chromatography has been investigated. RP-18 stationary phase and methanol-acetate buffer aqueous mobile phases at pH 4 and 7.4 have been used. In the case of the mobile phase at pH 7.4, higher concentrations of water can be applied than at pH 4. The studied compounds showed regular retention behavior, their log k values decreasing linearly with an increasing concentration of methanol in the mobile phase. On the basis of these relationships, the lipophilicity (log kw), specific hydrophobic surface area (S), and isocratic chromatographic hydrophobicity index (psi0) were determined. Similar log kw values and sensitivity to changes in the structure of compounds studied for both mobile phases have been found. Moderate correlations between the chromatographic parameters and the calculated octanol water log P values were found. Finally, the lipophilicity parameters were compared with the fungistatic properties of compounds expressed by log MIC (minimum inhibitory concentration) values to find quantitative structure activity relationship equations. PMID- 15287655 TI - Rapid fluorescence screening assay for tetracyclines in chicken muscle. AB - A simple, rapid fluorescence assay was developed for screening tetracyclines in chicken muscle at the U.S. tolerance level (2 mg/kg). The method requires only a homogenization of the tissue in acetonitrile-ammonium hydroxide, centrifugation, addition of Mg+2, and another centrifugation before fluorescence of the supernatant is measured at 505 nm (excitation at 385 nm). Comparison of the fluorescence of control chicken muscle extracts with extracts from muscle fortified with either 2 mg/kg tetracycline, oxytetracycline, or chlortetracycline showed no overlap. A threshold level set at the average fluorescence for a series of fortified 2 mg/kg samples minus 3sigma minimized false-negative responses to provide a successful screening method. The method was tested with blinded samples as controls or samples fortified with tetracycline, oxytetracycline, or chlortetracycline in order to demonstrate its utility. This approach can provide an alternative to microbial screening assays. PMID- 15287656 TI - Application of derivative spectrophotometry to simultaneous determination of indomethacin and 5-methoxy-2-methyl-3-indoleacetic acid in metindol injections. AB - Derivative spectrophotometry was employed to develop a rapid and accurate method for simultaneous determination of indomethacin and 5-methoxy-2-methyl-3 indoleacetic acid as its possible impurity in Metindol injections. At the selected wavelengths, 233.04 and 284.65 nm, no interference between the components determined was observed. Under the established experimental conditions, recoveries of the particular components were from 96.14 to 98.17%. Linearity was maintained over a broad range of concentrations, from 11.88 x 10( 3) to 35.64 x 10(-3) mg/mL for indomethacin and 0.4 to 1.2 mg/mL for 5-methoxy-2 methyl-3-indoleacetic acid. The limit of detection was found to be 6.0 x 10(-3) mg/mL for indomethacin and 0.04 x 10(-3) mg/mL for 5-methoxy-2-methyl-3 indoleacetic acid. The limits of quantitation were found to be 10.0 x 10(-3) mg/mL and 0.20 x 10(-3) mg/mL, respectively. PMID- 15287657 TI - Development, validation, and standardization of polymerase chain reaction-based detection of E. coli O157. AB - A diagnostic polymerase chain reaction assay was developed for the detection of E. coli O157 as the first part of a multicenter validation and standardization project. The assay is based on amplification of sequences of the rfbE O157 gene and includes an internal amplification control. The selectivity of the assay was evaluated against 155 strains, including 32 E. coli O157, 38 E. coli non-O157, and 85 non-E. coli. It was shown to be highly inclusive (100%) and exclusive (100%). The assay has a 100% detection probability of approximately 2 x 10(3) cells per reaction. PMID- 15287658 TI - A medium for the presumptive detection of Enterobacter sakazakii in infant formula: interlaboratory study. AB - A standard method for the detection of Enterobacteriaceae was modified for the presumptive detection of Enterobacter sakazakii, and the modified method was validated in an interlaboratory trial with 16 laboratories from 8 European countries. The modification included a differential-elective medium for the isolation of E. sakazakii, consisting of nutrient agar (NA) supplemented with 4 methyl-umbelliferyl alpha-D-glucoside (alpha-MUG). A 25 g sample was added to 225 mL buffered peptone water. After incubation at 35 degrees or 37 degrees C for 16 or 20 h, 10 mL nonselective enrichment was transferred into 90 mL selective enrichment. The selective enrichment was streaked on violet-red bile glucose agar (VRBGA) and incubated at 37 degrees C for 24 h. It was streaked in parallel on NA plates supplemented with alpha-MUG at 50 mg/L and incubated at 25 degrees C for 16 h, and afterwards for an additional 24 h at room temperature in the dark. E. sakazakii appeared as vivid yellow colonies under normal light and showed blue/violet fluorescence under UV light on NA + alpha-MUG plates. Validation samples represented powdered infant formula without E. sakazakii (blanks) and with low (1-10 colony-forming units [CFU]/25 g) and medium (1-10 CFU/g) contamination levels. All samples contained Pseudomonas aeruginosa and Lactobacillus spp. as background flora. The specificity for blank samples was 100%. The sensitivity of the low contamination level was similar for VRBGA and NA + alpha-MUG, i.e., 66.7% (66.7% accordance, 53.9% concordance). For the medium level the sensitivities were 96.7% (93.3% accordance, 93.5% concordance) for VRBGA and 98.3% (96.9% accordance, 96.9% concordance) for NA + alpha-MUG. PMID- 15287659 TI - Biosensor analysis of beta-lactams in milk: comparison with microbiological, immunological, and receptor-based screening methods. AB - Two recently developed surface plasmon resonance biosensor assays for detection of beta-lactams in milk were used to screen raw producer milk samples. Both assays use a beta-lactam receptor protein with carboxypeptidase activity for detection. The results of the biosensor assays were compared with those of various commercial screening tests, i.e., the Delvotest SP, Penzym S, Beta-STAR, SNAP, and Parallux. The results of the 2 biosensor assays showed good agreement with those of the other screening tests. Of 195 analyzed milk samples, the results of only 5 samples differed between the assays. Additionally, 30 milk samples with both negative and positive results in the screening assays were analyzed by liquid chromatography for identification and quantification of any beta-lactam residues. All screening tests showed 0% false-negative results with 15 incurred samples containing between 4.0 and 268 microg/kg penicillin G. The biosensor assays showed 27% positive results (false violatives) with 15 producer milk samples containing penicillin G concentrations between 0 and 3.6 microg/kg, i.e., below maximum residue limit. This figure varied between 27 and 53% for the other screening tests. PMID- 15287660 TI - Determination of the 13C/12C ratio of ethanol derived from fruit juices and maple syrup by isotope ratio mass spectrometry: collaborative study. AB - A collaborative study of the carbon-13 isotope ratio mass spectrometry (13C-IRMS) method based on fermentation ethanol for detecting some sugar additions in fruit juices and maple syrup is reported. This method is complementary to the site specific natural isotope fractionation by nuclear magnetic resonance (SNIF-NMR) method for detecting added beet sugar in the same products (AOAC Official Methods 995.17 and 2000.19), and uses the same initial steps to recover pure ethanol. The fruit juices or maple syrups are completely fermented with yeast, and the alcohol is distilled with a quantitative yield (>96%). The carbon-13 deviation (delta13C) of ethanol is then determined by IRMS. This parameter becomes less negative when exogenous sugar derived from plants exhibiting a C4 metabolism (e.g., corn or cane) is added to a juice obtained from plants exhibiting a C3 metabolism (most common fruits except pineapple) or to maple syrup. Conversely, the delta13C of ethanol becomes more negative when exogenous sugar derived from C3 plants (e.g., beet, wheat, rice) is added to pineapple products. Twelve laboratories analyzed 2 materials (orange juice and pure cane sugar) in blind duplicate and 4 sugar adulterated materials (orange juice, maple syrup, pineapple juice, and apple juice) as Youden pairs. The precision of that method for measuring delta13C was similar to that of other methods applied to wine ethanol or extracted sugars in juices. The within-laboratory (Sr) values ranged from 0.06 to 0.16%o (r = 0.17 to 0.46 percent per thousand), and the among-laboratories (SR) values ranged from 0.17 to 0.26 percent per thousand (R = 0.49 to 0.73 percent per thousand). The Study Directors recommend that the method be adopted as First Action by AOAC INTERNATIONAL. PMID- 15287661 TI - Instrumental factors influencing absorption measurements for fluid food color determination. AB - The differences in color coordinates obtained from the use of different spectral features in the calculation are studied. Seven groups of food (olive oil, vanilla milkshake, brandy, honey, grape juice, vinegar, and orange juice) and an anthocyanin petunidin solution (natural pigment present in several vegetables) with different pH values were selected. Tristimulus values were calculated by considering the different sources of errors (truncation, abridgement, or different bandwidths). Results obtained were corrected by using the methods recommended (see References). These methods of abridgment work well in general, although in some circumstances (20 nm bandwidth or larger), specific spectral weighting functions have to be used to obtain a negligible error. Therefore, it is interesting to know how much difference can be expected from those factors in order to avoid confusion between color differences attributable to instruments and those attributable to actual color changes. PMID- 15287662 TI - Suitability of real-time quantitative polymerase chain reaction and enzyme-linked immunosorbent assay for cry9C detection in Mexican corn tortillas: fate of DNA and protein after alkaline cooking. AB - Alkaline-cooked corn, called nixtamal, is the basis for many traditional corn products such as tortillas, chips, and taco shells that are used widely in Mexico and Central America and in the preparation of snack foods that are consumed globally. To assess the effects of alkaline and thermal treatments on the detectability of DNA and protein for the presence of genetically modified sequences, various nixtamalized products were prepared from blends of conventional white corn containing 0.1, 1.0, and 10% transgenic corn (event CBH 351, StarLink). Real-time quantitative polymerase chain reactions (RTQ-PCR) and immunoassays were used to determine the cry9C gene and protein, respectively, in unprocessed corn kernels, freshly prepared alkaline-cooked and ground corn (masa), masa flour, tortillas prepared from masa by heat treatment, chips prepared from damp masa dough by deep frying, and from tortillas processed at high (200 degrees C) and low temperatures (70 degrees C). In spite of progressive degradation of genomic DNA during processing, RTQ-PCR genetic analysis allowed detection and quantification of the cry9C gene in all products prepared from 10, 1, and 0.1% StarLink corn, except deep-fried chips containing 0.1% StarLink. Enzyme-linked immunosorbent assays readily detected <1 ppm cry9C protein in all blends of unprocessed corn (10, 1, and 0.1% StarLink) as well as in nonfried tortilla and masa products. This technique was not suitable for thermally treated nixtamalized products containing <1% transgenic corn. PMID- 15287663 TI - Estimation of the measurement uncertainty of the high-performance anion-exchange chromatographic determination of carbohydrates in soluble (instant) coffee. AB - The measurement uncertainty of the determination of free and total carbohydrates in soluble (instant) coffee using high-performance anion exchange chromatography with pulsed amperometric detection according to AOAC Method 995.13 and ISO standard 11292 was calculated. This method is important with regard to monitoring several carbohydrate concentrations and is used to assess the authenticity of soluble coffee. We followed the recommendations of the ISO, Eurachem, and Valid Analytical Measurement (VAM) guides: individual uncertainty contributions u(x) were identified, quantified, and expressed as relative standard deviations related to each specific source u(x)/x or RSD(x). Eventually, they were combined to yield the standard uncertainty and the relative standard uncertainty of a given carbohydrate concentration, c, that is respectively u(c) and u(c)/c. As a result of our study, we could demonstrate that the overall repeatability of the carbohydrate determination in duplicate, RSD(r); the repeatability of the integration of the peak area of the carbohydrate standards, RSD(r(area)(ST)); and the uncertainty of the linear calibration model used in our laboratory, RSD (linST), are the most significant contributions to the total uncertainty. The u(c)/c values thus determined differ for each carbohydrate and depend on their concentrations. The least standard uncertainties that can be achieved are about 2.5%. The question of trueness in the total carbohydrate assay (determination of monosaccharides obtained upon hydrolysis of coffee oligo- and polysaccharides) was addressed. For this purpose, we analyzed the data of 2 different collaborative trials in which our laboratory took part. PMID- 15287664 TI - A novel method for the determination of synthetic colors in ice cream samples. AB - A simple method has been developed for the extraction, separation, and determination of synthetic colors in ice cream samples. The process involves the breakdown of emulsion by neutral detergents (Triton X-100 and Tween 20) followed by extraction with petroleum ether for removal of fat. The aqueous colored solution obtained is treated with 5% acetic acid, and the uptake of color is carried out by a wool-dyeing technique. The color is eluted from the wool with 5% ammonia solution, the solution is evaporated to dryness, and the residue is dissolved in 60% ethanol for paper chromatography using trisodium citrate-ammonia water (2 + 5 + 95, w/v/v) as the mobile phase. The colored spots from the paper chromatogram are cut and eluted with 60% ethanol, and the absorbance is measured at the respective lambda maximum corresponding to the Rf value of the appropriate standard. The recoveries of 6 colors, including sunset yellow FCF (SSYFCF), tartrazine, carmoisine, ponceau 4R, brilliant blue FCF (BBFCF), and fast green FCF from spiked samples with either detergent were found to be >90%. However, recoveries of erythrosine were 21 and 65% with Triton X-100 and Tween 20, respectively. Indigo carmine could not be recovered at all because of its fugitive property in 5% ammonia solution, which is used to strip the color from the wool. The sensitivity of the method with the use of Tween 20 is 1 ppm (1 microg/g) for the colors in spiked ice cream samples. With this method, we analyzed samples of 20 branded colored ice cream. The results showed the presence of tartrazine (8.4-43.3 ppm), SSYFCF (23.5-117.6 ppm), carmoisine (traces-53.2 ppm), erythrosine (3.5 ppm), and BBFCF (4.1 ppm) in the ice cream samples. Apart from 2 samples of tuttifruity, all of the ice cream samples showed the presence of permitted synthetic colors below the permissible level of 100 ppm established by the Prevention of Food Adulteration Act of India. PMID- 15287665 TI - Determination of thiabendazole in orange juice and rind by liquid chromatography with fluorescence detection and confirmation by gas chromatography/mass spectrometry after extraction by matrix solid-phase dispersion. AB - A method was developed for the determination of thiabendazole (TBZ) in orange juice and rind based on matrix solid-phase dispersion (MSPD). TBZ was extracted with ethyl acetate and the extract was subsequently cleaned up on a solid-phase extraction column. Fungicide residues were determined by liquid chromatography with fluorescence detection. Recoveries through the method ranged from 87 to 97% with relative standard deviations < or = 11%. The detection and quantitation limits were 0.15 and 0.50 microg/kg, respectively. The confirmation of TBZ residues in positive samples was performed by solid-phase microextraction followed by gas chromatography with mass spectrometric detection using selected ion monitoring. The developed method was applied to determine TBZ levels in commercial orange juices and in juice and rind of fresh oranges. The influence of storage and washing of fruits on TBZ residues was also studied. PMID- 15287666 TI - Differential electrolytic potentiometric determination of some amino acids in dosage forms. AB - The application of direct-current differential electrolytic potentiometry to the nonaqueous titration of amino acids was investigated. The basic character of amino acids in acetic acid was enhanced to permit their direct titration with perchloric acid. A pair of antimony electrodes was used as an indicating system. The shapes of the titration curves obtained were almost symmetrical with sharp peaks. The optimum current density for those titrations was found to be 1-2 microA/cm2. The procedure was applied successfully to the determination of certain amino acids in drug formulations, and the results were favorably compared statistically with those obtained by official methods. PMID- 15287667 TI - Further studies on the spectrophotometric determination of amprolium. AB - The official AOAC spectrophotometric analytical method for amprolium in feeds (961.24) is quantitatively selective for the intact drug in the presence of its primary degradation products. Concentrations evaluated included mixtures of the individual degradates in the presence of amprolium, as well as an equimolar mixture of the 2 degradates. Neither compound responds to the amprolium colorimetric derivatization reaction under any conditions, demonstrating that the official method can be used as an analytical technique for demonstrating the stability of amprolium in medicated feeds. Additionally, liquid chromatography conditions have been established to resolve amprolium from its degradation products. PMID- 15287668 TI - Novel dietary fibers: the importance of carbohydrates in the diet. PMID- 15287669 TI - Dietary fiber: the influence of definition on analysis and regulation. AB - Since 1953 when the term "dietary fiber" was coined, there has been concern about accurately defining this macronutrient component of the human diet. Proper and adequate analytical methodology and food labeling regulations are dependent upon an accurate definition. Health impact studies also depend upon an accurate and meaningful definition along with relevant methodology to provide data of adequate quality for epidemiological and clinical studies. The scientific communities associated with dietary fiber within AOAC INTERNATIONAL have been the leaders in bringing consensus to the dietary fiber definition and method validation for over a quarter of a century. The consensus definition and subsequent methodology have served as the base for regulations worldwide with regard to dietary fiber labeling and health claims. Recently, there has been renewed interest in reviewing the dietary fiber definition and updating it if the review indicates such a need. The American Association of Cereal Chemists completed an effort that provides a continuum for the historical scientific and regulatory efforts while allowing for inclusion of future discoveries into a framework based upon the knowledge gained in the past. Such a definition will provide for transparent and workable regulations with regard to dietary fiber, and will allow the dietary fiber scientific community of AOAC to validate relevant methods. The Food Nutrition Board of the Institute of Medicine of the National Academies has published a set of definitions disconnected from the historical scientific base that do not provide a relevant basis for either adequate methodology, health studies or workable regulations. PMID- 15287670 TI - Measurement of novel dietary fibers. AB - With the recognition that resistant starch (RS) and nondigestible oligosaccharides (NDO) act physiologically as dietary fiber (DF), a need has developed for specific and reliable assay procedures for these components. The ability of AOAC DF methods to accurately measure RS is dependent on the nature of the RS being analyzed. In general, NDO are not measured at all by AOAC DF Methods 985.29 or 991.43, the one exception being the high molecular weight fraction of fructo-oligosaccharides. Values obtained for RS, in general, are not in good agreement with values obtained by in vitro procedures that more closely imitate the in vivo situation in the human digestive tract. Consequently, specific methods for the accurate measurement of RS and NDO have been developed and validated through interlaboratory studies. In this paper, modifications to AOAC fructan Method 999.03 to allow accurate measurement of enzymically produced fructo-oligosaccharides are described. Suggested modifications to AOAC DF methods to ensure complete removal of fructan and RS, and to simplify pH adjustment before amyloglucosidase addition, are also described. PMID- 15287671 TI - Nondigestible oligosaccharides as dietary fiber. AB - An overview is presented of the most important oligosaccharides that are classified as dietary fiber. Their occurrence and structures as well as their various physiological effects are described. The scientific evidence for health effects, associated with these physiological effects, along with the applications in the food industry, are also discussed. PMID- 15287672 TI - Applications and uses of resistant starch. AB - For the past 30 years there has been a steady increase in our knowledge of the sources, uses and physiological effects of resistant starch. However, it has only been in the past decade that the use of ingredients with a high resistant starch content has occurred in foods, initially in Australia but now throughout the world. Foods containing these resistant starch-rich ingredients include not only staple foods, such as bread and breakfast cereals, but also foods designed for those with special physiological or medical needs, such as celiac sensitivity and ulcerative colitis, or for individuals who are seeking to manage energy intake and control weight. Resistant starch has other benefits when compared with traditional sources of dietary fiber in that the preparation and design of foods with additional health benefits have the appearance, taste, and texture characteristics that encourage people to consume these "better for you" foods. As our knowledge of the range of physiological effects that occur through the consumption of resistant starch increases, more applications will be found for their inclusion in an expanding range of foods around the world. PMID- 15287673 TI - Resistant starch: safe intakes and legal status. AB - Resistant starch (RS) can provide added values such as health benefits and fiber content to many processed foods without compromising taste and product quality. RS is defined as the total amount of starch and the products of starch degradation that resists digestion in the small intestine of healthy people, and the class is comprised of many chemically and physically distinct entities. Some forms of RS are present naturally in many foods, and average global consumption is estimated at 3-10 g/day. Among the best-characterized forms of RS are those derived from high-amylose maize (HAM). Animal and human studies confirm the health benefits and safety of HAM-derived RS as a food ingredient. Legally, RS can be sold in most countries if it falls under current food classifications. However, approved fiber measurements vary throughout the world, and different methods are required to measure RS derived from specific sources. A particular RS can only be claimed as fiber on the nutrition label (and as a source for a high fiber claim) if the method appropriate to that product is approved in the country of interest. Similarly, each individual country's regulations must be consulted prior to developing health or other nutrition label claims, such as calorie content for RS-containing products, as these regulations vary widely around the world. PMID- 15287674 TI - Synthesis of resistant starches in plants. AB - The increased incidence in many countries in lifestyle diseases such as colorectal cancer, cardiovascular disease, and diabetes has led to an enhanced interest in disease-prevention measures that can be delivered to target populations through diet. Resistant starch (RS) is emerging as an important dietary component that has the potential to reduce the incidence of bowel health disorders. However, the range of crop species that can serve as effective sources of RS is limited. In this paper the state of knowledge of the starch biosynthesis pathway is reviewed and opportunities to manipulate crop genetics in order to generate additional sources of RS are discussed. The need for a "whole of chain" approach to delivery of RS to the consumer is highlighted because of the impact that different food-processing technologies can have in maintaining, enhancing, or destroying the RS potential of a raw material or food. PMID- 15287675 TI - Physiological aspects of resistant starch and in vivo measurements. AB - Resistant starch (RS) is the sum of starch and products of starch degradation not absorbed in the small intestine of healthy individuals. There are a number of RS with different characteristics which may have a different fate in the colon. As a consequence, all RS should not be considered equivalent as far as physiological properties are concerned; indeed, they may have a different impact on colonic health. This statement may explain part of the apparent contradictions in the literature on RS and cancer or inflammatory disease prevention. RS is fermented in the large intestine into short-chain fatty acids and, among those, butyrate, which is recognized as the main nutrient of the colonocyte. This fermentation pattern seems to be responsible for most of the effects of RS on colonic health. Another important property is linked to its ability to lower colonic pH, which is usually considered as beneficial for mineral biovailability in the colon or cancer prevention. Due to their fate in the digestive tract, RS materials do not seem to have any significant impact on glucose absorption or metabolism. On the contrary, they may have a hypocholesterolemic effect, but available information is contradictory. PMID- 15287676 TI - Diet and metabolic syndrome: where does resistant starch fit in? AB - Metabolic syndrome is a term linking the clinical profiles of some of the world's major health problems today: obesity, heart disease, and diabetes. It is predicated on dietary patterns, and particularly on the delivery of fuel. The effects may be seen first in the development of abdominal obesity and insulin resistance leading to Type 2 diabetes mellitus and coronary heart disease. This review examines the role resistant starch might play in the prevention and management of these conditions. Beginning with a definition of resistant starch, a critical review of the scientific literature is presented. Current knowledge suggests that resistant starch in the diet may assist in the prevention and management of conditions associated with the metabolic syndrome via its potential effects on delaying the delivery of glucose as fuel with subsequent fat utilization and appetite control benefits. There is still a great deal of research to be undertaken in this area, but it is clearly warranted, given the position of starches in the global food supply and the potential impact on population health. PMID- 15287677 TI - Resistant starch: metabolic effects and potential health benefits. AB - Although there is strong evidence that the amount and type of fat in the diet can have dramatic effects on metabolism, the case for carbohydrate subtypes influencing metabolic parameters is emerging. By definition, resistant starch (RS) is any starch that is not digested in the small intestine but passes to the large bowel. Here, RS is a good substrate for fermentation which gives rise to an increase in short-chain fatty acid production. The differing rates of absorption between RS and digestible starch are thought to denote their differential metabolic responses. RS intake is associated with several changes in metabolism which may confer some health benefits. RS intake seems to decrease postprandial glycemic and insulinemic responses, lower plasma cholesterol and triglyceride concentrations, improve whole body insulin sensitivity, increase satiety, and reduce fat storage. These properties make RS an attractive dietary target for the prevention of diseases associated with dyslipidemia and insulin resistance as well as the development of weight loss diets and dietary therapies for the treatment of Type 2 diabetes and coronary heart disease. This review analyzes the body of literature examining the metabolic effects of RS consumption and discusses possible mechanisms whereby increased short-chain fatty acid production in the bowel could account for some of these effects. The effects of RS in the large bowel per se are the topic of other reviews and are not addressed in this paper. PMID- 15287678 TI - Resistant starches and health. AB - It was initially hypothesized that resistant starches, i.e., starch that enters the colon, would have protective effects on chronic colonic diseases, including reduction of colon cancer risk and in the treatment of ulcerative colitis. Recent studies have confirmed the ability of resistant starch to increase fecal bulk, increase the molar ratio of butyrate in relation to other short-chain fatty acids, and dilute fecal bile acids. However the ability of resistant starch to reduce luminal concentrations of compounds that are damaging to the colonic mucosa, including fecal ammonia, phenols, and N-nitroso compounds, still requires clear demonstration. As such, the effectiveness of resistant starch in preventing or treating colonic diseases remains to be assessed. Nevertheless, there is a fraction of what has been termed resistant (RS1) starch, which enters the colon and acts as slowly digested or lente carbohydrate in the small intestine. Foods in this class are low glycemic index and have been shown to reduce the risk of chronic disease. They have been associated with systemic physiological effects such as reduced postprandial insulin levels and higher HDL cholesterol levels. Consumption of low glycemic index foods has been shown to be related to reductions in risk of coronary heart disease and Type 2 diabetes. Type 2 diabetes has in turn been related to a higher risk of colon cancer. If carbohydrates have a protective role in colon cancer prevention this may lie partly in the systemic effects of low glycemic index foods. The colonic advantages of different carbohydrates, varying in their glycemic index and resistant starch content, therefore, remain to be determined. However, as recent positive research findings continue to mount, there is reason for optimism over the possible health advantages of those resistant starches, which are slowly digested in the small intestine. PMID- 15287679 TI - Resistant starch and colorectal neoplasia. AB - There are several approaches to examining the relationship between resistant starch (RS) and development of colorectal cancer (CRC). These include examination of epidemiological relationships, objective testing of effects of RS given to humans on biological events of relevance to CRC, and studies in animal models where protection and mechanisms of protection can be directly tested. Nine epidemiological studies have examined the relationship between starch and CRC and/or adenomas. Most show a significant protective effect. However, epidemiological tools for measuring consumption of RS are poorly developed and so a benefit for RS can only be inferred. On balance, the magnitude of protection by starch appears to be in the order of 25-50%. Human intervention studies have examined the effect of various types and amounts of RS consumption on colonic biology. To generalize from these studies, RS softens stools and increases stool bulk, decreases pH, increases short-chain fatty acids (SCFAs) including butyrate, reduces products of protein fermentation, and decreases bile salts in fecal water. Such changes seem to be achieved within about 4 weeks of commencing consumption. The greatest effects are seen with the highest doses where increased fecal starch recovery is observed. A modest number of animal studies have been undertaken. Those examining effects of RS on colonic biology and biomarkers for CRC confirm and extend the results in humans. RS modifies the lumenal environment, largely through altered fermentation of polysaccharides and proteins. RS also affects epithelial biology in that it increases apoptotic deletion of genetically damaged cells. More work is needed to define what types and combinations of RS, perhaps with probiotics, exert the greatest effects on colonic environment and epithelial biology, and then to test these in the cancer models for their protective effect. A few studies have examined effect of RS on cancer as an end point in several rodent models, but the results are not clear cut. In conclusion, consumption of RS dramatically affects the colonic lumenal environment and facilitates apoptotic deletion of genetically damaged cells in the colon, several of which are considered to be biomarkers associated with risk for CRC. These effects can be interpreted as reflecting improved colonic health, which might be of benefit in protection against CRC. Direct evidence for protection is still not available. PMID- 15287680 TI - Resistant starch as related to companion animal nutrition. AB - Companion animal diets may contain up to 50% starch, derived from cereal grains. The amount of resistant starch (RS) in an ingredient depends on the origin and form of the ingredient and on the processing conditions to which the ingredient has been exposed. Extrusion has proven to be a means of optimizing utilization of starch by companion animals. Although the RS fraction of starch typically decreases by extrusion, retrogradation can result in increased concentrations of this fraction. Limited research exists regarding the effects of RS in companion animal nutrition and gastrointestinal health. Existing in vitro and in vivo research indicates that certain RS sources are readily fermented in the large bowel, producing short-chain fatty acids, whereas others are less fermentable, resulting in excellent laxation properties. Feeding dogs a diet high in RS may result in an increase in fecal bulk due to an increased excretion of microbial matter in those cases where RS is highly fermentable, or to indigestibility of the RS source in other cases. RS has a role to play as a potential proxy for dietary fiber, especially for those companion animals fed diets high in protein and fat and devoid of traditional dietary fiber. PMID- 15287681 TI - Physiological functions of resistant proteins: proteins and peptides regulating large bowel fermentation of indigestible polysaccharide. AB - Animal studies have shown conclusively that feeding of resistant starch (RS) increases production of large bowel total short-chain fatty acids (SCFAs). However, fermentation products of RS may be affected considerably by other dietary ingredients. In rats fed a 20% high-amylose cornstarch (HAS) with casein as the sole protein source, greater cecal SCFAs production was observed compared with that in rats fed a regular cornstarch diet. However, with this diet, the cecal succinate production was also very high. In contrast, when rice or potato protein with lower digestibility was used in place of casein, cecal succinate production decreased with a concomitant increase in butyrate. These observations suggest that nondigested protein, namely resistant protein, might play a role in correcting an imbalance in the ratio of carbohydrate and nitrogen as fermentative substrates for cecal bacteria and in promoting butyrate production. Epidemiological and biochemical data indicate a possible linkage between the fermentation products of starch (butyrate in particular) and the prevention of colorectal cancer as well as ulcerative colits. Accordingly, a fermentation strategy of RS favoring SCFA production should be established to elucidate the potentially beneficial effects of SCFAs on large bowel physiology. PMID- 15287682 TI - [Collagenin and collagen]. PMID- 15287683 TI - [A new in star in antibiotic heaven--daptomycin]. PMID- 15287684 TI - [Antidepressives for children?]. PMID- 15287685 TI - [New asthma-associated gene identified]. PMID- 15287686 TI - [Curcumin as method of choice for cystic fibrosis?]. PMID- 15287687 TI - [Medicine and neurobiology: the of clinical of picture depression]. PMID- 15287688 TI - [Resumption-inhibition in presynaptic action: the medicinal]. PMID- 15287689 TI - [ Action mechanisms of modern antidepressives]. PMID- 15287690 TI - [Modern antidepressives: pharmacokinetics, interaction potentials and therapeutic drug monitoring]. PMID- 15287691 TI - [The treatment of major depression]. PMID- 15287692 TI - [Modern antidepressive in pharmaceutical care]. PMID- 15287693 TI - [ Pharmacoeconomics of modern antidepressives]. PMID- 15287694 TI - ["Targets, drugs and carriers--novel therapeutic approaches"]. PMID- 15287695 TI - Linear and nonlinear functions on modeling of aqueous solubility of organic compounds by two structure representation methods. AB - Several quantitative models for the prediction of aqueous solubility of organic compounds were developed based on a diverse dataset with 2084 compounds by using multi-linear regression analysis and backpropagation neural networks. The compounds were described by two different structure representation methods: (1) with 18 topological descriptors; and (2) with 32 radial distribution function codes representing the 3D structure of a molecule and eight additional descriptors. The dataset was divided into a training and a test set based on Kohonen's self-organizing neural network. Good prediction results were obtained for backpropagation neural network models: with 18 topological descriptors, for the 936 compounds in the test set, a correlation coefficient of 0.92, and a standard deviation of 0.62 were achieved; with 3D descriptors, for the 866 compounds in the test set, a correlation coefficient of 0.90, and a standard deviation of 0.73 were achieved. The models were also tested by using another dataset, and the relationship of the two datasets was examined by Kohonen's self organizing neural network. PMID- 15287696 TI - The effect of tightly bound water molecules on the structural interpretation of ligand-derived pharmacophore models. AB - The importance of the consideration of water molecules in the structural interpretation of ligand-derived pharmacophore models is explored. We compare and combine results from recently introduced methods for bound-water molecule identification in protein binding sites and ligand-superposition-based pharmacophore derivation, for the interpretation of ligand-derived pharmacophore models. In the analysis of thymidine kinase (HSV-1) and poly (ADP-ribose) polymerase (PARP), the concurrent application of both methods leads to an agreement in the prediction of tightly bound water molecules as key pharmacophoric points in the binding site of these proteins. This agreement has implications for approaching binding site analysis and consensus drug design, as it highlights how pharmacophore-based models of binding sites can include interaction features not only with protein groups but also with bound water molecules. PMID- 15287697 TI - Gaussian mapping of chemical fragments in ligand binding sites. AB - We present a new approach to automatically define a quasi-optimal minimal set of pharmacophoric points mapping the interaction properties of a user-defined ligand binding site. The method is based on a fitting algorithm where a grid of sampled interaction energies of the target protein with small chemical fragments in the binding site is approximated by a linear expansion of Gaussian functions. A heuristic approximation selects from this expansion the smallest possible set of Gaussians required to describe the interaction properties of the binding site within a prespecified accuracy. We have evaluated the performance of the approach by comparing the computed Gaussians with the positions of aromatic sites found in experimental protein-ligand complexes. For a set of 53 complexes, good correspondence is found in general. At a 95% significance level, approximately 65% of the predicted interaction points have an aromatic binding site within 1.5 A. We then studied the utility of these points in docking using the program DOCK. Short docking times, with an average of approximately 0.18 s per conformer, are obtained, while retaining, both for rigid and flexible docking, the ability to sample native-like binding modes for the ligand. An average 4-5-fold speed-up in docking times and a similar success rate is estimated with respect to the standard DOCK protocol. PMID- 15287698 TI - Comparison of a 3D-model of the classical alpha-scorpion toxin V from Leiurus quinquestriatus quinquestriatus with other scorpion toxins. AB - In the present paper, a study of classical and insect alpha-scorpion toxins is described. A homology model of the classical alpha-toxin LqqV from Leiurus quinquestriatus quinquestriatus was developed. The model was compared to stable and energetically favourable conformations of AaHII from Androctonus australis Hector and LqhalphaIT from Leiurus quinquestriatus hebraeus, which are the most active alpha-toxins in mammals and insects. The conformations were retrieved from molecular dynamics simulations of known structures. The model of LqqV shows a C terminal conformation similar to LqhalphaIT. This is mainly caused by electrostatic interactions between Lys10 /Lys60 and Glu59, which are comparable to the cation-pi interactions of Tyr10 and Arg64 in LqhalphaIT. During the simulations the structures of AaHII and LqqV were stabilised through electrostatic interactions between Glu32 and Lys50 and especially the loop adjacent to the alpha-helix is affected, which is in contrast to LqhalphaIT. When the molecular electrostatic potentials of the toxins were studied, a possibly important difference between the classical alpha-toxins and the insect alpha toxin LqhalphaIT was found in the area around Lys30 and Arg56 of AaHII, where a positive potential is missing in LqhalphaIT. A large negative potential caused by Asp3, Glu15 and Asp19 in LqhalphaIT is also unique for this toxin. It is proposed that Arg18, which is important for activity of LqhalphaIT, restricts the negative potential in this area and is not essential for toxins where negatively charged residues in comparable positions are not present. PMID- 15287699 TI - Quantitative structure-activity relationships from optimised ab initio bond lengths: steroid binding affinity and antibacterial activity of nitrofuran derivatives. AB - The present day abundance of cheap computing power enables the use of quantum chemical ab initio data in Quantitative Structure-Activity Relationships (QSARs). Optimised bond lengths are a new such class of descriptors, which we have successfully used previously in representing electronic effects in medicinal and ecological QSARs (enzyme inhibitory activity, hydrolysis rate constants and pKas). Here we use AM1 and HF/3-21G* bond lengths in conjunction with Partial Least Squares (PLS) and a Genetic Algorithm (GA) to predict the Corticosteroid Binding Globulin (CBG) binding activity of the classic steroid data set, and the antibacterial activity of nitrofuran derivatives. The current procedure, which does not require molecular alignment, produces good r2 and q2 values. Moreover, it highlights regions in the common steroid skeleton deemed relevant to the active regions of the steroids and nitrofuran derivatives. PMID- 15287701 TI - [Risk management of coronary heart disease]. PMID- 15287700 TI - Solvent effect on the synthesis of clarithromycin: a molecular dynamics study. AB - Clarithromycin (6-O-methylerythromycin A) is a 14-membered macrolide antibiotic which is active in vitro against clinically important gram-positive and gram negative bacteria. The selectivity of the methylation of the C-6 OH group is studied on erythromycin A derivatives. To understand the effect of the solvent on the methylation process, detailed molecular dynamics (MD) simulations are performed in pure DMSO, pure THF and DMSO:THF (1:1) mixture by using the anions at the C-6, C-11 and C-12 positions of 2',4"-[O-bis(TMS)]erythromycin A 9-[O (dimethylthexylsilyl)oxime] under the assumption that the anions are stable on the sub-nanosecond time scale. The conformations of the anions are not affected by the presence of the solvent mixture. The radial distribution functions are computed for the distribution of different solvent molecules around the 'O-' of the anions. At distances shorter than 5 A, DMSO molecules are found to cluster around the C-11 anion, whereas the anion at the C-12 position is surrounded by the THF molecules. The anion at the C-6 position is not blocked by the solvent molecules. The results are consistent with the experimental finding that the methylation yield at the latter position is increased in the presence of a DMSO:THF (1:1) solvent mixture. Thus, the effect of the solvent in enhancing the yield during the synthesis is not by changing the conformational properties of the anions, but rather by creating a suitable environment for methylation at the C-6 position. PMID- 15287702 TI - [Risk management of coronary heart disease-prevention]. AB - Cardiovascular disease is one of the leading causes of death worldwide and is responsible for 45% of deaths in the western world and 24.5% of deaths in the developing countries. In the 21st century these diseases will continue to dominate the disease spectrum and death statistics in both the industrialised and developing worlds. Since 1975 mortality from cardiovascular disease has decreased by about 24 to 28% in most countries. About 45% of this reduction can be attributed to an improvement in treatment of coronary heart disease and around 55% are attributable to a reduction in risk factors, in particular, stopping smoking and control of hypertension. However, especially in the case of ischaemic heart disease, it is not clear whether the reduction in mortality reflects a reduction in incidence of this disease. Due to the aging population and the reduction in age-related mortality, it is expected that the absolute number of people with heart disease will increase. Furthermore, the increase in prevalence of obesity, metabolic syndrome, type II diabetes as well as the higher prevalence of female smokers compared with thirty years ago could result in an increase in mortality over the next years and decades. It has been shown that prevention strategies, such as education campaigns aimed at the general public, can potentially greatly contribute to a reduction in incidence of cardiovascular disease at every stage. In order for such campaigns to be effective, it is necessary to understand and reduce the risk factors for cardiovascular disease. A large proportion of these risk factors are associated with lifestyle and are therefore modifiable. These modifiable risk factors include smoking, hypertension, poor diet, dyslipidemia, lack of exercise, overweight, adiposity and diabetes mellitus and optimisation of these should be a key aim for all adults. Gender differences also play a role in the incidence and prevention of cardiovascular disease. Incidence of myocardial infarction in women increases significantly after the menopause, and mortality through coronary heart disease is higher amongst women than men. Hormonal status, use of oral contraceptives and pregnancy all influence risk for cardiovascular disease in women. Due to the enormity of the problem that cardiovascular disease presents to society and the great potential for management of risk factors for cardiovascular disease through preventive medicine, a number of health promotion and prevention programmes have been initiated by various national and global organisations. This paper presents an analysis of modifiable risk factors for cardiovascular disease together with a review of targeted prevention programmes aiming at reducing these risks. PMID- 15287703 TI - [Risk management of coronary artery disease--pharmacological therapy]. AB - Treatment of coronary artery disease primarily aims at reducing the severity and frequency of cardiac symptoms and improving prognosis. Both goals can be achieved by the administration of beta-receptor blockers, which are now used as first-line therapy in these patients. Calcium channel blockers or nitrates should be given in the event of contraindications or severe intolerance to beta-receptor blocking therapy. Only long-acting calcium channel blockers should be used in this setting. Another indication for additional treatment with calcium channel blockers and nitrates is given when the efficacy of beta-blocker therapy is not sufficient to relieve symptoms. Nitroglycerin and nitrates are the drugs of choice for the treatment of the acute angina pectoris attack. Calcium channel blockers are used as first-line treatment in patients with vasospastic angina. In patients with syndrome X, nitrates as well as calcium channel blockers or beta receptor blockers can be administered. In the absence of contraindications, every patient with coronary artery disease should be given aspirin. A daily dosage of 75 to 150 mg is sufficient to reduce the rate of future cardiac events. Clopidogrel should be given in every patient with intolerance or contraindications for aspirin. Increased plasma homocystein levels seem to be a risk factor for coronary artery disease. Homocystein levels can be reduced by dietary means as well as supplementation of folic acid or vitamin B complex. There is no evidence from controlled randomised studies that a decrease of homocystein is beneficial for the prognosis of patients with coronary artery disease. PMID- 15287704 TI - [Modification of conventional risk factors in coronary artery disease]. AB - The conventional cardiovascular risk factors such as smoking, hyperlipoproteinemia, arterial hypertension and diabetes are responsible for nearly 75% of myocardial infarction events. Since obesity is associated with a two- or threefold increased risk for arterial hypertension and diabetes, the reduction of body weight presents a basic and causal approach. Indeed 60% of the German population is overweight, and every fifth person has obesity. A low calorie diet and higher quality nutrition as well as increased physical activity is the main therapeutic strategy. The maximum fat supply in a 1200 kcal/d diet should be less than 70 g. Training should be of low intensity, below the anaerobic threshold (50-70% of VO2max), in order to obtain optimal metabolic effect in combination with maximal fat reduction. Should the newly adopted lifestyle not result in a satisfactory loss of weight, medication can be applied in addition. Sibutramin (Reductil) or Orlistat (Xenical) can in individual cases be of help and lead to a further weight loss of up to 10%. It has been demonstrated that such weight loss can evoke the same positive effects of glucose metabolism in patients with impaired glucose tolerance as can metformin. Nevertheless, from a prognostic point of view, in patients with coronary artery disease and manifest diabetes, insulin therapy is required. Although arterial hypertension carries with it four times the risk of stroke and twice that of myocardial infarction, the majority of the population does not receive adequate treatment. Even after an acute cardiac event, in every second patient, an elevated blood pressure of > 140/90 mm Hg at the beginning of the rehabilitation period is found. In approximately 80% of the patients, a guideline-based therapy can be achieved during the follow-up phase. Comparable results apply to LDL cholesterol patients as well. For patients with chronic coronary artery disease, it is highly important that medication and change in lifestyle be continued. Patients need to receive standardized information and ongoing medical care. PMID- 15287705 TI - German-language paediatric websites. AB - BACKGROUND: Among medical subspecialties, paediatrics is relatively well represented on the Internet, as evidenced by the number of topical pages retrieved with different search engine queries. We sought to identify the kind of information that is readily available and to evaluate those resources for the quality of patient education from the consumer's perspective. METHODS: A review was conducted of paediatric material found on German-language websites. In November 2002, we used a popular method of searching information on the Internet to locate German-language websites identified by the search string "kinderarzt". The first 200 websites listed in the most popular search engine 'Google' were recorded in a database and compared with one another. RESULTS: A search of the Internet identified 40.900 potential paediatric sites. The most common features of the reviewed websites included information about a specific paediatric practice (71%); general health-care information (52%); and a description of vaccinations (30%). E-mail consulting was offered by 44% of the sites. Only a few sites conform to the code of the Health On the Net (HON) Foundation, clearly state authorship, or exactly identify the most recent material update. CONCLUSIONS: The Internet is a tool capable of providing an unprecedented amount of information to patients interested in paediatric health topics. Our assessment of the status of German-language paediatric information on the Internet indicates that many colleagues use the Internet only to promote their private practice. Our findings indicate that Internet-based information regarding paediatric health care is extensive at present, but is also poorly organised and hard to find. PMID- 15287706 TI - "Above the neck" warning signs of diabetes may help identify those at risk. PMID- 15287708 TI - Blatant nepotism, But good for you! PMID- 15287710 TI - MTAD. PMID- 15287711 TI - A necessary evolution. PMID- 15287712 TI - Pensions: do they still make sense? PMID- 15287713 TI - What's a dentist to do? PMID- 15287714 TI - The new era: comprehensive disease management. PMID- 15287715 TI - Tackling obesity in managed markets. PMID- 15287716 TI - Dual atypical neuroleptic use: prescribing patterns at a Veterans Affairs outpatient clinic. AB - The use of two atypical neuroleptics (ANs) in a single patient is rising. In the study described herein, 57 patients receiving dual ANs were retrospectively evaluated at a Veterans Affairs outpatient medical center in South Carolina. Only 3% of all patients receiving ANs were prescribed two such drugs, and only 58% of patients taking dual ANs met the age-specific criteria for addition of a second drug. In 90% of cases, both prescriptions were written by the same prescriber. No evidence was noted of lack of communication between two prescribers treating the same patient. PMID- 15287717 TI - Voluntary participation in disease management programs. PMID- 15287718 TI - Preliminary findings on the effectiveness of the "healthy living with chronic conditions" workshop in a managed care plan. AB - The objective of this study was to evaluate the effectiveness of an educational program in teaching enrollees of an MCO to manage their medical conditions and consequently improve their quality of life. Treatment and control groups were evaluated before entering the program and after completing it. Statistically significant improvements for participants were found in five areas: (1) health distress, (2) self-rated general health, (3) illness intrusiveness, (4) self efficacy, and (5) activities limitation. PMID- 15287719 TI - Added value in health care with six sigma. AB - Six sigma is the structured application of the tools and techniques of quality management applied on a project basis that can enable organizations to achieve superior performance and strategic business results. The Greek character sigma has been used as a statistical term that measures how much a process varies from perfection, based on the number of defects per million units. Health care organizations using this model proceed from the lower levels of quality performance to the highest level, in which the process is nearly error free. PMID- 15287720 TI - Use of aspirin and its effect on adenoma risk. PMID- 15287721 TI - Disparity in allosteric interactions of monastrol with Eg5 in the presence of ADP and ATP: a difference FT-IR investigation. AB - Eg5 is a kinesin-like motor protein required for mitotic progression in higher eukaryotes. It is thought to cross-link antiparallel microtubules, and provides a force required for the formation of a bipolar spindle. Monastrol causes the catastrophic collapse of the mitotic spindle through the allosteric inhibition of Eg5. Utilizing a truncated Eg5 protein, we employ difference infrared spectroscopy to probe structural changes that occur in the motor protein with monastrol in the presence of either ADP or ATP. Difference FT-IR spectra of Eg5 monastrol-nucleotide complexes demonstrate that there are triggered conformational changes corresponding to an interconversion of secondary structural elements in the motor upon interaction with nucleotides. Notably, conformational changes elicited in the presence of ADP are different from those in the presence of ATP. In Eg5-monastrol complexes, exchange of ADP is associated with a decrease in random structure and an increase in alpha-helical content. In contrast, formation of the Eg5-monastrol-ATP complex is associated with a decrease in alpha-helical content and a concomitant increase in beta-sheet content. One or more carboxylic acid residues in Eg5 undergo unique changes when ATP, but not ADP, interacts with the motor domain in the presence of monastrol. This first direct dissection of inhibitor-protein interactions, using these methods, demonstrates a clear disparity in the structural consequences of monastrol in the presence of ADP versus ATP. PMID- 15287723 TI - Protein dissection experiments reveal key differences in the equilibrium folding of alpha-lactalbumin and the calcium binding lysozymes. AB - The alpha-lactalbumins and c-type lysozymes have virtually identical structure but exhibit very different folding behavior. All alpha-lactalbumins form a well populated molten globule state, while most of the lysozymes do not. alpha Lactalbumin consists of two subdomains, and the alpha-subdomain is considerably more structured in the molten globule state than the beta-subdomain. Constructs derived from the alpha-subdomain of human alpha-lactalbumin containing the A, B, D, and 3(10) helices are known to form a molten globule state in the absence of the rest of the protein (Demarest, S. et al. (1999) J. Mol. Biol. 294, 213-221). Here we reported comparative studies of constructs derived from the same regions of canine and equine lysozymes. These proteins form two of the most stable molten globule states among all the lysozymes. A construct containing the A, B, D, and 3(10) helices of equine lysozyme is partially helical but is less structured than the corresponding human alpha-lactalbumin peptide. Addition of the C-helix leads to a construct that is still less structured and less stable than the alpha lactalbumin construct. The corresponding construct from canine lysozyme is also less structured and less stable than the alpha-lactalbumin peptide. Thus, molten globule formation in human alpha-lactalbumin can be driven by the isolated alpha subdomain, while more extensive interactions are required to generate a stable molten globule in the two lysozymes. The stability of the canine and equine lysozyme constructs is similar, indicating that the extraordinary stability of the canine lysozyme molten globule is not due to an unusually stable isolated alpha-subdomain. PMID- 15287722 TI - Catalysis and function of the p38 alpha.MK2a signaling complex. AB - The p38 mitogen-activated protein kinase (p38) pathway is required for the production of proinflammatory cytokines (TNFalpha and IL-1) that mediate the chronic inflammatory phases of several autoimmune diseases. Potent p38 inhibitors, such as the slow tight-binding inhibitor BIRB 796, have recently been reported to block the production of TNFalpha and IL-1beta. Here we analyze downstream signaling complexes and molecular mechanisms, to provide new insight into the function of p38 signaling complexes and the development of novel inhibitors of the p38 pathway. Catalysis, signaling functions, and molecular interactions involving p38alpha and one of its downstream signaling partners, mitogen-activated protein kinase-activated protein kinase 2 (MK2), have been explored by steady-state kinetics, surface plasmon resonance, isothermal calorimetry, and stopped-flow fluorescence. Functional 1/1 signaling complexes (Kd = 1-100 nM) composed of activated and nonactivated forms of p38alpha and a splice variant of MK2 (MK2a) were characterized. Catalysis of MK2a phosphorylation and activation by p38alpha was observed to be efficient under conditions where substrate is saturating (kcat(app) = 0.05-0.3 s(-1)) and nonsaturating (kcat(app)/KM(app) = 1-3 x 10(6) M(-1) s(-1)). Specific interactions between the carboxy-terminal residues of MK2a (370-400) and p38alpha precipitate formation of a high-affinity complex (Kd = 20 nM); the p38alpha dependent MK2a phosphorylation reaction was inhibited by the 30-amino acid docking domain peptide of MK2a (IC50 = 60 nM). The results indicate that the 30 amino acid docking domain peptide of MK2a is required for the formation of a tight, functional p38alpha.MK2a complex, and that perturbation of the tight docking interaction between these signaling partners prevents the phosphorylation of MK2a. The thermodynamic and steady-state kinetic characterization of the p38alpha.MK2a signaling complex has led to a clear understanding of complex formation, catalysis, and function on the molecular level. PMID- 15287724 TI - Backbone dynamics of an oncogenic mutant of Cdc42Hs shows increased flexibility at the nucleotide-binding site. AB - Cdc42Hs, a member of the Ras superfamily of GTP-binding signal transduction proteins, binds guanine nucleotides, and acts as a molecular-timing switch in multiple signal transduction pathways. The structure of the wild-type protein has been solved (Feltham et al. (1997) Biochemistry 36, 8755-8766), and the backbone dynamics have been characterized by NMR spectroscopy (Loh et al. (1999) Biochemistry 38, 12547-12557). The F28L mutation of Cdc42Hs is characterized by an increased rate of cycling between the GTP and GDP-bound forms leading to cell transformation (Lin et al. (1997) Curr. Biol. 7, 794-797). Here, we describe the backbone dynamics of Cdc42Hs(F28L)-GDP using 1H-15N NMR measurements of T1, T1rho, and steady-state NOE at two magnetic field strengths. Residue-specific values of the generalized order parameters (Ss2 and Sf2), local correlation time (tau(e)), and exchange rate (R(ex)) were obtained using the Lipari-Szabo formalism. Chemical-shift perturbation analysis suggested that very little structural change was evident outside of the nucleotide-binding site. However, residues comprising the nucleotide-binding site, as well as the nucleotide itself, exhibit increased dynamics over a wide range of time scales in Cdc42Hs(F28L) relative to the wild type. In addition to changes in dynamics measured by relaxation methods, hydrogen-deuterium exchange indicated a substantial disruption of the hydrogen-bonding network within the nucleotide binding site. Thus, local dynamic changes introduced by a single-point mutation can affect important aspects of signaling processes without disrupting the conformation of the whole protein. PMID- 15287725 TI - Identification of sequences in apolipoprotein(a) that maintain its closed conformation: a novel role for apo(a) isoform size in determining the efficiency of covalent Lp(a) formation. AB - We have previously demonstrated that, in the presence of the lysine analogue epsilon-aminocaproic acid, apolipoprotein(a) [apo(a)] undergoes a conformational change from a closed to an open structure that is characterized by a change in tryptophan fluorescence, an increase in the radius of gyration, an alteration of domain stability, and an enhancement in the efficiency of covalent lipoprotein(a) [Lp(a)] formation. In the present study, to identify sequences within apo(a) that maintain its closed conformation, we used epsilon-aminocaproic acid to probe the conformational status of a variety of recombinant apo(a) isoforms using analytical ultracentrifugation, differential scanning calorimetry, intrinsic fluorescence, and in vitro covalent Lp(a) formation assays. We observed that the closed conformation of apo(a) is maintained by intramolecular interaction(s) between sequences within the amino- and carboxyl-terminal halves of the molecule. Using site-directed mutagenesis, we have identified the strong lysine-binding site present within apo(a) kringle IV type 10 as an important site within the C terminal half of the molecule, which is involved in maintaining the closed conformation of apo(a). Apo(a) exhibits marked isoform size heterogeneity because of the presence of varying numbers of copies of the kringle IV type-2 domain located within the amino-terminal half of the molecule. Using recombinant apo(a) species containing either 1, 3, or 8 copies of kringle IV type 2, we observed that, while apo(a) isoform size does not alter the affinity of apo(a) for low density lipoprotein, it affects the conformational status of the protein and therefore influences the efficiency of covalent Lp(a) assembly. The inverse relationship between apo(a) isoform size and the efficiency of covalent Lp(a) formation that we report in vitro may contribute to the inverse relationship between apo(a) isoform size and plasma Lp(a) concentrations that has been observed in vivo. PMID- 15287726 TI - Zinc ions trigger conformational change and oligomerization of hepatitis B virus capsid protein. AB - Assembly of virus particles in infected cells is likely to be a tightly regulated process. Previously, we found that in vitro assembly of hepatitis B virus (HBV) capsid protein is highly dependent on protein and NaCl concentration. Here we show that micromolar concentrations of Zn2+ are sufficient to initiate assembly of capsid protein, whereas other mono- and divalent cations elicited assembly only at millimolar concentrations, similar to those required for NaCl-induced assembly. Altered intrinsic protein fluorescence and highly cooperative binding of at least four Zn2+ ions (KD approximately 7 microM) indicated that binding induced a conformational change in capsid protein. At 37 degrees C, Zn2+ enhanced the initial rate of assembly and produced normal capsids, but it did not alter the extent of assembly at equilibrium. Assembly mediated by high zinc concentrations (> or =300 microM) yielded few capsids but produced a population of oligomers recognized by capsid-specific antibodies, suggesting a kinetically trapped assembly reaction. Comparison of kinetic simulations to in vitro assembly reactions leads us to suggest that kinetic trapping was due to the enhancement of the nucleation rate relative to the elongation rate. Zinc-induced HBV assembly has hallmarks of an allosterically regulated process: ligand binding at one site influences binding at other sites (cooperativity) indicating that binding is associated with conformational change, and binding of ligand alters the biological activity of assembly. We conclude that zinc binding enhances the kinetics of assembly by promoting formation of an intermediate that is readily consumed in the reaction. Free zinc ions may not be the true in vivo activator of assembly, but they provide a model for regulation of assembly. PMID- 15287727 TI - Proteolytic antibody light chains alter beta-amyloid aggregation and prevent cytotoxicity. AB - Beta-amyloid (Abeta), a peptide generated by proteolytic cleavage of the amyloid precursor protein (APP), is a major constituent of the neuritic plaques associated with Alzheimer's disease (AD). Up-regulation of alpha-secretase, which can hydrolyze Abeta between Lys16 and Leu17, has been proposed as a potential therapeutic strategy in the treatment of AD. Previously, we identified two light chain antibody fragments that had proteolytic activity against Abeta, one with alpha-secretase-like activity and one with carboxypeptidase-like activity. Here we show that cleavage of Abeta40 by hk14, the light-chain antibody having carboxypeptidase-like activity, alters aggregation of Abeta and neutralizes any cytotoxic effects of the peptide. Cleavage of Abeta40 with c23.5, the light chain having alpha-secretase-like cleavage, substantially increases the aggregation rate of Abeta; however, it does not show any corresponding increase in cytotoxicity. The increase in aggregation resulting from hydrolysis by c23.5 can be attributed to the decreased solubility of the hydrolyzed products relative to the parent Abeta40, primarily the Abeta17-40 fragment. These results demonstrate that antibody fragment mediated proteolytic degradation of Abeta peptide can be a potential therapeutic route to control Abeta aggregation and toxicity in vivo. Our results also suggest that increasing alpha-secretase activity as a therapeutic route must be approached with some caution because this can lead to a substantial increase in aggregation. PMID- 15287728 TI - Crystal structure of a high-affinity variant of rat alpha-parvalbumin. AB - In model peptide systems, Ca2+ affinity is maximized in EF-hand motifs containing four carboxylates positioned on the +x and -x and +z and -z axes; introduction of a fifth carboxylate ligand reduces the affinity. However, in rat beta parvalbumin, replacement of Ser-55 with aspartate heightens divalent ion affinity [Henzl, M. T., et al. (1996) Biochemistry 35, 5856-5869]. The corresponding alpha parvalbumin variant (S55D/E59D) likewise exhibits elevated affinity [Henzl, M. T., et al. (2003) Anal. Biochem. 319, 216-233]. To determine whether these mutations produce a variation on the archetypal EF-hand coordination scheme, we have obtained high-resolution X-ray crystallographic data for alpha S55D/E59D. As anticipated, the aspartyl carboxylate replaces the serine hydroxyl at the +z coordination position. Interestingly, the Asp-59 carboxylate abandons the role it plays as an outer sphere ligand in wild-type rat beta, rotating away from the Ca2+ and, instead, forming a hydrogen bond with the amide of Glu-62. Superficially, the coordination sphere in the CD site of alpha S55D/E59D resembles that in the EF site. However, the orientation of the Asp-59 side chain is predicted to stabilize the D-helix, which may contribute to the heightened divalent ion affinity. DSC data indicate that the alpha S55D/E59D variant retains the capacity to bind 1 equiv of Na+. Consistent with this finding, when binding measurements are conducted in K(+)-containing buffer, divalent ion affinity is markedly higher. In 0.15 M KCl and 0.025 M Hepes-KOH (pH 7.4) at 5 degrees C, the macroscopic Ca2+ binding constants are 1.8 x 10(10) and 2.0 x 10(9) M(-1). The corresponding Mg2+ binding constants are 2.7 x 10(6) and 1.2 x 10(5) M(-1). PMID- 15287729 TI - Selectivity of metal binding and metal-induced stability of Escherichia coli NikR. AB - NikR from Escherichia coli is a nickel-responsive transcription factor that regulates the expression of a nickel ion transporter. Metal analysis reveals that NikR can bind a variety of divalent transition metals, including Ni(II), Cu(II), Zn(II), Co(II), and Cd(II). The selectivity of metal binding to NikR was investigated by using electronic absorption spectroscopy and small-molecule competitors. The relative affinities, Mn(II) < Co(II) < Ni(II) < Cu(II) > or = Zn(II), follow the Irving-Williams series of metal-complex stabilities. Similar metal affinities were measured for the isolated metal-binding domain of NikR. To determine if any of these metal ions confer a differential effect on NikR, the stability of the metal-bound complexes was examined. In both thermal and chemical denaturation experiments, nickel binding stabilizes the protein more than any of the other metals tested. Thermal denaturation experiments indicate that metal dissociation occurs after loss of secondary structure, but there was no evidence for metal binding to unfolded protein following reversible chemical denaturation. These experiments demonstrate that, although several different metals can bind to NikR, nickel exerts a selective allosteric effect. The implications of these experiments on the in vivo role of NikR as a nickel metalloregulator are discussed. PMID- 15287730 TI - Metal-selective DNA-binding response of Escherichia coli NikR. AB - The NikR transcription factor from Escherichia coli is a Ni(II)-dependent repressor that regulates production of the nickel ion transporter encoded by the nik operon. In the previous paper in this issue (Wang, S. C., Dias, A., Bloom, S. L., and Zamble, D. B. (2004) Selectivity of Metal Binding and Metal-Induced Stability of Escherichia coli NikR, Biochemistry 43, 10018-10028) we demonstrated that NikR can bind 1 equiv of Ni(II) or several other divalent transition metals with similar affinities, but that the Ni(II)-loaded protein is less susceptible to thermal or chemical denaturation than other divalent metal complexes. Here, we investigate the metal selectivity of the DNA-binding activity of NikR. Stoichiometric nickel induces binding of nanomolar NikR to the recognition sequence in the nik promoter, but single equivalents of other divalent metals such as Cd(II), Co(II), and Cu(II) also induce a similar DNA-binding affinity. In the presence of excess nickel, DNA-binding experiments indicate that NikR binds to the nik promoter as a tetramer with much higher affinity (20 pM), and it is this response that is selective for nickel. The DNA binding induced by an excess of other divalent metals is weaker, and is enhanced by the addition of stoichiometric nickel. Nickel titrations into a DNA-binding assay reveal a nickel affinity of 30 nM for a second metal-binding site, and in the presence of 30 nM metal only nickel induces detectable DNA binding by Ni(II)-NikR. These experiments support the hypothesis that there are two metal-binding sites and that both contribute to the nickel-selective DNA-binding response. A model for the in vivo activity of NikR is discussed. PMID- 15287731 TI - Structural studies on the Ca2+-binding domain of human nucleobindin (calnuc). AB - Nucleobindin, also known as calnuc, participates in Ca2+ storage in the Golgi, as well as in other biological processes that involve DNA-binding and protein protein interactions. We have determined the three-dimensional solution structure of the Ca(2+)-binding domain of nucleobindin by NMR showing that it consists of two EF-hand motifs. The NMR structure indicates that the phi and psi angles of residues in both motifs are very similar, despite the noncanonical sequence of the C-terminal EF-hand, which contains an arginine residue instead of the typical glycine at the sixth position of the 12-residue loop. The relative orientation of the alpha-helices in the N-terminal EF-hand falls within the common arrangement found in most EF-hand structures. In contrast, the noncanonical EF-hand deviates from the average orientation. The two helix-loop-helix moieties are in the open conformation characteristic of the Ca(2+)-bound state. We find that both motifs bind Ca2+ with apparent dissociation constants of 47 and 40 microM for the noncanonical and the canonical EF-hand, respectively. The Ca(2+)-binding domain of nucleobindin is unstructured in the absence of Ca2+ and folds upon Ca2+ addition. NMR relaxation data and structural studies of the folded domain indicate that it undergoes slow dynamics, suggesting that it is floppier and less compact than a globular domain. PMID- 15287732 TI - Role of disulfide bonds for the structure and folding of proguanylin. AB - The intestinal peptide hormone guanylin circulates mainly as its corresponding prohormone of 94 amino acids and is the first identified endogenous ligand of intestinal guanylyl cyclase C. While the prohormone is biologically inactive, it is processed to the fully functional form with 15 amino acid residues corresponding to the COOH terminus of the precursor protein. In addition to this inactivation of the hormone region, the prosequence makes an essential contribution to the disulfide-coupled folding of the hormone. On the basis of the recently determined solution structure of proguanylin, explanations for these functions of the prosequence were found, indicating that interstrand contacts between the NH2-terminal beta-hairpin of the prosequence and the COOH-terminal hormone region are crucial for formation of the correct disulfide bonds of guanylin. To further investigate the role of individual disulfide bonds upon stabilization of the overall three-dimensional structure of proguanylin and to test the assumption of a direct effect of the prosequence on the structure of the hormone region, we studied the cysteine double mutant proteins proguanylin C48S/C61S and proguanylin-C86S/C94S. Disulfide determination as well as CD and NMR spectroscopy of proguanylin-C48S/C61S reveals an essential function of the Cys48-Cys61 disulfide bond for the stability of the hydrophobic core and thereby for the stability of the overall three-dimensional structure of proguanylin. Furthermore, sequence specific resonance assignment of the second disulfide deletion mutant, proguanylin-C86S/C94S, and comparison of the NMR spectra of this protein with those of the wild-type protein demonstrate that the rigid helical core structure of proguanylin is not affected by the mutation. Additionally, analysis of the interstrand contacts between the termini reveals a direct effect of the prosequence on the conformation of the hormone region. On the basis of these results, we propose a cooperative mechanism that leads to formation of the correct disulfide pattern of guanylin. PMID- 15287733 TI - Alanine-scanning mutagenesis in the signature disulfide loop of the glycine receptor alpha 1 subunit: critical residues for activation and modulation. AB - The glycine receptor enables the generation of inhibitory postsynaptic currents at synapses via neurotransmitter-dependent activation. These receptors belong to the ligand-gated ion channel gene superfamily, in which all members are comprised of five subunits, each of which possesses a signature 13-residue disulfide loop (Cys loop) in the extracellular domain. In this study, we used alanine-scanning mutagenesis of the residues between C138 and C152 of the Cys loop of the glycine receptor alpha1 subunit to identify residues critical for receptor activation and allosteric modulation. Mutation of L142, F145, or P146 to alanine produced decreases in the potency, maximal amplitude, and Hill coefficient for currents elicited by glycine and impaired receptor activation by the agonist taurine. These residues, along with D148, are positionally conserved in the family of LGIC subunits. Mutation at several other positions had little or no effect. The inhaled anesthetics halothane and isoflurane potentiate submaximal agonist responses at wild-type receptors, via an allosteric site. The mutations L142A, F145A, P146A, and D148A abolished positive modulation by these anesthetics, in some cases revealing a small inhibitory effect. A molecular model of the glycine receptor alpha1 subunit suggests that the Cys loop is positioned in a region of the receptor at the interface between the extracellular and transmembrane domains and that the critical functional residues identified here lie along the face of a predominantly hydrophobic surface. The present data implicate the Cys loop as an important functional moiety in the process of glycine receptor activation and allosteric regulation by anesthetics. PMID- 15287734 TI - Tryptophan scanning mutagenesis in the TM3 domain of the Torpedo californica acetylcholine receptor beta subunit reveals an alpha-helical structure. AB - We used tryptophan substitutions to characterize the beta M3 transmembrane domain (betaTM3) of the acetylcholine receptor (AChR). We generated 15 mutants with tryptophan substitutions within the betaTM3 domain, between residues R282W and I296W. The various mutants were injected into Xenopus oocytes, and expression levels were measured by [125I]-alpha-bungarotoxin binding. Expression levels of the M288W, I289W, L290W, and F293W mutants were similar to that of wild type, whereas the other mutants (R282W, Y283W, L284W, F286W, I287W, V291W, A292W, S294W, V295W, and I296W) were expressed at much lower levels than that of wild type. None of these tryptophan mutants produced peak currents larger than that of wild type. Five of the mutants, L284W, F286W, I287W, V295W, and I296W, were expressed at levels <15% of the wild type. I296W had the lowest expression levels and did not display any significant ACh-induced current, suggesting that this position is important for the function and assembly of the AChR. Tryptophan substitution at three positions, L284, V291, and A292, dramatically inhibited AChR assembly and function. A periodicity analysis of the alterations in AChR expression at positions 282-296 of the betaTM3 domain was consistent with an alpha-helical structure. Residues known to be exposed to the membrane lipids, including R282, M285, I289, and F293, were all found in all the upper phases of the oscillatory pattern. Mutants that were expressed at lower levels are clustered on one side of a proposed alpha-helical structure. These results were incorporated into a structural model for the spatial orientation of the TM3 of the Torpedo californica beta subunit. PMID- 15287735 TI - Localization of the voltage-sensor toxin receptor on KvAP. AB - A variety of venomous animals produce small protein toxins that impair the function of voltage-dependent cation channels by affecting the motions of the voltage-sensor domains and altering the energetics of the opening of the channel. In this study, we investigate the location of the receptor for tarantula venom voltage-sensor toxins on the voltage-dependent K+ channel from Aeropyrum pernix (KvAP), an archeabacterial channel that is functionally inhibited by members of this toxin family. We show that it is possible to purify the same set of toxins from venom of the tarantula Grammostola spatulata using either the purified KvAP voltage-sensor domain or the full-length KvAP channel. The equivalence of toxin retention profiles for the two channel proteins implies that the tarantula voltage-sensor toxin receptor resides exclusively on the voltage-sensor domain and that the pore is not required for the toxin-channel interaction. We have identified and characterized the functional properties of a subset of the tarantula toxins that bind to the KvAP voltage-sensor domain. Some of these toxins, VSTX1 and GSMTX4, have been previously isolated, while others, VSTX2 and VSTX3, are new members of the tarantula voltage-sensor toxin family. Some but not all toxins that bind to the voltage-sensor domain affect voltage-dependent gating of KvAP channels in lipid membranes. PMID- 15287736 TI - Recombinant expression and enzymatic characterization of PttCel9A, a KOR homologue from Populus tremula x tremuloides. AB - PttCel9A is a membrane-bound, family 9 glycosyl hydrolase from Populus tremula x tremuloides that is upregulated during secondary cell wall synthesis. The catalytic domain of PttCel9A, Delta(1-105)PttCel9A, was purified, and its activity was compared to TfCel9A and TfCel9B from Thermobifida fusca. Since aromatic amino acids involved in substrate binding at subsites -4, -3, and -2 are missing in PttCel9A, the activity of TfCel9A mutant enzymes W256S, W209A, and W313G was also investigated. Delta(1-105)PttCel9A hydrolyzed a comparatively narrow range of polymeric substrates, and the preferred substrate was (carboxymethyl)cellulose 4M. Moreover, Delta(1-105)PttCel9A did not hydrolyze oligosaccharides shorter than cellopentaose, whereas TfCel9A and TfCel9B hydrolyzed cellotetraose and cellotriose, respectively. These data suggest that the preferred substrates of PttCel9A are long, low-substituted, soluble cellulosic polymers. At 30 degrees C and pH 6.0, the kcat for cellohexaose of Delta(1-105)PttCel9A, TfCel9A, and TfCel9B were 0.023 +/- 0.001, 16.9 +/- 2.0, and 1.3 +/- 0.2, respectively. The catalytic efficiency (kcat/Km) of TfCel9B was 39% of that of TfCel9A, whereas the catalytic efficiency of Delta(1-105)PttCel9A was 0.04% of that of TfCel9A. Removing tryptophan residues at subsites -4, -3, and -2 decreased the efficiency of cellohexaose hydrolysis by TfCel9A. Mutation of W313 to G had the most drastic effect, producing a mutant enzyme with 1% of the catalytic efficiency of TfCel9A. The apparent narrow substrate range and catalytic efficiency of PttCel9A are correlated with a lack of aromatic amino acids in the substrate binding cleft and may be necessary to prevent excessive hydrolysis of cell wall polysaccharides during cell wall formation. PMID- 15287737 TI - Glycosaminoglycan disaccharide alters the dimer dissociation constant of the chemokine MIP-1 beta. AB - Chemokines are immune system proteins that recruit and activate leukocytes to sites of infection. This recruitment is believed to involve the establishment of a chemokine concentration gradient by the binding of chemokines to glycosaminoglycans (GAGs). In previous studies, we elucidated the GAG binding site of the chemokine MIP-1beta and implicated the involvement of the chemokine dimer in GAG binding through residues across the dimer interface. In the present studies, nuclear magnetic resonance spectroscopy was used to investigate the effect of GAG binding on MIP-1beta dimerization. Using several dimerization impaired variants of MIP-1beta (F13Y, F13L, L34W, and L34K), these studies indicate that the addition of disaccharide to the mutants increases their dimerization affinities. For MIP-1beta F13Y, the presence of the disaccharide increases the chemokine dimerization affinity about 9-fold as evidenced by a decrease in the dimer dissociation constant from 610 to 66 microM. Even more dramatically, the dimerization affinity of MIP-1beta L34W also increases upon addition of disaccharide, with the dimer dissociation constant decreasing from 97 to 6.5 microM. After this effect for the mutants of MIP-1beta was shown, similar experiments were conducted with the CC chemokine RANTES, and it was demonstrated that the presence of disaccharide increases its dimerization affinity by almost 7 fold. These findings provide further evidence of the importance of the dimer in chemokine function and provide the first quantitative investigation of the role of GAGs in the manipulation of the MIP-1beta quaternary structure. PMID- 15287739 TI - 1H NMR characterization of the solution active site structure of substrate-bound, cyanide-inhibited heme oxygenase from Neisseria meningitidis: comparison to crystal structures. AB - Heme oxygenase, HO, from the pathogenic bacterium Neisseria meningitidis catabolizes heme for the iron necessary for infection. The enzyme, labeled HemO, exhibits less sequence homology to mammalian HO than another studied HO from Corynebacterium diphtheriae. Solution 1H NMR has been utilized to define the active site molecular and electronic structure of the cyanide-inhibited, substrate-bound complex for comparison with those provided by several crystal structures. Extensive assignments by solely 1H NMR 2D methods reveal a structure that is very strongly conserved with respect to the crystal structure, although 1H/2H exchange indicates dynamically much more stable distal and proximal helices than those for other HOs. Several residues found with alternate orientations in crystal structures of water- and NO-ligated complexes were shown to occupy positions found solely in the NO complex, confirming that there are structural accommodations in response to ligating the substrate complex with a diatomic, H bond acceptor ligand. The observed dipolar shifts allow the determination of the magnetic axes that show that the Fe-CN unit is tilted approximately 10 degrees toward the alpha-meso position, thereby facilitating the alpha-stereoselectivity of the enzyme. Numerous labile protons with larger than usual low-field bias are identified and, in common with the other HO complexes, shown to participate in an extended, distal side H-bond network. This H-bond network orders several water molecules, most, but not all, of which have been detected crystallographically. A series of three C-terminal residues, His207-Arg208-His209, are not detected in crystal structures. However, 1H NMR finds two residues, His207 and likely Arg208 in contact with pyrrole D, which in crystal structures is exposed to solvent. The nature of the NOEs leads us to propose a H-bond between the proximally oriented His207 ring and the carboxylate of Asp27 and a salt-bridge between the terminus of Arg208 and the reoriented 7-propionyl carboxylate. While numerous ordered water molecules are found near both propionates in the crystal structure, we find much larger water NOEs to the 6- than 7-propionate, suggesting that water molecules near the 7-propionate have been expelled from the cavity by the insertion of Arg208 into the distal pocket. The conversion of the 7-propionate link from the N-terminal region (Lys16) to the C-terminal region (Arg208) in the ligated substrate complex both closes the heme cavity more tightly and may facilitate product exit, the rate-limiting step in the enzyme activity. PMID- 15287738 TI - pH-dependent perturbation of Ras-guanine nucleotide interactions and Ras guanine nucleotide exchange. AB - p21Ras (Ras) proteins cycle between active GTP-bound and inactive GDP-bound states to mediate signal transduction pathways that promote cell growth, differentiation, and apoptosis. To better understand how cellular regulatory factors, such as guanine nucleotide exchange factors (GEFs) and nitric oxide (NO), modulate Ras-guanine nucleotide binding interactions, we have conducted NMR and kinetic studies to investigate the pH dependence of Ras-GDP interactions and Ras-guanine nucleotide exchange (GNE). pH-sensitive amide protons were identified and found to be associated with residues in the switch I (Phe28-Asp30) and switch II (Asp57 and Thr58) regions of Ras. Furthermore, most of the residues that interact with Mg2+ exhibit pH-sensitive amide proton chemical shifts which appear to be coupled to pH-dependent Ras Mg2+ binding and guanine nucleotide binding affinity. These results suggest that perturbation of Mg2+ interactions within the Ras-guanine nucleotide complex is critical for pH-dependent dissociation of guanine nucleotide ligands from Ras. Notably, these same regions undergo conformational changes upon association with the Ras GEF, SOS. In addition, although we have recently shown that addition of NO to Ras in the presence of oxygen produces a Ras thiyl radical intermediate that promotes Ras GNE, we have also postulated that another byproduct of this reaction, a H+, may contribute to NO-mediated GNE. However, the results presented herein suggest that the H+ byproduct of the reaction is unlikely to be involved in the NO-mediated Ras GNE. PMID- 15287741 TI - Human pancreatic lipase-related protein 2 is a galactolipase. AB - Human pancreatic lipase-related protein 2 (HPLRP2) was found to be expressed in the pancreas, but its biochemical properties were not investigated in detail. A recombinant HPLRP2 was produced in insect cells and the yeast Pichia pastoris and purified by cation exchange chromatography. Its substrate specificity was investigated using pH-stat and monomolecular film techniques and various lipid substrates (triglycerides, diglycerides, phospholipids, and galactolipids). Lipase activity of HPLRP2 on trioctanoin was inhibited by bile salts and poorly restored by adding colipase. In vivo, HPLRP2 therefore seems unlikely to show any lipase activity on dietary fat. In human pancreatic lipase (HPL), residues R256, D257, Y267, and K268 are involved in the stabilization of the open conformation of the lid domain, which interacts with colipase. These residues are not conserved in HPLRP2. When the corresponding mutations (R256G, D257G, Y267F, and K268E) are introduced into HPL, the effects of colipase are drastically reduced in the presence of bile salts. This may explain why colipase has such weak effects on HPLRP2. HPLRP2 displayed a very low level of activity on phospholipid micelles and monomolecular films. Its activity on monogalactosyldiglyceride monomolecular film, which was much higher, was similar to the activity of guinea pig pancreatic lipase related-protein 2, which shows the highest galactolipase activity ever measured. The physiological role of HPLRP2 suggested by the present results is the digestion of galactolipids, the most abundant lipids occurring in plant cells, and therefore, in the vegetables that are part of the human diet. PMID- 15287740 TI - NMR backbone dynamics of the human type I interferon binding subunit, a representative cytokine receptor. AB - The antiviral and antiproliferative activities of type I interferons (IFNs) are mediated by a common receptor, and its second subunit (IFNAR2) exhibits nanomolar affinity to both IFNalpha and IFNbeta subtypes. We have previously determined the structure of the IFN-binding extracellular domain of IFNAR2 (IFNAR2-EC) using multidimensional NMR [Chill, J. H., Quadt, S. R., Levy, R., Schreiber, G. E., and Anglister, J. (2003) Structure 11, 791-802], showing it to comprise two fibronectin domains linked by a hinge. As the first cytokine receptor structure determined in the unliganded state and in solution, IFNAR2-EC offers an opportunity to characterize the dynamics of the cytokine receptor family and their correlation to biological function. Backbone dynamics of IFNAR2-EC were investigated using 15N relaxation at 11.74 and 18.79 T, and measurements of residual dipolar couplings (RDCs). Dynamics of the binding site distinguish between rigid structural domains, which stabilize the binding site conformation, and a more flexible binding interface which interacts with the ligand. Measurements of diffusional anisotropy and RDCs and model-free analysis all show that the backbone of the hinge interdomain region of IFNAR2-EC is rigid on the picosecond to nanosecond time scale. Signal transduction in cytokines receptors is initiated by ligand-induced juxtaposition of the two receptor subunits, triggering the mutual phosphorylation of kinases associated to their cytoplasmic domains. The rigidity of the hinge ensures correct positioning of the receptor subunits in the ternary signaling complex and modulates the interaction between kinases in the cytoplasm, thereby controlling the rate and efficiency of phosphorylation. PMID- 15287742 TI - Mechanism-based discovery of small molecules that prevent noncompetitive inhibition by cocaine and MK-801 mediated by two different sites on the nicotinic acetylcholine receptor. AB - The nicotinic acetylcholine receptor (nAChR) belongs to a group of five structurally related membrane proteins that play a major role in the communication between approximately 10(12) cells of the mammalian nervous system. The receptor is inhibited by both abused drugs and therapeutic agents. During the past two decades, many attempts have been made to find compounds that prevent cocaine inhibition of this protein. The use of newly developed transient kinetic techniques in investigations of the inhibition of the receptor by cocaine and MK 801 led to an inhibition mechanism not previously proposed. It was observed that the receptor contains two inhibitory sites: one that equilibrates with the tested noncompetitive inhibitors within approximately 50 ms, and a second site that equilibrates with inhibitors within approximately 1 s. The mechanism of inhibition of the rapidly equilibrating inhibitory site has been investigated, and based on that mechanism, the first evidence that small organic molecules exist that prevent inhibition of the rapidly equilibrating inhibitory site was obtained. These compounds did not prevent the inhibition due to the slowly equilibrating inhibitory site. Here, we present the first evidence that a compound (3-acetoxy ecgonine methyl ester) exists that prevents inhibition of the slowly equilibrating inhibitory site and that the mechanism of inhibition of this site differs from that of the rapidly equilibrating site. BC3H1 cells containing a fetal mouse muscle-type nAChR were used, and the receptor was activated by carbamoylcholine. The resulting whole-cell current due to the nondesensitized nAChR was determined. Because the nAChR desensitizes rapidly, the measurements required the use of a transient kinetic technique with a time resolution of 10 ms; the cell-flow technique was used. Inhibitors and compounds that alleviate inhibition were tested by determining their effects on the whole-cell current due to activation of the nAChR by carbamoylcholine. PMID- 15287743 TI - The identification of phosphorylation sites of pp32 and biochemical purification of a cellular pp32-kinase. AB - The versatile phosphoprotein pp32 is involved in important physiological processes, including cell proliferation, apoptosis, mRNA transport, and transcription. We have previously reported that pp32, through histone masking, inhibits histone acetylation and transcriptional activation by histone acetyltransferases. However, how pp32 itself is regulated remained largely unknown. Although pp32 is a phosphoprotein, neither the phosphorylation sites nor the cellular kinase has been identified. In this report, utilizing an in vitro kinase assay and a biochemical purification scheme, we identify casein kinase II as a cellular pp32-kinase. Our deletion and site-specific mutagenesis studies identify serines 158 and 204 as the sites of phosphorylation. Generation and utilization of antibodies with higher affinity for phospho-pp32 demonstrate that pp32 is indeed phosphorylated in vivo at these two sites. Mutagenesis studies on pp32 suggest a role for serines 158 and 204 in its function. The identification of the pp32 kinase and the sites of pp32 phosphorylation as well as the generation of antibodies with higher affinity for phospho-pp32 should now provide key information and tools for future studies on pp32 regulation. PMID- 15287744 TI - Catalytic mechanism of S-ribosylhomocysteinase (LuxS): stereochemical course and kinetic isotope effect of proton transfer reactions. AB - S-ribosylhomocysteinase (LuxS) catalyzes the cleavage of the thioether bond in S ribosylhomocysteine (SRH) to produce homocysteine and 4,5-dihydroxy-2,3 pentanedione (DPD), the precursor of type II bacterial quorum sensing molecule. The proposed mechanism involves a series of proton-transfer reactions, which are catalyzed by an Fe2+ ion and two general acids/bases in the LuxS active site, resulting in the migration of the ribose carbonyl group from its C1 to C3 position. Subsequent beta-elimination at C4 and C5 positions completes the catalytic cycle. In this work, the regiochemistry and stereochemical course of the proton transfer reactions were determined by carrying out the reactions using various specifically deuterium-labeled SRH as substrate and analyzing the reaction products by 1H NMR spectroscopy and mass spectrometry. Our data indicate a suprafacial transfer of the ribose C2 proton to its C1 position and the C3 proton to the C2 position during catalysis, whereas the ribose C4 proton is completely washed into solvent. The primary deuterium kinetic isotope effect suggests that the conversion of 2-keto intermediate to 3-keto intermediate is partially rate limiting. However, mutation of Glu-57, the putative second general acid/base in catalysis, to an aspartic acid renders the final beta-elimination step rate limiting. PMID- 15287746 TI - Bound peptide-dependent thermal stability of major histocompatibility complex class II molecule I-Ek. AB - We used differential scanning calorimetry to study the thermal denaturation of murine major histocompatibility complex class II, I-E(k), accommodating hemoglobin (Hb) peptide mutants possessing a single amino acid substitution of the chemically conserved amino acids buried in the I-Ek pocket (positions 71 and 73) and exposed to the solvent (position 72). All of the I-Ek-Hb(mut) molecules exhibited greater thermal stability at pH 5.5 than at pH 7.4, as for the I-Ek Hb(wt) molecule, which can explain the peptide exchange function of MHC II. The thermal stability was strongly dependent on the bound peptide sequences; the I-Ek Hb(mut) molecules were less stable than the I-Ek-Hb(wt) molecules, in good correlation with the relative affinity of each peptide for I-Ek. This supports the notion that the bound peptide is part of the completely folded MHC II molecule. The thermodynamic parameters for I-Ek-Hb(mut) folding can explain the thermodynamic origin of the stability difference, in correlation with the crystal structural analysis, and the limited contributions of the residues to the overall conformation of the I-Ek-peptide complex. We found a linear relationship between the denaturation temperature and the calorimetric enthalpy change. Thus, although the MHC II-peptide complex could have a diverse thermal stability spectrum, depending on the amino acid sequences of the bound peptides, the conformational perturbations are limited. The variations in the MHC II-peptide complex stability would function in antigen recognition by the T cell receptor by affecting the stability of the MHC II-peptide-T cell receptor ternary complex. PMID- 15287745 TI - Mnb/Dyrk1A phosphorylation regulates the interaction of dynamin 1 with SH3 domain containing proteins. AB - Mnb/Dyrk1A is a proline-directed serine/threonine kinase implicated in Down's syndrome. Mnb/Dyrk1A was shown to phosphorylate dynamin 1 and alter its interactions with several SH3 domain-containing endocytic accessory proteins. To determine the mechanism of regulation, we mapped the Mnb/Dyrk1A phosphorylation sites in dynamin 1. Using a combination of deletion mutants and synthetic peptides, three potential Mnb/Dyrk1A phosphorylation sites (S778, S795, and S857) were first identified. Phosphorylation at S795 and S857 was confirmed in full length dynamin 1, and S857 was subsequently determined to be the major Mnb/Dyrk1A phosphorylation site in vitro. Phosphorylation at S857 was demonstrated to be the basis for altering the binding of dynamin 1 to amphiphysin 1 and Grb 2 by site directed mutants mimicking phosphorylation. Furthermore, S857 of dynamin 1 is phosphorylated by the endogenous kinase in brain extracts and in PC12 cells. In PC12 cells, the state of S857 phosphorylation is dependent on membrane potentials. These results suggest that S857 phosphorylation is a physiological event, which regulates the binding of dynamin 1 to SH3 domain-containing proteins. Since S857 is unique to dynamin 1xa isoforms, Mnb/Dyrk1A regulation of dynamin 1 is expected to be specific to these spliced variants. PMID- 15287747 TI - Interactions of histone H1 with phospholipids and comparison of its binding to giant liposomes and human leukemic T cells. AB - Due to its net positive charge histone H1 readily associates with liposomes containing acidic phospholipids, such as phosphatidylserine (PS). Interestingly, circular dichroism reveals that while histone H1 in aqueous solutions appears as a random coil, its binding to liposomes containing PS is associated with a pronounced increase in alpha-helicity and beta-sheet content, estimated at 7% and 24%, respectively. This interaction further results in vesicle aggregation and lipid mixing. Fluorescence microscopy revealed rapid binding of Texas Red-labeled H1 (TR-H1) to giant liposomes composed of phosphatidylcholine and PS (SOPC/brain PS, 9/1 molar ratio), followed by lateral segregation and subsequent translocation of the membrane-bound H1 into the giant liposome. The above processes in giant liposomes did depend on the presence of the negatively charged PS. Comparison of the behavior of H1 in giant liposomes to that in cultured leukemic T cells demonstrated very similar patterns. More specifically, fluorescence microscopy revealed binding of TR-H1 to the plasma membrane as lateral segregated microdomains, followed by translocation into the cell. H1 also triggered membrane blebbing and fragmentation of the nuclei of these cells, thus suggesting induction of apoptosis. Our findings indicate that histone H1 and acidic phospholipids form supramolecular aggregates in the plasma membrane of T cells, subsequently resulting in major rearrangements of cellular membranes. Our results allow us to conclude that the minimal requirement for the interaction of histone H1 with the leukemia cell plasma membrane is reproduced by giant liposomes composed of unsaturated phosphatidylcholine and phosphatidylserine, the latter being mandatory for the observed changes in the secondary structure of H1 as well as the macroscopic consequences of the H1-PS interactions. PMID- 15287748 TI - Spectroscopic characterization of the soluble guanylate cyclase-like heme domains from Vibrio cholerae and Thermoanaerobacter tengcongensis. AB - Soluble guanylate cyclase (sGC) is a nitric oxide- (NO-) sensing hemoprotein that has been found in eukaryotes from Drosophila to humans. Prokaryotic proteins with significant homology to the heme domain of sGC have recently been identified through genomic analysis. Characterization of two of these proteins is reported here. The first is a 181 amino acid protein cloned from Vibrio cholerae (VCA0720) that is encoded in a histidine kinase-containing operon. The ferrous unligated form of VCA0720 is 5-coordinate, high-spin. The CO complex is low-spin, 6 coordinate, and the NO complex is high-spin and 5-coordinate. These ligand binding properties are very similar to those of sGC. The second protein is the N terminal 188 amino acids of Tar4 (TtTar4H), a predicted methyl-accepting chemotaxis protein (MCP) from the strict anaerobe Thermoanaerobacter tengcongensis. TtTar4H forms a low-spin, 6-coordinate ferrous-oxy complex, the first of this sGC-related family that binds O2. TtTar4H has ligand-binding properties similar to those of the heme-containing O2 sensors such as AxPDEA1. sGC does not bind O2 despite having a porphyrin with a histidyl ligand like the globins. The results reported here, with sequence-related proteins from prokaryotes but in the same family as the sGC heme domain, show that these proteins have evolved to discriminate between ligands such as NO and O2; hence, we term this family H-NOX domains (heme-nitric oxide/oxygen). PMID- 15287749 TI - UDP-galactose 4-epimerase from Kluyveromyces fragilis: analysis of its hysteretic behavior during catalysis. AB - UDP-galactose 4-epimerase serves as a prototype model of class II oxidoreductases that use bound NAD as a cofactor. This enzyme from Kluyveromyces fragilis is a homodimer with a molecular mass of 75 kDa/subunit. Continuous monitoring of the conversion of UDP-galactose (UDP-gal) to UDP-glucose (UDP-glu) by the epimerase in the presence of the coupling enzyme UDP-glucose dehydrogenase and NAD shows a kinetic lag of up to 80 s before a steady state is reached. The disappearance of the lag follows first-order kinetics (k = 3.22 x 10(-2) s(-1)) at 25 degrees C at enzyme and substrate concentrations of 1.0 nM and 1 mM, respectively. The observed lag is not due to factors such as insufficient activity of the coupling enzyme, association or dissociation or incomplete recruitment of NAD by epimerase, product activation, etc., but was a true expression of the activity of the prepared enzyme. Dissociation of the bound ligand(s) by heat followed by analysis with reverse-phase HPLC, TLC, UV-absorption spectrometry, mass spectrometry, and NMR showed that in addition to 1.78 mol of NAD/dimer, the epimerase also contains 0.77 mol of 5'-UMP/dimer. The latter is a strong competitive inhibitor. Preincubation of the epimerase with the substrate UDP-gal or UDP-glu replaces the inhibitor and also abolishes the lag, which reappeared after the enzyme was treated with 5'-UMP. The lag was not observed as long as the cells were in the growing phase and galactose in the growth medium was limiting, suggesting that association with 5'-UMP is a late log-phase phenomenon. The stoichiometry and conserved amino acid sequence around the NAD binding site of multimeric class I (classical dehydrogenases) and class II oxidoreductases, as reported in the literature, have been compared. It shows that each subunit is independently capable of being associated with one molecule of NAD, suggestive of two NAD binding sites of epimerase per dimer. PMID- 15287751 TI - Cold shock domain of the human Y-box protein YB-1. Backbone dynamics and equilibrium between the native state and a partially unfolded state. AB - The three-dimensional structure of the central cold shock domain (CSD) of the human Y-box protein (YB-1 CSD) is virtually identical to those available for the bacterial cold shock proteins (Csp's). We have further characterized YB-1 CSD by studying its dynamics by nuclear magnetic resonance. The observed structural similarity is reflected in the backbone dynamics, which for YB-1 CSD is very similar to that of the Escherichia coli protein CspA. The rotational correlation time of YB-1 CSD shows that it is a monomer. This indicates that the dimerization observed for the YB-1 protein is not caused by its CSD, but involves other parts of this protein. The YB-1 CSD is only marginally stable as are the mesophilic bacterial Csp's. In contrast to the rapid two-state folding of the bacterial Csp's, the formation of the native form of YB-1 CSD is slow and at least a three state process. The NMR experiments revealed the presence of a second state of YB 1 CSD in equilibrium with the native form. The exchange rates from and to the folded state are in the order of 0.2 and 0.5 s(-1), respectively. Relaxation experiments indicated that the second state is a highly flexible, partly structured molecule. PMID- 15287750 TI - Modified bases in RNA reduce secondary structure and enhance hybridization. AB - Secondary structure in RNA targets is a significant barrier to short DNA probes. However, when such targets are the end product of an in vitro amplification scheme, it is possible to carry out transcription in the presence of nucleoside triphosphate analogues that reduce secondary structure of the RNA without impairing subsequent hybridization. Here we show that nucleoside triphosphates of 2-aminoadenine (nA) and 2-thiouracil (sU) are taken up by T7 RNA polymerase and that the resulting RNA possesses reduced secondary structure and improved accessibility to DNA probes. The hybridization properties of short RNA transcripts were studied using a new gel mobility shift assay from which melting temperatures were determined. RNA hairpins that contained nA and sU were able to hybridize to DNA probes under conditions where the unmodified hairpins did not. DNA-RNA hybrids that contained nA and sU in the RNA strand exhibited enhanced specificity, increased stability, and greater equality of base pairing strength than the same hybrids without modifications. Substitution of guanine (G) with inosine (I) further reduced secondary structure, but RNA with this base hybridized nonselectively. The high stability of nA-T and A-sU base pairs in DNA RNA hybrids, combined with the destabilizing effect of the nA-sU couple in RNA targets, accounts for the improved hybridization properties. These results suggest that incorporation of nA and sU during in vitro transcription is a promising strategy for enhancing the performance of oligomeric DNA probes with an RNA target. PMID- 15287752 TI - Baculovirus-mediated overexpression of the phosphorylase b kinase holoenzyme and alpha gamma delta and gamma delta subcomplexes. AB - Recombinant baculoviruses were created and used to coexpress rat phosphorylase kinase (Phk) alpha, gamma, and delta subunits and rabbit beta subunit in insect cells. Coexpression allowed creation of the (alphabetagammadelta)4 hexadecamer, the alphagammadelta heterotrimer, and the gammadelta heterodimeric subcomplexes. Neither the individual alpha, beta, or gamma subunit nor any complex containing the beta subunit other than the hexadecameric holoenzyme was obtained in soluble form. The expressed complexes exhibited pH- and [Ca2+]-dependent specific activities that were similar to those of the Phk holoenzyme purified from rabbit skeletal muscle (SkM Phk). SkM Phk, expressed Phk, and the alphagammadelta subcomplex were activated by exogenous calmodulin and underwent Ca(2+)-dependent autophosphorylation. In some of these features there were subtle differences that could likely be attributed to differences in the covalent modification state of the baculovirus-driven expressed protein. Our results provide an important avenue to probe the detailed characterization of the structure of Phk and the function of the individual domains of the subunits using baculovirus-mediated expression of Phk and Phk subcomplexes. PMID- 15287753 TI - Photoreactions of metarhodopsin III. AB - Meta III is an inactive intermediate thermally formed following light activation of the visual pigment rhodopsin. It is produced from the Meta I/Meta II photoproduct equilibrium of rhodopsin by a thermal isomerization of the protonated Schiff base C=N bond of Meta I, and its chromophore configuration is therefore all-trans 15-syn. In contrast to the dark state of rhodopsin, which catalyzes exclusively the cis to trans isomerization of the C11=C12 bond of its 11-cis 15-anti chromophore, Meta III does not acquire this photoreaction specificity. Instead, it allows for light-dependent syn to anti isomerization of the C15=N bond of the protonated Schiff base, yielding Meta II, and for trans to cis isomerizations of C11=C12 and C9=C10 of the retinal polyene, as shown by FTIR spectroscopy. The 11-cis and 9-cis 15-syn isomers produced by the latter two reactions are not stable, decaying on the time scale of few seconds to dark state rhodopsin and isorhodopsin by thermal C15=N isomerization, as indicated by time resolved FTIR methods. Flash photolysis of Meta III produces therefore Meta II, dark state rhodopsin, and isorhodopsin. Under continuous illumination, the latter two (or its unstable precursors) are converted as well to Meta II by presumably two different mechanisms. PMID- 15287754 TI - Mutational analysis of protein splicing, cleavage, and self-association reactions mediated by the naturally split Ssp DnaE intein. AB - The ability to separately purify the naturally split Synechocystis sp. PCC6803 (Ssp) DnaE intein domains has allowed detailed examination of both universal and Ssp DnaE intein-specific steps in the protein splicing pathway. By engineering substitutions at both the +1 and penultimate intein positions, we have further characterized intein reaction kinetics in this system. Replacement of the crucial +1Cys with serine decreased N-terminal cleavage and trans-splicing rates; however, this substitution did not prevent splicing or the ability of ZnCl2 to inhibit it. Substitution of the penultimate intein residue (alanine) with a typically conserved histidine did not increase the rate or extent of trans splicing or cleavage under typical assay conditions. Despite the observation that this histidine aids in asparagine cyclization for other inteins, it did not encourage C-terminal cleavage for the Ssp DnaE intein or uncouple it from N terminal cleavage. Both the +1Ser and Ala to His mutants were insensitive to ZnCl2 during trans-cleavage experiments, uncoupling a previously linked inhibition in asparagine cyclization from an inhibition in trans thioesterification detected for the wild-type intein. PMID- 15287755 TI - Stacking-unstacking dynamics of oligodeoxynucleotide trimers. AB - The structure and dynamics of DNA trimers are experimentally assessed using the fluorescent purine analogue 2-aminopurine (2AP), incorporating 2AP between purine and pyrimidine bases to form 5'dXp2APpY3' molecules. Circular dichroism and fluorescence quenching of the 2AP show that the bases are stacked; at the same time, fluorescence decay lifetimes are heterogeneous, indicative of conformational sampling. 2AP does not exhibit the long fluorescence decay time characteristic of the free nucleoside, suggesting that its motions in the trimers bring it into proximity of the neighboring bases, resulting in efficient charge transfer and average fluorescence lifetimes on the order of 1-2 ns. PMID- 15287757 TI - Studies in heteroelement-based synthesis. AB - An account of studies in our group over 30 years is presented, with highlights taken from events that began with our plans for medium ring synthesis, but were soon diverted to organophosphorus chemistry, and that eventually resulted in the investigation of sulfur ylide ring expansions, thiocarbonyl chemistry, azomethine ylides, organotin and organoboron methodology, and control of relative and absolute configuration. PMID- 15287756 TI - Human glycolipid transfer protein: probing conformation using fluorescence spectroscopy. AB - Glycolipid transfer protein (GLTP) is a soluble 24 kDa protein that selectively accelerates the intermembrane transfer of glycolipids in vitro. Little is known about the GLTP structure and dynamics. Here, we report the cloning of human GLTP and characterize the environment of the three tryptophans (Trps) of the protein using fluorescence spectroscopy. Excitation at 295 nm yielded an emission maximum (lambda(max)) near 347 nm, indicating a relatively polar average environment for emitting Trps. Quenching with acrylamide at physiological ionic strength or with potassium iodide resulted in linear Stern-Volmer plots, suggesting accessibility of emitting Trps to soluble quenchers. Insights into reversible conformational changes accompanying changes in GLTP activity were provided by addition and rapid dilution of urea while monitoring changes in Trp or 1-anilinonaphthalene-8 sulfonic acid fluorescence. Incubation of GLTP with glycolipid liposomes caused a blue shift in the Trp emission maximum but diminished the fluorescence intensity. The blue-shifted emission maximum, centered near 335 nm, persisted after separation of glycolipid liposomes from GLTP, consistent with formation of a GLTP glycolipid complex at a glycolipid-liganding site containing Trp. The results provide the first insights into human GLTP structural dynamics by fluorescence spectroscopy, including global conformational changes that accompany GLTP folding into an active conformational state as well as more subtle conformational changes that play a role in GLTP-mediated transfer of glycolipids between membranes, and establish a foundation for future studies of membrane rafts using GLTP. PMID- 15287758 TI - New heterocyclic beta-sheet ligands with peptidic recognition elements. AB - A detailed and comprehensive overview is presented about the design, modeling, and synthesis, as well as spectroscopic characterization, of a new class of beta sheet ligands. The characteristic feature of these compounds is a peptidic chimeric structure formed from a specific combination of aminopyrazolecarboxylic acids with naturally occurring alpha-amino acids. These hybrid peptides are designed with the aid of molecular modeling to exist mainly in an extended conformation. All their hydrogen bond donors and acceptors can be aligned at the bottom face in such a way that a perfect complementarity toward beta-sheets is obtained. Thus the aminopyrazoles impart rigidity and a highly efficient DAD sequence for the recognition of whole dipeptide fragments, whereas the natural alpha-amino acids are designed to mimick recognition sites in proteins, ultimately leading to sequence-selective protein recognition. The synthetic protocols either rely upon solution phase peptide coupling with a PMB protecting group strategy or solid-phase peptide coupling based on the Fmoc strategy, using the same protecting group. In solution, a key building block was prepared by catalytic reduction of a nitropyrazolecarboxylic acid precursor. Subsequently, it was (N-1)-protected with a PMB group, and elongated by HCTU- or T3P-assisted peptide coupling with dipeptide fragments, followed by PyClop-assisted coupling with another nitropyrazolecarboxylic acid building block. Final simultaneous deprotection of all PMB groups with hot TFA completed the high-yield protocol, which works racemization-free. After preparing a similar key building block with an Fmoc protection at N-3, we developed a strategy suitable for automated synthesis of larger hybrid ligands on a peptide synthesizer. Attachment of the first amino acid to a polystyrene resin over the Sieber amide linker is followed by an iterative sequence consisting of Fmoc deprotection with piperidine and subsequent coupling with natural alpha-amino acid via HATU/HOAt. High yields of free hybrid peptides are obtained after mild acidic cleavage from the resin, followed by deprotection of the PMB groups with hot TFA. The new aminopyrazole peptide hybrid compounds were characterized by various spectroscopic measurements including CD spectra, VT, and ROESY NMR experiments. All these accumulated data indicate the absence of any intramolecular hydrogen bonds and strongly support an extended conformation in solution, ideal for docking on to solvent-exposed beta sheets in proteins. Initial results from aggregation tests of pathological proteins with these and related ligands look extremely promising. PMID- 15287759 TI - From branched hydrocarbon propellers to C3-symmetric graphite disks. AB - Branched hydrocarbon propellers (9 and 12) were prepared by stepwise palladium catalyzed Hagihara-Sonogashira coupling reactions and Diels-Alder cycloadditions and submitted to oxidative cyclodehydrogenation by FeCl3 to afford two new giant graphite disks 2 and 3 with 3-fold symmetry in high yield. One of the precursors, the polyphenylene dendrimer 9 containing 150 carbon atoms, was crystallographically characterized. The poorly soluble graphite disks were characterized by isotope-resolved MALDI-TOF mass spectroscopy, and their electronic and vibrational properties were investigated by solid-state UV-vis and Raman spectroscopy. The effect of molecular size and geometry on their properties was discussed. The molecule 2 represents the largest, 3-fold-symmetric all benzenoid graphite disk, and the five-membered rings in molecule 3 open the opportunity to synthesize large bowl-shaped PAHs. PMID- 15287760 TI - Electronic control of chiral quaternary center creation in the intramolecular asymmetric heck reaction. AB - The Boehringer-Ingelheim phosphinoimidazoline (BIPI) ligands were applied to the formation of chiral quaternary centers in the asymmetric Heck reaction. Several different substrates were examined in detail, using more than 70 members of this new ligand class. Hammett relationships were determined through systematic variation of the ligand electronics. All substrates showed essentially the same Hammett behavior, where enantioselectivity increased as the ligands were made more electron-deficient. Ligand optimization has led to catalysts which give the highest enantioselectivities reported to date for these difficult systems. PMID- 15287761 TI - Design and synthesis of a C2-symmetric self-complementary hydrogen-bonding cleft molecule based on the bicyclo[3.3.1]nonane and 4-oxo-5-azaindole framework. formation of channels and inclusion complexes in the solid state. AB - The synthesis of a C2-symmetric cleft molecule 2 based on the fused framework between bicyclo[3.3.1]nonane and 4-oxo-5-azaindole, incorporating a self complementary hydrogen-bonding motif, in both racemic and enantiomerically pure forms is reported. This cleft molecule is reminiscent of analogues of Troger's base though with different cleft dimensions and tilt angles. The framework of 2 provides a building block for the construction of self-assembled hydrogen-bonded supramolecular structures. The solid-state structure of 2 is highly influenced by the limited solubility of (+/-)-2 and (-)-2. The solvents interact with the potential hydrogen-bonding motifs of (+/-)-2 and (-)-2, forming different three dimensional structures as revealed by X-ray diffraction analysis. In the solid state (+/-)-(2)2 x 5DMF forms hydrogen-bonded pleated band structures that build up three-dimensional pens between adjacent bands in which two molecules of DMF are trapped. In contrast, the aggregate obtained from (-)-2, (-)-2 x 2AcOH, showed infinite bands of complex constitution. PMID- 15287762 TI - Transition-state effects in acid-catalyzed aryl epoxide hydrolyses. AB - The hydronium ion-catalyzed hydrolyses of 5-methoxyindene 1,2-oxide and of 6 methoxy-1,2,3,4-tetrohydronaphthalene-1,2-epoxide were each found to yield 75-80% of cis diol and only 20-25% of trans diol as hydrolysis products. The relative stabilities of the cis and trans diols in each system were determined by treating either cis or trans diols with perchloric acid in water solutions and following the approach to an equilibrium cis/trans mixture as a function of time. These studies establish that the trans diol in each system is more stable than the corresponding cis diol. Thus, acid-catalyzed hydrolysis of each epoxide, which proceeds via a carbocation intermediate, yields the less stable cis diol as the major product. Transition-state effects, presumably of a hydrogen-bonding nature, selectively stabilize the transition state for attack of water on the intermediate 2-hydroxy-1-indanyl carbocation leading to the less stable cis diol in this system. Transition-state effects must also be responsible for formation of the less stable cis diol as the major product in the acid-catalyzed hydrolysis of 5-methoxy-1,2,3,4-tetrahydronaphthalene 1,2-epoxide. However, in this system steric effects at the transition state may be more important than hydrogen bonding in determining the cis/trans diol product ratio. The synthesis of 5 methoxyindene 1,2-oxide and a study of its rate of reaction as a function of pH in water and dioxane-water solutions are reported. Both an acid-catalyzed reaction leading to only diol products and a pH-independent reaction yielding 71% of 5-methoxy-2-indanone and 29% of diols are observed; the half-life of its pH independent reaction in water is only 2.4 s. PMID- 15287763 TI - Dioxygen-promoted regioselective oxidative heck arylations of electron-rich olefins with arylboronic acids. AB - Arylations of electron-rich heteroatom-substituted olefins were performed with arylboronic acids. This appears to constitute the first example of palladium(II) catalyzed internal Heck arylations. The novel protocol exploits oxygen gas for environmentally benign reoxidation and a stable 1,10-phenanthroline bidentate ligand to promote the palladium(II) regeneration and to control the regioselectivity. Internal arylation is strongly favored with electron-rich arylboronic acids. DFT calculations support a charge-driven selectivity rationale, where phenyls substituted with electron-donating groups prefer the electron-poor alpha-carbon of the olefin. Experiments, verified by calculations, confirm the cationic nature of the catalytic route. This Heck methodology provides a facile and mild access to functionalized enamides. Controlled microwave heating and increased oxygen pressure were used to further reduce the reaction time to 1 h. PMID- 15287764 TI - Asymmetric total synthesis of (+)-cannabisativine. AB - The asymmetric total synthesis of natural (+)-cannabisativine 1 was completed in 19 steps and 7% overall yield. The key synthetic intermediate 29 was prepared with a high degree of stereocontrol in 12 steps starting from chiral 1 acylpyridinium salt 10. Addition of zinc enolate 11 to pyridinium salt 10 furnished dihydropyridone 12 containing two contiguous stereocenters of the correct absolute configuration. Luche reduction of ketone 16 afforded diol 17 in high yield (96%) and excellent diastereoselectivity. The Mukaiyama-Michael reaction of pyridones 27a/b with O-silyl ketene acetal 32 gave phenyl selenyl ketones 33a/b with complete stereoselectivity. Elimination of cis-beta hydroxyselenides 34 and 35 effected the regiocontrolled preparation of tetrahydropyridine derivative 29. Several approaches to the macrocyclic ring closure of the 13-membered ring were investigated, ultimately leading to the completion of an asymmetric synthesis of the target compound with a high degree of stereocontrol. PMID- 15287765 TI - Kinetic and thermodynamic acidities of pentacarbonyl(cyclobutenylidene)chromium complexes. Effect of antiaromaticity in the conjugate anion. An experimental and computational study. AB - The deprotonation of pentacarbonyl[(3-diethylamino-2,4-dimethyl)cyclobut-2-ene-1 ylidene]chromium (1d) and pentacarbonyl[(3-diethylamino-4-methyl-2 phenyl)cyclobut-2-ene-1-ylidene]chromium (1e) leads to antiaromatic conjugate anions by virtue of their being cyclobutadiene derivatives. Rate constants for the deprotonation of 1d and 1e by P2-Et and pKa values were determined in acetonitrile. Gas-phase B3LYP calculations of 1d, 1e, and their respective conjugate anions, using a generalized basis set, were also performed. Furthermore, for purposes of comparison with carbene complexes of similar structures, but having conjugate anions that are not antiaromatic, corresponding calculations were performed on pentacarbonyl[3-diethylamino-2,5 dimethyl)cyclopent-2-ene-1-ylidene]chromium (5), [dimethylamino(methyl)carbene]pentacarbonylchromium (3a), and [dimethylamino(iso propyl)carbene]pentacarbonylchromium (3b) and their respective conjugate anions, and solution-phase pKa and kinetic measurements were carried out for 3a and 3b. Major points of interest include the effect of antiaromaticity on the kinetic and thermodynamic acidities of 1d and 1e, the large effect of the phenyl group on the gas-phase acidity of 1e, the strong attenuation of the acidities and the effect of the phenyl group in acetonitrile, and the position of the C=C double bonds in the cyclobutadiene ring of the conjugate anion of 1e. PMID- 15287767 TI - Photolysis and oxidation of azidophenyl-substituted radicals: delocalization in heteroatom-based radicals. AB - 2-(4-Azidophenyl)-4,4,5,5-tetramethyl-4,5-dihydro-1H-imidazole-3-oxide-1-oxyl (14), 2-(4-azidophenyl)benzimidazole-1-oxide-3-oxyl (16), 2-(4-azidophenyl)-1,2,6 triphenylverdazyl (19), 2-(3-azidophenyl)-4,4,5,5-tetramethyl-4,5-dihydro-1H imidazole-3-oxide-1-oxyl (21), and (3-azidophenyl)-N-tert-butyl-N-aminoxyl (25) were photolyzed in frozen solution to give S = 3/2 state ESR spectra of the corresponding nitrenophenyl radicals with the following zero-field splitting parameters: |D/hc| = 0.277 cm(-1), |E/hc| < or = 0.002 cm(-1) (7 from 14); |D/hc| = 0.256 cm(-1), |E/hc| < or = 0.002 cm(-1) (8 from 16); |D/hc| = 0.288 cm(-1), |E/hc| < or = 0.002 cm(-1) (9 from 19); |D/hc| = 0.352 cm(-1), |E/hc| = 0.006 cm( 1) (10 from 21); |D/hc| = 0.336 cm(-1), |E/hc| = 0.004 cm(-1) (11 from 25). UB3LYP/6-31G computations and ESR spectroscopic analyses suggest that these are nitreno radicals, even para-linked systems with possible quinonoidal resonance forms. Neat samples of azidophenyl radicals 14 and 21 showed bulk paramagnetic behavior, consistent with the lack of close contacts in their crystal structures. Efforts to make photolabile coordination complexes of 14 and 21 with paramagnetic transition metal ions were unsuccessful: Cu(ClO4)2 x 6H2O instead oxidized them to the corresponding diamagnetic nitrosonium perchlorate salts. PMID- 15287766 TI - Total synthesis of flavocommelin, a component of the blue supramolecular pigment from Commelina communis, on the basis of direct 6-C-glycosylation of flavan. AB - We succeeded in a first total synthesis of flavocommelin (1), a component of the blue supramolecular pigment, commelinin (2), from Commelina communis, by direct 6 C-glycosylation of the flavan 4 using perbenzylglucosyl fluoride 8 in the presence of MS 5 angstroms in CH2Cl2 and a catalytic amount of BF3 x Et2O. After 6-C-glycosylation of 4, oxidation with CAN to flavanone 18 and subsequent 4'-O glycosylation, promoted with a combination of BF3 x Et2O and DTBMP, afforded diglucosylflavanone 20. DDQ oxidation of 20 and deprotection successively gave 1. PMID- 15287768 TI - A novel method for the synthesis of dinucleoside boranophosphates by a boranophosphotriester method. AB - 2'-Deoxyribonucleoside-3'-boranophosphates (nucleotide monomers), including four kinds of nucleobases, were synthesized in good yields by the use of new boranophosphorylating reagents. We have explored various kinds of condensing reagents as well as nucleophilic catalysts for the boranophosphorylation reaction with nucleosides. In the synthesis of dinucleoside boranophosphates, undesirable side reactions occurred at the O-4 of thymine and the O-6 of N2 phenylacetylguanine bases. To avoid these side reactions, additional protecting groups, benzoyl (Bz) and diphenylcarbamoyl (Dpc) groups, were introduced to thymine and guanine bases, respectively. As a result, the condensation reactions proceeded smoothly without any side reactions, and the dimers including four kinds of nucleobases were obtained in excellent yields. In the deprotection of the 5'-DMTr group, Et3SiH was found to be effective as a scavenger for the DMTr cation which caused a P-B bond cleavage. After removal of the other protecting groups by the conventional procedure, four kinds of dinucleoside boranophosphates were obtained in good yields. PMID- 15287769 TI - Divergence of carbonyl ylide reactions as a function of diazocarbonyl compound and aldehyde substituent: dioxolanes, dioxolenes, and epoxides. AB - The products from dirhodium(II) acetate-catalyzed reactions between diazocarbonyl compounds and a series of benzaldehydes demonstrate the extent of competition between intramolecular and intermolecular trapping of carbonyl ylide intermediates and the electronic effects that govern these transformations. With dimethyl diazomalonate, competition exists between dioxolane and epoxide formation so that with p-anisaldehyde only epoxide formation is observed and with p-nitrobenzaldehyde only 1,3-dioxolane products are formed. With methyl diazoacetoacetate, intramolecular trapping of the intermediate carbonyl ylide results in the sole production of dioxolenes. However, the vinyldiazoacetate analogue of methyl diazoacetoacetate, as its tert-butyldimethlsilyloxy derivative, only produces epoxides in its reactions with substituted benzaldehydes. PMID- 15287770 TI - Cp2TiCl-promoted isomerization of trisubstituted epoxides to exo-methylene allylic alcohols on carvone derivatives. AB - The ring-opening reaction of trisubstituted epoxides promoted by Cp2TiCl led to exo-methylene allylic alcohols as major compounds when 0.5 M solutions of the epoxides were added to 0.1 M solutions of the reagent at room temperature in THF. In most cases, the allylic alcohols were contaminated with saturated alcohols. Normal and reverse addition modes led to the alternate product being favored. The different stereochemical outcome of cis- and trans-epoxy acetates is rationalized in terms of mechanistically biased elimination processes. PMID- 15287771 TI - Photolysis of 1-alkylcycloalkanols in the presence of (diacetoxyiodo)benzene and I2. Intramolecular selectivity in the beta-scission reactions of the intermediate 1-alkylcycloalkoxyl radicals. AB - The C-C beta-scission reactions of 1-alkylcycloalkoxyl radicals, generated photochemically by visible light irradiation of CH2Cl2 solutions containing the parent 1-alkylcycloalkanols, (diacetoxy)iodobenzene (DIB), and I2, have been investigated through the analysis of the reaction products. The 1 alkylcycloalkoxyl radicals undergo competition between ring opening and C-alkyl bond cleavage as a function of ring size and of the nature of the alkyl substituent. With the 1-propylcycloheptoxyl, 1-propylcyclooctoxyl,and 1 phenylcyclooctoxyl radicals, formation of products deriving from an intramolecular 1,5-hydrogen atom abstraction reaction from the cycloalkane ring has also been observed. The results are discussed in terms of release of ring strain associated to ring opening, stability of the alkyl radical formed by C alkyl cleavage, and with cycloheptoxyl and cyclooctoxyl radicals, also in terms of the possibility of achieving a favorable geometry for intramolecular hydrogen atom abstraction. PMID- 15287772 TI - Complexation of metals with piperazine-containing azamacrocyclic fluorophores. AB - Azamacrocyclic fluorophores containing piperazine units were synthesized using sequential rhodium-catalyzed regioselective hydroformylation-reductive amination. A piperazine unit is introduced into the macrocycles to act simultaneously as electron donor and binding site. The macrocycles chelate divalent cations, either Zn2+ or Co2+, which considerably enhanced fluorescence. Complexation with Zn2+ was additionally confirmed by NMR. PMID- 15287773 TI - A rapid and stereoselective route to the trans-hydrindane ring system. AB - A short and stereoselective route to the trans-hydrindane derivative, a potential building block for the synthesis of steroidal and related molecules, was achieved by the operation of indium, tin, and ruthenium based reagents, starting from a tetrabromo norbornyl derivative. PMID- 15287774 TI - Electrochemical partial fluorination of organic compounds. 74. Efficient anodic synthesis of 2-fluoro- and 2,3-difluoro-2,3-dihydrobenzofuran derivatives. AB - Anodic fluorination of 3-substituted benzofuran derivatives in a variety of fluoride salts resulted in the formation of three fluorinated products; two stereoisomers of 2,3-difluoro-2,3-dihydrobenzofuran (cis and trans) and cis-2 fluoro-3-hydroxy-2,3-dihydrobenzofuran derivatives. Dehydrofluorination of the main products, cis-difluoro derivatives, furnished the nonaromatic 2-fluoro-3 benzofuranyledene derivatives instead of the aromatic 2-fluorobenzofuran derivatives. PMID- 15287775 TI - Highly stereoselective approach to alk-2-yne-1,4-diols by oxazaborolidine mediated reduction of alk-2-yne-1,4-diones. AB - We performed the borane-mediated reduction of a series of symmetrical alk-2-yne 1,4-diones (5) in the presence of the oxazaborolidine (R)-6 to afford (R,R)-alk-2 yne-1,4-diols ((R,R)-1) in good yields and high stereoselectivities (up to 99.9% ee). In some cases, the stereochemical purity of 1 was improved by a two-step process: (i) temporary transformation of 1 into its vic-dibromo derivatives 9, which allowed us to remove the minor meso isomer by chromatography, and (ii) regeneration of the enantioenriched diols 1 with SmI2. Reduction of the hexacarbonyldicobalt complexes 8 derived from 5 was also successful. PMID- 15287776 TI - RCM approaches toward the diastereoselective synthesis of vicinal trans diaminocyclitols from L-serine. AB - Starting from L-serine, the asymmetric synthesis of four diaminocyclitol derivatives as sugar-based glycosidase inhibitors has been achieved using ring closing metathesis (RCM) as a key step. Introduction of vicinal trans-diamino functionality onto the acyclic precursors was accomplished by highly diastereoselective addition of Grignard reagent to imine, and the elaboration of polyhydroxylic groups was effected via diastereoselective olefin epoxidation or dihydroxylation. The absolute configurations of final products were confirmed by 2D NMR studies. PMID- 15287777 TI - Probes for narcotic receptor-mediated phenomena. 33. Construction of a strained trans-5,6-ring system by displacement of a nitro-activated aromatic fluorine. synthesis of the penultimate oxide-bridged phenylmorphans. AB - The synthesis of the ortho- and para-e isomers in the oxide-bridged 5 phenylmorphan series of rigid tetracyclic compounds was accomplished via rac-5-(2 fluoro-5-nitrophenyl)-2-methyl-2-azabicyclo[3.3.1]nonan-9beta-ol ((+/-)-10), an intermediate containing an aromatic nitro-activated fluorine atom. The fluorine atom was used as the leaving group for the formation of the strained tetracyclic trans-fused 5,6-ring system in rac-(1alpha,4aalpha,9aalpha)-1,3,4,9a-tetrahydro-2 methyl-6-nitro-2H-1,4a-propanobenzofuro[2,3-c]pyridine ((+/-)-11), although preference for cis ring fusion during the formation of tricyclic tetra- and hexahydrodibenzofurans has been well-documented. Single-crystal X-ray crystallographic study of the desired para-e isomer ((+/-)-2), as well as of two intermediates in its synthesis, provided assurance of the correct structures. The e-isomers are among the last of the 12 oxide-bridged 5-phenylmorphans to be synthesized. We envisioned the syntheses of these rigid, tetracyclic compounds in order to determine the three-dimensional pattern of a ligand that would enable interaction with opioid receptors as agonists or antagonists. PMID- 15287778 TI - 7-substituted 2-azabicyclo[2.2.1]heptanes as key intermediates for the synthesis of novel epibatidine analogues; synthesis of syn-and anti-isoepiboxidine. AB - Neighboring group participation by the 2-nitrogen in anti-7-bromo-2-benzyl-2 azabicyclo[2.2.1]heptane allows ready nucleophilic substitution at the 7-position by C, N, O, and halogen nucleophiles and opens the way to a range of novel 7 substituted 2-azabicyclo[2.2.1]heptanes. Conversion of an anti-7-ethoxycarbonyl group into a methylisoxazole ring provides anti-isoepiboxidine, a conversion that is possible even without protection of the secondary bicyclic nitrogen. Successful base-induced epimerization alpha to the carbonyl of the anti-7 ethoxycarbonyl derivative gives the syn-stereoisomer and hence syn isoepiboxidine. PMID- 15287779 TI - A convenient protocol for the synthesis of ligands from a 4-methyl-3,5 diacylaminophenyl platform. AB - The synthesis of stable and highly organized phenanthroline, terpyridine, and pyridino-oxazoline ligands bearing one or two 4-methyl-3,5-diacylaminophenyl modules equipped with two lateral dialkoxyphenyl groups has been performed using EDC.HCl and DMAP reagents in the final coupling reaction. Evidently, in the final ligands and in the solid state intermolecular hydrogen bonding maintains the coherence of the tridimensional structure as clearly evidenced by FT-IR and X-ray diffraction spectroscopy in the cases of the methoxy ligands. The supramolecular packing is also maintained by additional pi-pi stacking interactions. PMID- 15287780 TI - Palladium-catalyzed transformation of cyclobutanone O-benzoyloximes to nitriles via C-C bond cleavage. AB - Palladium-catalyzed transformation of cyclobutanone O-benzoyloximes to a variety of nitriles is described. The reaction may proceed via two important steps, that is, (i) oxidative addition of the N-O bond of oximes to Pd(0) to give a cyclobutylideneaminopalladium(II) species and (ii) beta-carbon elimination of this species to afford a reactive alkylpalladium species. The kind of products is very dependent on the nature of substituents on the cyclobutane ring. The direction of the C-C bond cleavage is controlled by the kind of ligand employed. The sequential reaction composed of the C-C bond cleavage and the subsequent intra- and intermolecular C-C bond formations via the corresponding alkylpalladium species is also demonstrated. For example, an oxime having an alkynyl moiety at a suitable position reacts with a variety of alkenes to afford nitriles bearing dienylcyclopentane moiety in moderate to good yields. PMID- 15287781 TI - Rapid oligosaccharide synthesis using a fluorous protective group. AB - The Bfp-OH, a novel fluorous protecting reagent, was able to be easily prepared. The Bfp group was readily introduced to a carbohydrate, removed in high yield, and recyclable after cleavage. The use of the Bfp group made it possible to synthesize a pentasaccharide by minimal column chromatography purification. Each synthetic intermediate was able to be easily purified only by simple fluorous organic solvent extraction and monitored by TLC, NMR, and MS. PMID- 15287782 TI - Boron-complexation strategy for use with 1-acyldipyrromethanes. AB - 1-Acyldipyrromethanes are important precursors in rational syntheses of diverse porphyrinic compounds. 1-Acyldipyrromethanes are difficult to purify, typically streaking upon chromatography and giving amorphous powders upon attempted crystallization. A solution to this problem has been achieved by reacting the 1 acyldipyrromethane with a dialkylboron triflate (e.g., Bu2B-OTf or 9-BBN-OTf) to give the corresponding B,B-dialkyl-B-(1-acyldipyrromethane)boron(III) complex. The reaction is selective for a 1-acyldipyrromethane in the presence of a dipyrromethane. The 1-acyldipyrromethane-boron complexes are stable to routine handling, are soluble in common organic solvents, are hydrophobic, crystallize readily, and chromatograph without streaking. The 1-acyldipyrromethane can be liberated in high yield from the boron complex upon treatment with 1-pentanol. Alternatively, the 1-acyldipyrromethane-boron complex can be used in the formation of a trans-A2B2-porphyrin. In summary, the boron-complexation strategy has broad scope and greatly facilitates the isolation of 1-acyldipyrromethanes. PMID- 15287783 TI - Dual behavior of masked o-benzoquinones in intermolecular Diels-Alder reactions with acyclic dienes: a rapid entry to polyfunctionalized bicyclo[2.2.2]oct-5-en-2 ones and cis-decalins. AB - The potentiality of the masked o-benzoquinones, i.e., 6,6-dimethoxy-2,4 cyclohexadienones 5-8, to react both as dienes and dienophiles in their intermolecular reactions has been demonstrated. The masked o-benzoquinones (MOBs) 5-8 generated in situ from 2-methoxyphenols 1-4 underwent intermolecular Diels Alder cycloadditions with acyclic 1,3-dienes 9a-e to provide bicyclo[2.2.2]octenones 10a-f-13a-f along with cis-decalin derivatives 14a-f-17a f with regio- and stereoselectivity, except in the case of MOB 8. The formation of cis-decalins in these Diels-Alder reactions illustrates the dienophilic character of MOBs, in addition to their general behavior as dienes. The ratio of the two cycloadducts obtained in each reaction as a result of the dual character of MOBs depends on the nature and/or position of the substituents on both the cyclohexadienone moiety and the added conjugated acyclic diene. All of the cycloadducts resulted from the diene property of MOBs in intermolecular Diels Alder reactions smoothly underwent Cope rearrangement to furnish cis-decalins as sole products in excellent to quantitative yields. PMID- 15287784 TI - Asymmetric conjugate addition for the preparation of syn-1,3-dimethyl arrays: synthesis and structure elucidation of capensifuranone. AB - The synthesis of capensifuranone (1) has been achieved by the application of developments for asymmetric conjugate addition reactions of organocopper reagents with nonracemic N-enoyl-4-phenyl-1,3-oxazolidinones for the preparation of 1,3 syn-dimethyl arrays. The assignment of relative and absolute stereochemistry of 1 has been made following the high-field NMR characterizations of synthetic diol derivatives. The previously unassigned C4 stereochemistry of 1 was determined to be of the (S)-configuration. The thermodynamic equilibration of capensifuranone and its C4 diastereomer has been examined. PMID- 15287785 TI - Stereoselective synthesis of optically active disilanes and selective functionalization by the cleavage of silicon-naphthyl bonds with bromine. AB - Optically active disilanes with one chiral silicon center, (R)-1,2-dimethyl-1 (naphth-1-yl)-1,2,2-triphenyldisilane and (R)-1,2,2-trimethyl-2-(4-methoxynaphth 1-yl)-1-(naphth-1-yl)-1-phenyldisilane, were obtained by the reaction of (S) methyl(naphth-1-yl)phenylchlorosilane (> 99% ee) with methyldiphenylsilyllithium or by the reaction of methyldiphenylchlorosilane with optically active (S) methyl(naphth-1-yl)phenylsilyllithium and by the reaction of (S)-methyl(naphth-1 yl)phenylchlorosilane (> 99% ee) with dimethyl(4-methoxynaphth-1-yl)silyllithium. Under the optimized conditions, the reactions proceeded with almost complete inversion for the cholorosilanes and retention for the silyl anions. Optically active disilanes with two chiral centers, (1R,2R)-1,2-dimethyl-1,2-di(naphth-1 yl)-1,2-diphenyldisilane and (1S,2S)-1,2-di(4-methoxynaphth-1-yl)-1,2-dimethyl 1,2-diphenyldisilane, were obtained in high optical purity by the reactions of corresponding optically active halogenosilanes (Cl or F) with optically active silyllithiums. The silicon-silicon bond and the silicon-naphthyl bond of (R) 1,1,2-trimethyl-1,2-di(naphth-1-yl)-2-phenyldisilane and (1R,2R)-1,2-dimethyl-1,2 di(naphth-1-yl)-1,2-diphenyldisilane were cleaved without selectivity on bromination. The silicon-(4-methoxynaphth-1-yl) bond of (R)-1,2,2-trimethyl-2-(4 methoxynaphth-1-yl)-1-(naphth-1-yl)-1-phenyldisilane was regiospecifically cleaved, followed by the stereoselective cleavage of the remaining chiral silicon naphthyl bond (94% inversion). Although the silicon-(4-methoxynaphth-1-yl) bonds of (1S,2S)-1,2-di(4-methoxynaphth-1-yl)-1,2-dimethyl-1,2-diphenyldisilane (> 99% ee) were regioselectively cleaved without silicon-silicon bond scission, remarkable racemization could not be avoided during the one-pot reaction. PMID- 15287786 TI - Competing pathways in the [2 + 2] cycloadditions of cyclopentyne and benzyne. A DFT and ab initio study. AB - The [2 + 2] cycloadditions of cyclopentyne and benzyne to ethylene are explored at the B3LYP and CASSCF levels, supplemented by CCSD(T) and CAS-MP2 calculations at the stationary points. The biradical path in the benzyne system is computed to be about 4.1 kcal/mol lower than the concerted path, consistent with the experimentally observed loss of original stereochemistry in this cycloaddition. However, computations fail to confirm the 99% stereoretention in the corresponding reaction of cyclopentyne. The concerted and biradical paths in the latter reaction are found to involve nearly isoenergetic barriers, thus predicting only about 75% stereoretention. More sophisticated theoretical methods seem to be needed to resolve the issue in the cyclopentyne system. PMID- 15287787 TI - Gas electron diffraction analysis on S-methyl thioacetate, CH3C(O)SCH3. AB - The molecular structure of S-methyl thioacetate, CH3C(O)SCH3, was determined by gas electron diffraction (GED) with the assistance of quantum chemical calculations (B3LYP/6-31G and MP2/6-31G). Experimental and theoretical methods result in a structure with syn conformation (C=O double bond syn with respect to the S-C(H3) single bond). The following skeletal geometric parameters were derived from the GED analysis (ra values with 3sigma uncertainties): C=O = 1.214(3), C-C = 1.499(5), S-C(sp2) = 1.781(6), S-C(sp3) = 1.805(6) angstroms, O=C C = 123.4(8) degrees, O=C-S = 122.8(5) degrees and C-S-C = 99.2(9) degrees. PMID- 15287788 TI - Kinetics and mechanism of the pyridinolysis of s-4-nitrophenyl 4-substituted thiobenzoates in aqueous ethanol. AB - The pyridinolysis of S-4-nitrophenyl 4-X-substituted thiobenzoates (X = H, Cl, and NO2; 1, 2, and 3, respectively) is studied kinetically in 44 wt % ethanol water, at 25.0 degrees C and an ionic strength of 0.2 M (KCl). The reactions are measured spectrophotometrically (420-425 nm) by following the appearance of 4 nitrobenzenethiolate anion. Pseudo-first-order rate coefficients (kobsd) are obtained throughout, under excess of amine over the substrate. Plots of kobsd vs [free amine] at constant pH are linear with the slope (kN) independent of pH. The Brnsted-type plot (log kN vs pKa0 of the conjugate acids of the pyridines) for the reactions of thiolbenzoate 1 is curved with a slope at high pKa, beta1 = 0.20, and slope at low pKa0, beta2 = 0.94. The pKa value for the center of the Brnsted curvature is pKa0 = 9.7. The pyridinolysis of thiolbenzoates 2 and 3 show linear Brnsted-type plots of slopes 0.94 and 1.0, respectively. These results and other evidence indicate that these reactions occur with the formation of a zwitterionic tetrahedral intermediate (T+/-). For the pyridinolysis of thiolbenzoate 1, breakdown of T+/- to products (k2 step) is rate-limiting for weakly basic pyridines and T+/- formation (k1 step) is rate-determining for very basic pyridines. The k2 step is rate-limiting for the reactions of thiolbenzoates 2 and 3. The smallest pKa0 value for the reaction of 1 is due to the weakest electron withdrawal of H (relative to Cl and NO2) in the acyl group, which results in the smallest k-1/k2 ratio. The pKa0 values for the title reactions are smaller than those for the reactions of secondary alicyclic amines with thiolbenzoates 1-3. This is attributed to a lower leaving ability from the T+/- of pyridines than isobasic alicyclic amines. The lower p value found for the pyridinolysis of 2,4-dinitrophenyl benzoate (pKa0 = 9.5), compared with that for the pyridinolysis of 1, is explained by the greater nucleofugality from T+/- of 2,4-dinitrophenoxide than 4-nitrobenzenethiolate, which renders the k-1/k2 ratio smaller for the reactions of the benzoate relative to thiolbenzoate 1. The title reactions are also compared with the aminolysis of similar thiolbenzoates in other solvents to assess the solvent effect. PMID- 15287789 TI - A soluble-polymer system for the asymmetric transfer hydrogenation of ketones. AB - The appropriate combination of methacrylate polymers permits the synthesis of a soluble polymer for use in ruthenium(II)-catalyzed asymmetric transfer hydrogenation reactions. Using a 7:3 copolymer of a poly(ethylene glycol) ester and a hydroxyethyl ester, a derived ruthenium(II)/norephedrine complex catalyses reduction of acetophenone in up to 95% yield and 81% ee. PMID- 15287790 TI - New approach to indole alkaloids based on the intramolecular Pauson-Khand reaction. AB - The synthesis of indoles bearing alkenyl and alkynyl moieties in different positions of the nucleus is described. These compounds are used as substrates for the intermolecular Pauson-Khand reaction leading to tetracyclic cyclopentenones with formation of additional five- to seven-membered rings. Products are related to alkaloids such as mitosenes, clausines, ergotamines, or apogeissochizines. PMID- 15287791 TI - Efficient proton-templated synthesis of 18- to 38-membered tetraimino(amino)diphenol macrocyclic ligands: structural features and spectroscopic properties. AB - A whole range of Robson-type tetraiminodiphenol macrocyclic ligands have been prepared as their perchlorate salts [H4L](ClO4)2 in high yield (ca. 90%) by a single-step [2 + 2] condensation reaction between 4-methyl(or tert-butyl)-2,6 diformyl(or diacyl)phenols and alpha,omega-diaminoalkanes (C2-C12) in the presence of acetic acid and NaClO4. The reduction of these 18- to 38-membered macrocyclic salts with NaBH4 have afforded corresponding tetraaminodiphenol macrocycles H2L'. The X-ray crystal structures of two of the tetraiminodiphenol macrocycles with the C2 and C4 lateral chains have been determined, and the optimized configurations for all of the macrocycles have been obtained by molecular mechanics calculations. The macrocycles have been characterized by elemental analysis and by IR, absorption, emission, and NMR spectroscopic study. The protonated tetraiminodiphenol macrocycles exhibit strong fluoroscence in methanol, acetonitrile, and nitromethane and undergo quenching when treated with triethylamine. The neutral macrocycles H2L, isolated by treating [H4L](ClO4)2 with excess of triethylamine, lack luminescence, as do the reduced tetraaminodiphenol macrocycles H2L'. The hydrolytic cleavage of [H4L](ClO4)2 has been studied. PMID- 15287792 TI - A copper- and amine-free sonogashira reaction employing aminophosphines as ligands. AB - An efficient Pd-catalyzed Sonogashira coupling reaction was achieved in the absence of a copper salt or amine with an inorganic base and easily prepared, air stable aminophosphine ligands in commonly used organic solvents; good to excellent yields were obtained. Under optimized reaction conditions, the Sonogashira coupling reaction occurred selectively when an enyne substrate was employed and no Heck reaction product was detected; acetone-masked acetylene and trimethylsilylacetylene can also be efficiently coupled, providing a method to make terminal alkynes. PMID- 15287793 TI - Synthesis of l-lyxo-phytosphingosine and its 1-phosphonate analogue using a threitol acetal synthon. AB - The first synthesis of an isosteric phosphonate analogue of the aminotriol lipid phytosphingosine (3), together with an improved synthesis of (2S,3S,4S) phytosphingosine (2), are described. A key intermediate is 3-pentylidene acetal 9, which was prepared in two steps from dimethyl 2,3-O-benzylidene-d-tartrate (7). PMID- 15287794 TI - Studies on the chemical stability and synthetic utility of an oxazolidine linker for solid-phase chemistry. AB - A chemical stability study on the oxazolidine linker system has been carried out using a dual-linker analytical construct within a parallel reaction scan. The study established the compatibility of the oxazolidine platform with a wide range of commonly employed synthetic reaction conditions including nucleophilic, oxidizing, and reducing conditions. The scan was further used to probe and optimize acidic conditions under which the oxazolidine could release the substrate from the solid-support and to identify reagents that could cleave while retaining other acid-labile groups. The solid-phase synthesis of a small molecular array established the utility of oxazolidine aldehyde 1 as a building block for asymmetric chemistry while exploiting the data generated by the reaction scan. PMID- 15287795 TI - Energetic preferences for alpha,beta versus beta,gamma unsaturation. AB - Density functional theory has been applied at the B3LYP/6-311+G(d,p)//B3LYP/6 31G(d) level to examine the energetics of alpha,beta- versus beta,gamma unsaturation for some common organic functional groups. Specifically, the relative stabilities of allyl-X (H2C=CHCH2X) and 1-propenyl-X (H3CCH=CHX) isomers have been computed for X = methyl, vinyl, phenyl, formyl, acetyl, methoxy, methylthio, methylsulfinyl, methylsulfonyl, sulfamoyl, and methoxysulfonyl, and the results are compared to available experimental data. The intrinsic preference of 3 kcal/mol for the 1-propenyl isomer when X = CH3 is exceeded by 2-4 kcal/mol for first-row conjugating groups, but it is not met for the sulfur-containing groups. In particular, alpha,beta-unsaturation is favored by less than 1 kcal/mol for the sulfone and sulfonamide analogues, while it is preferred by 8 kcal/mol for the vinyl-substituted case. Detailed structural results and torsional energy profiles are also reported. PMID- 15287796 TI - Thermochemistry of 1,3-dithiacyclohexane 1-oxide (1,3-dithiane sulfoxide): calorimetric and computational study. AB - The enthalpies of combustion and sublimation of 1,3-dithiacyclohexane 1-oxide (1,3-dithiane sulfoxide, 2) were measured by a rotating-bomb combustion calorimeter and the Knudsen effusion technique, and the gas-phase enthalpy of formation was determined, DeltafH degrees m(g) = -98.0 +/- 1.9 kJ mol(-1). This value is not as large (negative) as could have been expected from comparison with thermochemical data available for the thiane/thiane oxide reference system. High level ab initio molecular orbital calculations at the MP2(FULL)/6-31G(3df,2p) level were performed, and the optimized molecular and electronic structures of 2 afforded valuable information on (1) the relative conformational energies of 2 axial and 2-equatorial--the latter being 7.1 kJ mol(-1) more stable than 2-axial, (2) the possible involvement of nS --> sigma*(C-S(O)) hyperconjugation in 2 equatorial, (3) the lack of computational evidence for sigma(S-C) --> sigma*(S-O) stereoelectronic interaction in 2-equatorial, and (4) the relevance of a repulsive electrostatic interaction between sulfur atoms in 1,3-dithiane sulfoxide, which apparently counterbalances any nS --> sigma*(C-S(O)) stabilizing hyperconjugative interaction and accounts for the lower than expected enthalpy of formation for sulfoxide 2. PMID- 15287797 TI - Water effect on the o-h dissociation enthalpy of para-substituted phenols: a DFT study. AB - The effect of water on the O-H bond dissociation enthalpy (BDE) of para substituted phenols has been investigated by means of DFT calculations. It is shown that the experimental BDE values are fairly well-reproduced by simple B3LYP/6-31G* calculations carried out on the phenol/phenoxyl-water complexes taking into account only hydrogen-bonding (HB) interactions of water molecules with molecular sites (HB model). On the contrary, the BDE values computed with the polarizable continuum model (PCM/B3LYP/6-31G*)8 are overestimated by about 3 4 kcal/mol. Discrepancy between theory and experiment increases using the PCM method in addition to the HB model. Calculations show that, in general, the HB interaction with water molecules decreases the BDE of phenols bearing electron releasing groups while increasing the BDE of phenols bearing electron-withdrawing substituents. This opposite effect is explained by considering the resonance structures with charge separation both in phenols and in phenoxyl radicals. With electron donors, the phenoxyl radical is preferentially stabilized by the HB acceptor interaction with two water molecules, while with electron acceptors the phenol is preferentially stabilized by the HB donor interaction with one water molecule. PMID- 15287798 TI - A Highly stereospecific and efficient synthesis of homopentafluoro- phenylalanine. AB - A short and efficient synthesis of homopentafluorophenylalanine (6) from oxazolidine aldehyde 1 in 57% overall yield and in > 98% ee is described. The enantiomeric excess of the product was determined by 19F NMR analysis of the coupling product derived from 5 and L-Ser(O-t-Bu)-OCH3, by comparison to a dipeptide obtained from racemic 5. PMID- 15287799 TI - First tandem free-radical cyclization reaction of alkylenecyclopropanes: a novel and efficient method for the preparation of 2-(3,4-dihydronaphthalen-2-yl)malonic acid diethyl esters. AB - Alkylidenecyclopropanes undergo Mn(OAc)3-mediated addition with malonate, leading to dihydronaphthalene derivatives in moderate yields. PMID- 15287801 TI - Efficient method for the synthesis of hetarenoindanones based on 3-arylhetarenes and their conversion into hetarenoindenes. AB - A series of hetarenoindanones have been prepared by direct double metalation of the appropriate 3-phenylhetarene with butyllithium in the presence of TMEDA followed by treatment of the resulting dilithium compound with ethyl N,N dimethylcarbamate. All hetarenoindanones were reduced according to Wolff-Kishner by hydrazine in the presence of KOH to the corresponding hetarenoindenes. PMID- 15287800 TI - A direct, efficient method for the preparation of n6-protected 15n-labeled adenosines. AB - N6-Protected adenosines have been prepared from inosines by activation of the C6 position and Pd-catalyzed coupling with amides. An efficient route to [6-15NH2] N6-benzoyladenosine and [1-15N,6-15NH2]-N6-benzoyladenosine has been achieved. PMID- 15287802 TI - Synthetic approach to substituted cyclopropanes based on the coupling reaction of lithiated chloroalkyloxazolines with Fischer carbene complexes. AB - Regioselective addition of lithiated oxazoline 2a, easily available from 2-(1 chloroethyl)-4,4-dimethyl-2-oxazoline 1a (LDA, THF, -98 degrees C), to alpha,beta unsaturated Fischer carbene complexes 3 afforded cyclopropylcarbene complexes 4 as sole diastereoisomers. Exposure of carbene complexes 4a-c (M = Cr) to air and sunlight gave cyclopropane carboxylate derivatives 5a-c. A plausible mechanistic explanation is proposed. Moreover, when lithiated oxazoline 2b was generated from 1b in the presence of the carbene complex 3a,b, the oxazolinylcyclopropane carboxylates 6a,b formed as a 1:1 mixture of diastereoisomers. Chiral lithiated oxazoline 2c added regioselectively and diastereoselectively to chromium complexes 3a,b and to tungsten complexes 3d,e, leading, after oxidation of the metal fragment, to esters 7a,b with good diastereoselectivity (dr = 4:1). The reaction of lithiated oxazoline 2d with chromium complex 3b and tungsten complex 3e proceeded less diastereoselectively, furnishing, in both cases, after oxidation, the ester 7c as a 3:2 diastereoselective mixture. PMID- 15287803 TI - Polysulfane antitumor agents from o-benzyne. An odd-even alternation found in the stability of products o-C6H4Sx (x = 1-8). AB - Benzyne is shown to add elemental sulfur and give rise to a series of polysulfane compounds. A computational and experimental study is presented. Odd-membered o C6H4Sx rings (x = 1-8), except x = 1, which suffers from ring strain, have enhanced stability compared to even-membered rings. The acquisition of "odd-even" data may shed new light, revealing patterns on polysulfane stability and structure. PMID- 15287804 TI - Regioselective and stereoselective nucleophilic ring opening of trifluoromethylated cyclic sulfates: asymmetric synthesis of both enantiomers of syn-(3-trifluoromethyl)isoserine. AB - A novel and efficient enantioselective synthesis of both enantiomers of syn-(3 trifluoromethyl)isoserine was achieved. Ring opening of trifluoromethylated cyclic sulfates 3, derived from enantiopure trifluoromethylated vicinal diols 2, with various nucleophiles occurred exclusively at C2 with inversion of chirality. Treatment of 4c and 4d, obtained by nucleophilic opening of 3a and 3b with PhCO2NH4, with (CF3SO2)2O followed by substitution with sodium azide, Jones oxidation, and hydrogenolysis furnished (2S,3S)-(N-benzoyl)-3 (trifluoromethyl)isoserine 9a and (2R,3R)-(N-benzoyl)-3 (trifluoromethyl)isoserine 9b, respectively. PMID- 15287806 TI - Hydrophosphorylation of alkenes with dialkyl phosphites catalyzed by Mn(III) under air. AB - A facile method for the synthesis of organophosphonates from alkenes and dialkyl phosphites was developed by the use of Mn(II) under air. Thus, the reaction of 1 octene with diethyl phosphite in the presence of Mn(OAc)2 (5 mol %) under air at 90 degrees C led to diethyl octylphosphonate (78%) and diethyl (2 hexyl)decylphosphonate (6%). Internal alkenes such as cis-2-octene gave a regioisomeric mixture of the corresponding hydrophosphorylation products in 84% yields. PMID- 15287805 TI - Birch reduction of (-)-ephedrine. Formation of a new, versatile intermediate for organic synthesis. AB - The reduction of (-)-ephedrine by lithium in liquid ammonia resulted in the formation of S-1-(1,4-cyclohexadien-1-yl)-N-methyl-2-propanamine. In addition to the reduction of the aromatic ring, the hydroxy group was reduced as well. The resulting 1,4-cyclohexadienyl group is a potentially versatile intermediate for further synthetic transformations. The ozonolysis of this group was investigated, producing derivatives of beta-keto-delta-methylamino esters and beta-keto aldehydes which could be subsequently converted to heterocycles. The restriction to rotation of the C-N bond in N-benzoyl-1-(1,4-cyclohexadien-1-yl)-N-methyl-2 propanamine is described. PMID- 15287808 TI - Synthesis of 1,4-annulated cyclooctatetraenophanes based on a novel cubane building block approach. AB - A general synthetic approach to strained 1,4-annulated cyclooctatetraene-based cyclophanes is described. A key feature in this approach is exploitation of the cubane core as a masked cyclooctatetraene synthon. Thus, 1,4-disubstituted cubanes 3 and 4 used as precursors to cyclooctatetraenophanes have been prepared in four steps from the readily available 1,4-cubanedicarboxaldehyde (5). The synthesis of 3 was effected by palladium/copper-mediated coupling of 1,4 bis[(Z,Z)-2-iodovinyl]cubane (6) and 1,4-bis[(Z,Z)-but-1-en-3-ynyl]cubane (8). For the synthesis of 4, on the other hand, modified Eglington-Glaser coupling was applied for the macrocyclization step. The general characteristic of Rh(I) to induce [2 + 2] cycloreversion of the cubane core to syn tricyclo[4.2.0.0(2,5)]octa-3,7-diene followed by thermal rearrangement to cyclooctatetraene was applied as a key structural transformation toward targeted cyclooctatetraenophanes 1 and 2. PMID- 15287809 TI - A new, efficient method for direct alpha-alkenylation of beta-dicarbonyl compounds and phenols using alkenyltriarylbismuthonium salts. AB - Direct alpha-alkenylation of beta-keto esters, beta-diketone, and phenols with alkenyltriarylbismuthonium salts proceeded smoothly in the presence of 1,1,3,3 tetramethylguanidine to afford the corresponding alpha-alkenylated carbonyl compounds (beta,gamma-unsaturated carbonyl compounds) in good yields. The high leaving ability of the triarylbismuthonio group is a key driving force to achieve the C-C bond formation at the vinylic carbon under mild conditions. PMID- 15287807 TI - Facile synthesis of saponins containing 2,3-branched oligosaccharides by using partially protected glycosyl donors. AB - Two natural saponins 1 and 2, isolated from Solanum indicum L., and containing 2,3-branched sugar moieties, have been efficiently synthesized. Partially protected monosaccharide and disaccharide donors were used to facilitate target synthesis. Stereo factors were critical in incorporating 2,3-branched sugars on steroid aglycones. Saponin 1 was synthesized in five steps and 30% overall yield, while saponin 2 was obtained using six straightforward sequential reactions in 31% overall yield. Saponin 2 shows promising cytotoxic activity toward human hepatocellular carcinoma BEL-7402 with an IC50 of <6 microg/mL. PMID- 15287810 TI - Synthesis of alpha-ketoamides from a carbamoylsilane and acid chlorides. AB - Treatment of acid chlorides with a carbamoylsilane affords alpha-ketoamides. In some instances, in situ reaction of additional carbamoylsilane with these products yielded alpha-organyl-alpha-siloxymalonamides. PMID- 15287811 TI - Samarium(III) carbenoid as a competing reactive species in samarium-promoted cyclopropanation reactions. AB - The trivalent samarium carbenoid I2SmCH2I-promoted cyclopropanation reactions with ethylene have been investigated and are predicted to be highly reactive, similarly to the divalent samarium carbenoid ISmCH2I. The methylene transfer and carbometalation pathways were explored and compared with and without coordination of THF solvent molecules to the carbenoid. The methylene transfer was found to be favored, with the barrier to reaction going from 12.9 to 9.2 kcal/mol compared to barriers of 15.4-17.5 kcal/mol for the carbometalation pathway upon the addition of one THF molecule. PMID- 15287815 TI - Genetic risk factors in eating disorders. AB - Eating disorders such as anorexia nervosa and bulimia nervosa involve complex and interacting mechanisms. Formal genetic studies suggest that there is a substantial genetic influence for these disorders. Animal models of eating disorders are scarce. Candidate gene studies have initially focused on the serotonergic and other central neurotransmitter systems and on genes involved in body weight regulation. Most of the studies, including meta-analysis, have yielded negative results; only a single positive finding has been replicated independently. Recently, systematic genome-wide scans based on families with two or more individuals with an eating disorder (anorexia nervosa or bulimia nervosa) revealed initial linkage regions on chromosomes 1, 3, and 4 (anorexia nervosa) and 10p (bulimia nervosa). Fine mapping of one of these regions led to the identification of genes where an association with anorexia nervosa was detected. Currently treatment of patients with eating disorders can not rely on results of molecular genetic studies. PMID- 15287816 TI - Gene expression profiling as a diagnostic tool in acute myeloid leukemia. AB - The standard methods for establishing the diagnosis of acute leukemias are cytomorphology and cytochemistry in combination with multiparameter immunophenotyping. Cytogenetics, fluorescence in situ hybridization, and PCR based assays add important information regarding biologically defined and prognostically relevant subgroups, and allow a comprehensive diagnosis of well defined subentities. In the clinical setting, a better understanding of the clinical course of distinct, biologically defined disease subtypes is the basis for a selection of disease-specific therapeutic approaches. As knowledge of deregulated pathways in leukemia increases and accelerates the development of new therapeutics, a detailed and comprehensive diagnostic tool is required. Microarray technology, which quantifies gene expression intensities of thousands of genes in a single analysis, has the potential to become an essential tool for the molecular classification of leukemias. It may, therefore, be used as a routine method for diagnostic purposes in the near future. Furthermore, gene expression profiling may also lead to the detection of new biologically defined and clinically relevant subtypes in leukemia and guide therapeutic decision making in the future. PMID- 15287817 TI - Genetic variation and lactose intolerance: detection methods and clinical implications. AB - The maturational decline in lactase activity renders most of the world's adult human population intolerant of excessive consumption of milk and other dairy products. In conditions of primary or secondary lactase deficiency, the lactose sugars in milk pass through the gastrointestinal tract undigested or are partially digested by enzymes produced by intestinal bacterial flora to yield short chain fatty acids, hydrogen, carbon dioxide, and methane. The undigested lactose molecules and products of bacterial digestion can result in symptoms of lactose intolerance, diarrhea, gas bloat, flatulence, and abdominal pain. Diagnosis of lactose intolerance is often made on clinical grounds and response to an empiric trail of dietary lactose avoidance. Biochemical methods for assessing lactose malabsorption in the form of the lactose breath hydrogen test and direct lactase enzyme activity performed on small intestinal tissue biopsy samples may also be utilized. In some adults, however, high levels of lactase activity persist into adulthood. This hereditary persistence of lactase is common primarily in people of northern European descent and is attributed to inheritance of an autosomal-dominant mutation that prevents the maturational decline in lactase expression. Recent reports have identified genetic polymorphisms that are closely associated with lactase persistence and nonpersistence phenotypes. The identification of genetic variants associated with lactase persistence or nonpersistence allows for molecular detection of the genetic predisposition towards adult-onset hypolactasia by DNA sequencing or restriction fragment length polymorphism analysis. The role for such genetic detection in clinical practice seems limited to ruling out adult-onset hypolactasia as a cause of intolerance symptoms but remains to be fully defined. Attention should be paid to appropriate interpretation of genetic detection in order to avoid potentially harmful reduction in dairy intake or misdiagnosis of secondary lactase deficiency. PMID- 15287818 TI - The impact of structural genomics on the protein data bank. AB - The advent of structural genomics presents new challenges to the archive of biomacromolecular structures--the Protein Data Bank (PDB). As technologies involved in structure determination have advanced, both the number and size of structures available in the PDB have increased rapidly. The structural genomics initiatives are creating a large amount of data that needs to be tracked, archived, and made easily available. The PDB has developed tools to facilitate the rapid deposition of data produced by the structural genomics initiatives and has created databases to track the progress of the work. PMID- 15287819 TI - Genome-wide linkage disequilibrium and haplotype maps. AB - There is currently a broad effort to produce genome-wide high-density linkage disequilibrium (LD) maps with single nucleotide polymorphisms. The hope is that the resulting maps can be exploited to find genes that affect the onset and severity of at least some common human diseases. These maps may also be useful for identifying genes that affect drug response or the likelihood of drug toxicities. The goal of this review is to provide a broad overview of some of the key concerns motivating the design of a major international project called the International Haplotype Map Project. The process of map production requires the identification of very large numbers of polymorphic sites, implementation of facile, highly accurate and inexpensive genotyping production pipelines, and provision for public access to the genotype data. Great progress has been made recently in genotyping methods and these advances are allowing very large-scale data collection. A major goal of these efforts is to enable the selection of subsets of markers that capture useful genetic information in short genomic intervals, while optimally reducing the number of markers that must be genotyped. Standard measures of LD provide a starting point but may not fully capture the complexity of the information inherent in the data. Extremely dense genotype data in several broadly representative populations (European, Chinese, Japanese, and Yoruba) should yield important insights into the genetic structure of most genes. Further study is required to determine how broadly applicable the data will be to other population groups. Significant challenges lie ahead in determining the best methods for the selection of markers in disease/phenotype studies, large-scale genotyping, and analysis of the resulting genetic data. PMID- 15287820 TI - Technological advances in high-throughput screening. AB - High-throughput screening (HTS) is the process of testing a large number of diverse chemical structures against disease targets to identify 'hits'. Compared to traditional drug screening methods, HTS is characterized by its simplicity, rapidness, low cost, and high efficiency, taking the ligand-target interactions as the principle, as well as leading to a higher information harvest. As a multidisciplinary field, HTS involves an automated operation-platform, highly sensitive testing system, specific screening model (in vitro), an abundant components library, and a data acquisition and processing system. Various technologies, especially the novel technologies such as fluorescence, nuclear magnetic resonance, affinity chromatography, surface plasmon resonance, and DNA microarray, are now available, and the screening of more than 100,000 samples per day is already possible. Fluorescence-based assays include the scintillation proximity assay, time-resolved energy transfer, fluorescence anisotropy, fluorescence correlation spectroscopy, and fluorescence fluctuation spectroscopy. Fluorescence-based techniques are likely to be among the most important detection approaches used for HTS due to their high sensitivity and amenability to automation, giving the industry-wide drive to simplify, miniaturize, and speed up assays. The application of NMR technology to HTS is another recent trend in drug research. One advantage afforded by NMR technology is that it can provide direct information on the affinity of the screening compounds and the binding location of protein. The structure-activity relationship acquired from NMR analysis can sharpen the library design, which will be very important in furnishing HTS with well-defined drug candidates. Affinity chromatography used for library screening will provide the information on the fundamental processes of drug action, such as absorption, distribution, excretion, and receptor activation; also the eluting curve can give directly the possibility of candidate drug. SPR can measure the quantity of a complex formed between two molecules in real-time without the need for fluorescent or radioisotopic labels. SPR is capable of characterizing unmodified biopharmaceuticals, studying the interaction of drug candidates with macromolecular targets, and identifying binding partners during ligand fishing experiments. DNA microarrays can be used in HTS be used to further investigate the expression of biological targets associated with human disease, which then opens new and exciting opportunities for drug discovery. Without doubt, the addition of new technologies will further increase the application of HTS in drug screening and its related fields. PMID- 15287821 TI - Physiological roles of amyloid-beta and implications for its removal in Alzheimer's disease. AB - The underlying pathological cause of Alzheimer's disease has been postulated to be an excess of amyloid-beta (Abeta) which aggregates into toxic fibrillar deposits within the extracellular space of the brain, thereby disrupting neuronal and synaptic function and eventually leading to neuronal degeneration and dementia. As a result, therapeutic strategies have been developed that are designed to remove Abeta from the brain. Caution needs to be exercised concerning such strategies because, in addition to its presence in neuritic plaques, Abeta has a widespread distribution through the brain and body, even in cognitively normal individuals. Evidence indicates that instead of being a toxic peptide, soluble Abeta serves a variety of physiological functions, including modulation of synaptic function, facilitation of neuronal growth and survival, protection against oxidative stress, and surveillance against neuroactive compounds, toxins and pathogens. These physiological functions must be taken into account when strategies are developed to reduce Abeta load in Alzheimer's disease. Ideally, such strategies should target forms of Abeta that are not bioavailable, such as fibrillar Abeta, or forms that are regarded to be overexpressed in Alzheimer's disease (such as oligomers) while leaving normal soluble Abeta1-40 and Abeta1-42 intact. At present none of the available therapeutic strategies appears to have such selectivity. Until these technical limitations and the uncertainties regarding the effect of depletion of Abeta from the brain are resolved, it would not be prudent to begin further clinical trials. PMID- 15287822 TI - Inhaled corticosteroids, bone mineral density and fracture in older people. AB - The efficacy of inhaled corticosteroids in the treatment of asthma has been firmly established in a variety of settings. The majority of asthma management plans now recommend the use of inhaled corticosteroids at an early stage. This means that most patients with asthma will be prescribed an inhaled corticosteroid at some point in time and many patients with asthma will use these drugs for several years. Inhaled corticosteroids are also used in the treatment of other conditions, particularly chronic obstructive pulmonary disease (COPD). Since inhaled corticosteroids are absorbed into the systemic circulation, they can have systemic adverse effects, such as suppression of the hypothalamic-pituitary adrenal axis and increasing the risk of bruising. However, perhaps the greatest concern for patients is whether the regular use of inhaled corticosteroids has an adverse impact on the bone mineral density and increases the risk of fracture. There is now accumulating evidence from epidemiological studies that the use of inhaled corticosteroids is inversely related to bone mineral density in a dose dependent fashion. However, data from two clinical trials of moderately high doses of inhaled corticosteroids in patients with COPD have produced conflicting results and while the larger study of triamcinolone found a significant impact of this drug on bone mineral density, a smaller study of budesonide found no effect. Epidemiological research into the relationship between inhaled corticosteroids and fracture is at an early stage. To date, only three studies in this area have been reported, all of which have used different approaches to try to minimise the impact of bias and confounding. There is a lack of consistency between the final estimates of the impact of inhaled corticosteroids on fracture risk. However, taken together these data suggest that the short to medium term use of inhaled corticosteroids is associated with a small adverse effect on bone. Doctors and patients need to be aware of this risk and balance it against the known beneficial effects of inhaled corticosteroids. PMID- 15287823 TI - Poststroke epilepsy: epidemiology, pathophysiology and management. AB - Seizures and status epilepticus can be a presenting feature of acute stroke. They may occur in its early (<7 days) clinical course or be a remote (>7 days) complication. Most seizures are single, either partial or generalised. Early and remote seizures seem to have different predictors and pathogenesis. Seizures are more frequent in severe and disabling strokes, haemorrhagic strokes and those with cortical involvement. The risk of epilepsy is higher for patients with early seizures, cortical infarctions and lobar haemorrhages and in dependent patients. Early or remote seizures do not have a significant influence on dependency or mortality, although seizures and status epilepticus can be a direct cause of death. Treatment can be started after a first or a recurrent seizure. Treatment options include phenytoin, carbamazepine, valproic acid (valproate sodium) and the new antiepileptic drugs (AEDs). New AEDs can be used to decrease the likelihood of drug interactions and adverse effects in patients who do not tolerate the classic AEDs and in treatment failures with classic AEDs. Large observational studies to define prognostic factors for poststroke seizures in specific stroke subtypes are needed. Randomised controlled trials of AED prophylaxis for acute and remote seizures are essential to improve the evidence level of current guidelines and recommendations. PMID- 15287825 TI - Assessing the health and economic impact of galantamine treatment in patients with Alzheimer's disease in the health care systems of different countries. AB - INTRODUCTION: Cholinesterase inhibitors have been shown to improve cognitive function and improve or maintain global function. OBJECTIVE: To estimate the long term economic impact of treating patients with Alzheimer's disease with galantamine in seven healthcare systems: Australia, Canada, Finland, New Zealand, Sweden, the Netherlands and the UK. METHODS: The time until patients require full time care (FTC), defined as the consistent requirement for a significant amount of care giving and supervision each day, and the associated costs were evaluated using the 'Assessment of Health Economics in Alzheimer's Disease (AHEAD)' model. Efficacy data were obtained from three clinical trials comparing galantamine with placebo and local cost and resource use data were determined for each country. Forecast costs reported in Euros (2001 value), were made for up to 10 years in each healthcare system. All costs were determined from a perspective somewhat broader than that of a comprehensive payer, including social services. Both benefits and costs were discounted at 3%. RESULTS: Galantamine (16 mg/day) is predicted to delay the need for FTC by 6.8%, thus the cumulative cost of care over 10 years is expected to be reduced, and this offsets much or all of the cost of galantamine. Approximately five patients need to be treated to avoid 1 year of FTC. In each healthcare system, FTC was estimated to account for 61-92% of the cost. Savings were estimated for most of the countries. For those countries with an expected expense, there were reasonable costs per FTC month avoided (euro553, discounted) and costs per quality-adjusted life year gained (euro25,000). CONCLUSION: In addition to the clinical benefits associated with galantamine treatment, the savings predicted from delaying FTC may offset the treatment costs. PMID- 15287826 TI - The time to recommend antenatal HIV screening for all pregnant women has arrived. PMID- 15287824 TI - Sexual dysfunction in the older woman: an overview of the current understanding and management. AB - Sexuality is one of the most important quality of life issues for both men and women. Sexual dysfunction is a highly prevalent, age-related and progressive problem. The various physiological and psychological changes that occur with aging can have a significant impact on sexual function. The complexity of female sexual dysfunction remains distinct from that of a man. Thus, we cannot approach female patients or their sexual function problems in a similar fashion to that of male patients. A woman's motivation and ability to find and respond to sexual stimuli is largely influenced by her emotional intimacy with her partner. Frequently, the emotional and relationship well-being a woman experiences contributes more to her sexual enjoyment than does her physiological response. However, it is imperative to assess for possible physiological barriers a woman may have which impede a healthy and satisfying sexual life. Therefore, a comprehensive approach, addressing both the physiological and psychological factors is instrumental to the evaluation of female patients with sexual complaints. After years of ardent research and recent therapeutic advances in male sexual dysfunction, researchers have begun addressing the intricacy of female sexual complaints. Studies involving both pre- and postmenopausal women have reported that most women do experience some type of sexual dysfunction during their lifetime. The sexual complaints women experience in their younger years may follow them into older adulthood, but often times change considerably because of various age-related changes. In an effort to assist researchers and clinicians in designing studies and implementing appropriate evaluation and treatment options for women with sexual complaints, a classification system for female sexual dysfunction has been designed. The four categories of female dysfunction include: hypoactive sexual desire disorder, sexual arousal disorder, orgasmic disorder and sexual pain disorders. Evaluation of women with sexual complaints should include a detailed psychological, social and medical history and thorough physical examination including a hormonal profile. Current treatment options are dependent on the diagnosis and include physical therapy, psychological counselling, hormonal supplements, medication changes and sexual devices. There has also been a burgeoning interest in investigational medications for female sexual dysfunction, from centrally acting (e.g. serotonin agonists) to peripheral, localised treatment (e.g. vasodilating creams). The area of female sexuality and sexual dysfunction has been undergoing important critical changes within the last 10 years. Researchers and clinicians are continuing to recognise the need to try and understand both the psychological and physiological aspects of the female sexual experience and how they influence one another. PMID- 15287827 TI - Medical registry governance and patient privacy. PMID- 15287828 TI - UK health inequalities: the class system is alive and well. PMID- 15287829 TI - Severe traumatic brain injury in New South Wales: comparable outcomes for rural and urban residents. AB - OBJECTIVE: To compare differences in functional outcomes between urban and rural patients with traumatic brain injury (TBI). DESIGN: A longitudinal, prospective, multicentre study of a 2-year cohort from the Brain Injury Rehabilitation Program (BIRP) for New South Wales, with follow-up at 18 months after injury. PARTICIPANTS: 198 patients (147 urban, 51 rural) with severe TBI from the 11 participating rehabilitation units. MAIN OUTCOME MEASURES: Demographic and injury details collected prospectively using a standardised questionnaire, and measures from five validated instruments (Disability Rating Scale, Mayo-Portland Adaptability Inventory, Sydney Psychosocial Reintegration Scale, Medical Outcomes Study Short Form and the General Health Questionnaire--28-item version) administered at follow-up to document functional, psychosocial, emotional and vocational outcomes. RESULTS: Demographic details, injury severity, lengths of stay in intensive and acute care wards were similar for both rural and urban groups. There were no significant group differences in functional outcomes, including return to work, at follow-up. CONCLUSIONS: Our findings contrast with previous research that has reported poorer outcomes after TBI for rural residents, and suggest that the integrated network of inpatient, outpatient and outreach services provided throughout NSW through the BIRP provides effective rehabilitation for people with severe TBI regardless of where they live. PMID- 15287831 TI - Who are the kids who self-harm? An Australian self-report school survey. AB - OBJECTIVE: To determine the prevalence and types of deliberate self-harm (DSH) in adolescents, and associated factors. DESIGN: A cross-sectional questionnaire study. PARTICIPANTS AND SETTING: 3757 of 4097 Year 10 and Year 11 students (91.7%) from 14 high schools on the Gold Coast, Queensland, during September 2002. MAIN OUTCOME MEASURES: DSH behaviour, including descriptions of the last act, psychological symptoms, recent stressors, coping styles, help-seeking behaviour, lifestyle choices, and self-prescribing of medications. RESULTS: 233 students (6.2%) met the criteria for DSH in the previous 12 months, with DSH more prevalent in females than males (OR, 7.5; 95% CI, 5.1-10.9). The main methods were self-cutting (138 respondents; 59.2%) and overdosing with medication (69 respondents; 29.6%). Factors associated with DSH included similar behaviours in friends or family, coping by self-blame, and self-prescribing of medications. Most self-harmers did not seek help before or after their most recent action, with those who did primarily consulting friends. CONCLUSIONS: DSH is common in Australian youth, especially in females. Preventive programs should encourage young people to consult health professionals in stressful situations. PMID- 15287830 TI - Changing availability of neonatal intensive care for extremely low birthweight infants in Victoria over two decades. AB - OBJECTIVE: To determine the changes in availability of neonatal intensive care for extremely low birthweight (ELBW) infants, and the consequences of a lack of availability. DESIGN AND SETTING: Population-based cohort study of consecutive ELBW infants born in the state of Victoria during four distinct eras. PARTICIPANTS: All livebirths weighing 500-999 g in Victoria in the calendar years 1979-1980 (n = 351), 1985-1987 (n = 560), 1991-1992 (n = 429), and 1997 (n = 233). MAIN OUTCOME MEASURES: Changes over time in the proportions of ELBW infants offered intensive care, the proportions that were "outborn" (born outside level 3 perinatal centres), and their survival rates and quality of survival compared with "inborn" infants. RESULTS: The proportions of ELBW infants offered intensive care increased over time and were significantly higher in heavier infants. The proportion of outborn ELBW infants was 30% in 1979-1980, falling to 9% by 1997. The difference in survival rates between inborn and outborn infants widened progressively over time: the survival advantages for inborn infants over outborn infants were 12.0% in 1979-1980, 30.1% in 1985-1987, 36.5% in 1991-1992, and 43.6% in 1997. For survivors, the quality of life was significantly better for inborn infants in two of the four eras. CONCLUSIONS: Neonatal intensive care has been increasingly available for ELBW infants in Victoria over the period 1979 to 1997. The gap in survival rates between outborn and inborn infants has widened, and the quality of life of outborn survivors is inferior. PMID- 15287832 TI - Mobile phone interference with medical equipment and its clinical relevance: a systematic review. AB - OBJECTIVE: To conduct a systematic review of studies on clinically relevant digital mobile phone electromagnetic interference with medical equipment. DATA SOURCES: MEDLINE and SUMSEARCH were searched for the period 1966-2004. The Cochrane Library and Database of Abstracts of Reviews of Effects were also searched for systematic reviews. STUDY SELECTION: Studies were eligible if published in a peer-reviewed journal in English, and if they included testing of digital mobile phones for clinically relevant interference with medical equipment used to monitor or treat patients, but not implantable medical devices. DATA SYNTHESIS: As there was considerable heterogeneity in medical equipment studied and the conduct of testing, results were summarised rather than subjected to meta analysis. RESULTS: Clinically relevant electromagnetic interference (EMI) secondary to mobile phones potentially endangering patients occurred in 45 of 479 devices tested at 900 MHz and 14 of 457 devices tested at 1800 MHz. However, in the largest studies, the prevalence of clinically relevant EMI was low. Most clinically relevant EMI occurred when mobile phones were used within 1 m of medical equipment. CONCLUSIONS: Although testing was not standardised between studies and equipment tested was not identical, it is of concern that at least 4% of devices tested in any study were susceptible to clinically relevant EMI. All studies recommend some type of restriction of mobile phone use in hospitals, with use greater than 1 m from equipment and restrictions in clinical areas being the most common. PMID- 15287833 TI - Peripheral arterial disease: prognostic significance and prevention of atherothrombotic complications. AB - The prevalence of peripheral arterial disease (PAD) in people aged over 55 years is 10%-25% and increases with age; 70%-80% of affected individuals are asymptomatic; only a minority ever require revascularisation or amputation. Patients with PAD alone have the same relative risk of death from cardiovascular causes as those with coronary or cerebrovascular disease, and are four times more likely to die within 10 years than patients without the disease. The ankle brachial pressure index (ABPI) is a simple, non-invasive bedside tool for diagnosing PAD - an ABPI less than 0.9 is considered diagnostic of PAD. About half of patients with PAD (defined by an abnormal ABPI) have symptomatic coronary or cerebral vascular disease. The ABPI is an independent predictor of coronary and cerebrovascular morbidity and mortality. Patients with PAD require medical management to prevent future coronary and cerebral vascular events. There are currently insufficient data to recommend routine population screening for asymptomatic PAD using the ABPI. PMID- 15287834 TI - Ultraviolet radiation from welding and possible risk of skin and ocular malignancy. AB - Arc welding produces the full spectrum of ultraviolet radiation (UVR). It is possible that welders are at greater risk of developing skin cancer than the general population, but there is a dearth of well designed studies in this area. The only major study of the relationship between arc welding and skin cancer risk did not reveal an increased incidence of skin cancer in welders. As the welders examined were all well protected and the length-of-exposure period was limited, the findings cannot be generalised to all welders. Studies have demonstrated that welding increases the risk of ocular melanoma. Just as we urge the public to protect themselves from UVR, we need to consider similar advice for arc welders. PMID- 15287835 TI - Teaching on the run tips 4: teaching with patients. PMID- 15287836 TI - Adult chickenpox complicated by fatal necrotising pneumonia. PMID- 15287837 TI - Ethical and legal issues at the interface of complementary and conventional medicine. AB - Doctors should: Honestly answer patients' direct questions about CAM and elicit information about their use of it. Establish patients' understanding of the conventional and complementary therapies, both those available to them, and those that they may already be using. Establish why the patient uses CAM, and their goals for both complementary and conventional therapies. Reflect on whether information about CAM would be material for that patient at that time, taking into account the patient's burden of illness, his or her expressed preferences and the risks and benefits of both conventional and complementary therapy. Take steps to become adequately informed about available CAM that has consistently been shown to be safe and effective; has consistently been shown to be ineffective and/or harmful; or is consistently enquired about by patients. Become familiar with qualified and competent CAM practitioners (medical and non-medical) to whom referrals can be made when necessary. Continue a relationship with the patient, while continuing to monitor the patient conventionally and staying open to further discussions about CAM. PMID- 15287838 TI - Legal and ethical issues in complementary medicine: a United States perspective. AB - The way forward involves not only preventing negligence and fraud, but also facilitating therapeutic exchanges between various healthcare providers and their patients. PMID- 15287839 TI - Temporary protection visas and child refugees. PMID- 15287840 TI - Gouty arthritis in Australian Aboriginals: more common than previously suspected. PMID- 15287841 TI - Access block viewed as a medical model. PMID- 15287842 TI - Coronial autopsies: a rising tide of objections. PMID- 15287843 TI - Privacy: bad for your health? PMID- 15287844 TI - Metformin therapy and diabetes in pregnancy. PMID- 15287845 TI - Multisite, quality-improvement collaboration to optimise cardiac care in Queensland public hospitals. PMID- 15287846 TI - The upsurge of interest in Indigenous health in the 1950s and 1960s. Barry Christophers' letters to the MJA editor about Indigenous health. PMID- 15287847 TI - Vaccines: the new Australian best-practice schedule. PMID- 15287849 TI - Bacterial infections in cirrhosis. AB - Spontaneous bacterial peritonitis, urinary tract infections, respiratory infections and bacteremia are the most frequent infective complications in cirrhosis. These infections are due to the concomitant presence of different facilitating mechanisms including changes in the intestinal flora and in the intestinal barrier, depression of activity of the reticuloendothelial system, decreased opsonic activity of the ascitic fluid, neutrophil leukocyte dysfunction and iatrogenic factors among others. The fact, that the probability of having a microorganism responsible for the infection quinolone resistant is higher than 30% should be taken into account when treating any infection in a cirrhotic patient receiving selective intestinal decontamination with quinolones, and therefore, quinolones as empiric treatment are not indicated. PMID- 15287850 TI - Hepatic hydrothorax--pathophysiology, diagnosis and treatment--review of the literature. AB - Hepatic hydrothorax is defined as the accumulation of significant pleural effusion in a cirrhotic patient without primary pulmonary or cardiac disease. Hydrothorax is uncommon occurring in up to 4-6% of all patients with cirrhosis and up to 10% in patients with decompensated cirrhosis. Although ascites is usually present, hydrothorax can occur in the absence of ascites. Patients with hepatic hydrothorax usually have advanced liver disease with portal hypertension and most of them will require liver transplantation. Over the last few years, new insights into the pathogenesis of this entity have lead to improved treatment modalities such as portosystemic shunts (TIPS) and video-assisted thoracoscopy for closure of diaphragmatic defects. These modalities may be of help as a bridge to transplantation. The aim of this review is to describe recent developments in the pathogenesis, diagnosis and treatment of hepatic hydrothorax. PMID- 15287848 TI - Kinetic and structural optimization to catalysis at low temperatures in a psychrophilic cellulase from the Antarctic bacterium Pseudoalteromonas haloplanktis. AB - The cold-adapted cellulase CelG has been purified from the culture supernatant of the Antarctic bacterium Pseudoalteromonas haloplanktis and the gene coding for this enzyme has been cloned, sequenced and expressed in Escherichia coli. This cellulase is composed of three structurally and functionally distinct regions: an N-terminal catalytic domain belonging to glycosidase family 5 and a C-terminal cellulose-binding domain belonging to carbohydrate-binding module family 5. The linker of 107 residues connecting both domains is one of the longest found in cellulases, and optimizes substrate accessibility to the catalytic domain by drastically increasing the surface of cellulose available to a bound enzyme molecule. The psychrophilic enzyme is closely related to the cellulase Cel5 from Erwinia chrysanthemi. Both kcat and kcat/K(m) values at 4 degrees C for the psychrophilic cellulase are similar to the values for Cel5 at 30-35 degrees C, suggesting temperature adaptation of the kinetic parameters. The thermodynamic parameters of activation of CelG suggest a heat-labile, relatively disordered active site with low substrate affinity, in agreement with the experimental data. The structure of CelG has been constructed by homology modelling with a molecule of cellotetraose docked into the active site. No structural alteration related to cold-activity can be found in the catalytic cleft, whereas several structural factors in the overall structure can explain the weak thermal stability, suggesting that the loss of stability provides the required active-site mobility at low temperatures. PMID- 15287851 TI - Common heterozygous hemochromatosis gene mutations are risk factors for inflammation and fibrosis in chronic hepatitis C. AB - BACKGROUND: Chronic hepatitis C is frequently associated with increased hepatic iron stores. It remains controversial whether heterozygous mutations of hemochromatosis genes affect fibrosis progression. Therefore our aim was to assess associations between HFE mutations and hepatic inflammation and stage of fibrosis in German hepatitis C patients. METHODS: Liver biopsies from 166 patients were scored for inflammatory activity (A0-4) and hepatic fibrosis (F0 4). Gene mutations were determined by LightCycler, restriction fragment length polymorphism analysis, or direct sequencing. RESULTS: The frequencies of common HFE mutations C282Y and H63D are 4.2% and 21.3%, whereas the recently described S65C substitution and the Y250X mutation in the transferrin receptor 2 gene are very rare. In regression analysis, heterozygous carriers of C282Y or H63D mutations display significantly (P < 0.05) higher inflammatory activities and more advanced fibrosis than patients without mutations. For C282Y heterozygous patients, the odds ratios for marked inflammatory activity (A2-4) and advanced liver fibrosis or cirrhosis (F2-4) are 4.9 and 4.6, respectively, compared with patients carrying homozygous wild-type alleles. C282Y mutations are associated with significantly (P < 0.05) increased serum iron and aminotransferase levels, whereas H63D heterozygotes display higher transferrin saturation, serum iron, and ferritin concentrations compared to wild-type (P < 0.01). CONCLUSIONS: Common heterozygous hemochromatosis mutations are associated with higher grades of inflammation and more severe hepatic fibrosis. Our findings support a role of HFE mutations as primary risk factors for fibrogenesis and disease progression in chronic hepatitis C. PMID- 15287852 TI - Induction of oral immune regulation towards liver-extracted proteins for treatment of chronic HBV and HCV hepatitis: results of a phase I clinical trial. AB - BACKGROUND: Anti-viral immunity can be modulated via oral feeding of viral proteins. Hepatitis B and C viral (HBV, HCV)-associated hepatocellular injury is mediated by a defective host anti-viral immune response. AIMS: To determine the effect of oral administration of a mixture of liver-extracted proteins with HBV/HCV proteins, on viral load, liver injury, and the anti-viral T-cell response of chronic HBV/HCV patients. METHODS: Fourteen patients with chronic HBV and 15 patients with chronic HCV were treated orally with hepatocyte-extracted proteins and HBV or HCV viral proteins for 24 weeks, and followed for an additional 26 weeks. Patients were monitored for HBV-DNA or HCV-RNA levels, liver enzymes and liver histology. Viral-directed T-cell immunity was assessed by IFNgamma and IL10 ELISPOT, viral-specific T-cell proliferation, cytotoxicity, and cytokines assays, and followed for peripheral natural killer T-cell (NKT) number. RESULTS: In both chronic HBV and HCV patients, oral administration of a mixture of selected liver extracted proteins and viral proteins induced a favorable increase in viral specific T-cell proliferation, and IFNgamma-secreting clones, along with a significant decrease in the anti-viral IL10-secreting T-cell clones. However, the effects of modulation of the anti-viral immunity differed between the HBV and HCV patients. In both groups, no major adverse events were noted. In chronic HBV patients, a significant decrease in viral load was observed in 5/14 (35.7%) of patients. HB surface antigen/HB nucleocapsid antigen scores on liver biopsy improved in 46.1% and 50%, respectively, and the histological necroinflammatory score improved in 4/13 (30.7%). Forty percent of the patients with elevated liver enzymes showed a favorable biochemical response. In contrast, an improvement in the histological necroinflammatory score was observed in only 2/12 (17%) of the chronic HCV patients. No significant decrease in HCV RNA was noted in any of these patients. CONCLUSIONS: Immune regulation of the anti-HBV/HCV immune response via oral administration of a mixture of liver-extracted and viral proteins significantly altered the viral-specific immunity. This effect was associated with clinical and virological improvements in chronic HBV patients. PMID- 15287853 TI - Hepatitis B virus-specific T cell response in chronic hepatitis B patients treated with lamivudine and interferon-alpha. AB - AIMS: The goal of the present study was to assess the impact combination antiviral therapy has on immune responses in chronic hepatitis B. MATERIALS AND METHODS: T cell responses were studied in 16 chronically hepatitis B virus (HBV) infected patients treated with sequential, partially overlapping, lamivudine interferon (IFN)-alpha combination therapy. RESULTS: HBcAg-specific lymphoproliferative response (LPR) was transiently detected in four of five patients who achieved virus suppression (HBV DNA < 10(4) genome equivalents/ml) at end of dual therapy, and then reverted to pre-treatment viral load after therapy discontinuation. In contrast, no significant HBcAg-specific LPR was detected in 8 patients who did not attain profound HBV suppression, as well as in three patients who experienced no HBV DNA rebound after therapy discontinuation. CONCLUSIONS: This pilot study suggests that restored viral replication after pharmacological suppression drives the immune response to HBV in chronically infected patients. Further characterization of the adaptive immunity and its regulatory mechanisms at time of therapy discontinuation appears therefore necessary in controlled trials. PMID- 15287854 TI - The role of heat shock protein 27 expression in hepatocellular carcinoma in Japan: special reference to the difference between hepatitis B and C. AB - BACKGROUND: A recent report showed that heat shock protein (HSP)-27 expression was related to histological grade and survival of patients with hepatocellular carcinoma (HCC). AIMS: The aim of this study was to examine the effect of expression of HSP-27 on clinicopathological variables in Japanese patients with HCC. METHODS: An immunohistochemical study for HSP-27 was performed on 60 HCC cases using a monoclonal anti-HSP-27 antibody. We divided 60 patients into two groups, patients with a low expression of HSP-27 (n = 34) and those with a high expression of HSP-27 (n = 26). Forty patients tested positive for the hepatitis C virus (HCV) antibody and 20 tested positive for the hepatitis B surface antigen. RESULTS: There appeared to be no relationship between HSP expression and clinicopathologic factors and no differences were observed between the high expression group and the low expression group. In the hepatitis B virus (HBV) group (n = 20), HSP-27 expression correlated significantly with prognosis, disease-free survival (DFS) and overall survival. High expression was significantly associated with poor prognosis in the HBV group. In contrast, patients with a high expression tended to have a good prognosis in the HCV group (n = 40): DFS and overall survival. CONCLUSIONS: This study showed the possibility that HSP-27 plays different roles in HBV- and HCV-associated HCCs. PMID- 15287855 TI - Diagnostic and therapeutic implications of bile duct injury in autoimmune hepatitis. AB - BACKGROUND: Bile duct injury is not a feature of classical autoimmune hepatitis (AIH), but it has been described in variant forms of the disease. AIMS: Our goals were to assess the similarity of AIH with bile duct injury to classical disease and to evaluate the possibility of concurrent primary biliary cirrhosis (PBC). METHODS: Fifteen patients with bile duct injury were compared with 151 patients with classical AIH. Patterns of nuclear immunofluorescence and the frequency and nature of autoantibodies associated with AIH and PBC were determined. RESULTS: Patients with bile duct injury had the same nuclear-staining patterns, frequency and nature of autoantibodies, and genetic risk factors as the comparison group. Features specific for PBC, including the multiple nuclear dot pattern of immunofluorescence and antibodies to the M2 antigens, Sp100 and nuclear pore complex antigen, gp210, did not distinguish them from classical disease. Remission and treatment failure occurred with similar frequencies in both groups. CONCLUSIONS: Patients with AIH and bile duct injury lack features of PBC, and they respond as well to corticosteroid therapy as patients with classical disease. Background bile duct changes should not alter the diagnosis or treatment of AIH. PMID- 15287856 TI - Serum levels of soluble molecules associated with evasion of immune surveillance: a study in biliary disease. AB - BACKGROUND: Biliary carcinoma cells produce the transmembrane proteins, Fas, FasL, and RCAS1. It has been demonstrated that the Fas/FasL and RCAS1 systems induce apoptosis of activated immune cells and that the soluble isoforms of these proteins (sFas, sFasL, and sRCAS1) also exhibit this function. METHODS: We measured serum levels of these soluble-types in patients with biliary disease by ELISA and investigated their clinical significance. RESULTS: In some cases of cholangitis and autoimmune biliary disease, serum sFasL values were over 0.1 ng/ml but the protein was undetectable in any patients with biliary carcinoma. sFas levels were significantly higher in the autoimmune disease (mean, 6.83 ng/ml) and cancer (mean, 6.42 ng/ml) groups than in the cholangitis group (mean, 4.23 ng/ml) and normal controls (mean, 2.93 ng/ml). However, the sFas values in malignancy did not correlate with the progression of clinical stage. The percentage positive for serum sRCAS1 was 9.7% in benign disease but was 63.4% in cancer. CONCLUSIONS: Our data suggest that serum sFasL in biliary disease may be derived predominantly from activated immune cells and not from cancer cells and that autoimmune biliary disease may be mediated by the Fas/FasL apoptotic system. sRCAS1 is highly tumor-specific and may be of value in the diagnosis of malignancy. PMID- 15287857 TI - Hepatocyte growth factor and c-Met expression in rat and human liver fibrosis. AB - BACKGROUND: Hepatocyte growth factor (HGF) is a potent mitogen for hepatocytes in vitro. AIMS: Substitution of HGF was suggested for human liver disease on the basis of animal experiments. The cellular sources of HGF and its receptor, c-Met, in liver disease in vivo are not well defined. METHODS: We characterised HGF and c-Met expression in normal and cirrhotic human livers and rat livers at various time points after CCl4 administration by in situ hybridisation and immunohistology. HGF transcripts were restricted to resting and activated stellate cells in rat and human liver. RESULTS: In rat liver, HGF showed peak levels 6-12 h following acute intoxication, and remained increased after repeated CCl4 injury. HGF transcript levels were very low in normal human liver, but excessively raised in fibrosis/cirrhosis. In contrast, HGF immunoreactivity was found not only in perisinusoidal/periductular cells but also in cholangiocytes of proliferating ductules. c-Met RNA and protein was expressed in hepatocytes, cholangiocytes, and arteriolar endothelial cells. CONCLUSIONS: The HGF-specific immunostaining of proliferating cholangioles in the absence of HGF RNA suggests c Met-mediated uptake of HGF and paracrine stimulation of cholangiocellular proliferation. Mitogenic effects of HGF on hepatocytes may therefore be accompanied by undesired cholangiogenesis and angiogenesis limiting its therapeutic value in chronic liver disease. PMID- 15287858 TI - Nitric oxide synthase 1 is partly compensating for nitric oxide synthase 3 deficiency in nitric oxide synthase 3 knock-out mice and is elevated in murine and human cirrhosis. AB - BACKGROUND: The role of endothelial nitric oxide synthase 3 (NOS-3) in the hyperdynamic circulation associated with cirrhosis is established but not that of the neuronal (NOS-1) isoform. We therefore investigated aortic NOS-1 levels in NOS-3 knock-out (KO) and wildtype (WT) mice and in hepatic arteries of patients. METHODS: Mice rendered cirrhotic by bile duct ligation (BDL) were compared with sham-operated controls. Hepatic arteries of cirrhotic patients were collected during liver transplantation; donor vessels served as controls. mRNA levels were quantified by real-time PCR, protein levels by Western blotting and NO production by Nomega-nitro-L-arginine methyl ester inhibitable arginine-citrulline assay. RESULTS: Aortae of NOS-3 KO mice exhibited higher NOS-1mRNA (5.6-fold, P < 0.004) and protein levels (8.8-fold) compared with WT. NO production in aortae of NOS-3 KO mice was 52% compared with WT (P = 0.002). BDL increased NOS-1 mRNA (2.4-fold, P = 0.01) and protein (7.1-fold) levels in aortae of WT, but no further in the NOS-3 KO mice. Hepatic artery NOS-1 mRNA levels in cirrhotic patients were markedly increased compared with controls (24.5-fold, P = 0.0007). CONCLUSIONS: Increased NOS-1 mRNA and protein levels and partially maintained in vitro NO production in aortae of NOS-3 KO mice suggest that NOS-1 may partially compensate for NOS-3 deficiency. BDL-induced increase in aortic NOS-1 mRNA and protein levels hint that not only NOS-3, but also NOS-1 may be involved in the regulation of systemic hyperdynamic circulation and portal hypertension. Upregulation of NOS 1 mRNA levels in hepatic arteries of portal hypertensive patients suggests possible clinical significance for these experimental findings. PMID- 15287859 TI - Characteristics of murine histidinaemia and its potential for genetic manipulation. AB - BACKGROUND: Histidinaemia is an autosomal recessive disorder affecting the hepatic enzyme histidine ammonia lyase (histidase) resulting in elevated plasma and urinary histidine and is prototypic of a series of hepatic cytosolic enzyme defects. AIMS: To characterise the physiology of murine histidinaemia with respect to histidine excretion and catabolism, and explore the potential for manipulating cellular and whole body histidase metabolism by gene transfer. MATERIALS AND METHODS: We studied his/his mice which have a G to A substitution in the gene encoding histidase, using both in vitro transduction of isolated hepatocytes by lipofection with wild-type histidase cDNA, and in vivo transduction of whole liver using a retroviral construct. RESULTS AND CONCLUSION: Histidase cDNA expression restored histidase activity in vivo and in vitro towards normal levels, demonstrated both at the cellular level and by whole body metabolic studies, establishing the potential of this model for the development of new gene therapeutic approaches. PMID- 15287860 TI - Analysis of the functional integrity of cryopreserved human liver cells including xenografting in immunodeficient mice to address suitability for clinical applications. AB - BACKGROUND: The availability of well-characterized human liver cell populations that can be frozen and thawed will be critical for cell therapy. We addressed whether human hepatocytes can recover after cryopreservation and engraft in immunodeficient mice. METHODS: We isolated cells from discarded human livers and studied the properties of cryopreserved cells. The viability of thawed cells was established with multiple in vitro assays, including analysis of liver gene expression, ureagenesis, cytochrome P450 activity, and growth factor-induced cell proliferation. The fate of transplanted cells was analysed in immunodeficient NOD SCID mice. RESULTS: After thawing, the viability of human hepatocytes exceeded 60%. Cells attached to culture dishes, proliferated following growth factor stimulation and exhibited liver-specific functions. After transplantation in NOD SCID mice, cells engrafted in the peritoneal cavity, a heterologous site, as well as the liver itself, retained hepatic function and proliferated in response to liver injury. Transplanted hepatocytes were integrated in the liver parenchyma. Occasionally, transplanted cells were integrated in bile ducts. CONCLUSIONS: Cryopreserved human liver cell showed the ability to retain functional integrity and to reconstitute both hepatic and biliary lineages in mice. These studies offer suitable paradigms aimed at characterizing liver cells prior to transplantation in people. PMID- 15287861 TI - Tissue collection, transport and isolation procedures required to optimize human hepatocyte isolation from waste liver surgical resections. A multilaboratory study. AB - BACKGROUND: The European Center for Validation of Alternative Methods (ECVAM) has funded a prevalidation study in three laboratories (France, USA and UK) on the use of human hepatocyte cultures to predict cytochrome P-450 induction. AIMS AND METHODS: As first stage of this prevalidation study, the purpose of the present work was to set criteria for optimization and harmonization of hepatocyte isolation from human tissue among laboratories to establish a routine procedure. This was achieved by combining and/or comparing the data generated by the two independent European laboratories (France and UK). RESULTS: The results confirmed that surgical waste material is a valuable source for obtaining high quality hepatocytes under certain pre-, intra- and post-operative conditions: cell yield of viable hepatocytes was not significantly affected by age and sex of patients, nor indications for resection, steatosis or cholestasis. Cold ischeamia up to 5 hours did not influence viable cell yield allowing transport of material. CONCLUSION: The use of biopsy sizes between 50-100 g, cannulation with 2-4 cannulae, digestion with collagenase-containing digestion medium at a flow rate of 25 ml/cannula for 20 minutes, with cut surface being glued in order to reform Glisson's capsule, should optimize the total yield of viable human hepatocytes obtained per preparation of waste liver surgical resections. PMID- 15287862 TI - Increased levels of tissue inhibitor of metalloproteinase-1 in human hepatocellular carcinoma. AB - BACKGROUND: Invasion and metastasis of hepatocellular carcinoma (HCC) are regulated by matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs). However, the role of TIMPs in these processes is not clear. AIM: To examine the potential involvement of TIMP-1 in HCC and the association between TIMP-1 and clinical outcome of patients with HCC. METHODS: The study included 91 patients who underwent surgical removal of HCC. TIMP-1 concentrations in the supernatant of tissue homogenates of HCC and non-neoplastic liver were measured by enzyme immunoassay. The relationships between TIMP-1 concentration and various clinicopathological features and recurrence of HCC after surgical operation were examined. RESULTS: The mean level of TIMP-1 in HCC (486 +/- 610 ng/mg protein, +/- SD) was significantly higher than in the non neoplastic liver (75 +/- 69, P < 0.0001). The median level of TIMP-1 in poorly differentiated HCCs (701 ng/mg protein) was significantly higher than in well- (80) and moderately (172) differentiated HCCs (P = 0.0047 and P = 0.0082, respectively). TIMP-1 level in liver cirrhosis was higher than in chronic hepatitis (P = 0.0015). TIMP-1 levels in HCC did not influence the recurrence rate of HCC. CONCLUSIONS: TIMP-1 concentration in HCC was higher than in non neoplastic liver and correlated with the differentiation grade of HCCs. However, tissue TIMP-1 concentration does not seem to be an important determinant of HCC recurrence. PMID- 15287863 TI - Inhibition of tumor necrosis factor-induced apoptosis in transgenic mouse liver expressing creatine kinase. AB - BACKGROUND: The mitochondrion acts as a pivotal decision center in many types of apoptotic responses. To clarify the effects of the enhanced mitochondrial function on tumor necrosis factoralpha (TNFalpha)-induced apoptosis, we studied hepatic injuries in transgenic mice whose livers express creatine kinase (CK). METHODS: Mice fed a diet containing 10% creatine, came to accumulate phosphocreatine and to enhance hepatic ATP levels and mitochondrial oxidative phosphorylation activities. TNFalpha-mediated hepatic apoptosis in normally fed and Cr-feeding CK transgenic mice were assessed. RESULTS: TNFalpha and actinomycin D cause severe liver failure in normally fed transgenic mice, and in the wild-type mice. In contrast, no significant elevations in transaminase levels after injection were observed in Cr feeding transgenic mice. The disruption of the mitochondrial transmembrane potential at 2 h after TNFalpha injection, prior to ATP depletion, activation of caspase 3 like protease, and DNA fragmentation at 4-6 h after injection, were observed in normally fed transgenic mice. These were fully suppressed in Cr feeding transgenic mice. However, anti-Fas antibody induced apoptosis was not inhibited in both groups. CONCLUSIONS: The results indicate that TNFalpha-induced apoptosis was inhibited in CK transgenic mice livers by maintaining mitochondrial function. PMID- 15287865 TI - Detection of Helicobacter pylori in water by direct PCR. AB - AIMS: This paper demonstrates a rapid, simple method for the detection of Helicobacter pylori in water that eliminates the need for recovery of cells or DNA extraction prior to PCR. METHODS AND RESULTS: Direct polymerase chain reaction (DPCR) with primers specific for H. pylori ureA (urease, subunit A) were used to detect H. pylori added to groundwater. DPCR also detected H. pylori in a naturally contaminated water sample. CONCLUSIONS: DPCR should provide an improved method to assess contamination of water by H. pylori. SIGNIFICANCE AND IMPACT OF THE STUDY: This simple, rapid method for detection of H. pylori in water will provide an improved means to investigate the possible role of water as a disease vector. PMID- 15287866 TI - In vitro new dialysis protocol to assay the antiseptic properties of a quaternary ammonium compound polymerized with denture acrylic resin. AB - AIMS: To develop an in vitro protocol in order to assess the antiseptic properties of a quaternary ammonium compound polymerized with acrylic denture resin base, using experimental resin discs and dialysis membranes. METHODS AND RESULTS: Experimental acrylic resin discs were polymerized with Poly 202063A, an ammonium compound (2-50%). Antiseptic properties were assayed against two reference strains (Escherichia coli, Staphylococcus aureus) and a laboratory strain (Candida albicans), using three different conditions (test A, B and C). In test A, according to classical protocols the resin discs were first soaked in large volumes of microbial inoculum (45 ml). An original dialysis protocol was then designed to recreate the small biofilm volume on the prosthetic surface. In test B, discs and bacterial inoculum (600 microl) were introduced in a dialysis bag and dialysed against a sterile buffer. A bactericidal effect was observed against E. coli and Staph. aureus (<0.1% viable cells in initial bacterial suspension). A dose-dependent fungistatic effect was observed against C. albicans. Finally, in test C discs and sterile buffer (600 microl) were introduced in a dialysis bag and dialysed against microbial inoculum. Reduced activity was found outside the dialysis bag, demonstrating that free ammonium was able to diffuse through the dialysis membrane, displaying antiseptic properties. CONCLUSIONS: The present protocol demonstrated that a quaternary ammonium compound remains efficient after heat polymerization with an acrylic denture base resin, both in immediate and distant microbial environments. SIGNIFICANCE AND IMPACT OF THE STUDY: Such removable prosthetic devices with intrinsic antiseptic properties would contribute to improve the long-term management of denture stomatitis. PMID- 15287867 TI - Genetic diversity of Neisseria meningitidis strains isolated in Rio de Janeiro, Brazil, evaluated by multilocus enzyme electrophoresis. AB - AIMS: To analyse Neisseria meningitidis isolates from meningococcal meningitis cases in Rio de Janeiro (Brazil) from 1990 to 1993 and 1999-2002, to determine the genetic and relatedness with hypervirulent and epidemic strains. METHODS AND RESULTS: The isolates were analysed by multilocus enzyme electrophoresis (MEE) clustering into 83 electrophoretic types (ET). All isolates from 1999 to 2002, formed a cluster which included one strain of the ET-5 complex worldwide associated with epidemics. CONCLUSIONS: The overall results suggested a panmictic structure probably because of recombination events. The observation of a separated cluster including isolates from 1999 to 2002 and an ET-5 complex strain, also suggested the introduction of strains genetically related with this hypervirulent complex in the State of Rio de Janeiro (Brazil) over the last 5 years. SIGNIFICANCE AND IMPACT OF THE STUDY: The presence of strains related to the ET-5 complex in several states of Brazil was already described elsewhere, but this is the first time it was reported in the State of Rio de Janeiro. Our findings reinforce the necessity to genetically determine the clones which should be considered to produce a national vaccine against meningococcal meningitis. PMID- 15287868 TI - Comparative analysis of conserved genetic markers and adjacent DNA regions identified in beer-spoilage lactic acid bacteria. AB - AIMS: To conduct an inter-species comparative study on the nucleotide sequences of the conserved DNA regions surrounding ORF5, a genetic marker for differentiating beer-spoilage lactic acid bacteria. METHODS AND RESULTS: The conserved DNA regions surrounding ORF5 were examined by PCR analysis, using three beer-spoilage strains, Lactobacillus brevis ABBC45C, L. paracollinoides LA2T and Pediococcus damnosus ABBC478. As a result, the DNA regions containing ORF1-7, originally found in ABBC45C, appeared to be conserved among the three strains, while the downstream region was not found in L. paracollinoides LA2T and P. damnosus ABBC478. The sequencing analysis of the conserved DNA regions of LA2T and ABBC478 revealed ca 99% nucleotide sequence identities with that of ABBC45C. CONCLUSIONS: The nucleotide sequences of the ca 8.2 kb DNA regions containing ORF1-7 were virtually identical among the three strains belonging to different species. The internal organizations of the ORFs were found to be remarkably similar. SIGNIFICANCE AND IMPACT OF THE STUDY: The level of nucleotide sequence identities suggests the DNA regions surrounding ORF5 were horizontally acquired by these beer-spoilage strains belonging to the three different species of lactic acid bacteria. PMID- 15287869 TI - Use of a fluorescent viability stain to assess lethal and sublethal injury in food-borne bacteria exposed to high-intensity pulsed electric fields. AB - AIMS: To apply scanning electron microscopy, image analysis and a fluorescent viability stain to assess lethal and sublethal in food-borne bacteria exposed to high-intensity pulsed electric fields (PEF). METHODS AND RESULTS: A rapid cellular staining method using the fluorescent redox probes 5-cyano-2,3-ditolyl tetrazolium chloride (CTC) and 4',6-diamidino-2-phylindole was used for enumerating actively respiring cells of Listeria mononcytogenes, Bacillus cereus and Escherichia coli. This respiratory staining (RS) approach provided good agreement with the conventional plate count agar method for enumerating untreated and high-intensity PEF-treated bacteria suspended in 0.1% (w/v) peptone water. However, test organisms subjected to similar levels of lethality by heating at 56 degrees C resulted in ca 3-log-unit difference in surviving cell numbers ml(-1) when enumerated by these different viability indicators. PEF-treated bacteria were markedly altered at the cellular level when examined by scanning electron microscopy. CONCLUSIONS: While PEF-treatment did not produce sublethally injured cells (P < 0.05), substantial subpopulations of test bacteria rendered incapable of forming colonies by heating may remain metabolically active. SIGNIFICANCE AND IMPACT OF THE STUDY: The fluorescent staining method offers interesting perspectives on assessing established and novel microbial inactivation methods. Use of this approach may also provide a better understanding of the mechanisms involved in microbial inactivation induced by PEF. PMID- 15287870 TI - Some potential sources for transmission of Campylobacter jejuni to broiler chickens. AB - AIMS: The aim of the study was to determine Campylobacter jejuni contamination and prevalence on fomites moving between broiler farms and the processing plant in the period after cleaning and before departure to harvest chickens. In addition, changes in the proportion of contaminated fomites in the course of a day were assessed. METHODS AND RESULTS: Pooled swab samples were obtained from pallets, crates, wheels of trucks, tractors and forklifts, truck beds, and from drivers' and catchers' boots. After enrichment in Bolton's broth Campylobacter were recovered on modified blood-free Campylobacter selective agar (mCCDA). Isolates were identified using tests for phenotypic and biochemical characteristics. Of the 209 samples collected, 53% were positive for C. jejuni, with all fomites positive except tractor wheels. Pallets had the highest contamination rate at 75%. More than 50% of catchers' boots, drivers' boots, crates and truck wheels were positive. Forty-seven per cent and 31% of truck beds and forklift wheels, respectively, were contaminated. The proportion of contaminated fomites did not change significantly during the day. CONCLUSIONS: This study has identified trucks, forklifts, pallets, crates, drivers' and catchers' boots as potential sources of C. jejuni for broilers. SIGNIFICANCE AND IMPACT OF THE STUDY: Campylobacter jejuni contamination of broiler processing plant fomites was found to be extensive ranging from 31% for truck beds to 75% for pallets. The proportion of contaminated fomites was observed to be similar throughout the day. The impact of contaminated fomites as sources of colonization of broilers with C. jejuni is discussed. PMID- 15287871 TI - Phylogenetic analysis of methyl coenzyme-M reductase detected from the bovine rumen. AB - AIMS: The object of the present study is isolation of methyl coenzyme-M reductase (MCR) genes (mcrA) from the bovine rumen fluid and determination of phylogenetical placements of the genes to investigate mechanisms of methanogenesis in the rumen from a point of view of mcrA genes. METHODS: Genes for methanogen-specific MCR were isolated from the bovine rumen by PCR amplification. The deduced amino acid sequences were fitted to the alignments of mcrA gene products from the referred sequences. SIGNIFICANCE AND IMPACT OF THE STUDY: Although the deduced amino acid sequences of mcrA genes, isolated from the bovine rumen in the present study, were close to that of Methanobrevibacter ruminantium, these amino acid sequences did not fall into known clusters of MCR. The findings suggest that methanogenesis in the rumen would be partially carried out by unknown methanogens. PMID- 15287872 TI - The pattern and kinetics of substrate metabolism of Campylobacter jejuni and Campylobacter coli. AB - AIMS: The main aim was to investigate the patterns and kinetics of substrate oxidation by Campylobacter jejuni and C. coli. METHODS AND RESULTS: Substrate oxidation profiles by 100 strains were determined using oxygen electrode system. All the isolates tested oxidized formate, l-lactate, cysteine, glutamine and serine with high oxidation rates and high affinity but varied in their ability to oxidize citric acid cycle intermediates, aspartic acid and serine. CONCLUSIONS: Based on the oxidation ability of alpha-ketoglutarate, succinate, fumarate and aspartic acid, Campylobacter strains tested were divided into three distinct metabolic categories. The first group was able to metabolize alpha-ketoglutarate, succinate, fumarate and aspartic acid; the second group was unable to oxidize alpha-ketoglutarate; and the third group was unable to oxidize, succinate, fumarate, and aspartic acid. Furthermore, serine oxidation rate enabled the differentiation of C. jejuni and C. coli. SIGNIFICANCE AND IMPACT OF THE STUDY: Overall, the results highlights the extensive metabolic diversity between and within Campylobacter species. In addition, the kinetic data of oxidized substrates obtained may improve the isolation procedures of the organism. PMID- 15287873 TI - Development of a 16S rDNA-targeted PCR assay for monitoring of Lactobacillus plantarum and Lact. rhamnosus during co-cultivation for production of inoculants for silages. AB - AIMS: This paper reports a simple, rapid approach for the parallel detection of Lactobacillus plantarum and Lact. rhamnosus in co-culture in order to produce an inoculant mixture for silage purposes. METHODS AND RESULTS: The 16S rDNA-targeted PCR primers were established for parallel detection of Lact. plantarum and Lact. rhamnosus in a single multiplex PCR. A protocol for application of these primers in direct PCR as well as colony-direct (CD) PCR was developed. These primers were also applicable for the estimation of the relative amount of each DNA type in mixed probes (semi-quantitative PCR). CONCLUSIONS: The PCR assay presented in this study is a robust, fast and semi-quantitative approach for detection of Lact. plantarum and Lact. rhamnosus in liquid cultures as well as on agar plates. SIGNIFICANCE AND IMPACT OF THE STUDY: This method provides an effective tool for the establishment of a regime for co-cultivation of Lact. plantarum and Lact. rhamnosus. This would enable faster and thus cost-reduced production of ensiling inoculants. PMID- 15287874 TI - Enhanced resolution of random amplified polymorphic DNA genotyping of Pseudomonas aeruginosa. AB - AIMS: To develop a rapid, sensitive and reproducible screening test for the detection of nosocomial spreading of Pseudomonas aeruginosa. METHODS AND RESULTS: Ps. aeruginosa genomic DNA extraction, RAPD-PCR, electrophoresis on acrylamide gel and silver staining were performed by using standardized reagents and conditions. The results were compared with the agarose gel electrophoresis followed by ethidium bromide staining. CONCLUSIONS: The coupling of acrylamide gel electrophoresis and silver staining gave about 80% more DNA bands than the traditional method, allowing a finer discrimination among different Ps. aeruginosa strains. SIGNIFICANCE AND IMPACT OF THE STUDY: By enhancing the resolution of the electrophoretic separation and the sensitivity of the staining, random amplification could be easily applied to the surveillance and prevention of nosocomial infections by clinical microbiology laboratories. PMID- 15287876 TI - Barotolerance is inducible by preincubation under hydrostatic pressure, cold-, osmotic- and acid-stress conditions in Lactobacillus sanfranciscensis DSM 20451T. AB - AIMS: This study addresses the inducibility of barotolerance by preincubation of Lactobacillus sanfranciscensis DSM 20451T under various sublethal stress conditions. METHODS AND RESULTS: Stress conditions which reduce the growth rate of L. sanfranciscensis DSM 20451T to 10% of its maximum were determined. These conditions were met at 43, 12.5 degrees C, a pH value of 3.7, 1.9% NaCl, or 80 MPa respectively. In contrast to heat preincubation, other prestresses, including salt, cold and pressure led to an increase of barotolerance by hydrostatic pressure of 300 MPa for 30 min. Stationary-phase cells also showed an increased barotolerance. Sublethal pressure leads to enhanced heat tolerance. CONCLUSIONS: Stress response to salt, low temperature and acidic pH as well as starvation overlap with that one to high pressure by inducing barotolerance. SIGNIFICANCE AND IMPACT OF THE STUDY: Inactivation of bacteria by high pressure treatment is influenced by their history which modulates barotolerance. Mechanisms of barotolerance appear different from heat shock defence. PMID- 15287875 TI - Isolation of salt-sensitive mutants from Sinorhizobium meliloti and characterization of genes involved in salt tolerance. AB - AIMS: The purpose of our research is to isolate salt-sensitive mutants and to study the genes involved in salt tolerance of the salt-tolerant bacterium Sinorhizobium meliloti 042BM. METHODS: Wild type S. meliloti 042BM bacteria are able to grow at a NaCl concentration of 0.6 mol l(-1). A transposon Tn5-1063a mutagenesis library of S. meliloti 042BM was constructed and eight salt-sensitive mutants were isolated, which were unable to growth on FY plates containing 0.4 mol l(-1) NaCl. SIGNIFICANCE: Our interest is to provide information about the mechanism of salt tolerance in bacteria by studying the genes involved in salt tolerance. Here, seven different genes were identified. These genes include omp10 encoding a cell outer membrane protein, relA encoding (p)ppGpp synthetase, greA encoding a transcription cleavage factor, nuoL encoding NADH dehydrogenase I chain L transmembrane protein, a putative nuclease/helicase gene and two unknown genes. Based on these findings, we suggest that the regulation of salt tolerance of S. meliloti 042BM is complex and on several levels. PMID- 15287877 TI - Quantification of Listeria monocytogenes in fermented sausages by MPN-PCR method. AB - AIMS: To combine the principles of most-probable-number (MPN) statistics and the conventional PCR technique to enumerate Listeria monocytogenes in fermented sausages. METHODS AND RESULTS: A simple method to enumerate L. monocytogenes in fermented sausages was developed and compared with direct plating in Palcam agar. Species-specific MPN-PCR, but not direct plating, made the enumeration of L. monocytogenes possible in all assayed samples. CONCLUSIONS: MPN-PCR proved to be a rapid and reliable method for enumerating L. monocytogenes in fermented sausages, including low contaminated samples. SIGNIFICANCE AND IMPACT OF THE STUDY: This MPN-PCR technique may facilitate the enumeration of L. monocytogenes for routine analyses in fermented sausages without excessive work. PMID- 15287878 TI - The behaviour of log phase Escherichia coli at temperatures that fluctuate about the minimum for growth. AB - AIMS: To investigate the behaviour of cold-adapted, log phase Escherichia coli exposed to temperatures that fluctuate below and above the minimum for growth. METHODS AND RESULTS: Log phase E. coli cultures were incubated at a constant temperature of 2, 4 or 6 degrees C or with temperatures allowed to increase from those temperatures for 35 min, to 10 degrees C, at 6-, 12- or 24-h intervals, as commonly occurs during retail display of chilled foods. At suitable intervals for each culture, the optical absorbance value was determined using a spectrophotometer, the forward angle light scatter was determined using a flow cytometer, and portions were spread on plate count agar for enumeration of colony forming units (CFU). Numbers of CFU decreased by 3 log units or increased by 1 log unit for cultures incubated at 6 degrees C for 17 days without or with temperatures fluctuations at < or =12-h intervals, respectively. Cells elongated when cultures were incubated at 4 or 2 degrees C with temperatures fluctuating at 6-h intervals, and at 6 degrees C at constant or fluctuating temperatures, but cells did not elongate in cultures incubated at a constant temperature of 2 or 4 degrees C. SIGNIFICANCE AND IMPACT OF THE STUDY: The minimum growth temperature of E. coli is assumed to be > or =7 degrees C. Elongated cells were able to divide when temperatures rose from 6 degrees C to above 7 degrees C for <45 min at < or =12-h intervals. Such temperature fluctuations may be experienced by chilled foods during defrosting cycles of retail display cases. The finding that cells behave differently under fluctuating than at constant temperatures may significantly affect understanding of appropriate temperatures for the safe storage of chilled foods and for predictive modelling of bacterial growth in such foods. PMID- 15287879 TI - A fluorescence-based method for the detection of adhesive properties of lactic acid bacteria to Caco-2 cells. AB - AIMS: The ability of probiotic micro-organisms to adhere to the intestinal surface is regarded as a substantial advantage in terms of bacteria persistence in the gastrointestinal tract. The aim of the present study was the development of a method based on fluorescent staining of bacteria and subsequent spectrofluorimetric detection to quantify the adhesion of several strains of Lactobacillus and Bifidobacterium to Caco-2 cells. METHODS AND RESULTS: Lactic acid bacteria strains were subjected to fluorescent staining using the viable probe carboxyfluorescein diacetate and subsequently incubated on Caco-2 monolayers. The adhesion of the micro-organisms was determined by spectrofluorimetry following the lysis of the attached bacterial cells and expressed as adhesion percentage. The values obtained for the micro-organisms tested ranged from 4% for Bifidobacterium infantis Bi1 to 10% for a Bifidobacterium mixture containing three different strains. CONCLUSIONS: In the present study we successfully applied fluorescent labelling and fluorimetric detection to investigate the adhesive properties of some Lactobacillus and Bifidobacterium strains and a Bifidobacterium mixture to Caco-2 cells. SIGNIFICANCE AND IMPACT OF THE STUDY: The results proved that fluorescent labelling is suitable for adhesion studies and provides a reliable and safer alternative to radioactive labelling. PMID- 15287880 TI - Kinetics and effects of trace elements and electron complexes on 2-keto-4 methylthiobutyric acid-dependent biosynthesis of ethylene in soil. AB - AIMS: 2-Keto-4-methylthiobutyric acid (KMBA) is an established intermediate in microbial biosynthesis of ethylene from methionine. This study demonstrates the kinetics and effects of trace elements and electron complexes on substrate (KMBA) derived C2H4 biosynthesis in soil. METHODS AND RESULTS: We have previously reported KMBA-dependent C2H4 production in soil. We studied the kinetics and effects of various trace elements and electron complexes on KMBA-derived C2H4 biosynthesis in soil by gas chromatography. Kinetic analysis revealed that ethylene forming enzyme (EFE) reaction was linear (R2 = 0.9448) when velocity of reaction (V) was plotted against substrate [S] over the range from 2.5 to 10 mmol l(-1) and thus followed a first order reaction. Application of three linear transformations of the Michaelis-Menten equation indicated high affinity of EFE for the substrate because Km values ranged between 5.4 and 6.67 mmol l(-1) and Vmax of reaction was between 22.4 and 35.7 nmol kg(-1) soil 120 cm(-1). Most of the trace elements exhibited positive effects on KMBA-dependent C2H4 production in soil. Maximum stimulatory effect on C2H4 biosynthesis was observed in response to Co(II) application, while Fe(III) inhibited the biotransformation of KMBA into C2H4. Contrarily, most of the tested electron complexes inhibited KMBA-derived C2H4 biosynthesis in the soil. However, lower concentrations (1.0 mmol l(-1)) of mannitol and hydroquinone were stimulatory to C2H4 production in soil compared with controls (substrate only). CONCLUSIONS: The results revealed that both kind and concentration of trace elements and electron complexes affected the substrate dependent production of C2H4 in soil with different degrees of efficacy. SIGNIFICANCE AND IMPACT OF THE STUDY: The C2H4 in the root environment could be physiologically active even at low concentrations, so knowledge regarding various factors which regulate C2H4 biosynthesis in soil could be of significance for plant growth and development. PMID- 15287881 TI - MAP kinases in the mammalian circadian system--key regulators of clock function. AB - Over the past 7 years, there has been spectacular progress in our understanding of the molecular basis of the circadian pacemaker in many species, from yeast to mammals. However, the biochemical signalling mechanisms that underpin synchronization of the clock to environmental cues are still poorly understood. Recently, attention has been focused on the role of mitogen-activated protein (MAP) kinase in biological timekeeping. It has been proposed that signal transduction via the MAP kinase cascades allows environmental information to be assimilated intracellularly within the circadian clock to produce changes in the phasing of clock gene expression, which, in turn, underlies clock-controlled phase-resetting of biological rhythms. This review examines the evidence for MAP kinase, particularly extracellular regulated kinases 1/2, involvement in the circadian clock and looks at the putative upstream regulators and downstream substrates of this signalling system. PMID- 15287882 TI - N-acetylaspartate: a literature review of animal research on brain ischaemia. AB - The present review examines and discusses the changes in N-acetylaspartate (NAA) concentration in the ischaemic brain from the existing literature of animal research. By summarizing the current knowledge on NAA metabolic pathways and reviewing the data obtained in animal models of global and focal ischaemia, the following conclusions emerge from this compilation: (i) both magnetic resonance spectroscopy (MRS) and the absolute HPLC method of NAA quantification give converging information; (ii) decreases in brain NAA concentration in acute stroke can be considered as an index of neuronal loss or dysfunction, although NAA redistribution by glial cells and NAA trapping in cell debris restrict its use as a quantitative neuronal marker; (iii) further studies on NAA metabolism in pathologic brain are required before using NAA measurement in the chronic stage of ischaemia for evaluating neuroprotective strategies. PMID- 15287883 TI - Up-regulation of Na(+)-Ca2+ exchange activity by interferon-gamma in cultured rat microglia. AB - The Na(+)-Ca2+ exchanger (NCX) plays a role in regulating intracellular Ca2+ concentration, but little is known about NCX in microglia. We examined mRNA expression of NCX isoforms in cultured rat microglia and the effect of interferon gamma (IFN-gamma) on NCX activity. RT-PCR showed that all of the known NCX isoforms, NCX1-3, are expressed in cultured microglia. Ouabain and monensin increased 45Ca2+ uptake and intracellular Ca2+ concentration in microglia, suggesting the presence of NCX activity in the reverse mode. Treatment with IFN gamma (100 U/mL) caused a biphasic increase in NCX activity. The transient increase in NCX activity by IFN-gamma for 1 h was blocked by the protein kinase C (PKC) inhibitors, staurosporine and GF109203X, and the tyrosine kinase inhibitor, herbimycin A. The delayed increase in NCX activity by IFN-gamma for 24 h was blocked by the protein synthesis inhibitor cycloheximide and actinomycin D. Treatment with IFN-gamma for 24 h increased NCX mRNA and protein levels. The increase in NCX activity and NCX protein by IFN-gamma for 24 h was blocked by staurosporine, GF109203X, herbimycin A and the extracellular signal-regulated kinase inhibitor, PD98059. These findings suggest that NCX is up-regulated by IFN gamma in a biphasic manner in microglia. Moreover, PKC and tyrosine kinase are involved in the up-regulation of NCX and the extracellular signal-regulated protein kinase is also involved in the delayed increase in NCX activity. PMID- 15287884 TI - Dopamine and cyclic AMP-regulated phosphoprotein-32 phosphorylation pattern in cocaine and morphine-sensitized rats. AB - This study reports some of the modifications in dopaminergic signalling that accompany cocaine and morphine behavioural sensitization. Cocaine-sensitized rats showed increased phosphorylation of dopamine- and cyclic AMP-regulated phosphoprotein Mr 32 kDa (DARPP-32) at threonine-75 (Thr75) and decreased DARPP 32 phosphorylation at Thr34, in the caudate-putamen (CPu) and nucleus accumbens (NAc) 7 days after sensitization assessment. Conversely, in morphine-sensitized rats, no apparent modifications in DARPP-32 phosphorylation pattern were observed. Morphine-sensitized rats have increased binding and coupling of micro opioid receptors and increased dopaminergic transmission in striatal areas and, upon morphine challenge, exhibit dopamine D1 receptor-dependent stereotypies. Thus, the DARPP-32 phosphorylation pattern was studied in morphine-sensitized rats at different times after morphine challenge. Morphine challenge increased levels of phospho-Thr75 DARPP-32 and decreased levels of phospho-Thr34 DARPP-32 in a time-dependent manner in the CPu and NAc. In order to assess whether these modifications were related to modified cyclic AMP-dependent protein kinase (PKA) activity, the phosphorylation levels of two other PKA substrates were examined, the GluR1 and NR1 subunits of alpha-amino-3-hydroxy-5-methylisoxazole-4 propionate and NMDA receptors respectively. The phosphorylation levels of GluR1 and NR1 subunits decreased in parallel with those of phospho-Thr-34 DARPP-32, supporting the hypothesis that morphine challenge elicited a decrease in PKA activity in morphine-sensitized rats. PMID- 15287885 TI - Presenilin-directed inhibitors of gamma-secretase trigger caspase 3 activation in presenilin-expressing and presenilin-deficient cells. AB - The amyloid beta peptide (Abeta) is generated by subsequent cleavages by beta- and gamma-secretases. Therefore, these two enzymes are putative therapeutic targets to prevent Abeta production, and hopefully to slow down or even stop the Alzheimer's disease (AD) neurodegenerative process. Several studies have revealed that gamma-secretase hydrolyses other important substrates besides beta-amyloid precursor protein (betaAPP) thus adding another level of complexity to designing fully AD-specific interfering drugs. Here we demonstrate that three distinct presenilin-directed gamma-secretase inhibitors as well as JLK compounds indirectly potentiate caspase 3 activity, the effector caspase of the apoptotic cascade. Thus, inhibitors were shown to drastically stimulate caspase 3 activity in wild-type mice blastocyst-derived and fibroblast cells. Interestingly, some of these inhibitors known to interact with presenilins also trigger caspase activation in presenilin-deficient cells. However, inhibitors do not affect recombinant caspase 3 activity, indicating that the effect on this enzyme was indirect. Furthermore, we established that caspase 3 activation was not due to an effect of gamma-secretase inhibitors on calpains, a family of proteolytic enzymes able to modulate caspase 3 activity. Altogether, our data demonstrate that presenilin-directed gamma-secretase inhibitors affect caspase 3 activity in a presenilin-independent manner. Therefore, as presenilin-dependent gamma-secretase activity is not specific for betaAPP and because its inhibitors clearly affect other vital cell functions, care should be taken in considering 'gamma-secretase' inhibitors as putative therapeutic tools to interfere with AD pathology. PMID- 15287886 TI - AMPA protects cultured neurons against glutamate excitotoxicity through a phosphatidylinositol 3-kinase-dependent activation in extracellular signal regulated kinase to upregulate BDNF gene expression. AB - The signal transduction and molecular mechanisms underlying alpha-amino-3-hydroxy 5-methyl-4-isoxazole propionate (AMPA)-mediated neuroprotection are unknown. In the present study, we determined a major AMPA receptor-mediated neuroprotective pathway. Exposure of cerebellar granule cells to AMPA (500 microM) + aniracetam (1 microM), a known blocker of AMPA receptor desensitization, evoked an accumulation of brain-derived neurotropic factor (BDNF) in the culture medium and enhanced TrkB-tyrosine phosphorylation following the release of BDNF. AMPA also activated the src-family tyrosine kinase, Lyn, and the downstream target of the phosphatidylinositol 3-kinase (PI3-K) pathway, Akt. Extracellular signal regulated kinase (ERK), a component of the mitogen-activated protein kinase (MAPK) pathway, was also activated. K252a, a selective inhibitor of neurotrophin signaling, blocked the AMPA-mediated neuroprotection. The involvement of BDNF release in protecting neurons by AMPA was confirmed using a BDNF-blocking antibody. AMPA-mediated neuroprotection is blocked by PP1, an inhibitor of src family kinases, LY294002, a PI3-K inhibitor, or U0126, a MAPK kinase (MEK) inhibitor. Neuroprotective concentrations of AMPA increased BDNF mRNA levels that was blocked by the AMPA receptor antagonist, 1,2,3,4-tetrahydro-6-nitro-2,3-dioxo benzo[f]quinoxaline-7-sulfonamide (NBQX). The increase in BDNF gene expression appeared to be the downstream target of the PI3-K-dependent activation of the MAPK cascade since MEK or the PI3-K inhibitor blocked the AMPA receptor-mediated increase in BDNF mRNA. Thus, AMPA receptors protect neurons through a mechanism involving BDNF release, TrkB receptor activation, and a signaling pathway involving a PI3-K dependent activation of MAPK that increases BDNF expression. PMID- 15287887 TI - Metalloendopeptidase EC3.4.24.15 is constitutively released from the exofacial leaflet of lipid rafts in GT1-7 cells. AB - Metalloendopeptidase EC3.4.24.15 (EP24.15) is a physiologically important neuropeptide-degrading enzyme involved in the metabolism of multiple neuropeptides. The mechanism of release of EP24.15 from neuronal cells is multimodal, being both constitutive and stimulatable. Previous studies have characterized stimulated EP24.15 secretion, yet little is understood concerning constitutive release of the peptidase. Utilizing the mouse hypothalamic neuronal GT1-7 cell line, we demonstrate that EP24.15 exists within lipid rafts in the plasma membrane, and that the enzyme is localized to the exofacial leaflet of lipid rafts. Further, we have found that biotinylated EP24.15 on the extracellular surface is released into the cell media in a fashion similar to constitutive release. In addition, classical and non-classical secretion pathway inhibitors were employed to understand the release of EP24.15 into surrounding cell media. The non-classical secretion inhibitor glyburide, a blocker of ATP sensitive K+ channels, decreased the amount of constitutively released EP24.15 in cell media of GT1-7 cells. With these data, we conclude that EP24.15 association with lipid rafts on the extracellular surface precedes constitutive release of the peptidase into the extracellular milieu for its action on neuropeptides. PMID- 15287888 TI - Phosphorylated p38MAPK specific antibodies cross-react with sarkosyl-insoluble hyperphosphorylated tau proteins. AB - Neurofibrillary tangles (NFT) accumulated in Alzheimer's diseases and related disorders contain hyperphosphorylated tau and display immunoreactivity for active forms of various kinases. To understand the role of p38MAPK (mitogen-activated protein kinase) in NFT formation, we have studied a transgenic (Tg) mouse model of tauopathy, JNPL3, that expresses P301L mutant tau, and bigenic mice, TAPP, generated by cross-breeding of JNPL3 with Tg2576 mice. Age-matched non-Tg mice (NTg), wild-type human tau Tg mice (JN25), and Tg2576 mice were used as controls. Phosphorylated p38MAPK (active form) immunoreactivity was consistently located in NFT and granulovaculolar degeneration in JNPL3 and TAPP mice older than 5 months of age. Unphosphorylated/total-p38MAPK was not detectable in spinal cord and brain sections from 2- to 11-month-old mice, even though JNPL3 mice, but not controls had an age-dependent increase of total-p38MAPK by western blotting. Spinal cord/brain extracts from mice and human with tauopathy were demonstrated to have insignificant amount of active-p38MAPK. However, they contained antiactive-p38MAPK cross-reactive proteins insoluble in sarkosyl and similar to phosphorylated tau in size. Consistently, antiactive-p38MAPK immunoprecipitates displayed tau immunoreactivity, but not total-p38MAPK, and antitau immunoprecipitates displayed active-p38MAPK immunoreactivity. Together, the results indicate that the cross-reactivity of antiactive-p38MAPK antibody with phosphorylated tau is responsible for the immunolabeling of tau-positive inclusion. PMID- 15287889 TI - Effects of repeated cocaine on medial prefrontal cortical GABAB receptor modulation of neurotransmission in the mesocorticolimbic dopamine system. AB - Increased excitatory output from medial prefrontal cortex is an important component in the development of cocaine sensitization. Activation of GABAergic systems in the prefrontal cortex can decrease glutamatergic activity. A recent study suggested that sensitization might be associated with a decrease in GABAB receptor responsiveness in the medial prefrontal cortex. Therefore, the present study examined whether repeated exposure to cocaine-modified neurochemical changes in the mesocorticolimbic dopamine system induced by infusion of baclofen into the medial prefrontal cortex. In vivo microdialysis studies were conducted to monitor dopamine, glutamate and GABA levels in the medial prefrontal cortex and glutamate levels in the ipsilateral nucleus accumbens and ventral tegmental area during the infusion of baclofen into medial prefrontal cortex. Baclofen minimally affected glutamate levels in the medial prefrontal cortex, nucleus accumbens or ventral tegmental area of control animals, but dose-dependently increased glutamate levels in each of these regions in animals sensitized to cocaine. This effect was not the result of changes in GABAB receptor-mediated modulation of dopamine or GABA in the medial prefrontal cortex. The data suggest that alterations in GABAB receptor modulation of medial prefrontal cortical excitatory output may play an important role in the development of sensitization to cocaine. PMID- 15287890 TI - Changes occurring in cortical NO release and brain NO-synthases during a paradoxical sleep deprivation and subsequent recovery in the rat. AB - Variations occurring in cortical nitric oxide (NO) release were analysed with a voltametric method in rats (i) placed in control conditions, (ii) while being paradoxical sleep deprived (PSD), or (iii) recovering from a PSD. Activities of neuronal (nNOS) and inducible (iNOS) NO-synthases as well as nNOS expression were also determined in several brain regions. In baseline conditions, circadian variations in nNOS expression and activity were maximal during the dark period and minimal during the light one for all the structures analysed (frontal cortex, pons and medulla). In the same way, cortical NO release occurred through a circadian rhythm exhibiting maxima and minima during dark and light periods, respectively. In the same experimental conditions, iNOS activity did not exhibit time-dependent changes. The correlative changes observed in baseline conditions between NO release, nNOS expression and activity within the frontal cortex were disrupted during PSD and subsequent recovery. Still again, iNOS activity remained unchanged. Results obtained point out that the tight coupling existing in control conditions between nNOS expression-activity and NO release is disrupted by a PSD and remains affected during the subsequent 24 h recovery. Their significance is discussed. PMID- 15287891 TI - Phosphorylation of Ser19 increases both Ser40 phosphorylation and enzyme activity of tyrosine hydroxylase in intact cells. AB - We have previously shown that the phosphorylation of Ser19 in tyrosine hydroxylase can increase the rate of phosphorylation of Ser40 in tyrosine hydroxylase threefold in vitro. In this report we investigated the role of Ser19 on Ser40 phosphorylation in intact cells. Treatment of bovine chromaffin cells with anisomycin produced a twofold increase in Ser19 phosphorylation with no increase in Ser31 phosphorylation and only a small increase in Ser40 phosphorylation. Treatment of bovine chromaffin cells with forskolin produced a fourfold increase in Ser40 phosphorylation but no significant increase in either Ser19 or Ser31 phosphorylation. When chromaffin cells were first treated with anisomycin, the level of Ser40 phosphorylation after treatment by forskolin was 76% greater than the level of Ser40 phosphorylation in cells treated with forskolin alone. This potentiation of Ser40 phosphorylation by anisomycin could be completely blocked by the p38 MAP (mitogen-activated protein) kinase inhibitor SB 203580. The potentiation of Ser40 phosphorylation by anisomycin was not due to an increase in Ser40 kinase activity. Anisomycin treatment of chromaffin cells potentiated the forskolin-induced increase in tyrosine hydroxylase activity by 50%. This potentiation of activity was also blocked by SB 203580. These data provide the first evidence that the phosphorylation of Ser19 can potentiate the phosphorylation of Ser40 and subsequent activation of tyrosine hydroxylase in intact cells. PMID- 15287893 TI - Activation of AP-1 and CRE-dependent gene expression via mu-opioid receptor. AB - Addiction to opiates depend on drug-induced neuroplastic changes and are underlain by alterations of gene expression. Transcription factors Ca2+/cAMP responsive element binding protein (CREB) and activator protein 1 (AP-1) may constitute a direct link between the opioid-regulated signal transduction pathways and modulation of gene expression. Acute treatment of Neuro2a MOR neuroblastoma cells with opioids stimulated CREB activity; prolonged treatment normalized it, while withdrawal from the drug again elicited an increase in phosphorylated CREB levels. Protein kinase C was responsible for the activation of transcription following acute opioid administration whereas the cAMP pathway activated similar mechanisms during withdrawal, making CREB a kind of 'a trigger' reacting to the presence or withdrawal of the opioid signal. Apart from the elevated CREB phosphorylation, CRE binding activity and expression of luciferase reporter gene regulated by CRE elements were increased after single administration and during withdrawal from the prolonged opioid treatment. Along with CREB, AP-1 binding activity and AP-1-directed transcription were stimulated after single administration and during withdrawal from the opioid. These results provide evidence that both single opioid administration and opioid withdrawal activate CREB and CRE-dependent transcriptional mechanisms via distinct intracellular signaling pathways. PMID- 15287892 TI - Regulation of dopamine D-receptor activation in vivo by protein phosphatase 2B (calcineurin). AB - Mice lacking dopamine D2 receptors exhibit a significantly decreased agonist promoted forebrain neocortical D1 receptor activation that occurs without changes in D1 receptor expression levels. This raises the possibility that, in brains of D2 mutants, a substantial portion of D1 receptors are uncoupled from their G protein, a phenomenon known as receptor desensitization. To test this, we examined D1-agonist-stimulated [35S]GTPgammaS binding (in the presence and absence of protein phosphatase inhibitors) and cAMP production (in the presence and absence of pertussis toxin) in forebrain neocortical tissues of wild-type mice and D2-receptor mutants. These studies revealed a decreased agonist stimulated G-protein activation in D2 mutants. Moreover, whereas protein phosphatase 1/2A (PP1/2A) and 2B (PP2B) inhibitors decrease [35S]GTPgammaS binding in a concentration-dependent manner in wild type, they have either no (PP2B) or only partial (PP1/2A) effects in D2 mutants. Furthermore, for D2 mutants, immunoprecipitation experiments revealed increased basal and D1-agonist stimulated phosphorylation of D1-receptor proteins at serine residues. Finally, D1 immunoprecipitates of both wild type and D2 mutants also contain protein kinase A (PKA) and PP2B immunoreactivities. In D2 mutants, however, the catalytic activity of the immunoprecipitated PP2B is abolished. These data indicate that neocortical D1 receptors are physically linked to PKA and PP2B and that the increased phosphorylation of D1 receptors in brains of D2 mutants is due to defective dephosphorylation of the receptor rather than increased kinase-mediated phosphorylation. PMID- 15287894 TI - Distribution of CNT2 and ENT1 transcripts in rat brain: selective decrease of CNT2 mRNA in the cerebral cortex of sleep-deprived rats. AB - Nucleoside transport processes regulate the levels of adenosine available to modulate neurotransmission, vascular tone and other physiological events. However, although equilibrative transporter transcripts or proteins have been mapped in the central nervous system of rats and humans, little is known about the presence and distribution of the complete family of nucleoside transporters in brain. In this study, we analysed the distribution of the transcript encoding the high affinity adenosine-preferring concentrative transporter CNT2 in the rat central nervous system and compared it with that of the equilibrative transporter ENT1. Furthermore, we evaluated the changes in expression of these two transporters in a situation of increased extracellular levels of adenosine, such as sleep deprivation. CNT2 mRNA was widespread in rat brain, although most prevalent in the amygdala, the hippocampus, specific neocortical regions and the cerebellum. The distribution of CNT2 mRNA only partially overlapped that of ENT1. Most of the cells labelled were neurones. Total sleep deprivation dramatically diminished the amounts of CNT2 mRNA, whereas ENT1 mRNA remained unchanged. This specific decrease in CNT2 transcript suggests a new physiological role for the transporter in the modulation of extracellular adenosine levels and the sleep/wakefulness cycle. PMID- 15287895 TI - Nomifensine amplifies subsecond dopamine signals in the ventral striatum of freely-moving rats. AB - The present experiment examined the effect of the dopamine transporter blocker nomifensine on subsecond fluctuations in dopamine concentrations, or dopamine transients, in the nucleus accumbens and olfactory tubercle. Extracellular dopamine was measured in real time using fast-scan cyclic voltammetry at micron dimension carbon fibers in freely-moving rats. Dopamine transients occurred spontaneously throughout the ventral striatum in the absence of apparent sensory input or change in behavioral response. The frequency of dopamine transients increased at the presentation of salient stimuli to the rat (food, novel odors and unexpected noises). Administration of 7 mg/kg nomifensine amplified spontaneous dopamine transients by increasing both amplitude and duration, consistent with its known action at the dopamine transporter and emphasizing the dopaminergic origin of the signals. Moreover, nomifensine increased the frequency of detected dopamine transients, both during baseline conditions and at the presentation of stimuli, but more profoundly in the nucleus accumbens than in the olfactory tubercle. This difference was not explained by nomifensine effects on the kinetics of dopamine release and uptake, as its effects on electrically evoked dopamine signals were similar in both regions. These findings demonstrate the heterogeneity of dopamine transients in the ventral striatum and establish that nomifensine elevates the tone of rapid dopamine signals in the brain. PMID- 15287897 TI - Differential expression of N-methyl-D-aspartate receptor NR2 isoforms in Alzheimer's disease. AB - We have previously shown that the expression of NMDA receptor NR1 subunit mRNA splice variants in Alzheimer's disease (AD) brain varies according to regional susceptibility to pathological damage. Here we investigated the expression of the modulatory NR2 subunits of the NMDA receptor using quantitative RT-PCR to assay all NR2 isoforms. Significantly lower expression of NR2A and NR2B transcripts was found in susceptible regions of AD brain, whereas expression of NR2C and NR2D transcripts did not differ from that in controls. Western blot analysis confirmed a lower expression of the NR2A and NR2B isoforms at the protein level. The results suggest that NR2 subunit composition may modulate NMDA receptor-mediated excitotoxicity. NMDA receptor dysfunction might give rise to the regionally selective pattern of neuronal loss that is characteristic of AD. PMID- 15287896 TI - Focal adhesion kinase mediates endothelin-induced cyclin D3 expression in rat cultured astrocytes. AB - Focal adhesion kinase (FAK), a non-receptor type tyrosine kinase, is involved in the G1/S phase cell cycle transition of astrocytes induced by endothelin-1 (ET 1). In this study, the roles of FAK in the expression of cyclin D1 or D3, which are pivotal in G1/S phase transition, were examined in cultured astrocytes. Accompanied with increases in bromodeoxyuridine (BrdU) incorporation, ET-1 (100 nm) increased the numbers of cyclin D1- and D3-positive astrocytes. PD98059 (a MEK inhibitor) and PP-2 (a Src kinase inhibitor) inhibited ET-induced cyclin D1 expression and BrdU incorporation without affecting cyclin D3 expression. In contrast, cytochalasin D, lovastatin (a 3-hydroxy-3-methylglutaryl-CoA reductase inhibitor) and Y-27632 (a rho-kinase inhibitor) prevented both cyclin D3 expression and BrdU incorporation. FAK phosphorylation by ET-1 was inhibited by cytochalasin D, lovastatin and Y-27632, but not by PD98059 or PP-2. Transfection with wild-type FAK increased expression of cyclin D3 in astrocytes, while that of cyclin D1 was not affected. Dominant-negative FAK mutants prevented an ET-induced increase in cyclin D3 expression, but not D1. These results suggest that activation of FAK causes cyclin D3 expression in cultured astrocytes, which would underlie the FAK-mediated astrocytic G1/S phase transition by ET-1. PMID- 15287898 TI - Intracellular lithium and cyclic AMP levels are mutually regulated in neuronal cells. AB - In this work, we studied the effect of intracellular 3',5'-cyclic adenosine monophosphate (cAMP) on Li+ transport in SH-SY5Y cells. The cells were stimulated with forskolin, an adenylate cyclase activator, or with the cAMP analogue, dibutyryl-cAMP. It was observed that under forskolin stimulation both the Li+ influx rate constant and the Li+ accumulation in these cells were increased. Dibutyryl-cAMP also increased Li+ uptake and identical results were obtained with cortical and hippocampal neurons. The inhibitor of the Na+/Ca2+ exchanger, KB R7943, reduced the influx of Li+ under resting conditions, and completely inhibited the effect of forskolin on the accumulation of the cation. Intracellular Ca2+ chelation, or inhibition of N-type voltage-sensitive Ca2+ channels, or inhibition of cAMP-dependent protein kinase (PKA) also abolished the effect of forskolin on Li+ uptake. The involvement of Ca2+ on forskolin-induced Li+ uptake was confirmed by intracellular free Ca2+ measurements using fluorescence spectroscopy. Exposure of SH-SY5Y cells to 1 mm Li+ for 24 h increased basal cAMP levels, but preincubation with Li+, at the same concentration, decreased cAMP production in response to forskolin. To summarize, these results demonstrate that intracellular cAMP levels regulate the uptake of Li+ in a Ca(2+)-dependent manner, and indicate that Li+ plays an important role in the homeostasis of this second messenger in neuronal cells. PMID- 15287899 TI - Organic anion transporter 3 is involved in the brain-to-blood efflux transport of thiopurine nucleobase analogs. AB - Thiopurines are used as antileukemic drugs. However, during chemotherapy CNS relapses occur due to the proliferation of leukemic cells in the CNS resulting from restricted drug distribution in the brain. The molecular mechanism for this limited cerebral distribution remains unclear. The purpose of this study was to identify the transporter responsible for the brain-to-blood transport of thiopurines across the blood-brain barrier (BBB) using the brain efflux index method. [14C]6-Mercaptopurine (6-MP) and [3H]6-thioguanine were eliminated from rat brain in a time-dependent manner. The elimination of [14C]6-MP was inhibited by substrates of rat organic anion transporters (rOATs), including indomethacin and benzylpenicillin. rOAT1 and rOAT3 exhibited 6-MP uptake, while benzylpenicillin inhibited rOAT3-mediated uptake, but not that by rOAT1. rOAT3 mediated [14C]6-MP uptake was also inhibited by other thiopurine derivatives. Although methotrexate inhibited rOAT3-mediated [14C]6-MP uptake, the Ki value was 17.5-fold greater than the estimated brain concentration of methotrexate in patients receiving chemotherapy. Accordingly, 6-MP would undergo efflux transport by OAT3 from the brain without any inhibitory effect from coadministered methotrexate in the chemotherapy. In conclusion, rOAT3 is involved in the brain to-blood transport of thiopurines at the BBB and is one mechanism of limited cerebral distribution. PMID- 15287900 TI - Lack of mitochondrial nitric oxide production in the mouse brain. AB - Based on our initial finding that the nitric oxide (NO) sensitive fluorochrome diaminofluorescein (DAF) was localized to mitochondria in cultured primary neurons, we investigated whether brain mitochondria produce NO through a mitochondrial NO synthase (mtNOS) enzyme. Isolated brain mitochondria were loaded with DAF and subjected to flow cytometry analysis. Neither the application of NOS inhibitors nor the genetic disruption of either NOS gene diminished the DAF fluorescence. However, peroxynitrite scavengers reduced the mitochondrial DAF fluorescence, indicating that the DAF signal is not specific to NO. Chemiluminescence detection in the head space gas and a Clark-type NO-sensitive electrode in the solution failed to detect NO release in brain mitochondria. NOS activity in mitochondria was only 1% of the whole brain NOS activity level, which may be attributed to extramitochondrial contamination. Extensive immunoblotting and immunoprecipitation experiments failed to show the presence of endothelial, neuronal, or inducible NOS in mouse brain mitochondria using a variety of primary antibodies. Arginine, calmodulin or 2,5-ADP affinity purification protocols successfully concentrated eNOS and nNOS from full brain tissue but failed to show any signal in mitochondria. We conclude that mouse brain mitochondria do not contain NOS isoforms, nor do they produce NO through a NOS-dependent mechanism. PMID- 15287901 TI - Effects of NGF on acetylcholine, acetyl-CoA metabolism, and viability of differentiated and non-differentiated cholinergic neuroblastoma cells. AB - Nerve growth factor (NGF) is a peptide displaying multiple cholinotropic activities. The aim of this work was to explain mechanisms of the positive and negative effects of NGF on phenotypic properties and viability of cholinergic cells. To discriminate these effects we used two p75NTR receptor-positive lines of cholinergic neuroblastoma cells, SN56 and T17 that are devoid of or express high affinity NGF (TrkA) receptors, respectively. cAMP and retinoic acid caused differentiation of both cell lines. In addition to the morphologic maturation, the increase of choline acetyltransferase activity, acetylcholine, Ca and cytoplasmic acetyl-CoA levels and decrease of mitochondrial acetyl-CoA and cell viability were observed. NGF caused similar effects in non-differentiated T17 cells but had no influence on non-differentiated SN56 cells. On the contrary, in both cAMP/all-trans-retinoic acid (RA) differentiated cell lines, NGF resulted in a similar suppression of cholinergic phenotype along with an increase of mitochondrial acetyl-CoA and cell susceptibility to nitric oxide and amyloid beta25-35. These effects of NGF were prevented by an antibody against the p75NTR receptor. Data indicate that: (i) positive cholinotrophic effects of NGF required activation of both TrkA and p75NTR receptors; (ii) cAMP/RA-evoked differentiation inhibited NGF effects mediated by TrkA receptors and activated its p75NTR dependent suppressing influences and (iii) a differentiation-evoked decrease of mitochondrial acetyl-CoA and an elevation of mitochondrial Ca could augment impairment of cholinergic neurons by neurotoxic signals. PMID- 15287902 TI - Cross-linking of glycine receptor transmembrane segments two and three alters coupling of ligand binding with channel opening. AB - Contact points between transmembrane segments (TMs) two and three of the glycine receptor are undefined and may play an important role in channel gating. We tested whether two amino acids in TM2 (S267) and TM3 (A288), known to be critical for alcohol and volatile anesthetic action, could cross-link by mutating both to cysteines and expressing the receptors in Xenopus laevis oocytes. In contrast with the wild-type receptor and single cysteine mutants, the S267C/A288C double mutant displayed unusual responses, including a tonic leak activity that was closed by strychnine and a run-down of the response upon repeated applications of glycine. We hypothesized that these characteristics were due to cross-linking of the two cysteines on opposing faces of these adjacent, alpha helical TMs. This would alter the movement of these two regions required for normal gating. To test this hypothesis, we used dithiothreitol to reduce the putative S267C-A288C disulfide bond. Reduction abolished the leak current and provided normal responses to glycine. Subsequent application of the cross-linking agent mercuric chloride caused the initial characteristics to return. These data demonstrate that S267 and A288 are near-neighbors and provide insight towards the location and role of the TM2-TM3 interface in ligand-gated ion channels. PMID- 15287903 TI - Effects of phosphorylation by protein kinase A on binding of catecholamines to the human tyrosine hydroxylase isoforms. AB - Tyrosine hydroxylase (TyrH), the catalyst for the key regulatory step in catecholamine biosynthesis, is phosphorylated by cAMP-dependent protein kinase A (PKA) on a serine residue in a regulatory domain. In the case of the rat enzyme, phosphorylation of Ser40 by PKA is critical in regulating the enzyme activity; the effect of phosphorylation is to relieve the enzyme from inhibition by dopamine and dihydroxyphenylalanine (DOPA). There are four isoforms of human tyrosine hydroxylase (hTyrH), differing in the size of an insertion after Met30. The effects of phosphorylation by PKA on the binding of DOPA and dopamine have now been determined for all four human isoforms. There is an increase of about two-fold in the Kd value for DOPA for isoform 1 upon phosphorylation, from 4.4 to 7.4 microM; this effect decreases with the larger isoforms such that there is no effect of phosphorylation on the Kd value for isoform 4. Dopamine binds more much tightly, with Kd values less than 3 nM for all four unphosphorylated isoforms. Phosphorylation decreases the affinity for dopamine at least two orders of magnitude, resulting in Kd values of about 0.1 microM for the phosphorylated human enzymes, due primarily to increases in the rate constant for dissociation of dopamine. Dopamine binds about two-fold less tightly to the phosphorylated isoform 1 than to the other three isoforms. The results extend the regulatory model developed for the rat enzyme, in which the activity is regulated by the opposing effects of catecholamine binding and phosphorylation by PKA. The small effects on the relatively high Kd values for DOPA suggest that DOPA levels do not regulate the activity of hTyrH. PMID- 15287904 TI - Docosahexaenoic acid promotes neurite growth in hippocampal neurons. AB - Docosahexanoic acid (22:6n-3; DHA) deficiency during development is associated with impairment in learning and memory, suggesting an important role of DHA in neuronal development. Here we provide evidence that DHA promotes neuronal differentiation in rat embryonic hippocampal primary cultures. DHA deficiency in vitro was spontaneously induced by culturing hippocampal cells in chemically defined medium. DHA supplementation improved DHA levels to values observed in freshly isolated hippocampus. We found that DHA supplementation in culture increased the population of neurons with longer neurite length per neuron and with higher number of branches. However, supplementation with arachidonic, oleic or docosapentaenoic acid did not have any effect, indicating specificity of the DHA action on neurite growth. Furthermore, hippocampal cultures obtained from n-3 fatty acid deficient animals contained a lower DHA level and a neuronal population with shorter neurite length per neuron in comparison to those obtained from animals with adequate n-3 fatty acids. DHA supplementation to the deficient group recovered the neurite length to the level similar to n-3 fatty acid adequate cultures. Our data demonstrates that DHA uniquely promotes neurite growth in hippocampal neurons. Inadequate neurite development due to DHA deficiency may contribute to the cognitive impairment associated with n-3 fatty acid deficiency. PMID- 15287905 TI - Biosynthesis of NAAG by an enzyme-mediated process in rat central nervous system neurons and glia. AB - The peptide transmitter N-acetylaspartylglutamate (NAAG) is present in millimolar concentrations in mammalian spinal cord. Data from the rat peripheral nervous system suggest that this peptide is synthesized enzymatically, a process that would be unique for mammalian neuropeptides. To test this hypothesis in the mammalian CNS, rat spinal cords were acutely isolated and used to study the incorporation of radiolabeled amino acids into NAAG. Consistent with the action of a NAAG synthetase, inhibition of protein synthesis did not affect radiolabel incorporation into NAAG. Depolarization of spinal cords stimulated incorporation of radiolabel. Biosynthesis of NAAG by cortical astrocytes in cell culture was demonstrated by tracing incorporation of [3H]-glutamate by astrocytes. In the first test of the hypothesis that NAA is an immediate precursor in NAAG biosynthesis, [3H]-NAA was incorporated into NAAG by isolated spinal cords and by cell cultures of cortical astrocytes. Data from cerebellar neurons and glia in primary culture confirmed the predominance of neuronal synthesis and glial uptake of NAA, leading to the hypothesis that while neurons synthesize NAA for NAAG biosynthesis, glia may take it up from the extracellular space. However, cortical astrocytes in serum-free low-density cell culture incorporated [3H]-aspartate into NAAG, a result indicating that under some conditions these cells may also synthesize NAA. Pre-incubation of isolated spinal cords and cultures of rat cortical astrocytes with unlabeled NAA increased [3H]-glutamate incorporation into NAAG. In contrast, [3H]-glutamine incorporation in spinal cord was not stimulated by unlabeled NAA. These results are consistent with the glutamate glutamine cycle greatly favoring uptake of glutamine into neurons and glutamate by glia and suggest that NAA availability may be rate-limiting in the synthesis of NAAG by glia under some conditions. PMID- 15287906 TI - Effects of post-injury hypothermia and nerve growth factor infusion on antioxidant enzyme activity in the rat: implications for clinical therapies. AB - The pathological sequelae of traumatic brain injury (TBI) include increased oxidative stress due to the production of reactive oxygen species (ROS). Regulation of ROS levels following TBI is determined primarily by antioxidant enzyme activity that in turn can be influenced by nerve growth factor (NGF). Hypothermia is one of the current therapies designed to combat the deleterious effects of TBI. However, it has been shown to suppress post-trauma increases in NGF levels in rat brain. The present study sought to determine whether post injury hypothermia also impairs the antioxidant response to injury, and if such an effect could be reversed by infusion of exogenous NGF. We employed a lateral controlled cortical impact injury model in rat, followed by moderate hypothermia treatment with supplemental intracerebroventricular infusion of NGF or vehicle. The time course of changes in post-injury/intervention levels of NGF and activity of three major enzymes responsible for ROS scavenging, catalase (CAT), glutathione peroxidase (GPx) and superoxide dismutase (SOD), was determined in the hippocampus. Relative to levels in injured, normothermic animals, hypothermia treatment not only suppressed NGF levels, but also attenuated CAT and GPx activity, and increased SOD activity. Infusion of NGF in injured, hypothermia treated animals was ineffective in restoring hippocampal antioxidant enzymes activity to levels produced after injury under normothermic conditions, although it was able to increase septal cholinergic (choline acetyltransferase) enzyme activity. These results have implications for clinical treatment of TBI, demonstrating that moderate hypothermia suppresses NGF and the antioxidant response after TBI; the latter cannot be countered by exogenous NGF administration. PMID- 15287907 TI - Atorvastatin-induced activation of Alzheimer's alpha secretase is resistant to standard inhibitors of protein phosphorylation-regulated ectodomain shedding. AB - Studies of metabolism of the Alzheimer amyloid precursor protein (APP) have focused much recent attention on the biology of juxta- and intra-membranous proteases. Release or 'shedding' of the large APP ectodomain can occur via one of two competing pathways, the alpha- and beta-secretase pathways, that are distinguished both by subcellular site of proteolysis and by site of cleavage within APP. The alpha-secretase pathway cleaves within the amyloidogenic Abeta domain of APP, precluding the formation of toxic amyloid aggregates. The relative utilization of the alpha- and beta-secretase pathways is controlled by the activation of certain protein phosphorylation signal transduction pathways including protein kinase C (PKC) and extracellular signal regulated protein kinase [ERK/mitogen-activated protein kinase (MAP kinase)], although the relevant substrates for phosphorylation remain obscure. Because of their apparent ability to decrease the risk for Alzheimer disease, the effects of statins (HMG CoA reductase inhibitors) on APP metabolism were studied. Statin treatment induced an APP processing phenocopy of PKC or ERK activation, raising the possibility that statin effects on APP processing might involve protein phosphorylation. In cultured neuroblastoma cells transfected with human Swedish mutant APP, atorvastatin stimulated the release of alpha-secretase-released, soluble APP (sAPPalpha). However, statin-induced stimulation of sAPPalpha release was not antagonized by inhibitors of either PKC or ERK, or by the co-expression of a dominant negative isoform of ERK (dnERK), indicating that PKC and ERK do not play key roles in mediating the effect of atorvastatin on sAPPalpha secretion. These results suggest that statins may regulate alpha-secretase activity either by altering the biophysical properties of plasma membranes or by modulating the function of as-yet unidentified protein kinases that respond to either cholesterol or to some intermediate in the cholesterol metabolic pathway. A 'phospho-proteomic' analysis of N2a cells with and without statin treatment was performed, revealing changes in the phosphorylation state of several protein kinases plausibly related to APP processing. A systematic evaluation of the possible role of these protein kinases in statin-regulated APP ectodomain shedding is underway. PMID- 15287908 TI - Aging, gender and APOE isotype modulate metabolism of Alzheimer's Abeta peptides and F-isoprostanes in the absence of detectable amyloid deposits. AB - Aging and apolipoprotein E (APOE) isoform are among the most consistent risks for the development of Alzheimer's disease (AD). Metabolic factors that modulate risk have been elusive, though oxidative reactions and their by-products have been implicated in human AD and in transgenic mice with overt histological amyloidosis. We investigated the relationship between the levels of endogenous murine amyloid beta (Abeta) peptides and the levels of a marker of oxidation in mice that never develop histological amyloidosis [i.e. APOE knockout (KO) mice with or without transgenic human APOEepsilon3 or human APOEepsilon4 alleles]. Aging-, gender-, and APOE-genotype-dependent changes were observed for endogenous mouse brain Abeta40 and Abeta42 peptides. Levels of the oxidized lipid F2 isoprostane (F2-isoPs) in the brains of the same animals as those used for the Abeta analyses revealed aging- and gender-dependent changes in APOE KO and in human APOEepsilon4 transgenic KO mice. Human APOEepsilon3 transgenic KO mice did not exhibit aging- or gender-dependent increases in F2-isoPs. In general, the changes in the levels of brain F2-isoPs in mice according to age, gender, and APOE genotype mirrored the changes in brain Abeta levels, which, in turn, paralleled known trends in the risk for human AD. These data indicate that there exists an aging-dependent, APOE-genotype-sensitive rise in murine brain Abeta levels despite the apparent inability of the peptide to form histologically detectable amyloid. Human APOEepsilon3, but not human APOEepsilon4, can apparently prevent the aging-dependent rise in murine brain Abeta levels, consistent with the relative risk for AD associated with these genotypes. The fidelity of the brain Abeta/F2-isoP relationship across multiple relevant variables supports the hypothesis that oxidized lipids play a role in AD pathogenesis, as has been suggested by recent evidence that F2-isoPs can stimulate Abeta generation and aggregation. PMID- 15287909 TI - Circadian rhythms in the retina of rats with photoreceptor degeneration. AB - Previous studies have demonstrated that the mammalian retina contains a circadian clock system that controls several retinal functions. In mammals the location of the retinal circadian clock is unknown whereas, in non-mammalian vertebrates, earlier work has demonstrated that photoreceptor cells contain the circadian clock. New experimental evidence has suggested that in mammals the retinal circadian clock may be located outside the photoreceptor cells. In this study we report that circadian rhythms in Aa-nat mRNA (in vivo) and melatonin synthesis (in vitro) are still present in the retina of rats lacking photoreceptors. The circadian pacemaker(s) controlling such rhythms is probably located in kainic acid sensitive neurons in the inner retina since kainic acid injections abolished the rhythmicity. These data are the first direct demonstration that circadian rhythmicity in the mammalian retina can be generated independently from the photoreceptors and the suprachiasmatic nuclei of the hypothalamus. PMID- 15287910 TI - Bone marrow biopsy in Hodgkin's lymphoma. AB - In the study of patients with Hodgkin's lymphoma (HL) the evaluation of bone marrow biopsy (BMB) can be difficult. In this review we analyze the main diagnostic features and the clinical risk factors of BM involvement. Although the role of BMB is criticized by some authors, its value is irreplaceable in the staging of HL and in the diagnosis of primary medullary HL. The Ann Arbor staging committee criteria should be revised and updated in the light of the current immunohistochemical studies that give a fundamental help in the diagnostic process. A single BMB should be adequate for diagnosis in most instances. In cases of suspicious involvement a controlateral BMB could be performed. PMID- 15287911 TI - A systematic approach to molecular quantitative determination of mixed chimaerism following allogeneic bone marrow transplantation: an analysis of its applicability in a group of patients with severe aplastic anaemia. AB - Mixed chimaerism (MC) following allogeneic bone marrow transplantation (allo-BMT) is defined as the persistent cohabitation of haematopoietic cells from recipients and donors. Its kinetics, clinical implications and more efficient laboratory approaches for MC detection are the object of ongoing research in view of the possibility of developing useful markers. Here we describe a sequential analysis of chimaerism using variable number of tandem repeat (VNTR) polymerase chain reaction (PCR) followed by quantitative, fluorescent labelled, short tandem repeat (STR) PCR. A set of four, highly discriminative VNTR and four STR markers was used to assess chimaerism. Sensitivity and regression analysis indicated that this approach was reliable for routine application in a single BMT centre. We studied 12 patients with severe aplastic anaemia (SAA) who had received allo-BMT, and had been conditioned with cyclosphosphamide (Cy) with or without anti thymocyte globulin (ATG). We found a 50% prevalence of MC in the whole group, with levels between 4% and 37% of recipient cells. A sustained stable MC pattern after BMT was characteristic of the Cy-only conditioned patients but was also recorded in one patient treated with the Cy + ATG regime who showed a sustained MC pattern over a period of 24 months post-BMT. In none of our patients, MC was associated with an increased risk of graft rejection in a median follow-up of 39.5 months. PMID- 15287912 TI - Mixed chimerism is frequent after allogeneic peripheral blood stem cell transplantation with positive CD34 selection, and is not reverted by low doses of donor T-cells add-back. AB - The incidence of full donor chimerism (full DC) after CD34+ -selected peripheral blood stem cell transplantation (CD34+ -PBSCT) is controversial. Whereas the initial reports suggested a high incidence of full DC (hypothetically because of the high number of CD34+ cells infused) more recent works describe a high incidence of mixed lymphoid chimerism. There are no data concerning the ability of low-dose donor T-lymphocyte add-back on conversion to full DC. METHODS: We prospectively monitored the chimerism status of 25 patients undergoing CD34+ PBSCT and the effect on chimerism of delayed low doses of donor T-cell add-back (TCAB). One, two or three doses of TCAB were administered on days +28 (2 x 10(5) CD3+/kg), +60 (2 x 10(5) CD3+/kg) and +90 (2 x 10(6) CD3+/kg), respectively, when on cyclosporine A prophylaxis. RESULTS: Incidence of full DC on day +20 was 56%. However, all but two patients progressed to MC. Fifteen patients were scheduled to TCAB. Six patients with initial MC did not convert to full DC after TCAB. Moreover, seven patients with full DC status progressed to mixed chimerism. CONCLUSIONS: Our results indicate that low doses of TCAB administered under cyclosporine A prophylaxis have no effect on the eradication of the recipient cells. We believe that a high dose of CD34+ cells in the grafts of CD34+ -PBSCT is not enough to achieve stable full DC unless a minimum number of CD3+ cells are infused, or more intensified transplant conditioning regimens are employed. PMID- 15287913 TI - High-dose chemotherapy and autologous peripheral blood stem cell transplantation in high-risk multiple myeloma. AB - We compared the effect of high-dose therapy together with autologous peripheral blood stem cell transplantation (autoPBSCT) in 60 patients with multiple myeloma (MM) with 90 patients who underwent conventional chemotherapy. We scored the prognostic factors according to our reported classification system that includes measurements of serum albumin, serum beta-2-microglobulin, and morphology of myeloma cells selected by multivariate analysis. We separated the patients into three risk groups at stratification level I (low, intermediate and high) and into two risk groups at stratification level II (low and high). AutoPBSCT tended to be as effective for high, as for low-risk patients in level I, and was obviously as helpful for high, as for low-risk patients in stratification II. In conclusion, high-risk patients with MM should be treated with high-dose therapy accompanied with autoPBSCT like low-risk patients. PMID- 15287914 TI - Invasive fungal infections in autologous stem cell transplant recipients: a nation-wide study of 1188 transplanted patients. AB - Based on small single-centre series, the risk of invasive fungal infections (IFI) has been considered small in autologous stem cell transplant (ASCT) recipients. PURPOSE: To analyse epidemiological and clinical features of (IFI) among ASCT recipients in Finland 1990-2001. PATIENTS: During the study period, 1188 adult patients received high-dose therapy supported by ASCT in six centres. Altogether, 1112 patients (94%) received blood progenitor cells. The graft was CD34+ selected in 261 patients (22%). The major diagnostic groups were non-Hodgkin's lymphoma (n = 417), multiple myeloma (n = 395), breast cancer (n = 132) and Hodgkin's lymphoma (n = 53). RESULTS: Eighteen patients (1.5%) with IFI were identified. The incidence of proven or probable invasive aspergillosis was 0.8%, followed by candidaemia with an incidence of 0.3%. The median time to the diagnosis of IFI was 35 d (6-162) from the progenitor cell infusion. In fourteen patients (78%) IFI was diagnosed during lifetime and they were treated with antifungal therapy for a median of 50 d. Nine patients (64%) were cured. CONCLUSIONS: IFI appears to be a rare event after ASCT and Aspergillus infections seem to be predominant. These epidemiological features have an impact in planning prophylactic and empirical antifungal strategies in ASCT recipients. PMID- 15287915 TI - ICE (ifosfamide, carboplatin, etoposide) as second-line chemotherapy in relapsed or primary progressive aggressive lymphoma--the Nordic Lymphoma Group experience. AB - OBJECTIVE: To evaluate ICE (ifosfamide, carboplatin, etoposide) as second-line chemotherapy in relapsed or primary progressive aggressive lymphoma, in terms of objective response rate (ORR) and peripheral blood stem cell (PBSC) harvest mobilization rate. PATIENT POPULATION: A total of 40 patients were included, with a median age of 57 yr. The major histopathological subgroup was diffuse large B cell lymphoma (n = 27). The indication for ICE was relapse in 23 patients, primary progressive disease in 11, transformation in four and adjuvant primary chemotherapy in one patient. RESULTS: After three cycles of ICE, the ORR was 59%. Among patients with primary progressive disease, ORR was 36% (four of 11). A PBSC harvest after ICE could be performed in 11 of 20 patients, and was sufficient for stem cell rescue in 10 of 20. The median number of collected CD34+ cells was 3.6 x 10(6) (range 1.4-12.5). In six of 10 patients, an adequate PBSC harvest could be performed with a second mobilization regimen. CONCLUSION: In this patient population, the rate of response to ICE was comparable with other second-line regimens used in aggressive lymphoma. The rate of harvest failure (45%) was disappointingly high, compared with previous reports, possibly because of patient selection or differences in granulocyte-colony stimulating factor (G-CSF) dosage. PMID- 15287916 TI - Microenvironment factors do not afford myeloma cell lines protection from simvastatin. AB - BACKGROUND: The intensive interactions of myeloma cells (multiple myeloma, MM) with microenvironmental components of the bone marrow contribute significantly to their proliferation and survival. It has been shown that these signals confer drug resistance, delineating their circumvention as a primary objective in disease treatment. This study was designed to assess the effect of some major extracellular factors on the previously established anti-neoplastic response of myeloma cells to simvastatin (Sim). STUDY DESIGN: RPMI8226, U266, and ARH77 seeded in culture plates precoated with fibronectin (FN)/agarose/none were treated with Sim, insulin-like growth factor-I (IGF-I), interleukin-6 (IL-6) or combinations for 5 d. Then we assessed cell morphology, viability (WST1), cell cycle (propidium iodide, PI, staining and flow cytometric analysis), total cell count, and cell death (trypan blue exclusion), and DNA fragmentation. RESULTS AND CONCLUSIONS: Reduced viability was demonstrated with Sim in all treated cell lines with and without co-administration of IGF-I or IL-6 (P < 0.05). The extent of inhibition did not vary between Sim only and combinations (NS). FN did not influence cell response to Sim alone or combined with IL-6/IGF-I (NS). We conclude that IL-6, IGF-I, and FN do not afford myeloma cell lines protection from Sim modulation. PMID- 15287917 TI - Initial expression of interferon alpha receptor 2 (IFNAR2) on CD34-positive cells and its down-regulation correlate with clinical response to interferon therapy in chronic myelogenous leukemia. AB - In order to investigate the mechanism of interferon-alpha (IFNalpha) action in the treatment of chronic myelogenous leukemia (CML), we examined surface expressions of both type I interferon receptor 1 (IFNAR1) and 2 (IFNAR2) subunits on CD34-positive cells in bone marrow (BM) in a total of 57 CML patients. Initial cell-surface IFNAR2 expression at diagnosis assessed by flow cytometry widely distributed but showed overall significantly higher expression in CML patients when compared with normal controls. In 15 fresh patients who subsequently received IFNalpha therapy, IFNAR2 expression at diagnosis was significantly higher in cytogenetic good responders than in poor responders. Down-regulation of IFNAR2 expression during IFNalpha therapy was observed only in good responders but not in poor responders. In addition to protein level, both initial high IFNAR2c mRNA expression level and its down-regulation during IFNalpha therapy, in purified CD34-positive cells, were also observed only in good responders. In contrast to IFNAR2, cell-surface IFNAR1 expression was generally lower than IFNAR2, and correlation between either the pretreatment level or down-regulation of IFNAR1 and clinical response was not evident. With in vitro IFNalpha stimulation, CD34-positive cells showed down-regulations of cell-surface IFNAR2, and IFNAR1 to a lesser extent, in one good-responder patient, but not in one poor responder patient. Serum soluble interferon receptor (sIFNR) was higher in untreated CML patients than in normal controls, without any correlation with clinical response to IFNalpha. Thus, the pretreatment protein and mRNA expression levels of IFNAR2 and their down-regulations during IFNalpha therapy correlate well with IFNalpha response in CML patients. PMID- 15287918 TI - Alemtuzumab-induced remission of both severe paraneoplastic pemphigus and leukaemic bone marrow infiltration in a case of treatment-resistant B-cell chronic lymphocytic leukaemia. AB - BACKGROUND: Alemtuzumab (MabCampath; ILEX Pharmaceuticals, Geneva, Switzerland) is a humanised monoclonal antibody directed against CD52. It belongs to a new group of monoclonal antibodies with anti-neoplastic effects used in chronic lymphocytic leukaemia (CLL) either as first-line treatment or in those cases resistant to alkylating drugs. Paraneoplastic pemphigus (PNP) is a severe mucocutaneus disease mostly associated with B-cell lymphoproliferative disorders. Independent of the course of the underlying malignancy, this disease is often resistant to conventional immunosuppressive treatment and may lead to death as a result of infectious complications. CASE PRESENTATION: We report a case where an ongoing long-term remission of PNP has been induced by alemtuzumab in a patient with an underlying B-CLL. A 68-yr-old male with a 4-yr history of B-CLL presented with a widespread blistering eruption on the extremities and trunk and a severe stomatitis. The diagnosis of PNP relied on the clinical, histological and direct immunofluorescence findings. Despite intensive treatment strategies with various immunosuppressive drugs and antibiotics, blisters continued to develop and the patient was deteriorating. When treated with alemtuzumab the mucocutaneous lesions healed almost completely within a few weeks and the patients' general condition improved significantly. After 12 wk of treatment with alemtuzumab, the CLL infiltration of the bone marrow previously quantified at 75-80% remitted completely. Twelve months later, the patient was still in remission with only a small residual ulceration on the lip and one on the penis. CONCLUSIONS: Based on this case report we recommend treatment with alemtuzumab to severe cases of PNP in CLL. However, further follow-up of this case is needed in order to assess the long-term effect of alemtuzumab treatment in PNP. PMID- 15287919 TI - Clonally related splenic marginal zone lymphoma and Hodgkin lymphoma with unmutated V gene rearrangements and a 15-yr time gap between diagnoses. AB - Hodgkin lymphoma (HL) can rarely occur during the course of B-cell non-Hodgkin lymphoma (B-NHL), where both the HL and NHL tumours have been reported to be clonally related in most of the few combination lymphomas so far investigated. We here investigated a case that developed HL 15 yr after being diagnosed with an indolent B-cell lymphoma, classified as a splenic marginal zone lymphoma (SMZL). Analysis of rearranged immunoglobulin genes in the SMZL clone and in single Hodgkin Reed-Sternberg cells revealed presence of identical V gene rearrangements, thus demonstrating a clonal relationship. In contrast to previously described B-NHL/HL combinations, in this case both types of tumour cells carried unmutated V gene rearrangements. We conclude that the HL evolved from an unmutated tumour precursor, either the SMZL clone itself or a common earlier precursor. PMID- 15287920 TI - Zoledronate-induced remission of acute panmyelosis with myelofibrosis. AB - Acute panmyelosis with myelofibrosis is a rare and aggressive form of acute myeloid leukemia. We describe a new case with a huge proliferation of megakaryocytes, blast cells and reticulin fibers. The patient was treated with zoledronate, a third-generation bisphosphonate, and a gradual recovery from pancytopenia was observed. A new bone marrow biopsy performed 4 months later showed a surprising disappearance of the leukemic infiltration. Ten months after the diagnosis, the patient is still in healthy condition. This may support the recently described anti-tumor activity of zoledronate. PMID- 15287921 TI - Cytogenetic and hematological spontaneous remission in a case of acute myelogenous leukemia. AB - Several cases of spontaneous remission (SR) interrupting the invariably progressive course of untreated acute myeloblastic leukemia (AML) have been reported so far. We shall add to this series the hematological and cytogenetic SR occurring in a 72-yr-old man affected by AML following myelodysplastic syndrome. At diagnosis cytogenetic analysis showed the 48, xy, del (6) (p22-pter), +13, +14 karyotype. Owing to a lobar pneumonia, the chemotherapy was deferred and a broad spectrum antibiotic therapy was established. Supportive care included red cells and platelet transfusions and low-dose corticosteroid. Two months later, after the pneumonia had completely disappeared, a complete remission, lasting about 5 months, was documented on bone marrow morphological and cytogenetical examination, although some degree of myeloid dysplasia persisted. Possible mechanisms of the various SRs described during the course of AML are discussed with a review of the literature. PMID- 15287922 TI - Anagrelide-induced visual hallucinations in a patient with essential thrombocythemia. PMID- 15287923 TI - Successful treatment of steroid resistant autoimmune thrombocytopenia associated with chronic lymphocytic leukemia with alemtuzumab. PMID- 15287924 TI - Beta-cell mass plasticity in type 2 diabetes. PMID- 15287925 TI - Changes in serum leptin concentrations in overweight Japanese men after exercise. AB - AIM: To investigate the link between serum leptin concentrations and exercise. DESIGN: Cross-sectional and longitudinal studies of an exercise intervention. SUBJECTS: 110 Japanese overweight men aged 32-59 years were recruited. At baseline, the average body mass index (BMI) was 28.5 +/- 2.5 kg/m2. From this group, we used data of 36 overweight men (BMI, 28.9 +/- 2.3) for a 1-year exercise programme. MEASUREMENTS: Leptin was measured at baseline and after 1 year. Fat distribution was evaluated by visceral fat (V) and subcutaneous fat (S) areas measured with computed tomography (CT) scanning at umbilical levels. Anthropometric parameters, aerobic exercise level, muscle strength and flexibility were also investigated at baseline and after 1 year. RESULTS: In the first analysis, using cross-sectional data, leptin was significantly correlated with total body fat (r = 0.760, p < 0.01), V (r = 0.383, p < 0.01) and S (r = 0.617, p < 0.01) areas. In the second analysis, using longitudinal data, leptin was significantly reduced after 1 year (pre 6.7 +/- 4.0 ng/ml vs. post 5.1 +/- 3.1 ng/ml, p < 0.01). Results showed that steps per day were increased, and aerobic exercise level, weight-bearing index (WBI) and insulin resistance were significantly improved. Although, there was a positive correlation between Delta leptin(positive changes in leptin after 1 year) and anthropometric measurements such as Delta body weight, Delta BMI and Delta body fat, leptin/body weight, leptin/BMI and leptin/body fat ratios were significantly reduced during exercise intervention. CONCLUSION: The present study indicated exercise significantly lowers serum leptin concentrations, and thus it may improve the leptin resistance observed in overweight Japanese men. PMID- 15287926 TI - Lipids and lipoprotein(a) concentrations in Pakistani patients with type 2 diabetes mellitus. AB - AIM: The aim of the present study was to analyze serum lipoprotein(a) [Lp(a)] levels in Pakistani patients with type 2 diabetes mellitus (DM) and to find correlations between clinical characteristics and dyslipidaemias in these patients. METHODS: Fasting blood samples were analyzed for Lp(a), total cholesterol, triglycerides, low-density lipoprotein cholesterol (LDL-c), high density lipoprotein cholesterol (HDL-c), glucose and glycosylated haemoglobin (HbA1c) in 68 Pakistani patients with type 2 DM and 40 non-diabetic healthy control subjects. RESULTS: Lp(a) levels were significantly raised in diabetics as compared to the control group. No correlation of Lp(a) was seen with age, body mass index (BMI), systolic blood pressure (SBP), diastolic blood pressure (DBP) and fasting glucose. There was a positive correlation of BMI to SBP and DBP. There was a significant positive correlation between Lp(a) and total cholesterol and LDL-c. No correlation of Lp(a) was observed with HDL-c, triglycerides and glycosylated haemoglobin (HbA1c). CONCLUSION: The present study led us to conclude that serum Lp(a) levels are significantly raised in type 2 DM and have a positive correlation with serum total and LDL-c levels. PMID- 15287927 TI - Nateglinide alone or with metformin safely improves glycaemia to target in patients up to an age of 84. AB - AIM: To assess the effect of nateglinide on efficacy [fasting plasma glucose (FPG), postprandial plasma glucose (PPG) plasma glucose and HbA1c], tolerability and safety in patients with type 2 diabetes mellitus (T2Dm) on diet alone or on metformin in subjects up to an age of 84. METHODS: In an open-labelled 12-week, parallel study of 358 patients, aged 35-84 years with T2Dm, nateglinide was given as either monotherapy in patients previously on diet alone or low-dose sulfonylureas, which required washout before the study (group 1), or as an addition therapy in patients on steady dose of metformin (group 2). Nateglinide 120 mg was given before main meals. HbA1c, FPG and PPG values were taken at the time of breakfast at the beginning and the end of the study. RESULTS: HbA1c fell by a mean of 0.83%, 95% confidence interval (CI) (-0.97, -0.69) (p < 0.001) in group 1, and 0.67%, 95% CI (-0.77, -0.58) (p < 0.001) in group 2. There was a significant improvement in PPG in group 1 by a mean reduction of -3.47 mmol/l, 95% CI (-4.08, -2.87) (p < 0.0001) and in group 2 of -2.41 mmol/l, 95% CI (-2.84, -1.99) (p < 0.0001). There was an improvement in FPG of -1.2 mmol/l, 95% CI ( 1.49, -0.81) (p < 0.0001) and -0.8 mmol/l, 95% CI -(1.07, -0.53) (p < 0.0001) in group 1 and 2 respectively. 44% of patients in group 1 and 34% in group 2 achieved target of HbA1c < 7.0 and 66% in group 1 and 59% in group 2 achieved of HbA1c < 7.5%. Only one subject on nateglinide and metformin was withdrawn due to the side effect of hypoglycaemia. No patient required third-party assistance nor was admitted to hospital due to hypoglycaemia. CONCLUSION: These data demonstrate that nateglinide is a safe and effective agent in treatment to target in patients with T2Dm up to an age of 84 years. PMID- 15287928 TI - Characteristics of US adults with the metabolic syndrome and therapeutic implications. AB - BACKGROUND: The third Adult Treatment Panel (ATP III) of the National Cholesterol Education Program defines clinical criteria for diagnosis of the metabolic syndrome, which increases cardiovascular risk and is a target for therapy. AIM: We analysed the third National Health and Nutrition Examination Survey (NHANES III; 1988-94) to determine how many US adults meet these criteria and are recommended for lipid-modifying drug therapy by ATP III. METHODS: NHANES III data were used to estimate the number of individuals with the metabolic syndrome and the number recommended for treatment by ATP III, based on 1990 census data. RESULTS: An estimated 36.3 million (23%) US adults have the metabolic syndrome. Of these, 84% met the criterion for obesity, 76% for blood pressure, 75% for HDL C, 74% for triglycerides and 41% for glucose. Most (54%) are in the higher risk categories of ATP III, yet only 39% overall are recommended for drug therapy by ATP III cutpoints; of these, most will achieve LDL-C targets with reductions of 35-40%. Of the 15.3 million individuals with the metabolic syndrome and triglycerides > or = 2.26 mmol/l (200 mg/dl), non-HDL-C is above ATP III recommendations in 11.6 million. CONCLUSIONS: Of the large number of Americans with the metabolic syndrome, ATP III recommends drug therapy for only a minority, because LDL-C typically is not substantially elevated. Instead, high triglycerides and low HDL-C are more common; clinical trial data are needed to determine whether optimal therapy should focus on reductions in LDL-C or on comprehensive improvements to the lipid profile. PMID- 15287929 TI - Outcomes of initiation of therapy with once-daily combination of a thiazolidinedione and a biguanide at an early stage of type 2 diabetes. AB - CONTEXT: Utilization of the biguanide metformin and a thiazolidinedione (TZD) with new onset diabetes has the benefit of lowering A1cs into the normal range without the problem of severe hypoglycaemia. OBJECTIVE: To assess the effectiveness of once-daily combined metformin and TZD therapy compared with other therapeutic regimens typically utilized at later stages of type 2 diabetes. METHODS: A random chart review of 300 type 2 diabetic patients and extraction of data for body mass index (BMI), duration of diabetes and C-peptide and A1c was performed. In the 210 type 2 diabetic subjects in whom this information was currently available, the data were analysed. RESULTS: Eighty-six patients on once daily metformin and rosiglitazone had an average A1c of 6.2% (group A), and 58 subjects on triple therapy (metformin, rosiglitazone and glimepiride) (group B) had an average haemoglobin A1c (HbA1c) of 6.9%. The 22 subjects on one injection of insulin per day in addition to triple therapy (group C) had an average HbA1c of 7.6%, and the 44 subjects on more than one insulin injection per day plus metformin and/or rosiglitazone (group D) had an average HbA1c of 8.3%. HbA1cs below 7.0% were found in 91.9% of group A, 21.7% of group B, 36.4% of group C and 56.8% of group D. HbA1cs below 6.5% were found in 78.2% in group A, 15.5% in group B, 22.7% in group C and 31.8% in group D. HbA1cs below 6.0% were found in 41.9% in group A, 6.9% in group B, 9.1% in group C and 13.6% in group D. On univariate analyses, the HbA1c was positively associated with the duration of diabetes and the BMI and negatively associated with random C-peptide levels. Alternatively, on multiple regression analysis, there was no statistical correlation between the duration of diabetes or BMI with the HbA1c. However, there was a strong statistical correlation between the random C-peptide level and the HbA1c (p = 0.002). CONCLUSION: Early initiation of therapy for type 2 diabetes with a once-daily combination of metformin and rosiglitazone provides the greatest opportunity to achieve A1cs within the normal range. The level of achieved glycaemic control is not dependent on the number or potency of the therapies utilized but is dependent on the level of endogenous insulin production. The use of a TZD as part of initial therapy of type 2 diabetes with its documented ability to preserve or improve beta-cell function has the potential to achieve prolonged normoglycaemia in the type 2 diabetic patient. PMID- 15287930 TI - Atrial fibrillation and its association with type 2 diabetes and hypertension in a Swedish community. AB - AIM: To explore the prevalence of atrial fibrillation in patients with hypertension and type 2 diabetes and to identify possible mechanisms for the development of atrial fibrillation. METHODS: A community-based, cross-sectional observational study was conducted in the primary health care in Skara, Sweden, and 1739 subjects (798 men, 941 women) were surveyed. Patients were categorized as those with hypertension only (n = 597); those with both hypertension and type 2 diabetes (n = 171), and those with type 2 diabetes only (n = 147). In the reference population, 824 normotensive subjects without diabetes were identified and used as controls. Participants were examined for cardiovascular risk factors including fasting blood glucose, serum insulin, blood pressure, lipids and anthropometric measures. Resting electrocardiogram (ECG) was recorded and Minnesota-coded. Insulin resistance was measured by the homeostasis model assessment (HOMA). RESULTS: Age-adjusted prevalence of atrial fibrillation was 2% in patients with hypertension only, 6% in patients with both hypertension and type 2 diabetes, 4% in patients with type 2 diabetes only and 2% in controls, respectively. Age and sex adjusted odds ratios (OR) (95% CI) were; hypertension 0.7 (0.30-1.5), combined hypertension and type 2 diabetes 3.3 (1.6-6.7), and type 2 diabetes 2.0 (0.9-4.7). The association with combined hypertension and type 2 diabetes remained significant when adjusted for cardiovascular disease (CVD) risk factors and body mass index (BMI), was attenuated with adjustment for ischemic ECG; 2.4 (1.1-5.0) and lost significance with adjustment for insulin resistance; 1.3 (0.5-3.1). CONCLUSIONS: Atrial fibrillation is associated with the combined occurrence of type 2 diabetes and hypertension. Insulin resistance may be a common underlying mechanism. PMID- 15287931 TI - Effects of orlistat on obesity-related diseases - a six-month randomized trial. AB - AIM: To assess the effect of orlistat on body weight and concomitant diseases in patients with body mass index (BMI) of > 28 kg/m2 and poorly controlled type 2 diabetes, hypertension or hypercholesterolaemia. METHODS: This trial was a six month, randomized, double-blind, placebo-controlled study of orlistat 120 mg three times daily plus a mildly reduced-calorie diet. 1004 obese patients (BMI 28 40 kg/m2) were included by 253 private endocrinologists and received orlistat (n = 499) or placebo (n = 505). Patients were stratified by concomitant disorder (type 2 diabetes, n = 193; hypertension, n = 614; hypercholesterolaemia, n = 197). Body weight, anthropometry, lipid and glycaemic control parameters and blood pressure. RESULTS: After six months, orlistat produced a significantly greater weight loss than placebo in type 2 diabetes (-4.2% vs. -1.4%), hypertension (-6.2% vs. -1.9%) and hypercholesterolaemia (-5.5% vs. -2.3%) groups (p < 0.0001 for all). There was a greater decrease in HbA(1c) in the type 2 diabetes group (-0.54 vs. -0.18%; p = 0.002) and low-density lipoprotein (LDL) cholesterol in the hypercholesterolaemia group (-11.7% vs. -4.5%; p = 0.004) with orlistat vs. placebo. Early weight loss (> or = 5% at 12 weeks) was associated with the highest weight loss in each group, and the highest decreases in HbA1c, LDL-cholesterol and diastolic blood pressure in patients with type 2 diabetes, hypercholesterolaemia and hypertension, respectively, at six months. The incidence of adverse events was similar for orlistat and placebo, except for certain generally well-tolerated gastrointestinal events that were more common with orlistat. CONCLUSION: Orlistat plus a mildly reduced-calorie diet produced clinically meaningful weight loss and improvements in risk factors in overweight and obese patients with poorly controlled type 2 diabetes, hypertension or hypercholesterolaemia. PMID- 15287932 TI - Acarbose lowers serum triglyceride and postprandial chylomicron levels in type 2 diabetes. AB - AIM: This study was designed to examine the therapeutic effect of acarbose on serum triglyceride (TG), free fatty acid (FFA), very low-density lipoprotein (VLDL) and chylomicron (CM) in the meal tolerance test (MTT) before and after acarbose treatment in type 2 diabetes mellitus (DM2). METHODS: Effects of acarbose on postprandial lipid metabolism were examined in DM2 patients. The subjects with normotriglyceridaemia (TG > or = 1.7 mmol/l, n = 60) were divided to three groups (A, B and C), and DM2 patients with hypertriglyceridaemia (TG > 1.7 mmol/l, n = 20) were designated group D. Group A was a control, and group B was designed to examine the one-dose effect of acarbose (100 mg) on lipid levels in MTT using the balanced food of 400 kcal. In groups C and D, acarbose 300 mg/day was administered for 8 weeks, and MTT with the one-dose acarbose administration was performed. We determined the levels of fasting and postprandial levels of glucose, insulin, FFA and TG-rich lipoproteins such as CM and VLDL. RESULTS: Acarbose treatment lowered plasma glucose levels and insulin secretion. In comparison among study groups A, B and C, acarbose significantly lowered serum TG levels in postprandial state. In group D, after the 8-week acarbose administration, fasting or postprandial FFA, TG and VLDL levels were also lowered. Interestingly, postprandial increase in CM was suppressed by acarbose administration in group B, C or D. CONCLUSIONS: Acarbose lowers postprandial TG and CM levels in DM2 with either normotriglyceridaemia or hypertriglyceridaemia. Improvement of insulin resistance with acarbose may also reduce fasting TG levels in DM2 with hypertriglyceridaemia. Acarbose is a beneficial therapeutic agent to reduce TG levels in DM2 patients, thereby leading to suppression of cardiovascular events. PMID- 15287933 TI - The diagnostic value of determining the hydroxy metabolite of glimepiride (M1) in blood serum in cases of severe hypoglycaemia associated with glimepiride therapy. PMID- 15287935 TI - Cerebral mucormycosis in a patient with myelodysplastic syndrome taking corticosteroids. PMID- 15287936 TI - Tracing the origin of cytogenetic anomalies in acute myeloid leukaemia following multiple myeloma. PMID- 15287937 TI - Hormones and pregnancy: thromboembolic risks for women. AB - During their lifetimes, women face several unique situations with an increased risk of venous thromboembolism (VTE). Doctors in a variety of specialties must advise women on the risks of oral contraceptives (OC), hormone replacement or pregnancy. Modern 'low dose' OC are associated with a three to sixfold increased relative risk of VTE. Hormone replacement and selective oestrogen receptor modulators confer a similar two to fourfold increase in thrombotic risk. However, because the baseline incidence of thrombosis is higher in older postmenopausal women, the absolute risk is higher than in younger OC users. The risk of venous thrombosis is six to 10-fold higher during pregnancy than in non-pregnant women of similar age. Thrombophilic disorders increase the thrombotic risk of OC, hormone replacement and pregnancy, especially in women with homozygous or combined defects. This review focuses on recent data estimating the thrombotic risk of hormonal therapies and pregnancy in women with and without other thrombotic risk factors. PMID- 15287938 TI - Guidelines for the diagnosis and management of hereditary spherocytosis. AB - Hereditary spherocytosis (HS) is a heterogeneous group of disorders with regard to clinical severity, protein defects and mode of inheritance. It is relatively common in Caucasian populations; most affected individuals have mild or only moderate haemolysis. There is usually a family history, and a typical clinical and laboratory picture so that the diagnosis is often easily made without additional laboratory tests. Atypical cases may require measurement of erythrocyte membrane proteins to clarify the nature of the membrane disorder and in the absence of a family history, occasionally molecular genetic analysis will help to determine whether inheritance is recessive or non-dominant. It is particularly important to rule out stomatocytosis where splenectomy is contraindicated because of the thrombotic risk. Mild HS can be managed without folate supplements and does not require splenectomy. Moderately and severely affected individuals are likely to benefit from splenectomy, which should be performed after the age of 6 years and with appropriate counselling about the infection risk. In all cases careful dialogue between doctor, patient and the family is essential. Laparoscopic surgery, when performed by experienced surgeons, can result in a shorter hospital stay and less pain. PMID- 15287939 TI - Impaired osteoblastogenesis in myeloma bone disease: role of upregulated apoptosis by cytokines and malignant plasma cells. AB - Bone remodelling is severely affected in myeloma bone disease as a consequence of skeletal metastatization of malignant plasma cells. We investigated whether defective bone replacement is dependent on increased osteoblast apoptosis and/or on deregulated events within the bone microenvironment. Circulating tumour necrosis factor (TNF)-alpha, interferon-gamma, interleukin (IL)-1beta, and IL-6 levels were higher in myeloma patients with overt bone disease, whose osteoblasts constitutively overexpressed Fas, DR4/DR5 complex as receptors to TNF-related apoptosis inducing ligand, intercellular adhesion molecule-1 (ICAM-1), and monocyte chemotactic protein-1 (MCP-1). They were functionally exhausted and promptly underwent apoptosis in vitro, in contrast to the minor tendency to death detected in control osteoblasts from patients without bone involvement and normal donors. Osteoblasts dramatically enhanced their apoptosis in co-cultures with MCC 2 myeloma cells and upregulated both ICAM-1 and MCP-1 in a manner similar to control osteoblasts. Pretreating MCC-2 cells with soluble ICAM-1 led to a striking inhibition of their adhesion to osteoblasts, suggesting that the ICAM 1/lymphocyte function-associated antigen-1 system plays a role in the reciprocal membrane contact to trigger apoptogenic signals. Our data suggest that, in the myeloma bone microenvironment, both high cytokine levels and physical interaction of malignant plasma cells with osteoblasts drive the accelerated apoptosis in these cells leading to defective new bone formation. PMID- 15287940 TI - High frequencies of chromosomal aberrations in multiple myeloma and monoclonal gammopathy of undetermined significance in direct chromosome preparation. AB - Although many cases of multiple myeloma (MM) and monoclonal gammopathy of undetermined significance (MGUS) are cytogenetically normal, interphase fluorescence in situ hybridization (FISH) analyses reveal aberrations in the majority of the cases. Most likely, non-neoplastic cells are more prone to divide in culture than neoplastic cells. Direct chromosome preparations (DCP) would be one way to circumvent this methodological problem. We have investigated 47 bone marrow samples from 39 patients by DCP. A median of 58 metaphases (range 9-158) was analysed per sample. Interphase FISH analyses using probes to detect IGH rearrangements, -13/13q-, +3, +7, and +11 were also performed. Abnormal karyotypes were detected in 15 (63%) of 24 MM and in 4 (50%) of eight MGUS/smouldering MM (SMM) cases that could be successfully cytogenetically analysed. Age, sex, or degree of bone marrow plasma cell (PC) infiltration did not influence the karyotypic patterns (P > 0.05). However, the frequencies of aberrant karyotypes varied in relation to the Colcemide concentrations used - 7% (30 ng/ml) versus 69% and 67% (100 and 200 ng/ml, respectively) (P = 0.01). Combining the G-banding and FISH results, abnormalities were detected in 29 of 31 (94%) MM and in six of eight (75%) MGUS/SMM patients. Thus, cytogenetic and FISH analyses after DCP using 100-200 ng Colcemide/ml identified aberrations in most MM/MGUS/SMM, irrespective of PC percentages. PMID- 15287941 TI - Translocations involving 6p22 in acute myeloid leukaemia at relapse: breakpoint characterization using microarray-based comparative genomic hybridization. AB - The detection of chromosomal aberrations is essential for the diagnosis and therapy of acute myeloid leukaemia (AML). We report two cases of de novo AML with translocations involving the breakpoint 6p22 first detected at relapse. Chromosomes were identified by conventional and molecular cytogenetics. At diagnosis, one patient presented a normal karyotype and the other one a trisomy 11 and a del(7)(q31q36). In the first case, cytogenetic analyses at relapse revealed a t(3;6)(q21;p22). The second patient showed a t(1;6)(q21;p22) at relapse. Detailed characterization of the breakpoints on the short arm of chromosome 6 was performed using array comparative genomic hybridization (CGH) on a platform specific for chromosome 6. In both cases, array CGH showed a terminal deletion and a small internal duplication of the short arm of chromosome 6. The region 6p22 is involved in several aberrations in tumours. Translocation partners are distributed throughout the human genome. We identified 3q21, a recurrent breakpoint in AML, for the first time as a translocation partner. The fragile site FRA6C, located in 6p22.2, and possibly the genes that reside within it, may play a role in tumorigenesis. The occurrence of translocations involving 6p22 after chemotherapy or radiation therapy suggests that one or more therapeutic agents might play a role in their origin. PMID- 15287942 TI - In patients with myelodysplastic syndromes response to rHuEPO and G-CSF treatment is related to an increase of cytogenetically normal CD34 cells. AB - The in vivo effect of recombinant human erythropoietin (rHuEpo) and granulocyte colony-stimulating factor (G-CSF) combined treatment on CD34(+) cells was evaluated by fluorescence in situ hybridization (FISH) in 13 myelodysplastic syndrome (MDS) patients with known cytogenetic abnormalities. After treatment, responsive patients presented a significantly lower proportion of FISH abnormal CD34(+) cells than before treatment (P = 0.003), and in comparison with unresponsive cases (P = 0.007). Response to treatment was associated with a reduced degree of apoptosis in CD34(+) cells (P = 0.021): however, no difference in telomere length was observed in responsive patients after growth factor administration. Although the number of patients analysed was relatively small, the present data suggest that, in MDS patients, response to rHuEpo and G-CSF may be related to the proliferation of karyotypically normal but potentially defective CD34(+) progenitor cells. PMID- 15287943 TI - Low expression of the putative tumour suppressor gene gravin in chronic myeloid leukaemia, myelodysplastic syndromes and acute myeloid leukaemia. AB - The putative tumour suppressor gene gravin is down-regulated in several solid tumours and is implicated in tumorigenesis. We have evaluated the expression levels of the gravin gene in the CD34(+)/blast cells of a range of myeloid malignancies as compared with controls using real-time quantitative polymerase chain reaction (PCR). Gravin was markedly down-regulated in 41 of 41 patients with acute myeloid leukaemia (AML), nine of 10 patients with myelodysplastic syndromes (MDS) and 33 of 33 patients with chronic myeloid leukaemia (CML), of whom 24 were in blast crisis (BC). We have shown that gravin is consistently down regulated in the CD34(+)/blast cells of myeloid malignancies and may play a role in the molecular pathogenesis of these disorders. PMID- 15287944 TI - Autologous cytomegalovirus-specific T cells as effector cells in immunotherapy of B cell chronic lymphocytic leukaemia. AB - B-cell chronic lymphocytic leukaemia (B-CLL) cells express low levels of co stimulatory molecules and therefore fail to induce activation and differentiation of tumour-specific T cells. We have shown that patients with B-CLL have considerably expanded numbers of cytomegalovirus (CMV) reactive CD8(+) T cells. This study demonstrated that B-CLL cells loaded with CMV peptide not only promoted the ex vivo expansion of autologous, in vivo-generated virus-specific T cells, but also constituted excellent target cells for these cytotoxic T cells, even without ex vivo re-stimulation. Directing virus-specific T cells to B-CLL may overcome the inadequate immunostimulatory capacity of these cells, which could be exploited for T-cell mediated immunotherapy. PMID- 15287945 TI - Tolerance associated with cord blood transplantation may depend on the state of host dendritic cells. AB - Allogeneic cord blood (CB) transplantation is associated with less severe graft versus-host disease (GvHD), thought to be due to the immaturity of CB T cells, but how T cells interact with host and donor-derived dendritic cells (DCs) to initiate GvHD has not been elucidated. We therefore investigated the responses of CB and adult blood CD4(+) T cells co-cultured with adult host DCs of different maturities. Primed by adult host DCs, CB and adult blood CD4(+) T cells underwent similar changes in the expression of CD45RA/45RO, CD25, CD40L and CTLA-4. However, CB CD4(+) T cells, when primed by either immature or Bacillus Calmette Guerin mycobacteria-treated adult host DCs, produced lower interferon-gamma (IFN gamma) and higher interleukin-10 (IL-10), which is a regulatory T cell-like cytokine profile, as compared with adult blood CD4(+) T cells. In contrast, lipopolysaccharide (LPS)-treated adult host DCs significantly up-regulated IFN gamma and down-regulated IL-10 production levels from CB CD4(+) T cells to that from adult blood CD4(+) T cells. The sustained low IFN-gamma and high IL-10 production from CB CD4(+) T cells co-cultured with adult blood DCs might account for the less severe GvHD occurrence after CB transplantation, which could be reversed by LPS-treated adult blood DCs. PMID- 15287946 TI - CD34 cell selection of peripheral blood progenitor cells using the CliniMACS device for allogeneic transplantation: clinical results in 102 patients. AB - The present study investigated the effects of CD34(+) cell selection in 102 patients using the CliniMACS device. Patients were at high risk for the development of graft versus host disease (GvHD) because of age, or the use of a haploidentical, mismatched or unrelated donor (UD). The median age of the patients was 44 years. The CliniMACS procedure yielded 8.0 x 10(6) CD34(+) cells/kg and the number of residual T cells was 1.3 x 10(4)/kg (median). The median follow up was 20.6 months. The probability of graft failure was 7%. The rate of acute GvHD was low (compatible family donors 10%, UDs 17%, and haploidentical donors 26%) with no patient enduring more than grade II disease. The cumulative incidence of chronic GvHD at the median follow up after transplant was 15% for the compatible family donor group, 40% for the UD group and 78% in the group transplanted from a haploidentical donor Treatment failure was mainly because of transplant-related mortality, especially aspergillus infection, and not due to relapse. The probability of disease-free survival, stratified for the risk of treatment failure, was 27% for the high risk, 46% for the intermediate risk and 83% for the low risk group. PMID- 15287947 TI - Improved haematopoietic recovery following transplantation with ex vivo-expanded mobilized blood cells. AB - Infusions of ex vivo-expanded (EXE) mobilized blood cells have been explored to enhance haematopoietic recovery following high dose chemotherapy (HDT). However, prior studies have not consistently demonstrated improvements in trilineage haematopoietic recovery. Three cohorts of three patients with breast cancer received three cycles of repetitive HDT supported by either unmanipulated (UM) and/or EXE cells. Efficacy was assessed by an internal comparison of each patient's consecutive HDT cycles, and to 106 historical UM infusions. Twenty-one cycles were supported by EXE cells and six by UM cells alone. Infusions of EXE cells resulted in fewer days with an absolute neutrophil count (ANC) <0.1 x 10(9)/l (median 2 vs. 4 d, P = 0.002) and 3 d faster ANC recovery to >0.1 x 10(9)/l (median 5 vs. 8 d, P = 0.0002). This resulted in a major reduction in the incidence of febrile neutropenia compared with UM cycles (0% vs. 83%; P = 0.008) and in 66% of historical UM cycles (P = 0.01) and a marked reduction in hospital re-admission. There were also fewer platelet transfusions required (43% vs. 100%; P = 0.009). We conclude that EXE cells enhance both neutrophil and platelet recovery and reduce febrile neutropenia, platelet transfusion and hospital re admission. PMID- 15287948 TI - Compound heterozygous mutations in the gamma-glutamyl carboxylase gene cause combined deficiency of all vitamin K-dependent blood coagulation factors. AB - Hereditary combined deficiency of the vitamin K-dependent coagulation factors II, VII, IX, X, protein C, S and protein Z (VKCFD) is a very rare autosomal recessive inherited bleeding disorder. The phenotype may result from functional deficiency of either the gamma-glutamyl carboxylase (GGCX) or the vitamin K epoxide reductase (VKOR) complex. We report on the third case of VKCFD1 with mutations in the gamma-glutamyl carboxylase gene, which is remarkable because of compound heterozygosity. Two mutations were identified: a splice site mutation of exon 3 and a point mutation in exon 11, resulting in the replacement of arginine 485 by proline. Screening of 100 unrelated normal chromosomes by restriction fragment length polymorphism and denaturing high-performance liquid chromatography analysis excluded either mutation as a frequent polymorphism. Substitution of vitamin K could only partially normalize the levels of coagulation factors. It is suggested that the missense mutation affects either the propeptide binding site or the vitamin K binding site of GGCX. PMID- 15287949 TI - Activated protein C levels in Behcet's disease and risk of venous thrombosis. AB - Behcet's disease is a multi-systemic inflammatory disorder of unknown cause. Most abnormalities have been associated with endothelial injury caused by vasculitis. Thrombosis occurs in about 25% of patients, although the mechanism is unknown. The objective of this study was to evaluate the protein C activation system in Behcet's disease and its correlation with venous thromboembolism (VTE). Thirty nine patients (12 with VTE) and 78 age- and sex-matched controls were included in the study, and levels of protein C, protein S, activated protein C (APC), protein C inhibitor (PCI), soluble thrombomodulin (TM), antithrombin (AT), alpha(1) antitrypsin, fibrinogen, factor VIII, von Willebrand factor (VWF) and C-reactive protein (CRP) were measured. APC and TM levels were significantly lower in patients than in controls, whereas protein S, AT, alpha(1)-antitrypsin, fibrinogen, factor VIII, VWF and CRP levels were significantly higher in patients than in controls. APC, PCI and TM levels were lower in patients with VTE (0.65 +/ 0.19 ng/ml, 86% +/- 22% and 15.5 +/- 7.1 ng/ml respectively) than in those without VTE (0.78 +/- 0.17 ng/ml, 100% +/- 15% and 22.1 +/- 15.3 ng/ml) (P < 0.05). In patients, APC levels below 0.75 ng/ml (10th percentile of the control group) increased the risk of VTE about fivefold (odds ratio = 5.1; 95% confidence interval = 1.1-23.4). These results show that reduced APC levels are associated with the high incidence of VTE in Behcet's disease. PMID- 15287950 TI - The value of education and self-monitoring in the management of warfarin therapy in older patients with unstable control of anticoagulation. AB - Of 125 patients aged 65 years or over, with atrial fibrillation taking warfarin for at least 12 months, with a standard deviation (SD) of prothrombin time, expressed as the International Normalized Ratio (INR) >0.5 over the previous 6 months, 40 were randomized to continue with usual clinic care and 85 to receive education about warfarin. Of these, 44 were randomized to self-monitor their INR and 41 returned to clinic. Compared with the previous 6 months there was a significant increase in percentage time within the therapeutic range for the 6 months following education [61.1 vs. 70.4; mean difference 8.8; 95% confidence interval (CI): -0.2-17.8; P = 0.054] and following education and self-monitoring (57 vs. 71.1; mean difference 14.1; 95% CI: 6.7-21.5; P < 0.001), compared with those patients following usual clinic care (60.0 vs. 63.2; mean difference 3.2; 95% CI: -7.3-13.7). Using the same comparative periods, the INR SD fell by 0.24 (P < 0.0001) in the group allocated to education and self-monitoring, 0.26 (P < 0.0001) in the group receiving education alone and 0.16 (P = 0.003) in the control group. Inter-group differences were not statistically significant (intervention groups 0.26 +/- 0.30 vs. control 0.16 +/- 0.3, P = 0.10). Quality of-life measurements and health beliefs about warfarin were unchanged (apart from emotional role limitation) with education or education and self-monitoring. Patient education regarding anticoagulation therapy could be a cost-effective initiative and is worthy of further study. PMID- 15287951 TI - Effects of genetic fusion of factor IX to albumin on in vivo clearance in mice and rabbits. AB - Individuals with haemophilia B require replacement therapy with recombinant or plasma-derived coagulation factor IX (fIX). More benefit per injected dose might be obtained if fIX clearance could be slowed. The contribution of overall size to fIX clearance was explored, using genetic fusion to albumin. Recombinant murine fIX (MIX), and three proteins with C-terminal epitope tags were expressed in HEK 293 cells: tagged MIX (MIXT), tagged mouse serum albumin (MSAT) and MFUST, in which MIX and MSAT were fused in a single polypeptide chain. Proteins MFUST and MIXT were two- to threefold less active in clotting assays than MIX. In mice, the area under the clearance curve (AUC) was reduced for MFUST compared with MSAT or plasma-derived MSA (pd-MSA); the terminal catabolic half-life (t(0.5)) did not differ amongst the three proteins. Two minutes after injection, >40% of the injected MFUST was found in the liver, compared with <10% of either MSAT or pd MSA. In rabbits, the AUC for MFUST was reduced compared to MIXT, MSAT, or pd-MSA, while the t(0.5) of the fusion protein fell between that of MIXT and MSAT or pd MSA. Similar results were obtained with non-radioactive fused or non-fused recombinant human fIX in fIX knockout mice. The clearance behaviour of the fusion protein thus more closely resembled that of fIX than that of albumin despite a modest increase in terminal half-life, suggesting that fIX-specific interactions that are important in determining clearance were maintained in spite of the increased size of the fusion protein. PMID- 15287952 TI - Novel 2'-substituted, 3'-deoxy-phosphatidyl-myo-inositol analogues reduce drug resistance in human leukaemia cell lines with an activated phosphoinositide 3 kinase/Akt pathway. AB - Activation of the phosphoinositide 3-kinase (PI3-K)/Akt signalling pathway has been linked with resistance to chemotherapeutic drugs, and its down-regulation, by means of pharmacological inhibitors of PI3-K, considerably lowers resistance to various types of therapy in cell lines derived from solid tumours. Recently, a new class of Akt inhibitors, referred to as phosphatidylinositol ether lipids (PIAs), have been synthesized. We tested whether two new PIAs could lower the sensitivity threshold to chemotherapeutic drugs of human leukaemia cell lines with an activated PI3-K/Akt network. We used HL60AR (for apoptosis resistant), K562 and U937 cells. The two pharmacological inhibitors, used at 5 micromol/l, down-regulated Akt kinase activity and phosphorylation. Neither of the two chemicals affected the activity of other signalling proteins in the Akt pathway, such as phosphoinositide-dependent protein kinase-1 or PTEN. When employed at 5 micromol/l, the Akt inhibitors markedly reduced the resistance of the leukaemic cell lines to etoposide or cytarabine. Remarkably, a 5 micromol/l concentration of the inhibitors did not negatively affect the survival rate of human cord blood CD34(+) cells. Overall, our results indicate that new selective Akt pharmacological inhibitors might be used in the future for overcoming Akt mediated resistance to therapeutic treatments of acute leukaemia cells. PMID- 15287953 TI - Human gammadelta T cells as mediators of chimaeric-receptor redirected anti tumour immunity. AB - Human peripheral blood gammadelta T cells (Vgamma9(+) Vdelta2(+)) can be selectively expanded in vivo by the systemic administration of aminobisphosphonates without prior antigen priming. To assess the potential of human gammadelta T cells to serve as effector cells of specific anti-tumour immunity, we expanded peripheral blood-derived gammadelta T cells and transduced them with recombinant retrovirus encoding G(D2)- or CD19-specific chimaeric receptors. Flow cytometric analysis of T cells from four individual donors cultured in the presence of zoledronate at day 14 of culture showed selective enrichment of the gammadelta T cell population (Vgamma9(+) Vdelta2(+) CD3(+) CD4( ) CD8(-)) to 73-96% of total CD3(+) T cells. Retroviral gene transfer resulted in chimaeric receptor surface expression in 73 +/- 12% of the population. Transduced gammadelta T cells efficiently recognized antigen-expressing tumour cell targets, as demonstrated by target-specific upregulation of CD69 and secretion of interferon-alpha. Moreover, transduced gammadelta T cells efficiently and specifically lysed the antigen-expressing tumour targets. They could be efficiently expanded in vitro and maintained in culture for prolonged periods. Zoledronate-activated human gammadelta T cells expressing chimaeric receptors may thus serve as potent and specific anti-tumour effector cells. Their responsiveness to stimulation with aminobisphosphonates may enable the selective re-expansion of adoptively transferred T cells in vivo, permitting long lasting anti-tumour immune control. PMID- 15287954 TI - Dendritic cells fused with core binding factor-beta positive acute myeloid leukaemia blast cells induce activation of cytotoxic lymphocytes. AB - Several reports have described various strategies of dendritic cell (DC) vaccination to induce specific T-cell responses in patients with acute myeloid leukaemia (AML). About 50-60% of AML cases blasts have chromosomal abnormalities, such as inv(16) or t(8,21), which could encode for leukaemia-specific antigenic peptide sequences, possibly presented in the context of self-major histocompatibility complex (MHC) molecules. As the co-culture of AML blasts with T lymphocytes seldom resulted in T-cell stimulation, we fused AML blasts with autologous DC to enhance this effect. The fusion cells expressed MHC class I and II, CD40, B7-1, B7-2, CD209 and several adhesion molecules. In a mixed lymphocyte hybrid reaction, the fusion cells induced the proliferation of autologous T cells. Moreover, in the special case of fusion cells established from AML blasts with the chromosomal abnormality inv(16), the autologous T lymphocytes could be primed to induce cytotoxicity against up to 70% autologous AML blasts in a effector:target ratio of 20:1. Blocking assays demonstrated that the lysis was chiefly mediated by CD8(+), CCR7(-) T lymphocytes, which could be further expanded in the form of effector memory CD8(+) T cells by repeated co-cultures with the autologous fusion cells. PMID- 15287955 TI - A single, multiplex analysis for all relevant activating NRAS gene mutations using heteroduplex generators. AB - We describe a multiplex polymerase chain reaction (PCR)-based test that detected all relevant NRAS activating mutations using a single PCR followed directly by electrophoresis. The test uses a Universal Heteroduplex Generator (UHG) to detect exon-2 (codon 61) NRAS mutations in multiplex with an UHG for exon-1 (codons 12 and 13). The method differentiated all 19 relevant mutations in these exons and showed a mutation independent sensitivity of approximately 6%. The sensitive, specific detection of all NRAS activating mutations using this single rapid test represents a minimum workload and could be applied readily for large-scale screening and for routine analysis. PMID- 15287956 TI - Are there clinical phenotypes of homozygous sickle cell disease? AB - The distribution of clinical features was examined in subjects with homozygous sickle cell (SS) disease in the Jamaican Cohort Study to determine whether there is evidence of distinct clustering of symptoms or clinical phenotypes. A twofold model yielded groups that could be interpreted as painful crisis or leg ulcer phenotypes and 78% of patients were classified with 95% confidence into one of these. The painful crisis phenotype also manifested higher frequencies of dactylitis, meningitis/septicaemia, acute chest syndrome and stroke. Attempts to define a three-group model were less convincing although 43% of patients could be allocated with 95% confidence. The three-group model essentially divided subjects with the leg ulcer phenotype into subgroups with higher and lower frequencies of painful crisis, dactylitis, meningitis/septicaemia and acute chest syndrome. In the three-group model, the painful crisis phenotype had lower total haemoglobin, fetal haemoglobin, mean cell volume and higher reticulocytes but there was no apparent influence of alpha thalassaemia or beta globin haplotype. Both environmental and genetic factors are likely to contribute to most manifestations of SS disease and the evidence for different clinical phenotypes suggests that a search for associated genetic polymorphisms may provide insights into the mechanisms of clinical variability in SS disease. PMID- 15287957 TI - Rapamycin-mediated induction of gamma-globin mRNA accumulation in human erythroid cells. AB - The present study aimed to determine whether rapamycin could increase the expression of gamma-globin genes in human erythroid cells. Rapamycin is a macrocyclic lactone that possesses immunosuppressive, antifungal and anti-tumour properties. This molecule is approved as an immunosuppressive agent for preventing rejection in patients receiving organ transplantation. To verify the activity of rapamycin, we employed two experimental cell systems, the human leukaemia K562 cell line and the two-phase liquid culture of human erythroid progenitors isolated from normal donors and patients with beta-thalassaemia. The results suggested that rapamycin, when compared with cytosine arabinoside, mithramycin and cisplatin, is a powerful inducer of erythroid differentiation and gamma-globin mRNA accumulation in human leukaemia K562 cells. In addition, when normal human erythroid precursors were cultured in the presence of rapamycin, gamma-globin mRNA accumulation and fetal haemoglobin (HbF) production increased to levels that were higher than those obtained using hydroxyurea. These effects were not associated with inhibition of cell growth. Furthermore, rapamycin was found to increase HbF content in erythroid precursor cells from four beta thalassaemia patients. These results could have practical relevance, because pharmacologically mediated regulation of the expression of human gamma-globin genes, leading to increased HbF, is considered a potential therapeutic approach in haematological disorders, including beta-thalassaemia and sickle cell anaemia. PMID- 15287958 TI - Immune thrombocytopenic purpura: investigation of the role of cytomegalovirus infection. PMID- 15287959 TI - Successful reduced-intensity allogeneic transplantation utilizing a fludarabine based preparative regimen in a patient with chronic lymphocytic leukaemia and a history of fludarabine-associated autoimmune haemolytic anaemia. PMID- 15287962 TI - Retrieving sequences of enzymes experimentally characterized but erroneously annotated : the case of the putrescine carbamoyltransferase. AB - BACKGROUND: Annotating genomes remains an hazardous task. Mistakes or gaps in such a complex process may occur when relevant knowledge is ignored, whether lost, forgotten or overlooked. This paper exemplifies an approach which could help to resuscitate such meaningful data. RESULTS: We show that a set of closely related sequences which have been annotated as ornithine carbamoyltransferases are actually putrescine carbamoyltransferases. This demonstration is based on the following points : (i) use of enzymatic data which had been overlooked, (ii) rediscovery of a short NH2-terminal sequence allowing to reannotate a wrongly annotated ornithine carbamoyltransferase as a putrescine carbamoyltransferase, (iii) identification of conserved motifs allowing to distinguish unambiguously between the two kinds of carbamoyltransferases, and (iv) comparative study of the gene context of these different sequences. CONCLUSIONS: We explain why this specific case of misannotation had not yet been described and draw attention to the fact that analogous instances must be rather frequent. We urge to be especially cautious when high sequence similarity is coupled with an apparent lack of biochemical information. Moreover, from the point of view of genome annotation, proteins which have been studied experimentally but are not correlated with sequence data in current databases qualify as "orphans", just as unassigned genomic open reading frames do. The strategy we used in this paper to bridge such gaps in knowledge could work whenever it is possible to collect a body of facts about experimental data, homology, unnoticed sequence data, and accurate informations about gene context. PMID- 15287963 TI - Function and evolution of the serotonin-synthetic bas-1 gene and other aromatic amino acid decarboxylase genes in Caenorhabditis. AB - BACKGROUND: Aromatic L-amino acid decarboxylase (AADC) enzymes catalyze the synthesis of biogenic amines, including the neurotransmitters serotonin and dopamine, throughout the animal kingdom. These neurotransmitters typically perform important functions in both the nervous system and other tissues, as illustrated by the debilitating conditions that arise from their deficiency. Studying the regulation and evolution of AADC genes is therefore desirable to further our understanding of how nervous systems function and evolve. RESULTS: In the nematode C. elegans, the bas-1 gene is required for both serotonin and dopamine synthesis, and maps genetically near two AADC-homologous sequences. We show by transformation rescue and sequencing of mutant alleles that bas-1 encodes an AADC enzyme. Expression of a reporter construct in transgenics suggests that the bas-1 gene is expressed, as expected, in identified serotonergic and dopaminergic neurons. The bas-1 gene is one of six AADC-like sequences in the C. elegans genome, including a duplicate that is immediately downstream of the bas-1 gene. Some of the six AADC genes are quite similar to known serotonin- and dopamine-synthetic AADC's from other organisms whereas others are divergent, suggesting previously unidentified functions. In comparing the AADC genes of C. elegans with those of the congeneric C. briggsae, we find only four orthologous AADC genes in C. briggsae. Two C. elegans AADC genes - those most similar to bas 1 - are missing from C. briggsae. Phylogenetic analysis indicates that one or both of these bas-1-like genes were present in the common ancestor of C. elegans and C. briggsae, and were retained in the C. elegans line, but lost in the C. briggsae line. Further analysis of the two bas-1-like genes in C. elegans suggests that they are unlikely to encode functional enzymes, and may be expressed pseudogenes. CONCLUSIONS: The bas-1 gene of C. elegans encodes a serotonin- and dopamine-synthetic AADC enzyme. Two C. elegans AADC-homologous genes that are closely related to bas-1 are missing from the congeneric C. briggsae; one or more these genes was present in the common ancestor of C. elegans and C. briggsae. Despite their persistence in C. elegans, evidence suggests the bas-1-like genes do not encode functional AADC proteins. The presence of the genes in C. elegans raises questions about how many 'predicted genes' in sequenced genomes are functional, and how duplicate genes are retained or lost during evolution. This is another example of unexpected retention of duplicate genes in eukaryotic genomes. PMID- 15287964 TI - A drop in the bucket. AB - With 151 billion dollars, all world-wide AIDS programs could be fully funded for the next 12 years. PMID- 15287965 TI - Molecular tumor profiling: translating genomic insights into clinical advances. AB - Molecular profiling of the transcripts or proteins within an individual tumor may in future provide important prognostic and therapeutic clinical information both for the affected individual and for their extended family, but for the time being traditional genetics and pathology retain their place in the clinic. PMID- 15287966 TI - Chipping away at 'stemness'. AB - Global gene-expression analyses of human embryonic stem cells confirm the involvement of some known genes in stem-cell function and identify some new candidate regulators of stem-cell growth. Support remains elusive, however, for the concept of 'stemness'--a pattern of expression of genes that is common to all stem cells. PMID- 15287967 TI - The imbalanced supertree of flowering-plant phylogeny. AB - Two contrasting approaches have been used to construct the overall tree of life from molecular data: one involves the analysis of single large datasets, while the other involves joining many independent smaller analyses into a supertree. A recent study uses the latter approach to produce the most complete phylogeny yet of flowering plant families. PMID- 15287968 TI - Success for gene therapy: render unto Caesar that which is Caesar's. AB - Reports that two young children developed leukemia after being treated for immunodeficiency with their own retrovirally modified bone-marrow cells delivered a severe blow to confidence in gene therapy as a treatment. Two reports, published since the trial was initiated, now take away some of the mystery as to why these events happened and allay fears for the safety of gene therapy across all therapeutic applications. PMID- 15287969 TI - What makes us human? AB - The sequence of chimpanzee chromosome 22 is starting to help us to define the set of genetic attributes that are unique to humans, but interpreting the biological consequences of these remains a major challenge. PMID- 15287970 TI - Addressing the age-old question of old age. PMID- 15287971 TI - Bacterial comparative genomics. PMID- 15287972 TI - Recent advances in Drosophila genomics. PMID- 15287973 TI - Call for an enzyme genomics initiative. AB - I propose an Enzyme Genomics Initiative, the goal of which is to obtain at least one protein sequence for each enzyme that has previously been characterized biochemically. There are 1,437 enzyme activities for which Enzyme Commission (EC) numbers have been assigned but no sequence can be found in public protein sequence databases. PMID- 15287974 TI - Comprehensive de novo structure prediction in a systems-biology context for the archaea Halobacterium sp. NRC-1. AB - BACKGROUND: Large fractions of all fully sequenced genomes code for proteins of unknown function. Annotating these proteins of unknown function remains a critical bottleneck for systems biology and is crucial to understanding the biological relevance of genome-wide changes in mRNA and protein expression, protein-protein and protein-DNA interactions. The work reported here demonstrates that de novo structure prediction is now a viable option for providing general function information for many proteins of unknown function. RESULTS: We have used Rosetta de novo structure prediction to predict three-dimensional structures for 1,185 proteins and protein domains (<150 residues in length) found in Halobacterium NRC-1, a widely studied halophilic archaeon. Predicted structures were searched against the Protein Data Bank to identify fold similarities and extrapolate putative functions. They were analyzed in the context of a predicted association network composed of several sources of functional associations such as: predicted protein interactions, predicted operons, phylogenetic profile similarity and domain fusion. To illustrate this approach, we highlight three cases where our combined procedure has provided novel insights into our understanding of chemotaxis, possible prophage remnants in Halobacterium NRC-1 and archaeal transcriptional regulators. CONCLUSIONS: Simultaneous analysis of the association network, coordinated mRNA level changes in microarray experiments and genome-wide structure prediction has allowed us to glean significant biological insights into the roles of several Halobacterium NRC-1 proteins of previously unknown function, and significantly reduce the number of proteins encoded in the genome of this haloarchaeon for which no annotation is available. PMID- 15287975 TI - Phylogenetic profiling of the Arabidopsis thaliana proteome: what proteins distinguish plants from other organisms? AB - BACKGROUND: The availability of the complete genome sequence of Arabidopsis thaliana together with those of other organisms provides an opportunity to decipher the genetic factors that define plant form and function. To begin this task, we have classified the nuclear protein-coding genes of Arabidopsis thaliana on the basis of their pattern of sequence similarity to organisms across the three domains of life. RESULTS: We identified 3,848 Arabidopsis proteins that are likely to be found solely within the plant lineage. More than half of these plant specific proteins are of unknown function, emphasizing the general lack of knowledge of processes unique to plants. Plant-specific proteins that are membrane-associated and/or targeted to the mitochondria or chloroplasts are the most poorly characterized. Analyses of microarray data indicate that genes coding for plant-specific proteins, but not evolutionarily conserved proteins, are more likely to be expressed in an organ-specific manner. A large proportion (13%) of plant-specific proteins are transcription factors, whereas other basic cellular processes are under-represented, suggesting that evolution of plant-specific control of gene expression contributed to making plants different from other eukaryotes. CONCLUSIONS: We identified and characterized the Arabidopsis proteins that are most likely to be plant-specific. Our results provide a genome-wide assessment that supports the hypothesis that evolution of higher plant complexity and diversity is related to the evolution of regulatory mechanisms. Because proteins that are unique to the green plant lineage will not be studied in other model systems, they should be attractive priorities for future studies. PMID- 15287976 TI - System-based proteomic analysis of the interferon response in human liver cells. AB - BACKGROUND: Interferons (IFNs) play a critical role in the host antiviral defense and are an essential component of current therapies against hepatitis C virus (HCV), a major cause of liver disease worldwide. To examine liver-specific responses to IFN and begin to elucidate the mechanisms of IFN inhibition of virus replication, we performed a global quantitative proteomic analysis in a human hepatoma cell line (Huh7) in the presence and absence of IFN treatment using the isotope-coded affinity tag (ICAT) method and tandem mass spectrometry (MS/MS). RESULTS: In three subcellular fractions from the Huh7 cells treated with IFN (400 IU/ml, 16 h) or mock-treated, we identified more than 1,364 proteins at a threshold that corresponds to less than 5% false-positive error rate. Among these, 54 were induced by IFN and 24 were repressed by more than two-fold, respectively. These IFN-regulated proteins represented multiple cellular functions including antiviral defense, immune response, cell metabolism, signal transduction, cell growth and cellular organization. To analyze this proteomics dataset, we utilized several systems-biology data-mining tools, including Gene Ontology via the GoMiner program and the Cytoscape bioinformatics platform. CONCLUSIONS: Integration of the quantitative proteomics with global protein interaction data using the Cytoscape platform led to the identification of several novel and liver-specific key regulatory components of the IFN response, which may be important in regulating the interplay between HCV, interferon and the host response to virus infection. PMID- 15287977 TI - Origins of chromosomal rearrangement hotspots in the human genome: evidence from the AZFa deletion hotspots. AB - BACKGROUND: The origins of the recombination hotspots that are a common feature of both allelic and non-allelic homologous recombination in the human genome are poorly understood. We have investigated, by comparative sequencing, the evolution of two hotspots of non-allelic homologous recombination on the Y chromosome that lie within paralogous sequences known to sponsor deletions resulting in male infertility. RESULTS: These recombination hotspots are characterized by signatures of concerted evolution, which indicate that gene conversion between paralogs has been predominant in shaping their recent evolution. By contrast, the paralogous sequences that surround the hotspots exhibit little evidence of gene conversion. A second feature of these rearrangement hotspots is the extreme interspecific sequence divergence (around 2.5%) that places them among the most divergent orthologous sequences between humans and chimpanzees. CONCLUSIONS: Several hominid-specific gene conversion events have rendered these hotspots better substrates for chromosomal rearrangements in humans than in chimpanzees or gorillas. Monte Carlo simulations of sequence evolution suggest that extreme sequence divergence is a direct consequence of gene conversion between paralogs. We propose that the coincidence of signatures of concerted evolution and recurrent breakpoints of chromosomal rearrangement (mapped at the sequence level) may enable the identification of putative rearrangement hotspots from analysis of comparative sequences from great apes. PMID- 15287978 TI - Identifying combinatorial regulation of transcription factors and binding motifs. AB - BACKGROUND: Combinatorial interaction of transcription factors (TFs) is important for gene regulation. Although various genomic datasets are relevant to this issue, each dataset provides relatively weak evidence on its own. Developing methods that can integrate different sequence, expression and localization data have become important. RESULTS: Here we use a novel method that integrates chromatin immunoprecipitation (ChIP) data with microarray expression data and with combinatorial TF-motif analysis. We systematically identify combinations of transcription factors and of motifs. The various combinations of TFs involved multiple binding mechanisms. We reconstruct a new combinatorial regulatory map of the yeast cell cycle in which cell-cycle regulation can be drawn as a chain of extended TF modules. We find that the pairwise combination of a TF for an early cell-cycle phase and a TF for a later phase is often used to control gene expression at intermediate times. Thus the number of distinct times of gene expression is greater than the number of transcription factors. We also see that some TF modules control branch points (cell-cycle entry and exit), and in the presence of appropriate signals they can allow progress along alternative pathways. CONCLUSIONS: Combining different data sources can increase statistical power as demonstrated by detecting TF interactions and composite TF-binding motifs. The original picture of a chain of simple cell-cycle regulators can be extended to a chain of composite regulatory modules: different modules may share a common TF component in the same pathway or a TF component cross-talking to other pathways. PMID- 15287979 TI - Modular decomposition of protein-protein interaction networks. AB - We introduce an algorithmic method, termed modular decomposition, that defines the organization of protein-interaction networks as a hierarchy of nested modules. Modular decomposition derives the logical rules of how to combine proteins into the actual functional complexes by identifying groups of proteins acting as a single unit (sub-complexes) and those that can be alternatively exchanged in a set of similar complexes. The method is applied to experimental data on the pro-inflammatory tumor necrosis factor-alpha (TNF-alpha)/NFkappaB transcription factor pathway. PMID- 15287980 TI - POSaM: a fast, flexible, open-source, inkjet oligonucleotide synthesizer and microarrayer. AB - DNA arrays are valuable tools in molecular biology laboratories. Their rapid acceptance was aided by the release of plans for a pin-spotting microarrayer by researchers at Stanford. Inkjet microarraying is a flexible, complementary technique that allows the synthesis of arrays of any oligonucleotide sequences de novo. We describe here an open-source inkjet arrayer capable of rapidly producing sets of unique 9,800-feature arrays. PMID- 15287981 TI - Statistical modeling for selecting housekeeper genes. AB - There is a need for statistical methods to identify genes that have minimal variation in expression across a variety of experimental conditions. These 'housekeeper' genes are widely employed as controls for quantification of test genes using gel analysis and real-time RT-PCR. Using real-time quantitative RT PCR, we analyzed 80 primary breast tumors for variation in expression of six putative housekeeper genes (MRPL19 (mitochondrial ribosomal protein L19), PSMC4 (proteasome (prosome, macropain) 26S subunit, ATPase, 4), SF3A1 (splicing factor 3a, subunit 1, 120 kDa), PUM1 (pumilio homolog 1 (Drosophila)), ACTB (actin, beta) and GAPD (glyceraldehyde-3-phosphate dehydrogenase)). We present appropriate models for selecting the best housekeepers to normalize quantitative data within a given tissue type (for example, breast cancer) and across different types of tissue samples. PMID- 15287982 TI - Model selection and efficiency testing for normalization of cDNA microarray data. AB - In this study we present two novel normalization schemes for cDNA microarrays. They are based on iterative local regression and optimization of model parameters by generalized cross-validation. Permutation tests assessing the efficiency of normalization demonstrated that the proposed schemes have an improved ability to remove systematic errors and to reduce variability in microarray data. The analysis also reveals that without parameter optimization local regression is frequently insufficient to remove systematic errors in microarray data. PMID- 15287983 TI - Optimizing the HIV/AIDS informed consent process in India. AB - BACKGROUND: While the basic ethical issues regarding consent may be universal to all countries, the consent procedures required by international review boards which include detailed scientific and legal information, may not be optimal when administered within certain populations. The time and the technicalities of the process itself intimidate individuals in societies where literacy and awareness about medical and legal rights is low. METHODS: In this study, we examined pregnant women's understanding of group education and counseling (GEC) about HIV/AIDS provided within an antenatal clinic in Maharashtra, India. We then enhanced the GEC process with the use of culturally appropriate visual aids and assessed the subsequent changes in women's understanding of informed consent issues. RESULTS: We found the use of visual aids during group counseling sessions increased women's overall understanding of key issues regarding informed consent from 38% to 72%. Moreover, if these same visuals were reinforced during individual counseling, improvements in women's overall comprehension rose to 96%. CONCLUSIONS: This study demonstrates that complex constructs such as informed consent can be conveyed in populations with little education and within busy government hospital settings, and that the standard model may not be sufficient to ensure true informed consent. PMID- 15287984 TI - Who will lose weight? A reexamination of predictors of weight loss in women. AB - BACKGROUND: The purpose of this study was to analyze pretreatment predictors of short-term weight loss in Portuguese overweight and obese women involved in a weight management program. Behavioral and psychosocial predictors were selected a priori from previous results reported in American women who participated in a similar program. METHODS: Subjects were 140 healthy overweight/obese women (age, 38.3 +/- 5.9 y; BMI, 30.3 +/- 3.7 kg/m2) who participated in a 4-month lifestyle weight loss program consisting of group-based behavior therapy to improve diet and increase physical activity. At baseline, all women completed a comprehensive behavioral and psychosocial battery, in standardized conditions. RESULTS: Of all starting participants, 3.5% (5 subjects) did not finish the program. By treatment's end, more than half of all women had met the recomended weight loss goals, despite a large variability in individual results (range for weight loss = 19 kg). In bivariate and multivariate correlation/regression analysis fewer previous diets and weight outcome evaluations, and to a lesser extent self motivation and body image were significant and independent predictors of weight reduction, before and after adjustment for baseline weight. A negative and slightly curvilinear relationship best described the association between outcome evaluations and weight change, revealing that persons with very accepting evaluations (that would accept or be happy with minimal weight change) lost the least amount of weight while positive but moderate evaluations of outcomes (i.e., neither low nor extremely demanding) were more predictive of success. Among those subjects who reported having initiated more than 3-4 diets in the year before the study, very few were found to be in the most successful group after treatment. Quality of life, self-esteem, and exercise variables did not predict outcomes. CONCLUSIONS: Several variables were confirmed as predictors of success in short term weight loss and can be used in future hypothesis-testing studies and as a part of more evolved prediction models. Previous dieting, and pretreatment self motivation and body image are associated with subsequent weight loss, in agreement with earlier findings in previous samples. Weight outcome evaluations appear to display a more complex relationship with treatment results and culture specific factors may be useful in explaining this pattern of association. PMID- 15287985 TI - Identification of proteases employed by dendritic cells in the processing of protein purified derivative (PPD). AB - Dendritic cells (DC) are known to present exogenous protein Ag effectively to T cells. In this study we sought to identify the proteases that DC employ during antigen processing. The murine epidermal-derived DC line Xs52, when pulsed with PPD, optimally activated the PPD-reactive Th1 clone LNC.2F1 as well as the Th2 clone LNC.4k1, and this activation was completely blocked by chloroquine pretreatment. These results validate the capacity of XS52 DC to digest PPD into immunogenic peptides inducing antigen specific T cell immune responses. XS52 DC, as well as splenic DC and DCs derived from bone marrow degraded standard substrates for cathepsins B, C, D/E, H, J, and L, tryptase, and chymases, indicating that DC express a variety of protease activities. Treatment of XS52 DC with pepstatin A, an inhibitor of aspartic acid proteases, completely abrogated their capacity to present native PPD, but not trypsin-digested PPD fragments to Th1 and Th2 cell clones. Pepstatin A also inhibited cathepsin D/E activity selectively among the XS52 DC-associated protease activities. On the other hand, inhibitors of serine proteases (dichloroisocoumarin, DCI) or of cystein proteases (E-64) did not impair XS52 DC presentation of PPD, nor did they inhibit cathepsin D/E activity. Finally, all tested DC populations (XS52 DC, splenic DC, and bone marrow-derived DC) constitutively expressed cathepsin D mRNA. These results suggest that DC primarily employ cathepsin D (and perhaps E) to digest PPD into antigenic peptides. PMID- 15287986 TI - Autoimmune progesterone dermatitis in a patient with endometriosis: case report and review of the literature. AB - Autoimmune progesterone dermatitis (APD) is a condition in which the menstrual cycle is associated with a number of skin findings such as urticaria, eczema, angioedema, and others. In affected women, it occurs 3-10 days prior to the onset of menstrual flow, and resolves 2 days into menses. Women with irregular menses may not have this clear correlation, and therefore may be missed. We present a case of APD in a woman with irregular menses and urticaria/angioedema for over 20 years, who had not been diagnosed or correctly treated due to the variable timing of skin manifestations and menses. In addition, we review the medical literature in regards to clinical features, pathogenesis, diagnosis, and treatment options. PMID- 15287987 TI - Increased contractile responses to 5-Hydroxytryptamine and Angiotensin II in high fat diet fed rat thoracic aorta. AB - BACKGROUND: Feeding normal rats with high dietary levels of saturated fat leads to pathological conditions, which are quite similar to syndrome X in humans. These conditions such as hypertriglyceridemia, hypercholesterolemia, obesity, and hyperglycemia might induce hypertension through various mechanisms. Metabolic syndrome and the resulting NIDDM represent a major clinical challenge because implementation of treatment strategies is difficult. Vascular abnormalities probably contribute to the etiology of many diabetic complications including nephropathy, neuropathy, retinopathy, and cardiomyopathy. It has been shown that in Streptozotocin induced diabetic animals there is an increase in maximal responses to 5-Hydroxytryptamine and Angiotensin II. The purpose of this study was to evaluate High fat diet fed rats for the development of hypertriglyceridemia, hypercholesterolemia, hyperinsulinemia and hyperglycemia and to assess their vascular responses to 5-Hydroxytryptamine and Angiotensin II. METHODS: Male Sprague Dawley rats were used for this study and were divided into two equal groups. One of the groups was fed with normal pellet diet and they served as the control group, whereas the other group was on a high fat diet for 4 weeks. Body weight, plasma triglycerides, plasma cholesterol, and plasma glucose were measured every week. Intraperitoneal glucose tolerance test was performed after 4 weeks of feeding. At the end of fourth week of high fat diet feeding, thoracic aortae were removed, and cut into helical strips for vascular reactivity studies. Dose-response curves of 5-Hydroxytryptamine and Angiotensin II were obtained. RESULTS: There was no significant difference in pD2, with 5 Hydroxytryptamine and Angiotensin II in both groups but Emax was increased. CONCLUSIONS: These results suggest that hypertension in high fat diet rats is associated with increased in vitro vascular reactivity to 5-HT and Ang II. PMID- 15287989 TI - Lymphoedema management knowledge and practices among patients attending filariasis morbidity control clinics in Gampaha District, Sri Lanka. AB - BACKGROUND: Little information is available on methods of treatment practiced by patients affected by filarial lymphoedema in Sri Lanka. The frequency and duration of acute dematolymphangioadenitis (ADLA) attacks in these patients remain unclear. This study reports the knowledge, practices and perceptions regarding lymphoedema management and the burden of ADLA attacks among patients with lymphoedema. METHODS: A semi-structured questionnaire was used to assess morbidity alleviation knowledge, practices and perceptions. The burden of ADLA attacks was assessed using one-year recall data. RESULTS: 66 patients (22 males, 44 females) with mean age 51.18 years (SD +/- 13.9) were studied. Approximately two thirds of the patients were aware of the importance of skin and nail hygiene, limb elevation and use of footwear. Washing was practiced on a daily and twice daily basis by 40.9% and 48.5% respectively. However, limb elevation, exercise and use of footwear were practiced only by 21-42.4% (while seated and lying down), 6% and 34.8% respectively. The majority of patients considered regular intake of diethylcarbamazine citrate (DEC) important. Approximately two thirds (65.2%) had received health education from filariasis clinics. Among patients who sought private care (n = 48) the average cost of treatment for an ADLA attack was Rs. 737.91. Only 18.2% had feelings of isolation and reported community reactions ranging from sympathy to fear and ridicule. CONCLUSIONS: Filariasis morbidity control clinics play an essential role in the dissemination of morbidity control knowledge. Referral of lymphoedema patients to morbidity control clinics is recommended. PMID- 15287988 TI - Activity of telithromycin and comparators against bacterial pathogens isolated from 1,336 patients with clinically diagnosed acute sinusitis. AB - BACKGROUND: Increasing antimicrobial resistance among the key pathogens responsible for community-acquired respiratory tract infections has the potential to limit the effectiveness of antibiotics available to treat these infections. Since there are regional differences in the susceptibility patterns observed and treatment is frequently empirical, the selection of antibiotic therapy may be challenging. PROTEKT, a global, longitudinal multicentre surveillance study, tracks the activity of telithromycin and comparator antibacterial agents against key respiratory tract pathogens. METHODS: In this analysis, we examine the prevalence of antibacterial resistance in 1,336 bacterial pathogens, isolated from adult and paediatric patients clinically diagnosed with acute bacterial sinusitis (ABS). RESULTS AND DISCUSSION: In total, 58.0%, 66.1%, and 55.8% of S. pneumoniae isolates were susceptible to penicillin, cefuroxime, and clarithromycin respectively. Combined macrolide resistance and reduced susceptibility to penicillin was present in 200/640 (31.3 %) of S. pneumoniae isolates (128 isolates were resistant to penicillin [MIC > or = 2 mg/L], 72 intermediate [MIC 0.12-1 mg/L]) while 99.5% and 95.5% of isolates were susceptible to telithromycin and amoxicillin-clavulanate, respectively. In total, 88.2%, 87.5%, 99.4%, 100%, and 100% of H. influenzae isolates were susceptible to ampicillin, clarithromycin, cefuroxime, telithromycin, and amoxicillin clavulanate, respectively. In vitro, telithromycin demonstrated the highest activity against M. catarrhalis (MIC50 = 0.06 mg/L, MIC90 = 0.12 mg/L). CONCLUSION: The high in vitro activity of against pathogens commonly isolated in ABS, together with a once daily dosing regimen and clinical efficacy with 5-day course of therapy, suggest that telithromycin may play a role in the empiric treatment of ABS. PMID- 15287990 TI - Use and improvement of microbial redox enzymes for environmental purposes. AB - Industrial development may result in the increase of environmental risks. The enzymatic transformation of polluting compounds to less toxic or even innocuous products is an alternative to their complete removal. In this regard, a number of different redox enzymes are able to transform a wide variety of toxic pollutants, such as polynuclear aromatic hydrocarbons, phenols, azo dyes, heavy metals, etc. Here, novel information on chromate reductases, enzymes that carry out the reduction of highly toxic Cr(VI) to the less toxic insoluble Cr(III), is discussed. In addition, the properties and application of bacterial and eukaryotic proteins (lignin-modifying enzymes, peroxidases and cytochromes) useful in environmental enzymology is also discussed. PMID- 15287991 TI - Preliminary inventory and classification of indigenous afromontane forests on the Blyde River Canyon Nature Reserve, Mpumalanga, South Africa. AB - BACKGROUND: Mixed evergreen forests form the smallest, most widely distributed and fragmented biome in southern Africa. Within South Africa, 44% of this vegetation type has been transformed. Afromontane forest only covers 0.56 % of South Africa, yet it contains 5.35% of South Africa's plant species. Prior to this investigation of the indigenous forests on the Blyde River Canyon Nature Reserve (BRCNR), very little was known about the size, floristic composition and conservation status of the forest biome conserved within the reserve. We report here an inventory of the forest size, fragmentation, species composition and the basic floristic communities along environmental gradients. RESULTS: A total of 2111 ha of forest occurs on Blyde River Canyon Nature Reserve. The forest is fragmented, with a total of 60 forest patches recorded, varying from 0.21 ha to 567 ha in size. On average, patch size was 23 ha. Two forest communities - high altitude moist afromontane forest and low altitude dry afromontane forest - are identified. Sub-communities are recognized based on canopy development and slope, respectively. An altitudinal gradient accounts for most of the variation within the forest communities. CONCLUSION: BRCNR has a fragmented network of small forest patches that together make up 7.3% of the reserve's surface area. These forest patches host a variety of forest-dependent trees, including some species considered rare, insufficiently known, or listed under the Red Data List of South African Plants. The fragmented nature of the relatively small forest patches accentuates the need for careful fire management and stringent alien plant control. PMID- 15287993 TI - 'Gammon's law and a change in thinking?'. PMID- 15287992 TI - Are p.I148T, p.R74W and p.D1270N cystic fibrosis causing mutations? AB - BACKGROUND: To contribute further to the classification of three CFTR amino acid changes (p.I148T, p.R74W and p.D1270N) either as CF or CBAVD-causing mutations or as neutral variations. METHODS: The CFTR genes from individuals who carried at least one of these changes were extensively scanned by a well established DGGE assay followed by direct sequencing and familial segregation analysis of mutations and polymorphisms. RESULTS: Four CF patients (out of 1238) originally identified as carrying the p.I148T mutation in trans with a CF mutation had a second mutation (c.3199del6 or a novel mutation c.3395insA) on the p.I148T allele. We demonstrate here that the deletion c.3199del6 can also be associated with CF without p.I148T. Three CBAVD patients originally identified with the complex allele p.R74W-p.D1270N were also carrying p.V201M on this allele, by contrast with non CF or asymptomatic individuals including the mother of a CF child, who were carrying p.R74W-p.D1270N alone. CONCLUSION: These findings question p.I148T or p.R74W-p.D1270N as causing by themselves CF or CBAVD and emphazises the necessity to perform a complete scanning of CFTR genes and to assign the parental alleles when novel missense mutations are identified. PMID- 15287994 TI - Prevalence of overactive bladder and incontinence in Canada. AB - AIMS OF STUDY: The main objectives of the study were to measure the prevalence of overactive bladder (OAB) with its sub-types (wet OAB, dry OAB, mixed OAB) in the Canadian population and to assess prevalence variations in according to gender and age. METHODS: The prevalence of OAB in Canada was investigated via a validated, computer-assisted telephone interview (CATI) system. The interviews were conducted in two steps: an initial questionnaire evaluating if respondents suffered from OAB, followed by a detailed questionnaire completing the assessment. The final data were weighted according to the census of metropolitan areas and by gender, on the basis of Canadian population statistics. RESULTS: A sample of 3249 adults, aged 35 years and older, was interviewed with the CATI system. Six hundred and three respondents were found to suffer from wet OAB, dry OAB or mixed OAB. The overall prevalence of OAB in this Canadian population was evaluated to be 18.1%. It was lower in men (14.8%) than in women (21.2%). Dry OAB was assessed to be the highest sub-type, with a rate of 13.6%, 11.7% in men and 15.6% in women. The prevalence of wet OAB was estimated to be 2.3%, 2% in men and 2.6% in women. Finally, the prevalence of mixed OAB was found to be 1.2% in this population, with a much lower prevalence in men (0.3%) than in women (2.1%). Dry OAB increased with age in both men and women; wet OAB was markedly higher in both men and women over the age of 75 years. No correlation was observed between age and prevalence of mixed OAB in women, whereas a linear relationship was noted between the two variables in men. CONCLUSIONS: This is the first large study of OAB symptoms prevalence in the Canadian population aged 35 years and older. We established that OAB prevalence was higher in women than in men, with women experiencing more dry OAB and mixed OAB than men. PMID- 15287995 TI - Restoring sexual function in prostate cancer patients: an innovative approach. AB - It has been extremely difficult for men with prostate cancer to obtain reasonable estimates of the likelihood of remaining potent after first line therapy, partly because of differences in defining potency. If, as in more recent studies, the definition requires that men are usually (not just occasionally) able to get and sustain an erection, then the picture is not encouraging. Additional strategies are needed to help men sustain sexual activity. In this paper we draw on the experiences of a rather remarkable prostate cancer patient to help consider the possibilities for a different kind of intervention for men with ED--use of a strap-on dildo (an external prosthetic penis fastened by a harness around the hips). The dildo is a simple and inexpensive strategy for dealing with impotence and in certain circumstances it can work better than more established medical treatments for ED. Use of a dildo potentially removes the fear of erectile failure, allows for increased stimulation of the glans, facilitates full-body contact between partners, and offers potential satisfaction to one's partner. Urologists (and other health professionals) are encouraged to explore dildos as an option during discussions with patients about sexual rehabilitation. The potential benefits are discussed of specialty sexuality clinics that facilitate introduction of innovative approaches like dildos. PMID- 15287996 TI - A survey of urological manpower, technology, and resources in Canada. AB - INTRODUCTION: Knowledge of the current status of manpower and resources is important in understanding the state of any medical specialty, and critical in planning for future recruitment, funding and infrastructure development. METHODS: In 2003, the Canadian Urological Association (CUA) conducted two nationwide surveys examining manpower, resources, and the technology available. One survey went only to academic and hospital leaders across the country (the resources survey), while the other was sent to the entire general membership of the CUA. RESULTS: The response rate for the resources survey was 67%, while that for the membership survey was 50.4%. The respondents' ages were evenly distributed, with the modal 5-year range being 51 to 55 years of age. Eighty-eight percent of respondents were Canadian-trained. Two-thirds of respondents spent over 80% of their practice time in direct patient care, and most practiced general urology. The majority of respondents practiced in smaller hospitals: 57.6% in centres with 300 or fewer inpatient beds, and 47.2% of centres reported < 500 procedures/year. Community hospitals (62% of responses to the resources survey) generally had fewer advanced technologies than academic centres. A quarter of the cystoscopy equipment used by respondents was over 15 years old. CONCLUSIONS: The results of these surveys present a snapshot of the current state of urology resources and manpower across Canada, potentially allowing better planning and negotiations with hospitals and governments. PMID- 15287997 TI - Multiple bilateral perivascular epithelioid cell tumor (PEComa) of the kidneys. AB - Perivascular Epithelial Cell (PEComa) Tumors are extremely rare. These tumors are often regarded as low grade Sarcomas and treated as such. We report a case of a 70 year old female with a history of nonspecific complaints and on routine CT scan had bilateral multiple renal masses with no other extra renal disease. The largest mass on the right was 11 cm x 7 cm x 11 cm and the left there were multiple smaller masses. She underwent a right radical nephrectomy that showed a PEComa and an adjacent renal cell carcinoma. Her contralateral kidney was followed for 3 years with no evidence of growth or metastasis. PMID- 15287998 TI - McMaster experience with laparoscopic pyeloplasty. AB - PURPOSE: Laparoscopic pyeloplasty has been developed as a minimally invasive alternative to open pyeloplasty for the treatment of ureteropelvic junction obstruction (UPJO). Several series have been published with similar success rates for the two procedures. We present our initial experience with laparoscopic pyeloplasty. MATERIAL AND METHODS: A retrospective review of 29 consecutive patients (mean age 37 years) who underwent Laparoscopic dismembered Hynes Anderson pyeloplasty in our institution between January 2001 to April 2003 was performed. All patients had flank pain with radiologic findings consistent with ureteropelvic junction obstruction and impaired drainage on diuretic renal scan. Patients were assessed at 6 weeks with an ultrasound and assessment of pain, then an intravenous pyelogram (i.v.p.) and diuretic renogram were completed at 6 months along with a repeat clinical assessment. RESULTS: Twenty-nine patients underwent the procedure with one patient converted to an open procedure due to difficulties with the anastomosis. Mean operating time was 225 minutes, which decreased with experience. Mean blood loss was 50 cc and no patient required transfusion. Mean hospital stay was 2.5 days. Mean follow-up was 12 months. Twenty-six patients had complete resolution of their pain and an improvement on ultrasound was demonstrated, but only six patients showed improvement in function on i.v.p. or renogram at 6 months. In five patients with 25% or less differential renal function preoperatively, the function was worse or negligible despite complete resolution of symptoms. One patient developed stent migration requiring repositioning and another developed calcification on the distal end of the stent requiring cystolithalopaxy prior to stent removal. CONCLUSIONS: In our experience, laparoscopic pyeloplasty offers excellent symptomatic relief in a minimally invasive fashion with low morbidity for adult patients with ureteropelvic junction obstruction. In patients with borderline function (25% or less) preoperatively and with a normal functioning contralateral kidney, nephrectomy should be a consideration. PMID- 15287999 TI - Parameters of successful sacral root neuromodulation of the pelvic floor: a retrospective study. AB - INTRODUCTION/OBJECTIVE: Neuromodulation of the pelvic floor (InterStim) is a relatively new technique in the field of urology. We present our observations for effective neuromodulation on our patient population. MATERIALS AND METHODS: In a retrospective case review study, we studied the charts of 67 patients, who underwent InterStim operations between the years 1993 to 2002. All patients had a good response to InterStim. Patients with inefficient or inconclusive responses were not included in the study. All the relevant patient data was recorded from their charts. For each patient, the following was recorded; the amplitude in volts, the pulse width (in microseconds) and rate, the mode (cycling versus continuous), the electrodes and their position, the load impedance, and the change in amplitude over time. RESULTS: Amplitude over time showed an initial plateau, followed by a small increase that gets larger. CONCLUSIONS: Long-term management of InterStim recipients requires increasing amplitude following the implantation of the IntreStim to maintain the same satisfactory levels of urinary control. PMID- 15288000 TI - Laparoscopic resection of a juxta-adrenal schwannoma. AB - A seven centimeter supra-renal mass was discovered in a 62-year old patient who presented with gross hematuria and a superficial bladder tumor. The supra-renal mass was resected laparoscopically and the final pathology revealed a benign schwannoma. The epidemiology, diagnostic features and treatment options for this rare peripheral nerve sheath tumor are reviewed. PMID- 15288001 TI - Viruses and autoimmune diseases--adapting Koch's postulates. AB - Viral hypotheses for the aetiopathogenesis of autoimmune diseases are unproven. Indeed, a primary role for virus-induced autoimmune disease has yet to be fully established even in experimental systems. However new insights into virus-host interactions and improved technology justify fresh evaluation of these issues. We suggest modified Koch's postulates to test the viral hypothesis. PMID- 15288002 TI - Epstein-Barr virus and rheumatoid arthritis. AB - The cause of rheumatoid arthritis (RA) is still unknown. Both genetic and environmental factors may help its development. For 25 years, the Epstein-Barr Virus (EBV) has been suspected to contribute to RA pathogenesis. RA patients have higher levels of anti-EBV antibodies than healthy controls. EBV-specific suppressor T cell function is defective in RA. HLA-DRB1*0404, an RA predisposing allele, is associated with low frequencies of T cells specific for EBV gp110, a replicative phase glycoprotein critical for the control of EBV infection. Patients with RA have higher EBV load in peripheral blood lymphocytes (median 8.84 copies per 500 ng DNA) than healthy controls (median 0.6 copies/500 ng DNA). EBV, a widespread virus, highly recognized by antibodies but never eliminated, is an ideal candidate to trigger chronic immune complex disease. Anti-EBV antibody responses should be considered as one of the chronic autoantibody responses that are most relevant to the development of RA. PMID- 15288003 TI - BLyS--an essential survival factor for B cells: basic biology, links to pathology and therapeutic target. AB - A paradigm shift in our understanding of autoimmune disease pathology is underway; B cells are now considered to play a central role in disease pathogenesis. Targeting B cells may prove to be an effective route for the development of novel therapeutics. BLyS, a member of the TNF family of cytokines, is an essential survival factor for B cells. Constitutive BLyS overexpression in mice leads to an autoimmune phenotype similar to lupus nephritis. Clinically, BLyS is elevated in patients with systemic autoimmune diseases including rheumatoid arthritis and systemic lupus erythematosus. BLyS ablation results in a block of B cell development in which mature B cells are absent. BLyS binds to three receptors, BR3, TACI and BCMA. Analysis of the receptors suggests that the major pro-survival signals are mediated by BR3, while TACI is involved in negative signaling. BCMA is required for survival of long-lived plasma cells. BLyS signaling results in upregulation of anti-apoptotic bcl-2 family members. In animal models of autoimmune disease, BLyS antagonists reduce disease severity and delay disease progression. BLyS is an attractive target for antagonism in autoimmune diseases. Multiple approaches are being taken to antagonize BLyS including a fully human antibody and soluble BLyS receptors. These approaches are currently being tested in clinical trials. PMID- 15288004 TI - Systemic autoimmune diseases in elderly patients: atypical presentation and association with neoplasia. AB - The possible consequences of the progressive 'ageing' of the immune system are the increase in autoimmune phenomenon, incidence of neoplasia and predisposition to infections. This review analyses the clinical expression of the systemic autoimmune diseases in older patients, focusing on three specific characteristics: the frequent atypical presentation of autoimmune diseases in the elderly, the higher morbidity and mortality of these patients and the frequent association with neoplasic processes. The study of autoimmune manifestations in elderly populations should be considered a priority for future medical research because of increasing life expectation, especially in developed countries. PMID- 15288005 TI - Autoimmune uveitis and antigenic mimicry of environmental antigens. AB - Autoimmunity directed against antigens of immune privileged sites, which are hidden from the immune system by blood-organ-barriers, is difficult to explain: it would require already activated cells to enter the tissue where the respective autoantigens are sequestered. Autoimmune uveitis, a sight-threatening inflammatory disease of the eye, is such an example. To induce disease autoreactive T cells must have been activated outside the eye to pass the blood retina-barrier and then crossreact with retinal autoantigen. We have described two environmental peptides mimicking a highly pathogenic epitope from retinal S antigen. One mimicry antigen is from rotavirus, a common pathogen causing gastroenteritis, the other from bovine milk alpha s2casein, a frequent nutritional protein ought to induce oral tolerance. Lewis rats develop uveitis after immunization with both mimicry peptides and casein protein. However, these mimicry antigens failed to induce oral tolerance for protection from uveitis, suspecting that they rather induce immunity than tolerance. Humoral and cellular immune responses to these antigens are enhanced and more frequent in patients with uveitis compared to healthy individuals. Our findings suggest that multiple environmental antigens mimic autoantigens and might cause autoimmune diseases by eliciting defensive immune responses, however, they are not necessarily useful for therapeutic tolerance induction. PMID- 15288006 TI - Interferon induced Autoimmune Thyroid Disease (AITD): a model for human autoimmunity. AB - Interferon (IFN) alpha treatment for various conditions has been associated with thyroid autoimmunity. The incidence of interferon induced thyroid autoimmunity has been reported to range from 2.5% to 42%, possibly depending upon dose and duration of medical therapy and patient characteristics. It is not known whether IFN-alpha initiates autoimmune thyroid disease (AITD) or simply exacerbates AITD in individuals with subclinical AITD. PMID- 15288007 TI - Inflammatory bowel disease: the role of environmental factors. AB - Environmental factors are essential components of the pathogenesis of inflammatory bowel disease (IBD) and primarily responsible for its growing incidence around the globe. Epidemiological, clinical and experimental evidence support an association between IBD and a large number of seemingly unrelated environmental factors, which include smoking, diet, drugs, geographical and social status, stress, microbial agents, intestinal permeability and appendectomy. Data supporting the involvement of each of these factors in predisposing to, triggering, or modulating the course or outcome of IBD vary from strong to tenuous. Smoking and the enteric bacterial flora are the ones for which the most solid evidence is currently available. Smoking increases the risk of Crohn's disease (CD) and worsens its clinical course, but has a protective effect in ulcerative colitis (UC). Presence of enteric bacteria is indispensable to develop gut inflammation in most animal models of IBD, and modulation of the quantity or quality of the flora can be beneficial in patients with IBD. Surprisingly, evidence for a major role of the diet in inducing or modifying IBD is limited, while that for nonsteroidal anti-inflammatory drugs is more convincing than for oral contraceptives. Northern geographic location, and a high social, economical, educational or occupational status increase the risk of IBD, an observation fitting the hygiene hypothesis for allergic and autoimmune diseases. Stress is also associated with IBD, but more as a modifier than an inducing factor, and its contribution is more obvious in IBD animal models than human IBD. Finally, an increased intestinal permeability may increase the risk for developing CD, whereas an appendectomy lowers the risk of developing UC. PMID- 15288008 TI - Inflamm-aging: autoimmunity, and the immune-risk phenotype. AB - Aging of the immune system, or immunosenescence, is a complex subject best defined as a decline in cell-mediated immunity, particularly with respect to T cell function. Paradoxically with the decline in immune function is an increase in autoantibody frequency. It has been postulated that the accumulation of anamnestic cells over time and/or environmental/infectious mimics leads to the production of autoantibodies, sometimes accompanied by autoimmune disease. This specific phenotype has given rise to the concept of a specific cluster of cytokine profiles, coined an immune-risk phenotype (IRP). The IRP is likely dictated by not only cytokine production, but also defects in activation-induced cell death and also a shift in T cell subsets. These concepts are an important bridge between basic immune function and clinical immunology in the hopes for generation of effective reconstitution to improve immune function in the elderly. PMID- 15288009 TI - Hormones and the genome, HLSC 3rd event, April 29-May 1, 2004 endocrine malignancies and autoimmunity--Molecular biological events and targets for clinical application, Reisensburg Castle, Guenzburg, Germany. PMID- 15288010 TI - Liver transplantation for hepatitis B. AB - Recurrent HBV is almost universal post-LT and is accompanied by significant graft and patient loss in the absence of effective immunoprophylaxis. Hepatitis B immunoglobulin (HBIG) monotherapy and lamivudine monotherapy significantly reduce the rate of recurrent hepatitis B, but recurrent hepatitis B still occurs in up to 25% due to the emergence of resistant mutants. Combination administration of HBIG and lamivudine is more efficacious in preventing recurrent hepatitis B, decreasing recurrence rates of hepatitis B to 0-18% in studies. Future studies to determine the optimal dosing regimen and duration of HBIG and lamivudine and to evaluate the efficacy of newer antivirals such as adefovir dipivoxil in preventing recurrent HBV are needed. Treatment of established recurrent hepatitis B remains problematic. Lamivudine has shown promise during the initial treatment period, but is plagued by rapid development of viral resistance with longer treatment duration. Adefovir dipivoxil appears very promising, and further studies are needed to evaluate its efficacy in the post-liver transplant patient. Interferon-alfa's use in the post-liver transplant patient is limited given the availability of the newer antiviral drugs. Famciclovir and ganciclovir have shown some promise in treating recurrent HBV, but have been replaced by newer agents such as lamivudine and adefovir. PMID- 15288011 TI - Plasma osteopontin levels in patients with fulminant hepatitis. AB - Fulminant hepatitis is characterized by massive or submassive liver necrosis. Massive liver necrosis can be induced by activated macrophages infiltrating into the liver. Osteopontin, an extracellular matrix, is a secretory glycoprotein as well essential for Th1 immune response, contributing to macrophage activation and infiltration. To know the significance of osteopontin in the development of fulminant hepatitis, plasma osteopontin levels were measured in patients with fulminant hepatitis. The levels were significantly greater in patients with fulminant hepatitis than in those with acute or chronic hepatitis as well as healthy adults. Among patients with fulminant hepatitis except one in whom bacterial infection was complicated, plasma osteopontin levels were elevated especially in the patients who developed hepatic encephalopathy of grade II or more within 10 days of the disease onset, a clinical type characteristic of massive liver necrosis. Immunohistochemical examination revealed that osteopontin was stained in macrophages positive for CD68, a marker for macrophages, in necrotic areas of the liver in a patient with fulminant hepatitis. In conclusion, plasma osteopontin levels were elevated in patients with fulminant hepatitis, probably reflecting production of osteopontin in Kupffer cells and hepatic macrophages, which might be involved in the development of massive liver necrosis in fulminant hepatitis. PMID- 15288012 TI - Phagocytic function of neutrophils of patients with decompensated liver cirrhosis is restored by oral supplementation of branched-chain amino acids. AB - It has been reported that phagocytic function of neutrophils is impaired in cirrhotic patients. We examined the effects of oral supplementation of branched chain amino acids (BCAA) on phagocytic function of neutrophils in peripheral blood of patients with decompensated cirrhosis. Five patients with decompensated cirrhosis received 12g of BCAA daily for 3 months. Phagocytic function of neutrophils and NK activities of lymphocytes in peripheral blood as well as serum albumin levels and Fisher's ratios were determined before and at 1 and 3 months of BCAA supplementation. Phagocytic function of neutrophils was significantly improved by 3-month BCAA oral supplementation. NK activity of lymphocytes was improved in four of five patients at 3 months of BCAA supplementation, although the changes were not statistically different. In conclusion, BCAA supplementation improved phagocytic function of neutrophils in cirrhotic patients. BCAA supplementation may reduce the risk of bacterial infection in patients with decompensated liver cirrhosis. PMID- 15288013 TI - Estimation of the mutation rate of hepatitis E virus based on a set of closely related 7.5-year-apart isolates from Sapporo, Japan. AB - Since hepatitis E virus (HEV) does not persist in infected hosts or in cultured cell lines, it has been difficult to know its spontaneous mutation rate. Recently, we identified an HEV isolate in stored serum from a patient having developed hepatitis E in 1995 (JSM-Sap95), nucleotide sequence of which showed a strong resemblance to those obtained from three patients who contracted hepatitis E in 2000 and 2002 (JKK-Sap00, JYW-Sap02, and JTS-Sap02). The remarkable nucleotide similarity together with the fact that all these patients were residents of the same city, Sapporo, prompted us to hypothesize that JKK-Sap00, JYW-Sap02 and JTS-Sap02 are descendants of JSM-Sap95. Then, the mutation rate of HEV was calculated to be 1.72, 1.41, or [Formula: see text] base substitutions per site per year, from JSM-Sap95 to JKK-Sap00, JYW-Sap02 or JTS-Sap02, respectively. Interestingly, these values were very similar to those ( [Formula: see text] to [Formula: see text] ) reported for hepatitis C virus. Because it remains possible that JSM-Sap95 is not the direct ancestor of the other three isolates but was merely a relative of the true ancestor, HEV mutation rate may be a little lower than [Formula: see text] base substitutions per site per year. PMID- 15288014 TI - Prospective randomized crossover trial of combination therapy with bezafibrate and UDCA for primary biliary cirrhosis. AB - The aim of this study was to evaluate the effects of the combination therapy with bezafibrate and ursodeoxycholic acid (UDCA) for primary biliary cirrhosis (PBC), compared to UDCA monotherapy. Sixteen patients with compensated PBC were divided randomly into two groups. Group A received treatment with bezafibrate and UDCA for 6 months, while group B received UDCA alone, treatment protocols were then exchanged for another 6 months. The laboratory data was followed every month. The mean levels of alkaline phosphatase (ALP) decreased significantly more in group A than in group B in the first half of the study. Then serum ALP levels were elevated in group A after exchanged the therapy, but fell down in group B. Serum levels of gamma-glutamyl transferase (GGT), immunoglobulin M and triglycerides values were significantly lower in group B than in group A, after changing therapies from monotherapy to combination therapy with bezafibrate and UDCA. The mean levels of ALP, GGT and triglycerides were significantly lower at the end of the combination therapy than those at the end of the monotherapy. The combination therapy with bezafibrate and UDCA significantly improves the laboratory data that specific for PBC in comparison with UDCA monotherapy. PMID- 15288015 TI - Clinicopathologic characteristics of small intrahepatic cholangiocarcinomas of mass-forming type. AB - We investigated the clinicopathologic characteristics of and outcome after treatment for small intrahepatic cholangiocarcinomas of mass-forming type (3cm or smaller) in nine patients to study appropriate treatment. In seven of the nine patients, the cancer was detected during follow-up for hepatitis C virus-related disease. One patient was seropositive for anti-hepatitis B core antibody alone. One patient had alcoholic cirrhosis. Three patients also had hepatocellular carcinoma. Seven patients underwent liver resection and two other patients underwent microwave coagulation therapy. In the resection specimens, no vascular or lymphatic invasion, or invasion of perineural spaces, was evident pathologically. No patient had intrahepatic metastasis or lymph node metastasis; all had hepatic fibrosis. Lung metastases developed after surgery in one patient whose surgical margin was positive for cancer. In one patient with both type of carcinoma, the hepatocellular carcinoma recurred. Screening for not only hepatocellular carcinoma but also intrahepatic cholangiocarcinoma is necessary in patients with hepatic fibrosis, especially those infected with hepatitis C virus. For small intrahepatic cholangiocarcinomas of mass-forming type located peripherally in the liver, partial resection or ablation therapy with care to obtain a negative surgical margin may be appropriate. PMID- 15288016 TI - Clinicopathologic analysis of risk factors for distant metastasis of hepatocellular carcinoma. AB - Background: Numbers of patients with extrahepatic metastasis of hepatocellular carcinoma (HCC) are increasing. As characteristics of HCC patients who develop distant metastasis remain uncertain, we analyzed potential risk factors. Methods: Among patients followed up after hepatic resection for HCC, we identified 31 patients with extrahepatic metastasis (extrahepatic metastasis group, EM). In 46 other patients we found intrahepatic metastasis synchronously with the primary HCC. In these patients no extrahepatic metastasis occurred during 5 years of follow-up (intrahepatic metastasis group, IM). During the 5-year follow-up period, 14 patients have remained alive with no metastasis (no metastasis group, NM). We compared age, gender, hepatitis virus infection status, grade of liver dysfunction, serum alpha-fetoprotein (AFP) concentration, tumor size, tumor differentiation, presence of portal or hepatic venous infiltration, and surgical strategy among these three groups. Results: No significant difference in clinicopathologic factors was noted among the three groups except for immunohistochemical staining for CD44. A significantly higher proportion of tumors was immunoreactive for CD44v3 in the EM group than in the other groups ( [Formula: see text] ). By multivariate analysis, CD44v3 expression (risk ratio, 9.28; [Formula: see text] ) was a significant independent risk factor for extrahepatic metastasis. Conclusion: CD44v3 expression in HCC may be a useful predictor of likelihood of distant metastasis. PMID- 15288017 TI - Usefulness of contrast-enhanced ultrasonography with a new contrast mode, Agent Detection Imaging, in evaluating therapeutic response in hepatocellular carcinoma treated with radio-frequency ablation therapy. AB - Objective: The therapeutic response of radio-frequency ablation (RFA) for hepatocellular carcinoma (HCC) was evaluated by contrast-enhanced ultrasonography (US) using a new contrast mode, agent detection imaging (ADI), and its usefulness was compared with that of dynamic computed tomography (CT). Materials and methods: Forty patients with 64 nodules diagnosed as HCC histologically and/or by various imaging modalities were evaluated with ADI (1 and 7 days after treatment) and Dynamic CT (7 days after treatment). Results: ADI and dynamic CT revealed residual lesions in 15 and 12 nodules, and no residual lesions in 45 and 52 nodules, respectively. ADI yielded indeterminate findings in four nodules; although the vascular phase showed minute residual tumor vessels, the tumor appeared to be completely contained within the ablated area in the delayed phase. In these indeterminate cases, US-guided biopsies of the areas of residual vascularity revealed no viable cancer cells; only degenerated tissues. Conclusion: In assessing the therapeutic response of RFA for HCC, ADI has a similar competence to dynamic CT. Even if the vascular phase of contrast-enhanced US shows minute residual vessels, as long as the tumor is safely contained within the ablated area in the delayed phase, the cancer cells are likely to be degenerated. PMID- 15288018 TI - Proinflammatory cytokines up-regulate synthesis and secretion of urinary trypsin inhibitor in human hepatoma HepG2 cells. AB - Background and Aims: The urinary trypsin inhibitor (UTI), a wide range protein inhibitor synthesized by hepatocytes, is considered to play an important role not only in the protection of organ injury during severe inflammation but also in the inhibition of tumor invasion and metastasis. However, the precise mechanisms underlying control of its synthesis, secretion, and processing remain unclarified. The aim of this study is to determine whether human hepatoma HepG2 cells secrete UTI in free form and whether its synthesis and secretion are regulated by proinflammatory cytokines. Methods: Cultured HepG2 cells were stimulated using different concentrations of interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta). The concentration of free UTI in the medium was measured by ELISA and the intracellular UTI precursor was identified by western blotting. UTI mRNA expression was studied by reverse transcription polymerase chain reaction (RT-PCR). Results: HepG2 cells constantly secreted free UTI and this secretion was significantly up-regulated by IL-6, IL 1beta, and TNF-alpha. IL-6, IL-1beta, and TNF-alpha enhanced the synthesis of the intracellular UTI precursor protein, and IL-1beta up-regulated UTI mRNA expression. Conclusions: HepG2 cells constantly secrete free UTI. The proinflammatory cytokines, IL-1beta, IL-6, and TNF-alpha, up-regulate UTI synthesis and secretion by up-regulating UTI mRNA expression. PMID- 15288019 TI - A comparative genomic analysis of the calcium signaling machinery in Neurospora crassa, Magnaporthe grisea, and Saccharomyces cerevisiae. AB - A large number of Ca2+ -signaling proteins have been previously identified and characterized in Saccharomyces cerevisiae but relatively few have been discovered in filamentous fungi. In this study, a detailed, comparative genomic analysis of Ca2+ -signaling proteins in Neurospora crassa, Magnaporthe grisea, and S. cerevisiae has been made. Our BLAST analysis identified 48, 42, and 40 Ca2+ signaling proteins in N. crassa, M. grisea, and S. cerevisiae, respectively. In N. crassa, M. grisea, and S. cerevisiae, 79, 100, and 13% of these proteins, respectively, were previously unknown. For N. crassa, M. grisea, and S. cerevisiae, respectively, we have identified: three Ca2+ -permeable channels in each species; 9, 12, and 5 Ca2+/cation-ATPases; eight, six, and four Ca2+ exchangers; four, four, and two phospholipase C's; one calmodulin in each species; and 23, 21, and 29 Ca2+/calmodulin-regulated proteins. Homologs of a number of key proteins involved in the release of Ca2+ from intracellular stores, and in the sensing of extracellular Ca2+, in animal and plant cells, were not identified. The greater complexity of the Ca2+ -signaling machinery in N. crassa and M. grisea over that in S. cerevisiae probably reflects their more complex cellular organization and behavior, and the greater range of external signals which filamentous fungi have to respond to in their natural habitats. To complement the data presented in this paper, a comprehensive web-based database resource (http://www.fungalcell.org/fdf/) of all Ca2+ -signaling proteins identified in N. crassa, M. grisea, and S. cerevisiae has been provided. PMID- 15288020 TI - Key differences between lateral and apical branching in hyphae of Neurospora crassa. AB - We examined in fine detail growth kinetics and intracellular events during lateral and apical branching in hyphae of Neurospora crassa. By high-resolution video-enhanced light microscopy, we found remarkable differences in the events preceding lateral vs apical branching. While apical branching involved a significant disturbance in the apical growth of the parental hypha, lateral branching occurred without any detectable alterations in the growth of the parental hypha. Prior to the emergence of a lateral branch, an incipient Spitzenkorper was formed about 12-29 microm behind the apex. Lateral branch formation did not interfere with the elongation rate of the primary hypha, the shape of its apex or the behavior of its Spitzenkorper. In sharp contrast, apical branching was preceded by marked changes in physiology and morphology of the parental hypha and by a sharp drop in elongation rate. The sequence involved a cytoplasmic contraction, followed by a retraction, dislocation, and disappearance of the Spitzenkorper; hyphal elongation decreased sharply and a transient phase of isotropic growth caused the hyphal apex to round up. Growth resumed with the formation of two or more apical branches, each one with a Spitzenkorper formed by gradual condensation of phase-dark material (vesicles) around an invisible nucleation site. The observed dissimilarities between lateral and apical branching suggest that these morphogenetic pathways are triggered differently. Whereas apical branching may be traced to a sudden discrete disruption in cytoplasmic organization (cytoplasmic contraction), the trigger of lateral branching probably stems from the subapical accumulation of wall precursors (presumably vesicles) reaching a critical concentration. PMID- 15288021 TI - Five hydrophobin genes in Fusarium verticillioides include two required for microconidial chain formation. AB - Five hydrophobin genes have been identified in the fungal corn pathogen Fusarium verticillioides. HYD1, HYD2, and HYD3 encode Class I hydrophobins. The predicted structures of Hyd1p and Hyd2p are 80% similar, while Hyd3p has an unusually small number of amino acids between the third and fourth cysteines. HYD4 and HYD5 encode Class II hydrophobins. Mutants with HYD1-5 individually deleted and a hyd1deltahyd2delta double mutant were similar to wild-type strains in the amount of disease caused in a corn seedling infection assay and in the number of microconidia produced. Microconidial chains were rare in hyd1delta and hyd2delta mutants as microconidia were present almost exclusively as false heads. Transformation of hyd1delta and hyd2delta mutants with HYD1 and HYD2, respectively, restored microconidial chain formation, but transformation with HYD1::AcGFP and HYD2::AcGFP did not complement the mutation. HYD1::AcGFP and HYD2::AcGFP localized to the outside of conidia in false heads and in chains. PMID- 15288022 TI - The mitochondrial plasmid pAL2-1 reduces calorie restriction mediated life span extension in the filamentous fungus Podospora anserina. AB - Calorie restriction is the only life span extending regimen known that applies to all aging organisms. Although most fungi do not appear to senesce, all natural isolates of the modular filamentous fungus Podospora anserina have a limited life span. In this paper, we show that calorie restriction extends life span also in Podospora anserina. The response to glucose limitation varies significantly among 23 natural isolates from a local population in The Netherlands, ranging from no effect up to a 5-fold life span extension. The isolate dependent effect is largely due to the presence or absence of pAL2-1 homologous plasmids. These mitochondrial plasmids are associated with reduced life span under calorie restricted conditions, suggesting a causal link. This has been substantiated using three combinations of isogenic isolates with and without plasmids. A model is proposed to explain how pAL2-1 homologues influence the response to calorie restriction. PMID- 15288023 TI - Biomechanical evidence for convergent evolution of the invasive growth process among fungi and oomycete water molds. AB - Diverse microorganisms traditionally called fungi are recognized as members of two kingdoms: mushroom-forming species and their relatives in the Fungi, and oomycete water molds in the Stramenopila. Phylogenetic analysis suggests that these kingdoms diverged early in the evolution of eukaryotes. The phylogenetic detachment of the fungi and oomycetes is reflected in radical differences in their biochemistry, cell structure, and development. In terms of their biological activities, however, they show great similarity, because both groups form colonies of filamentous hyphae that invade and decompose solid food sources. Here we present biomechanical evidence of the convergent evolution of the invasive growth process in these microorganisms. Using miniature strain gauges to measure the forces exerted by single hyphae, we show that the hyphae of species in both kingdoms exert up to 2 atmospheres of hydrostatic pressure as they extend at their tips. No other eukaryotes have adopted this process for meeting their nutritional needs. PMID- 15288024 TI - Cloning of genes expressed early during cellulase induction in Hypocrea jecorina by a rapid subtraction hybridization approach. AB - The cellulase system of the filamentous fungus Hypocrea jecorina (Trichoderma reesei) is encoded by several cellobiohydrolase, endoglucanase and beta glucosidase genes, which are co-ordinately expressed upon induction by cellulose or the disaccharide sophorose. To identify genes, which are specifically expressed under these inducing conditions and possibly related to the induction process, we applied rapid subtraction hybridization (RaSH) to sophorose induced mRNAs from the wild-type strain H. jecorina QM9414 and a mutant strain H. jecorina QM9978, which is defective in the induction of cellulase gene expression. From a total of 224 clones, 22 gene fragments representing 20 different genes were analyzed. These included one gene encoding a PAS-domain protein with similarity to the Neurospora clock modulator VIVID; one gene similar to Podospora anserina ami1 involved in nuclear migration and the genes encoding translation elongation factor 1alpha, the transcriptional activator Hap5, and myo inositol-1-phosphate synthase; in addition, several genes were detected, whose function is unknown. Some of them did not even have potential homologues in the Neurospora or Fusarium genome databases. The differential regulation of expression of those 20 genes by sophorose in wild-type and mutant was verified by Northern blotting. Their consistent response to additional inducing conditions (cellulose) confirms their interconnection with cellulase formation. PMID- 15288026 TI - Vaginal douching among adolescent and young women: more challenges than progress. AB - The practice of vaginal douching dates back centuries. Numerous studies have shown that douching is quite prevalent and often begins during adolescence. Motivation for the initiation and maintenance of this practice appears complex, and presents challenges to the intervention efforts. The practice of douching remains controversial. Douching has been implicated in numerous adverse reproductive health outcomes such as increased risk for pelvic inflammatory disease, ectopic pregnancy, reduced fertility, and bacterial vaginosis. However, recent studies in developing countries have suggested that in certain circumstances, douching may actually be beneficial. We summarize key findings from the review of published literature and ongoing research, as well as highlight research challenges to our understanding of the role of vaginal douching in reproductive health. PMID- 15288027 TI - Non-gynecologic causes of unexplained lower abdominal pain in adolescent girls: two clinical cases and review of the literature. PMID- 15288028 TI - Having a good relationship with their mother: a protective factor against sexual risk behavior among adolescent females? AB - STUDY OBJECTIVE: To assess whether having a good relationship with their mother was a protective factor against risky sexual behavior for female adolescents and whether it was independent of family structure. DESIGN: Cross-sectional survey of in-school adolescents aged 14-19 years. SETTING: Catalonia, in northeast Spain. PARTICIPANTS: A total of 3677 females divided according on whether they had a good (n=3335) or a bad (n=342) relationship with their mother. MAIN OUTCOME MEASURES: Rates of sexual activity and sexual behavior. RESULTS: Adolescents in the good relationship group were significantly younger, more likely to live in an intact family, to have a good relationship with their father and siblings, and to talk about sexuality and their partner with their mother. They were also less likely to have ever had sexual intercourse. Among those sexually experienced, they were significantly older at first intercourse and less likely to have multiple partners or a history of STI. After adjusting for potential confounders, females in the good relationship group were less likely to be sexually active and to have had multiple partners, independently of family structure. CONCLUSIONS: Having a good relationship with their mother is a protective factor against sexual intercourse and having multiple sexual partners independently of family structure. Communication between generations and having a good relationship with their father and siblings also play an important role. PMID- 15288029 TI - Teenage mother's predictions of their support levels before and actual support levels after having a child. AB - OBJECTIVES: To evaluate to what extent teenage mothers are able to predict their postnatal support networks in the antenatal period, and the extent to which support correlates with depressive symptomatology and social class. DESIGN: A prospective cohort study. SETTING: Teenage antenatal clinic. PARTICIPANTS: Teenage mothers (n=124) aged less than 18 years. INTERVENTION: Women were surveyed in the antenatal period and again 6 months postpartum about their anticipated and actual levels of support using the Maternal Social Support Index. The questionnaire covered key issues involved in discharge planning such as the availability of help with household tasks, emergency respite, and communication. Subjects also completed social class data and the Edinburgh Postnatal Depression Scale. MAIN OUTCOME MEASURE: Ability of pregnant teenagers to predict postnatal support and correlation of support with social class and depressive symptomatology. RESULTS: The key finding was that teenage mothers significantly overestimated their support networks in the antenatal period compared to the reality experienced 6 months postpartum (P=0.0001). Dissatisfaction was most marked in the communication and daily activity support subscales. The level of support at 6 months postpartum correlated significantly with the Edinburgh Postnatal Depression score (P=0.0001) and with social class (P=0.017). CONCLUSION: Services for teenage mothers that provide assistance with communication and daily support activities are required to help overcome the discrepancies between idealization and reality. Poor support correlates with depressive symptomatology and social class. PMID- 15288030 TI - Young Latinas recall contraceptive use before and after pregnancy: a focus group study. AB - STUDY OBJECTIVE: To understand factors influencing use and nonuse of contraception in young, low-income Latina adolescents through focus group conversations. DESIGN: We conducted seven focus group discussions with Latino females in an outpatient clinic and community setting. Qualitative data analyses techniques were used. SETTING: Participants were recruited from the outpatient gynecology clinic at the University of Illinois at Chicago, and from the Easter Seals Day Care Center. PARTICIPANTS: Women were recruited if they were Latina and between the ages of 18-26 (n=40). RESULTS: Due to lack of information and family beliefs prohibiting birth control use and sexual activity, sexually active teenagers had high rates of contraceptive non-use and relied on irregular use of condoms and withdrawal for pregnancy prevention. Following pregnancy, participants had greater access to contraception and more determination to use it. Yet, persistent use of withdrawal and irregular use of hormonal contraception may place Latina mothers at risk for repeat pregnancy. CONCLUSIONS: Different approaches are needed for prevention of the first pregnancy compared to subsequent ones among Latino adolescents and young adults. PMID- 15288033 TI - Management quandary. An unusual case of STI in a teenager. PMID- 15288032 TI - Early recognition and treatment of staphylococcal and streptococcal toxic shock. PMID- 15288034 TI - Ovarian cysts. PMID- 15288035 TI - Adoption as an option. PMID- 15288036 TI - 3 Tesla magnetic resonance imaging of the prostate with combined pelvic phased array and endorectal coils; Initial experience(1). AB - RATIONALE AND OBJECTIVES: High-resolution magnetic resonance imaging of the prostate at 1.5T has gained acceptance for pretherapeutic staging of prostate cancer. The aim of this study was to evaluate the potential clinical utility of combined pelvic phased-array and endorectal coils at 3T. MATERIALS AND METHODS: Six volunteers were examined on 1.5T and 3T scanners with pelvic phased-array surface coil combined with a disposable endorectal prostate coil. RESULTS: We were able to acquire T2-W fast spin echo images with 1.5 mm slices, field of view 12, matrix 320 x 192, (voxel size 0.35 mm(3)), with excellent anatomic detail and good T2 contrast. A 1.5 mm axial slice thickness permitted high-quality multiplanar reconstructions with clear visualization of small patho-anatomic structures. Dynamic contrast-enhanced gradient echo images showed excellent spatial resolution (voxel size, 0.38 mm(3)) and temporal resolution. With this level of anatomic information in dynamic images we could clearly distinguish between intracapsular and extracapsular contrast enhancement. CONCLUSION: Using modified T2-fast spin echo and dynamic contrast-enhanced gradient echo sequences, we obtained whole gland coverage with 35-38 microm(3) resolution, without interfering artifacts, in reasonable acquisition times and staying well below the specific absorption rate guidelines. The high spatial resolution in the axial plane allowed meaningful multiplanar reconstructions. The initial results show the clinical utility of endorectal 3T for the noninvasive evaluation of the prostate with image features and quality not achievable at 1.5 T. PMID- 15288037 TI - Quantitative evaluation of a pulmonary contour segmentation algorithm in X-ray computed tomography images. AB - RATIONALE AND OBJECTIVES: Pulmonary contour extraction from thoracic x-ray computed tomography images is a mandatory preprocessing step in many automated or semiautomated analysis tasks. This study was conducted to quantitatively assess the performance of a method for pulmonary contour extraction and region identification. MATERIALS AND METHODS: The automatically extracted contours were statistically compared with manually drawn pulmonary contours detected by six radiologists on a set of 30 images. Exploratory data analysis, nonparametric statistical tests, and multivariate analysis were used, on the data obtained using several figures of merit, to perform a study of the interobserver variability among the six radiologists and the contour extraction method. The intraobserver variability of two human observers was also studied. RESULTS: In addition to a strong consistency among all of the quality indexes used, a wider interobserver variability was found among the radiologists than the variability of the contour extraction method when compared with each radiologist. The extraction method exhibits a similar behavior (as a pulmonary contour detector), to the six radiologists, for the used image set. CONCLUSION: As an overall result of the application of this evaluation methodology, the consistency and accuracy of the contour extraction method was confirmed to be adequate for most of the quantitative requirements of radiologists. This evaluation methodology could be applied to other scenarios. PMID- 15288038 TI - Diagnostic accuracy of Fischer Senoscan Digital Mammography versus screen-film mammography in a diagnostic mammography population. AB - RATIONALE AND OBJECTIVES: To compare the diagnostic accuracy of the Fischer Senoscan Digital Mammography System with that of standard screen-film mammography in a population of women presenting for screening or diagnostic mammography. MATERIALS AND METHODS: Enrollment of patients took place at six different breast imaging centers between 1997 and 1999. A total of 247 cases were selected for inclusion in the final reader study. All known cancer cases were included (111) from all six participating sites representing 45% of the total cases. The remaining 136 cases (55%) were randomly selected from all available benign or negative cases from three of the six sites. A complete case consisted of both a (unilateral or bilateral) digital and screen-film mammogram of the same patient. Eight radiologists interpreted the cases in laser-printed digital and screen-film hardcopy formats. The study was designed to detect differences of 0.05 in the ROC area under the curve (AUC) between digital and screen-film radiologist interpretation performance. RESULTS: The average AUC for the Senoscan digital was 0.715 for the 8 readers. The average AUC for screen-film was 0.765. The difference AUC of -0.05 falls within the 95% confidence interval (-0.101, 0.002). The average sensitivity was 66% and specificity 67% for SenoScan full-field digital mammography. The average screen-film mammography sensitivity and specificity were 74% and 60%, respectively. CONCLUSION: No statistically significant difference in diagnostic accuracy between the Fischer Senoscan and screen-film mammography was detected in this study. PMID- 15288040 TI - A preliminary report on the role of spatial frequency analysis in the perception of breast cancers missed at mammography screening. AB - RATIONALE AND OBJECTIVES: Because several factors are involved in cancer detection, a malignant lesion that is visible on a mammogram will not necessarily be reported by the radiologist reading the case. Indeed, a significant fraction of screening-detected cancers are visible in retrospect, and were perceived by the radiologist when the case was read, but were either reported as benign findings or dismissed as variations of normal breast tissue. In this preliminary report the spatial frequency characteristics of clinically missed lesions are investigated by analyzing the mammogram acquired when the lesion was sent for biopsy and the most recent prior mammogram. For control purposes, the contralateral breast is also analyzed, when this breast is lesion free. MATERIALS AND METHODS: A database of 70 mammogram cases was assembled. Each case contained eight films: craniocaudal (CC) and mediolateral oblique (MLO) of the breast where a biopsy-proven lesion was found, CC and MLO of the contralateral breast, and CC and MLO of both breasts in the most recent prior mammogram. The dictated reports for all of these cases were obtained. Both benign and malignant lesions were used. The films were digitized and an region of interest surrounding each lesion was segmented from the image for processing using wavelet packets to extract spatial frequency information. The corresponding area was also segmented from the prior mammogram and from the contralateral breast, when this breast was lesion free. Analysis of variance was used to determine if statistically significant differences existed between the derived features of cancer in the current and prior mammograms. RESULTS: The data suggests that malignant lesions reported in the prior mammogram as being benign differed from correctly reported malignant lesions and from correctly reported benign lesions. They also differed from nonreported malignant lesions. In addition, the spatial frequency representation of cancer significantly differed in the current and prior cases from the representation of normal breast tissue. CONCLUSION: Spatial frequency analysis may be useful to differentiate malignant lesions that are reported as benign and correctly reported benign lesions. PMID- 15288039 TI - Computer-assisted diagnosis by temporal subtraction in postoperative brain tumor patients: a feasibility study. AB - RATIONALE AND OBJECTIVES: To introduce and evaluate a novel, image fusion-based technique that can be used to compare the findings of primary and control brain magnetic resonance imaging scans, with special attention to the differences found in this comparison. MATERIALS AND METHODS: A new technique named "colored difference mapping" was applied to the brain examinations of five patients. The possible changes in the magnetic resonance imaging findings were analyzed by the colored difference mapping technique and by using conventional film reading and the results were compared. RESULTS: Colored difference mapping accurately depicts the differences between successive magnetic resonance images and reveals small changes that are difficult to perceive in a visual evaluation. CONCLUSION: Colored difference mapping is suitable for comparison of images between two different radiologic examinations and helps to show even minimal changes in brain tissues. PMID- 15288041 TI - Patient, faculty, and self-assessment of radiology resident performance: a 360 degree method of measuring professionalism and interpersonal/communication skills. AB - RATIONALE AND OBJECTIVES: To develop and test the reliability, validity, and feasibility of a 360-degree evaluation to measure radiology resident competence in professionalism and interpersonal/communication skills. MATERIALS AND METHODS: An evaluation form with 10 Likert-type items related to professionalism and interpersonal/communication skills was completed by a resident, supervising radiologist and patient after resident-patient interactions related to breast biopsy procedures. Residents were also evaluated by faculty, using an end-of rotation global rating form. Residents, faculty, and technologists were queried regarding their reaction to the assessments after a 7-month period. RESULTS: Fifty-six complete 360-degree data sets (range, 2-14 per resident) and seven rotational evaluations for seven residents were analyzed and compared. Internal consistency reliability estimates were 0.85, 0.86, and 0.87 for resident, patient, and faculty 360-degree evaluations, respectively. Correlations between resident-versus-patient, resident-versus-faculty, and patient-versus-faculty ratings for the 56 interactions were -0.06 (P =.64), 0.31 (P <.02), and 0.45 (P <.0006), respectively. Pearson correlation coefficients approached significant correlation (0.70) between the faculty global rating and patient 360-degree scores (P =.08) but not with faculty 360-degree scores. Residents and faculty felt that completing the 360-degree forms was easy, but the requirement for faculty presence during the consent process was burdensome. CONCLUSION: Results from this pilot study suggest that self, faculty, and patient evaluations of resident performance constitutes a valid and reliable assessment of resident competence. Additional data are needed to determine whether the 360-degree assessment should be incorporated into residency programs and how frequently the assessment should be performed. Requiring only a specified number of assessments per rotation would make the process less burdensome for residents and faculty. PMID- 15288046 TI - Molecular systematics of African 20-chromosome toads (Anura: Bufonidae). AB - Intrageneric lineages and the historical biogeography of toads, genus Bufo, are poorly resolved due to their conservative morphology, their highly conserved karyotypes (typically 2N = 22), and erratic patterns of interspecific hybridisation. Here, we use mitochondrial and nuclear DNA sequence data to reconstruct relationships of the 20-chromosome toads, a major component of the African bufonid fauna. Mitochondrial 12S and 16S sequences from 29 species revealed two independent transitions between phenotypically distinct savannah and forest adapted forms. Analyses of mitochondrial 12S, 16S, and ND2, along with nuclear ACTC and Rhodopsin sequences from 12 species greatly increased bootstrap, and likelihood support for internal branches including a basal split into two pan African 20-chromosome clades. Hybridisation is a weak indicator of phylogenetic relationship as it occurs across these deeply divergent clades, between Bufo rangeri and Bufo gutturalis. These analyses suggest a secondary reversion to 22 chromosomes in Bufo pardalis, within the 20-chromosome group, although we could not reject an alternative hypothesis that this lineage forms a sister to all 2N = 20 toads. Other informally recognised 22-chromosome groups form independent phylogenetic lineages outside the 20-chromosome group, such as the Angusticeps and Vertebralis divisions. Bufo lindneri, from the Taitanus division, is closely related to Stephopaedes anotis, and these species should be considered congeners. PMID- 15288047 TI - Evidence for independent Hox gene duplications in the hagfish lineage: a PCR based gene inventory of Eptatretus stoutii. AB - Hox genes code for transcription factors that play a major role in the development of all animal phyla. In invertebrates these genes usually occur as tightly linked cluster, with a few exceptions where the clusters have been dissolved. Only in vertebrates multiple clusters have been demonstrated which arose by duplication from a single ancestral cluster. This history of Hox cluster duplications, in particular during the early elaboration of the vertebrate body plan, is still poorly understood. In this paper we report the results of a PCR survey on genomic DNA of the pacific hagfish Eptatretus stoutii. Hagfishes are one of two clades of recent jawless fishes that are an offshoot of the early radiation of jawless vertebrates. Our data provide evidence for at least 33 distinct Hox genes in the hagfish genome, which is most compatible with the hypothesis of multiple Hox clusters. The largest number, seven, of distinct homeobox fragments could be assigned to paralog group 9, which could imply that the hagfish has more than four clusters. Quartet mapping reveals that within each paralog group the hagfish sequences are statistically more closely related to gnathostome Hox genes than with either amphioxus or lamprey genes. These results support two assumptions about the history of Hox genes: (1) The association of hagfish homeobox sequences with gnathostome sequences suggests that at least one Hox cluster duplication event happened in the stem of vertebrates, i.e., prior to the most recent common ancestor of jawed and jawless vertebrates. (2) The high number of paralog group 9 sequences in hagfish and the phylogenetic position of hagfish suggests that the hagfish lineage underwent additional independent Hox cluster/-gene duplication events. PMID- 15288048 TI - Polyploid origins in a circumpolar complex in Draba (Brassicaceae) inferred from cloned nuclear DNA sequences and fingerprints. AB - Polyploid evolution has been of major importance in the arctic flora, but rarely addressed on the full circumpolar scale. Herein we study the allopolyploid Draba lactea and its close allies, which form a taxonomically intricate arctic-alpine complex including diploids, tetraploids, and hexaploids. Based on samples from the entire circumpolar area, we inferred the origins of polyploids in this complex using cloned DNA sequences from two nuclear regions (one intron from a gene encoding a second largest subunit in the RNA polymerase family, RPD2, and the ribosomal internal transcribed spacer region, ITS) and DNA fingerprints (random amplified polymorphic DNAs, RAPDs). Although D. lactea and all other polyploids examined in Draba are genetic alloploids showing fixed heterozygosity, the data obtained in the present study suggest that each of the polyploids analyzed here may have originated from a single diploid lineage: hexaploid D. lactea via tetraploid D. lactea from the D. palanderiana lineage (not from the D. fladnizensis and D. nivalis lineages as previously hypothesized), the tetraploid D. turczaninovii from the D. fladnizensis lineage, the tetraploid D. porsildii from the D. lonchocarpa lineage, and a tetraploid here named Draba spB from the D. nivalis lineage. Draba lactea has probably originated several times in the Beringian area, and it is not necessary to invoke complex origins based on a combination of different species lineages as previously suggested. PMID- 15288049 TI - Critical analysis of the topology and rooting of the parabasalian 16S rRNA tree. AB - The morphological classification of the protozoan phylum Parabasala is not in absolute agreement with the 16S rRNA phylogeny. However, there are strong indications that tree-construction artifacts play a considerable role in the shaping of the 16S rRNA tree. We have performed rigorous analyses designed to minimize such artifacts using the slow-fast and taxa-exclusion methods. The analyses, which included new sequences from the genera Monocercomonas and Hexamastix, in most respects confirmed the previously suggested tree topology and polyphyly of Hypermastigida and Monocercomonadidae but detected one artificial cluster of long branches (Trichonymphidae, Pseudotrichonymphidae, Hexamastix, and Tricercomitus). They also indicated that the rooting of the phylum on the trichonymphid branch is probably wrong and that reliable rooting on the basis of current data is likely impossible. We discuss the tree topology in the view of anagenesis of cytoskeletal and motility organelles and suggest that a robust taxonomic revision requires extensive analysis of other gene sequences. PMID- 15288050 TI - Molecular phylogeny and character evolution in the Western Palaearctic Helicidae s.l. (Gastropoda: Stylommatophora). AB - In this study, we present a molecular phylogeny for the west Palaearctic Helicidae sensu lato based on sequence data from two mitochondrial (COI, 16S rDNA) and two nuclear (ITS-1, 18S rDNA) genes. Maximum likelihood analysis and Bayesian inference revealed well supported monophyletic clades partly conflicting traditional classifications. Based on these results, we propose the following system. The Western Palaearctic Helicidae s.l. consist of two families, Helicidae and Hygromiidae. Within the Helicidae, three well supported subfamilies can be recognised: the Helicinae, Ariantinae, and Helicodontinae. The Hygromiidae consist of three clades: the Hygromiinae, the Helicellinae, and a yet unnamed clade comprising the genera Sphincterochila and Cochlicella. We then used the phylogeny to study the evolution of anatomical, and ecological characters traditionally used for systematic classification. In the Helicidae s.l., two independent evolutionary transitions to life in xeric environments occurred, which allowed the occupation of new niches with a subsequent radiation of the Helicellinae-Cochlicella/Sphincterochila clade and the Helicinae. Whereas, the multiplication of the Glandulae mucosae is a synapomorphy of the Hygromiidae, the lovedart sac apparatus is present in all groups and thus, the trait cannot provide a synapomorphy for either families or subfamilies. Additionally, we evaluated the use of structural molecular genetic characters for taxonomic assessment. The presence of an unique loop region of the 16S rDNA gene and a short tandem repeat in the ITS-1 region provide independent evidence for the monophyly of these major two groups, and can be used for preliminary classification. PMID- 15288051 TI - Population structure and history of a phenotypically variable teiid lizard (Ameiva chrysolaema) from Hispaniola: the influence of a geologically complex island. AB - Ameiva chrysolaema is distributed across the island of Hispaniola in the West Indies. The species is restricted to dry lowlands between major mountain ranges and along the southern and eastern coasts. Phylogenetic and phylogeographic analyses of mtDNA sequence variation from 14 sampling localities identify at least three independent evolutionary lineages, separated from one another by major mountain ranges. Nested clade phylogeographic analysis (NCPA) suggests a complex history of population fragmentation, consistent with geological evidence of seawater incursions into the Azua and Enriquillo basins during the Pliocene/Pleistocene (approximately 1.6 mya). Significantly negative Fu's F(S) values and parameters of mismatch distributions suggest that formerly fragmented populations have recently expanded their ranges. Significantly large average population clade distances (APCD) for two sampling localities in the Azua basin suggest secondary contact at these localities of previously separated populations. The distribution of haplotypes among polymorphic populations of A. chrysolaema suggests that variation in dorsal pattern represents a polymorphism within evolutionary lineages. Ameiva leberi is ecologically indistinguishable from and syntopic with A. chrysolaema. Genetic data suggest that A. leberi is a junior synonym of A. chrysolaema. PMID- 15288052 TI - A molecular test of alternative hypotheses of tetraodontiform (Acanthomorpha: Tetraodontiformes) sister group relationships using data from the RAG1 gene. AB - Two primary competing hypotheses regarding the identity of the sister group of the order Tetraodontiformes exist. The first hypothesis holds that some or all acanthuroid fishes represent the sister of Tetraodontiformes. The second, proposed in 1984 by Rosen, holds that the order Zeiformes is sister to Tetraodontiformes and that the family Caproidae is sister to this Zeiformes + Tetraodontiformes clade. These two hypotheses were tested using data from the single-copy nuclear gene RAG1. Representatives of most major orders of acanthomorph fishes were included to provide an appropriate context in which to place Tetraodontiformes and its hypothesized sister groups. The results of an unweighted parsimony analysis indicate that Zeiformes is not the sister group of Tetraodontiformes. In addition, Caproidae appears unrelated to Zeiformes. A monophyletic Tetraodontiformes was recovered as the sister group of the clade Ephippidae + Drepanidae and was more distantly related to the included zeiform and caproid representatives. PMID- 15288053 TI - Evolution of Galapagos Island Lava Lizards (Iguania: Tropiduridae: Microlophus). AB - Nucleotide sequences of mitochondrial genes (ND1, ND2, COI, and tRNAs) were determined for 38 samples representing 15 taxa of tropidurid lizards from the Galapagos Islands and mainland South America. Phylogenetically informative characters (759 of 1,956) were analyzed under Bayesian, maximum likelihood, and parsimony frameworks. This study supports the hypothesis that tropidurid lizards dispersed to the Galapagos on at least two separate occasions. One dispersal event involved an eastern Galapagos clade (Microlophus habelii and M. bivittatus, on Marchena and San Cristobal islands, respectively) the sister taxon of which is M. occipitalis from coastal Ecuador and Peru; the closest mainland relative of the western Galapagos clade was not unambiguously identified. The wide-ranging M. albemarlensis is revealed to be a complex of weakly divergent lineages that is paraphyletic with respect to the insular species M. duncanensis, M. grayii, and M. pacificus. PMID- 15288054 TI - Phylogenetic relationships within the Mysidae (Crustacea, Peracarida, Mysida) based on nuclear 18S ribosomal RNA sequences. AB - Species of the order Mysida (Crustacea, Peracarida) are shrimp-like animals that occur in vast numbers in coastal regions of the world. The order Mysida comprises 1,053 species and 165 genera. The present study covers 25 species of the well defined Mysidae, the most speciose family within the order Mysida. 18S rRNA sequence analysis confirms that the subfamily Siriellinae is monophyletic. On the other hand the subfamily Gastrosaccinae is paraphyletic and the subfamily Mysinae, represented in this study by the tribes Mysini and Leptomysini, consistently resolves into three independent clades, and hence is clearly not monophyletic. The tribe Mysini is not monophyletic either, and forms two clades of which one appears to be closely related to the Leptomysini. Our results are concordant with a number of morphological differences urging a taxonomic revision of the Mysidae. PMID- 15288055 TI - An investigation of the "Ancyloplanorbidae" (Gastropoda, Pulmonata, Hygrophila): preliminary evidence from DNA sequence data. AB - The Planorbidae is the largest family of freshwater pulmonate snails, yet an understanding of their intrafamily phylogenetic relationships is lacking and existing inferences are tentative. Moreover, it has been suggested that the Ancylidae, limpet-like freshwater pulmonates, should be merged with Planorbidae according to analysis of internal organ morphology. The present study explicitly tests this hypothesis by phylogenetic inference from partial DNA sequences of three molecular markers, nuclear ribosomal small subunit 18S and the mitochondrial cytochrome oxidase, and large subunit 16S. A molecular phylogeny was inferred based upon 22 taxa representing 12 ancylid and planorbid genera; additional taxa were included from the authors' database and from available sequences from GenBank, to further explore this basic data set. Taxa from Acroloxidae, Lymnaeidae, and Physidae were used as outgroups. Ancylidae and Planorbidae were found to be paraphyletic, with Planorbidae including some members of Ancylidae. "Ancyloplanorbidae" was also found to be paraphyletic because Acroloxus (Acroloxidae) surprisingly was included. Burnupia was found to be ancestral to "Ancyloplanorbidae" (including Acroloxus). The following clades of Planorbidae were supported: Bulininae and Planorbinae, Biomphalarini (including Helisoma and Planorbarius), and Planorbini and Segmentini. PMID- 15288056 TI - Phylogeographic history of the yellow-necked fieldmouse (Apodemus flavicollis) in Europe and in the Near and Middle East. AB - The exact location of glacial refugia and the patterns of postglacial range expansion of European mammals are not yet completely elucidated. Therefore, further detailed studies covering a large part of the Western Palearctic region are still needed. In this order, we sequenced 972 bp of the mitochondrial DNA cytochrome b (mtDNA cyt b) from 124 yellow-necked fieldmice (Apodemus flavicollis) collected from 53 European localities. The aims of the study were to answer the following questions: Did the Mediterranean peninsulas act as the main refuge for yellow-necked fieldmouse or did the species also survive in more easterly refugia (the Caucasus or the southern Ural) and in Central Europe? What is the role of Turkey and Near East regions as Quaternary glacial refuges for this species and as a source for postglacial recolonisers of the Western Palearctic region? The results provide a clear picture of the impact of the quaternary glaciations on the genetic and geographic structure of the fieldmouse. This species survived the ice ages in two main refuges, the first one in the Italo-Balkan region; the second one in Turkey and the Near East regions. It is from the Balkan refuge that it recolonised all European regions at the end of the last glaciation. The Turkish and Near East populations are distinct from the European ones and they did not recolonise the Palearctic region probably because: (i) they were blocked by the Black Sea and the Caucasus, (ii) the long term presence of fieldmice populations in the Balkans prevented their expansion. These are genetically differentiated from the European and Russian ones and could be described as a particular subspecies. This result emphasises the importance of Turkey and the Near and Middle East regions as a refuge for Palearctic mammals. PMID- 15288057 TI - Discordant temporal and geographic patterns in maternal lineages of eastern North American frogs, Rana catesbeiana (Ranidae) and Pseudacris crucifer (Hylidae). AB - Whether the Pleistocene has had a disproportionate impact on the recent diversification of temperate species, or played a lesser role in a more protracted process, has been a prominent evolutionary debate for the past decade. We used cytochrome b sequences to reconstruct the evolutionary histories of two widely co-distributed, and ecologically divergent frogs (Rana catesbeiana and Pseudacris crucifer) to examine the role of the Pleistocene in structuring these species. Results for R. catesbeiana reflect a pattern of allopatric fragmentation, likely in Coastal Plain refugia on either side of the Mississippi River dating to the mid to early Pleistocene. In contrast, P. crucifer contains numerous divergent lineages, including one west of the Mississippi River in the Interior Highlands, and in the east, multiple lineages that likely expanded from a number of southern Appalachian refugia with lineage sundering originating in the late Pliocene. Large-scale phylogeographic comparisons between these and other eastern North American species reflect both congruent and independent patterns of diversification, possibly reflecting the relative importance of dispersal ability and habitat associations. Although intra-lineage diversification has been structured by repeated Pleistocene glaciations, lineage sundering likely dates at least to the Pliocene in most (but not all) northern temperate amphibian and reptile species studied to date. PMID- 15288058 TI - Molecular systematics of Acarus siro s. lat., a complex of stored food pests. AB - The astigmatid mite Acarus siro (Linnaeus 1758) is an important agricultural pest and environmental allergen. However, it is likely that many mites described in the literature as A. siro, collected from both outdoor and stored product habitats, may belong to one of its sibling species, A. farris [Ent. Ber. Amst. 2 (26) (1905) 20] or A. immobilis [Bull. Br. Mus. Nat. Hist. 11 (1964a) 413; Acarologia. 6 (Suppl) (1964) 101]. The three species are difficult to separate morphologically, gene exchange between some of them is possible and, although each species displays environmental preferences, they occur together in some environments. This raises a question about their separate species status. In a pilot study, we investigated whether genetic data supported the separate species status of these forms. Both nuclear (the second internal transcribed spacer region [ITS-2] of the ribosomal cistron) and mitochondrial (cytochrome oxidase subunit I, mtcoxI hereafter) loci were employed for this purpose. Mtcox1 data does not conflict the differentiation into three separate species and while the ITS2 data were problematic for this group of mites it suggested that a congener, Acarus gracilis [Ann. Mag. Nat. Hist. 10 (1957) 753], is basal to the A. siro species complex. PMID- 15288059 TI - Molecular phylogeny of Demospongiae: implications for classification and scenarios of character evolution. AB - An analysis of the phylogenetic relationships of the 13 orders of Demospongiae, based on 18S and C1, D1 and C2 domains of 28S rRNA (for, respectively, 26 and 32 taxa) has been performed. The class Demospongiae as traditionally defined is not found to be monophyletic. Instead, a clade comprising all demosponges except Homoscleromorpha is well-supported, and we define phylogenetically the name Demospongiae in this more restricted sense to preclude the possibility of drastic alterations of the meaning of Demospongiae in the future, depending on the position of Homoscleromorpha. Within this clade Demospongiae s.s., ceractinomorphs and tetractinomorphs are polyphyletic, implying homoplastic evolution of characters such as reproductive strategies (viviparity vs. oviparity) and skeleton architecture (reticulate vs. radiate). The topology derived from our molecular data provides a basis for proposing a new classification of Demospongiae s.s., and suggests a reverse polarity of some characters, with respect to traditional conceptions: viviparity, presence of monaxon spicules and of spongin appear to be ancestral, whereas oviparity, and presence of tetraxon spicules appear as derived characters. PMID- 15288060 TI - Molecular phylogenetics and biogeography of Neotropical tanagers in the genus Tangara. AB - Species in the genus Tangara are distributed throughout the New World tropics and vary in their morphology, behavior, and ecology. We used data from the cytochrome b and ND 2 genes to provide the first phylogenetic perspective on the evolution of this diversity. Reconstructions based on parsimony, maximum likelihood, and Bayesian approaches were largely congruent. The genus is monophyletic and consists of two main clades. Within these clades, DNA sequence data confirm the monophyly of most previously recognized species groups within Tangara, indicating general concordance between molecular data and impressions based on geographic distribution, morphology, and behavior. Within some currently recognized species, levels of DNA sequence variation are larger than expected, suggesting multiple taxa may be involved. In contrast, some currently recognized species are only weakly differentiated from their sister species. Biogeographic analyses indicate that many early speciation events occurred in the Andes. More recently, dispersal events followed by subsequent speciation have occurred in other geographic areas of the Neotropics. Assuming a molecular clock, most speciation events occurred well before Pleistocene climatic cycles. The time frame of Tangara speciation corresponds more closely to a period of continued uplift in the Andes during the late Miocene and Pliocene. PMID- 15288061 TI - Phylogeny and host-specificity of European seed beetles (Coleoptera, Bruchidae), new insights from molecular and ecological data. AB - We used partial sequences of three mitochondrial genes (12S rRNA, cytochrome b, and cytochrome c oxidase subunit I) to reconstruct the phylogeny of European seed beetles (Bruchidae) belonging to the genera Bruchus Linnaeus and Bruchidius Schilsky. Adult beetles examined in this study were obtained from larvae bred from seeds directly collected in the field. Parsimony, maximum likelihood, and Bayesian inference were used to infer phylogenetic relationships among species. Both genera, Bruchidius and Bruchus, formed monophyletic groups in all analyses. Our results were partially in discrepancy with existing taxonomic groups (Borowiec, 1987). Critical analysis of relationships among taxa, and exhaustive review of host-plants data highlight the very high level of specialization of these seed beetles. Phylogenetically related insects were associated with host plants belonging to the same botanical tribes. PMID- 15288062 TI - MtDNA phylogeny and biogeography of Copelatinae, a highly diverse group of tropical diving beetles (Dytiscidae). AB - Copelatinae is a diverse lineage of diving beetles (Dytiscidae) frequently encountered in wet tropical and subtropical forests, but phylogenetic relationships are very poorly understood. We performed a phylogenetic and biogeographic analysis of this worldwide distributed group based on 50 species including a representative sample of major taxonomic groups and biogeographical regions. DNA sequences were obtained for the mitochondrial genes cytochrome oxidase I, cytochrome b, and 16S rRNA, for a total of 1575 aligned nucleotide positions. We found Copelatinae to be monophyletic, placed in a derived position and not sister to all remaining dytiscids, as had been suggested by earlier authors. The largest genus, Copelatus with some 460 known species was paraphyletic with respect to the smaller genera Lacconectus and Aglymbus. Among the major lineages of Copelatus, the subgenus Papuadytes was consistently recovered as sister to all other species (including Lacconectus and Aglymbus) with the possible exception of two western Palearctic taxa. We propose that the subgenus Papuadytes is removed from Copelatus and assigned generic status. Likewise, the two western Palearctic Copelatus are removed from this genus, and assigned the available genus name Liopterus. Our best phylogenetic hypothesis retrieved Afrotropical and New Guinean plus Australian species of Copelatus as monophyletic. Asian species were paraphyletic with respect to a species from Sulawesi which grouped with the species from New Guinea. Asian species were also paraphyletic with respect to Oriental Lacconectus, which was grouped with a clade of Neotropical species. Neotropical Copelatus form at least two separate lineages. The biogeographical evolution of Papuadytes is consistent with the relative age of the landmasses in the Austral region. Basal species are Australian, and successively derived ones are from New Caledonia and New Guinea. One species apparently dispersed from New Caledonia to China. Assuming a molecular clock and using a standard calibration of 2% divergence/MY the origin of Copelatinae is estimated to be between 85 and 95 MY. PMID- 15288063 TI - Phylogenetic investigations of Antarctic notothenioid fishes (Perciformes: Notothenioidei) using complete gene sequences of the mitochondrial encoded 16S rRNA. AB - The Notothenioidei dominates the fish fauna of the Antarctic in both biomass and diversity. This clade exhibits adaptations related to metabolic function and freezing avoidance in the subzero Antarctic waters, and is characterized by a high degree of morphological and ecological diversity. Investigating the macroevolutionary processes that may have contributed to the radiation of notothenioid fishes requires a well-resolved phylogenetic hypothesis. To date published molecular and morphological hypotheses of notothenioids are largely congruent, however, there are some areas of significant disagreement regarding higher-level relationships. Also, there are critical areas of the notothenioid phylogeny that are unresolved in both molecular and morphological phylogenetic analyses. Previous molecular phylogenetic analyses of notothenioids using partial mtDNA 12S and 16S rRNA sequence data have resulted in limited phylogenetic resolution and relatively low node support. One particularly controversial result from these analyses is the paraphyly of the Nototheniidae, the most diverse family in the Notothenioidei. It is unclear if the phylogenetic results from the 12S and 16S partial gene sequence dataset are due to limited character sampling, or if they reflect patterns of evolutionary diversification in notothenioids. We sequenced the complete mtDNA 16S rRNA gene for 43 notothenioid species, the largest sampling to-date from all eight taxonomically recognized families. Phylogenetic analyses using both maximum parsimony and maximum likelihood resulted in well-resolved trees with most nodes supported with high bootstrap pseudoreplicate scores and significant Bayesian posterior probabilities. In all analyses the Nototheniidae was monophyletic. Shimodaira-Hasegawa tests were able to reject two hypotheses that resulted from prior morphological analyses. However, despite substantial resolution and node support in the 16S rRNA trees, several phylogenetic hypotheses among closely related species and clades were not rejected. The inability to reject particular hypotheses among species in apical clades is likely due to the lower rate of nucleotide substitution in mtDNA rRNA genes relative to protein coding regions. Nevertheless, with the most extensive notothenioid taxon sampling to date, and the much greater phylogenetic resolution offered by the complete 16S rRNA sequences over the commonly used partial 12S and 16S gene dataset, it would be advantageous for future molecular investigations of notothenioid phylogenetics to utilize at the minimum the complete gene 16S rRNA dataset. PMID- 15288064 TI - Phylogeny of mysticete whales based on mitochondrial and nuclear data. AB - Mysticetes or baleen whales are comprised of four groups: Eschrichtiidae, Neobalaenidae, Balaenidae, and Balaenopteridae. Various phylogenetic hypotheses among these four groups have been proposed. Previous studies have not satisfactorily determined relationships among the four groups with a high degree of confidence. The objective of this study is to determine the relationships among the mysticete whales. Mitochondrial and nuclear DNA were sequenced for phylogenetic analysis. Most species relationships determined using these data were well resolved and congruent. Balaenidae is the most basal group and Neobalaenidae is the second most basal and sister group to the balaenopterid eschrichtiid clade. In this phylogenetic study, the resolution of Eschrichtiidae with two main lineages of Balaenopteridae was problematic. Some data partitions placed this group within the balaenopterids, and other partitions placed it as a sister taxon to the balaenopterids. An additive likelihood approach was used to determine the most optimal trees. Although it was not found in the combined phylogenetic analyses, the "best" tree found under the additive likelihood approach was one with a monophyletic Balaenopteridae. PMID- 15288065 TI - Mitochondrial DNA phylogenetic structuring suggests similarity between two morphologically plastic genera of Australian freshwater mussels (Unionoida: Hyriidae). AB - In the Lake Eyre Basin, the Australian hyriid genus Velesunio is represented by three undescribed species, each of which are highly divergent genetically, but morphologically similar to Velesunio wilsonii (Lea 1859). A fourth species, Velesunio ambiguus (Philippi 1847), occurs not only in the Lake Eyre Basin but throughout much of eastern Australia, including the Murray-Darling Basin. In this study, we show that another hyriid, Alathyria jacksoni (Iredale 1934), which is sympatric with V. ambiguus, is genetically deeply nested within the Velesunio species complex, such that the genus Velesunio is paraphyletic with respect to A. jacksoni. Moreover, our mitochondrial phylogenetic reconstructions indicate that A. jacksoni is closely allied to one of the cryptic Velesunio species occurring in the Lake Eyre Basin, but distinct from V. ambiguus and the two other Velesunio species. These data suggest that the genera Alathyria and Velesunio are in need of revision. The shells of A. jacksoni and Velesunio spp. vary with local conditions and sometimes are difficult to distinguish. Our analyses also show that shell characters of these taxa do not closely match the phylogenetic data, and it appears that the traditional taxonomic emphasis on these plastic characters has obscured evolutionary relationships between these, and possibly other, Hyriidae. PMID- 15288066 TI - Relative character-state space, amount of potential phylogenetic information, and heterogeneity of nucleotide and amino acid characters. AB - We examined a broad selection of protein-coding loci from a diverse array of clades and genomes to quantify three factors that determine whether nucleotide or amino acid characters should be preferred for phylogenetic inference. First, we quantified the difference in observed character-state space between nucleotides and amino acids. Second, we quantified the loss of potential phylogenetic signal from silent substitutions when amino acids are used. Third, we used the disparity index to quantify the relative compositional heterogeneity of nucleotides and amino acids and then determined how commonly convergent (rather than unique) shifts in nucleotide and amino acid composition occur in a phylogenetic context. The greater potential phylogenetic signal for nucleotide characters was found to be enormous (on average 440% that of amino acids), whereas the greater observed character-state space for amino acids was less impressive (on average 150.4% that of nucleotides). While matrices of amino acid sequences had less compositional heterogeneity than their corresponding nucleotide sequences, heterogeneity in amino acid composition may be more homoplasious than heterogeneity in nucleotide composition. Given the ability of increased taxon sampling to better utilize the greater potential phylogenetic signal of nucleotide characters and decrease the potential for artifacts caused by heterogeneous nucleotide composition among taxa, we suggest that increased taxon sampling be performed whenever possible instead of restricting analyses to amino acid characters. PMID- 15288067 TI - A molecular phylogeny of aquatic gastropods provides a new perspective on biogeographic history of the Snake River Region. AB - Mitochondrial DNA sequences of aquatic gastropods of the subgenus Pyrgulopsis (Natricola) were analyzed to test a commonly accepted hypothesis concerning the early history of the Snake River in the northwestern US. Distributions of Natricola and other regional biota were previously used to infer that the Snake River flowed to the Pacific through southeastern Oregon and northern California during the Neogene prior to its capture by the Columbia River in the late Pliocene (2 Ma). A molecular phylogeny based on partial sequences of COI and NDI (1149 bp) indicates that the Natricola clade is restricted to the modern Snake Columbia River Basin and the Oregon Lakes region whereas northern California populations previously assigned to this subgenus belong to other lineages. The Natricola clade is not deeply subdivided into Oregon Lakes and Snake River Basin units consistent with late Pliocene fragmentation of the hypothesized paleodrainage, but instead is shallowly structured and contains multiple transitions among these two geographic areas. The strongly supported sister relationship between Natricola and a species from northwest Nevada (P. imperialis) is consistent with a recent proposal that the ancestral Snake River did not flow through southeast Oregon but instead flowed south to the Humboldt River. Within the context of this hypothesis, the multiple transitions between the Snake River Basin and the Oregon Lakes region that occurred within Natricola may be attributed to a late Pleistocene connection between these areas that was unrelated to the early course of the Snake River. PMID- 15288068 TI - A molecular perspective on the phylogeny of the Hyla pulchella species group (Anura, Hylidae). AB - A molecular phylogenetic analysis of the Hyla pulchella species group was performed to test its monophyly, explore the interrelationships of its species, and evaluate the validity of the taxa that were considered subspecies of H. pulchella. Approximately 2.8 kb from the mitochondrial genes 12s, tRNA valine, 16s, and Cytochrome b were sequenced. The analysis included 50 terminals representing 10 of the 14-15 species currently recognized in the H. pulchella group, including samples from several localities for some taxa, several outgroups, as well as two species previously suspected to be related with the group (Hyla guentheri and Hyla bischoffi). The results show that the H. pulchella and Hyla circumdata groups are distantly related, and, therefore, should be recognized as separate groups. As currently defined, the H. pulchella group is paraphyletic with respect to the Hyla polytaenia group; therefore, we recognize the Hyla polytaenia clade in the H. pulchella group. Two subspecies of H. pulchella recognized by some authors are considered full species including Hyla pulchella riojana because it is only distantly related to H. pulchella, and Hyla pulchella cordobae because molecular and non-molecular evidence suggests that it is specifically distinct. With the inclusion of the H. polytaenia clade, H. guentheri, and H. bischoffi, and the recognition of the two former subspecies of H. pulchella as distinct species, the H. pulchella group now comprises 25 described species. All representatives of the H. pulchella group with an Andean distribution are monophyletic and nested within a clade from the Atlantic forest from south-southeastern Brazil/northeastern Argentina, and Cerrado gallery forest from central Brazil. PMID- 15288069 TI - Generic delimitation and phylogenetic relationships within the subtribe Chironiinae (Chironieae: Gentianaceae), with special reference to Centaurium: evidence from nrDNA and cpDNA sequences. AB - To better understand the evolutionary history of the genus Centaurium and its relationship to other genera of the subtribe Chironiinae (Gentianaceae: Chironieae), molecular analyses were performed using 80 nuclear ribosomal ITS and 76 chloroplast trnLF (both the trnL UAA intron and the trnL-F spacer) sequences. In addition, morphological, palynological, and phytochemical characters were included to a combined data matrix to detect possible non-molecular synapomorphies. Phylogenetic reconstructions support the monophyly of the Chironiinae and an age estimate of ca. 22 million years for the subtribe. Conversely, both molecular data sets reveal a polyphyletic Centaurium, with four well-supported main clades hereafter treated as separate genera. The primarily Mediterranean Centaurium s.s. is closely related to southern African endemics Chironia and Orphium, and to the Chilean species Centaurium cachanlahuen. The resurrected Mexican and Central American genus Gyrandra is closely related to Sabatia (from eastern North America). Lastly, the monospecific genus Exaculum (Mediterranean) forms a monophyletic group together with the two new genera: Schenkia (Mediterranean and Australian species) and Zeltnera (all other indigenous American centauries). Several biogeographical patterns can be inferred for this group, supporting a Mediterranean origin followed by dispersals to (1) North America, Central America, and South America, (2) southern Africa (including the Cape region), and (3) Australia and Pacific Islands. PMID- 15288070 TI - The mitochondrial genome of the firefly, Pyrocoelia rufa: complete DNA sequence, genome organization, and phylogenetic analysis with other insects. AB - The complete nucleotide sequences of the mt genome from the firefly, Pyrococelia rufa (Coeleoptera: Lampyridae) was determined. The circular genome is 17,739-bp long, and contains a typical gene complement, order, and arrangement identical to Drosophila yacuba. The presence of 1,724-bp long intergenic spacer in the P. rufa mt genome is unique. The putative initiation codon for ND1 gene appears to be TTG, instead of frequently found ATN. All tRNAs showed stable canonical clover leaf structure of other mt tRNAs, except for tRNA(Ser) (AGN), DHU arm of which could not form stable stem-loop structure. Phylogenetic analysis among insect orders confirmed a monophyletic Endopterygota, a monophyletic Mecopterida, a monophyletic Diptera, a monophyletic Lepidoptera, and a monophyletic Coleoptera, suggesting that the complete insect mt genome sequence has a resolving power in the diversification events within Endopterygota. However, internal relationships among three coleopteran species are not clear, and the inclusion of some insect orders (i.e., apterygotan T. gertschi) in the analysis provided inconsistent results compared to other molecular studies. PMID- 15288071 TI - Molecular phylogeny of the hexagrammid fishes using a multi-locus approach. AB - Ideally, organisms are grouped into monophyletic assemblages reflecting their evolutionary histories. Single (molecular) markers can reflect the evolutionary history of the marker, rather than the species in question, therefore, phylogenetic relationships should be inferred from adequate sampling of characters. Because the use of multiple loci greatly improves the resolving power of the molecular assay, we constructed a molecular phylogeny of the family Hexagrammidae based on six loci, including two mitochondrial and four nuclear loci. The resulting molecular phylogeny, from the combined data, was significantly different from the morphological topology suggested by Shinohara [Memoirs of the Faculty of Fisheries, Hokkaido University 41 (1994) 1]. Our data support a monophyletic assemblage for the genera Hexagrammos and Pleurogrammus. However, other taxa traditionally included in the family Hexagrammidae did not form a monophyletic assemblage. The monotypic genus Ophiodon was more closely associated with cottids than with other hexagrammids. Our data concur with the morphological topology in that the genera Zaniolepis and Oxylebius formed a monophyletic clade, which was distinct and basal to the remaining hexagrammids, seven cottids and one agonid. PMID- 15288072 TI - Linking phylogenetics with population genetics to reconstruct the geographic origin of a species. AB - Reconstructing ancestral geographic origins is critical for understanding the long-term evolution of a species. Bayesian methods have been proposed to test biogeographic hypotheses while accommodating uncertainty in phylogenetic reconstruction. However, the problem that certain taxa may have a disproportionate influence on conclusions has not been addressed. Here, we infer the geographic origin of Drosophila simulans using 2,014 bp of the period locus from 63 lines collected from 18 countries. We also analyze two previously published datasets, alcohol dehydrogenase related and NADH:ubiquinone reductase 75 kDa subunit precursor. Phylogenetic inferences of all three loci support Madagascar as the geographic origin of D. simulans. Our phylogenetic conclusions are robust to taxon resampling and to the potentially confounding effects of recombination. To test our phylogenetically derived hypothesis we develop a randomization test of the population genetics prediction that sequences from the geographic origin should contain more genetic polymorphism than those from derived populations. We find that the Madagascar population has elevated genetic polymorphism relative to non-Madagascar sequences. These data are corroborated by mitochondrial DNA sequence data. PMID- 15288073 TI - Molecular architecture of Pipistrellus pipistrellus/Pipistrellus pygmaeus complex (Chiroptera: Vespertilionidae): further cryptic species and Mediterranean origin of the divergence. AB - Previous genetic analyses have demonstrated that two phonic types of one of the most common European bats, the Common pipistrelle, belong to distinct species, although they are almost identical morphologically (45 kHz Pipistrellus pipistrellus and 55 kHz Pipistrellus pygmaeus). To reconstruct the history of the species complex and explain the codistribution of both forms in Europe and the Mediterranean, we performed phylogenetic analysis based on a 402-bp portion of the cytochrome b gene. Particular attention was paid to the eastern and southern parts of the range where no data were available. We found further distinctive allopatric haplotypes from Libya and Morocco. The difference of about 6-7% described in the Libyan population suggests the occurrence of a new species in the southern Mediterranean. The species status of Moroccan population is also discussed. The phylogeographic patterns obtained and analysis of fossil records support the hypothesis of expansion of both species into Europe from the Mediterranean region during the Holocene. The allopatric speciation model fits our data best. The paleobiographic scenario envisaged is corroborated also by molecular clock estimations and correlations with Late Neogene environmental changes in the Mediterranean region which ended with the Messinian salinity crisis. PMID- 15288074 TI - Contribution of RPB2 to multilocus phylogenetic studies of the euascomycetes (Pezizomycotina, Fungi) with special emphasis on the lichen-forming Acarosporaceae and evolution of polyspory. AB - Despite the recent progress in molecular phylogenetics, many of the deepest relationships among the main lineages of the largest fungal phylum, Ascomycota, remain unresolved. To increase both resolution and support on a large-scale phylogeny of lichenized and non-lichenized ascomycetes, we combined the protein coding-gene RPB2 with the traditionally used nuclear ribosomal genes SSU and LSU. Our analyses resulted in the naming of the new subclasses Acarosporomycetidae and Ostropomycetidae, and the new class Lichinomycetes, as well as the establishment of the phylogenetic placement and novel circumscription of the lichen-forming fungi family Acarosporaceae. The delimitation of this family has been problematic over the past century, because its main diagnostic feature, true polyspory (numerous spores issued from multiple post-meiosis mitoses) with over 100 spores per ascus, is probably not restricted to the Acarosporaceae. This observation was confirmed by our reconstruction of the origin and evolution of this form of true polyspory using maximum likelihood as the optimality criterion. The various phylogenetic analyses carried out on our data sets allowed us to conclude that: (1) the inclusion of phylogenetic signal from ambiguously aligned regions into the maximum parsimony analyses proved advantageous in reconstructing phylogeny; however, when more data become available, Bayesian analysis using different models of evolution is likely to be more efficient; (2) neighbor-joining bootstrap proportions seem to be more appropriate in detecting topological conflict between data partitions of large-scale phylogenies than posterior probabilities; and (3) Bayesian bootstrap proportion provides a compromise between posterior probability outcomes (i.e., higher accuracy, but with a higher number of significantly supported wrong internodes) vs. maximum likelihood bootstrap proportion outcomes (i.e., lower accuracy, with a lower number of significantly supported wrong internodes). PMID- 15288075 TI - Molecular phylogeny of two lineages of Leuciscinae cyprinids (Telestes and Scardinius) from the peri-Mediterranean area based on cytochrome b data. AB - We examined phylogenetic relationships in two lineages of Leuciscinae cyprinid fishes based on the sequence data of the complete mitochondrial DNA region coding for the cytochrome b gene (1140 bp). Telestes includes obligate riverine, moderately cold water-adapted species whereas Scardinius comprises warm-adapted species living in lowland lakes and still waters of rivers and streams. We also analysed selected representatives of Leuciscus and Phoxinellus because the taxonomic status of some species belonging to these genera is dubious and they could be placed in the genus Telestes. The study includes 18 species, 43 populations, and 111 individuals from 9 of the 14 peri-Mediterranean ichthyogeographic districts. Clades recovered from the phylogenetic analyses do not support previous taxonomic assumptions based on morphology. Telestes, Leuciscus, and Phoxinellus do not form monophyletic assemblages; phylogenetic analyses suggest that L. polylepis, L. turskyi, P. croaticus, and P. metohiensis should be included in Telestes. Similarly, populations of Scardinius erythrophthalmus do not cluster together and the endangered S. scardafa, endemic to central Italy and surviving in a single locality, is nested within them. The radiations of Telestes and Scardinius occurred in different time periods. A major diversification of Telestes is consistent with a sea dispersal during the freshwater Messinian "Lago Mare" phase of the Mediterranean Sea. Cladogenetic events within Scardinius are likely related to the extension and confluence of river drainages in lowlands following multiple lowering of the sea level during the Quaternary glaciations. PMID- 15288076 TI - The criterion of reciprocal monophyly and classification of nested diversity at the species level. PMID- 15288077 TI - [Treatment of acute infantile diarrhoea: practice is still far from recommendations]. PMID- 15288078 TI - [Management of acute infantile diarrhoea: a study on community pharmacy counseling in the Midi-Pyrenees region]. AB - Counselling by community pharmacists is becoming an accepted standard for pharmacy practice. However, drugs available in children without prescription form are scarce, and most of the over-the-counter drugs have not been tested and approved in children. OBJECTIVES: The aim of this study was to investigate attitude and knowledge of community pharmacists about advice and treatment in children with acute diarrhoea. MATERIAL AND METHODS: We sent a postal questionnaire to a sample of 176 community pharmacies in the Midi-Pyrenees area (South western France), asking what they would give as advice and/or drugs in a simulated case of acute diarrhoea in an eight-month-old baby. For each question (interview of the mother, counselling about hygiene and dietetics, monitoring and drugs), we compared pharmacists answers to available evidence-based data and/or guidelines in the literature and to Summary Products Characteristics (SPC) for each reported drug. Forty one percent of pharmacies answered, giving 101 exploitable questionnaires. RESULTS: Only 48.5% of subjects have recommended a rehydration solution. 71.3% recommended an inadequate beverage (soda) and 40% recommended stopping food intake despite WHO guidelines. Most of pharmacists (77%) noticed a drug with an appropriate indication and a paediatric mention in the SPC. However, in 12.9% of cases, drugs were contra-indicated or inadequate (loperamide, nifuroxazide, microorganisms available in capsules). CONCLUSION: Even if an appropriate advice was given by most of the responders, improvements in advice are needed: too many pharmacists recommended anti-diarrhoeal drugs and withholding milk despite evidence about their lack of effectiveness on dehydration prevention. Conversely, rehydration solutions, which have been proved their effectiveness since many years, are not sufficiently proposed. PMID- 15288079 TI - [Treatment of acute diarrhea: prescription patterns by private practice pediatricians]. AB - Acute gastroenteritis remains a frequent illness in infants and children with still important morbidity and mortality rates. Oral rehydratation solutions (ORS) and early refeeding are the main recommendations. Indication of drugs remains limited. OBJECTIVE: To evaluate the management of acute diarrhea by private practice pediatricians of France. METHODS: A questionnaire concerning ORS, dietary formula, antidiarrheal diet, antibiotherapy, antidiarrheal drugs was sent to all 2907 private pediatricians of France. RESULTS: Six hundred twenty-nine questionnaires were analyzed (22%). Three hundred and ninety-seven pediatricians (63%) prescribed systematically an ORS, 294 (47%) changed formula, 412 (66%) prescribed a regimen. Antibiotic was prescribed after coproculture (81%), when glairy and bloody diarrhea (65%), associated infectious disease (63%), toxi infectious syndrome (42%) or immunodeficiency were present (28%). Most pediatricians (97%) prescribed at least one drug: diosmectite (84%), Lactobacillus acidophilus (63%), Saccharomyces boulardii (62%), racecadotril (62%), loperamide (28%), attapulgite de Mormoiron (26%), nifuroxazide (20%). Drugs were prescribed more often for their effectiveness than for comfort. CONCLUSION: This study demonstrates the discrepancies that remain between recommendations and practical care in the treatment of acute diarrhea in children. Private French pediatricians often prescribe drugs. PMID- 15288080 TI - [Nosocomial viral infections in a pediatric service: example of rotaviral gastroenteritis and respiratory syncytial viral bronchiolitis]. AB - Nosocomial infections are a preoccupation in a pediatric hospital mainly during the winter with bronchiolitis and gastroenteritis epidemics. We have examined the risk factors of nosocomial infections. MATERIAL AND METHODS: A prospective study was conducted between November, 1999 and March, 2000 in the infants units of the Le Havre hospital. We systematically listed the admissions and contacted the family after their discharge by phone. A geographic information system was implemented to display the epidemiological data; this software is able to illustrate the sectors at risk. RESULTS: During the study, 687 infants were hospitalized of whom 458 for bronchiolitis and community-acquired gastroenteritis. Mean age was 5.4 months old. No nosocomial bronchiolitis occurred. Prevalence of nosocomial gastroenteritis was 10% (68 cases including nine after discharge). Infants with nosocomial infection were younger than those with community-acquired infection (6.6 months vs. 11.2 months, P < 0.01). The mean length of stay was longer in nosocomial infection (7.7 vs. 4.1 days, P < 0.05). Among the infants with bronchiolitis, 16% have developed nosocomial intestinal infections (RR = 2.65, IC: 1.59-4.4; P < 0.01). The geographic analysis pointed the area with nosocomial risk (bedroom without water, nearness of nurse office and games room). CONCLUSION: Geographic information system is a part of the quality control system and may have some interaction effect on final decision making. Incidence of nosocomial infections showed the need for a prevention strategy in a pediatric hospital. PMID- 15288081 TI - [Mortality in pediatric renal transplants: 15 years' experience]. AB - The aim of the study was to analyse retrospectively the patient survival and mortality risk factors in renal transplant children at Edouard-Herriot hospital, Lyon. PATIENTS AND METHODS: One hundred and ninety four patients 0 to less than 18 years of age received a renal transplant between April 1987 and January 2002. We collected data concerning the age at transplantation, the mean duration of dialysis prior to transplantation, the type of donor (living related: LRD, or cadaver: CAD) as well as long term follow-up of all the patients. Eleven patients died during the study period and we collected mortality and morbidity factors. RESULTS: Mean age at transplantation was 9.5 years (range: 0.6-17.9 years); 16 children were transplanted before 2 years of age. Eighty six percent received a first graft, 10% a second graft, and 3% a third graft. One third of the patients had preemptive transplantation. The graft was carried out respectively from a LRD in 18% and from CAD in 82% of the cases. Eleven patients (5.7%) died during this period; four of them were transplanted twice. Death occurred within 2.4 years after transplantation (range: 2 days-6.3 years). Five of these deceased children developed an acute rejection episode treated with methylprednisolone pulses and OKT3 monoclonal antibodies. Four had a cytomegalovirus (CMV) infection and three required repeated surgery. The causes of death were bacterial infections (4/11), CMV infection (2/11), neurological involvement (2/11), Burkitt lymphoma (1/11) and unexplained sudden death (2/11). CONCLUSION: The main causes of deaths in renal transplant children are bacterial infections. The improvement in patient survival probably will come from both better preventive approach to nosocomial infections and a less aggressive immunosuppression. PMID- 15288082 TI - [Analgesic effects of Emla cream and saccharose solution for subcutaneous injections in preterm newborns: a prospective study of 265 injections]. AB - OBJECTIVE: To compare the analgesic effects of non nutritive pacifier sucking, oral administration of a 30% saccharose solution, local application of Emla and their association for subcutaneous injection of erythropoietin (EPO) in preterm infants. METHODS: Our study was a randomised, prospective study conducted over 5 months. Neonates with a gestational age below 33 weeks of gestation and older than 8 days of life were included if they were treated with EPO (three subcutaneous injections per week during 6 weeks). For each consecutive EPO injection, patients were randomised between four groups of intervention: non nutritive pacifier sucking (T), oral administration of 0.2-0.5 ml of a 30% saccharose solution with non nutritive pacifier sucking (S), local application of Emla with non nutritive pacifier sucking (E), and oral administration of 0.2-0.5 ml of a 30% saccharose solution with local application of Emla and with non nutritive pacifier sucking (S + E). Each child was its own control. Pain was assessed with the Newborn Acute Pain scale (DAN) and with the Neonatal Facial Coding System (NFCS). RESULTS: Thirty-three neonates were included, representing 265 injections. Distribution was: 41 in group T, 71 in group E, 86 in group S and 67 in group E + S. Mean DAN and NFCS scores were statistically different between groups T, E and S. Analgesic effect of saccharose (-1.05) was greater than Emla ( 0.56). Used together, effects were adding up without potentialisation. CONCLUSION: This study shows that the association of non nutritive pacifier sucking with oral administration of saccharose and local application of Emla has a better analgesic effect than each of these three interventions alone for subcutaneous injection of EPO. PMID- 15288083 TI - [Congenital toxoplasmosis: a new case of infection during pregnancy in an previously immunized and immunocompetent woman]. AB - Congenital toxoplasmosis is a potentially serious infection which usually affects infants born to non immune women. CASE REPORT: Our case report focuses on a baby born to a normally immunocompetent woman previously immunized against toxoplasmosis. To our knowledge only three similar cases have been published until now. CONCLUSION: We conclude that in front of a patient neonatal congenital infection picture, toxoplasmosis cannot be excluded on the ground of maternal immunity status and must be quickly investigated, given the emergency of appropriate treatment. PMID- 15288084 TI - [Delayed discovery of congenital diaphragmatic hernia: diagnostic difficulties. A report of two cases]. AB - Delayed revelation of congenital diaphragmatic hernias (CDH) is not uncommon and can represent 5-30% of total CDHs. Time before diagnosis may be prolonged, sometimes to the adult period. Respiratory and gastrointestinal symptoms are frequent but not specific. The clinical presentation of delayed CDH may thus mislead the practitioner. Diagnosis can be approached and/or confirmed by plain radiography. Outcome is usually favorable after surgery. We report two cases of delayed CDH and we discuss the difficulty of diagnosis. PMID- 15288085 TI - [Neonatal accidental burn by isopropyl alcohol]. AB - Antiseptic solutions are commonly used for skin care but are not always safe. In preterm infants, application of antiseptic solutions can lead to serious burns. We report the case of a premature newborn who developed severe burns at 35 weeks post-conceptional age, after his mother used disposable antiseptic towels containing isopropyl alcohol for his skin care. Burns outcome led to diffuse skin burn and death of the baby. Several cases of isopropyl alcohol poisoning through skin absorption have been reported in neonate and infants. Because of its neonatal toxicity, isopropyl alcohol has been excluded from composition of antiseptic solutions commonly used in neonatology. However, isopropyl alcohol is still available in many housecleaning and cosmetic products, while its toxic effects in children are not clearly mentioned. A specific mention "toxic for infants and children" should appear on mass consumption products containing isopropyl alcohol. Moreover, health workers may individually inform parents about possible hazards of poisoning through skin absorption. PMID- 15288086 TI - [Neonatal lupus: different clinical neonatal expression in siblings]. AB - Neonatal Lupus Syndrome is a rare disease caused by placental passage of maternal autoantibodies. Pathogenesis is partially unknown and many clinical manifestations are possible. We report on newborn siblings who presented with different symptoms of Neonatal Lupus Syndrome. One patient presented with congenital heart block and another with hepatic and haematologic involvement. Cases of Neonatal Lupus among siblings are very rare, because of the high risk of pregnancy in affected women. Various clinical expressions may be explained by a different specificity of Anti-Ro autoantibodies among siblings. The reported cases are commented with regard to recent literature, trying to explain their pathogenesis. PMID- 15288087 TI - [Disseminated lupus erythematosus in children: guidelines about investigations during the initial evaluation and follow-up]. AB - Childhood-onset systemic lupus erythematosus (SLE) is often severe and has a serious long-term morbidity. Pediatric guidelines about its management do not exist. The French study group of childhood-onset SLE proposes recommendations about the investigation which are needed at diagnosis and during follow-up of SLE, in order to adjust the treatment according to the severity of the disease and to avoid unnecessary investigations. PMID- 15288088 TI - [Treatment of pulmonary arterial hypertension in children]. AB - Treatment strategies for pulmonary hypertension in children have dramatically evolved. Traditional therapy with calcium channel blockers and pulmonary transplantation is only indicated in selected patients and does not reduce mortality very significantly. New pulmonary vasodilators are emerging from recent trials in the adult population. Their indications are based on the patient's NYHA classification. The epoprostenol (prostacyclin, Flolan) has shown reduction in mortality and improvement in functional symptoms in pediatric patients. The frequent side effects and continuous intravenous infusion limit the indication of prostacyclin in NYHA class IV children. The endothelin receptor blocker bosentan (Tracleer) is an orally given agent. It improves functional symptoms in adults and hemodynamic measures in children. It can be started in children with moderate functional symptoms (NYHA class II and III). The type V phosphodiesterase inhibitor sildenafil (Viagra) is being evaluated and may represent a promising therapy in the future. Invasive strategies like catheter-based atrial septostomy may be useful in particular cases. Randomized-controlled studies are urgently needed to evaluate the safety and efficacy of these new therapies. PMID- 15288089 TI - [Movement disorders in childhood: therapeutic update]. AB - Abnormal movements are not uncommon in childhood. Due to the severity of the abnormal movements or to the functional disability, a medical treatment is often required; the wide range of available pharmacological molecules and the absence of therapeutic consensus highlight the limited efficacy of the medical treatment on dystonic or athetoid movements, or severe tic disorders. The recent identification of the enzymatic defect implicated in metabolic diseases led to the development of specific treatment for newly recognized disorders, with more or less interesting results (creatine ou biotine supplementation). Recent progress in functional neurosurgery opened new fields in the treatment of movement disorders. Intrathecal baclofen was proved effective in the treatment of secondary dystonia, especially in patients with cerebral palsy. Deep brain stimulation is now an established therapy for patients with a generalized dystonic syndrome. Given the successful results of pallidal stimulation in dystonia, the indication of this procedure has been discussed in other types of abnormal movements. PMID- 15288092 TI - [Kawasaki syndrome and Mycoplasma pneumoniae infection]. PMID- 15288093 TI - [Megaesophagus associated with growth hormone deficiency in a 14-year-old adolescent]. PMID- 15288094 TI - [Should diagnosis of attention deficit/hyperactivity disorder be limited to criteria of DSM IV classification?]. PMID- 15288095 TI - [Are IgE-independent food hypersensitivity and chronic fatigue syndrome related?]. PMID- 15288096 TI - [Compliance to atopia preventive regimes after discharge from maternity services]. PMID- 15288097 TI - [Rubinstein-Taybi syndrome: a familial form ]. PMID- 15288098 TI - [Jacques Cartier Conference: Vaccines. 1-2 December 2003, Lyon, France. Proceedings]. PMID- 15288113 TI - [Treatment in 2003 of septic shock in children in the first two hours (excluding newborns)]. AB - Despite new understandings in pathophysiology, sepsis mortality remains high in children. Recently, it has been demonstrated that early goal directed therapy may decrease septic shock mortality. The aim of this paper is to propose practical clinical guidelines based on earlier consensus recommendations. Septic shock must be rapidly suspected and early recognized. Bases of treatment are maintenance of adequate oxygenation with use of artificial ventilation if necessary, larger and faster volume resuscitation than recommended before, empiric antibiotherapy and early use of vasopressive agents associated with corticosteroids in particular situations. Treatment efficacy must be regularly assessed during first hours of resuscitation. Taking into account pediatric particularities and results of adult studies, pediatricians who take care of children at beginning of septic shock may reasonably hope to decrease mortality if they keep in mind specific therapeutic goals. PMID- 15288114 TI - [Climate change and bacterial disease]. PMID- 15288115 TI - Role of oxidative damage in the genotoxicity of arsenic. AB - Arsenic is a well-established human carcinogen and is ubiquitous in the environment. For decades, arsenic has been considered to be a nongenotoxic carcinogen because it is only weakly active or, more often, completely inactive in bacterial and mammalian cell mutation assays. In this review, evidence is presented that when assayed using model systems in which both intragenic and multilocus mutations can readily be detected, arsenic is, indeed, found to be a strong, dose-dependent mutagen which induces mostly multilocus deletions. Furthermore, the roles of reactive oxygen and reactive nitrogen species in mediating the genotoxic response are presented in a systematic and logical fashion in support of a working model. The data suggest that antioxidants may be a useful interventional treatment in reducing the deleterious effects of arsenic. PMID- 15288116 TI - Oxidative stress and apoptosis in metal ion-induced carcinogenesis. AB - Epidemiological evidence suggests that exposure to certain metals causes carcinogenesis. The mechanisms of metal-induced carcinogenesis have been pursued in chemical, biochemical, cellular, and animal models. Significant evidence has accumulated that oxidative stress may be a common pathway in cellular responses to exposure to different metals. For example, in the last few years evidence in support of a correlation between the generation of reactive oxygen species, DNA damage, tumor promotion, and arsenic exposure has strengthened. This article summarizes the current literature on metal-mediated oxidative stress, apoptosis, and their relation to metal-mediated carcinogenesis, concentrating on arsenic and chromium. PMID- 15288117 TI - Introduction to serial reviews on 4-hydroxy-2-nonenal as a signaling molecule. PMID- 15288118 TI - 4-hydroxynonenal and regulation of cell cycle: effects on the pRb/E2F pathway. AB - The hypothesis that 4-hydroxynonenal (HNE), a product of lipid peroxidation, might negatively affect cell proliferation, arose from the observation that lipid peroxidation is very low in tumors. In leukemic cells HNE inhibited cell growth and reduced c-myc and c-myb expression. HNE also induced differentiation in different leukemic cell lines. In HL-60 human leukemic cells, HNE induced the accumulation of cells in the G(0)/G(1) phase of the cell cycle accompanied by a decrease of cyclins D1, D2, and A. Moreover, HNE caused an increase in p21 expression. As cyclin D/CDK2 and cyclin A/CDK2 phosphorylate pRB, these findings suggested that pRb phosphorylation could be affected by HNE. Hypophosphorylated pRb binds and inactivates the E2F transcription factors. HNE induced the dephosphorylation of pRb and the increase in pRb/E2F1 complexes, whereas pRb/E2F4 complexes were reduced, because HNE downregulated E2F4 protein expression. The analysis of E2F binding to the P2 c-myc promoter revealed that HNE caused a decrease in "free" E2F, as well as an increase in pRb (and pRB family members) bound to E2F, with consequent repression of the transcription. In conclusion, HNE reduces E2F transcriptional activity by modifying a number of genes involved in regulation of the pRb/E2F pathway. PMID- 15288119 TI - Regulation of 4-hydroxynonenal-mediated signaling by glutathione S-transferases. AB - 4-Hydroxynonenal (HNE), one of the major end products of lipid peroxidation, has been shown to be involved in signal transduction and available evidence suggests that it can affect cell cycle events in a concentration-dependent manner. Glutathione S-transferases (GSTs) can modulate the intracellular concentrations of HNE by affecting its generation during lipid peroxidation by reducing hydroperoxides and also by converting it into a glutathione conjugate. We have recently demonstrated that overexpression of the Alpha class GSTs in cells leads to lower steady-state levels of HNE, and these cells acquire resistance to apoptosis induced by lipid peroxidation-causing agents such as H(2)O(2), UVA, superoxide anion, and pro-oxidant xenobiotics, suggesting that signaling for apoptosis by these agents is transduced through HNE. Cells with the capacity to exclude HNE from the intracellular environment at a faster rate are relatively more resistant to apoptosis caused by H(2)O(2), UVA, superoxide anion, and pro oxidant xenobiotics as well as by HNE, suggesting that HNE may be a common denominator in mechanisms of apoptosis caused by oxidative stress. We have also shown that transfection of adherent cells with HNE-metabolizing GSTs leads to transformation of these cells due to depletion of HNE. These recent studies from our laboratories, which strongly suggest that HNE is a key signaling molecule and that GSTs, being determinants of its intracellular concentrations, can regulate stress-mediated signaling, are reviewed in this article. PMID- 15288120 TI - Alpha-tocopherol amplifies phosphorylation of endothelial nitric oxide synthase at serine 1177 and its short-chain derivative trolox stabilizes tetrahydrobiopterin. AB - Alpha-tocopherol has been shown to increase nitric oxide (NO)-dependent relaxation but the underlying mechanisms have not been fully characterized. The present study investigates the effect of alpha-tocopherol and its derivative trolox on the synthesis of NO in human umbilical vein endothelial cells. NO was assayed as citrulline (co-product of NO) and cGMP (product of the NO-activated soluble guanylate cyclase) on ionomycin stimulation of cells. Ionomycin induced citrulline and cGMP formation partially through phosphorylation of endothelial NO synthase (eNOS) at its serine residue 1177, which was mediated mainly by calmodulin-dependent kinase II. Preincubation of cells with alpha-tocopherol or trolox increased eNOS activity in a concentration-dependent manner without changing eNOS expression. The effect of the water-soluble trolox was due to chemical stabilization of the eNOS cofactor tetrahydrobiopterin. On the contrary, alpha-tocopherol, located mainly in cellular membranes, did not affect tetrahydrobiopterin but increased ionomycin-induced eNOS phosphorylation at serine 1177. The effects of alpha-tocopherol on citrulline and cGMP formation and eNOS phosphorylation were amplified by co-incubation with ascorbate, which is suggested to regenerate oxidized alpha-tocopherol and to act synergistically with alpha-tocopherol. Our data describe a new vasoprotective function of alpha tocopherol that may contribute to the prevention of endothelial dysfunction in vivo. PMID- 15288121 TI - Glutamate-cysteine ligase attenuates TNF-induced mitochondrial injury and apoptosis. AB - Glutathione (GSH) is important in free radical scavenging, maintaining cellular redox status, and regulating cell survival in response to a wide variety of toxicants. The rate-limiting enzyme in GSH synthesis is glutamate-cysteine ligase (GCL), which is composed of catalytic (GCLC) and modifier (GCLM) subunits. To determine whether increased GSH biosynthetic capacity enhances cellular resistance to tumor necrosis factor-alpha- (TNF-alpha-) induced apoptotic cell death, we have established several mouse liver hepatoma (Hepa-1) cell lines overexpressing GCLC and/or GCLM. Cells overexpressing GCLC alone exhibit modest increases in GCL activity, while cells overexpressing both subunits have large increases in GCL activity. Importantly, cells overexpressing both GCL subunits exhibit increased resistance to TNF-induced apoptosis as judged by a loss of redox potential; mitochondrial membrane potential; translocation of cytochrome c to the cytoplasm; and activation of caspase-3, caspase-8, and caspase-9. Analysis of the effects of TNF on these parameters indicates that maintaining mitochondrial integrity mediates this protective effect in GCL-overexpressing cells. PMID- 15288122 TI - Aberrant neuronal and mitochondrial proteins in hippocampus of transgenic mice overexpressing human Cu/Zn superoxide dismutase 1. AB - Mutations of Cu/Zn superoxide dismutase 1 (SOD1), a metalloenzyme catalyzing the conversion of superoxide anion to hydrogen peroxide (H(2)O(2)), are linked to motor neuron degeneration. Transgenic mouse strains overexpressing wild-type human SOD1 (Tg-SOD1) were shown to have mitochondrial swelling, vacuolization, or learning and memory deficits and are widely used for biochemical, genetic, and cognitive studies; this, along with the advent of advanced proteomic methods, made us investigate protein expression in hippocampus. Hippocampal tissues of wild-type, hemizygous, and homozygous Tg-SOD1 mice were isolated and used for two dimensional gel electrophoresis with subsequent matrix-assisted laser desorption/ionization-time of flight identification. We identified several synaptosomal, neuronal, antioxidant, and mitochondrial proteins in hippocampus, and expression levels of syntaxin-binding protein 1, N-ethylmaleimide-sensitive factor, synaptosomal-associated protein 25, dynamin-1, neurofilament triplet L protein, neurofilament triplet M protein, neuronal tropomodulin, and neuronal protein 25 were significantly decreased in Tg-SOD1. None of the antioxidant proteins were altered except mouse SOD1. Mitochondrial ATP synthase alpha/beta chain and elongation factor Tu were aberrant in Tg-SOD1. We conclude that derangement of neuronal and mitochondrial proteins may indicate synaptosomal and neuronal loss in Tg-SOD1 hippocampus, already reported in morphological terms. This observation is of relevance to understanding brain deficits in Down syndrome, as SOD1 is encoded on chromosome 21. PMID- 15288123 TI - Beta-carotene inhibits UVA-induced matrix metalloprotease 1 and 10 expression in keratinocytes by a singlet oxygen-dependent mechanism. AB - UVA exposure causes skin photoaging by singlet oxygen (1)O(2)-mediated induction of, e.g., matrix metalloproteases (MMPs). We assessed whether pretreatment with beta-carotene, a (1)O(2) quencher and retinoic acid (RA) precursor, interferes with UVA-induced gene regulation. HaCaT keratinocytes were precultured with beta carotene at physiological concentrations (0.5, 1.5, and 3.0 microM) prior to exposure to UVA from a Honle solar simulator (270 kJ/m(2)). HaCaT cells accumulated beta-carotene in a time- and dose-dependent manner. UVA irradiation massively reduced the cellular beta-carotene content. Beta-carotene suppressed UVA-induction of MMP-1, MMP-3, and MMP-10, three major matrix metalloproteases involved in photoaging. We show that regulation by not only MMP-1, but also MMP 10, involves (1)O(2)-dependent mechanisms. Beta-carotene dose-dependently quenched (1)O(2)-mediated induction of MMP-1 and MMP-10. Thus, as in chemical solvent systems, beta-carotene quenches (1)O(2) also in living cells. Vitamin E did not cooperate with beta-carotene to further inhibit MMP induction. HaCaT cells produced weak retinoid activity from beta-carotene, as demonstrated by mild upregulation of RAR beta and activation of an RARE-dependent reporter gene. Beta carotene did not regulate the genes encoding other RARs, RXRs, or the two beta carotene cleavage enzymes. These results demonstrate that beta-carotene acts photoprotectively, and that this effect is mediated by (1)O(2) quenching. PMID- 15288124 TI - Nitration and nitrosation of N-acetyl-L-tryptophan and tryptophan residues in proteins by various reactive nitrogen species. AB - Proteins are targets of reactive nitrogen species such as peroxynitrite and nitrogen dioxide. Among the various amino acids in proteins, tryptophan residues are especially susceptible to attack by reactive nitrogen species. We carried out experiments on the reactions of peroxynitrite and other reactive nitrogen species with N-acetyl-L-tryptophan under various conditions. Four major products were identified as 1-nitroso-N-acetyl-L-tryptophan, 1-nitro-N-acetyl-L-tryptophan, 6 nitro-N-acetyl-L-tryptophan, and N-acetyl-N'-formyl-L-kynurenine on the basis of their mass and UV spectra. The reactions with SIN-1 (a peroxynitrite generator), Angeli's salt (a nitroxyl donor), and spermine NONOate (a nitric oxide donor) generated the nitroso derivative but not the nitro derivatives. A myeloperoxidase H(2)O(2)-NO(2)(-) system generated the nitro derivatives but not the nitroso derivative. Under physiological conditions 6-nitro-N-acetyl-L-tryptophan was stable, whereas the 1-nitroso and 1-nitro derivatives decomposed with half-lives of 1.5 and 18 h, respectively. After treatment with various reactive nitrogen species, bovine serum albumin was enzymatically hydrolyzed and analyzed for 6 nitro-L-tryptophan and 3-nitro-L-tyrosine by HPLC with electrochemical detection. Levels of 6-nitro-L-tryptophan and 3-nitro-L-tyrosine were similar in the nitrated protein. 6-Nitro-L-tryptophan in proteins can be measured as an additional biomarker of protein nitration. PMID- 15288125 TI - Effect on endothelial cell gene expression of shear stress, oxygen concentration, and low-density lipoprotein as studied by a novel flow cell culture system. AB - A new cell culture system has been developed that reflects the vascular microenvironment. By means of this system the cultured cells are exposed not only to shear stress by the circulating culture medium, but also to an oxygen concentration gradient and certain critical blood components such as low-density lipoprotein (LDL) and monocytes. DNA microarray analysis was performed for human umbilical vein endothelial cells cultured in this system in the absence and presence of laminar flow at a low shear stress, 0.2 dyn/cm(2). In addition to shear stress, either an oxygen concentration gradient, or LDL (1 mg/ml), or both were applied. Many Nrf-2-regulating genes, such as heme oxygenase 1, NAD(P)H quinone oxidoreductase 1, solute carrier family 7 No. 11, and glutamate-cysteine ligase modifier subunit, were induced by laminar flow at very low shear stress regardless of the additional conditions. Certain genes were specifically affected by exposure to the oxygen gradient and/or LDL under shear stress, but the degree was very low. These results suggest that shear stress is the most critical factor affecting gene expression in endothelial cells and that Nrf-2-regulating proteins may contribute to protecting endothelial cells against other vascular stress. This system should provide highly relevant and useful information about both vascular physiology and pathology, in the latter on such urgent matters as the specific steps involved in atherogenesis. PMID- 15288126 TI - Weight loss and race modulate nitric oxide metabolism in overweight women. AB - Weight reduction is associated with a decrease in the risk of developing cardiovascular disease. We hypothesized that, given the central role of reactive oxygen and nitrogen species in vascular biology, changes in nitric oxide (NO) metabolism contribute to benefits of weight loss. In a controlled weight loss trial involving overweight (body mass index (BMI) = 27-30 kg/m(2)), otherwise healthy premenopausal Caucasian and African-American women, serum levels of nitrite and nitrate, as an index of NO production, and protein 3-nitrotyrosine and myeloperoxidase (MPO), as markers of inflammation, were determined. Testing was performed before and after reduction to normal body weight (BMI < 25) under standardized conditions, with controlled diet, and following 1 month of weight maintenance. After weight loss there was an increase in nitrite and nitrate, and levels were higher among African-American women relative to Caucasian counterparts. Whereas weight loss was associated with a decrease in 3 nitrotyrosine in Caucasian women, no change was observed among African-Americans. Furthermore, MPO levels increased in response to weight loss for African Americans, but did not change in Caucasian women. These data indicate that vascular production of reactive nitrogen species can be modulated by race and weight loss and highlight important racial differences in these responses and are discussed in the context of risk for developing vascular disease. PMID- 15288127 TI - CYP3A induction aggravates endotoxemic liver injury via reactive oxygen species in male rats. AB - We carried out this experiment to evaluate the relationship between isoforms of cytochrome P450 (P450) and liver injury in lipopolysaccharide (LPS)-induced endotoxemic rats. Male rats were intraperitoneally administered phenobarbital (PB), a P450 inducer, for 3 days, and 1 day later, they were intravenously given LPS. PB significantly increased P450 levels (200% of control levels) and the activities (300-400% of control) of the specific isoforms (CYP), CYP3A2 and CYP2B1, in male rats. Plasma AST and ALT increased slightly more in PB-treated rats than in PB-nontreated (control) rats with LPS treatment. Furthermore, either troleandomycin or ketoconazole, specific CYP3A inhibitors, significantly inhibited LPS-induced liver injury in control and PB-treated male rats. To evaluate the oxidative stress in LPS-treated rats, in situ superoxide radical detection using dihydroethidium (DHE), hydroxy-2-nonenal (HNE)-modified proteins in liver microsomes and 8-hydroxydeoxyguanosine (8-OHdG) in liver nuclei were measured in control and PB-treated rats. DHE signal intensity, levels of HNE modified proteins, and 8-OHdG increased significantly in PB-treated rats. LPS further increased DHE intensity, HNE-modified proteins, and 8-OHdG levels in normal and PB-treated groups. CYP3A inhibitors also inhibited the increases in these items. Our results indicate that the induction or preservation of CYP isoforms further promotes LPS-induced liver injury through mechanisms related to oxidative stress. In particular, CYP3A2 of P450 isoforms made an important contribution to this LPS-induced liver injury. PMID- 15288128 TI - Inhibition of the yeast metal reductase heme protein fre1 by nitric oxide (NO): a model for inhibition of NADPH oxidase by NO. AB - Nitric oxide (NO) has been found to inhibit the actions of the transmembrane metal reductase Fre1 in the yeast Saccharomyces cerevisiae. This membrane spanning heme protein is homologous to the gp91(PHOX) protein of the NADPH oxidase enzyme complex and is responsible for reducing extracellular oxidized metals (i.e., ferric and cupric ions) before high-affinity uptake. Consistent with its role in metal metabolism, inhibition of Fre1 by NO also inhibited yeast growth in low-iron medium. Inhibition by NO was found to be O(2)-dependent and irreversible. Further examination of the chemistry responsible for activity loss shows that the generation of N(2)O(3) via NO-O(2) chemistry was responsible for the activity loss, possibly via nitrosation of the protein followed by loss of the heme prosthetic group. PMID- 15288129 TI - Plasma F2-isoprostanes are elevated in newborns and inversely correlated to gestational age. AB - F(2)-isoprostanes, prostaglandin F(2)-like compounds formed by free radical catalyzed lipid peroxidation, are considered the most reliable markers of oxidative stress. It has been repeatedly suggested that newborns are exposed to conditions of oxidative stress resulting from the change from a low oxygen pressure in utero to a high oxygen pressure at birth. We measured the levels of F(2)-isoprostanes in plasma of newborns by gas chromatography/mass spectrometry and we found that F(2)-isoprostanes are significantly higher in term newborns compared to healthy adults. The greatest values were found in preterm newborns in whom F(2)-isoprostanes are even higher than in term babies. Moreover a significant inverse correlation was found between the plasma levels of isoprostanes and the gestational age. A quite normal level of isoprostanes was found in the mothers both at delivery and during pregnancy. Placental total F(2) isoprostanes (sum of free plus esterified) were significantly higher in preterm compared to term deliveries and such a difference might account for the difference in plasma isoprostanes. Plasma non-protein-bound iron is higher in preterm than in term newborns, even if no correlation was found with plasma F(2) isoprostanes. Erythrocyte desferrioxamine-chelatable iron content (0 time) and release (24 h of aerobic incubation) are higher in newborns than in adults and in preterm than in term newborns, but again no correlation was found with plasma F(2)-isoprostanes. The marked increase in plasma isoprostanes suggests that oxidative stress is a feature of the physiopathological changes seen in the perinatal period. PMID- 15288130 TI - Effect of anesthesia on the signal intensity in tumors using BOLD-MRI: comparison with flow measurements by Laser Doppler flowmetry and oxygen measurements by luminescence-based probes. AB - BOLD-contrast functional magnetic resonance imaging (MRI) was used to assess the evolution of tumor oxygenation and blood flow after administration of four different anesthetics: pentobarbital (60 mg/kg), ketamine/xylazine (80/8 mg/kg), fentanyl/droperidol (0.078/3.9 mg/kg), and isoflurane (1.5%). Gradient echo sequences were carried out at 4.7 Tesla in a TLT tumor model implanted in the muscle of NMRI mice. In parallel experiments, tumor blood flow and tumor pO2 were measured using the OxyLite/OxyFlo probe system. A comparison was made with the changes occurring in the skeletal muscle (host tissue). The signal intensity was dramatically decreased in tumors after administration of anesthetics, except isoflurane. These results correlated well with measurements of oxygenation and blood perfusion. Isoflurane produced constant muscle pO2 and blood perfusion although large transient fluctuations in pO2 and blood flow were reported in some tumors. Our results emphasize the need for careful monitoring of the effects of anesthesia when trying to identify new therapeutic approaches that are aimed at modulating tumor hemodynamics. PMID- 15288131 TI - What the little differences between men and women tells us about the BOLD response. AB - We analyzed the functional MRI signal of 15 men and 15 women. All had been presented with a flashed and a reversing, radial checkerboard stimulus. We investigated both positive and negative blood oxygenation level-dependent (BOLD) responses. The extent of activation and the change in the neuronal activity were examined. The former, by counting the number of activated voxels, the latter by using deltaR2* as an indicator of the change in the local deoxyhemoglobin (HbR) concentration. We examined both the positive and the negative BOLD response. Positive BOLD response: The flashed checkerboard gave rise to a larger number of activated voxels than for the reversing checkerboard. The mean number of activated pixels did not differ between men and women. The peak deltaR2* was significantly larger to the flashed than the reversing checkerboard, but did not reveal a gender-related difference. We noted an attenuation of the BOLD signal amplitude with time. This attenuation was larger in women than in men. Negative BOLD response: The attenuation was also larger for the flashed than the reversing stimulus and more pronounced in the chromatic contrast compared to the luminance contrast stimulus. The extent of activation was larger for the flashed than the reversing checkerboard, but did not differ between the sexes. The deltaR2* for the chromatic contrast checkerboard was larger in men than in women. No other significant differences were found. We conclude that the difference in the extent of activation between men and women is the result of our ability to detect activated pixels using statistical methods and not the result of a difference in the processing between the sexes. PMID- 15288132 TI - How much luxury is there in 'luxury perfusion'? An analysis of the BOLD response in the visual areas V1 and V2. AB - We re-analyzed the functional magnetic resonance imaging data from a study involving awake, adult, human volunteers in order to examine the influence of vascular density on the blood oxygenation level-dependent (BOLD) response. We employed a flashed and reversing stimulus paradigm where the latter stimulated twice the number of receptive fields and with it doubled the neuronal metabolic load (CMRO2) compared to the former stimulus. The blood flow increase to these stimuli was identical, so that differences in the BOLD response are due to differences in the oxygen extraction fraction. By comparing the BOLD response in human striate cortex (V1) and its neighbor, extra-striate area V2 to the two stimuli, we were able to determine the influence of the higher vascular density of striate cortex on the BOLD response. In striate cortex, the extent of activation, as measured by the number of activated voxels, was larger for the flashed than for the reversing stimulus. In extra-striate area V2, no such difference in the extent of activation was noted. Gauging the local concentration of HbR using deltaR2*, we found it to be significantly lower for the flashed than for the reversing checkerboard. We estimated the HbR concentration in extra striate area V2 to be double that of striate cortex independent of the stimulus presented. A frequency distribution of the deltaR2* values for the flashed and reversing checkerboard revealed a shift consistent with an increase in the HbR concentration between areas V1 and V2. The metabolically most demanding stimulus, the reversing checkerboard was associated with the highest HbR concentration and with the largest number of voxels with a negative BOLD response. PMID- 15288133 TI - Effects of echo time variation on perfusion assessment using dynamic susceptibility contrast MR imaging at 3 tesla. AB - The implications of changing the echo time of a gradient-echo echo planar imaging sequence applied to dynamic susceptibility contrast magnetic resonance imaging (DSC-MRI) for perfusion imaging at 3T were investigated. Four echo times in the range of 21 to 45 ms were examined in a total of 17 patients who received a dose of 0.1 mmol/kg bodyweight Gadobutrol (Gadovist, 1.0 mmol/ml). As the primary optimization parameter, the concentration-to-noise ratio (SNRc) was selected as it takes effects of variations in baseline as well as in signal drop into account. In an analysis of gray matter, white matter and arterial regions of interest, SNRc showed the highest values for the shortest applied echo time in all cases. Maps of regional cerebral blood volume (rCBV) and blood flow (rCBF) were calculated using deconvolution based on singular value decomposition. The quality of rCBF and rCBV images was judged to be good or excellent in all cases, independent of the echo time. Calculated gray matter/white matter ratios of rCBF and rCBV displayed no significant dependence on the applied echo time. Considering the better SNRc and arterial signal saturation aspects, we found that the shortest investigated echo time was the superior one. We thus suggest that short echo times should be applied, taking technical limitations and clinical demands into consideration. PMID- 15288135 TI - High resolution MRI relaxation measurements of water in the articular cartilage of the meniscectomized rat knee at 4.7 T. AB - Measurements by magnetic resonance imaging (MRI) of the spin-spin (T2), spin lattice (T1) and spin-density (M0) parameters of water protons, optimized by using the Cramer-Rao Lower Bound (CRLB) theory, were made to quantify the effect of surgically induced osteoarthritis on rat knee cartilage at 4.7 T. Partial meniscectomy was performed on the right medial condyle of four Sprague Dawley rats, leaving the left medial condyle as a control. The animals were euthanized 3 weeks after the operation; the entire limbs were removed and T2 and T1 relaxation measurements and M0 measurements of the protons of water were obtained using conventional Carr-Purcell-Meiboom-Gill (CPMG) and saturation recovery methods. M0 was normalized with respect to a water phantom, to obtain the relative spin density M0%. Weight-bearing cartilage areas on the meniscectomized medial condyles exhibited a significant increase of T2 relaxation time (p < 0.001) and of M0% (p < 0.01) with respect to the control; T1 relaxation times did not show any statistically significant changes. CRLB-based sampling optimization offered an insight to improved measurement precision and a reduction of scanning time against conventional sampling methods methods. Quantitative MRI assessment of the meniscectomized rat knee shows that cartilage exhibits changes in T2 and M0 values 3 weeks after operation. PMID- 15288134 TI - Contrast-enhanced MR angiography in rats with hepatobiliary contrast agents. AB - The aim of this animal study was to evaluate the feasibility of contrast-enhanced magnetic resonance (MR) angiography with two hepatobiliary contrast agents, Gd DTPA-DeA and Gd-EOB-DTPA. Coronal images of the rat abdomen were acquired using a three-dimensional spoiled gradient recalled sequence before and after the administration of Gd-DTPA-DeA, Gd-EOB-DTPA, or Gd-DTPA. Four sets of postcontrast images were collected every 90 s. Contrast ratios were calculated for the abdominal aorta on the source images, and the retention index was defined as the ratio of the contrast ratio on the last imaging to that on the first postcontrast imaging. Partial minimum intensity projection (MIP) images covering the abdominal aorta were generated from the first and second postcontrast imagings, and the image quality was visually evaluated. The contrast ratio on the first postcontrast imaging was the highest for Gd-DTPA-DeA, followed by Gd-EOB-DTPA and Gd-DTPA. Retention indices were higher with Gd-DTPA-DeA than with Gd-EOB-DTPA and Gd-DTPA, implying a prolonged contrast effect with Gd-DTPA-DeA. On the MIP image from the first postcontrast imaging, delineation of the abdominal aorta tended to be better with Gd-DTPA-DeA and Gd-EOB-DTPA than with Gd-DTPA, and the difference was evident at low injection doses. Image quality for the second postcontrast imaging was higher with Gd-DTPA-DeA than with the other two agents, suggesting a longer imaging window for Gd-DTPA-DeA. In conclusion, Gd-DTPA-DeA and Gd-EOB-DTPA showed stronger contrast enhancement for the rat abdominal aorta and provided MR angiograms of higher image quality when compared with Gd-DTPA at the same injection dose. These hepatobiliary agents may make it possible to perform contrast-enhanced MR angiography even at a low injection dose. PMID- 15288136 TI - Comparison of high-resolution MRI, optical microscopy and SEM for quantitation of trabecular architecture in the rat femur. AB - Magnetic resonance imaging (MRI) has been used to analyze trabecular bone architecture in femur heads taken from adult Wistar rats. The aim of this study was to validate the use of MRI in assessing trabecular structure and morphology by comparing standard measures of bone morphology in the rat femur obtained from high resolution MRI with those obtained by conventional optical microscopy and by scanning electron microscopy (SEM). MR images were obtained on a Bruker 4.7 T micro-imaging system using a three-dimensional spin echo sequence with spatial resolution of 23 microm in-plane and a slice thickness of 39 microm. Optical images were obtained by de-calcifying the bone in EDTA and then sectioning 5 microm-thick slices. SEM images were obtained from bone embedded in epoxy resin with surface preparation by diamond polishing. Values of standard bone morphological parameters were compared and correlation coefficients between the MRI and the optical- and SEM-derived measures of morphology were calculated. Partial volume effects in MRI were minimized in this study by the use of very thin slices, yielding better agreement with optical- and SEM-derived measures of trabecular bone morphology than have been obtained in previous studies. Correlations between the MRI and optical data were significantly lower than those between the MRI and SEM data. Effects of de-calcification were also investigated. The results indicate that comparison of MRI with thin (de-calcified) optical images may be inherently flawed due to the destructive de-calcification and sectioning process used to prepare samples for the optical imaging. PMID- 15288137 TI - Apparent diffusion coefficient of intervertebral discs related to matrix composition and integrity. AB - While tremendous work has been performed to characterize degenerative disc disease through gross morphologic, biochemical, and histologic grading schemes, the development of an accurate and noninvasive diagnostic tool is required to objectively detect changes in the matrix with aging and disc degeneration. In the present study, quantitative magnetic resonance was used to determine if the quality of the nutritional supply to the intervertebral disc at various ages and levels of degeneration could be assessed through measurement of the apparent diffusion coefficients (ADCs). Modifications of the nucleus pulposus matrix content, specifically of water and glycosaminoglycan contents, with age and disc degeneration, were reflected in correlating changes in the ADCs. From unforced stepwise linear regression analyses, relations were established showing that decreases in glycosaminoglycan or water contents in the nucleus pulposus resulted in direct decreases in the ADCs. Relations obtained for the ADCs of the nucleus pulposus were direction dependent, in conformity with the anisotropic diffusion in the intervertebral discs. Changes in matrix integrity, as evidenced by the percentage of denatured collagen, were also detected in the nucleus pulposus with a low positive correlation to the ADC along the height of the disc and an inverse statistically significant regression to the ADC along the anterior to posterior axis of the disc. Correlations between the matrix content and integrity of the annulus fibrosus and its ADCs were not as evident, with only the ADC in the lateral direction of the disc of the anterior annulus fibrosus able to reflect changes in matrix content. The information obtained by the ADCs, particularly of the nucleus pulposus, can potentially be used in combination with quantitative T1, T2, and MT parameters to noninvasively obtain a quantitative assessment of the disc matrix composition and structural integrity. PMID- 15288138 TI - Investigation of microenvironmental factors influencing the longitudinal relaxation times of drugs and other compounds. AB - The aim of this study was to investigate the microenvironmental factors likely to influence the longitudinal relaxation time of MR visible drugs or compounds in vivo at 1.5 T. The relative influence that viscosity, albumin and paramagnetic contrast agent concentrations have on the observed longitudinal relaxation times of three 19F MR detectable drugs and compounds have been investigated. Our data show that for 5-fluorouracil, flucloxacillin and tetrafluorosuccinic acid containing phantoms, the presence of albumin at normal physiological concentrations will have relaxation effects of the same order of magnitude as that of a commonly clinically administered contrast agent, gadolinium diethylenetriamine pentaacetic acid. The contribution of viscosity is shown, in the examples studied here, to be of minor importance, contributing less than 6.5% to the observed relaxation effects. It is also demonstrated that in the presence of competitive binding of other ligands for common binding sites on albumin, the 19F longitudinal relaxation time of 5-fluorouracil can increase by up to 340% from its value in the absence of the competing ligand. The relevance of the findings to in vivo studies is discussed. PMID- 15288139 TI - Sodium T2*-weighted MR imaging of acute focal cerebral ischemia in rabbits. AB - Changes in T2*-weighted tissue sodium (23Na) signal following acute ischemia may help to identify necrotic tissue and estimate the duration of ischemia. Sodium signal was monitored in a rabbit model of acute (0-4 h) focal cerebral ischemia, using gradient echo 23Na MR images (echo time = 3.2 ms) acquired continuously in 20-min intervals on a 4-Tesla MRI. 2,3,5-Triphenyl-tetrazolium chloride staining was used to identify regions of necrosis. In necrotic tissue, average 23Na image signal intensity decreased by 11% +/- 8% during the first 40 min of ischemia followed by a linear increase (0.19%/min) to 25% +/- 14% greater than baseline after 4 h of ischemia. The time course of 23Na signal change observed in necrotic tissue following focal ischemia in this rabbit model is consistent with an initial decrease in 23Na T2* relaxation time followed by an increase in tissue sodium concentration and provides further evidence that tissue 23Na signal may offer unique information regarding tissue viability that is complementary to other MR imaging techniques. PMID- 15288140 TI - Using nine degrees-of-freedom registration to correct for changes in voxel size in serial MRI studies. AB - Quantitative longitudinal brain magnetic resonance (MR) studies may be confounded by scanner-related drifts in voxel sizes. Total intracranial volume (TIV) normalisation is commonly used to correct serial cerebral volumetric measurements for these drifts. We hypothesised that automated rigid-body registration of whole brain incorporating automatic scaling correction might also correct for such fluctuations, and might be a more practical alternative. Twenty-three subjects (12 patients with Alzheimer's disease [AD] and 11 controls) had at least two serial T1-weighted volumetric brain MR scans. Ten scans from the control subjects were artificially scaled (stretched) by 1.5, 3.0, 4.6 and 6.1%. A 9-degrees-of freedom (9dof) registration was used to register the scaled scans back onto the original scans and corresponding scaling factors compared to TIV measurements. A further nine 1-year repeat scans from the AD subjects were artificially scaled and registered (9dof) to baseline. The two correction methods were further assessed using multiple serial scans for each of the 23 subjects (resulting in 49 scan pairs). All serial scans were registered (9dof) to baseline. TIV was measured on all scans. It was found that the 9dof registration successfully recovered the artificially generated scaling changes. Scaling correction using 9dof registration did not alter the amount of brain atrophy measured over the 1 year period in the AD subjects. The 9dof volume scaling factors were very similar to the TIV ratios (repeat TIV over baseline TIV), but less variable (p < 0.001), in both artificial and 'real' scenarios. In the latter, the volume scaling factors allowed identification of two time-points in which a 3% change in voxel size had occurred. Both the 9dof brain registration and TIV correction were successfully able to correct for these fluctuations. Significant shifts in voxel size are a problem in longitudinal brain imaging studies. It is important that such changes are adjusted for: 9dof registration, which is automated and computationally inexpensive, may be superior to the more labour-intensive TIV correction for this purpose. PMID- 15288141 TI - Realignment of myocardial first-pass MR perfusion images using an automatic detection of the heart-lung interface. AB - Magnetic resonance first-pass imaging of a bolus of contrast agent is well adapted to distinguish normal and hypoperfused areas of the myocardium. In most cases, the signal intensity-time curves in user defined regions of interest (ROI) are studied. As image acquisition is ECG-gated, the images are acquired at the same moment in the cardiac cycle, and the basic shape of the heart is similar from one view to the next. However, superficial respiratory motion can displace the heart in the short-axis plane. The aim of this study is to correct for the respiratory motion of the heart by performing an automatic realignment of the myocardial ROI based on a method tracking the movement of the lung-myocardium interface. Visual and quantitative analyses performed on 120 curves show a very good concordance between two automatic methods and the manual one. PMID- 15288142 TI - Stereology versus planimetry to estimate the volume of malignant liver lesions on MR imaging. AB - Liver tumor volume measurements are clinically useful in patients undergoing cancer treatment. The techniques of planimetry and stereology were applied for this purpose on magnetic resonance (MR) imaging. Fifty-eight malignant liver lesions were depicted on MR images in 20 consecutive patients. The volume of all lesions was estimated using stereology technique, based on point counting. Stereological tumor volume estimations were compared with those determined by manual planimetry. The repeatability of both techniques was assessed. Tumor volumes estimated by the two techniques were highly correlated (r = 0.98, p < 0.0001). The 95% limits of agreement showed that the stereological volume estimations may differ from the planimetric assessments by less than 23%. Both techniques presented comparable intra- and interobserver variability. The planimetry was 1.5 times faster than the stereology. Both volumetric techniques may provide reliable and reproducible liver tumor volume estimations. The planimetry may be the method of choice because of its superior speed. PMID- 15288143 TI - In vivo quantification of the metabolites in normal brain and brain tumors by proton MR spectroscopy using water as an internal standard. AB - Metabolite concentrations in normal adult brains and in gliomas were quantitatively analyzed by in vivo proton magnetic resonance spectroscopy (MRS) using the fully relaxed water signal as an internal standard. Between January 1998 and October 2001, 28 healthy volunteers and 18 patients with gliomas were examined by in vivo proton MRS. Single voxel spectra were acquired using the point-resolved spectroscopic pulse sequence with a 1.5-T scanner (TR/TE/Ave = 3000 ms/30 ms/64). The calculated concentrations of N-acetyl-aspartate (NAA), creatine (Cre), choline (Cho), and water (H2O) in the normal hemispheric white matter were 23.59 +/- 2.62 mM (mean +/- SD), 13.06 +/- 1.8 mM, 4.28 +/- 0.8 mM, and 47280.96 +/- 5414.85 mM, respectively. The metabolite concentrations were not necessarily uniform in different parts of the brain. The concentrations of NAA and Cre decreased in all gliomas (p < 0.001). The NAA/Cho and NAA/H2O ratios can distinguish the normal brain from gliomas, and low-grade astrocytoma from high grade group (p < 0.001). The concentration of taurine (Tau) in medulloblastomas was 29.64 +/- 5.76 mM. This is the first quantitative analysis of Tau in medulloblastoma in vivo and confirms earlier in vitro findings. PMID- 15288144 TI - Parallel magnetic resonance imaging using coils with localized sensitivities. AB - The purpose of this study was to present clinical examples and illustrate the inefficiencies of a conventional reconstruction using a commercially available phased array coil with localized sensitivities. Five patients were imaged at 1.5 T using a cardiac-synchronized gadolinium-enhanced acquisition and a commercially available four-element phased array coil. Four unique sets of images were reconstructed from the acquired k-space data: (a) sum-of-squares image using four elements of the coil; localized sum-of-squares images from the (b) anterior coils and (c) posterior coils and a (c) local reconstruction. Images were analyzed for artifacts and usable field-of-view. Conventional image reconstruction produced images with fold-over artifacts in all cases spanning a portion of the image (mean 90 mm; range 36-126 mm). The local reconstruction removed fold-over artifacts and resulted in an effective increase in the field-of-view (mean 50%; range 20-70%). Commercially available phased array coils do not always have overlapping sensitivities. Fold-over artifacts can be removed using an alternate reconstruction method. When assessing the advantages of parallel imaging techniques, gains achieved using techniques such as SENSE and SMASH should be gauged against the acquisition time of the localized method rather than the conventional sum-of-squares method. PMID- 15288145 TI - Rapid and accurate measurement of transverse relaxation times using a single shot multi-echo echo-planar imaging sequence. AB - Methods for making rapid and accurate measurements and maps of the transverse relaxation time from a single free induction decay (FID) are proposed. The methods use a multi-echo sequence in combination with B1 insensitive (hyperbolic secant or BIREF2b) refocusing pulses and rapid echo-planar imaging techniques. The results were calibrated against a single spin echo echo-planar imaging sequence using a phantom containing a range of CuSO4 concentrations. The mean percentage absolute difference between the multi-echo and single-echo results was 3% for the multi-echo sequence using the hyperbolic secant refocusing pulse, and 7% for the multi-echo sequence using the BIREF2b refocusing pulse, compared to 13% for a multi-echo sequence using a nonselective sinc refocusing pulse. The use of the sequences in vivo has been demonstrated in studies of gastric function, i.e., the measurement of gastric dilution and monitoring of formation of a raft of alginate polysaccharide within the stomach. PMID- 15288146 TI - Proton magnetic resonance imaging of diffusion of high- and low-molecular-weight contrast agents in opaque porous media saturated with water. AB - Besides their use in contrast-enhanced proton magnetic resonance imaging (MRI), contrast agents were found to be useful as tracer molecules. Since paramagnetic ions in water have the ability to reduce the T1 of protons around them, MRI can determine the locations of Mn2+ and Gd3+ of ppm concentration in water. In opaque porous media saturated with water, MRI revealed diffusional motions of three contrast agents: MnCl2 (molecular-weight [M.W.], 126), gadolinium-diethylene triaminepenta acetic acid (Gd-DTPA) (M.W., 743) and albumin (Gd-DTPA) (M.W., 94,000) at a diffusional displacement ratio of 9:5:2. With the aid of these contrast agents, the transport of low- to high-molecular-weight molecules in opaque water media such as living bodies can be observed using MRI. PMID- 15288147 TI - Adult mesoblastic nephroma: appearances on magnetic resonance imaging. AB - Mesoblastic nephroma presenting in an adult is extremely unusual. The magnetic resonance imaging (MRI) appearances of this tumor in adulthood have not been widely reported. We present a 55-year-old patient who was diagnosed with this rare neoplasm and describe the MRI findings. PMID- 15288148 TI - Ganglioglioma of the right optic tract: case report and review of the literature. AB - We describe a case of ganglioglioma of the right optic tract in a 52-year-old woman. Review of the general literature discloses only 11 cases of tumors of the visual pathway that meet the histological criteria for such neoplasm. A detailed radiological description of our case and a review of the literature is provided. PMID- 15288149 TI - Immunoblastic large B-cell lymphoma of the peripancreatic head region: MR findings. AB - We report the magnetic resonance (MR) appearance of a large B-cell lymphoma in the peripancreatic head region, in a 38-year-old male who presented with a 1 month history of pruritus and jaundice. Routine laboratory examination at presentation revealed an elevated bilirubin. The tumor was a large, solitary well defined mass with no evidence of necrosis, which showed mild diffuse heterogeneous enhancement. The tumor was closely applied to the lateral margin of the head of the pancreas. The constellation of MR findings was interpreted as consistent with the correct eventual diagnosis of lymphoma. PMID- 15288150 TI - In search of the grail: the never-ending story of biomarkers for coronary risk prediction. PMID- 15288151 TI - Pharmacological cardioversion of recent onset atrial fibrillation. PMID- 15288152 TI - The emerging role of delayed contrast-enhanced magnetic resonance imaging in the peri-operative evaluation of patients undergoing coronary revascularisation. PMID- 15288153 TI - Obstructive sleep apnoea and plasma homocysteine: an overview. PMID- 15288154 TI - BNP in acute coronary syndromes: the heart expresses its suffering. PMID- 15288155 TI - Associations between differential leucocyte count and incident coronary heart disease: 1764 incident cases from seven prospective studies of 30,374 individuals. AB - AIMS: We aimed to assess potential associations between different leucocyte components and coronary heart disease (CHD) in a prospective cohort study, and to put these findings in context of other relevant prospective studies in a meta analysis. METHODS AND RESULTS: We report data on differential leucocyte count and CHD derived from the first National Health and Nutrition Examination Survey (NHANES I) and the NHANES 1 Epidemiologic Follow-up Study (NHEFS) involving 4625 individuals followed, on average, for 18 years. The NHEFS involved 914 incident CHD cases and yielded an adjusted risk ratio of 1.09 (0.93-1.29) comparing individuals with neutrophil counts in the top third versus those in the bottom third of the population. In a meta-analysis involving the NHEFS and four other studies comprising a total of 1764 incident CHD cases, the association of CHD with neutrophil counts was somewhat stronger than those with other specific leucocyte components (combined risk ratio=1.33 [1.17-1.50]) but there was substantial heterogeneity between the separate studies (Chi2(4), p <0.001). CONCLUSIONS: Although the present synthesis provides the most comprehensive assessment so far of specific leucocyte components in CHD, additional prospective data will be needed to resolve whether neutrophil counts are much stronger predictors of CHD risk than other components. PMID- 15288156 TI - Visualisation and quantification of peri-operative myocardial infarction after coronary artery bypass surgery with contrast-enhanced magnetic resonance imaging. AB - AIMS: To evaluate if elevated biochemical marker levels after coronary artery bypass grafting (CABG) correspond to the amount of peri-operatively infarcted myocardium, quantified by magnetic resonance imaging (MRI) post-operatively. METHODS AND RESULTS: A total of 23 patients without evidence of previous myocardial infarction or myocarditis and with normal pre-operative ECG and left ventricular function and who underwent elective, primary CABG, without any other concomitant cardiac surgery, were included. Plasma creatinine kinase MB (CK-MB) and troponin I and T were measured on the first, second and fourth post-operative days. Between the fourth and ninth post-operative days, cardiac MRI was carried out. Infarctions were found in 18 patients. The infarction mass at MRI was numerically largest in patients with transmural infarctions, all of whom had a CK MB more than five times the upper normal limit. All three cardiac markers correlated to the mass of infarction. CONCLUSION: Elevated biochemical markers after CABG correspond to the amount of peri-operatively infarcted myocardium. PMID- 15288158 TI - Heart rate is a predictor of success in the treatment of adults with symptomatic paroxysmal supraventricular tachycardia. AB - AIMS: To analyse whether heart rate may affect the efficacy of adenosine, verapamil and carotid sinus massage in terminating symptomatic episodes of paroxysmal supraventricular tachycardia (PSVT) in adults. METHODS AND RESULTS: The study population was selected among 175 adult patients, affected by symptomatic PSVT. One hundred and six of them were considered eligible for the study. Each subject received one of the following treatments: verapamil, 5 mg intravenously (IV) in 5-10 min, followed by 1-5 microg/kg/min; adenosine, 6 mg IV, followed by 12 mg IV after 2-3 min; carotid sinus massage. Adenosine and verapamil were similarly effective in terminating PSVT (74.4% vs 81.8%, P=0.45). The efficacy of carotid sinus massage was significantly lower in comparison with the other two groups (32.4%, P=0.00032 vs adenosine and P=0.000044 vs verapamil group). At logistic regression, PSVT rate showed a positive association with the percentage of sinus rhythm restoration in the group who received adenosine (P=0.0004). The probability of success in resolving the tachycardia following treatment with adenosine was > 75% for heart rates over 166 beats per minute (bpm), but rapidly decreased at lower frequencies, reducing to 25% at 138 bpm. In the verapamil group, PSVT rate was negatively related to the percentage of sinus rhythm restoration (P=0.018). The probability of success in terminating PSVT following administration of verapamil was > 75% for heart rates lower than 186 bpm, but tended to decrease at faster rates, reducing to 25% at 241 bpm. No significant effects of heart rate were observed in the carotid sinus massage group (P=0.17). The probability curves obtained in the adenosine and verapamil group crossed at a point corresponding to 173 bpm, which may represent a cut-off value to predict which treatment could ensure higher rate of success. CONCLUSIONS: Heart rate predicts restoration of sinus rhythm in adult subjects with symptomatic episodes of PSVT treated with adenosine and verapamil. Adenosine is highly effective in PSVT characterised by fast rates, whereas the efficacy of verapamil is increased in patients with low-frequency PSVT. PMID- 15288157 TI - Efficacy, safety and tolerability of beta-adrenergic blockade with metoprolol CR/XL in elderly patients with heart failure. AB - AIM: To study the efficacy and tolerability of beta-blockade in elderly patients with heart failure in the MERIT-HF study. METHODS AND RESULTS: Cox proportional hazards model was used to calculate hazard ratios (HR) with 95% confidence intervals (CI). Risk reduction was defined as (1-HR). In patients > or = 65 years total mortality was reduced by 37% (95% CI 17% to 52%; p=0.0008), sudden death by 43% (95% CI 17% to 61%; p=0.0032), and death from worsening heart failure by 61% (95% CI 32% to 77%; p=0.0005). Hospitalisations for worsening heart failure was reduced by 36% (p=0.0006). Elderly patients with severe heart failure (NYHA class III/IV with ejection fraction < 0.25; n=425, and patients above 75 years (n=490) showed similar risk reductions. Metoprolol CR/XL was safe and well tolerated both during initiating therapy and during long-term follow-up. CONCLUSIONS: Metoprolol CR/XL was easily instituted, safe and well tolerated in elderly patients with systolic heart failure. The data suggest that these are the patients in whom treatment will have the greatest impact as shown by number of lives saved and number of hospitalisations avoided. The time has come to overcome the barriers that physicians perceive to beta-blocker treatment, and to provide it to the large number of elderly patients with heart failure in need of this therapy. PMID- 15288159 TI - Flecainide versus ibutilide for immediate cardioversion of atrial fibrillation of recent onset. AB - AIMS: This study compared the efficacy and safety of intravenous flecainide and ibutilide for immediate cardioversion of atrial fibrillation (AF). METHODS AND RESULTS: We conducted a prospective, randomised trial, including 207 patients with AF of recent onset (< or = 48 h). Flecainide was given over 20 min at a dose of 2 mg/kg body weight (maximum 200 mg), ibutilide was infused at a dose of 1 mg (or 0.01 mg/kg if less than 60 kg) over 10 min, followed by a 10 min observation period and an identical second dose if AF did not convert to sinus rhythm (SR). Treatment was considered successful if SR occurred within 90 min of starting medication. The conversion rates were 56.4% in patients given flecainide and 50.0% in patients given ibutilide (P=0.34). Multivariate analysis revealed that a lower age for women independently increased the probability of conversion. None of the other variables, including left atrial size, left ventricular systolic function, presence of left ventricular hypertrophy, plasma levels of potassium or magnesium at baseline, or concomitant use of digoxin, beta-blocker, diltiazem or verapamil were predictors of conversion. The frequency of adverse events was comparable in the two treatment groups. CONCLUSIONS: There was no significant difference in the cardioversion efficacy or in the risk of adverse events between flecainide and ibutilide in patients with AF of recent onset. In patients without contraindications to both medications, the physician's choice has to be governed by other factors. PMID- 15288160 TI - Plasma homocysteine in obstructive sleep apnoea. AB - AIMS: Whether increased homocysteine is one mechanism linking obstructive sleep apnoea (OSA) to cardiovascular abnormalities is unclear. We hypothesised that plasma homocysteine would be higher in OSA patients than in control subjects, would increase further during sleep, and decrease after treatment with continuous positive airway pressure (CPAP). METHODS AND RESULTS: For study A, homocysteine was measured in 22 OSA patients and 20 controls first before sleep, then after 5 h of untreated OSA, and then in the morning after CPAP treatment. Homocysteine was similar in the OSA and control subjects at all three time points, and declined overnight in both groups (P=0.0017, P=0.036, respectively). To further assess this diurnal variation, we studied plasma homocysteine under a full-night protocol in 10 OSA patients and 12 controls (study B). Homocysteine was measured before sleep, in the morning after sleep, and at noon. Results in both OSA and control groups showed an overnight decline in homocysteine which was reversed by noon (repeated measures ANOVA: OSA, P=0.04; controls, P=0.02). Study C showed that disturbed sleep did not affect homocysteine levels in normal subjects. CONCLUSION: There is a significant diurnal variation in plasma homocysteine, so that homocysteine is lower in the morning after waking. Neither OSA nor disturbed sleep elicit acute or chronic changes in homocysteine. PMID- 15288161 TI - Long-term assessment of a novel biodegradable paclitaxel-eluting coronary polylactide stent. AB - AIM: The aim of this study was to assess technical feasibility, biocompatibility, and impact on coronary stenosis of a new biodegradable paclitaxel-loaded polylactide stent. Due to high rates of in-stent restenosis and permanent nature of metal stent implants, synthetic polymers have been proposed as surrogate materials for stents and local delivery systems for drugs. Paclitaxel was shown to inhibit vascular smooth muscle cell proliferation and migration. METHODS AND RESULTS: A novel biodegradable double-helical stent was manufactured using controlled expansion of saturated polymers (CESP) for the moulding of a bioresorbable poly(D,L)-lactic acid (PDLLA). A modified balloon catheter for stent deployment was developed according to the mechanical stent properties. Twelve paclitaxel-loaded (170 microg) polylactide stents, 12 unloaded polylactide stents, and 12 316L bare metal stents were deployed in porcine coronary arteries of 36 animals. Six pigs of each group were sacrificed after 3 weeks and 3 months, respectively, for every setting. Drug release kinetics as well as histomorphometrical and histopathological analyses were performed. A slow paclitaxel release kinetic for more than 2 months and therapeutic tissue concentrations were demonstrated. Coronary stenosis after implantation of paclitaxel-loaded stents (30+/-5% or 49+/-4%) was significantly inhibited compared to unloaded PDLLA stents (65+/-10%, P=0.021 or 71+/-4%, P=0.004) and metal stents (53+/-6% or 68+/-8%, P=0.029 and P=0.020) after 3 weeks or 3 months. Early complete endothelialisation was shown. Nevertheless, a local inflammatory response to the polylactide as a result of the polymer resorption process was observed. CONCLUSIONS: This novel polylactide stent showed sufficient mechanic stability, and by incorporation of paclitaxel, a significant potential to reduce restenosis development after vascular intervention was seen. PMID- 15288162 TI - Expert consensus document on beta-adrenergic receptor blockers. PMID- 15288163 TI - Tissue Doppler and cardiac resynchronisation therapy: a new challenge for the optimal choice of candidates. PMID- 15288165 TI - Long-term clopidogrel therapy in the drug-eluting stent era: beyond CREDO and PCI CURE. PMID- 15288167 TI - Cardiovascular disease, periodontitis, and the monocyte relationship. PMID- 15288169 TI - Direct epicardial mapping predicts the recovery of left ventricular dysfunction in chronic ischaemic myocardium. PMID- 15288171 TI - Heart failure clinics have decreased mortality and hospitalisation rates in Sweden. PMID- 15288173 TI - Contrast agent nephropathy. PMID- 15288175 TI - Proceedings of the 'HIPPOKRATION' Congress on Reproductive Immunology (Fourth Congress of the European Society for Reproductive and DEvelopmental Immunology). June 4-6, 2003. Rhodes Island, Greece. PMID- 15288177 TI - Human decidual lymphocytes and the immunobiology of pregnancy. PMID- 15288176 TI - Differential down-modulation of HLA-G and HLA-A2 or -A3 cell surface expression following human cytomegalovirus infection. AB - During pregnancy, the non-classical major histocompatibility complex (MHC) class I HLA-G molecule is specifically expressed in trophoblast cells at the materno fetal interface and may exert a local control of the immune response against viral infections. Human cytomegalovirus (HCMV) infection, which is the major cause of congenital defects, encodes multiple glycoproteins (US2, US3, US6, US10 and US11) that interrupt the MHC class I pathway of antigen presentation. The effect of some of these unique short (US) proteins on HLA-G expression has been previously studied, but little is known about the modulation of HLA-G cell surface expression during the course of HCMV infection which ensures expression of all of these US proteins. Using flow cytometry analysis, HLA-G cell surface expression was evaluated in HCMV-infected U373-HLA-G transfectant cells and compared with the modulation of the endogenous classical HLA-A2 molecules. The results indicated that HCMV infection down-modulated HLA-G cell surface expression, but later after infection and to a lesser extent than HLA-A2. Using various HLA-G/HLA-A2 chimeras, we showed that the unique structure of HLA-G cytoplasmic tail was partly involved in the resistance of HLA-G to viral down modulation. Such limited down-modulation of HLA-G may have functional consequences in term of innate immunity against congenital HCMV infection. PMID- 15288178 TI - Role of a KIR/HLA-C allorecognition system in pregnancy. AB - Decidual natural killer (NK) cells are thought to play a significant role in the allorecognition mechanisms during pregnancy. Through their activating and inhibitory receptors they may recognize selectively class I HLA alleles expressed on invading trophoblast and provide self-signals to control NK responses, thus regulating the maternal immune response at the fetomatenal interface. Killer immunoglobulin-like receptors (KIR) constitute one of the families of class I MHC receptors which are expressed on NK cells. Their repertoire includes both activating and inhibitory receptors, most of which recognize specific epitopes on HLA-C molecules and can either activate NK cell responses or abort activating signals and inhibit NK cell functions. Since KIRs are expressed on decidual NK cells and the HLA-C molecules that they recognize are also expressed on invading trophoblast, KIR receptors may play a regulatory role in pregnancy by interacting with their trophoblastic HLA-C counterparts and providing trophoblast damage evading signals (KIR/HLA-C allorecognition system). Our hypothesis that the KIR/HLA-C system might be ineffective in some unsuccessful pregnancies, has been investigated in women with unexplained spontaneous abortions. Our results suggest that a limited maternal repertoire of inhibiting KIRs (inhKIRs) and/or lack of maternal inhKIR-fetal HLA-C epitope matching may predispose to miscarriage. PMID- 15288179 TI - Is there a place for immunomodulation in assisted reproduction techniques? AB - We briefly review the history of the concepts of the materno foetal relationship, and the (various) rationales which have been used to justify lymphocyte alloimmunisation (LA) as a treatment for recurrent spontaneous abortion of putative immune origins. The effectiveness of the treatment is at best dubious and limited to a small number of women for which there is no real positive selection rationale, at worst it is not efficient. The rationales themselves are rather "evolutive". The present one is to use the Th1:Th2 paradigm and, thus, to propose to "dampen NK activity" in abortion prone women and this concept has been extended by some to implantation failure. We briefly review why the Th1:Th2 paradigms is no longer fully valid, describe briefly why it is inappropriate for implantation, and conclude that alloimmunisation should no longer be proposed for RSA, hence, more for implantation failure. We, however, do not reject immunotherapy, but we believe that molecular and cellular defects specific approaches should be used, tailored for the specific pathway whose disruption cause the clinical symptom. PMID- 15288180 TI - Active or passive immunization in unexplained recurrent miscarriage. AB - Controversy exists as to whether active immunotherapy with allogeneic lymphocyte transfusions (ALT) or passive immunotherapy with intravenous immunoglobulin (IvIg) improve the chance of live birth in women with unexplained recurrent miscarriages (RM). Meta-analyses of the placebo-controlled trials carried out as Cochrane reviews have concluded than none of the different forms of immunotherapy has proved effective in the total RM population. However, the included trials have generally been small and very heterogenous with respect to the clinical histories of patients and the immunization protocols. Thus, other meta-analyses which have looked at the efficacy in subgroups of RM patients have reported that ALT and IvIg may be effective in women with primary RM and secondary RM, respectively. In RM clinics in Denmark, ALT with donor lymphocytes or IvIg immunotherapy have been tested in several placebo-controlled trials since 1986 in which greater doses than used in other trials have been administered, and both treatments are now used for routine therapy. Our results have convinced us that using the correct immunization protocols on the right subsets of RM patients is effective, but we admit that new placebo-controlled trials focusing on subsets of RM patients are now urgently needed. Furthermore, treated patients should be extensively monitored for changes in a series of immune parameters that may predict pregnancy success and be of importance for our understanding of the mechanisms of action of immunotherapy in RM. PMID- 15288181 TI - Endometrial and placental CRH as regulators of human embryo implantation. AB - Epithelial cells of the human endometrium and differentiated endometrial stromal cells express the corticotropin-releasing hormone (CRH) gene. CRH is also produced by human placental cytotrophoblast. Endometrial and placental CRH are under the endocrine control of gonadal steroids as well as under autocrine/paracrine regulation by prostanoids and interleukins. Human endometrium, myometrium and placenta express the relevant receptors. Human trophoblast and decidualized endometrial cells also express Fas ligand (FasL), a pro-apoptotic molecule. These findings suggest that intra-uterine CRH may participate in local inflammatory phenomena associated with blastocyst implantation, while FasL may assist with maternal immune tolerance to the semi allograft embryo. A nonpeptidic CRH receptor type 1 (CRH-R1)-specific antagonist decreased the expression of FasL by human trophoblasts, suggesting that CRH regulates the pro-apoptotic potential of these cells in an auto-paracrine fashion. Invasive trophoblasts promoted apoptosis of activated Fas-expressing human T lymphocytes, an effect potentiated by CRH and inhibited by the CRH antagonist. Female rats treated with the CRH antagonist in the first 6 days of gestation had a dose-dependent decrease of endometrial implantation sites and live embryos as well as markedly diminished endometrial FasL expression. However, embryos of mothers lacking T cells (nude rats) and embryos of syngeneic matings were not rejected when mothers were treated with antalarmin, suggesting that the effect of antalarmin on embryonic implantation is not due to a nonspecific toxicity of this compound but a specific effect on T cells. Our data suggest important physiological roles of endometrial and placental CRH in the regulation of blastocyst implantation and early maternal tolerance. PMID- 15288182 TI - Stress and the female reproductive system. AB - The hypothalamic-pituitary-adrenal (HPA) axis, when activated by stress, exerts an inhibitory effect on the female reproductive system. Corticotropin-releasing hormone (CRH) inhibits hypothalamic gonadotropin-releasing hormone (GnRH) secretion, and glucocorticoids inhibit pituitary luteinizing hormone and ovarian estrogen and progesterone secretion. These effects are responsible for the "hypothalamic" amenorrhea of stress, which is observed in anxiety and depression, malnutrition, eating disorders and chronic excessive exercise, and the hypogonadism of the Cushing syndrome. In addition, corticotropin-releasing hormone and its receptors have been identified in most female reproductive tissues, including the ovary, uterus, and placenta. Furthermore, corticotropin releasing hormone is secreted in peripheral inflammatory sites where it exerts inflammatory actions. Reproductive corticotropin-releasing hormone is regulating reproductive functions with an inflammatory component, such as ovulation, luteolysis, decidualization, implantation, and early maternal tolerance. Placental CRH participates in the physiology of pregnancy and the onset of labor. Circulating placental CRH is responsible for the physiologic hypercortisolism of the latter half of pregnancy. Postpartum, this hypercortisolism is followed by a transient adrenal suppression, which may explain the blues/depression and increased autoimmune phenomena observed during this period. PMID- 15288183 TI - How far from a hormone-based contraceptive vaccine? AB - Antibodies of appropriate specificity are able to block the action of hormones which are obligatory for successful reproduction. Thus, if immunisation using such hormones can provoke adequate titres of bioneutralizing antibodies in sexually mature individuals, the vaccinee becomes infertile ('immunocontraception') for as long as sufficient titres of the antibodies are maintained. In the case of hormones that are required for the development of sexual maturity in the male, immunisation of young animals can prevent sexual maturation ('immunocastration'). The hormones which have been targeted are gonadotropin-releasing hormone (GnRH) for both immunocastration and immunocontraception, and follicle-stimulating hormone (FSH) and human chorionic gonadotropin (hCG) for immunocontraception. PMID- 15288184 TI - Update on zona pellucida glycoproteins based contraceptive vaccine. AB - Zona pellucida (ZP) glycoproteins, due to their critical role in mammalian fertilization, have been proposed as candidate immunogens for development of a contraceptive vaccine. Active immunization studies in a variety of animal species, employing either native or recombinant zona proteins, has established their contraceptive potential. Hence, ZP glycoprotein-based contraceptive vaccines have a very good potential for controlling wild life population. To make it a realistic proposition, additional research inputs are required to develop new potent adjuvants and novel practical strategies for vaccine delivery. The observed ovarian dysfunction, often associated with immunization by ZP glycoproteins, is one of the major obstacles for their application in the control of human population. Ongoing studies to delineate epitopes of ZP glycoproteins that will generate an immune response capable of inhibiting fertility without any untoward effects on ovarian functions will help in determining their feasibility for human use. PMID- 15288185 TI - Possible presence of O-linked carbohydrate in the human male reproductive tract CD52. AB - Male reproductive tract CD52 (mrtCD52) is an antigen recognized by a complement dependent sperm-immobilizing monoclonal antibody (SI-Abs) derived in an infertile patient. The molecule has been shown to contain a unique N-linked carbohydrate that does not cross-react with other tissues. In this study, we have investigated whether O-linked carbohydrate as well as N-linked carbohydrate is present in mrtCD52 using specific lectins and anti-CD52 core peptide antiserum. The lectin PNA, which recognizes O-linked carbohydrate [Galbeta1-3GalNAc], reacted with mrtCD52 and showed a similar polymorphic reaction pattern to that of the anti peptide antiserum in western blotting analysis on two-dimensional SDS-PAGE. The PNA-reactive spots disappeared after removal of O-linked carbohydrate, but not after removal of N-linked carbohydrate. These results suggest that O-linked carbohydrate is present in mrtCD52. The moiety may possibly contribute to a specific antigenic epitope of mrtCD52. PMID- 15288186 TI - Antisperm immunity in assisted reproduction. AB - Antisperm antibodies (ASA) can impair the fertilising capacity of human spermatozoa, acting negatively on sperm motility and cervical mucus penetration, and at the level of in vitro gamete interaction. Several methods attempt to improve the potentially deleterious effects of ASA-mediated infertility: by decreasing ASA production, by removing ASA already bound to sperm, artificial insemination (AIH) and fertilisation (IVF, ICSI). Only ICSI seems able to overcome the problem, with fertilisation and pregnancy rates of ASA-positive patients undergoing this technique in the same range as ASA-negative patients. As immunological infertility is relatively rare, literature in the field is quite scarce and more studies need to be conducted to confirm that embryo quality is not impaired. PMID- 15288187 TI - Male genital tract infection: an influence of leukocytes and bacteria on semen. AB - We have studied the oxidative status of 155 semen samples, 95 originating from healthy individuals and 60 from infertile patients, which were subdivided into two groups: (a) normozoospermic with genitourinary tract infection (GTI); and (b) with pathological spermiogram and GTI. Several phases of infection were observed: with bacterial presence only, bacteria and leukocytes, and leukocytes only, following the routine inflammatory pattern. Leukocyte numbers, bacterial strains, pro- and anti-oxidants, and selected pro-inflammatory cytokines (IL-1 beta, IL-6, IL-8 and TNF-alpha) were studied. Additionally, two oxido-sensitive indices were created (SOD/XO and CAT/XO) in order to follow particular phases of semen infection in two subgroups of patients. Different patterns of activities of pro- and anti-oxidant substances, as well as cytokines, were observed in the studied populations. It was reflected mainly by elevated XO activity in a group of patients with a pathological spermiogram while, in a group of patients with GTI and normozoospermia, xanthine oxidase was normal. In the latter group, oxido sensitive indices were elevated in favour of anti-oxidants; similarly, this occurred with IL-6 levels in comparison to healthy controls. It appears therefore that normozoospermic semen recovers better after infection than pathological semen. Perhaps, IL-6 secretion might be helpful in the observed recovery? PMID- 15288188 TI - Comparison of decidual leukocytes following spontaneous vaginal delivery and elective cesarean section in uncomplicated human term pregnancy. AB - The aim of this study was to quantify and compare leukocyte populations in term decidua basalis and parietalis obtained after spontaneous vaginal delivery (SVD) or elective cesarean section (CS) without labor. Decidua basalis and parietalis samples were obtained from placentas after SVD (n = 20) and after CS (n = 30). Following mechanical disaggregation, leukocytes were purified and stained with monoclonal antibodies. Percentages of leukocyte subclasses within the CD45(+) cell fraction and activated T cells were determined by flow cytometry. No differences were found in the percentages of CD45(+) cells or CD56(bright)CD16(-) uterine natural killer (NK) cells between decidua basalis from SVD and CS or between decidua parietalis from SVD and CS. In decidua basalis and parietalis from SVD, a significantly higher number of CD56(dim)CD16(+) NK cells was found compared to CS. In decidua basalis from SVD, there was a significantly lower percentage of CD14(+) cells and higher percentage of CD19(+) cells compared to CS. The percentage of CD3(+) T cells expressing CD25 or human leukocyte antigen (HLA)-DR was significantly decreased in decidua basalis and parietalis from SVD compared to CS. Comparison of decidua collected after SVD or CS suggests that labor is associated with dynamic changes in the distribution of decidual leukocytes, specifically NK and T cell subpopulations. In particular, the disappearance of the CD4(+)CD25(+) T cell population, which possibly contains a subpopulation of regulatory T cells, may contribute to the initiation of labor. Further investigation into factors affecting decidual leukocytes may expand our understanding of the immunological events at the maternal-fetal interface. PMID- 15288189 TI - Expression of SLAM as a functional and phenotypic marker in women with recurrent miscarriage. AB - In the present work, we investigated the Th1 and Th2 cytokine patterns secreted by infiltrating endometrial lymphocytes from fertile women and from patients with recurrent spontaneous miscarriage (RSM). Moreover, we also analyzed the expression of cytokines in the whole endometrium from fertile and RSM women. Furthermore, we investigated the expression of the activation marker signaling lymphocytic activation molecule (SLAM) a cell surface glycoprotein expressed on lymphocytes that upon engagement boosts IFN-gamma production. Our results showed a slight increase in IL-10 expression in the endometrium of some fertile women, although no significant differences were found in IFN-gamma and IL-5 expression. In contrast, analysis of IFN-gamma production by polyclonal activated lymphocytes from endometrium and/or peripheral blood from fertile women showed a significant increase compared to RSM. Analysis of SLAM protein expression in luteal phase endometrial samples showed a significant increase in the levels of the receptor in RSM women compared to fertile women. These results correlated with a significant augmentation of SLAM levels in peripheral blood T-lymphocytes from RSM patients. Interestingly, after treatment of RSM patients with paternal mononuclear cells, surface-SLAM-expression in T-cells from RSM patients significantly decreased up to levels comparable to those of fertile women. Taken together, our results suggest that endometrial cells have not a defined pattern cytokine-production under pre-implatatory conditions, and SLAM might be a potential marker for the diagnosis of RSM and an indicator useful to follow up the patient response to allogeneic immunotherapy. PMID- 15288190 TI - Cytokines in recurrent pregnancy loss. AB - Cytokines seem to play a critical role in the pathogenesis of unexplained recurrent pregnancy loss (RPL). Th1 cytokines have been shown to exert deleterious effects on pregnancy, inhibiting foetal growth and development. On the other hand, Th2 cytokines have been associated with successful pregnancy. The purpose of this study was to evaluate cytokine production in women with RPL. The studied group comprised 29 women with RPL, with at least three consecutive spontaneous abortions. The control group included 27 women with a history of successful pregnancies and no miscarriage. We determined IL-6 and TNF-alpha production in peripheral blood cultured with LPS, as well as IFN-gamma and TGF beta induced by PHA stimulation. Cytokines were measured by enzyme-linked immunoabsorbant assay (ELISA) using commercial kits (RD, Amersham-Pharmacia). Mann-Whitney test was applied to compare differences between groups. The level of significance was defined at P < 0.05. We observed significantly higher levels of IFN-gamma (355.8 pg/ml versus 98.0 pg/ml; P = 0.01) and a trend toward increased TNF-alpha production (2410.2 pg/ml versus1980.2 pg/ml; P = 0.07) in RPL women as compared to controls. In relation to IL-6 and TGF-beta, no significant difference was detected between RPL and control groups. In agreement with experimental observations, our data support the hypothesis of Th1 cytokine involvement in the pathogenesis of RPL. PMID- 15288191 TI - The promoter region (-800, -509) polymorphisms of transforming growth factor beta1 (TGF-beta1) gene and recurrent spontaneous abortion. AB - Recurrent spontaneous abortion (RSA) is regarded as a common pregnancy complication in southern Iran. The exact causes of RSA are not yet known. Transforming growth factor-beta1 (TGF-beta1) is produced by T regulatory lymphocytes (Treg), which play an important role in the physiology of pregnancy. Several polymorphisms of the TGF-beta1 gene have been reported, some with important correlation with disease severity. In this investigation, the polymorphism of the TGF-beta1 gene at promoter region positions -800 (G/A) and 509 (C/T) was studied in 111 RSA and 110 normal female subjects from southern Iran by PCR-RFLP. Results indicated that at position -800 (G/A) polymorphism, 75.7% of RSA cases and 77.3% of normals were homozygote GG. In addition, 23.4% of cases and 22.7% of normal individuals were heterozygote AG. Only one of the patients appeared to be homozygote AA. None of the normal individuals were found to be homozygote AA at this position. In the case of the -509 (C/T) polymorphism, 38.7% of patients and 28.2% of controls were homozygote CC. While 40.6% of cases and 50.9% of normal individuals were heterozygote CT, 20.7% of RSA cases and 20.9% of controls were homozygote TT. The results indicate that there are no statistically significant differences of genotype distribution and allele frequency between RSA cases and controls at both polymorphic sites. In conclusion, the promoter region polymorphisms of TGF-beta1 at positions -800 (G/A) and -509 (C/T) may not be associated with RSA. PMID- 15288192 TI - Mannan-binding proteins from boar seminal plasma. AB - The interaction of boar seminal plasma proteins and sperm with yeast mannan was investigated by the enzyme-linked binding assay (ELBA) and specific detection of proteins after SDS electrophoresis and blotting using biotinylated derivative of the polysaccharide. Heparin-binding proteins (especially AQN 1 and DQH proteins) and their aggregated forms showed affinity to yeast mannan. Besides that, these proteins were shown to bind to oviductal epithelium. The mannan-binding activity of boar proteins and sperm was inhibited most efficiently by ovomucoid, ovalbumin and N-glycans released from ovalbumin, but not with d-glucose, d-mannose and their phosphates. On the other hand, yeast mannan inhibited both the interaction of boar seminal plasma and sperm with heparin and the binding of these proteins to porcine oviductal epithelium. Yeast mannan immobilized to divinyl sulfone activated Sepharose was used for the isolation of mannan-binding proteins. Proteins adsorbed to the immobilized polysaccharide were analyzed by RP-HPLC, SDS electrophoresis and N-terminal amino acid sequencing. AQN and AWN spermadhesins and DQH protein (names are derived from the N-terminal amino acid sequence) were identified as components of the isolated fraction. The results suggest an involvement of mannan-binding proteins in the formation of the sperm oviductal reservoir in pig. The ability of these proteins to interact both the complex d mannose-containing saccharide structures and the heparin may also play an important role in sperm release from the oviductal reservoir or the capacitation process. PMID- 15288193 TI - The use of potassium dichromate and ethyl alcohol as blood preservatives for analysis of organochlorine contaminants. AB - The "gold standard" for preserving and shipping of human tissue samples for analysis of organochlorine contaminants is freezing. This method can be difficult, costly if using heavy dry ice for shipping, and often unfeasible, especially in less developed countries where electricity and dry ice are frequently rare or absent. Therefore, it is essential that more convenient and practical methods for preservation of blood samples are found. As an alternative to freezing, there have been studies employing potassium dichromate as a preservative for human or cow's milk or ethyl alcohol preservation of blood for dioxin analysis. In this study, four methods were compared to investigate the effectiveness of ethyl alcohol and potassium dichromate as blood preservatives for analysis of dioxins, dibenzofuran, and polychlorinated biphenyl (PCB) congeners. Samples of whole blood from a Dallas, Texas hospital were collected and pooled. Freezing, ethyl alcohol in two concentrations (20% and 40% per volume of sample), and potassium dichromate were used for blood preservation. The blood samples containing potassium dichromate or ethyl alcohol were stored and sent to ERGO laboratory for dioxin analysis and comparison with results from the frozen sample, which was kept frozen at all times until analyzed. This study suggests that potassium dichromate is a suitable alternative to freezing for preservation of whole blood for dioxin, dibenzofuran, and PCB measurements when either lipid or wet weight based results are reported. Potassium dichromate tablets were very easy and convenient to use--two 100 mg tablets (with a dichromate content of about 33 mg each) were added to each bottle containing 65 ml of blood. However, ethyl alcohol at 20% and 40% concentration under the conditions of this pilot study and the analytical method employed did not appear to provide satisfactory preservation when lipid based results are given or when the fat content has to be determined (wet or whole weight). Further research with a larger number of samples, inclusion also of other groups of persistent organic pollutants such as organochlorine pesticides or brominated flame retardants, a longer duration of storage time, and at temperatures greater than US or German room temperature is indicated in order to recommend the routine use of potassium dichromate as preservative for whole blood intended for dioxin, dibenzofuran, and PCB analysis. PMID- 15288194 TI - Polychlorinated biphenyls in Narragansett Bay surface sediments. AB - Polychlorinated biphenyls (PCBs) were evaluated in 41 surface sediments collected from Narragansett Bay, RI in 1997-1998. Highest concentrations of total PCBs (1760 ng/g) were in rivers at the head of the bay and the values decreased southward toward the mouth of the bay, with elevated concentrations in some of the coves. The PCB levels in approximately 43% of the samples exceeded the effects range median (ERM) guideline [Environ. Manage. 19 (1995) 81] indicating possible adverse biological effects at these stations. Principal component analysis (PCA) of the surface sediment PCBs separated the Taunton River samples from the rest of the samples. This result suggests that this river has a different PCB composition and sources than the other areas investigated. It also showed that this river has a limited influence on other bay stations as the adjacent samples downstream did not have the same chemical signature. Congener ratios derived from the PCA were useful in distinguishing stations that had different sources of PCBs than the bulk of the bay sediments. Although Aroclor 1268 and 1270 accounted for <1% of all PCB production, their major components, CB206 and CB209, account for 3-6% of the CBs in most bay samples. This may reflect more local use of these Aroclor mixtures and/or be indicative of their relative stability, compared to less chlorinated mixtures. Using linear alkyl benzenes (LABs) as a marker for sewage derived PCBs suggested that up to 95% of the PCBs at the most contaminated sites in the Seekonk, Providence, and Taunton Rivers were sewage derived. This analysis also showed that there is a high background level (167 ng/g) of PCBs in the Seekonk and Providence River, while the Taunton River had a relatively low background level (23.7 ng/g). At the furthest stations south in the Providence River, the sewage derived PCBs only accounted for 23% of the total which suggests that PCB associated with sewage particles are rapidly deposited and are therefore not the most significant source of these compounds to the lower reaches of Narragansett Bay. PMID- 15288195 TI - Characterisation and analysis of persistent organic pollutants and major, minor and trace elements in Calabash chalk. AB - Analysis of Calabash chalk has been done using energy dispersive X-ray fluorescence spectroscopy (EDXRF), X-ray diffraction (XRD) and pressurised fluid extraction (PFE) followed by gas chromatography (GC) with mass selective detection (MSD). It was found by XRD that the composition of Calabash chalk was an aluminium silicate hydroxide from the kaolin clay group with the possible formula Al(2)Si(2)O(5)(OH)(4). Multi-elemental analysis by EDXRF was able to quantify 22 elements in Calabash chalk including lead at a mean concentration of approximately 40 mg/kg. A range of persistent organic pollutants were identified and quantified in Calabash chalk including alpha lindane, endrin, endosulphan II and p,p'-DDD using PFE-GC-MSD. PMID- 15288196 TI - Present status of POP contamination in Lake Maggiore (Italy). AB - In the last decade, Lake Maggiore has been subject to heavy DDT contamination due to a chemical plant located near the main influent of the Baveno Bay. The freshwater bivalve zebra mussel (D. polymorpha) was used as a bioindicator of several POPs (DDTs, PCBs, HCB, HCHs) to follow their concentration trend after a heavy flood in autumn 2000. Sampling of mollusc specimens were carried out monthly from April 2001 to October 2002 at two different stations in and outside the bay. Results showed worsening of DDT pollution due to the transport of insecticide from contaminated sediments and soils still present in the closed chemical plant site. Levels of about 4.5 microg/g lipids in soft tissues of specimens from the most contaminated site and 2.0 microg/g lipids outside it were found, which are twice those measured before the flood. HCHs and HCB values were always very low, but it was noticed since winter 2001-2002 a sharp increase of PCB pollution, with values of about 3-4 microg/g lipids, not due to the 2000 flood, but probably to improper discharge or release of contaminated sediments from numerous dams located in the watershed. Concentrations of total dioxin-like PCBs reached dangerous levels for the water community. Fish consumption may be a risk for human health especially for resident population, bearing in mind that fish usually have a higher POP concentration than zebra mussel. PMID- 15288197 TI - Residues of dioxins and PCBs in fat of growing pigs and broilers fed contaminated feed. AB - To investigate the kinetics of PCBs and dioxins, 3 week old broilers and 3 month old pigs were fed with a 10-fold diluted feed from the Belgium crisis for one week, followed by a period on clean feed. In the case of broilers this resulted in levels for dioxins, non-ortho and mono-ortho PCBs in fat of 102, 84 and 216 ng TEQ/kg, summarized to 402 ng TEQ/kg. Total levels decreased to 217 and 109 ng TEQ/kg after 1 and 3 weeks on clean feed. Indicator PCB levels decreased from an initial 6.2 mg/kg fat to respectively 3.2 and 1.5 mg/kg. The ratio of indicator PCBs to dioxins TEQs was stable over this period. Levels in back fat of pigs at the end of the exposure period were 26, 15, 82 and 123 ng TEQ/kg for respectively dioxins, non-ortho PCBs, mono-ortho PCBs and the sum. Total TEQ levels decreased to respectively 95, 70, 40, 22 and 12 ng TEQ/kg after 1, 2, 4, 8 or 12 weeks on clean feed. After 12 weeks dioxin levels were around 1 ng TEQ/kg. Indicator PCB levels decreased from 3.48 mg/kg to 2.65, 2.01, 1.25, 0.76 and 0.45 mg/kg fat, again after 1, 2, 4, 8 or 12 weeks on clean feed. Levels of dioxins decreased more rapidly than those of indicator PCBs, also reflected by the ratio of indicator PCBs to dioxins, being 133,000 at the end of the exposure period and 357,000 after week 12. It is concluded that the use of indicator PCBs for dioxins, in the case of a combined exposure, is a safe alternative for screening and in the case of pigs results in an overestimation rather than underestimation of the dioxin levels. PMID- 15288198 TI - Monitoring dioxins and furans in a population living near a hazardous waste incinerator: levels in breast milk. AB - In this study, the concentrations of polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/PCDFs) in breast milk from mothers living in the vicinity of a new hazardous waste incinerator (HWI) were determined. Monitoring was performed after three years of regular operations in the facility and the present results were compared with baseline concentrations obtained in a pre-operational program. PCDD/PCDF levels were determined by HRGC/HRMS in 15 samples. In the present study, PCDD/PCDF concentrations ranged from 4.9 to 39.9 pg I-TEQ/g fat (5.1-46.8 pg WHO-TEQ/g fat), with a median value of 7.7 pg I-TEQ/g fat (9.1 pg WHO-TEQ/g fat). In the baseline survey, PCDD/PCDF concentrations ranged between 5.9 and 17.1 pg I-TEQ/g fat, with a median value of 11.7 pg I-TEQ/g fat. In relation to this, a percentage of reduction of 34.2% was noted. This decrease is in agreement with the relevant reduction found in the dietary intake of PCDD/PCDFs between both surveys. The results of the present study, as well as other recent environmental and biological data, indicate that living in the vicinity of this HWI should not mean additional health risks due to PCDD/PCDFs for the general population. PMID- 15288199 TI - Effect of temperature on the disappearance of four triazine herbicides in environmental waters. AB - The influence of temperature on the disappearance of four s-triazine herbicides, terbuthylazine, simazine, atrazine and prometryn was studied in sea, river and groundwaters spiked with approx. 5 mg l(-1) of each during long-term laboratory incubation. Residues were analyzed by GC-NPD and confirmed by GC-MSD. No clean-up was necessary and a micro on-line method for the determination of herbicide residues was used. The results showed that temperature had little effect on the behaviour of the four herbicides in river and seawaters but strongly affected their behaviour in groundwater. Simazine was the most readily affected compound in sea, river and groundwaters, while terbuthylazine and atrazine were the most persistent in all cases, especially in riverwater. Half-lives ranged from 41 days (constant rate = 0.017 days(-1)) to 196 days (constant rate = 0.003 days(-1)) for simazine (40 degrees C) and terbuthylazine (20 degrees C), respectively, in riverwater. Only for terbuthylazine in riverwater was the remaining percentage at the end of the experiment higher than 50% (58%, 3.21 mg l(-1)). In the other cases, the remaining percentage varied from 4% (0.20 mg l(-1), 40 degrees C) to 43% (2.25 mg l(-1), 20 degrees C) for simazine and terbuthylazine, respectively, in groundwater. PMID- 15288200 TI - First assessment of dioxin emissions from coal-fired power stations in Spain. AB - In this work, the findings of the first assessment of polychlorinated dibenzo-p dioxins and dibenzofurans (PCDD/Fs) in emissions releases to the atmosphere from coal-fired power plants are given. A total of five plants were selected for the study located at different provinces in Spain. In all the cases, the results revealed very low levels, in the range of 0.41 pg I-TEQ/Nm(3). The profile indicated in the majority of the cases predominance of highly chlorinated congeners being OCDD the most important contributor. The findings were also used to estimate contribution of PCDDs/PCDFs emitted from coal-fired power plants in Spain. Individual plant results revealed values below 0.02 g I-TEQ per year and plant. Nevertheless, considering the total coal consumption in Spain in 1997, the values are comparable to those reported in other countries in the range of 0.6 0.7 g I-TEQ per year. Moreover, emission factors were determined considering operating conditions of evaluated plants. In general, large variability was observed and values below 1-5 pg I-TEQ/kg coal were early reached. PMID- 15288201 TI - Role of copper chloride in the formation of polychlorinated dibenzo-p-dioxins and dibenzofurans during incineration. AB - Combustion experiments in a laboratory-scale fluidized-bed reactor were performed to elucidate the role of copper chloride in formation of polychlorinated dibenzo p-dioxins (PCDDs) and dibenzofurans (PCDFs) during model waste incineration. The amounts of PCDDs and PCDFs formed, the homologue profiles, and the isomer distributions were measured in the flue gas from incineration of model wastes containing various levels of copper. A correlation was found between the Cu content of the waste and the proportion of each congener. An increase in copper enhanced the formation of certain congeners, showing that copper acts as a catalyst for formation of PCDDs and PCDFs. An increase in the copper content of the waste decreased the CO concentration in the flue gas and reduced the formation of PCDDs and PCDFs during incineration. This indicates that copper also works as an oxidation catalyst to promote combustion, leading to lower concentrations of products of incomplete combustion. It is indispensable to consider both roles of the catalyst, i.e., enhancement and suppression, in the formation of PCDDs and PCDFs during waste incineration, which are estimated separately from the isomer distributions and the amounts of PCDDs and PCDFs formed. PMID- 15288202 TI - Isolation of an imaginal disc growth factor homologue from Pieris rapae and its expression following parasitization by Cotesia rubecula. AB - Endoparasitoid insects introduce maternal factors into the body of their host at oviposition to suppress cellular defences for the protection of the developing parasitoid. We have shown that transient expression of polydnavirus genes from a hymenopteran parasitoid Cotesia rubecula (CrPDV) is responsible for the inactivation of hemocytes from the lepidopteran host Pieris rapae. Since the observed downregulation of CrPDV genes in infected host tissues is not due to cis regulatory elements at the CrV1 gene locus, we speculated that the termination of CrPDV gene expression may be due to cellular inactivation caused by the CrV1 mediated immune suppression of infected tissues. To test this assumption, we isolated an imaginal disc growth factor (IDGF) that is expressed in fat body and hemocytes, the target of viral infection and expression of CrPDV genes. Time course experiments showed that the level of P. rapae IDGF is not affected by parasitization and polydnavirus infection. However, the amount of highly expressed genes, such as storage proteins, arylphorin and lipophorin, are significantly reduced following parasitization. PMID- 15288203 TI - Dissecting chill coma recovery as a measure of cold resistance: evidence for a biphasic response in Drosophila melanogaster. AB - Cold resistance in insects has traditionally been measured in terms of survival following a stress, but alternative methods are increasingly being used because of their relevance to the ecology of organisms and their utility in characterizing variation among species, populations and individuals. One such method capable of discriminating among Drosophila species and conspecific Drosophila populations from different environments is adult chill coma recovery time, the time taken for adults to become active again after being knocked down by a cold stress. Here we characterized the chill coma response of D. melanogaster in detail. Adults were exposed to a range of temperatures and stressful periods prior to measuring recovery. Recovery from chill coma in D. melanogaster was biphasic; as flies were stressed under cooler temperatures, recovery times leveled off and then decreased before sharply increasing again as mortality starts to occur. This biphasic response has previously been observed in D. subobscura where it has a somewhat different shape. A second mechanism therefore acts at relatively lower temperatures to ameliorate the effects of the cold stress. When D. melanogaster were reared at 19 and 25 degrees C for two generations, the shape of the curve relating temperature to recovery time was similar, but flies from the warmer temperature had longer recovery times and showed responses that leveled off and then decreased at relatively higher temperatures. As exposure time to cold stress was increased, recovery times also increased except at mild stress levels. Chill coma recovery in D. melanogaster is a complex trait and likely to reflect multiple underlying components. PMID- 15288204 TI - Octopamine receptors in the honeybee (Apis mellifera) brain and their disruption by RNA-mediated interference. AB - Octopamine plays important neuromodulatory roles in the honeybee brain. Accordingly, mRNA from a recently identified honeybee octopamine receptor (AmOA1) is distributed throughout the brain. We have evaluated the occurrence of AmOA1 in the antennal lobe (AL) as well as rest of the brain (RB) by western blotting using an antiserum raised against a peptide selected from AmOA1 sequence. In addition to an expected band (78 kDa in the AL), one additional band (72 kDa) was identified from the AL and four bands (48, 60, 72 and 78 kDa) were observed in the RB. These bands were also recognized with antiserum against a different peptide segment from an octopamine receptor ortholog from the fruitfly (OAMB). Significant sequence identity with the peptide segment used to generate the antiserum was only found with OAMB and its splice variants in fruitfly; it was less conserved in other biogenic amine receptors from honeybee and other insects. Furthermore, western blot analysis performed on brains with dsRNA-treated antennal lobes showed a decrease in the intensity of all four bands. This suggests that AmOA1 antiserum specifically recognizes one or more types of AmOA1 receptors in the honeybee brain. We extend our earlier study of RNAi to quantify the rate of spread of dsRNA from a localized injection to other neuropils. PMID- 15288205 TI - Plasticity in juvenile hormone in male burying beetles during breeding: physiological consequences of the loss of a mate. AB - Burying beetles, Nicrophorus orbicollis, have facultative biparental care. They bury and prepare small vertebrate carcasses that provide food for their young. Here we establish the juvenile hormone (JH) profiles of paired females, paired males and single males and investigate some of the environmental and social factors that may affect these profiles. Before larvae hatch JH profiles of paired males and females were similar. However, after larvae hatch and during brood care, JH titers of females were very high and those of single males were significantly higher than those of paired males. We tested the hypothesis that higher JH was a response to the need for increased parental care by manipulating brood size. Although JH titers of single males caring for small versus large broods were not significantly different, when comparing JH titers and larval growth (a measure of parental effort), a significant positive correlation emerged. In contrast, we found that food quality had no effect on JH levels suggesting that increased feeding by males and females after carcass discovery cannot explain the elevation of JH. The regulation of JH in male burying beetles appears thus to be dependent on the presence of a mate and on critical stimuli from young. PMID- 15288206 TI - Behavioural correlates of phenotypic plasticity in mouthpart chemoreceptor numbers in locusts. AB - Rearing locusts in an impoverished chemosensory environment leads to fewer chemoreceptors developing on the mouthparts and antennae as adults but the behavioural relevance of these changes remains unknown. To address this question, locusts were reared for the final two larval stadia on either a single, nutritionally near-optimal synthetic food ('plain' pretreatment), or a diet comprising two nutritionally complementary foods containing two added flavours ('mix' pretreatment). Insects reared on the 'mix' diet had a mean 20% more chemosensilla on the maxillary palps than those fed on the 'plain' diet. Following an equilibration period, when all newly moulted adults could feed on two nutritionally complementary foods, insects were food deprived for 2 or 6 h, and then given a test meal of a single balanced food at one of two dilutions whilst their behaviour was recorded. 'Mix'-pretreated locusts had a shorter latency to feed and were more likely to reject the test food upon first contact if deprived for only 2 h; but if they did take a meal it lasted longer and contained fewer pauses. Using sensilla number as a covariate removed the statistical significance of pretreatment regime, indicating that sensilla number, or some close correlate of it, can largely account for the variation in behaviour. This suggests that sensilla numbers are behaviourally relevant; particularly where locusts are not greatly food deprived and faced with marginally acceptable foods. PMID- 15288207 TI - Cadmium effects on development and reproduction of Oncopeltus fasciatus (Heteroptera: Lygaeidae). AB - Newly hatched nymphs of the insect Oncopeltus fasciatus were exposed to various concentrations of CdCl2 administered in drinking water until the end of adult life. Significant nymphal mortalities were observed at concentrations above 30 mg Cd/l (corresponding to the LC50). The duration of the nymphal stages increased in proportion to the Cd concentration; at the lowest Cd concentration of 10 mg Cd/l, the median duration was significantly prolonged by one day, while at the highest concentration of 100 mg Cd/l it was increased by 10 days over the control group. The weight of newly emerged adults lineally decreased with Cd concentration, being reduced to half the weight of controls at 100 mg Cd/l. In addition, a proportionality between delay in development and weight reduction was found in Cd treated adults. Survival of adult females was decreased at concentrations higher than 10 mg Cd/l, while males were only affected at 30 mg Cd/l or higher doses. Reproduction was the most affected parameter. Oviposition rate, fecundity and fertility of females exposed to 10 mg Cd/l were significantly lower than controls (73%, 58% and 55% relative to controls, respectively). Hatchability of the eggs laid by treated females was also reduced. These results show that development and reproduction of O. fasciatus are seriously impaired at sublethal Cd concentrations. PMID- 15288208 TI - Artificial parthenogenesis and control of voltinism to manage transgenic populations in Bombyx mori. AB - In order to improve the management of transformed populations in a routine application of transgenesis technology in Bombyx mori, we modified its mode of reproduction and its voltinism. On one hand, after a stable integration of the gene of interest by transgenesis, it is preferable to maintain this gene in an identical genomic context through successive generations. This can be obtained by artificial parthenogenetic reproduction (ameiotic parthenogenesis) giving isogenic females identical to their transformed mother. On the other hand, it is essential to obtain continuous generations (polyvoltinism) after microinjection, in order to screen positive transgenic insects and study genetics and insertion of the transgene. Thereafter, it is more convenient to store these populations, as diapause eggs before their use in biotechnology application. We obtained such polyvoltine parthenoclones, first by selection for a parthenogenetic character in polyvoltine races, and second, by selection for a polyvoltine character in a parthenogenetic, but diapausing clone of B. mori. As diapause was directly under the control of diapause hormone (DH), we also tested direct injection of DH in female pupae of polyvoltine strains, as well as anti-DH antibody treatment to eliminate diapause in univoltine strains. We discussed the advantages and limitations of these methods and proved the feasibility in obtaining polyvoltine parthenoclones and determining the voltinism in B. mori. These methods would permit us to improve the management of populations used in transgenesis technology. PMID- 15288209 TI - Chemical basis for inter-colonial aggression in the stingless bee Scaptotrigona bipunctata (Hymenoptera: Apidae). AB - Inter-colonial aggression was tested using three colonies of Scaptotrigona bipunctata in a natural setting when their nests were moved and by artificial contact between individuals. Examination of the cuticular lipids of individuals from two colonies kept under identical conditions showed clear differences in their cuticular hydrocarbon profiles. The cuticular lipids were a mixture of hydrocarbons (saturated and unsaturated alkanes and alkenes) within the range of C23-C29. The use of multivariate analysis (PCA and discriminant analysis) showed that seven of the identified surface compounds are enough to separate workers from colonies A and B from each other. PMID- 15288210 TI - The 2005 United States budget: wasteful expenditures, foregone opportunities. PMID- 15288211 TI - Contraceptive failure in the United States. AB - This review provides an update of previous estimates of first-year probabilities of contraceptive failure for all methods of contraception available in the United States. Estimates are provided of probabilities of failure during typical use (which includes both incorrect and inconsistent use) and during perfect use (correct and consistent use). The difference between these two probabilities reveals the consequences of imperfect use; it depends both on how unforgiving of imperfect use a method is and on how hard it is to use that method perfectly. These revisions reflect new research on contraceptive failure both during perfect use and during typical use. PMID- 15288212 TI - Effect of four oral contraceptives on hemostatic parameters. AB - This is the first double-blind, controlled, randomized study comparing the effect of different estrogen components in oral contraceptives (OCs) on hemostasis variables. Four groups of 25 women each were treated for six cycles with monophasic combinations containing 21 tablets with either 30 microg ethinylestradiol (EE) + 2 mg dienogest (DNG) (30EE/DNG), 20 microg EE + 2 mg DNG (20EE/DNG), 10 microg EE + 2 mg estradiol valerate (EV) + 2 mg DNG (EE/EV/DNG) or 20 microg EE + 100 microg levonorgestrel (LNG) (EE/LNG). Blood samples were taken on Days 21-26 of the control cycle and on Days 18-21 of the first, third and sixth treatment cycle. Treatment with all four OCs caused an increase in levels of fibrinogen, prothrombin fragment 1+2, D-dimer, plasminogen, plasmin antiplasmin complex and an increase in protein C activity, a decrease in antithrombin activity, tissue-plasminogen activator (t-PA) and plasminogen activator inhibitor (PAI), and a slight decrease in the sensitivity to activated protein C, but no significant change in that of the thrombin-antithrombin complex. In users of the DNG-containing OCs, the reduction in total and free protein S, and in t-PA and PAI was dependent on the EE dose, while factor VII activity was elevated, but not significantly different from EE/LNG. The results are in agreement with those of previous studies. The effects of EE/EV/DNG on total and free protein S and on t-PA and PAI were lower than those of 20EE/DNG, suggesting that the impact of 2 mg EV on several hemostasis variables is less than that of 10 microg EE. The results show an antagonistic effect of LNG on the EE-induced rise of factor VII activity and fragment 1+2 and on the EE-dependent reduction of total and free protein S. PMID- 15288213 TI - Lubricants containing N-9 may enhance rectal transmission of HIV and other STIs. AB - It has been shown that men who have sex with men actively seek lubricants that contain nonoxynol-9 (N-9) because they believe that N-9 may help to prevent infection by HIV. However, indirect evidence suggests that N-9 may actually enhance infection. Microscopic examination of rectal lavage and biopsy specimens collected at different time points following rectal application of a lubricant containing 2% N-9 showed rapid exfoliation of the rectal epithelium. Because the rectal epithelium protects target cells in the submucosa from HIV, we conclude that lubricants containing N-9 should be avoided during rectal sex. PMID- 15288214 TI - Transcervical polidocanol as a nonsurgical method of female sterilization: a pilot study. AB - We have studied the effects of transcervically administered polidocanol on uterine and fallopian tube morphology in Wistar rats and Rhesus monkeys. Polidocanol is a synthetic, long-chain fatty acid that is widely used as a sclerosing agent in Europe. The goal of the study was to determine whether polidocanol would safely cause tubal occlusion in an animal model. Twenty female Wistar rats and three female Rhesus monkeys underwent transcervical injection of polidocanol or physiological saline. The animals were followed for 30 days and then a lower laparotomy was performed, with excision of the entire upper reproductive tract. Specimens were subsequently examined for macroscopic and microscopic changes. Only cyclic changes were observed in the control animals of both species. Fifteen macroscopic and 37 microscopic changes were observed in the uterine horns of the 10 rats treated with polidocanol. There was no observed effect in the monkey fallopian tube. These results suggest that species differences that exist between rodents and primates may influence the effects of transcervical polidocanol. Experiments using a primate model are needed as proof of concept prior to human studies of candidate agents for transcervical tubal sterilization. PMID- 15288215 TI - A randomized comparison of sublingual and vaginal misoprostol for cervical priming before suction termination of first-trimester pregnancy. AB - This randomized trial compares the efficacy and side effects of sublingual and vaginal misoprostol for cervical priming before first-trimester pregnancy termination. One-hundred pregnant women between 6 and 12 weeks of gestation opting for termination of pregnancy by suction evacuation were included in this study. The women were randomly allocated into two groups. Group 1 received 400 microg of sublingual misoprostol and group 2 received 400 microg of vaginal misoprostol 2 h prior to suction evacuation. The abortion was carried out by suction evacuation using a Karman's cannula attached to an electrically operated suction machine under intravenous analgesia. Baseline cervical dilatation, duration of the procedure, operative blood loss, side effects and complications were noted in both groups. There was a significant difference between the sublingual and vaginal misoprostol groups with respect to mean cervical dilatation (8.6 mm vs. 6.8 mm, p < 0.05). However, the duration of the procedure (3.03 min vs. 3.16 min) and the amount of blood loss (29 mL vs. 31.2 mL) were not significantly different between the two groups. The women in the sublingual group experienced significantly more shivering and preoperative vaginal bleeding (68% vs. 56%, p < 0.05). None of the women in the two groups had either uterine perforation or excessive hemorrhage. In our study, sublingual misoprostol (400 microg) was significantly more effective in facilitating cervical dilation prior to surgical abortion than vaginal misoprostol. PMID- 15288216 TI - Misoprostol as the primary agent for medical abortion in a low-income urban setting. AB - The purpose of this study was to assess the outcomes of early medical abortion in an inner-city hospital abortion service, using misoprostol as the primary agent. This was a retrospective chart review from July 2001 through December 2002. Women were eligible if they had a viable pregnancy with gestational age 8 weeks or less by transvaginal ultrasound and no medical contraindications. Two doses of 800 microg misoprostol were administered vaginally, 24 h apart. Initial follow-up was scheduled 2-3 days later. Of the 440 women who underwent medical abortion, 373 (90.8%, 95% confidence interval (CI) 88-94%) completed abortion medically, 38 (9.2%) had uterine aspiration and the remainder had incomplete or no follow-up. Of uterine aspirations, 11 were medically indicated, giving a rate of indicated aspiration of 2.7%. Gestational age, age, gravidity, parity, past abortion history, ethnic group and payer did not significantly correlate with overall rate of aspiration or rate of follow-up, but gestational age was correlated with medically indicated aspiration. Among 57 women who reported a time of tissue passage, the mean time from initial misoprostol dose was 8.5 h (95% CI 6.5-13 h). PMID- 15288217 TI - Physicians' knowledge and opinions about medication abortion in four Latin American and Caribbean region countries. AB - To examine physicians' knowledge and attitudes in regard to medication abortion, we conducted focus-group discussions with general practice physicians and obstetrician-gynecologists in Honduras, Mexico, Nicaragua and Puerto Rico. Physicians were familiar with the practice of several types of medication and surgical abortion methods. Medication abortion with misoprostol is most common among women of higher socioeconomic status and is prescribed by physicians, pharmacists or self-administered. Conflicting opinions regarding safety, efficacy, cost, potential for self-medication and acceptability emerged; some participants expressed hope that medical abortion would reduce the risks associated with unsafe abortion, while others contended that drug distribution and self-medication without proper counseling could be problematic. Participants noted a lack of reliable sources of information for both providers and women, and expressed interest in strategic dissemination of information. PMID- 15288218 TI - Teaching sex education improves medical students' confidence in dealing with sexual health issues. AB - Medical students at the University of Edinburgh in Scotland can volunteer to participate in an undergraduate options project that gives them the opportunity to provide sex education for secondary school (high school) pupils. Using a questionnaire presenting a set of fictional case histories, we assessed medical students' theoretical confidence at dealing with sexual health consultations. Students who had participated in delivering peer-led sex education felt significantly more confident at discussing sexual health issues with patients of all age groups (p = 0.001) than students who had not participated in the project. All students felt more comfortable seeing patients of the same gender as themselves but more than half felt that their training left them generally ill equipped to handle sexual health consultations. PMID- 15288219 TI - HIV diagnostic tests: an overview. AB - This article provides an overview of different HIV diagnostic tests, describing how they work and the advantages and limitations of the various tests. This article also briefly reviews the structure and genetic diversity of HIV, the mechanism the virus uses to infect host cells and the different phases of HIV infection. This information is included to facilitate understanding of how HIV diagnostic tests work and what molecular markers these tests use to pinpoint an HIV infection. PMID- 15288220 TI - Comparison of surface morphology and composition of intrauterine devices in relation to patient complaints. AB - The morphology and the related composition of T-type intrauterine devices (IUDs) made of gold-plated copper were studied in relation to patient complaints. The removed IUDs were examined without pretreatment by scanning electron microscopy and energy dispersive x-ray spectroscopy. Besides the previously found incrustation formation, lamination cracks were observed on the surface of gold plated copper IUDs causing various side effects (abdominal pain, inflammatory complication). A possible explanation for lamination cracks could be the difference in the heat expansion of the copper and its gold-plated surface and the great difference between the strong acidic galvanizing pH during production and the in utero pH. Stability tests in combination with morphology screening of such IUDs could be required during their development phase to avoid such side effects. PMID- 15288221 TI - The release of cupric ion in simulated uterine: New material nano-Cu/low-density polyethylene used for intrauterine devices. AB - With the development of IUDs, a number of copper-bearing devices are now commercially available, including the copper-T, the Multiload and the copper-T in various other forms, so-called "the second-generation" IUDs. In this article, we report on nano-Cu/low-density polyethylene composite as a potential copper carrier in IUD. Two issues for the new material are addressed: the effectiveness of polymers in reducing the initial burst in cupric ion release and the amount and pattern of continuing release. The aim of this study was to investigate copper ion release from this composite as a basis for considering its used in an IUD. PMID- 15288222 TI - The role and comparison of two techniques of paracervical block for pain relief during suction evacuation for first-trimester pregnancy termination. AB - This prospective study assessed the role and compared two techniques of paracervical block (PCB) for pain relief during suction evacuation for first trimester termination of pregnancy following cervical priming with misoprostol. One-hundred and thirty-five women undergoing suction evacuation up to 12 weeks of gestation were randomized into three groups: (a) 5 mL of 1% lignocaine injected at the 4 and 8 o'clock positions of the vaginal vault; (b) 5 mL of 1% lignocaine injected at the 4 and 8 o'clock positions of the cervix and (c) no PCB. Pain scores during PCB, cervical dilatation and during and after suction evacuation were compared among the three groups. The sedation and satisfaction levels were also compared. There were no statistically significant differences in the pain levels during PCB, cervical dilatation and suction evacuation and in the satisfaction levels among the three groups. Patients with a lighter sedation level experienced more pain. In conclusion, PCB did not improve the pain levels during first-trimester suction termination of pregnancy after cervical priming with misoprostol and use of intravenous sedation, regardless of whether the local anaesthetic was injected into the cervix or the vaginal vault. PMID- 15288223 TI - Assessment of menstrual blood loss in women with ideopathic menorrhagia using the frameless levonorgestrel-releasing intrauterine system. AB - OBJECTIVE: This study was conducted to evaluate the effect of a "low-dose" levonorgestrel (LNG)-releasing intrauterine system (IUS) on the amount of menstrual blood loss (MBL) in women with ideopathic menorrhagia. METHODS: Menstrual blood loss was assessed with the visual assessment technique in 12 Belgian FibroPlant-LNG users with menorrhagia. In addition, ferritin levels were measured. RESULTS: The median MBL, evaluated by the visual scoring technique, decreased by more than 90%. The ferritin levels increased significantly during treatment with the levonorgestrel system. CONCLUSION: This study confirms previous MBL studies conducted with the FibroPlant-LNG IUS demonstrating the efficacy of the LNG-IUS to significantly reduce the amount of MBL in women with menorrhagia. The strong endometrial suppression is the principal mechanism explaining the effect on MBL. The therapeutic effect of this contraceptive method is highly desirable, particularly in women with heavy bleeding or anemia, as other treatment modalities are less effective, more costly, more invasive or inaccessible. The simple design characteristics and anchoring system account for minimizing the occurrence of complaints of pain and expulsion. PMID- 15288224 TI - Assessment of menstrual blood loss in Belgian users of the frameless copper releasing IUD with copper surface area of 200 mm2 and users of a copper levonorgestrel-releasing intrauterine system. AB - OBJECTIVE: This study was conducted to evaluate the effect of a miniaturized frameless copper IUD (GyneFix 200 small) and a copper-levonorgestrel (GynePlant) intrauterine system (IUS) on the amount of menstrual blood loss (MBL). METHODS: In 60 Belgian women using GyneFix 200 and 21 using GynePlant, MBL was assessed with the visual assessment technique. RESULTS: MBL scores in GyneFix 200 users did not change from baseline during the mean observation period of 31 months. In GynePlant users, mean MBL scores decreased by at least 50% in all but one user. CONCLUSION: The impact of copper IUDs on MBL can be minimized by reducing the surface area of the foreign body. Reduction of MBL, without causing amenorrhea, can be obtained by adding levonorgestrel. PMID- 15288225 TI - Assessment of menstrual blood loss in Brazilian users of the frameless copper releasing IUD with copper surface area of 330 mm2 and the frameless levonorgestrel-releasing intrauterine system. AB - OBJECTIVE: This study was conducted to evaluate the effect of two types of IUDs on the amount of menstrual blood loss (MBL): the frameless copper-releasing intrauterine device (IUD) with copper surface area of 330 mm2 (GyneFix; Contrel Research, Ghent, Belgium) and the frameless levonorgestrel (LNG)-releasing intrauterine system (IUS) releasing 14 microg per day (FibroPlant-LNG; Contrel Research). Heavy and abnormal MBL is the main reason for discontinuation of intrauterine devices. METHODS: In 20 Brazilian women using GyneFix 330 and 32 using FibroPlant-LNG, respectively, MBL was measured by the quantitative alkaline hematin technique. In addition, ferritin levels were measured in GyneFix 330 and FibroPlant-LNG users. RESULTS: MBL with GyneFix 330, measured over a 24-month period, increased but was less when compared with TCu380A. Ferritin levels with GyneFix 330 were not affected in contrast with TCu380A. In FibroPlant-LNG users, mean MBL decreased by about 90% and ferritin levels increased significantly. CONCLUSIONS: The authors confirm earlier reports that, especially for women with low body iron stores and heavy menstrual bleeding, there is an order of preference for IUD use to minimize MBL. The choice should first be a progestin releasing IUS, then a copper IUD, which has the least effect on menstrual bleeding, such as the frameless GyneFix IUD. PMID- 15288226 TI - An additive factor analysis of the effect of depression on the reaction time of old patients. AB - Depressed participants display longer reaction times (RTs) than control participants. The present study was aimed at deciphering which stages of processing are affected by depression in old adults. Sixteen old depressed patients and 16 old healthy volunteers performed a two-choice visual RT task. Signal intensity, stimulus-response mapping and foreperiod duration were manipulated so as to affect the stages of stimulus preprocessing, response selection and motor adjustment, respectively. Reaction time data suggest that depression spares the stage of stimulus preprocessing but affects the stage of motor adjustment. An analysis of the error rate leaves open the possibility that depression alters the stage of response selection. PMID- 15288227 TI - Kinesthetic estimation of the main orientations from the upright and supine positions. AB - This work investigated the accuracy of the perception of the main orientations (i.e., vertical and horizontal orientations) with the kinesthetic modality--a modality not previously used in this field of research. To further dissociate the influence of the postural and physical verticals, two body positions were explored (supine and upright). Twenty-two blindfolded participants were asked to set, as accurately as possible, a rod to both physical orientations while assuming one of the two body positions. The horizontal was perceived more accurately than the vertical orientation in the upright position but not in the supine position. Essentially, there were no differences in the supine position because the adjustments to the physical vertical were much more accurate than they were in the upright position. The lower accuracy in the estimation of the vertical orientation observed in the upright position might be linked to the dynamics associated with the maintenance of posture. PMID- 15288228 TI - Total sleep deprivation increases the costs of shifting between simple cognitive tasks. AB - In two experiments we studied the effects of one night of total sleep deprivation on task-shift costs. In different conditions shifts were between types of judgment (extradimensional shifts) and between stimulus-response mappings (intradimensional shifts). In addition, with an alternating-runs procedure we used short and long response-to-stimulus intervals and also external precues to vary the opportunities for advance configuration of task sets. Under all conditions sleep deprivation increased shift costs derived from the 20% slowest reaction times, which were insensitive to the opportunities for advance configuration. Shift costs derived from the 20% fastest reaction times were increased only for extradimensional shifts. As indicated by congruency effects, the increase of shift costs after a night without sleep cannot be attributed to increased interference between competing task sets. The findings suggest that total sleep deprivation increases task-set instability and thus lapsing, in particular in conditions with long stimulus-to-response intervals and in shift trials. In addition total sleep deprivation seems to increase the duration of an exogenously controlled process involved in extradimensional shifts. PMID- 15288229 TI - Mental rotation depends on the number of objects rather than on the number of image fragments. AB - It is often intuitively assumed that disconnected image fragments result in a representation of separate objects. When objects are partly occluded, disconnected image fragments can still result in a representation of a single object, based on visual completion. In a simultaneous matching task, displays showing one object, partly occluded objects, or two objects were compared with each other. When only a translation was required to match pairs of displays, one object displays were matched faster than both occluded-object and two-object displays, which did not differ significantly from each other. When mental rotation and translation were required, the one-object displays were again matched the fastest. In addition, an advantage for occluded-object displays compared with two-object displays was found. We conclude that when the generation of a mental representation is likely, object-based connectedness determines object matching. Mental rotation then seems to depend on the number of objects rather than on the number of image fragments. PMID- 15288230 TI - Gender differences on the mental rotations test: a factor analysis. AB - The purpose of the present study was to examine possible gender differences in strategy when completing the mental rotations test. Two experiments examined gender differences and the factor structure on outcomes that can be obtained on this test. Experiment 1 involved large groups testing and Experiment 2 used small groups. Factor analytic results in both experiments generally supported the notion that items with one wrong and one blank response or one correct and one blank reflect reluctance to guess, whereas one correct and one wrong or two wrong answers reflect propensity to guess. Even though the factor structure was the same in males and females, the data provided mitigated support for the hypothesis that males have a higher propensity to guess and females show a greater reluctance to guess. Findings are discussed in terms of their implications for the interpretation of gender differences on the MRT. PMID- 15288231 TI - Detection of linear temporal drift in sound sequences: empirical data and modelling principles. AB - Three experiments determined the perceptual threshold (JND) for detecting tempo change; i.e. linear continuous increase or decrease in inter onset interval (IOI) in sequences with 2-9 brief sounds. JND decreased as a power function of the number of intervals presented (Expt. 1). JND increased with IOI and exhibited breakpoints in this respect close to 1 and 1.4 s (Expt. 3), in agreement with previous results for serial interval production. No interaction was apparent between IOI and number of intervals (Expt. 2). None of the experiments showed any effect of direction (increasing or decreasing intervals). The results are inconsistent with several conceivable principles for perceiving tempo change, except for one in which the external intervals are compared with the intervals generated by an internal, periodic process. The plausibility of this principle is discussed in the light of recent research on sensorimotor synchronisation. PMID- 15288232 TI - An ageing problem. PMID- 15288233 TI - Involving children in paediatric oncology decision-making. PMID- 15288234 TI - Issues undermining provision of diagnostic imaging in the UK. PMID- 15288235 TI - Gene-based therapy in prostate cancer. AB - Prostate cancer is one of the commonest causes of illness and death from cancer. Radical prostatectomy, radiotherapy, and hormonal therapy are the main conventional treatments. However, gene therapy is emerging as a promising adjuvant to conventional strategies, and several clinical trials are in progress. Here, we outline several approaches to gene therapy for prostate cancer that have been investigated. Methods of gene delivery are described, particularly those that have commonly been used in research on prostate cancer. We discuss efforts to achieve tissue-specific gene delivery, focusing on the use of tissue-specific gene promoters. Finally, the present use of gene therapy for prostate cancer is evaluated. The ability to deliver gene-therapy vectors directly to prostate tissue, and to regulate gene expression in a tissue-specific manner, offers promise for the use of gene therapy in prostate cancer. PMID- 15288236 TI - Amyloidosis presenting as malignant disease. PMID- 15288237 TI - Role of liver transplantation for hepatobiliary malignant disorders. AB - The role of liver transplantation for hepatobiliary malignant disorders remains controversial and will remain so until several crucial issues are resolved, the main difficulty being the shortage of organ donors. Furthermore, a consensus needs to be reached within the transplantation community on the tumour stage at which each disorder is too advanced to be salvaged by liver transplantation. Despite these limitations, there are generally accepted criteria that define when transplantation can, and should, be offered for hepatobiliary malignant disorders. PMID- 15288238 TI - Drug interactions in oncology. AB - Drug interactions are an ongoing concern in treatment of cancer, especially when cytotoxic drugs are being used. However, the clinical relevance of these interactions is not always investigated. Drug interactions can be pharmaceutical, pharmacokinetic, or pharmacodynamic. They can also be wanted (eg, use of ciclosporin to enhance the oral bioavailability of paclitaxel); unwanted (eg, combination of the antiviral agent sorivudine and oral fluorouracil analogues can lead to fatal complications); between cytotoxic drugs, cytotoxic drugs and non cytotoxic drugs; or with pharmaceutical vehicles. Potential interactions between anticancer drugs and over-the-counter or alternative medicines and herbs should not be underestimated. More attention should be given to the recognition of potential drug interactions in the preclinical and early clinical development phase of a new anticancer drug. Here, we provide a comprehensive overview of drug interactions, with selected examples. PMID- 15288239 TI - The present and future role of photodynamic therapy in cancer treatment. AB - It is more than 25 years since photodynamic therapy (PDT) was proposed as a useful tool in oncology, but the approach is only now being used more widely in the clinic. The understanding of the biology of PDT has advanced, and efficient, convenient, and inexpensive systems of light delivery are now available. Results from well-controlled, randomised phase III trials are also becoming available, especially for treatment of non-melanoma skin cancer and Barrett's oesophagus, and improved photosensitising drugs are in development. PDT has several potential advantages over surgery and radiotherapy: it is comparatively non-invasive, it can be targeted accurately, repeated doses can be given without the total-dose limitations associated with radiotherapy, and the healing process results in little or no scarring. PDT can usually be done in an outpatient or day-case setting, is convenient for the patient, and has no side-effects. Two photosensitising drugs, porfirmer sodium and temoporfin, have now been approved for systemic administration, and aminolevulinic acid and methyl aminolevulinate have been approved for topical use. Here, we review current use of PDT in oncology and look at its future potential as more selective photosensitising drugs become available. PMID- 15288240 TI - Induction chemotherapy followed by radiotherapy in Merkel-cell carcinoma. PMID- 15288242 TI - Taking cover. PMID- 15288241 TI - Development of gliomas: potential role of asymmetrical cell division of neural stem cells. AB - Asymmetrical cell division is a mechanism that gives rise to two daughter cells with different proliferative and differentiative fates. It occurs mainly during development and in adult stem cells. Accumulating evidence suggests that tumour cells arise from the transformation of normal stem cells. Here, we propose that the asymmetrical mitosis potential of stem cells is associated with the generation of migrating tumour progenitors. Application of this speculative model to glioma proposes that the sites where tumour-initiating stem cells reside are indolent and distinct from the tumour mass, and implies that the tumour mass is continuously replenished with new migrating tumour cells from these clinically silent regions. This hypothesis offers explanations for our inability to cure glioblastoma and points to asymmetrical division as a new potential therapeutic target. PMID- 15288243 TI - Integrating virtual screening in lead discovery. AB - Target- and ligand-based virtual screening have emerged as resource-saving techniques that have been successfully applied to identify novel chemotypes in biologically active molecules. Eight confirmed virtual screening hits have recently been described and are discussed in this review, with focus on the workflow. These are then evaluated in the light of pharmacokinetics prediction (e.g. Caco-2 permeability, cytochrome P450 inhibition and hERG binding). We anticipate problems for five of these hits (e.g. cardiac toxicity), which warrant further experiments. Future challenges include dynamic tautomer/protonation treatment for both ligands and targets and improved pre- and post- virtual screening filters. PMID- 15288244 TI - Target-biased scoring approaches and expert systems in structure-based virtual screening. AB - Structure-based virtual screening followed by selection of a top fraction of the rank-ordered result list suffers from many false positives and false negatives because the general scoring functions are not accurate enough. Many approaches have emerged to address this problem by including knowledge about the specific target in the scoring and selection steps. This target bias can include requirements for critical interactions, use of pharmacophore patterns or interaction patterns found in known co-crystal structures, and similarity to known ligands. Such biases are implemented in methods that vary from filtering tools for pre- or post-processing, to expert systems, quantitative (re)scoring functions, and docking tools that generate target-biased poses. PMID- 15288245 TI - High-throughput docking as a source of novel drug leads. AB - Receptor-based virtual screening has become a viable source of novel leads in the pharmaceutical industry. The rapidly growing availability of structural information across protein families, the accessibility to increased computational power at affordable cost, as well as an improved understanding on how to effectively apply virtual screening technologies has contributed to their emergence. Nonetheless, continued improvement in the accuracy of scoring functions and a greater understanding of protein mobility is critical to advance the technology further. PMID- 15288246 TI - Tools for target identification and validation. AB - Reliable technologies for addressing target identification and validation are the foundation of successful drug development. Microarrays have been well utilized in genomics/proteomics approaches for gene/protein expression profiling and tissue/cell-scale target validation. Besides being used as an essential step in analyzing high-throughput experiments such as those involving microarrays, bioinformatics can also contribute to the processes of target identification and validation by providing functional information about target candidates and positioning information to biological networks. Antisense technologies (including RNA interference technology, which is recently very 'hot') enable sequence-based gene knockdown at the RNA level. Zinc finger proteins are a DNA transcription targeting version of knockdown. Chemical genomics and proteomics are emerging tools for generating phenotype changes, thus leading to target and hit identifications. NMR-based screening, as well as activity-based protein profiling, are trying to meet the requirement of high-throughput target identification. PMID- 15288247 TI - Predictive ADMET studies, the challenges and the opportunities. AB - Predictive ADMET is the new 'hip' area in drug discovery. The aim is to use large databases of ADMET data associated with structures to build computational models that link structural changes with changes in response, from which compounds with improved properties can be designed and predicted. These databases also provide the means to enable predictions of human ADMET properties to be made from human in vitro and animal in vivo ADMET measurements. Both methods are limited by the amount of data available to build such predictive models, the limitations of modelling methods and our understanding of the systems we wish to model. The current failures, successes and opportunities are reviewed. PMID- 15288248 TI - Using NMR for ligand discovery and optimization. AB - Several recent technology-driven advances in the area of NMR have rekindled an interest in the application of the technology to problems in drug discovery and development. A unique aspect of NMR is that it has applicability in broadly different areas of the drug discovery and optimization processes. NMR techniques for screening aimed at the discovery of novel ligands or low molecular weight structures for fragment-based build up procedures are being applied commonly in the industry. Application of NMR in structure-guided drug design and metabonomics are also becoming routine. We present an overview of some of the most recent NMR developments in these areas. PMID- 15288249 TI - Flow cytometry for high-throughput, high-content screening. AB - Flow cytometry is a mature platform for quantitative multi-parameter measurement of cell fluorescence. Recent innovations allow up to 30-fold faster serial processing of bulk cell samples. Homogeneous discrimination of free and cell bound fluorescent probe eliminates wash steps to streamline sample processing. Compound screening throughput may be further enhanced by multiplexing of assays on color-coded bead or cell suspension arrays and by integrating computational techniques to create smaller, focused compound libraries. Novel bead-based assay systems allow studies of real-time interactions between solubilized receptors, ligands and molecular signaling components that recapitulate and extend measurements in intact cells. These new developments, and its broad usage, position flow cytometry as an attractive analysis platform for high-throughput, high-content biological testing and drug discovery. PMID- 15288250 TI - Making drugs on proteins: site-directed ligand discovery for fragment-based lead assembly. AB - Rapid progress in genomics and proteomics has provided a wealth of new targets for the pharmaceutical industry, even as many older targets still remain challenging for small-molecule drug discovery. Fragment-based lead discovery, in which leads are built progressively by expanding or combining small fragments, is a rapidly growing field that offers potential advantages over traditional lead discovery processes. However, identifying and assembling the fragments themselves can be challenging. Here, we review the concept of site-directed ligand discovery, in which a covalent bond is used to stabilize the interaction between a low-affinity fragment and a target protein. We also describe how this technique can facilitate fragment-based lead discovery and help overcome some of the limitations of traditional screening methods. PMID- 15288251 TI - Integrating cheminformatic analysis in combinatorial chemistry. AB - Cheminformatic analysis of drug-related compound databases has enabled the identification of the physicochemical properties that have the greatest influence on determining the drug-like characteristics of a compound. This enables definition of the parameters and profiles used in constructing a high-quality combinatorial library. Awareness of the multi-objective nature of combinatorial library construction has also given rise to techniques designed to enhance the likelihood of including the best compounds in a given library. PMID- 15288252 TI - Exploring the chemogenomic knowledge space with annotated chemical libraries. AB - The recent human genome initiatives have led to the discovery of a multitude of genes that are potentially associated with various pathologic conditions and, thus, have opened new horizons in drug discovery. Simultaneously, annotated chemical libraries have emerged as information-rich databases to integrate biological and chemical data. They can be useful for the discovery of new pharmaceutical leads, the validation of new biotargets and the determination of the structural basis of ligand selectivity within target families. Annotated libraries provide a strong information basis for computational design of target directed combinatorial libraries, which are a key component of modern drug discovery. Today, the rational design of chemical libraries enhanced with chemogenomics data is a new area of progressive research. PMID- 15288253 TI - High-throughput characterization and quality control of small-molecule combinatorial libraries. AB - To fully realize the potential of combinatorial synthesis and high-throughput screening for increasing the efficiency of the drug discovery and development process, issues related to compound purity must be addressed. Impurities, often present after synthesis, can lead to ambiguous screening results and inhibit the development of quality structure-activity relationships. The demand for high throughput analytical characterization of combinatorial libraries has prompted the development of more rapid methods to keep pace with compound production. Recent progress has focused upon the development of parallel separation methods, multiplexed detector interfaces, and synergistic combinations of different detectors possessing complementary selectivities. PMID- 15288254 TI - Inhibitors of angiogenesis and cancer-related receptor tyrosine kinases. AB - Inhibition of tumor angiogenesis is an attractive target in cancer therapy. In this context, receptor tyrosine kinases play a pivotal role. Extensive efforts have been made to identify and develop small-molecule inhibitors of these central signaling proteins. Some of these compounds have already passed or are currently in clinical trials to investigate their applicability as anti-cancer drugs. However, the high expectations that are set in antiangiogenic therapy have not yet been accomplished. But there are also new and exciting opportunities for cancer treatment by combining antiangiogenic molecules with newly emerging therapeutics. PMID- 15288255 TI - Emerging classes of protein-protein interaction inhibitors and new tools for their development. AB - Protein-protein interactions play a key role in the signal transduction pathways that regulate cellular function. Three years ago, few descriptions of small molecule protein-protein interaction inhibitors (SMPPIIs) existed in the literature. Today, the number of examples of both the biology and chemistry of such interaction inhibitors is growing rapidly. This growth occurs at the convergence of medicinal chemistry, signaling biology and novel assay technology for profiling emerging compound classes and modes of action. Protein translocation assays provide a unique new tool for identifying, profiling, and optimizing SMPPIIs. This review summarizes recent work in the field, and outlines future developments we can anticipate. PMID- 15288256 TI - Notch signaling and cancer: emerging complexity. PMID- 15288257 TI - Notch signaling in development and disease. AB - Cells in multicellular organisms need to decipher extracellular cues into appropriate responses including correct differentiation choices. A considerable portion of this information is relayed through a surprisingly small number of signaling pathways, which are highly evolutionarily conserved and used in many different cell types. This "ivy league" of signaling mechanisms comprises the Wnt/wingless, BMP/TGF-beta, Sonic Hedgehog, receptor tyrosine kinases, nuclear receptors, JAK/STAT and, the subject of this review, the Notch signaling pathway. The aim of this article is to provide an overview of the Notch signaling pathway. The role of Notch in various types of cancers is discussed in the accompanying articles in this issue of SCBI. PMID- 15288258 TI - Notch and T cell malignancy. AB - Notch signaling is required for normal T cell development. However, Notch expression must be precisely regulated as constitutive Notch signaling leads to T cell lymphomas. Recent evidence has provided insights into potential mechanisms of Notch-mediated lymphomagenesis and its relationship to T cell development. The evidence suggests that Notch likely interacts with several important cellular pathways and can cooperate with other oncogenes during lymphomagenesis. In particular, Notch appears to modulate pre-TCR signaling, inhibit the E2A pathway, and in murine leukemia models, frequently cooperates with Myc, E2A-PBX and dominant negative Ikaros dysregulation. This review will present current knowledge in these areas and explore theories on Notch-mediated T cell lymphomagenesis. PMID- 15288259 TI - Notch in mammary gland development and breast cancer. AB - Notch signaling has been implicated in many processes including cell fate determination and oncogenesis. In mice, the Notch1 and Notch4 genes are both targets for insertion and rearrangement by the mouse mammary tumor virus and these mutations promote epithelial mammary tumorigenesis. Moreover, expression of a constitutively active form of Notch4 in mammary epithelial cells inhibits epithelial differentiation and leads to tumor formation in this organ. These data implicate the Notch pathway in breast tumorigenesis and provide the foundation for future experiments that will aid in our understanding of the role of Notch in human breast cancer development. Here, we review studies of mammary tumorigenesis induced by Notch in mouse and in vitro culture models providing evidence that Notch activation is a causal factor in human breast cancer. PMID- 15288260 TI - Modulation of Notch signaling by mastermind-like (MAML) transcriptional co activators and their involvement in tumorigenesis. AB - Notch signaling is mediated by cell-cell interactions and is critical for cell fate determination in many species. Recently, a family of mastermind-like (MAML) transcriptional co-activator genes was identified that encode proteins that cooperate with Notch and CSL to activate transcription. Here, we review our current understanding of the roles of the MAML proteins in Notch signaling, and their involvement in certain human cancers. The mounting biochemical and functional evidence indicate that the MAML genes are critical components of the Notch signaling pathway, likely regulating cellular events involved in both normal development and oncogenesis. PMID- 15288261 TI - Notch in lung development and lung cancer. AB - Although data regarding the role of the Notch pathway in human lung cancer are still limited, fetal lung developmental studies suggest that Notch signaling plays a critical role in regulating airway epithelial development. The moderate hypotrophic phenotype of lungs from animals bearing a Hes1 mutation, and the expression of Notch components in the distal lung bud during branching morphogenesis, together suggest that Notch may play a role in normal lung growth, especially in Clara cell precursors. Non-small cell lung cancers, including adenocarcinoma, appear to actively utilize this conserved developmental pathway. Pharmacologic inhibition of the Notch pathway is a potential experimental approach to lung cancer treatment. PMID- 15288262 TI - Notch signaling in neuroblastoma. AB - Neuroblastoma is a pediatric tumor that originates from precursor cells of the sympathetic nervous system that have discontinued their normal differentiation program. This review is focused on involvement of the Notch signaling cascade in the process of differentiation in neuroblastoma cells and normal cells of the sympathetic nervous system. Hypoxia induces dedifferentiation of neuroblastoma cells in vivo and in vitro, and under oxygen-compromised conditions the Notch cascade is activated. This activation might promote development of the dedifferentiated phenotype. The implications of these observations for tumor biology are also discussed. PMID- 15288263 TI - Notch signaling in the integrated control of keratinocyte growth/differentiation and tumor suppression. AB - Oncogenesis is closely linked to abnormalities in cell differentiation. Notch signaling provides an important form of intercellular communication involved in cell fate determination, stem cell potential and differentiation. Here we review the role of this pathway in the integrated growth/differentiation control of the keratinocyte cell type, and the maintenance of normal skin homeostasis. In parallel with the pro-differentiation function of Notch1 in keratinocytes, we discuss recent evidence pointing to a tumor suppressor function of this gene in both mouse skin and human cervical carcinogenesis. The possibility that Notch signaling elicits signals with a duality of growth positive and negative function will be discussed. PMID- 15288264 TI - Viral interactions with the Notch pathway. AB - The Notch signaling pathway influences cell fate decisions, proliferation versus differentiation and cell survival. Viruses both utilize and manipulate the differentiation state of infected cells, promote or block cell cycling and employ a variety of mechanisms to evade innate cellular anti-viral responses and promote cell survival. In light of these commonalities, it is perhaps not surprising that several viruses have tapped into the Notch pathway to advance their own life cycles. This first became apparent from studies showing targeting of Epstein-Barr virus proteins to the nuclear effector of Notch signaling CSL (CBF1/RBPJk). More recently the Kaposi's sarcoma-associated herpesvirus RTA protein has been found to bind CSL. Notch pathway interactions have also been described for adenovirus SV40 and human papilloma virus. This review focuses on the herpesvirus protein interactions with the Notch pathway and the insights that these interactions have provided. PMID- 15288265 TI - Greenhouse gas emission during storage of pig manure on a pilot scale. AB - The greenhouse gas emissions (CO2, CH4, N2O) from a 2 ton (4.4 m3) deep litter pig manure pile (storage time 113 days during winter season) were quantified by using a tent, which covered the whole pile during the measuring periods only. The emissions were calculated in CO2 equivalents per kilogram dry matter by. Additionally the retention time (use of tracer gas SF6) and the concentrations of the gases in different parts of the pile were determined. The average retention time of the gases in the pile was less than 2 h. CH4 is assumed to have been generated only in the centre of the pile, whereas CO2 was assumed to have been generated in a wider zone. The emissions of CH4, CO2 and N2O were at the highest in the beginning when nearly the whole pile had temperatures in the range of thermophilic microorganisms. This leads to the assumption that mainly thermophilic microorganisms formed the gases. The most important gas for global warming was found to be nitrous oxide. PMID- 15288266 TI - Aerobic thermophilic treatment of farm slurry and food wastes. AB - The review discusses the aerobic treatments for farm slurry and food wastes and concentrates in particular on the thermophilic aerobic treatments. Methods are discussed under the heading of chemical, physical and other treatments. From those methods considered, the most suitable physical-microbiological treatment are aerobic thermophilic treatments. The main problem faced in aerobic thermophilic treatments could be the foaming formation during the process, and this could be solved by using different methods, mainly mechanical control method. Aerobic thermophilic treatments are also simple, economical and environmentally accepted. This method is known to have effects, and could be used to assist decontaminations on farms, as such technologies are already used in routine slurry treatment in many farms. PMID- 15288267 TI - Removal of nitrate-nitrogen from drinking water using bamboo powder charcoal. AB - The adsorption effectiveness of bamboo powder charcoal (BPC); made from the residual of Moso bamboo manufacturing; in removing nitrate-nitrogen from water has been investigated. Commercial activated carbon (CAC) was also used to compare the effectiveness of adsorption in removal of nitrate-nitrogen. The adsorption effectiveness of BPC was higher than that of CAC; regardless of the concentration of nitrate-nitrogen; in the range of 0-10 mg/l. The effect of temperature on adsorption by BPC and CAC in the range of 10-20 degrees C was also investigated. From the results, it was found that the temperature dependency of the adsorption effectiveness of BPC was weaker than that of CAC. This fact indicates that BPC can be an attractive option for the in situ treatment by adsorption of nitrate nitrogen-contaminated underground and surface water. PMID- 15288268 TI - Influence of salts and phenolic compounds on olive mill wastewater detoxification using superabsorbent polymers. AB - For a selection of nine commercially available superabsorbent polymers, the absorption capacity was evaluated for the principal absorption-inhibition constituent of OMW, mineral salts and for phytotoxic-components, the phenolic compounds. A double exponential model was established for electrical conductivities ranging 4.2-25,000 microS cm(-1). For solutions of phenolic compounds ranging 0-0.5 g l(-1), a distribution coefficient near unit was achieved, while for OMW, the phenolic compounds were concentrated inside the gel as the distribution coefficient was 1.4. Correction of OMW pH towards neutrality was found to increase the absorption capacity by up to 35%. The phytotoxicity was assessed by the germination of Lepidium sativum. Inhibition in plant growth occurred for all OMW dilutions without superabsorbent polymers application. For 5% of OMW (COD 5 gl(-1) and 200 ppm of phenolic compounds) immobilised in PNa2 (1 gl(-1)), plant growth was promoted being observed a 120% growth germination, thus indicating that olive mill wastewater detoxification occurred. PMID- 15288269 TI - Case study: design, operation, maintenance and water quality management of sustainable storm water ponds for roof runoff. AB - The purpose of this case study was to optimise design, operation and maintenance guidelines, and to assess the water treatment potential of a storm water pond system after 15 months of operation. The system was based on a combined silt trap, attenuation pond and vegetated infiltration basin. This combination was used as the basis for construction of a roof water runoff system from a single domestic property. United Kingdom Building Research Establishment and Construction Industry Research and Information Association, and German Association for Water, Wastewater and Waste design guidelines were tested. These design guidelines failed because they did not consider local conditions. The infiltration function for the infiltration basin was logarithmic. Algal control techniques were successfully applied, and treatment of rainwater runoff from roofs was found to be largely unnecessary for recycling (e.g., watering plants). However, seasonal and diurnal variations of biochemical oxygen demand, dissolved oxygen and pH were recorded. PMID- 15288270 TI - Biodegradation of diesel oil by an Arabian Sea sediment culture isolated from the vicinity of an oil field. AB - Laboratory scale batch studies were performed to test the diesel oil biodegradation ability of ES1 cultures isolated from Arabian Sea sediments obtained from the vicinity of an oil field. This culture could utilize diesel as the sole source of carbon and energy. Under aerobic conditions, 39% loss of diesel oil was observed over 8 days where 80% of the loss was due to aliphatic constituents. Under anoxic nitrate reducing conditions the rate and extent of degradation was significantly lower, i.e., 18% over 50 days. Salt acclimatized cultures could tolerate salinities up to 3.5% and demonstrated optimal performance at a salinity of 0.5%. The optimum N/P ratio for these cultures was found to be in the range of 2:1-5:1. Addition of two trace elemental substance formulations exhibited a significant inhibitory effect on culture growth. This culture has good potential for decontamination of oil-contaminated marine and subsurface environments. PMID- 15288271 TI - Single cell oil (SCO) production by Mortierella isabellina grown on high-sugar content media. AB - Mortierella isabellina cultivated in nitrogen-limited media presented remarkable cell growth (up to 35.9 g/l) and high glucose uptake even with high initial sugar concentrations (e.g. 100 g/l) in media. After nitrogen depletion, significant fat quantities were accumulated inside the fungal mycelia (50-55%, wt/wt oil in dry biomass), resulting in a notable single cell oil production of 18.1 g/l of culture medium. Total dry biomass and lipid yields presented greatly increased values (0.34 and 0.17 g respectively per gram of glucose consumed). The microbial lipid produced contained gamma-linolenic acid (GLA) at a concentration of 3.5+/ 1.0%, wt/wt, which corresponded to 16-19 mg GLA per gram of dry microbial mass and a maximum concentration of 0.801 g GLA per liter of culture medium. PMID- 15288272 TI - Production of high molecular weight pullulan by Aureobasidium pullulans HP-2001 with soybean pomace as a nitrogen source. AB - The production of pullulan by Aureobasidium pullulans HP-2001 was enhanced by yeast extract as a nitrogen source as well as soybean pomace. The highest production of pullulan by A. pullulans HP-2001 with yeast extract was 5.5 g/l whereas that of pullulan with soybean pomace was 7.5 g/l. The gas chromatogram of pullulan produced by A. pullulans HP-2001 with soybean pomace as a nitrogen source showed that the major and minor components were glucose and mannose. The FTIR spectra of pullulans produced with yeast extract, a mixture of yeast extract and soybean pomace, and soybean pomace alone exhibited similar features. The increase in content of reducing sugars after pullulanase treatment of pullulans produced with different nitrogen sources indicated that all the pullulans had alpha-(1,6) glucosidic linkages of alpha-(1,4) linked maltotriose units. The average molecular weights of pullulans produced with various concentrations of yeast extract and soybean pomace ranged from 0.17 to 1.32x10(6) and from 1.32 to 5.66x10(6), respectively. All pullulans produced by A. pullulans HP-2001 in this study had the same basic structures, but their ratios of monomeric components were a little different, which might result in the production of pullulans with different molecular weights. PMID- 15288273 TI - Analysis of the performance of an anaerobic digestion system at the Regina Wastewater Treatment Plant. AB - From the performance analysis of the anaerobic digestion system at the Regina Wastewater Treatment Plant, it was found that the anaerobic digestion system at the Regina plant was generally operated in a stable condition as indicated by pH, volatile acids and alkalinity levels. The operation of the anaerobic digestion system was not optimal because of the low volatile solids concentration and low volatile solids loading rate, especially because of high HRT. Two options, thickening the primary sludge and increasing the volatile solids loading rate, were recommended for the optimal operation of the digestion system. After examining a number of kinetic models, it was found that the Chen-Hashimoto model could be used to predict the volumetric methane production rate and the first order model could be used to predict the efficiency of volatile solids reduction. The study showed that utilization of digester gas for power production was the best alternative for the excess digester gas. 13.3% of the electrical demand and 35.5% of the plant's total energy could be met based on digester gas wasted, assuming 25% as the conversion efficiency. PMID- 15288274 TI - Characterization of the radical scavenging activity of lignins--natural antioxidants. AB - The present work is devoted to studies of the radical scavenging properties of lignins, which are recognized as efficient antioxidants of natural origin. Radical scavenging efficiency of a series of lignins isolated from deciduous and coniferous wood species and 10 lignin related monomeric compounds were examined against 1,1-diphenyl-2-picrylhydrazyl (DPPH*) radical in homogeneous conditions using ESR and spectrophotometry methods. Some structure-activity relationships are proposed, pointing out the importance of the non-etherified OH phenolic groups, ortho-methoxy groups, hydroxyl groups and the double bond between the outermost carbon atoms in the side chain for increasing scavenger activity. Analysis of rate constants for the lignins-DPPH* interaction revealed the contribution of polymer molecular weight and pi-polyconjugation systems. The pi conjugation systems of lignins operate as catalysts/activators of the interaction with DPPH*. Heterogeneity in terms of component composition (carbohydrate admixtures) and polydispersity is the factor which can decrease drastically the antioxidant efficiency of isolated lignins. The connection of the antibacterial effect of kraft lignin with radical scavenging activity of its soluble fraction was assumed. PMID- 15288275 TI - Performance characteristics of three aeration systems in the swine manure composting. AB - Pilot composting of swine manure mixed with rice straw was carried out to evaluate performance characteristics of three aeration systems: forced aeration, passive aeration and natural aeration. It was expected to provide academic basis for farmers to select an advisable aeration system. The results showed that the thermophilic durations were long enough to satisfy the sanitary standard, and swine manure could reach maturity. The indexes of the composting, including physical changes, pH value, TOC, OM, TKN, WSC, WSN, solid C/N ratio, water soluble C/N ratio, TOM, NH4+-N, (NO3(-) + NO2(-))-N, and GI had no significant difference among the treatments (P > 0.05) except the average temperature profiles (P12 = 0.001, P13 = 0.036). Economic analysis showed that a passive aeration system was suitable for a small-scale swine farm, and forced aeration system should be considered to apply in the middle and large-scale swine farms with a high extent of industrialization. But, in order to avoid too high temperature occurring during composting, an active aeration control system needed to be developed. PMID- 15288276 TI - Production of biodegradable plastics from activated sludge generated from a food processing industrial wastewater treatment plant. AB - Most of the excess sludge from a wastewater treatment plant (60%) is disposed by landfill. As a resource utilization of excess sludge, the production of biodegradable plastics using the sludge has been proposed. Storage polymers in bacterial cells can be extracted and used as biodegradable plastics. However, widespread applications have been limited by high production cost. In the present study, activated sludge bacteria in a conventional wastewater treatment system were induced, by controlling the carbon: nitrogen ratio to accumulate storage polymers. Polymer yield increased to a maximum 33% of biomass (w/w) when the C/N ratio was increased from 24 to 144, where as specific growth yield decreased with increasing C/N ratio. The conditions which are required for the maximum polymer accumulation were optimized and are discussed. PMID- 15288277 TI - Combustion behaviors of a compression-ignition engine fueled with diesel/methanol blends under various fuel delivery advance angles. AB - A stabilized diesel/methanol blend was described and the basic combustion behaviors based on the cylinder pressure analysis was conducted in a compression ignition engine. The study showed that increasing methanol mass fraction of the diesel/methanol blends would increase the heat release rate in the premixed burning phase and shorten the combustion duration of the diffusive burning phase. The ignition delay increased with the advancing of the fuel delivery advance angle for both the diesel fuel and the diesel/methanol blends. For a specific fuel delivery advance angle, the ignition delay increased with the increase of the methanol mass fraction (oxygen mass fraction) in the fuel blends and the behaviors were more obvious at low engine load and/or high engine speed. The rapid burn duration and the total combustion duration increased with the advancing of the fuel delivery advance angle. The centre of the heat release curve was close to the top-dead-centre with the advancing of the fuel delivery advance angle. Maximum cylinder gas pressure increased with the advancing of the fuel delivery advance angle, and the maximum cylinder gas pressure of the diesel/methanol blends gave a higher value than that of the diesel fuel. The maximum mean gas temperature remained almost unchanged or had a slight increase with the advancing of the fuel delivery advance angle, and it only slightly increased for the diesel/methanol blends compared to that of the diesel fuel. The maximum rate of pressure rise and the maximum rate of heat release increased with the advancing of the fuel delivery advance angle of the diesel/methanol blends and the value was highest for the diesel/methanol blends. PMID- 15288278 TI - Acetylation of sugarcane bagasse using NBS as a catalyst under mild reaction conditions for the production of oil sorption-active materials. AB - Sugarcane bagasse was esterified with acetic anhydride using N-bromosuccinimide as a catalyst under mild conditions in a solvent free system. The extent of acetylation was measured by weight percent gain, which varied from 2.1% to 24.7% by changing the reaction temperature (25-130 degrees C) and duration (0.5-6.0 h). N-Bromosuccinimide was found to be a novel and highly effective catalyst for acetylation of hydroxyl groups in bagasse. At a concentration of 1% of the catalyst in acetic anhydride, a weight percent gain of 24.7% was achieved at 120 degrees C for 1 h, compared with 5.1% for the un-catalyst reaction under the same reaction condition. FT-IR and CP-MAS 13C-NMR studies produced evidence for acetylation. The thermal stability of the products decreased slightly upon chemical modification, but no significant decrease in thermal stability was observed for WPG > or = 24.7%. More importantly, the acetylation significantly increased hydrophobic properties of the bagasse. The oil sorption capacity of the acetylated bagasse obtained at 80 degrees C for 6 h, was 1.9 times higher than the commercial synthetic oil sorbents such as polypropylene fibres. Therefore, these oil sorption-active materials can be used to substitute non-biodegradable materials in oil spill cleanup. PMID- 15288279 TI - Old and new markers for breast cancer prognosis: the need for integrated research on quantitative issues. PMID- 15288280 TI - Evaluation of epidermal growth factor receptor tyrosine kinase inhibitors combined with chemotherapy: Is there a need for a more rational design? PMID- 15288281 TI - The Hamburg statement: the partnership driving the European agenda on breast cancer. PMID- 15288282 TI - Isolated hepatic perfusion for the treatment of colorectal metastases confined to the liver: recent trends and perspectives. AB - Isolated hepatic perfusion (IHP) involves a method of complete vascular isolation of the liver to allow treatment of liver tumours with toxic systemic doses. The recent clinical studies mainly employed IHP with melphalan with or without tumour necrosis factor-alpha (TNF-alpha) and mild hyperthermia. The results of these studies show that high response rates and high survival rates can be achieved by IHP. In this article, the current status, recent developments and future perspectives of IHP are discussed. PMID- 15288283 TI - Re-evaluating the role of dacarbazine in metastatic melanoma: what have we learned in 30 years? AB - Since dacarbazine was approved for treating metastatic melanoma in the 1970s, numerous studies have evaluated whether different schedules and dacarbazine-based combinations improve clinical outcomes. This evidence-based review shows that combining dacarbazine with other drugs having single-agent activity and/or hormonal or immunotherapeutic compounds fails to provide clinically meaningful improvements in survival, and may increase toxicity. In patients with metastatic melanoma, dacarbazine was previously administered in cycles of multiple consecutive daily infusions per cycle. The introduction of potent antiemetics, together with concerns relating to patient comfort and clinic utilisation time, has enabled regimens involving single-dose dacarbazine, administered at the same total dose per cycle. These appear to be as effective as multiple-dose schedules, are well tolerated, and are more straightforward to administer. Single administration dacarbazine (850-1000 mg/m2), once every 3 weeks, is currently the standard reference therapy in patients with advanced melanoma. New effective therapies are urgently needed for this treatment-refractory disease. PMID- 15288284 TI - "Good Old" clinical markers have similar power in breast cancer prognosis as microarray gene expression profilers. AB - We compared the power of gene expression measurements with that of conventional prognostic markers, i.e., clinical, histopathological, and cell biological parameters, for predicting distant metastases in breast cancer patients using both established prognostic indices (e.g., the Nottingham Prognostic Index (NPI)) and novel combinations of conventional markers. We used publicly available data on 97 patients, and the performance of metastasis prediction was represented by receiver operating characteristic (ROC) areas and Kaplan-Meier plots. The gene expression profiler did not perform noticeably better than indices constructed from the clinical variables, e.g., the well established NPI. When analysing separately subgroups, according to the oestrogen receptor (ER) status both approaches could predict clinical outcome more easily for the ER-positive than for the ER-negative cohort. Given the time it may take before microarray processing is used worldwide, particularly due to the costs and the lack of standards, it is important to pursue research using conventional markers. Our analysis suggests that it might be possible to improve the combination of different conventional prognostic markers into one prognostic index. PMID- 15288285 TI - Impact of surgical staging in patients with macroscopic "stage I" ovarian borderline tumours: analysis of a continuous series of 101 cases. AB - The aim of this study was to assess the patient's clinical outcome following complete or incomplete surgical staging in cases treated for an early stage low malignant-potential ovarian tumour (LMPOT). One-hundred and one patients treated between 1965 and 1998 for a early stage I LMPOT were reviewed according to whether the initial surgical staging was complete (Group 1/defined by peritoneal cytology + peritoneal biopsies + infracolic omentectomy) or incomplete (Group 2/omission of at least one of the peritoneal staging procedures described above). Complete and incomplete surgical stagings were carried out in 48 (48%) and 53 (52%) patients, respectively. Four (8%) LMPOT recurrences were observed in Group 2, all following conservative management, but there were no recurrences in Group 1. No relapses with invasive carcinoma or peritoneal disease and no tumour related deaths were observed. The absence of complete peritoneal staging in patients with an apparent "stage I" LMPOT increased the recurrence rate. However, this surgical restaging (in cases of incomplete initial surgery) does not modify the survival of patients with apparent "stage I" LMPOT misdiagnosed during the initial surgery. This procedure could probably be omitted: (1) if the peritoneum is clearly reported as "normal" during the initial surgery; (2) in the absence of a micropapillary pattern; and (3) if the patient agrees to be carefully followed up. PMID- 15288286 TI - Information given to cancer patients on diagnosis, prognosis and treatment: the clinical oncologist's perspective. AB - The extent of information to cancer patients is, in general, culture-dependent. Information mainly refers to three aspects, namely diagnosis (Dx), prognosis (Px) and treatment (Rx), but the relative contribution of each domain to the information given overall is not available. To address this issue, we e-mailed a questionnaire to 9893 members of the American Society of Clinical Oncology (ASCO) asking whether they agree that information about Dx, Px and Rx contribute differently to the information given to the cancer patient overall and, if so, to what extent, both in the adjuvant and advanced settings. 857 questionnaires were evaluable. There was no statistically significant difference between the contribution of these 3 domains in the adjuvant setting (33%, 34% and 33%, respectively). In subgroup analysis, medical oncologists and haematologists attributed a significantly higher contribution of Px information compared with other specialists (P < 0.05). In the advanced setting, respondents estimated a higher contribution of Px (41%) to patient information overall compared with Dx and Rx (28% and 31%, respectively; P < 0.05). This finding was more pronounced in North America than in Europe (P < 0.0001), and in Germanic-language than in Romance-language countries (P = 0.005). In conclusion, information on Dx, Px and Rx are believed to contribute differently to the information delivered to cancer patients overall, depending on the stage of disease, the cultural environment and the specialty of the physician. PMID- 15288287 TI - LH-RH agonists offer very good protection against the adverse gynaecological effects induced by tamoxifen. AB - This study was initiated to evaluate the efficacy of luteinizing hormone releasing hormone (LH-RH) agonists in protecting premenopausal patients against the adverse gynaecological effects induced by tamoxifen. Between January 1998 and January 2000, 85 premenopausal breast cancer patients were included in this prospective study. All were to receive LH-RH agonists and tamoxifen for a minimum of two years. All patients underwent a pretreatment gynaecological evaluation and annual follow-up. Bone density was also measured at the start of treatment and then after 2, 3 and 4 years. Pretreatment evaluation revealed 2 polyps. At one and two years of follow-up, no abnormal symptoms were noted and echographic findings were normal. At three years of follow-up, a polyp associated with adnexal masses was discovered. Histology revealed ovarian and endometrial metastases of infiltrating lobular breast carcinoma. Bone density evaluation after 2, 3 and 4 years of treatment showed no significant bone loss. LH-RH agonists offer safe protection against the gynaecological side-effects of tamoxifen in premenopausal breast cancer patients. PMID- 15288288 TI - Quality of life after palliative treatment for oesophageal carcinoma -- a prospective comparison between stent placement and single dose brachytherapy. AB - Metal stent placement and single dose brachytherapy are commonly used treatment modalities for the palliation of inoperable oesophageal carcinoma. We investigated generic and disease-specific health-related quality of life (HRQoL) after these palliative treatments. Patients with dysphagia from inoperable oesophageal carcinoma were randomised to placement of a covered Ultraflex stent (n = 108) or single dose (12 Gy) brachytherapy (n = 101). We obtained longitudinal data on disease-specific (dysphagia score, European Organisation for Research and Treatment of Cancer (EORTC) OES-23, visual analogue pain scale) and generic (EORTC Quality of Life-Core 30 Questionnaire (QLQ-C30), Euroqol (EQ)-5D) HRQoL at monthly home visits by a specially-trained research nurse. We compared HRQoL between the two treatments and analysed changes in HRQoL during follow-up. Dysphagia improved more rapidly after stent placement than after brachytherapy, but long-term relief of dysphagia was better after brachytherapy. For generic HRQoL, there was an overall significant difference in favour of brachytherapy on four out of five functional scales of the EORTC QLQ-C30 (role, emotional, cognitive and social) (P < 0.05). Generic HRQoL deteriorated over time on all functional scales of the EORTC QLQ C-30 and EQ-5D, in particular physical and role functioning (on average -23 and -24 on a 100 points scale during 0.5 years of follow-up). This decline was more pronounced in the stent group. Major improvements were seen on the dysphagia and eating scales of the EORTC OES-23, in contrast to other scales of this disease-specific measure, which remained almost stable during follow-up. Reported levels of chest or abdominal pain remained stable during follow-up in both treatment groups, general pain levels increased to a minor extent. The effects of single dose brachytherapy on HRQoL compared favourably to those of stent placement for the palliation of oesophageal cancer. Future studies on palliative care for oesophageal cancer should at least include generic HRQoL scales, since these were more responsive in measuring patients' functioning and well-being during follow-up than disease-specific HRQoL scales. PMID- 15288289 TI - Phase II studies of BBR3464, a novel tri-nuclear platinum complex, in patients with gastric or gastro-oesophageal adenocarcinoma. AB - BBR3464, a novel tri-nuclear platinum complex, forms long-range DNA adducts and is highly potent when compared with cisplatin in vitro. Preclinical studies demonstrated activity in cisplatin-resistant tumours and tumours with mutated p53 status. Phase I & II clinical studies gave preliminary indications of activity in melanoma, pancreatic, lung and ovarian cancers. The aim of this study was to determine the efficacy and confirm the toxicity of BBR3464 when given either as first- or second-line treatment for advanced disease in patients with gastric and gastro-oesphageal adenocarcinoma. Two multicentre, open label, Gehan design studies were conducted; one study used BBR3464 as first-line and the other as second-line treatment for metastatic or locally advanced disease. Nineteen first line and 26 second-line patients were enrolled receiving a total of 74 and 53 infusions, respectively. Initially, seven patients in the second-line study received BBR3464 using the planned schedule of 1.1 mg/m2 every 4 weeks; however, 5 of these patients experienced dose-limiting grade 3 or 4 febrile neutropenia; subsequent patients in both studies were treated using the modified schedule of 0.9 mg/m2, every 21 days. In 1 of 17 evaluable, previously untreated patients, regression of multiple skin lesions was noted with stabilisation of lung metastases and maxillary sinus mass, lasting 155 days. In the first-line study, the median time to progression was 85 days [95% Confidence Interval (CI): 42, 127] (2.8 months) and in the second-line study, the median time to progression was 71 days [95% CI: 42, 109] and 38 days [95% CI: 32, 73] in the 1.1 and 0.9 mg/m2 dose level groups, respectively. Toxicity data were available for 45 patients. Neutropenia was the main toxicity seen (G3: 40%, G4: 40%). Febrile neutropenia was observed in six patients (15%) treated with 0.9 mg/m2 compared with five patients (71%) treated with 1.1 mg/m2 BBR3464. Other drug-related toxicities (G3/4) included: anaemia, thrombocytopenia, nausea, vomiting, diarrhoea, mucositis and fatigue. Diarrhoea and nausea/ vomiting were adequately controlled by the use of loperamide and antiemetics, respectively. Recruitment to the second-line study was closed early due to the poor response rate (1/17 evaluable, 6%; 95% CI: 1%, 27%) and short time to progression noted in the first line study. Further studies with BBR3464 in this tumour type are not recommended. PMID- 15288290 TI - Late events occurring five years or more after successful therapy for childhood rhabdomyosarcoma: a report from the Soft Tissue Sarcoma Committee of the Children's Oncology Group. AB - The aim of our study was to describe late failures in children who initially survived event-free five years from a diagnosis of rhabdomyosarcoma. Charts of children enrolled in the Intergroup Rhabdomyosarcoma Study Group (IRSG) trials III, IV pilot and IV (1984-1997) who survived five years event-free and subsequently experienced an adverse event (disease recurrence, second malignant neoplasm or death from other causes) were reviewed. Of the 2534 enrolled patients, 1160 were event-free at five years and 48 subsequently experienced a late event. The estimated 10-year event rate for the 1160 patients who were alive and event-free at five years was 9% (95% Confidence Interval (CI) 5%, 13%). Patients with both advanced disease (Group III/IV) and large primary tumours at diagnosis (> 5 cm) were at the highest risk for late events (19%; 95% CI 8%, 30%). Late events after successful treatment for rhabdomyosarcoma occur in 9%. Those with advanced disease and large primary tumours have the highest risk of late events. PMID- 15288291 TI - European collaboration in trials of new agents for children with cancer. AB - Childhood cancer is a relatively rare disease, representing just 1% of all malignancies. Within Europe, this represents some 12,000 new cases each year, with approximately 1600 a year in the United Kingdom and 1800 in France. International collaboration in phase III trials of childhood cancer has been the norm for many years, traditionally within Europe, but, largely because of organisational considerations, phase I and II trials have only been conducted on a national basis. With overall cure rates in the region of 70%, relatively few children are available for these early drug trials. Access to new drugs is also a major problem. Against this background, a United Kingdom (UK)/French 'new agent' collaboration was established, expanding subsequently into a wider European grouping. This paper documents the history of that collaboration, the outcomes and future challenges. PMID- 15288292 TI - Commentary on "European collaboration in trials of new agents for children with cancer" by Ablett et al. AB - Recent progress in establishing a European network to conduct paediatric oncology phase I/II clinical trials calls attention to the challenges facing researchers developing new agents for children with cancer. These challenges include: ensuring that effective infrastructures are in place to safely and efficiently conduct early phase clinical trials in children while meeting all ethical and regulatory requirements associated with such trials; obtaining timely access to new agents from pharmaceutical sponsors for both preclinical testing and for phase I and phase II testing; and effectively prioritizing new agents for evaluation in children so that those agents most likely to benefit children with specific cancers are brought forward for clinical testing. The use of public funds to develop and maintain clinical trials infrastructures devoted to paediatric oncology drug development can help in addressing these challenges and can facilitate the timely paediatric evaluation of new agents, thereby contributing to the goal of identifying more effective treatments for children with cancer. PMID- 15288293 TI - E-Cadherin (CDH1) and p53 rather than SMAD4 and Caspase-10 germline mutations contribute to genetic predisposition in Portuguese gastric cancer patients. AB - Approximately 30% of all hereditary diffuse gastric cancer (HDGC) families carry CDH1 germline mutations. The other two thirds remain genetically unexplained and are probably caused by alterations in other genes. Using polymerase chain reaction (PCR)/single-strand conformation polymorphism (SSCP)/sequencing, we screened 32 Portuguese families with a history of gastric cancer and 23 patients with early onset gastric cancer for CDH1 germline mutations. In probands negative for CDH1 mutations, we screened genes involved in hereditary cancer syndromes in which gastric cancer may be one of the component tumours, namely p53 (Li-Fraumeni Syndrome) and hMLH1 and hMSH2 (HNPCC). We also screened in these patients for mutations in Caspase-10, a gene inactivated in sporadic gastric cancer, and SMAD4, a gene whose inactivation in mice is associated with signet-ring cell carcinoma of the stomach. One of the families fulfilling the HDGC criteria harboured a CDH1 germline mutation, and one of the families with incomplete criteria harboured a p53 germline mutation. No mutations were identified in hMLH1 and hMSH2, and only sequence variants were found in SMAD4 and Caspase-10. The present work reports for the first time CDH1 germline mutations in Portuguese gastric cancer families, and highlights the need for p53 mutation screening in families lacking CDH1 germline mutations, in a country with one of the highest incidences of gastric cancer in the world. No evidence was found for a role of germline mutations in SMAD4 and Caspase-10 in families lacking CDH1 mutations. PMID- 15288294 TI - Expression of hypoxia-inducible factor (HIF)-1alpha is associated with vascular endothelial growth factor expression and tumour angiogenesis in human oesophageal squamous cell carcinoma. AB - The purpose of this study was to examine the relationship between hypoxia inducible factor (HIF)-1alpha expression, vascular endothelial growth factor (VEGF) expression, and tumour vascularity in squamous cell carcinoma of the oesophagus. Expression of HIF-1alpha and VEGF was examined in two oesophageal squamous cell carcinoma cell lines (TE2, TE3) and 82 archival surgical specimens of human oesophageal squamous cell carcinoma tissue. In both cell lines, the levels of HIF-1alpha protein and VEGF mRNA were increased under hypoxic conditions. Thirty-two of the 82 (39%) tumour specimens showed high levels of HIF 1alpha immunoreactivity in the nuclei and/or cytoplasm of cancer cells. HIF 1alpha expression correlated significantly with venous invasion, VEGF expression, and microvessel density. Among the 47 patients who did not receive pre-operative chemotherapy, the outcome of those with high HIF-1alpha-expressing tumours was significantly poorer than that of those with low HIF-1alpha-expressing tumours. These results suggest that HIF-1alpha and VEGF expression are important determinants of survival in squamous cell carcinoma of the oesophagus. PMID- 15288295 TI - An open randomised trial of second-line endocrine therapy in advanced breast cancer: comparison of the Aromatase inhibitors letrozole and anastrozole. PMID- 15288296 TI - Long-term behavior of passively aerated compost methanotrophic biofilter columns. AB - The methane oxidation potential of several types of compost methanotrophic biofilter columns were compared in the laboratory over a period of 220 days. The results indicate an increase in methanotrophic activity over a period of about 100 days, up to a maximum of 400 g m(-2) day(-1), and a gradual decline to about 100 g m(-2) day(-1) within the next 120 days. High methane oxidation rates appear to be restricted to a small area of the column, 10-15 cm thick. Based on the laboratory investigations carried out to determine the cause for the decline in methane oxidation rate, it was concluded that the formation of exopolymeric substances (EPS), at the zones of maximum methane oxidation, was responsible for this decline. In monitoring methane oxidation in a column for up to 600 days, it was observed that mixing of the medium after formation of EPS enabled the column to temporarily recover high performance. The results suggest that stable, homogenous compost, with a low C/N and low ammonium content, mixed on a regular basis, could achieve and maintain high methane oxidation efficiencies. PMID- 15288297 TI - Decentralised composting of urban waste--an overview of community and private initiatives in Indian cities. AB - The national waste legislation, introduced in India in 2000, endorses the principle of "Recycle Before Disposal" and clearly stipulates composting as an option for organic waste treatment. It also recommends waste separation as prerequisite for treatment. Although various composting schemes of different scale, type and organisational structure currently exist in the country, a general overview is lacking and very little independent site-specific information is available. This paper presents the results of a study assessing 17 decentralised systems from the cities of Bangalore, Chennai, Pune, and Mumbai. The schemes were classified according to their organisational setup into: (1) citizens' and community initiatives; (2) business and institution initiatives operating on their premises; and (3) small and medium-size private sector initiatives. These categories also coincide with different operational scales. Community initiatives have developed from unreliable collection services, and composting emerged mainly as a spin-off activity from the collection system to reduce waste delivery to the communal containers emptied by the municipal services. The potential to launch and sustain decentralised composting schemes is dependent on the municipal provision of adequate space. This paper summarises further key issues pertaining to the assessed schemes and reveals overall deficiencies in the field of accounting and transparency, composting technique and marketing, as well as municipal authority involvement. PMID- 15288298 TI - Cryo-comminution of plastic waste. AB - Recycling of plastics is a big issue in terms of environmental sustainability and of waste management. The development of proper technologies for plastic recycling is recognised as a priority. To achieve this aim, the technologies applied in mineral processing can be adapted to recycling systems. In particular, the improvement of comminution technologies is one of the main actions to improve the quality of recycled plastics. The aim of this work is to point out suitable comminution processes for different types of plastic waste. Laboratory comminution tests have been carried out under different conditions of temperature and sample pre-conditioning adopting as refrigerant agents CO2 and liquid nitrogen. The temperature has been monitored by thermocouples placed in the milling chamber. Also different internal mill screens have been adopted. A proper procedure has been set up in order to obtain a selective comminution and a size reduction suitable for further separation treatment. Tests have been performed on plastics coming from medical plastic waste and from a plant for spent lead batteries recycling. Results coming from different mill devices have been compared taking into consideration different indexes for representative size distributions. The results of the performed tests show as cryo-comminution improves the effectiveness of size reduction of plastics, promotes liberation of constituents and increases specific surface size of comminuted particles in comparison to a comminution process carried out at room temperature. PMID- 15288299 TI - Biodegradable organic matter in municipal solid waste incineration bottom ash. AB - For investigation of the behavior of municipal solid waste incineration bottom ash in landfill, we have analysed bottom ash samples taken after the quench tank as well as after five months of storage in the laboratory for elements and organic constituents. Water extractable organic carbon, particulate organic carbon, amino acids, hexosamines and carbohydrates considerably decreased during the five months of storage and their spectra revealed microbial reworking. This shows that the organic matter present in the bottom ash after incineration can provide a substrate for microbial activity. The resulting changes of the physico chemical environment may effect the short-term behavior of the bottom ash in landfill. PMID- 15288300 TI - Parametric sensitivity analysis of leachate transport simulations at landfills. AB - This paper presents a case study in simulating leachate generation and transport at a 2000 ton/day landfill facility and assesses leachate migration away from the landfill in order to control associated environmental impacts, particularly on groundwater wells down gradient of the site. The site offers unique characteristics in that it is a former quarry converted to a landfill and is planned to have refuse depths that could reach 100 m, making it one of the deepest in the world. Leachate quantity and potential percolation into the subsurface are estimated using the Hydrologic Evaluation of Landfill Performance (HELP) model. A three-dimensional subsurface model (PORFLOW) was adopted to simulate ground water flow and contaminant transport away from the site. A comprehensive sensitivity analysis to leachate transport control parameters was also conducted. Sensitivity analysis suggests that changes in partition coefficient, source strength, aquifer hydraulic conductivity, and dispersivity have the most significant impact on model output indicating that these parameters should be carefully selected when similar modeling studies are performed. PMID- 15288301 TI - Conventional and fast pyrolysis of automobile shredder residues (ASR). AB - This work aims at comparing performance and product yields in conventional pyrolysis and fast pyrolysis of automotive shredded residues. In both processes, carbon conversion to gaseous and liquid products was more than 80%. Gas production was maximised in conventional pyrolysis (about 35% by weight of the initial ASR weight), while fast pyrolysis led to an oil yield higher than 55%. Higher heating values (HHV) of both conventional pyrolysis gas and fast pyrolysis oil increased from 8.8 to 25.07 MJ/Nm3 and from 28.8 and 36.27 MJ/kg with increasing pyrolysis temperature. PMID- 15288302 TI - A study of disposed fly ash from landfill to replace Portland cement. AB - The landfills of fly ash are the problem of all power plants because this disposed fly ash is not used in any work. This research studies the potential of using disposed fly ashes which have disposal time of 6-24 months from the landfill of Mae Moh power plants in Thailand to replace Portland cement type I. Median particle sizes of disposed fly ashes between 55.4 and 99.3 microm were ground to reduce the sizes to about 7.1-8.4 microm. Both original and ground disposed fly ashes were investigated on physical and chemical properties. Compressive strengths of disposed fly ash mortars were determined when Portland cement type I was replaced by disposed fly ashes at the rate of 10%, 20%, and 30% by weight of cementitious material (Portland cement type I and disposed fly ash). The results presented that most particles of original disposed fly ashes were solid and sphere with some irregular shape while those of ground disposed fly ashes were solid and irregular shape. CaO and LOI contents of disposed fly ashes with different disposal times had high variation. The compressive strengths of original disposed fly ash mortars were low but those of ground disposed fly ash mortars at the age of 7 days were higher than 75% of the standard mortar and increased to be higher than 100% after 60 days. From the results, it could be concluded that ground disposed fly ashes were excellent pozzolanic materials and could be used as a partial replacement of cement in concrete, even though they were exposed to the weather for 24 months. PMID- 15288303 TI - The investigation of a class of capacitated arc routing problems: the collection of garbage in developing countries. AB - The collection, transport and disposal of solid waste, which is a highly visible and important municipal service, involves a large expenditure but receives, scant attention. This problem is even more crucial for large cities in developing countries due to the hot weather. A constructive heuristic which takes into account the environmental aspect as well as the cost is proposed to solve the routing aspect of garbage collection. This is based on a look-ahead strategy which is enhanced by two additional mechanisms. Interesting results were obtained when tested on instances with and without the presence of the effect of the environment. PMID- 15288304 TI - Influence of landfill leachate suspended solids on clog (biorock) formation. AB - Laboratory column tests were performed to evaluate the role of leachate-suspended solids in clogging a granular material permeated with Keele Valley Landfill leachate. The development of the clog material was a result of biological, chemical, and physical processes occurring within the column. The increase in volatile solids, which contributed to clog development over time, was primarily due to the retention of volatile suspended solids and growth of a biofilm capable of removing acetate, propionate, and butyrate from the leachate. Acetate fermentation was primarily responsible for precipitation of calcium within the column. The precipitated calcium and retention of inorganic suspended solids contributed to the increase in clog inorganic solids. Over the duration of the experiment, 3.7 times more calcium was precipitated in the column (due to acid fermentation) than was retained with inorganic suspended solids. Clogging resulted in a greater than 60% reduction in drainable porosity and a six-order magnitude decrease in hydraulic conductivity. The potential practical implications with respect to pipe cleaning and leachate recirculation were discussed. PMID- 15288305 TI - Assessment of performance of solid waste management contractors: a simple techno social model and its application. AB - Despite being in operation for more than 7 years now, privatised solid waste management (SWM) service in Tanzania has not been formally assessed. This is the premise for the techno-social performance assessment model for SWM contractors, which is presented in this paper. This paper outlines the background of SWM privatisation in Dar es Salaam City in addition to describing and demonstrating the application of the assessment model. The model has three components: service beneficiaries' assessment sub-model, service area technical assessment sub-model, and contractors' attributes sub-model. The model can be used to assess the performance of SWM contractors and in so doing provide data for assessing SWM privatisation as a whole. In the application of the model presented in this paper scores for 5 assessed contractors are in the range 24-65%. The contractor who scored highest is also the most experienced and also the best equipped. Nevertheless, all contractors scored poorly with the respect to their ability and attributes in SWM mainly because most of them are new to the solid waste management field. PMID- 15288306 TI - A review of clinical failures associated with macrolide-resistant Streptococcus pneumoniae. AB - The emerging reports of clinical failures using macrolides and their associations with macrolide-resistant Streptococcus pneumoniae prompted us to review the literature describing these cases. Thirty-three cases reporting macrolide treatment failure during treatment of pneumococcal infections were available for review. The most prevalent diagnosis (24/27 or 88.8% of available diagnoses) was community-acquired pneumonia (CAP). Previous medical history included cardiopulmonary disease in eight (24.2%) and immunocompromised states in five (15.1%) patients. The majority, 31/33 (93.9%) of patients received oral macrolide treatment in an outpatient setting. S. pneumoniae was isolated from the blood in 26 (78.8%) of 33 patients, three (9.1%) patients had bacteria present in both blood and cerebrospinal fluid, two (6%) patients grew S. pneumoniae from blood and bronchial washings and two (6%) patients had positive sputum cultures. The MLS(B) phenotype was the most predominant phenotype present in 12 (63.2%) of 19 patients. After failing initial macrolide treatment, 26 (78.8%) of 33 patients received parenteral antibiotic treatment. Of 33 patients admitted to hospital, 29 (87.8%) had their outcome described as 'survived'. PMID- 15288307 TI - A view on antimicrobial resistance in developing countries and responsible risk factors. AB - Antimicrobial resistance is one of the biggest challenges facing global public health. Although antimicrobial drugs have saved many lives and eased the suffering of many millions, poverty, ignorance, poor sanitation, hunger and malnutrition, inadequate access to drugs, poor and inadequate health care systems, civil conflicts and bad governance in developing countries have tremendously limited the benefits of these drugs in controlling infectious diseases. The development of resistance in the responsible pathogens has worsened the situation often with very little resource to investigate and provide reliable susceptibility data on which rational treatments can be based as well as means to optimise the use of antimicrobial agents. The emergence of multi-drug-resistant isolates in tuberculosis, acute respiratory infections and diarrhoea, often referred to as diseases of poverty, has had its greatest toll in developing countries. The epidemic of HIV/AIDS, with over 30 million cases in developing countries, has greatly enlarged the population of immunocompromised patients. The disease has left these patients at great risk of numerous infections and even greater risks of acquiring highly resistant organisms during long periods of hospitalisation. This review discusses antimicrobial resistance in developing countries and the risk factors responsible. PMID- 15288308 TI - Assessment of pathogen occurrences and resistance profiles among infected patients in the intensive care unit: report from the SENTRY Antimicrobial Surveillance Program (North America, 2001). AB - Originating from 25 selected intensive care units (ICUs) in North America, a total of 1,321 bacterial strains from blood, respiratory tract, urine and wound sites were processed at a central laboratory as part of the SENTRY Antimicrobial Surveillance Program (2001) to assess their occurrence rates and antimicrobial susceptibility profiles. The rank order of pathogens recovered was Staphylococcus aureus (24.1%), Pseudomonas aeruginosa (12.2%), Escherichia coli (10.1%), Klebsiella spp. (8.9%), Enterococcus spp. (7.2%), coagulase-negative staphylococci (7.0%) and Enterobacter spp. (7.0%). Although oxacillin resistance among S. aureus was 51.4%, no resistance was detected to vancomycin, linezolid and quinupristin/dalfopristin. The most active agents tested against P. aeruginosa were amikacin, cefepime, tobramycin, meropenem and piperacillin/tazobactam (3.1-13.0% resistance). Among agents tested against the Enterobacteriaceae, amikacin, cefepime, imipenem and meropenem showed greatest in vitro activity (0.0-3.4% resistance). Extended-spectrum beta-lactamase-producing phenotype rates were 11.2 and 16.2% in E. coli and Klebsiella spp., respectively. Linezolid was most active against enterococci (1.1% resistance; G2576U ribosomal mutation) whereas 28.4% of isolates were resistant to vancomycin. Cefepime and the carbapenems (imipenem or meropenem) for Gram-negative isolates and linezolid for Gram-positive isolates, provided the broadest spectrum of in vitro activity against contemporary ICU pathogens in North America. PMID- 15288309 TI - Determining incidence of extended spectrum beta-lactamase producing Enterobacteriaceae, vancomycin-resistant Enterococcus faecium and methicillin resistant Staphylococcus aureus in 38 centres from 17 countries: the PEARLS study 2001-2002. AB - The PEARLS study prospectively monitored selected nosocomial pathogens from 38 centres in 13 European, three Middle Eastern countries and South Africa during 2001-2002. Extended spectrum beta-lactamase (ESBL) production rates among Escherichia coli, Klebsiella pneumoniae, and Enterobacter spp. were 5.4% (142/2609), 18.2% (401/2,206) and 8.8% (204/2,328), respectively, for all study sites. The overall ESBL production rate for the combined Enterobacteriaceae was 10.5% (747/7,143), highest in Egypt, 38.5%, and Greece, 27.4%, and lowest in The Netherlands, 2.0%, and Germany, 2.6%. IEF, PCR and DNA sequencing determined 10.7% false positives among Enterobacter spp. when using NCCLS guidelines to screen for ESBL production. The prevalence of nosocomial methicillin-resistant Staphylococcus aureus and vancomycin-resistant Enterococcus faecium was 32.4% (294/908) and 8.7% (83/949), respectively. PEARLS provides baseline data against which prospective changes in resistant determinants and outcomes can be measured in this ongoing study. PMID- 15288310 TI - Antimicrobial susceptibilities and analysis of genes related to penicillin or macrolide resistance in Streptococcus pneumoniae. AB - One hundred and seventy-seven strains of Streptococcus pneumoniae derived from respiratory specimens between 1987 and 2001 were evaluated for their antimicrobial susceptibilities and distribution of genes related to penicillin and macrolide resistance. Resistance rates tended to be higher for the 1996-2001 isolates than for the 1987-1995 isolates for all beta-lactams tested. For benzylpenicillin the MIC(90) value of the isolates derived between 1996 and 2001 was 1.56 mg/L, while that of strains isolated between 1987 and 1990 was 0.05 mg/L. Furthermore, the number of strains susceptible to macrolides also decreased, but only two strains isolated in 1993 were resistant to levofloxacin of the 177 S. pneumoniae strains tested. When of genes relating to penicillin resistance were analysed using PCR with primers specific to susceptible alleles, although more than 50% of strains from 1987 to 1990 and 1991 to 1995 revealed no mutations in the pbp 1a, 2x and 2b genes, only 30.0% of strains derived between 1996 and 2001 showed no mutations in the pbp gene. Strains having mutations in all three pbp genes (1a, 2x and 2b) by the PCR method increased from only 2.2% in the 1987-1990 derived strains to 27.5% in the 1996-2001 strains. Furthermore, 64.1 and 60.0% of the isolates from 1987 to 1990 and 1991 to 1995, respectively, did not possess either the mefA or ermB by PCR analysis. Conversely, 75.0% of isolates from 1996 to 2001 possessed mefA and/or ermB. These genetic changes may explain the increase in the number of penicillin and macrolide resistant strains. We believe that it is important to evaluate changes in MIC as well as genetic mutations in order to select the most appropriate therapy for S. pneumoniae infections. PMID- 15288311 TI - Evaluation of genetic determinants involved in beta-lactam- and multiresistance in a surgical ICU. AB - Antibiotic resistance is a major and well-known problem in intensive care units (ICUs) world-wide and previously susceptible isolates become resistant through the acquisition of resistance determinants from other bacteria or the development of mutations, as is the case in beta-lactam resistance. We evaluated the presence of resistance determinants involved in beta-lactam resistance and multi resistance in order to establish the contribution of horizontal gene transfer to the spread of resistance in a surgical ICU during an antibiotic rotation study. Pseudomonas aeruginosa and Enterobacteriaceae isolates were selected and iso electric focusing (IEF), DNA-typing methods such as specific beta-lactamase and specific integron PCRs were performed to determine the presence of beta lactamases. The PCRs specific for IMP-1, OXA-1, and VIM-type beta-lactamases performed on the selected P. aeruginosa and Enterobacteriaceae isolates with MICs for cephalosporins >1 mg/l did not demonstrate any of these beta-lactamases. IEF for 14 pseudomonads, representing 7 genotypes from 9 patients, showed a beta lactamase with a pI larger than 8.5 in 13 of the isolates. The integrase PCR was positive for only five isolates from three patients and conserved segment PCR showed integrons of variable sizes (700, 900, 1,400 and 1,500 bp). Each patient had its own integron types. It can be concluded that integrons and associated resistance determinants played only a minor role in the surgical ICU and beta lactam resistance among P. aeruginosa isolates was most likely due to the derepression of its AmpC gene. PMID- 15288312 TI - A Bacteroides thetaiotamicron porin that could take part in resistance to beta lactams. AB - The aim of this study was to investigate porin absence or deficiency in two Bacteroides thetaiotaomicron strains resistant to amoxicillin combined with clavulanic acid. Their outer membrane protein (OMP) extracts and those of two susceptible strains were analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and compared to detect differences between the strains. A protein band of interest at around 70 kDa electro-eluted for each strain, was tested in a liposome swelling assay. A decrease in initial absorbency was noted for the two susceptible strains but not for the two resistant strains. The liposome swelling of the two susceptible strains was directly visualized by photon microscopy and then photographed. This suggested a B. thetaiotaomicron porin of around 70 kDa could take part in resistance to beta-lactams. PMID- 15288313 TI - Streptococcus pneumoniae in vitro development of resistance to amoxicillin/clavulanic acid, cefaclor, levofloxacin and azithromycin. AB - The purpose of this study was to determine effect of repeated exposure to sub inhibitory concentrations of amoxicillin/clavulanic acid on the development of resistance in Streptococcus pneumoniae. Other agents, azithromycin, cefaclor and levofloxacin, were also tested. Twenty S. pneumoniae were passaged for 9 days in the presence of sub-inhibitory concentrations of each antimicrobial agent and MICs determined by NCCLS macro-dilution method. There was a four-fold increase in amoxicillin/clavulanic acid MICs for 2 of 20 isolates. Three of 9 tested against cefaclor, 11 of 13 tested against azithromycin and 9 of 20 tested against levofloxacin showed > or =4-fold increase. Amoxicillin/clavulanic acid was the most stable of the agents tested. Cefaclor MICs were also fairly stable. Azithromycin and levofloxacin MICs were most affected. PMID- 15288314 TI - In vitro activities of mutant prevention concentration-targeted concentrations of fluoroquinolones against Staphylococcus aureus in a pharmacodynamic model. AB - To test the validity of the mutant selection window, we simulated mutant prevention concentration-targeted fluoroquinolone concentrations using an in vitro model with infected fibrin clots. Therapeutic ciprofloxacin (peak 5 microg/mL; t(1/2) 4 h), gatifloxacin (3.5 microg/mL; 8h), gemifloxacin (1.25 microg/mL; 8 h), levofloxacin (6 microg/mL; 6 h) and moxifloxacin (4.5 microg/mL; 12 h) were tested against methicillin-susceptible and -resistant Staphylococcus aureus, as were mutant prevention concentration (MPC)-targeted regimens achieving a trough of 1/4x or 2x MPC. MIC/MPC for MSSA K553 were 0.125/2, 0.03/0.125, 0.03/0.063, 0.125/1 and 0.015/0.25 microg/mL for ciprofloxacin, gatifloxacin, gemifloxacin, levofloxacin and moxifloxacin, respectively. Corresponding values for MRSA 494 were 0.125/1, 0.063/0.125, 0.03/0.063, 0.125/0.5 and 0.063/0.125 microg/mL. All regimens produced efflux mutants of MSSA K553. For MRSA 494, therapeutic and 1/4x MPC levofloxacin regimens produced resistance, whereas only 1/4x MPC regimens of gatifloxacin, gemifloxacin, and moxifloxacin produced resistance. All ciprofloxacin regimens produced resistance. Ciprofloxacin 1/4x MPC and therapeutic levofloxacin caused outgrowth of GrlA mutants (S80Y amino acid substitution); efflux mutants were isolated in all other cases. Overall, gatifloxacin, gemifloxacin, and moxifloxacin displayed a lesser propensity to select resistant isolates of S. aureus than ciprofloxacin and levofloxacin. The mutant selection window premise appeared valid for MRSA only. Additional studies are necessary to define the applicability of the MPC. PMID- 15288315 TI - Comparison of minimal inhibitory and mutant prevention drug concentrations of 4 fluoroquinolones against clinical isolates of methicillin-susceptible and resistant Staphylococcus aureus. AB - Staphylococcus aureus remains an important human pathogen affecting both outpatients and those hospitalized. Increasing antimicrobial resistance is global but prevalence rates are variable for different geographical areas. Fluoroquinolones have been used to treat S. aureus infections and the newer quinolones have enhanced in vitro activity against this organism. The mutant prevention concentration (MPC) defines the antimicrobial drug concentration threshold that would require an organism to simultaneously possess two mutations for growth in the presence of the drug. We tested clinical isolates of methicillin-susceptible (MSSA) and methicillin-resistant (MRSA) S. aureus by minimum inhibitory concentration (MIC) and MPC against gatifloxacin, gemifloxacin, levofloxacin and moxifloxacin. For MSSA strains, the rank order of potency based on MIC(90) values were gemifloxacin (0.063 mg/l) = moxifloxacin (0.063 mg/l) > gatifloxacin (0.05 mg/l) = levofloxacin (0.25 mg/l) and by MPC values moxifloxacin (0.25 mg/l) > gemifloxacin (0.5 mg/l) > gatifloxacin (1 mg/l) = levofloxacin (1mg/l). For 87% of the isolates the MPC value was 0.5 mg/l for gatifloxacin. The rank order of potency based on the time the serum drug concentration exceeded the MPC(90), was as follows: moxifloxacin (>24 h) > levofloxacin (>18 h) > gatifloxacin (12 h) > gemifloxacin (9 h). Serum drug concentration remained in excess of the MPC(87) for 24 h for gatifloxacin. Both MIC(90) and MPC(90) values were higher against MRSA strains and the time above the MPC(90) was significantly shorter for all agents. PMID- 15288316 TI - Urinary concentrations and bactericidal activities of newer fluoroquinolones in healthy volunteers. AB - Eleven healthy male subjects participated in a crossover study to compare the urine concentrations and bactericidal activities of newer fluoroquinolones against common uropathogens. Each volunteer received a single oral dose of gatifloxacin (400 mg), levofloxacin (250 mg), moxifloxacin (400 mg) and trovafloxacin (200 mg), and a urine sample was obtained at 2, 6, 12 and 24 h after the dose. Urine concentrations were highest with gatifloxacin and levofloxacin and lowest with trovafloxacin. Each drug concentration was studied against a levofloxacin susceptible and moderately-susceptible strain of Escherichia coli (minimal inhibitory concentration, MICs: 0.125 and 4 mg/l), K. pneumoniae (MICs: 0.125 and 4 mg/l), Pseudomonas aeruginosa (MICs: 0.5 and 4 mg/l) and Enterococcus faecalis (MICs: 0.25 and 4 mg/l). The duration of urine bactericidal activity (UBA) was based upon the median bactericidal titre at each time period. Both gatifloxacin and levofloxacin exhibited prolonged (> or = 6 h) UBA against all of the study isolates. Moxifloxacin exhibited prolonged UBA against both isolates of E. coli, K. pneumoniae and E. faecalis but not against either strain of P. aeruginosa. Prolonged UBA was not observed for trovafloxacin against the moderately-susceptible strains with the exception of E. faecalis. Furthermore, UBA was not observed for trovafloxacin against the susceptible strain of P. aeruginosa. Although these newer fluoroquinolones exhibited similar in vitro activity against these uropathogens, only those compounds with the highest urinary concentrations (gatifloxacin and levofloxacin) produced prolonged UBA against both strains of P. aeruginosa. The findings from this study suggest that both microbiological activity and urinary concentrations are important parameters to consider when choosing a fluoroquinolone for empirical treatment of urinary tract infections (UTIs). PMID- 15288318 TI - In vitro antagonism between beta-lactam and macrolide in Streptococcus pneumoniae: how important is the antibiotic order? AB - We found that the in vitro interaction between penicillin or cefotaxime and erythromycin against Streptococcus pneumoniae varies depending on the order of antibiotic exposure. Time-kill experiments were performed with penicillin, cefotaxime, erythromycin and different order combinations of both beta-lactams with erythromycin. The mean difference between the colony count at 0 and 6h for penicillin, cefotaxime and erythromycin tested separately was 3.5 log cfu/mL, 2.4 and 1.5 respectively for susceptible strains. The mean difference for the combination of beta-lactam and erythromycin studied simultaneously was 1.8 log cfu/mL for these strains. The association of penicillin or cefotaxime with erythromycin added two hours later showed an activity similar to those of beta lactam alone (mean difference was 3.0 for this association with penicillin and 2.5 with cefotaxime). Therefore, the antagonistic effect of macrolide activity could be less important if erythromycin was administrated after beta-lactam. PMID- 15288317 TI - Comparative pharmacodynamics of the new fluoroquinolone ABT492 and ciprofloxacin with Escherichia coli and Pseudomonas aeruginosa in an in vitro dynamic model. AB - The killing kinetics of Escherichia coli and Pseudomonas aeruginosa were compared when exposed to ABT492 and ciprofloxacin. E. coli ATCC 25922 and a clinical isolate of P. aeruginosa 4226 were exposed to ABT492 (single dose) and ciprofloxacin (two 12 h doses) at the ratios of area under the curve (AUC) to MIC varying from 60 to 480 h and at clinically achievable AUC/MIC ratios of ABT492 (1,740 and 140 h, respectively) and ciprofloxacin (2,200 and 120 h, respectively) that correspond to a 400 mg dose of ABT492 and two 500 mg doses of ciprofloxacin. In addition, a double dose of ABT492 (800 mg; AUC/MIC 280 h) and two 12 h doses of ABT492 (2 x 400 mg) were used with P. aeruginosa. Maximal reductions in the starting inoculum of E. coli and P. aeruginosa were greater with ABT492 than with ciprofloxacin at a given AUC/MIC ratio (60-480 h), whereas the times to regrowth were shorter with ABT492. A specific AUC/MIC relationship of the antimicrobial effect was inherent in each quinolone-pathogen pair. With both E. coli and P. aeruginosa, AUC/MIC plots of the area between the control growth and the time kill curves (I(E)) were steeper for ciprofloxacin than ABT492 and they were species-independent. The effect of ABT492 on E. coli at the clinically achievable AUC/MIC ratio (1740h) was more pronounced than the respective AUC/MIC of ciprofloxacin (2,200 h). With P. aeruginosa, a 140 h AUC/MIC of ABT492 (400 mg as a single dose) provided 1.8-fold less effect than a 120 h AUC/MIC of ciprofloxacin (2 x 500 mg). However, two 12 h doses of ABT492 (AUC/MIC 2 x 140 h) but not a double single dose (800 mg) were more efficient than ciprofloxacin. These findings predict comparable efficacies of clinically achievable AUC/MICs of ABT492 and ciprofloxacin against E. coli (q.d. versus b.i.d. quinolone dosing) and P. aeruginosa at b.i.d. but not at q.d. ABT492. PMID- 15288319 TI - Azithromycin versus comparative therapy for the treatment of community acquired pneumonia. AB - A 3-day course of azithromycin was compared with the 10 days of other antibiotics, which general practitioners routinely use as therapy for community acquired pneumonia (CAP). The study was a prospective open labelled, randomised, multicentre, comparative study from five family clinics. Patients with clinical and radiological evidence of pneumonia were included. The pneumonia resolved in 98.4% (61/62) of patients treated with azithromycin and in 87% (40/46) of patients treated with other antibiotics (P < 0.017). Restitution of normal function at home (2.3 +/- 1.2 and 4.3 +/- 2.6 days) and return to work (3.4 +/- 2.0 and 5.5 +/- 3.1 days) was more rapid among the group treated with azithromycin ( P < 0.001). Three days of azithromycin was more convenient and cost effective than the comparators used to treat pneumonia. PMID- 15288320 TI - In vitro activity of older and newer fluoroquinolones against efflux-mediated high-level ciprofloxacin-resistant Streptococcus pneumoniae. AB - The effect of high-level efflux activity on the MICs of fluoroquinolones against Streptococcus pneumoniae in the absence of topoisomerase mutations leading to fluoroquinolones resistance was investigated. A S. pneumoniae ATCC 46619-derived strain with high-level efflux activity was obtained (SP-25A). Both the parent and obtained strains were tested against efflux substrates acriflavine (Acr) and ethidium bromide (EtBr), and against norfloxacin (NFX), ciprofloxacin (CFX), levofloxacin (LFX), moxifloxacin (MFX), trovafloxacin (TVX) and sitafloxacin (SFX), in presence and absence of the efflux pump inhibitor reserpine. gyrA, gyrB, parC and parE QRDR genes were amplified by PCR and sequenced. MICs of NFX and CFX against SP-25A were 64-fold higher than parent strain MICs (256 mg/L versus 4 mg/L and 64 mg/L versus 1mg/L, respectively). MIC of LFX increased from 1 to 4 mg/L and MICs of MFX, TVX and SFX remained virtually unchanged (0.1-0.2 mg/L). MICs of Acr and EtBr against SP-25A were 8- and 16-fold higher than against parent strains. In both cases, reserpine reverted MICs to the parent strain values (1 and 0.2 mg/L). Only parE showed two mutations leading to a Pro(454) --> Ser and Glu(443) changes, which have previously been shown not to lead to significant fluoroquinolones MIC increases. SP-25A showed a significant increase of MICs of the hydrophilic fluoroquinolones, apparently derived only from efflux activity. Efflux activity, at these high levels, can lead to high level resistance to older hydrophilic fluoroquinolones, but does affect newer fluoroquinolones such as moxifloxacin, trovafloxacin and sitafloxacin. PMID- 15288321 TI - Comparative randomized pilot study of azithromycin and doxycycline efficacy in the treatment of prostate infection caused by Chlamydia trachomatis. AB - The study included 125 adult patients (> 18years of age) who had symptoms of chronic prostatitis and proven presence of Chlamydia trachomatis. The presence of C. trachomatis was confirmed in expressed prostatic secretion or in voided bladder urine collected immediately after prostatic massage by a DNA/RNA hybridization method and/or by isolation on McCoy culture and then by immunofluorescent typing with monoclonal antibodies. The patients were randomized in the ratio 2/1; azithromycin/doxycycline, to receive a total of 4.0 g azithromycin over 4 weeks, given as a single dose of 1 x 1000 mg weekly for 4 weeks or doxycycline 100 mg b.i.d. for 28 days. Patients' sexual partners were treated at the same time. Clinical and bacteriological efficacy was evaluated 4-6 weeks after the end of therapy. In the group of patients with chlamydial infection of the prostate, there was no significant difference between the eradication rates (azithromycin 65/82, doxycycline 33/43; P = 0.82) and the clinical cure rates (azithromycin 56/82, doxycycline 30/43; P = 0.94) of the two antimicrobials. PMID- 15288322 TI - Ciprofloxacin resistant Neisseria gonorrhoeae in South Africa. PMID- 15288323 TI - In vitro activity of oxacillin, vancomycin and teicoplanin against staphylococci isolated from patients on surgical and coronary intensive care units. PMID- 15288324 TI - In vitro susceptibility of Coxiella burnetii to azithromycin, doxycycline, ciprofloxacin and a range of newer fluoroquinolones. PMID- 15288325 TI - Blood stream isolates in Austria and Germany--results of three multi-centre studies. PMID- 15288326 TI - National Academy of Neuropsychology: President's address. The future of neuropsychology. PMID- 15288327 TI - A meta-analysis of the sensitivity and specificity of the Stroop Color and Word Test with children. AB - The purpose of this paper is to examine the extent to which the Stroop Color-Word Test demonstrates sensitivity and specificity for the identification of executive function deficits in children and adolescents. Meta-analytic methods were used to identify executive function deficits associated with attention deficit hyperactivity disorder (ADHD) and other developmental disorders. Weighted effect sizes were calculated for all studies found that compared groups of children on the Stroop task. Results indicated that across studies, children and adolescents with ADHD fairly consistently exhibited poorer performance when compared to individuals without clinical diagnoses on the Stroop task as measured by the weighted Word, Color, Color-Word, and Interference scores. The Stroop task did not discriminate ADHD groups from other clinical groups consistently across studies. In conclusion, while impaired performance of the Stroop task may be indicative of an underlying neurological disorder related to frontal lobe dysfunction, poor performance is not sufficient for a diagnosis of ADHD. PMID- 15288328 TI - The neuropsychological profile of vascular cognitive impairment--no dementia: comparisons to patients at risk for cerebrovascular disease and vascular dementia. AB - Hachinski and co-workers have used the term vascular cognitive impairment-no dementia (VaCIND) to represent the earliest stages of cognitive decline associated with vascular changes [Neurology 57 (4) (2001) 714]. However, the neuropsychological profile of vascular CIND remains unclear. Twenty-five healthy elders, 29 individuals at risk for cerebrovascular disease (R-CVD), 18 individuals with VaCIND, and 26 individuals with vascular dementia (VaD) were examined to determine whether patterns of neuropsychological assessment performance can assist in the differentiation of patients at varying levels of risk and severity for cerebrovascular disease and VaD. The R-CVD group performed within normal expectations on most cognitive measures as compared to the elderly control sample and published clinical norms. Relative to elderly controls, the VaCIND group demonstrated significant difficulties on measures of cognitive flexibility, verbal retrieval, and verbal recognition memory, but not on measures of confrontational naming or verbal fluency. The VaD group was impaired on all cognitive measures assessed. The current findings suggest that poor cognitive flexibility and verbal retrieval in the context of preserved function in other domains may characterize the prodromal stage of VaD. PMID- 15288329 TI - Speed and memory in the WAIS-III Digit Symbol--Coding subtest across the adult lifespan. AB - The primary role of speed in determining Digit Symbol scores is well established. Among the important questions that remain to be resolved are: (1) whether speed accounts for all of the age-related decline in Digit Symbol scores, and (2) whether memory ability makes any significant contribution to Digit Symbol performance, especially after controlling for speed. We analyzed data from the WAIS-III/WMS-III standardization sample to resolve these issues. As expected, speed (Digit Symbol-Copy) correlated very strongly with Digit Symbol--Coding. Memory (Digit Symbol--Incidental Learning or WMS-III index scores) correlated more moderately with Digit Symbol-Coding. Even after controlling for variance in Coding explained by Copying, a statistically significant proportion of the residual variance was explicable in terms of memory functions. The contribution of memory to Digit Symbol--Coding, while relatively small, is real. In addition, a small portion of the age-associated decline in Coding scores cannot be accounted for by Copying scores. PMID- 15288330 TI - Estrogen replacement therapy: effects on the cognitive functioning and clinical course of women with Alzheimer's disease. AB - This study examined the effects of unopposed estrogen replacement therapy (ERT) on the cognition and clinical course of women who have been diagnosed with probable Alzheimer's disease. The records of 99 women diagnosed with probable Alzheimer's disease according to NINCDS-ADRDA criteria, and in some cases autopsy confirmed as well, were analyzed. Depression, education, and age were all considered in the analysis. Results suggest that after an average of four years after symptoms appeared, women on ERT at the time of their diagnoses had similar cognitive functioning compared to those who are not on ERT. There were, however, significant differences in the clinical course of the illness between the two groups, with the non-ERT group reporting more "non-memory" problems as initial symptoms in addition to a having greater degree of overall behavioral deficit in activities of daily living. PMID- 15288331 TI - Remote memory in advanced Alzheimer's disease. AB - The present study investigated remote memory functioning in advanced dementia of Alzheimer type (DAT). Remote memory measures included the Autobiographical Memory Enquiry (AME) and autobiographical fluency (AF), as well as recognition of remote public events and famous faces. Ten DAT patients were selected with Mini-Mental State Exam (MMSE) scores less than 15. Compared to healthy controls, DAT patients were impaired on all measures of remote memory, although autobiographical fluency for names was only borderline impaired. There was no clear evidence of a monotonic temporal gradient with the AME; however, both subjects groups showed better preserved early compared to more recent public-event memory. A measure of semantic fluency was correlated with autobiographical and remote public-event memory among the patients only. Overall, results indicate a minimal outline of preserved remote memory in advanced DAT, with evidence of an association between remote memory performance and executive functioning. PMID- 15288332 TI - Evidencing inhibitory deficits in Alzheimer's disease through interference effects and shifting disabilities in the Stroop test. AB - To investigate the contribution of inhibitory deficits in the deterioration of executive function abilities in Alzheimer's disease (AD), a modified version of the Stroop test was submitted to 44 AD patients and 44 elderly controls. Half of the subjects performed successively the Interference Stroop task, the two control tasks and the Reverse Stroop task, and half performed the Reverse Stroop task, the control tasks and finally the Interference Stroop task. This experimental design allowed to assess inhibitory deficits by measuring classical interference effects but also by measuring the ability to shift between tasks instructions. Results confirmed AD patients' difficulty in suppressing the automatic response of reading in the Interference Stroop task. Moreover, AD patients presented worsened performances in the Interference task when administered after the Reverse task, and a Reverse Stroop effect was found in the patients revealing their difficulty in suppressing a previously relevant rule in order to learn a new one. PMID- 15288333 TI - A longitudinal, controlled study of patient complaints following treated mild traumatic brain injury. AB - This study provided 3-month follow-up data to a previous paper that compared symptom complaints of patients with mild traumatic brain injury (MTBI) with those of non-injured control participants within 1 month of injury. The 110 MTBI patients and 118 control participants were group-matched on age, gender, education level, and socioeconomic status. As a group, MTBI patients no longer endorsed significantly more symptoms (M = 14.09, S.D. = 10.77) than did the control group (M = 12.56, S.D. = 8.46, P = .232). Only 3 of the 43 queried symptoms were endorsed by significantly more (Bonferroni-corrected P < .00116) MTBI patients than controls. Using the same Bonferroni-corrected criteria, 10 of the 43 symptoms were endorsed at a significantly higher severity level by MTBI patients. Overall, the treated MTBI group's symptom complaints diminished from baseline to 3 months post-injury, with relatively few differences remaining between the two groups. PMID- 15288334 TI - Effects of incentive and preparation time on performance and classification accuracy of standard and malingering-specific memory tests. AB - The effects of incentive and preparation on performance and classification accuracy of standard and malingering-specific tests were investigated in a simulation study using a 2 (no incentive vs. a $20 incentive) x 2 (immediate vs. delayed preparation) factorial design. Eighty undergraduate students and 15 individuals with traumatic brain injury were administered standard (viz., Digit Span and Visual Memory Span from the WMS-R) and malingering-specific (viz., the Rey 15-Item Memory Test and the Multi-Digit Memory Test) memory tests. Preparation time was found to have a significant effect on performance and classification accuracy on a number of these tests, but incentive was found to have a significant effect on the performance but not the classification accuracy of one test (viz., the Multi-Digit Memory Test). These findings suggest that extra-test variables such as incentive and preparation time should be taken into consideration in evaluating the utility of standard and malingering-specific memory tests in detecting malingering. PMID- 15288335 TI - RBANS analysis of verbal memory in multiple sclerosis. AB - Patients with neurodegenerative diseases that cause mainly subcortical pathology often exhibit impairment when required to recall lists of unrelated words, but their memories are supposedly improved by test procedures that promote retrieval such as recognition or improve the organization of the to-be-remembered materials. Difficulties with floor effects on free recall and ceiling effects on recognition and other methodological concerns raise doubts about the validity of existing studies that tested these ideas. Using the verbal memory subtests of the RBANS, we [Arch. Clin. Neuropsychol. 18 (2003) 509] expressed each patient's performances on Story Memory, List Learning, Story Recall, List Recall, and List Recognition as Z scores relative to his or her age group. Then, the Z scores were subtracted pairwise to test hypotheses about the nature of memory in Parkinson's disease (PD). Contrary to expectation, patients with PD did not show better immediate or delayed recall of stories relative to lists and they did not show better recognition than recall. In the present investigation, the same methodology was used to study verbal memory in multiple sclerosis, a disease that primarily affects subcortical structures. In contrast to previous results for patients with PD, the patients with MS exhibited better recall of stories than of lists and better List Recognition than Recall. Differences in the pathology of entorhinal regions in PD and MS may contribute to the differing patterns of memory impairment of these patients. The results emphasize that most patients with MS with memory impairments have deficits that are relatively mild and potentially remediable. PMID- 15288336 TI - The Biber Cognitive Estimation Test. AB - Normative data from 113 participants, and cross-validation data from 49 additional participants, are presented for the Biber Cognitive Estimation Test (BCET), a 20-item test with five estimation questions in each of four categories: time/duration, quantity, weight, and distance. In Study 1, the range of normal answers is provided for each item, and a cut-off for impaired performance is suggested. Although very low IQ or education levels would be expected to invalidate this test as a measure of estimation skills, participants in the current sample made few errors. In Study 2, a cross-validation suggested a slightly more conservative cut-off score for abnormality. Study 3 examined cognitive estimation in demented (dementia of the Alzheimer's type and dementia syndrome of Parkinson's disease) versus intact elderly participants. Results indicated that the BCET was able to distinguish between demented and intact elderly participants. PMID- 15288338 TI - Neutron activation-based gamma scintigraphy in pharmacoscintigraphic evaluation of an Egalet constant-release drug delivery system. AB - This paper is a report from a pharmacoscintigraphic study with an Egalet constant release system containing caffeine and natural abundance samarium oxide. First the formulation was tested in vitro to clarify integrity during irradiation in the nuclear reactor. Then six healthy male volunteers were enrolled into the in vivo study. The in vitro release of caffeine obeyed all the time linear zero order kinetics. The in vivo release of radioactive Sm2O3 consisted of three consequent linear phases with different slopes. The release rate was fastest while the product was in the small intestine and slowest when the product was in the descending colon. In terms of the bioavailability of caffeine, the most important factor seemed to be the residence time in the ascending and transverse colon. A long residence time in these sections led to high AUC values for caffeine. PMID- 15288339 TI - Nasal insulin delivery in the chitosan solution: in vitro and in vivo studies. AB - The effects of chitosan concentrations, osmolarity, medium and absorption enhancers in the chitosan solution on nasal insulin delivery were studied in vitro and in vivo. The penetration of insulin through the mucosa of rabbit nasal septum was investigated by measuring the transmucosal flux in vitro, while the nasal absorption of insulin in vivo was assessed by the efficiency in lowering the blood glucose levels in normal rats. It was demonstrated that increasing concentrations of chitosan up to 1.5% (w/v) caused an increase in the permeability of insulin across the nasal mucosa. Insulin given intranasally in hypo- or hyperosmotic formulation showed a higher hypoglycemic effect than insulin delivered in isoosmotic formulation. Insulin formulation in chitosan solution prepared with deionized water brought to a higher relative pharmacological bioavailability (Fr) value than that prepared with 50 mM pH 7.4 phosphate buffer. A formulation containing both 1% chitosan and 0.1% ethylenediaminetetraacetic acid (EDTA), 5% polysorbate 80 (Tween 80) or 1.2% beta cyclodextrin (beta-CD) did not lead to a higher Fr than insulin formulated with 1% chitosan alone. The formulation containing both 5% hydroxypropyl-beta cyclodextrin (HP-beta-CD) and 1% chitosan was more effective at reducing blood glucose levels than the formulation containing 5% HP-beta-CD or 1% chitosan alone. The studies indicated that chitosan concentrations, osmolarity, medium and absorption enhancers in chitosan solution have significant effect on the insulin nasal delivery. The results of in vitro experiments were in good agreement with that of in vivo studies. PMID- 15288340 TI - Factors governing the in vivo tissue uptake of transferrin-coupled polyethylene glycol liposomes in vivo. AB - Liposomes, coated with transferrin (Tf)-coupled polyethylene glycol are considered to be potent carriers for drug delivery to various organs via receptor mediated endocytosis. Since Tf receptors were ubiquitously expressed in various organs, additional perturbation of the liposomes such as regulation of the size may be required to exhibit the tissue selectivity. In the present study, the effect of size on the uptake of transferrin-coupled polyethylene glycol liposomes (Tf-PEG-L) to various organs was investigated. In liver and brain, Tf-dependent uptake was found to be dependent on the size of the liposomes used. In small liposomes with a diameter of 60-80 nm, Tf-PEG-L was taken up to these organs more efficiently than PEG-L. This Tf-dependent uptake for small liposomes decreased by the high dose administration, suggested that Tf-PEG-L is taken up via Tf receptor mediated endocytosis even under the physiological condition, in which plasma concentration of endogenous Tf remains high. On the other hand, Tf receptor mediated uptake was also observed in the heart, but size-dependency was not observed in this case. Collectively, these results indicate that size dependency in the uptake of Tf-PEG-L is tissue-dependent and therefore, controlling the size of Tf-PEG-L may be useful for the success of tissue targeting. PMID- 15288341 TI - Development and in vitro evaluation of furosemide transdermal formulations using experimental design techniques. AB - The in vitro skin permeation of furosemide, a commonly used loop diuretic, through human epidermis, as a preliminary step towards the development of a transdermal therapeutic system, was examined. A screening study was carried out, in order to estimate the effects of the type, the concentration of enhancer and the concentration of gelling agent on the cumulative amount of furosemide permeated through human epidermis, using a 3(3) factorial design. The type and the concentration of enhancer were further evaluated as they were found to affect significantly furosemide permeation. In order to further increase the amount of the drug permeated, the combination of two enhancers, Azone and oleyl alcohol, at three concentration levels was employed, using an optimization technique. The results indicated that higher amounts of furosemide permeated were observed when Azone was used at 5.0-6.5% (v/v) and oleyl alcohol at 7.5-9% (v/v), in the gels used. These formulations seem to be suitable for possible transdermal delivery of furosemide for pediatric use. PMID- 15288342 TI - The depolymerization of chitosan: effects on physicochemical and biological properties. AB - Chitosan has been extensively used as an absorption enhancer for macromolecules and as gene delivery vehicle. Both properties are molecular weight (MW) dependent. Here, we investigate factors affecting the oxidative depolymerization of chitosan and physicochemical properties of the resulting polymer fractions including their cytotoxicity. The molecular weight of the depolymerized chitosan was influenced by the initial concentration and the source of chitosan. At constant initial concentrations, the molecular weight decreased linearly with the chitosan/NaNO2 ratio and was a function of logarithm of the reaction time. Chitosan with larger molecular weight was more sensitive to depolymerization. No structural change was observed during the depolymerization process by infrared and proton nuclear magnetic resonance spectroscopy. In addition, thermal properties of chitosan fragments were studied by thermal gravimetric analysis and it was found that the decomposition temperature was molecular weight dependent. Furthermore, the solubility of different molecular weight chitosan was assayed as a function of pH and it increased with decreasing molecular weight. The cytotoxicity of chitosan was concentration dependent but almost molecular weight independent according to MTT assay using L929 cell line recommended by USP26. In summary, low molecular weight fractions of chitosan may potentially useful for the design of drug delivery systems due to the improved solubility properties. PMID- 15288343 TI - Evaluation of SCF-engineered particle-based lactose blends in passive dry powder inhalers. AB - The objective of this study was to assess the performance of SCF-engineered budesonide and albuterol sulfate powder blends in passive dry powder inhalers (DPI) relative to micronized drug blends. A number of lactose grades for inhalation were screened and the appropriate carrier and drug-to-lactose blending ratio were selected based on drug content and emitted dose uniformity. Aerosol performance was characterized by Andersen cascade impaction. Blend formulations of SEDS (solution enhanced dispersion by supercritical fluids) budesonide and albuterol exhibited a significant drug content uniformity (7-9% RSD) improvement over micronized drug blends (16-20% RSD). Further, the SEDS formulations demonstrated higher emitted dose and reduced emitted dose variability (10-12% RSD) compared to micronized powders (21-25% RSD) in the Turbospin, albeit without significant enhancement of the fine particle fraction. In contrast, SEDS powders exhibited increased fine particle fractions over micronized blends in the Clickhaler; improvements were more pronounced with albuterol sulfate. The performance enhancements observed with the SEDS powders are attributed to their increased surface smoothness and reduced surface energy that are presumed to minimize irreversible drug-carrier particle interactions, thus resulting in more efficient drug detachment from the carrier particle surface during aerosolization. As demonstrated for budesonide and albuterol, SEDS may enhance performance of lactose blends and thus provide an attractive particle engineering option for the development of blend formulations for inhalation delivery. PMID- 15288344 TI - Influence of lipolysis and droplet size on tocotrienol absorption from self emulsifying formulations. AB - A single dose comparative bioavailability study was conducted to evaluate the bioavailability of tocotrienols from two self-emulsifying formulations, one of which produced an emulsion that readily lipolysed under in vitro condition (SES A), while the other produced a finer dispersion with negligible lipolysis (SES-B) in comparison with that of a non-self-emulsifying formulation in soya oil. The study was conducted according to a three-way crossover design using six healthy human volunteers. Statistically significant differences were observed between the logarithmic transformed peak plasma concentration (Cmax) and total area under the plasma concentration-time curve (AUC(0-infinity)) values of both SES-A and -B compared to NSES-C indicating that SES-A and -B achieved a higher extent of absorption compared to NSES-C. Moreover, the 90% confidence interval of the AUC(0 infinity) values of both SES-A and -B over those of NSES-C were between 2-3 suggesting an increase in bioavailability of about two-three times compared to NSES-C. Both SES-A and -B also achieved a faster onset of absorption. However, both SES-A and -B had comparable bioavailability, despite the fact that SES-B was able to form emulsions with smaller droplet size. Thus, it appeared that both droplet sizes as well as the rate and extent of lipolysis of the emulsion products formed were important for enhancing the bioavailability of the tocotrienols from the self-emulsifying systems. PMID- 15288345 TI - Validation of an ocular microdialysis technique in rabbits with permanently implanted vitreous probes: systemic and intravitreal pharmacokinetics of fluorescein. AB - The purpose of this work is to validate a novel ocular microdialysis sampling technique in rabbits with permanently implanted vitreous probes. This objective is achieved by studying the vitreous pharmacokinetics of fluorescein following systemic and intravitreal administration. The rabbits were divided into two groups (groups I and II) based on whether or not they were allowed a recovery period following surgical implantation of probes. The integrity of the blood retinal barrier was determined by the vitreal protein concentrations and the fluorescein permeability index. Vitreal protein concentrations returned to baseline 48 h after probe implantation and therefore experiments were conducted 72 h post-implantation of probes in rabbits where recovery period was allowed. The permeability indices for fluorescein after systemic administration in group I (without recovery period) and group II (with recovery period) indicated that the integrity of the blood-retinal barrier was maintained and were found out to be 0.55 +/- 0.27 and 0.71 +/- 0.38%, respectively, for the vitreous chamber. Following microdialysis probe implantation in the group II rabbits, the blood retinal barrier integrity was not compromised. A novel microdialysis technique in rabbits with permanently implanted probes for studying the pharmacokinetics of posterior segment has been developed and characterized. PMID- 15288346 TI - Solid state NMR perspective of drug-polymer solid solutions: a model system based on poly(ethylene oxide). AB - Poly(ethylene oxide) (PEO) was tested as a polymer matrix for solid dispersion to enhance drug bioavailability. Solid state nuclear magnetic resonance (NMR), X-ray diffraction (XRD), and transmission electron microscopy (TEM) were utilized to characterize the high miscibility between PEO and ketoprofen, a model for crystalline drugs with poor water solubility. The experimental data demonstrated that ketoprofen in the melt-processed blend formed a complete molecular dispersion within the amorphous domain of PEO, resulting in high molecular mobility of ketoprofen in the melt-processed blend that leads to enhanced dissolution rate of ketoprofen in aqueous media. Hydrogen bonds between the carboxylic group of ketoprofen and the ether oxygen of PEO, as detected by solid state NMR, are the likely source for the high miscibility between ketoprofen and PEO. Such drug/polymer molecular interactions promote dispersion of ketoprofen into amorphous phase of PEO at temperatures well below melting points of both crystalline ketoprofen and PEO. Consequently, melt-processing temperatures can be reduced significantly to avoid thermal degradation. The processing conditions can be also flexible while maintaining reproducibility of the physico-chemical properties of the blend. Furthermore, the high degree of drug/polymer molecular interactions stabilizes the morphology of the blend during storage. PMID- 15288347 TI - Paclitaxel loaded poly(L-lactic acid) (PLLA) microspheres. II. The effect of processing parameters on microsphere morphology and drug release kinetics. AB - The kinetics of solvent removal in microsphere preparation and their effect on the morphology and release characteristics of paclitaxel-loaded PLLA microspheres were determined. Microspheres were analyzed by SEM and DSC and in vitro paclitaxel release was monitored by HPLC. During manufacture, dichloromethane evaporated at a constant rate, which increased with dispersion stirring speed and decreased with increasing paclitaxel content. Paclitaxel-loaded microspheres had a dimpled surface, due to surface deposition of the drug, while controls were smooth. In the formation of larger microspheres, the deposition of drug in the surface slowed the solidification process resulting in drug-loading dependent thermal properties. Paclitaxel release did not follow diffusion kinetics, rather it was characterized by a large burst followed by a linear phase. We speculate that non-uniform (surface-rich) drug distribution in the microspheres may contribute to the deviation from the theoretical pattern of kinetics for diffusion from a sphere. PMID- 15288348 TI - Hydration of an amphiphilic excipient, Gelucire 44/14. AB - The incorporation of drugs into Gelucires has been reported to increase the dissolution rate of poorly soluble drugs, often leading to improved drug bioavailability. In pharmaceutical applications, it is important to know how the excipient interacts with the drug, and how the mixture behaves during manufacturing, storage as well as during administration. The uptake of water by an amphiphilic excipient, Gelucire 44/14, has been investigated in two ways: storage in humid air and addition of liquid water. During exposure to humid air, the uptake goes in stages that correspond to the dissolution of the components of the excipient, starting with the most hydrophilic ones: glycerol, then polyethylene glycol (PEG), PEG esters (PEG monolaurate and PEG dilaurate), and finally glycerides (trilaurin). At each stage, the remaining crystals are in equilibrium with an interstitial solution made of water and the dissolved components. In this range of hydrations, the total uptake is close to the sum of the equilibrium hydrations of the components. In the pharmaceutical formulation, the active ingredient could dissolve in the liquid phase. At larger hydrations, obtained through addition of liquid water, the state of Gelucire 44/14 differs from those of its components. Gelucire 44/14 forms a lamellar phase and this phase melts at 30 degrees C whereas the pure PEG esters form hexagonal and cubic mesophases. The cubic mesophases do not melt until the temperature exceeds 40 degrees C. At body temperature, all crystals in Gelucire 44/14 melt to an isotropic fluid as soon as the total water content exceeds 5%. Therefore the formulation of amphiphilic excipients can be optimized to avoid the formation of mesophases that impede dissolution of the excipient at body temperature. PMID- 15288349 TI - Determination of tackiness of chitosan film-coated pellets exploiting minimum fluidization velocity. AB - The tackiness of aqueous chitosan film coatings and effects of anti-sticking agents on sticking tendency, were evaluated. A novel rapid method exploiting minimum fluidization velocity to determine tackiness was introduced and tested. The pressure difference over the miniaturized fluidized-bed was precisely recorded as a function of velocity of fluidization air. High molecular weight chitosan plasticized with glycerol was used as a film-forming agent. Magnesium stearate, titanium dioxide, colloidal silicon dioxide and glyceryl-1-monostearate (GMS) were studied as anti-sticking agents. Film coatings were performed in a miniaturized top-spray coater. The incorporation of anti-sticking agents led to a clear decrease in tackiness of the chitosan films, and magnesium stearate and GMS were shown the most effective. Film-coated pellets containing magnesium stearate and GMS as an anti-sticking agent were very easily fluidized (showing very low values of minimum fluidization velocity) and were thus classified as the best flowing and the least sticking samples. Both these additives were found anti sticking agents of choice for aqueous chitosan film coatings. Determination of the experimental minimum fluidization velocity in a fluidized bed, is a useful and sensitive method of measuring the tackiness tendency of film-coated pellets. PMID- 15288350 TI - In vitro and in vivo evaluation of a melamine dendrimer as a vehicle for drug delivery. AB - Cell-based and acute and subchronic in vivo toxicity profiles of a dendrimer based on melamine reveal that this class of molecules warrants additional study as vehicles for drug delivery. In cell culture, a substantial decrease in viability was observed at 0.1 mg/mL. For the acute studies, mice were administered 2.5, 10, 40 and 160 mg/kg of dendrimer via i.p. injection. At 160 mg/kg, 100% mortality was seen 6-12 h after injection. For the other cohorts, blood chemistry work revealed no renal damage was taking place at 48 h. Liver enzyme activity nearly doubled for the mice treated at 40 mg/kg suggesting hepatotoxicity. For the subchronic studies, three i.p. injections of 2.5-40 mg/kg of dendrimers were administered at 3-week intervals. No mortality was observed. Forty-eight hours following the last administration, blood chemistry revealed no renal damage, but liver damage was indicated by elevated serum enzyme activity at the highest dose. Histopathological data further confirms that doses up to 10 mg/kg show no hepatic damage at subchronic doses. However, subchronic doses at 40 mg/kg lead to extensive liver necrosis. PMID- 15288351 TI - Could ECT be effective in autism? AB - Autism is increasingly diagnosed, but therapeutic options are limited in many children. ECT is considered as a safe, effective, and life-saving treatment in people of all ages who suffer from affective disorders, acute psychosis, and, in particular, catatonia. There are recent speculations that certain types of autism may be the earliest expression of catatonia and that both disorders have identical risk factors. Therefore, ECT may improve autism and, if started early enough, may prevent further development of autistic symptoms in some children. The use of ECT in autism has never been systematically assessed. There have been two large ECT studies in children in the 1940s. Autism was not assessed in these studies because the autistic syndrome was just then being recognized as a separate entity. Findings from these studies add little to the hypothesis that ECT may be effective in autistic children, but attest to the safety and feasibility of ECT in children. Another limitation is the use of older ECT techniques. What may well be the greatest deterrent to use ECT in autism is widespread anti- ECT sentiment not only among the public but within the medical community as well. All child specialists--psychiatrists, neurologists, psychologists, and developmental pediatricians--should independently review the feasibility, potential, and risk of using ECT in autism. Unless anti-ECT prejudice can be overcome, it is unlikely that any ECT trial in autism is forthcoming. Research areas that may support the hypothesis that ECT is effective in autism should be pursued. First, any link between autism and catatonia should be further explored in clinical and biochemical studies. A GABA theory of autism and catatonia may be pivotal. Second, the role of abnormal GABAA receptor subunit genes in autism and catatonia should be further assessed. Candidate loci for autism and catatonia have been found on the long arm of chromosome 15 where three GABAA receptor subunits genes are located. The GABAA receptor beta 3 subunit gene (GABRB3) was the leading candidate gene for a subgroup of autism in two independent studies. Third, a novel genetic mouse model of autism should be tested. Mutant mice with a targeted deletion of the GABRB3 gene have a complete deficit of the beta 3 subunit of the GABAA receptor. This knockout mouse model seems promising to study developmental effects of altered GABAA receptor function as it relates to certain developmental disorders including autism. PMID- 15288352 TI - Just how happy is the happy puppet? An emotion signaling and kinship theory perspective on the behavioral phenotype of children with Angelman syndrome. AB - The favored level of parental investment in a child may differ for genes of maternal and paternal origin in the child. This conflict can be expressed in the phenomenon of genomic imprinting that refers to situations in which the same gene is differentially expressed depending on its parent of origin. Two disorders that show the effects of genomic imprinting--both at 15q11-q13--are Angelman Syndrome (AS) which is due to the absence of expression of maternally-inherited genes and Prader-Willi syndromes (PWS) which is due to the absence of expression of paternally-inherited genes. However, although both disorders can arise from the deletion of the same genetic region, the gustatory, behavioral, and affective characteristics of AS and PWS children are remarkably distinct. Recent research inspired by kinship theory has suggested the origins of these phenotypic differences may lie in the differential investment of each parent's genome in the AS or PWS child. Specifically, it is thought that each set of parental genes have different 'ideas' regarding how the child should behave towards the mother and how much investment they should look to extract. In normal cases, the trade-off between the competing parental genomes produces a behavioral equilibrium in the child. However, in pathological instances, particularly where gene expression is one-sided, the evolved behavioral strategies favored by the contributing genome will dominate the child's behavior. To date, research in the area of genomic conflict in AS and PWS children has primarily focusing on differences in post natal nutrition-related behaviors. The current paper extends this framework by offering an emotion and evolutionary signaling interpretation of the affective characteristics of AS children. A review of the affective characteristics of the two syndromes (PWS and AS) is presented before kinship and emotions theory are used to examine the functions that differential affect expression may serve in altering maternal investment. We expected that because the ultimate goal of paternal genes is to increase the child rearing burden of mothers, the Angelman behavioral phenotype should exhibit the emotion signaling characteristics that elicit levels of investment more consistent with paternal genetic interests. AS children display more positive, relative to negative, affect expressions (i.e. AS children laugh and smile more frequently than PWS children). In affect signaling theories, positive affect signals (i.e., smiling, laughing) have evolved to manipulate the sensory systems of receivers to increase social resources. In contrast, because the expression of some negative affects may indicate to the mother that the infant is not viable, negative affect expression is characteristically low among AS children. However, AS children may nonetheless have high levels of non-expressed anxiety because of its role in assisting the child (and its paternal genome) to maintain vigilance for changes in investment on the part of the mother. Overall, our kinship and emotion signaling analysis of AS children suggests that their global pattern of affect signaling represents one manifestation of an array of possible evolved strategies within the parental genome. Specifically, because AS exhibits the effects of paternally-inherited genes unhindered by the expression of maternally-inherited genes, the AS infant manifests a pattern of expression and non-expression that maximize maternal investment and thus paternal fitness. This theory is a significant departure from the standard but erroneous conjecture that a mother and child's inclusive fitness interests are one and the same. PMID- 15288353 TI - The poor man's epidural: systemic local anesthetics for improving postoperative outcomes. AB - The inflammatory response is an important determinant of outcome after major surgery. Perioperative excessive stimulation of the inflammatory and hemostatic systems plays a role in the development of postoperative ileus, ischemia reperfusion syndromes (e.g. myocardial infarction), hypercoagulation syndromes (e.g. deep venous thrombosis) and pain; together, these represent a significant fraction of major postoperative disorders. Epidurally administered local anesthetics prevent or modulate many of these processes. Since local anesthetics show inflammatory modulating properties in vitro and in vivo, they might also prevent these postoperative inflammatory processes when administered intravenously. Clinical studies have shown that perioperative local anesthetic administration significantly reduces the incidence of thrombosis and postoperative pain, shortens postoperative ileus and decreases hospital stay. On this basis we hypothesize that continuous intravenous administration of local anesthetic perioperatively might prevent or reduce several postoperative disorders resulting from excessive stimulation of inflammatory and hemostatic systems, and thereby improve surgical outcome. PMID- 15288354 TI - Obstetric denervation-gynaecological reinnervation: disruption of the inferior hypogastric plexus in childbirth as a source of gynaecological symptoms. AB - Difficult vaginal deliveries damage branches of the inferior hypogastric plexus as they are distributed to the pelvic viscera. The initial denervatory injury is often asymptomatic. Nerve fibre proliferation along the course of damaged nerve trunks results in chaotic reinnervation of the visceral stroma and subsequent sensory pelvic symptoms including chronic pelvic pain, painful heavy periods, painful intercourse, urinary frequency and urgency, irritable bowel symptoms and vulval pain. Each symptom may be described as pain, or discomfort, in response to light touch (allodynia). In other visceral sites allodynic syndromes are frequently associated with aberrant neural repair. Nerve fibre proliferation has now been reported in every viscus in the female pelvis though its antecedents have not been established. It is proposed that clinical presentations with one, or more, sensory pelvic symptoms occur some years after a difficult intrapartum episode; the initial denervatory injury is succeeded by nerve fibre proliferation in the affected organs. The precise pathoanatomy, mechanisms of intrapartum injury and clinical presentation remote from the intrapartum episode, may have obscured the common aetiology of many sensory pelvic syndromes. PMID- 15288355 TI - Syncope: facts and fiction. AB - According to the current state-of-art on the brainstem functional anatomy and reticular formation, authors believe that nucleus tractus solitarii (NTS) is the neural structure, which meets all the conditions of the hypothetical syncope generating, reflex centre. The afferent branch of this reflex arc represents information from different visceral sources including the brain itself. The efferent branch of this reflex arc is reticular activating system (RAS). The executive mechanism of syncope is deactivation of RAS done with the active engagement of NTS through solitarioreticular pathway (SRT) and parabrachial nuclear complex (PBC). The biological purpose of syncope would be resetting of the NTS in case of an unbearable vegetative input, which is code for triggering the mechanism described. PMID- 15288356 TI - Postoperative delirium: the tryptophan dyregulation model. AB - A model previously presented by Uchida in this journal [Med. Hypotheses 53 (1997) 103] described a mechanism for postoperative delirium. It described an increased level of melatonin resulting in a central "serotonin shortage". This construct adequately explained only the hypoactive type of delirium. Recently it has been shown that a reduction in urinary metabolites of melatonin is associated with hyperactive delirium, whereas urinary metabolites were increased in the hypoactive variant. These findings suggest that this initial paradigm requires modification. We propose that both the agitation seen in hyperactive delirium, and the somnolence associated with the hypoactive form could be explained by a disturbance of central tryptophan homeostasis. It is postulated that intervention in the form of melatonin administration may restore tryptophan levels, and prevent delirium. PMID- 15288358 TI - Asthma medication may influence the psychological functioning of children. AB - Asthma is a global health problem with up to 15% of children suffering from the disease. It has been shown by various researchers that symptomatic asthmatic patients have increased levels of free serotonin in plasma when compared with asymptomatic patients. Thus, some researchers suggest that reducing the concentration of free serotonin in plasma might be useful in treating patients with asthma. Low levels of serotonin, has however, been linked to various psychological conditions like depression, oppositional defiant disorder, ADHD and even conduct disorder. Research has indicated that products like methylphenidate (also known by the brand names as e.g., Ritalin, Concerta, Metadate and others) and other stimulants used for these conditions, particularly ADHD, exert their paradoxical calming effects by boosting serotonin levels in the brain. Therefore, the hypothesis suggest that some children using asthma medication that lowers serotonin levels, might present with symptoms of depression ADHD, oppositional defiant disorder and even conduct disorder. They may be using asthma medication that lowers serotonin and additionally use methylphenidate that boosts serotonin levels for e.g., ADHD. The hypothesis therefore suggests that asthmatic children presenting with psychological complaints, be treated holistically and serotonin levels measured before coming to conclusions regarding their psychological functioning. PMID- 15288357 TI - Consideration of Rituximab for fibrodysplasia ossificans progressiva. AB - Fibrodysplasia ossificans progressiva (FOP) is a severe, progressive disease of the musculoskeletal system. Muscles, tendons and other connective tissues ossify after minor trauma, and patients often become encased in a second immobile skeleton. There is no known cure or treatment for FOP. It has been found that lymphocytes from FOP patients elaborate excess levels of bone morphogenic protein 4 (BMP-4). Given this, it has been suggested that allogenic bone marrow transplantation (BMT) possibly could be a cure for FOP, and drawn attention to a previously unappreciated case of an FOP patient who had successful BMT for aplastic anemia with apparent short- and medium-term arresting of the FOP disease process. However, BMT has non-trivial associated morbidity and mortality. Here, it is noted that if B cells are found to be the lymphocytes responsible for excess BMP-4 production in FOP, use of Rituximab, a monoclonal anti-CD20 antibody which effectively targets B cells, could be a less permanent and less risky treatment alternative for FOP. PMID- 15288359 TI - Supplementary taurine may stabilize atheromatous plaque by antagonizing the activation of metalloproteinases by hypochlorous acid. AB - The rupture of atherosclerotic plaque, responsible for triggering the majority of myocardial infarctions, presumably requires proteolysis of collagen fibers and other protein components of the intercellular matrix. This is achieved by activated matrix metalloproteases (MMPs) secreted by intimal macrophages and foam cells. MMPs are synthesized as inactive pro-enzymes in which coordinate binding of the thiol group of a key cysteine residue to the active-site zinc atom blocks proteolytic activity. Physiological activation of MMPs is mediated, in large measure, by phagocyte-derived hypochlorous acid (HOCL), which can oxidize the zinc-bound thiol to sulfinic acid, thus freeing the active-site zinc. HOCL also encourages proteolysis of ground substance by inactivating proteins such as TIMP 1 that are physiological inhibitors of MMPs. In vivo, the unrestrained oxidant activity of HOCL is opposed by taurine, which reacts spontaneously with HOCL to generate taurine chloramine, much more stable than HOCL. Taurine chloramine has less impact than HOCL on MMP activation, and does not impair the activity of TIMP 1. Since tissue levels of taurine can be boosted via supplementation, taurine may thus have potential for stabilizing plaque and thereby warding off infarction--an effect that should be reinforced by taurine's platelet-stabilizing activity. In light of recent epidemiological evidence that increased expression of myeloperoxidase - the enzyme which generates HOCL--is an important risk factor for coronary disease, supplemental taurine may indeed have broader utility for suppressing both the genesis and the rupture of atherosclerotic plaque. PMID- 15288360 TI - A taurine-supplemented vegan diet may blunt the contribution of neutrophil activation to acute coronary events. AB - Neutrophils are activated in the coronary circulation during acute coronary events (unstable angina and myocardial infarction), often prior to the onset of ischemic damage. Moreover, neutrophils infiltrate coronary plaque in these circumstances, and may contribute to the rupture or erosion of this plaque, triggering thrombosis. Activated neutrophils secrete proteolytic enzymes in latent forms which are activated by the hypochlorous acid (HOCl) generated by myeloperoxidase. These phenomena may help to explain why an elevated white cell count has been found to be an independent coronary risk factor. Low-fat vegan diets can decrease circulating leukocytes--neutrophils and monocytes--possibly owing to down-regulation of systemic IGF-I activity. Thus, a relative neutropenia may contribute to the coronary protection afforded by such diets. However, vegetarian diets are devoid of taurine - the physiological antagonist of HOCl- and tissue levels of this nutrient are relatively low in vegetarians. Taurine has anti-atherosclerotic activity in animal models, possibly reflecting a role for macrophage-derived myeloperoxidase in the atherogenic process. Taurine also has platelet-stabilizing and anti-hypertensive effects that presumably could reduce coronary risk. Thus, it is proposed that a taurine-supplemented low-fat vegan diet represents a rational strategy for diminishing the contribution of activated neutrophils to acute coronary events; moreover, such a regimen would work in a number of other complementary ways to promote cardiovascular health. Moderate alcohol consumption, the well-tolerated drug pentoxifylline, and 5-lipoxygenase inhibitors--zileuton, boswellic acids, fish oil--may also have potential in this regard. PMID- 15288361 TI - Sub-optimal taurine status may promote platelet hyperaggregability in vegetarians. AB - Although vegan diets typically have a very favorable effect on a range of vascular risk factors, several independent groups have reported that the platelets of vegetarians are more sensitive to pro-aggregatory agonists than are those of omnivores. In light of clear and convincing evidence that platelet function has an important impact on risk for thromboembolic events, it is important to clarify the basis of platelet hyperaggregability in vegetarians. A dietary deficit of long-chain omega-3 fatty acids is not likely to explain this phenomenon, since most omnivore diets do not include enough of these fats to discernibly influence platelet function. A more plausible possibility is that relatively poor taurine status--a function of the facts that plants are devoid of taurine and the human capacity for taurine synthesis is limited - is responsible. Plasma taurine levels are lower, and urinary taurine excretion is substantially lower, in vegetarians than in omnivores. Platelets are rich in taurine, which functions physiologically to dampen the calcium influx evoked by aggregating agonists--thereby down-regulating platelet aggregation. Supplemental intakes of taurine as low as 400 mg daily have been reported to markedly decrease the sensitivity of platelets to aggregating agonists ex vivo. Although the average daily intake of taurine from omnivore diets may be only about 150 mg, it is credible to speculate that a supplemental intake of this magnitude could normalize the platelet function of vegetarians in the long term; in any case, this thesis is readily testable clinically. Taurine is just one of a number of nutrients found almost solely in animal products--"carninutrients"--which are rational candidates for supplementation in vegans. PMID- 15288362 TI - Nitric oxide and prostacyclin mediate coronary shear force-induced alterations in cardiac electrophysiology. AB - Previous studies have shown that an increase in coronary flow or shear force enhances the oxygen consumption and contractility of myocardium (the "Gregg effect"). Recent studies have indicated that an increase in coronary shear force also affects atrioventricular conduction and ventricular repolarization. The mechanisms of "Gregg effect" and the shear force-induced alterations in cardiac electrophysiology are unclear. It has been proposed that the distension of the coronary arteries and the stretch of the surrounding myocytes may play a key role but this is not supported by the latest studies. Intracoronary perfusion increases coronary endothelial synthesis of nitric oxide and prostacyclin. Emerging evidence has indicated that both nitric oxide and prostacyclin prolong the action potential duration or effective refractory period in the in vitro and in vivo animal heart. We therefore hypothesized that shear force-induced alterations in cardiac function and electrophysiology are largely attributed to the enhanced endothelial release of nitric oxide and prostacyclin. PMID- 15288363 TI - Left cardiac sympathetic denervation as the first-line therapy for congenital long QT syndrome. AB - Congenital long QT syndrome (LQTS) is a cardiac electrophysiological disorder due to genetic mutations. Patients with LQTS, if untreated, have a high incidence of ventricular tachycardia and cardiac arrest. Adrenergic activities are believed to play a major role in triggering the onset of cardiac events. The current mainstay of therapy for LQTS is oral beta-blockers, which improves clinical symptoms and reduces the incidence of sudden cardiac death in approximately 70% of the patients. Left cardiac sympathetic denervation (LCSD) is an alternative therapy for patients who are resistant to beta-blockers. Its clinical use, however, has been hindered by the complexity of the procedure and complications after the surgery. Video-assisted thoracoscopic sympathectomy has been used to treat patients with palmar and axillary hyperhidrosis. We suggest that the use of the microinvasice thoracoscopic technique may greatly simplify the LSCD procedure, making it the first-line therapy for LQTS. PMID- 15288364 TI - Rheumatoid arthritis increases the risk of coronary heart disease via vascular endothelial injuries. AB - Patients with rheumatoid arthritis (RA) have an increased prevalence of coronary heart disease and a high cardiovascular mortality rate. The causes of increased coronary heart disease in RA patients are poorly understood. Conventional cardiovascular risk factors, such as inactivity, overweight or dyslipidemia may play a role, but they do not seem to be wholly responsible for the increased cardiovascular risk. RA is associated with a high incidence of inflammation and vascular endothelial injuries. Endothelial dysfunction is one of the key steps in the pathogenesis of atherosclerosis in non-RA patients. Therefore, we hypothesized that inflammation-induced vascular endothelial injuries may be responsible for the increased risk of coronary heart disease and high rates of cardiovascular mortality in patients with RA. PMID- 15288365 TI - Can local ventricular fibrillation interval predict ventricular refractory period in human hearts? AB - Assessment of the spatial dispersion of ventricular refractory periods has become an important part of electrophysiological study in both experimental and clinical settings, because inhomogeneity of ventricular refractoriness is associated with an increased risk of life-threatening ventricular arrhythmias. Previous animal studies in dog and sheep have demonstrated that local ventricular fibrillation (VF) intervals measured from the heart surface correlate well with the ventricular effective refractory periods measured from the same ventricular sites. We hypothesise that local VF intervals may also predict the ventricular refractory periods in human hearts, hence, can be used to assess the spatial dispersion of refractoriness and to predict the risk of ventricular arrhythmias. PMID- 15288366 TI - TAGE (toxic AGEs) hypothesis in various chronic diseases. AB - The advanced stage of the glycation process (one of the post-translational modifications of proteins) leads to the formation of advanced glycation end products (AGEs) and plays an important role in the pathogenesis of angiopathy in diabetic patients, in aging, and in neurodegenerative diseases. However, it is still not clear which AGEs subtypes play a pathogenetic role and which of several AGEs receptor mediate AGEs effects on cells. We have provided direct immunochemical evidence for the existence of six distinct AGEs structures (AGEs-1 to -6) within the AGEs-modified proteins and peptides that circulate in the serum of diabetic patients. Recently we demonstrated for the first time that glyceraldehyde-derived AGEs (AGEs-2) and glycolaldehyde-derived AGEs (AGE-3) have diverse biological activities on vascular wall cells, mesangial cells, Schwann cells, malignant melanoma cells and cortical neurons. We also demonstrated for the first time that acetaldehyde (AA)-derived AGEs (AA-AGE) have cytotoxic activity on cortical neurons and the AA-AGE epitope was detected in human brain of alcoholics. These results indicate that of the various types of AGEs structures that can form in vivo, the toxic AGEs (TAGE) structures (AGEs 2, 3, and AA-AGE), but not non-toxic AGEs (N-carboxymethyllysine, pentosidine, pyrraline etc.) are likely to play an important role in the pathophysiological processes associated with AGEs formation. PMID- 15288367 TI - Alternative routes for the formation of glyceraldehyde-derived AGEs (TAGE) in vivo. AB - The advanced stage of the glycation process (one of the post-translational modifications of proteins) leads to the formation of advanced glycation end products (AGEs) and plays an important role in the pathogenesis of angiopathy in diabetic patients, and in Alzheimer's disease (AD). Recently we have provided direct immunochemical evidence for the existence of six distinct AGEs structures, designated AGEs-1 to -6, within the AGEs-modified proteins and peptides that circulate in the serum of diabetic patients. We found for the first time that glyceraldehyde-derived AGEs (AGE-2), which comprise main structure of TAGE (toxic AGEs), in the serum of diabetic patients have diverse biological activities on vascular wall cells and cortical neurons. These results suggest a causal role for AGE-2 in the pathogenesis of diabetic complications and AD in vivo. In AD brains, AGE-2 epitope was mainly present in the cytosol of neurons in the hippocampus and para-hipocampal gyrus. We propose three pathways for the in vivo formation of AGE 2 precursor, glyceraldehyde, by: (i) glycolytic pathway, (ii) polyol pathway, and (iii) fructose metabolic pathway. Glyceraldehyde can be transported or can leak passively across the plasma membrane. It can react non-enzymatically with proteins to lead to accelerated formation of AGE-2 at both intracellular and extracellular region. PMID- 15288368 TI - Could oxytocin administration during labor contribute to autism and related behavioral disorders?--A look at the literature. AB - This literature review summarizes recent potential evidence, most of which is at the molecular/mechanistic level, in support of Hollander's hypothesis that excess oxytocin (OT), possibly through OT administration at birth, could contribute to the development of autistic spectrum disorders and related syndromes by proposed down regulation of the OT receptor (OTR). In this review, recent molecular evidence for OTR internalization by excess OT is related to OT's reported effects on animal social behavior, favoring social bondage, notably in sheep, voles, rats and especially mice. Adding indications for OT's capability of crossing the maternal placenta and OT's possibility of crossing an underdeveloped or stressed infantile blood brain barrier at birth, a causal connection between OT excess and behavioral disorders such as autism can be supported from a molecular perspective. Possible strategies such as a thorough statistical analysis of numerous birth records as well as molecular studies such as radiotracing using labeled OT are proposed to test this hypothesis. PMID- 15288369 TI - Anticardiolipin antibodies as markers of physiologic stress: a case study of idiopathic stroke in a young adult. AB - Although elevated anticardiolipin antibodies have been associated with idiopathic thromboembolic events, the disorder is not well understood. A case study of stroke in an otherwise healthy young adult spanning 6 years is presented. In this case, there was a correlation between increased serum concentrations of the antibodies, symptoms and periods of physiologic or emotional stress. Stress free intervals were accompanied by normal serum concentrations. The author proposes that anticardiolipin antibodies and their subtypes may be stress-induced markers and fluctuations in levels may correlate with periods of physiologic stress. PMID- 15288370 TI - Mania as a dysfunction of reentry: application of Edelman's and Tononi's hypothesis for consciousness in relation to a psychiatric disorder. AB - The concept of reentry as the most important element in a hypothesis for consciousness proposed by Edelman and Tononi is reviewed. Reentry, is a process of ongoing parallel and recursive signalling between separate neuronal groups along parallel reciprocally fibers that link these groups anatomically. Reentry alters the activity of the target areas it interconnects until a synchronous activity across these areas is created, this may be the direct biological mechanism of consciousness. The repetitive process of reentry may explain how the millisecond time scale of neural signalling is turned into the time scale of seconds characterizing our impression of the duration of a given content of consciousness. It is suggested that reentry may be faster in mania, and specifically that the repetitive recursive signalling is faster in mania, hereby allowing reentry to produce a conscious state, faster than usual. Faster reentry may on a molecular level be caused by faster propagation of nerve impulses, which may be in accordance with a number of hypotheses where mania is seen as a disorder of ionic conductance, nerve cell excitability, action potential firing, membrane abnormalities, and cortical instability. Also the antiepileptic drugs used to treat mania may point to reentry as a factor in this disorder. On a more integrated level faster reentry processes may explain several of the core symptoms of the manic state. Also the drug induced switch from depression to mania in bipolar patients may be explained by the concept of reentry. PMID- 15288371 TI - Negative socio-emotional resonance in schizophrenia: a functional magnetic resonance imaging hypothesis. AB - The aim of the present study is to use neuroscience theories about brain function (mirror-neurons MN) to draw inferences about the mechanisms supporting emotional resonance in two different groups of schizophrenia patients (with flat affect FA+ n = 13 and without flat affect FA- n = 11). We hypothesize that FA+ will not activate key brain areas involved in emotional processing. Conversely, FA- will have a functional mirror system for emotional resonance confirmed by activation of the prefrontal cortex and behavioral results. To test this hypothesis, we compared the two groups using blood oxygenation level-dependent (BOLD) functional magnetic resonance imaging (fMRI) displaying a passive visual task (44 negative IAPS pictures and 44 neutral pictures). A random-effects analysis, for schizophrenia patients FA-, revealed significant loci of activation in the left mesial prefrontal (MPFC), right orbitofrontal (OFC) and left anterior cingulate cortices (ACC). Correlational analyses carried out between self-report ratings of negative feelings and BOLD signal changes revealed the existence of positive correlation in the LACC, LMPFC and ROFC. Conversely, FA+ did not show significant activation in the prefrontal cortex. We propose that negative emotional resonance induced by passively viewing negative pictures may be a form of "mirroring" that grounds negative feelings via an experiential mechanism. Hence, it could be argued that FA- were able to 'feel' emotions through this resonance behavior. Conversely, we suggest that the dysfunction seen in the FA+ group is a failure or distortion in the development of the MN system. This could be due to genetic or other endogenous causes, which affected prefrontal cortex MN involved in emotional resonance. PMID- 15288372 TI - Metabolic rate determines haematopoietic stem cell self-renewal. AB - The number of haematopoietic stem cells (HSCs) per animal is conserved across species. This means the HSCs need to maintain hematopoiesis over a longer period in larger animals. This would result in the requirement of stem cell self renewal. At present the three existing models are the stochastic model, instructive model and the third more recently proposed is the chiaro-scuro model. It is a well known allometric law that metabolic rate scales to the three quarter power. Larger animals have a lower metabolic rate, compared to smaller animals. Here it is being hypothesized that metabolic rate determines haematopoietic stem cell self-renewal. At lower metabolic rate the stem cells commit for self renewal, where as at higher metabolic rate they become committed to different lineages. The present hypothesis can explain the salient features of the different models. Recent findings regarding stem cell self-renewal suggest an important role for Wnt proteins and their receptors known as frizzleds, which are an important component of cell signaling pathway. The role of cGMP in the Wnts action provides further justification for the present hypothesis as cGMP is intricately linked to metabolic rate. One can also explain the telomere homeostasis by the present hypothesis. One prediction of the present hypothesis is with reference to the limit of cell divisions known as Hayflick limit, here it is being suggested that this is the result of metabolic rate in laboratory conditions and there can be higher number of cell divisions in vivo if the metabolic rate is lower. PMID- 15288373 TI - Architectural organisation of neuronal activity-associated magnetic fields: a hypothesis for memory. AB - Despite intensive investigation into the mechanisms underlying the memory process, the physical bases for this superior cognitive function remain elusive. Neuronal activity-associated magnetic fields (NAAMFs) hypothesis of memory suggests that items of information are stored as three-dimensional bundles of magnetic fields associated to the complex but extremely organised cerebral cortex. The present paper proposes a plausible architectural organisation of neuronal activity-associated magnetic fields that may explain how information could be stored in the human cerebral cortex. Magnetic fields generated as consequence of neuronal minicolumns activation could modify the basal "electromagnetic status" of the closest astrocytes allowing codification and storage of information. PMID- 15288374 TI - Alveolar septal collapse in the transitional infant lung: a possible common mechanism in sudden unexpected death in infancy. AB - Sudden unexpected death in infancy (SUDI) is a category used to represent the largest single group of infant deaths. Although there are several theories, the cause of SUDI remains unknown and the mechanism of co-sleeping associated deaths are also undetermined. We investigate a possible biomechanical mechanism which may be common in SUDI and may provide an explanation for the association of the known risk factors for SUDI such as co-sleeping, prematurity, prone sleeping position, overwrapping, overheating and maternal smoking. The neonatal lung has few, if any, true septa but from about four weeks of age, a period of rapid alveolarisation commences. The developing alveolar walls (septae) have little fibre support against surface tension forces as they grow but are supported by a double layer of capillaries. Until the elastin/collagen supporting network is laid down these nascent septal walls are vulnerable to collapse against sac or duct walls during this transitional period. We hypothesise that such collapse will prevent one side of the septa, and the wall it overlays, from alveolar gas exchange and a functional left-right shunt is formed which may result in hypoxia. Furthermore, lung stretch receptors in bronchi running through or adjacent to collapsed regions will be activated, falsely signalling lung inflation to the brain stem with resultant respiratory inhibition, so precipitating further collapse. The process will continue until lung volume falls below residual capacity, when normal tidal breathing efforts will no longer result in significant air flow, even if stretch receptor signals have not produced complete apnoea. Large inspiratory efforts are then required to break the surface tension seal, which damages capillaries to produce petechial haemorrhages. Many epidemiological risk factors for SUDI could influence such a mechanism, leading to the proposal that Alveolar Septal Collapse in Infancy (ASCI) is a core mechanism via which these factors act. PMID- 15288375 TI - Polymorphism of estrogen metabolism genes and cataract. AB - Cataract is the leading cause of visual impairment in older adults in the world. Age-related lens opacities are common and are frequent causes of loss of vision. The incidence of cataract increases significantly with increasing age in women only. The onset coincides with estrogen deficiency that occurs after menopause. Hormone replacement therapy has proven beneficial to selected postmenopausal women. Estrogen effects on biological system are modulated via the estrogen receptors (ER) and/or estrogen metabolites. Although ER have been detected in ocular tissue, whether ER polymorphism is related to cataract is not known at present. The polymorphisms of estrogen metabolizing enzymes are also related to the serum concentration and activity of estrogen. Polymorphism such as cytochrome P450c17 (A2/A2), cytochrome P450c1A (vt/vt) will result in increased formation of catechol estrogen, while people with catechol-O-methyltransferase (COMT) polymorphism COMT (L/L) will have decreased metabolism of catechol estrogen and decreased level of methoxyestradiol. COMT was also involved in tamoxifen metabolism which may further decrease the activity of COMT in breast cancer patients treated with tamoxifen. It is known that a 4-7% increase in cataract was found in tamoxifen-treated breast cancer patients than non-user. The 7.0% COMT (L/L) genotype in general population corresponded well with the 4-7% of cataract formation in tamoxifen-treated breast cancer patients. Our hypothesis is that breast cancer patients with COMT (L/L) genotype may be at increased risk of cataract formation after tamoxifen treatment. PMID- 15288376 TI - Questions concerning low intensity reduction in lung compliance. AB - From the very early stages of reduction in lung compliance, the thoracic organs may be forcibly displaced, be subject to attraction or be prevented from fully carrying out their normal movement. By evaluating these changes on a chest radiogram we may assess, at least broadly, the existence of reduced compliance and to some extent its degree. Given that reduction of lung compliance can constitute the initial sign of establishment of a secondary neoplasm or other pathological condition in the lung, the potential of chest radiograms becomes wider both in the diagnostic sphere and in that of pathophysiological research. The more frequent displacement of the trachea to the right is, we believe, due to the greater overall mass of the right lung, so that in homogenous reduction of the lung compliance the total attraction to the right may be greater. The rising of the hemidiaphragms usually starts in their frontal region. This, we consider, is due to the different resistance to the reduction in lung compliance of the adjacent areas of the lung resulting from their different structure. The reduction of compliance in segments or subsegments adjacent to the hemidiaphragms produces local rising of them. When these segments are adjacent to the osseous thoracic wall they cause pleural thickening by activating the suction mechanism. The reduction in lung compliance may cause S-shaped distortion of the bronchovascular bundles of the lower lobes, displacement of the hili, blunting of the angle of the carina, curvature of the main bronchi and of the intrathoracic part of the trachea. All these changes can be seen on a chest radiogram. PMID- 15288377 TI - Disruption of hydrophobic stability of biomembranes is the earliest event in several clinical disorders. AB - Hydrophobic attractive force is the major force in maintaining the stability of biomembranes, yielding coordinated functionality to the embedded proteins that they contain. This force between the composite linear hydrocarbons of the biomembranes is a function of their length and their mutual parallel distance from each other, and is extremely sensitive to this distance. Extracellular, natural linear hydrocarbons of certain length and shape can intercalate into lipid matrix of the biomembranes, reducing their innate hydrophobic net strength in a concentration-dependent manner, making them loose, leaky, and thus gaining the credence of stimulus-generating agents. In physiological circulatory concentration, these molecules may have a role for the maintenance of cellular homeostasis. However, in stagnating physiological excess, these same agents can become acutely or chronically stimulating and, therefore, disease-precipitating. Such situations do exist in the clinical disorders of acne, atherosclerosis, acute pancreatitis, diabetes, diabetic retinopathy, homocysteinemea, and stress. A systematic approach, beginning with surface film studies with the suspect linear hydrocarbons, can be followed up with in vitro and in vivo studies. This should substantiate or negate the view presented here. Isolated information, along these lines, already exist in literature. The example of acne is a suitable starting point to elaborate this view, for sebaceous gland of the human pilosebaceous unit (PSU) contains all the exemplary, stimulus-inducing linear hydrocarbons to generate surface-reaction on the pilosebaceous ductal surface. PMID- 15288378 TI - High altitude sickness. Is acute cortisol deficiency involved in its pathophysiology? AB - High altitude illness (HAI) affects 42% of individuals climbing above 3000 m. The pathophysiology of HAI, including water retention remains unclear. Although decreased nitric oxide (NO) production is implicated in the pathophysiology, a recent study reported increased NO in breathes of high altitude inhabitants, apparently produced to combat the high altitude hypoxia. NO binds heme generally and impairs cytochrome P450 steroidogenic enzymes. A consequence of increased NO production may be decreased steroidogenesis. An acute cortisol deficiency may thus be the reason for water retention and oedema, and explains why dexamethasone is effective in treating some aspects of HAI. PMID- 15288379 TI - A hypothesis on the role of insulin-like growth factor I in testicular germ cell tumours. AB - It has already been established that the growth effects of growth hormone (GH) are mediated through insulin-like growth factor I (IGF-I). Recent studies demonstrated a relationship between IGF-I levels and various types of cancer, namely colon, prostate, breast, brain and lung cancer. In addition, many experimental observations documented a participation of the IGF-I system in tumourigenesis through enhanced cell proliferation rate, anti-apoptotic functions and stimulation of neovascularization. With the present known biological mechanisms, implicated in the pathogenesis of testicular germ cell tumours (GCT), it is difficult to interpret the consistently increasing incidence of this tumour over the last decades. On the other hand, unpublished data of our department are in accordance with previous published studies, suggesting that GCT may be positively associated with body height. Scattered publications report development of GCT secondary to acromegaly or long-term GH replacement therapy. Thus, it is possible that the IGF-I system may be implicated in this pathogenesis, thereby predisposing to an increased risk of testicular GCTs. If IGF-I and IGFBP-3 are found to correlate with a high incidence of testicular GCT, they might be useful surrogate markers for diagnosis and surveillance of tumour growth, and an early screening method to identify an increased risk of this type of cancer in the first degree young male relatives of these patients. PMID- 15288380 TI - Replicative homeostasis: a mechanism of viral persistence. AB - Acute viral infection is characterised by high-level replication before prompt decline of viraemia and, commonly, viral clearance. This kinetic pattern is generally held to be due to immune control. However, infection with some viruses, notably hepatitis C (HCV), hepatitis B (HBV) and the human immunodeficiency virus (HIV), often results in chronic stable low-level spontaneously fluctuating viraemia, kinetics that are difficult to rationalize on this basis. The persistence of HCV, an RNA virus, is especially problematic and its stability, occurring despite rapid, genomic mutation is highly paradoxical. This paper outlines the hypothesis, and evidence, that viruses autoregulate replication and mutation and describes a mechanism--replicative homeostasis--explaining viral stability. Replicative homeostasis results in stable, but reactive, replicative equilibria that drive quasispecies expansion and immune escape and explain all observed viral behaviours and host responses. This paradigm implies new approaches to antiviral therapy and is broadly relevant to modulation of gene expression. PMID- 15288381 TI - Association of Helicobacter pylori with central serous chorioretinopathy: hypotheses regarding pathogenesis. AB - Central serous chorioretinopathy (CSC) is a serous macular detachment that usually affects young people and leads fortunately to a spontaneous resolution and a good visual prognosis in most patients. Nevertheless, although in a small percentage of subjects only, it may also develop a chronic or progressive disease with widespread decompensation of the retinal pigment epithelium (RPE) and severe vision loss. The aetiopathogenesis of the disease is still not completely understood and no effective treatment is available at this time. However, an interesting association has been recently highlighted between CSC and the Helicobacter pylori infection. In particular, in a first case report recurrences of the disease were always associated with HP-positivity whereas improvements of both retinal findings and visual acuity were significantly correlated with a successful eradication of the bacterium using the conventional antimicrobial triple-therapy. In a second study, the prevalence of HP infection was found to be significantly higher in CSC-affected subjects compared to age- and sex-matched controls from the same country. Much speculation surrounds the role potentially played by HP in determining CSC. In particular, CSC seems not to be more a merely RPE disease but the final result of a general involvement of the choroidal microcirculation. In fact, several vascular abnormalities, such as localized vasoconstriction and impaired fibrinolysis, have been demonstrated during CSC whose "end-points" might be a focal occlusion of the choriocapillaries with decreased foveal choroidal blood flow, secondary RPE defects and serous macular detachment. Moreover, a HP-dependent immune mechanism, based on a "molecular mimicry" between pathogenic antigens expressed on the bacterium and homologous host proteins (e.g., those of the endothelial vascular wall), might also be involved in the pathophysiology of CSC. In this case, a genetically determined susceptibility of the subject could be an important and limiting factor. Although further multicenter, randomized, case-control trials are necessary to confirm the role potentially played by the HP infection in the pathogenesis of CSC, if this hypothesis would be confirmed in the near future, a novel antimicrobial approach to the disease might be possible waiting for a successful vaccine therapy that will surely stimulate the scientific interest of many authors. PMID- 15288382 TI - Pedophilia: neuropsychological evidence encouraging a brain network perspective. AB - Although the vast majority of current pathogenetic theories support a neurobiological understanding of psychiatric disorders, the brain functional correlates of pedophilia are largely unknown. Based on prior behavior genetics research on human sexual orientation and phenomenology as well as the phenotypical intersection of pedophilia with other psychiatric spectrum disorders, we hypothesize the involvement of striato-thalamo-cortical processing loops in the formation of pedophilic urges and behaviors. Data from a current neuropsychological pilot study in four pedophiles encourage our brain functional perspective. As deduced from the network model, all four patients exhibited pronounced and circumscribed deficits in cognitive domains mediated by striato thalamically controlled areas of the frontal cortex. All patients were especially impaired in neuropsychological functions associated with the prefrontal and motor processing loops (e.g., response inhibition, working memory and cognitive flexibility), with a performance level located up to five standard deviations below the normative data. Contrary to this, neuropsychological performances in cognitive domains without a comparable high frontal loading were in all participants unobtrusive. In future, studying gene by environment interactions in combination with functional neuroimaging and neuropsychological assessment is promising to elucidate the pathophysiological relationship of psychiatric disorders that are characterized by inadequate urges and poor behavioral inhibition. PMID- 15288383 TI - Maldaptation of the link between inflammation and bone turnover may be a key determinant of osteoporosis. AB - Currently the etiology of osteoporosis is attributed to various endocrine, metabolic, and mechanical factors. We hypothesize that many cases of osteoporosis are also partially attributable to a maladaptation of the link between inflammation and bone turnover. We explore the spatial and temporal link between inflammation and osteoporosis in conditions such as aging, menopause, reflex sympathetic dystrophy, HIV, pregnancy, transplantation, and steroid administration. While nutritional and mechanical factors clearly play a role in many of these situations, the spatial and temporal concordance of osteoporosis and inflammation is buttressed by emerging molecular evidence. Modern bone biology of humans may reflect dual functional legacies of mineral storage and structural support. Osteoporosis may result from disequilibrium between structural demand for key minerals and their biologic demand during maladaptive states of inflammation. PMID- 15288384 TI - Narcotic antagonists in drug dependence: pilot study showing enhancement of compliance with SYN-10, amino-acid precursors and enkephalinase inhibition therapy. AB - We decided to test the hypothesis that possibly by combining a narcotic antagonist and amino-acid therapy consisting of an enkephalinase inhibitor (D phenylalanine) and neurotransmitter precursors (L-amino- acids) to promote neuronal dopamine release might enhance compliance in methadone patients rapidly detoxified with the narcotic antagonist Trexan (Dupont, Delaware). In this regard, Thanos et al. [J. Neurochem. 78 (2001) 1094] and associates found increases in the dopamine D2 receptors (DRD2) via adenoviral vector delivery of the DRD2 gene into the nucleus accumbens, significantly reduced both ethanol preference (43%) and alcohol intake (64%) of ethanol preferring rats, which recovered as the DRD2, returned to baseline levels. This DRD2 overexpression similarly produced significant reductions in ethanol non-preferring rats, in both alcohol preference (16%) and alcohol intake (75%). This work further suggests that high levels of DRD2 may be protective against alcohol abuse [JAMA 263 (1990) 2055; Arch, Gen. Psychiatr. 48 (1991) 648]. The DRD2 A1 allele has also been shown to associate with heroin addicts in a number of studies. In addition, other dopaminergic receptor gene polymorphisms have also associated with opioid dependence. For example, Kotler et al. [Mol. Phychiatr. 3 (1997) 251] showed that the 7 repeat allele of the DRD4 receptor is significantly overpresented in the opioid-dependent cohort and confers a relative risk of 2.46. This has been confirmed by Li et al. [Mol. Psychiatry 2 (1997) 413] for both the 5 and 7 repeat alleles in Han Chinese case control sample of heroin addicts. Similarly Duaux et al. [Mol. Psychiatry 3 (1998) 333] in French Heroin addicts, found a significant association with homozygotes alleles of the DRD3-Bal 1. A study from NIAAA, provided evidence which strongly suggests that DRD2 is a susceptibility gene for substance abusers across multiple populations (2003). Moreover, there are a number of studies utilizing amino-acid and enkephalinase inhibition therapy showing reduction of alcohol, opiate, cocaine and sugar craving behavior in human trials (see Table 1). Over the last decade, a new rapid method to detoxify either methadone or heroin addicts utilizing Trexan sparked interest in many treatment centers throughout the United States, Canada, as well as many countries on a worldwide basis. In using the combination of Trexan and amino-acids, results were dramatic in terms of significantly enhancing compliance to continue taking Trexan. The average number of days of compliance calculated on 1000 patients, without amino-acid therapy, using this rapid detoxification method is only 37 days. In contrast, the 12 subjects tested, receiving both the Trexan and amino acid therapy was relapse-free or reported taking the combination for an average of 262 days (p < 0.0001F). Thus coupling amino-acid therapy and enkephalinase inhibition while blocking the delta-receptors with a pure narcotic antagonist may be quite promising as a novel method to induce rapid detox in chronic methadone patients. This may also have important ramifications in the treatment of both opiate and alcohol-dependent individuals, especially as a relapse prevention tool. It may also be interesting too further test this hypothesis with the sublingual combination of the partial opiate mu receptor agonist buprenorphrine. PMID- 15288385 TI - Idiopathic intracranial hypertension: priapism of the brain? AB - It has long been recognized that a clinical syndrome similar to idiopathic intracranial hypertension can occur secondary to venous sinus obstruction with the elevation in sinus pressure causing a reversal in the pressure gradient across the arachnoid granulations and therefore elevated CSF pressure. There remains, however, a group of predominantly obese female patients with elevated CSF pressures who have either no apparent abnormality of venous outflow or a tapering, apparently extrinsic compression, of their dominant transverse sinuses on catheter venography. This suggests that venous collapse may be associated in some way with the cause of the elevated pressure but clearly something else must be initiating the pressure rise or a circular argument ensues. Elevated CSF pressure compresses the veins, which then elevates CSF pressure. However, collapse of the venous sinuses secondary to the elevated CSF pressure once initiated may exacerbate the condition. It has been suggested that the initiating event leading to the elevated pressures in the idiopathic group of patients may be caused by cerebral hyperaemia and cerebral dysautoregulation. Priapism is a condition of pathological elevation of venous pressure of the male genitalia in which there are two forms: (1) venous out flow obstruction and (2) hyperaemia due to a to loss of regulation of blood flow and secondary venous out flow compression. Review of the literature suggests the pathophysiology of idiopathic intracranial hypertension may be analogous to that of priapism. PMID- 15288386 TI - Effects of very low-fat diets on anginal symptoms. PMID- 15288387 TI - A possible natural antibacterial compound in the human metabolism. PMID- 15288388 TI - Antibiotic therapy may induce cancers in the colon and breasts through a mechanism involving bile acids and colonic bacteria. PMID- 15288389 TI - The communal coping model and interpersonal context: problems or process? PMID- 15288390 TI - Painful beginnings. PMID- 15288391 TI - Active despite pain: the putative role of stop-rules and current mood. PMID- 15288392 TI - HIV-1 gp120 stimulates proinflammatory cytokine-mediated pain facilitation via activation of nitric oxide synthase-I (nNOS). AB - It has become clear that spinal cord glia (microglia and astrocytes) importantly contribute to the creation of exaggerated pain responses. One model used to study this is peri-spinal (intrathecal, i.t.) administration of gp120, an envelope protein of HIV-1 known to activate glia. Previous studies demonstrated that i.t. gp120 produces pain facilitation via the release of glial proinflammatory cytokines. The present series of studies tested whether spinal nitric oxide (NO) contributes to i.t. gp120-induced mechanical allodynia and, if so, what effect NO has on spinal proinflammatory cytokines. gp120 stimulation of acutely isolated lumbar dorsal spinal cords released NO as well as proinflammatory cytokines (tumor necrosis factor-alpha, interleukin-1beta (IL1), interleukin-6 (IL6)), thus identifying NO as a candidate mediator of gp120-induced behavioral effects. Behaviorally, identical effects were observed when gp120-induced mechanical allodynia was challenged by i.t. pre-treatment with either a broad-spectrum nitric oxide synthase (NOS) inhibitor (L-NAME) or 7-NINA, a selective inhibitor of NOS type-I (nNOS). Both abolished gp120-induced mechanical allodynia. While the literature pre-dominantly documents that proinflammatory cytokines stimulate the production of NO rather than the reverse, here we show that gp120-induced NO increases proinflammatory cytokine mRNA levels (RT-PCR) and both protein expression and protein release (serial ELISA). Furthermore, gp120 increases mRNA for IL1 converting enzyme and matrix metalloproteinase-9, enzymes responsible for activation and release of proinflammatory cytokines. PMID- 15288393 TI - Pain in primary erythromelalgia--a neuropathic component? AB - Erythromelalgia is a condition characterized by attacks of red, hot, painful extremities with relief of symptoms by cooling and aggravation by warmth. Although the main emphasis has been on pathophysiological mechanisms related to circulatory changes, recent reports have focused on an involvement of efferent small nerve fibers indicating a neuropathic component. Since the symptoms resemble those described in neuropathic pain, we wanted to investigate the possible affection of afferent nerve fibers. Twenty-five patients with primary erythromelalgia were examined by neurological testing, neurography and quantitative sensory testing. Thresholds for heat, cold, heat-pain and cold-pain detection were compared with those of a group of 29 healthy controls. The patients had significantly higher median heat (39.5 (36.1-40.8) and cold (29.3 (27.1-30.8)-detection thresholds at the dorsal aspects of their feet compared to the controls (37.0 (35.4-37.7) and 31.2 (30.3-31.5) respectively). These findings show an impaired small fiber function inside or close to the symptomatic area in this group of erythromelalgia patients. Seven patients had brush-evoked allodynia and fourteen had punctate hyperalgesia inside or close to the symptomatic areas in their feet. When comparing the individual results, there is a tendency to clustering of patients in two separate groups; reduced small fiber input/no hyperalgesia and normal thermal thresholds/hyperalgesia. Our results showing an affection of afferent small nerve fibers together with the nature of the symptoms, suggest that the pain experienced by erythromelalgia patients could have a neuropathic component. PMID- 15288394 TI - Effects of spouse-assisted coping skills training and exercise training in patients with osteoarthritic knee pain: a randomized controlled study. AB - This study tested the separate and combined effects of spouse-assisted pain coping skills training (SA-CST) and exercise training (ET) in a sample of patients having persistent osteoarthritic knee pain. Seventy-two married osteoarthritis (OA) patients with persistent knee pain and their spouses were randomly assigned to: SA-CST alone, SA-CST plus ET, ET alone, or standard care (SC). Patients in SA-CST alone, together with their spouses, attended 12 weekly, 2-h group sessions for training in pain coping and couples skills. Patients in SA CST + ET received spouse-assisted coping skills training and attended 12-weeks supervised ET. Patients in the ET alone condition received just an exercise program. Data analyses revealed: (1) physical fitness and strength: the SA-CST + ET and ET alone groups had significant improvements in physical fitness compared to SA-CST alone and patients in SA-CST + ET and ET alone had significant improvements in leg flexion and extension compared to SA-CST alone and SC, (2) pain coping: patients in SA-CST + ET and SA-CST alone groups had significant improvements in coping attempts compared to ET alone or SC and spouses in SA-CST + ET rated their partners as showing significant improvements in coping attempts compared to ET alone or SC, and (3) self-efficacy: patients in SA-CST + ET reported significant improvements in self-efficacy and their spouses rated them as showing significant improvements in self-efficacy compared to ET alone or SC. Patients receiving SA-CST + ET who showed increased self-efficacy were more likely to have improvements in psychological disability. An intervention that combines spouse-assisted coping skills training and exercise training can improve physical fitness, strength, pain coping, and self-efficacy in patients suffering from pain due to osteoarthritis. PMID- 15288395 TI - Prognosis of non-specific musculoskeletal pain in preadolescents: a prospective 4 year follow-up study till adolescence. AB - Musculoskeletal pain is common in children but studies on the outcome and predictive factors for persistence/recurrence of these symptoms are scarce. A baseline cross-sectional survey of 1,756 schoolchildren (mean age 10.8) identified 564 (32.1%) children with musculoskeletal pain. At baseline, these children were evaluated using a structured questionnaire and examined for hypermobility and physical fitness. The children were re-evaluated after one year, and four years (at adolescence) using the same pain questionnaire. At 1 year follow-up, 53.8% (95% CI 48.8-58.8) of the children reported pain persistence (persistent preadolescent musculoskeletal pain). At 4-year follow-up, 63.5% (95% CI 58.7-68.1) of them had musculoskeletal pain. Neck was the site with most persistent/recurrent musculoskeletal pain. Those with persistent preadolescent musculoskeletal pain had approximately three times higher risk of pain recurrence (OR=2.90 [95% CI 1.9-4.4]). In the univariate analysis, female gender, older age group (11+), hypermobility, co-existence of psychosomatic symptoms, having high disability index, and reporting combined musculoskeletal pain at baseline predicted pain recurrence at adolescence. In the multivariate analysis, age, headache, hypermobility and having combined musculoskeletal pain were found as independent predictors. Statistically significant sex interactions were found for age, depressive feelings, waking up during nights and hypermobility. More psychosomatic symptoms predicted pain recurrence in girls than in boys, and hypermobility was a strong predictor in females only. Musculoskeletal pain in preadolescents is not a self-limiting phenomenon and more studies are still warranted to explore its determinants aiming to improve the long-term outcome of these symptoms. PMID- 15288396 TI - Hyperalgesia in an animal model of multiple sclerosis. AB - Many individuals with multiple sclerosis (MS) experience clinically significant pain, yet the underlying neural mechanisms for MS pain are not understood. Experimental autoimmune encephalomyelitis (EAE) is a well-studied disease in rodents that mimics many clinical and pathological features of MS, including central nervous system inflammation and demyelination. To determine whether EAE is an appropriate model for MS-related pain, nociceptive responses in both male and female SJL mice were measured before and after immunization with myelin proteolipid protein peptide 139-151 (PLP(139-151)) in complete Freund's adjuvant to induce 'active' EAE. To determine if changes in nociception were due to direct effects of encephalitogenic T cells, nociceptive responses in female SJL mice were measured following the transfer of activated, PLP(139-151) specific T cells to induce 'passive' EAE. Both forepaw and tail withdrawal latencies to a radiant heat stimulus were measured. In both active and passive EAE, there was an initial increase in tail withdrawal latency (hypoalgesia) that peaked several days prior to the peak in motor deficits during the acute disease phase. During the chronic disease phase, tail withdrawal latencies decreased and were significantly faster than control latencies for up to 38 days post-immunization. This hyperalgesia was seen in both sexes and in both active and passive EAE models. Forepaw withdrawal latencies remained within 1-2 s of baseline latencies for the entire testing period, indicating that the hypoalgesia and hyperalgesia were most pronounced in clinically affected body regions. These results suggest that both active and passive EAE are useful models of MS-related pain. PMID- 15288397 TI - Gender differences in pain, coping, and mood in individuals having osteoarthritic knee pain: a within-day analysis. AB - This study examined gender differences in prospective within-day assessments of pain, pain coping, and mood in men and women having OA, and analyzed gender differences in dynamic relations between pain, mood, and pain coping. A sample of 64 women and 36 men diagnosed as having pain due to osteoarthritis of the knee(s) rated their pain, pain coping, and mood two times each day (once in the afternoon and once in the evening) for 30 days using a booklet format. Two gender differences were found in between person-analyses: women used more problem focused coping than men, and women who catastrophized were less likely than men to report negative mood. Several within-day and across-day gender differences were noted. First, women were much more likely to show a significant increase in pain over the day. Second, men were more likely than women to experience an increase in coping efficacy over the day. Third, men were more likely than women to use emotion-focused coping when their mood was more negative. Finally, men were more likely than women to experience an increase in negative mood and a decrease in positive mood in the morning after an evening of increased pain. Taken together, these findings underscore the importance of obtaining multiple daily assessments when studying gender differences in the pain experience. PMID- 15288398 TI - Role of TNF-alpha in sensitization of nociceptive dorsal horn neurons induced by application of nucleus pulposus to L5 dorsal root ganglion in rats. AB - Herniation of the nucleus pulposus (NP) from lumbar intervertebral discs commonly results in radiculopathic pain and paresthesia (sciatica). While traditionally considered the result of mechanical compression of the dorsal root ganglion (DRG) and/or spinal nerve root, recent studies implicate pro-inflammatory mediators released from or evoked by NP, a possibility that was presently investigated. Single-unit recordings were made from L5 wide dynamic range dorsal horn neurons in pentobarbital-anesthetized rats. Autologous NP was harvested from a coccygeal disc and placed onto the exposed L5 DRG. A control group had subcutaneous adipose tissue or saline placed similarly. To test involvement of tumor necrosis factor alpha (TNF-alpha), a third group received autologous NP plus local soluble TNF alpha receptor type 1 (0.013 microg) which binds TNF-alpha to prevent its action. In each group, neuronal responses to graded heat (38-50 degrees C) and mechanical (von Frey filaments 4-76 g) stimuli were recorded prior to and at three successive hourly intervals following each treatment. Responses to noxious heat and mechanical stimuli were significantly enhanced 1 h post-NP and remained elevated thereafter. Thermally and mechanically evoked responses were not significantly affected in control rats or those treated with NP + soluble TNF alpha receptor type 1. These results indicate that sensitization of nociceptive spinal neuronal responses develops quickly following exposure of the DRG to NP, and that TNF-alpha is involved. This electrophysiological model of herniated NP may prove useful in further characterizing the role of inflammatory mediators in hyperalgesia and allodynia resulting from lumbar disc herniation. PMID- 15288399 TI - Characterization of basal and re-inflammation-associated long-term alteration in pain responsivity following short-lasting neonatal local inflammatory insult. AB - Recently, several studies have suggested that neonatal noxious insult could alter future responses to painful stimuli. However, the manifestations, mechanisms, and even developmental nature of these alterations remain a matter of controversy. In part, this is due to the lack of detailed information on the neonatal sensitive period(s) during which noxious stimulation influences future nociception, and the time-course and distribution of the resultant abnormalities. The present paper describes these parameters in a rat model of short-lasting ( approximately 24 h) neonatal local inflammation of a hindpaw produced by injection of 0.25% carrageenan (1 microl/g). Examinations of paw withdrawal responses to thermal and mechanical stimulations in adult animals, which as neonates were subjected to this insult, showed that the previously-reported long-term hypoalgesia and hyperalgesia are not mutually exclusive outcomes of early noxious experience. Long-term hypoalgesia was apparent at the basal conditions and was equally strong in the previously injured and uninjured paws, which suggests a globally-driven deficit. In contrast, long-term excessive hyperalgesia had the strongest manifestation in the neonatally-injured paw after re-inflammation, indicating significant segmental involvement in its generation. The differences between mechanisms underlying the observed hypoalgesia and hyperalgesia are further underscored by the finding that, while the former is detectable only after animals reach the second month of life, the latter is elicitable immediately upon cessation of the initial neonatal inflammation. Nevertheless, we detected a significant overlap in the neonatal sensitive periods for generation of these effects (both occurring within the first postnatal week). Also, neither the basal hypoalgesia nor excessive re-inflammation-associated hyperalgesia subsided with age and were detectable in 120-125-day-old rats. These finding provide a framework within which the entire complex of long-term effects of early noxious experience can be understood and examined. PMID- 15288400 TI - Pain catastrophizing and interpersonal problems: a circumplex analysis of the communal coping model. AB - Using the circumplex model of interpersonal behavior [Handbook of research methods in clinical psychology, 1982], this study tested the communal coping model of catastrophizing (CCM) in a large (N = 179) sample of patients with irritable bowel syndrome (IBS), a common, benign chronic pain disorder associated with significant painful extraintestinal comorbidity (e.g. headache, low back pain). Patients completed the Coping Strategies Questionnaire, the Brief Symptom Inventory, and the Inventory of Interpersonal Problems. The main findings were: (1) individuals who reported higher levels of catastrophizing described greater interpersonal problems; (2) the interpersonal problems described by catastrophizers fell within the friendly and friendly submissive quadrants of the circumplex supporting the notion that they have an interpersonal style demanding support and care-taking [Pain 103 (2003) 151]; (3) the pain coping behavior most strongly associated with interpersonal problems was catastrophizing; and (4) the relationship between interpersonal problems and catastrophizing remained after removing the influence of general symptomatic distress (i.e. an overall tendency to complain of psychological problems in general). In general, data provide evidence supporting the interpersonal distinctiveness of pain catastrophizing as postulated by the CCM. Advantages of a circumplex model and of interpersonal theory for understanding and testing the CCM are discussed. PMID- 15288401 TI - Rapid deterioration of pain sensory-discriminative information in short-term memory. AB - The assessment of pain and analgesic efficacy sometimes relies on the retrospective evaluation of pain felt in the immediate, recent or distant past, yet we have a very limited understanding of the processes involved in the encoding, maintenance and intentional retrieval of pain. We examine the properties of the short-term memory of thermal and pain sensation intensity with a delayed-discrimination task using pairs of heat pain, warm and cool stimulation in healthy volunteers. Performance decreased as a function of the inter-stimulus interval (ISI), indicating a robust deterioration of sensory information over the test period of 4-14 s. As expected, performance also decreased with smaller temperature differences (Delta-T) and shorter stimulus durations (6-2 s). The relation between performance and Delta-T was adequately described by a power function, the exponent of which increased linearly with longer ISI. Importantly, performance declined steadily with increasing ISI (from 6 to 14 s)--but only for pairs of heat pain stimuli that were relatively difficult to discriminate (Delta T < or = 1.0 degree C; perceptual difference < or = 32/100 pain rating units) while no deterioration in performance was observed for the largest temperature difference tested (Delta T = 1.5 degrees C; perceptual difference of 50 units). These results are consistent with the possibility that short-term memory for pain and temperature sensation intensity relies on a transient analog representation that is quickly degraded and transformed into a more resistant but less precise categorical format. This implies that retrospective pain ratings obtained even after very short delays may be rather inaccurate but relatively reliable. PMID- 15288402 TI - Somatostatin modulates the transient receptor potential vanilloid 1 (TRPV1) ion channel. AB - Activation of peripheral somatostatin receptors (SSTRs) inhibits sensitization of nociceptors, thus having a short term or phasic effect [Pain 90 (2001) 233] as well as maintaining a tonic inhibitory control over nociceptors [J Neurosci 21 (2001) 4042]. The present study provides several lines of evidence that an important mechanism underlying SSTR modulation of nociceptors is regulation of the transient receptor potential vanilloid 1 ion channel (TRPV1, formerly the VR1 receptor). Double labeling of L5 dorsal root ganglion cells demonstrates that approximately 60% of SSTR2a-labeled cells are positive for TRPV1. Conversely, approximately 33% of TRPV1-labeled cells are positive for SSTR2a. In vivo behavioral studies demonstrate that intraplantar injection of 20.0 but not 2.0 microM octreotide (OCT, SSTR agonist) significantly reduces capsaicin (CAP, a ligand for TRPV1) -induced flinching and lifting/licking behaviors. This occurs through local activation of SSTRs in the injected hindpaw and is reversed following co-application of the SSTR antagonist cyclo-somatostatin (c-SOM). In vitro studies using a skin-nerve preparation demonstrate that activation of peripheral SSTRs on nociceptors with 20.0 microM OCT significantly reduces CAP induced activity and can prevent CAP-induced desensitization. Furthermore, blockade of peripheral SSTRs with c-SOM dramatically enhances CAP-induced behaviors and nociceptor activity, demonstrating SSTR-induced tonic inhibitory modulation of TRPV1. Finally, TRPV1 does not appear to be under tonic opioid receptor control since the opioid antagonist naloxone does not change CAP-induced excitation and does not effect OCT-induced inhibition of CAP responses. These data strongly suggest that SSTRs modulate nociceptors through phasic and tonic regulation of peripheral TRPV1 receptors. PMID- 15288403 TI - Pregabalin for the treatment of painful diabetic peripheral neuropathy: a double blind, placebo-controlled trial. AB - A randomized, double-blind, placebo-controlled, parallel-group, multicenter, 8 week trial (with subsequent open-label phase) evaluated the effectiveness of pregabalin in alleviating pain associated with diabetic peripheral neuropathy (DPN). For enrollment, patients must have had at baseline: 1- to 5-year history of DPN pain; pain score > or =40 mm (Short-Form McGill Pain Questionnaire [SF MPQ] visual analogue scale); average daily pain score of > or =4 (11-point numerical pain rating scale [0 = no pain, 10 = worst possible pain]). One hundred forty-six (146) patients were randomized to receive placebo (n = 70) or pregabalin 300 mg/day (n = 76). Primary efficacy measure was endpoint mean pain score from daily patient diaries (11-point numerical pain rating scale). Secondary measures included SF-MPQ scores; sleep interference scores; Patient and Clinical Global Impression of Change (PGIC and CGIC); Short Form-36 (SF-36) Health Survey scores; and Profile of Mood States (POMS) scores. Safety assessment included incidence and intensity of adverse events, physical and neurological examinations, and laboratory evaluations. Pregabalin produced significant improvements versus placebo for mean pain scores (P < 0.0001); mean sleep interference scores SF-36 Bodily Pain subscale (P < 0.0001); total SF-MPQ score (P < 0.01); SF-36 Bodily Pain subscale (P < 0.03); PGIC (P = 0.001); and Total Mood Disturbance and Tension-Anxiety components of POMS (P < 0.03). Pain relief and improved sleep began during week 1 and remained significant throughout the study (P < 0.01). Pregabalin was well tolerated despite a greater incidence of dizziness and somnolence than placebo. Most adverse events were mild to moderate and did not result in withdrawal. Pregabalin was safe and effective in decreasing pain associated with DPN, and also improved mood, sleep disturbance, and quality of life. PMID- 15288404 TI - Prognostic factors for neck pain in general practice. AB - Prognostic studies on neck pain are scarce and are typically restricted to short term follow-up only. In this prospective cohort study, indicators of short- and long-term outcomes of neck pain were identified that can easily be measured in general practice. Patients between 18 and 70 years of age, suffering for at least 2 weeks from neck pain were recruited by 42 general practitioners (GPs). Perceived recovery, pain intensity and neck dysfunction after 7 and 52 weeks were considered as outcome measures. Indicators of prognosis were identified by means of logistic regression analyses (perceived recovery) and linear regression analyses (pain intensity and neck dysfunction). In total, 183 patients were included. After 1 year, 63% had recovered. The prognostic models showed differences between short- and long-term indicators. At the short term, besides the baseline values of the respective outcome measurements, only older age (> or =40) and concomitant low back pain and headache were associated with poor outcome. At the long term, in addition to age and concomitant low back pain, previous trauma, a long duration of neck pain, stable neck pain during the 2 weeks prior to baseline measurement, and previous neck pain predicted poor prognosis. The predictive power of the models was weak: the explained variance (R(2)) varied from 24 to 36%. Patient history and physical examination give GPs little handholds to predict the prognosis for patients with sub-acute and chronic neck pain. A few indicators of a less favourable prognosis of neck pain were identified, of which older age and concomitant low back pain was the most consistent. PMID- 15288405 TI - Experimental pain perception remains equally active over all sleep stages. AB - The literature on sensory perception during sleep suggests that light sleep (Stage 2) is more responsive to external sensory stimulation (e.g. sound, electrical shock) than deep sleep (Stages 3 and 4) and REM sleep. The main objective of this study was to characterize the specificity of nociceptive stimulation to trigger sleep arousal-awakening over all sleep stages. Thirteen healthy adults (e.g. without pain or sleep problems; six female and seven male of a mean age of 24.2+/-1.3 years) were included in the study. The responses to noxious intramuscular 5% hypertonic infusion were compared to innocuous vibrotactile and to respective control stimulations: isotonic infusion and auditory stimulations. These stimulations were applied during wakefulness and were repeated during sleep. Polygraphic signals (e.g. brain activity, heart rate) signals were recorded to score sleep arousal over all sleep stages. A subjective assessment of sleep quality was made on next morning. No overnight sensitization or habituation occurred with any of the experimental stimulations. The vibratory auditory stimulations and the noxious hypertonic infusions triggered significantly (P < 0.05) more awakenings in sleep Stage 2 and in REM than their respective control stimulations. In sleep Stage 2, both vibratory + auditory stimulations and the noxious hypertonic infusions has the same awakening response frequency (approximately 30%), however, with the noxious infusions the response frequency were similar in sleep Stages 3 and 4 (P < 0.05) and in REM (trend). Compared to the baseline night, sleep quality was lower following the night with noxious stimulation (90.1+/-2.7 and 73.3+/-7.4 mm, respectively; P < 0.03. These data suggest that pain during sleep could trigger a sleep awaking response over all sleep stages and not only in light sleep. PMID- 15288406 TI - Pain catastrophizing and social support in married individuals with chronic pain: the moderating role of pain duration. AB - In the current study, 96 married chronic pain patients were recruited from the community to test hypotheses about the roles of catastrophizing and psychological distress in relation to perceived support from close others. It was expected that pain duration would moderate the relationship between catastrophizing and perceived support and between catastrophizing and psychological distress. In addition, distress was hypothesized to mediate the relationship between the pain duration-catastrophizing interaction and support. Hierarchical regression analyses showed that pain duration interacted with catastrophizing such that at shorter pain durations, pain catastrophizing was related to more perceived solicitous spouse responses; however no such relationship existed for patients with longer pain durations. In contrast, catastrophizing was significantly related to less perceived spousal support (i.e. support not specific to pain) in patients with longer durations of pain whereas no significant relationship existed for patients with shorter pain durations. Pain duration did not interact with catastrophizing in relating to psychological distress, which precluded the examination of distress as a mediator between the pain duration-catastrophizing interaction and support. Moreover, psychological distress did not significantly mediate the relationships between pain catastrophizing and perceived support. These findings are discussed in the context of cognitive-behavioral and interpersonal perspectives of pain. PMID- 15288407 TI - Differential morphine effects on short- and long-latency laser-evoked cortical responses in the rat. AB - Evoked potential and ensemble neuronal activities were used to study the responses of the primary sensorimotor cortex (SmI) to noxious CO(2) laser irradiation of the middle part of the tail in conscious behaving rats. The hypothesis that systemic morphine treatment preferentially attenuates the longer latency laser-evoked cortical responses was also tested. Laser-evoked potentials (LEPs) and multiple single-unit (SU) activities were, respectively, recorded from chronically implanted stainless-steel screws and microwire electrodes. When examined individually, many SmI neurons showed either short-latency (<100 ms) or long-latency (300-500 ms) responses to laser irradiation. These neurons are widely dispersed in the tail region and hind limb region of the SmI, and also in the forelimb and head regions of the primary motor cortex (MI). Quantitatively, a higher percentage of neurons in the SmI tail region responded with shorter latencies compared to those in the SmI hind limb region or in the MI. When responses of many simultaneously recorded SU were examined together, short latency and long-latency SmI ensemble activities matched the LEP1 and LEP2, respectively. Systemic morphine significantly attenuated the long-latency but not the short-latency component in both LEPs as well as ensemble neuronal activity in the tail region of the SmI. These effects were blocked by naloxone pretreatment. PMID- 15288408 TI - Glyceryl trinitrate triggers premonitory symptoms in migraineurs. AB - Studying attacks of migraine is considerably hampered by its fundamentally episodic nature. Developing approaches to triggering migraine reliably is important for advancing understanding of the disorder by facilitating its study. Based on the work of the Copenhagen Group we administered an intravenous infusion of 0.5 microg/kg/min glyceryl trinitrate (GTN) to 44 migraineurs, 23 migraine without aura, 21 migraine with aura, and to 12 healthy controls. We sought to characterise the GTN-induced migraine in terms of the clinical features of the attacks and reproducibility of triggering, and included a non-migraine control group for the purpose of comparing any effects to exclude an ordering effect. Of the 44 patients administered GTN, 33 had a migraine attack fulfilling International Headache Society criteria. Thirty-two attacks were of migraine without aura and one of migraine with aura. Twelve patients described typical premonitory symptoms, which have not been previously documented with GTN-induced migraine. A repeat attack was triggered in all subjects but one. In one case a visual aura was also triggered both times. Our study shows that GTN-induced triggering is common in our patients, and remarkably reproducible. The data will facilitate the use of the GTN model in studies requiring extensive planning, such as brain imaging, or where preventive questions are at issue. We also report the first patient with a reproducible GTN-triggered migraine with aura. PMID- 15288409 TI - Reproducibility and responsiveness of the Whiplash Disability Questionnaire. AB - This study continued the validation of a Whiplash Specific Disability Questionnaire (WDQ) that was developed from the Neck Disability Index (NDI) using self-reported disabilities in a group of participants experiencing whiplash associated disorders [J Manipulative Physiol Ther 14 (1991) 409]. Previous research has established the content, construct and face validity and internal consistency of the WDQ. The aim of this study was to establish the short-term and medium-term test-retest reliability and responsiveness of the WDQ. Participants (n = 63) receiving physiotherapy treatment for WAD were recruited from 30 private physiotherapy practices in Melbourne, Australia. Each participant completed three WDQ questionnaires over a 1-month period, the first two separated by 24 h. The third questionnaire contained an additional item that asked respondents to rate their perceived change in condition over the month. Reproducibility was determined using an intra-class correlation co-efficient. Responsiveness was assessed via correlation with participant perceived change, the effect size, standardised response mean (SRM) and the responsiveness statistic. Results demonstrated excellent short-term test-retest reliability (ICC 0.96). Reproducibility over 1 month was excellent (ICC 0.93). Correlation between change in WDQ score over 1 month and participant perceived change was r(s) = 0.64, the effect size was 0.03, the SRM was 0.08 and the responsiveness statistics were 0.90 (participants who improved) and -1.60 (participants who deteriorated). The minimal detectable change of the WDQ was established at 15 points. These results demonstrate that the WDQ has excellent short- and medium-term reproducibility and responsiveness in a population seeking treatment for WAD. PMID- 15288410 TI - Maintenance of windup of second pain requires less frequent stimulation in fibromyalgia patients compared to normal controls. AB - Many chronic pain syndromes, including fibromyalgia (FM), show evidence of central nervous system hyperexcitability related to central sensitization. Windup (WU) of second pain reflects increased excitability of spinal cord neurons that is related to central sensitization. Psychophysical testing can help characterize this important central nervous system phenomenon because of the parallels between electrophysiological WU and WU of second pain. Animal experiments have shown that once WU has been established, only low frequency tonic nociceptive input is required to maintain the sensitized state of dorsal horn neurons (WU-maintenance or WU-M). The stimulus frequency necessary to maintain the hyperexcitability of spinal cord neurons can provide a measure of central sensitization. Because central sensitization plays an important role in many chronic pain syndromes including FM, we compared WU-M in 72 normal controls (NC) and 104 FM subjects. WU of second pain was produced by a train of 0.7 s duration thermal pulses applied to the glabrous surface of the hands at a frequency of 0.3 Hz. Enhanced second pain associated with WU could, thereafter, be maintained in FM but not NC subjects for up to 120 s by stimuli delivered at 0.16 and 0.08 Hz (WU-M stimuli). These two frequencies of stimulation do not produce WU when delivered alone. Thus, unlike NC subjects, FM subjects showed enhanced second pain during WU-M stimuli at very low stimulus frequencies, indicating central sensitization. Increased WU sensitivity, enhanced WU-M, and increased WU-related aftersensations help account for persistent pain conditions in FM subjects. In addition to WU, WU M appears to be a useful tool to study mechanisms of pain in patients with characteristics of central sensitization. PMID- 15288411 TI - Venlafaxine extended release in the treatment of painful diabetic neuropathy: a double-blind, placebo-controlled study. AB - To evaluate the efficacy and safety of 6 weeks of venlafaxine extended-release (ER) (75 mg and 150-225 mg) treatment in patients with painful diabetic neuropathy. This multicenter, double-blind, randomized, placebo-controlled study included 244 adult outpatients with metabolically stable type 1 or 2 diabetes with painful diabetic neuropathy. Primary efficacy measures were scores on the daily 100 mm Visual Analog Pain Intensity (VAS-PI) and Pain Relief (VAS-PR) scales. Secondary efficacy measures included the Clinical Global Impressions Severity of Illness and the Clinical Global Impressions-Improvement, Patient Global Rating of Pain Relief, and percentage of patients achieving 50% reduction in pain intensity. Baseline pain intensity was 68.7 mm (moderately severe). At week 6, the percentage reduction from baseline in VAS-PI was 27% (placebo), 32% (75 mg), and 50% (150-225 mg; P < 0.001 vs placebo). Mean VAS-PR scores in the 150-225 mg group were significantly greater than placebo at week 6 (44 vs 60 mm; P < 0.001). The number needed to treat (NNT) for 50% pain intensity reduction with venlafaxine ER 150-225 mg was 4.5 at week 6. Nausea and somnolence were the most common treatment-emergent adverse events. Seven patients on venlafaxine had clinically important ECG changes during treatment. Venlafaxine ER appears effective and safe in relieving pain associated with diabetic neuropathy. NNT values for higher dose venlafaxine ER are comparable to those of tricyclic antidepressants and the anticonvulsant gabapentin. PMID- 15288412 TI - Pre-emptive analgesia using intravenous fentanyl plus low-dose ketamine for radical prostatectomy under general anesthesia does not produce short-term or long-term reductions in pain or analgesic use. AB - The aim of the study was to evaluate post-operative pain and analgesic use after pre-operative or post-incisional i.v. fentanyl plus low dose i.v. ketamine vs. a standard treatment receiving i.v. fentanyl but not ketamine. Men undergoing radical prostatectomy under general anesthesia were randomly assigned in a double blinded manner to one of three groups. Patients received i.v. fentanyl before incision followed by an i.v. bolus dose (0.2 ml kg(-1)) and an i.v. infusion (0.0025 ml kg(-1)min(-1)) of 1 mg ml(-1) ketamine (group 1) or normal saline (groups 2 and 3). Seventy minutes after incision, patients received i.v. fentanyl followed by an i.v. bolus dose (0.2 ml kg(-1)) and an i.v. infusion (0.0025 ml kg(-1)min(-1)) of saline (groups 1 and 3) or ketamine (group 2). Pain, von Frey pain thresholds, and cumulative morphine consumption using patient-controlled analgesia (PCA) were assessed up to 72 h after surgery. 143 patients completed the study (group 1, n = 47, group 2, n = 50, group 3, n = 46). Cumulative PCA morphine (mean+/-SD) did not differ significantly among groups (group 1, 92.3+/ 45.9 mg; group 2, 107.2+/-58.4 mg; group 3, 103.6+/-50.4 mg; P = 0.08 for groups 1 vs. 2, and groups 1 vs. 3). On day 3, the hourly rate (mean+/-SEM) of morphine consumption was significantly lower (p < 0.0009) in group 1 (0.61+/-0.013 mg h( 1)) than group 2 (0.86+/-0.011 mg h(-1)) and group 3 (0.89+/-0.008 mg h(-1)). Pain scores and von Frey pain thresholds did not differ significantly among groups. Two-week and 6-month follow-ups did not reveal significant group differences in pain incidence, intensity, disability or mental health. Pre operative, low-dose administration of i.v. ketamine did not result in a clinically meaningful reduction in pain or morphine consumption when compared with post-incisional administration of ketamine or a saline control condition. PMID- 15288413 TI - Sensory fibers resistant to the actions of tetrodotoxin mediate nocifensive responses to local administration of endothelin-1 in rats. AB - Endothelin-1 (ET-1) applied to the sciatic nerve or injected into the plantar hindpaw of rats induces pain behavior (ipsilateral hindpaw flinching) and selective excitation of nociceptors by activation of endothelin-A (ET(A)) receptors. To determine the pharmacological profile of the sensory fibers that mediate this pain behavior, we administered lidocaine (LID, a non-selective conduction blocker) or tetrodotoxin (TTX) prior to ET-1. LID (1 or 2%, 0.1 ml) was injected percutaneously into the sciatic notch, or TTX (10 microM, 4 microl) was injected into the sciatic nerve prior to the more distal application of ET-1 (400 microM, 40 microl) onto the sciatic nerve or subcutaneously into the plantar hindpaw (400 microM, 10 microl). LID inhibited ET-1-induced flinching in a dose dependent manner; the mean total number of flinches was reduced by 39% for 1% LID and by 87% for 2% LID. In contrast, TTX failed to inhibit flinching behavior induced by sciatic nerve application of ET-1 despite a similar magnitude of motor and sensory blockade as that observed with 2% LID. Partial blockade of flinching behavior by intraneural TTX (mean total flinches were reduced by 51%) was observed after subcutaneous injection of ET-1. Unexpectedly, ET-1 prolonged the actions of 1% LID and, even when applied alone, produced clear signs of motor and sensory conduction block. These results are evidence that ET-1-induced pain is transmitted to the central nervous system via sensory fibers using tetrodotoxin resistant sodium channels, and that ET-1 has analgesic actions that exist despite the activation of local pain pathways. PMID- 15288414 TI - Experimental malignancy in the rat induces early hypersensitivity indicative of neuritis. AB - Cancer pain mechanisms involve multiple factors including the accompanying inflammatory process neural effects. Inflammation along a nerve trunk (neuritis) has been shown to induce hypersensitivity at the innervated target organ. In this experiment the neural effects of MAT B mammary adenocarcinoma cells implanted adjacent to sciatic nerve (MAT group) were compared to those induced by thymus cell extract (THY), saline (SAL) and the potent proinflammatory agent Complete Freund's adjuvant (CFA). Significant pain behavior was detected only in the ipsilateral hindpaw of MAT and CFA rats lasting seven days post-operatively (dpo). On the ninth dpo MAT rats developed significant hyposensitivity to mechanical and electrical stimuli. Interleukin (IL)-6 levels (ELISA) from MAT and CFA exposed nerves were significantly elevated at dpo 2 and remained so in MAT at dpo 8. Indomethacin (1 mg/kg i.p.) abolished the observed pain responses in MAT and CFA rats exposed nerves. Light microscopy of the MAT nerves at the second dpo revealed neural infiltration of immune and malignant cells with mild edema. By the seventh dpo there was nerve damage and at dpo 14 nerve tissue was largely replaced by malignant and immune cells. Electrophysiology of saphenous nerves exposed to MAT, CFA or SAL revealed significantly increased spontaneous activity in MAT and CFA groups. Spike response to hindpaw mechanical stimulation was significantly reduced only in the MAT group (dpo 6-9) suggestive of nerve damage. Inflammatory neuritis is an early expression of malignancy and may play a role in chronic cancer-related pain initiation and additionally may offer diagnostic opportunities. PMID- 15288415 TI - c-fos and CRF receptor gene transcription in the brain of acetic acid-induced somato-visceral pain in rats. AB - We aimed to characterize neuronal and corticotrophin-releasing (CRF) pathways in a model of somato-visceral pain in rats. Male rats received an intraperitoneal (i.p.) injection of either vehicle (controls) or acetic acid (AA) and were sacrificed 1, 2, 3, 4, or 6 h later. Coronal frozen sections of the brain were cut and mRNAs encoding the rat c-fos, CRF(1), CRF(2 alpha,beta) receptors were assayed by in situ hybridisation histochemistry. Localization of these transcripts within CRF-immunoreactive (i.r.) neurons of the paraventricular nucleus (PVN) of the hypothalamus was also determined. AA i.p. induced c-fos mRNA expression in brain nuclei involved in the autonomic, behavioural and neuroendocrine response to pain. Some of these nuclei are involved in the control of digestive motility, as represented by the PVN, locus coeruleus and nucleus tractus solitarius. CRF pathways, in particular in the PVN, are activated in this model. Indeed, a robust signal of c-fos and CRF(1) transcripts was observed in the PVN and numerous CRF-i.r. neurons expressed c-fos or CRF(1) transcripts in the PVN of AA-treated animals. In contrast, no expression of CRF(2) transcripts was observed in the PVN either in basal conditions or after AA i.p. These data argue for an activation of CRF pathways within the PVN in this model of somato visceral pain. The use of CRF antagonists, particularly of the CRF(1) type, should have an interest in somato-visceral pain pathology. PMID- 15288416 TI - In vitro opioid induced proliferation of peripheral blood immune cells correlates with in vivo cold pressor pain tolerance in humans: a biological marker of pain tolerance. AB - There is substantial evidence for bidirectional communication between the immune system and the central nervous system, as the cells and signalling molecules of the immune system influence many central nervous system functions, for instance nociception. Opioids, such as morphine, produce analgesia and numerous other central and peripheral effects including sedation and euphoria, while their effects on the immune system are wide-ranging. There is considerable interindividual variability in basal nociception and response to opioids, however, the physiological and biological mechanisms underlying this are unclear. Therefore, we investigated the relationship between the immune system and basal nociceptive thresholds, using the proliferative response of isolated peripheral blood mononuclear cells and cold pressor pain tolerance. Here we show that the percent increase in proliferation of peripheral immune cells from 13 healthy subjects incubated with morphine ex vivo is highly correlated with the subjects' tolerance to noxious cold stimuli (Pearson r = 0.92, P < 0.0001). These pilot data provide evidence of a novel objective biological marker of pain tolerance in humans, which also links the immune and opioid systems with basal pain tolerance. PMID- 15288417 TI - Impaired self-perception of the hand in complex regional pain syndrome (CRPS). AB - To investigate neglect, extinction, and body-perception in patients suffering from complex regional pain syndrome (CRPS). So-called 'neglect-like' symptoms have been reported in CRPS, however no studies have yet analyzed this phenomenon which might substantiate the theory of the central nervous system involvement in the pathophysiology of CRPS. A total of 114 patients with CRPS of the upper limb underwent bedside neurological examination. 'Neglect-like' symptoms were determined by asking all patients what kind of feeling they had toward the affected hand (feeling of foreignness). Hemispatial neglect was tested with the line bisection task in 29 patients and sensory extinction to simultaneous stimulation in 40 patients. The ability to identify fingers after tactile stimulation was tested in 73 patients. Independently of the affected side and disease duration, 54.4% of the patients reported that their hand felt 'foreign' or 'strange'. The ability to identify fingers was impaired in 48% on the affected hand and in 6.5% on the unaffected hand ( X(2) = 33.52, df = 1, p < 0.0001). These findings were related to pain intensity, illness duration and the extent of sensory deficits. No typical abnormalities indicating neglect were found in the line bisection test. Sensory extinction was normal in all patients. A large proportion of CRPS patients have disturbances of the self-perception of the hand, indicating an alteration of higher central nervous system processing. There are no indicators that classic neglect or extinction contribute to these findings. Physical therapy of such patients should take this observation into consideration. PMID- 15288418 TI - Safety and efficacy of intranasal ketamine in a mixed population with chronic pain. PMID- 15288419 TI - Comment on: Bell RF, Kalso K. Is intranasal ketamine an appropriate treatment for chronic non-cancer breakthrough pain? Pain 2004;108:1-2. PMID- 15288421 TI - Intra-nasal ketamine for somatization? PMID- 15288422 TI - Different behavioral observation methods serve different purposes. PMID- 15288424 TI - Primary cultures of rat astrocytes respond to thiamine deficiency-induced swelling by downregulating aquaporin-4 levels. AB - Imaging studies indicate that cerebral edema is an important consequence of Wernicke's encephalopathy (WE), a disorder caused by thiamine deficiency (TD). We have investigated this problem using a recently developed in vitro astrocyte model of TD. Measurement of cell volume using the 3-O-methylglucose uptake method revealed a dose-dependent swelling of astrocytes during exposure to TD conditions. Time course studies indicated a progressive volume increase up to a maximum of 93% above controls after 4 days of treatment. This swelling then partially resolved, and remained stable for up to 10 days following commencement of TD treatment. Measurement of aquaporin-4 (AQP-4) levels showed a 44% loss after 10 days and a temporal profile consistent with an important role for this water channel protein in astrocyte cell volume changes during TD. Our findings of astrocyte swelling in TD are consistent with previous reports of focal brain edema in cases of WE, and indicate that AQP-4 may be an important target for ameliorating some of the clinical problems associated with this disorder. PMID- 15288425 TI - The processing of speech and non-speech sounds in aphasic patients as reflected by the mismatch negativity (MMN). AB - This study compared the discrimination of speech and non-speech sounds in left hemisphere stroke patients with aphasia and healthy controls by recording event related potentials (ERPs) and behavioural responses. It was found that the mismatch negativity (MMN) amplitude for vowel and duration changes in speech sounds was diminished in the patients. In contrast, there was no significant difference between the groups in the MMN for comparable frequency and duration changes in non-speech sounds. In the behavioural session, the patients were slower than the control subjects in discriminating duration changes of both types of sounds. These results suggest that left-hemisphere lesions have differential effects on the discrimination of speech and acoustic features indicating that they have separate neural substrates. PMID- 15288426 TI - Regulation of calcitonin gene-related peptide release from rat trigeminal nucleus caudalis slices in vitro. AB - Calcitonin gene-related peptide (CGRP) released from trigeminal primary afferents has been implicated in the pathophysiology of migraine. Here, we have used an in vitro slice preparation to investigate its release from nerve terminals in the rat trigeminal nucleus caudalis. Extracellular-calcium dependent CGRP release was stimulated by both capsaicin and neuronal depolarization with KCl. The capsaicin (1 microM)-evoked CGRP release was blocked by capsazepine and was also attenuated in the presence of the cyclooxygenase inhibitor, indomethacin, an effect that was reversed when slices were stimulated with capsaicin in the presence of the cyclooxygenase metabolite, prostaglandin E(2). Taken together, these data further highlight the importance of prostaglandins as enhancers of neuropeptide release and suggest that CGRP released from the central terminals of trigeminal neurones has the potential to be involved in the transmission of nociceptive information of relevance to migraine headache. PMID- 15288427 TI - Does support surface coefficient of friction influence postural dynamics and isometric force productions of the upper extremities performed by seated subjects? AB - The purpose of this study is to determine if increasing the potential risk of slipping on support surface materials decreases postural dynamics and task performance. Seated subjects were asked to exert maximum horizontal two-handed isometric pushes as quickly as possible on a dynamometric bar. Reaction forces exerted at seat (Rxs and Rzs) and footrest (Rxf and Rzf) were measured using force sensors. A dynamometric bar measured horizontal push force (Fx). Global adherence ratio (micro) was calculated. Three support surface materials, characterised by different coefficients of friction, were tested. The same surfaces were placed on the seat and the footrest during a series of pushes. It was shown that the different support surfaces significantly influence Fx, Rxs, Rzs, Rzf and micro maximum values obtained at the end of the push. Even when the instructions given to the subject are to produce a maximum push, the maximum horizontal force applied to the bar differs according to the support surface material. Dynamic postural adjustments are influenced by characteristics of slipperiness of support surfaces. The coefficient of friction contributes an essential element to the program of postural dynamics, which in turn modulates the task performance. During brief isometric pushes, seated subjects have the capacity to control the potential risk of slipping. PMID- 15288428 TI - Conditioning Ia-afferent stimulation reduces the soleus Hoffman reflex in humans when muscle spindles are assumed to be inactive. AB - Despite higher neural activation during active as compared to passive muscle shortening, Hoffman reflexes (H-reflexes) are similar. This may be explained by homosynaptic post-activation depression (HPAD) of Ia-afferents being present during active shortening. Accordingly, it was investigated whether conditioning electrical stimulation of the tibial nerve reduced the H-reflex less during active than passive shortening. The effects of two conditioning modes (0.2 and 1 Hz) were compared to a control mode without conditioning. H-reflexes and M-waves were elicited as the ankle passed 90 degrees with the soleus muscle undergoing passive or active (20% MVC) lengthening or shortening. Conditioning had no effect during active shortening. In contrast, during passive shortening, the H:M of the 1 Hz mode was significantly less than that of the 0.2 Hz and control modes. In lengthening, H:M was unaffected by conditioning. These findings support that HPAD reduces the synaptic efficacy of Ia-afferents during active shortening, active and passive lengthening, but not passive shortening. PMID- 15288429 TI - Electrophysiological correlates of grapheme-phoneme conversion. AB - The cortical processes underlying grapheme-phoneme conversion were investigated by event-related potentials (ERPs). The task consisted of silent reading or vowel matching of three Japanese hiragana characters, each representing a consonant vowel syllable. At earlier latencies, typical components of the visual ERP, namely, P1 (110 ms), N1 (170 ms) and P2 (300 ms), were elicited in the temporo occipital area for both tasks as well as control task (observing the orthographic shapes of three Korean characters). Following these earlier components, two sustained negativities were identified. The earlier sustained negativity, referred here to as SN1, was found in both the silent-reading and vowel-matching task but not in the control task. The scalp distribution of SN1 was over the left occipito-temporal area, with maximum amplitude over O1. The amplitude of SN1 was larger in the vowel-matching task compared to the silent-reading task, consistent with previous reports that ERP amplitude correlates with task difficulty. SN2, the later sustained negativity, was only observed in the vowel-matching task. The scalp distribution of SN2 was over the midsagittal centro-parietal area with maximum amplitude over Cz. Elicitation of SN2 in the vowel-matching task suggested that the vowel-matching task requires a wider range of neural activities exceeding the established conventional area of language processing. PMID- 15288430 TI - Cortico-muscular coupling in a patient with postural myoclonus. AB - We investigated the cortico-muscular coherence in a patient with posturally induced cortically originating negative myoclonus. We recorded simultaneously 50 channels EEG and EMG from quadriceps and biceps femoris muscles of the left upper leg. Three experimental conditions were investigated with the patient in a seated position: (i) recording during rest (Rest), (ii) recording while the patient had to hold his left leg horizontally stretched out (Postural), and (iii) recording while the patient had to hold his left leg horizontally stretched out against a vertical force (Postural against force). Coherence, phase difference and cumulant density were computed as indicators for cortico-muscular coupling. The cortical component preceding the silent period was shown by averaging and was reconstructed. During postural and postural against force conditions, the EEG over the vertex was significantly coherent with EMG, in alpha (7-15 Hz) and beta range (15-30 Hz). The strongest coherence peak was at 21 Hz. No high-frequency coherence was observed. The phase difference and the cumulant density estimate corresponded to a 32 ms time lag between motor cortex and muscles, with EEG leading. The broadening of the coherence spectrum at which the motor cortex drives the muscles together with the excessive coherence levels and the giant SEP could reflect the hyperexcitability of the sensorimotor cortex. The frequency content of the coherence may be characteristic for this type of myoclonus. The results lend support to the view that the frequency analysis may have some diagnostic potential in cortical myoclonus. PMID- 15288431 TI - Accumulation of aluminum in ferritin isolated from rat brain. AB - The neurotoxic effects of aluminum have been widely reported but the mechanism of action and detoxification is poorly understood. To investigate the toxic potential of aluminum, we found it necessary to detail the behavior of absorbed aluminum in brain. The aim of this study was to clarify the distribution of aluminum in the brain. Rats were exposed to aluminum lactate intraperitoneally for 7 weeks. Although no marked differences in aluminum content was observed in brain regions, aluminum was eluted by gel filtration chromatography of the ferritin fraction from aluminum-loaded brain extracts; 5.9% of the total brain aluminum was recovered in purified ferritin from aluminum-loaded rat brains. These results suggest that ferritin may function as an aluminum detoxicant in the cell. PMID- 15288432 TI - Association of ABCA1 with late-onset Alzheimer's disease is not observed in a case-control study. AB - Genetic association of ABCA1 or the ATP-binding cassette A1 transporter with late onset Alzheimer's disease (LOAD) has recently been proposed for a haplotype comprised of three single nucleotide polymorphisms (SNPs). We have genotyped these and other ABCA1 SNPs in a LOAD case-control series of 796 individuals (419 cases versus 377 controls) collected at Washington University. While our sample series is larger and thus presumably has greater power than any of the series used to implicate ABCA1, we were unable to replicate the published association, using either single markers or multiple marker haplotypes. Further, we did not observe significant and replicated association of other ABCA1 SNPs we examined with the disease, thus these ABCA1 variants do not appear to influence the risk of LOAD in this study. PMID- 15288433 TI - Very delayed neuronal loss occurs in the glomerular layer of the main olfactory bulb following transient ischemia in gerbils. AB - Olfactory dysfunction could happen following various insults such as ischemic hypoxic state. Neurons of the main olfactory bulb (MOB) are resistant to ischemic damage. In the present study, we investigated the ischemia-related changes of neurons and glial cells in the glomerular layer (GL) of the gerbil MOB after transient ischemia. The number of NeuN-immunoreactive neurons became to decrease from 10 days after ischemic insult. Fifteen days after ischemic insult, astrocytes and microglia were increased in number. By 60 days after ischemia, NeuN-immunoreactive neurons were significantly decreased by 42% per glomerulus. At this time period, astrocytes and microglia were pronouncedly increased. This result indicates that neuronal loss must be much delayed in the GL following transient ischemia. PMID- 15288434 TI - Age-related changes in the levels of voltage-dependent calcium channels and other synaptic proteins in rat brain cortices. AB - Neurotransmitter release from synapses is one of the most important interneuronal signaling in the nervous system. We previously reported that aging decreases depolarization-induced acetylcholine release in rat brain synaptosomes. To investigate the mechanisms underlying the age-related decrements of neurotransmission, we determined the levels of the alpha1 subunit proteins of voltage-dependent calcium channels (VDCCs) and three synaptic proteins that relate to exocytotic processes using synaptosomes prepared from cerebral cortices of young (6-month-old) and aged (27-month-old) rats. Immunoblotting analyses revealed that the protein levels of alpha1A (P/Q-type) and alpha1B (N-type) subunits in aged rats were 38% and 43% lower than the levels of young rats, respectively, but the levels of the alpha1C (L-type) subunit were not different between young and aged. On the contrary, the levels of synaptotagmin-1, synaptophysin and syntaxin were not significantly different between the two age groups in the synaptosomal preparations. These results suggest that synaptic density does not change much in the cerebral cortex in normal aging, and that the reduction of P/Q-type and N-type VDCCs, both of which participate in neurotransmitter release, is one of the causes for the decrease of neurotransmission at aged synapses. PMID- 15288435 TI - Gene-environment interaction in hyperkinetic conduct disorder (HD + CD) as indicated by season of birth variations in dopamine receptor (DRD4) gene polymorphism. AB - Recently, an interaction between season of birth and the expression of candidate genes has been suggested. Season of birth variations in tryptophan hydroxylase (TPH), the serotonin transporter (5-HTTLPR) and the dopamine D4 receptor (DRD4) gene polymorphisms are different for affective disorders and schizophrenia. The DRD4 gene has been postulated as a candidate gene for attention-deficit hyperactivity disorder (ADHD), equivalent to hyperkinetic disorder (HD). The seven-repeat long variant of this gene (DRD4*7) in comparison to the short repeat variants of the DRD4 gene polymorphism, has been found to be associated with ADHD. A seasonal pattern of birth has also been proposed for different subtypes of ADHD. Therefore, in a subgroup of children with HD and conduct disorder (CD) and in healthy controls, we investigated a possible association between the DRD4*7 allele and HD + CD in association with the season of birth. Supporting this hypothesis, we found an interaction between the seasons of birth and the expression of the DRD4 candidate gene in children with HD + CD as well as in controls, which differ significantly from each other. Depending on the season of birth, children carrying the DRD4*7R allele showed different relative risks for developing HD + CD. PMID- 15288436 TI - Intravenously injected neural progenitor cells of transgenic rats can migrate to the injured spinal cord and differentiate into neurons, astrocytes and oligodendrocytes. AB - Transplantation of neural progenitor cells (NPCs) has been reported recently to promote regeneration of the injured spinal cord. In the majority of these reports, cell transplantation was performed by local injection with a needle. However, direct injection might be too invasive for clinical use; therefore, the authors investigated a new method of delivering NPCs for the treatment of spinal cord injury. In this study, NPCs were obtained from E15 fetal hippocampus of transgenic rats expressing green fluorescent protein and 100,000 cells were transplanted intravenously into each animal 24h after contusion injury. It was found that the injected NPCs migrated to the lesion site widely and demonstrated nestin at an early phase after transplantation. These NPCs differentiated into neurons, astrocytes and oligodendrocytes, and survived at least for 56 days. These results indicated that intravenously injected neural stem cells migrated into the spinal cord lesion while preserving their potential as NPCs, and that this procedure is a potential method of delivering cells into the lesion for the treatment of spinal cord injury. PMID- 15288437 TI - Status epilepticus induced by pilocarpine and Ca2+ transport by microsome in the hippocampus of rats. AB - An increase in intra-neuronal Ca(2+) concentration has been associated to status epilepticus (SE). Ca(2+) is stored in the endoplasmic reticulum, mediated by the Ca(2+)-ATPases (SERCAs). Here we studied the Ca(2+)-ATPase activity and the SERCA2b distribution in the hippocampus of rats submitted to 5h of SE. The Ca(2+) uptake was measured using [45Ca]CaCl(2) and the hippocampal distribution of SERCA2b was analyzed by immunohistochemistry. A reduction in the amount of cells expressing SERCA2b in CA1, CA3 and dentate gyrus was observed. However, the surviving cells of these regions increased the SERCA2b immunoreactivity, when compared with control tissues. The Ca(2+)-ATPase activity measured in all hippocampal formation was not modified by SE. These results suggest that SE promotes a redistribution of SERCA2b in the hippocampus as a compensatory Ca(2+) transport mechanism. PMID- 15288438 TI - The distribution of c-myb immunoreactivities in the adult mouse retina. AB - The myb gene family is composed of three different myb-related genes, A-, B- and c-myb. Among these, the presence of c-myb mRNA in developing and adult retina was previously reported. However, further study on the expression of c-myb in the retina is warranted because the previous study was only performed on developing retinal tissues by in situ hybridization technique. Therefore, in this study, we tried to perform immunohistochemical study, with a focus on the c-myb protein expression in adult retina. Although the cell types were unconfirmed, c-myb protein was likely to have been found in the ganglion, amacrine, horizontal and photoreceptor cells, judging from their locations and morphologies. The experimental data suggested that c-myb immunoreactivities were expressed by the cells in the neural retina, even in the adult stage, especially within some types of cells in specified retinal layers. This suggests that c-myb might play a role in the physiology of the retinal cells, not only in differentiation during retinal development. PMID- 15288439 TI - Different response to exogenous L-arginine in nitric oxide production between hippocampus and striatum of conscious rats: a microdialysis study. AB - We previously showed that systemic administration of a nitric oxide (NO) precursor, L-arginine (L-Arg), failed to reverse suppression by NO synthase (NOS) inhibitors of chemically induced shaking behavior in rats, leading to the hypothesis that exogenous L-Arg might be non-uniformly supplied to brain regions susceptible to NOS inhibitors. In the present study, therefore, we examined the effect of exogenous L-Arg on the extracellular levels of the oxidative nitric oxide (NO) products, nitrite (NO2-) and nitrate (NO3-), in two different brain regions, the hippocampus and the striatum, of conscious rats by means of in vivo brain microdialysis. The basal NO2- levels in the two brain regions were comparable, while the NO3- level was significantly lower in the hippocampus than the striatum. The addition of 10 mM L-Arg, but not D-Arg, to the perfusing solution significantly increased NO2- and NO3- in the hippocampus and NO2- alone in the striatum. These increases were abolished by 1 mM N(omega)-nitro-L arginine, an NOS inhibitor. L-Arg at 1mM was able to significantly increase NO2-, but not NO3-, in the hippocampus to a level comparable with that at 10 mM L-Arg, while it had no effect in the striatum. L-Arg (500 mg/kg, i.p.) induced a significant increase in NO2- and NO3- in the hippocampus, but not in the striatum. These results suggest that the striatum may have a lower ability to enhance NO production by utilising exogenous L-Arg than the hippocampus, despite higher basal NO production. PMID- 15288440 TI - The Ebbinghaus illusion affects on-line movement control. AB - Changes in the planning and control of discrete aiming movements in response to the introduction and removal of the Ebbinghaus size-contrast illusion were examined. Movements were executed faster to targets that appeared larger following movement initiation. The differences in movement time were associated with the portion of the movement associated with on-line control. The results are inconsistent with the assumptions of the planning and control model [Behav. Brain Sci. (in press); J. Exp. Psychol. Hum. Perc. Perf. 27 (2001) 560], and the perception and action dissociation model of goal-directed movement [A.D. Milner, M.A. Goodale, The Visual Brain in Action, Oxford University Press, New York, Oxford, 1995]. PMID- 15288441 TI - The chemokine receptor CCR5-Delta32 gene mutation is not protective against Alzheimer's disease. AB - Chronic local inflammatory reaction involving reactive microglia is one of the major pathological events in Alzheimer's disease (AD). There is growing evidence that the chemokine receptor CCR5 is up-regulated in AD brain and plays a role in the recruitment and accumulation of microglia in senile plaques. A 32-base pair deletion in the CCR5 gene (CCR5-Delta32 mutant allele) confers resistance to HIV 1 infection by preventing expression of the receptor on the cell surface. Several other reports have shown a similar protective effect of CCR5-Delta32 mutation towards certain chronic inflammatory diseases. Given the potential importance of CCR5 in brain inflammation, we hypothesized that individuals carrying the CCR5 Delta32 allele would show a reduced risk of AD. So, we performed a case-control study in 376 Spanish AD patients and 369 healthy controls. The frequency of the CCR5-Delta32 allele in our AD sample was 7.8%, not significantly different from our control sample group (5.8%). The present study indicates that the CCR5 Delta32 allele is not a preventive factor for AD. PMID- 15288442 TI - Direction-selective visual responses in macaque superior colliculus induced by behavioral training. AB - In a previous report, we described a heretofore undetected population of neurons in the intermediate and deep layers of the monkey superior colliculus (SC) that yielded directionally selective visual responses to stimuli presented within the central 4 degrees of the visual field. We observed these neurons in three monkeys that had been extensively trained to perform a visual direction discrimination task in this region of the visual field. The task required the monkeys to report the perceived direction of motion by making a saccadic eye movement to one of two targets aligned with the two possible directions of motion. We hypothesized that these neurons reflect a learned association between visual motion direction and saccade direction formed through extensive training on the direction discrimination task. We tested this hypothesis by searching for direction selective visual responses in two monkeys that had been trained to perform a similar motion discrimination task in which the direction of stimulus motion was dissociated from the direction of the operant saccade. Strongly directional visual responses were absent in these monkeys, consistent with the notion that extensive training can induce highly specific visual responses in a subpopulation of SC neurons. PMID- 15288443 TI - Amyloid beta-protein induced electrophysiological changes are dependent on aggregation state: N-methyl-D-aspartate (NMDA) versus non-NMDA receptor/channel activation. AB - Alzheimer's disease (AD) is a progressive neurodegenerative disease, however, the underlying mechanism driving this condition is unknown. Unexplored is the possibility that the time-dependent generation of different Abeta assemblies may underlie the pathogenic cascade with biophysically distinct structures interacting with unique biological targets. Thus, the presence of subtle alterations in synaptic function during the earliest clinical phase of AD may be mediated by diffusible assemblies of the amyloid beta-protein (Abeta). Using primary neocortical cultures, here we compare the synaptic responses induced by two different Abeta assemblies, protofibrils (PFs) and fibrils (FBs), and demonstrate for the first time that neuronal activation was selectively dependent on the assembly state of Abeta. PF-induced activity was specifically attenuated by the N-methyl-D-aspartate (NMDA) receptor antagonist, D-APV. In contrast, the non-NMDA glutamate receptor antagonist, NBQX, preferentially reduced FB-induced activity. In support, removal of Mg(2+) from the medium, which enhances NMDA channels, increased both PF- or FB-induced activation, but D-APV was more effective in attenuating PF-induced excitatory activity. These findings suggest that PFs may activate neurons differently than fibrils and lend support to the hypothesis that pre-fibrillar assemblies of Abeta may play an important role in the development of AD-type synaptic deficits. PMID- 15288444 TI - Case-only study of interactions between genetic polymorphisms of GSTM1, P1, T1 and Z1 and smoking in Parkinson's disease. AB - Current opinion contends that complex interactions between genetic and environmental factors play a role in the etiology of Parkinson's disease (PD). Cigarette smoking is thought to reduce risk of PD, and emerging evidence suggests that genetic factors may modulate smoking's effect. We used a case-only design, an approach not previously used to study gene-environment interactions in PD, specifically to study interactions between glutathione-S-transferase (GST) gene polymorphisms and smoking in relation to PD. Four-hundred PD cases (age at onset: 60.0 +/- 10.7 years) were genotyped for common polymorphisms in GSTM1, P1, T1 and Z1 using well-established methods. Smoking exposure data were collected in face to-face interviews. The independence of the studied GST genotypes and smoking exposure was confirmed by studying 402 healthy, aged individuals. No differences were observed in the distributions of GSTM1, T1 or Z1 polymorphisms between ever smoked and never-smoked PD cases using logistic regression (all P > 0.43). However, GSTP1 *C haplotypes were over-represented among PD cases who ever smoked (odds ratio for interaction (ORi) = 2.00 (95% CI: 1.11-3.60, P = 0.03)). Analysis revealed that ORi between smoking and the GSTP1-114Val carrier status increased with increasing smoking dose (P = 0.02 for trend). These data suggest that one or more GSTP1 polymorphisms may interact with cigarette smoking to influence the risk for PD. PMID- 15288445 TI - Acute and chronic ethanol does not affect incisional pain in neonatal rats. AB - We have previously found that in post-natal day 7 rats withdrawal from acute and chronic ethanol (EtOH) exposure lowers mechanical thresholds during withdrawal and exacerbates spontaneous pain responses to an inflammatory injury 4 days post withdrawal. These findings suggested alterations in somatosensory pathways following EtOH exposure during the third trimester developmental equivalent. In this study we wanted to determine whether EtOH exposure during the third trimester equivalent exacerbates mechanical allodynia and thermal hyperalgesia produced by an incisional model of post-operative pain at post-natal day 21. The extent and duration of mechanical allodynia and thermal hyperalgesia following incision was measured and found to be unaffected by prior EtOH exposure. PMID- 15288446 TI - Lack of a genetic association between the frizzled-3 gene and schizophrenia in a British population. AB - In recent studies, the frizzled-3 (FZD3) locus was found to be associated with schizophrenia in both Japanese and Chinese populations. To validate the initial finding, we detected three single nucleotide polymorphisms (SNPs) present in a 10 kb segment of DNA at the FZD3 locus, as described in a previous study with a Chinese population. We totally recruited 120 British family trios consisting of fathers, mothers and affected offspring with schizophrenia. The transmission disequilibrium test (TDT) did not show allelic association between these three SNPs and schizophrenia. The 3-SNP haplotype system was composed of only 3 individual haplotypes among the 120 family trios and these 3 SNPs were mainly carried by two distinct haplotypes, suggesting that these 3 SNPs may result from a single founding event in history. No association was shown between the 3-SNP haplotypes and schizophrenia. The present results imply that the FZD3 gene is less evolutionary in the British population than in the Chinese population. This may be a possible reason for the failure to replicate the FZD3 finding with the British sample. PMID- 15288447 TI - Opioid receptors in fast and slow skeletal muscles of normal and dystrophic mice. AB - The density of beta-endorphin receptors and the proportions of fibres that expressed the receptors was assessed in fast extensor digitorum longus muscle and slow soleus muscles of normal and dystrophic mice using [125I]beta-endorphin and autoradiography. In the EDL the density was approximately 3.5 times higher and the proportion of labelled fibres approximately 2.6 times higher in dystrophic mice than normal mice. In the soleus the density was approximately 6.4 times higher and the proportion of labelled fibres approximately 1.5 times higher in the dystrophic mice than the normal mice. The receptors were of the delta-opioid subtype. PMID- 15288448 TI - Human auditory evoked mismatch field amplitudes vary as a function of vowel duration in healthy first-language speakers. AB - Previous auditory studies demonstrated that vowel shortening elicited a more prominent mismatch component than its lengthening in event-related potentials (ERP) and magnetoencephalography (MEG). Based on these findings, the current study investigated whether the magnetic mismatch field (MMF) component would be generated by vowel shortening of various degrees to determine a neuronal response threshold of pre-attentive deviation detection. Behavioral pre-test data revealed that while listening to Japanese short-duration (100%: reference), long-duration (180%), and other in-between duration-synthesized types, healthy native speakers of Japanese failed to clearly categorize 140-124% durations as either short or long words, while categorizing 108-116% durations as short words and 148-172% durations as long. Following these results, MEG responses were recorded with a whole-head 148-channel magnetometer, as subjects listened to 100% standard and five deviant durations (124, 132, 140, 148, 180%). MEG results showed that the above-32% duration decrements (180-->100%, 148-->100%) elicited a more prominent MMF than the others, the MMF amplitudes increasing linearly to the degree of duration deviance, and that neuronal responses correlated with behavioral word categorization accuracy. PMID- 15288449 TI - Multidirectional neck strength and electromyographic activity for normal controls. AB - OBJECTIVE: To assess the multidirectional force and indwelling electromyographic activity during maximal effort isometric actions of the neck. DESIGN: A descriptive study involving maximal effort isometric actions of the neck and bilateral electromyographic activity. BACKGROUND: This study extends previous efforts to assess the isometric strength of the neck, but with greater precision with respect to the intermediate angles between the frontal and sagittal planes. METHODOLOGY: Participants (n = 18) generated a maximal isometric force in twelve directions in the horizontal plane. All exertions were realized in neutral position. A load cell measured forces and intramuscular fine-wire electrodes were used to record the bilateral electromyographic activity of the sternocleidomastoid, scalenus medius, trapezius (middle fibers), semispinalis capitis, and splenius capitis. RESULTS: Strength in the anterolateral directions were similar, but exhibited right lateral dominance in extension. The sternocleidomastoid and trapezius (middle fibers) exhibited bilateral symmetry while the scalenus medius, semispinalis capitis, and splenius capitis did not. Furthermore, the agonist, synergist, and antagonist action of the individual muscles was clear. The direction of force that resulted in the greatest electromyographic activity was consistent with what has been shown in anatomy texts. This was not true for the scalenus medius. We showed that the scalenus medius contributes to extension, with synergistic activity in the lateral bending direction. CONCLUSIONS: The greater precision revealed novel information about the isometric strength of the neck and its musculature. PMID- 15288450 TI - A protocol for the assessment of 3D movements of the head in persons with cervical dystonia. AB - OBJECTIVE: To design and test a protocol for the assessment of neck movements in patients affected by cervical dystonia by using an electromagnetic system. This approach could overcome the limits of the current assessment scales in this specific field. BACKGROUND: Initial assessment and function recovery during treatments are diagnosed by the clinician using outcome scales which present many drawbacks in terms of easiness of use, sensitivity, and reliability. DESIGN: A three-dimensional motion analysis system was used to record six different head movements. METHODS: Six able-bodied subjects and 10 subjects affected by cervical dystonia participated in this study. For the different head movements three kinematic parameters (a symmetry index and two indexes related to the reduction of the range of motion) have been extracted in order to compare the performance of able-bodied and disabled persons. RESULTS: The features selected allowed highlighting of the differences between able-bodied and disabled subjects for the degrees of freedom of the neck. CONCLUSIONS: Using a motion analysis system, three kinematic features were extracted from head movements. They seem to allow a more objective assessment of the disability and a more appropriated strategy for the management of patients affected by cervical dystonia. PMID- 15288451 TI - Musculoskeletal parameters of muscles crossing the shoulder and elbow and the effect of sarcomere length sample size on estimation of optimal muscle length. AB - BACKGROUND: Knowledge of musculoskeletal parameters is essential to understanding and modeling a muscle's force generating capability. A study of musculoskeletal parameters was conducted in two parts: (I) Empirical measurement of upper extremity musculoskeletal parameters. (II) Computational bootstrap simulation to examine statistical power of detecting optimal muscle length as a function of sarcomere length sample size and effect size. METHODS: Parameters were determined with a cadaver model. Sarcomere lengths were measured for 120 samples per muscle using laser diffraction and the mean sarcomere length used to estimate optimal muscle length. A bootstrap computational simulation was conducted to estimate variance in mean sarcomere length as a function of sample size. Statistical power for detecting optimal muscle length as a function of sample size and effect size was then determined. FINDINGS: Parameters are reported in tabular format. Power is 80% at approximately 85, 50, 40 and 25 samples for effect sizes of 0.5, 0.75, 1.0 and 1.5 mm respectively. INTERPRETATION: Musculoskeletal parameters for predicting muscle forces can be adequately measured in a cadaver model. Measurement of 40-60 sarcomere lengths per muscle is sufficient to calculate mean sarcomere length for estimating optimal muscle length with power of 80% for an effect size of 0.75-1.0 mm. PMID- 15288452 TI - Measurement of angular wrist neutral zone and forearm muscle activity. AB - OBJECTIVES: To determine the forearm muscles activity in different wrist deviated positions and wrist neutral zone, and to assess the self-selected resting position without visual feedback. BACKGROUND: Wrist deviation occurs in almost all industrial and office jobs. This has been deemed hazardous for carpal tunnel syndrome. Proper resting wrist position is likely to decrease the hazard for carpal tunnel pressure. METHODS: Twenty blindfolded subjects without history of hand/forearm musculoskeletal disorders participated in the study. The EMG of the forearm muscles (flexor carpi radialis, flexor carpi ulnaris, extensor carpi radialis and, extensor carpi ulnaris) in deviated and neutral wrist postures was recorded at a sampling rate of 1 kHz. Also, wrist neutral zone at rest was measured using a custom-made calibrated uniaxial electrogoniometer. One-way ANOVA with repeated measures was used in order to find the impact of wrist deviation on muscles activity. RESULTS: The participants positioned their wrist in rest at 7 degrees -9 degrees extension and 5 degrees -7 degrees ulnar deviation. Significantly higher EMG activity was recorded for each muscle in the wrist deviated postures when compared to neutral position (P < 0.001). CONCLUSIONS: Self selected wrist neutral posture decreased the muscle activity significantly. Placement of wrists in neutral zone is expected to reduce risk of injuries. PMID- 15288453 TI - The effect of cane use on the compensatory step following posterior perturbations. AB - OBJECTIVE: The compensatory step is a critical component of the balance response and is impaired in older fallers. The purpose of this research was to examine whether utilization of a cane modified the compensatory step response following external posterior perturbations. DESIGN: Single subject withdrawal design was employed. Single subject statistical analysis--the standard deviation bandwidth method--supplemented visual analysis of the data. METHODS: Four older adults (range: 73-83 years) with balance impairment who habitually use a cane completed this study. Subjects received a series of sudden backward pulls that were large enough to elicit compensatory stepping. We examined the following variables both with and without cane use: timing of cane loading relative to step initiation and center of mass acceleration, stability margin, center of mass excursion and velocity, step length and width. RESULTS: No participant loaded the cane prior to initiation of the first compensatory step. There was no effect of cane use on the stability margin, nor was there an effect of cane use on center of mass excursion or velocity, or step length or width. CONCLUSIONS: These data suggest that cane use does not necessarily improve balance recovery following an external posterior perturbation when the individual is forced to rely on compensatory stepping. Instead these data suggest that the strongest factor in modifying step characteristics is experience with the perturbation. PMID- 15288454 TI - One-leg stance in healthy young and elderly adults: a measure of postural steadiness? AB - OBJECTIVE: To investigate postural steadiness during 30 s of one-leg stance in healthy young and elderly adults, by analysing the pattern of the ground reaction force variability. DESIGN: A laboratory set-up was used to analyse the variability of the ground reaction forces in relation to time as a measure of postural steadiness. BACKGROUND: The one-leg stance test is a measure considered to assess postural steadiness in a static position by a temporal measurement. The common notion is that a better postural steadiness, i.e. less force variability, allows for longer time standing on one leg. However, there is lack of evidence how postural steadiness during one-leg stance changes over time. METHODS: Twenty eight healthy elderly and 28 healthy young adults were tested by means of force plates assessing ground reaction forces while performing one-leg stance. RESULTS: During one-leg stance, two phases could be identified in both groups: First a dynamic phase, a rapid decrease of force variability, and thereafter a static phase, maintaining a certain level of force variability. During the first 5 s of one-leg stance the force variability decreased significantly more in the young group resulting in a lower force variability level during the static phase than in the elderly. CONCLUSIONS: The difficulties in maintaining the static position in elderly seems dependent on the reduced initial decrease in force variability and/or musculoskeletal components. We suggest that the first 5 s are crucial when assessing balance during one-leg stance. PMID- 15288455 TI - A conical-collared intramedullary stem can improve stress transfer and limit micromotion. AB - OBJECTIVE: The objective of this study was to quantify the effect of collar geometry on stress transfer and micromotion in idealized models of a cementless implant having an intramedullary stem. BACKGROUND: Intramedullary stems exist on several types of orthopaedic implants, including the femoral component of hip arthroplasties and segmental replacements used in the surgical treatment of a tumor or trauma in the diaphysis of a long bone. METHODS: Using three-dimensional finite element analysis, we compared four idealized, straight-stemmed, axisymmetric prostheses: flat-collared (0 degrees), conical-collared (30 degrees and 60 degrees), and collarless tapered (80 degrees). We simulated axial and non axial (20 degrees oblique) loads as well as non-ingrown and ingrown interface conditions. RESULTS: Without bone ingrowth, stress transfer to bone adjacent to the collar increased with collar angle. Micromotion at the distal stem increased moderately with collar angle from 0 degrees through 60 degrees, then increased markedly from 60 degrees to 80 degrees. With simulated bony ingrowth, the effect of the collar was greatly reduced. CONCLUSIONS: The results of this study suggest that the selection of collar angle represents a tradeoff between initial stress transfer and micromotion. Stems with conical collar angles in the range of 30-60 degrees can provide increased stress transfer compared to a flat collar design and reduced micromotion compared to a collarless tapered design. PMID- 15288456 TI - Patella kinematics and patello-femoral contact areas in patients with genu varum and mild osteoarthritis. AB - BACKGROUND: Patients with genu varum of the knee and moderate to severe osteoarthritis often suffer from additional symptoms of the patello-femoral joint. These patients have a poor prognosis following high tibial osteotomy. It is unclear whether varus knees with only mild femoro-tibial osteoarthritis are also associated with alterations of patella biomechanics, and affect the prognosis of intended high tibial osteotomy. METHODS: Fifteen patients with genu varum and mild osteoarthritis and 15 healthy volunteers were assessed in an open MRI-scanner. 3D-GRE sequences of the knee were obtained in 0 degrees, 30 degrees and 90 degrees with and without activity of the extensor muscles. After segmentation of patella, femur, tibia and the adjacent cartilage, a patella-based local coordinate system was established. Femoral and tibial reference points allowed definition of the spatial position of the patella. Contact areas were defined by intersection of opposing cartilage volumes. FINDINGS: No significant differences in patella kinematics and patello-femoral contact areas could be found (P > 0.05) between varus knees with mild osteoarthritis and healthy knees either at different flexion angles or under extending muscle activity. INTERPRETATION: In knees with genu varum and mild medial osteoarthritis we could detect no alterations in patello-femoral kinematics. Since the alterations of patients with genu varum and mild osteoarthritis are restricted to the medial femoro-tibial joint high tibial osteotomy might be successful. PMID- 15288457 TI - Performance specification for lower limb orthotic devices. AB - OBJECTIVE: To establish the range of forces and moments applied to lower limb orthoses during ambulation by routine users. DESIGN: Well-established gait analysis techniques were used to determine the loading at the major joints. It was assumed that the joint moments were transmitted by the orthosis encompassing any particular joint. Two hundred and five assessments of 164 patients were successfully completed by a consortium of four gait laboratories in Europe. The orthosis specification and patient clinical data were also recorded. BACKGROUND: The design and development of orthoses has occurred largely by evolution rather than by formal engineering methods. In particular, formal design has been hampered by a lack of information on the forces and moments applied during ambulation. METHODS: A standard gait analysis procedure was employed to capture the data. In-house biomechanical models were used to calculate the joint loading. Data were normalised with respect to patient weight and leg length. RESULTS: It was found that the median maximum normalised ankle moment transmitted by an ankle foot orthosis was 0.15 and the maximum knee moment was 0.09. The greatest moment transmitted by the hip joint of a hip knee ankle foot orthosis was also 0.09. There was a wide variation in the data due to differences in the impairments of the test subjects. CONCLUSION: It is possible to estimate the loads transmitted by an orthosis using established gait analysis procedures without the need for load measurement transducers. There is now a need both to collect a larger representative dataset and to perform validation studies with transducers. PMID- 15288458 TI - Influence of a mono-centric knee brace on the tension of the collateral ligaments in knee joints after sectioning of the anterior cruciate ligament--an in vitro study. AB - OBJECTIVE: To analyze the influence of knee bracing on the tension of the medial and lateral collateral ligaments in anterior cruciate ligament deficiency. DESIGN: The tension of the collateral ligaments in anterior cruciate ligament deficient knees was measured with and without knee bracing using an in vitro model. BACKGROUND: Anterior cruciate ligament deficiency increases the tension in both collateral ligaments at the knee joint. Therefore knee braces should reduce that tension increase. However, that effect has never been proven quantitatively. METHODS: After anterior cruciate ligament-transection, the forces of the medial (anterior/posterior part) and lateral collateral ligament were measured in ten fresh human cadaver knees at 0 degrees, 20 degrees, 40 degrees, 60 degrees, 80 degrees and 100 degrees of flexion, with and without application of a mono centric knee brace. To quantify the ligament forces, strain gauges were fixed at the bony origins of the ligaments. RESULTS: Bracing led to a significant decrease of ligament forces (20-100 degrees: P < 0.0001) in the anterior part of the medial collateral ligament in all joint positions. In the posterior aspect, this effect was observed only at 40 degrees (P < 0.0001) and 80 degrees (P = 0.001) of flexion. In the lateral collateral ligament, bracing caused a strain reduction from 60 degrees to 100 degrees of flexion (P < 0.0001). Therefore a flexion angle dependent effect of knee bracing on the strain was seen in the posterior aspect of the medial and in the lateral collateral ligament in anterior cruciate ligament deficient knee joints. CONCLUSIONS: Application of a mono-centric knee brace leads to a significant position dependent reduction of collateral ligament tension after anterior cruciate ligament-rupture. PMID- 15288459 TI - The effect of localized leg muscle fatigue on tibial impact acceleration. AB - OBJECTIVE: The objective of this study was to assess the effect of localized leg muscle fatigue on tibial acceleration following impact. BACKGROUND: Peak tibial accelerations measured just distal to the knee joint during running have been shown to increase with general body fatigue. However, the role that local leg muscle fatigue plays in shock attenuation is not clear. METHODS: The unshod, dominant foot of 24 healthy women in two different age groups was impacted into a vertical force plate, using a human pendulum method. Impact velocity and force approximated that found in running. EMG activity of the gastrocnemius and tibialis anterior muscles was monitored during fatiguing isometric exercise to assess fatigue, and quantified at impact as a measure of muscle activation level. A skin-mounted uni-axial accelerometer, was located at the tibial tubercle under pre-load, to measure tibial acceleration at impact. RESULTS: There was a significant decrease in peak tibial acceleration (P = 0.008) and acceleration slope (P = 0.033) following fatigue. There were no significant main effects or interactions for age group or muscle group for all tibial response variables. CONCLUSION: The reduction in peak tibial acceleration following fatigue (for the test performed) is contrary to the response documented following whole-body fatigue. PMID- 15288460 TI - Distraction-resisting force during tibial diaphyseal lengthening and consolidation--a study on a rabbit model. AB - BACKGROUND: Distraction-resisting force is generated in the soft tissues and callus during limb lengthening. Monitoring this force may offer a method of studying the behaviour of soft tissue and detecting the distraction osteogenesis related problems, and help to prevent complications. Changes in the post distraction period have not been previously investigated and there are no reports on the contribution of gastrocnemius to the distraction-resisting force. METHODS: Sixteen immature New Zealand White rabbits underwent 30% (left) tibial diaphyseal lengthening at a rate of two 0.4 mm increments per day. Using an instrumented bilateral fixator, the passive distraction-resisting force and the contribution made by gastrocnemius were measured at the end of lengthening and 5 weeks after lengthening. FINDINGS: The distraction-resisting force at the end of lengthening (mean 44 N (SD 10)) was statistically higher (p < 0.01) than that five weeks after lengthening (mean 20 N (SD 8)), so was the contribution of the gastrocnemius to the force (mean 11 N (SD 5 N) or 25% (SD 7) at the end of lengthening and 3 N (SD 1) or 13% (SD 5.2) five weeks later). INTERPRETATION: The callus rather than the surrounding muscles generates most of the passive DRF and its share of the force increased during consolidation period. PMID- 15288461 TI - Mechanical tests and finite element models for bone holding power of tibial locking screws. AB - OBJECTIVE: To investigate the bone holding power of tibial locking screws. DESIGN: The bone holding power was assessed by mechanical testing and finite element analysis. BACKGROUND: Screw loosening might threaten fracture fixation and bone healing. METHODS: In mechanical tests, six types of different tibial locking screws were inserted into low-density polyurethane foam tubes, which simulated osteoporotic bone. The screws were pushed out of the foam bone by an axial load, and the maximal pushout load was recorded. In finite element analysis, three-dimensional finite element models with a nonlinear contact interface between the screws and the bones were created to simulate the mechanical testing. The total strain energy of the bone and total reaction force of the screws were recorded. The contribution of the design factors was analyzed by the Taguchi method. RESULTS: In the mechanical tests, foam bone was stripped by the screw threads without screw deformation. The testing results were closely related to those of finite element analysis. The Taguchi analysis showed that the descending order of contribution of the design factors was outer diameter, pitch, half angle, and inner diameter. Root radius and thread width had minimal effects. CONCLUSIONS: The bone holding power of the screws could be reliably assessed by finite element models, which could analyze the effects of all the design factors independently and were potentially applicable to screws with irregular thread patterns. PMID- 15288462 TI - Avoiding the material nonlinearity in an external fixation device. AB - BACKGROUND: External fixation devices are widely used for treating unstable bone fractures because of their attractive features including minimal invasiveness, maximum tailorability, and extreme versatility. In one type of these devices (i.e., the fine-wire fixators), these unique features are made possible by the use of tensioned wires to support bone fragments. The major problem with these wires is their yielding. Once the wires yield, the fracture healing process will be adversely affected. A recent study showed that the nonlinear behavior observed in these tensioned wires can be geometric and material, and the geometric nonlinearity will stiffen the wires while the material nonlinearity will cause the wires to yield. This study is to investigate if it is possible to avoid the material nonlinearity in order to retain the elastic and repeatable performance for the wires. METHODS: Nonlinear and large deformation finite element analyses were conducted. Models of a bone segment transfixed by pairs of cross-aligned wires subjected to various levels of pre-tension were developed. The bone segment was subjected to a vertical load, and the load-displacement curves, wire tensions and wire tensile stresses were obtained under a full cycle of loading and unloading regime. FINDINGS: Pre-tensioning the wires is beneficial for stiffening a fixation device, but is disadvantageous to maintaining the wire elasticity. By limiting the level of the pre-tension, we can avoid the material nonlinearity. Doing so we will be able to stiffen the fixation device and retain elastic and predictable mechanical performance at the same time. INTERPRETATION: The findings will lead to a new paradigm toward enhancing the performance of external fixation devices. PMID- 15288463 TI - Initial stability of ankle arthrodesis with three-screw fixation. A finite element analysis. AB - OBJECTIVE: Compare the initial stability at the fusion site of ankle arthrodesis fixed with two and three screws. DESIGN: Finite element models of ankle arthrodesis were developed from computed tomography images. Two-screw constructs were augmented with a third screw in different orientations and subjected to loads likely to affect the ankle postoperatively. BACKGROUND: More stable fixation seems to increase the chance of fusion, as it minimises the motion between the tibiotalar interfaces. METHODS: Non-linear elastic finite element analyses were performed in external torsion and dorsiflexion. The micromotions at the tibiotalar interface were computed to compare the two- and three-screw fixation in intact and flat-cut arthrodesis. RESULTS: Adding a third screw reduced the micromotions at the fusion site. Inserting the third screw anteriorly predicted lower peak micromotions than inserting the screw posteriorly, except for the intact arthrodesis tested in dorsiflexion. Three-screw intact arthrodesis predicted lower peak micromotions than flat-cut arthrodesis. CONCLUSIONS: Better stability was predicted for three-screw ankle arthrodesis. In flat-cut arthrodesis, a third screw inserted anteriorly performed better than a posterior screw. In intact arthrodesis, a posterior screw seemed a better option when flexion stability was the main concern. Even with three-screw fixation, the configuration of the first two-crossed screws may still be important to improve the stability at the fusion site. PMID- 15288464 TI - Ankle joint evertor-invertor muscle torque ratio decrease due to recurrent lateral ligament sprains. AB - OBJECTIVE: To determine an ankle inversion-eversion movement range part where evertor/invertor muscles torque ratio is altered after recurrent ankle lateral ligament sprains. DESIGN: The ankle evertor/invertor muscles torque ratios were determined in the different range parts of angular positions by the muscle isokinetic movements. BACKGROUND: It is important to determine the movement range part where the evertor muscle weakness after the ankle lateral ligament sprains is more expressed and a repeated trauma of the lateral ligaments is more probable. METHODS: Twenty-eight male handball players participated in the tests using an ankle isokinetic inversion-eversion movement investigation dynamometer system. Thirty-three ankle joints were uninjured, but 23 underwent recurrent lateral ligament sprains. RESULTS: The ankle evertor/invertor muscles torque ratio for the sprained ankles was significantly lower in comparison with the uninjured joints in inversion positions at 50 degrees and 60 degrees of the range of movements in all applied velocities, except the slowest movement (30 degrees /s). CONCLUSIONS: The recurrent ankle lateral ligament sprains reduced the evertor/invertor muscles torque ratio in the inversion positions of the range of movements and therefore the evertor muscle weakness was more expressed in the beginning of the eversion movement. PMID- 15288465 TI - Familial amyloid polyneuropathy and liver transplantation. PMID- 15288466 TI - Gallstone disease: physicochemical research sheds new light on an old disease and points the way to medical therapy. PMID- 15288467 TI - Differential expression of bile salt and organic anion transporters in developing rat liver. AB - BACKGROUND/AIMS: Differentiated hepatocytes express distinct transport systems at their basolateral and canalicular membrane domains. Here, we investigated the ontogenesis of the polar expression of hepatocellular organic anion and bile salt transport systems in rat liver. METHODS: mRNA levels (real time PCR) and protein expression (immunofluorescence microscopy) were investigated for the Na(+) taurocholate cotransport protein (Ntcp), the organic anion transporting polypeptides (Oatp1a1, Oatp1a4, Oatp1b2), the multidrug resistance associated proteins (Mrp2, Mrp6) and the bile salt export pump (Bsep). RESULTS: Expression of mRNA and protein was detected first for Oatp1b2, Mrp2 and Mrp6 at embryonic day 16 (E16), followed by Ntcp, Oatp1a1 and Bsep at E20 and by Oatp1a4 at postnatal day 5 (P5). Intracellular localization of Oatps (e.g. Oatp1b2) preceded expression at the plasma membrane. Approximate adult phenotypes of polarized expression were achieved for Ntcp by P5, for Bsep, Mrp2 and Mrp6 by P12 and for Oatp1a1, Oatp1a4 and Oatp1b2 by P29. CONCLUSIONS: The data demonstrate that full maturation of polarized transporter expression in rat liver requires several weeks. The findings provide a molecular explanation for the previously observed chronology of the functional maturation of bile salt-independent and dependent bile formation and of hepatic detoxification functions in developing rat liver. PMID- 15288468 TI - Association of matrix metalloproteinase-1 and -3 promoter polymorphisms with clinical subsets of Norwegian primary sclerosing cholangitis patients. AB - BACKGROUND/AIMS: Primary sclerosing cholangitis (PSC) is considered an immune mediated liver disease of multifactorial and multigenetic aetiology. Concomitant ulcerative colitis (UC) is seen in many PSC patients, but the pathogenetic link between these disorders is unknown. Due to association with inflammation, fibrosis, and cancer development, the matrix metalloproteinases MMP-1 and MMP-3 are candidate genes for predisposition to both PSC, UC and cholangiocarcinoma. METHODS: We investigated the association of MMP-1 and MMP-3 promoter polymorphisms in 165 Norwegian PSC patients compared to 118 UC patients and 346 healthy controls. RESULTS: There were no differences in MMP-1 and MMP-3 frequencies between PSC patients and UC patients or healthy controls. PSC patients with UC showed an increased frequency of the MMP-3 allele 5A compared to PSC patients without UC (60% vs. 45%; P(c)=0.01). All patients (100%) with cholangiocarcinoma carried MMP-1 allele 1G, compared to only 72% of PSC patients without cholangiocarcinoma. CONCLUSIONS: We found no general associations of the MMP-1 and MMP-3 genes to PSC or UC among Norwegian patients, but specific alleles were associated to subsets of PSC patients with UC and cholangiocarcinoma. The results support the theory of genetic heterogeneity among PSC patients. PMID- 15288469 TI - Colonization of albumin-producing hepatocytes derived from transplanted F344 rat bone marrow cells in the liver of congenic Nagase's analbuminemic rats. AB - BACKGROUND/AIMS: We investigated whether bone marrow cells (BMCs) of normal rats can be transformed in albumin-producing hepatocytes in analbuminemic rat livers. METHODS: BMCs (2 x 10(7)) from F344 rats (F344) were infused via the portal vein into the livers of congenic Nagase's analbuminemic rats (F344alb) immediately after 70% hepatectomy (PH). Alternatively, F344alb were hematopoietically reconstituted with F344 BMCs by whole body irradiation and BMC transplantation before PH. The recipients were examined for albumin positive (alb +) hepatocytes and albumin mRNA in the livers as well as serum albumin levels 4 weeks later. Sry3 in situ hybridization was done for the livers of female F344alb that received male F344 BMCs. RESULTS: Livers of untreated F344alb contained a few single and double alb+hepatocytes, but these did not form clusters after PH. Clusters (>3 alb + hepatocytes) were detected in livers of the recipients which were transplanted with BMCs immediately after PH as well as the reconstituted F344alb with or without PH. Normal albumin mRNA was detected in the recipient livers, and serum albumin levels were increased. Sry3 was identified in the alb+clusters in the female recipients. CONCLUSIONS: Transplanted BMCs from normal rats can increase clusters of albumin-producing hepatocytes within the liver of analbuminemic rats. PMID- 15288470 TI - Long term transgene expression by hepatocytes transduced with retroviral vectors requires induction of immune tolerance to the transgene. AB - BACKGROUND/AIMS: Gene therapy for inherited liver diseases requires permanent expression of the therapeutic gene. However, in vivo liver transduction with retroviral vectors triggers an immune elimination of transduced hepatocytes. Here we investigated whether immune response could be prevented by treatment with compounds known to induce tolerance in organ transplantation: CTLA4Ig and LF-15 0195. METHODS: CTLA4Ig was administered either via i.p. injection of the drug or by i.m. injection of recombinant adenoviruses encoding CTLA4Ig. LF-15-0195 was administered i.p. All animals were subjected to partial hepatectomy and received beta-galactosidase retroviral vectors intravenously. Appearance of anti-beta galactosidase antibodies was monitored and the number of positive hepatocytes was assessed at day 7 and at sacrifice. RESULTS: No beta-galactosidase antibodies were detected as long as CTLA4Ig was detectable in serum. Short-term treatment with CTLA4Ig induced tolerance in a significant proportion of animals only at high dose (1 mg/kg). Administration of CTLA4Ig adenovectors resulted in prolonged secretion of CTLA4Ig and permanent absence of anti-beta-galactosidase antibodies. LF-15-0915 administration achieved tolerance in some animals. CONCLUSIONS: In conclusion, manipulation of the immune system at the time of virus delivery using clinically relevant tolerance-inducing protocols is a promising approach to achieve long term expression after retrovirus-mediated gene transfer to the liver. PMID- 15288471 TI - Soluble TNF-R1, but not tumor necrosis factor alpha, predicts the 3-month mortality in patients with alcoholic hepatitis. AB - BACKGROUND/AIMS: In alcoholic hepatitis (AH), soluble TNF alpha receptor-1 (sTNF R1) is increased. Elevated TNF alpha predicts mortality, but infection influences TNF alpha values. In patients with AH, we determined the prognostic value of TNF alpha, sTNF-R1, and lipopolysaccharide binding protein (LBP) and CD14, both involved in endotoxemia-associated inflammation. METHODS: One hundred and eight cirrhotic patients (Pugh score 10 [6-13]) and biopsy-proven AH (Maddrey's DF <32: n=46; > or =32: n=62) without associated infection were included within 8 days of admission and followed-up for 3 months. Cytokines were measured using specific immunoassays. Patients with severe AH received steroids. RESULTS: Twenty four patients died at a median time of 35 days (range: 3-89). The overall survival was 78%. Multivariate Cox regression analysis showed that sTNF-R1 was an independent predictor of mortality, (OR 4.33: 95% CI [1.12-16.75]). Pugh's score (P=0.618), Maddrey's DF (P=0.182), creatinine (P=0.197), TNF alpha (P=0.319), LBP (P=0.362), and CD14 (P=0.347) were not related to survival. CONCLUSIONS: In patients with AH, sTNF-R1 measured at admission is an independent predictor of survival at 3 months. Provided that TNF-R1 mediates the cytotoxic actions of TNFalpha, these results support the concept of dysregulated TNF alpha metabolism in AH. PMID- 15288472 TI - Treatment of thioacetamide-induced liver cirrhosis by the Ras antagonist, farnesylthiosalicylic acid. AB - BACKGROUND/AIMS: Several studies have indicated increased expression of the Ras protooncogenes in liver cirrhosis. In a previous study in rats, we have shown that a synthetic Ras antagonist, S-farnesylthiosalicylic acid (FTS), could inhibit the development of liver cirrhosis. The aim of the current study was to examine whether FTS will accelerate the resolution of liver cirrhosis induced in rats by thioacetamide. METHODS: Cirrhosis was induced in male Wistar rats by intraperitoneal (i.p.) administration of thioacetamide (200 mg/kg twice weekly for 12 weeks). In the treated group, the Ras antagonist FTS (5 mg/kg, i.p./3 times/week) was administered for 8 weeks after liver cirrhosis has already been established. Control cirrhotic rats received PBS injections for 8 weeks. RESULTS: Rats treated with FTS for 8 weeks had lower histopathologic score of fibrosis (P = 0.01), lower hepatic hydroxyproline levels (P = 0.0002) and lower spleen weight (P = 0.02) than the cirrhotic rats treated with PBS. Following FTS treatment, the MMP-2 and MMP-9-induced collagenolytic activity and TIMP-2 expression, were increased in FTS-compared to PBS-treated rats. TUNEL assay of liver sections performed 8 weeks after thioacetamide withdrawal showed increased apoptotic figures in both groups (P = NS). CONCLUSIONS: These results indicate that the Ras antagonist FTS accelerates the regression of experimentally-induced hepatic cirrhosis. The mechanism may involve increased collagenolytic activity. PMID- 15288473 TI - Herb medicine Inchin-ko-to (TJ-135) regulates PDGF-BB-dependent signaling pathways of hepatic stellate cells in primary culture and attenuates development of liver fibrosis induced by thioacetamide administration in rats. AB - BACKGROUND/AIMS: We studied the effect of Inchin-ko-to (TJ-135), a herb medicine that has been clinically used for liver cirrhosis in Japan, on liver fibrosis in a rat model and on the function of stellate cells. METHODS: Rat liver fibrosis was generated by thioacetamide (TAA) administration. DNA synthesis was assessed by 5-bromo-2'-deoxyuridine incorporation assay. Protein expression was analysed by western blotting. Collagen and fibronectin mRNA expression were analysed by reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: TJ-135 improved liver fibrosis induced in rats by TAA administration. TJ-135 reduced collagen deposition and the expression of smooth muscle alpha-actin in fibrotic liver tissues and decreased the serum level of hyaluronic acid. In primary cultured stellate cells, TJ-135 suppressed DNA synthesis and the expression of collagen alpha 1(I), collagen III, and fibronectin mRNAs. It hampered DNA synthesis and migration of PDGF-BB-stimulated stellate cells through inhibiting phosphorylation of PDGF receptor-beta and down-stream signaling pathways. Among TJ-135 components, 3-methyl-1,6,8-trihydroxyanthraquinone (emodin) derived from Rhei rhizoma was found to be the most active molecule. CONCLUSIONS: TJ-135 and emodin regulate PDGF-dependent events in stellate cells and attenuate the development of liver fibrosis. Their clinical use may be beneficial for the therapy of human liver fibrosis. PMID- 15288474 TI - Prostaglandin E2 inhibits transforming growth factor beta 1-mediated induction of collagen alpha 1(I) in hepatic stellate cells. AB - BACKGROUND/AIMS: Cyclooxygenase-2 (COX-2) has been implicated in a number of hepatic stellate cell (HSC) functions but its relationship to transforming growth factor-beta 1 (TGF-beta 1)-mediated fibrogenesis is unknown. We assessed the impact of COX-2 inhibition and PGE(2) on the regulation of TGF-beta 1-stimulated matrix synthesis in an immortalized human HSC line, LX-1 and corroborated these findings in primary stellate cells. METHODS: Expression of COX-2 was assessed by Western blotting and real time quantitative PCR. The effect of NS398, a selective COX-2 inhibitor, and PGE(2) on TGF-beta 1-mediated fibrogenesis was examined by measuring mRNA levels of collagen alpha1(I). PGE(2) receptor expression was analyzed by RT-PCR. RESULTS: Under basal conditions, NS398 suppressed PGE(2) synthesis and induced collagen alpha 1(I) whereas exogenous PGE(2) suppressed expression of collagen alpha1(I). TGF-beta 1 induced COX-2 mRNA, COX-2 protein and PGE(2) biosynthesis. Importantly, TGF-beta 1-mediated induction of collagen alpha 1(I) was markedly suppressed by the addition of exogenous PGE(2). All four major PGE(2) receptors were expressed in LX-1 cells. CONCLUSIONS: These results suggest that COX-2-derived PGE(2) inhibits both basal and TGF-beta 1-mediated induction of collagen synthesis by HSC. Based on these findings, it will be important to determine whether inhibiting COX-derived PGE(2) synthesis alters the progression of liver fibrosis in vivo. PMID- 15288475 TI - Endothelial cell differentiation in hepatocellular adenomas: implications for histopathological diagnosis. AB - BACKGROUND/AIMS: Little information is available about the patterns of endothelial cell differentiation observed in hepatocellular adenomas. We therefore aimed to analyze the endothelial cell immunophenotype in a large series of these tumors and evaluate its possible diagnostic relevance. METHODS: The expression of continuous and sinusoidal endothelial cell markers and of extracellular matrix proteins was analyzed by immunoperoxidase in 56 adenomas, as compared with 30 cases of focal nodular hyperplasia (FNH), 2 cases of telangiectatic FNH and 40 cases of hepatocellular carcinoma (HCC). RESULTS: Twenty-eight adenomas (50%) presented a sinusoidal pattern of endothelial cell differentiation, characterized by the expression of specific sinusoidal endothelial cell markers and the presence of a subendothelial matrix resembling the normal perisinusoidal matrix. Eleven tumors (19.5%) presented a continuous pattern of endothelial cell differentiation. Seventeen tumors (30.5%) showed a mixed pattern. Twenty-eight FNH presented a sinusoidal pattern of endothelial cell differentiation; all HCC presented a continuous pattern of endothelial cell differentiation. CONCLUSIONS: In hepatocellular adenomas, intra-tumoral vessels usually present a sinusoidal pattern of endothelial cell differentiation. However, the vascular phenotypic heterogeneity observed in our study questions the potential relevance of endothelial cell immunophenotyping for the differential diagnosis between hepatocellular adenomas and well differentiated HCC. PMID- 15288476 TI - SU5416 is a potent inhibitor of hepatocyte growth factor receptor (c-Met) and blocks HGF-induced invasiveness of human HepG2 hepatoma cells. AB - BACKGROUND/AIMS: SU5416 is a potent inhibitor of receptor tyrosine kinases, including those of the vascular endothelial growth factor receptor, stem cell factor receptor, and platelet-derived growth factor receptor. Because of the overwhelming evidence favoring the role of aberrant hepatocyte growth factor (HGF)/Met signaling in the pathogenesis of various human cancers, various inhibitor strategies have been employed to therapeutically target this receptor. METHODS: Cell proliferation was determined by incorporation of [(3)H] thymidine. Invasiveness was assayed in Boyden Chambers with 8 microm Matrigel coated filters. Phosphorylation of ERK1/2, Akt by HGF stimulation was detected by Western blotting. RESULTS: We found that SU5416 inhibited motility scattering and the invasive activity of a hepatocellular carcinoma cell line HepG2 in vitro and growth in primary cultured hepatocytes induced by HGF. Consequently, tyrosine autophosphorylation of the c-met induced by HGF was inhibited in these cells by SU5416 in a dose-dependent manner. Furthermore, ERK1/2 and Akt phosphorylation, the signaling events down-stream of c-met activation were reduced. Moreover, SU5416 caused reversion in NIH3T3 fibroblasts transformed by the oncogenic form of the receptor, Tpr-Met. CONCLUSIONS: Inhibition of various solid tumors growth and metastasis by SU5416 may be partially attributed to blocking activation of the hepatocyte growth factor receptor. PMID- 15288477 TI - Disruption of hepatocellular tight junctions by vascular endothelial growth factor (VEGF): a novel mechanism for tumor invasion. AB - BACKGROUND/AIMS: Vascular endothelial growth factor (VEGF) is expressed by many tumors, including hepatocellular carcinoma (HCC) and is involved in tumor angiogenesis. Little is known about its role for HCC infiltration into normal liver parenchyma. METHODS: The effects of VEGF on the integrity of tight junctions were studied in HepG2 cells and human HCC by means of confocal laser scanning microscopy. RESULTS: VEGF induced within 45 min a marked loss of pseudocanaliculi and disruption of occludin-delineated tight junctions. This effect of VEGF was mimicked by phorbol-12-myristate-13-acetate (PMA) and was sensitive to protein kinase C (PKC) inhibition by Go6850. VEGF induced within 15 min the translocation of the PKC alpha-isoform to the plasma-membrane, but had no effect on the activity of Erks and p38(MAPK). Sections from surgically removed HCC showed expression of VEGF in the tumor and occludin disassembly in normal liver parenchyma next to the tumor. CONCLUSIONS: VEGF induces disruption of tight junctions in a PKC-alpha dependent manner. In addition to its known angioneogenic properties, VEGF may promote HCC spreading into normal liver parenchyma. The data may provide another rationale for the use of VEGF antagonists for tumor therapy. PMID- 15288478 TI - Molecular-based prediction of early recurrence in hepatocellular carcinoma. AB - BACKGROUND/AIMS: Hepatocellular carcinoma (HCC) has a very poor prognosis, due to the high incidence of tumor recurrence. As the current morphological indicators are often insufficient for therapeutic decisions, we sought to identify additional biologic indicators for early recurrence. METHODS: We analyzed gene expression using a PCR-based array of 3,072 genes in 100 HCC patients. Informative genes predicting early intrahepatic recurrence were selected by random permutation testing, and a weighted voting prediction method was constructed. Following estimation of prediction accuracy, a multivariate Cox analysis was performed. RESULTS: By permutation testing, we selected 92 genes demonstrated distinct expression patterns differing significantly between recurrence cases and recurrence-free cases. Our prediction method, using the 20 top-ranked genes, correctly predicted the early intrahepatic recurrence for 29 of 40 cases within the validation group, and the odds ratio was 6.8 (95%CI 1.7-27.5, P = 0.010). The 2-year recurrence rates in the patients with the good signature and those with the poor signature were 29.4 and 73.9%, respectively. Multivariate Cox analysis revealed that molecular-signature was an independent indicator for recurrence (hazard ratio 3.82, 95%CI 1.44-10.10, P = 0.007). CONCLUSIONS: Our molecular-based prediction method using 20 genes is clinically useful to predict early recurrence of HCC. PMID- 15288479 TI - Clinical and molecular analysis of combined hepatocellular-cholangiocarcinomas. AB - BACKGROUND/AIMS: Combined hepatocellular-cholangiocarcinoma (HCC-CC) show dual hepatocellular and biliary epithelial differentiation. To better understand the relations between cholangiocarcinoma (CC), HCC-CC and hepatocellular carcinoma (HCC), we screened for genetic alterations. METHODS: A series of nine CC, 15 HCC CC and three separated HCC and CC lesions ('collision tumors') were screened for loss of heterozygosity (LOH) using 400 microsatellite markers and for p53 and beta-catenin mutations. A comparison with a previously characterized series of 137 HCC was performed. RESULTS: In six cases of CC and HCC-CC, we identified TP53 gene mutations. A CTNNB1/beta-catenin was identified in two patients presenting collision tumors, but no mutations were found in CC or in HCC-CC. A high level of chromosome instability in both CC and HCC-CC was found. Recurrent specific LOH were identified at 3p and 14q in more than 50% of the CC and the HCC-CC cases, whereas these chromosomal regions were deleted in less than 10% of the HCC cases (P<10(-5)). Minimal common regions of deletion (MCRD) were defined at 3p24-p14 and 14q24-q32, respectively. CONCLUSIONS: These results suggest that combined HCC CC are genetically closer to CC than HCC and common carcinogenesis pathways may be altered in HCC-CC and CC. PMID- 15288480 TI - The delay of rearterialization after initial portal reperfusion in living donor liver transplantation significantly determines the development of microvascular graft dysfunction. AB - BACKGROUND/AIMS: Graft reperfusion in liver transplantation is usually performed by initial portal reperfusion (IPR) and delayed rearterialization. Its influence on graft microcirculation is unknown. This study aimed to assess sinusoidal perfusion in dependence to this reperfusion technique during human living-donor liver transplantation. METHODS: Hepatic microcirculation was measured both in the donor and the recipient (n=14) by using the orthogonal polarization spectral imaging technique. By using initial portal reperfusion, the mean time interval between portal venous and hepatic arterial reperfusion was 27.7+/-13.3 min. RESULTS: Hepatic nutritive perfusion, as given by the functional sinusoidal density and the volumetric blood flow, was found significantly decreased during portal reperfusion when compared to baseline. Rearterialization resulted in hyperperfusion of individual sinusoids at a decreased density of the sinusoidal network. Interestingly, the time interval between portal venous and hepatic arterial reperfusion significantly correlated with the changes of the liver grafts' microcirculation. CONCLUSIONS: The study indicates graft microcirculatory dysfunction as a major determinant of postischemic liver injury. Moreover, microvascular impairment was significantly influenced by the interval between portal venous and hepatic arterial reperfusion, which suggests the reinforcement of the pathomechanism of injury involving hypoxia and rapid graft rewarming due to initial portal reperfusion. PMID- 15288481 TI - Evaluation of donor hepatic iron concentration as a factor of early fibrotic progression after liver transplantation. AB - BACKGROUND/AIMS: Hepatic iron may act as an important co-morbid factor in non hemochromatotic liver diseases, but whether it may favour fibrogenesis after liver transplantation is not known. To verify whether the hepatic iron concentration of the graft might play a role in the rapid fibrotic progression frequently observed after liver transplantation for chronic hepatitis C. METHODS: The hepatic iron concentration, measured at the time of the donor operation, was retrospectively related to the histological follow-up data of 68 recipients (49 males, 19 females), of whom 38 were hepatitis C virus positive. RESULTS: The hepatic iron concentration in donor liver biopsies ranged from 25 to 7,100 microg/gdw. After a median follow-up of 19 months, nine patients (five HCV positive) had a staging score >3. There was a significant association between a higher frequency of increasing staging and donor age >50 years. In female HCV positive recipients, a graft hepatic iron concentration >1,200 microg/gdw was associated with fibrosis progression >0.15 fibrosis units per month (4/4 vs. 1/7, p<0.01). CONCLUSIONS: The graft hepatic iron concentration may be one of the factors involved in early fibrosis progression due to recurrent hepatitis C in female recipients. PMID- 15288482 TI - A randomized, controlled trial of triple antiviral therapy as initial treatment of chronic hepatitis C in HIV-infected patients. AB - BACKGROUND/AIMS: Interferon and ribavirin combination therapy for chronic hepatitis C induces a low response rate in human immunodeficiency virus (HIV) infected patients. To assess the impact of intensification of interferon administration and of the addition of amantadine on the efficacy and safety of standard anti-hepatitis C virus (HCV) treatment in HIV-infected patients. METHODS: Multicentre, prospective, open-label, randomized, phase III clinical trial. Eighty co-infected patients were randomized to receive ribavirin 800-1,000 mg/day in combination with, group A: interferon alpha 2a 3MIU thrice weekly; group B: IFN alpha 2a 3MIU daily, plus amantadine 200 mg/day; treatment duration was 24-48 weeks according to HCV genotype. RESULTS: Forty-one patients were randomized in group A and 39 in group B. Intention-to-treat analysis showed a sustained virological response, defined as HCV-RNA negativization, 6 months after stopping treatment in 22% of patients from group A and 13% from group B (P>0.05). The lack of a 2-log drop in HCV-RNA levels after 12 weeks of treatment showed a 100% predictive value of lack of sustained response. CONCLUSIONS: Amantadine addition and interferon intensification do not improve the low efficacy of combination of interferon alfa plus ribavirin in HIV/HCV co-infected patients. Patients with no early virologic response did not have any probability of sustained response. PMID- 15288483 TI - A study of some hepatic immunological markers, iron load and virus genotype in chronic hepatitis C. AB - BACKGROUND/AIMS: Host factors that may influence progression of hepatitis C infection to chronic hepatitis include T-cell responses and iron accumulation. We evaluated the hepatic expression of immunological markers relevant for a cytotoxic response in relation to viral and HFE genotype. METHODS: Frozen liver biopsies were obtained at diagnosis from 28 HFE genotyped patients. Sections stained for CD8, MHC-I, beta(2)m, HFE and CD68 were analyzed blind by morphometry. Response to therapy was available in 12 cases. RESULTS: A negative correlation was found between the number of CD8(+) cells and fibrosis. CD8(+) cells localized as clusters in portal tracts and sinusoids and were seen interacting with MHC-I positive lining cells. MHC-I and beta(2)m were expressed mainly in the endothelial and Kupffer cells. HFE was expressed in most, but not all, round and dendritic CD68(+) cells. Patients with virus genotype 3a had higher hepatic MHC-I and HFE expression, and a better-sustained response to IFN therapy than other patients. CONCLUSIONS: In chronic hepatitis C virus infection MHC-I expression in the liver seems to relate to viral-genotype. In addition, the expression of MHC-I molecules by Kupffer cells places them as probable important players in the host response to HCV. PMID- 15288484 TI - Genetics of autoimmune and viral liver diseases; understanding the issues. PMID- 15288485 TI - Gene expression profiles in hepatocellular carcinoma: not yet there. PMID- 15288486 TI - Prognostic models including the Child-Pugh, MELD and Mayo risk scores--where are we and where should we go? PMID- 15288487 TI - Diagnosing of peliosis hepatis by magnetic resonance imaging. PMID- 15288488 TI - Oral glucose tolerance deteriorates in rats fed with methionine choline deficient diet. PMID- 15288490 TI - The rising health and economic burden of chronic hepatitis C. PMID- 15288493 TI - Executive and prefrontal dysfunction in unipolar depression: a review of neuropsychological and imaging evidence. AB - This paper reviews recent empirical findings related to prefrontal and executive function in unipolar depression. While a number of reviews have dealt with either the neuropsychological literature or findings from imaging studies, the present review addresses both, as well as findings from studies that have combined brain imaging techniques with neuropsychological measures. This combined approach is of great interest as the performance of a structured task may act to load the areas of interest and reduce variance, thus making the imaging evidence more valuable; while the use of imaging provides a check that the neuropsychological tasks are indeed engaging the structures whose performance they are intended to assess. Prominent models of the neurobiology of depression implicate involvement of the anterior cingulate cortex (ACC) and the dorsolateral prefrontal cortex (DLPFC). The evidence from combined imaging and neuropsychological studies supports the involvement of the ACC, but is less clear in the case of the DLPFC. However, the limited number of such studies conducted to date means that conclusions must be tentative and further studies employing this combined approach may be of great value. PMID- 15288494 TI - Development of stellate and basket cells and their apoptosis in mouse cerebellar cortex. AB - Stellate and basket cells in the molecular layer (ML) of the cerebellar cortex proliferate within the white matter (WH) during development. Developmental neuronal death has been documented on granule cells but has not been demonstrated on other GABAergic neurons. We investigated the migration and the cell death of stellate/basket cells further in glutamic acid decarboxylase 67/green fluorescent protein (GFP) knock-in mouse in which every GABAergic neuron was identified by its GFP fluorescence. Analyses were made in the first three postnatal weeks. In the WM, GFP-positive cells were abundant on postnatal day (P) 5-15 but scarce in P21. Stellate/basket cells increased in number in the ML until P15, corresponding to the growth of the ML. Administration of 5-bromo-2'deoxyuridine (BrdU) at P2-8 labeled many cells in the WM within 1h. After BrdU administration at P5, many BrdU-labeled GFP-positive cells were observed in the WM and the internal granular layer at P7, and in the ML at P9. These results support the proliferation of stellate/basket cells in the WM and their migration to the ML. Apoptosis of GABAergic interneurons was demonstrated in the ML and WM during the first two weeks. Their apoptotic loss will contribute to the adjustment of neuron number or elimination of any improper populations. PMID- 15288495 TI - Estrogen reduces the neurite growth of serotonergic cells expressing estrogen receptors. AB - Serotonergic innervation of the central nervous system has a sexual dimorphism. The serotonin level in the hypothalamus was modulated by estrogen, and the formation of sexual dimorphism of serotonergic fiber innervation in the hypothalamus has been shown by the effect of sexual hormone during the critical perinatal period. In this study, we examined the direct effect of estrogen on the neurite growth of serotonergic neurons in primary culture from embryonic day 14 (E14) of rat mesencephalon. The total neurite length of serotonin-immunoreactive (IR) cells was significantly decreased by estradiol benzoate (E2, 10(-8)M) treatment for 7 days, compared with the case of no treatment. Moreover, the presence of estrogen receptor (ER) alpha and ERbeta mRNA in the E14 mesencephalon with reverse transcription-polymerase chain reaction (RT-PCR), and the ERalpha or ERbeta protein in the cultured serotonin-IR cells with double fluorescence immunohistochemistry were also demonstrated. Our results suggest that the inhibitory effects of E2 on the neurite growth of serotonergic cells expressing ERalpha or ERbeta might be involved in the formation of the sexual dimorphic distribution of serotonergic innervation. PMID- 15288496 TI - Striosomes are enriched in glutamic acid decarboxylase in primates. AB - The compartmental distribution of glutamic acid decarboxylase (GAD) in the striatum was investigated in squirrel monkeys and rats with antibodies raised against the two isoforms of this enzyme (GAD65 and GAD67) and with calbindin D 28k (CB) and/or micro-opiate receptor (MOR) as striosomal markers. In primates, immunostaining for both GAD65 and GAD67 was much more intense in striosomes than in the surrounding matrix. A thin immunoreactive strip of GAD labeling was also present in the dorsolateral part of both caudate nucleus and putamen. This narrow band appears to correspond to the so-called subcallosal streak (SS) found in rodent striatum. Although the immunostaining intensity for the two enzymes was similar at pallidal level, that for GAD65 was more intense than that for GAD67 at the striatal level. The GAD immunostaining was more uniformly distributed in the rat striatum, which did not display GAD-rich patches that corresponded to MOR positive striosomes. Moreover, in contrast to the findings obtained in monkeys, the subcallosal streak in rats was less intensely stained for GAD than for the remaining regions of the striatum. These results reveal that GAD65 and GAD67 are faithful markers of striosomes in primates but not in rodents. They suggest the existence of a significant species difference between rodents and primates in respect to the chemical organization of the striatum, a difference that should be taken into account when using rodents as animal models to study the functional organization of the basal ganglia and the pathogenesis of neurodegenerative diseases that affect the striatum. PMID- 15288497 TI - Seizure susceptibility to various convulsant stimuli in dystrophin-deficient mdx mice. AB - In the present study, the susceptibility of the mdx mouse, a dystrophin-deficient genetic model of Duchenne muscular dystrophy (DMD), to various convulsant stimuli has been evaluated and compared to three related mice strains (C57BL/6J, C57BL/10 and DBA/2 mice). Animals were treated with chemical convulsants impairing gamma aminobutyric acid (GABA) neurotransmission [pentylenetetrazole, picrotoxin, bicuculline, methyl-6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate (DMCM), methyl-beta-carboline-3-carboxylate (beta-CCM)], enhancing glutamatergic neurotransmission [N-methyl-d-aspartate (NMDA), alpha-amino-3-hydroxy-5-methyl-4 isoxazole propionate (AMPA) and kainic acid (KA)] or a K(+) channel blocker (4 aminopyridine). Occurrence of clonic and/or tonic seizures was evaluated to observe possible differences in seizure susceptibility. In addition, all strains of mice were repeatedly treated with a subconvulsant dose of pentylenetetrazole (PTZ) for possible differences in kindling development. The mdx mice exhibited no difference in seizure susceptibility for all convulsant drugs with the exception of a significantly lower sensitivity to AMPA and KA than the other mice strains. This study demonstrates that mdx mice possess a decreased susceptibility to some convulsant stimuli. However, mdx mice showed an enhanced seizure severity and a shorter latency in the development of chemical kindling produced by administration of PTZ. The present data suggests that the dystrophin deficiency in mdx mice affects the pathophysiology and pharmacology of acute and chronic epileptic seizures in an opposite manner. PMID- 15288498 TI - Intrinsic signal recording from a monkey whose behavior was maintained by a schedule of reinforcement. AB - Optical recording of cortical activity in awake monkeys has enhanced our understanding of the functional anatomy of the primary visual cortex (V1). However, cortical representation of visual cognition has not been studied by optical recording, even though the greatest merit of using awake animals is that they can offer advantages in studying cognitive function that anesthetized animals cannot. Thus far, the optical recording method has not been combined with tasks that accompany body movements because of concern about movement noise, although behavioral tasks are helpful in the study of animal cognition. Here, I tested the influence of body movements during the signal acquisition period on the resultant images. I recorded the intrinsic signals associating with different orientations from V1 of a monkey who was emitting behavior during the signal acquisition period. Although the monkey's behavior was maintained on a variable interval schedule that typically induces a high rate of response, orientation maps were consistently obtained. Therefore, a successful recording under this operant regimen implies the applicability of the optical recording method to other behavioral tasks. Several constraints in applying optical recording to studies using behaving animals are also discussed. PMID- 15288499 TI - A morphometric study of the progressive changes on NADPH diaphorase activity in the developing rat's barrel field. AB - The distribution of NADPH diaphorase (NADPH-d)/nitric oxide synthase (NOS) neurons was evaluated during the postnatal development of the primary somatosensory cortex (SI) of the rat. Both cell counts and area measurements of barrel fields were carried out throughout cortical maturation. In addition, NADPH d and cytochrome oxidase (CO) activities were also compared in both coronal and tangential sections of rat SI between postnatal days (P) 10 and 90. Throughout this period, the neuropil distributions of both enzymes presented a remarkable similarity and have not changed noticeably. Their distribution pattern show the PMBSF as a two-compartmented structure, displaying a highly reactive region (barrel hollows) flanked by less reactive regions (barrel septa). The number of NADPH-d neurons increased significantly in the barrel fields between P10 and P23, with peak at P23. The dendritic arborization of NADPH-d neurons became more elaborated during barrel development. In all ages evaluated, the number of NADPH d cells was always higher in septa than in the barrel hollows. Both high neuropil reactivity and differential distribution of NADPH-d neurons during SI development suggest a role for nitric oxide throughout barrel field maturation. PMID- 15288501 TI - Immunohistochemical and immunoblot study of GABA(A) alpha1 and beta2/3 subunits in the prefrontal cortex of subjects with schizophrenia and bipolar disorder. AB - A number of investigations have provided a growing body of evidence of the involvement of the gamma-aminobutyric acid (GABA) transmitter system in the pathophysiology of schizophrenia and bipolar disorder. In this study, immunohistochemical and immunoblot techniques were employed in order to examine alterations of the GABA(A) receptor alpha1 and beta2/3 subunits in the prefrontal cortex from postmortem subjects with schizophrenia and bipolar disorder. alpha1 immunoreactivity was observed in the neuropil of the prefrontal cortex and in the neuronal soma in specimens from both groups, as well as from normal controls. alpha1 immunolabeling in the neuronal soma from the schizophrenic group was more intense than that of the other two groups. The distribution of beta2/3 immunoreactivity was similar to that of alpha1. beta2/3 immunolabeling in the neuronal soma from the schizophrenia and bipolar disorder groups was more intense than that of the normal controls. The densitometry measurements, as well as the immunoblot analysis for alpha1 and beta2/3 were highly consistent with the alpha1 and beta2/3 immunohistochemistry results. The present study suggests that the expression of these two GABA(A) receptor subunits was altered in subjects with schizophrenia and bipolar disorder, but that the patterns of change differed between those with these two disorders. PMID- 15288500 TI - Perinatal inflammatory cytokine challenge results in distinct neurobehavioral alterations in rats: implication in psychiatric disorders of developmental origin. AB - Maternal stress, viral infection, and obstetric complications, which trigger cytokine signaling, are hypothesized to be involved in schizophrenia and its related disorders. The etiologic contribution of individual cytokines to such psychiatric disorders, however, remains to be evaluated. To estimate the impact of peripheral cytokine challenge on neurobehavioral development, we examined effects of four proinflammatory cytokines on rat neonates and their later behavioral performance. Sublethal doses of interleukin-1 alpha, interleukin-2, interleukin-6, or interferon-gamma were subcutaneously administered to rat pups for 9 days. These animals displayed alterations in physical development, including lower weight gain and/or accelerated eyelid opening. In addition, behavioral abnormalities related to fear/anxiety levels and sensorimotor gating emerged at different developmental stages, depending on the cytokine species administered. During juvenile stages, neonatal interleukin-2 treatment increased exploratory locomotor activity, whereas other cytokine treatments did not. At the post-puberty stage, however, the interleukin-2-induced abnormal motor activity became undetectable, whereas interleukin-1 alpha-treated rats developed abnormalities in startle response, prepulse inhibition (PPI), and social interaction. Subchronic treatment of an anti-psychotic drug, clozapine, ameliorated the impairment of prepulse inhibition without altering startle responses. These animal experiments illustrate that, during early postnatal development, inflammatory cytokine challenge in the periphery can induce future psycho-behavioral and/or cognitive impairments with various latencies, although the pathologic mechanisms underlying these abnormalities remain to be determined. PMID- 15288502 TI - Genetic and functional characteristics of the human in vivo LRP1/A2MR receptor suggested as a risk marker for Alzheimer's disease and other complex (degenerative) diseases. AB - LDL receptor-related protein/alpha2-macroglobulin receptor (LRP1/A2MR) a multiligand receptor is considered as not only being a possible risk factor of neurodegenerative diseases like Alzheimer's disease but also as determining the progression of other complex diseases like atherosclerosis and cancer. Although a large number of in vitro studies have highlighted its functional importance, as yet not enough is known about the clinical importance of the genetic background of LRP1 in human diseases. The aim of this ex vivo/in vivo study of 448 subjects was to present data on genetic LRP1 variants of healthy European Caucasians from Central Germany. Genotype-dependent LRP1 expression was analyzed in a representative subgroup (gene expression: n = 127, protein expression: n = 44). These data were evaluated in comparison to other published clinical LRP1 studies. For 15 functionally interesting genetic variants the genotype and allele distributions of the German Caucasians were presented in relation to their in vivo LRP1 gene and protein expression. A direct influence of the LRP1 promoter polymorphism c.1-25C>G on the human in vivo LRP1 expression level was demonstrated. In an analysis of 48 further studies genomic and functional results were evaluated. The analysis especially on Alzheimers's disease partly highlighted contradictory results, but suggested that ethnic as well as genomic characteristics determine LRP1 expression and must be considered in clinical investigations on human LRP1. PMID- 15288503 TI - Neurons regulating the duration of forward locomotion in Caenorhabditis elegans. AB - The locomotory behavior of Caenorhabditis elegans consists of four simple events, forward and backward movements, omega-shaped turns and rests. The wide variety of behaviors of this worm is achieved through a combination of these simple locomotions. To gain insight into the neuronal mechanisms regulating this locomotion, we analyzed the locomotory behavior of C. elegans over a long time period. By using an automatic worm tracking system, we revealed the existence of at least two distinct behavioral states -- pivoting and traveling -- in the forward locomotion of C. elegans in the absence of food. Pivoting is characterized by pronounced directional switching and resulting in short-duration forward movement, whereas in the traveling state forward movement is of longer duration. Pivoting occurred when we transferred a well-fed worm to an unseeded plate, and then the transition to traveling occurred, successively. We showed that, by laser ablation, antagonistic neuronal pathways consisting of nine classes of sensory neurons and four classes of interneurons were involved in this regulation. Loss of any one of these neurons altered the locomotory behavior. PMID- 15288504 TI - Sound sequence discrimination learning is dependent on cholinergic inputs to the rat auditory cortex. AB - In rat auditory cortex (AC) slices, synaptic potentiation following heterosynaptic stimulation is affected by the stimulus sequence used for induction. It was hypothesized that this sequence-dependent plasticity might be partly involved in the cellular mechanisms underlying sound sequence discrimination. Sequence dependence is abolished by muscarinic receptor antagonists. Therefore, dependence of sound sequence discrimination learning on cholinergic inputs to the rat AC was investigated. Rats were trained to discriminate the sequences of two sound components and a licking behavior in response to one of two possible sequences was rewarded with water. Atropine, a muscarinic receptor antagonist, attenuated sound sequence discrimination learning. The acquired sound sequence discrimination was not affected by atropine. Injections of the cholinergic immunotoxin 192IgG-saporin into the AC suppressed sound sequence discrimination learning, while discrimination between the two sound components was not affected. An inhibitor of M-current, linopirdine, restores the sequence dependence of synaptic potentiation in the AC slices suppressed by atropine. In this study, sound sequence discrimination learning attenuated by 192IgG-saporin was also restored by linopirdine. These similarities between sequence dependent plasticity in the AC slices and sound sequence discrimination learning support the hypothesis that the former is involved in the cellular mechanisms underlying the latter. PMID- 15288505 TI - Expression of connexin 26 in the ganglionic eminence of preterm infants after bleedings. AB - The ganglionic eminence being a prominent part of the telencephalic proliferative zone is the most common site of bleedings in preterm infants. This immunohistochemical study demonstrates that connexin 26 involved in forming intercellular gap junctions is expressed in ganglionic eminence cells up to 500 microm from the bleeding. In controls, no positive cells are present. It is discussed that an increase of gap junctional communication may result in spreading of the primary injury. PMID- 15288506 TI - Disruption of intraneuronal divalent cation regulation by methylmercury: are specific targets involved in altered neuronal development and cytotoxicity in methylmercury poisoning? AB - Methylmercury is an environmental contaminant which causes relatively specific degeneration of the granular layer of the cerebellum, despite its ability to bind thiol groups in proteins of all cell types. The mechanisms underlying the specific targeting of cells during MeHg poisoning may depend on specific receptors and other targets related to divalent cation homeostasis, particularly intracellular calcium (Ca(2+)(i) signaling. MeHg disrupts Ca(2+)(i) homeostasis in a number of neuronal models, including cerebellar granule cells in primary culture, and contributes to MeHg-induced cell death, impaired synaptic function and disruption of neuronal development. Interestingly, the disruption of [Ca(2+)](i) regulation occurs through specific pathways which affect Ca(2+) regulation by organelles, particularly mitochondria and the smooth endoplasmic reticulum (SER). Cholinergic pathways which affect [Ca(2+)](i) signaling also appear to be critical targets, particularly muscarinic acetylcholine (ACh) receptors which are linked to Ca(2+) release through inositol-1,4,5-triphosphate (IP(3)) receptors. [Ca(2+)](i) dysregulation may also underlie observed alterations in cerebellar neuron development through interaction with specific target(s) in the developing axon. In this review, we examine the hypothesis that MeHg affects specific targets to cause disruption of neuronal development and cell death. PMID- 15288507 TI - Mice lacking alpha-synuclein have an attenuated loss of striatal dopamine following prolonged chronic MPTP administration. AB - The functional role of alpha-synuclein in the pathogenesis of Parkinson's disease (PD) is not fully understood. Systemic exposure of alpha-synuclein-deficient mice to neurotoxins provides a direct approach to evaluate how alpha-synuclein may mediate cell death in a common murine model of PD. To this end, wild-type and homozygous alpha-synuclein knock-out mice were treated with sub-chronic and prolonged, chronic exposure to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). In the sub-chronic model, wild-type and alpha-synuclein knock-out mice were treated for five consecutive days with MPTP (1-25 mg/kg, s.c.) or vehicle, and sacrificed 3 days following the last injection. The prolonged, chronic model consisted of two injections of MPTP (1-20 mg/kg, s.c.) per week for 5 weeks, with co-administration of probenecid (250 mg/kg, i.p.), and animals were sacrificed 3 weeks following the last injection. Sub-chronic administration of MPTP caused a dramatic, dose-dependent decrease in striatal dopamine (DA) concentrations, while an attenuated response was observed in alpha-synuclein knock-out mice. Similarly, prolonged, chronic administration of MPTP produced a dose-dependent decrease in striatal DA concentrations, and a corresponding loss of striatal vesicular monoamine transporter (VMAT-2) protein in wild-type mice. However, mice lacking alpha-synuclein had an attenuated loss of striatal DA concentrations, while no loss of striatal VMAT-2 protein was observed. Both sub-chronic and prolonged, chronic administration of MPTP caused an increase in the 3,4 dihydroxyphenylacetic acid (DOPAC) to DA ratio in wild-type mice, but not in mice lacking alpha-synuclein. Despite attenuated toxicity, elevated lactate concentrations were observed in alpha-synuclein knock-out mice following prolonged, chronic MPTP administration. The results of this study provide evidence that alpha-synuclein null mice have an attenuated response to the toxic effects of MPTP exposure, even over prolonged periods of time and that the biochemical sequela of a protracted insult to nigrostriatal DA neurons are distinct between mice with and without alpha-synuclein expression. PMID- 15288508 TI - Cerebrospinal fluid S100B increases reversibly in neonates of methyl mercury intoxicated pregnant rats. AB - Methylmercury (MeHg), an organic methylated form of mercury, is one of the most hazardous environmental pollutants. MeHg is a potent neurotoxin, particularly during brain development. Neurotoxicity-induced by MeHg in prenatal age can cause mental disorders, cerebral palsy and seizures. We investigated cerebrospinal fluid (CSF) and brain tissue contents of S100B, a calcium binding protein produced and secreted by astrocytes, which has trophic and toxic activity on neurons depending on concentration. Pregnant rats were exposed to MeHg (5 mg/kg per day, on the 12th, 13th and 14th days of pregnancy). CSF and brain tissue (hippocampus, cerebral cortex and cerebellum) were obtained from neonate rats on 1, 15 and 30 days postnatal. MeHg accumulation was measured in brain tissue after birth and on the 30th postnatal day. An increase of CSF S100B was observed on the 15th, but not on the 30th postnatal day. Hippocampal tissue demonstrated increased S100B (and reduction in glial fibrillary acidic protein) immediately after birth, but not later. No changes in the S100B content were observed in cerebellum and cerebral cortex. No changes were observed in the spatial learning of these rats at adult age. These specific and reversible changes in the hippocampus could be related to the cognitive and epileptic disorders attributed to MeHg. Our results further indicate the glial involvement in the MeHg-induced neurotoxicity. The increment of CSF S100B in neonates exposed to MeHg reinforces the view that increased S100B is related to damage in the nervous system and that S100B could be a marker for MeHg-neurotoxicity. Although the cellular mechanism related to MeHg-induced increase in S100B content in CSF remains unknown, our results suggest the use of S100B as a peripheral marker of brain damage induced by MeHg. PMID- 15288509 TI - Overexpression of metallothionein protects cultured motor neurons against oxidative stress, but not mutant Cu/Zn-superoxide dismutase toxicity. AB - Mutations in Cu/Zn-superoxide dismutase 1 (SOD1) are responsible for a familial form of amyotrophic lateral sclerosis (FALS). It has been proposed that oxidative stress and abnormal metal homeostasis contribute to death of motor neurons in this disease. Also, inability of motor neurons to upregulate protective proteins under stress may contribute to their preferential vulnerability to toxicity. Metallothioneins (MT) are low molecular weight, metal-binding proteins with established antioxidant capabilities. This study investigated the ability of motor neurons to upregulate MT isoforms in response to expression of mutant SOD1(G93A) or exposure to other neurotoxicants, and the ability of MT-I gene transfer to protect motor neurons from these stresses. MT isoform-I and -II were expressed constitutively in astrocytes and other non-neuronal cells of dissociated spinal cord cultures, but not in motor neurons. MT-I/II was upregulated in astrocytes, but not motor neurons, following treatment with ZnCl(2) or excitotoxic concentrations of glutamate. MT-III expression was restricted to neurons and was unaffected by treatment with ZnCl(2), paraquat, or glutamate. Overexpression of MT-I in motor neurons by gene transfer reduced the toxicity of ZnCl(2) and paraquat, but failed to protect them against glutamate or SOD1(G93A). These data are evidence against metal-catalyzed, oxidative stress being the primary mechanisms of toxicity conferred by disease-causing mutations in SOD1. PMID- 15288510 TI - Phosphatidylinositol-3 kinase/Akt and GSK-3 mediated cytoprotective effect of epigallocatechin gallate on oxidative stress-injured neuronal-differentiated N18D3 cells. AB - Epigallocatechin gallate (EGCG) is one of most famous compounds of green tea. EGCG suppresses apoptosis induced by oxidative radical stress through several mechanisms. This study was designed to investigate whether EGCG plays a cytoprotective role by activating phosphatidylinositol-3 kinase (PI3K)/Akt dependent anti-apoptotic pathway and inhibiting glycogen synthase kinase-3 (GSK 3) activity in oxidative stressed N18D3 neural cells. N18D3 cells, mouse neuroblastoma X dorsal root ganglion hybrid cell line, were pre-treated with EGCG or z-VAD-fmk, non-selective caspase inhibitor used as a control substance, for 2 h. The N18D3 cells were then exposed to low concentration of H(2)O(2) (100 microM) for 30 min, and further incubated for 24 h. MTT (3,[4,5-dimethylthiazol] 2-yl) assay and trypan blue staining were used to identify cell viability. Immunoreactivity (IR) of PI3K, Akt, and GSK-3 beta were measured by Western blotting. MTT assay and trypan blue staining showed that EGCG and z-VAD-fmk significantly increased cell viability, and IR of PI3K, phospho-Akt and phospho GSK-3 beta was significantly increased in the cells treated with EGCG, but not in z-VAD-fmk treated. These results imply that EGCG has neuroprotective effect by increasing PI3K/Akt-dependent anti-apoptotic signals. PMID- 15288511 TI - Impairments of colour vision induced by organic solvents: a meta-analysis study. AB - The impairment of colour discrimination induced by occupational exposure to toluene, styrene and mixtures of organic solvents is reviewed and analysed using a meta-analytical approach. Thirty-nine studies were surveyed covering a wide range of exposure conditions. Those studies using the Lanthony Panel D-15 desaturated test (D-15d) were further considered. From these for 15 samples data on colour discrimination ability (Colour Confusion Index, CCI) and exposure levels were provided, required for the meta-analysis. In accordance with previously reported higher CCI values for the exposed groups, the computations yielded positive effect sizes for 13 of the 15 samples, indicating that in the great majority of the studies the exposed groups showed inferior colour discrimination. However, the meta-analysis showed great variation in effect sizes across the studies. Possible reasons for inconsistency among the reported findings are discussed. These pertain to exposure-related parameters, as well as to confounders such as conditions of test administration and characteristics of subject samples. Those factors vary considerably among the studies and might have greatly contributed to divergence in measured colour vision capacity, thereby obscuring consistent effects of organic solvents on colour discrimination. PMID- 15288512 TI - S-methylcysteine may be a causal factor in monohalomethane neurotoxicity. AB - S-methylcysteine (SMC) is formed after exposure to monohalomethanes in rodents as well as in humans. The present study was performed to study whether SMC, directly or indirectly, contributes to the well-known neurotoxicity of monohalomethanes. We have investigated the effects of acute exposure to SMC by means of electrophysiolocal measurements in freshly prepared hippocampal slices and dissociated hippocampal neurons in culture. For longer-term exposures (24 h) we have used organotypic cultures (2 weeks in culture), taking electrophysiologic recordings and assessing membrane integrity with propidium iodide (PI) fluorescence. We found that only high concentrations of SMC (10(-2) M; exposure time 30 min) in freshly isolated slices of adult rats reduce synaptically evoked population spikes in the CA1 region. This effect was at least partially reversible. In organotypic cultures, at 5 x 10(-5) M after 24 h of exposure, SMC compromises membrane integrity as revealed by PI fluorescence, only in the dentate gyrus, spreading to pyramidal cell layers at 50 x 10(-4) M. At 5 x 10(-6) and 2 x 10(-5) M, under the same experimental conditions, no changes were seen with the PI method, but we recorded increased population spike amplitudes, repetitive discharges and frequency potentiation (at a stimulus repetition rate of 0.05 Hz). Using whole-cell patch clamp in hippocampal dissociated neurons we have found that SMC (applied for approximately 1s) reduces GABA-induced currents ( IC(50) = 4.4 x 10(-4) M) without having an effect of its own, acting like a competitive antagonist at GABA(A) receptors. Our findings are in line with the view that the ability of monohalomethanes to induce the formation of SMC is an important factor for their neurotoxicity, provided that SMC is allowed to act at least for several hours. The effects exerted by SMC seem to be due, at least in part, to its interaction with GABA receptors. PMID- 15288513 TI - The modulatory effect of pyrethroids on acetylcholine release in the hippocampus of freely moving rats. AB - The peripheral effects of pyrethroids on Na(+) channels are well known but the effects on CNS neurotransmission are less known. In the present study, type I and II pyrethroids were found to affect the release of acetylcholine (ACh) from hippocampus in freely moving rats as measured by in vivo microdialysis. The basal release of ACh from the hippocampus of untreated rats was 6.6 pmol/10 microl/10 min. Allethrin had an interesting dual effect on ACh release, increasing ACh efflux (to about 300% of baseline) at the lower dose of 20 mg/kg i.p. with a peak time of 60 min and decreasing the efflux (to about 40% of baseline) at the higher dose of 60 mg/kg i.p. up to 3 h after administration. Cyhalothrin 20 and 60 mg/kg i.p. inhibited the release (to about 30% of baseline) dose-dependently, with a peak time of 50-60 min after administration. Deltamethrin 20 mg/kg i.p. increased the efflux (to about 250% of baseline) with a peak time of 30 min after administration and 60 mg/kg i.p. increased the efflux (to about 450% of baseline) and remained at a steady level during the rest of the 3 h experiment. Control vehicle injections had no effect on the efflux of ACh in any of the experiments. This is the first report, using in vivo microdialysis, that pyrethroids modulate the ACh release in the hippocampus of rat brain. PMID- 15288514 TI - Differential neuroprotective effects of the NMDA receptor-associated glycine site partial agonists 1-aminocyclopropanecarboxylic acid (ACPC) and D-cycloserine in lithium-pilocarpine status epilepticus. AB - The status epilepticus (SE) induced in rats by lithium-pilocarpine (Li-pilo) shares many common features with soman-induced SE including a glutamatergic phase that is inhibited by NMDA antagonists. The present study determined whether 1 aminocyclopropanecarboxylic acid (ACPC) or D-cycloserine (DCS), both partial agonists of the strychnine-insensitive glycine site on the NMDA receptor ionophore complex, exerted anticonvulsant or neuroprotectant activity in Li-pilo SE. ACPC or DCS were administered either immediately following pilocarpine (exposure treatment) or 5 min after the onset of SE as determined by ECoG activity. SE was allowed to proceed for 3 h before termination with propofol. The rats were sacrificed 24 h following pilocarpine administration. Neither drug had an effect on the latency to seizure onset or the duration of seizure activity. ACPC administered 5 min after SE onset produced significant neuroprotection in cortical regions, amygdala and CA1 of the hippocampus. In contrast, when administered as exposure treatment ACPC enhanced the neural damage in the thalamus and CA3 of the hippocampus suggesting the neuropathology in those regions is mediated by a different subset of NMDA receptors. DCS had no neuroprotectant activity in Li-pilo SE but exacerbated neuronal damage in the thalamus. Neither drug affected the cholinergic convulsions but both had differential effects on neural damage. This suggests that the SE-induced seizure activity and subsequent neuronal damage involve independent mechanisms. PMID- 15288515 TI - Effects of methylmercury on the microvasculature of the developing brain. AB - The study, undertaken with the aim of further investigating the effects of methylmercury (MeHg) exposure on the developing brain, was performed in the cerebellum of chick embryos, chronically treated with a MeHgCl solution dropped onto the chorioallantoic membrane, and in control embryo cerebella. Quantitative evaluations, performed by cold vapour atomic absorption spectrophotometry, demonstrated a high mercury content in the chorioallantoic membrane, encephalon, liver and kidney of the treated embryos. The morphological observations showed severe neuronal damage consisting of degenerative changes of the granules and Purkinje neurons. The effects on astrocytes were even more severe, since they were extremely rare both in the neuropil and around the vessel wall. Compared with the controls, the cerebellar vessels of MeHg-treated embryos showed immature morphology, poor differentiation of endothelial barrier devices, and high permeability to the exogenous protein horseradish peroxidase. These findings support the hypothesis that MeHg-related neuronal sufferance may be secondary to astrocytic damage and suggest that the developmental neurotoxicity of this compound could also be related to astrocyte loss-dependent impairment of blood brain barrier (BBB) differentiation. PMID- 15288516 TI - Aluminum maltolate-induced toxicity in NT2 cells occurs through apoptosis and includes cytochrome c release. AB - Aluminum (Al) compounds are neurotoxic and have been shown to induce experimental neurodegeneration although the mechanism of this effect is unclear. In order to study this neurotoxic effect of Al, we have developed an in vitro model system using Al maltolate and human NT2 cells. Al maltolate at 500 microM caused significant cell death with a 24-h incubation and this toxicity was even more evident after 48 h. Lower doses of Al maltolate were also effective, but required a longer incubation for cell death. Nuclear fragmentation suggestive of apoptosis was observed as early as three hours and increased substantially through 24 h. Chromatin condensation and nuclear fragmentation were confirmed by electron microscopy. In addition, TUNEL positive nuclei were also observed. The release of cytochrome c was demonstrated with Western blot analysis. This in vitro model using human cells adds to our understanding of Al neurotoxicity and could provide insight into the neurodegenerative processes in human disease. PMID- 15288517 TI - Exposure to cadmium during gestation and lactation decreases cocaine self administration in rats. AB - This investigation examined the effects of perinatal cadmium exposure on subsequent self-administration of cocaine during the adult cycle. Female Sprague Dawley rats were gavaged daily with 0.0 (14% sucrose solution, w/v) or 5.0 mg cadmium chloride (dissolved in 14% sucrose solution, w/v) for 30 days prior to breeding with non-exposed males. Dams continued to experience cadmium exposure through gestation and until pups were weaned at postnatal day (PND) 21. On PND 70, offspring were anesthetized and chronic indwelling jugular catheters were implanted. Following recovery, test subjects were trained in operant chambers to self-administer 0.500 mg/kg infusion (inf) intravenous cocaine on a fixed-ratio (FR) 2 schedule of reinforcement. Following acquisition, self-administration rates were tested for saline, 0.030, 0.060, 0.125, 0.250, and 0.500 mg/kg inf cocaine. Rats exposed developmentally to cadmium self-administered significantly less than controls at saline, 0.030, and 0.060 mg/kg inf cocaine. These data indicate that early-life cadmium exposure, a common exposure vector of which is the use of tobacco products, may affect cocaine sensitivity. PMID- 15288518 TI - Effects of norepinephrine depletion in rats during cerebral post-ischemic reperfusion. AB - The present paper reports the effects of norepinephrine depletion in rats, after treatment with N-(2-chloroethyl)-N-ethyl 2-bromobenzylamine (DSP-4) neurotoxin, on partial cerebral ischemia and reperfusion. Histological observations made under experimental conditions of noradrenergic (NA)-depletion demonstrated that neuronal lesions were not exacerbated; in fact, in DSP-4-treated ischemic animals, a minor number of neurons appeared damaged. Our results suggest that neuronal recovery after post-ischemic reperfusion is not affected by NA depletion. DSP-4 neurotoxin does not induce 5-hydroxy-triptamine (5-HT) depletion. PMID- 15288519 TI - The soy isoflavone, genistein, protects human cortical neuronal cells from oxidative stress. AB - Genistein, a soy isoflavone, has been shown to mimic the pharmacological actions of the endogenous steroid estrogen with which it has structural similarities. There is now evidence that the genistein can prevent disorders-like heart diseases, cancer and diabetes as well. However, very few studies have looked at the effect of genistein on the central nervous system. Published studies also show conflicting conclusions regarding the effects of genistein in the brain. The current study was conducted in the human cortical cell lines HCN1-A and HCN2 in order to determine the neuroprotective efficacy of genistein. It was observed that pre-treatment with 50 or 10 microM genistein was able to protect HCN1-A and HCN2 cells from the cell death induced by 100 microM or 1 mM tertiary butylhydroperoxide (t-BuOOH; a free radical generating toxin). The morphological disruption caused by t-BuOOH was also prevented by genistein in HCN2 cells. Moreover, genistein was able to prevent the down-regulation of the anti-apoptotic protein bcl-2 that was caused by t-BuOOH treatment. These results indicate that genistein may have neuroprotective effect in cortical cells, which may be mediated by its regulation of the anti-apoptotic protein bcl-2. PMID- 15288521 TI - The Pleistocene colonization of northeastern Europe: a report on recent research. AB - Recent studies have shown that northeastern Europe was occupied by humans significantly earlier than previously thought. Some traces of human presence in the European Arctic even date back to about 35-40 ka. This paper discusses the Middle and early Upper Palaeolithic (EUP) assemblages from this area within the local context of their environmental characteristics, as well as their implications for our views on the occupational history of northern environments. PMID- 15288522 TI - Early Natufian remains: evidence for physical conflict from Mt. Carmel, Israel. AB - Prior to the establishment of farming communities direct physical evidence for human conflict was rarely reported from archaeological contexts. Here we present a case of an Early Natufian (14,500-13,000 cal B.P.) projectile, classified as Helwan lunate, embedded inside the seventh or eighth thoracic vertebra sequence of a mature middle age adult male. Due to calcareous concretion four vertebras were still in anatomical connection when uncovered by F. Turville-Petre, during his excavations at Kebara cave (Mt. Carmel) in 1931. PMID- 15288523 TI - A chronology for late prehistoric Madagascar. AB - A database has been assembled with 278 age determinations for Madagascar. Materials 14C dated include pretreated sediments and plant macrofossils from cores and excavations throughout the island, and bones, teeth, or eggshells of most of the extinct megafaunal taxa, including the giant lemurs, hippopotami, and ratites. Additional measurements come from uranium-series dates on speleothems and thermoluminescence dating of pottery. Changes documented include late Pleistocene climatic events and, in the late Holocene, the apparently human caused transformation of the environment. Multiple lines of evidence point to the earliest human presence at ca. 2300 14C yr BP (350 cal yr BC). A decline in megafauna, inferred from a drastic decrease in spores of the coprophilous fungus Sporormiella spp. in sediments at 1720+/-40 14C yr BP (230-410 cal yr AD), is followed by large increases in charcoal particles in sediment cores, beginning in the SW part of the island, and spreading to other coasts and the interior over the next millennium. The record of human occupation is initially sparse, but shows large human populations throughout the island by the beginning of the Second Millennium AD. Dating of the "subfossil" megafauna, including pygmy hippos, elephant birds, giant tortoises, and large lemurs, demonstrates that most if not all the extinct taxa were still present on the island when humans arrived. Many taxa overlapped chronologically with humans for a millennium or more. The extinct lemurs Hadropithecus stenognathus, Pachylemur insignis, Mesopropithecus pithecoides, and Daubentonia robusta, and the elephant birds Aepyornis spp. and Mullerornis spp., were still present near the end of the First Millennium AD. Palaeopropithecus ingens, Megaladapis edwardsi, and Archaeolemur sp. (cf. edwardsi) may have survived until the middle of the Second Millennium A.D. One specimen of Hippopotamus of unknown provenance dates to the period of European colonization. PMID- 15288524 TI - Prevalence and the duration of linear enamel hypoplasia: a comparative study of Neandertals and Inuit foragers. AB - As a dental indicator of generalized physiological stress, enamel hypoplasia has been the subject of several Neandertal studies. While previous studies generally have found high frequencies of enamel hypoplasia in Neandertals, the significance of this finding varies with frequencies of enamel hypoplasia in comparative samples. The present investigation was undertaken to ascertain if the enamel hypoplasia evidence in Neandertals suggests a high level of physiological stress relative to a modern human foraging group, represented here by an archaeological sample of Inuit from Point Hope, Alaska. Unlike previous studies, this study focused specifically on linear enamel hypoplasia (LEH), emphasizing systemic over localized causes of this defect by considering LEH to be present in an individual only if LEH defects occur on two anterior teeth with overlapping crown formation periods. Moreover, this study is the first to evaluate the average growth disruption duration represented by these defects in Neandertals and a comparative foraging group. In the prevalence analysis, 7/18 Neandertal individuals (from Krapina and southern France) and 21/56 Neandertal anterior teeth were affected by LEH, or 38.9% and 37.5% respectively. These values do not differ significantly from those of the Inuit sample in which 8/21, or 38.1% of individuals, and 32/111, or 28.8% of anterior teeth were affected. For the growth disruption duration analysis, 22 defects representing separate episodes of growth disruption in Neandertals were compared with 22 defects in the Inuit group using three indicators of duration: the number of perikymata (growth increments) in the occlusal walls of LEH defects, the total number of perikymata within them, and defect width. Only one indicator, the total number of perikymata within defects, differed significantly between the Inuit and Neandertal groups (an average of 13.4 vs. 7.3 perikymata), suggesting that if there is any difference between them, the Inuit defects may actually represent longer growth disruptions than the Neandertal defects. Thus, while stress indicators other than linear enamel hypoplasia may eventually show that Neandertal populations were more stressed than those of modern foragers, the evidence from linear enamel hypoplasia does not lend support to this idea. PMID- 15288525 TI - Locomotor mechanics of the slender loris (Loris tardigradus). AB - The quadrupedal walking gaits of most primates can be distinguished from those of most other mammals by the presence of diagonal-sequence (DS) footfall patterns and higher peak vertical forces on the hindlimbs compared to the forelimbs. The walking gait of the woolly opossum (Caluromys philander), a highly arboreal marsupial, is also characterized by diagonal-sequence footfalls and relatively low peak forelimb forces. Among primates, three species--Callithrix, Nycticebus, and Loris--have been reported to frequently use lateral-sequence (LS) gaits and experience relatively higher peak vertical forces on the forelimbs. These patterns among primates and other mammals suggest a strong association between footfall patterns and force distribution on the limbs. However, current data for lorises are limited and the frequency of DS vs. LS walking gaits in Loris is still ambiguous. To test the hypothesis that patterns of footfalls and force distribution on the limbs are functionally linked, kinematic and kinetic data were collected simultaneously for three adult slender lorises (Loris tardigradus) walking on a 1.25 cm horizontal pole. All subjects in this study consistently used diagonal-sequence walking gaits and always had higher peak vertical forces on their forelimbs relative to their hindlimbs. These results call into question the hypothesis that a functional link exists between the presence of diagonal sequence walking gaits and relatively higher peak vertical forces on the hindlimbs. In addition, this study tested models that explain patterns of force distribution based on limb protraction angle or limb compliance. None of the Loris subjects examined showed kinematic patterns that would support current models proposing that weight distribution can be adjusted by actively shifting weight posteriorly or by changing limb stiffness. These data reveal the complexity of adaptations to arboreal locomotion in primates and indicate that diagonal-sequence walking gaits and relatively low forelimb forces could have evolved independently. PMID- 15288526 TI - The effect of lower limb length on the energetic cost of locomotion: implications for fossil hominins. AB - The consequences of the relatively short lower limbs characteristic of AL 288-1 have been widely discussed, as have the causes and consequences of the short limbs of Neanderthals. Previous studies of the effect of limb length on the energetic cost of locomotion have reported no relationship; however, limb length could have accounted for as much as 19% of the variation in cost and gone undetected (Steudel and Beattie, 1995; Steudel, 1994, 1996). Kramer (1999) and Kramer and Eck (2000) have recently used a theoretical model to predict the effect of the shorter limbs of early hominids, concluding that the shorter limbs may actually have been energetically advantageous. Here, we took an experimental approach. Twenty-one human subjects, of varying limb lengths, walked on a treadmill at 2.6, 2.8, 3.0 and 3.2m.p.h., while their expired gases were analyzed. The subjects walked for 12 minutes at each speed and their rates of oxygen consumption (VO2) over four minutes were averaged to estimate VO2. We also measured each subject's height, weight and lower limb length. Lean body mass and % fat were determined using dual-energy x-ray absorptiometry. ANCOVA with total VO2 at either speed as the dependent variable and total lean mass, % fat and lower limb length as covariates resulted in all three covariates having a significant positive effect on VO2 at p<0.01. Subjects with relatively longer lower limbs had lower locomotor costs. Thus the short lower limbs characteristic of some hominid taxa would have resulted in more costly locomotion, barring some physiological anomaly. The magnitude of this effect is substantial; Neanderthals are estimated to have had locomotor costs 30% greater than those of contemporary anatomically modern humans. By contrast the increase in lower limb length seen in H. erectus would have mitigated the increase in locomotor costs produced by the increase in body size. PMID- 15288528 TI - An introduction to the short-term toxicology of respirable industrial fibres. AB - Fibrous materials, exemplified by asbestos, that release respirable fibres are in common use and there is considerable knowledge regarding the toxicology of these common fibres. Newer materials or those that are under development, such as synthetic organic fibres and carbon nanotubes may have a different toxicology paradigms. The existing paradigm for silicate fibres suggests that respirable fibre types vary in their ability to cause lung disease and that this can be understood on the basis of the length of the fibres and their biopersistence in the lungs. Because fibres are regulated on a fibre number basis and the hazard is understood on the basis of the number of long fibres, in fibre testing the dose should always be expressed as fibre number, not mass and the length and diameter distribution need to be known. Short-term biological tests are likely to produce false positives in the case of long non-biopersistent fibres, because whilst they may have effects in vitro, they do not persist long enough in the lungs for sufficient dose to build up and produce effects in vivo. The biopersistence of fibres is therefore a key factor that needs to be known in order to interpret short-term tests that may claim to predict fibre pathogenicity. PMID- 15288529 TI - Research needs to improve risk assessment of fiber toxicity. AB - Risk characterization of exposure to toxic compounds requires information on the intrinsic toxic properties, including toxic mechanism and toxicokinetics, on dose response at the most critical targets for identification of the NOEL or for extrapolation from high to low dose, and on human exposure. Abundant information is available on the intrinsic properties of MMMF, on the three D's (dose, dimension, durability) and on the toxic mechanisms. However, only a few of these studies provide information on the dose response of the effects or of the mechanisms investigated. Moreover, in many cases single high doses exceeding the MTD have been applied and are difficult to interpret for lower exposure scenarios. Risk characterization is further hampered by the still open question whether MMMF are directly genotoxic or induce secondary genotoxicity via inflammation. Finally, there is disagreement about the relevance of animal studies on MMMF for humans and thus about the most rational extrapolation of the dose response of toxic effects observed in animals to man. These deficits are briefly described and discussed from a toxicological point of view. PMID- 15288530 TI - Signal transduction pathways relevant for neoplastic effects of fibrous and non fibrous particles. AB - Apart from their genotoxic effects, both fibrous and non-fibrous particles are known to induce signalling pathways involved in the development of malignant lung diseases. Different direct effects of particles as well as indirect cellular effects are believed to induce changes in apoptosis or proliferation in target cells. Signalling events, e.g. the induction of mitogen-activated protein kinase (MAPK) cascades resulting in the activation of the transcription factor AP-1, as well as the induction of the transcription factor NFkappaB which mainly mediates the expression of pro-inflammatory genes are discussed. There is some insight into the molecular mechanisms eliciting these pathways. Therefore, this review aims to give an overview on signalling pathways as well as initial events including effects of reactive oxygen and nitrogen species, membrane receptors and particle uptake. PMID- 15288531 TI - Approaches for the development of occupational exposure limits for man-made mineral fibres (MMMFs). AB - Occupational exposure limits (OELs) are an essential tool in the control of exposure to hazardous chemical agents, and serve to minimise the occurrence of occupational diseases associated with such exposure. The setting of OELs, together with other associated measures, forms an essential part of the European Community's strategy on health and safety at work, upon which the legislative framework for the protection of workers from risks related to chemical agents is based. The European Commission is assisted by the Scientific Committee on Occupational Exposure Limits (SCOEL) in its work of setting OELs for hazardous chemical agents. The procedure for setting OELs requires information on the toxic mechanisms of an agent that should allow to differentiate between thresholded and non-thresholded mechanisms. In the first case, a no-observed adverse effect level (NOAEL) can be defined, which can be the basis for a derivation of an OEL. In the latter case, any exposure is correlated with a certain risk. If adequate scientific data are available, SCOEL estimates the risk associated with a series of exposure levels. This can then be used for guidance, when setting OELs at European level. Man-made mineral fibres (MMMFs) are widely used at different worksites. MMMF products can release airborne respirable fibres during their production, use and removal. According to the classification of the EU system, all MMMF fibres are considered to be irritants and are classified for carcinogenicity. EU legislation foresees the use of limit values as one of the provisions for the protection of workers from the risks related to exposure to carcinogens. In the following paper, the research requirements identified by SCOEL for the development of OELs for MMMFs will be presented. PMID- 15288532 TI - Man-made mineral (vitreous) fibres: evaluations of cancer hazards by the IARC Monographs Programme. AB - Man-made vitreous (glass-like) fibres are non-crystalline, fibrous inorganic substances (silicates) made primarily from rock, slag, glass or other processed minerals. These materials, also called man-made mineral fibres, include glass fibres (used in glass wool and continuous glass filament), rock or stone wool, slag wool and refractory ceramic fibres. They are widely used for thermal and acoustical insulation and to a lesser extent for other purposes. These products are potentially hazardous to human health because they release airborne respirable fibres during their production, use and removal. Man-made mineral fibres and man-made vitreous fibres have been the subject of reviews by IARC Monographs Working Groups in 1987 and 2001, respectively, which resulted in evaluations of the carcinogenic hazard to humans from exposure to these materials. These reviews and evaluations have been published as Volumes 43 and 81 of the IARC Monographs series [IARC Monographs on the Evaluation of Carcinogenic Risks to Humans, vol. 43, Man-made Mineral Fibres and Radon (1988); IARC Monographs on the Evaluation of Carcinogenic Risks to Humans, vol. 81, Man-made Vitreous Fibres (2002)]. The re-evaluation in 2001 was undertaken because there have been substantial improvements in the quality of the epidemiological information available on the carcinogenicity to humans of glass fibres, continuous glass filament and rock/slag wool. The new evaluations have addressed the limitations of earlier cohort studies, particularly concerning the lack of adjustment with respect to concomitant risk factors such as smoking and other sources of occupational exposure. In addition, the evaluation of the evidence for carcinogenicity of glass fibres to experimental animals has been refined, by making a distinction between insulation glass wool and special-purpose glass fibres. The results of the evaluations in 1987 and 2001 are thus different in several aspects. In this paper, the reviews and evaluations of the carcinogenic hazards of exposure to man-made mineral fibres (MMMF, Monograph volume 43, [1]) and man-made vitreous fibres (MMVF, Monograph volume 81, [2]) are summarised, and the differences explained. In particular, the considerations of the respective IARC Monographs Working Groups (1987, 2001) in reaching their conclusions are discussed in some detail. PMID- 15288533 TI - Induction of 8-hydroxydeoxyguanosine by man made vitreous fibres and crocidolite asbestos administered intraperitoneally in rats. AB - Inhaled fibres with certain physico-chemical properties are known to induce mesothelioma in humans. The induction of reactive oxygen (ROS) or nitrogen species (RNS) have been suggested as molecular mechanism of fibre induced carcinogenesis. In earlier studies we were able to demonstrate that crocidolite asbestos in vivo induces mutations in transgenic rats with a specific molecular spectrum that indicates the involvement of 8-hydroxydeoxyguanosine (8-OHdG) as pre-mutagenic adduct. 8-OHdG may be induced by primary (direct) and/or secondary (cellular mediated) mechanisms. Therefore, the induction of 8-OHdG as well as the inflammatory response of animals treated with fibre samples significantly differing in their physico-chemical characteristics was investigated. As appropriate system to study mesothelioma carcinogenesis, intraperitoneal injection in rats was used with samples of UICC crocidolite, crocidolite with reduced iron content, and a vitreous fibre (MMVF 11). Equal numbers of carcinogenic fibres from each sample revealed significant comparable increases in 8-OHdG induction. Parameters of inflammation (percentage of macrophages and TNF alpha secretion) correlated significantly with the induction of 8-OHdG, 10 weeks after treatment. PMID- 15288534 TI - Mutagenesis by asbestos in the lung of lambda-lacI transgenic rats. AB - In order to get more insight into the mechanism of asbestos-related lung cancer, the mutagenic potential of asbestos was examined in vivo in rat lung. Groups of five transgenic lambda-lacI (Big Blue) rats were intratracheally instilled with single doses of 1 or 2mg, or with four weekly doses of 2mg, per animal of the amosite asbestos. Sixteen weeks after instillation, the mutation frequency was found to be increased in lung DNA by 2-fold at doses of 2 mg (P = 0.035) and of 4 x 2 mg (P = 0.007) amosite. No significant changes were observed after 4 weeks of exposure. In separate experiments, wild-type F344 rats were treated by the same regimen as described above and markers of inflammation, genotoxicity, cell proliferation and lung tissue damage were analysed. Our results indicate a weak but persistent inflammation and cell proliferation which possibly plays a major role in the observed mutagenic effect. PMID- 15288535 TI - Benzo[a]pyrene-enhanced mutagenesis by asbestos in the lung of lambda-lacI transgenic rats. AB - To study the suspected mechanism of the interaction between tobacco smoking and asbestos exposure in the modulation of cancer risk, the mutagenic potential of asbestos in combination with the tobacco smoke carcinogen benzo[a]pyrene (B[a]P) was examined in vivo in the rat lung. B[a]P was administered intratracheally in one set of experiments, or by two daily intraperitoneal injections in another set of experiments, to lambdalacI transgenic rats, together with 1, 2 or 4 x 2 mg amosite in one experiment. In the first experiment, the combined action of amosite and B[a]P caused a synergistic (superadditive) increase of mutation frequency in the lung, as compared to groups treated only with asbestos or B[a]P. In the second experiment, i.p. treatment with B[a]P did not significantly alter the mutation frequency induced by amosite, neither after 4 nor after 16 weeks of exposure. The B[a]P-DNA adduct levels were unaffected by amosite co-treatment in both experiments. We assume that the synergistic increase of mutation frequency after intratracheal treatment was due to the mitogenic activities of B[a]P and of amosite. In conclusion, our findings indicate that a weak and delayed mutagenic effect of amosite in rat lung observed in another study was strongly enhanced by the concomitant action of B[a]P. The striking enhancement effect of B[a]P may provide a basis for understanding the suspected synergism of smoking on asbestos carcinogenesis. PMID- 15288536 TI - Genotoxic effects of asbestos in humans. AB - Risks of carcinogenic and non-carcinogenic effects from asbestos continue owing to the persistence of the fibres in building materials and other products. For this reason, epidemiological and mechanistic research on the toxic effects of asbestos and mineral fibres is still needed. The present molecular epidemiological study was conducted in a former asbestos cement plant in Slovakia. Altogether 82 subjects were investigated, 61 exposed subjects (24 smokers and 37 non-smokers), and 21 factory controls (8 smokers and 13 non smokers). Workers were exposed to asbestos for between 5 and 40 years. Though the exposure to asbestos during past 40 years was relatively high, at the time of sampling the concentrations of asbestos in the production hall exceeded the Slovak occupational limit (0.001 fibre/cm3) by a factor of only 3-5. The office area levels were below this limit. Biomarkers of exposure, effect and individual susceptibility were measured, including DNA damage (strand breaks [SBs], base oxidation and alkylation, using the comet assay); cytogenetic parameters; and individual DNA repair capacity (incision at 8-oxoguanine measured using a modified comet assay). Oxidised pyrimidines were significantly higher in exposed men compared with non-exposed (P = 0.04). There was also a positive association between SBs (P = 0.04) and age, and alkylation damage to DNA (P = 0.04) and age. Moreover, oxidised pyrimidines (P = 0.01) and alkylated bases (P = 0.001) strongly correlated with years of occupational exposure. Micronucleus frequency did not differ between exposed and control subjects. Repair capacity overall did not show any effect of exposure, though female controls had higher incision rates than did female exposed subjects. However, exposed asbestos workers had significantly higher numbers of chromosomal aberrations (P = 0.01) compared with control group. This finding is consistent with the known association of chromosome aberrations with cancer-risk. PMID- 15288537 TI - Does occupational exposure to mineral fibres cause DNA or chromosome damage? AB - Markers of genetic stability were monitored in lymphocytes from 98 workers employed in rockwool manufacture in a factory in the Slovak Republic, and 43 controls (administrative employees in the same factory). Strand breaks in lymphocyte DNA were higher in exposed compared to control non-smokers, but there was no effect of exposure on specific damage to bases in DNA, nor on chromosome aberrations. The frequency of micronuclei was higher in women in the control group than in rockwool-exposed women. DNA repair (8-oxoguanine DNA glycosylase activity) was unaffected by exposure, but was negatively correlated with micronucleus frequency, implying that unrepaired 8-oxoguanine contributes to micronucleus formation. The conclusion from this study is that, overall, rockwool exposure has no deleterious effect on genetic stability in humans. PMID- 15288538 TI - Immunomodulatory effects of mineral fibres in occupationally exposed workers. AB - In the context of a large-scale molecular epidemiology study, the possible immunomodulatory effects of mineral fibres, in workers occupationally exposed to asbestos, rockwool and glass fibres, were examined. In each plant, 61, 98 and 80 exposed workers and 21, 43 or 36 control clerical subjects, respectively, were recruited. In the case of the asbestos-exposed subjects, an additional town control group of 49 people was included. Evidence of pulmonary fibrosis was found in 42% of the asbestos-exposed workers, while evidence of pleural fibrosis was found in 24%. The asbestos-exposed cohort had significantly decreased forced vital capacity of lungs as well as forced expiratory volume per first second. Our findings indicate that exposure to all three types of fibres examined modulates to different degrees the immune response. Suppression of T-cell immunity and to a lesser extent, B-cell immunity was found in the case of workers from a former asbestos cement plant, while stimulation of T-cell response was observed in rockwool workers, and stimulation of T- and B-cell response was seen in glass fibre workers. Depression of the percentage of lymphocyte subpopulation of CD 16+56 (natural killer cells) in peripheral blood was found in glass fibre workers. Statistical analysis showed increased levels of proinflammatory cytokines (IL-6 asbestos; IL-8 all three fibres), expression of adhesion molecule L-selectin on granulocytes and monocytes (asbestos), levels of soluble adhesion molecules (SAMs) in sera (ICAM-1 all three fibres; E-selectin glass fibres), increased levels of immunoglobulin E (asbestos and rockwool) and elevated expression of activation markers on eosinophils (CD66b asbestos, glass fibres; CD69 asbestos). Significant correlations were observed between lymphocyte proliferation and markers of DNA damage and repair. Increased levels of proinflammatory cytokines, SAMs, immunoglobulin E and elevated expression of activation markers on eosinophils was found in people with symptoms of hypersensitivity and an elevated inflammatory status. PMID- 15288539 TI - Gene expression fingerprints of largemouth bass (Micropterus salmoides) exposed to pulp and paper mill effluents. AB - Effluents from pulp and paper mills that historically have used elemental chlorine in the bleaching process have been implicated in inhibiting reproduction in fish. Compounds with estrogenic and androgenic binding affinities have been found in these effluents, suggesting that the impairment of reproduction is through an endocrine-related mode of action. To date, a great deal of attention has been paid to phytoestrogens and resin acids that are present in mill process streams as a result of pulping trees. Estrogen and estrogen mimics interact directly with the estrogen receptor and have near immediate effects on gene transcription by turning on the expression of a unique set of genes. Using differential display (DD) RT-PCR, we examined changes in gene expression induced by exposure to paper mill effluents. Largemouth bass were exposed to 0, 10, 20, 40, and 80% paper mill effluent concentrations in large flow-through tanks for varied periods of time including 7, 28 or 56 days. Plasma hormone levels in males and females and plasma vitellogenin (Vtg) in females decreased with dose and time. Measurements of changes in gene expression using DD RT-PCR suggest that the gene expression patterns of male fish do not change much with exposure, except for the induction of a few genes including CYP 1A, a protein that is induced through the action of the Ah receptor in response to dioxin and similar polyaromatic hydrocarbons. However, in the case of females, exposure to these effluents resulted in an up-regulation of CYP 1A that was accompanied by a generalized down-regulation of genes normally expressed during the reproductive season. These antiestrogenic changes are in agreement with previous studies in bass exposed to these effluents, and could result in decreased reproductive success in affected populations. PMID- 15288540 TI - Temporal changes in gene expression in the liver of male plaice (Pleuronectes platessa) in response to exposure to ethynyl oestradiol analysed by macroarray and Real-Time PCR. AB - Suppression subtractive hybridisation (SSH) was used to generate cDNA libraries representing genes differentially-expressed in liver from male plaice (Pleuronectes platessa) exposed to ethynyl oestradiol (EE2). BLAST analysis and alignments of the clones with database sequence suggested at least three vitellogenin (VTG) genes and three zona radiata protein (ZRP) genes were represented. Clones with unique sequence (62 up-, 13 down-regulated) were arrayed as probes on nylon membranes to investigate temporal expression of oestrogen responsive genes in experimental animals. Arrays were hybridised with radiolabelled cDNAs prepared from hepatic mRNA from animals treated with EE2 for various times upto 21 days and from treated animals transferred to clean water for upto a further 31 days. By day 21 of treatment 11 out of 17 probes from unidentified genes, 21/22 VTG, 13/14 ZRP, 2/2 liver aspartic proteinase (LAP) and 8/10 other gene sequences were induced by EE2 exposure. Of the down-regulated sequences, only three showed significant, decreased expression and these encode cytochrome b and two with cryptic functions. Based on the pattern of temporal response the up-regulated probes fell into two classes. Pattern A reached maximum expression by day 16 of exposure and then declined prior to removal of EE2 at 21 days. Pattern B genes reached maximal expression between day 16 and 22, declining only after removal of EE2. Independent investigation of the expression patterns of selected probes using quantitative Real-Time PCR reproduced the distinctive patterns. The results indicate a previously unrecognised mechanism for oestrogenic toxicity in which there is a selective down-regulation of some egg proteins, potentially diminishing the quality of eggs and this may contribute to reproductive failure described elsewhere. PMID- 15288541 TI - Functional analysis of chemically-induced mutations at the flounder TP53 locus, the FACIM assay. AB - A functional assay was developed in yeast to identify mutations induced by DNA damaging agents at the flounder TP53 locus. This assay named FACIM for functional analysis of chemically-induced p53 mutations, is based on the assumption that most genotoxin-induced mutations inactivate transcriptional activity of the TP53 protein. The functional status of the protein expressed in yeast was measured using a p53-responsive reporter gene. The FACIM assay was used to evaluate the mutagenesis of the flounder TP53 exposed in vitro to benzo[a]pyrene diol epoxide (BPDE). A dose-dependent increase of p53 mutation rate was observed with increasing concentrations of BPDE and extension of exposure time. Flounder TP53 gene appeared highly sensitive to point mutations since most of those identified targeted different nucleotides. Mutated base-pairs corresponded predominantly to guanines located on the non-transcribed strand of the DNA. The general distribution of mutations along the flounder TP53 protein was different from that identified in the human homologue suggesting species-differences in mutagenesis of the TP53 gene. Most of flounder TP53 mutants were defective for transactivation and cell growth regulation but some maintained a partial wild type phenotype. This functional assay in yeast could be used for both evaluation of the genotoxic potency of chemicals or environmental samples and screening of p53 mutations in fish tumours. PMID- 15288542 TI - A HECT E3 ubiquitin-protein ligase with sequence similarity to E6AP does not target p53 for degradation in the softshell clam (Mya arenaria). AB - Numerous reports have raised the level of national concern that chemicals found in the environment may have adverse effects on the health of humans and wildlife. Environmental exposure to pollutants, such as dioxin, has been implicated in gonadal tumor formation in Maine softshell clams (Mya arenaria). Prevalence of these tumors is as high as 40% in some populations. Although their etiology is still unknown, investigations into the mechanisms of tumor formation have revolved around a hypothesis of dioxin-induced toxicity. The aryl hydrocarbon receptor (AHR) was initially investigated, but was later determined to not bind the prototypical ligand, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), suggesting that dioxin toxicity is mediated through an AHR-independent pathway. An alternative mechanism of tumor formation has been investigated, involving a protein with significant sequence similarity to mammalian E6AP, a HECT (homologous to E6AP carboxy terminus) E3 ubiquitin-protein ligase. E6AP, in association with the high-risk human papillomavirus (HPV) E6 protein, is involved in the abnormal degradation of the p53 tumor suppressor protein in human cervical cancer. Tumorigenic clam reproductive tissue revealed higher M. arenaria E3 (MaE3) protein levels concomitant with lower M. arenaria p53 (Map53) levels. While the function of MaE3 as a HECT E3 was verified, results from three methods agree that MaE3 does not associate with Map53. However, alteration in Map53 levels may still play a role in clam gonadal tumorigenesis. Due to upregulation of MaE3 in neoplastic reproductive tissue, further investigations will focus on determining the proteolytic targets of MaE3. In conjunction with our previous findings that dioxin toxicity in the softshell clam is not mediated by AHR, the results from our current investigation suggest a complex etiology for the clam germinomas. PMID- 15288543 TI - Resistance to contaminants in North American fish populations. AB - Fish from urban and industrialized estuaries are exposed among the highest levels of contaminants of any vertebrate populations. As a result, they serve as especially relevant models for determining the toxic effects and mechanisms through which environmental toxicants work. In controlled laboratory experiments, fish from highly contaminated locales sometimes exhibit resistance to contaminant induced toxicity. Resistance may be due to genetic adaptation or physiological acclimations. Distinguishing between these possibilities is important in predicting the persistence of resistance and its potential costs to affected populations and communities. Along the Atlantic coast of North America, populations of two estuarine species, Atlantic killifish (mummichog) Fundulus heteroclitus and Atlantic tomcod Microgadus tomcod, exhibit phenotypes that are resistant to aromatic hydrocarbon (AH) contaminants, including polychlorinated biphenyls (PCBs), polychlorinated dibenzo-p-dioxins (PCDDs), and polycyclic aromatic hydrocarbons (PAHs). Populations of these species exhibit resistance to AH-induced lethality, early life-stage toxicities, and expression of cytochrome P4501A (CYP1A). However, some differences among populations in the occurrence and type (genetic or physiological) of AH-resistant phenotypes have been observed. In some instances, resistance was obviously genetic and resulted in its transmission to at least the F2 generation, in others, resistance had a physiological or yet to be identified epigenetic basis. In some cases, resistance was observed for all AH compounds tested, in others, it was seen only for halogenated AHs. As toxic responses to AHs are believed to be mediated by the aryl hydrocarbon receptor pathway (AHR), several studies compared the structure and expression of AHR pathway molecules between resistant and sensitive fish populations. However, no obvious differences in these molecular parameters were observed between resistant and sensitive populations at the transcriptional level. Further studies at the protein level are recommended to further evaluate the role of the AHR pathway in conferring resistance. Open-ended microarray and proteomic approaches may provide additional resolution in determining the molecular mechanisms of resistance. Also, studies that evaluate the prevalence and ecosystem cost of resistance are needed. PMID- 15288544 TI - Emerging contaminants--pesticides, PPCPs, microbial degradation products and natural substances as inhibitors of multixenobiotic defense in aquatic organisms. AB - The environmental presence of chemosensitizers or inhibitors of the multixenobiotic resistance (MXR) defense system in aquatic organisms could cause increase in intracellular accumulation and toxic effects of other xenobiotics normally effluxed by MXR transport proteins (P-glycoprotein (P-gps), MRPs). MXR inhibition with concomitant detrimental effects has been shown in several studies with aquatic organisms exposed to both model MXR inhibitors and environmental pollutants. The presence of MXR inhibitors has been demonstrated in environmental samples from polluted locations at concentrations that could abolish P-gp transport activity. However, it is not clear whether the inhibition observed after exposure to environmental samples is a result of saturation of MXR transport proteins by numerous substrates present in polluted waters or results from the presence of powerful MXR inhibitors. And are potent environmental MXR inhibitors natural or man-made chemicals? As a consequence of these uncertainties, no official action has been taken to monitor and control the release and presence of MXR inhibitors into aquatic environments. In this paper we present our new results addressing these critical questions. Ecotoxicological significance of MXR inhibition was supported in in vivo studies that demonstrated an increase in the production of mutagenic metabolites by mussels and an increase in the number of sea urchin embryos with apoptotic cells after exposure to model MXR inhibitors. We also demonstrated that MXR inhibitors are present among both conventional and emerging man-made pollutants: some pesticides and synthetic musk fragrances show extremely high MXR inhibitory potential at environmentally relevant concentrations. In addition, we emphasized the biological transformation of crude oil hydrocarbons into MXR inhibitors by oil-degrading bacteria, and the risk potentially caused by powerful natural MXR inhibitors produced by invasive species. PMID- 15288545 TI - Transgenerational genomic instability as revealed by a somatic mutation assay using the medaka fish. AB - We previously established a somatic mutation assay of the medaka wl (white leucophores) locus based on visual inspection, and showed that somatic mutations at paternally derived alleles frequently arise during the development of F1 embryos fertilized by sperm/late spermatids that had been exposed to gamma-rays. To further study such delayed mutations, we determined the frequency of mutant embryos obtained from three different crosses between irradiated males and non irradiated females. When sperm and late spermatids were irradiated, the mutant frequency within non-irradiated maternally derived alleles was approximately 3 times higher than in the control group. In the F2 generation, however, no increase in mutant frequency was observed. Similarly, there was no significant increase in the F1 mutant frequency when stem spermatogonia were irradiated. These data suggest that irradiation of sperm and late spermatids can induce indirect mutations in F1 somatic cells, supporting the idea that genomic instability arises during F1 embryonic development. Moreover, such instability apparently arises most frequently when eggs are fertilized just after the sperm are irradiated. PMID- 15288546 TI - The random amplified polymorphic DNA (RAPD) assay to determine DNA alterations, repair and transgenerational effects in B(a)P exposed Daphnia magna. AB - The random amplified polymorphic DNA (RAPD) is a useful assay for the detection of genotoxin-induced DNA damage and mutations. In this study, we have further evaluated the potential of this assay to measure benzo(a)pyrene [B(a)P]-induced DNA changes, and repair (in kinetic experiments) as well as transgenerational effects in the water fleas, Daphnia magna. The organisms, which reproduce parthenogenetically, were exposed to 50 microg L(-1) B(a)P for 3 or 6 days and were allowed to recover in clean medium for 12 or 9 days, respectively. Qualitative and quantitative changes were observed in RAPD profiles generated not only from the B(a)P exposed Daphnia but also from previously treated organisms during the recovery experiments. The fact that some of the RAPD changes disappeared at the end of both recovery experiments suggested that the DNA effects were fully repaired or reversed. In addition, some of the B(a)P-induced RAPD alterations detected in parental D. magna were also observed in the offspring patterns. This suggested that DNA alterations that occurred in germ cells were probably transmitted to the next cohorts. The present study shows that the RAPD method can be useful to qualitatively assess the kinetics of DNA changes, repair and transgenerational effects and such effects could potentially be linked to survival and reproductive success at higher levels of biological organisation. In addition, the water fleas have efficient capabilities to repair or reverse B(a)P-induced DNA effects. Finally, unrepaired or misrepaired genetic damage induced by genotoxins such as B(a)P could be transmitted to next generations in these parthenogenetically reproducing organisms. PMID- 15288547 TI - In vitro assessment of DNA damage after short- and long-term exposure to benzo(a)pyrene using RAPD and the RTG-2 fish cell line. AB - Genotoxins present in the aquatic environment are often associated with the decline or disappearance of many wild populations. The hazard assessment of chemicals requires sensitive and specific tests to study the genotoxic effects in order to establish the maximum allowable chemical concentrations prior to the release to the environment. We have previously shown that an established fish cell line (RTG-2) together with the random amplified polymorphic DNA (RAPD) technique, can be used to detect alterations in the DNA caused by direct acting genotoxins. The current study takes this a step further and examines in the same system the effect of a pro-mutagen benzo(a)pyrene (B(a)P) at different concentrations (0.05, 0.1, and 0.5 microg/ml) and at different exposure periods (1, 2, 3, 15, and 30 days). After comparing DNA fingerprints from control and exposed cells, both qualitative and quantitative analysis show an increase in the instability in the DNA fingerprint of exposed cells over a time- and concentration-dependent manner. At the higher concentration (0.5 microg/ml) three out the four primers showed altered bands after 1 day of exposure, while after 3 days all used primers showed an altered pattern. At the lower concentration of B(a)P (0.05 microg/ml) the appearance of new bands was observed with a 100% level of reproducibility after 30 days of exposure suggesting an inheritance of the altered DNA. We conclude that this in vitro system is useful to evaluate genotoxic effects, both after acute and chronic exposures and of direct and non direct acting genotoxins. Cultured cells can be considered as genetically homogenous populations. Therefore, in vitro systems permits us to undertake mechanistic studies avoiding the interference of polymorphisms inherent in the in vivo systems. Furthermore, the RTG-2 fish cell line combined with a RAPD assay could be used in studies of hazard identification in risk assessment protocols of chemicals. PMID- 15288548 TI - Genotoxicity biomarkers in Mytilus galloprovincialis: wild versus caged mussels. AB - A biomonitoring programme of wild and caged mussels (Mytilus galloprovincialis) was carried out at four selected sites along the Ligurian coast: Cornigliano, Voltri, Zinola, and Sanremo (Italy). Mussels of a very narrow size range were left in situ for 30 days. Adult specimen of mussels from natural substrates were collected in the same areas. Animals from a mussel farm located in La Spezia were used as controls. Micronucleus frequency and DNA single strand breaks, evaluated by alkaline elution, were used as biomarkers of genotoxicity. Mussels were also analyzed for polycyclic aromatic hydrocarbons (PAH) and heavy metals (Hg and Cd). Different gradients of PAH and metal concentrations were detected in tissues of mussels from different samplings sites. A weak correlation was found between single strand breaks and PAH content while MN frequency correlated with Hg concentration (r = 0.28, P < 0.002). A clear distinction between the sites, allowing classification along a pollution gradient (Sanremo < Zinola < Voltri < Cornigliano) was demonstrated by the analysis of genotoxicity parameters. The obtained results suggested that the micronucleus assay compared with DNA damage determination by alkaline elution allow to better discriminate the selected sites. DNA damage expressed as constant of elution (k ml(-1) x 10(3)) ranges from 30 +/- 9.6 to 89.60 +/- 40.10, and micronuclei frequency from 1.78 +/- 1.04 to 24.4 +/- 12.9, in control animals and in mussels from the most polluted site, respectively. Wild mussels accumulated significant concentrations of chemicals and showed a higher induction of chromosomal damage than caged mussels, expressed as micronuclei frequency. Caged mussels showed higher level of DNA damage than wild mussels, probably as a result of recent exposure. DNA damage was higher in September than in May, as opposed to micronuclei frequency being higher in May than in September. Endogenous and exogenous factors, such as change of pollutant input levels or compositions, could be considered the cause of such variability. PMID- 15288549 TI - DNA strand breaks and adducts determined in feral and caged chub (Leuciscus cephalus) exposed to rivers exhibiting variable water quality around Birmingham, UK. AB - This study forms part of an investigation into the effects on fish of immersion in three rivers around Birmingham, UK. The rivers Blythe, Cole and Tame exhibit relatively high, intermediate and poor overall water quality, respectively, according to combined levels of polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs) and organochlorine pesticides (OCPs), as well as heavy metals. Specifically, biomarkers of genotoxicity (DNA strand breaks and adducts) were measured in feral and caged chub (Leuciscus cephalus), complementing another study in which data were presented for a number of other hepatic biomarkers measured in the same animals. In both feral and caged chub, there was a general elevation of DNA strand breaks with a decrease in chemical water quality, with some time points exhibiting significantly higher levels at the most (Tame) compared with least polluted sites (Blythe), particularly in the cage-held animals. Combined-season DNA adduct data suggested a higher degree of toxic insult in the feral compared with caged chub and revealed particularly high levels of adducts in fish caught from the Cole. The pattern of adducts shown was typical of exposure to a complex mixture of PAHs which were relatively high, and similar, in both the Cole and Tame. Overall, these data are consistent with exposure of both feral and caged chub to contaminants which are able to induce specific, moderately genotoxic effects. PMID- 15288550 TI - DNA adduct analysis and histopathological biomarkers in European flounder (Platichthys flesus) sampled from UK estuaries. AB - The presence of genotoxic and potentially carcinogenic chemical contaminants in the estuarine and coastal marine environment is well documented. In this study, European flounder (Platichthys flesus) sampled from eight UK estuaries were analysed for hepatic DNA adducts, using the 32P-postlabelling assay and liver histopathology as part of an on going survey to establish the health status of UK estuaries. Fish were collected from the estuaries Tyne, Mersey, Thames, Alde (reference site), Belfast, Forth, Clyde and Southampton. At the majority of contaminated sites (Southampton, Thames, Clyde, Tyne and Mersey) the predominant DNA adduct profile consisted of diagonal radioactive zones (DRZs). In contrast, flounder collected from the Forth, Alde and Belfast lacked DRZs with only background levels of DNA damage being observed. Statistically significant differences were observed between several of the sites sampled with the hepatic DNA adduct levels detected in flounder from Southampton, Thames and Clyde statistically elevated (P < 0.05) over those levels detected at the Tyne (Southampton and Thames only), Forth, Alde and Belfast. Histological analysis of these samples revealed a range of lesions including foci of cellular alteration, hepatocellular fibrillar inclusions, nuclear pleomorphisms along with non toxicopathic changes/alterations, such as those associated with cell turnover (apoptosis, necrosis, regeneration) and immune-related functions (melanomacrophage aggregates, inflammation). Although it is difficult to associate higher prevalence of these lesion types with specific sites, generally, the lowest prevalence was seen in flounder captured from the Alde estuary, with higher prevalence (particularly of melanomacrophage aggregates, inflammation and necrotic foci) seen in fish from the contaminated sites. PMID- 15288551 TI - DNA damage in eelpout (Zoarces viviparus) from Goteborg harbour. AB - The relationship between DNA damage and the exposure of marine organisms to environmental contaminants was examined in the Goteborg harbour area. This research is part of a wider ecotoxicological study planned to evaluate the biological impact of chemical contamination in the River Gota estuary, following a bunker oil (10-100 tonnes) spill occurred in June 2003. Here we present data on the DNA strand breaks derived using the comet assay and the presence of apoptotic cells using the diffusion assay in nucleated erythrocytes of the eelpout (Zoarces viviparus) from the study area and at a clean reference site. Polycyclic aromatic hydrocarbon metabolites were also analyzed in the bile of exposed fish. The results showed a high level of damaged DNA, paralleled by a peak in bile PAH metabolites, in fish from the most impacted site, 3 weeks after the oil spill. A significant recovery was observed in specimens from the spill site, 5 months later, but not in fish caught in the middle part of Goteborg harbour, which is chronically subjected to heavy chemical pollution. The levels of apoptic cells did not show any marked variations, but a significant recovery was observed in fish from the oil impacted site 5 months after the spill. PMID- 15288552 TI - Measurement of DNA adducts and strand breaks in dab (Limanda limanda) collected in the field: effects of biotic (age, sex) and abiotic (sampling site and period) factors on the extent of DNA damage. AB - In the Eastern English Channel, the potential application of the comet assay and post-labelling technique in dab was evaluated for genotoxicity monitoring of the marine environment. The effects of biotic (age, sex) and abiotic (sampling site and period) factors on the extent of DNA lesions were also studied. Female and male dab of two class of size (juvenile and adult) were collected by trawling in different sites in Seine Bay and Somme Bay during September 2001. Single-strand breaks and adducts were, respectively, measured in erythrocytes and the liver. Results obtained for the adult female were compared with those collected during a first cruise in March 2001 [Akcha et al., Mutat Res. 534 (1-2) (2003) 21]. Significant effects of sex and age were demonstrated on the level of strand breaks. Moreover, a significant interaction between age and sex was shown that might indicate the complex influence of other factors on the extent of DNA damage (i.e. reproduction status). In the adult dab, the level of breaks is higher in the male than in the female, whereas the opposite trend was observed for the juvenile. Whatever the sex, the number of DNA breaks is higher in the adult than in the juvenile. For the female dab, significant differences were observed with the comet assay between the Seine Bay and the Somme Bay in March but not in September. This may be due to seasonal variations in the formation of DNA lesions related to variations in lipid content and levels of biotransformation activities and/or to spawning cycles. The presence of genotoxic substances in the study areas was also confirmed by the detection of DNA adducts in each sample analysed. Whereas no effect was shown on the total level of adducts for the tested biotic and abiotic factors, qualitative differences in adduct profiles were observed for each of these factors. For the female dab, comparison of adduct profiles obtained in March and September with one generated by hepatic microsomal activation in dab of a PAH mixture indicated a PAH contamination of the study areas in autumn. These results show the importance of studying the effects of biotic and abiotic factors on the genotoxic endpoints considered to correctly assess the contribution of chemical contamination to the measured biological responses. PMID- 15288553 TI - Interspecies differences in DNA single strand breaks caused by benzo(a)pyrene and marine environment. AB - The presence of DNA single strand breaks in untreated specimens of selected species, mosquito fish Gambusia affinis, painted comber Serranus scriba, blue mussel Mytilus galloprovincialis, spiny crab Maja crispata and sea cucumber Holothuria tubulosa as well as in 10 microg/g benzo(a)pyrene (BaP) treated mosquito fish, blue mussel and spiny crab was measured, using alkaline filter elution. Interspecies differences in alkaline elution profiles were observed and attributed to different lengths of DNA from different sources and to differences in the number of strand breaks present during normal cellular events in different phyla. Spiny crab hemocytes are more sensitive to action of BaP then blue mussel hemocytes and mosquito fish hepatocytes that could be explained by differences in the rates of distinct metabolic reactions and DNA repair among the investigated species. In field study, DNA single strand breaks were measured in hepatocytes of painted comber and in hemocytes of blue mussel and spiny crab from natural population specimens collected at eight sampling sites along Istrian coast, Croatia. Spatial variations in DNA integrity for each species were detected and revealed for the first time that spiny crab is responsive to different environmental conditions. Interspecies variations in the DNA integrity due to environmental conditions, confirmed species specific susceptibility to genotoxicity of certain environment that in long-term may modify the structure of marine communities. The multi-species approach in designing biomonitoring studies was suggested. PMID- 15288554 TI - Applicability of the black slug Arion ater for monitoring exposure to polycyclic aromatic hydrocarbons and their subsequent bioactivation into DNA binding metabolites. AB - The applicability of terrestrial black slugs Arion ater (Mollusca, Gastropoda) was studied for biomonitoring environmental exposure to polycyclic aromatic hydrocarbons (PAHs). In laboratory experiments, slugs were orally exposed to benzo[a]pyrene (BaP) for a short term (3 days) or a long term (119 days) period. Test animals were collected in the field, or were reared under laboratory conditions to ensure that they had no history of PAH-exposure. Benzo[a]pyrene hydroxylase (BPH) activity was measured in the digestive gland as a biomarker for BaP exposure. Bulky DNA adduct formation in kidney was measured as an effect biomarker for BaP bioactivation into DNA-binding metabolites. Although success of clutching was relatively low (5 out of 18 slugs produced egg packages), sufficient number of slugs were obtained to perform exposure experiments due to high hatching (89%) and survival rates (79%). After a short exposure to a relatively high BaP doses of 20 and 200 microg/g fresh feed, a dose-dependent and significant increase of BPH activity and bulky DNA adduct levels could be demonstrated in A. ater. Induction factors were low (two times control level), but optimization of the test conditions yielded a higher BPH induction factor of 4.8 times control level. BPH activity and bulky DNA adduct levels, however, did not increase after a long-term exposure to environmentally relevant BaP doses (upto 0.25 microg/g fresh feed). Based on this lack of response after realistic exposure it is concluded that A. ater is not sensitive to BaP exposure and, therefore, not suitable for monitoring environmental exposure to PAHs. PMID- 15288555 TI - The effects of hydrostatic pressure change on DNA integrity in the hydrothermal vent mussel Bathymodiolus azoricus: implications for future deep-sea mutagenicity studies. AB - Comet and agarose gel electrophoresis (AGE) assays were used to show that haemocytes (blood cells) and gill tissues of vent mussels, Bathymodiolus azoricus, are sensitive to hydrostatic pressure change, but can repair DNA damage induced by retrieval from 840 m to the sea surface. In contrast, animals collected from 1700 m survived for only a few days in the laboratory, which was reflected in their poor DNA quality. These findings support the hypothesis of a physiological barrier to survival around 1000-1500 m depth, which these results show affects both vent and non-vent species alike. Based on in vitro experimental exposures to hydrogen peroxide and MMC, vent mussels appear to have sensitivities to the environmental mutagens that are not significantly different from those of coastal mussels. PMID- 15288556 TI - An integrated biomarker-based strategy for ecotoxicological evaluation of risk in environmental management. AB - Environmental impacts by both natural events and man-made interventions are a fact of life; and developing the capacity to minimise these impacts and their harmful consequences for biological resources, ecosystems and human health is a daunting task for environmental legislators and regulators. A major challenge in impact and risk assessment, as part of integrated environmental management (IEM), is to link harmful effects of pollution (including toxic chemicals) in individual sentinel animals to their ecological consequences. This obstacle has resulted in a knowledge-gap for those seeking to develop effective policies for sustainable use of resources and environmental protection. Part of the solution to this problem may lie with the use of diagnostic clinical-type laboratory-based ecotoxicological tests or biomarkers, utilising sentinel animals as integrators of pollution, coupled with direct immunochemical tests for contaminants. These rapid and cost-effective ecotoxicological tools can provide information on the health status of individuals and populations based on relatively small samples of individuals. In the context of ecosystem status or health of the environment, biomarkers are also being used to link processes of molecular and cellular damage through to higher levels (i.e., prognostic capability), where they can result in pathology with reduced physiological performance and reproductive success. Complex issues are involved in evaluating environmental risk, such as the effects of the physico-chemical environment on the speciation and uptake of pollutant chemicals and inherent inter-individual and inter-species differences in vulnerability to toxicity; and the toxicity of complex mixtures. Effectively linking the impact of pollutants through the various hierarchical levels of biological organisation to ecosystem and human health requires a pragmatic integrated approach based on existing information that either links or correlates processes of pollutant uptake, detoxication and pathology with each other and higher level effects. It is further proposed here that this process will be facilitated by pursuing a holistic or whole systems approach with the development of computational simulation models of cells, organs and animals in tandem with empirical data (i.e., the middle-out approach). In conclusion, an effective integrated environmental management strategy to secure resource sustainability requires an integrated capability for risk assessment and prediction. Furthermore, if such a strategy is to influence and help in the formulation of environmental policy decisions, then it is crucial to demonstrate scientific robustness of predictions concerning the long-term consequences of pollution to politicians, industrialists and environmental managers; and also increase stakeholder awareness of environmental problems. PMID- 15288557 TI - Adversarial safety analysis: borrowing the methods of security vulnerability assessments. AB - INTRODUCTION: Safety and security share numerous attributes. The author, who heads the (Security) Vulnerability Assessment Team at Los Alamos National Laboratory, therefore argues that techniques used to optimize security might be useful for optimizing safety. OPTIMIZING SECURITY: There are three main ways to attempt to improve security-security surveys, risk assessment (or "design basis threat"), and vulnerability assessments. The latter is usually the most effective. SAFETY ANALOGS: Vulnerability assessment techniques used to improve security can be applied to safety analysis--even though safety is not ordinarily viewed as having malicious adversaries (other than hazards involving deliberate sabotage). Thinking like a malicious adversary can nevertheless have benefits in identifying safety vulnerabilities. SUGGESTIONS: The attributes of an effective safety vulnerability assessment are discussed, and recommendations are offered for how such an adversarial assessment might work. CONCLUSION: A safety vulnerability assessment can potentially provide new insights, a fresh and vivid perspective on safety hazards, and increased safety awareness. PMID- 15288558 TI - The ergonomics of children in playground equipment safety. AB - INTRODUCTION: Annually, some 200,000 U.S. children require hospital emergency room treatment from injuries suffered on playground equipment. Most of these injuries (70%) are the result of falls. While standards for protective surfacing within the play zone areas are very adequate, standards for the prevention of falls are very inadequate. "Climbers" are responsible for the greatest percentage of injuries with most of these injuries the result of falls. Current standards are woefully lacking in regard to these climbers. METHOD: This paper reviews the history of climber research and climber standards. RESULTS: It then presents recommendations for improved standards in this area based on the ergonomics of the users, children. PMID- 15288559 TI - Whole-body vibration and postural stress among operators of construction equipment: a literature review. AB - INTRODUCTION: Operators of construction equipment perform various duties at work that expose them to a variety of risk factors that may lead to health problems. A few of the health hazards among operators of construction equipment are: (a) whole-body vibration, (b) awkward postural requirements (including static sitting), (c) dust, (d) noise, (e) temperature extremes, and (f) shift work. It has been suggested that operating engineers (OEs) are exposed to two important risk factors for the development of musculoskeletal disorders: whole-body vibration and non-neutral body postures. METHOD: This review evaluates selected papers that have studied exposure to whole-body vibration and awkward posture among operators of mobile equipment. There have been only few studies that have specifically examined exposure of these risk factors among operators of construction equipment. Thus other studies from related industry and equipment were reviewed as applicable. CONCLUSION: In order to better understand whole-body vibration and postural stress among OEs, it is recommended that future studies are needed in evaluating these risk factors among OEs. PMID- 15288560 TI - Increasing the safety of children's vehicle travel: from effective risk communication to behavior change. AB - INTRODUCTION: When installed and used correctly, child safety seats reduce the risk of fatal injury by 71% for infants and 54% for toddlers. However, four out of five safety seats are unintentionally misused. Yet, parents fail to participate in safety-seat checks and other child seat interventions aimed at correcting misuse. METHOD: Such lack of participation is the focus of this article, which argues that most caregivers are naive to their own vulnerability for misusing their child's seat. Research on risk perception is discussed as a guide to understanding both the high misuse rate and the lack of participation in interventions designed to correct this public safety problem. RESULTS AND IMPACT: A comprehensive intervention plan that incorporates risk communication techniques for maximum parental participation is proposed that includes three essential components: (a) establishing community locations for parents to turn for safety seat advice, (b) making these locations well known to the public, and (c) increasing caregivers' perceptions of risk of misusing their children's seats. PMID- 15288561 TI - Investigating factors that influence individual safety behavior at work. AB - INTRODUCTION: A qualitative study was conducted to investigate the factors that influence individual safety behavior at work. METHOD: Semi-structured interviews were conducted with participants from a variety of occupations. RESULTS: The analysis revealed several organizational and social factors that explain why individuals engage in unsafe work practices. CONCLUSIONS: The influence of organizational/social factors on safety behavior were discussed. The results suggest that important organizational factors, in addition to job design and engineering systems, may be overlooked when identifying the causes of workplace accidents. Such factors include early socialization, and the need to portray a positive image. IMPACT ON INDUSTRY: The implications for management and industry are discussed. PMID- 15288562 TI - An examination of the characteristics and traffic risks of drivers suspended/revoked for different reasons. AB - PROBLEM: Prior research has demonstrated that suspended/revoked drivers pose a significant traffic risk, but until now little has been known about whether, and if so how, this risk varies as a function of the reason for suspension/revocation. METHOD: This study classifies suspended/revoked drivers into subgroups based on their reason for suspension/revocation, and then develops demographic and driving record profiles for each group. Separate driving record profiles are developed for the following traffic safety indicators, measured 3 years prior to the suspension/revocation action: (a) total crashes, (b) fatal/injury crashes, (c) total traffic convictions, and (d) total incidents (crashes + convictions). RESULTS: The findings clearly show that: (a) suspended/revoked drivers are a heterogeneous group, both demographically and in their driving behavior; (b) some suspended drivers, such as those suspended/revoked for a non-driving offense, have low traffic risks that are not much higher than those of validly-licensed drivers, and; (c) all suspended groups have elevated crash and conviction rates, compared to validly-licensed drivers. DISCUSSION: The implications of these findings for current laws and policies targeting suspended/revoked drivers are discussed, and recommendations for improving these laws/policies are presented. IMPACT ON INDUSTRY: None. PMID- 15288563 TI - Project safety as a sustainable competitive advantage. AB - INTRODUCTION: To be consistently profitable, a construction company must complete projects in scope, on schedule, and on budget. At the same time, the nature of the often high-risk work performed by construction companies can result in high accident rates. Clients and other stakeholders are placing increasing pressure on companies to decrease those accident rates. Clients routinely demand copies of safety plans and evidence of past results at the "pre-qualification" or "request for proposal" stages of the procurement process. Are high accident rates and the associated costs just a part of business? FINDINGS: Companies that deliver on scope, schedule, and budget have a competitive advantage. Is it possible for projects with low accident rates to use it as a competitive advantage? Is the value added by safety just a temporary or parity issue, or does a successful safety program offer significant advantage to the company and the client? IMPACT ON INDUSTRY: This article concludes that in the case of a high-risk industry, such as the construction industry, an organization with a successful safety program can promote safety performance as a sustainable competitive advantage. It is a choice the company can make. PMID- 15288564 TI - Using state administrative data to study nonfatal worker injuries: challenges and opportunities. AB - PROBLEM: Administrative data from states have the potential to capture broader representation of worker injury, facilitating examination of trends, correlates, and patterns. While many states use their workers' compensation (WC) data to document frequency and type of injury, few conduct in-depth examinations of patterns of injury and other etiologies. Administrative data are generally an untapped resource. METHOD: Comparisons are made among four state databases used in a study linking worker injuries and patient outcomes in hospitals and nursing homes. RESULTS: Worker injury data varies in terms of inclusion criteria, variables, and coding schemes used. Linkages to organizational level characteristics can be difficult. CONCLUSIONS: Despite limitations, data can be used to study injury patterns and etiologies. Users must be knowledgeable and recognize how database characteristics may influence results. PMID- 15288565 TI - Computer-based training for food services workers at a hospital. AB - INTRODUCTION: Interactive computer-based safety instruction (CBI) was given to 73 workers in the food services department of an urban hospital. RESULTS: Post-test accuracy (95%) improved significantly (p < or = 0.0001) from the pre-test (74.5 %), d = 1.09. Generalization was confirmed by increased accuracy in answering questions, posed on-the-job, that required application of knowledge to the work setting (from 46% to 79%; p < 0.0001). Problematic kitchen conditions such as puddles increased slightly after training, but adjustment for increasing production/workload revealed an overall post-training decline in problems from 0.58 to 0.32 (p = 0.0001, d = 0.89). Work practice improvement was seen in 79% of workers (p < 0.0001, d = 1.00). Effect sizes (d) of knowledge, location, and work practice improvements are large and demonstrate that the benefits of CBI extend to the workplace floor. Further, the decrease between knowledge and behavior change (d = 0.09-0.2) is less than reported following other forms of training. PMID- 15288566 TI - Factorial structure of recklessness: to what extent are older drivers different? AB - PROBLEM: This study tests whether the original factorial structure of a recklessness questionnaire can be maintained for the current Spanish population of older drivers. JUSTIFICATION: Our recent interest in dedicating special attention to senior citizen mobility (Monterde, 2001), is due to the impending increase of the aging population in Western countries; this has led us to reinitiate the psychometric study of the construct validity, revising and including older drivers in the psychometric aspects of those evaluation instruments that will then be used in the Spanish psycho-medical check of drivers and in research. METHODS: Factorial analysis was used to determine validity. RESULTS: There was an appearance of a different psychological pattern in elderly drivers, specifically, a psychological anxiety trait related to the task of driving. Furthermore, interesting data were found about the attitude of this sector toward the "traffic society" and toward some of the measures included in Spanish legislation that affect especially older drivers (such as the psycho medical check). IMPACT ON INDUSTRY, RESEARCH AND PRACTICE: These results suggest the possible existence of some kind of "compensation phenomenon," which could have influence over the scores obtained and their interpretation. Consequently, the evaluation instruments should be tested and, if necessary, adapted or specifically created for use with this age group. PMID- 15288567 TI - Michigan's graduated driver licensing program: evaluation of the first four years. AB - INTRODUCTION: To evaluate the four-year outcome of Michigan's graduated driver licensing (GDL) program, motor-vehicle crash data for 16-year-old drivers in 1996 (pre-GDL), and 1998-2001 (post-GDL) were analyzed. METHOD: Relative risks and 95% confidence intervals for several crash types were computed, and pre-post-GDL population-based crash rates were compared. Reductions in crash risks among 16 year-olds previously found in 1998 and 1999 were generally maintained in 2000 and 2001. RESULTS: Reductions in crash risk among 16-year-olds from 1996 to 2001 were 29% for all, 44% for fatal, 38% each for nonfatal-injury and fatal-plus-nonfatal injury, 32% for day, 31% for evening, 59% for night, 32% for single-vehicle, and 28% for multi-vehicle crashes. Even after adjusting for more general population wide changes among drivers 25 years and older that might have contributed to changes in 16-year-old crash risk, reductions remained impressive (19% for all crashes in 2001). IMPACT ON INDUSTRY: As one approach to reducing teenage motor vehicle morbidity and mortality, GDL remains promising. PMID- 15288568 TI - Crisis management of SARS in a hospital. AB - INTRODUCTION: A large general hospital was suddenly disabled by an in-hospital outbreak of Severe Acute Respiratory Distress Syndrome (SARS). METHOD: The crisis was successfully managed by a Standard Operation Procedure (SOP) that included: (a) containment of SARS patients on a special floor and evacuation of the patients from the infected and near-around floors; (b) sorting of the hospital into areas and floors to avoid cross contact of people; (c) triage of patients into groups according to risks; (d) closure of the emergency room and outpatient clinics; and (e) set up of an outdoor fever screening station and emergency service. RESULTS: The situation was quickly controlled after the implementation of these procedures. The central argument in this case is that crisis managerial behavior is the result of how managers channel and distribute the attention of their crisis sense. IMPACT ON INDUSTRY: What managers should do depends on what risk issues and actions related to risk independency, efficiency, safety priority, and transparency they take. What risk issues and actions they take depends on the crisis sense and on how management responds to leadership, resource, and execution. PMID- 15288569 TI - Characterisation of GEA 3175 on human platelets; comparison with S-nitroso-N acetyl-D,L-penicillamine. AB - By comparing the effect of two nitric oxide (NO)-containing compounds, we found that S-nitroso-N-acetyl-D,L-penicillamine (SNAP), but not GEA 3175 (1,2,3,4 Oxatriazolium,3-(3-chloro-2-metylphenyl)-5-[[(4-methylphenyl)sulfonyl]amino]-, hydroxide inner salt), released NO. Despite this, both drugs elevated cyclic guanosine 3',5'-monophosphate (cGMP) levels in human platelets. However, SNAP was more effective after short exposure times (5 and 20 s). The compounds also inhibited thrombin-induced rises in cytosolic Ca2+. Time studies revealed that the action of SNAP rapidly declined by increasing the length of incubation (from 5 s to 30 min). This desensibilisation phenomenon mainly involved the release of Ca2+ from intracellular stores. In comparison, GEA 3175-induced inhibition of cytosolic Ca2+ signalling was much more long-lasting. The soluble guanylyl cyclase (sGC) inhibitor 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) reversed the effect of GEA 3175 on cytosolic Ca2+. Consequently, this inhibition depends solely on the increase in cGMP. In summary, differences between GEA 3175 and SNAP were observed in NO releasing, cGMP elevating and Ca2+ suppressive properties. PMID- 15288570 TI - Terfenadine induces thymocyte apoptosis via mitochondrial pathway. AB - The treatment of rat thymocytes with 10 microM terfenadine resulted in a significant increase in DNA fragmentation. The DNA fragmentation induced by terfenadine was dependent on its concentration and incubation time. In terfenadine-treated cells, the translocation of phosphatidylserine from the inside of plasma membrane to the outside, an early event of the apoptotic process, and chromatin condensation, the morphological characterization of apoptotic cell death, were observed. Terfenadine stimulated caspase-8, -9 and -3 like activities in an incubation time-dependent manner in thymocytes. The active forms of caspase-3 and -9 were detected in the extract from terfenadine-treated cells by immunoblotting analysis using specific antibodies to caspases, but active caspase-8 was not found in this fraction. Decrease in mitochondrial membrane potential and the release of cytochrome c from mitochondria to cytosol were observed in terfenadine-treated thymocytes. These results suggest that terfenadine induces apoptosis in rat thymocytes via mitochondrial pathway. PMID- 15288571 TI - Penicillin blocks human alpha 1 beta 1 and alpha 1 beta 1 gamma 2S GABAA channels that open spontaneously. AB - We used the open-channel blocker, penicillin (10 mM), as a tool to investigate if the human alpha1beta1 or alpha1beta1gamma2S gamma-aminobutyric acid type A (GABAA) receptor channels opened in the absence of GABA. Application of penicillin to cells expressing the receptors resulted in a transient inward whole cell current, the off-current, upon penicillin removal. The amplitude of the off current was dependent on the duration of the penicillin application, it reversed in polarity at depolarized potentials and exhibited "run-down" similar to the GABA-activated currents. Bicuculline (100 microM) blocked the off-current response. Pentobarbital (50 microM) enhanced the peak off-current amplitude by 2.8 and 3.4 in alpha1beta1 and alpha1beta1gamma2S receptors, respectively. Diazepam (1 microM) only enhanced the off-current peak response in alpha1beta1gamma2S receptors (1.6) and induced the development of an inward current when applied alone. The results are consistent with that the alpha1beta1 or alpha1beta1gamma2) GABAA receptors can open in the absence of GABA and raise the question of what role spontaneous channel openings have in the function of GABAA receptors. PMID- 15288572 TI - Anandamide transport inhibitor AM404 and structurally related compounds inhibit synaptic transmission between rat hippocampal neurons in culture independent of cannabinoid CB1 receptors. AB - N-(hydroxyphenyl)-arachidonamide (AM404) is an inhibitor of endocannabinoid transport. We examined the effects of AM404 on glutamatergic synaptic transmission using network-driven increases in intracellular Ca2+ concentration ([Ca2+] spikes) as an assay. At a concentration of 1 microM AM404 inhibited [Ca2+]i spiking by 73+/-8%. The cannabinoid CB1 receptor antagonist N-(piperidin 1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3 carboxamide hydrochloride (SR141716A), the vanilloid VR1 receptor antagonist capsazepine (CPZ), and treatment with pertussis toxin failed to block AM404 mediated inhibition. AM404 (3 microM) inhibited action-potential-evoked Ca2+ influx by 58+/-3% but failed to affect calcium influx evoked by depolarization with 30 mM K+, suggesting that the inhibition of electrically evoked [Ca2+]i increases and that [Ca2+]i spiking was due to inhibition of Na+ channels. Palmitoylethanolamide (PMEA), capsaicin (CAP) and (5Z,8Z,11Z,14Z)-N-(4-hydroxy-2 methylphenyl)-5,8,11,14-eicosatetraenamide (VDM11), compounds structurally similar to AM404, inhibited [Ca2+]i spiking by 34+/-10%, 42+/-18% and 67+/-12%, respectively. Thus, AM404 and related compounds inhibit depolarization-induced Ca2+ influx independent of cannabinoid receptors, suggesting caution when using these agents as pharmacological probes to study synaptic transmission. PMID- 15288573 TI - Concentration-dependent differential effects of quercetin on rat aortic smooth muscle cells. AB - Quercetin is one of the most ubiquitous bioflavonoids in foods of plant origin. Although quercetin is generally considered to provide protection against oxidative injury and inflammation, recent studies have demonstrated that its cytoprotective effects occur within a narrow concentration range. We attempted to examine the concentration-dependent effect on proliferation and inflammation in the primary culture of rat aortic smooth muscle cells. We demonstrate that quercetin inhibited [3H]thymidine incorporation into rat aortic smooth muscle cells only at concentrations < or =50 microM in a concentration-dependent manner. Nevertheless, quercetin, at concentrations > or =100 microM, reduced cell viability; this was further characterized as being due to apoptosis, which occurred through the proteolytic activation of pro-caspase-3. Additionally, the phosphorylation of c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (p38MAPK) substantially increased in rat aortic smooth muscle cells exposed to 100 microM quercetin, results which differ from observations by others and ourselves of cells exposed to < or =50 microM quercetin. Unlike P-JNK and P-p38, the phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/ERK2) was not significantly affected by the concentration-dependent effects of quercetin. Surprisingly, the adverse effects of higher concentrations of quercetin could be ameliorated by adding the antioxidants, catalase, and N acetylcysteine (NAC). Furthermore, the electrophoretic mobility shift assay (EMSA) showed that rat aortic smooth muscle cells exposed to quercetin at concentrations of < or =50 microM caused concentration-dependent inhibition of nuclear factor kappa B (NF-kappaB) activity, whereas concentrations of > or =100 microM resulted in increased NF-kappaB binding activity. We demonstrate for the first time that quercetin at low concentrations has antiproliferative and antiinflammatory effects, but at concentrations of > or =100 microM, is likely to induce the opposite effects on rat aortic smooth muscle cells. PMID- 15288574 TI - Effects of amiodarone on mutant Na+/Ca2+ exchangers expressed in CCL 39 cells. AB - Using the whole cell voltage clamp, we reported previously that amiodarone acutely inhibits Na+/Ca2+ exchange current (INCX) in guinea pig cardiac ventricular myocytes. Intracellular application of trypsin via the patch pipette attenuated the blocking effect of amiodarone, suggesting that amiodarone affects the Na+/Ca2+ exchanger (NCX) from the cytoplasmic side. Here, we attempted to detect the site of amiodarone inhibition using wild type NCX1, mutants, and NCX3 expressed in CCL39 fibroblasts. INCX was recorded by ramp pulses. Amiodarone at 30 microM inhibited INCX by 80% in cells expressing wild type NCX1. However, 30 microM amiodarone inhibited INCX by about 55% in cells expressing mutant NCX1 with amino acids 217-671 (DeltaXIP) or 247-671 (Delta247-671) deleted in the long intracellular loop between the transmembrane segments (TM) 5 and 6. INCXs from NCX mutants deleted of cytoplasmic TM1-2, TM3-4 or the C-terminus were inhibited by amiodarone to a similar extent as the wild type. Amiodarone also inhibited INCX of NCX3 by 76%. These results suggest that a long intracellular loop may be involved in the inhibition of NCX1 by amiodarone, but that other intracellular loops, XIP region or C terminus are not involved in the amiodarone inhibition of NCX1. PMID- 15288575 TI - In vitro comparative assessment of the scavenging activity against three reactive oxygen species of non-steroidal anti-inflammatory drugs from the oxicam and sulfoanilide families. AB - The study of the interaction of non-steroidal anti-inflammatory drugs (NSAIDs) with several reactive oxygen species is of great interest in inflammatory conditions where an uncontrolled release of these potentially damaging intermediates has been documented. This study focused on the scavenging of three species (hydroxyl radical, hydrogen peroxide and hypochlorous acid) with several members of the oxicam family and with the sulfoanilide nimesulide. Reaction with hydroxyl radical was assessed by the modified deoxyribose assay, and rate constants were calculated showing values between 0.8 and 1.1 x 10(10) M(-1) s(-1) for oxicams and of about 0.9 x 10(10) M(-1) s(-1) for nimesulide and ibuprofen. These were consistent with those of the literature but in the same range as those for other NSAIDs and for several thiol-containing molecules. The study of hydrogen peroxide scavenging by the horseradish peroxidase (HRP) assay lacked specificity but no interaction could be evidenced by the glutathione peroxidase assay. The scavenging of hypochlorous acid was finally investigated by the recently developed para-aminobenzoic acid assay which demonstrated better performances for meloxicam (1.7 x 10(4) M(-1) s(-1)) as compared to the other oxicams (tenoxicam: 4.0 x 10(4) M(-1) s(-1), piroxicam: 3.6 x 10(4) M(-1) s(-1), lornoxicam: 4.3 x 10(4) M(-1) s(-1)) and nimesulide (2.3 x 10(3) M(-1) s(-1)). These rate constants were, however, lower than those for thiol-containing molecules and ascorbate. These results suggest that the antioxidant properties of NSAIDs could be influenced by a proper pharmacomodulation as far as the scavenging of hypochlorous acid is concerned while the interest is quite limited for the scavenging of hydroxyl radical. PMID- 15288576 TI - Deglycosylation of Fas receptor and chronic morphine treatment up-regulate high molecular mass Fas aggregates in the rat brain. AB - This study was designed to immunodetect and characterize Fas receptor aggregates (oligomerization) in the brain and to assess its possible modulation in opiate addiction. High molecular mass, sodium dodecyl sulfate (SDS)- and beta mercaptoethanol-resistant Fas aggregates (approximately 110/120 and approximately 203 kDa specific peptides) were immunodetected with a cytoplasmic domain-specific antibody in brain tissue (rat, mouse and human) and SH-SY5Y cells by Western blot analysis. Preincubation of rat cortical membranes with N-ethylmaleimide (NEM; 1 mM for 1 h at 37 degrees C) reduced the immunodensity of approximately 203 kDa Fas aggregates (51%) and increased that of 35 kDa native Fas (172%) and 51/48 kDa glycosylated Fas (47%), indicating that disulfide bonds are involved in Fas dimerization. Enzymatic N-deglycosylation of Fas receptor increased the content of Fas aggregates (approximately 110/120 kDa: five- to sixfold, and approximately 203 kDa: two- to threefold), suggesting that Fas glycosylation is involved in regulating receptor dimerization. Chronic (10-100 mg/kg for 5 days), but not acute (30 mg/kg for 2 h), treatment with morphine (a micro-opioid peptide receptor agonist) induced up-regulation of Fas aggregates in the brain (approximately 110/120 kDa: 39%, and approximately 203 kDa: 89%). The acute and/or chronic treatments with delta- and kappa-opioid peptide receptor agonists and with a sigma1-receptor agonist did not readily alter the content of Fas aggregates in the rat brain. The results indicate that Fas aggregates are natively expressed in the brain and that its density is regulated by the state of Fas glycosylation. These forms of Fas (receptor homodimerization) are functionally relevant because they were up-regulated in the brain of morphine dependent rats. PMID- 15288577 TI - Xenon suppresses nociceptive reflex in newborn rat spinal cord in vitro; comparison with nitrous oxide. AB - Although analgesic action of xenon has been reported, little is known about the effect of xenon at the spinal cord, which plays a crucial role in nociceptive transmission. We studied the effect of xenon on nociceptive reflex (the slow ventral root potential) and the monosynaptic reflex in neonatal rat spinal cord in vitro in comparison with nitrous oxide. Xenon (30%) and nitrous oxide (30%) were applied for 17 min through superfusing artificial cerebrospinal fluid. Xenon and nitrous oxide significantly reduced the amplitude of nociceptive reflex by approximately 70% and approximately 25%, respectively. Xenon and nitrous oxide also significantly reduced the amplitude of the monosynaptic reflex by approximately 35% and approximately 15%, respectively. These results indicate that xenon suppressed the synaptic transmission at the spinal cord, especially those of the slow ventral root potential, which reflect nociceptive transmission. PMID- 15288578 TI - Endothelin ETB receptors inhibit articular nociception and priming induced by carrageenan in the rat knee-joint. AB - The participation of the endothelin system on nociception and priming induced by carrageenan in the knee-joint was investigated. Intra-articular (i.a.) carrageenan (300 microg) caused long-lasting nociceptive effects (i.e., increases in paw elevation time [PET]), which were potentiated by endothelin-1 (dual endothelin ETA/ETB receptor agonist) and inhibited by sarafotoxin S6c (endothelin ETB receptor agonist; both at 30 pmol, i.a., 24 h beforehand). Priming the naive joint with carrageenan augmented nociceptive responses to a second carrageenan challenge, 72 h later. Carrageenan-induced priming, but not nociception, was potentiated by local BQ-788 (10 nmol, i.a., 15 min before priming; endothelin ETB receptor antagonist; N-cis-2,6-dimethylpiperidinocarbonyl-L-gamma-methylleucyl-D 1-methoxycarbonyl-tryptophanil-D-norleucine), but BQ-123 (endothelin ETA receptor antagonist; cyclo [D-Asp-Pro-D-Val-Leu]) was ineffective. Sarafotoxin S6c markedly suppressed carrageenan-induced priming to nociception triggered by carrageenan, endothelin-1 or sarafotoxin S6c, and BQ-788 prevented this action. Thus, selective endothelin ETB receptor agonists inhibit carrageenan-induced nociception and priming in the naive joint. This priming effect of carrageenan to nociception evoked by subsequent inflammatory insults is limited by an endothelin ETB receptor-operated mechanism. PMID- 15288579 TI - Evidence that GABAergic neurons in the spinal trigeminal nucleus are involved in the transmission of inflammatory pain in the rat: a microdialysis and pharmacological study. AB - The aim of this experiment was to investigate the role of the gamma-aminobutyric acid (GABA)-ergic transmission in the nociception within the spinal trigeminal nucleus. The formalin test was used as an animal model of inflammatory pain. Two groups of six rats were used. The behavioural response to the labial injection of formaldehyde (50 microl of a 5% solution) (group 1) or saline (group 2) was evaluated by recording the time spent in facial grooming during a period of 8 min (one period before and seven consecutive periods after the injection). The extracellular concentration of GABA in the trigeminal caudalis nucleus was evaluated, during the formalin test, on samples of 30 microl each (one sample before and three samples after the labial injection) obtained by microdialysis and analysed by HPLC with electrochemical detection of the o-phtalaldeyde pre column derivate. Subsequently, three more groups of six rats each were injected with saline, muscimol (GABAa receptor agonist), or bicuculline (GABAa receptor antagonist) in the trigeminal caudalis nucleus, before performing the formalin test. The injection of formaldehyde induced a biphasic behavioural response and an increase of the GABA levels at 15-45 min. The injection of bicuculline, but not muscimol or saline, strongly decreased the behavioural response of the formalin test. These findings suggest that GABAergic neurons in the trigeminal caudalis nucleus are involved in the transmission of nociceptive information. PMID- 15288580 TI - Alpha 2-adrenoceptors and 5-HT receptors mediate the antinociceptive effect of new pyrazolines, but not of dipyrone. AB - In this study, we investigated whether spinal noradrenergic and serotonergic systems are involved in the antinociception induced by the novel pyrazolines 3 methyl- and 3-phenyl-5-hydroxy-5-trichloromethyl-4,5-dihydro-1H-1-pyrazole-1 carboxyamide (MPCA and PPCA, respectively), and the pyrazolinone dipyrone in the acetic acid writhing (stretching) test in mice. Intrathecal (i.t.) administration of methysergide (3 and 10 microg) and yohimbine (3 microg), but not of prazosin (0.3 and 1 microg) prevented the antinociceptive action of MPCA and PPCA (500 micromol/kg, s.c.). Dipyrone-induced antinociception (500 micromol/kg, s.c.) was not affected by methysergide or adrenoceptor antagonists. These results suggest that spinal 5-HT receptors and alpha2-adrenoceptors are involved in the antinociception induced by MPCA and PPCA, but not in that elicited by dipyrone. PMID- 15288581 TI - Reversal of delta 9-tetrahydrocannabinol-induced tolerance by specific kinase inhibitors. AB - Tolerance develops to the pharmacological effects of Delta9-tetrahydrocannabinoid (THC) following repetitive administration. Adaptations in signaling pathways involved in tolerance to THC-induced behaviors are not understood. The objective of our study was the evaluation of kinase involvement in the expression of tolerance to the above four THC-induced behaviors. Kinase inhibitors that specifically inhibit cyclic AMP-dependent protein kinase (PKA), cyclic GMP dependent protein kinase (PKG), calmodulin-dependent protein kinase (PKC) and src tyrosine kinase were tested for reversal of tolerance to THC's effects. PKG and PKC inhibitors did not reverse tolerance in any behavioral measure. Src tyrosine kinase inhibition reversed tolerance to only the hypoactive effects of THC. PKA inhibition reversed tolerance to all measures, although the doses of inhibitor and time-course of inhibition varied among behaviors. Thus, our data suggest that PKA activity plays a major role in THC-induced tolerance, and that THC produces its multiple effects through different signaling pathways. PMID- 15288582 TI - Depletion of Ca2+ from intracellular stores potentiates spontaneous contractions of the rat portal vein. AB - Spontaneous contractions of the rat portal vein were potentiated in magnitude by phenylephrine, cyclopiazonic acid, ryanodine or caffeine. All these drugs can deplete Ca2+ from intracellular stores, which stimulates store-operated cation entry in some tissues. The possibility that depletion of Ca2+ from intracellular stores potentiates the spontaneous contractions was therefore investigated using functional experiments. Phenylephrine or cyclopiazonic acid was added to tissues in Ca2+-free Krebs solution, followed by a 30-min washout. After addition of extracellular Ca2+, the spontaneous contractions were potentiated. This showed the stimulus for potentiating the contractions remained so long as intracellular Ca2+ stores were depleted. Following phenylephrine washout in normal Krebs solution, potentiation of the spontaneous contractions was attenuated with time. This attenuation was abolished by the protein kinase C inhibitor calphostin C. These results show depletion of Ca2+ from intracellular stores potentiates spontaneous contractions of the portal vein. Protein kinase C may inhibit this mechanism. PMID- 15288583 TI - Involvement of nitric oxide in amiodarone- and dronedarone-induced coronary vasodilation in guinea pig heart. AB - Amiodarone, a powerful antiarrhythmic compound, possesses coronary and peripheral vasodilator properties. The mechanisms responsible for these effects remain incompletely understood. In the present study, the coronary effects of amiodarone and dronedarone, a non-iodinated amiodarone-like compound, were investigated in isolated guinea pig hearts perfused at constant flow with high K+ solution (40 mM). Amiodarone (0.01-10 microM), dronedarone (0.01-1 microM) and verapamil (0.01 10 microM) induced concentration-dependent decreases in coronary perfusion pressure. Amiodarone- and dronedarone-mediated reductions in coronary perfusion pressure were not modified by a cyclooxygenase inhibitor, indomethacin (3 microM). L-Nitro-L-arginine (L-NOARG; 3-100 microM) caused a rightward shift of concentration-response curves to amiodarone and dronedarone in a dose-dependent way; L-arginine, a nitric oxide (NO) precursor, reversed this effect. Furthermore, when guinea pigs were treated with NG-nitro-L-arginine methyl ester (L-NAME; 20 mg/kg), amiodarone could not reduce coronary perfusion pressure. NO synthase inhibition did not affect the coronary perfusion pressure response to verapamil. 1H-[1,2,4]Oxadiazole (4,3-a) quinoxalin-1-one (ODQ), a specific inhibitor of the guanylyl cyclase, inhibited the effects of amiodarone but not those of verapamil. In the presence of L-NOARG and ODQ, and in hearts from animals treated with L-NAME, a decrease in coronary perfusion pressure was still observed at the highest concentration of dronedarone. These results show that, in guinea pig hearts, the coronary vasodilation induced by amiodarone highly depends on nitric oxide. Dronedarone differs from amiodarone by a remaining relaxant effect, refractory to inhibition of NO synthase pathway, probably due to its Ca+ antagonist properties. PMID- 15288584 TI - The orally active nonpeptide selective endothelin ETA receptor antagonist YM598 prevents and reverses the development of pulmonary hypertension in monocrotaline treated rats. AB - We investigated the preventive and therapeutic effects of the selective endothelin ETA receptor antagonist potassium(E)-N-[6-methoxy-5-(2-methoxyphenoxy) 2-(pyrimidin-2-yl)pyrimidin-4-yl]-2-phenylenthenesulfonamidate (YM598) on the development of pulmonary hypertension in monocrotaline-induced pulmonary hypertensive and hypoxemic rats. In the prevention study, oral administration of YM598 (0.1 and 1 mg/kg) or bosentan (30 mg/kg) for 4 weeks was started on the day following monocrotaline (60 mg/kg) injection. In the therapeutic study, oral administration of YM598 (0.1, 0.3 and 1 mg/kg) for 2 weeks was started 3 weeks after monocrotaline injection. In the prevention study, a marked increase in pulmonary arterial pressure and right ventricular hypertrophy, a decrease in right cardiac function and hypoxemia were observed. Histopathological examination indicated the presence of pulmonary remodeling, including medial wall thickening of the pulmonary microvasculature and alveolar disorders. YM598 suppressed the increase in pulmonary arterial pressure, right ventricular hypertrophy and systemic congestion, and improved the hypoxemia, but bosentan had only modest effects. Histopathological disorders were also ameliorated by YM598. In the therapeutic study, YM598 also ameliorated the pulmonary hypertension and hypoxemia in monocrotaline-treated rats. These results suggest that YM598 effectively prevented and reversed the development of pulmonary hypertension, and reduced the pulmonary vascular remodeling and parenchymal injury in monocrotaline treated rats. YM598 also improved hypoxemia which accompanied with the severe pulmonary hypertension in these rats. PMID- 15288585 TI - Aldehyde dehydrogenase, nitric oxide synthase and superoxide in ex vivo nitrate tolerance in rat aorta. AB - The role of aldehyde dehydrogenase (ALDH) in ex vivo tolerance to transdermal glyceryl trinitrate was explored in rat aorta. ALDH activity, measured by aldehyde-induced NADH formation, was strongly depressed in the tolerant arteries. ALDH inhibitors, chloral hydrate (0.3 mM) and cyanamide (0.1-1 mM) inhibited relaxation to glyceryl trinitrate in non-tolerant and tolerant arteries. The inhibition differed from tolerance in that (a) the glyceryl trinitrate concentration-response curve was sigmoidal cf. biphasic in tolerance, (b) the potentiating effect of nitric oxide synthase (eNOS) inhibition was unchanged cf. increased in tolerance and (c) superoxide inhibited the response cf. no significant effect in tolerant or non-tolerant arteries. Hence, reduced ALDH activity does not account fully for ex vivo tolerance. The discrepancies are consistent with evidence that (a) organic nitrates, unlike chloral and cyanamide, irreversibly inactivate ALDH (hence reduced enzyme saturability can explain the biphasic curve) and (b) eNOS contributes to tolerance by a mechanism independent of glyceryl trinitrate metabolism. PMID- 15288586 TI - Cocaine-induced relaxation of isolated rat aortic rings and mechanisms of action: possible relation to cocaine-induced aortic dissection and hypotension. AB - Cocaine HCl is well known for its toxic effects on the cardiovascular system, but little is known about its effects on different regional blood vessels. We designed experiments to determine if cocaine HCl could influence the tension of isolated aortic rings, i.e., induce contraction or relaxation. Surprisingly, cocaine HCl (1 x 10(-5) to 6 x 10(-3) M) relaxed isolated aortic rings precontracted by phenylephrine in a concentration-dependent manner. No significant differences were found between intact or denuded isolated aortic rings (P>0.05). The maximal % relaxations of intact vs. denuded isolated aortic rings were 108.9+/-24.3% vs. 99.5+/-8.3% (P>0.05). Cocaine HCl, 2 x 10(-3) M, was found to inhibit contractions by phenylephrine; EC50s were increased (P<0.01) and Emax's were decreased (51.3+/-16.4% vs. 89.8+/-10.6%, P<0.01). A variety of amine antagonists could not inhibit the relaxant effects of cocaine HCl (P>0.05). The cyclooxygenase-1 inhibitor, indomethacin, also failed to inhibit relaxations induced by cocaine HCl (P>0.05). Neither L-arginine, NG-monomethyl-L-arginine (L NMMA), nor methylene blue could inhibit the relaxations induced by cocaine HCl (P>0.05), suggesting cocaine HCl does not relax isolated aortic rings by inducing the synthesis or release of nitric oxide (NO) or prostanoids from either endothelial or vascular muscle cells. Inhibitors of cAMP, cGMP and protein kinase G (PKG) also failed to inhibit cocaine-induced relaxations. Cocaine HCl (1 x 10( 5) to 6 x 10(-3) M) could also relax isolated aortic rings precontracted by phenylephrine in high K+ depolarizing buffer. Surprisingly, calyculin A, an inhibitor of myosin light chain (MLC) phosphatase, inhibited cocaine-induced relaxations in a concentration-dependent manner, suggesting the probable importance of cocaine-induced MLC phosphatase activation in rat aortic smooth muscle cells. It was also found that cocaine HCl could dose-dependently inhibit Ca2+-induced contractions of isolated aortic rings in high K+-Ca2+-free buffer, suggesting that cocaine HCl may inhibit Ca2+ influx and/or intracellular release. PMID- 15288587 TI - The cardiovascular and renal effects of a highly potent mu-opioid receptor agonist, cyclo[N epsilon,N beta-carbonyl-D-Lys2,Dap5]enkephalinamide. AB - Investigation of the acute cardiovascular and renal effects of cyclo[Nepsilon,Nbeta-carbonyl-D-Lys2,Dap5]enkephalinamide (cUENK6), the most potent mu-opioid receptor agonist, revealed dose-related effects, but most pronounced during the first hour post i.v. injections. During first hour, cUENK6 (3 microg/rat) stimulated (P<0.001) excretion of urine (1.1+/-0.2 vs. 3.3+/-0.3 ml/h), sodium (60+/-10 vs. 124+/-12 microeq/h), potassium and cGMP (1.76+/-0.19 vs. 4.92+/-0.80 nmol/h). These effects were inhibited by naloxone (4 mg/kg i.v.), but not by naloxonazine (35 mg/kg s.c.), or 4 mg/kg i.v. naloxone methiodide. cUENK6 stimulated urinary atrial natriuretic peptide (ANP)-like activity (113+/ 12 vs. 167+/-20 pg/h, P<0.02) and the effect was totally abolished by naloxone. cUENK6 also suppressed the transient stress-induced elevation in blood pressures and heart rate that occurred over the first 30-min post-injection, an effect attenuated by naloxone. Plasma ANP increased 2-h post-injection (123+/-11 vs. 192+/-21 pg/ml, P<0.005), and was associated with augmented ANP mRNA levels in right atria and left ventricles. Thus, cUENK6 evokes renal effects by enhancing activity of the renal natriuretic peptide system. PMID- 15288588 TI - Urocortin does not reduce the renal injury and dysfunction caused by experimental ischaemia/reperfusion. AB - Recent evidence indicates that activators of the serine/threonine kinase pathway protect against ischaemia/reperfusion. Here, we investigate the effects of renal ischaemia/reperfusion on the degree of renal dysfunction and injury with urocortin in rats. Rats treated with urocortin or its vehicle (saline) were subjected to bilateral renal artery occlusion (45 min) and reperfusion (6 h). At the end of experiments, the following indicators and markers of renal injury and dysfunction were measured: plasma urea, creatinine and aspartate aminotransferase, urine flow and creatinine clearance. Urocortin (1 or 15 microg/kg i.v.), administered 5 min prior to reperfusion, was not able to significantly reduce plasma urea, creatinine and aspartate aminotransferase indicating a non-protective effect on the renal dysfunction and reperfusion injury caused by ischaemia/reperfusion. In addition, 15 microg/kg urocortin significantly depressed urine flow and creatinine clearance, which was associated with a significant depression in mean arterial pressure, indicating reduced renal perfusion. Thus, we propose that the pharmacological application of urocortin does not reduce the renal injury caused by bilateral renal ischaemia/reperfusion. PMID- 15288589 TI - Time-dependent reduction of acetylcholine-induced relaxation in corpus cavernosum of cholestatic rats: role of nitric oxide and cyclooxygenase pathway. AB - The endothelium-dependent relaxation of corpus cavernosum smooth muscle and the roles of nitric oxide (NO) and arachidonic acid products of cyclooxygenase were investigated in non-operated, SHAM-operated, and bile duct-ligated rats. We further investigated the time-dependent alterations of corpus cavernosum relaxation in 2-, 7-, and 14-day bile duct-ligated animals. Acetylcholine produced concentration-dependent relaxation in phenylephrine-precontracted strips of corpus cavernosum. A significant reduction in the acetylcholine-induced relaxation was observed 2 days after bile duct ligation, and a greater reduction was observed on subsequent days. Incubation with 20 microM indomethacin reduced the acetylcholine-induced relaxation of the corpus cavernosum of unoperated rats while it had no effect in the corpus cavernosum of bile duct-ligated rats. Chronic treatment with Nomega-Nitro-L-Arginine Methyl Ester (L-NAME, 3 mg/kg/day, intraperitoneally) reduced the relaxation responses in the unoperated group while it had no effect in the bile duct-ligated group. These results show that acetylcholine-induced corporal relaxation is impaired in cholestatic rats, and this may be related to deficient nitric oxide production by the endothelium. The involvement of prostaglandins in this impairment seems unlikely. PMID- 15288590 TI - In vitro antibacterial and anti-inflammatory effects of honokiol and magnolol against Propionibacterium sp. AB - Honokiol and magnolol, two major phenolic constituents of Magnolia sp., have been known to exhibit antibacterial activities. However, until now, their antibacterial activity against Propionibacterium sp. has not been reported. To this end, the antibacterial activities of honokiol and magnolol were detected using the disk diffusion method and a two-fold serial dilution assay. Honokiol and magnolol showed strong antibacterial activities against both Propionibacterium acnes and Propionibacterium granulosum, which are acne-causing bacteria. The minimum inhibitory concentrations (MIC) of honokiol and magnolol was 3-4 microg/ml (11.3-15 microM) and 9 microg/ml (33.8 microM), respectively. In addition, the killing curve analysis showed that magnolol and honokiol killed P. acnes rapidly, with 10(5) organisms/ml eliminated within 10 min of treatment with either 45 microg (169.2 microM) of magnolol or 20 microg (75.2 microM) of honokiol per ml. The cytotoxic effect of honokiol and magnolol was determined by a colorimetric (3-(4,5-dimetyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide) (MTT) assay using two animal cell lines, human normal fibroblasts and HaCaT. In this experiment, magnolol exhibited lower cytotoxic effects than honokiol at the same concentration, but they showed similar cytotoxicity when triclosan was employed as an acne-mitigating agent. In addition, they reduced secretion of interleukin-8 and tumor necrosis factor alpha (TNF-alpha) induced by P. acnes in THP-1 cells indicating the anti-inflammatory effects of them. When applied topically, neither phenolic compound induced any adverse reactions in a human skin primary irritation test. Therefore, based on these results, we suggest the possibility that magnolol and honokiol may be considered as attractive acne mitigating candidates for topical application. PMID- 15288591 TI - Tumour necrosis factor-alpha mediates neutrophil migration to the knee synovial cavity during immune inflammation. AB - Tumour necrosis factor (TNF)-alpha, interleukin-1beta, interleukin-8 and leukotriene B4 have an important role on neutrophil recruitment during immune inflammation. Here we evaluated the participation of several inflammatory mediators on ovalbumin-induced neutrophil recruitment in the knee articular space of immunized rats. Ovalbumin administration in immunized, but not in control, rats induced a dose- and time-dependent neutrophil accumulation, which was inhibited by dexamethasone, pentoxifylline or thalidomide, but not by selective inhibitors of nitric oxide (nitro-L-arginine), platelet-activating factor (BN50730 or UK74505), prostaglandins (indomethacin), histamine (meclisine) or leukotriene B4 (MK 886 and CP105,696). Anti-TNF-alpha antiserum, but not anti interleukin-1beta or anti-CINC-1 (cytokine-induced neutrophil chemoattractant 1) antisera, impaired ovalbumin-induced neutrophil accumulation. High amounts of TNF alpha were detected in the exudates, which was inhibited by dexamethasone, pentoxifylline and thalidomide. These results suggest a specific role for TNF alpha in this model, and the ability of pentoxifylline and thalidomide to inhibit both neutrophil influx and TNF-alpha release may have therapeutic implications in arthritis. PMID- 15288592 TI - Treatment of type IIb familial combined hyperlipidemia with the combination pravastatin-piperazine sultosilate. AB - The risk of coronary heart disease is increased for any given low-density lipoprotein (LDL) cholesterol level in patients with high levels of triglycerides because some triglyceride-rich lipoproteins are atherogenic. This paper reports the results of a pilot clinical trial aimed to evaluate a novel triglyceride lowering drug in combination with pravastatin to treat combined hyperlipidemia. Twenty-six patients with type 2b hyperlipoproteinemia were randomized to receive pravastatin 40 mg/day or pravastatin 40 mg/day plus piperazine-sultosilate 1000 mg/day for 12 weeks if their cholesterol levels, but not triglyceride levels, had responded to therapeutic lifestyle changes and treatment with 40 mg/day of pravastatin. Concentrations of triglycerides, cholesterol and apolipoproteins A and B were measured in duplicate before and after the intervention. There were no significant differences between groups in the change from baseline in the concentration of serum triglycerides. Conversely, significant differences were found for LDL cholesterol, which increased slightly with pravastatin alone but decreased with the combination (12.605+/-22.777% vs. -6.396+/-13.157%, respectively; p=0.022). Apolipoprotein-B levels increased with pravastatin alone but remained stable with the combined treatment (10.464+/-8.446% vs. 0.767+/ 12.335%; P=0.028). The increase in the pravastatin group was significant. Although sultosilate was not efficacious in reducing triglycerides, it helped to decrease the concentration of small, dense, atherogenic LDL particles that are less receptor-sensitive and which could accumulate during long-term statin therapy in patients with high levels of triglycerides. PMID- 15288593 TI - GLUT1-deficient mice exhibit impaired endothelium-dependent vascular relaxation. AB - We tested the hypothesis that decreased glucose transporter 1 (GLUT1) expression alters endothelial function. Nitric oxide-dependent endothelial relaxation, but not endothelium-independent relaxation, was significantly reduced in aortas from transgenic mice expressing GLUT1 antisense mRNA, compared to aortas from nontransgenic littermates. These data suggest that GLUT1-dependent glucose metabolism may play an important role in regulating endothelial function. PMID- 15288595 TI - Metabolic adaptation to hypoxia: cost and benefit of being small. AB - Following metabolic size allometry, the specific metabolic rate of mammals increases with decreasing body mass, resulting in a steeper metabolic fall-off and a faster exhaustion of energy reserves under hypoxic conditions. However, both mammalian hibernators and fetuses are able to temporarily "switch-off" Kleiber's rule as an adaptation to limited food or oxygen supply. Further exceptions to the usual metabolic size relationship are observed in newborn mammals. For instance, neonatal mouse hearts exhibit slower calorimetric "dying curves" under conditions of ischemia, although their aerobic tissue metabolic rates are higher than in adult samples. This is apparently due to a transient reduction of metabolic rate back to the former feto-maternal level. A continuing deviation from metabolic size allometry is found in newborn marsupials (Monodelphis domestica) where the "inappropriately" low specific metabolic rate is a precondition of efficient growth and tissue aerobiosis in spite of extreme immaturity. Obviously, adaptive suppression of elevated metabolism in organisms of small size results in a dramatic improvement of oxygen supply. Vice-versa, the overall increase in specific metabolic rate with decreasing body size might be regarded as one of several phylogenetic adaptations to protect tissues from hyperoxygenation. PMID- 15288596 TI - Hypoxic tolerance in air-breathing invertebrates. AB - Terrestrial invertebrates experience hypoxia in many habitats and under a variety of physiological conditions. Some groups (at least insects) are much more capable of recovery from anoxia than most vertebrates, but there is still a tremendous unexplained variation in hypoxia/anoxia tolerance among terrestrial invertebrates. Crustaceans and arachnids may be less often confronted with hypoxic environments than insects and myriapods and also seem to be less hypoxia/anoxia tolerant. Tracheated groups, especially those that are able to ventilate their tracheal system like many insects, cope with lower critical PO2 than nontracheated groups. Modulation of oxygen carrier proteins is normally not important in hypoxia resistance. Recent application of genetic and cellular tools are revealing that many of the same pathways documented for mammals (e.g. HIF, nitric oxide) function to regulate morphological and biochemical responses to hypoxia/anoxia in insects. PMID- 15288597 TI - Oxygen limited thermal tolerance in fish?--Answers obtained by nuclear magnetic resonance techniques. AB - In various phyla of marine invertebrates limited capacities of both ventilatory and circulatory performance were found to set the borders of the thermal tolerance window with limitations in aerobic scope and onset of hypoxia as a first line of sensitivity to both cold and warm temperature extremes. The hypothesis of oxygen limited thermal tolerance has recently been investigated in fish using a combination of non-invasive nuclear magnetic resonance (NMR) methodology with invasive techniques. In contrast to observations in marine invertebrates arterial oxygen tensions in fish were independent of temperature, while venous oxygen tensions displayed a thermal optimum. As the fish heart relies on venous oxygen supply, limited cardio-circulatory capacity is concluded to set the first level of thermal intolerance in fish. Nonetheless, maximized ventilatory capacity is seen to support circulation in maintaining the width of thermal tolerance windows. The interdependent setting of low and high tolerance limits is interpreted to result from trade-offs between optimized tissue functional capacity and baseline oxygen demand and energy turnover co-determined by the adjustment of mitochondrial densities and functional properties to a species-specific temperature range. At temperature extremes, systemic hypoxia will elicit metabolic depression, thereby widening the thermal window transiently sustained especially in those species preadapted to hypoxic environments. PMID- 15288598 TI - Hypometabolism in reptiles: behavioural and physiological mechanisms that reduce aerobic demands. AB - During exposure to hypoxia all vertebrates utilize a suite of cardiovascular and ventilatory responses that, in combination, strive to maintain adequate delivery of oxygen to the metabolizing tissues. In addition to maintaining oxygen delivery through cardio-respiratory responses, oxygen demands in the tissues can also be reduced. Reptiles use this alternative strategy during periods of moderate to severe hypoxia by behavioural reductions in preferred body temperature and by active down-regulation of aerobic metabolism. Below we review these two different strategies and discuss their possible mechanisms and physiological significance. PMID- 15288599 TI - Acid-base balance during hypoxic hypometabolism: selected vertebrate strategies. AB - An important functional advantage of hypoxic hypometabolism is that it blunts the acid-base consequences of hypoxia. Hypoxia can lead to anaerobiosis and metabolic acidosis and, in animals that are apneic, to respiratory acidosis. A fall in blood and tissue pH is a major limiting factor in hypoxic tolerance and a variety of strategies occur in vertebrates, in concert with hypometabolism, to respond to this acid-base challenge. These include sequestering of lactic acid away from the circulating blood during the hypoxic exposure, either in underperfused tissues or in mineralized tissues, supplementing extracellular buffering by releasing bone mineral into the circulation, and utilizing alternative metabolic pathways for anaerobiosis to produce ethanol rather than lactate as the principal end-product. For submerged air-breathing ectotherms, effective cutaneous O2 and CO2 exchange can also allow an animal to avoid or minimize both anaerobiosis and respiratory acidosis. These responses serve to maintain a viable acid-base state in the body and to extend the time that the hypoxic stress can be endured. PMID- 15288600 TI - Vertebrate brains at the pilot light. AB - While the brains of most vertebrates are unable to tolerate more than a few minutes of anoxia, some freshwater turtles (Trachemys and Chrysemys), crucian carp (Carassius carassius) and frogs (Rana pipens and Rana temporaria) can survive anoxia for hours to months. Obviously, anoxia tolerance has evolved separately several times and this is also reflected in the divergent strategies these animals utilize to survive without oxygen. The turtles and crucian carp defend their brain ATP levels and avoid a loss of ion homeostasis by reducing ATP use. In the turtles, the early release of adenosine and the activation of K(ATP) channels, a progressive release of GABA and a drastic reduction in electric activity and ion fluxes send the brain into a comatose like state. The crucian carp displays a more modest depression of ATP use, probably achieved through a moderated release of GABA and adenosine, allowing the animal to maintain physical activity in anoxia. The anoxic frog, on the other hand, seems to rely on mechanisms that greatly retard the anoxia induced fall in ATP levels and loss of ion homeostasis, so that the brain can be saved as long as the anoxia is limited to a few hours. The sequence of events characterizing the anoxic frog brain is similar to that of failing anoxic mammalian brain, although over a greatly extended time frame, allowing the frog to die slowly in anoxia, rather than survive. By contrast the only factor that limits anoxic survival in turtles and crucian carp may be the final depletion of their glycogen reserves. PMID- 15288601 TI - Metabolic regulation in diving birds and mammals. AB - Ducks, fur seals, Weddell seals and probably most cetaceans seem to be able to dive and remain aerobic for durations that are consistent with their elevated stores of usable oxygen and their metabolic rate while diving being similar to that when they are resting at the surface of the water. Ducks, in fact, have a high metabolic rate while diving, mainly because of their large positive buoyancy, but other species have relatively low buoyancy, are better streamlined and use lift-based rather than drag-based propulsion. However, species such as the larger penguins, grey seals and elephant seals seem to achieve the impossible by performing a substantial proportion of their dives for periods longer than would be expected on the above assumptions, and yet remaining aerobic. The logical conclusion is that during such dives these species reduce their metabolic rate below the resting level (hypometabolism) and, in some of them, there is a regional reduction in body temperature (hypothermia) which may contribute to the reduction in metabolic rate. PMID- 15288602 TI - Natural hypometabolism during hibernation and daily torpor in mammals. AB - Daily torpor and hibernation are the most powerful measures of endotherms to reduce their energy expenditure. During entrance into these torpid states metabolic rate is suppressed to a fraction of euthermic metabolism, paralleled by reductions in ventilation and heart rate. Body temperature gradually decreases towards the level of ambient temperature. In deep torpor body temperature as well as metabolic rate are controlled at a hypothermic and hypometabolic level. Torpid states are terminated by an arousal where metabolic rate spontaneously returns to normal levels again and euthermic body temperature is established by a burst of heat production. In recent years some of the cellular mechanisms which contribute to hypometabolism have been disclosed. Transcription, translation, as well as protein synthesis are largely suppressed. Cell proliferation in highly proliferating epithelia like the intestine is suspended. ATP production from glucose is reduced and lipids serve as the major substrate for remaining energy requirements. All these changes are rapidly reverted to normometabolism during arousal. Hibernation and daily torpor are found in small mammals inhabiting temperate as well as tropical climates. It indicates that this behaviour is not primarily aimed for cold defense, instead points to a general role of hypometabolism, as a measure to cope with a timely limited or seasonal bottleneck of energy supply. PMID- 15288603 TI - Avian embryos in hypoxic environments. AB - Avian embryos at high altitude do not benefit of the maternal protection against hypoxia as in mammals. Nevertheless, avian embryos are known to hatch successfully at altitudes between 4,000 and 6,500 m. This review considers some of the processes that bring about the outstanding modifications in the pressure differences between the environment and mitochondria of avian embryos in hypoxic environments. Among species, some maintain normal levels of oxygen consumption ( VO2) have a high oxygen carrying capacity, lower the air cell-arterial pressure difference ( PAO2 - PaO2 ) with a constant pH. Other species decrease VO2, increase only slightly the oxygen carrying capacity, have a higher PAO2 - PaO2 difference than sea-level embryos and lower the PCO2 and pH. High altitude embryos, and those exposed to hypoxia have an accelerated decline of erythrocyte ATP levels during development and an earlier stimulation of 2,3-BPG synthesis. A higher Bohr effect may ensure high tissue PO2 in the presence of the high affinity hemoglobin. Independently of the strategy used, they serve together to promote suitable rates of development and successful hatching of high altitude birds in hypoxic environments. PMID- 15288604 TI - Implications of hypoxic hypometabolism during mammalian ontogenesis. AB - During hypoxia, many newborn mammals, including the human infant, decrease metabolic rate, therefore adopting a strategy common to many living creatures of all classes, but usually not adopted by adult humans and other large mammals. In acute hypoxic conditions, hypometabolism largely consists in actively dropping mechanisms of thermoregulation. One implication is a decrease in body temperature. This is a safety mechanism, which favours hypoxic survival. Indeed, artificial warming during hypoxia can be counterproductive. Because carbon dioxide is an important stimulus for pulmonary ventilation, the drop in its metabolic production may tilt the balance of ventilatory control in favor of respiratory inhibition. Some experimental data support this view. In conditions of sustained hypoxia, the newborn's hypometabolism also results from a depression of tissue growth and differentiation. Some organs are affected more than others. To what extent the blunted organ growth will be compatible with survival depends not only on the severity and duration of hypoxia, but also on the timing of its occurrence during development. Upon termination of hypoxia, the newborn's metabolic rate recovers and growth resumes at higher rate. Even if body weight may be completely regained, alterations in the respiratory mechanical properties and in aspects of ventilatory control can persist into adulthood, a phenomenon not seen when the hypoxia was experienced at later stages of development. Some of the long-term respiratory effects of neonatal hypoxia are reminiscent of those observed in adult animals and humans native and living in high altitude regions. PMID- 15288605 TI - Detection and quantitation of parvovirus B19. PMID- 15288606 TI - Evaluation of the Roche LightCycler parvovirus B19 quantification kit for the diagnosis of parvovirus B19 infections. AB - BACKGROUND: The rapid and quantitative detection of viral DNA is important in the diagnosis of parvovirus B19 infection in immunocompromised patients and in congenital infection. It is also valuable for monitoring progress following therapeutic interventions. OBJECTIVES: To evaluate the diagnostic sensitivity and specificity of the Roche LightCycler (LC) parvovirus B19 quantification kit in comparison with previously described nested PCR and dot blot hybridisation assays. STUDY DESIGN: Two hundred and twenty eight clinical samples and two standard B19 DNA sera were tested to assess the diagnositic performance of the Roche LC kit. RESULTS: Ten clinical samples (4.3%) gave invalid LC results, including three of five bone marrow samples but only two of 165 serum samples. In the remaining 218 samples, the LC assay detected B19 DNA in 97.5% (79/81) samples that were positive by the nested PCR. The two samples (from the same patient) that were LC negative were sequenced in a 511-nucleotide region of the NS gene and 42 nucleotide changes were found. The Roche LC assay detected B19 DNA in 9.5% (13/137) samples that were negative by nested PCR. Analysis of the available clinical and serological data associated with these samples suggested that the LC results in the majority of these cases were true positive. In patients with resolving persistent infection, the LC assay remained positive for longer than nested PCR. CONCLUSIONS: The Roche LC assay was more sensitive than the nested PCR used in this study. The additional sensitivity and the quantitative DNA measurements were valuable for monitoring patients with persistent B19 infection. Practical advantages of the LC assay include a short running time and the possibility to automate the assay. The LC assay provides a controlled and standardised method for quantitative detection of viral DNA for the diagnosis and monitoring of parvovirus B19 infections but failed to detect a variant strain. PMID- 15288607 TI - No detection of parvovirus B19 or herpesvirus DNA in giant cell arteritis. AB - BACKGROUND: Compelling arguments exist for a role of infectious agent in giant cell arteritis (GCA). Parvovirus B19 and several herpesviruses have focussed the attention in recent years, but the few studies to date have yielded inconsistent results. OBJECTIVES: To study the relationship between the presence of parvovirus B19 DNA or major known herpesviruses and the histopathological features of GCA. STUDY DESIGN: Between January 1997 and March 2002, 147 consecutive temporal artery biopsies were performed in our center because of a clinical suspicion of GCA. Using polymerase chain reaction (PCR) procedures validated by the World Health Organization and employed routinely by our laboratory, we examined the paraffin-embedded specimens for DNA from parvovirus B19, herpes simplex viruses (HSV) 1 and 2, Epstein-Barr virus (EBV), varicella-zoster virus (VZV), human cytomegalovirus (HCMV), and human herpesvirus 6 (HHV-6). We investigated positive results further with immunohistochemistry studies. RESULTS: Fifty of the 147 temporal artery biopsies (34%) showed histological features of GCA. Three biopsies (2.5%) were initially PCR positive for parvovirus B19. None of the herpesvirus PCR assays were positive. Upon repeat testing by both PCR and immunohistochemistry, none of the three initially positive parvovirus B19 assays were confirmed. The results of both positive and negative control assays in these studies validated these findings. We confirmed the presence of amplifiable DNA in the temporal artery biopsy specimens using PCR primers for beta-globin and indoleamine 2,3-dioxygenase (IDO). CONCLUSIONS: The results of our study do not support a role in the etiopathogenesis of GCA for either parvovirus B19 or any of these six herpesviruses. PMID- 15288608 TI - Long-term parvovirus B19 viraemia associated with pure red cell aplasia after allogeneic bone marrow transplantation. AB - BACKGROUND: Parvovirus B19 infection is associated with a variety of symptoms like erythema infectiosum, anaemia and arthritis. In immunocompetent persons, viraemia is usually cleared a few weeks after infection. OBJECTIVE: An immunocompromised adult female patient was persistently infected with B19 after allogenic bone marrow transplantation (BMT) and developed chronic anaemia. STUDY DESIGN: B19-specific antibodies were determined by ELISA and viral load was assessed using a quantitative real time B19 PCR. The patient was evaluated clinically. RESULTS: Two years after successful BMT, the patient received intensified immunosuppressive treatment, erythropoetin and erythrocyte concentrates due to chronic graft-versus-host disease with renal failure. Despite of this treatment, the aplastic anaemia worsened. PCR revealed B19 viraemia with 10(12) geq/ml serum. After 7 months of repeated applications of immunoglobulins and reduction of immunosuppressive treatment, reticulocyte counts and haemoglobin levels normalized and the viral load finally dropped to 10(3) geq/ml serum. One of the back-up samples of the erythrocyte concentrates tested positive, the respective transfusion had been applied 2 months after the beginning of viraemia. CONCLUSIONS: The source of the primary infection remained unclear, but at least re-infection by blood transfusion is likely. Treatment did not result in virus elimination from peripheral blood but in resolvement of symptoms. PMID- 15288609 TI - Prevalence of human parvovirus B19 DNA in cardiac tissues of patients with congenital heart diseases indicated by nested PCR and in situ hybridization. AB - BACKGROUND: Infection with parvovirus B19 (B19) was reported to be correlated with myocarditis, cardiomyopathy, and kawasaki disease. But no information is available about the relationship between inutero B19 infection and congenital heart disease (CHD). OBJECTIVE: To explore whether there is relationship between B19 infection and CHD. STUDY DESIGN: Retrospective investigation of biopsy samples from CHD patients from January 1996 to December 1998. METHODS: Parvovirus B19 was detected in biopsy samples from 42 cases of CHD patients and 38 cases of biopsy or autopsy samples from patients with other diseases (controls) by nested polymerase chain reaction (PCR) and in situ hybridization (ISH) technique. HE staining was also performed to observe the morphology of these cardiac tissue samples. RESULTS: Nested PCR assay indicated that seven of 42 (16.7%) CHD cardiac tissue were B19 DNA positive, while all the 38 controls were B19 DNA negative, the difference is significant (P = 0.012). ISH assay indicated that five of 42 (11.7%) CHD cardiac tissues were positive for B19 DNA and none of the control cardiac tissue were positive, all B19 DNA positive signals were located in the nucleuses of cardiac cells. HE staining showed that there was no inflammatory change in B19 DNA positive cardiac tissue. CONCLUSIONS: Parvovirus B19 DNA was presented in part of CHD cardiac tissues and located in nucleus, which suggested that inutero B19 infection might be correlated with CHD. PMID- 15288610 TI - Replicative multivirus infection with cytomegalovirus, herpes simplex virus 1, and parvovirus B19, and latent Epstein-Barr virus infection in the synovial tissue of a psoriatic arthritis patient. AB - BACKGROUND: Psoriatic arthropathy occurs as complicating feature in about 5-7% of psoriasis patients. Infectious mechanisms including viral antigens have been suggested by serologic data as CD8 T cellular specifity towards viral epitopes. OBJECTIVE AND RESULTS: We here reported a case of a 32-year-old male psoriatic arthritis patient, where we could demonstrate simultaneous infection with cytomegalovirus (CMV), herpes simplex virus type I (HSV1) and parvovirus B19 (B19), as well as latent Epstein-Barr virus (EBV) infection within the synovial tissue by immunohistochemistry (CMV, parvovirus B19, HSV1, EBV-LMP) and DNA-in situ-hybridization (CMV). Serologic examination revealed positive EBV and parvovirus B19-IgG-antibodies, but no antibody response to HSV1 and CMV. CONCLUSION: This case is of special interest, since replicative viral infections have not yet been demonstrated localised in the psoriatic arthritis synovia. Thus, with particular regard to the limited information of the serologic data and the possible need of immuno suppressive therapy direct synovial testing for viral antigenes may be considered in psoriatic arthritis patients. PMID- 15288611 TI - Analysing myocardial tissue from explanted hearts of heart transplant recipients and multi-organ donors for the presence of parvovirus B19 DNA. AB - BACKGROUND: Parvovirus B19 (PVB19) is an erythrovirus causing diverse clinical manifestations ranging from asymptomatic or mild to more severe outcomes, dependent on the haematological and/or immunological status of the host. Reports of PVB19 infection as a causative agent of paediatric or adult inflammatory cardiac diseases, or of cardiac transplant rejection are rare. OBJECTIVES: To identify PVB19 and other cardiotropic viruses in the myocardium of heart transplant (HTx) recipients and multi-organ donors (MOD). Furthermore, to assess the prevalence of cardiotropic viral infection in inflammatory heart disease. STUDY DESIGN: Heart tissue samples from 110 explants were analysed for PVB19 using primers and a 5'-nuclease probe designed to amplify a 160-basepair PCR product from the VP1/NS1 gene region. Samples tested included those obtained from patients undergoing HTx or from MODs. The findings were correlated with clinical course, histologic analysis and serologic testing. Confirmation of the positive PCR-results was done by sequencing and in situ hybridisation. RESULTS: The new assay described here allows precise quantitation of viral load over 7 orders of magnitude (10(6) to 10(0)IU/assay). Measurable amounts of parvoviral genomes were detected in 4/56 (7%) explanted HTx-hearts and in 5/54 (9%) explanted MOD-hearts. CONCLUSIONS: The newly developed real-time PCR is a rapid, sensitive and specific method to detect PVB19 infection in heart tissue. It will be a useful tool to address important questions regarding viral infections transmitted by transplantation, acute infections, relapses and complications involving late or chronic rejection. PMID- 15288612 TI - Virological surveillance of influenza-like illness in the community using PCR and serology. AB - BACKGROUND: Surveillance of winter respiratory viral illness has been carried out for nearly 30 years using a clinical diagnosis by general practitioners as part of the Scottish Sentinel General Practice (SSGP) network. Contemparaneous laboratory diagnosis has not been available previously. OBJECTIVES: To assess the proportion of influenza-like illness (ILI) attributable to influenza, respiratory syncytial virus (RSV) and picornavirus infection during the winter season. To compare the influenza PCR data with serology of paired blood samples. STUDY DESIGN: Combined nose and throat swabs, from patients with ILI attending 15 general practices across Scotland, were submitted to the laboratory in virus PCR sample solution (VPSS). The extracted nucleic acid was tested using a multiplex reverse-transcription polymerase chain reaction (RT-PCR) assay. Serological analysis was performed on paired serum samples using complement fixation assays. The rate of influenza virus positivity was compared with reports of ILI obtained from the SSGP network. RESULTS: Of 240 samples received at the laboratory, 132 (55%) were PCR positive for influenza A virus. There were nine (3.8%) picornavirus and three (1.2%) RSV PCR positives, two (0.8%) were dual influenza A/picornavirus infections. Ninety four (39.2%) were negative for all viruses tested. Results on paired sera from 89 patients showed a rising titre to influenza A in 48 of the 57 PCR positive samples (84.2%). One PCR negative patient displayed a significant rising titre to influenza A. Virological data paralleled the SSGP data but was available at least a week earlier. CONCLUSIONS: Influenza A infection was detected in the majority of patients with ILI; picornavirus infection was also shown to be an important cause of illness. PCR is a rapid and sensitive method for respiratory virus surveillance. Serology is slow, insensitive and difficult to interpret at low titres. PMID- 15288613 TI - Molecular epidemiology of hepatitis B virus infections in Denmark. AB - BACKGROUND: Denmark has a low incidence of acute hepatitis B (HBV) infections but the impact of an increasing number of immigrants with chronic HBV infection on HBV transmission is unknown. OBJECTIVES: To characterise individuals with chronic and acute HBV infection in a defined region and to examine the importance of different risk groups for the current HBV transmission. METHODS: During 2000-2001 all consecutive HBV infected individuals routinely diagnosed through the regional HBV serology laboratory in the County of Funen were classified according to ethnicity, presumed route of transmission and stage of infection based on clinical data mainly supplied by the requesting physician. HBV DNA was sequenced and subjected to phylogenetic analysis. RESULTS: Of 309 identified cases, 91 (29%) were classified as acute infection. HBV DNA sequencing was possible in 54 (59%) of these cases. Phylogenetic analysis showed that HBV isolated from injecting drug users (IDUs) was identical or closely related. Among acute cases acquired in Denmark 89% (74/83) were seen in IDUs (65) or in individuals presumably exposed to IDUs (nine) and phylogenetic analysis corroborated the assumption of IDU related transmission in every case with available sequence data. Among 83 ethnic Danes who acquired their HBV infection in Denmark, no new cases of transmission from immigrants were detected. CONCLUSION: Injecting drug use was the single most important factor for hepatitis B transmission in Denmark. The current Danish vaccination strategy is unable to protect IDUs from HBV infection and IDUs pose a greater risk of HBV transmission to the general population than immigrants. PMID- 15288614 TI - Hepatic failure due to fibrosing cholestatic hepatitis in a patient with pre surface mutant hepatitis B virus and mixed connective tissue disease treated with prednisolone and chloroquine. AB - Fibrosing cholestatic hepatitis (FCH) is a severe variant of hepatitis B infection that has until recently been described almost exclusively in the setting of organ transplantation and HIV infection. This case report describes a patient with pre-surface (pre-S) mutant hepatitis B virus (HBV) infection who developed a fatal form of FCH after high dose prednisolone for mixed connective tissue disease (MCTD). The role of corticosteroids and pre-S viral mutation in the pathogenesis of the disease is discussed, and the importance of early diagnosis is emphasised. This report alerts the physician to the need for close monitoring of LFTs and HBV DNA of hepatitis B carriers during immunosuppressive therapy regardless of the indication. As in the transplantation setting, viral DNA levels should be kept to undetectable if viral replication or recurrence is to be prevented. PMID- 15288615 TI - Quantification of Epstein-Barr virus load in Argentinean transplant recipients using real-time PCR. AB - BACKGROUND: Post-transplant lymphoproliferative disease (PTLD) is a frequent and severe Epstein-Barr virus (EBV)-associated complication in transplant recipients that is caused by suppression of T-cell function. OBJECTIVE: Evaluation of the diagnostic value of EBV DNA load in non-fractionated whole blood samples (n = 297) from 110 pediatric transplant patients by real-time PCR. RESULTS AND CONCLUSIONS: Patients with PTLD had a median viral load of 1.08 x 10(5) copies/ml blood (n = 24), which was significantly higher compared with patients without PTLD (median: 50 copies/ml blood, n = 273, P < 0.0001). From receiver operating characteristic (ROC) curve analysis we obtained a cut-off value of 6215 copies/ml blood with a sensitivity of 95.8%, specificity of 71.4%, negative predictive value (NPV) of 99.5% and positive predictive value (PPV) of 22.8%. Thus, real time PCR proved to be more useful in ruling out than in indicating the presence of PTLD. Further analysis showed that patients without PTLD but developing a post transplant EBV-primary infection had associated high viral loads that were indistinguishable from those of the PTLD group (statistically not significant). Similarly, the presence of clinical symptoms of disease in patients without PTLD was associated with higher viral loads than in patients that were asymptomatic (P < 0.0001), but the difference was much less significant when compared with the PTLD group of patients (P = 0.0391). These patients who had a high viral load may benefit from a close follow-up of the viral burden. PMID- 15288616 TI - Antibody responses against SARS-coronavirus and its nucleocaspid in SARS patients. AB - BACKGROUND: SARS-Cov is the etiologic agent of severe acute respiratory syndrome. An understanding of the antibody responses to the viral components is very important for diagnosis and vaccine development. OBJECTIVE: The spectrum of SARS specific antibody profiles in SARS patients was investigated from 7 to 210 days after the onset of the symptoms. STUDY DESIGN: Serial serum samples from 14 SARS patients were isolated from 7 to 210 days after the onset of the symptoms, and were tested for anti-viral IgG and IgM by indirect immunofluorescence tests (IFA), anti-nucleocaspid antibody by ELISA tests and viral neutralization. RESULTS: Anti-viral (IgG) and anti-nucleocaspid antibodies were observed in 13 of 14 patients at 14 days after the onset of symptoms, and in all 14 patients at 30 210 days thereafter. Anti-viral antibody (IgM) was detected maximally at 30 days, later than that for the IgG class. IgM antibody declined and became undetectable between 60 to 180 days after the onset of the symptoms. Neutralizing viral antibodies were demonstrated in the sera from all of the patients with SARS symptoms. CONCLUSIONS: Anti-viral IgG, IgM, and anti-nucleocaspid antibodies were detected 7-30 days in patients after the onset of SARS symptoms. Anti-viral IgM antibodies disappeared earlier than IgG. Viral neutralization was demonstrated in the sera from the convalescent patients. PMID- 15288617 TI - In vitro susceptibility of 10 clinical isolates of SARS coronavirus to selected antiviral compounds. AB - Effective antiviral agents are urgently needed to combat the possible return of severe acute respiratory syndrome (SARS). Commercial antiviral agents and pure chemical compounds extracted from traditional Chinese medicinal herbs were screened against 10 clinical isolates of SARS coronavirus by neutralisation tests with confirmation by plaque reduction assays. Interferon-beta-1a, leukocytic interferon-alpha, ribavirin, lopinavir, rimantadine, baicalin and glycyrrhizin showed antiviral activity. The two interferons were only active if the cell lines were pre-incubated with the drugs 16 h before viral inoculation. Results were confirmed by plaque reduction assays. Antiviral activity varied with the use of different cell lines. Checkerboard assays for synergy were performed showing combinations of interferon beta-1a or leukocytic interferon-alpha with ribavirin are synergistic. Since the clinical and toxicity profiles of these agents are well known, they should be considered either singly or in combination for prophylaxis or treatment of SARS in randomised placebo controlled trials in future epidemics. PMID- 15288618 TI - A novel pattern (sW195a) in surface gene of HBV DNA due to YSDD (L180M plus M204S) mutation selected during lamivudine therapy and successful treatment with adefovir dipivoxil. PMID- 15288619 TI - Prevalence of infection by HHV-8, HIV, HCV and HBV among pregnant women in Burkina Faso. PMID- 15288620 TI - Animal models in the analysis of Candida host-pathogen interactions. AB - An increasingly diverse array of clinically relevant animal models of candidiasis have been established that mimic both the immune perturbations of the host and tissue-specific features of candidiasis in humans. Cause-and-effect analysis of Candida host-pathogen interactions using these animal models has made a quantum leap forward in the genomic era, with the concurrent construction of C. albicans mutants with targeted mutations of putative virulence factors, the application of microarrays and other emerging technologies to comprehensively assess C. albicans gene expression in vivo, and construction of transgenic and knockout mice to simulate specific host immunodeficiencies. The opportunity to combine these powerful tools will yield an unprecedented wealth of new information on the molecular and cellular pathogenesis of candidiasis. PMID- 15288621 TI - From commensal to pathogen: stage- and tissue-specific gene expression of Candida albicans. AB - Candida albicans is both a successful commensal and pathogen of humans that can infect a broad range of body sites. The transition from commensalism to parasitism requires a susceptible host but it is also an active process. Gene expression of C. albicans is regulated by an interplay between host and pathogen and at least one transcriptional program associated with the yeast-to-hyphal transition. This not only allows immediate adaptation to changing environmental conditions, but also prepares cells for subsequent steps of infection. Recent work using transcript profiling has begun to shed light on infection strategies of pathogenic fungi. PMID- 15288622 TI - RNA interference for analysis of gene function in trypanosomatids. AB - Gene-specific silencing by RNA interference is a valuable tool for analysis of gene function in the protozoan parasite Trypanosoma brucei. The development of tetracycline-regulated vectors for production of double-stranded RNA has facilitated its widespread use. RNA interference provides a fast and efficient method for determining whether a gene is essential for growth and viability, reveals mechanistic information on gene function, and has greatly enhanced our understanding of complex biological processes. Finally, the creation of an RNA interference-based library has allowed, for the first time, an approach for conducting forward genetic experiments in this organism. PMID- 15288623 TI - Antigenic variation in Trypanosoma brucei: facts, challenges and mysteries. AB - Antigenic variation allows African trypanosomes to develop chronic infections in mammalian hosts. This process results from the alternative occurrence of transcriptional switching and DNA recombination targeted to a telomeric locus that contains the gene of the variant antigen and is subjected to mono-allelic expression control. So far, the identification of mechanisms and factors involved still resists technological developments and genome sequencing. PMID- 15288624 TI - Quality of life after repair of ruptured abdominal aortic aneurysm. AB - BACKGROUND: Ruptured abdominal aortic aneurysm (AAA) continues to be associated with high operative mortality. Though survivors can expect to return to a normal life expectancy, their postoperative health related quality of life (HRQoL) remains uncertain. This review examines HRQoL following operative repair of ruptured AAA. METHODS: PreMedline, Medline and Embase databases were searched for clinical studies relating to quality of life following repair of ruptured AAA. Reference lists of relevant papers were also reviewed. RESULTS: Fourteen retrospective-observational studies of postoperative quality of life following repair of ruptured AAA were identified. Both validated and non-validated tools for generic HRQoL assessment were used. All but one study showed no significant difference in overall HRQoL following ruptured AAA repair when compared to both the normal age-adjusted population and patients undergoing elective repair of intact AAA. However, survivors of ruptured AAA did exhibit significant reductions in the isolated domains of physical function, social behaviour and general well being. CONCLUSIONS: There are few studies of HRQoL following repair of ruptured AAA. These reports are retrospective, have small sample sizes and use generic instruments for HRQoL assessment. The findings suggest that survivors of ruptured AAA may attain a similar functional outcome to patients undergoing elective AAA repair and the age-matched healthy population. However, these results must be interpreted with caution and further prospective study is required. PMID- 15288625 TI - Ischaemia-reperfusion injury and regional inflammatory responses in abdominal aortic aneurysm repair. AB - OBJECTIVES: The inflammatory response to abdominal aortic aneurysm repair is likely to result in response to an ischaemia-reperfusion injury (IRI) to the lower-limbs and gastrointestinal tract. This paper reviews the pathogenesis of the inflammatory response to abdominal aortic aneurysm repair, with specific reference to the levels of evidence in the current literature regarding the potential origin of the inflammatory response. DESIGN: Review article. METHODS: The current literature (1966 to August 2003) was reviewed specifically for all articles employing techniques of regional blood sampling from the venous drainage of the lower limbs or gastrointestinal tract during abdominal aortic aneurysm repair. RESULTS: Ten relevant studies were identified. These demonstrated that regional blood sampling techniques could be easily performed, and provided useful information regarding the potential sites of origin of the inflammatory response. CONCLUSIONS: Regional blood sampling techniques provide useful information regarding the potential sites of origin of the inflammatory response. Current evidence suggests that both the lower limbs and gastrointestinal tract are clearly important in their roles, however more work is now required to compare directly the roles and contributions of the lower limbs and gastrointestinal tract to the inflammatory response during abdominal aortic aneurysm repair. PMID- 15288626 TI - Reduction of neoreflux after correctly performed ligation of the saphenofemoral junction. A randomized trial. AB - BACKGROUND: Neoreflux at the sapheno-femoral junction (SFJ) is an important cause of recurrent great saphenous varicose veins. This study compares four surgical methods of ligating the SFJ with the aim to reduce the rate of neoreflux. METHOD: In a prospective study, 379 patients (500 SFJ ligations) were randomised to one of four surgical procedures at the SFJ (125 groins each). In group A (control group) the SFJ was ligated in standard fashion with Vicryl (absorbable ligature); in group B, after Vicryl ligation continuous Prolene (non-absorbable) was sutured over the stump endothelium to prevent any contact with surrounding tissue; in group C. SFJ ligation was done with Ethibond (non-absorbable); in group D Ethibond ligation was followed by Prolene oversewing. The final study group included 114 patients (152 groins) who were all known to be free from recurrent groin reflux 3 months postoperatively and had colour duplex venous imaging 2 years after operation. RESULTS: Duplex imaging identified neoreflux at the SFJ in 10 out of 114 groins after 2 years (7%). There were differences in the rates between the four groups: Group A 3/31 (10%), Group B 0/32, Group C 5/44 (11%) and Group D 2/45 (4%). Neoreflux was significantly reduced in the two groups with endothelial closure (B and D): 2/70 (3%) versus 8/75 (11%, p<0.025). CONCLUSION: Recurrent reflux in the groin was reduced by over sewing the ligated SFJ in patients having varicose vein surgery. This adds weight to the theory of neovascularisation as a cause of recurrent veins and offers a means to reduce clinical recurrence rates. PMID- 15288627 TI - The effect of superficial venous surgery on generic health-related quality of life. AB - BACKGROUND: Superficial venous surgery (SVS) is associated with a significant improvement in disease-specific health related quality of life (HR-QoL) but the effect on generic HR-QoL remains uncertain. The aim of this study was to determine the effect of SVS on responses to the Short Form [SF]-36, the most widely used generic HR-QoL instrument. METHOD: Two hundred and three patients undergoing SVS completed the SF-36 pre-operatively and 24 months post operatively. Scores for the 8 SF-36 domains [physical (PF) and social functioning (SF), role limitation due to physical (RP) and emotional (RE) problems, mental health (MH), vitality (V), pain (P), and general health perception (HP)] were calculated and normalised using UK standard data. RESULTS: Pre-operatively, patients scored significantly lower (worse) than the general UK population in PF, RP and P. Surgery was associated with a significant improvement in PF and P (45.3 vs. 42.5 and 48.9 vs. 43.8 postop vs. preop, p<0.001, WSR) at 2 years. CONCLUSION: SVS leads to a statistically and clinically significant improvement in the physical components of the SF-36. These data will allow the clinical benefits of SVS to be compared with other interventions so helping informing decisions about how venous surgery should be prioritised appropriately within the NHS. PMID- 15288628 TI - Segmental hypoplasia of the great saphenous vein and varicose disease. AB - OBJECTIVES: Primitive narrowing of great saphenous vein segments (saphenous hypoplasia) has been described in healthy limbs. The aim of the present study was to detect great saphenous vein segmental hypoplasia in limbs with varicose veins and to evaluate the local anatomical and haemodynamic patterns. MATERIALS AND METHODS: The incidence of saphenous hypoplasia and the local haemodynamic rearrangement were evaluated by duplex ultrasonography in 676 normal limbs and in 320 limbs with varicose veins. RESULTS: Segmental hypoplasia was demonstrated in 84 normal limbs and in 79 limbs with saphenous reflux. In the latter, the retrograde flow leaves the GSV at the proximal end of the hypoplastic segment to feed tributary veins. CONCLUSIONS: Saphenous hypoplasia occurs in varicose limbs more frequently than in healthy ones (p= >0.001). It greatly influences the path of the reflux and the anatomy of the varicose veins. GSV segmental hypoplasia can be detected preoperatively by duplex ultrasonography. Its occurrence may influence surgical management for two main reasons: in about 68% of varicose limbs with segmental hypoplasia, the distal GSV is competent. If the distal GSV is varicose, its size and flow direction is normalised by treating the accessory vein that bypasses the hypoplastic segment. PMID- 15288629 TI - Quality of life and long-term results after ruptured abdominal aortic aneurysm. AB - OBJECTIVES: Quality of life as an endpoint of surgery and the long-term prognosis for patients who have survived surgery for a ruptured abdominal aortic aneurysm (RAAA) is not well-documented. PATIENTS AND METHODS: The records of all patients from 1993 to 2000 who underwent resection of RAAA were reviewed. Survival data were calculated from direct contact with the patients or follow-up records. All patients who were alive at the time of our study were invited to participate in follow-up investigations. They received the internationally comparable WHO-QOL BREF-test. RESULTS: In a period of 7 years, 80 patients underwent surgery for RAAA. The average follow-up time was 5.1 years (1-7.9 years). Our data show that 51% of our patients died within 6 months postoperatively because of the complications of the aortic rupture (in-hospital mortality 39%). Patients who survived the first 6 months after surgery died for the same reasons as the normal population. However, patients who were younger than 75 at the time of RAAA had a higher relative survival rate than a matched sample of the population. There was no significant difference in the quality of life between the study group and the general population. CONCLUSIONS: RAAA survivors had no difference in long-term survival as compared to the general population and also had very few long-term complications. The WHOQOL-BREF-test suggests that the quality of life of survivors of RAAA is similar to the general population. PMID- 15288630 TI - Late survival after elective repair of aortic aneurysms detected by screening. AB - BACKGROUND: The aim of this study was to examine whether there was any survival advantage in men following elective repair of an abdominal aortic aneurysm (AAA) detected by ultrasound screening compared to those with an AAA detected incidentally. METHODS: A total of 424 men underwent elective AAA repair between 1990 and 1998; 181 were detected in an aneurysm screening programme and 243 were diagnosed incidentally. Follow-up survival data were collected until 2003 (minimum 5 years) and survival curves were compared using regression analysis. RESULTS: The postoperative 30-day mortality rate was significantly lower in men whose aneurysms were detected by screening (4.4%), compared with those detected incidentally (9.0%). Similarly, 5-year survival (78% vs. 65%) and 10-year survival rates (63% vs. 40%) were better after repair of a screen-detected AAA (p<0.0003 at all time intervals, by log rank testing). Multivariate analysis showed that this was largely due to the older age of men who had repair of an incidental AAA (71.2 vs. 67.1 years). CONCLUSION: Men who had elective repair of an AAA detected by screening had a better late survival rate than men whose aneurysm was discovered incidentally because they were younger at the time of surgery. PMID- 15288631 TI - Cytokines, their genetic polymorphisms, and outcome after abdominal aortic aneurysm repair. AB - BACKGROUND: Excessive cytokine production has been implicated in the development of organ failure. Polymorphic sites in cytokine genes have been shown to affect levels of production in vitro and may influence cytokine production in vivo. The aims of this study were to determine if cytokines or their genetic polymorphisms were related to outcome after abdominal aortic aneurysm (AAA) repair. METHODS: A prospective study of 135 patients undergoing open AAA repair. Plasma levels of TNF-alpha, IL-1beta, IL-6 and IL-10 were measured 24 h post-operatively and genotypes for the TNF-alpha -308, IL-1beta+3953, IL-6 -174, IL-10 -1082 and IL-10 -592 polymorphisms were determined for each patient. RESULTS: After elective AAA high levels of IL-10 were associated with both prolonged critical care (P<0.001) and hospital stay (P=0.001). The presence of a G allele at the IL-6 -174 locus was associated with a higher incidence of organ failure (P=0.04) and an A allele at TNF-alpha -308 with prolonged critical care stay (P=0.03). After ruptured AAA the development of multi-organ failure was associated with high levels of IL-6 (P=0.01) and TNF-alpha (P=0.04). High TNF-alpha levels were also associated with mortality (P=0.01). CONCLUSION: Post-operative cytokine levels are related to outcome after AAA repair. Cytokine gene polymorphisms may provide a method for determining which patients are at high risk of complications. PMID- 15288632 TI - Are laparoscopic staplers effective for ligation of large intraabdominal arteries? AB - OBJECTIVES: To evaluate ligation of aortoiliac arteries with laparoscopic staplers in order to develop specifically designed staplers. METHODS: Cadaveric study. Seven human cadaver aortas were stapled using EndoGIA60 staplers. Efficiency was evaluated macroscopically and on a hydrodynamic bench. Clinical study. Twelve patients had ligation of 14 large abdominal arteries (aorta: nine, iliac artery: four, hepatic artery: one) using a laparoscopic stapler. Stapling efficiency was judged on peroperative clinical and postoperative CT scan criteria. RESULTS: Cadaveric study. Stapling was performed perfectly on four moderately calcified aortas, without leakage with a pulsatile pressure of >250 mmHg. For three aortas with severe calcification, stapling was not efficient and major leakage occurred. Clinical study. Stapling appeared clinically efficient on all arteries but one aorta: this severely calcified aorta was ligated conventionally. The staplers are not easy to use due to their shape and their lack of articulation. After a mean follow-up of 31.3 months, all the other stapled arteries were effectively ligated. CONCLUSION: The commercially available staplers can be used securely on moderately calcified arteries but stapling of severely calcified arteries should be avoided. These devices should be redesigned to facilitate their use in vascular surgery. PMID- 15288633 TI - Optimal follow-up strategies after aorto-iliac prosthetic reconstruction: a decision analysis and cost-effectiveness analysis. AB - OBJECTIVE: The primary aim of ultrasound follow-up after aorto-iliac prosthetic reconstruction is to correct false aneurysms before rupture occurs. We investigated whether follow-up improves the life expectancy of patients and sought to identify the most cost-effective follow-up strategy. DESIGN OF THE STUDY: A Monte Carlo Markov decision model was constructed. The occurrence of false aneurysms was modelled as a time-dependent process for each anastomotic site, based on published series. Using this model, the impact of various follow up strategies was investigated for three types of prostheses, aorto-distal tube, aorto-bi-iliac, and aorto-bi-femoral prostheses. Main outcome measures were discounted quality adjusted life years (dQALYs), discounted costs, and (discounted) cost-effectiveness (CE) ratios. RESULTS: Follow-up of patients with aorto-distal tube and aorto-bifemoral prostheses did not result in an improvement life expectancy and was not cost-effective, QALYs 7.53 and 7.62 years, respectively. The results for aorto-distal tube and aorto-bifemoral prostheses were not sensitive to any variation in the model parameters. In the base case analysis, the life expectancy of patients with aorto-bi-iliac prostheses was 7.50 QALYs (95% confidence interval 7.46-7.54) whether or not they underwent routine follow-up. However, patients aged 54 years or younger gained 0.11 QALYs with annual follow-up (p<0.05). The most cost-effective strategy was annual follow-up that starts 10 years after the initial operation, and continues up to 30 years after surgery (4600 Euro; CE ratio 21,000 Euro per QALY). When perioperative mortality of elective reconstruction of false aneurysms is 2% or lower (e.g. when endovascular treatment is used), a small improvement is observed (7.56 vs. 7.50 QALYs; p<0.05; CE ratio 35,000 Euro per QALY). CONCLUSIONS: Annual follow-up of aorto-bi-iliac prostheses should be restricted to patients aged 54 or younger and not start before 10 years after surgery. The same strategy can only be considered for older patients if mortality for secondary intervention is lower than 2%. Since patients with aorto-distal tube and aorto-bi-femoral prostheses do not benefit from follow-up for the detection of false aneurysms, this practice should be discouraged in these patient groups. PMID- 15288634 TI - Use of a fibrin preparation in the engineering of a vascular graft model. AB - OBJECTIVE: Morphological and functional characterization of cocultured endothelial cells (EC) and myofibroblasts (MFB) seeded on a matrix composed of a fibrin preparation mimicking the microenvironment of a vascular wall. METHODS: MFB and EC were isolated from human saphenous veins and expanded separately in vitro. MFB were seeded on a composite matrix consisting of a fibrin preparation (with or without transforming growth factor-beta2) and a polyglactin-mesh to form a 3-dimensional structure, which was consecutively reseeded with EC. Seeded matrices were incubated in a bioreactor. Characterization was done including fluorescence staining, live-/dead-assay and immunohistochemistry. RESULTS: High density cocultures in hierarchical structure mimicking the formation of a vascular wall were obtained with nearly complete coverage of the surface with EC. Distribution of preseeded MFB in a 519+/-27 microm thick layer (day 14) was achieved. Cell viability was shown in fluorescence staining for at least 19 days. In deeper layers, no viable cells could be detected within the fibrin preparation. EC covered the surface, had uniform morphology, and their preserved viability was shown for at least 5 days. No EC-ingrowth was found into the fibrin preparation. Neoformation of the matrix proteins laminin and collagen IV was observed. CONCLUSION: A structured coculture of MFB and EC was obtained mimicking the formation of a vascular wall with preserved viability utilizing a fibrin preparation. Nutrition problems seem to limit the maximal extent of MFB in the matrix. PMID- 15288635 TI - Speed rather than distance: a novel graded treadmill test to assess claudication. AB - OBJECTIVE: To evaluate a new treadmill test, determining pain threshold speed (PTS) for use in assessment and measuring rehabilitation of patients with intermittent claudication. METHODS AND DESIGN: Twenty-nine patients with claudication were evaluated, and the ankle-brachial index (ABI) was assessed. PTS was determined with a treadmill protocol based on level walking, low starting speed, and progressive increments at a predetermined distance up to the onset of pain. Repeatability and equivalence with a time-based protocol were verified. PTS was compared to pain-free walking distance, 6-minute walking distance, and ABI. RESULTS: PTS was measured in all patients (3.6+/-1.1 km/h). Repeatability and equivalence between established tests were demonstrated. PTS showed a significant correlation with pain-free walking distance (r=0.833; P=0.0001), with 6-minute walking distance (r=0.724; P=0.005), and with ABI in the more ischemic limb (r=0.641; P=0.0001). CONCLUSIONS: PTS is a reliable parameter that correlates well with other established measures. It is useful for determining the degree of functional handicap and for designing and guiding rehabilitation protocols. PMID- 15288636 TI - Evaluation of flow hemodynamics by color-Doppler following two different brachial arterial repair techniques. AB - OBJECTIVES: We compared the clinical and hemodynamic results following surgical repair of traumatic brachial artery injury using two different techniques micro- and macrovascular repair. MATERIALS AND METHODS: This was a retrospective study of 27 patients who had sustained penetrating, clean cut injuries of the brachial artery. Macrovascular techniques and a saphenous vein graft was used in 13 patients, while 14 patients were treated by primary microsurgical technique. Postoperatively, patients were followed for a mean of 26 months. All patients had color Doppler examination of the brachial artery, digital artery pressures and transcutaneous oxygen saturation determined. RESULTS: Clinical results based on distal pulses, Allens test and digital pressures were similar in the two groups. Color Doppler showed 8/13 anastomotic stenoses in macrovascular vein grafted repairs and 2/14 in microvascular repairs (p<0.05). The ratio of flow velocity proximal compared distal to the injury was significantly decreased in patients who had macrovascular repairs. CONCLUSION: Using ratio between proximal and distal site of anastomosis maximal peak systolic velocity as a objective color Doppler parameter, we were able to demonstrate differences in the hemodynamic status following macrovascular repair with vein grafts and microvascular primary repair. The results emphasize the importance of using a standard repair technique for similar injuries rather than the preference of the surgeon. PMID- 15288637 TI - A new approach for the screening of carotid lesions: a 'fast-track' method with the use of new generation hand-held ultrasound devices. AB - OBJECTIVES: We assessed the usefulness of fast-track neck sonography with a new generation hand-held ultrasound scanner in the detection of > or =60% carotid stenosis. DESIGN: Patients with a past history of atherosclerotic disease or presence of risk factors were enrolled. All had fast-track carotid screening with a hand-held ultrasound scanner. METHODS: Initial assessment was performed with our quick imaging protocol. A second examiner performed a conventional complete carotid duplex as gold-standard. RESULTS: We enrolled 197 consecutive patients with a mean age of 67 years (range 35-94). A carotid stenosis >60% was detected in 13 cases (6%). The sensitivity, specificity, positive and negative predictive value of fast-track sonography was 100%, 64%, 17% and 100%, respectively. Concomitant power Doppler imaging during the fast-track method did not improve accuracy. CONCLUSIONS: The use of a fast-track method with a hand-held ultrasound device can reduce the number of unnecessary carotid Duplex and enhance the screening efficiency without missing significant carotid stenoses. PMID- 15288638 TI - Homocysteine levels, haemostatic risk factors and restenosis after carotid thrombendarterectomy. AB - OBJECTIVES: To investigate the effect of elevated serum homocysteine and haemostatic as well as clinical risk factors on the tendency to restenosis after carotid artery thrombendarterectomy. DESIGN: A prospective, observational study. PATIENTS AND METHODS: In the period from October 1999 to October 2002, 86 patients were subjected to 96 carotid endarterectomies because of internal carotid artery stenoses. The carotid stenoses were symptomatic in 86 cases (90%). Fasting plasma homocysteine, fibrinogen, D-dimer and activated protein C resistance were measured the day before surgery. Follow-up was done 1, 3, 6, 12 and 18 months postoperatively and yearly thereafter with clinical assessment and triplex ultrasonography. The median follow-up time was 17 months (range 9-42 months). Freedom from restenosis was estimated with Kaplan-Meier curves, using log-rank test for comparison between groups. Variables found to be significantly related to restenosis rates were included in a multivariate analysis performed with the Cox proportional hazards model. Comparison of means of continuous data between two groups was done with Student's t-test and more than two groups with one-way analysis of variance. RESULTS: Restenoses within 12 months of the operation occurred in 11 cases (11%). Univariate analysis revealed that plasma homocysteine values < or =10 micromol/l and freedom from ischaemic heart disease were both significantly associated with an increased risk of restenosis (p=0.0076 and 0.0059). However, multivariate analysis showed that only plasma homocysteine values <10 micromol/l were independently and significantly associated with an increased risk of restenosis (p=0.046). There were no associations between the degree of atherosclerotic affection of the precerebral circulation or symptoms on one hand and the levels of homocysteine, fibrinogen, D-dimer and activated protein C resistance on the other. CONCLUSION: There seems to be an independent, significant association between homocysteine values within the lower two thirds of the normal range and restenosis after carotid endarterectomy. Studies on the biological properties of human endothelial cells from different types of vasculature and different locations, specifically with respect to homocysteine metabolism and its effect, are warranted. PMID- 15288639 TI - Acute mesenteric venous thrombosis: a better outcome achieved through improved imaging techniques and a changed policy of clinical management. AB - OBJECTIVE: To analyse and compare the results obtained from acute mesenteric venous thrombosis (MVT) patients before and after the change of the clinical management principle, to assess the factors responsible for the recent better outcome and determine the best management for this disease. MATERIALS AND METHODS: We retrospectively reviewed 41 patients treated for acute MVT admitted in our hospital between 1978 and 2003. Before 1995 (Group I), our policy was to perform surgery in patients with suspected acute MVT. After 1995 (Group II), we changed our policy to a medical approach when achievable. Each patient in this study was assessed for diagnosis, initial management (operative or non operative), mortality, duration of hospitalisation, and outcome. RESULTS: There were 13 in Group I, 28 in Group II. The mean duration of diagnoses made after admission was 7.3 S.D. 2.6 days for patients in Group I, and 1.5 S.D. 1.2 days for those in Group II (p<0.01, Student's t-test). Eleven patients underwent operations and two patients received non-operative treatment initially in group I, the mortality was 39%; while nine patients underwent operations and 19 patients received non-operative management in group II, the mortality was 11% (p<0.05). No death occurred in the patients with initial non-operative management. The mean duration of hospitalisation was 26 S.D. 6.8 days in Group I and 12.6 S.D. 4.6 days in Group II (p<0.01, Student's t-test). No significant difference in 2-year survival rate between the two groups. CONCLUSION: Recent improvements in imaging techniques and better understanding of the aetiology have led to a dramatic change in the principle and policy of clinical management for acute MVT, which leads to a more favourable outcome of acute MVT. PMID- 15288640 TI - Neurovascular compression of the common peroneal nerve by varicose veins. AB - Compression of the common peroneal nerve occurs sometimes, but compression caused by varicose veins has not been reported before. We report a case of common peroneal nerve compression syndrome which was confirmed and treated surgically. A 63-year-old woman complained of paraesthesia on the lateral aspect of the right leg, which was worse in the evening. A primary varicose vein arising from non saphenous tributaries was seen in the posterior calf. Her symptoms resolved with the wearing of compression hosiery for 2 weeks. At operation, the common peroneal nerve was found to be surrounded by tortuous varicosities. After decompression the paraesthesia on the lateral aspect of the right leg resolved completely with no evidence of residual neuralgia. PMID- 15288641 TI - Superior mesenteric arterial blood flow during infrarenal aortic surgery. PMID- 15288642 TI - Electron microscopy studies on DNA recognition by DNA-PK. AB - Advances in transmission electron microscopy coupled to increasingly powerful biocomputing techniques are opening enormous possibilities to understand the structure and function of complex biological processes performed by large multi protein assemblies. This is an exciting time for electron microscopists because we can combine our efforts with X-ray crystallographers and NMR spectroscopists to reach the prospect of studying the structure and dynamics of the so-called 'molecular machines'. One of these fascinating systems is the macromolecular complex formed around double-stranded DNA breaks (DSBs). Non-homologous end joining (NHEJ) is the main DSBs repair pathway in mammalian cells, where a collection of proteins interact to rejoin two broken DNA ends. During NHEJ, DNA dependent protein kinase (DNA-PK) binds damaged DNA with high affinity and acts as the main scaffold for other repair factors. Several studies have made use of the electron microscope to reveal the three-dimensional architecture of DNA-PK and the structural basis for the recognition of damaged DNA and the activation of DNA-PK's kinase activity. PMID- 15288643 TI - Some trends in microscope image processing. AB - The present review tries to identify some trends among the multitude of ways followed by image processing developments in the field of microscopy. Nine topics were selected. They cover the fields of: signal processing, statistical analysis, artificial intelligence, three-dimensional microscopy, multidimensional microscopy, multimodality microscopy, theory, simulation and multidisciplinarity. A specific topic is dedicated to a trend towards semi-automation instead of full automation in image processing. PMID- 15288644 TI - Bizarre alterations of the morphology of pineal synaptic bodies under constant light and an evaluation of suitable 3D-reconstruction software. AB - Three dimensional (3D) reconstruction and modelling software was evaluated to find a procedure suitable for visualization of small subcellular structures in transmission electron microscope images. The method applied in this study demonstrates bizarre alterations of the structure of synaptic bodies (SBs) in pinealocytes of the guinea-pig pineal gland caused by constant illumination. It can, in general, be used for any 3D reconstruction from serial sections. Pineal glands of five guinea-pigs (two kept under a LD cycle of 12:12 h; three kept in constant light, for 4 months) were investigated. SBs consist of an electron-dense centre with attached vesicles. Under normal lighting conditions most SBs are flat plates (about 35 nm in thickness), which eventually may be bent. The proteins comprising the molecular basis of SBs, mainly RIBEYE A and B are polymerised in a regular manner in these plates. This is not the case in other SBs, which appear as spheres or irregular lumps. SBs lie in groups in which usually some of the plates are arranged in parallel arrays Constant illumination caused different changes in morphology: many of the SBs lie in 'paired fields', i.e. appear in groups attached to the cell membranes of two pinealocytes directly opposite to each other. Some of the SBs in such groups are strongly bent, showing blebs and irregular thickened areas, others seem to aggregate and show inclusions of cytoplasm. Further goblet-like, shield-like and other bizarre forms of SBs occurred and the relative number of spheroid and lump-like SBs increased. Protrusions on larger SBs suggest detachment or fusion of SB material to a greater extent than in the control animals. There is a reduction of areas in which the polymerisation of the SB proteins remains well ordered, i.e. where the typical thickness of 35 nm is maintained. It remains unclear why this polymerisation pattern is only partly affected by constant light. PMID- 15288645 TI - Antennal sensilla of some forensically important flies in families Calliphoridae, Sarcophagidae and Muscidae. AB - Antennal sensilla of some forensically important fly species in the families Calliphoridae (Chrysomya megacephala, Chrysomya rufifacies, Chrysomya nigripes and Lucilia cuprina), Sarcophagidae (Parasarcophaga dux) and Muscidae (Musca domestica) were studied using scanning electron microscopy. Five types of sensilla were observed: trichoid, basiconic, coeloconic, styloconic and sensory pit. Only trichoid sensilla are found on the scape of the antenna, while both trichoid and styloconic sensilla are located on the antennal pedicels of all species studied. Basiconic sensilla are the most numerous of the sensilla found on the antennae of both sexes of all fly species studied and are comprised of two subtypes: large and small basiconic sensilla. Coeloconic sensilla are characterized by short pegs, with either grooved or smooth surfaces, that are sunken into deep depressions. No marked difference was observed in the number, morphological structure or distributional pattern of any of the sensilla among the species studied, with the exception of there being more numerous sensory pits detected in female P. dux compared to the other species. The suggested function of each antennal sensillum was based on comparison with results of other investigations on similar sensilla. PMID- 15288646 TI - Imaging of growth banding of minerals using 2-stage sectioning: application to accessory zircon. AB - In the past, investigations of mineral growth were routinely carried out by the application of light-microscopic methods and, later on, by the production of single crystal sections and their documentation using specific imaging techniques (CL, BSEI, etc.). In the present work, a method is described which enables the precise sectioning of elongate crystals parallel and perpendicular to their longest axis. By examining backscattered electron images of parallel and perpendicular sections of the same grain, growth of all faces may be evaluated without major geometric correction. The new technique is applied to zircon crystals of a granitoid exposed in the southwestern Bohemian Batholith, Austria. For the studied zircon population, pyramidal and prism development during crystal growth is worked out very clearly by the imaging procedure. Besides its significance in crystal studies, the introduced method could also find a use in material science for the growth study of synthetic mineral phases. PMID- 15288647 TI - Histochemical and ultrastructural study of collagen fibers in mouse pubic symphysis during late pregnancy. AB - Reference is usually made to the parallel orientation towards the main line of exerted tension at the pubic joint in mice, for supporting forces applied to the joint. Despite the wealth of morphological information about the extracellular matrix in this joint, little is known regarding the involvement of the crimp of collagen fibers in the dramatic transformations occurring in this region during the last 3 days of pregnancy. Examination of the collagenous architecture suggests that the biomechanical properties are directly related to fibril diameters, composition of ground substance and changes in the bundle morphology, particularly in the crimp structure. The purpose of this study was to further describe the transformation of the collagen fibers of the pubic symphysis during late mouse pregnancy. We examined the architecture of collagen fibers in the symphysis and pubic ligament through the Picrosirius-polarization method and also through scanning electron microscopy to directly visualize and measure the crimping from pregnant and virgin mice. The crimp angle and the length of five consecutive crimps were measured according to Patterson-Kane et al. [Connect. Tissue Res. 36 (1997) 253]. It could be demonstrated that the angles progressively decreased and the crimp length increased, denoting that the fibers have untwisted during the relaxation process. Our findings suggest that a disruption of the helical arrangement of the collagen containing fibers may contribute to explaining the rapid remodeling that occurs at the end of pregnancy and that is responsible for an increase in pliancy and length of the pubic ligament in mice. PMID- 15288648 TI - Bulk interfaces in a Ni-rich Ni-Au alloy investigated by high-resolution Z contrast imaging. AB - Interfaces between Au-rich precipitates and the Ni-rich matrix in a decomposed Ni 10 at.% Au alloy were investigated by low-magnification and high-resolution Z contrast imaging. During aging at 923 K, the originally single crystalline sample decomposed and recrystallized resulting in a microstructure consisting of subgrains separated by small-angle grain boundaries. These small-angle grain boundaries are decorated by Au-rich precipitates. The interfaces between the Au rich precipitates and the Ni-rich matrix were characterized with respect to the orientation relationship between precipitates and matrix, misfit dislocations and concentration gradients. Two transformation modes were identified that are involved in the decomposition of bulk Ni-rich Ni-Au alloys. While in the first mode the interface is semi-coherent, in the second mode the interface corresponds to an incoherent twin boundary. It is further shown that strain fields around misfit dislocations can result in systematic errors in the determination of the concentration gradients across interfaces between precipitates and matrix. PMID- 15288649 TI - Collection and X-ray microanalysis of airway surface liquid in the mouse using ion exchange beads. AB - The airway surface liquid (ASL) is a thin layer of liquid covering the airway epithelium. The ionic composition of the ASL is assumed to be important for airway function and may be altered in diseases such as cystic fibrosis and exercise-induced asthma. A method for collection of ASL is presented in which the fluid is collected using small dextran ion-exchange beads. The beads are equilibrated with the ASL in a humidity chamber, collected under silicon oil, dried and analyzed by X-ray microanalysis. Analysis of standard beads prepared by exposure to different salt solutions shows that linear calibration lines can be obtained, but that beads absorb different elements to a different extent. The results show that the ASL in mice is hypotonic, and that the mucus component of the ASL has an elemental composition that is different from that of the periciliary fluid. PMID- 15288650 TI - Ocadaic acid treatment causes tyrosine phosphorylation of caveolin-2 and induces internalization of caveolae in rat peritoneal macrophages. AB - In this paper, we provide evidences that protein phosphatases could regulate the internalization cycle of caveolae in rat peritoneal cells. Ocadaic acid (OA)--a serine/threonine phosphatase inhibitor--was used in various concentrations (4 and 100 nM) to study the internalization of horseradish peroxidase (HRP) in resident and elicited macrophages. We have found that OA in both concentrations has significantly decreased HRP uptake in resident and elicited cells. The results of our morphometrical analysis showed that in OA-treated cells, the number of surface-connected caveolae has been dramatically decreased. Simultaneously large, endosome-like vacuoles containing small vesicles appeared in the cytoplasm. The membrane of these small vesicles was labeled with anti-caveolin-1 antibody. Immunoprecipitation and Western blot analysis revealed that in OA-treated cells an approximately 29 kDa protein identified as caveolin-2 in macrophages was phosphorylated on tyrosine residues. These data support the idea that there is a close correlation between the phosphorylation of caveolin-2 and endocytosis of caveolae: the tyrosine phosphorylation of this approximately 29 kDa protein can drive caveolae to pinch off from the plasma membrane and causes accumulation of caveolae in a multivesicular body-like cellular compartment, which was never found to contain lysosomal enzymes. As a result of OA treatment caveolin-2 remains phosphorylated and the phosphorylation of these protein might inhibit the recycling of caveolae. PMID- 15288651 TI - Karyotype, banding and rDNA FISH in the scarab beetle Anoplotrupes stercorosus (Coleoptera Scarabaeoidea: Geotrupidae). Description and comparative analysis. AB - Six specimens of Anoplotrupes stercorosus (Coleoptera Scarabaeoidea: Geotrupidae) were analysed using conventional staining, banding techniques and fluorescent in situ hybridization with a ribosomal probe (rDNA FISH). Detailed karyotype description was also joined to a comparative analysis between present data and those previously reported for Thorectes intermedius [Chromosome Res. 7 (1999) 1]. The two species, both belonging to the tribe Geotrupini, show the same modal number but different autosomal morphology which is in contrast with the high chromosome stability argued for Geotrupinae during the last three decades. Moreover, a detailed comparison reveals the occurrence of a plesiomorphic condition in A. stercorosus with respect to the apomorphic one of T. intermedius. This finding agrees with phylogenetic relationships proposed for the two genera based on morphological and anatomical characters. PMID- 15288652 TI - Implementation and evaluation of a detector for forward propagated second harmonic signals. AB - Second harmonic generation (SHG) is emerging as an alternative non-linear imaging method. The fact that most commercial multi-photon microscopes can be easily adapted to image SHG makes it appealing to explore the kind of sample information given by SHG. Here we describe an SHG detection implementation designed to optimize the collection of forward propagating light. A Hamamatsu H957-08 PMT is inserted at the back-focal plane of the condenser on an inverted Nikon TE300, and controlled by the existing electronics of a BioRad 1024MP system. Evaluation of the performance was done on common SHG generating preparations, KH2PO4 crystals and collagen. We concluded that positioning a detector at the back focal plane of the condenser provides a highly efficient detection system for second harmonic signals, with many advantages over a detector sited at the lamp housing. PMID- 15288672 TI - Ordering and interpreting diagnostic studies em leader but who is caring for the patient? PMID- 15288673 TI - Fabry's disease: long-term study of a family. AB - Fabry's disease is the second most prevalent lysosomal storage disorder after Gaucher's disease. It occurs as the result of a deficit in the alpha galactosidase A enzyme. The gene coding for it is located on the long arm of the X chromosome (Xq22.1). This deficit causes the gradual accumulation of a glycosphingolipid. The main substance accumulated is globotriaosylceramide (Gb3). This accumulation leads to pain and angiokeratomas, and to cardiac, cerebral, and vascular involvement as the disease progresses. The treatment of Fabry's disease has so far only been symptomatic; however, new advances have now made it possible to prescribe alpha-galactosidase replacement therapy, which not only improves symptoms, but also enhances these patients' quality of life and lowers mortality. In this paper we review the status of Fabry's disease and we report the follow-up of a family with Fabry's disease, with some members receiving replacement therapy with alpha-galactosidase A and demonstrating good progress. PMID- 15288674 TI - Supervised echocardiography in internal medicine. An integrated model in a community hospital. AB - Background: The management of patients admitted to an internal medicine ward frequently requires echocardiography, which may often be delayed because of overburdened specialist cardiologist services. The availability of appropriate echocardiography may be improved if internists first perform autonomous echocardiography on their cardiac patients. Our 5-year experience with such a model shows how it can exploit the complementary role of internists and cardiologists. Methods: We analysed data collected prospectively over 5 years, including patient characteristics, indications for investigation, time of execution, echocardiographic findings, incidence of technical failures and incomplete reports, and need for expert consultation, supervision, and review. Results: Out of 6035 admitted patients, 1943 (32%) had a primary cardiac discharge diagnosis and 1158 (19%) underwent transthoracic echocardiography (54% male, mean age 70.2+/-10.3 years). Heart failure, atrial fibrillation/flutter, and chest pain were the most frequent indications (19%, 14%, and 12% of cases, respectively). Technical failure occurred in 31 cases (2.7%) and incomplete information was provided in 127 cases (11%). Valvular and coronary heart diseases were the most frequent echocardiographic diagnoses (27% and 15%, respectively). Expert supervision, consultation, or review was required in 21 of the examinations (1.8%). Conclusion: Internists with training in echocardiography and adequate access to expert consultation can provide timely and clinically profitable echocardiographic information for the majority of their cardiac patients. The improved selection of their referrals allows cardiologists more time to devote themselves to detecting expertise-demanding pathology using special echocardiographic procedures. This integrated model can be applied in various clinical settings. PMID- 15288675 TI - Is cardiology consultation required before cardiac catheterization? AB - Background: In many hospitals, internists have begun to directly refer patients for cardiac catheterization without a prior cardiology consultation. The purpose of this study is to compare the results of a policy of mandatory consultation prior to catheterization with one of optional consultation. Methods: One hundred seventy-five consecutive patients who underwent catheterization with a prior cardiology consultation (closed group) were compared to 175 patients who underwent the procedure without a prior mandatory consultation (open group). The primary outcomes were defined as whether significant coronary disease was found and what therapy the patient received. Results: There was no difference in the percentage of patients who were found to have coronary disease in each group (72% in the closed group and 77% in the open group, P=NS), and there was also no difference in the percentage of patients who received revascularization therapy (43% in the closed group and 44% in the open group, P=NS). Conclusions: Allowing internists to refer patients directly for catheterization resulted in equivalent results as compared to requiring cardiology consultation. This study supports the policy of allowing direct referral for catheterization, but further studies are needed to compare the outcomes of cardiac patients cared for by hospitalists without cardiology consultation. PMID- 15288676 TI - Comparison of the effects of amlodipine and verapamil on autonomic activity in hypertensive patients. AB - Background: Many studies have shown that autonomic activation is one of the major factors in the etiology of hypertension. Furthermore, sympathovagal imbalance may be responsible for arrhythmias and sudden cardiac death. The aim of the present study was to compare and to evaluate the effects of short-term therapy with amlodipine and verapamil on heart rate variability (HRV) in patients with essential hypertension. Methods: Forty patients with essential hypertension (11 men and 29 women, mean age 50.5+/-10.4 years) were included in the study. Patients with cardiac, metabolic, or any other systemic disease were excluded. Patients were randomized to receive either amlodipine (10 mg; n=20) or verapamil (240 mg; n=20). Patients underwent 24-h Holter monitoring assessment before treatment and after the 4-week treatment period. Standard deviation of normal RR intervals (SDNN), standard deviation of all 5-min mean normal RR intervals (SDANN), square root of the mean of the sum of the squares of differences between adjacent RR intervals (r-MSSD), and pNN50 (time domain variables) and TF, high frequency power (HF), low-frequency power (LF), and sympathovagal balance (LF/HF; frequency domain variables) were analyzed before and after treatment. Results: Blood pressure (BP) was reduced to a similar degree, from 182/104 to 128/85 mmHg with verapamil and from 174/100 to 124/86 mmHg with amlodipine (verapamil p<0.001; amlodipine p<0.001). This study revealed that amlodipine had no significant effect on any of the time or frequency domain parameters. In contrast, in patients on verapamil, there were significant increases in all time domain parameters, and the LF/HF ratio was significantly decreased (p<0.05). Conclusions: These results suggest that verapamil may have additional positive effects on sympathico-parasympathetic control beyond lowering blood pressure compared with amlodipine, even after short-term treatment in hypertensive patients. PMID- 15288677 TI - Serum lipids and estrogen receptor gene polymorphisms in male-to-female transsexuals: effects of estrogen treatment. AB - The effects of chronic administration of estrogens on the lipid profile in males are not fully understood. We have studied the effect of chronic administration of estrogens on the lipid profile in a group of transsexual (TS) Canarian men who were taking estrogens and anti-androgens for a minimum of 3 years. In this cross sectional study of cases (n=27) and controls (n=26), plasma lipid profile and selected biochemical and hormonal features were studied. TS subjects had shorter stature than controls, and, after adjusting for height and weight, we found that they had lower values of serum free testosterone (FT) and higher estradiol (E2) levels than controls. The TS group had lower total and low-density lipoprotein (LDL) cholesterol and lower apoprotein B (Apo B) levels than the control group. Biochemistry was similar in both groups. The distribution of estrogen receptor gene polymorphisms (ER-Pvu and ER-Xba) was also similar in both groups. Serum Apo B concentration was related to ER-Xba polymorphism. No other association between lipid profile and the distribution of ER-Pvu and ER-Xba was found. We conclude that the chronic administration of estrogens in men could produce an increase in serum estradiol, a decrease in free testosterone levels, and a reduction in total cholesterol, LDL-cholesterol, and Apo B levels. The ER-Xba polymorphism may influence the Apo B response to exogenous estrogen in males. PMID- 15288678 TI - Phenotype and genotype of French cystic fibrosis patients with long survival and follow-up. AB - Background: The aim of this study was to assess the clinical features and the genotype characteristics of French patients diagnosed with cystic fibrosis (CF) before their fifth year and who were still alive after 30 years. It is the first descriptive study of 114 CF patients with long survival and follow-up. We compared this subgroup of French CF patients with the overall French CF population and with French adult (> 18 years) CF patients regardless of their age at diagnosis. Methods: Data were obtained from the French CF registry. Results: The 67 men and 47 women studied were 30-59 years old. Some 56% of the patients had DeltaF508 homozygous genotype, 90% had a pancreatic insufficiency, and 81% were colonized with Pseudomonas aeruginosa. The mean body mass index (BMI) was 19.5 for both female and male patients. Mean forced expiratory volume in 1 s was 46% (S.D. 29.2) of the predicted value for men and 53% (S.D. 20.6) for women. Eleven patients underwent a lung transplantation. Conclusions: The data on the patients with long survival and long follow-up were very similar to the data of the overall CF adult population in terms of clinical status. Therefore, criteria such as DeltaF508 homozygous genotype, pancreatic insufficiency, and P. aeruginosa colonization are not sufficient to serve as prognostic criteria for life expectancy in CF patients. PMID- 15288679 TI - Efficacy and tolerability of ebastine 10 mg plus pseudoephedrine 120 mg in the symptomatic relief of the common cold. AB - Background: The objective of this multicenter, parallel, double-blind, placebo controlled clinical trial was to determine the efficacy and tolerability of the combination of ebastine 10 mg plus pseudoephedrine 120 mg once daily after 3 days of treatment in the symptomatic relief of patients with a common cold. Methods: The principal variable studied was the evaluation of overall efficacy and secondary variables were improvement of the patient, evolution of symptoms, disposition of the patient to take the medication again, and variation in nasal peak flow. Results: The percentage of subjects showing a good or excellent treatment efficacy was significantly higher in the group treated with ebastine plus pseudoephedrine (75.8%) than in the group treated with placebo (57.6%; p<0.001). Statistically significant differences were also found in favor of ebastine plus pseudoephedrine when comparing the changes in the sum of scores for nasal and ocular symptoms (p<0.006) or total symptoms (p<0.0016). The tolerability of the active treatment studied was good; that is, no significant differences were found between ebastine 10 mg plus pseudoephedrine 120 mg and placebo. Most adverse events described were slight or moderate in intensity, and no serious adverse events were reported. Conclusions: The combination of ebastine 10 mg immediate release and pseudoephedrine 120 mg sustained release was found effective in the symptomatic treatment of patients suffering from a common cold and as safe as placebo. PMID- 15288680 TI - Acute chest syndrome in sickle-cell trait; Two case reports in persons of Mediterranean origin and review of the literature. AB - Sickle-cell trait has been characterized as a benign condition. However, life threatening complications sometimes develop. Acute chest syndrome (ACS) is usually described in homozygous sickle cell disease, but it rarely develops in individuals with sickle-cell trait. Two cases of ACS in patients with sickle-cell trait are reported here. Factors such as advanced age at the time of presentation and absent past medical and family history can be misleading. Although ACS in sickle-cell trait has thus far only been reported in persons of Afro-American origin, persons of Mediterranean origin can, on rare occasions, also experience the syndrome. PMID- 15288681 TI - Anomaly of the inferior vena cava causing recurrent deep vein thrombosis in a young male. AB - Anomaly of the inferior vena cava (IVC) as a cause of (recurrent) deep vein thrombosis (DVT) is uncommon. We report on a 33-year-old patient suffering from this unusual cause of DVT without the presence of known predisposing factors (immobilization, trauma, surgery, or underlying thrombophilia). Thus, in young patients with recurrent DVT, anomaly of the IVC should be regarded as an independent risk factor. PMID- 15288682 TI - Nevirapine-induced toxic epidermal necrolysis and toxic hepatitis treated successfully with a combination of intravenous immunoglobulins and N acetylcysteine. AB - We describe a case of an HIV-seropositive patient presenting with a severe stomatitis that initially improved with anti-infective agents. Only 13 days after the onset of the stomatitis, the patient developed rapidly progressive constitutional symptoms and a cutaneous eruption. He was diagnosed with a Stevens Johnson syndrome (SJS) caused by the antiretroviral drug nevirapine (NVP). Despite meticulous supportive care and withdrawal of all drugs, his situation worsened and developed into a toxic epidermal necrolysis (TEN), or Lyell's syndrome, complicated by a toxic hepatitis. Treatment with a novel combination of intravenous immunoglobulins (IVIG) and N-acetylcysteine (NAC) resulted in an exceptionally fast recovery. A literature research revealed no other cases of patients treated with both NAC and IVIG for the combination of TEN and toxic hepatitis. Because of the rapid clinical recovery, this approach merits further investigation. This case report also illustrates the importance of early suspicion of SJS when an HIV-infected patient treated with nevirapine presents with stomatitis. PMID- 15288683 TI - Obstructive jaundice and hematemesis: two cases with unusual presentations of intra-abdominal tuberculosis. AB - Extrapulmonary tuberculosis (TB) is notorious for its many manifestations, which can lead to delayed diagnosis and treatment. In particular, abdominal tuberculosis is easily overlooked because the incidence is low and because it can mimic common noninfectious abdominal syndromes. We describe here a 37-year-old Moroccan man presenting with obstructive jaundice and a 42-year-old man originating from the Philippines who presented with recurrent hematemesis due to portal hypertension. In both patients, a retroperitoneal mass was found and tuberculosis was first diagnosed after a diagnostic laparotomy. Tuberculosis should be included in the differential diagnosis of intra-abdominal mass lesions, especially in persons originating from regions where tuberculosis is endemic. Invasive procedures are often required to obtain adequate diagnostic samples. PMID- 15288684 TI - Thrombotic thrombocytopenic purpura due to alcohol binge drinking. AB - Binge alcohol drinking is a pattern of alcohol abuse that is common among young males worldwide. It has been found to be associated with an increased likelihood of injury as a cause of death. Chronic alcohol abuse is known to cause some common hematological manifestations such as macrocytosis, thrombocytopenia, sideroblastic anemia, global marrow suppression, and folic acid deficiency anemia. We present a rare case involving an unusual and severe hematological manifestation of binge alcohol drinking: thrombotic thrombocytopenic purpura (TTP). The patient we present had severe and prolonged TTP necessitating prolonged treatment with plasmapheresis and plasma exchange. We discuss the relevant medical literature and the possible physiopathology of this complication. PMID- 15288685 TI - Acute hepatitis due to brucellosis in a laboratory technician. PMID- 15288686 TI - Interferon-alpha therapy and autoimmune manifestations. PMID- 15288687 TI - The Bcl-2 protein family and its role in the development of neoplastic disease. AB - Programmed cell death is the physiological process responsible for shaping organs during embryogenesis, maintaining tissue homeostasis and allowing controlled deletion of potentially harmful cells within the adult organism. The genetics of apoptosis are well conserved in all metazoans and although the evolution of humans and worms separated more than 600 million years ago, basic signaling concepts in apoptosis are highly related in both species. More crucial to humans than worms is the fact that abnormalities in cell death control can contribute to the development of cancer. While C.elegans can easily survive with additional somatic cells that should normally be deleted during development humans may suffer pathological consequences, ranging from tumorigenesis to autoimmunity, as a result of mutations in cell death regulatory genes. Despite the high degree of evolutionary conservation in cell death control, apoptosis signaling in mammals is much more complex than in C.elegans. In mammalian cells, programmed cell death can be induced either by ligand-mediated activation of certain members of the tumor necrosis factor receptor family--so-called 'death receptors'--such as Fas (CD95/Apo-1) and TRAIL or it can be induced in a cell autonomous manner in response to certain stress signals by pro-apoptotic members of the Bcl-2 family. In this review, we focus on general concepts of how the Bcl-2 protein family regulates cell death and how deregulation of this 'intrinsic' apoptotic signaling pathway impinges on the pathogenesis of malignant disease, the major cause of death in the aging population. PMID- 15288688 TI - Metabolic rate is not reduced by dietary-restriction or by lowered insulin/IGF-1 signalling and is not correlated with individual lifespan in Drosophila melanogaster. AB - The link between resting metabolic rate and aging, measured as adult lifespan, was investigated in Drosophila melanogaster by (i) comparing lifespan and metabolic rate of individual flies, (ii) examining the effect of dietary restriction on the metabolic rate of adult flies, and (iii) comparing the metabolic rate of wild-type and insulin/IGF-1 signalling mutant chico1 flies. The resting oxygen consumption of 65 individually housed and fully fed Drosophila was measured weekly throughout their lifetime. There was no significant difference in the mass-specific rate of oxygen consumption between cohorts that differed in lifespan. Nor was there any statistical correlation between mass-specific oxygen consumption and lifespan of individual Drosophila. The average mass-specific rate of oxygen consumption at 25 degrees C was 3.52+/-0.07 microl O2 mg(-1) h(-1). Variation in mass-specific metabolic rate explained only 4% of variation in individual life span in these flies. Contrary to predictions from the 'rate of living' theory of aging lifetime oxygen consumption was not constant and the lifespan of individual flies accounted for 91% of their lifetime oxygen consumption. An average Drosophila consumes about 3 ml O2 during its adult life. Dietary-restriction had no effect on mass-specific resting metabolic rate both when measured as oxygen consumption by respirometry and when measured as heat production by microcalorimetry. The mass-specific resting heat production of fully fed adult flies at 25 degrees C averaged 17.3+/-0.3 microW mg(-1). Similarly there was no difference in mass-specific metabolic rate of wild-type flies and longliving chico1 insulin/IGF-1 signalling mutant flies, either when measured as oxygen consumption or heat production. Thus, individual variation in lifespan in wild-type flies, and life extension by dietary-restriction and reduced insulin/IGF-1 signalling is not attributable to differences in metabolic rate. PMID- 15288689 TI - Krebs cycle enzymes from livers of old mice are differentially regulated by caloric restriction. AB - Krebs cycle enzyme activities and levels of five metabolites were determined from livers of old mice (30 months) maintained either on control or on long-term caloric restriction (CR) diets (28 months). In CR mice, the cycle was divided into two major blocks, the first containing citrate synthase, aconitase and NAD dependent isocitrate dehydrogenase which showed decreased activities, while the second block, containing the remaining enzymes, displayed increased activity (except for fumarase, which was unchanged). CR also resulted in decreased levels of citrate, glutamate and alpha-ketoglutarate, increased levels of malate, and unchanged levels of aspartate. The alpha-ketoglutarate/glutamate and malate/alpha ketoglutarate ratios were higher in CR, in parallel with previously reported increases with CR in pyruvate carboxylase activity and glucagon levels, respectively. The results indicate that long-term CR induces a differential regulation of Krebs cycle in old mice and this regulation may be the result of changes in gene expression levels, as well as a complex interplay between enzymes, hormones and other effectors. Truncation of Krebs cycle by CR may be an important adaptation to utilize available substrates for the gluconeogenesis necessary to sustain glycolytic tissues, such as brain. PMID- 15288691 TI - Modulation of gene expression of SMP-30 by LPS and calorie restriction during aging process. AB - Senescence marker protein-30 (SMP-30) has been proposed as an important aging marker and is now functionally identified as a Ca2+ binding protein. SMP-30 has been shown to blunt cell death caused by intracellular Ca2+ accumulation by enhancing plasma membrane Ca(2+)-pumping activity. Although SMP-30 is reported decrease during aging, at present, neither has the mechanism underlying this decrease been fully defined, nor have the mechanisms related to the modulation of SMP-30 been extensively explored. In the current study, we used the well-known anti-aging action of the calorie restriction (CR) paradigm to explore age-related changes in SMP-30 gene expression. The thrust of our investigation was based on CR's ability to defend against age-related oxidative stress and the inflammatory process. The kidney and liver from Fischer 344 rats at 6, 12, 18 and 24 months of age were utilized for this study. The rats were divided into two groups, ad libitum (AL)-fed and 40% restricted CR. Results showed that SMP-30 expression declined with age and that this decline was clearly blunted by CR. To correlate changes between SMP-30 gene expression and the oxidative status, SMP-30 expression and the production of reactive oxygen species (ROS) during aging and free-radical generating lipopolysaccharide (LPS) were monitored. Our data showed that the down-regulation of SMP-30 was accompanied by increased ROS generation and LPS-induced ROS. The potent anti-aging and anti-oxidative action of CR effectively suppressed the age-related down-regulation of SMP-30 by ROS reduction. PMID- 15288690 TI - Proteomic analysis of post-mitochondrial fractions of young and old rat kidney. AB - Proteomic analysis is defined as the characterization of the entire set of proteins encoded by a genome. Two-dimensional (2D) electrophoresis and matrix assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) are key technologies used in proteomic analysis to gain information about protein expression profiles and post-translational modifications. Knowledge about aging processes can be gained by recognizing changes in protein expression. Thus, to better understand the aging process through protein profiling, post mitochondrial (PM) fractions of young (13-month) and old (31-month) male Fischer 344 rat kidney were differentially analyzed by 2D. We detected a total number of 380 spots on 2D gel images. Among them, 167 spots showed 2-fold significant alterations (p<0.05) between young and old PM fractions. Further, 103 proteins were identified by MALDI-TOF MS. The PM fraction of aged rat kidney showed increases in antioxidative and proteolytic proteins and decreases in cytoskeletal proteins. In addition, we found age-related changes in transport and homeostasis proteins. Thus, our results demonstrated that proteomic analysis can be effectively applied to the assessment of the age status of protein expression, and thereby provide valuable information on age-related changes of proteome. PMID- 15288692 TI - Analyses of hind leg skeletons in human growth hormone transgenic rats. AB - Growth hormone (GH) is essential in the development and growth of the skeleton and for the maintenance of bone mass and density, and its secretion is known to decline with aging. We have previously produced transgenic rats with low circulating GH that represent several age-associated phenotypes such as obesity, insulin-resistance and leptin-resistance. In the present study, the cross sectional area, bone mineral density, and strength indexes of the hind leg skeletons of the transgenic rats were examined by an X-ray computed tomography scanning. The mean cross-sectional area of the transgenic rats showed no increase after 2 months old up to 8 months old and the strength indexes were significantly lower than their non-transgenic siblings at all ages examined. The trabecular bone mineral density in the transgenic rats drastically decreased at 8 months old, while the cortical bone mineral density was comparable to the non-transgenic rats, suggesting the onset of osteoporosis at this period. The results obtained in this study indicate that the transgenic rats could be useful model to gain insight into the complex mechanism leading to osteoporosis with aging. PMID- 15288693 TI - Dietary fat type (virgin olive vs. sunflower oils) affects age-related changes in DNA double-strand-breaks, antioxidant capacity and blood lipids in rats. AB - This study was designed to investigate the possible effect on DNA double-strand breaks, antioxidant capacity and blood lipids of feeding rats lifelong with two different dietary fat sources: virgin olive oil (rich in the monounsaturated oleic acid) or sunflower oil (rich in the polyunsaturated linoleic acid). No changes in mean or maximal lifespan were observed. Overall, aging led to increased levels of plasma cholesterol, triglycerides, phospholipids, total lipids, polyunsaturated fatty acids and DNA double-strand breaks. All these parameters were higher in animals fed on sunflower oil diet. Aging diminished total antioxidant capacity with both diets, but in a lower extension for virgin olive oil diet. A very good inverse correlation (r= -0.715; P < 0.01, for sunflower oil group and r= -0.535; P < 0.01 for virgin olive oil group) between DNA damage and total antioxidant capacity was found. These results allow to conclude that dietary fat type should be considered in studies on aging, since the intake of oils with different polyunsaturation levels directly modulates total antioxidant capacity of plasma, DNA damage to peripheral blood lymphocytes and lead to important changes at the lipid metabolism level. In the present study best results were found after intake of virgin olive oil, which suggest the possible use of that edible oil to provide a healthier aging. PMID- 15288694 TI - Effect of age and caloric restriction on coenzyme Q and alpha-tocopherol levels in the rat. AB - Alterations in the amount of coenzyme Q and alpha-tocopherol during aging and in response to 40% reduction in caloric intake were determined in homogenates and mitochondria of liver, heart and kidney of the rat. A comparison among 4-, 19- and 28-month-old ad libitum fed (AL) rats indicated an age-related loss in the amount of CoQ9 and alpha-tocopherol in mitochondria of all the three tissues. Depletion of alpha-tocopherol, but not of CoQ, was also detectable in tissue homogenates, apparently due to the preferential sequestration of CoQ in the mitochondrial fraction. Comparison of 19-month-old AL and calorically restricted (CR) rats indicated that CR elevates the level of mitochondrial CoQ, but greatly diminishes the alpha-tocopherol content. Activity of DT-diaphorase, a quinone reductase, increased with age as well as in response to CR. Altogether, results are interpreted to suggest that the widely observed age-related increase in mitochondrial oxidative damage may be associated with depletion of CoQ and alpha tocopherol, which are known to act in tandem to prevent oxidative damage to membranes. PMID- 15288695 TI - Potential involvement of NOS and arginase in age-related behavioural impairments. AB - The present study investigated age-related changes in nitric oxide synthase (NOS) and arginase, which shares a substrate with NOS, in the hippocampus and parahippocampal region and the relationship between NOS/arginase and age associated behavioural impairments. Aged rats (24 months old) displayed reduced exploratory activity, enhanced anxiety, poorer spatial learning and memory, and impaired object recognition memory relative to the young adults (4 months old). There were significant increases in total NOS activity in the aged hippocampus and perirhinal, postrhinal and temporal cortices and a dramatic decrease in endothelial NOS expression in the aged postrhinal cortex. Activity and protein expression of inducible NOS were not detected in any region from either group and a significant increase in total arginase activity was found in the aged perirhinal cortex. Multiple regression analysis revealed significant correlations between NOS/arginase and behavioural measures in both groups. The present findings provide further support for a contribution of nitric oxide to the normal aging process and suggest a potential involvement of arginase in aging and learning and memory. PMID- 15288696 TI - Investigation of KIR diversity in immunosenescence and longevity within the Irish population. AB - Natural killer (NK) cells play a pivotal role in the innate immune response. During the ageing process, variations occur in NK cell number and function. The cytolytic activity of NK cells is controlled by an array of activating and inhibitory cell surface receptors, including the killer cell immunoglobulin-like receptors (KIRs). In the present study, genetic diversity of the KIR loci was analysed with respect to successful ageing in the Irish population. A PCR-SSOP KIR gene identification system was employed to determine the frequency of the named KIR genes/pseudogenes and KIR genotypes within a healthy aged cohort and young control group. Although, two KIR genes (2DS3, 2DL5) displayed an initial increased frequency in the aged group, the significance of this association was lost when repeated in a second cohort. In view of the lack of studies to date, investigating the role of the KIR gene system in healthy ageing, further analysis of KIR diversity is required to fully elucidate it's role in respect to age related disease and longevity. PMID- 15288697 TI - Different contribution of EBV and CMV infections in very long-term carriers to age-related alterations of CD8+ T cells. AB - Aging is accompanied by a complex dynamics of CD8+ T cell subsets whose origin is unclear. To evaluate the impact of Epstein-Barr virus (EBV) and cytomegalovirus (CMV) chronic infections on CD8+ T cells in far advanced age, we studied CD8+ T cells frequencies and phenotype in nonagenarians and centenarians by HLA-A*0201- and HLA-B*0702-tetramers incorporating epitopes specific of both viruses along with viral replication. The results demonstrate that EBV and CMV infections induce quantitatively and qualitatively different CD8+ T-cell responses in advanced aging. The frequency and absolute number of CD8+ T cells specific for one lytic and two latent EBV-epitopes, were relatively low and mostly included within CD8+ CD28+ cells. By contrast, CMV infection was characterized by highly variable numbers of CD8+ T cells specific for two differently restricted CMV epitopes that, in some subjects, were strikingly expanded. Moreover, the great majority of anti-CMV CD8+ T cells did not bear CD28 antigen. Notwithstanding the expansion of CMV-specific CD8+ lymphocytes, CMV-DNA detection in blood samples was invariably negative. Altogether, we suggest that CMV, but not EBV, can sustain chronic activation of the HLA-class I restricted effector arm in elderly that might have detrimental effects on age-associated diseases. PMID- 15288698 TI - Basal cerebral blood flow is dependent on the nitric oxide pathway in elderly but not in young healthy men. AB - OBJECTIVE: Brain perfusion is tightly regulated over a wide range of blood pressures by local regulation of cerebral blood flow (CBF). Ageing is associated with impaired CBF and impaired nitric oxide mediated vasodilator responses. The role of nitric oxide in the regulation of basal CBF in young and older subjects was investigated, using the nitric oxide synthase inhibitor L-NMMA as pharmacological tool. METHODS: We used a gradient echo phase-contrast magnetic resonance imaging technique to investigate the role of nitric oxide in the regulation of cerebral blood flow in young (25+/-7.1 years; n=8) and old (78+/ 6.6 years; n=7) volunteers. The study was performed in a double-blinded fashion and consisted of two study days. On one day the effects of the intravenously infused L-NMMA on CBF and blood pressure was measured and on the other day the effects of a matching placebo. RESULTS: Basal CBF was significantly lower in old compared to young subjects (590+/-20 vs 704+/-20 ml/min), while the cerebral vascular resistance (CVR) levels were significantly higher (0.15+/-0.01 (arbitrary units) vs 0.12+/-0.01, respectively). Infusion of L-NMMA significantly increased mean arterial pressure in both groups (2.8+/-1.2 mmHg; p=0.02 in the young and in the old subjects 5.6+/-1.1 mmHg; p<0.001). Infusion of L-NMMA significantly decreased CBF (49+/-12 ml/min; p<0.001) and increased CVR (0.02+/ 0.004; p<0.001) in the old subjects but did not significantly influence cerebral circulation in the young subjects. CONCLUSION: We conclude that compared to young subjects, in old people CBF is impaired, and dependent on the intactness of the nitric oxide pathway. PMID- 15288699 TI - Monocyte chemoattractant protein-1 (MCP-1) gene polymorphism and risk of Alzheimer's disease in Italians. AB - Monocyte chemoattractant protein-1 (MCP-1) is a key molecule for monocyte chemotaxis and tissue extravasation and for the modulation of leukocyte function during inflammation. Upregulation of MCP-1 may occur in the brain of subjects affected by Alzheimer's disease (AD) and MCP-1 levels in plasma and cerebrospinal fluid have been proposed as biological markers for the inflammatory process that accompanies AD pathogenesis. Importantly, serum levels and biological activity of MCP-1 protein are strongly influenced by a single nucleotide polymorphism occurring at position -2518 of the MCP-1 gene promoter. A recent study has investigated the possible association between this gene polymorphism and AD in a Spanish population, with negative results. Here, we performed a case-control study to test whether the risk for AD might be influenced by the -2518 A/G polymorphism of the MCP-1 gene in an ethnically homogeneous Italian population. The GG genotype and the G allele of the MCP-1 gene polymorphism were significantly more common in the AD group than in control individuals (P<0.0001) A logistic regression analysis indicated that the GG genotype was an independent risk factor for AD in our population. This effect was not influenced by the presence of the APOE 4 high-risk allele, nor by the presence of other gene variations associated with a pro-inflammatory phenotype. These findings indicate that the -2518 A/G polymorphism of the MCP-1 gene is associated with AD in Italians and confirm that inflammatory gene variations may be important contributors in the development and progression of neurodegenerative disorders. PMID- 15288700 TI - Age-dependent modifications of expression level of VEGF and its receptors in the inner ear. AB - The mechanisms responsible for age-associated hearing loss are still incompletely characterized. In this study, we used a murine model of age-dependent hearing loss and evaluated whether this condition is associated with vascular modifications of the structures of the inner ear. We used old C57BL/6J mice that are affected by rapid and severe age-related hearing loss, and analyzed the expression pattern of vascular endothelial growth factor (VEGF), a prototypical angiogenic cytokine, and its receptors Flt-1 and Flk-1 in the inner ear. We report for the first time morphological and quantitative data about the expression of these crucial angiogenic molecules in the murine cochlea. We also show that in this animal model, cochlear VEGF expression is significantly reduced as a function of age. Our findings provide new evidence of possible interdependent relationships between aging, VEGF, and presbycusis, suggesting that vascular abnormalities might play a role in aging-associated hearing loss, with potentially important fundamental and clinical implications. PMID- 15288701 TI - Effects of acute and chronic ketoconazole administration on hypothalamo- pituitary--adrenal axis activity and brain corticotropin-releasing hormone. AB - We have been investigating the effects of ketoconazole on cocaine reward in rats for several years now. However, we recently confirmed that ketoconazole-induced changes in cocaine self-administration and reinstatement do not always correspond with decreases in plasma corticosterone, which suggests that other mechanisms must be underlying the behavioral effects that we observe. This experiment was therefore designed to determine the effects of acute, repeated and chronic ketoconazole administration on corticotropin-releasing hormone (CRH) content in hypothalamic and extra-hypothalamic brain sites in rats following the same dosing regimen that we use in our behavioral studies. Although ketoconazole significantly increased the concentration of ACTH in trunk blood, there were no significant effects on plasma cortisol, corticosterone or testosterone. There was also a significant increase in CRH content in the median eminence after the acute administration of ketoconazole that just failed to reach statistical significance following repeated or chronic administration. However, acute, repeated and chronic treatment with ketoconazole each significantly increased CRH content in the medial prefrontal cortex (MPC), but did not consistently affect the peptide in any other brain region studied. Since the MPC and CRH have been implicated in the neurobiology of cocaine, CRH-induced alterations in dopaminergic neurotransmission may play an important role in this peptide's effects on cocaine responsiveness. Taken together with the results from previous studies, these data suggest that ketoconazole may affect cocaine reward, at least in part, through interactions with dopamine and CRH within the MPC. PMID- 15288702 TI - The "trouble" with salivary testosterone. AB - In a series of studies, we identify several specific issues that can limit the value of integrating salivary testosterone in biosocial research. Salivary testosterone measurements can be substantially influenced during the process of sample collection, are susceptible to interference effects caused by the leakage of blood (plasma) into saliva, and are sensitive to storage conditions when samples have been archived. There are gender differences in salivary testosterone levels and variance, the serum-saliva association, the relationship of salivary testosterone to age and pubertal development, and the stability of individual differences in salivary testosterone levels over time. The findings have important implications at several levels of analysis for research that aims to test biosocial models of testosterone--behavior relationships. Recommendations are provided to steer investigators around these "troubles" with salivary testosterone. PMID- 15288703 TI - Chronic fluoxetine suppresses circulating estrogen and the enhanced spatial learning of estrogen-treated ovariectomized rats. AB - We are interested in developing animal models to evaluate cognitive processes as influenced by the interplay of steroidal hormones and drugs commonly used in psychotherapy. Two experiments with female rats were conducted to evaluate the interaction of estrogen with the serotonin specific reuptake inhibitor (SSRI) fluoxetine on spatial learning and memory and on the endocrine system. In experiment 1, estrogen (50 microg estradiol benzoate/kg body weight) was administered SC to young adult, ovariectomized (OVX) rats either alone or in combination with fluoxetine (2 mg/kg SC). After a month, the groups were compared with appropriate OVX and gonadally intact controls on trials to criterion in a hole board spatial memory task using massed training trials. Experiment 2 was a dose-response study of the influence of fluoxetine (0.5-5 mg/kg) on circulating estrogen in OVX, estrogen treated females. Results were that the OVX females administered estrogen only reached the learning criterion significantly faster than the other groups. All other groups, including the estrogen + fluoxetine animals, performed no better than the controls. Combining fluoxetine with estrogen also lowered circulating estrogen titers, with the least estrogen reductions being in the group receiving the highest dosage of fluoxetine. No differences among groups were found on measures of activity in an open field or for anxiety in a plus maze. Conclusions were that administration of estrogen improved spatial learning and memory in OVX rats, whereas concurrent fluoxetine exposure suppressed the levels of estrogen in circulation and eliminated the gains in spatial performance obtained from chronic estrogen exposure. PMID- 15288704 TI - Uncoupling of social zeitgebers and diurnal cortisol secretion in clinical depression. AB - Daily activities (zeitgebers) such as waking, eating, and exercising, done alone or in the presence of others, may help to entrain biological rhythms. To examine whether the relationship between zeitgebers and biological rhythms is altered in depression, this study tracked daily activity and cortisol secretion in 50 depressed and 50 control participants using a daily diary methodology. The groups reported similar levels of regular daily activities. Among control participants, regular daily activities were associated with a normative decline in cortisol secretion. Among depressed participants, daily activities and cortisol secretion were unrelated, consistent with the hypothesis that these activities are less able to entrain diurnal rhythms. This lack of social entrainment may underlie some of the circadian disturbances in depression. PMID- 15288705 TI - Tone probe event-related potential differences during a face recognition task in prepubertal children and Turner Syndrome girls. AB - Hormones have been shown to play a role in both cerebral development and neurocognitive function. Turner Syndrome (TS) provides the opportunity to study the effect of the lack of estrogen on neurocognitive development. In this study, event-related potential (ERP) differences were examined among 12 TS girls, 20 prepubertal control girls, and 20 prepubertal control boys during a face recognition memory task. Stage of puberty was determined by Tanner Scale rating and hormonal assay. ERPs to pairs of auditory probe stimuli were recorded from eight scalp sites while participants performed a faced recognition memory (FRM) task. For the N2 component of the ERP (which has previously been associated with evaluation of stimulus information, categorization difficulty, and attention), control boys displayed greater right versus left hemisphere amplitude, control girls displayed greater left versus right hemisphere amplitude, and there was no amplitude asymmetry for TS girls. Further, control girls had greater left hemisphere N2 amplitude than control boys and TS girls, and greater right hemisphere N2 amplitude than TS girls. The results suggest more right hemisphere activation during face recognition in boys, while the opposite pattern was present in control girls. In contrast, TS girls displayed no asymmetry, indicative of more uniform involvement of the left and right hemispheres during face recognition. These findings are consistent with differences in cortical organization related to face recognition memory processing among prepubertal control boys, girls, and TS girls. They also support the notion that sex differences in cognitive function are present prior to pubertal onset, and that lack of endogenous sex hormones (e.g., estrogen) during prenatal/perinatal development (i.e., for TS girls) may influence brain organization and, in turn, neurocognitive processes that relate to face recognition. PMID- 15288706 TI - Neuroendocrine response to casino gambling in problem gamblers. AB - Problematic gambling is thought to be influenced by neurobiological mechanisms. However, the neuroendocrine response to gambling is largely unknown. Therefore, the effect of casino gambling on the sympathoadrenal system, the HPA-axis, and pituitary hormones were analyzed. Fourteen male problem gamblers and 15 non problem gamblers were examined in a balanced cross-over design. In the experimental session, participants played blackjack in a casino wagering their own money. During the control session, subjects played cards for accumulation of points. Heart rate and endocrine measures were recorded at baseline, at 30, 60 and 90 min during gambling/card playing, and after the game. Heart rate and norepinephrine levels increased with the onset of blackjack in both groups, with problem gamblers showing significantly higher levels across the entire gambling session. In addition, dopamine levels were significantly higher in problem gamblers during casino gambling compared to non-problem gamblers. Cortisol levels were transiently increased with the onset of blackjack in both groups. Casino gambling as a "real life" situation induces activation of the HPA-axis and the sympathoadrenergic system, with significantly more pronounced changes in problem gamblers. These findings may contribute to a better understanding of neuroendocrine disturbances in problem gambling. PMID- 15288707 TI - Increased adrenocorticotropin suppression following dexamethasone administration in sexually abused adolescents with posttraumatic stress disorder. AB - Evidence suggests that individuals with posttraumatic stress disorder (PTSD) have an enhanced sensitization of the hypothalamic--pituitary--adrenocortical (HPA) axis. However, few studies in adolescents have been performed. Fourteen sexually abused adolescent inpatients with DSM-IV PTSD (12 female, two male; mean +/- SD age, 16.2 +/- 1.9 years) were compared with 14 adolescent hospitalized controls (11 female, three male; mean age, 15.7 +/- 2.0 years). All subjects underwent a standard dexamethasone suppression test (DST, 1 mg given orally at 2300 h) five days after admission. Baseline blood samples were obtained at 0800 h, and the following day, adrenocorticotropin (ACTH) and cortisol levels were measured at 0800, 1600, and 2300 h. Clinical assessment included the Impact of Event Scale, Stanford Acute Stress Reaction Questionnaire, Beck Depression Inventory, and Coping Inventory for Stressful Situations. Post-DST ACTH levels were significantly lower in PTSD than in control adolescents (at 0800 h: P < 0.005; at 1600 h: P < 0.001; at 2300 h: P < 0.05). In patients, post-DST cortisol levels were reduced but not significantly. No correlations were found between ACTH and cortisol levels and time elapsed since trauma. These results demonstrate that sexually abused adolescents with PTSD show ACTH hypersuppression to DST suggesting enhanced glucocorticoid receptor sensitivity in the pituitary. PMID- 15288708 TI - Sniffing a human sex-steroid derived compound affects mood and autonomic arousal in a dose-dependent manner. AB - The effects of sniffing different concentrations of the human sex-steroid derived compound 4,16-androstadien-3-one (AND) on autonomic nervous system function and mood were measured in 60 subjects. The effects were sex-specific and concentration-dependent. Only high concentrations of AND (0.00625 M) increased positive mood (p < 0.03) and decreased negative mood (p < 0.05) in women compared to men, and had sympathetic-like effects in women (p < 0.003), and parasympathetic-like effects in men (p < 0.05). These findings further implicate AND in chemical communication between humans, but pose questions as to the path by which AND is transduced, whether through chemical sensing or transdermal diffusion. PMID- 15288709 TI - Personality characteristics and basal cortisol concentrations in adult male rhesus macaques (Macaca mulatta). AB - Although data show that psychosocial factors can regulate physiological processes, few data have been collected on normative populations. Studies in humans have suggested that personality characteristics might be related to regulation of the hypothalamic--pituitary--adrenal (HPA) axis. We explored the relationship between personality characteristics and plasma cortisol concentrations in adult male rhesus macaques. Two sets of blood samples were obtained from monkeys using a procedure with which they were very familiar; thus, cortisol concentrations reflected basal values. Analyses indicated high-excitable animals had lower basal cortisol concentrations during the afternoon period, and that low-confidence was associated with lower cortisol in the morning period, and lack of a circadian decline in the afternoon period. Sociability and equability were unrelated to cortisol levels. Our data confirm and extend some results found in human studies, and suggest that even in normal populations, personality characteristics are related to measures of HPA function. We propose that comparative studies of personality in nonhuman primates that parallel studies in humans can increase our understanding of mechanisms whereby personality may relate to mental and physical health outcomes. PMID- 15288710 TI - Effect of long-term estrogen therapy on dopaminergic responsivity in post menopausal women--a preliminary study. AB - Females have a higher prevalence than men of neuropsychiatric disorders in which dopaminergic abnormalities play a prominent role, e.g. very late-onset schizophrenia and Parkinson's disease (PD). The biological basis of these sex differences is unknown but may include modulation of the dopaminergic system by sex hormones, as there is preliminary evidence that estrogen modulates treatment response in these disorders. Furthermore, sex differences in dopamine-mediated cognitive decline suggest estrogen may also play a role in healthy aging. However, the effects of estrogen on the dopaminergic system are poorly understood, and nobody has examined the effect of long-term estrogen therapy (ET) on this system. We compared dopaminergic responsivity (growth hormone (GH) response to apomorphine) in post-menopausal women on ET to women who were ET naive. GH response to subcutaneous apomorphine (0.005 mg/kg) was measured in two groups of healthy post-menopausal women aged between 55 and 70 years: those taking ET (n = 13) and those who had never taken ET (n = 13). Neither group was taking any other medication. GH was measured at 15 min intervals from -30 min before administration of apomorphine to 90 min post-administration. GH response was measured in two ways: area under the curve (AUC) and maximum response over baseline (GH). There were no between-group differences in demographic or baseline variables. The ET treated women had a significantly greater (p = 0.03) AUC than ET naive women (mean +/- S.D.; 5.3 +/- 4.7 vs. 2.6 +/- 2.3). However, (GH) did not differ significantly between groups (6.1 mU/l +/- 6.2 vs. 2.7 mU/l +/- S.D. = 4.1). Also, analysis of GH response over time revealed a significant main effect of time (p < 0.0005), and a group by time interaction (p = 0.004) , but no significant main effect of group. Our results suggest that ET may enhance dopaminergic responsivity in post-menopausal women. Estrogen deficiency following menopause may partly explain age and gender differences in late-onset neuropsychiatric disorders. PMID- 15288711 TI - Neonatal lesions of the ventral hippocampal formation disrupt neuroendocrine responses to auditory stress in the adult rat. AB - Lesioning the ventral hippocampal formation (vHF) in the neonatal rat with an excitotoxin replicates several features of schizophrenia. Similar lesions in the adult rat disrupt the normal constraint of neuroendocrine responses to environmental stressors, which is of potential interest because the enhanced HPA axis and antidiuretic hormone activity in schizophrenia is linked to acute stress and hippocampal formation (HF) pathology. In the current study, we investigated the effects of neonatal ventral hippocampal formation lesions (NVHFL) on plasma adrenocorticotropin hormone (ACTH) and arginine vasopressin (AVP) responses following a 2-min acoustic stressor in the adult rat. Levels of the two hormones did not differ between SHAM-operated and NVHFL animals in their home cages. ACTH levels doubled in SHAM-operated animals immediately following stress, but increased more than six-fold in the NVHFL group. AVP levels were halved immediately following stress in SHAM-operated animals, but did not change significantly in NVHFL. Findings could not be attributed to intervening factors known to influence neuroendocrine activity. Thus, NVHFL appear to disrupt the HF mediated constraint of neuroendocrine responses to stress, and model the neuroendocrine dysfunction seen in schizophrenia. We posit that clarification of how NVHFL alters relatively "simple", well characterized, and phylogenetically preserved systems, such as the neuroendocrine system, may provide insight into the mechanism of hippocampal pathology in schizophrenia. PMID- 15288712 TI - Intranasal insulin improves memory in humans. AB - Previous studies have suggested an acutely improving effect of insulin on memory function. To study changes in memory associated with a prolonged increase in brain insulin activity in humans, here we used the intranasal route of insulin administration known to provide direct access of the substance to the cerebrospinal fluid compartment. Based on previous results indicating a prevalence of insulin receptors in limbic and hippocampal regions as well as improvements in memory with systemic insulin administration, we expected that intranasal administration of insulin improves primarily hippocampus dependent declaration memory function. Also, improvements in mood were expected. We investigated the effects of 8 weeks of intranasal administration of insulin (human regular insulin 4 x 40 IU/d) on declarative memory (immediate and delayed recall of word lists), attention (Stroop test), and mood in 38 healthy subjects (24 males) in a double blind, between-subject comparison. Blood glucose and plasma insulin levels did not differ between the placebo and insulin conditions. Delayed recall of words significantly improved after 8 weeks of intranasal insulin administration (words recalled, Placebo 2.92 +/- 1.00, Insulin 6.20 +/- 1.03, p < 0.05). Moreover, subjects after insulin reported signs of enhanced mood, such as reduced anger (p < 0.02) and enhanced self-confidence (p < 0.03). Results indicate a direct action of prolonged intranasal administration of insulin on brain functions, improving memory and mood in the absence of systemic side effects. These findings could be of relevance for the treatment of patients with memory disorders like in Alzheimer's disease. PMID- 15288713 TI - Acute reduction in secretory immunoglobulin A following smoking cessation. AB - Smokers report an increase in upper respiratory infections in the early phase of stopping smoking. One possible cause is a depletion in secretory immunoglobulin A (S-IgA) which has been observed in one study. The present study sought to establish this finding in smokers using nicotine patches. Ninety-two smokers, trying to stop smoking, were assessed whilst smoking and for up to six weeks of abstinence. All smokers were prescribed 15 mg 16-h nicotine patches. Among abstinent smokers, changes in S-IgA and saliva volume were assessed. During the preliminary analyses, we observed that for the pre-smoking cessation measure a longer time since the last cigarette was significantly related to lower S-IgA levels (P = 0.006). Consequently, the main analysis, of changes in S-IgA from pre cessation to post-cessation, was confined to those who had smoked within 0.5-1.5 h of the pre-cessation measure (n = 51). There was a significant decline in S IgA, relative to pre-smoking abstinence levels, following abstinence of one day (P = 0.027), but levels returned to pre-abstinence values after one week. There was no evidence of any significant changes in saliva volume following smoking cessation, relative to pre-cessation levels. Users of 15 mg patches are likely to experience a decline in S-IgA levels on the first day of smoking cessation, independent of saliva volumes, and this decline in S-IgA is likely to occur acutely, within the first few hours of smoking abstinence. This acute drop in S IgA appears to stem from a factor other than depletion of nicotine from the body. The observed decrease in S-IgA may help to explain the increased susceptibility of smokers to upper respiratory tract infections in the immediate post-cessation period. PMID- 15288714 TI - Effects of PhD examination stress on allopregnanolone and cortisol plasma levels and peripheral benzodiazepine receptor density. AB - Peripheral benzodiazepine receptor (PBR) density in blood platelets and plasma allopregnanolone concentration in humans were determined following acute stress as represented by PhD examination. Fifteen healthy PhD students participated. Heart rate, blood pressure, plasma allopregnanolone, plasma cortisol, and PBR density were measured at different time points. Allopregnanolone and cortisol concentration and PBR density were significantly increased during examination. A positive correlation between allopregnanolone and PBR density was found. PMID- 15288716 TI - Sudan and its new weapons of war. PMID- 15288715 TI - Why the world needs another malaria initiative. PMID- 15288717 TI - How much of China's success in tuberculosis control is really due to DOTS? PMID- 15288718 TI - Beyond the evidence in clinical guidelines. PMID- 15288719 TI - Anthrax and bioterrorism: are we prepared? PMID- 15288720 TI - Evidence-based staging system for malignant melanoma: is new necessarily better? PMID- 15288721 TI - Emerging pathogens: is Mycobacterium avium subspecies paratuberculosis zoonotic? PMID- 15288722 TI - Turning the health system 90 degrees down under. PMID- 15288723 TI - Why are 4 million newborn babies dying each year? PMID- 15288724 TI - Why is Ian Kennedy's Healthcare Commission damaging NHS care? PMID- 15288725 TI - Ombudsman's eighth report. PMID- 15288726 TI - Over-the-counter statins. PMID- 15288727 TI - Over-the-counter statins. PMID- 15288728 TI - Statins for multiple sclerosis. PMID- 15288729 TI - Statins for multiple sclerosis. PMID- 15288732 TI - Diuretics in the LIFE study. PMID- 15288733 TI - Enterobacter sakazakii in factories and households. PMID- 15288735 TI - Pupil blown by a puffer. PMID- 15288737 TI - Merck Sharpe and Dohme versus Laporte. PMID- 15288738 TI - What symbol should represent the medical profession? PMID- 15288739 TI - The effect of tuberculosis control in China. AB - BACKGROUND: China has 1.4 million new cases of tuberculosis every year, more than any country except India. A new tuberculosis control project based on short course chemotherapy was introduced in half the country in 1991, after a national survey of tuberculosis prevalence in 1990. Another survey was done in 2000 to re evaluate the national tuberculosis burden, providing the opportunity to assess the effect of the control project. METHODS: The survey in 2000 identified 375599 eligible individuals at 257 investigation points chosen from all 31 mainland provinces by stratified random sampling. Children (aged 0-14 years) were suspected of having tuberculosis if they had an induration of 10 mm or greater after a tuberculin skin test, and an abnormal fluorograph. Adults were suspected if they had a persistent cough, abnormal fluorograph, or both. Tuberculosis was diagnosed by chest radiograph, sputum-smear microscopy, and culture. FINDINGS: 365097 people were examined (97% of those eligible). Prevalences of pulmonary, culture-positive, and smear-positive tuberculosis in 2000 were 367 (95% CI 340 397), 160 (144-177), and 122 (110-137) per 100000 population, respectively. Between 1990 and 2000, prevalences of these three forms of the disease had fallen, respectively, by 32% (5-68), 37% (7-66), and 32% (9-51) more in areas in which the project was implemented than in non-project areas. For culture-positive disease, a 30% (9-48) reduction was directly attributable to the project. INTERPRETATION: Between 1991 and 2000, prevalence of tuberculosis was reduced significantly in areas of China by use of short-course chemotherapy following WHO guidelines. We estimate that in 2000, in a population of more than half a billion, there were 382000 fewer prevalent culture-positive cases and 280000 fewer prevalent smear-positive cases than there would otherwise have been. PMID- 15288740 TI - Prevalence, care, and outcomes for patients with diet-controlled diabetes in general practice: cross sectional survey. AB - BACKGROUND: Tight glycaemic control reduces microvascular complications in patients with type 1 and type 2 diabetes. We aimed to establish the proportion with type 2 diabetes treated through diet only and to determine levels of complications and quality of care received compared with patients on hypoglycaemic medication. METHODS: We undertook a cross-sectional study of 7870 patients with type 2 diabetes from a population of 253,618 patients from 42 general practices in the UK. Our primary outcome was process of care measures and diabetes-related complications. FINDINGS: 31.3% of all patients with type 2 diabetes are being managed with diet only (1% of the total population). More than four-fold variation between practices exists (range 15.6-73.2%). Patients treated with diet only are much less likely to have HbA1c (glycosylated haemoglobin) measurements, blood pressure, cholesterol, smoking, microalbuminuria testing, or screening for foot pulses recorded. 38.4% of patients with type 2 diabetes on medication have a HbA(1c) above 7.5% compared with 17.3% of those treated with diet only. Compared with those on medication, patients treated by diet only are more likely to have raised blood pressure and less likely to be on anti hypertensive medication; they are 45% more likely to have raised cholesterol and less likely to be prescribed lipid-lowering medication. Although fewer of those treated by diet (68%) have diabetes-related complications compared with those on medication (80%), the rate is much higher than for the population without diabetes. INTERPRETATION: Diabetics treated by diet only have significant rates of complications and are less likely than those on medication to be adequately monitored. There is great scope for improved management within general practice. PMID- 15288741 TI - An experimental study of determinants of group judgments in clinical guideline development. AB - BACKGROUND: Clinical guidelines for improving the quality of care are a familiar part of clinical practice. Formal consensus methods such as the nominal group technique are often used as part of guideline development, but little is known about factors that affect the statements produced by nominal groups, and on their consistency with the research evidence. METHODS: Cognitive behavioural therapy, behavioural therapy, brief psychodynamic interpersonal therapy, and antidepressants for irritable bowel syndrome, chronic fatigue syndrome, and chronic back pain were selected for study. 16 nominal groups in a factorial design allowed comparison of GP-only with mixed groups of GPs and specialists, provision of a literature review with no provision, and ratings made in the context of realistic or ideal levels of health-care resources. Participants rated appropriateness independently, and again after a facilitated meeting. Audiotapes of four group discussions were analysed. FINDINGS: There was agreement with the research evidence for 51% of 192 scenarios. Agreement was more likely if the group was GP-only, if a literature review was provided, or if the evidence was in accordance with clinicians' beliefs. Assumptions about the level of resources available had no impact. Clinical and social cues had mixed effects, irrespective of the research evidence. Qualitative analysis showed the modifying effect of clinical experience and beliefs about research evidence. INTERPRETATION: Guidelines cannot be based on data alone; judgment is unavoidable. The nominal group technique is a method of eliciting and aggregating judgments in a transparent and structured way. It can provide important information on levels of agreement between experts. However, conclusions can be at odds with the published literature. If they are, reasons need to be explicit. PMID- 15288743 TI - Seal finger. PMID- 15288742 TI - Mefloquine resistance in Plasmodium falciparum and increased pfmdr1 gene copy number. AB - BACKGROUND: The borders of Thailand harbour the world's most multidrug resistant Plasmodium falciparum parasites. In 1984 mefloquine was introduced as treatment for uncomplicated falciparum malaria, but substantial resistance developed within 6 years. A combination of artesunate with mefloquine now cures more than 95% of acute infections. For both treatment regimens, the underlying mechanisms of resistance are not known. METHODS: The relation between polymorphisms in the P falciparum multidrug resistant gene 1 (pfmdr1) and the in-vitro and in-vivo responses to mefloquine were assessed in 618 samples from patients with falciparum malaria studied prospectively over 12 years. pfmdr1 copy number was assessed by a robust real-time PCR assay. Single nucleotide polymorphisms of pfmdr1, P falciparum chloroquine resistance transporter gene (pfcrt) and P falciparum Ca2+ ATPase gene (pfATP6) were assessed by PCR-restriction fragment length polymorphism. FINDINGS: Increased copy number of pfmdr1 was the most important determinant of in-vitro and in-vivo resistance to mefloquine, and also to reduced artesunate sensitivity in vitro. In a Cox regression model with control for known confounders, increased pfmdr1 copy number was associated with an attributable hazard ratio (AHR) for treatment failure of 6.3 (95% CI 2.9-13.8, p<0.001) after mefloquine monotherapy and 5.4 (2.0-14.6, p=0.001) after artesunate-mefloquine therapy. Single nucleotide polymorphisms in pfmdr1 were associated with increased mefloquine susceptibility in vitro, but not in vivo. INTERPRETATION: Amplification in pfmdr1 is the main cause of resistance to mefloquine in falciparum malaria. RELEVANCE TO PRACTICE: Multidrug resistant P falciparum malaria is common in southeast Asia, but difficult to identify and treat. Genes that encode parasite transport proteins maybe involved in export of drugs and so cause resistance. In this study we show that increase in copy number of pfmdr1, a gene encoding a parasite transport protein, is the best overall predictor of treatment failure with mefloquine. Increase in pfmdr1 copy number predicts failure even after chemotherapy with the highly effective combination of mefloquine and 3 days' artesunate. Monitoring of pfmdr1 copy number will be useful in epidemiological surveys of drug resistance in P falciparum, and potentially for predicting treatment failure in individual patients. PMID- 15288744 TI - Clinical predictors of bioterrorism-related inhalational anthrax. AB - Limitation of a bioterrorist anthrax attack will require rapid and accurate recognition of the earliest victims. To identify clinical characteristics of inhalational anthrax, we compared 47 historical cases (including 11 cases of bioterrorism-related anthrax) with 376 controls with community-acquired pneumonia or influenza-like illness. Nausea, vomiting, pallor or cyanosis, diaphoresis, altered mental status, and raised haematocrit were more frequently recorded in the inhalational anthrax cases than in either the community-acquired pneumonia or influenza-like illness controls. The most accurate predictor of anthrax was mediastinal widening or pleural effusion on a chest radiograph. This finding was 100% sensitive (95% CI 84.6-100.0) for inhalational anthrax, 71.8% specific (64.8 78.1) compared with community-acquired pneumonia, and 95.6% specific (90.0-98.5) compared with influenza-like illness. Our findings represent preliminary efforts toward identifying clinical predictors of inhalational anthrax. PMID- 15288746 TI - Child sexual abuse. AB - Child sexual abuse is a worldwide concern. It is an insidious, persistent, and serious problem that, depending on the population studied and definition used, affects 2-62% of women and 3-16% of men as victims. Pain and tissue injury from child sexual abuse can completely heal in time, but psychological and medical consequences can persist through adulthood. Associated sexually transmitted diseases (such as HIV) and suicide attempts can be fatal. All physicians who treat children should be aware of the manifestations and consequences of child sexual abuse, and should be familiar with normal and abnormal genital and anal anatomy of children. This aim is best accomplished through training and routine examination of the anus and genitalia of children. Because as many as 96% of children assessed for suspected sexual abuse will have normal genital and anal examinations, a forensic interview by a trained professional must be relied on to document suspicion of abuse. PMID- 15288745 TI - Borderline personality disorder. AB - Borderline personality disorder is characterised by a pervasive pattern of instability in affect regulation, impulse control, interpersonal relationships, and self-image. Clinical signs of the disorder include emotional dysregulation, impulsive aggression, repeated self-injury, and chronic suicidal tendencies, which make these patients frequent users of mental-health resources. Causal factors are only partly known, but genetic factors and adverse events during childhood, such as physical and sexual abuse, contribute to the development of the disorder. Dialectical behaviour therapy and psychodynamic partial hospital programmes are effective treatments for out-of-control patients, and drug therapy can reduce depression, anxiety, and impulsive aggression. More research is needed for the understanding and management of this disabling clinical condition. Current strategies are focusing on the neurobiological underpinnings of the disorder and the development and dissemination of better and more cost-effective treatments to clinicians. PMID- 15288747 TI - Ganser's syndrome. PMID- 15288748 TI - A boy with a mediastinal mass. PMID- 15288749 TI - Weapon-carrying at Swiss schools? A gender-specific typology in context of victim and offender related violence. AB - After reviewing prevalence rates, this work tries to identify gender specific groups in which weapon-carrying occurs in the context of victim and offender related violent behaviours. k-Means cluster and logistic regression analyses were calculated, based on a cross-sectional survey of a national representative sample of 1549 15-year-olds as part of the Health Behaviour in School-Aged Children (HBSC) international collaborative study. The victims and offenders of violence concern mainly female bullies and physically violent boys. In the male victim offender group, 17% had already taken a weapon to school. Another male group emerged in which 95% had already taken a weapon to school. Members of this group revealed an elevated level of acting violently but a low level of being the victim. Among the minority girls who were offenders and victims of physical violence, everyone has already taken a weapon to school. Based on these results, homogenous risk groups can be targeted for violence prevention. PMID- 15288750 TI - Linkages among adolescent girls' romantic relationships, best friendships, and peer networks. AB - This study examined the linkages among girls' best friendships and romantic relationships and accounted for the level of dating involvement as a moderating variable. Social exchange and Sullivan's socioemotional theories served as guides in this process. Questionnaires were administered to 446 girls aged 15-19 years. Results showed that: (a) dating is associated with more positive and less negative interactions with best friends; (b) increasing age and dating involvement are linked with increased reliance on romantic partners; (c) romantic relationships have more negative interactions than best friendships; and (d) peer network size and structure is related to dating behaviour. Mid to late adolescence is a time when girls shift attention away from friends and towards romantic partners. PMID- 15288751 TI - The psychosocial needs of young offenders and adolescents from an inner city area. AB - To date, assessments of the prevalence of mental health problems in young offenders have largely focused on incarcerated samples. This paper describes a quantitative study of a sample of 47 male young offenders under the supervision of an inner city Youth Offending Team. A semi-structured interview, modified from previous studies, was used to investigate the type and frequency of psychosocial and health problems they experienced. The results were compared with data from a random community sample of 38 male adolescents interviewed in a comparable way. Young offenders reported more psychosocial problems at a statistically significant level (including depression/misery, excessive worry and problematic substance use), as well as higher life-time rates of head injury, than adolescents in the community sample. Levels of problematic substance use and exposure to head injury continued to differentiate the groups when offenders in custody were removed from the analyses. The findings are discussed in relation to previous research and implications for service delivery. PMID- 15288752 TI - Trends in adolescent alcohol and other drug use: findings from three sentinel sites in South Africa (1997-2001). AB - This paper aims to provide surveillance information about the extent and consequences of alcohol and other drug (AOD) use by adolescents for three sentinel sites in South Africa (Cape Town, Durban and Gauteng province). From 1997 to 2001, data were gathered from multiple sources, including specialist treatment centres, trauma units, school students, rave party attenders, and arrestees. Since the start of surveillance, an increasing proportion of South African adolescents are using AODs. Surveys point to high levels of alcohol misuse among high school students, with alcohol being the most common substance of abuse. Cannabis is the most frequently reported illicit drug of abuse among adolescents. This is reflected in the large proportion of adolescents receiving treatment for cannabis, cannabis-positive arrestees, and cannabis-positive trauma patients. Cannabis smoked together with methaqualone is the second most common primary drug of abuse in Cape Town. Arrestee data highlights the potentially negative effect of adolescent methaqualone use. Cocaine and heroin are emerging as problem drugs of abuse among adolescents in large metropolitan centres. Ecstasy (MDMA) use occurs mainly among adolescents who attend rave parties and clubs. The study points to the need for AOD intervention programmes that target young people and the need for continued monitoring of adolescent AOD use in the future. PMID- 15288753 TI - Challenging times: a study to detect Irish adolescents at risk of psychiatric disorders and suicidal ideation. AB - Suicide rates in young Irish males have risen markedly in the past 10 years, and suicide is now the leading cause of death in young men in the 15-24-year-old age range. This is the first large-scale study in Ireland that set out to identify young people at risk of psychiatric disorders, including depressive disorders, and suicidal ideation. Seven hundred and twenty three school-going adolescents aged 12-15 years were screened using the Children's Depression Inventory and the Strengths and Difficulties Questionnaire. 19.4% were identified as being 'at risk' of having a mental health disorder. Of this 'at risk' group, 12.1% expressed possible suicidal intent and 45.7% expressed suicidal ideation. Of the 583 adolescents identified as being 'not at risk', 13% expressed suicidal ideation while none expressed suicidal intent. Being 'at risk' was not related to gender or to socio-economic status. Those living with two parents were significantly more likely to be in the 'not at risk' group. Girls attending co educational schools were twice as likely to be in 'at risk' group compared with those attending all girls schools, while school type was not a risk factor for boys. This study shows that, as in other western countries, there are large numbers of young Irish people at risk of mental health disorders and suicidal ideation in the community, and raises the question of the importance of mental health promotion in our education system. PMID- 15288754 TI - The role of parental and peer support in adolescents well-being: a comparison of adolescents with and without a visual impairment. AB - In the present study we examined the importance of parental and peer support for well-being of adolescents with and without a visual impairment. The sample included 178 adolescents who are blind or visually impaired and 338 adolescents without visual impairments. Peer and parental support proved to be important for well-being of both adolescents with a visual impairment and sighted adolescents. Whereas in the group of adolescents with a visual impairment, a positive linear relationship exists between peer support and well-being, in the group of adolescents without an impairment well-being appears not be affected by peer support. Parental support is more strongly related to well-being of adolescents without impairments than of adolescents who are blind or visually impaired. PMID- 15288755 TI - Adolescent lottery and scratchcard players: do their attitudes influence their gambling behaviour? AB - This paper examines the link between attitudes and behaviour in relation to adolescent participation on the National Lottery and scratchcards by applying the theory of planned behaviour. A questionnaire constructed by the authors was administered to a sample of 1195 adolescents between the ages of 11 and 15 years (550 male, 641 female, 4 unspecified). In this paper, all data relating to factors associated with adolescent attitudes toward the National Lottery and scratchcards, and the links with any subsequent gambling behaviour, were analysed. Results revealed that young people's attitudes are an accurate predictor of their gambling behaviour on these activities, and that social cognitive theory provides an explanation of how these attitudes may develop. In light of these findings, a number of suggestions are given on how to discourage young people from gambling. PMID- 15288756 TI - Construct validity of two identity status measures: the EIPQ and the EOM-EIS-II. AB - The present study was designed to examine construct validity of two identity status measures, the Ego Identity Process Questionnaire (EIPQ; J. Adolescence 18 (1995) 179) and the Extended Objective Measure of Ego Identity Status II (EOM-EIS II; J. Adolescent Res. 1 (1986) 183). Construct validity was operationalized in terms of how identity status classifications generated by the EIPQ and EOM-EIS related to a measure of psychological agency, the Multi-Measure Agentic Personality Scale (MAPS; J. Adolescence 20 (1997) 421). Results indicated that, for the most part, the agentic personality scales differentiated the EIPQ diffused and moratorium statuses from the foreclosed and achieved statuses, whereas the agentic personality scales differentiated the EOM-EIS-II achieved status from the other three statuses. The EIPQ is recommended as the preferred instrument for making contrasts between or among identity statuses, whereas the EOM-EIS-II is recommended when the objective is to consider continuous measures of the identity statuses. PMID- 15288757 TI - Parents' life events and substance use among German adolescent girls: testing a family mediation model. AB - In this cross-sectional study, data from 202 German adolescent girls (ages 13-17 years) and their parents were utilized to test whether the relationship between parents' stressful life events and their daughter's substance use was mediated by higher levels of parents' depressive symptoms, lower quality of the relationship between the parents, and poor parenting behaviour. Structural equation modelling revealed that these family characteristics did not account for the link between parents' stressful life events and adolescent girl's substance use. Further research is needed to better understand the mechanisms that account for such linkages. PMID- 15288758 TI - Ligand-gated ion channels: mechanisms underlying ion selectivity. AB - Anion/cation selectivity is a critical property of ion channels and underpins their physiological function. Recently, there have been numerous mutagenesis studies, which have mapped sites within the ion channel-forming segments of ligand-gated ion channels that are determinants of the ion selectivity. Site directed mutations to specific amino acids within or flanking the M2 transmembrane segments of the anion-selective glycine, GABA(A) and GABA(C) receptors and the cation-selective nicotinic acetylcholine and serotonin (type 3) receptors have revealed discrete, equivalent regions within the ion channel that form the principal selectivity filter, leading to plausible molecular mechanisms and mathematical models to describe how ions preferentially permeate these channels. In particular, the dominant factor determining anion/cation selectivity seems to be the sign and exposure of charged amino acids lining the selectivity filter region of the open channel. In addition, the minimum pore diameter, which can be influenced by the presence of a local proline residue, also makes a contribution to such ion selectivity in LGICs with smaller diameters increasing anion/cation selectivity and larger ones decreasing it. PMID- 15288759 TI - HERG binding specificity and binding site structure: evidence from a fragment based evolutionary computing SAR study. AB - We describe the application of genetic programming, an evolutionary computing method, to predicting whether small molecules will block the HERG cardiac potassium channel. Models based on a molecular fragment-based descriptor set achieve an accuracy of 85-90% in predicting whether the IC(50) of a 'blind' set of compounds is <1 microM. Analysis of the models provides a 'meta-SAR', which predicts a pharmacophore of two hydrophobic features, one preferably aromatic and one preferably nitrogen-containing, with a protonatable nitrogen asymmetrically situated between them. Our experience of the approach suggests that it is robust, and requires limited scientist input to generate valuable predictive results and structural understanding of the target. PMID- 15288760 TI - Inter-residue interactions in protein folding and stability. AB - During the process of protein folding, the amino acid residues along the polypeptide chain interact with each other in a cooperative manner to form the stable native structure. The knowledge about inter-residue interactions in protein structures is very helpful to understand the mechanism of protein folding and stability. In this review, we introduce the classification of inter-residue interactions into short, medium and long range based on a simple geometric approach. The features of these interactions in different structural classes of globular and membrane proteins, and in various folds have been delineated. The development of contact potentials and the application of inter-residue contacts for predicting the structural class and secondary structures of globular proteins, solvent accessibility, fold recognition and ab initio tertiary structure prediction have been evaluated. Further, the relationship between inter residue contacts and protein-folding rates has been highlighted. Moreover, the importance of inter-residue interactions in protein-folding kinetics and for understanding the stability of proteins has been discussed. In essence, the information gained from the studies on inter-residue interactions provides valuable insights for understanding protein folding and de novo protein design. PMID- 15288761 TI - Glutamate-induced deregulation of calcium homeostasis and mitochondrial dysfunction in mammalian central neurones. AB - Delayed neuronal death following prolonged (10-15 min) stimulation of Glu receptors is known to depend on sustained elevation of cytosolic Ca(2+) concentration ([Ca(2+)](i)) which may persist far beyond the termination of Glu exposure. Mitochondrial depolarization (MD) plays a central role in this Ca(2+) deregulation: it inhibits the uniporter-mediated Ca(2+) uptake and reverses ATP synthetase which enhances greatly ATP consumption during Glu exposure. MD-induced inhibition of Ca(2+) uptake in the face of continued Ca(2+) influx through Glu activated channels leads to a secondary increase of [Ca(2+)](i) which, in its turn, enhances MD and thus [Ca(2+)](i). Antioxidants fail to suppress this pathological regenerative process which indicates that reactive oxygen species are not involved in its development. In mature nerve cells (>11 DIV), the post glutamate [Ca(2+)](i) plateau associated with profound MD usually appears after 10-15 min Glu (100 microM) exposure. In contrast, in young cells (<9 DIV) delayed Ca(2+) deregulation (DCD) occurs only after 30-60 min Glu exposure. This difference is apparently determined by a dramatic increase in the susceptibility of mitochondia to Ca(2+) overload during nerve cells maturation. The exact mechanisms of Glu-induced profound MD and its coupling with the impairment of Ca(2+) extrusion following toxic Glu challenge is not clarified yet. Their elucidation demands a study of dynamic changes in local concentrations of ATP, Ca(2+), H(+), Na(+) and protein kinase C using novel methodological approaches. PMID- 15288762 TI - Proceedings of the Third International Meeting on Rapid Responses to Steroid Hormones. September 12-14, 2003. Florence, Italy. PMID- 15288763 TI - Rapid signalling pathway activation by androgens in epithelial and stromal cells. AB - Estradiol rapidly activates Src as well as the Src-dependent pathway in human mammary cancer-derived MCF-7 cells, in human prostate cancer-derived LNCaP cells and in Cos cells transiently expressing hERs [EMBO J. 15 (1996) 1292; EMBO J. 17 (1998) 2008]. In addition, estradiol immediately stimulates, yes, an ubiquitous member of the Src kinase family, in human colon carcinoma-derived Caco-2 cells [Cancer Res. 56 (1996) 4516]. Progestins and androgens activate the same pathway in human mammary and prostate cancer-derived cells [EMBO J. 17 (1998) 2008; EMBO J. 19 (2000) 5406]. We observed that estradiol also stimulates the phosphatidylinositol-3-kinase (PI3K)/AKT pathway in MCF-7 cells [EMBO J. 20 (2001) 6050]. In these cells, activation of the Src- and the PI3 K-dependent pathways is simultaneous and mediated by direct interactions of the two kinases with ERalpha. The signalling pathway activation by sex-steroid hormones leads to DNA synthesis and cell growth in human mammary and prostate cancer-derived cells [EMBO J. 19 (2000) 5406; EMBO J. 20 (2001) 6050; EMBO J. 18 (1999) 2500]. Furthermore, androgen stimulation of NIH3T3 fibroblasts activates the same pathways triggered by this hormone in LNCaP cells and promotes the S-phase entry or cytoskeleton changes in these cells [J. Cell Biol. 161 (2003) 547]. All the described effects are rapid and require classic steroid receptors, but, surprisingly, not their transcriptional activity. Indeed, a transcriptionally inactive mutant of hER mediates the estrogen-stimulated DNA synthesis of NIH3T3 fibroblasts [EMBO J. 18 (1999) 2500]. Furthermore, AR in NIH3T3 cells does not enter nuclei and is unable to respond to the hormone with transcription stimulation, whereas it activates signaling pathways and triggers important biological responses. Signaling pathway activation by steroids has also been described by other groups under different experimental conditions and/or in different cell types. In these cells, steroid stimulation triggers various effects, such as neuroprotection, vasorelaxation or bone protection [J. Neurosci. Res. 60 (2000) 321; Nature 407 (2000) 538; J. Cell Biochem. 76 (1999) 206]. Analysis of the mechanisms responsible for the hormone-dependent and steroid receptor-mediated pathway activation in epithelial as well as stromal cells reveals immediate association of steroid receptors with extranuclear signaling effectors [EMBO J. 17 (1998) 2008; Cancer Res. 56 (1996) 4516; EMBO J. 19 (2000) 5406; EMBO J. 20 (2001) 6050; J. Cell Biol. 161 (2003) 547]. These results further highlight the central role of the hormone-regulated protein-protein interactions in the steroid action. They also offer the possibility of interfering with important activities of hormones, such as proliferation or survival, cytoskeleton changes as well as invasiveness and vasorelaxation, without affecting the steroid effects that depend on receptor transcriptional activity. PMID- 15288764 TI - The role of adapter protein Shc in estrogen non-genomic action. AB - Breast cancer is one of the most common malignancies in the United States. Seventy percent of breast cancers are hormone-responsive due to the presence of estrogen receptors ERalpha and ERbeta, which are important diagnostic and therapeutic targets in cancer treatment. Estrogen acts through its receptors, which reside on the cell membrane as demonstrated recently and in the nucleus, leading to cancer cell proliferation and protection from cell death. The membrane ERalpha has been reported in MCF-7 human breast cancer cells and is believed to mediate estrogen effects to activate mitogen-activated protein (MAP) kinase and phosphoinositide 3-kinase (PI3-kinase). Activation of many growth factor receptors require adapter proteins to delivery the upstream signals to downstream kinases, such as MAP kinase. Both Shc and the p85alpha subunit of PI3-kinase are adapter proteins. In addition to their roles in transducing signals from membrane growth factor receptors, they have been demonstrated to interact with ERalpha in an estrogen dependent manner. In this review, the role of Shc in mediating estrogen effects on MAP Kinase regulation, cell growth and anti-apoptosis will be discussed. The possible role of PI3-kinase in estrogen rapid action is also reviewed in brief. PMID- 15288765 TI - Estrogen and xenoestrogen actions on endocrine pancreas: from ion channel modulation to activation of nuclear function. AB - 17beta-Estradiol elicits a rapid opposite effect on [Ca2+]i in alpha- and beta cells within intact islets of Langerhans. In beta-cells, physiological concentrations of the gonadal hormone decreases KATP channel activity in synergy with glucose, leading to a membrane depolarization that opens voltage-gated Ca2+ channels, potentiating Ca2+ signals. As a consequence insulin release is enhanced and transcription factor CREB is activated in a Ca(2+)-dependent manner. In glucagon-containing alpha-cells, 17beta-estradiol provokes the abolishment of Ca2+ oscillations generated by low glucose, a situation that should decrease glucagon release. In both types of cells the second messenger involved is cGMP. The estrogen receptor involved is located in the plasma membrane and has a pharmacological profile unrelated to classical estrogen receptors ERalpha and ERbeta. For that reason, it has been named non-classical membrane estrogen receptor (ncmER). Although the physiological roles of this receptor are still unknown, it may be implicated in the responses of the endocrine pancreas to the physiological and pathological changes of 17beta-estradiol. PMID- 15288766 TI - Genomic and non-genomic effects of estrogens on endothelial cells. AB - Estrogen receptors act via the regulation of transcriptional processes, involving nuclear translocation and binding on specific response elements, thus leading to regulation of target gene expression. However, novel non-transcriptional mechanisms of signal transduction through steroid hormone receptors have been identified. These so-called "non-genomic" effects are independent by gene transcription or protein synthesis and involve steroid-induced modulation of cytoplasmic or of cell membrane-bound regulatory proteins. Relevant biological actions of steroids have been associated with this signaling in different tissues. Ubiquitary regulatory cascades such as mitogen-activated protein kinases (MAPK), the phosphatidylinositol 3-OH kinase (PI3K) and tyrosine kinases are modulated through non-transcriptional mechanisms by steroid hormones. Furthermore, steroid hormone receptors modulation of cell membrane-associated molecules such as ion channels and G-protein-coupled receptors has been shown in diverse tissues. The vascular wall is a site where non-genomic steroid hormones actions are particularly prominent. For instance, estrogens and glucocorticoids trigger rapid vasodilatation due to rapid induction of nitric oxide synthesis in endothelial cells via the estrogen receptor-dependent activation of MAPK and PI3K, leading to relevant pathophysiological consequences, in vitro and in vivo. The growing amount of evidence collected in the last years claims that non transcriptional signaling mechanisms play a primary role in the generation of the effects of steroids on endothelial cells, which may turn out to be of relevance for clinical purposes. PMID- 15288767 TI - Integrating rapid responses to 1,25-dihydroxyvitamin D3 with transcriptional changes in osteoblasts: Ca2+ regulated pathways to the nucleus. AB - It is well established that 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) treatment of target cells including osteoblasts activates both membrane-initiated rapid Ca2+ responses linked to influx through voltage sensitive Ca2+ channels (VSCCs) and longer term nuclear receptor-mediated changes in gene expression. We recently reported use of a cDNA microarray strategy to identify transcriptional changes after 3 and 24h of treatment with 1,25(OH)2D3 and with an analog of 1,25(OH)2D3 (25(OH)-16ene-23yne-D3 [AT]) that activates Ca2+ influx without binding to the nuclear receptor. Among 5000 different clones on the array filters, we identified families of genes in osteoblasts that were altered two-fold or greater following treatment with 1,25(OH)2D3 or analog AT for 3h. Cluster analysis further revealed complex patterns of changes in gene expression, indicative of multiple pathways to the nucleus. Evidenced by changes in target gene expression, activation of a Ca2+/CaMK/CREB/CRE pathway clearly occurs and modulates expression of a variety of genes associated with changes in protein secretion including those involved in paracrine regulation of bone resorption, RANKL and osteoprotegerin (OPG). The changes in gene expression can be inhibited by L-type VSCC channel blockers, confirming the role of Ca2+ entry in pathway activation. These findings provide clear evidence of rapid changes in gene expression associated with Ca2+ influx after treatment with 1,25(OH)2D3, and open the door to novel nuclear receptor independent signaling pathways that affect gene transcription. PMID- 15288768 TI - The androgen receptor associates with the epidermal growth factor receptor in androgen-sensitive prostate cancer cells. AB - Many recent evidences indicate that androgen-sensitive prostate cancer cells have a lower malignant phenotype that is in particular characterized by a reduced migration and invasion. We previously demonstrated that expression of androgen receptor (AR) by transfection of the androgen-independent prostate cancer cell line PC3 decreases invasion and adhesion of these cells (PC3-AR) through modulation of alpha6beta4 integrin expression. The treatment with the synthetic androgen R1881 further reduced invasion of the cells without, however, modifying alpha6beta4 expression on the cell surface, suggesting an interference with the invasion process in response to EGF. We investigated whether the presence of the AR could affect EGF receptor (EGFR)-mediated signaling in response to EGF by evaluating autotransphosphorylation of the receptor as well as activation of downstream signalling pathways. Immunoprecipitation studies demonstrated a reduction of EGF-induced tyrosine phosphorylation of EGFR in PC3-AR cells. In addition, EGF-stimulated PI3K activity, a key signalling pathway for invasion of these cells, was decreased in PC3-AR cells and further reduced by treatment with R1881, indicating decreased functionality of EGFR. An interaction between EGFR and AR has been demonstrated by immunoconfocal and co-immunoprecipitation analysis in PC3-AR cells, suggesting a possible interference of AR on EGFR signalling by interaction of the two proteins. In conclusion, our results suggest that the expression of AR by transfection in PC3 cells confers a less malignant phenotype by interfering with EGFR autophosphorylation and signalling in response to EGF leading to invasion through a mechanism involving an interaction between AR and EGFR. PMID- 15288769 TI - Human spermatozoa as a model for studying membrane receptors mediating rapid nongenomic effects of progesterone and estrogens. AB - In the past few years, besides the classical genomic effects of steroid hormones, a plethora of so called rapid non genomic effects have been described in different cell types, which are too rapid to be due to activation of gene expression. Although some of these effects might involve the same nuclear steroid receptors acting on different cellular signalling, others have been ascribed to poorly characterized membrane receptors. Several rapid nongenomic effects of progesterone (P) and estrogens (E) have been recently demonstrated in human spermatozoa. They seem to be mediated by the steroid binding to specific receptors on plasma membrane different from the classical ones. In particular, P has been demonstrated to stimulate calcium influx, tyrosine phosphorylation of sperm proteins, including extracellular signaling regulated kinases, chloride efflux and cAMP increase, finally resulting in activation of spermatozoa through induction of capacitation, hyperactivated motility and acrosome reaction. Conversely, E, by acting rapidly on calcium influx and on protein tyrosine phosphorylation, seem to modulate sperm responsiveness to P. Several attempts have been used to characterize the putative membrane receptors for P (mPR) and E (mER) in spermatozoa, however their isolation still remains elusive. However, in the past few years our laboratory has obtained several evidences supporting the existence and functional activity of mPR and mER in human spermatozoa. To characterize these membrane receptors, we used two antibodies directed against the ligand binding domains of the classical receptors, namely c262 and H222 antibodies for PR and ER respectively, hypothesizing that these regions should be conserved between nongenomic and genomic receptors. In western blot analysis of sperm lysates the antibodies detected a band of about 57 kDa for PR and of 29 kDa for ER, excluding the presence of the classical receptors. On live human spermatozoa, both antibodies were able to block the calcium and AR response to P and E respectively, whereas, antibodies directed against different domains of the classical PR and ER were ineffective. Moreover, c262 antibody also blocks in vitro human sperm penetration of hamster oocytes. Taken together all these data strongly support the existence of mPR and mER different from the classical ones, mediating rapid effects of these steroid hormones in human spermatozoa. PMID- 15288770 TI - Electrical responses to 1alpha,25(OH)2-Vitamin D3 and their physiological significance in osteoblasts. AB - Osteoblasts are a main target for the steroid 1alpha,25(OH)2-Vitamin D3 (1,25D3), where a major outcome is the modulation of the bone remodeling process. 1,25D3 deficiency leads to clinical disorders such as osteomalacia and osteoporosis, characterized by a state of insufficiently calcified tissue and bone loss, respectively. In the osteoblast nucleus, 1,25D3 modulates gene transcription for the synthesis of bone matrix proteins via the Vitamin D receptor (VDR). At the plasma membrane level, 1,25D3 potentiates ion channel functions, activates signal transduction pathways, and increases cytoplasmic calcium concentrations. So far, no clear physiological significance has been attributed to membrane-initiated 1,25D3 actions in single cells. To investigate if (a) 1,25D3 is a modulatory agent of secretion in osteoblasts and (b) the classical VDR is involved in rapid electrical events in the cell membrane, we studied hormone effects on ion channel activities in relation to exocytosis in osteoblasts isolated from VDR knockout (KO) and wild-type (WT) mice. This paper is a retrospect of the electrophysiological studies done in our laboratory to date. We found that 1,25D3 promoted ion channel responses are coupled to secretion in calvarial osteoblasts, and develop only in the presence of a functional nuclear steroid VDR. This 1,25D3 regulated exocytosis in osteoblasts, which takes place within minutes of hormone application, seems to be the natural complement of genomic actions that evolve at a longer time scale. The absence of both 1,25D3 membrane and nuclear effects in VDR KO osteoblasts may explain bone abnormalities typically found in VDR KO mice. PMID- 15288771 TI - Multiple rapid progestin actions and progestin membrane receptor subtypes in fish. AB - Progestin hormones exert rapid, nongenomic actions on a variety of target tissues in fish. The induction of oocyte maturation and the progestin membrane receptor (mPR) that mediates this action of progestins have been well characterized in fishes. Progestins also act on Atlantic croaker spermatozoa via an mPR to rapidly increase sperm motility. Preliminary results indicate that progestins can also exert rapid actions in the preoptic anterior hypothalamus (POAH) in this species to down-regulate gonadotropin-releasing hormone (GnRH) secretion. Recently, we reported the cloning, sequencing and characterization of a novel cDNA in a closely related species, spotted seatrout, that has the characteristics of the mPR involved in the progestin induction of oocyte maturation. Three distinct mPR subtypes, named alpha, beta, and gamma, have been identified in both fishes and mammals. The tissue distribution of the mPRalpha protein in seatrout suggests the alpha-subtype mediates progestin actions on GnRH secretion, sperm motility and oocyte maturation. However, mPRbeta antisense experiments in zebrafish oocytes suggest the beta-subtype also participates in the control of oocyte maturation in zebrafish. PMID- 15288772 TI - Rapid effects of aldosterone on vascular cells: clinical implications. AB - Aldosterone has attracted considerable interest as an independent cardiovascular risk marker, which has been demonstrated in a number of studies. Furthermore, recent studies revealed the prevalence of hyperaldosteronism to be about tenfold higher than previously assumed, which underlines its clinical importance. Aldosterone affects virtually any part of the cardiovascular system, namely cardiac fibroblasts and myocytes, and vascular endothelial and smooth muscle cells. In the latter cells, our laboratory has demonstrated a variety of rapid effects of the steroid, e.g. on intracellular calcium, inositol trisphosphate, and cAMP. There is also evidence for a modulation of genomic events by rapid aldosterone effects that occur via phosphorylation of transcription factors such as CREB. Furthermore, rapid tyrosine phosphorylation has been observed in vascular cells. The majority of rapid responses reported to date are insensitive towards the classic mineralocorticoid receptor (MR) antagonist, spironolactone. The in vitro experiments are complemented by a series of clinical studies in healthy volunteers, which could demonstrate rapid modulation of cardiovascular parameters after aldosterone administration, e.g. of systemic vascular resistance. In addition, an interaction of aldosterone with the adrenergic system has been observed. Most recently, rapid aldosterone induced contraction of resistance arteries has been reported. In general, the rapid in vivo effects of aldosterone are likely to participate in the pathogenesis of cardiovascular disorders. As many rapid and thus nonclassic aldosterone responses cannot be blocked by spironolactone, further research is required in order to provide adequate inhibitors to interfere with these pathways. PMID- 15288773 TI - Rapid actions of progesterone on granulosa cells. AB - Ovarian granulosa cells are responsive to progesterone but do not express the nuclear progesterone receptor. In an attempt to identify a receptor for progesterone (P4) in granulosa cells (GCs), an antibody built against the ligand binding site of the P4 receptor (i.e. C-262) was used. This antibody detected a 60 kDa protein in GCs as well as spontaneously immortalized granulosa cells (SIGCs). This C-262 detectable protein localizes to the plasma membrane and binds P4. Importantly, this C-262 detectable protein appears to be involved in mediating P4's biological actions. This is based on the findings that C-262 1) blocks P4's ability to inhibit mitogen-induced mitosis and apoptosis and 2) FITC BSA-conjugated P4 binding to granulosa cells. A C-262 detectable protein was isolated using a C-262 affinity column and sequenced. This analysis identified an unnamed protein referred to as RDA288 that was found in the rat genome (Accession number: XM216160). A nearly identical unnamed protein was found in a cDNA library of mouse lung (Accession number: AK004678). Whether RDA288 functions as a membrane receptor for P4 remains to be determined. PMID- 15288774 TI - Rapid effects of androgens in macrophages. AB - We investigated the existence of membrane receptors for testosterone (mAR) in mouse macrophages of the cell lines IC-21 and RAW 264.7 as well as their roles in nongenomic pathways, gene expression and cell functioning. Both cell lines lack intracellular androgen receptors (iARs) and respond to testosterone with rapid rises in [Ca2+]i. These rises in [Ca2+]i can neither be inhibited by iAR- nor by iER blockers, but are rather mediated through mAR. Pharmacological approaches suggest that the mAR belongs to the class of membrane receptors which are coupled to phospholipase C via pertussis toxin (PTX) sensitive G-proteins. The mAR can be localized as specific surface binding sites for testosterone-BSA-FITC by confocal laser scanning microscopy (CLSM)and flow cytometry, and are characterized by their agonist-sequestrability. In order to examine a possible role of the testosterone-induced rise in [Ca2+]i on gene expression, a c-fos promoter reporter gene construct was transfected into RAW 264.7 macrophages. The increase in [Ca2+]i induced by testosterone cannot significantly activate the c-fos promoter directly. Also, no significant activation of ERK1/2, JNK/SAPK and p38 can be observed following testosterone-stimulation alone. However, testosterone induced rises in [Ca2+]i do have specific effects on gene expression in context with lipopolysaccharide (LPS)-induced genotropic signaling: testosterone specifically down-regulates LPS-induced activation of c-fos promoter, p38 MAPK and NO production. In fetal calf serum (FCS)-induced genotropic signaling, the situation is reversed, i.e. testosterone augments the activation of c-fos promoter and ERK1/2. Our studies demonstrate a cross-talk between the testosterone-induced nongenomic Ca2+ signaling and the genotropic signaling induced by LPS and FCS in macrophages. PMID- 15288775 TI - Rapid vitamin D-dependent PKC signaling shares features with estrogen-dependent PKC signaling in cartilage and bone. AB - Our work is based on the hypothesis that steroid hormones regulate cells through traditional cytoplasmic and nuclear receptor-mediated mechanisms, as well as by rapid effects that are mediated by membrane-associated pathways. We have used the rat costochondral growth plate chondrocyte culture model to study the signaling mechanisms used by steroid hormones to elicit rapid responses and to modulate gene expression in target cells. Our studies show that the secosteroids 1,25 dihydroxyvitamin D3 [1alpha,25(OH)2D3] and 24,25-dihydroxyvitamin D3 [24R,25(OH)2D3], and the steroid hormone 17beta-estradiol, cause rapid increases in protein kinase C alpha (PKCalpha) activity, and many of the physiological responses of the cells to these regulators are PKC-dependent. Target cell specificity and the mechanisms by which PKCalpha is activated vary with each hormone. PKC activation initiates a signaling cascade that results in activation of the ERK1/2 family of mitogen activated protein kinases (MAPK), providing an alternate method for the steroids to modulate gene expression other than by traditional steroid hormone receptor-mediated pathways. In addition to their effects on growth plate chondrocytes, steroid hormones secreted by the cells also control events in the extracellular matrix through direct non-genomic regulation of matrix vesicles. PMID- 15288776 TI - In vivo selection of combinatorial libraries and designed affinity maturation of polydactyl zinc finger transcription factors for ICAM-1 provides new insights into gene regulation. AB - Zinc finger DNA-binding domains can be combined to create new proteins of desired DNA-binding specificity. By shuffling our repertoire of modified zinc finger domains to create randomly generated polydactyl zinc finger proteins with transcriptional regulatory domains, we developed large combinatorial libraries of zinc finger transcription factors (TFZFs). Millions of TFZFs can then be simultaneously screened in mammalian cells. Here, we successfully isolated specific TFZFs that significantly positively and negatively modulate the transcription of the ICAM-1 gene in primary and cancer cells, which are relevant to ICAM-1 biology and tumor development. We show that TFZFs can work in a general and in a cell-type specific manner depending on the regulatory domain and the zinc finger protein. We show that a TFZF that interacts directly with the ICAM-1 promoter at an overlapping NF-kappaB binding enhancer can overcome or synergistically cooperate with NF-kappaB induction of ICAM-1. For this TFZF, rational design was used to optimize the binding of the zinc finger protein to its DNA element and the resulting TFZF demonstrated a direct correlation between increased affinity and efficiency of target gene regulation. Thus, combining library and affinity maturation approaches generated superior TFZFs that may find further applications in therapeutic research and in ICAM-1 biology, and also provided novel mechanistic insights into the biology of transcription factors. Transcription factor libraries provide genome-wide approaches that can be applied towards the development of TFZFs specific for virtually any gene or desired phenotype and may lead to the discovery of new genetic functions and pathways. PMID- 15288777 TI - Activation mechanisms of transcriptional regulator CooA revealed by small-angle X ray scattering. AB - CooA, a heme-containing transcriptional activator, binds CO to the heme moiety and then undergoes a structural change that promotes the specific binding to the target DNA. To elucidate the activation mechanism coupled to CO binding, we investigated the CO-dependent structural transition of CooA with small-angle X ray scattering (SAXS). In the absence of CO, the radius of gyration Rg and the second virial coefficient (A2) were 25.3(+/-0.5)A and -0.39(+/-0.25) x 10(-4)ml mol g(-2), respectively. CO binding caused a slight increase in Rg (by 0.5A) and a marked decrease in A2 (by 5.09 x 10(-4)ml mol g(-2)). The observed decrease in A2 points to higher attractive interactions between CO-bound CooA molecules in solution compared with CO-free CooA. Although the minor alternation of Rg rules out changes in the overall structure, the marked change in the surface properties points to a CO-induced conformational transition. The experimental Rg and SAXS curves of the two states did not agree with the crystal structure of CO-free CooA. We thus simulated the solution structures of CooA based on the experimental data using rigid-body refinements as well as low-resolution model reconstructions. Both results demonstrate that the hinge region connecting the N terminal heme domain and C-terminal DNA-binding domain is kinked in CO-free CooA, so that the two domains are positioned close to each other. The CO-dependent structural change observed by SAXS corresponds to a slight swing of the DNA binding domains away from the heme domains coupled with their rotation by about 8 degrees around the axis of 2-fold symmetry. PMID- 15288778 TI - Different expression strategy: multiple intronic gene clusters of box H/ACA snoRNA in Drosophila melanogaster. AB - The high degree of rRNA pseudouridylation in Drosophila melanogaster provides a good model for studying the genomic organization, structural and functional diversity of box H/ACA small nucleolar RNAs (snoRNAs). Accounting for both conserved sequence motifs and secondary structures, we have developed a computer assisted method for box H/ACA snoRNA searching. Ten snoRNA clusters containing 42 box H/ACA snoRNAs were identified from D.melanogaster. Strikingly, they are located in the introns of eight protein-coding genes. In contrast to the mode of one snoRNA per intron so far observed in all animals, our results demonstrate for the first time a novel polycistronic organization that implies a different expression strategy for a box H/ACA snoRNA gene when compared to box C/D snoRNAs in D.melanogaster. Mutiple isoforms of the box H/ACA snoRNAs, from which most clusters are made up, were observed in D.melanogaster. The degree of sequence similarity between the isoforms varies from 99% to 70%, implying duplication events in different periods and a trend of enlarging the intronic snoRNA clusters. The variation in the functional elements of the isoforms could lead to partial alternation of snoRNA's function in loss or gain of rRNA complementary sequences and probably contributes to the great diversity of rRNA pseudouridylation in D.melanogaster. PMID- 15288779 TI - Deprotonation stimulates productive folding in allosteric TRAP hammerhead ribozymes. AB - Hammerhead ribozymes in crystals change conformation in response to deprotonation of the nucleophilic 2' OH, thereby aligning the hydroxyl for in-line displacement at the scissile phosphate. Published data do not address whether deprotonation affects folding in solution. Allosteric hammerhead "TRAPs," when activated by the appropriate oligonucleotide, show the expected log-linear relation between initial cleavage rate and pH. In contrast, attenuated TRAPs shows biphasic kinetics in which a rapid burst is followed by slow cleavage that is nearly independent of pH. Attenuated ribozymes are stimulated by urea at both low and high pH, confirming that rearrangement of secondary structure is rate-limiting for the attenuated ribozymes once they have folded. Plots of burst magnitude versus pH in the absence of urea show a sharp transition around pH 8.3, which is near the kinetic pKa for the cleavage reaction in Mg2+. Raising the pH after folding at pH 7.5 did not activate attenuated ribozymes even when the RNA was incubated at the elevated pH for extended periods prior to addition of Mg2+. In contrast, lowering the pH after folding at pH 9.5 rapidly re-established attenuation. Deprotonation of the ribozyme-substrate complex thus appears to alter the folding landscape such that a metastable "pre-activated" complex forms before the thermodynamically more stable attenuated state can be attained. From the initial partition into active and inactive conformers, we estimate that this deprotonation contributes approximately 1.2 kcal/mol toward stabilization of the active fold at a crucial step during folding of the TRAP. Assuming that the nucleophilic 2' OH is the relevant acid, its deprotonation would thus serve a dual role of favoring productive fold and enhancing the nucleophilicity of this oxygen. PMID- 15288781 TI - Class II photolyase in a microsporidian intracellular parasite. AB - Photoreactivation is the repair of DNA damage induced by ultraviolet light radiation using the energy contained in visible-light photons. The process is carried out by a single enzyme, photolyase, which is part of a large and ancient photolyase/cryptochrome gene family. We have characterised a photolyase gene from the microsporidian parasite, Antonospora locustae (formerly Nosema locustae) and show that it encodes a functional photoreactivating enzyme and is expressed in the infectious spore stage of the parasite's life cycle. Sequence and phylogenetic analyses show that it belongs to the class II subfamily of cyclobutane pyrimidine dimer repair enzymes. No photolyase is present in the complete genome sequence of the distantly related microsporidian, Encephalitozoon cuniculi, and this class of photolyase has never yet been described in fungi, the closest relatives of Microsporidia, raising questions about the evolutionary origin of this enzyme. This is the second environmental stress enzyme to be found in A.locustae but absent in E.cuniculi, and in the other case (catalase), the gene is derived by lateral transfer from a bacterium. It appears that A.locustae spores deal with environmental stress differently from E.cuniculi, these results lead to the prediction that they are more robust to environmental damage. PMID- 15288780 TI - Design of a highly reactive HDV ribozyme sequence uncovers facilitation of RNA folding by alternative pairings and physiological ionic strength. AB - The hepatitis delta virus (HDV) ribozyme is a self-cleaving RNA that resides in the HDV genome and regulates its replication. The native fold of the ribozyme is complex, having two pseudoknots. Earlier work implicated four non-native pairings in slowing pseudoknot formation: Alt 1, Alt 2, Alt 3, and Alt P1. The goal of the present work was design of a kinetically simplified and maximally reactive construct for in vitro mechanistic and structural studies. The initial approach chosen was site-directed mutagenesis in which known alternative pairings were destabilized while leaving the catalytic core intact. Based on prior studies, the G11C/U27Delta double mutant was prepared. However, biphasic kinetics and antisense oligonucleotide response trends opposite those of the well-studied G11C mutant were observed suggesting that new alternative pairings with multiple registers, termed Alt X and Alt Y, had been created. Enzymatic structure mapping of oligonucleotide models supported this notion. This led to a model wherein Alt 2 and the phylogenetically conserved Alt 3 act as "folding guides", facilitating folding of the major population of the RNA molecules by hindering formation of the Alt X and Alt Y registers. Attempts to eliminate the strongest of the Alt X pairings by rational design of a quadruple mutant only resulted in more complex kinetic behavior. In an effort to simultaneously destabilize multiple alternative pairings, studies were carried out on G11C/U27Delta in the presence of urea or increased monovalent ion concentration. Inclusion of physiological ionic strength allowed the goal of monophasic, fast-folding (kobs approximately 60 min(-1)) kinetics to be realized. To account for this, a model is developed wherein Na+, which destabilizes secondary and tertiary structures in the presence of Mg2+, facilitates native folding by destabilizing the multiple alternative secondary structures with a higher-order dependence. PMID- 15288782 TI - Solution structure and DNA binding of the zinc-finger domain from DNA ligase IIIalpha. AB - DNA ligase IIIalpha carries out the final ligation step in the base excision repair (BER) and single strand break repair (SSBR) mechanisms of DNA repair. The enzyme recognises single-strand nicks and other damage features in double stranded DNA, both through the catalytic domain and an N-terminal domain containing a single zinc finger. The latter is homologous to other zinc fingers that recognise damaged DNA, two in the N terminus of poly(adenosine ribose)polymerase and three in the N terminus of the Arabidopsis thaliana nick sensing DNA 3'-phosphoesterase. Here, we present the solution structure of the zinc-finger domain of human DNA ligase IIIalpha, the first structure of a finger from this group. It is related to that of the erythroid transcription factor GATA 1, but has an additional N-terminal beta-strand and C-terminal alpha-helix. Chemical shift mapping using a DNA ligand containing a single-stranded break showed that the DNA-binding surface of the DNA-ligase IIIalpha zinc finger is substantially different from that of GATA-1, consistent with the fact that the two proteins recognise very different features in the DNA. Likely implications for DNA binding are discussed. PMID- 15288783 TI - Exploring rare conformational species and ionic effects in DNA Holliday junctions using single-molecule spectroscopy. AB - The four-way DNA (Holliday) junction is an essential intermediate in DNA recombination, and its dynamic characteristics are likely to be important in its cellular processing. In our previous study we observed transitions between two antiparallel stacked conformations using a single-molecule fluorescence approach. The magnesium concentration-dependent rates of transitions between stacking conformers suggested that an unstacked open structure, which is stable in the absence of metal ions, is an intermediate. Here, we sought to detect possible rare species such as open and parallel conformations and further characterized ionic effects. The hypothesized open intermediate cannot be resolved directly due to the limited time resolution and sensitivity, but our study suggests that the open form is achieved very frequently, hundreds of times per second under physiologically relevant conditions. Therefore despite being a minority species, its frequent formation raises the probability that it could become stabilized by protein binding. By contrast, we cannot detect even a transient existence of the junctions in a parallel form, and the probability of such forms with a lifetime greater than 5 ms is less than 0.01%. Stacking conformer transitions are observable in the presence of sodium or hexammine cobalt (III) ions as well as magnesium ions, but the transition rates are higher for lower valence ions at the same concentrations. This further supports the notion that electrostatic stabilization of the stacked structures dictates the interconversion rates between different structural forms. PMID- 15288784 TI - Complete sequence and genetic organization of pDTG1, the 83 kilobase naphthalene degradation plasmid from Pseudomonas putida strain NCIB 9816-4. AB - The complete 83,042 bp sequence of the circular naphthalene degradation plasmid pDTG1 from Pseudomonas putida strain NCIB 9816-4 was determined in order to examine the process by which the nah and sal operons may have been compiled and distributed in nature. Eighty-nine open reading frames were predicted using computer analyses, comprising 80.0% of the pDTG1 DNA sequence. The most distinctive feature of the plasmid is the upper and lower naphthalene degradation operons, which occupy 9.5 kb and 13.4 kb regions, respectively, bordered by numerous defective mobile genetic element fragments. Identified on this plasmid were homologues of genes required for large plasmid replication, maintenance, and conjugation, as well as transposases, resolvases, and integrases, suggesting an evolution that involved the lateral transfer of DNA between bacterial species. Also found were genes that contain a high degree of sequence similarity to other known degradation genes, as well as genes involved in chemotaxis. Although the incompatibility group designation of pDTG1 remains unresolved, striking sequence organization and homology exists between the plasmid backbones of pDTG1 and the IncP-9 toluene-degradation plasmid pWW0, which suggests a divergent evolution from a progenitor plasmid prior to degradative gene incorporation. PMID- 15288785 TI - A highly unusual palindromic transmembrane helical hairpin formed by SARS coronavirus E protein. AB - The agent responsible for the recent severe acute respiratory syndrome (SARS) outbreak is a previously unidentified coronavirus. While there is a wealth of epidemiological studies, little if any molecular characterization of SARS coronavirus (SCoV) proteins has been carried out. Here we describe the molecular characterization of SCoV E protein, a critical component of the virus responsible for virion envelope morphogenesis. We conclusively show that SCoV E protein contains an unusually short, palindromic transmembrane helical hairpin around a previously unidentified pseudo-center of symmetry, a structural feature which seems to be unique to SCoV. The hairpin deforms lipid bilayers by way of increasing their curvature, providing for the first time a molecular explanation of E protein's pivotal role in viral budding. The molecular understanding of this critical component of SCoV may represent the beginning of a concerted effort aimed at inhibiting its function, and consequently, viral infectivity. PMID- 15288786 TI - Crystal structure of the 270 kDa homotetrameric lignin-degrading enzyme pyranose 2-oxidase. AB - Pyranose 2-oxidase (P2Ox) is a 270 kDa homotetramer localized preferentially in the hyphal periplasmic space of lignocellulolytic fungi and has a proposed role in lignocellulose degradation to produce the essential co-substrate, hydrogen peroxide, for lignin peroxidases. P2Ox oxidizes D-glucose and other aldopyranoses regioselectively at C2 to the corresponding 2-keto sugars; however, for some substrates, the enzyme also displays specificity for oxidation at C3. The crystal structure of P2Ox from Trametes multicolor has been determined using single anomalous dispersion with mercury as anomalous scatterer. The model was refined at 1.8A resolution to R and Rfree values of 0.134 and 0.171, respectively. The overall fold of the P2Ox subunit resembles that of members of the glucose methanol-choline family of long-chain oxidoreductases, featuring a flavin-binding Rossmann domain of class alpha/beta and a substrate-binding subdomain with a six stranded central beta sheet and three alpha helices. The homotetramer buries a large internal cavity of roughly 15,000 A3, from which the four active sites are accessible. Four solvent channels lead from the surface into the cavity through which substrate must enter before accessing the active site. The present structure shows an acetate molecule bound in the active site with the carboxylate group positioned immediately below the flavin N5 atom, and with one carboxylate oxygen atom interacting with the catalytic residues His548 and Asn593. The entrance to the active site is blocked by a loop (residues 452 to 461) with excellent electron density but elevated temperature factors. We predict that this loop is dynamic and opens to allow substrate entry and exit. In silico docking of D-glucose in the P2Ox active site shows that with the active-site loop in the closed conformation, monosaccharides cannot be accommodated; however, after removing the loop from the model, a tentative set of protein-substrate interactions for beta-D-glucose have been outlined. PMID- 15288787 TI - High-resolution structures reveal details of domain closure and "half-of-sites reactivity" in Escherichia coli aspartate beta-semialdehyde dehydrogenase. AB - Two high-resolution structures have been determined for Eschericia coli aspartate beta-semialdehyde dehydrogenase (ecASADH), an enzyme of the aspartate biosynthetic pathway, which is a potential target for novel antimicrobial drugs. Both ASADH structures were of the open form and were refined to 1.95 A and 1.6 A resolution, allowing a more detailed comparison with the closed form of the enzyme than previously possible. A more complex scheme for domain closure is apparent with the subunit being split into two further sub-domains with relative motions about three hinge axes. Analysis of hinge data and torsion-angle difference plots is combined to allow the proposal of a detailed structural mechanism for ecASADH domain closure. Additionally, asymmetric distortions of individual subunits are identified, which form the basis for the previously reported "half-of-the-sites reactivity" (HOSR). A putative explanation of this arrangement is also presented, suggesting the HOSR system may provide a means for ecASADH to offset the energy required to remobilise flexible loops at the end of the reaction cycle, and hence avoid falling into an energy minimum. PMID- 15288788 TI - Expression improvement and mechanistic study of the retro-Diels-Alderase catalytic antibody 10F11 by site-directed mutagenesis. AB - Antibody 10F11 catalyzes the retro-Diels-Alder reaction of the bicyclic prodrug 1 releasing HNO and anthracene 4 (kcat/kuncat=2500). Earlier X-ray crystal structures of Fab 10F11 showed that tryptophan H104 at the bottom of the binding pocket interacts by pi-stacking with the aromatic ring of the substrate. Antibody 10F11 was expressed as a chimeric Fab and subjected to site-directed mutagenesis. Expression was improved by substituting a serine for a phenylalanine residue on the Fv-domain surface. Nine active-site mutants were then prepared including replacements at TrpH104, PheH101 and SerH100. Catalysis depends mainly on TrpH104 and PheH101. Catalysis is most likely caused by a combination of shape complementarity and specific electronic interactions between transition state and the aromatic residue H104. Medium and de-solvation effects have no effect on the reaction rate. Catalysis was improved to (kcat/kuncat=6300) by substituting phenylalanine for LeuL101 to indirectly enhance pi-stacking between transition state and TrpH104. PMID- 15288789 TI - Structural Basis of allosteric regulation and substrate specificity of the non phosphorylating glyceraldehyde 3-Phosphate dehydrogenase from Thermoproteus tenax. AB - The non-phosphorylating glyceraldehyde-3-phosphate dehydrogenase (GAPN) of the hyperthermophilic Archaeum Thermoproteus tenax is a member of the superfamily of aldehyde dehydrogenases (ALDH). GAPN catalyses the irreversible oxidation of glyceraldehyde 3-phosphate (GAP) to 3-phosphoglycerate in the modified glycolytic pathway of this organism. In contrast to other members of the ALDH superfamily, GAPN from T.tenax (Tt-GAPN) is regulated by a number of intermediates and metabolites. In the NAD-dependent oxidation of GAP, glucose 1-phosphate, fructose 6-phosphate, AMP and ADP increase the affinity for the cosubstrate, whereas ATP, NADP, NADPH and NADH decrease it leaving, however, the catalytic rate virtually unaltered. As we show here, the enzyme also uses NADP as a cosubstrate, displaying, however, unusual discontinuous saturation kinetics indicating different cosubstrate affinities and/or reactivities of the four active sites of the protein tetramer caused by cooperative effects. Furthermore, in the NADP dependent reaction the presence of activators decreases the overall S0.5 and increases Vmax by a factor of 3. To explore the structural basis for the different effects of both pyridine nucleotides we solved the crystal structure of Tt-GAPN in complex with NAD at 2.2 A resolution and compared it to the binary Tt GAPN-NADPH structure. Although both pyridine nucleotides show a similar binding mode, NADPH appears to be more tightly bound to the protein via the 2' phosphate moiety. Moreover, we present four co-crystal structures with the activating molecules glucose 1-phosphate, fructose 6-phosphate, AMP and ADP determined at resolutions ranging from 2.3 A to 2.6 A. These crystal structures reveal a common regulatory site able to accommodate the different activators. A phosphate-binding pocket serves as an anchor point ensuring similar binding geometry. The observed conformational changes upon activator binding are discussed in terms of allosteric regulation. Furthermore, we present a crystal structure of Tt-GAPN in complex with the substrate D-GAP at 2.3 A resolution, which allows us to analyse the structural basis for substrate binding, the mechanism of catalysis as well as the stereoselectivity of the enzymatic reaction. PMID- 15288790 TI - The crystal structure of ZapA and its modulation of FtsZ polymerisation. AB - FtsZ is part of a mid-cell cytokinetic structure termed the Z-ring that recruits a hierarchy of fission related proteins early in the bacterial cell cycle. The widely conserved ZapA has been shown to interact with FtsZ, to drive its polymerisation and to promote FtsZ filament bundling thereby contributing to the spatio-temporal tuning of the Z-ring. Here, we show the crystal structure of ZapA (11.6 kDa) from Pseudomonas aeruginosa at 2.8 A resolution. The electron density reveals two dimers associating via an extensive C-terminal coiled-coil protrusion to form an elongated anti-parallel tetramer. In solution, ZapA exists in a dimer tetramer equilibrium that is strongly correlated with concentration. An increase in concentration promotes formation of the higher oligomeric state. The dimer is postulated to be the predominant physiological species although the tetramer could become significant if, as FtsZ is integrated into the Z-ring and is cross linked, the local concentration of the dimer becomes sufficiently high. We also show that ZapA binds FtsZ with an approximate 1:1 molar stoichiometry and that this interaction provokes dramatic FtsZ polymerisation and inter-filament association as well as yielding filaments, single or bundled, more stable and resistant to collapse. Whilst in vitro dynamics of FtsZ are well characterised, its in vivo arrangement within the ultra-structural architecture of the Z-ring is yet to be determined despite being fundamental to cell division. The ZapA dimer has single 2-fold symmetry whilst the bipolar tetramer displays triple 2-fold symmetry. Given the symmetry of these ZapA oligomers and the polar nature of FtsZ filaments, the structure of ZapA carries novel implications for the inherent architecture of the Z-ring in vivo. PMID- 15288791 TI - Monitoring the transition from the T to the R state in E.coli aspartate transcarbamoylase by X-ray crystallography: crystal structures of the E50A mutant enzyme in four distinct allosteric states. AB - A detailed description of the transition that allosteric enzymes undergo constitutes a major challenge in structural biology. We have succeeded in trapping four distinct allosteric states of a mutant enzyme of Escherichia coli aspartate transcarbomylase and determining their structures by X-ray crystallography. The mutant version of aspartate transcarbamoylase in which Glu50 in the catalytic chains was replaced by Ala destabilizes the native R state and shifts the equilibrium towards the T state. This behavior allowed the use of substrate analogs such as phosphonoacetamide and malonate to trap the enzyme in T like and R-like structures that are distinct from the T-state structure of the wild-type enzyme (as represented by the structure of the enzyme with CTP bound and the R-state structure as represented by the structure with N-(phosphonacetyl) L-aspartate bound). These structures shed light on the nature and the order of internal structural rearrangements during the transition from the T to the R state. They also suggest an explanation for diminished activity of the E50A enzyme and for the change in reaction mechanism from ordered to random for this mutant enzyme. PMID- 15288792 TI - Structure of the coat protein in Pf1 bacteriophage determined by solid-state NMR spectroscopy. AB - The atomic resolution structure of Pf1 coat protein determined by solid-state NMR spectroscopy of magnetically aligned filamentous bacteriophage particles in solution is compared to the structures previously determined by X-ray fiber and neutron diffraction, the structure of its membrane-bound form, and the structure of fd coat protein. These structural comparisons provide insights into several biological properties, differences between class I and class II filamentous bacteriophages, and the assembly process. The six N-terminal amino acid residues adopt an unusual "double hook" conformation on the outside of the bacteriophage particle. The solid-state NMR results indicate that at 30 degrees C, some of the coat protein subunits assume a single, fully structured conformation, and some have a few mobile residues that provide a break between two helical segments, in agreement with structural models from X-ray fiber and neutron diffraction, respectively. The atomic resolution structure determined by solid-state NMR for residues 7-14 and 18-46, which excludes the N-terminal double hook and the break between the helical segments, but encompasses more than 80% of the backbone including the distinct kink at residue 29, agrees with that determined by X-ray fiber diffraction with an RMSD value of 2.0 A. The symmetry and distance constraints determined by X-ray fiber and neutron diffraction enable the construction of an accurate model of the bacteriophage particle from the coordinates of the coat protein monomers. PMID- 15288793 TI - It just didn't work: the realities of quality assessment in the English health care context. AB - AIMS: Assessment of care quality is integral to health and palliative care provision and there is a need to develop and implement outcome measures to assess quality. This study aimed to: (1) describe the implementation of a palliative care outcome measure in non-specialist palliative care settings and (2) to understand the implementation of the measure. METHOD: Twenty-five non-specialist palliative care settings were purposely sampled and invited to implement a palliative care outcome measure. Fifteen settings agreed to take part. The research team provided training and support in the use of the measure. Data were collected on actual use of the measure and, via interviews with patients and nurses, on their experiences. RESULTS: The number of assessments was low (21 patients assessed against an anticipated minimum of 240). The analysis of nurses' accounts identified important considerations in understanding the low response. Although nurses saw the implementation of the outcome measure as bringing opportunities for themselves, including their own professional development, and for the organisations they represent, including raising organisational profiles, there were a number of factors that acted as impediments. These include: perceived time to administer the paperwork; competence and confidence in recruiting patients and proceeding with informed consent; concerns about the effects of completing the measure with very ill patients; and the effects of nurses raising palliative care issues on their relationships with patients. CONCLUSIONS: It is difficult to integrate outcome measures into routine clinical practice. Future interventions should consider how to tailor the implementation of outcome measures within existing working structures and provide education and training to enable nurses to deal with potentially sensitive palliative care issues. PMID- 15288794 TI - The impact of bachelor degree completion programs on career development of nurses in Israel. AB - The study compares aspects of the career development of working nurses, graduates of a BA degree program, which runs on two tracks--university (UG) and college (CG). Two hundred nurses participated in the study, 86 of them (43%) UG and 114 (57%) CG. Participants were asked about their academic backgrounds, career advancement and satisfaction with their studies. Results indicated no major differences in professional advancement between the graduates of the two tracks. College-graduates were significantly older with lower previous academic achievements. Both BA programs promoted nurses and enabled career advancement, especially so for the college-graduates who would not, otherwise, have been able to make this progress. PMID- 15288795 TI - Perceptions of nursing: confirmation, change and the student experience. AB - Research has identified a number of negative societal perceptions of nursing related to gendered stereotyping, subordination to doctors, low academic standards, limited career opportunities and poor pay and conditions, and importantly how these perceptions may affect levels of recruitment into nursing. Focusing specifically on nurses, research has also considered the extent to which these societal perceptions are realities in their workplaces, and the direct experiences that contribute to attrition from both nursing courses and jobs. However, to date, few research has actually bridged the above approaches and considered the perceptions that nursing students hold as they first enter their education and how these change, or are confirmed, as a result of their experiences. In this context, the current study uses a combined questionnaire (n = 650), interview (n = 30) and focus group (n = 7) methodology to investigate the experiences of students based at two British Universities. The findings suggest that many students were surprised, yet not overwhelmed, by the high academic standards required of them and came to recognize and value the tremendous knowledge, skills set and responsibilities of nurses as they acquired them. However, their experiences reinforced both society's and their own image of an underpaid, overworked profession that lacks respect and has low morale. The findings support media initiatives that emphasize nurses' skills in order to influence public opinion. They also support a range of subtle changes in nurse education at the institutional level to make student life easier. Nevertheless, it is acknowledged that these may have a limited impact unless pay and conditions are adequately addressed at the national level. PMID- 15288796 TI - Perceptual adjustment levels: patients' perception of their dignity in the hospital setting. AB - Dignity is an important concept which lies at the heart of nursing. Despite the statements made by various international bodies, as a concept it remains complex and under researched. This is because it needs to be understood from the patient's point of view and experience. This empirical study explores the concept in terms of the actual circumstances of patients in hospital. Through a large number of semi-structured interviews it reveals the relationship between 'normal' ideas of self-respect and self-esteem, and those experienced during the trauma of being ill and helpless, and the ways in which patients adjust to these circumstances. Through the realisation of the importance of 'necessary submission' to the power of others, the evidence creates a concept of 'perceptual adjustment level' through which we can understand the importance of matching patient needs to nursing care. This has crucial relevance to nursing. PMID- 15288797 TI - Development of the choices and acquisition of antibiotics model from a descriptive study of a lay Honduran population. AB - Antibiotic resistance is a global public health problem that is accelerated by overuse and misuse of antibiotics. In today's world of increasing international travel and exchange of goods, the spread of antibiotic resistant organisms is a growing threat. Despite significant antibiotic use in developing nations, research to describe and curtail inappropriate use is limited. In this study, the investigators developed a model of antibiotic use, choices and acquisition of antibiotics model, from a study of a lay population in Honduras. A representative sample of 939 rural and urban Hondurans completed the Preguntas Para El Uso de Antibiotics questionnaire to determine how the participant made choices about antibiotic use. The study indicated that the rural participants used significantly fewer antibiotics than the urban participants and that the demographic indicators did not show a significant difference in antibiotic use in those of lower socioeconomic status. In addition, the participants reported that they seek out professional advice and care rather than self-prescribing. Implications for educational and empowerment programs based on the model are discussed. PMID- 15288798 TI - Culture and communication in Thai nursing: a report of an ethnographic study. AB - Most nurses live and work in multicultural settings. Given the need for all nurses and health-care workers to communicate--with patients, with families and with other health-care professionals--the study of the relationship between culture and communication can help to inform practice. This paper offers the findings from an ethnographic study of culture and communication, carried out in Thailand. The aim of the study was to address the question: 'in what, if any, ways do Thai cultural issues influence interpersonal communication patterns in Thai nursing and Thai nursing education?'. Data were collected from a variety of sources, including direct and indirect observation, interviews and discussions and the literature on the topic. For the interviews, the sample was a convenience and purposive one made up of clinical nurses and nurse educators (n = 14). Those data were analysed with the aid of a computerised, qualitative data analysis program. Findings reported in this paper include those relating to 'Thainess', Buddhism, the nursing profession and nurse--patient/doctor--patient relationships. The report ends with a 'portrait' of Thai nursing communication. It is suggested that understanding the cultural aspects of nursing in various contexts can help nurses, internationally. PMID- 15288799 TI - Impact of the stage of dementia on the time required for bathing-related care: a pilot study in a Japanese nursing home. AB - Time required for bathing-related care for nursing home residents with various stages of severe dementia were observed. Time required for each resident, including guiding to the bathroom, undressing, and dressing were plotted in graphs in order to make comparisons. The situations and conversations observed for the instances when additional time was needed were analyzed. Stage of dementia affected the amount of time required for the task of guiding to the bathroom, but did not appear to affect time required for dressing or undressing. For dressing and undressing, additional time was required when caregivers failed to keep to a specific routine. PMID- 15288800 TI - The Strain of Care for Delirium Index: a new instrument to assess nurses' strain in caring for patients with delirium. AB - This study evaluated content validity, internal consistency and construct validity of the Strain of Care for Delirium Index (SCDI), a newly constructed instrument to measure the strain nurses experience in caring for patients with delirium. Content validity, evaluated by eight experts, reduced the initial pool of items from 38 to 28. Using a convenience sample of 190 nurses, Cronbach's alpha for the 28-item version was 0.88. Using non-linear principal components analysis another eight items were eliminated and a four-factor structure was identified. The proportion of variance explained by the remaining 20 items was 61.51%. Preliminary psychometric evaluation of the SCDI supported content validity, internal consistency and construct validity; however additional psychometric evaluation is warranted. PMID- 15288801 TI - The guideline contradiction: health visitors' use of formal guidelines for identifying and assessing families in need. AB - The aim of this paper is to examine health visitors' use of formal guidelines in identifying health needs and prioritizing families requiring extra health visiting support. With the increasing emphasis on targeted health visiting, a case study was used to explore the extent to which health visitors in three case sites use needs assessment guidelines in the assessment of family health need. The findings indicate how the presence of core visiting protocols hints at elements of control by managers, leading to conflicts in the relationship between professional judgements and official guidelines. Despite a management ethos of guideline formulation, several contradictions exist for which these guidelines are a focus. These include: little involvement of health visitors in guideline development, some staff not informed about the existence of formal guidelines, little evidence of guidelines contributing to improved client outcomes and their limited use by many health visitors in practice. Thus, even when guidelines exist, no accurate predictions can be made about health visitors' knowledge of or use of such guidelines in practice. PMID- 15288803 TI - Guidelines for clinical practice: development, dissemination and implementation. AB - Clinical guidelines are one of the most promising and effective advances for defining and improving the quality of care. However, their development, dissemination and implementation in practice are rarely straightforward. Within nursing practice, guidelines have the potential to ensure the clinical application of research findings, thus ensuring that the profession rejects ineffective practices while employing those shown to work. Nevertheless, the benefits and limitations of clinical guidelines should be carefully considered by practitioners, managers and consumers of health care alike. PMID- 15288802 TI - An application of the mini review to a complex methodological question: how best to research public health nursing and service quality? AB - This paper describes a mini review which enabled the identification of a suitable methodology to undertake a study about quality in the public health nursing in the Republic of Ireland. Reviews of literature increasingly adopt the methods of systematic review. In general, these methods have been developed to answer clearly focussed clinical questions. In this paper, we adopt the key elements of systematic review, comprehensive identification of relevant material and selection based on objectively defined validity, to a different type of question, that of an appropriate methodology to examine quality in the public health nursing service. In doing so, we demonstrate that questions of clinical effectiveness are but one application for systematic review. PMID- 15288804 TI - Active-site characteristics of CYP2C19 and CYP2C9 probed with hydantoin and barbiturate inhibitors. AB - Three series of N-3 alkyl substituted phenytoin, nirvanol, and barbiturate derivatives were synthesized and their inhibitor potencies were tested against recombinant CYP2C19 and CYP2C9 to probe the interaction of these ligands with the active sites of these enzymes. All compounds were found to be competitive inhibitors of both enzymes, although the degree of inhibitory potency was generally much greater towards CYP2C19. Inhibitor stereochemistry did not markedly influence K(i) towards CYP2C9, and log P adequately predicted inhibitor potency for this enzyme. In contrast, stereochemistry was an important factor in determining inhibitor potency towards CYP2C19. (S)-(+)-N-3-Benzylnirvanol and (R) (-)-N-3-benzylphenobarbital emerged as the most potent and selective CYP2C19 inhibitors, with K(i) values of < 250nM--at least two orders of magnitude greater inhibitor potency than towards CYP2C9. Both inhibitors were metabolized preferentially at their C-5 phenyl substituents, indicating that CYP2C19 prefers to orient the N-3 substituents away from the active oxygen species. These features were incorporated into expanded CoMFA models for CYP2C9, and a new, validated CoMFA model for CYP2C19. PMID- 15288805 TI - A compensatory double mutation of the alanine-86 to leucine mutant located in the hinge region of the iron-sulfur protein of the yeast cytochrome bc1 complex. AB - Mutations in the hinge region connecting the membrane anchor to the extra membranous head-group of the iron-sulfur protein can impede proper assembly and function of the cytochrome bc(1) complex. Mutating the conserved alanines, residues 86, 90, and 92, located in the hinge region resulted in a 30-50% decrease in enzymatic activity without loss of the iron-sulfur protein [J. Bioenerg. Biomembr. 31 (1999) 215]. The lowered enzymatic activity in the A86L mutant was shown to result from steric interference between the side chains of Leu-86 and Leu-89 [Biochemistry 40 (2001) 327]. The compensatory double mutant A86L/L89A restored activity to wild type levels and relieved the steric hindrance; however, the L89A mutant did not assemble properly into the bc(1) complex. Molecular modeling studies of these mutants compared to the wild type have suggested that the hydrophobic residues located in the hinge region are critical to the motion of the head group of the iron-sulfur protein during catalysis. PMID- 15288807 TI - The oxidative mechanism of action of ortho-quinone inhibitors of protein-tyrosine phosphatase alpha is mediated by hydrogen peroxide. AB - Here, we report the identification and characterization of five ortho-quinone inhibitors of PTPalpha. We observed that the potency of these compounds in biochemical assays was markedly enhanced by the presence of DTT. A kinetic analysis suggested that they were functioning as irreversible inhibitors and that the inhibition was targeted to the catalytic site of PTPalpha. The inhibition observed by these compounds was sensitive to superoxide dismutase and catalase, suggesting that reactive oxygen species may be mediators of their inhibition. We observed that in the presence of DTT, these compounds would produce up to 2.5mM hydrogen peroxide (H(2)O(2)). The levels of H(2)O(2) produced were sufficient to completely inactivate PTPalpha. In contrast, without a reducing agent the compounds did not generate H(2)O(2) and showed little activity towards PTPalpha. In addition, these compounds inhibited PTPalpha-dependent cell spreading in NIH 3T3 cells at concentrations that were similar to their activity in biochemical assays. The biological implications of these results are discussed as they support growing evidence that H(2)O(2) is a key regulator of PTPs. PMID- 15288806 TI - Nuclear localization of the DOCK180/ELMO complex. AB - DOCK180 protein plays a key role during development, cell motility, and phagocytosis. It forms a complex with another protein ELMO, and this complex acts as a guanine nucleotide exchange factor (GEF) for Rac. However, DOCK180 containing complexes have not been purified by unbiased biochemical approaches, and the nature and subcellular localization of these complexes remain unclear. Here, we show that a large fraction of endogenous DOCK180 is present as a 700kDa nuclear complex with ELMO proteins. In addition, this nuclear DOCK180/ELMO complex has functional Rac-GEF activity. Furthermore, endogenous DOCK180 could be found in complexes with different ELMO isoforms (ELMO1, 2 or 3) in different cell lines, depending on the ELMO isoforms expressed. These studies suggest that DOCK180 may associate with different ELMO proteins to form cell-type specific complexes and may have functions in both the nucleus and the cytoplasm. PMID- 15288808 TI - CyP40, but not Hsp70, in rabbit reticulocyte lysate causes the aryl hydrocarbon receptor-DNA complex formation. AB - Upon ligand binding, the aryl hydrocarbon receptor (AhR) translocates into the nucleus and dimerizes with its partner aryl hydrocarbon receptor nuclear translocator (Arnt). The AhR-Arnt heterodimer binds to the dioxin response element (DRE) to regulate target gene expression. Using baculovirus expressed human AhR and Arnt, we showed that the formation of the ligand-dependent AhR-Arnt DRE complex requires protein factors in vitro. Recently, we provided evidence that p23, an Hsp90-associated protein, is involved in the complex formation. The aim of this study was to determine whether two other Hsp90-associated proteins present in rabbit reticulocyte lysate (RRL), namely CyP40 and Hsp70, play any role in forming the AhR-Arnt-DRE complex. Fractionation and immunodepletion experiments revealed that Hsp70 is not necessary for the formation of this complex. In contrast, CyP40 is involved in forming the complex since (1) immunodepletion of CyP40 from a RRL fraction reduces the intensity of the AhR Arnt-DRE complex by 48% and (2) recombinant human CyP40 alone causes the formation of this complex. In addition, CyP40-interacting proteins appear to be essential for the full CyP40 effect on the AhR gel shift complex. PMID- 15288809 TI - Human 17beta-hydroxysteroid dehydrogenases types 1, 2, and 3 catalyze bi directional equilibrium reactions, rather than unidirectional metabolism, in HEK 293 cells. AB - Human 17beta-hydroxysteroid dehydrogenases (17betaHSDs) catalyze the interconversion of weak and potent androgen and estrogen pairs. Although the reactions using purified enzymes can be driven in either direction, these enzymes appear to function unidirectionally in intact cells: only reductive reactions for 17betaHSD1 and 17beta HSD3 and only oxidative reactions for 17betaHSD2. We show that, after exhaustive incubations with either 17beta-hydroxy- or 17-ketosteroid, the medium for HEK-293 cells expressing 17betaHSD1 or 17betaHSD3 contains a 92:8 ratio of reduced:oxidized steroid. Similarly, 17betaHSD2 yields a >95:5 ratio of oxidized:reduced steroids for both androgens and estrogens. Dual-isotope kinetic measurements show that the rates of the forward and reverse reactions are identical at these functional equilibrium states in intact cells for all three 17betaHSD isoforms, and these rates are much faster than those estimated from single-isotope flux studies. Mutation L36D converts 17betaHSD1 to an oxidative enzyme in intact cells, reversing the equilibrium distribution of estradiol:estrone to 5:95; however, the rates of the forward and reverse reactions at equilibrium are equal and comparable to those of the wild-type enzymes. The co-expression of 17betaHSD2 paradoxically increases the potency of estrone in transactivation assays, demonstrating the physiological relevance of "backwards" metabolism to estradiol. We conclude that 17betaHSD types 1, 2, and 3 catalyze both oxidative and reductive reactions in HEK-293 cells at intrinsic rates that are much faster than those estimated from single-isotope studies. These 17betaHSD isoforms do not drive steroid flux in one direction but rather may achieve functional equilibria in intact cells, reflecting thermodynamically driven steroid distributions. PMID- 15288810 TI - Mechanism-based inactivation of human leukocyte elastase via an enzyme-induced sulfonamide fragmentation process. AB - We describe herein the design and in vitro biochemical evaluation of a novel class of mechanism-based inhibitors of human leukocyte elastase (HLE) that inactivate the enzyme via an unprecedented enzyme-induced sulfonamide fragmentation cascade. The inhibitors incorporate in their structure an appropriately functionalized saccharin scaffold. Furthermore, the inactivation of the enzyme by these inhibitors was found to be time-dependent and to involve the active site. Biochemical, HPLC, and mass spectrometric studies show that the interaction of these inhibitors with HLE results in the formation of a stable acyl complex and is accompanied by the release of (L) phenylalanine methyl ester. The data are consistent with initial formation of a Michaelis-Menten complex and subsequent formation of a tetrahedral intermediate with the active site serine (Ser(195)). Collapse of the tetrahedral intermediate with tandem fragmentation results in the formation of a highly reactive conjugated sulfonyl imine which can either react with water to form a stable acyl enzyme and/or undergo a Michael addition reaction with an active site nucleophilic residue (His(57)). It is also demonstrated herein that this class of compounds can be used in the design of inhibitors of serine proteases having either a neutral or basic primary substrate specificity. Thus, the results suggest that these inhibitors constitute a potential general class of mechanism-based inhibitors of (chymo)trypsin-like serine proteases. PMID- 15288811 TI - Ascorbate in thylakoid lumen functions as an alternative electron donor to photosystem II and photosystem I. AB - The roles of ascorbate (Asc) in the thylakoid lumen to support photosynthetic electron transport are investigated. Asc can be photooxidized in photosystem (PS) II and PSI. When the water oxidase complex (WOC) was inactivated by acidic pH or by UV-B, Asc was photooxidized in PSII with Asc replacing water as the electron donor. An apparent Km was 2.5mM. At 20mM Asc, the electron transport rate reached 50 micromol NADP+ -reduced mgChl-1h-1. Asc was not oxidized by the PSII reaction center complex having an active WOC, and hence was suggested to function as an emergency donor only when WOC is inactivated. In the presence of 3-(3,4 dichlorophenyl)-1,1'-dimethylurea, Asc was photooxidized by PSI, with a lower affinity and an electron transport rate of 70 micromol NADP+ -reduced mgChl-1h-1 in 50mM Asc. Thus, Asc can support a PSI-mediated electron flow at a reasonably high rate at Asc concentrations in the physiologically relevant range. During the photooxidation of Asc by PSI, we observed the production of the monodehydroascorbate radical (MDA) that was unscavenged by the exogenously added MDA reductase. Reduction of P700+ by Asc was not affected by the inactivation of plastocyanin with cyanide. These results indicated that Asc was univalently oxidized on the lumenal side of PSI, directly by P700+. The electron transport Asc --> PSI --> NADP+ did not form a proton gradient across the thylakoid membrane, as determined by 9-aminoacridine fluorescence. Based on these results, we propose that the Asc-dependent cyclic electron flow around PSI, and that through both PSI and PSII can operate when the linear electron transport is partially impaired. PMID- 15288812 TI - A molecular thermodynamic view of DNA-drug interactions: a case study of 25 minor groove binders. AB - Developing a molecular view of the thermodynamics of DNA recognition is essential to the design of ligands for regulating gene expression. In a first comprehensive attempt at sketching an atlas of DNA-drug energetics, we present here a detailed thermodynamic view of minor-groove recognition by small molecules via a computational study on 25 DNA-drug complexes. The studies are configured in the MMGBSA (Molecular Mechanics-Generalized Born-Solvent Accessibility) framework at the current state of the art and facilitate a structure-energy component correlation. Analyses were conducted on both energy minimized structures of DNA drug complexes and molecular dynamics trajectories developed for the purpose of this study. While highlighting the favorable role of packing, shape complementarity, and van der Waals and hydrophobic interactions of the drugs in the minor groove in conformity with experiment, the studies reveal an interesting annihilation of favorable electrostatics by desolvation. Structural modifications attempted on the ligands point to the requisite physico-chemical factors for obtaining improved binding energies. Hydrogen bonds predicted to be important for specificity based on structural considerations do not always turn out to be significant to binding in post facto analyses of molecular dynamics trajectories, which treat thermal averaging, solvent, and counterion effects rigorously. The strength of the hydrogen bonds retained between the DNA and drug during the molecular dynamics simulations is approximately 1kcal/mol. Overall, the study reveals the compensatory nature of the diverse binding free energy components, possible threshold limits for some of these properties, and the availability of a computationally viable free energy methodology which could be of value in drug design endeavors. PMID- 15288813 TI - A 50-kDa isoform of mouse brain acyl-CoA hydrolase: expression and molecular properties. AB - Brain acyl-CoA hydrolase (BACH) is responsible for most of the long-chain acyl CoA hydrolyzing activity in the brain and is localized exclusively in neurons. There are two BACH isoforms: the major isoform, a 43-kDa BACH, and a lesser isoform, a 50-kDa BACH. In our previous work [Brain Res. Mol. Brain Res. 98 (2002) 81], a possibility was raised that these BACH isoforms might be generated from a single mRNA species via a mechanism of alternative use of translation start sites. However, the results obtained in the current study indicated that the 43-kDa BACH and 50-kDa BACH are not generated from a single mRNA species, but from distinct mRNA species transcribed by alternative use of transcription start sites. The molecular properties of the 50-kDa BACH were compared to those of the 43-kDa BACH. Palmitoyl-CoA hydrolase activity and protein stability were almost the same between both BACH isoforms. In addition, both 43-kDa BACH and 50-kDa BACH that were fused to green fluorescent protein showed cytosolic distribution. These results suggest that the 50-kDa BACH plays a similar role as the 43-kDa BACH. Therefore, since the 43-kDa BACH is expressed at higher levels than 50-kDa BACH, the 43-kDa BACH should largely contribute to understanding the physiological functions of the BACH gene in neurons. PMID- 15288814 TI - Nitric oxide-induced oxidant stress in endothelial cells: amelioration by ascorbic acid. AB - Nitric oxide has multiple beneficial effects in the blood vessel wall. However, high concentrations of nitric oxide in the presence of hydroperoxides have been shown to damage cultured cells. In this work, the effect of relatively high concentrations of nitric oxide alone on the function and antioxidant status of a human endothelial cell line (EA.hy926) was tested. Nitric oxide generated from 0.1 to 0.5mM spermine NONOate generated reactive species in the cells detected by triazole formation from diaminofluorescein and by oxidation of dihydrofluorescein. Intracellular ascorbic acid decreased this oxidant stress. Spermine NONOate also decreased intracellular ascorbate concentrations, although reduced glutathione was not affected unless cells had also been caused to reduce dehydroascorbic acid to ascorbate. Nitric oxide predictably inhibited both endothelial nitric oxide synthase and glyceraldehyde 3-phosphate dehydrogenase, and ascorbate partially prevented inhibition of the latter enzyme. These results suggest that relatively high concentrations of nitric oxide can cause oxidant stress in endothelial cells that is ameliorated by ascorbic acid. PMID- 15288815 TI - A new public health world order. PMID- 15288816 TI - SARS, emerging infections, and bioterrorism preparedness. PMID- 15288817 TI - Yersinial F1 antigen and the cause of Black Death. PMID- 15288818 TI - Was the Black Death yersinial plague? PMID- 15288819 TI - Grand round. PMID- 15288821 TI - Pathogenesis of filoviral haemorrhagic fevers. AB - The filoviruses, marburgvirus and ebolavirus, cause epidemics of haemorrhagic fever with high case-fatality rates. The severe illness results from a complex of pathogenetic mechanisms that enable the virus to suppress innate and adaptive immune responses, infect and kill a broad variety of cell types, and elicit strong inflammatory responses and disseminated intravascular coagulation, producing a syndrome resembling septic shock. Most experimental data have been obtained on Zaire ebolavirus, which causes uniformly lethal disease in experimentally infected non-human primates but produces a broader range of outcomes in naturally infected human beings. 10-30% of patients can survive the illness by mobilising adaptive immune responses, and there is limited evidence that mild or symptomless infections also occur. The other filoviruses that have caused human disease, Sudan ebolavirus, Ivory Coast ebolavirus, and marburgvirus, produce a similar illness but with somewhat lower case-fatality rates. Variations in outcome during an epidemic might be due partly to genetically determined differences in innate immune responses to the viruses. Recent studies in non human primates have shown that blocking of certain host responses, such as the coagulation cascade, can result in reduced viral replication and improved host survival. PMID- 15288822 TI - Schistosomal appendicitis. PMID- 15288823 TI - Confronting the avian influenza threat: vaccine development for a potential pandemic. AB - Sporadic human infection with avian influenza viruses has raised concern that reassortment between human and avian subtypes could generate viruses of pandemic potential. Vaccination is the principal means to combat the impact of influenza. During an influenza pandemic the immune status of the population would differ from that which exists during interpandemic periods. An emerging pandemic virus will create a surge in worldwide vaccine demand and new approaches in immunisation strategies may be needed to ensure optimum protection of unprimed individuals when vaccine antigen may be limited. The manufacture of vaccines from pathogenic avian influenza viruses by traditional methods is not feasible for safety reasons as well as technical issues. Strategies adopted to overcome these issues include the use of reverse genetic systems to generate reassortant strains, the use of baculovirus-expressed haemagglutinin or related non pathogenic avian influenza strains, and the use of adjuvants to enhance immunogenicity. In clinical trials, conventional surface-antigen influenza virus vaccines produced from avian viruses have proved poorly immunogenic in immunologically naive populations. Adjuvanted or whole-virus preparations may improve immunogenicity and allow sparing of antigen. PMID- 15288824 TI - Immunogenicity and efficacy of childhood vaccines in HIV-1-infected children. AB - Children infected by HIV-1 are particularly vulnerable to severe, recurrent, or unusual infections by vaccine-preventable pathogens. Routine immunisations seem to be generally safe for HIV-1-infected children, but responses may be suboptimal. Early HIV-1-induced immune attrition associated with viral replication may particularly interfere with the development of memory responses. In high HIV-1 prevalence regions, the accumulation of susceptible hosts may compromise disease-control efforts. Although early control of viral replication through treatment with highly active therapy may preserve immune function and responses to routine childhood vaccines, availability is limited in the areas most affected. In this review, we provide an overview of the immunogenicity and efficacy of childhood vaccines in HIV-1-infected children. The possible immunological bases for defective responses are discussed; unanswered questions and the need for further research are delineated. PMID- 15288825 TI - Lemierre's variant. PMID- 15288826 TI - Does combination antimicrobial therapy reduce mortality in Gram-negative bacteraemia? A meta-analysis. AB - The use of combination antimicrobial therapy for bacteraemia caused by Gram negative bacilli is controversial. We did a meta-analysis of published studies to determine whether a combination of two or more antimicrobials reduces mortality in patients with Gram-negative bacteraemia. Criteria for inclusion were: analytic studies of patients with documented Gram-negative bacteraemia that included patients receiving a single antibiotic (monotherapy) and patients receiving two or more antibiotics (combination therapy). Data on mortality (outcome) had to be provided. A pooled odds ratio was calculated with the random effects model of DerSimonian and Laird. Assessment of heterogeneity was done with the Breslow-Day test and reasons for heterogeneity were explored. 17 studies met the inclusion criteria, five prospective cohort studies, two prospective randomised trials, and ten retrospective cohort studies. Most studies used beta-lactams or aminoglycosides alone and in combination. The summary odds ratio was 0.96 (95% CI 0.70-1.32), indicating no mortality benefit with combination therapy. Subgroup analyses adjusting for year of publication, study design, and severity of illness did not change the results. Considerable heterogeneity was present in the main analyses. Analysis of only Pseudomonas aeruginosa bacteraemias showed a significant mortality benefit (OR 0.50, 95% CI 0.30-0.79). Our analysis does not support the routine use of combination antimicrobial therapy for Gram-negative bacteraemia, beyond settings where infection by P aeruginosa is strongly suspected or more than one drug would be desirable to assure in-vitro efficacy. PMID- 15288827 TI - Peripheral neuropathy associated with prolonged use of linezolid. AB - We present a case of a woman who developed severe, painful peripheral neuropathy while receiving linezolid therapy for 6 months. Nerve conduction studies indicated a sensory-motor axonal neuropathy. Extensive assessment did not show alternative explanations for her neuropathy. At the time of death 1 month after discontinuing linezolid, the neuropathy had not resolved. A review of published material shows a growing body of evidence that long-term use of linezolid may be associated with severe peripheral and optic neuropathy. 21 cases have been reported. In most cases, optic neuropathies resolved after stopping linezolid but peripheral neuropathies did not. The duration of therapy rather than indication for treatment seems to be the most important factor. The mechanism of toxicity is unknown but certain pharmacological properties of linezolid that may play a part are proposed. This report highlights the importance of post-approval surveillance and reporting of serious adverse drug effects, and potential consequences of off label use of pharmaceuticals. It further demonstrates the critical role clinicians have in communicating awareness of emerging drug toxicities. PMID- 15288828 TI - Spawning an idea. PMID- 15288829 TI - Radioimmunodetection and radioimmunotherapy of head and neck cancer. AB - Radiolabeled monoclonal antibodies (MAbs) can add a dimension to diagnostic imaging and staging of metastatic head and neck cancer, as well as in eradication of this disease. The vast majority of malignancies arising in the oral cavity, pharynx and larynx are squamous cell carcinomas. This common cellular origin makes it attractive to search for appropriate tumor-associated antigens, which are preferentially expressed in these neoplasms. Radiolabeled MAbs directed against these antigens can be used for tumor detection and selective therapy, known as radioimmunoscintigraphy and radioimmunotherapy, respectively. The combination of MAbs with positron emission tomography (PET) is an attractive novel option to improve tumor detection and to facilitate MAb quantification in a therapeutic setting. Basic aspects of tumor targeting with MAbs, as well as a review of the clinical trials reported in the literature, including own results, are presented. PMID- 15288830 TI - Merkel cell carcinoma of the head and neck. AB - Merkel cell carcinoma (MCC) is a rare neuroendocrine neoplasm of the skin. The tumor most frequently affects elderly patients, with a preference for the head and neck. Incidence rates increase with sun exposure and after immunosuppression and organ transplantation. A significant proportion of MCC have been reported to occur in intimate association with malignant epithelial neoplasms. The genetic mechanisms underlying the development and tumor progression of MCC are poorly understood, sharing pathogenetic mechanisms with other neoplasms of neural crest derivation. MCC has a propensity for spreading to regional lymph nodes, either at presentation or as a first site of relapse. Sentinel lymph node positivity is helpful in predicting the risk of recurrence or metastasis in patients with MCC. Complete surgical resection is the mainstay of treatment of the primary tumor. Tumor resections are recommended to include a 2-3-cm tumor-free margin around the primary lesion when possible, but this is often difficult to achieve in the head and neck, where Mohs micrographic surgery has proved to be effective. The role of adjuvant radiation therapy is controversial. The role of adjuvant chemotherapy in diminishing the risk of subsequent systemic recurrence in patients with positive nodes remains undefined. Overall response rates to combination chemotherapy for surgically unresectable distant metastatic disease are generally high, although responses are transient. Overall survival of head and neck MCC at 5 years postoperatively ranks between 40% and 68%. MCC has a high incidence of locoregional recurrences, but even after a locoregional failure, a substantial proportion of patients achieve long-term disease-free survival. Finally, factors generally associated with survival are the stage of disease at presentation, distant recurrence usually being the most adverse predictor of survival. PMID- 15288831 TI - Predictive factors of occult metastasis and prognosis of clinical stages I and II squamous cell carcinoma of the tongue and floor of the mouth. AB - The incidence of occult neck metastasis in early stage tumours of the tongue and floor of the mouth varies from 20% to 30%, and the survival rates in 5 years from 73% to 97%. This study analyzes the rates of occult metastasis and prognostic factors for clinical stages I and II squamous cell carcinoma of the tongue and floor of the mouth. The records of patients with squamous cell carcinoma of the tongue and floor of the mouth, without prior treatment and treated by surgery between 1965 and 1998 were reviewed. All cases were re-staged and the surgical specimens were reviewed. This study included 193 patients, 145 men (75.1%), with ages ranging from 29 to 89 years old (mean, 60 years). The tumour site was the tongue in 132 cases (68.4%), the floor of the mouth in 45 (23.3%) and both in 16 (8.3%). With regard to stage, 85 cases were at clinical stage I (44.0%) and 108, clinical stage II (56.0%). One hundred and seventeen patients (60.6%) were submitted to a neck dissection and 27 (23.1%) had metastasic lymph nodes (pN+). The only factor associated with the presence of occult metastasis for all patients was the presence of muscular infiltration (p = 0.020); for tongue tumours the presence of vascular embolization (p = 0.043) and the presence of desmoplastic reaction (p = 0.050); for floor of the mouth tumours and T2 tumors, the histological grade (p = 0.025 and p = 0.035, respectively). Disease-free survival in 5 years was 66.4% and overall survival in 5 years 68.5%. The only factor associated with disease-free survival was the presence of muscular infiltration (p = 0.019) and with overall survival were gender (p = 0.002) and clinical stage (p = 0.031). Tumours of the tongue and floor of the mouth in the initial stages, which had muscular infiltration showed a higher probability of occult metastasis and lower disease-free survival; T2 tumours showed a worse survival as did patients of the male gender. PMID- 15288832 TI - Detection of photodynamic therapy-induced early apoptosis in human salivary gland tumor cells in vitro and in a mouse tumor model. AB - We studied the detection of apoptosis of malignant human salivary gland tumor cells induced by photodynamic therapy (PDT) using the photosensitizer mono-L aspartyl chlorin e6 (NPe6) in vitro and in vivo in mice receiving transplanted human salivary gland tumor (HSG) cells. An immunohistocytochemical method using a monoclonal antibody (MoAb), M30, which reacts with the product resulting from the cleavage of cytokeratin (CK) 18 by activated caspase, was applied to detect the apoptosis of HSG cells induced by PDT. Significant amounts of immunoreactive products were observed in the cytoplasm of HSG cells after PDT. In vitro, M30 positive cells increased from 2 h after PDT, increased rapidly from 8 h and reached a peak 24 h after PDT. In vivo, a peak of early apoptosis was confirmed two hours after PDT. In comparison with DNA fragmentation detected by the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) method, the destroyed tumor cells were observed sporadically 24 h after PDT. These results suggest that immunohistocytochemical staining with the MoAb M30 may be useful for detecting early apoptosis induced by PDT. Futhermore, PDT using NPe6 is effective in inducing apoptosis of HSG cells at an early stage, which suggests the possibility of the therapy being ideal for treatment of human malignant neoplasms. PMID- 15288833 TI - Treatment results of oral verrucous carcinoma and its biological behavior. AB - The biologic behavior of and optimal treatment for oral verrucous carcinoma (VC) remain controversial. We analyzed the clinicopathological characteristics of 12 patients with oral VC. Immunohistochemical techniques were used to evaluate p53 protein, CD44 variant 9, and proliferating cell nucleus antigen. The TNM classification (UICC, 1997) was T1 in 1 patient, T2 in 3, T3 in 4, and T4 in 4. All patients were classified as N0M0. Four patients were treated by surgery alone and 8 by surgery after chemotherapy, radiotherapy, or both. After surgery, two patients had primary recurrence of disease. Immunohistochemically, the proliferative activity of tumor cells as evaluated by proliferating cell nuclear antigen labeling index and p53 protein expression was similar in VC and well differentiated squamous cell carcinoma. However, CD44 varient 9 expression was positive in 8 of 10 VC, suggesting that oral VC is associated with a low risk of lymph node metastasis. Positive CD44 variant 9 expression by most oral VCs, indicating a low risk of cervical lymph node metastasis, suggests that most cases can be controlled by surgical intervention. PMID- 15288834 TI - Expression of MMP-9, TIMP-1, CD-34 and factor-8 as prognostic markers for squamous cell carcinoma of the tongue. AB - Squamous cell carcinoma (SCC) of the tongue is characterized by an unpredictable course, ranging from relatively benign to a high degree of locally aggressive growth and metastasis. Treatment guidelines have been developed according to TNM stage, but they do not always accurately predict clinical outcome. The aim of the present study was to evaluate the expression of the matrix metalloproteinases (MMPs) that degrade the extracellular matrices, their inhibitors (TIMPs), and angiogenic factors (factor-8 and CD-34) in tumor cells and to correlate these findings with the clinicopathological features and patient outcome. Tissue specimens from 23 patients with primary SCC of the tongue were immunohistochemically stained for these markers. High expressions of MMP-9 and TIMP-1 were detected in 60.9% and 65.2% of the specimens, respectively. Tumor invasion to adjacent muscle, lymph node metastasis, and disease status at the end of follow-up were positively correlated with the microvessel count using the CD 34 marker, but not with high expression of MMP-9 or TIMP-1. Expression of MMP-9 and TIMP-1 fails to predict aggressiveness in SCC of the tongue. However, the degree of vascularization in tumor tissue is indicative of disease aggressiveness and might be used as a basis for patient selection for more intensive therapy. PMID- 15288835 TI - Expression of tumor necrosis factor-alpha and its soluble receptors in betel-quid chewing patients at different stages of treatment for oral squamous cell carcinoma. AB - Tumor necrosis factor-alpha (TNF-alpha) can inhibit tumor progression. It can be regulated by its soluble receptors (sTNF-Rs). We examined the expression of TNF alpha and sTNF-Rs and the TNF-alpha/sTNF-R ratios in betel-quid-chewing patients with oral squamous cell carcinoma (OSCC) to see if these parameters are associated with disease progression according to the treatment stage. Peripheral blood mononuclear cells from 116 OSCC patients at different treatment stages and 19 betel-quid chewers with normal mucosa were assayed with ELISA. Levels of sTNF RII in the OSCC patients were significantly higher than normal controls, with the recurrence group having the highest levels. After controlling for age and use of alcohol and tobacco, the TNF-alpha/sTNF-RII ratio showed significant differences comparing OSCC patients at each treatment stage with normal controls. Our results suggest that sTNF-RII and TNF-alpha/sTNF-RII ratio may be informative for the diagnosis and prognosis of OSCC. PMID- 15288836 TI - Severe oral chronic graft-versus-host disease following allogeneic bone marrow transplantation: highly effective treatment with topical tacrolimus. AB - Oral involvement of chronic graft-versus-host disease (GvHD) is a most distressing and disabling complication of hematopoietic cell transplantation, for which systemic immunosuppression as well as topical corticosteroid treatment may offer only limited symptomatic relief. Here we report encouraging preliminary results with the application of tacrolimus (FK-506) as a 0.1% ointment in three patients with severe oral chronic GvHD, heavily pretreated without success, who experienced rapid, consistent, complete or at least marked, subjective and objective improvement with topical tacrolimus. PMID- 15288837 TI - Effects of individual characteristics on healthy oral mucosa autofluorescence spectra. AB - Autofluorescence spectroscopy is a tool for detecting tissue alterations in vivo. In a previous study, we found spectral differences between clinically normal mucosa of different patient groups. These are possibly caused by associated patient characteristics. In the present study, we explore the influences of volunteer characteristics on healthy oral mucosa autofluorescence. Autofluorescence spectra were recorded in 96 volunteers with no clinically observable oral lesions. We applied principal components analysis to extract the relevant information. We used multivariate linear regression techniques to estimate the effect of volunteer characteristics on principal component scores. Statistically significant differences were found for all factors but age. Skin color strongly affected autofluorescence intensity. Gender differences were found in blood absorption. Alcohol consumption was associated with porphyrin-like peaks. However, all differences but those associated with skin color were of the same order of magnitude as standard deviations within categories. The effects of volunteer characteristics on autofluorescence spectra of the oral mucosa are measurable. Only the effects of skin color were large. Therefore, in lesion classification, skin color should be taken into account. PMID- 15288838 TI - The sensitivity and specificity of the OralCDx technique: evaluation of 103 cases. AB - In this study, we compared 103 OralCDx results with the histological findings of 96 clinical sites in 80 patients (33 females; 64.3+/-13.7 years and 47 males; 53.2+/-11.5 years). The histological findings were classified as follows: compatible with oral leukoplakia (OL; n = 60) or oral lichen planus (OLP; n = 17), both without dysplasia; dysplasia in OL or OLP (n = 9); and oral squamous cell carcinoma (OSCC; n = 17). There were seven (6.8%) specimens with an inadequate cell count. Overall, the sensitivity of the OralCDx technique to detect dysplasia and OSCC was 92.3% (95% CI: 74.9-99.1%), and the specificity was 94.3% (95% CI: 86.0-98.4%). The positive likelihood ratio (LR+) was 16.2 (95% CI: 6.2-42.1) and the negative likelihood ratio (LR-) was 0.08 (95% CI: 0.02-0.31). In conclusion, these figures are in agreement with previously published data and support the use of OralCDx as a screening tool of oral lesions, but further trials are still necessary. PMID- 15288839 TI - An audit of the efficacy of the oral brush biopsy technique in a specialist Oral Medicine unit. AB - The diagnosis of oral epithelial dysplasia has traditionally been based upon histopathological evaluation of a full thickness biopsy specimen from lesional tissue. It has recently been proposed that cytological examination of "brush biopsy" samples is a non-invasive method of determining the presence of cellular atypia, and hence the likelihood of oral epithelial dysplasia. The present audit determined, retrospectively the sensitivity, specificity and positive and negative predictive values of the oral brush biopsy technique in the diagnosis of potentially malignant disease in a group of 112 patients attending a specialist Oral Medicine unit. The sensitivity of detection of oral epithelial dysplasia or squamous cell carcinoma of the oral brush biopsy system was 71.4% while the specificity was 32%. The positive predictive value of an abnormal brush biopsy result (positive or atypical) was 44.1%, while the negative predictive value was 60%. It is concluded that not all potentially malignant disease is detected with this non-invasive investigative procedure. PMID- 15288840 TI - Proliferative verrucous vs conventional leukoplakia: no significantly increased risk of HPV infection. AB - Proliferative verrucous leukoplakia (PVL) is a very aggressive form of oral leukoplakia (OL) with high morbidity and mortality rates, hypothesised to be linked to HPV infection. This study aimed to determine the presence of HPV DNA in PVL in comparison with OL, and in relation to social-demographical variables (age, gender, smoking and drinking habits) in an Italian multi-centric hospital based study. The study group consisted of 58 cases of PVL and 90 cases of OL as controls (47 homogeneous (H) and 43 non-homogeneous (non-H) form), both recruited from four Italian cohorts. HPV DNA was identified in exfoliated mucosal cells by nested PCR (nPCR) with MY09/MY11 and GP5+/GP6+ primer pairs and the HPV genotype determined by direct DNA sequencing. HPV DNA was found in 24.1% (14/58)of PVL and in 25.5% (23/90) of OL; there was thus no significant difference found between PVL and OL (both forms) for risk of HPV infection (OR=0.93; 95% IC:0.432-1.985). Similarly, in both groups of PVL and OL lesions, no statistic association was found between any demographical variable considered and HPV infection. HPV-18 was the most frequently detected genotype in all tissues, being found in 78.5% and 60.8% of HPV+ve PVL and OL, respectively. Other more rarely detected genotypes were HPV-16 (28.6% in PVL and 13% in OL), HPV-6 (17.4% in OL) and HPV-53 (8.8% in OL). PVL does not appear more likely to be associated to HPV infection than conventional OL lesions. PMID- 15288841 TI - Ameloblastin gene (AMBN) mutations associated with epithelial odontogenic tumors. AB - Ameloblastin (AMBN, MIM *601259) gene expresses an important protein (AMBN), present in the organic matrix of enamel. The AMBN protein has an important role in the differentiation of ameloblast cells and epithelium-mesenchyme signaling during odontogenesis which prompted us to investigate this gene in aggressive epithelial odontogenic tumors, such as ameloblastomas, and in some non-aggressive ones, such as the adenomatoid odontogenic tumor and the squamous odontogenic tumor. Six cases of epithelial odontogenic tumors were studied and normal cells of the patient's mucosa were used as negative controls. The results demonstrated novel mutations in all tumors, while mucosal cells showed the wild type DNA sequence. Our data demonstrates that AMBN gene has an important role in the tumorigenesis of subtypes of epithelial odontogenic tumors and that this phenotypic heterogeneity could be caused by genetic heterogeneity. PMID- 15288842 TI - Important prognostic factors of long-term oropharyngeal carcinoma survivors in Taiwan. AB - In Taiwan, a clear gender difference emerges for rates of oropharyngeal carcinoma incidence. The purpose of this study was to identify the gender differences and clinical factors associated with oropharyngeal carcinoma survival rates in Taiwan. We analyzed the 5-year survival rates of 8114 subjects diagnosed with oropharyngeal carcinoma between 1987 and 1994. The Cox proportional-hazards model identified clinical characteristics for gender according to oropharyngeal carcinoma death and all-cause death outcomes. The 5-year survival rates were significantly lower for males than females (p < 0.0001). The adjusted hazard ratio of males versus females was 1.54 (95% CI: 1.36-1.74) for oropharyngeal carcinoma death and 1.44 (95% CI: 1.31-1.58) for all-cause death. Gender and other clinical characteristics (i.e. diagnostic age, anatomic site, morphologic type, and treatment modality) play important roles in oropharyngeal carcinoma survival. We suggested that Taiwanese males have high proportion of betel quid chewing and that this is associated with the gender differences. PMID- 15288843 TI - Keratosis follicularis of the oral mucosa with oral squamous cell carcinoma. AB - Darier's disease or keratosis follicularis is a genodermatosis which may involve the oral mucosa. Malignant degeneration is rare. We report the first case of the combined manifestation of oral keratosis follicularis and oral squamous cell carcinoma and discuss the possible involvement of ATP2A2 (located in 12q23-24.1) which encodes the sarco/endoplasmic reticulum Ca(2+)-ATP isoform 2 (SERCA2), in the pathogenesis. PMID- 15288847 TI - Is dosage reduction appropriate in patients who respond well to anti-TNF-alpha agents? PMID- 15288848 TI - Strontium: a new treatment for osteoporosis. PMID- 15288849 TI - Bone density measurement in older women: who and why? PMID- 15288850 TI - Acetaminophen as symptomatic treatment of pain from osteoarthritis. AB - Osteoarthritis is a major public health burden. The incidence of osteoarthritis increases with advancing age. Symptomatic treatments aimed at alleviating the pain and thereby restoring joint function form the basis of the treatment. The chronic course requires long-term treatment with special attention to minimizing the side effects of drugs. Acetaminophen has a good risk/benefit ratio that has prompted international consensus panels to recommend its use as first-line therapy in dosages of up to 4 g/day. This review discusses safety and efficacy data from randomized double-blind trials of acetaminophen used to alleviate pain caused by osteoarthritis. PMID- 15288851 TI - Chronic autoimmune thyroiditis and rheumatic manifestations. AB - A variety of rheumatic manifestations have been described in association with autoimmune thyroiditis. In the past, most of these manifestations were attributed to the underlying thyroid dysfunction, in particular hypothyroidism. However, a responsibility of the mechanisms involved in the autoimmunity rather than a direct action of thyroid hormones seems supported by the evidences that some rheumatic manifestations may occur even in euthyroid patients, or that they are more frequent in hypothyroid patient with autoimmune thyroiditis than in those without this disease. Rheumatic manifestations could be sometimes attributable to the autoimmune rheumatic diseases frequently associated with autoimmune thyroiditis, such as Sjogren's syndrome, rheumatoid arthritis, systemic lupus erythematosus, or scleroderma. Among the most important or frequent rheumatic manifestations there are a mild non-erosive variety of arthritis, polyarthralgia, myalgia, and sicca syndrome without a true Sjogren's syndrome. Although the possible pathogenesis of these manifestations is not completely established, some hypotheses may be proposed, including a role of autoantibodies characteristics of autoimmune thyroiditis, a possible overlap between autoimmune thyroiditis and some autoimmune rheumatic diseases, and a systemic inflammatory reaction associated with thyroiditis. PMID- 15288852 TI - Tumor necrosis factor receptor-associated periodic syndrome (TRAPS): definition, semiology, prognosis, pathogenesis, treatment, and place relative to other periodic joint diseases. AB - Tumor necrosis factor receptor-associated periodic syndrome (TRAPS) is an autosomal dominant inherited condition of periodic fever and pain. Most patients are of northern European descent. The attacks manifest as fever and pain in the joints, abdomen, muscles, skin, or eyes, with variations across patients. An acute-phase response occurs during the attacks. Patients with TRAPS are at risk for AA amyloidosis, the most common targets being the kidneys and liver. Soluble TNFRSF1A is usually low between the attacks and may be normal during the attacks, when TNF levels are high. TNFRSF1A is found in abnormally high numbers on leukocyte cell membranes. TRAPS is the first condition for which naturally occurring mutations in a TNF receptor were found; the mutations affect the soluble TNFRSF1A gene in the 12p13 region. In some patients, the pathogenesis involves defective TNFRSF1A shedding from cell membranes in response to a given stimulus. Thus, TRAPS is a model for a novel pathogenic concept characterized by failure to shed a cytokine receptor. This review compares TRAPS to other inherited periodic febrile conditions, namely, familial Mediterranean fever, Muckle-Wells syndrome, cold urticaria, and hyper-IgD syndrome. The place of TRAPS relative to other intermittent systemic joint diseases is discussed. Colchicine neither relieves nor prevents the attacks, whereas oral glucocorticoid therapy is effective when used in dosages greater than 20 mg/day. The pathogenic hypothesis involving defective TNFRSF1A shedding suggests that medications targeting TNF may be effective in TRAPS. PMID- 15288853 TI - Osteoporosis and ankylosing spondylitis. AB - Ankylosing spondylitis is a chronic inflammatory joint disease in which inflammation of the entheses gradually causes local ossification and ankylosis. Diffuse osteoporosis responsible for bone fragility is another feature established by recent data. The bone loss predominates at the spine and can cause vertebral fractures that are rare but nonnegligible sources of mortality and morbidity late in the course of the disease. PMID- 15288854 TI - Joint lavage for treating recurrent knee involvement in patients with juvenile idiopathic arthritis. AB - OBJECTIVE: To retrospectively evaluate the benefits of knee joint lavage with intraarticular glucocorticoid injection in patients who have juvenile idiopathic arthritis with knee involvement unresponsive to repeated intraarticular glucocorticoid injections. PATIENTS: Seventeen knees in 10 children (eight girls and two boys) were treated from 1997 to 2000. Mean age was 14 years 9 months and mean disease duration was 7.2 years. The diagnoses were juvenile oligoarthritis (n = 6, including two with extended disease), systemic arthritis (n = 2), juvenile spondyloarthropathy (n = 1), and juvenile dermatomyositis (n = 1). Repeated intraarticular triamcinolone hexacetonide injections had been performed in all the patients, the mean number of injections being 2.2 per patient within the last 30 months. Plain radiographs were normal in six of the eight patients. Mean erythrocyte sedimentation rate was 21.7 mm/h and mean C-reactive protein level was 20.6 mg/l. Joint fluid was obtained from 10 knees and had a mean cell count of 12?660 mm(-3). Second-line therapy was with methotrexate alone or combined with cyclosporine or azathioprine. Oral glucocorticoids and/or nonsteroidal antiinflammatory drugs were used for symptom relief. TREATMENT PROCEDURE: Lavage was performed under strict aseptic conditions with simple analgesia, on a day-hospital basis. After aspiration of the joint, lavage was performed with saline, and a delayed-action glucocorticoid was injected. The knee joint was immobilized in the extended position for 48 h. Efficacy criteria were presence of effusion, presence of pain, and presence of a systemic treatment sparing effect. RESULTS: Freedom from effusion and pain was noted in all 17 knees after 1 month, in eight (47%) knees after 6 months, and in seven (41%) knees after 12 months. The patients with the longest lasting improvements had systemic polyarthritis. After joint lavage, second-line treatment was reduced in two patients and oral glucocorticoid therapy was stopped in two others. None of the variables studied (age, sex, disease duration, inflammatory syndrome, or joint fluid cytology) predicted a good response. No adverse effects were recorded. CONCLUSION: These preliminary results show that joint lavage with glucocorticoid injection is safe in children. The improvements were modest, but the patients had a history of arthritis refractory to multiple triamcinolone hexacetonide injections. Thus, joint lavage may have a place in the treatment pyramid just before synovectomy. PMID- 15288855 TI - Is inflammatory joint disease in HIV-infected patients a form of spondyloarthropathy? AB - In Congo-Brazzaville, the seroprevalence of HIV infection is in the range 7-8%, and AIDS is the leading cause of aseptic arthritis (60% of cases). PATIENTS, MATERIAL AND METHODS: The ESSG and Amor's criteria for spondyloarthropathy were evaluated in 83 patients with HIV infection admitted for aseptic arthritis to the rheumatology department of the Brazzaville teaching hospital, over an 8-year period. RESULTS: All 83 patients were CDC stage IV; 66 (80%) had polyarthritis and 17 (20%) oligoarthritis. A single patient met ESSG and Amor's criteria, with six points; one patient had five points and 15 had three points. The joint involvement was asymmetrical and nonerosive. Nonsteroidal antiinflammatory therapy ensured resolution of the manifestations within 4-8 weeks. The most common sites of involvement were the knees (84%), ankles (59%), and great toes (23%) at the lower limbs and the wrists (41%), elbows (29%), and metacarpophalangeal and interphalangeal joints (25%) at the upper limbs. CONCLUSION: Inflammatory arthritis in HIV patients does not meet ESSG or Amor's criteria for spondyloarthropathy. PMID- 15288856 TI - Streptococcal septic arthritis in adults. A study of 55 cases with a literature review. AB - OBJECTIVES: To evaluate the rate of occurrence and characteristics of streptococcal septic arthritis. METHODS: Retrospective single-center study of patients with bacteriologically documented septic arthritis admitted to a rheumatology department over a 20-year period. RESULTS: Of 303 cases of septic arthritis, 55 (18%) were due to streptococci and 166 (55%) to Staphylococcus aureus (55%). As compared to patients with S. aureus arthritis, patients with streptococcal arthritis was more likely to be in female (56% vs. 36%, P < 0.006) and older than 60 years of age (71% vs. 58%), less likely to have comorbidities (36% vs. 56%), rheumatoid arthritis (5% vs. 19%, P < 0.01), or diabetes (2% vs. 15%, P < 0.01), and more likely to have cancer (13% vs. 7%). Involved joints and proportions of patients with arthritis in multiple joints were similar in two groups. Mortality was lower in the group with streptococcal infection (3.6% vs. 7.8%). The streptococci were distributed as follows: group A (n = 7), group B (n = 12), group C (n = 4), group D (n = 7), group F (n = 1), group G (n = 2), nongroupable (n = 14), nontypable (n = 1), and Streptococcus pneumoniae (n = 7). Groups A and B and nongroupable strains mainly affected women; group A selectively involved younger patients and group B very elderly patients. Comorbidity, most notably cancer, was common in patients with S. pneumoniae or group D streptococci. The portal of entry was often a skin lesion for groups A and B and a medical procedure for group D. Multiple joint involvement was common with groups A and B and prosthetic joint infection with groups B and C. Group A and S. pneumoniae were associated with severe systemic symptoms and extra articular foci of infection, whereas a smoldering course was more common with groups D and G and with nongroupable strains. Residual joint abnormalities were noted in half the patients, with no differences across groups. CONCLUSIONS: The features of streptococcal septic arthritis vary according to the group of the causative organism and differ from those of S. aureus arthritis. PMID- 15288857 TI - The results of rehabilitation on motor and functional improvement of the spinal tuberculosis. AB - OBJECTIVES: To evaluate the result of rehabilitation on motor and functional improvement in spinal tuberculosis. METHOD: Prospective case study. Data were collected from 47 patients with spinal tuberculosis medically and/or surgically treated, and rehabilitated over 6 months of period, after spinal decompression and fusion. The main outcome measures were motor development of the patients who were evaluated at the beginning, in the 1st week, in the 3rd month, and in the 6th month. Functional development of the patients was evaluated at the beginning and in the 6th month. Functional assessment was made according to Modified Barthel Index (MBI), and motor examination was made according to American Spinal Injury Association (ASIA). RESULTS: The study population consisted of 47 patients (22 males and 25 females) mean aged 37.9 +/- 18.3 years (range 5-76 years). The most common site of spinal tuberculosis was the thoracic region. Localized back pain, paraparesis, sensory dysfunction and fever were typical clinical manifestations. Surgical management was performed as anterior or posterior drainage of abscess and/or stabilization of the spine. The rehabilitation program was performed in all patients during the preoperative, early postoperative and late postoperative 6 month periods. Muscle-strengthening exercises on necessary localization such as pectoral, abdominal, lower extremities; truncal and sacrospinal extensors were started for the rehabilitation. The motor score for the lower limbs and the MBI scores for activities of daily living (ADL) and mobility improved significantly (P < 0.001). The self-care and mobility categories of the MBI on admission; were 14.8% severely dependent and 10.6% independent. However, at the end of the rehabilitation program, 4.2% were severely dependent and 70.2% independent. IN CONCLUSION: Early diagnosis and appropriate medical and/or surgical treatment together with a rehabilitation program will improve the life quality of patients with spinal tuberculosis. PMID- 15288858 TI - The ability to change of three questionnaires for neck pain. AB - OBJECTIVE: To assess the sensitivity to change of three algofunctional scales for neck pain. METHODS: Observational, prospective study. Patients with neck disorders were included. Pain and patients' perceived handicap assessed on visual analogue scales (VAS Pain, VAS Handicap), and functional disability measures (Neck Disability Index, Neck Pain and Disability Scale, Northwick Park Neck Pain Questionnaire) were recorded twice, at baseline and at an 11-month follow-up assessment. Sensitivity to change was assessed using the effect size (ES) and the standardised response mean (SRM), and the non-parametric Spearman's rank correlation coefficient (r) was used to assess the correlation between quantitative variable changes and patients' overall opinion. Analysis of variance (ANOVA) followed by a Tukey-test was performed to determine if the scales distinguished improved, stable, and deteriorated patients. RESULTS: Seventy-one patients (43 women, mean age 49 years) were included and evaluated twice at an interval of 11 +/- 2 months. The three scales showed good sensitivity to change. The ANOVA showed a group effect, and individual changes in the scales scores were statistically different in two-by-two comparisons (improved, stable or deteriorated patients). Changes in NPDS scores had the highest correlation with patient's overall assessment (r = 0.592). CONCLUSION: The three scales can detect changes in patients with neck disorders. Changes in NPDS score had the best correlation with patients' overall opinion concerning their neck disorder and this questionnaire should be given preference in clinical trials. PMID- 15288859 TI - Unusual and aggressive thrombotic onset in two children with antiphospholipid antibody syndrome and systemic lupus erythematosus. AB - We report two pediatric patients with unusual, aggressive initial manifestation of antiphospholipid antibody syndrome secondary to systemic lupus erythematosus. The first patient, a 13-year-old girl, presented with bilateral amaurosis and ischemic cerebral lesions. The second, another 13-year-old girl, presented with cerebral venous sinus thrombosis and membranous glomerulonephritis. Both patients improve after treatment with anticoagulants and immunosuppressive drugs, two therapies that are aimed at modulating the immune response or towards preventing thromboembolic events. However, there is no consensus regarding the duration and intensity of oral anticoagulation in children with antiphospholipid antibody syndrome. PMID- 15288860 TI - IgD kappa myeloma: a new case. AB - IgD myeloma, which is particularly severe, accounts for only 1-3% of all myeloma cases, and the kappa subtype contributes only 10-30% of IgD myelomas. We report a new case. CASE-REPORT: A 59-year-old man was admitted for inflammatory low back pain with L5 sciatica and diffuse bone pain. The symptoms had been present for 3 months, during which he had experienced a severe decline in general health. Laboratory test abnormalities included an erythrocyte sedimentation rate of 70 mm/h, normochromic normocytic aplastic anemia, hypercalcemia (3.5 mmol/l), and serum creatinine elevation to 583 micromol/l. Tests were negative for cryoglobulinemia. Serum protein electrophoresis showed hypergammaglobulinemia but no monoclonal peak. Immunoelectrophoresis, however, detected a faint IgD kappa band in the blood and a homogeneous kappa band in the urine. Bone marrow aspirated from the sternum was found to contain 30% of malignant plasma cells. Biopsies for amyloidosis were negative. Radiographs disclosed multiple punched out lesions with no evidence of spinal cord compression. Symptomatic treatment was given to correct the hypercalcemia, and combination chemotherapy was started. DISCUSSION: IgD kappa myeloma is a severe variant of myeloma often associated with extraosseous lesions, renal failure, and amyloidosis. The monoclonal component is absent or faint by serum protein electrophoresis, making the diagnosis difficult. The pathogenesis is unclear and the prognosis grim. PMID- 15288861 TI - Caudal spinal cord ischemia after lumbar vertebral manipulation. AB - Neurological complications after lumbar spine manipulation are uncommon. The cause is usually a herniated disk or displaced bony structure. We report a case of paraplegia that developed a few hours after manipulation of the lumbar spine. Magnetic resonance imaging was consistent with ischemia of the caudal spinal cord. No disk fragment or bony structure impinging on the spinal cord was seen. Spinal cord ischemia may deserve to be added to the list of possible adverse events after lumbar spine manipulation. PMID- 15288862 TI - Group B streptococcal spondylodiscitis in adults: 2 case reports. AB - Streptococcus agalactiae, or group B streptococcus (GBS), has been traditionally considered an infrequent etiologic agent of disease in adults except for urinary tract infection in pregnant women. Attention has recently been drawn to other adult infections caused by GBS such as skin and soft tissue infections, bacteriemias, pneumonia, meningitis, endocarditis, peritonitis, and bone and joint infections. We present two adult patients with GBS spondylodiscitis and review 30 cases of GBS spinal infection previously reported in the literature. This series clearly illustrates that GBS has recently been recognized as an emerging cause of vertebral infections in adults, particularly in those with chronic underlying diseases, although it can also affect immunocompetent patients without debilitating conditions. Although uncommon, GBS should be considered in the differential diagnosis of infective spondylodiscitis in nonpregnant adults, whatever the patient's immunological status. PMID- 15288863 TI - Systemic lupus erythematosus with celiac disease: a report of five cases. AB - Systemic lupus erythematosus and celiac disease (CD) are rarely reported in combination. We report five cases seen over a 4-year period. The two conditions occurred concomitantly in one patient, whereas the CD antedated the lupus in one patient and postdated the lupus in the remaining three patients. Villous atrophy on duodenal biopsy specimens with a favorable response to a gluten-free diet was noted in all five patients. Only four patients had positive serological tests for CD and only three had abdominal symptoms. PMID- 15288864 TI - Bilateral lumbosacral plexopathy after femoral vein dialysis: synopsis of a case. AB - Femoral vein dialysis is a technique applied in many clinics. Hemorrhagic complications following the procedure either directly due to the femoral catheterization itself in the early period or less with concomitant late neurological impairments may pose serious challenges to the clinician. Likewise in this report, we are presenting a dialysis patient with bilateral retroperitoneal hematomas causing bilateral lumbosacral plexopathies-to our best knowledge the first in the literature. We have also touched upon its prompt diagnosis and treatment. PMID- 15288865 TI - Compressive injury of the brachial plexus after axillary arteriography and its further consequences. AB - A patient suffering from a brachial plexus injury after axillary arteriography is discussed. In the pertinent literature, local complications such as hematomas or pseudoaneurysms have been reported. Herein, rendering this patient, we introduce a new type of a delayed complication after the angiography-compressive injury of the brachial plexus. PMID- 15288866 TI - Acute pseudoseptic arthritis after intraarticular sodium hyaluronan. AB - A 70-year-old woman with a history of knee osteoarthritis was admitted for acute arthritis 9 days after a second intraarticular injection of sodium hyaluronan (Ostenil). The joint fluid was purulent, with no crystals, and laboratory tests showed marked inflammation, leading to antibiotic treatment for suspected septic arthritis. Incapacitating symptoms persisted, prompting surgical lavage of the knee, which failed to relieve the severe pain. The persistent symptoms and negative results of joint fluid and blood cultures led to discontinuation of the antibiotic therapy after 10 days. Antiinflammatory therapy relieved the symptoms, and the patient was discharged home 1 month after her admission. Nevertheless, the pain persisted, requiring rehabilitation therapy of the knee. Aseptic arthritis induced by repeated sodium hyaluronan injection is the most likely diagnosis. Physicians should be aware of this extremely severe complication. PMID- 15288867 TI - Is chromatin remodeling required to build sister-chromatid cohesion? AB - Chromosome segregation during mitosis and meiosis depends on the linkage of sister DNA molecules after replication. These links, known as sister-chromatid cohesion, are provided by a multi-subunit complex called cohesin. Recent papers suggest that chromatin-remodeling complexes also have a role in the generation of sister-chromatid cohesion. It remains unclear whether they do so by facilitating the recruitment of cohesin to specific chromosomal sequences or by modifying an event at replication forks giving rise to cohesion between sister DNAs. PMID- 15288868 TI - BTLCP proteins: a novel family of bacterial transglutaminase-like cysteine proteinases. AB - Using sequence similarity searches and top-of-the-range fold-recognition methods, we have identified a novel family of bacterial transglutaminase-like cysteine proteinases (BTLCPs) with an invariant Cys-His-Asp catalytic triad and a predicted N-terminal signal sequence. This family of previously uncharacterized hypothetical proteins encompasses sequences of unknown function from DUF920 (in the Pfam database) and COG3672. BTLCPs are predicted to possess the papain-like cysteine proteinase fold and catalyze post-translational protein modification through transamidase, acetylase or hydrolase activity. Inspection of neighboring genes encoding BTLCPs suggests a link between this predicted activity and a type I secretion system resembling ATP-binding cassette exporters of toxins and proteases involved in bacterial pathogenicity. PMID- 15288869 TI - GIFT domains: linking eukaryotic intraflagellar transport and glycosylation to bacterial gliding. AB - We describe GIFT [for GldG, intraflagellar transport (IFT)] domains in the flavobacterial gliding protein GldG and eukaryotic IFT-52. In eukaryotes, domain homologues are also found in the eukaryotic oligosaccharyltransferase complex and in subtilisin kexin isozyme-1 (SKI-1 or S1P). A distant evolutionary relationship to periplasmic-binding proteins hints that GIFT domains might possess oligosaccharide-binding functions. PMID- 15288870 TI - The right position at the right time: mobility makes a difference. PMID- 15288871 TI - Practical considerations in the administration of intravenous vasoactive drugs in the critical care setting. Part II--how safe is our practice? AB - INTRODUCTION: Vasoactive drugs (e.g. inotropes), namely adrenaline and noradrenaline, are frequently used in critical care to maintain cardiovascular function. This is achieved by ensuring that a continuous infusion of the vasoactive drug is administered so that when one infusion is about to finish another infusion is commenced. This is known as "double pumping" or "piggy backing". Failure to administer these drugs appropriately may result in haemodynamic instability (hypotension and hypertension) and in extreme cases death. AIMS: The aim of this study was to evaluate current practice and identify the safest method for inotrope administration. METHODS: A series of three consecutive audits were undertaken to determine which 'Method' and 'Syringe Driver' were associated with the least adverse effects to patient blood pressure. RESULTS: The findings suggest that Modified Method 2, when used in conjunction with a high-risk syringe driver and guidelines, proved to be the safest method for 'double pumping' inotrope drugs. Modified Method 2 instructed nurses to: 'Run both syringe drivers together until a rise in systolic blood pressure is seen (>5 mmHg), then turn the near empty infusion off'. CONCLUSION: As a direct result of these audits, and the development of guidelines, inotrope administration practice on the unit has improved. PMID- 15288872 TI - Reclaiming the everyday world: how long-term ventilated patients in critical care seek to gain aspects of power and control over their environment. AB - Critical care nurses are increasingly seeking to base patient care on evidence derived from research studies. The purpose of this study was to explore the meanings former patients attributed to being on long-term mechanical ventilation in a critical care unit (CCU) in Australia. Using Heideggerian phenomenology, unstructured interviews were undertaken with nine participants. Data were analysed thematically using the method developed by van Manen. Thematic analysis revealed four major themes. This article presents the findings from the theme titled: Reclaiming the everyday world, which describes how the study participants gained comfort from the presence of nurses and their families, sought control over their treatments, and questioned and interpreted the environment, in order to reclaim self. The study highlighted the central role of nurses in patient care, and served as a basis for a number of recommendations, which include recognising the significant role of nurses and family in patient care, and being aware that patients may want more control over their environment and instigate ways to facilitate this. Further research is warranted to examine CCU patients' perceived level of control and power, and to investigate the extent and type of involvement CCU patients would like to have in their care. PMID- 15288873 TI - An analysis of information available to relatives in intensive care. AB - The Comprehensive Critical Care Review published by the Government in 2000 acknowledges that patients are part of family units and critical illness has an extended impact. It outlines information that should be provided to relatives and suggests recommendations be implemented within 3-5 years. The aim of this study was to gauge an overall view of provisions available across general Intensive Care Units in England for relatives, by conducting an analysis of information available and unit policies, and to see the extent that government guidelines have been adhered to. Two hundred and ten units were approached for copies of policy documents and leaflets. There was a 56% response rate. Results were collated and analysed for basic descriptive statistics using software package SPSS version 11.5. The Gunning's Fog Index was performed on 20% of leaflets to measure readability. All leaflets measured above the recommended level. Sixteen percent of units do not have a leaflet and therefore do not comply with the Department of Health recommendations. Huge variation exists nationally over the amount and quality of information that relatives have access to and receive. Only 9% of units had an official policy on how to deal with relatives. The implications of this are discussed. PMID- 15288874 TI - Screening for delirium in an adult intensive care unit. AB - Delirium is an acute, reversible disorder of attention and cognition and may be viewed as cerebral dysfunction similar to the failure of any other organ. The development of delirium is associated with increased morbidity and mortality, extended length-of-stay in the intensive care unit and longer time spent sedated and ventilated. Nearly every clinical, pharmacological and environmental factor present and necessary in the ICU setting has the potential to cause delirium. Since all of these factors cannot be removed, it is paramount to increase the awareness amongst health care professionals so as to minimise under-recognition and encourage future research into factors that may improve the long-term outcome for ICU patients. There is a need for user-friendly, validated assessment tools for the intubated and ventilated ICU patient, which can be applied at the time of ICU admission without the need for lengthy psychiatric assessment. Nursing professionals are at the forefront of those who are able to provide holistic care through meaningful conversation and empathetic touch. A 6-month Quality Improvement (QI) project screening patients for signs of delirium provided a foundation for discussion. All patients admitted to ICU for more than 72 h, with a hospital length-of-stay less than 96 h prior to ICU admission were screened. Patients admitted following neurological insults or with pre-existing altered mental state were excluded. The QI project showed the incidence of delirium to be 40% of the total sample (n = 73) in a mixed medical/surgical and elective/emergency patient population. PMID- 15288875 TI - An insight into Australian nurses' experience of withdrawal/withholding of treatment in the ICU. AB - BACKGROUND: The success of biotechnology has created moral and ethical dilemmas concerning end-of-life care in the Intensive Care Unit (ICU). Whilst the competent individual has the right to refuse or embrace treatment, ICU patients are rarely able to exercise this right. Thus, decision-making is left to medical professionals and family/significant others. AIM: This study aimed to explore the lived experience of ICU nurses caring for clients having treatment withdrawn or withheld, and increase awareness and understanding of this experience amongst other health professionals. METHODS: Van Manens' (1990) phenomenological framework formed the basis of this study as it provided an in-depth insight into the human experience. A convenience sample of ten ICU Nurses participated in the study. Conversations were transcribed verbatim and analysed using a process of thematic analysis. RESULTS: Five major themes emerged during the analysis. These were: (1) comfort and care, (2) tension and conflict, (3) do no harm, (4) nurse family relationships and (5) invisibility of grief and suffering. CONCLUSION: The experience of providing care for the adult having treatment withdrawn or withheld in the ICU represents a significant personal and professional struggle. Improvements in communication between health professionals, debriefing and education about the process of withdrawing or withholding treatment would be beneficial to both staff and families and has the potential to improve patient care and reduce burden on nurses. PMID- 15288876 TI - Reducing family members' anxiety and uncertainty in illness around transfer from intensive care: an intervention study. AB - INTRODUCTION: This intervention study examines anxiety and uncertainty in illness in families transferring from intensive care to a general ward. METHODS: The pre test, post-test design purposively allocated family members to a control (n = 80) and intervention group (n = 82). The intervention group experienced a structured individualised transfer method whereas the control group received existing ad hoc transfer methods. Families were surveyed before and after transfer. RESULTS: Families' uncertainty was significantly related to their state anxiety (P < 0.000), the relationship to the patient (P = 0.022), and the unexpected nature of patients' admission (P < 0.000). Anxiety increased significantly with reduced social support (P = 0.002). Following transfer, anxiety reduced significantly for both groups whereas uncertainty reduced significantly for the intervention group (P = 0.03). CONCLUSION: Families at the time of transfer experience uncertainty and anxiety, which are significantly related in this study. The intervention significantly reduced uncertainty scores. When the family member was a parent, when admissions were unexpected, and those with fewer social supports represent potential 'at risk' groups whose adaptation to transfer may limit their coping ability. The structured individualised method of transfer is recommended with further research of ICU families to further examine the dimension of uncertainty and how it affects patient outcomes. PMID- 15288877 TI - Clinical research 1: research questions and design. PMID- 15288878 TI - Ball C, Kirby M, Williams S. Effect of the critical care outreach team on patient survival to discharge from hospital and readmission to critical care: non randomised population based study. British Medical Journal 2003; 327:1014-1017. PMID- 15288880 TI - Fractional diastolic and systolic pressure in the ascending aorta are related to the extent of coronary artery disease. AB - BACKGROUND: Ascending aortic fractional pulse pressure and fractional systolic pressure (FSP) were demonstrated to differentiate patients with and without coronary artery disease. However, no study so far has analyzed the relationship between FSP and fractional diastolic pressure (FDP) and the extent of coronary artery disease. Therefore, we investigated the relationship between ascending aortic FSP and FDP and the extent of coronary atherosclerosis in unselected patients with angiographically confirmed coronary artery disease. METHODS: The study group consisted of 445 patients (350 men and 95 women, mean age 58.5 +/- 9.7 years) with angiographically confirmed coronary artery disease and ejection fraction > 55%. Invasive ascending aortic blood pressure during catheterization and conventional sphygmomanometer measurements were taken. RESULTS: Pulse pressure (PP), FSP, and FDP derived from intraaortic measurements differentiated patients with one-, two-, and three-vessel coronary artery disease (PP, 62.8 +/- 15.8 v 64.8 +/- 17.9 v 71.7 +/- 19.1 [P < .0001]; FSP, 1.45 +/- 0.09 v 1.46 +/- 0.10 v 1.51 +/- 0.12 [P < .0001]; FDP 0.77 +/- 0.05 v 0.77 +/- 0.05 v 0.75 +/ 0.06 [P < .0001]). After multivariate stepwise adjustment, the odds ratio (OR) and confidence interval (CI) of having three-vessel disease was as follows: PP per 10 mm Hg, OR = 1.15, 95% CI = 1.01 to 1.30; FSP per 0.1, OR = 1.28, 95% CI = 1.03 to 1.60; and FDP per 0.1, OR = 0.61, 95% CI = 0.39 to 0.95. None of the brachial pressure indices was independently related to the extent of coronary atherosclerosis. CONCLUSIONS: Pulse pressure (PP), FSP, and FDP of the ascending aorta are related to the risk of three-vessel disease in patients with coronary artery disease and preserved left ventricular function. PMID- 15288881 TI - Determinants of radial artery pulse wave analysis in asymptomatic individuals. AB - BACKGROUND: Noninvasive techniques to evaluate arterial stiffness include noninvasive radial artery pulse contour analysis. Diastolic pulse contour analysis provides a separate assessment of large (C1) and small artery (C2) elasticity. Analysis of the systolic pulse contour identifies two pressure peaks (P1 and P2) that relate to incident and reflected waves. This study aimed to compare indices from systolic and diastolic pulse contour analysis from the radial pressure waveform and to correlate these indices with traditional risk factors in asymptomatic individuals screened for cardiovascular disease. METHODS: In 298 consecutive subjects (206 male and 92 female healthy subjects with a mean age of 50 +/- 12 years), noninvasive radial artery pressure waveforms were acquired with a piezoelectric transducer and analyzed for 1) diastolic indices of C1 and C2 from the CR-2000 CVProfiler, and 2) systolic indices of augmentation as defined by augmentation pressure (AP), augmentation index (AIx), and systolic reflective index (SRI = P2/P1). These indices were then correlated to each other as well as to individual traditional risk factors and the Framingham Risk Score. RESULTS: Diastolic indices were significantly and inversely correlated to systolic indices with C2 showing a stronger inverse association than C1. C2 and Alx were significantly correlated with height, weight, and body mass index in men but not in women. All indices correlated better to blood pressure in women than men. In women, only systolic indices were significantly correlated to HDL cholesterol and only diastolic indices were significantly correlated to LDL cholesterol. All indices were significantly correlated to the Framingham Risk Score, which was stronger in women then men, but when adjusted for age only diastolic indices remained significant in women. CONCLUSIONS: Diastolic and systolic indices of pulse contour analysis correlate differently with traditional risk factors in men and women. PMID- 15288882 TI - Reduced arterial elasticity is associated with endothelial dysfunction in persons of advancing age: comparative study of noninvasive pulse wave analysis and laser Doppler blood flow measurement. AB - BACKGROUND: Endothelial dysfunction is the earliest marker for age-related abnormalities in vascular function, and examination of endothelial function has important clinical relevance. The present study was performed to evaluate effects of aging on arterial elasticity by using pulse waveform analysis and to investigate whether the changes in arterial elasticity might be used as a noninvasive measure for endothelial dysfunction. METHODS: A total of 24 healthy male volunteers were divided into young (n = 12) and elderly (n = 12) groups. Endothelial function was evaluated by delivering acetylcholine (Ach) and sodium nitroprusside (SNP) to the forearm vessels using iontophoresis, respectively, and measured blood flow using laser Doppler fluximetry. Large and small artery elasticity indices were noninvasively assessed using pulse wave analysis. RESULTS: Basal blood flow was similar between the young and elderly groups (14.58 +/- 3.4 v 13.52 +/- 3.41 PU, P = NS). Peak blood flow induced by Ach was significantly reduced in the elderly group compared with the young group (83.4 +/ 11.9 v 93.75 +/- 10.87 PU, P < .05). However, peak blood flow induced by SNP was similar in the two groups (119.17 +/- 16.76 v 128.33 +/- 21.29 PU, P = NS). In parallel, C1 large artery elasticity and C2 small artery elasticity indices were significantly reduced in the elderly group compared with the young group (11.42 +/- 1.67 v 16.75 +/- 2.09 mL/mm Hg x 10, P < .001; and 7.67 +/- 1.56 v 10.75 +/- 1.86 mL/mm Hg x 100, P < .001, respectively). The Ach-induced peak blood flow correlated with C1 large and C2 small artery elasticity indices. CONCLUSIONS: Advancing age is associated with endothelial dysfunction and reduced arterial elasticity. Reduced arterial elasticity parallels changes in impaired endothelium dependent vasodilation. It appears that reduced arterial elasticity may be used as a noninvasive measure for the determination of endothelial function. PMID- 15288883 TI - Morning blood pressure surge and hypertensive cerebrovascular disease: role of the alpha adrenergic sympathetic nervous system. AB - BACKGROUND: The morning surge of blood pressure (BP) is associated with alpha adrenergic activity. We studied the association between the alpha-adrenergic morning surge in BP and silent cerebrovascular disease in elderly patients with hypertension. METHODS: We conducted ambulatory BP monitoring three times (twice at baseline and after nighttime dosing of the alpha1-blocker doxazosin) in 98 elderly hypertensive patients in whom the presence of silent cerebral infarcts (SCI) was assessed by brain magnetic resonance imaging. The morning BP surge (MBPS) was calculated as the mean systolic BP during the 2 h after waking minus the mean systolic BP during 1 h that included the lowest sleep BP. The alpha adrenergic MBPS was calculated as the reduction of MBPS by doxazosin. RESULTS: The prevalence of multiple SCI was higher in the Surge group (top quartile: MBPS > or = 45 mm Hg, n = 24) than in the Nonsurge group (MBPS < 45 mm Hg, n = 74) (54% v 31%, P = .04), and in the higher alpha-adrenergic surge group (top quartile: alpha-adrenergic MBPS > or = 28 mm Hg, n = 25) than in the lower alpha adrenergic surge group (< 28 mm Hg, n = 73) (68% v 26%, P < .0001). In the Surge group, subjects with higher alpha-adrenergic surge (n = 17) had a markedly higher frequency of multiple SCI, whereas none in the lower alpha-adrenergic surge group had multiple SCI (n = 7) (77% v 0%, P = .001). The alpha-adrenergic MBPS was closely associated with multiple SCI (10 mm Hg increase: OR = 1.96, P = .006), independently of age, MBPS, 24-h systolic BP, and other confounding factors. CONCLUSION: The morning BP surge, particularly that dependent on alpha-adrenergic activity, is closely associated with advanced silent hypertensive cerebrovascular disease in elderly individuals. PMID- 15288884 TI - Roles of angiotensin type 1 and 2 receptors in pregnancy-associated blood pressure change. AB - BACKGROUND: Activation of the renin-angiotensin system with increased levels of renin and angiotensin (Ang) II in pregnancy has been reported, but the vascular responsiveness to Ang II seems to be decreased, thereby keeping maternal blood pressure (BP) constant. We postulated that the balance of angiotensin type 1 (AT1) and angiotensin type 2 (AT2) receptor expression, which would exert antagonistic actions on vasoconstriction and cell growth, might control BP in pregnancy. METHODS: Using wild type (C57BL/6J), AT1a receptor null and AT2 receptor null mice, we examined the changes in BP, expression and localization of AT1 and AT2 receptors in placenta, umbilical cord, and uterus by immunohistochemical staining and urinary albumin measurement during pregnancy. RESULTS: Wild type mice did not show any significant change in BP throughout pregnancy. The BP in AT1a receptor null mice declined significantly in the second trimester of pregnancy, whereas BP in AT2 receptor null mice increased significantly in the third trimester. We did not observe any significant differences in albuminuria, litter size, or body weight of neonates among the three groups. Vascular smooth muscle cells in blood vessels of the umbilical cord and placenta specifically expressed AT2 receptors, which are minimally expressed in adult vessels. In contrast, AT1 receptors were dominantly expressed in the cytotrophoblast and chorionic plate as well as blood vessels in placenta and umbilical cord. CONCLUSIONS: Our results suggested that disturbance of the balance of the AT1 and AT2 receptors could trigger pregnancy induced hypertension. PMID- 15288885 TI - Randomized controlled trial of sour milk on blood pressure in borderline hypertensive men. AB - BACKGROUND: A double-blind randomized controlled trial was carried out to assess the effect of sour milk, containing two tripeptides (valine-proline-proline and isoleucine-proline-proline), on blood pressure (BP). METHODS: A total of 46 borderline hypertensive men aged 23 to 59 years were recruited at their workplace for this trial. Subjects were randomly allocated into two groups; sour milk drink group (S-group, n = 23) and placebo (acidified milk) drink group (P-group, n = 23) for 4 weeks. Blood pressure was measured twice at each occasion by a physician, at the health center of the company, with a mercury at baseline, 2 and 4 weeks. Statistical analysis was performed by SPSS 10.0J. RESULTS: The S-group and P-group showed no significant difference in baseline systolic BP (mean [SD], S: 147.6 [9.6], P: 145.3 [13.0]) or diastolic BP (S: 95.3 [9.9], P: 91.5 [9.6]). In the S-group, change in systolic BP at 2 and 4 weeks were -4.3 mm Hg (95% confidence interval [CI] -8.3 to -0.4; P = .032) and -5.2 mm Hg (95% CI -10.1 to 0.3; P = .039), both statistically significant. Diastolic BP showed change from 1.7 mm Hg (95% CI -5.4 to 2.0) at 2 weeks and -2.0 (95% CI -5.4 to 1.5) at 4 weeks, respectively. In the P-group, change in systolic BP were -0.5 (95% CI -5.8 to 4.8) at 2 weeks and -3.7 (95% CI -8.3 to 0.9) and change in diastolic BP were 0.6 (95% CI -4.7 to 3.6) and -0.3 (95% CI -3.9 to 3.3), which were not statistically significant. CONCLUSIONS: This trial demonstrated the beneficial effect of sour milk on BP in borderline hypertensive men who were not taking antihypertensive medication. PMID- 15288886 TI - Cyclic strain stimulates L-proline transport in vascular smooth muscle cells. AB - BACKGROUND: The increase in vessel wall strain in hypertension contributes to arterial remodeling by stimulating vascular smooth muscle cell (SMC) proliferation and collagen synthesis. Because L-proline is essential for the synthesis of collagen and cell growth, we examined whether cyclic strain regulates the transcellular transport of L-proline by vascular SMC. METHODS: Cultured rat aortic SMCs were subjected to mechanical strain using the Flexercell 3000 Strain Unit. RESULTS: Cyclic strain increased L-proline transport in a time- and strain-degree-dependent manner that was inhibited by cycloheximide or actinomycin D. Kinetic studies indicated that cyclic strain-induced L-proline uptake was mediated by an increase in transport capacity independent of any change in the affinity for L-proline. Cyclic strain stimulated the expression of system A amino acid transporter 2 mRNA in a time-dependent fashion that paralleled the increase in L-proline transport. Cyclic strain also induced the release of transforming growth factor-beta1 in a time- and strain-dependent manner. Moreover, conditioned media from SMCs exposed to cyclic strain stimulated the transport of L-proline in control, static SMCs and this was significantly attenuated by a transforming growth factor-beta1 neutralizing antibody. CONCLUSIONS: These results demonstrate that cyclic strain stimulates L-proline transport by inducing system A amino acid transporter 2 gene expression through the autocrine release of transforming growth factor-beta1. The ability of cyclic strain to induce system A amino acid transporter 2 expression may promote arterial remodeling in hypertension by providing vascular SMCs with the necessary intracellular levels of L-proline required for collagen synthesis and cell growth. PMID- 15288887 TI - A new view of the primary visual cortex. PMID- 15288888 TI - Feedforward, feedback and inhibitory connections in primate visual cortex. AB - Visual cortical circuits are organized at multiple levels of complexity including cortical areas, layers and columns, and specific cell types within these modules. Making sense of the functions of these circuits from anatomical observations requires linking these circuits to function at each of these levels of complexity. Observations of these relationships have become increasingly sophisticated over the last several decades, beginning with correlations between the connectivities and functions of various visual cortical areas and progressing toward cell type-specificity. These studies have informed current views about the functional interactions between cortical areas and modules and the mechanisms by which fine scale microcircuits influence interactions at more coarse levels of organization. PMID- 15288889 TI - Electrophysiological classes of neocortical neurons. AB - Neocortical network behavior and neocortical function emerge from synaptic interactions among neurons with specific electrophysiological and morphological characteristics. The intrinsic electrophysiological properties of neurons define their firing patterns and their input-output functions with critical consequences for their functional properties within the network. Understanding the role played by the active non-linear properties caused by ionic conductances distributed in the soma and the dendrites is a critical step towards understanding cortical function. Here I present a brief description of electrophysiological and morphological characteristics of neocortical cells that allow their classification in categories. I review some examples of differences in functional properties among different electrophysiological cell classes in the visual cortex, as well as the role played by specific ionic conductances in defining firing and accommodation properties of neocortical neurons. PMID- 15288890 TI - A simple cell model with dominating opponent inhibition for robust image processing. AB - The extraction of oriented contrast information by cortical simple cells is a fundamental step in early visual processing. The orientation selectivity originates at least partly from the input of lateral geniculate nuclei neurons with properly aligned receptive fields. In the present article, we investigate the feedforward interactions between on- and off-pathways. Based on physiological evidence we propose a push-pull model with dominating opponent inhibition (DOI). We show that the model can account for empirical data on simple cells, such as contrast-invariant orientation tuning, sharpening of orientation tuning with increasing inhibition, and strong response decrements to stimuli with luminance gradient reversal. With identical parameter settings, we apply the model for the processing of synthetic and real world images. We show that the model with DOI can robustly extract oriented contrast information from noisy input. More important, noise is adaptively suppressed, i.e. the model simple cells do not respond to homogeneous regions of different noise levels, while remaining sensitive to small contrast changes. The image processing results reveal a possible functional role of the strong inhibition as observed empirically, namely to adaptively suppress responses to noisy input. PMID- 15288891 TI - Nonlinear V1 responses to natural scenes revealed by neural network analysis. AB - A key goal in the study of visual processing is to obtain a comprehensive description of the relationship between visual stimuli and neuronal responses. One way to guide the search for models is to use a general nonparametric regression algorithm, such as a neural network. We have developed a multilayer feed-forward network algorithm that can be used to characterize nonlinear stimulus-response mapping functions of neurons in primary visual cortex (area V1) using natural image stimuli. The network is capable of extracting several known V1 response properties such as: orientation and spatial frequency tuning, the spatial phase invariance of complex cells, and direction selectivity. We present details of a method for training networks and visualizing their properties. We also compare how well conventional explicit models and those developed using neural networks can predict novel responses to natural scenes. PMID- 15288892 TI - The contribution of vertical and horizontal connections to the receptive field center and surround in V1. AB - Here we review the results of anatomical and physiological studies in tree shrew visual cortex which focus on the contribution of vertical and horizontal inputs to receptive field center and surround properties of layer 2/3 neurons. A fundamental feature of both sets of connections is the arrangement of axon arbors in a fashion that respects both the orientation preference and retinotopic displacement of the target site. As a result, layer 2/3 neurons receive convergent input from populations of layer 4 and other layer 2/3 neurons whose receptive fields are displaced along an axis in visual space that corresponds to their preferred orientation. Although, horizontal connections extend for greater distances across the cortical surface than vertical connections, the majority of these inputs link neurons with overlapping receptive fields, emphasizing that both feed-forward and recurrent circuits are likely to play a constructive role in generating properties (such as orientation selectivity) that define the receptive field center. Both within and beyond the dimensions of the receptive field center, the distribution of horizontal connections accords remarkably well with the magnitude and axial tuning of length summation effects. Taken together, these results suggest a continuum of functional properties that transcends the traditional designation of receptive field center and surround. By extension, we suggest that the perceptual effects of stimulus context may arise from stimulus interactions within the receptive field center as well as between center and surround. PMID- 15288893 TI - Perceptual grouping and the interactions between visual cortical areas. AB - Visual perception involves the grouping of individual elements into coherent patterns, such as object representations, that reduce the descriptive complexity of a visual scene. The computational and physiological bases of this perceptual remain poorly understood. We discuss recent fMRI evidence from our laboratory where we measured activity in a higher object processing area (LOC), and in primary visual cortex (V1) in response to visual elements that were either grouped into objects or randomly arranged. We observed significant activity increases in the LOC and concurrent reductions of activity in V1 when elements formed coherent shapes, suggesting that activity in early visual areas is reduced as a result of grouping processes performed in higher areas. In light of these results we review related empirical findings of context-dependent changes in activity, recent neurophysiology research related to cortical feedback, and computational models that incorporate feedback operations. We suggest that feedback from high-level visual areas reduces activity in lower areas in order to simplify the description of a visual image-consistent with both predictive coding models of perception and probabilistic notions of 'explaining away.' PMID- 15288894 TI - A model of contextual interactions and contour detection in primary visual cortex. AB - A new model of contour extraction and perceptual grouping in the primary visual cortex is presented and discussed. It differs from previous models since it incorporates four main mechanisms, according to recent physiological data: a feed forward input from the lateral geniculate nucleus, characterized by Gabor elongated receptive fields; an inhibitory feed-forward input, maximally oriented in the orthogonal direction of the target cell, which suppresses non-optimal stimuli and warrants contrast invariance; an excitatory cortical feedback, which respects co-axial and co-modularity criteria; and a long-range isotropic feedback inhibition. Model behavior has been tested on artificial images with contours of different curvatures, in the presence of considerable noise or in the presence of broken contours, and on a few real images. A sensitivity analysis has also been performed on the role of intracortical synapses. Results show that the model can extract correct contours within acceptable time from image presentation (30-40 ms). The feed-forward input plays a major role to set an initial correct bias for the subsequent feedback and to ensure contrast-invariance. Long-range inhibition is essential to suppress noise, but it may suppress small contours due to excessive competition with greater contours. Cortical excitation sharpens the initial bias and improves saliency of the contours. Model results support the idea that contour extraction is one the primary steps in the visual processing stream, and that local processing in V1 is able to solve this task even in difficult conditions, without the participation of higher visual centers. PMID- 15288895 TI - Using afterimages to test neural mechanisms for perceptual filling-in. AB - Many theories of visual perception propose that brightness information spreads from edges to define the perceived intensity of the interior of visual surfaces. Several theories of visual perception have hypothesized that this filling-in process is similar to a diffusion of information where the signals coding brightness spread to nearest neighbors. This paper shows that diffusive mechanisms fail to account for the characteristics of certain afterimage percepts that seem to be dependent on the filling-in process. A psychophysical experiment tests a key property of diffusion-based filling-in mechanisms and finds data that rejects this class of models. A non-diffusive based filling-in mechanism is proposed and is shown to act much like the diffusive based mechanism in many instances, but also produces afterimage percepts that match the experimental data. PMID- 15288896 TI - Hue geometry and horizontal connections. AB - Primate visual systems support an elaborate specialization for processing color information. Concentrating on the hue component, we observe that, contrary to Mondrian-like assumptions, hue varies in a smooth manner for ecologically important natural imagery. To represent these smooth variations, and to support those information processing tasks that utilize hue, a piecewise smooth hue field is postulated. The geometry of hue-patch interactions is developed analogously to orientation-patch interactions in texture. The result is a model for long-range (horizontal) interactions in the color domain, the power of which is demonstrated on a number of examples. Implications for computer image processing, computer vision, visual neurophysiology and psychophysics are discussed. PMID- 15288898 TI - A neuromorphic model for achromatic and chromatic surface representation of natural images. AB - This study develops a neuromorphic model of human lightness perception that is inspired by how the mammalian visual system is designed for this function. It is known that biological visual representations can adapt to a billion-fold change in luminance. How such a system determines absolute lightness under varying illumination conditions to generate a consistent interpretation of surface lightness remains an unsolved problem. Such a process, called 'anchoring' of lightness, has properties including articulation, insulation, configuration, and area effects. The model quantitatively simulates such psychophysical lightness data, as well as other data such as discounting the illuminant, and lightness constancy and contrast effects. The model retina embodies gain control at retinal photoreceptors, and spatial contrast adaptation at the negative feedback circuit between mechanisms that model the inner segment of photoreceptors and interacting horizontal cells. The model can thereby adjust its sensitivity to input intensities ranging from dim moonlight to dazzling sunlight. A new anchoring mechanism, called the Blurred-Highest-Luminance-As-White rule, helps simulate how surface lightness becomes sensitive to the spatial scale of objects in a scene. The model is also able to process natural color images under variable lighting conditions, and is compared with the popular RETINEX model. PMID- 15288897 TI - A theory of the Benham Top based on center-surround interactions in the parvocellular pathway. AB - A model color-opponent neuron was used to investigate the subjective colors evoked by the Benham Top (BT). Color-opponent inputs from cone-selective parvocellular (P) pathway neurons with center-surround receptive fields were subtracted with a short relative delay, yielding a small transient input in response to a white spot. This transient input was amplified by BT-like stimuli, modeled as a thin dark bar followed by full-field illumination. The narrow bar produced maximal activation of the P-pathway surrounds but only partial activation of the P-pathway centers. Due to saturation, subsequent removal of the bar had little effect on the P-pathway surrounds, whereas the transient input from the P-pathway centers was amplified via disinhibition. Responses to BT-like stimuli became weaker as surround sensitivity recovered, producing an effect analogous to the progression of perceived BT colors. Our results suggest that the BT-illusion arises because cone-selective neurons convey information about both color and luminance contrast, allowing the two signals become confounded. PMID- 15288899 TI - Integration of form and motion within a generative model of visual cortex. AB - One of the challenges faced by the visual system is integrating cues within and across processing streams for inferring scene properties and structure. This is particularly apparent in the inference of object motion, where psychophysical experiments have shown that integration of motion signals, distributed across space, must also be integrated with form cues. This has led several to conclude that there exist mechanisms which enable form cues to 'veto' or completely suppress ambiguous motion signals. We describe a probabilistic approach which uses a generative network model for integrating form and motion cues using the machinery of belief propagation and Bayesian inference. We show, using computer simulations, that motion integration can be mediated via a local, probabilistic representation of contour ownership, which we have previously termed 'direction of figure'. The uncertainty of this inferred form cue is used to modulate the covariance matrix of network nodes representing local motion estimates in the motion stream. We show with results for two sets of stimuli that the model does not completely suppress ambiguous cues, but instead integrates them in a way that is a function of their underlying uncertainty. The result is that the model can account for the continuum of bias seen for motion coherence and perceived object motion in psychophysical experiments. PMID- 15288900 TI - Associative learning in early vision. AB - Sensory discriminations often improve with practice (perceptual learning). Recent results show that practice does not necessarily lead to the best possible performance on the task. It was shown that learning a task (contrast discrimination) that has already reached saturation could be enabled by a contextual change in the stimulus (the addition of surrounding flankers) during practice. Psychophysical results with varying context show a behavior that is described by a network of local visual processors with horizontal recurrent interactions. We describe a mathematical learning rule for the modification of cortical synapses that is inspired by the experimental results and apply it to recurrent cortical networks that respond to external stimuli. The model predicts that repeated presentation of the same stimulus leads to saturation of synaptic modification, such that the strengths of recurrent connections depend on the configuration of the stimulus but not on its amplitude. When a new stimulus is introduced, the modification is rekindled until a new equilibrium is reached. This effect may explain the saturation of perceptual learning when practicing a certain task repeatedly. We present simulations of contrast discrimination in a simplified model of a cortical column in the primary visual cortex and show that performance of the model is reminiscent of context-dependent perceptual learning. PMID- 15288901 TI - Recognition invariance obtained by extended and invariant features. AB - In performing recognition, the visual system shows a remarkable capacity to distinguish between significant and immaterial image changes, to learn from examples to recognize new classes of objects, and to generalize from known to novel objects. Here we focus on one aspect of this problem, the ability to recognize novel objects from different viewing directions. This problem of view invariant recognition is difficult because the image of an object seen from a novel viewing direction can be substantially different from all previously seen images of the same object. We describe an approach to view-invariant recognition that uses extended features to generalize across changes in viewing directions. Extended features are equivalence classes of informative image fragments, which represent object parts under different viewing conditions. This representation is extracted during learning from images of moving objects, and it allows the visual system to generalize from a single view of a novel object, and to compensate for large changes in the viewing direction, without using three-dimensional information. We describe the model, its implementation and performance on natural face images, compare it to alternative approaches, discuss its biological plausibility, and its extension to other aspects of visual recognition. The results of the study suggest that the capacity of the recognition system to generalize to novel conditions in an efficient and flexible manner depends on the ongoing extraction of different families of informative features, acquired for different tasks and different object classes. PMID- 15288902 TI - Attentional states and the degree of visual adaptation to gratings. AB - We studied top-down attentional effects on adaptation to two aspects of sinusoidal gratings: contrast (CTE: contrast threshold elevation for detection) and orientation (TAE: tilt aftereffect, bias in perceived orientation). Adaptation was examined under five different behavioral conditions designed to assess the effect of alertness, spatial attention and the dimension attended. Alertness increased CTE, but had no effect on TAE. Spatial attention increased TAE, but had no effect on CTE. TAE (but not CTE) was also sensitive to the attended dimension. It was greater when gratings' contrast rather than orientation was attended. The different patterns of top-down effects on CTE compared with TAE are consistent with these two types of adaptation taking place at different levels along the visual hierarchy: CTE occurs at very low-levels, where activity is affected by alertness but not by spatial attention, whereas TAE occurs at subsequent stages, which are modulated by selective attention. PMID- 15288903 TI - Separate neural definitions of visual consciousness and visual attention; a case for phenomenal awareness. AB - What is the relation between visual attention and visual awareness? It is difficult to imagine being aware of something without attending to it, and by some, visual consciousness is simply equated to what is in the focus of attention. However, findings from psychological as well as from neurophysiological experiments argue strongly against equating attention and visual consciousness. From these experiments clearly separate neural definitions of visual attention and visual consciousness emerge. In the model proposed here, visual attention is defined as a convolution of sensori-motor processing with memory. Consciousness, however, is generated by recurrent activity between cortical areas. The extent to which these recurrent interactions involve areas in executive or mnemonic space depends on attention and determines whether a conscious report is possible about the sensory experience, not whether the sensory experience is there. This way, a strong case can be made for a pure non cognitive form of seeing, independent of attentional selection, called phenomenal awareness. This can be dissociated from the reportable form, depending on attention, called access awareness. The hypothesis explains why attention and consciousness seem so intricately related, even though they are fully separate phenomena. PMID- 15288904 TI - A model of active visual search with object-based attention guiding scan paths. AB - When a monkey searches for a colour and orientation feature conjunction target, the scan path is guided to target coloured locations in preference to locations containing the target orientation [Vision Res. 38 (1998b) 1805]. An active vision model, using biased competition, is able to replicate this behaviour. As object based attention develops in extrastriate cortex, featural information is passed to posterior parietal cortex (LIP), enabling it to represent behaviourally relevant locations [J. Neurophysiol. 76 (1996) 2841] and guide the scan path. Attention evolves from an early spatial effect to being object-based later in the response of the model neurons, as has been observed in monkey single cell recordings. This is the first model to reproduce these effects with temporal precision and is reported here at the systems level allowing the replication of psychophysical scan paths. PMID- 15288905 TI - Who will train the trainers? PMID- 15288906 TI - Sitting habits in elementary schoolchildren: a traditional versus a "Moving school". AB - This study evaluated differences in sitting habits in the classroom between the project "Moving school" and a traditional school in 8-year-old children. Twenty two children, since 1.5 years involved in the project were compared to 25 children in a traditional school. Making use of the Portable Ergonomic Observation (PEO) method, it was observed that children from a traditional school spend an average of 97% of the lesson time sitting statically, from which one third with the trunk bend over 45 degrees. In the "Moving school" this posture was replaced by dynamic sitting (53%), standing (31%) and walking around (10%), while trunk flexion over 45 degrees was nearly not observed. Children from the "Moving school" also showed significantly less neck and trunk rotation. Additionally, accelerometric data showed significantly more physical activity in lessons of the "Moving school". Rates of self-reported back or neck pain did not differ significantly between both study groups. Results show that sitting habits are more favourable in a "Moving school". Further research is needed to study the impact of implementing "Moving school" concepts in traditional schools on sitting habits. PMID- 15288908 TI - Meeting between experts: evaluation of the first UK forum for lay and professional experts in intersex. AB - At present the clinical management of intersex is in turmoil. The policy of non disclosure of diagnosis is widespread and cosmetic genital surgery is routinely performed on infants throughout the world. Some clinicians feel such practices are in the interest of the intersex child and the family, but some intersex adults are calling for a moratorium on sex assignment genital surgery. These widely opposing views have led to distrust between groups. One way to begin to address these critical issues is to facilitate dialogue with equal input from clinicians and intersex people and families. Clinicians are experts by training, but patients and families are experts through lived experiences. Our paper reports the rationale, process and outcome of the first UK forum bringing together these different experts to address some of the most complex issues in clinical services. In communicating our experiences, we hope that it will provide a useful reference point for those seeking similar service-user/provider collaboration in other areas of medicine. PMID- 15288907 TI - Measuring patient knowledge of the risks and benefits of prostate cancer screening. AB - This manuscript describes the development and validation of measures assessing patient knowledge about the risks and benefits of prostate cancer (CaP) screening. The measures described include a 10-item knowledge index and four single-item measures, used in previous studies, that assess knowledge of: CaP natural history and treatment efficacy, expert disagreement over the value of CaP screening, and the accuracy of the prostate specific antigen (PSA) test for CaP. We assessed the validity and reliability of these measures on a sample of 1152 male veteran patients age 50 and older. All knowledge index items had acceptable levels of discrimination, difficulty, and reliability. The index demonstrated strong evidence for construct and criterion validity. Much weaker validity evidence was found for the four single-item knowledge questions. The 10-item index developed in this study provides a valid and reliable tool for assessing patient knowledge of the risks and benefits of CaP screening. PMID- 15288909 TI - Copying referral letters to patients: prepare for change. AB - The National Health Service (NHS) Plan for England has directed that from April 2004 clinicians will offer patients the opportunity to receive copies of letters that are written about them. Patients like to have more information and patients who have received copies of letters have found them useful. It is hoped that copying letters will improve relationships between doctors and patients, encourage patients to be better informed, and improve the quality of information provided to patients. Relatively little empirical research has been performed in this area but what exists is generally supportive. Attention will need to be paid to issues of confidentiality, the language and content of letters, and individuals who may have difficulty obtaining information from letters. This initiative is one of many that the NHS has introduced to enhance openness, honesty and the quality of information provided to patients. PMID- 15288910 TI - Patient-centred communication: videotaped consultations. AB - The aim of this study is to analyse the way in which orthopaedic physicians manage consultations, and to identify those factors associated with patient experienced satisfaction/dissatisfaction. This was explored both using a descriptive method and by analysing comments from patients. Consultations were videotaped; 18 physicians and 18 patients participated. Approximately 1 week after the consultation, the patient was shown the video recording and asked for his/her points of view and spontaneous reactions. Each time, the patient wished to say something, the video was stopped and the comments recorded. According to the patients' comments of the videotaped consultations four consultations were mainly positive, seven negative and seven neither completely positive nor completely negative. We analysed the positive and negative consultations using the Consultation Map (CM) method. The pattern in the positive consultations shows a greater flexibility. Statements regarding initial history and aetiology often move to and fro between other items, and the consultation as a whole was often characterised by this rapid change between items. The pattern in the negative consultations seems to indicate a slower motion; with longer time spent with few items and fewer items covered. The positive consultations were characterised by a greater prevalence of the items 'Sharing Understanding' and 'Involving the Patient in Management'. On the other hand, the negative consultations were characterised by more time spent with the items 'History of problems', and 'Patient Ideas'. This might be due to the patient having tried to express him/herself in order to present his/her views but the physician not following them up. In this study, the CM has been helpful in clarifying the difference between encounters experienced as satisfactory or dissatisfactory. PMID- 15288911 TI - Feelings related to first patient experiences in medical school. A qualitative study on students' personal portfolios. AB - Feelings and thoughts of medical students related to first patient experiences during the first clinical year were examined. Twenty-two volunteer third and fourth year medical students (15 women and 7 men) of the University of Helsinki participated in a portfolio course for 1 year. Their reflective learning diaries and writings on specific themes were analyzed by qualitative content analysis. First patient encounters were strong emotional experiences for medical students. The first patient examination was often described as an anxiety-provoking and confusing incident. Other emotionally significant encounters included helplessness when faced with serious illness and death, and role confusion when examining patients of one's own age but opposite sex. Students felt guilty for using patients for their own learning purposes. Portfolios as learning tools may help in recognizing key experiences and support professional development of medical students. PMID- 15288912 TI - The information-seeking behaviours of partners of men with prostate cancer: a qualitative pilot study. AB - This pilot study explores in depth the information-seeking behaviours of partners of men with prostate cancer. Six men with prostate cancer and their partners participated in one mini focus group discussion or four couple interviews. Theme analysis by two independent analysts produced three related themes: partners' information-seeking behaviours; partners' information-avoiding behaviours; and the conflict between seeking and avoiding information. The information-seeking behaviours of partners were individualistic, with some partners seeking voluminous information and others avoiding information. Partners sought information to help reduce their feelings of anxiety and uncertainty, to help them participate in the decision-making process, to help them care for their partner and to ensure that they had their information needs met. Partners avoided information to reduce their levels of fear and worry and to maintain a sense of normality. They failed to seek information from healthcare professionals because they felt disempowered and pressurised for time during patient-physician consultations. The information-seeking behaviours of partners changed over time and across situations and their behaviours were sometimes different from those of their partners (the patients), with some partners exhibiting more information seeking behaviour than patients. The findings within each of these themes and their practice implications are discussed in this paper. PMID- 15288913 TI - Using information technology for patient education: realizing surplus value? AB - Computer-based patient information systems are introduced to replace traditional forms of patient education like brochures, leaflets, videotapes and, to a certain extent, face-to-face communication. In this paper, we claim that though computer based patient information systems potentially have many advantages compared to traditional means, the surplus value of these systems is much harder to realize than often expected. By reporting on two computer-based patient information systems, both found to be unsuccessful, we will show that building computer-based patient information systems for patient education requires a thorough analysis of the advantages and limitations of IT compared to traditional forms of patient education. When this condition is fulfilled, however, these systems have the potential to improve health status and to be a valuable supplement to (rather than a substitute for) traditional means of patient education. PMID- 15288914 TI - The patients' written word: a simple communication aid. AB - Effective doctor-patient communication is an essential part of good medical practice. Question asking is one way for patients to have greater participation. In this feasibility study, patients were given the opportunity to list questions or discussion topics on a proforma before seeing the doctor in an oncology outpatient clinic. The items listed were reviewed by the clinic doctor. Eighty eight of 100 patients approached agreed to participate. Biomedical questions (mean 2.0 per patient) predominated over psychosocial ones (mean 0.1 per patient). Possible reasons for this are the study researcher being a doctor, patients attending for test results or that patients consider the format of listing questions on a proforma inappropriate for psychosocial issues. The listing of questions and discussion topics by patients gave the doctor valuable information about the patients' understanding and use of language. While the importance of doctors being adequately trained in communication skills should not be ignored, this simple intervention has potential for improving the outcome of clinic consultations. PMID- 15288915 TI - Evaluation of the Dutch AIDS information helpline: an investigation of information needs and satisfaction of callers. AB - AIDS telephone hotlines have an important function in AIDS education, HIV prevention and counselling. In this study, consults of the Dutch AIDS information helpline were evaluated to determine the AIDS information needs of the callers and callers' satisfaction with the telephone-delivered information and consultation. Immediately after their telephone consult, callers (N = 309) were redirected to co-workers of an independent telephone survey. They participated in an interview on content and evaluation of the telephone consult. This study shows that most telephone calls concerned questions about personal risk of HIV transmission, HIV transmission in general and HIV testing. Callers were very satisfied with the services of the helpline. Furthermore, helpline employees' counselling and conversation skills were evaluated very positively. These results are discussed within the scope of the professional organization of the Dutch AIDS information helpline. PMID- 15288917 TI - A participatory process for developing quality assurance tools for health education programs. AB - There is a constant need to find methods and tools for achieving high quality in health education and health promotion programs. The aim of this initiative was to develop standards and criteria for quality assessment of health education programs. Health educators participated in a process to develop these standards and criteria. A consensus was reached regarding the list of standards by participation of the health educators in two workshops. This list consisted of the three main stages of health education programs: planning, implementation and evaluation. Operational criteria for each standard were developed in order to be able to measure the standards. Four different subjects were chosen for the development of the criteria: menopause, smoking cessation, cardiac rehabilitation and prenatal care. A consensus regarding these criteria was reached by discussions in workshops. The tools developed were tested by interviewing health educators who were planning and running these programs. The tools built were found acceptable by the health educators as they provided information on the quality of the programs. PMID- 15288916 TI - Validation of the EORTC QLQ-SAT32 cancer inpatient satisfaction questionnaire by self- versus interview-assessment comparison. AB - Doubts are expressed on the validity of patient satisfaction questionnaires. High satisfaction levels are consistently reported. Within the European Organisation for Research and Treatment of Cancer (EORTC) quality of life group, we developed a cancer inpatient satisfaction questionnaire (QLQ-SAT32), adopting several precautions to overcome the ceiling effect commonly reported in satisfaction ratings. Since patients are often more critical when expressing themselves in an interview, in order to study the validity of the QLQ-SAT32, we assessed the agreement between self- and interview-administered QLQ-SAT32 responses. One hundred and twenty three patients were asked to complete the QLQ-SAT32 at home within 2 weeks of hospital discharge and were randomly allocated to participate in a telephone interview-administration of the QLQ-SAT32, either before or after self-completing the QLQ-SAT32. One hundred and four of them completed both modalities of questionnaire administration. Correlation and agreement between self- and interview-administered QLQ-SAT32 ratings were examined, for each subscale and the general satisfaction item of the QLQ-SAT32, using Spearman correlation, intra-class correlation coefficients (ICC) or weighted kappa coefficients. Agreement showed excellent for the doctors' and nurses' subscales and satisfactory for the services' subscale and the general satisfaction single item. The telephone interview-administration modality did not prove an adequate procedure to assess the validity of a patient satisfaction questionnaire. PMID- 15288918 TI - A focus group study of health professionals' views on phantom sensation, phantom pain and the need for patient information. AB - Focus groups with 32 health professionals from pre- and post-amputation care in central Scotland were used to explore perceptions of phantom sensation and pain, and current practice and potential improvements to patient information. Findings were compared to our parallel study of patients' experiences of phantom phenomena and information needs. Professionals' perceptions of phantom phenomena did not always match patient experiences: few professionals were fully aware of the nature, or the problems associated with phantom pain. There was uncertainty about who provided information and reported information was inconsistent and only weakly grounded in theory and mechanism-based management. Whilst there was awareness of the benefits of information, content, mode of delivery and co ordination were all identified as areas for improvement. Our findings suggest that the information given to patients on phantom phenomena is inconsistent and insufficient. Possible solutions are the development of minimum standards of information and specifically targeted interprofessional education. PMID- 15288919 TI - Consultation in general practice: a standard operating procedure? AB - The objectives of this study were to describe the features of consultation within general practice with special attention to the differences between short, moderate and long consultations. An analysis of 2801 videotaped consultations of 183 General Practitioners from six countries participating in the Eurocommunication Study was made. The communicative behaviour was gauged by means of the Roter Interaction Analysis System. The consultation can be seen as a "standard operating procedure" consisting of 8% social behaviour, 15% agreement, 4% rapport building, 10% partnership building, 11% giving directions, 28% giving information, 14% asking questions and 7% counselling. A short consultation can be described as an encounter with a little bit of social behaviour to set the contact, medical questioning, giving directions for the further consultation and advises in order to solve the problem(s) mentioned. In a long consultation doctors take more time for a social talk, they give more attention to the relation or contact with the patient, they listen more extensively, especially to psychosocial problems, and they give more information. PMID- 15288920 TI - Development of a questionnaire to evaluate the information needs of cancer patients: the EORTC questionnaire. AB - Information disclosure is one of the key areas of support that patients may receive. The European Organisation for Research and Treatment of Cancer (EORTC) Quality of Life Group has a group working on the development of a questionnaire that evaluates information received by cancer patients at different stages of their disease--EORTC QLQ-INFO30. This instrument is being developed by professionals from most European regions. The questionnaire aims to evaluate the information received by cancer patients on different areas of the disease, diagnosis, treatment and care. Besides, the instrument also assesses qualitative aspects of the information they have received. The present paper presents the first two phases of the module development process that include literature review, interviews with patients and professionals, and formulation of the items. All these steps have been carried out in different countries and have been approved by the EORTC QLG. PMID- 15288921 TI - Coronavirus infection of spotted hyenas in the Serengeti ecosystem. AB - Sera from 38 free-ranging spotted hyenas (Crocuta crocuta) in the Serengeti ecosystem, Tanzania, were screened for exposure to coronavirus of antigenic group 1. An immunofluorescence assay indicated high levels of exposure to coronavirus among Serengeti hyenas: 95% when considering sera with titer levels of > or = 1:10 and 74% when considering sera with titer levels of > or = 1:40. Cubs had generally lower mean titer levels than adults. Exposure among Serengeti hyenas to coronavirus was also confirmed by a serum neutralisation assay and an ELISA. Application of RT-PCR to 27 fecal samples revealed viral RNA in three samples (11%). All three positive fecal samples were from the 15 juvenile animals (<24 months of age) sampled, and none from the 12 adults sampled. No viral RNA was detected in tissue samples (lymph node, intestine, lung) from 11 individuals. Sequencing of two amplified products from the S protein gene of a positive sample revealed the presence of coronavirus specific RNA with a sequence homology to canine coronavirus of 76 and 78% and to feline coronavirus type II of 80 and 84%, respectively. Estimation of the phylogenetic relationship among coronavirus isolates indicated considerable divergence of the hyena variant from those in European, American and Japanese domestic cats and dogs. From long-term observations of several hundred known individuals, the only clinical sign in hyenas consistent with those described for coronavirus infections in dogs and cats was diarrhea. There was no evidence that coronavirus infection in hyenas caused clinical signs similar to feline infectious peritonitis in domestic cats or was a direct cause of mortality in hyenas. To our knowledge, this is the first report of coronavirus infection in Hyaenidae. PMID- 15288922 TI - The effect of a killed porcine reproductive and respiratory syndrome virus (PRRSV) vaccine treatment on virus shedding in previously PRRSV infected pigs. AB - Two experiments were conducted to investigate if virus shedding could be reduced following a killed porcine reproductive and respiratory syndrome virus (PRRSV) vaccination (KV) of PRRSV infected pigs. In experiment 1, PRRSV infected pigs were vaccinated with KV on days 14 and 28 following infection. Viremia and serum neutralizing (SN) antibody were compared to infected pigs with no KV. The second experiment was conducted in an identical manner. In addition to viremia and SN antibody, virus in oropharyngeal scrapings and interferon gamma (IFN-gamma) producing cells were monitored. Magnitude and duration of viremia were not different between KV vaccinated and non-vaccinated groups. No virus was detected in oropharyngeal scraping from any pig, nor was there a difference in the detection of viral RNA. In both experiments, however, increases in SN titer and number of IFN-gamma producing cells were observed. The SN titer was significantly higher in KV vaccinated groups than in non-vaccinated group on days 42 and 42-56 following infection in experiments 1 and 2, respectively. The number of IFN-gamma producing cells was slightly higher in KV vaccinated groups than in non vaccinated group on days 42 and 63. These observations suggest that KV had no effect on virus shedding. However, previously infected pigs responded immunologically to KV, as demonstrated by increases in SN antibody titers and IFN gamma producing cells. PMID- 15288923 TI - Characterisation of a type 2 bovine viral diarrhoea virus isolated from cattle in the UK. AB - Two genotypes of bovine viral diarrhoea virus (BVDV) are recognised. Type 2 was first recognised when virulent strains caused significant losses among cattle in North America. Subsequently, BVDV type 2 has been found in many other countries, but recent studies have shown that only type 1 BVDV is circulating in the UK herds (sheep and cattle) with type 1a predominating. During routine genotyping of UK BVDV isolates, a type 2 isolate was identified. Phylogenetic analysis of the 5'-untranslated region of the viral genome showed it to be a BVDV type 2a, most similar to a low virulent US strain of BVDV type 2. Antigenic typing with a panel of monoclonal antibodies verified this classification. This is the first confirmed isolation of BVDV type 2 found circulating in the UK. PMID- 15288924 TI - Comparison of methods for antimicrobial susceptibility testing and MIC values for pleuromutilin drugs for Brachyspira hyodysenteriae isolated in Germany. AB - In Germany treatment of swine dysentery is hampered by Brachyspira hyodysenteriae strains showing elevated MIC values to the few antibiotics licensed. Therefore, susceptibility testing of clinical isolates is an important service to the swine practitioner. This study compares the established agar dilution procedure for antimicrobial susceptibility testing of this fastidious anaerobe to the broth microdilution test newly developed [Anim. Health Res. 2 (2001) 59; Vet. Microbiol. 84 (2002) 123; J. Clin. Microbiol. 41 (2003) 2596]. A total of 221 isolates were examined twice with either test procedure using tiamulin and valnemulin as antibiotics. Both methods gave reproducible results, and the MIC values for the reference strains B. hyodysenteriae B204 and Staphylococcus aureus ATCC 29213 corresponded to previously published data. However, the results for individual strains differed significantly for both tests (P < 0.001) with MIC values being on average one dilution step lower in the broth dilution method. The 221 strains used for comparing test procedures were isolated between 1989 and 2001. An additional 102 strains isolated in 2002 were tested only with the broth dilution procedure. A significant rise in the average MIC value for both pleuromutilins could be demonstrated when comparing earlier isolates to those from 2000 to 2001 (P < 0.05), while in 2002 the average MIC significantly decreased when compared to the value in 2000 (P < 0.05). However, strains with MIC values for tiamulin as high as 8 microg/ml (broth dilution) could still be isolated. PMID- 15288925 TI - Antimicrobial susceptibility of mastitis pathogens from first lactation and older cows. AB - Increasing antimicrobial resistance has become a serious concern worldwide and antimicrobial use in animal agriculture is currently under scrutiny. Mastitis is the most common reason for antibiotic use in dairy herds and thus, antimicrobial resistance of mastitis pathogens has received recent attention. The purpose of this study was to describe and compare antimicrobial susceptibility of mastitis pathogens isolated at calving from first lactation and older cows. A total of 202 bacteria were isolated from intramammary infections (IMIs) within 3 days after calving over a 16-month study period in the Krauss Dairy Research Herd at the Ohio Agricultural Research and Development Center (OARDC) in Wooster, OH. Of these IMIs, 78% were caused by coagulase-negative staphylococci (CNS). Forty-four percent of them were resistant to at least one antibiotic. Most resistance was observed against penicillin, 39% of the isolates from older cows and 26% from first lactation cows being resistant to penicillin (P > 0.05). Also MIC90 for penicillin was higher among isolates from older cows. On the other hand, resistance to tetracycline was more common and MIC90 higher among isolates from first lactation cows than from older cows. Differences in the proportions of resistant isolates between first lactation and older cows were not statistically significant, though. The resistance patterns of the CNS isolated during the study are concordant with antimicrobial usage in the study herd. This is in agreement with the generally accepted notion that selection pressure from the use of antibiotics is a main factor in development of antibiotic resistance. PMID- 15288926 TI - Characterisation of attaching-effacing Escherichia coli isolated from animals at slaughter in England and Wales. AB - Escherichia coli isolates were recovered from faecal samples taken from cattle, sheep and pigs at slaughter in England and Wales. Isolates (n = 1227) selected at random from this collection were each hybridised in colony dot-blot experiments with an eae gene probe that presumptively identified attaching-effacing E. coli (AEEC). Of the 99 (8.1%) eae positive isolates 72 were of ovine origin, 24 were of bovine origin and three of porcine origin. None were typed as O157:H7 whereas 78 were assigned to 23 serogroups and 21 were untypable. The most frequently isolated eae positive serogroups were O156 (10), O26 (8), O103 (8), O108 (7) O56 (6) and O168 (6) of which serogroups O103 and O156 only were recovered from all three animal species. In tissue culture adherence assays, 36 representatives of eae positive isolates of all serogroups and host of origin tested induced intimate attachment with varying degrees of actin accumulation and pedestal formation in the HEp-2 cells. The identity of the eae type for these 36 was determined by specific PCR and the most prevalent intimin types were eaebeta (15), eaegamma (12) and eae (4). Isolates were examined by PCR for the presence of other virulence determinants and five possessed stx1 but none possessed stx2. One O115 eae isolate possessed cnf1 and 2, hlyA, etpD and katP genes which is a novel combination of virulence determinants. PMID- 15288927 TI - Development of two real-time PCR assays for the detection of Mycoplasma hyopneumoniae in clinical samples. AB - In order to improve the diagnosis of enzootic pneumonia (EP) in pigs two real time polymerase chain reaction (rtPCR) assays for the detection of Mycoplasma hyopneumoniae in bronchial swabs from lung necropsies were established and validated in parallel. As a gold standard, the current "mosaic diagnosis" was taken, including epidemiological tracing, clinical signs, macro- and histopathological lesions of the lungs and immunofluorescence. One rtPCR is targeting a repeated DNA element of the M. hyopneumoniae genome (REP assay), the other a putative ABC transporter gene (ABC assay). Both assays were shown to be specific for M. hyopneumoniae and did not cross react with other bacteria and mollicutes from pig. With material from pigs of defined EP-negative farms the two assays showed to be 100% specific. When testing lungs from pig farms with EP, the REP assay detected 50% and the ABC assay 90% of the farms as positive. Both tests together detected all positive farms. Within a positive herd the two assays tested similarly with on average over 90% of the lung samples analysed from a single farm showing positive scores. A series of samples with suspicion of EP and samples from pigs with diseases other than respiratory taken from current routine diagnostic was assayed. None of the assays showed false positive results. The sensitivities in this sample group were 50% for the REP and 70% for the ABC assays and for both assays together 85%. The two assays run in parallel are therefore a valuable tool for the improvement of the current diagnosis of EP. PMID- 15288928 TI - Lack of influence of the anaerobic [NiFe] hydrogenase and L-1,2 propanediol oxidoreductase on the outcome of Actinobacillus pleuropneumoniae serotype 7 infection. AB - The genes for the large subunit of [NiFe] hydrogenase 2, (hybB) and for L-1,2 propanediol oxidoreductase (fucO), were identified in an Actinobacillus (A.) pleuropneumoniae serotype 7 strain. Based on the hypothesis that adaptation to anaerobic conditions in damaged lung tissue may play a role in A. pleuropneumoniae persistence in host tissues, deletion mutants with a deletion in the hybB or the fucO gene were constructed and examined in an aerosol infection model. Deletion of the hybB or fucO genes appeared to have no significant effect on A. pleuropneumoniae virulence. PMID- 15288929 TI - The O-antigen of Salmonella enterica serotype Enteritidis PT4: a significant factor in gastrointestinal colonisation of young but not newly hatched chicks. AB - The lipopolysaccharide of Salmonella and other Gram negative pathogenic species has been implicated as a major virulence determinant and in this study we report the role of LPS of S. Enteritidis in the colonisation and persistent gastrointestinal infection of young poultry. The gene encoding the unique O antigen ligase, waaL, was mutated by insertional inactivation in a well characterised S. Enteritidis strain, S1400/94. The waaL mutant, designated PCP, produced rough colonies on agar medium, did not agglutinate O9 antiserum, did not produce an LPS ladder on silver stained gels and was serum sensitive. PCP and a nalidixic acid marked derivative of S1400/94 (S1400/94 Nalr) were used to orally challenge young chicks, separately and together in competitive index experiments. At post-mortem examination of 1-day-old chicks challenged S1400/94 Nalr and PCP separately there were no significant differences in the numbers of S1400/94 Nalr and PCP bacteria in tissues sampled on days 1, 2, and 5. By day 42 after challenge S1400/94 Nalr bacteria were recovered in significantly higher numbers than PCP from the caecal contents (P < 0.001). In competitive index studies in the 1-day-old chick PCP colonised, invaded and persisted in lower numbers than S1400/94 Nalr. In 4-week-old chicks challenged separately, PCP bacteria were recovered from all tissues examined in significantly lower numbers than S1400/94 Nalr. In competitive index experiments in 4-week-old chicks, PCP was not detected at any site and at any time point. Therefore, the O-antigen of S. Enteritidis plays an important role in poultry infections although this role is less important in the newly hatched chick. PMID- 15288930 TI - Cloning and purification of the Streptococcus suis serotype 2 glyceraldehyde-3 phosphate dehydrogenase and its involvement as an adhesin. AB - Streptococcus suis serotype 2 is a swine pathogen responsible for diverse diseases and may be present in the tonsils of pigs which show no sign of illness. Because adhesion to host cells may be important in the carrier state, this study was undertaken to characterize a 39 kDa surface protein identified as a glyceraldehyde-3-phosphate dehydrogenase (GAPDH), possibly implicated in the adhesion of the bacteria. The gene encoding for the GAPDH of S. suis was cloned and sequenced. The DNA sequence contained an open reading frame encoding for a 336 amino acid polypeptide exhibiting 95% sequence identity with the GAPDH from Streptococcus pyogenes and from other streptococci. Using the Qiaexpress expression plasmids, the gapdh gene was inducibly overexpressed in E. coli to produce GAPDH with a hexahistidyl N-terminus to permit its purification. The (His)6GAPDH protein was found to possess functional GAPDH enzymatic activity after the purification. An adherence assay with S. suis and porcine tracheal rings pre-incubated with (His)6GAPDH and non-incubated rings was showed a significant reduction in the adhesion of S. suis in the (His)6GAPDH pre-incubated rings compared to the non-incubated rings. The GAPDH protein of S. suis seems to be involved in the first steps of the bacterial adhesion to host cells. PMID- 15288931 TI - Virulence-associated genes in Escherichia coli isolates from poultry with colibacillosis: correction. AB - Several virulence genes of avian Escherichia coli were detected in 200 colibacillosis isolates from our region by PCR. However, the genes sfaDE and facA were not detected in that study. In this work we correct those data, showing by colony hybridization that sfaDE and facA are present in 40% and 30% of those isolates, respectively. PMID- 15288932 TI - Protection of pigs from swine dysentery by vaccination with recombinant BmpB, a 29.7 kDa outer-membrane lipoprotein of Brachyspira hyodysenteriae. AB - Swine dysentery (SD) is an important endemic infection in many piggeries, and control can be problematic. In this study the efficacy of BmpB, a 29.7 kDa outer membrane lipoprotein of Brachyspira hyodysenteriae, was evaluated as an SD vaccine. Non-lipidated BmpB was expressed in Escherichia coli as a histidine tagged protein (His6-BmpB), or as an 8 kDa carboxy-terminal portion fused to maltose-binding protein (MBP-BmpB-F604). The purified proteins were emulsified with oil-based adjuvants for intramuscular (im) administrations. In experiment 1, 20 weaner pigs were vaccinated im with 1 mg of His6-BmpB. After 3 weeks, 10 received 1 mg of the protein orally (im/oral), and 10 received 1 mg im (im/im). Ten acted as unvaccinated controls. In experiment 2, 12 pigs were vaccinated im with 1 mg of His6-BmpB, and 12 with 1 mg of MBP-BmpB-F604. Three weeks later, each was given 1 mg of the same protein orally. Twelve pigs acted as unvaccinated controls. All pigs were challenged orally with B. hyodysenteriae 2 weeks after their second vaccination. In both experiments, all pigs vaccinated with His6-BmpB developed serum antibodies to BmpB, and oral administration provided boosting of im-induced serum antibody titres. In experiment 1, seven non-vaccinated control pigs developed dysentery and severe colitis. Three pigs vaccinated im/oral developed diarrhoea; two had severe colitis and one had mild lesions. Four pigs vaccinated im/im developed diarrhoea; one had severe colitis and the others had mild lesions. In experiment 2, six control pigs developed SD with severe colitis. Two His6-BmpB vaccinated pigs developed SD with mild colitis. Nine pigs vaccinated with MBP-BmpB-F604 developed SD and severe colitis. Overall, 50-70% of controls and 17-40% of His6-BmpB vaccinated pigs developed disease. Vaccination with MBP-BmpB-F604 did not induce serum titres against BmpB, nor confer protection. The incidence of disease for the three His6-BmpB vaccinated groups was significantly less (P = 0.047) than for the control groups, with a approximately 50% reduction. BmpB appears to have potential as an SD vaccine component. PMID- 15288933 TI - Role of catalase in the virulence of Brucella melitensis in pregnant goats. AB - An isogenic katE mutant derived from virulent Brucella melitensis 16M displays hypersensitivity to hydrogen peroxide in disk sensitivity assays but retains the capacity to colonize pregnant goats and induce abortion. These experimental findings indicate that although the sole periplasmic catalase of Brucella melitensis functions as an antioxidant, this enzyme does not play a critical role in virulence in the natural host. PMID- 15288934 TI - Differentiation of Pasteurella multocida isolates from cases of atrophic rhinitis in pigs from Zimbabwe by RAPD and ribotyping. AB - Atrophic rhinitis in pigs is rarely reported in Southern Africa. To determine the relationship between Pasteurella multocida clones from clinical cases of atrophic rhinitis, twenty-one strains were characterised by selected phenotypic and genotypic methods. Biochemical analysis classified 18 strains as P. multocida subspecies multocida, whilst the remainder were grouped into separate unassigned biotypes. Capsular groups A (16/21) and D (l/21) were found among the isolates by PCR. Four ribotype patterns were obtained following HpaII ribotyping, whilst random amplification of polymorphic DNA (RAPD) revealed three main clusters. However, subclusters were also noted for each RAPD cluster. Our results indicate that RAPD offers a better discrimination of strains than ribotyping and that none of the phenotypic characters were directly related to the genotypic clusters. PMID- 15288936 TI - Single molecule force spectroscopy on ligand-DNA complexes: from molecular binding mechanisms to biosensor applications. AB - Recent developments in single molecule force spectroscopy (SMFS) allow direct observation and measurements of forces that hold protein-DNA complexes together. Furthermore, the mechanics of double-stranded (ds) DNA molecules in the presence of small binding ligands can be detected. The results elucidate molecular binding mechanisms and open the way for ultra sensitive and powerful biosensor applications. PMID- 15288937 TI - Theoretical analysis of dynamic force spectroscopy experiments on ligand-receptor complexes. AB - The forced rupture of single chemical bonds in biomolecular compounds (e.g. ligand-receptor systems) as observed in dynamic force spectroscopy experiments is addressed. Under the assumption that the probability of bond rupture depends only on the instantaneously acting force, a data collapse onto a single master curve is predicted. For rupture data obtained experimentally by dynamic AFM force spectroscopy of a ligand-receptor bond between a DNA and a regulatory protein we do not find such a collapse. We conclude that the above mentioned, generally accepted assumption is not satisfied and we discuss possible explanations. PMID- 15288938 TI - Detection and manipulation of biomolecules by magnetic carriers. AB - The detection and manipulation of single molecules on a common platform would be of great interest for basic research of biological or chemical systems. A promising approach is the application of magnetic carriers. The principles are demonstrated in this contribution. It is shown that paramagnetic beads can be detected by highly sensitive magnetoresistive sensors yielding a purely electronic signal. Different configurations are discussed. The capability of the sensors to detect even single markers is demonstrated by a model experiment. In addition, the paramagnetic beads can be used as carriers for biomolecules. They can be manipulated on-chip via currents running through specially designed line patterns. Thus, magnetic markers in combination with magnetoresistive sensors are a promising choice for future integrated lab-on-a-chip systems. PMID- 15288939 TI - Analysing a magnetic molecule detection system--computer simulation. AB - The detection of single molecules, e.g. in biology is possible by marking the interesting molecules with magnetic beads and detect the influence of the beads on giant magnetoresistance (GMR)/tunnel magnetoresistance (TMR)/spin valve (SV) sensors. The development of suitable multilayers has been studied experimentally as well as theoretically in order to optimize the sensor parameters. A finite difference (FD) method including the usually used contributions to the total energy [exchange, antiferromagnetically (af) coupling, anisotropy and magnetostatic] is used for the simulation with additional contributions to the local field according to the stray fields of the beads. In this work, we will show the results of micromagnetic calculations of the magnetization behavior of GMR/TMR sensors considering also the interaction between the domains in the magnetic layers of the sensor and the bead area. We can present first calculations where the bead particles (signal source) and the magnetic layers (sensor device) are considered as a whole magnetic ensemble. PMID- 15288940 TI - New magnetic nanoparticles for biotechnology. AB - Paramagnetic carriers, which are linked to antibodies enable highly specific biological cell separations. With the colloidal synthesis of superparamagnetic Co and FeCo nanocrystals with superior magnetic moments the question about their potential to replace magnetite as the magnetically responsive component of magnetic beads is addressed. Starting from a magnetic analysis of the corresponding magnetophoretic mobility of Co and FeCo based alloys their synthesis and resulting microstructural and magnetic properties as function of the underlying particle size distribution are discussed in detail. The stability of the oleic acid ligand of Co nanocrystals has been investigated. The oxidation kinetics were quantified using magnetic measurements. As a result, this ligand system provides sufficient protection against oxidation. Furthermore, the kinetics of the synthesis of Fe(50)Co(50) nanoparticles has been monitored employing Fourier transform infra red (FT-IR) spectroscopy and is modeled using a consecutive decomposition and growth model. This model predicts the experimentally realized FeCo nanoparticle composition as a function of the particle size fairly well. High-resolution transmission electron microscopy (HRTEM) was performed to uncover the resulting microstructure and composition on a nanometer scale. PMID- 15288941 TI - Towards single molecule analysis in PDMS microdevices: from the detection of ultra low dye concentrations to single DNA molecule studies. AB - In this paper, we report on the performance of electrophoretical separation and laser-induced fluorescence (LIF) detection of dyes and fluorescently labeled biomolecules in poly(dimethylsiloxane) (PDMS) microdevices. The dyes fluorescein and fluorescein isothiocyanate (FITC) have been separated effectively in nM concentrations. Fluorescein injections gave linear concentration response in the range from 4 to 100 pM. As ultimate detection sensitivity, 100 fM injected fluorescein was obtained. Further, 100 fM injected fluorescein could be detected. This is to our knowledge the smallest electrokinetically injected dye concentration detected on a microchip. Injection studies of fluorescently labeled avidin revealed a theoretical detection limit of 25 nM for laser-induced fluorescence detection in good agreement with separations in glass chips. Furthermore, the injection of several and even one single DNA molecule using a PDMS cross injector has been demonstrated as well as free solution separation of lambda- and T2-DNA (60 pM each) in periodically structured channels. PMID- 15288942 TI - An improved method for the solution cyclization of peptides under pseudo-high dilution conditions. AB - Depending on the ring size, the cyclization of peptides often is accompanied by dimerization or cyclodimerization. Hence, these macrocyclizations have to be performed under high dilution conditions. Efficient cyclization of peptides in solution with a minimum amount of solvent succeeds, when a dual syringe pump is used to simultaneously add the linear peptide precursor and a coupling reagent from two separate syringes. PMID- 15288943 TI - Mechanisms of DNA separation in entropic trap arrays: a Brownian dynamics simulation. AB - Using Brownian dynamics simulations, we study the migration of long charged chains in an electrophoretic microchannel device consisting of an array of microscopic entropic traps with alternating deep regions and narrow constrictions. Such a device has been designed and fabricated recently by Han and Craighead [Science 288 (2000) 1026] for the separation of DNA molecules. Our simulation reproduces the experimental observation that the mobility increases with the length of the DNA. A detailed data analysis allows to identify the reasons for this behavior. Two distinct mechanisms contribute to slowing down shorter chains. One has been described earlier by Han and Craighead [Science 288 (2000) 1026]: the chains are delayed at the entrance of the constriction and escape with a rate that increases with chain length. The other, actually dominating mechanism is here reported for the first time: some chains diffuse out of their main path into the corners of the box, where they remain trapped for a long time. The probability that this happens increases with the diffusion constant, i.e., the inverse chain length. PMID- 15288944 TI - Absence of intrinsic electric conductivity in single dsDNA molecules. AB - The intrinsic dc conductivity of long, individual lambda phage dsDNA molecules has been investigated by ultrasensitive low current-voltage-spectroscopy (IV) under ambient conditions and controlled low humidity inert gas atmosphere on microfabricated metal-insulator-metal gap structures. We found a strong dependence of the measured conductivity on the apparent humidity, which we attribute to capillary condensation of water to the immobilized DNA molecules, giving rise to additional ionic currents. Additional IV-spectroscopy experiments under controlled argon atmosphere always revealed a significant drop in electrical conductivity to 4 x 10(-15)AV(-1)microm(-1), indicating almost no considerable contribution of electrical long range charge transport. PMID- 15288945 TI - Controlled three-dimensional immobilization of biomolecules on chemically patterned surfaces. AB - We used electron-beam lithography to fabricate chemical nanostructures, i.e. amino groups in aromatic self-assembled monolayers (SAMs) on gold surfaces. The amino groups are utilized as reactive species for mild covalent attachment of fluorescently labeled proteins. Since non-radiative energy transfer results in strong quenching of fluorescent dyes in the vicinity of the metal surfaces, different labeling strategies were investigated. Spacers of varying length were introduced between the gold surface and the fluorescently labeled proteins. First, streptavidin was directly coupled to the amino groups of the SAMs via a glutaraldehyde linker and fluorescently labeled biotin (X-Biotin) was added, resulting in a distance of approximately 2 nm between the dyes and the surface. Scanning confocal fluorescence images show that efficient energy transfer from the dye to the surface occurs, which is reflected in poor signal-to-background (S/B) ratios of approximately 1. Coupling of a second streptavidin layer increases the S/B-ratio only slightly to approximately 2. The S/B-ratio of the fluorescence signals could be further increased to approximately 4 by coupling of an additional fluorescently labeled antibody layer. Finally, we introduced tetraethylenepentamine as functional spacer molecule to diminish fluorescence quenching by the surface. We demonstrate that the use of this spacer in combination with multiple antibody layers enables the controlled fabrication of highly fluorescent three-dimensional nanostructures with S/B-ratios of >20. The presented technique might be used advantageously for the controlled three dimensional immobilization of single protein or DNA molecules and the well defined assembly of protein complexes. PMID- 15288947 TI - Structural organization of DMPC lipid layers on chemically micropatterned self assembled monolayers as biomimetic systems. AB - The growth structure of DMPC lipid layers on hydrophobic and hydrophilic alkylsilane-based self-assembled monolayers adsorbed on silicon has been investigated by means of X-ray reflectometry and atomic force microscopy. Hydrophilic modification of hydrophobically terminated ODS-SAMs has been achieved by dose-controlled irradiation with DUV light. While island formation of small DMPC bilayer islands is observed on hydrophobic SAM surfaces, closed layers of DMPC monolayers are formed on hydrophilic SAM surfaces. Furthermore, DMPC adsorption on chemically micropatterned substrates with alternating hydrophobic/hydrophilic surface properties has been studied by imaging ellipsometry and photoemission microscopy. Indication for at least partial bridging of hydrophobic areas by an adsorbed DMPC monolayer has been found. PMID- 15288946 TI - Solid phase synthesis of an amphiphilic peptide modified for immobilisation at the C-terminus. AB - The solid phase peptide synthesis (SPPS) of the amphiphilic peptide Ac-(Leu-Ala Arg-Leu)(3)-linker, which is modified at the C-terminus with 1,8-diamino-3,6 dioxaoctane as linker moiety, has been investigated. Two different approaches that allow for the synthesis of C-terminally modified, side-chain protected peptides were examined. The solid phase peptide synthesis using aliphatic safety catch resin followed by activation and aminolysis with the mono-Boc protected linker was compared with the synthesis on 1,8-diamino-3,6-dioxaoctane loaded 2 chlorotrityl resin. PMID- 15288948 TI - Synthesis of photoswitchable amino acids based on azobenzene chromophores: building blocks with potential for photoresponsive biomaterials. AB - Three novel derivatives of azobenzene substituted amino acids have been synthesized. The compounds may serve as photoswitchable building blocks in the synthesis of bicyclic peptides or peptide strands interconnected with a photoisomerizable group. PMID- 15288949 TI - Large-scale homogeneous molecular templates for femtosecond time-resolved studies of the guest-host interaction. AB - Self-assembled monolayer films based on iodobenzoyloxy-functionalized resorc[4]arenes were prepared on gold substrates to serve as model systems for future time-resolved studies of molecular recognition, a mechanism of outstanding importance in bioorganic systems. The film properties were tested using X-ray photoelectron spectroscopy (XPS), atomic force microscopy (AFM) and imaging ellipsometry. An apparatus for time-resolved electron spectroscopy utilizing femtosecond soft X-ray pulses is capable of detecting iodine core-level photolines and the photoinduced dissociation after ultraviolet illumination. The developed technique holds promise for tracking the temporal evolution of chemical shifts of atomic markers as local probes for the dynamics of the guest-host interaction. PMID- 15288950 TI - The use of surface plasmon resonance (SPR) and fluorescence resonance energy transfer (FRET) to monitor the interaction of the plant G-proteins Ms-Rac1 and Ms Rac4 with GTP. AB - Using an RT-PCR approach a cDNA clone, designated Ms-Rac4 and putatively coding for a small GTPase was isolated from Medicago sativa. Ms-Rac4 and the earlier described Ms-Rac1 [Mol. Gen. Genet. 263 (2000) 761] belong to the class of GTP binding Rho of plants (Rop) proteins. At the amino acid level they display all conserved regions that are common to GTP-binding proteins. Phylogenetically both are located in the group Ia, but within this group they are well-separated. Computed structure models of both proteins revealed a high degree of structural conservation. Particularly the switch I and switch II region are 100% conserved between Ms-Rac1 and Ms-Rac4 and highly conserved as compared to other Rac-like G proteins. Both GTPases differ in structure within the fourth loop and the fourth helix. GTP-binding properties of the heterologously expressed Ms-Rac1 and Ms-Rac4 was shown by fluorescence resonance energy transfer (FRET) using mantGTP and by surface plasmon resonance (SPR). By this method the specificity of the G protein/GTP interaction was shown and the inhibitory effect of GTP, EDTA and Mg(2+) on the Ms-Rac1 and Ms-Rac4 binding to immobilized GTP was characterized. Ms-Rac1 and Ms-Rac4 exhibited the same affinity to GTP and are similarly affected by GTP, EDTA and Mg(2+). Thus, the predicted structural differences do not result in different GTP-binding properties of Ms-Rac1 and Ms-Rac4. PMID- 15288951 TI - Colocalization and FRET-analysis of subunits c and a of the vacuolar H+-ATPase in living plant cells. AB - The proton-translocating plant vacuolar H(+)-ATPase (VHA) is of prime importance for acidification of intracellular compartments and is essential for processes such as secondary activated transport, maintenance of ion homeostasis, and adaptation to environmental stress. Twelve genes have been identified that encode subunits of the functional V-ATPase complex. In this study, subunits c and a of the V-ATPase from the plant Mesembryanthemum crystallinum were fused to cyan fluorescent protein (CFP) and yellow fluorescent protein (YFP), respectively, and were transiently coexpressed in protoplasts. Two-colour scanning confocal fluorescence microscopy demonstrates that the fusion proteins VHA-c-CFP and VHA-a YFP are colocalized at the tonoplast, the plasmamembrane, and at endoplasmic membrane structures indicating expression in cytoplasmic vesicles. Furthermore, fluorescence resonance energy transfer (FRET) was used to visualize the interaction of VHA-c and VHA-a in vivo on the nanometer length scale. Excitation of CFP as donor fluorophore caused increased emission of YFP-fluorescence in protoplasts due to FRET. Our results give strong evidence for physical interaction of subunits c and a in living plant cells. PMID- 15288952 TI - Classification of hyper-variable Corynebacterium glutamicum surface-layer proteins by sequence analyses and atomic force microscopy. AB - The structural S-layer proteins of 28 different Corynebacterium glutamicum isolates have been analyzed systematically. Treatment of whole C. glutamicum cells with detergents resulted in the isolation of S-layer proteins with different apparent molecular masses, ranging in size from 55 to 66 kDa. The S layer genes analyzed were characterized by coding regions ranging from 1,473 to 1,533 nucleotides coding for S-layer proteins with a size of 490-510 amino acids. Using PCR techniques, the corresponding S-layer genes of the 28 C. glutamicum isolates were all cloned and sequenced. The deduced amino acid sequences of the S layer proteins showed identities between 69 and 98% and could be grouped into five phylogenetic classes. Furthermore, sequence analyses indicated that the S layer proteins of the analyzed C. glutamicum isolates exhibit a mosaic structure of highly conserved and highly variable regions. Several conserved regions were assumed to play a key role in the formation of the C. glutamicum S-layers. Especially the N-terminal signal peptides and the C-terminal anchor sequences of the S-layer proteins showed a nearly perfect amino acid sequence conservation. Analyses by atomic force microscopy revealed a committed hexagonal structure. Morphological diversity of the C. glutamicum S-layers was observed in a class specific unit cell dimension (ranging from 15.2 to 17.4 nm), which correlates with the sequence similarity-based classification. It could be demonstrated that differences in the primary structure of the S-layer proteins were reflected by the S-layer morphology. PMID- 15288953 TI - Regulation of gene expression with pyrrole-imidazole polyamides. AB - The pyrrole-imidazole (Py-Im) polyamides represent the only available class of small molecules that can be designed to recognize virtually any predetermined DNA sequence. These molecules have affinities and specificities that equal or exceed natural eukaryotic transcriptional regulatory proteins. Studies with model gene systems, and a variety of eukaryotic and viral transcription factors, have shown that these molecules are potent inhibitors of protein-DNA interactions. Polyamides have been shown to regulate gene expression in simple in vitro systems using defined DNA templates and nuclear extracts as a source of the transcriptional machinery. Activation of gene expression has also been achieved in vitro with polyamide-activator peptide conjugates. Most importantly, polyamides are cell permeable and localize in the nucleus in various cultured cell lines and are capable of down regulating target genes in these cells. Polyamides have been shown to bind to their target sites in chromosomal DNA and both gain- and loss-of-function have been observed by targeting repeated DNA sequences in developing Drosophila embryos. PMID- 15288954 TI - Detection of hepatitis B virus core mutants by PCR-RFLP in chronically infected patients. AB - HBV chronic infection is an important health problem. The HBV core antigen carries several epitopes for T and B cell recognition and the immune response is crucial for determining the outcome of viral infection. Using PCR-RFLP several point mutations were detected in the HBV core ORF of HBV extracted from the serum of 140 chronically infected patients and 86 samples from another 37 patients followed-up in a longitudinal study. Mutations at position 2248 and 2147 (A3) and at 2038 (M2) were found most frequently. The wild type core genotype was found in about 50% of the samples. PCR-RFLP results were confirmed by direct sequencing of amplified products from HBV DNA present in chronically infected patients. The method is rapid and reliable and may be particularly useful for a rapid detection of viral mutants in a large number of patients. PMID- 15288955 TI - Simultaneous quantitation and genotyping of hepatitis B virus by real-time PCR and melting curve analysis. AB - Hepatitis B virus (HBV) genotype and HBV DNA levels have been implicated in clinical evaluation and prognosis of patients with chronic HBV infection. The aim of the present study was to develop a rapid and sensitive method for simultaneous HBV DNA quantitation and differentiation between HBV genotypes B and C in a single-step reaction by real-time PCR and melting curve analysis using SYBR Green I fluorescent dye. The genotypes obtained by this method were compared with those examined by PCR-RFLP and direct sequencing on 52 serum samples of patients with chronic HBV infection. Using the results obtained by direct sequencing and phylogenetic analysis as the reference, the accuracy of HBV genotyping by PCR RFLP and melting curve analysis was 90.38 and 92.31%, respectively. The geometric mean of HBV DNA levels was 3.42 x 10(6), 2.10 x 10(6), 1.19 x 10(5) and 3.10 x 10(4) copies/microl in asymptomatic carriers, patients with chronic hepatitis, cirrhosis and hepatocellular carcinoma, respectively. It is concluded that this method has the advantages of rapidity, reproducibility and accuracy, which would be feasible and attractive for large-scale analysis, particularly in regions where HBV genotypes B and C are prevalent. PMID- 15288956 TI - Rapid preparative purification of West Nile and Sindbis virus PCR products utilizing a microbore anion-exchange column. AB - Analysis and purification of specific PCR products from PCR reactions can be problematic due to several issues relating to amplification and low product yield. The use of HPLC as a preparative tool in PCR product analysis is common but has not replaced traditional electrophoretic techniques for purifying DNA to be used in subsequent experiments. Gel purification of PCR products can result in a net loss greater than 50% of the starting DNA amount. Thus, this method of recovery can become the limiting factor in the overall cloning protocol. This paper describes a simple and relatively inexpensive micro-preparative HPLC method to purify and analyze nM quantities of DNA. A microbore polyethyleneimine-based anion-exchange column fractionates PCR mixtures in less than 40 min with a recovery of the purified specific product as high as 80%, thus eliminating the need for gel purification. Using this method, nested PCR products from Sindbis virus differing by 18 bp in some cases and a 277 bp fragment from West Nile virus were resolved and quantified. This method differs from existing methodologies because separation is based on size and charge as well as the overall G + C content of the PCR product. PMID- 15288957 TI - Detection of economically important viruses in boar semen by quantitative RealTime PCR technology. AB - The objective of this study was to develop quantitative real-time polymerase chain reaction (ReTi-PCR) tests for the detection of five economically important viruses in swine semen namely, pseudorabies virus (PRV), classical swine fever virus (CSFV), foot-and-mouth disease virus (FMDV), swine vesicular disease virus (SVDV), and porcine reproductive and respiratory syndrome virus (PRRSV). Each ReTi-PCR test was validated for specificity, analytical sensitivity (detection limits), and experimental infection studies were performed to compare the conventional virus isolation methods with the newly developed ReTi-PCR tests. All five developed ReTi-PCR tests are very rapid compared to virus isolation, highly specific, and even more sensitive (lower detection limits) than conventional virus isolation methods for the detection of mentioned viruses in semen. In semen of experimentally infected boars, viruses were detected much earlier after infection and more frequently by ReTi-PCR tests than by virus isolations. The high throughput of these rapid ReTi-PCR tests makes it possible to screen large number of semen samples for the presence of viruses prior to insemination. This is a substantial advantage, in particular for boar semen the quality of which deteriorates quickly after storage. In general, the newly developed ReTi-PCR tests are valuable tools for the early, reliable and rapid detection of five economically important viruses, namely PRV, CSFV, FMDV, SVDV, and PRRSV in boar semen. These ReTi-PCR tests will improve the control of viral diseases transmitted via semen. PMID- 15288958 TI - Inactivation of HSV-2 by ascorbate-Cu(II) and its protecting evaluation in CF-1 mice against encephalitis. AB - Ascorbate is an important antioxidant. However, in the presence of transition metals such as Cu(II) or Fe(III), it also has pro-oxidant capabilities. The effect of ascorbate-Cu(II) in the in vitro infection of herpes simplex virus type 2 (HSV-2) and its protecting effect in a murine model was investigated. HSV-2 was treated with different concentrations of ascorbate in the presence of Cu(II). A group of CF-1 mice were treated with the inactivated virus and other treated with maintenance medium containing only ascorbate-Cu(II). Weeks later, mice were challenged intranasally with infectious viruses. HSV-2 was completely inactivated by 2mM ascorbate plus 1mM Cu(II). Ascorbate or Cu(II) alone did not inactivate the virus. Compared with the control group, 60% of the immunized animals did not show any sign of encephalitis and survived the herpes virus infection, while a 7% survival rate was observed in the control group (P = 0.056). We concluded that the in vitro treatment of HSV-2 with ascorbate-Cu(II) is not only able to inactivate the virus, but also suggested that the viral particles induced a protective response against herpes encephalitis. This inactivation may provide an alternative method to develop new agents therapeutics. PMID- 15288959 TI - Development of a simple restriction fragment length polymorphism assay for subtyping of coxsackie B viruses. AB - Coxsackie B viruses (genus, Enterovirus; family, Picornaviridae) can cause aseptic meningitis, encephalitis, pleurodynia, myocarditis and are implicated in the pathogenesis of dilated cardiomyopathy. The differentiation of the group B coxsackieviruses into their subtypes has potential clinical and epidemiological implications. In the present study, a simple restriction fragment length polymorphism (RFLP) assay was developed for typing of group B coxsackieviruses into subtypes 1-6. It is a two step process, first, virus isolation and identification by virus neutralization assay, using pools of polyclonal antisera, second, the reverse transcription polymerase chain reaction (RT-PCR) using a single primer pair selected from the conserved 5'-untranslated region (5'-UTR) of enterovirus genome followed by RFLP. A 440 bp product was amplified from the reference strains of each subtype of group B coxsackievirus and 29 clinical isolates (positive for group B coxsackieviruses by neutralization assay). The amplified products were subjected to restriction endonuclease digestion by enzyme BsaJI. The assay was able to distinguish all six serotypes of coxsackie B viruses. The results were comparable to serotyping and showed that due to the relatively conserved nature of 5'-UTR in enterovirus genome, this region can be used for subgeneric molecular identification of enteroviruses. PMID- 15288961 TI - Comparisons of physical separation methods of Kunjin virus-induced membranes. AB - The two sets of connected membranes induced in Kunjin virus-infected cells are characterized by the presence of NS3 helicase/protease in both, and by RNA dependent RNA polymerase (RdRp) activity plus the associated double-stranded RNA (dsRNA) template in vesicle packets (VP), or by the absence of both the VP specific markers in the convoluted membranes/paracrystalline arrays (CM/PC). Attempts were made to separate flavivirus-induced membranes by sedimentation or flotation analyses in density gradients of sucrose or iodixanol, respectively, after treatment of cell lysates by sonication, osmotic shock, or tryptic digestion. Only osmotic shock treatment provided suggestive evidence of separation. This was explored by flow cytometry analysis (FCA) of RdRp active membrane fractions from a sucrose gradient, using dual fluorescent labelling via antibodies to NS3 and dsRNA. FCA revealed the presence of a dual labelled membrane population indicative of VP, and in a faster sedimenting fraction a membrane population able to be labelled only in NS3, representative of CM/PC and associated (R)ER. It was postulated that osmotic shock ruptured the bounding membrane of the VP, releasing the enclosed small vesicles associated with the Kunjin virus replication complex characterized previously. Notably, the presence of the full spectrum of nonstructural proteins in some membrane fractions was not a reliable marker for RdRp activity. These experiments may provide the opportunity for isolation of relatively pure flavivirus replication complexes in their native membrane-associated state by fluorescence-activated cell sorting. PMID- 15288960 TI - Total HCV core antigen assay. A new marker of HCV viremia and its application during treatment of chronic hepatitis C. AB - The present study assesses the clinical usefulness of the hepatitis C core antigen assay for monitoring of patients being treated for chronic hepatitis C virus (HCV) infection. Eighty-six serum samples were selected at random from 16 patients and levels of HCV RNA and HCV core antigen were determined simultaneously and in parallel to compare both techniques. The data obtained were compared by Pearson's correlation and the coefficients calculated by Fisher transformation and by calculating the difference and standard error. A good linear correlation was observed between both techniques. Maximum correlation, with significant difference, was found between patients infected with the 1a genotype and other genotypes. In conclusion, the HCV core antigen assay is useful for the diagnosis of early infection; however, its use for determining the exact timing of viral elimination during treatment is clearly unsuitable. PMID- 15288962 TI - Retroviral vector targeting through insertion of epidermal growth factor into receptor binding deficient influenza A hemagglutinin results in fusion defective particles. AB - Targeting retroviral entry is a central theme in the development of vectors for gene therapy. The host range of a retrovirus is dependent upon the interaction of its envelope glycoprotein (Env) with a specific cell surface receptor protein, which allows viral entry. In contrast, the pH-dependent viruses enter cells through receptor-mediated endocytosis and the subsequent acidification produces conformational changes in the viral envelope protein(s) which lead to membrane fusion. We attempted to redirect retroviral vectors to epidermal growth factor (EGF) receptor expressing cells by using the pH-dependent influenza A virus hemagglutinin (HA). Wild type receptor binding was avoided either by point mutations or by deletion of the globular head structure of HA and also inserted EGF into HA. Replacement of the whole head domain was not tolerated. Two of the EGF-HA proteins bearing point mutations could be incorporated into retroviral particles, but unfortunately their fusion activity was lost. The data indicate that care must be taken when mutating multiple sites in HA, and that targeting HA requires further analysis of appropriate sites for the insertion of foreign sequences. PMID- 15288963 TI - Few modifications of the Cobas Amplicor HIV Monitor 1.5 test allow reliable quantitation of HIV-1 proviral load in peripheral blood mononuclear cells. AB - Recent studies have suggested that monitoring the amount of HIV provirus in peripheral blood mononuclear cells (PBMCs) may be a useful end point for HAART where, in combination with plasma viral load, it provides additional information as to the possibility of virus eradication. In the present study, a modified version of the Cobas Amplicor HIV-1 Monitor test (CAHIM), currently used to quantify plasma viremia, have been evaluated to also measure the amount of proviral DNA in PBMCs. The analytical and clinical performance of the modified CAHIM test was assessed by quantifying different amounts of a standard HIV-DNA preparation obtained from the 8E5 cell line and by analysing 165 patients and controls samples. In these experiments, the modified test, that showed a linear dynamic range from 1.7 to 4.7 log10 copies/10(6) cells (r = 0.99) with a maximum CV of 20%, proved able to detect and quantify HIV-DNA in all but one clinical samples, with concentrations varying from 1.3 to 3.8 log10 copies/10(6) cells. During anti-retroviral treatment, the assay revealed different proviral DNA time courses associated with viral load changes and inversely correlated with CD4+ cells count. As expected, HIV-DNA was always detectable even when plasma viremia fell below the CAHIM cut-off. The modified CAHIM test specificity was confirmed by testing 20 HIV-negative samples in triplicates. Taken together, the data showed that the modified CAHIM test can be used to monitor HIV proviral DNA changes during HAART and can help in investigating further the clinical use of this marker. PMID- 15288964 TI - Development of an efficient fluorescence-based microneutralization assay using recombinant human cytomegalovirus strains expressing green fluorescent protein. AB - MedImmune Vaccines has created four, live, attenuated human cytomegalovirus (HCMV) vaccine candidates, each derived from defined portions of the parental strains, Towne and Toledo. To determine each candidate's ability to induce HCMV specific immunity, a fluorescence-based microneutralization assay was developed using recombinants of Toledo and Towne which express enhanced green fluorescent protein (EGFP). Replication of the EGFP recombinants in cell culture was the same as the respective parental strains. Using the EGFP recombinants, this fluorescence-based microneutralization assay was compared with the traditional plaque reduction assay. Serum samples were analyzed by both the fluorescence microneutralization and plaque reduction assays and regression analysis showed a correlation of R2 > or = 0.90 between the two assays. As an alternative to measuring fluorescence, infected cells were examined microscopically and the number of green fluorescent cells was counted automatically. Regression lines between fluorescent cell counting and fluorescence in the well also showed a high correlation (R2 > or = 0.92). An excellent linear concordance in titers was observed between the two assays. Using the plaque reduction assay, serum samples were identified that preferentially neutralized the Toledo strain compared to the Towne strain. The same preferences were observed with the fluorescence-based microneutralization assay. This new assay is adaptable to rapid, automated collection of neutralization data and would therefore be suitable for the examination of large numbers of clinical serum samples. PMID- 15288965 TI - Development and use of a biotinylated 3ABC recombinant protein in a solid-phase competitive ELISA for the detection of antibodies against foot-and-mouth disease virus. AB - A biotinylated 3ABC recombinant protein was developed and used in a competitive ELISA (cELISA) to detect foot-and-mouth disease virus (FMDV) antibodies in cattle, sheep and pigs. In this report, we describe the cloning and expression of 3ABC protein in Escherichia coli cells as fusion protein with 6xHis and biotin. This cELISA uses streptavidin to capture bacterially expressed and in vivo biotinylated 3ABC antigen. The antigen capture strategy provides a simple and reliable method, which does not require purification of recombinant antigen before the serological assay. An hyperimmune guinea pig antiserum produced against purified 6xHis-3ABC was used as competitor in the test. The potential use of this cELISA for the identification of antibodies induced by FMD virus infection from those induced by vaccination is discussed. PMID- 15288966 TI - Specific detection of Nipah virus using real-time RT-PCR (TaqMan). AB - Nipah and Hendra viruses belong to the novel Henipavirus genus of the Paramyxoviridae family. Its zoonotic circulation in bats and recent emergence in Malaysia with fatal consequences for humans that were in close contact with infected pigs, has made the reinforcement of epidemiological and clinical surveillance systems a priority. In this study, TaqMan RT-PCR of the Nipah nucleoprotein has been developed so that Nipah virus RNA in field specimens or laboratory material can be characterized rapidly and specifically and quantitated. The linearity of the standard curve allowed quantification of 10(3) to 10(9) RNA transcripts. The sensitivity of the test was close to 1 pfu. The kinetics of Nipah virus production in Vero cells was monitored by the determination of infectious virus particles in the supernatant fluid and by quantitation of the viral RNA. Approximately, 1000 RNA molecules were detected per virion, suggesting the presence of many non-infectious particles, similar to other RNA viruses. TaqMan real-time RT-PCR failed to detect Hendra virus DNA. Importantly, the method was able to detect virus despite a similar ratio in viremic sera from hamsters infected with Nipah virus. This standardized technique is sensitive and reliable and allows rapid detection and quantitation of Nipah RNA in both field and experimental materials used for the surveillance and specific diagnosis of Nipah virus. PMID- 15288967 TI - Mouse antibody production test: can we do without it? AB - Introduction of microbiologically contaminated materials into mice can cause infections of the recipients and jeopardize experimental protocols. As such, the methods used to screen biological materials should be sensitive, reliable and suitable for routine diagnostic work. In this report, the sensitivity of the viral plaque assay, mouse antibody production test and polymerase chain reaction (PCR) for detection of MHV-A59 and MMVp, two of the most prevalent pathogenic viruses in experimental mouse facilities, was compared. Analysis of serial tenfold dilutions of virus stocks revealed that the sensitivity of the mouse antibody production test on day 28 (10(-10) dilution) was at least 10 times higher than that of the viral plaque assay (10(-9) dilution) and 10(4) times more than that of the RT-PCR (10(-6) dilution) for detection of MHV-A59. For detection of MMVp, the PCR (10(-10) dilution) proved to be 10(6) times more sensitive than the viral plaque assay (10(-4) dilution) and the mouse antibody production test on day 28 (10(-4) dilution) which were equally sensitive. Based on the present study, it was shown that the method for diagnosis of viruses in biological materials should be employed only after the sensitivity has been determined for the viruses of interest implying that the most sensitive method needs to be determined independently for each virus. PMID- 15288968 TI - Glucocorticoid-induced osteoporosis: considerations in ophthalmology. PMID- 15288969 TI - Prevalence of open-angle glaucoma and ocular hypertension in Latinos: the Los Angeles Latino Eye Study. AB - OBJECTIVE: To estimate age- and gender-specific prevalences of ocular hypertension and open-angle glaucoma (OAG) in adult Latinos. DESIGN: Population based, cross-sectional study. PARTICIPANTS: Six thousand three hundred fifty seven Latinos 40 years and older from 6 census tracts in Los Angeles, California. METHODS: The study cohort consisted of all self-identified Latinos of primarily Mexican ancestry 40 years and older residing in 6 census tracts in La Puente, California. All participants underwent a complete ophthalmologic examination, including measurement of intraocular pressure (IOP), visual field (VF) testing using an automated field analyzer, and simultaneous stereoscopic fundus photography of the optic disc. Ocular hypertension was defined as IOP of >21 mmHg and the absence of optic disc damage or abnormal VF test results. Open-angle glaucoma was defined as the presence of an open angle and various criteria that included a glaucomatous VF abnormality and/or evidence of glaucomatous optic disc damage in at least one eye. MAIN OUTCOME MEASURES: Prevalence of open-angle glaucoma and ocular hypertension. RESULTS: For the 6142 participants who underwent a complete ophthalmologic examination at the clinical center, the prevalence of OAG was 4.74% (95% confidence interval [CI], 4.22%-5.30%). The prevalence of ocular hypertension was 3.56% (95% CI, 3.12%-4.06%). The prevalences of OAG and ocular hypertension were higher in older Latinos than in younger Latinos (P<0.0001). No gender-related differences in prevalences of OAG and ocular hypertension were present. The mean IOP, mean deviation, and mean vertical cup-disc ratio in persons with OAG were 17 mmHg, -9.6 decibels, and 0.6, respectively. Seventy-five percent of Latinos with OAG and 75% of Latinos with ocular hypertension were previously undiagnosed. Further, 17% of Latinos with OAG and 23% of Latinos with ocular hypertension had received treatment for "glaucoma." CONCLUSION: Our data suggest that the prevalence of OAG is high among Latinos of Mexican ancestry. The higher prevalence of OAG in older Latinos emphasizes the public health importance of providing eye care services for the early diagnosis and management of this condition in Latinos. PMID- 15288970 TI - Prevalence of lens opacities in Latinos: the Los Angeles Latino Eye Study. AB - OBJECTIVE: To estimate the age- and gender-specific prevalence of posterior subcapsular (PSC), nuclear, cortical, and mixed lens opacities in a population based sample of Latinos 40 years and older. DESIGN: Population-based, cross sectional study. PARTICIPANTS: Six thousand three hundred fifty-seven Latinos 40 years and older from 6 census tracts in Los Angeles, California. METHODS: A population-based sample of Latinos underwent a complete eye examination, including assessment of presence and severity of lens opacification, using the slit lamp-based Lens Opacities Classification System II (LOCS II). All lens changes (including pseudophakia/aphakia); any nuclear, PSC, and cortical opacities; and nuclear-only, PSC-only, and cortical-only opacities were evaluated. Frequency distributions and chi-square test analyses were used to determine the age- and gender-specific prevalences for each opacity type. MAIN OUTCOME MEASURES: Prevalences of cortical, nuclear, and posterior subcapsular opacities. RESULTS: Of the 7789 eligible subjects, 6357 completed a clinical examination (82% participation rate). Of all participants with LOCS II grading, 20% had all lens changes, 7.6% had cortical-only opacities, 3.5% had nuclear-only opacities, 0.4% had PSC-only opacities, and 5.9% had mixed-type opacities. The prevalence of all types of lens opacities increased with age (P<0.0001). Of all participants with mixed opacities, 49% had monocular visual impairment and 20% had binocular impairment. Of all 6357 participants, 3.9% had undergone cataract extraction in at least one eye. CONCLUSION: Our data provide the first population based estimates of the prevalence and severity of lens opacities in Latinos. Cortical opacities were the most common type. The high rate of visual impairment from lens opacities suggests that programs that increase access to cataract surgery for older Latinos could help to reduce the burden of visual impairment in the United States. PMID- 15288971 TI - Prevalence of ocular diagnoses found on screening 1539 adults with intellectual disabilities. AB - OBJECTIVE: To study the prevalence of ocular disorders in adults with intellectual disabilities (IDs) in the Netherlands. DESIGN: Cross-sectional survey. SUBJECTS: A stratified random sample (for age more than 50 years and Down syndrome [DS]) of 1598 participants drawn from a base population of 9012 adult users of ID services with mild to profound intellectual disabilities in the Netherlands. METHODS: Participants underwent on-site visual screening on the basis of a protocol. Results were related to degree of ID, occurrence of DS, age, and a diagnosis of visual impairment or blindness. Referral to ophthalmologists followed when visual impairment was diagnosed. MAIN OUTCOME MEASURES: Diagnosis of ocular disorders and their prevalence. RESULTS: Refractive errors were most prevalent (60.6%), followed by strabismus (44.1%) and lens opacities (18.1%). Besides these, in participants diagnosed as visually impaired, cerebral visual impairment was the most common untreatable disorder (12.6%), followed by macular degeneration (5.4%). Compared with known figures from general populations, the prevalence of ocular diagnoses in adults with ID was significantly higher. The occurrence of refractive errors and strabismus was significantly related to DS (odds ratio [OR], 2.16; 95% confidence interval [CI], 1.56 to 3.00; and OR, 2.47; 95% CI, 1.93 to 3.17, respectively). Lens opacities had an independent relation with age more than 50 years (OR, 4.23; 95% CI, 3.04 to 5.88) and DS (OR, 8.27; 95% CI, 5.95 to 11.49). Keratoconus was independently related to DS (OR, 7.65; 95% CI, 3.91 to 14.96) and degree of ID (OR, 5.56; 95% CI, 2.79 to 11.06). Corneal opacities also were related to DS (OR, 2.70; 95% CI, 1.41 to 5.18) and degree of ID (OR, 5.53; 95% CI, 2.66 to 11.48). The risk of ocular hypertension was increased by age more than 50 years (OR, 2.54; 95% CI, 1.16 to 5.57) and severe or profound ID (OR, 4.86; 95% CI, 2.06 to 10.63); DS decreased the risk (OR, 0.21; 95% CI, 0.05 to 0.94). CONCLUSIONS: In 1539 adults with ID in the Netherlands, high prevalences of ocular disorders were found. Adults with ID in general have an increased risk of severe myopia, strabismus, and lens opacities; DS, older age, or severe ID further increase the risk of specific ocular disorders. PMID- 15288972 TI - Long-term outcomes in asians after acute primary angle closure. AB - PURPOSE: To determine the long-term outcome of Asian eyes with an acute attack of primary angle closure (APAC) and to identify risk factors at presentation associated with the development of glaucomatous optic nerve damage. DESIGN: Cross sectional observational case series. PARTICIPANTS: Ninety individuals who were initially seen with APAC 4 to 10 years previously at 2 Singapore hospitals. METHODS: All subjects underwent a complete eye examination, including visual acuity, visual field testing, dilated eye examination, and optic nerve head photography. The optic discs were judged clinically and photographically as to whether there was glaucomatous optic neuropathy present, and visual fields were assessed for corresponding visual field loss. All visual fields and optic nerve photographs underwent a second evaluation by an experienced, but masked, glaucoma specialist, who assessed whether the changes were compatible with glaucoma. MAIN OUTCOME MEASURES: The main outcome measures were blindness (defined as best corrected visual acuity worse than 6/60 and/or central visual field of less than 20 degrees in the attack eye) and glaucomatous optic neuropathy (GON). RESULTS: A total of 90 of 170 eligible subjects (65.2%) were examined. All subjects were Asian and were predominantly Chinese (78 subjects [86.7%]). There were 61 females (67.8%), and the age of the subjects was 62.0+/-9.0 years (mean +/- standard deviation) at the time of APAC, with a mean duration of 6.3+/-1.5 years from the time of the APAC episode to the study examination. Sixteen (17.8%) subjects were blind in the attack eye; half of the cases of blindness were caused by glaucoma. Forty-three subjects (47.8%) had GON, with 13 eyes (15.5%) having markedly cupped optic discs (cup-to-disc ratio >0.9). Thirty-eight eyes (58%) had best-corrected vision worse than 6/9, with cataract responsible for close to half the cases of poor vision. There were no identifiable risk factors related to the APAC episode that were significantly associated with the presence of GON. CONCLUSIONS: Several years after being seen with APAC, 17.8% of subjects examined were blind in the attack eye, and almost half had glaucomatous optic nerve damage. Vision was also reduced in a large number of individuals, largely from unoperated cataract. Subjects with APAC would benefit from regular follow-up to monitor for visual field decline and glaucoma development. PMID- 15288973 TI - Acute primary angle closure: configuration of the drainage angle in the first year after laser peripheral iridotomy. AB - PURPOSE: To evaluate the changes in the configuration of the drainage angle in the first year after acute primary angle closure (APAC). DESIGN: Prospective observational case series. PARTICIPANTS: Forty-four Asian subjects with APAC. METHODS: Acute primary angle closure cases were treated with medical therapy followed by laser peripheral iridotomy (LPI). Static and dynamic gonioscopies were performed in APAC-affected and fellow eyes before LPI (baseline) and then at 2 weeks, 4 months, and 12 months after presentation. The angles were graded in each quadrant according to the Shaffer scheme, and the number of clock hours of peripheral anterior synechiae (PAS) was recorded. Patients who underwent intraocular surgery at any point during follow-up were excluded from the study. Intraocular pressure (IOP) and medical treatment were documented at each visit, and gonioscopic changes were correlated with the development of elevation in IOP requiring medical treatment. MAIN OUTCOME MEASURES: Average Shaffer grade and the number of clock hours of PAS. RESULTS: The majority of subjects were Chinese (84%) and female (64%), and the mean age was 60.2+/-10.7 years. At presentation, 73% of both affected and fellow eyes had very narrow angles (average Shaffer grade < or = 1), with affected eyes having more extensive PAS (P<0.001), a third of whom had > or =8 clock hours of PAS. In APAC eyes, there was a significant increase in angle width from baseline to 2 weeks after LPI (P = 0.045), but no change in angle width subsequently. Fellow eyes showed a widening of the angle between baseline and week 2 (P = 0.01) and from week 2 to month 4 (P = 0.001). There was no significant change in PAS in either affected or fellow eyes over the 12 months of follow-up. Of the 44 subjects, 19 (41.3%) subsequently developed IOP elevation during follow-up that required treatment. However, there was no difference in angle width or amount of PAS between eyes with and without a subsequent rise in IOP, and the angle configuration did not change significantly in either group over 1 year. CONCLUSION: In Asian eyes with APAC, the angle widened in the first 2 weeks after LPI, but did not change thereafter over 1 year, and the amount of PAS remained stable throughout. The results indicate the effectiveness of LPI in preventing progressive closure of the angle in the first year after APAC. PMID- 15288974 TI - Changes in retinal nerve fiber layer thickness after acute primary angle closure. AB - PURPOSE: To evaluate the changes in retinal nerve fiber layer (RNFL) thickness in the first 16 weeks after acute primary angle closure (APAC) using scanning laser polarimetry (SLP). DESIGN: Prospective, observational case series. PARTICIPANTS: Thirty-seven Asian subjects with APAC. METHODS: For all cases, APAC had resolved after treatment, and the study was conducted during the follow-up period after the acute episode. Using the GDx Nerve Fiber Analyzer (Laser Diagnostic Technologies, San Diego, CA), the RNFL was assessed in both eyes 2 weeks after APAC, and again after 16 weeks. The SLP parameters were compared between week 2 and week 16 within affected and fellow eyes. A multiple logistic regression analysis was carried out to analyze factors likely to influence RNFL outcome. MAIN OUTCOME MEASURES: Superior and inferior average RNFL thickness. RESULTS: The mean age of study subjects was 60.1+/-10.3 years (range, 46-91 years), and most subjects were female (68%) and Chinese (86%). In APAC eyes, the superior average RNFL thickness decreased from 63.8+/-13.6 microm to 61.4+/-11.2 microm (P = 0.04) and the inferior average RNFL thickness decreased from 69.5+/-14.4 microm to 66.3+/-12.6 microm (P = 0.005). There was also a decrease in inferior ratio (P = 0.008) and ellipse modulation (P = 0.02). In the fellow eyes, there was no difference found between week 2 and week 16 for any of the SLP parameters studied. Logistic regression analysis showed no significant association between developing a 10% reduction in either superior or inferior RNFL thickness with age, gender, history of ischemic risk factors, duration of symptoms during APAC, the level of presenting intraocular pressure (IOP), or the development of a rise in IOP between study visits. CONCLUSIONS: After an episode of APAC, superior and inferior average RNFL thickness was found to decrease significantly from week 2 to week 16. PMID- 15288975 TI - Low incidence of iris pigmentation and eyelash changes in 2 randomized clinical trials with unoprostone isopropyl 0.15%. AB - OBJECTIVE: To assess whether iris color and eyelash changes occur with the use of unoprostone for 2 years. DESIGN: The 2 clinical trials described herein were prospective, randomized, double-masked, active-controlled, parallel group, multicenter studies. PARTICIPANTS: A total of 1131 patients with primary open angle glaucoma or ocular hypertension participated in 2 clinical trials and received either unoprostone isopropyl 0.15% (659), timolol maleate 0.5% (331), or betaxolol hydrochloride 0.5% (141), 1 drop per eye twice daily for up to 24 months. METHODS: Color photographs (1:1 magnification) were taken of the iris and eyelid of each patient at baseline and at regular intervals thereafter through month 24 using a standardized camera system. Photography included 7 views of each eye plus a calibration photograph and a patient identification photograph, for a total of 16 photographs per patient per visit. Two independent (masked) readers subjectively compared baseline iris colors to subsequent visits. Side view photographs of the upper and lower eyelashes were used for the eyelash length analysis, with each having sufficient depth of field and a sufficient number of eyelashes in focus. Similarly, frontal eyelash views were used for the eyelash density analysis. MAIN OUTCOME MEASURES: Changes from baseline in iris color and eyelash length and density within and between treatment groups. RESULTS: Seven cases of iris color change (1.06%) were confirmed in patients treated with unoprostone for up to 24 months; no confirmed cases were reported in the timolol or betaxolol groups. In the unoprostone group, cases of iris color change were confirmed at months 12 (1 case), 18 (2 cases), and 24 (4 cases). No clinically relevant differences were observed among treatment groups for changes from baseline in eyelash length or density. CONCLUSION: Although iris hyperpigmentation and abnormal eyelash changes may occur after treatment with unoprostone, the incidence of these events appears to be low in the 2-year clinical study. PMID- 15288976 TI - Retinal venous pulsation in glaucoma and glaucoma suspects. AB - PURPOSE: To determine whether changes in central retinal vein pulsation characteristics occur in glaucoma, and how these are related to indices of glaucoma severity. DESIGN: A large, consecutive, prospective, case-controlled study. PARTICIPANTS: Ninety-four consecutive glaucoma patients and 105 glaucoma suspects seen in a tertiary referral clinic were examined. Forty-one age-matched normal subjects also were examined. METHODS: The presence or absence of spontaneous venous pulsation was observed in these 3 groups. The ophthalmodynamometric force (ODF) required to induce venous pulsation at the optic disc was measured in those without spontaneous pulsation. Optic disc photographs were obtained and visual field testing was performed for all subjects. MAIN OUTCOME MEASURES: The prevalence of spontaneous venous pulsation between these 3 groups was compared. The relationship between ODF and visual field mean deviation, neuroretinal rim area, age, intraocular pressure (IOP), gender, and diagnosis of glaucoma was investigated using linear mixed models fitted by Gibb's sampling. RESULTS: Significantly fewer (chi-square, 27.7; P<0.001) glaucoma patients (54%) were observed to have spontaneous venous pulsation than suspects (75%) or normals (98%). A worse visual field mean deviation was shown to be the most significant predictor of a higher ODF (P<0.000), with younger age (P<0.000) also predictive of a higher ODF. A strong relationship between ODF and mean deviation was found in the glaucoma patients (r = 0.59; n = 52; P<0.001). CONCLUSIONS: Spontaneous venous pulsation is less common in glaucoma. The ODF required to induce venous pulsation is increased in glaucoma, and this ODF is greater in those with more severe field loss. PMID- 15288977 TI - Bleb-associated endophthalmitis: clinical characteristics and visual outcomes. AB - PURPOSE: To analyze the clinical characteristics and treatment outcomes of patients with bleb-associated endophthalmitis (BAE). DESIGN: Retrospective, noncomparative, interventional case series. PARTICIPANTS: Consecutive patients treated at one institution for BAE. INTERVENTIONS: Prompt pars plana vitrectomy (PPV) with intravitreal injection of antibiotics, or prompt vitreous biopsy and intravitreal injection of antibiotics (tap and inject). METHODS: Retrospective analysis of 68 consecutive cases of BAE between July 1, 1989 and June 30, 2001. Clinical presentation, treatment modality, microbiologic data, and clinical course were analyzed. Visual outcomes were compared between vitrectomy and tap and-inject groups, culture-positive and culture-negative groups, and early and late times. MAIN OUTCOME MEASURES: Snellen visual acuities (VAs) at 3 months and 12 months after treatment and at most recent follow-up. RESULTS: The incidence of no light perception (NLP) at 12 months after treatment for BAE was 35%. Vitreous isolates included streptococcal species (32% of positive cultures), Staphylococcus epidermidis (26%), Enterococcus, and Serratia (12% each). Patients with a positive vitreous culture had significantly worse VA (median, hand movements [HM] at 3 and 12 months after treatment) and a higher rate of NLP vision. Patients treated with tap-and-inject had a significantly worse final VA (medians, HM at 3 months and LP at 12 months) and a significantly higher rate of NLP vision than patients treated with PPV. One third of patients who underwent PPV achieved a final VA of 20/100 or better 12 months after treatment (P = 0.09). CONCLUSIONS: Bleb-associated endophthalmitis causes significant visual morbidity. Patients with culture-negative BAE and patients treated with prompt PPV may achieve better visual outcome. PMID- 15288979 TI - Impact of cataract on the results of frequency-doubling technology perimetry. AB - PURPOSE: To determine the effect of cataract on the results of frequency-doubling technology (FDT) perimetry. DESIGN: Consecutive cohort study. PARTICIPANTS: Forty four patients with normal ophthalmic examinations, with the exception of cataract, scheduled to undergo phacoemulsification and posterior chamber lens implantation were prospectively identified and completed the study. METHODS: All subjects underwent FDT perimetry using the full-threshold C-20 strategy. Both eyes were tested within 1 month before cataract surgery and up to 3 months after surgery. The unoperated fellow eyes served as controls. MAIN OUTCOME MEASURES: Changes in visual acuity (VA), mean deviation (MD), and pattern standard deviation (PSD) were evaluated. For each subject, the change in MD and PSD in the eye that underwent cataract surgery was adjusted for change in the control eye that is thought to occur due to a learning effect. RESULTS: Among the eyes that underwent cataract surgery, the median preoperative VA was 20/60 (range, 20/30 20/800) and the mean preoperative MD was -4.00+/-3.72 decibels (dB). Postoperatively, the median VA improved to 20/30 (range, 20/20-20/70) and the mean postoperative MD was -0.26+/-3.09 dB (P<0.001). Among the control eyes, MDs were -1.74+/-3.71 dB preoperatively and -0.94+/-3.85 dB postoperatively (P = 0.019). The adjusted improvement in MD among eyes that underwent cataract surgery was 2.94+/-3.44 dB (P<0.001). There was no significant change in PSD. Preoperative VA correlated significantly with preoperative MD (r = 0.39, P = 0.01). The improvement in VA correlated significantly with the adjusted improvement in MD (r = 0.38, P = 0.01). CONCLUSIONS: Cataract has an adverse effect on the MD but not the PSD in FDT perimetry. Among eyes with visually significant cataract, the MD correlates significantly with VA. After cataract surgery, the change in VA correlates significantly with the adjusted change in MD. PMID- 15288980 TI - Detection of undiagnosed glaucoma by eye health professionals. AB - PURPOSE: To examine the clinical features of undiagnosed open-angle glaucoma (OAG) in people who have attended an eye care provider within the previous 12 months and to suggest strategies to assist in the early detection of glaucoma. DESIGN: Population based cross-sectional study. PARTICIPANTS: Permanent residents aged 40 years and older at recruitment during 1992 through 1996. METHODS: A cluster-stratified random sample of 4744 participants from the urban and rural cohorts was studied. Structured standardized interviews and dilated ocular examinations were conducted in all eligible participants. Data on demographic characteristics, prior knowledge of eye disease, use of eye care services, intraocular pressures, cup-to-disc ratios, visual fields, and photography of optic discs were obtained. All suspected glaucoma cases were submitted to a panel of 6 ophthalmologists to determine glaucoma diagnosis. MAIN OUTCOME MEASURES: Clinical features of participants seen by eye health professionals within the previous 12 months who have previously undiagnosed OAG, previously diagnosed OAG, and no glaucoma. RESULTS: Thirty-five previously undiagnosed and 43 previously diagnosed participants had visited an optometrist or ophthalmologist or both in the previous 12 months. Age and gender were not significantly different between the undiagnosed and diagnosed glaucoma cases. After logistic regression, the type of eye professional seen (odds ratio [OR], 45.17; 95% confidence interval [95% CI], 5.89-346.17; P = 0.0002) and the presence of visual field defects (OR, 0.06; 95% CI, 0.01-0.69, P = 0.020) were the only statistically significant variables between the diagnosed and undiagnosed glaucoma groups. CONCLUSIONS: Raised intraocular pressure should not be relied on as the only triggering factor in glaucoma investigations. PMID- 15288981 TI - Laserinterferometric assessment of pilocarpine-induced movement of an accommodating intraocular lens: a randomized trial. AB - PURPOSE: To measure the axial movement of an accommodating intraocular lens (IOL) induced by ciliary muscle contraction after application of pilocarpine. DESIGN: Randomized, controlled, patient- and examiner-masked trial with intrapatient comparison. PARTICIPANTS AND CONTROLS: One hundred ten eyes of 55 patients with age-related bilateral cataract. METHODS: This study was divided into 3 parts. In the first, the accommodating IOL (1CU) was compared with a 3-piece open-loop acrylic IOL that served as the control. In the second, to assess the effect of capsule fibrosis on the potential accommodating performance of the accommodating IOL, extensive polishing of the anterior capsule with a slit cannula was compared with standard surgery. In the third, the effect of a posterior capsulorhexis was compared with that of standard surgery. Anterior chamber depth (ACD) was assessed with partial coherence interferometry, measured before and after topical application of pilocarpine 2%, and near visual acuity (VA) was evaluated 3 months after surgery. MAIN OUTCOME MEASURE: Pilocarpine-induced change in ACD. RESULTS: The accommodating IOL showed a forward movement under pilocarpine with a median amplitude of movement of -314 microm (95% confidence interval [CI]: -148 to 592), compared with the backward movement of 63 microm (95% CI: 161 to -41) for the open-loop control IOL (P = 0.001). Capsule polishing and a posterior capsulorhexis had no effect on IOL movement with the accommodating IOL. The median near VA with distance correction was 20/60. CONCLUSION: Pilocarpine induced a small but significant forward movement of the accommodating IOL. However, the amount of movement was calculated to result in a refractive change of <0.5 diopters (D) in most patients, reaching 1 D or slightly more in only single cases, with a large variability of movement. Neither polishing of the capsule bag nor a posterior capsulorhexis could enhance the accommodative ability. PMID- 15288982 TI - Long-term effects of mitomycin C in pterygium surgery on scleral thickness and the conjunctival epithelium. AB - PURPOSE: To evaluate the long-term effects of intraoperative application of mitomycin C on the scleral thickness and the conjunctival epithelium at the surgical site of pterygium excision. DESIGN: Prospective observational case series. PARTICIPANTS: Twenty-four patients who underwent excision of primary pterygium with intraoperative mitomycin C in our department during the year 1996. METHODS: Patients were evaluated by slit-lamp biomicroscopy, impression cytology, and high-frequency ultrasonography. Impression cytology was performed by applying a small nitrocellulose filter paper for a few seconds at the excision area and for a few seconds at the opposite perilimbal area, and subjecting the specimens to the periodic acid-Schiff-Gill modified Papanicolaou staining protocol. The morphology of the conjunctival epithelium and goblet cell density (GCD) were recorded. High-frequency ultrasound was performed at the same sites, and the scleral thickness was measured at a distance of 1 mm from the limbus. MAIN OUTCOME MEASURES: Goblet cell density, conjunctival epithelial morphology, and the scleral thickness at the operated and nonoperated sites. RESULTS: All patients had successful pterygium removal with no corneal recurrence after a mean follow-up of 77.2+/-3.9 months (range, 72-84). Impression cytology revealed normal nongoblet conjunctival epithelial cells at the excision area, with a 4 fold decrease in the GCD at the excision area when compared with the contralateral nonoperated site (296+/-120 cells/mm(2) and 1183+/-310 cells/mm(2), respectively; P = 0.0036). No differences were noted between the scleral thicknesses at the operated site (750+/-70 microm) and the opposite site (740+/ 80 microm) (P = 0.84). CONCLUSIONS: A single application of mitomycin C after pterygium excision is not associated with reduction in scleral thickness more than 6 years postoperatively. The conjunctival epithelium retains its normal phenotype, with a marked reduction of the GCD. PMID- 15288983 TI - Penetrating limbo-keratoplasty for granular and lattice corneal dystrophy: survival of donor limbal stem cells and intermediate-term clinical results. AB - PURPOSE: To assess clinical follow-up data, and to identify donor epithelial cells after homologous penetrating central limbo-keratoplasty in patients with granular and lattice corneal dystrophies compared with patients who underwent conventional penetrating keratoplasty (PK). DESIGN: Mixed retrospective and prospective nonrandomized comparative case series. PARTICIPANTS AND CONTROLS: Twenty-six patients who underwent 33 limbo-keratoplasty procedures for granular or lattice corneal dystrophy since May 1995 and a historical control group of 24 patients who underwent 36 PK procedures between November 1986 and May 1995. METHODS: Postoperatively, all but 2 limbo-keratoplasty patients were treated with systemic immunosuppressants for 6 months. All patients received long-term topical immune prophylaxis with prednisolone-21-acetate 1% (2 drops per day). After obtaining informed consent, epithelial cells were harvested from 10 limbo keratoplasty eyes of 8 patients with granular dystrophy and 7 limbo-keratoplasty eyes of 7 patients with lattice dystrophy. Conjunctival epithelium or buccal mucosal epithelium for recipient identification and corneal epithelial cells from 3 graft sites were harvested. Deep-frozen donor corneoscleral rims were analyzed to characterize donor features. Genetic analysis was performed by polymerase chain reaction of short tandem repeat (STR) loci. MAIN OUTCOME MEASURES: The ratio of dystrophy recurrences in the graft was clinically assessed. Donor features in epithelial cells were genetically established if at least 1 STR profile differed from that of the recipient. RESULTS: There were 5 recurrences in limbo-keratoplasty eyes with granular dystrophy and 2 recurrences in limbo keratoplasty eyes with lattice dystrophy, compared with 15 and 6 recurrences in PK eyes, respectively. The differences between limbo-keratoplasty and PK were not statistically significant over time (log-rank test; P = 0.14 for granular dystrophy and P = 0.56 for lattice dystrophy; alpha error, 0.05). For genetic analysis, 12 of 17 samples were evaluated. Donor epithelial cells were detected in 5 of the 12 samples (42%). CONCLUSIONS: Limbo-keratoplasty tended to be associated with fewer recurrences of granular and lattice dystrophies. However, the difference was not yet statistically significant, probably due to the disappearance of the transplanted limbal stem cells over time. Genetic analysis confirmed the survival of transplanted limbal stem cells over several years in some limbo-keratoplasty eyes, which might correlate with less recurrence. Limbo keratoplasty, therefore, is likely to represent a first step towards long-term recurrence-free survival of corneal grafts in patients with granular and lattice dystrophies. PMID- 15288985 TI - The giant fornix syndrome: an unrecognized cause of chronic, relapsing, grossly purulent conjunctivitis. AB - AIM: To describe a group of elderly patients presenting with chronic, relapsing, copiously purulent conjunctivitis, in which the condition was often perpetuated by the sequestration of a large number of bacteria on a protein coagulum lodged in the recesses of a large upper conjunctival fornix. PATIENTS AND METHODS: Retrospective review of a noncomparative case series, drawn from patients attending the lacrimal clinic at Moorfields Eye Hospital. OUTCOME MEASURES: Characterization of this unrecognized syndrome and its response to treatment. RESULTS: Twelve patients (10 female) presented between the ages of 77 and 93 years (mean, 85; median, 86) with a history of chronic relapsing bacterial conjunctivitis affecting, with 2 exceptions, just one eye. All had experienced multiple episodes of markedly purulent conjunctivitis and chronic ocular discharge for between 8 and 48 months (mean, 23.5; median, 24) before referral, and the patients had received multiple courses of treatment. Three had successful external dacryocystorhinostomy (for nasolacrimal duct occlusion) before the final diagnosis of giant fornix syndrome was made, 9 had developed corneal vascularization and scarring before referral, and 5 had suffered prior spontaneous corneal perforation or thinning. All patients had deep upper conjunctival fornices in association with the changes of age-related dehiscence of the levator muscle aponeurosis. Copious amounts of thick, purulent debris and a yellow coagulum were lodged in the depths of the upper fornix-this debris universally culturing Staphylococcus aureus. The condition settled rapidly on appropriate systemic antibiotics (ciprofloxacin or ofloxacin), intensive topical antibiotics, and high-dose, high-potency steroids; some patients required repeated treatment or needed to continue the use of a single drop of a combined steroid-antibiotic to prevent relapse. CONCLUSION: The capacious upper fornix of the elderly may harbor a coagulum colonized by S. aureus, leading to chronic conjunctivitis that may lead to severe sight impairment due to toxic keratopathy and secondary corneal vascularization. PMID- 15288984 TI - Influence of donor storage time on corneal allograft survival. AB - PURPOSE: To investigate the influence of graft storage time on corneal allograft survival in high-risk and low-risk patients. DESIGN: Comparative retrospective nonrandomized clinical trial. PARTICIPANTS: Overall, 193 patients with 210 corneal allografts were classified as high risk or low risk for corneal allograft rejection on the basis of recipient corneal neovascularization and number of ipsilateral transplants. METHODS: Data from 3 groups were evaluated. The first group received fresh (no storage in culture medium) corneas, the second received corneas of donor storage time less than 7 days in minimum essential medium (MEM) at 37 degrees C, and the final group received corneas stored in MEM longer than 7 days. Recipients were analyzed for development of immune rejection according to storage time of the corneal tissue. Corneal allograft survival rate was determined with Kaplan-Meier survival analysis. The log-rank test was used to determine statistical significance. MAIN OUTCOME MEASURES: Risk of reversible and irreversible allograft rejection in corneas stored at various intervals in high corneal and low-corneal transplant patients. RESULTS: High-risk corneal allograft recipients had a significantly prolonged allograft survival when the tissue was stored for 7 days or greater, compared with recipients receiving fresh tissues. Patients at low risk of corneal allograft rejection also showed a tendency for prolonged survival, although not statistically significant (P = 0.06). CONCLUSIONS: Storage of corneal tissue may reduce the frequency of allograft rejection, especially in high-risk patients. PMID- 15288986 TI - Corneal and conjunctival toxicity from hydrogen peroxide: a patient with chronic self-induced injury. AB - PURPOSE: To report a patient with severe corneal and conjunctival toxicity from long-term, habitual use of hydrogen peroxide as an eye wash. DESIGN: Observational case report. INTERVENTION AND TESTING: Serial examinations of the cornea, conjunctiva, and ocular adnexa were done. Penetrating keratoplasty with amniotic membrane transplantation was performed. MAIN OUTCOME MEASURES: Ocular inflammation, pain, and visual acuity outcome. RESULTS: Bilateral corneal and conjunctival inflammation and scarring mimicking ocular-cicatricial pemphigoid were noted. Formation of a descemetocele after starting treatment with low-dose topical steroids required emergent penetrating keratoplasty with amniotic membrane transplantation. CONCLUSIONS: This is the first reported case of ocular surface toxicity in a patient after deliberate chronic use of high-dose hydrogen peroxide. It highlights the value of obtaining a thorough medical and social history and the importance of direct questioning about the use of any medications or agents on the eyes before making a diagnosis or initiating therapy. PMID- 15288987 TI - The emotional impact of amblyopia treatment in preschool children: randomized controlled trial. AB - PURPOSE: To investigate the emotional status of children undergoing active treatment for amblyopia. DESIGN: Postal survey, in the context of a prospective, multicenter, randomized controlled trial. PARTICIPANTS: Parents of 177 children with a unilateral visual impairment referred from preschool vision screening. The children had been recruited to a randomized controlled trial of treatment for unilateral visual impairment and randomly assigned to receive either glasses with or without patches, glasses alone, or treatment deferred for 1 year. METHODS: A self-completion questionnaire, including a psychometric behavioral scale, was sent to the parents of all children recruited to the trial at age 4 years, to 66 whose deferred treatment began at age 5 years, and finally to 151 remaining in the trial at the end of follow-up. MAIN OUTCOME MEASURES: Mean scores per treatment group on the Revised Rutter Parent Scale for Preschool Children. Comparison of parent responses to questions assessing the child's general well being and difficulties associated with treatment. RESULTS: Completed questionnaires were returned for 144 of 177 (81%) children at a mean age (standard deviation) of 48 months (5.0), for 45 of 66 (68%) at a mean age of 61 months (5.8), and for 78 of 151 (52%) at a mean age of 67 months (5.0). Most parents reported having difficulty with patching their child regardless of age (77% at age 4 years and 73% at age 5 years), with fewer reporting difficulties with glasses alone (42% and 53%, respectively). Children were significantly more upset by patching than by glasses only (chi-square test, P = 0.03 for age 4 years and P = 0.01 for age 5 years), as were the parents of 4-year-olds (chi-square test, P = 0.01). Most parents thought their children were happy, cooperative, and good tempered, and behavioral scores did not differ between treatment groups. CONCLUSIONS: Treatment for unilateral visual impairment is not easy to implement and is commonly associated with some degree of distress. Despite this, no impact on the child's global well-being or behavior was seen either during or after the treatment period. PMID- 15288988 TI - Orbital irradiation for Graves' ophthalmopathy: Is it safe? A long-term follow-up study. AB - PURPOSE: We evaluated the frequency of long-term complications of orbital irradiation (radiation-induced tumors, cataract, and retinopathy) in comparison with glucocorticoids. DESIGN: We conducted a follow-up study in a cohort of 245 Graves' ophthalmopathy patients who had been treated with retrobulbar irradiation (20 Gy in 2 weeks) and/or oral glucocorticoids between 1982 and 1993 in our institution. Irradiated patients were compared with nonirradiated patients. METHODS: Data on mortality and cause of death were obtained. Living patients were invited to participate in a follow-up study. Possible retinopathy was assessed in a masked fashion and defined as the presence of > or =1 hemorrhages and/or microaneurysms on red-free retina photographs. If >5 lesions were present, patients were categorized as suffering from definite retinopathy. Cataract was assessed using the Lens Opacity Classification System II score. MAIN OUTCOME MEASURES: Mortality, prevalence of retinopathy, prevalence of cataract, and type of cataract. RESULTS: Thirty-seven of the 245 patients had died, none of them from an intracranial tumor. Mortality was similar in the irradiated (27/159 [17%]) and nonirradiated patients (10/86 [12%]; P = 0.264). One hundred fifty seven of the 208 living patients (75%) consented to participate in a follow-up ophthalmologic investigation; the mean follow-up time (+/- standard deviation) was 11+/-3 years. Possible retinopathy was present in 15% of patients, 22 of the irradiated and 1 of the nonirradiated patients (P = 0.002). In 5 patients (all had been irradiated), definite retinopathy (i.e., >5 retinal lesions) was present. Of these, 3 had diabetes mellitus, and 1 had hypertension. Diabetes was associated with both possible (P = 0.029) and definite (P = 0.005) retinopathy, with a relative risk of 21 (95% confidence interval, 3-179). The prevalence and severity of cataract were similar in the radiotherapy group (29%) and the glucocorticoid group (34%); it should be noted that 88 of 104 of the irradiated patients were also treated with oral glucocorticoids. CONCLUSION: The data suggest that orbital irradiation for Graves' ophthalmopathy is a safe treatment modality, except possibly for diabetic patients. PMID- 15288989 TI - Muscle surgery in patients with Graves' disease using topical anesthesia. AB - PURPOSE: To report our experience in extraocular muscle surgery for Graves' disease using topical anesthesia. DESIGN: Retrospective, noncomparative, observational case series. PARTICIPANTS: In 135 patients with Graves' disease, a total of 200 ocular muscles were operated during the past 20 years at the Department of Ophthalmology, Ludwig-Maximilian-University, Munich, Germany. METHODS: Surgery was performed under topical anesthesia with tetracaine hydrochloride 1% eyedrops. Because of the restrictive nature of the motility impairment, recession of the muscles was used in all patients. The exact amount of recession was determined during the operation with active cooperation from the patient. MAIN OUTCOME MEASURES: Binocular single vision and the angle of deviation were evaluated preoperatively and postoperatively. RESULTS: Postoperative binocular single vision in the primary position was achieved by 78.7% of the patients on the first postoperative day. Subsequent evaluation demonstrated binocular single vision in 91.9% of all patients and in 96.4% of the group with only 1 muscle (inferior rectus) operated. CONCLUSIONS: The authors have demonstrated that topical anesthesia is a feasible and reliable method for performing extraocular muscle surgery in patients with Graves' disease. Intraoperative patient discomfort seemed insignificant, and the active cooperation of the patient in finding the appropriate extent of surgery was advantageous. The overall results showed that deviation surgery with the use of topical anesthesia is highly successful in restoring binocular single vision in patients with endocrine orbitopathy. PMID- 15288990 TI - Droperidol and dolasetron alone or in combination for prevention of postoperative nausea and vomiting after vitrectomy. AB - PURPOSE: Droperidol and the new serotonin-3 antagonists are effective drugs for the prophylaxis of postoperative nausea and vomiting (PONV). The aim of this trial was to evaluate whether dolasetron could be a substitute for droperidol, because the Food and Drug Administration has required a Black Box warning on the droperidol package insert. DESIGN: Randomized, placebo-controlled, double-blinded trial. PARTICIPANTS: Inpatients undergoing vitreoretinal surgery (standard 3-port pars plana vitrectomy for proliferative diabetic vitreoretinopathy, complicated retinal detachment, or macular disease, such as macular pucker, macular hole, or choroidal neovascularization). INTERVENTION: Two hundred forty patients (3x80) receiving droperidol (10 microg. kg(-1)), dolasetron (12.5 mg), or the combination of both drugs administered 5 to 10 minutes before the end of surgery. CONTROL: Eighty patients received saline placebos as controls. METHODS: Standardized general anesthesia was performed, including benzodiazepine premedication, propofol, atracurium or vecuronium, desflurane in N(2)O/O(2), and a continuous infusion of remifentanil. Postoperative analgesia and antiemetic rescue medication were standardized. Episodes of vomiting, retching, nausea, and the need for additional antiemetics were recorded for 24 hours. MAIN OUTCOME MEASURES: Severity of PONV (rated by a standardized scoring algorithm) was analyzed as the main end point of the study using the Kruskal-Wallis test. RESULTS: Data of 304 patients could be analyzed. Mean severity scores in the placebo, dolasetron, droperidol, and combination groups were 1.21, 0.76, 0.47, and 0.30. Incidences of PONV of any severity were 56%, 40%, 28%, and 18%, respectively. The reduction of the incidence of PONV and its severity was statistically significant in the droperidol group and in the combination group relative to the placebo. Dolasetron alone failed to reduce the incidence of PONV. The combination of dolasetron and droperidol showed an additive antiemetic efficacy. CONCLUSION: Low-dose droperidol (10 microg. kg(-1)) but not dolasetron (12.5 mg) reduced postoperative nausea and vomiting after vitreoretinal surgery. Dolasetron (12.5 mg) is not an equivalent substitute for droperidol. PMID- 15288991 TI - Photodynamic therapy with verteporfin for symptomatic polypoidal choroidal vasculopathy: one-year results of a prospective case series. AB - OBJECTIVE: To determine the efficacy of photodynamic therapy (PDT) with verteporfin as a treatment for symptomatic polypoidal choroidal vasculopathy (PCV). DESIGN: Prospective consecutive, 2-centered, noncomparative interventional case series. PARTICIPANTS: Twenty-one Asian patients with 22 eyes presenting with serosanguinous maculopathy due to PCV and an initial best-corrected visual acuity (BCVA) of 20/40 or worse were recruited prospectively. All patients had angiographic leakage seen on fluorescein angiograms (FAs) and features of PCV seen with indocyanine green (ICG) angiography. METHODS: Intravenous infusion of verteporfin at a dose of 6 mg/m(2) of body surface area over 10 minutes was administered. Five minutes after the completion of infusion, a 689-nm laser was applied for 83 seconds, with a light dose of 50 J/cm(2). The laser spot size was chosen to cover the polyps and the surrounding abnormally dilated choroidal vessels shown on ICG angiography plus an extra 1000-microm margin. Photodynamic therapy retreatment was performed if leakage from the polyps was found on both repeat FAs and ICG angiography at regular 3-month follow-up intervals. MAIN OUTCOME MEASURES: The proportion of eyes with stable or improved vision at a 1 year follow-up. Secondary outcome measures included change in mean BCVA and the changes in clinical and angiographic features in FAs and ICG angiography. The total number of PDT sessions and any complications were also recorded. RESULTS: Stable or improved vision was achieved in 21 (95%) of the 22 eyes at the 1-year follow-up. Ten (45%) eyes had a moderate gain in vision (improved by > or =3 lines), whereas 1 (5%) eye suffered a moderate visual loss (decrease by > or =3 lines). The mean BCVA improved from a logarithm of the minimum angle of resolution(logMAR) of 0.73 to 0.60, an equivalent of 1.3 lines of improvement. The change in logMAR BCVA at 12 months was statistically significant (Wilcoxon signed-ranks test, P = 0.009). Complete absence of leakage in FAs and total regression of the polyps in ICG angiography were observed in 20 (91%) and 21 (95%) eyes, respectively. Severe loss of vision due to massive subretinal hemorrhage occurred in 1 eye; otherwise, there were no other serious treatment related adverse events. CONCLUSIONS: The 1-year results of PDT in treating PCV of the macular type with serosanguinous presentations are encouraging. Further studies with longer follow-up and randomized controlled trials are warranted to assess the long-term safety and efficacy of PDT relative to observation or other treatment modalities. PMID- 15288993 TI - Retinotopic mapping of the visual cortex using functional magnetic resonance imaging in a patient with central scotomas from atrophic macular degeneration. AB - PURPOSE: To describe retinotopic mapping of the visual cortex when a central scotoma is present. DESIGN: Single observational case report. METHODS: Scanning laser ophthalmoscope perimetry was used to define the site and stability of fixation and the area of dense scotoma. Functional magnetic resonance imaging of the visual cortex was performed while the patient viewed an expanding annular stimulus. RESULTS: Retinotopic mapping of the visual cortex for a patient with a horseshoe scotoma from geographic atrophy involving the macular region showed a loss of stimulation to the cortical areas representing the site of the atrophic lesion. CONCLUSIONS: Cortical retinotopic mapping can be performed successfully in patients with central scotomas from macular disease. This study can serve as a basis for the future investigation of cortical plasticity in visual cortex. PMID- 15288992 TI - Lack of fundus autofluorescence to 488 nanometers from childhood on in patients with early-onset severe retinal dystrophy associated with mutations in RPE65. AB - PURPOSE: Fundus autofluorescence is due to accumulation of lipofuscin in the retinal pigment epithelium (RPE) resulting from incomplete digestion of N retinylidene-phosphatidyl-ethanolamine from shed photoreceptor outer segment discs. Alteration in autofluorescence reflects changes in lipofuscin content of the RPE. Mutations on both alleles of RPE65 result in absent or largely decreased formation of rhodopsin, due to a defect in all-trans retinol isomerization in the RPE. Autofluorescence could therefore be altered. This study was conducted to evaluate fundus autofluorescence in patients with early-onset severe retinal dystrophy (EOSRD, or early-onset rod-cone dystrophy) associated with mutations on both alleles of RPE65. DESIGN: Case series. PARTICIPANTS AND CONTROLS: Ten 10- to 55-year-old patients with EOSRD and compound heterozygous or homozygous mutations in RPE65. For comparison, 6 heterozygous parents and 2 patients with other forms of EOSRD were examined. METHODS: Participants underwent, in addition to standard clinical and electrophysiological examination, autofluorescence imaging using a confocal scanning laser ophthalmoscope. Three of the patients were also examined by optical coherence tomography (OCT) to evaluate the status of retinal degeneration. Mutations in 7 patients have been reported previously; the other patients were investigated by polymerase chain reaction-single-strand conformation polymorphism and direct sequencing for mutations in RPE65 and lecithin retinol acyltransferase (LRAT). MAIN OUTCOME MEASURES: Fundus autofluorescence and OCT. RESULTS: Absent or minimal autofluorescence was found in all patients with compound heterozygous or homozygous RPE65 mutations. Autofluorescence was normal in the heterozygous parents. Autofluorescence was present in 2 children with EOSRD not associated with mutations in RPE65 or LRAT, another gene involved in retinol recycling. Optical coherence tomography in younger patients revealed an intraretinal appearance similar to that of their healthy, heterozygous parents. CONCLUSIONS: Lack of autofluorescence in patients with EOSRD associated with mutations in RPE65 is in accordance with the biochemical defect and can be used as a clinical marker of this genotype. Optical coherence tomography results in younger patients would indicate still viable photoreceptors despite the absence of autofluorescence. PMID- 15288994 TI - Milder ocular findings in Hermansky-Pudlak syndrome type 3 compared with Hermansky-Pudlak syndrome type 1. AB - PURPOSE: To compare clinically 2 different subtypes of Hermansky-Pudlak syndrome (HPS), type 1 (HPS-1) and type 3 (HPS-3). DESIGN: Cross-sectional study of a series of patients. PARTICIPANTS: Sixteen patients with HPS-1 and 14 patients with HPS-3 were studied. METHODS: Complete eye examination, including best corrected visual acuity and photographs and photographic grading of iris transillumination and macular transparency using a previously established grading system. RESULTS: Snellen visual acuity was 20/160-2 in the HPS-1 group and 20/125+2 in the HPS-3 group (P = 0.017). Iris grading was statistically significant for less translucence in the HPS-3 patients. The HPS-3 patients also tended to have less transparent maculas, but the difference was not statistically significant. CONCLUSIONS: Patients with HPS-3 have less severe ophthalmic manifestations than patients with HPS-1. Ophthalmologists treating patients with albinism should consider HPS in their differential diagnosis even in the case of mild iris and macular hypopigmentation. PMID- 15288995 TI - LASIK for hyperopia, hyperopic astigmatism, and mixed astigmatism: a report by the American Academy of Ophthalmology. AB - OBJECTIVE: To describe LASIK for hyperopia, hyperopia with astigmatism, and mixed astigmatism and to examine the evidence to answer questions about the safety and efficacy of the procedure. METHODS: A literature search conducted for the years 1968 to 2002 retrieved 118 citations. During review and preparation of this article, an additional 2 articles were included. The panel members selected 36 articles for the panel methodologist to review and rate according to the strength of evidence. A level I rating is assigned to properly conducted, well-designed, randomized clinical trials; a level II rating to well-designed cohort and case control studies; and a level III rating to case series, case reports, and poorly designed prospective and retrospective studies. RESULTS: This assessment describes 5 nonrandomized interventional trials (level II), 3 nonrandomized comparative trials (level III), and 20 noncomparative case series (level III). Additionally, 6 single-case reports (level III) were included because they reported relevant complications, and 2 theoretical analyses (level III) were also considered. This assessment does not compare studies because many variables such as range of hyperopia, follow-up periods, lasers, microkeratomes, techniques, and surgeon experience have not been controlled. CONCLUSIONS: For low (<3 diopters [D]) to moderate (3-5 D) hyperopia, results from published studies (levels II and III evidence) have shown that LASIK is effective and predictable in achieving very good to excellent uncorrected visual acuity, achieving postoperative refractions within 1 D of emmetropia, and is safe in terms of minimal loss of best-corrected spectacle vision. Although there are fewer data for hyperopic astigmatism, the results available seem to mirror the data for low to moderate hyperopia (levels II and III evidence). The postoperative results for both uncorrected vision and safety are less compelling, as greater amounts of hyperopia are treated (>4 to 5 D). Utilizing hyperopic LASIK for the treatment of consecutive hyperopia and astigmatism is also effective, although the ability to reduce hyperopic astigmatism after radial keratotomy is limited. Although a variety of ablation profiles can be used to treat mixed astigmatism, very good visual results have been reported (levels II and III evidence). Serious adverse complications leading to permanent visual loss are possible but, fortunately, very rare. There are insufficient data to compare one laser system with another or one ablation profile with another. PMID- 15288996 TI - Medication use before cataract surgery. PMID- 15288997 TI - Oxybuprocaine induces a false positive. PMID- 15288999 TI - Night vision complaints after LASIK. PMID- 15289001 TI - Night vision complaints after LASIK. PMID- 15289003 TI - Trypan blue staining in vitreoretinal surgery. PMID- 15289005 TI - Retinal detachment and uveitis. PMID- 15289006 TI - The somatodendritic release of dopamine in the ventral tegmental area and its regulation by afferent transmitter systems. AB - The release of dopamine in the ventral tegmental area (VTA) plays an important role in the autoinhibition of the dopamine neurons of the mesocorticolimbic system through the activation of somatodendritic dopamine D2 autoreceptors. Accordingly, the intra-VTA application of dopamine D2 receptor agonists reduces the firing rate and release of dopamine in the VTA, and this control appears to possess a tonic nature because the corresponding antagonists enhance the somatodendritic release of the transmitter. In addition, the release of dopamine in the VTA is increased by potassium or veratridine depolarization and abolished by tetrodotoxin and calcium omission. Overall, it appears that the somatodendritic release of dopamine is consistently lower than that in nerve endings. Apart from intrinsic dopaminergic mechanisms, other transmitter systems such as serotonin, noradrenaline, acetylcholine, GABA and glutamate play a role in the control of the activity of dopaminergic neurons of the VTA, although the final action depends on the particular receptor involved as well as the neuronal type where it is localized. Given the involvement of the mesocorticolimbic dopaminergic systems in the pathogenesis of severe neuropsychiatric disorders such as schizophrenia, the knowledge of the factors that regulate the release of dopamine in the VTA could provide new insight into the ethiogenesis of the disease as well as its implication on the mechanisms of action of therapeutic drugs. PMID- 15289007 TI - Statistical and conceptual issues in defining post-operative cognitive dysfunction. AB - The occurrence of post-operative cognitive dysfunction is a distressing complication following surgery. In an effort to gain a more complete understanding of patients' cognitive recovery following surgical procedures common neuropsychological assessment tools have been adopted in a repeated measures design. It is widely regarded that this represents the most comprehensive method of determining cognitive status in this population but it has resulted in a number of statistical and conceptual difficulties in attempting to infer significant change. The current paper outlines these core difficulties and provides some potential methods to overcome these. PMID- 15289008 TI - Reinforcement-related brain potentials from medial frontal cortex: origins and functional significance. AB - The development of the field of cognitive neuroscience has inspired a revival of interest in the brain mechanisms involved in the processing of rewards, punishments, and abstract performance feedback. One fruitful line of research in this area was initiated by the report of an electrophysiological brain potential in humans that was differentially sensitive to negative and positive performance feedback [J. Cogn. Neurosci. 9 (1997) 788]. Here we review current knowledge regarding the neural basis and functional significance of this feedback-evoked 'error-related negativity' (ERN). Our review is organized around a set of predictions derived from a recent theory, which holds that the ERN is associated with the arrival of a negative reward prediction error signal in anterior cingulate cortex. PMID- 15289009 TI - Telomeres and telomerase in normal and leukemic hematopoietic cells. AB - Telomere length and telomerase have an important role in normal and malignant hematopoiesis. Telomere erosion can lead to chromosome end fusion and thereby contribute to genomic instability during tumorigenesis. Thus, like complex chromosomal aberrations, telomere length may be a prognostic factor in hematopoietic malignancies. A paper by Sieglova et al. in this issue of Leukemia Research reports on the prognostic impact of telomere shortening in bone marrow (BM) and peripheral blood (PB) specimens of myelodysplastic syndrome (MDS) and MDS converted-AML patients (pts). Their results underline the importance to study telomere biology together with cytogenetics, genomic and proteomic profiling as prognostic factors, in order to improve risk-adapted therapy of MDS and AML pts. PMID- 15289010 TI - Extramedullary acute myeloid leukemia infiltrates. PMID- 15289011 TI - Extramedullary infiltrates of AML are associated with CD56 expression, 11q23 abnormalities and inferior clinical outcome. AB - We evaluated the frequency and prognostic significance of extramedullary infiltrates (EMI) at presentation of acute myeloid leukemia (AML) in adult patients. Of 331 cases with de novo AML, 101(30.5%) had extramedullary infiltrates at diagnosis. The extramedullary manifestations included: lymphadenopathy, splenomegaly, hepatomegaly, gingival hypertrophy, skin infiltrates and involvement of central nervous system (CNS). Patients with EMI had a high initial WBC count and a high proportion of M4/M5 morphological variants. The complete remission rate (CR) with induction chemotherapy was lower in patients with EMI (P=0.0077) and their overall survival was also inferior (P=0.0017). Flow cytometric evaluation of the surface antigens expressed by the leukemic blasts for CD34, TdT, HLADR, CD7, CD19 and CD56 found that only CD56 expression was associated with EMI. The association of CD56 expression with lymphadenopathy was statistically significant (P=0.035). Abnormal karyotypes were found in 50.6% of patients with EMI and 49.7% of patients without EMI. Only 11q23 abnormalities were associated with specific sites of EMI; lymphadenopathy (P=0.0111) and gingival hypertrophy (P=0.0016). Our study of adult AML patients demonstrates that EMI at diagnosis is associated with CD56 expression by leukemic blasts, 11q23 karyotypic abnormalities, low complete remission rate and poor overall survival. PMID- 15289012 TI - Dynamics of telomere erosion and its association with genome instability in myelodysplastic syndromes (MDS) and acute myelogenous leukemia arising from MDS: a marker of disease prognosis? AB - Telomere length was evaluated by terminal repeat fragment method (TRF) in 50 patients with myelodysplastic syndromes (MDS) and acute myelogenous leukemia (AML) arising from MDS and in 21 patients with untreated primary AML to ascertain, whether telomere erosion was associated with progression of MDS towards overt leukemia. Heterogeneity of TRF among MDS FAB subgroups (P=0.004) originated from its shortening in increased number of patients during progression of the disease. Chromosomal aberrations were present in 32% MDS patients with more eroded telomeres (P=0.022), nevertheless a difference between mean TRF in the subgroups with normal and abnormal karyotype diminished during progression of MDS. A negative correlation between individual TRF and IPSS value (P=0.039) showed that telomere dynamics might serve as a useful prognostic factor for assessment of an individual MDS patient's risk and for decision of an optimal treatment strategy. PMID- 15289013 TI - Growth and apoptosis of human natural killer cell neoplasms: role of interleukin 2/15 signaling. AB - Interleukin (IL)-15 plays an important role in the survival of human natural killer (NK) cells. We investigated IL-2/15 signaling in NK cell neoplasms from five patients and in five cell lines (NK-92, KHYG-1, SNK-6, HANK1 and MOTN-1) compared to mature peripheral NK cells from 10 healthy subjects. Apoptosis of NK cell lines was prevented by addition of IL-15 in vitro. Blocking IL-2/15Rbeta on IL-2-stimulated NK-92 cells resulted in reduced expression of Bcl-X(L) and phosphorylated Stat5, which paralleled early apoptosis without altering Bcl-2 expression. These data add IL-2/15Rbeta to the list of factors important for the survival of NK cell neoplasms. PMID- 15289014 TI - High frequency of t(12;21)(p13;q22) in children with acute lymphoblastic leukemia and known clinical outcome in southern Brazil. AB - The presence of the t(12;21)(p13;q22) distinguishes a subset of children with acute lymphoblastic leukemia (ALL) that present a favorable prognosis. This is a cryptic translocation difficult to detect through conventional cytogenetics. In this study, bone marrow samples from 30 children with ALL from southern Brazil were evaluated by fluorescence in situ hybridization (FISH) for the t(12;21), using locus specific probes to detect the TEL/AML1 rearrangement. The selection criteria included: age (0-12 years old); FAB classification (L1 or L2), absence of specific clonal chromosomal aberrations; and adequate cellular integrity to perform FISH analysis. A frequency of 40% of the t(12;21) was observed, in addition to extra copies of the AML1 gene in 7.5% of patients. These findings were analyzed in relation to the patient's clinical parameters and compared with other pediatric populations. PMID- 15289015 TI - Methylenetetrahydrofolate reductase genotype does not play a role in adult T-cell leukemia/lymphoma pathogenesis among human T-lymphotrophic virus type 1 carriers. AB - Human T-lymphotropic virus type 1 (HTLV-1) is associated with adult T-cell leukemia/lymphoma (ATL). However, the incidence of ATL is low among HTLV-1 carriers suggesting additional mechanisms are involved in the progression of the disease. A recent study found that polymorphisms in the methylenetetrahydrofolate reductase (MTHFR) gene influence the susceptibility to malignant lymphoma. We have analyzed the MTHFR genotype in 81 HTLV-1 carriers and 87 ATL patients. No statistically significant associations were found between MTHFR genotype and development of ATL. These data suggest that genetic polymorphisms in the MTHFR locus do not play a role in ATL pathogenesis among HTLV-1 carriers. PMID- 15289016 TI - Elevated levels of soluble CD44 are associated with advanced disease and in vitro proliferation of neoplastic lymphocytes in B-cell chronic lymphocytic leukaemia. AB - PURPOSE: Increased expression of the adhesion molecule CD44 has been associated with an unfavourable clinical outcome in lymphomas. We evaluated the prognostic value of soluble CD44 in B-cell chronic lymphocytic leukaemia (B-CLL) and analysed the source and regulation of CD44 secretion in B-CLL clones in vitro. PATIENTS AND METHODS: Levels of soluble CD44 standard (sCD44s) and of the soluble variant isoform CD44v6 (sCD44v6) were analysed by enzyme linked immuno sorbent assay. Highly purified B-CLL cells (98% CD19 + CD3 - cells) were stimulated in vitro by different combinations of thioredoxin (Trx), Staphylococcus aureus Cowan strain 1 (SAC), IL-2, IL-4, IL-10, 12-O-tetradecanoyl-phorbol-13-acetate (TPA) and by anti-CD40 mAbs presented on irradiated CD32L cells. RESULTS: Serum levels of sCD44s and of sCD44v6 are significantly elevated in B-CLL patients (n = 90) in comparison with normal persons (n = 44) (P < 0.001). Elevated levels of sCD44s and sCD44v6 are associated with an advanced disease as reflected by an extended lymph node involvement (P < 0.02), an advanced Binet (P < 0.03) and Rai stage (P < 0.04) and chemotherapy requirement (P < 0.02). High levels of sCD44s are associated with high leukocyte counts (P < 0.04) and increased sCD44v6 is significantly associated with splenomegaly (P < 0.002). In B-CLL sCD44s as well as sCD44v6 is shed from leukaemia cells as shown by in vitro cultures. Stimulation of B-CLL clones results in a proliferation-associated increased secretion of sCD44s (rho = 0.7; P = 0.0001) and of sCD44v6 (rho = 0.5; P = 0.005). B-CLL clones from advanced stage patients are characterised by an increased capacity for proliferation and CD44 production in comparison with early stage patients. CONCLUSIONS: Both sCD44s and sCD44v6 represent a reliable prognostic marker in B-CLL and may be involved in the pathogenesis of B-CLL. PMID- 15289017 TI - An association study of the tumor necrosis factor alpha C-850T polymorphism and childhood acute lymphoblastic leukemia in a population from northern Greece. AB - Excessive production of tumor necrosis factor alpha (TNFalpha) may influence the risk and/or progression of hematologic malignancies and has been associated with febrile episodes at diagnosis. We have examined the putative association of the C 850T polymorphism of the human TNFalpha gene with childhood acute lymphoblastic leukemia (ALL) and with clinical and laboratory findings at diagnosis, in 58 childhood ALL patients from northern Greece. No statistically significant associations have emerged between this polymorphism and either the risk for childhood ALL or presence of fever, anemia, leukocytosis and leukopenia at diagnosis, in this study. PMID- 15289018 TI - Addition of cyclosporin A to the combination of mitoxantrone and etoposide to overcome resistance to chemotherapy in refractory or relapsing acute myeloid leukaemia: a randomised phase II trial from HOVON, the Dutch-Belgian Haemato Oncology Working Group for adults. AB - Cyclosporin A (CsA) inhibits the P-gp pump that can be responsible for failure of cytostatic treatment in acute myeloid leukaemia (AML). Eighty patients with relapsing/refractory AML were randomly assigned to mitoxantrone (M) and etoposide (VP) (MVP) in unmitigated antileukaemic doses with or without CsA, to investigate if toxicity was manageable and if antileukaemic therapy could be improved. CsA did not delay haematological recovery, but fewer CsA patients received post induction therapy because of haematological and non-haematological toxicity. CR rate was 43% for MVP and 53% for CsA; DFS was 9 and 8 months, and OS 8 and 9 months, respectively. Seventeen of 38 CR patients proceeded to stem cell transplantation (SCT). After a median follow-up of 66 months, six patients were still alive. Addition of CsA did not improve treatment outcome, possibly due to inadequate post-induction therapy as a result of increased toxicity. PMID- 15289019 TI - Evolution of FLT3-ITD and D835 activating point mutations in relapsing acute myeloid leukemia and response to salvage therapy. AB - Internal tandem duplications (ITDs) of the juxtamembrane region of the FLT3 tyrosine kinase receptor are the most frequent genetic alterations in acute myeloid leukemia (AML). The presence of this mutation has been recognized as an independent poor prognostic factor. In this study, we compared the FLT3 mutational status between diagnosis and subsequent relapses in 31 patients with AML. At diagnosis, seven patients were identified to contain FLT3-ITD mutations and one patient to harbor the D835 mutation. Five patients remained FLT3-ITD positive throughout the disease course (+/+). Three patients lost the FLT3 gene mutation at first (one FLT3-ITD, one D835 mutation), or second relapse (one FLT3 ITD) (+/-). One additional patient lost a small FLT3-ITD positive clone at relapse and at the same time gained an apparently unrelated FLT3-ITD positive clone. One patient without FLT3 mutation at diagnosis relapsed with an FLT3-ITD mutation (-/+). A shorter median duration of first remission (6 months versus 11.5 months) and a higher relapse rate after salvage therapy (e.g. allogeneic peripheral blood stem cell transplantation) resulted in a lower leukemia-free survival in the FLT3 mutated group (11% versus 31%). The loss of a clone with a mutation in the FLT3 gene at relapse did not improve the prognosis. PMID- 15289021 TI - Overexpression of delta-like (Dlk) in a subset of myelodysplastic syndrome bone marrow trephines. AB - The mRNA expression level of the gene delta-like (Dlk), coding for a signal transducer related to the Delta-Notch family, was measured using real-time PCR methodology in a large series of control biopsies (n = 61) and bone marrow trephines from patients with myelodysplastic syndrome (MDS) (n = 93) and related myeloid disorders (AML, n = 16 and chronic myeloproliferative disease (CMPD), n = 38). It turned out that dlk is strongly overexpressed in a subset of MDS (16%) cases, however with higher frequency in blast rich MDS cases. Therefore, the quantitative detection of dlk mRNA can support the morphological diagnosis of MDS in selected cases. PMID- 15289020 TI - Pathogenesis of jumping translocations: a molecular cytogenetics study. AB - BACKGROUND/AIMS: Jumping translocations are rare cytogenetic aberrations in haematological malignancies, the pathogenesis of which remains to be fully characterised. We investigated the mechanism of formation of jumping translocations in a case of adult common acute lymphoblastic leukaemia (ALL) positive for the Ph translocation. METHODS: Interphase and metaphase fluorescence in situ hybridisation (FISH) was performed using several probe systems. Results were correlated with findings on conventional cytogenetics. Granulocytes, T-cells and leukaemic B-cells in peripheral blood were sorted by immunomagnetic method and the terminal restriction fragment (TRF) length of these cellular populations was determined by Southern blot analysis. RESULTS: Duplicated BCR-ABL fusion signals were found on a dic(14;22)der(22)t(9;22) chromosome. Clonal jumping translocations, existing as evolutionary changes, involved the donor chromosomal segment distal to 1q12 jumping onto the telomere ends of 11q, 15p, 19p and 20p. Telomere length was decreased in the neoplastic B-cell population and contributed to the formation of the dicentric chromosome that showed absence of telomere repeats at fusion ends. Subsequent pericentromeric heterochromatin decondensation of chromosome 1q occurred, and this donor segment was randomly fused to the shortened telomere ends of non-homologous chromosomes. CONCLUSIONS: Both telomere shortening and pericentromeric heterochromatin decondensation contribute to the formation of jumping translocations, which is most probably a multi-stage process. PMID- 15289022 TI - Enhanced antitumor immunity by murine cytokine activated T lymphocytes after cocultured with bone marrow derived dendritic cells pulsed with whole tumor lysates. AB - Human cytokine induced killer cells (CIK) have shown potent non-major histocompatibility (MHC)-restricted antitumor activity in vitro in recent years. Using a similar protocol, we generated murine cytokine activated T lymphocytes (CAT) from splenocytes with the sequential addition of IFN-gamma, IL-1beta, anti CD3 and IL-2. The NK depleted CATs did not possess significant antitumor activity both in vitro and in vivo. However, after cocultured with dendritic cells (DC) pulsed with whole tumor lysates, significant specific antitumor activity was observed both in vitro and in vivo. Our data demonstrated that the CATs cocultured with dendritic cells pulsed with whole tumor lysates have potent specific antitumor effects and might be useful in the treatment of malignancies. PMID- 15289023 TI - HI44a, an anti-CD44 monoclonal antibody, induces differentiation and apoptosis of human acute myeloid leukemia cells. AB - CD44 is a cell surface antigen that expresses on leukemia blasts from most acute myeloid leukemia (AML) patients. It has been reported that ligation of CD44 with some specific anti-CD44 monoclonal antibodies can reverse the differentiation blockage of leukemia cell lines. In this study, the differentiation and apoptosis inducing effects of HI44a, another anti-CD44 monoclonal antibody (IgG2a), were investigated on leukemia cells obtained from 31 patients with AML-M2, AML-M3, AML M4 or AML-M5. When the AML cells were treated with HI44a, the percentage of nitroblue tetrazolium (NBT)+ cells was significantly increased. The expression of CD11b, CD14 and CD15 on treated AML cells was also increased compared to control AML cells. In addition, HI44a was found to induce apoptosis of leukemia cells, as evidenced by an annexin-V assay. The mean percentage of apoptotic cells in HI44a treated AML cells was significantly increased compared to that in control AML cells. Moreover, the level of c-myc transcript expression on AML cells was found to be obviously decreased in all detected patients. These results indicate that HI44a effectively induces both differentiation and apoptosis of AML cells and suggest that this activity of the anti-CD44 antibody may be associated with its inhibitory effect on c-myc transcript expression. PMID- 15289024 TI - Bone and bone marrow interactions: hematological activity of osteoblastic growth peptide (OGP)-derived carboxy-terminal pentapeptide III. Action on human megakaryocytopoiesis: focus on essential thrombocythemia. AB - The increase of megakaryocytes and platelets that characterizes essential thrombocythemia (ET) appears to be secondary to a deregulation of megakaryocytopoiesis. The carboxy-terminal fragment of osteogenic growth peptide (OGP10-14) promotes bone formation and hemopoiesis, while it inhibits megakaryocytopoiesis. In this paper we show that treatment with synthetic OGP10 14 (sOGP10-14) induces a significant reduction of mid and large colony-forming unit-megakaryocytes (CFU-Mk) in ET patients as well as in controls, and is associated with a significant inhibition of thrombopoietin (TPO)-primed MO-7e megakaryoblastic cells proliferation. These actions appear to be related to sOGP10-14 modulation of TGF-beta(1) synthesis and/or secretion, although a direct effect on TGF-beta receptor expression cannot be excluded. PMID- 15289025 TI - Acute leukemia of donor origin arising after stem cell transplantation for acute promyelocytic leukemia. AB - We report a patient with PML/RARalpha-positive acute promyelocytic leukemia (APL) who developed PML/RARalpha-negative acute myeloid leukemia 37 months after allogeneic bone marrow (BMT) transplant for molecular relapse. Features of myelodysplasia were noted 11 months earlier, chimerism testing by analysis of short tandem repeats was consistent with development of myelodysplasia and acute leukemia within cells of donor origin. To our knowledge, this is the first report of donor cell leukemia following BMT for APL. We hypothesize that replicative stress may lead to the development of some cases of donor cell acute leukemia. PMID- 15289026 TI - Familial association of leukemia with colorectal cancer. PMID- 15289027 TI - Monitoring of plasma imatinib concentration for the effective treatment of CML patients. PMID- 15289028 TI - Early neurosensory visual development of the fetus and newborn. AB - Neurosensory development of the visual system has its origins long before birth. The genetic processes of basic structure formation are followed by endogenous retinal ganglion cell activation in the form of spontaneous synchronous waves of stimulation. These waves of stimulation are required to establish the topographic relationship among retina, lateral geniculate nucleus, and visual cortex. This process prepares the visual system for visual experience. Visual experience ultimately stimulates creation of columns of neurons in the visual cortex, which are needed to see and interpret patterns, lines, movement, and color. Spontaneous synchronous retinal waves occur in preterm infants in the neonatal intensive care unit and must be protected, as they are critical for visual development. PMID- 15289029 TI - Emergence and influences of circadian rhythmicity in infants. AB - Recent evidence shows that the circadian system of primate infants is responsive to light at very premature stages and that low intensity lighting can regulate the developing clock. After birth, there is progressive maturation of the circadian system outputs, with pronounced rhythms in sleep-wake and hormone secretion generally developing after 2 months of age. Showing the importance of photic regulation of circadian phase in infants, exposure of premature infants to low-intensity cycled lighting results in the early establishment of rest-activity patterns that are in phase with the 24-hour light-dark cycle. With the continued elucidation of circadian system development and influences on human physiology and illness, it is anticipated that consideration of circadian biology will become an increasingly important component of neonatal care. PMID- 15289030 TI - Lighting for caregivers in the neonatal intensive care unit. AB - The primary aim of this article is to define good lighting for caregivers(both medical staff and families) working in the hospital neonatal intensive care unit (NICU) and other areas associated with the critical care environment. Defining good lighting requires understanding that lighting is not only important for the infants in the NICU but that it also plays significant roles for adults in the NICU. First, lighting supports visual processes (eg, acuity, color vision, visual performance). Second, lighting affects circadian regulation (eg, alertness, sleeping, hormone production). Finally,lighting communicates a message to professional staff as well as parents and visitors about the level of care and sophistication provided by the hospital. By thoughtfully addressing all three roles, the lighted environment in the NICU can support the productivity and well being of the professional staff, the health and safety of patients, as well as the profitability of the NICU. A secondary aim of this chapter is to provide practical guidance to health care professionals on how to articulate good lighting objectives to application engineers and designers responsible for the lighting in the NICU. PMID- 15289031 TI - Effects of the neonatal intensive care unit on auditory attention and distraction. AB - A theory is proposed that attention to acoustic signals is important for normal development and varies with background masking sounds. Specifically, the theory states that distractibility increases with decreasing predictability of the acoustic environment and with decreasing age. Literature from premature neonates, normal infants, preadolescent children, children with attention deficit disorders, and adults is reviewed. One conclusion is that an environment perceived by adults as predictable may be distracting for preterm infants. One recommendation for future research is to include measures of background acoustic predictability as independent variables or covariates in developmental studies. PMID- 15289032 TI - Olfaction in the fetal and premature infant: functional status and clinical implications. AB - This article considers olfaction as a functioning source of information for the fetus and the neonate, born on term or prematurely. It aims to present how odors are involved in the sensory continuity between the prenatal and postnatal environments and how they influence the earliest adaptive responses of newborns in the realms of self-regulation, emotional balance, feeding, and social interactions.Finally, it evaluates odors as sensory means to ameliorate the physiologic and behavioral responses of preterm infants to the adverse impacts of separation from mother, nonoral feeding, or iatrogenic distress. PMID- 15289033 TI - Early relationship environments: physiology of skin-to-skin contact for parents and their preterm infants. AB - Skin-to-skin care involves the mother placing her diaper-clad infant upright between her breasts in direct skin contact. The practice has evolved worldwide to be an intervention strategy in neonatal intensive care units for premature infants and their mothers. Few adverse outcomes have been noted in thermoregulation, cardiovascular changes, or behavioral organization. Findings have been positively related to better infant physiologic and neurobehavioral outcomes, maternal breastfeeding success, and positive attachment relationships. The early, intimate, and physiologically stabilizing benefits of skin-to-skin care provide for a new conceptualization of the optimal environment for preterm infants in intensive care. PMID- 15289034 TI - Evidence-based design for infants and staff in the neonatal intensive care unit. AB - Conscientious architects are becoming increasingly aware of the impact of design decisions on the sensory environment of the neonatal intensive care unit (NICU). This article addresses the relevance of theories of environmental psychology to NICU design. Design research on infants and staff in NICUs is summarized, and future research directions are identified. PMID- 15289035 TI - Planning a developmentally appropriate neonatal intensive care unit. AB - Neonatal intensive care units (NICUs) are now being redesigned in the wake of growing evidence that the physical environment of the NICU has a profound impact on all who live and work there. These new units reflect the universal trend of bringing families directly into the center of the medical care team. More than ever,such projects are bound to change how staff care for babies, how families interact with babies, how staff and families interact with one another, and how staff interact among themselves. When a NICU decides that a more developmentally appropriate environment should be initiated, the question of funding and obtaining other resources inevitably arises. This chapter identifies the essential components that should be included when planning to seek investments in NICU facilities. PMID- 15289036 TI - Lighting design in the neonatal intensive care unit: practical applications of scientific principles. AB - Meeting the varied lighting needs of infants, caregivers, and families has become more complex as our understanding of visual development and perception and the effect of light on circadian rhythms advances. Optimal lighting strategies are discussed for new unit construction, as well as modifications to consider for existing units. In either case, the key concept is that lighting should be provided for the individual needs of each person, rather than the full-room lighting schemes previously used. Ideas gleaned from nonhospital settings, re introduction of natural light into the neonatal intensive care unit, and new devices such as light-emitting diodes will dramatically change the lighting and visual environment of future neonatal intensive care units. PMID- 15289037 TI - Planning the acoustic environment of a neonatal intensive care unit. AB - This article addresses general principles of designing a quiet neonatal intensive care unit (NICU) and describes basic aspects of room acoustics as these apply to the NICU. Recommended acoustical criteria for walls, background noise, vibration, and reverberation are included as appendices. Crowding in open, multiple-bed NICUs is the major factor in designs that inevitably produce noisy nurseries with limited space for parents. Quiet infant spaces with appropriate sound sources rely on isolation of the infant from facility and operational noise sources (eg, adult work spaces, supply delivery, and travel paths) and extended contact with family members.However, crowding has been an important influence on the clinical practice and social context of neonatology. It allows clinicians to rely on wide visual and auditory access to many patients for monitoring their well-being. It also allows immediate social contact with other adults, both staff and families. Giving up this wide access and relying on other forms of communication in order to provide for increased quiet and privacy for staff, infants, and parents is a challenge for some design teams. Studies of the effects of various nursery designs on infants, parents, clinicians, and the delivery of services are proposed as a means of advancing the field of design. PMID- 15289038 TI - Designing the neonatal intensive care unit for optimal family involvement. AB - The design of a new neonatal intensive care unit provides an opportunity to ensure that the new facility best meets the needs of the infants and families whom the unit serves. In design planning,administrators, staff, family members, and the architect must work together in a self-education process that entails examining current design standards, exploring exemplary facilities at other institutions,defining the priorities and needs of infants, families, and staff, and deciding how to respond to them. The involvement of family members in this important work can help ensure that the facility is responsive to families and supports the family as the primary caregiver and decision maker for the infant. Such an environment will lead to improved health and developmental outcomes for infants and greater family and staff satisfaction. PMID- 15289039 TI - Mothers' arms--the past and future locus of neonatal care? AB - With the advent of neonatal intensive care, medical professionals inserted themselves between the baby and its family. Even for healthy newborn, the mother could only get to her infant with permission from the medical staff and then only for limited periods and in a very restricted manner. Family-centered care restored the family's right to full access to their baby, but as generally practiced(and certainly as NICUs are currently designed) medical professionals still view the infant as a solitary individual who sleeps most of the time in a bed. Future NICU design should recognize that the baby must spend most of its time in its mother's arms to get the full benefit of her sensory environment as experienced throughout our evolution. NICUs must therefore be planned to facilitate this extended proximity as much as possible (ie, not just providing for parents at the bedside with the bed as the locus of care, but transforming the preferred and predominant locus of care from the bed to the parents' arms, with the design changes inherent to that concept). Designing our units to facilitate this interaction will not assure that it will always occur, but it will guarantee that we have not created permanent structural features that interfere with this crucial relationship. PMID- 15289041 TI - Sperm cryopreservation of a live-bearing fish, the platyfish Xiphophorus couchianus. AB - The objectives of this study were to evaluate the effects of cryoprotectant, osmotic pressure, cooling rate, equilibration time, and sperm-to-extender ratio, as well as somatic relationships of body length, body weight, and testis weight to sperm density in the platyfish Xiphophorus couchianus. Sperm motility and survival duration after thawing were significantly different between cryopreservation with dimethyl sulfoxide (DMSO) and glycerol, with the highest motility at 10 min after thawing obtained with 14% glycerol. With subsequent use of 14% glycerol as cryoprotectant, the highest motility after thawing was observed with Hanks' balanced salt solution (HBSS) across a range of 240-300 mOsm/kg. Samples cooled from 5 to -80 degrees C at 25 degrees C/min yielded the highest post-thaw motility, although no significant difference was found for cooling rates across the range of 20-30 degrees C/min. In addition, the highest motility after thawing was found in samples equilibrated from 10 to 30 min with 14% glycerol and cooled at 25 degrees C/min. The post-thaw motility declined rapidly with use of 10% glycerol and cooling at 5 degrees C/min across the equilibration range of 10 min to 2h. Sperm motility with a dilution ratio of sperm to extender of 1:10 was not different at 10 min after thawing with those samples at greater dilutions, but declined significantly from Day 1 after thawing and showed lower survival duration when stored at 4 degrees C. However, the additional dilution of sperm solutions with HBSS (300 mOsm/kg) immediately after thawing significantly slowed the decline of motility and prolonged the duration of survival. Based on the above findings, the highest average sperm motility (78+/-3 %) at 10 min after thawing was obtained when sperm were suspended in HBSS at 300 mOsm/kg with 14% glycerol as cryoprotectant, diluted at a ratio of sperm to HBSS-glycerol of 1:20, equilibrated for 10 min, cooled at 25 degrees C/min from 5 to -80 degrees C before plunging into liquid nitrogen, and thawed at 40 degrees C in a water bath for 7 s. If diluted within 5 h after thawing, sperm frozen by the above protocol retained continuous motility for 15 days when stored at 4 degrees C. PMID- 15289042 TI - Transcervical artificial insemination in sheep: effects of a new transcervical artificial insemination instrument and traversing the cervix on pregnancy and lambing rates. AB - Cervical anatomy limits the use of transcervical intrauterine artificial insemination (TC AI) in sheep. We have developed an instrument to cope atraumatically with the cervix; although this instrument has not affected fertilization rate or pregnancy rate through Day 3, the effects on sperm transport and pregnancy after Day 3 are not known. The objective of the present study was to determine whether our TC AI instrument affected sperm transport, pregnancy rates, or lambing rate. In Experiment 1, ewes were assigned to two treatments: TC AI using the new TC AI instrument (n=10) or AI via laparotomy using a laparoscopic AI instrument (n=10). Twenty hours after artificial insemination, the uterine horns and oviducts were recovered and flushed to collect spermatozoa. Sperm transport did not differ (P>0.05) between the two treatments. In Experiment 2, ewes were assigned to three treatments: TC AI using the new TC AI instrument+sham intrauterine AI via laparotomy (n=29); sham TC AI+intrauterine AI via laparotomy using a laparoscopic AI instrument (n=29); and sham TC AI+intrauterine AI via laparotomy using the new TC AI instrument (n=30). On Day 14 after AI, uteri were collected and flushed to recover blastocysts. Transcervical deposition of semen reduced (P<0.05) Day 14 pregnancy rate (17.2% versus 61%), but intrauterine deposition of semen using the TC AI instrument via midventral laparotomy increased (P<0.05) Day 14 pregnancy rate (76.6% versus 44.8%). In Experiment 3, ewes were assigned to two treatments: sham cervical manipulation (n=40) or cervical manipulation to mimic TC AI (n=40). Immediately after treatment, each ewe was mated with a ram and watched until the ram mounted and ejaculated into the ewe. Treatment did not affect Day 30 or 50 pregnancy rate (67.5 and 66.2%, respectively), determined ultrasonically, or lambing rate (62.5%). The differences between Days 30 and 50 pregnancy rates and lambing rate were not significant. In Experiment 4, ewes were assigned to two treatments: TC AI (n=99) or laparoscopic AI (n=99). Transcervical AI reduced (P<0.01) Day 30 (TC AI versus laparoscopic AI; 5.0% versus 46.0%) and Day 50 pregnancy rates (4.0% versus 41.0%), determined ultrasonically, and lambing rate (4.0% versus 41.0%). Although the TC AI procedure significantly reduced pregnancy and lambing rates, large numbers of spermatozoa deposited at natural insemination seemed to compensate. Because our TC AI procedure has all but eliminated any visual evidence of trauma, and because the procedure does not seem to affect sperm transport or embryonal survival until Day 3, we speculate that cervical manipulation associated with TC AI may activate pathways that interrupt pregnancy between Days 3 and 14. PMID- 15289043 TI - Changes in ovarian, follicular, and oocyte morphology immediately after the onset of puberty are not accompanied by an increase in oocyte developmental competence in the pig. AB - This study was conducted to determine whether ovarian morphology and developmental competence of in vitro-matured (IVM) oocytes is immediately affected by the onset of puberty in the pig. Ovaries of peri-pubertal pigs were sorted into two groups according to the presence or absence of corpora lutea presence (CL and NCL, respectively. Ovary dimensions, follicle diameter and number, and oocyte diameter (with and without zona pellucidae) were determined. The developmental competence of in vitro-matured oocytes from these two groups was evaluated following parthenogenetic activation and culture in vitro. CL ovaries were significantly (P<0.01) larger than NCL ovaries (width: 22.3+/-0.9 mm versus 15.9+/-0.4 mm, length: 33.2+/-1 mm versus 24.1+/-0.4 mm). Although CL ovaries had fewer antral follicles in total compared with NCL ovaries (21.1+/-1.8 mm versus 46.8+/-2.2 mm), they had a similar number of follicles 3-8mm in diameter. The mean diameter of follicles that were aspirated was greater for CL ovaries than for NCL ovaries (4.5+/-0.1 mm versus 3.3+/-0.02 mm). Oocytes from CL ovaries were greater in diameter compared with those from NCL ovaries (zona retained: 159+/-1.3 microm versus 146.1+/-1.5 microm, zona free: 124.7+/-1.8 microm versus 113.1+/-1.6 microm). No differences were found between oocytes from CL and NCL ovaries for rates of meiotic maturation (91.6+/-3.2% versus 92.4+/ 3.2%), cleavage (88.4+/-11% versus 90.7+/-2.6%) and blastocyst formation (21.0+/ 3.7% versus 23.7+/-5.7%). Therefore, the onset of puberty coincides with immediate changes in ovarian morphology, increased ovary size, follicle and oocyte diameter, but not with improved oocyte developmental competence. This suggests that the higher developmental competence usually observed in adult oocytes is acquired gradually and requires exposure to multiple estrus cycles. PMID- 15289044 TI - Prenatal toxicity of cyanide in goats--a model for teratological studies in ruminants. AB - Although exposure to cyanogenic plants or cyanide during pregnancy has adverse effects, no teratological study with cyanide has been conducted in goats or any other ruminant. The objective of the present study was to evaluate the effects of the maternal exposure to potassium cyanide (KCN) during pregnancy on both dams and offspring and furthermore, to develop a model for prenatal toxicological studies in ruminants. Twenty-six pregnant goats were allocated into four groups and given 0, 1.0, 2.0, or 3.0mg KCN/kg body weight per day orally (administered via twice-daily gavage) from Day 24 of pregnancy to term. However, one control dam and another from the 3.0mg KCN/kg per day group were sacrificed on Day 120. At birth, the kids were examined carefully for gross abnormalities. Three months after birth, the male kids and one dam from each group were sacrificed for histopathological study. Although clinical signs of poisoning were observed in dams, cyanide treatment did not alter the length of gestation or the number of live kids. Two prognata kids were born in the 3.0mg KCN/kg group, and one dam from the same group aborted two fetuses. There were histological lesions only in the KCN-treated dam (and its fetuses) sacrificed on Day 120; these consisted of an increased number of resorption vacuoles of thyroid follicular colloid, and status spongiosis of nervous white matter. This study proposes a new animal model for teratogenic trials that could be important to evaluate the effects of chemicals throughout pregnancy in goats and potentially other ruminants. PMID- 15289045 TI - Distribution of spermatozoa in the female reproductive tract of the domestic cat in relation to ovulation induced by natural mating. AB - The purposes of this study were to demonstrate the localization of spermatozoa in the reproductive tract of female domestic cats before (30 min and 3 h after mating) and after ovulation (48 and 96 h after mating), and to evaluate the efficiency of two techniques for studying sperm distribution. Estrus was induced in twenty-four female cats using 100 IU eCG and the females were divided into four groups with six females per group. The same male cat was used for mating with all the females. One group of six females was mated once; the others were mated four times in 1 h. Ovariohysterectomy was performed at 30 min, 3 h, 48 h, and 96 h after mating and the excised reproductive tracts were divided into seven segments on each side: infundibulum, ampulla, isthmus, uterotubal junction (UTJ), cranial and caudal uterine horn, and uterine body. The vagina and the lumina of the segments from one side were flushed with 0.5 ml PBS. The flushed and the non flushed segments from the contralateral side were then fixed in 3% neutral buffered formalin and processed for routine histology. The numbers of spermatozoa in the flushings and in 40 histological sections from each segment were counted. Before ovulation, the majority of spermatozoa was detected in the vagina and the uterine segments, whereas after ovulation, significantly higher numbers of spermatozoa were present in the uterine tubal segments. The decreasing gradient in sperm numbers at 30 min and 3 h after mating between the vagina, the uterine segments, including the UTJ, and the uterine tubal segments indicated that the cervix and the UTJ served as barriers for sperm transport in the cat. The UTJ and the uterine crypts acted as sperm reservoirs before ovulation whereas the isthmus was a sperm reservoir around the time of ovulation. There was no difference in sperm numbers in the tissue sections between flushed and non-flushed segments, implying that the flushing technique only recovered some intraluminal spermatozoa while most of the spermatozoa remained in the epithelial crypts. This was further supported by the finding that significantly higher numbers of spermatozoa were recovered in the flushings at 30 min and 3 h after mating, when more spermatozoa were free in the lumina, than at 48 and 96 h after mating, when the majority of the spermatozoa were entrapped in the uterine epithelial crypts. PMID- 15289046 TI - Follicular dynamics and concentrations of steroids and gonadotropins in lactating cows and nulliparous heifers. AB - Differences in follicular development and circulating hormone concentrations, between lactating cows and nulliparous heifers, that may relate to differences in fertility between the groups, were examined. Multiparous, cyclic, lactating Holstein cows (n=19) and cyclic heifers (n=20) were examined in the winter, during one estrous cycle. The examinations included ultrasound monitoring and daily blood sampling. Distributions of two-wave and three-wave cycles were similar in the two groups: 79 and 21% in cows, 70 and 30% in heifers, respectively. Cycle lengths were shorter by 2.6 days in heifers than in cows, and in two-wave than in three-wave cycles. The ovulatory follicle was smaller in heifers than in cows (13.0+/-0.3 mm versus 16.5+/-0.05 mm). The greater numbers of large follicles in cows than in heifers corresponded well to the higher concentrations of FSH in cows. The duration of dominance of the ovulatory follicle tended to be longer in cows than in heifers. Estradiol concentrations around estrus and the preovulatory LH surge were higher in heifers than in cows (20 versus 9 ng/ml). Progesterone concentrations were higher in heifers than in cows from Day 3 to Day 16 of the cycle. Circulating progesterone did not differ between two-wave and three-wave cycles. The results revealed differences in ovarian follicular dynamics, and in plasma concentrations of steroids and gonadotropins; these may account for the differences in fertility between nulliparous heifers and multiparous lactating cows. PMID- 15289047 TI - Evaluation of alternative cryoprotectants for preserving stallion spermatozoa. AB - Although use of cryopreserved stallion spermatozoa is currently accepted by many breed registries, utilization of this technique remains limited due to poor fertility for some stallions. One reason for these results is osmotic stress that spermatozoa experiences when the cryoprotectant (glycerol) is added to the cells prior to freezing and removal from the cells after thawing. In an effort to minimize osmotic damage, alternative cryoprotectants, having lower molecular weights and greater membrane permeability than glycerol, were evaluated to determine their effectiveness for cryopreserving stallion spermatozoa. In the first experiment, equal molar concentrations of several amides were compared to determine if they could preserve the motility of sperm as well as glycerol. At 0.55 M concentration, addition of glycerol to a skim milk-egg yolk (SMEY) diluent resulted in higher percentages of motile sperm (61%) than methyl formamide (40%) or dimethyl formamide (38%, P<0.05), while formamide, acetamide, and methyl acetamide resulted in recovery of less than 20% motile cells (P<0.05). When methyl formamide or dimethyl formamide were increased to 0.6 or 0.9 M they resulted in percentages of motile cells (48-54%) similar to that achieved with glycerol (52%). Similarly, 0.9 M ethylene glycol also resulted in similar percentages of motile cells (43%). Replacing the glucose and fructose in the SMEY diluent with either raffinose or trehalose did not result in higher percentages of motile sperm (65 and 66%, respectively) than the control SMEY (63%). Similarly, addition of methyl cellulose also did not increase the percentages of motile spermatozoa in the samples, after cryopreservation (P>0.05). In conclusion, both methyl formamide and dimethyl formamide protected stallion spermatozoa from cryodamage as effectively as glycerol. Since these compounds permeate the plasma membrane more effectively than glycerol, they should cause less osmotic damage to stallion spermatozoa than glycerol. Therefore, these compounds may prove very effective in the cryopreservation of stallion spermatozoa, and may be particularly useful for spermatozoa from stallions that produce spermatozoa that have poor post-thaw characteristics when glycerol is used as the cryoprotectant. PMID- 15289048 TI - Effects of granulosa coculture on in-vitro oocyte meiotic maturation within a putatively less competent murine model. AB - A less competent murine in vitro maturation (IVM) model was achieved by shortening the standard duration of in vivo PMSG stimulation from 48 to 24 h and selecting only naked/partially naked GV oocytes from a mixture of large and small follicles. Porcine granulosa coculture enhanced meiotic maturation within such a less competent model (37.3% versus 23.1%, P<0.05), while no significant enhancement was observed with macaque and murine granulosa coculture. Culture of porcine granulosa on extracellular matrix (ECM) gel resulted in a more differentiated morphology, but did not significantly further enhance the beneficial effects it already had on meiotic maturation. Increased concentrations of serum as well as the supplementation of gonadotrophins and follicular fluid within the culture milieu did not enhance IVM under both cell-free and coculture conditions. Porcine granulosa-conditioned medium also enhanced meiotic maturation (36.5% versus 26.7%, P<0.05), which was not diminished upon freeze-thawing (35.8% versus 22.6%, P<0.05). Enhancement of meiotic maturation by porcine granulosa coculture did not however translate to significant improvements in developmental competence, as assessed by in vitro fertilization (IVF) and embryo culture to the blastocyst stage, followed by total cell counts. ECM gel had a detrimental effect on fertilization and developmental competence, even though it had no detrimental effect on meiotic maturation itself. PMID- 15289049 TI - Effect of prematuration, meiosis activating sterol and enriched maturation medium on the nuclear maturation and competence to development of calf oocytes. AB - New strategies were proposed to improve the developmental competence of calf oocytes through in vitro technologies. Cumulus-oocyte complexes were first prematured for 24 h in the presence of meiosis inhibitors. Both Roscovitine alone (50 microM) or in combination with Butyrolactone-I (12.5 microM Rosco+6.25 microM BL-I) prevented the progression of meiosis. Their effect on nuclear maturation was reversible after a further 17 or 24 h maturation step. However, a dramatic decrease in embryo development was observed after fertilization (abattoir oocytes: 4-9% blastocyst rate versus 14-17% for control embryos). Similar results were obtained with oocytes collected by Ovum Pick Up from living donors. No pregnancy was obtained after single transfer of two blastocysts obtained from prematured oocytes (0/2 versus 4/12 for control embryos). Adding low concentrations (1, 3 or 10 microM) of follicular fluid-meiosis activating sterol (FF-MAS) during the maturation step had a beneficial effect on nuclear maturation (73-86% metaphase II versus 58% for control oocytes). However, subsequent embryo development was not improved. Enriching the maturation medium, namely with hormones, growth factors and precursors of glutathione, induced a sixfold increase in glutathione in the oocyte and had a beneficial effect on embryo development (38% increase in blastocyst rate). In conclusion, in opposition to the results reported with adult oocytes, prematuring calf oocytes had a negative impact on their developmental potential. Although FF-MAS improved nuclear maturation, its addition in the maturation medium did not increase embryo development. However, enriching the maturation medium had a positive effect on embryo development, indicating that cytoplasmic maturation was improved. PMID- 15289050 TI - A comparison of diagnosis of pregnancy in the goat via transrectal ultrasound scanning, progesterone, and pregnancy-associated glycoprotein assays. AB - Real-time ultrasound scanning (US) via the transrectal route, progesterone (P4) assay, and pregnancy-associated glycoprotein (PAG) detection can be used to diagnose pregnancy at around 3 weeks after breeding. Although several studies have been carried out to evaluate each of these different methods individually, it is difficult to establish adequate comparisons due to differences, such as the breed of goat, age, and farming conditions, among others. The aim of the present paper is to compare the accuracy of diagnosis of pregnancy using transrectal US, P4 assay and PAG detection at the same time and on the same animals. Canary dairy goats (n=143) were synchronized with an 11-day fluorogestone acetate (FGA) intravaginal sponge followed by PGF2alpha and eCG 2 days before the FGA withdrawal. Blood samples were collected on Days 20, 22, 24, and 26 after mating to determine P4 and PAG concentrations. Transrectal US examinations were performed at the same time. There were 79 pregnant goats and another 64 non pregnant. The US via the transrectal method and the determination of PAG concentrations provide very accurate pregnancy diagnosis at 24-26 days after breeding; on the contrary, P4 assay on plasma samples performed on Day 22 after breeding was accurate, in this case, in detecting pregnant animals but did not always detect the non-pregnant does. PMID- 15289051 TI - GnRH immunocontraception of male cats. AB - The development of nonsurgical contraceptives for cats may facilitate population control of the species. The purpose of this study was to investigate the utility of GnRH for immunocontraception of male cats. Male cats (n=12) were divided into groups of three and were immunized once with 0 (sham), 50, 200, or 400 microg synthetic GnRH coupled to keyhole limpet hemocyanin and combined with a mycobacterial adjuvant to enhance immunogenicity. GnRH antibody titer, serum testosterone concentration, and scrotal size were determined monthly. At 6 months, semen was collected by electroejaculation and testes were examined histologically. GnRH antibodies were detected in all cats receiving GnRH vaccine by 1 month post-treatment and persisted throughout the study. No dose effect of GnRH was observed; titers were not different among cats treated with 50, 200, or 400 microg GnRH (P=0.5). Six of nine treated cats were classified as responders based on high GnRH antibody titers (>32,000). By 3 months post-treatment, responder cats had undetectable testosterone concentrations and testicular atrophy. Nonresponder cats had GnRH titers of 4000-32,000 and testosterone concentrations intermediate between responder and sham-treated cats. At 6 months, total sperm counts were similar for sham-treated cats (3.1+/-1.8 x 10(6) sperm) and nonresponder cats (3.4+/-1.6 x 10(6) sperm; P=0.7). Only one of the six responder cats produced sperm, none of which were motile. Combined testicular weights of responder cats (1.3+/-0.1 g) were lower than sham-treated controls (5.3+/-1.3 g; P=0.02) and nonresponder cats (2.9+/-0.3 g; P=0.02). Histologic evaluation of the testes revealed that in responder cats, the interstitial cells that were present were pale and shrunken compared to the plump, polyhedral eosinophilic cells in sham-treated cats. GnRH responder cats had marked tubular atrophy with vacuolated Sertoli cells and a paucity of germ cells. Single-dose GnRH treatment resulted in testosterone concentrations and semen quality consistent with immunocastration in a majority of cats treated. PMID- 15289053 TI - In vitro development following one-step dilution of OPS-vitrified porcine blastocysts. AB - The objective of this experiment was to compare the in vitro survival and hatching rates of OPS-vitrified porcine blastocysts obtained after conventional (three-step dilution) or direct (one-step dilution) warming procedures. Expanded blastocysts were collected by laparotomy from weaned crossbred sows (n=7) on Day 6 of the cycle (D0: onset of estrus). Vitrification was performed as described by Berthelot et al. [Cryobiology 41 (2000) 116] using 17% (v/v) ethylene glycol and 17% (v/v) dimethyl-sulfoxide in the second vitrification medium. Conventional warming was carried out by plunging straws containing embryos in 800 microl of TCM199 Hepes containing 20% new born calf serum (TCM-NBCS) and 0.13 M sucrose for 1 min. Embryos were then transferred to another well with the same medium for 5 min, washed in TCM-NBCS with 0.075 M sucrose for 5 min and transferred to TCM NBCS for 5 min. In one-step dilution, embryos were placed in 400 microl TCM-NBCS containing 0.13 M sucrose. To evaluate in vitro development, embryos warmed by conventional (n=59) or direct (n=58) procedures were cultured for 96 h. Non vitrified embryos were used as controls (n=20). No significant (P>0.05) differences were observed in the in vitro development of vitrified and non vitrified embryos. The survival and hatching rates obtained by three-step dilution (84.8 and 71.2%, respectively) and one-step dilution (86.2 and 74.1%, respectively) procedures were not different (P>0.05). The average diameter of expanded blastocysts from each donor was significantly different (P<0.001) among embryo donors. The embryo diameter or the interactions among the factors evaluated did not affect (P>0.05) the embryo survival and hatching of the vitrified/warmed blastocysts. However, the donor of embryos had a significant effect (P<0.001) on these parameters, confirming previous experiments. This experiment shows that porcine embryo vitrification and one-step dilution are promising procedures to be used under field conditions. However, the good results obtained in vitro must be confirmed also by in vivo experiments. PMID- 15289052 TI - Metabolic changes in follicular fluid of the dominant follicle in high-yielding dairy cows early post partum. AB - Characteristics of the intrafollicular environment to which the preovulatory oocyte is exposed may be one of the major factors determining subsequent fertility. The aim of our study was to examine to what extent metabolic changes that occur in early post partum high-yielding dairy cows are reflected in the follicular fluid (FF) of the dominant follicle (>8 mm). Nine blood samples were taken per cow from nine high-yielding dairy cows between 7 days before and 46 days after parturition. From Day 14 post partum on and together with blood sampling, FF samples of the largest follicle were collected from the same cows by means of transvaginal follicle aspiration. Serum and FF samples were analyzed using commercial clinical and photometric chemistry assays for glucose, beta hydroxybutyrate (beta-OHB), urea, total protein (TP), triglycerides (TG), non esterified fatty acids (NEFA) and total cholesterol (TC). All cows lost body condition during the experimental period (0.94+/-0.09 points) illustrating a negative energy balance during the experimental period. In FF, glucose concentrations were significantly higher and the TP, TG, NEFA and TC concentrations were significantly lower than in serum (P<0.05). The concentrations of glucose, beta-OHB, urea and TC in serum and in FF changed significantly over time (P<0.05). Throughout the study, changes of all metabolites in serum were reflected by similar changes in FF. Especially for glucose, beta-OHB and urea the correlations were remarkably high. The results from the present study confirm that the typical metabolic adaptations which can be found in serum of high-yielding dairy cows shortly post partum, are reflected in follicular fluid and, therefore, may affect the quality of both the oocyte and the granulosa cells. PMID- 15289054 TI - Methanol as a cryoprotectant for equine embryos. AB - Equine embryos (n=43) were recovered nonsurgically 7-8 days after ovulation and randomly assigned to be cryopreserved in one of two cryoprotectants: 48% (15M) methanol (n=22) or 10% (136 M) glycerol (n=21). Embryos (300-1000 microm) were measured at five intervals after exposure to glycerol (0, 2, 5, 10 and 15 min) or methanol (0, 15, 35, 75 and 10 min) to determine changes (%) in diameter over time (+/-S.D.). Embryos were loaded into 0.25-ml plastic straws, sealed, placed in a programmable cell freezer and cooled from room temperature (22 degrees C) to -6 degrees C. Straws were then seeded, held at -6 degrees C for 10 min and then cooled to -33 degrees C before being plunged into liquid nitrogen. Two or three embryos within a treatment group were thawed and assigned to be either cultured for 12 h prior to transfer or immediately nonsurgically transferred to a single mare. Embryo diameter decreased in all embryos upon initial exposure to cryoprotectant. Embryos in methanol shrank and recovered slightly to 76+/-8 % of their original diameter; however, embryos in glycerol continued to shrink, reaching 57+/-6 % of their original diameter prior to cryopreservation. Survival rates of embryos through Day 16 of pregnancy were 38 and 23%, respectively (P>0.05) for embryos cryopreserved in the presence of glycerol or methanol. There was no difference in pregnancy rates of mares receiving embryos that were cultured prior to transfer or not cultured (P>0.05). Preliminary experiments indicated that 48% methanol was not toxic to fresh equine embryos but methanol provided no advantage over glycerol as a cryoprotectant for equine blastocysts. PMID- 15289055 TI - Effects of egg yolk during the freezing step of cryopreservation on the viability of goat spermatozoa. AB - Four experiments were conducted to investigate the effects of egg yolk during the freezing step of cryopreservation (namely, the process except for the cooling step), on the viability of goat spermatozoa. The effects of egg yolk on sperm motility and acrosome integrity during the freezing step were investigated in Experiment 1. Spermatozoa diluted with Tris-citric acid-glucose (TCG) solution containing 20% (v/v) egg yolk were cooled to 5 degrees C, washed, and then frozen in TCG with egg yolk (TCG-Y), TCG without egg yolk (TGG-NY), 0.370 M trehalose with egg yolk (TH-Y), or trehalose without egg yolk (TH-NY). All extenders contained glycerol. In frozen-thawed spermatozoa, the inclusion of egg yolk in the freezing extenders increased (P<0.05) percentages of motile sperm, progressively motile sperm, and the recovery rate (ratio of post-thaw to pre freeze values), but decreased (P<0.05) acrosomal integrity. Moreover, extenders with trehalose had better (P<0.05) post-thaw sperm viability. In Experiment 2, the effects of egg yolk on acrosome status before and after freezing were studied. Egg yolk significantly decreased the proportion of intact acrosomes before freezing, leading to fewer (P<0.05) intact acrosomes post-thaw and lower (P<0.05) recovery rates for intact acrosomes. In Experiment 3, including sodium dodecyl sulfate (SDS) in a diluent containing egg yolk tended to preserve the acrosome compared with the egg yolk containing diluent free of SDS, however, spermatozoa had a lower (P<0.05) proportion of intact acrosomes than those in a yolk-free diluent. However, after cooling, spermatozoa were diluted with a glycerolated extender containing egg yolk. Therefore, the objective of Experiment 4 was to explore whether the egg yolk or glycerol was responsible for the reduced intact acrosome percentage. In this experiment, after cooling and washing the spermatozoa were diluted in TCG with glycerol and/or egg yolk. The combination of glycerol and egg yolk in the extender reduced (P<0.05) the proportion of intact acrosomes compared with egg yolk or glycerol alone. In conclusion, the inclusion of egg yolk significantly improved sperm motility, indicating its beneficial effects during the freezing step of cryopreservation; trehalose appeared to synergistically increase its cryoprotective effects. Furthermore, although neither glycerol nor egg yolk per se affected the proportion of intact acrosomes, the combination of the two significantly reduced the proportion of acrosome intact spermatozoa. PMID- 15289056 TI - Gene site saturation mutagenesis: a comprehensive mutagenesis approach. PMID- 15289057 TI - Single-stranded DNA family shuffling. PMID- 15289058 TI - Module shuffling. PMID- 15289059 TI - SCHEMA-guided protein recombination. PMID- 15289060 TI - Staggered extension process in vitro DNA recombination. PMID- 15289061 TI - Using incremental truncation to create libraries of hybrid enzymes. PMID- 15289062 TI - Random multirecombinant polymerase chain reaction. PMID- 15289063 TI - Gene library synthesis by structure-based combinatorial protein engineering. PMID- 15289064 TI - New enzymes from combinatorial library modules. PMID- 15289065 TI - Construction of protein fragment complementation libraries using incremental truncation. PMID- 15289066 TI - GigaMatrix: a novel ultrahigh throughput protein optimization and discovery platform. PMID- 15289067 TI - High throughput microplate screens for directed protein evolution. PMID- 15289068 TI - Periplasmic expression as a basis for whole cell kinetic screening of unnatural enzyme reactivities. PMID- 15289069 TI - Engineering the thermotolerance and pH optimum of family 11 xylanases by site directed mutagenesis. PMID- 15289070 TI - Screening for oxidative resistance. PMID- 15289071 TI - Consensus-based engineering of protein stability: from intrabodies to thermostable enzymes. PMID- 15289072 TI - Improving the functional expression of N-carbamoylase by directed evolution using the green fluorescent protein fusion reporter system. PMID- 15289073 TI - Directed evolution of lipases and esterases. PMID- 15289074 TI - Protein engineering of the cytochrome P450 monooxygenase from Bacillus megaterium. PMID- 15289075 TI - Directed evolution of aldolases. PMID- 15289076 TI - Changing the enantioselectivity of enzymes by directed evolution. PMID- 15289077 TI - Control of stereoselectivity in phosphotriesterase. PMID- 15289078 TI - Manipulation and analysis of polyketide synthases. PMID- 15289079 TI - Reactions catalyzed by mature and recombinant nonribosomal peptide synthetases. PMID- 15289080 TI - Engineering carotenoid biosynthetic pathways. PMID- 15289081 TI - Directed evolution of bacteriorhodopsin for device applications. PMID- 15289082 TI - Engineering antibody affinity by yeast surface display. PMID- 15289083 TI - A conditionally replicating virus as a novel approach toward an HIV vaccine. PMID- 15289085 TI - Effects of vitamin E on the NF-kappaB pathway in rats treated with the peroxisome proliferator, ciprofibrate. AB - Peroxisome proliferators (PPs) are a diverse group of nongenotoxic compounds, which induce hepatic tumors in rodents. The mechanisms leading to hepatic tumors have not been elucidated, but oxidative stress may play a role in the process. Previous studies in our laboratory have shown that peroxisome proliferators activate the transcription factor nuclear factor-kappa B (NF-kappaB) and that this activation is mediated at least in part by oxidative stress. We therefore hypothesized that increased dietary vitamin E would decrease NF-kappaB DNA binding in rodents treated with ciprofibrate (CIP). In this study, 36 male Sprague-Dawley rats were fed a purified diet containing varying levels of vitamin E (10, 50, 250 ppm alpha-tocopherol acetate). After 28 days on the purified diet, seven animals per vitamin E group received 0.01% CIP in the diet for 10 days. Electrophoretic mobility shift assays (EMSAs) showed that CIP treatment increased DNA binding of NF-kappaB. Increased dietary alpha-tocopherol acetate inhibited CIP-induced NF-kappaB DNA binding. Because NF-kappaB translocates to the nucleus upon the phosphorylation and degradation of inhibitor of IkappaB, we also used Western blots to measure cytosolic protein levels of IkappaBalpha and IkappaBbeta, and the IkappaB kinases, IKKalpha and IKKbeta. IkappaBalpha protein levels were decreased in all three CIP-treated groups, with the 10 ppm vitamin E diet also decreasing IkappaBalpha levels in control rats. No difference in IkappaBbeta protein levels was observed among any of the groups. The CIP-treated rats generally had lower protein levels of IKKalpha and IKKbeta. This study supports our working hypothesis that an increased antioxidant environment can inhibit CIP-mediated NF-kappaB induction. PMID- 15289086 TI - Mitochondrial oxidative stress in human hepatoma cells exposed to stavudine. AB - The toxicity of nucleoside reverse transcriptase inhibitors (NRTIs) is linked to altered mitochondrial DNA (mtDNA) replication and subsequent disruption of cellular energetics. This manifests clinically as elevated concentrations of lactate in plasma. The mechanism(s) underlying how the changes in mtDNA replication lead to lactic acidosis remains unclear. It is hypothesized that mitochondrial oxidative stress links the changes in mtDNA replication to mitochondrial dysfunction and ensuing NRTIs toxicity. To test this hypothesis, changes in mitochondrial function, mtDNA amplification efficiency, and oxidative stress were assessed in HepG2-cultured human hepatoblasts treated with the NRTI stavudine (2',3'-didehydro-2',3'-deoxythymidine or d4T) for 48 h. d4T produced significant mitochondrial dysfunction with a 1.5-fold increase in cellular lactate to pyruvate ratios. In addition, d4T caused a dose-dependent decrease in mtDNA amplification and a correlative increase in abundance of markers of mitochondrial oxidative stress. Manganese (III) meso-tetrakis (4-benzoic acid) porphyrin, MnTBAP, a catalytic antioxidant, ameliorated or reversed d4T-induced changes in cell injury, energetics, mtDNA amplification, and mitochondrial oxidative stress. In conclusion, d4T treatment elevates mitochondrial reactive oxygen species (ROS), enhances mitochondrial oxidative stress, and contributes mechanistically to NRTI-induced toxicity. These deleterious events may be potentiated in acquired immunodeficiency syndrome (AIDS) by human immunodeficiency virus (HIV) infection itself, coinfection (e.g., viral hepatitis), aging, substance, and alcohol use. PMID- 15289087 TI - gamma-Diketone neuropathy: axon atrophy and the role of cytoskeletal protein adduction. AB - Multifocal giant neurofilamentous axonal swellings and secondary distal degeneration have been historically considered the hallmark features of gamma diketone neuropathy. Accordingly, research conducted over the past 25 years has been directed toward discerning mechanisms of axonal swelling. However, this neuropathological convention has been challenged by recent observations that swollen axons were an exclusive product of long-term 2.5-hexanedione (HD) intoxication at lower daily dose-rates (e.g., 175 mg/kg/day); that is, higher HD dose-rates (e.g., 400 mg/kg/day) produced neurological deficits in the absence of axonal swellings. The observation that neurological toxicity can be expressed without axonal swelling suggests that this lesion is not an important pathophysiological event. Instead, several research groups have now shown that axon atrophy is prevalent in nervous tissues of laboratory animals intoxicated over a wide range of HD dose-rates. The well-documented nerve conduction defects associated with axon atrophy, in conjunction with the temporal correspondence between this lesion and the onset of neurological deficits, strongly suggest that atrophy has pathophysiological significance. In this commentary, we present evidence that supports a pathognomonic role for axon atrophy in gamma-diketone neuropathy and suggests that the functional consequences of this lesion mediate the corresponding neurological toxicity. Previous research has demonstrated that HD interacts with proteins via formation of pyrrole adducts. We therefore discuss the possibility that this chemical process is essential to the mechanism of atrophy. Evidence presented in this review suggests that "distal axonopathy" is an inaccurate classification and future nosological schemes should be based on the apparent primacy of axon atrophy. PMID- 15289088 TI - Metavanadate causes cellular accumulation of copper and decreased lysyl oxidase activity. AB - Selected indices of copper metabolism in weanling rats and fibroblast cultures were progressively altered in response to increased levels of sodium metavanadate. In diets, vanadium was added in amounts ranging from 0 to 80 microg V/g of diet, that is, 0-1.6 micromol V/g of diet. In fibroblast cultures, vanadium ranged from 0 to 400 nmol V/ml. The inhibition of P-ATPase-7A activity by metavanadate, important to copper egress from cells, was a primary focus. In skin, and tendon, the copper concentration was increased in response to increased dietary levels of metavanadate, whereas lysyl oxidase activity, a secreted cuproprotein, was reduced. The reduction in lysyl oxidase activity was also accompanied by reduced redox cycling potential of isolated fractions of lysyl oxidase, presumably due to reduced lysyltyrosyl quinone (LTQ) formation at the active site of lysyl oxidase. In contrast, liver copper concentrations and plasma ceruloplasmin activity were not affected by metavanadate exposure. However, semicarbazide-sensitive benzylamine oxidase (SCBO) activity, which was taken as an indirect measure of vascular adhesive protein-1 (VAP-1), was increased. In cultured fibroblasts, cellular copper was also increased and lysyl oxidase decreased in response to metavanadate. Moreover, the steady-state levels of atp7a and lysyl oxidase mRNAs were not affected by addition of metavanadate to culture medium up to 200 nmol/ml. Taken together, these data suggest that pathways involving copper egress and lysyl oxidase activation are particularly sensitive to metavanadate exposure through processes that are predominately posttranslational. PMID- 15289089 TI - Disruption of mitochondrial membrane potential during apoptosis induced by PSC 833 and CsA in multidrug-resistant lymphoid leukemia. AB - Previous findings from our laboratory demonstrated that when used at low concentration (0.1 microg ml(-1)), CsA as well as its analog PSC 833 were able to revert the MDR phenotype, while at high concentration (1 microg ml(-1)) were able to induce apoptosis. CsA induced apoptosis in leukemia cell lines sensitive (LBR ) and resistant to vincristine (LBR-V160), and doxorubicin (LBR-D160), while PSC 833 only induced apoptosis in vincristine-resistant cell line (LBR-V160). In this work, we investigated mitochondrial-associated mechanisms during CsA- and PSC 833 induced apoptosis. Mitochondrial function was evaluated by recording changes in its transmembrane potential, cytochrome c release, and caspase activation cascade. Results showed that CsA- and PSC 833-induced apoptosis was associated with mitochondrial depolarization, through potentiometric measurements with JC-1 and DiOC(6) probes. Collapse of mitochondrial potential in these cell lines after CsA treatment was followed by cytochrome c release to the cytosol, reaching an increase of 2.61-fold in LBR-, 1.98-fold in LBR-V160, and 3.01-fold in the case of LBR-D160. However, in the case of PSC 833 treatment, induction of apoptosis in LBR-V160 was associated with mitochondrial depolarization followed by a lower cytochrome c release of 1.15-fold as compared with untreated cells. Caspase 3 activation was clearly observed in LBR-, LBR-V160, and LBR-D160 after CsA treatment, while in LBR-V160, PSC 833 was less effective inducing activation of this caspase. Neither caspase 6 nor 8 activity was observed in these three cell lines. Our results suggest that during CsA- and PSC 833-induced apoptosis, mitochondrial dysfunction occurs. This is mediated through mitochondrial events, associated with an evident decrease in DeltaPsi(m), cytochrome c release and caspase 3 activation. PMID- 15289090 TI - Determination of myocardium to plasma concentration ratios of five antipsychotic drugs: comparison with their ability to induce arrhythmia and sudden death in clinical practice. AB - Reviewing available data shows that most of antipsychotic drugs are associated with arrhythmia and sudden death. Experimental studies have shown a HERG channel blockade, a dose-dependent increase in duration of action potential or of QT interval, with various degrees of indicators of serious arrhythmogenicity. However, it seems difficult to relate these in vitro and in vivo preclinical models to clinical findings, in part, because the relationship between concentrations used and in vivo tissue concentrations during treatment in man is not known. Consequently, we established the myocardium to plasma concentration ratios for a series of antipsychotic drugs by intraperitoneal administration of different level doses to the guinea pig. Then, we compared these values to their ability to induce arrhythmia or torsade de pointes in clinical practice. The myocardium to plasma concentration ratios were 2.2 for clozapine, 2.7 for olanzapine, 3.1 for sertindole, 4.5 for risperidone, and 6.4 for haloperidol. These data suggest that when the ratio is higher than 4, arrhythmia and sudden death may be expected. On the contrary, when the ratio is less than 3, little effect may be predicted. These results underscore the importance of interpreting HERG channel data and electrophysiological data in the context of other pharmacokinetic parameters such as myocardium to plasma distribution. PMID- 15289091 TI - Symposium summary: children's health risk--what's so special about the developing immune system? AB - In recent years, there has been increasing regulatory pressure to protect the health of children, with the basic tenet being that children differ significantly from adults in their biological or physiological responses to chemical exposures. In a regulatory context, this has been translated to mean a requirement for an additional 10-fold safety factor for environmental contaminants, specialized tests, or both. Much of the initial focus has been on the developing endocrine and nervous systems; but increasingly, the developing immune system has been identified as a potential target organ for chemically mediated toxicity. More recently, the question has been raised regarding whether the current state of science supports the creation of developmental immunotoxicology (DIT) test guidelines. What is needed is a risk-based evaluation of the biology associated with the proposed differential sensitivity between children and adults and the impact of that assessment on additional regulatory measures to protect children in risk assessment analyses. Additionally, an understanding of whether the developing immune system shows greater susceptibility, either qualitatively or quantitatively, to chemical perturbation is critical. To address the question "What's so special about the developing immune system?" a symposium was organized for the 2003 Society of Toxicology annual meeting that brought together risk assessors, clinicians, immunologists, and toxicologists. PMID- 15289092 TI - Biological agents with potential for misuse: a historical perspective and defensive measures. AB - Biological and chemical agents capable of producing serious illness or mortality have been used in biowarfare from ancient times. Use of these agents has progressed from crude forms in early and middle ages, when snakes and infected cadavers were used as weapons in battles, to sophisticated preparations for use during and after the second World War. Cults and terrorist organizations have attempted the use of biological agents with an aim to immobilize populations or cause serious harm. The reasons for interest in these agents by individuals and organizations include relative ease of acquisition, potential for causing mass casualty or panic, modest financing requirement, availability of technology, and relative ease of delivery. The Centers for Disease Control and Prevention has classified Critical Biological Agents into three major categories. This classification was based on several criteria, which include severity of impact on human health, potential for delivery in a weapon, capacity to cause panic and special needs for development, and stockpiling of medication. Agents that could cause the greatest harm following deliberate use were placed in category A. Category B included agents capable of producing serious harm and significant mortality but of lower magnitude than category A agents. Category C included emerging pathogens that could be developed for mass dispersion in future and their potential as a major health threat. A brief description of the category A bioagents is included and the pathophysiology of two particularly prominent agents, namely anthrax and smallpox, is discussed in detail. The potential danger from biological agents and their ever increasing threat to human populations have created a need for developing technologies for their early detection, for developing treatment strategies, and for refinement of procedures to ensure survival of affected individuals so as to attain the ultimate goal of eliminating the threat from intentional use of these agents. International treaties limiting development and proliferation of weapons and continuing development of defense strategies and safe guards against agents of concern are important elements of plans for eliminating this threat. PMID- 15289093 TI - Spatial and temporal risk factors for the early detection of Trypanosoma brucei rhodesiense sleeping sickness patients in Tororo and Busia districts, Uganda. AB - We have carried out a study of risk factors for early detection of Trypanosoma brucei rhodesiense sleeping sickness. Records of sleeping sickness patients from 1987 to 2001 from Tororo and Busia districts in Uganda were reviewed for their village of origin and clinical stage (early or late). All villages that reported sleeping sickness and fixed post-diagnostic sleeping sickness health units in Tororo and Busia districts were geo-referenced. The spatial distribution of early and late stage patient detection by health units was analysed using Geographical Information Systems (GIS). Of 1316 sleeping sickness patients admitted at the Livestock Health Research Institute and Busolwe hospitals and Lumino health centre from Tororo and Busia districts, 471 (35.8%) were early stage, 825 (62.7%) were late stage, while 20 (1.5%) were not staged. Five hundred and eighty-five (44.5%) came from within a 10 km radius of the reporting health units. After multivariate analysis, the proportion of early stage patients detected was found to be significantly associated with patients originating from within a 10 km radius of the health unit (P < 0.01), with adults (>19 years) (P < 0.01), and with annual parish incidence (P < 0.01). Application of GIS and the early to late stages ratio are an informative and powerful means of determining efficiency of surveillance of sleeping sickness. PMID- 15289094 TI - Prevalence of rabies and LPM paramyxovirus antibody in non-hematophagous bats captured in the Central Pacific coast of Mexico. AB - To investigate if non-hematophagous bats play a role in outbreaks of rabies and blue eye disease (LPMV), we studied the seroprevalence against both agents in several species of non-hematophagous bats on the sub-tropical Pacific coast of the state of Colima, Mexico. The survey covered a predominantly agricultural area (disturbed), and an area dominated by semideciduous dry forest (undisturbed). A total of 151 non-hematophagous bats of 16 species were captured from the two areas. Fifty-six (37%) had antirabic antibodies (Ab) while 87 (58%) did not and 8 samples (5%) had to be discarded because of hemolysis. A much lower (P<0.05) prevalence of antirabic Ab was found in bats caught in disturbed areas (22.7%) compared with those from undisturbed areas (51.9%). The presence of antirabic Ab was not related to sex, genera or feeding habits. The higher prevalence found in bats in the undisturbed area may be the result of more frequent interspecies encounters. Of the 108 sera analyzed for antibodies against LPMV, only one was positive (a male Rhogeessa parvula major, captured in the undisturbed area). This suggests that bats in the surveyed localities do not play a role in the epidemiology of LPMV. PMID- 15289095 TI - Risk of Plasmodium vivax malaria reintroduction in Uzbekistan: genetic characterization of parasites and status of potential malaria vectors in the Surkhandarya region. AB - Plasmodium vivax malaria was eradicated from Uzbekistan in 1961. Due to resurgence of the disease in neighbouring states and massive population migration, there has been an increase of P. vivax malaria, imported from Tajikistan, resulting in a number of indigenous cases being identified in areas bordering that country. A molecular study using the merozoite surface protein 1 (msp-1) gene as a marker was performed on 24 P. vivax genomic isolates from 12 indigenous and 10 imported malaria cases that occurred in the Surkhandarya region during the summer of 2002. Results have shown a significant difference in the frequency of msp-1 types between indigenous and imported isolates, the latter showing greater genetic heterogeneity. An entomological investigation in the area suggested that three Anopheles species, namely A. superpictus, A. pulcherrimus and A. hyrcanus may have a potential role in the endemic transmission of P. vivax. PMID- 15289096 TI - A potential disruptive technology in vaccine development: gene-based vaccines and their application to infectious diseases. AB - Vaccine development requires an amalgamation of disparate disciplines and has unique economic and regulatory drivers. Non-viral gene-based delivery systems, such as formulated plasmid DNA, are new and potentially disruptive technologies capable of providing 'cheaper, simpler, and more convenient-to-use' vaccines. Typically and somewhat ironically, disruptive technologies have poorer product performance, at least in the near-term, compared with the existing conventional technologies. Because successful product development requires that the product's performance must meet or exceed the efficacy threshold for a desired application, the appropriate selection of the initial product applications for a disruptive technology is critical for its successful evolution. In this regard, the near term successes of gene-based vaccines will likely be for protection against bacterial toxins and acute viral and bacterial infections. Recent breakthroughs, however, herald increasing rather than languishing performance improvements in the efficacy of gene-based vaccines. Whether gene-based vaccines ultimately succeed in eliciting protective immunity in humans to persistent intracellular pathogens, such as HIV, malaria and tuberculosis, for which the conventional vaccine technologies have failed, remains to be determined. A success against any one of the persistent intracellular pathogens would be sufficient proof that gene based vaccines represent a disruptive technology against which future vaccine technologies will be measured. PMID- 15289097 TI - Clinical outcomes in a randomized controlled study comparing azathioprine and prednisolone versus prednisolone alone in the treatment of severe leprosy type 1 reactions in Nepal. AB - The ILEP nerve function impairment and reaction research programme (INFIR 2) was a group of clinical trials conducted to identify second-line treatments for severe leprosy type 1 reactions (T1R). This paper presents the clinical results of one of these trials in which azathioprine was used in combination with short course prednisolone to ascertain if the combination was effective in controlling the symptoms and signs of reaction. Forty patients were alternately assigned to a 12-week treatment with either AP (12 weeks azathioprine at 3mg/kg/d plus 8 week reducing course prednisolone starting at 40mg/d) or P (12-week reducing course prednisolone starting at 40mg/d). Evaluation included serial quantitative clinical assessments. The overall frequency of side effects was similar in both groups. Results show that there was no difference in clinical outcome in the AP and P groups and a similar number of patients in each group required extra prednisolone for worsening clinical features. We conclude that a 12-week course of azathioprine at 3mg/kg/day plus an 8 week reducing course of prednisolone starting at 40mg/d is as effective as a 12 week reducing course of prednisolone starting at 40mg/d and that the combination therapy is well-tolerated in severe leprosy T1R patients. PMID- 15289098 TI - Community-level analysis of risk of vector-borne disease. AB - Ecological community structure is particularly important in vector-borne zoonotic diseases with complex life cycles. Qualitative community model analysis may provide a meaningful alternative to standard population-based models of vector borne disease. We built on recent mathematical developments in qualitative community modeling coupled with conventional biomathematical models of vector borne disease transmission, to provide a procedure to analyze risk. Using this procedure, we can hypothesize changes in risk of vector-borne disease from disturbances, such as control measures, habitat alteration, or global warming. We demonstrate the application of this procedure to an oak forest community to predict the risk of Lyme disease. Our predictions of Lyme disease risk in an oak forest community confirm reports of positive associations between deer abundance and risk of disease and are consistent with published observations. PMID- 15289099 TI - Development of a single tube hemi-nested PCR for genus-specific detection of Plasmodium in oligoparasitemic patients. AB - Primers targeting the Plasmodium small-subunit (SSU) rDNA were designed to amplify DNA from P. vivax, P. falciparum, P. malariae, and P. ovale, using conventional PCR, two-step nested PCR (HNPCR), and single tube hemi-nested PCR (STHNPCR). The limit of detection of parasite DNA for the conventional PCR, HNPCR, and STHNPCR were 10 pg, 0.01 pg, and 0.1 pg, respectively, indicating that the STHNPCR is 100-fold more sensitive than conventional PCR, and only 10 times less sensitive than HNPCR. In addition, the detection limit was also defined using blood from a patient infected with P. falciparum. Using the saponin method, the detection limit of the conventional PCR, HNPCR, and STHNPCR were 70, 0.7, and 0.07 parasites/microl, respectively. Finally, the three techniques were evaluated using blood from 30 patients receiving antimalarial treatment, and negative by microscopy and conventional PCR. The HNPCR could still detect specific DNA in 16/30 patients, whereas STHNPCR detected parasite DNA in 10/30 patients, but the difference was not statistically significant. No significant correlation was found between presence of clinical manifestations and presence of parasite DNA, detected by either HNPCR or STHNPCR. We conclude that these sensitive molecular diagnostic systems can be used for the diagnosis of asymptomatic oligoparasitemic patients. PMID- 15289100 TI - Sequence context modulates the stability of a GxxxG-mediated transmembrane helix helix dimer. AB - To quantify the relationship between sequence and transmembrane dimer stability, a systematic mutagenesis and thermodynamic study of the protein-protein interaction residues in the glycophorin A transmembrane helix-helix dimer was carried out. The results demonstrate that the glycophorin A transmembrane sequence dimerizes when its GxxxG motif is abolished by mutation to large aliphatic residues, suggesting that the sequence encodes an intrinsic propensity to self-associate independent of a GxxxG motif. In the presence of an intact GxxxG motif, the glycophorin A dimer stability can be modulated over a span of 0.5 kcal mol(-1) to +3.2 kcal mol(-1) by mutating the surrounding sequence context. Thus, these flanking residues play an active role in determining the transmembrane dimer stability. To assess the structural consequences of the thermodynamic effects of mutations, molecular models of mutant transmembrane domains were constructed, and a structure-based parameterization of the free energy change due to mutation was carried out. The changes in association free energy for glycophorin A mutants can be explained primarily by changes in packing interactions at the protein-protein interface. The energy cost of removing favorable van der Waals interactions was found to be 0.039 kcal mol(-1) per A2 of favorable occluded surface area. The value corresponds well with estimates for mutations in bacteriorhodopsin as well as for those mutations in the interiors of soluble proteins that create packing defects. PMID- 15289101 TI - Expression, purification and the 1.8 angstroms resolution crystal structure of human neuron specific enolase. AB - Human neuron-specific enolase (NSE) or isozyme gamma has been expressed with a C terminal His-tag in Escherichia coli. The enzyme has been purified, crystallized and its crystal structure determined. In the crystals the enzyme forms the asymmetric complex NSE x Mg2 x SO4/NSE x Mg x Cl, where "/" separates the dimer subunits. The subunit that contains the sulfate (or phosphate) ion and two magnesium ions is in the closed conformation observed in enolase complexes with the substrate or its analogues; the other subunit is in the open conformation observed in enolase subunits without bound substrate or analogues. This indicates negative cooperativity for ligand binding between subunits. Electrostatic charge differences between isozymes alpha and gamma, -19 at physiological pH, are concentrated in the regions of the molecular surface that are negatively charged in alpha, i.e. surface areas negatively charged in alpha are more negatively charged in gamma, while areas that are neutral or positively charged tend to be charge-conserved. PMID- 15289102 TI - Apo and Holo structures of an NADPH-dependent cinnamyl alcohol dehydrogenase from Saccharomyces cerevisiae. AB - The crystal structure of Saccharomyces cerevisiae ScAdh6p has been solved using the anomalous signal from the two zinc atoms found per subunit, and it constitutes the first structure determined from a member of the cinnamyl alcohol dehydrogenase family. ScAdh6p subunits exhibit the general fold of the medium chain dehydrogenases/reductases (MDR) but with distinct specific characteristics. In the three crystal structures solved (two trigonal and one monoclinic), ScAdh6p molecules appear to be structural heterodimers composed of one subunit in the apo and the second subunit in the holo conformation. Between the two conformations, the relative disposition of domains remains unchanged, while two loops, Cys250 Asn260 and Ile277-Lys292, experience large movements. The apo-apo structure is disfavoured because of steric impairment involving the loop Ile277-Lys292, while in the holo-holo conformation some of the hydrogen bonds between subunits would break apart. These suggest that the first NADPH molecule would bind to the enzyme much more tightly than the second. In addition, fluorimetric analysis of NADPH binding demonstrates that only one cofactor molecule binds per dimer. Therefore, ScAdh6p appears to function according to a half-of-the-sites reactivity mechanism, resulting from a pre-existing (prior to cofactor binding) tendency for the structural asymmetry in the dimer. The specificity of ScAdh6p towards NADPH is mainly due to the tripod-like interactions of the terminal phosphate group with Ser210, Arg211 and Lys215. The size and the shape of the substrate-binding pocket correlate well with the substrate specificity of ScAdh6p towards cinnamaldehyde and other aromatic compounds. The structural relationships of ScAdh6p with other MDR structures are analysed. PMID- 15289103 TI - Crystal structures of novel non-peptidic, non-zinc chelating inhibitors bound to MMP-12. AB - Human macrophage elastase (MMP-12) plays an important role in inflammatory processes and has been implicated in diseases such as emphysema and chronic obstructive pulmonary disease (COPD). It is therefore an attractive target for therapeutic agents. As part of a structure-based drug design programme to find new inhibitors of MMP-12, the crystal structures of the MMP-12 catalytic domain (residues 106-268) complexed to three different non-peptidic small molecule inhibitors have been determined. The structures reveal that all three ligands bind in the S1' pocket but show varying degrees of interaction with the Zn atom. The structures of the complexes with inhibitors CP-271485 and PF-00356231 reveal that their central morpholinone and thiophene rings, respectively, sit over the Zn atom at a distance of approximately 5A, locating the inhibitors halfway down the S1' pocket. In both of these structures, an acetohydroxamate anion, an artefact of the crystallisation solution, chelates the zinc atom. By contrast, the acetohydroxamate anion is displaced by the ligand in the structure of MMP-12 complexed to PD-0359601 (Bayer), a potent zinc chelating N-substituted biaryl butyric acid, used as a reference compound for crystallisation. Although a racemate was used for the crystallisation, the S enantiomer only is bound in the crystal. Important hydrophobic interactions between the inhibitors and residues from the S1' pocket are observed in all of the structures. The relative selectivity displayed by these ligands for MMP-12 over other MMP family members is discussed. PMID- 15289104 TI - Crystallographic analysis of synechocystis cyanoglobin reveals the structural changes accompanying ligand binding in a hexacoordinate hemoglobin. AB - The crystal structures of cyanide and azide-bound forms of the truncated hemoglobin from Synechocystis are presented at 1.8 angstroms resolution. A comparison with the structure of the endogenously liganded protein reveals a conformational shift unprecedented in hemoglobins, and provides the first picture of a hexacoordinate hemoglobin in both the bis-histidyl and the exogenously coordinated states. The structural changes between the different conformations are confined to two regions of the protein; the B helix, and the E helix, including the EF loop. A molecular "hinge" controlling movement of the E helix is observed in the EF loop, which is composed of three principal structural elements: Arg64, the heme-d-propionate, and a three-residue extension of the F helix. Additional features of the structural transition between the two protein conformations are discussed as they relate to the complex ligand-binding behavior observed in hexacoordinate hemoglobins, and the potential physiological function of this class of proteins. PMID- 15289105 TI - History of FIGO. PMID- 15289107 TI - The Presidents of FIGO. PMID- 15289108 TI - General assemblies of FIGO. PMID- 15289109 TI - FIGO's Constitution and Bye-Laws. PMID- 15289110 TI - FIGO'S finances. PMID- 15289111 TI - FIGO administration: insight into activities. PMID- 15289112 TI - FIGO Triennial Congress: the World Congress in Gynecology and Obstetrics. PMID- 15289113 TI - Societies in relationship with FIGO. PMID- 15289114 TI - History of the International Journal of Gynecology & Obstetrics. PMID- 15289115 TI - The FIGO manual of human reproduction. PMID- 15289116 TI - International Medical Ethics: The FIGO Committee for the Ethical Aspects of Human Reproduction and Women's Health. PMID- 15289117 TI - FIGO educational activities. PMID- 15289118 TI - The FIGO save the mothers initiative. PMID- 15289119 TI - History of the FIGO Committee for Women's Sexual and Reproductive Rights. PMID- 15289120 TI - FIGO initiative for the prevention and treatment of vaginal fistula. PMID- 15289121 TI - The WHO/FIGO Alliance for Women's Health. PMID- 15289122 TI - A FIGO initiative for the 21st century: eliminate all forms of violence against women worldwide. PMID- 15289123 TI - FIGO in the future. PMID- 15289124 TI - Treatment of conjunctival squamous cell carcinoma with photodynamic therapy. AB - PURPOSE: The aim of this study was to describe the clinical and angiographic response of squamous cell carcinoma (SCC) of the conjunctiva to treatment with photodynamic therapy (PDT). DESIGN: Interventional case series. METHODS: In a prospective study, three patients (62 to 86 years old) with SCC of the conjunctiva were treated with PDT. Patients received one to three treatments of verteporfin (6 mg/m(2) body surface area, intravenously). The light dose was calculated as 50 J/cm(2). All tumors were irradiated 1 minute after injection. The mean follow-up time was 8.6 months (7 to 12 months). Main outcome measurements were clinical and angiographic response and treatment-related side effects. RESULTS: One week after treatment, angiographic occlusion of tumor vasculature and normal conjunctival vessels was observed in all patients. Tumor regression was noted in all patients 1 month after treatment. Two patients had complete regression (clinical and angiographic observation) after one or two treatments for the entire follow-up time. One tumor involved large aspects of the conjunctiva and cornea. In this case, only the treated areas showed tumor regression. PDT caused minimal temporary local irritation in two patients, and small conjunctival hemorrhages and mild transient chemosis in the three eyes directly after treatment. One patient had infusion-related back pain. CONCLUSION: The preliminary results of this study suggest that PDT may be a valuable addition to the treatment of patients with SCC of the conjunctiva. However, longer follow up is necessary to assess the duration and degree of tumor control. PMID- 15289125 TI - The effect of corneal flap on optical aberrations. AB - PURPOSE: To evaluate the changes in ocular aberrations induced by corneal flap creation. DESIGN: Prospective interventional nonrandomized clinical trial. METHODS: This study included 15 patients who were scheduled for laser in situ keratomileusis. A nasal hinge flap was created, using the Nidek MK-2000 microkeratome and then replaced without performing laser ablation. The ocular aberrations were measured before and after flap creation using the Nidek Optical Path Difference Scanning System ARK-10000. RESULTS: The root mean square wavefront errors of the higher-order optical aberrations (third-, fourth-, fifth , and sixth-order aberrations) were not significantly altered at 1 week postsurgery compared with the preoperative values (P >.35). CONCLUSIONS: Creating a corneal flap with the Nidek MK-2000 microkeratome did not induce changes in higher-order optical aberrations as measured with the Nidek Optical Path Difference Scanning System ARK-10000 during the early postoperative period. PMID- 15289126 TI - Indocyanine green-assisted internal limiting membrane removal in epiretinal membrane surgery: a clinical and histologic study. AB - PURPOSE: To evaluate the clinical outcome and quantify histologically the amount of microscopic internal limiting membrane (ILM) and epiretinal membrane (ERM) present in ILM and ERM specimens obtained from ERM surgery. DESIGN: Interventional consecutive case series. METHODS: Patients scheduled for ERM surgery were recruited prospectively. Pars plana vitrectomy, removal of ERM, and ILM peeling with indocyanine green (ICG) staining were performed in all patients. Epiretinal membrane and ILM specimens were sent for histologic examination. The amount of ERM present in ILM specimens and the amount of ILM present in ERM specimens were quantified by manual counting. Outcome measures include change in best-corrected visual acuity (BCVA), proportion of cases with 2 or more lines of visual improvement, anatomic outcome, proportions of microscopic ERM within ILM, and microscopic ILM within ERM. RESULTS: Eighteen eyes in 18 patients were operated with a mean follow-up of 19.3 months. There were 13 primary ERMs and five secondary ERMs. The mean logMAR BCVA improved from 0.83 preoperatively to 0.49 postoperatively (P <.001). The mean lines of improvement in BCVA was 3.3 lines with 14 patients (77.8%) who had 2 or more lines of BCVA improvement. Histologic evaluation of the specimens showed no significant correlation with the final BCVA of 20/50 or better. Eleven (61.1%) of the ILM specimens showed various amount of microscopic ERM and 16 (88.9%) of the ERM specimens showed various amount of ILM fragments. The mean proportion of ERM within ILM specimens was 4.69% and that of ILM within ERM specimens was 51.5%. No significant recurrence of ERM was found. CONCLUSION: Recurrence of ERM may be minimized by removing residual microscopic ERM present on the ILM. Indocyanine green-assisted ILM peeling in ERM surgery appears to have favorable visual and anatomic outcomes. PMID- 15289127 TI - The long-term results of keratoplasty in eyes with a glaucoma drainage device. AB - PURPOSE: To study the outcome of penetrating keratoplasty (PK) in eyes with a glaucoma drainage device (GDD). DESIGN: Retrospective case-controlled study. METHODS: We reviewed all patients who underwent PK from December 1986 to September 2002 at the University of California, Davis (n = 1,974). We identified 33 patients (40 grafts) who were treated with a GDD and followed up for 6 months or more after PK. Graft survival and glaucoma control were compared with grafts in patients without glaucoma (n = 40) and patients with medically controlled glaucoma (n = 17). Kaplan-Meier survival analysis, log rank test, repeated measures analysis of variance (ANOVA), Fisher exact test, and chi(2) were used in group comparisons. Multivariate analysis was performed using the Cox proportional hazards model. RESULTS: The percentages of clear grafts in the glaucoma drainage device group were 58.5% and 25.8% at 1 and 2 years, respectively. At these time points, glaucoma was controlled in 74.0% and 63.1% of the eyes, respectively. Both medically controlled glaucoma patients and nonglaucomatous patients had higher graft survival percentages at comparable time points. The presence of a GDD was an important factor influencing graft survival (Hazard ratio = 6.8). CONCLUSION: A GDD implant is an independent risk factor for graft failure. Although these devices are effective in controlling intraocular pressure (IOP) in the majority of eyes in the presence of PK, corneal graft clarity is often compromised. PMID- 15289128 TI - Inflow of ocular surface fluid through clear corneal cataract incisions: a laboratory model. AB - PURPOSE: To evaluate the self-sealing properties of standard clear corneal cataract incisions during two events: (1) application of mechanical external pressure, or (2) controlled fluctuation of intraocular pressure (IOP). DESIGN: Laboratory investigation. METHODS: Eight fresh human donor globes were prepared for Miyake video microscopy. A standard two-plane 3-mm clear corneal incision was created and a 3 x 3-mm sponge soaked with India ink was placed on the wound surface. One globe with a sutured corneal wound served as the control. A transcleral cannula was inserted and connected to a bottle of saline. Intraocular pressure was modified varying the bottle height. External pressure was applied through manual contact on different regions of cornea. RESULTS: Four of seven eyes demonstrated intraocular presence of ink, three of them after external manipulation and another after varying the IOP. CONCLUSION: Self-sealing properties of unsutured clear corneal wounds were compromised in our model. These data may give insight into the possible mechanisms involved in the inflow of extraocular fluid into the eye through clear corneal cataract incisions. PMID- 15289129 TI - Endothelial cell density after posterior lamellar keratoplasty (Melles techniques): 3 years follow-up. AB - PURPOSE: To report the midterm endothelial cell density measurements after posterior lamellar keratoplasty (Melles techniques). DESIGN: Cohort study. METHODS: Fifteen consecutive eyes of 15 patients in whom a posterior lamellar keratoplasty procedure was performed for pseudophakic bullous keratopathy or Fuchs' endothelial dystrophy were evaluated. In 11 corneas the donor tissue was inserted through a 9.0-mm sclerocorneal pocket incision (technique A); in four cases the donor was folded and inserted through a 5.0-mm incision (technique B). Specular microscopy was performed at 6, 12, 24, and 36 months after surgery, to measure the endothelial cell density. RESULTS: Mean postoperative endothelial cell density averaged 2,126 cells/mm(2) (+/-548) at 6 months, 1,859 cells/mm(2) (+/-477) at 12 months, 1,385 cells/mm(2) (+/-451) at 24 months, and 1,047 cells/mm(2) (+/-425) at 36 months. CONCLUSION: In posterior lamellar keratoplasty, the donor corneal endothelium showed a decrease in cell density similar to that after conventional full-thickness penetrating keratoplasty. PMID- 15289130 TI - Optical coherence tomography (OCT) macular and peripapillary retinal nerve fiber layer measurements and automated visual fields. AB - PURPOSE: To investigate the structure-function relationship between optical coherence tomography (OCT) macular retinal and peripapillary nerve fiber layer (NFL) thickness and automated visual field (VF) findings. DESIGN: Cross-sectional observational study. METHODS: Retrospective institutional study where 150 consecutive eyes (101 subjects) from a glaucoma service were included. All the participants had full ophthalmic evaluation, VF testing and prototype OCT scanning at the same visit. Orthogonal OCT macular analysis was obtained to maximize the sampling of the area of interest. Pearson age-adjusted correlation was determined between macular retinal thickness and peripapillary NFL thickness. Area under the receiver operator characteristics (AROC) curves for the association between macular retinal thickness and peripapillary NFL thickness and VF findings were calculated in a subgroup of eyes without VF defect and eyes with VF defect confined to one hemifield. RESULTS: The correlation between macular retinal and peripapillary NFL measurements ranged between r =.27 to.54 for quadrants,.44 to.55 for hemiretina, and.52 for the overall mean. Areas under the receiver operator characteristics for macular thickness were higher in areas corresponding to the VF defect location than the noncorresponding locations. Areas under the receiver operator characteristics for peripapillary NFL thickness were higher than for the macular retinal thickness. Including both macular retinal thickness and peripapillary NFL thickness measurements in the logistic regression model yielded AROCs (range:.69 -.77) similar to those found for the peripapillary NFL alone. CONCLUSION: Macular retinal thickness, as measured by OCT, was capable of detecting glaucomatous damage and corresponded with peripapillary NFL thickness; however, peripapillary NFL thickness had higher sensitivity and specificity for the detection of VF abnormalities. PMID- 15289131 TI - A laboratory evaluation of antibiotic therapy for ciprofloxacin-resistant Pseudomonas aeruginosa. AB - PURPOSE: The emergence of ciprofloxacin-resistant Pseudomonas aeruginosa (CRPA) has created a new therapeutic challenge in ophthalmology. We evaluated ophthalmic antibiotics in vitro and in a rabbit keratitis model to determine effective therapy. DESIGN: Experimental laboratory investigation. METHODS: The susceptibilities of 12 CRPA isolates were determined in vitro for amikacin, ceftazidime, tobramycin, polymyxin B, gentamicin, ticarcillin, and the fluoroquinolones (that is, ciprofloxacin, ofloxacin, levofloxacin, gatifloxacin, and moxifloxacin) using E-tests and National Committee of Clinical Laboratory Standards. A rabbit keratitis model was used to determine the reduction in colony counts of CRPA and ciprofloxacin-susceptible P. aeruginosa (CSPA) isolates following topical treatment with polymyxin B/trimethoprim, tobramycin (14 mg/ml), ceftazidime (50 mg/ml), and ciprofloxacin (3 mg/ml). RESULTS: For 12 CRPA isolates, the susceptibilities and median minimum inhibitory concentrations ([MIC]microg/ml) were as follows: amikacin (92%, 14.0), ceftazidime (75%, 4.0), tobramycin (67%, 1.75), polymyxin B (42%, 7.0), gentamicin (17%, 7.0), ticarcillin (0%, >32.0), and all fluoroquinolones (0%, >32.0). While no antibiotic regimen reduced colony counts in the time frame of the animal model for CRPA, ciprofloxacin alone demonstrated a significant decrease in colony counts for CSPA. Comparing CRPA with CSPA, both tobramycin and ciprofloxacin demonstrated a significant decrease in colony counts for CSPA. CONCLUSION: Our laboratory studies suggest that current antibiotics may be suboptimal in treating CRPA keratitis. Until new antibiotics are available, combination therapy such as fortified tobramycin and ticarcillin, and others may prove effective in aggressive topical long-term therapy. PMID- 15289132 TI - Endophthalmitis caused by Streptococcus pneumoniae. AB - PURPOSE: To investigate clinical settings, management strategies, antibiotic sensitivities, and visual acuity outcomes of endophthalmitis caused by Streptococcus pneumoniae. DESIGN: Retrospective, observational case series. METHODS: Records were reviewed of all patients with culture-positive endophthalmitis caused by Streptococcus pneumoniae treated at the Bascom Palmer Eye Institute between January 1, 1989 and December 31, 2003. MAIN OUTCOME MEASURES: Visual acuity and antibiotic sensitivities. RESULTS: Twenty-seven eyes of 27 patients met study inclusion criteria. The median follow-up was 7 months (range, 3 months to 10 years). Clinical settings included acute postoperative (10 eyes), corneal stitch abscess (5), corneal ulcer (3), bleb-associated (4), post trauma (3), and endogenous (2). Eighteen cases (67%) were acute-onset (less than 3 weeks from event), with a median interval between event and presentation of endophthalmitis of 5 days (range, 1 day to 16 days). Nine cases (33%) were delayed-onset (median, 27 months; range, 3 to 121 months). Initial visual acuity was hand motions or better in 11 cases (41%). Initial therapeutic procedures included vitreous tap and injection of intravitreal antibiotics in 15 eyes (56%), pars plana vitrectomy and injection of intravitreal antibiotics in 10 eyes (37%), and evisceration in 2 eyes (7%). Seventeen (68%) of 25 eyes received intravitreal dexamethasone. Twelve patients (48%) received additional doses of intraocular antibiotics, and 11 patients (44%) underwent secondary surgical intervention within one week of diagnosis. The Streptococcus pneumoniae isolates showed sensitivity patterns as follows: 27/27 vancomycin, 13/13 clindamycin, 6/6 cefazolin, 11/11 ciprofloxacin, 14/14 moxifloxacin, 24/26 (92%) ofloxacin, 12/14 (86%) levofloxacin, 13/14 (93%) gatifloxacin, and 1/13 (8%) gentamicin. The organism was sensitive to at least one antibiotic administered initially in all cases. Final visual acuity was 20/400 or better in 8/27 (30%) cases, but 10 eyes (37%) had a final vision of no light perception. CONCLUSION: Despite prompt treatment with appropriate antibiotics, endophthalmitis caused by Streptococcus pneumoniae is associated with a poor visual prognosis. PMID- 15289133 TI - Treatment and long-term outcome of patients with orbital cavernomas. AB - PURPOSE: To evaluate the long-term prognosis of visual function in patients with orbital cavernomas after conservative and surgical treatment. DESIGN: Interventional case series. METHODS: The authors describe the outcome of 20 patients with cavernous hemangiomas of the orbit treated in their departments between 1988 and 2003. This prospective study included five cases followed by clinical and radiologic observation and 15 cases of symptomatic tumors that were completely removed by means of a frontotemporal or by means of a transconjunctival approach. The clinical characteristics of orbital cavernomas were analyzed together with their appropriate treatment. Furthermore, the authors present the unusual case of a patient suffering from progressive visual deterioration from a cerebral cavernoma compressing the optic nerve. RESULTS: The follow-up period was between 3 and 10 years. All orbital cavernomas in the group of conservatively managed patients remained stable. The transconjunctival excision of a medially located lesion was uncomplicated. The frontotemporal approach was chosen for large tumors situated in the proximity of the orbital apex and was associated with a higher number of complications. A good overall outcome of visual function and patient satisfaction was achieved in 11 of 14 cases operated on by craniotomy. CONCLUSION: The combination of clinical signs and magnetic resonance imaging (MRI) is highly sensitive and specific for the diagnosis of orbital cavernomas. In the presence of visual deterioration clearly attributable to the tumor we recommend immediate surgery, while lesions producing solely exophthalmos can safely be followed by observation. The transcranial approach offers excellent exposure and a rewarding cosmetic result and may be considered for large lesions superior and medial to the optic nerve, especially if they involve the orbital apex. PMID- 15289134 TI - Predictors of structural findings in old disciform lesions. AB - PURPOSE: To analyze the baseline findings predictive of the size and type of the late exudative age-related macular degeneration (AMD) lesions. DESIGN: Observational case series. METHODS: Retrospective study. SETTING: University clinic referral practice. STUDY POPULATION: The records of 167 consecutive patients initially diagnosed with recent choroidal neovascularization (CNV) related to AMD were analyzed 4.8 to 9.2 (mean 6.8) years after baseline. Of 121 patients still living, 74 attended the reexamination. After exclusions, data from 61 patients were analyzed. OBSERVATION PROCEDURES: From the fundus photographs and fluorescein angiographic images taken at baseline and at follow-up the size and components of AMD lesion were measured. The presence of hemorrhage, pigment epithelial detachment (PED), classic CNV, laser treatment, and involvement of the fellow eye were recorded at baseline. MAIN OUTCOME MEASURES: Lesion size, increase in lesion size, persistent exudative process, subretinal fibrosis, and chorioretinal anastomosis at the follow-up examination. RESULTS: Large lesion (P <.001) and CNV (P =.003) sizes, and the presence of occult, no classic CNV (P =.013) at baseline predict a large lesion at follow-up. Other factors predicting a large lesion were subfoveal (P =.019) and bilateral lesions (P =.017) and the presence of hemorrhage (P =.012) at baseline. A large (P =.040) and bilateral lesion (P =.040) and hemorrhage (P =.011) at baseline were correlated with subretinal fibrosis at follow-up. Occult lesions (P =.002) at baseline were correlated with chorioretinal anastomosis at follow-up. CONCLUSION: The type and size of the late AMD lesion can partly be predicted from baseline characteristics. PMID- 15289135 TI - Eye involvement in inherited epidermolysis bullosa: experience of the National Epidermolysis Bullosa Registry. AB - PURPOSE: To determine the frequency of ocular manifestations in inherited epidermolysis bullosa (EB) within the continental United States and to define the estimated cumulative risks of developing nonscarring (blisters or erosions) and scarring corneal manifestations within each major EB subtype over time. DESIGN: Observational (cross-sectional and longitudinal). METHODS: Up to 16 years of longitudinal follow-up was conducted on 3,280 consecutively enrolled patients in the National EB Registry, an epidemiologic study funded by the National Institutes of Health. Data were stratified by major EB type and subtype. Frequencies of occurrence were determined for eight variables (corneal erosions or blistering; corneal scarring; symblepharons; blepharitis; ectropions; lacrimal duct obstruction; impaired vision; blindness) by contingency tables, and cumulative risks were generated by life table analysis technique. RESULTS: The most common ocular manifestations were corneal erosions and blisters. Frequencies mirrored relative severity of skin disease, with 74.10% of all patients with recessive dystrophic EB, Hallopeau-Siemens (RDEB-HS) and 47.50% of all patients with junctional EB, Herlitz (JEB-H) experiencing at least one episode. Lower frequencies were noted for corneal scarring. Symblepharons and ectropions were most commonly seen in inversa RDEB and JEB-H, respectively. Blindness was reported in 6.47% of RDEB-HS patients. The cumulative risks of nonscarring and scarring corneal lesions in JEB-H at age 5 are 83.18% and 27.08% and at age 25 are 83.18% and 72.22%. With time, the cumulative risk of each in RDEB-HS approached that reported in JEB-H patients. CONCLUSION: Ocular disease activity, particularly corneal, is common in some EB subtypes. Careful ophthalmologic examination should become an integral part of the management of all patients with inherited EB. PMID- 15289136 TI - Photodynamic therapy with verteporfin for subfoveal choroidal neovascularization associated with multifocal choroiditis. AB - PURPOSE: Evaluation of visual acuity outcomes of photodynamic therapy with verteporfin for subfoveal choroidal neovascularization (CNV) secondary to multifocal choroiditis. DESIGN: Open-label, prospective, interventional case series. METHODS: Thirteen patients (13 eyes) diagnosed with subfoveal CNV associated with multifocal choroiditis at the Eye Clinics of Trieste and Udine were considered for the study. Inclusion criteria were the presence of subfoveal CNV no larger than 5,400 microm in greatest linear dimension and best-corrected visual acuity, Snellen equivalent, of approximately 20/400 or better. The primary outcome was the number of eyes that had fewer than 8 letters lost (less than approximately 1.5 lines) at the 12-month examination compared with the baseline examination. Secondary outcomes included fluorescein angiographic features such as progression and CNV size. RESULTS: Baseline and final best-corrected visual acuity were 0.52 logarithm of the minimal angle of resolution (20/62(-2) Snellen equivalent) and 0.55 logarithm of the minimal angle of resolution (20/62(-2) Snellen equivalent), respectively. By the 12-month visit, one patient (7.7%) had gained at least 1.5 lines, two patients (15.4%) had lost 1.5 or more lines, and no patient lost 3 or more lines of visual acuity, whereas 10 patients (84.6%) showed less than 1.5-line change. Mean CNV area was 0,69 mm(2) and 0.63 mm(2) at baseline and at the 12-month visit, respectively. By the month 12 examination, patients had received an average of 1.7 treatments. CONCLUSIONS: Photodynamic therapy may be considered a viable therapeutic option for subfoveal CNV associated with multifocal choroiditis at least for a 1-year period. Further studies with longer follow-up are needed to confirm these results. PMID- 15289137 TI - Expression of ephrinB2 and its receptors on fibroproliferative membranes in ocular angiogenic diseases. AB - PURPOSE: To determine whether ephrinB2 plays a role in ocular angiogenesis, we investigated the expression of ephrinB2 and EphB receptors on retinal fibroproliferative membranes. DESIGN: Experimental study of the expression of ephrinB2 and EphB receptors within fibroproliferative membranes in patients with ocular angiogenic diseases collected during vitrectomy. METHODS: Fibroproliferative membranes were obtained at the time of vitrectomy from 20 patients with proliferative diabetic retinopathy (PDR) and from 40 patients who had stage 5 retinopathy of prematurity. Specimens were investigated with immunohistochemistry using polyclonal antibodies directed against ephrinB2 and the EphB2, EphB3, and EphB4 receptors. Immunoreactivity for von Willebrand factor (factor VIII) and alpha-smooth muscle actin (alpha-SMA) was also determined to confirm the identity of the target vascular endothelial cells. RESULTS: Positive staining for ephrinB2 was observed on fibroproliferative membranes that were obtained from patients with PDR (65.0%) and retinopathy of prematurity (25.0%). Specifically, ephrinB2 was found to be present on endothelial cells, as confirmed by its colocalization with factor VIII and alpha-SMA staining. EphB2 and EphB3 expression was observed on fibroproliferative membranes that were harvested from patients with PDR (EphB2, 90.0%; EphB3, 70.0%) and retinopathy of prematurity (EphB2, 35.0%; EphB3, 45.0%). However, EphB4 expression was not observed in any of the membranes derived from patients with PDR or retinopathy of prematurity. The rate of ephrinB2 expression in patients with PDR was significantly higher than that seen in patients with retinopathy of prematurity, which probably reflected differences in the vascular density of their fibroproliferative membranes. CONCLUSION: These data suggest that the ephrinB2-EphB2/B3 system may play an important role in ocular angiogenesis. PMID- 15289138 TI - Determination of the incidence and clinical characteristics of subsequent retinal tears following treatment of the acute posterior vitreous detachment-related initial retinal tears. AB - PURPOSE: To determine the incidence of subsequent retinal tears (SRT) in patients who were previously treated for acute posterior vitreous detachment (PVD)-related retinal tears. DESIGN: Retrospective chart review. METHODS: A retrospective chart review of 155 eyes of 137 consecutive patients treated for acute PVD-related retinal tears was performed. RESULTS: During a median follow-up of 13 months (range 3 to 157 months) after treatment, SRT developed in 19 (12.2%) of 155 treated eyes. Most of the SRT occurred during the first 6 months (12 of 19 eyes) after initial treatment. Subsequent vetinal tears developed within the first 12 months in 15 of 19 treated eyes, however only six of 15 of these eyes were symptomatic. Of those patients who developed SRT after 12 months, all had new visual symptoms. CONCLUSION: Patients who present with acute PVD-related retinal tears are at a low but significant risk for developing SRT. A significant number of patients treated for PVD-associated retinal tears present within 1 year with SRT without symptoms. PMID- 15289139 TI - Dorzolamide-induced choroidal detachment in a surgically untreated eye. AB - PURPOSE: Choroidal detachment is a known complication of topical hypotensive agents when used to treat eyes sensitized by prior surgery. We document the abrupt development of an extensive choroidal detachment after initiation of dorzolamide therapy in a surgically untreated eye with primary open-angle glaucoma. DESIGN: Observational case report. METHODS: A 76-year-old woman with primary open-angle glaucoma and no history of ocular surgery developed a choroidal detachment 12 hours after initiation of therapy with dorzolamide eye drops. Choroidal detachment was diagnosed clinically and confirmed by echography. RESULTS: Withdrawal of the drug and initiation of corticosteroid drops resulted in prompt resolution of the choroidal detachment. CONCLUSIONS: Choroidal detachment can occur in surgically untreated eyes after use of a topical carbonic anhydrase inhibitor. PMID- 15289140 TI - Early rapid rise in intraocular pressure after intravitreal triamcinolone acetonide injection. AB - PURPOSE: To report the occurrences of early rapid increases in intraocular (IOP) after intravitreal glucocorticoid injection. DESIGN: Observational case series. METHODS: We retrospectively reviewed the records of three patients seen and treated at Duke Eye Center. RESULTS: In all three cases, a significant rise in IOP occurred within 1 week of intravitreal triamcinolone injection for refractory macular edema. In one patient, a white material was found in the angle on gonioscopy. All three cases required surgical intervention to reduce the IOP. CONCLUSIONS: Considering the early rapid rise in IOP in these three cases, we suggest that clinicians closely monitor patients after intravitreal triamcinolone injections for the development of acute glaucoma. Additionally, it may be advisable to perform gonioscopic examinations to look for any abnormal accumulation of material in the angle. PMID- 15289141 TI - Removal of retained subfoveal perfluoro-n-octane liquid. AB - PURPOSE: To report a novel technique to remove retained submacular perfluorocarbon liquid. DESIGN: Retrospective cases series. METHODS: Two patients with retained subfoveal perfluorocarbon liquid were treated with this technique. With a three-port pars plana approach, a 39-gauge flexible cannula was used to perform a retinotomy adjacent to the subfoveal perfluorocarbon bubble. The tip of the cannula was inserted into the bubble, which was removed with active suction. RESULTS: Both patients underwent successful removal of the perfluorocarbon liquid without complications. In one patient, visual acuity improved from 20/70 to 20/30, with resolution of his preoperative central scotoma. The second patient required removal of subfoveal perfluorocarbon liquid at the time of silicone oil removal. The preoperative vision of finger counting improved to 20/60. CONCLUSIONS: Subretinal perfluorocarbon can be removed using a flexible 39-gauge cannula without inducing retinal detachment or making a large retinotomy. This technique may benefit patients with retained subfoveal perfluorocarbon liquid. PMID- 15289142 TI - Intravitreal triamcinolone acetonide for the treatment of cystoid macular edema secondary to Behcet disease. AB - PURPOSE: To evaluate the safety and effectiveness of intravitreal triamcinolone acetonide in patients with cystoid macular edema (CME) associated with Behcet disease. DESIGN: Interventional case series. METHODS: Ten eyes of 10 patients with CME from Behcet disease made up the study population. All eyes had persistent CME despite medical treatment for at least 2 months. Intravitreal injection of 4 mg (0.1 ml) of triamcinolone acetonide was offered to treat macular edema. The visual and anatomic responses were observed. RESULTS: At 1 month follow-up, a reduction in mean foveal thickness of 37.4%, from 416 microm to 260.5 microm, was attained. At 3-month follow-up, mean foveal thickness was 286.2 microm and at 6 months, 263.7 microm. No eyes had lost vision at 1 month, and eight eyes (80%) showed improvement in visual acuity. At 3-month and 6-month follow-up, three eyes (30%) remained stable and the other eyes had maintained the improved acuity. CONCLUSION: Intravitreal triamcinolone acetonide is a promising therapeutic method for CME from Behcet disease. PMID- 15289143 TI - Bilateral lid margin ulcers as the initial manifestation of Crohn disease. AB - PURPOSE: To report an unusual bilateral ulcerative lid involvement as the presenting manifestation of a severe Crohn disease. DESIGN: Observational case report. METHOD: Description of an otherwise healthy woman who initially presented with bilateral ulcerative lid involvement before the discovery of an extensive ulcerative intestinal inflammatory disease. RESULTS: A 32-year-old woman presented with bilateral ulcerative blepharitis. She also complained of aphthous oral lesions and diarrheic episodes for the previous 3 days. Impression cytology of the lid ulcers showed conjunctival cells, together with the presence of lymphocytes and macrophages. Colonoscopy and colonic biopsy were characteristic of Crohn disease. The treatment with systemic corticosteroids healed bowel, oral, and lid margin ulcerative lesions. CONCLUSION: The simultaneous appearance of ulcerative lesions in the intestinal mucosa and in the mucocutaneous lid margin and the comparable features encountered in the cytologic studies indicate that ulcerative lid margin disease could be an ocular manifestations of Crohn disease. PMID- 15289144 TI - Adult-onset foveomacular vitelliform dystrophy with OCT 3. AB - PURPOSE: To report the morphologic data of adult-onset foveomacular vitelliform dystrophy (AFVD) provided by third-generation optical coherence tomography (OCT 3). DESIGN: Observational case report. METHODS: An 85-year-old woman presenting with AFVD underwent fundus biomicroscopy, fluorescein angiography, and OCT examination. RESULTS: Fundus examination disclosed a round, elevated yellowish lesion, centered by a pigmented spot, in the macula of the left eye. Examination with OCT disclosed an area of hyperreflectivity located between the retinal pigment epithelium layer and the photoreceptor layer, compatible in size with the yellowish elevated lesion. The hyperreflective line corresponding to the photoreceptor layer was elevated by the material and separated from the retinal pigment epithelium layer. CONCLUSIONS: In AFVD, examination with OCT 3 demonstrates elevation of the photoreceptor layer by the material and supports the previous hypothesis that the material is located between the photoreceptor and the retinal pigment epithelium layer. PMID- 15289145 TI - Decrease of blue cone sensitivity in acute idiopathic blind spot enlargement syndrome. AB - PURPOSE: To determine the sensitivity of the blue cone system by static perimetry in patients with unilateral acute idiopathic blind spot enlargement (AIBSE) syndrome. DESIGN: Observational case series. METHODS: Four patients with AIBSE syndrome, aged 16 to 30 years, were studied. Diagnosis of the AIBSE syndrome was based on clinical findings including an enlarged blind spot without corresponding ophthalmoscopic changes of the ocular fundus and with depressed multifocal electoretinograms. The visual sensitivity and mean deviations (MD) were measured by white-on-white (W/W) and blue-on-yellow (B/Y) automated perimetry. RESULTS: The average difference in the MD between the affected and unaffected eyes was 4.87 +/- 1.51 dB and 13.65 +/- 4.19 dB for W/W and B/Y perimetries, respectively. The difference between the two groups of eyes was significantly greater for B/Y perimetry than that for W/W perimetry (P =.01). CONCLUSIONS: Our results suggest that the decrease of blue cone sensitivity is diffusely present over the retina in eyes with AIBSE syndrome. We recommend B/Y perimetry as a sensitive measure to detect retinal dysfunction that is not detected by W/W perimetry. PMID- 15289146 TI - Prevention of ligneous conjunctivitis by topical and subconjunctival fresh frozen plasma. AB - PURPOSE: To present a case of ligneous conjunctivitis where the recurrence of membranous conjunctivitis was prevented by subconjunctival and topical instillation of fresh frozen plasma. DESIGN: Interventional case report. METHODS: A case of ligneous conjunctivitis with multiple recurrences since the age of 3 years developed recurrent membranous conjunctivitis after transconjunctival levator recession. Blood plasminogen activity was determined. The membrane was excised, and the membrane reappeared 4 days later. The patient was treated with excision of the membrane and subconjunctival injection of fresh frozen plasma and topical fresh frozen plasma. Plasminogen activity of the fresh frozen plasma was normal. RESULTS: Plasminogen blood functional activity was 52% (normal is 80% 120%). The patient had complete remission with no recurrences of membranous conjunctivitis after topical and subconjunctival fresh frozen plasma. CONCLUSIONS: Prophylactic use of topical and subconjunctival fresh frozen plasma may help in the prevention of membranes in susceptible patients with plasminogen deficiency. PMID- 15289147 TI - Group B streptococcus endogenous endophthalmitis presenting as septic arthritis and a homonymous hemianopsia due to embolic stroke. AB - PURPOSE: To report a case of group B streptococcus (GBS) endogenous endophthalmitis in a patient presenting with septic arthritis and a homonymous hemianopsia due to embolic stroke. DESIGN: Observational case report. METHODS: A 75-year-old woman with septic arthritis and a homonymous hemianopsia due to embolic stroke was examined and found to have endogenous GBS endophthalmitis due to bacterial endocarditis. RESULTS: Magnetic resonance imaging showed many septic emboli to the brain, including the left occipital lobe. Fundus examination showed evidence for endogenous endophthalmitis. Blood and urine cultures were positive for GBS. A mitral valve vegetation was presumed to be the cause of the intracranial septic emboli and was demonstrated on transesophageal echocardiography. CONCLUSIONS: Septic arthritis and embolic stroke are suggestive of endocarditis and sepsis. Endogenous endophthalmitis in this case led to transesophageal echocardiography, which was diagnostic for endocarditis. PMID- 15289148 TI - Histopathological abnormalities in ocular blood vessels of CADASIL patients. AB - PURPOSE: To assess histopathological findings in patients with cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL). DESIGN: Case reports and histopathological evaluation of enucleated eyes. METHODS: Four eyes from two CADASIL patients were enucleated at autopsy and prepared for histopathological analysis using light and electron microscopy. RESULTS: Thickening of arterial walls with fibrosis, eosinophilic Periodic acid Schiff-positive basement membrane material and loss of vascular smooth muscle cells (VSMC) in the central retinal artery and its branches, the leptomeninges, the ocular adnexa, and the optic disk were observed. On electron microscopy, numerous deposits of granular, osmiophilic material in arterial walls as well as VSMC and pericyte degeneration were noted. In contrast to retinal vessels, the choroid was not affected. CONCLUSION: Our findings suggest a differential involvement of small blood vessels in CADASIL, depending on the angioarchitecture and support autopsy data of nervous tissue describing that loss of VSMCs is most pronounced in tissues depending on blood-tissue barriers. PMID- 15289149 TI - Latanoprost exposure in pregnancy. AB - PURPOSE: To observe pregnancies exposed to latanoprost, a prostaglandin analog administered in the treatment of glaucoma. Its prescription is limited in pregnancy, because reproduction studies in animals report a high incidence of abortion and human investigations are not adequate. As a consequence it is classified as category C drug according to the United States Food and Drug Administration's use-in-pregnancy ratings. DESIGN: Observational study. METHODS: We collected data, referred to our Teratology Information Service, relative to latanoprost exposure in pregnancy. We followed by phone interviews women treated with latanoprost during the first trimester, and we evaluated whether there had been any adverse effects on the fetus. RESULTS: Eleven cases of latanoprost exposure in pregnancy were referred to our Teratology Information Service. One case was lost to follow-up, and one case was complicated by miscarriage. Nine cases had a complete follow-up without congenital anomalies. CONCLUSIONS: Our series is too small to perform statistical significance; however, we found no evidence of adverse effects of latanoprost on pregnancy or neonatal outcomes. PMID- 15289150 TI - Bilaterally identical monoclonality in a case of primary intraocular lymphoma. AB - PURPOSE: To report a case of primary intraocular lymphoma possessing bilaterally identical monoclonal rearrangements of the immunoglobulin heavy chain (IgH) gene. DESIGN: Observational case report. METHODS: A 78-year-old woman with bilateral vitritis, subretinal infiltrates in the right eye, and no involvement of the central nervous system (CNS) was diagnosed histologically with diffuse large B cell lymphoma by a transvitreal subretinal biopsy of the right eye. One month later, vitrectomy was performed on the left eye due to increased vitreous opacity. DNA was extracted from a vitrectomy specimen from each eye, and the third complementarity-determining region of IgH gene was analyzed by polymerase chain reaction and direct sequencing. RESULTS: Monoclonal rearrangements of IgH gene with identical DNA sequence were detected in both eyes. CONCLUSIONS: Bilaterally identical monoclonality was detected in a case of primary intraocular lymphoma with no CNS involvement. PMID- 15289151 TI - Irregularity of photoreceptor layer after successful macular hole surgery prevents visual acuity improvement. AB - PURPOSE: To investigate the association between a photoreceptor irregularity and visual acuity (VA) after macular hole (MH) surgery. DESIGN: Prospective observational case series. METHODS: Twenty-four eyes with an idiopathic MH that resolved after vitreous surgery were examined by new optical coherence tomography (OCT 3000). The images were divided into regular photoreceptor, detected as a straight line above the retinal pigment epithelium reflex, and irregular, which cannot be detected by it. RESULTS: Regular images were observed in 12 eyes (80%) and irregular images in 3 eyes (20%) with good VA (>or=0.7). Regular images were seen in 3 eyes (33%) and irregular images in 6 eyes (67%) with poor VA (<0.7). The percentage of regular images in the group with good VA was significantly higher than that in the group with poor VA (P <.05). CONCLUSION: A photoreceptor irregularity after MH surgery may prevent VA improvement. PMID- 15289152 TI - Conjunctival hyperemia associated with bimatoprost use: a histopathologic study. AB - PURPOSE: To evaluate histopathologic signs of conjunctival inflammation in patients with conjunctival hyperemia induced by bimatoprost treatment. DESIGN: Prospective interventional study. METHODS: The study included 15 eyes of 15 patients scheduled for cataract surgery. Patients in the treatment group (n = 9) exhibited trace to moderate conjunctival hyperemia when treated with bimatoprost 0.03% every day for 15 to 30 days before surgery. The control group (n = 6) included untreated patients with no ocular disease other than cataract. Conjunctival biopsies were obtained for histologic evaluation with light microscopy. RESULTS: Vascular congestion was observed in biopsies from 7 patients (78%) in the bimatoprost group and 5 patients (83%) in the control group. Signs of inflammation were found in biopsies from 2 patients (22%) in the bimatoprost group and 2 patients (33%) in the control group. CONCLUSIONS: Histopathologic signs of inflammation were no more frequent in conjunctival specimens from bimatoprost-treated patients with trace to moderate hyperemia than in those from untreated control subjects. PMID- 15289153 TI - Herpes simplex virus-1--associated congenital cataract. AB - PURPOSE: To report association of herpes simplex virus-type 1 (HSV-1) in four cases of congenital cataract. DESIGN: Prospective interventional case series. METHODS: Four infants younger than 12 months, presenting with unilateral or bilateral congenital cataract, were included. The cases were clinically evaluated by the pediatric ophthalmologist. The lens aspirates collected at the time of cataract surgery were processed for HSV-1 culture in rabbit corneal epithelial (SIRC) cell line and for HSV-1 DNA by polymerase chain (PCR). The sera of the children and the mother were tested for HSV-1 immunoglobulin (Ig) M and IgG by enzyme linked immunosorbent assay (ELISA). RESULTS: HSV-1 was isolated in tube cultures in three of four lens aspirates, and all four lens aspirates were positive for HSV-1 DNA by PCR. Serum HSV-1 IgM was positive in all babies and in three cases HSV-1 IgM was positive in the mother's serum. CONCLUSION: Based on a computerized literature search, we believe this may be the first report of HSV-1 associated congenital cataract. PMID- 15289154 TI - In vivo observation of internal limiting membrane in an eye with macular hole. PMID- 15289156 TI - The need to decide if all estrogens are intrinsically similar. AB - We used gene expression profiling to investigate whether the molecular effects induced by estrogens of different provenance are intrinsically similar. In this article we show that the physiologic estrogen 17-beta-estradiol, the phytoestrogen genistein, and the synthetic estrogen diethylstilbestrol alter the expression of the same 179 genes in the intact immature mouse uterus under conditions where each chemical has produced an equivalent gravimetric and histologic uterotrophic effect, using the standard 3-day assay protocol. Data are also presented indicating the limitations associated with comparison of gene expression profiles for different chemicals at times before the uterotrophic effects are fully realized. We conclude that the case has yet to be made for regarding synthetic estrogens as presenting a unique human hazard compared with phytoestrogens and physiologic estrogens. Key words: diethylstilbestrol, estrogen, gene expression, genistein, microarray, phytoestrogen, toxicogenomics, uterus. PMID- 15289157 TI - Using human disease outbreaks as a guide to multilevel ecosystem interventions. AB - Human health often depends on environmental variables and is generally subject to widespread and comprehensive surveillance. Compared with other available measures of ecosystem health, human disease incidence may be one of the most useful and practical bioindicators for the often elusive gauge of ecologic well-being. We argue that many subtle ecosystem disruptions are often identified only as a result of detailed epidemiologic investigations after an anomalous increase in human disease incidence detected by routine surveillance mechanisms. Incidence rates for vector-mediated diseases (e.g., arboviral illnesses) and direct zoonoses (e.g., hantaviruses) are particularly appropriate as bioindicators to identify underlying ecosystem disturbances. Outbreak data not only have the potential to act as a pivotal warning system for ecosystem disruption, but may also be used to identify interventions for the preservation of ecologic health. With this approach, appropriate ecologically based strategies for remediation can be introduced at an earlier stage than would be possible based solely on environmental monitoring, thereby reducing the level of "ecosystem distress" as well as resultant disease burden in humans. This concept is discussed using local, regional, and global examples, thereby introducing the concept of multilevel ecosystem interventions. Key words: bioindicators, disease control, disease outbreaks, ecologic management, ecosystem health, surveillance. PMID- 15289158 TI - Determinants of bone and blood lead levels among minorities living in the Boston area. AB - We measured blood and bone lead levels among minority individuals who live in some of Boston's neighborhoods with high minority representation. Compared with samples of predominantly white subjects we had studied before, the 84 volunteers in this study (33:67 male:female ratio; 31-72 years of age) had similar educational, occupational, and smoking profiles and mean blood, tibia, and patella lead levels (3 microg/dL, 11.9 microg/g, and 14.2 microg/g, respectively) that were also similar. The slopes of the univariate regressions of blood, tibia, and patella lead versus age were 0.10 microg/dL/year (p < 0.001), 0.45 microg/g/year (p < 0.001), and 0.73 microg/g/year (p < 0.001), respectively. Analyses of smoothing curves and regression lines for tibia and patella lead suggested an inflection point at 55 years of age, with slopes for subjects greater than or equal to 55 years of age that were not only steeper than those of younger subjects but also substantially steeper than those observed for individuals > 55 years of age in studies of predominantly white participants. This apparent racial disparity at older ages may be related to differences in historic occupational and/or environmental exposures, or possibly the lower rates of bone turnover that are known to occur in postmenopausal black women. The higher levels of lead accumulation seen in this age group are of concern because such levels have been shown in other studies to predict elevated risks of chronic disease such as hypertension and cognitive dysfunction. Additional research on bone lead levels in minorities and their socioeconomic and racial determinants is needed. PMID- 15289159 TI - Age-related differences in susceptibility to carcinogenesis: a quantitative analysis of empirical animal bioassay data. AB - In revising cancer risk assessment guidelines, the U.S. Environmental Protection Agency (EPA) analyzed animal cancer bioassay data over different periods of life. In this article, we report an improved analysis of these data (supplemented with some chemical carcinogenesis observations not included in the U.S. EPA's original analysis) and animal bioassay studies of ionizing radiation. We use likelihood methods to avoid excluding cases where no tumors were observed in specific groups. We express dosage for animals of different weights on a metabolically consistent basis (concentration in air or food, or per unit body weight to the three-quarters power). Finally, we use a system of dummy variables to represent exposures during fetal, preweaning, and weaning-60-day postnatal periods, yielding separate estimates of relative sensitivity per day of dosing in these intervals. Central estimate results indicate a 5- to 60-fold increased carcinogenic sensitivity in the birth-weaning period per dose divided by(body weight(0.75)-day) for mutagenic carcinogens and a somewhat smaller increase- centered about 5-fold--for radiation carcinogenesis per gray. Effects were greater in males than in females. We found a similar increased sensitivity in the fetal period for direct-acting nitrosoureas, but no such increased fetal sensitivity was detected for carcinogens requiring metabolic activation. For the birth-weaning period, we found an increased sensitivity for direct administration to the pups similar to that found for indirect exposure via lactation. Radiation experiments indicated that carcinogenic sensitivity is not constant through the "adult" period, but the dosage delivered in 12- to 21-month-old animals appears a few-fold less effective than the comparable dosage delivered in young adults (90 105 days of age). PMID- 15289160 TI - Behavioral alterations in response to fear-provoking stimuli and tranylcypromine induced by perinatal exposure to bisphenol A and nonylphenol in male rats. AB - The purpose of this study was to examine whether perinatal exposure to two major environmental endocrine-disrupting chemicals, bisphenol A (BPA; 0.1 mg/kg/day orally) and nonylphenol [NP; 0.1 mg/kg/day (low dose) and 10 mg/kg/day (high dose) orally] daily from gestational day 3 to postnatal day 20 (transplacental and lactational exposures) would lead to behavioral alterations in the male offspring of F344 rats. Neither BPA nor NP exposure affected behavioral characteristics in an open-field test (8 weeks of age), in a measurement of spontaneous motor activity (12 weeks of age), or in an elevated plus-maze test (14 weeks of age). A passive avoidance test (13 weeks of age) showed that both BPA- and NP-treated offspring tended to delay entry into a dark compartment. An active avoidance test at 15 weeks of age revealed that BPA-treated offspring showed significantly fewer avoidance responses and low-dose NP-treated offspring exhibited slightly fewer avoidance responses. Furthermore, BPA-treated offspring significantly increased the number of failures to avoid electrical unconditioned stimuli within 5-sec electrical shock presentation compared with the control offspring. In a monoamine-disruption test using 5 mg/kg (intraperitoneal) tranylcypromine (Tcy), a monoamine oxidase inhibitor, both BPA-treated and low dose NP-treated offspring at 22-24 weeks of age failed to show a significant increment in locomotion in response to Tcy, whereas control and high-dose NP treated offspring significantly increased locomotion behavior after Tcy injection. In addition, when only saline was injected during a monoamine disruption test, low-dose NP-treated offspring showed frequent rearing compared with the control offspring. The present results indicate that perinatal low-dose BPA or NP exposure irreversibly influenced the reception of fear-provoking stimuli (e.g., electrical shock), as well as monoaminergic neural pathways. Key words: behavior, bisphenol A, fear, learning, monoamine, nonylphenol. PMID- 15289161 TI - Hair mercury levels in U.S. children and women of childbearing age: reference range data from NHANES 1999-2000. AB - Exposure to methyl mercury, a risk factor for neurodevelopmental toxicity, was assessed in U.S. children 1-5 years of age (n = 838) and women 16-49 years of age (n = 1,726) using hair mercury analysis during the 1999-2000 National Health and Nutrition Examination Survey (NHANES). The data are nationally representative and are based on analysis of cross-sectional data for the noninstitutionalized, U.S. household population. The survey consisted of interviews conducted in participants' homes and standardized health examinations conducted in mobile examination centers. Distributions of total hair mercury levels expressed as micrograms per gram hair Hg and the association of hair Hg levels with sociodemographic characteristics and fish consumption are reported. Geometric mean (standard error of the geometric mean) hair mercury was 0.12 microg/g (0.01 microg/g) in children, and 0.20 microg/g (0.02 microg/g) in women. Among frequent fish consumers, geometric mean hair mercury levels were 3-fold higher for women (0.38 vs. 0.11 micro g/g) and 2-fold higher for children (0.16 vs. 0.08 microg/g) compared with nonconsumers. The NHANES 1999-2000 data provide population-based data on hair mercury concentrations for women and children in the United States. Hair mercury levels were associated with age and fish consumption frequency. PMID- 15289162 TI - The effect of arsenic mitigation interventions on disease burden in Bangladesh. AB - Many interventions have been advocated to mitigate the impact of arsenic contamination of drinking water in Bangladesh. However, there are few data on the true magnitude of arsenic-related disease in Bangladesh nationally. There has also been little consideration given to possible adverse effects of such interventions, in particular, diarrheal disease. The purpose of this study was to estimate and compare the likely impacts of arsenic mitigation interventions on both arsenic-related disease and water-borne infectious disease. We found that arsenic-related disease currently results in 9,136 deaths per year and 174,174 disability-adjusted life years (DALYs; undiscounted) lost per year in those exposed to arsenic concentrations > 50 microg/L. This constitutes 0.3% of the total disease burden in Bangladesh in terms of undiscounted DALYs. We found intervention to be of overall benefit in reducing disease burden in most scenarios examined, but the concomitant increase in water-related infectious disease significantly reduced the potential benefits gained from intervention. A minimum reduction in arsenic-related DALYs of 77% was necessary before intervention achieved any reduction in net disease burden. This is assuming that interventions were provided to those exposed to > 50 microg/L and would concomitantly result in a 20% increase in water-related infectious disease in those without access to adequate sanitation. Intervention appears to be justified for those populations exposed to high levels of arsenic, but it must be based on exposure levels and on the effectiveness of interventions not only in reducing arsenic but in minimizing risk of water-related infections. Key words: arsenic/adverse effects, Bangladesh, burden of disease, diarrhea, risk assessment, water pollutants, water supply. PMID- 15289163 TI - Lead, diabetes, hypertension, and renal function: the normative aging study. AB - In this prospective study, we examined changes in renal function during 6 years of follow-up in relation to baseline lead levels, diabetes, and hypertension among 448 middle-age and elderly men, a subsample of the Normative Aging Study. Lead levels were generally low at baseline, with mean blood lead, patella lead, and tibia lead values of 6.5 microg/dL, 32.4 microg/g, and 21.5 microg/g, respectively. Six percent and 26% of subjects had diabetes and hypertension at baseline, respectively. In multivariate-adjusted regression analyses, longitudinal increases in serum creatinine (SCr) were associated with higher baseline lead levels but these associations were not statistically significant. However, we observed significant interactions of blood lead and tibia lead with diabetes in predicting annual change in SCr. For example, increasing the tibia lead level from the midpoints of the lowest to the highest quartiles (9-34 microg/g) was associated with an increase in the rate of rise in SCr that was 17.6-fold greater in diabetics than in nondiabetics (1.08 mg/dL/10 years vs. 0.062 mg/dL/10 years; p < 0.01). We also observed significant interactions of blood lead and tibia lead with diabetes in relation to baseline SCr levels (tibia lead only) and follow-up SCr levels. A significant interaction of tibia lead with hypertensive status in predicting annual change in SCr was also observed. We conclude that longitudinal decline of renal function among middle-age and elderly individuals appears to depend on both long-term lead stores and circulating lead, with an effect that is most pronounced among diabetics and hypertensives, subjects who likely represent particularly susceptible groups. PMID- 15289164 TI - Pesticide spraying for West Nile virus control and emergency department asthma visits in New York City, 2000. AB - Pyrethroid pesticides were applied via ground spraying to residential neighborhoods in New York City during July-September 2000 to control mosquito vectors of West Nile virus (WNV). Case reports link pyrethroid exposure to asthma exacerbations, but population-level effects on asthma from large-scale mosquito control programs have not been assessed. We conducted this analysis to determine whether widespread urban pyrethroid pesticide use was associated with increased rates of emergency department (ED) visits for asthma. We recorded the dates and locations of pyrethroid spraying during the 2000 WNV season in New York City and tabulated all ED visits for asthma to public hospitals from October 1999 through November 2000 by date and ZIP code of patients' residences. The association between pesticide application and asthma-related emergency visits was evaluated across date and ZIP code, adjusting for season, day of week, and daily temperature, precipitation, particulate, and ozone levels. There were 62,827 ED visits for asthma during the 14-month study period, across 162 ZIP codes. The number of asthma visits was similar in the 3-day periods before and after spraying (510 vs. 501, p = 0.78). In multivariate analyses, daily rates of asthma visits were not associated with pesticide spraying (rate ratio = 0.92; 95% confidence interval, 0.80-1.07). Secondary analyses among children and for chronic obstructive pulmonary disease yielded similar null results. This analysis shows that spraying pyrethroids for WNV control in New York City was not followed by population-level increases in public hospital ED visit rates for asthma. PMID- 15289165 TI - Increased risk of hepatocellular carcinoma and liver cirrhosis in vinyl chloride workers: synergistic effect of occupational exposure with alcohol intake. AB - Hepatocellular carcinoma (HCC) and liver cirrhosis (LC) are not well-established vinyl chloride monomer (VCM)-induced diseases. Our aim was to appraise the role of VCM, alcohol intake, and viral hepatitis infection, and their interactions, in the etiology of HCC and LC. Thirteen cases of HCC and 40 cases of LC were separately compared with 139 referents without chronic liver diseases or cancer in a case-referent study nested in a cohort of 1,658 VCM workers. The odds ratios (ORs) and the 95% confidence intervals (CIs) were estimated by common methods and by fitting models of logistic regression. We used Rothman's synergy index (S) to evaluate interactions. By holding the confounding factors constant at logistic regression analysis, each extra increase of 1,000 ppm times years of VCM cumulative exposure was found to increase the risk of HCC by 71% (OR = 1.71; 95% CI, 1.28-2.44) and the risk of LC by 37% (OR = 1.37; 95% CI, 1.13-1.69). The joint effect of VCM exposure above 2,500 ppm times years and alcohol intake above 60 g/day resulted in ORs of 409 (95% CI, 19.6-8,553) for HCC and 752 (95% CI, 55.3-10,248) for LC; both S indexes suggested a synergistic effect. The joint effect of VCM exposure above 2,500 ppm times years and viral hepatitis infection was 210 (95% CI, 7.13-6,203) for HCC and 80.5 (95% CI, 3.67-1,763) for LC; both S indexes suggested an additive effect. In conclusion, according to our findings, VCM exposure appears to be an independent risk factor for HCC and LC interacting synergistically with alcohol consumption and additively with viral hepatitis infection. PMID- 15289166 TI - The relationship between levels of PCBs and pesticides in human hair and blood: preliminary result. AB - Human hair as a biologic measure of exposure to persistent organic pollutants (POPs) has some advantages over the more commonly used blood and adipose tissue samples. However, one of the primary limitations is the difficulty in distinguishing between exogenous and endogenous contamination. In addition, there are currently no standardized methods for hair sample collection, washing, and chemical analysis. There is also very limited information describing the correlation between levels of organic contaminants in hair and other body compartments. To explore levels of POPs in blood and hair, samples from 10 volunteers were collected and analyzed for select organochlorine pesticides and 57 individual polychlorinated biphenyl (PCB) congeners. We demonstrated that the method for analyzing organic contaminants in human hair was reliable and reproducible. Washing hair with shampoo decreased levels of PCBs, pesticides, and lipids by 25-33% on average and up to 62% for low-chlorinated congeners. The percentage of lipids and the levels of organochlorines in hair were higher than in serum. We found strong correlation (r = 0.8) between p,p -DDE (dichlorodiphenyldichloroethylene) levels in hair and blood and moderate correlations for the more persistent PCB congeners, but no correlations or weak correlations for other organochlorines. The present study provides preliminary evidence on the utility of hair analysis for POPs; however, further larger studies are recommended before hair analysis can be successfully applied in epidemiologic studies on POPs. PMID- 15289167 TI - The association between environmental lead exposure and bone density in children. AB - Osteoporosis is a decrease in bone mineral density (BMD) that predisposes individuals to fractures. Although an elderly affliction, a predisposition may develop during adolescence if a sufficient peak BMD is not achieved. Rat studies have found that lead exposure is associated with decreased BMD. However, human studies are limited. We hypothesized that the BMD of children with high lead exposure would be lower than the BMD of children with low lead exposure. We collected data on 35 subjects; 16 had low cumulative lead exposure (mean, 6.5 microg/dL), and 19 had high exposure (mean, 23.6 micro g/dL). All were African American; there was no difference between the groups by sex, age, body mass index, socioeconomic status, physical activity, or calcium intake. Significant differences in BMD between low and high cumulative lead exposure were noted in the head (1.589 vs. 1.721 g/cm2), third lumbar vertebra (0.761 vs. 0.819 g/cm2), and fourth lumbar vertebra (0.712 vs. 0.789 g/cm2). Contrary to our hypothesis, subjects with high lead exposure had a significantly higher BMD than did subjects with low lead exposure. This may reflect a true phenomenon because lead exposure has been reported to accelerate bony maturation by inhibiting the effects of parathyroid hormone-related peptide. Accelerated maturation of bone may ultimately result in a lower peak BMD being achieved in young adulthood, thus predisposing to osteoporosis in later life. Future studies need to investigate this proposed model. PMID- 15289168 TI - Perfluorooctane sulfonate (PFOS) and related perfluorinated compounds in human maternal and cord blood samples: assessment of PFOS exposure in a susceptible population during pregnancy. AB - Fluorinated organic compounds (FOCs), such as perfluorooctane sulfonate (PFOS), perfluoro-octanoate (PFOA), and perfluorooctane sulfonylamide (PFOSA), are widely used in the manufacture of plastic, electronics, textile, and construction material in the apparel, leather, and upholstery industries. FOCs have been detected in human blood samples. Studies have indicated that FOCs may be detrimental to rodent development possibly by affecting thyroid hormone levels. In the present study, we determined the concentrations of FOCs in maternal and cord blood samples. Pregnant women 17-37 years of age were enrolled as subjects. FOCs in 15 pairs of maternal and cord blood samples were analyzed by liquid chromatography-electrospray mass spectrometry coupled with online extraction. The limits of quantification of PFOS, PFOA, and PFOSA in human plasma or serum were 0.5, 0.5, and 1.0 ng/mL, respectively. The method enables the precise determination of FOCs and can be applied to the detection of FOCs in human blood samples for monitoring human exposure. PFOS concentrations in maternal samples ranged from 4.9 to 17.6 ng/mL, whereas those in fetal samples ranged from 1.6 to 5.3 ng/mL. In contrast, PFOSA was not detected in fetal or maternal samples, whereas PFOA was detected only in maternal samples (range, < 0.5 to 2.3 ng/mL, 4 of 15). Our results revealed a high correlation between PFOS concentrations in maternal and cord blood (r2 = 0.876). However, we did not find any significant correlations between PFOS concentration in maternal and cord blood samples and age bracket, birth weight, or levels of thyroid-stimulating hormone or free thyroxine. Our study revealed that human fetuses in Japan may be exposed to relatively high levels of FOCs. Further investigation is required to determine the postnatal effects of fetal exposure to FOCs. Key words: cord blood, fluorinated organic compounds, human, PFOA, PFOS, PFOSA, pregnancy. PMID- 15289169 TI - Fetal exposure to PCBs and their hydroxylated metabolites in a Dutch cohort. AB - Polychlorinated biphenyls (PCBs) are still the most abundant pollutants in wildlife and humans. Hydroxylated PCB metabolites (OH-PCBs) are known to be formed in humans and wildlife. Studies in animals show that these metabolites cause endocrine-related toxicity. The health effects in humans have not yet been evaluated, especially the effect on the fetus and newborn. The aim of this study is to measure the levels of PCBs and OH-PCBs in maternal and cord blood samples in a population with background levels of PCBs. We analyzed 51 maternal and corresponding cord blood samples in the northern part of the Netherlands. The PCB concentrations in maternal plasma ranged from 2 to 293 ng/g lipid, and OH-PCB concentrations from nondetectable (ND) to 0.62 ng/g fresh weight. In cord plasma, PCB concentrations were 1-277 ng/g lipid, and OH-PCB concentrations, ND to 0.47 ng/g fresh weight. The cord versus maternal blood calculated ratio was 1.28 +/- 0.56 for PCBs and 2.11 +/- 1.33 for OH-PCBs, expressed per gram of lipid. When expressed per gram fresh weight, the ratios are 0.32 +/- 0.15 and 0.53 +/- 0.23 for PCBs and OH-PCBs, respectively. A significant correlation between the respective maternal and cord levels for both PCBs and OH-PCBs was found. Our results indicate that OH-PCBs and PCBs are transferred across the placenta to the fetus in concentrations resulting in levels of approximately 50 and 30%, respectively, of those in maternal plasma. More research in humans is needed to evaluate potential negative effects of these endocrine disruptors on the fetus. PMID- 15289170 TI - Drinking water contaminants, gene polymorphisms, and fetal growth. AB - There are still many uncertainties regarding the risk of adverse pregnancy outcomes associated with exposure to drinking water disinfection by-products. In Montreal, Quebec, Canada, we carried out a hospital-based case-control study including 493 cases of intrauterine growth restriction defined as birth weight below the 10th percentile for gestational age and sex, according to Canadian standards. Controls were babies (n = 472) delivered at the same hospital whose birth weight was at or above the 10th percentile, matched for gestational age, race, and sex. Exposure to total and specific trihalomethanes was measured using regulatory data collected by municipalities and the provincial Ministry of Environment. Residential history, water drinking, and shower habits during pregnancy, as well as known risk factors for intrauterine growth restriction, were measured with a face-to-face interview with all mothers. Mothers and newborns were characterized for two genetic polymorphisms, one in the CYP2E1 gene (G1259C), and another in the 5,10-methylenetetrahydrofolate reductase (MTHFR) gene (C677T). Exposure to specific and total trihalomethanes from drinking water, determined for 458 cases and 426 controls, did not result in an increased risk of intrauterine growth restriction. However, significant effect modification was observed between newborns with and without the CYP2E1 variant; among newborns with the variant, the adjusted odds ratio for intrauterine growth restriction associated with exposure to average total trihalomethanes above the 90th percentile (corresponding to 29.4 microg/L) was 13.20 (95% confidence interval, 1.19-146.72). These findings suggest that exposure to trihalomethanes at the highest levels can affect fetal growth but only in genetically susceptible newborns. PMID- 15289171 TI - Olden times: looking back on a career at the NIEHS. PMID- 15289172 TI - Impacts of our built environment on public health. PMID- 15289173 TI - Studying human fertility and environmental exposures. PMID- 15289174 TI - Studying human fertility. PMID- 15289176 TI - The WTC disaster and asbestos regulations. PMID- 15289177 TI - Trichloroethylene and cardiac malformations. PMID- 15289180 TI - Mother's thyroid, baby's health. PMID- 15289181 TI - Fighting obesity through the built environment. PMID- 15289182 TI - Sprawl: the new manifest destiny? PMID- 15289183 TI - Vehicular manslaughter: the global epidemic of traffic deaths. AB - The number of motorized vehicles is climbing exponentially in developing countries, and so is the number of people killed in accidents involving those vehicles. Traffic crashes in poorer nations tend to be fatal more often than those in developed countries because they often involve pedestrians and riders in less protected vehicles such as rickshaws, bicycles, or motorcycles. Such accidents are now a leading cause of death in the developing world, and policy makers and citizen advocates are searching for effective ways to help solve this growing public health problem. PMID- 15289184 TI - Formula for a new foam. AB - As the 2010 phaseout date for chlorofluorocarbons draws nearer, materials engineers are working to find replacements for these ozone-depleting chemicals in the production of plastics and other products. One team of engineers is focusing on a combination of two low-cost and environmentally benign substances- supercritical carbon dioxide and clay nanoparticles--to meet these needs. The result is a strong yet lightweight alternative that retains all the beneficial qualities of solid plastic. PMID- 15289185 TI - Multiple episodes of mild traumatic brain injury result in impaired cognitive performance in mice. AB - OBJECTIVES: Results from recent studies on animal models of concussion suggest that multiple, rather than single, episodes of mild traumatic brain injury result in impaired cognitive performance in mice. The objective of the present study was to administer multiple impacts to the heads of mice while directly measuring the force of the impacts to determine how these parameters are related to transient loss of consciousness, cognitive deficits, and potential neuropathologic effects. METHODS: even-week-old male C57BL/6 mice were randomly assigned to experimental conditions involving three impacts (weight-drop method) to the head to induce mild traumatic brain injury or to sham control procedures. Some impacted (n = 10) and sham control (n = 10) mice were evaluated behaviorally and tested for spatial learning using the Morris water maze (MWM), whereas other impacted (n = 10) and sham control (n = 5) mice were used for histopathologic analysis. RESULTS: The mean ( +/- SD) force of impact was 19 ( +/- 3.5) N. Impacted mice took longer to regain consciousness compared with sham control mice (p < 0.0005). Behavioral test results showed that the groups did not differ on activity or sensorimotor tests or during cued trials in the MWM. Impacted mice exhibited impaired spatial learning performance during place trials in the MWM (p < 0.05). Silver staining revealed a contra-coup type of injury involving ventral brain structures in contact with or in close proximity to the skull. CONCLUSIONS: This multiple impact model, delivered within a specifiable force range, results in transient, reversible loss of consciousness, a contra-coup brain injury, and cognitive impairment. PMID- 15289186 TI - Effects of 4-methylpyrazole on ethanol neurobehavioral toxicity in CD-1 mice. AB - OBJECTIVES: 4-Methylpyrazole (4-MP), an alcohol dehydrogenase (ADH) antagonist, is used for the treatment of ethylene glycol and methanol ingestions. However, ethanol is frequently co-ingested by those who ingest these more toxic alcohols. Several in vitro and in vivo studies have shown a decrease in the elimination rate of ethanol after the administration of 4-MP, but none has evaluated the effects of 4-MP administration on the neurobehavioral toxicity of ethanol. This was a study to determine whether ADH blockade with 4-MP prolongs ethanol neurobehavioral toxicity in a murine model. METHODS: D-1 mice were pretreated with 4-MP, with observation of its effect on ethanol dose-response curves. 4-MP (25 mg/kg) or an equal volume of saline was administered intraperitoneally. Ten minutes later, incremental ethanol doses of 1-5 g/kg were administered intraperitoneally. Pretreated and control groups were composed of ten mice each for each dose of ethanol tested. Outcomes for assessing ethanol neurobehavioral toxicity were successful performance on the rotarod test and presence of the righting reflex, two established and validated outcome measures for ethanol induced neurobehavioral toxicity in mice. RESULTS: The dose of ethanol at which 50% of the animals failed a particular outcome test (toxic dose 50 [TD(50)]) was decreased with 4-MP administration for both the rotarod test and the righting reflex. The TD(50) intergroup differences (control vs. 4-MP) were statistically significant at 60, 120, and 180 minutes (p < 0.05). CONCLUSIONS: Pretreatment with 4-MP significantly prolonged ethanol neurobehavioral toxicity in CD-1 mice, presumably by inhibiting its metabolism by ADH. Further investigation is warranted to evaluate this interaction. PMID- 15289187 TI - Patient and primary care physician satisfaction with chest pain unit and routine care. AB - OBJECTIVES: The chest pain unit (CPU) has been developed to improve care for patients with acute, undifferentiated chest pain. The authors aimed to measure patient and primary care physician (PCP) satisfaction with CPU care and routine care and to determine whether patient satisfaction predicted PCP satisfaction. METHODS: A CPU was established, and 442 days were randomly allocated to either CPU care or routine care. Consenting patients presenting with acute, undifferentiated chest pain were recruited and followed at two days and one month. All were given a self-completed patient satisfaction questionnaire two days after attendance (N = 972). Each patient's PCP was sent a self-completed satisfaction questionnaire during days 171-442 of the trial (N = 601). Analysis determined whether CPU care was associated with improved patient or PCP satisfaction and whether patient satisfaction predicted PCP satisfaction for three questions relating to diagnosis, treatment, and overall care. RESULTS: CPU care was consistently associated with higher scores across all patient satisfaction questions, from the perceived thoroughness of examination to care received to an overall assessment of the service received. However, CPU care achieved small improvements in only two of ten PCP satisfaction questions, concerning overall management of the patient and the amount of information about investigations performed. Furthermore, patient satisfaction did not predict PCP satisfaction in relation to diagnosis (p = 0.456), treatment (p = 0.256), or overall care (p = 0.085). CONCLUSIONS: CPU care is associated with substantial improvements in all dimensions of patient satisfaction but only minimal improvements in PCP satisfaction. Patient satisfaction was not a strong predictor of PCP satisfaction with emergency care. PMID- 15289188 TI - Initial emergency department trauma scores from the OPALS study: the case for the motor score in blunt trauma. AB - OBJECTIVES: To compare the predictive accuracy of the Revised Trauma Score (RTS), the Glasgow Coma Scale (GCS), and their components in blunt trauma patients. METHODS: This multicenter prospective cohort study was conducted in 20 communities as part of the Ontario Prehospital Advanced Life Support (OPALS) Study. It included adult trauma patients with Injury Severity Scores >12. The assessments made by trauma team leaders for the RTS, GCS, and their subscales were analyzed: 1) receiver operating characteristic (ROC) curve areas and Kendall's tau c correlation coefficient (Tc) for survival to hospital discharge, 2) Mann-Whitney U test and Tc correlations for intensive care unit admission, and 3) Spearman correlations with the disability measure Glasgow Outcome Scale. RESULTS: The authors analyzed data from 795 blunt trauma patients with these characteristics: median age of 40 years, 70% male, and 18% mortality. The scores that best predicted survival were the RTS (ROC = 0.83, Tc = 0.39), the GCS (ROC = 0.82, Tc = 0.38), the motor component of the GCS (ROC = 0.81, Tc = 0.37), and the verbal component of the GCS (ROC = 0.81, Tc = 0.36). Only scores for the RTS (p = 0.03), the GCS (p = 0.02), and the motor component of the GCS (p = 0.03) showed a significant association with admission to the intensive care unit. The associations with disability were weak in all scores. CONCLUSIONS: The initial emergency department motor score showed the highest predictive validity among all of the other components. These results suggest its validity for blunt trauma triage when compared with the GCS or RTS. PMID- 15289190 TI - Society for Academic Emergency Medicine survey of the Association of American Medical Colleges Council of Academic Societies. AB - The Society for Academic Emergency Medicine (SAEM) surveyed the Council of Academic Societies (CAS) organizations to obtain useful information to project SAEM goals into the year 2010. The objective of this work was to understand common and varying organizational operations and identify opportunities. The authors reviewed CAS organizations' mission statements, operating budgets, modes of communications, products, meeting formats, foundations, endowment funds, staff structures, headquarters, advocacy activities (services), top challenges, and most significant changes anticipated the next ten years. The survey methodology was used to gain insight into modes of operation of CAS organizations and enable SAEM to review its own operations and identify potential organizational changes based on the experiences of others. Individual CAS organizations might similarly benefit by reviewing the results of the survey and comparing themselves with others. PMID- 15289191 TI - Emergency department orientation utilizing web-based streaming video. AB - To assure a smooth transition to their new work environment, rotating students and housestaff require detailed orientations to the physical layout and operations of the emergency department. Although such orientations are useful for new staff members, they represent a significant time commitment for the faculty members charged with this task. To address this issue, the authors developed a series of short instructional videos that provide a comprehensive and consistent method of emergency department orientation. The videos are viewed through Web based streaming technology that allows learners to complete the orientation process from any computer with Internet access before their first shift. This report describes the stepwise process used to produce these videos and discusses the potential benefits of converting to an Internet-based orientation system. PMID- 15289192 TI - Clinical factors predicting fractures associated with an anterior shoulder dislocation. AB - OBJECTIVES: To identify risk factors for fractures associated with an anterior shoulder dislocation treated in an emergency department (ED). METHODS: A retrospective case-control study over five years of patients with an anterior shoulder dislocation was accomplished in a university-affiliated ED. Chart review identified possible predictors of fractures. Comparing the profile of patients having a clinically important fracture associated with their shoulder dislocation (cases) with those sustaining a noncomplicated dislocation (controls) provided the outcome measure. RESULTS: A total of 334 patients were included in the study. Eighty-five (25.5%) had a clinically important fracture-dislocation, and the remaining 249 (74.5%) sustained a noncomplicated shoulder dislocation. Chi square, logistic regression, and recursive partitioning analysis showed three significant factors for the presence of fracture-dislocation: 1) age 40 years or older, 2) a first episode of dislocation, and 3) mechanism of injury (i.e., a fall greater than one flight of stairs, a fight/assault episode, or a motor vehicle crash). A multiple logistic regression model estimated the significant adjusted odds ratios (and their 95% confidence intervals [95% CIs]) for each of the three factors: 5.18 (95% CI = 2.74 to 9.78), 4.23 (95% CI = 1.82 to 9.87), and 4.06 (95% CI = 1.95 to 8.48), respectively. A predictive model using any one of the three factors reached a sensitivity of 97.7% (95% CI = 91.8% to 99.4%), a specificity of 22.9% (95% CI = 18.1% to 28.5%), and a negative predictive value of 96.6% (95% CI = 88.3% to 99.6%). CONCLUSIONS: Three risk factors predict clinically important fractures that are associated with shoulder dislocation: age, first episode, and mechanism of dislocation. A prospective validation may lead to standardized use of prereduction radiographs of the shoulder in the ED. PMID- 15289193 TI - Research fundamentals: follow-up of subjects in clinical trials: addressing subject attrition. AB - Many published clinical trials have less than adequate follow-up. When conducting a clinical trial, researchers attempt to minimize data loss; however, some data may not be collected, particularly when subjects are lost to follow-up. Careful planning of research protocols, including comprehensive initial data collection, identification of locators, flexible scheduling, systematic subject tracking, monitoring subject loss, and systematically approaching problem cases can ensure high follow-up rates. This article presents a compendium of techniques and procedures that researchers can use to enhance follow-up and address attrition in their studies. Finally, this article outlines statistical techniques that can be used to address the effects of missing data, particularly when patients are lost to follow-up. PMID- 15289194 TI - Bench to bedside: HMGB1-a novel proinflammatory cytokine and potential therapeutic target for septic patients in the emergency department. AB - Overwhelming gram-negative bacterial infection and life-threatening systemic inflammation are widespread problems in critically ill emergency department patients. Currently, the treatment of these patients is largely supportive, focusing on antibiotics, fluids, hemodynamic and ventilatory support, and intensive monitoring. The only Food and Drug Administration-approved pharmaceutical agent for the treatment of sepsis is activated protein C, with its use largely relegated to the intensive care unit. The subject thus remains an active area of exploration for emergency medicine research. During sepsis and inflammation, innate immune cells release excessive amounts of proinflammatory cytokines such as tumor necrosis factor (TNF) and interleukin-1beta. If delivered early enough, anti-TNF antibodies can be an effective therapy in experimental models of septic shock. Anti-TNF antibodies have been developed for clinical use in rheumatoid arthritis and Crohn's disease. However, anti-TNF treatment for sepsis has been difficult to achieve in the clinical setting, perhaps because TNF's early release and transient appearance in the serum create a narrow therapeutic window. An alternative strategy would be to identify "late" mediators that may be clinically more accessible. High mobility group box 1 (HMGB1), a protein previously known only as a nuclear transcription factor, is now implicated as a late mediator of sepsis. Targeting late mediators of lethal systemic inflammation represents a novel approach that may widen the therapeutic window and lead to new strategies for inhibiting the deleterious effects of the inflammatory cascade. Here the authors review the studies that led to the discovery of HMGB1 as a late mediator of systemic inflammation and discuss the possibility of HMGB1 as a therapeutic target for septic patients in the emergency department. PMID- 15289195 TI - Cocaine use in elder patients presenting to an inner-city emergency department. AB - OBJECTIVES: To determine the prevalence of cocaine use in a population of elder patients presenting to an inner-city academic emergency department (ED). METHODS: This was a prospective, blinded observational study of patients aged 60 years or older who presented to a large urban ED over a six-month period. A urine drug screen was performed on patients who had a sample obtained during treatment for routine analysis. Patients' demographic data were collected and compared. RESULTS: A total of 5,677 visits met the inclusion criteria. Urine samples were obtained in 911 (16%) of these visits with 852 unique individuals. There were 18 cocaine-positive results among the 911 visits, for a rate of 2.0%. The rate of positive subjects was also 2.0% (17/852). The cocaine users were younger (66.4 +/ 7.2 vs. 76.0 +/- 8.7 years), predominantly male (88.9% vs. 46.6%), and more likely to be diagnosed with drug or alcohol abuse as compared with the cocaine negative patients. However, there were no significant differences in disposition between the cocaine-positive and cocaine-negative groups. CONCLUSIONS: Elder patients may have a higher prevalence of cocaine use than previously estimated by national registries. PMID- 15289196 TI - Basic cardiac life support providers checking the carotid pulse: performance, degree of conviction, and influencing factors. AB - The American Heart Association recently abolished the carotid pulse check during cardiopulmonary resuscitation for lay rescuers, but not for health care providers. OBJECTIVES: The aim of the study was to evaluate health care providers' performance, degree of conviction, and influencing factors in checking the carotid pulse. METHODS: Sixty-four health care providers were asked to check the carotid pulse for 10 or 30 seconds on a computerized mannequin simulating three levels of pulse strength (normal, weak, and absent). Health care providers were asked whether they felt a pulse and how certain were they that they felt a pulse. Performance was evaluated, as well as degree of conviction about the answer, using a visual analog scale. Data were compared by using a general linear model procedure. RESULTS: In the pulseless situations, the answers were correct in 58% and 50% when checking the pulse for 10 and 30 seconds, respectively. In the situation with a weak pulse, the answer was correct in 83% when checking the pulse for 10 seconds. In situations with a normal pulse, the answers were correct in 92%, 84%, and 84%, respectively, when checking the pulse for 10 (twice) and 30 seconds. The exactitude of the answer was correlated with the pulse strength (p < 0.05). The degree of conviction about the answer was correlated with the exactitude of the answer (p < 0.01) and the pulse strength (p < 0.0001). CONCLUSIONS: These results question the routine use of the carotid pulse check during cardiopulmonary resuscitation, including for health care providers. PMID- 15289197 TI - Quantitative validation of a general competency composite assessment evaluation. AB - OBJECTIVES: The authors sought to modify and validate a composite assessment evaluation process that assesses resident acquisition of the Accreditation Council for Graduate Medical Education (ACGME) general competencies (GCs). METHODS: This study critically analyzed the evaluation process used in a multicenter study (150 emergency medicine resident evaluations) to determine whether the procedure was psychometrically valid. For each GC, principal component analysis (PCA) was used to determine whether certain evaluation items could be eliminated, as well as to determine the magnitude of variability explained by up to three linear combinations or "principal components." The factor proportions (factor loadings) of various eigenvectors were measured to determine the degree of variability (determined by the square of the factor proportion) within a data or item set. The factor proportions essentially measure the length of the eigenvector as determined from a correlation matrix. RESULTS: The first three principal components are reported as factor proportion sum (% of total variability) as follows: patient care 0.91 (83%), medical knowledge 0.87 (76%), practice-based learning and improvement 0.90 (81%), interpersonal and communication skills 0.84 (71%), professionalism 0.74 (55%), and systems-based practice 0.80 (64%). PCA showed that evaluating certain traditional categories such as medical knowledge seemed to capture a single element, whereas professionalism appeared to measure a more complex, multidimensional phenomenon. CONCLUSIONS: By using a structured development process, the authors were able to create valid evaluation items for determining resident acquisition of the ACGME GCs. PMID- 15289199 TI - The saliva strip test is an accurate method to determine blood alcohol concentration in trauma patients. AB - OBJECTIVES: To determine the accuracy of alcohol saliva testing (AST) in trauma patients. METHODS: Blood alcohol concentration (BAC) was measured by using both AST (QED A350; STC Technologies, Bethlehem, PA) and blood serum levels in 100 trauma patients admitted to the emergency department of an urban Level 1 trauma center. RESULTS: All 41 patients who tested positive for BAC on AST (mean [+/ SD]: 167.9 +/- 16.16; range: 20-350 mg/dL) also tested positive on serum determination (mean: 197.6 +/- 13.79; range: 22-446 mg/dL). Correlation between the two positive tests was significant (0.879, p < 0.001). Of the remaining 61 patients, 59 tested negative on both tests, while two patients with BACs of <30 mg/dL tested negative on the AST. For 18 patients with blood in the oropharynx, there was a correlation of 0.976 (p < 0.001, two-tailed) between serum and AST tests. CONCLUSIONS: The AST method of measuring BAC in trauma patients is accurate. Blood in the oral cavity did not appear to affect the accuracy of the test. PMID- 15289200 TI - Comparison of responses to a US 2 dollar bill versus a chance to win 250 US dollars in a mail survey of emergency physicians. AB - OBJECTIVES: To evaluate whether small monetary incentives improve physicians' responses to surveys. To the best of the authors' knowledge, no one has evaluated emergency physicians' response rate and cost per participant of a small monetary incentive relative to a chance to win a more substantial sum. The authors compared emergency physicians' responses and per-participant costs between a US 2 dollar bill and a 250 US dollars lottery. METHODS: Two groups of 288 emergency physicians were randomly selected and mailed a survey. Within each group of 288, half received a US 2 dollar bill and the other half received an offer that respondents would be entered into a drawing to win 250 US dollars. Nonresponders received a reminder postcard one week later, and persistent nonresponders received a second mailing of the survey three weeks after the initial mailing. RESULTS: Of the 576 surveys that were mailed, nine (2%) subjects were ineligible or undeliverable, leaving 567 eligible subjects, of whom 301 (53%) participated in the survey. The US 2 dollar bill had a substantially higher response rate: 170 (56%) of those receiving a US 2 dollar bill participated versus 131 (44%) of those receiving a chance to win 250 US dollars (95% confidence interval = 5% to 22%; p < 0.001). The US 2 dollar bill offer was less expensive per participant than the 250 US dollars offer. The cost of postage and incentives was 997.33 US dollars for 170 participants, or 5.87 US dollars per participant, for the US 2 dollar bill and 979.29 US dollars for 131 participants, or 7.48 US dollars per participant, for the chance to win 250 US dollars. CONCLUSIONS: Mailing a US 2 dollar bill incentive produces a better response rate with lower cost per participant than offering a chance to win 250 US dollars. PMID- 15289202 TI - Ethanol modulates neuropeptide-degrading aminopeptidases at synapse level in calcium-dependent conditions. AB - AIMS: To investigate the role of aminopeptidases in the pathways to peptides neurotransmission/neuromodulation ending in the actions of ethanol (EtOH) on the brain. METHODS: The effects of EtOH on alanyl-, arginyl-, cystyl-, leucyl- and tyrosyl-aminopeptidase activities were studied under basal/resting and K+ stimulated conditions at the synapse level, using mouse frontal cortex synaptosomes and their incubation supernatant in a Ca2+-containing or Ca2+-free medium. RESULTS: Under basal conditions, synaptosome aminopeptidase activities showed an inhibitory or biphasic response depending on the concentration of EtOH used and the aminopeptidase assayed, whereas supernatant activities showed a more complex response. Under K+-stimulated conditions, EtOH inhibited all synaptosome aminopeptidases assayed in presence of Ca2+. However, in absence of Ca2+, different responses were obtained depending on the concentration of EtOH used. In the supernatant, the highest concentration of EtOH inhibited the K+-stimulated increase on aminopeptidase activities, although the lowest concentration enhanced the release in presence of Ca2+. In absence of it, EtOH blocked the K+-stimulated decrease or increased the activity depending on the concentration of EtOH used. CONCLUSIONS: The changes on aminopeptidase activities induced by EtOH may reflect the functional status of their corresponding endogenous substrates. EtOH may influence opioid peptides, oxytocin, vasopressin and the brain renin-angiotensin system through their degrading enzymes. PMID- 15289203 TI - Problems with the graduated frequency approach to measuring alcohol consumption: results from a pilot study in Toronto, Canada. AB - AIMS: To evaluate advantages and disadvantages of the graduated frequency (GF) approach, which asks about the frequency of alcohol consumption at mutually exclusive quantity levels (i.e. 12 or more drinks, at least eight drinks but less than 12, etc.). METHODS: Telephone survey of 464 adults aged 18 and older in Toronto, Canada, using random digit dialing and computer-assisted telephone interviewing. RESULTS: Respondents reported higher frequency and volume of drinking on the GF compared to overall and beverage-specific quantity-frequency type measures; however, at least 16% of GF responses included double counting on their frequency estimates using the GF. When these cases were excluded or corrected, differences between the GF and quantity-frequency measures mostly disappeared. The GF was superior to quantity-frequency measures for identifying heavy episodic drinkers. However, the GF had little advantage over the weekly recall method except for identifying very infrequent (i.e. less often than twice a month) heavy drinkers. CONCLUSIONS: Because the GF has a high rate of response errors in terms of measuring frequency of alcohol consumption, other combinations of measures, including alternate measures of heavy episodic drinking should be considered. PMID- 15289204 TI - High alcohol preferring (HAP) and low alcohol preferring (LAP) rats show altered proopiomelanocortin (POMC) messenger RNA expression in the arcuate nucleus. AB - AIMS AND METHODS: We have investigated proopiomelanocortin (POMC) and neuropeptide Y (NPY) gene expression in the arcuate nucleus of the hypothalamus comparing high alcohol preference (HAP) rats and low alcohol preference (LAP) rats under basal conditions using in situ hybridization histochemistry. RESULTS: A significantly higher expression of POMC mRNA was observed in HAP rats compared with LAP rats. In contrast, no difference in NPY mRNA expression was observed between the two rat lines. CONCLUSIONS: These data suggest that an endogenous opioid system may contribute to alcohol preference in HAP/LAP rat lines. PMID- 15289205 TI - CUGE: a screening instrument for alcohol abuse and dependence in students. AB - OBJECTIVES: The prevalence of alcohol abuse on college campuses ranges from 7 to 17%. Frequent heavy drinkers place themselves and others at risk for a variety of adverse consequences and frequently remain undetected. Brief individual interventions result in a significant reduction on the number of drinks. Therefore, detection of students at risk is useful and desirable. The CUGE has been elsewhere described as a promising screening device for problem drinking in students. In order to determine the diagnostic value of this new questionnaire, we set up a validation study in a new and independent population of freshmen. METHODS: A cross-sectional diagnostic study. Participants were college freshmen of the Katholieke Universiteit Leuven. All students received a questionnaire, containing the CUGE, being the test of interest, and the CIDI as the reference test. RESULTS: The CUGE combines a very high sensitivity of 91% with a reasonable specificity of 76.3% in this validation group. CONCLUSIONS: The CUGE is an excellent screening device in this population of students. In addition, it is a short questionnaire with only yes or no questions. This makes the CUGE easily applicable as a part of broad routine questionnaires. PMID- 15289206 TI - Ethyl glucuronide: a biomarker to identify alcohol use by health professionals recovering from substance use disorders. AB - AIMS: Physicians recovering from substance-related disorders are usually allowed to return to practice if they agree to remain abstinent from drugs, including alcohol, and to undergo random urine testing. Over 9000 physicians are currently involved in such monitoring programs in the US. To date, it has been difficult to adequately monitor abstinence from alcohol due to the short half-life of alcohol and no other highly specific marker. Ethyl glucuronide (EtG), a direct metabolite of alcohol, offers an extended window for assessment of drinking status (up to 5 days). Our aim was to assess the potential value of EtG testing in abstinence based monitoring programs. PATIENTS AND METHODS: Urine samples were obtained from 100 participants in a physician monitoring program and additional samples were subsequently obtained 'for cause', 'to verify positive urine alcohol, when drinking was denied' and 'in high risk individuals'. All participants had signed contracts agreeing to remain abstinent from mood-altering drugs, including alcohol, and had agreed to random urine testing. EtG was determined using LC/MS MS in addition to standard testing. The main outcome measure were urine specimens positive for EtG versus those positive based on standard testing for alcohol and other drugs. RESULTS: Among the initial 100 random samples collected, no sample was positive for alcohol using standard testing; however, seven were positive for EtG (0.5-196 mg/l), suggesting recent alcohol use. Subsequent EtG testing was performed clinically during the course of monitoring. Of the 18 tests performed to date, eight of eight tests performed 'for cause' were positive for EtG but negative for all other drugs including urine alcohol. All eight were confirmed positive by self reported drinking by the patient when confronted regarding the positive test result. Of six tests performed to 'confirm a positive urine alcohol' two were positive for EtG and confirmed positive by self reported drinking. For the other four samples, especially as two are from a diabetic, in vitro fermentation of ethanol is discussed. CONCLUSIONS: These data suggest that physicians in monitoring programs have a higher rate of unrecognized alcohol use than previously reported. Incorporation of EtG testing into alcohol abstinence monitoring can strengthen these programs. PMID- 15289207 TI - Role of angiotensin II and the subfornical organ in the pharmacological actions of ethanol. AB - AIMS: The current study was designed to evaluate if angiotensin II mediates the hypothermic effects of ethanol, and to determine if the effects of angiotensin are mediated centrally. We also tested the hypothesis that the subfornical organ (SFO) is a site responsible for the alterations in body temperature and aerial righting reflex mediated by ethanol and for the modulation of ethanol consumption in rats. METHODS: Male Sprague-Dawley rats were used in a series of experiments to evaluate the role of both peripheral and central administration of losartan, a selective angiotensin type 1 receptor antagonist on ethanol-induced hypothermia. Subsequent studies were undertaken in SFO-lesioned rats to evaluate the effects of SFO-lesion on alcohol intake, the thermal response to alcohol and angiotensin, and the aerial righting reflex. RESULTS: Selective antagonism of the angiotensin II type 1 receptor, administered either peripherally or centrally, attenuated not only the fall in colonic temperature but also attenuated the transient rise in tail skin temperature that was associated with administration of ethanol. The thermal responses to both angiotensin and ethanol were similarly attenuated in SFO-lesioned rats. Likewise the aerial righting reflex, which has previously been shown to be impaired by losartan treatment, was also significantly attenuated in SFO-lesioned animals. Alcohol intake, as determined by a 48 h, two-bottle preference test also revealed that SFO-lesioned animals consumed significantly less alcohol (ethanolic beer) than did controls. CONCLUSION: Collectively, the results demonstrate that ethanol-induced temperature responses are mediated by the renin-angiotensin system and that this interaction is mediated centrally. In addition, the results demonstrate that the SFO is a site that mediates several neurobiological effects of ethanol, possibly via the renin-angiotensin system. PMID- 15289208 TI - Gender differences in the relationships between alcohol, tobacco and mental health in patients attending an emergency department. AB - AIMS: There is evidence of a non-linear relationship between alcohol consumption and mental health status, and of an association between tobacco use and poor mental health. This paper examines the nature of the association between usual alcohol consumption, tobacco use and symptoms of anxiety and depression in Emergency Department patients in Queensland, Australia. METHODS: A cross sectional survey of patients aged 16-84 presenting for treatment over a 14 day period to Gold Coast Hospital Emergency Department using socio-demographic items, the Alcohol Use Disorders Identification Test (AUDIT) to measure moderate, hazardous and harmful alcohol consumption, and the Hospital Anxiety and Depression Scale (HADS) to measure state anxiety and depression. RESULTS: 812 patients were interviewed. Gender differences in results were evident. For men, there was a U-shaped relationship between alcohol consumption and anxiety/depression, and a linear association between smoking and anxiety. For women, alcohol consumption and anxiety/depression showed a more linear relationship, but there was no significant relationship between tobacco use and anxiety/depression. CONCLUSIONS: There may be important gender differences in the relationships between alcohol consumption, tobacco use and mental health status. This study supports previous evidence that mental health status of non-drinkers is worse than that of moderate drinkers, but only among males. PMID- 15289209 TI - Changes over time in the self-reported level of response to alcohol. AB - AIMS: A low level of response to alcohol, or the need for a higher number of drinks for an effect, is a risk factor for alcohol use disorders. The response to alcohol is usually measured in young subjects, and changes in this phenomenon over time have rarely been evaluated. Reports that, overall, individuals are likely to become more reactive to alcohol with advancing age led to the current evaluation to determine whether the number of drinks needed for an effect decreased between the teens and age 40 in a group of men. METHODS: Data were available from the 20-year follow-up of 202 men who had originally been chosen at age 20 as nonalcoholic subjects from families at high and low risk for alcoholism. The number of drinks required for effects was determined through the Self-Report of the Effects of Alcohol questionnaire (SRE) regarding their recollection of their intensity of reaction early in their drinking careers, as well as reports regarding the recent (e.g. prior 3 month) response to alcohol at the 15- and 20-year follow-ups. RESULTS: Overall, there was a slight decrease in the drinks required for effects (SRE scores) across the three time points which became significant when recent drinking and depressant medication use were evaluated as covariates. When nonalcoholic or light-drinking subjects were evaluated separately, the decrease in the number of drinks needed for effects was more prominent. Among heavier drinkers, there was an increase in the number of drinks required for effects over time. The findings were generally similar for men with and without alcoholic relatives. CONCLUSIONS: The development of a more intense reaction to alcohol, or the need for fewer drinks for an effect, with advancing age may only be relevant to lighter drinkers. Among heavier drinkers, the finding that a higher number of drinks are required for effects may be relatively stable over time. PMID- 15289210 TI - Chronic alcohol consumption yields sex differences in whole-body glucose production in rats. AB - AIMS: The effects of chronic alcohol consumption (8 weeks) on glucose kinetics, in the absence (water, 4 g/kg) and presence of an acute ethanol dose (4 g/kg), were examined in 48 h fasted male and female Wistar rats. METHODS: Primed continuous infusions of [6-3H]- and [U-14C]glucose were employed to assess rates of glucose appearance (Ra), glucose disappearance (Rd), and apparent glucose carbon recycling. RESULTS: After injecting the male and female controls with water, there were no significant alterations in glucose kinetics. Compared to controls, chronic alcohol-fed female animals (injected with water) demonstrated significantly lower: glucose Ra, blood glucose concentration, and apparent glucose carbon recycling for a majority of the experimental period. In separate groups injected with ethanol, the glucose Ra fell by 31% for male rats fed the control diet (MC), 43% for male rats fed the ethanol diet (ME), 29% for female rats fed the control diet (FC), and 42% for female rats fed the ethanol diet (FE). Further, compared to controls (MC and FC), the blood glucose concentration was significantly lower prior to and following the ethanol injection for FE. In addition, FE animals had significantly lower rates of glucose Ra and glucose carbon recycling compared to controls prior to and after the ethanol injection. ME animals demonstrated similar declines in glucose Ra (compared to FE), but only after the ethanol injection. Conversely, ME were able to match the decrease in glucose Ra with comparable declines in glucose Rd resulting in blood glucose concentrations that did not differ from controls. CONCLUSIONS: Chronic alcohol consumption results in sex differences in whole-body glucose production and glucose regulation. PMID- 15289211 TI - Chronic alcoholism causes deleterious conditioning of innate immunity. AB - AIMS: To examine the immune consequences of chronic alcoholism in man, in relation to the known association between alcoholism and raised incidence and severity of infections. METHODS: In 36 alcoholics without liver disease, at the point of commencing withdrawal from alcohol, the following measures of immune competence were measured: the immunophenotypes of cells, acute phase proteins, the endotoxin-neutralizing capacity (ENC) of the serum, titers of anti lipopolysaccharide (LPS) antibodies, and ex vivo cytokine inducibility in T cells and monocytes (TNFalpha, IL1beta, IL1RA, IL4, IL6, IL8, IL10 and IL12). The results were compared to those from healthy volunteers (day controls). Measures were repeated after 8-13 days of abstinence. RESULTS: LPS-binding protein (LBP) and soluble CD14 (sCD14) were significantly increased in patients' sera at the outset of withdrawal, whereas reduced titers of anti-LPS IgG (P = 0.012) and a reduced ENC (P = 0.001) were measured. Only ENC rapidly returned to normal values after withdrawal therapy. Cytokine induction with phorbol ester showed no significant alterations in patients' T cells. Patients' monocytes, however, responded to LPS stimulation with enhanced IL1beta-, but reduced TNFalpha- and IL12-production (P = 0.004, P = 0.0042 and P = 0.001, respectively). While IL1- and TNFalpha-responses normalized after the withdrawal period, impairment of the IL12 response persisted throughout the observation period of 2 weeks. CONCLUSIONS: Alcoholism results in a prolonged LPS-mediated hypoinflammatory conditioning of the innate but not the adaptive immune system, which is not reversed immediately after withdrawal. This alcohol-induced status of the immune system predisposes to infections and sepsis by blunting initial response to the pathogens. PMID- 15289212 TI - Effect of ethanol on morphine state-dependent learning in the mouse: involvement of GABAergic, opioidergic and cholinergic systems. AB - AIMS: We have studied the effect of acute administration of ethanol when it replaced morphine in step-down passive avoidance task on the test day and the effects of antagonists of GABAergic, opioidergic and cholinergic systems on ethanol actions. METHODS: Morphine (5 mg/kg, s.c.) was administered as pre training and 24 h later as pre-test drug, and the latencies were measured in mice. Ethanol (0.125, 0.25, 1 and 2 g/kg, i.p.) was administered instead of pre test morphine. Antagonists of GABAergic (bicuculline 0.5, 1 and 2 mg/kg, i.p.), opioidergic (naloxone 0.06, 0.25 and 1 mg/kg, i.p.) and cholinergic (atropine 0.625 and 1.25 mg/kg, i.p. and mecamylamine 0.5, 1 and 2 mg/kg, i.p.) systems were co-administered with ethanol (0.25 g/kg, i.p.) on the test day. Locomotor activity was measured as well. RESULTS: Pre-training morphine impaired the memory on the test day which was restored when the same dose of morphine was used as pre test drug. All four doses of ethanol replaced pre-test morphine and enhanced the memory. This effect was prevented by all of the above antagonists. No significant changes were seen in the locomotor activity of the animals treated with ethanol or antagonists compared to the proper controls. CONCLUSIONS: GABAergic, endogenous opioidergic and cholinergic systems are involved in the memory recall improvement by ethanol when it replaced morphine on the test day. A review of the literature suggests other possibilities such as the release of intermediate neurotransmitters. PMID- 15289213 TI - Sildenafil increased exercise capacity during hypoxia at low altitudes and at Mount Everest base camp: a randomized, double-blind, placebo-controlled crossover trial. AB - BACKGROUND: Alveolar hypoxia causes pulmonary hypertension and enhanced right ventricular afterload, which may impair exercise tolerance. The phosphodiesterase 5 inhibitor sildenafil has been reported to cause pulmonary vasodilatation. OBJECTIVE: To investigate the effects of sildenafil on exercise capacity under conditions of hypoxic pulmonary hypertension. DESIGN: Randomized, double-blind, placebo-controlled crossover study. SETTING: University Hospital Giessen, Giessen, Germany, and the base camp on Mount Everest. PARTICIPANTS: 14 healthy mountaineers and trekkers. MEASUREMENTS: Systolic pulmonary artery pressure, cardiac output, and peripheral arterial oxygen saturation at rest and during assessment of maximum exercise capacity on cycle ergometry 1) while breathing a hypoxic gas mixture with 10% fraction of inspired oxygen at low altitude (Giessen) and 2) at high altitude (the Mount Everest base camp). INTERVENTION: Oral sildenafil, 50 mg, or placebo. RESULTS: At low altitude, acute hypoxia reduced arterial oxygen saturation to 72.0% (95% CI, 66.5% to 77.5%) at rest and 60.8% (CI, 56.0% to 64.5%) at maximum exercise capacity. Systolic pulmonary artery pressure increased from 30.5 mm Hg (CI, 26.0 to 35.0 mm Hg) at rest to 42.9 mm Hg (CI, 35.6 to 53.5 mm Hg) during exercise in participants taking placebo. Sildenafil, 50 mg, significantly increased arterial oxygen saturation during exercise (P = 0.005) and reduced systolic pulmonary artery pressure at rest (P < 0.001) and during exercise (P = 0.031). Of note, sildenafil increased maximum workload (172.5 W [CI, 147.5 to 200.0 W]) vs. 130.6 W [CI, 108.8 to 150.0 W]); P < 0.001) and maximum cardiac output (P < 0.001) compared with placebo. At high altitude, sildenafil had no effect on arterial oxygen saturation at rest and during exercise compared with placebo. However, sildenafil reduced systolic pulmonary artery pressure at rest (P = 0.003) and during exercise (P = 0.021) and increased maximum workload (P = 0.002) and cardiac output (P = 0.015). At high altitude, sildenafil exacerbated existing headache in 2 participants. LIMITATIONS: The study did not examine the effects of sildenafil on normoxic exercise tolerance. CONCLUSIONS: Sildenafil reduces hypoxic pulmonary hypertension at rest and during exercise while maintaining gas exchange and systemic blood pressure. To the authors' knowledge, sildenafil is the first drug shown to increase exercise capacity during severe hypoxia both at sea level and at high altitude. PMID- 15289214 TI - Physician reports of terminal sedation without hydration or nutrition for patients nearing death in the Netherlands. AB - BACKGROUND: Terminal sedation in patients nearing death is an important issue related to end-of-life care. OBJECTIVE: To describe the practice of terminal sedation in the Netherlands. DESIGN: Face-to-face interviews. SETTING: The Netherlands. PARTICIPANTS: Nationwide stratified sample of 482 physicians; 410 responded and 211 of these reported characteristics of their most recent terminal sedation case. MEASUREMENTS: Physician reports of frequency of terminal sedation (defined as the administration of drugs to keep the patient in deep sedation or coma until death, without giving artificial nutrition or hydration), characteristics of the decision-making process, drugs used, the estimated life shortening effect, and frequency of euthanasia discussions. RESULTS: Of respondents, 52% (95% CI, 48% to 57%) had ever used terminal sedation. Of the 211 most recent cases, physicians used terminal sedation to alleviate severe pain in 51% of patients (CI, 44% to 58%), agitation in 38% (CI, 32% to 45%), and dyspnea in 38% (CI, 32% to 45%). Physicians reported discussing with patients the decision to use deep sedation in 59% of the 211 most recent cases (CI, 52% to 66%) and the decision to forgo artificial nutrition or hydration in 34% (CI, 28% to 41%). Hastening death was partly the intention of the physician in 47% (CI, 41% to 54%) of cases and the explicit intention in 17% (CI, 13% to 22%) of cases. LIMITATIONS: The generalizability of physician reports about their most recent cases to all terminal sedation cases is uncertain. In addition, the findings are subject to recall bias and may not apply to other geographic settings. CONCLUSIONS: Terminal sedation precedes a substantial number of deaths in the Netherlands. In about two thirds of most recently reported cases, physicians indicated that in addition to alleviating symptoms, they intended to hasten death. PMID- 15289215 TI - The effect of routine, early invasive management on outcome for elderly patients with non-ST-segment elevation acute coronary syndromes. AB - BACKGROUND: Although increasing age is an important risk factor for adverse outcome among patients with acute coronary syndromes, elderly patients are more often managed conservatively. OBJECTIVE: To examine outcome according to age and management strategy for patients with unstable angina and non-ST-segment elevation myocardial infarction (MI). DESIGN: Randomized, controlled trial conducted from December 1997 to June 2000. SETTING: 169 community and tertiary care hospitals in 9 countries. PATIENTS: 2220 patients hospitalized with unstable angina and non-ST-segment elevation MI who were randomly assigned to an early invasive or conservative management strategy. INTERVENTIONS: Medical therapy and coronary angiography at 4 to 48 hours versus medical therapy and predischarge exercise testing. MEASUREMENTS: Rates of 30-day and 6-month mortality, nonfatal MI, rehospitalization, stroke, and hemorrhagic complications. RESULTS: Among patients 65 years of age and older, the early invasive strategy compared with the conservative strategy yielded an absolute reduction of 4.8 percentage points (8.8% vs. 13.6%; P = 0.018) and a relative reduction of 39% in death or MI at 6 months. Outcomes of the 2 strategies were similar, however, among patients younger than 65 years of age (6.1% vs. 6.5%; P > 0.2). Among patients older than 75 years of age, the early invasive strategy conferred an absolute reduction of 10.8 percentage points (10.8% vs. 21.6%; P = 0.016) and a relative reduction of 56% in death or MI at 6 months. The additional cost per death or MI prevented with the early invasive strategy was lower for elderly patients, but major bleeding rates were higher with this strategy in patients older than 75 years of age (16.6% vs. 6.5%; P = 0.009). LIMITATIONS: Because this study involved patients in the Treat Angina with Aggrastat and Determine Cost of Therapy with an Invasive or Conservative Strategy-Thrombolysis in Myocardial Infarction (TACTICS TIMI) 18 trial, its generalizability to elderly patients with excluded comorbid conditions is unknown. CONCLUSION: Despite an increased risk for major bleeding in patients older than 75 years of age, a routine early invasive strategy can significantly improve ischemic outcomes in elderly patients with unstable angina and non-ST-segment elevation MI. PMID- 15289216 TI - The impact of peer management on test-ordering behavior. AB - BACKGROUND: Laboratory testing of hospitalized patients, although essential, can be expensive and sometimes excessive. Attempts to reduce unnecessary testing have often been difficult to implement or sustain. OBJECTIVE: Use of peer management through a resource utilization committee (RUC) to favorably modify test-ordering behavior in a large academic medical center. DESIGN: Interrupted time-series study. SETTING: Medical center with inpatient care provider order entry (CPOE) system and database of ordered tests. PARTICIPANTS: Predominantly housestaff physicians but all clinical staff (attending physicians, housestaff, medical students, nurses, advance practice nurses, and other clinical staff) at Vanderbilt University Hospital who used CPOE systems. INTERVENTION: The RUC analyzed the ordering habits of providers during previous years and made 2 interventions by modifying software for the CPOE system. The committee first initiated a daily prompt in the system that asked providers whether they wanted to discontinue tests scheduled beyond 72 hours. After evaluating this first intervention, the committee further constrained testing options by unbundling serum metabolic panel tests (sodium, potassium, chloride, bicarbonate, glucose, blood urea nitrogen, and creatinine tests) into single components and by reducing the ease of repeating targeted tests (including electrolyte, blood urea nitrogen, creatinine, and glucose tests; electrocardiography; and portable chest radiography). MEASUREMENTS: Pre- and postintervention volumes of tests; proportion of patients with abnormal targeted chemistry levels after 48 hours; rates of repeated admission, transfer to intensive care units, and mortality; adjusted coefficient of variation for test ordering; and length of stay. RESULTS: Voluntary reduction of testing beyond 72 hours (first intervention) decreased orders for metabolic panel component tests by 24% (P = 0.02) and electrocardiograms by 57% (P = 0.006) but not orders for portable chest radiographs. Prospective constraints on recurrent test ordering with panel unbundling (second intervention) produced an additional decrease of 51% for metabolic panel component tests (P < 0.001) and 16% for portable chest radiographs (P = 0.03). Incidence of patients with abnormal targeted blood chemistry levels after 48 hours decreased after the intervention (P = 0.02). Postintervention-adjusted coefficients of variation decreased for metabolic panel component tests (P = 0.03) and electrocardiography (P = 0.04). Rates of (adjusted) monthly readmission, transfers to intensive care units, hospital length of stay, and mortality were unchanged. LIMITATIONS: Other activities occurring during the time period of the interventions might have influenced some test-ordering behaviors, and we assessed effects on only a limited number of commonly ordered tests. CONCLUSIONS: Peer management reduced provider variability by addressing the imperfect ability of clinicians to rescind testing in a timely manner. Hospitals with growing health care costs can improve their resource utilization through peer management of testing behaviors by using CPOE systems. PMID- 15289218 TI - Update in general internal medicine. PMID- 15289217 TI - Compensation and advancement of women in academic medicine: is there equity? AB - BACKGROUND: Women have been entering academic medicine in numbers at least equal to their male colleagues for several decades. Most studies have found that women do not advance in academic rank as fast as men and that their salaries are not as great. These studies, however, have typically not had the data to examine equity, that is, do women receive similar rewards for similar achievement? OBJECTIVE: To examine equity in promotion and salary for female versus male medical school faculty nationally. DESIGN: Mailed survey questionnaire. SETTING: 24 randomly selected medical schools in the contiguous United States. PARTICIPANTS: 1814 full time U.S. medical school faculty in 1995-1996, stratified by sex, specialty, and graduation cohort. MEASUREMENTS: Promotion and compensation of academic medical faculty. RESULTS: Among the 1814 faculty respondents (response rate, 60%), female faculty were less likely to be full professors than were men with similar professional roles and achievement. For example, 66% of men but only 47% of women (P < 0.01) with 15 to 19 years of seniority were full professors. Large deficits in rank for senior faculty women were confirmed in logistic models that accounted for a wide range of other professional characteristics and achievements, including total career publications, years of seniority, hours worked per week, department type, minority status, medical versus nonmedical final degree, and school. Similar multivariable modeling also confirmed gender inequity in compensation. Although base salaries of nonphysician faculty are gender comparable, female physician faculty have a noticeable deficit (-11,691 dollars; P = 0.01). Furthermore, both physician and nonphysician women with greater seniority have larger salary deficits (-485 dollars per year of seniority; P = 0.01). LIMITATIONS: This is a cross-sectional study of a longitudinal phenomenon. No data are available for faculty who are no longer working full-time in academic medicine, and all data are self-reported. CONCLUSIONS: Female medical school faculty neither advance as rapidly nor are compensated as well as professionally similar male colleagues. Deficits for female physicians are greater than those for nonphysician female faculty, and for both physicians and nonphysicians, women's deficits are greater for faculty with more seniority. PMID- 15289219 TI - From unequal treatment to quality care. PMID- 15289220 TI - The Patient's role in reducing disparities. PMID- 15289221 TI - Diversifying the racial and ethnic composition of the physician workforce. PMID- 15289222 TI - Will racial and ethnic disparities in health be resolved primarily outside of standard medical care? PMID- 15289223 TI - Racial and ethnic disparities in health care: a position paper of the American College of Physicians. AB - Disparities clearly exist in the health care of racial and ethnic minorities. This position paper of the American College of Physicians (ACP) provides ample evidence illustrating that minorities do not always receive the same quality of health care, do not have the same access to health care, are less represented in the health professions, and have poorer overall health status than nonminorities. The ACP finds this to be a major problem in our nation's health system that must be addressed. The ACP is dedicated to working toward eliminating all disparities in health care. This position paper sets forth specific positions for reducing these disparities and will be the foundation for public policy advocacy by ACP for eliminating racial and ethnic disparities in health care. PMID- 15289224 TI - Sildenafil for enhanced performance at high altitude? PMID- 15289225 TI - Terminal sedation: an acceptable exit strategy? PMID- 15289226 TI - Unequal pay for equal work: the gender gap in academic medicine. PMID- 15289227 TI - Of ears and E-mails. PMID- 15289228 TI - Metabolic markers of insulin resistance in overweight persons. PMID- 15289229 TI - Albuminuria and mortality in hypertension. PMID- 15289230 TI - Physician support for covering and caring for the uninsured. PMID- 15289231 TI - Screening and interventions for obesity in adults. PMID- 15289232 TI - Successful octreotide treatment of chylous pleural effusion and lymphedema in the yellow nail syndrome. PMID- 15289233 TI - Summaries for patients. Sildenafil increases exercise capacity in low-oxygen settings. PMID- 15289234 TI - Summaries for patients. Doctors' reports of terminal sedation without hydration or nutrition for patients nearing death in the Netherlands. PMID- 15289235 TI - Summaries for patients. Invasive versus conventional management of elderly patients with acute coronary artery disease: which is better? PMID- 15289236 TI - Summaries for patients. Altering test-ordering behavior of hospital staff. PMID- 15289237 TI - Leprosy accidentally transmitted from a patient to a surgeon in a nonendemic area. PMID- 15289238 TI - Male breast cancer: is the incidence increasing? AB - BACKGROUND: Male breast cancer is rare, and little is known about state population-level patterns of incidence. The primary objective of this study was to determine the incidence of MBC in Florida in comparison with the Surveillance, Epidemiology, and End RESULTS (SEER) program data. METHODS: Study data were obtained from the Florida Cancer Data System (FCDS). All males with pathologically confirmed invasive breast carcinoma diagnosed from 1985 to 2000 were included. Age-adjusted incidence rates, regional incidence rates, and descriptive statistics were calculated. Annual percent change (APC) for the study period was calculated with a linear model. Results were compared with the SEER data. RESULTS: A total of 1396 cases of MBC were identified. Age-adjusted incidence rates increased from 0.9 cases per 100,000 in 1990 to 1.5 cases per 100,000 in 2000. In 2000, the highest rates were in the age groups of 70 to 75 years (7.9) and > or =85 years (12.5). Infiltrating ductal was the most common subtype (92%); less common subtypes included mucinous (2%) and papillary (2%). Localized disease accounted for 45% of all cases, with regional disease in 33%, distant metastases in 7%, and unstaged in 15%. Most incident cases were diagnosed in the Palm Beach-Broward region (23%). The number of cases increased from 56 in 1985 to 132 new cases in 2000. The APC for this 16-year period was 2.0% (95% confidence interval [CI], 1.05-3.01; P <.005). SEER data indicated no change in MBC incidence rates (APC, 0.5; NS). CONCLUSIONS: The incidence of MBC in Florida increased significantly between 1985 and 2000. This finding is discordant with SEER incidence data. Further epidemiologic studies are warranted to investigate regional variation. PMID- 15289239 TI - Preoperative magnetic resonance imaging evaluation for breast cancers after sonographically guided core-needle biopsy: a comparison study. AB - BACKGROUND: The aim of the study was to evaluate the efficacy of contrast enhanced magnetic resonance imaging (MRI) for preoperative assessment of palpable breast cancer after sonographically guided percutaneous core-needle biopsy. METHODS: Thirty-six breast cancers in 35 women that had been diagnosed by sonographically guided core-needle biopsy prior to subsequent MRI were evaluated in this retrospective study. Radiological and pathological reports, multiplicity, retroareolar involvement, and the size of the breast cancers were reviewed. The cancer sizes, as derived from sonography and enhanced MRI, were correlated with histological size in greatest diameter by means of Pearson's correlation. The threshold value for significance was set at P <.05. RESULTS: Synchronous breast cancers were revealed in the index cases by means of enhanced MRI (10), sonography (8), and mammography (7). Two of the 36 index cancers (5.6%) benefited from MRI assessment. Retroareolar cancer extension was observed with enhanced MRI in five index cancers. Of these, one was also noted on both a sonogram and a mammogram. Four of the index cancers (11.1%) benefited from the enhanced MRI. Overall, five index cancers (13.9%) benefited from the enhanced MRI. With a gold standard of histology, the mean cancer sizes were underestimated by sonography and overestimated by enhanced MRI. In comparison with sonography, a stronger association was noted between MRI and histological measurements, with coefficients of 0.657 and 0.882, respectively (P <.001). CONCLUSIONS: In a clinical setting, MRI for preoperative assessment of breast cancers is warranted. Minimally invasive, percutaneous core-needle biopsy did not alter the clinical efficacy of the MRI evaluation. PMID- 15289240 TI - Rapid genetic diagnosis with the transcription-reverse transcription concerted reaction system for cancer micrometastasis. AB - BACKGROUND: Detection of cancer micrometastases is required for improvement of cancer therapy. The aim of this study was to establish a rapid and practical genetic assay to detect micrometastasis in gastric cancer and to assess its clinical significance with respect to prognosis. METHODS: A novel RNA amplification system with transcription-reverse transcription concerted reaction (TRC) was introduced for quantitative detection of cancer-specific carcinoembryonic antigen messenger RNA. The sensitivity and quantitative aspects of the assay were assessed with the full-length carcinoembryonic antigen messenger RNA, a gastric cancer cell line (MKN-45), and metastatic lymph nodes obtained from patients with gastric cancer. Peritoneal lavage fluid specimens that were collected from gastric cancer surgery were subjected to the assay, and the clinical significance of the results was examined for prediction of recurrence and survival. RESULTS: The quantification, sensitivity, and reproducibility of the assay with the TRC reaction were equal to those of quantitative reverse transcriptase-polymerase chain reaction with LightCycler. The most important advantages of the assay were its simplicity and rapidity. Molecular diagnosis of peritoneal lavage fluid by the TRC reaction significantly correlated with depth of invasion, peritoneal metastasis, clinical stage, overall survival, and peritoneal recurrence-free survival. CONCLUSIONS: Molecular diagnosis of peritoneal lavage fluid with the TRC reaction could be a useful prognostic indicator for peritoneal recurrence and survival. Because the TRC reaction is more rapid and simpler than reverse transcriptase-polymerase chain reaction as a format for detecting RNA sequences, it may enhance the genetic diagnosis of cancer micrometastasis and may improve cancer therapy. PMID- 15289241 TI - Impact of 18F-fluoro-2-deoxy-D-glucose positron emission tomography (FDG-PET) in patients with biochemical evidence of recurrent or residual medullary thyroid cancer. AB - BACKGROUND: Conventional imaging such as with (99m)Tc(V)dimercaptosunnic acid (DMSA), (111)In-octreotide scintigraphy, computed tomography (CT), and magnetic resonance imaging (MRI) rarely localizes occult medullary thyroid cancer (MTC). The role of (18)F-fluoro-2-deoxy-D-glucose positron emission tomography (FDG-PET) is not well defined. The aim of this study was to examine the usefulness of postoperative FDG-PET in localizing MTC metastases. METHODS: FDG-PET was performed in 26 patients with elevated serum tumor markers after total thyroidectomy with central compartment dissection and additional neck dissection on indication. Patient- and lesion-based results were compared with the findings of conventional nuclear imaging and validated by morphological imaging (CT, MRI, ultrasonography), including bone scintigraphy and pathology when possible. Clinical impact was evaluated. RESULTS: FDG-PET detected foci in 50% of patients with lesion-based sensitivity of 96%. (111)In-octreotide detected foci in 19% with sensitivity of 41%, and (99m)Tc(V)DMSA scintigraphy and morphological imaging detected foci in 21% and 40%, respectively, with sensitivity of 57% and 87%. No lesions were found in 11 patients (42%). Positive FDG-PET findings led to surgical intervention in nine patients (35%). They all underwent surgery for removal of residual tumor or metastases. One patient achieved disease-free status. In all patients who underwent surgery, serum calcitonin levels were reduced by an average of 58 +/- 31%. CONCLUSIONS: For detection of occult MTC lesions, FDG-PET is superior to conventional nuclear imaging and is the best detection method yet available. FDG-PET in postoperative follow-up has clinical value and may be used for guiding reoperation and additional morphological imaging preoperatively. PMID- 15289242 TI - Combined effect of mothers' and fathers' mental health symptoms on children's behavioral and emotional well-being. AB - BACKGROUND: Few studies have examined the effects on children of maternal mental health symptoms other than depressive symptoms or have examined the joint effects of mothers' and fathers' mental health symptoms. OBJECTIVE: To examine whether the father's mental health symptoms may modify the association between the mother's mental health and the child's behavioral and emotional health. METHODS: Cross-sectional data from a national longitudinal survey of families provided information on 822 children aged 3 to 12 years who were living with both parents. The main child outcomes were the Behavior Problems Index-Externalizing (BPI-EXT) and-Internalizing (BPI-INT) subscales. The mother's and father's mental health were each assessed by self-report using the K10, a new, validated 10-item screen for serious mental illness, including mood or anxiety disorder. Parents with scores in the upper quartile were considered to be in poorer mental health and those with scores in the other 3 quartiles were considered to be in better mental health. RESULTS: Adjusted for covariates, having both parents in poorer mental health was associated with a 1-SD increase in the children's BPI-EXT scores (beta coefficient, 5.2; SE, 0.9; P<.001) compared with neither parent reporting poorer mental health. This effect was substantially weakened if the mother was in poorer mental health but the father was not (beta coefficient, 1.8; SE, 0.5; P<.01). There was no statistically significant effect if only the father was in poorer mental health (beta coefficient, 0.1; SE, 0.6; P =.88). The risk of a child having a high BPI-EXT score (>or=90th percentile for the cohort) was elevated if both parents reported poorer mental health (odds ratio, 9.2; 95% confidence interval [CI], 4.8-17.8), but was less elevated if only the mother reported poorer mental health (odds ratio, 2.3; 95% CI, 1.1-4.9), and was not elevated if only the father reported poorer mental health (odds ratio, 0.6; 95% CI, 0.2-1.9). Similar patterns emerged for children's BPI-INT scores. CONCLUSIONS: A father in better mental health may buffer the influence of a mother's poorer mental health on a child's behavioral and emotional problems, and these problems seem to be most severe for children who have 2 parents with poorer mental health. The form and intensity of pediatric approaches to mothers with poorer mental health may need to consider the mental health of fathers. PMID- 15289244 TI - Prevalence of school bullying in Korean middle school students. AB - BACKGROUND: School bullying is the most common type of school violence. Victimization by or perpetration of school bullying has frequently been associated with a broad spectrum of behavioral, emotional, and social problems. OBJECTIVE: To investigate the prevalence and demographic characteristics of victims, perpetrators, and victim-perpetrators in a Korean middle school sample. METHODS: We evaluated 1756 middle school students in this cross-sectional study. Students provided demographic information and completed the Korean-Peer Nomination Inventory. Descriptive statistics and the Pearson chi(2) test were used. RESULTS: We found that 40% of all children participated in school bullying. By category, the prevalence of victims, perpetrators, and victim-perpetrators was 14%, 17%, and 9%, respectively. The most common subtypes of victimization were exclusion (23%), verbal abuse (22%), physical abuse (16%), and coercion (20%). Boys were more commonly involved in both school bullying and all 4 types of victimization. The prevalence of bullying was greater in students with either high or low socioeconomic status and in nonintact families. CONCLUSIONS: School bullying is highly prevalent in Korean middle school students. Demographic characteristics can help identify students at greater risk for participation in school bullying. PMID- 15289243 TI - Cross-national consistency in the relationship between bullying behaviors and psychosocial adjustment. AB - OBJECTIVE: To determine whether the relationship between bullying and psychosocial adjustment is consistent across countries by standard measures and methods. DESIGN: Cross-sectional self-report surveys were obtained from nationally representative samples of students in 25 countries. Involvement in bullying, as bully, victim, or both bully and victim, was assessed. SETTING: Surveys were conducted at public and private schools throughout the participating countries. PARTICIPANTS: Participants included all consenting students in sampled classrooms, for a total of 113 200 students at average ages of 11.5, 13.5, and 15.5 years. MAIN OUTCOME MEASURES: Psychosocial adjustment dimensions assessed included health problems, emotional adjustment, school adjustment, relationships with classmates, alcohol use, and weapon carrying. RESULTS: Involvement in bullying varied dramatically across countries, ranging from 9% to 54% of youth. However, across all countries, involvement in bullying was associated with poorer psychosocial adjustment (P<.05). In all or nearly all countries, bullies, victims, and bully-victims reported greater health problems and poorer emotional and social adjustment. Victims and bully-victims consistently reported poorer relationships with classmates, whereas bullies and bully-victims reported greater alcohol use and weapon carrying. CONCLUSIONS: The association of bullying with poorer psychosocial adjustment is remarkably similar across countries. Bullying is a critical issue for the health of youth internationally. PMID- 15289245 TI - Six-year intervention outcomes for adolescent children of parents with the human immunodeficiency virus. AB - HYPOTHESIS: Having a parent with the human immunodeficiency virus has a significant negative impact on an adolescent child's adjustment. OBJECTIVE: To assess the adjustment of adolescent children to having a parent with the human immunodeficiency virus over 6 years, following the delivery of a coping skills intervention. DESIGN: A randomized controlled trial with repeated evaluations that was analyzed with an intention-to-treat analysis. A skill-based intervention was delivered in 3 modules over 24 sessions, with the third module being delivered only if parents died. SETTING AND PATIENTS: A representative sample of parents with the human immunodeficiency virus (n = 307) and their adolescent children (n = 423) was recruited from the Division of AIDS Services in New York City; 51.5% (n = 158) of the parents died. MAIN OUTCOME MEASURES: Employment and school enrollment, receiving public welfare support, early parenthood, mental health symptoms, and the quality of romantic relationships. RESULTS: Over 6 years, significantly more adolescents in the intervention condition than the control condition were employed or in school (82.58% vs 68.94%), were less likely to receive public welfare payments (25.66% vs 36.65%), were less likely to have psychosomatic symptoms (mean, 0.24 vs 0.31), were more likely to report better problem-solving and conflict resolution skills in their romantic relationships (mean score, 4.38 vs 4.20), expected to have a partner with a good job (mean, 4.57 vs 4.19), and expected to be married when parenting (mean, 3.05 vs 2.40). With marginal significance, the percentage of parents in the intervention condition (34.6%) was less than in the control condition (44.1%). CONCLUSION: Physicians must consider the psychosocial consequences of illness-related challenges on children and provide interventions. PMID- 15289246 TI - Adolescent psychiatric hospitalization and mortality, distress levels, and educational attainment: follow-up after 11 and 20 years. AB - BACKGROUND: Adolescents with early psychiatric hospitalization are likely to be at a significant risk for long-term difficulties. OBJECTIVE: To examine early adulthood outcomes of psychiatrically hospitalized adolescents. DESIGN: Inception cohort recruited from 1978 to 1981 and observed until 2002. SETTING: Northeastern United States. PARTICIPANTS: Adolescents (aged 12-15 years) from 2 matched cohorts were recruited and assessed repeatedly across 20 years: 70 psychiatrically hospitalized youths and 76 public high school students. MAIN OUTCOME MEASURES: Death, emotional distress, high school completion, and educational attainment. RESULTS: Psychiatrically hospitalized youths were significantly more likely to die and to report higher levels of emotional distress. Hospitalized youths were significantly less likely to graduate from high school and complete college and graduate school. CONCLUSIONS: The association between psychiatric symptoms sufficient to result in psychiatric hospitalization during adolescence and later mortality, emotional distress, high school completion, and educational attainment is striking. Further study is needed to identify and understand linkages between adolescent psychiatric impairment and decrements in adult functioning, particularly the processes that may underlie these linkages. Increasing school completion and educational attainment among hospitalized youths may minimize decrements in adult adaptation. PMID- 15289247 TI - New users of antipsychotic medications among children enrolled in TennCare. AB - BACKGROUND: The use of antipsychotic medications in children and adolescents for indications other than psychosis or Tourette syndrome is controversial. Newer atypical antipsychotics with profiles of adverse effects that differ from those of traditional antipsychotics may lead providers to prescribe antipsychotics more frequently than in the past for behavioral indications not strongly supported by clinical study. OBJECTIVE: To identify population-based new use of antipsychotics among patients aged 2 to 18 years. DESIGN: Retrospective cohort study, January 1, 1996, through December 31, 2001. SETTING: Tennessee's managed care program for Medicaid enrollees and the uninsured (TennCare). MAIN OUTCOME MEASURES: New use of antipsychotic medications and indications for use by the child's diagnosis, adjusted for age, sex, race, county of residence, enrollment category, and income. RESULTS: The proportion of TennCare children who were new users of antipsychotics, adjusted for demographic characteristics, nearly doubled from 23/10 000 in 1996 to 45/10 000 in 2001 (adjusted incidence rate ratio, 1.98; 95% confidence interval, 1.82-2.16). In 1996, 6.8% of new users received an atypical antipsychotic; by 2001, this had increased to 95.9%. New use for attention deficit/hyperactivity disorder and affective disorders increased 2.5-fold. New use of antipsychotics for schizophrenia, acute psychotic reaction, Tourette syndrome, and mental retardation or autism remained relatively constant. Secular trends of increasing use were most pronounced for those aged 6 to 12 years (93% increase) and 13 to 18 years (116% increase), although use among preschool children increased 61% during the study period. CONCLUSION: The proportion of TennCare children who became new users of antipsychotics nearly doubled from 1996 to 2001, with a substantial increase in use of antipsychotics for attention deficit/hyperactivity disorder, conduct disorder, and affective disorders. PMID- 15289248 TI - Prevalence of and risk factors for depressive symptoms among young adolescents. AB - OBJECTIVE: To determine the prevalence, risk factors, and risk behaviors associated with depressive symptoms in a nationally representative, cross sectional sample of young adolescents. DESIGN: A school-based survey collected through self-administered questionnaires in grades 6, 8, and 10 in 1996. SETTING: Schools in the United States. PARTICIPANTS: 9863 students in grades 6, 8, and 10 (average ages, 11, 13, and 15). MAIN OUTCOME MEASURES: Depressive symptoms, substance use, somatic symptoms, scholastic behaviors, and involvement in bullying. RESULTS: Eighteen percent of youths reported symptoms of depression. A higher proportion of females (25%) reported depressive symptoms than males (10%). Prevalence of depressive symptoms increased by age for both males and females. Among American Indian youths, 29% reported depressive symptoms, as compared with 22% of Hispanic, 18% of white, 17% of Asian American, and 15% of African American youths. Youths who were frequently involved in bullying, either as perpetrators or as victims, were more than twice as likely to report depressive symptoms than those who were not involved in bullying. A significantly higher percentage of youths who reported using substances reported depressive symptoms as compared with other youths. Similarly, youths who reported experiencing somatic symptoms also reported significantly higher proportions of depressive symptoms than other youths. CONCLUSIONS: Depression is a substantial and largely unrecognized problem among young adolescents that warrants an increased need and opportunity for identification and intervention at the middle school level. Understanding differences in prevalence between males and females and among racial/ethnic groups may be important to the recognition and treatment of depression among youths. PMID- 15289249 TI - Parent identification of early emerging child behavior problems: predictors of sharing parental concern with health providers. AB - OBJECTIVES: To better understand the predictors of parental discussions with pediatric care providers (pediatricians, psychologists/psychiatrists, social workers, early intervention providers, or other medical specialists) regarding early child behavior problems and to suggest strategies for eliciting early identification from parents in health care settings. DESIGN: A cross-sectional survey of parents of children from a representative healthy birth cohort. The survey included the Infant-Toddler Social Emotional Assessment, measurement of parental worry regarding problematic behavior, and demographic factors. SETTING: Fifteen urban and suburban towns in the northeastern United States. PARTICIPANTS: The study sample consisted of all parents of 11- to 39-month-olds (n = 269) who exceeded the 90th percentile on 1 or more Infant-Toddler Social Emotional Assessment problem domain scores (representing elevated problematic behavior symptoms) from an original sample of 1278. RESULTS: Few parents (17.7%) who reported elevated problematic behavior spoke to a provider about such problems. In adjusted models, speaking to a provider was associated with reported worry about behavior (odds ratio [OR], 3.47 [95% confidence interval (CI), 1.74-6.92]) and with low reported child social-emotional competence (OR, 2.68 [95% CI, 1.23 5.84]). In adjusted models, worry was most likely among parents who reported low child competence (OR, 2.18 [95% CI, 1.07-4.22]) and disruption in family routines attributed to the child's behavior (OR, 2.38 [95% CI, 1.31-4.33]). CONCLUSIONS: Parental worry is a robust predictor of help seeking among parents of children with behavioral problems. Further, lags in social competence contribute to both parental worry and help seeking. These findings, in conjunction with previous evidence that child behavior problems amenable to early intervention are often unidentified, suggest that systematic inquiry by health care providers about parental concerns is important in the identification of early emerging behavioral health problems. PMID- 15289250 TI - Age effects on antidepressant-induced manic conversion. AB - BACKGROUND: Antidepressant drug therapy can precipitate mania in vulnerable individuals, but little is known about the effects of age on this phenomenon. OBJECTIVE: To pharmacoepidemiologically evaluate the risk of conversion to mania by antidepressant class and patient age. DESIGN, SETTING, AND PATIENTS: Using an administrative national database of more than 7 million privately insured individuals, linked outpatient and pharmacy claims were analyzed for mental health users aged 5 to 29 years (N = 87,920). MAIN OUTCOME MEASURES: The proportion and cumulative hazard of manic conversion were analyzed by antidepressant class and subject age among children, adolescents, and young adults with an anxiety or nonbipolar mood disorder in the United States between January 1, 1997, and December 31, 2001. Manic conversion was defined as a new diagnosis of bipolar illness. RESULTS: During median follow-up of 41 weeks (range, 8-251 weeks), manic conversion occurred in 4786 patients (5.4%). Multivariate analyses using time-dependent Cox proportional hazards models indicated that an increased risk of manic conversion was associated with antidepressant category vs no antidepressant exposure (hazard ratios: 2.1 for selective serotonin reuptake inhibitors, P<.001; 3.8 for "other" antidepressants, P<.001; and 3.9 for tricyclic antidepressants, P =.002). Antidepressant x age interactions revealed inverse age effects for selective serotonin reuptake inhibitors and other antidepressants (beta = -.05; P<.001 for both) but not for tricyclic antidepressants (beta = -.02; P =.25). Peripubertal children exposed to antidepressants were at highest risk of conversion (number needed to harm: 10 [95% confidence interval, 9-12] among 10- to 14-year-olds vs 23 [95% confidence interval, 21-25] among 15- to 29-year-olds). CONCLUSIONS: Patient age is an effect modifier on the risk of antidepressant-associated manic conversion. Treatment with antidepressants is associated with highest conversion hazards among children aged 10 to 14 years. PMID- 15289251 TI - Mental illness hospitalizations of youth in Washington State. AB - OBJECTIVE: To determine if mental health hospitalizations have increased among youth. DESIGN: A retrospective cross-sectional time trend study. The Washington State Comprehensive Hospital Abstract Reporting System data set was used to examine hospitalizations among youth (aged 5-19 years) from January 1, 1990, through December 31, 1999. The yearly rates of youth hospitalized for mental illness were calculated, as were the proportions of hospitalizations due to mental illness. Chi(2) tests of trend were computed to assess for significant change over time. Additional analyses examined trends in hospital days due to mental illness and repeated hospitalizations and compared mental illness with other major causes of child and adolescent hospitalization. RESULTS: The rate of school-aged children (aged 5-14 years) hospitalized for mental illness increased by 22% during the 1990s (P =.004). The proportion of hospitalizations due to mental illness in school-aged children increased from 7.8% in 1990 to 12.8% in 1999 (P<.001). Among adolescents (aged 15-19 years), no significant change occurred in the rate of mental illness hospitalizations, but the proportion of hospitalizations due to mental illness increased from 14.5% in 1990 to 21.5% in 1999 (P<.001). Although injuries were the leading cause of hospitalizations among youth in 1990, mental illness has since surpassed injuries as a cause for hospitalization. Mental illness accounted for one third of all hospital days for youth in 1999. CONCLUSIONS: Mental illness hospitalizations account for an increasing proportion of admissions and hospital days among children and adolescents in Washington State. During the past decade, mental illness has surpassed injury as a leading cause of hospitalization for Washington youth. PMID- 15289252 TI - Efficacy of treatment for child and adolescent traumatic stress. AB - BACKGROUND: Despite the expenditure of large sums of public monies to ameliorate the consequences of childhood trauma, little is known about the efficacy of treatment for traumatized children and their families. OBJECTIVE: To review the efficacy of treatment for child and adolescent traumatic stress. DATA SOURCES: An extensive literature search identified 102 studies addressing child and adolescent trauma treatment. STUDY SELECTION: Only 8 studies met the minimal inclusion criteria of (1) using a comparison group and (2) including symptoms of traumatic stress as a treatment outcome. DATA EXTRACTION: These studies are critically evaluated for adherence to standards of good efficacy research using formal criteria of treatment research quality. DATA SYNTHESIS: Treatment for traumatic stress appears to lead to greater improvement than either no treatment or routine community care. CONCLUSIONS: Child and adolescent posttraumatic stress disorder treatment research lags behind both adult posttraumatic stress disorder treatment research and other child treatment research. There is considerable need to establish a programmatic approach to developing evidence-based child trauma treatment. Barriers to conducting child trauma treatment research include sensitivity to the rights of victims and child service models that perceive research as intruding on vulnerable children at critically sensitive points in their development. PMID- 15289253 TI - Correlations between family meals and psychosocial well-being among adolescents. AB - OBJECTIVE: To determine the association between frequency of family meals and multiple indicators of adolescent health and well-being (tobacco, alcohol, and marijuana use; academic performance; self-esteem; depressive symptoms; and suicide involvement) after controlling for family connectedness. METHODS: Data come from a 1998-1999 school-based survey of 4746 adolescents from ethnically and socioeconomically diverse communities in the Minneapolis/St Paul, Minn, metropolitan area. Logistic regression, controlling for family connectedness and sociodemographic variables, was used to identify relationships between family meals and adolescent health behaviors. RESULTS: Approximately one quarter (26.8%) of respondents ate 7 or more family meals in the past week, and approximately one quarter (23.1%) ate family meals 2 times or less. Frequency of family meals was inversely associated with tobacco, alcohol, and marijuana use; low grade point average; depressive symptoms; and suicide involvement after controlling for family connectedness (odds ratios, 0.76-0.93). CONCLUSIONS: Findings suggest that eating family meals may enhance the health and well-being of adolescents. Public education on the benefits of family mealtime is recommended. PMID- 15289254 TI - Headache, stomachache, backache, and morning fatigue among adolescent girls in the United States: associations with behavioral, sociodemographic, and environmental factors. AB - BACKGROUND: Data on the prevalence and co-occurrence of multiple somatic symptoms among US adolescent females as they are influenced by sociodemographic, behavioral, and environmental factors is limited. OBJECTIVES: To describe the health status of adolescent US females measured by the prevalence, frequency, and co-occurrence of headache, stomachache, backache, and morning fatigue and to investigate associations between selected risk and protective factors. DESIGN, SETTING, AND PARTICIPANTS: School-based, cross-sectional, nationally representative survey of adolescents in the 6th through 10th grades in the US. Data collected between 1997 and 1998. MAIN OUTCOME MEASURES: Prevalence of headache, stomachache, backache, and morning fatigue. RESULTS: Among US adolescent girls, 29.1% experience headaches, 20.7% report stomachaches, 23.6% experience back pain, and 30.6% report morning fatigue at the rate of more than once a week. Co-occurrence of somatic complaints is common. Among girls who experienced headaches more than once a week, 3.2 million (53.3%) also reported stomach pain more than once a week and 4.1 million (74.3%) reported morning fatigue more than once a week. Heavy alcohol use, high caffeine intake, and smoking cigarettes every day were strongly associated with all symptoms, while parent and teacher support served as protective factors. CONCLUSIONS: Somatic complaints of headache, stomachache, backache, and morning fatigue are common among US adolescent girls and co-occur often. Effective clinical treatment of this population requires comprehensive assessment of all female adolescents presenting with seemingly isolated somatic complaints. PMID- 15289255 TI - Childhood overweight and parent- and teacher-reported behavior problems: evidence from a prospective study of kindergartners. AB - OBJECTIVES: To determine if there is a relationship between overweight and behavior problems among children as young as 5 years old by studying the association between overweight and behavioral health at entry into kindergarten and to determine whether overweight status is a risk factor for the onset of new behavior problems during the first 2 years in school. DESIGN: We use data from a nationally representative sample of kindergartners in the United States-the Early Childhood Longitudinal Study-Kindergarten class. Data on height, weight, and parent- and teacher-reported behavior problems were collected 3 times during their first 2 years in school for 9949 children. We use a multivariate regression analysis that controls for sociodemographic characteristics, parent-child interaction, birth weight, and mother's mental health. RESULTS: Among girls, but not boys, there is a significant association between overweight and teacher reported externalizing behavior problems (odds ratio [OR], 1.81; 95% confidence interval [CI], 1.23-2.68), teacher-reported internalizing behavior problems (OR, 1.54; 95% CI, 1.09-2.17), and parent-reported internalizing behavior problems (OR, 1.49; 95% CI, 1.08-2.06) at the beginning of kindergarten. However, overweight status was not a risk factor for the onset of new behavior problems over time for either girls (teacher-reported externalizing behavior problems: OR, 0.58 [95% CI, 0.25-1.33]; teacher-reported internalizing behavior problems: OR, 1.34 [95% CI, 0.88-2.03]; and parent-reported internalizing behavior problems: OR, 1.29 [95% CI, 0.82-2.01]) or boys (teacher-reported externalizing behavior problems: OR, 1.02 [95% CI, 0.67-1.57]; teacher-reported internalizing behavior problems: OR, 1.02 [95% CI, 0.68-1.52]; and parent-reported internalizing behavior problems: OR, 1.42 [95% CI, 0.94-2.15]), whereas low family income and maternal depression were strong predictors of such problems. CONCLUSIONS: Childhood overweight is already associated with behavior problems when girls start school, but not boys. In contrast to common belief, overweight status does not predict the onset of new internalizing or externalizing behavior problems during the first 2 years of school. PMID- 15289256 TI - Identification and management of psychosocial problems among toddlers in Dutch preventive child health care. AB - OBJECTIVES: To assess the degree to which preventive child health professionals (CHPs) identify and manage psychosocial problems among preschool children in the general population and to determine the association with parent-reported behavioral and emotional problems, sociodemographic factors, and mental health history of children. DESIGN: The CHPs examined the child and interviewed the parents and child during their routine health assessments. The Child Behavior Checklist (CBCL) was completed by the parents. SETTING: Sixteen child health care services across the Netherlands that routinely provided well-child care to nearly all preschool children. PATIENTS: Of 2354 children aged 21 months to 4 years who were eligible for a routine health assessment, 2229 (94.7%) participated. MAIN OUTCOME MEASURES: Identification and management of psychosocial problems by CHPs. RESULTS: In 9.4% of all children, CHPs identified psychosocial problems. Two in 5 of the CHP-identified children were referred for additional diagnosis and treatment. Identification of psychosocial problems and subsequent referral were much more likely in children with a clinical CBCL total problems score than in others (identification: 29% vs 7%; odds ratio [95% confidence interval], 5.40 [3.45-8.47]; referral: 15% vs 3%; odds ratio [95% confidence interval], 6.50 [3.69-11.46]). CONCLUSIONS: The CHPs frequently identify psychosocial problems in preschool children, although less than among school-aged children, but they miss many cases of parent-reported problems as measured by a clinical CBCL score. This general population study shows substantial room for improvement in the early identification of psychosocial problems. PMID- 15289258 TI - The rewards of reducing risk. PMID- 15289257 TI - The relation between physical activity and mental health among Hispanic and non Hispanic white adolescents. AB - OBJECTIVE: To assess the relation of physical activity (PA) with feelings of sadness and suicidal thoughts and behaviors among Hispanic and non-Hispanic white adolescent boys and girls. DESIGN: Cross-sectional study using a modified 2001 Youth Risk Behavior Survey. PARTICIPANTS: One thousand eight hundred seventy Hispanic and non-Hispanic white adolescents, aged 14 to 18 years, attending high school in Nueces County, Texas. MAIN OUTCOME MEASURE: Logistic regression analysis was used to assess the relation between PA, including moderate and vigorous PAs, strength and toning, total PA, physical education class, and participation in team sports, and the dependent variables feelings of sadness and considering, planning, and attempting suicide. RESULTS: More boys reported participating in PA than girls (P<.001), and more girls than boys reported feelings of sadness and considering and planning suicide (P<.001). Greater attendance in physical education class was inversely related to feelings of sadness (odds ratio [OR], 0.80 [95% confidence interval (CI), 0.68-0.94]); participation in more total PA sessions per week was associated with a lower risk of considering suicide (OR, 0.72 [95% CI, 0.65-0.79]); and higher levels of vigorous PA (OR, 0.73 [95% CI, 0.57-0.93]), total PA (OR, 0.65 [95% CI, 0.48 0.87]), and strength and toning activity (OR, 0.64 [95% CI, 0.42-0.99]) were associated with a lower risk of planning suicide. CONCLUSIONS: These findings are consistent with a beneficial effect of PA on feelings of sadness and suicidal behaviors in Hispanic and non-Hispanic white boys and girls. Physical activity may be considered as part of an intervention strategy to improve adolescent health as a whole. PMID- 15289259 TI - Mental health and obesity in pediatric primary care: a gap between importance and action. PMID- 15289260 TI - Prescribing more psychotropic medications for children: what does the increase mean? PMID- 15289261 TI - International trends in bullying and children's health: giving them due consideration. PMID- 15289262 TI - Integrating pediatrics and mental health: the reality is in the relationships. PMID- 15289264 TI - Impaired recognition of anger following damage to the ventral striatum. AB - Comparative neuropsychology has identified a role for the ventral striatum (VS) in certain forms of aggression. To address whether the homologous region in humans also contributes to the emotion anger, we studied a case series of four human subjects with focal lesions affecting the VS. All four demonstrated a disproportionate impairment in recognizing human signals of aggression. By contrast, a control group of individuals with damage to more dorsal basal ganglia (BG) regions showed no evidence of an anger impairment. Our findings demonstrate that the VS makes a significant contribution to coding signals of aggression in humans, and emphasize the importance of an approach to human affective neuroscience based on cross-species homologies. The results are discussed in relation to the ventral striatal dopamine system's role in the pursuit of biological resources in general. We propose that the role of the VS in the recognition of human signals of anger may reflect a more general role in the coordination of behaviour relevant to the acquisition and protection of valued resources, including detection of signals of conspecific challenge (anger). PMID- 15289265 TI - Notch1 and Jagged1 are expressed after CNS demyelination, but are not a major rate-determining factor during remyelination. AB - The reasons for the eventual failure of repair mechanisms in multiple sclerosis are unknown. The presence of precursor and immature oligodendrocytes in some non repairing lesions suggests a mechanism in which these cells either receive insufficient differentiation signals or are exposed to differentiation inhibitors. Jagged signalling via Notch receptors on oligodendrocyte precursor cells (OPCs) inhibits their differentiation during development and the finding that both notch and jagged are expressed in multiple sclerosis lesions has fostered the view that this signalling pathway may explain remyelination failure. In this study, we show that Notch1 is expressed on adult OPCs and that there are multiple cellular sources of its ligand Jagged1 in a rodent model of remyelination. However, despite their expression, the lesions undergo complete remyelination. To establish whether Notch-jagged signalling regulates the rate of remyelination we compared their expression profiles in young animals with those in older animals, where remyelination occurs more slowly, but could find no correlation between expression and remyelination rate. Finally we found that OPC targeted Notch1 ablation in cuprizone-treated Plp-creER Notch1(lox/lox) transgenic mice yielded no significant differences in remyelination parameters between knock-out and control mice. Thus, in contrast to developmental myelination, adult expression of Notch1 and Jagged1 neither prevents nor plays a major rate-determining role in remyelination. More generally, the re-expression of developmentally expressed genes following injury in the adult does not per se imply similar function. PMID- 15289266 TI - MRI prognostic factors for relapse after acute CNS inflammatory demyelination in childhood. AB - The prognostic factors for relapse of the initial MRI findings after a first episode of acute CNS inflammatory demyelination are unclear in children. In this study we aimed to identify initial MRI factors that are predictive of a second attack and disability after a first episode of acute CNS inflammatory demyelination in childhood. A cohort of 116 children who had a first episode of acute CNS inflammatory demyelination between 1990 and 2002 was studied using survival analysis methods. The initial MRI data were reviewed in a systematic, standardized, double-blind manner. The average follow-up was 4.9 +/- 3 years. Multivariate analysis showed that the rate of second attack was higher in patients with corpus callosum long axis perpendicular lesions (34 out of 116 patients, 30%) on the initial MRI [hazard ratio (HR) 2.89; 95% confidence interval (CI) 1.65-5.06] and/or with the sole presence of well-defined lesions (46 out of 116 patients, 40%) (HR 1.71; 95% CI 1.29-2.27). Both criteria were more specific predictors (100%) of relapse, demonstrating conversion to multiple sclerosis, than the three Barkhof criteria (63%), but were less sensitive (21% compared with 52%). None of the MRI criteria was predictive of severe disability. Using initial MRI and survival analysis methods, we identified two specific predictors of relapse and conversion to multiple sclerosis after a first episode of acute CNS inflammatory demyelination in childhood. Their low sensitivity, however, shows that this prediction remains difficult. PMID- 15289267 TI - Follow-up study and response to treatment in 23 patients with Lewis-Sumner syndrome. AB - Lewis-Sumner syndrome (LSS) is a dysimmune peripheral nerve disorder, characterized by a predominantly distal, asymmetric weakness mostly affecting the upper limbs with sensory impairment, and by the presence of multifocal persistent conduction blocks. The nosological position of this neuropathy in relation to multifocal motor neuropathy (MMN) and chronic inflammatory demyelinating polyradiculoneuropathy (CIDP) is still debated. We report the clinical, biological and electrophysiological features, the course and the response to treatment in 23 LSS patients. The initial symptoms started in the distal part of an upper limb in 70% of patients. They were sensorimotor in 65% and purely sensory in 35% of patients. A cranial nerve involvement was observed in 26% of patients and a distal limb amyotrophy in 52%. The CSF protein level was normal in 67% of patients and mildly elevated in the remainder. None had serum anti-GM1 antibodies. There were multiple motor conduction blocks (average of 2.87/patient), predominantly located in the forearm, whereas demyelinating features outside the blocked nerves were rare. Abnormal distal sensory potentials were found in 87% of patients. The electrophysiological pattern suggests a very focal motor fibre demyelination sparing the nerve endings, whereas sensory fibre involvement was widespread. The course was chronic progressive in 71% of patients and relapsing-remitting in the others. During the follow-up study (median duration of 4 years), half of the patients progressed with a multifocal pattern and the distribution of the motor deficit remained similar to the initial presentation. The other patients showed a progression to the other limbs, suggesting a more diffuse process. Fifty-four percent of the patients treated with intravenous immunoglobulin showed an improvement, compared with 33% of the patients treated with oral steroids. Overall, 73% of patients had a positive response to immune-mediated therapy. LSS may be distinguished from MMN by the presence of sensory involvement, the absence of serum anti-GM1 antibodies and, in some cases, a positive response to steroids. In some of the patients in our study, LSS evolved into a more diffuse neuropathy sharing similarities with CIDP. Others had a clinical course characterized by a striking multifocal neuropathy, which suggests underlying mechanisms different from CIDP. Overall, whatever the clinical course, LSS responded to immune-mediated treatment in a manner similar to CIDP. PMID- 15289268 TI - Corpus callosum size and very preterm birth: relationship to neuropsychological outcome. AB - Thinning of the corpus callosum (CC) is often observed in individuals who were born very preterm. Damage to the CC during neurodevelopment may be associated with poor neuropsychological performance. This study aimed to explore any evidence of CC pathology in adolescents aged 14-15 years who were born very preterm, and to investigate the relationship between CC areas and verbal skills. Seventy-two individuals born before 33 weeks of gestation and 51 age- and sex matched full-term controls received structural MRI and neuropsychological assessment. Total CC area in very preterm adolescents was 7.5% smaller than in controls, after adjusting for total white matter volume (P = 0.015). The absolute size of callosal subregions differed between preterm and full-term adolescents: preterm individuals had a 14.7% decrease in posterior (P < 0.0001) and an 11.6% decrease in mid-posterior CC quarters (P = 0.029). Preterm individuals who had experienced periventricular haemorrhage and ventricular dilatation in the neonatal period showed the greatest decrease in CC area. In very preterm boys only, verbal IQ and verbal fluency scores were positively associated with total mid-sagittal CC size and mid-posterior surface area. These results suggest that very preterm birth adversely affects the development of the CC, particularly its posterior quarter, and this impairs verbal skills in boys. PMID- 15289269 TI - Anti-disialoside antibodies kill perisynaptic Schwann cells and damage motor nerve terminals via membrane attack complex in a murine model of neuropathy. AB - Anti-disialoside antibodies (Abs) that bind NeuAc(alpha2-8) NeuAc epitopes on GQ1b and related gangliosides are found in human autoimmune neuropathy sera and are considered to be pathogenic. In a model system in mice, one mechanism by which anti-disialoside Abs have been demonstrated to induce paralysis is through a complement dependent blocking effect on transmitter release at the neuromuscular junction, similar to the effects of alpha-latrotoxin. Although direct targeting of presynaptic neuronal membranes occurs in this model, concomitant injury to perisynaptic Schwann cells (pSC) could indirectly contribute to this paralytic effect by influencing nerve terminal function and survival. To examine this possibility and the specific complement components that might mediate these effects, we exposed neuromuscular junctions in vivo and in vitro to an anti-disialoside Ab in conjunction with intact and selectively deficient complement sources. Using immuno-electron microscopy, we observed Ab deposits equally distributed on both neuronal and pSC membranes, and ultrastructural evidence of injury at both sites. Presynaptic neuronal injury was demonstrated functionally with microelectrode recordings and histologically as neurofilament loss. As hypothesized, concomitant pSC injury occurred, as indicated by abnormal uptake of ethidium dimer into pSC nuclei. The pSC and nerve terminal damage indicators correlated well with deposition of the pore-forming terminal complement component, membrane attack complex (MAC) in pSC and nerve terminal membranes. Furthermore, both neuronal and pSC injury were exacerbated in tissues from mice lacking the inhibitory complement regulator, CD59, where MAC formation is increased. These data demonstrate that both presynaptic neuronal membranes and pSCs are targets for anti-disialoside Abs, and that the injury to both sites is mediated by MAC and further regulated by CD59. This is the first demonstration that complement mediated pSC injury occurs in a model of autoimmune neuropathy and provides a rationale for investigating the possibility of pSC injury in equivalent conditions in man. PMID- 15289270 TI - Neurofibromatosis 1-associated neuropathies: a reappraisal. AB - Neurofibromatosis 1 (NF1) is a common disease which is a source of various multisystemic manifestations related either to the accumulation of neurofibromas or to specific developmental abnormalities. The neurofibroma is the hallmark lesion of NF1 and develops from peripheral nerves. However, to date, the description of peripheral neuropathies of NF1 has not been investigated. To examine this question, we have evaluated 688 NF1 patients for the presentation, prognosis and associated morbidity of peripheral neuropathies in two hospital based series. We collected 18 patients (four women and 14 men) with diffuse peripheral neuropathy (2.3%). Eight patients had a paucisymptomatic or an asymptomatic neuropathy detected only on electrophysiological study, two had minor sensory manifestations, five had moderate motor and sensory manifestations and three had severe motor and sensory manifestations. Superimposed radicular changes were observed in seven cases. Two patients had a subacute and 16 a chronic polyneuropathy. Fourteen patients had a demyelinating neuropathy with either severe axonal changes (three), moderate or minor axonal changes (four) or no axonal changes (seven). Four patients had axonal neuropathies. There was a strong association between the presence of a peripheral neuropathy and large root diffuse neurofibromas (P < 0.03) and subcutaneous neurofibromas (P < 0.0001). Severe morbidity and mortality of patients with NF1 and peripheral neuropathies was 50%, much higher than what is observed in the general population of patients with NF1, and 100% in patients with the most severe symptoms and electrophysiological changes (demyelination with severe axonal features). Four patients out of 18 (22%) developed a malignant peripheral nerve sheath tumour (MPNST), a much higher proportion than in the whole population of NF1. Two patients died. Peripheral neuropathy constitutes a potentially severe complication in patients with NF1 associated with a frequent morbidity related to spinal complications and MPNSTs. Association of proximal large neurofibromas, peripheral neuropathies and subcutaneous neurofibromas may constitute a phenotype of NF1 with a severe prognosis. PMID- 15289271 TI - Accelerated diabetic neuropathy in axons without neurofilaments. AB - Diabetic neuropathy is characterized by slowing of conduction velocity and axonal atrophy. Both of these cardinal features of neuropathy might be linked to impaired neurofilament investment of axons. Since neurofilaments form the critical structural latticework of axons, their importance in neuropathy is of interest. We tested directly the relationship of neurofilaments to diabetic neuropathy by superimposing streptozotocin-generated diabetes on a unique but viable transgenic mouse described by Eyer and Peterson. These mice express a fusion protein in which the carboxyl terminus of the high molecular weight neurofilament protein (Nf-H) was replaced by beta-galactosidase, in turn blocking normal neurofilament export and rendering axons completely lacking neurofilaments. Despite similar levels of hyperglycaemia, diabetic mice lacking neurofilaments developed progressive slowing of conduction velocity in their motor and sensory fibres between 4 and 8 weeks after the onset of diabetes (P < 0.05), unlike diabetic mice with normal neurofilaments, who developed only mild evidence of neuropathy over the same time-frame. Diabetic mice without neurofilaments, but not those with neurofilaments, had a progressive decline in the amplitude of the caudal nerve compound action potential and there were trends toward increased axonal atrophy in diabetics lacking neurofilaments. Single daily doses of insulin that restored normoglycaemia (0.1 IU subcutaneous insulin daily 5 of 7 days weekly for 4 weeks) reversed conduction slowing and restored sensory axon calibre. Our findings indicate that abnormalities in neurofilament export or transport alone cannot account for features of diabetic neuropathy. Instead, neurofilaments may allow axons to better resist the ravages of diabetes. Our findings also confirm the impact of insulin on reversing the phenotype. PMID- 15289272 TI - The human spinal cord interprets velocity-dependent afferent input during stepping. AB - We studied the motor response to modifying the rate of application of sensory input to the human spinal cord during stepping. We measured the electromyographic (EMG), kinematic and kinetic patterns of the legs during manually assisted or unassisted stepping using body weight support on a treadmill (BWST) in eight individuals with spinal cord injury (SCI). At various treadmill speeds (0.27-1.52 m/s), we measured the EMG activity of the soleus (SOL), medial gastrocnemius (MG), tibialis anterior (TA), medial hamstrings (MH), vastus lateralis (VL), rectus femoris (RF) and iliopsoas (ILIO); the hip, knee and ankle joint angles; the amount of body weight support (BWS); and lower limb loading. The EMG amplitude and burst duration of the SOL, MG, TA, MH, VL, RF and ILIO were related to the step cycle duration during stepping using BWST. EMG mean amplitudes increased at faster treadmill speeds, and EMG burst durations shortened with decreased step cycle durations. Muscle stretch of an individual muscle could not account for the EMG amplitude modulation in response to stepping speed. The effects on the EMG amplitude and burst duration were similar in subjects with partial and no detectable supraspinal input. We propose that the human spinal cord can interpret complex step-related, velocity-dependent afferent information to contribute to the neural control of stepping. PMID- 15289273 TI - Corticosteroids for severe sepsis and septic shock: a systematic review and meta analysis. AB - OBJECTIVE: To assess the effects of corticosteroids on mortality in patients with severe sepsis and septic shock. DATA SOURCES: Randomised and quasi-randomised trials of corticosteroids versus placebo (or supportive treatment alone) retrieved from the Cochrane infectious diseases group's trials register, the Cochrane central register of controlled trials, Medline, Embase, and LILACS. REVIEW METHOD: Two pairs of reviewers agreed on eligibility of trials. One reviewer entered data on to the computer and four reviewers checked them. We obtained some missing data from authors of trials and assessed methodological quality of trials. RESULTS: 16/23 trials (n = 2063) were selected. Corticosteroids did not change 28 day mortality (15 trials, n = 2022; relative risk 0.92, 95% confidence interval 0.75 to 1.14) or hospital mortality (13 trials, n = 1418; 0.89, 0.71 to 1.11). There was significant heterogeneity. Subgroup analysis on long courses (> or = 5 days) with low dose (< or = 300 mg hydrocortisone or equivalent) corticosteroids showed no more heterogeneity. The relative risk for mortality was 0.80 at 28 days (five trials, n = 465; 0.67 to 0.95) and 0.83 at hospital discharge (five trials, n = 465, 0.71 to 0.97). Use of corticosteroids reduced mortality in intensive care units (four trials, n = 425, 0.83, 0.70 to 0.97), increased shock reversal at 7 days (four trials, n = 425; 1.60, 1.27 to 2.03) and 28 days (four trials, n = 425, 1.26, 1.04 to 1.52) without inducing side effects. CONCLUSIONS: For all trials, regardless of duration of treatment and dose, use of corticosteroids did not significantly affect mortality. With long courses of low doses of corticosteroids, however, mortality at 28 days and hospital morality was reduced. PMID- 15289274 TI - Dead mother I. PMID- 15289275 TI - Neural and behavioral responses to tryptophan depletion in unmedicated patients with remitted major depressive disorder and controls. AB - CONTEXT: An instructive paradigm for investigating the relationship between brain serotonin function and major depressive disorder (MDD) is the response to tryptophan depletion (TD) induced by oral loading with all essential amino acids except the serotonin precursor tryptophan. OBJECTIVE: To determine whether serotonin dysfunction represents a trait abnormality in MDD in the context of specific neural circuitry abnormalities involved in the pathogenesis of MDD. DESIGN: Randomized double-blind crossover study. SETTING: Outpatient clinic. PARTICIPANTS: Twenty-seven medication-free patients with remitted MDD (18 women and 9 men; mean +/- SD age, 39.8 +/- 12.7 years) and 19 controls (10 women and 9 men; mean +/- SD age, 34.4 +/- 11.5 years). INTERVENTIONS: We induced TD by administering capsules containing an amino acid mixture without tryptophan. Sham depletion used identical capsules containing hydrous lactose. Fluorodeoxyglucose F 18 positron emission tomography studies were performed 6 hours after TD. Magnetic resonance images were obtained for all participants. MAIN OUTCOME MEASURES: Quantitative positron emission tomography of regional cerebral glucose utilization to study the neural effects of sham depletion and TD. Behavioral assessments used a modified (24-item) version of the Hamilton Depression Rating Scale. RESULTS: Tryptophan depletion induced a transient return of depressive symptoms in patients with remitted MDD but not in controls (P<.001). Compared with sham depletion, TD was associated with an increase in regional cerebral glucose utilization in the orbitofrontal cortex, medial thalamus, anterior and posterior cingulate cortices, and ventral striatum in patients with remitted MDD but not in controls. CONCLUSION: The pattern of TD-induced regional cerebral glucose utilization changes in patients with remitted MDD suggests that TD unmasks a disease-specific, serotonin system-related trait dysfunction and identifies a circuit that probably plays a key role in the pathogenesis of MDD. PMID- 15289276 TI - Serologic evidence of prenatal influenza in the etiology of schizophrenia. AB - CONTEXT: Some, but not all, previous studies suggest that prenatal influenza exposure increases the risk of schizophrenia. These studies used dates of influenza epidemics and maternal recall of infection to define influenza exposure, suggesting that discrepant findings may have resulted from exposure misclassification. OBJECTIVE: To examine whether serologically documented prenatal exposure to influenza increases the risk of schizophrenia. DESIGN: Nested case-control study of a large birth cohort, born from 1959 through 1966, and followed up for psychiatric disorders 30 to 38 years later. SETTING: Population-based birth cohort. PARTICIPANTS: Cases were 64 birth cohort members diagnosed as having schizophrenia spectrum disorders (mostly schizophrenia and schizoaffective disorder). Controls were 125 members of the birth cohort, had not been diagnosed as having a schizophrenia spectrum or major affective disorder, and were matched to cases on date of birth, sex, length of time in the cohort, and availability of maternal serum. MAIN OUTCOME MEASURES: Archived maternal serum was assayed for influenza antibody in pregnancies giving rise to offspring with schizophrenia and matched control offspring. RESULTS: The risk of schizophrenia was increased 7-fold for influenza exposure during the first trimester. There was no increased risk of schizophrenia with influenza during the second or third trimester. With the use of a broader gestational period of influenza exposure-early to midpregnancy-the risk of schizophrenia was increased 3-fold. The findings persisted after adjustment for potential confounders. CONCLUSIONS: These findings represent the first serologic evidence that prenatal influenza plays a role in schizophrenia. If confirmed, the results may have implications for the prevention of schizophrenia and for unraveling pathogenic mechanisms of the disorder. PMID- 15289277 TI - Anomalous prefrontal-subcortical activation in familial pediatric bipolar disorder: a functional magnetic resonance imaging investigation. AB - BACKGROUND: The neurobiological features of pediatric bipolar disorder (BD) are largely unknown. Children and adolescents with BD may be important to study with functional neuroimaging techniques because of their unique status of early-onset BD and high familial loading for the disorder. Neuroimaging studies of adults with BD have implicated the dorsolateral prefrontal cortex (DLPFC) and anterior cingulate cortex (ACC) in the development of this disorder. OBJECTIVES: To study children and adolescents with BD via functional magnetic resonance imaging using cognitive and affective tasks and to examine possible abnormalities in the DLPFC and ACC, as well as selected subcortical areas, in pediatric familial BD. DESIGN: We evaluated 12 male subjects aged 9 to 18 years with BD who had at least 1 parent with BD as well as 10 age- and IQ-matched healthy male controls. Stimulants were discontinued for at least 24 hours; other medications were continued. Subjects underwent functional magnetic resonance imaging at 3 T while performing a 2-back visuospatial working memory task and an affective task involving the visualization of positively, neutrally, or negatively valenced pictures. SETTING: An academic referral setting, drawing from the Bay Area of San Francisco, Calif. RESULTS: Compared with controls, for the visuospatial working memory task, subjects with BD had greater activation in several areas including the bilateral ACC, left putamen, left thalamus, left DLPFC, and right inferior frontal gyrus. Controls had greater activation in the cerebellar vermis. In viewing negatively valenced pictures, subjects with BD had greater activation in the bilateral DLPFC, inferior frontal gyrus, and right insula. Controls had greater activation in the right posterior cingulate gyrus. For positively valenced pictures, subjects with BD had greater activation in the bilateral caudate and thalamus, left middle/superior frontal gyrus, and left ACC, whereas controls had no areas of greater activation. CONCLUSIONS: Children and adolescents with BD may have underlying abnormalities in the regulation of prefrontal-subcortical circuits. Further functional magnetic resonance imaging studies of attention and mood with greater sample sizes are needed. PMID- 15289278 TI - Ventromedial prefrontal cortex and amygdala dysfunction during an anger induction positron emission tomography study in patients with major depressive disorder with anger attacks. AB - CONTEXT: Although a variety of functional neuroimaging studies have used emotion induction paradigms to investigate the neural basis of anger in control subjects, no functional neuroimaging studies using anger induction have been conducted in patient populations. OBJECTIVE: To study the neural basis of anger in unmedicated patients with major depressive disorder with anger attacks (MDD + A), unmedicated patients with MDD without anger attacks (MDD - A), and controls. DESIGN: We used positron emission tomography, psychophysiologic measures, and autobiographical narrative scripts in the context of an anger induction paradigm. SETTING: Academic medical center. PARTICIPANTS: Thirty individuals, evenly divided among the 3 study groups. INTERVENTIONS: In separate conditions, participants were exposed to anger and neutral autobiographical scripts during the positron emission tomography study. Subjective self-report and psychophysiologic data were also collected. MAIN OUTCOME MEASURES: Voxelwise methods were used for analyses of regional cerebral blood flow changes for the anger vs neutral contrast within and between groups. RESULTS: Controls showed significantly (P<.001) greater regional cerebral blood flow increases in the left ventromedial prefrontal cortex during anger induction than patients with MDD + A, whereas these differences were not present in other between-group analyses. Also, in controls, an inverse relationship was demonstrated between regional cerebral blood flow changes during anger induction in the left ventromedial prefrontal cortex and left amygdala, whereas in patients with MDD + A there was a positive correlation between these brain regions during anger induction. There was no significant relationship between these brain regions during anger induction in patients with MDD - A. CONCLUSION: These results suggest a pathophysiology of MDD + A that is distinct from that of MDD - A and that may be responsible for the unique clinical presentation of patients with MDD + A. PMID- 15289279 TI - Prevalence and co-occurrence of substance use disorders and independent mood and anxiety disorders: results from the National Epidemiologic Survey on Alcohol and Related Conditions. AB - BACKGROUND: Uncertainties exist about the prevalence and comorbidity of substance use disorders and independent mood and anxiety disorders. OBJECTIVE: To present nationally representative data on the prevalence and comorbidity of DSM-IV alcohol and drug use disorders and independent mood and anxiety disorders (including only those that are not substance induced and that are not due to a general medical condition). DESIGN: Face-to-face survey. SETTING: The United States. PARTICIPANTS: Household and group quarters' residents. MAIN OUTCOME MEASURES: Prevalence and associations of substance use disorders and independent mood and anxiety disorders. RESULTS: The prevalences of 12-month DSM-IV independent mood and anxiety disorders in the US population were 9.21% (95% confidence interval [CI], 8.78%-9.64%) and 11.08% (95% CI, 10.43%-11.73%), respectively. The rate of substance use disorders was 9.35% (95% CI, 8.86% 9.84%). Only a few individuals with mood or anxiety disorders were classified as having only substance-induced disorders. Associations between most substance use disorders and independent mood and anxiety disorders were positive and significant (P<.05). CONCLUSIONS: Substance use disorders and mood and anxiety disorders that develop independently of intoxication and withdrawal are among the most prevalent psychiatric disorders in the United States. Associations between most substance use disorders and independent mood and anxiety disorders were overwhelmingly positive and significant, suggesting that treatment for a comorbid mood or anxiety disorder should not be withheld from individuals with substance use disorders. PMID- 15289280 TI - Relationships among plasma dehydroepiandrosterone sulfate and cortisol levels, symptoms of dissociation, and objective performance in humans exposed to acute stress. AB - CONTEXT: Recently, a growing body of research has provided evidence that dehydroepiandrosterone sulfate (DHEA-S) is involved in an organism's response to stress and that it may provide beneficial behavioral and neurotrophic effects. OBJECTIVE: To investigate plasma DHEA-S and cortisol levels, psychological symptoms of dissociation, and military performance. DESIGN: Prospective study. SETTING AND PARTICIPANTS: Twenty-five healthy subjects enrolled in military survival school. RESULTS: The DHEA-S-cortisol ratios during stress were significantly higher in subjects who reported fewer symptoms of dissociation and exhibited superior military performance. CONCLUSIONS: These data provide prospective, empirical evidence that the DHEA-S level is increased by acute stress in healthy humans and that the DHEA-S-cortisol ratio may index the degree to which an individual is buffered against the negative effects of stress. PMID- 15289281 TI - Effects of primary care depression treatment on minority patients' clinical status and employment. AB - BACKGROUND: The response of ethnic minorities to mental health care is largely unstudied. OBJECTIVE: To determine the effect of appropriate care for depression on ethnic minorities. DESIGN: Observational analysis of the effects of evidence based depression care over 6 months on clinical outcomes and employment status is examined for ethnic minorities and nonminorities. Selection into treatment is accounted for using instrumental variables techniques, with randomized assignment to the quality improvement intervention as the identifying instrument. SETTING: Six managed care organizations across the United States. Patients One thousand three hundred fifty-six depressed adults, including 601 white, 258 Latino, 56 African American, and 24 Asian or Native American patients. Intervention Quality improvement interventions aimed at increasing guideline-concordant depression care. RESULTS: At 6 months, minority patients who received appropriate care, compared with those who did not receive it, had lower rates of probable depressive disorder (20.5% vs 70.5%); the findings were similar for nonminority patients (24.3% vs 71.2%). Nonminority patients who received appropriate care were found to have higher rates of employment than were those who did not receive appropriate care (71.4% vs 52.4%). This was not true of minority patients (68.2% vs 56.5%). CONCLUSIONS: Evidence-based care for depression is equally effective in reducing depressive disorders for minority and nonminority patients. However, functional outcomes of care, such as continued employment, may be more limited for minority than nonminority patients. Because minority members are less likely to get appropriate care, efforts should be made to engage minority members in effective care for depression. PMID- 15289282 TI - Prenatal smoking and early childhood conduct problems: testing genetic and environmental explanations of the association. AB - BACKGROUND: Extensive evidence now supports a statistical association between prenatal smoking and increased risk for antisocial outcomes in offspring. Though this statistical link may signal a causal association, commentators have urged caution in interpreting findings because of the likelihood of confounding. METHODS: We used data from the Environmental Risk Longitudinal Twin Study, a representative British sample of 1116 twin pairs studied at ages 5 and 7 years, to assess associations between prenatal smoking and early childhood conduct problems net of the effects of both heritable and environmental risks for child antisocial outcomes. RESULTS: Prenatal smoking showed a strong, dose-response relationship with child conduct problems at ages 5 and 7 years. Around half of this association was attributable to correlated genetic effects. Mothers who smoked during pregnancy differed from other mothers in a number of ways. They were more likely to be antisocial, had children with more antisocial men, were bringing up their children in more disadvantaged circumstances, and were more likely to have had depression. Controlling for antisocial behavior in both parents, depression in mothers, family disadvantage, and genetic influences, estimates for the effects of prenatal smoking were reduced by between 75% and the entire initial effects. CONCLUSIONS: Observed associations between prenatal smoking and childhood conduct problems are likely to be heavily confounded with other known risks for children's behavioral development. As a result, tests of any causal influence of prenatal smoking must await findings from experimental studies. PMID- 15289283 TI - Novel diacylglycerol kinase inhibitor selectively suppressed an U46619-induced enhancement of mouse portal vein contraction under high glucose conditions. AB - 1. Diacylglycerol kinase (DG kinase) is a key enzyme in vascular contraction; however, alterations of the regulatory mechanisms in vascular dysfunction are poorly understood. In this study, the effect of a novel DG kinase inhibitor, stemphone, on vascular contraction was investigated. 2. The conventional DG kinase inhibitor, 6-[2-(4-[(4-fluorophenyl)phenyl-methylene]-1-piperidinyl)ethyl] 7-methyl-5H-thiazolo [3,2-alpha] pyrimidine-5-one (R59022) (0.1-30 microm), inhibited thromboxane A(2) analogue 9,11-dideoxy-11alpha,9alpha epoxymethanoprostaglandin F(2alpha) (U46619)-induced sustained contractions in mouse aorta and porcine coronary artery in a dose-dependent manner. Treatment with stemphone did not affect contractions in these tissues. However, stemphone significantly inhibited (>0.3 microm) U46619-induced spontaneous phasic contraction in mouse portal vein. This inhibitory effect was not detected following R59022 treatment in portal vein. Therefore, stemphone demonstrated selectivity in terms of portal vein contraction. 3. Under high glucose (22.2 mm) conditions, U46619-induced contraction was enhanced in these three types of vascular tissue. Inhibitory effects of R59022 were attenuated under these conditions; however, effects of stemphone were observed. These results indicated that stemphone could inhibit portal vein contraction under high glucose conditions, for example, diabetes. These data suggested the possibility that DG kinase may be a target of hyperportal pressure. 4. Total mass of DG was enhanced under high glucose conditions. DG was derived from incorporated glucose via de novo synthesis in the absence of phospholipase C pathway mediation. This enhanced DG under high glucose conditions activated a calcium-independent protein kinase C (PKC). This PKC was associated with calcium-independent DG kinase activation. Treatment with stemphone also inhibited calcium-independent DG kinase. These signal transduction pathways were distinguishable from a DG-PKC pathway under normal glucose conditions. 5. The present investigation suggested that stemphone selectively inhibited overcontraction of portal vein induced by high glucose levels. This phenomenon was attributable to inhibition of calcium-independent DG kinase activation that occurred under high glucose conditions mediated by both DG synthesized from glucose and calcium-independent PKC activation. PMID- 15289284 TI - GEA 3162 decomposes to co-generate nitric oxide and superoxide and induces apoptosis in human neutrophils via a peroxynitrite-dependent mechanism. AB - 1. GEA 3162 (1,2,3,4,-oxatriazolium, 5-amino-3-(3,4-dichlorophenyl)-chloride), has powerful effects on neutrophil function and apoptosis, but the underlying mechanisms are unclear, particularly with respect to the possible roles of nitric oxide (NO) and/or peroxynitrite (ONOO(-)). 2. Our hypothesis was that GEA 3162 is a generator of ONOO(-) and that its biological effects on neutrophil apoptosis differ from those of a conventional NO donor. The effects of GEA 3162 were compared to those of the established ONOO(-) donor, 3-morpholinosydnonimine (SIN 1), and the NO donor, diethylamine diazeniumdiolate (DEA/NO) in neutrophils from healthy volunteers. Electrochemical detection and electron paramagnetic resonance were used to define the NO-related species generated from these agents. 3. GEA 3162 and SIN-1 influence neutrophil apoptosis differently from DEA/NO. All three compounds induced morphological neutrophil apoptosis. However, both GEA 3162 and SIN-1 paradoxically inhibited internucleosomal DNA fragmentation, whereas DEA/NO induced fragmentation compared to control. 4. In contrast to DEA/NO, generation of free NO was not detectable in solutions of GEA 3162 or SIN-1 (100 microm). However, Cu/Zn superoxide dismutase (SOD; 50-750 U ml(-1)) unmasked NO generated from these compounds in a concentration-dependent manner. GEA 3162 and SIN-1 oxidised the O(2)(-)- and ONOO(-)-sensitive dye, dihydrorhodamine 123 (DHR 123; 1 microm), suggesting that ONOO(-) released from these compounds is responsible for oxidation of DHR 123. 5. We conclude that GEA 3162 is an ONOO(-) donor with pro apoptotic properties that more closely resemble SIN-1 than the NO donor, DEA/NO. Moreover, unlike NO, ONOO(-) induces apoptosis in neutrophils via a mechanism that does not require DNA fragmentation. PMID- 15289285 TI - Aspirin induces nitric oxide release from vascular endothelium: a novel mechanism of action. AB - 1. The study was designed to test the hypothesis that aspirin may stimulate nitric oxide (NO) release from vascular endothelium, a pivotal factor for maintenance of vascular homeostasis. 2. Clinical evidence suggests that low-dose aspirin may improve vascular endothelial function. Since other cyclooxygenase (COX) inhibitors showed no beneficial vascular effects, aspirin may exhibit a vasculoprotective, COX-independent mechanism. 3. Luminal NO release was monitored in real time on dissected porcine coronary arteries (PCA) by an amperometric, NO selective sensor. Additionally, endothelial NO synthase (eNOS) activity was measured in EA.hy 926 cell homogenates by an l-[(3)H]citrulline/l-[(3)H]arginine conversion assay. Superoxide scavenging capacity was assessed by lucigenin enhanced luminescence. 4. Aspirin induced an immediate concentration-dependent NO release from PCA with an EC(50) of 50 nm and potentiated the NO stimulation by the receptor-dependent agonist substance P. These effects were independent of an increase in intracellular calcium and could be mimicked by stimulation with acetylating aspirin derivatives. The aspirin metabolite salicylic acid or the reversible cyclooxygenase inhibitor indomethacin failed to modulate NO release. Incubation of soluble eNOS for 15 min with 100 microm aspirin or acetylating aspirin analogues increased the l-[(3)H]citrulline yield by 40-80%, while salicylic acid had no effect. Aspirin and salicylic acid showed a similar, but only modest, magnitude and velocity of superoxide scavenging. 5. Our findings demonstrate that therapeutically relevant concentrations of aspirin elicit NO release from vascular endothelium. This effect appears to be due to a direct acetylation of the eNOS protein, but is independent of COX inhibition or inhibition of superoxide-mediated NO degradation. PMID- 15289286 TI - The NOP (ORL1) receptor antagonist Compound B stimulates mesolimbic dopamine release and is rewarding in mice by a non-NOP-receptor-mediated mechanism. AB - 1. Compound B (1-[(3R, 4R)-1-cyclooctylmethyl-3-hydroxymethyl-4-piperidyl]-3 ethyl-1,3-dihydro-2H-benzimidazol-2-one, CompB) is a nociceptin/orphanin FQ (N/OFQ) antagonist showing high selectivity for the NOP (ORL1) receptor over classical opioid receptors. We studied the effect of subcutaneous CompB administration on the release of mesolimbic dopamine (DA) and the expression of hedonia in mice. 2. CompB (0.3-30 mg kg(-1)) dose dependently stimulated mesolimbic DA release as measured by in vivo freely moving microdialysis, without any change in locomotor activity. However, intracerebroventricular administered N/OFQ (endogenous agonist of the NOP receptor, 6 nmol) did not influence CompB- (10 mg kg(-1)) induced DA release, despite clearly suppressing release when administered alone. 3. Studies using NOP receptor knockout mice and no-net-flux microdialysis revealed mildly, but not statistically significantly higher endogenous DA levels in mice lacking the NOP receptor compared to wild-type mice. Administration of CompB (10 mg kg(-1)) induced identical increases in mesolimbic DA release in wild-type and NOP receptor knockout mice. 4. CompB was rewarding in approximately the same dose range in which CompB induced major increases in mesolimbic DA release when assayed using a conditioned place preference paradigm. The rewarding effect of CompB (30 mg kg(-1)) was maintained in NOP receptor knockout mice. 5. These results show that CompB stimulates mesolimbic DA release and is rewarding by an action independent of the NOP receptor, the precise site of which is unclear. Consequently, caution should be exercised when interpreting the results of studies using this drug, particularly when administered by a peripheral route. PMID- 15289287 TI - Influence of agonist intrinsic activity on the desensitisation of beta2 adrenoceptor-mediated responses in mast cells. AB - 1. The aim of the present study was to determine whether the intrinsic activity of an agonist influences the extent of desensitisation of beta(2)-adrenoceptor mediated responses in human lung mast cells. 2. The effects of a wide range of beta-adrenoceptor agonists (10(-10)-10(-5) m) on the IgE-mediated release of histamine from mast cells were determined. The intrinsic activity of agonists was established by comparing the maximal inhibitory response (E(max)) of an agonist relative to the maximal response obtained with the full agonist, isoprenaline. The intrinsic activity order for the inhibition of histamine release was isoprenaline (1.0)>formoterol (0.94)>fenoterol (0.89)>terbutaline (0.84)>salbutamol (0.69)>clenbuterol (0.65)>salmeterol (0.30)>dobutamine (0.20). 3. There was a significant (P<0.05) positive correlation (r=0.81) between the extent to which beta-adrenoceptor agonists inhibited histamine release and the degree to which the agonists caused elevations in cAMP in mast cells. 4. Further studies investigated the effects of long-term (24 h) incubation of mast cells with beta-adrenoceptor agonists on the subsequent ability of isoprenaline to inhibit histamine release. At concentrations of agonists selected to occupy a large percentage (88%) of beta(2)-adrenoceptors, there was a significant (P<0.05) correlation (r=0.73) between the relative intrinsic activity of agonists as inhibitors of histamine release and the extent of functional desensitisation induced by the agonists. At lower receptor occupancies, however, there was no correlation between the relative intrinsic activity of agonists and the extent of agonist-induced desensitisation. 5. These data indicate that, under experimental conditions where high receptor occupancies prevail, agonist intrinsic activity influences the extent of desensitisation of beta(2)-adrenoceptor-mediated responses in mast cells. PMID- 15289288 TI - Serum albumin induces iNOS expression and NO production in RAW 267.4 macrophages. AB - 1. We investigated the effects of serum albumin on inducible nitric oxide synthase (iNOS) expression in RAW 267.4 macrophages. Crude fraction-V type albumin as well as bovine serum albumin filtrated for endotoxin induced concentration-dependent iNOS expression in macrophages. Accordingly, NO production (estimated by supernatant nitrite) was markedly (up to 10-fold) increased in the presence of albumin. 2. Albumin-induced expression of iNOS protein was inhibited by cycloheximide and NO production was abolished after incubation of the cells with an iNOS inhibitor, N(G)-monomethyl-l-arginine (LNMMA). 3. An inhibitor of the NF-kappaB pathway, pyrrolidine dithiocarbamate (PDTC), as well as inhibitors of JAK2/STAT and ERK, AG490 and U0126, respectively, significantly reduced albumin-induced iNOS expression and NO production, while an inhibitor of the p38 pathway, SB203580, did not significantly affect NO production induced by albumin. 4. Both types of serum albumin were contaminated with traces of endotoxin. The endotoxin levels were found not to be sufficient for the observed induction of nitrite production in RAW 267.4 cells. In addition, the albumin-stimulated induction of iNOS was not reduced by preincubation of albumin-containing media with polymyxin B, a LPS inhibitor. 5. Polymerised albumin fractions were detected in the commercially available albumin tested in this study. A monomeric albumin-rich fraction, separated by ultrafiltration, showed a potent inducing effect on iNOS expression and NO production, while a polymer-rich fraction showed a smaller effect. 6. Advanced glycation endproducts (AGE) of albumin were not formed by interaction with glucose in incubation medium, as AGE was not increased even after long-time (4 weeks) incubation in albumin-containing media [3.2-4.4 microg ml(-1) (basal) vs 4.8-5.6 microg ml(-1) (in glucose-containing media)]. However, the duration of albumin exposure to glucose influenced the basal stimulatory properties of albumin. 7. Our results suggest that serum albumin fractions, as gained by cold alcoholic extraction, may include determinants that stimulate or further enhance stimulation of RAW 267.4 cells and are different from endotoxin, polymeric albumin and AGE. PMID- 15289289 TI - Nephroprotection in Zucker diabetic fatty rats by vasopeptidase inhibition is partly bradykinin B2 receptor dependent. AB - 1. Vasopeptidase inhibition (i.e., the simultaneous inhibition of both angiotensin-converting enzyme (ACE) and neutral endopeptidase) can ameliorate diabetic nephropathy. We investigated whether this nephroprotection is mediated by the bradykinin B2 receptor. 2. In all, 43 obese Zucker diabetic fatty (ZDF/Gmi fa/fa) rats aged 21 weeks were separated into four groups and treated for 26 weeks with either placebo, the bradykinin B2 receptor antagonist icatibant (500 microg kg(-1) day(-1) s.c. infusion), the vasopeptidase inhibitor AVE7688 (45 mg kg(-1) day(-1) in chow), or AVE7688 plus icatibant. Nephropathy was assessed as albuminuria at age 31 and 39 weeks, and by histopathologic scoring at the end of the treatment period. 3. All animals had established diabetes mellitus (blood glucose >20 mmol l(-1)) and marked albuminuria at baseline. Blood glucose was not influenced by any treatment. Icatibant alone did not influence albuminuria (8.6+/ 1.6 vs placebo 9.5+/-1.3 mg kg(-1) h(-1)). AVE7688 reduced albuminuria at week 31 markedly to 1.1+/-0.1 mg kg(-1) h(-1) and reduced glomerular and tubulo interstitial kidney damage at week 47. In the AVE7688 plus icatibant group, proteinuria was significantly higher than in the AVE7688 only group (2.0+/-0.6 mg kg(-1) h(-1)), but still reduced compared to placebo. In addition, icatibant partly antagonized the tubulo-interstitial protection mediated by AVE7688. 4. We conclude that vasopeptidase inhibition provides nephroprotection in rats with type II diabetic nephropathy, which is partly mediated by bradykinin B2 receptor activation. PMID- 15289290 TI - Sodium-calcium exchanger contributes to membrane hyperpolarization of intact endothelial cells from rat aorta during acetylcholine stimulation. AB - 1. The role of sodium-calcium exchanger in acetylcholine (Ach)-induced hyperpolarization of intact endothelial cells was studied in excised rat aorta. The membrane potential was recorded using perforated patch-clamp technique. 2. The mean resting potential of endothelial cells was -44.1+/-1.4 mV. A selective inhibitor of sodium-calcium exchanger benzamil (100 microm) had no significant effect on resting membrane potential, but reversibly decreased the amplitude of sustained Ach-induced endothelial hyperpolarization from 20.9+/-1.4 to 5.7+/-1.1 mV when applied during the plateau phase. 3. The blocker of reversed mode of the exchanger KB-R7943 (2-[2-[4-(4-nitrobenzyloxy)phenyl]ethyl]isothiourea methanesulfonate, 20 microm) reversibly decreased the amplitude of sustained Ach induced hyperpolarization from 20.5+/-2.9 to 7.5+/-1.8 mV. 4. Introduction of tetraethylammonium (10 mm) in the continuous presence of Ach decreased the sustained phase of hyperpolarization from 17.9+/-1.5 by 12.9+/-0.9 mV. Subsequent addition of 20 microm KB-R7943 further depolarized endothelial cells by 4.8+/-1.1 mV. 5. Substituting external sodium with N-methyl d-glucamine during the plateau phase of Ach-evoked hyperpolarization reversibly decreased the hyperpolarization from -61.8+/-2.7 to -54.2+/-1.9 mV. In the majority of preparations, the initial response to removal of external sodium was a transient further rise in the membrane potential of several mV. Sodium ionophore monensin hyperpolarized endothelium by 10.3+/-0.7 mV. 6. The inhibitory effect of benzamil on Ach-induced endothelial sustained hyperpolarization was observed in endothelium mechanically isolated from smooth muscle. 7. These results suggest that the sodium-calcium exchanger of intact endothelial cells is able to operate in reverse following stimulation by Ach, contributing to sustained hyperpolarization. Myoendothelial electrical communications do not mediate the effect of blockers of sodium-calcium exchanger. PMID- 15289292 TI - Mutual amplification of apoptosis by statin-induced mitochondrial stress and doxorubicin toxicity in human rhabdomyosarcoma cells. AB - Besides their cholesterol-lowering effect, 3-hydroxy-3-methyl-glutaryl-CoA reductase inhibitors (statins) show antiproliferative behaviour, which has been suggested as a promising anticancer strategy. However, the signalling cascades leading to statin-induced death of cancer cells are poorly characterized. Here we show that statins activate the mitochondrial pathway of apoptosis in rhabdomyosarcoma RD cells via translocation of Bax from the cytosol to mitochondria. The prototypical representative of statins, simvastatin, induced consecutive activation of caspase 9 and 3 in a concentration-dependent manner. The permeability transition pore inhibitor bongkrekic acid was capable of completely preventing simvastatin-induced caspase 9 and 3 activity, corroborating the mitochondrial pathway of apoptosis as the sole mechanism of statin action. Alternative pathways via death receptors, that is, caspase 8 or calpain activation, were not triggered by simvastatin. Simvastatin-treated RD cells could be completely rescued from apoptosis by the co-application of mevalonic acid, indicating that deprivation of cholesterol precursors is essential for statin induced apoptosis. However, pretreatment with subthreshold concentrations of simvastatin was sufficient to augment doxorubicin toxicity via the mitochondrial apoptotic machinery. Moreover, the presence of doxorubicin increased the potency of simvastatin to trigger caspase activation. Taken together, these data highlight the therapeutic anticancer potential of statins and their additivity and mutual sensitization, in combination with doxorubicin in human rhabdomyosarcoma cells. PMID- 15289291 TI - The evolving role of lipid rafts and caveolae in G protein-coupled receptor signaling: implications for molecular pharmacology. AB - The many components of G-protein-coupled receptor (GPCR) signal transduction provide cells with numerous combinations with which to customize their responses to hormones, neurotransmitters, and pharmacologic agonists. GPCRs function as guanine nucleotide exchange factors for heterotrimeric (alpha, beta, gamma) G proteins, thereby promoting exchange of GTP for GDP and, in turn, the activation of 'downstream' signaling components. Recent data indicate that individual cells express mRNA for perhaps over 100 different GPCRs (out of a total of nearly a thousand GPCR genes), several different combinations of G-protein subunits, multiple regulators of G-protein signaling proteins (which function as GTPase activating proteins), and various isoforms of downstream effector molecules. The differential expression of such protein combinations allows for modulation of signals that are customized for a specific cell type, perhaps at different states of maturation or differentiation. In addition, in the linear arrangement of molecular interactions involved in a given GPCR-G-protein-effector pathway, one needs to consider the localization of receptors and post-receptor components in subcellular compartments, microdomains, and molecular complexes, and to understand the movement of proteins between these compartments. Co-localization of signaling components, many of which are expressed at low overall concentrations, allows cells to tailor their responses by arranging, or spatially organizing in unique and kinetically favorable ways, the molecules involved in GPCR signal transduction. This review focuses on the role of lipid rafts and a subpopulation of such rafts, caveolae, as a key spatial compartment enriched in components of GPCR signal transduction. Recent data suggest cell-specific patterns for expression of those components in lipid rafts and caveolae. Such domains likely define functionally important, cell-specific regions of signaling by GPCRs and drugs active at those GPCRs. PMID- 15289293 TI - Actions of two naturally occurring saturated N-acyldopamines on transient receptor potential vanilloid 1 (TRPV1) channels. AB - Four long-chain, linear fatty acid dopamides (N-acyldopamines) have been identified in nervous bovine and rat tissues. Two unsaturated members of this family of lipids, N-arachidonoyl-dopamine (NADA) and N-oleoyl-dopamine, were shown to potently activate the transient receptor potential channel type V1 (TRPV1), also known as the vanilloid receptor type 1 for capsaicin. However, the other two congeners, N-palmitoyl- and N-stearoyl-dopamine (PALDA and STEARDA), are inactive on TRPV1. We have investigated here the possibility that the two compounds act by enhancing the effect of NADA on TRPV1 ('entourage' effect). When pre-incubated for 5 min with cells, both compounds dose-dependently enhanced NADA's TRPV1-mediated effect on intracellular Ca(2+) in human embryonic kidney cells overexpressing the human TRPV1. In the presence of either PALDA or STEARDA (0.1-10 microm), the EC(50) of NADA was lowered from approximately 90 to approximately 30 nm. The effect on intracellular Ca(2+) by another endovanilloid, N-arachidonoyl-ethanolamine (anandamide, 50 nm), was also enhanced dose dependently by both PALDA and STEARDA. PALDA and STEARDA also acted in synergy with low pH (6.0-6.7) to enhance intracellular Ca(2+) via TRPV1. When co-injected with NADA (0.5 micrograms) in rat hind paws, STEARDA (5 micrograms) potentiated NADA's TRPV1-mediated nociceptive effect by significantly shortening the withdrawal latencies from a radiant heat source. STEARDA (1 and 10 micrograms) also enhanced the nocifensive behavior induced by carrageenan in a typical test of inflammatory pain. These data indicate that, despite their inactivity per se on TRPV1, PALDA and STEARDA may play a role as 'entourage' compounds on chemicophysical agents that interact with these receptors, with possible implications in inflammatory and neuropathic pain. PMID- 15289294 TI - Nucleoside transporter subtype expression and function in rat skeletal muscle microvascular endothelial cells. AB - 1. Microvascular endothelial cells (MVECs) form a barrier between circulating metabolites, such as adenosine, and the surrounding tissue. We hypothesize that MVECs have a high capacity for the accumulation of nucleosides, such that inhibition of the endothelial nucleoside transporters (NT) would profoundly affect the actions of adenosine in the microvasculature. 2. We assessed the binding of [(3)H]nitrobenzylmercaptopurine riboside (NBMPR), a specific probe for the inhibitor-sensitive subtype of equilibrative NT (es), and the uptake of [(3)H]formycin B (FB), by MVECs isolated from rat skeletal muscle. The cellular expression of equilibrative (ENT1, ENT2, ENT3) and concentrative (CNT1, CNT2, CNT3) NT subtypes was also determined using both qualitative and quantitative polymerase chain reaction techniques. 3. In the absence of Na(+), MVECs accumulated [(3)H]FB with a V(max) of 21+/-1 pmol microl(-1) s(-1). This uptake was mediated equally by es (K(m) 260+/-70 microm) and ei (equilibrative inhibitor insensitive; K(m) 130+/-20 microm) NTs. 4. A minor component of Na(+)-dependent cif (concentrative inhibitor-insensitive FB transporter)/CNT2-mediated [(3)H]FB uptake (V(i) 0.008+/-0.005 pmol microl(-1) s(-1) at 10 microm) was also observed at room temperature upon inhibition of ENTs with dipyridamole (2,6 bis(diethanolamino)-4,8-dipiperidinopyrimido-[5,4-d]pyrimidine)/NBMPR. 5. MVECs had 122,000 high-affinity (K(d) 0.10 nm) [(3)H]NBMPR binding sites (representing es transporters) per cell. A lower-affinity [(3)H]NBMPR binding component (K(d) 4.8 nm) was also observed that may be related to intracellular es-like proteins. 6. Rat skeletal muscle MVECs express es/ENT1, ei/ENT2, and cif/CNT2 transporters with characteristics typical of rat tissues. This primary cell culture model will enable future studies on factors influencing NT subtype expression, and the consequent effect on adenosine bioactivity, in the microvasculature. PMID- 15289295 TI - Expression of matrix metalloproteinase-9 in human platelets: regulation of platelet activation in in vitro and in vivo studies. AB - 1. The aim of this study was to identify the presence of matrix metalloproteinase 9 (MMP-9) in human platelets and systematically examine its inhibitory mechanisms of platelet activation. 2. In this study, we report on an efficient method for the quantitative analysis of pro-MMP-9 in human platelets using capillary zone electrophoresis (CZE). To elucidate subcellular localization of MMP-9 in human platelets, we investigated intraplatelet MMP-9 by immunogold labeling and visualized it using electron microscopy. In an in vivo thrombotic study, platelet thrombus formation was induced by irradiation of mesenteric venules with filtered light in mice pretreated with fluorescein sodium. 3. MMP-9-gold labeling was observed on the plasma membrane, alpha-granules, open canalicular system, and within the cytoplasma both in resting and activated platelets. Furthermore, activated MMP-9 concentration-dependently (15-90 ng ml(-1)) inhibited platelet aggregation stimulated by agonists. Activated MMP-9 (21 and 90 ng ml(-1)) inhibited phosphoinositide breakdown, intracellular Ca(2+) mobilization, and thromboxane A(2) formation in human platelets stimulated by collagen (1 microg ml(-1)). In addition, activated MMP-9 (21 and 90 ng ml(-1)) significantly increased the formation of nitric oxide/cyclic GMP. 4. Rapid phosphorylation of a platelet protein of Mr 47,000 (P47), a marker of protein kinase C activation, was triggered by phorbol-12, 13-dibutyrate (PDBu) (60 nm). This phosphorylation was markedly inhibited by activated MMP-9 (21 and 90 ng ml(-1)). Activated MMP-9 (1 microg g(-1)) significantly prolonged the latency period of inducing platelet plug formation in mesenteric venules. 5. These results indicate that the antiplatelet activity of activated MMP-9 may be involved in the following pathways. (1) Activated MMP-9 may inhibit the activation of phospholipase C, followed by inhibition of phosphoinositide breakdown, protein kinase C activation, and thromboxane A(2) formation, thereby leading to inhibition of intracellular Ca(2+) mobilization. (2) Activated MMP-9 also activated the formation of nitric oxide/cyclic GMP, resulting in inhibition of platelet aggregation. These results strongly indicate that MMP-9 is a potent inhibitor of aggregation. It may play an important role as a negative feedback regulator during platelet activation. PMID- 15289296 TI - P2Y2-receptor-mediated activation of a contralateral, lanthanide-sensitive calcium entry pathway in the human airway epithelium. AB - 1. Receptor-mediated calcium entry (RMCE) was examined in well-differentiated cultures of normal human bronchial epithelial cells (HBECs). Changes in intracellular free Ca(2+) ([Ca(2+)](i)) were quantified using fluorescence ratio imaging of Fura-2-loaded cells during perfusion with Ca(2+) mobilizing agonists. 2. Initial studies revealed an agonist potency of ATP=uridine triphosphate (UTP) >ADP=uridine diphosphate, consistent with purinergic activation of an apical P2Y(2)-receptor mediating the increase in [Ca(2+)](i) in HBECs. 3. Apical UTP (30 microm) induced a sustained period of elevated [Ca(2+)](i) between 300 and 600 s following agonist stimulation that extracellular Ca(2+) free studies indicated was dominated by Ca(2+) influx. 4. RMCE was inhibited by 100 nm La(3+) (83+/-3%) or Gd(3+) (95+/-7%) (P<0.005, n=4-11) and was partially attenuated by Ni(2+) (1 mm) (58.7+/-5.0%, P<0.005, n=9). 5. RMCE was also partially sensitive (< 25% inhibition, P<0.01) to the cation channel blockers SKF96365 (30 microm) and econazole (30 microm), but was insensitive to both verapamil (1 microm) and ruthenium red (10 microm). 6. Using either a sided Ca(2+) readdition protocol or unilateral La(3+), established that the RMCE pathway was located exclusively on the basolateral membrane. 7. The pharmacological sensitivity of the P2Y(2) receptor activated Ca(2+) entry pathway in the human airway epithelium is inconsistent with the established profile of TRP channel families and is therefore likely to be of an as-yet uncharacterized molecular identity. PMID- 15289297 TI - Preclinical characterisation of 111In-DTPA-trastuzumab. AB - Trastuzumab (Herceptin) is a recombinant humanised IgG1 monoclonal antibody against the human epidermal growth factor receptor 2 (HER2), used for metastatic breast cancer treatment. Radiolabelled trastuzumab may have several future applications for diagnostic use. The aim of the present study was to develop clinical grade (111)Indium ((111)In) radiolabelled trastuzumab, to evaluate the stability and immunoreactivity of the tracer and to perform a biodistribution study in human tumour-bearing mice. Trastuzumab was radiolabelled with (111)In using DTPA as a chelator. (111)In-DTPA-trastuzumab (labelling yield 92.3+/-2.3%, radiochemical purity 97.0+/-1.5%) is stable in PBS when stored at 4 degrees C for more than 14 days. The immunoreactive fraction determined by cell-binding assays, using the HER2-overexpressing human ovarian SK-OV-3 tumour cell line, was 0.87+/ 0.06. Biodistribution and tumour targeting were studied in HER2 receptor-positive and -negative tumour-bearing athymic mice. The HER2-positive tumour showed (9.77+/-1.14% injected dose per gram (ID g(-1))) substantial uptake of the labelled antibody already after 5 h. The difference in uptake between HER2 positive versus -negative tumours was even more pronounced 3 days after injection (16.30+/-0.64% ID g(-1)), and was visualised by radioimmunoscintigraphy. Liver, spleen and kidney showed marked tracer uptake. In summary, trastuzumab can be efficiently radiolabelled with (111)In with high labelling yields and high stability. (111)In-DTPA-trastuzumab selectively binds to the human HER2 receptor both in vitro and in vivo in animals. Therefore, (111)In-DTPA-trastuzumab appears suitable for clinical use. PMID- 15289298 TI - Sigma receptors and cancer: possible involvement of ion channels. AB - The sigma (sigma) receptor and its agonists have been implicated in a myriad of cellular functions, biological processes and diseases. Whereas the precise molecular mechanism(s) of sigma receptors and their involvement in cancer cell biology have not been elucidated, recent work has started to shed some light on these issues. A molecular model has been proposed for the cloned sigma1 receptor; the precise molecular nature of the sigma2 receptor remains unknown. sigma receptors have been found to be frequently up-regulated in human cancer cells and tissues. sigma2 receptor drugs particularly have been shown to have antiproliferative effects. An interesting possibility is that sigma and/or sigma1 drugs could produce anticancerous effects by modulating ion channels. As well as proliferation, a variety of other metastatic cellular behaviors such as adhesion, motility, and secretion may also be affected. Other mechanisms of sigma receptor action may involve interaction with ankyrin and modulation of intracellular Ca(2+) and sphingolipid levels. Although more research is needed to further define the molecular physiology of sigma receptors, their involvement in the cellular pathophysiology of cancer raises the possibility that sigma drugs could be useful as novel therapeutic agents. PMID- 15289299 TI - Bortezomib as a potential treatment for prostate cancer. AB - Androgen ablation and chemotherapy provide effective palliation for most patients with advanced prostate cancer, but eventually progressing androgen-independent prostate cancer threatens the lives of patients usually within a few years, mandating improvement in therapy. Proteasome inhibition has been proposed as a therapy target for the treatment of solid and hematological malignancies. The proteasome is a ubiquitous enzyme complex that is a hub for the regulation of many intracellular regulatory pathways; because of its essential function, this enzyme has become a new target for cancer treatment. Studies with bortezomib (VELCADE, formerly known as PS-341) and other proteasome inhibitors indicate that cancer cells are especially dependent on the proteasome for survival, and several mechanisms used by prostate cancer cells require proteasome function. Bortezomib has been studied extensively in vitro and in vivo, and anticancer activity has been seen in cell and animal models for several solid tumor types, including prostate cancer. A Phase I trial to determine the maximum tolerated dose of once weekly bortezomib has been completed. This trial included a large fraction of patients with androgen-independent prostate cancer. The maximum tolerated dose was reached at 1.6 mg/m(2). A correlation was seen among bortezomib dose, proteasome inhibition, and positive modulation of serum prostate-specific antigen. There was also evidence of down-regulation of serum interleukin 6, a downstream nuclear factor kappaB effector. This Phase I trial and preclinical studies support additional testing of bortezomib in combination with radiation or chemotherapy for androgen-independent prostate cancer. PMID- 15289300 TI - In vivo flow cytometry: a new method for enumerating circulating cancer cells. AB - The fate of circulating tumor cells is an important determinant of their ability to form distant metastasis. Here, we demonstrate the use of in vivo flow cytometry as a powerful new method for detecting quantitatively circulating cancer cells. We specifically examine the circulation kinetics of two prostate cancer cell lines with different metastatic potential in mice and rats. We find that the cell line and the host environment affect the circulation kinetics of prostate cancer cells, with the intrinsic cell line properties determining the initial rate of cell depletion from the circulation and the host affecting cell circulation at later time points. PMID- 15289301 TI - Mutations of PIK3CA in anaplastic oligodendrogliomas, high-grade astrocytomas, and medulloblastomas. AB - The phosphatidylinositol 3'-kinase pathway is activated in multiple advanced cancers, including glioblastomas, through inactivation of the PTEN tumor suppressor gene. Recently, mutations in PIK3CA, a member of the family of phosphatidylinositol 3'-kinase catalytic subunits, were identified in a significant fraction (25-30%) of colorectal cancers, gastric cancers, and glioblastomas and in a smaller fraction of breast and lung cancers. These mutations were found to cluster into two major "hot spots" located in the helical and catalytic domains. To determine whether PIK3CA is genetically altered in brain tumors, we performed a large-scale mutational analysis of the helical and catalytic domains. A total of 13 mutations of PIK3CA within these specific domains were identified in anaplastic oligodendrogliomas, anaplastic astrocytomas, glioblastoma multiforme, and medulloblastomas, whereas no mutations were identified in ependymomas or low-grade astrocytomas. These observations implicate PIK3CA as an oncogene in a wider spectrum of adult and pediatric brain tumors and suggest that PIK3CA may be a useful diagnostic marker or a therapeutic target in these cancers. PMID- 15289302 TI - Increased progesterone receptor expression in benign epithelium of BRCA1-related breast cancers. AB - The study of pathologically normal breast epithelium of BRCA mutation carriers may yield insights into the early natural history of breast tumorigenesis. Hormone receptor expression was assessed in 24 cases of invasive breast cancer associated with a mutation in BRCA1 (n = 15) or BRCA2 (n = 9) and in 39 sporadic cases matched for patient age and tumor hormone receptor status. Expression of progesterone receptor was significantly (P = 0.0003) more common in normal breast epithelium adjacent to invasive breast carcinoma in BRCA1-linked cases compared with sporadic cases. The wild-type BRCA allele was retained in normal epithelium of all cases tested. We conclude that deregulation of progesterone receptor expression, as a result of BRCA1 haploinsufficiency, may represent an early event in BRCA1-linked breast tumorigenesis. PMID- 15289303 TI - Inducible activation of oncogenic K-ras results in tumor formation in the oral cavity. AB - Mouse models for cancer represent powerful tools to analyze the causal role of genetic alterations in cancer development. We have developed a novel mouse model that allows the focal activation of mutations in stratified epithelia. Using this system, we demonstrate that activation of an oncogenic K-rasG12D allele in the oral cavity of the mouse induces oral tumor formation. The lesions that develop in these mice are classified as benign squamous papillomas. Interestingly, these tumors exhibit changes in the expression pattern of keratins similar to those observed in human premalignant oral tumors, which are reflective of early stages of tumorigenesis. These results demonstrate a causal role for oncogenic K-ras in oral tumor development. The inducible nature of this model also makes it an ideal system to study cooperative interactions between mutations in oncogenes and/or tumor suppressor genes that are similar to those observed in human tumors. To our knowledge, this is the first reported inducible mouse model for oral cancer. PMID- 15289304 TI - Uric acid promotes tumor immune rejection. AB - Uric acid released from dying cells has been shown recently to act as a danger signal for the immune system, stimulating dendritic cell maturation and enhancing T-cell responses to foreign antigens. Stimulation of dendritic cell maturation by uric acid has been proposed as a mechanism by which the immune system could generate responses against tumors. We show here that uric acid levels are elevated in tumors undergoing immune rejection and that the inhibition of uric acid production, by systemic administration of allopurinol, or the removal of uric acid, by administration of uricase, delayed tumor immune rejection, whereas subcutaneous administration of crystalline uric acid enhanced the rejection process. PMID- 15289305 TI - Peloruside A does not bind to the taxoid site on beta-tubulin and retains its activity in multidrug-resistant cell lines. AB - Peloruside A (peloruside), a microtubule-stabilizing agent from a marine sponge, is less susceptible than paclitaxel to multidrug resistance arising from overexpression of the P-glycoprotein efflux pump and is not affected by mutations that affect the taxoid binding site of beta-tubulin. In vitro studies with purified tubulin indicate that peloruside directly induces tubulin polymerization in the absence of microtubule-associated proteins. Competition for binding between peloruside, paclitaxel, and laulimalide revealed that peloruside binds to a different site on tubulin to paclitaxel. Moreover, laulimalide was able to displace peloruside, indicating that peloruside and laulimalide may compete for the same or overlapping binding sites. It was concluded that peloruside and laulimalide have binding properties that are distinct from other microtubule stabilizing compounds currently under investigation. PMID- 15289306 TI - Overexpression of the anti-adhesin podocalyxin is an independent predictor of breast cancer progression. AB - Podocalyxin is a CD34-related cell surface molecule with anti-adhesive qualities. We probed a tissue microarray (n = 272) linked to long-term outcome data and found that podocalyxin was highly overexpressed in a distinct subset of invasive breast carcinomas (n = 15; 6%). Univariate disease-specific (P < 0.01) and multivariate regression (P < 0.0005) analyses indicated that this overexpression is an independent indicator of poor outcome. Forced podocalyxin expression perturbed cell junctions between MCF-7 breast carcinoma cells, and it caused cell shedding from confluent monolayers. Therefore, podocalyxin overexpression is a novel predictor of breast cancer progression that may contribute to the process by perturbing tumor cell adhesion. PMID- 15289307 TI - CpG oligodeoxynucleotide enhances tumor response to radiation. AB - CpG oligodeoxynucleotides (ODNs) are synthetic DNA sequences containing unmethylated cytosine-guanine motifs with potent immunomodulatory effects. Via Toll-like receptor 9 agonism of dendritic cells and B cells, CpG ODNs induce cytokines, activate natural killer cells, and elicit vigorous T-cell responses that lead to significant antitumor effects, including improved efficacy of chemotherapeutic agents. On the basis of these properties of CpG ODNs, we tested whether they also could enhance tumor response to radiotherapy. Using an immunogenic mouse tumor, designated FSa, the response to radiotherapy was assayed by tumor growth delay and tumor cure rate (TCD(50), radiation dose yielding 50% tumor cure rate). Treatments were initiated when established tumors were either 6 or 8 mm in diameter. CpG ODN as a single agent given s.c. peritumorally had little effect on tumor growth; however, it dramatically enhanced tumor growth delay in response to single-dose radiation by a factor of 2.58-2.65. CpG ODN also dramatically improved tumor radiocurability, reducing the TCD(50) by a factor of 1.93, from 39.6 (36.1-43.1) Gy to 20.5 (14.3-25.7) Gy. The CpG ODN-induced enhancement of tumor radioresponse was diminished in tumor-bearing mice immunocompromised by sublethal whole-body radiation. Tumors treated with CpG ODN and radiation showed histologic changes characterized by increased necrosis, heavy infiltration by host inflammatory cells (lymphocytes and granulocytes), and reduced tumor cell density. These results show that CpG ODNs are potent enhancers of tumor radioresponse and as such have potential to improve clinical radiotherapy. PMID- 15289308 TI - p53 upregulates death receptor 4 expression through an intronic p53 binding site. AB - Death receptor 4 (DR4) is one of the tumor necrosis factor-related apoptosis inducing ligand (TRAIL) receptors and triggers apoptosis on ligation with TRAIL or overexpression. Our previous study demonstrated that DR4 expression could be regulated in a p53-dependent fashion. In the present study, we have demonstrated that DR4 is a p53 target gene and is regulated by p53 through a functional intronic p53 binding site (p53BS) based on the following lines of evidence: (a) the p53BS in the DR4 gene is almost identical to the one found in the first intron of the DR5 gene in terms of their locations and sequences; (b) DR4 p53BS bound to p53 protein in intact cells upon p53 activation as demonstrated by a chromatin immunoprecipitation assay; (c) a luciferase reporter vector carrying the DR4 p53BS upstream of an SV40 promoter exhibited enhanced luciferase activity when transiently cotransfected with a wild-type p53 expression vector in p53-null cell lines or stimulated with DNA-damaging agents in a cell line having wild-type p53; and (d) when the DR4 p53BS, together with its own corresponding promoter region in the same orientation as it sits in its natural genomic locus, was cloned into a basic luciferase vector without a promoter element, its transcriptional activity was strikingly increased by cotransfection of a wild type p53 expression vector or treatment with DNA-damaging agents. However, wild type p53 or DNA-damaging agents completely lost their activity to increase transcriptional activity of a reporter construct with deleted DR4 p53BS. Thus, we conclude that p53 directly regulates the expression of the DR4 gene via the novel intronic p53BS. PMID- 15289309 TI - Mutation-selective tumor remission with Ras-targeted, whole yeast-based immunotherapy. AB - Activating mutations in Ras oncoproteins represent attractive targets for cancer immunotherapy, but few vectors capable of generating immune responses required for tumor killing without vector neutralization have been described. Whole recombinant yeast heterologously expressing mammalian mutant Ras proteins were used to immunize mice in a carcinogen-induced lung tumor model. Therapeutic immunization with the whole recombinant yeast caused complete regression of established Ras mutation-bearing lung tumors in a dose-dependent, antigen specific manner. In combination with the genomic sequencing of tumors in patients, the yeast-based immunotherapeutic approach could be applied to treat Ras mutation-bearing human cancers. PMID- 15289310 TI - Autoantibodies to Annexin XI-A and Other Autoantigens in the Diagnosis of Breast Cancer. AB - We report on the identification of autoantigens commonly recognized by sera from patients with breast cancer. We selected ten sera from patients with invasive ductal carcinoma (IDC) of the breast with high titer IgG autoantibodies for biopanning of a T7 phage breast cancer cDNA display library. A high throughput method involved the assembly of 938 T7 phages encoding potential breast cancer autoantigens. Microarrays of positive phages were probed with sera from 90 patients with breast cancer [15 patients with ductal carcinoma in situ (DCIS) and 75 patients with IDC of the breast], with 51 non-cancer control sera and with sera from 21 patients with systemic autoimmune diseases. A 12-phage breast cancer predictor group was constructed with phage inserts recognized by sera from patients with breast cancer and not by non-cancer or autoimmune control sera (P < 0.0001). Several autoantigens including annexin XI-A, the p80 subunit of the Ku antigen, ribosomal protein S6, and other unknown autoantigens could significantly discriminate between breast cancer and non-cancer control sera. Biopanning with three different sera led to the cloning of partial cDNA sequences identical to annexin XI-A. IgG autoantibodies reacting with the amino acid 41-74 sequence of annexin XI-A were found in 19% of all women with breast cancer but in 60% of sera from women with DCIS of the breast. In addition, partial sequences identical to annexin XI-A, nucleolar protein interacting with the forkhead-associated (FHA) domain of pKi-67, the KIAA1671 gene product, ribosomal protein S6, cyclin K, elongation factor-2, Grb2-associated protein 2, and other unknown proteins could distinguish DCIS from IDC of the breast and appear to be potential biomarkers for the diagnosis of breast cancer. PMID- 15289311 TI - Expression of chicken ovalbumin upstream promoter-transcription factor II enhances invasiveness of human lung carcinoma cells. AB - Chicken ovalbumin upstream promoter-transcription factor II (COUP-TFII) plays an essential role in angiogenesis and development. It is differentially expressed in tumor cell lines, but its role in carcinogenesis is largely unknown. We demonstrate here that noninvasive human lung cancer cells become invasive when COUP-TFII was expressed. The expression of extracellular matrix degrading proteinases, such as matrix metalloproteinase 2 and urokinase-type plasminogen activator, was up-regulated in these cells. This finding was confirmed by transduction of different human lung cancer cell lines with COUP-TFII protein and also by using antisense expression. We observed disorganization of actin filaments and focal adhesion kinase phosphorylation in COUP-TFII-transfected human lung cancer cells in addition to the increase in extracellular metalloproteinase activity. These results suggest that COUP-TFII may be considered as a new target for anticancer therapies. PMID- 15289312 TI - Heat shock protein 90 regulates the metaphase-anaphase transition in a polo-like kinase-dependent manner. AB - We have shown previously that the molecular chaperone heat shock protein 90 (Hsp90) is required for a proper centrosome function. Indeed, this Hsp90 function seems to be reflected in Polo-like kinase stability. Inhibition of Hsp90 in HeLa cells results in cell cycle arrest either in G2 stage or at the metaphase anaphase transition. Here, we show that this inhibition leads to inactivation of the anaphase-promoting complex or cyclosome by both dephosphorylation and induction of the spindle assembly checkpoint. Hsp90 inhibition compromises two of the main mitotic kinases, Polo-like kinase 1 (Plk1) and cdc2. Interestingly, this mitotic arrest does not occur in certain tumor cell lines where Hsp90 and Plk1 are not associated. Those cells are able to process mitosis successfully and have an active Plk1 despite Hsp90 inactivation. Therefore, it seems that Hsp90 regulates completion of mitosis depending on its association with Plk1. PMID- 15289313 TI - Multiple familial trichoepithelioma caused by mutations in the cylindromatosis tumor suppressor gene. AB - The recessive oncogene cylindromatosis (CYLD) mapping on 16q12-q13 is generally implicated in familial cylindromatosis, whereas a gene region for multiple familial trichoepithelioma has been assigned to 9p21. Markers from both chromosome intervals were subjected to linkage analysis in a large family with multiple hereditary trichoepithelioma (TE) from Algeria. Linkage to 9p21 was excluded, whereas CYLD remained as a candidate. Mutation analysis identified a single bp germ-line deletion expected to result in truncation or absence of the encoded protein, which segregated with the multiple TE phenotype. In individual tumors, loss of heterozygosity at 16q or a somatic point mutation in the CYLD gene was detected. Hence, mutations of the tumor suppressor gene CYLD at 16q12 q13 may give rise to familial TE indistinguishable from the phenotype assigned to 9p21. PMID- 15289314 TI - p53 heterozygosity results in an increased 2-acetylaminofluorene-induced urinary bladder but not liver tumor response in DNA repair-deficient Xpa mice. AB - Both nucleotide excision repair (NER) and the p53 tumor suppressor protein play crucial roles in the prevention of cells becoming cancerous. This is clearly demonstrated by the fact that NER-deficient xeroderma pigmentosum patients and Li Fraumeni patients who carry a germ-line p53 mutation are highly tumor prone. The NER-deficient Xpa and the p53(+/-) mouse models clearly mimic their human counterparts, because they are both tumor prone as well. The aim of the study presented here was to analyze the relative contribution of these two pathways in tumor suppression and to analyze a possible link between NER and p53 activation in vivo. For this, we exposed Xpa, p53(+/-), and Xpa/p53(+/-) mice to 2 acetylaminofluorene (2-AAF). We show that 2-AAF-induced urinary bladder tumor suppression is dependent on p53 status, because p53(+/-) mice were highly tumor prone. Xpa/p53(+/-) mice were even more tumor prone, whereas no increased tumor response was found in Xpa mice. Short-term assays revealed a decreased apoptotic response in Xpa/p53(+/-) mice, pointing in vivo toward a link between NER and p53 mediated apoptosis. In contrast, liver tumor response was primarily dependent on appropriate DNA repair, because Xpa-deficient mice were liver tumor prone. p53 heterozygosity had no influence on liver tumor incidences, in line with the results obtained from the short-term 2-AAF studies revealing no altered cellular response in p53(+/-) or Xpa/p53(+/-) mice. Interestingly, however, mice completely deficient in both NER and p53 (Xpa/p53(-/-) mice) showed a dramatic increase of hepatocellular proliferation accompanied by lacZ reporter gene mutations. PMID- 15289315 TI - Protein Kinase C theta (PKCtheta) expression and constitutive activation in gastrointestinal stromal tumors (GISTs). AB - KIT expression is a key diagnostic feature of gastrointestinal stromal tumors (GISTs), and virtually all of the GISTs express oncogenic forms of the KIT or PDGFRA receptor tyrosine kinase proteins, which serve as therapeutic targets of imatinib mesylate (Gleevec; Novartis, Basel, Switzerland). However, KIT expression can be low in PDGFRA-mutant GISTs, increasing the likelihood of misdiagnosis as other types of sarcoma. We report that the signaling intermediate protein kinase C theta (PKCtheta) is a diagnostic marker in GISTs, including those that lack KIT expression and/or contain PDGFRA mutations. PKCtheta is strongly activated in most GISTs and hence may serve, along with KIT/PDGFRA, as a novel therapeutic target. PMID- 15289316 TI - TBX3 and its isoform TBX3+2a are functionally distinctive in inhibition of senescence and are overexpressed in a subset of breast cancer cell lines. AB - TBX3 is a transcription factor of the T-box gene family. Mutations of TBX3 cause ulnar-mammary syndrome (MIM 181450) in humans, an autosomal dominant disorder characterized by the absence or underdevelopment of the mammary glands and other congenital anomalies. It recently was found that TBX3 was able to immortalize mouse embryo fibroblast (MEF) cells. In addition, TBX2, a homologue of TBX3, is active in preventing senescence in rodent cells and was found to be amplified in some human breast cancers, suggesting TBX3 plays a role in breast cancer. This study examined the function of TBX3 and its isoform, TBX3 + 2a. TBX3 + 2a differs from TBX3 in the DNA binding domain with an extra 20 amino acids produced by alternative splicing. We first examined the tissue expression and alternative splicing patterns of these two isoforms. We found that TBX3 and TBX3 + 2a are widely expressed in humans and mice, and alternative splicing could be tissue specific and species specific. Overexpression of TBX3 is able to immortalize MEF cells, whereas TBX3 + 2a shows an acceleration of senescence, a functional difference that may be explained by the fact that these two isoforms may have different downstream targets. TBX3, but not TBX3 + 2a, is able to bind to the previously identified T-box binding site in a gel shift assay. A subset of human breast cancer cell lines overexpresses TBX3. Our results indicate that TBX3 and TBX3 + 2a are functionally distinctive in inhibition of senescence of MEF cells and may play a role in breast cancer. PMID- 15289317 TI - Loss of heterozygosity occurs via mitotic recombination in Trp53+/- mice and associates with mammary tumor susceptibility of the BALB/c strain. AB - Loss of heterozygosity (LOH) occurs commonly in cancers causing disruption of tumor suppressor genes and promoting tumor progression. BALB/c-Trp53(+/-) mice are a model of Li-Fraumeni syndrome, exhibiting a high frequency of mammary tumors and other tumor types seen in patients. However, the frequency of mammary tumors and LOH differs among strains of Trp53(+/-) mice, with mammary tumors occurring only on a BALB/c genetic background and showing a high frequency of LOH, whereas Trp53(+/-) mice on a 129/Sv or (C57BL/6 x 129/Sv) mixed background have a very low frequency of mammary tumors and show LOH for Trp53 in only approximately 50% of tumors. We have performed studies on tumors from Trp53(+/-) mice of several genetic backgrounds to examine the mechanism of LOH in BALB/c Trp53(+/-) mammary tumors. By Southern blotting, 96% (24 of 25) of BALB/c Trp53(+/-) mammary tumors displayed LOH for Trp53. Karyotype analysis indicated that cells lacking one copy of chromosome 11 were present in all five mammary tumors analyzed but were not always the dominant population. Comparative genomic hybridization analysis of these five tumors indicated either loss or retention of the entire chromosome 11. Thus chromosome loss or deletions within chromosome 11 do not account for the LOH observed by Southern blotting. Simple sequence length polymorphism analysis of (C57BL/6 x BALB/c) F1-Trp53(+/-) mammary tumors showed that LOH occurred over multiple loci and that a combination of maternal and paternal alleles were retained, indicating that mitotic recombination is the most likely mechanism of LOH. Nonmammary tumors of BALB/c mice also showed a high frequency of LOH (22 of 26, 85%) indicating it was not a mammary tumor specific phenomenon but rather a feature of the BALB/c strain. In (C57BL/6 x BALB/c) F1 Trp53(+/-) mice LOH was observed in 93% (13 of 14) of tumors, indicating that the high frequency of LOH was a dominant genetic trait. Thus the high frequency of LOH for Trp53 in BALB/c-Trp53(+/-) mammary tumors occurs via mitotic recombination and is a dominant genetic trait that associates with the occurrence of mammary tumors in (C57BL/6 x BALB/c) F1-Trp53(+/-) mice. These results further implicate double-strand DNA break repair machinery as important contributors to mammary tumorigenesis. PMID- 15289318 TI - Effect of N-acetyl cysteine on oxidative DNA damage and the frequency of DNA deletions in atm-deficient mice. AB - Ataxia telangiectasia (AT) is a hereditary human disorder resulting in a wide variety of clinical manifestations, including progressive neurodegeneration, immunodeficiency, and high incidence of lymphoid tumors. Cells from patients with AT show genetic instability, hypersensitivity to radiation, and a continuous state of oxidative stress. Oxidative stress and genetic instability, including DNA deletions, are involved in carcinogenesis. We examined the effect of dietary supplementation with the thiol-containing antioxidant N-acetyl-l-cysteine (NAC) on levels of oxidative DNA damage and the frequency of DNA deletions in Atm deficient (AT-mutated) mice. We confirmed that Atm-deficient mice display an increased frequency of DNA deletions (Bishop et al., Cancer Res 2000;60:395). Furthermore, we found that Atm-deficient mice have significantly increased levels of 8-OH deoxyguanosine, an indication of oxidative DNA damage. Dietary supplementation with NAC significantly reduced 8-OH deoxyguanosine level and the frequency of DNA deletions in Atm-deficient mice. These levels were similar to the levels in wild-type mice. Our findings demonstrate that NAC counteracts genetic instability and suggest that genetic instability may be a consequence of oxidative stress in Atm-deficient mice. PMID- 15289319 TI - Truncated RON tyrosine kinase drives tumor cell progression and abrogates cell cell adhesion through E-cadherin transcriptional repression. AB - RON is a tyrosine kinase receptor that triggers scattering of normal cells and invasive growth of cancer cells on ligand binding. We identified a short RON mRNA, which is expressed in human lung, ovary, tissues of the gastrointestinal tract, and also in several human cancers, including ovarian carcinomas and cell lines from pancreatic carcinomas and leukemias. This transcript encodes a truncated protein (short-form RON; sf-RON), lacking most of the RON receptor extracellular domain but retaining the whole transmembrane and intracellular domains. Sf-RON shows strong intrinsic tyrosine kinase activity and is constitutively phosphorylated. Epithelial cells transduced with sf-RON display an aggressive phenotype; they shift to a nonepithelial morphology, are unable to form aggregates, grow faster in monolayer cultures, show anchorage-independent growth, and become motile. We show that in these cells, E-cadherin expression is lost through a dominant transcriptional repression pathway likely mediated by the transcriptional factor SLUG. Altogether, these data show that expression of a naturally occurring, constitutively active truncated RON kinase results in loss of epithelial phenotype and aggressive behavior and, thus, it might contribute to tumor progression. PMID- 15289320 TI - Inhibition of activator protein 1 activation, vascular endothelial growth factor, and cyclooxygenase-2 expression by 15-deoxy-Delta12,14-prostaglandin J2 in colon carcinoma cells: evidence for a redox-sensitive peroxisome proliferator-activated receptor-gamma-independent mechanism. AB - Cyclooxygenase (COX)-2 and vascular endothelial growth factor (VEGF) are significantly associated with tumor growth and metastasis. Here we show that phorbol ester-mediated induction of VEGF and COX-2 expression in colon carcinoma cells is inhibited by 15-deoxy-Delta(12,14)-prostaglandin J(2) (15d-PGJ(2)). This cyclopentenone was able to inhibit activator protein1 (AP-1)-dependent transcriptional induction of COX-2 and VEGF promoters induced by phorbol 12 myristate 13-acetate (PMA) or c-Jun overexpression. 15d-PGJ(2) interfered with at least two steps within the signaling pathway leading to AP-1 activation. First, 15d-PGJ(2) impaired AP-1 binding to a consensus DNA sequence. Second, 15d-PGJ(2) selectively inhibited c-Jun NH(2) terminal kinase (JNK) but not extracellular signal-regulated kinase or p38 mitogen-activated protein kinase activation induced by PMA. This led to a decreased ability of JNK to phosphorylate c-Jun and to activate its transactivating activity. Inhibition of AP-1 activation and COX-2 or VEGF transcriptional induction by this cyclopentenone was found to be independent of peroxisome proliferator-activated receptor-gamma (PPARgamma) because it was not affected by either expression of a dominant negative form of PPARgamma or the use of a PPARgamma antagonist. In contrast, we have found that the effects of 15d-PGJ(2) on AP-1 activation may occur through its ability to induce intracellular oxidative stress. The antioxidant N-acetylcysteine significantly reversed the inhibition by 15d-PGJ(2) of AP-1 activity and COX-2 or VEGF transcriptional induction. Together, these findings provide new insight into the antitumoral properties of 15d-PGJ(2) through the inhibition of the induction of AP-1-dependent genes involved in tumor progression, such as COX-2 and VEGF. PMID- 15289321 TI - Apoptotic speck protein-like, a highly homologous protein to apoptotic speck protein in the pyrin domain, is silenced by DNA methylation and induces apoptosis in human hepatocellular carcinoma. AB - We have identified a novel gene encoding a pyrin domain protein of 89 amino acids that is expressed in various tissues including liver, brain, and spleen. The protein is highly homologous to the pyrin domain of apoptosis-associated speck like protein (ASC). Therefore, we termed it ASC-like (ASCL). We found that ASCL gene was densely and frequently (80%) methylated in hepatocellular carcinoma (HCC) cell lines. In contrast, normal liver samples did not show any significant methylation. This aberrant methylation correlated well with the suppression of RNA expression. Furthermore, a demethylating agent, 5-aza-2'-deoxycytidine, reactivated the ASCL expression in the methylation-silenced cells, indicating that ASCL is silenced by the associated DNA methylation. ASCL methylation was also found in primary HCC (4 of 17 samples), although the frequency was less than that in cell lines. In addition, we found that ASC was also methylated in primary samples (6 of the 17). Interestingly, either ASCL or ASC methylation was observed in 53% (9 of the 17) of primary HCC samples. Significantly, the restoration of ASCL in the methylation-silenced cells demonstrated growth suppression in colony formation assay. This growth suppression effect of ASCL was supported by apoptotic changes observed in ASCL-transfected cells in which annexin-V binding was positive and caspase-3 was activated. Based on the methylation-silencing and the growth suppression activity, we propose that ASCL plays a significant role in the development of HCC. PMID- 15289322 TI - Host stromal bradykinin B2 receptor signaling facilitates tumor-associated angiogenesis and tumor growth. AB - We evaluated the significance of the host kallikrein-kinin system in tumor angiogenesis and tumor growth using two rodent models genetically deficient in a kallikrein-kinin system. Inoculation of Walker 256 carcinoma cells into the s.c. tissues of the back of normal Brown Norway Kitasato rats (BN-Ki rats) resulted in the rapid development of solid tumors with marked angiogenesis. By contrast, in kininogen-deficient Brown Norway Katholiek rats (BN-Ka rats), which cannot generate intrinsic bradykinin (BK), the weights of the tumors and the extent of angiogenesis were significantly less than those in BN-Ki rats. Daily administration of B(2) receptor antagonists significantly reduced angiogenesis and tumor weights in BN-Ki rats to levels similar to those in BN-Ka rats but did not do so in BN-Ka rats. Angiogenesis and tumor growth were significantly suppressed in B(2) receptor knockout mice bearing sarcoma 180 compared with their wild-type counterparts. Immunoreactive vascular endothelial growth factor (VEGF) was localized in Walker tumor stroma more extensively in BN-Ki rats than in BN-Ka rats, although immunoreactive B(2) receptor also was detected in the stroma to the same extent in both types of rats. Cultured stromal fibroblasts isolated from BN-Ki rats and BN-Ka rats produced VEGF in response to BK (10(-8)-10(-6) m), and this stimulatory effect of BK was abolished with a B(2) receptor antagonist, Hoe140 (10(-5) m). These results suggest that BK generated from kininogens supplied from the host may facilitate tumor-associated angiogenesis and tumor growth by stimulating stromal B(2) signaling to up-regulate VEGF production mainly in fibroblasts. PMID- 15289323 TI - Reduction in Raf kinase inhibitor protein expression is associated with increased Ras-extracellular signal-regulated kinase signaling in melanoma cell lines. AB - Mutations in the Raf signaling pathway are known to play a pivotal role in the progression of malignant melanoma. In this study, we provide evidence that the Raf-1 kinase inhibitory protein (RKIP) and its effects on Raf-1-mediated activation of mitogen-activated protein/extracellular signal-regulated kinase kinase are important for the metastatic potential of malignant melanoma. Screening nine melanoma cell lines at mRNA and protein levels, we detected significant down-regulation of RKIP expression in comparison with normal melanocytes. Loss of RKIP expression in transformed cells in vivo was confirmed in immunohistochemical analyses demonstrating reduction of RKIP expression already in primary melanoma and even stronger down-regulation or complete loss in melanoma metastases. Stable transfection of the melanoma cell line Mel Im with an RKIP expression plasmid blocked the Raf kinase pathway, resulting in down regulation of extracellular signal-regulated kinase 1/2 and activator protein 1 activity. In very good agreement with the in vivo finding that down-regulation of RKIP expression is most obvious in melanoma metastasis, overexpression of RKIP in the highly invasive Mel Im cell line leads to a significant inhibition of invasiveness in vitro. Taken together, our results suggest that loss of RKIP in malignant melanoma contributes to enhanced invasiveness of transformed cells and therefore to progression of the disease. PMID- 15289324 TI - Progression of pregnancy-dependent mouse mammary tumors after long dormancy periods. Involvement of Wnt pathway activation. AB - Mouse mammary tumor virus (LA) induces pregnancy-dependent mammary tumors that progress toward autonomy. Here we show that in virgin females, pregnancy dependent tumor transplants are able to remain dormant for up to 300 days. During that period, these tumors synthesize DNA, express high levels of estrogen and progesterone receptors (ER+PR+) and are able to resume growth after hormone stimulation. Surprisingly, in a subsequent transplant generation, all these tumors are fully able to grow in virgin females, they express low levels of ER and PR (ER-PR-) and have a monoclonal origin; i.e., show all of the features we have described previously in pregnancy-independent tumors. Histologically, mouse mammary tumor virus (LA)-induced tumors are morphologically similar to genetically engineered mouse (GEM) mammary tumors that overexpress genes belonging to the Wnt pathway. Interestingly, in the virus-induced neoplasias, pregnancy-independent passages arising after a dormant phase usually display a lower level of glandular differentiation together with epithelial cell trans differentiation, a specific feature associated to Wnt pathway activation. In addition, dormancy can lead to the specific selection of Int2/Fgf3 mutated and overexpressing cells. Therefore, our results indicate that during hormone dependent tumor dormancy, relevant changes in cell population occur, allowing rapid progression after changes in the animal internal milieu. PMID- 15289325 TI - Targeting endogenous transforming growth factor beta receptor signaling in SMAD4 deficient human pancreatic carcinoma cells inhibits their invasive phenotype1. AB - Transforming growth factor-beta (TGF-beta) suppresses tumor formation by blocking cell cycle progression and maintaining tissue homeostasis. In pancreatic carcinomas, this tumor suppressive activity is often lost by inactivation of the TGF-beta-signaling mediator, Smad4. We found that human pancreatic carcinoma cell lines that have undergone deletion of MADH4 constitutively expressed high endogenous levels of phosphorylated receptor-associated Smad proteins (pR-Smad2 and pR-Smad3), whereas Smad4-positive lines did not. These elevated pR-Smad levels could not be attributed to a decreased dephosphorylation rate nor to increased expression of TGF-beta type I (TbetaR-I) or type II (TbetaR-II) receptors. Although minimal amounts of free bioactive TGF-beta1 and TGF-beta2 were detected in conditioned medium, treatment with a pan-specific (but not a TGF beta3 specific) TGF-beta-neutralizing antibody and with anti-alpha(V)beta(6) integrin antibody decreased steady-state pSmad2 levels and activation of a TGF beta-inducible reporter gene in neighboring cells, respectively. Thus, activation of TGF-beta at the cell surface was responsible for the increased autocrine endogenous and paracrine signaling. Blocking TbetaR-I activity using a selective kinase inhibitor (SD-093) strongly decreased the in vitro motility and invasiveness of the pancreatic carcinoma cells without affecting their growth characteristics, morphology, or the subcellular distribution of E-cadherin and F actin. Moreover, exogenous TGF-beta strongly stimulated in vitro invasiveness of BxPC-3 cells, an effect that could also be blocked by SD-093. Thus, the motile and invasive properties of Smad4-deficient pancreatic cancer cells are at least partly driven by activation of endogenous TGF-beta signaling. Therefore, targeting the TbetaR-I kinase represents a potentially powerful novel therapeutic approach for the treatment of this disease. PMID- 15289326 TI - Class IV semaphorins promote angiogenesis by stimulating Rho-initiated pathways through plexin-B. AB - The semaphorins are a large family of secreted and cell surface proteins that provide attractive and repulsive cues for axon guidance during neuronal development. Semaphorins share a conserved NH(2)-terminal Sema domain with their receptors, the plexins, which mediate neuronal cell adhesion, axon guidance, and maintenance of established neuronal pathways in the adult. Both semaphorins and plexins share structural homology with the extracellular domain of c-Met, a member of the scatter factor family of receptors. However, the highly conserved cytoplasmic region of plexins has no homology with the c-Met tyrosine kinase or with any other known protein. Using a recently developed antibody and RNA analysis, we found that high levels of plexin-B1 are expressed in endothelial cells. Whereas c-Met, with which plexin-B1 can interact, is known to be a potent promoter of angiogenesis, the effects of semaphorin-mediated plexin activation in endothelial cells are still poorly understood. Here, we examined the role of plexin-B1 activation in angiogenesis using a purified, secreted form of its ligand, Semaphorin 4D (Sema4D). Sema4D potently induced chemotaxis and tubulogenesis in endothelial cells and enhanced blood vessel formation in an in vivo mouse model. Interestingly, responses to Sema4D did not require c-Met activation. Instead, the use of chimeric plexin-B1 receptors, Rho inhibitors, and lentiviral gene delivery of interfering molecules revealed that these proangiogenic effects are dependent on a COOH-terminal PDZ-binding motif of plexin-B1, which binds two guanine nucleotide exchange factors for the small GTPase Rho, PDZ-RhoGEF and LARG, and are mediated by the activation of Rho initiated pathways. PMID- 15289327 TI - Cytoplasmic mislocalization of p27Kip1 protein is associated with constitutive phosphorylation of Akt or protein kinase B and poor prognosis in acute myelogenous leukemia. AB - Cyclin-dependent kinase inhibitor p27Kip1 functions at the nuclear level by binding to cyclin E/cyclin-dependent kinase-2. It was shown that Akt or protein kinase B (Akt/PKB)-dependent phosphorylation of p27Kip1 led to the cytoplasmic mislocalization of p27Kip1, suggesting the potential abrogation of its activity. Here, we evaluated the localization of p27Kip1 protein in leukemic blasts in relation to Akt/PKB phosphorylation and clinical outcomes in acute myelogenous leukemia (AML). Western blot analysis of the nuclear and cytoplasmic fractions revealed a heterogenous localization pattern of p27Kip1 in AML. Cytoplasmic mislocalization of p27Kip1 was significantly associated with the constitutive serine(473) Akt/PKB phosphorylation in AML cells (P < 0.05). Transfection of U937 cells with an expression construct encoding the constitutively active form of Akt/PKB resulted in a remarkable increase in the levels of cytoplasmic p27Kip1. Whereas the transfection of U937 cells with a construct encoding dominant negative Akt/PKB resulted in a recovery of nuclear localization of p27Kip1. Both the disease-free survival and overall survival are significantly shorter in AML cases with high cytoplasmic to nuclear ratio of p27Kip1 localization compared with the cases with low cytoplasmic to nuclear ratio (P = 0.0353, P = 0.0023, respectively). Multivariate analysis indicated that the cytoplasmic to nuclear ratio of p27Kip1 localization was an independent prognostic variable for both disease-free survival and overall survival (P = 0.043, P = 0.008, respectively). These findings additionally extend our understanding of the role of p27Kip1 in AML, and buttress the case of p27Kip1 mislocalization as a prognostic indicator and Akt/PKB/p27Kip1 pathway as a ready target for antileukemia therapy. PMID- 15289328 TI - Phosphorylation of Akt (Ser473) is an excellent predictor of poor clinical outcome in prostate cancer. AB - We previously showed, by immunohistochemistry with phospho-specific antibodies, increased phosphorylation (activation) of Akt (Ser(473)) [phosphorylated Akt (pAkt)] in high-Gleason grade prostate cancer (Malik SN, et al., Clin Cancer Res 2002;8:1168-71). Elevation of pAkt was accompanied by decreased phosphorylation of extracellular signal-regulated kinase (ERK) 1/2 (Thr(202)/Tyr(204)) [phosphorylated ERK (pERK)], indicative of inactivation. In this report, we determined whether increased pAkt and decreased pERK predicted clinical outcome. Prostate-specific antigen (PSA) failure (detectable and rising PSA) versus PSA non-failure (undetectable PSA 5 years after prostatectomy) was used as a surrogate for clinical outcome. Prostate tumors from cases of PSA failure versus non-failure were stained for pAkt and pERK. A significant increase in mean pAkt staining (P < 0.001) in the PSA failures versus non-failures was seen based on the Wilcoxon signed ranks test [222.18 +/- 33.9 (n = 37) versus 108.79 +/- 104.57 (n = 16)]. Using the best-fitting multiple logistic regression equation, a 100 point increase in pAkt staining resulted in a 160% increase in the odds of being a PSA failure. There was decreased staining for pERK in PSA failures versus non failures: a 100-point decrease resulted in an 80% increase in the odds of being a PSA failure. Each of these effects assumed the other biomarker was held constant. The area under the receiver-operating characteristic curve for these two biomarkers predicting PSA failure was 0.84, indicating excellent discrimination between PSA failure and non-failure cases. These data indicate that increased pAkt, alone or together with decreased pERK, is an important predictor of probability of PSA failure. However, pERK alone was not a significant predictor of PSA failure. PMID- 15289329 TI - IRSp53/Eps8 complex is important for positive regulation of Rac and cancer cell motility/invasiveness. AB - IRSp53 has been characterized as an adaptor protein that links Rho-family small GTPases, such as Rac, to reorganization of the actin cytoskeleton. Here, we search for other binding partners for the IRSp53 SH3 domain and identify Eps8 as the major binding protein in fibroblasts and various cancer cell lines. Eps8 has been shown to form a Rac-specific guanine nucleotide exchange factor complex with Abi-1 and Sos-1, which seems essential for ruffling formation induced by oncogenic Ras. We confirm the IRSp53/Eps8 complex formation in vivo and the direct association between Eps8 NH(2)-terminal proline-rich sequence and IRSp53 SH3 domain. This complex synergistically activates Rac by reinforcing the formation of the Eps8/Abi-1/Sos-1 Rac-guanine nucleotide exchange factor complex, which mediates positive regulation of Rac activity. In addition, IRSp53/Eps8 complex formation as determined by fluorescent resonance energy transfer analysis, occurs at the leading edge of motile cells, and the motility and invasiveness of HT1080 fibrosarcoma cells are suppressed by inhibiting complex formation. These findings implicate the importance of the IRSp53/Eps8 complex in Rac activation and metastatic behavior of the malignant tumor cells. PMID- 15289330 TI - Normalization of real-time quantitative reverse transcription-PCR data: a model based variance estimation approach to identify genes suited for normalization, applied to bladder and colon cancer data sets. AB - Accurate normalization is an absolute prerequisite for correct measurement of gene expression. For quantitative real-time reverse transcription-PCR (RT-PCR), the most commonly used normalization strategy involves standardization to a single constitutively expressed control gene. However, in recent years, it has become clear that no single gene is constitutively expressed in all cell types and under all experimental conditions, implying that the expression stability of the intended control gene has to be verified before each experiment. We outline a novel, innovative, and robust strategy to identify stably expressed genes among a set of candidate normalization genes. The strategy is rooted in a mathematical model of gene expression that enables estimation not only of the overall variation of the candidate normalization genes but also of the variation between sample subgroups of the sample set. Notably, the strategy provides a direct measure for the estimated expression variation, enabling the user to evaluate the systematic error introduced when using the gene. In a side-by-side comparison with a previously published strategy, our model-based approach performed in a more robust manner and showed less sensitivity toward coregulation of the candidate normalization genes. We used the model-based strategy to identify genes suited to normalize quantitative RT-PCR data from colon cancer and bladder cancer. These genes are UBC, GAPD, and TPT1 for the colon and HSPCB, TEGT, and ATP5B for the bladder. The presented strategy can be applied to evaluate the suitability of any normalization gene candidate in any kind of experimental design and should allow more reliable normalization of RT-PCR data. PMID- 15289331 TI - Differential signaling pathways are activated in the Epstein-Barr virus associated malignancies nasopharyngeal carcinoma and Hodgkin lymphoma. AB - EBV is associated with the epithelial cancer, nasopharyngeal carcinoma (NPC), and the lymphoid malignancy, Hodgkin lymphoma (HL). The EBV latent membrane proteins 1 and 2A are expressed in these tumors. These proteins activate the phosphatidylinositol 3'-OH kinase (PI3K)/Akt pathway, which is commonly activated inappropriately in malignancy. In this study, the status of Akt activation and its targets, glycogen synthase kinase-3beta (GSK-3beta) and beta-catenin, was investigated in NPC and HL clinical specimens. In the majority of HL and NPC specimens, Akt was activated, indicating an important role for this kinase in the development and/or progression of these tumors. Akt phosphorylates and inactivates GSK-3beta, a negative regulator of the proto-oncoprotein beta-catenin that is aberrantly activated in many cancers. GSK-3beta was phosphorylated and inactivated with concomitant nuclear beta-catenin accumulation in the majority of NPC specimens. The malignant cells of the majority of HL cases, however, did not have inactivated GSK-3beta and lacked nuclear beta-catenin expression. These data indicate that this signaling arm of PI3K/Akt is universal and important in NPC pathogenesis but is apparently not affected in HL. These findings point to a divergence in pathways activated by EBV in different cellular contexts. PMID- 15289332 TI - Rolling of human bone-metastatic prostate tumor cells on human bone marrow endothelium under shear flow is mediated by E-selectin. AB - Prostate tumor cells preferentially adhere to bone marrow endothelial cells (BMECs) compared with endothelial linings from other tissue microvessels, implicating the importance of BMEC adhesion in the predilection of prostate tumor metastasis to bone. E (endothelial)-selectin, which functions as an initiator of leukocyte adhesion to target tissue endothelium, is constitutively expressed on BMECs, suggesting that prostate tumor cells could use this adhesive mechanism to initiate their migration into bone. In this report, we demonstrate for the first time that human bone-metastatic prostate tumor cells roll on human BMECs under physiological flow conditions. We show that these dynamic adhesive interactions are dependent on the expression of BMEC E-selectin and sialylated glycoconjugates on bone-metastatic prostate tumor cells. We also establish the importance of both glycoprotein(s) and glycosphingolipid structures displaying sialyl Lewis X epitopes as potential E-selectin ligands on bone-metastatic prostate tumor cells. Coexpression of sialylated glycoproteins and glycolipids on bone-metastatic prostate tumor cells triggers robust E-selectin binding activity, which is identical to that observed on human hematopoietic progenitor cells. By Western blot analysis, we identify candidate E-selectin glycoprotein ligand(s); distinct sialyl Lewis X (or HECA-452 antigen)-bearing membrane proteins were resolved at M(r) 130,000 and M(r) 220,000 as well as others ranging from M(r) 100,000 to M(r) 220,000. Immunohistochemical analysis of HECA-452 antigen expression on normal prostate tissue and on low- and high-grade prostate adenocarcinoma shows that HECA-452 antigen expression is directly associated with prostate tumor progression and may indicate acquisition of E-selectin ligand expression. These findings provide novel insight into potential adhesive mechanisms promoting hematogenous dissemination of prostate tumor cells into bone. PMID- 15289333 TI - Expression profiling reveals novel pathways in the transformation of melanocytes to melanomas. AB - Affymetrix and spotted oligonucleotide microarrays were used to assess global differential gene expression comparing normal human melanocytes with six independent melanoma cell strains from advanced lesions. The data, validated at the protein level for selected genes, confirmed the overexpression in melanoma cells relative to normal melanocytes of several genes in the growth factor/receptor family that confer growth advantage and metastasis. In addition, novel pathways and patterns of associated expression in melanoma cells not reported before emerged, including the following: (a) activation of the NOTCH pathway; (b) increased Twist expression and altered expression of additional transcriptional regulators implicated in embryonic development and epidermal/mesenchymal transition; (c) coordinated activation of cancer/testis antigens; (d) coordinated down-regulation of several immune modulation genes, in particular in the IFN pathways; (e) down-regulation of several genes implicated in membrane trafficking events; and (f) down-regulation of growth suppressors, such as the Prader-Willi gene NECDIN, whose function was confirmed by overexpression of ectopic Flag-necdin. Validation of differential expression using melanoma tissue microarrays showed that reduced ubiquitin COOH-terminal esterase L1 in primary melanoma is associated with worse outcome and that increased expression of the basic helix-loop-helix protein Twist is associated with worse outcome. Some differentially expressed genes reside on chromosomal regions displaying common loss or gain in melanomas or are known to be regulated by CpG promoter methylation. These results provide a comprehensive view of changes in advanced melanoma relative to normal melanocytes and reveal new targets that can be used in assessing prognosis, staging, and therapy of melanoma patients. PMID- 15289334 TI - HB-EGF is a potent inducer of tumor growth and angiogenesis. AB - Heparin-binding epidermal growth factor-like growth factor (HB-EGF) has been shown to stimulate the growth of a variety of cells in an autocrine or paracrine manner. Although HB-EGF is widely expressed in tumors compared with normal tissue, its contribution to tumorigenicity is unknown. HB-EGF can be produced as a membrane-anchored form (pro-HB-EGF) and later processed to a soluble form (s-HB EGF), although a significant amount of pro-HB-EGF remains uncleaved on the cell surface. To understand the roles of two forms of HB-EGF in promoting tumor growth, we have studied the effects of HB-EGF expression in the process of tumorigenesis using in vitro and in vivo systems. We demonstrate here that in EJ human bladder cancer cells containing a tetracycline-regulatable s-HB-EGF or pro HB-EGF expression system, s-HB-EGF expression increased their transformed phenotypes, including growth rate, colony-forming ability, and activation of cyclin D1 promoter, as well as induction of vascular endothelial growth factor in vitro. Moreover, s-HB-EGF or wild-type HB-EGF induced the expression and activities of the metalloproteases, MMP-9 and MMP-3, leading to enhanced cell migration. In vivo studies also demonstrated that tumor cells expressing s-HB-EGF or wild-type HB-EGF significantly enhanced tumorigenic potential in athymic nude mice and exerted an angiogenic effect, increasing the density and size of tumor blood vessels. However, cells expressing solely pro-HB-EGF did not exhibit any significant tumorigenic potential. These findings establish s-HB-EGF as a potent inducer of tumor growth and angiogenesis and suggest that therapeutic intervention aimed at the inhibition of s-HB-EGF functions may be useful in cancer treatment. PMID- 15289335 TI - Activation of phosphatidylinositol 3-kinase and extracellular signal-regulated kinase is required for glial cell line-derived neurotrophic factor-induced migration and invasion of pancreatic carcinoma cells. AB - Pancreatic carcinoma cells exhibit a pronounced tendency to invade along and into intra- and extrapancreatic nerves, even at early stages of the disease. The neurotrophic factor glial cell line-derived neurotrophic factor (GDNF) has been shown to promote pancreatic cancer cell invasion. Here, we demonstrate that pancreatic carcinoma cell lines, such as PANC-1, expressed the RET and GDNF family receptor alpha receptor components for GDNF and that primary pancreatic tumor samples, derived from carcinomas with regional lymph node metastasis, exhibited marked expression of the mRNA encoding the RET51 isoform. Moreover, GDNF was an efficacious and potent chemoattractant for pancreatic carcinoma cells as examined in in vitro and in vivo model systems. Treatment of PANC-1 cells with GDNF resulted in activation of the monomeric GTPases N-Ras, Rac1, and RhoA, in activation of the mitogen-activated protein kinases extracellular signal regulated kinase (ERK) and c-Jun NH(2)-terminal kinase (JNK) and in activation of the phosphatidylinositol 3-kinase/Akt pathway. Both inhibition of the Ras-Raf-MEK (mitogen-activated protein/ERK kinase)-ERK cascade by either stable expression of dominant-negative H-Ras(N17) or addition of the MEK1 inhibitor PD98059 as well as inhibition of the phosphatidylinositol 3-kinase pathway by LY294002 prevented GDNF-induced migration and invasion of PANC-1 cells. These results demonstrate that pancreatic tumor cell migration and possibly perineural invasion in response to GDNF is critically controlled by activation of the Ras-Raf-MEK-ERK and the phosphatidylinositol 3-kinase pathway. PMID- 15289336 TI - Sensitization to the lysosomal cell death pathway upon immortalization and transformation. AB - Tumorigenesis is associated with several changes that alter the cellular susceptibility to programmed cell death. Here, we show that immortalization and transformation sensitize cells in particular to the cysteine cathepsin-mediated lysosomal death pathway. Spontaneous immortalization increased the susceptibility of wild-type murine embryonic fibroblasts (MEFs) to tumor necrosis factor (TNF) mediated cytotoxicity >1000-fold, whereas immortalized MEFs deficient for lysosomal cysteine protease cathepsin B (CathB) retained the resistant phenotype of primary cells. This effect was specific for cysteine cathepsins, because also lack of cathepsin L (a lysosomal cysteine protease), but not that of cathepsin D (a lysosomal aspartyl protease) or caspase-3 (the major executioner protease in classic apoptosis) inhibited the immortalization-associated sensitization of MEFs to TNF. Oncogene-driven transformation of immortalized MEFs was associated with a dramatic increase in cathepsin expression and additional sensitization to the cysteine cathepsin-mediated death pathway. Importantly, exogenous expression of CathB partially reversed the resistant phenotype of immortalized CathB-deficient MEFs, and the inhibition of CathB activity by pharmacological inhibitors or RNA interference attenuated TNF-induced cytotoxicity in immortalized and transformed wild-type cells. Thus, tumorigenesis-associated changes in lysosomes may counteract cancer progression and enhance therapeutic responses by sensitizing cells to programmed cell death. PMID- 15289337 TI - Gene expression of angiogenic factors correlates with metastatic potential of prostate cancer cells. AB - We hypothesize that expression of proangiogenic genes correlates with the metastatic potential of prostate cancer cells. LNCaP, DU-145, and PC-3 are prostate cancer cell lines with low, moderate, and high metastatic potential, respectively, as we demonstrated by their capacity to invade an extracellular matrix, an established tumor invasion assay. The constitutive gene expression of the proangiogenic factors, vascular endothelial growth factor, intercellular adhesion molecule-1, interleukin-8, and transforming growth factor-beta2, was significantly greater in the more metastatic DU-145 and PC-3 cells as compared with LNCaP cells. Matrix metalloproteinase (MMP)-9 is thought to contribute to the invasive phenotype of tumor cells. PC-3 cells showed increased expression of MMP-9 and membrane type 4-MMP as compared with LNCaP and DU-145. Tissue inhibitors of metalloproteinase 1 and 4 gene expression were elevated in DU-145 and PC-3 cells, but paradoxically, LNCaP cells had undetectable levels of these genes. We transfected and overexpressed MMP-9 in poorly metastatic LNCaP cells and measured their invasive activity. Transient expression of human MMP-9 in LNCaP cells produced a 3-5-fold increase in MMP-9 activity with a comparable increase in invasiveness. Antisense ablation of the expression of MMP-9 in DU-145 and PC-3 cells produced concomitant inhibition of the gene expression of the proangiogenic factors, vascular endothelial growth factor, and intercellular adhesion molecule-1 (ICAM-1). Treatment of DU-145 and PC-3 cells with a selective chemical inhibitor of MMP-9 proteinase activity also inhibited their invasive activity. These results support our hypothesis that metastatic potential of prostate cancer cells correlates with expression of proangiogenic factors. PMID- 15289338 TI - Effect of mutational inactivation of tyrosine kinase activity on BCR/ABL-induced abnormalities in cell growth and adhesion in human hematopoietic progenitors. AB - Chronic myelogenous leukemia (CML) results from transformation of a primitive hematopoietic cell by the BCR/ABL gene. The specific BCR/ABL signaling mechanisms responsible for transformation of primitive human hematopoietic cells are not well defined. Previous studies have suggested that constitutively activated tyrosine kinase activity plays an important role for in abnormal proliferation of CML progenitors but has not clearly defined its role in abnormal adhesion and migration. We established a human progenitor model of CML by ectopic expression of BCR/ABL in normal CD34+ cells using retrovirus-mediated gene transfer. CD34+ cells expressing BCR/ABL demonstrated several features characteristic of primary CML progenitors including increased proliferation in committed and primitive progenitor culture, reduced adhesion to fibronectin, and reduced chemotaxis to stroma-derived factor-1alpha. We expressed a kinase-inactive BCR/ABL gene to directly investigate the role of kinase activity in abnormal progenitor function. Abnormalities in proliferation were completely reversed, whereas defects in adhesion and migration were significantly improved but not completely reversed in cells expressing a kinase-inactive BCR/ABL. Furthermore, the BCR/ABL kinase inhibitor imatinib mesylate markedly inhibited proliferation of BCR/ABL expressing progenitors but did not fully correct the adhesion and migration defects. Expression of BCR/ABL genes with deletions of either the COOH-terminal actin binding or proline-rich domains resulted in enhanced adhesion and chemotaxis compared with wild-type BCR/ABL but did not affect progenitor proliferation. We conclude that abnormal kinase activity is essential for abnormal proliferation and survival of CML progenitors but that abnormal adhesion and migration result from both kinase-dependent and -independent mechanisms. PMID- 15289339 TI - Epstein-Barr virus (EBV)-encoded RNA promotes growth of EBV-infected T cells through interleukin-9 induction. AB - EBV associates with various T-cell-proliferating diseases such as chronic active EBV infection and nasal lymphoma. In contrast to B cells, which are highly susceptible to EBV infection in vitro, T cells are refractory to EBV infection in vitro, and it has been difficult to examine the effects of EBV infection on T cells. We recently generated EBV recombinants with a selectable marker, which made it possible to select EBV-infected cells even when the efficiency of infection was low. Using the recombinant virus, we found that a human T-cell line, MT-2, was susceptible to EBV infection, and we succeeded in isolating EBV infected cell clones with type II EBV latency, which was identical with those seen in EBV-infected T cells in vivo. EBV-infected MT-2 cell clones had shorter cell doubling times and higher saturation density than non-EBV-infected counterparts. We found that EBV-positive MT-2 cells expressed higher levels of interleukin (IL)-9 than EBV-negative MT-2 cells at the transcriptional level. It was also demonstrated that EBV-encoded small RNA was responsible for IL-9 expression. Addition of recombinant IL-9 accelerated the growth of MT-2 cells, whereas growth of the EBV-converted MT-2 cells was blocked by treatment with an anti-IL-9 antibody. These results suggest that IL-9 induced by EBV-encoded small RNA acts as an autocrine growth factor for EBV-infected T cells. Analysis of nasal lymphoma biopsies indicated that three of four specimens expressed IL-9. The present findings suggest that EBV directly affects the pathogenesis of EBV associated T-cell diseases. PMID- 15289340 TI - Silencing of the hypoxia-inducible cell death protein BNIP3 in pancreatic cancer. AB - Hypoxic conditions exist within pancreatic adenocarcinoma, yet pancreatic cancer cells survive and replicate within this environment. To understand the mechanisms involved in pancreatic cancer adaptation to hypoxia, we analyzed expression of a regulator of hypoxia-induced cell death, Bcl-2/adenovirus E1B 19 kDa interacting protein 3 (BNIP3). We found that BNIP3 was down-regulated in nine of nine pancreatic adenocarcinomas compared with normal pancreas despite the up regulation of other hypoxia-inducible genes, including glucose transporter-1 and insulin-like growth factor-binding protein 3. Also, BNIP3 expression was undetectable even after hypoxia treatment in six of seven pancreatic cancer cell lines. The BNIP3 promoter, which was remarkably activated by hypoxia, is located within a CpG island. The methylation status of CpG dinucleotides within the BNIP3 promoter was analyzed after bisulfite treatment by sequencing and methylation specific PCR. Hypermethylation of the BNIP3 promoter was observed in all BNIP3 negative pancreatic cancer cell lines and eight of 10 pancreatic adenocarcinoma samples. Treatment of BNIP3-negative pancreatic cancer cell lines with a DNA methylation inhibitor, 5-aza-2' deoxycytidine, restored hypoxia-induced BNIP3 expression. BNIP3 expression was also restored by introduction of a construct consisting of a full-length BNIP3 cDNA regulated by a cloned BNIP3 promoter. Restoration of BNIP3 expression rendered the pancreatic cancer cells notably more sensitive to hypoxia-induced cell death. In conclusion, down-regulation of BNIP3 by CpG methylation likely contributes to resistance to hypoxia-induced cell death in pancreatic cancer. PMID- 15289341 TI - Neural progenitor cell lines inhibit rat tumor growth in vivo. AB - Current therapies for gliomas often fail to address their infiltrative nature. Conventional treatments leave behind small clusters of neoplastic cells, resulting in eventual tumor recurrence. In the present study, we have evaluated the antitumor activity of neural progenitor cells against gliomas when stereotactically injected into nucleus Caudatus of Fisher rats. We show that the rat neural progenitor cell lines HiB5 and ST14A, from embryonic hippocampus and striatum primordium, respectively, are able to prolong animal survival and, in 25% of the cases, completely inhibit the outgrowth of N29 glioma compared with control animals. Delayed tumor outgrowth was also seen when HiB5 cells were inoculated at the site of tumor growth 1 week after tumor inoculation or when a mixture of tumor cells and HiB5 cells were injected s.c. into Fisher rats. HiB5 cells were additionally coinoculated together with two alternative rat gliomas, N32 and N25. N32 was growth inhibited, but rats inoculated with N25 cells did not show a prolonged survival. To evaluate the possibility of the involvement of the immune system in the tumor outgrowth inhibition, we show that HiB5 cells do not evoke an immune response when injected into Fisher rats. Furthermore, the rat neural progenitor cells produce all transforming growth factor beta isotypes, which could explain the observed immunosuppressive nature of these cells. Hence, some neural progenitor cells have the ability to inhibit tumor outgrowth when implanted into rats. These results indicate the usefulness of neural stem cells as therapeutically effective cells for the treatment of intracranial tumors. PMID- 15289342 TI - Dual-agent molecular targeting of the epidermal growth factor receptor (EGFR): combining anti-EGFR antibody with tyrosine kinase inhibitor. AB - Molecular inhibition of epidermal growth factor receptor (EGFR/HER1) signaling is under active investigation as a promising cancer treatment strategy. We examined the potency of EGFR inhibition achieved by combining anti-EGFR monoclonal antibody and tyrosine kinase inhibitor, which target extracellular and intracellular domains of the receptor, respectively. We specifically studied the combination of cetuximab (Erbitux, C225; ImClone Systems, New York, NY) with either gefitinib (Iressa, ZD1839; AstraZeneca, Macclesfield, UK) or erlotinib (Tarceva, OSI-774; Genentech, South San Francisco, CA) across a variety of human cancer cells. The combination of cetuximab plus gefitinib or erlotinib enhanced growth inhibition over that observed with either agent alone. As measured by immunostaining, inhibition of EGFR phosphorylation with the combination of cetuximab plus gefitinib or erlotinib was augmented over that obtained with single-agent therapy in head and neck (H&N) cancer cell lines. Phosphorylation inhibition of downstream effector molecules [mitogen-activated protein kinase (MAPK) and AKT] also was enhanced in tumor cells treated with the combination of cetuximab plus gefitinib or erlotinib. Flow cytometry and immunoblot analysis demonstrated that treatment of H&N tumor cells with cetuximab in combination with either gefitinib or erlotinib amplified the induction of apoptosis. Following establishment of cetuximab-resistant cell lines, we observed that gefitinib or erlotinib retained the capacity to inhibit growth of lung and H&N tumor cells that were highly resistant to cetuximab. Treatment with gefitinib or erlotinib, but not cetuximab, also could further inhibit the activation of downstream effectors of EGFR signaling in cetuximab-resistant cells, including MAPK and AKT. These data suggest that tyrosine kinase inhibitors may further modulate intracellular signaling that is not fully blocked by extracellular anti-EGFR antibody treatment. Finally, animal studies confirmed that single EGFR inhibitor treatment resulted in partial and transient tumor regression in human lung cancer xenografts. In contrast, more profound tumor regression and regrowth delay were observed in mice treated with the combination of cetuximab and gefitinib or erlotinib. Immunohistochemical staining, which demonstrated significant reduction of the proliferative marker proliferating cell nuclear antigen in mice treated with dual EGFR inhibitors, further supported this in vivo observation. Together, these data suggest that combined treatment with distinct EGFR inhibitory agents can augment the potency of EGFR signaling inhibition. This approach suggests potential new strategies to maximize effective target inhibition, which may improve the therapeutic ratio for anti-EGFR-targeted therapies in developing clinical trials. PMID- 15289343 TI - Phosphorylated histone H2AX in spheroids, tumors, and tissues of mice exposed to etoposide and 3-amino-1,2,4-benzotriazine-1,3-dioxide. AB - We reported recently that exposure of hamster V79 fibroblasts to 6 drugs that varied in their ability to produce DNA double-strand breaks stimulated formation of phosphorylated histone H2AX (serine 139 phosphorylated histone H2AX; gammaH2AX). Using flow cytometry to analyze gammaH2AX antibody-stained cells 1 h after a 30-min drug treatment, the fraction of cells that showed the control levels of gammaH2AX correlated well with the fraction of cells that survived to form colonies. This observation is now extended to V79 and SiHa human cervical carcinoma cells grown as multicell spheroids and SiHa xenografts and SCCVII tumors in mice. Animals were injected with etoposide, a topoisomerase-II inhibitor that targets proliferating cells or 3-amino-1,2,4-benzotriazine-1,3 dioxide (tirapazamine), a bioreductive cytotoxin that targets hypoxic cells. For spheroids, gammaH2AX intensity predicted clonogenic cell survival for cells recovered 90 min after drug injection, regardless of position of the cells within the spheroid. Similar results were obtained for etoposide in tumors; however, the gammaH2AX signal for tirapazamine was smaller than expected for the observed amount of cell killing. Frozen sections of tumors confirmed the greater intensity of gammaH2AX staining in cells close to blood vessels of tumors soon after treatment with etoposide and the opposite pattern for tumors exposed to tirapazamine. Analysis of cells or frozen sections from mouse spleen and kidney suggests that information can also be obtained on initial damage in normal tissues. These results support the possibility of using gammaH2AX antibody staining as a method to aid in prediction of tumor and normal tissue response to treatment. PMID- 15289344 TI - A synthetic 15-mer peptide (PCK3145) derived from prostate secretory protein can reduce tumor growth, experimental skeletal metastases, and malignancy-associated hypercalcemia. AB - In previous studies, we have shown that prostate secretory protein (PSP-94) can reduce prostate cancer growth in vivo. In the current study, we identified the amino acid sequence of PSP-94 that is required for eliciting this response. For these studies, we used rat prostate cancer Mat Ly Lu cells overexpressing parathyroid hormone-related protein (PTHrP), which is the main pathogenetic factor responsible for hypercalcemia of malignancy. Synthetic peptides corresponding to amino acids 7-21 (PCK721), 31-45 (PCK3145), and 76-94 (PCK7694) of PSP-94 were synthesized. Only PCK3145 showed a significant reduction in tumor cell proliferation. For in vivo studies, syngenic male Copenhagen rats were inoculated s.c. with Mat Ly Lu cells overexpressing PTHrP into the right flank or into the left ventricle via intracardiac injection, which results in experimental metastases to the lumbar vertebrae causing hind-limb paralysis. Animals were infused with different doses (1, 10, and 100 microg/kg/day) of peptides for 15 days, and the effect of these treatments on tumor volume, skeletal metastases, or development of hind-limb paralysis was determined. Treatment with PCK3145 resulted in a dose-dependent decrease in tumor volume and delay in the development of skeletal metastases. Bone histomorphometry showed that after intracardiac inoculation of tumor cells, the highest dose of PCK3145 (100 microg/kg/day) resulted in reducing skeletal tumor burden, which delayed the development of hind-limb paralysis. Treatment with PCK3145 led to reduction of plasma calcium and PTHrP levels and a significant decrease in PTHrP levels in the primary tumors and in vertebrae of experimental animals. These effects of PCK3145 were due to its ability to promote tumor cell apoptosis. Collectively, the results of these studies have demonstrated the ability of a small peptide derived from PSP-94 to reduce tumor volume and experimental skeletal metastases-results that will be highly beneficial in the continued development of this peptide as a novel therapeutic agent for patients with hormone refractory, late-stage prostate cancer. PMID- 15289345 TI - Colony-stimulating factor-1 blockade by antisense oligonucleotides and small interfering RNAs suppresses growth of human mammary tumor xenografts in mice. AB - Colony-stimulating factor (CSF)-1 is the primary regulator of tissue macrophage production. CSF-1 expression is correlated with poor prognosis in breast cancer and is believed to enhance mammary tumor progression and metastasis through the recruitment and regulation of tumor-associated macrophages. Macrophages produce matrix metalloproteases (MMPs) and vascular endothelial growth factor, which are crucial for tumor invasion and angiogenesis. Given the important role of CSF-1, we hypothesized that blockade of CSF-1 or the CSF-1 receptor (the product of the c-fms proto-oncogene) would suppress macrophage infiltration and mammary tumor growth. Human MCF-7 mammary carcinoma cell xenografts in mice were treated with either mouse CSF-1 antisense oligonucleotide for 2 weeks or five intratumoral injections of either CSF-1 small interfering RNAs or c-fms small interfering RNAs. These treatments suppressed mammary tumor growth by 50%, 45%, and 40%, respectively, and selectively down-regulated target protein expression in tumor lysates. Host macrophage infiltration; host MMP-12, MMP-2, and vascular endothelial growth factor A expression; and endothelial cell proliferation within tumors of treated mice were decreased compared with tumors in control mice. In addition, mouse survival significantly increased after CSF-1 blockade. These studies demonstrate that CSF-1 and CSF-1 receptor are potential therapeutic targets for the treatment of mammary cancer. PMID- 15289346 TI - An anti-Wnt-2 monoclonal antibody induces apoptosis in malignant melanoma cells and inhibits tumor growth. AB - Activation of the Wnt/beta-catenin signaling pathway has been associated with human cancers. To test whether Wnt-2 signal is a survival factor in human melanoma cells and thus represents a potential therapeutic target, we investigated the effects of inhibition of Wnt-2 signaling in human melanoma cell lines. We have developed a novel monoclonal antibody against the NH(2) terminus of the human Wnt-2 ligand that induces apoptosis in human melanoma cells overexpressing Wnt-2. Whereas incubation of this antibody with normal cells lacking Wnt-2 expression does not induce apoptosis, Wnt-2 signaling blockade by the ligand-binding antibody is confirmed by down-regulation of Dishevelled and beta-catenin. Wnt-2 small interfering RNA treatment in these cells yielded similar apoptotic effects and downstream changes. Down-regulation of an inhibitor of apoptosis family protein, survivin, was observed in both the Wnt-2 antibody treated and small interfering RNA-treated melanoma cell lines, suggesting that the antibody induces apoptosis by inactivating survivin. In an in vivo study, this monoclonal anti-Wnt-2 antibody suppresses tumor growth in a xenograft model. These findings suggest that the anti-Wnt-2 monoclonal antibody may be useful for the treatment of patients with malignant melanoma. PMID- 15289347 TI - Effective gene-viral therapy for telomerase-positive cancers by selective replicative-competent adenovirus combining with endostatin gene. AB - Gene-viral therapy, which uses replication-selective transgene-expressing viruses to manage tumors, can exploit the virtues of gene therapy and virotherapy and overcome the limitations of conventional gene therapy. Using a human telomerase reverse transcriptase-targeted replicative adenovirus as an antiangiogenic gene transfer vector to target new angiogenesis and making use of its unrestrained proliferation are completely new concepts in tumor management. CNHK300-mE is a selective replication transgene-expressing adenovirus constructed to carry mouse endostatin gene therapeutically. Infection with CNHK300-mE was associated with selective replication of the adenovirus and production of mouse endostatin in telomerase-positive cancer cells. Endostatin secreted from a human gastric cell line, SGC-7901, infected with CNHK300-mE was significantly higher than that infected with nonreplicative adenovirus Ad-mE in vitro (800 +/- 94.7 ng/ml versus 132.9 +/- 9.9 ng/ml) and in vivo (610 +/- 42 ng/ml versus 126 +/- 13 ng/ml). Embryonic chorioallantoic membrane assay showed that the mouse endostatin secreted by CNHK300-mE inhibited angiogenesis efficiently and also induced distortion of pre-existing vasculature. CNHK300-mE exhibited a superior suppression of xenografts in nude mice compared with CNHK300 and Ad-mE. In summary, we provided a more efficient gene-viral therapy strategy by combining oncolysis with antiangiogenesis. PMID- 15289348 TI - Differential activation of the phosphatidylinositol 3'-kinase/Akt survival pathway by ionizing radiation in tumor and primary endothelial cells. AB - Ionizing radiation induces an intracellular stress response via activation of the phosphatidylinositol 3'-kinase (PI3K)/Akt survival pathway. In tumor cells, the PI3K/Akt pathway is induced through activation of members of ErbB receptor tyrosine kinases. Here, we investigated the receptor dependence of radiation induced PI3K/Akt activation in tumor cells and in endothelial cells. The integrity of both the ErbB and the vascular endothelial growth factor (VEGF) ligand-activated PI3K/Akt pathway in endothelial cells was demonstrated using specific ErbB and VEGF receptor tyrosine kinase inhibitors. Irradiation of endothelial cells resulted in protein kinase B (PKB)/Akt activation in a similar time course as observed in response to VEGF. More importantly, radiation-induced PKB/Akt phosphorylation in endothelial cells was strongly down-regulated by the VEGF receptor tyrosine kinase inhibitor, whereas the ErbB receptor tyrosine kinase inhibitor did not affect PKB/Akt stimulation in response to irradiation. An opposite receptor dependence for radiation-induced PKB/Akt phosphorylation was observed in ErbB receptor-overexpressing A431 tumor cells. Furthermore, direct VEGF receptor phosphorylation was detected after irradiation in endothelial cells in absence of VEGF, which was almost completely inhibited after irradiation in presence of the VEGF receptor tyrosine kinase inhibitor. These data demonstrate that ionizing radiation induces VEGF ligand-independent but VEGF receptor dependent PKB/Akt activation in endothelial cells and that PI3K/Akt pathway activation by radiation occurs in a differential cell type and receptor-dependent pattern. PMID- 15289349 TI - Two distinct pathways of immuno-modulation improve potency of p53 immunization in rejecting established tumors. AB - The p53 gene product is overexpressed by almost 50% of cancers, making it an ideal target for cancer immunotherapy. We previously demonstrated rejection of established p53-overexpressing tumors without stimulating autoimmunity by immunization with modified vaccinia Ankara-expressing murine p53 (MVAp53). Tumor rejection was enhanced through antibody-mediated CTL-associated antigen 4 (CTLA 4) blockade. We examined the role of synthetic oligodeoxynucleotides (ODN) containing unmethylated cytosine-phosphate-guanine (CpG) motifs (CpG ODN) in enhancing MVAp53-mediated tumor rejection. CpG ODN with MVAp53 resulted in tumor rejection in BALB/c mice bearing poorly immunogenic 11A-1 murine mammary carcinomas or Meth A sarcomas and C57Bl/6 mice bearing MC-38 colon carcinomas. The effect was similar to that seen in tumor-bearing mice treated with MVAp53 along with CTLA-4 blockade. Monoclonal antibody depletion experiments demonstrated that the adjuvant effects of CpG ODN and CTLA-4 blockades were CD8 dependent. CpG ODN were partially natural killer cell dependent and ineffective in Toll-like Receptor 9(-/-) and interleukin 6(-/-) mice, whereas CTLA-4 blockade was partially CD4 dependent and functional in Toll-like Receptor 9(-/-) and interleukin 6(-/-) mice. In addition, when administered with MVAp53, both adjuvants enhanced p53-specific cytotoxicity and demonstrated an additive effect when combined. The combination of CpG ODN and CTLA-4 blockade worked synergistically to reject palpable 11A-1 and MC-38 tumors. These experiments demonstrate the potential for augmenting MVAp53-mediated antitumor immunity using CpG ODN and CTLA-4 blockade. This cell-free immunotherapy approach is a candidate for evaluation in cancer patients. PMID- 15289350 TI - Fenretinide cytotoxicity for Ewing's sarcoma and primitive neuroectodermal tumor cell lines is decreased by hypoxia and synergistically enhanced by ceramide modulators. AB - Patients with disseminated Ewing's family of tumors (ESFT) often experience drug resistant relapse. We hypothesize that targeting minimal residual disease with the cytotoxic retinoid N-(4-hydroxyphenyl) retinamide (4-HPR; fenretinide) may decrease relapse. We determined the following: (a) 4-HPR cytotoxicity against 12 ESFT cell lines in vitro; (b) whether 4-HPR increased ceramide species (saturated and desaturated ceramides); (c) whether physiological hypoxia (2% O(2)) affected cytotoxicity, mitochondrial membrane potential (DeltaPsi(m)) change, or ceramide species or reactive oxygen species levels; (d) whether cytotoxicity was enhanced by l-threo-dihydrosphingosine (safingol); (e) whether physiological hypoxia increased acid ceramidase (AC) expression; and (f) the effect of the AC inhibitor N-oleoyl-ethanolamine (NOE) on cytotoxicity and ceramide species. Ceramide species were quantified by thin-layer chromatography and scintillography. Cytotoxicity was measured by a fluorescence-based assay using digital imaging microscopy (DIMSCAN). Gene expression profiling was performed by oligonucleotide array analysis. We observed, in 12 cell lines tested in normoxia (20% O(2)), that the mean 4-HPR LC(99) (the drug concentration lethal to 99% of cells) = 6.1 +/- 5.4 microm (range, 1.7-21.8 microm); safingol (1-3 microm) synergistically increased 4-HPR cytotoxicity and reduced the mean 4-HPR LC(99) to 3.2 +/- 1.7 microm (range, 2.0-8.0 microm; combination index < 1). 4-HPR increased ceramide species in the three cell lines tested (up to 9-fold; P < 0.05). Hypoxia (2% O(2)) reduced ceramide species increase, DeltaPsi(m) loss, reactive oxygen species increase (P < 0.05), and 4-HPR cytotoxicity (P = 0.05; 4-HPR LC(99), 19.7 +/- 23.9 microm; range, 2.3-91.4). However, hypoxia affected 4-HPR + safingol cytotoxicity to a lesser extent (P = 0.04; 4-HPR LC(99), 4.9 +/- 2.3 microm; range, 2.0-8.2). Hypoxia increased AC RNA expression; the AC inhibitor NOE enhanced 4-HPR-induced ceramide species increase and cytotoxicity. The antioxidant N-acetyl-l-cysteine somewhat reduced 4-HPR cytotoxicity but did not affect ceramide species increase. We conclude the following: (a) 4-HPR was active against ESFT cell lines in vitro at concentrations achievable clinically, but activity was decreased in hypoxia; and (b) combining 4-HPR with ceramide modulators synergized 4-HPR cytotoxicity in normoxia and hypoxia. PMID- 15289351 TI - Up-regulation of Egr1 by 1,25-dihydroxyvitamin D3 contributes to increased expression of p35 activator of cyclin-dependent kinase 5 and consequent onset of the terminal phase of HL60 cell differentiation. AB - Advances in differentiation therapy of cancer are likely to depend on improved understanding of molecular events that underlie cell differentiation. We reported recently that cyclin-dependent kinase (Cdk)5 and p35Nck5a (p35) are expressed in human leukemia HL60 cells induced to differentiate to monocytes by an exposure to 1,25-dihydroxyvitamin D(3) (1,25D(3)), form a complex, and this complex has kinase activity (F. Chen and G. P. Studzinski, Blood 2001;97:3763). This laboratory has also provided evidence that the extracellular signal-regulated kinase/mitogen-activated protein kinase pathway is active in the early (24-48 h) stages of HL60 cell differentiation induced by 1,25D(3) but declines in the later, terminal phase of this form of differentiation (X. Wang and G. P. Studzinski, J Cell Biochem 2001;80:471). We examine now the hypothesis that Egr1 protein contributes to the up-regulation of p35 gene transcription and, thus, activated Cdk5/p35 kinase phosphorylates and inactivates mitogen-activated protein/extracellular signal-regulated kinase kinase 1 (MEK1). Our data show that in 1,25D(3)-treated cells, p35 and Egr1 protein levels are elevated in a dose dependent manner at the onset of the late stage of differentiation. We show also that 1,25D(3) treatment of HL60 cells markedly increases the binding of Egr1 to an element in the p35 gene promoter, whereas transfection of an excess of this Egr1-binding oligonucleotide ("promoter decoy") reduces p35 gene transcription and cell differentiation. Additionally, Cdk5/p35 phosphorylates MEK1 and inhibits its ability to phosphorylate its downstream target Erk2. These data suggest that in 1,25D(3)-treated HL60 cells, Egr1 up-regulates p35 gene transcription and that Cdk5/p35 kinase inactivates the extracellular signal-regulated kinase/mitogen activated protein kinase pathway by phosphorylation of MEK1, and this contributes to terminal differentiation of these cells. PMID- 15289352 TI - The human cervical cancer oncogene protein is a biomarker for human hepatocellular carcinoma. AB - Human cervical cancer oncogene (HCCR) was identified and appeared to function as a negative regulator of p53 gene. The objective of this study was to validate HCCR expression as a candidate marker for human hepatocellular carcinoma. HCCR epitope was identified as Y(355)LGTRR(360). According to immunofluorescence study, HCCR was predominantly localized in the plasma membrane and cytoplasm of hepatocellular carcinoma. HCCR proteins were overexpressed in the tumorous compared with the nontumorous cirrhosis tissues. However, HCCR was not detected in normal liver tissue. Concentration of HCCR protein in the serum was measured in a total of 570 subjects, and comparisons were made to alpha-fetoprotein. Serological studies revealed 78.2% sensitivity of HCCR (cutoff value, 15 microg/ml), which was significantly higher than 64.6% of alpha-fetoprotein (P = 0.0098) and 95.7% specificity for hepatocellular carcinoma. Forty of 52 (76.9%) patients with carcinoma negative for alpha-fetoprotein showed positive values for HCCR. A positive rate of 69.2% in carcinoma patients with tumor sizes <2 cm was found to be a higher rate than measurement of alpha-fetoprotein. Furthermore, HCCR expression was also detected in liver cirrhosis at an intermediate level between carcinoma and normal groups, which gave 88.1% sensitivity and 79.0% specificity using 8 microg/ml as a cutoff value. In summary, the HCCR assay may have an advantage over the alpha-fetoprotein assay in that it is elevated according to disease progression from liver cirrhosis to carcinoma, and it is more frequently positive in patients with early, small hepatocellular carcinoma. PMID- 15289353 TI - Identification of human autologous cytotoxic T-lymphocyte-defined osteosarcoma gene that encodes a transcriptional regulator, papillomavirus binding factor. AB - The prognosis for patients with osteosarcoma who do not respond to current chemotherapy protocols still remains poor. Toward the goal of establishing efficacious peptide-based immunotherapy for those patients, we previously developed an autologous pair of CTLs and an osteosarcoma cell line. In the current study, we screened the cDNA library of this osteosarcoma cell line using an autologous CTL clone and identified cDNA encoding an antigen. The isolated cDNA was identical to papillomavirus binding factor (PBF), which was recently reported as a DNA binding transcription factor cooperating with RUNX1. Reverse transcription-PCR analysis revealed that PBF was expressed in 16 of 19 cases of bone and soft-tissue sarcoma cell lines (5 of 6 of osteosarcoma lines) and 57 of 76 sarcoma tissue samples (11 of 14 of osteosarcoma tissues). Also, PBF was expressed in 10 of 13 epithelial cancer cell lines and 20 of 34 of cancer tissues. In contrast, PBF was detected in some normal organs including ovary, pancreas, spleen, and liver by reverse transcription-PCR but was restricted in the cytoplasm by immunostaining and undetectable by Western blotting. Furthermore, a 12-mer peptide, CTACRWKKACQR, located at the COOH terminus of PBF, was found to be a minimum requirement for recognition by the CTL clone in the context of the HLA-B*5502 molecule. These findings suggest that PBF is a shared tumor-associated antigen, which may serve as a source of peptides applicable to peptide-based immunotherapy for osteosarcoma and other malignant tumors. PMID- 15289354 TI - Human papillomavirus type 16-positive cervical cancer is associated with impaired CD4+ T-cell immunity against early antigens E2 and E6. AB - Cervical cancer is the possible outcome of genital infection with high-risk human papillomavirus (HPV) and is preceded by a phase of persistent HPV infection during which the host immune system fails to eliminate the virus. Fortunately, the majority of genital HPV infections are cleared before the development of (pre)malignant lesions. Analysis of CD4+ T-helper (Th) immunity against the E2, E6, and E7 antigens of HPV16 in healthy women revealed strong proliferative E2- and E6-specific responses associated with prominent IFN-gamma and interleukin 5 secretion. This indicates that the naturally arising virus-induced immune response displays a mixed Th1/Th2 cytokine profile. Of all HPV16+ cervical cancer patients, approximately half failed to mount a detectable immune response against the HPV16-derived peptides. The other half of the patients showed impaired HPV16 specific proliferative responses, which generally lacked both IFN-gamma and interleukin 5. This indicates that the HPV16-specific CD4+ T-cell response in cervical cancer patients is either absent or severely impaired, despite a relatively good immune status of the patients, as indicated by intact responses against recall antigens. It is highly conceivable that proper CD4+ T-cell help is important for launching an effective immune attack against HPV because infection of cervical epithelia by this virus is, at least initially, not accompanied by gross disturbance of this tissue and/or strong proinflammatory stimuli. Therefore, our observations concerning the lack of functional HPV16-specific CD4+ T-cell immunity in patients with cervical cancer offer a possible explanation for the development of this disease. PMID- 15289355 TI - Immunogenicity of constitutively active V599EBRaf. AB - Activating BRAF somatic missense mutations within the kinase domain are present in 60-66% of melanomas. The vast majority of these represent a single substitution of glutamate for valine (V599E). Here, we demonstrate spontaneous HLA-B*2705-restricted cytotoxic T-cell responses against an epitope derived from (V599E)BRaf. These T-cell responses were mutation specific as the corresponding epitope derived from wild-type BRaf was not recognized. The loss of the (V599E)BRAF genotype during progression from primary to metastatic melanoma in patients with (V599E)BRaf specific T-cell responses suggests an active immune selection of nonmutated melanoma clones by the tumor-bearing host. PMID- 15289356 TI - Expression of toll-like receptor 4 on dendritic cells is significant for anticancer effect of dendritic cell-based immunotherapy in combination with an active component of OK-432, a streptococcal preparation. AB - A lipoteichoic acid-related molecule OK-PSA is an active component of OK-432, a Streptococcus-derived anticancer immunotherapeutic agent. In the present study, we first examined the effect of OK-PSA on the maturation of dendritic cells (DCs) in vitro by using the DCs derived from 5 healthy donors and 10 patients with head and neck cancer with or without expression of toll-like receptor 4 (TLR4) or MD-2 mRNA. OK-PSA treatment effectively increased the surface expression of MHC class II, CD80, CD83, and CD86. OK-PSA-stimulated DCs secreted the cytokines that can induce helper T-cell 1 (Th1)-type T-cell response, and stimulated allogeneic T cells to produce IFN-gamma and to elicit an allogeneic antigen-specific cytotoxicity. These activities almost depended on expression of TLR4 and MD-2 genes. We next investigated the in vivo anticancer effect of intratumoral administration of syngeneic DCs followed by OK-PSA against established tumors in mice. C57BL/6 mice, which express wild-type TLR4, and C57BL/6-derived TLR4 knockout (TLR4(-/-)) mice were used. Although OK-PSA accelerated the antitumor effect of intratumoral DC administration in wild-type mice bearing syngeneic tumors, the antitumor effect of OK-PSA as well as of the combination therapy with DCs and OK-PSA was not significant in TLR4(-/-) mice. Interestingly, an administration of wild-type-mouse-derived DCs followed by OK-PSA exhibited a marked antitumor effect even in the TLR4(-/-) mice. These findings suggest that OK-PSA may be a potent adjuvant for local DC therapy, and that DC therapy followed by OK-PSA is able to elicit anticancer activity even in a TLR4-deficient host when TLR4 is expressed only in DCs injected intratumorally. PMID- 15289357 TI - PT-100, a small molecule dipeptidyl peptidase inhibitor, has potent antitumor effects and augments antibody-mediated cytotoxicity via a novel immune mechanism. AB - The amino boronic dipeptide, PT-100 (Val-boro-Pro), a dipeptidyl peptidase (DPP) inhibitor, has been shown to up-regulate gene expression of certain cytokines in hematopoietic tissue via a high-affinity interaction, which appears to involve fibroblast activation protein. Because fibroblast activation protein is also expressed in stroma of lymphoid tissue and tumors, the effect of PT-100 on tumor growth was studied in mice in vivo. PT-100 has no direct cytotoxic effect on tumors in vitro. Oral administration of PT-100 to mice slowed growth of syngeneic tumors derived from fibrosarcoma, lymphoma, melanoma, and mastocytoma cell lines. In WEHI 164 fibrosarcoma and EL4 and A20/2J lymphoma models, PT-100 caused regression and rejection of tumors. The antitumor effect appeared to involve tumor-specific CTL and protective immunological memory. PT-100 treatment of WEHI 164-inoculated mice increased mRNA expression of cytokines and chemokines known to promote T-cell priming and chemoattraction of T cells and innate effector cells. The role of innate activity was further implicated by observation of significant, although reduced, inhibition of WEHI 164 and A20/2J tumors in immunodeficient mice. PT-100 also demonstrated ability to augment antitumor activity of rituximab and trastuzumab in xenograft models of human CD20(+) B-cell lymphoma and HER-2(+) colon carcinoma where antibody-dependent cytotoxicity can be mediated by innate effector cells responsive to the cytokines and chemokines up-regulated by PT-100. Although CD26/DPP-IV is a potential target for PT-100 in the immune system, it appeared not to be involved because antitumor activity and stimulation of cytokine and chemokine production was undiminished in CD26(-/-) mice. PMID- 15289358 TI - Fine specificity of high molecular weight-melanoma-associated antigen-specific cytotoxic T lymphocytes elicited by anti-idiotypic monoclonal antibodies in patients with melanoma. AB - HLA-A2-restricted CTLs, which lysed high molecular weight (HMW)-melanoma associated antigen (MAA)(+) melanoma cells, were induced in patients with melanoma immunized with MELIMMUNE, a combination of the murine anti-idiotypic (anti-id) monoclonal antibodies (mAb) MEL-2 and MF11-30 (MW Pride et al., Clin Cancer Res 1998;4:2363.). In the present study we investigated whether CTL epitopes are present in anti-id mAb MF-11-30 and activate T cells to recognize HMW-MAA on melanoma cells. One candidate epitope in the mAb MF11-30 VH chain, VH (3-11), was selected based on the presence of HLA-A2 anchor residues and partial homology with the HMW-MAA epitope, HMW-MAA (76-84). Lymphocytes from HLA-A2(+) immunized patients proliferated to VH (3-11) peptide and to a variant HMW-MAA peptide to a significantly greater extent than autologous lymphocytes stimulated with an irrelevant peptide and lymphocytes from nonimmunized patients. No proliferative response was detected to the wild-type HMW-MAA peptide (76-84). Significant increase in IFN-gamma production but not in interleukin 10 production in response to VH (3-11) and to variant HMW-MAA peptide (76-84) was observed in lymphocytes from the immunized patients. Stimulation of lymphocytes from HLA A2(+) patients with the two peptides induced CTL, which lysed HMW-MAA(+)/HLA A2(+) A375SM melanoma cells. This is the first report documenting the presence of immunogenic peptides in a murine anti-id mAb for a defined epitope expressed by a human melanoma-associated antigen. These results may be relevant for development of novel vaccines based on homology between anti-id mAb and tumor-associated antigen amino acid sequences. PMID- 15289359 TI - NE-10 neuroendocrine cancer promotes the LNCaP xenograft growth in castrated mice. AB - Increases in neuroendocrine (NE) cells and their secretory products are closely correlated with tumor progression and androgen-independent prostate cancer. However, the mechanisms by which NE cells influence prostate cancer growth and progression, especially after androgen ablation therapy, are poorly understood. To investigate the role of NE cells on prostate cancer growth, LNCaP xenograft tumors were implanted into nude mice. After the LNCaP tumors were established, the NE mouse prostate allograft (NE-10) was implanted on the opposite flank of these nude mice to test whether NE tumor-derived systemic factors can influence LNCaP growth. Mice bearing LNCaP tumors with or without NE allografts were castrated 2 weeks after NE tumor inoculation, and changes in LNCaP tumor growth rate and gene expression were investigated. After castration, LNCaP tumor growth decreased in mice bearing LNCaP tumors alone, and this was accompanied by a loss of nuclear androgen receptor (AR) localization. In contrast, in castrated mice bearing both LNCaP and NE-10 tumors, LNCaP tumors continued to grow, had increased levels of nuclear AR, and secreted prostate-specific antigen. Therefore, in the absence of testicular androgens, NE secretions were sufficient to maintain LNCaP cell growth and androgen-regulated gene expression in vivo. Furthermore, in vitro experiments showed that NE secretions combined with low levels of androgens activated the AR, an effect that was blocked by the antiandrogen bicalutamide. Because an increase in AR level has been reported to be sufficient to account for hormone refractory prostate cancers, the NE cell population ability to increase AR level/activity can be another mechanism that allows prostate cancer to escape androgen ablation therapy. PMID- 15289360 TI - The potential role of hypoxia inducible factor 1alpha in tumor progression after hypoxia and chemotherapy in hepatocellular carcinoma. AB - This study investigates the possible molecular basis leading to failure in a treatment that is composed of hypoxia and chemotherapy in a rat orthotopic hepatoma model. Hypoxia was induced by hepatic artery ligation, whereas chemotherapeutic effect was achieved by intraportal injection of cisplatin. High dose sodium salicylate was administered to achieve transcriptional blockade. Significant prolongation of animal survival was observed in the groups receiving hepatic artery ligation with cisplatin or sodium salicylate. Massive tumor cell necrosis and apoptosis were found in the ligation and all of the combined treatment groups. Up-regulation of hypoxia inducible factor 1alpha (HIF-1alpha) and vascular endothelial growth factor (VEGF) at both mRNA and protein levels were detected in the groups receiving ligation and ligation with cisplatin, whereas a decreased level of von Hippel-Lindau tumor suppressor protein was identified in the group receiving ligation with cisplatin. Sodium salicylate enhanced expression of von Hippel-Lindau tumor suppressor protein but down regulated HIF-1alpha and VEGF levels after ligation with or without cisplatin. An increased number of activated hepatic stellate cells in the tumors were observed in the ligation and ligation with cisplatin groups, whereas they were greatly reduced by sodium salicylate. In vitro study revealed that under hypoxic condition, both cisplatin and sodium salicylate could remarkably augment P53 and caspase 3 levels. Cisplatin stimulated HIF-1alpha up-regulation, whereas sodium salicylate suppressed HIF-1alpha expression. In conclusion, tumor progression after hypoxia and chemotherapy might be related to up-regulation of HIF-1alpha and subsequent VEGF production, and transcriptional blockade by sodium salicylate could enhance the therapeutic efficacy of hypoxia and chemotherapy. PMID- 15289361 TI - An autoantibody-mediated immune response to calreticulin isoforms in pancreatic cancer. AB - The identification of circulating tumor antigens or their related autoantibodies provides a means for early cancer diagnosis as well as leads for therapy. We have used a proteomic approach to identify proteins that commonly induce a humoral response in pancreatic cancer. Aliquots of solubilized proteins from a pancreatic cancer cell line (Panc-1) were subjected to two-dimensional PAGE, followed by Western blot analysis in which sera of individual patients were tested for primary antibodies. Sera from 36 newly diagnosed patients with pancreatic cancer, 18 patients with chronic pancreatitis, 33 patients with other cancers, and 15 healthy subjects were analyzed. Autoantibodies were detected against either one or two calreticulin isoforms identified by mass spectrometry in sera from 21 of 36 patients with pancreatic cancer. One of 18 chronic pancreatitis patients and 1 of 15 healthy controls demonstrated autoantibodies to calreticulin isoform 1; none demonstrated autoantibodies to isoform 2. None of the sera from patients with colon cancer exhibited reactivity against either of these two proteins. One of 14 sera from lung adenocarcinoma patients demonstrated autoantibodies to calreticulin isoform 1; 2 of 14 demonstrated autoantibodies to isoform 2. Immunohistochemical analysis of calreticulin in pancreatic/ampullary tumor tissue arrays using an isoform nonspecific antibody revealed diffuse and consistent cytoplasmic staining in the neoplastic epithelial cells of the pancreatic and ampullary adenocarcinomas. The detection of autoantibodies to calreticulin isoforms may have utility for the early diagnosis of pancreatic cancer. PMID- 15289362 TI - Quantitative detection of promoter hypermethylation of multiple genes in the tumor, urine, and serum DNA of patients with renal cancer. AB - Aberrant promoter hypermethylation of several known or putative tumor suppressor genes occurs frequently during the pathogenesis of human cancers and is a promising marker for cancer detection. We investigated the feasibility of detecting aberrant DNA methylation in the urine and serum samples of renal cancer patients. We examined the tumor and the matched urine and serum DNA for aberrant methylation of nine gene promoters (CDH1, APC, MGMT, RASSF1A, GSTP1, p16, RAR beta2, and ARF) from 17 patients with primary kidney cancer by quantitative fluorogenic real-time PCR. An additional 9 urine samples (total, 26) and 1 serum sample (total, 18) also were tested from renal cancer patients. Urine from 91 patients without genitourinary cancer and serum from 30 age-matched noncancer individuals were used as controls. Promoter hypermethylation of at least two of the genes studied was detected in 16 (94%) of 17 primary tumors. Aberrant methylation in urine and serum DNA generally was accompanied by methylation in the matched tumor samples. Urine samples from 91 control subjects without evidence of genitourinary cancer revealed no methylation of the MGMT, GSTP1, p16, and ARF genes, whereas methylation of RAR-beta2, RASSF1A, CDH1, APC, and TIMP3 was detected at low levels in a few control subjects. Overall, 23 (88%) of 26 urine samples and 12 (67%) of 18 serum samples from cancer patients were methylation positive for at least one of the genes tested. By combination of urine or serum analysis of renal cancer patients, hypermethylation was detected in 16 of 17 patients (94% sensitivity) with high specificity. Our findings suggest that promoter hypermethylation in urine or serum can be detected in the majority of renal cancer patients. This noninvasive high-throughput approach needs to be evaluated in large studies to assess its value in the early detection and surveillance of renal cancer. PMID- 15289363 TI - Modern criteria to establish human cancer etiology. AB - The Cancer Etiology Branch of the National Cancer Institute hosted a workshop, "Validation of a causal relationship: criteria to establish etiology," to determine whether recent technological advances now make it possible to delineate improved or novel criteria for the rapid establishment for cancer causation. The workshop was held in Washington, D.C., December 11-12, 2003, and participants were among the international leaders in the fields of epidemiology, chemistry, biochemistry, microbiology, virology, environmental and chemical carcinogenesis, immunology, pathology, molecular pathology, genetics, oncology, and surgical oncology. There was a general consensus that the rapid identification of human carcinogens and their removal (when possible) or the establishment of specific preventive and therapeutic measures was the most desirable and effective way to have a rapid and positive impact in the fight against cancer. From a clinical perspective, it may be as important to target initiators, cocarcinogens and promoters, if by removing any one of them tumor growth can be prevented. Future studies should focus on interactions among and between different biological, chemical, and physical agents. Analyses of single agents can at times miss their carcinogenic potential when such agents are carcinogenic only in subgroups of individuals because of their genetic background, diet, exposure to other carcinogens, or microbial infection. Epidemiology, molecular pathology (including chemistry, biochemistry, molecular biology, molecular virology, molecular genetics, epigenetics, genomics, proteomics, and other molecular-based approaches), and animal and tissue culture experiments should all be seen as important integrating evidence in the determination of human carcinogenicity. Concerning the respective roles of epidemiology and molecular pathology, it was noted that epidemiology allows the determination of the overall effect of a given carcinogen in the human population (e.g., hepatitis B virus and hepatocellular carcinoma) but cannot prove causality in the individual tumor patient. Molecular pathology cannot determine the overall impact of a carcinogen in the population but can at times prove causality in the individual tumor patient [such as the detection of high-risk human papillomavirus (HPV) in a cervical carcinoma biopsy]. This is possible when molecular techniques have shown that the agent is required for transformation or malignant growth of human cells (such as antisense HPV strategies showing the requirement for the expression of HPV proteins for tumor cell growth) and when there is supportive experimental animal evidence. Ideally, epidemiology and molecular pathology information together with experimental evidence in animals should be available for the most reliable identification of human carcinogens. All sets of data are not always available, and a rapid identification of human carcinogens is in the best public health interest. Swift validation of a causal relationship when followed by a rapid deployment of preventive and therapeutic approaches should lead to a favorable public health impact (such as hepatitis B virus vaccination to prevent hepatocellular carcinoma). PMID- 15289364 TI - NOD2 and colorectal cancer: guilt by non-association. PMID- 15289365 TI - Electrocardiographic predictors of arrhythmic death and total mortality in the multicenter unsustained tachycardia trial. AB - BACKGROUND: Stratifiers of sudden and total mortality risk are needed to optimally target preventive therapies in patients with coronary artery disease and impaired ventricular function. We assessed the prognostic significance of ECG markers of conduction abnormalities and left ventricular hypertrophy in the Multicenter Unsustained Tachycardia Trial (MUSTT). METHODS AND RESULTS: We analyzed the ECGs of 1638 patients from MUSTT who did not receive antiarrhythmic therapy (antiarrhythmic medication or implantable cardioverter-defibrillator). After adjustment for other significant factors, left bundle-branch block and intraventricular conduction delay were associated with a 50% increase in the risk of both arrhythmic and total mortality. Right bundle-branch block was not associated with arrhythmic or total mortality. Left ventricular hypertrophy was the only ECG predictor of arrhythmic (hazard ratio, 1.35; 95% CI, 1.08 to 1.69) but not total mortality. CONCLUSIONS: In patients with coronary artery disease, depressed left ventricular function, and nonsustained ventricular tachycardia, QRS prolongation resulting from left bundle-branch block or intraventricular conduction delay but not right bundle-branch block provided prognostic information about the risk of arrhythmic and total mortality independently of electrophysiological evaluation and ejection fraction. Left ventricular hypertrophy was associated with increased arrhythmic but not total mortality. PMID- 15289366 TI - Mineralocorticoid receptor antagonism prevents the electrical remodeling that precedes cellular hypertrophy after myocardial infarction. AB - BACKGROUND: Cardiac hypertrophy underlies arrhythmias and sudden death, for which mineralocorticoid receptor (MR) activity has recently been implicated. We sought to establish the sequence of ionic events that link the initiating insult and MR to hypertrophy development. METHODS AND RESULTS: Using whole-cell, patch-clamp and quantitative reverse transcription-polymerase chain reaction techniques on right ventricular myocytes of a myocardial infarction (MI) rat model, we examined the cellular response over time. One week after MI, no sign of cellular hypertrophy was found, but action potential duration (APD) was lengthened. Both an increase in Ca2+ current (I(Ca)) and a decrease in K+ transient outward current (I(to)) underlay this effect. Consistently, the relative expression of mRNA coding for the Ca2+ channel alpha1C subunit (Ca(v)1.2) increased, and that of the K+ channel K(v)4.2 subunit decreased. Three weeks after MI, AP prolongation endured, whereas cellular hypertrophy developed. I(Ca) density, Ca(v)1.2, and K(v)4.2 mRNA levels regained control levels, but I(to) density remained reduced. Long-term treatment with RU28318, an MR antagonist, prevented this electrical remodeling. In a different etiologic model of abdominal aortic constriction, we confirmed that APD prolongation and modifications of ionic currents precede cellular hypertrophy. CONCLUSIONS: Electrical remodeling, which is triggered at least in part by MR activation, is an initial, early cellular response to hypertrophic insults. PMID- 15289367 TI - Prior statin therapy is associated with a decreased rate of severe sepsis. AB - BACKGROUND: Statins have anti-inflammatory properties that are independent of their lipid-lowering abilities. We hypothesized that statin therapy before the onset of an acute bacterial infection may have a protective effect against severe sepsis. The aim of this study was to determine whether patients treated with statins develop severe sepsis less frequently. METHODS AND RESULTS: In this prospective observational cohort study, consecutive patients admitted with presumed or documented acute bacterial infection were enrolled. The primary outcomes were the rate of severe sepsis and intensive care unit (ICU) admission. Of the 361 patients enrolled, 82 (22.7%) were treated with statins before their admission. Both groups had a similar severity of illness on admission. Severe sepsis developed in 19% of patients in the no-statin group and in only 2.4% of the statin group (P<0.001). Statin treatment was associated with a relative risk of developing severe sepsis of 0.13 (95% CI, 0.03 to 0.52) and an absolute risk reduction of 16.6%. The overall ICU admission rate was 10.2% (37/361): 12.2% of the no-statin group required ICU admission, whereas in the statin group only 3.7% were admitted to the ICU (P=0.025), reflecting a relative risk of ICU admission of 0.30 (95% CI, 0.1 to 0.95). CONCLUSIONS: Prior therapy with statins may be associated with a reduced rate of severe sepsis and ICU admission. If supported by prospective controlled trials, statins may have a role in the primary prevention of sepsis. PMID- 15289368 TI - Randomized, placebo-controlled trial of dalteparin for the prevention of venous thromboembolism in acutely ill medical patients. AB - BACKGROUND: Considerable variability exists in the use of pharmacological thromboprophylaxis among acutely ill medical patients, partly because clinically relevant end points have not been fully assessed in this population. We undertook an international, multicenter, randomized, double-blind, placebo-controlled trial using clinically important outcomes to assess the efficacy and safety of dalteparin in the prevention of venous thromboembolism in such patients. METHODS AND RESULTS: Patients (n=3706) were randomly assigned to receive either subcutaneous dalteparin 5000 IU daily or placebo for 14 days and were followed up for 90 days. The primary end point was venous thromboembolism, defined as the combination of symptomatic deep vein thrombosis, symptomatic pulmonary embolism, and asymptomatic proximal deep vein thrombosis detected by compression ultrasound at day 21 and sudden death by day 21. The incidence of venous thromboembolism was reduced from 4.96% (73 of 1473 patients) in the placebo group to 2.77% (42 of 1518 patients) in the dalteparin group, an absolute risk reduction of 2.19% or a relative risk reduction of 45% (relative risk, 0.55; 95% CI, 0.38 to 0.80; P=0.0015). The observed benefit was maintained at 90 days. The overall incidence of major bleeding was low but higher in the dalteparin group (9 patients; 0.49%) compared with the placebo group (3 patients; 0.16%). CONCLUSIONS: Dalteparin 5000 IU once daily halved the rate of venous thromboembolism with a low risk of bleeding. PMID- 15289369 TI - Troponin I isoform expression in human and experimental atrial fibrillation. AB - BACKGROUND: Atrial fibrillation (AF) is accompanied by re-expression of fetal genes and activation of proteolytic enzymes. In this study both aspects were addressed with respect to troponin I (TnI) isoform expression. METHODS AND RESULTS: Western blotting and real-time reverse transcription-polymerase chain reaction were used to study TnI isoform expression in patients with paroxysmal or chronic AF and in goats after 1, 2, 4, 8, and 16 weeks of AF. In addition to cardiac TnI (cTnI), low expression of slow-twitch skeletal TnI (ssTnI) protein was found in 60% of patients in sinus rhythm or paroxysmal AF and in 8% of patients with chronic AF. In adult goat atrium, ssTnI protein expression was undetectable. Calcium-dependent degradation of cTnI protein was found in 1 or 2 of 6 animals after 1 to 4 weeks of AF. Although always low, ssTnI mRNA levels were significantly higher in patients who expressed ssTnI protein than in those who did not. Relative ssTnI mRNA expression was significantly lower in patients with paroxysmal AF and chronic AF than in those in sinus rhythm. In goats there was a tendency toward higher relative levels of ssTnI at the onset of AF followed by a normalization when AF had become sustained. CONCLUSIONS: Atrial re expression of ssTnI during paroxysmal AF in patients and during the first 2 weeks of pacing-induced AF in goats does not seem to be part of the process of AF associated cardiomyocyte dedifferentiation but seems to result from transient cardiomyocyte stress at the onset of AF. PMID- 15289370 TI - Possible inhibition of focal cerebral ischemia by angiotensin II type 2 receptor stimulation. AB - BACKGROUND: The role of angiotensin II receptor subtypes was investigated in focal brain ischemia induced by middle cerebral artery (MCA) occlusion. METHODS AND RESULTS: In Agtr2+ (wild-type) mice, MCA occlusion induced focal ischemia of approximately 20% to 30% of the total area in coronal section of the brain. The ischemic area was significantly larger in angiotensin II type 2 receptor deficient (Agtr2-) mice than in Agtr2+ mice. The neurological deficit after MCA occlusion was also greater in Agtr2- mice than in Agtr2+ mice. The decrease in surface cerebral blood flow after MCA occlusion was significantly exaggerated in the peripheral region of the MCA territory in Agtr2- mice. Superoxide production and NADPH oxidase activity were enhanced in the ischemic area of the brain in Agtr2- mice. An AT1 receptor blocker, valsartan, at a nonhypotensive dose significantly inhibited the ischemic area, neurological deficit, and reduction of cerebral blood flow as well as superoxide production and NADPH oxidase activity in Agtr2+ mice. These inhibitory actions of valsartan were weaker in Agtr2- mice. CONCLUSIONS: These results suggest that AT2 receptor stimulation has a protective effect on ischemic brain lesions, at least partly through the modulation of cerebral blood flow and superoxide production. PMID- 15289371 TI - Endothelial dysfunction and mild renal insufficiency in essential hypertension. AB - BACKGROUND: Mild to moderate renal insufficiency in individuals with essential hypertension is currently considered the expression of a renal microvasculopathy characterized by preglomerular arteriolar involvement and tubulo-interstitial changes. Whether endothelial dysfunction plays a role in this alteration is still undefined. METHODS AND RESULTS: We investigated the relationship between endothelial function (hemodynamic response to acetylcholine [ACh] in the forearm) and renal function in 500 patients with uncomplicated, never-treated, essential hypertension and serum creatinine within the normal range (ie, < or =1.5 mg/dL). Serum creatinine, creatinine clearance, and estimated glomerular filtration rate (GFR, by the Modification of Diet in Renal Disease formula) were related to the forearm blood flow response to ACh (all P< or =0.003), and these relationships held true in multiple regression analyses that included age, gender, systolic pressure, serum cholesterol and glucose, smoking, and body mass index. Accordingly, on multiple logistic regression analysis, the risk of moderate renal dysfunction (ie, an estimated GFR <60 mL x min(-1) x 1.73 m(-2)) was 64% lower (OR 0.36, 95% CI 0.18 to 0.70) in patients in the third ACh tertile (ie, those showing the higher vasodilatory response) than in those in the first tertile (ie, showing the lower response). C-reactive protein was related directly to serum creatinine and inversely to GFR and vasodilatory response to ACh, which suggests that endothelial dysfunction is a possible mechanism linking inflammation and impaired renal function in essential hypertension. CONCLUSIONS: An impaired vasodilatory response to ACh appears to be associated with renal function loss in patients with essential hypertension. This association suggests that systemic endothelial dysfunction is involved in mild to moderate renal insufficiency in patients with uncomplicated essential hypertension. PMID- 15289372 TI - Antioxidant effects of statins via S-nitrosylation and activation of thioredoxin in endothelial cells: a novel vasculoprotective function of statins. AB - BACKGROUND: HMG-CoA reductase inhibitors (statins) are lipid-lowering drugs that also exert pleiotropic vasculoprotective effects via activation of the endothelial NO synthesis. NO induces S-nitrosylation of target proteins. S Nitrosylation of the antioxidant enzyme thioredoxin was recently shown to enhance its activity, thereby reducing intracellular reactive oxygen species. Therefore, we investigated whether statins may exert an antioxidant activity in endothelial cells via S-nitrosylation of thioredoxin. METHODS AND RESULTS: Statins dose- and time-dependently increased the overall level of S-nitrosylated proteins in endothelial cells (atorvastatin 0.1 micromol/L, 206+/-30% increase; simvastatin 1 micromol/L, 214+/-19% increase; mevastatin 1 micromol/L, 191+/-10% increase). The increased S-nitrosylation was blocked by an NO-synthase inhibitor and mevalonate. Moreover, S-nitrosylation of thioredoxin was also significantly augmented after atorvastatin treatment. The atorvastatin-mediated increase in S-nitrosylation was associated with an enhanced enzymatic activity of thioredoxin (atorvastatin, 157+/-9% increase). This resulted in a significant reduction of intracellular reactive oxygen species within the endothelial cells. In contrast, in endothelial cells overexpressing a thioredoxin construct in which the S-nitrosylation acceptor amino acid cysteine 69 was replaced by serine [TRX(C69S)], atorvastatin did not activate the redox-regulatory activity of thioredoxin. Moreover, overexpression of the non-nitrosylatable thioredoxin TRX(C69S) abolished atorvastatin-mediated reduction of reactive oxygen species. CONCLUSIONS: Here, we demonstrate a novel antioxidant mechanism by which statins reduce reactive oxygen species in endothelial cells. Statin-mediated S-nitrosylation of thioredoxin enhanced the enzymatic activity of thioredoxin, resulting in a significant reduction in intracellular reactive oxygen species. PMID- 15289373 TI - Neuregulins regulate cardiac parasympathetic activity: muscarinic modulation of beta-adrenergic activity in myocytes from mice with neuregulin-1 gene deletion. AB - BACKGROUND: Neuregulins are required for maintenance of acetylcholine receptor inducing activity of nicotinic receptors in neurons and skeletal muscle, but effects of neuregulins on muscarinic receptors are not known. In the normal heart, parasympathetic activation counterbalances beta-adrenergic activation. To test the hypothesis that neuregulins modify parasympathetic function in the heart, we studied cardiomyocytes from mice heterozygous for neuregulin-1 gene deletion (NRG-1+/-) and examined the effects of beta-adrenergic stimulation on contractility in the presence and absence of the muscarinic agonist carbachol. METHODS AND RESULTS: We evaluated contraction and intracellular Ca2+ transients ([Ca2+]i) in left ventricular (LV) myocytes loaded with Fluo-3 from NRG-1+/- and wild-type (WT) mice. Under baseline conditions (0.5 Hz, 1.5 mmol/L [Ca2+]o, 25 degrees C), characteristics of myocyte contraction/relengthening and systolic/diastolic [Ca2+]i were not different between WT and NRG-1+/- mice. The steady-state increases in fractional shortening (FS) and peak-systolic [Ca2+]i in response to isoproterenol were similar in both groups. In WT myocytes stimulated with isoproterenol, carbachol decreased FS, peak-systolic [Ca2+]i, and cAMP levels. In NRG-1+/- myocytes, carbachol did not attenuate either FS or peak systolic [Ca2+]i, associated with the failure to decrease cAMP levels. Investigation of muscarinic receptor signaling showed no difference of LV protein levels of muscarinic M2 receptors or G protein Galpha(i1,2), Galpha(i3), and Galpha(o) subunits. CONCLUSIONS: Cardiomyocytes deficient in neuregulin signaling are unable to adequately counterbalance beta-adrenergic activation by inhibitory parasympathetic activity. This mechanism may contribute to the known increased risk of heart failure in injured human hearts when neuregulin signaling is suppressed. PMID- 15289374 TI - Myocardial first-pass perfusion magnetic resonance imaging: a multicenter dose ranging study. AB - BACKGROUND: MRI can identify patients with obstructive coronary artery disease by imaging the left ventricular myocardium during a first-pass contrast bolus in the presence and absence of pharmacologically induced myocardial hyperemia. The purpose of this multicenter dose-ranging study was to determine the minimally efficacious dose of gadopentetate dimeglumine injection (Magnevist Injection; Berlex Laboratories) for detecting obstructive coronary artery disease. METHOD AND RESULTS: A total of 99 patients scheduled for coronary artery catheterization as part of their clinical evaluation were enrolled in this study. Patients were randomized to 1 of 3 doses of gadopentate dimeglumine: 0.05, 0.10, or 0.15 mmol/kg. First-pass perfusion imaging was performed during hyperemia (induced by a 4-minute infusion of adenosine at a rate of 140 microg x kg(-1) x min(-1)) and then again in the absence of adenosine with otherwise identical imaging parameters and the same contrast dose. Perfusion defects were evaluated subjectively by 4 blinded reviewers. Receiver-operating curve analysis showed that the areas under the receiver-operating curve were 0.90, 0.72, and 0.83 for the low-, medium-, and high-contrast doses, respectively, compared with quantitative coronary angiography (diameter stenosis > or =70%). For the low-dose group, mean sensitivity was 93+/-0%, mean specificity was 75+/-7%, and mean accuracy was 85+/-3%. CONCLUSIONS: First-pass perfusion MRI is a safe and accurate test for identifying patients with obstructive coronary artery disease. A low dose of 0.05 mmol/kg gadopentetate dimeglumine is at least as efficacious as higher doses. PMID- 15289375 TI - Outcomes in children with idiopathic pulmonary arterial hypertension. AB - BACKGROUND: Treatment for idiopathic pulmonary arterial hypertension in children includes calcium channel blockade (CCB) for acute responders with vasodilator testing and chronic epoprostenol for nonresponders. We sought to determine parameters associated with survival and treatment success. METHODS AND RESULTS: A previously identified cohort of 77 children diagnosed between 1982 and 1995 with idiopathic pulmonary arterial hypertension was followed up through 2002. For acute responders treated with CCB (n=31), survival at 1, 5, and 10 years was 97%, 97%, and 81%, respectively; treatment success was 84%, 68%, and 47%, respectively. Survival for all children treated with epoprostenol (n=35) at 1, 5, and 10 years was 94%, 81%, and 61%, respectively; treatment success was 83%, 57%, and 37%, respectively. Because of the inconsistent availability of epoprostenol before 1995, we defined a "recent medical era" subset by excluding children from the total 77 patient cohort for whom epoprostenol was recommended but was unavailable. Survival in the recent medical era (n=44) at 1, 5, and 10 years was 97%, 97%, and 78%; treatment success was 93%, 86%, and 60%, respectively. Treatment success on CCB decreased significantly when acute responders became nonresponders. Age at diagnosis predicted treatment success in the recent medical era. CONCLUSIONS: Survival for children with idiopathic pulmonary arterial hypertension has significantly improved with CCB and epoprostenol. Children who are acute responders are treated with CCB; they are treated with epoprostenol if they become nonresponders. The decrease in survival and in treatment success after 5 years in all children supports the role for transplant evaluation before treatment failure. PMID- 15289376 TI - Prolonged endoplasmic reticulum stress in hypertrophic and failing heart after aortic constriction: possible contribution of endoplasmic reticulum stress to cardiac myocyte apoptosis. AB - BACKGROUND: The endoplasmic reticulum (ER) is recognized as an organelle that participates in folding secretory and membrane proteins. The ER responds to stress by upregulating ER chaperones, but prolonged and/or excess ER stress leads to apoptosis. However, the potential role of ER stress in pathophysiological hearts remains unclear. METHODS AND RESULTS: Mice were subjected to transverse aortic constriction (TAC) or sham operation. Echocardiographic analysis demonstrated that mice 1 and 4 weeks after TAC had cardiac hypertrophy and failure, respectively. Cardiac expression of ER chaperones was significantly increased 1 and 4 weeks after TAC, indicating that pressure overload by TAC induced prolonged ER stress. In addition, the number of terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL)-positive cells increased, and caspase-3 was cleaved in failing hearts. The antagonism of angiotensin II type 1 receptor prevented upregulation of ER chaperones and apoptosis in failing hearts. On the other hand, angiotensin II upregulated ER chaperones and induced apoptosis in cultured adult rat cardiac myocytes. We also investigated possible signaling pathways for ER-initiated apoptosis. The CHOP- (a transcription factor induced by ER stress), but not JNK- or caspase-12-, dependent pathway was activated in failing hearts by TAC. Pharmacological ER stress inducers upregulated ER chaperones and induced apoptosis in cultured cardiac myocytes. Finally, mRNA levels of ER chaperones were markedly increased in failing hearts of patients with elevated brain natriuretic peptide levels. CONCLUSIONS: These findings suggest that pressure overload by TAC induces prolonged ER stress, which may contribute to cardiac myocyte apoptosis during progression from cardiac hypertrophy to failure. PMID- 15289377 TI - Targeted inactivation of cystic fibrosis transmembrane conductance regulator chloride channel gene prevents ischemic preconditioning in isolated mouse heart. AB - BACKGROUND: Recent evidence suggests that chloride channels may be involved in ischemic preconditioning (IPC). In this study, we tested whether the cystic fibrosis transmembrane conductance regulator (CFTR) chloride channels, which are expressed in the heart and activated by protein kinase A and protein kinase C, are important for IPC in isolated heart preparations from wild-type (WT) and CFTR knockout (CFTR-/-) mice. METHODS AND RESULTS: Hearts were isolated from age matched WT or CFTR-/- (B6.129P2-Cftr(tm1Unc) and STOCKCftr(tm1Unc)-TgN 1Jaw) mice and perfused in the Langendorff or working-heart mode. All hearts were allowed to stabilize for 10 minutes before they were subjected to 30 or 45 minutes of global ischemia followed by 40 minutes of reperfusion (control group) or 3 cycles of 5 minutes of ischemia and reperfusion (IPC group) before 30 or 45 minutes of global ischemia and 40 minutes of reperfusion. Hemodynamic indices were recorded to evaluate cardiac functions. Release of creatine phosphate kinase (CPK) in the samples of coronary effluent and infarct size of the ventricles were used to estimate myocardial tissue injury. In WT adult hearts, IPC protected cardiac function during reperfusion and significantly decreased ischemia-induced CPK release and infarct size. A selective CFTR channel blocker, gemfibrozil, abrogated the protective effect of IPC. Furthermore, targeted inactivation of the CFTR gene in 2 different strains of CFTR-/- mice also prevented IPC's protection of cardiac function and myocardial injury against sustained ischemia. CONCLUSIONS: CFTR Cl- channels may serve as novel and crucial mediators in mouse heart IPC. PMID- 15289378 TI - Measures of insulin resistance add incremental value to the clinical diagnosis of metabolic syndrome in association with coronary atherosclerosis. AB - BACKGROUND: Whether measures of insulin resistance provide incremental information regarding atherosclerotic cardiovascular disease beyond current National Cholesterol Education Program (NCEP) Adult Treatment Panel III metabolic syndrome (MetSyn) criteria or inflammatory markers is uncertain. METHODS AND RESULTS: We examined the association of insulin resistance and MetSyn with coronary artery calcification (CAC) in 840 asymptomatic nondiabetic subjects. Both NCEP and World Health Organization-defined MetSyn were associated (ordinal regression odds ratio [OR] and 95% confidence intervals for NCEP-defined MetSyn) with CAC after controlling for age, non-MetSyn risk factors, and plasma CRP levels (OR, 1.93 [1.43 to 2.60], P<0.001) and after further controlling for homeostasis model assessment index (HOMA) (OR, 1.56 [1.14 to 2.15], P=0.006). Conversely, HOMA was significantly associated with CAC after adjusting for age, non-MetSyn risk factors, and CRP levels (OR, 1.62 [1.31 to 2.01], P<0.001) and after further adjusting for NCEP-defined MetSyn (OR, 1.45 [1.16 to 1.82], P=0.007). Addition of HOMA to the NCEP MetSyn significantly improved the association with CAC, but addition of CRP data to MetSyn or HOMA did not. CONCLUSIONS: Both MetSyn and HOMA index were associated with coronary atherosclerosis independent of established risk factors, including CRP. These findings support the use of biomarkers of insulin resistance in addition to NCEP MetSyn criteria in assessing cardiovascular disease risk. PMID- 15289379 TI - Gene therapy ameliorates cardiovascular disease in dogs with mucopolysaccharidosis VII. AB - BACKGROUND: Mucopolysaccharidosis VII (MPS VII) is a lysosomal storage disease caused by deficient beta-glucuronidase (GUSB) activity resulting in defective catabolism of glycosaminoglycans (GAGs). Cardiac disease is a major cause of death in MPS VII because of accumulation of GAGs in cardiovascular cells. Manifestations include cardiomyopathy, mitral and aortic valve thickening, and aortic root dilation and may cause death in the early months of life or may be compatible with a fairly normal lifespan. We previously reported that neonatal administration of a retroviral vector (RV) resulted in transduction of hepatocytes, which secreted GUSB into the blood and could be taken up by cells throughout the body. The goal of this study was to evaluate the effect on cardiac disease. METHODS AND RESULTS: Six MPS VII dogs were treated intravenously with an RV-expressing canine GUSB. Echocardiographic parameters, cardiovascular lesions, and biochemical parameters of these dogs were compared with those of normal and untreated MPS VII dogs. CONCLUSIONS: RV-treated dogs were markedly improved compared with untreated MPS VII dogs. Most RV-treated MPS VII dogs had mild or moderate mitral regurgitation at 4 to 5 months after birth, which improved or disappeared when evaluated at 9 to 11 and at 24 months. Similarly, mitral valve thickening present early in some animals disappeared over time, whereas aortic dilation and aortic valve thickening were absent at all times. Both myocardium and aorta had significant levels of GUSB and reduction in GAGs. PMID- 15289380 TI - Role of mitochondrial aldehyde dehydrogenase in nitroglycerin-induced vasodilation of coronary and systemic vessels: an intact canine model. AB - BACKGROUND: It has recently been shown that mitochondrial aldehyde dehydrogenase 2 (mtALDH) catalyzes the formation of 1,2-glyceryl dinitrate and nitrite from nitroglycerin (glyceryl trinitrate [GTN]) within mitochondria, leading to production of cGMP and vasorelaxation. However, whether this mechanism operates in the systemic and coronary beds that subserve the antianginal action of GTN is not known. In this study, we address this question in an intact canine model. METHODS AND RESULTS: Fourteen healthy mongrel dogs (weight, 20 to 25 kg) were studied. Coronary blood flow and hemodynamics were continuously monitored by a pulse Doppler flow probe implanted around the left circumflex coronary artery and with catheters in left ventricle and aorta, respectively. Each dog was given a 1 mL bolus injection of GTN, sodium nitroprusside (SNP), or adenosine through a catheter in the left atrium before and 30 minutes after infusion of cyanamide (17 mg/kg), an inhibitor of mtALDH. Cyanamide significantly inhibited both the classic dehydrogenase and GTN reductase activities of mtALDH in situ and attenuated the coronary blood flow increase and declines in blood pressure and left ventricular end-diastolic pressure produced by GTN in vivo. In contrast, mtALDH inhibition had no effect on the coronary and systemic effects of SNP and adenosine. CONCLUSIONS: Our data suggest that mtALDH contributes to GTN biotransformation in vivo and thus at least partly underlies the antianginal mechanism of drug action. Our findings also highlight the differences in biometabolism of clinically relevant nitrosovasodilators. PMID- 15289381 TI - Raf-1 kinase is required for cardiac hypertrophy and cardiomyocyte survival in response to pressure overload. AB - BACKGROUND: Cardiac hypertrophy is a common response to pressure overload and is associated with increased mortality. Mechanical stress in the heart results in the activation of the small GTPase ras and the Raf-1/MEK/ERK signaling cascade in addition to other signaling pathways. METHODS AND RESULTS: In an attempt to determine the requirement for the serine/threonine kinase Raf-1 in the pathogenesis of cardiac hypertrophy, we generated transgenic mice with cardiac specific expression of a dominant negative form of Raf-1 (DN-Raf). DN-Raf mice appeared normal at birth, were fertile, and had normal cardiac structure and function in the absence of provocative stimulation. In response to pressure overload, cardiac extracellular signal-regulated kinase (ERK) activation was inhibited, but c-Jun N-terminal kinase (JNK) activation and p38 mitogen-activated protein kinase (MAPK) activation were normal. DN-Raf mice were sensitized to pressure overload and the development of cardiomyocyte apoptosis, and >35% of animals died within 7 days of aortic banding. Surviving DN-Raf animals were markedly resistant to the development of cardiac hypertrophy and hypertrophic gene induction in response to transverse aortic constriction. CONCLUSIONS: These results establish that Raf-1 kinase activity is essential for cardiac hypertrophy and cardiomyocyte survival in response to pressure overload. PMID- 15289382 TI - Profiling of aortic smooth muscle cell gene expression in response to chronic inhibition of nitric oxide synthase in rats. AB - BACKGROUND: Chronic inhibition of nitric oxide (NO) synthesis by N(omega)-nitro-L arginine methyl ester (L-NAME) induces hypertension associated with remodeling of the arterial wall. In this study, we aimed at identifying genes and pathways involved in this process in aortic smooth muscle cells from Fischer 344 rats, which exhibit an accelerated hypertension after administration of L-NAME. METHODS AND RESULTS: We studied the transcriptional profile of aortic media after 15 days (moderate hypertension) and 30 days (accelerated hypertension) of L-NAME administration (50 mg x kg(-1) x d(-1)) by using rat Affymetrix Genechips, and we present a large-scale validation of the DNA chip results by real-time reverse transcription-polymerase chain reaction (RT-PCR). We observed, in aortic media, a progressive increase in the number of modulated genes during L-NAME administration, with 53 genes significantly modulated after 15 days and 147 genes after 30 days. These expression changes were confirmed at 87% by RT-PCR. We found 28 known genes regulated at both 15 and 30 days (96% confirmation by RT-PCR). The functional classification of the regulated genes highlights 3 major biological pathways modulated in aortic media during L-NAME administration: genes regulating cell proliferation, genes involved in the extracellular matrix remodeling, and genes of the NO/cGMP signaling pathway. CONCLUSIONS: As a consequence of the genomic approach, we observed a large increase in modulation of gene expression along the evolution of the model and the progressive implication of compensatory mechanisms, making expression profile analysis more complex. PMID- 15289383 TI - National patterns of use and effectiveness of angiotensin-converting enzyme inhibitors in older patients with heart failure and left ventricular systolic dysfunction. AB - BACKGROUND: Although ACE inhibitors are underprescribed for heart failure, factors associated with their use are not well described. Furthermore, the effectiveness of ACE inhibitors has been questioned in some populations, potentially contributing to underuse. Our objectives were to assess the correlates of ACE inhibitor use and the relationship between ACE inhibitor prescription and mortality in older patients with heart failure. METHODS AND RESULTS: We studied a national sample aged > or =65 years who had survived hospitalization for heart failure between April 1998 and March 1999 or July 2000 and June 2001, restricting the analysis to patients with left ventricular systolic dysfunction and without a documented contraindication to use of ACE inhibitors (n=17 456). Factors associated with ACE inhibitor prescription at discharge and the relationship between ACE inhibitor prescription and death within 1 year were assessed with hierarchical logistic models. Secondary analyses assessed therapeutic substitution with angiotensin receptor blockers (ARBs). ACE inhibitors were prescribed to only 68% of this ideal cohort, and 76% received either an ACE inhibitor or an ARB. Patient, physician, and hospital factors were weak predictors of prescription, except for serum creatinine (RR for 133 to 221 micromol/L=0.87, 95% CI 0.85 to 0.89; RR for > or =222 micromol/L=0.53, 95% CI 0.49 to 0.57 compared with < or =132 micromol/L). ACE inhibitor prescription was associated with lower crude 1-year mortality (33.0% versus 42.1%, P<0.001), lower risk of death after adjustment (RR 0.86, 95% CI 0.82 to 0.90), and lower mortality regardless of patient gender, age, race, or serum creatinine level. CONCLUSIONS: ACE inhibitors were widely underprescribed despite evidence of a favorable impact on survival in a broad range of patients with heart failure. These results emphasize the importance of ongoing efforts to translate clinical trial results into practice. PMID- 15289384 TI - Comparison of dobutamine stress magnetic resonance, adenosine stress magnetic resonance, and adenosine stress magnetic resonance perfusion. AB - BACKGROUND: Dobutamine stress MR (DSMR) is highly accurate for the detection of inducible wall motion abnormalities (IWMAs). Adenosine has a more favorable safety profile and is well established for the assessment of myocardial perfusion. We evaluated the diagnostic value of IWMAs during dobutamine and adenosine stress MR and adenosine MR perfusion compared with invasive coronary angiography. METHODS AND RESULTS: Seventy-nine consecutive patients (suspected or known coronary disease, no history of prior myocardial infarction) scheduled for cardiac catheterization underwent cardiac MR (1.5 T). After 4 minutes of adenosine infusion (140 microg x kg(-1) x min(-1) for 6 minutes), wall motion was assessed (steady-state free precession), and subsequently perfusion scans (3 slice turbo field echo-echo planar imaging; 0.05 mmol/kg Gd-BOPTA) were performed. After a 15-minute break, rest perfusion was imaged, followed by standard DSMR/atropine stress MR. Wall motion was classified as pathological if > or =1 segment showed IWMAs. The transmural extent of inducible perfusion deficits (<25%, 25% to 50%, 51% to 75%, and >75%) was used to grade segmental perfusion. Quantitative coronary angiography was performed with significant stenosis defined as >50% diameter stenosis. Fifty-three patients (67%) had coronary artery stenoses >50%; sensitivity and specificity for detection by dobutamine and adenosine stress and adenosine perfusion were 89% and 80%, 40% and 96%, and 91% and 62%, respectively. Adenosine IWMAs were seen only in segments with >75% transmural perfusion deficit. CONCLUSIONS: DSMR is superior to adenosine stress for the induction of IWMAs in patients with significant coronary artery disease. Visual assessment of adenosine stress perfusion is sensitive with a low specificity, whereas adenosine stress MR wall motion is highly specific because it identifies only patients with high-grade perfusion deficits. Thus, DSMR is the method of choice for current state-of-the-art treatment regimens to detect ischemia in patients with suspected or known coronary artery disease but no history of prior myocardial infarction. PMID- 15289385 TI - Identification of the ventricular tachycardia isthmus after infarction by pace mapping. AB - BACKGROUND: Ventricular tachycardia (VT) isthmuses can be defined by fixed or functional block. During sinus rhythm, pace mapping near the exit of an isthmus should produce a QRS similar to that of VT. Pace mapping at sites proximal to the exit may produce a similar QRS with a longer stimulus-to-QRS interval (S-QRS). The aim of the study was to determine whether a VT isthmus could be identified and followed by pace mapping. METHODS AND RESULTS: Left ventricular pace mapping during sinus rhythm was performed at 819 sites in 11 patients with VT late after infarction, and corresponding CARTO maps were reconstructed. An isthmus site was defined by entrainment and/or VT termination by ablation. Pace-mapping data were analyzed from the identified isthmus site and from sites at progressively increasing distances from this initial isthmus site. Sites where pace mapping produced the same QRS with different S-QRS delays were identified to attempt to trace the course of the isthmus. In 11 patients, 13 confluent low-voltage infarct regions were present. In all these regions, parts of VT isthmuses were identified by pace mapping. In 11 of 13 of the identified isthmus parts, the QRS morphology of the pace map matched a VT QRS. In 10 of 11 patients, radiofrequency ablation rendered clinical VTs noninducible. Successful ablation sites were localized within an isthmus identified by pace mapping in all of these 10 patients. CONCLUSIONS: VT isthmuses can be identified and part of their course delineated by pace mapping during sinus rhythm. This method could help target isthmus sites for ablation during stable sinus rhythm. PMID- 15289386 TI - Economics of sirolimus-eluting stents: drug-eluting stents have really arrived. PMID- 15289387 TI - Comparing warfarin with aspirin after biological aortic valve replacement: a prospective study. AB - BACKGROUND: Patients with prosthetic heart valves have a higher risk of developing valve thrombosis and arterial thromboembolism. Antithrombotic therapy in the early postoperative period after biological aortic valve replacement (BAVR) is controversial. The American College of Cardiology/American Heart Association and European Society of Cardiology guidelines recommend the use of warfarin for the first 3 months after BAVR, although the American College Chest Physician guidelines suggest that the recommendations are very weak and that the risk/benefit is unclear. This prospective study investigated the efficacy of postoperative warfarin compared with aspirin in patients after aortic valve replacement. METHODS AND RESULTS: Patients undergoing BAVR between 2001 and 2002 received 2 antithrombotic therapies: 141 patients received warfarin for the first 3 months, and 108 patients received only aspirin. The major end points evaluated were the rate of cerebral ischemic events, bleeding, and survival. There were 3 and 5 postoperative cerebral ischemic events between 24 hours and 3 months for patients treated with aspirin and warfarin, respectively. After 3 months, the incidence of cerebral ischemic events did not differ between the 2 groups. The rate of major bleeding events, the stroke-free survival, and the overall survival rates were not statistically significant between the warfarin and aspirin groups. CONCLUSIONS: There seem to be no advantages in performing early anticoagulation therapy compared with a low-antiplatelet regimen with regard to early cerebral ischemic events, bleeding, and survival. Currently there is no evidence to support the fact that warfarin is more effective than aspirin. PMID- 15289388 TI - ACC/AHA guidelines for the management of patients with ST-elevation myocardial infarction--executive summary: a report of the American College of Cardiology/American Heart Association Task Force on Practice Guidelines (Writing Committee to Revise the 1999 Guidelines for the Management of Patients With Acute Myocardial Infarction). PMID- 15289389 TI - Antioxidant vitamin supplements and cardiovascular disease. PMID- 15289390 TI - Should radial arteries be used routinely for coronary artery bypass grafting? PMID- 15289391 TI - Marked malapposition and aneurysm formation after sirolimus-eluting coronary stent implantation. PMID- 15289392 TI - 8-iso-prostaglandin F2alpha as a risk marker in patients with coronary heart disease. PMID- 15289393 TI - Bubbles in the heart. PMID- 15289394 TI - Optimal therapy for acute coronary syndromes: the more the better? PMID- 15289395 TI - Further confirmation that a conduction disturbance underlies the electrocardiographic pattern of the so-called Brugada syndrome. PMID- 15289396 TI - Primary prevention of sudden cardiac death: the time of your life. PMID- 15289397 TI - Carbon dioxide insufflation on the number and behavior of air microemboli in open heart surgery. PMID- 15289398 TI - Genetic variability of von Willebrand factor and atherosclerosis. PMID- 15289399 TI - Walnuts and endothelial function in hypercholesterolemic subjects. PMID- 15289400 TI - Pound of prevention, ounce of cure? PMID- 15289403 TI - Treatment of locally advanced breast cancer. PMID- 15289404 TI - Treatment of locally advanced breast cancer. PMID- 15289405 TI - Treatment of locally advanced breast cancer. PMID- 15289407 TI - Crisis in orthopedic care: surgeon and resource shortage. PMID- 15289409 TI - Czech Republic fears EU membership will lure doctors. PMID- 15289416 TI - New directions in drug approval. PMID- 15289417 TI - New outbreaks of polio: WHO issues travel warning. PMID- 15289419 TI - Pulmonary abscess with bacteremia in a young man. PMID- 15289420 TI - Determinants of overdose incidents among illicit opioid users in 5 Canadian cities. AB - BACKGROUND: Drug overdose is a major cause of death and illness among illicit drug users. Previous research has indicated that most illicit drug users experience nonfatal overdoses and has suggested a variety of factors that are associated with risk of overdose. In this study, we examined the occurrence of and the factors associated with nonfatal overdoses within a Canadian sample of illicit opioid users not enrolled in treatment at the time of study recruitment. METHODS: Interviewers used a standard questionnaire to collect data on sociodemographic characteristics, drug use, health and health care, experience in the criminal justice system and treatment for drug problems; they also performed standard assessments for mental health and infectious disease. The association between overdose and sociodemographic and drug-use factors was examined with chi(2) and t test analyses; marginally significant variables were examined with logistic regression to determine independent effects. RESULTS: A total of 679 subjects were interviewed; 651 provided answers sufficient for this analysis. One hundred and twelve (17.2%) of the 651 respondents reported an overdose episode in the previous 6 months. In the logistic regression analysis (after adjustment for sociodemographic factors), homelessness, noninjection use of hydromorphone in the past 30 days and involvement in drug treatment in the past 12 months were predictors of overdose (p < 0.05). INTERPRETATION: Overdose poses a considerable health risk for illicit opioid users. We found that a diverse set of factors was associated with overdose episodes. Prevention efforts will likely be more effective if they can be directed to specific causal factors. PMID- 15289421 TI - Prevalence of overweight and obesity in a provincial population of Canadian preschool children. AB - BACKGROUND: More and more school-aged children in Canada and elsewhere are becoming overweight or obese. Many countries are now reporting a similar trend among preschool children. However, little information is available on the prevalence of overweight and obesity among preschool children in Canada. In addition, available data are based on reported rather than measured heights and weights. We conducted this study to determine the prevalence of overweight and obesity, using measured heights and weights, in the 1997 cohort of children aged 3-5 years born in Newfoundland and Labrador. METHODS: We calculated the body mass indices (BMIs) using heights and weights measured by public health nurses during the province-wide Preschool Health Check Program conducted between October 2000 and January 2003. Descriptive data on the children's BMIs and prevalence estimates were generated and analyzed by sex and age with the use of the classification system recommended by the International Obesity Task Force. RESULTS: Data were available for 4161 of the 5428 children born in 1997; boys and girls were equally represented (50.1% and 49.9% respectively). Overall, 25.6% of the preschool children in the cohort were overweight or obese. The rates did not differ significantly by sex or age group. INTERPRETATION: These results indicate that a high proportion of children aged 3-5 years in Newfoundland and Labrador are overweight or obese. It appears that prevention measures should begin before the age of 3 years. PMID- 15289422 TI - Early childhood obesity: a call for early surveillance and preventive measures. PMID- 15289423 TI - Are the recommendations to use perioperative beta-blocker therapy in patients undergoing noncardiac surgery based on reliable evidence? PMID- 15289424 TI - Ethics review procedures for research in developing countries: a basic presumption of guilt. PMID- 15289425 TI - Vitamin B12 (cobalamin) deficiency in elderly patients. AB - Vitamin B12 or cobalamin deficiency occurs frequently (> 20%) among elderly people, but it is often unrecognized because the clinical manifestations are subtle; they are also potentially serious, particularly from a neuropsychiatric and hematological perspective. Causes of the deficiency include, most frequently, food-cobalamin malabsorption syndrome (> 60% of all cases), pernicious anemia (15%-20% of all cases), insufficient dietary intake and malabsorption. Food cobalamin malabsorption, which has only recently been identified as a significant cause of cobalamin deficiency among elderly people, is characterized by the inability to release cobalamin from food or a deficiency of intestinal cobalamin transport proteins or both. We review the epidemiology and causes of cobalamin deficiency in elderly people, with an emphasis on food-cobalamin malabsorption syndrome. We also review diagnostic and management strategies for cobalamin deficiency. PMID- 15289432 TI - How to make an eye. AB - The eye is an organ of such remarkable complexity and apparently flawless design that it presents a challenge to both evolutionary biologists trying to explain its phylogenetic origins, and developmental biologists hoping to understand its formation during ontogeny. Since the discovery that the transcription factor Pax6 plays a crucial role in specifying the eye throughout the animal kingdom, both groups of biologists have been converging on the conserved mechanisms behind eye formation. Their latest meeting was at the Instituto Juan March in Madrid, at a workshop organized by Walter Gehring (Biozentrum, Basel, Switzerland) and Emili Salo (Universitat de Barcelona, Spain), entitled 'The genetic control of eye development and its evolutionary implications'. The exchange of ideas provided some new insights into the construction and history of the eye. PMID- 15289433 TI - Control of Arabidopsis flowering: the chill before the bloom. AB - The timing of the floral transition has significant consequences for reproductive success in plants. Plants gauge both environmental and endogenous signals before switching to reproductive development. Many temperate species only flower after they have experienced a prolonged period of cold, a process known as vernalization, which aligns flowering with the favourable conditions of spring. Considerable progress has been made in understanding the molecular basis of vernalization in Arabidopsis. A central player in this process is FLC, which blocks flowering by inhibiting genes required to switch the meristem from vegetative to floral development. Recent data shows that many regulators of FLC alter chromatin structure or are involved in RNA processing. PMID- 15289435 TI - Integration of anteroposterior and dorsoventral regulation of Phox2b transcription in cranial motoneuron progenitors by homeodomain proteins. AB - Little is known about the molecular mechanisms that integrate anteroposterior (AP) and dorsoventral (DV) positional information in neural progenitors that specify distinct neuronal types within the vertebrate neural tube. We have previously shown that in ventral rhombomere (r)4 of Hoxb1 and Hoxb2 mutant mouse embryos, Phox2b expression is not properly maintained in the visceral motoneuron progenitor domain (pMNv), resulting in a switch to serotonergic fate. Here, we show that Phox2b is a direct target of Hoxb1 and Hoxb2. We found a highly conserved Phox2b proximal enhancer that mediates rhombomere-restricted expression and contains separate Pbx-Hox (PH) and Prep/Meis (P/M) binding sites. We further show that both the PH and P/M sites are essential for Hox-Pbx-Prep ternary complex formation and regulation of the Phox2b enhancer activity in ventral r4. Moreover, the DV factor Nkx2.2 enhances Hox-mediated transactivation via a derepression mechanism. Finally, we show that induction of ectopic Phox2b expressing visceral motoneurons in the chick hindbrain requires the combined activities of Hox and Nkx2 homeodomain proteins. This study takes an important first step to understand how activators and repressors, induced along the AP and DV axes in response to signaling pathways, interact to regulate specific target gene promoters, leading to neuronal fate specification in the appropriate developmental context. PMID- 15289436 TI - Disrupted gonadogenesis and male-to-female sex reversal in Pod1 knockout mice. AB - Congenital defects in genital and/or gonadal development occur in 1 in 1000 humans, but the molecular basis for these defects in most cases remains undefined. We show that the basic helix-loop-helix transcription factor Pod1 (capsulin/epicardin/Tcf21) is essential for normal development of the testes and ovaries, and hence for sexual differentiation. The gonads of Pod1 knockout (KO) mice were markedly hypoplastic, and the urogenital tracts of both XX and XY mice remained indistinguishable throughout embryogenesis. Within Pod1 KO gonads, the number of cells expressing the cholesterol side-chain cleavage enzyme (Scc) was increased markedly. Biochemical and genetic approaches demonstrated that Pod1 transcriptionally represses steroidogenic factor 1 (Sf1/Nr5a1/Ad4BP), an orphan nuclear receptor that regulates the expression of multiple genes (including Scc) that mediate sexual differentiation. Our results establish that Pod1 is essential for gonadal development, and place it in a transcriptional network that orchestrates cell fate decisions in gonadal progenitors. PMID- 15289434 TI - Ribosomal protein L24 defect in belly spot and tail (Bst), a mouse Minute. AB - Ribosomal protein mutations, termed Minutes, have been instrumental in studying the coordination of cell and tissue growth in Drosophila. Although abundant in flies, equivalent defects in mammals are relatively unknown. Belly spot and tail (Bst) is a semidominant mouse mutation that disrupts pigmentation, somitogenesis and retinal cell fate determination. Here, we identify Bst as a deletion within the Rpl24 riboprotein gene. Bst significantly impairs Rpl24 splicing and ribosome biogenesis. Bst/+ cells have decreased rates of protein synthesis and proliferation, and are outcompeted by wild-type cells in C57BLKS<-->ROSA26 chimeras. Bacterial artificial chromosome (BAC) and cDNA transgenes correct the mutant phenotypes. Our findings establish Bst as a mouse Minute and provide the first detailed characterization of a mammalian ribosomal protein mutation. PMID- 15289437 TI - The T-Box transcription factor Tbx5 is required for the patterning and maturation of the murine cardiac conduction system. AB - We report a critical role for the T-box transcription factor Tbx5 in development and maturation of the cardiac conduction system. We find that Tbx5 is expressed throughout the central conduction system, including the atrioventricular bundle and bundle branch conduction system. Tbx5 haploinsufficiency in mice (Tbx5(del/+)), a model of human Holt-Oram syndrome, caused distinct morphological and functional defects in the atrioventricular and bundle branch conduction systems. In the atrioventricular canal, Tbx5 haploinsufficiency caused a maturation failure of conduction system morphology and function. Electrophysiologic testing of Tbx5(del/+) mice suggested a specific atrioventricular node maturation failure. In the ventricular conduction system, Tbx5 haploinsufficiency caused patterning defects of both the left and right ventricular bundle branches, including absence or severe abnormalities of the right bundle branch. Absence of the right bundle branch correlated with right bundle-branch block by ECG. Deficiencies in the gap junction protein gene connexin 40 (Cx40), a downstream target of Tbx5, did not account for morphologic conduction system defects in Tbx5(del/+) mice. We conclude that Tbx5 is required for Cx40-independent patterning of the cardiac conduction system, and suggest that the electrophysiologic defects in Holt-Oram syndrome reflect a developmental abnormality of the conduction system. PMID- 15289438 TI - Iodotyrosine dehalogenase 1 (DEHAL1) is a transmembrane protein involved in the recycling of iodide close to the thyroglobulin iodination site. AB - In the thyroid, iodotyrosine dehalogenase acts on the mono and diiodotyrosines released during the hydrolysis of thyroglobulin to liberate iodide, which can then reenter the hormone-producing pathways. It has been reported that the deiodination of iodotyrosines occurs predominantly in the microsomes and is mediated by NADPH. Recently, two cDNAs, 7401- and 7513-base pairs long that encode proteins with a conserved nitroreductase domain were published in GenBank as iodotyrosine dehalogenase 1 (DEHAL1) and iodotyrosine dehalogenase 1B (DEHAL1B), respectively. We report here our investigation of the localization and activity of one of these isoforms, DEHAL1. DEHAL1 mRNA is highly expressed in the thyroid, is up-regulated by cAMP, and encodes a transmembrane protein that efficiently catalyzes the NADPH-dependent deiodination of mono (L-MIT) and diiodotyrosine (L-DIT), with greater activity vs. L-MIT. Iodotyrosine deiodinase was active in HEK293 cells transfected by DEHAL1 cDNA, but not in CHO cells. A fraction of DEHAL1 protein is exposed to the cell surface, as indicated by biotinylation experiments. Immunohistochemistry studies showed that DEHAL1 proteins accumulate at the apical pole of thyrocytes. Taken together, these findings indicate that the deiodination reaction occurs at the apical pole of the thyrocyte and is involved in a rapid iodide recycling process at and/or close to the organification site. PMID- 15289439 TI - Differential role of estrogen receptor isoforms in sex-specific brain organization. AB - Transient activation of estrogen receptors (ER) in the developing brain during a limited perinatal "window of time" is recognized as a key mechanism of defeminization of neural control of reproductive function and sexual behavior. Two major ER isoforms, alpha and beta, are present in neural circuits that govern ovarian cycle and sexual behavior. Using highly selective ER agonists, this study provides the first evidence for distinct contribution of individual ER isoforms to the process of estrogen dependent defeminization. Neonatal activation of the ERalpha in female rats resulted in abrogation of cyclic ovarian activity and female sexual behavior in adulthood. These effects are associated with male-like alterations in the morphology of the anteroventral periventricular (AVPV) and sexually dimorphic nucleus of the preoptic area (SDN-POA), as well as refractoriness to estrogen-mediated induction of sexual receptivity. Exposure to an ERbeta-selective agonist induced persistent estrus and had a strong defeminizing effect on the hypothalamic gonadotropin "surge generator" AVPV. However, neonatal ERbeta activation failed to alter female sexual behavior, responsiveness to estrogens and morphometric features of the behaviorally relevant SDN-POA. Thus, although co-present in several brain regions involved in the control of female reproductive function, ER isoforms convey different, and probably not synergistic, chemical signals in the course of neonatal sex-specific brain organization. PMID- 15289440 TI - Inhibition of p38 MAP kinase- and RICK/NF-kappaB-signaling suppresses inflammatory bowel disease. AB - Ulcerative colitis and Crohn's disease are the two entities of chronic inflammatory bowel diseases (IBD). One of the main pathogenic mechanisms is probably a dysregulated immune response triggered by products of the enteric bacterial flora. The goal of this study was to evaluate the effects of the p38 mitogen-activated protein kinase (MAPK) inhibitor SB203580 on inflammatory responses using the DSS-induced experimental colitis model in mice reflecting human IBD. We found that SB203580 improved the clinical score, ameliorates the histological alterations, and reduces the mRNA levels of proinflammatory cytokines. In addition to p38 kinase activity, the "classical" and the "alternative" NF-kappaB pathways were also strongly activated during colitis induction. All three pathways were drastically down-regulated by SB203580 treatment. An analysis of the molecular basis of NF-kappaB activation revealed that Rip-like interacting caspase-like apoptosis-regulatory protein kinase (RICK), a key component of a pathway leading to NF-kappaB induction, is also strongly inhibited by SB203580. Since RICK is an effector kinase of NOD2, an intracellular receptor of bacterial peptidoglycan, these results support the notion that NOD signaling could play a pivotal role in the IBD pathogenesis. Thus, RICK could represent a novel target for future therapies in human IBD. PMID- 15289441 TI - Altered excitation-contraction coupling with skeletal muscle specific FKBP12 deficiency. AB - The immunophilin FKBP12 binds the skeletal muscle Ca2+ release channel or ryanodine receptor (RyR1), but the functional consequences of this interaction are not known. In this study, we have generated skeletal muscle specific FKBP12 deficient mice to investigate the role of FKBP12 in skeletal muscle. Primary myotubes from these mice show no obvious change in either Ca2+ stores or resting Ca2+ levels but display decreased voltage-gated intracellular Ca2+ release and increased L-type Ca2+ currents. Consistent with the decreased voltage-gated Ca2+ release, maximal tetanic force production is decreased and the force frequency curves are shifted to the right in extensor digitorum longus (EDL) muscles of the mutant mice. In contrast, there is no decrease in maximal tetanic force production in the mutant diaphragm or soleus muscle. The force frequency curve is shifted to the left in the FKBP12-deficient diaphragm muscle compared with controls. No changes in myosin heavy chain (MHC) phenotype are observed in EDL or soleus muscle of the FKBP12-deficient mice, but diaphragm muscle displays an increased ratio of slow to fast MHC isoforms. Also, calcineurin levels are increased in the diaphragm of the mutant mice but not in the soleus or EDL. In summary, FKBP12 deficiency alters both orthograde and retrograde coupling between the L-type Ca2+ channel and RyR1 and the consequences of these changes depend on muscle type and activity. In highly used muscles such as the diaphragm, adaptation to the loss of FKBP12 occurs, possibly due to the increased Ca2+ influx. PMID- 15289442 TI - Estrogen exerts neuroprotective effects via membrane estrogen receptors and rapid Akt/NOS activation. AB - The neuroprotective role of estrogen (E2) is supported by a multitude of experimental and epidemiological data, although its mode of action is not fully understood. The present work was conducted to study the underlying mechanisms of its neuroprotective action, using the rat cell line PC12, an established model for neuronal cell apoptosis and survival. Our results show that E2 (but not androgens or progestins) prevent growth inhibition and apoptosis of PC12 cells, induced by serum deprivation. Several mechanisms of action were investigated: 1) intracellular estrogen receptors (ERs) have been identified but do not appear to mediate the protective effect of E2. 2) The antioxidant properties of E2 cannot explain their protective actions at the concentrations used (10(-12)-10(-6) M). 3) Finally, membrane sites for E2 have been identified, and the underlying initial signaling cascade (2-30 min after E2) has been tested, showing Ca(2+) mobilization-->PI3K activation-->Akt phosporylation-->NOS activation. Inhibition of PI3K or NOS completely reversed the anti-apoptotic effect of E2. These results suggest a new mechanism of neuroprotective action of estrogen. PMID- 15289443 TI - Uncomplicated human obesity is associated with a specific cardiac transcriptome: involvement of the Wnt pathway. AB - A dramatic increase in obesity prevalence and cardiovascular morbidity is expected for the coming years. However, with relevance to the heart, little is known about the specific contribution of obesity on associated morbidity. Consequently, global analysis of gene regulations in human heart was undertaken to monitor molecular regulations related to obesity or to obesity-related hypertension. Transcriptome analysis using cDNA arrays was performed in right appendage biopsies from obese patients (n=5), from patients with arterial hypertension with (n=5) or without obesity (n=5), and from 5 leans. All biopsies came from patients that had cardiac surgery and coronary bypass. Statistical analysis of the data revealed 2686 differentially expressed genes out of 11,500 when compared with lean tissues. Differential expression was verified by real time PCR in 84% of 50 randomly chosen genes. Among genes encountered, 397 were specifically regulated in obese, 1,299 in non-obese hypertensive, and 355 in obese hypertensive patients, respectively, whereas an additional set of 153 genes was differentially expressed in all these situations. Ontology analysis, hierarchical clustering, and molecular pathway analysis indicated that the heart molecular picture of obesity differs clearly from that observed for obesity related hypertension or arterial hypertension. Clearly, the Wnt pathway known to be involved in cardiac hypertrophy mechanisms, showed opposite regulation in obese heart compared with hypertensive heart and potentially prevented the development of cardiac remodeling in obese patients. All over, this work shows that uncomplicated obesity has a strong impact on cardiac gene expression, which could be considered as precursor signs for future cardiac disease and also demonstrates that obesity-related hypertension generates a heart-molecular distinct phenotype that cannot be predicted by a simple sum of the impact of obesity and arterial hypertension on gene expression. PMID- 15289444 TI - P2Y-mediated Ca2+ response is spatiotemporally graded and synchronized in sensory neurons: a two-photon photolysis study. AB - ATP is thought to be an initiator and modulator of noxious pain sensation. We employed two-photon photolysis to apply ATP locally and transiently, thus mimicking ATP release upon cell damage or exocytosis. Using this technique, an increase in intracellular Ca2+ concentration ([Ca2+]i) was induced via P2Y receptors in individual sensory neurons, or in a neurite region. The ATP-induced [Ca2+]i rise was attenuated by applications of either a phospholipase C inhibitor, or inhibitors for IP3 or ryanodine receptors. These results indicate that intracellular Ca2+ stores play a major role in contributing to the increase in [Ca2+]i. Spatiotemporal analysis revealed that local and transient applications of ATP increased [Ca2+]i by release from intracellular stores, but in a unique, graded, and synchronized manner. 1) As the duration of local ATP application was prolonged, the latency decreased and the magnitude of the [Ca2+]i rise increased; 2) The time course of the rising phase of the [Ca2+]i response to ATP was essentially the same over the cell body, once [Ca2+]i had started to rise. It is anticipated that sensory responses may be modulated variably, depending on the spatiotemporal characteristics of the ATP-related [Ca2+]i profile. PMID- 15289445 TI - Hyperglycemia upregulates translation of the fibroblast growth factor 2 mRNA in mouse aorta via internal ribosome entry site. AB - Fibroblast growth factor 2 (FGF-2) is normally synthesized at low levels but is elevated in various pathophysiological conditions including diabetes-associated vascular diseases. FGF-2 expression is regulated translationally through an internal ribosome entry site (IRES) located in its mRNA, which allows a nonclassical cap-independent translation. We addressed the pathophysiological regulation of the IRES in vivo by using a streptozotocin-induced hyperglycemic model known to suppress markedly overall translation. Evaluation of FGF-2 IRES dependent translation was performed with transgenic mice expressing dual luciferase bicistronic mRNA containing the FGF-2 IRES. FGF-2 IRES-dependent reporter activity increased 240% of control in the diabetic aorta although the reporter mRNA levels significantly decreased. Expression of endogenous FGF-2 protein in the aorta closely correlated with the IRES activity but not with FGF-2 mRNA levels. Moreover, the biosynthesis of endogenous FGF-2 protein was stimulated in an IRES-dependent manner by high glucose that significantly suppressed global protein synthesis in aortic smooth muscle cells from the transgenic mice. These results suggest that IRES-dependent translational regulation could play a pathological role in FGF-2 expression in vivo, especially in the cardiovascular consequences of diabetes. PMID- 15289446 TI - Identification of the pregnancy hormone relaxin as glucocorticoid receptor agonist. AB - The insulin-like peptide relaxin is a central hormone of pregnancy, but it also produces anti-fibrotic, myocardial, renal, central-nervous, and vascular effects. Recently, two G protein-coupled receptors, LGR7 and LGR8, have been identified as relaxin receptors. Prompted by reports on immunoregulatory effects of relaxin, we investigated possible interactions with the human glucocorticoid receptor (GR). Relaxin blunted the endotoxin-induced production of inflammatory cytokines (IL-1, IL-6, TNF-alpha) by human macrophages--an effect that was suppressed by the GR antagonist RU-486. In three different cell lines, relaxin induced GR activation, nuclear translocation, and DNA binding as assessed in GRE-luciferase assays. Co immunoprecipitation experiments revealed physical interaction of endogenous and exogenous relaxin with cytoplasmic and nuclear GR. Relaxin competed with GR agonists for GR binding, both in vivo in whole-cell assays, and in vitro in fluorescence polarization assays. Relaxin was shown to up-regulate GR protein expression as well as the number of functionally active GR sites. In LGR7/8-free cells, the relaxin-mediated activation of GR was preserved. In conclusion, relaxin acts as GR agonist--a pathway pivotal to its effects on cytokine secretion by human macrophages. These findings may deepen our understanding of relaxin's abundant physiological actions, as well as our insights into general principles of hormone signaling. PMID- 15289447 TI - Alterations of gene expression in skin and lung of mice exposed to light and cigarette smoke. AB - We previously showed that sunlight-mimicking light induces genotoxic damage not only in skin but also even in lung, bone marrow, and peripheral blood of hairless mice. Moreover, light and smoke acted synergically in the respiratory tract. To clarify the mechanisms involved, we investigated by cDNA-arrays the expression of 746 toxicologically relevant genes in skin and lungs of mice exposed for 28 days to light and/or environmental cigarette smoke. Glutathione-S-transferase-Pi and catalase were overexpressed in the lungs of mice exposed to light only. Moreover, the light induced in skin the expression of genes involved in carcinogenesis, photoaging, and production of genotoxic and oxidizing derivatives traveling at a distance. Smoke induced the expression of multiple genes in both skin and lung, which reflect adaptive responses and mechanisms related to cancer and, possibly, to emphysema and stroke. As shown in mice exposed to both light and smoke, the light tended to increase smoke-induced gene expression in lungs, while smoke tended to attenuate light-induced gene expression in skin. The oral administration of the nonsteroidal anti-inflammatory drug sulindac inhibited the light-induced overexpression of cyclooxygenase-2 and oxidative stress-related genes in skin, and down-regulated smoke-induced genes involved in oxidative stress, removal of damaged proteins, inflammation, and immune response in lung. These results provide a mechanistic insight explaining the systemic alterations induced by both light and smoke in mouse skin and lungs. PMID- 15289448 TI - A new strategy to block tumor growth by inhibiting endocannabinoid inactivation. AB - Endocannabinoid signaling has been shown to be enhanced in several cancer tissues and malignant cells, and studies in cell lines have shown that this up-regulation might serve the purpose of providing transformed cells with a further means to inhibit their proliferation. Here we investigated the effect of inhibitors of endocannabinoid degradation on the growth of rat thyroid tumor xenografts induced in athymic mice. VDM-11, a selective inhibitor of endocannabinoid cellular re uptake, and arachidonoyl-serotonin (AA-5-HT), a selective blocker of endocannabinoid enzymatic hydrolysis, both inhibited the growth in vivo of tumor xenografts induced by the subcutaneous injection of rat thyroid transformed (KiMol) cells. This effect was accompanied by significantly enhanced endocannabinoid concentrations in the tumors excised at the end of the in vivo experiments. Endocannabinoids, as well as VDM-11 and AA-5-HT, inhibited the growth in vitro of the transformed rat thyroid cells used to induce the tumors in vivo, and their effect was reversed at least in part by the cannabinoid CB1 receptor antagonist SR141716A. This compound, however, when administered alone, did not enhance, but instead slightly inhibited, the growth of rat thyroid transformed cells both in vitro and in tumor xenografts induced in vivo. These findings indicate that endocannabinoids tonically control tumor growth in vivo by both CB1-mediated and non-CB1-mediated mechanisms and that, irrespective of the molecular mechanism of their anti-proliferative action, inhibitors of their inactivation might be used for the development of novel anti-cancer drugs. PMID- 15289449 TI - Glutathione depletion up-regulates Bcl-2 in BSO-resistant cells. AB - Glutathione depletion by inhibition of its synthesis with buthionine sulfoximine (BSO) is a focus of the current research in antitumor therapy, BSO being used as chemosensitizer. We had previously shown that two human tumor cell lines (U937 and HepG2) survive to treatment with BSO: BSO can elicit an apoptotic response, but the apoptotic process is aborted after cytochrome c release and before caspase activation, suggesting the development of an adaptive response (FASEB J., 1999, 13, 2031-2036). Here, we investigate the mechanisms of such an adaptation. We found that following BSO, U937 up-regulate Bcl-2 mRNA and protein levels, by a mechanism possibly involving NF-kappaB transcription factor; the increase in protein level is limited by a rapid decay of Bcl-2 in BSO-treated cells, suggesting that redox imbalance speeds up Bcl-2 turnover. BSO-dependent Bcl-2 up regulation is associated with the ability to survive to BSO. Indeed, 1) its abrogation by CAPE or protein synthesis inhibition sensitizes U937 to BSO; 2) in a panel of four tumor lines, BSO-resistant (U937, HepG2, and HGB1) but not BSO sensitive (BL41) cells can up-regulate Bcl-2 following GSH depletion; remarkably, only the latter are chemosensitized by BSO. PMID- 15289450 TI - Stearoylethanolamide exerts anorexic effects in mice via down-regulation of liver stearoyl-coenzyme A desaturase-1 mRNA expression. AB - Given the recent demonstration that oleoylethanolamide (OEA), a cannabinoid receptor-inactive N-acylethanolamine, decreases food intake by activating the nuclear receptor PPARalpha (peroxisome proliferator-activated receptor alpha) in the periphery, we here evaluated the effects of both saturated and unsaturated C18 N-acylethanolamides (C18:0; C18:1; C18:2) in mice feeding behavior after overnight starvation. Our results show stearoylethanolamide (SEA, C18:0) exerts, unlike other unsaturated C18 homologs, a marked dose-dependent anorexic effect evident already at 2 h after its intraperitoneal administration. In addition, oral administration of SEA (25 mg/kg) was also effective in reducing food consumption, an effect ascribed to the molecule itself and not to its catabolites. Moreover, although the anorexic response to oral administered SEA was not associated with changes in the levels of various hematochemical parameters (e.g., glucose, cholesterol, triglycerides, leptin) nor in liver mRNA expression of peroxisome proliferator-activated receptors (PPARs) including PPARalpha, the anorexic effect of SEA was interestingly accompanied by a reduction in liver stearoyl-CoA desaturase-1 (SCD-1) mRNA expression. As SCD-1 has been recently proposed as a molecular target for the treatment of obesity, the novel observation provided here that SEA reduces food intake in mice in a structurally selective manner, in turn, correlated with downregulation of liver SCD-1 mRNA expression, has the potential of providing new insights on a class of lipid mediators with suitable properties for the pharmacological treatment of over-eating dysfunctions. PMID- 15289451 TI - Degradation of BACE by the ubiquitin-proteasome pathway. AB - The amyloid beta protein (Abeta) is derived from beta-amyloid precursor protein (APP). Cleavage of APP by beta-secretase generates a C-terminal fragment (APPCTFbeta or C99), which is subsequently cleaved by gamma-secretase to produce Abeta. BACE (or BACE1), the major beta-secretase involved in cleaving APP, has been identified as a Type 1 membrane-associated aspartyl protease. In this study, we found that treatment with proteasome inhibitors resulted in an increase in APP C99 levels, suggesting that APP processing at the beta-secretase site may be affected by the ubiquitin-proteasome pathway. To investigate whether the degradation of BACE is mediated by the proteasome pathway, cells stably transfected with BACE were treated with lactacystin. We found that BACE protein degradation was inhibited by lactacystin in a time- and dose-dependent manner. Non-proteasome protease inhibitors had no effect on BACE degradation. BACE protein is ubiquitinated. Furthermore, lactacystin increased APP C99 production and Abeta generation. Our data demonstrate that the degradation of BACE proteins and APP processing are regulated by the ubiquitin-proteasome pathway. PMID- 15289452 TI - Alpha-synuclein induces apoptosis by altered expression in human peripheral lymphocyte in Parkinson's disease. AB - Though the etiology of Parkinson's disease (PD) remains unclear, alpha-synuclein (alpha-SN) is regarded as a major causative agent of PD. Several lines of evidence indicate that immunological abnormalities are associated with PD for unknown reasons. The present study was performed to assess whether peripheral blood mononuclear cells (PBMCs) show altered alpha-SN expression in PD patients and to identify its functions, which may be related to peripheral immune abnormalities in PD. alpha-SN was found to be expressed more in 151 idiopathic PD (IPD) patients than in 101 healthy controls, who nevertheless showed as age dependent increases. By in vitro transfection, alpha-SN expression was shown to be correlated with glucocorticoid sensitive apoptosis, possibly caused by the enhanced expression of glucocorticoid receptor (GR), caspase activations (caspase 8, caspase-9), CD95 up-regulation, and reactive oxygen species (ROS) production. An understanding of the correlation between alpha-SN levels and apoptosis in the presence of the coordinated involvement of multiple processes would provide an insight into the molecular basis of the disease. The present study provides a clue that the alpha-SN may be one of the primary causes of the immune abnormalities observed in PD and offers new targets for pharmacotherapeutic intervention. PMID- 15289453 TI - Functional links between telomeres and proteins of the DNA-damage response. AB - In response to DNA damage, cells engage a complex set of events that together comprise the DNA-damage response (DDR). These events bring about the repair of the damage and also slow down or halt cell cycle progression until the damage has been removed. In stark contrast, the ends of linear chromosomes, telomeres, are generally not perceived as DNA damage by the cell even though they terminate the DNA double-helix. Nevertheless, it has become clear over the past few years that many proteins involved in the DDR, particularly those involved in responding to DNA double-strand breaks, also play key roles in telomere maintenance. In this review, we discuss the current knowledge of both the telomere and the DDR, and then propose an integrated model for the events associated with the metabolism of DNA ends in these two distinct physiological contexts. PMID- 15289454 TI - Mutually exclusive mutations of the Pten and ras pathways in skin tumor progression. AB - Pten heterozygous (Pten+/-) mice develop increased papilloma numbers and show decreased carcinoma latency time in comparison with controls after skin treatment with dimethyl benzanthracene (DMBA) and tetradecanoyl-phorbol acetate (TPA). H ras mutation is normally a hallmark of DMBA-TPA-induced skin tumors, but 70% of carcinomas from Pten+/- mice do not exhibit this mutation, and in all cases have lost the wild-type Pten allele. Tumors that retain the Pten wild-type allele also have H-ras mutations, indicating that activation of H-ras and complete loss of Pten are mutually exclusive events in skin carcinomas. Mitogen-activated protein kinase (MAPK) is consistently activated in the tumors with H-ras mutations, but is strongly down-regulated in Pten-/- tumors, suggesting that this pathway is dispensable for skin carcinoma formation. These data have important implications in designing individual therapeutic strategies for the treatment of cancer. PMID- 15289455 TI - Primitive neural stem cells from the mammalian epiblast differentiate to definitive neural stem cells under the control of Notch signaling. AB - Basic fibroblast growth factor (FGF2)-responsive definitive neural stem cells first appear in embryonic day 8.5 (E8.5) mouse embryos, but not in earlier embryos, although neural tissue exists at E7.5. Here, we demonstrate that leukemia inhibitory factor-dependent (but not FGF2-dependent) sphere-forming cells are present in the earlier (E5.5-E7.5) mouse embryo. The resultant clonal sphere cells possess self-renewal capacity and neural multipotentiality, cardinal features of the neural stem cell. However, they also retain some nonneural properties, suggesting that they are the in vivo cells' equivalent of the primitive neural stem cells that form in vitro from embryonic stem cells. The generation of the in vivo primitive neural stem cell was independent of Notch signaling, but the activation of the Notch pathway was important for the transition from the primitive to full definitive neural stem cell properties and for the maintenance of the definitive neural stem cell state. PMID- 15289456 TI - Distinct roles of c-Abl and Atm in oxidative stress response are mediated by protein kinase C delta. AB - c-Abl and Atm have been implicated in cell responses to DNA damage and oxidative stress. However, the molecular mechanisms by which they regulate oxidative stress response remain unclear. In this report, we show that deficiency of c-Abl and deficiency of ATM differentially altered cell responses to oxidative stress by induction of antioxidant protein peroxiredoxin I (Prx I) via Nrf2 and cell death, both of which required protein kinase C (PKC) delta activation and were mediated by reactive oxygen species. c-abl-/- osteoblasts displayed enhanced Prx I induction, elevated Nrf2 levels, and hypersusceptibility to arsenate, which were reinstated by reconstitution of c-Abl; Atm-/- osteoblasts showed the opposite. These phenotypes correlated with increased PKC delta expression in c-abl-/- osteoblasts and decreased PKC delta expression in Atm-/- cells, respectively. The enhanced responses of c-abl-/- osteoblasts could be mimicked by overexpression of PKC delta in normal cells and impeded by inhibition of PKC delta, and diminished responses of Atm-/- cells could be rescued by PKC delta overexpression, indicating that PKC delta mediated the effects of c-Abl and ATM in oxidative stress response. Hence, our results unveiled a previously unrecognized mechanism by which c-Abl and Atm participate in oxidative stress response. PMID- 15289457 TI - BMP signaling mediated by ALK2 in the visceral endoderm is necessary for the generation of primordial germ cells in the mouse embryo. AB - Deletion of various bone morphogenetic proteins (BMPs) and their downstream Smads in mice have clearly shown that BMP signaling is essential for the formation of primordial germ cells (PGCs). However, the molecular mechanism through which this takes place is still unclear. Here, we demonstrate that BMP4 produced in the extraembryonic ectoderm signals through ALK2, a type I BMP receptor, in the visceral endoderm (VE) to induce formation of PGCs from the epiblast. Firstly, embryonic day 5.5-6.0 (E5.5-E6.0) embryos cultured on fibronectin formed PGCs in the presence of VE, but not in its absence. Secondly, Alk2-deficient embryos completely lacked PGCs and the heterozygotes had reduced numbers, resembling Bmp4 deficient phenotypes. Thirdly, expression of constitutively active ALK2 in the VE, but not in the epiblast, was sufficient to rescue the PGC phenotype in Bmp4 deficient embryos. In addition, we show that the requirement for the VE at E5.5 E6.0 can be replaced by culturing embryos stripped of VE on STO cells, indicating that STO cells provide or transduce signals necessary for PGC formation that are normally transmitted by the VE. We propose a model in which direct signaling to proximal epiblast is supplemented by an obligatory indirect BMP-dependent signal via the VE. PMID- 15289458 TI - N-terminal polyubiquitination and degradation of the Arf tumor suppressor. AB - Unknown mechanisms govern degradation of the p19Arf tumor suppressor, an activator of p53 and inhibitor of ribosomal RNA processing. Kinetic metabolic labeling of cells with [3H]-leucine indicated that p19Arf is a relatively stable protein (half-life approximately 6 h) whose degradation depends upon the ubiquitin-proteasome pathway. Although p19Arf binds to the Mdm2 E3 ubiquitin protein ligase to activate p53, neither of these molecules regulates p19Arf turnover. In contrast, the nucleolar protein nucleophosmin/B23, which binds to p19Arf with high stoichiometry, retards its turnover, and Arf mutants that do not efficiently associate with nucleophosmin/B23 are unstable and functionally impaired. Mouse p19Arf, although highly basic (22% arginine content), contains only a single lysine residue absent from human p14ARF, and substitution of arginine for lysine in mouse p19Arf had no effect on its rate of degradation. Mouse p19Arf (either wild-type or lacking lysine) and human p14ARF undergo N terminal polyubiquitination, a process that has not as yet been documented in naturally occurring lysine-less proteins. Re-engineering of the p19Arf N terminus to provide consensus sequences for N-acetylation limited Arf ubiquitination and decelerated its turnover. PMID- 15289459 TI - Reprogramming of a melanoma genome by nuclear transplantation. AB - We have used nuclear transplantation to test whether the reprogramming activity of oocytes can reestablish developmental pluripotency of malignant cancer cells. We show here that the nuclei of leukemia, lymphoma, and breast cancer cells could support normal preimplantation development to the blastocyst stage but failed to produce embryonic stem (ES) cells. However, a blastocyst cloned from a RAS inducible melanoma nucleus gave rise to ES cells with the potential to differentiate into multiple cell types in vivo including melanocytes, lymphocytes, and fibroblasts. Chimeras produced from these ES cells developed cancer with higher penetrance, shorter latency, and an expanded tumor spectrum when compared with the donor mouse model. These results demonstrate that the secondary changes of a melanoma nucleus are compatible with a broad developmental potential but predispose mice to melanomas and other malignant tumors on reactivation of RAS. Our findings serve as a paradigm for studying the tumorigenic effect of a given cancer genome in the context of a whole animal. PMID- 15289460 TI - Role of the Escherichia coli RecQ DNA helicase in SOS signaling and genome stabilization at stalled replication forks. AB - The RecQ protein family is a highly conserved group of DNA helicases that play roles in maintaining genomic stability. In this study, we present biochemical and genetic evidence that Escherichia coli RecQ processes stalled replication forks and participates in SOS signaling. Cells that carry dnaE486, a mutation in the DNA polymerase III alpha-catalytic subunit, induce an RecA-dependent SOS response and become highly filamented at the semirestrictive temperature (38 degrees C). An recQ mutation suppresses the induction of SOS response and the filamentation in the dnaE486 mutant at 38 degrees C, causing appearance of a high proportion of anucleate cells. In vitro, RecQ binds and unwinds forked DNA substrates with a gap on the leading strand more efficiently than those with a gap on the lagging strand or Holliday junction DNA. RecQ unwinds the template duplex ahead of the fork, and then the lagging strand is unwound. Consequently, this process generates a single-stranded DNA (ssDNA) gap on the lagging strand adjacent to a replication fork. These results suggest that RecQ functions to generate an initiating signal that can recruit RecA for SOS induction and recombination at stalled replication forks, which are required for the cell cycle checkpoint and resumption of DNA replication. PMID- 15289461 TI - DNA trajectory in the Gal repressosome. AB - The Gal repressosome is a higher-order nucleoprotein complex that represses transcription of the gal operon in Escherichia coli. During the repressosome assembly, a DNA loop is formed by the interaction of two GalR dimers, bound to two spatially separated operators, OE and OI, flanking the gal promoters. Structure-based genetic analysis indicated that GalR homodimers interact directly and form a V-shaped stacked tetramer in repressosome, further stabilized by HU binding to an architecturally critical position on the DNA. In this scheme of GalR tetramerization, the alignment of the operators in the DNA loop could be in either parallel (PL) or antiparallel (AL) mode. As each mode can have two alternative geometries differing in the mutual stacking of the OE- and OI-bound GalR dimers, it is possible to have four different DNA trajectories in the repressosome. Feasibilities of these trajectories were tested by in vitro transcription repression assays, first by isolating GalR mutants with altered operator specificity and then by constructing four different potential loops with mutant GalR heterodimers bound to specifically designed hybrid operators in such a way as to give rise to only one of the four putative trajectories. Results show that OE and OI adopt a mutual antiparallel orientation in an under-twisted DNA loop, consistent with the energetically optimal structural model. In this structure the center of the HU-binding site is located at the apex of the DNA loop. The approach reported here can be used to distinguish between otherwise indistinguishable DNA trajectories in complex nucleoprotein machines. PMID- 15289462 TI - A role for centromere pairing in meiotic chromosome segregation. AB - In meiosis I, exchanges provide a connection between homologous chromosome pairs that facilitates their proper attachment to the meiotic spindle. In many eukaryotes, homologous chromosomes that fail to become linked by exchanges exhibit elevated levels of meiotic errors, but they do not segregate randomly, demonstrating that mechanisms beyond exchange can promote proper meiosis I segregation. The experiments described here demonstrate the existence of a meiotic centromere pairing mechanism in budding yeast. This centromere pairing mediates the meiosis I bipolar spindle attachment of nonexchange chromosome pairs and likely plays the same role for all homologous chromosome pairs. PMID- 15289463 TI - The X-ray structure of the papillomavirus helicase in complex with its molecular matchmaker E2. AB - DNA replication of the papillomaviruses is specified by cooperative binding of two proteins to the ori site: the enhancer E2 and the viral initiator E1, a distant member of the AAA+ family of proteins. Formation of this prereplication complex is an essential step toward the construction of a functional, multimeric E1 helicase and DNA melting. To understand how E2 interacts with E1 to regulate this process, we have solved the X-ray structure of a complex containing the HPV18 E2 activation domain bound to the helicase domain of E1. Modeling the monomers of E1 to a hexameric helicase shows that E2 blocks hexamerization of E1 by shielding a region of the E1 oligomerization surface and stabilizing a conformation of E1 that is incompatible with ATP binding. Further biochemical experiments and structural analysis show that ATP is an allosteric effector of the dissociation of E2 from E1. Our data provide the first molecular insights into how a protein can regulate the assembly of an oligomeric AAA+ complex and explain at a structural level why E2, after playing a matchmaker role by guiding E1 to the DNA, must dissociate for subsequent steps of initiation to occur. Building on previously proposed ideas, we discuss how our data advance current models for the conversion of E1 in the prereplication complex to a hexameric helicase assembly. PMID- 15289465 TI - Regulation of vascular tone: the fat connection. PMID- 15289464 TI - Effect of a C/EBP gene replacement on mitochondrial biogenesis in fat cells. AB - CCAAT/enhancer-binding proteins, C/EBPalpha and C/EBPbeta, are required for fat cell differentiation and maturation. Previous studies showed that replacement of C/EBPalpha with C/EBPbeta, generating the beta/beta alleles in the mouse genome, prevents lipid accumulation in white adipose tissue (WAT). In this study, beta/beta mice lived longer and had higher energy expenditure than their control littermates due to increased WAT energy oxidation. The WAT of beta/beta mice was enriched with metabolically active, thermogenic mitochondria known for energy burning. The beta/beta allele exerted its effect through the elevated expression of the G protein alpha stimulatory subunit (Galphas) in WAT. Galphas, when overexpressed in fat-laden 3T3-L1 cells, stimulated mitochondrial biogenesis similar to that seen in the WAT of beta/beta mice, and effectively diminished the stored lipid pool. PMID- 15289466 TI - Hypertensive response to acute stress is attenuated in interleukin-6 knockout mice. AB - This study tested the hypothesis that the inflammatory cytokine, interleukin-6, contributes to the hypertensive response to acute psychosocial stress, caused by switching male mice to a cage previously occupied by a different male mouse. Male C57BL6 (WT) and interleukin-6 (IL-6) knockout (KO) mice were implanted with biotelemetry devices to monitor mean arterial pressure, heart rate, and motor activity in the unrestrained state. Baseline mean arterial pressure was 98+/-1 and 103+/-1 for WT and IL-6 KO mice. Cage switch increased mean arterial pressure by 42+/-2 mm Hg in WT mice, but this was blunted significantly in KO mice (31+/-3 mm Hg peak increase). Area under the curve for the first 90 minutes also was significantly less. Heart rate and motor activity increased similarly, and there also were no differences in the increases in plasma renin activity or plasma norepinephrine concentration between WT and KO mice. Thus, the acute hypertensive response to psychosocial stress depends significantly on IL-6, and the effect appears to be specific for blood pressure rather than to a global impairment in the response to stress. However, because perfusion of the isolated mesenteric bed with phenylephrine and chronic infusion of angiotensin II caused similar responses in WT and IL-6 KO mice, it is clear that future studies are needed to determine to what extent the acute blood pressure effect of IL-6 is stress specific. PMID- 15289468 TI - Differences in circadian pattern of ambulatory pulse pressure between healthy and complicated pregnancies. AB - With the use of ambulatory monitoring, a circadian blood pressure pattern has been shown to characterize normotensive as well as hypertensive pregnant women. However, the potential differences between healthy and complicated pregnancies in pulse pressure, an independent marker of cardiovascular risk in the general population, have not yet been investigated. We analyzed 2523 blood pressure series sampled for 48 hours once every 4 weeks from the first obstetric visit until delivery in 245 women with uncomplicated pregnancies, 140 with gestational hypertension, and 49 who developed preeclampsia. Compared with uncomplicated pregnancies, a statistically significant elevation in the 24-hour mean of pulse pressure is found in complicated pregnancies in all trimesters (P<0.001). Results further indicate similar 24-hour mean of pulse pressure between gestational hypertension and preeclampsia in the first trimester of pregnancy (P=0.158). The increase in pulse pressure among women who developed preeclampsia compared with women with gestational hypertension, although small, was statistically significant in the second trimester (1.4 mm Hg; P=0.010) and, to a larger extent, in the third trimester of pregnancy (1.8 mm Hg; P<0.001). The differences in pulse pressure between healthy and complicated pregnancies, observed already in the first trimester of gestation, are found when systolic and diastolic blood pressure for women with a later diagnosis of gestational hypertension or preeclampsia are within the accepted range of normotension. Moreover, ambulatory pulse pressure provides higher sensitivity than clinic measurements for the diagnosis of hypertension in pregnancy. PMID- 15289467 TI - Intracellular and extracellular angiotensin II enhance the L-type calcium current in the failing heart. AB - The influence of intracellular and extracellular angiotensin II (Ang II) on the L type calcium current of cardiomyocytes isolated from cardiomyopathic hamsters was investigated. The results indicated that Ang II (10(-8) mmol/L), added to the bath, increased the peak inward calcium current (I(Ca)) density by 37+/-3.4% (P<0.05), an effect that depends on the activation of protein kinase C. Intracellular administration of the same dose of Ang II (10(-8) mmol/L) also elicited an increase of peak I(Ca) density but enhanced the rate of I(Ca) inactivation, an effect not seen with extracellular Ang II. Moreover, in control animals, no change in the rate of I(Ca) inactivation was seen with intracellular Ang II. Thapsigargin (1 micromol/L), a potent inhibitor of sarcoplasmic reticulum (SR) ATPase, which depletes the SR, decreased the rate of I(Ca) inactivation elicited by intracellular Ang II, although the cytoplasmic calcium concentration was highly buffered with 10 mmol/L EGTA. These findings might indicate that intracellular Ang II releases calcium from the SR and inactivates I(Ca). The effect of intracellular Ang II on peak I(Ca) was not altered by extracellular losartan (10(-7) mmol/L), supporting the notion that the peptide acted intracellularly. Other studies showed that intracellular Ang I administration (10(-8) mmol/L) enhanced the peak I(Ca) density and the rate of I(Ca) inactivation, an effect that was reduced by intracellular enalaprilat (10(-8) mmol/L). Moreover, intracellular enalaprilat by itself reduced the peak I(Ca) density. These observations might indicate that endogenous Ang II is contributing to I(Ca) modulation in the failing heart. PMID- 15289469 TI - Treatment of dilated cardiomyopathy with electroporation of hepatocyte growth factor gene into skeletal muscle. AB - Hepatocyte growth factor (HGF) is a potent angiogenic and antifibrotic factor. Cardioprotective effects of HGF for idiopathic dilated cardiomyopathy were examined in hamsters with electroporation of plasmid DNA into skeletal muscle. We used hamster skeletal muscle as a protein producer of HGF gene. A plasmid vector encoding HGF (HGF group, n=12) or empty plasmid (placebo group, n=12) was transferred with in vivo electroporation into tibialis anterior muscles of hamsters with inherited dilated cardiomyopathy (TO-2 strain). The HGF group had greater serum HGF levels (21.6+/-2.2 versus 0.11+/-0.07 ng/mL, P<0.05), higher left ventricular ejection fraction (47.9+/-9.4% versus 28.8+/-11.2%, P<0.05), and greater wall thickening (31.6+/-6.3% versus 19.7+/-6.1%, P<0.05) when compared with the placebo group. The HGF group had smaller areas of ventricular fibrosis (11.8+/-3.4% versus 17.1+/-3.5%, P<0.05) and lower hydroxyproline content (3.7+/ 0.7 versus 5.1+/-0.9 micromol/g, P<0.05) than did the placebo group. The HGF group also had higher capillary density (1885+/-232 versus 1447+/-182 vessel/mm(2), P<0.05) and higher matrix metalloprotease-1 activity (13.1+/-3.5 versus 8.1+/-3.6 microg/collagen degraded per hour per gram tissue, P<0.05) than did the placebo group. Exogenous HGF might improve the deleterious changes in myocardial function and structure in the hamster with dilated cardiomyopathy. Systemic delivery of gene products with in vivo electroporation into skeletal muscle seemed to be an alternative means of direct gene delivery. PMID- 15289470 TI - The complete genome and proteome of Mycoplasma mobile. AB - Although often considered "minimal" organisms, mycoplasmas show a wide range of diversity with respect to host environment, phenotypic traits, and pathogenicity. Here we report the complete genomic sequence and proteogenomic map for the piscine mycoplasma Mycoplasma mobile, noted for its robust gliding motility. For the first time, proteomic data are used in the primary annotation of a new genome, providing validation of expression for many of the predicted proteins. Several novel features were discovered including a long repeating unit of DNA of approximately 2435 bp present in five complete copies that are shown to code for nearly identical yet uniquely expressed proteins. M. mobile has among the lowest DNA GC contents (24.9%) and most reduced set of tRNAs of any organism yet reported (28). Numerous instances of tandem duplication as well as lateral gene transfer are evident in the genome. The multiple available complete genome sequences for other motile and immotile mycoplasmas enabled us to use comparative genomic and phylogenetic methods to suggest several candidate genes that might be involved in motility. The results of these analyses leave open the possibility that gliding motility might have arisen independently more than once in the mycoplasma lineage. PMID- 15289471 TI - Regional patterns of gene expression in human and chimpanzee brains. AB - We have analyzed gene expression in various brain regions of humans and chimpanzees. Within both human and chimpanzee individuals, the transcriptomes of the cerebral cortex are very similar to each other and differ more between individuals than among regions within an individual. In contrast, the transcriptomes of the cerebral cortex, the caudate nucleus, and the cerebellum differ substantially from each other. Between humans and chimpanzees, 10% of genes differ in their expression in at least one region of the brain. The majority of these expression differences are shared among all brain regions. Whereas genes encoding proteins involved in signal transduction and cell differentiation differ significantly between brain regions within individuals, no such pattern is seen between the species. However, a subset of genes that show expression differences between humans and chimpanzees are distributed nonrandomly across the genome. Furthermore, genes that show an elevated expression level in humans are statistically significantly enriched in regions that are recently duplicated in humans. PMID- 15289472 TI - Segmental phylogenetic relationships of inbred mouse strains revealed by fine scale analysis of sequence variation across 4.6 mb of mouse genome. AB - High-density SNP screening of panels of inbred mouse strains has been proposed as a method to accelerate the identification of genes associated with complex biomedical phenotypes. To evaluate the potential of these studies, a more detailed understanding of the fine structure of sequence variation across inbred mouse strains is needed. Here, we use high-density oligonucleotide arrays to discover an extremely dense set of SNPs in 13 classical and two wild-derived inbred strains in five genomic intervals totaling 4.6 Mb of DNA sequence, and then analyze the segmental haplotype structure defined by these high-density SNPs. This analysis reveals segments ranging from 12 to 608 kb in length within which the inbred strains have a simple and distinct phylogenetic relationship with typically two or three clades accounting for the 13 classical strains examined. The phylogenetic relationships among strains change abruptly and unpredictably from segment to segment, and are distinct in each of the five genomic regions examined. The data suggest that at least 12 strains would need to be resequenced for exhaustive SNP discovery in every region of the mouse genome, that approximately 97% of the variation among inbred strains is ancestral (between clades) and approximately 3% private (within clades), and provides critical insights into the proposed use of panels of inbred strains to identify genes underlying quantitative trait loci. PMID- 15289473 TI - Genetic divergence of the rhesus macaque major histocompatibility complex. AB - The major histocompatibility complex (MHC) is comprised of the class I, class II, and class III regions, including the MHC class I and class II genes that play a primary role in the immune response and serve as an important model in studies of primate evolution. Although nonhuman primates contribute significantly to comparative human studies, relatively little is known about the genetic diversity and genomics underlying nonhuman primate immunity. To address this issue, we sequenced a complete rhesus macaque MHC spanning over 5.3 Mb, and obtained an additional 2.3 Mb from a second haplotype, including class II and portions of class I and class III. A major expansion of from six class I genes in humans to as many as 22 active MHC class I genes in rhesus and levels of sequence divergence some 10-fold higher than a similar human comparison were found, averaging from 2% to 6% throughout extended portions of class I and class II. These data pose new interpretations of the evolutionary constraints operating between MHC diversity and T-cell selection by contrasting with models predicting an optimal number of antigen presenting genes. For the clinical model, these data and derivative genetic tools can be implemented in ongoing genetic and disease studies that involve the rhesus macaque. PMID- 15289475 TI - Comparative analysis of gene expression for convergent evolution of camera eye between octopus and human. AB - Although the camera eye of the octopus is very similar to that of humans, phylogenetic and embryological analyses have suggested that their camera eyes have been acquired independently. It has been known as a typical example of convergent evolution. To study the molecular basis of convergent evolution of camera eyes, we conducted a comparative analysis of gene expression in octopus and human camera eyes. We sequenced 16,432 ESTs of the octopus eye, leading to 1052 nonredundant genes that have matches in the protein database. Comparing these 1052 genes with 13,303 already-known ESTs of the human eye, 729 (69.3%) genes were commonly expressed between the human and octopus eyes. On the contrary, when we compared octopus eye ESTs with human connective tissue ESTs, the expression similarity was quite low. To trace the evolutionary changes that are potentially responsible for camera eye formation, we also compared octopus eye ESTs with the completed genome sequences of other organisms. We found that 1019 out of the 1052 genes had already existed at the common ancestor of bilateria, and 875 genes were conserved between humans and octopuses. It suggests that a larger number of conserved genes and their similar gene expression may be responsible for the convergent evolution of the camera eye. PMID- 15289474 TI - Evolution of eukaryotic transcription: insights from the genome of Giardia lamblia. AB - The Giardia lamblia genome sequencing project affords us a unique opportunity to conduct comparative analyses of core cellular systems between early and late diverging eukaryotes on a genome-wide scale. We report a survey to identify canonical transcription components in Giardia, focusing on RNA polymerase (RNAP) subunits and transcription-initiation factors. Our survey revealed that Giardia contains homologs to 21 of the 28 polypeptides comprising eukaryal RNAPI, RNAPII, and RNAPIII; six of the seven RNAP subunits without giardial homologs are polymerase specific. Components of only four of the 12 general transcription initiation factors have giardial homologs. Surprisingly, giardial TATA-binding protein (TBP) is highly divergent with respect to archaeal and higher eukaryotic TBPs, and a giardial homolog of transcription factor IIB was not identified. We conclude that Giardia represents a transition during the evolution of eukaryal transcription systems, exhibiting a relatively complete set of RNAP subunits and a rudimentary basal initiation apparatus for each transcription system. Most class-specific RNAP subunits and basal initiation factors appear to have evolved after the divergence of Giardia from the main eukaryotic line of descent. Consequently, Giardia is predicted to be unique in many aspects of transcription initiation with respect to paradigms derived from studies in crown eukaryotes. PMID- 15289476 TI - Stress-induced DNA duplex destabilization (SIDD) in the E. coli genome: SIDD sites are closely associated with promoters. AB - We present the first analysis of stress-induced DNA duplex destabilization (SIDD) in a complete chromosome, the Escherichia coli K12 genome. We used a newly developed method to calculate the locations and extents of stress-induced destabilization to single-base resolution at superhelix density sigma = -0.06. We find that SIDD sites in this genome show a statistically highly significant tendency to avoid coding regions. And among intergenic regions, those that either contain documented promoters or occur between divergently transcribing coding regions, and hence may be inferred to contain promoters, are associated with strong SIDD sites in a statistically highly significant manner. Intergenic regions located between convergently transcribing genes, which are inferred not to contain promoters, are not significantly enriched for destabilized sites. Statistical analysis shows that a strongly destabilized intergenic region has an 80% chance of containing a promoter, whereas an intergenic region that does not contain a strong SIDD site has only a 24% chance. We describe how these observations may illuminate specific mechanisms of regulation, and assist in the computational identification of promoter locations in prokaryotes. PMID- 15289477 TI - Early genetic mechanisms underlying the inhibitory effects of endostatin and fumagillin on human endothelial cells. AB - A tumor needs to initiate angiogenesis in order to develop its own blood supply, to grow, to invade, and to spread. Angiogenesis, under normal conditions, is a tightly regulated balance between endogenous pro- and antiangiogenic factors. In this study, we investigated, by microarray analysis, the effects of two known antiangiogenic agents (endostatin and fumagillin) on the gene expression profiles of human umbilical vein endothelial cells (HUVEC) in order to elucidate pathways common to the effects of these agents. We observed a majority of gene expression changes within 1 and 2 h of treatment. The genes demonstrating these early expression changes are involved in cell proliferation, gene transcription, and a number have unknown functions. We selected four genes (DOC1, KLF4, TC-1, ID1) from the microarray profile that showed a similar pattern of expression for both of the antiangiogenic agents we tested. We then used small interfering RNAs (siRNA) in an attempt to better understand the role of these selected genes in the inhibitory activity of these agents. Because the gene expression changes occurred within 1 and 2 h of treatment, these genes might be involved in the initial pathways of angiogenesis inhibition. PMID- 15289478 TI - Fosmid-based physical mapping of the Histoplasma capsulatum genome. AB - A fosmid library representing 10-fold coverage of the Histoplasma capsulatum G217B genome was used to construct a restriction-based physical map. The data obtained from three restriction endonuclease fingerprints, generated from each clone using BamHI, HindIII, and PstI endonucleases, were combined and used in FPC for automatic and manual contig assembly builds. Concomitantly, a whole-genome shotgun (WGS) sequencing of paired-end reads from plasmids and fosmids were assembled with PCAP, providing a predicted genome size of up to 43.5 Mbp and 17% repetitive DNA. Fosmid paired-end sequences in the WGS assembly provide anchoring information to the physical map and result in joining of existing physical map contigs into 84 clusters containing 9551 fosmid clones. Here, we detail mapping the Histoplasma capsulatum genome comprehensively in fosmids, resulting in an efficient paradigm for de novo sequencing that uses a map-assisted whole genome shotgun approach. PMID- 15289479 TI - Indel-based evolutionary distance and mouse-human divergence. AB - We propose a method for estimating the evolutionary distance between DNA sequences in terms of insertions and deletions (indels), defined as the per site number of indels accumulated in the course of divergence of the two sequences. We derive a maximal likelihood estimate of this distance from differences between lengths of orthologous introns or other segments of sequences delimited by conservative markers. When indels accumulate, lengths of orthologous introns diverge only slightly slower than linearly, because long indels occur with substantial frequencies. Thus, saturation is not a major obstacle for estimating indel-based evolutionary distance. For introns of medium lengths, our method recovers the known evolutionary distance between rat and mouse, 0.014 indels per site, with good precision. We estimate that mouse-human divergence exceeds rat mouse divergence by a factor of 4, so that mouse-human evolutionary distance in terms of selectively neutral indels is 0.056. Because in mammals, indels are approximately 14 times less frequent than nucleotide substitutions, mouse-human evolutionary distance in terms of selectively neutral substitutions is approximately 0.8. PMID- 15289480 TI - A non-EST-based method for exon-skipping prediction. AB - It is estimated that between 35% and 74% of all human genes can undergo alternative splicing. Currently, the most efficient methods for large-scale detection of alternative splicing use expressed sequence tags (ESTs) or microarray analysis. As these methods merely sample the transcriptome, splice variants that do not appear in deeply sampled tissues have a low probability of being detected. We present a new method by which we can predict that an internal exon is skipped (namely whether it is a cassette-exon) merely based on its naked genomic sequence and on the sequence of its mouse ortholog. No other data, such as ESTs, are required for the prediction. Using our method, which was experimentally validated, we detected hundreds of novel splice variants that were not detectable using ESTs. We show that a substantial fraction of the splice variants in the human genome could not be identified through current human EST or cDNA data. PMID- 15289481 TI - Optimal haplotype block-free selection of tagging SNPs for genome-wide association studies. AB - It is widely hoped that the study of sequence variation in the human genome will provide a means of elucidating the genetic component of complex diseases and variable drug responses. A major stumbling block to the successful design and execution of genome-wide disease association studies using single-nucleotide polymorphisms (SNPs) and linkage disequilibrium is the enormous number of SNPs in the human genome. This results in unacceptably high costs for exhaustive genotyping and presents a challenging problem of statistical inference. Here, we present a new method for optimally selecting minimum informative subsets of SNPs, also known as "tagging" SNPs, that is efficient for genome-wide selection. We contrast this method to published methods including haplotype block tagging, that is, grouping SNPs into segments of low haplotype diversity and typing a subset of the SNPs that can discriminate all common haplotypes within the blocks. Because our method does not rely on a predefined haplotype block structure and makes use of the weaker correlations that occur across neighboring blocks, it can be effectively applied across chromosomal regions with both high and low local linkage disequilibrium. We show that the number of tagging SNPs selected is substantially smaller than previously reported using block-based approaches and that selecting tagging SNPs optimally can result in a two- to threefold savings over selecting random SNPs. PMID- 15289482 TI - The use of MPSS for whole-genome transcriptional analysis in Arabidopsis. AB - We have generated 36,991,173 17-base sequence "signatures" representing transcripts from the model plant Arabidopsis. These data were derived by massively parallel signature sequencing (MPSS) from 14 libraries and comprised 268,132 distinct sequences. Comparable data were also obtained with 20-base signatures. We developed a method for handling these data and for comparing these signatures to the annotated Arabidopsis genome. As part of this procedure, 858,019 potential or "genomic" signatures were extracted from the Arabidopsis genome and classified based on the position and orientation of the signatures relative to annotated genes. A comparison of genomic and expressed signatures matched 67,735 signatures predicted to be derived from distinct transcripts and expressed at significant levels. Expressed signatures were derived from the sense strand of at least 19,088 of 29,084 annotated genes. A comparison of the genomic and expression signatures demonstrated that approximately 7.7% of genomic signatures were underrepresented in the expression data. These genomic signatures contained one of 20 four-base words that were consistently associated with reduced MPSS abundances. More than 89% of the sum of the expressed signature abundances matched the Arabidopsis genome, and many of the unmatched signatures found in high abundances were predicted to match to previously uncharacterized transcripts. PMID- 15289483 TI - Elucidation of gene interaction networks through time-lagged correlation analysis of transcriptional data. AB - The photosynthetic cyanobacterium Synechocystis sp. strain PCC 6803 uses a complex genetic program to control its physiological response to alternating light conditions. To study this regulatory program time-series experiments were conducted by exposing Synechocystis sp. to serial perturbations in light intensity. In each experiment whole-genome DNA microarrays were used to monitor gene transcription in 20-min intervals over 8- and 16-h periods. The data was analyzed using time-lagged correlation analysis, which identifies genetic interaction networks by constructing correlations between time-shifted transcription profiles with different levels of statistical confidence. These networks allow inference of putative cause-effect relationships among the organism's genes. Using light intensity as our initial input signal, we identified six groups of genes whose time-lagged profiles possessed significant correlation, or anti-correlation, with the light intensity. We expanded this network by using the average profile from each group of genes as a seed, and searching for other genes whose time-lagged profiles possessed significant correlation, or anti-correlation, with the group's average profile. The final network comprised 50 different groups containing 259 genes. Several of these gene groups possess known light-stimulated gene clusters, such as Synechocystis sp. photosystems I and II and carbon dioxide fixation pathways, while others represent novel findings in this work. PMID- 15289484 TI - Large-scale validation of single nucleotide polymorphisms in gene regions. AB - Genome-wide association studies using large numbers of bi-allelic single nucleotide polymorphisms (SNPs) have been proposed as a potentially powerful method for identifying genes involved in common diseases. To assemble a SNP collection appropriate for large-scale association, we designed assays for 226,099 publicly available SNPs located primarily within known and predicted gene regions. Allele frequencies were estimated in a sample of 92 CEPH Caucasians using chip-based MALDI-TOF mass spectrometry with pooled DNA. Of the 204,200 designed assays that were functional, 125,799 SNPs were determined to be polymorphic (minor allele frequency > 0.02), of which 101,729 map uniquely to the human genome. Many of the commonly available RefSNP annotations were predictive of polymorphic status and could be used to improve the selection of SNPs from the public domain for genetic research. The set of uniquely mapping, polymorphic SNPs is located within 10 kb of 66% of known and predicted genes annotated in LocusLink, which could prove useful for large-scale disease association studies. PMID- 15289485 TI - cpnDB: a chaperonin sequence database. AB - Type I chaperonins are molecular chaperones present in virtually all bacteria, some archaea and the plastids and mitochondria of eukaryotes. Sequences of cpn60 genes, encoding 60-kDa chaperonin protein subunits (CPN60, also known as GroEL or HSP60), are useful for phylogenetic studies and as targets for detection and identification of organisms. Conveniently, a 549-567-bp segment of the cpn60 coding region can be amplified with universal PCR primers. Here, we introduce cpnDB, a curated collection of cpn60 sequence data collected from public databases or generated by a network of collaborators exploiting the cpn60 target in clinical, phylogenetic, and microbial ecology studies. The growing database currently contains approximately 2000 records covering over 240 genera of bacteria, eukaryotes, and archaea. The database also contains over 60 sequences for the archaeal Type II chaperonin (thermosome, a homolog of eukaryotic cytoplasmic chaperonin) from 19 archaeal genera. As the largest curated collection of sequences available for a protein-encoding gene, cpnDB provides a resource for researchers interested in exploiting the power of cpn60 as a diagnostic or as a target for phylogenetic or microbial ecology studies, as well as those interested in broader subjects such as lateral gene transfer and codon usage. We built cpnDB from open source tools and it is available at http://cpndb.cbr.nrc.ca. PMID- 15289486 TI - Individual patient data-based meta-analysis of patients aged 16 to 60 years with core binding factor acute myeloid leukemia: a survey of the German Acute Myeloid Leukemia Intergroup. AB - PURPOSE: To evaluate prognostic factors for relapse-free survival (RFS) and overall survival (OS) and to assess the impact of different postremission therapies in adult patients with core binding factor (CBF) acute myeloid leukemias (AML). PATIENTS AND METHODS: Individual patient data-based meta analysis was performed on 392 adults (median age, 42 years; range, 16 to 60 years) with CBF AML (t(8;21), n = 191; inv(16), n = 201) treated between 1993 and 2002 in prospective German AML treatment trials. RESULTS: RFS was 60% and 58% and OS was 65% and 74% in the t(8;21) and inv(16) groups after 3 years, respectively. For postremission therapy, intention-to-treat analysis revealed no difference between intensive chemotherapy and autologous transplantation in the t(8;21) group and between chemotherapy, autologous, and allogeneic transplantation in the inv(16) group. In the t(8;21) group, significant prognostic variables for longer RFS and OS were lower WBC and higher platelet counts; loss of the Y chromosome in male patients was prognostic for shorter OS. In the inv(16) group, trisomy 22 was a significant prognostic variable for longer RFS. For patients who experienced relapse, second complete remission rate was significantly lower in patients with t(8;21), resulting in a significantly inferior survival duration after relapse compared with patients with inv(16). CONCLUSION: We provide novel prognostic factors for CBF AML and show that patients with t(8;21) who experience relapse have an inferior survival duration. PMID- 15289487 TI - Chemotherapy sensitivity and resistance assays: a systematic review. AB - PURPOSE: This systematic review evaluates evidence comparing therapy guided by chemotherapy sensitivity and resistance assays with empiric chemotherapy, emphasizing survival outcomes. METHODS: Prospective studies were sought comparing patients treated contemporaneously by assay-guided chemotherapy and empiric therapy. An initial MEDLINE search and a search performed by a Working Group of the American Society of Clinical Oncology were reviewed with attention to prespecified study selection criteria. RESULTS: This review identified 10 studies meeting selection criteria, plus one retrospective study, using seven different assays. Only two studies randomly assigned patients to assay-guided treatment or empiric treatment. Five of nine nonrandomized studies found significantly higher response rates for patients who received assay-guided therapy compared with those treated empirically. One of the two randomized trials found a significantly higher response rate in the assay-guided group. Four additional studies found response rates favoring assay-guided therapy, but comparisons did not achieve statistical significance. Two nonrandomized studies found overall survival to be significantly improved with assay-guided therapy. One randomized study used a cross-over design that made it difficult to determine whether survival differed between groups, while the other randomized trial found no difference in survival. Six studies provided no comparison of groups on baseline patient characteristics. Only one study reported adverse events data. CONCLUSION: While higher response rates for assay-guided therapy have been observed, differences may be attributable to bias or confounding. Little evidence on survival is available. These results do not establish the relative effectiveness of assay-guided treatment and empiric treatment. Randomized trials are needed. PMID- 15289488 TI - American Society of Clinical Oncology Technology Assessment: chemotherapy sensitivity and resistance assays. AB - PURPOSE: To develop a technology assessment of chemotherapy sensitivity and resistance assays in order to define the role of these tests in routine oncology practice. METHODS: The American Society of Clinical Oncology (ASCO) established a Working Group to develop the technology assessment. The Working Group collaborated with the Blue Cross and Blue Shield Association (BCBSA) Technology Evaluation Center. The Working Group developed independent criteria for selecting articles for inclusion in the ASCO assessment, and developed a structured data abstraction tool to facilitate review of selected manuscripts. One Working Group member and an ASCO staff member independently reviewed the 1,139 abstracts identified by the BCBSA comprehensive literature search, and by an updated literature search performed by ASCO using the BCBSA search strategy (1966 to January 2004). Of the 12 articles included in this technology assessment, eight were identified by the original BCBSA systematic review, one was provided by industry, and three were identified by the ASCO updated literature review. RESULTS: Review of the literature does not identify any CSRAs for which the evidence base is sufficient to support use in oncology practice. RECOMMENDATIONS: The use of chemotherapy sensitivity and resistance assays to select chemotherapeutic agents for individual patients is not recommended outside of the clinical trial setting. Oncologists should make chemotherapy treatment recommendations on the basis of published reports of clinical trials and a patient's health status and treatment preferences. Because the in vitro analytic strategy has potential importance, participation in clinical trials evaluating these technologies remains a priority. PMID- 15289489 TI - Crossing the quality chasm in breast cancer care. PMID- 15289490 TI - PC-SPES: hope or hype? PMID- 15289491 TI - Compliance with consensus recommendations for systemic therapy is associated with improved survival of women with node-negative breast cancer. AB - PURPOSE: The impact of consensus recommendations for systemic therapy on outcome of disease is unclear. We evaluated if compliance with guidelines for systemic adjuvant treatment is associated with improved survival of women with node negative breast cancer. PATIENTS AND METHODS: The study population included women diagnosed with invasive node-negative breast cancer in Quebec, Canada, in 1988 to 1989, 1991 to 1992, and 1993 to 1994. Information was collected by chart review, linkage with administrative databases, and queries to attending physicians. Guidelines from the 1992 St Gallen conference were used as standard of care. Survival was estimated by Kaplan-Meier and Cox proportional hazards analyses. RESULTS: Among 1,541 women, 358 died before December 1999. Median follow-up was 6.8 years. Seven-year event-free and overall survivals were 66% and 81%, respectively. Survival was 88%, 84%, and 74% in women at minimal, moderate, or high risk of recurrence. Virtually all women at minimal risk were treated according to the consensus (98.4% of 370). In comparison, adjusted hazard ratios of death were 1.0 (95% CI, 0.6 to 1.7) and 2.3 (95% CI, 1.3 to 4.0) among women at moderate risk treated according to the consensus or not, respectively. Among women at high risk, adjusted hazard ratios of death were 2.0 (95% CI, 1.4 to 2.8) and 2.7 (95% CI, 1.9 to 3.9), respectively. Both risk category (P <.0005) and compliance with guidelines (P <.0005) were independent significant predictors of survival. CONCLUSION: Treatment according to consensus recommendations is associated with improved survival of women with breast cancer in the community. Promoting the adoption of guidelines for treatment is an effective strategy for disease control. PMID- 15289492 TI - Prospective, multicenter, randomized phase II trial of the herbal supplement, PC SPES, and diethylstilbestrol in patients with androgen-independent prostate cancer. AB - PURPOSE: To evaluate the herbal combination, PC-SPES, and diethylstilbestrol (DES) in patients with androgen independent prostate cancer (AIPC). PATIENTS AND METHODS: A randomized phase II study was conducted with cross-over design. Patients were randomly assigned to receive either three PC-SPES capsules orally three times a day or DES 3 mg orally once a day. Prophylactic warfarin was administered. At clinical or prostate-specific antigen progression, patients received the other therapy. The study closed prematurely after PC-SPES was withdrawn from the market. Chemical analyses were performed on multiple lots of PC-SPES. RESULTS: Ninety patients were enrolled, of whom 85 were assessable for response. Prostate-specific antigen declines > or = 50% were noted in 40% (95% CI, 25% to 56%) with PC-SPES, and 24% (95% CI, 12% to 39%) with DES. Median response duration was not reached with PC-SPES, and was 3.8 months with DES. Median time to progression for randomly assigned patients was 5.5 months for PC SPES and 2.9 months for DES. Common toxicities included mild fatigue, gynecomastia, and mastodynia. Five thromboembolic events occurred (one PC-SPES, four DES). Responses in the cross-over phase were inconclusive. Four lots of PC SPES had measurable quantities of DES, ranging from 0.01% to 3.1% of the dose used in the DES arm. Ethinyl estradiol was also detected in PC-SPES lots. CONCLUSION: PC-SPES and DES demonstrate activity in AIPC and are well tolerated. However, the synthetic estrogens, DES and ethinyl estradiol, were detected in various lots of PC-SPES, including those used in this trial. Clinical trials that utilize herbal therapies must account for issues of purity and consistency. PMID- 15289493 TI - Yeast cell death during DNA damage arrest is independent of caspase or reactive oxygen species. AB - CDC13 encodes a telomere-binding protein that prevents degradation of telomeres. cdc13-1 yeast grown at the nonpermissive temperature undergo G2/M arrest, progressive chromosome instability, and subsequent cell death. Recently, it has been suggested that cell death in the cdc13-1 mutant is an active process characterized by phenotypic hallmarks of apoptosis and caspase activation. In this work, we show that cell death triggered by cdc13-1 is independent of the yeast metacaspase Yca1p and reactive oxygen species but related to cell cycle arrest per se. Inactivating YCA1 or depleting reactive oxygen species does not increase viability of cdc13-1 cells. In turn, caspase activation does not precede cell death in the cdc13-1 mutant. Yca1p activity assayed by cell binding of mammalian caspase inhibitors is confounded by artifactual labeling of dead yeast cells, which nonspecifically bind fluorochromes. We speculate that during a prolonged cell cycle arrest, cdc13-1 cells reach a critical size and die by cell lysis. PMID- 15289494 TI - Active Rho is localized to podosomes induced by oncogenic Src and is required for their assembly and function. AB - Transformation of fibroblasts by oncogenic Src causes disruption of actin stress fibers and formation of invasive adhesions called podosomes. Because the small GTPase Rho stimulates stress fiber formation, Rho inactivation by Src has been thought to be necessary for stress fiber disruption. However, we show here that Rho[GTP] levels do not decrease after transformation by activated Src. Inactivation of Rho in Src-transformed fibroblasts by dominant negative RhoA or the Rho-specific inhibitor C3 exoenzyme disrupted podosome structure as judged by localization of podosome components F-actin, cortactin, and Fish. Inhibition of Rho strongly inhibited Src-induced proteolytic degradation of the extracellular matrix. Furthermore, development of an in situ Rho[GTP] affinity assay allowed us to detect endogenous Rho[GTP] at podosomes, where it colocalized with F-actin, cortactin, and Fish. Therefore, Rho is not globally inactivated in Src transformed fibroblasts, but is necessary for the assembly and function of structures implicated in tumor cell invasion. PMID- 15289495 TI - Beta-catenin is required for endothelial-mesenchymal transformation during heart cushion development in the mouse. AB - During heart development endocardial cells within the atrio-ventricular (AV) region undergo TGFbeta-dependent epithelial-mesenchymal transformation (EMT) and invade the underlying cardiac jelly. This process gives rise to the endocardial cushions from which AV valves and part of the septum originate. In this paper we show that in mouse embryos and in AV explants TGFbeta induction of endocardial EMT is strongly inhibited in mice deficient for endothelial beta-catenin, leading to a lack of heart cushion formation. Using a Wnt-signaling reporter mouse strain, we demonstrated in vivo and ex vivo that EMT in heart cushion is accompanied by activation of beta-catenin/TCF/Lef transcriptional activity. In cultured endothelial cells, TGFbeta2 induces alpha-smooth muscle actin (alphaSMA) expression. This process was strongly reduced in beta-catenin null cells, although TGFbeta2 induced smad phosphorylation was unchanged. These data demonstrate an involvement of beta-catenin/TCF/Lef transcriptional activity in heart cushion formation, and suggest an interaction between TGFbeta and Wnt signaling pathways in the induction of endothelial-mesenchymal transformation. PMID- 15289496 TI - NFkappaB activation by Fas is mediated through FADD, caspase-8, and RIP and is inhibited by FLIP. AB - Fas (APO-1/CD95) is the prototypic death receptor, and the molecular mechanisms of Fas-induced apoptosis are comparably well understood. Here, we show that Fas activates NFkappaB via a pathway involving RIP, FADD, and caspase-8. Remarkably, the enzymatic activity of the latter was dispensable for Fas-induced NFkappaB signaling pointing to a scaffolding-related function of caspase-8 in nonapoptotic Fas signaling. NFkappaB was activated by overexpressed FLIPL and FLIPS in a cell type-specific manner. However, in the context of Fas signaling both isoforms blocked FasL-induced NFkappaB activation. Moreover, down-regulation of both endogenous FLIP isoforms or of endogenous FLIPL alone was sufficient to enhance FasL-induced expression of the NFkappaB target gene IL8. As NFkappaB signaling is inhibited during apoptosis, FasL-induced NFkappaB activation was most prominent in cells that were protected by Bcl2 expression or caspase inhibitors and expressed no or minute amounts of FLIP. Thus, protection against Fas-induced apoptosis in a FLIP-independent manner converted a proapoptotic Fas signal into an inflammatory NFkappaB-related response. PMID- 15289497 TI - Differential transactivation of sphingosine-1-phosphate receptors modulates NGF induced neurite extension. AB - The process of neurite extension after activation of the TrkA tyrosine kinase receptor by nerve growth factor (NGF) involves complex signaling pathways. Stimulation of sphingosine kinase 1 (SphK1), the enzyme that phosphorylates sphingosine to form sphingosine-1-phosphate (S1P), is part of the functional TrkA signaling repertoire. In this paper, we report that in PC12 cells and dorsal root ganglion neurons, NGF translocates SphK1 to the plasma membrane and differentially activates the S1P receptors S1P1 and S1P2 in a SphK1-dependent manner, as determined with specific inhibitors and small interfering RNA targeted to SphK1. NGF-induced neurite extension was suppressed by down-regulation of S1P1 expression with antisense RNA. Conversely, when overexpressed in PC12 cells, transactivation of S1P1 by NGF markedly enhanced neurite extension and stimulation of the small GTPase Rac, important for the cytoskeletal changes required for neurite extension. Concomitantly, differentiation down-regulated expression of S1P2 whose activation would stimulate Rho and inhibit neurite extension. Thus, differential transactivation of S1P receptors by NGF regulates antagonistic signaling pathways that modulate neurite extension. PMID- 15289498 TI - Selective modulation of type 1 insulin-like growth factor receptor signaling and functions by beta1 integrins. AB - We show here that beta1 integrins selectively modulate insulin-like growth factor type I receptor (IGF-IR) signaling in response to IGF stimulation. The beta1A integrin forms a complex with the IGF-IR and insulin receptor substrate-1 (IRS 1); this complex does not promote IGF-I mediated cell adhesion to laminin (LN), although it does support IGF-mediated cell proliferation. In contrast, beta1C, an integrin cytoplasmic variant, increases cell adhesion to LN in response to IGF-I and its down-regulation by a ribozyme prevents IGF-mediated adhesion to LN. Moreover, beta1C completely prevents IGF-mediated cell proliferation and tumor growth by inhibiting IGF-IR auto-phosphorylation in response to IGF-I stimulation. Evidence is provided that the beta1 cytodomain plays an important role in mediating beta1 integrin association with either IRS-1 or Grb2-associated binder1 (Gab1)/SH2-containing protein-tyrosine phosphate 2 (Shp2), downstream effectors of IGF-IR: specifically, beta1A associates with IRS-1 and beta1C with Gab1/Shp2. This study unravels a novel mechanism mediated by the integrin cytoplasmic domain that differentially regulates cell adhesion to LN and cell proliferation in response to IGF. PMID- 15289499 TI - Switch from alphavbeta5 to alphavbeta6 integrin expression protects squamous cell carcinomas from anoikis. AB - Stratified squamous epithelia express the alphavbeta5 integrin, but in squamous cell carcinomas (SCCs) there is down-regulation of alphavbeta5 and up-regulation of alphavbeta6. To investigate the significance of this finding, we transduced an alphav-negative human SCC line with retroviral vectors encoding alphav integrins. alphavbeta5-expressing cells underwent suspension-induced apoptosis (anoikis), whereas alphav-negative cells and cells expressing alphavbeta6 did not. Resistance to anoikis correlated with PKB/Akt activation in suspension, but not with changes in PTEN or p110alpha PI3 kinase levels. Anoikis was induced in parental and alphavbeta6-expressing cells by inhibiting PI3 kinase. Conversely, activation of Akt or inhibition of caspases in alphavbeta5-expressing cells suppressed anoikis. Caspase inhibition resulted in increased phosphoAkt, placing caspase activation upstream of decreased Akt activation. Anoikis required the cytoplasmic domain of beta5 and was independent of the death receptor pathway. These results suggest that down-regulation of alphavbeta5 through up-regulation of alphavbeta6 may protect SCCs from anoikis by activating an Akt survival signal. PMID- 15289500 TI - A contribution of mouse dendritic cell-derived IL-2 for NK cell activation. AB - Dendritic cells (DCs) play a predominant role in activation of natural killer (NK) cells that exert their functions against pathogen-infected and tumor cells. Here, we used a murine model to investigate the molecular mechanisms responsible for this process. Two soluble molecules produced by bacterially activated myeloid DCs are required for optimal priming of NK cells. Type I interferons (IFNs) promote the cytotoxic functions of NK cells. IL-2 is necessary both in vitro and in vivo for the efficient production of IFNgamma, which has an important antimetastatic and antibacterial function. These findings provide new information about the mechanisms that mediate DC-NK cell interactions and define a novel and fundamental role for IL-2 in innate immunity. PMID- 15289501 TI - Mesothelin-specific CD8(+) T cell responses provide evidence of in vivo cross priming by antigen-presenting cells in vaccinated pancreatic cancer patients. AB - Tumor-specific CD8(+) T cells can potentially be activated by two distinct mechanisms of major histocompatibility complex class I-restricted antigen presentation as follows: direct presentation by tumor cells themselves or indirect presentation by professional antigen-presenting cells (APCs). However, controversy still exists as to whether indirect presentation (the cross-priming mechanism) can contribute to effective in vivo priming of tumor-specific CD8(+) T cells that are capable of eradicating cancer in patients. A clinical trial of vaccination with granulocyte macrophage-colony stimulating factor-transduced pancreatic cancer lines was designed to test whether cross-presentation by locally recruited APCs can activate pancreatic tumor-specific CD8(+) T cells. Previously, we reported postvaccination delayed-type hypersensitivity (DTH) responses to autologous tumor in 3 out of 14 treated patients. Mesothelin is an antigen demonstrated previously by gene expression profiling to be up-regulated in most pancreatic cancers. We report here the consistent induction of CD8(+) T cell responses to multiple HLA-A2, A3, and A24-restricted mesothelin epitopes exclusively in the three patients with vaccine-induced DTH responses. Importantly, neither of the vaccinating pancreatic cancer cell lines expressed HLA-A2, A3, or A24. These results provide the first direct evidence that CD8 T cell responses can be generated via cross-presentation by an immunotherapy approach designed to recruit APCs to the vaccination site. PMID- 15289503 TI - Prevention and treatment of experimental autoimmune encephalomyelitis by soluble CD83. AB - CD83 is up-regulated on the surface of dendritic cells (DCs) during maturation and has been widely used as a marker for mature DCs. Recently, we reported the recombinant expression of the extracellular immunoglobulin domain of human CD83 (hCD83ext). Using this soluble form of CD83, allogeneic as well as specific cytotoxic T lymphocyte proliferation could be blocked in vitro. Here we report the functional analysis of soluble CD83 in vivo, using murine experimental autoimmune encephalomyelitis (EAE) as a model. Strikingly, only three injections of soluble CD83 prevented the paralysis associated with EAE almost completely. In addition, even when the EAE was induced a second time, CD83-treated mice were protected, indicating a long-lasting suppressive effect. Furthermore, soluble CD83 strongly reduced the paralysis in different therapeutic settings. Most important, even when the treatment was delayed until the disease symptoms were fully established, soluble CD83 clearly reduced the paralyses. In addition, also when EAE was induced a second time, soluble CD83-treated animals showed reduced disease symptoms. Finally, hCD83ext treatment almost completely reduced leukocyte infiltration in the brain and in the spinal cord. In summary, this work strongly supports an immunosuppressive role of soluble CD83, thereby indicating its therapeutic potential in the regulation of immune disorders in vivo. PMID- 15289502 TI - Limited T cell receptor diversity of HCV-specific T cell responses is associated with CTL escape. AB - Escape mutations are believed to be important contributors to immune evasion by rapidly evolving viruses such as hepatitis C virus (HCV). We show that the majority of HCV-specific cytotoxic T lymphocyte (CTL) responses directed against viral epitopes that escaped immune recognition in HCV-infected chimpanzees displayed a reduced CDR3 amino acid diversity when compared with responses in which no CTL epitope variation was detected during chronic infection or with those associated with protective immunity. Decreased T cell receptor (TCR) CDR3 amino acid diversity in chronic infection could be detected long before the appearance of viral escape mutations in the plasma. In both chronic and resolved infection, identical T cell receptor clonotypes were present in liver and peripheral blood. These findings provide a deeper understanding of the evolution of CTL epitope variations in chronic viral infections and highlight the importance of the generation and maintenance of a diverse TCR repertoire directed against individual epitopes. PMID- 15289504 TI - Intracellular triggering of Fas aggregation and recruitment of apoptotic molecules into Fas-enriched rafts in selective tumor cell apoptosis. AB - We have discovered a new and specific cell-killing mechanism mediated by the selective uptake of the antitumor drug 1-O-octadecyl-2-O-methyl-rac-glycero-3 phosphocholine (ET-18-OCH(3), Edelfosine) into lipid rafts of tumor cells, followed by its coaggregation with Fas death receptor (also known as APO-1 or CD95) and recruitment of apoptotic molecules into Fas-enriched rafts. Drug sensitivity was dependent on drug uptake and Fas expression, regardless of the presence of other major death receptors, such as tumor necrosis factor (TNF) receptor 1 or TNF-related apoptosis-inducing ligand R2/DR5 in the target cell. Drug microinjection experiments in Fas-deficient and Fas-transfected cells unable to incorporate exogenous ET-18-OCH(3) demonstrated that Fas was intracellularly activated. Partial deletion of the Fas intracellular domain prevented apoptosis. Unlike normal lymphocytes, leukemic T cells incorporated ET-18-OCH(3) into rafts coaggregating with Fas and underwent apoptosis. Fas-associated death domain protein, procaspase-8, procaspase-10, c-Jun amino-terminal kinase, and Bid were recruited into rafts, linking Fas and mitochondrial signaling routes. Clustering of rafts was necessary but not sufficient for ET-18-OCH(3)-mediated cell death, with Fas being required as the apoptosis trigger. ET-18-OCH(3)-mediated apoptosis did not require sphingomyelinase activation. Normal cells, including human and rat hepatocytes, did not incorporate ET-18-OCH(3) and were spared. This mechanism represents the first selective activation of Fas in tumor cells. Our data set a framework for the development of more targeted therapies leading to intracellular Fas activation and recruitment of downstream signaling molecules into Fas enriched rafts. PMID- 15289505 TI - Tumor necrosis factor (TNF) receptor shedding controls thresholds of innate immune activation that balance opposing TNF functions in infectious and inflammatory diseases. AB - Tumor necrosis factor (TNF) is a potent cytokine exerting critical functions in the activation and regulation of immune and inflammatory responses. Due to its pleiotropic activities, the amplitude and duration of TNF function must be tightly regulated. One of the mechanisms that may have evolved to modulate TNF function is the proteolytic cleavage of its cell surface receptors. In humans, mutations affecting shedding of the p55TNF receptor (R) have been linked with the development of the TNFR-associated periodic syndromes, disorders characterized by recurrent fever attacks and localized inflammation. Here we show that knock-in mice expressing a mutated nonsheddable p55TNFR develop Toll-like receptor dependent innate immune hyperreactivity, which renders their immune system more efficient at controlling intracellular bacterial infections. Notably, gain of function for antibacterial host defenses ensues at the cost of disbalanced inflammatory reactions that lead to pathology. Mutant mice exhibit spontaneous hepatitis, enhanced susceptibility to endotoxic shock, exacerbated TNF-dependent arthritis, and experimental autoimmune encephalomyelitis. These results introduce a new concept for receptor shedding as a mechanism setting up thresholds of cytokine function to balance resistance and susceptibility to disease. Assessment of p55TNFR shedding may thus be of prognostic value in infectious, inflammatory, and autoimmune diseases. PMID- 15289507 TI - Highly selective escape from KSHV-mediated host mRNA shutoff and its implications for viral pathogenesis. AB - During Kaposi's sarcoma (KS)-associated herpesvirus (KSHV) lytic infection, many virus-encoded signaling molecules (e.g., viral G protein-coupled receptor [vGPCR]) are produced that can induce host gene expression in transiently transfected cells, and roles for such induced host genes have been posited in KS pathogenesis. However, we have recently found that host gene expression is strongly inhibited by 10-12 h after lytic reactivation of KSHV, raising the question of whether and to what extent de novo host gene expression induced by viral signaling molecules can proceed during the lytic cycle. Here, we show by microarray analysis that expression of most vGPCR target genes is drastically curtailed by this host shutoff. However, rare cellular genes can escape the host shutoff and are potently up-regulated during lytic KSHV growth. Prominent among these is human interleukin-6, whose striking induction may contribute to the overexpression of this cytokine in several disease states linked to KSHV infection. PMID- 15289508 TI - Multi-layered representation for cell signaling pathways. AB - To understand complex signaling pathways and networks, it is necessary to develop a formal and structured representation of the available information in a format suitable for analysis by software tools. Due to the complexity and incompleteness of the current biological knowledge about cell signaling, such a device must be able to represent cellular pathways at differing levels of details, one level of information abstract enough to convey an essential signaling flow while hiding its details and another level of information detailed enough to explain the underlying mechanisms that account for the signaling flow described at a more abstract level. We have defined a formal ontology for cell-signaling events that allows us to describe these cellular pathways at various levels of abstraction. Using this formal representation, ROSPath (reactive oxygen species-mediated signaling pathway) database system has been implemented and made available on the web (rospath.ewha.ac.kr). ROSPath is a database system for reactive oxygen species (ROS)-mediated cell signaling pathways and signaling processes in molecular detail, which facilitates a comprehensive understanding of the regulatory mechanisms in signaling pathways. ROSPath includes growth factor-, stress-, and cytokine-induced signaling pathways containing about 500 unique proteins (mostly mammalian) and their related protein states, protein complexes, protein complex states, signaling interactions, signaling steps, and pathways. It is a web-based structured repository of information on the signaling pathways of interest and provides a means for managing data produced by large-scale and high throughput techniques such as proteomics. Also, software tools are provided for querying, displaying, and analyzing pathways, thus furnishing an integrated web environment for visualizing and manipulating ROS-mediated cell-signaling events. PMID- 15289506 TI - Early growth response gene 1-mediated apoptosis is essential for transforming growth factor beta1-induced pulmonary fibrosis. AB - Fibrosis and apoptosis are juxtaposed in pulmonary disorders such as asthma and the interstitial diseases, and transforming growth factor (TGF)-beta(1) has been implicated in the pathogenesis of these responses. However, the in vivo effector functions of TGF-beta(1) in the lung and its roles in the pathogenesis of these responses are not completely understood. In addition, the relationships between apoptosis and other TGF-beta(1)-induced responses have not been defined. To address these issues, we targeted bioactive TGF-beta(1) to the murine lung using a novel externally regulatable, triple transgenic system. TGF-beta(1) produced a transient wave of epithelial apoptosis that was followed by mononuclear-rich inflammation, tissue fibrosis, myofibroblast and myocyte hyperplasia, and septal rupture with honeycombing. Studies of these mice highlighted the reversibility of this fibrotic response. They also demonstrated that a null mutation of early growth response gene (Egr)-1 or caspase inhibition blocked TGF-beta(1)-induced apoptosis. Interestingly, both interventions markedly ameliorated TGF-beta(1) induced fibrosis and alveolar remodeling. These studies illustrate the complex effects of TGF-beta(1) in vivo and define the critical role of Egr-1 in the TGF beta(1) phenotype. They also demonstrate that Egr-1-mediated apoptosis is a prerequisite for TGF-beta(1)-induced fibrosis and remodeling. PMID- 15289511 TI - Prescribing cannabis: freedom, autonomy, and values. AB - In many Western jurisdictions cannabis, unlike most other psychoactive drugs, cannot be prescribed to patients even in cases where medical professionals believe that it would ease the patient's pain or anxiety. The reasons for this prohibition are mostly ideological, although medical and moral arguments have been formulated to support it. In this paper, it is argued that freedom, properly understood, provides a sound ethical reason to allow the use of cannabis in medicine. Scientific facts, appeals to harm and autonomy, and considerations of symbolic value cannot consistently justify prohibitions. PMID- 15289512 TI - Ethical issues in using a cocaine vaccine to treat and prevent cocaine abuse and dependence. AB - A "cocaine vaccine" is a promising immunotherapeutic approach to treating cocaine dependence which induces the immune system to form antibodies that prevent cocaine from crossing the blood brain barrier to act on receptor sites in the brain. Studies in rats show that cocaine antibodies block cocaine from reaching the brain and prevent the reinstatement of cocaine self administration. A successful phase 1 trial of a human cocaine vaccine has been reported. The most promising application of a cocaine vaccine is to prevent relapse to dependence in abstinent users who voluntarily enter treatment. Any use of a vaccine to treat cocaine addicts under legal coercion raises major ethical issues. If this is done at all, it should be carefully trialled first, and only after considerable clinical experience has been obtained in using the vaccine to treat voluntary patients. There will need to be an informed community debate about what role, if any, a cocaine vaccine may have as a way of preventing cocaine addiction in children and adolescents. PMID- 15289513 TI - Further ethical and social issues in using a cocaine vaccine: response to Hall and Carter. AB - Evaluation of the potential of a cocaine vaccine requires a detailed understanding of the intended and unintended social consequences of its use. Prospective technology assessment is always difficult, but in the case of treatment and prevention of cocaine addiction we need to understand not only the neuroscience and pharmacology of cocaine addiction, but also social attitudes to drug use and addiction, the social context of drug use, and the factors which make drug use a rational strategy for an addict and make treatment seeking or relapse more or less likely. By considering different scenarios related to differing levels of effectiveness of the vaccine, the authors argue that vaccination will be at best a useful adjunct to existing methods of treatment, rather than a substitute for them. PMID- 15289514 TI - Nicotine conjugate vaccine: is there a right to a smoking future? AB - Tobacco consumption is believed to be one of the world's greatest preventable health problems. According to the World Health Organisation, 1.1 billion people worldwide are addicted to nicotine with tobacco causing an estimated four million premature deaths every year. The development of a nicotine conjugate vaccine suggests that immunisation may hold promise as a future therapeutic and preventive strategy for tobacco smoking and nicotine addiction. Allowing parents to immunise their children against smoking could be an infringement of children's right to an open future, however, and is not ethically unproblematic PMID- 15289515 TI - The patient who refuses nursing care. AB - OBJECTIVES: The aim of this paper is to examine the way in which nurses manage patients who refuse nursing care procedures. DESIGN: This paper reports on a qualitative study which was undertaken to explore the way in which nurses obtain consent prior to nursing care procedures. Focus groups were carried out to obtain background data concerning how consent is obtained. Critical incidents were collected through in depth interviews as a means of focusing on specific incidents in clinical practice. SETTING: Two teaching hospitals in England. PARTICIPANTS: Purposive sample of qualified nurses. RESULTS: When a patient refuses nursing care, nurses respond by giving information until the patient finally accedes to the procedure. Nurses will go to great lengths to achieve patients' agreement to the procedure, but the extent to which the agreement remains voluntary cannot be ascertained by the data collected in this study. If the patient does not eventually agree to a procedure, there is evidence that nurses will administer the care in the absence of consent. CONCLUSIONS: Nurses are concerned to obtain the patient's consent prior to the administration of nursing care but if this cannot be achieved do not regard obtaining consent as an absolute requirement. Consent is preferred, but not considered essential. Nurses have some understanding of the principles of informed consent but do not apply them to everyday clinical nursing practice. PMID- 15289516 TI - Scale of levels of care versus DNR orders. PMID- 15289517 TI - Is the clock ticking for terminally ill patients in Israel? Preliminary comment on a proposal for a bill of rights for the terminally ill. AB - This paper presents and discusses a recent Israeli proposal to legislate on the rights of the dying patient. A gap exists between elitist biases of the committee proposing the law, and popular values and sentiments. The proposed law divides the dying patients into two groups: "those who wish to go on living" and "those who wish to die". The former will have a right to life prolonging extraordinary care. It is not clear who would foot the bill for this care. Also it is hard to see how this munificence could fail to discriminate against all other patients. Both the secular ethicists and the rabbis involved in drawing up the proposal accepted the assumption that it is good for some terminal patients to die. The rabbis objected, however, to direct and active interventions that shorten life. The solution arrived at was to install timers in the ventilators so as to allow them to expire automatically unless the patient wishes for their resetting. PMID- 15289519 TI - Medical ethics, logic traps, and game theory: an illustrative tale of brain death. AB - Decision making and choices are frequent themes in medical ethics. Game theory is based upon modelled decision making. Game theory, and associated logic traps, may have relevance to the clinical practice of medicine and medical ethics. The "prisoner's dilemma" is one logic trap from game theory in which "rational" decision making on the part of participating individuals can lead to "suboptimal" situations. An example of such a situation involving brain death is presented and discussed from the perspective of the prisoner's dilemma. PMID- 15289520 TI - End of life decisions: attitudes of Finnish physicians. AB - OBJECTIVES: This study investigated Finnish physicians' experiences of decisions concerning living wills and do not resuscitate (DNR) orders and also their views on the role of patients and family members in these decisions. DESIGN: A questionnaire was sent to 800 physicians representing the following specialties: general practice (n = 400); internal medicine (n = 207); neurology (n = 100), and oncology (n = 93). RESULTS: The response rate was 56%. Most of the respondents had a positive attitude toward (92%), and respect for (86%) living wills, and 72% reported situations in which such a will would have been helpful, although experience with their use was limited. The physicians reported both benefits and problems with living wills. Thirteen per cent had completed a living will of their own. Half did not consider living wills to be reliable if they were several years old. Do not resuscitate orders were interpreted in two ways: resuscitation forbidden (70%) or only palliative (symptom oriented) care required (30%). The respondents also documented DNR orders differently. Seventy two per cent discussed DNR decisions always or often with patients able to communicate, and even 76% discussed DNR orders with the family members of patients unable to communicate. Most respondents were able to approach a dying patient without difficulty. They also felt that education in general was needed. CONCLUSIONS: In general Finnish physicians accept living wills, but find they are accompanied by several problems. Many problems could be avoided if physicians and patients conducted progressive discussions about living wills. The differing interpretations of DNR orders are a matter of concern in that they may affect patient treatment. The promotion of patient autonomy with respect to treatment seems rather good, but the limitations of the study need to be kept in mind. PMID- 15289521 TI - Evolutionary ethics: can values change. AB - The hypothesis that values change and evolve is examined by this paper. The discussion is based on a series of examples where, over a period of a few decades, new ethical issues have arisen and values have changed. From this analysis it is suggested that there are a series of core values around which most people would agree. These are unlikely to change over long time periods. There are then a series of secondary or derived values around which there is much more controversy and within which differences of view occur. Such changes need to be documented if we are to understand the process involved in the evolution of differences in ethical views. PMID- 15289523 TI - Prenatal sex and race determination is a slippery slope. PMID- 15289522 TI - Students' attitudes and potential behaviour to a competent patient's request for withdrawal of treatment as they pass through a modern medical curriculum. AB - OBJECTIVE: To examine students' attitudes and potential behaviour to a competent patient's request for withdrawal of treatment as they pass through a modern medical curriculum. DESIGN: Cohort design. SETTING: University of Glasgow Medical School, United Kingdom. SUBJECTS: A cohort of students entering Glasgow University's new learner centred, integrated medical curriculum in October 1996. METHODS: Students' responses before and after year 1, after year 3, and after year 5 to the assisted suicide vignette of the Ethics in Health Care Survey instrument, were examined quantitatively and qualitatively. Analysis of students' multichoice answers enabled measurement of the movement towards professional consensus opinion. Analysis of written justifications helped determine whether their reasoning was consistent with professional consensus and enabled measurement of change in knowledge content and recognition of the values inherent in the vignette. Themes on students' reasoning behind their decision to withdraw treatment or not were also identified. RESULTS: Students' answers were found to be consistent with professional consensus opinion precurriculum and remained so throughout the curriculum. There was an improvement in the knowledge content of the written responses following the first year of the curriculum, which was sustained postcurriculum. However, students were found to analyse the section mainly in terms of autonomy, with few responses considering the other main ethical principles or the wider ethical perspective. Students were unclear on their legal responsibilities. CONCLUSIONS: Students should be encouraged to consider all relevant ethical principles and the wider ethical perspective when deliberating ethical dilemmas. Students should have a clear understanding of their legal responsibilities. PMID- 15289525 TI - A rational cure for prereproductive stress syndrome. PMID- 15289526 TI - Human reproduction: irrational but in most cases morally defensible. PMID- 15289527 TI - Response from Dundee Medical Student Council to "media misinterpretation". PMID- 15289528 TI - Why it is not strongly irrational to have children. PMID- 15289529 TI - Response to: A rational cure for pre-reproductive stress syndrome. AB - This response to "A rational cure for pre-reproductive stress syndrome" first suggests it is existence that is essential and prerequisite to everything good or bad, therefore it deserves to be protected and respected. Secondly, it argues that every life is worth living, even if it is worse than some other lives, if the only alternative is non-existence. Finally, it takes a critical view of and challenges Hayry's suggestion that in a good clinical situation, the idea of the irrationality of having children could be a legitimate part of the guidance given, since it is not the counsellor's or doctor's duty to advise a couple who wish to have children that it is irrational or even immoral to bring a child into life. PMID- 15289530 TI - Woman wants dead fiance's baby: who owns a dead man's sperm. AB - The Brisbane Supreme Court has denied an Australian woman's request to harvest and freeze her dead fiance's sperm for future impregnation. After she was denied access to the sperm, the woman learnt that her fiance may have been a sperm donor and she began checking to find out if his sperm was still available. Given what we know, there is a good ethical argument that the woman should have access to the sperm and should be allowed to have her dead fiance's child. Another aspect of this case is that it illustrates the way in which ethics, law, and personal opinion can differ. PMID- 15289532 TI - Who owns a dead man's sperm? PMID- 15289533 TI - 'Til death us do part: the ethics of postmortem gamete donation. PMID- 15289534 TI - Response to Orr and Siegler--collective intentionality and procreative desires: the permissible view on consent to posthumous conception. AB - Orr and Siegler have recently defended a restrictive view concerning posthumous sperm retrieval and conception, which would limit insemination to those cases where the deceased man has provided explicit consent for such a procedure. The restrictive view dominates current law and practice. A permissible view, in contrast, would allow insemination and conception in all but those cases where the posthumous procedure has been explicitly refused, or where there is no reasonable evidence that the deceased person desired children. I describe a phenomenology of procreative desires which supports the permissible view, and which is compatible with requirements concerning the interests of the decedent, concepts of medical infertility, and the welfare of the future child. The account illustrates how our current obsession with individual rights and autonomy can be self-defeating and repressive. PMID- 15289535 TI - A wrongful existence in the Netherlands. PMID- 15289536 TI - Gaps, conflicts, and consensus in the ethics statements of professional associations, medical groups, and health plans. AB - BACKGROUND: Patients today interact with physicians, physician groups, and health plans, each of which may follow distinct ethical guidelines. METHOD: We systematically compared physician codes of ethics with ethics policies at physician group practices and health plans, using the 1998-99 policies of 38 organisations-18 medical associations (associations), nine physician group practices (groups), and 12 health plans (plans)-selected using random and stratified purposive sampling. A clinician and a social scientist independently abstracted each document, using a 397-item health care ethics taxonomy; a reconciled abstraction form was used for analysis. This study focuses on ethics policies regarding professional obligation towards patients, resource allocation, and care for the vulnerable in society. RESULTS: A majority in all three groups mention "fiduciary obligations" of one sort or another, but associations generally address physician/patient relations but not health plan obligations, while plans rarely endorse physicians' obligations of advocacy, beneficence, and non-maleficence. Except for occasional mentions of cost effectiveness or efficiency, ethical considerations in resource allocation rarely arise in the ethics policies of all three organisational types. Very few associations, groups, or plans specifically endorse obligations to vulnerable populations. CONCLUSIONS: With some important exceptions, we found that the ethics policies of associations, groups, and plans are narrowly focused and often ignore important ethical concerns for society, such as resource allocation and care for vulnerable populations. More collaborative work is needed to build integrated sets of ethical standards that address the aims and responsibilities of the major stakeholders in health care delivery. PMID- 15289537 TI - Ethics of refusing parental requests to withhold or withdraw treatment from their premature baby. AB - In the United Kingdom women have access to termination of pregnancy for maternal reasons until 24 weeks' completed gestation, but it is accepted practice for children born at or beyond 25 weeks' gestation to be treated according to the child's perceived best interests even if this is not in accordance with parental wishes. The authors present a case drawn from clinical practice which highlights the discomfort that parents may feel about such an abrupt change in their rights over their child, and argue that parents should have greater autonomy over treatment decisions regarding their prematurely born children. PMID- 15289540 TI - Stem cells, embryos, and the environment: a context for both science and ethics. AB - Debate on the potential and uses of human stem cells tends to be conducted by two constituencies-ethicists and scientists. On many occasions there is little communication between the two, with the result that ethical debate is not informed as well as it might be by scientific insights. The aim of this paper is to highlight those scientific insights that may be of relevance for ethical debate. Environmental factors play a significant role in identifying stem cells and their various subtypes. Research related to the role of the microenvironment has led to emphasis upon "plasticity", which denotes the ability of one type of stem cell to undergo a transition to cells from other lineages. This could increase the value given to adult stem cells, in comparison with embryonic stem cell research. Any such conclusion should be treated with caution, however, since optimism of this order is not borne out by current research. The role of the environment is also important in distinguishing between the terms totipotency and pluripotency. We argue that blastocysts (early embryos) and embryonic stem cells are only totipotent if they can develop within an appropriate environment. In the absence of this, they are merely pluripotent. Hence, blastocysts in the laboratory are potentially totipotent, in contrast to their counterparts within the human body which are actually totipotent. This may have implications for ethical debate, suggesting as it does that arguments based on potential for life may be of limited relevance. PMID- 15289541 TI - Law and policy in the era of reproductive genetics. AB - The extent to which society utilises the law to enforce its moral judgments remains a dominant issue in this era of embryonic stem cell research, preimplantation genetic diagnosis, and human reproductive cloning. Balancing the potential health benefits and diverse moral values of society can be a tremendous challenge. In this context, governments often adopt legislative bans and prohibitions and rely on the inflexible and often inappropriate tool of criminal law. Legal prohibitions in the field of reproductive genetics are not likely to reflect adequately the depth and diversity of competing stakeholder positions. Rather, a comprehensive and readily responsive regulatory policy is required. Such a policy must attend to the evolving scientific developments and ethical considerations. We outline a proposal for effective, responsive, and coherent oversight of new reproductive genetic technologies. PMID- 15289542 TI - Disability, identity and the "expressivist objection". AB - The practice of prenatal screening for disability is sometimes objected to because of the hurt and offence such practices may cause to people currently living with disabilities. This objection is commonly termed "the expressivist objection". In response to the objection it is standardly claimed that disabilities are analogous to illnesses. And just as it would be implausible to suppose reduction of the incidence of illnesses such as flu sends a negative message to ill people, so it is not plausible to suppose prevention of disability sends a negative message to disabled people. The expressivist objection hinges, however, upon a view of the relationship between disability and self identity which sees disability as part of the identity of the disabled person, in a way in which illnesses such as flu cannot be. This possibility is generally not considered in critiques of the expressivist objection. In this paper, an "identity claim" to the effect that disabilities can be identity constituting is accepted and the force of the expressivist argument is reconsidered in the light of its acceptance. It is concluded that even when such an identity claim is accepted, the expressivist objection is still not morally compelling. PMID- 15289544 TI - Genome update: alignment of bacterial chromosomes. PMID- 15289545 TI - Sequence similarity between multidrug resistance efflux pumps of the ABC and RND superfamilies. PMID- 15289546 TI - Cyclic di-GMP as a bacterial second messenger. AB - Environmental signals trigger changes in the bacterial cell surface, including changes in exopolysaccharides and proteinaceous appendages that ultimately favour bacterial persistence and proliferation. Such adaptations are regulated in diverse bacteria by proteins with GGDEF and EAL domains. These proteins are predicted to regulate cell surface adhesiveness by controlling the level of a second messenger, the cyclic dinucleotide c-di-GMP. Genetic evidence suggests that the GGDEF domain acts as a nucleotide cyclase for c-di-GMP synthesis while the EAL domain is a good candidate for the opposing activity, a phosphodiesterase for c-di-GMP degradation. PMID- 15289547 TI - Controlled expression of CluA in Lactococcus lactis and its role in conjugation. AB - CluA is a 136 kDa surface-bound protein encoded by the chromosomally located sex factor of Lactococcus lactis MG1363 and is associated with cell aggregation linked to high-frequency transfer of the sex factor. To further investigate the involvement of CluA in these phenomena, the cluA gene was cloned on a plasmid, downstream from the lactococcal nisA promoter. In a sex-factor-negative MG1363 derivative, nisin-controlled CluA expression resulted in aggregation, despite the absence of the other genes of the sex factor. Therefore, CluA is the only sex factor component responsible for aggregation. The direct involvement of CluA in the establishment of cell-to-cell contact for aggregate formation was observed by electron microscopy using immunogold-labelled CluA antibodies. Inactivation of cluA in an MG1363 background led to a dramatic decrease in sex factor conjugation frequency compared to the parental strain. Increasing levels of CluA expressed in trans in the cluA-inactivated donor strain facilitated a gradual restoration of conjugation frequency, reaching that of the parental strain. In conclusion, CluA is essential for efficient sex factor transfer in conjugation of L. lactis. PMID- 15289548 TI - Involvement of a Rab8-like protein of Dictyostelium discoideum, Sas1, in the formation of membrane extensions, secretion and adhesion during development. AB - Establishment of cell-cell adhesions, regulation of actin, and secretion are critical during development. Rab8-like GTPases have been shown to modulate these cellular events, suggesting an involvement in developmental processes. To further elucidate the function of Rab8-like GTPases in a developmental context, a Rab8 related protein (Sas1) of Dictyostelium discoideum was examined, the expression of which increases at the onset of development. Dictyostelium cell lines expressing inactive (N128I mutant) and constitutively active (Q74L mutant) Sas1 as green fluorescent protein (GFP)-Sas1 chimeras were generated. Cells expressing Sas1Q74L displayed numerous actin-rich membrane protrusions, increased secretion, and were unable to complete development. In particular, these cells demonstrated a reduction in adhesion as well as in the levels of a cell adhesion molecule, gp24 (DdCAD-1). In contrast, cells expressing Sas1N128I exhibited increased cell cell adhesion and increased levels of gp24. Counting factor is a multisubunit signalling complex that is secreted in early development and controls aggregate size by negatively regulating the levels of cell adhesion molecules, including gp24. Interestingly, the Sas1Q74L mutant demonstrated increased levels of extracellular countin, a subunit of counting factor, suggesting that Sas1 may regulate trafficking of counting factor components. Together, the data suggest that Sas1 may be a key regulator of actin, adhesion and secretion during development. PMID- 15289549 TI - Killer toxin of Pichia membranifaciens and its possible use as a biocontrol agent against grey mould disease of grapevine. AB - The use of Pichia membranifaciens CYC 1106 killer toxin against Botrytis cinerea was investigated. This strain exerted a broad-specificity killing action against other yeasts and fungi. At pH 4, optimal killer activity was observed at temperatures up to 20 degrees C. At 25 degrees C the toxic effect was reduced to 70%. The killer activity was higher in acidic medium. Above about pH 4.5 activity decreased sharply and was barely noticeable at pH 6. The killer toxin protein from P. membranifaciens CYC 1106 was purified to electrophoretic homogeneity. SDS PAGE of the purified killer protein indicated an apparent molecular mass of 18 kDa. Killer toxin production was stimulated in the presence of non-ionic detergents. The toxin concentrations present in the supernatant during optimal production conditions exerted a fungicidal effect on a strain of B. cinerea. The symptoms of infection and grey mould observed in Vitis vinifera plants treated with B. cinerea were prevented in the presence of purified P. membranifaciens killer toxin. The results obtained suggest that P. membranifaciens CYC 1106 killer toxin is of potential use in the biocontrol of B. cinerea. PMID- 15289550 TI - Kluyveromyces phaffii killer toxin active against wine spoilage yeasts: purification and characterization. AB - The killer toxin secreted by Kluyveromyces phaffii (KpKt) is active against spoilage yeast under winemaking conditions and thus has potential applications in the biocontrol of undesired micro-organisms in the wine industry. Biochemical characterization and N-terminal sequencing of the purified toxin show that KpKt is a glycosylated protein with a molecular mass of 33 kDa. Moreover, it shows 93% and 80% identity to a beta-1,3-glucanase of Saccharomyces cerevisiae and a beta 1,3-glucan transferase of Candida albicans, respectively, and it is active on laminarin and glucan, thus showing a beta-glucanase activity. Competitive inhibition of killer activity by cell-wall polysaccharides suggests that glucan (beta-1,3 and beta-1,6 branched glucans) represents the first receptor site of the toxin on the envelope of the sensitive target. Flow cytometry analysis of the sensitive target after treatment with KpKt and K1 toxin of S. cerevisiae, known to cause loss of cell viability via formation of pores in the cell membrane, suggests a different mode of action for KpKt. PMID- 15289551 TI - Effects of mutations in the rpoS gene on cell viability and global gene expression under nitrogen starvation in Escherichia coli. AB - Escherichia coli bearing an rpoS amber or disrupted mutation exhibited a significant decrease in the number of colony-forming units (c.f.u.) when exposed to nitrogen starvation, which was not observed in cells bearing a functional rpoS allele. The decrease in the number of c.f.u. that was observed about 25 h after initiation of nitrogen starvation was prevented by the addition of nitrogen within 3 h but not by the addition of nitrogen at more than 7 h after the initiation of nitrogen starvation, suggesting that a process leading to a decline in c.f.u. starts within this period. DNA microarray analysis of the rpoS mutant showed that a large number of genes including many functionally undefined genes were affected by nitrogen starvation. The expression levels of sigma(S) and sigma(H) regulon genes encoding acid-resistant proteins (hdeA, hdeB, gadA and gadB), DNA-binding protein (dps), chaperones (dnaK, ibpA, ibpB, dnaJ and htpG), chaperonins (mopB and mopA) and energy-metabolism-related proteins (hyaABCDF and gapA), and those of other genes encoding nucleotide-metabolism-related proteins (deoC and deoB), cell-division protein (ftsL), outer-membrane lipoprotein (slp) and DNA-binding protein (stpA) were significantly decreased by 10 h nitrogen starvation. The genes encoding transport/binding proteins (nac, amtB, argT, artJ, potF and hisJ) and amino acid-metabolism-related proteins (glnA, trpB, argG, asnB, argC, gdhA, cstC, ntrB, asd and lysC) were significantly up-regulated under the same condition, some of which are known Ntr genes expressed under nitrogen limitation. On the basis of these results, possible causes of the decrease in the number of c.f.u. under nitrogen starvation are discussed. PMID- 15289552 TI - The impact of different intensities of green light on the bacteriochlorophyll homologue composition of the Chlorobiaceae Prosthecochloris aestuarii and Chlorobium phaeobacteroides. AB - Members of the Chlorobiaceae and Chloroflexaceae are unique among the phototrophic micro-organisms in having a remarkably rich chlorophyll pigment diversity. The physiological regulation of this diversity and its ecological implications are still enigmatic. The bacteriochlorophyll composition of the chlorobiaceae Prosthecochloris aestuarii strain CE 2404 and Chlorobium phaeobacteroides strain UdG 6030 was therefore studied by both HPLC with photodiode array (PDA) detection and liquid chromatography-mass spectrometry (LC MS). These strains were grown in liquid cultures under green light (480-615 nm) at different light intensities (0.2-55.7 micromol photons m(-2) s(-1)), simulating the irradiance regime at different depths of the water column of deep lakes. The specific growth rates of Ptc. aestuarii under green light achieved a maximum of 0.06 h(-1) at light intensities exceeding 6 micromol photons m(-2) s( 1), lower than the maximum observed under white light (approx. 0.1 h(-1)). The maximal growth rates of Chl. phaeobacteroides under green light were slightly higher (0.07 h(-1)) than observed for Ptc. aestuarii and were achieved at 3.5 and 4.3 micromol photons m(-2) s(-1). LC-MS/MS analysis of pigment extracts revealed most (>90 %) BChl c homologues of Ptc. aestuarii to be esterified with farnesol. The homologues differed in mass by multiples of 14 Da, reflecting different alkyl subsituents at positions C-8 and C-12 on the tetrapyrrole macrocycle. The relative proportions of the individual homologues varied only slightly among different light intensities. The specific content of BChl c was maximal at 3-5 micromol photons m(-2) s(-1) [400+/-150 nmol BChl c (mg protein)(-1)]. In the case of Chl. phaeobacteroides, the specific content of BChl e was maximal at 4.3 micromol photons m(-2) s(-1) [115 nmol BChl e (mg protein)(-1)], and this species was characterized by high carotenoid (isorenieratene) contents. The major BChl e forms were esterified with a range of isoprenoid and straight-chain alcohols. The major isoprenoid alcohols comprised mainly farnesol and to a lesser extent geranylgeraniol. The straight-chain alcohols included C(15), C(15 : 1), C(16), C(16 : 1) and C(17). Interestingly, the proportion of straight alkyl chains over isoprenoid esterified side chains shifted markedly with increasing light intensity: the isoprenoid side chains dominated at low light intensities, while the straight-chain alkyl substituents dominated at higher light intensities. The authors propose that this phenomenon may be explained as a result of changing availability of reducing power, i.e. the highly reduced straight-chain alcohols have a higher biosynthetic demand for NADPH(2) than the polyunsaturated isoprenoid with the same number of carbon atoms. PMID- 15289553 TI - Characterization of vaginal microbial communities in adult healthy women using cultivation-independent methods. AB - The normal microbial flora of the vagina plays an important role in preventing genital and urinary tract infections in women. Thus an accurate understanding of the composition and ecology of the ecosystem is important to understanding the aetiology of these diseases. Common wisdom is that lactobacilli dominate the normal vaginal microflora of post-pubertal women. However, this conclusion is based on methods that require cultivation of microbial populations; an approach that is known to yield a biased and incomplete assessment of microbial community structure. In this study cultivation-independent methods were used to analyse samples collected from the mid-vagina of five normal healthy Caucasian women between the ages of 28 and 44. Total microbial community DNA was isolated following resuspension of microbial cells from vaginal swabs. To identify the constituent numerically dominant populations in each community 16S rRNA gene libraries were prepared following PCR amplification using the 8f and 926r primers. From each library, the DNA sequences of approximately 200 16S rRNA clones were determined and subjected to phylogenetic analyses. The diversity and kinds of organisms that comprise the vaginal microbial community varied among women. Species of Lactobacillus appeared to dominate the communities in four of the five women. However, the community of one woman was dominated by Atopobium sp., whereas a second woman had appreciable numbers of Megasphaera sp., Atopobium sp. and Leptotrichia sp., none of which have previously been shown to be common members of the vaginal ecosystem. Of the women whose communities were dominated by lactobacilli, there were two distinct clusters, each of which consisted of a single species. One class consisted of two women with genetically divergent clones that were related to Lactobacillus crispatus, whereas the second group of two women had clones of Lactobacillus iners that were highly related to a single phylotype. These surprising results suggest that culture-independent methods can provide new insights into the diversity of bacterial species found in the human vagina, and this information could prove to be pivotal in understanding risk factors for various infectious diseases. PMID- 15289554 TI - The novel extracellular Streptomyces reticuli haem-binding protein HbpS influences the production of the catalase-peroxidase CpeB. AB - The Gram-positive soil bacterium and cellulose degrader Streptomyces reticuli synthesizes the mycelium-associated enzyme CpeB, which displays haem-dependent catalase and peroxidase activity, as well as haem-independent manganese peroxidase activity. Downstream of the cpeB gene, a so far unknown gene was identified. The new gene and its mutated derivatives were cloned in Escherichia coli as well as in Streptomyces lividans and a gene-disruption mutant within the chromosome of the original S. reticuli host was constructed, comparative physiological, biochemical and immunological studies then allowed the deduction of the following characteristics of the novel gene product. (i) The protein was found extracellularly; the substitution of twin arginines within the signal peptide abolished its secretion. (ii) The highly purified protein interacted specifically with haem and hence was designated HbpS (haem-binding protein of Streptomyces). (iii) HbpS contained three histidine residues surrounded by hydrophobic amino acids; one of them was located within the motif LX(3)THLX(10)AA, which is related to the motif within the yeast cytochrome c peroxidase LX(2)THLX(10)AA whose histidine residue interacts with haem. (iv) The addition of haemin (Fe(3+) oxidized form of haem) to the Streptomyces cultures led to enhanced levels of HbpS which correlated with increased haemin-resistance. (v) The presence of HbpS increased synthesis of the highly active catalase peroxidase CpeB containing haem. In this process HbpS could act as a chaperone that binds haem and then delivers it to the mycelium-associated CpeB; HbpS could also interact with membrane-associated proteins involved in a signal transduction cascade regulating the expression of cpeB. (vi) HbpS shared varying degrees of amino acid identities with bacterial proteins of so far unknown function. This report contributes to the elucidation of the biological function of these proteins. PMID- 15289555 TI - Identification and transcriptional organization of a gene cluster involved in biosynthesis and transport of acinetobactin, a siderophore produced by Acinetobacter baumannii ATCC 19606T. AB - In order to assimilate iron, Acinetobacter baumannii ATCC 19606(T) produces a siderophore named acinetobactin (Ab) that is composed of equimolar quantities of 2,3-dihydroxybenzoic acid (DHBA), L-threonine and N-hydroxyhistamine. Application of the Fur titration assay system to A. baumannii genomic libraries, followed by further cloning of the regions surrounding the candidate genes, led to the identification of the Ab cluster, which harbours the genetic determinants necessary for the biosynthesis and transport of the siderophore. However, an entA homologue essential for DHBA biosynthesis was not found in this cluster. Functions of potential biosynthetic genes inferred by homology studies suggested that the precursors, DHBA, l-threonine and N-hydroxyhistamine, are linked in steps resembling those of bacterial non-ribosomal peptide synthesis to form Ab. Genes responsible for the two-step biosynthesis of N-hydroxyhistamine from histidine were also identified in this cluster. Their genetic organization suggests that five genes involved in the transport system of ferric Ab into the cell cytosol form an operon. Construction of disruptants of some selected genes followed by phenotypic analysis supported their predicted biological functions. Interestingly, three additional genes probably involved in the intracellular release of iron from ferric Ab and the secretion of nascent Ab are contained in this cluster. Primer extension and RT-PCR analyses suggested that the Ab cluster, which includes 18 genes, is organized in seven transcriptional units originating from respective Fur-regulated promoter-operator regions. PMID- 15289556 TI - Partial purification and characterization of a non-cyanobacterial cyanophycin synthetase from Acinetobacter calcoaceticus strain ADP1 with regard to substrate specificity, substrate affinity and binding to cyanophycin. AB - This study reports, for the first time, purification and biochemical characterization of a cyanophycin synthetase from a non-cyanobacterial strain. Cyanophycin synthetase of Acinetobacter calcoaceticus strain ADP1 was purified 69 fold from recombinant Escherichia coli by two chromatographic steps and one novel affinity step utilizing the Mg(2+)-dependent binding of the enzyme to cyanophycin. Unlike cyanobacterial cyanophycin synthetases characterized so far, the purified enzyme from A. calcoaceticus strain ADP1 did not accept lysine as an alternative substrate to arginine. The apparent K(m)-values for arginine (47 microM) and aspartic acid (240 microM) were similar to those of known cyanophycin synthetases from cyanobacteria, but this enzyme had a slightly higher affinity for aspartic acid. In addition, the two different ATP-binding sites of the enzyme were characterized independently of each other with respect to K(m) values for ATP. The ATP-binding site responsible for the addition of arginine was found to have a much higher affinity for ATP (38 microM) than that responsible for the addition of aspartate (210 mM). Furthermore, the binding of the enzyme to the two possible forms of cyanophycin granule polypeptide (CGP), CGP-Asp and CGP-Arg, was studied. While both forms bound around 30-40 % of the enzyme activity present under the assay conditions, binding was Mg(2+)-dependent in the case of CGP-Asp. Two-dimensional gel electrophoresis revealed that both forms of cyanophycin were equally abundant in cyanophycin-accumulating cells of A. calcoaceticus ADP1. PMID- 15289557 TI - Characterization of the Bacillus subtilis YxdJ response regulator as the inducer of expression for the cognate ABC transporter YxdLM. AB - The genome of Bacillus subtilis, like those of some other AT-rich Gram-positive bacteria, has the uncommon feature of containing several copies of arrangements in which the genes encoding two-component and cognate ABC transporter systems are adjacent. As the function of one of these systems, the product of the yxd locus, is still unknown, it was analysed further in order to get some clues on the physiological role of the gene products it encodes. The yxdJ gene was shown to encode a DNA-binding protein that directly controls transcription of the neighbouring operon encoding the ABC transporter YxdLM. Primer extension and DNase protection experiments allowed precise definition of the yxdLM transcription start and controlling region. Two putative direct repeats were identified that are proposed to be the YxdJ response regulator binding sites. Whole-cell transcriptome analyses revealed that the YxdJ regulon is extremely restricted. In addition to the yxdJKLMyxeA operon, only a few genes involved in modifications of the bacterial cell wall were shown to be regulated by YxdJ. PMID- 15289558 TI - Post-transcriptional regulation of the Bacillus subtilis pst operon encoding a phosphate-specific ABC transporter. AB - During phosphate starvation, Bacillus subtilis regulates genes in the PhoP regulon to reduce the cell's requirement for this essential substrate and to facilitate the recovery of inorganic phosphate from organic sources such as teichoic and nucleic acids. Among the proteins that are highly induced under these conditions is PstS, the phosphate-binding lipoprotein component of a high affinity ABC-type phosphate transporter. PstS is encoded by the first gene in the pst operon, the other four members of which encode the integral membrane and cytoplasmic components of the transporter. The transcription of the pst operon was analysed using a combination of methods, including transcriptional reporter gene technology, Northern blotting and DNA arrays. It is shown that the primary transcript of the pst operon is processed differentially to maintain higher concentrations of PstS relative to other components of the transporter. The comparative studies have revealed limitations in the use of reporter gene technology for analysing the transcription of operons in which the messenger RNA transcript is differentially processed. PMID- 15289559 TI - Genomic organization and in vivo characterization of proteolytic activity of FtsH of Mycobacterium smegmatis SN2. AB - The ftsH gene of Mycobacterium smegmatis SN2 (MsftsH) was cloned from two independent partial genomic DNA libraries and characterized, along with the identification of ephA and folE as the neighbouring upstream and downstream genes respectively. The genomic organization of the MsftsH locus was found to be identical to that of the Mycobacterium tuberculosis ftsH gene (MtftsH) and similar to that of other bacterial genera, but with divergence in the upstream region. The MsftsH gene is 2.3 kb in size and encodes the AAA (ATPases Associated with diverse cellular Activities) family Zn(2+)-metalloprotease FtsH (MsFtsH) of 85 kDa molecular mass. This was demonstrated from the expression of the full length recombinant gene in Escherichia coli JM109 cells and from the identification of native MsFtsH in M. smegmatis SN2 cell lysates by Western blotting with anti-MtFtsH and anti-EcFtsH antibodies respectively. The recombinant and the native MsFtsH proteins were found localized to the membrane of E. coli and M. smegmatis cells respectively. Expression of MsFtsH protein in E. coli was toxic and resulted in growth arrest and filamentation of cells. The MsftsH gene did not complement lethality of a DeltaftsH3 : : kan mutation in E. coli, but when expressed in E. coli cells, it efficiently degraded conventional FtsH substrates, namely sigma(32) protein and the protein translocase subunit SecY, of E. coli cells. PMID- 15289560 TI - The Candida albicans pH-regulated KER1 gene encodes a lysine/glutamic-acid-rich plasma-membrane protein that is involved in cell aggregation. AB - Immunoscreening of a Candida albicans cDNA library with a polyclonal germ-tube specific antibody (pAb anti-gt) resulted in the isolation of a gene encoding a lysine/glutamic-acid-rich protein, which was consequently designated KER1. The nucleotide and deduced amino acid sequences of this gene displayed no significant homology with any other known sequence. KER1 encodes a 134 kDa lysine (14.5%)/glutamic acid (16.7%) protein (Ker1p) that contains two potential transmembrane segments. KER1 was expressed in a pH-conditional manner, with maximal expression at alkaline pH and lower expression at pH 4.0, and was regulated by RIM101. A Deltaker1/Deltaker1 null mutant grew normally but was hyperflocculant under germ-tube-inducing conditions, yet this behaviour was also observed in stationary-phase cells grown under other incubation conditions. Western blotting analysis of different subcellular fractions, using as a probe a monospecific polyclonal antibody raised against a highly antigenic domain of Ker1p (pAb anti-Ker1p), revealed the presence of a 134 kDa band in the purified plasma-membrane fraction from the wild-type strain that was absent in the homologous preparation from Deltaker1/Deltaker1 mutant. The pattern of cell-wall protein and mannoprotein species released by digestion with beta-glucanases, reactive towards pAbs anti-gt and anti-Ker1p, as well as against concanavalin A, was also different in the Deltaker1/Deltaker1 mutant. Mutant strains also displayed an increased cell-surface hydrophobicity and sensitivity to Congo red and Calcofluor white. Overall, these findings indicate that the mutant strain was affected in cell-wall composition and/or structure. The fact that the ker1 mutant had attenuated virulence in systemic mouse infections suggests that this surface protein is also important in host-fungus interactions. PMID- 15289561 TI - Identification of two new genes involved in twitching motility in Pseudomonas aeruginosa. AB - Mu transposition complexes were used for transposon mutagenesis of Pseudomonas aeruginosa strain PA68. Mu DNA transposition complexes were assembled with MuA transposase and an artificial mini-Mu transposon in vitro, and introduced into Pseudomonas aeruginosa by electroporation. Eight mutants deficient in twitching motility were isolated. Southern blotting confirmed that the insertions had occurred as single events. DNA sequencing of the region flanking the insertion in the twitching-motility mutants revealed that the mini-Mu transposon had inserted into six different genes, PAO171, PA1822, PAO413, PA4959, PA4551 and PA5040. Four of these have previously been proven to be needed for twitching motility, whereas the PA1822 and PA0171 genes have not previously been shown to be required for twitching motility. The twitching-motility defect in the PA1822 mutant was partially complemented by providing the PA1822 gene in trans, and the defect in the PA0171 mutant was fully complemented when PA0171 was provided. A PA0171 mutant and a PA1822 mutant were constructed by gene replacement in the P. aeruginosa PAO1 strain. These mutants were deficient in twitching motility, showing that both the PA1822 and the PA0171 gene are involved in twitching motility. PMID- 15289562 TI - Expression of mptC of Listeria monocytogenes induces sensitivity to class IIa bacteriocins in Lactococcus lactis. AB - Sensitivity to class IIa bacteriocins from lactic acid bacteria was recently associated with the mannose phosphotransferase system (PTS) permease, in Listeria monocytogenes. To assess the involvement of this protein complex in class IIa bacteriocin activity, the mptACD operon, encoding, was heterologously expressed in an insensitive species, namely Lactococcus lactis, using the NICE double plasmid system. Upon induction of the cloned operon, the recombinant Lc. lactis became sensitive to leucocin A. Pediocin PA-1 and enterocin A also showed inhibitory activity against Lc. lactis cultures expressing mptACD. Furthermore, the role of the three genes of the mptACD operon was investigated. Derivative plasmids containing various combinations of these three genes were made from the parental mptACD plasmid by divergent PCR. The results showed that expression of mptC alone is sufficient to confer sensitivity to class IIa bacteriocins in Lc. lactis. PMID- 15289563 TI - Attenuated virulence and protective efficacy of a Burkholderia pseudomallei bsa type III secretion mutant in murine models of melioidosis. AB - Melioidosis is a severe infectious disease of animals and humans caused by the Gram-negative intracellular pathogen Burkholderia pseudomallei. An Inv/Mxi-Spa like type III protein secretion apparatus, encoded by the B. pseudomallei bsa locus, facilitates bacterial invasion of epithelial cells, escape from endocytic vesicles and intracellular survival. This study investigated the role of the Bsa type III secretion system in the pathogenesis of melioidosis in murine models. B. pseudomallei bipD mutants, lacking a component of the translocation apparatus, were found to be significantly attenuated following intraperitoneal or intranasal challenge of BALB/c mice. Furthermore, a bipD mutant was attenuated in C57BL/6 IL 12 p40(-/-) mice, which are highly susceptible to B. pseudomallei infection. Mutation of bipD impaired bacterial replication in the liver and spleen of BALB/c mice in the early stages of infection. B. pseudomallei mutants lacking either the type III secreted guanine nucleotide exchange factor BopE or the putative effectors BopA or BopB exhibited varying degrees of attenuation, with mutations in bopA and bopB causing a significant delay in median time to death. This indicates that bsa-encoded type III secreted proteins may act in concert to determine the outcome of B. pseudomallei infection in mice. Mice inoculated with the B. pseudomallei bipD mutant were partially protected against subsequent challenge with wild-type B. pseudomallei. However, immunization of mice with purified BipD protein was not protective. PMID- 15289564 TI - Identification of an extracellular matrix protein adhesin, EmaA, which mediates the adhesion of Actinobacillus actinomycetemcomitans to collagen. AB - Actinobacillus actinomycetemcomitans is an aetiologic agent in the development of periodontal and some systemic diseases in humans. This pathogen localizes to the underlying connective tissue of the oral cavity in individuals with periodontal disease. The adhesion of A. actinomycetemcomitans to extracellular matrix components of the connective tissue prompted this study to identify gene products mediating the interaction of A. actinomycetemcomitans to these molecules. A transposon mutagenesis system was optimized for use in A. actinomycetemcomitans and used to generate an insertional mutant library. A total of 2300 individual insertion transposon mutants were screened for changes in the adhesion to collagen and fibronectin. Mutants were identified which exhibited the following phenotypes: a decrease in collagen binding; a decrease in fibronectin binding; a decrease in binding to both proteins; and an increase in binding to both collagen and fibronectin. The identification of mutants defective in adhesion to the individual proteins indicates that distinct adhesins are expressed by this organism. Molecular analysis of these mutants implicated 11 independent loci in protein adhesion. One gene, emaA, is likely to encode a direct mediator of collagen adhesion, based on predicted protein features homologous to the collagen binding protein YadA of Yersinia enterocolitica. EmaA was localized to the outer membrane, as expected for an adhesin. Reduction in fibronectin adhesion appeared to be influenced by abrogation of proteins involved in molybdenum-cofactor biosynthesis. Several other loci identified as reducing or increasing adhesion to both collagen and fibronectin are suggested to be involved in regulatory cascades that promote or repress expression of collagen and fibronectin adhesins. Collectively, the results support the hypothesis that A. actinomycetemcomitans host colonization involves afimbrial adhesins for extracellular matrix proteins, and that the expression of adhesion is modulated by global regulatory mechanisms. PMID- 15289565 TI - Genetic and functional analysis of the cytK family of genes in Bacillus cereus. AB - CytK is a pore-forming toxin of Bacillus cereus that has been linked to a case of necrotic enteritis. PCR products of the expected size were generated with cytK primers in 13 of 29 strains. Six strains were PCR-positive for the related gene hly-II, which encodes haemolysin II, a protein that is 37 % identical to the original CytK. Five of the strains were positive for both genes. The DNA sequences of putative cytK genes from three positive strains were determined, and the deduced amino acid sequences were 89 % identical to that of the original CytK. The authors have designated this new cytK variant cytK-2, and refer to the original cytK as cytK-1. The CytK-2 proteins from these three strains were isolated, and their identity was verified by N-terminal sequencing. blast analysis using the cytK-2 gene sequences revealed very high homology with two cytK-2 sequences in the genomes of B. cereus strains ATCC 14579 and ATCC 10987. The differences between CytK-1 and the CytK-2 proteins were clustered to certain regions of the proteins. The isolated CytK-2 proteins were haemolytic and toxic towards human intestinal Caco-2 cells and Vero cells, although their toxicity was about 20 % of that of CytK-1. Both native and recombinant CytK-2 proteins from B. cereus 1230-88 were able to form pores in planar lipid bilayers, but the majority of the channels observed were of lower conductance than those created by CytK-1. It is likely that CytK-2 toxins contribute to the enterotoxicity of several strains of B. cereus, although not all of the CytK-2 toxins may be as harmful as the CytK-1 originally isolated. PMID- 15289566 TI - Comparison of cytotoxin cytK promoters from Bacillus cereus strain ATCC 14579 and from a B. cereus food-poisoning strain. AB - The cytotoxin CytK produced by Bacillus cereus is believed to be involved in food borne diseases. The transcriptional activity of the cytK promoter region in a food-poisoning strain was studied using a reporter gene and compared with that in the reference B. cereus strain ATCC 14579. In the food-poisoning strain, cytK is more strongly transcribed, possibly explaining the pathogenicity. The global regulator PlcR in B. cereus controls several putative virulence factors. It was found that PlcR regulates cytK in this clinical strain despite a mismatch in the PlcR recognition site, as currently defined. This suggests that the PlcR box consensus should be reconsidered and that the PlcR regulon might be larger than suspected. It is also shown that the high level of cytK transcription is not caused by a modification in the PlcR recognition site. PMID- 15289567 TI - Expression of bacteriophage phiEa1h lysozyme in Escherichia coli and its activity in growth inhibition of Erwinia amylovora. AB - A 3.3 kb fragment from Erwinia amylovora phage Ea1h in plasmid pJH94 was previously characterized and found to contain an exopolysaccharide depolymerase (dpo) gene and two additional ORFs encoding 178 and 119 amino acids. ORF178 (lyz) and ORF119 (hol) were found to overlap by 19 bp and they resembled genes encoding lysozymes and holins. In nucleotide sequence alignments, lyz had structurally conserved regions with residues important for lysozyme function. The lyz gene was cloned into an expression vector and expressed in Escherichia coli. Active lysozyme was detected only when E. coli cells with the lyz gene and a kanamycin resistance cassette were grown in the presence of kanamycin. Growth of Erw. amylovora was inhibited after addition of enzyme exceeding a threshold for lysozyme to target cells. When immature pears were soaked in lysates of induced cells, symptoms such as ooze formation and necrosis were retarded or inhibited after inoculation with Erw. amylovora. PMID- 15289568 TI - Topological and deletion analysis of CorS, a Pseudomonas syringae sensor kinase. AB - A modified two-component regulatory system consisting of two response regulators, CorR and CorP, and the histidine protein kinase CorS, regulates the thermoresponsive production of the phytotoxin coronatine (COR) in Pseudomonas syringae PG4180. COR is produced at the virulence-promoting temperature of 18 degrees C, but not at 28 degrees C, the optimal growth temperature of PG4180. Assuming that the highly hydrophobic N-terminus of CorS might be involved in temperature-signal perception, the membrane topology of CorS was determined using translational phoA and lacZ fusions, leading to a topological model for CorS with six transmembrane domains (TMDs). Interestingly, three PhoA fusions located downstream of the sixth TMD showed a thermoresponsive phenotype. Enzymic activity, immunoblot, and protease-sensitivity assays were performed to localize the CorS derivatives, to analyse the expression level of hybrid proteins and to examine the model. In-frame deletions of the last four, or all six TMDs gave rise to non-functional CorS. The results indicated that the transmembrane region is important for CorS to function as a temperature sensor, and that the membrane topology of CorS might be involved in signal perception. PMID- 15289569 TI - AlgR functions in algC expression and virulence in Pseudomonas syringae pv. syringae. AB - Pseudomonas syringae pv. syringae strain FF5 is a phytopathogen associated with a rapid dieback on ornamental pear trees. P. syringae and the human pathogen Pseudomonas aeruginosa produce the exopolysaccharide alginate, a copolymer of mannuronic and guluronic acid. In P. aeruginosa, the response regulator AlgR (AlgR1) is required for transcription of algC and algD, which encode key enzymes in the alginate biosynthetic pathway. In P. syringae FF5, however, algR is not required for the activation of algD. Interestingly, algR mutants of P. syringae remain nonmucoid, indicating an undefined role for this response regulator in alginate biosynthesis. In the current study, the algC promoter region was cloned from P. syringae pv. syringae strain FF5, and sequence analysis of the algC promoter indicated the presence of potential binding sites for AlgR and sigma(54), the alternative sigma factor encoded by rpoN. The algC promoter from P. syringae FF5 (PsalgC) was cloned upstream of a promoterless glucuronidase gene (uidA), and the PsalgC-uidA transcriptional fusion was used to monitor algC expression in strains FF5.32 (algR mutant of P. syringae FF5) and PG4180.K2 (rpoN mutant of P. syringae pv. glycinea PG4180). Expression of the PsalgC-uidA fusion was fourfold lower in both the algR and rpoN mutants as compared to respective wild-type strains, indicating that both AlgR and sigma(54) are required for full activation of algC transcription in P. syringae pv. syringae. AlgR from P. syringae was successfully overproduced in Escherichia coli as a C-terminal translational fusion to the maltose-binding protein (MBP). Gel shift experiments indicated that MBP-AlgR binds strongly to the algC promoter region. Biological assays demonstrated that the algR mutant was significantly impaired in both pathogenicity and epiphytic fitness as compared to the wild-type strain. These results, along with the gene expression studies, indicate that AlgR has a positive role in the activation of algC in P. syringae and contributes to both virulence and epiphytic fitness. Furthermore, the symptoms observed with wild type P. syringae FF5 suggest that this strain can move systemically in leaf tissue, and that a functional copy of algR is required for systemic movement. PMID- 15289570 TI - NblA is essential for phycobilisome degradation in Anabaena sp. strain PCC 7120 but not for development of functional heterocysts. AB - Phycobilisomes (PBS) are the major light-harvesting complexes of cyanobacteria. These usually blue-coloured multiprotein assemblies are rapidly degraded when the organisms are starved for combined nitrogen. This proteolytic process causes a colour change of the cyanobacterial cells from blue-green to yellow-green ('bleaching'). As is well documented for the unicellular, non-diazotrophic cyanobacteria Synechococcus elongatus PCC 7942 and Synechocystis sp. PCC 6803, a gene termed nblA plays a key role in PBS degradation. Filamentous, diazotrophic cyanobacteria like Anabaena adapt to nitrogen deprivation by differentiation of N(2)-fixing heterocysts. However, during the first hours after nitrogen deprivation all cells degrade their PBS. When heterocysts mature and nitrogenase becomes active, vegetative cells resynthesize their light-harvesting complexes while in heterocysts the phycobiliprotein content remains very low. Expression and function of nblA in Anabaena sp. PCC 7120 was investigated. This strain has two nblA homologous genes, one on the chromosome (nblA) and one on plasmid delta (nblA-p). Northern blot analysis indicated that only the chromosomal nblA gene is up-regulated upon nitrogen starvation. Mutants with interrupted nblA and nblA-p genes, respectively, grew on N(2) and developed functional heterocysts. Mutant DeltanblA-p behaved like the wild-type. However, mutant DeltanblA was unable to degrade its PBS, which was most obvious in non-bleaching heterocysts. The results show that NblA, encoded by the chromosomal nblA gene, is required for PBS degradation in Anabaena but is not essential for heterocyst differentiation. PMID- 15289571 TI - Biofilms promote altruism. AB - The origin of altruism is a fundamental problem in evolution, and the maintenance of biodiversity is a fundamental problem in ecology. These two problems combine with the fundamental microbiological question of whether it is always advantageous for a unicellular organism to grow as fast as possible. The common basis for these three themes is a trade-off between growth rate and growth yield, which in turn is based on irreversible thermodynamics. The trade-off creates an evolutionary alternative between two strategies: high growth yield at low growth rate versus high growth rate at low growth yield. High growth yield at low growth rate is a case of an altruistic strategy because it increases the fitness of the group by using resources economically at the cost of decreased fitness, or growth rate, of the individual. The group-beneficial behaviour is advantageous in the long term, whereas the high growth rate strategy is advantageous in the short term. Coexistence of species requires differences between their niches, and niche space is typically divided into four 'axes' (time, space, resources, predators). This neglects survival strategies based on cooperation, which extend the possibilities of coexistence, arguing for the inclusion of cooperation as the fifth 'axis'. Here, individual-based model simulations show that spatial structure, as in, for example, biofilms, is necessary for the origin and maintenance of this 'primitive' altruistic strategy and that the common belief that growth rate but not yield decides the outcome of competition is based on chemostat models and experiments. This evolutionary perspective on life in biofilms can explain long-known biofilm characteristics, such as the structural organization into microcolonies, the often-observed lack of mixing among microcolonies, and the shedding of single cells, as promoting the origin and maintenance of the altruistic strategy. Whereas biofilms enrich altruists, enrichment cultures, microbiology's paradigm for isolating bacteria into pure culture, select for highest growth rate. PMID- 15289572 TI - Heterologous production of the antifungal polyketide antibiotic soraphen A of Sorangium cellulosum So ce26 in Streptomyces lividans. AB - The antifungal polyketide soraphen A is produced by the myxobacterium Sorangium cellulosum So ce26. The slow growth, swarming motility and general intransigence of the strain for genetic manipulations make industrial strain development, large scale fermentation and combinatorial biosynthetic manipulation of the soraphen producer very challenging. To provide a better host for soraphen A production and molecular engineering, the biosynthetic gene cluster for this secondary metabolite was integrated into the chromosome of Streptomyces lividans ZX7. The upstream border of the gene cluster in Sor. cellulosum was defined by disrupting sorC, which is proposed to take part in the biosynthesis of methoxymalonyl coenzyme A, to yield a Sor. cellulosum strain with abolished soraphen A production. Insertional inactivation of orf2 further upstream of sorC had no effect on soraphen A production. The genes sorR, C, D, F and E thus implicated in soraphen biosynthesis were then introduced into an engineered Str. lividans strain that carried the polyketide synthase genes sorA and sorB, and the methyltransferase gene sorM integrated into its chromosome. A benzoate-coenzyme A ligase from Rhodopseudomonas palustris was also included in some constructs. Fermentations with the engineered Str. lividans strains in the presence of benzoate and/or cinnamate yielded soraphen A. Further feeding experiments were used to delineate the biosynthesis of the benzoyl-coenzyme A starter unit of soraphen A in the heterologous host. PMID- 15289573 TI - Characterization of a multicopper oxidase gene cluster in Phanerochaete chrysosporium and evidence of altered splicing of the mco transcripts. AB - A cluster of multicopper oxidase genes (mco1, mco2, mco3, mco4) from the lignin degrading basidiomycete Phanerochaete chrysosporium is described. The four genes share the same transcriptional orientation within a 25 kb region. mco1, mco2 and mco3 are tightly grouped, with intergenic regions of 2.3 and 0.8 kb, respectively, whereas mco4 is located 11 kb upstream of mco1. All are transcriptionally active, as shown by RT-PCR. Comparison of cDNAs and the corresponding genomic sequences identified 14-19 introns within each gene. Based on homology and intron composition, two subfamilies of mco sequences could be identified. The sequences have copper-binding motifs similar to ferroxidase proteins, but different from fungal laccases. Thus, these sequences constitute a novel branch of the multicopper oxidase family. Analysis of several cDNA clones obtained from poly(A) RNA revealed the presence of transcripts of various lengths. Splice variants from mco2, mco3 and mco4 were characterized. They generally exhibited the presence of one to five introns, whereas other transcripts lacked some exons. In all cases, the presence of introns leads to frame shifts that give rise to premature stop codons. In aggregate, these investigations show that P. chrysosporium possesses a novel family of multicopper oxidases which also feature clustering and incomplete processing of some of their transcripts, a phenomenon referred to in this paper as 'altered splicing'. PMID- 15289574 TI - Green fluorescent protein (GFP) as a vital marker for pathogenic development of the dermatophyte Trichophyton mentagrophytes. AB - Skin infections by dermatophytes of the genus Trichophyton are widespread, but methods to investigate the molecular basis of pathogenicity are only starting to be developed. The initial stages of growth on the host can only be studied by electron microscopy, which requires fixing the tissue. This paper shows that restriction-enzyme-mediated integration (REMI) provides stable expression of the green fluorescent protein (GFP) in a clinical isolate of Trichophyton mentagrophytes. Under control of a constitutively active fungal promoter, GFP renders the hyphae fluorescent both in culture and in a recently developed model using human skin explants. Stages of infection and penetration into the skin layers were visualized by confocal microscopy. The stages of infection can thus be followed using GFP as a vital marker, and this method will also provide, for the first time, a means to follow gene expression during infection of skin by dermatophyte fungi. PMID- 15289575 TI - AdoMet radical proteins--from structure to evolution--alignment of divergent protein sequences reveals strong secondary structure element conservation. AB - Eighteen subclasses of S-adenosyl-l-methionine (AdoMet) radical proteins have been aligned in the first bioinformatics study of the AdoMet radical superfamily to utilize crystallographic information. The recently resolved X-ray structure of biotin synthase (BioB) was used to guide the multiple sequence alignment, and the recently resolved X-ray structure of coproporphyrinogen III oxidase (HemN) was used as the control. Despite the low 9% sequence identity between BioB and HemN, the multiple sequence alignment correctly predicted all but one of the core helices in HemN, and correctly predicted the residues in the enzyme active site. This alignment further suggests that the AdoMet radical proteins may have evolved from half-barrel structures (alphabeta)4 to three-quarter-barrel structures (alphabeta)6 to full-barrel structures (alphabeta)8. It predicts that anaerobic ribonucleotide reductase (RNR) activase, an ancient enzyme that, it has been suggested, serves as a link between the RNA and DNA worlds, will have a half barrel structure, whereas the three-quarter barrel, exemplified by HemN, will be the most common architecture for AdoMet radical enzymes, and fewer members of the superfamily will join BioB in using a complete (alphabeta)8 TIM-barrel fold to perform radical chemistry. These differences in barrel architecture also explain how AdoMet radical enzymes can act on substrates that range in size from 10 atoms to 608 residue proteins. PMID- 15289576 TI - Quantitative modeling of DNA-protein interactions: effects of amino acid substitutions on binding specificity of the Mnt repressor. AB - Understanding DNA-protein recognition quantitatively is essential to developing computational algorithms for accurate transcriptional binding site prediction. Using a quantitative, multiple fluorescence, relative affinity (QuMFRA) assay, we determine the binding specificity of 11 different position 6 variants of the Mnt repressor for operators containing all 16 possible dinucleotides at operator positions 16 and 17. We show that the wild-type and all variant proteins interact with the two positions in a non-independent manner, but that a simple independent model provides a close approximation to the true binding affinities. The wild type His at amino acid 6 is the only protein to prefer the AC sequence of the wild-type operator, whereas most of the variant proteins prefer TA. H6R is unique in having a strong preference for C at position 16. A comparison of the quantitative binding data for all of the protein variants with a model for recognition of the early growth response (EGR) zinc finger family suggests that interactions of Mnt with positions 16 and 17 are similar to interactions of EGR with positions 1 and 2, respectively. This information leads to an augmented model for the interaction of Mnt with its operator. PMID- 15289577 TI - Integron integrase binds to bulged hairpin DNA. AB - Gene cassettes are short, monogenic DNA elements that translocate between integrons through site-specific excision and integration. These events require that an integron-coded tyrosine recombinase forms a reactive complex with two sites, at least one of which belongs to the attC class. An attC site can be divided into two pairs of short repeats flanking a palindromic central region. The nucleotide sequence of attC among different cassettes varies extensively, implying that the site contains a structural recognition determinant with low sequence constraints. Oligonucleotides representing many different sequence modifications in either strand of the site were examined for integrase binding by using an electrophoresis mobility shift assay. The inner repeats, a central triplet and two single-nucleotide asymmetries in the site had the strongest influence on binding strength and strand choice. Our data show that the recombinase binds to a bulged hairpin in attC and that the hairpin distortion due to bulging could define the appropriate orientation of the otherwise symmetrical site. This is consistent with the strong bias for binding of recombinase to the bottom-strand oligonucleotides in vitro. Moreover, it was observed that the mobility-shifted complexes persisted under protein-denaturing assay conditions, indicating that a covalent link is indeed formed between integrase and DNA. Upon substitution of the presumed DNA-attacking residue, Y312, with a phenylalanine, DNA binding remained but there was no covalent linkage. PMID- 15289578 TI - Origin of the intrinsic rigidity of DNA. AB - The intrinsic rigidities of DNA and RNA helices are generally thought to arise from some combination of vertical base-stacking interactions and intra-helix phosphate-phosphate charge repulsion; however, the relative contributions of these two types of interaction to helix rigidity have not been quantified. To address this issue, we have measured the rotational decay times of a 'gapped duplex' DNA molecule possessing a central, single-stranded region, dT24, before and after addition of the free purine base, N6-methyladenine ((me)A). Upon addition of (me)A, the bases pair with the T residues, forming a continuous stack within the gap region. Formation of the gapped duplex is accompanied by a nearly 2-fold increase in decay time, to values that are indistinguishable from the full duplex control for monovalent salt concentrations up to 90 mM. These results indicate that at least 90% of the rigidity of the dT(n)-dA(n) homopolymer derives from base pair stacking effects, with phosphate-phosphate interactions contributing relatively little to net helix rigidity at moderate salt concentrations. PMID- 15289579 TI - Enhanced nucleic acid binding to ATP-bound hepatitis C virus NS3 helicase at low pH activates RNA unwinding. AB - The molecular basis of the low-pH activation of the helicase encoded by the hepatitis C virus (HCV) was examined using either a full-length NS3 protein/NS4A cofactor complex or truncated NS3 proteins lacking the protease domain, which were isolated from three different viral genotypes. All proteins unwound RNA and DNA best at pH 6.5, which demonstrate that conserved NS3 helicase domain amino acids are responsible for low-pH enzyme activation. DNA unwinding was less sensitive to pH changes than RNA unwinding. Both the turnover rate of ATP hydrolysis and the K(m) of ATP were similar between pH 6 and 10, but the concentration of nucleic acid needed to stimulate ATP hydrolysis decreased almost 50-fold when the pH was lowered from 7.5 to 6.5. In direct-binding experiments, HCV helicase bound DNA weakly at high pH only in the presence of the non hydrolyzable ATP analog, ADP(BeF3). These data suggest that a low-pH environment might be required for efficient HCV RNA translation or replication, and support a model in which an acidic residue rotates toward the RNA backbone upon ATP binding repelling nucleic acid from the binding cleft. PMID- 15289580 TI - Cleavage of deoxyoxanosine-containing oligodeoxyribonucleotides by bacterial endonuclease V. AB - Oxanine (O) is a deamination product derived from guanine with the nitrogen at the N1 position substituted by oxygen. Cytosine, thymine, adenine, guanine as well as oxanine itself can be incorporated by Klenow Fragment to pair with oxanine in a DNA template with similar efficiency, indicating that oxanine in DNA may cause various mutations. As a nucleotide, deoxyoxanosine may substitute for deoxyguanosine to complete a primer extension reaction. Endonuclease V, an enzyme known for its enzymatic activity on uridine-, inosine- and xanthosine-containing DNA, can cleave oxanosine-containing DNA at the second phosphodiester bond 3' to the lesion. Mg2+ or Mn2+, and to a small extent Co2+ or Ni2+, support the oxanosine-containing DNA cleavage activity. All four oxanosine-containing base pairs (A/O, T/O, C/O and G/O) were cleaved with similar efficiency. The cleavage of double-stranded oxanosine-containing DNA was approximately 6-fold less efficient than that of double-stranded inosine-containing DNA. Single-stranded oxanosine-containing DNA was cleaved with a lower efficiency as compared with double-stranded oxanosine-containing DNA. A metal ion enhances the binding of endonuclease V to double-stranded and single-stranded oxanosine-containing DNA 6- and 4-fold, respectively. Hypothetic models of oxanine-containing base pairs and deaminated base recognition mechanism are presented. PMID- 15289582 TI - 'You have to know a little in order to know what you don't know'. PMID- 15289581 TI - Single amino acid changes in AspRS reveal alternative routes for expanding its tRNA repertoire in vivo. AB - Aminoacyl-tRNA synthetases (aaRSs) are enzymes that are highly specific for their tRNA substrates. Here, we describe the expansion of a class IIb aaRS-tRNA specificity by a genetic selection that involves the use of a modified tRNA displaying an amber anticodon and the argE(amber) and lacZ(amber) reporters. The study was performed on Escherichia coli aspartyl-tRNA synthetase (AspRS) and amber tRNA(Asp). Nine AspRS mutants able to charge the amber tRNA(Asp) and to suppress the reporter genes were selected from a randomly mutated library. All the mutants exhibited a new amber tRNA(Asp) specificity in addition to the initial native tRNA(Asp). Six mutations were found in the anticodon-binding site located in the N-terminal OB-fold. The strongest suppressor was a mutation of residue Glu-93 that contacts specifically the anticodon nucleotide 34 in the crystal structure. The other mutations in the OB-fold were found at close distance from the anticodon in the so-called loop L45 and strand S1. They concern residues that do not contact tRNA(Asp) in the native complex. In addition, this study shows that suppressors can carry mutations located far from the anticodon binding site. One such mutation was found in the synthetase hinge-module where it increases the tRNA(Asp)-charging rate, and two other mutations were found in the prokaryotic-specific insertion domain and the catalytic core. These mutants seem to act by indirect effects on the tRNA acceptor stem binding and on the conformation of the active site of the enzyme. Altogether, these data suggest the existence of various ways for modifying the mechanism of tRNA discrimination. PMID- 15289583 TI - Occupational health hazards in mining: an overview. AB - This review article outlines the physical, chemical, biological, ergonomic and psychosocial occupational health hazards of mining and associated metallurgical processes. Mining remains an important industrial sector in many parts of the world and although substantial progress has been made in the control of occupational health hazards, there remains room for further risk reduction. This applies particularly to traumatic injury hazards, ergonomic hazards and noise. Vigilance is also required to ensure exposures to coal dust and crystalline silica remain effectively controlled. PMID- 15289584 TI - Noise-induced hearing loss and hearing conservation in mining. AB - BACKGROUND: Noise exposure is prevalent in mining, and as the prevalence of noise induced hearing loss has not changed much in the past two decades, hearing conservation is an important issue. OBJECTIVES: To review the literature and highlight important developments in the field. METHODS: A review of the literature using OVID as the primary search engine, using the search terms as: noise, occupational; hearing loss, noise induced; ear protective devices; and mining. RESULTS: A total of 66 articles were found, but only 11 were in the English language and few were published in the past 10 years. This is disappointing, because neither noise exposure nor the consequent risk of noise induced hearing loss seems to have changed much in the past 20 years. Noise is, however, a generic hazard, and this article reviews current best practice in prevention. PMID- 15289585 TI - Ergonomics in mining. AB - Like other areas of occupational health and safety (OHS) ergonomics is evolving and becoming more integrated into overall work management systems. As we learn more about the complex interaction between psychosocial and physical factors in the aetiology of work-related illness and injury the more we rely on managers to 'get it right' if we are to prevent these conditions. Risks to health and safety in the mining industry posed by longer shift lengths, higher work loads, less task variation and decision latitude have not really been well researched. Heavy physical workloads and stresses are still areas of concern, but are likely to be intermittent rather than constant. Recent research confirms current thinking rather than shedding new light on the subject. The contribution of slips, trips and falls and increasing age of miners to manual handling injuries is still not clear. In some cases sedentary work and the operation of machinery has completely replaced heavy physical work. The issues of machinery design for operations and maintenance and whole-body vibration exposures when operating machines and vehicles are becoming more critical. The link between prolonged sitting, poor cab design and vibration with back and neck pain is being recognized but has yet to be addressed in any systematic way by the mining industry. On the plus side some mining companies have well-developed participative approaches to problem solving and these need to be extended to areas such as ergonomics. PMID- 15289586 TI - Occupational respiratory disease in mining. AB - This review is based on research-based literature on occupational lung disease in the mining and related industries, focusing on conditions of public health importance arising from asbestos, coal and silica exposure. Both 'traditional' and 'new' concerns about occupational respiratory disease in miners are addressed, with the inclusion of practical evidence-based findings relevant to practitioners working in developed and developing countries. Mining is not a homogeneous industry since current miners work in formal and informal operations with numerous, and often multiple, air-borne exposures. A further occupational health challenge facing primary care practitioners are ex-miners presenting with disease only after long latency. The sequelae of silica exposure remain an occupational health priority, particularly for practitioners who serve populations with concomitant HIV and tuberculosis infection and even when exposure is apparently below the statutory occupational exposure level. Coal workers' pneumoconiosis, asbestos related diseases, lung cancer and other occupational respiratory diseases remain of considerable importance even after mining operations cease. While mining exposures contribute significantly to lung disease, smoking is a major factor in the development of lung cancer and chronic obstructive airways disease necessitating a comprehensive approach for prevention and control of mining-related occupational lung disease. PMID- 15289587 TI - Occupational safety risk management in Australian mining. AB - In the past 15 years, there has been a major safety improvement in the Australian mining industry. Part of this change can be attributed to the development and application of risk assessment methods. These systematic, team-based techniques identify, assess and control unacceptable risks to people, assets, the environment and production. The outcomes have improved mine management systems. This paper discusses the risk assessment approach applied to equipment design and mining operations, as well as the specific risk assessment methodology. The paper also discusses the reactive side of risk management, incident and accident investigation. Systematic analytical methods have also been adopted by regulatory authorities and mining companies to investigate major losses. PMID- 15289588 TI - Attitudes and access to electronic exchange of information on occupational disease. AB - BACKGROUND: THOR is a network of work-related disease surveillance schemes dependent on volunteer case reporting by medical specialists. Data collection and dissemination has hitherto been paper-based. AIMS: To elicit the opinion of existing reporters in THOR on electronic exchange of information and to assess the practical capabilities of the same reporters to participate in electronic communication. METHODS: A mail-based questionnaire of randomly selected THOR reporters using closed format questions. RESULTS: The response rate to the questionnaire was 66% (253/383). Almost half (47%) of the responders wanted dissemination of information solely in an electronic form, 35% favoured paper based reports, while 16% wanted both paper and electronic reports. Two-thirds (66%) would make use of electronic archives of reports and 59% would use this facility to resolve questions by accessing accumulated data. The majority (82%) read e-mail more than once a week and 34% browsed the web as frequently. However, 5% did not have e-mail and 6% never browsed the web. Most responders judged their internet connectivity to be rapid (68%) and convenient (83%), and 91% could receive e-mail attachments. CONCLUSIONS: Most responders have the skills and infrastructure required to engage in electronic information exchange, and are favourably disposed to electronic means of communication. However it is also relevant to note that one-third of responders have a preference for the existing paper-based system. PMID- 15289589 TI - Bladder tumours in rubber workers: a factory study 1946-1995. AB - BACKGROUND: Prior to December 1949, some British rubber industry workers were inadvertently exposed to the human bladder carcinogen beta-naphthylamine, which was present as a contaminant (at 0.25%) in antioxidants used in manufacturing. This study follows a composite cohort of 6450 men employed at a large tyre factory either during the 'at-risk' period or just after it. METHODS: A group of 2090 at-risk men (employed 1945-1949) and 3038 men, first employed only after January 1950, when the carcinogen had been removed, were followed for their bladder cancer morbidity and mortality experiences. RESULTS: Fifty-eight tumours were registered for those at risk, whereas only 33.9 were expected at national standardized registration rates [SRRN = 171 and 95% confidence interval (CI) = 130-221]. Thirty-nine bladder tumours were reported for the post-1950 intake, whereas 38.3 were expected (SRRN = 102 and 95% CI = 72-139). The use of mortality data did not reveal any underlying hazard because 12 of the 58 at-risk workers with tumours were still alive at the study end date. In only 16 instances was bladder cancer actually certified as the underlying cause of death. Plotting cases by their location of work on a factory plan assisted the interpretation. CONCLUSIONS: A statistically significant elevated risk of bladder cancer for the exposed workforce was evident, but this reversed when the carcinogen was removed from processing in October 1949. The use of morbidity (incidence) data in long term studies of occupational bladder cancer should be the required methodology if the hazard and risk are not to be underestimated. PMID- 15289590 TI - A study of mortality patterns at a tyre factory 1951-1985: a reference statistic dilemma. AB - BACKGROUND: The general and cancer mortalities of rubber workers at a large tyre factory were studied in an area of marked regional variation in death rates. METHODS: Three quinquennial intakes of male rubber workers engaged between January 1946 and December 1960 formed a composite cohort of 6454 men to be followed up. Over 99% were successfully traced by December 1985. The cohort analysis used both national and local rates as reference statistics for several causes. RESULTS: Between 1951 and 1985, a national standardized mortality ratio (SMRN) of 101 for all causes (based on 2556 deaths) was noted, whereas the local standardized mortality ratio (SMRL) was only 79. For all cancers, the figures were 115 (SMRN) and 93 (SMRL), for stomach cancer they were 137 (SMRN) and 84 (SMRL), and for lung cancer they were 121 (SMRN) and 94 (SMRL). No outright excesses against the national norm were observed for other cancers except for larynx, brain and central nervous system and thyroid cancer and the leukaemias. Excesses were statistically significant for cancer of the gallbladder and the bile ducts, for silicotuberculosis (SMRN = 1000) and for the pneumoconioses (SMRN = 706). Deaths from cerebrovascular diseases, chronic bronchitis and emphysema showed statistically significant deficits using either norm. CONCLUSIONS: These results from a large factory cohort study of rubber workers, followed for over three decades, demonstrate the marked discrepancy that can result from using only one reference statistic in areas of significant variation in mortality patterns. PMID- 15289591 TI - Meta-analysis in occupational epidemiology: a review of practice. AB - OBJECTIVES: To describe past practice in meta-analyses found in occupational epidemiology, identifying the major issues that should be considered by researchers planning a meta-analysis in this setting. METHODS: An electronic search of relevant online databases was undertaken. Papers were included in the review if they contained a statistical synthesis of risks in an occupational health setting. RESULTS: Sixty reports of meta-analyses were identified, mostly in cancer. The number of meta-analyses has increased consistently over the last 20 years. A majority of studies focused on a mean overall effect, although more than half of them also investigated heterogeneity of results. Both fixed effect and random effects meta-analysis models were employed, the former more often, and in eight studies used despite a statistically significant test for heterogeneity. A large proportion of the meta-analyses included different effect measures in the statistical synthesis, for example, including standardized mortality ratios (SMRs) and standardized incidence ratios. Most meta-analyses limited to a single type of effect measure focused on SMRs. The vast majority of meta-analyses combined all studies regardless of variation in the extent of information on exposures. CONCLUSIONS: Meta-analyses in occupational epidemiology should properly explore and incorporate heterogeneity among studies. The meta-SMR is an important construct in this field, evidenced by a large proportion of cohort studies in the meta-analyses we identified. Controversy remains over the definition and validity of the meta-SMR. In addition, several other issues, notably dealing with heterogeneity in exposure, warrant further consideration. PMID- 15289592 TI - An update of a systematic review of controlled clinical trials on the primary prevention of back pain at the workplace. AB - OBJECTIVE: To update the evidence on the effectiveness of lumbar supports, education and exercise in the primary prevention of low back pain at the workplace. METHODS: A computerized search for controlled clinical trials published between 1997 and 2002 was conducted, and the methodological quality of the studies was assessed using a criteria list. The available evidence was graded with a rating system for the level of evidence. Effect sizes of individual studies were combined if the studies were sufficiently similar. RESULTS: Five new papers were identified for the update. These trials were added to the previously available trials (n = 11). The methodological quality of most studies was low. Since three of four RCTs on lumbar supports reported no effect, there is no evidence for the effectiveness of lumbar supports. No evidence for education could be found either, since all six RCTs showed negative results. The four RCTs on exercise consistently reported a positive effect, indicating limited evidence for the effectiveness of exercise. CONCLUSION: There is no evidence for the effectiveness of lumbar supports or education in the primary prevention of low back pain at the workplace. There is limited evidence for the efficacy of exercise, and the effect that can be obtained is moderate. There is still a need for methodologically sound studies and studies on the cost-effectiveness of exercise. Also the possible effect of lumbar supports in the treatment of back pain needs further investigation. PMID- 15289593 TI - Computerized cognitive behavioural therapy at work: a randomized controlled trial in employees with recent stress-related absenteeism. AB - BACKGROUND: Emotional distress has major implications for employees and employers. Cognitive behavioural therapy (CBT) is a recommended treatment, but demand outstrips supply. CBT is well suited to computerization. Most employee assistance programmes have not been systematically evaluated and computerized CBT has not previously been studied in the workplace. AIMS: To evaluate the effect of an 8 week computerized cognitive behavioural therapy programme, 'Beating The Blues', on emotional distress in employees with recent stress-related absenteeism, and to explore the reasons for non-participation. METHODS: An open, randomized trial in a London NHS occupational health department. Forty-eight public sector employees, with 10 or more cumulative days stress-related absenteeism in the last 6 months, randomized equally to 'Beating The Blues' plus conventional care, or conventional care alone. Main outcome measures were Hospital Anxiety and Depression Scale and Attributional Style Questionnaire scores at end of treatment and 1, 3 and 6 months later; and reasons for non participation. RESULTS: At end of treatment and 1 month later, adjusted mean depression scores and adjusted mean negative attributional style scores were significantly lower in the intervention group. One month post-treatment, adjusted mean anxiety scores were also significantly lower in the intervention group. The differences were not statistically significant at 3 and 6 months post-treatment. Non-participation was common and related to access problems, preference for other treatments, time commitment, scepticism about the intervention and the employer connection. CONCLUSIONS: 'Beating The Blues' may accelerate psychological recovery in employees with recent stress-related absenteeism. Greater flexibility and accessibility might improve uptake. PMID- 15289594 TI - Preventing baker's asthma. PMID- 15289595 TI - Monitor. The role of physical activity on health. PMID- 15289596 TI - Monitor. How often as occupational physicians do we see people with low back pain remain off work whilst waiting for magnetic resonance imaging (MRI)? PMID- 15289597 TI - How cholesterol homeostasis is regulated by plasma membrane cholesterol in excess of phospholipids. AB - How do cells sense and control their cholesterol levels? Whereas most of the cell cholesterol is located in the plasma membrane, the effectors of its abundance are regulated by a small pool of cholesterol in the endoplasmic reticulum (ER). The size of the ER compartment responds rapidly and dramatically to small changes in plasma membrane cholesterol around the normal level. Consequently, increasing plasma membrane cholesterol in vivo from just below to just above the basal level evoked an acute (<2 h) and profound ( approximately 20-fold) decrease in ER 3 hydroxy-3-methylglutaryl-CoA reductase activity in vitro. We tested the hypothesis that the sharply inflected ER response to cholesterol is governed by the thermodynamic activity (fugacity) of plasma membrane cholesterol. The following two independent measures of plasma membrane cholesterol activity in human red cells and fibroblasts were used: susceptibility to cholesterol oxidase and cholesterol transfer to cyclodextrin. Both indicators revealed a threshold at the physiologic set point of plasma membrane cholesterol. Incrementing the phospholipid compartment in the plasma membrane with lysophosphatidylcholine, previously shown to decrease cholesterol oxidase susceptibility, reduced the transfer of plasma membrane cholesterol to cyclodextrin and to the ER. Conversely, the membrane intercalator, n-octanol, increased cholesterol oxidation, transfer, and ER pool size, perhaps by displacing cholesterol from plasma membrane phospholipids. We conclude that the activity of the fraction of cholesterol in excess of other plasma membrane lipids sets the cholesterol level in the ER. Cholesterol-sensitive elements therein respond by nulling the active plasma membrane pool, thereby keeping the cholesterol matched to the other plasma membrane lipids. PMID- 15289598 TI - Inhibitor design by wrapping packing defects in HIV-1 proteins. AB - Two viral proteins, HIV-1 protease and HIV-1 integrase, have been targeted for inhibitor design to prevent assembly and maturation of HIV-1 virions. The enzymatic mechanism of these proteins involves side-chain groups that serve as general acids or bases. Furthermore, catalytic activity requires that water be removed from the microenvironment surrounding the chemical reaction site or be constrained to serve as an activated nucleophile. Here, we identify previously unrecognized structural features that promote water removal from polar catalytic regions. Packing defects in the form of hydrogen bonds that are insufficiently dehydrated intramolecularly, named "dehydrons," are strategically placed in the structure to induce an anhydrous enzymatic pathway. Dehydrons become electrostatically enhanced and stabilized upon further desolvation. Thus, packing defects act synergistically with the polar active groups to enhance the enzymatic electrostatics. However, because dehydrons are sticky, they constitute targets for inhibitor design. We noticed that inhibitors attach to polar surfaces by further desolvating dehydrons, thus blocking the active sites or the sites involved in harnessing the substrate. The dehydrons are thus required for functional reasons, making them suitable targets. The differences in success when targeting HIV-1 protease, feline immunodeficiency virus protease, and HIV-1 integrase are rationalized in terms of the dehydron distribution, revealing possible improvements in the targeting strategy. Principles of design optimization are proposed to create an inhibitor that can be neutralized only at the expense of the loss of catalytic function. The possibility of using drugs that wrap dehydrons to block protein-protein associations is also discussed. PMID- 15289599 TI - Marek's disease virus-encoded Meq gene is involved in transformation of lymphocytes but is dispensable for replication. AB - Marek's disease virus (MDV) causes an acute lymphoproliferative disease in chickens, resulting in T cell lymphomas in visceral organs and peripheral nerves. Earlier studies have determined that the repeat regions of oncogenic serotype 1 MDV encode a basic leucine zipper protein, Meq, which structurally resembles the Jun/Fos family of transcriptional activators. Meq is consistently expressed in MDV-induced tumor cells and has been suggested as the MDV-associated oncogene. To study the function of Meq, we have generated an rMd5DeltaMeq virus by deleting both copies of the meq gene from the genome of a very virulent strain of MDV. Growth curves in cultured fibroblasts indicated that Meq is dispensable for in vitro virus replication. In vivo replication in lymphoid organs and feather follicular epithelium was also not impaired, suggesting that Meq is dispensable for lytic infection in chickens. Reactivation of the rMd5DeltaMeq virus from peripheral blood lymphocytes was reduced, suggesting that Meq is involved but not essential for latency. Pathogenesis experiments showed that the rMd5DeltaMeq virus was fully attenuated in chickens because none of the infected chickens developed Marek's disease-associated lymphomas, suggesting that Meq is involved in lymphocyte transformation. A revertant virus that restored the expression of the meq gene, showed properties similar to those of the parental virus, confirming that Meq is involved in transformation but not in lytic replication in chickens. PMID- 15289600 TI - Targeting cell division: small-molecule inhibitors of FtsZ GTPase perturb cytokinetic ring assembly and induce bacterial lethality. AB - FtsZ, the ancestral homolog of eukaryotic tubulins, is a GTPase that assembles into a cytokinetic ring structure essential for cell division in prokaryotic cells. Similar to tubulin, purified FtsZ polymerizes into dynamic protofilaments in the presence of GTP; polymer assembly is accompanied by GTP hydrolysis. We used a high-throughput protein-based chemical screen to identify small molecules that target assembly-dependent GTPase activity of FtsZ. Here, we report the identification of five structurally diverse compounds, named Zantrins, which inhibit FtsZ GTPase either by destabilizing the FtsZ protofilaments or by inducing filament hyperstability through increased lateral association. These two classes of FtsZ inhibitors are reminiscent of the antitubulin drugs colchicine and Taxol, respectively. We also show that Zantrins perturb FtsZ ring assembly in Escherichia coli cells and cause lethality to a variety of bacteria in broth cultures, indicating that FtsZ antagonists may serve as chemical leads for the development of new broad-spectrum antibacterial agents. Our results illustrate the utility of small-molecule chemical probes to study FtsZ polymerization dynamics and the feasibility of FtsZ as a novel therapeutic target. PMID- 15289601 TI - Scaling theory for quasibrittle structural failure. AB - This inaugural article has a twofold purpose: (i) to present a simpler and more general justification of the fundamental scaling laws of quasibrittle fracture, bridging the asymptotic behaviors of plasticity, linear elastic fracture mechanics, and Weibull statistical theory of brittle failure, and (ii) to give a broad but succinct overview of various applications and ramifications covering many fields, many kinds of quasibrittle materials, and many scales (from 10(-8) to 10(6) m). The justification rests on developing a method to combine dimensional analysis of cohesive fracture with second-order accurate asymptotic matching. This method exploits the recently established general asymptotic properties of the cohesive crack model and nonlocal Weibull statistical model. The key idea is to select the dimensionless variables in such a way that, in each asymptotic case, all of them vanish except one. The minimal nature of the hypotheses made explains the surprisingly broad applicability of the scaling laws. PMID- 15289602 TI - Breed distribution and history of canine mdr1-1Delta, a pharmacogenetic mutation that marks the emergence of breeds from the collie lineage. AB - A mutation in the canine multidrug resistance gene, MDR1, has previously been associated with drug sensitivities in two breeds from the collie lineage. We exploited breed phylogeny and reports of drug sensitivity to survey other purebred populations that might be genetically at risk. We found that the same allele, mdr1-1Delta, segregated in seven additional breeds, including two sighthounds that were not expected to share collie ancestry. A mutant haplotype that was conserved among affected breeds indicated that the allele was identical by descent. Based on breed histories and the extent of linkage disequilibrium, we conclude that all dogs carrying mdr1-1Delta are descendants of a dog that lived in Great Britain before the genetic isolation of breeds by registry (ca. 1873). The breed distribution and frequency of mdr1-1Delta have applications in veterinary medicine and selective breeding, whereas the allele's history recounts the emergence of formally recognized breeds from an admixed population of working sheepdogs. PMID- 15289603 TI - A single-amino-acid lid renders a gas-tight compartment within a membrane-bound transporter. AB - Proteins undergo structural fluctuations between nearly isoenergetic substates. Such fluctuations are often intimately linked with the functional properties of proteins. However, in some cases, such as in transmembrane ion transporters, the control of the ion transport requires that the protein is designed to restrict the motions in specific regions. In this study, we have investigated the dynamics of a membrane-bound respiratory oxidase, which acts both as an enzyme catalyzing reduction of O(2) to H(2)O and as a transmembrane proton pump. The segment of the protein where proton translocation is controlled ("gating" region) overlaps with a channel through which O(2) is delivered to the catalytic site. We show that the replacement of an amino acid residue with a small side chain (Gly) by one with a larger side chain (Val), in a narrow part of this channel, completely blocks the O(2) access to the catalytic site and results in formation of a compartment around the site that is impermeable to small gas molecules. Thus, the protein motions cannot counter the blockage introduced by the mutation. These results indicate that the protein motions are restricted in the proton-gating region and that rapid O(2) delivery to the catalytic site requires a gas channel, which is confined within a rigid protein body. PMID- 15289605 TI - The mouse juvenile spermatogonial depletion (jsd) phenotype is due to a mutation in the X-derived retrogene, mUtp14b. AB - The recessive juvenile spermatogonial depletion (jsd) mutation results in a single wave of spermatogenesis, followed by failure of type A spermatogonia to differentiate, resulting in adult male sterility. We have identified a jsd specific rearrangement in the mouse homologue of the Saccharomyces cerevisiae gene UTP14, termed mUtp14b. Confirmation that mUtp14b underlies the jsd phenotype was obtained by transgenic bacterial artificial chromosome (BAC) rescue. We also identified a homologous gene on the Mus musculus X chromosome (MMUX) (mUtp14a) that is the strict homologue of the yeast gene, from which the intronless mUtp14b has been derived by retrotransposition. Expression analysis showed that mUtp14b is predominantly expressed in the germ line of the testis from zygotene through round spermatids, whereas mUtp14a, although well expressed in all somatic tissues, could be detected only in the germ line in round spermatids. In yeast, depletion of the UTP proteins impedes production of 18S rRNA, leading to cell death. We propose that the retroposed autosomal copy mUtp14b, having acquired a testis-specific expression pattern, could have provided a mechanism for increasing the efficiency and/or numbers of germ cells produced by meeting the need for more 18S rRNA and protein. Such a mechanism would be of obvious reproductive advantage and be strongly selected for in evolution. Consistent with this hypothesis is the finding of a similar X-autosome retroposition of UTP14 in human which seems to have arisen independently of that in rodents. In jsd homozygotes, which lack a functional copy of Utp14b, insufficient production of rRNA quickly leads to a cessation of spermatogenesis. PMID- 15289604 TI - Advanced glycation endproduct (AGE) receptor 1 is a negative regulator of the inflammatory response to AGE in mesangial cells. AB - Advanced glycation endproducts (AGE) contribute to kidney disease due to diabetes or aging by means of mesangial cell (MC) receptors, such as the receptor for AGE (RAGE), which promote oxidant-stress-dependent NF-kappaB activation and inflammatory gene expression. MC also express scavenger receptors SR-I and SR-II and AGE receptors 1, 2, and 3 (AGE-R1, -R2, and -R3), some of which are linked to AGE turnover. Because AGE-R1 expression is found suppressed in severe diabetic kidney disease, as other receptors increase, we investigated whether his molecule has a protective role against AGE-induced MC injury. A stable murine MC line overexpressing AGE-R1 (R1-MC) was generated, exhibiting a 1.8- to 2.7-fold increase in (125)I-AGE-specific binding, uptake, and degradation, compared with mock-MC. However, AGE-stimulated NF-kappaB activity and mitogen-activated protein kinase (MAPK) (p44/42) phosphorylation were found markedly suppressed in R1-MC. Additionally, AGE-stimulated macrophage chemotaxis protein 1 and RAGE overexpression were abolished in R1-MC. The effect of R1 on RAGE signaling was investigated after overexpressing RAGE in Chinese hamster ovary cells, which lack RAGE. AGE stimulation elicited NF-kappaB and MAPK activities in RAGE-Chinese hamster ovary cells; however, after cotransfection with R1, these responses were suppressed. Also, after silencing endogenous R1 in wild-type MC by R1 small interfering RNA, AGE-mediated MAPK/p44/42 activation exceeded by >2-fold that of mock-MC, consistent with loss of the activation-inhibitory properties of native AGE-R1. AGE-R1, although enhancing AGE removal, is also a distinct receptor in that it suppresses AGE-mediated MC inflammatory injury through negative regulation of RAGE, a previously uncharacterized pathway that may protect renal and other tissue injury due to diabetes and aging. PMID- 15289606 TI - In different organisms, the mode of interaction between two signaling proteins is not necessarily conserved. AB - Although interfaces mediating protein-protein interactions are thought to be under strong evolutionary constraints, binding of the chemotaxis histidine kinase CheA to its phosphorylation target CheY suggests otherwise. The structure of Thermotoga maritima CheA domain P2 in complex with CheY reveals a different association than that observed for the same Escherichia coli proteins. Similar regions of CheY bind CheA P2 in the two systems, but the CheA P2 domains differ by an approximately 90 degrees rotation. CheA binds CheY with identical affinity in T. maritima and E. coli at the vastly different temperatures where the respective organisms live. Distinct sets of P2 residues mediate CheY binding in the two complexes; conservation patterns of these residues in CheA and compensations in CheY delineate two families of prokaryotic chemotaxis systems. A protein complex that has the same components and general function in different organisms, but an altered structure, indicates unanticipated complexity in the evolution of protein-protein interactions and cautions against extrapolating structural data from homologs. PMID- 15289607 TI - The mouse kinome: discovery and comparative genomics of all mouse protein kinases. AB - We have determined the full protein kinase (PK) complement (kinome) of mouse. This set of 540 genes includes many novel kinases and corrections or extensions to >150 published sequences. The mouse has orthologs for 510 of the 518 human PKs. Nonorthologous kinases arise only by retrotransposition and gene decay. Orthologous kinase pairs vary in sequence conservation along their length, creating a map of functionally important regions for every kinase pair. Many species-specific sequence inserts exist and are frequently alternatively spliced, allowing for the creation of evolutionary lineage-specific functions. Ninety seven kinase pseudogenes were found, all distinct from the 107 human kinase pseudogenes. Chromosomal mapping links 163 kinases to mutant phenotypes and unlocks the use of mouse genetics to determine functions of orthologous human kinases. PMID- 15289608 TI - Evolvability is a selectable trait. AB - Concomitant with the evolution of biological diversity must have been the evolution of mechanisms that facilitate evolution, because of the essentially infinite complexity of protein sequence space. We describe how evolvability can be an object of Darwinian selection, emphasizing the collective nature of the process. We quantify our theory with computer simulations of protein evolution. These simulations demonstrate that rapid or dramatic environmental change leads to selection for greater evolvability. The selective pressure for large-scale genetic moves such as DNA exchange becomes increasingly strong as the environmental conditions become more uncertain. Our results demonstrate that evolvability is a selectable trait and allow for the explanation of a large body of experimental results. PMID- 15289609 TI - Evolutionary genomics of ecological specialization. AB - We used a combination of genomic techniques to monitor chromosomal evolution across hundreds of generations as Escherichia coli adapted to growth-limiting concentrations of either lactulose, methyl-galactoside, or a 72:28 mixture of the two. DNA microarrays identified 8 unique duplications and 16 unique deletions among 42 evolvants from 23 chemostat experiments. Each mutation was confirmed by sequencing PCR-amplified flanking genomic DNA and, except for one deletion, an insertion sequence was found at the break point. vPCR of insertion sequences identified these same mutations and 16 additional insertions (all confirmed by sequencing). The pattern of genomic evolution is highly reproducible. Statistical analyses show that duplications at lac and mutations in mgl are adaptations specific to lactulose and to methyl-galactoside, respectively. Adaptation to mixed sugars is characterized by similar mutations, but lac duplications and mgl mutations usually arise in different backgrounds, producing ecological specialists for each sugar. This suggests that an antagonistic pleiotropic tradeoff between duplications at lac and mutations in mgl retards the evolution of generalists. Other mutations that repeatedly appear in replicate experiments are adaptations to the chemostat environment and are not specific to one or the other sugar. PMID- 15289610 TI - Dose-dependent response of FGF-2 for lymphangiogenesis. AB - Spatio-temporal studies on the growth of capillary blood vessels and capillary lymphatic vessels in tissue remodeling have suggested that lymphangiogenesis is angiogenesis-dependent. We revisited this concept by using fibroblast growth factor 2 (FGF-2) (80 ng) to stimulate the growth of both vessel types in the mouse cornea. When we lowered the dose of FGF-2 in the cornea 6.4-fold (12.5 ng), the primary response was lymphangiogenic. Further investigation revealed that vascular endothelial growth factor-C and -D are required for this apparent lymphangiogenic property of FGF-2, and when the small amount of accompanying angiogenesis was completely suppressed, lymphangiogenesis remained unaffected. Our findings demonstrate that there is a dose-dependent response of FGF-2 for lymphangiogenesis, and lymphangiogenesis can occur in the absence of a preexisting or developing vascular bed, i.e., in the absence of angiogenesis, in the mouse cornea. PMID- 15289611 TI - Genes encoding candidate pheromone receptors in a moth (Heliothis virescens). AB - The remarkable responsiveness of male moths to female released pheromones is based on the extremely sensitive and selective reaction of highly specialized sensory cells in the male antennae. These cells are supposed to be equipped with male-specific receptors for pheromonal compounds, however, the nature of these receptors is still elusive. By using a combination of genomic sequence analysis and cDNA-library screening, we have cloned various cDNAs of the tobacco budworm Heliothis virescens encoding candidate olfactory receptors. A comparison of all identified receptor types not only highlighted their overall high degree of sequence diversity but also led to the identification of a small group of receptors sharing >40% identity. In RT-PCR analysis it was found that distinct members of this group were expressed exclusively in the antennae of male moths. In situ hybridization experiments revealed that the male-specific expression of these receptor types was confined to antennal cells located beneath sensillar hair structures (sensilla triochoidea), which have been shown to contain pheromone-sensitive neurons. Moreover, two-color double in situ-hybridization approaches uncovered that cells expressing one of these receptor types were surrounded by cells expressing pheromone-binding proteins, as expected for a pheromone-sensitive sensillum. These findings suggest that receptors like Heliothis receptor 14-16 (HR14-HR16) may render antennal cells responsive to pheromones. PMID- 15289612 TI - The structure of human parvovirus B19. AB - Human parvovirus B19 is the only parvovirus known to be a human pathogen. The structure of recombinant B19-like particles has been determined to approximately 3.5-A resolution by x-ray crystallography and, to our knowledge, represents the first near-atomic structure of an Erythrovirus. The polypeptide fold of the major capsid protein VP2 is a "jelly roll" with a beta-barrel motif similar to that found in many icosahedral viruses. The large loops connecting the strands of the beta-barrel form surface features that differentiate B19 from other parvoviruses. Although B19 VP2 has only 26% sequence identity to VP3 of adeno-associated virus, 72% of the C(alpha) atoms can be aligned structurally with a rms deviation of 1.8 A. Both viruses require an integrin as a coreceptor, and conserved surface features suggest a common receptor-binding region. PMID- 15289613 TI - Crystal structure of Caenorhabditis elegans HER-1 and characterization of the interaction between HER-1 and TRA-2A. AB - HER-1 is a secreted protein that promotes male development in the nematode Caenorhabditis elegans. HER-1 inhibits the function of TRA-2A, a multipass integral membrane protein thought to serve as its receptor. We report here the 1.5-A crystal structure of HER-1. The structure was solved by the multiwavelength anomalous diffraction method by using selenomethionyl-substituted HER-1 produced in Chinese hamster ovary cells. The HER-1 structure consists of two all-helical domains and is not closely homologous to any known structure. Sites of amino acid substitutions known to impair HER-1 function were mapped on the HER-1 structure and classified according to the likely mechanism by which they affect HER-1 activity. A subset of these and other amino acid substitutions on the HER-1 surface were assayed for their ability to disrupt interactions between HER-1 and TRA-2A-expressing cells, and a localized region on the HER-1 surface important for mediating this interaction was identified. PMID- 15289614 TI - Rhodopsin kinase activity modulates the amplitude of the visual response in Drosophila. AB - A feature shared between Drosophila rhodopsin and nearly all other G protein coupled receptors is agonist-dependent protein phosphorylation. Despite extensive analyses of Drosophila phototransduction, the identity and function of the rhodopsin kinase (RK) have been elusive. Here, we provide evidence that G protein coupled receptor kinase 1 (GPRK1), which is most similar to the beta-adrenergic receptor kinases, G protein-coupled receptor kinase 2 (GRK2) and GRK3, is the fly RK. We show that GPRK1 is enriched in photoreceptor cells, associates with the major Drosophila rhodopsin, Rh1, and phosphorylates the receptor. As is the case with mammalian GRK2 and GRK3, Drosophila GPRK1 includes a C-terminal pleckstrin homology domain, which binds to phosphoinositides and the Gbetagamma subunit. To address the role of GPRK1, we generated transgenic flies that expressed higher and lower levels of RK activity. Those flies with depressed levels of RK activity displayed a light response with a much larger amplitude than WT. Conversely, the amplitude of the light response was greatly suppressed in transgenic flies expressing abnormally high levels of RK activity. These data point to an evolutionarily conserved role for GPRK1 in modulating the amplitude of the visual response. PMID- 15289615 TI - High-throughput sensing and noninvasive imaging of protein nuclear transport by using reconstitution of split Renilla luciferase. AB - Nucleocytoplasmic trafficking of functional proteins plays a key role in regulating gene expressions in response to extracellular signals. We developed a genetically encoded bioluminescent indicator for monitoring the nuclear trafficking of target proteins in vitro and in vivo. The principle is based on reconstitution of split fragments of Renilla reniformis (Rluc) by protein splicing with a DnaE intein (a catalytic subunit of DNA polymerase III). A target cytosolic protein fused to the N-terminal half of Rluc is expressed in mammalian cells. If the protein translocates into the nucleus, the Rluc moiety meets the C terminal half of Rluc, and full-length Rluc is reconstituted by protein splicing. We demonstrated quantitative cell-based in vitro sensing of ligand-induced translocation of androgen receptor, which allowed high-throughput screening of exo- and endogenous agonists and antagonists. Furthermore, the indicator enabled noninvasive in vivo imaging of the androgen receptor translocation in the brains of living mice with a charge-coupled device imaging system. These rapid and quantitative analyses in vitro and in vivo provide a wide variety of applications for screening pharmacological or toxicological compounds and testing them in living animals. PMID- 15289616 TI - Recruitment of the ArgR/Mcm1p repressor is stimulated by the activator Gcn4p: a self-checking activation mechanism. AB - Transcription of the arginine biosynthetic gene ARG1 is repressed by the ArgR/Mcm1p complex in arginine-replete cells and activated by Gcn4p, a transcription factor induced by starvation for any amino acid. We show that all four subunits of the arginine repressor are recruited to ARG1 by Gcn4p in cells replete with arginine but starved for isoleucine/valine. None of these proteins is recruited to the Gcn4p target genes ARG4 and SNZ1, which are not regulated by ArgR/Mcm1p. Mcm1p and Arg80p were found in a soluble complex lacking Arg81p and Arg82p, and both Mcm1p and Arg80p were efficiently recruited to ARG1 in wild-type cells in the presence or absence of exogenous arginine, and also in arg81Delta cells. By contrast, the recruitment of Arg81p and Arg82p was stimulated by exogenous arginine. These findings suggest that Gcn4p constitutively recruits an Mcm1p/Arg80p heterodimer and that efficient assembly of a functional repressor also containing Arg81p and Arg82p occurs only in arginine excess. By recruiting an arginine-regulated repressor, Gcn4p can precisely modulate its activation function at ARG1 according to the availability of arginine. PMID- 15289617 TI - In vitro transcription system delineates the distinct roles of the coactivators pCAF and p300 during MyoD/E47-dependent transactivation. AB - The transcriptional coactivators p300 and pCAF are necessary for the myogenic factor MyoD to initiate the expression of skeletal muscle genes. In addition to mediating histone acetylation, both of these factors can acetylate MyoD; however, the complexity of cellular systems used to study MyoD has impeded delineation of the specific roles of these two acetyltransferases. Therefore, we established a MyoD-dependent in vitro transcription system that permits us to determine the roles of p300 and pCAF during MyoD-dependent transcriptional activation. Consistent with results from cellular systems, we demonstrate that maximal levels of transactivation in vitro require both p300 and pCAF, as well as the cofactor acetyl CoA. Dissection of the steps leading to transcription initiation revealed that the activities of p300 and pCAF are not redundant. During the initial stages of transactivation, p300 acetylates histone H3 and H4 within the promoter region and then recruits pCAF to MyoD. Once tethered to the promoter, pCAF acetylates MyoD to facilitate the transactivation process. Thus, we have established that pCAF and p300 carry out sequential and functionally distinct events on a promoter leading to transcriptional activation. Further dissection of this in vitro transcription system should be highly useful toward elucidating the mechanism by which coactivators facilitate differential gene expression by MyoD. PMID- 15289618 TI - Lightoid and Claret: a rab GTPase and its putative guanine nucleotide exchange factor in biogenesis of Drosophila eye pigment granules. AB - To elucidate the biogenetic pathways for the generation of lysosome-related organelles, we have chosen to study the Drosophila eye pigment granules because they are lysosome-related and the fruit fly provides the advantages of a genetic system in which many mutations affect eye color. Here, we report the molecular identification of two classic Drosophila eye-color genes required for pigment granule biogenesis, claret and lightoid; the former encodes a protein containing seven repeats with sequence similarity to those that characterize regulator of chromosome condensation 1 (RCC1, a guanine nucleotide exchange factor for the small GTPase, Ran), and the latter encodes a rab GTPase, Rab-RP1. We demonstrate in transfected cells that Claret, through its RCC1-like domain, interacts preferentially with the nucleotide-free form of Rab-RP1, and this interaction involves Claret's first three RCC1-like repeats that are also critical for Claret's function in pigment granule biogenesis in transgenic rescue experiments. In addition, double-mutant analyses suggest that the gene products of claret and lightoid function in the same pathway, which is different from that of garnet and ruby (which encode the delta- and beta-subunit of the tetrameric adaptor protein 3 complex, respectively). Taken together, our results suggest that Claret functions as a guanine nucleotide exchange factor for Lightoid/Rab-RP1 in an adaptor protein 3-independent vesicular trafficking pathway of pigment granule biogenesis. PMID- 15289620 TI - Analysis of clinical incidents: a window on the system not a search for root causes. AB - Incident reporting lies at the heart of many initiatives to improve patient safety. The UK National Patient Safety Agency (NPSA)1 has recently launched a national reporting and learning system following substantial piloting and testing across the National Health Service (NHS). In the USA the Agency for Healthcare Research and Quality (AHRQ) made incident reporting the centrepiece of its first patient safety funding programme, investing $25 million in the first year into research in incident reporting systems.2 The Australian incident monitoring system has amassed a massive database of reports over 15 years.3 New risk management and patient safety programmes-whether local or national-rely on incident reporting to provide data on the nature of safety problems and to provide indications of the causes of those problems and the likely solutions. PMID- 15289619 TI - Coactivator AIB1 links estrogen receptor transcriptional activity and stability. AB - Agonist-mediated degradation of estrogen receptor alpha (ERalpha) has been associated with its transcriptional activity. However, the mechanism by which ERalpha is targeted for degradation and whether there is a direct functional link between ERalpha stability and ERalpha-mediated transactivation have not been elucidated. Here we provide evidence that the p160 coactivator, AIB1, uniquely mediates agonist-induced, but not antagonist-induced, ERalpha degradation. We show that AIB1 recruitment by ERalpha is not only necessary but also sufficient to promote degradation. Suppression of AIB1 levels leads to ERalpha stabilization in the presence of 17beta-estradiol and, despite increased ERalpha levels, reduced recruitment of ERalpha to endogenous target gene promoters. In addition, association of RNA polymerase II with ERalpha target promoters is lost when AIB1 is suppressed, leading to inhibition of target gene transcription. AIB1 thus plays a dual role in regulating ERalpha activity, one in recruiting transcription factors including other coactivators involved in gene activation and the other in regulating ERalpha protein degradation mediated by the ubiquitin-proteosome machinery. PMID- 15289621 TI - Using statistical process control to improve the quality of health care. PMID- 15289622 TI - Waking up to chronic care. PMID- 15289623 TI - Developing and implementing organisational practice that delivers better, safer care. PMID- 15289624 TI - Developing quality indicators to assess quality of care. PMID- 15289625 TI - FMEA and RCA: the mantras of modern risk management. PMID- 15289626 TI - Use of medical emergency team responses to reduce hospital cardiopulmonary arrests. AB - BACKGROUND: Medical emergency team (MET) responses have been implemented to reduce inpatient mortality, but data on their efficacy are sparse and there have been no reports to date from US hospitals. OBJECTIVES: To determine how the incidence and outcomes of cardiac arrests have changed following increased use of MET. METHODS: Objective criteria for MET activation were created and disseminated as part of a crisis management program, after which there was a rapid and sustained increase in the use of MET. A retrospective analysis of clinical outcomes was performed to compare the incidence and mortality of cardiopulmonary arrest before and after the increased use of MET. RESULTS: A retrospective analysis of 3269 MET responses and 1220 cardiopulmonary arrests over 6.8 years showed an increase in MET responses from 13.7 to 25.8 per 1000 admissions (p<0.0001) after instituting objective activation criteria. There was a coincident 17% decrease in the incidence of cardiopulmonary arrests from 6.5 to 5.4 per 1000 admissions (p = 0.016). The proportion of fatal arrests was similar before and after the increase in use of MET. CONCLUSIONS: Increased use of MET may be associated with fewer cardiopulmonary arrests. PMID- 15289627 TI - Use of medical emergency team (MET) responses to detect medical errors. AB - BACKGROUND: No previous studies have investigated whether medical emergency team (MET) responses can be used to detect medical errors. OBJECTIVES: To determine whether review of MET responses can be used as a surveillance method for detecting medical errors. METHODS: Charts of all patients receiving MET responses during an 8 month period were reviewed by a hospital based Quality Improvement Committee to establish if the clinical deterioration that prompted the MET response was associated with a medical error (defined as an adverse event that was preventable with the current state of medical knowledge). Medical errors were categorized as diagnostic, treatment, or preventive errors using a descriptive typology based on previous published reports. RESULTS: Three hundred and sixty four consecutive MET responses underwent chart review and 114 (31.3%) were associated with medical errors: 77 (67.5%) were categorized as diagnostic errors, 68 (59.6%) as treatment errors, and 30 (26.3%) as prevention errors. Eighteen separate hospital care processes were identified and modified as a result of this review, 10 of which involved standardization. CONCLUSIONS: MET review may be used for surveillance to detect medical errors and to identify and modify processes of care that underlie those errors. PMID- 15289628 TI - Developing quality indicators for older adults: transfer from the USA to the UK is feasible. AB - BACKGROUND: Measurement of the quality of health care is essential for quality improvement, and patients are an underused source of data about quality of care. We describe the adaptation of a set of USA quality indicators for use in patient interview surveys in England, to measure the extent to which older patients receive a broad range of effective health care interventions in both primary and secondary care. METHOD: One hundred and nineteen quality indicators covering 16 clinical areas, based on a set of indicators for the care of vulnerable elderly patients in the USA, were reviewed by a panel of 10 clinical experts in England. A modified version of the RAND/UCLA appropriateness method was used and panel members were supplied with literature reviews summarising the evidence base for each quality indicator. The indicators were sent for comment before the panel meeting to UK charitable organisations for older people. RESULTS: The panel rated 102 of the 119 indicators (86%) as valid for use in England; 17 (14%) were rejected as invalid. All 58 indicators about treatment or continuity and follow up were rated as valid compared with just over half (13 of 24) of the indicators about screening. CONCLUSIONS: These 102 indicators are suitable for use in patient interview surveys, including the English Longitudinal Study of Ageing (ELSA). The systematic measurement of quality of care at the population level and identification of gaps in quality is essential for quality improvement. There is potential for transfer of quality indicators between countries, at least for the health care of older people. PMID- 15289629 TI - Design of a safer approach to intravenous drug infusions: failure mode effects analysis. AB - OBJECTIVES: A set of standard processes was developed for delivering continuous drug infusions in order to improve (1) patient safety; (2) efficiency in staff workflow; (3) hemodynamic stability during infusion changes, and (4) efficient use of resources. Failure modes effects analysis (FMEA) was used to examine the impact of process changes on the reliability of delivering drug infusions. SETTING: An 11 bed multidisciplinary pediatric ICU in the children's hospital of an academic medical center staffed by board certified pediatric intensivists. The hospital uses computerized physician order entry for all medication orders. METHODS: A multidisciplinary team characterized key elements of the drug infusion process. The process was enhanced to increase overall reliability and the original and revised processes were compared using FMEA. Resource consumption was estimated by reviewing purchasing and pharmacy records for the calendar year after full implementation of the revised process. Staff satisfaction was evaluated using an anonymous questionnaire administered to staff nurses in the ICU and pediatric residents who had rotated through the ICU. RESULTS: The original process was characterized by six elements: selecting the drug; selecting a dose; selecting an infusion rate; calculating and ordering the infusion; preparing the infusion; programming the infusion pump and delivering the infusion. The following practice changes were introduced: standardizing formulations for all infusions; developing database driven calculators; extending infusion hang times from 24 to 72 hours; changing from bedside preparation by nurses to pharmacy prepared or premanufactured solutions. FMEA showed that the last three elements of the original process had high risk priority numbers (RPNs) of >225 whereas the revised process had no elements with RPNs >100. The combined effect of prolonging infusion hang times, preparation in the pharmacy, and purchasing premanufactured solutions resulted in 1500 fewer infusions prepared by nurses per year. Nursing staff expressed a significant preference and pediatric residents unanimously expressed a strong preference for the revised process. CONCLUSIONS: Standardization of infusion delivery reduced the frequency for completing the most unreliable elements of the process and reduced the riskiness of the individual elements. Both contribute to a safer system. PMID- 15289630 TI - Sources of variability in uncertain medical decisions in the ICU: a process tracing study. AB - BACKGROUND: Consistency of medical decision making (equity) is an important component of quality of care. When patients with chronic obstructive pulmonary disease (COPD) present with an exacerbation needing respiratory support they may die if it is not provided. However, if the disease has reached its terminal stage, ventilation will prolong the process of dying. The ventilation outcome is uncertain and there is evidence of variability when this decision is made, the sources of which are not well understood. OBJECTIVES: To identify sources of variability and propose ways of tackling them in order to promote equity in this type of medical decision. METHODS: Six case histories were selected from hospital records of COPD patients. Fourteen senior doctors from seven hospitals in the West Midlands participated. A process tracing approach was used which consisted of (1) withholding case information until specifically requested by the doctors, (2) estimating survival during the decision making process, and (3) concurrent questioning regarding information interpretation and its impact on survival estimates and decisions. RESULTS: The observed decision variability was attributed to doctors attaching importance to different information, gathering different information, and interpreting information differently. There were significant differences between doctors in the amount of information requested. CONCLUSIONS: Differences in information gathering and interpretation by clinicians can result in different decisions being made about the same patient. This variation may exist for other uncertain medical decisions and may be tackled by providing clinicians with prognostic models in the form of usable decision aids. PMID- 15289631 TI - Electronic reporting to improve patient safety. AB - BACKGROUND: Limited data are available on the experiences of voluntary event reporting systems to improve patient safety. OBJECTIVE: Development and implementation of educational initiatives to facilitate the use of an electronic reporting system (ERS) in an academic medical center to measure the impact on knowledge of the ERS on reporting behavior and safety attitudes and to evaluate the accuracy of the information being reported. METHODS: A voluntary internal confidential electronic system for reporting safety events was implemented which involved patients and visitors. A multifaceted educational program was developed to promote safety awareness and use of the ERS system. The safety event detail reported for the calendar year 2002 was tracked and trended and central event analyses were performed for five high event clinical areas. A survey was administered to assess safety knowledge and attitudes of patient care personnel. RESULTS: 2843 safety events were entered into the ERS during 2002 with an increase during the course of the year (p = 0.055, linear trend) for all events. Nurses entered 73% of the events and physicians only 2%. 453 events (16%) were unsafe conditions or near misses and 623 (22%) were associated with patient harm. System factors were considered by the reporter as contributing to the event in only a few cases (5%). Central event analysis revealed that 39% of events had coding errors either in event classification, level of impact, or location; significant underreporting was also present. Although survey response rates were low (10.3%), responders showed a high degree of knowledge on general questions of patient safety and an increase in knowledge on use of the ERS (p = 0.0015, linear trend). CONCLUSIONS: Knowledge on the use of the reporting system and the frequency of reported events increased over the first year of the study. More work is needed to involve physicians in reporting, to improve the accuracy of submitted information, and to better prioritize, organize, and streamline event analysis. PMID- 15289632 TI - Assessing organisational development in primary medical care using a group based assessment: the Maturity Matrix. AB - OBJECTIVE: To design and develop an instrument to assess the degree of organisational development achieved in primary medical care organisations. DESIGN: An iterative development, feasibility and validation study of an organisational assessment instrument. SETTING: Primary medical care organisations. PARTICIPANTS: Primary care teams and external facilitators. MAIN OUTCOME MEASURES: Responses to an evaluation questionnaire, qualitative process feedback, hypothesis testing, and quantitative psychometric analysis (face and construct validity) of the results of a Maturity Matrix assessment in 55 primary medical care organisations. RESULTS: Evaluations by 390 participants revealed high face validity with respect to its usefulness as a review and planning tool at the practice level. Feedback from facilitators suggests that it helped practices to prioritise their organisational development. With respect to construct validity, there was some support for the hypothesis that training and non-training status affected the degree and pattern of organisational development. The size of the organisation did not have a significant impact on the degree of organisational development. CONCLUSION: This practice based facilitated group evaluation method was found to be both useful and enjoyable by the participating organisations. Psychometric validation revealed high face validity. Further developments are in place to ensure acceptability for summative work (benchmarking) and formative feedback processes (quality improvement). PMID- 15289633 TI - Sustaining better diabetes care in remote indigenous Australian communities. AB - PROBLEM: Inhabitants of Torres Strait Islands have the highest prevalence of diabetes in Australia and many preventable complications. In 1999, a one year randomised cluster trial showed improved diabetes care processes and reduced admissions to hospital when local indigenous health workers used registers, recall and reminder systems, and basic diabetes care plans, supported by a specialist outreach service. This study looked at whether those improvements were sustained two years after the end of the trial. DESIGN: Three year follow up clinical audit of 21 primary health care centres, and review of admissions to hospital in the previous 12 months. BACKGROUND AND SETTING: Remote indigenous communities in far north east Australia, population about 9600, including 921 people with diabetes. KEY MEASURES FOR IMPROVEMENT: Number of people on registers, care processes (regular measures of weight, blood pressure, haemoglobin A1c, urinary protein concentration, and concentrations of serum lipids and creatinine), appropriate clinical interventions (drug treatment and vaccinations), and intermediate patient outcome measures (weight, blood pressure, and glycaemic control). Admissions to hospital. STRATEGIES FOR CHANGE: Audit and feedback to clinicians and managers; provision of clinical guidelines and a clear management structure; workshops and training. EFFECTS OF CHANGE: The number of people on registers increased from 555 in 1999 to 921 in 2002. Most care processes and clinical interventions improved. The proportion of people with good glycaemic control (haemoglobin A1c 7%) increased from 18% to 25% in line with increased use of insulin (from 7% to 16%). The proportion of those with well controlled hypertension (< 140/90) increased from 40% to 64%. The proportion admitted to hospital with a diabetes related condition fell from 25% to 20%. Mean weight increased from 87 kg to 91 kg. LESSONS LEARNT: In remote settings, appropriate management structures and clinical support for people with diabetes can lead to improvements in care processes, control of blood pressure, and preventable complications that result in admission to hospital. Control of weight and glycaemia are more difficult and requires more active community engagement. Priorities now include increasing the availability and affordability of good food, achieving weight loss, and increasing appropriate use of hypoglycaemic agents, including insulin. PMID- 15289634 TI - Improving the quality of health care for chronic conditions. AB - Chronic conditions are increasingly the primary concern of health care systems throughout the world. In response to this challenge, the World Health Organization has joined with the MacColl Institute for Healthcare Innovation to adapt the Chronic Care Model (CCM) from a global perspective. The resultant effort is the Innovative Care for Chronic Conditions (ICCC) framework which expands community and policy aspects of improving health care for chronic conditions and includes components at the micro (patient and family), meso (health care organisation and community), and macro (policy) levels. The framework provides a flexible but comprehensive base on which to build or redesign health systems in accordance with local resources and demands. PMID- 15289636 TI - Shuffle the deck, flip that coin: randomization comes to medicine. PMID- 15289635 TI - Adverse drug events and medication errors: detection and classification methods. AB - Investigating the incidence, type, and preventability of adverse drug events (ADEs) and medication errors is crucial to improving the quality of health care delivery. ADEs, potential ADEs, and medication errors can be collected by extraction from practice data, solicitation of incidents from health professionals, and patient surveys. Practice data include charts, laboratory, prescription data, and administrative databases, and can be reviewed manually or screened by computer systems to identify signals. Research nurses, pharmacists, or research assistants review these signals, and those that are likely to represent an ADE or medication error are presented to reviewers who independently categorize them into ADEs, potential ADEs, medication errors, or exclusions. These incidents are also classified according to preventability, ameliorability, disability, severity, stage, and responsible person. These classifications, as well as the initial selection of incidents, have been evaluated for agreement between reviewers and the level of agreement found ranged from satisfactory to excellent (kappa = 0.32-0.98). The method of ADE and medication error detection and classification described is feasible and has good reliability. It can be used in various clinical settings to measure and improve medication safety. PMID- 15289638 TI - NIH-Supported National Islet Transplantation Registry. AB - An estimated 300,000 to 500,000 cases of type 1 diabetes exist today in the United States. Despite strict monitoring and attempts at control, people with type 1 diabetes still face the prospect of diminished health and earlier death than the general population. Islet transplantation offers an alternative to insulin usage and a potential treatment for type 1 diabetes mellitus. There are more than 30 islet transplant centers in the world focusing their efforts on the challenges and methods of this procedure. As the field of islet transplantation matures and the number of islet transplants performed increases, detailed analyses on factors that predict patient and graft survival are needed. This increased amount of data will allow for a better understanding of the safety and efficacy of islet transplantation. In response to the need for more complete information in the field, the National Institute of Diabetes and Digestive and Kidney Diseases is sponsoring the North American Collaborative Islet Transplant Registry (CITR). The mission of CITR is to expedite progress and promote safety in islet/beta-cell transplantation through the collection, analysis, and communication of comprehensive and current data on all islet/beta-cell transplants performed in North America. Compiling and analyzing data from all transplant centers in North America will accelerate the identification of both critical risk factors and key determinants of success, and thereby guide transplant centers in developing and refining islet/beta-cell transplant protocols, leading to an advancement in the field of islet transplantation. Participation in CITR is voluntary, and more than 22 transplant centers have been invited to join. Seven centers are actively participating in CITR, with an additional 11 centers in the process of joining. Both an executive committee and a scientific advisory committee guide CITR. All islet transplants performed in North America since January 1, 1996, are captured by the CITR database. Through an electronic, Internet-based data capture system, quality control procedures, and minimization of duplicate efforts at the transplant center, the most relevant and succinct information are entered. From these data a comprehensive report will be published annually. In addition, special analyses will be performed and published periodically. PMID- 15289639 TI - FDA regulation of allogeneic islets as a biological product. AB - This article describes the Food and Drug Administration's recent manufacturing review experience with investigational new drug applications submitted for allogeneic pancreatic islets of Langerhans for the treatment of type 1 diabetes mellitus. In addition, considerations of islet preparation issues that will need to be resolved before the submission of a biologics license application are discussed. PMID- 15289640 TI - Request for applications: islet cell replacement in type 1 diabetes. AB - The American Diabetes Association announces funding for clinical and basic science studies in three major areas of research that focus on cell-based insulin replacement in type 1 diabetes: 1. Genetic engineering of nonpancreatic cells into glucose-sensitive insulin-producing cells; 2. Transforming stem cells, pancreatic ductal cells, or pancreatic endocrine cell lines into glucose sensitive insulin-producing cells; and 3. Xenografts of nonhuman islet cells. PMID- 15289641 TI - New strategies in immune tolerance induction. AB - Induction of tolerance in clinical organ transplantation that will obviate the use of chronic immunosuppression and preserve host immune response to other antigens remains the goal of transplant research. The thymus plays a critical role in the ability of the immune system to discriminate between self- and nonself-antigens or harmful and harmless alloantigens. We now know that multiple factors determine how the immune system responds to a self-antigen or foreign antigen. These determinants include developmental stage of the host, stage of T cell maturity, site of antigen encounter, type and maturity of antigen-presenting cells, and presence and type of costimulatory molecules. Our understanding of the mechanisms of T-cell interactions with peptide/ major histocompatibility complex in peripheral lymphoid organs has led to experiments that translate into peripheral T-cell tolerance. The induction of high-avidity peripheral alloreactive T cells in the early phase of organ transplantation makes it difficult to achieve long-term alloantigen-specific tolerance without the use of transient perioperative immunosuppression. Therefore, protocols that induce robust tolerance in rodent and nonhuman primate models involve the use of donor antigen combined with a short course of perioperative immunosuppression. These studies suggest that the underlying mechanisms of peripheral tolerance include deletion, anergy, immune deviation, and regulatory T cells. This review focuses on recent advances in tolerance induction in experimental animal models and discusses their relevance to the development of protocols for the induction and maintenance of clinical transplant tolerance. PMID- 15289642 TI - Advances in long-term islet culture: the Memphis experience. AB - Long-term culture of human islets provides opportunity for improving results of islet transplantation. The techniques of long-term culture are reproducible and can result in improved function of the islet after transplantation into NOD-SCID mice. We have been able to cure streptozotocin-induced diabetes by islets cultured for more than 6 mo. Culture conditions play an important role in the success of the procedure. Culture success is dependent on the media type, additives, type of colloid or protein used, purity of the islets, and concentration and volume of the tissue. Cellular and structural changes occur over time in culture. These changes may explain the improved efficacy of the islet graft after short and intermediate culture periods. Further research into long-term culture of islets is necessary to fully explore the potential of the technique. PMID- 15289643 TI - Promoting islet cell function after transplantation. AB - Engraftment (i.e., the adaptation of transplanted pancreatic islets to their new surroundings with regard to revascularization, reinnervation, and reorganization of other stromal compartments) is of crucial importance for the survival and function of the endocrine cells. Previous studies suggest that transplantation induces both vascular and stromal dysfunctions in the implanted islets when compared with endogenous islets. Thus the vascular density and the blood perfusion of islet grafts is decreased and accompanied with a capillary hypertension. This leads to hypoxic conditions, with an associated shift toward anaerobic metabolism in grafted islets. An improved engraftment will prevent or compensate for the vascular/stromal dysfunction seen in transplanted islets and thereby augment survival of the islet implant. By such means the number of islets needed to cure the recipient will be lessened. This will increase the number of patients that can be transplanted with the limited material available. PMID- 15289644 TI - The role of GLP-1 in the regulation of islet cell mass. AB - Glucagon-like peptide-1 (GLP-1) is an incretin hormone capable of restoring euglycemia in glucose-intolerant subjects and improving glucose control in individuals with type 2 diabetes. Whether the antidiabetic properties of GLP-1 are exclusively the result of its acute postprandial action is being investigated. A GLP-1-dependent differentiation of pancreatic precursor cells into mature beta-cells has been proposed. In addition, GLP-1 has been shown to have antiapoptotic activity in cultured insulin-secreting cells and in an animal model in which diabetes occurs as a consequence of an excessive rate of beta-cell apoptosis. Studies from our laboratory, and others, lead us to propose that GLP-1 is a growth factor for pancreatic cells and it is a regulator of islet cell mass. The aim of this article is to review those reports that have emphasized the role of GLP-1 as a regulator of islet cell mass as well as its insulin secretory action. PMID- 15289645 TI - Islet morphogenesis and stem cell markers. AB - The mechanism of islet neogenesis remains poorly understood, despite its potential applications in regenerative or replacement therapies for the treatment of insulin-dependent diabetes. During fetal development of the mouse or rat, the majority of islet cells are formed in late gestation (E18-21) by the process of neogenesis from precursor cells. The precursor cells are organized as ducts that actively proliferate and express high levels of specific cytokeratin (CK) proteins. Transitional cells coexpressing islet hormones and CK are frequent and disappear shortly after birth, to reappear only in conditions in which pancreas or islet regeneration has been induced. Islet morphogenesis is thought to operate mainly through the budding of islet cells from ducts, followed by their migration away from the duct to form clusters. Single islet cells are indeed frequent in the fetal and regenerating pancreas, but they also occur in normal tissue, especially in the human pancreas. A different neogenic mechanism, observed in the fetal rat, consists in the proliferation of ductal cells resulting in large aggregates. Starting from the middle of the aggregate, cells differentiate into islet cells and gradually lose their proliferative activity and other ductal characteristics. In adult pancreas, islets are in close contact with at least one duct or ductule. Such a direct duct-islet axis becomes even more evident in regeneration models, such as duct ligation. In these models, a metaplastic transformation of the exocrine pancreas to so-called pseudoductal complexes is seen. Surviving exocrine cells acquire a metaplastic phenotype, which resembles the fetal protodifferentiated state. They start to express CK, the beta-cell transcription factor Pdx1, the neuroendocrine/islet cell markers PGP9.5 and the CCKB receptor for gastrin, and they show pronounced proliferative activity and islet neogenesis. We hypothesize that these de-differentiated or metaplastic exocrine cells (acinar and ductal), acquire a multipotential state and can serve as islet precursors. PMID- 15289646 TI - Islet-derived multipotential cells/progenitor cells. AB - The pancreatic beta-cell has a pivotal role in the regulation of glucose homeostasis; its death leads to type I diabetes. Neogenesis of beta-cells, the differentiation of beta-cells from non-beta-cells, could be an important mechanism of islet cell repopulation. To examine the ability of the adult pancreas to generate new beta-cells, we characterized the phenotype of beta precursor cells in embryos and then determined that cells expressing embryonic traits appeared in islets of adult mouse pancreas following deletion of preexisting insulin cells by streptozotocin, a specific beta-cell toxin. These precursor cells generated new beta-cells (NBCs) that repopulated the islets. The number of NBCs increased dramatically after restoration of normoglycemia by insulin therapy. Future studies will seek to identify the source of the NBCs and to examine the mechanisms that lead to their differentiation. PMID- 15289647 TI - Sources of beta-cells for human cell-based therapies for diabetes. AB - Recent progress in islet transplantation coupled with the extremely limited supply of primary human islets has spurred the search for alternative sources of beta-cells for transplantation therapy in treating diabetes. Many potential sources of cells are being explored, including embryonic and adult stem cells, identification of intrapancreatic precursor cells, and human beta-cell lines. Here, we review the promise and problems with those cell sources, focusing on our studies in developing functional human beta-cell lines. Those efforts involve a two-step process in which the first is to introduce growth stimulatory genes that induce human beta-cells to enter the cell cycle. Immortalization can then be achieved by expressing the hTERT telomerase subunit. The second step is to induce differentiation. This involves a complex set of manipulations, including the expression of the important beta-cell transcription factor PDX-1. Although PDX-1 is critical for promoting beta-cell differentiation, we do not find increased expression of the glucagon-like peptide-1 receptor, a gene that has been reported to be induced by PDX-1. Further understanding of the factors governing beta-cell development are likely to be required before a robust cell-based therapy is available for the treatment of diabetes. PMID- 15289648 TI - Generation of new islets from stem cells. AB - Spain ranks number one in organ donors (35 per million per yr). Although the prevalence of diabetes is low (100,000 type 1 diabetic patients and 2 million type 2 diabetic patients), the expected number of patients receiving islet transplants should be estimated at 200 per year. Islet replacement represents a promising cure for diabetes and has been successfully applied in a limited number of type 1 diabetic patients, resulting in insulin independence for periods longer than 3 yr. However, it has been difficult to obtain sufficient numbers of islets from cadaveric donors. Interesting alternatives include acquiring renewable sources of cells using either embryonic or adult stem cells to overcome the islet scarcity problem. Stem cells are capable of extensive proliferation rates and are capable of differentiating into other cell types of the body. In particular, totipotent stem cells are capable of differentiating into all cell types in the body, whereas pluripotent stem cells are limited to the development of a certain number of differentiated cell types. Insulin-producing cells have been obtained from both embryonic and adult stem cells using several approaches. In animal models of diabetes, the therapeutic application of bioengineered insulin secreting cells derived from stem cells has delivered promising results. This review will summarize the different approaches that have been used to obtain insulin-producing cells from embryonic and adult stem cells and highlights the key points that will allow in vitro differentiation and subsequent transplantation in the future. PMID- 15289649 TI - Developmental biology of the pancreas. AB - In this review, I summarize some aspects of murine pancreas development, with particular emphasis on the analysis of the ontogenetic relationships between different pancreatic cell types. Lineage analyses allow the identification of the progenitor cells from which mature cell types arise. The identification and successful in vitro culture of putative pancreatic stem cells is highly relevant for future cell replacement therapies in diabetic patients. PMID- 15289650 TI - (Pro)Insulin processing: a historical perspective. AB - Insulin, the major secreted product of the beta-cells of the islets of Langerhans, is initially synthesized as a precursor (preproinsulin), from which the mature hormone is excised by a series of proteolytic cleavages. This review provides a personal narrative of some of the key research projects leading to the identification of the central processing enzymes as proprotein convertase 1, proprotein convertase 2, and carboxypeptidase E. It also discusses the central roles of the intragranular environment and chaperone-like proteins in modulating processing activity. PMID- 15289651 TI - Understanding of basic mechanisms of beta-cell function and survival: prelude to new diabetes therapies. AB - Type 1 and type 2 diabetes are both diseases of insulin insufficiency, although they develop by distinct pathways. The recent surge in the incidence of type 2 diabetes and the chronic ailments confronted by patients with either form of the disease highlight the need for better understanding of beta-cell biology. In this review, we present recent work focused on this goal. Our hope is that basic research being conducted in this and other laboratories will ultimately contribute to the development of methods for enhancing beta-cell function and survival in the context of both major forms of diabetes. Our strategy for understanding the beta-cell involves a multidisciplinary approach in which tools from the traditional fields of biochemistry, enzymology, and physiology are teamed with newer technologies from the fields of molecular biology, gene discovery, cell and developmental biology, and biophysical chemistry. We have focused on two important aspects of beta-cell biology in our studies: beta-cell function, specifically the metabolic regulatory mechanisms involved in glucose stimulated insulin secretion, and beta-cell resistance to immune attack, with emphasis on resistance to inflammatory cytokines and reactive oxygen species. PMID- 15289652 TI - Secretory protein trafficking: genetic and biochemical analysis. AB - Acute insulin secretion from stimulated pancreatic beta-cells is derived from the intracellular pool of insulin secretory granules wherein insulin is packaged in a highly concentrated (and in some species, crystalline) state. Here we review experimental work, principally from our laboratory, on the question of biogenesis of mature secretory granules within the broader context of intracellular protein trafficking. Events occurring in the lumen of organelles at various stages of intracellular transport within the secretory pathway and events at the limiting membrane of newly forming secretory granules each contribute to formation of the insulin storage compartment comprising the readily releasable pool. PMID- 15289653 TI - Imaging glucose-regulated insulin secretion and gene expression in single islet beta-cells: control by AMP-activated protein kinase. AB - The mechanisms by which changes in glucose concentration regulate gene expression and insulin secretion in pancreatic islet beta-cells are only partly understood. Here we describe the development of new technologies for examining these processes at the level of single living beta-cells. We also present recent findings, made using these and other techniques, which implicate a role for adenosine 5'-monophosphate-activated protein kinase in glucose signaling in these cells. PMID- 15289655 TI - The role of A-Kinase anchoring proteins in cAMP-mediated signal transduction pathways. AB - Compartmentalization of signal transduction enzymes is an important mechanism of cellular signaling specificity. This occurs through the interaction of enzymes with scaffolding or anchoring proteins. To date, one of the best-studied examples of kinase anchoring is the targeting of protein kinase A to cellular locations through its association with A-kinase anchoring proteins (AKAPs). AKAPs mediate a high-affinity interaction with the type II regulatory subunit of protein kinase A for the purpose of localizing the kinase to pools of cyclic adenosine monophosphate and within proximity of preferred substrates. Furthermore, AKAPs can organize entire signaling complexes made up of kinases, phosphatases, signaling enzymes, and additional regulatory proteins. PMID- 15289654 TI - MAP kinases and their roles in pancreatic beta-cells. AB - We discuss our work examining regulation and functions of mitogen-activated protein kinases, particularly ERK1 and ERK2, in pancreatic beta-cells. These enzymes are activated by glucose, other nutrients, and insulinogenic hormones. Their activation by these agents is calcium-dependent. A number of other stimuli also activate ERK1/2, but by mechanisms distinct from those involved in nutrient sensing. Inhibition of ERK1/2 has no apparent effect on insulin secretion measured after 2 h. On the other hand, ERK1/2 activity is required for maximal glucose-dependent activation of the insulin gene promoter. The primary effort has focused on INS-1 cell lines, with supporting and confirmatory studies in intact islets and other beta-cell lines, indicating the generality of our findings in beta-cell function. Thus ERK1/2 participate in transmitting glucose-sensing information to beta-cell functions. These kinases most likely act directly and indirectly on multiple pathways that regulate beta-cell function and, in particular, to transduce an elevated glucose signal into insulin gene transcription. PMID- 15289656 TI - Protein tyrosine phosphatases: potential role in beta-cell insulin signal transduction. AB - Insulin receptor signal transduction plays a critical role in regulating pancreatic beta-cell function, notably the acute first-phase insulin release in response to glucose. The basis for insulin resistance in pancreatic beta-cells is not well understood but may be related to abnormal regulation of tyrosine phosphorylation events, which, in turn, may alter organization of insulin signaling molecules in space and time. Members of the protein tyrosine phosphatase (PTPase) family are both functionally and structurally diverse; and within the past few years data have emerged from many laboratories that suggest selectivity of the PTPase catalytic domains toward cellular substrates. Of significance, a subset of PTPases has been implicated in the regulation of insulin signaling in a number of insulin-sensitive tissues. Alteration in PTPase expression or activity has been associated with abnormal regulation of tyrosine phosphorylation events and is accompanied by modulation of insulin sensitivity in vivo. Manipulations aimed at reducing expression of physiologically relevant PTPases acting at a step proximal to the insulin receptor are accompanied by normalization of blood glucose levels and improved insulin sensitivity in both normal and diabetic animals. Hence, the development of tissue-specific gene inactivation strategies should facilitate the study of the potential role of PTPases in beta-cell insulin signaling transduction. PMID- 15289659 TI - Centromeres, kinetochores and the segregation of chromosomes. Foreword. PMID- 15289661 TI - The roles of histone modifications and small RNA in centromere function. AB - Here, epigenetic regulation of centromeric chromatin in fission yeast (Schizosaccharomyces pombe) is reviewed, focussing on the role of histone modifications and the link to RNA interference (RNAi). Fission yeast centromeres are organized into two structurally and functionally distinct domains, both of which are required for centromere function. The central core domain anchors the kinetochore structure while the flanking heterochromatin domain is important for sister centromere cohesion. The chromatin structure of both domains is regulated epigenetically. In the central core domain, the histone H3 variant Cnp1(CENP-A) plays a key role. In the flanking heterochromatin domain, histones are kept underacetylated by the histone deacetylases (HDACs) Clr3, Clr6 and Sir2, and methylated by Clr4 methyltransferase (HMTase) to create a specific binding site for the Swi6 protein. Swi6 then directly mediates cohesin binding to the centromeric heterochromatin. Recently, a surprising link was made between heterochromatin formation and RNAi. PMID- 15289660 TI - Kinetochore and heterochromatin domains of the fission yeast centromere. AB - Fission yeast centromeres are composed of two distinctive chromatin domains. The central domain nucleosomes contain the histone H3-like protein CENP-A(Cnp1). In contrast, the flanking repeats are coated with silent chromatin in which Swi6 (HP1) binds histone H3 methylated on lysine 9 that is induced by the action of the RNA interference pathway on non-coding centromeric transcripts. The overall structure is similar to that of metazoan centromeres where the kinetochore is embedded in surrounding heterochromatin. Kinetochore specific proteins associate with the central domain and affect silencing in that region. The flanking heterochromatin is required to recruit cohesin and mediate tight physical cohesion between sister centromeres. The loss of silencing that accompanies defects in heterochromatin has been invaluable as a tool in the investigation of centromere function. Both the heterochromatin and kinetochore regions are required for the de novo assembly of a functional centromere on DNA constructs, suggesting that heterochromatin may provide an environment that promotes kinetochore assembly within the central domain. The process is clearly epigenetically regulated. Fission yeast kinetochores associate with 2-4 microtubules, and flanking heterochromatin may be required to promote the orientation of multiple microtubule binding sites on one kinetochore towards the same pole and thus prevent merotelic orientation. PMID- 15289662 TI - The role of CENP-B and alpha-satellite DNA: de novo assembly and epigenetic maintenance of human centromeres. AB - The centromere is an essential functional domain responsible for the correct inheritance of eukaryotic chromosomes during cell division. Eukaryotic centromeres include the highly conserved centromere-specific histone H3 variant, CENP-A, which has provided a powerful tool for investigating the recruitment of centromere components. However, the trigger that targets CENP-A to a specific genomic locus during centromere assembly remains unknown. Although, on rare occasions, CENP-A chromatin may assemble at non-centromeric DNA, all normal human centromeres are assembled and maintained on alpha-satellite (alphoid) DNA. The importance of alphoid DNA and CENP-B binding sites (CENP-B boxes), typical of normal human centromere DNA configurations, has been demonstrated through their requirement in de novo centromere assembly and Human Artificial Chromosome (HAC) assays. Mechanisms to link the centromere tightly to specific genomic sequences exist in humans and the two yeast species. PMID- 15289663 TI - Centromere DNA, proteins and kinetochore assembly in vertebrate cells. AB - The centromere is a specialized region of the chromosome that is essential for faithful chromosome segregation during mitosis and meiosis in eukaryotic cells. It is the site at which the kinetochore, the functional nucleoprotein complex responsible for microtubule binding and chromosome movement, is assembled through complex molecular mechanisms. Herein, I review recent advances in our understanding of centromeric DNAs as sites for kinetochore assembly and the mechanisms underlying kinetochore assembly in vertebrate cells. PMID- 15289664 TI - Topoisomerase II: untangling its contribution at the centromere. AB - Topoisomerase II (topo II) is a major component of mitotic chromosomes and its unique decatenating activity has been implicated in many aspects of chromosome dynamics including DNA replication, transcription, recombination, chromosome condensation and segregation. Of these, chromosome segregation is the most seriously affected by loss of topo II expression or activity in living cells, most likely because of residual catenations between sister chromatids. At metaphase, vertebrate chromatids are attached to each other principally through their centromeric regions, and we review here evidence that topo II has a specific role at the centromere. Despite strong evidence for the centromere specific accumulation of topo II protein and the cytogenetic and molecular mapping of topo II catalytic activity to active centromeres, there is so far relatively little evidence for an overt role in centromere function (as judged by the effects of topo II inactivation on kinetochore assembly, bipolar microtubule attachment and chromosome separation). Nevertheless, recent data linking the post translational modification of topo II to the regulation of sister centromere cohesion suggest that topo II may indeed contribute to the timely separation of centromeres at anaphase. PMID- 15289665 TI - Kinetochore-microtubule interactions during cell division. AB - Proper segregation of chromosomes during cell division is essential for the maintenance of genetic stability. During this process chromosomes must establish stable functional interactions with microtubules through the kinetochore, a specialized protein structure located on the surface of the centromeric heterochromatin. Stable attachment of kinetochores to a number of microtubules results in the formation of a kinetochore fibre that mediates chromosome movement. How the kinetochore fibre is formed and how chromosome motion is produced and regulated remain major questions in cell biology. Here we look at some of the history of research devoted to the study of kinetochore-microtubule interaction and attempt to identify significant advances in the knowledge of the basic processes. Ultrastructural work has provided substantial insights into the structure of the kinetochore and associated microtubules during different stages of mitosis. Also, recent in-vivo studies have probed deep into the dynamics of kinetochore-attached microtubules suggesting possible models for the way in which kinetochores harness the capacity of microtubules to do work and turn it into chromosome motion. Much of the research in recent years suggests that indeed multiple mechanisms are involved in both formation of the k-fibre and chromosome motion. Thus, rather than moving to a unified theory, it has become apparent that most cell types have the capacity to build the spindle using multiple and probably redundant mechanisms. PMID- 15289666 TI - The spindle checkpoint: a quality control mechanism which ensures accurate chromosome segregation. AB - The centromere defines where on a chromosome the kinetochores assemble. Kinetochores, large protein structures, mediate chromosome segregation during mitosis and meiosis by performing three key functions. Firstly, kinetochores attach chromosomes to the microtubule spindle apparatus. Secondly, kinetochores co-ordinate microtubule dynamics to allow chromosomes to move along the spindle. Lastly, kinetochores generate the 'wait' signal which prevents anaphase onset until all the chromosomes are correctly aligned on the spindle. This signal forms part of the spindle checkpoint mechanism, a highly conserved cell cycle checkpoint which maintains the accuracy of the chromosome segregation process. This article provides a brief historical overview before focusing on some of the outstanding issues and more recent developments in the field. PMID- 15289667 TI - Chromosomal dynamics of human neocentromere formation. AB - Neocentromeres are rare human chromosomal aberrations where a new centromere has formed in a previously non-centromeric location. The emergence of new centromeres on a chromosome that already contains an endogenous centromere would be a highly deleterious event which would lead to dicentricity and mitotic instability. Nonetheless, neocentromere formation appears to provide a mechanism for the acquisition of a new centromere. Neocentromeres are most often observed on chromosomal arm fragments that have separated from an endogenous centromere, and therefore actually lead to mitotic stability of what would have been an acentric fragment. Neocentromeres have recently also been observed on apparently unrearranged chromosomes where the endogenous centromere has been inactivated. Furthermore, the process of centromere repositioning during primate chromosomal evolution may depend on the acquisition and subsequent fixation of neocentromeres. This remarkable plasticity in the position of centromeres has important implications for human cytogenetics and chromosome evolution, and provides an opportunity to further our understanding of the process of centromere formation and structure. PMID- 15289668 TI - Centric fission--simple and complex mechanisms. AB - Centric fission describes a rather poorly molecularly defined process of the transverse division of a functional centromere to result in two new centric chromosomes. While centric fission occurs rarely in humans, this process has been attributed an important role in eukaryotic karyotype evolution. Recent studies have given insight into the complex molecular mechanisms that lead to apparent centric fission events, including evidence in support of a mechanism driven by centric preduplication. These studies suggest that the traditional definition of centric fission, based on gross cytogenetic and molecular cytogenetic observations, needs to be broadened. It is necessary to distinguish between simple centric fissions, that involve the direct transverse breakage of a functional centromere, and other more complex fission events at or around a centromere that may be preceded by chromosomal rearrangements. PMID- 15289669 TI - "Holo"er than thou: chromosome segregation and kinetochore function in C. elegans. AB - Kinetochores are proteinaceous organelles that assemble on centromeric DNA to direct chromosome segregation in all eukaryotes. While many aspects of kinetochore function are conserved, the nature of the chromosomal domain upon which kinetochores assemble varies dramatically between different species. In monocentric eukaryotes, kinetochores assemble on a localized region of each chromosome. In contrast, holocentric species such as the nematode Caenorhabditis elegans have diffuse kinetochores that form along the entire length of their chromosomes. Here, we discuss the nature of chromosome segregation in C. elegans. In addition to reviewing what is known about kinetochore function, chromosome structure, and chromosome movement, we consider the consequences of the specialized holocentric architecture on chromosome segregation. PMID- 15289671 TI - Degradation of microbial polyesters. AB - Microbial polyhydroxyalkanoates (PHAs), one of the largest groups of thermoplastic polyesters are receiving much attention as biodegradable substitutes for non-degradable plastics. Poly(D-3-hydroxybutyrate) (PHB) is the most ubiquitous and most intensively studied PHA. Microorganisms degrading these polyesters are widely distributed in various environments. Although various PHB degrading microorganisms and PHB depolymerases have been studied and characterized, there are still many groups of microorganisms and enzymes with varying properties awaiting various applications. Distributions of PHB-degrading microorganisms, factors affecting the biodegradability of PHB, and microbial and enzymatic degradation of PHB are discussed in this review. We also propose an application of a new isolated, thermophilic PHB-degrading microorganism, Streptomyces strain MG, for producing pure monomers of PHA and useful chemicals, including D-3-hydroxycarboxylic acids such as D-3-hydroxybutyric acid, by enzymatic degradation of PHB. PMID- 15289670 TI - Plant neocentromeres: fast, focused, and driven. AB - Plant neocentromeres are large heterochromatic domains that associate with microtubules and move rapidly poleward during meiotic cell division. In maize, neocentromeres are part of a process that leads to the preferential recovery (meiotic drive) of knobs in progeny. These 'classical' plant neocentromeres differ from animal neocentromeres by their morphology, inability to mediate sister chromatid cohesion, and their rates of movement on the spindle. We provide a comprehensive review of classical neocentromeres with emphasis on their origin and mechanisms of motility. The data support the view that most, if not all, classical neocentromeres are the outcome of selection by meiotic drive. In addition, we compare and contrast neocentromere-mediated meiotic drive with a recently proposed meiotic drive model for centromere evolution. PMID- 15289672 TI - Correlation between the physicochemical properties of organic solvents and their biocompatibility toward epoxide hydrolase activity in whole-cells of a yeast, Rhodotorula sp. AB - Epoxides are often highly hydrophobic substrates and the presence of an organic co-solvent within an aqueous bioreactor is in such cases indicated. The effect of 40 water-miscible and -immiscible organic solvents on epoxide hydrolase activity in whole-cells of the yeast Rhodotorula sp. UOFS Y-0448 was investigated. No formal correlation between solvent biocompatibility and physicochemical properties was deductible, although the introduction of hydroxyl groups increased biocompatibility. 1-Pentanol, 2-methylcyclohexanol and 1-octanol were the most biocompatible resulting in relatively low activity losses when used at up to 20% (v/v). PMID- 15289673 TI - Hydrolytic kinetic resolution of the enantiomers of the structural isomers trans 1-phenylpropene oxide and (2,3-epoxypropyl)benzene by yeast epoxide hydrolase. AB - Kinetic resolution of the enantiomers of trans -1-phenylpropene oxide and (2,3 epoxypropyl)benzene was achieved by yeasts from the genus Rhodotorula. The resolution of trans -1-phenylpropene oxide by Rhodotorula glutinis UOFS Y-0123 yielded (1R,2R)-epoxide (ee >98%, yield 30%) and (1R,2S)-diol (ee 95%, yield 40%). The highest enantio- and regioselectivity toward (2,3-epoxypropyl)benzene resided in Rhodotorula sp. UOFS Y-0448 (E = 6.16), yielding (S)-epoxide (ee 64%, yield 33%) and (R)-diol (ee 67%, yield 28%). This confirms the superiority of yeasts from the Basidiomycetes genera in the enantioselective hydrolysis of epoxides from different structural classes. PMID- 15289674 TI - Detection method of the adjacent motor neuronal death in an in vitro co-culture model of familial ALS-associated Cu/Zn superoxide dismutase. AB - Mutations in Cu/Zn-superoxide dismutase (SOD1) gene have been identified in familial amyotrophic lateral sclerosis. Motor neuron degeneration subsequently spreads to contiguous neurons of the motor systems. We developed an in vitro disease model with motor neuron-neuroblastoma hybrid cells (VSC4.1) constitutively expressing a mutant (G93A) SOD1. The extracellular effect upon adjacent motor neurons was determined using the substratum culture insert. The viability of VSC 4.1 was lowered by 26% in a co-culture of VSC 4.1 and G93A, which was reversed by Trolox, an antioxidant. This in vitro disease model confirmed the extracellular toxicity of the mutant SOD1 cells on the adjacent neurons by generating oxidative stress. PMID- 15289675 TI - Improved method of analysis of biomass sugars using high-performance liquid chromatography. AB - The precise quantitative analysis of biomass derived sugars is a very important step in the conversion of biomass feedstocks to fuels and chemicals. However, the most accurate method of biomass sugar analysis is based on the gas chromatography analysis of derivatized sugars either as alditol acetates or trimethylsilanes. The derivatization method is time-consuming but the alternative HPLC method cannot resolve most sugars found in biomass hydrolysates. We have demonstrated for the first time that by careful manipulation of the HPLC mobile phase, biomass monomeric sugars (arabinose, xylose, fructose, glucose, mannose, and galactose) can be analyzed quantitatively and there is excellent baseline resolution of all the sugars. This was demonstrated for both standard sugars and corn stover hydrolysates. Our method can also be used to analyze dimmeric sugars (cellobiose and sucrose). PMID- 15289676 TI - Chemo-enzymatic synthesis of N-arachidonoyl glycine. AB - N-Arachidonoyl glycine was synthesized in a chemo-enzymatic process where glycine tert -butyl ester was acylated by arachidonic acid and the resulted ester was then de-protected to give the final product. Among various lipases tested and chosen for their ability to cleave fatty amides, that from Candida antarctica B gave the best results resulting in a 39% hydrolysis after 24 h. This enzyme was then used for the reverse N-acylation synthesis and gave a 75% product formation after 24 h using methyl ester of arachadonic acid as acyl donor and acetonitrile as solvent. Direct acylation of glycine gave less than 10% yield. PMID- 15289677 TI - Biocatalytic resolution of glycidyl aryl ethers by Trichosporon loubierii : cell/substrate ratio influences the optical purity of (R) - epoxides. AB - Glycidyl aryl ethers were resolved by using lyophilized cells of Trichosporon loubierii ECU1040 having epoxide hydrolase activity. The activity and enantioselectivity depended on the structure of the aryl group. Different cell/substrate ratios also influenced the optical purity of remaining substrate. An additional stability test of the whole-cell enzyme suggests that rapid deactivation of the epoxide hydrolase was the potential reason. (R)-Epoxides were prepared in gram amounts with optical purity of 87% - 99% ee. PMID- 15289678 TI - H(2) production and carbon utilization by Thermotoga neapolitana under anaerobic and microaerobic growth conditions. AB - H(2) production by Petrotoga miotherma, Thermosipho africanus, Thermotoga elfii, Fervidobacterium pennavorans, and Thermotoga neapolitana was compared under microaerobic conditions. Contrary to these previously reported strains being strict anaerobes, all tested strains grew and produced H(2) in the presence of micromolar levels of O(2). T. neapolitana showed the highest H(2) production under these conditions. Microscopic counting techniques were used to determine growth curves and doubling times, which were subsequently correlated with optical density measurements. The Biolog anaerobic microtiter plate system was used to analyze the carbon source utilization spectrum of T. neapolitana and to select non-metabolized or poorly metabolized carbohydrates as physiological buffers. Itaconic acid was successfully used as a buffer to overcome pH-induced limitations of cell growth and to facilitate enhanced production of CO-free H(2). PMID- 15289679 TI - Single-step recovery of ephedrine hydrochloride from raw materials using expanded bed adsorption. AB - Expanded bed adsorption, using a cation resin 001 x 7 Styrene-DVB, was used to recover and purify ephedrine hydrochloride from a powdered herb. The axial liquid phase dispersion coefficient was about 10(-5) m(2) s(-1) and the recovery yield and purification reached 86% and 22, respectively. Compared with using conventional extraction with dimethylbenzene, this method is safer and also more efficient. PMID- 15289680 TI - Fermentative capacity of baker's yeast exposed to hyperbaric stress. AB - Baker's yeast suspensions were incubated at different pressures (from 1 bar to 6 bar) and different gases [air, O(2) and a mixture of 8% (v/v) CO(2), 21% O(2) and N(2)]. Raising the air pressure from 1 bar to 6 bar stimulated cell growth but had no effect on leavening ability or viability of the cells. A 50% reduction of the CO(2) produced in dough occurred with 6 bar O(2) which also stopped growth. The fermentative capacity of the cells was stimulated by the cells exposure to increased CO(2) partial pressure up to 0.48 bar. PMID- 15289681 TI - Production of sophorolipids by Candida bombicola grown on soy molasses as substrate. AB - Sophorolipids (SLs) were produced from Candida bombicola using soy molasses and oleic acid as co-substrates. The purified SLs were obtained at 21 g l(-1) and were 97% in lactone form. The major SL constituent (81% relative abundance) of the product mixture contains an oleoyl chain. The surface properties of the SLs obtained from the soy molasses/oleic acid fermentation had minimum surface tension values of 37 mN m(-1) (pH 6) and 38 mN m(-1) (pH 9), and critical micelle concentration values of 6 mg l(-1) (pH 6) and 13 mg l(-1) (pH 9). PMID- 15289682 TI - Cloning and characterization of the bovine testicular PH-20 hyaluronidase core domain. AB - The core nucleotide sequence of bovine (Bos taurus) testicular PH-20 hyaluronidase was cloned using one step RT-PCR. The 5' and 3' regions were cloned separately and a sequence overlap of 124 bp facilitated the fusion of these two fragments by overlapping PCR, resulting in a concatenated sequence of 1422 bp. This nucleotide sequence and its deduced amino acid sequence were compared to homologous sequences from eight other mammal species. The bovine sequences were most similar to those of the pig, Sus scrofa (swine Spam1: 79.1% nucleotide and 70.1% amino acid similarity) and least similar to sequences from the Norway rat, Rattus norvegicus (murine Spam1: 61% nucleotide and 53.3% amino acid similarity). A phylogenetic analysis joined the red fox (Vulpes vulpes) sequence as sister to the bull-pig pair. Twelve cysteine residues were conserved among all nine aligned amino acid sequences and five proposed glycosylation sites have been identified. The feasibility of developing an effective, low-cost bovine PH-20 expression system is discussed in light of these new data. PMID- 15289683 TI - Rotational malalignment of the tibia following reamed intramedullary nail fixation. AB - OBJECTIVE: To determine the incidence and severity of tibial malrotation following reamed intramedullary nail fixation as measured by computerized tomography and to determine the repeatability of computed tomography measurement in the assessment of rotational malreduction. DESIGN: Prospective cohort. SETTINGS: Level 1 trauma center. PARTICIPANTS: Twenty-five consecutive patients with 25 tibial shaft fractures. INTERVENTION: All patients were treated with reamed intramedullary nailing. Appropriate radiographs and a postoperative lower extremity computed tomography scan were obtained for each patient who consented to the study. MAIN OUTCOME MEASURE: Rotational alignment of affected tibia as compared to a version of the normal contralateral limb. Malrotation was defined as an internal/external rotation deformity greater than 10degrees. RESULTS: Malrotation, comparing the fractured limb to normal limb, was determined using a similar measurement method previously described in the literature. Two patients declined inclusion, and in one case, the computed tomography was not acceptable for analysis. Malrotation, comparing the fractured limb to the normal limb, was determined using the measurements from axial computed tomography images. Results revealed a mean absolute rotational difference of 6.7degrees (SD +/-6.3degrees). Rotational malreduction ranged from 15degrees of internal rotation to 22degrees of external rotation. Five of the 22 tibia (22%) were malrotated greater than 10degrees. A larger degree of deformity was seen with certain injury patterns. The intraobserver and interobserver repeatability testing revealed a mean absolute difference between paired malrotation calculations of 3.4degrees and 3.9degrees, respectively, and a repeatability coefficient of 8degrees for both. CONCLUSION: Computed tomography measurement is a repeatable method of assessing tibial torsion and in this study revealed a significantly higher incidence of rotational malreduction than that previously reported in the literature. PMID- 15289684 TI - Rotational malalignment after intramedullary nailing of femoral fractures. AB - OBJECTIVES: Intramedullary nailing has been accepted as the treatment of choice for femoral shaft fractures. The aim of our study was to determine the incidence and implications of rotational malalignment after intramedullary nailing using computed tomography measurements. DESIGN: Cohort study. SETTING: Patients who postoperatively visited the orthopaedic outpatient and radiology clinics. PATIENTS: Seventy-six patients, 59 men and 17 women, with a mean age of 28.4 years (15-88). INTERVENTION: Patients treated on a fracture table with an antegrade reamed AO nail (n = 46) or Grosse Kempf nail (n = 30) for a unilateral femoral shaft fracture between 1988 and 1998 were included in the study. MAIN OUTCOME MEASUREMENTS: Patients filled out a questionnaire concerning pain, daily activities, and sport. Oxford, Western Ontario and McMaster University osteoarthritis index, and Harris Hip and Knee Society scores were obtained. Physical exams and computed tomography measurements were established. RESULTS: Twenty-one patients (28%) were found to have a rotational malalignment of 15degrees or more. There was no significant difference in rotational deformity with either the AO or Grosse Kempf nail. The incidence of malrotation was independent of the fracture level. Patients with a torsional deformity had difficulties with more demanding activities like running, sports, and climbing stairs. Patients with an external rotational malalignment (n = 12) have more functional problems than patients with an internal rotational malalignment (n = 9). Clinically determined rotation differences are not accurate (+/-20degrees) compared with the established computed tomography measurements. CONCLUSIONS: Rotational malalignment after intramedullary nailing for femoral fractures is found in 28% of the patients in this study. These patients have difficulties with more demanding activities, especially when they have an external torsional deformity. PMID- 15289685 TI - Mismatch of current intramedullary nails with the anterior bow of the femur. AB - OBJECTIVES: The anterior curvature of the femur affects intramedullary nail insertion, revision prosthesis design, and the biomechanics of the proximal femur. Two previous studies, using small numbers of femurs, determined femoral curvature and showed that it was significantly greater than that of the several intramedullary nails they evaluated. In this study, the curvature of 948 femurs (474 matched pairs) was determined and compared with current intramedullary nails. The correlation of femoral curvature to age, gender, femoral size, and race was also evaluated. SETTING: Museum skeletal collections and a hospital biomechanics laboratory. METHODS: The curvature of 892 femurs (446) from the skeletal collections of 2 museums was measured by processing the digital images of the femurs with a computer curve-fitting program. Fifty-six additional, embalmed femurs (28 pairs) from our collection were also digitally imaged and then radiographed and their medullary curvatures similarly determined for comparison. Curvatures of 8 current antegrade intramedullary nails and 3 long stemmed femoral hip prostheses were obtained from manufacturers and confirmed by measurements from their templates after digitization. RESULTS: We found the average femoral anterior radius of curvature was 120 cm (+/- 36 cm). There was no effect of age on femoral curvature nor was there a correlation between femoral width or femoral length to curvature. Black donor femurs had less curvature than white donor femurs (P < 0.001). There was close correlation (r = 0.967) between the femoral curvatures determined from the digital images and the radiographs. Radii of curvature of the intramedullary nails ranged from 186 to 300 cm (eg, straighter than the femurs). CONCLUSIONS: There was a large mismatch between the curvature of some current antegrade intramedullary nails and the average femur. Although this is only 1 factor affecting nail insertion, the mismatch warrants a reappraisal of these intramedullary nail designs. CLINICAL RELEVANCE: Because ease of intramedullary nail insertion and possibility of cortical comminution are determined by a number of factors including insertion point and fracture location, it appears that a decrease in radii of curvature (less straight) of current nail designs is warranted, particularly for those larger diameter nails designed for hip fracture stabilization that have greater rigidity due to design or material. PMID- 15289686 TI - Intramedullary nailing of femoral fractures in children through the lateral aspect of the greater trochanter using a modified rigid humeral intramedullary nail: preliminary results of a new technique in 15 children. AB - OBJECTIVES: To evaluate the clinical results of intramedullary nailing of femoral shaft fractures using a rigid intramedullary nail placed through the lateral aspect of the greater trochanter in older children and adolescents. DESIGN: A retrospective study was carried out evaluating all skeletally immature patients with femoral shaft fractures treated using a modified rigid humeral intramedullary nail. PATIENTS/PARTICIPANTS: Fifteen children and adolescents with displaced femoral diaphyseal fractures and open physes. INTERVENTION: Femoral shaft fractures in children and adolescents were stabilized using a modified humeral intramedullary nail placed through the lateral aspect of the greater trochanter. MAIN OUTCOME MEASUREMENTS: Patients were evaluated to determine time to union, final fracture alignment, hospital stay, complications, clinical outcome, and proximal femoral changes including avascular necrosis or proximal femoral valgus with femoral neck narrowing. RESULTS: Fifteen patients were followed for a minimum of 1 year (range 70-157 weeks). The average age of the patients was 12 years and 5 months (range 8 years and 2 months-17 years and 1 month). All fractures healed at a mean of 7 weeks (range 5-14 weeks) after fracture. The average hospital stay for patients with isolated femur fractures (8/15) was 2.8 days (range 1-5 days). At an average follow-up of 141 weeks (range 70-326 weeks), no patient had developed avascular necrosis, femoral neck valgus, femoral neck narrowing, or other complications. CONCLUSIONS: The technique of intramedullary nailing in children through the lateral aspect of the greater trochanter seems to be safe, effective, and well tolerated by patients. PMID- 15289688 TI - Femoroacetabular impingement as a factor in the development of nonunion of the femoral neck: a report of three cases. AB - OBJECTIVES: Description of an anatomic condition where a femoroacetabular impingement was identified as the cause for the development of nonunion of the femoral neck. DESIGN: Retrospective analysis. SETTING: University hospital. PATIENTS: Three patients, aged 27 to 74 years, in whom, after exclusion of other known factors, a femoroacetabular impingement was identified as the cause for the nonunion of a femoral neck fracture. INTERVENTION: Surgical correction of the femoroacetabular impingement. MAIN OUTCOME MEASUREMENT: Intraoperative verification of femoroacetabular impingement. Healing of the femoral neck non union. RESULTS: In all 3 patients, femoroacetabular impingement was confirmed at surgery. After elimination of the cause for impingement, all nonunions went on to uneventful healing. CONCLUSIONS: A femoroacetabular impingement mechanism is proposed as a cause for nonunion of femoral neck fractures. Predisposing factors such as bulging at the fracture site or decreased femoral-neck offset should be addressed at the time of initial fracture treatment. PMID- 15289689 TI - Operative treatment for hip fractures in patients 100 years of age and older: is it justified? AB - OBJECTIVES: To evaluate a unique group of elderly patients over 100 years of age who had hip fractures. DESIGN: Retrospective database analysis. SETTINGS: Academic teaching hospital. PATIENTS: All patients who had hip fractures between January 1990 and December 2001 and were over 100 years old were included. INTERVENTION: In this study, we evaluated the age, gender, type of fracture, type of treatment, background disease, rehabilitation, and time until death of all patients over 100 years, whether treated operatively or nonoperatively. RESULTS: Twenty-three patients (17 females and 6 males) were identified with ages ranging from 100 to 107 (mean: 101.8). The group had 4 subcapital and 19 pertrochanteric fractures and between 1 and 4 major background diseases. Four patients were treated nonoperatively (1 due to major pneumonia and 3 refused the operative procedure). Three of those 4 patients died in the same month of admission, and 1 patient died during the second month. Among the 19 patients who underwent operation, 17 patients have died, living between 0 and 78 months (mean: 13.8) postoperatively. Two are still alive (21 and 45 months) after the operation. Eight patients died prior to 6 months, and 11 lived more than a year after the operation. A comparison between these 2 groups showed greater major background disease in the patients who died prior to 6 months (P < 0.05). CONCLUSIONS: Most hip fractures in patients over 100 years of age are pertrochanteric. Patients with 2 or more major background diseases have an increased risk for dying in the first 6 months after the operation. Most patients having operations in this age group had a postoperative reduction in mobility status and in performing basic activities of daily living. PMID- 15289690 TI - Osteogenic protein-1 induces bone formation in the presence of bacterial infection in a rat intramuscular osteoinduction model. AB - OBJECTIVE: Evaluate the ability of osteogenic protein-1 to induce formation of de novo bone in the presence of bacterial infection and metal in an intramuscular osteoinduction model in the rat. DESIGN: Prospective experimental design with assessment time points of up to 4 weeks. SETTING: Intramuscular pocket surgically created along each side of the spine. ANIMALS: One-hundred-twenty adult male Sprague-Dawley rats. INTERVENTIONS: Each intramuscular pocket received 0, 10, or 25 microg of osteogenic protein-1 combined with a lyophilized collagen carrier, and 0 or 5 x 10 colony-forming units of Staphylococcus aureus. Pockets in 48 animals received a metal implant. Animals were killed at 1, 2, 3, or 4 weeks. MAIN OUTCOME MEASUREMENTS: High-resolution radiographs of resulting nodules of bone/soft tissue were digitized, and areas of newly formed bone were quantified using an image analysis workstation. The nodules were decalcified for histology, and calcium content of the decalcifying solution was quantified by flame atomic absorption spectrophotometry. RESULTS: There were minimal levels of calcium and area of new bone formation in nodules from pockets containing collagen carrier without osteogenic protein-1, for both infection and noninfection conditions. Calcium content and area of newly formed bone were significantly greater: 1) in infected pockets with osteogenic protein-1, compared to infected pockets without osteogenic protein-1; and 2) in noninfected pockets with osteogenic protein-1, compared to infected pockets with osteogenic protein-1. The presence of metal did not have a significant effect. CONCLUSION: Osteogenic protein-1 maintained its osteoinductive capability in a contaminated intramuscular pocket in the rat, albeit at a lower level than without infection. This finding supports further study using a more clinically realistic model. PMID- 15289691 TI - Avoiding malunion with 95degrees fixed-angle distal femoral implants. AB - Malunion of the distal end segment of a fractured femur is a common complication after open reduction and internal fixation when using 95degrees fixed-angle devices. Incorrect positioning of the alignment Kirschner wires can result in implant malposition and subsequent malunion. This article discusses the relevant distal femoral anatomy and the technical points necessary to avoid a malunion when using 95degrees fixed-angle devices. PMID- 15289692 TI - Olecranon osteotomy for exposure of fractures and nonunions of the distal humerus. AB - Although olecranon osteotomy provides excellent exposure of the distal humerus, enthusiasm for this approach has been limited by reports suggesting numerous complications. It has been suggested that specific techniques for creating and repairing an olecranon osteotomy may help limit complications. This paper describes a technique for olecranon osteotomy using an apex, distal, chevron shaped osteotomy, Kirschner wires directed out the anterior ulnar cortex distal to the coronoid process and bent 180degrees and impacted into the olecranon proximally, and two 22- gauge, figure-of-eight, stainless steel tension wires. A single surgeon used this technique for exposure of a fracture (16 patients) or nonunion (29 patients) of the distal humerus in 45 consecutive patients. One patient returned to activity too soon, had loosening of the wire fixation, and required a second operation for plate fixation of the ulna. The remaining 44 osteotomies (98%) healed with good alignment within 6 months. There were no broken or migrated wires prior to healing. Twelve patients (27%) had removal of the wires used to repair the olecranon: in 6 patients, this was for symptoms related to the wires (13%); 1 for septic olecranon bursitis, and 5 at the time of another procedure (elbow capsular release in 4 patients and submuscular ulnar nerve transposition in 1). Olecranon osteotomy can be used for exposure of the distal humerus with a low rate of complications when specific techniques are used. PMID- 15289693 TI - Secondary neurologic deficit due to unrecognized spine instability in multitrauma casualties: a report of three cases. AB - We present three cases of major multiple injuries in which delayed diagnosis of spinal instability resulted in neurologic deficit. The spinal injuries, once diagnosed, were treated by urgent surgical stabilization, which resulted in prompt and full recovery. A set of recommendations is proposed to prevent these potentially disastrous consequences. PMID- 15289694 TI - Atrophic femoral nonunion with bone loss: treatment with monorail transport: a case report. AB - Nonunions are an uncommon outcome of femoral fractures. Atrophic nonunions with a leg length discrepancy secondary to bone loss are often the most difficult to treat, and the treatment options are limited. We present a case that uses concomitant monolateral external fixation and intramedullary nailing to heal a nonunion and perform a simultaneous 7-cm lengthening procedure in a 33-year-old female. PMID- 15289695 TI - Management of pathologic fractures of the proximal femur: state of the art. AB - Metastases to bone are the most common cause of a destructive lesion of the skeleton in an adult. The proximal femur is the most commonly affected bone with metastatic disease in the appendicular skeleton. A systematic approach to patient management is critical, so as to avoid complications that will delay systemic therapy. The orthopedic traumatologist is often the first physician to see the patient with a pathologic fracture. As a result, the surgeon must be aware of the indications for resection versus internal fixation, as well as options for reconstruction. Polymethyl methacrylate and curettage can be useful in the appropriately selected patient. Postoperative external beam irradiation can significantly reduce disease progression and subsequent loss of fixation. A multidisciplinary approach to this patient group will help optimize prognosis as well as function. This article reviews the evaluation, management, and complications of treatment of pathologic fractures of the proximal femur, with an emphasis on metastatic disease and multiple myeloma. PMID- 15289696 TI - Infected nonunion of the ulna: Opinion: ilizarov external fixation and bone transport. PMID- 15289697 TI - Infected nonunion of the ulna: Opinion: open reduction and internal fixation and resection tricortical bone grafting. PMID- 15289698 TI - Fractures of the shaft of the ulna. AB - BACKGROUND: The optimal management for ulnar shaft fractures remains debatable. Investigators have advocated nonoperative management as well as internal fixation of these fractures. OBJECTIVE: The primary objective of this study was to determine the effect of alternative management strategies of fractures of the ulnar shaft on rates of union, infection, and functional outcomes. A secondary objective was to examine outcomes after alternative strategies in managing patients with bone defects. PMID- 15289699 TI - Oral risperidone plus oral lorazepam versus standard care with intramuscular conventional neuroleptics in the initial phase of treating individuals with acute psychosis. AB - Although atypical antipsychotics are now considered first line treatments for schizophrenia, intramuscular (i.m.) conventional neuroleptics are often still considered necessary in emergency treatment of acute psychoses. This European, multicentre, open-label, active-controlled trial compared oral risperidone plus oral lorazepam to standard care with i.m. conventional neuroleptics with or without lorazepam in the emergency treatment of acutely psychotic patients. Patients were allowed to choose either oral risperidone (a single dose of 2 mg and 2.0-2.5 mg lorazepam; 121 patients) or standard i.m. treatment (conventional neuroleptic with or without lorazepam; 105 patients). No additional treatment was allowed for 2 h. Primary outcome was the percentage of patients with treatment success (asleep or at least much improved on Clinical Global Impression-global improvement scale) 2 h after treatment initiation. Baseline characteristics were similar in both treatment groups. Oral risperidone plus oral lorazepam was more successful at 2 h (66.9%) and significantly non-inferior compared to standard i.m. care (54.3%; P=0.0003), and the incidence of extrapyramidal symptoms (EPS) was lower (1.7%) compared to standard i.m. care (9.5%). In acutely psychotic patients requiring emergency treatment, oral risperidone/oral lorazepam was at least as effective as i.m. conventional neuroleptic treatment with or without lorazepam. Oral risperidone plus lorazepam rapidly reduces symptoms, including aggression, and causes fewer EPS. PMID- 15289700 TI - Absence of discontinuation symptoms with agomelatine and occurrence of discontinuation symptoms with paroxetine: a randomized, double-blind, placebo controlled discontinuation study. AB - The effects of an abrupt interruption of agomelatine, a new melatonergic/serotonergic antidepressant, were explored in a double-blind, placebo-controlled study. Paroxetine was used as active control. After 12 weeks of double-blind treatment with agomelatine 25 mg/day or paroxetine 20 mg/day, sustained remitted depressed patients were randomized for 2 weeks, under double blind conditions, to placebo or to their initial antidepressant treatment. Discontinuation symptoms were assessed at the end of the first and second week of discontinuation with the Discontinuation Emergent Signs and Symptoms (DESS) checklist. One hundred and ninety-two sustained remitted patients were randomized to the 2-week discontinuation period. Patients who discontinued agomelatine did not experience more discontinuation symptoms than those who continued on agomelatine. Patients who discontinued paroxetine for placebo experienced significantly more DESS discontinuation symptoms, during the first week, compared to those who continued with paroxetine (respective mean number of emergent symptoms: 7.3+/-7.1 and 3.5+/-4.1, P<0.001). No significant difference was shown between the continuing and interrupting groups in the second week of discontinuation. By contrast to paroxetine, abrupt cessation of agomelatine is not associated with discontinuation symptoms. PMID- 15289701 TI - Maintenance of long-term efficacy and safety of quetiapine in the open-label treatment of schizophrenia. AB - As schizophrenia is a chronic disorder, it is important that treatment be given over a long period of time to avoid relapse. Quetiapine, an atypical antipsychotic, has established efficacy and good tolerability in the short-term treatment of schizophrenia. This study investigated the long-term efficacy and safety of quetiapine in 674 patients with schizophrenia using combined data from the open-label extension phase of four Phase IIIa trials. The results showed that quetiapine, at a mean daily dose of 472.4 mg, provided progressive improvement and maintenance in the Brief Psychiatric Rating Scale total, positive- and negative-symptoms cluster, Clinical Global Impression Severity of Illness, and Scale for the Assessment of Negative Symptoms total scores over 208 weeks and beyond. Furthermore, quetiapine was well tolerated throughout the study period, with a low incidence of extrapyramidal symptom-related adverse events. In conclusion, quetiapine may be a suitable therapy in the long-term treatment of schizophrenia. PMID- 15289702 TI - Antipsychotic drugs in mania: factors predicting use of antipsychotic medication following inpatient treatment of mania. AB - Inpatients treated for mania are frequently discharged on antipsychotics. There has been disagreement in clinical guidelines concerning their use, and there is little knowledge of the factors predicting this. The aim of the present study was to identify those factors predicting discharge on typical antipsychotic medication after hospital admission for mania. Data were collected retrospectively from the hospital notes of 74 patients who had 100 consecutive psychiatric hospital admissions for mania during the 1990s. Univariate and multivariate analyses were used to estimate odds ratios. The rate of discharge on typical antipsychotics was 71.6% (53 out of the 74 episodes included in the analyses) (95% confidence interval 61.3-81.9). Three factors were found to predict this with odds ratios significant at the P<0.05 level: (i) use of typical antipsychotics during the first week of admission [odds ratio (OR)=13.3]; (ii) voluntary admission (OR=4.2); and (iii) male gender (OR=8.4). There was no evidence that psychotic symptoms or aggressive behaviour were associated. In conclusion, the use of antipsychotics early in the episode strongly predicted discharge on antipsychotics. It appears that clinicians use antipsychotics for their anti-manic effects and patients are often thought to still need the drugs at the time of discharge. PMID- 15289703 TI - Risperidone in combination with mood stabilizers for acute mania: a multicentre, open study. AB - The primary aims of this study were (i) a replication of the effectiveness and tolerability of risperidone in the treatment of patients with acute mania in a very large cohort in a naturalistic treatment setting and (ii) to extend the data on the effect and tolerability of risperidone in the treatment of patients with acute mania to an Asian population. A total of 909 patients fulfilling DSM-IV criteria of bipolar disorder (current manic and hypomanic episode) entered this large, open, multicentre study. The Young Mania Rating Scale (YMRS), Clinical Global Impression (CGI) and Simpson-Angus Rating Scale (SARS) were measured at baseline and weeks 1, 3 and 6, for the assessment of effectiveness and extrapyramidal symptoms. This study showed a statistically significant reduction of scores on the YMRS and CGI-severity (mean change=-23.5+/-11.8, P<0.0001; mean change=-2.7+/-1.5, P<0.0001, respectively) from baseline to endpoint (week 6). The number of patients with a 50% reduction or more in the YMRS and CGI-severity scores was 693 (77.8%) and 630 (70.7%) at endpoint, respectively. There were no statistically significant increments of scores on SARS. Risperidone was generally well tolerated. The present larger open study indicates that risperidone add-on therapy is effective and tolerable in treatment of bipolar disorder, replicating results in various controlled and uncontrolled studies from Western countries. PMID- 15289704 TI - Evidence based review of escitalopram in treating major depressive disorder in primary care. AB - The study aimed to summarize clinical data for escitalopram in the treatment of major depressive disorder in primary care. Medline, Embase and Cochrane databases were searched for randomized controlled trials of escitalopram (10-20 mg/day for 8 weeks) versus other antidepressants in therapeutic doses or placebo. Patients were required to have had moderate/severe depression, with Montgomery-Asberg Depression Rating Scale (MADRS) scores recorded at baseline and 8 weeks. Outcomes examined were remission rates (MADRS/=50% decrease from baseline in MADRS at week 8). Data were combined using a random effects meta analytic model. Of the 15 studies identified, 11 were rejected (five not primary care, four duplicate reports, one lacked 8-week MADRS scores, one not depression) and four were accepted (n=1472 patients). The four studies had nine arms, four for escitalopram (n=654), two for citalopram (n=333), one for venlafaxine-XR (n=142) and two for placebo (n=343). Remission rates for escitalopram were superior to placebo (48.7% versus 37.6%, P=0.003) and citalopram (52.8% versus 43.5%, P=0.003) but similar to venlafaxine-XR (P=0.97). Response rates were superior to placebo (48.7% versus 43.1%, P<0.001) and citalopram (62.5% versus 49.5%, P=0.001) but not venlafaxine-XR (P=0.52). Adverse events were comparable among active drugs (P<0.05). Remission rates for escitalopram were superior to placebo (48.7% versus 37.6%, P=0.003) and citalopram (52.8% versus 43.5%, P=0.003) but similar to venlafaxine-XR (P=0.97). Response rates were superior to placebo (48.7% versus 43.1%, P<0.001) and citalopram (62.5% versus 49.5%, P=0.001) but not venlafaxine-XR (P=0.52). Adverse events were comparable among active drugs (P>0.05). Remission and response rates of escitalopram in primary care are clinically superior to placebo and citalopram, but similar to venlafaxine-XR. Further head-to-head trials are warranted to verify these findings. A pharmacoeconomic analysis is also required to determine whether these clinical advantages for the patients translate into economic advantages for the health care system. PMID- 15289705 TI - Lormetazepam in depressive insomnia: new evidence of phase-response effects of benzodiazepines. AB - Benzodiazepines can shift the phase of circadian rhythms in mammalian species, but few data are available on their phase-response effects in humans, and on possible links between timing of administration and hypnotic efficacy. Using a placebo-controlled, cross-over design, we evaluated the hypnotic effect of lormetazepam 0.03 mg/kg and placebo in 38 inpatients who were affected by a major depressive episode. Patients were divided into three groups, receiving treatment at 18.00 h, 20.00 h or 22.00 h, respectively. Sleep and psychiatric symptoms were evaluated with self-administered scales and a sleep diary. The results demonstrate that active treatment significantly improved insomnia independent of the severity of depression, which remained unchanged. Timing of treatment influenced changes in timing of sleep observed with active treatment. Although sleep duration was equally improved in all treatment groups, patients who received treatment at 20.00 h showed an acute advance of sleep onset, with no changes in morning awakening. Patients who received treatment at 22.00 h showed an acute delay in morning awakening, with no changes of sleep onset. Finally, patients who received treatment at 18.00 h showed a non-significant trend in the same direction. These effects reverted with cross-over return to placebo. The perceived degree of improvement of insomnia was proportional to the advance in timing of sleep onset obtained with treatment. Our results suggest that the effects of lormetazepam on the subjective sleep of patients affected by a major depressive episode depend upon the timing of administration, and that improvement in subjective sleep is related to advance of sleep onset, and not to delay of morning awakening. PMID- 15289706 TI - Medical errors: lessons learned from industry. PMID- 15289715 TI - Electrotherapy promotes healing and microcirculation of infrapopliteal ischemic wounds: a prospective pilot study. AB - OBJECTIVE: To determine if high-voltage pulsed current (HVPC) electrotherapy augments ischemic wound healing and increases periwound microcirculation. DESIGN: A prospective, randomized, single-blinded, sham-controlled clinical trial was conducted on a homogenous subset of quasi-stable ischemic wounds. INTERVENTION: Active HVPC or sham HVPC was applied to wounds for a 14-week period. MAIN OUTCOME MEASURE: Wounds were monitored every 4 weeks, except 2 weeks between weeks 12 and 14, for wound area, wound appearance, and microcirculation, which was measured by transcutaneous oxygen (TcPO2) levels and laser Doppler flow. RESULTS: Ischemic wounds treated with active HVPC decreased in size, contrary to the expected increase in ischemic wound size that was observed in wounds in the control group (P <.05, Student t test; week 4). A trend toward smaller wound area occurred in wounds in the HVPC group compared with wounds in the control group (week 14). Among the HVPC group, an improvement in periwound microcirculation occurred at weeks 8 (P <.05, TcPO2; P <.01, laser Doppler) and 12 (P <.05, laser Doppler). These increases suggest that HVPC promotes arteriolar vasodilation and dermal capillary formation. HVPC was well tolerated. CONCLUSION: The results of this study demonstrate that HVPC decreased the area of ischemic wounds, reversing the expected increase in wound size, and improved microcirculation. The promising results of this pilot study require a larger Phase II study to confirm and generalize these findings. CLINICAL RELEVANCE: Electrotherapy may prove to be a relatively safe and effective complement to surgical revascularization to improve the odds of healing ischemic wounds and promoting limb salvage. PMID- 15289717 TI - Reversal of diabetic peripheral neuropathy and new wound incidence: the role of MIRE. AB - OBJECTIVE: To determine if improved foot sensitivity to the Semmes-Weinstein 10-g (5.07) monofilament, originally impaired because of diabetic peripheral neuropathy, might be associated with a reduced incidence of new diabetic foot wounds. DESIGN: Retrospective cohort study using a health status questionnaire. SUBJECTS: Sixty-eight individuals over age 64 with diabetes, diabetic peripheral neuropathy, and loss of protective sensation who had clinically demonstrable increases in foot sensation to the Semmes-Weinstein monofilament after treatment with monochromatic near infrared photo energy. MAIN RESULTS: After reversal of diabetic peripheral neuropathy following treatment with monochromatic near infrared photo energy, only 1 of 68 patients developed a new diabetic foot wound, for an incidence of 1.5%. Comparatively, the incidence previously reported in the Medicare-aged population with diabetes was 7.3%. CONCLUSIONS: Improved foot sensitivity to the Semmes-Weinstein monofilament in patients previously suffering from loss of protective sensation due to diabetic neuropathy appears to be associated with a lower incidence of new diabetic foot ulcers when compared with the expected incidence in the Medicare-aged population with diabetes. CLINICAL RELEVANCE: Therapeutic interventions that effectively improve foot sensitivity that has been previously diminished due to diabetic peripheral neuropathy may substantially reduce the incidence of new foot wounds in the Medicare-aged population with diabetes. PMID- 15289718 TI - Recognizing and managing venous leg ulcers. AB - PURPOSE: To provide physicians and nurses with an overview of the pathophysiology, assessment, diagnosis, and treatment of venous insufficiency and ulceration. TARGET AUDIENCE: This continuing education activity is intended for physicians and nurses with an interest in managing patients with venous insufficiency and ulceration. OBJECTIVES: After reading the article and taking the test, the participant will be able to: describe the anatomy, physiology, and pathophysiology of the lower extremity venous system; describe the assessment and diagnosis of venous insufficiency and ulceration; identify treatment options and teaching considerations for patients with venous insufficiency and ulceration. PMID- 15289720 TI - Decision on national coverage of electromagnetic therapy for wounds. PMID- 15289721 TI - Closing the gap: how to provide protein without increasing total calories. PMID- 15289722 TI - Experience with conformal proton therapy for early prostate cancer. AB - A study was conducted to evaluate the use of proton beam therapy for the treatment of organ-confined prostate cancer. This is a preliminary assessment of treatment-related morbidity and tumor response. Sixteen patients with T1-T2b prostate cancer underwent proton beam therapy. Acute and late toxicity was scored according to the National Cancer Institute Common Toxicity Criteria Grading System (version 2.0, April 1999) and to the Radiation Therapy Oncology Group grading system, respectively. Local control was assessed using magnetic resonance imaging (MRI) and prostate-specific antigen (PSA) values. Although skin toxicity and bladder irritability were commonly observed, none of the patients developed grade III or IV toxicity. Of 9 patients in whom the primary lesion was detected by MRI, partial response and no change (NC) was observed in 6 (66.7%) and 3 (33.3%) patients, respectively. Four patients presented normal PSA value before treatment due to the previous endocrine therapy. However, the other 12 patients with elevated PSA value before treatment showed complete response. No patients showed PSA failure within the median follow-up period of 11.9 months. Although longer follow-up is necessary, minimum toxicity and good short-term clinical responses were observed following proton beam therapy in T1-T2 prostate cancer patients. PMID- 15289723 TI - Oral fluconazole for empiric treatment of prolonged Fever in neutropenic patients: prospective study in 250 consecutive patients after stem cell transplantation. AB - Neutropenic patients who continue to be febrile despite adequate broad-spectrum antibacterial treatment require empirical antifungal therapy. The aim of the present study was to evaluate the safety and efficacy of oral fluconazole for empirical antifungal therapy in neutropenic patients with persistent fever. A prospective cohort design was used. The study sample included 250 consecutive patients with high-risk stage II, III, or responding metastatic breast cancer who received high-dose chemotherapy (HDC) with autologous peripheral blood progenitor stem cell transplantation. Those with neutropenic fever lasting more than 72 hours despite broad-spectrum antibacterial coverage were treated with fluconazole. Treatment was continued until fever dropped and/or neutrophil count recovered with blood cultures remaining negative. Antifungal treatment was required in 173 patients (69%). There were no cases of documented deep systemic fungal infection. Two patients (<1%) had positive blood cultures for fungi. None of the patients experienced toxicity related to fluconazole. There was one transplant-related death. Thirty-one patients (18%) were unable to complete the oral fluconazole protocol because of severe mucositis, and they received intravenous fluconazole at the same dose, with similar efficacy. Oral fluconazole is a safe and effective alternative to amphotericin B for empirical early antifungal treatment in persistent neutropenic fever in breast cancer patients undergoing HDC with autologous stem cell support. Further study of oral fluconazole and amphotericin B as empirical agents in other groups of patients with persistent neutropenic fever is warranted. PMID- 15289724 TI - Breast cancer associated with idiopathic thrombocytopenic purpura: a single center series of 10 cases. AB - The aim of this study was to define the characteristics of patients with idiopathic thrombocytopenic purpura (ITP) and breast cancer and discuss the relationship between these two diseases. Ten patients treated for breast cancer and presenting with ITP were screened for this study. The diagnosis of breast cancer was confirmed by biopsy or surgical sample. The diagnosis of ITP was defined by 1) platelet count less than 140.10(9)/l with normal or increased number of megakaryocytes on bone marrow aspirate, 2) after exclusion of thrombocytopenia-induced medication or disorders, and 3) absence of splenomegaly. ITP was diagnosed before breast cancer in three cases, concomitantly in three, and after the diagnosis of breast cancer in four cases. Platelet count and breast cancer showed an independent course in seven cases, and appeared to be correlated in the other three patients. No correlation was found between the development of ITP and tumor characteristics. In contrast, the median platelet count was 15.10(9)/l (range 3-26) for the 3 patients with a correlation between the course of ITP and breast cancer evolution and 70.10(9)/l (range 20-90) for the other cases (p = 0.05, Mann-Whitney U test). Breast cancers are associated with ITP, with a parallel course of the two diseases in one third of cases. This may suggest tumor-induced immunologic thrombocytopenia. PMID- 15289725 TI - ZD9331 as second- or third-line therapy in patients with advanced colorectal cancer: a phase II multicenter trial. AB - This study investigated the efficacy and tolerability of ZD9331 as second- or third-line treatment for patients with advanced colorectal cancer (aCRC). One hundred patients were recruited to the study: 45 in group 1 (failed first-line 5 FU-based regimen) and 55 in group 2 (failed first-line 5-FU-based regimen and second-line irinotecan). Patients received ZD9331 as a 30-minute intravenous infusion on days 1 and 8 of a 3-week cycle, and treatment continued until disease progression (PD) or withdrawal. After a median of 4 cycles of treatment, there were no objective responses in group 1 (N = 37), 25 (67.6%) patients had a best overall response of stable disease (SD), and 12 (32.4%) had PD. After a median of 3 cycles of treatment, there were 2 (4.5%) partial responses in group 2 (N = 44), 21 (47.7%) patients had a best overall response of SD, 20 (45.4%) had PD, and 1 (2.3%) had clinical progression. At data cut-off, 59.5% and 77.3% of patients in groups 1 and 2, respectively, had PD. The main adverse events were neutropenia (69%), fatigue (53%), nausea (46%), and diarrhea (40%), and most (72.3%) were grade I/II. ZD9331 demonstrated minimal antitumor activity, and manageable toxicity, in the second- or third-line treatment of aCRC. PMID- 15289726 TI - Neoadjuvant chemoradiation with tegafur in cancer of the pancreas: initial analysis of clinical tolerance and outcome. AB - The early institutional experience in the neoadjuvant treatment of potentially resectable pancreatic carcinoma using oral Tegafur as radioenhancing agent is analyzed. Fifteen patients (10 male and 5 female, mean age of 61 years) were treated over a 30-month period. Tegafur dose was 1,200 mg/d along the external radiotherapy period (45-55 consecutive days). Preoperative radiotherapy achieved a total dose of 45 to 50 Gy (1.8 Gy/d). Intraoperative electron boost (10-15 Gy) was delivered at the time of surgery. Hematologic tolerance showed a significant decrease of neutrophil and platelet counts from the outset to the end of the neoadjuvant period (p = 0.001 and p = 0.004, respectively). Five grade III vomiting episodes (33%) were also registered. In 9 patients (60%), surgical resection was performed after chemoradiation. Three complete pathologic responses (pT0 specimens) were identified; in seven cases, the resection achieved tumor free surgical margins of the specimen. With a median follow-up of 21 months, median survival time was 17 months, with actuarial rates of 45% at 1 year and 24% at 3 years. Median survival for the resected patients was 23 months, and for the unresected patients median survival was 8 months (p = 0.02). The overall median survival in completely resected patients was 28 months, with a 71% survival rate at 1 and 3 years. It is concluded that the treatment scheme described is feasible and acceptably tolerated. The use of oral Tegafur seems to induce results similar to those of other therapeutic protocols using intravenous radioenhancing chemotherapy. PMID- 15289727 TI - Stage III non-small-cell lung cancer treated with high-dose hyperfractionated radiation therapy and concurrent low-dose daily chemotherapy with or without weekend chemotherapy: retrospective analysis of 301 patients. AB - We investigated the outcome in patients with stage III non-small-cell lung cancer (NSCLC) treated with high-dose hyperfractionated radiation therapy (Hfx RT) and concurrent chemotherapy (CHT) consisting of carboplatin (C) and etoposide (E). During three prospective randomized phase III and one prospective phase II study enrolling a total of 536 patients, 301 patients were treated with high-dose Hfx RT (69.6 Gy) and either low-dose daily CE (50 mg each) (n = 163) or daily CE (30 mg each) accompanied by "weekend" CE (100 mg of each on Saturdays and Sundays) (n = 138). The median survival time for all 301 patients is 22 months and 5-year survival is 24%. Median local recurrence-free survival (LRFS) time is 21 months and 5-year local recurrence-free survival is 32%. The median time to distant metastasis is 25 months, and 5-year distant metastasis-free survival (DMFS) is 35%. Only the type/schedule of CHT administration did not influence overall survival, LRFS, and DMFS. On multivariate analyses using these three endpoints, age stage, interfraction interval, and type/schedule of CHT administration did not predict survival, LRFS, and DMFS, while gender, KPS, and weight loss did. Only high grade hematologic toxicity was more frequent in weekend CHT group. High dose Hfx RT and concurrent low-dose daily CE with or without weekend CE is an active treatment approach in stage III NSCLC that led to high overall survival, LRFS, and DMFS rates. PMID- 15289728 TI - Severe radiation dermatitis is related to Staphylococcus aureus. AB - Acute radiation dermatitis commonly occurs following local radiation therapy for various cancers and, when severe, may necessitate disruption of treatment. Intense inflammatory reaction may result in a breakdown of the skin's barrier function and accompanying bacterial colonization. Bacterial superantigens may exacerbate inflammation through activation of T-cells and subsequent cytokine release. We report six cases of severe radiation dermatitis in cancer patients. Four of the six grew pathogenic bacteria, and three had psoriasiform or eczematous reactions at distant sites. Both the radiation dermatitis and the distant cutaneous reactions resolved rapidly on a combination of topical steroids and oral plus topical antibiotic therapy. We suggest that staphylococcal superinfection of acute radiodermatitis intensifies the inflammatory process and hinders repair of the epidermal barrier. Patients with acute radiation dermatitis should be investigated for secondary infection. We emphasize the importance of including topical and oral antibiotic therapy in conjunction with topical corticosteroids to eradicate infection as well as hasten repair of the skin's barrier function. These cases are presented to call attention to the role of Staphylococcus aureus in the pathogenesis of severe radiation dermatitis and the need to include appropriate antibiotic therapy based on culture in the management of acute radiation dermatitis. PMID- 15289729 TI - Effect of capecitabine on mean corpuscular volume in patients with metastatic breast cancer. AB - Capecitabine is a novel oral chemotherapy agent designed to generate 5 fluorouracil (5-FU) preferentially in tumor tissue, and is the most effective therapy for anthracycline and taxane-resistant breast cancer. Macrocytosis has not been previously reported in association with capecitabine therapy. We performed a retrospective review of consecutive metastatic breast cancer (MBC) patients receiving standard 21-day cycles of oral capecitabine therapy at a single center during the year 2000. Patients were assessed prior to each cycle with clinical examinations and complete blood counts. Seventy-six women (median age 52 years, median follow-up 273 days) met inclusion criteria for the study. Prior to treatment, the average mean corpuscular volume (MCV) was 91.6 fl (normal range 80-100 fl). During chemotherapy, MCV increased in a dose-dependent and time dependent manner. Fifty-seven percent of study patients developed macrocytosis (MCV > 100 fl) while on capecitabine therapy; 85% of women who received at least nine cycles of therapy exhibited macrocytosis. Development of macrocytosis was independent of anemia, thrombocytopenia, neutropenia, liver metastasis, and hepatic dysfunction; however, increases in MCV were more pronounced in 5-FU-naive patients. Alternative causes of macrocytosis were not identified in patients without coexisting anemia. We conclude that capecitabine therapy produces time dependent and dose-dependent macrocytosis in MBC patients. However, macrocytosis was not associated with anemia or overt myelosuppression. When capecitabine treated breast cancer patients develop macrocytosis in the absence of anemia, investigations of other causes of macrocytosis are not warranted. PMID- 15289730 TI - Prospective phase II trial of PFL-induction chemotherapy followed by definitive local treatment for advanced squamous cell carcinoma of the head and neck: 10 year follow-up. AB - Reported is an analysis of overall survival at 10 years of 102 patients with advanced squamous cell carcinoma of the head and neck (SCCHN) who were enrolled in a prospective phase II trial of high-dose cisplatin, 5-fluorouracil (5-FU), and high-dose leucovorin (PFL) induction chemotherapy followed by surgery and/or definitive radiation therapy (RT) between 1987 and 1991. Initially, 14 patients underwent primary site (PS) and neck surgery irrespective of the clinical response to PFL. The high rate of clinical and pathologic complete response (CR) to PFL prompted a switch from PS surgery to definitive RT. Of 102 patients, 18 (17.6%) who completed PFL and local-regional treatment for SCCHN between 1988 to 1991 were alive in December 2000. Among these, 1 of 14 patients (7%) who had undergone PS resection and 17 of 85 (20%) who were treated after PFL with definitive RT but without PS surgery were alive at 10 years. Median survival time was higher in the nonsurgical group (98.9 vs. 51.9 months). Subset analysis suggested that patients with oropharyngeal PS had the longest median survival (108.6 months). The oropharyngeal patients represented the 61% (11/18) of the long-term survivors with organ preservation. An organ preservation approach for patients with advanced SCCHN who demonstrated PS CR to chemotherapy demonstrated a trend to improved overall survival. PMID- 15289731 TI - Hepatic artery infusion chemotherapy for metastatic colorectal cancer to the liver at the lahey clinic: comparison between two methods of treatment, surgical versus percutaneous catheter placement. AB - This study updates our experience with hepatic artery infusion chemotherapy for colorectal liver metastases at the Lahey Clinic. It compares surgical versus percutaneous catheter methods, employing an external pump. The surgical series (SS) consisted of 58 patients (1970-1995) treated with floxuridine (FUDR), 20 mg/d for 4 to 5 weeks (modified in 1985; 2-week cycles). Percutaneous series (PS) consisted of 42 patients (1976-1995) treated with fluorouracil (5-FU), 20 mg/d for 10 days followed by a floxuridine (FUDR) schedule as with SS. Analysis consisted of tumor response, survival, and toxicity data between the two methods. Response rates showed no significant difference, SS (34%) and PS (48%) (P = 0.22). There were no significant differences in survival from treatment until death in SS (n = 58) of 13 months versus PS (n = 42) of 10.6 months (P = 0.39), from diagnosis until death, SS being 28.4 months versus PS of 26.4 months (P = 0.71) and from metastases until death, SS being 17.4 months versus PS of 22.2 months (P = 0.35). Hepatic toxicity was similar, but there was increased bone marrow toxicity, mucositis, and diarrhea for the PS. Response rates are similar for both our SS and PS and to that reported in recently randomized surgical trials. Hepatic artery infusion chemotherapy with FUDR by percutaneous catheter placement may be as effective as surgical catheter placement for colorectal liver metastases, but further study is needed. PMID- 15289732 TI - Comparison of the antiproliferative properties of antiestrogenic drugs (nafoxidine and clomiphene) on glioma cells in vitro. AB - The antitumoral activity of nonsteroidal antiestrogens on C6 and low passage of human glioma cells was investigated. Tamoxifen and its metabolite, 4 hydroxytamoxifen, did not influence viability of the human cells, but tamoxifen had a limited antiproliferative effect on C6 cells (IC50: 49 micromol/l). The derivatives of tamoxifen, nafoxidine and clomiphene, caused reduction of living cell number in a dose-dependent manner. These two drugs showed differences in their potency following 24-hour incubation in a humidified atmosphere with 37 degrees C and 5% CO2. Obtained from a tetrazolium-formazan growth rate assay, IC50 of nafoxidine for C6 cells was calculated as 44 micromol/l and for the human cells as 16.5 micromol/l. The calculated IC50 dose of clomiphene for C6 is 16 micromol/l and for the human cells 13 micromol/l. Compared to the other drugs we used, it is clear that clomiphene is the most efficient inhibitor of C6 and the human glioma cells. These preliminary results suggest that nafoxidine and clomiphene possess antiproliferative effect on two different sources of glioma cells and therefore, instead of tamoxifen, multiple activities of these drugs may enable their use in combination therapy of glioblastoma malignancies. PMID- 15289733 TI - Acute and chronic results of adjuvant radiotherapy after mastectomy and Transverse Rectus Abdominis Myocutaneous (TRAM) flap reconstruction for breast cancer. AB - A retrospective review of the treatment of 15 breast cancer patients who received postoperative radiotherapy after a mastectomy and transverse rectus abdominis myocutaneous (TRAM) flap breast reconstruction was undertaken to determine the effects of postoperative irradiation on flap viability and cosmesis. Fourteen patients had pedicle TRAM flaps, and one patient had a free TRAM flap. Surgical complications, acute and chronic side effects of radiotherapy, and cosmetic outcome were evaluated. The median interval between the TRAM flap procedure and radiotherapy was 7 months. The median total radiation dose was 60 Gy. All patients underwent three-dimensional radiotherapy treatment planning to determine the optimal dose distribution. Mild erythema developed in 9 patients (60%), moderate erythema developed in 2 (13%), and severe erythema developed in 1 (7%). Dry desquamation developed in 6 patients (40%), whereas moist desquamation developed in none. At median follow-up of 26.4 months, only 2 (13%) of the 15 patients had fat necrosis within the TRAM flap that was not present before radiotherapy. Fourteen patients (93%) retained their flap, and 13 patients (87%) rated their cosmetic outcome as "good" to "excellent." We conclude that TRAM flaps can be irradiated with few complications and acceptable cosmetic results. PMID- 15289735 TI - Thymidine labeling index: prognostic role in breast cancer. AB - The aim of this study is to evaluate the prognostic role of thymidine labeling index in patients with breast cancer. Cellular proliferation rates in 155 breast cancer specimens were investigated by 3H-thymidine labeling index (3H-TLI). Median age was 47 years (range: 23-76). At presentation, 11 patients (7.1%) had stage I disease, 76 (49%) had stage II, 64 (41.3%) had stage III disease, and 4 (2.6%) had metastatic involvement. Patients were placed in 2 groups based on their proliferative indices. The cut-off level was assigned as the median TLI value of the whole group. Correlations between proliferative activity of the tumors based on 3H-TLI levels and various previously established prognostic factors, as well as the influence of proliferative activity on survival as a clinical outcome, were analyzed. The mean and median TLI values for the whole group of patients were 4.36 +/- 4.96% and 2.76% (range: 0-23.6), respectively. There was a significant association of nuclear grade with TLI (P = 0.04). Patients who were alive with no sign of disease at the final follow-up examination had a significantly lower median TLI rate than those who were either alive with disease or those who had eventually died with disease progression (3.7% versus 1.9%, respectively; P = 0.04). Patients with locally advanced disease (N2 + N3 involvement) had a significantly higher median TLI rate than those with local nodal involvement (N1) (3.4% versus 1.7%, respectively, P = 0.026). Furthermore, TLI levels showed a significant association with overall survival in patients with node-negative disease (P = 0.02). Based on the results of this study, it can be concluded that TLI plays a significant prognostic role in a subset of patients with node-negative breast cancer. Furthermore, TLI appears to have a predictive value for the clinical outcome of patients with breast cancer. These findings may justify a more aggressive therapeutic approach in patients with high TLI levels. Further large-scale, prospective studies are required before a definite conclusion can be reached.reached. PMID- 15289734 TI - Dose escalation of docetaxel concomitant with hypofractionated, once weekly chest radiotherapy for non-small-cell lung cancer: a phase I study. AB - Hypofractionated chest radiotherapy has been used as an alternative when standard fractionated schedules are neither practical nor feasible. To explore docetaxel as radiosensitizer in a hypofractionated chest irradiation schedule, a docetaxel dose escalation study was conducted in which 26 patients with advanced non-small cell lung cancer (NSCLC) (stages III and IV) were enrolled. Docetaxel was administered 24 hours prior to irradiation (starting dose 10mg/m2; escalating in 5mg/m2 increments). Radiation was administered at 500 cGy (one fraction) once/wk for 10 consecutive weeks (5000 cGy total). The docetaxel dose was escalated up to 45 mg/m2/wk. The treatment was well tolerated over 10 consecutive weeks without requiring dose reductions or interruptions. Toxicities were mainly docetaxel related. One of 19 evaluable patients had a complete radiographic response within the radiation treatment port, 13 had a partial response, and 5 had stable disease. No patient recurred within the radiation field. Three patients who underwent surgical resection following treatment were pathologically down staged to stage I. This trial of a small group of patients supports, in selected patients, synchronous administration of effective hypofractionated, radiosensitized radiation therapy and optimized systemic chemotherapy. PMID- 15289736 TI - Evaluation of the internal mammary lymph nodes by FDG-PET in locally advanced breast cancer (LABC). AB - The presence of internal mammary (IM) lymph node metastases in breast cancer predicts outcome and may alter treatment. Standard imaging has limited usefulness for evaluation of the IM chain because of low sensitivity. Our preliminary studies suggested that [F-18]-2-fluoro-d-glucose positron emission tomography (FDG-PET) improves the detection of IM and mediastinal metastases. We therefore performed a retrospective review of women who underwent FDG-PET prior to treatment to determine the benefit of PET for imaging IM disease. The records of 28 consecutive patients undergoing FDG-PET prior to neoadjuvant chemotherapy for suspected locally advanced breast cancer (LABC) were reviewed. The presence of abnormal IM uptake on FDG-PET was noted. IM uptake on FDG PET was compared with standard radiographic imaging and was correlated with putative risk factors for IM involvement and with clinical patterns of failure. Patients did not undergo IM biopsy; however, patterns of failure were assessed to validate the FDG-PET findings. Clearly abnormal FDG uptake in the IM nodes was seen in 7 of 28 women (25%). Prospective conventional chest imaging failed to identify IM metastases in any patient. IM uptake on PET was associated with large size of the primary tumor (P = 0.03) and with inflammatory disease (P = 0.04). The presence of IM FDG uptake predicted failure by a pattern consistent with spread from IM lymph node metastasis. FDG-PET appears to be a useful noninvasive modality to detect IM metastases in LABC. Pathologic verification in a prospective study is necessary to confirm these findings. PMID- 15289737 TI - Nonocular second primary tumors after retinoblastoma: retrospective study of 111 patients treated by electron beam radiotherapy with or without TEM. AB - The objective of this study was to assess the role of various clinical and treatment factors involved in the long-term incidence of nonocular second primary tumors following retinoblastoma. The study was based on 111 patients treated between 1963 and 1977 according to the same radiotherapy protocol (electron beam radiotherapy) alone or in combination with triethylene melamine (TEM). Various statistical methods were used to obtain the actuarial survival curve, the cumulative incidence of second primary tumors, and comparisons of patient groups and subgroups. The 5-, 10-, 20-, and 30-year survival rates were 75%, 70%, 63%, and 55% with a follow-up of 23 to 35 years. The study reports the various parameters concerning 111 children and 17 second primary tumors: sex, age at treatment, histology of the retinoblastoma and second primary tumors, site of second tumors (anatomic and compared with irradiation field), radiation dose, time to onset, and chemotherapy with or without TEM. The results are discussed and compared with the data reported in the literature. Electron beam radiotherapy at a dose of 45 Gy does not eliminate the risk of nonocular second primary tumors. TEM also does not modify survival or the overall incidence of second primary tumors, but significantly increases the risk of second primary tumors outside the irradiation field. PMID- 15289738 TI - Survival in relation to radiotherapeutic modality for brain metastasis: whole brain irradiation vs. gamma knife radiosurgery. AB - The purpose of this report is to evaluate and compare the survival of patients with brain metastasis (BRM) treated by whole brain irradiation (WBI) using linear energy accelerator (LINAC) and by stereotactic radiosurgery using gamma knife. This study consists of a series of 67 patients with BRM treated with WBI between 1998 and 1999 and 53 patients with BRM treated with gamma knife radiosurgery (GKRS) between 2000 and 2001. A retrospective study of the data was performed and the overall survival between these 2 groups was analyzed. The comparability of these 2 groups was tested by chi2 and t test values. Log-rank test was used in the survival comparison. The 1-year survival rate was 26.3% and 22.6%, and corresponding mean survival was 7.8 months and 6.7 months for WBI and GKRS groups, respectively. There was no statistically significant difference between these 2 groups' survival. It was evident from imaging defined lesions that with GKRS the lesions were reduced, stabilized, or disappeared in 89% of cases. Survival of patients with BRM treated with WBI or GKRS was similar in these series. The present study suggests that good tumor response by GKRS does not translate in longer patient survival. PMID- 15289739 TI - Prognostic impact of survivin, cyclin D1, integrin beta1, and VEGF in patients with small adenocarcinoma of stage I lung cancer. AB - The purpose of this study was to investigate the impact of survivin, cyclin D1, integrin beta1, and vascular endothelial growth factor (VEGF) in tumor on survival of patients with small adenocarcinoma of the lung. Seventy-two patients with pathologic stage I resected tumors <2 cm in diameter were entered into the study. Each patient underwent curative surgical resection for lung cancer between July 1992 and November 1999. The resected tumors were subjected to immunostaining for each gene. Thirty-five, 26, 6, and 16 patients had tumors with >10% survivin , >20% cyclin D1-, >10% integrin beta1-, and >10% VEGF-positive cells, respectively. When the survival of 72 patients was compared according to each gene expression, the overall survival of patients with positive expression of survivin, cyclin D1, and integrin [beta]1 was significantly worse than that of individuals whose tumors had negative expression of each gene. By multivariate analysis controlling for each gene expression, no gene expression was an independent marker of poor prognosis, however, the overall survival of the complex gene expression (2 or more gene-positive) group (n = 35) was significantly worse than that of 0 or 1 gene-positive group (n = 37; log-rank test, P = 0.0011; Wilcoxon test, P = 0.0011). When the association between survival and pathologic factors, including lymphatic invasion, venous invasion, type of bronchioalveolar carcinoma, and complex gene positive expression was analyzed, only complex gene-positive expression was found to be a significant independent factor (hazard ratio = 0.085, P = 0.0299). It can be concluded that multiple increased expression of oncogene is a poor prognostic factor in patients with small adenocarcinoma of the lung. PMID- 15289740 TI - Gemcitabine combined with epirubicin in the treatment of patients with locally advanced or metastatic breast cancer: a phase II study. AB - This phase II study of gemcitabine and epirubicin evaluated the activity and toxicity in advanced breast cancer. Female patients with stage IIIB or IV breast cancer received gemcitabine 1000 mg/m2 and then epirubicin 15 mg/m2 on days 1, 8, and 15 of 28-day cycles. Thirty-five patients with stage IV disease, a median age of 59 years (range, 39-73), and a median Karnofsky performance status of 90 (range, 60-100) were enrolled. Fourteen (40.0%) patients received prior chemotherapy (12 adjuvant, 4 metastatic, 2 both). Of 35 evaluable patients, 10 had PR, for an overall RR of 28.6%, and 12 (34.3%) patients had SD. Median time to progression and overall survival were 5.8 months (95% CI, 3.4-9.5 months) and 17.1 months (95% CI, 11.2-19.9 months), respectively. WHO grade 3/4 neutropenia occurred in 51.5% of patients without febrile neutropenia, and grade 3 thrombocytopenia in 29.4% of patients without hemorrhage or platelet transfusions. The most common nonhematologic toxicities were grade 3 alopecia (38.2%) and nausea/vomiting (11.4%). There were no grade 4 nonhematologic toxicities. Gemcitabine plus epirubicin is active and well tolerated in patients with metastatic breast cancer. Future studies should continue to evaluate the impact of various schedules on outcome. PMID- 15289741 TI - Shielding of the contralateral breast during tangential irradiation. AB - The purpose of this study was to investigate both optimal and practical contralateral breast shielding during tangential irradiation in young patients. A shaped sheet of variable thickness of lead was tested on a phantom with rubber breasts, and an optimized shield was created. Testing on 18 consecutive patients 50 years or younger showed shielding consistently reduced contralateral breast dose to at least half, with small additional reduction after removal of the medial wedge. For younger patients in whom radiation exposure is of considerable concern, a simple shield of 2 mm lead thickness proved practical and effective. PMID- 15289743 TI - Do NSAIDs interfere with the cardioprotective effects of aspirin? PMID- 15289742 TI - Identification of myocardial ischemia in the diabetic patient. Joint ALFEDIAM and SFC recommendations. PMID- 15289744 TI - Methamphetamine abuse. PMID- 15289745 TI - Trospium chloride (Sanctura): another anticholinergic for overactive bladder. PMID- 15289746 TI - Obstructive sleep apnea syndromes. PMID- 15289747 TI - Classification and definition of respiratory disorders during sleep. AB - Growing interest in sleep disorders has led to increased research in this direction. Increasingly sophisticated instrumental tests have disclosed new breathing patterns and complex syndromes. The initial identification of obstructive apneas was followed by studies characterizing snoring, hypopneas, respiratory effort-related arousal and flow limitation events. Since Pickwickian syndrome, an historical term currently deemed obsolete and confusing, sleep investigations have differentiated secondary hypoventilation, central hypoventilation and syndromes resulting from narrowing of the upper airways (snoring, upper airway resistance syndrome, obstructive sleep apnea-hypopnea syndrome). Inevitably, this has given rise to some confusion in the classification of events and syndromes which recent studies have attempted to clarify. PMID- 15289748 TI - Epidemiology of obstructive sleep apnea syndrome. AB - The association of obstructive sleep apnea (OSA) syndrome with obesity, hypertension and cardiovascular disease has highlighted the broad public health importance of this condition. OSA affects at least 9% to 15% of middle-aged adults. Both epidemiological and sleep clinic-based studies indicate that OSA is more common in men than in women. However, the ratio of men to women with OSA in clinical studies appears to be considerably higher than in the community: up to 8:1 versus 2 to 3:1. Cross sectional studies on OSA prevalence showed effects of age, independently from the unfortunate propensity for a rising body mass index (BMI) with age: an approximate doubling of apnea-hypopnea index (AHI) every 10 years has been reported. A recent prospective study with 4-year follow-up showed that a 10% weight gain predicted a 32% increase in AHI, whereas a 10% loss in weight predicted a 26% decrease in AHI. Another 5-year prospective study found that longitudinal change in AHI varies nonuniformly with age, sex and weight: older heavier may experience the highest rate of AHI increase over time and, thus, may benefit most from prospective monitoring. PMID- 15289749 TI - The genetics of sleep disorders. AB - Most sleep disorders result from complex interactions between genes and the environment. Modern molecular techniques are increasingly applied to determine the contribution of genes to sleep and its disorders. The genetic basis of circadian rhythms has been explored using Drosophila and rodent models. This culminated in the identification of the molecular basis of one autosomal dominant form of familial advanced sleep phase syndrome: mutations in the human period 2 gene. Genetic studies in an autosomal recessive canine model of narcolepsy and in gene-targeted mice have identified the hypothalamic hypocretin (orexin) neuropeptide system as a key target for human narcolepsy. In this case, animal models have provided important clues to a human disorder with complex genetics. The study of the role of genes in the obstructive sleep apnoea syndrome is likely to provide important clues to a phenotype associated with respiratory, cardiovascular, and metabolic dysfunction. This brief review will present the role of genetic factors in the obstructive sleep apnoea syndrome, restless leg syndrome, narcolepsy, and circadian rhythm disorders. PMID- 15289750 TI - Diagnosis of sleep apnea. AB - Obstructive sleep apnea syndrome (OSAS) is characterized by repeated oropharyngeal occlusions occurring during sleep. The prevalence of moderate OSAS (with an apnea-hypopnea index = or >15/h) is 9% and 4% in male and female, respectively. It is associated with an abnormally high frequency of cardiovascular disease (hypertension, stroke, coronary heart disease) and excessive daytime sleepiness responsible for an increased frequency of work and road accidents. Because the treatment of OSAS provides many benefits to patients and society, it is very important to obtain an early diagnosis. The diagnosis of OSAS is based on the combination of characteristic clinical features plus compatible findings on instrumental tests in which multiple physiologic signals are monitored simultaneously during a night of sleep. A full night polysomnography, conducted by a technologist in a sleep laboratory, is the gold standard for the diagnosis of suspected OSAS, but the capacity for performing polysomnography is limited. On the basis of the high incidence and prevalence of OSAS, of the limited number of sleep laboratories, long waiting times and high costs recommendations have been formulated for the use of unattended portable systems in the assessment of OSAS. The main clinical aspects of of OSAS, the diagnostic approach with full night polysomnography and unattended portable systems, the differential diagnosis and some examples of cardiorespiratory portable monitoring are presented in this paper. PMID- 15289751 TI - [Preclinical and clinical results with the epidermal growth factor receptor inhibitor Gefitinib (ZD1839, Iressa)]. AB - Since epidermal growth factor receptor (EGFR) is involved in tumor proliferation and angiogenesis, and in the mechanisms of resistance to chemo- and hormono therapy, it represents a unique promising target for anticancer treatment. Gefinitib (Iressa, ZD1839), an inhibitor of the EGFR tyrosine kinase activity able to bind the intracellular domain of the receptor, is at present in clinical development. In preclinical models Gefitinib induced a dose-dependent response rate in tumor xenografts obtained from different human cancer cells lines. The expression of EGFR in the prior tumor did not appear to be a predictive marker for Gefitinib sensitivity. Furthermore, long-term drug use was well tolerated in mice without inducing resistance. However, tumors started to grow again after treatment interruption. Laboratory findings and in vivo data have prompted the evaluation of Gefitinib administered as a single oral daily dose alone or in combination with conventional anticancer treatment. PMID- 15289752 TI - [Celiac disease and its endocrine and nutritional implications on male reproduction]. AB - The problem of the interference of celiac disease (CD) with the male reproductive system is made evident both by the recognized adverse effects on female reproduction and by the multifactorial nature of the disease. It is important to consider CD as a multifactorial condition since its diverse effects can be modulated, besides gluten, by different concurrent genetic and environmental factors. The male CD patient has a greater risk of infertility and other reproductive disturbances, as well as a greater incidence of hypoandrogenism. In this paper the problems of CD associated to endocrine disorders and to deficiencies of micronutrients are discussed. Affected males show a picture of tissue resistance to androgens. Moreover, attention should be paid to increases of FSH and prolactin; these are not associated to infertility and/or impotence, but they may indicate an imbalance at hypothalamus-pituitary level, with general effects on health: an example is the increased risk of male osteoporosis in CD patients. Hormone alterations are reversible upon start of the gluten-free diet, emphasizing the importance of early diagnosis; this should be performed in the case of clinical suspicion, e.g., unexplained hypoandrogenism. As regards nutritional aspects, the folic acid deficiency of CD can affect rapidly proliferating tissues, such as the embryo and the seminiferous epithelium. More attention should be paid to deficiencies of fat-soluble vitamins, such as A and E, observed in CD. Vitamin A is important for Sertoli cell function as well as for early spermatogenetic phases. Vitamin E supports the correct differentiation and function of epidydimal epithelium, spermatid maturation and secretion of proteins by the prostate. Therefore, CD male patients should be considered as vulnerable subjects; thus, the detection of early biomarkers of andrological or endocrinological dysfunctions should trigger timely strategies for prevention and treatment. PMID- 15289753 TI - Hepatitis C virus and hepatic abnormalities in Bulgarian patients with primary Sjogren's syndrome. PMID- 15289754 TI - Generational evolution and the future of pediatrics. PMID- 15289755 TI - Protecting our children from environmental hazards in the face of limited data-a precautionary approach is needed. PMID- 15289756 TI - Short bowel and long life: no longer mutually exclusive. PMID- 15289757 TI - Symptomatic cerebral edema in diabetic ketoacidosis: the mechanism is clarified but still far from clear. PMID- 15289758 TI - Treatment of acute wheezing episodes in young children. PMID- 15289759 TI - Scientific advances provide opportunities to improve pediatric environmental health. PMID- 15289760 TI - Long-term parenteral nutritional support and intestinal adaptation in children with short bowel syndrome: a 25-year experience. AB - OBJECTIVE: To analyze the outcome of children with short bowel syndrome (SBS) who required long-term parenteral nutrition (PN). STUDY DESIGN: Retrospective analysis of children (n=78) with SBS who required PN >3 months from 1975 to 2000. STATISTICS: univariate analysis, Kaplan-Meier method, and Cox proportional regression model were used. RESULTS: We identified 78 patients. Survival was better with small bowel length (SBL) >38 cm, intact ileocecal valve (ICV), intact colon, takedown surgery after ostomy (all P <.01), and primary anastomosis (P <.001). PN-associated early persistent cholestatic jaundice (P <.001) and SBL of <15 cm (P <.01) were associated with a higher mortality. Intestinal adaptation was less likely if SBL <15 cm (P <.05), ICV was removed, colonic resection was done (both P <.001), >50% of colon was resected (P <.05), and primary anastomosis could not be accomplished (P <.01). Survival was 73% (57), and 77% (44) of survivors had intestinal adaptation. CONCLUSIONS: SBL, intact ICV, intestinal continuity, and preservation of the colon are important factors for survival and adaptation. Adaptation usually occurred within the first 3 years. Need for long term PN does not preclude achieving productive adulthood. Patients with ICV even with <15 cm of SBL and patients with SBL >15 cm without ICV have a chance of intestinal adaptation. PMID- 15289761 TI - Mechanism of cerebral edema in children with diabetic ketoacidosis. AB - OBJECTIVES: Cerebral edema during diabetic ketoacidosis (DKA) has been attributed to osmotic cellular swelling during treatment. We evaluated cerebral water distribution and cerebral perfusion during DKA treatment in children. STUDY DESIGN: We imaged 14 children during DKA treatment and after recovery, using both diffusion and perfusion weighted magnetic resonance imaging (MRI). We assessed the apparent diffusion coefficients (ADCs) and measures reflecting cerebral perfusion. RESULTS: The ADC was significantly elevated during DKA treatment (indicating increased water diffusion) in all regions except the occipital gray matter. Mean reductions in the ADC from initial to postrecovery MRI were: basal ganglia 4.7 +/- 2.5 x 10(-5) mm(2)/s (P=.002), thalamus 3.7 +/- 2.8 x 10(-5) mm(2)/s, (P=.002), periaqueductal gray matter 4.3 +/- 5.1 x 10(-5) mm(2)/s (P=.03), and frontal white matter 2.0 +/- 3.1 x 10(-5) mm(2)/s (P=.03). In contrast, the ADC in the occipital gray matter increased significantly from the initial to postrecovery MRI (mean increase 3.9 +/- 3.9 x 10(-5) mm(2)/s, P=.004). Perfusion MRI during DKA treatment revealed significantly shorter mean transit times (MTTs) and higher peak tracer concentrations, possibly indicating increased cerebral blood flow (CBF). CONCLUSIONS: Elevated ADC values during DKA treatment suggests a vasogenic process as the predominant mechanism of edema formation rather than osmotic cellular swelling. PMID- 15289762 TI - beta-agonists through metered-dose inhaler with valved holding chamber versus nebulizer for acute exacerbation of wheezing or asthma in children under 5 years of age: a systematic review with meta-analysis. AB - OBJECTIVE: To compare the efficacy of beta-agonists given by metered-dose inhaler with a valved holding chamber (MDI+VHC) or nebulizer in children under 5 years of age with acute exacerbations of wheezing or asthma in the emergency department setting. STUDY DESIGN: Published (1966 to 2003) randomized, prospective, controlled trials were retrieved through several different databases. The primary outcome measure was hospital admission. RESULTS: Six trials (n=491) met criteria for inclusion. Patients who received beta-agonists by MDI+VHC showed a significant decrease in the admission rate compared with those by nebulizer (OR, 0.42; 95% CI, 0.24-0.72; P=.002); this decrease was even more significant among children with moderate to severe exacerbations (OR, 0.27; 95% CI, 0.13-0.54; P=.0003). Finally, measure of severity (eg, clinical score) significantly improved in the group who received beta-agonists by MDI+VHC in comparison to those who received nebulizer treatment (standardized mean difference, -0.44; 95% CI, -0.68 to -0.20; P=.0003). CONCLUSIONS: The use of an MDI+VHC was more effective in terms of decreasing hospitalization and improving clinical score than the use of a nebulizer in the delivery of beta-agonists to children under 5 years of age with moderate to severe acute exacerbations of wheezing or asthma. PMID- 15289763 TI - Near-miss apparent SIDS from adrenal crisis. AB - OBJECTIVE: Adrenal crisis from salt-losing congenital adrenal hyperplasia (CAH) typically occurs in the first 2 weeks of life. We evaluated 3 infants with adrenal crisis who presented at 6 to 8 months of age with near-miss sudden infant death syndrome (SIDS). SUBJECTS: Three 46,XY phenotypic female infants presented near death at 6 to 8 months of age with adrenal crisis and unmeasurable steroid hormones consistent with congenital lipoid adrenal hyperplasia (lipoid CAH). METHODS: We sequenced genes potentially causing this phenotype: steroidogenic acute regulatory protein (StAR), the cholesterol side-chain cleavage enzyme, adrenodoxin reductase, adrenodoxin, and steroidogenic factor 1 (SF1). Site directed mutagenesis and functional assays were performed for the missense mutation. RESULTS: Hormonal values showed complete absence of adrenal and gonadal steroids. Patient 1 was a compound heterozygote for missense mutation R140P and an mRNA splice donor site mutation in the StAR gene. The R140P mutation was wholly inactive in vitro. Patient 2 was homozygous for a 7 base pair StAR deletion causing a frameshift. No mutations were found in Patient 3, suggesting a novel disease. CONCLUSIONS: Although genetic disorders of steroidogenesis typically present in the first month of life, some defects, especially those in StAR, can present in mid-infancy, when adrenal hyperplasias are rarely considered. Adrenal insufficiency is a subtle disorder that may cause cardiovascular collapse, causing unexplained infant death that resembles SIDS. PMID- 15289764 TI - Intracranial hemorrhage in premature neonates treated with extracorporeal membrane oxygenation correlates with conceptional age. AB - OBJECTIVE: To determine the effect of patient age on the risk of intracranial hemorrhage (ICH) in premature neonates treated with extracorporeal membrane oxygenation (ECMO). STUDY DESIGN: This was a retrospective cohort study of neonates of <37 weeks' gestation treated with ECMO in the years 1992 through 2000 and reported to the Extracorporeal Life Support Organization Registry (n=1524). The relation between ICH and patient age, defined as gestational age, postnatal age (PNA), and postconceptional age (PCA), was determined with the use of multiple logistic regression analysis. RESULTS: PNA was inversely correlated with ICH in the univariate analysis (P=.01) but not in the multivariate analysis (P=.36). PCA showed a strong univariate correlation with decreasing ICH: 26% of patients A, with high predictivity for impaired function of cytochrome P450 2D6 in white subjects. AB - BACKGROUND: Individuals with the cytochrome P450 (CYP) 2D6 intermediate metabolizer (IM) phenotype have low residual enzyme activity and compose about 10% to 15% of white populations. Their identification is clinically relevant but remains unsatisfactory because of incomplete characterization of the major allele involved, termed CYP2D6*41 (-1584C, R296C, S486T). METHODS: To search for novel mutations, resequencing of the entire CYP2D6 gene was performed in selected individuals. Genotype-phenotype correlation analysis was done in a population sample of 308 white subjects phenotyped with sparteine and previously genotyped for all major alleles. RESULTS: A total of 16 novel polymorphic positions were identified, of which 7 were located within 2.4 kilobases of previously uncharacterized 2D7-2D6 intergenic sequence and 9 were located within intronic regions. The novel mutation 2988G>A in intron 6 appeared to be specifically associated with the IM phenotype. Further analysis in the population sample demonstrated that 2988G>A was strongly linked to allele *41 but not to any other alleles including *1, *2, *2xN, *4, *6, *7, *8, *9, *10, and *35. The overall frequency of the novel polymorphism was 8.4% in the normal white population. Compared with conventionally determined *41, 2988G>A was shown to have improved predictivity for the IM phenotype. With 2988G>A being taken into account, alleles *1, *2, and *35 (-1584G, V11M, R296C, S486T) were found to be phenotypically equivalent. CONCLUSIONS: CYP2D6 genotyping can be considerably simplified by using 2988G>A as a marker for *41 and by omitting genotyping for the functionally equivalent alleles *2 and *35. PMID- 15289791 TI - Human sympathetic activation by alpha2-adrenergic blockade with yohimbine: Bimodal, epistatic influence of cytochrome P450-mediated drug metabolism. AB - BACKGROUND: alpha2-Adrenergic blockade responses suggest adrenergic dysfunction in hypertension. alpha2-Blockade is also used to treat autonomic dysfunction. However, pharmacokinetic determinants of yohimbine disposition are not understood. METHODS: We evaluated alpha2-blockade with intravenous yohimbine in 172 individuals. Specific cytochrome P450 (CYP) isoform-mediated metabolism was investigated. Results were evaluated by ANOVA and by maximum likelihood analysis for bimodality of response distributions. RESULTS: Yohimbine metabolism to 11 hydroxy-yohimbine displayed greater than 1000-fold variability, with 17 individuals showing no metabolism. Nonmetabolizers differed from others in ethnicity but not in age, sex, body habitus, blood pressure, heart rate, or family history of hypertension. Bimodality of metabolism was suggested by frequency histogram, as well as maximum likelihood and cluster analysis. Among ethnic groups, subjects of European ancestry had the highest frequency of nonmetabolism. In vitro oxidation suggested that the major route of metabolism (lowest Michaelis-Menten constant and greatest intrinsic clearance) was likely via CYP2D6 to 11-hydroxy-yohimbine. In vivo genotypes at both CYP2D6 and CYP3A4 were necessary to predict metabolism (overall F = 3.03, P =.005); an interaction of alleles at these 2 loci (interaction F = 3.05, P =.033) suggested an epistatic effect on drug metabolism in vivo. Nonmetabolizers had greater activation of sympathetic nervous system activity. Yohimbine increased blood pressure, an effect mediated hemodynamically by elevation of cardiac output rather than systemic vascular resistance. Blood pressure and cardiac output responses did not differ by metabolizer group. CONCLUSIONS: We conclude that heterogeneous, bimodally distributed yohimbine metabolism depends on common genetic variation in both CYP2D6 and CYP3A4 and contributes to differences in sympathetic neuronal response to alpha2-blockade. These results have implications for both diagnostic and therapeutic uses of this alpha2-antagonist. PMID- 15289792 TI - Ketoconazole, a cytochrome P450 3A4 inhibitor, markedly increases concentrations of levo-acetyl-alpha-methadol in opioid-naive individuals. AB - BACKGROUND: Levo-acetyl-alpha-methadol (LAAM) exerts most of it mu-agonist activity through the action of its 2 N-demethylation metabolites, norLAAM and dinorLAAM. The N-demethylation of LAAM to norLAAM and norLAAM to dinorLAAM is primarily performed by cytochrome P450s (CYP) in the 3A family. No previous studies have been conducted to determine the effect of in vivo inhibition of CYP3A on the pharmacokinetics and pharmacodynamics of LAAM. METHODS: Oral LAAM (5 mg/70 kg) was administered on 2 occasions in a single-blind, randomized crossover design to 13 opioid-naive subjects (6 women and 7 men) 1 hour after pretreatment with 400 mg ketoconazole or placebo. Blood and urine samples were collected at defined intervals over 240- and 96-hour periods, respectively; LAAM, norLAAM, and dinorLAAM concentrations were determined by liquid chromatography-tandem mass spectrometry. Physiologic and subjective measures were collected for up to 72 hours. RESULTS: Results are presented as the geometric mean with 90% confidence intervals of individual ratios of ketoconazole to placebo sessions. Coadministration of ketoconazole and LAAM resulted in a 3.22-fold (2.53-4.10, P <.001) and 5.29-fold (4.24-6.61, P <.001) increase in the maximum plasma concentration (Cmax) and area under the curve (AUC) of LAAM. The values for time to Cmax (tmax) of norLAAM and dinorLAAM were increased 2.43-fold (1.92-3.08, P <.001) and 11.6-fold (8.36-16.1, P <.001), with 0.77-fold (0.67-0.87, P <.005) and 0.55-fold (0.49-0.60, P <.001) decreases in their respective Cmax values. The AUCs of norLAAM and dinorLAAM were increased 2.25-fold (1.96-2.58, P <.001) and 1.21-fold (1.12-1.32, P <.005), respectively. Pupil diameter was significantly decreased by LAAM after both placebo and ketoconazole pretreatment; ketoconazole increased the tmax for miosis 2.92-fold (2.01-4.25, P <.001). Other physiologic measures and numerous subjective effects measures were significantly affected by LAAM; however, few significant effects of ketoconazole pretreatment were observed on these outcomes. CONCLUSION: A single dose of ketoconazole causes a significant pharmacokinetic drug interaction with a single dose of LAAM that results in increased LAAM concentrations relative to norLAAM and dinorLAAM at early time points. Coadministration also results in prolongation of the appearance of its active metabolites and a concomitant prolongation of miosis, a sensitive dynamic index of mu-opioid action. The clinically relevant increase in LAAM concentrations and prolongation of plasma LAAM metabolites may affect physiologic function, such as QT intervals, suggesting that coadministration of LAAM and CYP3A4 inhibitors should be contraindicated. PMID- 15289793 TI - Rosuvastatin pharmacokinetics in heart transplant recipients administered an antirejection regimen including cyclosporine. AB - BACKGROUND: Cyclosporine (INN, ciclosporin) increases the systemic exposure of all statins. Therefore rosuvastatin pharmacokinetic parameters were assessed in an open-label trial involving stable heart transplant recipients (> or =6 months after transplant) on an antirejection regimen including cyclosporine. Rosuvastatin has been shown to be a substrate for the human liver transporter organic anion transporting polypeptide C (OATP-C). Inhibition of this transporter could increase plasma concentrations of rosuvastatin. Therefore the effect of cyclosporine on rosuvastatin uptake by cells expressing OATP-C was also examined. METHODS: Ten subjects were assessed while taking 10 mg rosuvastatin for 10 days; 5 of these were then assessed while taking 20 mg rosuvastatin for 10 days. Rosuvastatin steady-state area under the plasma concentration-time curve from time 0 to 24 hours [AUC(0-24)] and maximum observed plasma concentration (Cmax) were compared with values in controls (historical data from 21 healthy volunteers taking 10 mg rosuvastatin). Rosuvastatin uptake by OATP-C-transfected Xenopus oocytes was also studied by use of radiolabeled rosuvastatin with and without cyclosporine. RESULTS: In transplant recipients taking 10 mg rosuvastatin, geometric mean values and percent coefficient of variation for steady-state AUC(0 24) and Cmax were 284 ng. h/mL (31.3%) and 48.7 ng/mL (47.2%), respectively. In controls, these values were 40.1 ng. h/mL (39.4%) and 4.58 ng/mL (46.9%), respectively. Compared with control values, AUC(0-24) and Cmax were increased 7.1 fold and 10.6-fold, respectively, in transplant recipients. In transplant recipients taking 20 mg rosuvastatin, these parameters increased less than dose proportionally. Rosuvastatin had no effect on cyclosporine blood concentrations. The in vitro results demonstrate that rosuvastatin is a good substrate for OATP-C mediated hepatic uptake (association constant, 8.5 +/- 1.1 micromol/L) and that cyclosporine is an effective inhibitor of this process (50% inhibition constant, 2.2 +/- 0.4 micromol/L when the rosuvastatin concentration was 5 micromol/L). CONCLUSIONS: Rosuvastatin exposure was significantly increased in transplant recipients on an antirejection regimen including cyclosporine. Cyclosporine inhibition of OATP-C-mediated rosuvastatin hepatic uptake may be the mechanism of the drug-drug interaction. Coadministration of rosuvastatin with cyclosporine needs to be undertaken with caution. PMID- 15289794 TI - Time response of cytochrome P450 1A2 activity on cessation of heavy smoking. AB - BACKGROUND AND OBJECTIVE: Cytochrome P450 (CYP) 1A2 activity is induced by cigarette smoking. Thus smoking cessation in patients while they are undergoing therapy with a CYP1A2 substrate such as theophylline or clozapine increases its concentrations and may cause adverse effects. Our objective was to determine the time course of CYP1A2 activity changes after smoking cessation in heavy smokers as the basis for dosing adaptation schemes. METHODS: The study was conducted in 8 men and 4 women (all white) who smoked 20 cigarettes or more per day. Sudden smoking cessation was carried out after a 14-day run-in period. Subjects were phenotyped for CYP1A2 activity at 6, 4, and 1 day before smoking cessation and at 0, 1, 2, 3, 6, 8, 10, and 13 days thereafter by use of the paraxanthine-to caffeine ratio in plasma 6 hours after a 148-mg caffeine test dose. A monoexponential decay of CYP1A2 activity to a residual value was fitted to the data by nonlinear regression analysis. RESULTS: On cessation of smoking, initial caffeine clearance (estimated geometric means and 95% confidence intervals) decreased significantly (P <.01), by 36.1% (30.9%-42.2%), from 2.47 mL. min(-1). kg(-1) body weight (2.03-3.00 mL. min(-1). kg(-1) body weight) to a new steady state of 1.53 mL. min(-1). kg(-1) body weight (1.24-1.89 mL. min(-1). kg(-1) body weight). The apparent half-life of CYP1A2 activity decrease was 38.6 hours (27.4 54.4 hours). CONCLUSION: Doses of CYP1A2 substrates with a narrow therapeutic range should be decreased immediately on cessation of heavy smoking. As a rule of thumb, a stepwise daily dose reduction of approximately 10% until the fourth day after smoking cessation is proposed, which should be accompanied by therapeutic drug monitoring. PMID- 15289795 TI - Caution with beta1-adrenergic receptor genotyping. PMID- 15289796 TI - Recovery of hippocampal cell proliferation and BDNF levels, both of which are reduced by repeated restraint stress, is accelerated by chronic venlafaxine. AB - The present study investigated the poststress (PS) cellular and molecular changes in the hippocampus of rats subjected to repeated restraint stress (RS) and the effects of chronic administration of an antidepressant drug, venlafaxine, on these changes. It was found that RS suppressed hippocampal cell proliferation, decreased brain-derived neurotrophic factor (BDNF) levels, and increased both the levels of copper/zinc superoxide dismutase (Cu/Zn-SOD) and the number of Cu/Zn SOD immunostained hippocampal interneurons. In venlafaxine-treated rats, the changes in cell proliferation, BDNF levels, and the number of Cu/Zn-SOD interneurons returned to control levels on PS Days 21, 14, 7, respectively. In vehicle-injected rats, BDNF and the number of Cu/Zn-SOD interneurons returned to control levels on PS Days 21 and 14, respectively, but cell proliferation was still suppressed on PS Day 21. The stress-induced elevation of Cu/Zn-SOD protein remained during the 3-week PS period, and it was further increased by about 20% after 3 weeks of venlafaxine administration. PMID- 15289797 TI - Differential qualitative responses to rivastigmine in APOE epsilon 4 carriers and noncarriers. AB - This retrospective analysis of two double-blind, placebo-controlled studies in patients with mild to moderately severe AD investigated the efficacy of rivastigmine 6-12 mg/day on cognitive outcomes in patients with or without the apolipoprotein (APOE) epsilon4 allele. APOE data were collected from patients who consented to pharmacogenetic testing. Treatment differences within each subgroup were compared, using the Observed Case (OC) population. The APOE epsilon4 and non APOE epsilon4 subgroups comprised 246 and 121 patients, respectively. Overall, APOE epsilon4 noncarriers showed greater decline than carriers (P<0.05). However, at 26 weeks, placebo-treated APOE epsilon4 patients declined 3.04 points below baseline on the cognitive subscale of the Alzheimer's Disease Assessment Scale (ADAS-cog), and rivastigmine-treated patients improved by 1.67 points. Non-APOE epsilon4 placebo-treated patients declined by 4.59 points and rivastigmine treated patients declined by 0.48 points. Thus, non-APOE epsilon4 carriers showed a less favorable course under either placebo or rivastigmine, but both genotype defined subgroups showed quantitatively similar responses to therapy (both P<0.05 vs placebo). PMID- 15289798 TI - Valproic acid inhibits histone deacetylase activity and suppresses excitotoxicity induced GAPDH nuclear accumulation and apoptotic death in neurons. AB - Valproic acid (VPA), used to treat bipolar mood disorder and seizures, also inhibits histone deacetylase (HDAC). Here, we found that VPA and other HDAC inhibitors, butyrate and trichostatin A, robustly protected mature cerebellar granule cell cultures from excitotoxicity induced by SYM 2081 ((2S, 4R)-4 methylglutamate), an inhibitor of excitatory amino-acid transporters and an agonist of low-affinity kainate receptors. These neuroprotective effects required protracted treatment and were correlated with enhanced acetylated histone levels, indicating HDAC inhibition. SYM-induced excitotoxicity was blocked by MK-801 ((5R,10S)-(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine hydrogen maleate), supporting that the toxicity was largely N-methyl-D-aspartate receptor dependent. SYM excitotoxicity had apoptotic characteristics and was prevented by a caspase inhibitor. SYM-induced apoptosis was associated with a rapid and robust nuclear accumulation of glyceraldehyde-3-phosphate dehydrogenase (GAPDH), a housekeeping gene previously shown to be proapoptotic. VPA pretreatment suppressed SYM 2081-induced GAPDH nuclear accumulation, concurrent with its neuroprotective effects. Chromatin immunoprecipitation (ChIP) revealed that GAPDH is copresent with acetylated histone H3, including Lys9-acetylated histone, and that VPA treatment caused a time-dependent decrease in the levels of nuclear GAPDH with a concomitant increase in acetylated histones in the ChIP complex. Our results strongly suggest that VPA protects neurons from excitotoxicity through inhibition of HDAC activity and that this protective effect may involve suppression of excitotoxicity-induced accumulation of GAPDH protein in the nucleus. PMID- 15289799 TI - Optimal exercise intensities for fat metabolism in handbike cycling and cycling. AB - STUDY DESIGN: Energy expenditure (EE) and fat oxidation in handbike cycling compared to cycling in order to determine the intensity that elicits maximal fat oxidation in handbike cycling. OBJECTIVE: To establish the exercise intensity with the highest fat oxidation rate in handbike cycling compared with cycling (control group) in order to give training recommendations for spinal cord-injured (SCI) athletes performing handbike cycling. SETTING: Institute of Sports Medicine, Swiss Paraplegic Centre, Nottwil, Switzerland. METHODS: Eight endurance trained handbike cyclists (VO2 peak(handbike cycling) 37.5+/-7.8 ml/kg/min) and eight endurance trained cyclists (VO2 peak(cycling) 62.5+/-4.5 ml/kg/min) performed three 20-min exercise blocks at 55, 65 and 75% VO2 peak in handbike cycling on a treadmill or in cycling on a cycling ergometer, respectively, in order to find the intensity with the absolutely highest fat oxidation. RESULTS: The contribution of fat to total EE was highest (39.1+/-16.3% EE) at 55% VO2 peak in handbike cycling compared to cycling, where highest contribution of fat to EE (50.8+/-13.8%) was found at 75% VO2 peak. In handbike cycling, the highest absolute fat oxidation (0.28+/-0.10 g/min) was found at 55% VO2 peak compared to cycling, where highest fat oxidation (0.67+/-0.20 g/min) was found at 75% VO2 peak. CONCLUSION: Well-trained handbike cyclists have their highest fat oxidation at 55% VO2 peak(handbike cycling) compared to well-trained cyclists at 75% VO2 peak(cycling). Handbike cyclists should perform endurance exercise training at 55% VO2 peak(handbike cycling), whereas well-trained cyclists should be able to exercise at 75% VO2 peak(cycling). For training recommendations, the heart rate at 55% VO2 peak(handbike cycling) lies at 135+/-6 bpm in handbike cycling in SCI compared to 147+/-14 bpm at 75% VO2 peak(cycling) in well-trained cyclists. We presume that the reduced muscle mass involved in exercise during handbike cycling is the most important factor for impaired fat oxidation compared to cycling. But also other factors as fitness level and haemodynamic differences should be considered. Our results are only applicable to well-trained handbike cyclists with SCI and not for the general SCI population. PMID- 15289800 TI - Time/duration effectiveness of sildenafil versus tadalafil in the treatment of erectile dysfunction in male spinal cord-injured patients. AB - STUDY DESIGN: A randomized, blinded, crossover clinical trial comparing sildenafil versus tadalafil for erectile dysfunction (ED) in male spinal cord injured (SCI) patients. OBJECTIVES: To compare the safety, time/duration effectiveness, and the impact on the quality of life (QoL) of tadalafil 10 mg versus sildenafil 50 mg. SETTING: Neurourology Section, Careggi Hospital, Florence, Italy. METHODS: During a screening (visit 1), a diary card was distributed, in which the subjects assessed, after each attempt at intercourse the quality of their erection, responding (Yes/No) to both Sexual Encounter Profile Questions 2 (SEP2) and 3 (SEP3). The subjects made at least four attempts at intercourse. At visit 2, 15 patients (group 1) were assigned sildenafil and 15 (group 2) started with tadalafil. Responses to baseline International Index of Erectile Function 5 items (IIEF-5), Questions 13-14 (IIEF 15 items) and SEP diary were recorded. Patients attempted intercourse on four separate occasions: within 4 h of taking the first tablet, within 12 h for the second tablet, 24 h for the third, and the fourth from 24 to 36 h. At visit 3, the investigators evaluated the effectiveness with the same measures used at baseline. After a wash-out period, at visit 4, Group 1 was given tadalafil, and Group 2 was given sildenafil. Patients were required to observe the same criteria in taking the four tablets as in visit 2. After 4 weeks (visit 5), we evaluated the patients as we did in visit 3. RESULTS: Overall, 28 patients completed the study. No subjects discontinued the drugs due to drawbacks. Tadalafil allowed a majority of men in this trial to achieve both normal sexual functioning up to 24 h postdosing compared to sildenafil (P<0.01) and improved overall sex life satisfaction as well as sexual relations with partner. CONCLUSION: Based on these data, tadalafil may have the potential to become an important treatment option for ED in SCI patients. SPONSORSHIP: This study was not sponsored. PMID- 15289801 TI - Neuropathic pain after traumatic spinal cord injury--relations to gender, spinal level, completeness, and age at the time of injury. AB - STUDY DESIGN: Retrospective register study. OBJECTIVE: To investigate the predictive value of age at the time of injury, gender, level of injury, and completeness of injury for the development of at level and below level neuropathic pain. SETTING: "Spinalis", a postacute spinal cord injury (SCI) outpatient clinic, serving the greater Stockholm area (Sweden). METHOD: All patients who visited the clinic in 1995-2000 (402 patients) for the first time were examined. The following items were selected: at-level and below-level neuropathic pain according to the International Association for the Study of Pain (IASP) criteria, age at the time of injury, gender, level of injury according to ASIA, and completeness of injury. Mean time of 6 years after the injury. Results were analysed with chi(2) analysis and logistic regression. RESULTS: Of all patients examined, 13% had at level pain and 27% had below level pain. Neuropathic pain was less than half as frequent (26%) in the group aged less than 20 years at the time of injury as in the oldest group (58%). The increasing trend was mainly due to below-level pain up to 39 years of age, and due to at-level pain at ages 40 and above at the time of injury. No correlation was observed to gender, level of injury or completeness of injury, except for below level pain, which was associated with complete injury. CONCLUSION: The results show that neuropathic pain after SCI is common and occurs much more often in patients injured at higher ages. This indicates the importance of neuroanalgetic intervention, in particular for patients injured in higher ages. PMID- 15289802 TI - Outcomes in patients admitted for rehabilitation with spinal neurological lesions following intervertebral disc herniation. AB - BACKGROUND: Little information is available about the survival, neurological recovery, and length of stay in hospital for rehabilitation (LOS) of patients with spinal neurological deficit following disc herniation (DH). STUDY DESIGN: Retrospective cohort study. OBJECTIVE: To report on outcomes and factors affecting these. SETTING: The Spinal Research Laboratory, Loewenstein Rehabilitation Hospital, Israel. SUBJECTS: A total of 158 patients with DH spinal neurological lesions (DHSNL). METHOD: Data were collected retrospectively. Survival was assessed using the Kaplan-Meier method; relative mortality risk by the Cox proportional hazard model. Neurological recovery was evaluated by calculating the change in Frankel grades, and factors that affect it were assessed by logistic regression. LOS associations were analyzed with ANOVA. RESULTS: The median age at lesion onset was 48 years, and the median survival 29 years. Age and gender had a significant effect on survival, but not so lesion severity, level, or decade of onset. Of the 69 patients who had Frankel grades A, B, or C on admission, 72% achieved useful recovery to grades D or E. The severity and level of the spinal neurological lesion (SNL) had a significant effect on recovery. The mean LOS was 87 days; it was significantly affected by lesion severity and level and by the decade of admission to rehabilitation, and decreased with time. CONCLUSIONS: Patients with DHSNL who were admitted for rehabilitation have favorable survival and recovery rates compared with previously studied patients with other types of SNL. Their LOS is probably a function of medical requirements, but is decreasing with time. PMID- 15289803 TI - Spinal chronic subdural hematoma in a patient with ventriculo-peritoneal shunting after a minor trauma. AB - STUDY DESIGN: Case report and review of literature. OBJECTIVES: Intracranial chronic subdural hematoma (SDH) is a well-recognized complication of ventriculoperitoneal (VP) shunt. Spinal chronic SDH is very rarely associated with VP shunt. SETTING: Kaohsiung Medical University Hospital, Kaohsiung, Taiwan. CASE REPORT: We describe a spinal chronic SDH, developing after a minor trauma, in a patient who underwent a VP shunt surgery for hydrocephalus 6 months previously. A good outcome was achieved after decompressive surgery. CONCLUSION: Spinal chronic SDH should be considered in the diagnosis of progressive spinal compression, especially in the patients with VP shunt after minor trauma. PMID- 15289804 TI - Transcutaneous functional electrical stimulation for grasping in subjects with cervical spinal cord injury. AB - STUDY DESIGN: Case series. OBJECTIVES: To evaluate the benefit, shortcomings and acceptance of a new transcutaneous functional electrical stimulation (FES) technology aimed at improving the grasp function in tetraplegic subjects in acute and postacute rehabilitation. SETTING: Spinal cord injury (SCI) centre, university hospital. METHODS: : Subjects (N=11) with complete or incomplete SCI at C4/5-C7 who started FES 1-67 months after their accident were included. Hand function tests, analysis of video recordings and of written documentation of FES sessions, status of muscle strength, and follow-up query were used as outcome measures. RESULTS: Nine subjects used FES as a neuroprosthesis. Eight demonstrated improved grasp function and performance in activities of daily living. In one subject, no benefit from FES was observed. Two other subjects showed improvements in muscle strength and facilitation of active movement with FES. Four subjects successfully integrated FES as neuroprosthesis in everyday life within the rehabilitation centre. Three received the system for home use. The most relevant reasons for stopping the FES application were: (i) improvement of voluntary grasp function, (ii) physical and psychological problems, (iii) no available stimulator for home use, and (iv) insufficient assistance for electrode placement at home. Shortcomings related to the transcutaneous surface technology (eg pain or coactivation of neighbouring muscles) could usually be reduced, or did not limit the efficiency or acceptance of FES. Individually designed digital or analogue control devices were preferred. CONCLUSION: Tetraplegic subjects in acute and postacute rehabilitation can profit from a new transcutaneous FES system with respect to functional use and independence. It can be implemented in the rehabilitation programme for muscle strengthening and facilitation of voluntary activity. For a successful application of FES, there is a need for individual electrode placement, stimulation programmes, and FES control devices. PMID- 15289805 TI - Intramedullary inflammatory mass dorsal to the Klippel-Feil deformity: error in development or response to an abnormal motion segment? AB - STUDY DESIGN: Case study on a 45-year-old female with progressive weakness and paresthesias in her lower extremities and a magnetic resonance image of an intramedullary mass at the level of C5-6 and a Klippel-Feil (KF) deformity ventral to the lesion. OBJECTIVE: Present an interesting case of an intramedullary mass coexisting with a ventral KF deformity with review of the medical literature on intramedullary masses and cervical spine biomechanics. SETTING: New York city, New York, USA. METHODS: Case study with discussion of neurosurgical and neuropathological findings and review of the literature. RESULTS: The patient underwent open neurosurgical excisional biopsy of the intramedullary mass which revealed a non-neoplastic inflammatory mass that slowly resolved with medical management. CONCLUSIONS: While no definitive etiology was found in this case we offer two interesting mechanisms: (1) maldevelopment of the cervical spine or (2) this inflammatory mass is in response to an abnormal motion at the level of the Klippel-Feil. PMID- 15289806 TI - Reused silicone catheter for clean intermittent catheterization (CIC): is it safe for spinal cord-injured (SCI) men? AB - STUDY DESIGN: Study of reusable catheter. OBJECTIVE: To investigate whether a silicone cathether reused over years for clean intermittent catheterization (CIC) was safe for spinal cord injured (SCI) men. SETTING: Maharaj Hospital, Chiang Mai, Thailand. METHOD: A cross-sectional study was obtained from SCI men who had used CIC with a reusable silicone catheter for more than a year. Demographic data, urological management and urinary tract complications focusing on the radiologic status of the urethra were reviewed and analyzed. In addition, two reused and one new catheters were studied under electron microscope for catheter morphology (surface and lumen) and stiffness. RESULTS: There were 28 SCI men included in this study. The average duration of CIC use was 4.8 years and the average time of usage for each catheter was 3 years (range 1-7 years). In all, 26 men previously used indwelling catheterization (ID) during the acute phase. In all, 23 men performed self-catheterization. Regarding urinary complications, three reported urethral bleeding, five had episodes of pus per urethra, five had epididymitis, four had passing stones, 18 had occasional foul smelly urine, 10 developed fever and cloudy urine during the past year. Of 17 patients who had ultrasonography done, four had pathologic findings in kidney and one had bladder calculi. Demographic data, urinary management and complications did not have significant relation to the abnormality of the urethrogram or urinary tract infection. However, where the frequency of CIC was higher, the abnormality of the urethra was lower (P<0.05). All had serum Cr level < or =1.3 mg/dl. Electron microscopic findings of reused catheters for 2 years revealed encrustation but no obstruction in the lumens and 20% increase in stiffness. CONCLUSION: The clinical outcome, especially with regard to urethral abnormalities with this reusable silicone catheter is as good as with a disposable one. However, to reuse urinary catheters, one should consider the increasing risk of infection. For SCI patients in developing countries, CIC with a reusable silicone catheter may be a suitable and safe choice if one cleans and applies it properly to reduce infection. In order to answer the question how long a person in a developing country should use the same silicone catheter, further research should be conducted. PMID- 15289807 TI - Mobility aids and transport possibilities 10-45 years after spinal cord injury. AB - STUDY DESIGN: A cross-sectional survey with retrospective data. OBJECTIVE: Follow up information on the use of mobility aids and transportation possibilities in a chronic traumatic spinal cord injury (SCI) population. SETTING: Clinic for Para- and Tetraplegia at Rigshospitalet, University hospital, Denmark (CPT). The uptake area is East Denmark with a population of 2.5 million inhabitants. METHODS: Survey on date of birth, gender, time of SCI, cause of SCI, neurological level and functional classification from medical files were combined with information concerning mobility aids and transport possibilities at the time of follow-up from a mailed questionnaire. MATERIAL: Individuals with traumatic SCI before 1 January 1991 were still in regular follow-up at CPT, and with sufficient medical record. A total of 279 were included, out of which 236 answered the questionnaire. Of the 193 men and 43 women injured from 1956 to 1990 the response rate was 84.6%. Age at the time of follow-up was 50.5 years in mean, and follow up time was 24.1 years in mean. In all, 126 were paraplegic and 110 tetraplegic. Responders and nonresponders were comparable. RESULTS: In all, 3.4% used no special mobility aids at all. In total, 49 used crutches or rolling walkers and 26 lower extremities bracing, but mostly in combination with a wheelchair. Standing frame and stand-up wheelchair were used by men only. Manual wheelchair was used by 83.5% and electrical wheelchair by 27%, and the latter more by the tetraplegics. In all, 9.3% had neither a manual nor an electrical wheelchair. Overall, 86.4% had a passenger van or another car. Women used a car less often. Passenger vans were more often used by tetraplegics. CONCLUSION: Nearly all SCI participants had mobility aids of some sort, and 90.7% had either a manual or an electrical wheelchair or both. Most had a passenger van or another type of car for transportation. These facilities are important for the individuals to obtain an independent living. PMID- 15289808 TI - Patient perception of the impact of sporting activity on rehabilitation in a spinal cord injuries unit. AB - OBJECTIVE: To establish patients' perception of the effect of sport on rehabilitation. DESIGN: Single investigator telephone questionnaire survey. SETTING: Spinal Cord Injuries Unit, Musgrave Park Hospital, Belfast, Northern Ireland, UK. METHOD: A telephone questionnaire was administered to patients admitted over a 12-month period. Information on patient demographics, sports participation before and after injury and patients' perception of the impact of sports on rehabilitation was recorded. RESULTS: In all, 33 (84.6%) of the 39 patients could be contacted and all completed the questionnaire. Of the respondents 27 (81.8%) had a diagnosis of spinal cord injury. Six (18.2%) were female and 20 (60.6%) were below 45 years.A total of 15 (45.5%) patients previously participated in regular sporting activity. At least 24 (72.7%) tried one sport during admission. In all, 14 reported taking part in regular sporting activity after discharge, although 23 expressed a desire to do so. Of those 60% who had regularly exercised continued to do so and 27% of those who reported no regular sport, before injury, commenced regular active exercise. A general benefit of sporting activity was recognised by 78.8% and a rehabilitation benefit by 69.7%. Self-reported benefits included increase in fitness, quality of life, confidence and social contact. CONCLUSION: A high proportion of patients perceived sporting activity as beneficial. Patients exercising before injury were more likely to exercise after injury. All inpatients were introduced to sport by staff, emphasising the importance of this rehabilitation opportunity in planning in-patient care. PMID- 15289809 TI - Reflex reciprocal facilitation of antagonist muscles in spinal cord injury. AB - STUDY DESIGN: Electromyographic study in complete and incomplete spinal cord injury (SCI). OBJECTIVE: To examine the changes in the pattern of reciprocal inhibition between agonist and antagonist muscles in SCI. SETTINGS: Sensory Motor Performance Program, Rehabilitation Institute of Chicago, IL, USA. METHODS: Tendon taps were delivered manually with an instrumented hammer to the tendons of the tibialis anterior and soleus muscle in positions of full-ankle dorsiflexion and plantarflexion in eight subjects with complete SCI and eight subjects with incomplete SCI. Electromyographic activity (EMG) was recorded from ankle dorsiflexor and plantarflexor muscles. Tapping force was also recorded by a force sensor mounted to the tendon hammer, indicating the stimulus onset. Measures of reflex EMG magnitude and reflex latency were obtained for both agonist and antagonist muscles. The ratio of antagonist to agonist EMG was computed based on normalized EMG. RESULTS: Substantial reflex responses occurred in both the stretched muscle and in its antagonist. The reflex in antagonist, which we term 'reciprocal facilitation (RF)', was most evident in subjects with incomplete SCI. The magnitude of RF was consistently greater than reflex responses in agonist muscles under all test conditions. The latency of the RF was comparable to that of monosynaptic reflex response. CONCLUSIONS: Following SCI, reciprocal organization of segmental reflexes at the ankle is often partially or completely suppressed, allowing reflex activation in antagonist muscles to be manifested. Possible mechanisms underlying these changes in neural organization are discussed. SPONSORSHIP: This study was supported by Spinal Cord Research Foundation, the Paralyzed Veterans of America. PMID- 15289810 TI - Prostate cancer detection and tumor characteristics in men with multiple biopsy sessions. AB - PURPOSES: To address prostate cancer (PCa) detection with respect to the number of biopsy sessions performed, to identify risk factors for detection after a negative biopsy, and to analyze the clinical characteristics of the detected tumors. SCOPE: Only biopsied men (sextant) were included. A total of 1011 biopsy sessions were carried out in 770 men; 172 underwent a second prostate biopsy and 51 a third biopsy. During the first biopsy round, 111 cancers were found (14.4%), 27 in the second (15.7%), and five during the third round (9.8%), P=0.156. Only high-grade PIN or atypia were identified as independent predictors or PCa detection in subsequent biopsies (P=0.008). A nonsignificant increase of clinically localized tumors, and a decrease of metastatic and poorly differentiated cases were found when more biopsy sessions were needed for detection. CONCLUSIONS: A nonsignificant trend to lower cancer detection rates and less clinical relevance of the tumors detected can be observed when more biopsy rounds are needed for detection. PMID- 15289811 TI - The 100-day PSA: usefulness as surrogate end point for biochemical disease-free survival after definitive radiotherapy of prostate cancer. AB - Overall and biochemical disease-free (bNED) survival data after definitive radiotherapy (RT) for prostate cancer (CaP) requires decades of patient follow up. Surrogates involving dynamics of prostate-specific antigen (PSA) decline, PSA nadir and time thereto have been unrewarding. This study investigated the metric of the PSA value 100 days after RT (PSA(100)), analyzed with respect to 8-y bNED survival. A total of 214 patients with T1-3 CaP were treated with definitive RT (defined as dose >66 Gy) in our institution between 1/1/1988 and 12/31/2000. All were subject to continuous follow-up with routine PSA levels. Biochemical failure (77 patients) was defined by the ASTRO criteria (n=67) or by the date of first hormonal therapy for a rising PSA, which did not meet the ASTRO criteria (n=10). No patients were included if they received postoperative radiation, or if hormones were administered prior to bNED recurrence, if any. Patients were stratified by PSA(100) values 4.0 ng/ml, and 4.0 ng/ml had 20% 8-y bNED survival (P<0.001). Use of a PSA(100) cutoff of 2.5 ng/ml yielded no significant difference in 8-y bNED survival (P=0.229). Cox proportional analysis revealed that initial PSA (P=0.006), stage (P=0.001) and PSA(100)4 cm). Changes in the AFP values, and both CT and ultrasonography (US) findings before and after treatment were investigated. All 24 tumors received 1 or more treatments (average, 3.3 treat-ments) of PHoT. The injection volume ranged from 3-26 ml (average, 11.2 ml). The total volume of the injection per tumor ranged from 10-37.2 ml (average, 37.2 ml). The AFP values decreased in all patients who initially showed high values. On CT scanning, all lesions receiving PHoT became hypodense. The disappearance of the tumor was also confirmed by contrast-enhanced CT. No severe complications, excluding mild abdominal pain and skin burning, were observed during the procedure. In conclusion, PHoT shows good anti-tumor effects despite a small number of punctures and holds promise as a curative local treatment method for small HCCs. PMID- 15289840 TI - Comparison of effects of As2O3 and As4O6 on cell growth inhibition and gene expression profiles by cDNA microarray analysis in SiHa cells. AB - An arsenical compound, As2O3 has been reported to be effective for treating acute leukemia and induce apoptosis in many different tumor cell types. In this study we designed a novel arsenical compound, As4O6, and compared its ability to induce cell growth inhibition as well as gene expression profiles along with As2O3 in HPV16 infected SiHa cervical cancer cells. Both As2O3 and As4O6 induced apoptosis in SiHa cells, as determined by a DNA ladder formation. As4O6 was more effective in suppressing the growth of SiHa cells in vitro, as compared to As2O3. To further compare gene expression profiles between these two drugs, we used a 384 cDNA microarray system. The gene expression profiles were also classified into the Gene Ontology (GO) to investigate apoptosis-related cellular processes. In the case of As2O3, 41 genes were up- or down-regulated at least 2-fold, as compared to non-treatment, whereas, 65 genes were up- or down-regulated by As4O6 treatment. In particular, 27 genes were commonly regulated by both arsenic compounds. The GO analysis also indicated that down-regulation of cell-regulatory functions, such as cell cycle, protein kinase activity and DNA repair, induces an anti-tumor effect. Taken together, these data support that As4O6 could be more effective than As2O3 in inhibiting the growth of HPV16 infected cervical cancer cells. This appears to be mediated through a unique but overlapping regulatory mechanism(s), suggesting that the regulated genes and cellular processes could be used for a new potential drug approach for treating cervical cancer in clinical settings. PMID- 15289841 TI - Prevention of irinotecan-induced diarrhoea by oral carbonaceous adsorbent (Kremezin) in cancer patients. AB - Irinotecan (CPT-11) treatments induce severe diarrhoea at a rate of >40%. In clinical trials, we evaluated the preventing effects of oral alkalization, which has been reported previously, and oral carbonaceous adsorbent (Kremezin trade mark ) on diarrhoea possibly induced by CPT-11. Evaluation was made by counting the maximum number of bowel motions in each patient. Five patients out of 7 treated with CPT-11 had bowel motions of >5 times daily, and maximum number of bowel motions reached 20 times in 1 patient. Oral alkalization (2 g sodium bicarbonate, 2 g magnesium oxide and 300 mg ursodeoxycholic acid daily for 4 days) decreased bowel motions from 20 to 8 thereafter in the patient. Maximum number of bowel motions in other 3 patients treated in a combination of CPT-11 and oral alkalization was <3. Oral adsorbent (2 g Kremezin x 3 times, during and after CPT-11 treatment) also decreased maximum number of bowel motions from 7 (without oral adsorbent) to 3 in 1 patient. Also, the maximum number of bowel motions in other 3 patients treated with oral adsorbent was <3 (p<0.05, vs CPT-11 alone). Effect of oral Kremezin on plasma concentrations of CPT-11 and its related compounds after a 1-h CPT-11 infusion, evaluated in a patient, was small. These results suggested that the oral Kremezin is effective in ameliorating CPT 11-induced diarrhoea without decreasing much the plasma clearance of CPT-11. PMID- 15289842 TI - Relationship between HPV typing and the status of G2 cell cycle regulators in cervical neoplasia. AB - We examined human papillomavirus (HPV) typing and the status of ATM, chk2, CDC25C, cdc2 and cyclinB1 in cervical intraepithelial neoplasia (CIN) and invasive cancer (IC). A total of 93 samples [normal: 10; CIN: 34 (CINI:9, CINII:12, CINIII:13); IC: 49 (stage I:10, stage II:21, stage III:15, stage IV:3)] were included in this study. HPV status was evaluated by the PCR non-radioactive HPV detection system. We analyzed ATM, chk2, CDC25C, cdc2 and cyclinB1 protein expression by immunohistochemistry. HPV DNA was detected in 73.5% of 34 CINs and 89.8% of 49 ICs. Detection of HPV subtypes 16 and 18 was more frequent in ICs (46.9%) than in CINs (23.5%) (p=0.0387). Abnormal expression of ATM, chk2, CDC25C, cdc2 and cyclinB1 were 2.9%, 32.4%, 2.9% 20.6% and 0% in CINs and 8.2%, 30.6%, 10.2%, 46.9% and 12.2% in ICs. The alteration of cdc2 was higher in ICs than in CINs (p=0.0198). Altered expression of cdc2 was higher in HPV16 and 18 cases (69.6%) than in other cases (26.9%) (p=0.0042). However, the relationship between HPV typing and ATM, chk2, CDC25C and cyclinB1 expression was not significant. Cdc2 is implicated in cervical carcinogenesis and may be related to p53 inactivation by HPV. PMID- 15289843 TI - Amplification/overexpression of Aurora-A in human gastric carcinoma: potential role in differentiated type gastric carcinogenesis. AB - Aurora-A encodes a cell cycle regulated serine/threonine kinase that has essential functions for centrosome maturation and chromosome segregation. Aurora A is amplified and overexpressed in various human carcinomas and is suggested to be a potential oncogene. To clarify the potential role of Aurora-A in human gastric carcinoma, we examined the amplification and expression in both tumor cell lines and primary carcinoma. We examined the amplification and overexpression of Aurora-A in 9 gastric carcinoma cell lines and 88 primary gastric carcinomas using Southern and Northern blot analysis, and confirmed a protein expression by immunohistochemical staining. We also investigated the relationship between Aurora-A overexpression and clinicopathological features of the tumors. Aurora-A amplification and overexpression was observed in 29% and 44.4% of cell lines and 12.5% and 41% of primary carcinomas, respectively. There was discordance between gene amplification and transcript expression, since in a previous study DNA amplification was the main mechanism for Aurora-A activation. Aurora-A overexpression exhibited significant association with increasing age and differentiated type gastric carcinoma. It was also detected in early stage gastric cancer as well as in gastric intestinal metaplasia, which is considered as a common precursor lesion for the differentiated type gastric carcinoma, and severe dysplastic cells showed stronger protein expression. We concluded that Aurora-A overexpression may well be involved in differentiated type gastric carcinogenesis. Further evaluation of the possible roles of Aurora-A and the regulation of Aurora-A expression in malignant cells will be critically important for the development of new strategies aimed at controlling the growth of malignant cells. PMID- 15289844 TI - Prostate-related antigen-derived new peptides having the capacity of inducing prostate cancer-reactive CTLs in HLA-A2+ prostate cancer patients. AB - Prostate-related antigens, including prostate-specific membrane antigen (PSMA) and prostatic acid phosphatase (PAP), can be targets in specific immunotherapy for prostate cancer. In this study, we attempted to newly identify epitope peptides from these 2 antigens, which are immunogenic in human histocompatibility leukocyte antigen (HLA)-A2+ prostate cancer patients. Twenty-nine peptides (PSMA with 15 and PAP with 14) were prepared based on the HLA-A2 binding motif. Based on our previous finding that antigenic peptides recognized by both cellular and humoral immune systems are useful for peptide-based immunotherapy, peptide candidates were screened first by their ability to be recognized by immunoglobulin G (IgG), and then by their ability to induce peptide-specific cytotoxic T lymphocytes (CTLs). As a result, PSMA441-450 and PAP112-120 peptides were found to be frequently recognized by IgG in plasma from prostate cancer patients. These 2 candidates effectively induced HLA-A2-restricted and prostate cancer-reactive CTLs in HLA-A2+ prostate cancer patients with several HLA-A2 subtypes. In addition, their cytotoxicity was mainly dependent on peptide specific and CD8+ T cells. These results indicate that these PSMA441-450 and PAP112-120 peptides could be promising candidates for peptide-based immunotherapy for HLA-A2(+) prostate cancer. PMID- 15289845 TI - Histologic tumor growth pattern is significantly associated with disease-related survival in muscle-invasive transitional cell carcinoma of the urinary bladder. AB - Tumor stage and lymph node status are currently the most established prognostic factors in muscle-invasive bladder cancer. However, there is still a need for new parameters allowing to assess the prognosis more accurately. A recently introduced morphologic classification of tumor growth, which distinguishes three growth patterns (nodular, trabecular, and infiltrative type), was retrospectively applied to a collective of 153 muscle-invasive bladder carcinomas in order to examine its prognostic relevance. Nodular pattern was found in 34 cases (22%), trabecular pattern in 97 cases (63%) and infiltrative pattern in 22 cases (14%) as the dominant type of tumor growth. The majority of cases (54%) displayed more than one tumor growth pattern. A two-tiered classification considering the worst type of tumor growth (non-infiltrative or infiltrative type) yielded 74 non infiltrative tumors (55%) and 69 infiltrative tumors (45%). The former group was associated with a significantly better disease-related survival than the latter (p=0.007 by univariate Cox regression analysis). Moreover, the two-tiered classification was identified, besides tumor stage (p=0.036) and lymph node status (p=0.0001), as an independent predictor of disease-related survival in a multivariate analysis (p=0.029). In conclusion, our data suggest that the morphologic pattern of tumor growth is significantly associated with the clinical course of advanced transitional cell carcinomas of the bladder. The tumor growth classification proposed here may serve as an additional tool to predict the prognosis of patients undergoing radical cystectomy for bladder cancer. PMID- 15289846 TI - Cell growth after withdrawal of gefitinib ("Iressa", ZD1839) in human lung cancer cells. AB - Gefitinib ("Iressa", ZD1839) is a selective epidermal growth factor receptor tyrosine kinase inhibitor, which blocks signal transduction pathways implicated in proliferation and survival of cancer cells. However, in vitro and in vivo studies concerning cell growth after withdrawal of gefitinib are limited. To determine whether cancer cells would resume proliferation upon removal of gefitinib, an in vitro study was undertaken using 4 human non-small cell lung cancer cell lines. With immunocytochemical staining and Western blot analysis, we confirmed positive expression of EGFR in these cell lines. We next evaluated growth inhibition before and after withdrawal of gefitinib. After incubation with 0-100 microM gefitinib for 24 h, medium containing gefitinib was removed, followed by addition of fresh growth medium to cell cultures. MTT assays were performed daily over a 6-day time course. Even at very low levels of gefitinib (<1 microM), these 4 TKB cells had 1-10% growth inhibition, respectively. With these levels of gefitinib, continued inhibitory effects after withdrawal of getitinib were observed in 3 of the 4 cell lines. Furthermore, none of the 4 cell lines, including the cell line which had abolished growth inhibition, showed accelerated re-growth rate after the withdrawal of gefitinib even in these exposure conditions. Based on our in vitro experiments, additional in vivo studies, which can compare the pre- and post-treatment growth rate, will be necessary to help better understand the mechanisms behind cell growth recovery after temporary gefitinib treatment. PMID- 15289847 TI - Immunohistochemistry and microsatellite instability testing for selecting MLH1, MSH2 and MSH6 mutation carriers in hereditary non-polyposis colorectal cancer. AB - Hereditary non-polyposis colorectal cancer (HNPCC) represents 1-3% of all colorectal cancers. HNPCC is caused by a constitutional defect in a mismatch repair (MMR) gene, most commonly affecting the genes MLH1, MSH2 and MSH6. The MMR defect results in an increased cancer risk, with the greatest lifetime risk for colorectal cancer and other cancers associated to HNPCC. The HNPCC-associated tumor phenotype is generally characterized by microsatellite instability (MSI) and immunohistochemical loss of expression of the affected MMR protein. The aim of this study was to determine the sensitivity of IHC for MLH1, MSH2 and MSH6, and MSI analysis in tumors from known MMR gene mutation carriers. Fifty-eight paired normal and tumor samples from HNPCC families enrolled in our high-risk colorectal cancer registry were studied for the presence of germline mutations in MLH1, MSH2 and MSH6 by DGGE and direct sequencing. MSI analysis and immunostaining for MLH1, MSH2 and MSH6 were evaluated. Of the 28 patients with a real pathogenic mutation, loss of immunohistochemical expression for at least 1 of these MMR proteins was found, and all except 1 have MSI-H. Sensitivity by MSI analysis was 96%. IHC analysis had a sensitivity of 100% in detecting MMR deficiency in carriers of a pathogenic MMR mutation, and can be used to predict which gene is expected to harbor the mutation for MLH1, MSH2 and MSH6. This study suggests that both analyses are useful for selecting high-risk patients because most MLH1, MSH2 and MSH6 gene carriers will be detected by this 2-step approach. This practical method should have immediate application in the clinical work of patients with inherited colorectal cancer syndromes. PMID- 15289848 TI - Methylation of multiple genes in prostate cancer and the relationship with clinicopathological features of disease. AB - Promoter methylation plays an important role in the inactivation of tumor suppressor genes during tumorigenesis. We examined the methylation status of glutathione s-transferase Pi1 (GSTP1), retinoic acid receptor beta (RARB), CD44, E-cadherin (ECAD), RAS association domain family protein 1A (RASSF1A) and endothelin B receptor (EDNRB) genes in 81 prostate cancer and 42 benign prostatic hyperpasia specimens. Genomic DNA was isolated from archived formaldehyde-fixed and paraffin-embedded tissue blocks. Methylation-specific PCR (MSP) was carried out after bisulfite treatment of genomic DNA. Methylation frequencies in prostate cancer and benign prostatic hyperplasia were 72% and 5% for GSTP1, 40% and 0% for RARB, 72% and 38% for CD44, 61% and 14% for ECAD, 49% and 19% for RASSF1A and 72% and 62% for EDNRB, respectively. Methylation of GSTP1, RARB, CD44, ECAD and RASSF1A, but not of EDNRB was detected at a statistically higher frequency in prostate cancer than in the benign prostatic hypertrophy specimens. Methylation of RARB occurred more frequently in early onset (age <55 years) as compared to late onset disease (age >70 years) (odds ratio, 8.6; 95% CI, 1.4-51.4; P=0.02). Methylation of RARB also occurred more frequently in stage III as compared to stage II disease (odds ratio, 3.2; 95% CI, 1.1-8.8; P=0.03). A methylation index (MI) was calculated as the total number of genes methylated, excluding EDNRB. A trend toward higher MI was noted in stage III as compared to stage II disease, and in Gleason score 7 as compared to Gleason score 6 tumors. Our results suggest that the methylation of selected genes in prostate cancers correlates with clinicopathological features of poor prognosis. PMID- 15289849 TI - Foscan-based photodynamic treatment in vivo: correlation between efficacy and Foscan accumulation in tumor, plasma and leukocytes. AB - The tumoricidal effect of Foscan-mediated photodynamic therapy may involve both vessel and tumor cell destruction. The relevant importance of each mechanism seems to be defined by the time interval between photosensitizer administration and illumination (drug-light interval, DLI). Short drug-light intervals favor vascular damage due to the preferential photosensitizer accumulation in the tumor vasculature, whereas long drug-light intervals trigger direct tumor cell damage due to the dye localization in the tumor. The purpose of this study was to investigate the influence of tumor, plasma and leukocyte concentrations of Foscan at different times after photosensitizer delivery on PDT response. Both pharmacokinetic and tumor-response studies were carried out in nude mice bearing s.c. Colo26 tumors. One to 96 h after i.v. injection of 0.5 mg/kg Foscan, animals were exposed to 10 J/cm(2) 652-nm light delivered at 30 mW/cm(2). Mean tumor regrowth time was determined for each schedule of treatment and correlated to Foscan distribution in the compartments of interest at the time of illumination. PDT efficacy was greatest for irradiations performed at 6 and 12 h post Foscan injection and limited at 96 h. Unlike tumor and plasma Foscan concentrations, photosensitizer accumulation in leukocytes exhibited a good correlation with PDT efficacy. The results suggest that leukocytes could play an important role in the mechanism of PDT-induced vascular damage either by being one of the main effector compartments or by better reflecting Foscan accumulation in endothelial cells compared to plasma. The prevalence of indirect damage was highlighted by the fact that PDT efficacy was not modified by the use of a higher fluence rate of irradiation (160 mW/cm(2)), which depleted intratumor oxygen and did not restrain PDT-induced cell toxicity. PMID- 15289850 TI - Is survival following post-operative radiotherapy in pN1-non-small cell lung cancer reduced? Results of a multivariate analysis. AB - Prognostic factors and the role of post-operative radiotherapy (PORT) in patients with pN1 nodal stage following surgery for NSCLC were identified. The clinical course of 211 patients with pN1 nodal involvement following thoracic surgery were reviewed, 97 of them received PORT. Multi-variate survival analysis with respect to prognostic factors (including treatment) was performed. The most frequent site of recurrence was the ipsilateral bronchus-stump or hilus (63% of recurrences). The 5-year rate of intercurrent deaths for PORT was 1% vs 6% in the group without PORT. The 5-year rate of locoregional recurrence was similar (24% vs 19%) for PORT vs no PORT (p=0.97). PORT patients had a higher rate of distant metastases (p=0.04). The 5-year rate of overall survival was 45% without PORT and 25% with PORT (p=0.003). Multivariate survival analysis identified 4 prognostic factors associated with decreased survival rate: age, extended pneumectomy, number of involved nodes and PORT dose. A PORT dose of 50 Gy corresponds to an increase in relative risk of death in the range of 1.5. Patients with PORT do not have an increased rate of intercurrent deaths. However in this cohort, PORT in pN1 patients was associated with a decreased survival rate due to distant metastases. Even after correction with respect to accepted prognostic factors in multivariate survival analysis, PORT was not able to improve or equalize prognosis of these negatively selected patients. The main site of recurrence is the bronchial stump and hilus. If PORT is applied in pN1 patients, a reduction of the target volume should be discussed since local control in high-risk patients may be of relevance. PMID- 15289851 TI - Correlation of pre-operative coexistence of organ dysfunction with overall survival of patients with squamous cell carcinoma of the thoracic esophagus. AB - Coexistence of organ dysfunction has been regarded as a risk factor in the surgical treatment of digestive diseases. The aim of this study was to investigate the correlation between coexistence of organ dysfunction and post operative outcome in patients with thoracic esophageal squamous cell carcinoma (SCC). One hundred and fifty-eight patients with SCC of the thoracic esophagus, who had been surgically treated were examined, including 37 cases associated with organ dysfunction. The proportion of post-operative complication in patients with organ dysfunction (32.4%, 12/37) was significantly higher than that in patients without organ dysfunction (15.7%, 19/121; p=0.025). The overall survival of patients with organ dysfunction was significantly more unfavorable than that of patients without organ dysfunction (p=0.031), and multivariate analysis demonstrated that coexistence of organ dysfunction (p=0.004), depth of tumors (p=0.012), lymph node metastasis (p=0.0003), blood vessel invasion (p=0.008) and the incidence of post-operative complications (p=0.026) were factors independently associated with worse overall survival of the patients with SCC of thoracic esophagus. Coexistence of organ dysfunction was found to be an independent indicator of worse overall survival in patients with SCC of the thoracic esophagus. PMID- 15289852 TI - Ganoderma lucidum spore extract inhibits endothelial and breast cancer cells in vitro. AB - This study was conducted to investigate the anti-proliferative activities of medicinal mushroom Ganoderma lucidum (Rei-shi or Mannentake). We have identified an alcohol extract from the spore of Ganoderma lucidum that inhibits the in vitro proliferation of human umbilical vein endothelial cells and MDA-MB231 human breast cancer cells. Further fractionation of the alcohol extract revealed that the ethyl acetate fraction inhibited both cell lines in a dose-dependent manner from 2 to 40 micro g/ml. Our results suggest that the alcohol extract from the spore of Ganoderma lucidum may possess potential anti-tumor and anti-angiogenic activities. PMID- 15289853 TI - Deletion and aberrant CpG island methylation of Caspase 8 gene in medulloblastoma. AB - Aberrant methylation of promoter CpG islands in human genes is an alternative genetic inactivation mechanism that contributes to the development of human tumors. Nevertheless, few studies have analyzed methylation in medulloblastomas. We determined the frequency of aberrant CpG island methylation for Caspase 8 (CASP8) in a group of 24 medulloblastomas arising in 8 adult and 16 pediatric patients. Complete methylation of CASP8 was found in 15 tumors (62%) and one case displayed hemimethylation. Three samples amplified neither of the two primer sets for methylated or unmethylated alleles, suggesting that genomic deletion occurred in the 5' flanking region of CASP8. Our findings suggest that methylation commonly contributes to CASP8 silencing in medulloblastomas and that homozygous deletion or severe sequence changes involving the promoter region may be another mechanism leading to CASP8 inactivation in this neoplasm. PMID- 15289854 TI - The human myotubularin-related protein suppresses the growth of lung carcinoma cells. AB - Herein, we report that the human myotubularin-related protein (KIAA0371) induced growth suppression of lung cancer cells. Colony formation assays demonstrated that the exogenous expression of the human myotubularin-related gene induced a significant growth inhibition of H1299 cells in which the expression of the human myotubularin-related gene was undetectable. Colony formation of these cells was reduced by 39% after transfection with this gene, as compared with the control vector. Cells stably transfected with this gene showed a reduction in growth of 48% compared with those cells transfected with the empty vector. We also showed that this negative regulation of the myotubularin-related protein is correlated with cell-cycle arrest at G1. In addition, we found that the cyclin-dependent kinase inhibitor, p27, is induced by increased expression of the human myotubularin-related gene. These results suggest that the human myotubularin related protein can negatively regulate the growth of lung cancer cells. PMID- 15289855 TI - Effects of potent VEGF blockade on experimental Wilms tumor and its persisting vasculature. AB - We characterized the effect of potent vascular endothelial growth factor (VEGF) blockade on early-stage Wilms tumor xenograft growth, vasculature and metastasis. VEGF is a key mediator of both physiologic and tumor angiogenesis. We recently described that potent VEGF blockade induces regression of established Wilms tumor xenografts and vessels, also reducing the size but not the incidence of pre existing metastases. In these studies, we examined the effects of potent VEGF blockade on earlier stages of experimental Wilms tumors, focusing on tumor growth, vasculature and metastasis. Athymic mice received intrarenal human Wilms tumor cell implants. Biweekly treatment with vehicle or the VEGF-Trap, a high affinity soluble decoy receptor incorporating regions of VEGFR1 and VEGFR2, was begun 1 week later (100 or 500 micrograms/dose, n=20 in each group). Mice were euthanized at week 6 to examine tumor weight, incidence of lung metastases, vascularity and expression of angiogenic factors. A cohort of mice was examined 2 weeks after cessation of treatment. Compared to controls, VEGF-Trap treated tumors were significantly smaller (100 micrograms/dose: 92.7% smaller, p=0.0017; 500 micro g/dose: 99.0% smaller, p=0.0009). The incidence of lung metastasis also decreased significantly (p<0.0055). VEGF-Trap nearly eradicated tumor vasculature. Rare persisting vessels were characterized by large caliber, quiescence (lacking proliferation/apoptosis) and arterialization (both phenotypic and molecular). Potent VEGF blockade caused near-arrest of experimental Wilms tumor growth, resulted in nearly avascular tumors, and also decreased the incidence and size of metastases. Persistent vessels in tumors treated with VEGF Trap displayed specific morphologic and molecular features, suggestive of arterialization. Future strategies that target these persisting vessels may enhance the efficacy of VEGF blockade therapy. PMID- 15289856 TI - Diosgenin induces cell cycle arrest and apoptosis in HEL cells with increase in intracellular calcium level, activation of cPLA2 and COX-2 overexpression. AB - Many natural components of plant extracts are studied for their beneficial effects for health and particularly on carcinogenesis chemoprevention. In the present study, we investigated the effects of diosgenin on erythroleukemia HEL cells. Our results demonstrated that diosgenin induced G2/M arrest of cell cycle progression through p21 up-regulation in a p53-independent pathway and strong induction of apoptosis in HEL cells. Apoptosis induction was accompanied by an increase in Bax/Bcl-2 ratio, PARP cleavage and DNA fragmentation. Moreover, we showed for the first time that diosgenin provoked a collapse of mitochondrial membrane potential with an increase in intracellular calcium levels. It is well known that [Ca2+]i increase is one of the major activators of cytosolic PLA2. In our study, we demonstrated that diosgenin treatment induced cPLA2 activation through translocation to the cellular membrane. Moreover, arachidonic acid metabolism activation led to cyclooxygenase-2 (COX-2) but not lipoxygenase overexpression. Surprisingly, we observed a COX-2 up-regulation associated with apoptosis induction by diosgenin. These findings suggest that diosgenin has a potential chemopreventive effect; future studies should evaluate the mechanism of COX-2 activation during diosgenin-induced apoptosis in cancer cell lines. PMID- 15289857 TI - Growth suppression of esophageal squamous cell carcinoma induced by heavy carbon ion beams combined with p53 gene transfer. AB - Heavy carbon-ion beam therapy has revealed several potential advantages over X rays. Heavy-ion therapy has been applied for various solid tumors including esophageal squamous cell carcinoma (SCC). Although the local control rate in carbon ion radiotherapy for esophageal cancer has revealed better rates than conventional radiotherapy, severe mucosal damage was observed in adjacent normal mucosa. A suitable treatment strategy is required to reduce irradiation dose by introducing combined local therapy. Recently, we initiated clinical p53 gene therapy for esophageal SCC. We herein evaluate the cytotoxic effects of heavy carbon-ion beams combined with p53 gene transfer on human esophageal SCC. We assessed the induction of apoptosis and growth suppression with the use of recombinant adenoviral vector Ad.p53 or heavy carbon-ion beam irradiation or both. Growth suppression was significantly potentiated by combined treatment with heavy carbon-ion beams and Ad.p53 as compared to that treated with either of them alone. Western blot analysis confirmed the expression of both exogenous p53 and p21 proteins after irradiation of Ad.p53 infected cells. Enhanced apoptotic cell death was observed with a terminal deoxynucleotidyl transferase-mediated nick end labeling assay. These data suggest that heavy carbon-ion beam irradiation combined with Ad.p53 may be a potentially effective therapeutic strategy for locally advanced esophageal SCC. PMID- 15289858 TI - A novel combination antimetabolite, TAS-102, exhibits antitumor activity in FU resistant human cancer cells through a mechanism involving FTD incorporation in DNA. AB - TAS-102 is a new antimetabolite agent composed of a alpha, alpha, alpha trifluorothymidine (FTD; 1 M) and thymidine phosphorylase inhibitor (TPI; 0.5 M). Here, we investigated the antitumor effect and mechanism of TAS-102 against 5-FU, or FdUrd, resistant human cancer cell lines. The respective tumor growth inhibition rate of orally administered FTD against 5-FU-resistant NUGC-3 was about 70% at a dose level of 200 mg/kg/day; this value was comparable to that against the parental NUGC-3. On the other hand, the tumor inhibition rates of 5 FU, FdUrd, and TS-1 against 5-FU-resistant NUGC-3 were lower than those against parental NUGC-3. Similar observations were made in an FdUrd-resistant human colorectal cancer cell line (DLD-1). TAS-102 was also effective in 5-FU-less sensitive human pancreatic cancer cell lines (PAN-12 and BxPC-3) and human esophagus cancer (T.T.) when compared with 5-FU or UFT. Our hypothesis was that a relatively short and high dosage of TAS-102 results in an additional mechanism of FTD incorporation into DNA other than thymidylate synthase (TS) inhibition. We then examined the effects of FTD on DNA at the cellular level. After treatment with FTD or FdUrd, the DNA fragmentation pattern was examined using filter elution and in situ nick translation. Treatment with FTD for 2 h resulted in marked DNA fragmentation. When the tumor cells were treated with FTD for 72 h or with FdUrd for 2 or 72 h, only a small amount of DNA fragmentation was observed, and the appearance of the tumor cells did not differ markedly from that of untreated cells. Moreover, the DNA fragmentation rate in the TAS-102 treatment group was significantly higher than that in the control group in vivo. These results suggest that when tumor cells are exposed to high concentrations of FTD for short periods of time, FTD manifests its antitumor activity primarily through the induction of DNA fragmentation after FTD incorporation into the DNA. We conclude that TAS-102 is expected to manifest antitumor effects against 5-FU resistant tumors that are similar to those exerted in 5-FU-sensitive tumors. PMID- 15289859 TI - Cyclin D1 overexpression is not a specific grouping marker, but may collaborate with CDC37 in myeloma cells. AB - Cyclin D1 is a positive-regulator of the cell cycle and is overexpressed in myeloma cells with t(11;14)(q13;q32). First, we analyzed whether there was a correlation between cyclin D1 overexpression and the presence of Ki67-positive myeloma cells in multiple myeloma (MM). Cyclin D1 overexpression was examined by competitive RT-PCR. Then we found these two markers were present independently in a given case. FISH analysis revealed that cyclin D1 over-expression was caused by t(11;14)(q13;q32) or extra copies of B-cell leukemia/lymphoma-1 (BCL-1/CCND1), and unknown mechanism without them. We compared the gene expression between myeloma cells with cyclin D1 overexpression and those without it using cDNA microarray analysis. Analysis of the expression profiles showed that the significantly up-regulated genes included cyclin D1, cell division cycle 37 (CDC37) and B-cell leukemia/lymphoma-2 (BCL-2), while the down-regulated genes included cyclin D2 and CD9 antigen (p24) in MM cases with cyclin D1 overexpression. However, hierarchical clustering analysis of the data showed that myeloma cells of MM cases with cyclin D1 overexpression could not be distinguished clearly from those without it. Real-time RT-PCR showed that the expression of CDC37 gene was significantly up-regulated in MM patients with cyclin D1 overexpression compared with those without it (p=0.0418). However, there was no significant difference in BCL-2 gene (p=0.5748). These results suggested that MM cases with cyclin D1 overexpression do not constitute a specific group, and cyclin D1 overexpression may not be caused only by abnormality of the BCL-1/CCND1 gene. The CDC37 may collaborate with cyclin D1 in progression of MM. PMID- 15289860 TI - Application of RT-PCR to clinical diagnosis of micrometastasis of colorectal cancer: A translational research study. AB - We previously reported in a retrospective study that CEA-based RT-PCR was useful for predicting the prognosis of patients with node-negative colorectal cancer. RT PCR is well established for laboratory use, but many issues remain to be resolved prior to its clinical application. In addition to the false positive rate of RT PCR, we addressed several issues, including the timing of lymph node sampling, stability of RNA after surgery, and reproducibility of results. After appropriate modification, including development of a tissue sampling kit, a multi institutional clinical study was commenced prospectively from November 2001, and 100 patients were enrolled for examination of micrometastasis. RNA was stable in lymph nodes for up to 3 h after surgical resection. This range of sampling time was acceptable to the surgeons. RNA was well preserved in RNA later at -20 degrees C for 3 weeks. Dilutions of MKN45 and LoVo cells served as positive controls for conventional PCR since these controls were found to be highly stable and ensured reproducibility. Moreover, simultaneous use of quantitative PCR (Light Cycler) ensured double confirmation of the results. Our clinical study showed that the quality of RNA was excellent or good in most samples (98 of 100; 98%). Twenty-four of 98 (24.5%) cases were judged to be micrometastasis-positive. In conclusion, the current translational research study established a clinically feasible RT-PCR system for micrometastasis. Our system could potentially be useful as a clinical tool. PMID- 15289861 TI - Classification of distinct subtypes of peripheral T-cell lymphoma unspecified, identified by chemokine and chemokine receptor expression: Analysis of prognosis. AB - WHO classification for malignant lymphoma was recently proposed. However, PTCL is heterogeneous. Chemokines and its receptors are closely associated with the T cell subtypes. To clarify the T-cell subtype in PTCL, we conducted DNA chips of chemokine, its receptor (R) and cytokines. Angioimmunoblastic T-cell lymphoma (AILD, n=4), anaplastic large cell lymphoma (ALCL, n=4), adult T-cell leukemia lymphoma (ATLL, n=7), NK-cell lymphoma (NKL, n=2) and PTCL, unspecified (PTCL-U, n=6) were analyzed using DNA chips. In addition, immunological stainings were performed in 280 cases. In DNA chip, AILD, ALCL, NKL and ATLL showed a tendency for respective clusters, otherwise, PTCL-U clustered with AILD, ALCL and ATLL. From the gene expression profiling, CCR4, CCR3, MIG, CXCR3 and BLC were selected for immunohistochemistry. ATLL (n=48) expressed CCR4. ALCL (n=26) expressed CCR3, NKL (n=20) expressed MIG, and AILD (n=29) expressed CXCR3 and/or BLC. From the expression patterns, PTCL-U (n=134) were classified into three groups; CCR4 type (CCR4(+), n=42), CCR3 type (CCR3(+), n=31) and CXCR3 type (CXCR3(+) BLC(+/-), n=54). The prognosis was poor for ATLL, intermediate for AILD and favorable for ALCL (P=0.0014). Among PTCL-U, CCR4 type, CXCR3 type and CCR3 type had prognoses equivalent to ATLL, AILD and ALCL, respectively (P<0.0001). PMID- 15289862 TI - Two subtypes of mucinous colorectal carcinoma characterized by laser scanning cytometry and comparative genomic hybridization. AB - To data, there have been no comprehensive cytogenetic studies of mucinous colorectal carcinomas (MUCs). We used comparative genomic hybridization (CGH) and laser scanning cytometry (LSC), to analyze cytogenetic changes in 21 MUCs and to compare the results with those of our previous study of 60 non-MUCs. Six of 21 MUCs were aneuploid and 15 were diploid. Gains of 13q, 8q, 2q, 12p and 18p were more frequent aberrations. Recurrent decreases in DNA copy number were found frequently at 17p, 22q, 1p, 16p and 8p. Amplifications of 8q, 5p, 12, 18p, 13q and 20p were observed in aneuploid tumors. The average number of DNA sequence copy number aberrations (DSCNAs) was significantly higher in aneuploid MUCs than in diploid ones. Aneuploid MUCs were clinicopathologically more aggressive, with greater lymph node involvement, distant organ metastasis, recurrence after surgery, higher stage and poorer prognoses. Gain or amplification of 18p was detected in 5 of 6 aneuploid MUCs but not in diploid MUCs or non-MUCs. When the average number of DSCNAs was compared among MUCs and well, moderately, and poorly differentiated adenocarcinomas, the average number of DSCNAs was significantly lower in diploid MUCs; however, with aneuploid tumors, the average number of DSCNAs in MUCs was similar to that in poorly differentiated adenocarcinomas but higher than that in well and moderately differentiated cancers. Moreover, tumor cells were well differentiated in diploid MUCs but poorly differentiated in aneuploid MUCs. These data suggest that MUCs have two types with different genetic pathways, histologic characteristics, and behavior. PMID- 15289863 TI - Evaluation of Hypocrellin B in a human bladder tumor model in experimental photodynamic therapy: biodistribution, light dose and drug-light interval effects. AB - Hypocrellin B (HB), a monomeric perylenequinone pigment, is a promising second generation photosensitizer for photodynamic therapy. We have evaluated the efficacy of HB mediated PDT by experimenting with various drug-light intervals, based on the biodistribution analysis in human bladder tumor (MGH cell line) models. Tumor growth rates were assessed at 10-day post treatment followed by morphometric analysis. Biodistribution of HB was evaluated using spectrofluorophotometry analysis (Ex: 480 nm, Em: 620-630 nm). The level of HB peaked at 6 h postinjection in tumor, peritumoral skin and normal muscle followed by a decline over the next 42 h. Concurrently, the ratio of drug in tumor versus skin was relatively low at all times in comparison to tumor to muscle ratio. In serum, concentration of HB peaked at 1 h. Almost 88% of its original uptake level was cleared at 48 h. The level of PDT response revealed a strong dependence on the drug-light intervals (DLI) and light dose. For both high and low fluence/fluence rate, comparable tumor response was observed at 1 h DLI; treated tumors exhibited significant tumor regression compared to 6 and 24 h DLI. The absence of tumor response was observed at 24 h DLI even at high light dose (100 J/cm(2); 100 mW/cm(2)). Tumor response detected at low light dose (12 J/cm(2); 12 mW/cm(2)) at short DLI suggests that the tumor vasculature is a more sensitive target compared to the cellular compartment of the tumor, correlating significantly with the bioavailability of the drug in serum. Therefore, HB mediated PDT effect is characteristics of a predominantly vascular mediated effect. This study confirms that for short drug-light intervals, PDT seems to target tumor vasculature, which contributes to tumor destruction. PMID- 15289864 TI - Thiazolidinedione, a peroxisome proliferator-activated receptor-gamma ligand, inhibits growth and metastasis of HT-29 human colon cancer cells through differentiation-promoting effects. AB - Peroxisome proliferator-activated receptor-gamma (PPAR-gamma) ligands inhibit the growth of PPAR-gamma expressing cancer cells through terminal differentiation. However, there are few studies examining the effect of a PPAR-gamma ligand on metastatic potential of cancer cells in an animal model and the underlying molecular mechanisms. We have recently developed a rectal cancer xenograft animal model in which anti-tumor and anti-metastatic efficacy of agents can be evaluated. This study was designed to examine whether a representative PPAR-gamma ligand, thiazolidinedione (TZD), could inhibit growth and metastasis of PPAR gamma positive HT-29 human colon cancer cells through the induction of terminal differentiation. TZD caused G1 arrest in association with a marked increase in p21Waf-1, Drg-1, and E-cadherin expression. In untreated cancer cells, fluorescence immunostaining demonstrated beta-catenin in the nucleus and/or cytoplasm; in TZD-treated cancer cells, beta-catenin localization shifted to the plasma membrane, in association with increased E-cadherin at this site and reduced tyrosine phosphorylation of beta-catenin. In addition, TZD completely inhibited lymph node and lung metastases in the xenograft animal model, and TZD inhibited growth of primary xenografts by 40%. These results suggest that TZD can function as a cytostatic anti-cancer agent to inhibit growth and metastasis of HT 29 colon cancer cells through differentiation-promoting effects. These effects involve not only modulation of the E-cadherin/beta-catenin system, but also up regulation of Drg-1 gene expression. PMID- 15289865 TI - Loss of SFRP1 is associated with breast cancer progression and poor prognosis in early stage tumors. AB - Aberrant activation of the Wnt signaling pathway plays an important role in the development of solid tumors such as breast and colon cancer. Secreted Frizzled related protein 1 (SFRP1) is a negative regulator of the Wnt pathway. It has been described that SFRP1 mRNA is strongly down-regulated in breast cancer and a putative tumor suppressor function has been postulated. We have generated and characterized an SFRP1 specific antibody to analyze its expression on protein level and to investigate the association of SFRP1 expression with clinicopathological parameters and patient survival. Analysis of >2000 invasive breast tumors and 56 carcinoma in situ revealed similar frequencies of SFRP1 loss in these tumors (46% and 43% respectively). Therefore, we propose that loss of SFRP1 expression is an early event in breast tumorigenesis. SFRP1 expression was inversely correlated with tumor stage (p<0.001) but not with tumor grade (p=0.14) or lymph node status (p=0.84). Performing a multivariate analysis we could confirm the association between tumor stage and SFRP1 expression (p=0.029). In particular, loss of SFRP1 expression in early stage breast tumors (pT1) was associated with poor prognosis (p=0.04). In conclusion, expression of SFRP1 is commonly lost in breast cancer. SFRP1 expression might be useful as a novel prognostic marker in early stage breast cancer. PMID- 15289866 TI - Bid sensitizes apoptosis induced by chemotherapeutic drugs in hepatocellular carcinoma. AB - Hepatocellular carcinoma (HCC) is one of the most common malignancies in Asia. HCC is often resistant to chemotherapy and the mechanism remains unclear. Mitochondrion-mediated pathway is critical in hepatocyte apoptosis, which suggests Bcl-2 family genes may play a role in the regulation of chemotherapy in HCC. In the present study, we investigated the role of BH3 domain-only protein Bid in HCC tissues, HCC-derived cell lines and how the expression of Bid was related to chemotherapeutic agent-induced apoptosis. Bid was differently expressed in HCC tissues and hepatoma cell lines. Hep3B, a Bid-abundant HCC cell line, was more sensitive to drug-induced cytotoxicity than PLC/PRF/5, a Bid insufficient HCC cell line. The level of caspase activity induced by 5 fluorouracil (5-FU) was higher in Hep3B than in PLC/PRF/5 and a significant increase in the activity occurred at a rather late stage, after 48 h of the treatment. Similar to the activation of caspase, Bid cleavage and activation was only significant at 72 h after the treatment. Overexpression of Bid or tBid sensitized HCC cells to 5-FU and doxorubicin (Dox) treatments. We further demonstrated that such a sensitive effect could be offset by Bcl-xL, as Bid- or tBid-induced apoptosis was completely blocked by the over-expression of Bcl-xL. These results indicate the level of Bid expression is closely associated with the sensitivity of HCC cells to chemotherapeutic drugs, suggesting that Bid plays an important role in HCC management. PMID- 15289867 TI - Flavone inhibition of tumor growth via apoptosis in vitro and in vivo. AB - Colorectal carcinoma is a human malignant tumor, which is very resistant to currently available methods of treatment. Therefore, developing an effective agent with anti-colorectal carcinoma activity is important. In the present study, 8 structurally related flavones including flavone, 3-OH flavone, 5-OH flavone, 7 OH flavone, quercetin, kaempferol, quercetin, and morin were used to study their effects on colorectal carcinoma cells (HT29, COLO205, COLO320-HSR). Results of MTT assay indicated that flavone shows the most potent cytoxic effect among them on these three cell types. The cytotoxicity induced by flavone is mediated by inducing the occurrence of apoptosis characterized by the appearance of DNA ladders, apoptotic bodies and hypodiploid cells. Activation of caspase 3 protein procession and enzyme activity with inducing cleavage of caspase 3 substrates PARP was identified in flavone-treated cells, and an inhibitory peptide Ac-DEVD FMK for caspase 3, but not Ac-YVAD-FMK for caspase 1, attenuates the cytotoxic effect of flavone in COLO205 and HT29 cells. Elevation of p21 but no p53 protein was observed in flavone-treated cells. Increasing intracellular peroxide level was detected in flavone-treated cells by DCHF-DA assay, and antioxidants such as tiron, catalase, SOD, PDTC, but not DPI, suppress flavone-induced cytotoxic effect. In vivo anti-tumor study indicates that flavone exhibits ability to inhibit tumor formation elicited by s.c. injection of COLO205 cells in nude mice, and apoptotic cells and an increase in p21, but not p53, protein were observed in tumor tissues derived from flavone-treated group. Additionally, flavone induced apoptosis in primary colon carcinoma cells COLO205-X with appearance of DNA ladders, caspase 3 protein procession, PARP protein cleavage, and an increase in p21 (not p53) protein. These data provide evidence to suggest that flavone is an effective agent to induce apoptosis in colorectal carcinoma cells in vitro and in vivo; activation of caspase 3, ROS production, and increasing p21 protein are involved. PMID- 15289868 TI - Deregulation of PKB influences antiapoptotic signaling by PKC in breast cancer cells. AB - We have previously shown that novel protein kinase C (nPKC) isozymes, such as nPKCepsilon, negatively regulate TNF-induced apoptosis in breast cancer cells although the level on nPKCs did not correlate with cellular sensitivity to TNF. In the present study, we examined if the level/activation status of Akt/PKB influences antiapoptotic signaling by nPKCepsilon. While MCF-7 cells overexpressed PKB, BT-20 and SKBR-3 cells expressed constitutively phosphorylated PKB, and MDA-MB-231 cells expressed unphosphorylated PKB. Ly294002, an inhibitor of PI-3 kinase, induced cell death in SKBR-3 cells, which contained little nPKCs. Although Ly294002 by itself had only a modest effect on cell death in BT-20 and MCF-7 cells, it potentiated sensitivity of these cells to TNF. In contrast, Ly294002 either alone or in combination with TNF had little effect on cell death in MDA-MB-231 cells. These results suggest that the status of PKB in breast cancer cells influences antiapoptotic signaling by PKC. PMID- 15289869 TI - Doxorubicin potentiates TRAIL cytotoxicity and apoptosis and can overcome TRAIL resistance in rhabdomyosarcoma cells. AB - Doxorubicin (DOX) and ifosfamide (IFO) are the most active single agents in soft tissue sarcomas (STS). Tumour necrosis factor-alpha (TNF-alpha) is used for STS in the setting of isolated limb perfusions. Like TNF-alpha, TNF-related apoptosis inducing ligand (TRAIL) induces apoptosis. In contrast to TNF-alpha preliminary studies suggest that TRAIL lacks systemic side effects. The effects of TRAIL alone and in combination with DOX or 4-hydroxy-IFO were evaluated in the TNF alpha sensitive rhabdomyosarcoma cell line KYM-1, its 5-fold TNF-alpha sensitive subline KD4 and its >150-fold TNF-alpha resistant subline 37B8R. Membrane expression of TRAIL-receptors DR4 (death receptor 4), DR5 (pro-apoptotic), DcR1 (decoy receptor 1), DcR2 (anti-apoptotic) was assessed by flow cytometry. Cytotoxicity was determined by microculture tetrazolium assays. Apoptosis assays were performed with acridine orange. DOX (doxorubicin) and 4-OH-IFO decreased survival in all cell lines; a 2-fold resistance was observed for both drugs in 37B8R. All cell lines expressed DR4 and DR5, but hardly any DcR1 or DcR2. TRAIL was cytotoxic in KYM-1, even more in KD4 and induced massive apoptosis; 37B8R was >500-fold resistant to TRAIL and little apoptosis could be observed. TRAIL plus DOX showed synergistic cytotoxicity in KYM-1 and 37B8R. TRAIL plus 4-OH-IFO showed addition in all three cell lines. DOX plus TRAIL-induced more cytotoxicity and apoptosis in all cell lines compared to TRAIL alone. In 37B8R, DOX overcame resistance to TRAIL. In KYM-1, KD4 and 37B8R, sensitivity and resistance to TNF alpha and TRAIL parallels. TRAIL-resistance was independent from expression of TRAIL-receptors. DOX with TRAIL could overcome TRAIL-resistance in 37B8R cells. PMID- 15289870 TI - Prevalent hyper-methylation of the CDH13 gene promoter in malignant B cell lymphomas. AB - CDH13 (H-cadherin) is a member of the cadherin superfamily, which plays an important role in cell recognition and adhesion. We examined the expression and methylation status of the CDH13 gene in diffuse large B cell lymphomas (B-DLCLs). We found decreased expression of the CDH13 gene in all of 6 hematopoietic cell lines by reverse transcription-polymerase chain reaction (RT-PCR). Promoter hyper methylation of the gene was detected in all 6 cell lines and in 13 of 19 (68%) B DLCL samples by methylation-specific PCR. Interestingly, the methylation frequency of the CDH13 gene was comparable to those of the tumor suppressor genes p15 (68%) and p16 (74%) detected in B-DLCLs. Sequencing of bisulfite-treated DNA revealed hyper-methylation of the CpG islands of the CDH13 promoter in B-DLCLs and the cell lines. Treatment with 5-aza-2'-deoxycytidine restored CDH13 gene expression in a cell line in which promoter hyper-methylation and impaired expression of the CDH13 gene were observed. Loss of heterozygosity (LOH) around the CDH13 gene on chromosome 16q24 was detected in 6 of 15 (40%) informative cases with microsatellite marker D16S507 and in 6 of 15 (40%) cases with D16S422 in B-DLCLs. In all of 4 B-DLCL cases which showed both promoter methylation and LOH at the two marker loci, expression of the CDH13 gene was significantly low. These results suggest that silencing of the CDH13 gene by aberrant promoter methylation and allelic deletion is associated with tumorigenesis in a subset of B-DLCL. PMID- 15289871 TI - Multiple EBNA1-binding sites within oriPI are required for EBNA1-dependent transactivation of the Epstein-Barr virus C promoter. AB - The transactivating function of the oriPI-EBNA1 complex is essential for activation of the Epstein-Barr virus (EBV) C promoter (Cp) in lymphoblastoid cell lines expressing the viral growth programme. Furthermore, the oriPI-EBNA1 complex is believed to play an important role during promoter switching upon primary infection of B-lymphocytes and establishment of latent infection in vivo. Previously, it was shown that six EBNA1-binding sites within oriPI were required for transactivation of the heterologous thymidine kinase promoter. Here, we define the number of EBNA1-binding sites within oriPI necessary for its biological function as EBNA1-dependent Cp enhancer. We show that four EBNA1 binding sites within oriPI lead to significant upregulation of Cp in response to EBNA1 and eight or more to full activation. Thus, multiple EBNA1 homodimers at oriPI are required for the formation of a transcriptionally active Cp complex, a process that involves EBNA1-induced changes in the chromatin structure including DNA looping and nucleosome destabilization. PMID- 15289872 TI - Inhibition of the ubiquitin-proteasome pathway activates stress kinases and induces apoptosis in renal cancer cells. AB - The ubiquitin-proteasome pathway plays a critical role in the degradation of cellular proteins related to signal transduction. Cytokine and growth factor dependent aberrant proliferation has been implicated in renal cell carcinoma (RCC). We hypothesized that inhibiting the proteasome function might activate a proapoptotic signal transduction by modulating the cytokine and growth factor related signal transduction pathway. We therefore investigated the effectiveness of a proteasome inhibitor in the treatment of RCC regarding the involvement of Mitogen-activated protein kinases (MAP kinases), because MAP kinases are major signal transduction molecules that are known to play a pivotal role in cancer cell proliferation or apoptosis triggered by extra-cellular cytokines and growth factors. A proteasome inhibitor, MG132 inhibited the proliferation of RCC cell lines, 786-O and KU20-01 in a time and dose-dependent manner. 786-O cells have truncated von-Hippel Lindau (VHL) tumor suppressor gene protein due to a one base pair deletion at exon 1, whereas KU20-01 cells have a wild-type VHL protein. MG132 induced apoptosis in both cell lines. The inhibition of the ubiquitin proteasome pathways was confirmed by the accumulation of ubiquitin-tagged proteins. MG132 induced the phosphorylation of ERK at 4 h and thereafter persisted for 8 to 16 h. In contrast, JNK and p38 activation persisted for longer periods and remained enhanced until 24 h. The concomitant activation of effector caspases, caspase-3 and caspase-7 was observed in 786-O cells. The inhibition of the proteasome function can induce apoptosis in RCC irrespective of the VHL protein status. The persistence of JNK and p38 activation may therefore be a unique mechanism underlying MG132 induced apoptosis. PMID- 15289873 TI - Anti-EpCAM monoclonal antibody (MAb17-1A) based treatment combined with alpha interferon, 5-fluorouracil and granulocyte-macrophage colony-stimulating factor in patients with metastatic colorectal carcinoma. AB - Monoclonal antibodies (MAbs) have different modes of action and toxicity profile compared to chemotherapeutics, which makes it interesting to combine these drugs. Addition of cytokines to MAb therapy may also augment immune effector functions utilized by MAb. In an effort to improve the therapeutic effect of a MAb-based regimen in colorectal carcinoma (CRC) patients, the effects of a combination of alpha-interferon (alpha-IFN), 5-fluorouracil (5-FU), granulocyte-macrophage colony-stimulating factor (GM-CSF) and mouse MAb17-1A was evaluated in 27 patients with metastatic disease. alpha-IFN was given s.c. once daily for 5 consecutive days and at days 4 and 5, 5-FU was administered as a daily i.v. bolus injection. After 2 days rest, GM-CSF was given s.c. once daily, days 8-14 and on day 10, MAb17-1A was given i.v. The treatment cycle was repeated every 4th week. One patient achieved a partial remission and 13 patients showed a minor response or stable disease >3 months, inducing an overall response rate of 54%. Responding patients survived significantly longer than non-responding patients (p=0.021). Median overall survival time for all patients was 75 weeks and progression-free survival time 15 weeks. Adverse events related to alpha-IFN, GM-CSF and 5-FU were as expected. The frequency of patients with an immediate-type allergic reaction (ITAR) against MAb17-1A at the 1st, 2nd, 3rd and 4th treatment cycles was 11%, 52%, 62% and 64% respectively. The planned MAb17-1A dose had to be reduced by repeated infusions. No patient received full dose of MAb17-1A from the 3rd cycle and onward. Compared to historical control patients treated with MAb17-1A alone, the present combination regimen seemed to improve the response rate (54% vs 15%) as well as progression-free survival (15 vs 7 weeks; p<0.05). PMID- 15289874 TI - Targeting hypoxic cancer cells with a protein prodrug is effective in experimental malignant ascites. AB - Tumor hypoxia in a solid tumor mass has long been recognized as a cause of resistance to current cancer therapies, and has also been suggested to be a potent driving force towards malignancy. Recent progress in the understanding of the molecular mechanism of the tumor response to hypoxia has increased attention on targeting hypoxia for cancer therapy. We have generated a hypoxia-targeting fusion protein, TOP3, which is composed of a protein transduction domain (PTD) of HIV TAT, an oxygen-dependent degradation domain (ODD) of HIF-1 alpha, and procaspase-3. Here, we examine the effects of TOP3 in a rat ascites model. First, we clarified that the fluid in ascites from MM1 cells, which are derivatives of AH130 rat ascites hepatoma cells, was highly hypoxic. In vitro, MM1 cells retained protein degradation machinery through the ODD domain, and TOP3 effectively impaired MM1 cell growth in culture under hypoxic conditions by inducing apoptosis. Intraperitoneal administration of TOP3 prolonged the life span of rats bearing a significant amount of malignant ascites, and 60% of the treated animals were cured without recurrence of ascites. Thus, TOP3 had a dramatic effect on malignant ascites and, hence, we propose that rodent malignant ascites is an appropriate platform for testing hypoxia-targeted drugs. PMID- 15289875 TI - Loss of caspase-8 activation pathway is a possible mechanism for CDDP resistance in human laryngeal squamous cell carcinoma, HEp-2 cells. AB - Cisplatin (CDDP) is among the most widely used and most effective chemotherapeutic agent for many types of human cancer. Because killing cancer cells by chemotherapy is principally executed by apoptosis, a defective apoptotic program might acquire drug resistance. Flow cytometric Annexin V assay demonstrated that HEp-2 cells (human laryngeal cancer) were persistently resistant to CDDP as compared to HeLa cells (human uterine cervical cancer), despite the same histological type and wild-type p53 status. CDDP treatment caused steady induction of p53 protein in both cancer cell types, although it was more dramatic in CDDP-resistant HEp-2 cells, which was correlated well with p53 Ser15 phosphorylation, but not with the expression level of HPV type 18 E6 oncoprotein in these cells. Importantly, CDDP differently activated caspase cascades between HEp-2 and HeLa cells. CDDP activated the caspase-8 pathway through TNFR superfamily receptors such as Fas, but not caspase-9 in HeLa cells. On the other hand, the caspase-9 pathway was significantly activated in HEp-2 cells, although the activation of caspase-8 by CDDP was deficient. This different response to CDDP in caspase-8 activation was not related with the expression level of either Fas or FasL in these cells. We concluded from these results that loss of the caspase-8 activation pathway in HEp-2 cells was a possible mechanism for its resistance to CDDP-induced apoptosis. The caspase-8 pathway might play an important role in CDDP-induced apoptosis in HPV-positive human squamous cell carcinomas. PMID- 15289876 TI - Topoisomerase inhibitors enhance the cytocidal effect of AAV-HSVtk/ganciclovir on head and neck cancer cells. AB - Adeno-associated virus (AAV) is a non-pathogenic virus with a single-strand DNA genome. AAV vectors have several unique properties suited for gene therapy applications. However, an obstacle to their application is a low efficiency of transgene expression, mainly due to a limited second-strand synthesis. Previously, we reported that gamma-rays enhanced the transduction efficiency and cytocidal effect of AAV vector harboring the herpes simplex virus-thymidine kinase (AAVtk) and ganciclovir (GCV) system. In the present study, we investigated whether topoisomerase inhibitors (etoposide and camptothecin) enhance the AAV vector-mediated transgene expression and the killing effect by AAVtk/GCV system. The enhancement of transgene expression was observed in a concentration-dependent manner on human laryngeal carcinoma cells (HEp-2 cells) and HeLa cells. Southern analysis confirmed that etoposide enhanced the double strand synthesis of the AAV vector genome in HEp-2 cells and HeLa cells. The cells were efficiently killed by AAVtk/GCV system, as expected. More importantly, both etoposide and camptothecin augmented the cytocidal effect of the AAVtk/GCV system. These findings suggest that the combination of AAV-mediated suicide gene therapy and treatment with topoisomerase inhibitors may have synergistic therapeutic effects in the treatment of cancers. PMID- 15289878 TI - The interaction of tenascin-C with fibronectin modulates the migration and specific metalloprotease activity in human mesothelioma cell lines of different histotype. AB - Malignant mesothelioma (MM) is a tumor with local invasive behaviour. Tenascin-C (TN-C) with fibronectin (FN) are associated extracellular matrix (ECM) molecules frequently neo-expressed in stromal remodeling during neoplastic progression, mostly at the invasive edge of these tumors. Tenascin-C alone or in association with other ECM molecules, could play an important role in the process of tumor invasion, acting as substrate for movement or modulating the migration on FN or promoting the degradation of ECM. Three mesothelioma cell lines of different histotype were analysed for the adhesive capacity on TN-C. The haptotactic activity on TN and the TN modulation of migration on a substrate of FN were analysed by a Boyden modified chamber. The effects of TN on proteolytic activity was evaluated by zymography. None of the lines adhered to tenascin. TN-C was not haptotactic for the three cell lines. Soluble or solid TN reduced the migration on FN of epithelial (E-MM) and sarcomatous cell line (S-MM), whereas enhanced the movement of a byphasic cell line (B-MM). When the cells were pretreated with anti integrin blocking antibodies we observed a different pattern of inhibition of migration on FN respect to FN plus TN. Finally, no difference of metalloprotease (MMP-2, MMP-3, MMP-7, MMP-9) activity was observed between cells plated on FN and on FN plus TN, except for B-MM which showed an increased MMP-7 activity when TN was added to FN. Although TN is not a substrate for movement of MM cell lines, it interacts with FN by modulating differently the migration, according to the different histotype and to the integrin involved, and increasing specific metalloprotease activity. PMID- 15289877 TI - Docosahexaenoic acid modulates different genes in cell cycle and apoptosis to control growth of human leukemia HL-60 cells. AB - Although previous studies have shown that docosahexaenoic acid (DHA; 22:6 omega 3) from fish oils inhibits growth of different cancers, safety issues have been raised repeatedly about contaminations of toxins in these oils. Cultured microalgae are suggested recently as an alternative cleaner and safer source of the fatty acid. We investigated in this study the function of DHA from the enriched microalga Crypthecodinium cohnii (ADHA) in cell-growth control and its mechanism in human leukemia HL-60 cells. ADHA retarded proliferation of the leukemia cells dose-dependently by 4-93% of the control level, after 72-h incubations with 10-160 micro M of the fatty acid; and the 50% inhibitory concentration (IC50) was estimated as 74 micro M. DNA-flow cytometry study showed that ADHA arrested G0/G1 cells by 12-22% and induced apoptotic cells by 569-906% of their controls, after incubation with the IC50 of ADHA for 24, 48 and 72 h. The modes of cell-cycle arrest and pro-apoptotic actions of ADHA were further elucidated. Gene-array analysis illustrated that ADHA modulated a number of cell cycle and apoptosis genes to control the cell growth; in particular, the fatty acid up-regulated the transcriptional repressor E2F-6 and pro-apoptotic Bax by 1435 and 4172% respectively, after 24 h of incubation. Semi-quantitative RT-PCR study further showed that ADHA induced elevation of the Bax mRNA transcript time dependently. In meanwhile, ADHA also induced phosphorylation and thus inactivation of Rb protein in the leukemia cells. All these results suggest that ADHA up-regulates Bax and inactivates Rb protein to induce the cell-growth control and apoptosis in human leukemia HL-60 cells. PMID- 15289879 TI - Identification and characterization of a novel human FOXK1 gene in silico. AB - We describe here the identification and characterization of a human forkhead box gene, FOXK1, that encodes predicted proteins most homologous to the mouse myocyte nuclear factor (MNF)/forkhead box K1 (FoxK1). Human FOXK1 is located at the chromosomal position 7p22. Two transcript variants, FOXK1a and FOXK1b, containing the divergent 5'-ends are identified. Sequence comparison indicated that the human FOXK1 proteins share 66-74% amino acid identity (99% identity in the forkhead domain) with their mouse ortholog. The in silico expression analysis showed FOXK1 expression in immature tissues of brain, eye, heart, lung and thymus. In adults, FOXK1 was predominantly expressed in many malignant tissues, in particular in the tumors of brain, colon and lymph node. This gene is therefore implicated in the process of both normal and neoplastic development. PMID- 15289880 TI - Identification and characterization of TMEM24 family genes in silico. AB - MLL gene at human chromosome 11q23.3 is frequently rearranged or amplified in hematological malignancies, while PHLDB1, BCL9L, FOXN5 (FOXR1), RNF26, and MFRP genes linked to MLL gene are deleted in neuroblastoma. Here, we characterized the TMEM24 gene family by using bioinformatics. KIAA0285 gene within the 11q23.3 commonly deleted region of neuroblastoma was designated TMEM24, because KIAA0285 gene product was a 707-aa (or 706-aa) protein with N-terminal short cytoplasmic region, single transmembrane domain, and C-terminal large extracellular region. C21orf25 (NM_199050.1) encoded an N-terminally truncated 541-aa protein homologous to TMEM24. Complete coding sequence of C21orf25 was determined by assembling exons 1 and 2 within human genome sequence AP001745 and NM_199050.1 cDNA. Full-length C21orf25 encoded a 696-aa TMEM24-related protein with similar membrane topology. Exon-intron structure was conserved between TMEM24 and C21orf25 genes. TMEM24-ABCG4 locus at human chromosome 11q23.3 and C21orf25-ABCG1 locus at human chromosome 21q22.3 were paralogous regions (paralogons) with insertions of other genes due to recombination during evolution. Mouse 1300006O23 (BC060156.1) and 5830404H04 (NM_174847.1) cDNAs were derived from orthologs of human TMEM24 and C21orf25 genes, respectively. TMEM24 homologous domains (TM24H1 and TM24H2) were identified as novel domains conserved among TMEM24, C21orf25, 1300006O23, and 5830404H04. Human TMEM24, C21orf25 and their mouse homologs were type II transmembrane proteins with extracellular TM24H1 and TM24H2 domains. This is the first report on identification and characterization of the TMEM24 family. PMID- 15289881 TI - Identification and characterization of human DFNA5L, mouse Dfna5l, and rat Dfna5l genes in silico. AB - An evolutionary recombination hotspot around the GSDML-GSDM locus at human chromosome 17q21 is closely linked to an oncogenomic recombination hotspot around the PPP1R1B-STARD3-TCAP-PNMT-PERLD1 (MGC9753)-ERBB2-C17orf37 (MGC14832)-GRB7 locus at human chromosome 17q12. Here, we identified DFNA5L (GSDMDC1) gene related to GSDM and GSDML genes by using bioinformatics. Human DFNA5L gene at chromosome 8q24.3 was linked to ZC3HDC3, PP3856, EEF1D, and TIGD5 genes. NM_024736.4 (AK127941.1), AK022212.1, BC008904.2, and BC069000.1 cDNAs were derived from human DFNA5L gene. BC008904.2 was the representative human DFNA5L cDNA, while NM_024736.4 was an aberrant human DFNA5L cDNA with frame shifts due to the retention of introns 1, 3, 4, 5 and 8. Human DFNA5L mRNA was expressed in placenta, pancreatic cancer, prostate cancer, melanoma, salivary gland tumor, Jarkat T cells, and Ramos B cells. Complete coding sequence of rat Dfna5l cDNA was determined by assembling 11 exons of rat Dfna5l gene within AC120830.4 genome sequence, and that of mouse Dfna5l cDNA was derived from 1810036L03 (NM_026960.1). Exon-intron boundaries were conserved among human DFNA5L and rodent Dfna5l genes. Human DFNA5L (484 aa) showed 59.5% total-amino-acid identity with rat Dfna5l (488 aa), and 58.7% total-amino-acid identity with mouse Dfna5l (487 aa). DFNA5L orthologs were DNFA5 (GSDM) domain containing DFNA5 DC or GSDMDC proteins with Coiled-coil and Leucine zipper domains. Human DFNA5L, GSDM, GSDML, MLZE, DFNA5 and their mammalian orthologs were found to constitute the DFNA5 DC (GSDMDC) family. Because DFNA5 and MLZE are cancer-associated genes, DFNA5L, GSDM, and GSDML are predicted cancer associated genes. PMID- 15289882 TI - Allelic variation of BAT-25 and BAT-26 mononucleotide repeat loci in tumours from a group of young women with breast cancer. AB - Genomic instability in the form of repeat number variation at microsatellite loci has been reported in several human cancers. To resolve current confusion regarding frequency of microsatellite instability (MSI), standardised protocols have proposed use of a consensus set of informative loci; it is claimed that analysis of 2 apparently quasi-homozygous, quasi-monomorphic, mononucleotide repeats (BAT-25 and BAT-26) is sufficiently accurate to define MSI (obviating need for corresponding constitutive DNA). We examined these loci in 163 breast cancers, 108 of which had previously been analysed at 11-24 other loci and found to have MSI in 38% of them. For BAT-26 only 1/153 (homozygous) tumours showed a contracted allele, with minor size variations (1-6 bp) between individuals. For BAT-25 repeat contractions were unambiguously observed in 12 (7.4%) tumours; only 4 of these were previously designated MSI+. DNA from normal individuals showed significant allelic variation in 8/159 (5%) cases for BAT-25; no instance of heterozygosity was seen for BAT-26. Subsequently, we analysed normal DNA from the 12 individuals whose tumours had MSI at BAT-25; in 2 cases there was germline heterozygosity. We conclude that analysis with BAT-26 (in contrast to other loci) was not a useful detector of MSI. With BAT-25, a low frequency of MSI not much greater than germline polymorphism, also limits the utility of this marker for determining MSI in breast cancer. PMID- 15289883 TI - Radioresistance is associated to increased Hsp70 content in human glioblastoma cell lines. AB - Radiation therapy is routinely prescribed for high-grade malignant gliomas. However, the efficacy of this therapeutic modality is often limited by the occurrence of radioresistance, reflected as a diminished susceptibility of the irradiated cells to undergo apoptosis. Heat-shock proteins (Hsps) synthesis can be increased by cellular insults, such as radiation-induced damage. Inducible Hsp70 has been suggested to have multiple roles in cytoprotection against apoptosis. Accordingly, high levels of Hsp70 prevent stress-induced apoptosis. In the present study, we investigated whether the content of Hsp70 is associated to glioblastoma cell radioresistance. To this end, the U-87MG, U-251MG and MO59J human glioblastoma cell lines were irradiated at 2, 5 and 10 Gy and their relative radioresistance and Hsp70 were determined. Following 5 Gy irradiation, in MO59J and U-251MG a significant decrease in colony formation was found, whereas the U-87MG was relatively radioresistant. Three hours after the irradiation (at 5 Gy) Hsp70 contents increased 110% in the U-87 MG cells, but did not significantly change in the U-251MG and MO59J cells. Thus, our results suggest that Hsp70 protection against radiation-induced apoptosis might underlie glioblastoma radioresistance. PMID- 15289884 TI - Regularity of distribution of immunoreactive pulmonary surfactant protein A in rat tissues. AB - Existing data has shown that SP-A-like protein or mRNA is widely distributed in lamellar bodies such as tissues and mucosal surfaces. Using immunohistochemistry method with a polyclonal antibody against human SP-A, in this study we investigated distribution of immunoreactive pulmonary surfactant protein A (IR-SP A) in a number of rat tissues. The SP-A-like immunoreactivity was found in alveolar, parenchyma, pleura of lung; myelin sheath of brain; epithelia of Bowman's capsule, glomerulus and renal tubules of kidney; epithelia of colon, stomach, duct of salivary gland, pharynx; and blood vessel wall and connective tissue of extracellular matrix. The positive signal was blocked by pre-absorbed SP-A antigen from recombinant or bronchoalveolar lavage (BAL). SP-A has long been considered as an important frontier host defense molecule which participates in immune and inflammatory regulation of lung. With every inhalation, small particles, viruses, bacteria, and antigens from environment are continuously deposited onto the vast pulmonary epithelial surface. While a proper host defense is required to protect the lung, an over-exuberant response can disrupt the appropriate balance between pro- and anti-inflammatory. Traditional Chinese medicine believes that body is an open system relevant to the external environment. The physical, chemical and biological environmental factors constantly affect the open system, and the body properly reacts to maintain homeostasis of body machinery. The Chinese traditional medicine scholars have thus hypothesized that 'Qi' (meaning air) is the communication way between the body and external environment. What is 'Qi'? The results from our study suggest that IR-SP-A is a candidate of 'Qi'. It is compatible with the sites, theoretically containing collagenous and lectin domain molecules, also compatible with the primary injury sites of some autoimmune diseases. SP-A may be as one of 'Qi' molecules mentioned in traditional Chinese medicine that trigger some of autoimmune diseases. PMID- 15289885 TI - Native LDL and oxidized LDL modulate cyclooxygenase-2 expression in HUVECs through a p38-MAPK, NF-kappaB, CRE dependent pathway and affect PGE2 synthesis. AB - Native low density lipoproteins (n-LDL) and oxidized low density lipoproteins (Ox LDL) play a central role in atherogenesis and possess a wide variety of biological properties. We investigated whether n-LDL or Ox-LDL modulate cyclooxygenase-1 and -2 (Cox-1 and Cox-2) expression and prostaglandins release in human endothelial cells via an MAPK-dependent pathway. HUVECs were incubated in the presence of n-LDL or Ox-LDL (30 micro g/ml for both) for 2-15 h. Real-time PCR, western blotting and immunocytochemistry were used to investigate Cox-1 and Cox-2 expression. N-LDL and Ox-LDL induced Cox-2 expression in a time- and dose dependent manner. The Cox-2 protein was strongly induced 2 h after exposure to n LDL or Ox-LDL, the induction was maximal after 4 h and sustained for at least 8 h. The effect was specific for Cox-2, as Cox-1 expression was not modulated either by n-LDL or by Ox-LDL. The induction of Cox-2 expression was mainly dependent on the activation of p38 MAPK. Transient transfection analysis using a Cox-2 promoter showed that n-LDL and Ox-LDL exert their effects at the transcriptional level via NF-kappaB and CREB activation. N-LDL and Ox-LDL increased PGE2 release in a Cox-2-dependent manner while TXA2 and PGI2 release were not affected either by n-LDL or Ox-LDL. The finding that n-LDL and Ox-LDL induces Cox-2 in human endothelial cells through a p38 MAPK, NF-kappaB, CREB dependent pathway thus modulating PGE2 release, suggests a new mechanism by which these lipoproteins induce endothelial dysfunction, sustaining inflammatory processes in the arterial wall. PMID- 15289886 TI - Structural organization of the mouse neurochondrin gene. AB - Neurochondrin is a brain and bone specific leucine-rich protein. We previously cloned the two types of mRNAs (neurochondrin-1; 729 amino acids and neurochondrin 2; 712 amino acids) from mouse and human species. As a first step, to better understand the mechanism of the bone and brain specific and developmentally regulated expression of the neruochondrin gene, the genomic organization of murine neurochondrin was determined. It consists of 7 exons and spans about 10 kb; all splice junctions conform to the GT/AG rule. It codes for two alternatively spliced messenger RNAs, neurochondrin-1 containing all 7 exons and neurochondrin-2 lacking exon 1b but containing the other exons. Cap site analysis showed that the major transcription initiation occurs at 765 bp upstream of the ATG start codon of neurochondrin-1. The promoter region has no TATA and CAAT box like sequence but contains potential AP-1 and SP-1 binding sites. The neurochondrin gene is localized to mouse chromosome 4D1 and rat chromosome 5q36.11. PMID- 15289887 TI - New experimental models for the in vitro reconstitution of human bladder mucosa. AB - This study describes two experimental models for the in vitro reconstitution of the human bladder mucosa (neo-bladder): human urothelial stabilized cell lines were cultured on three-dimensional matrices, collagen or platelet-fibrin gels, containing murine fibroblast 3T3-J2 cells. Low-density seeding (2x10(4) cells/ml) of both normal (TCA-48) and neoplastic cell lines (TCA-47) on collagen matrix gave rise to isolated papillar colonies, while high-density seeding (3.75x10(6) cells/ml) led to the formation of wide pluristratified epithelial sheets, resembling the normal transitional epithelium. In contrast, high-density seeding (5x10(5) cells/ml) on platelet-fibrin matrix did not allow the formation of epithelial sheets: only isolated voluminous colonies of normal TCA-48 cells, and sparse and small colonies of neoplastic TCA-47 could be observed. Growth assays and cytotoxicity reduction tests showed that the growth inhibitory effect of platelet-fibrin gel on urothelial cells was probably due to the aspecific activation of the complement contained in the plasmatic fraction, whose precipitation forms fibrin-glue. Collectively, these findings allow us to draw the following conclusions: i) neobladders obtained by culturing urothelial cells on collagen matrix reproduce normal bladder mucosa and could be utilized in pharmacological studies; and ii) platelet-fibrin gels, that specifically inhibit neoplastic urothelial cell growth, could be used as scaffolds in surgical bladder reconstitution. PMID- 15289888 TI - Imatinib mesylate (STI571) interference with growth of neuroectodermal tumour cell lines does not critically involve c-Kit inhibition. AB - A therapeutic role of STI571 (imatinib mesylate) has been anticipated in patients with c-Kit positive neuroectodermal tumors. We examined the efficacy of STI571 to inhibit expansion of c-Kit positive neuroectodermal tumor cell lines in vitro and in a mouse model inoculated with ES (Ewing sarcoma) derived tumor cells, and investigated the molecular mechanism of STI571 action. Eleven tumor lines of ES, PNET (primitive neuroectodermal tumors) and NB (neuroblastoma) were assayed in the presence of 1, 5, 10, 15, 20 or 30 micro M STI571 for 24, 48, 72 h and 7 days. The mechanism of STI571 action was investigated using a microphysiometer cytosensor that determines cellular metabolic rates in the presence of STI571. c Kit and global protein phosphorylation was assayed by immunoprecipitation and a direct enzyme-linked immunoadsorbent assay after 72 h of 10 micro M STI571. Apoptosis was investigated by propidium iodide (PI), Annexin V staining and by enzymatic activity of caspase-3. Moreover, apoptotic gene expression was investigated using microarray technology. In nude mice, tumor volume and histology were analyzed in STI571 treated and untreated mice, and apoptotic gene expression analysis was performed on tumor masses. A decrease in cell proliferation and increase of cell apoptosis was caused by STI571 in a concentration- and time-dependent manner. Cytosensor microphysiometer and immunoprecipitation experiments demonstrated a time- and concentration-dependent decrease of cellular metabolic activity and global protein dephosphorylation after STI571 exposure. The inhibition by STI571 appeared at least to some extent independent of c-Kit inhibition since cells remained sensitive to SCF stimulation. Tumor volume was significantly reduced in STI571-treated mice compared to tumors from control inoculated non-treated mice. The apoptosis pathway in response to STI571 appeared not to be dependent on caspase activation, while gene expression profiles suggested accumulation of reactive oxygen species (ROS) resulting in cell death after exposure to STI571. The results point to the potential relevance of STI157 for neuroectodermal tumor therapy in view of its inhibitory effect on tumor cell growth, in spite of the observation that the inhibition of the c-Kit signaling pathway is not critically involved. PMID- 15289889 TI - Generation and characterization of dimeric small immunoproteins specific for neuroblastoma associated antigen GD2. AB - GD2 is a disialoganglioside expressed at high density on the surface of malignant cells of neuroectodermal origin, especially in neuroblastoma (NB) and melanoma. Since its expression in normal tissues is very restricted, GD2 represents an excellent target for neuroectodermal tumor targeting. Mini-antibody technology allows the production of dimeric single-chain antibodies, also called small immunoproteins (SIPs), which are composed of a scFv fused to a dimerizing domain of immunoglobulin heavy chains. Dimerization results in an increase of the total apparent affinity and a slower clearance in vivo than scFvs. These properties make SIPs very attractive molecules for tumor targeting. We isolated the variable regions from an anti-GD2 monoclonal antibody and exploited the SIP technology to generate two novel anti-GD2 SIPs. The first anti-GD2 SIP is a fully murine molecule containing the CH3 domain of mouse IgG1, whereas the second construct is a hybrid mouse-human molecule containing the CH4 domain of human IgE. Both mini antibodies were successfully produced and shown to retain binding specificity as well as an affinity similar to that of the original antibody. PMID- 15289890 TI - The expression of vascular endothelial growth factor-A and -C, and receptors 1 and 3: correlation with lymph node metastasis and prognosis in tongue squamous cell carcinoma. AB - We determined whether the expression of VEGF-A and -C and their receptors, Flt-1 and Flt-4, are associated with primary tumor size, regional lymph node metastasis, distant metastasis and prognosis in patients with tongue carcinoma. The expression of VEGF-A and -C, and their receptors, Flt-1 and Flt-4, in biopsy specimens taken from 73 patients with tongue carcinoma were examined by immunohistochemical staining. VEGF-A expression was associated with distant failure and VEGF-C expression correlated with locoregional recurrence and distant failure. Furthermore, VEGF-C expression was associated with lymph node recurrence in N0 cases. Multivariate analysis revealed that VEGF-C expression was an exclusively independent factor influencing lymph node metastasis. In terms of the overall 5-year survival rate, there was no significance correlation between the overall 5-year survival rate and expression of VEGF-A, Flt-1 and Flt-4 expression, whereas there was a significant difference between VEGF-C-positive and VEGF-C-negative cases (VEGF-C-positive, 51.7% vs VEGF-C-negative, 94.2%). Furthermore, there was a significant difference between positive and negative expression for both VEGF-A and VEGF-C. Multivariate analysis revealed that lymph node metastasis and VEGF-C expression were exclusive, independent factors influencing the overall survival rate. VEGF-C expression may be a predictive factor of regional lymph node recurrence and prognosis in patients with tongue carcinoma. PMID- 15289891 TI - Immunocytochemical characteristics of human osteosarcoma cell line HS-Os-1: possible implication in apoptotic resistance against irradiation. AB - In our previous study, we examined reactive oxygen species (ROS) formation in T lymphocytes following 5 Gy irradiation. We found that ROS formation occurred immediately after irradiation, continued for several hours, and resulted in oxidative DNA damage. Therefore, the origin of the hyper-radiosensitivity of T lymphocytes seemed to be the high production of ROS in the mitochondrial DNA following irradiation. In the succeeding study, we examined radiation-induced ROS formation, oxidative DNA damage, early apoptotic changes, and mitochondrial membrane dysfunction in the human osteosarcoma cell line HS-Os-1. We found that ROS formation and oxidative DNA damage were actually scarcely seen after irradiation of up to 30 Gy in these cells, that mitochondrial membrane potential was preserved, and that apoptotic changes were not demonstrated despite the relatively high-dose irradiation of 30 Gy. In the present study, we examined the immunocytochemical characteristics of the apoptotic-resistance of the HS-Os-1 cell line against irradiation in order to clarify its possible implications regarding radiosensitivity. The results showed that these cells lack P53 and Bax protein expression, and strong peroxidase activity was confirmed in the nuclei of the cells. Moreover, SODII (manganese superoxide dismutase II) protein expression was gradually increased in spite of irradiation of up to 30 Gy. Therefore, it is concluded that HS-Os-1 cells are originally apoptotic-resistant and that the cells possess a strong ability to scavenge for free radicals. To convert these cells to a state of apoptotic-susceptibility, a powerful oxidant such as hydrogen peroxide might exert such an effect in terms of the production of hydroxyl radicals in lysosomes in the cells as shown in our previous studies. The origin of the radioresistance of the human osteosarcoma cell line HS-Os-1 is considered to to be low degree of ROS formation following irradiation, reflecting the strong scavenging ability of these cells for free radicals including hydroxyl radicals. PMID- 15289892 TI - A combination of selective COX-2 inhibitors and hydrogen peroxide increase the reactive oxygen species formation in osteosarcoma cells after X-ray irradiation. AB - We investigated whether a combination of selective COX-2 inhibitors and hydrogen peroxide increases the effect of X-ray irradiation, with regard to reactive oxygen species (ROS) formation in an osteosarcoma cell line. COX-2 inhibitor did not induce ROS formation when combined with irradiation. A low dose concentration of COX-2 inhibitor in combination with hydrogen peroxide and irradiation did affect ROS formation in the intracellular compartment; however, this same combination of agents at high doses did not modulate the effect of irradiation. Therefore, low doses of COX-2 inhibitor and hydrogen peroxide together, in combination with irradiation, is a potentially useful alternative form of radiotherapy for apoptotic-resistant neoplasms such as osteosarcoma. PMID- 15289893 TI - Transcriptional gene expression profile of human nasopharynx. AB - Inflammation in the nasopharynx is very common worldwide and nasopharyngeal carcinoma (NPC) results in significant morbidity and mortality in southeast Asia and north Africa. To facilitate the understanding of pathogenesis of these diseases as well as normal nasopharynx biology, transcriptional gene expression profile of normal human nasopharynx, from undissected biopsies, was established in this study by generating a large amount of ESTs, followed by bioinformatics analysis. A total of 27,209 ESTs generated from human nasopharynx cDNA library were integrated into 8,420 non-redundant clusters, of which, 6,070 (72.09%) corresponded to known genes, 156 (1.85%) to known ESTs, and 2,194 (26.06%) to novel sequences, respectively. Functional classification revealed that transcripts constituting enzymes (2,284, 28.30%) and those participating in cell growth/maintenance (3,306, 46.33%) were the largest population expressed in the nasopharynx, followed by genes coding for binding proteins (2,135, 26.45%) and involved in cell communication process (1,832, 25.68%). In addition, through comparison of the nasopharynx gene expression profile with those of 7 other human tissues, 59 transcripts preferentially expressed and 22 transcripts unique in the nasopharynx were identified. Our study acquired an initial assessment of qualitative diversity of gene expression in the nasopharynx, providing the first step toward comprehensive characterization of the molecular features of the nasopharyngeal disorders. PMID- 15289894 TI - Possible association of interleukin 1B C-31T polymorphism among Helicobacter pylori seropositive Japanese Brazilians with susceptibility to atrophic gastritis. AB - Associations between anti-Helicobacter pylori seropositivity (HP+) and interleukin 1B (IL-1B) C-31T polymorphism have been reported and little is known about the host factors involved in the development of atrophic gastritis (AG) among infected individuals. This study aimed to examine the IL-1B C-31T polymorphism among anti-HP antibody seropositive Japanese descendants with AG in Brazil and to investigate the interactions with lifestyle factors. Subjects were 455 seropositive from four cities in Brazil, aged 33-69 years. Sex-age-adjusted odds ratio (OR) of AG for 61 current smokers was 1.29, 95% confidence interval (CI): 0.18-9.26 for T/T, while that for 325 never smokers was 0.52, 95% CI: 0.27 0.98. A negative association for AG and never alcohol drinking, every day fruit consumption and milk drinking, and less frequently coffee drinking in individuals with C/T or T/T genotype was observed. This study suggested that host genetic constitution and lifestyle factors may influence the protective effect for AG development. PMID- 15289895 TI - Characterization of protein tyrosine phosphatase activity in rat liver microsomes: suppressive effect of endogenous regucalcin in transgenic rats. AB - The role of regucalcin, a regulatory protein in intracellular signaling system, in the regulation of protein phosphatase activity in rat liver microsomes was investigated. Protein phosphatase activity torward phosphotyrosine, phosphoserine, and phosphothreonine was assayed in a reaction mixture containing the microsomal protein. Protein phosphatase activity toward phosphotyrosine was strong as compared with that of the enzyme activity toward phosphoserine and phosphothreonine, indicating the existence of protein tyrosine phosphatase. Protein phosphatase activity toward three phosphoaminoacids was significantly enhanced by the addition of both calcium chloride (10 micro M) and calmodulin (2.5 or 5 micro g/ml) in the reaction mixture. The presence of ethylene glycol bis (2-amino-ethylether) N, N, N', N'-tetracetic acid (EGTA; 0.1, 1 or 2 mM) or trifluoperazine (TFP; 10, 20 or 50 micro M), an antagonist of calmodulin, did not have a significant effect on protein phosphatase activity toward phosphotyrosine without calcium addition. Microsomal protein tyrosine phosphatase activity was not changed by okadaic acid (10(-6)-10(-4) M). The enzyme activity was significantly decreased by vanadate (10, 50 or 100 micro M). The addition of regucalcin (0.25 or 0.5 micro M) in the reaction mixture caused a significant inhibition of protein tyrosine phosphatase activity in liver microsomes. Western blot analysis showed a remarkable increase in regucalcin protein level in the liver microsomes of regucalcin transgenic (TG) rats. Protein tyrosine phosphatase activity was significantly suppressed in the liver microsomes of TG rats. This study demonstrates that protein tyrosine phosphatase activity is found in the liver microsomes, and that the enzyme activity is suppressed by regucalcin. PMID- 15289896 TI - Blood antioxidant parameters in patients with diabetic retinopathy. AB - It has been postulated that enhanced generation of reactive oxygen species (ROS) may take part in a pathogenesis of diabetic microvascular complication - retinopathy. There are two types of diabetic retinopathy, non-proliferative (simplex) and proliferative. ROS are anihilated by an intracelluar enzymatic system composed mainly of glutathione peroxidase (GPx), superoxide dismutase (SOD) and catalase (CAT). Beta-carotene, tocopherols and ascorbic acid are major components of serum antioxidants. All serum antioxidants are usually measured together as total antioxidant status (TAS). Erythrocyte activities of GPx, SOD, CAT and TAS were measured in diabetic patients without retinopathy, with non proliferative and proliferative retinopathy. Obtained results were correlated with a period of diabetic history and a period of insulin treatment. SOD was significantly elevated in diabetics with non-proliferative retinopathy compared to patients without retinopathy. TAS was significantly lower in patients with proliferative retinopathy than in diabetics not developing retinopathy. Only CAT was significantly negatively correlated with the period of insulin treatment. This significant negative correlation was also observed in a subgroup of patients with proliferative retinopathy. PMID- 15289897 TI - Relatively common mutations of the Bloom syndrome gene in the Japanese population. AB - Bloom syndrome (BS) is a rare autosomal recessive genetic disorder characterized by lupus-like erythematous facial telangiectasia, sun sensitivity, infertility, stunted growth and a high predisposition to various types of cancer. Chromosomal abnormalities are hallmarks of this disorder, and high frequencies of sister chromatid exchanges and quadriradial configurations in lymphocytes and fibroblasts are diagnostic features. BLM is the causative gene for BS. We investigated the mutation in the BLM gene in 4 Japanese BS kindreds. Taken together with previously documented mutations, 2 kindreds were homozygous for 631delCAA and 2 were compound heterozygous for 631delCAA. The silent mutation of A1055C (Thr to Thr) was detected in control Japanese individuals. The 6-bp deletion/7-bp insertion at position 2,281, which most Askenazi Jewish BS patients carry, was not detected in 200 Japanese alleles. These results suggest that 631delCAA is a relatively common mutation among the Japanese BS patients. PMID- 15289898 TI - Nitric oxide as a possible mechanism for understanding the therapeutic effects of osteopathic manipulative medicine (Review). AB - Throughout the history of medicine we have seen the progression of medical therapies from the empirical to the counter-intuitive, with much pressure being placed upon the scientific community to distinguish the two. This exercise has proven the effectiveness of numerous modern therapeutic techniques that have been adapted into modern medicine with remarkable success. While it is certain that many of these techniques yield beneficial results, the mechanisms by which these results are achieved have not been fully realized. In the present report, we consider the case of osteopathic manipulative medicine (OMM), which represents a therapeutic technique developed over a century ago as a means of non-invasive treatment for numerous ailments. Our intention is to use current findings from our laboratory, as well as those of our colleagues in the area of nitric oxide (NO) research to explain the mechanism through which osteopathic manipulations aid the patient. These reports demonstrate that fluidic motions applied to vascular and nerve tissue in a manner comparable to manipulations can cause a remarkable increase in NO concentration within the blood and vasculature. These findings combined with the overwhelming amount of research into the beneficial effects of constitutive NO provide a dynamic theoretical framework to explain the therapeutic effects of OMM. PMID- 15289900 TI - Beacon[47-73] inhibits glucocorticoid secretion and growth of cultured rat and human adrenocortical cells. AB - Evidence has been recently provided that beacon, an ubiquitin-like protein overexpressed in the hypothalamus of Israeli sand rat, is also expressed in several endocrine glands of the Wistar rat, including adrenal cortex. Moreover, it has been shown that the in vivo administration of beacon[47-73] (hereinafter, beacon) evokes within 60 min a marked decrease in the plasma concentrations of ACTH and corticosterone. Hence, we have investigated the effect of beacon (4x10( 9) or 4x10(-7) M) on the secretion and growth of cultured rat and human zona fasciculata/reticularis (ZF/R) cells. Reverse transcription-polymerase chain reaction detected beacon mRNA in all human adrenal cortexes examined. A 3-h exposure to beacon was ineffective, but prolonged (24 and 96 h) exposures significantly lowered basal corticosterone and cortisol secretion from cultured rat and human ZF/R cells, respectively. Moreover, beacon (4x10(-7) M) counteracted the secretagogue action of 10(-8) M ACTH on cultured cells. The 96-h exposure to beacon concentration-dependently decreased basal proliferation rate of cultured cells, without inducing significant changes in the number of apoptotic and necrotic cells. Beacon (4x10(-7) M) significantly inhibited the proliferogenic effect of 10(-8) M adrenomedullin. In light of the involvement of ubiquitin-like proteins in the control of cell cycle and protein sorting and degradation, the hypothesis is advanced that the inhibitory effect of beacon on the secretion and growth of cultured rat ZF/R cells may be connected to its stimulating effect on proteolysis of steroidogenic enzymes and proteins involved in cell replication. PMID- 15289899 TI - Bone loss in regucalcin transgenic rats: enhancement of osteoclastic cell formation from bone marrow of rats with increasing age. AB - Bone loss was previously shown to be induced in the femoral tissue of regucalcin transgenic (TG) rats. Regucalcin is expressed in rat bone marrow cells and its expression is enhanced in regucalcin TG rats. This study was undertaken to determine the change in osteoclastic bone resorption in regucalcin TG rats with increasing age. Femoral-diaphyseal and -metaphyseal tissues were obtained from normal (wild-type) and regucalcin TG rats aged 5, 14, 25 or 50 weeks. Calcium content in the femoral-diaphyseal and -metaphyseal tissues was significantly decreased in regucalcin TG male and female rats aged 5, 14, 25 or 50 weeks as compared with the value obtained from normal rats with each age. When the marrow cells obtained from normal or regucalcin TG rats were cultured for 7 days, the number of tartrate-resistant acid phosphatase (TRACP), a marker of osteoclasts, positive multinucleated cells (MNCs) were significantly increased in the marrow culture of regucalcin TG male and female rats aged 5, 14, 25 or 50 weeks. The effect of parathyroid hormone [human PTH (1-34); 10(-7) M] or 1,25 dihydroxyvitamin D3 [1,25(OH)2D3; 10(-7) M] in stimulating TRACP-positive MNC formation was significantly enhanced in regucalcin TG male and female rats aged 14 or 25 weeks. This study demonstrates that osteoclastic bone resorption is stimulated in regucalcin TG male and female rats with increasing age. PMID- 15289901 TI - Germ-line mutation of Foxn5 gene in mouse lineage. AB - Amplified region (amplicon) around MLL gene is closely linked to the 11q23.3 commonly deleted region of neuroblastoma, which includes cancer-associated genes such as PHLDB1 (LL5A), BCL9L, FOXN5 (FOXR1), CBL, MFRP, and PVRL1 (Nectin) genes. FOXN6 (FOXR2) gene at human chromosome Xp11.21 is generated due to retrotransposition of ancestral Foxn5 gene during evolution. FOXN5 and FOXN6 orthologs share the common domain structure consisting of FN56 and Forhead-box (FOX) domains. Here, we identified and characterized mouse Foxn5 gene by using bioinformatics. Mouse Foxn5, consisting of six exons, was located within mouse genome sequences AC122428.4 and AC125129.5. Foxn5 locus at mouse chromosome 9B was synthenic to rat chromosome 8q22 and human chromosome 11q23.3. Mouse Foxn5 (180 aa) was C-terminally truncated compared with rat Foxn5 and human FOXN5. Mouse 'Foxn5' protein without FOX domain was generated due to a frame shift introduced by germ-line one-base deletion within exon 3. Mouse Foxn5 mRNA was expressed in embryonic germ cells and fertilized eggs. Germ-line mutation of Foxn5 gene in the mouse lineage might lead to divergent scenario of early embryogenesis between mouse and rat through the deregulation of Foxn5 target genes in mouse early embryos, and explain the difficulty in manipulation of rat embryonic stem (ES) cells based on the mouse equivalent system. This is the first report on identification and characterization of mouse Foxn5 gene as well as on species specific germ-line mutation of the Fox family gene. PMID- 15289902 TI - Characterization of FMN2 gene at human chromosome 1q43. AB - Mouse Formin (Fmn1) is an actin regulator interacting with Profilin, SRC, EMS1, FNBP1, FNBP2, FNBP3, FNBP4, WBP4 and alpha-catenin. FMN1, FHOD1, FHOD3, GRID2IP and FHDC1 are non-FDD-type Formin homology proteins, while FMNL1, FMNL2, FMNL3, DIAPH1, DIAPH2, DIAPH3, DAAM1 and DAAM2 are FDD-type Formin homology proteins. Here, we characterized human FMN2 gene by using bioinformatics. Complete coding sequence of human FMN2 cDNA was determined by assembling AL359918, AL513342, AL590490, AL646016 genome sequences, AF218941 partial cDNA, and AF218942 partial cDNA. FMN2 mRNA was expressed in fetal brain, adult whole brain, hypothalamus, retina, pancreatic islet and germinal-center B cells. Among various human tumors, FMN2 mRNA was expressed in parathyloid tumor, glioblastoma, retinoblastoma and chondrosarcoma. Human FMN2 (1722 aa) showed 74.7% total-amino-acid identity with mouse Fmn2, and 31.9% total-amino-acid identity with human FMN1. Although N terminal half was divergent between FMN2 orthologs and FMN1 orthologs, FH1 and FH2 domains were conserved among FMN2 and FMN1 orthologs. Exon-intron structure was conserved between FMN2 and FMN1 genes. RYR2-FMN2-CKTSF1B2 (PRDC) locus at human chromosome 1q43 and RYR3-FMN1-CKTSF1B1 (Gremlin) locus at human chromosome 15q13-q14 were paralogous regions (paralogons) within the human genome. This is the first report on comprehensive characterization of the human FMN2 gene. PMID- 15289903 TI - Evaluation of Fas gene promoter polymorphism in cervical cancer patients. AB - Transduction of signalling through Fas receptor has been implicated in physiological regulation of apoptosis process as well as pathogenesis of various human diseases. The gene encoding Fas receptor contains single nucleotide polymorphism at -670 position, which influences the expression by different transcriptional efficiency of this gene. The aim of this study was to determine the distribution of -670 A/G Fas gene promoter polymorphism in cervical cancer patients and healthy control group in Poland in order to evaluate the potential association between Fas genotype and cervical carcinogenesis. Our results do not confirm the hypothesis that AA genotype in Fas gene promoter may be engaged in the development of cervical neoplasia. PMID- 15289906 TI - [Cost effectiveness of intraoperative three-dimensional imaging with a mobile surgical C-arm]. AB - DRGs lead to a new definition of refunding medical treatment. The calculated DRG covers all costs of the individual hospital stay of each patient. Any revision surgery is not refunded separately. Especially in spine surgery and in articular fractures, postoperative X-ray control sometime reveals malposition of implants that later require operative revision. The latter causes additional costs that can be in the range of 50% of the whole refund as shown here for tibia plateau and calcaneus fractures. New intraoperative 3D fluoro techniques provide CT-like images of bones and joints that enable the surgeon to optimize implant positions immediately. Despite higher initial costs as compared to conventional imaging devices, these technologies help to increase process quality in the OR and clearly save costs. PMID- 15289912 TI - [Acute airway obstruction in acquired hemophilia A]. AB - We report the case of a 67 year old patient suffering from acute airway obstruction caused by hemorrhage localized to the tongue, mouth cavity and hypopharynx, with no evidence of bleeding in his history. The patient presented initially with a globus feeling of the neck, dysphagia and a sore throat. CT scan revealed a swelling of the lingual and sublingual areas and the pharyngeal wall. Next day, there was an immediate life-threatening event caused by progressive bleeding with airway obstruction and an inability to intubate requiring coniotomy. The etiology of the hemorrhage was confirmed by finding a depletion of factor VIII and the presence of auto-antibody directed against this factor. Based on this case report and a review of the literature, we discuss the diagnosis and treatment of acquired hemophilia. PMID- 15289920 TI - [Special legal features of treatment of minors]. AB - According to current jurisdiction any intervention of the physical integrity of the body, even medical treatment and the administration of medication, constitutes a physical injury. The legal authority for this primarily comes from the consent of the patient. The problem for the medical doctor is whose consent is necessary when the patient is a minor. According to the jurisdiction and the literature, the consent of a minor to medical treatment is a legally binding decision not dependent on reaching the age of majority. Nevertheless, according to the overwhelming opinion it can be assumed that minors under the age of 14 years old are not yet capable of consent. The authority for consent lies with the parents or guardians. Because children are normally only accompanied by one of the parents when visiting a doctor, in routine practice the 3-stage theory developed by the Federal High Court pays a deciding role. According to this theory, for routine cases the doctor can assume that one of the parents is basically a representative of the other: for more complicated or severe cases the doctor must satisfy himself that this is true. For patients over 14 years old, the doctor must ascertain whether the patient is capable of consent. Even though a 15-year-old patient for example, can possess the necessary power of judgement and therefore be considered capable of consenting to routine measures or simple interventions, such as taking a blood sample, the yardstick for higher-risk operations, even routine ones is much higher. PMID- 15289921 TI - On the three-dimensional physiological position of the temporomandibular joint. AB - Despite intensive research regarding the position of the temporomandibular joint, only few evidence-based facts are known about a three-dimensional physiological condyle-fossa relationship. The aim of this systematic literature review was therefore to summarize the existing literature regarding the condyle position and to draw conclusions about a standardized three-dimensional condyle position. An extended search profile based on the search strategy of the Cochrane Oral Health Group and applied to twelve medical databases was used to identify existing studies. Articles which were not electronically accessible (mainly those published before 1960) were identified from reference lists and bibliographies. These publications were assessed independently by two clinicians with reference to points based on specified criteria. The interexaminer agreement was assessed on the basis of the kappa coefficient.1903 articles published in the period 1899 2001 and relating to condyle position were identified. These studies showed a pronounced variability in their methodological approaches, with only 49 of them meeting the inclusion criteria. The interexaminer agreement yielded a kappa-value of 0.92. Although numerous studies used three-dimensional data, no physiological three-dimensional condyle position was determined. The most frequent analytic method was two-dimensional projection of the temporomandibular joint onto a subjectively selected sagittal plane. This evaluation method revealed a noticeable shift over time in condyle position from posterior to anterior, suggesting a clear-cut publication bias. Publications with the highest evidence level favored no specific position (p > 0.05). Simplification of the three dimensional structure of the temporomandibular joint to a two-dimensional projection is questionable for therapeutic positioning of the condyle in relation to the fossa. PMID- 15289922 TI - Sagittal and vertical growth of the jaws in Class II, Division 1 and Class II, Division 2 malocclusions during prepubertal and pubertal development. AB - AIM: The aim of the present study was to evaluate the sagittal and vertical development of the jaws in Class II, Division 1 (II/1) and Class II, Division 2 (II/2) malocclusions. In addition, facial morphology was to be investigated in probands with these malocclusions. PROBANDS AND METHODS: Maxillary and mandibular development was investigated with reference to lateral cephalograms of orthodontically untreated probands from the Belfast Growth Study at 7, 9, 11, 13 and 15 years of age. Moreover, development of facial width was assessed from the associated posteroanterior cephalograms, with radiographic magnifications being corrected in both the lateral and the posteroanterior cephalograms. A Class II/1 group (n = 17) and a Class II/2 group (n = 12) were compared with two control groups: a group with good occlusion (n = 18) and a Class I group (n = 37). RESULTS AND CONCLUSIONS: With respect to the sagittal position of the maxilla, no significant differences between the Class II groups and the controls were found. In the Class II/1 group, mandibular retrognathism was observed. The posterior position of the mandible present at 15 years of age had been present even at 7 years of age, and growth increments in the Class I and Class II/1 subjects were similar. In the Class II/2 groups no uniform pattern with respect to mandibular position was found. With respect to vertical development, a deficit in lower anterior facial height was found in the Class II/2 groups. In addition, between 7 and 15 years of age, growth increments in lower anterior facial height were significantly smaller in the Class II/2 subjects than in the controls. Furthermore, the Class II/2 groups displayed a more euryprosopic facial form on average. The cause of this characteristic facial morphology was the vertical deficit in lower anterior facial height. Overall, however, the broad variability and the small sample sizes, in particular of the Class II/2 groups, in the present study have to be seen as limitations. PMID- 15289923 TI - Load-deflection behavior of transpalatal bars supported on orthodontic palatal implants. An in vitro study. AB - Within the scope of indirect orthodontic implant anchorage, the transpalatal bar (TPB) transmits the forces applied at the anchorage teeth to an implant inserted into the palate. It is thus the connecting element between teeth and palatal implant. The aim of the present study was to analyze the extent of sagittal deflection of TPBs of different dimensions attached to an implant abutment. For this purpose, commercially available stainless steel wires measuring 1.2 mm x 1.2 mm (n = 25) and 0.8 mm x 0.8 mm (n = 25) respectively were bent to comply with a representative bend specified with respect to casts, fixed with clamp caps to the implant abutment, and exposed for forces of 50, 100, 150, 200 and 300 cN. The mean deflection of the 1.2 mm x 1.2 mm TPB was lower by a factor of 4.5 than that of the 0.8 mm x 0.8 mm TPB. With a clinically relevant load of 200 cN, for example, the mean deflection of the 1.2 mm x 1.2 mm (0.051" x 0.051") TPB was 301 microm, whereas that of the 0.8 mm x 0.8 mm (0.032" x 0.032") TPB was 1337 microm. When maximum anchorage is required, 1.2 mm x 1.2 mm (0.051" x 0.051") TPBs should be used as a matter of principle. PMID- 15289924 TI - Stability of the bonded lingual wire retainer-a study of the initial bond strength. AB - Bonded lingual retainers (individually adjusted multistranded wires with one bond site per tooth) are used extensively to maintain the orthodontic treatment result. Failure or loss often leads to a relapse. The bond strength of bonded lingual retainers has not yet been studied in respect of the loads that can be withstood by them through deflection of the interdental archwire region. Furthermore, human anterior teeth have never before been used for a study of this kind. Six different wire/composite combinations were studied (wires: Dentaflex co axial 0.018", Dentaflex multistranded 0.018", and Respond Dead Soft straight, length 0.0175"; composites: Tetric Flow and Heliosit Orthodontic) by bonding 1 cm lengths of wire to the lingual surfaces of 360 extracted lower anterior teeth. Using an Instron 6025 universal testing machine, vertical shear bond strength tests at the bond site as well as vertical shear bond strength tests and horizontal tensile strength tests were performed. The failure characteristics after failure at maximum force were evaluated by light microscopy, scanning electron microscopy, and morphometry. Most failures were observed at the enamel/composite interface. The selected wires displayed no significant differences; Tetric Flow proved to be the most stable resin; and no enamel tear outs were observed. PMID- 15289925 TI - Effect of loading rate on bond strength. AB - The purpose of this study was to examine the effect of loading rate on the bond strength of brackets bonded to enamel. Forty premolars were used and brackets were bonded with a chemically cured or a light-cured adhesive, mounted on a testing machine, and debonded under shear stress at two loading velocities: a standard 1 mm/min, and a fast 200 mm/min which better approximates the actual jaw velocity during chewing. Bond strength results (N) were analyzed with two-way analysis of variance (loading rate, adhesive) (alpha = 0.05). An increased loading rate resulted in decreased bond strength, probably due to the induction of a stiff body response and elimination of the viscoelastic properties of the resin. The results indicated a similar effect on both chemically cured and light cured adhesives, thus emphasizing the need for standardized test conditions in bond strength protocols. The implication of loads generated during chewing must be considered when estimating the long-term survival of the bond. PMID- 15289926 TI - Forced eruption for preservation of a deeply fractured molar. AB - BACKGROUND: The treatment of deeply fractured and irreparably damaged teeth often confronts the dentist with a complex problem: Besides extraction, re-implantation and surgical crown lengthening, forced eruption is a reasonable differential alternative. HISTORY AND CLINICAL FINDINGS: In the following case a second lower molar was deeply fractured as a result of an internal granuloma. The tooth had undergone endodontic treatment and had been shortened to alveolar bone level, as the patient had refused extraction. THERAPY: A forced eruption with fibrotomy was performed using an unphysiologically high force. The buccal teeth (canine to first molar) served as the anchorage unit. The mechanics consisted of a lever arm activated to 230 cN. The duration of extrusion was 8 weeks. The tooth was subsequently hemisectioned and provided with an abutment, and the two posts were distalized because of root crowding and moved apart from each other. This was followed by an 8-week retention period, after which the finished crown could be inserted. CONCLUSIONS: The technique of forced eruption accompanied by fibrotomy enabled the roots to be elongated by ca. 4 mm without apposition of alveolar bone. This permitted restoration with physiologic gingival conditions, eliminating the need for surgical crown lengthening with marginal ostectomy or tooth extraction. PMID- 15289927 TI - Nanoscale science: a big step towards the Holy Grail of single molecule biochemistry and molecular biology. AB - Light scattering from metal nanoparticles and fluorescence from quantum dots offer distinct advantages over traditional fluorophores when it comes to detection of single molecules in living cells. PMID- 15289928 TI - Triadin: a multi-protein family for which purpose? AB - Triadin is a protein first identified as a member of the muscle calcium release complex, involved in calcium release for muscle contraction. However, its precise function in this complex is still undefined. Recently, triadin has been shown to be a multi-protein family, with different distribution of the various splice variants within the sarcoplasmic reticulum, raising the possibility of multiple functions for this family of polypeptides. Such functions may include involvement in excitation-contraction coupling, in triad targeting, in structural function or in muscle differentiation. The putative role(s) of triadin(s) will be discussed here. PMID- 15289929 TI - Dependence receptors: between life and death. AB - The recently described family of dependence receptors is a new family of functionally related receptors. These proteins have little sequence similarity but display the common feature of inducing two completely opposite intracellular signals depending on ligand availability: in the presence of ligand, these receptors transduce a positive signal leading to survival, differentiation or migration, while in the absence of ligand, the receptors initiate or amplify a negative signal for apoptosis. Thus, cells that express these proteins manifest a state of dependence on their respective ligands. The mechanisms that trigger cell death induction in the absence of ligand are in large part unknown, but typically require cleavage by specific caspases. In this review we will present the proposed mechanisms for cell death induction by these receptors and their potential function in nervous system development and regulation of tumorigenesis. PMID- 15289932 TI - The role of apolipoprotein E in lipid metabolism in the central nervous system. AB - The critical roles of apolipoprotein E (apoE) in regulating plasma lipid and lipoprotein levels have been extensively studied for over 2 decades. However, an understanding of the roles of apoE in the central nervous system (CNS) is less certain. This review will summarize the available experimental results on the role of apoE in CNS lipid homeostasis with respect to its modulation of sulfatide trafficking, alteration of CNS cholesterol homeostasis and apoE-induced changes in phospholipid molecular species in specialized subcellular membrane fractions. The results indicate that apoE mediates sulfatide trafficking and metabolism in the CNS. Moreover, although apoE does not affect the cholesterol mass content or the phospholipid mass levels and composition in the CNS as a whole, apoE modulates cholesterol and phospholipid homeostasis in selective subcellular membrane compartments. Through elucidating the roles of apoE in CNS lipid metabolism, new insights into overall functions of apoE in neurobiology can be accrued ultimately, leading to an increased understanding of CNS lipid metabolism and the identification of novel therapeutic targets for CNS diseases. PMID- 15289931 TI - Structure-based models of cadherin-mediated cell adhesion: the evolution continues. AB - Cadherins are glycoproteins that are responsible for homophilic, Ca2+-dependent cell-cell adhesion and play crucial roles in many cellular adhesion processes ranging from embryogenesis to the formation of neuronal circuits in the central nervous system. Many different experimental approaches have been used to unravel the molecular basis for cadherin-mediated adhesion. In particular, several high resolution structures have provided models for cadherin-cadherin interactions that are illuminative in many respects yet contradictory in others. This review gives an overview of the structural studies of cadherins over the past decade while focusing on recent developments that reconcile some of the earlier findings. PMID- 15289930 TI - Apoptosis regulation in the mammary gland. AB - Epithelial apoptosis has a key role in the development and function of the mammary gland. It is involved with the formation of ducts during puberty and is required to remove excess epithelial cells after lactation so that the gland can be prepared for future pregnancies. Deregulated apoptosis contributes to malignant progression in the genesis of breast cancer. Since epithelial cell apoptosis in the lactating mammary gland can be synchronised by forced weaning, it has been possible to undertake biochemical analysis of the pathways involved. Together with the targeted overexpression or deletion of candidate genes, these approaches have provided a unique insight into the complex mechanisms of apoptosis regulation in vivo. This review explores what is currently known about the triggers for apoptosis in the normal mammary gland, and how they link with the intrinsic apoptotic machinery. PMID- 15289933 TI - Molecular mechanisms in male determination and germ cell differentiation. AB - Sex determination and gametogenesis are key processes in human reproduction, and any defect can lead to infertility. We describe here the molecular mechanisms of male sex determination and testis formation; defects in sex determination lead to a female phenotype despite the presence of a Y chromosome, more rarely to a male phenotype with XX chromosomes, or to intersex phenotypes. Interestingly, these phenotypes are often associated with other developmental malformations. In testis, spermatozoa are produced from renewable stem cells in a complex differentiation process called spermatogenesis. Gene expression during spermatogenesis differs to a surprising degree from gene expression in somatic cells, and we discuss here mechanistic differences and their effect on the differentiation process and male fertility. PMID- 15289935 TI - Differential chemotactic potential of mouse platelet basic protein for thymocyte subsets. AB - Mouse platelet basic protein (CXCL7/mPBP) was cloned from thymic stromal cells and further identification indicated that it was expressed in thymic monocytes/macrophages (Mo/Mphis). Recombinant mPBP was chemoattractive for target cells of polymorphonuclear leucocytes, peritoneal Mo/Mphis and splenic lymphocytes with distinct potencies. CXCR2 was identified to be a cognate receptor for mPBP. Mouse thymocyte subsets of CD4-CD8- double-negative (DN), CD4+CD8+ double-positive (DP), CD4+CD8- single-positive (CD4SP) and CD4-CD8+ single-positive (CD8SP) expressed cell surface CXCR2 with different positive percentages and expression levels. mPBP was chemoattractive for thymocyte subsets with the potency order DN>DP> CD8SP>CD4SP, consistent with the levels of CXCR2 expressed on the respective cells. Thus, mPBP in thymus is functionally redundant with chemokine CXCL12/ SDF-1. Moreover, our finding that thymic Mo/Mphis can produce mPBP implies that they may have other functions apart from acting as scavengers in thymus. PMID- 15289934 TI - Lactacystin-induced apoptosis of cultured mouse cortical neurons is associated with accumulation of PTEN in the detergent-resistant membrane fraction. AB - The tumor suppressor function of PTEN is attributed to its phospholipid phosphatase activity that dephosphorylates the plasma membrane phosphatidylinositol-(3,4,5)-triphosphate [PtdIns(3,4,5)P3]. Implicit in this notion is that PTEN needs to be targeted to the plasma membrane to dephosphorylate PtdIns(3,4,5)P3. However, the recruitment of PTEN to the plasma membrane is not fully understood. Here, we demonstrate PTEN accumulation in the detergent-insoluble fraction of neuronal cells in response to treatment by the proteasome inhibitor lactacystin. First, lactacystin induces apoptosis and the activation of caspase-3 in cultured cortical neurons. Second, PTEN undergoes proteolysis to form a truncated 50-kDa form that lacks parts of its C-terminal tail. Third, the truncated PTEN is stably associated with the detergent-insoluble fraction in which the plasma membrane marker protein flotillin-1 resides. Taken together, our results suggest that truncation and accumulation of PTEN to the detergent-insoluble membrane fraction are two events associated with the apoptotic signals of the proteasome inhibitor in cortical neurons. PMID- 15289936 TI - The trefoil protein TFF1 is bound to MUC5AC in human gastric mucosa. AB - The trefoil protein TFF1 is expressed principally in the superficial cells of the gastric mucosa. It is a small protein and forms homo- and hetero-dimers via a disulphide bond through Cys58 which is located three amino acids from the C terminus. TFF1 is co-expressed with the secreted mucin MUC5AC in superficial cells of the gastric mucosa suggesting that it could be involved in the packaging or function of gastric mucus. We have previously shown that TFF1 co-sediments with mucin glycoproteins on caesium chloride gradients. To extend this observation we have now used gel filtration under physiological conditions, immunoprecipitation and Western transfer analysis to characterise the interaction of TFF1 with gastric mucin glycoproteins. We show that TFF1 co-elutes with MUC5AC but not MUC6 on gel filtration and that immunoprecipitation and Western transfer analysis confirms that TFF1 interacts with MUC5AC. We also demonstrate that the TFF1 dimer is the predominant molecular form bound to MUC5AC. Salt and chelators of divalent cations such as EDTA and EGTA disrupted the TFF1- MUC5AC interaction and increased the degradation of MUC5AC, whereas calcium increased the amount of TFF1 bound to MUC5AC. These data support the contention that TFF1 is pivotal in the packaging and function of human gastric mucosa. PMID- 15289937 TI - TRAIL promotes the survival, migration and proliferation of vascular smooth muscle cells. AB - Human and rat primary sub-cultured vascular smooth muscle cells (VSMCs) showed clear expression of the death receptors TRAIL-R1 and TRAIL-R2; however, recombinant soluble TRAIL did not induce cell death when added to these cells. TRAIL tended to protect rat VSMCs from apoptosis induced either by inflammatory cytokines tumor necrosis factor-alpha + interleukin-1beta + interferon-gamma or by prolonged serum withdrawal, and promoted a significant increase in VSMC proliferation and migration. Of note, all the biological effects induced by TRAIL were significantly inhibited by pharmacological inhibitors of the ERK pathway. Western blot analysis consistently showed that TRAIL induced a significant activation of ERK1/2, and a much weaker phosphorylation of Akt, while it did not affect the p38/MAPK pathway. Taken together, these data strengthen the notion that the TRAIL/TRAIL-R system likely plays a role in the biology of the vascular system by affecting the survival, migration and proliferation of VSMCs. PMID- 15289938 TI - Interaction of volkensin with HeLa cells: binding, uptake, intracellular localization, degradation and exocytosis. AB - Among two-chain ribosome-inactivating proteins (RIPs), volkensin is the most toxic to cells and animals, and is retrogradely axonally transported in the rat central nervous system, being an effective suicide transport agent. Here we studied the binding, endocytosis, intracellular routeing, degradation and exocytosis of this RIP. The interaction of volkensin with HeLa cells was compared to that of nigrin b, as an example of a type 2 RIP with low toxicity, and of ricin, as a reference toxin. Nigrin b and volkensin bound to cells with comparable affinity (approx. 10(-10) M) and had a similar number of binding sites (2 x 10(5)/cell), two-log lower than that reported for ricin. The cellular uptake of volkensin was lower than that reported for nigrin b and ricin. Confocal microscopy showed the rapid localization of volkensin in the Golgi stacks with a perinuclear localization similar to that of ricin, while nigrin b was distributed between cytoplasmic dots and the Golgi compartment. Consistently, brefeldin A, which disrupts the Golgi apparatus, protected cells from the inhibition of protein synthesis by volkensin or ricin, whereas it was ineffective in the case of nigrin b. Of the cell-released RIPs, 57% of volkensin and only 5% of ricin were active, whilst exocytosed nigrin b was totally inactive. Despite the low binding to, and uptake by, cells, the high cytotoxicity of volkensin may depend on (i) routeing to the Golgi apparatus, (ii) the low level of degradation, (iii) rapid recycling and (iv) the high percentage of active toxin remaining after exocytosis. PMID- 15289939 TI - Interaction of metastasis-inducing S100A4 protein in vivo by fluorescence lifetime imaging microscopy. AB - Elevated levels of the calcium-binding regulatory protein, S100A4, have been shown to be causative of a metastatic phenotype in models of cancer metastasis and to be associated with reduced patient survival in breast cancer patients. Recombinant S100A4 protein interacts in vitro in a calcium-dependent manner with the heavy chain of non-muscle myosin isoform A at a protein kinase C phosphorylation site. At present, the mechanism of metastasis induction by S100A4 in vivo is almost completely unknown. The binding of S100A4 to a C-terminal recombinant fragment of non-muscle myosin heavy chain in living HeLa cells has now been shown using confocal microscopy, fluorescence lifetime imaging microscopy and time-correlated single-photon counting. The association between S100A4 and non-muscle myosin heavy chain was studied by determining fluorescence resonance energy transfer-derived changes in the fluorescence lifetime of enhanced cyan fluorescent protein fused to S100A4 in the presence of a recombinant fragment of the C-terminal region of non-muscle myosin heavy chain (rNMMHCIIA) fused to enhanced yellow fluorescent protein. There was no interaction between the non-muscle myosin heavy chain fragment and a calcium binding-deficient mutant of S100A4 protein which has been shown to be defective in the induction of metastasis in model systems in vivo. The results demonstrate, for the first time, not only direct interaction between S100A4 and a target rNMMHCIIA in live mammalian cells, but also that the interaction between S100A4 and the non-muscle myosin heavy chain in vivo could contribute to the mechanism of metastasis induction by a high level of S100A4 protein. PMID- 15289942 TI - Langerhans' cell histiocytosis: pathology, imaging and treatment of skeletal involvement. AB - Langerhans' cell histiocytosis (LCH) is manifested in a variety of ways, the most common being the eosinophilic granuloma, a localized, often solitary bone lesion that occurs predominantly in the pediatric age group. The hallmark of LCH is the proliferation and accumulation of a specific histiocyte: the Langerhans' cell. In bone this may cause pain and adjacent soft-tissue swelling, but some lesions are asymptomatic. LCH can involve any bone, but most lesions occur in the skull (especially the calvarium and temporal bones), the pelvis, spine, mandible, ribs, and tubular bones. Imaging diagnosis of the disease in bone is first based on the plain radiographic appearance, which is usually a central destructive, aggressive looking lesion. In the skull, the lesions develop in the diploic space, are lytic, and their edges may be beveled, scalloped or confluent (geographic), or show a "button sequestrum." Vertebral body involvement usually causes collapse, resulting in vertebra plana. With significant recent improvements in the quality of gamma cameras, imaging techniques, and in studying children, bone scintigraphy at diagnosis and on follow-up usually reveals the sites of active disease, especially when the involvement is polyostotic. CT and MR imaging are very useful in providing detailed cross-sectional anatomic detail of the involved bone, including the bone marrow and the adjacent soft tissues. CT is better suited for demonstrating bone detail and MR imaging for bone marrow and soft-tissue involvement. PMID- 15289943 TI - Megacystis-microcolon-intestinal hypoperistalsis and prune belly: overlapping syndromes. AB - Megacystis-microcolon-intestinal hypoperistalsis syndrome (MMIHS) is a rare, often fatal condition. Infants present with a functional obstruction of the gastrointestinal tract (GI), malrotation, microcolon, and a large nonobstructed bladder. Several features common to both MMIHS and Eagle-Barrett or prune belly syndrome (PBS) include hydronephrosis, bladder distension and laxity of the abdominal wall musculature. Additionally, MMIHS and PBS have been reported in the same family, suggesting the possibility of a common pathogenesis. MMIHS usually presents in female infants. We present a male infant diagnosed with both MMIHS and PBS. This is a unique case in which both MMIHS and true PBS are present in the same infant. PMID- 15289951 TI - Endodermal sinus tumour of the omentum in a child. AB - Endodermal sinus tumour usually arises in a gonad; extragonadal endodermal sinus tumours are rare. We report a 3-year-old boy with an endodermal sinus tumour arising in the greater omentum, which may be the second reported case in the English literature. He presented with a solid mass in the upper abdomen and a markedly raised serum alpha-fetoprotein (AFP) level. Following percutaneous needle biopsy and omentectomy, histological examination revealed classic morphological features of an endodermal sinus tumour. This rare diagnosis is possible from the imaging features in association with a raised serum level of AFP. PMID- 15289955 TI - Cerebellar pleomorphic xanthoastrocytoma in a patient with neurofibromatosis type 1. AB - We describe a case of cerebellar pleomorphic xanthoastrocytoma (PXA) occurring in a patient with neurofibromatosis type 1 (NF1). The histomorphology of this uncommon glial (astrocytic) neoplasm is discussed. The occurrence of this tumor within the posterior fossa is extremely rare. To our knowledge, this is the first reported case of a cerebellar PXA in a patient with NF1. PMID- 15289956 TI - Intrathecal gadolinium (gadopentetate dimeglumine)-enhanced MR cisternography used to determine potential communication between the cerebrospinal fluid pathways and intracranial arachnoid cysts. AB - This study was designed to assess the feasibility of intrathecal gadolinium enhanced magnetic resonance cisternography (MRC) for the evaluation of the presence or absence of communication of cranial arachnoid cysts with the cerebrospinal fluid (CSF) pathways of the central nervous system (CNS). This prospective study included 20 patients (12 males and 8 females) with a mean age of 37 years, who had, as a group, 22 intracranial arachnoid cysts identified on prior CT and/or MR examinations. Routine pre-contrast cranial MR imaging was followed by the intrathecal administration of 0.5 cc gadopentetate dimeglumine (GD) (Magnevist, Schering, Germany). Immediate and delayed (24 h) MR cisternography was then carried out. Eleven of 22 arachnoid cysts showed immediate CSF communication by the demonstration GD-contrast enhancement of the cyst fluid on the immediate post-injection scan. Contrast enhancement of the cyst was observed only on delayed MRC in six patients. MR imaging in five patients demonstrated no contrast enhancement of the arachnoid cysts on either immediate or delayed imaging. Six patients had mild transient post-procedure headache that was relieved by oral analgesics within 24 h. No serious immediate or chronic adverse effects or complications relating to the intrathecal contrast administration were observed. This study showed the relative safety, feasibility and reliability of low-dose intrathecal GD MR imaging in the demonstration of the communication or non-communication of intracranial arachnoid cysts with the CSF pathways of the CNS. In the future, this may have bearing on the selection for surgery of patients with intracranial arachnoid cysts presenting with clinical signs and symptoms potentially related to the location and mass effect of the cyst. PMID- 15289957 TI - Magnetic resonance imaging findings in Kimura's disease. AB - Although early diagnosis of Kimura's disease, a rare chronic inflammatory disorder most commonly presenting with asymmetric swelling in the head and neck region, is helpful in avoiding unnecessary diagnostic tests and starting prompt treatment, only a few reports emphasized radiological findings in detail. Magnetic resonance imaging findings showing the infiltrative nature of the disease and diffuse loss of fat tissue even in nonpalpable normally appearing regions of the head and neck in a young man with Kimura's disease are presented in this report. PMID- 15289958 TI - Extracranial-intracranial arterial bypass surgery improves total brain blood supply in selected symptomatic patients with unilateral internal carotid artery occlusion and insufficient collateralization. AB - It remains controversial whether extracranial-intracranial (EC-IC) arterial bypass surgery leads to a significant increase in brain blood supply, allowing the reversal of regional cerebral hypoperfusion in symptomatic patients with occlusive cerebrovascular disease and hemodynamic impairment. The aim of the present study was to determine the effects of EC-IC bypass surgery on cerebral brain-supplying blood volume flow (BVF; ml/min) from a purely hemodynamic point of view, using 2D cine phase-contrast MR imaging. Twenty-five patients with symptomatic, unilateral internal carotid artery (ICA) occlusion and hemodynamic compromise received EC-IC arterial bypass surgery. All patients underwent quantitative BVF measurements of brain-supplying arteries preoperatively and postoperatively, including the direct BVF measurement in the established EC-IC bypass after surgery. Preoperatively, total brain BVF was reduced in comparison to normal controls (595 +/- 89 vs 663 +/- 49 ml/min; [mean +/- SEM]; p = 0.039). Mean BVF through the EC-IC bypass reached 84 +/- 32 ml/min (range: 14-177 ml/min), leading to a significant net increase in total BVF of 78 +/- 43 ml/min (range: 7-136 ml/min) when compared with BVF prior to surgery (p < 0.001), with resulting postoperative BVF reaching values obtained in normal controls. EC-IC arterial bypass surgery increases total brain blood supply, allowing restoration of local perfusion in hemodynamically compromised brain tissue in patients with symptomatic ICA occlusion. PMID- 15289959 TI - Population pharmacokinetic analysis of mirtazapine. AB - OBJECTIVE: Mirtazapine belongs to the new generation of antidepressants that is commonly used in clinical routine. Therefore, we feel it mandatory to control compliance in the context of non-response, adverse events or other clinical situations by means of plasma concentration measurements. While controlled clinical studies have evaluated the effect of individual covariates on the pharmacokinetics of mirtazapine, our analysis aims to identify covariates within a naturalistic clinical setting. METHODS: We performed non-linear mixed-effects modelling with data from 65 depressed inpatients whose plasma concentrations were measured weekly during their stay in hospital. Each patient's age, height, weight, co-medication, alcohol, coffee and cigarette consumption, weekly serum creatinine concentrations, liver enzyme activity, blood pressure and pulse was noted. From 49 patients, the genotype of cytochrome P450 (CYP) isoenzymes 2D6, 2C9 and 2C19 was analysed. RESULTS: The clearance of CYP2D6 intermediate metabolisers was reduced by 26% compared with extensive metabolisers. No other factor significantly influenced the clearance of these patients. CONCLUSION: The variability of mirtazapine plasma concentrations in clinical routine is caused to a relevant degree by CYP2D6. This should be taken into account when therapeutic drug monitoring is carried out to check treatment adherence or when a special clinical situation, such as co-morbidity and add-on medication, demands careful dosing of this drug. PMID- 15289960 TI - Ursodeoxycholic acid does not affect ethinylestradiol bioavailability in women taking oral contraceptives. AB - OBJECTIVE: Contraception is recommended for female patients during ursodeoxycholic acid (UDCA) treatment for the potential teratogenic effect of this bile acid, and the aim of our study was to determine whether this treatment affects the bioavailability of ethinylestradiol (EE2). METHODS: In this double blind, randomised study, we measured EE2 pharmacokinetics in eight healthy volunteers randomly allocated to receive oral contraceptive (30 microg EE2 and 75 microg gestodene) plus either UDCA (8-10 mg/kg per day) or placebo for 21 days during the first of three consecutive menstrual cycles. After a washout period during the second cycle, the subjects received the alternative treatment during the third menstrual cycle. Serum EE2 and UDCA were measured using radioimmunoassay and gas chromatography-mass spectrometry, respectively. RESULTS: The profile for serum EE2 concentration was similar during UDCA (mean maximum serum concentration 177 pg/ml, SEM 59) and during placebo treatment (153 pg/ml, SEM 62), and mean area under the curve (AUC) was 1374 pg/h per ml (SEM 580) and 1320 pg/h per ml (SEM 551) during the two regimens, respectively. The point estimates and 90% confidence intervals of UDCA/placebo ratios for EE2 AUC and for maximum serum concentration were 1.1 (0.8-1.5) and 1.2 (1.0-1.4), respectively. Mean serum triglycerides concentration increased from 58.3 mg/dl (SEM 6.8) at enrolment to 91.4 mg/dl (SEM 10.7) during placebo (P < 0.01) and to 88.6 mg/dl (SEM 13.7) during UDCA treatment (P < 0.05). During UDCA treatment, serum enrichment with this bile acid and with the metabolite iso-UDCA was 29% (16%) and 3% (2%), respectively. CONCLUSION: Co-administration with UDCA does not affect the bioavailability of EE2 in healthy volunteers, indicating that contraceptive efficacy is not affected. PMID- 15289962 TI - Action-induced blindness with lateralized stimuli and responses. AB - Previous dual-task studies showed that the selection and/or execution of a response interfere with concurrent visual encoding (action-induced blindness). Four experiments examined how the lateralization of stimuli and responses might affect action-induced blindness. Participants responded to tones (S1) by pressing keys with the left or right hand (R1), and simultaneously identified stimuli (S2) presented to the left or right visual field. Results revealed a complex pattern of cross-talk effects between response preparation and visual encoding. Firstly, preparing a response generally impaired concurrent visual encoding. Secondly, action-induced blindness was equally present for ipsilaterally and contralaterally presented stimuli. Thirdly, response preparation facilitated processing of visual stimuli at ipsilateral locations, probably a case of action centered attention. Finally, the facilitatory effect of R1-S2 correspondence on visual encoding was complemented by a S2-R1 correspondence effect on response execution. Thus, acting while seeing can have both beneficial and detrimental effects on identification performance at the same time. PMID- 15289963 TI - Longer storage of auditory than of visual information in the rabbit brain: evidence from dorsal hippocampal electrophysiology. AB - Whereas sensory memory in humans has been found to store auditory information for a longer time than visual information, it is unclear whether this is the case also in other species. We recorded hippocampal event-related potentials (ERPs) in awake rabbits exposed to occasional changes in a repeated 50-ms acoustic (1000 versus 2000 Hz) and visual (vertical versus horizontal orientation) stimulus. Three intervals (500, 1500, or 3000 ms) between stimulus repetitions were applied. Whereas acoustic changes significantly affected ERPs with the repetition intervals of 500 and 1500 ms, visual changes did so only with the repetition interval of 500 ms. Our finding, thus, suggests a similarity in sensory processing abilities between human and non-human mammals. PMID- 15289964 TI - Relationship between plantar-flexor torque generation and the magnitude of the movement-related potentials. AB - This study investigates whether rate of torque development (RTD) and/or torque amplitude are reflected in the movement-related potentials (MRPs) preceding and accompanying isometric activation of plantar flexor muscles. Subjects were asked to perform six different tasks involving the left ankle joint. The tasks consisted of voluntary isometric plantar flexions at three different RTDs (two fixed rates and a 'ballistic' task) ending at two different torque amplitudes. The main observations from the analysis of the MRPs were: 1) the readiness potentials (RP) demonstrated a statistically significant discrimination between low and high torque amplitudes; 2) the RP, the motor potentials (MP) and the movement-monitoring potentials (MMP) could be statistically differentiated among the different RTDs; and 3) in general the MRPs demonstrated an ipsilateral tendency in relation to the involved limb. The results indicate that RP is a suitable parameter for differentiation between levels of isometric plantar flexion torque and MP and MMP are sensitive to a differentiation between RTDs. The correlation between MRPs and motor tasks involving different rates of torque development and levels of torque suggests that MRPs may comprise a potential solution for programming of intended movements to be executed by systems based on neural rehabilitation technology. PMID- 15289965 TI - Role of capsaicin-sensitive primary afferent inputs from the masseter muscle in the C1 spinal neurons responding to tooth-pulp stimulation in rats. AB - The aim of the present study was to demonstrate the convergence of inputs from masseter muscle (MM) and tooth pulp (TP) onto C1 spinal neurons and to determine whether the afferent fibers express the functional vanilloid receptor (VR1). Extracellular single-unit recordings were made from 61 C1 units responding to TP electrical stimulation with a constant temporal relationship to a digastric electromyogram signal in pentobarbital anesthetized rats. Eighty-four percent of C1 neurons responding to TP stimulation also responded to the ipsilateral MM stimulation. Of these neurons, 61% were considered to be afferent inputs from Adelta-fibers and the remaining units (39%) were C-fibers, based on calculation of the nerve conduction velocity. Intramuscular injection of capsaicin (0.05 and 0.1%) produced a reduction in a MM-induced C1 neuronal activity in a dose dependent manner and this effect was antagonized by pretreatment with an antagonist of VR1, capsazepine. Some of these units were also excited by noxious heat stimulation (> 43 degrees C). The trigeminal root ganglion (TRG) neurons that innervated the MM were retrogradely labeled with Fluorogold (FG) and the small-diameter FG-labeled TRG neurons expressed the immunoreactivity for VR1. After intramuscular mustard oil injection (noxious chemical stimulation), the C1 neuronal activity induced by both touch and pinch stimuli was enhanced and their receptive field sizes were significantly expanded. These changes were reversed within 15-20 min. These results suggest that there may be the convergence of noxious afferents inputs from the MM and TP afferents on the same C1 neurons in rats, and that the afferent fibers expressing the functional VR1 may contribute to the hyperalgesia and/or referred pain associated with temporomandibular joint disorder. PMID- 15289966 TI - Ocular oscillations generated by coupling of brainstem excitatory and inhibitory saccadic burst neurons. AB - The human saccadic system is potentially unstable and may oscillate if the burst neurons, which generate saccades, are not inhibited by omnipause neurons. A previous study showed that combined saccade vergence movements can evoke oscillations in normal subjects. We set out to determine: 1) whether similar oscillations can be recorded during other paradigms associated with inhibition of omnipause neurons; 2) whether lesions of the fastigial nuclei disrupt such oscillations; and 3) whether such oscillations can be reproduced using a model based on the coupling of excitatory and inhibitory burst neurons. We recorded saccadic oscillations during vergence movements, combined saccade-vergence movements, vertical saccades, pure vergence and blinks in three normal subjects, and in a patient with saccadic hypermetria due to a surgical lesion affecting both fastigial nuclei. During combined saccade-vergence, normal subjects and the cerebellar patient developed small-amplitude (0.1 - 0.5 degrees), high-frequency (27-35 Hz), conjugate horizontal saccadic oscillations. Oscillations of a similar amplitude and frequency occurred during blinks, pure vergence and vertical saccades. One normal subject could generate saccadic oscillations voluntarily (approximately 0.7 degrees amplitude, 25 Hz) during sustained convergence. Previous models proposed that high-frequency eye oscillations produced by the saccadic system (saccadic oscillations), occur because of a delay in a negative feedback loop around high-gain, excitatory burst neurons in the brainstem. The feedback included the cerebellar fastigial nuclei. We propose another model that accounts for saccadic oscillations based on 1) coupling of excitatory and inhibitory burst neurons in the brainstem and 2) the hypothesis that burst neurons show post-inhibitory rebound discharge. When omnipause neurons are inhibited (as during saccades, saccade-vergence movements and blinks), this new model simulates oscillations with amplitudes and frequencies comparable to those in normal human subjects. The finding of saccadic oscillations in the cerebellar patient is compatible with the new model but not with the recent models including the fastigial nuclei in the classic negative-feedback loop model. Our model proposes a novel mechanism for generating oscillations in the oculomotor system and perhaps in other motor systems too. PMID- 15289967 TI - Spatial orientation of optokinetic nystagmus and ocular pursuit during orbital space flight. AB - On Earth, eye velocity of horizontal optokinetic nystagmus (OKN) orients to gravito-inertial acceleration (GIA), the sum of linear accelerations acting on the head and body. We determined whether adaptation to micro-gravity altered this orientation and whether ocular pursuit exhibited similar properties. Eye movements of four astronauts were recorded with three-dimensional video oculography. Optokinetic stimuli were stripes moving horizontally, vertically, and obliquely at 30 degrees/s. Ocular pursuit was produced by a spot moving horizontally or vertically at 20 degrees/s. Subjects were either stationary or were centrifuged during OKN with 1 or 0.5 g of interaural or dorsoventral centripetal linear acceleration. Average eye position during OKN (the beating field) moved into the quick-phase direction by 10 degrees during lateral and upward field movement in all conditions. The beating field did not shift up during downward OKN on Earth, but there was a strong upward movement of the beating field (9 degrees) during downward OKN in the absence of gravity; this likely represents an adaptation to the lack of a vertical 1-g bias in-flight. The horizontal OKN velocity axis tilted 9 degrees in the roll plane toward the GIA during interaural centrifugation, both on Earth and in space. During oblique OKN, the velocity vector tilted towards the GIA in the roll plane when there was a disparity between the direction of stripe motion and the GIA, but not when the two were aligned. In contrast, dorsoventral acceleration tilted the horizontal OKN velocity vector 6 degrees in pitch away from the GIA. Roll tilts of the horizontal OKN velocity vector toward the GIA during interaural centrifugation are consistent with the orientation properties of velocity storage, but pitch tilts away from the GIA when centrifuged while supine are not. We speculate that visual suppression during OKN may have caused the velocity vector to tilt away from the GIA during dorsoventral centrifugation. Vertical OKN and ocular pursuit did not exhibit orientation toward the GIA in any condition. Static full-body roll tilts and centrifugation generating an equivalent interaural acceleration produced the same tilts in the horizontal OKN velocity before and after flight. Thus, the magnitude of tilt in OKN velocity was dependent on the magnitude of interaural linear acceleration, rather than the tilt of the GIA with regard to the head. These results favor a 'filter' model of spatial orientation in which orienting eye movements are proportional to the magnitude of low frequency interaural linear acceleration, rather than models that postulate an internal representation of gravity as the basis for spatial orientation. PMID- 15289979 TI - Silica xerogels as a delivery system for the controlled release of different molecular weight heparins. AB - In this work, we investigated a sol-gel derived silica matrix as a delivery system for the prolonged release of different molecular weight heparins, which allows the glycosaminoglicons to retain their whole biological activity. Several xerogels were obtained by embedding different molecular weight heparins into matrices prepared by using different amount of NH4OH as a catalyst during gel formation. Gel synthesis parameters, drug release properties, and xerogels surface area were evaluated. Unfractionated, low and oligo-molecular weight heparins were embedded into xerogels and the effect of the molecular weight on the release kinetics and the retained biological activity has been investigated. The results show that the surface area of the matrix is a determinant parameter affecting drug release kinetics. This structural feature can be modified by varying the catalyst tetraethoxysilane molar ratio used during the matrix synthesis. In most cases release kinetics fitted the Higuchi diffusive model and a lower diffusion rate was observed from silica matrices characterized by a smaller surface area. In the case of matrices with lower surface area, loaded with unfractionated heparin, zero order kinetics was observed. In this paper, we have defined a heparin release silica xerogel system and we have pointed out how modulation of its synthesis parameters allows adjusting the release of heparin according to therapeutic needs. PMID- 15289980 TI - Nuclear magnetic resonance and high-performance liquid chromatographic evaluation of polymer-based stationary phases immobilized on silica. AB - Three poly(ethylene-co-acrylic) acid copolymers (-CH(2)CH(2)-)(x)[CH(2)CH(CO(2)H) ](y) with different chain lengths and mass fractions of acrylic acid were covalently immobilized as stationary phases on silica via two variants of spacer molecules (3-aminopropyltriethoxysilane and 3-glycidoxypropyltrimethoxysilane). Different mobilities of the alkyl chains in the stationary phases were observed using (13)C solid-state NMR spectroscopy. The stationary phases with more rigid trans-ordered alkyl chains had better selectivity for geometric beta-carotene and xanthophyll isomers (provitamin A derivatives). Also, all the separations of the analytes were affected by polar interactions with the chromatographic sorbent. This was further proved by separating more polar cis/trans retinoic acid isomers (vitamin A derivatives). (13)C high-resolution/magic-angle spinning (HR/MAS) NMR measurements of the chromatographic sorbents suspended in the mobile phase confirmed a dependence of molecular shape recognition ability on alkyl chain conformation. PMID- 15289986 TI - Electrochemical vapor generation of selenium species after online photolysis and reduction by UV-irradiation under nano TiO2 photocatalysis and its application to selenium speciation by HPLC coupled with atomic fluorescence spectrometry. AB - An online UV photolysis and UV/TiO2 photocatalysis reduction device (UV-UV/TiO2 PCRD) and an electrochemical vapor generation (ECVG) cell have been used for the first time as an interface between high-performance liquid chromatography (HPLC) and atomic fluorescence spectrometry (AFS) for selenium speciation. The newly designed ECVG cell of approximately 115 microL dead volume consists of a carbon fiber cathode and a platinum loop anode; the atomic hydrogen generated on the cathode was used to reduce selenium to vapor species for AFS determination. The noise was greatly reduced compared with that obtained by use of the UV-UV/TiO2 PCRD-KBH4-acid interface. The detection limits obtained for seleno-DL: -cystine (SeCys), selenite (Se(IV)), seleno-DL: -methionine (SeMet), and selenate (Se(VI)) were 2.1, 2.9, 4.3, and 3.5 ng mL(-1), respectively. The proposed method was successfully applied to the speciation of selenium in water-soluble extracts of garlic shoots cultured with different selenium species. The results obtained suggested that UV-UV/TiO2 PCRD-ECVG should be an effective interface between HPLC and AFS for the speciation of elements amenable to vapor generation, and is superior to methods involving KBH4. PMID- 15289993 TI - Effect of glucose on tobacco craving. Is it mediated by tryptophan and serotonin? AB - RATIONALE: Oral glucose has been shown to decrease tobacco craving in many but not all previous studies. Glucose ingestion may facilitates entry of tryptophan (TRP), the unique source of brain serotonin, into the brain, glucose's action seems to be opposite of rapid TRP depletion. Therefore, the aim was to assess the effect of high doses of oral glucose on tobacco craving, withdrawal symptoms, plasma TRP and blood serotonin concentrations in temporarily abstinent smokers. METHODS: Aspartame 0.6 g/200 ml (A, placebo), glucose 32.5 g/200 ml (G32.5) and 75 g/200 ml water (G75) were administered to 12 healthy smokers after an overnight abstinence in a crossover, double blind study. Tobacco craving (short version of the Tobacco Craving Questionnaire, TCQ), withdrawal symptoms, choice reaction time, affect, blood glucose, plasma insulin, nicotine, cotinine, free and total TRP, and blood serotonin concentrations were assessed during a period of 5 h after administration. RESULTS: Blood glucose and plasma insulin increased after G32.5, G75 and remained unchanged after A. TCQ score increased with A and remained almost unchanged with both doses of glucose (conditionxtime interaction: P=0.023). Total withdrawal score increased differently according to sex and condition (P<0.05). Motor reaction time increased with A and decreased with glucose (P=0.016). The overall decrease in plasma TRP was 0.31+/-17, 0.49+/-0.19 and 1.44+/-0.24 mg/l with A, G32.5 and G75, respectively (P<0.001). Baseline blood serotonin was lower in women (n=5) than in men; it showed a condition by time (P=0.007) and a condition by time by sex interaction (P=0.023). CONCLUSIONS: Glucose attenuates tobacco craving and withdrawal symptoms in temporarily abstinent smokers. This is accompanied by a decrease in plasma TRP and a sex dependent increase in blood serotonin. Further studies assessing the direct effect of glucose on brain serotonin are needed to ascertain whether a glucose induced reduction in craving is associated with an increase in brain serotonin. PMID- 15289996 TI - Prolactin levels in male schizophrenic patients treated with risperidone and haloperidol: a double-blind and randomized study. AB - RATIONALE: There are few data from systematic, double-blind clinical trials that have compared the effect of the typical and the atypical antipsychotics on serum prolactin (PRL) levels in patients with schizophrenia. OBJECTIVES: The goal of this study was to compare the effect of risperidone and haloperidol on serum PRL and investigate the relationship between serum PRL levels and clinical response in patients with schizophrenia. METHODS: Seventy-eight inpatients with a diagnosis of schizophrenia (according to DSM-III-R) were randomly assigned to 12 weeks of treatment with 6 mg/day of risperidone or 20 mg/day of haloperidol after a 2-week washout period, using a randomized, double-blind design. Clinical efficacy was determined using the positive and negative syndrome scale (PANSS). Their serum PRL was assayed by means of radioimmunometric assay (RIA) between pre treatment and post-treatment, and compared with 30 sex-matched and age-matched normal subjects. RESULTS: Both risperidone and haloperidol treatment significantly increased serum PRL levels in drug-free chronic schizophrenia patients (both P<0.001). Hyperprolactinemia induced by risperidone 6 mg/kg was comparable to levels produced by haloperidol 20 mg/day. Considering dose-adjusted serum PRL levels, risperidone treatment induced a significant elevation of PRL levels compared with haloperidol treatment at the haloperidol equivalent (P<0.001). Change in PRL levels at pre-treatment and post-treatment were related to positive symptom improvement seen in the risperidone group (r=0.51, P=0.016), but not in the haloperidol group (P>0.05). Female patients showed both a higher baseline and post-treatment PRL level and a greater increase in PRL than men (all P<0.05). CONCLUSIONS: Risperidone is associated with a robust effect on prolactin secretion in contrast to the conventional antipsychotic haloperidol. Prolactin monitoring during risperidone treatment should be performed. PMID- 15289997 TI - Adherence to nicotine replacement therapy versus quitting smoking among Chinese smokers: a preliminary investigation. AB - RATIONALE: There are over 300 million Chinese smokers, but use of nicotine replacement therapy (NRT) is rare. On the other hand, data on the factors associated with quitting and adherence to NRT use are scarce in the East. OBJECTIVES: To describe adherence and other predictors of quitting smoking at the 12-month follow-up amongst Chinese smokers in Hong Kong. METHODS: Chinese smokers (1186) who attended the Smoking Cessation Health Centre from August 2000 through January 2002 were studied. Trained counsellors provided individual counselling and carried out follow-up interviews. We used structured questionnaires at baseline and at 1, 3 and 12 months and an intention-to-treat approach for analysis. RESULTS: Among those who received NRT (1051/1186), the prevalence of adherence (self-reported NRT use for at least 4 weeks) was 16% (95% confidence interval 14-18%). The 7-day point prevalence quit rate at 12 months (not smoking any cigarette during the past 7 days at the 12 month follow-up) was 27% (95% CI, CI 24-29%). Stepwise logistic regression model showed that adherence to NRT use, a higher income, good perceived health and having more confidence in quitting were significant predictors of quitting. The quit rate in the adherent group (40%) was greater than that of the non-adherent group (25%) (P<0.001). Older age, male, higher education, experience of NRT use, perceiving quitting as more difficult and willingness to pay were significant predictors of adherence. CONCLUSIONS: Clinically significant smoking cessation rates can be achieved among Chinese smokers in a clinic-based smoking cessation service. The NRT adherence was low and low adherence was associated with a lower quit rate. Trials of interventions to improve adherence and increase quit rates are needed. PMID- 15289998 TI - Bupropion effects on aggressiveness and anxiety in OF1 male mice. AB - RATIONALE: Bupropion is an antidepressant drug that is being used to help in giving up smoking. Its behavioral effects have been evaluated in different animal models, although limited information is available regarding its effects on aggressiveness, anxiety and exploratory behavior. OBJECTIVES: Evaluate acute effects of bupropion on locomotor activity, isolation-induced aggression, hole board and elevated plus-maze tests in OF1 male mice. METHODS: In the first experiment, effects of bupropion (2.5, 5, 10, 20 and 40 mg/kg) on locomotion were evaluated. In the second experiment, isolation-induced aggression was assessed in isolated male mice previously classified as short attack latency (SL) and long attack latency (LL). Mice were treated with bupropion or vehicle and confronted with standard opponents for 10 min. In experiments 3 and 4, mice were treated with bupropion or vehicle and 30 min later examined in the plus-maze or in the hole-board apparatus. RESULTS: In the actimeter, bupropion induced a dose dependent increase in locomotion. During agonistic encounters, bupropion (10 mg/kg and 40 mg/kg) increased time devoted to attack in LL mice. In the plus maze, no significant differences were found between bupropion-treated and vehicle treated mice in the percentage of entries or time spent in open arms. In the hole board, the highest dose of bupropion (40 mg/kg) significantly decreased number of head-dips and increased latency to the first head-dip. CONCLUSIONS: During agonistic encounters the two sub-groups of mice (SL and LL) may display differential sensitivity in drug-induced changes on aggressiveness, since bupropion increased attack only in mice with "long attack latency" in the pre screening test. In the plus-maze, this drug does not seem to have specific actions on anxiety and in the hole-board a high dose had similar effects to those induced by anxiogenic drugs. PMID- 15289999 TI - Anxiolytic properties of agomelatine, an antidepressant with melatoninergic and serotonergic properties: role of 5-HT2C receptor blockade. AB - RATIONALE: The novel antidepressant agent, agomelatine, behaves as an agonist at melatonin receptors and as an antagonist at serotonin (5-HT)(2C) receptors. OBJECTIVES: To determine whether, by virtue of its antagonist properties at 5 HT(2C) receptors, agomelatine elicits anxiolytic properties in rats. METHODS: Employing a combined neurochemical and behavioural approach, actions of agomelatine were compared to those of melatonin, the selective 5-HT(2C) receptor antagonist, SB243,213, and the benzodiazepine, clorazepate. RESULTS: In unfamiliar pairs of rats exposed to a novel environment, agomelatine enhanced the time devoted to active social interaction, an action mimicked by clorazepate and by SB243,213. In a Vogel conflict procedure, agomelatine likewise displayed dose dependent anxiolytic activity with a maximal effect comparable to clorazepate, and SB243,213 was similarly active in this procedure. In a plus-maze procedure in which clorazepate significantly enhanced percentage entries into open arms, agomelatine revealed only modest activity and SB243,213 was inactive. Further, like SB243,213, and in contrast to clorazepate, agomelatine did not suppress ultrasonic vocalizations emitted by rats re-exposed to an environment associated with an aversive stimulus. Whereas clorazepate reduced dialysate levels of 5-HT and noradrenaline in hippocampus and frontal cortex of freely moving rats, agomelatine did not affect extracellular levels of 5-HT and elevated those of noradrenaline. SB243,213 acted similarly to agomelatine. Melatonin, which did not modify extracellular levels of 5-HT or noradrenaline, was ineffective in all models of anxiolytic activity. Furthermore, the selective melatonin antagonist, S22153, did not modify anxiolytic properties of agomelatine in either the social interaction or the Vogel Conflict tests. CONCLUSIONS: In contrast to melatonin, and reflecting blockade of 5-HT(2C) receptors, agomelatine is active in several models of anxiolytic properties in rodents. The anxiolytic profile of agomelatine differs from that of benzodiazepines from which it may also be distinguished by its contrasting influence on corticolimbic monoaminergic pathways. PMID- 15290000 TI - Comparative effects of duloxetine and desipramine on sleep EEG in healthy subjects. AB - RATIONALE: Antidepressants are known to modify human sleep patterns. OBJECTIVES: Duloxetine is a new antidepressant with a mechanism of action involving reuptake inhibition of both serotonin (5-HT) and norepinephrine (NE). In this study, the effects of two dosing regimens of duloxetine on sleep electroencephalography (EEG) were investigated at steady-state plasma concentrations in young, healthy, male subjects. METHODS: Placebo (n=12), desipramine (50 mg BID; n=12) and two regimens of duloxetine (80 mg QD, n=6; or 60 mg BID, n=6) were compared in a randomized, double-blind, three-period crossover study, each treatment being administered from day 1 to day 7. Sleep polygraphic recordings took place at baseline (day -1) and day 6 of each period. The Leeds sleep evaluation questionnaire (LSEQ) was also administered on the morning of day 7. RESULTS: Both regimens of duloxetine produced a significant increase in the onset latency of REM sleep as well as a significant mean decrease in total REM sleep duration. Desipramine exhibited comparable effects. When compared to placebo, sleep continuity was significantly reduced with desipramine and duloxetine 60 mg BID whereas a significant improvement was observed with duloxetine 80 mg QD. On the LSEQ, duloxetine 80 mg QD produced a significant improvement in the "getting to sleep" subscale compared to placebo, whereas desipramine demonstrated a significant reduction (worsening) in the "quality of sleep" score versus placebo. CONCLUSIONS: The two dose regimens of duloxetine (80 mg QD and 60 mg BID) produced a REM sleep pattern comparable to that of most antidepressant medications. Duloxetine 80 mg QD appeared to exhibit less impact upon sleep quality than duloxetine 60 mg BID in healthy subjects. PMID- 15290001 TI - Alcohol impairs auditory processing of frequency changes and novel sounds: a combined MEG and EEG study. AB - RATIONALE: Alcohol has been shown to impair involuntary attention studied by event-related potentials using mismatch negativity (MMN) and P3a. OBJECTIVES: However, no studies have investigated whether alcohol affects the magnetic counterparts of N1 (N1m), MMN (MMNm) and P3a (P3am). METHODS: Auditory evoked potentials and magnetic fields elicited by infrequent deviant tones differing in frequency (5% and 20% change) and novel sounds were recorded with whole-head magnetoencephalography (MEG) and electroencephalography (EEG). Stimuli were presented separately to the left and right ear. Eleven right-handed subjects were studied in a double-blind, placebo-controlled (0.8 g/kg ethanol or juice), cross over design. N1m, MMNm, and P3am were calculated from the channel pair at the temporal cortex showing the strongest responses in the hemisphere contralateral to the stimulation. N1, MMN and P3a were analyzed from 12 electrodes at the midline frontocentral area. RESULTS: Alcohol reduced bilaterally N1, N1m, MMN and MMNm amplitudes. P3a amplitudes, but not P3am amplitudes were also significantly decreased. No effects of alcohol on the latencies of N1, MMN and P3a or their magnetic counterparts were observed. CONCLUSIONS: Alcohol impairs the processing of tones, frequency change and novel sounds at different phases of auditory processing similarly in both hemispheres. MEG provides us with additional information unobtainable with EEG about the effects of alcohol on the neural correlates of cognition. PMID- 15290002 TI - Fluvoxamine, a selective serotonin reuptake inhibitor, suppresses tetrahydrobiopterin levels and dopamine as well as serotonin turnover in the mesoprefrontal system of mice. AB - RATIONALE: Tetrahydrobiopterin (BH4) is a coenzyme of tyrosine hydroxylase (TH) and tryptophan hydroxylase (TPH), rate-limiting enzymes of monoamine biosynthesis. According to the monoamine hypothesis of depression, antidepressants will restore the function of the brain monoaminergic system, and BH4 concentration. OBJECTIVE: To investigate the effects of fluvoxamine on BH4 levels and dopamine (DA) and serotonin (5-HT) turnover in the mesoprefrontal system, incorporating two risk factors of depression, social isolation and acute environmental change. METHODS: Male ddY mice (6W) were divided into two housing groups, i.e. group-housing (eight animals per cage; 35 days), and isolation housing (one per cage; 35 days), SC injected with fluvoxamine (20 or 40 mg/kg; days 29-35), and exposed to 20-min novelty stress (day 35). The levels of BH4, DA, homovanilic acid (HVA), 5-HT, and 5-hydroxyindoleacetic acid (5-HIAA) were measured in the prefrontal cortex and midbrain. RESULTS: Under the group-housing condition, novelty stress significantly increased BH4 levels in both regions, and the HVA/DA ratio in the midbrain, whereas it did not change any parameters in either region under the isolation-housing condition. In the prefrontal cortex, fluvoxamine significantly decreased the 5-HIAA/5-HT ratio under the group-housing condition, and BH4 levels and the HVA/DA ratio under the isolation-housing condition. In the midbrain, fluvoxamine significantly decreased all parameters, except for an increasing in the 5-HIAA/5-HT ratio under the isolation-housing condition. CONCLUSION: Isolation-housing suppressed the increase of BH4 levels and DA turnover elicited by novelty stress. Fluvoxamine suppressed BH4 levels, and DA and 5-HT turnover. Fluvoxamine may have altered DA turnover by suppressing BH4 levels. PMID- 15290003 TI - Metamemory without the memory: are people aware of midazolam-induced amnesia? AB - RATIONALE: Midazolam is a benzodiazepine which produces a dense anterograde amnesia, while permitting relatively well-preserved short-term memory, semantic retrieval, and other higher cognitive functions. Given these preserved abilities, we were interested in whether or not participants given midazolam would be aware of this anterograde amnesia. METHOD: In the present experiment, participants were given midazolam in one testing session and a saline placebo in another. Participants provided judgments-of-learning (JOLs) immediately following study of cue-target pairs. During the test phase of the experiment, confidence levels and feeling-of-knowing (FOK) judgments were collected. RESULTS: Although cued recall performance was substantially impaired in the midazolam condition, mean JOLs were unaffected, indicating participants had little insight into their impairment during the study phase. Participants were relatively accurate in confidence levels and FOK judgments in the midazolam condition. CONCLUSION: When studying items under the influence of midazolam, participants are unaware that their memory will be impaired. Implications for clinical practice and pharmacological studies of amnesia are discussed. PMID- 15290004 TI - Atypical antipsychotic profile of flunarizine in animal models. AB - RATIONALE: Flunarizine is known as a calcium channel blocker commonly used in many countries to treat migraine and vertigo. Parkinsonism has been described as one of its side-effects in the elderly, which is in agreement with its recently characterized moderate D2 receptor antagonism. OBJECTIVES: To perform a pre clinical evaluation of flunarizine as a potential antipsychotic. METHODS: We evaluated the action of orally administered flunarizine in mice against hyperlocomotion induced by amphetamine and dizocilpine (MK-801) as pharmacological models of schizophrenia, induction of catalepsy as a measure for extrapyramidal symptoms and impairment induced by dizocilpine on the delayed alternation task for working memory. RESULTS: Flunarizine robustly inhibited hyperlocomotion induced by both amphetamine and dizocilpine at doses that do not reduce spontaneous locomotion (3-30 mg/kg). Mild catalepsy was observed at 30 mg/kg, being more pronounced at 50 mg/kg and 100 mg/kg. Flunarizine (30 mg/kg) improved dizocilpine-induced impairment on the delayed alternation test. CONCLUSIONS: These results suggest a profile comparable to atypical antipsychotics. The low cost, good tolerability and long half-life (over 2 weeks) of flunarizine are possible advantages for its use as an atypical antipsychotic. These results warrant clinical trials with flunarizine for the treatment of schizophrenia. PMID- 15290005 TI - Cannabinoid CB1 receptor-mediated impairment of visuospatial attention in the rat. AB - RATIONALE: CB1 receptors (CB1Rs) mediate many of the psychoactive effects of cannabinoids, and marijuana intoxication can produce neurocognitive deficits with a similarity to those seen in schizophrenia, including impairments of attention. OBJECTIVES: We thus sought to characterize the effects of a CB1R-selective agonist and antagonist on attention in the rat using a lateralized reaction time task (LRT). We hypothesized that CB1R agonists would impair performance and that CB1R antagonists might improve performance. METHODS: Subjects were trained to perform the LRT, a procedure that measured their ability to attend to and detect brief visual target stimuli. After training, we tested the effects of the CB1R agonist WIN55,212-2 (WIN; 0-2.5 mg/kg) or the CB1R antagonist SR141716A (SR; 0 1.0 mg/kg), administered alone or in combination, on visual attention performance using task conditions in which target stimulus salience was varied systematically across trials. RESULTS: The highest dose of WIN reduced correct choices in well trained rats, with impairment greatest at the shortest stimulus durations. The highest dose of WIN also increased omissions and slowed response times. By contrast, SR itself did not produce any measurable effects on performance but was able to prevent the impairment produced by WIN. CONCLUSIONS: These results suggest that CB1Rs mediate the attentional performance impairments caused by acute administration of cannabinoid agonists and begin to unravel the possible contribution of cannabinoid systems to the pathophysiological substrates of cognitive dysfunction in schizophrenia. PMID- 15290006 TI - Bimodal effects of MK-801 on locomotion and stereotypy in C57BL/6 mice. AB - RATIONALE: Systemic injection of the non-competitive NMDA (N-methyl-D-aspartate) receptor antagonist MK-801 (dizocilpine maleate) causes both increased locomotion in rodents and various stereotypic behaviors that are proposed to model certain aspects of schizophrenic symptoms in humans. OBJECTIVES: This study presents a comprehensive characterization of the bimodal effects of MK-801 on locomotion and stereotypy in the C57BL/6 mouse strain, a strain commonly used for genetically modified mice. RESULTS: We found that it is important to analyze both locomotion and stereotypy in parallel, as MK-801-induced stereotypy results in abnormal movements that are recorded as locomotion by automated beam detection systems. Furthermore, it is important to analyze the bimodal effects of MK-801 over an extended time span, rather than the commonly used narrower time window, as at higher doses (e.g., above 0.3 mg/kg) the hyperlocomotion phase develops only after the stereotypic phase subsides. We also observed that the apparent dose response curve is very sensitive to the particular time window chosen for analysis because MK-801 affects both the time course and maximum value of stimulated locomotion. We show that analyzing the absolute peak value of locomotion induced for each animal, rather than group-averaged time courses, provides a measure that is sensitive over a wider range of MK-801 doses. Interestingly, MK-801 even at a very low dose of 0.02 mg/kg suppressed rather than enhanced rearing behavior, differing in this regard from amphetamine. CONCLUSIONS: The non-competitive NMDA receptor antagonist MK-801 induces a complex pattern of behavioral modification in mice with respect to both the time course and the dose-response relationship of behavioral changes. The results of this study provide a foundation and frame of reference for the growing interest in studying MK-801-induced behavior in mice. PMID- 15290007 TI - Diabetes inhibits the DOI-induced head-twitch response in mice. AB - RATIONALE: Clinical studies suggest that the prevalence of psychiatric disorders is higher in diabetic patients than in the general population. It has been reported that central serotonin(2A) (5-HT(2A)) receptors may be involved in the pathogenesis and treatment of psychiatric disorders. OBJECTIVES: We examined the effect of streptozotocin-induced diabetes on the function of central 5-HT(2A) receptors in mice. METHODS: Male ICR mice were rendered diabetic by an injection of streptozotocin (200 mg/kg, i.v.). The experiments were conducted 2 weeks after the injection of streptozotocin. To evaluate the central 5-HT(2A) receptor function, head-twitch responses were measured for 15 min immediately after the treatment with (+/-)-2,5-dimethoxy-4-iodoamphetamine (DOI) (0.1-1 mg/kg, s.c.), a selective 5-HT(2) receptor agonist. RESULTS: Significantly fewer head-twitch responses were induced by DOI in diabetic mice than in non-diabetic mice. The number and affinity of 5-HT(2A) receptors in the mouse frontal cortex were not affected by diabetes. The corticosterone response to DOI (1 mg/kg and 3 mg/kg, s.c.) was not different between non-diabetic and diabetic mice, although the baseline of plasma corticosterone levels was significantly higher in diabetic than in non-diabetic mice. CONCLUSIONS: Our results suggest that a neuronal network that causes head-twitch responses by triggering by the activation of 5 HT(2A) receptors may be altered by type-1 diabetes in mice. PMID- 15290008 TI - The CB receptor agonist WIN 55,212-2 fails to elicit disruption of prepulse inhibition of the startle in Sprague-Dawley rats. AB - RATIONALE: A growing evidentiary body indicates cannabinoid exposure is conducive to cognitive impairment and psychotic phenomena in vulnerable individuals. In this respect, recent studies have displayed controversial results on the ability of cannabinoids to elicit sensorimotor gating alterations and attentional filtering, whose disruption is a distinctive feature of psychosis. OBJECTIVES: The goal of this study was to investigate the effects of acute, subchronic, and chronic treatment with the synthetic CB receptor agonist WIN 55,212-2 (WIN) on prepulse inhibition (PPI) of the acoustic startle reflex (ASR), a powerful paradigm for evaluation of sensorimotor gating. METHODS: Different groups of adult Sprague-Dawley rats were treated with 0.5, 1, and 2 mg/kg WIN (i.p.) acutely, as well as for 7 days and 21 days. All animals underwent testing 40 min after the last treatment and their evaluation was compared with that of animals treated with vehicle. In a separate group, the effects of WIN withdrawal were also analyzed, 24 h after discontinuation of a 21-day treatment. RESULTS: No variation in PPI was detected in any of the test groups when compared with controls, whatever the dosage and the treatment. CONCLUSIONS: These findings suggest WIN does not impair sensorimotor gating in Sprague-Dawley rats and confirm clinical evidence according to which cannabis is an unlikely causative of psychosis among non-vulnerable individuals. Nonetheless, since in other studies the same compound was shown to induce PPI alterations in Wistar rats, our results are also suggestive that genetic differences might be critical for the development of cannabis-induced cognitive disorders. PMID- 15290009 TI - Differential expression of human COMT alleles in brain and lymphoblasts detected by RT-coupled 5' nuclease assay. AB - RATIONALE: A common polymorphism, Val158Met, alters catechol- O-methyltransferase (COMT) enzyme activity and has been linked to psychiatric phenotypes. Bray et al. (2003) reported that COMT is subject to differential allele expression in brain, finding modest (13-22%) underexpression of a haplotype containing Val158. However, disparate findings by another group who used the same method, but in lymphoblasts, raise the issues of tissue specificity, magnitude of differential expression, and identity of loci altering expression. OBJECTIVES: We measured COMT allele expression ratios in heterozygous human lymphoblast cell lines and brains. METHODS: Using transcribed single nucleotide polymorphisms as endogenous reporters, we developed an RT-coupled 5' nuclease assay for allele expression ratios and applied it to 63 COMT rs4818(C>G) heterozygotes and 68 Val158Met [rs4680(G>A)] heterozygotes. RESULTS: For rs4818(C>G), the C allele was overexpressed relative to the G allele in 18 of 27 lymphoblast lines and 23 of 36 brains. For Val158Met, Met158 was overexpressed relative to Val158 in all (29 of 29) lymphoblast lines and all (39 of 39) brains. Each of the 22 rs4818 heterozygotes without differential allele expression was a Val158/Val158 homozygote. The Met158 allele was overexpressed by 65-77% when compared with Val158 in lymphoblasts and brain. Haplotype augmented ability to predict expression in brain only. However, the expression of the Val158 allele on the high-expressing haplotype was only 19% higher than Val158 alleles on the other haplotype background. CONCLUSIONS: COMT alleles are differentially expressed. The Met158 allele predicts higher mRNA expression in both brain and lymphoblasts. As exemplified here, the RT-coupled 5' nuclease assay is a reliable method for the quantitative evaluation of cis-acting regulatory effects. PMID- 15290043 TI - Molecular characterization of HMW glutenin subunit allele 1Bx14: further insights in to the evolution of Glu-B1-1 alleles in wheat and related species. AB - 1Bx14 is a member of the high molecular weight (HMW) glutenin subunits specified by wheat Glu-B1-1 alleles. In this work, we found that the full-length amino acid sequence of 1Bx14 and 1Bx20, the last two of the three cysteine residues, which are conserved in 1Bx7, 1Bx17 and homologous 1Ax and 1Dx subunits, were replaced by tyrosine residues. In the 5' flanking regions (-900 to -1,200 bp relative to the start codon), a novel miniature inverted-repeat transposable element insertion was present in 1Bx12 and 1Bx20 but not 1Bx7 and 1Bx17. 1Bx14 and 1Bx20 like alleles were readily found in tetraploid wheat subspecies but not several S genome containing Aegilops species. Phylogenetic analysis showed that the four molecularly characterized Glu-B1-1 alleles (1Bx7, 1Bx14, 1Bx17, 1Bx20) could be divided into two allelic lineages. The lineage represented by 1Bx7 and 1Bx17 was more ancient than the one represented by 1Bx14 and 1Bx20. Combined, our data establish that 1Bx14 and 1Bx20 represent a novel subclass of Glu-B1-1 alleles. Based on current knowledge, potential mechanism involved in the differentiation of two Glu-B1-1 lineages is discussed. PMID- 15290046 TI - Open-nucleus breeding strategies compared with population-wide positive assortative mating: II. Unequal distribution of testing effort. AB - This study compares population-wide positive assortative mating (PAM) with open nucleus breeding with an elite and main population when more effort is allocated to parents of the elite. A companion study showed that PAM is advantageous when testing effort is independent of parental value. In the present study,unbalanced testing was imposed by varying the number of crosses or the number of genotypes per cross. These unbalanced alternatives are compared with PAM, where the testing effort was varied so that better parents were mated more frequently. More effort allocated to parents of higher rank increased the additive effect and the additive variance and only slightly altered the group coancestry and inbreeding in the breeding population (BP) compared with completely balanced scenarios. Of particular interest to the breeder, large enhancement of the additive variance in the BP contributed to higher gains in the production population (PP). These simulations demonstrate that population-wide PAM leads to higher genetic gains compared with open-nucleus alternatives at any desired target level of diversity in the PP. This is true for both balanced (part I)and unbalanced distribution of testing effort (part II). PMID- 15290047 TI - Use of 2n gametes for the production of sexual polyploids from sterile Oriental x Asiatic hybrids of lilies (Lilium). AB - Sixteen Oriental and 12 Asiatic cultivars were crossed in 158 different combinations. A total of 708 F1 hybrids were obtained from 86 of the different combinations of 15 Oriental and 11 Asiatic cultivars. Because the Lilium cultivars (2n=2x=24) used for the production of these hybrids belong to two different taxonomic sections-Archelirion (0) and Sinomartagon (A), respectively the F1 hybrids (OA) could be obtained only through embryo, embryo sac rescue, ovary slice or ovule culture. Most of the F, hybrids were highly sterile (did not produce viable n gametes) due to the failure of chromosome pairing. However, in a few cases F1 plants were found that produced viable 2n pollen at variable frequencies. These 2n pollen grains were successfully used for the production of backcross progenies. Using genomic in situ hybridization we found intergenomic recombinant chromosomes in the sexual polyploid progenies. These results indicate that there are effective prospects for combining important horticultural traits from the two main groups of cultivars of lilies through sexual polyploidization. PMID- 15290049 TI - Identification and mapping of AFLP markers linked to peanut (Arachis hypogaea L.) resistance to the aphid vector of groundnut rosette disease. AB - Groundnut rosette disease is the most destructive viral disease of peanut in Africa and can cause serious yield losses under favourable conditions. The development of disease-resistant cultivars is the most effective control strategy. Resistance to the aphid vector, Aphis craccivora, was identified in the breeding line ICG 12991 and is controlled by a single recessive gene. Bulked segregant analysis (BSA) and amplified fragment length polymorphism (AFLP) analysis were employed to identify DNA markers linked to aphid resistance and for the development of a partial genetic linkage map. A F(2:3) population was developed from a cross using the aphid-resistant parent ICG 12991. Genotyping was carried out in the F2 generation and phenotyping in the F3 generation. Results were used to assign individual F2 lines as homozygous-resistant, homozygous susceptible or segregating. A total of 308 AFLP (20 EcoRI+3/MseI+3, 144 MluI+3/MseI+3 and 144 PstI+3/MseI+3) primer combinations were used to identify markers associated with aphid resistance in the F(2:3) population. Twenty putative markers were identified, of which 12 mapped to five linkage groups covering a map distance of 139.4 cM. A single recessive gene was mapped on linkage group 1, 3.9 cM from a marker originating from the susceptible parent, that explained 76.1% of the phenotypic variation for aphid resistance. This study represents the first report on the identification of molecular markers closely linked to aphid resistance to groundnut rosette disease and the construction of the first partial genetic linkage map for cultivated peanut. PMID- 15290050 TI - Development and assessment of microarray-based DNA fingerprinting in Eucalyptus grandis. AB - Development of improved Eucalyptus genotypes involves the routine identification of breeding stock and superior clones. Currently, microsatellites and random amplified polymorphic DNA markers are the most widely used DNA-based techniques for fingerprinting of these trees. While these techniques have provided rapid and powerful fingerprinting assays, they are constrained by their reliance on gel or capillary electrophoresis, and therefore, relatively low throughput of fragment analysis. In contrast, recently developed microarray technology holds the promise of parallel analysis of thousands of markers in plant genomes. The aim of this study was to develop a DNA fingerprinting chip for Eucalyptus grandis and to investigate its usefulness for fingerprinting of eucalypt trees. A prototype chip was prepared using a partial genomic library from total genomic DNA of 23 E. grandis trees, of which 22 were full siblings. A total of 384 cloned genomic fragments were individually amplified and arrayed onto glass slides. DNA fingerprints were obtained for 17 individuals by hybridizing labeled genome representations of the individual trees to the 384-element chip. Polymorphic DNA fragments were identified by evaluating the binary distribution of their background-corrected signal intensities across full-sib individuals. Among 384 DNA fragments on the chip, 104 (27%) were found to be polymorphic. Hybridization of these polymorphic fragments was highly repeatable (R2>0.91) within the E. grandis individuals, and they allowed us to identify all 17 full-sib individuals. Our results suggest that DNA microarrays can be used to effectively fingerprint large numbers of closely related Eucalyptus trees. PMID- 15290051 TI - The use of microsatellite markers for the detection of genetic similarity among winter bread wheat lines for chromosome 3A. AB - Previous studies with chromosome substitution and recombinant inbred chromosome lines identified that chromosome 3A of wheat cv. Wichita contains alleles that influence grain yield, yield components and agronomic performance traits relative to alleles on chromosome 3A of Cheyenne, a cultivar believed to be the founder parent of many Nebraska developed cultivars. This study was carried out to examine the genetic similarity among wheat cultivars based on the variation in chromosome 3A. Forty-eight cultivars, two promising lines and four substitution lines (in duplicate) were included in the study. Thirty-six chromosome 3A specific and 12 group-3 barley simple sequence repeat (SSR) primer pairs were used. A total of 106 polymorphic bands were scored. Transferability of barley microsatellite markers to wheat was 73%. The coefficient of genetic distance (D) among the genotypes ranged from 0.40 to 0.91 and averaged D=0.66. Cluster analysis by the unweighted pair-group method with arithmetic averages showed one large and one small cluster with eight minor clusters in the large cluster. Several known pedigree relationships largely corresponded with the results of SSR clusters and principal coordinate analysis. Cluster analysis was also carried out by using 22 alleles that separate Wichita 3A from Cheyenne 3A, and three clusters were identified (a small cluster related to Cheyenne of mainly western Nebraska wheat cultivars; a larger, intermediate cluster with many modern Nebraska wheat cultivars; a large cluster related to Wichita with many modern high-yielding or Kansas wheat cultivars). Using three SSR markers that identify known agronomically important quantitative trait loci (QTL) regions, we again separated the cultivars into three main clusters that were related to Cheyenne or Wichita, or had a different 3A lineage. These results suggest that SSR markers linked to agronomically important QTLs are a valuable asset for estimating both genetic similarity for chromosome 3A and how the chromosome has been used in cultivar improvement. PMID- 15290052 TI - A genetic linkage map of microsatellite, gene-specific and morphological markers in diploid Fragaria. AB - Diploid Fragaria provide a potential model for genomic studies in the Rosaceae. To develop a genetic linkage map of diploid Fragaria, we scored 78 markers (68 microsatellites, one sequence-characterised amplified region, six gene-specific markers and three morphological traits) in an interspecific F2 population of 94 plants generated from a cross of F.vesca f. semperflorens x F. nubicola. Co segregation analysis arranged 76 markers into seven discrete linkage groups covering 448 cM, with linkage group sizes ranging from 100.3 cM to 22.9 cM. Marker coverage was generally good; however some clustering of markers was observed on six of the seven linkage groups. Segregation distortion was observed at a high proportion of loci (54%), which could reflect the interspecific nature of the progeny and, in some cases, the self-incompatibility of F. nubicola. Such distortion may also account for some of the marker clustering observed in the map. One of the morphological markers, pale-green leaf (pg) has not previously been mapped in Fragaria and was located to the mid-point of linkage group VI. The transferable nature of the markers used in this study means that the map will be ideal for use as a framework for additional marker incorporation aimed at enhancing and resolving map coverage of the diploid Fragaria genome. The map also provides a sound basis for linkage map transfer to the cultivated octoploid strawberry. PMID- 15290053 TI - Mapping QTL associated with resistance to Fusarium head blight in the Nanda2419 x Wangshuibai population. I. Type II resistance. AB - Scab disease caused by Fusarium spp. has been a major concern for both wheat producers and consumers. Deployment of scab-resistant varieties is the major strategy to curb this disease. To identify the scab resistance genes in wheat cv. Wangshuibai, we produced a F(6:7) recombinant inbred line (RIL) population by crossing Wangshuibai with the scab-susceptible cultivar Nanda2419. The RILs were evaluated for scab resistance in the field by single floret inoculation in two replicates in 2002 and one replicate in 2003. The number of diseased spikelets (NDS) and the length of diseased rachides (LDR) were investigated to reflect the Type II resistance. Among 654 simple sequence repeat (SSR) markers surveyed, 326 were found to be polymorphic between the parents. A partial molecular map was constructed with these markers that covered over 2,210 cM of the wheat genome. Six chromosome regions showed association with both NDS and LDR in a one-way anova analysis, even though the variation explained by them varied between the two traits. Eight intervals were detected for their association with Type II resistance through interval mapping, five of which were not identified in single point analysis. The quantitative trait loci (QTL) with large effects were the ones in the interval of Xgwm533-3-Xgwm533-1 on chromosome 3B and in the interval of Xwmc539-Xbarc024 on chromosome 6B, whose alleles favoring resistance originate from Wangshuibai. In addition, a QTL whose resistance allele originated from Nanda2419 was consistently detected in the interval of Xs1021m-Xgwm47-1 on chromosome 2B. These results suggest that Wangshuibai is the major source for Type II resistance in this population. The markers associated with these QTL would facilitate the use of scab-resistant genes of Wangshuibai in scab resistance breeding programs of wheat. PMID- 15290063 TI - [Extraordinary high cerebrospinal fluid protein in two cases of intracranial hypotension syndrome]. AB - Intracranial hypotension is a rare cause of chronic headache. Although there is still debate about the aetiology, it is believed that the syndrome is caused by low cerebrospinal fluid volumes due to dural leakage. Such leakages can occur spontaneously after lumbar puncture or surgical or traumatic opening of the dura. In magnetic resonance contrast imaging, diffuse meningeal enhancement can be seen; usually the pressure at the cerebrospinal opening is lower than normal. Sometimes a pleocytosis and, in most cases, increased protein content can be identified in the CSF. These protein levels most frequently range between 0.5 g/l and 2 g/l. Here we describe two patients with typical clinical signs and neuroradiological alterations of intracranial hypotension syndrome but with extraordinarily high CSF protein levels (8.3 g/l and 9.63 g/l). On the basis of these findings, the putative causes of elevated CSF protein contents are discussed. PMID- 15290064 TI - Gyral abnormalities in the early stage of superior sagittal sinus thrombosis. AB - We report computed tomography (CT) features in a case of segmental thrombosis of the superior sagittal sinus. On the initial cranial CT scan, both frontal cortices showed focal areas of slightly increased attenuation. The lesions were isointense on magnetic resonance (MR) images no matter what pulse sequence was used, except on fluid-attenuated inversion recovery images, which showed curvilinear sulcal hyperintensity. On postcontrast T1-weighted images, curvilinearly enhancing structures were apparent in both frontal cortical sulci. No lesion appeared on follow-up CT or in MR images. We speculated that the areas of slightly increased attenuation in the cortices represented blood congestion in the cortical veins, venules and capillaries without serum leakage. Cranial CT images should be carefully interpreted to avoid overlooking subtle lesions. PMID- 15290065 TI - Remitting/relapsing idiopathic hypertrophic spinal pachymeningitidis: comprehensive imaging work-up and MR monitoring. AB - We present the initial MR and PET work-up in a diabetic 31-year-old female patient presenting with spinal cord compression because of abnormal meningeal tissue and highlight the close correlation between the clinical course and the monitoring of the lesions on MR images. The combination of non-specific inflammatory signs at histopathological examination and of the negative results of a comprehensive bacteriological and serological testing led to the diagnosis of idiopathic hypertrophic spinal pachymeningitidis. The abnormal tissue showed mild glucose uptake at whole-body FDG-PET examination. Repeated MR follow-up examinations during corticoid treatment only demonstrated partial shrinkage of the abnormal tissue. Symptomatic relapse and re-increase in lesion size on MR images occurred concomitantly after an 11-month symptom-free remitting interval under tapering corticoid treatment. PMID- 15290066 TI - Pediatric brain MRI in neurofibromatosis type I. AB - Neurofibromatosis (NF) is the most common of the phakomatoses, with a prevalence of 1 in 3-4,000. Many organ systems can be affected. In addition to multiple peripheral neurofibromas, NF I predisposed to CNS tumors including optic glioma, astrocytoma and plexiform neurofibroma. The purpose of this pictorial review is to illustrate characteristic brain MR imaging lesions in children with NF I and to give some recommendations about diagnostic imaging procedures in children suffering from NF I. Typical findings in brain MRI are hyperintense lesion on T2 weighted images, so-called unknown bright objects, which may be useful as an additional imaging criterion for NF I. Contrast administration is necessary in MR studies to maximize tumor detection and characterization, to add confidence to the diagnosis of benign probable myelin vacuolization, and to document stability of neoplasm on follow-up examinations. We recommend to perform serial MR imaging in children every 12 months. The frequency of follow-up in children with known brain tumors will vary with the tumor grade, biological activity and treatment. PMID- 15290072 TI - CT and MR imaging of unusual locations of extra-adrenal paragangliomas (pheochromocytomas). AB - Our review was undertaken to describe CT and MRI features of unusual extra adrenal paragangliomas (pheochromocytomas). We retrospectively reviewed CT and MRI findings in 29 patients with 39 extra-adrenal paragangliomas. For each tumour, site, size, MRI characteristics, CT appearances and enhancement after gadolinium and iohexol were recorded. There were 17 carotid body tumours, 1 mediastinal, 1 intra-cardiac, 15 retroperitoneal extra-adrenal paragangliomas, 2 bladder, 1 pelvic sidewall and 2 intra-spinal paragangliomas within the lumbo sacral spine. All 39 paragangliomas were shown on MRI. Of the 32 lesions studied by MRI and CT, CT detected 30. Of the two lesions missed on CT, one was an intra cardiac paraganglioma and the second a bladder wall paraganglioma. At detection, 25 tumours were larger than 4 cm, of which 20 were heterogeneous lesions on CT and MRI with variable contrast enhancement. The 14 smaller paragangliomas were smooth in contour and demonstrated avid, homogeneous contrast enhancement. Our review of extra-adrenal paragangliomas highlights their unusual sites and appearances. MRI demonstrated the greatest variability in the appearances of larger tumours, provided additional information compared to CT for surgical planning and is a useful screening tool for patients at high risk of extra adrenal paragangliomas. PMID- 15290067 TI - Accuracy of MRI in characterization of soft tissue tumors and tumor-like lesions. A prospective study in 548 patients. AB - The purpose of our study was to assess prospectively the value of MRI in characterization of soft tissue tumors (STT) and soft tissue tumor-like lesions in a multi-institutional setting by a group of experts. The material consisted of 548 untreated and proven STT or tumor-like lesions originating from a multi institutional database of STT in which 930 consecutive patients with STT examined by MRI were registered between 1 January 2001 and 28 April 2003. Based on MRI findings, a suitably ordered differential diagnosis was made in consensus by two radiologists (J.L.M.A.G and A.M.D.S). MRI diagnoses were compared with histology results (455 cases, 83%) and/or 6-month follow-up (93 cases, 17%) as reference standards. The correlation between the MRI and histological diagnosis and between the radiological and histological phenotype were statistically determined. One hundred twenty-three patients presented with a malignant STT; 425 patients presented with a benign one. Concerning differentiation between malignant and benign lesions (dignity), a sensitivity of 93%, specificity of 82%, negative predictive value (NPV) of 98% and positive predictive value (PPV) of 60% with accuracy of 85% were obtained. Concerning phenotype characterization, if only the first MRI diagnosis was taken into account, a sensitivity of 67%, specificity of 98%, NPV of 98%, PPV of 70% and accuracy of 96% were obtained. For benign lesions, sensitivity of 75%, specificity of 98%, NPV of 98%, PPV of 76% and accuracy of 97% were obtained. The phenotype's definition of malignant STT had a sensitivity of 37%, a specificity of 96%, NPV of 96%, PPV of 40% and an accuracy of 92%. A correct diagnosis compared with histological assessment was proposed in 227(50%) of the 455 histologically confirmed cases. Despite non-quantified MR parameter evaluation, the results of our prospective study were better than those reported in previous studies and demonstrated the need for a centralized approach to such rare pathology. PMID- 15290074 TI - Combined surgical-endovascular treatment of multiple splenic artery aneurysms. PMID- 15290084 TI - Transformation of Saussurea medusa for hairy roots and jaceosidin production. AB - Axenically grown Saussurea medusa plantlets were inoculated with four Agrobacterium rhizogenes strains, and hairy root lines were established with A. rhizogenes strain R1601 in N6 medium. PCR and Southern hybridization confirmed integration of the T-DNA fragment of the Ri plasmid from A. rhizogenes into the genome of S. medusa hairy roots. In N6 medium, maximum biomass of the hairy root cultures was achieved [8 g (dry weight) per liter; growth ratio 35-fold] after 21 days of culture. The amount of jaceosidin extracted from the hairy root cultures was 46 mg/l (production ratio of 37-fold) after 27 days of culture. The maximum jaceosidin content obtained using N6 medium was higher than that obtained with Modified White, MS or B5 medium. In N6 medium, the tip segments were more efficient for hairy root growth and jaceosidin production than the middle and basal regions of the root. PMID- 15290086 TI - Lp(a) lipoprotein and lipids in patients with rheumatoid arthritis: serum levels and relationship to inflammation. AB - OBJECTIVES: Changes in lipid profiles, Lp(a) lipoprotein, and acute phase reactants are associated with early atherosclerosis in rheumatoid arthritis (RA). The associations of Lp(a) levels with atherosclerotic disorders, diabetes, RA, and renal diseases suggest that Lp(a) might be involved in autoimmune reactions. METHODS: Eighty-seven women with RA diagnosed according to American Rheumatism Association criteria (mean age 45.4+/-9.4 years) were recruited and 50 healthy women (mean age 44+/-10.7 years) included as a control group. Serum Lp(a), total cholesterol (TC), triglyceride (TG), LDL cholesterol (LDL-C), HDL cholesterol (HDL-C), and C-reactive protein levels were analyzed. RESULTS: In the RA and C groups, serum Lp(a) levels were 39.2+/-20.6 mg/dl and 14.8+/-9.7 mg/dl, respectively (P<0.001). The TC levels were 188.4+/-41.8 mg/dl and 185.3+/-19.3 mg/dl (P>0.05), TG levels were 124.5+/-50.1 mg/dl and 94.6+/-24.9 mg/dl (P<0.01), HDL-C levels were 40.0+/-7.4 mg/dl and 52.8+/-4.8 mg/dl (P<0.01), and LDL-C levels were 123.4+/-24.6 mg/dl and 113.3+/-21.1 mg/dl (P>0.05). While serum CRP levels showed a positive correlation with Lp(a), they correlated negatively with HDL-C levels (r=0.83 and P<0.0001, r=-0.49 and P<0.0001, respectively). It was meaningful that Lp(a) correlated negatively with serum HDL-C level (r=-0.36, P<0.001). CONCLUSIONS: It is suggested that higher serum Lp(a), lower HDL-C, higher TG level, and a high ratio of TC/HDL-C might show high risk of atherosclerosis. Inflammation in RA may cause changes in HDL-C and Lp(a) metabolisms. PMID- 15290087 TI - A comparison of clinical findings of familial Mediterranean fever patients with and without amyloidosis. AB - OBJECTIVE: This study investigates the clinical and demographic characteristics of familial Mediterranean fever (FMF) patients with and without amyloidosis. PATIENTS AND METHODS: The clinical data of 503 patients with FMF (females:males 250:253) were reviewed. Fifty of these patients had amyloidosis (f:m 23:27). RESULTS: The ages of attack onset in patients with and without amyloidosis were 7.8+/-6.2 and 11.1+/-8.5, respectively (P<0.05). The time between disease onset and diagnosis was longer in patients with amyloidosis than those without (187.6+/ 99.4 months and 132.5+/-110.2 months, respectively, P<0.001). More patients in the amyloidosis group had positive family histories of FMF (68% vs 54%, P<0.05). The frequencies of chest pain (78% vs 51%, P<0.001), arthritis ( 80% vs 60%, P<0.01), and erysipelas-like erythema (44% vs 16%, P<0.001) were higher in the amyloidosis group. CONCLUSION: In the amyloidosis group, FMF-related manifestations of chest pain, arthritis, and erysipelas-like erythema are more frequent. Our results also support that long periods between disease onset and diagnosis are associated with a high risk of developing amyloidosis. PMID- 15290088 TI - Glucocorticoid iontophoresis for Achilles tendon enthesitis in ankylosing spondylitis: significant response documented by power Doppler ultrasound. PMID- 15290089 TI - [Mucinous sweat gland carcinoma of a toe with metastasis to the maxilla. A case report]. AB - Malignant tumors of the maxilla are rare. In addition to primary tumors metastatic disease has to be considered. Renal cell carcinomas, carcinomas of the breast, lung and adenocarcinomas are constituting the most frequent primary lesions for maxillary metastases. To the best of our knowledge, we present the first case of a mucinous sweat gland carcinoma metastasizing to the maxilla four years after primary diagnosis. Thirty-two months after resecting the hemimaxilla, there are no signs of local recurrence. Although skin tumors only rarely develop distant metastases, sweat gland carcinomas should be included in the differential diagnosis of tumors metastasizing to the maxilla. PMID- 15290094 TI - Phase I and pharmacokinetic study of KW-2170, a novel pyrazoloacridone compound, in patients with malignant tumors. AB - PURPOSE: The primary purposes of this study were to determine the dose-limiting toxicity (DLT) and maximum tolerated dose (MTD), to recommend a dose for phase II studies, and to analyze the pharmacokinetics of KW-2170. A secondary purpose was to assess tumor response to KW-2170. EXPERIMENTAL DESIGN: KW-2170 was given as a 30-min i.v. infusion every 4 weeks. Doses were escalated from 1.0 mg/m2 according to a modified Fibonacci method. RESULTS: A total of 45 cycles of KW-2170 were delivered to 41 patients at doses ranging from 1.0 to 53.0 mg/m2. The primary DLT was neutropenia which was observed in two of six patients treated at 32.0 mg/m2 and in two of two patients treated at 53.0 mg/m2; therefore, the MTD was 53.0 mg/m2. Although no patients showed a complete response (CR)or partial response (PR), 15 patients were evaluated a shaving freedom from progression at the 1 month time-point, with two demonstrating slight tumor shrinkage in their metastatic lesions. None of the patients experienced significant cardiotoxicity. The plasma concentration of KW-2170 declined in a triphasic manner. The half life, total clearance (CLtot) and volume of distribution (Vdss) were nearly constant and independent of dose, and showed a relatively small interpatient variability. A linear relationship was observed between dose and maximum plasma concentration (Cmax) and area under the concentration-time curve (AUC 0 infinity). In addition, there was a good correlation between neutropenia and AUC 0- infinity. This suggests that toxicity may be dependent on systemic exposure to the drug. Two oxi-dative metabolites were observed in the patients' plasma and urine. CONCLUSIONS: The primary DLT of KW-2170 in this study was neutropenia, with a MTD of 53 mg/m2.A significant linear relationship was observed between neutropenia and AUC 0- infinity. We estimate the recommended dose for phase II studies to be 41.0 mg/m2. PMID- 15290096 TI - Redox-cycling of anthracyclines by thioredoxin system: increased superoxide generation and DNA damage. AB - Anthracyclines such as doxorubicin and daunomycin undergo bioreductive activation by redox-cycling, and this is associated with generation of reactive oxygen species. Toxicity of anthracyclines is attributed to DNA intercalation by an anthracycline semiquinone radical that is generated via redox-cycling. Flavoprotein enzymes catalyze the bioreductive activation of anthracyclines. Thioredoxin reductase (TR), which is also a flavoprotein enzyme, participates in bioreductive activation of anthracyclines. In the present study we showed that addition of E. coli thioredoxin (Trx) enhances the rate of superoxide production by E. coli TR in the presence of anthracyclines. The superoxide generated in this redox-cycling process induced DNA damage as determined by an in vitro plasmid DNA damage assay. In addition, Trx-SH enhanced the activity of cyto-chrome P450 reductase and the redox-cycling of anthracyclines independently of NADPH. Furthermore,when A549 cells were incubated with E. coli Trx followed by doxorubicin treatment, increased levels of ROS generation were observed. Taken together, these results show a novel property of the Trx system in bioreductive activation of anthracyclines. PMID- 15290100 TI - Synergistic antitumor activity of TRAIL combined with chemotherapeutic agents in A549 cell lines in vitro and in vivo. AB - PURPOSE: To investigate the synergistic cytotoxicity of TRAIL in combination with chemotherapeutic agents in A549 cell lines, we systematically evaluated the cytotoxicity of TRAIL alone and TRAIL in combination with cisplatin, paclitaxel (Taxol) or actinomycin D in A549 cell lines in vitro and in vivo, and whether the sensitivity was correlated with the expression level of TRAIL receptors. METHODS: We investigated the cytotoxicity of TRAIL alone and the synergistic antitumor effects of TRAIL in combination with chemotherapeutic agents in A549 cells by crystal violet staining and FACS in vitro. The expression levels of DR4, DR5, DcR1 and DcR2 were measured in TRAIL-treated and chemotherapeutic agent-treated A549 cells by Western blotting. The growth inhibition of tumors was evaluated in terms of incidence, volume and weight in a A549-implanted nude mice model. RESULTS: Chemotherapeutic agents cisplatin (5.56 mug/ml), Taxol (10 and 30 mug/ml) or actinomycin D (9.26, 83.3 and 750 ng/ml) augmented the cytotoxicity of TRAIL in A549 cell lines within a range of concentrations of TRAIL (1.98-160 ng/ml) in vitro. The expression levels of DR4 and DR5 were not significantly different and the expression of DcR2 was slightly downregulated, but the expression of DcR1 was not detected in non-treated, TRAIL-treated and chemotherapeutic agent-treated A549 cells. The rates of tumor inhibition following treatment with TRAIL alone (15 mg/kg per day, daily for 10 days) and TRAIL/cisplatin (15 mg/kg per day TRAIL, daily for 10 days; 1.5 mg/kg per day cisplatin, daily for 10 days with 7-day intervals) were 28.3% and 76.8% by tumor weight ( P<0.05 for TRAIL alone versus control, P<0.05 for TRAIL/cisplatin versus cisplatin alone and TRAIL alone) on day 65 in vivo. CONCLUSION: TRAIL in combination with chemotherapeutic agents cisplatin, Taxol or actinomycin D exerted synergistic antitumor effects in A549 cell lines in vitro and TRAIL/cisplatin demonstrated synergistic antitumor effects in vivo. The expression levels of TRAIL receptors suggested that the synergistic effects of TRAIL in combination with chemotherapeutic agents are not at the receptor level in A549 cell lines. PMID- 15290102 TI - The spectrum of bleeding disorders in women with menorrhagia: a report from Western India. AB - In order to evaluate the incidence of hereditary bleeding disorders in patients presenting with menorrhagia, where the usual gynecological and endocrinal causes of bleeding were ruled out by various local ultrasonography (USG) and relevant endocrine investigations, 120 women aged between 18 and 35 years presenting with menorrhagia without any discernable cause were studied using an open design, where the investigators knew that these patients had menorrhagia. These patients were investigated for inherited coagulation defects. Of the 120 women investigated, 19.16% (23 cases) had an inherited coagulation disorder to account for their menorrhagia. Although a majority (11.6%) are patients with von Willebrand's disease (VWD), other rare platelet disorders such as Glanzmann's thrombasthenia (3.3%), Bernard-Soulier syndrome (0.83%), coagulation factor deficiencies such as factor VIII (0.83%), factor X (0.83%), and factor XI (0.83%), and immune thrombocytopenia (0.83%) were also found. Taking a detailed history for bleeding from other sites however minor, paternal consanguinity as well as family history of bleeding tendencies appeared as a very strong predictor for such kinds of disease in patients with menorrhagia. Patients with menorrhagia without a discernable cause, therefore, need evaluation for the congenital coagulation disorders. PMID- 15290103 TI - Histological study of the human temporo-mandibular joint and its surrounding muscles. AB - This is a histological study of the human temporo-mandibular joint and its surrounding muscles. Using a microscopic study of serial sections from anatomical specimens from six subjects, the detailed anatomy of the joint is presented with particular regard to the histology. This study has allowed, in particular, the description of the ligaments and capsule as well as the insertions of the masticatory muscles (temporalis, masseter, lateral pterygoid) on this joint. These observations are then compared with the anatomical and histological data already reported on this subject. This study shows that the bulk of the muscular fibres of the lateral pterygoid passes under the foot of the disc is attached over the whole height of the condylar, unite and extend as far as the medial pole of the joint under the insertion of the articular disc. An insertion of the temporo-masseter musculo-tendinous complex on the anterior and lateral capsulo discal structures was observed. The lateral pterygoid is composed of a succession of tendinous and fleshy fibres. This study confirms the thickening of the lateral capsule that corresponds to a lateral collateral ligament, and the absence of a medial collateral ligament. Medial stability is conferred by the lateral ligament of the contralateral joint. PMID- 15290104 TI - Quantitation of ligament laxity in anterior shoulder instability: an experimental cadaver model. AB - Three groups of cadaver specimens were studied. In group 1 (20 shoulders) glenohumeral ligaments were detached from the humerus until a permanent dislocation of the humeral head occurred in abduction plus external rotation. On the dislocated joint the ligament was reconstructed using a fascia lata lengthening plasty. After the plasty had been completed, the shoulder was reduced and instability checked in the same position. Then the capsule (including the plasty) was harvested and measured. In group 2 (20 shoulders), after the plasty had been completed in the same conditions as above, the capsule was progressively reduced by 2 mm steps until the instability disappeared. Then the capsule (including the plasty) was harvested and measured. In group 3 (12 shoulders), measurements of the head and of the capsule were done. To dislocate the shoulder the section of the three glenohumeral ligaments was required. Lengthening of the capsule in group 1 was 240-250%. In all cases shortening of the capsule led to the stabilization of the shoulder. After stabilization of the shoulder was reached a residual lengthening of 175-185% was recorded. In 3 out of 4 shoulders the amount of capsule shortening required to return to a stable shoulder was between 16 and 18 mm. This experiment did not reproduce the Bankart lesion; therefore it only concerns atraumatic instability. The main limitation of this model is the low lever force that may be used to dislocate the shoulder; consequently the elasticity of the glenohumeral ligament was not taken in account. The experimental values were likely over-estimated. Nevertheless the present results provide useful information for building an experimental model of atraumatic instability of the shoulder. PMID- 15290105 TI - Kinematics of the wrist using 2D and 3D analysis: biomechanical and clinical deductions. AB - Surgery of the wrist relies on the known notions of biomechanics of the wrist. But these notions are incomplete. For a better understanding of the movements of the wrist, we studied five wrists of healthy volunteers with CT scanning. Each wrist was studied in neutral position, and in the four extreme positions: flexion, extension, radial and ulnar deviations. Using oblique reformatted CT sections, we measured the angular displacements in frontal and sagittal views of every carpal bone in the different positions of the wrist. This allowed us to construct a table of intracarpal mobility. By comparing the angle values and the three-dimensional pictures of these wrists, we illustrate some fundamental points regarding intracarpal movement. The dynamics of the wrist are like those of two super-imposed mobile cups with different movements. The proximal row is malleable with flattening and torsion according to the transverse axis and its behavior is like that of an articular meniscus. The distal row, more rigid but deformable, behaves like a T-handle giving attachment to the hand and articulating under the proximal row around the head of the capitate and the proximal pole of the hamate. During radial and ulnar deviations of the wrist, the movement between the two rows is like an inverse pronation-supination shearing. During flexion-extension, the distortion of the two rows allows maximal congruence to be maintained between the different carpal bones. PMID- 15290106 TI - Anatomical variations of occipital bone impressions for dural venous sinuses around the torcular Herophili, with special reference to the consideration of clinical significance. AB - Venous blood flow through the cerebral dural sinus is variable and clinically significant. It has been investigated by cadaver dissection or radiology; however, we thought that osteology might be informative. A total of 160 dried skulls were macroscopically examined for impressions on the inner surface of the occipital bone in order to interpret the sinus flow around the torcular Herophili. The continuity between the grooves for the superior sagittal sinus (SSS) and the transverse sinuses was categorized into four types. Confluence type was noted in 56 specimens (35%), in which SSS drained into a common pool of venous sinuses. Bifurcation type was noted in 22 cases (14%), in which SSS was divided to drain into the bilateral transverse sinuses. Right dominant type was the most frequent one with 66 cases (41%), in which SSS drained only into the right transverse sinus. Left dominant type was the least frequent one with 16 cases (10%), in which SSS drained to the left, in a mirror image to the right dominant type. Clinical significance is discussed with our preliminary trial for the optimization of the inner skull surface and venous flow using computed tomography and magnetic resonance imaging, and demonstration of cerebrovascular disease. PMID- 15290107 TI - Anatomical study concerning the origin and course of the pectoral branch of the thoracoacromial trunk for the pectoralis major flap. AB - The patterns of the feeding vessels to each muscle determine the extent of their safe transposition and the muscle's value as a pedicled flap in reconstructive surgery. This study aimed to demonstrate the point of origin and the intra- and submuscular course of the pectoral branch of the thoracoacromial trunk (TAT) for pectoralis major (PM) flap surgery. Seventy sides of the PM were dissected based on a clinical reference line that has been used for several decades. The branching point of the TAT from the axillary artery was located lateral to the midclavicular line on the right-sided specimens (100%) and medial to the midclavicular line on the left sides (86%). The branching patterns of the pectoral branch to the PM muscle from the TAT were classified into three types. In type I the pectoral branches originated directly from the TAT (55 cases, 78.6%). In type II (11 cases, 15.7%) and type III (4 cases, 5.7%) the pectoral branch divided from the medial and lateral pedicle of the TAT, respectively. The course of the pectoral branch from the TAT in the PM was categorized into three patterns according to the degree of proximity to the midclavicular line. In 49 cases (70%), the pectoral branch in the PM ran within 1 cm of the midclavicular line. The other cases ran 2 cm (20 cases, 29%) and 3 cm (1 case, 1%) from the midclavicular line, respectively. These results provide topographic data of the pectoral branch based on anatomical landmarks, and will be useful in surgical planning as well as the procedure for PM flap surgery. PMID- 15290114 TI - Variability of left ventricular ejection fraction and volumes with quantitative gated SPECT: influence of algorithm, pixel size and reconstruction parameters in small and normal-sized hearts. AB - PURPOSE: Several software packages are commercially available for quantification of left ventricular ejection fraction (LVEF) and volumes from myocardial gated single-photon emission computed tomography (SPECT), all of which display a high reproducibility. However, their accuracy has been questioned in patients with a small heart. This study aimed to evaluate the performances of different software and the influence of modifications in acquisition or reconstruction parameters on LVEF and volume measurements, depending on the heart size. METHODS: In 31 patients referred for gated SPECT, 64(2) and 128(2) matrix acquisitions were consecutively obtained. After reconstruction by filtered back-projection (Butterworth, 0.4, 0.5 or 0.6 cycles/cm cut-off, order 6), LVEF and volumes were computed with different software [three versions of Quantitative Gated SPECT (QGS), the Emory Cardiac Toolbox (ECT) and the Stanford University (SU-Segami) Medical School algorithm] and processing workstations. Depending upon their end systolic volume (ESV), patients were classified into two groups: group I (ESV>30 ml, n=14) and group II (ESV<30 ml, n=17). Agreement between the different software packages and the influence of matrix size and sharpness of the filter on LVEF and volumes were evaluated in both groups. RESULTS: In group I, the correlation coefficients between the different methods ranged from 0.82 to 0.94 except for SU-Segami (r=0.77), and were slightly lower for volumes than for LVEF. Mean differences between the methods were not significant, except for ECT, with which LVEF values were systematically higher by more than 10%. Changes in matrix size had no significant influence on LVEF or volumes. On the other hand, a sharper filter was associated with significantly larger volume values though this did not usually result in significant changes in LVEF. In group II, many patients had an LVEF in the higher range. The correlation coefficients between the different methods ranged between 0.80 and 0.96 except for SU-Segami (r=0.49), and were slightly worse for volumes than for LVEF values. In contrast to group I, however, inter-method variability was quite large and most mean LVEF differences were significant. LVEF was systematically highest with ECT and lowest with SU Segami. With QGS, changes in matrix size from 64(2) to 128(2) were associated with significantly larger volumes as well as lower LVEF values. Increasing the filter cut-off frequency had the same effect. With SU-Segami, a larger matrix was associated with larger end-diastolic volumes and smaller ESVs, resulting in a highly significant increase in LVEF. Increasing the filter sharpness, on the other hand, had no influence on LVEF though the measured volumes were significantly larger. CONCLUSION: In patients with a normal-sized heart, LVEF and volume estimates computed from different commercially available software packages for quantitative gated SPECT are well correlated. LVEF and volumes are only slightly sensitive to changes in matrix size. Smoothing, by contrast, is associated with significant changes in volumes but usually not in LVEF values. However, owing to the specific characteristics of each algorithm, software should not be interchanged for follow-up in an individual patient. In small hearts, on the other hand, both the used software and the matrix size or smoothing significantly influence the results of quantitative gated SPECT. LVEF values in the higher range are frequently observed with all the studied software except for SU-Segami. A larger matrix or a sharper filter could be suggested to enhance the accuracy of most commercial software, more particularly in patients with a small heart. PMID- 15290115 TI - Relation between wall thickening on gated perfusion SPECT and functional recovery after coronary revascularization in patients with previous myocardial infarction. AB - PURPOSE: This study aimed to evaluate whether wall thickening analysis by gated perfusion single-photon emission computed tomography (SPECT) is useful in predicting functional recovery after revascularization. METHODS: Forty-one patients with previous myocardial infarction and left ventricular (LV) dysfunction (ejection fraction, EF, 36+/-6%) who were scheduled for revascularization underwent rest 99mTc-sestamibi gated SPECT. RESULTS: Of 131 akinetic or dyskinetic segments at baseline echocardiography, 82 (63%) recovered after revascularization. Compared with wall thickening analysis, perfusion imaging provided higher sensitivity (78% vs 50%, P<0.0001) and specificity (80% vs 71%, P<0.0005). Among segments with > or =55% sestamibi uptake (viable), those with detectable wall thickening had a higher likelihood of functional recovery than those with absent wall thickening (95% vs 77%, P<0.05). In segments with improved function, the absence of wall thickening was associated with lower sestamibi activity than was observed when detectable wall thickening was present (58+/-14% vs 71+/-13%, P<0.0005). An increase in EF of > or =5% was detectable in 22 (54%) patients. For the prediction of EF improvement, perfusion imaging provided a higher sensitivity than wall thickening analysis (68% vs 41%, P<0.05), while specificity was not significantly different (68% vs 74%). The prevalence of patients with functional recovery did not change when wall thickening analysis was considered in addition to perfusion status (73% in patients with detectable wall thickening and 70% in those without; P=NS). CONCLUSION: In patients with coronary artery disease, wall thickening analysis by gated perfusion SPECT provides additional information compared with perfusion data for the prediction of segmental functional recovery. However, on a patient basis, wall thickening assessment seems to be of more limited value than perfusion status. PMID- 15290118 TI - Tetrahydrobiopterin restores impaired coronary microvascular dysfunction in hypercholesterolaemia. AB - PURPOSE: Tetrahydrobiopterin (BH4) is an essential co-factor for the synthesis of nitric oxide (NO), and BH4 deficiency may cause impaired NO synthase (NOS) activity. We studied whether BH4 deficiency contributes to the coronary microcirculatory dysfunction observed in patients with hypercholesterolaemia. METHODS: Myocardial blood flow (MBF; ml min(-1) g(-1)) was measured at rest, during adenosine-induced (140 microg kg(-1) min(-1) over 7 min) hyperaemia (mainly non-endothelium dependent) and immediately after supine bicycle exercise (endothelium-dependent) stress in ten healthy volunteers and in nine hypercholesterolaemic subjects using 15O-labelled water and positron emission tomography. Measurements were repeated 60 min later, after intravenous infusion of BH4 (10 mg kg(-1) body weight over 30 min). Adenosine-induced hyperaemic MBF is considered to represent (near) maximal flow. Flow reserve utilisation was calculated as the ratio of exercise-induced to adenosine-induced hyperaemic MBF and expressed as percent to indicate how much of the maximal (adenosine-induced) hyperaemia can be achieved by bicycle stress. RESULTS: BH4 increased exercise induced hyperaemia in controls (2.96+/-0.58 vs 3.41+/-0.73 ml min(-1) g(-1), p<0.05) and hypercholesterolaemic subjects (2.47+/-0.78 vs 2.70+/-0.72 ml min(-1) g(-1), p<0.01) but had no influence on MBF at rest or during adenosine-induced hyperaemia in controls (4.52+/-1.10 vs 4.85+/-0.45 ml min(-1) g(-1), p=NS) or hypercholesterolaemic subjects (4.86+/-1.18 vs 4.53+/-0.93 ml min(-1) g(-1), p=NS). Flow reserve utilisation remained unchanged in controls (70+/-17% vs 71+/ 19%, p=NS) but increased significantly in hypercholesterolaemic subjects (53+/ 15% vs 66+/-14%, p<0.05). CONCLUSION: BH4 restores flow reserve utilisation of the coronary microcirculation in hypercholesterolaemic subjects, suggesting that BH4 deficiency may contribute to coronary microcirculatory dysfunction in hypercholesterolaemia. PMID- 15290119 TI - Brain perfusion abnormalities in patients with euthyroid autoimmune thyroiditis. AB - PURPOSE: Brain perfusion abnormalities have recently been demonstrated by single photon emission computed tomography (SPECT) in rare cases of severe Hashimoto's thyroiditis (HT) encephalopathy; moreover, some degree of subtle central nervous system (CNS) involvement has been hypothesised in HT, but no direct evidence has been provided so far. The aim of this study was to assess cortical brain perfusion in patients with euthyroid HT without any clinical evidence of CNS involvement by means of 99mTc-ECD brain SPECT. Sixteen adult patients with HT entered this study following informed consent. METHODS: The diagnosis was based on the coexistence of high titres of anti-thyroid auto-antibodies and diffuse hypoechogenicity of the thyroid on ultrasound in association with normal circulating thyroid hormone and TSH concentrations. Nine consecutive adult patients with non-toxic nodular goitre (NTNG) and ten healthy subjects matched for age and sex were included as control groups. All patients underwent 99mTc-ECD brain SPECT. Image assessment was both qualitative and semiquantitative. Semiquantitative analysis was performed by generation of four regions of interest (ROI) for each cerebral hemisphere--frontal, temporal, parietal and occipital- and one for each cerebellar hemisphere in order to evaluate cortical perfusion asymmetry. The Asymmetry Index (AI) was calculated to provide a measurement of both magnitude and direction of perfusion asymmetry. RESULTS: As assessed by visual examination, 99mTc-ECD cerebral distribution was irregular and patchy in HT patients, hypoperfusion being more frequently found in frontal lobes. AI revealed abnormalities in 12/16 HT patients, in three of the nine NTNG patients and in none of the normal controls. A significant difference in the mean AI was found between patients with HT and both patients with NTNG (p<0.003) and normal controls (p<0.001), when only frontal lobes were considered. CONCLUSION: These results show the high prevalence of brain perfusion abnormalities in euthyroid HT. These abnormalities are similar to those observed in cases of severe Hashimoto's encephalopathy and may suggest a higher than expected involvement of CNS in thyroid autoimmune disease. PMID- 15290120 TI - Assessment of right ventricular oxidative metabolism by PET in patients with idiopathic dilated cardiomyopathy undergoing cardiac resynchronization therapy. AB - PURPOSE: Right ventricular (RV) performance is known to have prognostic value in patients with congestive heart failure (CHF). Cardiac resynchronization therapy (CRT) has been found to enhance left ventricular (LV) energetics and metabolic reserve in patients with heart failure. The interplay between the LV and RV may play an important role in CRT response. The purpose of the study was to investigate RV oxidative metabolism, metabolic reserve and the effects of CRT in patients with CHF and left bundle brach block. In addition, the role of the RV in the response to CRT was evaluated. METHODS: Ten patients with idiopathic dilated cardiomyopathy who had undergone implantation of a biventricular pacemaker 8+/-5 months earlier were studied under two conditions: CRT ON and after CRT had been switched OFF for 24 h. Oxidative metabolism was measured using [11C]acetate positron emission tomography (Kmono). The measurements were performed at rest and during dobutamine-induced stress (5 microg/kg per minute). LV performance and interventricular mechanical delay (interventricular asynchrony) were measured using echocardiography. RESULTS: CRT had no effect on RV Kmono at rest (ON: 0.052+/-0.014, OFF: 0.047+/-0.018, NS). Dobutamine-induced stress increased RV Kmono significantly under both conditions but oxidative metabolism was more enhanced when CRT was ON (0.076+/-0.026 vs 0.065+/-0.027, p=0.003). CRT shortened interventricular delay significantly (45+/-33 vs 19+/-35 ms, p=0.05). In five patients the response to CRT was striking (32% increase in mean LV stroke volume, range 18-36%), while in the other five patients no response was observed (mean change +2%, range -6% to +4%). RV Kmono and LV stroke volume response to CRT correlated inversely (r=-0.66, p=0.034). None of the other measured parameters, including all LV parameters and electromechanical parameters, were associated with the response to CRT. In responders, RV Kmono with CRT OFF was significantly lower than in non-responders (0.036+/-0.01 vs 0.058+/-0.02, p=0.047). CONCLUSION: CRT appears to enhance RV oxidative metabolism and metabolic reserve during stress. Patients responding to CRT appear to have lower RV oxidative metabolism at rest, suggesting that the RV plays a significant role in the response to CRT. PMID- 15290121 TI - High-quality 124I-labelled monoclonal antibodies for use as PET scouting agents prior to 131I-radioimmunotherapy. AB - PURPOSE: Monoclonal antibodies (MAbs) labelled with 124I are an attractive option for quantitative imaging with positron emission tomography (PET) in a scouting procedure prior to 131I-radioimmunotherapy (131I-RIT). In this study, three important items in the labelling of MAbs with 124I were introduced to obtain optimal and reproducible product quality: restoration of radiation-induced inorganic deterioration of the starting 124I solution, radiation protection during and after 124I labelling, and synchronisation of the I/MAb molar ratio. METHODS: A new method was applied, using an NaIO3/NaI carrier mix, realising in one step >90% restoration of deteriorated 124I into the iodide form and chemical control over the I/MAb molar ratio. Chimeric MAb (cMAb) U36 and the murine MAbs 425 and E48 were labelled with 124I using the so-called Iodogen-coated MAb method, as this method provides optimal quality conjugates under challenging radiation conditions. As a standardising condition, NaIO3/NaI carrier mix was added at a stoichiometric I/MAb molar ratio of 0.9. For comparison, MAbs were labelled with 131I and with a mixture of 124I, 123I, 126I and 130I. RESULTS: Labelling with 124I in this setting resulted in overall yields of >70%, a radiochemical purity of >95%, and preservation of MAb integrity and immunoreactivity, including at the patient dose level (85 MBq). No significant quality differences were observed when compared with 131I products, while the iodine isotope mixture gave exactly the same labelling efficiency for each of the isotopes, excluding a different chemical reactivity of 124I-iodide. The scouting performance of 124I-cMAb U36 labelled at the patient dose level was evaluated in biodistribution studies upon co-injection with 131I-labelled cMAb U36, and by PET imaging in nude mice bearing the head and neck cancer xenograft line HNX-OE. 124I cMAb and 131I-cMAb U36 labelled with a synchronised I/MAb molar ratio gave fully concordant tissue uptake values. Selective tumour uptake was confirmed with immuno-PET, revealing visualisation of 15 out of 15 tumours. CONCLUSION: These results pave the way for renewed evaluation of the potential of 124I-immuno-PET for clinical applications. PMID- 15290124 TI - The impact of PET scanning on management of paediatric oncology patients. AB - PURPOSE: Limited information is available on the use of positron emission tomography (PET) in paediatric oncology. The aim of this study was to review the impact of PET on the management of paediatric patients scanned over a 10-year period. METHODS: One hundred and sixty-five consecutive oncology patients aged 11 months to 17 years were included. Two hundred and thirty-seven scans were performed. Diagnoses included lymphoma (60 patients), central nervous system (CNS) tumour (59), sarcoma (19), plexiform neurofibroma with suspected malignant change (13) and other tumours (14). A questionnaire was sent to the referring clinician to determine whether the PET scan had altered management and whether overall the PET scan was thought to be helpful. RESULTS: One hundred and eighty nine (80%) questionnaires for 126 patients were returned (63 relating to lymphoma, 62 to CNS tumours, 30 to sarcoma, 16 to plexiform neurofibroma and 18 to other tumours). PET changed disease management in 46 (24%) cases and was helpful in 141 (75%) cases. PET findings were verified by histology, clinical follow-up or other investigations in 141 cases (75%). The returned questionnaires indicated that PET had led to a management change in 20 (32%) lymphoma cases, nine (15%) CNS tumours, four (13%) sarcomas, nine (56%) plexiform neurofibromas and four (22%) cases of other tumours. PET was thought to be helpful in 47 (75%) lymphoma cases, 48 (77%) CNS tumours, 24 (80%) sarcomas, 11 (69%) neurofibromas and 11 (61%) cases of other tumours. PET findings were verified in 44 (70%) lymphoma cases, 53 (85%) CNS tumours, 21 (70%) sarcomas, 12 (75%) neurofibromas and 11 (61%) other tumour cases. CONCLUSION: PET imaging of children with cancer is accurate and practical. PET alters management and is deemed helpful (with or without management change) in a significant number of patients, and the results are comparable with the figures published for the adult oncology population. PMID- 15290127 TI - A cholesterol-containing foreign body granuloma presenting as an inter-metatarsal bursa. AB - A 68-year-old man presented with progressive forefoot swelling which coincided with the onset of type 2 diabetes mellitus. Imaging revealed a cystic inter metatarsal mass containing two foreign bodies, which had been present for many years. Following aspiration of the mass, cholesterol crystals were observed on polarised microscopy. It is postulated that the development of diabetes triggered the shedding of cholesterol crystals around a long-standing quiescent foreign body granuloma. PMID- 15290129 TI - D-hexosaminate production by oxidative fermentation. AB - Microbial production of D-hexosaminate was examined by means of oxidative fermentation with acetic acid bacteria. In most strains of acetic acid bacteria, membrane-bound D-glucosamine dehydrogenase (synonymous with an alternative D glucose dehydrogenase distinct from quinoprotein D-glucose dehydrogenase) oxidized D-hexosamines to the corresponding D-hexosaminates in a stoichiometric manner. Conversion of D-hexosamines to the corresponding D-hexosaminates was observed with growing cells of acetic acid bacteria, and D-hexosaminate was stably accumulated in the culture medium even though D-hexosamine was exhausted. Since the enzyme responsible is located on the outer surface of the cytoplasmic membrane, and the enzyme activity is linked to the respiratory chain of the organisms, resting cells, dried cells, and immobilized cells of acetic acid bacteria were effective catalysts for D-hexosaminate production. D-Mannosaminate and D-galactosaminate were also prepared for the first time by means of oxidative fermentation, and three different D-hexosaminates were isolated from unreacted substrate by a chromatographic separation. In this paper, D-hexosaminate production by oxidative fermentation carried out mainly with Gluconobacter frateurii IFO 3264 is exemplified as a typical example. PMID- 15290130 TI - Protein engineering of toluene ortho-monooxygenase of Burkholderia cepacia G4 for regiospecific hydroxylation of indole to form various indigoid compounds. AB - Previous work showed that random mutagenesis produced a mutant of toluene ortho monooxygenase (TOM) of Burkholderia cepacia G4 containing the V106A substitution in the hydroxylase alpha-subunit (TomA3) that changed the color of the cell suspension from wild-type brown to green in rich medium. Here, DNA shuffling was used to isolate a random TOM mutant that turned blue due to mutation TomA3 A113V. To better understand the TOM reaction mechanism, we studied the specificity of indole hydroxylation using a spectrum of colored TOM mutants expressed in Escherichia coli TG1 and formed as a result of saturation mutagenesis at TomA3 positions A113 and V106. Colonies expressing these altered enzymes ranged in color from blue through green and purple to orange; and the enzyme products were identified using thin-layer chromatography, high performance liquid chromatography, and liquid chromatography-mass spectroscopy. Derived from the single TOM template, enzymes were identified that produced primarily isoindigo (wild-type TOM), indigo (A113V), indirubin (A113I), and isatin (A113H and V106A/A113G). The discovery that wild-type TOM formed isoindigo via C-2 hydroxylation of the indole pyrrole ring makes this the first oxygenase shown to form this compound. Variant TOM A113G was unable to form indigo, indirubin, or isoindigo (did not hydroxylate the indole pyrrole ring), but produced 4 hydroxyindole and unknown yellow compounds from C-4 hydroxylation of the indole benzene ring. Mutations at V106 in addition to A113G restored C-3 indole oxidation, so along with C-2 indole oxidation, isatin, indigo, and indirubin were formed. Other TomA3 V106/A113 mutants with hydrophobic, polar, or charged amino acids in place of the Val and/or Ala residues hydroxylated indole at the C-3 and C-2 positions, forming isatin, indigo, and indirubin in a variety of distributions. Hence, for the first time, a single enzyme was genetically modified to produce a wide range of colors from indole. PMID- 15290132 TI - The effect of decreasing oxygen feed rates on growth and metabolism of Torulaspora delbrueckii. AB - The effect of decreasing oxygen feed rates on the growth and metabolism of Torulaspora delbrueckii and Saccharomyces cerevisiae in chemostat cultures was investigated. The biosynthetic oxygen requirement, i.e. the minimum specific oxygen consumption rate required for steady-state growth at a dilution rate of 0.10 h(-1), of T. delbrueckii was quantified to be less than 0.1 mmol O(2) g(-1) h(-1). Under strict anaerobiosis, washout of T. delbrueckii occurred, whereas for S. cerevisiae it did not. Under oxygen-limited conditions, the increase in fermentative ability of T. delbrueckii with diminishing oxygen supply was less pronounced than that of S. cerevisiae. These results indicate that T. delbrueckii was more disturbed in its energy balance than S. cerevisiae under strict anaerobiosis, and they may explain why T. delbrueckii exhibits poorer growth than S. cerevisiae under this condition. PMID- 15290133 TI - Degradation of estradiol and ethinyl estradiol by activated sludge and by a defined mixed culture. AB - The aerobic degradation of the natural hormone 17-beta-estradiol (E2) and the synthetic hormone 17-alpha-ethinyl estradiol (EE2) was investigated in batch experiments with activated sludge from a conventional and a membrane sewage treatment plant. E2 was converted to estrone (E1), the well known metabolite, and further completely transformed within 3 days. The turnover rates of E2 did not differ greatly between conventional and membrane activated sludge. EE2 was persistent in both sludges. By several transfers into fresh E2-medium an enrichment culture could be selected that used E2 as growth substrate. Further enrichment and isolation led to a defined mixed culture consisting of two strains, which were identified by a polyphasic approach as Achromobacter xylosoxidans and Ralstonia sp., respectively. The culture used E2 and E1 as growth substrates and transformed estriol (E3) and 16-alpha-hydroxyestrone but not the xenoestrogens bisphenol A, alpha-zearalenol, mestranol or EE2. The turnover rates of E2 were 0.025-0.1 microg h(-1) cfu(-1) and did not depend on the steroid concentration. PMID- 15290134 TI - Quorum-sensing antagonist (5Z)-4-bromo-5-(bromomethylene)-3-butyl-2(5H)-furanone influences siderophore biosynthesis in Pseudomonas putida and Pseudomonas aeruginosa. AB - Siderophore synthesis of Pseudomonas putida F1 was found to be regulated by quorum sensing since normalized siderophore production (per cell) increased 4.2 fold with cell density after the cells entered middle exponential phase; similarly, normalized siderophore concentrations in Pseudomonas aeruginosa JB2 increased 28-fold, and a 5.5-fold increase was seen for P. aeruginosa PAO1. Further evidence of the link between quorum sensing and siderophore synthesis of P. putida F1 was that the quorum-sensing-disrupter (5Z)-4-bromo-5 (bromomethylene)-3-butyl-2(5H)-furanone (furanone) from the marine red alga Delisea pulchra was found to inhibit the formation of the siderophore produced by P. putida F1 in a concentration-dependent manner, with 57% siderophore synthesis repressed by 100 microg/ml furanone. In contrast, this furanone did not affect the siderophore synthesis of Burkholderia cepacia G4 at 20-40 microg/ml, and stimulated siderophore synthesis of P. aeruginosa JB2 2.5- to 3.7-fold at 20-100 microg/ml. Similarly, 100 microg/ml furanone stimulated siderophore synthesis in P. aeruginosa PAO1 about 3.5-fold. The furanone appears to interact with the quorum-sensing machinery of P. aeruginosa PAO1 since it stimulates less siderophore synthesis in the P. aeruginosa qscR quorum-sensing mutant (QscR is a negative regulator of LasI, an acylated homoserine lactone synthase). PMID- 15290135 TI - Astaxanthin hyperproduction by Phaffia rhodozyma (now Xanthophyllomyces dendrorhous) with raw coconut milk as sole source of energy. AB - Natural carbon sources, such as those present in cane sugar molasses and grape juice, promote the synthesis of astaxanthin in different Phaffia rhodozyma yeasts. One of these, coconut milk, has a very rich nutrient composition. The aim of this work was to investigate the utility of coconut milk as sole source of energy for astaxanthin pigment production by P. rhodozyma strains. Currently, coconut pulp is widely used in industrial processes in Mexico for the production of shampoos, candies, food, etc. However, coconut milk is a waste product. We show that coconut milk enhances astaxanthin production. The fermentation yielded 850 microg/g yeast with the NRRL-10921 wild-type strain and 1850 microg/g yeast with the mutated R1 strain. Production was better than reported results employing other natural carbon sources. PMID- 15290136 TI - Chemotaxis of Pseudomonas stutzeri OX1 and Burkholderia cepacia G4 toward chlorinated ethenes. AB - The chemotactic responses of Pseudomonas putida F1, Burkholderia cepacia G4, and Pseudomonas stutzeri OX1 were investigated toward toluene, trichloroethylene (TCE), tetrachloroethylene (PCE), cis-1,2-dichloroethylene (cis-DCE), trans-1,2 dichloroethylene (trans-DCE), 1,1-dichloroethylene (1,1-DCE), and vinyl chloride (VC). P. stutzeri OX1 and P. putida F1 were chemotactic toward toluene, PCE, TCE, all DCEs, and VC. B. cepacia G4 was chemotactic toward toluene, PCE, TCE, cis DCE, 1,1-DCE, and VC. Chemotaxis of P. stutzeri OX1 grown on o-xylene vapors was much stronger than when grown on o-cresol vapors toward some chlorinated ethenes. Expression of toluene-o-xylene monooxygenase (ToMO) from touABCDEF appears to be required for positive chemotaxis attraction, and the attraction is stronger with the touR (ToMO regulatory) gene. PMID- 15290139 TI - Biosurfactant from Lactococcus lactis 53 inhibits microbial adhesion on silicone rubber. AB - The ability of biosurfactant obtained from the probiotic bacterium Lactococcus lactis 53 to inhibit adhesion of four bacterial and two yeast strains isolated from explanted voice prostheses to silicone rubber with and without an adsorbed biosurfactant layer was investigated in a parallel-plate flow chamber. The microbial cell surfaces and the silicone rubber with and without an adsorbed biosurfactant layer were characterized using contact-angle measurements. Water contact angles indicated that the silicone-rubber surface with adsorbed biosurfactant was more hydrophilic (48 degrees) than bare silicone rubber (109 degrees). The results showed that the biosurfactant was effective in decreasing the initial deposition rates of Staphylococcus epidermidis GB 9/6 from 2,100 to 220 microorganisms cm(-2) s(-1), Streptococcus salivarius GB 24/9 from 1560 to 137 microorganisms cm(-2) s(-1), and Staphylococcus aureus GB 2/1 from 1255 to 135 microorganisms cm(-2) s(-1), allowing for a 90% reduction of the deposition rates. The deposition rates of Rothia dentocariosa GBJ 52/2B, Candida albicans GBJ 13/4A, and Candida tropicalis GB 9/9 were far less reduced in the presence of the biosurfactant as compared with the other strains. This study constitutes a step ahead in developing strategies to prevent microbial colonization of silicone rubber voice prostheses. PMID- 15290142 TI - Enrichment of chitinolytic microorganisms: isolation and characterization of a chitinase exhibiting antifungal activity against phytopathogenic fungi from a novel Streptomyces strain. AB - Thirteen different chitin-degrading bacteria were isolated from soil and sediment samples. Five of these strains (SGE2, SGE4, SSL3, MG1, and MG3) exhibited antifungal activity against phytopathogenic fungi. Analyses of the 16S rRNA genes and the substrate spectra revealed that the isolates belong to the genera Bacillus or Streptomyces. The closest relatives were Bacillus chitinolyticus (SGE2, SGE4, and SSL3), B. ehimensis (MG1), and Streptomyces griseus (MG3). The chitinases present in the culture supernatants of the five isolates revealed optimal activity between 45 degrees C and 50 degrees C and at pH values of 4 (SSL3), 5 (SGE2 and MG1), 6 (SGE4), and 5-7 (MG3). The crude chitinase preparations of all five strains possessed antifungal activity. The chitinase of MG3 (ChiIS) was studied further, since the crude enzyme conferred strong growth suppression of all fungi tested and was very active over the entire pH range tested. The chiIS gene was cloned and the gene product was purified. The deduced protein consisted of 303 amino acids with a predicted molecular mass of 31,836 Da. Sequence analysis revealed that ChiIS of MG3 is similar to chitinases of Streptomyces species, which belong to family 19 of glycosyl hydrolases. Purified ChiIS showed remarkable antifungal activity and stability. PMID- 15290149 TI - The Nagel anomaloscope: its calibration and recommendations for diagnosis and research. AB - BACKGROUND: The Nagel anomaloscope Model I is the definitive clinical instrument for classifying phenotypic variations in X-linked color-vision disorders. Its system of classification is based on the Rayleigh equation: the relative amounts of red and green primary lights required to match a yellow primary. Our aim was to characterize how changes in mains voltage and ambient temperature influence the wavelength and intensity of each primary and alter the Rayleigh matches of normal and anomalous trichromats. METHODS: A Nagel Model I anomaloscope was calibrated in wavelength and intensity while varying the temperature of its prism housing and the mains voltage. Three normal, three protanomalous and three deuteranomalous trichromats made Rayleigh matches at various temperatures and voltages. RESULTS: The intensities of the green and red primaries show an exponential growth with mains voltage. Additionally, the wavelengths and intensities of all three primaries change with prism housing temperature. As a result, the R-G match midpoints of normal and anomalous trichromats shift with increasing mains voltage, and more markedly with increasing prism housing temperature, to higher R-G settings. CONCLUSIONS: Rayleigh matches obtained with the Nagel I anomaloscope are sensitive to changes in voltage supply and prism housing temperature, arising largely from thermal effects of the internal light sources. However, the instrument may still be safely used for diagnostic and research purposes provided that: (1) a stable voltage supply is used; (2) it is kept at a constant temperature; and (3) the match midpoint of the reference population has been established under identical conditions. PMID- 15290150 TI - Mesopic contrast sensitivity in the presence or absence of glare in a large driver population. AB - BACKGROUND: To evaluate mesopic contrast sensitivity in conditions of glare and no glare in a vehicle driver population, and to explore the effects of age, habitual spectacle correction, photopic visual acuity and driving exposure. METHODS: A cross-sectional study was performed on 297 drivers stratified by age into six groups. The mesopic contrast sensitivity was measured in the absence or presence of glare using the Mesotest II (Oculus, Germany) in each subject both with habitual and best spectacle correction. A questionnaire on the subject's driving habits was completed. RESULTS: There were no significant differences between contrast sensitivity measured with habitual or best spectacle correction. In conditions of no glare, the mesopic contrast sensitivity gradually got worse from 51 to 60 years onwards, and from 41 to 50 years onwards in the presence of glare. In both conditions, the total decrease in contrast sensitivity was 0.3 log units. The with-glare and without-glare mesopic contrast sensitivity improved as photopic visual acuity increased. Forty-five per cent of drivers who reported difficulties in driving at night were unable to perform any of the tests with glare, compared to 20% without glare. However, the effect of driving habits on contrast sensitivity was only significant in the oldest age group. CONCLUSIONS: The mesopic contrast sensitivity and glare sensitivity seem to be stable until the age of 50 years, from which point they start to decline at a rate of 0.1 log contrast sensitivity loss per decade. Drivers with poor visual acuity and/or older drivers who avoided night driving presented worse mesopic contrast sensitivity and greater glare sensitivity. PMID- 15290152 TI - Pars plana vitrectomy with internal limiting membranectomy for refractory diabetic macular edema without a taut posterior hyaloid. AB - BACKGROUND: This is a retrospective study designed to investigate the effect of pars plana vitrectomy (PPV) with internal limiting membrane (ILM) peeling on diabetic macular edema in eyes that do not have a taut hyaloid and have been refractory to standard laser treatment. METHODS: Review of 26 eyes of 20 patients consecutively were treated with PPV with ILM peel for refractory diabetic macular edema. Eyes were included if they had been unresponsive to conventional treatment defined as at least two focal laser applications by a retina specialist. Paired t testing was performed to determine if a change in both optical coherence tomography (OCT)-measured retinal thickness and logarithm of the minimum angle of resolution (logMAR) visual acuity occurred prior to and following PPV with epiretinal membrane vitrectomy. In addition, we performed multivariate regression analysis to determine if any clinical variables predicted a change in visual acuity. RESULTS: The mean age in the sample was 65 years (range 29-81 years). The mean follow-up time was 242 days (range 35-939). Sixteen of the 26 eyes were phakic and the remaining ten were pseudophakic. There was a statistically significant improvement of mean visual acuity from a preoperative logMAR vision of 1.0 to a best postoperative vision of 0.75 (p=0.016, paired t-test). Thirteen (50%) of the 26 eyes gained at least two lines of best-corrected Snellen acuity, three (11.5%) had a decline of at least two lines, and ten (38.5%) showed stable visual acuity. Regression analysis demonstrated that baseline worse visual acuity was the only clinical variable that was associated with improvement in visual acuity (beta=0.602, p=0.016; R (2)=28.7). Fourteen eyes had preoperative and postoperative OCT. Thirteen eyes (93%) had a significant decrease in foveal thickness; with an average preoperative thickness of 575 mum compared to a postoperative average of 311 mum (t=3.65, p=0.002). No surgical complications were observed during the follow-up period. CONCLUSIONS: Surgery for refractory diabetic edema without a taut hyaloid is associated with a significant improvement in visual acuity and diminution of retinal thickness as measured by OCT. Further investigations are warranted to define the role of surgery in the management of persistent diabetic macular edema. PMID- 15290153 TI - Characteristics of patients with a favorable natural course of myopic choroidal neovascularization. AB - OBJECTIVE: To identify the characteristics of patients with myopic choroidal neovascularization (CNV) who had a favorable visual prognosis without treatment. METHODS: We reviewed the medical records of 52 consecutive patients (57 eyes) with myopic CNV who were followed for at least 5 years after the onset of CNV. Clinical characteristics (patient age, CNV size and location, visual acuity at onset, chorioretinal atrophy development around CNV, and degree of myopia) were compared between patients whose visual acuity 5 years after CNV onset was better than 20/40 and those whose visual acuity was worse than 20/200. RESULTS: Among 57 eyes, eight eyes (14.0%; 8 patients) had a final visual acuity better than 20/40. On the other hand, 37 eyes (64.9%; 33 patients) had a final visual acuity worse than 20/200. Statistical analysis revealed that the patients with a good prognosis (final visual acuity better than 20/40) were significantly younger, had significantly smaller CNV, and significantly better initial visual acuity (Mann Whitney U-test, p<0.05). Juxtafoveal CNV was more frequently observed in patients with a good prognosis than in those with a poor prognosis (Fisher's exact probability test, p<0.05). Only one patient (12.5%) in the good prognosis group developed a very limited area of chorioretinal atrophy around the regressed CNV, while 91.9% of the patients in the poor prognosis group developed chorioretinal atrophy. Refractive status and the axial length measurements did not differ between the two groups. CONCLUSIONS: Some young patients with myopic CNV retain favorable vision over the long term without active treatment. These information might be useful to predict the visual outcome of patients with myopic CNV. PMID- 15290154 TI - Safety of posterior juxtascleral depot administration of the angiostatic cortisene anecortave acetate for treatment of subfoveal choroidal neovascularization in patients with age-related macular degeneration. AB - BACKGROUND: Anecortave acetate is a synthetic derivative of cortisol, but very specific and irreversible chemical modifications to the cortisol structure have resulted in the creation of a potent inhibitor of blood vessel growth with no evidence non-clinically or clinically of glucocorticoid receptor-mediated bioactivity. The clinical safety of Anecortave Acetate administered as a posterior juxtascleral depot every 6 months for up to 4 years is reviewed in this manuscript. METHODS: Clinical safety and efficacy of the novel angiostatic agent Anecortave Acetate for Depot Suspension was evaluated in patients with subfoveal exudative age-related macular degeneration (AMD) in a masked, randomized, dose duration clinical trial completed in June 2003. This safety and efficacy study enrolled and treated 128 patients at 18 clinical sites in the US and EU. This was the first clinical trial of Anecortave Acetate for Depot Suspension administered as a posterior juxtascleral depot. Assessments of clinical safety were made with general physical examinations including electrocardiograms and hematology/serum chemistry/urinalysis, detailed ophthalmic evaluations with fluorescein/indocyanine green angiography and assessments of best-corrected logMAR visual acuity. All safety reports have been reviewed periodically by an Independent Safety Committee responsible for overseeing these activities. RESULTS: No clinically relevant safety issues related to either Anecortave Acetate for Depot Suspension or the administration procedure have been identified by an Independent Safety Committee. The most frequent safety issues reported were cataractous changes, decreased visual acuity, ptosis, ocular pain, abnormal vision and subconjunctival hemorrhage, but the majority of these were assessed as unrelated to treatment. CONCLUSIONS: Anecortave Acetate for Depot Suspension (3, 15 and 30 mg) is clinically safe following administration and re-administration at 6-month intervals as a posterior juxtascleral depot using a specially designed curved cannula. PMID- 15290165 TI - Second-trimester genetic amniocentesis: 5-year experience. AB - AIMS: The purpose of this study is to evaluate the data related to the genetic amniocentesis performed in a single university hospital. METHODS: Medical records were used to analyze indications of amniocentesis, the results of chromosome analysis, complications and pregnancy outcomes from January 1998 through January 2002. Anomaly screening was performed to all patients attending to our Obstetrics and Gynecology Department between 16 and 20 weeks of pregnancy. The staff obstetricians-gynecologists performed all of the second-trimester genetic amniocentesis. RESULTS: Totally 2,686 patients attended to our department between 16 and 20 weeks of pregnancy during the index period. A total of 131 genetic amniocentesis were performed. The indications were advanced maternal age (cut off age 35) in 24, suspicion of genetic abnormality on ultrasound in 15, history of siblings with Down syndrome in 2 and abnormal triple screen in 90 patients, respectively. There were two pregnancy losses due to the procedure; thus the overall complication rate of second-trimester genetic amniocentesis was 1.5%. CONCLUSIONS: Being one of the most performed invasive techniques for prenatal diagnose, second-trimester genetic amniocentesis is a reliable and safe method. Although the size of this study is limited, our complication rates are similar to the related literature. PMID- 15290166 TI - Clinical and urodynamic effects of anterior colporraphy and approximation of pubococcygeus muscles in patients with severe cystocele. AB - OBJECTIVE: The objective was to evaluate in a homogeneous series of patients the clinical and urodynamic outcomes following anterior colporraphy and approximation of pubococcygeus muscles for correcting cystocele and genuine stress incontinence (SUI). METHODS: The authors analyzed the pre- and postoperative clinical and urodynamic data of 56 consecutive, not previously operated, patients underwent anterior colporraphy and approximation of pubococcygeus muscles by the same surgeon. The mean follow-up period was 20 months (range 12-32). Statistical analysis was performed with Wilcoxon matched pairs test and Fisher's exact test. RESULTS: Four (8%) patients experienced cystocele recurrence at a lesser degree than preoperatively and 2 (4%) patients developed grade II rectocele postoperatively. Recurrence of cystocele occurred only among those patients undergone sacrospinous colposuspension (4 out of 20 vs. 0 out of 32, P=0.018). Manifested and potential genuine stress incontinence (Pot-SUI) treatment was successful in 18 out of 30 (60%) patients. These patients showed a significant increase (P<0.001) of bladder-to-urethra pressure transmission ratio (PTR) values whereas, in those patients who presented recurrence, such an increase did not occur. CONCLUSION: Anterior colporraphy and approximation of pubococcygeus muscles appears, from our preliminary results to be a safe, effective technique for the primary treatment of cystocele without compromising the other vaginal profiles but should not be considered as the procedure of choice for patients presenting with manifested or Pot-SUI. Long-term follow-up is needed to evaluate whether these results are durable. PMID- 15290167 TI - Meckel's diverticulum complicating pregnancy. Case report and review of the literature. AB - INTRODUCTION: Symptomatic Meckel's diverticulum is a rare entity in pregnancy. The clinical presentation is variable and preoperative diagnosis is hampered by the various anatomical and physiological changes of pregnancy that can obscure serious underlying intra-abdominal pathology. CASE REPORT: We report a 14-year old who presented at 32 weeks' gestation with worsening abdominal pain, distension, and leukocytosis. Abdominal computed tomography showed a pelvic fluid collection with extraluminal air suggestive of a perforated viscus and mechanical small bowel obstruction with features of a closed-loop obstruction. Explorative laparotomy revealed a perforated Meckel's diverticulum. Multiple adhesions between the appendix and the perforated diverticulum, and phlegmon sequelae led to the incarceration of an adjacent segment of ileum. DISCUSSION: Meckel's diverticulum in pregnancy can have serious consequences. There is a high rate of perforation due to delayed diagnosis and surgical intervention. Our report and review of the literature suggest that a high index of clinical suspicion can lead to earlier diagnosis and help to keep maternal and fetal morbidity and mortality to a minimum. PMID- 15290168 TI - Intraepithelial G3 adenocarcinoma of the endometrium after tamoxifen treatment. AB - CASE REPORT: In this paper we describe a case of endometrial carcinoma observed in a post-menopausal patient who was treated with tamoxifen for 5 years after a mastectomy for cancer. She came to our department because of vaginal bleeding 2 years after the end of tamoxifen treatment. TREATMENT: She underwent hysteroscopy and a D and C. A polypoid endometrium completely filled the uterine cavity and was carefully removed by curettage; histology showed a highly undifferentiated neoplasia with a component of serous adenocarcinoma, which was likely to originate from endometrial polyps. OUTCOME: The patient underwent radical hysterectomy, but no residual tumor was found in the uterus or in the tubes, ovary, or pelvic nodes, in spite of its low differentiation grade and high potential aggressiveness, and even though the patient was already symptomatic. Two years after surgery the patient is disease free, which is consistent with the evaluation of the surgical specimen, but unusual in poorly differentiated neoplasms. PMID- 15290170 TI - Thalidomide upregulates macrophage inflammatory protein-1alpha in a herpes simplex virus-induced Behcet's disease-like animal model. AB - The mechanism of action of thalidomide in the treatment of patients with Behcet's disease (BD) is poorly understood. There is some evidence to suggest that certain immunological abnormalities are associated with the pathogenesis of BD. A BD-like mouse model induced by herpes simplex virus (HSV) inoculation shows similar immunological abnormalities. In this study, thalidomide was administered in order to understand the mechanism for the improvement in symptoms in BD-like mice. Eight out of ten thalidomide-treated mice showed improvement but none of ten placebo-treated mice (P < 0.005). The improvements were seen in mucocutaneous symptoms. The mice were sacrificed on the 6th day, and the spleens subjected to RT-PCR, FACS, Western blot and immunohistochemical analysis. IL-2, IL-4, IL-6, IL 10, IFN-gamma, TNFalpha, TGFbeta, MCP-1, RANTES, perforin, IP-10, FasL, FasR and MIP-lalpha were determined. Among these, TNFalpha, MIP-1alpha, perforin and Fas were influenced by thalidomide treatment. These results suggest that thalidomide can attenuate HSV-induced BD-like symptoms in mice through the downregulation of TNFalpha (P < 0.005) and the upregulation of MIP-1alpha (P < 0.005), perforin (P < 0.05) and FasR (P < 0.1). PMID- 15290172 TI - Synovial cysts of proximal tibiofibular joint causing peroneal nerve palsy: report of three cases and review of the literature. AB - INTRODUCTION: Synovial cyst of the proximal tibiofibular joint is a very rare condition, for which there is no consensus regarding treatment. CASE PRESENTATION: We present three patients who had synovial cysts of proximal tibiofibular joint that caused peroneal nerve palsy. We discuss the special features of synovial cysts and review the literature. CONCLUSION: We consider the best treatment of synovial cysts originating from proximal tibiofibular joint and causing peroneal nerve palsy to be total surgical removal as soon as possible after the diagnosis is made. It should be kept in mind that despite surgical treatment the neurological symptoms may not recover. PMID- 15290181 TI - Thoracoscopic repair of diaphragmatic eventration. AB - We report a 6-month-old child who presented with recurrent chest infections associated with a right diaphragmatic eventration. Failure in conservative management lead to thoracoscopic plication at 17 months of age and discharge on the third postoperative day. At one year followup he is completely free from all symptoms, and his chest x-ray demonstrates a marked improvement in the position of the diaphragm. We recommend thoracoscopy as a viable approach in treating this condition in children. PMID- 15290185 TI - Influence of penicillin-induced epileptic activity during pregnancy on postnatal hippocampal nestin expression in rats: light and electron microscopic observations. AB - OBJECTS: Current data concerning the effects of maternal epileptic phenomena on newborns are limited. In clinical practice, therefore, it is difficult to suggest proper guidelines on this issue. This study was carried out to investigate the morphological changes in the hippocampus of newborn pups of rats subjected to experimental epilepsy during pregnancy. METHODS: Eighteen Swiss Albino rats were randomly divided into three groups (n=6): experimental group, saline-injected sham surgery group, and intact control group. In the experimental group of rats, an acute grand mal epileptic seizure was induced by 400 IU penicillin-G administration into their intra-hippocampal CA3 region with a stereotaxic device during the 13th day of their pregnancy. On the first neonatal day, pups were perfused with intracardiac fixative solution under anesthesia, and newborn hippocampi were dissected surgically for light and electron microscopic examinations. In an immunohistochemical study using Rat-401 mono-clonal antibody and peroxidase, nestin expression was analyzed in the developing hippocampal tissue. RESULTS: Histologically, normal migration and hippocampal maturation were determined in the newborn rat hippocampus in the control and the sham-operated groups. It was observed that the morphological structure of hippocampus in the experimental group corresponded to the early embryonal period. Most importantly, it was found that nestin (+)cell density was increased in the experimental epilepsy group in contrast to the control and sham groups. CONCLUSION: It has been concluded that epileptic seizures during embryonic life may cause impaired hippocampal neurogenesis and maturation,explaining the potentially harmful effects of epileptic seizures on the embryo at the early stage of neuronal differentiation. This is the first report regarding the alterations in nestin expression in newborn rat hippocampus. In the light of such findings, it will also be necessary to evaluate the functional consequences of a va-riety of epileptic seizures on learning and memory in neonates. PMID- 15290187 TI - Prenatal diagnosis of a craniopharyngioma: a new case with radical surgery and review. AB - CASE REPORT: A case of the antenatal diagnosis of a craniopharyngioma with radical surgery in the neonatal period is reported. REVIEW OF THE LITERATURE: We have reviewed the literature of such cases in an attempt to isolate specific features in this age group and to determine the appropriate management. Only six cases of the truly antenatal diagnosis of craniopharyngiomas have been reported. Diagnosis has resulted from routine ultrasound during pregnancy or from polyhydramnios. Clinically, there is often macrocephaly due to hydrocephalus or a significant-sized tumor. CONCLUSIONS: Management of these rare cases is controversial with high postoperative mortality and significant morbidity, including panhypopituitarism, visual disturbance, and neuropsychological disorders. From the available literature, no conclusions concerning the management can be drawn at present, due to the rarity of early surgical intervention. Our case, despite the lack of important follow-up, seems to confirm the possibility of attempting radical surgery in the neonatal period as a result of advances both in surgical techniques and in neonatal intensive care. PMID- 15290193 TI - Mutism after evacuation of acute subdural hematoma of the posterior fossa. AB - CASE REPORT: A 7-year-old boy was involved in a road traffic accident. A computed tomography scan revealed an acute subdural hematoma (ASDH) of the posterior fossa, traumatic subarachnoid hemorrhage, and distortion of the brain stem. Removal of the ASDH was completed 3.5 h after injury. After extubation, the patient rapidly recovered consciousness. He was able to follow commands, although he did not speak. He began to utter 14 days after the injury. His speech became normal 39 days after injury. A magnetic resonance imaging scan revealed a post contusional change in the right cerebellum and an ischemic lesion in the pons. DISCUSSION: Immediate removal of the hematoma is the only therapy for patients with ASDH of the posterior fossa. Although any lesions of the dentate nucleus, red nucleus, thalamus, cerebral cortex, and pons, all of which are involved in this case, are able to cause mutism, his mutism was primarily caused by the severe ASDH of the posterior fossa. The transient nature of this syndrome suggests that the cause of the mutism is trauma-related edema and/or transient ischemia of these structures. PMID- 15290195 TI - [Intraindividual comparison of higher order aberrations after implantation of aspherical and spherical IOLs depending on pupil diameter]. AB - BACKGROUND: The implantation of aspherical IOLs with negative spherical aberration should equalize the positive spherical aberration of the cornea. The aim of our study was the intraindividual comparison of higher order aberrations (HOA) after implantation of an aspherical and spherical IOL. PATIENTS AND METHODS: In ten patients we randomly implanted an aspherical IOL in one eye and a spherical IOL in the other. After 3 months, we used a Hartmann-Shack sensor to measure total HOA, 3rd to 5th order as well as spherical aberration Z4.0, in mydriasis. We compared all aberrations intraindividually for pupil diameters of 3.5-6 mm. RESULTS: For both IOLs, all aberrations rose with increasing pupil diameter. However, after implantation of the aspherical IOL total HOA, 4th order and spherical aberration Z4.0 were lower, for 4th order aberration and Z4.0 even significantly. For 3rd and 5th order aberrations, there was no difference between both IOLs. CONCLUSIONS: With implantation of aspherical IOLs, total HOA, 4th order and spherical aberration Z4.0 could be reduced compared to spherical IOLs. PMID- 15290196 TI - [Extra-ocular muscle surgery using combined topical and subconjunctival anesthesia]. AB - A retrospective study of a consecutive series of 284 cases of strabismus surgery under local anesthesia is presented. A total of 284 operations on the extraocular muscles, performed with topical and subconjunctival application of local anesthetics-in some cases combined with anesthesiological sedation and monitoring were analyzed retrospectively for adverse effects or complications caused by the type of anesthesia chosen. No ophthalmological complications caused by the mode of anesthesia and no vital complications occurred. In three cases, the intended surgery could not be fully completed due to discomfort of the patient caused by pain perception. In this case series, combined topical and subconjunctival anesthesia in strabismus surgery had a low frequency of complications. PMID- 15290197 TI - [The Ahmed glaucoma valve. Medium term results]. AB - BACKGROUND: In complicated glaucoma, when classical filtrating surgery would be ineffective, aqueous shunts may be used. Complications due to hypotonia are reduced by valved systems, such as the Ahmed glaucoma valve (AGV). METHOD: In a retrospective case control study, 28 patients with complicated glaucoma were included. In addition to the clinical examination, we examined the size and function of the filtering area using ultrasound. RESULTS: The medium term follow up was 25+/-16 months, the preoperative intraocular pressure (IOP) 35.5 mmHg+/ 10.3 while 17 eyes were pseudophakic and nine aphakic. In the first weeks after AGV implantation, the mean IOP was 6.3+/-2.5 mmHg. In nine eyes, the pressure was less than 5 mmHg and five developed a temporary choroidal detachment. At the last visit, IOP was regulated in 22 eyes (82.1%). There was no correlation between IOP regulation and the size of the filtering bleb or the increase in the latter by digital pressure. CONCLUSION: In the management of complicated glaucoma, if there is a high risk of failure due to conjunctival scarring, AGV implantation can be used as a save procedure with a success rate comparable to other glaucoma implants. PMID- 15290198 TI - [Blinking activity during visual display terminal work. 2: reduced blinking and therapeutic approaches]. AB - New findings based on a noninvasive, automated long-term measurement method revealed interindividual differences in lid movement behavior, existence of blinking patterns, and the dominance of cognitive influence in the regulation of blinking frequency during increased concentration and especially visual attention. The development of an individual blinking animation promises long lasting increase and harmonization of lid movements during visual display work. Maintenance of the integrity of the ocular surface by preventing surface evaporation and providing sufficient precorneal environment eradicates important pathogenic factors of ocular discomfort. An animation program for stimulation of blinking has been developed. First results showed that an increase in blinking rate initiated by the computer itself is feasible in principle during work at a visual display terminal. Further improvement of this new approach is promising. PMID- 15290200 TI - [Evaluation of portable TGDc-01 tonometers and comparison with the Goldmann applanation tonometer]. AB - PURPOSE: The aim of this study was to evaluate patients' acceptance and intraocular pressure (IOP) readings of a new digital mobile tonometer (TGDc-01) and compare it to Goldmann applanation tonometry. METHOD: Measurements repeated five times with the TGDc-01 and three times with Goldmann tonometry were performed in 100 eyes of 100 patients by two independent investigators. Patients' acceptance of both techniques was evaluated by a visual analogue scale (VAS). RESULTS: The mean IOP with the TGDc-01 yielded 15.4 mmHg for investigator 1 and 12.7 mmHg for investigator 2 (range: 4-43 mmHg). Results of the measurements with Goldmann tonometry showed 17.6 mmHg for investigator 1 and 17.3 mmHg for investigator 2 (9-42 mmHg). The IOP difference of the two tonometry methods was highly significant (p<0.001). The intraobserver variability was 29% for investigator 1 and 8% for investigator 2. Mean IOP values of the two investigators taken with the TGDc-01 differed significantly (p<0.01) from each other by a mean of 2.6 mmHg. CONCLUSIONS: The new mobile tonometer TGDc-01 is better accepted by patients but IOP values are significantly lower compared to Goldmann tonometry and variability is high. Regarding glaucoma diagnostics it seems to be less suitable than Goldmann tonometry. PMID- 15290201 TI - [Non-contact donor cornea trephination with a flying spot excimer laser system]. AB - BACKGROUND: One of the most common complications after mechanical penetrating keratoplasty is the occurrence of irregular astigmatism. We developed a method to prepare donor grafts for transplantation applying excimer laser techniques. The quality of the cut edges and exposure time were evaluated. MATERIALS AND METHODS: The beam of an experimental excimer laser (OPTex, Lambda Physik) was homogenized and guided by an optical system including lenses and scanner mirrors. A special software was used to conduct the laser beam in a circular mode over the surface of the cornea. Corneas from porcine eyes were stabilized in an artificial anterior chamber. Twenty in vitro explants were trephinated applying our method and subsequently investigated by electron microscopy and histology. RESULTS: All experiments provided regular and smooth cut edges. The average exposure time until perforation was 10.5 min (SD 2.7 min), the diameter of the grafts was 8.0 mm. CONCLUSION: The experiments showed the possibility of non-contact trephination and generation of smooth cut edges with a rotating focussed excimer laser beam in a porcine cornea model. A major advantage of the system is the possibility of customized "tailored" grafts. PMID- 15290202 TI - Preoperative imaging in renal cell cancer. AB - Renal cell cancer (RCC) represents the fifth most common cancer in men, with a rising incidence. Radical cancer surgery remains the only curative treatment in localized and advanced RCC. Therefore, preoperative imaging is most important for the planning of the surgical approach and strategy. The aim of any preoperative imaging in RCC is to differentiate benign from malignant lesions, to adequately assess tumor size, localization and organ confinement, to identify lymph node and/or visceral metastases, and to reliably predict the presence and extent of any thrombus of the vena cava. It is our aim to review the current status of preoperative imaging modalities in RCC. Computed tomography (CT) remains the most appropriate imaging modality to differentiate benign from malignant lesions. Although RCC can appear as iso-, hyper- or hypodense lesions on native CT scans, it usually demonstrates a significant contrast enhancement of about 115 HU and intratumoral areas of necrosis following the intravenous application of contrast medium. Benign masses such as renal oncocytoma are most often homogenous lesions exhibiting hypodensity compared to the normal renal parenchyma following the i.v. application of contrast dye. CT accurately predicts the tumor size with only a 0.5 cm difference as compared to the pathological size of the lesion. The identification of lymph node metastases still remains a problem since the limiting size is 4 mm and CT will result in a false negative rate of about 10%, especially in the presence of micrometastases; the false positive rate of 3-43% is mainly due to reactive hyperplasia. New technologies, such as the multidetector CT with thin collimation and multiplanar reformatting, might result in a diagnostic improvement. The involvement of the adrenal gland can be accurately predicted by CT scans or MRI, allowing an adrenal sparing approach in the case of unsuspicious findings. The detection of visceral metastases appears to be crucial since it has been shown that even patients with metastatic disease might benefit from radical nephrectomy followed by systemic immunotherapy in the case of a good performance status, and the presence of lymph node and pulmonary metastases only. Involvement of the renal vein and the vena cava with tumor thrombus formation will change the surgical strategy. Preoperatively, the presence and the cranial extent of the thrombus need to be known in order to plan the surgical approach. With regard to the extent of renal vein thrombi, a three phase helical CT scan is most appropriate; for vena caval thrombi only a MRI examination is able to accurately identify any infra- or suprahepatic as well as intracardial extension of the thrombus. The identification of multifocal lesions remains another unsolved problem in preoperative imaging techniques for RCC. Compared to the pathohistological analysis of nephrectomy specimens, neither ultrasonography, color duplex sonography nor regular CT scans are able to identify multifocal lesions with acceptable sensitivity and specificity. The evaluation of unenhanced CT scans together with the enhanced corticomedullary and the nephrogenic phase result in a 100% sensitivity and might represent a valuable option. Angiography has basically been abandoned from the armory of routine imaging techniques. It has, however, a current role in terms of the embolization of large tumors to reduce intraoperative blood loss, and in the palliative management of pain and bleeding due to RCC not amenable to surgery. Finally, we present a diagnostic algorithm for the most informative imaging techniques in the evaluation of RCC. PMID- 15290203 TI - Imaging for kidney stones. AB - Imaging investigations play a vital role in the management of patients with kidney stones. The techniques available include plain x-ray of the abdomen, ultrasound scan, intravenous urogram, computed tomography (CT) and magnetic resonance imaging, amongst others. All of these techniques have their own individual roles to play and also have limitations. CT has been establishing itself as the imaging technique of choice and some exciting developments are on the way. However, renal stone disease is a complex condition. Furthermore, there are a variety of surgical techniques used to treat stones. It is therefore important that the strengths and weakness of each of the modalities are clearly understood and the investigations are tailored to address the problem in hand. PMID- 15290204 TI - The role of imaging in urinary tract infections. AB - The aim of imaging in urinary tract infections (UTI) is to detect conditions that must be corrected to avoid imminent deterioration of kidney function, or to prevent recurrent infections and long-term kidney damage. The most threatening conditions are obstruction of an infected upper tract and abscesses of the genitourinary system. An image-guided percutaneous drainage can be lifesaving. The role of imaging in small children with UTI is controversial in terms of the importance of anatomical and functional disorders in relation to the preventive measures to be taken. In newborns identified with hydronephrosis during pregnancy or by neonatal screening, vesicoureteral reflux (VUR) and renal scarring are congenital and not caused by infection. Most of these patients are males and the VUR is of a higher grade than VUR detected in girls after the first UTI. Imaging in children with UTI should only be ordered after a thorough evaluation of the risk of renal damage and the benefits of preventive measures. In adult females, no imaging is necessary in cystitis, while ultrasonography and plain films are recommended in acute pyelonephritis. Since uncomplicated UTI in men is rare, diagnostic imaging should be started early to rule out complicating factors in the urinary tract. In prostatitis, vesiculitis, epididymitis and orchitis the role of imaging is to rule out abscess formation and testicular malignancies. PMID- 15290205 TI - Sonography of the bladder. AB - Ultrasonography (US) is the method of choice for the diagnosis of bladder disease. It is superior to other imaging techniques, such as urography and cystography, in depicting certain structures and abnormalities. US examination of the bladder should include a study of the ureterovesical junction and the structures round the vesical neck. The examination technique may be transabdominal, transrectal or transvaginal, or transurethral. The bladder pathology that can be studied by US includes cystitis, calculi, clots, diverticula, trauma and tumors. The sensitivity and the specificity of the method are very high and sometimes superior to cystoscopy. Sonography can be used to explore patients with stress incontinence and those with abdominal trauma. The ureterovesical junction may be clearly examined by US and the pathology of the papilla clearly defined. PMID- 15290206 TI - Acid-base and electrolyte disorders after urinary diversion. AB - The use of bowel in urologic reconstructive procedures may result in numerous short and long-term complications, including well-described acid-base and electrolyte disorders. Many of these metabolic alterations are influenced by how solute absorption occurs across the particular bowel segment chosen for reconstruction. Solute absorption is impacted by: (1) the segment of bowel used, (2) the surface area of bowel used, (3) the time of retention of urine, (4) the concentration of solutes in the urine, (5) renal function, and (6) the pH and osmolality of the urine. These factors affect the type and amount of solutes absorbed, as well as the severity of metabolic complications that develop. PMID- 15290207 TI - Mucus production after transposition of intestinal segments into the urinary tract. AB - Following transposition into the urinary tract, intestinal segments continue to produce mucus and problems related to excessive production do not to diminish with time. Currently, 20 human mucin genes have been described and their protein products partially or fully characterised. As the use of transposed intestinal segments in urology increases, there is now a need for a better understanding of mucins at the gene and protein levels. There is also a need for urologists to be aware of the many complications related to excess mucus production. Whilst effective therapeutic measures to reduce mucus production and its related complications remain elusive, it is now clear that without such effective mucoregulatory agents the quality of life of patients will continue to be less than satisfactory. This review describes the biology of mucus and the problems related to continued production following transposition of intestinal segments into the urinary tract. Difficulties related to quantification of urinary mucus, in addition to the structural and mucin gene changes that occur in transposed segments, are addressed. PMID- 15290227 TI - Expression of P2 receptors at sites of chronic inflammation. AB - Extracellular nucleotides have been identified as important signaling molecules. These nucleotides act on the P2 family of receptors that respond by either forming an ion-channel or by activation of a signal transduction cascade, both of which enable a cellular response. Although a role for P2 receptors in inflammation has been implied, the local expression pattern and kinetics of these receptors at sites of inflammation are not known. Therefore, we have studied the expression of the P2 receptors expressed by inflammatory cells or by cells in the vasculature, with special attention of P2X(1), P2X(7)R, P2Y(1)R, and P2Y(2)R. As a suitable model for studying inflammatory reactions, we have employed the foreign body reaction (FBR), a sterile inflammatory reaction induced by implanting degradable cross-linked dermal sheep collagen disks subcutaneously in the rat. We show that, in the vasculature, the express of P2X(7)R, P2Y(1)R and P2Y(2)R increase until day 2. The expression of P2X(7)R and P2Y(1)R on macrophages and giant cells increased during the course of the inflammatory reaction which was studied for 21 days. The expression of the P2Y(2)R on macrophages and giant cells inside the foreign body increases with time, whereas the expression on macrophages in the surrounding tissue is maximal at day 5. The expression of P2X(1)R remains at a constant low level. The upregulation of P2X(7)R, P2Y(1)R, and P2Y(2)R over time suggests a regulatory function for these receptors in inflammation. PMID- 15290234 TI - Dynamics of PAF-induced conjunctivitis reveals differential expression of PAF receptor by macrophages and eosinophils in the rat. AB - The action of platelet-activating factor (PAF) toward the PAF receptor (PAF-R) plays an important role in inflammation. We employed immunohistochemistry and quantitative confocal immunofluorescence microscopy to examine the dynamic changes of PAF-R expression in the conjunctiva in response to PAF-induced conjunctivitis in Brown Norway rats within the first 2 h after topical administration of PAF. Instillation of PAF caused an alteration in pattern of recruitment of macrophages and eosinophils into the conjunctiva, as was visualized by immunohistochemical staining for the antigens ED 1 and ED 2 (markers of macrophages) and MBP (a marker of eosinophils). An increase in the number of PAF-R positive cells was alos detected. Quantitative colocalization analysis revealed ther strongest rise in the degree of PAF-R expression by macrophages within the first 6 h, whereas their infiltration increased throughout the period of observation. However, eosinophils showed a high degree of PAF-R expression during all 24 h of the experiment, although they infiltrated strongly only within the first 2 h. Thus, for the first time, the use of quantitative colocalization analysis software developed by us has reveal intrinsic details of the interaction of PAF anf PAF-R in conjunctivitis, findings not otherwise obtainable by using qualitative approaches alone. Our results provide a theoretical basis for a definition of the proper time-frame in which to prevent and control the infiltration of macrophages and eosinophils into the conjunctiva. PMID- 15290237 TI - The origin and spread of the HFE-C282Y haemochromatosis mutation. AB - The mutation responsible for most cases of genetic haemochromatosis in Europe (HFE C282Y) appears to have been originated as a unique event on a chromosome carrying HLA-A3 and -B7. It is often described as a "Celtic mutation"- originating in a Celtic population in central Europe and spreading west and north by population movement. It has also been suggested that Viking migrations were largely responsible for the distribution of this mutation. Two, initial estimates of the age of the mutation are compatible with either of these suggestions. Here we examine the evidence about HFE C282Y frequencies, extended haplotypes involving HLA-A and -B alleles, the validity of calculations of mutation age, selective advantage and current views on the relative importance of "demic diffusion" (population migration) and "adoption-diffusion" (cultural change) in the neolithic transition in Europe and since then. We conclude that the HFE C282Y mutation occurred in mainland Europe before 4,000 BC. PMID- 15290238 TI - Genomic rearrangements at the IGHMBP2 gene locus in two patients with SMARD1. AB - Autosomal recessive spinal muscular atrophy with respiratory distress type 1 (SMARD1) is caused by mutations in the immunoglobulin mu-binding protein 2 (IGHMBP2) gene. Patients affected by the infantile form of SMARD1 present with early onset respiratory distress. So far, patients with neither juvenile onset nor with larger deletions/rearrangements in IGHMBP2 have been reported. In this study, we investigated one patient with infantile (4 months) and another with juvenile (4.3 years) onset of respiratory distress. Direct sequencing of all exons and flanking intron sequences in both patients revealed a mutation on only one allele. In both patients, we identified genomic rearrangements of the other allele of IGHMBP2 by means of Southern blotting. Putative breakpoints were confirmed by polymerase chain reaction on genomic and cDNA. The patient with juvenile onset had an Alu/Alu mediated rearrangement, which resulted in the loss of aproximately 18.5 kb genomic DNA. At the mRNA level, this caused an in-frame deletion of exons 3-7. The patient with infantile onset had a complex rearrangement with two deletions and an inversion between intron 10 and 14. This rearrangement led to a frameshift at the mRNA level. Our results show that SMARD1 can be caused by genomic rearrangements at the IGHMBP2 gene locus. This may be missed by mere sequence analysis. Additionally, we demonstrate that juvenile onset SMARD1 may also be caused by mutations of IGHMBP2. The complex nature of the genomic rearrangement in the patient with infantile SMARD1 is discussed and a deletion mechanism is proposed. PMID- 15290239 TI - Concurrent analysis of loss of heterozygosity (LOH) and copy number abnormality (CNA) for oral premalignancy progression using the Affymetrix 10K SNP mapping array. AB - Like most human cancers, oral squamous cell carcinoma (SCC) is characterized by genetic instabilities. In this study, a single platform (Affymetrix 10K SNP mapping array) was used to generate both loss of heterozygosity (LOH) and DNA copy number abnormality (CNA) read-outs for precise and high-resolution genetic alteration profiles. As a proof of principle, we performed this concordant analysis on a panel of deletion and trisomy cell lines with known chromosomal alterations and the precise LOH and CNA regions were detected as expected. Using a previously described oral SCC progression model system, we identified a set of genomic regions that may be associated with the malignancy progression, including chromosome regions 3pter-3p35.3, 3p14.1-3p13, 11p, 11q14.3-11q22.2, and 11q13.5 11q14.1. These data show that it is feasible to utilize high-density SNP arrays to generate concordant LOH and CNA profiles at high resolution. PMID- 15290247 TI - The first report of Hepatozoon spp. (Apicomplexa, Hepatozoidae) in domestic cats from Sao Paulo state, Brazil. AB - Hepatozoon sp. was diagnosed in three naturally infected cats from Sao Paulo state, Brazil. The first animal was admitted to the veterinary clinic with renal failure. During the hematological examination, gamonts of Hepatozoon sp. were observed within polymorphonuclear cells. Another two cats, which lived in the same house as the first cat, were also positive for this hemoparasite. This is the first report of a Hepatozoon sp. infection in domestic cats from Brazil. PMID- 15290250 TI - The use of tetraphyll as food for snails increases the intensity of cercarial shedding in Galba truncatula infected with Fasciola hepatica. AB - Experimental infections of Galba truncatula with Fasciola hepatica were carried out under laboratory conditions to determine why the use of Tetraphyll, when provided every 3 days as food for snails, stimulated the intensity of cercarial shedding. In snails raised on cos lettuce and Tetraphyll, the number of metacercariae was significantly higher than that recorded in controls on lettuce only. In the former group, numerous metacercariae were counted on day 1 of every 3-day period (when snails fed on Tetraphyll), while the numbers of these larvae strongly decreased on day 2 and were very low on day 3. The repetition of this experiment using microalgae or modified Boray's diet every 3 days as food for snails gave the same results. However, the number of metacercariae recorded on day 1 of every 3-day period was significantly higher in snails raised on lettuce and Tetraphyll than in those reared on lettuce and algae (or Boray's diet). According to the authors, the first contact of an infected snail with Tetraphyll induces a faster differentiation of cercariae within the rediae and the accumulation of many free cercariae in the snail's hemocoel, so that a second contact of the snail with this food 3 days after allows free cercariae to complete their glycogen and fat reserves and to rapidly exit from the snail. However, a temporary immobilization of the infected snail by Tetraphyll, or rather its degradation products, and a massed exit of cercariae cannot be excluded. PMID- 15290254 TI - The role of B- and T-cell immunity in toltrazuril-treated C57BL/6 WT, microMT and nude mice experimentally infected with Neospora caninum. AB - Neospora caninum is a protozoan parasite predominantly known for causing abortion in cattle and neuromuscular disease in dogs. So far, no efficient metaphylactic chemotherapy has been developed. In preliminary studies, toltrazuril had been successfully used against experimental neosporosis in mice and calves. In the present study, we used immunocompetent and immunodeficient mouse strains to address the role of immunity in supporting the chemotherapy of experimental N. caninum infection. WT, microMT and athymic nude mice were intraperitoneally inoculated with 1x10(6) Nc-1 tachyzoites. The drug was administered in the drinking water for 6 consecutive days so as to obtain a daily dose of approximately 20 mg toltrazuril/kg body weight. The course of infection was monitored by clinical, histological and immunohistochemical means, as well as by the search for parasite DNA using PCR-analyses of various organs. In immunocompetent WT mice, treatment proved to be of high efficacy by abrogation of any lesion formation or PCR-positivity in medicated C57BL/6 mice and a significant reduction of lesion formation or PCR-positivity in BALB/c animals. Similarly, treated microMT mice exhibited a significant reduction in cerebral lesion formation as well as in parasite DNA detectability by PCR when compared to untreated animals. Athymic nude mice, however, did not respond to treatment in that only a delay of the parasite dissemination was achieved, and nude mice still showed the neosporosis disease symptoms, although later than untreated animals. We conclude that treatment with toltrazuril appears to act parasitostatically rather than parasitocidically. This is supported by the fact that: (1) although the lack of B-cells did not impair the effect of toltrazuril, (2) the lack of T cells did not allow for a full efficacy of treatment. Therefore, chemotherapy with toltrazuril against experimental infections with N. caninum requires the support of T-cell immunity in order to be successful. PMID- 15290259 TI - Percutanous closure of patent ductus arteriosus in small infants of less than 8 kg body weight using different devices. PMID- 15290260 TI - Diagnostic evaluation for asymmetry: consider genetic mosaicism. PMID- 15290262 TI - Inherited multicentric osteolysis with carpal-tarsal localisation mimicking juvenile idiopathic arthritis. AB - Five patients with multicentric carpal-tarsal osteolysis are presented: a mother and her three children with an autosomal dominant mode of inheritance and one of the children with nephropathy, the fifth a sporadic case also with renal involvement. The main findings common to these five patients are symptoms and signs simulating arthritis of the wrists and/or ankles starting at a young age and mimicking juvenile idiopathic arthritis. Early signs of osteolysis and shortening of the carpus or tarsus are radiological characteristic. The disease may be associated with a peculiar face, but most importantly with nephropathy. The pathogenesis is still unknown. CONCLUSION: Recognition of this disease and differentiation from juvenile idiopathic arthritis is important to avoid unnecessary investigations and treatment. Follow-up of renal function is indicated. PMID- 15290263 TI - Irritable bowel syndrome: the commonest cause of recurrent abdominal pain in children. AB - Apley, working in Bristol, UK, defined recurrent abdominal pain (RAP) in 1958. After extensive investigations, he found that 8% of children presenting to his clinic with RAP had an organic pathology. The aims of this study were to identify (1) causes of RAP using modern methodology, (2) factors associated with organic RAP and (3) children with non-organic RAP who fulfill the diagnostic criteria for irritable bowel syndrome (IBS). Children, aged over 3 years, presenting with RAP were prospectively recruited to this study. They had a detailed questionnaire completed, a full examination with screening tests (blood for coeliac screen, Helicobacter pylori antibody titre, inflammatory markers, serum amylase, liver function tests, and full blood count, urine and stool analyses and abdominal ultrasonography). Endoscopy and oesophageal pH monitoring were performed if clinically indicated. IBS was diagnosed if the child had no organic pathology and fulfilled the Rome II criteria. Out of 103 children (median age of 10 years, mean 10.04, SD +/-3.44), 31 children (30%) had organic pathologies. Factors associated with organic pain were nocturnal symptoms (P<0.01) and abdominal tenderness (P<0.005) and with non-organic pain were periumbilical locality (P<0.002), pain alleviation on defaecation (P<0.04) and low fibre diet (P<0.005). Of children with non-organic pain, 37/52 (51%) fulfilled the criteria for IBS (36% of the total). CONCLUSION: Of children presenting with recurrent abdominal pain in a hospital setting, 30% have a diagnosable organic aetiology compared to 8% in Apley's time. Irritable bowel syndrome, however, may be the commonest cause of recurrent abdominal pain and should be considered. PMID- 15290264 TI - Hearing impairment in familial X-linked hypophosphatemic rickets. PMID- 15290265 TI - Prognostic factors for mortality of oesophageal atresia patients: Waterston revived. PMID- 15290266 TI - Leukaemia presenting with fulminant hepatic failure in a child. PMID- 15290267 TI - Fluoride content of mineral waters on the Belgian market and a case report of fluorosis induced by mineral water use. PMID- 15290268 TI - The infrequent bowel movements in young infants who are exclusively breast-fed. PMID- 15290291 TI - Distribution of cell wall components in Sphagnum hyaline cells and in liverwort and hornwort elaters. AB - Spiral secondary walls are found in hyaline cells of Sphagnum, in the elaters of most liverworts, and in elaters of the hornwort Megaceros. Recent studies on these cells suggest that cytoskeletal and ultrastructural processes involved in cell differentiation and secondary wall formation are similar in bryophytes and vascular plant tracheary elements. To examine differences in wall structure, primary and secondary wall constituents of the hyaline cells of Sphagnum novo zelandicum and elaters of the liverwort Radula buccinifera and the hornwort Megaceros gracilis were analyzed by immunohistochemical and chemical methods. Anti-arabinogalactan-protein antibodies, JIM8 and JIM13, labeled the central fibrillar secondary wall layer of Megaceros elaters and the walls of Sphagnum leaf cells, but did not label the walls of Radula elaters. The CCRC-M7 antibody, which detects an arabinosylated (1-->6)-linked beta-galactan epitope, exclusively labeled hyaline cells in Sphagnum leaves and the secondary walls of Radula elaters. Anti-pectin antibodies, LM5 and JIM5, labeled the primary wall in Megaceros elaters. LM5 also labeled the central layer of the secondary wall but only during formation. In Radula elaters, JIM5 and another anti-pectin antibody, JIM7, labeled the primary wall. The distribution of arabinogalactan-proteins and pectic polysaccharides restricted to specific wall types and stages of development provides evidence for the developmental and functional regulation of cell wall composition in bryophytes. Monosaccharide-linkage analysis of Sphagnum leaf cell walls suggests they contain polysaccharides similar to those of higher plants. The most abundant linkage was 4-Glc, typical of cellulose, but there was also evidence for xyloglucans, 4-linked mannans, 4-linked xylans and rhamnogalacturonan-type polysaccharides. PMID- 15290270 TI - Chronic mucocutaneous candidiasis and primary hypothyroidism in two families. AB - We describe the clinical and immunological features of two families with chronic mucocutaneous candidiasis (CMC) and primary hypothyroidism. Family A includes three siblings with both candidiasis and hypothyroidism and four individuals with hypothyroidism only. Family B includes four members with candidiasis, of whom one (a male child) also had hypothyroidism. All individuals affected with CMC had suffered from oral candidiasis and onychomycosis since infancy. Facial seborrhoic dermatitis, general folliculitis and scaling blepharitis were main manifestations. Hypothyroidism became evident during childhood. No thyroid antibodies were present in the affected siblings in family A, while the male in family B with hypothyroidism had antibodies against thyroid peroxidase at diagnosis. Immunological evaluation revealed intra-individual variations in serum immunoglobulin levels, lymphocyte subsets and proliferative responses, but there were no consistent abnormalities. Vaccine responses were normal. AIRE gene region microsatellite markers did not segregate with disease nor were autoantibodies typical for autoimmune polyendocrine syndrome type 1 detected in the families. CONCLUSION: The link between hypothyroidism and chronic mucocutaneous candidiasis remains to be identified. PMID- 15290293 TI - Structural and biochemical bases of photorespiration in C4 plants: quantification of organelles and glycine decarboxylase. AB - In C(4) plants, photorespiration is decreased relative to C(3) plants. However, it remains unclear how much photorespiratory capacity C(4) leaf tissues actually have. We thoroughly investigated the quantitative distribution of photorespiratory organelles and the immunogold localization of the P protein of glycine decarboxylase (GDC) in mesophyll (M) and bundle sheath (BS) cells of various C(4) grass species. Specific differences occurred in the proportions of mitochondria and peroxisomes in the BS cells (relative to the M cells) in photosynthetic tissues surrounding a vein: lower in the NADP-malic enzyme (NADP ME) species having poorly formed grana in the BS chloroplasts, and higher in the NAD-malic enzyme (NAD-ME) and phosphoenolpyruvate carboxykinase (PCK) species having well developed grana. In all C(4) species, GDC was localized mainly in the BS mitochondria. When the total amounts of GDC in the BS mitochondria per unit leaf width were estimated from the immunogold labeling density and the quantity of mitochondria, the BSs of NADP-ME species contained less GDC than those of NAD ME or PCK species. This trend was also verified by immunoblot analysis of leaf soluble protein. There was a high positive correlation between the degree of granal development (granal index) in the BS chloroplasts and the total amount of GDC in the BS mitochondria. The variations in the structural and biochemical features involved in photorespiration found among C(4) species might reflect differences in the O(2)/CO(2) partial pressure and in the potential photorespiratory capacity of the BS cells. PMID- 15290294 TI - Isolation and characterization of PHYC gene from Stellaria longipes: differential expression regulated by different red/far-red light ratios and photoperiods. AB - We have cloned and characterized the phytochrome C ( PHYC) gene from Stellaria longipes. The PHYC gene is composed of a 110-bp 5'-untranslated leader sequence, a 3,342-bp coding region, and a 351-bp 3'-untranslated sequence. The Stellaria PHYC contains three long introns within the coding region at conserved locations as in most angiosperm PHY genes. DNA blot analysis indicates that the Stellaria genome contains a single copy of PHYC. Stellaria PHYC shares 60%, 58%, and 57% deduced amino acid identities with rice, Sorghum, and Arabidopsis PHYC, respectively. Phylogenetic analysis indicates that Stellaria PHYC is located in the dicot branch, but is divergent from Arabidopsis PHYC. The Stellaria PHYC is constitutively expressed in different plant organs, though the level of PHYC gene transcript in roots is slightly higher than in flowers, leaves, and stems. When 2 week old seedlings grown in the dark were exposed to constant white light, PHYC mRNA quickly accumulates within 1-12 h. When plants grown in darkness for 7 days were exposed to different red/far-red light (R/FR) ratios, the levels of PHYC mRNA at R/FR = 0.7 are much lower than under R/FR = 3.5. The levels of PHYC mRNA under short-day (SD) photoperiod are higher than under long-day (LD) photoperiod. Plants under SD conditions do not elongate, and are only about 1.7 cm tall at 19 days. In contrast, plants under LD conditions elongate with an average height of 21.2 cm at 19 days. The plants do not flower under SD conditions, but do so at 18 19 days under LD conditions. These results indicate that under SD conditions the high level of PHYC mRNA may inhibit stem elongation and flower initiation. In contrast, under LD conditions the high level of PHYC mRNA may promote stem elongation and flowering. PMID- 15290295 TI - Activities of fructan- and sucrose-metabolizing enzymes in wheat stems subjected to water stress during grain filling. AB - This study investigated if a controlled water deficit during grain filling of wheat (Triticum aestivum L.) could accelerate grain filling by facilitating the remobilization of carbon reserves in the stem through regulating the enzymes involved in fructan and sucrose metabolism. Two high lodging-resistant wheat cultivars were grown in pots and treated with either a normal (NN) or high amount of nitrogen (HN) at heading time. Plants were either well-watered (WW) or water stressed (WS) from 9 days post anthesis until maturity. Leaf water potentials markedly decreased at midday as a result of water stress but completely recovered by early morning. Photosynthetic rate and zeatin + zeatin riboside concentrations in the flag leaves declined faster in WS plants than in WW plants, and they decreased more slowly with HN than with NN when soil water potential was the same, indicating that the water deficit enhanced, whereas HN delayed, senescence. Water stress, both at NN and HN, facilitated the reduction in concentration of total nonstructural carbohydrates (NSC) and fructans in the stems but increased the sucrose level there, promoted the re-allocation of pre-fixed (14)C from the stems to grains, shortened the grain-filling period, and accelerated the grain filling rate. Grain weight and grain yield were increased under the controlled water deficit when HN was applied. Fructan exohydrolase (FEH; EC 3.2.1.80) and sucrose phosphate synthase (SPS; EC 2.4.1.14) activities were substantially enhanced by water stress and positively correlated with the total NSC and fructan remobilization from the stems. Acid invertase (EC 3.2.1.26) activity was also enhanced by the water stress and associated with the change in fructan concentration, but not correlated with the total NSC remobilization and (14)C increase in the grains. Sucrose:sucrose fructosyltransferase (EC 2.4.1.99) activity was inhibited by the water stress and negatively correlated with the remobilization of carbon reserves. Sucrose synthase (EC 2.4.1.13) activity in the stems decreased sharply during grain filling and showed no significant difference between WW and WS treatments. Abscisic acid (ABA) concentration in the stem was remarkably enhanced by water stress and significantly correlated with SPS and FEH activities. Application of ABA to WW plants yielded similar results to those for WS plants. The results suggest that the increased remobilization of carbon reserves by water stress is attributable to the enhanced FEH and SPS activities in wheat stems, and that ABA plays a vital role in the regulation of the key enzymes involved in fructan and sucrose metabolism. PMID- 15290297 TI - Developmental analyses reveal early arrests of the spore-bearing parts of reproductive organs in unisexual flowers of cucumber (Cucumis sativus L.). AB - To understand the regulatory mechanisms governing unisexual flower development in cucumber, we conducted a systematic morphogenetic analysis of male and female flower development, examined the dynamic changes in expression of the C-class floral organ identity gene CUM1, and assessed the extent of DNA damage in inappropriate carpels of male flowers. Accordingly, based on the occurrence of distinct morphological events, we divided the floral development into 12 stages ranging from floral meristem initiation to anthesis. As a result of our investigation we found that the arrest of stamen development in female flowers, which occurs just after the differentiation between the anther and filament, is mainly restricted to the primordial anther, and that it is coincident with down regulation of CUM1 gene expression. In contrast, the arrest of carpel development in the male flowers occurs prior to the differentiation between the stigma and ovary, given that no indication of ovary differentiation was observed even though CUM1 gene expression remained detectable throughout the development of the stigma like structures. Although the male and female reproductive organs have distinctive characteristics in terms of organ differentiation, there are two common features regarding organ arrest. The first is that the arrest of the inappropriate organ does not affect the entirety of the organ uniformly but occurs only in portions of the organs. The second feature is that all the arrested portions in both reproductive organs are spore-bearing parts. PMID- 15290298 TI - Recruitment of fibre types and quadriceps muscle portions during repeated, intense knee-extensor exercise in humans. AB - To investigate recruitment of slow-twitch (ST) and fast-twitch (FT) muscle fibres, as well as the involvement of the various quadriceps femoris muscle portions during repeated, intense, one-legged knee-extensor exercise, 12 healthy male subjects performed two 3-min exercise bouts at approximately 110% maximum thigh O2 consumption (EX1 and EX2) separated by 6 min rest. Single-fibre metabolites were determined in successive muscle biopsies obtained from the vastus lateralis muscle (n = 6) and intra-muscular temperatures were continuously measured at six quadriceps muscle sites (n = 6). Creatine phosphate (CP) had decreased (P < 0.05) by 27, 73 and 88% in ST fibres and 25, 71 and 89% in FT fibres after 15 and 180 s of EX1 and after 180 s of EX2, respectively. CP was below resting mean-1 SD in 15, 46, 84 and 100% of the ST fibres and 9, 48, 85 and 100% of the FT fibres at rest, after 15 and 180 s of EX1 and after 180 s of EX2, respectively. A significant muscle temperature increase (deltaTm) occurred within 2-4 s at all quadriceps muscle sites. DeltaTm varied less than 10% between sites during EX1, but was 23% higher (P < 0.05) in the vastus lateralis than in the rectus femoris muscle during EX2. DeltaTm in the vastus lateralis was 101 and 109% of the mean quadriceps value during EX1 and EX2, respectively. We conclude that both fibre types and all quadriceps muscle portions are recruited at the onset of intense knee-extensor exercise, that essentially all quadriceps muscle fibres are activated during repeated intense exercise and that metabolic measurements in the vastus lateralis muscle provide a good indication of the whole-quadriceps muscle metabolism during repeated, intense, one-legged knee extensor exercise. PMID- 15290299 TI - Acidic ATP activates lymphocyte outwardly rectifying chloride channels via a novel pathway. AB - Using whole-cell patch-clamp techniques we found that ATP activated an outwardly rectifying current in Daudi human B lymphoma cells under acidic conditions. The substitution of Cl- for gluconate(-) shifted the reversal potential, while Cl- channel blockers, 4,4'-diisothiocyanostibene-2,2'-disulfonic acid (DIDS) and 9 anthracene carboxylic acid (9-AC), blocked the current, indicating that ATP induces this current by activating the outwardly rectifying chloride channel (ORCC). The effect of ATP on ORCC was mimicked by ADP, but not by other P2 receptor agonists such as ATPgammaS (a poorly hydrolyzable analog of ATP), 2',3' O-benzoyl-4-benzoyl-ATP (BzATP), and UTP. The ATP-induced ORCC current was completely blocked by 100 microM suramin (a P2 receptor antagonist), and was partially blocked by 100 microM pyridoxal-phosphate-6-azophenyl-2',4'-disulfonic acid tetrasodium (PPADS), which is another P2 receptor antagonist. Neither inactivation of G proteins nor elimination of extracellular Ca2+ affected the ATP induced current, indicating that G protein-coupled P2Y receptors and Ca(2+) permeable P2X receptors are not involved. Based on the pharmacological profile and the fact that acidic conditions are required for ATP to activate the ORCC, we suggest that acidic ATP activates the lymphocyte ORCC via a novel pathway, which is not associated with any previously described purinergic receptors. PMID- 15290300 TI - Exercise training enhanced the expression of myocardial proteins related to cell protection in spontaneously hypertensive rats. AB - Exercise training could potentially exert beneficial effects on the signaling events associated with cardiac cell apoptosis. Spontaneously hypertensive rats (SHR) were trained 5 days/week on a treadmill (18 m/min for 120 min/day) between the ages of 4 weeks and 1 week, corresponding to the hypertensive accelerating phase. The effect of exercise training on the expression of anti-apoptotic proteins HSP-72, Bcl-2 and protein kinase B (PKB), and the apoptotic proteins Bax and glycogen synthase kinase-3 (GSK-3) was examined. Exercise had a significant acute lowering effect on blood pressure, but this decrease did not attenuate the progressive increase in blood pressure. In the left ventricles of exercised SHR, PKB phosphorylation of both Ser473 and Thr308 residues was significantly increased by 166% and 120%, respectively, compared to sedentary SHR. PKB phosphorylation significantly correlated with GSK-3beta phosphorylation. HSP-72 and Bcl-2 protein expression were increased in the left ventricle of exercised SHR, and associated with the concomitant increased expression of the protein Bax. Thus, the Bcl-2/Bax ratio was not changed by exercise training, suggesting that the anti-apoptotic mechanism was effective in compensating the increase in the expression of the pro-apoptotic protein Bax in the myocardium of the SHR. PMID- 15290301 TI - Electrophysiological characterization of the tetrodotoxin-resistant Na+ channel, Na(v)1.9, in mouse dorsal root ganglion neurons. AB - Small dorsal root ganglion neurons express preferentially the Na+ channel isoform Na(v)1.9 that mediates a tetrodotoxin-resistant (TTX-R) Na+ current. We investigated properties of the Na+ current mediated by Na(v)1.9 (I(NaN)) using the whole-cell, patch-clamp recording technique. To isolate I(NaN) from heterogeneous TTX-R Na+ currents that also contain another type of TTX-R Na+ current mediated by Na(v)1.8, we used Na(v)1.8-null mutant mice. When F- was used as an internal anion in the patch pipette solution, both the activation and inactivation kinetics for I(NaN) shifted in the hyperpolarizing direction with time. Such a time-dependent shift of the kinetics was not observed when Cl- was used as an internal anion. Functional expression of I(NaN) declined with time after cell dissociation and recovered during culture, implying that Na(v)1.9 may be regulated dynamically by trophic factors or depend on subtle environmental factors for its survival. During whole-cell recordings, the peak amplitude of I(NaN) increased dramatically after a variable delay, as if inactive or silent channels had been "kindled". Such an unusual increase of the amplitude could be prevented by adding ATP to the pipette solution or by recording with the nystatin perforated patch-clamp technique, suggesting that the rupture of patch membrane affected the behaviour of Na(v)1.9. These peculiar properties of I(NaN) may provide an insight into the plasticity of Na+ channels that are related to pathological functions of Na+ channels accompanying abnormal pain states. PMID- 15290302 TI - Direct effects of 9-anthracene compounds on cystic fibrosis transmembrane conductance regulator gating. AB - Anthracene-9-carboxylic acid (9-AC) has been reported to show both potentiation and inhibitory effects on guinea-pig cardiac cAMP-activated chloride channels via two different binding sites, and inhibition of Mg(2+)-sensitive protein phosphatases has been proposed for the mechanism of 9-AC potentiation effect. In this study, we examined the effects of 9-AC on wild-type and mutant human cystic fibrosis transmembrane conductance regulator (CFTR) chloride channels expressed in NIH3T3 or CHO cells. 9-AC inhibits whole-cell CFTR current in a voltage dependent manner, whereas the potentiation effect is not affected by membrane potentials. Anthracene-9-methanol, an electro-neutral 9-AC analog, fails to block CFTR, but shows a nearly identical potentiation effect, corroborating the idea that two chemically distinct sites are responsible, respectively, for potentiation and inhibitory actions of 9-AC. 9-AC also enhances the activity of deltaR-CFTR, a constitutively active CFTR mutant whose R-domain is removed. In excised inside-out patches, 9-AC increases Po by prolonging the mean burst durations and shortening the interburst durations. We therefore conclude that two different 9-AC binding sites for potentiation and inhibitory effects on CFTR channels are located outside of the R-domain. We also speculate that 9-AC potentiates CFTR activity by directly affecting CFTR gating. PMID- 15290304 TI - Nifedipine and diltiazem suppress ventricular arrhythmogenesis and calcium release in mouse hearts. AB - Ventricular arrhythmogenesis leading to sudden cardiac death remains responsible for significant mortality in conditions such as cardiac failure and the long-QT syndrome (LQTS). Arrhythmias may be accentuated by beta-adrenergic stimulation and, accordingly, the present study explored the possible effects of beta adrenergic stimulation and L-type Ca(2+) channel blockade on ventricular arrhythmogenesis and Ca(2+) handling using the mouse heart as an experimental system. Studies in whole, Langendorff-perfused hearts using programmed electrical stimulation protocols adapted from clinical practice demonstrated sustained ventricular tachycardia following addition of 0.1 microM isoprenaline (n=15), whilst no arrhythmias were observed in the absence of the drug (n=15). Arrhythmias were suppressed by nifedipine or diltiazem pre-treatment (both 1 microM) (n=8 and 4 respectively) and were also induced by elevating external [Ca(2+)] (n=3). At the cellular level, 0.1 microM isoprenaline significantly increased normalized fluorescence (F/F(0)) in field-stimulated fluo-3-loaded mouse ventricular myocytes imaged using confocal microscopy, reflecting increases in sarcoplasmic reticulum Ca(2+) release (n=8). Elevated external [Ca(2+)] also increased F/F(0) (n=4) whilst 0.1 microM nifedipine or 0.1 microM diltiazem significantly decreased F/F(0) (n=13 and 6 respectively). Pre-treatment with 0.1 microM nifedipine or 0.1 microM diltiazem suppressed the increases in F/F(0) induced by 0.1 microM isoprenaline alone (n=14 and 6 respectively). The findings thus paralleled suppression of isoprenaline-induced arrhythmias seen with nifedipine or diltiazem at the whole-heart level. Taken together, the findings may have implications for the use of L-type Ca(2+) channel blockade in conditions associated with beta-adrenergically driven ventricular arrhythmias such as cardiac failure and LQTS. PMID- 15290303 TI - Prostaglandins and nitric oxide in regional kidney blood flow responses to renal nerve stimulation. AB - We examined the roles of cyclooxygenase products and of interactions between the cyclooxygenase and nitric oxide systems in the mechanisms underlying the relative insensitivity of medullary perfusion to renal nerve stimulation (RNS) in anaesthetized rabbits. To this end we examined the effects of ibuprofen and N(G) nitro-L: -arginine (L-NNA), both alone and in combination, on the responses of regional kidney perfusion to RNS. Under control conditions, RNS produced frequency-dependent reductions in total renal blood flow (RBF; -82+/-3% at 6 Hz), cortical laser-Doppler flux (CLDF; -84+/-4% at 6 Hz) and, to a lesser extent, medullary laser-Doppler flux (MLDF; -46+/-7% at 6 Hz). Ibuprofen did not affect these responses significantly, suggesting that cyclooxygenase products have little net role in modulating renal vascular responses to RNS. L-NNA enhanced RBF (P=0.002), CLDF (P=0.03) and MLDF (P=0.03) responses to RNS. As we have shown previously, this effect of L-NNA was particularly prominent for MLDF at RNS frequencies < or = 1.5 Hz. Subsequent administration of ibuprofen, in L-NNA pretreated rabbits, did not affect responses to RNS significantly. We conclude that counter-regulatory actions of NO, but not of prostaglandins, partly underlie the relative insensitivity of medullary perfusion to renal nerve activation. PMID- 15290323 TI - Halobacterium noricense sp. nov., an archaeal isolate from a bore core of an alpine Permian salt deposit, classification of Halobacterium sp. NRC-1 as a strain of H. salinarum and emended description of H. salinarum. AB - Two rod-shaped haloarchaeal strains, A1 and A2, were isolated from a bore core from a salt mine in Austria. The deposition of the salt is thought to have occurred during the Permian period (225-280 million years ago). The 16S rDNA sequences of the strains were 97.1% similar to that of the type species of the genus Halobacterium, which was also determined in this work. Polar lipids consisted of C20-C20 derivatives of phosphatidylglycerol, methylated phosphatidylglycerol phosphate, phosphatidylglycerol sulfate, triglycosyl diether and sulfated tetraglycosyl diether. Optimal salinity for growth was 15-17.5% NaCl; Mg++ was tolerated up to a concentration of 1 M. The DNA-DNA reassociation value of strain A1T was 25% with H. salinarum DSM 3754T and 41% with Halobacterium sp. NRC-1, respectively. Based on these results and other properties, e.g. whole cell protein patterns, menaquinone content and restriction patterns of DNA, strains A1 and A2 are members of a single species, for which we propose the name H. noricense. The type strain is A1 (DSM 15987T, ATCC BAA-852T, NCIMB 13967T). Since we present evidence that Halobacterium sp. NRC-1 is a member of H. salinarum, an emended description of H. salinarum is provided. PMID- 15290324 TI - Digestion by serine proteases enhances salt tolerance of glutaminase in the marine bacterium Micrococcus luteus K-3. AB - Salt-tolerant glutaminase (Micrococcus glutaminase, with an apparent molecular mass of 48.3 kDa, intact glutaminase) from the marine bacterium Micrococcus luteus K-3 was digested using protease derived from M. luteus K-3. The digestion products were a large fragment (apparent molecular mass of 38.5 kDa, the glutaminase fragment) and small fragments (apparent molecular mass of 8 kDa). The digestion was inhibited by phenylmethanesulfonyl fluoride (PMSF). Digestion of intact glutaminase by serine proteases including trypsin, elastase, lysyl endopeptidase, and arginylendopeptidase also produced the glutaminase fragment. The N-terminus of the glutaminase fragment was the same as that of intact glutaminase. The N-termini of two small fragments were Ala394 and Ala396, respectively. The enzymological and kinetic properties of the glutaminase fragment were almost the same as those of intact glutaminase except for salt tolerant behavior. The glutaminase fragment was a higher salt-tolerant enzyme than the intact glutaminase, suggesting that Micrococcus glutaminase is digested in the C-terminal region by serine protease from M. luteus K-3 to confer salt tolerance on glutaminase. PMID- 15290326 TI - Bifunctional phosphoglucose/phosphomannose isomerase from the hyperthermophilic archaeon Pyrobaculum aerophilum. AB - ORF PAE1610 from the hyperthermophilic crenarchaeon Pyrobaculum aerophilum was first annotated as the conjectural pgi gene coding for hypothetical phosphoglucose isomerase (PGI). However, we have recently identified this ORF as the putative pgi/pmi gene coding for hypothetical bifunctional phosphoglucose/phosphomannose isomerase (PGI/PMI). To prove its coding function, ORF PAE1610 was overexpressed in Escherichia coli, and the recombinant enzyme was characterized. The 65-kDa homodimeric protein catalyzed the isomerization of both glucose-6-phosphate and mannose-6-phosphate to fructose-6-phosphate at similar catalytic rates, thus characterizing the enzyme as bifunctional PGI/PMI. The enzyme was extremely thermoactive; it had a temperature optimum for catalytic activity of about 100 degrees C and a melting temperature for thermal unfolding above 100 degrees C. PMID- 15290325 TI - Archaeal elongation factor 1alpha from Sulfolobus solfataricus interacts with the eubacterial antibiotic GE2270A. AB - The thiazolyl-peptide antibiotic GE2270A, an inhibitor of the elongation factor Tu from Escherichia coli (EcEF-Tu), was used to study the effects produced in the biochemical properties of the archaeal functional analogue elongation factor 1alpha from Sulfolobus solfataricus (SsEF-1alpha). GE2270A did not substantially affect the poly(U)-directed-polyPhe incorporation catalyzed by SsEF-1alpha and the formation of the ternary complex SsEF-1alpha.GTP.Phe-tRNAPhe. On the other hand, the antibiotic was able to increase the GDP/GTP exchange rate of SsEF 1alpha; nevertheless, this improvement was not associated with an increase in the catalytic activity of the enzyme. In fact, GE2270A inhibited both the intrinsic GTPase of SsEF-1alpha (GTPaseNa) and that stimulated by ribosomes. Interestingly, GTPaseNa of both intact and C-terminal-deleted SsEF-1alpha resulted in a greater sensitivity to the antibiotic with respect to SsEF-1alpha lacking both the M- and C-terminal domains. This result suggested that, similar to what is found for EcEF Tu, the M domain of SsEF-1alpha is the region of the enzyme most responsible for the interaction with GE2270A. The different behavior observed in the inhibition of protein synthesis with respect to EcEF-Tu can be ascribed to the different adaptive structural changes that have occurred in SsEF-1alpha during evolution. PMID- 15290334 TI - Polyamines and cancer: minireview article. AB - The naturally occurring polyamines, spermine, spermidine and the diamine putrescine are widespread in nature. They have been implicated in growth and differentiation processes. Polyamines accumulate in cancerous tissues and their concentration is elevated in body fluids of cancer patients. Assays of urinary and blood polyamines have been used to detect cancer and to determine the success of therapy. Drugs which inhibit the synthesis of polyamines can prevent cancer and may also be used for therapeutic purposes. Ornithine decarboxylase, which catalyzes the rate limiting step in polyamine synthesis, can serve as a marker of proliferation. Recently, a new in vitro chemosensitivity test, based on the disappearance of ornithine decarboxylase in drug-treated cancer cells has been developed. The increasing interest in polyamines and their physiological functions may lead to a more extensive application of these compounds or their derivatives in cancer diagnosis and treatment. PMID- 15290327 TI - A thermostable manganese-containing superoxide dismutase from the thermophilic fungus Thermomyces lanuginosus. AB - A thermostable superoxide dismutase (SOD) from a Thermomyces lanuginosus strain (P134) was purified to homogeneity by fractional ammonium sulfate precipitation, ion-exchange chromatography on DEAE-Sepharose, Phenyl-Sepharose hydrophobic interaction chromatography, and gel filtration on Sephacryl S-100. The molecular mass of a single band of the enzyme was estimated to be 22.4 kDa, using sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Using gel filtration on Sephacryl S-100, the molecular mass was estimated to be 89.1 kDa, indicating that this enzyme was composed of four identical subunits of 22.4 kDa each. The SOD was found to be inhibited by NaN3, but not by KCN or H2O2, suggesting that the SOD in T. lanuginosus was of the manganese superoxide dismutase type. The SOD exhibited maximal activity at pH 7.5. The optimum temperature for the activity was 55 degrees C. It was thermostable at 50 and 60 degrees C and retained 55% activity after 60 min at 70 degrees C. The half-life of the SOD at 80 degrees C was approximately 28 min and even retained 20% activity after 20 min at 90 degrees C. PMID- 15290335 TI - Mammary histidine decarboxylase vulnerability to enzyme antisense oligonucleotides: histamine and polyamine systems cross-talk. AB - Histamine system is suggested to have a role in mammary gland growth regulation, differentiation and functioning during pregnancy and lactation. Histidine decarboxylase activity undergoes significant changes during pregnancy and lactation. Pregnancy associated elevation of HDC activity and mRNA transcript in mouse mammary gland was successfully affected by enzyme antisense oligonucleotides treatment. The enzyme activity of resting mammae was unaffected as it lacked inducible pool of HDC. The short-term mammary histamine shortage evoked influenced the mRNA expression of histamine receptors (H1 and H2) and ornithine decarboxylase during pregnancy. There were essentially no morphological changes in the mammary gland upon the treatment, however, adipocytes neighbouring alveolar structures were more pronounced. These findings further substantiate the role of histamine in mammary gland physiology and emphasise presence of common motifs of biogenic amines and polyamine metabolism as well as mutual interferences implicating observed "cross-talk" phenomenon. PMID- 15290336 TI - How important is the oxidative degradation of spermine?: minireview article. AB - Spermine is a constituent of most eucaryotic cells, however, it is not of vital importance for the vertebrate organism, as is demonstrated by the existence of transgenic (Gy) mice that lack spermine and spermine synthase. In contrast its degradation appears to be of vital importance, since mice die after chronic administration of N1,N4-bis(2,3-butadienyl)-1,4-butanediamine (MDL 72517). Under this condition spermine accumulates in red blood cells and blood plasma. Lethal toxicity can be avoided by intervals of MDL 72527-free periods. During these periods spermine appears to be directly degraded to spermidine without an intermediary acetylation step within the red blood cells. Since this reaction is of enormous physiological significance, it will be important to characterise the red blood cell spermine oxidase, and it will be particularly important to determine whether this oxidase is identical with the FAD-dependent polyamine oxidase that is considered to be involved in the polyamine interconversion sequence, or whether it is one of the recently characterised spermine oxidase isoenzymes. PMID- 15290337 TI - Arginine pathways and the inflammatory response: interregulation of nitric oxide and polyamines: review article. AB - An early response to an acute inflammatory insult, such as wound healing or experimental glomerulonephritis, is the conversion of arginine to the cytostatic molecule nitric oxide (NO). This 'anti-bacterial' phase is followed by the conversion of arginine to ornithine, which is the precursor for the pro proliferative polyamines as well as proline for the production of extracellular matrix. This latter, pro-growth phase constitutes a 'repair' phase response. The temporal switch of arginine as a substrate for the cytostatic iNOS/NO axis to the pro-growth arginase/ ornithine/polyamine and proline axis is subject to regulation by inflammatory cytokines as well as interregulation by the arginine metabolites themselves. Arginine is also the precursor for another biogenic amine, agmatine. Here we describe the capacity of these three arginine pathways to interregulate, and propose a model whereby agmatine has the potential to serve in the coordination of the early and repair phase pathways of arginine in the inflammatory response by acting as a gating mechanism at the transition from the iNOS/NO axis to the arginase/ODC/polyamine axis. Due to the pathophysiologic and therapeutic potential, we will further examine the antiproliferative effects of agmatine on the polyamine pathway. PMID- 15290338 TI - Protein-polyamine conjugation by transglutaminase in cancer cell differentiation: review article. AB - Considerable and intense progress has been made in the understanding of the chemistry, molecular biology and cell biology of transglutaminases (TGases: EC 2.3.2.13). The knowledge that very different processes such as cell growth, reproduction and death are dependent on the presence of adequate levels of these enzymes and that the amount of both free and protein-conjugated polyamines, formed by the enzyme, are capable of modulating the differentiation and proliferative capability of several cell types, has prompted a multitude of researchers to study the role of these fascinating molecules in cancer cell differentiation. PMID- 15290339 TI - Biogenic amines and apoptosis: minireview article. AB - The programmed cell death is a very complex mechanism involving many factors, among them the intracellular concentration of biogenic amines (BA) appears to be important for apoptosis triggering. The mitochondrial damage is imputable to hydrogen peroxide and aldehydes, produced by amine oxidases (AO)-mediated oxidation of BA. On the other hands, the apoptosis protection observed by high BA concentration appears to be related to their scavenger effect of ROS and/or their interaction with membrane pores. Also monoamine oxidase (MAO) inhibitors, like propargylamines, preserve the mitochondria integrity by inhibiting MAO and therefore the production of H2O2 and aldehydes and, as cations, by regulating membrane pores, like BA. As general conclusion, apoptosis is protected by high concentration of BA and/or other cations while it is favoured by ROS produced by AOs or other mechanisms. PMID- 15290340 TI - Arginine revisited: minireview article. AB - Arginine is a precursor of proteins and employed in urea synthesis. It is also the precursor of many other compounds, such as creatine, nitric oxide, polyamines, agmatine, proline. In this review, its transport and that of other basic amino acids are examined, along with its transformation into nitric oxide, agmatine and proline, and the mutual regulation of the individual pathways. PMID- 15290341 TI - Inhibitors of polyamine metabolism: review article. AB - The identification of increased polyamine concentrations in a variety of diseases from cancer and psoriasis to parasitic infections has led to the hypothesis that manipulation of polyamine metabolism is a realistic target for therapeutic or preventative intervention in the treatment of certain diseases. The early development of polyamine biosynthetic single enzyme inhibitors such as alpha difluoromethylornithine (DFMO) and methylglyoxal bis(guanylhydrazone) showed some interesting early promise as anticancer drugs, but ultimately failed in vivo. Despite this, DFMO is currently in use as an effective anti-parasitic agent and has recently also been shown to have further potential as a chemopreventative agent in colorectal cancer. The initial promise in vitro led to the development and testing of other potential inhibitors of the pathway namely the polyamine analogues. The analogues have met with greater success than the single enzyme inhibitors possibly due to their multiple targets. These include down regulation of polyamine biosynthesis through inhibition of ornithine decarboxylase and S adenosylmethionine decarboxylase and decreased polyamine uptake. This coupled with increased activity of the catabolic enzymes, polyamine oxidase and spermidine/spermine N1-acetyltransferase, and increased polyamine export has made the analogues more effective in depleting polyamine pools. Recently, the identification of a new oxidase (PAO-h1/SMO) in polyamine catabolism and evidence of induction of both PAO and PAO-h1/SMO in response to polyamine analogue treatment, suggests the analogues may become an important part of future chemotherapeutic and/or chemopreventative regimens. PMID- 15290342 TI - The transglutaminase family: an overview: minireview article. AB - The knowledge that very different processes such as normal and neoplastic cell growth, reproduction and death are dependent on the presence of adequate levels of transglutaminases (TGase: EC 2.3.2.13) and that they are capable of affecting the differentiation and proliferative capability of several cell types, has prompted a multitude of researchers to study these fascinating molecules. In the following overview we intend to summarize the currently known information on the biological significance of these enzymes. PMID- 15290343 TI - The multifaceted role of transglutaminase in neurodegeneration: review article. AB - A critical role for transglutaminase [TGase] has been hypothesized in the pathogenesis of the CAG trinucleotide repeat diseases, characterized by proteins with abnormal expansions of a polyglutamine domain. In the last few years the involvement of TGase in neurodegenerative diseases [NDS], including its role in aggregate formation, has been broadened to include Alzheimer's [AD] and Parkinson's Disease [PD]. It is clear that reduction of TGase activity is beneficial for prolonged survival in mouse models of NDS. The pathological progression of these diseases might reflect in part increases of TGase induced aggregates, or changes in other pathways influenced by increases in TGase activity. Neurodegeneration may be influenced by increased TGase activity affecting apoptosis, modulation of GTPase activity and signal transduction. This review will focus on the leading hypotheses in relation to both old and new experimental results. PMID- 15290344 TI - Transglutaminase 2 in celiac disease: minireview article. AB - Celiac disease (CD) is an autoimmune pathology of the small intestine triggered, in genetically predisposed patients, by the exposition to gliadin, a flour protein, thus evoking local immune reactions and mucosal atrophy. The discovery that type 2 transglutaminase (TG2) is the main, if not the sole, target of the endomysium CD-specific autoantibodies assigned to this enzyme a master regulator role of CD. Two separated events, both based on the finding that gliadin is able to act as a TG2 substrate, have been described to indicate that TG2 is involved in both the humoral and cellular immune responses. In this paper we review the novel insights on the localization and enzymatic activity of TG2 in the small intestinal mucosa. Moreover, we report on the capability of gliadin and its peptides to act as TG2 substrates. PMID- 15290345 TI - Tissue transglutaminase in normal and abnormal wound healing: review article. AB - A complex series of events involving inflammation, cell migration and proliferation, ECM stabilisation and remodelling, neovascularisation and apoptosis are crucial to the tissue response to injury. Wound healing involves the dynamic interactions of multiple cells types with components of the extracellular matrix (ECM) and growth factors. Impaired wound healing as a consequence of aging, injury or disease may lead to serious disabilities and poor quality of life. Abnormal wound healing may also lead to inflammatory and fibrotic conditions (such as renal and pulmonary fibrosis). Therefore identification of the molecular events underlying wound repair is essential to develop new effective treatments in support to patients and the wound care sector. Recent advances in the understating of the physiological functions of tissue transglutaminase a multi functional protein cross-linking enzyme which stabilises tissues have demonstrated that its biological activities interrelate with wound healing phases at multiple levels. This review describes our view of the function of tissue transglutaminase in wound repair under normal and pathological situations and highlights its potential as a strategic therapeutic target in the development of new treatments to improve wound healing and prevent scarring. PMID- 15290346 TI - Overexpressed transglutaminase 5 triggers cell death. AB - Transglutaminases are a class of nine different proteins involved in many biological phenomena such as differentiation, tissue repair, endocytosis. Transglutaminase 5 was originally cloned from skin keratinocytes, and a partial biochemical characterization showed its involvement in skin differentiation. Here we demonstrate that transglutaminase 5 is able to induce cell death when intracellularly overexpressed. Transfected cells show enzymatic activity, as demonstrated by fluoresceincadaverine staining. Transfected cells died due to the formation of hypodiploid DNA content, indicating the induction of cell death under these pharmacological conditions. We also show that the primary sequence of transglutaminase 5 contains GTP binding domains which are similar to those in transglutaminase 2. This raises the possibility that transglutaminase 5 is regulated by GTP in a similar fashion to transglutaminase 2. PMID- 15290347 TI - Translational and post-translational modifications of proteins as a new mechanism of action of alpha-interferon: review article. AB - Interferon-alpha (IFNalpha) is a recombinant protein widely used in the therapy of several neoplasms such as myeloma, renal cell carcinoma, epidermoid cervical and head and neck tumours and melanoma. IFNalpha, the first cytokine to be produced by recombinant DNA technology, has emerged as an important regulator of cancer cell growth and differentiation, affecting cellular communication and signal transduction pathways. However, the way by which tumour cell growth is directly suppressed by IFNalpha is not well known. Wide evidence exists on the possibility that cancer cells undergo apoptosis after the exposure to the cytokine. Here we will discuss data obtained by us and others on the post translational regulation of the expression of proteins involved in the occurrence of apoptotic process such as tissue transglutaminase (tTG) or in the modulation of cell cycle such as the cyclin-dependent kinase inhibitor p27. This new way of regulation of p27 and tTG occurs through the modulation of their proteasome dependent degradation induced by the cytokine. We will also review the involvement of protein synthesis machinery in the induction of cell growth inhibition by IFNalpha. In details, we will describe the effects of IFNalpha on the expression and activity of the protein kinase dependent from dsRNA (PKR) and on the eukaryotic initiation factor of protein synthesis 5A (eIF-5A) and their correlations with the regulation of cancer cell growth. These data strongly suggest that the antitumour activity of IFNalpha against human tumours could involve still unexplored mechanisms based on post-translational and translational control of the expression of proteins that regulate cell proliferation and apoptosis. PMID- 15290348 TI - Producing transglutaminases by molecular farming in plants: minireview article. AB - Transglutaminases have a range of catalytic activities, most of which concern the post-translational modification of proteins. The most important of these activities, both in terms of biology and biotechnology, is the cross-linking of proteins into large supramolecular networks. The widespread use of transglutaminases in research, medicine and industry has increased the demand for an inexpensive, efficient and safe source of recombinant enzymes. We describe initial results concerning the production of a mammalian transglutaminase in transgenic rice plants as a first step towards the large-scale molecular farming of this enzyme. PMID- 15290349 TI - Transglutaminase 5 is acetylated at the N-terminal end. AB - Transglutaminases (TGases) are calcium-dependent enzymes that catalyse cross linking between proteins by acyl transfer reaction; they are involved in many biological processes including coagulation, differentiation, and tissue repair. Transglutaminase 5 was originally cloned from keratinocytes, and a partial biochemical characterisation showed its involvement in skin differentiation, in parallel to TGase 1 and TGase 3. Here, we demonstrate, by electrospray tandem mass spectrometry that TGase 5 is acetylated at the N-terminal end. Moreover, in situ measurement of TGase activity shows that endogenous TGase 5 is active upon treatment with phorbol acetate, and the enzyme co-localises with vimentin intermediate filaments. PMID- 15290350 TI - Biochemical mechanisms for a possible involvement of the transglutaminase activity in the pathogenesis of the polyglutamine diseases: minireview article. AB - Transglutaminases are a family of enzymes which show the common capacity to catalyse the cross-linking of protein substrates. Some members of this family of enzymes are also capable to catalyse other chemical reactions for the cell life. The distribution and the role of these enzymes have been studied in numerous cell types and tissues, but only recently their expression and functions started to be investigated in the Nervous System. One of the main biochemical properties of the Transglutaminase enzymes is to form large protein aggregates that are insoluble in all known protein detergents. Recently, the Transglutaminase activity has been hypothesised to be involved in the pathogenetic mechanisms responsible for the formation of cellular inclusions present in the Corea Major and in other polyglutamine diseases. In this review we describe the biochemical mechanisms by which the Transglutaminases could play a critical role in the physiopathology of the polyglutamine diseases. PMID- 15290351 TI - Acetylation of proteins as novel target for antitumor therapy: review article. AB - Imbalance in histone acetylation can lead to changes in chromatin structure and transcriptional dysregulation of genes that are involved in the control of proliferation, cell-cycle progression, differentiation and/or apoptosis. Histone acetyltransferases (HATs) and histone deacetylases (HDACs), are two classes of enzymes regulating histone acetylation and whose altered activity has been identified in several cancers. HATs and HDACs enzymes also target non histone protein substrates, including transcription factors, nuclear import factors, cytoskeleton and chaperon proteins. HDAC inhibitors are a novel class of anticancer agents which have been recently shown to induce growth arrest and apoptosis in a variety of human cancer cells by mechanism that cannot be solely attributed to the level of histone acetylation. Several clinical studies with HDAC inhibitors are ongoing, however the molecular basis for their tumour selectivity remains unknown and represent a challenge for the cancer research community. PMID- 15290352 TI - The translation elongation factor 1A in tumorigenesis, signal transduction and apoptosis: review article. AB - An increasing number of evidences suggest the involvement of the eukaryotic elongation factor 1A, a core component of the protein synthesis machinery, at the onset of cell transformation. In fact, eEF1A is shown to be up-regulated in cell death; moreover, it seems to be involved in the regulation of ubiquitin-mediated protein degradation. In addition, eEF1A undergoes several post-translational modifications, mainly phosphorylation and methylation, that generally influence the activity of the protein. This article summarizes the present knowledges on the several extra-translational roles of eEF1A also in order to understand as the protein synthesis regulatory mechanisms could offer tools for cancer intervention. PMID- 15290353 TI - Effect of human papillomavirus type 16 E6 and E7 oncogenes on the activity of the transforming growth factor-beta2 (TGF-beta2) promoter. AB - The effect of the human papillomavirus type 16 (HPV 16) E6 and E7 proteins was studied on the transcriptional activity of the human transforming growth factor beta2 (TGF-beta) promoter in different cell lines. Luciferase tests were performed after co-transfection of cells with TGF-beta2 reporter constructs and HPV 16 E6 or E7 expression vectors. HPV 16 E7, but not E6 significantly repressed TGF-beta2 promoter activity in NIH/3T3 cells, which have wild-type p53 and pRb proteins. The repressive effect of HPV 16 E7 on the transcriptional activity of the TGF-beta2 promoter could be localized to the promoter region -528 to -251 relative to the transcriptional start site. Ability of E7 to bind pRb was necessary to inhibit the TGF-beta2 promoter. Over-expression of the transcription factor E2F-1 had an effect on the TGF-beta2 promoter similar to that of E7, which may indicate that HPV 16 E7 represses the TGF-beta2 promoter by releasing E2F from pRb. PMID- 15290355 TI - RGD-mutants of B-lymphotropic polyomavirus capsids specifically bind to alpha(v)beta3 integrin. AB - Integrins are a family of cell surface proteins that function as receptors for extracellular matrix ligands and for some viruses. A subset of integrins recognises peptide sequences containing arginine-glycine-aspartic acid (RGD) motifs as ligands. The B-lymphotropic polyomavirus (LPV) has a non-enveloped capsid that recognises a sialylated cell surface receptor. To change the receptor binding specificity we have replaced sets of three amino acids in three predicted surface loops of the major capsid protein VP1 of the B-lymphotropic polyomavirus LPV by RGD. Ten mutants gave rise to the expected 40 kDa VP1 protein upon expression from a baculovirus vector in insect cells. Five of the VP1 mutants representing all three surface loops have retained the ability to spontaneously assemble to capsids in the nuclei of the insect cells. Structural changes of the mutant capsid surface were shown by differential reactivity with a set of 7 neutralising monoclonal antibodies that recognise conformational surface epitopes of wildtype LPV virions. In addition all mutant capsids had lost specific binding to the LPV receptor. Three mutant capsids of one loop (BC) showed specific binding to alpha(v)beta3 integrin but not to integrins alpha(v)beta5, alpha(v)beta6, or to alpha(IIb)beta3 known also to recognise RGD containing peptide sequences. This selective binding of the mutant capsids could be inhibited by synthetic peptides that specifically bind to alpha(v)beta3 integrin with IC50 values between 10 and 40 nM. PMID- 15290359 TI - Phylogenetic analysis of avian infectious bronchitis virus strains isolated in Japan. AB - To define the origin and evolution of recent avian infectious bronchitis virus (IBV) in Japan, a genetic analysis was performed. By phylogenetic analysis based on the S1 gene including the sequence of the hypervariable regions, IBV isolates in Japan were classified into five genetic groups, which included two already known groups (Mass and Gray). Among them, three major genetic groups were associated with the recent outbreaks of IB in Japan. One group is indigenous to Japan and could not be placed within the known existing groups in other countries. The remaining two groups, which have emerged recently, are related to isolates in China and Taiwan. PMID- 15290360 TI - Analysis of the beak and feather disease viral genome indicates the existence of several genotypes which have a complex psittacine host specificity. AB - A study was made of the phylogenetic relationships between fifteen complete nucleotide sequences as well as 43 nucleotide sequences of the putative coat protein gene of different strains belonging to the virus species Beak and feather disease virus obtained from 39 individuals of 16 psittacine species. The species included among others, cockatoos ( Cacatuini), African grey parrots ( Psittacus erithacus) and peach-faced lovebirds ( Agapornis roseicollis), which were infected at different geographical locations, within and outside Australia, the native origin of the virus. The derived amino acid sequences of the putative coat protein were highly diverse, with differences between some strains amounting to 50 of the 250 amino acids. Phylogenetic analysis demonstrated that the putative coat gene sequences form six clusters which show a varying degree of psittacine species specificity. Most, but not all strains infecting African grey parrots formed a single cluster as did the strains infecting the cockatoos. Strains infecting the lovebirds clustered with those infecting such Australasian species as Eclectus roratus, Psittacula kramerii and Psephotus haematogaster. Although individual birds included in this study were, where studied, often infected by closely related strains, infection by highly diverged trains was also detected. The possible relationship between BFD viral strains and clinical disease signs is discussed. PMID- 15290371 TI - Analysis and comparison of the 4b core protein gene of avipoxviruses from wild birds: evidence for interspecies spatial phylogenetic variation. AB - Avipoxviruses have been isolated from a wide variety of avian hosts, and yet little is known regarding the host-virus species variation of the genus Avipoxvirus. We have investigated the variations in the viral 4b core protein gene from six different avipoxviruses based on PCR, Southern blot and nucleotide sequence analysis to evaluate the suitability of this region for differentiation between avipoxvirus isolates. Southern blot and nucleotide sequence analysis revealed considerable interspecies variation between the different virus isolates. In the deduced amino acid sequences (of 142 residues) of the 4b core protein gene, fowlpox virus vaccine strain (FPV-VR250) was found to be similar to the three poxvirus isolates from great tit (GTV-A310, GTV-A311 and GTV-A256), sparrowpox virus (SPV-A468), and pigeonpox virus (PPV-B7) with similarities of 79.6%, 81%, 81%, 64.8% and 84.5%, respectively. Furthermore, comparative phylogenetic analysis of the aligned DNA sequences revealed divergence among the different viruses that can be consistently correlated to the host. PMID- 15290372 TI - Strains of Peru tomato virus infecting cocona (Solanum sessiliflorum), tomato and pepper in Peru with reference to genome evolution in genus Potyvirus. AB - Two isolates (SL1 and SL6) of Peru tomato virus (PTV, genus Potyvirus) were obtained from cocona plants (Solanum sessiliflorum) growing in Tingo Maria, the jungle of the Amazon basin in Peru. One PTV isolate (TM) was isolated from a tomato plant (Lycopersicon esculentum) growing in Huaral at the Peruvian coast. The three PTV isolates were readily transmissible by Myzus persicae. Isolate SL1, but not SL6, caused chlorotic lesions in inoculated leaves of Chenopodium amaranticolor and C. quinoa. Isolate TM differed from SL1 and SL6 in causing more severe mosaic symptoms in tomato, and vein necrosis in the leaves of cocona. Pepper cv. Avelar (Capsicum annuum) showed resistance to the PTV isolates SL1 and SL6 but not TM. The 5'- and 3'-proximal sequences of the three PTV isolates were cloned, sequenced and compared to the corresponding sequences of four PTV isolates from pepper, the only host from which PTV isolates have been previously characterised at the molecular level. Phylogenetic analyses on the P1 protein and coat protein amino acid sequences indicated, in accordance with the phenotypic data from indicator hosts, that the PTV isolates from cocona represented a distinguishable strain. In contrast, the PTV isolates from tomato and pepper were not grouped according to the host. Inclusion of the sequence data from the three PTV isolates of this study in a phylogenetic analysis with other PTV isolates and other potyviruses strengthen the membership of PTV in the so-called "PVY subgroup" of Potyvirus. This subgroup of closely related potyvirus species was also distinguishable from other potyviruses by their more uniform sizes of the protein-encoding regions within the polyprotein. PMID- 15290373 TI - Infectious bursal disease virus polyprotein expression arrests growth and mitogenic stimulation of B lymphocytes. AB - Infectious bursal disease virus (IBDV) causes lymphocytolysis and immunosuppression in infected poultry. The IBDV genome encodes a polyprotein VP243 that is post-translationally cleaved by the VP4 protease into the two structural proteins pVP2 and VP3. The objective of the present study was to determine if IBDV polyprotein induced suppression of bursal B lymphocyte growth and their capacity for proliferation. Bursal B cells were examined both for chickens infected with IBDV and for chickens orally inoculated with a DNA construct expressing IBDV VP243 polyprotein. Bursae were collected at 0, 12, 24 and 48 hours after inoculation. Proliferation of bursal B cells (purified AvBu1(+) cells) in response to concanavalin A mitogenic stimulation was significantly suppressed by infection at 1 day old with either the classical STC or variant E strains of IBDV. Oral administration of DNA constructs expressing the IBDV VP243 polyprotein from either the classical STC or variant E strains in the pCR3.1 vector resulted in persistent, moderate levels of construct in the bursa until at least 48 hours after inoculation. The VP243 DNA construct similarly induced suppression of proliferation for bursal lymphocytes independently of the virus infection. Expression of VP243 polyprotein in transiently transfected DT40 B lymphocyte culture also suppressed cell growth and proliferative responses to mitogen stimulation. Polyprotein expression did not affect cell viability and suppression of proliferation probably occurred by means of cell cycle arrest. The expression of the mature viral proteins VP2, VP4 or VP3 did not change the rate of cell proliferation or response of B cell cultures to mitogen. The results suggested that IBDV polyprotein is a mediator of immunosuppression. PMID- 15290374 TI - Biological and sequence analysis of a novel European isolate of Barley mild mosaic virus that overcomes the barley rym5 resistance gene. AB - A Barley mild mosaic virus (BaMMV) isolate from France (BaMMV-Sil) capable of overcoming rym5-controlled resistance was inoculated to barley genotypes carrying various genes for resistance to the barley mosaic viruses. BaMMV-Sil was unable to infect genotypes carrying rym1, rym4, rym8, rym9, or rym11 but genotypes carrying rym3, rym5, rym6 or no known bymovirus resistance gene were susceptible. Plants carrying rym7 or rym10 showed partial resistance with delayed virus accumulation. The two genomic RNAs of BaMMV-Sil were sequenced and compared to published sequences and those of a further common strain isolate from the UK. Four amino acid differences were observed between BaMMV-Sil and European common strain isolates in the polypeptide encoded by RNA1, the RNA species which determines pathogenicity on the rym5 genotypes. Only two of these differences are likely to be functionally important (His rather than Gln at position1217 in the VPg cistron; His rather than Asp at position 1776 in the NIb cistron). Comparisons with related viruses in the genera Bymovirus and Potyvirus suggest that the change in the VPg, which occurs within a motif conserved amongst all viruses within the family Potyviridae, is the more likely cause of rym5 resistance-breaking. PMID- 15290375 TI - Trapping of Tomato yellow leaf curl virus (TYLCV) and other plant viruses with a GroEL homologue from the whitefly Bemisia tabaci. AB - To avoid destruction in the haemolymph of their vector, many plant circulative viruses interact with GroEL homologues produced by insect endosymbiotic bacteria. We have exploited this phenomenon to devise tools allowing trapping of plant viruses by either GroEL purified from the whitefly Bemisia tabaci or by whitefly GroEL over-expressed in E. coli. PCR tubes or 96-well plates coated with a GroEL preparation were incubated with cleared sap of virus infected plant leaves or insect vectors. GroEL-bound viruses were then identified by PCR or RT-PCR using virus-specific primers or by ELISA with virus specific antibodies. In this way Tomato yellow leaf curl virus (TYLCV) - a whitefly-transmitted geminivirus - was detected in plant sap, in extracts of leaf squashes and in homogenates of individual viruliferous whiteflies. Anti-GroEL antibody prevented TYLCV binding to GroEL. GroEL-bound virus was also detected by ELISA. GroEL was much more potent in binding TYLCV than commercial anti-TYLCV antibodies. In addition to several other geminiviruses, these procedures allowed detecting a variety of RNA viruses such as Cucumber mosaic virus (CMV), Prune dwarf virus (PDV) and Tomato spotted wilt (TSWV), but not Potato virus X and Potato virus Y (PVX and PVY), Grapevine leafroll-associated viruses (GLRV) and Tobacco mosaic virus (TMV). Predictions pertaining to viruses that do, or do not bind to GroEL, and applications in plant virus diagnosis, are presented. PMID- 15290376 TI - The Tomato mosaic virus 30 kDa movement protein interacts differentially with the resistance genes Tm-2 and Tm-2(2). AB - In tomato plants ( Lycopersicon esculentum Mill.), the genes Tm-2 and Tm-2(2) confer resistance to Tomato mosaic virus (ToMV). Sequence analysis of ToMV strains able to break the Tm-2 or Tm-2(2) resistance revealed distinct amino acid exchanges in the viral 30 kDa protein, suggesting that the movement protein is recognized by both resistance genes to induce the plant defense reaction. To analyze the interactions between the ToMV movement protein and the Tm-2 and Tm 2(2) genes in detail, we generated transgenic tomato lines expressing various movement protein gene constructs. Crosses of the transgenic tomato lines with cultivars containing either the Tm-2 or the Tm-2(2) gene demonstrated that both genes are able to elicit a hypersensitive reaction in response to movement proteins from resistance inducing ToMV strains. However, the domains and the structural requirements for induction of the necrotic response by the ToMV movement protein are completely different for either resistance gene. In the context of the Tm-2 gene, the resistance determinant lies within the N-terminal 188 amino acids of the ToMV movement protein. Interaction of the 30 kDa protein with the Tm-2(2) gene requires two distinct domains localized at the C-terminus and in a different region of the protein, respectively. PMID- 15290378 TI - Pelargonium necrotic spot virus: a new member of the genus Tombusvirus. AB - A virus isolate from Pelargonium spp., provisionally designated UPEV (unknown pelargonium virus), had isometric particles 31-33 nm in diameter, with a granular surface structure similar to that of viruses in three genera of family Tombusviridae. Immunoelectron microscopy proved that UPEV was serologically distinct from all examined morphologically similar members of the family Tombusviridae. The induced cytopathology was characterized by large cytoplasmic virion aggregates and the formation of multivesicular bodies derived from mitochondria. Analysis of the complete ssRNA genome sequence revealed four open reading frames (ORFs) arranged like those of viruses in the genera Tombusvirus and Aureusvirus. Sequence comparisons indicated that three of the four ORFs had a high identity (52-97% identical amino acids) with the respective ORFs of tombusvirus species, especially with Carnation Italian ringspot virus, but not with those of viruses in other genera in Tombusviridae. On the contrary, UPEV coat protein had a low indentity (36-53% identical amino acids) with that of the aureusvirus Pothos latent virus. The data suggested that UPEV originated in a recombination event between a tombus- and an aureusvirus. According to its original host and symptom expression we proposed the new virus be named Pelargonium necrotic spot virus (PeNSV) and classified it as a distinct and new species in the genus Tombusvirus. PMID- 15290377 TI - High frequency of Epstein Barr virus latent membrane protein-1 30 bp deletion in a series of pediatric malignancies in Argentina. AB - Epstein Barr virus widely infects human populations and remains mostly asymptomatic; however, it has been associated with several malignancies. The EBV encoded latent membrane protein-1 has been involved in neoplasic transformation; a 30-bp deletion and several mutations in the COOH-terminal domain have been associated with histopathological and clinical disease features. OBJECTIVE: To analyze and correlate the presence of mutations and a 30-bp deletion with the influence of LMP-1 on tumorigenicity in a population of EBV+ pediatric malignancies. METHODS: We studied EBV presence by LMP-1 immunohistochemistry, EBERs in situ hybridization and PCR in fresh and formalin-fixed paraffin-embedded tissue samples from 10 Hodgkin's lymphomas, 6 non-Hodgkin's lymphomas, 4 undifferentiated nasopharyngeal carcinomas. Eighteen out of 20 samples were sequenced. Eight fresh normal lymphoid tissue samples and 3 peripheral blood samples were analyzed. RESULTS: All cases were EBV positive. EBV typing rendered 12 EBV-1 and 8 EBV-2. Del-LMP-1 was detected in 15/20 EBV related malignancies, as well as in 4/11 control tissues. A high percentage of patients showed point mutations previously described. The presence of del-LMP-1 and point mutations failed to correlate with clinical course. CONCLUSIONS: We found a marked incidence of del-LMP-1 (75%) in our series. However, we failed to find any correlation between histological aggressiveness of malignancies and the presence of del-LMP-1 and point mutations. PMID- 15290379 TI - Stable expression of foreign proteins in herbaceous and apple plants using Apple latent spherical virus RNA2 vectors. AB - Infectious cDNA clones of Apple latent spherical virus (ALSV)-RNA1 (pEALSR1) and RNA2 (pEALSR2) were constructed using an enhanced 35S promoter. A viral vector was constructed from pEALSR2 by creating artificial protease processing sites by duplicating the Q/G protease cleavage site between 42KP and Vp25. Eight RNA2 derived vectors expressing GFP with varied sizes of duplications around the 42KP/Vp25 junction were constructed and tested for infectivity in Chenopodium quinoa. The results indicated that greater than five aa from the C-terminus of 42KP and N-terminus of Vp25 in duplication are necessary for systemic infection. In infected C. quinoa plants, GFP fluorescence was observed in both inoculated and upper leaves. Serial passages of the viruses derived from the above vectors in C. quinoa showed that the size of duplications affected the stability of the GFP gene. The version of the RNA2-vector (pER2L5R5GFP) with the shortest duplications and its silent mutant version could stably express GFP in leaves even after at least nine serial passages. ALSV-RNA2 vector has a capacity to maintain a DNA insert as long as 1300 bp because Apple chlorotic leaf spot virus movement protein (50KP) gene could be expressed in C. quinoa. Inoculation of a virus derived from pER2L5R5GFP to apple seedlings resulted in the expression of GFP fluorescence in uninoculated upper leaves, indicating that the vector is available for the expression of foreign genes in apple trees. PMID- 15290380 TI - Molecular characterization of the nucleocapsid protein gene of Newcastle disease virus strains in Japan and development of a restriction enzyme-based rapid identifying method. AB - Nucleocapsid protein (NP) gene of Newcastle disease virus (NDV) strains mainly isolated in Japan from 1930 to 2001 was genetically characterized. By deduced amino acid sequence comparison, the N-terminal region (from 1 to 401 residues) of the NP protein was found to be highly conserved, while the C-terminal region was highly variable among the NDV isolates. A phylogenetic tree construct based on the nucleotide sequence of the complete NP gene revealed that the old (prior to 1970s) and the new (after 1980s) isolates could be classified into two major different groups, i.e., a group comprising virulent strains, and another group composed of avirulent strains. By restriction enzyme analysis using Pst I, none of the virulent strains were cleaved, while avirulent strains were cleaved. The results may be useful for simple primary screening test for differentiating NDV isolates. PMID- 15290381 TI - Prion protein allele A136 H154Q171 is associated with high susceptibility to scrapie in purebred and crossbred German Merinoland sheep. AB - Prion protein (PrP) genotypes were determined in eight sheep that have been tested positive for atypical scrapie from purebred or crossbred Merinoland sheep flocks in Germany and compared with the PrP genotypes of their flock mates. Two restriction fragment length polymorphism (RFLP) analyses were developed to determine all PRNP haplotypes occurring by variations at codons 136, 154 and 171. At least one copy of the A(136) H(154) Q(171) (AHQ) allele was found in all scrapie-positive sheep while the frequency of AHQ varied from over 23% to less than 3% in the whole flocks. There was a significant association between PrP genotype and a positive scrapie diagnosis over all flocks, suggesting a high scrapie susceptibility of PrP genotypes including the AHQ allele, at least in sheep of Merinoland type. These results argue that sheep with the AHQ allele are not generally less susceptible to scrapie and support the hypothesis that the influence of this allele on scrapie susceptibility may vary from flock to flock depending on genetic and/or epidemiological factors. This has to be considered when strategies for the eradication of scrapie in sheep are based on PrP genotypes. PMID- 15290382 TI - Bombyx mori nucleopolyhedrovirus orf8 encodes a nucleic acid binding protein that colocalizes with IE1 during infection. AB - This report describes the characterization of the Bombyx mori nucleopolyhedrovirus (BmNPV) orf8 gene. Immunoblot analyses demonstrated that orf8 was expressed as an early gene. The ORF8 protein accumulated in the nucleus, and was maintained at relatively constant levels from 4 to 24 h postinfection. Immunoblot analysis failed to detect ORF8 protein associated with budded virus and occlusion derived virus. In addition, immunohistochemical analysis by confocal microscopy showed that ORF8 protein colocalized with IE1 to specific nuclear foci throughout infection. To further examine the function of ORF8, a reporter gene was inserted into the orf8 reading frame. One orf8 disruption mutant (BmD8), which expressed the N-terminal half of ORF8, was isolated. However, it was not possible to isolate a null mutant, suggesting that orf8 may have an important role during viral infection. Single-step growth curves showed that BV production was reduced in BmD8 infected cells. Biochemical analyses indicated that ORF8 bound to nucleic acids. Together, these results suggest that BmNPV ORF8 may be involved in viral DNA replication and/or transcription. PMID- 15290383 TI - A whole genome perspective on the phylogeny of the plant virus family Tombusviridae. AB - Most current classifications of viruses are based on single gene analysis of capsid protein or polymerase. The comparison of entire genomes is a more balanced approach that should provide a more complete picture of relatedness. We have used a singular value decomposition (SVD)-based analysis to generate phylogenetic trees using whole genome protein sequences from a family of single-stranded RNA plant viruses. Our dataset includes the 26 species of the family Tombusviridae, 25 of which have complete genome sequences cataloged in GenBank. The resulting phylogenetic tree agrees well with current taxonomic classifications, but with significant exceptions. One previously unassigned virus within this family, Maize necrotic streak virus, is definitively placed within the genus Tombusvirus by this analysis. In addition, the analysis defines two distinct subsets within the genus Necrovirus. Future datasets will be expanded to include other icosahedral positive strand RNA plant viruses, and then perhaps all positive strand RNA plant viruses. PMID- 15290384 TI - Comparative analysis of adenoviral transgene delivery via tail or portal vein into rat liver. AB - The potential indications for gene therapy are expanding continuously. Currently, hepatotropic adenoviruses are useful vector systems for targeting liver in experimental animal models. Although this gene delivery technique is widely distributed, there is no common sense about how these viruses should be applied. In general, the local delivery into portal vein and the systemic application via tail vein induces above all substantial transgene expression. We here comparatively analysed both methods and found that the systemic administration of an adeno-virus expressing the green fluorescent protein resulted in a stronger infiltration, a more homogenous distribution, and a higher inter-individual reproducibility of reporter gene expression in rat liver than organ-specific administration via the portal vein. PMID- 15290385 TI - The nucleotide sequence of Watermelon mosaic virus (WMV, Potyvirus) reveals interspecific recombination between two related potyviruses in the 5' part of the genome. AB - Watermelon mosaic virus (WMV, Potyvirus) is a potyvirus with a worldwide distribution, mostly in temperate and mediterranean regions. According to the partial sequences that were available, WMV appeared to share high sequence similarity with Soybean mosaic virus (SMV), and it was almost considered as a strain of SMV in spite of its different and much broader host range. Like SMV, it was also related to legume-infecting potyviruses belonging to the " Bean common mosaic virus (BCMV) subgroup". In this paper we obtained the full-length sequence of WMV, and we confirmed that this virus is very closely related to SMV in most of its genome; however, there is evidence for an interspecific recombination in the P1 protein, as the P1 of WMV was 135 amino-acids longer than that of SMV, and the N-terminal half of the P1 showed no relation to SMV but was 85% identical to BCMV. This suggests that WMV has emerged through an ancestral recombination event, and supports the distinction of WMV and SMV as separate taxonomic units. PMID- 15290386 TI - Sequence diversity in the coat protein coding region of the genome RNA of Johnsongrass mosaic virus in Australia. AB - Sequence diversity in the coat protein coding region of Australian strains of Johnsongrass mosaic virus (JGMV) was investigated. Field isolates were sampled during a seven year period from Johnsongrass, sorghum and corn across the northern grain growing region. The 23 isolates were found to have greater than 94% nucleotide and amino acid sequence identity. The Australian isolates and two strains from the U.S.A. had about 90% nucleotide sequence identity and were between 19 and 30% different in the N-terminus of the coat protein. Two amino acid residues were found in the core region of the coat protein in isolates obtained from sorghum having the Krish gene for JGMV resistance that differed from those found in isolates from other hosts which did not have this single dominant resistance gene. These amino acid changes may have been responsible for overcoming the resistance conferred by the Krish gene for JGMV resistance in sorghum. The identification of these variable regions was essential for the development of durable pathogen-derived resistance to JGMV in sorghum. PMID- 15290387 TI - Analysis of an isolate of Mungbean yellow mosaic virus (MYMV) with a highly variable DNA B component. AB - One DNA A (KA30) and five different DNA B components (KA21, KA22, KA27, KA28 and KA34) of a geminivirus, Mungbean yellow mosaic virus-Vigna (MYMV-Vig) were cloned from a pooled sample of field-infected Vigna mungo plants from Vamban, South India. MYMV-Vig DNA A (KA30) and one of the DNA B components (KA27) exhibited 97% and 95% sequence identities, respectively, to those of MYMV reported from Thailand. However, the DNA B components KA21, KA22, KA28 and KA34 exhibited only 71 to 72% sequence identity to MYMV DNA B. Co-existence of multiple DNA B components in field-infected V. mungo was proved by Southern and PCR analyses. Each of the five DNA B components was infective together with the DNA A upon agroinoculation. Agroinoculation with mixed cultures of Agrobacterium with partial dimers of DNA A and all five DNA Bs proved that all five DNA B components can co-infect a single V. mungo plant. PMID- 15290388 TI - Selection of genetic inhibitors of rabies virus. AB - A cDNA library of short random fragments derived from four of the five genes of the rabies virus genome has been used to isolate genetic suppressor elements (GSEs) expressed intracellularly that inhibit rabies virus replication. Two nucleotide fragments, one from the rabies virus nucleocapsid protein (N) gene and the other from the phosphoprotein (P) gene, have been identified as inhibitors of rabies virus replication in cell culture. The N cDNA fragment is expressed in sense-orientation and could produce a dominant negative protein affecting virus replication. The P cDNA fragment is expressed in the inhibitory antisense direction. Inhibition of rabies virus replication was detected in cell culture using an ELISA for detection of rabies virus glycoprotein expression on the cell surface and immunofluorescence for detection of intracellular rabies virus N expression. Both the sense and antisense GSEs, because of their targeted inhibition of rabies virus replication, have possible uses in rational design of antiviral compounds for treatment of rabies. This approach could be applied to any virus, particularly to those that lyse their target host cell. PMID- 15290391 TI - Air-mattress motion therapy for respiratory failure after cardiothoracic surgery. AB - PURPOSE: The standard management of respiratory failure after cardiothoracic surgery involves prolonged mechanical ventilation, drug support, and chest physiotherapy. We report the effectiveness of a novel type of respiratory physiotherapy, the air mattress V-CUE unit, which utilizes a computerized automatic combination of three movements: rotation, percussion, and vibration. METHODS: Among 189 patients who underwent various cardiothoracic operations, 6 were eligible for V-CUE application, after the development of respiratory failure caused by early adult respiratory distress syndrome, massive atelectasis, or pulmonary infection. RESULTS: The V-CUE unit was used to treat six postsurgical patients, aged 58-73 years (mean, 65.8 years); four who had undergone coronary artery bypass grafting, one who had undergone mitral valve replacement, and one who had undergone a bilobectomy. The mean duration of supportive V-CUE was 3.9 days (range 2-6 days), and the mean duration of mechanical ventilation was 2.9 days. There was no need for reintubation or tracheotomy, and all six patients recovered uneventfully. CONCLUSION: Our preliminary results indicate that V-CUE dynamic air therapy is effective for managing postoperative respiratory failure, adjuvant to or complementary to standard therapy. PMID- 15290389 TI - Binding of the hemagglutinin from human or equine influenza H3 viruses to the receptor is altered by substitutions at residue 193. AB - Interactions of the hemagglutinin (HA) of influenza viruses with sialic acids (SA) are important for host range restriction. Most human H3s have a Ser193, while avian and equine H3s usually have an Asn or a Lys, respectively. To investigate the role of residue 193 in the recognition of SA, substitutions were introduced by mutagenesis within a human H3 and an equine H3. Hemadsorption assays performed on COS-1 cells expressing wt or mutated HAs, showed that a K193S substitution in the context of an equine H3 decreased its ability to bind several animal erythrocytes. Using de- and then alpha2,3 or alpha2,6 re-sialylated chicken erythrocytes we showed that for both human and equine H3s, substitution of a Serine by positively-charged Arginine or Lysine at position 193 increased binding to its preferred receptor, SAalpha2,6Gal and SAalpha2,3Gal, respectively. Moreover, when combined with the L194I substitution, the S193R substitution induced binding of the human H3 to NeuAcalpha2,3Gal. PMID- 15290392 TI - Use of bioresorbable membrane to prevent postoperative small bowel obstruction in transabdominal aortic aneurysm surgery. AB - PURPOSE: To evaluate the efficacy of Seprafilm (Genzyme, Cambridge, MA, USA), a bioresorbable membrane, in preventing or reducing early postoperative small bowel obstructions after transabdominal abdominal aortic aneurysm (AAA) surgery. METHODS: Fifty-one patients underwent aortic reconstruction via a midline transperitoneal approach for infrarenal AAAs. Twenty-one patients underwent surgery with Seprafilm (Seprafilm group) and the remaining 30 patients did not (control group). The incidence of early small bowel obstruction was examined, and the time before liquid and solid diet were resumed was also compared to assess postoperative paralytic ileus. RESULTS: Patients in the Seprafilm group resumed a liquid diet on postoperative day (POD) 2.4 +/- 1.1 and a solid diet on POD 4.0 +/ 1.3, whereas the patients in the control group resumed a liquid diet on POD 3.3 +/- 1.9 and a solid diet on POD 5.4 +/- 3.4. These values were not significantly different between the two groups; however, the incidence of early postoperative small bowel obstruction was significantly lower ( P < 0.05) in the Seprafilm group (0/21) than in the control group (6/30). CONCLUSION: These findings suggest that Seprafilm may help to prevent early postoperative small bowel obstructions after transabdominal AAA surgery. PMID- 15290393 TI - Initial management of acute type-A aortic dissection with a thrombosed false lumen: a retrospective cohort study. AB - PURPOSE: Acute type-A aortic dissection with a clotted false lumen is often managed conservatively; however, we found that surgery has a better outcome. METHODS: Enhanced computed tomography (CT) showed a clotted false lumen in the ascending aorta in 38 (33.3%) of 114 patients with acute type-A aortic dissection. After the exclusion of 8 patients whose condition was too critical for comparison, 13 patients who were hemodynamically stable and did not have pericardial effusion, organ ischemia, or a dilated ascending aorta greater than 50 mm in diameter, were managed conservatively (group C) and 17 were managed surgically (group S). We compared the early and late results of both groups. RESULTS: The early mortality rates were 23.1% in group C and 0% in group S ( P = 0.037). The early deaths in group C were caused by redissection in the acute phase. The actuarial survival rates and dissection-related event-free rates 5 years after onset in groups C and S were 64.1% and 80.8% ( P = 0.131) and 46.2% and 92.9% ( P = 0.002), respectively. CONCLUSION: The early mortality rate and dissection-related event-free rate were better after surgery than after conservative treatment. The indications for conservative management should be limited because redissection is usually fatal. PMID- 15290394 TI - Long-term results of aorta-superior mesenteric artery bypass using a new route. AB - PURPOSE: Several methods of revascularization after mesenteric ischemia have been proposed. Using a new route, we performed retrograde loop bypass grafting to the superior mesenteric artery (SMA) with a ringed expanded polytetrafluoroethylene (ePTFE) graft. METHODS: We anastomosed the graft to the infrarenal aorta, which ran behind the left renal hilum, turned ventral, and was anastomosed to the SMA in an antegrade fashion hemodynamically. Five patients underwent this procedure, which resulted in remarkable symptomatic relief. RESULTS: There were no postoperative deaths or serious complications, although some patients suffered paralytic ileus. All of the grafts remained patent during long-term follow-up, ranging from 17 to 72 months (mean: 37.8 months). Postoperative angiograms showed good configuration of the graft, which did not compress the renal vessels. CONCLUSION: Infrarenal aorta-SMA bypass relieved mesenteric ischemia and achieved good long-term graft patency. Thus, we consider it to be an effective and durable vascular procedure to reduce postoperative mortality and morbidity. PMID- 15290395 TI - Perioperative prostaglandin E1 treatment for the prevention of postoperative complications after esophagectomy: a randomized clinical trial. AB - PURPOSE: We conducted a prospective randomized clinical study to examine whether perioperative prostaglandin E1 (PGE1) could help in the prevention of postoperative complications after esophagectomy for esophageal cancer. METHODS: Forty patients with esophageal cancer eligible for radical esophagectomy were randomly assigned to an experimental group ( n = 20), given perioperative PGE1, or to a control group ( n = 20), given standard postoperative treatment. The main clinical endpoints examined were the incidence of postoperative complications, hospitalization, duration of systemic inflammatory response syndrome (SIRS), portal vein blood flow, and serum bilirubin levels. RESULTS: Severe postoperative complications developed in five patients in the control group and two in the PGE1 group. There was one surgery-related death in the control group. The duration of SIRS was significantly shorter in the PGE1 group than in the control group (5.74 days vs 7.50 days; P = 0.047). Portal vein flow was also significantly lower on postoperative day (POD) 1 in the control group than in the PGE1 group ( P = 0.042). Maximum postoperative serum bilirubin levels were significantly lower in patients treated with PGE1, at 2.91 vs 4.38 mg/dl in the control group ( P = 0.040). CONCLUSIONS: The perioperative administration of PGE1 helps maintain adequate portal blood flow, improves hyperbilirubinemia, and attenuates the duration of SIRS, thereby reducing the risk of postoperative complications after esophagectomy and lymphadenectomy for esophageal cancer. PMID- 15290396 TI - Changes in total serum glycosaminoglycan levels in patients undergoing renal transplantation: preliminary data. AB - PURPOSE: To study the involvement of glycosaminoglycans (GAGs) in the immunological process of renal disease. They are related to cytokines, which are known to play an important role in acute graft rejection after renal transplantation. METHODS: We studied 40 patients on maintenance hemodialysis for chronic renal failure, who underwent renal transplantation from a live donor. We measured serum GAGs preoperatively, intraoperatively, and on postoperative days (PODs) 2 and 10. The clinical outcome and other biochemical markers, such as blood urea nitrogen and serum creatinine, were also recorded. RESULTS: The total serum GAG levels decreased after renal transplantation in patients with normal graft function (group N). However, in patients with acute graft rejection post transplantation (group R), the GAG levels remained significantly elevated. CONCLUSIONS: Total serum GAGs fluctuate in renal transplantation and their association with graft rejection should be investigated further. PMID- 15290397 TI - Soluble and cell-associated forms of some yet to be identified factor in transfused blood which promotes solid tumor growth in mice. AB - PURPOSE: The mechanism underlying the immunomodulation caused by blood transfusion has yet to be elucidated. The aim of the present study was to determine whether the transfusion of a soluble or insoluble factor present in stored blood can induce immunomodulation, which would thereby promote solid tumor growth. METHODS: C57Bl/6J mice were subcutaneously inoculated with B16-CG melanoma cells, which secrete beta-human chorionic gonadotropin (beta-hCG). Following inoculation, each of three different products of allogeneic and syngeneic blood were transfused on days 0 and 1: fresh whole blood, stored whole blood, and supernatants from the stored blood. Tumor growth was then monitored by measuring urinary beta-hCG. All mice were killed on day 15, and the tumor weight and volume were measured. RESULTS: Transfusion of all allogeneic blood products enhanced tumor growth, as did the stored syngeneic whole blood. Neither fresh syngeneic blood nor the supernatant from stored syngeneic blood promoted tumor growth. Although the tumors were not visually detectable until day 10 after inoculation, by day 7 the levels of urinary beta-hCG were significantly higher in the mice that received allogeneic blood supernatant than in the mice that received saline. CONCLUSIONS: A soluble alloantigen enhances solid tumor growth, as does an insoluble factor present in stored syngeneic whole blood. The immunomodulation associated with this factor begins to enhance tumor growth within 7 days after transfusion. PMID- 15290398 TI - FK506 to prevent lung injury after hindlimb ischemia and reperfusion in a rat model: an electron microscopic study. AB - PURPOSE: Hindlimb ischemia and reperfusion leads to lung injury in various animal models. We investigated the effectiveness of FK506, an immunosuppressive agent, which also modulates neutrophilic infiltration, in preventing lung injury after hindlimb ischemia and reperfusion in a rat model. METHODS: Twenty-seven male Sprague-Dawley rats were randomized to received FK506 at doses of 0.3 mg/kg, 0.5 mg/kg, or 1 mg/kg body weight per day, or normal saline injections, as pretreatment, and there was also a sham group. On the 4th day, the animals were subjected to 2 h of ischemia induced by a tourniquet, followed by reperfusion of the extremities for 2 h. Lung tissue assays were performed for the lipid peroxidation product malondialdehyde (MDA) and total glutathione (GSH). Lung tissues were also examined histopathologically under light and electron microscopy. RESULTS: The MDA levels in the study groups were significantly lower than those in the control group ( P < 0.05), but the total GSH levels did not differ significantly among the groups. Histopathologically, there were no significant differences among the groups given different doses of FK506, but there was a significant difference between the control group and all the treatment groups. CONCLUSION: FK506 ameliorates the lung injury associated with ischemia and reperfusion of the lower limbs, and might have an inhibitory effect on the neutrophils that cause remote organ damage. PMID- 15290399 TI - Relationship between fibril length and tissue ingrowth in the healing of expanded polytetrafluoroethylene grafts. AB - PURPOSE: To determine whether fibril length is correlated with graft healing as well as cellular and capillary ingrowth in a canine carotid implantation model. METHODS: Expanded polytetrafluoroethylene (ePTFE) vascular grafts with three different fibril lengths (30, 60, and 90 microm) were implanted into the carotid artery in dogs. They were retrieved 4 weeks later, and subjected to histomorphometric analysis. RESULTS: Endothelial healing was best in the 60 microm grafts. Not only cellular ingrowth but also capillary ingrowth was most evident in the 60-microm grafts, followed by the 90-microm grafts and then the 30 microm grafts. CONCLUSION: Better endothelial healing of ePTFE vascular grafts is correlated with more cellular and capillary ingrowth, but more cellular and capillary ingrowth is not correlated with longer fibril length or higher air porosity. PMID- 15290400 TI - Omeprazole is more effective than famotidine for preventing acute gastritis in rats. AB - PURPOSE: Acute gastric mucosal lesions, which can develop within a few hours after polytrauma, shock, major operations, central nervous system lesions, or severe infection, cause about 33% of cases of gastrointestinal bleeding. We analyzed and compared the effectiveness of famotidine and omeprazole on acute gastric mucosal lesions. METHODS: Thirty male albino Wistar rats were given ketalar anesthesia after 12 h fasting, then immobilized and exposed to stress according to Brodie's protocol, without restricting their respiration. We divided the rats into three groups of ten according to whether they were given famotidine, omeprazole, or normal saline (control group). All rats were ulcer indexed according to the diameter of their ulcers. The stomach contents were aspirated for acid output and pH analysis, and sent to the laboratory. The total number of mast cells was also counted. RESULTS: Omeprazole was more effective than famotidine in keeping gastric pH high and lowering the total gastric acid output. Lower ulcer indexes in acute gastric mucosal erosions and better protected mucosal integrity were found in the omeprazole-treated rats. CONCLUSION: Omeprazole prevents acute gastric mucosal erosions in rats more effectively than famotidine. PMID- 15290401 TI - Concomitant mitral and tricuspid valve infective endocarditis: report of a case. AB - A rare case of native valve endocarditis affecting both the normal mitral and tricuspid valves is presented. A 25-year-old woman with an acute ischemic stroke was found to have vegetation secondary to infective endocarditis as the embolic source. One month after the onset of embolic cerebrovascular intervention, a valve repair with the implantation of artificial chordae, sliding commissuroplasty, and ring annuloplasty resulted in a complete recovery. PMID- 15290402 TI - Large perigraft seroma after aortoiliac bypass with an expanded polytetrafluoroethylene graft: report of a case. AB - A 63-year-old man who had undergone aortoiliac bypass with an expanded polytetrafluoroethylene (PTFE) graft was referred to our hospital for investigation and treatment of a possible pseudoaneurysm of the abdominal aorta. A tender, pulsatile, and bulging mass, about the size of an adult fist, was palpated around the navel. Enhanced computed tomography (CT) showed a large low density area around the abdominal aorta and PTFE graft, and aortography showed a patent graft with no anastomotic leakage. Operative inspection revealed that the pulsatile mass was a large perigraft seroma, and we replaced the PTFE graft with a new woven Dacron graft. The patient has been well with no sign of recurrence for 1 year, although close long-term follow-up is mandatory. PMID- 15290403 TI - Emergency pylorus-preserving pancreatoduodenectomy followed by second-stage pancreatojejunostomy for a gastrointestinal stromal tumor of the duodenum with an intratumoral gas figure: report of a case. AB - We report a case of a duodenal gastrointestinal stromal tumor (GIST) necessitating urgent surgery because of a gas figure on computed tomography (CT). A 46-year-old woman, complaining chiefly of upper abdominal pain and tarry stools, consulted a local doctor. A gastrointestinal fiberscopy revealed an ulcer in the second part of the duodenum, and the patient was admitted to our hospital where a dynamic CT scan showed a hypervascular solid tumor in the pancreatic head. A repeat CT scan done 4 days later showed a gas figure in the tumor, necessitating an emergency pylorus-preserving pancreatoduodenectomy (PpPD). First, we performed a tube pancreatostomy for complete external drainage of the pancreatic juice, and planned a second-stage pancreatojejunostomy for the near future. Histopathologically, the tumor was diagnosed as a GIST originating in the duodenum. The patient was discharged on postoperative day 23 after an uneventful postoperative recovery. Her local doctor completed the second-stage pancreatojejunostomy. PMID- 15290404 TI - Radiation-induced rectal cancer originating from a rectocutaneous fistula: report of a case. AB - This report describes a patient with radiation-induced rectal cancer with an unusual history. A 51-year-old man was admitted in 2000 because of ichorrhea of the skin on the left loin. The patient had received irradiation for a suspicious diagnosis of a malignant tumor in the pelvic cavity in 1975. A subcutaneous abscess in the right loin appeared in 1989, and rectocutaneous fistula was noted in 1992. Moreover, radiation-induced rectal cancer developed in 2000. Plain computed tomography and magnetic resonance imaging of the pelvis demonstrated a presacral mass and tumor in the rectum. Finally, we diagnosed the presacral mass to be an abscess attached to the center of the rectal cancer. The rectum was resected by Miles' operation and a colostomy of the sigmoid colon was also performed. Many cases of radiation-induced rectal cancer have been reported. However, this is a rare case of radiation-induced rectal cancer originating from a presacral abscess and rectocutaneous fistula. PMID- 15290405 TI - Successful resection of a ruptured hepatoblastoma prior to chemotherapy: report of a case. AB - A 12-year-old boy was referred to our hospital suffering from severe anemia and liver dysfunction. The laboratory data on admission confirmed severe anemia and an elevated alpha-fetoprotein level. Abdominal ultrasonography revealed a mass measuring 51 x 49 mm in size, and abdominal computed tomography showed a low density mass in S8 of Couinaud's segment and a low-density area in S7, thus suggesting bleeding in the tumor. Right subphrenic fluid collection and perirectal fluid collection were also observed. Celiac arteriography showed a faint tumor stain fed by A5-8 but no evidence of any extravasation. A diagnosis of pediatric liver carcinoma was made, and the case was classified as T2 C3 V0 N0 M0 Stage IIIA. Although there was no evidence of bleeding during angiography, because of the high risk of rebleeding, a laparotomy was performed before chemotherapy. At operation, the tumor rupture site and hematoma appeared to be in S7, and a right lobectomy was thus performed. Ascitic fluid cytology was class V. The cut surface of the resected specimen showed a tumor measuring 51 x 49 mm located in S8 and a hematoma located in S7. Histologically, the tumor was a well differentiated hepatoblastoma. The patient was transferred to the pediatric department and treated with six courses of intravenous chemotherapy followed by peripheral blood stem cell transplantation. The outcome has been favorable, with no recurrence as of 25 months after the operation. PMID- 15290406 TI - A functioning-desensitization paraganglioma which caused hemodynamic instability during tumoral resection: report of a case. AB - Paragangliomas are uncommon tumors arising from the neuroendocrine elements of the paraganglia. Their successful management is associated with many problems. We herein present the findings of a 22-year-old man in whom a paraganglioma was incidentally found and in which the clinical and previous operative behavior was functioning desensitization. As a result, preoperative medication was not performed; however, during the tumor resection the patient demonstrated hemodynamic instability. PMID- 15290407 TI - Treatment strategies for perianal rhabdomyosarcoma: report of two cases. AB - Intensive chemotherapy and high-dose radiation with complete excision of the tumor are the treatment of choice for rhabdomyosarcoma (RMS). However, because there are so few reports of perianal RMS, a mainstream treatment has not been established. We report two cases of perianal RMS and review 13 cases documented in Japan since 1990. Case 1 was a 7-year-old boy first treated by tumor excision followed by chemotherapy and external irradiation. He has since been tumor-free but suffered severe perianal erosion and ulceration. Case 2 was a 17-year-old girl first treated by preoperative chemotherapy to reduce the tumor size, after which the tumor was completely excised with anatomical guidance using three dimensional computed tomography via the posterior sagittal approach, followed by open intraoperative irradiation. She has since been tumor-free with preserved anal function. These case reports show the importance of cogitated treatments for preservation of anal function and optimal therapeutic outcome in patients with perianal RMS. PMID- 15290408 TI - The effects of epidural application of allografted nucleus pulposus in rats on cytokine expression, limb withdrawal and nerve root discharge. AB - This study investigated cytokine expression, behavioral and neurophysiologic changes in Lewis rats whose lumbar nerve roots were exposed to nucleus pulposus (NP). Allografted NP or fat was implanted over the left L5 nerve root. Sham rats had no NP or fat implantation. Control rats had no surgery. Rats were allowed to survive for 7 days and were tested daily for hind-paw mechanical and thermal withdrawal response (TWR). Granulation tissue was processed by immunohistochemistry for cytokines--interleukin 1 beta (IL-1beta), interleukin 6 (IL-6) and tumor necrosis factor (TNF). Neurophysiological response from the L5 nerve roots was also characterized after 7 days. Significant staining density for IL-1beta, IL-6 and TNF was observed in NP granulation tissue compared with fat and sham (p<0.05). However, there were no significant thermal and mechanical behavioral changes. TWR data computed as percentage-difference scores indicated no significant changes in withdrawal response between the four groups, although NP-treated group showed a trend of decreasing withdrawal latency. Comparison of combined percentage-difference scores revealed increased sensitivity in the NP group on days 4, 5 and 6, 7 when compared with control rats only, with no significant changes in the percentage-difference scores of fat and sham rats when compared to control. Neurophysiologically, the percentage increase in discharge rate in NP-treated rats was higher than control (p<0.05) but not higher than fat and sham rats. These results support the inflammatory nature of NP but offer limited support to NP-mediated thermal behavioral changes. PMID- 15290409 TI - Arum- and Paris-type arbuscular mycorrhizas in a mixed pine forest on sand dune soil in Niigata Prefecture, central Honshu, Japan. AB - Arbuscular mycorrhizas (AM) are the most widespread mycorrhiza in nature and form two morphologies, Arum- and Paris-type. The determining factors defining the two different morphologies are not well understood. In this study, the distribution of Arum- and Paris-type AM was determined in a mixed pine forest. A total of 35 plant species belonging to 20 families and 32 genera were identified and examined for AM colonization and morphological types. AM morphological types in 14 families were confirmed as follows: Arum-type in Rosaceae, Oleaceae, Lauraceae, Vitaceae and Compositae, Paris-type in Aquifoliaceae, Ulmaceae, Araliaceae, Theaceae, Magnoliaceae, Rubiaceae and Dioscoraceae, and both and/or intermediate types in Caprifoliaceae and Gramineae. Plant families whose AM morphological status was previously unknown were clarified as follows: Polygonaceae and Commelinaceae showed Arum-type morphology; Celastraceae, Menispermaceae and Elaeagnaceae had typical Paris-type morphology. The proportion of Arum-type to Paris-type species decreased in the following order: annuals > perennials > deciduous species > evergreen species, and pioneer group > early successional group > late successional group. Evergreen plants had a higher tendency to form Paris-type AM than annuals, perennials and deciduous plants. The results indicate that environmental changes, such as shade during plant succession, control the distribution of plant growth forms in mixed pine forest and may also play a part in the distribution of Arum- and Paris-type morphology. The identity of the plant seems to strongly influence AM morphology, though control by the fungal genome cannot be ruled out. PMID- 15290410 TI - Addition of epinephrine to intrathecal tetracaine augments depression of the bispectral index during intraoperative propofol sedation. AB - PURPOSE: Epinephrine added to local anesthetic agents for spinal anesthesia is frequently used to prolong the duration of anesthesia. Epinephrine stimulates the alpha-adrenoceptor, and it is known that the alpha2-adrenoceptor agonists have a central inhibitory effect. We investigated the effect of intrathecal epinephrine during propofol sedation with spinal anesthesia, using a bispectral index (BIS) monitor. METHODS: Twenty adult patients, scheduled for spinal anesthesia, were allocated to the control group ( n = 10) or epinephrine group ( n = 10). Patients in the control group received 14 mg of tetracaine, whereas the epinephrine group received 14 mg of tetracaine and 0.2 mg of epinephrine. Immediately after the pinprick test, propofol was administered at 0.5 mg x kg(-1) by infusion for the initial dose, then continuously at 2 mg x kg(-1) x h(-1) in both groups. BIS scores were recorded before subarachnoid block, and then every 5 min for 90 min after subarachnoid block. RESULTS: There were significant differences in the BIS score between the two groups at 45-55 min and at 60-70 min after subarachnoid block. CONCLUSION: Intrathecal epinephrine augments the sedative effect of propofol during spinal anesthesia. PMID- 15290411 TI - A comparison of postoperative sore throat after use of laryngeal mask airway and tracheal tube. AB - PURPOSE: We compared the degree of postoperative sore throat (PST) after use of a laryngeal mask airway (LMA; by two insertion techniques) and a tracheal tube (TT) in adult patients. METHODS: Eighty-six adult patients undergoing surgery of an extremity were randomized into three groups. The LMAs (size 4 for men, 3 for women) and TTs were lubricated with 2% lidocaine gel. After the induction of anesthesia, an LMA with the cuff deflated was inserted and then the cuff was inflated in group A, an LMA with the cuff inflated was inserted in group B, and the trachea was intubated using vecuronium in group C; staff anesthesiologists performed all these methods. LMA cuffs were inflated with the maximum recommended volume of air. TT cuffs were inflated with the minimum volume of air without gas leakage at 20 cm H(2)O pressure. The mode of ventilation depended on the individual anesthesiologists. Blood traces on the devices were examined after their removal. PST was rated immediately after anesthesia and on the first postoperative day, using a three-point score and a 100-mm visual analog scale, respectively. RESULTS: Most of the patients receiving an LMA breathed spontaneously and those receiving a TT underwent controlled ventilation. The ratio of positive blood traces on devices, as well as the degree of PST immediately after anesthesia, was similar in the three groups; however, on the first postoperative day, the severity of PST was greater in the LMA groups than in the TT group ( P = 0.016). The severity of PST was similar with the two LMA insertion techniques. CONCLUSION: In the conditions of our study, LMAs inserted with the cuff either fully inflated or deflated worsened PST compared with TTs. PMID- 15290412 TI - Efficacy of prophylactic intravenous granisetron in postoperative emesis in adults. AB - PURPOSE: This randomized, double-blind, placebo-controlled trial evaluated the efficacy, safety, and optimal dose of granisetron in the prophylactic control of postoperative nausea and vomiting in patients undergoing gynecologic surgery or cholecystectomy. METHODS: Three-hundred and fifteen patients (age, 20-65 years) received intravenous granisetron (1 mg or 3 mg) or placebo immediately before the end of anesthesia. After treatment, patients were observed for 24 h, and the occurrence of nausea and vomiting was recorded and safety was assessed. The no vomiting rate, time-to-first vomiting episode, and severity of nausea were recorded. RESULTS: The no-vomiting rates in patients receiving granisetron 1 mg and 3 mg were significantly higher than that in the placebo group (83.7%, 78.8%, and 57.9%, respectively; P = 0.0004 for 1 mg vs placebo, P = 0.001 for 3 mg). Time-to-first vomiting episode was longer in the granisetron 1-mg and 3-mg groups than in the placebo group (time-to-event analysis, Kaplan-Meier, log-rank test; 83.2%, 80.1%, and 59.1%, respectively; P = 0.0002 and P = 0.0010). The severity of nausea was also less in granisetron-treated patients (25.2%, 11.5%, and 15.4% severe nausea incidence for placebo, granisetron 1 mg, and granisetron 3 mg, respectively; P = 0.00003 and P = 0.002). Fewer rescue medications were required in the two granisetron-treated groups compared with those receiving placebo. Adverse events were similar in all groups. No differences in efficacy or safety were observed between granisetron doses. CONCLUSION: Granisetron is well tolerated and more effective than placebo in the prophylactic control of nausea and vomiting after surgery. This study suggests that the optimum dose of granisetron is 1 mg. PMID- 15290413 TI - Safety and beneficial effect on body core temperature of a prewarmed plasma substitute--hydroxyethyl starch--during anesthesia. AB - PURPOSE: We investigated, first, the safety of use and stability of a plasma substitute-hydroxyethyl starch (HES)-kept in a warming cabinet for a long period, and then the effect on body core temperature of the prewarmed HES in patients during urological surgery. METHODS: In the first part of the study, HES colloid solutions (500 ml per pack; Hespander) were kept in a warming cabinet (40 degrees C) for 3 months and were tested for biological and chemical safety and stability. In the second part of the study, 1000 ml of HES at room temperature (control group; n = 10) or kept in a warming cabinet for a few days (warmed group; n = 10) was infused via a central venous catheter for 30 min in patients undergoing urological surgery under general anesthesia with lumbar epidural anesthesia. Esophageal temperature was monitored as the core temperature. HES fluid temperatures in the pack and at the end of a 1-m intravenous tube connected to the central venous catheter were also measured. RESULTS: The test of HES products warmed for 3 months passed all inspections performed during the study period. In the warmed group, the pack and intravenous tube temperatures of HES were still high at 15 min after infusion (37.1 degrees +/- 1.5 degrees C [mean +/- SD] and 34.8 degrees +/- 2.2 degrees C, respectively). Core temperature in the warmed group decreased significantly, by 0.34 degrees +/- 0.06 degrees C, but was significantly higher than that in the control group (by 0.84 degrees +/- 0.13 degrees C) after 30 min of the infusion. CONCLUSIONS: The use of HES products kept in a warming cabinet prior to surgery can maintain warm body temperature, easily, safely, and effectively. PMID- 15290414 TI - Duration of vecuronium-induced neuromuscular block can be predicted by change of skin temperature over the thenar muscles. AB - PURPOSE: The purpose of this study was to clarify the relationship between skin temperature over the thenar muscles and the duration of action of vecuronium measured acceleromyographically at the thumb in anesthetized patients. METHODS: In 15 patients undergoing elective open abdominal surgery under propofol, fentanyl, and nitrous oxide anesthesia, train-of-four (TOF) stimuli were delivered over the ulnar nerve at 2 Hz every 15 s, and the degree of neuromuscular block was measured acceleromyographically at the thumb. Each patient received an intubating dose of vecuronium 0.1 mg x kg(-1), followed by maintenance doses of 0.02 mg x kg(-1) administered repeatedly whenever the first twitch of TOF responses had recovered to 25% of control. The interval between maintenance doses was defined as the clinical duration (DUR25). The median values of skin temperature (ST) over the ipsilateral thenar muscles and esophageal temperature (ET) were recorded during the action of the first and all subsequent maintenance doses. The relationships between change in temperature and change in DUR25 were analyzed. RESULTS: Whereas ET showed only minor changes (median, -0.3 degrees C), ST fluctuated markedly between +0.9 degrees and -6.3 degrees C (median, -1.4 degrees C). Increase and decrease were also seen in a series of DUR25s, as expected from the changes in ST. The median values of DUR25 produced by the first and last maintenance vecuronium doses were 21.5 and 32.3 min, respectively. A negative linear correlation was found between the change in DUR25 and that in ST, demonstrating that DUR25 increased by 20% of baseline with each 1 degrees C decrease in ST. CONCLUSION: Our results show that peripheral ST decreases considerably during open abdominal surgery without reduction in core temperature, and the decrease contributes to the potentiation of neuromuscular block in the periphery during propofol, fentanyl, and nitrous oxide anesthesia. PMID- 15290415 TI - Systemic ATP infusion improves spontaneous pain and tactile allodynia, but not tactile hypesthesia, in patients with postherpetic neuralgia. AB - PURPOSE: Activation of purinoceptors may improve neuropathic pain. Accordingly, the effects of systemic ATP infusion were assessed in patients with postherpetic neuralgia (PHN). METHODS: Eight patients with PHN lasting over 3 months were enrolled. Initially, patients received the vehicle (20% dextrose) or ATP (at a dose of 1 mg x kg(-1) in 20% dextrose) infused intravenously for 60 min on two separate occasions in a single-blinded manner. The levels of spontaneous continuous pain, paroxysmal pain, and tactile allodynia were assessed by a visual analogue scale (VAS), and tactile hypesthesia was assessed by Semmes-Weinstein monofilament before and after infusion. Subsequently, the eight patients received an ATP infusion (1 mg.kg(-1) in 20% dextrose) once a week for 5-12 weeks in an open-label manner, and changes in the above parameters were assessed. RESULTS: In the initial study, VAS for spontaneous continuous pain and tactile allodynia decreased significantly with ATP infusion but not with placebo infusion. After repeated ATP infusions for 5-12 weeks, the median VAS for spontaneous continuous pain, paroxysmal pain, and tactile allodynia decreased significantly from 32.1 to 13.0, from 46.9 to 17.5, and from 49.5 to 15.6 respectively. However tactile hypesthesia did not improve significantly. CONCLUSION: This study demonstrated that repetitive intravenous ATP infusion could improve spontaneous continuous pain and paroxysmal pain, as well as improving tactile allodynia, but did not influence tactile hypesthesia. PMID- 15290416 TI - A comparison of the absolute amplitude of motor evoked potentials among groups of patients with various concentrations of nitrous oxide. AB - PURPOSE: It has been shown in previous studies that nitrous oxide (N(2)O) suppresses the amplitude of motor evoked potentials (MEPs) in individual subjects. In the present study, we compared the absolute amplitude and latency of MEPs among groups of patients with various concentrations of N(2)O. METHODS: The subjects were 60 patients who were scheduled to undergo craniotomy with MEP monitoring. Anesthesia was induced and maintained with propofol and fentanyl. The patients were randomly assigned to one of three groups based on the concentration of N(2)O: 0% N(2)O (N0 group), 50% N(2)O (N50 group), and 66% N(2)O (N66 group). MEPs were elicited by transcranial electrical stimulation. The effect-site concentrations (ESCs) of anesthetics were calculated retrospectively. The effects of anesthetics on MEP were analyzed by analysis of covariance (ANCOVA) followed by Tukey's method. RESULTS: MEPs were elicited in all cases. The absolute amplitude of the MEP was significantly higher in the N0 group than in the N50 and N66 groups [4.16 +/- 0.42 vs 1.00 +/- 0.26 mV and 1.00 +/- 0.27 mV, respectively (mean +/- SD); P < 0.05]. In contrast, there was no significant difference in the latency of the MEP among the three groups of subjects (N0: 16.64 +/- 0.72, N50: 16.78 +/- 0.66, and N66: 16.82 +/- 0.63 ms). CONCLUSION: The results suggest that N(2)O can suppress the absolute amplitude of the MEP in patients under propofol and fentanyl anesthesia. Although monitoring of MEP as a trend is feasible even if N(2)O is used, the use of N(2)O may be better avoided. PMID- 15290417 TI - Effects of oral atenolol on volatile anesthetic induction with sevoflurane in adults. AB - PURPOSE: To determine whether premedication with a beta-blocker can bring about a more rapid and smooth induction of anesthesia, we investigated the effect of oral premedication with atenolol on volatile anesthetic induction with sevoflurane by monitoring the cardiac output (CO) and bispectral (BIS) index. METHODS: Twenty four patients undergoing general anesthesia with endotracheal intubation were randomly divided into two groups: a control group ( n = 12) and a beta-blocker group ( n = 12). Each patient in the beta-blocker group was premedicated with oral atenolol 25 mg 1 h before the induction of anesthesia. Anesthesia was induced by the repeated vital capacity technique with 5% sevoflurane and 66% nitrous oxide. The trachea was intubated 5 min after sevoflurane exposure. The CO and BIS index, as well as the induction time and specific side effects of induction (e.g., movement of limbs), were recorded. RESULTS: There were no significant differences in induction time and specific side effects between the groups. The downward-sloping part of the BIS index curve in the beta-blocker group (mean, 2.9 +/- 0.2) was significantly sharper than that in the control group (2.5 +/- 0.2), and the BIS index after induction of anesthesia was significantly lower in the beta-blocker group (21.0 +/- 2.2) than in the control group (24.2 +/- 2.0). CO in the beta-blocker group was significantly lower than in the control group during the study period. The hemodynamic changes caused by endotracheal intubation were inhibited in the beta-blocker group but not in the control group. CONCLUSION: Oral premedication with 25 mg of atenolol provides a more rapid decrease in BIS index and is recommended for use in stable volatile anesthetic induction with sevoflurane. PMID- 15290418 TI - Addition of noradrenaline to intrathecal morphine augments the postoperative suppression of natural killer cell activity. AB - PURPOSE: Intrathecal administration of morphine has been shown to suppress natural killer (NK) cell activity. We tested the hypothesis that combined administration of morphine and noradrenaline would further modify NK cell activity in patients undergoing hysterectomy. METHODS: Thirty female patients were randomly divided into three groups of ten patients each. Groups MN and M received intrathecal morphine (0.5 mg) dissolved in 5 ml of physiological saline with and without 5 micro g noradrenaline, respectively. Group C received saline alone. After the intrathecal administration, general anesthesia was induced. Blood samples were withdrawn before and 2 h after surgery and on postoperative days 1, 2, and 7 to determine the NK cell activity, the ratio of T-helper/inducer cells (CD4) to T-suppressor/cytotoxic cells (CD8), the levels of interleukin-6 (IL-6) and interleukin-8 (IL-8), and the plasma concentrations of catecholamines and cortisol. RESULTS: NK cell activity decreased on postoperative day 1 in groups MN (12.0 +/- 2.7%) and M (25.4 +/- 9.6%) compared with their respective baseline levels. In group MN, NK cell activity remained lower (23.7 +/- 8.0%) on postoperative day 2 than the baseline value before surgery. CONCLUSION: Intrathecal administration of morphine causes a decrease in NK cell activity, and its combined use with noradrenaline prolongs the suppression of NK cell activity. PMID- 15290419 TI - KB-R9032, newly developed Na(+)/H(+) exchange inhibitor, attenuates reperfusion induced arrhythmias in isolated perfused rat heart. AB - PURPOSE: This study was conducted to elucidate the effects of KB-R9032, a newly developed Na(+)-H(+) exchange inhibitor, on reperfusion-induced ventricular arrhythmia in the isolated perfused rat heart. METHODS: Male Wistar rat hearts ( n = 48; 12 for each group) were perfused with modified Krebs-Ringer's solution equilibrated with 5% carbon dioxide in oxygen by means of the Langendorff technique. An occluder was placed around the left anterior descending coronary artery (LAD). Heart rate, coronary flow, and ECG were monitored. Drug-free perfusate was used for 10 min before switching to a perfusate containing various concentrations of KB-R9032. The added concentrations of KB-R9032 varied in the range of 0 (control) to 1 x 10(-5) mol x l(-1). Each heart was subjected to regional ischemia (occlusion of LAD for 11 min) and to 3 min of reperfusion (release of the ligation). RESULTS: In the control group, reperfusion-induced ventricular fibrillation (VF) occurred in 91.7%, and the duration was 158.2 +/- 14.4 s (mean +/- SEM); however, 1 x 10(-7), 1 x 10(-6), and 1 x 10(-5) mol x l( 1) KB-R9032 reduced the incidence of VF to 75.0%, 42.9%, and 6.7%, respectively ( P < 0.05 at 1 x 10(-5) mol x l(-1) of KB-R9032) and reduced the duration of VF to 64.8 +/- 22.1, 16.8 +/- 10.1, and 1.2 +/- 1.2 s, respectively ( P < 0.05 at 1 x 10(-6) and 1 x 10(-5) mol x l(-1) of KB-R9032). CONCLUSION: It was shown in this study that the Na(+)/H(+) exchange inhibitor KB-R9032 suppresses reperfusion arrhythmias in the ischemia-reperfusion model of isolated rat heart. PMID- 15290420 TI - The volatile anesthetics halothane and isoflurane differentially modulate proinflammatory cytokine-induced p38 mitogen-activated protein kinase activation. AB - PURPOSE: Volatile anesthetics affect the cardiovascular and immune systems. Toward a better understanding of the molecular mechanisms behind the modulation exerted by these agents, we focused on the effects of halothane and isoflurane on the activation of p38 mitogen-activated protein kinase (MAPK), which plays a critical role in the cellular responses to extracellular stimuli such as lipopolysaccharide (LPS) and proinflammatory cytokines, including tumor necrosis factor (TNF) and interleukin 1 (IL-1). METHODS: Human umbilical vein endothelial cells and HeLa cells, an established cell line, were examined by molecular biological methods. Cells were treated with proinflammatory compounds with or without the volatile anesthetics. p38 MAPK activation was investigated by Western blotting analysis with phosphospecific anti-p38 MAPK antibodies. RESULTS: Isoflurane activated p38 MAPK by itself, but halothane did not. Halothane or isoflurane augmented the LPS- and TNF-induced activation of p38 MAPK. In contrast, neither halothane nor isoflurane enhanced the p38 MAPK activation induced by IL-1. Neither of the anesthetics affected H(2)O(2) or MAPK kinase 3 (MKK3)-induced p38 MAPK activation. CONCLUSION: Our in vitro results indicate that the volatile anesthetics used in the clinical field and in animal experiments modify the p38 MAPK signaling cascade and suggest that the target molecules of the anesthetics are not unique and the anesthetics regulate them differentially at clinically relevant doses. PMID- 15290421 TI - JTV-506, a new K(ATP) channel opener, relaxes pulmonary artery isolated from monocrotaline-treated pulmonary hypertensive rats. AB - PURPOSE: Vasodilators are considered effective in the treatment of pulmonary hypertension if the vascular reactivity remains reversible. This study was designed to investigate the possibility that JTV-506, a new adenosine trisphoshate-sensitive potassium channel opener, may serve as a useful vasodilator in the treatment of pulmonary hypertension. METHODS: After approval by the animal care committee, with the use of the isometric-force recording method, the effects of JTV-506 (1 nM-100 micro M) on the contractile response to norepinephrine (0.1 micro M) were examined in pulmonary arteries isolated from monocrotaline (MCT)-treated (i.e., presumed pulmonary hypertensive) and age matched control rats. The experiments were performed in the presence of endothelium with or without treatment with N(G)-nitro- l-arginine (L-NAME, 100 micro M), and in its absence. RESULTS: JTV-506 relaxed ( P < 0.05) norepinephrine preconstricted, endothelium-intact arteries from MCT-treated and control rats. However, the vasorelaxation was greater ( P < 0.05) in the arteries from MCT treated rats than in controls. L-NAME treatment attenuated ( P < 0.05) vasorelaxation in the arteries from both MCT-treated and control rats. However, endothelial removal attenuated ( P < 0.05) vasorelaxation only in the arteries from MCT-treated rats and not in control arteries. CONCLUSION: JTV-506 may possibly attenuate pulmonary vascular tone through its direct action on vascular smooth muscle cells. In the presence of MCT-induced pulmonary hypertension, JTV 506 may further attenuate pulmonary vascular tone through its direct action on endothelial cells, possibly by stimulating the endothelial release of NO. PMID- 15290422 TI - Changes in plasma total and ionized magnesium concentrations and factors affecting magnesium concentrations during cardiac surgery. AB - The purpose of this study was to measure blood total and ionized magnesium concentrations ([TMg] and [Mg(2+)], respectively) and to investigate factors that might be affecting their changes during cardiac surgery using hypothermic cardiopulmonary bypass. Eight patients were examined. All the patients received diuretics and predeposited autologous blood during surgery. No drugs containing Mg(2+) were administered. Nine blood samples and eight urine samples were collected from the pre-induction period to the end of surgery. Hematocrit, [TMg], [Mg(2+)], plasma concentrations of calcium ([Ca(2+)]), creatinine, parathyroid hormone (PTH), urinary concentrations of TMg, and creatinine were measured, and the fractional excretion of Mg (FEMg) was calculated. Both [TMg] and [Mg(2+)] decreased significantly in the prebypass period and remained significantly depressed thereafter. The ionized fraction of magnesium ([Mg(2+)]/[TMg]) was decreased during the postbypass period. Hematocrit decreased significantly from the prebypass period, and FEMg increased significantly after aortic cross clamping. In conclusion, hemodilution and renal loss were main causes of hypomagnesemia, and citrate in predeposited autologous blood may contribute to the decrease in [Mg(2+)]/[TMg] in the postbypass period. These results suggest that magnesium supplementation under close monitoring of [Mg(2+)] should be required during cardiac surgery. PMID- 15290423 TI - Anesthetic management of a patient undergoing cardioverter defibrillator implantation: usefulness of transesophageal echocardiography and near infrared spectroscopy. AB - A case of a patient with sustained ventricular tachycardia (VT) undergoing implantable cardiovertor defibrillator (ICD) implantation, using transesophageal echocardiography (TEE) and near infrared spectroscopy (NIR) is described. A 67 year-old man with sustained VT associated with old myocardial infarction underwent ICD implantation. Anesthesia was induced with fentanyl and propofol and maintained with nitrous oxide, oxygen, sevoflurane, and fentanyl. Global hypokinesis of the left ventricle was observed in the short-axis view provided by TEE. Intraoperative systolic blood pressure was maintained between 100 and 120 mmHg, and cerebral oxygenated hemoglobin (HbO2) was between 63% and 65%. During periods of induced ventricular fibrillation, systolic blood pressure decreased to 60 mmHg, HbO2 decreased to 59%, and TEE revealed cardiac arrest. These changes were transient; HbO2 returned to baseline values immediately after the restoration of normal rhythm. TEE confirmed no remarkable change in cardiac function after defibrillation testing. TEE and NIR were found to be beneficial for the anesthetic management of a patient with sustained VT who was underdoing ICD implantation. PMID- 15290424 TI - Acute thrombus formation in the left atrium after the termination of warfarin. AB - We report a case of acute thrombosis formation in the left atrium 3 days after the discontinuation of warfarin therapy prior to mitral valve replacement in a patient with mitral stenosis and atrial fibrillation. A 58-year-old Asian female patient was scheduled for mitral valve replacement for mitral stenosis. She had received warfarin therapy every day for 2 years. Warfarin therapy was discontinued 3 days before surgery. Using transesophageal echocardiography (TEE), we confirmed that there was no thromboembolism at the left atrium 10 days before surgery. No replacement anticoagulant therapy, such as heparin, was given after the discontinuation of warfarin. After the induction of anesthesia, a TEE probe was inserted through the esophagus to monitor left ventricular function. We found two thrombi (35 mm and 40 mm in diameter) in the left atrium. This case shows that discontinuation of warfarin therapy within a few days before operation carries a risk of thromboembolism formation. PMID- 15290425 TI - Isoflurane increases, but sevoflurane decreases blood concentrations of melatonin in women. AB - The blood concentrations of melatonin are elevated by stress-induced sympathetic nerve excitation and are affected by some anesthetics. Isoflurane has an effect to increase sympathetic nerve activity when compared with sevoflurane. This study was performed to investigate the effects of these two anesthetics on the blood concentrations of melatonin. Female patients were anesthetized with either isoflurane or sevoflurane. We obtained blood samples before and 5 min after 5% isoflurane (ISO group) or 7% sevoflurane (SEV group) anesthesia. The blood melatonin concentrations during anesthesia in the ISO group increased significantly, from 65 +/- 60 to 170 +/- 90 pg x ml(-l); mean +/- SD ( P < 0.05), whereas those in the SEV group decreased, from 60 +/- 50 to 30 +/- 30 pg x ml(-l) ( P < 0.05). In conclusion, isoflurane increases, but sevoflurane decreases blood melatonin concentrations. PMID- 15290426 TI - Changes in femoral vein blood flow velocity by intermittent pneumatic compression: calf compression device versus plantar-calf sequential compression device. AB - Intermittent pneumatic compression has become widely used to prevent deep venous thrombosis potentially causing fatal pulmonary embolism. Although uniform compression has been commonly applied, a new method of sequential compression from plantar to calf has recently been developed. In this report, changes in maximum blood flow velocity in the femoral vein were compared with compression of only the calf uniformly and compression from plantar to calf sequentially in 10 healthy adult volunteers. A compression pressure of 60 mm Hg was applied for 5 min, and the velocity was measured before and after this treatment by ultrasound echography. There was no statistically significant difference in the change in maximum velocity between calf compression and plantar-calf sequential compression. The maximum velocity increased significantly with both compressions. However, plantar-calf sequential compression tended to have a greater effect. Although the results did not demonstrate an advantage of plantar-calf sequential compression compared with calf compression only, if the former compression is applied for a long time, it may have a greater effect. PMID- 15290427 TI - Relationship between symptoms of ischemic heart disease and upper abdominal digestive organ disease. AB - We previously reported that percutaneous trans-arterial coronary angioplasty (PTCA) seemed to decrease cardiac complications in patients with ischemic heart disease (IHD) who underwent abdominal surgery. After the report, 1293 PTCAs were performed for patients with IHD in our institute. Of these 1293 patients, 6 patients underwent abdominal surgery under general anesthesia within 14-150 days after successful PTCA. We observed the relationship between symptoms of IHD and upper abdominal digestive organ disease (ADOD), which sometimes occurs concomitantly in IHD patients. In conclusion, the present study identified some features of the chief complaints and symptoms of IHD patients with concomitant upper ADOD. This information should prove useful for making a differential diagnosis and deciding treatment. PMID- 15290428 TI - Relationship between morphine and radiotherapy for management of symptomatic bone metastases from lung cancer. AB - To determine whether radiotherapy is effective for reducing morphine dose in patients with bone metastasis from lung cancer, a retrospective study was undertaken of 58 patients who had undergone palliative radiotherapy. Mean dose of radiotherapy was 20.26 (range 8-60) Gy. Daily morphine dose after start of radiotherapy was significantly greater than before radiotherapy, and dose of morphine did not significantly decrease. Radiotherapy appears ineffective for reducing morphine dose in patients with bone metastasis from lung cancer. PMID- 15290431 TI - Winter North Atlantic Oscillation, temperature and ischaemic heart disease mortality in three English counties. AB - As cold weather is an ischaemic heart disease (IHD) risk factor, year-to-year variations of the level of IHD mortality may be partly determined by inter-annual variations in winter climate. This paper investigates whether there is any association between the level of IHD mortality for three English counties and the winter North Atlantic Oscillation (NAO), which exerts a fundamental control on the nature of the winter climate over Western Europe. Correlation and regression analysis was used to explore the nature of the association between IHD mortality and a climate index (CI) that represents the interaction between the NAO and temperature across England for the winters 1974-1975 to 1989-1999. Statistically significant inverse associations between the CI and the level of IHD mortality were found. Generally, high levels of winter IHD mortality are associated with a negative CI, which represents winters with a strong negative phase of the NAO and anomalously low temperatures across England. Moreover, the nature of the CI in the early stages of winter appears to exert a fundamental control on the general level of winter IHD mortality. Because winter climate is able to explain a good proportion of the inter-annual variability of winter mortality, long-lead forecasting of winter IHD mortality appears to be a possibility. The integration of climate-based health forecasts into decision support tools for advanced general winter emergency service and capacity planning could form the basis of an effective adaptive strategy for coping with the health effects of harsh winters. PMID- 15290432 TI - Relationship between latex yield of Hevea brasiliensis and antecedent environmental parameters. AB - A study on the relationship between latex yield and antecedent environmental data was undertaken for five clones (RRII203, RRII118, RRIM600, RRII105 and GT1) of Hevea brasiliensis (rubber) in Agartala, northeast India, a region in which rubber is not traditionally cultivated. The explained variance for the regression equations based on parameters determined on the day of tapping and up to 3 days prior to it, varied from 72% to 37% during the NWT period and 94-83% during the WT period. Soil moisture storage, 1 and 3 days prior to tapping, was found to be the primary parameter affecting yield for the NWT and WT periods, respectively. It was observed that the clone RRII105, with a comparatively lower yield to that of RRIM600, was more susceptible to daily WD conditions during the non-winter season. RRIM600 and RRII105 being high-yielding clones were also found to be fairly dependent on the AT of the day prior to tapping. The mean lag period correlation of this parameter with yield was also found to be higher during the WT period than during the NWT period. As a whole, the mean lag period based on prior measurements of environmental variables showed optimum correlation with yield at 15-20 days prior to the day of tapping. The study also confirms that varied responses of yield with environmental factors in this non-traditional region of rubber cultivation depend on clonal character. PMID- 15290433 TI - Mortality impact of extreme winter temperatures. AB - During the last few years great attention has been paid to the evaluation of the impact of extreme temperatures on human health. This paper examines the effect of extreme winter temperature on mortality in Madrid for people older than 65, using ARIMA and GAM models. Data correspond to 1,815 winter days over the period 1986 1997, during which time a total of 133,000 deaths occurred. The daily maximum temperature (T(max)) was shown to be the best thermal indicator of the impact of climate on mortality. When total mortality was considered, the maximum impact occurred 7-8 days after a temperature extreme; for circulatory diseases the lag was between 7 and 14 days. When respiratory causes were considered, two mortality peaks were evident at 4-5 and 11 days. When the impact of winter extreme temperatures was compared with that associated with summer extremes, it was found to occur over a longer term, and appeared to be more indirect. PMID- 15290434 TI - Climate factors influencing bacterial count in background air samples. AB - Total (as opposed to culturable) bacterial number counts are reported for four sites in the United Kingdom measured during campaigns over four separate seasons. These are interpreted in relation to simple climatic factors, i.e. temperature, wind speed and wind direction. Temperature has a marked effect at all four sites with data for a rural coastal site conforming best to a simple exponential model. Data for the other rural and urban locations show a baseline similar to that determined at the coastal rural location, but with some very significant positive excursions. The temperature dependence of bacterial number is found to conform to that typical of bacterial growth rates. At the coastal rural location, bacterial numbers normalised for temperature show no dependence on wind speed whilst at the inland sites there is a decrease with increasing wind speed of the form expected for a large area source. Only one site appeared to show a systematic relationship of bacterial concentrations to wind direction that being a site in the suburbs of Birmingham with highest number concentrations observed on a wind sector approaching from the city centre. PCR techniques have been used to identify predominant types of bacteria and results are presented which show that Bacillus was the dominant genus observed at the three inland sites during the winter and summer seasons. Pseudomonas appeared with comparable frequency at certain sites and seasons. There was in general a greater diversity of bacteria at the coastal site than at the inland sites. PMID- 15290435 TI - [Pain prevention allows patients with chronic pruritus to itch from central sensitivity for itching]. PMID- 15290439 TI - One-trocar appendectomy. PMID- 15290436 TI - [Pain assessment and pain treatment in the geriatric patient. Part II: pain treatment]. AB - A primary goal of pain treatment in geriatric patients consists of maintaining physical and mental function, which is a precondition of activity and participation. In patients with chronic pain, multidisciplinary treatment without excluding invasive procedures is the most effective approach. The medication ladder, suggested by the WHO initially for cancer pain, provides a guideline for pharmacological treatment. Due to age-dependent alterations in the kinetics and dynamics of pharmaceuticals, the titration of the medication follows the rule "start low and go slow." The same principle holds true for the maintenance or augmentation of physical activity in order to escape from the activity deconditioning cycle. The training should be based on learning theories, include pain management strategies, and incorporate psychological approaches to facilitate the active participation of the patient in the treatment program. In hospitals and nursing homes, nurses play an important role in defining the need for pain treatment and in supervising the patient in the treatment process. Despite all these endeavors, a significant number of patients remains whose pain cannot be controlled sufficiently. Euthanasia on demand of the patient with untreatable pain is not admitted in Germany. PMID- 15290452 TI - Pathophysiology of pulmonary aspergillosis. AB - A description of the pathophysiology of aspergillosis is followed by a review of investigational considerations of animal models. Because a large body of invasive Aspergillus infection occurs as opportunistic infection, there is a large spectrum of the histopathological feature of lesions demonstrated at the site of infection. Histopathology of the lesions can be understood as a phenotypical representation of interaction between lowered defense mechanisms in the host and the virulence of invading fungi. Detailed observations with a consideration of previous pathological knowledge of infection and inflammation provide much important information useful in predicting the pathophysiology of the patient. Moreover, experimental studies can also provide much insight to elucidate pathogenesis of the infection that emerges from the clinical and pathological investigations. The importance of pathophysiology should be emphasized to understand the implications of radiographic images, clinical symptoms, and laboratory dates. By reviewing these, especially computed tomography (CT) images, we can see that they accurately mirror the histological features of the lesion that can be recognized as a phenotypical representation of pathophysiology of Aspergillus infection. This is also confirmed by the reports emphasizing the importance of CT scans to identify hallmark clinical signs and symptoms of the disease. PMID- 15290453 TI - In vitro antimicrobial activity of T-91825, a novel anti-MRSA cephalosporin, and in vivo anti-MRSA activity of its prodrug, TAK-599. AB - TAK-599 is a water-soluble prodrug of a cephalosporin compound, T-91825. In vitro and in vivo antibacterial activities of T-91825 and TAK-599, respectively, were examined. T-91825 was active against both gram-positive and gram-negative bacteria, unlike vancomycin and linezolid, which are inactive against gram negative bacteria. The 90% minimum inhibitory concentration of T-91825 against clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA) was 2 micro g/ml. This activity was comparable to those of vancomycin, linezolid, teicoplanin, and arbekacin. T-91825 was similarly active against vancomycin intermediate S. aureus. In a time-kill study, T-91825 showed more rapid and distinct decrease of viable cells of two MRSA strains than did vancomycin and linezolid in vitro. The effect of TAK-599 against systemic infection caused by clinical isolates of MRSA in mice was comparable or superior to that of vancomycin, linezolid, teicoplanin, and arbekacin. In addition, TAK-599 at a dose of 20 mg/kg significantly decreased bacterial counts in lungs of mice in an experimental pneumonia model caused by MRSA in which vancomycin and linezolid were totally ineffective at the same dose. These results suggest the usefulness of TAK-599 in the treatment of MRSA infections in humans. PMID- 15290454 TI - Influence of daily low-dose 14-membered-ring macrolide therapy on Helicobacter pylori infection in patients with chronic inflammatory disease of the airway. AB - Our aim was to evaluate the effects of eradication and the incidence of secondary resistance by long-term low-dose daily 14-membered-ring macrolide therapy on Helicobacter pylori ( H. pylori) infection in patients with chronic lower respiratory tract inflammatory disease. In a retrospective analysis, we studied the seroprevalence of H. pylori IgG in 90 patients with inflammation of the lower respiratory tract (68 had been treated with macrolide and 22 served as controls). Then, in a prospective analysis, we evaluated the eradication effect of macrolide therapy by the decline of IgG values and the (13)C-urea breath test. Only long term macrolide use significantly affected the seroprevalence of H. pylori IgG. However, macrolide therapy did not reduce the H. pylori IgG values in 24 patients and did not eradicate H. pylori in (13)C-urea breath tests. Chemosensitivity testing was performed on three H. pylori strains obtained by gastric biopsy from patients in whom the disease could not be eradicated. Only one strain demonstrated a resistant character. Daily long-term low-dose 14-membered-ring macrolide therapy for patients with lower respiratory inflammatory disease may not be sufficient to eradicate H. pylori, but some strains do not acquire a resistant nature. PMID- 15290455 TI - Incidence of sexually transmitted diseases in Hokkaido, Japan, 1998 to 2001. AB - The objective of this study was to provide precise data on the incidence of sexually transmitted diseases (STDs) in Hokkaido. The goal of this prospective surveillance, study was to clarify the STD incidence between 1998 and 2001 in Hokkaido, Japan. The incidence of gonococcal infection in men was found to be 127 199 per 100 000 people per year, which was three or four times higher than that for women. Female genital chlamydial infection had an incidence of 300-400 with a female to male ratio of two or three to one. Younger adults had higher incidences of gonococcal and chlamydial infections than older people. In conclusion, the current study of STDs revealed high incidences of gonococcal and chlamydial infections in the Hokkaido area, and there was no decreasing trend in STD incidence during these 4 years. PMID- 15290456 TI - Prophylactic antimicrobial agents in urologic laparoscopic surgery: 1-day versus 3-day treatments. AB - Although the incidence of surgical site infection (SSI) is generally low in laparoscopic urologic surgery, the standard protocol for prophylactic use of antimicrobial agents remains to be established. We retrospectively compared the incidence and severity of SSI after laparoscopic surgery between two different protocols for prophylactic use of antimicrobial agents. This study included 114 patients who underwent urologic laparoscopic surgery categorized as "clean" or "clean-contaminated" in Sapporo Medical University School Hospital between January 1996 and October 2002. As a prophylactic antimicrobial agent, one of the cephalosporins or penicillins was administered intravenously to all patients. For 46 consecutive patients between January 1996 and July 2000, an antimicrobial agent was given 30 min before operation and thereafter every 12 h on the same day and the next 2 days after operation (the 3-day group). For 68 consecutive patients from August 2000 to October 2002, an antimicrobial agent was given once 30 min before operation and was additionally given only in the evening or night of the day of operation (the 1-day group). The incidence of SSI was retrospectively investigated. There were two patients who developed SSI in each group (4.3% in the 3-day group and 2.9% in the 1-day group). The incidence of SSI was not significantly different between the two groups. The 1-day protocol has efficacy equal to that of the 3-day protocol in prophylaxis of SSI. The 1-day use of a prophylactic antimicrobial agent may be recommended for the clean or clean contaminated urologic laparoscopic surgery described above. PMID- 15290457 TI - Thoracic actinomycosis with mainly pleural involvement. AB - A 61-year-old man, who had been diagnosed with pleuritis 5 months earlier, was admitted to our hospital to determine if a tumor shadow that appeared in his right lower lung field on March 2002 was a localized pleural mesothelioma. Although a CT-guided lung biopsy was performed, no definite diagnosis was made. However, because the tumor shadow continued to increase in size, we could not rule out the possibility of a malignant thoracic tumor, and performed video assisted thoracoscopic surgery (VATS). Histological examination of the surgically resected tissue led to a diagnosis of thoracic actinomycosis in the main component of the pleura. Because the recurrence of pulmonary actinomycosis was also suspected after surgical treatment, penicillin G was administered intravenously and afterward amoxicillin was administered intraorally. Subsequently, the patient's clinical status improved. We considered a case of thoracic actinomycosis that was suspected to have spread directly from the lung to the chest wall with complicating pleural effusion and remained with organization because there was a pulmonary infiltration shadow in the right upper lobe on chest CT at the first admission. PMID- 15290458 TI - Strongyloidiasis in a patient with acquired immunodeficiency syndrome. AB - Rhabditiform larvae, transforming larvae from rhabditiform to filariform, and eggs of Strongyloides stercoralis were identified in the sputum of a Thai woman with acquired immunodeficiency syndrome (AIDS), and stool microscopy also showed a heavy load of rhabditiform larvae of S. stercoralis. She was treated with 12 mg ivermectin once a day for 2 days for the strongyloidiasis, with good therapeutic results being obtained. Strongyloidiasis may be a curable disease through the use of an appropriate therapy, even in a patient with AIDS. PMID- 15290459 TI - Augmented inhibition of Candida albicans growth by murine neutrophils in the presence of a tryptophan metabolite, picolinic acid. AB - The effects of picolinic acid (PLA), a product of tryptophan catabolism, on anti- Candida activity of neutrophils were studied. Casein-induced peritoneal neutrophils of C3H/He mice partially inhibited mycelial growth of Candida albicans when cultured with C. albicans for 16 h in vitro. The growth inhibition of Candida was augmented by a combination of neutrophils and more than 4 mM picolinic acid. Especially in the presence of 200 U/ml murine interferon-gamma (IFN-gamma), 2 mM picolinic acid augmented the anti- Candida activity of neutrophils. The physiological significance of the augmenting effects of picolinic acid is discussed. PMID- 15290460 TI - The activity of grepafloxacin in two murine models of Mycobacterium avium infection. AB - The activity against Mycobacterium avium complex (MAC) of varying doses of grepafloxacin (GRE; 25 mg/kg, 50 mg/kg, 100 mg/kg, and 200 mg/kg) were compared to clarithromycin (CLA; 100 mg/kg and 200 mg/kg), ethambutol (EMB; 100 mg/kg), and rifabutin (RBT; 10 mg/kg) using an intranasal (IN) infection model compared to an intravenous (IV) infection model. Beige mice (C57BL6/J-Lyst bg J/+) were infected intranasally with about 10(6) organisms and for the IV model about 10(7) organisms. Treatment for both models was started 1 week postinfection and given by gavage 5 days/week for 4 weeks. At the initiation of therapy, an early control group was killed to determine the initial organism load. Three days following the completion of therapy, drug-treated groups of mice and the late control group were killed and the response to therapy measured. The most effective agents were CLA and RBT. GRE and EMB had modest activities in both the IN and the IV models. A matched comparison between IN and IV challenges for each of the agents used revealed greater suppression of MAC in the IN model compared to the IV model. PMID- 15290461 TI - High-risk populations for nasal carriage of methicillin-resistant Staphylococcus aureus. AB - To determine the population at high risk of nasal carriage of methicillin resistant Staphylococcus aureus (MRSA) on hospital patients admission, a nasal swab was taken from the following patients: (1) those aged 70 years or over (age >or= 70), (2) non ambulatory receiving regular home visits by nurses and physicians (visiting), (3) residents of nursing homes (nursing home), (4) patients from other hospitals (another Hp), and (5) those scheduled for surgery (presurgery). Between March and July 2000, a total of 412 patients were admitted and 136 were enrolled. MRSA was isolated from 12 (8.8%) patients. The number of patients positive for MRSA in the five groups, age >or=70, visiting, nursing home, another Hp, and presurgery, were 3 of 68, 3 of 21, 2 of 3, 3 of 9, and 1 of 35, respectively. Multivariate analysis revealed that living in a nursing home [odds ratio (OR) = 32.82, P = 0.010] or coming from another hospital (OR = 14.55, P = 0.0043) were high risk factors with for nasal carriage of MRSA. Furthermore, patients' ages were further divided into three categories, or=90, and regarded as independent high risk factors (OR = 3.08, P = 0.043). The results were that advanced living in a age (>or=80, >or=90), living in a nursing home or coming from another hospital are high risk factors of nasal carriage of MRSA on hospital admission. PMID- 15290464 TI - Advanced imaging application for acute ischemic stroke. AB - With the approval of intravenous recombinant tissue plasminogen activator (r-TPA) in 1995, acute ischemic stroke therapy is increasingly being administered. Currently the approach to imaging these patients is very simplistic. Typically, noncontrast head computed tomography (CT) is the only study performed prior to treatment. Advanced imaging using CT or magnetic resonance imaging (MRI) can play a very important role in the triage and classification of patients with acute ischemic stroke. With knowledge of the location and size of the occlusion as well as the collateral circulation, the best treatment can be selected, minimizing any morbidity from treatment and maximizing the chance of success. The identification and stratification of patients according to their imaging and clinical features will further individualize treatment and allow tailored therapy. This review will discuss rapid imaging techniques that are easily available and the rationale for their use. PMID- 15290465 TI - Call back or else. PMID- 15290466 TI - Rapid CT scan visualization of the appendix and early acute non-perforated appendicitis using an improved oral contrast method. AB - The purpose of this study was to optimize detection of the normal appendix in the clinical exclusion of acute nonperforated appendicitis using an improved and rapid method of bowel opacification in conjunction with the CT examination. A prospective evaluation of 100 consecutive patients, ranging from 13 to 50 years in age, was performed over a 4-month period using water-soluble oral contrast medium consisting of a fixed dose of diatrizoate salts administered as a prepared beverage in the emergency ward 50 min prior to performing a CT scan to evaluate clinical signs and symptoms of early acute appendicitis. The appendix was visualized in 84% (84 of 100) of patients, with a mean transit time of 50 min. The appendix filled with oral contrast medium in 89% (75 of 84) patients, and this sign was reliable in excluding appendicitis. In no instance did a contrast filled appendix prove to represent appendicitis. The earliest signs of appendicitis were seen in 8% (8 of 100) patients. CT scan findings included absence of a contrast- or air-containing appendix with appendiceal thickening and infiltration of the periappendiceal mesenteric fat. CT scan utilizing a fixed dosage of orally administered water-soluble contrast containing diatrizoate salts, with a mean transit time of 50 min, provides a rapid and efficient means of visualizing the appendix in the clinical exclusion of appendicitis in the emergency setting. PMID- 15290467 TI - Controversies in emergency radiology: acute appendicitis in children--the case for CT. AB - Acute appendicitis is the most common condition requiring surgical intervention in childhood. The clinical diagnosis of acute appendicitis is often not straightforward and can be challenging. Approximately one-third of children with the condition have atypical clinical findings and are initially managed nonoperatively. There is currently great variability in the utilization of imaging for the assessment of suspected acute appendicitis in children. The principal imaging modalities utilized are graded-compression sonography and CT. Sonography has important diagnostic limitations that are addressed by CT. The principal advantages of CT include its operator independency, with resultant higher diagnostic accuracy, enhanced delineation of disease extent in perforated appendicitis, and improved patient outcomes including decreased negative laparotomy and perforation rates. PMID- 15290468 TI - Soft tissue injury protocol (STIP) using motion MRI for cervical spine trauma assessment. AB - The purpose of this study was to establish a noninvasive scoring method, using motion MRI, to determine the degree of clinical impairment in traumatized cervical spines. This method is called the soft tissue injury protocol (STIP) scoring method. The cervical spines of 100 adult accident victims were evaluated prospectively using motion MRI at 12 weeks following hyperflexion/hyperextension injury from rear, low-impact motor vehicle collisions. Subjects were scored for degree of functional impairment based on an eight-point scale derived from the following clinical criteria: hypolordosis, motion restriction, disk herniation, and spinal stenosis. Five classes of impairment, ranging from normal to severe impairment, were identified as a basis for therapeutic management. Using the STIP scoring method, 94% of patients (94 of 100) were determined to have nonsurgical injuries. Class 1 and 2 injuries indicated mild impairment and were found in 68% of patients, who were considered to have reached maximum medical improvement at 12 weeks after injury. A Class 3 injury indicated moderate impairment and was found 26% of patients, who required an additional 12 weeks of rehabilitative and medical treatment to achieve maximum medical improvement. Class 4 and 5 injuries indicated severe impairment; these were identified in 6% of patients and required surgical intervention. Five of the six patients requiring surgery (83%) achieved maximum medical improvement at 36 weeks after injury. The STIP scoring method is a practical, noninvasive method of determining the degree of clinical impairment, as a basis for distinguishing injury requiring medical treatment from injury requiring surgical treatment, in cases of subacute cervical spine trauma. PMID- 15290469 TI - Pelvic imaging in the stable trauma patient: is the AP pelvic radiograph necessary when abdominopelvic CT shows no acute injury? AB - The purpose of the study was to determine the utility of anteroposterior (AP) pelvic radiographs in stable trauma patients who will undergo or have undergone abdominopelvic CT as part of the initial trauma imaging evaluation. Radiology reports of all stable trauma patients who underwent both abdominopelvic CT and AP pelvic radiograph from 25 January through 30 April, 2003 were reviewed for findings of acute pelvic injuries. A total of 509 consecutive patients were included in this series. Of these, 449 patients (88.2%) had no acute pelvic injury revealed by abdominopelvic CT. CT showed 163 acute injuries in 60 patients. AP radiographs showed 132 acute injuries in 52 patients. No patients with a negative CT had an acute finding on the radiograph. There were eight false negative pelvic radiographs (negative predictive value 98.25%). CT is highly accurate in excluding acute osseous pelvic injuries. In the stable trauma patient whose CT does not reveal an acute pelvic injury or who is scheduled to undergo an abdominopelvic CT as part of the initial imaging evaluation, the pelvic radiograph may be unnecessary. PMID- 15290470 TI - Degeneration of the costovertebral articulation: a cause of pulmonary pseudolesion. AB - The objective was to report examples of degeneration of the costovertebral articulation producing a pulmonary pseudolesion. Three cases in which a nodular opacity seen on plain radiograph was determined to be secondary to degeneration of the costovertebral articulation were compiled, one of which was confirmed by CT. Pseudolesions produced by degenerative osteophytes of the vertebral spine and anomalous articulations between transverse processes are more commonly identified, but less well described is the pseudolesion produced by degeneration of the costovertebral articulation. Recognition of this etiology may prevent misconstruing the lesion as a significant finding. PMID- 15290471 TI - Medial clavicular epiphysiolysis in children: the so-called sterno-clavicular dislocation. AB - We retrospectively reviewed six pediatric cases of medial clavicular injury, i.e., epiphyseal separation (Salter/Harris type I or II injury), diagnosed between 1993 and 1997. The clavicular metaphysis was displaced posteriorly in three cases and anteriorly in three. On conventional radiographic views the diagnosis was initially missed in two of three retrosternal dislocations. A special X-ray projection (described by Heinig) or computed tomography (CT) permitted correct diagnosis. Anterior dislocations were immediately and correctly diagnosed. Closed reduction successfully treated retrosternal displacement in two of the three patients. The third patient needed open reduction and internal fixation. Open reduction and internal fixation had to be performed in all three patients with anterior displacement. Follow-up assessment showed perfect functional results in all cases. Direct visualization during open reduction, which was necessary in four of six cases, yielded clear evidence that the so called sternoclavicular dislocation in children and young adults is, in fact, a fracture of the medial growth plate with posterior or anterior displacement of the metaphysis. PMID- 15290472 TI - Current trends in imaging evaluation of acute cholecystitis. AB - This study was designed to retrospectively determine recent clinical trends of initial radiological evaluation in patients pathologically proven to have acute cholecystitis (AC) and to assess the methodology that led to its diagnosis. Over a 28-month period, the medical records and imaging studies of 117 consecutive patients who had pathologically confirmed AC were retrospectively analyzed. The sensitivities of ultrasound (US) and hepatobiliary 99mTc-iminodiacetic acid (HIDA) were computed. The false-negative scans were retrospectively reviewed by a blinded radiologist to determine the limitations and advantages of each modality. The 117 patients were grouped into six categories based on the type of imaging examination they underwent prior to cholecystectomy: initial US evaluation only (n=80, 68.4%), initial US followed by HIDA (n=17, 14.5%), initial HIDA only (n=2, 1.7%), initial HIDA followed by US (n=3, 2.6%), initial CT (n=5, 4.3%), and no imaging evaluation (n=10, 8.6%). HIDA scan had a calculated sensitivity of 90.9% (20 true-positive, 2 false-negative) while US had a sensitivity of 62% (62 true positive, 38 false-negative). Current practice in the initial radiological evaluation of acute cholecystitis remains outdated. The vast majority of patients in our study group were initially worked up using US, although HIDA scan has been shown to have greater sensitivity for the diagnosis of acute cholecystitis. PMID- 15290473 TI - Jacuzzi jet-induced pneumoperitoneum. AB - Pneumoperitoneum outside the setting of recent surgical intervention usually indicates perforation of the gastrointestinal tract. Following radiologic evidence of pneumoperitoneum, surgical exploration of the abdomen may be indicated depending on the nature of the patient's presentation. We present the radiological findings of a healthy young woman who presented with acute onset of abdominal pain and was found to have extensive pneumoperitoneum. No visceral disruption was evident by multidetector CT or by single-contrast barium fluoroscopic evaluation of the upper gastrointestinal tract. Knowledge of benign causes of pneumoperitoneum by the radiologist may avert an unnecessary laparotomy. PMID- 15290474 TI - Dense ascites: CT manifestations and clinical implications. AB - Ascites is a not infrequent finding on CT. Causes of ascites include congestive heart failure, hypoalbuminemia, cirrhosis, inflammation, and neoplasm. In most cases the attenuation of ascites is that of clear fluid, measuring around 0 HU. Rarely, however, a considerably higher density of ascites is seen. This finding may be a challenge for the radiologist, particularly since some of the conditions associated with it are of major clinical importance requiring prompt intervention. PMID- 15290475 TI - Emergency transjugular liver biopsies in post-liver-transplant patients: technical success and utility. PMID- 15290476 TI - Emergency suprarenal inferior vena cava filter placement in ovarian vein thrombosis. PMID- 15290477 TI - Superior mesenteric artery syndrome involving the duodenum and jejunum. AB - Superior mesenteric artery (SMA) syndrome is a rare cause of intestinal obstruction involving the duodenum. Diagnosis is based on clinical suspicion with radiologic confirmation. We report an unusual presentation of the SMA syndrome involving both the duodenum and jejunum initially not recognized on contrast enhanced CT. This case demonstrates the judicious use of multiple modalities in evaluating for this syndrome. PMID- 15290478 TI - Omental infarct: an unusual CT appearance after superior mesenteric artery occlusion. AB - We report a case of omental infarct resulting from superior mesenteric artery occlusion, which had an unusual appearance on computed tomography. PMID- 15290479 TI - Intravascular gas in the transplanted kidney: a sign of extensive graft necrosis. AB - We encountered a case of transplanted kidney necrosis, with computed tomography (CT) demonstrating multiple areas of intravascular gas within the allograft. The intravascular gas represented air emboli from gas liberated from fermentation by gas-forming organisms in a perinephric abscess. Arterial bleeding accelerated by the wound infection and the resultant large perinephric hematoma caused renal infarction. Gas-forming infection of transplanted organs is associated with a poor graft outcome, which can present as a fulminant clinical course. Intravascular gas should be distinguished from collecting system gas because the former could represent extensive necrosis of the transplanted kidney. PMID- 15290480 TI - Pulmonary atelectasis: a frequent alternative diagnosis in patients undergoing CT PA for suspected pulmonary embolism. AB - The purpose of the study was to evaluate the prevalence of atelectasis as an alternative diagnosis in patients who underwent computed tomographic pulmonary angiography (CT-PA) for suspected pulmonary embolism (PE), and to contrast the pathophysiology of pulmonary atelectasis and PE, both of which are associated with dyspnea and hypoxemia. We retrospectively identified 144 consecutive emergency department patients (n=49) and inpatients (n=95) admitted between July 2001 and June 2002 who were evaluated with CT-PA for suspected PE. There were 98 women and 46 men with a mean age of 58 years (range 27-95 years). Each CT report was reviewed for PE, the words "atelectasis," "collapse," and/or "volume loss," findings known to predispose to atelectasis, and alternative diagnoses. CT scans of those with PE and those with atelectasis were reviewed. Each case was categorized into one of three groups, as follows: group 1, PE; group 2, atelectasis of three or more segments and no PE; group 3, neither PE nor atelectasis. PaO2 was documented, when available (n=115), with PaO2 >100 mmHg recorded as 100 mmHg. Reports for group 3 were reviewed for alternative diagnoses. Thirteen percent of the study population (19/144, group 1) had PE, and two of them had concomitant atelectasis; mean PaO2 was 69 mmHg (range 38-100 mmHg). Nineteen percent of the study population (27/144, group 2) had atelectasis of three or more segments without PE; mean PaO2 was 73 mmHg (range 45-100 mmHg). Sixty-eight percent of the study population (98/144, group 3) had neither PE nor atelectasis; mean PaO2 was 79 mmHg (range 36-100 mmHg). There was a significant difference in the PaO2 between groups 1 and 3 (Student's t-test), with group 2 intermediate. Seventy percent of group 2 (19/27) had at least one finding predisposing to atelectasis: central bronchial abnormality (n=6), moderate or larger pleural effusion (n=11), pleural mass, pneumothorax, elevated hemidiaphragm, and severe kyphosis (the last four all n=1 each), versus 16% (3/19) of group 1 ( P<0.05). Sixty-three percent of group 3 (62/98) had one or more alternative diagnoses on CT that explained the patient's symptoms as follows: pneumonia (28%, 27/98), other lung disease (18%, 18/98), congestive heart failure (13%, 13/98), and malignancy (13%, 13/98). Pulmonary atelectasis was common in patients undergoing CT-PA for suspected PE, equaling pneumonia as the most common alternative diagnosis. Most patients with atelectasis had predisposing findings on CT. Pulmonary atelectasis and PE cause similar symptoms by different mechanisms of ventilation-perfusion mismatch. PMID- 15290481 TI - Sickle crisis. PMID- 15290482 TI - CT of orbital trauma. AB - In a patient with acute orbital trauma, visual acuity and extraocular muscle motility are the two most important ophthalmologic functions to be evaluated emergently. The assessment of these capabilities may sometimes be difficult due to the severity of the head injury, the extent of periorbital soft tissue edema, inadequate cooperation in alert patients, and a reduced level of consciousness in obtunded individuals. Consequently, computed tomography (CT) has come to play a major role in the orbital examination of acute trauma patients. In this study, in conjunction with clinical evaluation, we have sought to utilize CT to determine the various prevalences of the causes of decreased visual acuity and extraocular muscle motility resulting from orbital trauma. We retrospectively reviewed the records of all patients admitted to our emergency facility who, having suffered head trauma, underwent a CT study for diagnosis. CT examinations of the head using a multidetector scanner were performed from the base of the skull to the vertex at 5-mm intervals. Orbital CT was obtained when a routine CT of the head showed periorbital soft tissue edema and/or facial bone fractures. The orbital CT examination was performed using axial 1-mm and coronal 3-mm slices. Coronal reformation images were prepared if the patient was unable to tolerate the prone position for direct coronal imaging. The imaging findings were correlated with ophthalmologic observations. The orbit floor was the most common and the orbital roof the least common site of fracture of the bony coverings of the eye. Twenty three patients suffered decreased visual acuity. In order of declining frequency, the causes of reduced vision consequent to trauma were retrobulbar hemorrhage, optic nerve thickening presumably secondary to edema, intraorbital emphysema, optic nerve impingement, detached retina and ruptured globe. Five patients had visual impairment without demonstratable radiographic abnormalities. The most common finding associated with diminished extraocular muscle motility was muscle impingement by fracture fragments, followed in decreasing frequency by thickened muscle due to edema or contusion, intraconal emphysema, muscle entrapment, and retrobulbar fat herniation. Six patients with decreased extraocular muscle activity had no abnormalities demonstrated on CT images. The overwhelming majority of patients with decreased visual acuity or reduced extraocular muscle motility consequent to trauma had abnormalities demonstrated by orbital CT. Hence, CT examinations should play a major role in the evaluation of the intraorbital contents in patients with orbital trauma. PMID- 15290483 TI - Role of routine nonenhanced head computed tomography scan in excluding orbital, maxillary, or zygomatic fractures secondary to blunt head trauma. AB - The purpose of this paper is to determine the necessity of a dedicated facial bone/orbital computed tomography (CT) scan for fracture surveillance in patients who have suffered blunt head trauma and whose routine nonenhanced head CT scan is negative. It is based on a retrospective review of 115 patients presenting to the Emergency Department at a level I trauma center after blunt head trauma. Included patients underwent both a nonenhanced head CT scan and a dedicated facial bone or orbit CT. Standard nonenhanced head CT protocol was followed for each patient as per department protocol. A positive head CT scan is defined to include either an air-fluid level within the paranasal sinuses or fracture of the maxillary, orbital, or zygomatic osseous structures. A negative scan demonstrates none of these findings. Intracranial/parenchymal pathology was not evaluated in this study. Sixty-five of the 115 patients had a negative head CT scan as defined above. Of these 65 patients, none subsequently had a positive facial bone or orbit CT scan. The sensitivity and negative predictive values of a negative routine nonenhanced head CT scan for fracture surveillance are both 100%. In the setting of blunt trauma, a negative nonenhanced head CT scan precludes the need for a dedicated facial bone or orbital CT scan in the evaluation for orbital, maxillary, or zygomatic fractures. This saves the patient unnecessary radiation exposure, health care costs, and time spent in the emergency radiology department. PMID- 15290484 TI - Cervical spine trauma in children under 5 years: productivity of CT. AB - The aim of the study was to determine the value of CT in the evaluation of cervical spine trauma in children under 5 years in the Emergency Department. A retrospective review of 606 patients undergoing cervical spine examination in the emergency room was undertaken. The age and sex of the patients were documented, and in addition presence or absence of fracture-dislocation was noted on each of the plain film and CT studies. Of the 606 patients studied, 459 (75.7%) were cleared by a combination of clinical and plain film radiographic findings. The other 147 (24.3%) went on to CT imaging for clearing of the cervical spine. Of the 147 patients evaluated with CT, 143 (97.3%) had exams that were negative for fracture, dislocation, and instability. Only four (2.7%) demonstrated positive findings for fracture, dislocation, or instability. All of these patients had positive, diagnostic findings on initial plain film evaluation. Another five patients demonstrated new findings which were unrelated to trauma and of no clinical consequence. The yield of positive, clinically significant findings on CT of cervical spine injuries in children less than 5 years was low and showed significant findings only in patients where the same findings were seen on initial plain radiographs. PMID- 15290485 TI - Optimal patient position for lumbar puncture, measured by ultrasonography. AB - The purpose of this study was to identify the patient position for lumbar puncture associated with the widest interspinous distance utilizing ultrasound. Sixteen healthy adult volunteers were placed in three positions commonly used for lumbar puncture (lateral recumbent with knees to chest, sitting and bent forward over an adjustable bedside stand, and sitting with feet supported and chest to knees) and the distance between lumbar spinous processes was measured by ultrasound. Measurements were compared between the three positions. Differences were analyzed using Tukey's honestly significant difference test. The results showed that the interspinous distance was significantly greater in the "sitting, feet supported" position than in the other two positions ( P<0.001). The "sitting, feet supported" position may offer advantages for selected patients undergoing lumbar puncture. Ultrasonography may be a useful adjunct when performing lumbar puncture in the emergency department. PMID- 15290486 TI - Wrist injuries; diagnosis with multidetector CT. AB - The aim of the study was to assess acute-phase multidetector CT (MDCT) findings in wrist injuries. We retrieved all emergency room MDCT requests processed in the period from August 2000 to May 2003. All patients with a wrist injury who underwent MDCT initially were included. Imaging studies were evaluated in relation to injury mechanism, fracture location, and fracture type. A total of 6422 MDCT examinations were performed during this 34-month period, and 38 patients (24 male, 14 female, age range 21-73 years, mean age 40 years) met the inclusion criteria. MDCT revealed 56 fractures and 7 dislocations in 29 patients. In 9 patients (24%) MDCT findings were normal. Eleven patients (29%) underwent surgical procedures. The main injury mechanism was a fall (58%). In 33 cases the primary radiograph was available. Compared to primary radiographs, MDCT revealed 9 occult fractures, mainly in small carpal bones. In 14 cases a suspected fracture (of the scaphoid in 7 cases) was ruled out by MDCT. Due to high-quality two-dimensional reformatting, MDCT examinations were not dependent on the wrist's position in the CT gantry. In the comparison with radiography, MDCT detected occult fractures and ruled out suspected fractures, both mainly in the small carpal bones. High-quality two-dimensional reformats gave significant information about the fracture anatomy. MDCT provides fast and valuable information in assessing complex wrist fractures or when the primary radiograph is equivocal. PMID- 15290487 TI - Optimizing the patient positioning for PICC line tip determination. AB - Central venous catheters (CVCs) are used for both emergent and long-term vascular access for the infusion of numerous therapeutic agents such as chemotherapy, parenteral nutrition, antibiotics, and analgesics, as well as for temporary hemodialysis or hemoperfusion. Current standard of care dictates that CVC insertion should be followed by an immediate chest radiograph to confirm appropriate position. Radiographic confirmation of central venous line placement is important because it is not possible to determine CVC tip position clinically. Although many catheter tips can be localized on the standard frontal radiograph, there are occasions when a second radiograph is necessary to localize the position of the CVC tip accurately. We hypothesized that a right posterior obligue chest radiograph would more consistently enable the catheter tip to be seen as it reduces the superimposition of mediastinal structures. One hundred chest radiographs taken in an anteroposterior (AP) projection and 100 chest radiographs taken in a right posterior oblique (RPO) projection after a peripherally inserted central catheter (PICC) line placement at UCI Medical Center from June 2000 to November 2002 were read by two radiologists. Forty-one percent of AP readings were discrepant and 4% had the annotation "difficult to identify the position of the tip" although the identification of tip position was similar. Fifty-five percent of AP readings were in agreement with no note of any difficulty. Eighteen percent of RPO readings were discrepant and 2% had the annotation "difficult to identify the position of the tip" although the identification of tip position was similar. Eighty percent of RPO readings were in agreement with no note of any difficulty. PMID- 15290488 TI - Isolated severe renal injuries after minimal blunt trauma to the upper abdomen and flank: CT findings. AB - Renal injuries caused by blunt abdominal trauma are common in children. Serious renal trauma is associated with insult to other organs, whereas isolated renal injuries are usually minor. We present the cases of six male children (aged 7-17 years) with major isolated renal injuries due to minimal blunt trauma to the upper adbomen and/or the flank, out of a total of 21 children admitted with renal trauma in a 5-years period. On physical examination all patients had a painful, tender abdomen and/or flank with ipsilateral bruises and ecchymosis. Hematuria, either macro ( n=4) or micro ( n=2), was found in all. The injuries were left sided in five and were of a variable severity (grade III: n=2; grade IV: n=3; grade V: n=1 according to the kidney injury scale of the American Association for the Surgery of Trauma). Four children underwent nephrectomy. This small series underlines that major kidney insult can occur after a minimal blunt trauma localized to the flank or upper abdomen. Abdominal CT should be performed when clinical or laboratory findings or the mechanism of trauma suggest renal injury. PMID- 15290489 TI - Two cases of pseudoaneurysm of the renal artery following laparoscopic partial nephrectomy for renal cell carcinoma: CT angiographic evaluation. AB - Partial nephrectomy via a laparoscopic approach can be technically challenging, and associated vascular complications such as pseudoaneurysm may occur. CT with CT angiography is ideal for the noninvasive imaging of this process. This article reports two cases of pseudoaneurysm of the renal artery detected on CT as a complication of laparoscopic partial nephrectomy and demonstrates the usefulness of 3-D CT angiography in the evaluation of vascular pathology. PMID- 15290491 TI - CT findings of afferent loop syndrome after a subtotal gastrectomy with Roux-en-Y reconstruction. AB - A rare case of combined afferent loop syndrome following Roux-en Y reconstruction and small bowel obstruction due to adhesions at the enteroenterostomy is presented. The CT findings of the obstruction of both the afferent and the efferent limbs are demonstrated, with emphasis on the characteristic CT features of afferent loop syndrome. PMID- 15290490 TI - CT findings in patients with small bowel obstruction due to phytobezoar. AB - The role of CT in evaluating patients with small bowel obstruction (SBO) has been extensively described in the current literature. We present the CT findings of SBO due to a phytobezoar, afterwards surgically confirmed, in 5 men and 1 woman (aged 32-89 years) out of 95 patients diagnosed by CT as having SBO in a 44-month period. These six patients underwent abdominal CT prior to operation and the CT findings were retrospectively reviewed. All six patients presented with clinical symptoms and signs of SBO; three of them had undergone gastric surgery 13, 17, and 22 years earlier, respectively. In all six cases, CT showed an ovoid intraluminal mass, 3 x 5 cm in size and of a mottled appearance, at the transition zone between dilated and collapsed small bowel loops. This was in contrast to feces-like material (the "small bowel feces sign"), seen within dilated small bowel loops in nine patients with SBO, and was typically longer. As CT is frequently performed for suspected SBO, an ovoid, short intraluminal mottled mass seen at the site of an obstruction may be regarded as a pathognomonic preoperative sign of an obstructing phytobezoar. PMID- 15290492 TI - Acute gastrointestinal hemorrhage in infancy from gastric duplication: imaging findings. AB - We report two cases of duplications, occurring along the greater curvature of the stomach, both complicated by acute gastrointestinal hemorrhage, and review the findings on various imaging studies. Special attention is directed to the plain film findings, as this may be the initial study performed in the emergency department setting. PMID- 15290493 TI - Gas-forming abdominal wall abscess: unusual manifestation of perforated retroperitoneal appendicitis extending through the superior lumbar triangle. AB - We present a case of an extensive gas-forming abdominal wall abscess secondary to ruptured retroperitoneal appendicitis in a diabetic patient. Computed tomography (CT) demonstrated a retroperitoneal abscess extending to the abdominal subcutaneous tissue through the superior lumbar triangle pathway, a known anatomical defect of the lumbar musculature. This case not only represents an unusual manifestation of acute appendicitis, but also alerts us to the importance of anatomical considerations in the imaging interpretation of disease extent. The applied anatomy of the lumbar triangle as a conduit of intra-abdominal and retroperitoneal processes into the abdominal wall is reviewed. PMID- 15290494 TI - CT presentation of ruptured appendicitis in an adult with incomplete intestinal malrotation. AB - Intestinal malrotation is defined anatomically as a developmental anomaly. It may cause atypical clinical symptoms in relatively common intestinal disorders because of the altered anatomy. A 64-year-old man presented with acute mid abdominal pain. Underlying incomplete malrotation prevented the correct clinical diagnosis of ruptured appendicitis. Computer tomography demonstrated typical signs of malrotation, i.e., right-sided duodenojejunal junction, left position of cecum, inverted position of the superior mesenteric vessels, and pathology revealed a ruptured appendix with an abscess and a coincident mucinous cystadenoma. PMID- 15290495 TI - Cervical and mediastinal hematoma: presentation of an asymptomatic cervical parathyroid adenoma: case report and literature review. AB - The spontaneous rupture with extracapsular hemorrhage of a cervical parathyroid adenoma is a rare cause of cervical and mediastinal hematoma. We describe this case to emphasize that a failure to consider this diagnosis may result in delayed operative intervention with potentially fatal complications. PMID- 15290496 TI - Foreign esophageal body impaction: multimodality imaging diagnosis. AB - We report a case of a portion of bran bread impacted in the cervical esophagus in an 88-year-old woman. A complete radiologic study including ultrasonography, computed tomography, and barium swallow was performed. These techniques confirmed and localized the foreign body, which was endoscopically removed. A complete radiologic assessment can guarantee the usefulness of esophagoscopy to avoid significant morbidity from unnecessary procedures in a patient in poor clinical condition. Ultrasonography and computed tomography are attractive and profitable options in these cases. PMID- 15290497 TI - Emergency radiology coverage: technical and clinical feasibility of an international teleradiology model. AB - The purpose of the study was to determine the feasibility of international teleradiology, utilizing day-night time differences, for online interpretation of overnight computed tomography (CT) studies. One hundred and two consecutive Emergency Room patients who underwent CT examinations between the hours of 11 pm and 7 am were enrolled. All age groups and study types were included. CT studies were transmitted from the in-hospital PACS system (Kodak, Fremont, Calif.) to a web-based server (Medweb, San Francisco, Calif.). A radiologist in Bangalore, India, working an 8 amto 4 pm day shift, downloaded and reviewed the studies on a desktop PC using a 128-kbps internet connection at 10-20:1 wavelet compression and generated a report. The report was then uploaded to the server, noting the time at upload. Each study report was compared with the official in-house diagnostic report and concordance assessed on a three-point scale. Mean download time was 8.14 s per image. For head CT reports ( n=47), the mean turnaround time for a final transcribed report was 39.5 min. For abdomen/pelvis CT reports ( n=48) the mean turnaround time was 84.4 min. Out of 106 cases, there was discordance between the clinical diagnostic report and the study report in 20 (19%); however, on subsequent review the teleradiology report was found to be correct in 13 of these. Day-night time differences across the globe can be utilized to provide overnight emergency radiology coverage using web-based teleradiology. Scan download and report upload times are within acceptable limits. PMID- 15290498 TI - Radiology coverage 24/7-what can we do, who can we call? PMID- 15290499 TI - Supervision of residents by faculty radiologists using home workstations. AB - Our purpose was to determine if a home-based faculty radiologist equipped with a high-resolution workstation could add new information to residents' readings on overnight computed chest images that was equivalent to the new information generated by faculty reviewers inside the hospital. Teleconferencing software was installed on home workstations for online supervision of residents by faculty on chest images from a cardiothoracic intensive care unit. Critical observations that could affect patient care were recorded by first-year radiology residents before and after teleconferencing with the home-based radiologist. The amount of information added was compared with that which was added on the same 50 images through direct consultations with faculty inside the medical center. The amount of critical information that was added by teleconferencing with a chest radiologist at home was equivalent statistically to the information added through direct supervision of residents by faculty inside the hospital. Teleconferencing resulted in 149 changes in critical image findings as reported initially by the residents, out of 800 possible findings on 50 chest images, as opposed to 142 changes in residents' readings by faculty inside the medical center. Faculty subspecialists can supervise radiology residents effectively from their homes after hours, using high-resolution workstations and special teleconferencing software. PMID- 15290500 TI - Evaluation of occult cervical spine fractures on radiographs and CT. AB - The aim of this study was to evaluate occult cervical spine fractures on radiographs and CT. We reviewed radiographs of 68 patients with cervical spine fractures at a level 1 trauma center. Twenty-six patients showed occult fractures on initial radiographs. Of seven odontoid fractures, two were diagnosed only after thinner repeat CT reconstructions. Five facet fractures were best seen on CT reconstructions. Three occult hangman's fractures were seen by CT. A C1 fracture was diagnosed on CT only. Two fractures through the C2 body were seen only by CT. Eight fractures were obscured by patient's shoulders, including five clay shoveler's fractures, a C6 facet fracture, a bilateral C6 pedicle fracture, and a C7-T1 fracture-dislocation. Odontoid, facet, and lower cervical spine fractures are most commonly occult on radiographs. CT, especially high-quality CT reconstructions for odontoid and facet fractures, can improve the diagnosis of cervical spine fracture. PMID- 15290501 TI - Vascular lesions of the renal sinus. AB - The renal sinus contains within it the collecting system of the kidney as well as lymphatics, nerves, and renovascular structures. This area may be affected by a large variety of pathological conditions arising from the various tissues in this site. Vascular lesions of the renal sinus are uncommon and may present clinically with acute symptoms and on imaging as a mass lesion. Awareness of the different vascular lesions affecting this area is essential for establishing the correct diagnosis and for appropriate treatment. The role of computed tomography is emphasized because it is the most commonly used modality to evaluate acute abdominal conditions as well as suspected renal masses, and the diagnosis can usually be made without the need for additional imaging modalities. PMID- 15290502 TI - A case of bilateral dense middle cerebral arteries with CT angiographic confirmation of vascular occlusion. AB - Hyperattenuating middle cerebral arteries on CT in acute stroke should generally not be associated with presence of intraluminal clot when bilaterally seen. We report a case of a woman who underwent emergency CT 60 min after sudden onset of coma. Bilateral dense middle cerebral arteries without parenchymal hypoattenuating areas or indirect signs of cerebral edema were present. CT angiography confirmed occlusion of the right middle cerebral artery and left internal carotid artery and middle cerebral artery. PMID- 15290503 TI - MRI evaluation of an open globe injury. AB - The diagnosis of open globe injuries is often difficult to ascertain by physical examination and even with CT scans. MRI is used to evaluate a variety of globe pathologies and can also be used in certain patients with suspected injury of the globe. We report a case of traumatic globe injury, where MRI was utilized to accurately depict the presence and extent of open globe injury. PMID- 15290504 TI - Iatrogenic venous air embolism during contrast enhanced computed tomography: a report of two cases. AB - Venous air embolism (VAE) is a known complication of venous access procedures such as contrast-enhanced computed tomography (CECT). Although a massive VAE can be fatal, most iatrogenic VAE cases during CECT involve a few milliliters of air and are asymptomatic. We report two cases of massive and nonfatal VAE during CECT. Both cases involve the inadvertent injection of air instead of contrast by power injectors during the contrast phase. In both cases, the patients were stable and survived the event without permanent sequelae. We also discuss the pathophysiology, treatment, and prevention of VAE, especially during CECT. PMID- 15290505 TI - Unusual presentation of purulent pericarditis: diagnostic contribution of MRI. AB - Purulent bacterial pericarditis is an uncommon form of infectious pericarditis, occurring usually in immunocompromised patients. It may rarely present as a nodular or tumefactive lesion, and in such cases is liable to be confused with primary and metastatic tumors. We describe the imaging features and diagnostic contribution of CT and MRI in a 68-year-old woman with mass-like purulent pericarditis. PMID- 15290506 TI - Fatal abdominal aortic compression associated with thoracic vertebral fracture dislocation after a short fall. AB - Abdominal aortic injury after blunt trauma is a rare event, as these injuries occur much less frequently than do those of the thoracic aorta. We present an unusual case of fatal abdominal aortic compression with presumed compression associated with a fracture-dislocation of the T11 vertebral body after a short fall, which to our knowledge is the first reported case of this type. PMID- 15290507 TI - Fasting hypoglycemia in solitary fibrous tumor of the peritoneum: a diagnostic clue for an emergent situation. AB - Solitary fibrous tumors are rare fibrous tumors found in various anatomic sites. Extrathoracic neoplasms are often symptomatic at presentation, with paraneoplastic hypoglycemia being reported as one of the associated clinical features of such slow-growing tumors. As hypoglycemia can be an emergent clinical situation, we describe a patient with a giant abdominal tumor who presented with a symptomatic hypoglycemia, which might be regarded as a diagnostic clue. PMID- 15290508 TI - Lower cervical dermal sinus tract and associated intraspinal abscess causing meningitis in a child. AB - Congenital dermal sinus tracts are frequently a cause of recurrent meningitis. We present a case of a rare lower cervical dermal sinus and an associated intraspinal abscess causing meningitis. PMID- 15290509 TI - Ruptured splenic abscess: a cause of pneumoperitoneum in a patient with AIDS. AB - We encountered a case of pneumoperitoneum caused by a gas-forming splenic abscess in a patient with acquired immunodeficiency syndrome (AIDS). Plain abdominal films and computed tomography demonstrated a large amount of free air. Pneumoperitoneum was eventually shown to represent gas liberated from fermentation by gas-forming organisms within the splenic abscess. Gas-containing necrotic tissue from the ruptured spleen mimicked the spillage of feces from colon perforation. The authors emphasize that a ruptured abscess should be included in the differential diagnosis of acute abdomen in an immunocompromised patient. PMID- 15290510 TI - The angled buckle fracture in pediatrics: a frequently missed fracture. AB - Buckle (torus) fractures in childhood are very common, and most assume a typical configuration wherein the trabeculae across the fracture line are compressed and the corresponding cortex bulges outward (unilateral or bilateral). In other cases the fracture merely shows cortical angulation along one side of the bone and classical buckling is not present. These latter fractures frequently are subtle and easily overlooked. However, if one knows what they look like and where they are likely to occur, one is able to detect them with greater frequency and confidence. The purpose of this communication is to bring attention to this fracture, describe the mechanism by which it occurs, and indicate its most common sites. PMID- 15290512 TI - Occurrence of comminution (type IIA) in type II odontoid process fractures: a multi-slice CT study. AB - The classification of odontoid process fractures introduced by Anderson and D'Alonzo in 1974 has gained wide acceptance among researchers and clinicians. In type II fractures the injury is located in the base of the odontoid process. In 1988 Hadley et al. introduced a comminuted subtype of this fracture, type IIA. Since then several authors have recommended a more aggressive surgical approach in treatment of this subtype. However, only a few cases of comminuted type II fractures have been reported. The objective of this study is to assess the occurrence of comminution in type II fractures. Twenty-six type II fractures imaged by multislice CT were reviewed. Subtle comminution of the fracture was observed in 12 of the cases. We conclude that, when imaged by multislice CT, subtle comminution in type II odontoid fractures is seen more commonly than previously reported in the literature. PMID- 15290511 TI - Anterior rim tibial plateau fractures and posterolateral corner knee injury. AB - The aim of this study was to review MRI findings of clinically suspected posterolateral corner knee injuries and their associated internal derangements. Sixteen knees in 15 patients who had evidence of a posterolateral corner knee injury on the physical exam underwent MRI to evaluate the posterolateral corner of the knee and to look for associated injuries. Two musculoskeletal radiologists reviewed the scans. Surgery was performed on 10 of the knees. Tibial plateau fractures were present in 6 knees; 5 of the fractures were anteromedial rim tibial plateau fractures. The popliteus muscle was injured in 13 knees and the biceps femoris in 6 knees. The lateral collateral ligament was ruptured in 12 knees. The posterior cruciate ligament was completely ruptured in 7 knees and avulsed from its tibial attachment in 1 knee. Eleven knees had a complete anterior cruciate ligament rupture. The anterior cruciate ligament was edematous without complete disruption of all fibers in 3 knees. There was excellent correlation between the MRI results and operative results in regard to the presence of a posterolateral corner injury of the knee (9 of the 10 knees had a posterolateral corner injury). In our study MRI readily detected posterolateral corner injuries. Posterolateral corner injuries of the knee are frequently associated with a variety of significant injuries, including cruciate ligament tears, meniscus tears, and fractures. Fractures of the peripheral anteromedial tibial plateau are not common; however, given their relatively common occurrence in this study, they may be an indicator of a posterolateral corner injury to the knee. PMID- 15290513 TI - Evaluation of personal digital assistants as an interpretation medium for computed tomography of patients with intracranial injury. AB - The objective of the study was to assess the feasibility of using a personal digital assistant (PDA) as a medium for the interpretation of cranial CT scans of trauma patients. Twenty-one noncontrast cranial CT scans were transferred in their entirety to a PDA from the picture archiving and communications system (PACS) utilizing General Electric (GE) PathSpeed PACS Web Server interface and synchronization. All CT scans had been interpreted by board-certified radiologists prior to the study. Seven of the scans demonstrated subarachnoid hemorrhage, seven demonstrated subdural hematomas, and the remaining scans were normal. After transfer to the PDA, all images were separately reviewed in a blinded manner by a radiologist and a neurosurgeon. Images were graded for their quality and diagnostic utility in the evaluation of intracranial hemorrhage. Image quality was categorized as excellent, very good, acceptable for diagnosis, or not acceptable for diagnosis. Based on the radiologic diagnosis, recommendation for surgical management was made by the reviewing neurosurgeon. The accuracy rate for both the radiologist and the neurosurgeon in the detection of intracranial hemorrhage was 95%. There was one false negative which was attributed to error in judgment rather than poor image quality. This diagnostic error did not affect patient management. The sensitivity and specificity for detection of intracranial hemorrhage were 93% and 100%, respectively. Image quality was judged to be excellent in 90% of the cases and very good in the remaining 10%. Our results suggest that the PDA is a robust medium for interpretation of CT scans in patients with suspected hemorrhage following intracranial injury. In this setting, the PDA should be considered for teleradiology purposes. PMID- 15290514 TI - Characteristic pancreatic injuries secondary to child abuse. AB - A review of the most common visceral injuries which arise as a result of child abuse is presented. Duodenal and pancreatic injuries are the most characteristic injuries secondary to abuse. The clinical presentation may not be suggestive of the nature of the patient's injury, and these descriptions may help the radiologist to discern their true etiology. PMID- 15290515 TI - Internal hernia through a defect in the broad ligament: a rare cause of intestinal obstruction. AB - Internal hernias through the broad ligament are an extremely rare cause of intestinal obstruction. Clinical symptoms and imaging are often nonspecific, making preoperative diagnosis difficult. The authors report a case in which multidetector computed tomography proved useful in the preoperative diagnosis. PMID- 15290516 TI - The jet ski open-book pelvic fracture: diagnosis with multidetector CT. AB - A 10-year-old girl sustained a traumatic open-book pelvic fracture from a straddle injury in a jet ski accident. Plain films and computed tomography both demonstrated diastasis of the symphysis pubis and bilateral widening of the sacroiliac joints. The open-book fracture resulted from the patient's striking the steering column of the watercraft during a deceleration accident. The unusual cause of this injury is of clinical interest because with increasing popularity of personal watercraft and changes in the design of these vehicles, the incidence and prevalence of this type of injury may increase in the future. PMID- 15290517 TI - Imaging of bilateral ureteropelvic junction laceration from blunt trauma. AB - Bilateral ureteropelvic junction (UPJ) injury from blunt abdominal trauma is rare, with only seven previously reported cases, all of which were complete avulsions. Early and delayed computed tomography (CT) for visualization of the nephrographic and excretory renal phases, respectively, is essential to distinguish parenchymal from collecting system injury. Once UPJ injury is detected by CT, differentiation between laceration and avulsion is mandatory since laceration is treated nonoperatively, whereas avulsion requires surgical repair. In addition to CT, intravenous pyelography (IVP) or retrograde pyelography may be required for full characterization of the injury. Retrograde pyelography may permit better opacification of the ureters than IVP, enabling the urologist to determine whether stent placement is necessary. We report the first case of bilateral UPJ laceration secondary to blunt abdominal trauma and the imaging studies necessary to make the diagnosis. PMID- 15290518 TI - Stress reaction in the carpal bones caused by breakdancing. AB - Stress reactions in the bones of the lower extremities are a common finding on magnetic resonance (MR) imaging. The primary finding in the bone marrow is nonspecific edema without any visible fracture line that may even mimic tumor or infection. Continuing stress may eventually lead to a stress fracture. We present the case of a stress reaction related to breakdancing in a less typical localization, in the triquetral bone in the wrist. PMID- 15290519 TI - Bilateral asymmetric hip dislocation: case report and literature review. AB - Simultaneous anterior and posterior hip dislocation is an unusual injury. A unique case is presented, consisting of bilateral asymmetric hip dislocation with associated femoral head, femoral shaft, and acetabular fractures resulting from a motorcycle collision. The mechanisms of injury, management, role of imaging, and complications of this injury complex are discussed, with a review of the relevant literature. PMID- 15290521 TI - The emergency suite and the end of geography. PMID- 15290520 TI - Self-inflicted transorbital and intracranial injury from eyeglasses. AB - Orbital injuries are commonly seen in the emergency department, and if they are high-energy they can lead to concomitant intracranial injuries. Plain films, CT, MRI, and ultrasound are used in various combinations to evaluate the extent of these injuries. We describe a unique case of self-inflicted transorbital penetrating intracranial injury from the temporal wire rim of a pair of eyeglasses. Imaging well demonstrates the full course of the wire rim in situ, and pathoanatomic correlates are highlighted. PMID- 15290522 TI - Rings and things on upper extremity radiographs of emergency patients. AB - Rings, intravenous lines, and other objects on the injured upper extremities of trauma patients are frequently overlooked by radiology and emergency department (ED) personnel. This can impair proper radiologic evaluation of the injured extremity as well as negatively affect the quality of the patient's treatment. A 1-week sample of radiographs of injured upper extremities from the ED of University Medical Center (UMC), Tucson, Arizona, showed that 20% of the studies (19 of 95) contained at least one object on the injured upper extremity, but only one radiology report (1.1%) mentioned such an object. A review of 2489 upper extremity ED radiology reports from January to June 2002 showed only 47 reports (1.9%) that mentioned the presence of an overlying object. It is important to educate radiology department and ED personnel to remove upper extremity jewelry and place necessary medical devices on noninjured extremities. PMID- 15290523 TI - Radiological findings in Boerhaave's syndrome. AB - The aim was to define the diagnostic value of chest radiography, esophagography, and computed tomography (CT) in patients with Boerhaave's syndrome. CT findings in 14 patients (11 male, 3 female; mean age: 60 years; median age: 66 years; age range: 36-78 years) with spontaneous esophageal perforation were retrospectively reviewed and compared to those of esophagography ( n=11) and chest radiography ( n=14). In six patients unenhanced CT scans were available; in ten patients (2/10 patients had an unenhanced CT before) a contrast-enhanced CT was performed as an emergency investigation. In 5/14 patients additional oral contrast medium was given. Chest radiography revealed pleural effusions in 9/14 and infiltrates in 9/14 patients. Pneumothorax and pneumopericardium were observed in two patients each. Pneumomediastinum was seen in three patients. Two chest radiographs were normal. Esophagography performed in 11 patients demonstrated contrast medium extravasation at a supradiaphragmatic level in seven patients, indicating esophageal perforation with esophagopleural fistula, and a submucosal contrast medium collection in four cases. Unenhanced CT scans revealed four intramural hematomas with typical localization. Unenhanced and contrast-enhanced CT demonstrated periesophageal air collections indicating esophageal perforation in all patients. Pleural effusions were seen in 11/14 and infiltrates in 8/14 patients. Contrast medium extravasation was observed in 5/14 patients, whereas a thickening of the esophageal wall was depicted in 11/14 patients. Periaortic air tracks ( n=7), pneumothorax ( n=3), pneumomediastinum ( n=6), and mediastinal fluid collections ( n=7) were depicted in CT. By demonstrating periesophageal air tracks suggestive of esophageal perforation, CT reveals the decisive criteria for diagnosing Boerhaave's syndrome and should therefore be performed in the diagnostic work-up of patients in whom this syndrome is part of the differential diagnosis. PMID- 15290524 TI - The use of MRCP in the detection of pancreatic injuries after blunt trauma. AB - From January 2000 to November 2001, five consecutive, hemodynamically stable trauma patients (age range 8-69 years, mean age 34 years) with parenchymal injuries were evaluated by magnetic resonance imaging (MRI) with magnetic resonance cholangiopancreatography (MRCP). One patient also underwent a MRCP facilitated secretin test. MRI depicted pancreatic laceration in two patients, ductal disruption and a post-traumatic intraparenchymal pseudocyst in one, migrating pancreatic fluid collection in the mediastinal space with disruption in another, and main pancreatic duct rupture and dilatation in the patient evaluated with MRCP following secretin administration. MRI with MRCP is an effective noninvasive test for detecting and managing pancreatic injuries after blunt trauma. Secretin administration improves ductal visualization, particularly of nondilated ducts. Finally, MRI was useful in the follow-up studies of parenchymal damage and minor ductal injuries, providing high-quality images of the pancreatic duct and biliary tract. PMID- 15290525 TI - Lateral femoral condylar shearing fractures. AB - Lateral femoral condylar shearing fractures occur as the aftermath of acute patellar dislocation. The fracture fragment may be larger than originally appreciated on plain films. Diagnosis depends on identifying the fracture fragment somewhere within the knee joint, and the "donor site" on the lateral femoral condyle. Therefore, MRI or arthroscopy should be performed to identify the true size of the fracture fragment. The larger of these fragments may require surgical intervention with fixing of the fracture fragment onto the lateral condyle with screws. We present the radiographic and clinical findings in seven patients. PMID- 15290526 TI - Report on a new type of trauma full-body digital X-ray machine. AB - The purpose of this study was to evaluate the diagnostic equivalence, radiation dose, clinical usefulness and radiographic aspects of a low-dose, full-body digital X-ray machine in a busy trauma unit. A digital trauma X-ray machine known as "LODOX" was compared with conventional radiography between June 1999 and November 2001 in the Groote Schuur Hospital Trauma Unit, Cape Town. Digital images of a variety of body regions commonly imaged in trauma were compared for diagnostic image quality in a number of categories with equivalent conventional radiographs. A seven-point equivalence scoring system ranging from much inferior (-3) through equivalent (0) to much superior (+3) was used in each category. Radiation dose was recorded and compared with that in conventional measurements. Turnaround times of patients undergoing digital and conventional X-rays were evaluated. Clinical and radiographic issues were assessed by staff feedback. The digital images when compared with conventional film had an overall mean equivalence score of -0.429, with a standard deviation (SD) of 0.77. The best digital performance was in the mediastinum (mean 0.346, SD 0.49) and the weakest was for bony detail (mean -0.654, SD 0.81). Relative digital radiation dose compared to conventional varied from 72% (chest) to 2% (pelvis), with a simple average of 6%. Radiographic points included full-body imaging capability and differing positioning, penetration, workflow and practicality considerations. The digital images required overall patient times of 5-6 min, compared with 8-48 min for conventional X-rays. New installations are under way, and computed tomography and angiography applications are being explored. FDA approval is awaited. Projected cost is similar to that of flat-panel digital units. This digital unit was felt to be diagnostically substantially equivalent to conventional radiographs, with low-dose full-body imaging, improved workflow, digital technology and long-term cost benefits as potentially favourable contributions to trauma imaging. PMID- 15290528 TI - Cervical spine trauma: evaluation by multidetector CT and three-dimensional volume rendering. AB - Multidetector-row computed tomography (CT) offers important advantages over conventional imaging modalities in the evaluation of the post-trauma cervical spine. It allows for faster scanning times, critical for triaging post-trauma patients as well as for eliminating motion artifacts, and allows for thinner collimation and the ability to achieve an isotropic data set which can be reformatted in any plane without loss of spatial resolution. In addition, three dimensional volume-rendered reconstructions of images obtained using multidetector scanners can provide additional information in defining extent of injury, allowing neurosurgeons to see the fractures in any plane, simulating intraoperative views. 3D multidetector-row CT represents an advance in CT technology and can help ensure rapid, accurate evaluation of cervical spine injuries. PMID- 15290527 TI - Adrenal injuries: spectrum of CT findings. AB - Injury to the adrenal gland is often incidentally diagnosed with Computed Tomography (CT) following blunt abdominal trauma. In a high percentage of cases, it is accompanied by other intra-abdominal, retroperitoneal or intrathoracic injuries. Although not usually clinically significant, adrenal traumatic lesions can be a source of infection, as well as a cause of acute adrenal insufficiency. We discuss and illustrate typical and less common CT findings of adrenal injuries. PMID- 15290529 TI - Arteriovenous fistula following pacemaker lead removal: CT diagnosis. AB - Arteriovenous fistula formation is a very rare complication of pacemaker lead extraction. Rapid diagnosis is essential due to the life-threatening nature of this complication. CT angiography provides a noninvasive and quick method for assessment. PMID- 15290530 TI - Pulmonary embolism-the initial manifestation of renal cell carcinoma in a young woman. AB - We report a case of massive pulmonary embolus demonstrated on CT in a young woman presenting with dyspnea, with no known risk factors for embolism. Abdominal CT on further investigation showed a renal tumor invading the left renal vein and the inferior vena cava as the cause of the pulmonary embolus. In a patient presenting with pulmonary artery embolism without venous thrombosis, the differential diagnosis should include an occult tumor as the cause of the embolus. PMID- 15290531 TI - Finding of CT and MR evaluation of gallbladder hemobilia. AB - We report a case of gallbladder hemobilia secondary to blunt abdominal trauma. The diagnosis was initially based on findings on CT scan of the abdomen and correlated with MRI findings. The patient was found to have a distended gallbladder containing high-density material consistent with hemobilia. MRI revealed a gallbladder containing material of mixed signal intensity consistent with blood products. Gallbladder hemobilia is a very rare condition which is potentially fatal. PMID- 15290532 TI - Anaphylaxis due to a rupture of hydatid cyst: imaging findings of a 10-year-old boy. AB - Anaphylactic shock as a result of trauma is very rare. We report the clinical and radiologic findings of a 10-year-old boy who developed systemic anaphylaxis due to traumatic rupture of hepatic hydatid cyst into a pericystic blood vessel. In regions where hydatid disease is endemic, rupture of a hydatid cyst might be taken into consideration in patients who have post-traumatic anaphylactic shock. Likewise, this pathology should be kept in mind when evaluating an immigrant from an endemic area in nonendemic regions. PMID- 15290533 TI - Spontaneous rupture of the spleen detected on CT as the initial manifestation of infectious mononucleosis. AB - Spontaneous splenic rupture after infectious mononucleosis (IM) is a rare, potentially fatal complication of IM, occurring in 0.1-0.5% of patients with proven IM. It usually occurs several weeks after the onset of symptoms, but may, rarely, be the initial manifestation of the disease. The patient is usually examined as an emergency due to severe abdominal pain and a falling hematocrit. The radiologist should be aware of the pathologic conditions involving the spleen which may lead to its spontaneous rupture. PMID- 15290534 TI - Adult intussuception as a cause of abdominal symptoms: a case report and review of literature. AB - Intussusceptions are frequently encountered in children. In adults, they are uncommon and have a different etiology. Our case is one such example of a rare, pathologically proven, recto-rectal intussusception due to an adenocarcinoma with characteristic CT findings. PMID- 15290535 TI - Uterine rupture with peritonitis in a nongravid uterus eight weeks after cesarean section. AB - The CT findings of uterine rupture are critical to recognize so that early surgical intervention can improve survival and potentially avoid hysterectomy. We report a case of uterine rupture in a nongravid uterus. PMID- 15290536 TI - Postpartum ovarian vein thrombosis with simultaneous pyelocaliectasis: diagnosis and follow-up by MR imaging. Case report and literature review. AB - We describe the critical role of MR imaging in a case of postpartum ovarian vein thrombosis (OVT) with concomitant pyelocaliceal ectasia. MR imaging confirmed the diagnosis suspected on the basis of ultrasonography and computed tomography by demonstration of a subacute clot with high signal intensity within the right ovarian vein and its complete resolution after anticoagulant therapy. MR imaging is a useful noninvasive, accurate tool for the diagnosis and follow-up of this potentially life-threatening condition, providing information helpful for choosing a prompt medical treatment rather than a surgical therapy. To our knowledge, no previous case of OVT causing pyelocaliceal ectasia documented by MR imaging has been reported. However, even though our case is suggestive, a cause effect relationship between OVT and hydronephrosis could not be demonstrated with certainty. PMID- 15290537 TI - Delayed rupture of abdominal aortic false aneurysm following blunt trauma. AB - Blunt injury of the abdominal aorta resulting in pseudoaneurysm formation is very rare. Such a pseudoaneurysm may rupture at any time, usually with fatal outcome. We report the case of a 32-year-old man with a clinically unsuspected ruptured abdominal aorta pseudoaneurysm, which had probably formed 3 years earlier, and emphasize the CT features. PMID- 15290538 TI - Bamboo splinter: ultrasound localization. AB - Ultrasound was used to identify and localize a 2-cm hyperechoic bamboo splinter in the wrist of a patient with persistent symptoms, 1 month after the initial injury, following an initial negative workup by clinical examinations and X-ray. PMID- 15290539 TI - Scaphoid (navicular) fractures of the wrist in children: attention to the impacted buckle fracture. AB - We retrospectively analyzed the radiographic findings in 76 patients with proven scaphoid (navicular) fractures. The purpose of the study was to determine the incidence of the impacted buckle fracture of the scaphoid bone in children. Fourteen of the 76 patients (17%) demonstrated such a fracture, and all of the fractures occurred in the middle or distal third of the scaphoid bone. The mean age of this group of patients was 13.7 years, as compared to the mean age of 15.6 years in the patients with overt transverse fractures. One out of 14 of the patients with a buckle fracture was female. None developed complicating aseptic necrosis. The fractures were most clearly visualized on AP and oblique views of the wrist, and all cases demonstrated edematous thickening of the soft tissues and/or navicular fat pad obliteration adjacent to the scaphoid bone. The use of comparative views was extremely helpful in the detection of this fracture. PMID- 15290540 TI - Helical computed tomography characteristics of splenic and hepatic trauma in children subjected to nonoperative treatment. AB - The purpose of this study was to present the radiological characteristics of abdominal computed tomography (CT) in the follow-up of splenic and hepatic injury in children. Children ( n=24) less than 13 years old who had suffered blunt abdominal trauma and were diagnosed with splenic and hepatic injury by CT scan prospectively were enlisted in the study. The CT was performed immediately after the injury was suspected, and 7 and 60 days after the trauma. The clinical course of the patients was observed (red blood transfusion requirement, associated abdominal injuries, and hospital stay). The splenic and hepatic injuries varied from grade II to grade IV of the American Association for the Surgery of Trauma. The CT showed a reduction in the volume of the injury 60 days after the trauma. In this article the radiological findings will be shown and correlated with the clinical course of the patients. This study shows that CT is advantageous for detecting and grading splenic and hepatic injuries. These injuries can be managed nonoperatively in hospitals where CT is available for the evaluation of pediatric patients. PMID- 15290541 TI - Simultaneous ipsilateral elbow and forearm fractures in children: a retrospective review. AB - The aim is to evaluate the incidence of simultaneous ipsilateral forearm fractures in pediatric patients who present with elbow fractures and to attempt to identify patients who might be at greater risk of this type of injury. All pediatric patients with elbow radiographs during a 3-month period were retrospectively reviewed. One hundred sixty patients were identified. Sixty-one of the 160 were diagnosed with elbow fractures. Of these, 7 had a simultaneous ipsilateral forearm fracture (12.3%). Of the 7 patients with simultaneous fractures, all had supracondylar fractures; 4 were displaced elbow fractures and 3 were nondisplaced. Although our initial experience is limited, there appears to be a fairly high incidence of ipsilateral forearm fractures in pediatric patients with elbow fractures. All of these cases involved a supracondylar fracture, and over 50% showed displaced fractures. We conclude that in preadolescent patients with elbow fractures, simultaneous forearm fracture should be considered, and clinical and or radiographic evaluation of the forearm may be warranted. PMID- 15290542 TI - Bacterial aortitis resulting in rapid development of infective "mycotic" aneurysm. AB - Aortitis and aneurysms with an infective etiology, though uncommon, must be recognized at imaging so that early medical and surgical intervention can improve survival. We report a case in which serial CT scans illustrate the short-interval development of an infective "mycotic" aneurysm resulting from bacterial aortitis. PMID- 15290543 TI - False-negative contrast MRA in the setting of carotid artery dissection. AB - A case of false-negative contrast magnetic resonance angiography (MRA) in the setting of significant carotid stenosis caused by a carotid arterial intimal dissection is described. If arterial dissection is suspected but contrast MRA is normal, additional evaluation may be warranted. In this case the abnormality was detectable on 2D time-of-flight MRA and contrast digital subtraction arteriography. Since MRA has become a commonly used initial screening test for carotid stenosis, it is important to understand the possibility of false-negative results. PMID- 15290544 TI - Post-traumatic posterior sternoclavicular dislocation: case report and review of the literature. AB - Injuries to the sternoclavicular joint are an uncommon problem, but can be life threatening if not diagnosed and treated properly. Both anterior and posterior sternoclavicular dislocations or subluxations can occur, the latter being a rare injury. In this case report we report the imaging findings of both conventional radiography and multidetector CT on a 16-year-old man who developed a posterior sternoclavicular dislocation while participating in a football match. The current literature regarding this subject is also reviewed. PMID- 15290545 TI - Effort-induced thrombosis: diagnosis with three-dimensional MR venography. AB - A case of Paget-von Schrotter syndrome diagnosed with gadolinium-enhanced magnetic resonance (MR) venography is described. MR enabled a comprehensive evaluation with identification of the thrombus within the left subclavian vein and a hypertrophied anterior scalene muscle as the probable cause of the patient's condition. We describe the MR venography technique used for evaluation of the subclavian vein and the imaging findings in this entity. PMID- 15290546 TI - Long-standing unilateral jumped facets at C3-4 with no apparent history of antecedent trauma. AB - We report two cases of unilateral jumped facets at C3-4 in reliable historians with no apparent history of neck trauma. Lack of associated morphological abnormalities of the associated disc, adjacent vertebral bodies, and contralateral facet essentially exclude a developmental etiology. Based on the location of the injury and the presence of chronic, osteoarthritic changes we postulate that the jumped facets were due to remote childhood trauma that was forgotten. PMID- 15290547 TI - CT of small bowel obstruction secondary to a cocaine-filled condom. AB - Drug smuggling is prevalent in our society. It is now frequently seen in the emergency room as an acute life-threatening emergency. The following case describes one such patient with an emphasis on the CT findings in these cases. PMID- 15290548 TI - The use of flexion and extension MR in the evaluation of cervical spine trauma: initial experience in 100 trauma patients compared with 100 normal subjects. AB - The purpose of this study was to determine the value of flexion and extension MR in traumatized cervical spines following rear low-impact acceleration deceleration injury. The cervical spines of 100 consecutive uninjured normal asymptomatic adults and 100 adult accident victims following rear low-impact motor vehicle accidents were evaluated using rapid T2-weighted MRI. Subjects were matched for age but not gender. The age range was 18 to 53 years, with a mean of 35 years. Injured subjects were evaluated during the subacute period, at 12 to 14 weeks after injury, following clinically resolved muscle spasm. Imaging findings were compared between normal and injured subjects. The normal subjects showed a stepwise segmental motion pattern that started at C1-C2 and transmitted to the lower cervical segments. Loss of normal cervical lordosis (hypolordosis) was observed in 4% (4 of 100) patients. Normal range of motion (rounded to the nearest 5 masculine ) was quantified as 50 masculine flexion (range, 45-65 masculine; standard deviation, 6.5 masculine ) and 60 masculine extension (range, 50-70 masculine; standard deviation, 6.5 masculine ). Asymptomatic disk herniations were observed in 2% (2 of 100) patients. In the subacute post traumatic subjects, there was a loss of the normal segmental motion pattern, with hypolordosis in 98% (98 of 100) patients. Range of motion (rounded to the nearest 5 masculine ) was restricted, quantified as 25 masculine flexion (range, 5-40 masculine; standard deviation, 15 masculine ) and 35 masculine extension (range, 20-50 masculine; standard deviation, 10 masculine ). Disk herniations were observed in 28% (28 of 100) patients. Biomechanical changes in the herniated disk were noted, with mildly increased spinal stenosis following flexion. The authors conclude that flexion and extension MR can be a valuable adjunct examination in the evaluation of patients in the clinical setting of subacute cervical spine trauma. PMID- 15290549 TI - Is there need for thoracic spine radiographs following a negative chest CT in trauma patients? AB - The purpose of this study was to assess the need for conventional radiographs of the thoracic spine for routine clearance of trauma patients in whom chest CT has revealed no spinal trauma. The study was in the form of a retrospective review of trauma patients over the previous five years who underwent conventional radiographs of the thoracic spine following a chest CT that revealed no spinal trauma. Two hundred thirty-five trauma patients were found to have undergone conventional thoracic spine series following a chest CT that showed no spinal trauma. In 234 of the cases, the thoracic spine series was also negative. In one case, the thoracic spine series revealed mild anterior compression of the T7 vertebral body. This injury was stable and required no specific intervention. CT of the chest is an adequate evaluation of the thoracic spine in trauma patients who require routine thoracic spine clearance, making subsequent conventional radiographs of the thoracic spine unnecessary. PMID- 15290550 TI - Chest CT scanning for clinical suspected thoracic aortic dissection: beware the alternate diagnosis. AB - The aim of the study was retrospectively to evaluate the spectrum of chest diseases in patients presenting with clinical suspicion of thoracic aortic dissection in the emergency department. We performed a retrospective medical records review of 86 men and 44 women (ages ranging between 23 and 106 years) with clinically suspected aortic dissection, for CT scan findings and final clinical diagnoses dating between January 1996 and September 2001. All images were obtained by using a standard protocol for aortic dissection. We found aortic dissection in 32 patients (24.6%), 22 of which were Stanford classification type A and 10 Stanford type B. In 70 patients (53.9%), chest pain could not be explained by the CT scan findings. However, in 28 patients (21.5%), CT scanning did reveal an alternate diagnosis that, along with the clinical impression, probably explained the patients' presenting symptoms, including: hiatal hernia (7), pneumonia (5), intrathoracic mass (4), pericardial effusion/hemopericardium (3), esophageal mass/rupture (2), aortic aneurysm without dissection (2), pulmonary embolism (2), pleural effusion (1), aortic rupture (1), and pancreatitis (1). In cases where there is clinical suspicion of aortic dissection, CT scan findings of an alternate diagnosis for the presenting symptoms are only slightly less common than the finding of aortic dissection itself. Although the spectrum of findings will vary depending upon your patient population, beware the alternate diagnosis. PMID- 15290551 TI - Acute epiploic appendagitis: CT findings in 33 cases. AB - Acute epiploic appendagitis (AEA) is a benign self-limiting process presenting with acute abdominal pain often misdiagnosed clinically as either diverticulitis or appendicitis, but which has a pathognomonic CT appearance. The CT findings in 33 adult patients diagnosed by CT over a 33-month period as having AEA were retrospectively reviewed. The study group included 24 men and 9 women, with a mean age of 44.6 years. The mean age of the male patients was lower than that of the female patients, 40.9 vs 54.7 years. All patients presented with acute abdominal pain, mainly in the left ( n=21) and right ( n=9) lower quadrants, with localized tenderness in all patients and peritoneal irritation in 15 of them. Low grade fever was found in 8 patients and mild leukocytosis in 16. Characteristic CT findings of an oval fatty mass with central streaky densities and surrounded by mesenteric stranding adjacent to the serosal surface of the colon were seen in all cases. Additional findings included mural thickening of the juxtaposed colon in 16 patients and peritoneal fluid in 7. One patient underwent surgery on the basis of an erroneous diagnosis of acute appendicitis. As CT is often used nowadays to evaluate various acute abdominal complaints, it may be the first imaging modality by which AEA is diagnosed. AEA should be included in the differential diagnosis in young male patients with localized left lower abdominal pain and tenderness. PMID- 15290552 TI - Is the screening portable pelvis film clinically useful in multiple trauma patients who will be examined by abdominopelvic CT? Experience with 397 patients. AB - The multiple trauma patient is usually initially imaged with a portable "trauma series" consisting of a lateral cervical spine film, a portable chest film, and a portable pelvis film (PPF). An investigation was performed to determine whether the screening PPF could be eliminated for multiple trauma patients being examined by abdominopelvic CT scan (APCT). A retrospective investigation analyzed all patients evaluated in our level I trauma center from 1 January to 31 December 2000 who were examined with a "trauma series" followed by an APCT scan within 8 h. The numbers and types of fractures diagnosed by PPF and by APCT were compared and correlated with clinical follow-up. Of 397 patients imaged by both PPF and APCT, 43 patients were diagnosed with 109 individual fractures by CT scan. The PPF did not detect 51 of the 109 individual fractures (47%) and failed to diagnose 9 of the 43 patients (21%) with a pelvic fracture. The PPF most often failed to detect sacral and iliac fractures. The four cases in which the PPF reported a fracture not listed in the APCT report were due to reporting errors or film artifacts. No soft tissue injuries were seen by PPF that were not also seen by APCT. We conclude that the screening PPF appears to be an unnecessary exam in multiple trauma patients about to be imaged by APCT scan. PMID- 15290553 TI - Isolated injury of the cuboid bone. AB - The purpose of this study was to describe isolated injury of the cuboid bone as a potentially radiographically occult cause of foot pain. The imaging studies of 17 patients, 13 women and 4 men aged 17-79 years (average 45 years), who presented with pain over the lateral aspect of the midfoot were retrospectively reviewed. Frontal, lateral, and inversion-oblique radiographs were available for all patients. In addition, MR imaging was performed in eight patients, CT in two, conventional tomography in two, and bone scan in one. Conventional radiographs revealed cuboid fracture in seven patients. Of the remaining ten, eight underwent MR imaging which demonstrated four fractures, three bone bruises, and one stress reaction, and two had tomography, CT, and/or bone scan, all of which documented an isolated cuboid fracture. Isolated fracture of the cuboid may be radiographically occult. Other imaging modalities, particularly MR imaging, can document this injury as the source of pain. PMID- 15290554 TI - Periosteal reaction with normal-appearing underlying bone: a child abuse mimicker. AB - Any irritation or disruption to the underlying bone will cause a periosteal reaction and result in new periosteal bone deposition. Periosteal bone formation may be due to either physiologic or pathologic causes. Pathologic bone formation generally results from an adjacent inflammatory process or a hypoxic or toxic stimulus. Common causes of pathologic periosteal reaction in children include trauma to the underlying bone. However, other causes such as hypervitaminosis A, prostaglandin therapy, cortical hyperostosis (Caffey's disease), hypertrophic osteoarthropathy (primary and secondary), osteomyelitis, leukemia, trauma, and syphilis must also be considered. The last four are usually associated with some degree of bone destruction, while in the first four diseases the underlying bone is left radiologically intact. This paper will concentrate on those diseases that appear to leave the underlying bone intact. The clinical and radiological features that help to differentiate some of these entities are presented. PMID- 15290555 TI - Ultrasound in acute trauma of the ankle and hindfoot. AB - Ultrasound is a rapid, widely available and inexpensive imaging modality for the evaluation of the ankle and hindfoot. Ultrasonography can be performed in acute, semiacute and chronic conditions. Ankle injuries can be evaluated with ultrasound combined with X-rays. In the emergency room, acute trauma of ankle and hindfoot is an important indication for ultrasound. PMID- 15290556 TI - Dobutamine stress magnetic resonance imaging. AB - Cardiovascular magnetic resonance imaging (CMR) provides superior imaging of cardiac structure and function that makes it a useful tool in the diagnosis of cardiovascular disease. The heart's response to stress has long been used as a diagnostic aid for identifying a cardiovascular etiology for patient symptoms or for cardiovascular risk stratification. Historically, this has been done with stress nuclear scintigraphy or echocardiography techniques. A distinct advantage of CMR over these modalities is greater image quality resulting in better test characteristics; this review summarizes the techniques and principles involved in dobutamine stress MRI. PMID- 15290557 TI - "Cecal gangrene": a rare cause of right-sided inferior abdominal quadrant pain, fever, and leukocytosis. AB - We report on a 58-year-old man with known diabetes, congestive heart failure, and need for chronic hemodialysis presenting with right lower abdominal quadrant pain, fever, and leukocytosis. Although initial clinical findings were highly suggestive of acute appendicitis, CT revealed marked circumferential wall thickening of the cecum, which was interpreted as cecal infarction by the radiologist. Intraoperatively, cecal necrosis was confirmed, but the ileocecal valve and, especially, the appendix showed no ischemia. No vascular occlusions were found. Histopathologic analysis of the resected cecum demonstrated isolated transmural cecal necrosis with marked infiltration of the cecal wall by numerous bacteria and neutrophils. We present the CT features and histopathologic findings of isolated cecal gangrene, review the pathogenesis of occlusive and nonocclusive cecal ischemia or infarction, and discuss the role of bacterial superinfection as a potential cofactor in the pathogenesis of isolated cecal necrosis which should be included in the differential diagnosis of right-sided inferior abdominal quadrant pain. PMID- 15290558 TI - Postcatheterization arteriovenous fistula: CT, ultrasound, and arteriographic findings. AB - The most common etiology of arteriovenous fistulae (AVF) in the lower extremity is iatrogenic, usually from diagnostic or therapeutic angiographic procedures. The finding of a palpable groin thrill, early venous opacification on contrast enhanced abdominal-pelvic CT, and typical findings on duplex ultrasonography establish the diagnosis. Anatomic confirmation is then made by arteriography. A case of an incidentally discovered AVF in patient presenting to the Emergency Department is presented. Radiographic findings concerning and subsequent management of this patient are then discussed. PMID- 15290559 TI - Penetrating axillary trauma in a survivor of the World Trade Center disaster. AB - Penetrating axillary trauma is classically associated with gunshot wounds or stab wounds, and it is a cause for concern because of the high risk of vascular injury. We report a case of penetrating axillary trauma to a victim of the World Trade Center disaster. In this case report, we discuss the CT and angiographic findings of both blunt and penetrating axillary injury, and present a diagnostic algorithm. PMID- 15290560 TI - Can we rely on mediastinal widening on chest radiography to identify subjects with aortic injury? AB - Widening of the mediastinum on chest radiography is widely promoted as a useful criterion for detecting aortic injury. We sought to determine the reliability, sensitivity, and specificity of this sign. The initial chest radiographs from 30 subjects with aortic injury and 47 controls were independently reviewed by six radiologists, who were blinded to diagnosis. The radiologists were asked to decide whether the mediastinum was normal or not normal, as well as whether the mediastinum was widened. Agreement, sensitivity, and specificity were assessed. Agreement for overall assessment of the mediastinum was substantial (kappa = 0.64). Individual radiologists had sensitivity varying from 0.77 to 0.97 and specificity varying from 0.62 to 0.89. For "widening" of the mediastinum, agreement was moderate (kappa = 0.49). "Widening" was less sensitive than the radiologists' overall impression (P = 0.01), varying from 0.50 to 0.83, but no difference was detected in specificity (P = 0.36), varying from 0.81 to 0.94. Mediastinal width has unacceptable sensitivity for predicting aortic injury, with substantial inter-reader variability. Medical education has ingrained the widely promoted concept of mediastinum widening, which may be misleading. PMID- 15290561 TI - Can CT predict the source of arterial hemorrhage in patients with pelvic fractures? AB - The objective of the study was to evaluate the ability of hemorrhage site and location as demonstrated on pelvic CT to predict the source of arterial hemorrhage in patients with traumatic pelvic fractures. CT scans of 104 consecutive patients who had sustained traumatic pelvic fracture and undergone emergent pelvic angiography were digitized, and fracture-related hemorrhage area and volume were measured at multiple locations within the pelvis. Clots that measured greater than 10 cm(2) were compared to angiographic results. The chi(2) test was used to find locations on CT that were significantly associated with specific arterial injuries found on angiography. Sixty-one (58%) of the patients had arterial bleeding at angiography. The most commonly injured arteries were the internal pudendal and the superior gluteal. Specific locations on CT were statistically significant indicators of injury to the superior gluteal artery (relative risk=2.9, 95% CI 1.2-7.3, P=0.013), the anterior division of the internal iliac artery (relative risk=3.2, 95% CI 1.4-4.1, P=0.006), and the internal pudendal arteries (relative risk=2.0, 95% CI 1.1-4.0, P=0.037). More blood was visible on CT when an artery was injured (mean volume with negative angiogram=318 ml, mean volume with positive angiogram=554 ml, ( P=0.007)). The rectus sheath region at the top of the iliac crest ( P=0.004), pelvic sidewalls at the L5-S1 disk space level ( P=0.001), and gluteal regions also at the L5-S1 disk space level ( P=0.012) were significant indicators of a positive arteriogram. CT can help predict the specific bleeding artery to potentially guide angiographic intervention. PMID- 15290562 TI - Patterns of diagnostic error in trauma abdominal CT. AB - OBJECTIVE: To define patterns of diagnostic error in the interpretation of trauma abdominal CT. MATERIALS AND METHODS: Two hundred fifty-four out of 1751 abdominal CT scans performed for evaluation of trauma had a definite or equivocal diagnosis of an abdominal injury. Cases were re-read initially without reference to the original reports, in which 44 potential diagnostic errors were identified. A panel of two or three expert readers reviewed each of the 44 cases along with the original report to evaluate the diagnostic error and to search for patterns among the errors. RESULTS: Thirty-one of the 254 CT scans (12%) that were re-read contained non-trivial mistakes that could affect patient outcome. Seventeen were false negative and 14 were false positive. Diagnostic errors were found in the liver, spleen, kidney, retroperitoneum, and peritoneal cavity. Patterns of false negative diagnosis included missed vascular contrast extravasation, missed hemoperitoneum, and missed right retroperitoneal hematoma. Patterns of false positive diagnosis included: periportal edema or blood tracking, called a liver laceration; respiratory motion, called a splenic or renal injury; and linear or round lucencies in the spleen or liver, called a laceration. CONCLUSION: Diagnostic errors in interpreting trauma abdominal CT cluster in several recurring patterns. Awareness of these patterns may assist readers in avoiding future errors. PMID- 15290563 TI - Traumatic injury of the internal mammary artery: embolization versus surgical and nonoperative management. AB - The purpose of the study was to compare the outcomes, complications, and effectiveness of embolization versus surgical and nonoperative management in patients with injury to the internal mammary artery. Eighteen cases of angiographically proven internal mammary artery injury were identified by a retrospective review. Patient age range was 17-71 years (mean 34 years). Causes of vascular injury were equally divided (9 each) between penetrating and blunt trauma. Type of trauma, associated injury, plain film findings, treatment complications (immediate and delayed), and overall outcomes were assessed. Results of embolization versus surgical and nonoperative management were compared. Angiographic findings included occlusion, active hemorrhage, and pseudoaneurysm of the internal mammary artery. Of the 18 patients studied, 12 underwent embolization; 2 underwent surgical ligation, and 4 were managed by nonoperative observation. No patient died as a direct result of vascular injury; one died of renal failure unrelated to chest trauma and one other died of myocardial contusion. One patient who underwent embolization had delayed bleeding and two patients with conservative management developed a delayed hemothorax. This small series demonstrates that embolotherapy offers an effective, efficient, and safe alternative to conventional surgical management of internal mammary artery injuries. PMID- 15290564 TI - Autotransplantation for treatment of severe splenic lesions. AB - The aim of the study was to evaluate clinical and laboratory variables in patients undergoing spleen autotransplantation. We studied 29 patients with severe trauma of the spleen and its pedicle. Of these, 20 underwent autotransplantation (group I) and 9 underwent total splenectomy without preservation of splenic tissue (group II). Twenty-two additional subjects with an intact spleen were used as controls (group III). Immediate and late postoperative complications were investigated. Laboratory counts were performed during the late postoperative period (red blood cells, hemoglobin, white blood cells, platelets, and Howell-Jolly bodies). To investigate the immunological profiles of patients we performed B- and T-lymphocyte counts and determined IgA, IgG, and IgM levels. All patients underwent splenic scintigraphy with technetium 99m sulfur colloid. Groups I and III did not present abnormal blood bodies, and their hematological and immunological patterns were normal. Group II showed increased numbers of Howell-Jolly bodies and low IgM levels. Splenic scintigraphy indicated the viability and filtering function of the splenic remnant in group I. Autotransplantation is a good option to maintain splenic function when total splenectomy is necessary. PMID- 15290565 TI - Serious horse-riding accidents: imaging findings and evaluation with multi-slice CT. AB - PURPOSE: To assess acute phase multi-slice CT findings in horse-riding accidents in patients referred to a level one trauma center. METHODS: Using PACS, we retrieved all CT requests during a time period of 19 months. Patients who had a horse-riding accident and were examined with multi-slice CT were included. The imaging findings of multi-slice CT were retrospectively evaluated by location and injury mechanism. RESULTS: Forty-six patients (3 male, 43 female, age 16-55 years, mean age 30) were assessed. The injuries we found were five head, three facial, 13 spine, five body, two pelvic, and three lower extremity trauma. Multiple injuries were seen in five (11%) patients. In 21 patients (46%) the initial multi-slice CT examination(s) was normal. Three main injury mechanisms were established; falling off a horse (72%), a horse kick (11%), and crushing injury caused by a falling horse (13%). Fifteen (33%) patients had serious injuries, one lethal and they were associated more frequently with falling off a horse. CONCLUSION: Serious injuries with horse riding accidents are not uncommon. In two thirds of patients, the injury mechanism was falling off a horse. Multiple injuries were seen in 11% of cases. In the acute phase, multi-slice CT provided fast and valuable information in assessing these injuries. PMID- 15290566 TI - Pediatric hip pain. AB - This presentation deals with acute hip pain problems in children. These include: fractures, avulsion injuries, infection and inflammation, aseptic necrosis, slipped capital femoral epiphysis, and tumor and tumor-like conditions. Emphasis is on plain film findings and how not to miss them in their subtle forms. PMID- 15290567 TI - Abdominal involvement in tuberculosis. AB - Rising incidence of disseminated and extrapulmonary tuberculosis (TB), especially in immunocompromised hosts and patients with multi-drug-resistant tuberculosis, has resulted in an increase of unusual clinical and radiographic presentations of TB. With CT being a common part of emergency room (ER) evaluation of abdominal pain, it is imperative that radiologists be able to recognize abdominal presentations of TB. We discuss and illustrate typical and less common CT manifestations of tuberculosis in the abdomen to help ER radiologists in this task. PMID- 15290568 TI - Primary epiploic appendagitis. AB - The clinical presentation of primary epiploic appendagitis can mimic diverticulitis or appendicitis. Review of the pathologically confirmed cases in the English literature shows the majority of cases to arise from the sigmoid colon. Pathognomonic findings by computed tomography impact patient management, and it is thus important to recognize these in patients imaged in the emergency setting. PMID- 15290569 TI - Hepatic portal venous gas as a complication of diverticulitis with a colovenous fistula: a case report. AB - We report here a case of sigmoid diverticulitis complicated by a colovenous fistula, as a rare but also a nonfatal cause of hepatic portal venous gas. The different radiological examinations and their features will be discussed. PMID- 15290570 TI - Acute massive subdural hematoma caused by rupture of internal carotid artery aneurysm during angiography: a case report. AB - We present a case of acute, massive subdural hemorrhage caused by rupture of an internal carotid artery aneurysm during the procedure of cerebral angiography. To our knowledge, a case like the present one has been reported only once in the English-language literature. The incidence, mechanisms, treatment, and prognosis of (1) subdural hematoma, caused by rupture of cerebral aneurysm, and (2) re rupture of aneurysm during the angiography procedure are discussed. PMID- 15290571 TI - Spinal myelopathy resulting from decompression sickness: MR findings in a case and review of the literature. AB - This is a case report of diving related decompression sickness causing spinal cord symptoms and unique MR findings. The clinical and imaging manifestations are examined, while the pathophysiology of decompression sickness is reviewed. The variety of imaging findings from similar reported clinical cases have are also discussed. PMID- 15290572 TI - Unique foreign body injury: bamboo penetration of thigh and pelvis while skiing. AB - We present a case of traumatic bamboo foreign body penetration through the posterior thigh extending cephalad into the pelvis sustained during skiing. The unsuspected bamboo foreign body was missed prospectively on the initial portable trauma radiograph of the pelvis, but was retrospectively quite apparent as a linear 12 x 2-cm radiolucent region overlying the left pelvis and hip region. On CT examination, the bamboo stick appeared as a round cylindrical air-filled structure of high density compared to soft tissue - the typical appearance of bamboo. Bamboo foreign bodies can be recognized radiographically by this typical appearance, since it is one of the few wood products that causes high attenuation relative to soft tissue on CT. PMID- 15290573 TI - Blunt trauma to the gastrointestinal tract: CT findings with small bowel and colon injuries. AB - The CT diagnosis of bowel injury is difficult and warrants an organized approach. Careful scrutiny of CT images for extraluminal gas or fluid and bowel wall thickening is required. Review of images of the entire abdomen and pelvis using lung window settings is recommended, followed by the analysis using soft tissue window settings. Specific search for extraluminal fluid collections with a triangular or matted appearance will help detect bowel injury. PMID- 15290574 TI - Acute flank pain: comparison of unenhanced helical CT and ultrasonography in detecting causes other than ureterolithiasis. AB - Several conditions can clinically mimic renal colic. We assessed the accuracy of non-contrast-enhanced helical CT and of ultrasonography (US) in offering an alternative explanation for flank pain. In a 3-year period, 181 patients with acute flank pain underwent US and non-contrast-enhanced helical CT in a blinded sequence. Their efficacy in detecting both alternative causes of pain and additional findings unrelated to the pain was assessed in 160 cases with a confirmed diagnosis. An alternative cause was found in 23 cases (14%). US gave 4 false-negative results (1 acute appendicitis, 1 ovarian cyst torsion, 1 diverticulitis, and 1 papillary necrosis) and 2 false-positive results (1 acute pyelonephritis and 1 diverticulitis), with a 78% sensitivity and a 98% specificity for nonlithiasic causes. CT gave 5 false-negative results (1 complicated ovarian cyst, 1 pleuritis, 1 epididymitis, 1 acute pyelonephritis, and 1 papillary necrosis) and 1 false-positive (1 simple ovarian cyst described as a complicated lesion), resulting in a 74% sensitivity and a 99% specificity for diagnosing alternative causes. There were 130 additional US findings in 68 patients and 151 additional CT findings in 77 patients. A wide spectrum of findings can be identified in subjects imaged for flank pain. Non-contrast enhanced helical CT and US have comparable accuracy in diagnosing causes other than stone disease. PMID- 15290575 TI - Diagnosis of acute appendicitis with unenhanced helical CT: a study of 130 patients. AB - The aim of this study was to determine the sensitivity, specificity, and diagnostic accuracy of helical computed tomography (CT) without oral, intravenous, or rectal administration of contrast material in confirming the diagnosis of acute appendicitis in patients with suggestive clinical and laboratory findings. One hundred and thirty patients with suspected acute appendicitis underwent an unenhanced helical CT scan. Scans were obtained in a single breath-hold from the level of umbilicus to the pubic symphysis using a 5 mm collimation. Oral, intravenous, or rectal contrast materials were not used. The criteria for diagnosis of acute appendicitis included an enlarged diameter of appendix more than 6 mm with associated periappendiceal inflammation. The results yielded a sensitivity of 94.7%, a specificity of 91.7%, an accuracy of 93.8%, a positive predictive value of 96.7%, and a negative predictive value of 86.8%. Unenhanced helical CT accurately diagnoses acute appendicitis, and it protects the patients from unnecessary further time-consuming diagnostic procedures, the risks associated with contrast material administration, and unnecessary surgical interventions. PMID- 15290576 TI - Virtual endoscopy for evaluation of tracheal laceration. AB - Rigid or fiberoptic endoscopy is performed to evaluate the trachea in a patient with suspected tracheal injury. We present a case in which CT virtual endoscopy identified a tracheal laceration not seen on axial or sagittal reformations. This case highlights the usefulness of this modality in the diagnosis and preoperative planning of tracheal injury. PMID- 15290577 TI - Emergency percutaneous retrieval of a silicone port catheter fragment in pinch off syndrome by means of an Amplatz gooseneck snare. AB - Rupture of a silicone Port-a-Cath catheter may occur, especially with costoclavicular pinch-off syndrome (POS), which is a typical consequence of fatigue when the catheter is introduced in the subclavian vein too medially. This case report describes the percutaneous retrieval of a fractured silicone port catheter fragment, which had migrated into the internal jugular vein. Extraction was complicated by the presence of an internal jugular vein stenosis and the fact that the catheter fragment was looped upon itself. Several retrieval devices failed before an Amplatz gooseneck snare finally allowed retrieval of the fragment. We recommend this device for extraction of silicone port catheter fragments. Rerupture of the port catheter occurred 7 months after surgical reinsertion at the same infraclavicular site, as a consequence of constant compression by POS. Alternative approaches should be used after catheter failure due to POS. PMID- 15290578 TI - Osteomyelitis mimicking a primary bone tumor in a patient with pulmonary arteriovenous malformation and Osler-Weber-Rendu disease. AB - Extremity pain is a common presenting symptom seen by the practitioner; the etiology may be related to problems in the bone, joint, soft tissues, or neurovascular bundle. The differential diagnosis is extensive for each, with a careful history and physical exam narrowing the list. Imaging begins with plain radiographs, reserving CT and MRI for further work-up. We report an unusual presentation of osteomyelitis mimicking a bone tumor in a patient with Osler Weber-Rendu disease. PMID- 15290579 TI - Cocaine-induced mesenteric ischemia: treatment with intra-arterial papaverine. AB - Mesenteric vascular ischemia is a known complication of cocaine use. Although the majority of cases of cocaine-induced mesenteric ischemia present with ischemic colitis and rectal bleeding, several cases have been described presenting only with abdominal pain. We present a case of mesenteric vasoconstriction with angiographic documentation and treatment. PMID- 15290580 TI - Hepatic leiomyomas in two adult patients with AIDS: intravenous contrast-enhanced CT and MR imaging. AB - Multiple opportunistic infections and neoplasms have been described in the AIDS population over the past 20 years. Four cases of AIDS-related hepatic smooth muscle tumors (leiomyomas) have been described, and of these, three were in children. The fourth case was in a single patient with two hepatic lesions. We describe the imaging features of two adult cases with biopsy-proven hepatic leiomyomas. In one of the cases, the gadolinium-enhanced MRI findings are described, which have not previously been reported. PMID- 15290581 TI - Utilization of CT-PA in an emergency department with readily available V/Q scintigraphy. AB - PURPOSE: To describe the role of computed tomographic pulmonary angiography (CT PA) in the emergency department of an institution which utilizes ventilation perfusion (V/Q) scintigraphy as its primary imaging modality for the diagnosis of pulmonary embolism. METHODS AND MATERIALS: We retrospectively identified and reviewed the records of 24 consecutive patients evaluated in the emergency department from October 1998 to September 2000 who were suspected of having pulmonary embolism. The inclusion criterion was that the images from the emergency department and work-up included CT-PA, which was the initial imaging test after chest radiograph. There were 10 men and 14 women with a mean age of 63. Results of CT-PA were categorized as positive, negative, limited negative (no main or lobar pulmonary emboli but not all segmental arteries visualized), or nondiagnostic. Each chart was reviewed with reference to clinical presentation, relevant history, results of Doppler ultrasonography of the legs, V/Q scan and pulmonary angiography, and discharge diagnosis. During the same study period, approximately 400 V/Q scans were performed from the emergency department. RESULTS: Each patient had a clinical presentation consistent with pulmonary embolism. CT-PA diagnosed pulmonary emboli in 21% (5/24), was negative in 33% (8/24), was limited negative in 38% (9/24), and was nondiagnostic in 8% (2/24). Chest radiographs were abnormal in 71% (17/24). V/Q scans were performed in 17% (4/24; 1 near normal, 2 low probability, 1 intermediate probability). None of these four patients was discharged with a diagnosis of pulmonary embolism. Doppler ultrasound leg exam was performed in 38% (9/24). Among the 5 patients diagnosed with pulmonary embolism, 1/3 examined had Doppler evidence of deep vein thrombosis (DVT). Among the 19 patients not diagnosed with pulmonary embolism, 3/6 examined had Doppler evidence of DVT. No patient with a negative or limited negative CT-PA was discharged with a diagnosis of pulmonary embolism. CT-PA provided alternative diagnoses explaining the patient's symptoms in 68% (13/19) of those not diagnosed with pulmonary embolism. During the most recent 12 months of the study period, 210 V/Q scans were performed from the emergency department, with results available in 194 cases as follows: normal/near normal 32% ( n=62), low probability 47% ( n=92), intermediate probability 14% ( n=28), high probability 6% ( n=12). CONCLUSION: V/Q scintigraphy is the primary imaging modality for suspected pulmonary embolism in our emergency department. However, when utilized, CT-PA played an important role in patient management by confirming or excluding pulmonary embolism or providing an alternative diagnosis in the majority of patients suspected of having pulmonary embolism. PMID- 15290582 TI - Massive pulmonary embolism: a comparison of radiological and clinical characteristics and outcomes. AB - STUDY OBJECTIVES: To describe the clinical features of radiographically massive pulmonary embolism (MPE). DESIGN: Retrospective analysis. SETTING: A 1,368-bed teaching hospital. PATIENTS OR PARTICIPANTS: Patients with pulmonary embolism between June 1997 and December 1999. INTERVENTIONS: Radiographic reports of patients with a radiographic diagnosis of pulmonary embolism were reviewed to determine whether MPE (>50% vascular occlusion) was present. For patients with MPE, vital signs, respiratory and cardiac symptoms, medical history, arterial blood gases, electrocardiographic (ECG) and echocardiographic results, treatment, and hospital mortality were recorded. MEASUREMENTS AND RESULTS: Fifty-four patients with MPE were identified. Patient age range was 28-91 years (mean 71 years). Symptoms were: dyspnea in 38 (70%), chest pain in 21 (38%), syncope in 12 (22%), palpitations in 6 (11%), systolic blood pressure <90 mmHg in 12 (22%), tachycardia (>120 beats/min) in 15 (28%) and tachypnea (respiratory rate >30) in 15 (28%). Pa O(2) (arterial partial pressure of oxygen) was less than 60 mmHg in 28 (71%) and the alveolar-arterial oxygen gradient was always greater than 20. ECG had an S1Q3T3 pattern in 6 (12%). Echocardiography revealed right ventricular dilatation in 12/31 (38%). Forty-nine patients received anticoagulation treatment, 4 (7%) received thrombolytic therapy with anticoagulation, 5 had inferior vena cava filters (IVC) alone, 6 received IVC filters with anticoagulation, and 2 received thrombolytic therapy, anticoagulation, and IVC filters. Eighteen (33%) patients were treated in the intensive care unit, 3 (5.5%) with mechanical ventilation. Fifty (93%) patients were eventually discharged and 4 (7%) died. Two of the deaths were not attributable to MPE. CONCLUSIONS: Patients with MPE usually present with dyspnea and hypoxemia, and most survive without thrombolytic therapy. PMID- 15290583 TI - The graveyard shift: experience with a night float system. AB - The aim of this study is to describe the effects of a new night float system on the circadian rhythm and clinical judgment of our residents. In addition, the study looks at the residents' opinions of how to optimize the night float system in the future. All 20 of the radiology residents at our institution completed a questionnaire about the night float system after completing their night float coverage. The results of the questionnaire were then compiled and tabulated. It took our residents an average of 2.0 days to become acclimated to the night float and an average of 2.3 days to return to a normal daily routine after completing the night float. No residents perceived impairment in their clinical judgment while on the night float. However, 9 of the 20 residents (45%) stated that their clinical judgment was improved on the night float compared to that of a 24-hour call. Eighteen of 20 residents (90%) preferred the night float system to a 24 hour call system. On average, our residents believe that the optimal number of hours for a night float shift is 10.5 hours and the optimal numbers of days to do the night float consecutively is 6.8 days. In conclusion, a night float system can be a preferable means of evening coverage as it has a minimal effect on the circadian rhythm by allowing residents to become acclimated to working the night shift over the course of several days. The night float system also demonstrates no appreciable adverse effects on clinical judgment and may allow better clinical judgment than a 24-hour call system. PMID- 15290584 TI - Reappraisal of use of X-rays in childhood ankle and midfoot injuries. AB - OBJECTIVES: To determine whether the Ottawa ankle rules (OAR) can be applied in children and what the potential reduction in the use of X-ray studies might be. METHODS: Prospective, observational study of children attending emergency department with blunt ankle and midfoot injuries. X-ray studies were obtained after recording of the physical assessment. A single investigator who was blinded to the emergency physician's interpretations and to the radiologist's interpretations of the studies then applied the OAR to each patient. Sensitivity, specificity, and positive and negative predictive values of the OAR were calculated with 95% confidence intervals. RESULTS: The OAR were 100% sensitive (95% CI=81-100%) and 30% specific (95%CI=19-43%). Positive and negative predictive values were 28% (95%CI=17-41%) and 100% (95% CI=82-100%) respectively. The number of X-ray studies ordered would have been reduced by 24% if the OAR had been applied. CONCLUSIONS: The Ottawa ankle rules are very sensitive and can be applied in children, resulting in a reduction in the use of X-rays studies. PMID- 15290585 TI - CT findings of small bowel trichobezoar. AB - A 14-year-old girl presented to our emergency room with abdominal pain and persistent vomiting. A plain radiograph of the abdomen showed features of small bowel obstruction, with dilated loops of small bowel and a mottled gas and debris pattern in the stomach and right lower quadrant. A CT scan of the abdomen and pelvis obtained to exclude appendicitis showed the distinctive appearance of a trichobezoar. A preoperative diagnosis of gastric and intestinal trichobezoar was made. PMID- 15290586 TI - Portomesenteric vein gas: diagnostic and prognostic value. AB - Portomesenteric pneumatosis has been traditionally associated with intestinal infarction and poor outcome; however, recent studies have questioned its clinical value. To assess its diagnostic and prognostic significance we have retrospectively evaluated 47 patients correlating the CT finding of portomesenteric vein gas with clinical data and outcome. Thirty-nine patients (83%) had surgical evidence of intestinal infarction, four had necrotic small bowel volvulus (8.5%), two had blunt trauma, one had necrotic gastric volvulus, and one a gastric mucosal lesion induced by a nasogastric tube. Fifteen patients survived (31.9%); only 8/39 patients with intestinal infarction survived. Portomesenteric pneumatosis is a reliable marker of intestinal infarction and poor outcome; however, in trauma patients this sign is associated with a better prognosis. PMID- 15290587 TI - Reliability of vacuum phenomenon in the sacroiliac joint as a sign of traumatic injury. AB - OBJECTIVE: To determine whether a vacuum phenomenon in the sacroiliac joint is a reliable sign of pelvic injury in trauma patients. MATERIALS AND METHODS: Prospective data were collected over a 1-year period for 107 patients with pelvic trauma and 104 nontrauma patients. Age ranges were 13-93 years in the trauma group and 19-83 years in the nontrauma group. All the patients had pelvic CT scans. The cases were assessed with regard to gas in the sacroiliac joint, osseous pelvic injuries, and mechanism of injury, and demographic data were analyzed. Injuries were caused by motor vehicle accidents in 67 cases, pedestrians being struck by a motor vehicle in 20, falling from a height in 18, gunshot wound in 1, and crush injury in 1. The indications for CT scan in the nontrauma patients were pain in 33 cases, infection in 31, cancer in 29, transplant in 5, bleeding in 4, and abnormal liver function tests in 2. RESULTS: Gas in the sacroiliac joint was present in 11 out of 107 trauma patients (10%) and 12 out of 104 nontrauma patients (12%). There was no statistical difference in the incidence of the vacuum phenomenon between the two patient populations according to the Chi(2) test. Degenerative sacroiliac changes were evident in 18 out of 107 trauma patients (17%) and 32 out of 104 nontrauma patients (41%). CONCLUSION: Gas in the sacroiliac joint is not a reliable indicator of sacroiliac joint injury. PMID- 15290588 TI - Ureteral leak after an abdominal aortic aneurysm repair: case report. AB - Iatrogenic ureteral injuries are an infrequent complication of vascular reconstructive surgery, and if they are not suspected at the time of surgery the diagnosis is usually delayed. Diagnosing these injuries may be challenging, since patients usually show signs and symptoms appropriate to a normal postoperative course and usually do not develop hematuria or renal dysfunction. In the proper clinical setting, a fluid collection adjacent to the ureter on cross-sectional imaging studies should alert the emergency radiologist to the possibility of ureteral injury. A high clinical suspicion would allow earlier diagnosis and treatment, potentially reducing the morbidity and mortality associated with a delay in diagnosis. We present a case of a ureteral leak diagnosed 1 week after an abdominal aortic aneurysm repair. A fluid collection seen adjacent to the ureter on contrast-enhanced CT prompted the radiologist to obtain delayed images that demonstrated urinary extravasation. PMID- 15290589 TI - Hepatic transplant complicated by hepatic artery thrombosis and bile duct necrosis: case report and potential application of contrast-enhanced MR cholangiography following intravenous mangafodipir trisodium in the emergency room setting. AB - Biliary tract strictures and leaks are the second most common complications following orthotopic liver transplantation. Nonanastomotic bile duct complications are most often caused by hepatic artery thrombosis and can result in fulminant hepatic necrosis, bile duct strictures, and bile duct leaks that increase the risk of cholangitis, sepsis, and abscess. The emergency physician and radiologist should strongly suspect biliary disease in a post-transplant patient presenting with elevated liver function tests, jaundice, fever, and/or abdominal pain in order to achieve diagnosis and treatment rapidly. We present the case of a liver transplant patient who developed bile duct necrosis and hepatic infarction secondary to hepatic artery thrombosis 5 months after surgery. In addition, we discuss a new contrast-enhanced MR cholangiographic technique that has the potential to be performed in the emergency setting as the only diagnostic test prior to appropriate therapy. PMID- 15290590 TI - Recurrent gallstone ileus: a case report. AB - Gallstone ileus is a rare complication of recurrent gallstone cholecystitis and usually occurs in elderly female patients. Recurrent gallstone ileus occurs in 5% of patients with a previous episode of gallstone ileus and is associated with a mortality of 20%. We present a 52-year-old female with recurrent gallstone ileus 1 year after her initial episode. PMID- 15290591 TI - A hematoma of the esophagus causing the trachea to stenose. AB - We report a case of spontaneous intramural hematoma of the esophagus (SIHE) with severe dyspnea due to compression of the trachea. SIHE is a rare hematoma that commonly presents with chest pain, epigastralgia, hematemesis, and dysphagia. Dyspnea is not a common symptom; it has been reported in only one patient, who underwent surgery. In our case, intubation of the compressed trachea prevented it from becoming more stenosed, and an operation was not needed. Another unusual feature of this case is the endoscopic findings. Endoscopic examination in SIHE has often revealed the presence of a dark red, bluish, or purplish bulge, suggesting the presence of a clot or blood in the esophageal wall. In our case, the bulge revealed by endoscopy in the esophageal lumen was white at first, before later turning dark red. PMID- 15290592 TI - Blunt submandibular gland trauma: acute CT findings. AB - We describe an early CT study of a rare case of blunt traumatic injury to the right submandibular salivary gland, without mandibular bone fractures, in a 30 year-old man after a car accident. PMID- 15290593 TI - Blunt nonaortic chest trauma: radiographic and CT findings. AB - Following initial clinical evaluation and stabilization of a patient who has sustained blunt chest trauma, imaging has an important role in the evaluation of thoracic injuries. The initial study is the chest radiograph. However, chest CT is being used with increased frequency in the evaluation of blunt chest trauma. Although CT is used primarily to assess for traumatic aortic injuries, it is also useful in the evaluation of pulmonary and bronchial, airway, skeletal and diaphragmatic injury. The aim of this article is to review the characteristic imaging findings of pulmonary and bronchial, esophageal, thoracic, skeletal and diaphragmatic injuries. PMID- 15290594 TI - Traumatic aortic injury: an imaging review. AB - Traumatic aortic injury (TAI) is a major cause of fatality in high speed deceleration injuries. It accounts for 10-20% of fatalities in blunt chest trauma. These injuries are usually related to high-speed motor vehicle and motorcycle collisions, pedestrian-motor vehicle collisions, and falls. Only 10 20% of patients who suffer TAI survive the initial injury and reach the emergency department. If left untreated, 30% die within 6 h, 40-50% die within 24 h, and 90% die within 4 months. A chronic pseudoaneurysm will develop in 2-5% of patients whose injury is not diagnosed. It is imperative, therefore, that these injuries are detected promptly and accurately. Symptoms and physical examination findings are nonspecific. External evidence of chest wall injury is present in 7 90% of cases, so that in up to 30% of the cases no apparent chest injury is identified on physical examination. Chest radiographs are very sensitive in detecting mediastinal hemorrhage, but have a low positive predictive value for aortic injury. The positive predictive value for chest radiography ranges between 5% and 20% for TAI. Aortography has been considered the gold standard for many years in the evaluation of TAI, but is time-consuming, labor- and resource intensive, and invasive. Because of the shortcomings of physical examination and these more traditional imaging examinations, computed tomography (CT) has become increasingly utilized as a screening and diagnostic tool. Recent investigations have documented its high sensitivity and specificity in the detection of TAI. This article reviews the recent investigations of imaging evaluation of TAI, with a focus on helical CT. PMID- 15290595 TI - Pediatric thoracic trauma: imaging considerations. AB - Imaging plays an important role in the evaluation of the pediatric thorax following blunt and penetrating trauma. This essay reviews important differences between children and adults with respect to the pattern of thoracic injury. Additionally, the role of various imaging modalities in the assessment is discussed. Finally, the spectrum of pediatric thoracic injury is provided. PMID- 15290596 TI - Value of contrast-enhanced CT for managing mesenteric injuries after blunt trauma: review of five-year experience. AB - OBJECTIVE: The purpose of this retrospective study was to determine the value of contrast-enhanced computed tomography (CT) in the detection and management of mesenteric injuries after blunt trauma. MATERIALS AND METHODS: Between June 1995 and December 2000, 1,619 consecutive abdominal CT examinations were performed in the setting of major blunt trauma. Findings at CT were evaluated before patients were classified as having grade 1 or grade 2 lesions or none. Grade 1 represented the presence of minor injuries: mesenteric haziness, confined fluid, and/or small hematomas (<30 mm) within the mesenteric folds and abdominal injuries; grade 2 was appropriate to evidence of major injuries: moderate to large hematomas (>30 mm), active bleeding, hemoperitoneum, and further abdominal injuries. RESULTS: On the basis of the CT findings, 161 (9.9%) of 1,619 patients were classified as having grade 1 and 25 (1.5%) of 1,619 patients as having grade 2 injuries. Of the 161 (77.6%) patients with grade 1 injuries, 125 were managed conservatively, while 36 (22.4%) underwent surgery. Of the 25 (84%) patients with grade 2 injuries, 21 were treated surgically and 1 (4%) patient was followed medically. Three (12%) of the 25 patients underwent laparotomy after 24 h close clinical observation and monitoring. Initial CT findings in 1,433 (88%) of the 1,619 patients were negative for mesenteric injuries, and in 1,430 of these cases no delayed mesenteric hemorrhage was observed. CONCLUSION: Contrast-enhanced CT has a critical role in the identification and exclusion of mesenteric injuries. Persistent, active extravasation of contrast material, in isolation or associated with further abdominal lesions, is a sign of a high likelihood of injury requiring urgent laparotomy. Haziness, isolated confined clotted mesenteric hemorrhage, and small hematomas within the mesentery are nonspecific findings and should be considered in the appropriate clinical context. Close clinical observation, monitoring, and surgical expertise are mandatory for appropriate management. PMID- 15290597 TI - Bystander trauma in the World Trade Center disaster. AB - We present the CT and clinical findings in a 21-year old male who presented to the emergency department following bystander trauma during the rescue effort of the World Trade Center disaster. PMID- 15290598 TI - Emergent MRI utilizing a 5-inch surface coil to evaluate for acute penile fracture. AB - MR imaging is useful in rapidly detecting penile fractures and in guiding surgical planning. PMID- 15290599 TI - Cervical spine fractures in ankylosing spondylitis: MR findings. AB - PURPOSE: The purpose of this study is to evaluate the MR findings in patients with long-term ankylosing spondylitis (AS) and acute cervical spine fractures. MATERIALS AND METHODS: The magnetic resonance imaging (MRI) studies of five patients with AS and acute cervical spine fractures were retrospectively reviewed for the presence of cervical spine instability, spinal cord compression, and epidural hematoma. RESULTS: Spinal fractures were unstable in all five patients. Three patients had neurological symptoms and abnormal signal within the spinal cord. All patients with neurological deficits had epidural hematomas posterior to the dural sac. CONCLUSION: MRI is useful for assessment of the integrity of intervertebral disks and spinal ligaments and, therefore, of the instability of the spinal fracture. MRI is mandatory in patients with neurological symptoms, especially in those with a symptom-free interval and those with neurological deterioration after established spinal cord injury, when suspicion for epidural hematoma is high. PMID- 15290600 TI - ICU chest radiographs - ICU calamities: evaluation of the portable chest radiograph. AB - Familiarity with anatomy and anatomic variants, with tubes and catheters and their correct positioning, as well as complications of these positionings, is critical for the radiologist interpreting plain film radiographs taken in the intensive care unit. PMID- 15290601 TI - Comparing the interpretations of CT pulmonary angiograms by attending and resident radiologists: can residents identify life-threatening pulmonary emboli in hospitalized patients? AB - OBJECTIVE: CT pulmonary angiography is now often the first-line investigation for pulmonary emboli. When these studies are performed after hours in teaching hospitals, they are often initially interpreted by trainees. It is of great significance whether the interpretations of trainees and certified radiologists with regard to the presence of pulmonary emboli on CT pulmonary angiograms correspond, because of the morbidity and mortality of both the condition and its treatment. MATERIAL AND METHODS: Twenty-five consecutive CT pulmonary angiograms (CTPAs) of hospitalized patients were viewed at lung and soft tissue windows both on a workstation and on hard copies, at the observers' discretion. Each CTPA was divided into 28 arterial zones based on pulmonary anatomy (including the subsegmental arteries), giving a total of 700 arterial zones, and analyzed retrospectively and independently by two cross-sectional imaging specialists and four residents. Each arterial segment was rated with regard to pulmonary embolus as either high, intermediate, or low probability or not visualized. The kappa (Kappa) test, which tests for interobserver agreement, was used for statistical analysis. RESULTS: At the time of the scan all patients were hospitalized for underlying conditions. Of the 25 patients studied, 9 were referred from the ICU, 7 experienced severe acute shortness of breath and respiratory failure, 5 were post-partum women, 2 had had a recent stroke, 1 patient had antithrombin III deficiency, and 1 had a diagnosis of breast cancer. The incidence of pulmonary emboli was 44%. For the main pulmonary arteries interobserver agreement was good (Kappa=0.61) and for the segmental pulmonary arteries it was fair (Kappa=0.26). For the subsegmental arteries interobserver agreement was poor (Kappa=0.16). The zones where interobserver agreement was greatest (Kappa>0.4) were the left main, left lower lobe, and the right main pulmonary arteries. Interobserver agreement was poorest (Kappa<0.05) in the left interlobar, left lower lobe lateral basal segment, right lower lobe superior segment, and left lower lobe superior segment branches. None of the patients expired due to pulmonary emboli. CONCLUSION: Most life-threatening pulmonary emboli requiring urgent treatment are the more central emboli. This study demonstrates that trainees and certified radiologists can make similar conclusions regarding these central pulmonary emboli in hospitalized patients and that preliminary interpretations by trainees should not therefore adversely affect patient care. PMID- 15290602 TI - Ureterolithiasis: classical and atypical findings on unenhanced helical computed tomography. AB - Evaluation of patients with acute flank pain using helical computed tomography (CT) is a well-accepted, rapid, and safe procedure in the emergency setting. Various primary and secondary signs are described in the literature for evaluation of these patients. Our purpose is to demonstrate both the classical findings associated with ureteral calculi on unenhanced helical CT and atypical findings and potential pitfalls. We also provide readers with a systematic approach to interpreting unenhanced helical CT scans performed for acute flank pain. PMID- 15290609 TI - Causes, prevention, and surgical treatment of postherniorrhaphy neuropathic inguinodynia: triple neurectomy with proximal end implantation. AB - The recommended surgical treatment for chronic neuropathic pain after herniorrhaphy has been a two-stage operation including: (a) ilioinguinal and iliohypogastric neurectomies through an inguinal approach and (b) genital nerve neurectomy through a flank approach. Two hundred twenty-five patients underwent triple neurectomies with proximal end implantation to treat chronic postherniorrhaphy neuralgia. Four patients reported no improvement. Eighty percent of patients recovered completely, and 15% had transient insignificant pain with no functional impairment. These results are comparable to the results of the two-stage operation. Simultaneous neurectomy of the ilioinguinal, iliohypogastric, and genital nerves without mobilization of the spermatic cord is an effective one-stage procedure to treat postherniorrhaphy neuralgia. It can be performed under local anesthesia and avoids testicular complications. Proximal end implantation of the nerves prevents adherence of the cut ends to the aponeurotic structures of the groin, which can result in recurrence of the pain. A one-stage surgical procedure resecting all three nerves from an anterior approach avoids a second operation through the flank and successfully treats chronic neuralgia. PMID- 15290610 TI - Intestinal perforation as a long-term complication of plug and mesh inguinal hernioplasty: case report. AB - Tension-free and sutureless hernioplasty by plug and mesh of nonreabsorbable material is one of the most common techniques for inguinal hernia repair. It's a simple and quick procedure with a low cost and allows for a short hospital stay. It shows a low reoccurrence rate, but it can result, in very few cases, in complications strictly related to prosthetic material. The literature describes some cases of plug migration from its proper position, for example, to the scrotum, preperitoneal adipose tissue, and abdominal cavity. We report on a case of sigmoid colon perforation due to a plug of Trabucco hernioplasty performed 2 years previously. PMID- 15290611 TI - Comparison of two composite meshes using two fixation devices in a porcine laparoscopic ventral hernia repair model. AB - INTRODUCTION: Laparoscopic ventral hernia repair uses tacks to secure mesh. The mesh is designed to maximize tissue ingrowth while minimizing adhesions. We hypothesized: (1) a collagen-coated polyester mesh (PCO) will form fewer adhesions than an ePTFE-polypropylene composite (BC) and (2) absorbable tacks are equivalent to metal tacks. METHODS: In a porcine model of adhesion formation, three pieces of 10x15-cm mesh were placed on the anterior abdominal wall. PCO was secured with absorbable (PLA) or metal tacks (PT), BC with PT. At 28 days, adhesion formation, abdominal-wall adherence, and tissue ingrowth were analyzed. RESULTS: PCO induced fewer adhesions (14.5% vs 53.4%, P = 0.007). On an adhesion scale (0 5), BC scored 3.6 vs 1.75 for PCO (P < 0.03). There was no difference in adhesion strength, tack adhesions, or abdominal-wall peel force. Histology showed equal ingrowth. CONCLUSIONS: PCO induces fewer adhesions than BC. There is no difference in the ingrowth of the two mesh types. The PLA achieves equivalent mesh incorporation to the PT. PMID- 15290612 TI - Incarcerated vermiform appendix in a left-sided inguinal hernia. AB - We report here of a patient with an incarcerated vermiform appendix occurring in a left-sided indirect inguinal hernia. Occasionally, appendices are found in a hernial sac; however, the finding of an incarcerated vermiform appendix in an inguinal hernia on the left side is very unusual and has only been previously described once. The patient suffering this rare entity underwent appendectomy and repair of the hernia and experienced an uneventful postoperative recovery. The possibility of the presence of a situs inversus, or malrotation, as an underlying cause for the observed pathology was excluded by x-ray examination. PMID- 15290613 TI - Long-term results after modified Gallie technique for incisional hernia repair: results in 19 patients. AB - BACKGROUND: A modified forgotten technique for repairing large incisional hernias is described together with its long-term results in 19 patients. A synthetic mesh with 1-cm wide spokes radiating from the mesh is placed preperitioneally, overlapping the fascial defect. The spokes are pulled through rectus sheaths and muscle and sutured ventrally, thereby creating a solid reconstruction withstanding shrinking of the mesh. METHOD: Nineteen patients were operated on (13 primary incisional hernia, minimal fascial defect 10 cm). Notes on patients were reviewed, and the patients were contacted for follow-up examination. RESULTS: No major complications occurred. After a median of 49 months, 17 patients were reviewed at the outpatient clinic. Two possible recurrences were detected, of which one was operated on. This proved to be bulging of the mesh, resulting in a recurrence of 1 out of 17 (6%). CONCLUSION: From these results, it is concluded that Gallie's technique using synthetic mesh is a safe and effective repair for incisional hernia and deserves more attention, especially for large fascial defects. PMID- 15290614 TI - Prognosis factors in incisional hernia surgery: 25 years of experience. AB - BACKGROUND: Incisional hernia underwent a change from conventional techniques to mesh implantation. The relevance of different factors, like operative technique, mesh material, and patient-related parameters concerning the outcome following mesh repair, are still under debate. METHODS: In a comparative retrospective study of 421 incisional hernia operations on 348 patients, we investigated 241 Mayo procedures and 180 mesh repairs over a 25-year period. In addition to the quality of life following mesh implantation, the prognostic relevance of demographic, preoperative and intraoperative parameters, surgical technique, mesh material, and the surgeon's experience were analysed, both in a univariate and multivariate manner. RESULTS: With a mean follow-up of 9.7+/-8.8 years, the total recurrence rate following Mayo overlap was 37%, in contrast to 15% after mesh implantation (P=0.001). Mesh size was the only significant prognostic factor concerning quality of life following mesh implantation, and 86% of the patients with mesh repair were satisfied. The complication rate was determined significantly by patients' risk factors, size of hernia, operative technique, and the surgeon's experience, whereas the rate of recurrences was significantly influenced by the parameters obesity (BMI>25), size of hernia, and surgical experience. The recurrence rate decreased significantly with surgeon's experience a minimum of 16 mesh repairs led to a recurrence rate of less than 10%. CONCLUSIONS: Only the mesh repair revealed acceptable recurrence rates with high patient comfort. From a surgical point of view, the most important prognostic factor following mesh repair is the surgeon's experience. PMID- 15290615 TI - Further study of tissue adhesive. PMID- 15290636 TI - [Nursing education in Taiwan: the current situation and prospects for the future]. AB - Two factors which have a close bearing on nursing education are social and technological change. How nursing education meets social needs and how nursing students are best equipped with professional knowledge, attitudes, and skills in a rapidly changing society have become important issues. The purposes of this paper are to describe the current state of nursing education and to identify problems that need to be solved, including the complexity of the educational system, the quality and quantity of nursing manpower, the degree of compatibility between the aims of education and future social needs, the quality of nursing educators, courses and course delivery, and the clinical competence of nursing graduates. Based on the paradigm shifts of International Medical Care and higher education described by McBride, several suggestions for future educational reform are proposed, including simplifying the educational system, improving the development of both the quality and quantity of nursing manpower, matching educational aims with future social needs, bridging the gap between theory and clinical practice, promoting both instructional and clinical skills among nursing educators and enhancing the ability of graduates to deal with a contingency strain. It is expected that in order to meet the challenges presented by a rapidly changing society, nursing educators will have to develop a mixed role including clinical proficiency, outstanding research skills, and excellent leadership skills. PMID- 15290637 TI - [Current challenges facing the technological and vocational nursing education in Taiwan]. AB - The technological and vocational education nursing system (TVE) has been the major source of nursing labor in Taiwan. As educational reform progresses, entry into nursing practice will soon mean that all nurses hold at least an associate nursing degree, which will be a milestone in the nursing history of Taiwan. However, the ecology of nursing is fast-altering, so in order to produce competent nurses, endeavors are required to renovate the current TVE system of nursing. In this article, the author brings years of personal experience as a nursing educator to bear in raising several key issues relating to the TVE system of nursing education. PMID- 15290638 TI - [Taiwan nursing accreditation for nursing education]. AB - On 1st January 2003, with the support and assistance of Professor Kun-Yan Huang, the Ministry of Education assembled its University Nursing Departments Evaluation Planning Group. Professor Yu-Mei Yu Chao led the group in over a year of hard work and planned an accreditation of systematic evaluation of university nursing departments. The hope is to accumulate experience through this accreditation to promote nursing undergraduate programs evaluation to the Ministry of Education's technical and vocational training system. This article illustrates the 12 core disciplines that the evaluation system stresses as embodying the spirit and aims of nursing. Once the spirit and aims of education have been dealt with, the evaluation turns to the six important matters of conduct of departmental affairs, faculty, students, curriculum and teaching, teaching resources and educational outcomes. The system is expected to raise the quality of nursing education, help in the monitoring of the citizens' health care, and raise the success and satisfaction levels of students. PMID- 15290639 TI - [Future trends in nursing education in Taiwan in the light of globalization]. AB - The twenty-first century is the era of the knowledge-based economy. Its information networks developing rapidly, Taiwan has already entered an age of liberalization, diversity and globalization. Competition and change will be the norm. As globalization continues it will pose substantial problems for nursing education. Nursing is a service-oriented activity which has to develop constantly to meet the changing demands of the public as people start to live longer, society becomes more multi-cultural, the nature of diseases and other health problems changes and public policy, such as that on National Health Insurance, is modified. This article outlines the problems currently facing nursing education (i.e., the complexity of the educational system, shortcomings in the learning environment, curriculum design, the quality of faculty, evaluation methods, and the quality of students' English and Mathematics) to predict likely difficulties (i.e. student recruitment, the running of schools and the quality of clinical nurses) and trends in nursing education. (i.e. changes in the way schools are run in line with the impact of globalization, new teaching methods; faculty training and development, lifelong learning, and the internationalization of education.) The article should be of interest to nursing educators. PMID- 15290640 TI - [Transition from student nurse to staff nurse]. AB - When new staff nurses leave school and enter the working world they experience a shock that is all the greater because of their limited clinical experience. Difficulty in adjustment and a high attrition rate have been noted among new staff nurses as they navigate this challenging and stressful transition. New staff nurses experience excessive workloads, a shortage of practical and managerial skills, great differences between their educational priorities and the reality of clinical practice, and the feeling of a lack of support. Setting up a preceptorship system to support and guide the professional development, psychological adjustment and career planning of individual nurses would help to foster positive socialization, reduce levels of anxiety and increase job satisfaction among nurses. In turn, if this is accomplished, it will lower work related stress, reduce attrition rates and stabilize levels of nursing manpower. PMID- 15290641 TI - [Symptom distresses and coping strategies in breast cancer women with mastectomy]. AB - The purpose of this descriptive cross-sectional survey was to understand the correlation between symptom distresses and coping strategies in female victims of breast cancer with in two years of their undergoing a mastectomy. The instruments used in this study were the Physical Symptoms Scale, short form of Profile of Mood states (POMS-SF), Social Function Distress Questionnaire, Jalowiec Coping scale (JCS), and demographic data. The data were analyzed by descriptive statistic, t-test, one-way ANOVA, post Scheffe test and Pearson product-moment correlation. The essential results are summarized as follows. Degrees of biopsychosocial distresses varied from none to mild. Physical symptom distress, psychological distress, and social function distress were positively correlated. When patients were confronted with symptom distresses, they tended to adopt problem-focused coping strategies. The more problem-focused coping strategies, they used the less symptom distresses they experienced. The results of this study could be used to help to develop nursing interventions and efficient coping strategies. Patient may then be able to use the latter to solve symptom distresses and enhance their quality of life. PMID- 15290642 TI - [The relationship between dysmenorrhea and menstrual attitudes among female students in vocational nursing schools]. AB - This research involved a cross-sectional investigation. An Essential background questionnaire and the Menstrual Attitude Questionnaire (MAQ) were used in the study. The purpose was to explore the relationship between dysmenorrhea and menstrual attitudes among female students in vocational nursing schools. Data collection was performed by questionnaire in school classes. Data were recorded using Microsoft Excel, and analyzed by JMP5.0. Essential background information was compared by t-test and Chi-square test, and menstrual attitudes by t-test. Eight hundred and five students answered the questionnaire, and 760 responses (94.4%) were valid. Among the valid sample, 557 respondents (73.3%) had experienced dysmenorrhea during the previous half year. This group was named the dysmenorrhea group. The other 203 students (26.7%) who had experienced no dysmenorrhea during the previous half year were called the non-dysmenorrhea group. Results of the study showed that the dysmenorrhea group agreed that "Menstruation is a debilitating event," and on the importance of "Anticipation and prediction of the onset of menstruation" more than the non-dysmenorrhea group. Markedly more of the non-dysmenorrhea group than of the dysmenorrhea group that agreed, "Menstruation has no effect". As to the propositions that "Menstruation is a bothersome event," and "Menstruation is a natural event," there were no significant differences between the two groups. PMID- 15290643 TI - [Strategies to improve protection from glutaraldehyde in endoscopic examination]. AB - Safety of the working environment has emerged as an important issue in recent years. Of safe work place is a prerequisite for the practice of medicine. High level disinfection by glutaraldehyde (GAD) is the usual method for sterilizing endoscopes. But GAD is highly volatile and is an irritant. Prolonged exposure to GAD in a closed unit usually causes discomfort. Our study demonstrated that the main reasons for this are: (1) nursing staff are not sufficiently familiar with GAD, (2) high GAD concentration in the air, (3) insufficient training to use the facilities and disinfectant, (4) lack of safety precautions due to large amount of GAD in examination room. In order to diminish the dangers of unavoidable exposure, the following strategies are recommended: intensify training for the handling of GAD, decrease the amount of GAD, encourage the use of individual precautions and remove fuzzy carpets, and use appropriate disinfection facilities. Many improvements can be achieved with the introduction of these measures as, this article shows. These improvements include: (1) a 100% response rate and a 97% satisfaction rate from trainees, (2) a total reduction of 150 liters of GAD per month with atmospheric GAD markedly reduced, to 0.0054 ppm, (3) reduction in reported discomforts from between 80% and 100% to 10%, (4) a protection rate of 100% compared to the previous 10% - 30%. PMID- 15290644 TI - [A primary exploration of nursing practicum during epidemics of major contagious diseases]. AB - The purpose of this article was to investigate the issues related to nursing students' clinical practicum when severe contagious diseases occur. Based on the concept that patient-benefit must come first, the authors suggested that nursing students should remain on duty even if there were an outbreak of a severe contagious disease. The authors also discussed relevant issues in terms of professional roles and the safety of the learning environment. The problems identified and the proposed strategies for solving them are outlined in this article. There is no single strategy applicable to all conditions, however. Finally, the authors urge schools and hospitals to develop effective communications and close cooperation and to remain well informed about severe contagious diseases. PMID- 15290645 TI - [An alternative in childbirth education: vaginal delivery following earlier cesarean delivery]. AB - The cesarean rate in Taiwan, at 33.10 percent, is the third highest in the world. Many women who have had a cesarean section for one birth opt to do so for all subsequent deliveries. The VBAC (vaginal birth after c-section) is defined as a vaginal birth subsequent to a cesarean birth. Many women remain skeptical about VBAC, but education can do much to dispel their doubts. Currently, the childbirth education provided by most hospitals in Taiwan is designed solely for women expecting a vaginal birth; it takes no account of those who have previously had a cesarean birth and are considering a vaginal birth. This article describes the current situation as regards VBAC, the reasons why it needs to become a more widespread practice, the necessity for childbirth education to cover the aims and nature of VBAC, and the author's experiences, to serve as a reference for caregivers providing training in VBAC in order to benefit more women. PMID- 15290646 TI - [Analyses of criminal judgments as related to nursing services in the cases of the Supreme Court in Taiwan]. AB - This paper briefly reviewed literature relating to medical disputes and investigated the criminal judgments of the Supreme Court in Taiwan in cases involving nurses and the delivery of nursing services. Past studies of criminal judgments involving medical dispute have usually been presented in the form of case studies. By contrast, this study used a retrospective data, and analyzed the judgments by content analysis and the quantitative research design method. The original written verdicts were obtained from two major online law databases. There have been a total of 70 criminal judgments related to medical disputes since 1927. Among them, 7 cases involved nurses and the delivery of nursing services. These 7 cases were analysed, and the findings will be instructive to those interested in lawsuits related to medical disputes. PMID- 15290647 TI - [Using focus groups among the elderly]. AB - Focus groups are important tools for assessing markets, making public policy and researching applied science. The use of focus groups can enable us to gather large volumes of data in short periods of time. The need for focus groups, because of the economical and efficient way in which they generate information, has grown recently in nursing. Focus groups can be used among different groups within a population, including teenagers, healthy adults, elderly people, patients and nurses. The manner in which they are used, however, must be adjusted to take account of the characteristics of each group. This article discusses the preparation, implementation, analysis and reporting of the use of focus groups among elderly people. It was important to observe the elderly subjects' physical and psychological states, work and rest habits, and interaction, and carefully arrange the detail of each group in accordance with the findings of such observation. This article is intended to enable the experience of using focus groups among the elderly to be shared with other researchers. PMID- 15290648 TI - [An assessment of the health of foreign brides in Hsin-Tien County]. AB - The numbers of foreign brides are increasing and they have become a major source of caregivers to families in Taiwan. Because of cultural and linguistic barriers, however, they may not be able to handle either their own health needs or those of the families for which they work effectively. A comprehensive assessment of their own health needs is therefore the first step to solving these problems. The purpose of this paper was to understand the health care needs of foreign brides in Hsin-Tien County, using the model of the community as partner. Data were collected by means of literature review, windshield survey, interviews, and focus group discussion. The results showed that prenatal, postpartum and baby care, and infertility were their primary health care needs. In addition, mental disability in the family, congenital disease, and chronic disease were important issues. In conclusion, community health nurses need to be concerned not only with common needs, but also with the individual health needs of foreign brides in order to provide comprehensive family health care. PMID- 15290649 TI - [Application of therapeutic play in the process of nursing a preschool patient]. AB - This report describes the application of therapeutic play to a 4-year-old female patient with intestinal obstruction during the process of nursing. The patient suffered a Port-A in the clavicle and an ileostomy in the right upper abdomen. She cried incessantly and resisted nurses who attempted to change her dressing. Data were collected by participant observation, and it was found that the patient had problems dealing with anticipatory pain and body image and lacked an emotional outlet. Therapeutic play therefore was applied during the process of nursing in an effort to accomplish the following: (1) establish trust between the patient and the health care providers; (2) reduce the patient's fear of therapy; (3) help the patient to understand her own self-image; (4) provide an emotional outlet for her. During the nursing process, we improved her overall compliance, provided her with an emotional outlet, and helped her to understand her self image. Through this nursing process we experienced the nature of children's play. Pediatric nurses may consider employing play as a means to communicate with patients in an effort to reduce their stress during hospitalization. Therapeutic play may also improve patient compliance with the process of nursing. PMID- 15290650 TI - [Conquering the fear of death in a lung cancer patient by reminiscence therapy]. AB - The purpose of this paper is to share a successful nursing experience involving a hospitalized lung cancer patient who suffered a fear of death from the terminal disease. After witnessing the process of resuscitation on a neighboring patient while hospitalized, the subject patient developed a negative mental response, refusing any treatment, developing a negative attitude to life, and retreating into total social isolation. The experience of witnessing the resuscitation induced a sense of fear of death in facing the terminal disease. Utilizing a non structural and personal reminiscence therapy, the nurse provided individualistic and humane nursing care after realizing the unique needs of the terminal patient, and was able to help the patient to diminish the feeling of fear, regain self awareness, and be well prepared for death. PMID- 15290651 TI - [The application of the resiliency model to caring for the family of a head injury client]. AB - Head injury often occurs in accidents, with severe consequences affecting not only the injured individual but his/her entire family. The purpose of this paper is to describe a case of nursing care provided to the family of a head injury patient using the family resiliency model. From June 26 to July 10, 2002, one of the authors took care of the head injury patient as a primary nurse, then assessed the family in accordance with the five factors influencing the family crisis adaptation process identified in the family resiliency model. The major stressors for the family included the unknown prognosis for the patient, the lack of family income, and not knowing how to talk with the children about their father's condition. Three nursing problems were identified, as follows: the family's knowledge deficit concerning head injury treatment, compromised family coping, disruption of family life and lack of coping resources. Applying various nursing interventions (i.e. education, family conferences, and play therapy), the authors guided the family towards an understanding of the disease treatment plan and the stresses they were facing. Finally, family members were able to participate in the rehabilitation plan for the patient and to find the resources to solve the family problems. This demonstrated that the resiliency model is a good tool for assessing families in crisis and planning nursing care in order to facilitate the family's successful bonadaptation. PMID- 15290652 TI - A powerful strategy to account for multiple testing in the context of haplotype analysis. AB - Haplotypes--that is, linear arrangements of alleles on the same chromosome that were inherited as a unit--are expected to carry important information in the context of association fine mapping of complex diseases. In consideration of a set of tightly linked markers, there is an enormous number of different marker combinations that can be analyzed. Therefore, a severe multiple-testing problem is introduced. One method to deal with this problem is Bonferroni correction by the number of combinations that are considered. Bonferroni correction is appropriate for independent tests but will result in a loss of power in the presence of linkage disequilibrium in the region. A second method is to perform simulations. It is unfortunate that most methods of haplotype analysis already require simulations to obtain an uncorrected P value for a specific marker combination. Thus, it seems that nested simulations are necessary to obtain P values that are corrected for multiple testing, which, apparently, limits the applicability of this approach because of computer running-time restrictions. Here, an algorithm is described that avoids such nested simulations. We check the validity of our approach under two disease models for haplotype analysis of family data. The true type I error rate of our algorithm corresponds to the nominal significance level. Furthermore, we observe a strong gain in power with our method to obtain the global P value, compared with the Bonferroni procedure to calculate the global P value. The method described here has been implemented in the latest update of our program FAMHAP. PMID- 15290653 TI - Integrated evaluation of DNA sequence variants of unknown clinical significance: application to BRCA1 and BRCA2. AB - Many sequence variants in predisposition genes are of uncertain clinical significance, and classification of these variants into high- or low-risk categories is an important problem in clinical genetics. Classification of such variants can be performed by direct epidemiological observations, including cosegregation with disease in families and degree of family history of the disease, or by indirect measures, including amino acid conservation, severity of amino acid change, and evidence from functional assays. In this study, we have developed an approach to the synthesis of such evidence in a multifactorial likelihood-ratio model. We applied this model to the analysis of three unclassified variants in BRCA1 and three in BRCA2. The evidence strongly suggests that two variants (C1787S in BRCA1 and D2723H in BRCA2) are deleterious, three (R841W in BRCA1 and Y42C and P655R in BRCA2) are neutral, and one (R1699Q in BRCA1) remains of uncertain significance. These results provide a demonstration of the utility of the model. PMID- 15290654 TI - The multiple colorectal adenoma phenotype and MYH, a base excision repair gene. AB - We summarize the genetic and clinical features of the colorectal adenomas and cancers that occur in MYH-associated polyposis (MAP). MAP results from biallelic germline mutations in the base excision repair gene, mutY homologue (MYH). MAP has a phenotype that is often indistinguishable from classical or attenuated familial adenomatous polyposis (FAP), but the former is inherited as a recessive condition, whereas the latter is a dominantly inherited disease caused by germline mutations of the APC gene. MYH mutations seem to act by increasing the frequency of somatic APC mutations. MAP tumors may then progress to cancer along a distinct genetic pathway. MAP occurs in several different ethnic groups, the mutation spectrum appearing to differ among groups. It remains unknown, however, as to why carriers of MYH mutations specifically develop tumors of the gastrointestinal tract. In general, carriers of biallelic MYH mutations should be treated and followed up as for FAP patients with a similar phenotype. Relatives of MAP patients should be counseled as for any other recessive condition, although it remains possible that carriers of single mutations are at a modestly increased risk of colorectal cancer. PMID- 15290655 TI - Evaluation and management of cystic pancreatic tumors: emphasis on the role of EUS FNA. AB - Cystic lesions of the pancreas are increasingly recognized and usually represent pseudocysts or cystic pancreatic tumors (CPTs), but also include congenital cysts, acquired cysts, extrapancreatic cysts, or cystic degeneration of solid tumors. It is important to distinguish CPT lesions given their varied prognosis and therapy. Mucinous varieties of CPTs (mucinous cystic neoplasms and intraductal papillary mucinous tumors) are premalignant or malignant, and surgical resection is generally recommended in good operative candidates. In contrast, nonmucinous CPTs include serous cystadenomas with a very low malignant potential, or pseudocysts, which are always benign. As a result, nonmucinous CPTs are generally resected only when inducing symptoms or complications. Review of the clinical, imaging, laboratory, and pathology information may clarify the specific tumor type. The relatively limited accuracy of any one modality requires that we consider the combined results when making management decisions. PMID- 15290656 TI - Erosive esophagitis and NERD: Can we really classify patients with heartburn by endoscopic findings? PMID- 15290657 TI - Effectiveness of proton pump inhibitors in nonerosive reflux disease. AB - BACKGROUND & AIMS: Little information is available about the efficacy of proton pump inhibitors (PPIs) in patients with nonerosive reflux disease (NERD). We aimed to synthesize available data and determine the effectiveness of PPIs on symptom resolution in patients with NERD. METHODS: A systematic review of the literature identified studies reporting the effects of PPIs in patients with NERD. Heartburn resolution data were pooled across studies. The effectiveness of PPI therapy in inducing complete heartburn resolution was compared in patients with NERD vs. erosive esophagitis (EE). RESULTS: Seven trials evaluating heartburn resolution in NERD were identified. Higher proportions of patients reported achieving sufficient heartburn resolution compared with complete heartburn resolution. The effect of PPIs on sufficient heartburn resolution was observed sooner than was complete heartburn resolution. Therapeutic gain of PPI therapy over placebo ranged from 30% to 35% for sufficient heartburn control and from 25% to 30% for complete heartburn control. Pooled response rates at 4 weeks were significantly higher for patients with EE compared with NERD (56% vs. 37%, P < 0.0001). CONCLUSIONS: PPIs provide a more modest therapeutic gain in patients with NERD as compared with those with EE. A trend in increased therapeutic gain for NERD patients was shown throughout the 4 weeks, suggesting that 4 weeks of follow-up evaluation may be insufficient to show full therapeutic gain in this patient population. PMID- 15290658 TI - Dysphagia in patients with erosive esophagitis: prevalence, severity, and response to proton pump inhibitor treatment. AB - BACKGROUND & AIMS: Dysphagia is considered an alarm symptom, raising the question of stricture or malignancy. We sought to determine the prevalence and severity of dysphagia in patients with uncomplicated erosive esophagitis and its response to therapy. METHODS: A total of 11,945 patients with endoscopically confirmed erosive esophagitis (Los Angeles grades A-D) participated in 5 double-blind, randomized, clinical trials evaluating the efficacy of up to 8 weeks of treatment with either once-daily esomeprazole 40 mg (n = 5068), esomeprazole 20 mg (n = 1243), omeprazole 20 mg (n = 3018), or lansoprazole 30 mg (n = 2616). The severity of dysphagia (4-point scale) was rated at baseline and at week 4. Esophagitis was classified as mild (grade A or B) or severe (grade C or D). RESULTS: At baseline, 4449 of 11,945 patients (37%) had dysphagia-43% of patients with severe esophagitis, and 35% of patients with mild esophagitis (odds ratio, 1.39; 95% confidence interval, 1.27-1.51, P < 0.001). Dysphagia resolved in 83% of patients after 4 weeks of proton pump inhibitor (PPI) treatment. Resolution of dysphagia was associated with a mean healing rate of 90% across all treatments. Seventeen percent of patients reported persistent dysphagia, and in these patients the healing rates were decreased significantly (mean 72%; P < 0.0001). CONCLUSIONS: Dysphagia is common in patients with erosive esophagitis but is not a reliable clinical predictor of severe erosive esophagitis. Dysphagia resolved with PPI therapy in most cases, but persistent dysphagia may indicate failed healing. PMID- 15290659 TI - Clinical application of polymerase chain reaction to diagnose Clostridium difficile in hospitalized patients with diarrhea. AB - BACKGROUND & AIMS: Clostridium difficile is a common cause of diarrhea in hospitalized patients and is associated with significant morbidity and cost. The current diagnostic standard, enzyme immunoassay (EIA), has low sensitivity, leading to duplicate testing and empiric treatment. We sought to show the usefulness and potential cost effectiveness of polymerase chain reaction (PCR) amplification of toxin B gene for diagnosis of C. difficile-induced diarrhea. METHODS: A total of 148 stool samples from academic and community-based hospitals were sent for EIA testing and were evaluated prospectively for the presence of toxin B gene by PCR. Results were compared with EIA regarding sensitivity, specificity, and predictive values. Medical charts were reviewed to determine the following: (1) number of EIAs sent per admission, (2) number sent within a 24 hour time period, and (3) how caregivers practiced based on EIA results. RESULTS: The mean age of 130 patients was 55 years. EIA and PCR were positive in 6.8% and 13.6% of patients, respectively. EIA sensitivity was 40%, specificity was 98%, and positive and negative predictive values were 80% and 91%, respectively. The cost of the PCR was $22/sample. Empiric treatment for C. difficile was given unnecessarily in 42% of EIA-negative results. Thirty percent of patients had 3 or more EIAs sent during their hospital admission. Of patients with multiple samples sent, 57% had more than 1 sample sent in a 24-hour period. CONCLUSIONS: Many physicians do not conform to practice guidelines regarding recommended diagnosis and empiric treatment of C. difficile. Toxin B gene PCR represents a more sensitive and potentially cost-effective method to diagnose C. difficile-induced diarrhea than EIA and should be considered for use as an alternative diagnostic standard. PMID- 15290660 TI - Effect of alosetron on bowel urgency and global symptoms in women with severe, diarrhea-predominant irritable bowel syndrome: analysis of two controlled trials. AB - BACKGROUND & AIMS: The aim of this study was to assess the effect of alosetron on bowel urgency and irritable bowel syndrome (IBS) global improvement in diarrhea predominant IBS (D-IBS). METHODS: Women with a lack of satisfactory bowel urgency control at least 50% of the time during screening were randomized to receive alosetron 1 mg (n = 246) or placebo (n = 246) twice daily. The primary end point was the percentage of days with satisfactory control of bowel urgency. The response rate for the IBS global improvement scale (GIS) was a secondary end point. GIS responders were patients who recorded either moderate or substantial improvement in IBS symptoms relative to the way they felt before entering the study. Other end points included improvement in stool frequency, stool consistency, and percentage of days with incomplete evacuation. Further analyses were performed on a subset of patients who had at least 10 of 14 days during screening (>/=71% of days) with a lack of satisfactory control of bowel urgency. RESULTS: Patients had severe chronic IBS symptoms, and 89% of patients had D-IBS. Alosetron resulted in a greater percentage of days with satisfactory control of urgency compared with placebo (69% vs. 56%, respectively, P < 0.001). Greater percentages of alosetron-treated patients were GIS responders at 4, 8, and 12 weeks compared with placebo (59% vs. 41%, 63% vs. 41%, and 68% vs. 46%, respectively, P < 0.001). Patients with more frequent urgency had similar results. Constipation occurred in 28% and 9% of subjects in the alosetron- and placebo-treated groups, respectively. No cases of ischemic colitis were reported. CONCLUSIONS: Alosetron effectively manages bowel urgency and improves global symptoms in women with severe chronic D-IBS. PMID- 15290661 TI - Fecal excretion of human deoxyribonucleic acid as an index of inflammatory activity in ulcerative colitis. AB - BACKGROUND & AIMS: Several indices evaluate disease activity in ulcerative colitis, but most have drawbacks to their application (invasiveness, complexity, or lack of specificity), and discrepancies between them are frequent. Assuming that desquamation of epithelial and inflammatory cells increases in damaged colonic mucosa, we hypothesized that fecal excretion of human DNA is an index of mucosal inflammation and damage. The aim of our study was to determine whether excretion of human DNA is useful in the evaluation of disease activity in ulcerative colitis. METHODS: Thirty-one controls and 36 ulcerative colitis patients were included. Ulcerative colitis patients and controls underwent colonoscopic examination after preparation by gut lavage with polyethylene-glycol electrolyte solution. In patients, disease activity was established using the clinical index of Rachmilewitz and an endoscopic score. Feces and gut lavage fluid were obtained and DNA levels were measured by quantitative polymerase chain reaction of the human beta-globin gene. RESULTS: Fecal DNA excretion correlated with the clinical index (r = 0.59, P < 0.05) and the endoscopic score (r = 0.76, P < 0.01). Gut lavage fluid DNA levels also correlated with clinical and endoscopic activity scores (r = 0.41 and 0.51, respectively, P < 0.05). Fecal DNA excretion was significantly higher in patients with endoscopically or clinically active disease than in controls or patients in remission. Fecal DNA excretion discriminates between endoscopically active disease and remission (sensitivity 0.67, specificity 1.00, P < 0.01). CONCLUSIONS: Excretion of human DNA in feces, as an expression of cellular desquamation, is a novel noninvasive technique to objectively assess disease activity in ulcerative colitis. PMID- 15290662 TI - Chronic granulomatous disease caused by a deficiency in p47(phox) mimicking Crohn's disease. AB - We describe 2 cases of autosomal recessive chronic granulomatous disease (CGD) in 2 sisters presenting with a picture consistent with inflammatory bowel disease. The index case is a 10-year-old girl with a history of refractory Crohn's colitis treated with aggressive immunosuppressive therapy whose course subsequently was complicated by central nervous system aspergillosis. Additional evaluation showed a diagnosis of CGD, an underlying immunodeficiency in which phagocytes fail to produce microbicidal reactive oxygen intermediates because of inherited defects in the reduced form of nicotinamide-adenine phosphate dinucleotide (NADPH) oxidase. The diagnosis of a typically X-linked inherited disease in our female patient suggested that she had 1 of the 3 less common autosomal recessive forms of the disease. This was confirmed by studies showing the absence of the p47(phox) subunit of NADPH oxidase in her neutrophils and the presence of a homozygous dinucleotide deletion in the neutrophil cytosolic factor 1 gene that encodes p47(phox). Additional analyses of members of the patient's immediate family showed the same homozygous mutation in 2 siblings, 1 of whom also developed chronic colitis consistent with a diagnosis of Crohn's disease. These 2 cases emphasize the importance of high clinical suspicion for an alternative diagnosis of immune deficiency in the setting of presumed inflammatory bowel disease and opportunistic infection. PMID- 15290663 TI - Lactose intolerance associated with adjuvant 5-fluorouracil-based chemotherapy for colorectal cancer. AB - BACKGROUND & AIMS: Bowel mucosal injury associated with 5-fluorouracil (5-FU) treatment might result in secondary lactose intolerance. The frequency and clinical significance of 5-FU-related hypolactasia are unknown. METHODS: One hundred fifty patients randomly assigned to receive 1 of 2 adjuvant 5-FU-based chemotherapy regimens, the Mayo regimen or the simplified de Gramont regimen, were studied for lactose tolerance by using an oral lactose absorption test, a symptom questionnaire, treatment-related toxicity, and Subjective Global Assessment of Nutritional Status questionnaire before, during, and 2 and 6 months after chemotherapy for colorectal cancer. RESULTS: The frequency of hypolactasia increased from 24% before treatment to 35% during treatment (P < 0.0001). Therapy related hypolactasia was reversible on discontinuation of chemotherapy. Symptoms compatible with lactose intolerance occurred in 94% of patients with an abnormal lactose absorption test result during chemotherapy. The frequency of hypolactasia increased during chemotherapy in both treatment groups, but was detected more commonly in those for whom therapy included continuous 5-FU infusions (the de Gramont regimen; 45% vs. 25%; P = 0.006). The presence of hypolactasia during chemotherapy was associated with flatulence, diarrhea, and poor nutritional status. CONCLUSIONS: Reversible chemotherapy-related hypolactasia and lactose intolerance are not infrequent in patients treated with 5-FU-based adjuvant chemotherapy for colorectal cancer. Avoidance of lactose during chemotherapy may improve treatment tolerability in these patients. PMID- 15290664 TI - A prospective study of genetic polymorphisms in the cytochrome P-450 2C9 enzyme and the risk for distal colorectal adenoma. AB - BACKGROUND & AIMS: Regular aspirin use is associated with a reduced risk for colorectal adenoma, whereas smoking increases risk. The cytochrome P-450 (CYP) 2C9 (CYP2C9) enzyme is involved in the metabolism of several drugs, including possibly aspirin, and such carcinogens as smoking-related polycyclic aromatic hydrocarbons. Genetic variation in this enzyme may modulate the influence of aspirin and smoking on adenoma risk. METHODS: We examined the risk for incident distal colorectal adenoma according to CYP2C9 genotype, aspirin use, and smoking in a prospective nested case-control study of women. RESULTS: Among 394 cases and 396 controls, women with at least 1 variant CYP2C9 allele had a significantly greater risk for adenoma (multivariate odds ratio [OR], 1.56; 95% confidence interval [CI], 1.13-2.15; P = 0.007). Although women who used aspirin regularly (>/=2 standard tablets/wk) experienced a lower risk for adenoma compared with non regular users, the effect was similar irrespective of genotype. Women who smoked >20 pack-years had an OR of adenoma of 1.50 (95% CI, 1.07-2.12; P = 0.02) compared with those who smoked 20 pack-years, the OR of adenoma was 2.50 (95% CI, 1.44-4.38; P = 0.001) compared with women with no variant alleles who smoked 1 (P < 0.0001), >/=3 pancreatic injections (P < 0.00001), and precut sphincterotomy (P = 0.01) were significantly associated with post-ERCP pancreatitis. At multiple logistic regression analysis, >/=3 pancreatic injections (odds ratio [OR], 1.95; 95% confidence interval [CI], 1.45-2.63) and a Freeman score >1 (OR, 1.47; 95% CI, 1.11-1.94) retained their predictive power. CONCLUSIONS: Long-term (6.5-hr) administration of either somatostatin or gabexate mesylate is ineffective for the prevention of post-ERCP pancreatitis. Pancreatic injury seems to be related to difficulty in common bile duct access. PMID- 15290666 TI - Predicting outcome after cardiac surgery in patients with cirrhosis: a comparison of Child-Pugh and MELD scores. AB - BACKGROUND & AIMS: This study aims to quantify the risk of cardiac surgery in patients with cirrhosis. METHODS: Records of all adult patients with cirrhosis undergoing cardiac surgery using cardiopulmonary bypass at the Cleveland Clinic (Cleveland, OH) from January 1992 to June 2002 were analyzed for any relationship of Child-Pugh class and/or score and Model for End-Stage Liver Disease (MELD) score with outcome measures of hepatic decompensation and death during the first 3 months after surgery. RESULTS: Forty-four patients underwent coronary artery bypass grafting (16 patients), valve surgery (16 patients), a combination of the 2 procedures (10 patients), or pericardiectomy (2 patients). Twelve patients (27%) developed hepatic decompensation, and 7 patients (16%) died. Proportions of hepatic decompensation were 3 of 31, 8 of 12, and 1 of 1 patients, and death, 1 of 31, 5 of 12, and 1 of 1 patients in Child-Pugh classes A, B, and C, respectively. The association of hepatic decompensation and mortality with Child Pugh class, Child-Pugh score, and MELD score was significant (P < 0.005). Areas under the receiver operating characteristic curves for mortality were similar for Child-Pugh (0.84 +/- 0.09) and MELD scores (0.87 +/- 0.09). A cutoff Child-Pugh score >7 was found to have a sensitivity and specificity of 86% and 92% for mortality, with a negative predictive value of 97% (95% confidence interval [CI], 83-99) and positive predictive value of 67% (95% CI, 31-91), respectively. However, a similar cutoff value for MELD score could not be established. CONCLUSIONS: Child-Pugh score and/or class and MELD score are significantly associated with hepatic decompensation and mortality after cardiac surgery using cardiopulmonary bypass in patients with cirrhosis. Such surgery can be conducted safely in patients with a Child-Pugh score /=8 have a significant risk for mortality. PMID- 15290668 TI - An unusual cause of hematemesis. PMID- 15290669 TI - Chondroid tumors of the larynx: a clinicopathologic study of 19 cases, including two dedifferentiated chondrosarcomas. AB - We studied 19 cases of chondroid tumors of the larynx: two chondrometaplastic nodules, two chondromas and 15 chondrosarcomas (including two dedifferentiated chondrosarcomas). One of chondromas recurred 18 months after resection because of inadequate surgical treatment. Chondrosarcomas were separated as low-grade (nine cases), intermediate (three cases), high-grade (one case), and dedifferentiated (two cases) according to histologic appearance. Chondrosarcomas are nearly always histologically low grade, make up the largest numbers of the neoplasms, and arise from the cricoid cartilage. Conservative surgical management, when possible, is advocated to preserve the larynx. Chondrometaplastic nodules are to be distinguished from chondrosarcoma and the rarely occurring chondroma. The nodules are not neoplastic and have a low to nil recurrent potential. PMID- 15290671 TI - Management of melanotic neuroectodermal tumor of infancy. AB - Melanotic neuroectodermal tumor of infancy is a rare congenital neoplasm involving the head and neck in young patients. The clinical assessment, histologic diagnosis, and management is reviewed, with an emphasis on different treatment alternatives in two new case reports. PMID- 15290670 TI - Chest wall resection for invasive lung carcinoma, soft tissue sarcoma, and other types of malignancy. Pathologic aspects in a series of 107 patients. AB - With improvements in surgical techniques for resection and reconstruction of the chest wall, pathologists are confronted with complicated surgical specimens. There are no currently available guidelines specifically dedicated to the handling of these specimens. Extended resections of lung carcinoma chest wall invasions may change the clinical value of some TNM subsets. We reviewed a series of 107 consecutive malignant tumors involving the chest wall and resected in our institution during a 3-year period. The 107 patients included 39 females and 68 males aged 6 to 80 years (mean, 53 years). Ninety-eight cases (92%) were en bloc resection. There were 55 invasions by lung carcinomas including 19 Pancoast tumors. With the current TNM classification, five lung carcinomas, treated with vertebral body resection because of vertebral foramina invasion, were T3. Four lung carcinomas were N3 or M1 only because of supraclavicular or chest wall lymph node invasion. Other tumors included 20 primary soft-tissue tumors, 13 primary skeletal tumors, 12 metastases, four local invasions by breast tumors, and three miscellaneous lesions. Resected structures included one to six ribs (mean, 2.6; n = 89), thoracic inlet (n = 24), three or four vertebral bodies (n = 13), sternum (n = 17), clavicles (n = 15), shoulder blade (n = 4), upper limb (n = 2), skin (n = 29), lung (n = 64), diaphragm (n = 2), and mediastinum (n = 2). Ten cases were incomplete resections including five because of vertebral body or vertebral foramina tumor invasion. The study of surgical specimens resulting from resection of malignant tumors of the chest wall is complicated because of the variety of both tumor histologic types and involved anatomic structures. Specimen radiograms have a great informative value. Assessment of surgical margins, especially vertebral foramina, is imperative. In lung carcinomas invading the chest wall, we suggest that vertebral foramina invasion could be classified T4 and that the prognostic value of chest wall lymph nodes isolated invasions should be assessed for a possible N1 classification. PMID- 15290672 TI - Multicystic endometrial stromal sarcoma. AB - Endometrial stromal sarcoma usually has the gross appearance of a single nodule, multiple masses, or a poorly demarcated lesion with occasional cystic degeneration; however, a multilocular form has not been described in the literature. We report the case of a 25-year-old woman with a cystic multilocular lesion with thin septae measuring 8 cm, discovered by a pelvic ultrasonography. Grossly, it was a multicystic mass located in uterine fundus that was attached to myometrium and showed infiltrating borders. We propose that cystic endometrial stromal sarcoma should be included in the differential diagnosis of cystic uterine tumors. PMID- 15290673 TI - Cerebellar paraganglioma. AB - Paragangliomas arising as primary neoplasms in the intracranial portion of the central nervous system are relatively uncommon and only have been rarely reported. We report a case of apparent primary paraganglioma arising in the left cerebellar hemisphere in an 11-year-old girl who presented with a 6-month history of headaches. The tumor was marked by a nested architectural pattern (zellballen), delimited by S-100-positive sustentacular cells. Morphologic and immunohistochemical features of the neoplasm are discussed and literature is reviewed regarding involvement of the central nervous system by paraganglioma. PMID- 15290674 TI - Sebaceous carcinoma ex-pleomorphic adenoma: a rare phenotypic occurrence. AB - Primary sebaceous carcinoma of salivary glands is a rare entity with approximately 22 de novo documented cases. Similar tumor arising in a benign mixed tumor has only been reported once. We report a second case of sebaceous carcinoma in a pleomorphic adenoma and discuss the clinicopathologic features, histogenesis, and the differential diagnosis of this unusual tumor. PMID- 15290675 TI - Sclerosing extramedullary hematopoietic tumor involving lesser omentum and ligamentumteres in liver explant. AB - A 47-year-old patient who presented with a 2-year history of hepatitis B virus associated liver cirrhosis underwent a liver transplantation. The explanted liver was grossly enlarged and diffusely cirrhotic. The lesser omentum and ligamentum teres revealed a 4 cm ill-defined mass that, on sectioning, showed a gray-white appearance. Extramedullary hematopoiesis on a fibrosclerotic background constituted this mass, but there was also extramedullary hematopoiesis within the liver cords and possibly within the omentum. A bone marrow study also showed chronic idiopathic myelofibrosis. Acute rejection ensued despite immunosuppression and the patient continued to deteriorate and expired. Sclerotic extramedullary hematopoietic tumor is rare, has an unfavorable prognosis, and may mark the end stage of chronic myeloproliferative disorder. PMID- 15290676 TI - Primary synovial sarcoma of the parotid gland. AB - Synovial sarcoma typically develops in the deep soft tissues of the extremities in adolescents and young adults. Uncommonly, these tumors may arise (9%) in the head and neck region (9%), especially at the cervical and parapharyngeal sites. Primary synovial sarcoma of the parotid gland is a rare occurrence that may not uncommonly cause differential diagnostic difficulties. In these cases, an origin from within the gland rather than a secondary involvement by tumor from a surrounding structure must be confirmed. We report a new case of biphasic synovial sarcoma arising in the parotid gland and review the previously documented cases. PMID- 15290677 TI - Primary paraganglioma of the lung. AB - There are few reported cases of primary pulmonary paraganglioma in the pathology literature. Given the historical confusion surrounding bronchial tumors, widespread use of the term "chemodectoma" and classification of these lesions as paraganglioma in an outdated World Health Organization classification of lung tumors, the recognition of tumors arising from paraganglia within the lung has not been accepted by leading authorities. We present a well-documented case of a primary pulmonary paraganglioma with typical morphologic features and a supporting immunohistochemical profile. The 0.9 cm endobronchial tumor was submucosal and composed of nests of ovoid cells with abundant eosinophilic cytoplasm, cytoplasmic vacuoles, round to oval nuclei with speckled chromatin, and occasional conspicuous nucleoli. The nests of cells were surrounded by thin walled vascular channels and stellate spindle cells. The ovoid cells showed strong diffuse staining for chromogranin A, synaptophysin, and faint staining for S-100; they were negative for cytokeratin AE1/AE3, Cam 5.2, and epithelial membrane antigen. The stellate spindle cells stained intensely positive for S-100 protein. A critical review of reported cases of pulmonary chemodectomas and paragangliomas in the English literature features few, if any, well-documented examples. While this exceedingly rare tumor should be discerned from carcinoid tumor, it remains unknown if primary pulmonary paragangliomas behave aggressively like intra-abdominal extra-adrenal paragangliomas, or in a more indolent manner observed with extra-adrenal paragangliomas in other locations. PMID- 15290678 TI - Nonspecific (idiopathic) granulomatous prostatitis associated with low-grade prostatic adenocarcinoma. AB - Nonspecific granulomatous prostatitis (NSGP) is uncommon and may simulate carcinoma both clinically and microscopically. Concurrent NSGP and prostatic adenocarcinoma is rare. To our knowledge this association has been documented once and it was only rarely mentioned in two large series of NSGP. We describe a 67-year-old man who presented with a history of prostatism of 1 month's duration. Suprapubic prostatectomy revealed NSGP associated with nodular hyperplasia and low-grade prostatic adenocarcinoma. The pathologist should be aware of the rare association of NSGP and prostatic adenocarcinoma. Wide sampling of the prostatectomy specimens with NSGP is mandatory to exclude an occult prostatic adenocarcinoma. PMID- 15290679 TI - Polyostotic giant enostoses with strongly positive radionuclide bone scan. AB - For nearly three quarters of a century bone islands (enostoses) were considered scintigraphically inactive, hence, easily differentiated radiologically from clinically more significant primary or metastatic intraosseous lesions. However, enostoses' clinical significance has changed considerably since the first report of a case with increased radioactive uptake on bone scan in 1976. Consequently, any radiotracer-positive skeletal lesion, regardless of radiographic appearance and clinical presentation, is now generally viewed with some caution. A 23-year old woman presented with polyostotic enostoses discovered incidentally during pelvic radiographic examination. Both scintiscan and skeletal survey identified one or three fairly large densely radiopaque lesions in many bones, the largest measuring 7 x 4 cm. Except for a few departures from characteristic radiologic and scintigraphic changes, such as lesional border alterations and considerable increase in size and number of lesions involving many bones (polyostotic), the clinical findings and radiographic appearance of every bone island seem typical. Absent definitive roentgenologic diagnosis, both needle and open biopsies were performed. We found no previous report of a polyostotic enostoses with or without positive radionuclide bone scans reported in the literature. Distinctions from osteopoikilosis and osteopathia striata are briefly discussed. PMID- 15290680 TI - Brain in the middle ear or nasal cavity: heterotopia or encephalocele? AB - Brain in the middle ear or nasal cavity: heterotopia or encephalocele? The answer to this question is greatly influenced by clinical information. Sometimes, however, this information is insufficient and the pathologist's opinion may influence patient management. It seems that most pathologists tend to get the answer wrong. This article will enable you to arrive at the correct answer. PMID- 15290681 TI - Novel approach for the introduction of a C-1 oxygenated group on the decalin skeleton: first asymmetric total synthesis of 1S,6S-dihydroxy-eudesm-3-ene. AB - This paper describes a novel approach for the introduction of a C-1 hydroxyl on the decalin ring system, starting from (-)-carvone. Utilizing the substrate controlled Mukaiyama aldol reaction and alkaline cyclization as key steps, the C 1 oxygenated decalin eudesmane skeleton 2' and its four isomers were synthesized efficiently. Furthermore, X-ray structural analysis showed that the claimed structure for the natural product is incorrect. PMID- 15290682 TI - Effect of intramolecular interactions on circular dichroism of ortho-substituted 1-phenethylamines. AB - The synthesis of ortho-substituted (S)-1-phenethylamines via ortho-lithiation of its N,N-dimethyl derivative followed by reactions with different reagents is described. Circular dichroism spectra of synthesized compounds were measured. It is shown that the observed short-wave Cotton effects greatly depend on the existence of cyclic structure due to possible interactions such as hydrogen bond or electrostatic interaction. PMID- 15290683 TI - Memory effect of diethylamine mobile phase additive on chiral separations on polysaccharide stationary phases. AB - The existence of a memory effect for amine additives on polysaccharide chiral stationary phases has often been suggested, but not clearly demonstrated. Demonstration of this effect is made difficult by the uncertainty as to which analytes benefit from use of amine additives and, typically, an unclear history of column use. In this work, analytes were selected for differences in their behavior with and without additives. Columns were used with no prior history. A persistent memory effect was demonstrated on a CHIRALPAK AD-H column in hexane based mobile phases. This effect was short-lived, with polar organic mobile phases. Memory was short-lived on a CHIRALCEL OJ-H column. Flushing with isopropanol was shown to remove most of the memory effect. Compounds expected to require amine additives on CHIRALCEL OD-H column did not. Acid treatment of the AD-H and OD-H columns changed their performance, which was subsequently restored by the incorporation of amine. PMID- 15290684 TI - Online monitoring of preferential crystallization of enantiomers. AB - Polarimetry is used for continuous online monitoring of optical resolution by preferential crystallization. In combination with refractometry the liquid phase composition is determined, allowing one to follow the resolution progress quantitatively. The measurement techniques were calibrated up to relatively high solution concentrations and combined with the crystallizer. The resolution of DL threonine was performed by preferential crystallization experiments in aqueous solution varying several process parameters like supersaturation, seed amount, initial enantiomeric excess, and scale. The resolution progress can be conveniently described by profiles of the optical rotation (polarimetric signal) and the crystallization pathway in the corresponding ternary phase diagram. The method outlined is applicable for dynamic process optimization and control purposes in "quasi-continuous" chiral separation processes. PMID- 15290685 TI - Determination of binding constants of cyclodextrin inclusion complexes with amino acids and dipeptides by potentiometric titration. AB - Cyclodextrins are well known for their ability to separate enantiomers of drugs, natural products, and other chiral substances using HPLC, GC, or CE. The resolution of the enantiomers is due to the formation of diastereomeric complexes between the cyclodextrin and the pairs of enantiomers. The aim of this study was to determine the binding constants of the complexes between alpha- and beta cyclodextrin and the enantiomers of a series of aliphatic and aromatic amino acids, and dipeptides, using a potentiometric titration method. The results of this method are compared to other methods, and correlated to findings in cyclodextrin-modified capillary electrophoresis and possible complex structures. Potentiometric titration was found to be an appropriate tool to determine the binding constants of cyclodextrin inclusion complexes. PMID- 15290686 TI - Dirhodium-catalyzed enantioselective C-H insertion of N-(2-benzyloxyethyl)-N (tert-butyl)diazoacetamide and its application for the synthesis of chiral GABOB. AB - Rh(2)(4S-MEOX)(4) and ethereal solvent are the best catalytic system for the enantioselective intramolecular C-H insertion of N-(2-benzyloxyethyl)-N-(tert butyl)diazoacetamide 2. The highest enantiomeric excess obtained was 91%. A new route for the asymmetric synthesis of gamma-amino-beta-hydroxybutyric acid (GABOB) has been developed. PMID- 15290687 TI - Enantioselective renal excretion of albendazole metabolites in patients with neurocysticercosis. AB - The present study investigates the urinary excretion of the enantiomers of (+)- and (-)-albendazole sulfoxide (ASOX) and albendazole sulfone (ASON) in 12 patients with neurocysticercosis treated with albendazole for 8 days (7.5 mg/kg/12 h). Serial blood samples (0-12 h) and urine (three periods of 8 h) were collected after administration of the last dose of albendazole. Plasma and urine (+)-ASOX, (-)-ASOX, and ASON metabolites were determined by HPLC using a chiral phase column (Chiralpak AD) with fluorescence detection. The pharmacokinetic parameters (P < 0.05) for (+)-ASOX, (-)-ASOX, and ASON metabolites are reported as means (95% CI); amount excreted (Ae) = 3.19 (1.53-4.85) vs. 0.72 (0.41-1.04) vs. 0.08 (0.03-0.13) mg; plasma concentration-time area under the curve, AUC(0 24) = 3.56 (0.93-6.18) vs. 0.60 (0.12-1.08) vs. 0.38 (0.20-0.55) microg x h/ml, and renal clearance Cl(R) = 1.20 (0.66-1.73) vs. 2.72 (0.39-5.05) vs. 0.25 (0.13 0.37) l/h. Sulfone formation capacity, expressed as the Ae ratio ASON/ASOX + ASON, was 2.21 (1.43-2.99). These data point to enantioselectivity in the renal excretion of ASOX as a complementary mechanism to the metabolism responsible for the plasma accumulation of (+)-ASOX. The results also suggest that the metabolite ASON is partially eliminated as a reaction product of the subsequent metabolism. PMID- 15290688 TI - New chiral selectors: design and synthesis of 6-TBDMS-2,3-methyl beta cyclodextrin 2-2' thioureido dimer and 6-TBDMS-2,3-methyl (or 2-methyl-3-acetyl) beta-cyclodextrin bearing an (R) Mosher acid moiety. AB - Cyclodextrin (CD) derivatives are important selectors for analytical chiral recognition. Their enantioselectivities and chemical properties depend on ring size and on nature, number and location of substituents. This paper describes the synthesis of 6-O-TBDMS-2,3-O-methyl beta-cyclodextrins bearing in position 2 either a single (R)-Mosher acid moiety or a second CD unit, in view of their possible application as chiral selectors. Most synthetic steps were successfully carried out under high-intensity ultrasound using a new sonochemical reactor developed in the authors' laboratory. 6-O-TBDMS-2-O-methyl-3-[(S)-2-methylbutyl] beta-CD was also synthesized and tested with gas chromatography; the enantiorecognition power of the other CD derivatives is also being tested. A computational study of model structures to design these CD derivatives. PMID- 15290689 TI - Diastereotopic analysis of mesoridazine besylate (Serentil). AB - NMR studies were conducted with the aim of determining the diastereoisomeric ratio of a commercially supplied sample of mesoridazine (MES) and to compare the results with a freshly synthesised sample of MES. The results indicated that the commercially supplied MES consisted almost entirely of one diastereoisomeric pair, which was in agreement with previous findings reported by Eap et al. The synthesised sample of MES was analysed by NMR in two stages: 1) as the initial product isolated as the free base from the direct synthesis, and 2) as the free base isolated from the crystallised besylate salt of the synthetic product. The NMR results show that the initial synthetic product consisted of two equal pairs of diastereoisomers. The diastereoisomeric pairs were further separated by the addition of the chiral shift reagent (R)-(-)-N-(3,5 dinitrobenzoyl)-alpha benzylamine to reveal equal quantities of all four enantiomers, clearly observed at the methyl sulfoxide proton peak of the NMR scan. The sample obtained from the crystallisation of MES besylate, however, indicated a significant difference, with a diastereoisomeric ratio of 75:25. The results suggest that MES besylate undergoes preferential crystallisation of one pair of diastereoisomers, with the other pair remaining in solution. PMID- 15290690 TI - Physicochemical properties, binary and ternary phase diagrams of ketoprofen. AB - Compared to simulated moving bed (SMB) chromatography, fractional crystallization is a simple and economical method for enantioseparation. Therefore, the coupling of SMB chromatography and fractional crystallization is suggested for enantioseparation of racemic compounds. In this work, a nonsteroidal antiinflammatory drug, ketoprofen (Kp), was chosen to be studied. Kp was verified as a racemic compound by FTIR, PXRD, and thermodynamic calculations. To derive the condition where pure (S)-Kp could be crystallized from a solution, which was previously enantiomerically enriched, the binary melting phase diagram and the ternary solubility phase diagram in the mixed solvent of ethanol and water over a temperature range of 15-30 degrees C were obtained. From these phase diagrams the eutectic point was determined as 91.6% mole percent (S)-Kp from the binary phase diagram and 91% from the ternary phase diagram. The results may provide valuable experiment data for the possibility of coupling fractional crystallization with SMB for Kp separation. PMID- 15290691 TI - Determination of the absolute configuration of Anisotome irregular diterpenes: application of CD and NMR methods. AB - Several Anisotome diterpene derivatives were synthesized in an attempt to obtain a crystalline compound for X-ray analysis. Although we were unable to obtain a suitable crystal, the absolute configuration of the irregular diterpene skeleton was determined using two other techniques: a circular dichroism (CD) protocol based on a tetraarylporphyrin molecular tweezer that allowed prediction of the absolute stereochemistry on a microscale level, and a method employing differences in NMR shifts from derivatization of the naturally occurring acid 1 with enantiomers of a phenylglycine methyl ester (PGME) chiral anisotropic reagent. The excellent agreement between the CD and NMR methods led to the assignment of a 2S-absolute configuration for anisotomenoic acid 1. PMID- 15290692 TI - Stereochemical studies of chiral resolving agents, M9PP and H9PP acids. AB - The stereoselective Grignard reaction of (1R,2S,5R)-(-)-2-isopropyl-5 methylcyclohexyl pyruvate (menthyl pyruvate) with 9-phenanthrylmagnesium bromide yielded diastereomeric hydroxy-esters, where intramolecular OH em leader O=C hydrogen bond was observed in IR and (1)H NMR spectra. The alkaline hydrolysis of the major product gave (+)-2-hydroxy-2-(9-phenanthryl)propionic acid (H9PP acid (3)), whose absolute configuration was assigned as S based on the chemical correlation with (1R,2S,5R)-2-isopropyl-5-methylcyclohexyl ester of (S)-2-methoxy 2-(9-phenanthryl)propionic acid (M9PP acid (2)); the absolute configuration of 2 had been previously established by X-ray crystallography. The enantioresolution of (+/-)-6-methyl-5-hepten-2-ol, sulcatol, an insect pheromone, was carried out using (S)-(+)-M9PP acid 2. PMID- 15290693 TI - Personal disease management in dementia care. PMID- 15290694 TI - Dementia Care Mapping reconsidered: exploring the reliability and validity of the observational tool. AB - BACKGROUND: Dementia Care Mapping (DCM) is a widely used observational method for evaluating the service quality provided to people with dementia. However, there is little evidence concerning its reliability and validity when used by routine care staff for whom it was designed. METHOD: The study evaluated levels of inter observer agreement; The ability of the five-minute time frame to reflect the 'actual passing of time'; And the nature of the relationship between individual Well/Ill-Being values (WIB) and dependency levels. Data collected using DCM and continuous time sampling (CTS) were compared. The methods were used in parallel where the CTS coder and the DCM mapper(s) observed the same participants. Observations were carried out with 64 people with dementia within a day hospital and a continuing care ward. Inter-observer agreement was calculated across 20 participants. Dependency levels were measured using the Clifton Assessment Procedure for the Elderly (CAPE) (Pattie and Gilleard, 1979). RESULTS: Low levels of inter-observer agreement were found where 11 of the 25 Behaviour Category codes and all six Well/Ill-being Codes produced unacceptable kappas (<0.6). The Behaviour coding frame provided a meaningful picture of activities participants engaged in, but significantly underestimated participant levels of inactivity.A strong relationship was demonstrated between participants' WIB score and levels of dependency, thus DCM was unable to measure well-/ill-being as a separate construct from participants' levels of dependency. CONCLUSIONS: Questions were raised regarding the reliability and validity of DCM as used by routine care staff. Possible reasons for this, and suggestions for amendments are made. PMID- 15290695 TI - Smell identification test as an indicator for cognitive impairment in Alzheimer's disease. AB - OBJECTIVES: The aim of the present study was to assess olfactory dysfunction in patients with Alzheimer's disease (AD) and to compare utility of the olfactory tests as possible clinical markers. METHODS: Two olfactory identification tests (The Cross-Cultural Smell Identification Test [CC-SIT] and the Picture-based Smell Identification Test [P-SIT]) and the Mini Mental State Examination (MMSE) were administered to patients with AD and age-matched controls. Apolipoprotein E (Apo E) genotypes of patients with AD were identified. RESULTS: Patients with AD had significantly lower olfactory identification scores than age-matched non demented elderly subjects in both olfactory assessments. In the AD group, the coefficient of correlation between the MMSE scores and the P-SIT scores was higher than that between the MMSE scores and the CC-SIT scores. Receiver operating curve (ROC) analyses for both tests indicated that the P-SIT discriminated AD patients from controls more reliably than did the CC-SIT. Within AD patients, those who were carrying one or two ApoE epsilon4 alleles had a higher coefficient of correlation between the MMSE scores and the P-SIT scores than patients without the ApoE epsilon4 allele. CONCLUSIONS: The results suggest that a short and simple non-lexical olfactory identification test can be useful as a clinical marker of AD appropriate for Japanese elderly population. PMID- 15290696 TI - The prevalence of depressive symptoms among elderly Chinese private nursing home residents in Hong Kong. AB - BACKGROUND: Privately-owned Nursing homes (PNH) in Hong Kong present a unique setting of institutional care where elderly with the whole spectrum of health status live together. OBJECTIVES: This cross-sectional descriptive study aimed to determine the prevalence of significant depressive symptoms in a group of Cantonese-speaking Chinese private nursing home elderly living in Hong Kong, and to identify associated psychosocial and health factors. METHODOLOGICAL RESULTS: Two hundred and forty five residents fulfilled the inclusion criteria. Using the Chinese version of the Geriatric Depression Scale--Short Form (GDS-SF), we detected significant depressive symptoms in 29% of subjects. Univariate analysis revealed some associated socio-economic risk factors including current non Comprehensive Social Security Assistance (CSSA) recipients, education levels and low abilities for social activities. Low vision, swallowing difficulties and low levels of basic activities of daily living (BADL) as reflected by the total Modified Barthel Index of less than 61 were important health predictors. Depression was also associated with features of self-perception of financial inadequacy, life dissatisfaction, poor self-perceived health, poor attitudes towards living arrangement and suicidal thoughts. Stepwise logistic regression identified swallowing problems, current non-CSSA recipient and low BADL ability as independent risk factors. CONCLUSION: The high prevalence of depressive symptoms in the nursing home elderly requires the attention of Government authorities, health care and social service providers. PMID- 15290697 TI - Dementia, pain, depression, behavioral disturbances, and ADLs: toward a comprehensive conceptualization of quality of life in long-term care. AB - OBJECTIVES: Quality of life in long-term care settings is a multidimensional construct that includes functional, cognitive, behavioral, and psychological variables. Quality of life variables have been found to be related to one another, but directional influences have not been tested. METHODS: The purpose of this study was to develop and compare two competing path models composed of quality of life variables, including dementia, pain, behavioral disturbances, and ADLs. RESULTS: Path analytic results revealed that cognitive, emotional, and behavioral variables interact with one another to predict patients' activities of daily living. Pain levels did not influence activities of daily living directly, but rather influenced behavioral disturbances and depression, which in turn influenced activities of daily living. CONCLUSIONS: These preliminary findings suggest that in order to assist long-term care residents in improving their activities of daily living, decreasing pain is likely to yield the greatest overall improvements. Future research on the relationships between quality of life variables is recommended to further develop multidimensional treatment models for healthcare providers in long-term care. PMID- 15290698 TI - EEG and the Test for the Early Detection of Dementia with Discrimination from Depression (TE4D): a validation study. AB - OBJECTIVE: The Test for the Early Detection of Dementia with Discrimination from Depression (TE4D) was developed as a screening instrument for mild dementia. We investigated the convergent validity of the TE4D to EEG and other psychometric tests in patients suffering from dementia and depression. METHOD: In 47 patients suffering from Alzheimer's disease (ICD-10 F.00) and 16 patients with affective disorders (F30-F39) the tests TE4D, ADAS-cog, SKT, BCRS, MMSE were performed and an EEG recorded. Group differences were compared by t-tests and a regression analysis was calculated. RESULTS: The inter-test-correlations varied between rs = 0.77 and rs = 0.91. Significant differences between the diagnostic groups were found for all tests as well as for the frequency bands alpha and beta. For the qEEG, significant positive correlations were found between TE4D (Dementia subscore) and the mean frequency (r = 0.47), the peak frequency (r = 0.42), the frequency bands alpha (r = 0.59) and beta (r = 0.56) as well as negative correlations in the frequency bands delta (r = -0.23) and theta (r = -0.42). The mean frequency and the activity in the frequency bands alpha, beta2, delta and theta contributed to the regression equation. The correlation between regression equation and the TE4D was rs = 0.87. The other tests also correlated with the TE4D: ADAS rs = -0.75, MMST rs = 0.82, SKT rs = -0.74, BCRS rs = -0.83. CONCLUSION: The TE4D showed convergent validity with the EEG parameters. Both the TE4D-score and the EEG-alterations correlated significantly with the degree of severity of Alzheimer's disease. This result underlines the assumption that the TE4D will be a useful instrument for the diagnostic process in dementia. PMID- 15290699 TI - A comparison of side effects of selective serotonin reuptake inhibitors and tricyclic antidepressants in older depressed patients: a meta-analysis. AB - OBJECTIVE: To examine the relative tolerability and side effect profile of tricyclic antidepressants and selective serotonin reuptake inhibitors in older depressed people. METHODS: A systematic literature search generated 37 randomised controlled trials of TCAs and SSRIs of which 11 were entered into a meta analysis comparing withdrawal rates and side effect profiles. RESULTS: 537 TCA recipients and 554 SSRI recipients were compared. TCAs had an increased withdrawal rate (RR: 0.24, CI 1.04, 1.47). A similar result was found when comparing classical TCAs (451 patients) (amitriptyline, clomipramine, doxepin and dothiepin) with SSRIs (466 patients) (RR 1.30 CI: 1.02,1.64). These findings were reflected in the increased TCA prevalence of side effects including dry mouth, drowsiness, dizziness and lethargy. No differences were found when comparing TCA related drugs (mianserin and trazadone) with SSRIs (RR 1.07 CI 0.43, 2.70). CONCLUSIONS: Despite the relative low prevalence of side effects associated with SSRIs a significant minority of older people find these drugs intolerable and experience nausea, vomiting, dizziness and drowsiness. We conclude that TCA related drugs are comparable to SSRIs in terms of tolerability and may offer an alternative when SSRIs are either contra-indicated or clinically unacceptable. PMID- 15290700 TI - A failure of 'pop-out' in visual search tasks in dementia with Lewy Bodies as compared to Alzheimer's and Parkinson's disease. AB - OBJECTIVES: The pattern of neural damage in dementia with Lewy Bodies (DLB) led us to hypothesize that patients with DLB would be particularly impaired on parallel ('pop-out') search tasks, relative to serial search tasks, and that this would serve to distinguish DLB from other forms of neurodegenerative disease, particularly Alzheimer's disease (AD). METHODS: To explore this possibility we tested four groups of observers (DLB, Alzheimer's Disease, Parkinson's Disease and age-matched controls) on parallel and serial search tasks, and a choice reaction time task. RESULTS: The DLB participants performed in a quantitatively and qualitatively different manner to the other groups. As predicted, they were particularly impaired on the parallel search task relative to the other observer groups. CONCLUSIONS: This pattern of deficit may reflect damage in the occipital areas leading to deficits in figure-ground segregation, and can assist differential diagnosis of DLB from other patients groups such as AD. PMID- 15290701 TI - Correlates of resident psychosocial status in long-term care. AB - OBJECTIVES: This pilot study randomly selected five nursing homes, five assisted living facilities, and 16 family care homes from a South-Central state in the US to identify correlates of resident psychosocial status. METHODS: In-person and telephone interviews were conducted with administrators and resident-family-staff triads (n = 79) to gather information on setting, resident functional status, family involvement, sociodemographic context, and resident psychosocial status. RESULTS: Results indicated that type of facility, resident health conditions, resident race, and facility family orientation were significantly correlated with dimensions of resident psychosocial status. CONCLUSIONS: The findings suggest that multiple informants are necessary to determine the processes that lead to residents' quality of life, and the consideration of diverse settings offers greater insight into how positive resident adaptation is achieved in long-term care. PMID- 15290702 TI - A counseling intervention for caregivers: effect on neuropsychiatric symptoms. AB - BACKGROUND: In Thailand, family caregivers have an important role in delivering care to patients with dementia. Most patients with dementia in Thailand and also in Western societies live in the community. Training caregivers may improve care of dementia patients. OBJECTIVE: We performed a treatment study of a six-month caregiver intervention with group counseling and support with provision of techniques to cope with non-cognitive symptoms of patients with dementia. We hypothesized that this caregiver intervention with group counseling and support would reduce behavioral and neuropsychiatric symptoms in the demented patients. SUBJECTS AND METHODS: We conducted a parallel group intervention study. A manual for group counseling and support was developed focusing on education regarding dementia, behavioral analysis and intervention, and environmental adaptation. Fifty nonprofessional caregivers-25 from the control group and 25 from the study group-of patients with dementia from the memory clinic at Siriraj Hospital were alternately assigned to each group as they presented to the clinic if they met the inclusion criteria and agreed to participate. The Thai Mental State Examination (TMSE) was used to assess dementia severity. Forty-five minute counseling sessions were conducted every 6-8 weeks for 6 months and assessments were conducted at 3 months and 6 months. The primary outcome measure was the Neuropsychiatric Inventory (NPI). RESULTS: A paired samples analysis of the NPI scores demonstrated a significant change of the total NPI scores at the end of six month from baseline in the intervention group (P = 0.045). Change from baseline of the comparison group was not significant. There was a trend towards improvement of the TMSE scores between the two groups at month six (p = 0.061). The result favored the treatment group. CONCLUSION: This study provided evidence of the utility of a non-pharmacologic intervention using group counseling in an out-patient setting for caregivers of patients with dementia. PMID- 15290703 TI - The utility of the Visual Analogue Scale for the assessment of depressive mood in cognitively impaired patients. AB - BACKGROUND: Early detection of depression in elderly demented patients may assist in adequate treatment. The aim of this study was to evaluate the utility of Visual Analogue Scale for depression among demented and mild cognitively impaired elderly patients. METHODS: 157 Patients, aged from 65 to 92 years, in a memory clinic were divided into two groups according to scores on the Mini Mental State Examination (MMSE): Demented group (62 patients) with MMSE scores from 16 to 23, and Mild Cognitively Impaired group (MCI-95 patients) with MMSE scores from 24 to 29. Subjects were diagnosed for depression according to DSM-IV criteria, using Hamilton Depression Rating Scale (HDRS) and a comprehensive clinical evaluation. All were administered a Visual Analogue Scale (VAS) for depression. RESULTS: In the demented group, 25 subjects were diagnosed as depressive. In the MCI group, 46 subjects were diagnosed as depressive. Mean VAS scores for the demented and MCI groups were similar, both for depressive and non-depressive patients. Correlations between VAS scores with HDRS scores and with the clinical diagnosis were high, although somewhat lower for the demented group. Analyzing data with ROC curve technique yielded significantly different ROC curves. Optimal cutoff point for the demented group was VAS value of 60, and 50 for the MCI group. CONCLUSIONS: VAS seems to be a useful tool for evaluation of depression among cognitively impaired patients. Severity of cognitive decline in the elderly may influence the cutoff points on VAS for detecting depression. Further large-scale studies are needed to substantiate our observation. PMID- 15290704 TI - An audit of physical restraint and seclusion in five psychogeriatric admission wards in Victoria, Australia. PMID- 15290705 TI - The impact of NICE guidelines on the diagnosis and treatment of Alzheimer's disease among general medical hospital inpatients. PMID- 15290706 TI - Do not use 'BPSD' if you want to be cited. PMID- 15290707 TI - Depression and coronary heart mortality in older coronary heart disease patients. PMID- 15290708 TI - Have risk factors for very late-onset schizophrenia-like psychosis changed in the last 40 years? PMID- 15290709 TI - The potentials of MS-based subproteomic approaches in medical science: the case of lysosomes and breast cancer. AB - Because of the great number of women who are diagnosed with breast cancer each year, and though this disease presents the lowest mortality rate among cancers, breast cancer remains a major public health problem. As for any cancer, the tumorigenic and metastatic processes are still hardly understood, and the biochemical markers that allow either a precise monitoring of the disease or the classification of the numerous forms of breast cancer remain too scarce. Therefore, great hopes are put on the development of high-throughput genomic and proteomic technologies. Such comprehensive techniques should help in understanding the processes and in defining steps of the disease by depicting specific genes or protein profiles. Because techniques dedicated to the current proteomic challenges are continuously improving, the probability of the discovery of new potential protein biomarkers is rapidly increasing. In addition, the identification of such markers should be eased by lowering the sample complexity; e.g., by sample fractionation, either according to specific physico-chemical properties of the proteins, or by focusing on definite subcellular compartments. In particular, proteins of the lysosomal compartment have been shown to be prone to alterations in their localization, expression, or post-translational modifications (PTMs) during the cancer process. Some of them, such as the aspartic protease cathepsin D (CatD), have even been proven as participating actively in the disease progression. The present review aims at giving an overview of the implication of the lysosome in breast cancer, and at showing how subproteomics and the constantly refining MS-based proteomic techniques may help in making breast cancer research progress, and thus, hopefully, in improving disease treatment. PMID- 15290710 TI - Optical detection methods for mass spectrometry of macroions. AB - Detection of macroions has been a challenge in the field of mass spectrometry. Conventional ionization-based detectors, relying on production and multiplication of secondary electrons, are restricted to detection for charged particles of m/z < 1 x 10(6). While both energy-sensitive and charge-sensitive detectors have been developed recently to overcome the limitation, they are not yet in common use. Photon-sensitive detectors are suggested to be an alternative, with which detection of macroions (or charged particles) by either elastic light scattering (ELS) or laser-induced fluorescence (LIF) has been possible. In this article, we provide a critical review on the developments of novel optical detection methods for mass spectrometry of macroions, including both micron-sized and nano-sized synthetic polymers as well as high-mass biomolecules. Design and development of new spectrometers making possible observations of the mass spectra of macroions with sizes in the range of 10-10(3) nm or masses in the range of 1-10(6) MDa are illustrated. The potential and promise of this optical approach toward macroion detection with high efficiency are discussed in practical aspects. PMID- 15290712 TI - "Cellular astronomy"--a foreseeable future in cytometry. AB - Advances in electro-optic technology within the past 2 years, notably the development of high-intensity light-emitting diodes and highly efficient charge coupled device cameras, have made it feasible to produce small, simple, rugged, automated fluorescence image cytometers, with selling prices well below 10,000 US dollars, that can make measurements previously the exclusive domain of flow and scanning cytometers costing many times more. It should be feasible to apply the new cytometric technology in scientific and geographic areas for which a previous generation of instruments was too complex and too expensive, e.g., to problems of diagnosis and management of infectious diseases prevalent at critical levels in resource-poor areas, such as the human immunodeficiency virus, malaria, and tuberculosis. PMID- 15290713 TI - An analytical system based on a compact flow cytometer for DNA fragment sizing and single-molecule detection. AB - BACKGROUND: Previous reports have demonstrated accurate DNA fragment sizing of linear DNA fragments, from 564 to approximately 4 x 10(5) bp, in a flow system. B phycoerythrin (B-PE), commonly used in conventional cytometric applications that require high-sensitivity, was the first fluorophore detected in flow at the single-molecule level. METHODS: Dilute solutions of stained DNA fragments or B-PE were analyzed in a simplified, compact flow system, with enhanced performance and lower cost, utilizing a solid-state laser and a single-photon sensing avalanche photodiode detector (SSAPD). Extensive data processing and display software, developed specifically for the photon-counting data stream, extracts correlated height, width, and area features from bursts of photons due to discrete molecules passing through the sensing region in the flow channel. RESULTS: DNA fragment sizing in flow has now been demonstrated for SYTOX-orange-stained fragments ranging in size over 3.4 orders of magnitude, from 125 to 5 x 10(5) bp. For Lambda bacteriophage DNA (lambda DNA; 48.5 kbp) a CV of 1.2 % has been achieved. Analysis of a femtomolar B-PE solution demonstrates that the bursts of photons from individual molecules can be baseline-resolved with 0.5 mW of laser power at a signal to noise ratio (SNR) of approximately 30, with approximately 100 photons detected from each molecule. CONCLUSIONS: A compact, low-power, high-sensitivity system detects DNA fragments as small as 125 bp or individual B-PE molecules in a flowing liquid stream. Demonstrated linearity, sensitivity, and resolution indicate that <1.0 mW of laser power is optimal, permitting further miniaturization of the system and additional cost reduction. Comprehensive analytical software exploits the standard cytometric paradigm of multiple 2D graphs and gating to extract features from classes of individually analyzed biomolecules. This complete system is thus poised to engage high-sensitivity applications not amenable to conventional flow cytometric instrumentation. PMID- 15290714 TI - Multiplexed, particle-based detection of DNA using flow cytometry with 3DNA dendrimers for signal amplification. AB - BACKGROUND: Complex mixtures of DNA may be found in environmental and medical samples. There is a need for techniques that can measure low concentrations of target DNAs. For a multiplexed, flow cytometric assay, we show that the signal-to noise ratio for fluorescence detection may be increased with the use of 3DNA dendrimers. A single fluorescent DNA molecule per bead could be detected with conventional flow cytometry instrumentation. METHODS: The analyte consisted of single-stranded (ss) DNA amplicons that were hybridized to capture probes on the surface of fluorescent polystyrene microspheres (beads) and initially labeled with streptavidin-R-phycoerythrin (single-step labeling). These beads have a low reporter fluorescence background and high efficiency of DNA hybridization. The DNA/SA-RPE complex was then labeled with 3DNA dendrimers and SA-RPE. The bead complexes were detected with a Luminex 100 flow cytometer. Bead standards were developed to convert the intensity to the number of SA-RPE labels per bead and the number of dendrimers per bead. RESULTS: The dendrimer assay resulted in 10 fold fluorescence amplification compared with single-step SA-RPE labeling. Based on concentration curves of pure target ss-amplicons, the signal-to-noise ratio of the dendrimer assay was greater by a factor of 8.5 over single-step SA-RPE labeling. The dendrimer assay was tested on 16S ribosomal DNA amplified from filter retentates of contaminated groundwater. Multiplexed detection of a single dendrimer-labeled DNA molecule per bead was demonstrated. CONCLUSIONS: Multiplexed detection of DNA hybridization on a single molecule level per bead was achieved with conventional flow cytometry instrumentation. This assay is useful for detecting target DNAs at low concentrations. PMID- 15290715 TI - Flow cytometric analysis of isolated liver mitochondria to detect changes relevant to cell death. AB - BACKGROUND: Mitochondria are key players in many forms of cell death, and mitochondrial production of reactive oxygen species (ROS), membrane depolarization, permeability changes, and release of apoptogenic proteins are involved in these processes. Flow cytometric analysis of isolated mitochondria enables parallel analysis of mitochondrial structure and function in individual mitochondria, and small mitochondrial samples are sufficient for analysis. This article describes a well-characterized protocol for flow cytometric analysis of isolated liver mitochondria that can be used to detect mitochondrial alterations relevant to cell death. METHODS: Fluorescent probes were used to selectively stain mitochondria (nonyl acridine orange), and to measure membrane potential (tetramethylrhodamine-methyl-ester, 1,1',3,3,3',3'-hexamethylindodicarbocyanine iodide), as well as production of ROS (2',7'-dichlorodihydrofluorescein diacetate). Calcium-induced mitochondrial swelling was detected as a decrease in SSC. To ensure optimal concentrations of all probes, the effect on mitochondrial respiration was evaluated. RESULTS: This protocol can be used to determine the purity of the mitochondrial preparation, to detect calcium-induced morphological changes, small mitochondrial de- and hyperpolarizations, as well as physiological changes in ROS generation. CONCLUSIONS: Flow cytometry is a very useful tool to simultaneously analyze several mitochondrial parameters that are important in the induction of mitochondria-mediated cell death. PMID- 15290717 TI - Green fluorescent protein-propidium iodide (GFP-PI) based assay for flow cytometric measurement of bacterial viability. AB - BACKGROUND: Several staining protocols have been developed for flow cytometric analysis of bacterial viability. One promising method is dual staining with the LIVE/DEAD BacLight bacterial viability kit. In this procedure, cells are treated with two different DNA-binding dyes (SYTO9 and PI), and viability is estimated according to the proportion of bound stain. SYTO9 diffuses through the intact cell membrane and binds cellular DNA, while PI binds DNA of damaged cells only. This dual-staining method allows effective separation between viable and dead cells, which is far more difficult to achieve with single staining. Although SYTO9-PI dual staining is practical for various bacterial viability analyses, the method has a number of disadvantages. Specifically, the passage of SYTO9 through the cell membrane is a slow process, which is significantly accelerated when the integrity of the cell membrane is disrupted. As a result, SYTO9 binding to DNA is considerably enhanced. PI competes for binding sites with SYTO9 and may displace the bound dye. These properties diminish the reliability of the LIVE/DEAD viability kit. In this study, we investigate an alternative method for measuring bacterial viability using a combination of green fluorescent protein (GFP) and PI, with a view to improving data reliability. METHODS: Recombinant Escherichia coli cells with a plasmid containing the gene for jellyfish GFP were stained with PI, and green and red fluorescence were measured by FCM. For comparison, cells containing the plasmid from which gfp was removed were stained with SYTO9 and PI, and analyzed by FCM. Viability was estimated according to the proportion of green and red fluorescence. In addition, bioluminescence and plate counting (other methods to assess viability) were used as reference procedures. RESULTS: SYTO9-PI dual staining of bacterial cells revealed three different cell populations: living, compromised, and dead cells. These cell populations were more distinct when the GFP-PI combination was used instead of dual staining. No differences in sensitivity were observed between the two methods. However, substitution of SYTO9 with GFP accelerated the procedure. Bioluminescence and plate counting results were in agreement with flow cytometric viability data. CONCLUSIONS: In bacterial viability analyses, the GFP-PI combination provided better distinction between current viability stages of E. coli cells than SYTO9-PI dual staining. Additionally, the overall procedure was more rapid. No marked differences in sensitivity were observed. PMID- 15290716 TI - Proliferative response of bystander cells adjacent to cells with incorporated radioactivity. AB - BACKGROUND: In a recent study, we showed that cells irradiated with gamma-rays stimulate cell growth of unirradiated (bystander) cells, when the two populations are co-cultured as a mixture. Direct cell-to-cell contact appears to be a prerequisite for the proliferative response of the bystander cells. The aim of the current work is to investigate the possible proliferative bystander effects caused by intracellular irradiation with incorporated radionuclides, specifically the short-range beta particle emitter, tritium ((3)H). METHODS: Subconfluent monolayers of rat liver epithelial cells (WB-F344) were incubated in the presence of (methyl-(3)H)thymidine ((3)HTdR) at concentrations ranging between 5.2 kBq/ml and 57.8 kBq/ml for 18 h. Radiolabeled cells, containing between 0.7 x 10(-3) Bq/cell and 8.8 x 10(-3) Bq/cell were mixed with unlabeled (i.e., bystander) cells in a ratio of 1:1 and cultured together for 24 h followed by an flow cytometry (FCM) study of their proliferation. In order to discriminate the two populations of co-cultured cells, one cell population (unlabeled bystander cells) was stained with carboxyfluorescein diacetate, succinimidyl ester (CFDA SE), which metabolizes intracellularly. The absorbed doses received by the radiolabeled cells that contained 0.7 x 10(-3), 2.5 x 10(-3), and 8.8 x 10(-3) Bq/cell were 0.14, 0.49, and 1.7 Gy, respectively. RESULTS: Cells that were not treated with tritiated thymidine (unlabeled cells), in the presence of radiolabeled cells that received absorbed doses from 0.14-1.7 Gy, showed enhanced cell growth by approximately 9 to 10%. CONCLUSIONS: Cells labeled with (3)HTdR can induce increased proliferation in neighboring unlabeled bystander cells. FCM provides an excellent basis for characterization of proliferative bystander effects in co-culture systems. PMID- 15290718 TI - Chromatic shifts in the fluorescence emitted by murine thymocytes stained with Hoechst 33342. AB - BACKGROUND: Many methods in flow cytometry rely on staining DNA with a fluorescent dye to gauge DNA content. From the relative intensity of the fluorescence signature, one can then infer position in cell cycle, amount of DNA (i.e., for sperm selection), or, as in the case of flow karyotyping, to distinguish individual chromosomes. This work examines the staining of murine thymocytes with a common DNA dye, Hoechst 33342, to investigate nonlinearities in the florescence intensity as well as chromatic shifts. METHODS: Murine thymocytes were stained with Hoechst 33342 and measured in a flow cytometer at two fluorescence emission bands. In other measurements, cells were stained at different dye concentrations, and then centrifuged. The supernatant was then used for a second round of staining to test the amount of dye uptake. Finally, to test for resonant energy transfer, we measured fluorescence anisotropy at two different wavelengths. RESULTS: The fluorescence of cells stained with Hoechst 33342 is a nonlinear process that shows an overall decrease in intensity with increased dye uptake, and spectral shift to the red. Along with the spectral shift of the fluorescence to the longer wavelengths, we document decreases in the fluorescence anisotropy that may indicate resonant energy transfer. CONCLUSIONS: At low concentrations, Hoechst 33342 binds to the minor groove of DNA and shows an increase in fluorescence and a blue shift upon binding. At higher concentrations, at which the dye molecules can no longer bind without overlapping, the blue fluorescence decreases and the red fluorescence increases until there is approximately one dye molecule per DNA base pair. The ratio of the blue fluorescence to the red fluorescence is an accurate indicator of the cellular dye concentration. PMID- 15290719 TI - Automated counting of mammalian cell colonies by means of a flat bed scanner and image processing. AB - BACKGROUND: Clonogenic assays are used frequently to measure the cell killing and mutagenic effects of radiation and other agents. Clonogenic assays carried out manually are tedious and time-consuming and involve a significant element of subjectivity. However, several commercial automatic colony counters are available. Based on CCD video imaging and image analysis they are relatively expensive and can analyze only one petri dish at a time. METHOD: We have developed a cheaper and more efficient device, which employs a flat bed scanner to image 12 60-mm petri dishes at a time. Two major problems in automated colony counting are the clustering of colonies and edge effects. By using standard image analysis and implementing an inflection point algorithm, these problems were greatly diminished. The resulting system was compared with two manual colony counts, as well as with automated counts with the Oxford Optronix ColCount colony counter for cell lines V79 and HaCaT. RESULTS: Comparisons assuming the manual counts to be correct showed that our automatic counter was slightly more accurate than the commercial unit. CONCLUSIONS: As a whole, our automated colony counter performed significantly better than the commercial unit with regard to processing time, cost and accuracy. PMID- 15290720 TI - Slide-based cytometry and predictive medicine: the 8th Leipziger workshop and the 1st international workshop on slide-based cytometry. AB - Slide-based cytometry (SBC) and related techniques offer unique tools to perform complex diagnostic procedures at very early disease stages. Multicolor or polychromatic analysis of cells by SBC is of special importance, not only as a cytomics technology platform, but for patients with low blood volume such as neonates. The exact knowledge of the location of each cell on the slide allows the specimen to be restained and subsequently reanalyzed. These separate measurements can be fused to one data file (merging), increasing the information obtained per cell. Relocalization and optical evaluation of the cells, a typical feature of SBC, can be of integral importance for cytometric analysis. Predictive medicine is aimed at the detection of changes in the patient's state prior to the manifestation of deterioration or improvement. Such instances are concerned with multiorgan failure in sepsis or noninfectious posttraumatic shock in intensive care patients, or the pretherapeutic identification of high risk patients in cancer cytostatic therapy. Early anti-infectious or anti-shock therapy, as well as curative chemotherapy in combination with stem cell transplantation, may provide better survival chances for the patient as well as concomitant cost containment. Predictive medicine-guided, individualized, early reduction or cessation of therapy may lower or abrogate potential therapeutic side effects (individualized medicine). With the 8th Leipziger Workshop and the 1st International Workshop on Slide-Based Cytometry, cytomics technologies moved to more practical applications in the clinics and the clinical laboratory. This development will be continued in 2004, at the upcoming Leipziger Workshop and the International Workshop on Slide-Based Cytometry. PMID- 15290721 TI - The joy of citrulline: new insights into the diagnosis, pathogenesis, and treatment of rheumatoid arthritis. PMID- 15290722 TI - Antiphospholipid antibody syndrome: further evidence to guide clinical practice? PMID- 15290723 TI - Observational cohort studies and well controlled clinical trials--we need them both! PMID- 15290724 TI - Why I no longer accept pens (or other "gifts") from industry (and why you shouldn't either). PMID- 15290725 TI - Osteoprotegerin and receptor activator of nuclear factor-kappaB ligand mRNA expression in patients with rheumatoid arthritis and healthy controls. AB - OBJECTIVE: To further understand the role of osteoprotegerin (OPG) and receptor activator of nuclear factor-kappaB ligand (RANK-L) in rheumatoid arthritis (RA), we studied the levels of RANK-L and OPG mRNA in peripheral blood mononuclear cells (PBMC) and synovial tissue of patients with RA and controls. METHODS: RANK L and OPG mRNA levels were measured in PBMC and CD4+/CD8+ T cell subsets of patients with chronic RA, osteoarthritis (OA), and healthy controls, using quantitative real-time polymerase chain reaction. OPG and RANK-L mRNA levels were measured in paired blood and synovial tissue samples of patients with early, untreated RA at 2 timepoints with an interval of 16 weeks. RESULTS: RANK-L mRNA levels were significantly higher in PBMC of patients with early and chronic RA compared to healthy controls. Contrary to healthy controls, RANK-L mRNA levels in patients with chronic RA were mainly of CD4+ T cell origin. OPG mRNA was observed in the blood of all (17/17) early RA patients, but could not be detected in chronic RA patients (0/14) or in patients with OA (0/8). Three out of 17 healthy controls showed measurable levels of OPG mRNA. The OPG/RANK-L ratio tended to be higher in the synovium than in the PBMC of early RA patients. RANK-L mRNA in synovial tissue was mainly of non-T cell origin. CONCLUSION: Since RANK-L and OPG mRNA levels are elevated in PBMC of RA patients, and CD4+ T cells are the major contributors to RANK-L mRNA expression, mononuclear cells in patients with RA may be involved in the pathways that regulate bone metabolism. PMID- 15290726 TI - Serological bone markers and joint damage in early polyarthritis. AB - OBJECTIVE: To investigate the relationship between osteocalcin (OC), a marker of bone formation, and the recently developed serum marker of bone resorption, beta C-telopeptide (beta-CTx), and radiographic damage in patients with early oligo- and polyarthritis. METHODS: Patients with peripheral arthritis of > or = 2 joints and < 2 years of symptom duration were studied. The OC and beta-CTx concentrations at baseline were correlated with disease activity and radiographic damage at baseline, and with radiographic progressive disease after 2 years (delta Sharp/van der Heijde score > or = 5). The additional value of serum bone metabolism markers to predict radiographic progressive disease was compared to that of established prognostic factors by multivariate logistic regression analysis. RESULTS: Two hundred seventy-nine patients (67% female; median age 56 yrs, range 18-83) were included in the study, of whom 73% were diagnosed with rheumatoid arthritis (RA). Baseline levels of beta-CTx (p < 0.05) were significantly correlated with baseline radiographic damage whereas OC was not. beta-CTx was also significantly (p < 0.001) related to measures of disease activity like erythrocyte sedimentation rate, C-reactive protein, and the disease activity score DAS28. Radiographic progressive disease after 2 years corresponded univariately with increased levels of beta-CTx (p < 0.001), but not with OC. In multivariate analysis, beta-CTx was not superior to other measures of radiographic progressive disease such as autoantibodies and disease activity. CONCLUSION: Increased serum levels of the bone turnover marker beta-CTx are associated with radiographic damage at baseline and radiographic progression after 2 years. However, beta-CTX is less predictive than markers already in use. PMID- 15290727 TI - Immune reactivity to connective tissue antigens in pristane induced arthritis. AB - OBJECTIVE: Pristane induced arthritis (PIA) is a seropositive experimental murine model that closely resembles rheumatoid arthritis (RA). Immune reactivity to a broad spectrum of autoantigens has been recognized in this disease model. We investigated the specificity of the autoimmune response in PIA to determine whether reactivity to connective tissue antigens is associated with the development of arthritis. METHODS: DBA/1 mice were injected with pristane and evaluated for development of joint disease and autoimmunity. Lymph nodes, spleen, sera, and arthritic paws were investigated at 1, 2, 4, 6, 9, and 12 months postinjection. T cell responses to 16 different joint components were evaluated using proliferation assays, and sera were assayed by ELISA for antibodies to these joint antigens. Cytokine concentrations after antigenic stimulation were assessed by ELISA in cultured cell supernatants and by real-time polymerase chain reaction using mRNA from spleens and arthritic paws. RESULTS: ELISA revealed positive responses to glucose-6-phosphate isomerase, chondroitin sulfate B, collagen I, collagen II, aggrecan, and DNA between 4 and 12 months post-pristane injection. In vitro tumor necrosis factor-alpha (TNF-alpha), interferon-gamma (IFN-gamma), and interleukin 6 (IL-6) responses were detected during reactions to most antigens tested, while IL-4 responses were absent. Cytokine analysis in arthritic joints revealed consistent expression of IL-1, IL-4, IL-6, TNF-alpha, and IFN-gamma mRNA. CONCLUSION: These results indicate that PIA animals develop both T cell and antibody responses to a broad spectrum of connective tissue antigens. Biglycan, aggrecan, and decorin may be relevant antigens in the pathogenesis of PIA, but no specific reaction pattern could be associated with the occurrence of disease. The data suggest that the development of pristane arthritis is not dependent upon reactivity against a single connective tissue antigen, but is a polyspecific autoimmune response to joint components elicited in pristane injected mice. PMID- 15290728 TI - Increased CD40 expression on articular chondrocytes from patients with rheumatoid arthritis: contribution to production of cytokines and matrix metalloproteinases. AB - OBJECTIVE: To examine expression of CD40 on articular chondrocytes derived from patients with rheumatoid arthritis (RA) and osteoarthritis (OA) and to investigate roles of CD40 on articular chondrocytes in production of inflammatory mediators associated with degradation of articular cartilage. METHODS: Articular cartilage samples were obtained from patients with RA and OA at total knee arthroplasty. Expression of CD40 on chondrocytes was examined using immunohistochemistry, reverse transcriptase-polymerase chain reaction (RT-PCR), and flow cytometry. Tumor necrosis factor-alpha (TNF-alpha), interleukin 6 (IL 6), matrix metalloproteinase (MMP)-3, and tissue inhibitor of matrix metalloproteinase (TIMP)-1 in culture supernatants of chondrocytes were measured by ELISA. RESULTS: Immunohistochemical staining revealed that CD40 was expressed on RA chondrocytes in vivo, but not in OA chondrocytes. RT-PCR and flow cytometry showed that cultured articular chondrocytes from RA patients, but not from OA patients, constitutively expressed CD40, and that interferon-gamma (IFN-gamma) enhanced the expression of CD40 on chondrocytes from both RA and OA patients. Membrane fraction of mouse myeloma Ag8 cells that was transfected with human CD154 (Ag8-hCD154) induced production of TNF-alpha, IL-6, and MMP-3 in RA chondrocytes, and pretreatment of RA chondrocytes with IFN-gamma enhanced the response. OA chondrocytes did not respond to stimulation with the membrane fraction of Ag8-hCD154 cells alone, but the cells did produce TNF-alpha, IL-6, and MMP-3 after pretreatment with IFN-gamma. CONCLUSION: CD40-CD154 interaction augments the expression of inflammatory cytokines and MMP in chondrocytes and contributes to an intrinsic process of cartilage degradation in RA. PMID- 15290729 TI - Serum galactosyltransferase isoform changes in rheumatoid arthritis. AB - OBJECTIVE: To investigate glycosylation changes associated with rheumatoid arthritis (RA) by determining whether there are beta1,4-galactosyltransferase (GTase) isoforms specific to or altered in the serum of patients with RA. Methods. Serum GTase isoform profiles were determined using isoelectric focusing (IEF) in patients with active RA (n = 9), disease controls (DC; n = 9), and healthy individuals (HI; n = 10). RESULTS: There was a highly significant difference (p < 0.0001) between the IEF profiles. The RA IEF profile was significantly (p < 0.0001) different from that of the DC or the HI group. There was, however, no significant difference between the DC and HI profiles. Serum GTase samples from 8/9 RA, 9/9 DC, and 9/10 HI resolved into 2 distinct peaks of activity. The RA isoform profile was associated with an acidic shift. There were no significant differences in the pH value of the first peak; the second peak was found to be significantly more acidic in the RA group (mean pH 5.02) compared to the DC and HI group (mean pH 5.20; p < 0.05). The RA associated isoform constituted a significantly greater proportion of total enzymatic activity in the RA sera (16.1%) compared to DC and HI (13.5%; p < 0.05 and 12.6%; p < 0.01, respectively). RA and HI serum GTase desialylation resulted in an alkaline shift of the isoforms into similar pH bands: 5.25-5.50, 5.70-5.85, and 6.20-6.40. GTase was found to be on average 75% more active in its desialylated form than in its sialylated state. CONCLUSION: RA is associated with a differential expression of GTase isoforms. This may be due to increased hypersialylation, which has the potential to adversely affect the catalytic activity of the enzyme, thus providing a possible mechanism for posttranslational regulation of GTase activity in RA. PMID- 15290730 TI - Combination leflunomide and methotrexate (MTX) therapy for patients with active rheumatoid arthritis failing MTX monotherapy: open-label extension of a randomized, double-blind, placebo controlled trial. AB - OBJECTIVE: To obtain additional safety and efficacy data on leflunomide (LEF) treatment in combination with methotrexate (MTX) therapy in an open-label extension study in patients with rheumatoid arthritis (RA). METHODS: Following a 24 week, randomized, double-blind trial of adding placebo (PLA) or LEF to stable MTX therapy, patients could enter a 24 week extension. Subjects randomized to LEF and MTX continued treatment [(LEF/LEF) + MTX]. Subjects randomized to PLA and MTX switched to LEF (10 mg/day, no loading dose) and MTX [(PLA/LEF) + MTX]. The double-blind regarding initial randomization was maintained. RESULTS: For subjects in the extension phase, American College of Rheumatology 20% (ACR20) responder rates for the (LEF/LEF) + MTX group were maintained from Week 24 (57/96, 59.4%) to Week 48 (53/96, 55.2%). ACR20 responder rates improved in patients switched to LEF from PLA at Week 24 [(PLA/LEF) + MTX] from 25.0% (24/96) at Week 24 to 57.3% (55/96) at Week 48. Patients in the extension who switched from PLA to LEF without a loading dose exhibited a lower incidence of elevated transaminases compared to patients initially randomized to LEF. Diarrhea and nausea were less frequent during the open-label extension in patients who did not receive a LEF loading dose. CONCLUSION: Response to therapy was maintained to 48 weeks of treatment in patients who continued to receive LEF and MTX during the extension. Importantly, ACR20 response rates after 24 weeks of LEF therapy were similar between patients switched from PLA to LEF without loading dose, and those who received a loading does of LEF (100 mg/day x 2 days) at randomization. Fewer adverse events were reported in patients switched to LEF without a loading dose. PMID- 15290731 TI - Etanercept (Enbrel) in patients with rheumatoid arthritis with recent onset versus established disease: improvement in disability. AB - OBJECTIVE: To compare etanercept-induced improvement in disability of patients with recent onset of rheumatoid arthritis (RA) to that of patients with established RA. METHODS: Health Assessment Questionnaire (HAQ) scores were collected over 3 years in 2 groups of patients with RA who were treated with etanercept. The first group consisted of 207 patients with recent onset RA (mean duration of 1 year) who had not previously received methotrexate, and the second group consisted of 464 patients with established RA (mean duration of 12 years) who had failed one or more disease-modifying antirheumatic drugs. RESULTS: Baseline demographics and disease characteristics were similar in the 2 groups, except for HAQ scores and C-reactive protein levels, which were higher in the established RA group. Patients in both groups showed rapid and sustained clinical responses with etanercept therapy, but patients with recent onset RA showed significantly greater improvement in HAQ scores compared with patients with established RA. The difference in magnitude of HAQ score improvement between groups was observed as early as week 2 after initiation of etanercept and persisted throughout the 3-year time frame. At year 3, significantly more patients with recent onset RA had a HAQ score of zero (26%) versus those with established RA (14%, p = 0.0095). CONCLUSION: Although etanercept therapy significantly improved disability scores in both groups, patients with recent onset of RA showed greater benefit in HAQ scores than patients with established RA. These results support prompt treatment of RA at an early stage of disease to minimize patient disability. PMID- 15290732 TI - Infliximab dose and clinical status: results of 2 studies in 1642 patients with rheumatoid arthritis. AB - OBJECTIVE: Infliximab is an effective anti-tumor necrosis factor (TNF) agent widely used in the treatment of rheumatoid arthritis (RA). Initially recommended at a dose of 3 mg/kg, subsequent label revisions allowed doses up to 10 mg/kg or at 4-week intervals rather than the originally suggested 8-week intervals, if clinically indicated. The doses used have implications for efficacy and costs, but no data exist for actual dose used in the US. This study evaluates the dosage and rates of increase in infliximab-treated patients with RA. METHODS: Study 1: Review of patient charts and infusion records for 394 RA patients from 2 large rheumatology practices comprising 15 rheumatologists in Dallas, Texas. Study 2: Survey of 1324 RA patients using infliximab participating in a longitudinal study of RA outcomes. Patients completed a detailed questionnaire about clinical status and infliximab use. RESULTS: The results of the 2 studies were similar: the average infliximab dose was 5 mg/kg, increasing most rapidly until the end of the first years, after which the increase was slowed. Increases > 3 mg/kg occurred in 61% of patients in Study 1 and 56% in Study 2. The 8-week treatment interval was almost universally used, and more than 95% of infusions occurred in this interval. The most common reason for increase in dose was insufficient response. Among patients who completed 4 infusions, 75% remained on therapy at 2 years after infliximab start. The average improvement in Health Assessment Questionnaire disability score was 0.28. CONCLUSION: Infliximab dose increases are common, particularly during the first year of treatment. The average dose is 5 mg/kg. Seventy-five percent of patients continue using infliximab 2 years after treatment onset. PMID- 15290734 TI - The clinical effect of dietary supplementation with omega-3 fish oils and/or copper in systemic lupus erythematosus. AB - OBJECTIVE: To determine the effect of dietary supplementation with omega-3 fish oils with or without copper on disease activity in systemic lupus erythematosus (SLE). Fish oil supplementation has a beneficial effect on murine models of SLE, while exogenous copper can decrease the formation of lupus erythematosus cells in rats with a hydralazine-induced collagen disease. METHODS: A double blind, double placebo controlled factorial trial was performed on 52 patients with SLE. Patients were randomly assigned to 4 treatment groups. Physiological doses of omega-3 fish oils and copper readily obtainable by dietary means were used. One group received 3 g MaxEPA and 3 mg copper, another 3 g MaxEPA and placebo copper, another 3 mg copper and placebo fish oil, and the fourth group received both placebo capsules. Serial measurements of disease activity using the revised Systemic Lupus Activity Measure (SLAM-R) and peripheral blood samples for routine hematological, biochemical, and immunological indices were taken at baseline, 6, 12, and 24 weeks. RESULTS: There was a significant decline in SLAM-R score from 6.12 to 4.69 (p < 0.05) in those subjects taking fish oil compared to placebo. No significant effect on SLAM-R was observed in subjects taking copper. Laboratory variables were unaffected by either intervention. CONCLUSION: In the management of SLE, dietary supplementation with fish oil may be beneficial in modifying symptomatic disease activity. PMID- 15290733 TI - Association between systemic lupus erythematosus and Helicobacter pylori seronegativity. AB - OBJECTIVE: Helicobacter pylori is a gram negative spiral bacterium that is clearly associated with a variety of gastrointestinal pathologies. A number of non-gastrointestinal diseases have also been associated with H. pylori. We investigated the prevalence of H. pylori seropositivity as part of a larger serologic survey in a group of 466 patients with systemic lupus erythematosus (SLE) and 466 controls. METHODS: We studied subjects for seropositivity against 5 antigens including mumps, measles, rubella, varicella zoster, and H. pylori. The 466 SLE patients were taken from a total of 290 pedigrees multiplex for SLE and matched to 466 controls for age (+/- 3 yrs), sex, and ethnicity to non-SLE affected individuals, taken mostly from the same collection of pedigrees multiplex for SLE. Assays for seropositivity were performed using a heterogeneous immunoassay technique. Pearson's chi-square was used to test for association of categorical variables and Student t-test for continuous variables. Logistic regression was used to compute the odds ratio for H. pylori seropositivity in patients and controls. RESULTS: There was a significant difference only in H. pylori seropositivity between SLE cases and their controls. The results were not altered by intrafamilial correlation. Subset analysis by race and sex showed that the differences between the African-American female patients with SLE and their matched controls were responsible for this association. Female African-American patients with SLE had a lower prevalence of H. pylori seropositivity compared to controls (38.1% vs 60.2%, OR 0.41, p = 0.0009, 95% CI 0.24-0.69). Of the 113 African-American female SLE patients in the study group, 43 were seropositive for H. pylori. The mean age of onset for SLE was older in the seropositive group (34.4 yrs) compared to the seronegative SLE patients (28.0 yrs) (t = 2.11, p = 0.039). CONCLUSION: Of 5 serologic tests performed, only the frequency of H. pylori seropositivity was different between SLE cases and their controls, and then only in African-Americans. We found an association between being seronegative for H. pylori and the development of SLE in African-American women, who also tend to be younger at the time of disease onset. These findings suggest that there is a possible protective role for H. pylori infection against the development of SLE or that immunoregulatory events leading to H. pylori seropositivity are inversely related to the risk of SLE. PMID- 15290735 TI - Urine protein-to-creatinine ratio is a reliable measure of proteinuria in lupus nephritis. AB - OBJECTIVE: To evaluate the 24-hour urine protein-to-creatinine (U pr:cr) ratio compared to 24-h urine total protein excretion as a measure of proteinuria in patients with lupus nephritis. METHODS: Proteinuria in 8 patients with lupus nephritis treated with cyclophosphamide was monitored by total protein excretion and U pr:cr ratio in 24-h urine collections. A median of 16 measurements per patient were collected over a median of 47 months. Adequacy of the 24-h collection was assessed by comparing total urine creatinine to the predicted creatinine. Collections in which the difference between the predicted 24-h urine creatinine and the measured 24-h urine creatinine was greater than or equal to 20% were defined as inadequate collections. RESULTS: Timed 24-h urine collections were frequently inadequate (30.2% of total collections were under-collections, while 14.3% were over-collections). We found 87.5% of patients had at least one under-collection whereas 75% had at least one over-collection. Correlations between total protein and U pr:cr ratio for individual patients ranged from 0.87 to 0.99 (mean 0.95). For the entire sample, the correlation (R2 = 0.89) of total urine protein to Upr:cr ratio was excellent. Excluding the 38 under-collections led to improvement in the overall correlation (0.94). Excluding the 18 over collections led to a correlation of 0.89. Excluding both under-collections and over-collections led to a correlation of 0.94. CONCLUSION: In patients with lupus nephritis, the 24-h U pr:cr ratio is highly correlated with the 24-h urine protein excretion when the collections are adequate. The error of the estimate is higher when collections are poor. PMID- 15290736 TI - Antiphospholipid syndrome and asymptomatic carriers of antiphospholipid antibody: prospective analysis of 404 individuals. AB - OBJECTIVE: We carried out a prospective analysis of clinical and analytical findings in individuals with antiphospholipid antibodies (aPL). METHODS: We prospectively studied 404 individuals, classified in 2 groups: (1) patients with primary or secondary antiphospholipid syndrome (APS, n = 226); and (2) asymptomatic carriers of aPL (n = 178). Patients with APS and thrombosis were treated with dicumarin, and an international normalized ratio around 3.0 (range 2.5-3.5) was targeted. Asymptomatic carriers were not treated, but specific prophylaxis with low molecular weight heparin or aspirin was prescribed for the periods when individuals were at increased risk of thrombosis. Both groups of individuals were followed up at semester intervals for 36 months. RESULTS: Patients with APS presented with venous (n = 106, 46.9%) and/or arterial (n = 70. 31%) thrombosis or fetal loss (n = 58 out of 112 women of fertility age, 51.8%). At the time of the first thrombotic event, 50.0% of patients with APS had coincident risk factors for thrombosis (previous surgery and prolonged immobilization were significantly associated with venous thrombosis, and hypercholesterolemia and arterial hypertension with arterial thrombosis). Eighteen patients with APS died during the study period. Recurrence of thrombosis in patients with APS was linked to insufficient anticoagulation. During the followup, no episode of thrombosis was detected in any asymptomatic carrier. The proportion of subjects with aPL was similar in patients and in asymptomatic carriers. The proportion of subjects with aPL decreased during the followup, in both patients and carriers. CONCLUSION: Differences between patients and asymptomatic carriers with aPL are at least partially dependent on the proportion of coincident vascular risk factors. The decline in aPL during the followup defines a subgroup in which an anticoagulation suppression assay could be tried. PMID- 15290737 TI - Efficacy of methotrexate in ankylosing spondylitis: a randomized, double blind, placebo controlled trial. AB - OBJECTIVE: To evaluate the efficacy and safety of methotrexate (MTX) compared with placebo in patients with active ankylosing spondylitis (AS). METHODS: This 24 week, double bind, randomized, placebo controlled trial compared the response between MTX 7.5 mg/week or placebo in patients with active AS. The primary outcome measure was a composite index of improvement in 5 of the following scales: severity of morning stiffness, physical well being, the Bath Ankylosing Spondylitis Disease Activity Index (BASDAI), the Bath Ankylosing Spondylitis Functional Index (BASFI), the Health Assessment Questionnaire for Spondyloarthropathies (HAQ-S), and physician and patient global assessment of disease activity. RESULTS: Seventeen patients received MTX and 18 placebo. In the intention-to-treat analysis at 24 weeks, 53% of patients in the MTX group had a treatment response, compared with 17% in the placebo group (p = 0.03). We observed significant improvements with MTX in physical well being (p = 0.009), BASDAI (p = 0.02), BASFI (p = 0.02), physician global assessment (p < 0.001), patient global assessment (p = 0.03), and HAQ-S (p = 0.02). In the adjusted analysis only MTX determined the improvement in the primary outcome. At the end of the trial, one patient with MTX withdrew due to a lack of compliance, and one with placebo due to a lack of efficacy. We did not observe significant differences in rates of side effects between the 2 groups. CONCLUSION: MTX is safe and effective for patients with AS. Longterm studies are needed to evaluate the permanence of its benefit. PMID- 15290738 TI - Compliance with allopurinol therapy among managed care enrollees with gout: a retrospective analysis of administrative claims. AB - OBJECTIVE: Poor compliance with gout medications has been recognized, but seldom studied. We investigated compliance with allopurinol among managed care enrollees suspected to have gout. METHODS: This was a retrospective, administrative claims based analysis of patients with gout. Compliance with allopurinol was measured using prescription-fill dates and days-supplied amounts. Compliance was defined for each prescription period as the presumed use of allopurinol on at least 80% of the days of that period. RESULTS: Of 9482 patients identified, 65.9% filled at least one prescription for allopurinol during the 24 month followup period; 10.4% of allopurinol users filled one prescription and then discontinued use. Of the remaining patients, 13.7% never achieved compliance with therapy; 18% were compliant throughout the entire followup period. Patients were compliant with therapy for an average of 56% of their treatment periods and noncompliant for an average of 44%. In multivariate analysis, male sex was associated with decreased compliance (p < 0.01), although the effect was mitigated by increasing age. For subjects of both sexes, increasing age was associated with increased compliance (p < 0.05). CONCLUSION: Compliance with allopurinol in this population was low. Because untreated gouty arthritis can lead to serious adverse outcomes (such as recurrent gouty arthritis, chronic gouty arthropathy, tophi, and urolithiasis) that are usually avoidable with antihyperuricemic therapy, efforts to achieve better compliance are warranted. PMID- 15290739 TI - Increasing prevalence of gout and hyperuricemia over 10 years among older adults in a managed care population. AB - OBJECTIVE: To determine whether the prevalence of gout and/or clinically significant hyperuricemia increased in a managed care population over 10 years. METHODS: The study was a descriptive analysis utilizing an administrative claims database to ascertain 10-year trends in prevalence of gout and/or hyperuricemia. Prevalence rates were calculated cross-sectionally for each year (1990-99) and expressed/compared as rates per 1000 enrollees. RESULTS: The prevalence of gout and/or hyperuricemia in the overall population increased by about 2 cases per 1000 enrollees over 10 years. In the > 75 year age group, rates increased from 21 per 1000 persons in 1990 to 41 per 1000 in 1999. In the 65-74 year age group, prevalence increased from between 21 and 24 per 1000 persons in the years 1990-92 to over 31 per 1000 during the years 1997-99. Prevalence rates in younger age groups (< 65 years) stayed consistently low during the years under study. There were sex differences in most age groups, with men having the greater burden of disease at every time point. CONCLUSION: Prevalence of gout and/or hyperuricemia in the overall study population increased during the 10-year period. When stratified by age, there were increases in prevalence among groups over age 65 in both sexes. Although gout prevalence increased in both sexes over the 10-year period, men still had most of the burden of disease. In ages younger than 65, men had 4 times higher prevalence than women (4:1 ratio), but in the older age groups (> 65), the gender gap narrowed to 1 woman to every 3 men with gout and/or hyperuricemia (3:1 ratio). PMID- 15290740 TI - Reproducibility of the semiflexed (metatarsophalangeal) radiographic knee position and automated measurements of medial tibiofemoral joint space width in a multicenter clinical trial of knee osteoarthritis. AB - OBJECTIVE: To determine the baseline and longitudinal consistency in reproducibility of the semiflexed metatarsophalangeal (MTP) position in repeat examinations of patients with knee osteoarthritis (OA) recruited for a multicenter clinical trial that terminated within one year (mean duration 0.81 yr), based on precise measurements both of minimum medial tibiofemoral compartment joint space width (JSW) and of tibial inter-rim distance. METHODS: Two technologists from 8 and one technologist from 14 clinical radiology units had received previous training in performing nonfluoroscopic semiflexed MTP knee examinations and in quality control criteria for film acceptance. Patients (N = 402; F = 269) were recruited from 58 rheumatology sites and referred to 22 centers, or "x-ray hubs," across North America. At baseline and at study exit, both knees were x-rayed twice on the same day. All films had quality control, and accepted films were digitized at the Central Radiographic Facility and transmitted to the Central Analysis Facility for computerized measurement of minimum medial compartment JSW and tibial inter-rim distance. JSW loss was calculated in the placebo group for the study period. RESULTS: The median SD of the difference in JSW between same-day test/retest film pairs was 0.9 mm for 767 baseline film pairs (knees with JSW > 0 mm), and 0.08 mm for 631 exit film pairs. JSW reproducibility was unaffected by subject's sex, age, and degree of JSW loss. Among all x-ray hubs, JSW reproducibility was excellent in 14 (SD < 0.1 mm), good in 6 (0.1 < SD < 0.2 mm), and moderate in 2 hubs (0.2 < SD < 0.3 mm). No statistical difference was found in technologists' ability either in positioning OA knees or in their test/retest reproducibility in repositioning joints at baseline and at study exit. JSW did not alter significantly during the study period. CONCLUSION: The protocol for the semiflexed MTP knee position provides a highly reproducible method for anatomically repositioning the knee and for measuring JSW, necessary for OA clinical trials. It is a simple method that can be employed readily at clinical radiology units, as shown by the similarity in JSW precision between x-ray hubs. The results from this large dataset show that throughout the study precise measurements of JSW were obtained from same-day repeat radiographs, findings that together with previous single-center studies confirm the reliability of this method for clinical trial use. PMID- 15290741 TI - Role of EP3 and EP4 prostaglandin receptors in reorganization of the cytoskeleton in mature human osteoclasts. AB - OBJECTIVE: Osteoclasts are central to the pathophysiology of several bone diseases. Prostaglandin E2 (PGE2) is well known to influence osteoclasts indirectly, but its direct action on osteoclasts is still controversial and the relevant receptors are unknown. We investigated the distribution and function of EP receptors in human mature osteoclasts. METHODS: Osteoclasts were extracted from femurs and tibias of human fetuses obtained from legal abortions. In situ hybridization and immunohistochemistry were used to detect the presence of EP1, EP2, EP3, and EP4 receptors on these cells. Actin staining and fluorescent microscopy were used to detect the effects of receptor activation on the cytoskeleton. RESULTS: Only EP3 and EP4 receptors were detected at the RNA and protein level in osteoclasts. These receptors were functional: PGE2 decreased the number of osteoclasts presenting an actin ring; 11-deoxy-PGE1, an EP2 and EP4 agonist, also decreased the number of tartrate-resistant acid phosphatase positive cells with an actin ring; sulprostone, an EP3-specific agonist, had no effect on this variable but increased the number of cells with lamellipodia. CONCLUSION: Mature human osteoclasts present 2 subtypes of EP receptors, namely EP3 and EP4, that mediate different actions of PGE2 on these cells: activation of the EP4 receptors inhibits actin ring formation and activation of the EP3 receptors increases the number of lamellipodia. Activation or inhibition of these receptors by specific agents could be used to study and influence osteoclast function. PMID- 15290742 TI - Cost-effectiveness of hip protectors in the prevention of osteoporosis related hip fractures in elderly nursing home residents. AB - OBJECTIVE: Hip fracture is a common complication of osteoporosis, resulting in significant morbidity and mortality, with a high financial cost to the healthcare system. Hip protectors have been advocated as an effective method to prevent hip fractures in high-risk individuals. This study models the cost-effectiveness of hip protectors in the prevention of osteoporosis related hip fractures in elderly nursing home residents. METHODS: An incremental cost-effectiveness analysis was performed comparing hip protectors to "no treatment" and to "calcium and vitamin D supplements." The study population was a hypothetical cohort of 1000 nursing home residents. A societal perspective, with a lifetime time horizon, was adopted. Data regarding costs, effectiveness, and quality of life measures were collected from the current literature and from Peace Arch Hospital, a community hospital in White Rock, British Columbia, Canada. Sensitivity analysis was performed. RESULTS: Hip protector use was found to be a dominant strategy compared to no treatment and to calcium and vitamin D supplements. Dominance implies lower cost and higher effect, generating cost-effectiveness ratios less than zero. Dominance with respect to cost and effectiveness of hip protectors in preventing hip fractures persisted when the model was subjected to probabilistic sensitivity analysis. CONCLUSION: Cost-effectiveness analysis suggests that hip protectors could save money while preventing hip fractures and improving quality of life in nursing home residents. PMID- 15290743 TI - Celecoxib effectively treats patients with acute shoulder tendinitis/bursitis. AB - OBJECTIVE: Shoulder tendinitis and subacromial bursitis are acute, painful inflammatory musculoskeletal conditions that may recur as a result of overuse. We investigated the efficacy of celecoxib in managing patients with acute shoulder tendinitis/bursitis. METHODS: In this double blind, placebo controlled, parallel group study, patients with acute onset shoulder tendinitis and/or subacromial bursitis were randomized to receive one of: celecoxib 400 mg followed by 200 mg bid, naproxen 500 mg bid, or placebo bid for 14 days. The primary measure of efficacy was the mean reduction in Maximum Pain Intensity at Rest, measured using a 100 mm visual analog scale, from baseline to Days 7 and 14. RESULTS: Of the 306 patients randomized to treatment, 254 completed the study. On Day 7, the mean reduction from baseline in Maximum Pain Intensity at Rest was significantly greater in the celecoxib group compared with the placebo group (-27.7 +/- 2.75 mm vs -18.4 +/- 2.63 mm, respectively; p < 0.05). Similarly, on Day 14, the mean reduction from baseline in Maximum Pain Intensity at Rest was greater in the celecoxib group compared with placebo (-35.0 +/- 3.06 mm vs -25.0 +/- 3.05 mm; p < 0.05). The mean reduction from baseline in Maximum Pain Intensity at Rest was also greater in the naproxen group compared with the placebo group at Day 7 ( 26.4 +/- 2.70 mm vs -18.4 +/- 2.63 mm; p < 0.05), but not on Day 14. Secondary measures of efficacy also showed treatment with celecoxib to be significantly better than placebo treatment and similar to treatment with naproxen. In addition, celecoxib was well tolerated in these patients. CONCLUSION: Celecoxib showed comparable efficacy to naproxen in relieving the pain of patients with acute shoulder tendinitis and/or subacromial bursitis. PMID- 15290744 TI - Depression and fibromyalgia: treatment and cost when diagnosed separately or concurrently. AB - OBJECTIVE: Depression and fibromyalgia (FM) are often coincident. Both syndromes share common symptoms and impose significant economic burdens. This study compared claims for treatment and costs of FM plus depression with those for FM or depression alone. METHODS: Administrative claims data from a national Fortune 100 manufacturer were used to identify 3 mutually exclusive patient cohorts based on claims with a diagnosis for: FM only, depression only, and FM plus depression. A fourth cohort comprised a random sample of 10% of the employer's overall beneficiary population. Cohorts were compared for demographics, comorbid conditions, and healthcare resources utilization. Mean direct (treatment) costs were calculated and indirect (work loss) costs imputed, and these were assessed using Student's t test and Bonferroni adjustments. RESULTS: Mean annual employer payments (direct plus indirect costs) per patient were 5,163 dollars for FM only, 8,073 dollars for depression only, 11,899 dollars for FM plus depression, and 2,486 dollars for the overall sample. Mean incremental employer payments (i.e., above those for the random sample) per patient with FM plus depression were 9,413 dollars, an amount more than the sum of incremental costs for those with FM or depression alone (8,264 dollars). These costs are consistent with costs of other chronic diseases. CONCLUSION: Patients with FM plus depression are high users of healthcare services. As in studies that established relationships between depression and other medical conditions, incremental costs for patients with FM plus depression were more than additive of costs for each condition alone. PMID- 15290745 TI - Autoantibodies in juvenile idiopathic arthritis: glucose-6-phosphate isomerase is not a specific target. AB - OBJECTIVE: Antibodies recognizing the ubiquitous cytosolic enzyme glucose-6 phosphate isomerase (GPI) cause arthritis in the K/BxN mouse model. Studies have shown that these antibodies are not specific for rheumatoid arthritis (RA) in humans. We evaluated GPI as a target of autoantibodies in juvenile idiopathic arthritis (JIA). METHODS: We studied 324 serum and 48 synovial fluid (SF) samples from 103 patients with JIA, 36 with RA, and 8 with arthralgia and 11 controls. Anti-GPI antibodies were assessed by densitometrically evaluating immunoblots and ELISA using native and recombinant GPI. We determined the GPI activity of the soluble antigen in serum and SF. RESULTS: Although several samples contained anti GPI-IgG antibodies, this was not specific for JIA or its subgroups, or for RA. Other proteins in the GPI preparation were also frequently recognized by antibodies. Additionally, we observed increased GPI activity in patients with the systemic manifestation of JIA, but not in other patients. Neither anti-GPI concentrations nor GPI activity were associated with disease activity. CONCLUSION: In addition to the findings in RA, our results indicate that GPI is not a general target of autoantibodies in JIA. PMID- 15290746 TI - Oral health of children with juvenile idiopathic arthritis. AB - OBJECTIVE: s. To estimate dental disease indices and temporomandibular joint (TMJ) dysfunction in children with juvenile idiopathic arthritis (JIA). METHODS: Indices were recorded for dental caries, bacterial dental plaque, gingival inflammation, and TMJ dysfunction in children with JIA and matched controls. RESULTS: There was no significant difference in dental caries experience or the mean plaque score between children with JIA and controls. The mean gingivitis score for the permanent teeth only was significantly greater in the JIA children compared with the controls (p = 0.02). There was a significantly greater proportion of children with JIA with signs of both left and right TMJ dysfunction (p = 0.05, p = 0.02) and symptoms (p = 0.0001, p = 0.0001) compared with controls. CONCLUSION: The low caries rate was attributed to the fact that children with JIA had received preventive dental care from an early age combined with sugar free medication. PMID- 15290747 TI - Skin involvement in juvenile dermatomyositis is associated with loss of end row nailfold capillary loops. AB - OBJECTIVE: To determine associations of dermatological findings in children with juvenile dermatomyositis (JDM) with specific nailfold capillary (NFC) structural abnormalities. METHODS: Sixty newly diagnosed, previously untreated children who met the Bohan-Peter criteria for definite JDM were seen between 1993 and 2002. They were classified by duration of untreated disease and by a disease activity score (DAS) composed of separate subscores for dermatological (DAS skin) and musculoskeletal (DAS muscle) findings. Routine NFC measurements yielded the number of end row loops, arboreal (bushy), and dilated capillary loops. Laboratory testing included muscle enzymes, von Willebrand Factor Antigen, and neopterin. RESULTS: DAS skin, but not DAS muscle, was associated with NFC end row capillary loss (rs = -0.394, p = 0.008). End row capillary loss (reflecting avascularity), arboreal (bushy), and dilated capillary loops (reflecting change in vascular morphology) were each associated with longer untreated symptom duration (rs = -0.401, rs = 0.534, rs = 0.371). CONCLUSION: End row capillary loss measured by NFC was associated with the dermatological, but not musculoskeletal manifestations of JDM, suggesting that damage to skin and muscle may each have distinct disease pathophysiology. In JDM, skin involvement indicates a vasculopathy that progresses with increasing duration of untreated disease and is not revealed by standard serological laboratory tests. We propose that the cutaneous manifestations of JDM are associated with vascular disease and warrant aggressive therapy. PMID- 15290748 TI - Gender and ethnic origin have no effect on longterm outcome of childhood-onset systemic lupus erythematosus. AB - OBJECTIVE: To investigate the associations of gender and ethnic origin with longterm outcome in childhood-onset systemic lupus erythematosus (SLE). METHODS: The study cohort consisted of 51 patients (13 males and 38 females) with childhood-onset SLE followed for > or = 5 years at the British Columbia Children's Hospital in Vancouver. Fifteen patients were Caucasian, 14 Chinese, 9 East Indian, and 13 patients were of other ethnic backgrounds: none was African American or Hispanic. Outcome measures assessed retrospectively included Systemic Lupus International Collaborating Clinics/American College of Rheumatology Damage Index score (SDI), SLE-related death, need for dialysis or renal transplantation, and use of intensive immunosuppressive therapy. A SDI > or = 2 was assigned as poor outcome. RESULTS: The median age at diagnosis was 10.8 years and the median duration of followup was 7.2 years. Five-year survival was 100%; 10-year survival was 85.7% (12/14 patients). The median SDI score at last followup was 2.0 (range 0-9); 2.0 for male, 1.5 for female; 2.0 for Caucasian and 2.03 for non-Caucasian patients. Twenty-six out of 51 patients (51%) had poor outcome (SDI score > 2). Three female patients required dialysis: 2 had subsequent renal transplants. Thirty patients received intensive immunosuppressive therapy. The SDI scores, mortality, and need for intensive immunosuppressive therapy were not influenced by either gender or ethnic origin. CONCLUSION: The median SDI score was high for this cohort with childhood-onset SLE. In contrast to other published data, no association of male gender and/or non-Caucasian ethnicity with poor outcome was found in our study cohort. PMID- 15290749 TI - Granulocyte colony-stimulating factor induces disease flare in patients with antineutrophil cytoplasmic antibody-associated vasculitis. AB - Recombinant human granulocyte colony-stimulating factor (rhuG-CSF) is frequently given to patients with leukopenia or neutropenia caused by various underlying diseases. The treatment with rhuG-CSF is apparently safe, although cutaneous vasculitis and flares in patients with autoimmune diseases are described. We describe 2 patients with histologically proven antineutrophil cytoplasmic antibody-associated systemic vasculitis with disease flares after administration of rhuG-CSF, given to improve collection of stem cells prior to autologous stem cell transplantation. PMID- 15290750 TI - Pigmented villonodular synovitis: a disease in evolution. AB - Pigmented villonodular synovitis (PVNS) is a rare, benign, proliferative disease of the synovial membrane of joints, tendon sheaths, and bursas. Joint aspiration typically yields hemorrhagic or xanthochromic/serosanguinous (brown, murky) fluid. We describe a case of PVNS that presented as an acute, painless, nontraumatic right knee effusion with clear synovial fluid on arthrocentesis. Initial magnetic resonance imaging of the knee revealed no evidence for hemosiderin. A diagnostic arthroscopy and surgical arthrotomy revealed a unique case of PVNS evolving from local to diffuse involvement of the synovium. PMID- 15290751 TI - Septic arthritis caused by Actinobacillus ureae in a patient with rheumatoid arthritis receiving anti-tumor necrosis factor-alpha therapy. AB - Actinobacillus ureae, formerly known as Pasteurella ureae, is a rare human pathogen. We describe a case of septic arthritis and abscess formation caused by this unusual organism in a patient with rheumatoid arthritis, who was being treated with tumor necrosis factor-alpha inhibitors. PMID- 15290752 TI - Rheumatoid neutrophilic dermatitis: rare cutaneous manifestation of rheumatoid arthritis in a patient with palindromic rheumatism. PMID- 15290753 TI - Epidemiology of rheumatic diseases in Greece. PMID- 15290755 TI - Optimal spaced seeds for homologous coding regions. AB - Optimal spaced seeds were developed as a method to increase sensitivity of local alignment programs similar to BLASTN. Such seeds have been used before in the program PatternHunter, and have given improved sensitivity and running time relative to BLASTN in genome-genome comparison. We study the problem of computing optimal spaced seeds for detecting homologous coding regions in unannotated genomic sequences. By using well-chosen seeds, we are able to improve the sensitivity of coding sequence alignment over that of TBLASTX, while keeping runtime comparable to BLASTN. We identify good seeds by first giving effective hidden Markov models of conservation in alignments of homologous coding regions. We give an efficient algorithm to compute the optimal spaced seed when conservation patterns are generated by these models. Our results offer the hope of improved gene finding due to fewer missed exons in DNA/DNA comparison, and more effective homology search in general, and may have applications outside of bioinformatics. PMID- 15290756 TI - Generation of a large gene/protein lexicon by morphological pattern analysis. AB - The identification of gene/protein names in natural language text is an important problem in named entity recognition. In previous work we have processed MEDLINE documents to obtain a collection of over two million names of which we estimate that perhaps two thirds are valid gene/protein names. Our problem has been how to purify this set to obtain a high quality subset of gene/protein names. Here we describe an approach which is based on the generation of certain classes of names that are characterized by common morphological features. Within each class inductive logic programming (ILP) is applied to learn the characteristics of those names that are gene/protein names. The criteria learned in this manner are then applied to our large set of names. We generated 193 classes of names and ILP led to criteria defining a select subset of 1,240,462 names. A simple false positive filter was applied to remove 8% of this set leaving 1,145,913 names. Examination of a random sample from this gene/protein name lexicon suggests it is composed of 82% (+/-3%) complete and accurate gene/protein names, 12% names related to genes/proteins (too generic, a valid name plus additional text, part of a valid name, etc.), and 6% names unrelated to genes/proteins. The lexicon is freely available at ftp.ncbi.nlm.nih.gov/pub/tanabe/Gene.Lexicon. PMID- 15290757 TI - Dynamic integration of gene annotation and its application to microarray analysis. AB - Comprehensive and structured annotations for all genes on a microarray chip are essential for the interpretation of its expression data. Currently, most chip gene annotations are one-line free text descriptions that are often partial, outdated and unsuitable for large-scale data analysis. Therefore the interpretation of microarray gene expression clusters is often limited. Although researchers can manually navigate a collection of databases for better annotations, it is only practical for limited number of genes. Existing meta databases fail to provide comprehensive categorized annotations for hundreds of genes simultaneously. We have developed an automatic system to address this issue. GeneView system monitors various data sources, extracts gene information from a source whenever it is updated, comprehensively matches genes, and integrates them into a central database by categories, such as pathway, genetic mapping, phenotype, expression profile, domain structure, protein interaction, disease association, and references. The system consists of four major components: (1) relational database; (2) data processing; (3) user curation; (4) data query. We evaluated it by analyzing genes on cDNA and Affymetrix Oligo chips. In both cases, the system provided more accurate and comprehensive information than those provided by the vendors or the chip users, and helped identify new common functions among genes in the same expression clusters. PMID- 15290758 TI - Kernel-based self-organized maps trained with supervised bias for gene expression data analysis. AB - Self-Organized Maps (SOMs) are a popular approach for analyzing genome-wide expression data. However, most SOM based approaches ignore prior knowledge about functional gene categories. Also, Self Organized Map (SOM) based approaches usually develop topographic maps with disjoint and uniform activation regions that correspond to a hard clustering of the patterns at their nodes. We present a novel Self-Organizing map, the Kernel Supervised Dynamic Grid Self-Organized Map (KSDG-SOM). This model adapts its parameters in a kernel space. Gaussian kernels are used and their mean and variance components are adapted in order to optimize the fitness to the input density. The KSDG-SOM also grows dynamically up to a size defined with statistical criteria. It is capable of incorporating a priori information for the known functional characteristics of genes. This information forms a supervised bias at the cluster formation and the model owns the potentiality of revising incorrect functional labels. The new method overcomes the main drawbacks of most of the existing clustering methods that lack a mechanism for dynamical extension on the basis of a balance between unsupervised and supervised drives. PMID- 15290759 TI - Developing optimal prediction models for cancer classification using gene expression data. AB - Microarrays can provide genome-wide expression patterns for various cancers, especially for tumor sub-types that may exhibit substantially different patient prognosis. Using such gene expression data, several approaches have been proposed to classify tumor sub-types accurately. These classification methods are not robust, and often dependent on a particular training sample for modelling, which raises issues in utilizing these methods to administer proper treatment for a future patient. We propose to construct an optimal, robust prediction model for classifying cancer sub-types using gene expression data. Our model is constructed in a step-wise fashion implementing cross-validated quadratic discriminant analysis. At each step, all identified models are validated by an independent sample of patients to develop a robust model for future data. We apply the proposed methods to two microarray data sets of cancer: the acute leukemia data by Golub et al. and the colon cancer data by Alon et al. We have found that the dimensionality of our optimal prediction models is relatively small for these cases and that our prediction models with one or two gene factors outperforms or has competing performance, especially for independent samples, to other methods based on 50 or more predictive gene factors. The methodology is implemented and developed by the procedures in R and Splus. The source code can be obtained at http://hesweb1.med.virginia.edu/bioinformatics. PMID- 15290760 TI - Development of image processing program for yeast cell morphology. AB - Every living organism has its own species-specific morphology. Despite the relatively simple ellipsoidal shape of budding yeast cells, the global regulation of yeast morphology remains unclear. In the past, each mutated gene from many mutants with abnormal morphology had to be classified manually. To investigate the morphological characteristics of yeast in detail, we developed a novel image processing program that extracts quantitative data from microscope images automatically. This program extracts data on cells that are often used by yeast morphology researchers, such as cell size, roundness, bud neck position angle, and bud growth direction, and fits an ellipse to the cell outline. We evaluated the ability of the program to extract quantitative parameters. The results suggest that our image-processing program can play a central objective role in yeast morphology studies. PMID- 15290761 TI - Discovering protein-protein interactions. AB - The ongoing genomics and proteomics efforts have helped identify many new genes and proteins in living organisms. However, simply knowing the existence of genes and proteins does not tell us much about the biological processes in which they participate. Many major biological processes are controlled by protein interaction networks. A comprehensive description of protein-protein interactions is therefore necessary to understand the genetic program of life. In this tutorial, we provide an overview of the various current high-throughput methods for discovering protein-protein interactions, covering both the conventional experimental methods and new computational approaches. PMID- 15290762 TI - Dissecting the human spliceosome through bioinformatics and proteomics approaches. AB - The precise excision of introns from mRNAs is executed by the spliceosome, a cellular machinery composed by five small nuclear RNAs and hundreds of proteins. In the last few years, several groups have used proteomics and computational biology tools to characterize the components of the human spliceosome. These reports have identified basically all known splicing factors and several new proteins. The composition of the human spliceosome confirms the link between splicing and other steps in gene expression. Here we comment on these reports and discuss the perspectives for the coming years. PMID- 15290763 TI - An interactive tool for extracting exons and SNP from genomic sequence: isolation of HCN1 and HCN3 ion channel genes. AB - Genome Analyzer (GenoA) with a relational database back-end, was developed to extract information from mammalian genomic sequences. This data mining and visualization tool-set enables laboratory bench scientists to identify and assemble virtual cDNA from genomic exon sequences, and provides a starting point to identify potential alternative splice variants and polymorphisms in silico. The study described in this paper demonstrates the use of GenoA to study human brain hyperpolarization-activated cation channel genes HCN1 and HCN3. PMID- 15290764 TI - Cluster analysis of dynamic parameters of gene expression. AB - Cluster analysis has proven to be a valuable statistical method for analyzing whole genome expression data. Although clustering methods have great utility, they do represent a lower level statistical analysis that is not directly tied to a specific model. To extend such methods and to allow for more sophisticated lines of inference, we use cluster analysis in conjunction with a specific model of gene expression dynamics. This model provides phenomenological dynamic parameters on both linear and non-linear responses of the system. This analysis determines the parameters of two different transition matrices (linear and nonlinear) that describe the influence of one gene expression level on another. Using yeast cell cycle microarray data as test set, we calculated the transition matrices and used these dynamic parameters as a metric for cluster analysis. Hierarchical cluster analysis of this transition matrix reveals how a set of genes influence the expression of other genes activated during different cell cycle phases. Most strikingly, genes in different stages of cell cycle preferentially activate or inactivate genes in other stages of cell cycle, and this relationship can be readily visualized in a two-way clustering image. The observation is prior to any knowledge of the chronological characteristics of the cell cycle process. This method shows the utility of using model parameters as a metric in cluster analysis. PMID- 15290765 TI - Use of gene networks for identifying and validating drug targets. AB - We propose a new method for identifying and validating drug targets by using gene networks, which are estimated from cDNA microarray gene expression profile data. We created novel gene disruption and drug response microarray gene expression profile data libraries for the purpose of drug target elucidation. We use two types of microarray gene expression profile data for estimating gene networks and then identifying drug targets. The estimated gene networks play an essential role in understanding drug response data and this information is unattainable from clustering methods, which are the standard for gene expression analysis. In the construction of gene networks, we use the Bayesian network model. We use an actual example from analysis of the Saccharomyces cerevisiae gene expression profile data to express a concrete strategy for the application of gene network information to drug discovery. PMID- 15290766 TI - A comprehensive whole genome bacterial phylogeny using correlated peptide motifs defined in a high dimensional vector space. AB - As whole genome sequences continue to expand in number and complexity, effective methods for comparing and categorizing both genes and species represented within extremely large datasets are required. Methods introduced to date have generally utilized incomplete and likely insufficient subsets of the available data. We have developed an accurate and efficient method for producing robust gene and species phylogenies using very large whole genome protein datasets. This method relies on multidimensional protein vector definitions supplied by the singular value decomposition (SVD) of a large sparse data matrix in which each protein is uniquely represented as a vector of overlapping tetrapeptide frequencies. Quantitative pairwise estimates of species similarity were obtained by summing the protein vectors to form species vectors, then determining the cosines of the angles between species vectors. Evolutionary trees produced using this method confirmed many accepted prokaryotic relationships. However, several unconventional relationships were also noted. In addition, we demonstrate that many of the SVD-derived right basis vectors represent particular conserved protein families, while many of the corresponding left basis vectors describe conserved motifs within these families as sets of correlated peptides (copeps). This analysis represents the most detailed simultaneous comparison of prokaryotic genes and species available to date. PMID- 15290767 TI - BTEVAL: a server for evaluation of beta-turn prediction methods. AB - This paper describes a web server BTEVAL, developed for assessing the performance of newly developed beta-turn prediction method and it's ranking with respect to other existing beta-turn prediction methods. Evaluation of a method can be carried out on a single protein or a number of proteins. It consists of clean data set of 426 non-homologous proteins with seven subsets of these proteins. Users can evaluate their method on any subset or a complete set of data. The method is assessed at amino acid level and performance is evaluated in terms of Qtotal, Qpredicted, Qobserved and MCC measures. The server also compares the performance of the method with other existing beta-turn prediction methods such as Chou-Fasman algorithm, Thornton's algorithm, GORBTURN, 1-4 and 2-3 Correlation model, Sequence coupled model and BTPRED. The server is accessible from http://imtech.res.in/raghava/bteval/ PMID- 15290768 TI - Predicting risk of coronary artery disease from DNA microarray-based genotyping using neural networks and other statistical analysis tool. AB - This paper presents a novel approach for complex disease prediction that we have developed, exemplified by a study on risk of coronary artery disease (CAD). This multi-disciplinary approach straddles fields of microarray technology and genetics, neural networks (NN), data mining and machine learning, as well as traditional statistical analysis techniques, namely principal components analysis (PCA) and factor analysis (FA). A description of the biological background of the study is given, followed by a detailed description of how the problem has been modeled for analyses by neural networks and FA. A committee learning approach for NN has been used to improve generalization rates. We show that our NN approach is able to yield promising prediction results despite using only the most fundamental network structures. More interestingly, through the statistical analysis process, genes of similar biological functions have been clustered. In addition, a gene marker involved in breaking down lipids has been found to be the most correlated to CAD. PMID- 15290769 TI - Computational strategies for analyzing data in gene expression microarray experiments. AB - Microarray analysis has become a widely used method for generating gene expression data on a genomic scale. Microarrays have been enthusiastically applied in many fields of biological research, even though several open questions remain about the analysis of such data. A wide range of approaches are available for computational analysis, but no general consensus exists as to standard for microarray data analysis protocol. Consequently, the choice of data analysis technique is a crucial element depending both on the data and on the goals of the experiment. Therefore, basic understanding of bioinformatics is required for optimal experimental design and meaningful interpretation of the results. This review summarizes some of the common themes in DNA microarray data analysis, including data normalization and detection of differential expression. Algorithms are demonstrated by analyzing cDNA microarray data from an experiment monitoring gene expression in T helper cells. Several computational biology strategies, along with their relative merits, are overviewed and potential areas for additional research discussed. The goal of the review is to provide a computational framework for applying and evaluating such bioinformatics strategies. Solid knowledge of microarray informatics contributes to the implementation of more efficient computational protocols for the given data obtained through microarray experiments. PMID- 15290770 TI - Design and implementation of a domain specific language for phylogenetic inference. AB - Domain experts think and reason at a high level of abstraction when they solve problems in their domain of expertise. We present the design and motivation behind a domain specific language, called phi LOG, to enable biologists to program solutions to phylogenetic inference problems at a very high level of abstraction. The implementation infrastructure (interpreter, compiler, debugger) for the DSL is automatically obtained through a software engineering framework based on Denotational Semantics and Logic Programming. PMID- 15290771 TI - Bayesian network and nonparametric heteroscedastic regression for nonlinear modeling of genetic network. AB - We propose a new statistical method for constructing a genetic network from microarray gene expression data by using a Bayesian network. An essential point of Bayesian network construction is the estimation of the conditional distribution of each random variable. We consider fitting nonparametric regression models with heterogeneous error variances to the microarray gene expression data to capture the nonlinear structures between genes. Selecting the optimal graph, which gives the best representation of the system among genes, is still a problem to be solved. We theoretically derive a new graph selection criterion from Bayes approach in general situations. The proposed method includes previous methods based on Bayesian networks. We demonstrate the effectiveness of the proposed method through the analysis of Saccharomyces cerevisiae gene expression data newly obtained by disrupting 100 genes. PMID- 15290772 TI - Automated identification of single nucleotide polymorphisms from sequencing data. AB - The single nucleotide polymorphism (SNP) is the difference of the DNA sequence between individuals and provides abundant information about genetic variation. Large scale discovery of high frequency SNPs is being undertaken using various methods. However, the publicly available SNP data sometimes need to be verified. If only a particular gene locus is concerned, locus-specific polymerase chain reaction amplification may be useful. Problem of this method is that the secondary peak has to be measured. We have analyzed trace data from conventional sequencing equipment and found an applicable rule to discern SNPs from noise. The rule is applied to multiply aligned sequences with a trace and the peak height of the traces are compared between samples. We have developed software that integrates this function to automatically identify SNPs. The software works accurately for high quality sequences and also can detect SNPs in low quality sequences. Further, it can determine allele frequency, display this information as a bar graph and assign corresponding nucleotide combinations. It is also designed for a person to verify and edit sequences easily on the screen. It is very useful for identifying de novo SNPs in a DNA fragment of interest. PMID- 15290773 TI - Constrained multiple sequence alignment tool development and its application to RNase family alignment. AB - In this paper, we design a heuristic algorithm of computing a constrained multiple sequence alignment (CMSA for short) for guaranteeing that the generated alignment satisfies the user-specified constraints that some particular residues should be aligned together. If the number of residues needed to be aligned together is a constant alpha, then the time-complexity of our CMSA algorithm for aligning K sequences is O(alphaKn(4)), where n is the maximum of the lengths of sequences. In addition, we have built up such a CMSA software system and made several experiments on the RNase sequences, which mainly function in catalyzing the degradation of RNA molecules. The resulting alignments illustrate the practicability of our method. PMID- 15290774 TI - Exploring alternative transcript structure in the human genome using blocks and InterPro. AB - Understanding how alternative splicing affects gene function is an important challenge facing modern-day molecular biology. Using homology-based, protein sequence analysis methods, it should be possible to investigate how transcript diversity impacts protein function. To test this, high-quality exon-intron structures were deduced for over 8000 human genes, including over 1300 (17 percent) that produce multiple transcript variants. A data mining technique (DiffMotif) was developed to identify genes in which transcript variation coincides with changes in conserved motifs between variants. Applying this method, we found that 30 percent of the multi-variant genes in our test set exhibited a differential profile of conserved InterPro and/or BLOCKS motifs across different mRNA variants. To investigate these, a visualization tool (ProtAnnot) that displays amino acid motifs in the context of genomic sequence was developed. Using this tool, genes revealed by the DiffMotif method were analyzed, and when possible, hypotheses regarding the potential role of alternative transcript structure in modulating gene function were developed. Examples of these, including: MEOX1, a homeobox-containing protein; AIRE, involved in auto-immune disease; PLAT, tissue type plasminogen activator; and CD79b, a component of the B-cell receptor complex, are presented. These results demonstrate that amino acid motif databases like BLOCKS and InterPro are useful tools for investigating how alternative transcript structure affects gene function. PMID- 15290775 TI - Identification of biological relationships from text documents using efficient computational methods. AB - The biological literature databases continue to grow rapidly with vital information that is important for conducting sound biomedical research and development. The current practices of manually searching for information and extracting pertinent knowledge are tedious, time-consuming tasks even for motivated biological researchers. Accurate and computationally efficient approaches in discovering relationships between biological objects from text documents are important for biologists to develop biological models. The term "object" refers to any biological entity such as a protein, gene, cell cycle, etc. and relationship refers to any dynamic action one object has on another, e.g. protein inhibiting another protein or one object belonging to another object such as, the cells composing an organ. This paper presents a novel approach to extract relationships between multiple biological objects that are present in a text document. The approach involves object identification, reference resolution, ontology and synonym discovery, and extracting object-object relationships. Hidden Markov Models (HMMs), dictionaries, and N-Gram models are used to set the framework to tackle the complex task of extracting object-object relationships. Experiments were carried out using a corpus of one thousand Medline abstracts. Intermediate results were obtained for the object identification process, synonym discovery, and finally the relationship extraction. For the thousand abstracts, 53 relationships were extracted of which 43 were correct, giving a specificity of 81 percent. These results are promising for multi-object identification and relationship finding from biological documents. PMID- 15290776 TI - Fast large scale oligonucleotide selection using the longest common factor approach. AB - We present a fast method that selects oligonucleotide probes (such as DNA 25 mers) for microarray experiments on a truly large scale. For example, reliable oligos for human genes can be found within four days, a speedup of one to two orders of magnitude compared to previous approaches. This speed is attained by using the longest common substring as a specificity measure for candidate oligos. We present a space- and time-efficient algorithm, based on a suffix array with additional information, to compute matching statistics (lengths of longest matches) between all candidate oligos and all remaining sequences. With the matching statistics available, we show how to incorporate constraints such as oligo length, melting temperature, and self-complementarity into the selection process at a postprocessing stage. As a result, we can now design custom oligos for any sequenced genome, just as the technology for on-site chip synthesis is becoming increasingly mature. PMID- 15290777 TI - Computational assignment of protein backbone NMR peaks by efficient bounding and filtering. AB - NMR resonance assignment is one of the key steps in solving an NMR protein structure. The assignment process links resonance peaks to individual residues of the target protein sequence, providing the prerequisite for establishing intra- and inter-residue spatial relationships between atoms. The assignment process is tedious and time-consuming, which could take many weeks. Though there exist a number of computer programs to assist the assignment process, many NMR labs are still doing the assignments manually to ensure quality. This paper presents a new computational method based on the combination of a suite of algorithms for automating the assignment process, particularly the process of backbone resonance peak assignment. We formulate the assignment problem as a constrained weighted bipartite matching problem. While the problem, in the most general situation, is NP-hard, we present an efficient solution based on a branch-and-bound algorithm with effective bounding techniques using two recently introduced approximation algorithms. We also devise a greedy filtering algorithm for reducing the search space. Our experimental results on 70 instances of (pseudo) real NMR data derived from 14 proteins demonstrate that the new solution runs much faster than a recently introduced (exhaustive) two-layer algorithm and recovers more correct peak assignments than the two-layer algorithm. Our result demonstrates that integrating different algorithms can achieve a good tradeoff between backbone assignment accuracy and computation time. PMID- 15290778 TI - Prediction of protein coarse contact maps. AB - Prediction of topological representations of proteins that are geometrically invariants can contribute towards the solution of fundamental open problems in structural genomics like folding. In this paper we focus on coarse grained protein contact maps, a representation that describes the spatial neighborhood relation between secondary structure elements such as helices, beta sheets, and random coils. Our methodology is based on searching the graph space. The search algorithm is guided by an adaptive evaluation function computed by a specialized noncausal recursive connectionist architecture. The neural network is trained using candidate graphs generated during examples of successful searches. Our results demonstrate the viability of the approach for predicting coarse contact maps. PMID- 15290779 TI - Efficient reconstruction of haplotype structure via perfect phylogeny. AB - Each person's genome contains two copies of each chromosome, one inherited from the father and the other from the mother. A person's genotype specifies the pair of bases at each site, but does not specify which base occurs on which chromosome. The sequence of each chromosome separately is called a haplotype. The determination of the haplotypes within a population is essential for understanding genetic variation and the inheritance of complex diseases. The haplotype mapping project, a successor to the human genome project, seeks to determine the common haplotypes in the human population. Since experimental determination of a person's genotype is less expensive than determining its component haplotypes, algorithms are required for computing haplotypes from genotypes. Two observations aid in this process: first, the human genome contains short blocks within which only a few different haplotypes occur; second, as suggested by Gusfield, it is reasonable to assume that the haplotypes observed within a block have evolved according to a perfect phylogeny, in which at most one mutation event has occurred at any site, and no recombination occurred at the given region. We present a simple and efficient polynomial-time algorithm for inferring haplotypes from the genotypes of a set of individuals assuming a perfect phylogeny. Using a reduction to 2-SAT we extend this algorithm to handle constraints that apply when we have genotypes from both parents and child. We also present a hardness result for the problem of removing the minimum number of individuals from a population to ensure that the genotypes of the remaining individuals are consistent with a perfect phylogeny. Our algorithms have been tested on real data and give biologically meaningful results. Our webserver (http://www.cs.columbia.edu/compbio/hap/) is publicly available for predicting haplotypes from genotype data and partitioning genotype data into blocks. PMID- 15290780 TI - CUBIC: identification of regulatory binding sites through data clustering. AB - Transcription factor binding sites are short fragments in the upstream regions of genes, to which transcription factors bind to regulate the transcription of genes into mRNA. Computational identification of transcription factor binding sites remains an unsolved challenging problem though a great amount of effort has been put into the study of this problem. We have recently developed a novel technique for identification of binding sites from a set of upstream regions of genes, that could possibly be transcriptionally co-regulated and hence might share similar transcription factor binding sites. By utilizing two key features of such binding sites (i.e. their high sequence similarities and their relatively high frequencies compared to other sequence fragments), we have formulated this problem as a cluster identification problem. That is to identify and extract data clusters from a noisy background. While the classical data clustering problem (partitioning a data set into clusters sharing common or similar features) has been extensively studied, there is no general algorithm for solving the problem of identifying data clusters from a noisy background. In this paper, we present a novel algorithm for solving such a problem. We have proved that a cluster identification problem, under our definition, can be rigorously and efficiently solved through searching for substrings with special properties in a linear sequence. We have also developed a method for assessing the statistical significance of each identified cluster, which can be used to rule out accidental data clusters. We have implemented the cluster identification algorithm and the statistical significance analysis method as a computer software CUBIC. Extensive testing on CUBIC has been carried out. We present here a few applications of CUBIC on challenging cases of binding site identification. PMID- 15290781 TI - Efficient inference of haplotypes from genotypes on a pedigree. AB - We study haplotype reconstruction under the Mendelian law of inheritance and the minimum recombination principle on pedigree data. We prove that the problem of finding a minimum-recombinant haplotype configuration (MRHC) is in general NP hard. This is the first complexity result concerning the problem to our knowledge. An iterative algorithm based on blocks of consecutive resolved marker loci (called block-extension) is proposed. It is very efficient and can be used for large pedigrees with a large number of markers, especially for those data sets requiring few recombinants (or recombination events). A polynomial-time exact algorithm for haplotype reconstruction without recombinants is also presented. This algorithm first identifies all the necessary constraints based on the Mendelian law and the zero recombinant assumption, and represents them using a system of linear equations over the cyclic group Z2. By using a simple method based on Gaussian elimination, we could obtain all possible feasible haplotype configurations. A C++ implementation of the block-extension algorithm, called PedPhase, has been tested on both simulated data and real data. The results show that the program performs very well on both types of data and will be useful for large scale haplotype inference projects. PMID- 15290782 TI - Two notes on genome rearrangement. AB - A central problem in genome rearrangement is finding a most parsimonious rearrangement scenario using certain rearrangement operations. An important problem of this type is sorting a signed genome by reversals and translocations (SBRT). Hannenhalli and Pevzner presented a duality theorem for SBRT which leads to a polynomial time algorithm for sorting a multi-chromosomal genome using a minimum number of reversals and translocations. However, there is one case for which their theorem and algorithm fail. We describe that case and suggest a correction to the theorem and the polynomial algorithm. The solution of SBRT uses a reduction to the problem of sorting a signed permutation by reversals (SBR). The best extant algorithms for SBR require quadratic time. The common approach to solve SBR is by finding a safe reversal using the overlap graph or the interleaving graph of a permutation. We describe a family of signed permutations which proves a quadratic lower bound on the number of affected vertices in the overlap/interleaving graph during any optimal sorting scenario. This implies, in particular, an Omega(n3) lower bound for Bergeron's algorithm. PMID- 15290783 TI - RAPTOR: optimal protein threading by linear programming. AB - This paper presents a novel linear programming approach to do protein 3 dimensional (3D) structure prediction via threading. Based on the contact map graph of the protein 3D structure template, the protein threading problem is formulated as a large scale integer programming (IP) problem. The IP formulation is then relaxed to a linear programming (LP) problem, and then solved by the canonical branch-and-bound method. The final solution is globally optimal with respect to energy functions. In particular, our energy function includes pairwise interaction preferences and allowing variable gaps which are two key factors in making the protein threading problem NP-hard. A surprising result is that, most of the time, the relaxed linear programs generate integral solutions directly. Our algorithm has been implemented as a software package RAPTOR-RApid Protein Threading by Operation Research technique. Large scale benchmark test for fold recognition shows that RAPTOR significantly outperforms other programs at the fold similarity level. The CAFASP3 evaluation, a blind and public test by the protein structure prediction community, ranks RAPTOR as top 1, among individual prediction servers, in terms of the recognition capability and alignment accuracy for Fold Recognition (FR) family targets. RAPTOR also performs very well in recognizing the hard Homology Modeling (HM) targets. RAPTOR was implemented at the University of Waterloo and it can be accessed at http://www.cs.uwaterloo.ca/~j3xu/RAPTOR_form.htm. PMID- 15290784 TI - Recognizing complex, asymmetric functional sites in protein structures using a Bayesian scoring function. AB - The increase in known three-dimensional protein structures enables us to build statistical profiles of important functional sites in protein molecules. These profiles can then be used to recognize sites in large-scale automated annotations of new protein structures. We report an improved FEATURE system which recognizes functional sites in protein structures. FEATURE defines multi-level physico chemical properties and recognizes sites based on the spatial distribution of these properties in the sites' microenvironments. It uses a Bayesian scoring function to compare a query region with the statistical profile built from known examples of sites and control nonsites. We have previously shown that FEATURE can accurately recognize calcium-binding sites and have reported interesting results scanning for calcium-binding sites in the entire Protein Data Bank. Here we report the ability of the improved FEATURE to characterize and recognize geometrically complex and asymmetric sites such as ATP-binding sites and disulfide bond-forming sites. FEATURE does not rely on conserved residues or conserved residue geometry of the sites. We also demonstrate that, in the absence of a statistical profile of the sites, FEATURE can use an artificially constructed profile based on a priori knowledge to recognize the sites in new structures, using redoxin active sites as an example. PMID- 15290785 TI - Data mining tools for biological sequences. AB - We describe a methodology, as well as some related data mining tools, for analyzing sequence data. The methodology comprises three steps: (a) generating candidate features from the sequences, (b) selecting relevant features from the candidates, and (c) integrating the selected features to build a system to recognize specific properties in sequence data. We also give relevant techniques for each of these three steps. For generating candidate features, we present various types of features based on the idea of k-grams. For selecting relevant features, we discuss signal-to-noise, t-statistics, and entropy measures, as well as a correlation-based feature selection method. For integrating selected features, we use machine learning methods, including C4.5, SVM, and Naive Bayes. We illustrate this methodology on the problem of recognizing translation initiation sites. We discuss how to generate and select features that are useful for understanding the distinction between ATG sites that are translation initiation sites and those that are not. We also discuss how to use such features to build reliable systems for recognizing translation initiation sites in DNA sequences. PMID- 15290786 TI - Assessing the impact of predictive biosimulation on drug discovery and development. AB - Systems biology is creating a context for interpreting the vast amounts of genomic and proteomic data being produced by pharmaceutical companies in support of drug development. While major data collection efforts capitalize on technical advances in miniaturization and automation and represent an industrialization of existing laboratory research, the transition from mental models to predictive computer simulations is setting the pace for advances in this field. This article addresses current approaches to the mathematical modeling of biological systems and assesses the potential impact of predictive biosimulation on drug discovery and development. PMID- 15290787 TI - From immunoinformatics to immunomics. PMID- 15290788 TI - Bioinformatics meets proteomics--bridging the gap between mass spectrometry data analysis and cell biology. AB - Proteomics research programs typically comprise the identification of protein content of any given cell, their isoforms, splice variants, post-translational modifications, interacting partners and higher-order complexes under different conditions. These studies present significant analytical challenges owing to the high proteome complexity and the low abundance of the corresponding proteins, which often requires highly sensitive and resolving techniques. Mass spectrometry plays an important role in proteomics and has become an indispensable tool for molecular and cellular biology. However, the analysis of mass spectrometry data can be a daunting task in view of the complexity of the information to decipher, the accuracy and dynamic range of quantitative analysis, the availability of appropriate bioinformatics software and the overwhelming size of data files. The past ten years have witnessed significant technological advances in mass spectrometry-based proteomics and synergy with bioinformatics is vital to fulfill the expectations of biological discovery programs. We present here the technological capabilities of mass spectrometry and bioinformatics for mining the cellular proteome in the context of discovery programs aimed at trace-level protein identification and expression from microgram amounts of protein extracts from human tissues. PMID- 15290789 TI - [Cement-free hip cups. Current status]. AB - Important criteria for stable cup fixation are the type of anchor system and stabilizers,cup form and the material and surface structure. Different fixation systems are manifest in pressfit and threaded cups. Pressfit implants are oversized and lead to equatorial jamming. For additional fixation, and to improve stability, screws, pegs, rings, fins, spikes or hollow cylinders are used. In threaded cups, thread geometry is decisive for the cup's performance during the screw-in process and positioning. The hemispheric shape of the cups requires less bone resection and the position of the implant can be arbitrarily selected. The conical shape guarantees high tilting stability. Most implants are made of pure titanium or a titanium-aluminum alloy. A rough surface area - produced by corundum blasting, titanium-plasma spray, titanium balls, nets or other grid designs - is essential for osseointegration. The results of second generation pressfit and threaded cups with 10 year survival rates of 93-100% are persuasive. PMID- 15290790 TI - Estrogenic potential of halogenated derivatives of nonylphenol ethoxylates and carboxylates. AB - Halogenated derivatives of nonylphenol and of its alkylates are generated during drinking water disinfection and treatment procedures. In this paper we analyze the potential of these compounds to interact with the estrogen receptor and to activate hormone-regulated gene promoters. We used the recombinant yeast assay (RYA) and the human breast cancer cell MCF7 proliferation assay for both estrogenic and antiestrogenic activities and the enzyme-linked receptor assay to examine in vitro binding to the receptor. Many nonylphenol derivatives were very weak estrogens in our functional tests when compared to nonylphenol while retaining a substantial affinity for the estrogen receptor in vitro. Antiestrogenicity tests demonstrated that brominated nonylphenol and most of the carboxylated compounds studied here behaved as estrogenic antagonists in the RYA. We also detected an increased cytotoxicity for the carboxylated derivatives in both yeast and mammalian cells. We conclude that derivatization may mask the apparent estrogenicity of nonylphenol, but the resulting compounds still represent a potential hazard since they are still able to bind the estrogen receptor and to influence the physiological response to estrogens. Our results also illustrate the advantage of combining different methods to assay estrogenicity of unknown substances. PMID- 15290791 TI - Resolvins, docosatrienes, and neuroprotectins, novel omega-3-derived mediators, and their aspirin-triggered endogenous epimers: an overview of their protective roles in catabasis. AB - The molecular basis for the beneficial impact of essential omega-3 fatty acids is of considerable interest. Recently, novel mediators generated from eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) that displayed potent bioactions were first identified in resolving inflammatory exudates [J. Exp. Med. 192 (2000) 1197; J. Exp. Med. 196 (2002) 1025] and in tissues enriched with DHA [J. Exp. Med. 196 (2002) 1025; J. Biol. Chem. 278 (2003) 14677]. The trivial names Resolvin (resolution phase interaction products) and docosatrienes were introduced for the bioactive compounds belonging to these novel series because they demonstrate potent anti-inflammatory and immunoregulatory actions. The compounds derived from eicosapentaenoic acid carrying potent biological actions (i.e., 1-10 nM range) are designated E series, given their EPA precursor, and denoted as Resolvins of the E series (Resolvin E1 or RvE1), and those biosynthesized from the precursor docosahexaenoic acid are Resolvins of the D series (Resolvin D1 or RvD1). Bioactive members from DHA with conjugated triene structures are docosatrienes (DT) that are immunoregulatory [J. Exp. Med. 196 (2002) 1025; J. Biol. Chem. 278 (2003) 14677], and neuroprotective [J. Biol. Chem., 278 (2003) 43807; Proc. Natl. Acad. Sci. U.S.A. [submitted for publication]] and are termed neuroprotectins. The specific receptors for these novel bioactive products from omega-3 EPA and DHA are abbreviated Resolvin D receptors (i.e., ResoDR1), Resolvin E receptor (ResoER1; RER1), and neuroprotectin D receptors (NPDR), respectively, in recognition of their respective cognate ligands. Aspirin treatment impacts biosynthesis of these compounds and a related series by triggering endogenous formation of the 17R-D series Resolvins and docosatrienes. These novel epimers are denoted as aspirin triggered (AT)-RvDs and -DTs, and possess potent anti-inflammatory actions in vivo essentially equivalent to their 17S series pathway products. Here, we provide a syntomy overview of the formation and actions of these newly uncovered pathways and products as well as highlight their role(s) as endogenous protective mediators generated in anti-inflammation and catabasis. PMID- 15290792 TI - Invasive and angiogenic phenotype of MCF-7 human breast tumor cells expressing human cyclooxygenase-2. AB - To evaluate the direct effect of human cyclooxygenase-2 (hCox-2) on human breast tumor cell proliferation, invasion, and angiogenesis, hCox-2 cDNA was transfected into slow growing, non-metastatic MCF-7 human breast tumor cells that express low levels of Cox-2. Two stable transfectant clones, designated MCF-7/hCox-2 clones 8 and 10, had significantly decreased (P < 0.05) doubling time, with two-fold greater number of cells during exponential growth compared to the MCF-7/vector control. Proliferation of both of the MCF-7/hCox-2 clones was significantly inhibited in a time- and dose-dependent manner by celecoxib. The MCF-7/hCox-2 clones 8 and 10 formed larger and greater numbers of colonies in soft agar than the MCF-7/vector control, with a corresponding increased invasion across an artificial Matrigel basement membrane in response to recombinant human epidermal growth factor (hEGF). The MCF-7/hCox-2 clones 8 and 10 had higher mRNA levels of two splice variants of vascular endothelial growth factor (VEGF), V145 and V165. These results demonstrate that hCox-2 directly increases breast tumor cell proliferation, stimulates invasion across a basement membrane, and induces synthesis of specific heparin binding splice variants of VEGF. PMID- 15290793 TI - Unicystic ameloblastoma--use of Carnoy's solution after enucleation. AB - A retrospective analysis of 29 patients with a histologically confirmed diagnosis of unicystic ameloblastoma is reported with special emphasis on a treatment regime employing enucleation and application of Carnoy's solution. Despite the finding that 93% of lesions exhibited mural invasion, a recurrence rate of 10% after treatment is reported, suggesting a possible benefit of Carnoy's solution against recurrence. PMID- 15290794 TI - A note on the applied use of MDL approximations. AB - An applied problem is discussed in which two nested psychological mod-els of retention are compared using minimum description length (MDL).The standard Fisher information approximation to the normalized maximum likelihood is calculated for these two models, with the result that the full model is assigned a smaller complexity, even for moderately large samples. A geometric interpretation for this behavior is considered, along with its practical implications. PMID- 15290795 TI - Optimal reduced-set vectors for support vector machines with a quadratic kernel. AB - To reduce computational cost, the discriminant function of a support vector machine (SVM) should be represented using as few vectors as possible. This problem has been tackled in different ways. In this article,we develop an explicit solution in the case of a general quadratic kernel k(x. x') = (C + D xT x')2. For a given number of vectors, this solution provides the best possible approximation and can even recover the discriminant function if the number of used vectors is large enough. The key idea is to express the inhomogeneous kernel as a homogeneous kernel ona space having one dimension more than the original one and to follow the approach of Burges (1996). PMID- 15290797 TI - Mentors need help. PMID- 15290796 TI - Private land burial clarified. PMID- 15290798 TI - Nurses must know the steps to ensure safety patients. PMID- 15290799 TI - NMC investigates confusing titles. PMID- 15290800 TI - Guidance to cut blood transfusion errors. PMID- 15290801 TI - There is far more to Christian practices. PMID- 15290802 TI - As doctor writes prescription, drug company writes a check. PMID- 15290803 TI - Pfizer to pay $430 million over promoting drug to doctors. PMID- 15290804 TI - South Africa rejects use of AIDS drug for women. PMID- 15290805 TI - Choosing death. PMID- 15290806 TI - As gene test menu grows, who gets to choose? PMID- 15290807 TI - Results of drug trials can mystify doctors through omission: some professional groups demand public listing of all test outcomes. PMID- 15290808 TI - India emerges as new drug proving ground. PMID- 15290809 TI - To close gaps in care, more health plans ask about race. PMID- 15290810 TI - In kidney quest, new rules boost chances for blacks; reform seeks to close gap in transplant wait times; worries about a downside. PMID- 15290811 TI - After Medtronic lobbying push, the FDA had change of heart. PMID- 15290812 TI - John C. Fletcher; biomedical ethicist, former Episcopal priest. PMID- 15290814 TI - Infertile ground is sown in Brazil; politicians trade sterilizations for votes. PMID- 15290813 TI - Detainees' medical files shared; Guantanamo interrogators access criticized. PMID- 15290815 TI - Drugmakers prefer silence on test data; firms violate US law by not registering trials. PMID- 15290816 TI - How many is too many? Risks spur effort to curb multiple births. PMID- 15290817 TI - Elementary computation of object approach by wide-field visual neuron. AB - An essential function of the brain is to detect threats, such as those posed by objects or predators on a collision course. A wide-field, movement-sensitive visual neuron in the brain of the locust was studied by presenting simulated approaching, receding, and translating objects. The neuron's responses could be described simply by multiplying the velocity of the image edge (dtheta/dtau) with an exponential function of the size of the object's image on the retina (e-alpha theta). Because this product peaks before the image reaches its maximum size during approach, this neuron can anticipate collision. The neuron's activity peaks approximately when the approaching object reaches a certain angular size. Because this neuron receives distinct inputs about image size and velocity, the dendritic tree of a single neuron may function as a biophysical device that can carry out a multiplication of two independent input signals. PMID- 15290818 TI - Causes of thrombocytopenia in triplet pregnancies. PMID- 15290819 TI - Progesterone and preterm birth. PMID- 15290820 TI - Prophylactic progesterone to prevent preterm birth. PMID- 15290821 TI - Supplemental progesterone to prevent preterm birth. PMID- 15290822 TI - Progesterone and preterm birth. PMID- 15290823 TI - Computer modeling of shoulder dystocia. PMID- 15290824 TI - Improving the management of opioid-dependent pregnancies. PMID- 15290825 TI - Peritoneal nonclosure in cesarean section. PMID- 15290826 TI - Is first trimester measurement of sex hormone binding globulin a possible screening test for gestational diabetes mellitus. PMID- 15290827 TI - Re: Stage-based treatment of twin-twin transfusion syndrome. PMID- 15290829 TI - Selective bibliography of Manfred Zimmermann 1960-2004. PMID- 15290828 TI - Staple yourself to an order. 1992. PMID- 15290830 TI - Abstracts of the ISBER Annual Meeting: Biodiversity and International Regulations. May 4-7, 2003, Philadelphia, Pennsylvania, USA. PMID- 15290831 TI - Corticosteroids, Kounis syndrome and the treatment of refractory vasospastic angina. PMID- 15290832 TI - Tattooing and civilizing processes: body modification as self-control. PMID- 15290833 TI - Disability history: no longer hidden [Review of: Paul Longmore, Why I burned my book and other essays on disability. Philadelphia: Temple University Press, 2003]. PMID- 15290834 TI - Mark Twain's " How to remove warts and tattoo marks" authenticated. PMID- 15290835 TI - Henry James on the great war: a letter recovered from the Mercure de France. PMID- 15290836 TI - [Thomas Fincke and trigonometry]. AB - Thomas Fincke (January 6th, 1561 - April 24th, 1650), born in Flensburg (Germany), was one of the very most important and significant scientists in Denmark during the seventeenth century, a mathematician and astrologer and physician in the beginning of modern science, a representative of humanism and an influentual academic organizer. He studied in Strasbourg (since 1577) and Padua (since 1583) and received his M.D. in Basel (1587), he practised as a physician throughtout his life (since 1587 or 1590) and became a professor at Copenhagen (1591). But he was best known because of his Geometriae rotundi libri XIIII (1583), a famous book on plane and spherical trigonometry, based not on Euclid but on Petrus Ramus. In this influentual work, in which Fincke introduced the terms tangent and secant and probable first noticed the Law of Tangents and the so-called Newton-Oppel-Mauduit-Simpson-Mollweide-Gauss-formula, he showed himself to be ,,abreast of the mathematics of his time". PMID- 15290837 TI - Proceedings of the International Congress on Valvular Heart Disease. 8-10 May 2003, Krakow, Poland. PMID- 15290838 TI - Drugs and demons. PMID- 15290839 TI - Healthy design. PMID- 15290840 TI - Richard Darling, President and CEO of the FAIR Foundation. PMID- 15290841 TI - Thomas F Mancuso. PMID- 15290842 TI - Endocrine-related resources for the National Institutes of Health. PMID- 15290843 TI - Bibliography of toxinology. PMID- 15290844 TI - [LADD syndrome with QT prolongation]. PMID- 15290846 TI - Opalescent appearance of an IgG1 antibody at high concentrations and its relationship to noncovalent association. AB - PURPOSE: Therapeutic antibodies are often formulated at a high concentration where they may have an opalescent appearance. The aim of this study is to understand the origin of this opalescence, especially its relationship to noncovalent association and physical stability. METHODS: The turbidity and the association state of an IgG1 antibody were investigated as a function of concentration and temperature using static and dynamic light scattering, nephelometric turbidity, and analytical ultracentrifugation. RESULTS. The antibody had increasingly opalescent appearance in the concentration range 5-50 mg/ml. The opalescence was greater at refrigerated temperature but was readily reversible upon warming to room temperature. Turbidity measured at 25 degrees C was linear with concentration, as expected for Rayleigh scatter in the absence of association. In the concentration range 1-50 mg/ml, the weight average molecular weights were close to that expected for a monomer. Zimm plot analysis of the data yielded a negative second virial coefficient, indicative of attractive solute solute interactions. The hydrodynamic diameter was independent of concentration and remained unchanged as a function of aging at room temperature. CONCLUSIONS: The results indicate that opalescent appearance is not due to self-association but is a simple consequence of Rayleigh scatter. Opalescent appearance did not result in physical instability. PMID- 15290845 TI - RAGE: a novel target for drug intervention in diabetic vascular disease. AB - At high levels as seen in diabetes, glucose reacts with and forms adducts (advanced glycation end products; AGEs) on macromolecules including proteins and DNA, eliciting cellular dysfunction and leading to vascular disease. The major means is through cellular receptors; the best characterized is the receptor for advanced glycation end products (RAGE). Accumulation of both AGE/RAGE in addition to other identified ligands of RAGE, including S100/calgranulins, is the hallmark of this receptor in disease pathogenesis. Blockade of ligand-receptor interaction directly at the protein level, or transgenetically, prevents development of micro vascular (nephropathy) and macro vascular (atherosclerosis/restenosis) disease in small animal models. Furthermore, allelic variants of RAGE exist that alter the protein function and gene expression, which may further affect disease outcome. In conclusion, RAGE is a target for drug development to prevent vascular disease in diabetic and nondiabetic subjects. PMID- 15290847 TI - Rapid characterization of amyloid-beta side-chain oxidation by tandem mass spectrometry and the scoring algorithm for spectral analysis. AB - PURPOSE: Amyloid-beta (Abeta) is a self-aggregating protein found in senile plaques in Alzheimer's disease (AD) brain and is thought to play a major role in the disease process. Oxidative stress may be a predominant cause of the formation of these Abeta aggregates. This study aims at identifying possible sites of copper-catalyzed oxidation of Abeta1-40 using liquid chromatography tandem mass spectrometry (LC/MS/MS) and scoring algorithm for spectral analysis (SALSA). Traditionally, identification of post-translational modifications by tandem mass spectrometric analysis requires users to inspect manually thousands of MS/MS spectra, which can be a tedious and time-consuming process. With the use of SALSA, users can automatically search for post-translational modifications based on the spacing of the m/z values associated with the ion series of an amino acid sequence. METHODS: Abeta1-40 was subjected to copper-catalyzed oxidative stress. LC/MS/MS and SALSA analyses were used to determine the sites of post translational modification within the tryptic fragments. RESULTS: Oxidation was found to occur preferentially at the histidine residues Hisl3 and Hisl4 and at the methionine residue (Met35) of Abeta1-40. CONCLUSIONS: The combination of LC/MS/MS and SALSA searches could dramatically improve the efficiency and accuracy of determining the specific sites of oxidation of in vitro, copper oxidized Abeta1-40 as well as other oxidized proteins. PMID- 15290848 TI - HPLC detection and quantification of radiolytic products of eight beta-blockers irradiated in the solid state and hypotheses on their origins. AB - PURPOSE: The radiolytic products of eight beta-blockers were studied in order to understand the mechanisms of irradiation of drugs in the solid state. METHODS: The drugs were analyzed by high-performance liquid chromatography coupled to a diode array detector in order to observe the degradation of the main compound after irradiation and in order to study the nonvolatile final products on more concentrated solutions of irradiated drugs. RESULTS: The first test assessed that the main compound was not significantly degraded after gamma irradiation for any of the eight beta-blockers. A more complete study, which consisted on separating the nonvolatile products and on quantifying them, indicated first that the radiolytic products could reach the number of 14 and moreover that some could exceed the 0.1% threshold at 30 kGy. Eventually, radiolytic yields were compared with radical yields previously determined. CONCLUSIONS: The sensitivity of the first test can be discussed. It seems that, to study the feasibility of the radiosterilization, a complete study of the products of degradation is needed. Moreover, no correlation between radical and final products could be established, which denies that the former would be the precursors of the latter. PMID- 15290849 TI - Suppression of the progress of disseminated pancreatic cancer cells by NK4 plasmid DNA released from cationized gelatin microspheres. AB - PURPOSE: NK4, composed of the NH2-terminal hairpin and subsequent four-kringle domains of hepatocyte growth factor (HGF), acts as a potent angiogenesis inhibitor. This study is an investigation to evaluate the feasibility of controlled release of NK4 plasmid DNA in suppressing the tumor growth. Controlled release by a biodegradable hydrogel enabled the NK4 plasmid DNA to exert the tumor suppression effects. METHODS: Biodegradable cationized gelatin microspheres were prepared for the controlled release of an NK4 plasmid DNA. The cationized gelatin microspheres incorporating NK4 plasmid DNA were subcutaneously injected to tumor-bearing mice to evaluate the suppressive effects on tumor angiogenesis and growth. RESULTS: The cationized gelatin microspheres incorporating NK4 plasmid DNA could release over 28 days as a result of microspheres degradation. The injection of cationized gelatin microspheres incorporating NK4 plasmid DNA into the subcutaneous tissue of mice inoculated with pancreatic cancer cells prolonged their survival time period. An increase in the tumor number was suppressed to a significantly greater extent than free NK4 plasmid DNA. The controlled release of NK4 plasmid DNA suppressed angiogenesis and increased the cell apoptosis in the tumor tissue while it enhanced and prolonged the NK4 protein level in the blood circulation. CONCLUSIONS: We conclude that the controlled release technology is promising to enhance the tumor suppression effects of NK4 plasmid DNA. PMID- 15290850 TI - Pulmonary delivery of deslorelin: large-porous PLGA particles and HPbetaCD complexes. AB - PURPOSE: To compare the systemic delivery of deslorelin following intratracheal administration of different deslorelin formulations. The formulations included dry powders of deslorelin, large-porous deslorelin-poly(lactide-co-glycolide) (PLGA) particles, and small conventional deslorelin-PLGA particles. Also, solution formulations of deslorelin and deslorelin-hydroxy-propyl-beta cyclodextrin (HPbetaCD) complexes were tested. METHODS: Dry powders of deslorelin, large-porous (mean diameter, 13.8 microm; density, 0.082 g/cc), and small conventional (mean diameter, 2.2 microm; density, 0.7 g/cc) deslorelin-PLGA particles and solutions of deslorelin with or without HPbetaCD were administered intratracheally to Sprague-Dawley rats. Blood samples were collected at 3 h, 1, 3, and 7 days postdosing, and plasma deslorelin concentrations were determined using enzyme immunoassay. At the end of 7 days, lungs were isolated, and bronchoalveolar lavage fluid was collected and analyzed for deslorelin. RESULTS: At the end of 7 days, deslorelin plasma concentrations in the large-porous deslorelin-PLGA particle group were 120-fold and 2.5-fold higher compared to deslorelin powder and small conventional deslorelin-PLGA particles, respectively. Co-administration of HPbetaCD resulted in 2-, 3-, and 3-fold higher plasma deslorelin concentrations at 3 h, 1 and 3 days, respectively, compared to deslorelin solution. On day 7, deslorelin concentrations in bronchoalveolar lavage fluid as well as plasma were in the order: large porous particles > small conventional particles > deslorelin-HPbetaCD solution > deslorelin powder > deslorelin solution. CONCLUSIONS: Large-porous deslorelin PLGA particles can sustain deslorelin delivery via the deep lungs. Co-administration of HPbetaCD enhances the systemic delivery of deslorelin. The pulmonary route is useful as a noninvasive alternative for the systemic delivery of deslorelin. PMID- 15290851 TI - Cyclodextrins in nasal delivery of low-molecular-weight heparins: in vivo and in vitro studies. AB - PURPOSE: To test the hypothesis that cyclodextrins reversibly enhance nasal absorption of low-molecular-weight heparins (LMWHs) and to investigate the mechanisms by which cyclodextrins enhance LMWH absorption via the nose. METHODS: Absorption of LMWHs was studied by measuring plasma anti-factor Xa activity after nasal administration of various LMWH formulations to anesthetized rats. In vivo reversibility studies were performed to investigate if the effects of cyclodextrins are reversible and diminish with time. The absorption-enhancing mechanisms of cyclodextrins were investigated in cell culture model. The transport of enoxaparin and mannitol, changes in transepithelial electrical resistance (TEER), and distribution of tight junction protein ZO-1 were investigated. RESULTS: Formulations containing 5% dimethyl-beta-cyclodextrin (DMbetaCD) produced the highest increase in the bioavailability of LMWH preparations tested. In vivo reversibility studies with 5% DMbetaCD showed that the effect of the absorption enhancer at the site of administration diminished with time. Transport studies using 16HBE14o(-) cells demonstrated that the increase in the permeability of enoxaparin and mannitol, reduction in TEER, and the changes in the tight junction protein ZO-1 distribution produced by 5% DMbetaCD were much greater than those produced by beta-cyclodextrin (betaCD) or hydroxyl-propyl-beta-cyclodextrin (HPbetaCD). CONCLUSIONS: Of the cyclodextrins tested, DMbetaCD was the most efficacious in enhancing absorption of LMWHs both in vivo and in vitro. The study also suggests that cyclodextrins enhance nasal drug absorption by opening of cell-cell tight junctions. PMID- 15290852 TI - Transdermal delivery of the synthetic cannabinoid WIN 55,212-2: in vitro/in vivo correlation. AB - PURPOSE: The aim of the current investigation was to evaluate the percutaneous absorption of the synthetic cannabinoid WIN 55,212-2 in vitro and in vivo. METHODS: The in vitro permeation studies of WIN 55,212-2 in human skin, hairless guinea pig skin, a polymer membrane with adhesive, and a skin/polymer membrane composite were conducted in flowthrough diffusion cells. The pharmacokinetic parameters for WIN 55,212-2 were determined after intravenous administration and topical application of Hill Top Chambers and transdermal therapeutic systems (TTS) in guinea pigs. RESULTS: The in vitro permeation studies indicated that the flux of WIN 55,212-2 through hairless guinea pig skin was 1.2 times more than that through human skin. The flux of WIN 55,212-2 through human and guinea pig skin was not significantly higher than that through the corresponding skin/polymer membrane composites. The mean guinea pig steady-state plasma concentrations after topical 6.3 cm2 chamber and 14.5 cm2 TTS patch applications were 5.0 ng/ml and 8.6 ng/ml, respectively. CONCLUSIONS: The topical drug treatments provided significant steady-state plasma drug levels for 48 h. The observed in vivo results from the Hill Top Chambers and TTS patches in the guinea pigs were in good agreement with the predicted plasma concentrations from the in vitro data. PMID- 15290853 TI - Physicochemical evaluation, in vitro human skin diffusion, and concurrent biotransformation of 3-O-alkyl carbonate prodrugs of naltrexone. AB - PURPOSE: The purpose of this study was to evaluate the physicochemical properties and in vitro human skin diffusion of the 3-O-alkyl carbonate prodrugs of naltrexone (NTX). METHODS: Melting points and heats of fusion (deltaHf) were determined using differential scanning calorimetry. In vitro human skin permeation rates of NTX and its prodrugs were measured using a flowthrough diffusion cell system. Drug disposition in the skin was quantified at the end of the diffusion experiment. The solubilities of the drugs were determined in mineral oil and isotonic buffer. Partitioning of the prodrugs from vehicle to skin was determined using isolated sheets of human stratum corneum (SC). RESULTS: All the prodrugs hydrolyzed to NTX on passing through the skin, and the methyl NTX-3-O-carbonate (ME-NTX) provided the highest NTX flux, apparent permeability coefficient (Kp), and calculated relative thermodynamic activity from the melting point and deltaHf. The ME-NTX SC/vehicle partition coefficient was the highest of the prodrug series, although similar to the NTX SC/vehicle partition coefficient value. The shortest chain prodrugs underwent the highest extent of bioconversion to NTX upon passing through the skin. CONCLUSIONS: Within this 3-O-alkyl carbonate prodrug series, the shortest chain prodrug was the most skin-permeable compound with the highest partition coefficient and a significant extent of bioconversion. PMID- 15290854 TI - Technetium-99m-Labeled N-(2-hydroxypropyl) methacrylamide copolymers: synthesis, characterization, and in vivo biodistribution. AB - PURPOSE: To synthesize novel technetium-99m (99mTc)-labeled N-(2-hydroxypropyl) methacrylamide (HPMA) copolymers and characterize the effect of charge and molecular weight on their biodistribution in SCID mice. METHODS: Electronegative and neutral 7-kDa, 21-kDa, and 70-kDa HPMA copolymers containing a 99mTc chelating comonomer, bearing N-omega-bis(2-pyridylmethyl)-L-lysine (DPK), were synthesized by free-radical precipitation copolymerization. The copolymers were labeled via 99mTc tricarbonyl chelation to DPK-bearing comonomer. They were characterized by side-chain content, molecular weight, molecular weight distribution, radiochemical purity, and labeling stability. Scintigraphic images were obtained during the first 90 min and at 24 h postintravenous injection in SCID mice. At 24 h, organ radioactivity was determined from necropsy tissue counting. RESULTS: 99mTc-labeled HPMA copolymers showed greater than 90% stability over a 24-h challenge with cysteine and histidine. Scintigraphic images and the necropsy data showed that the negatively charged copolymers were eliminated from the body significantly faster than the neutral copolymers in a size-dependent manner. CONCLUSIONS: To facilitate clinical scintigraphic imaging, stable chelation of 99mTc may be achieved by incorporation of a DPK-bearing comonomer into the HPMA backbone. Electronegative and neutral 99mTc-labeled HPMA copolymers of 7, 21, and 70 kDa show significant variation in organ biodistribution in SCID mice. 99mTc-labeled HPMA copolymers could be used as diagnostic agents and to study pharmacokinetics of delivery systems based on these copolymers. PMID- 15290855 TI - Lectins as endocytic ligands: an assessment of lectin binding and uptake to rabbit conjunctival epithelial cells. AB - PURPOSE: To investigate the binding and uptake pattern of three plant lectins in rabbit conjunctival epithelial cells (RCECs) with respect to their potential for enhancing cellular macromolecular uptake. METHODS: Three fluorescein-labeled plant lectins (Lycoperison esculentum, TL; Solanum tuberosum, STL; and Ulex europaeus 1, UEA-1) were screened with respect to time-, concentration-, and temperature-dependent binding and uptake. Chitin (30 mg/ml) and L-alpha-fucose (10 mM) were used as inhibitory sugars to correct for nonspecific binding of TL or STL and UEA-1, respectively. Confocal microscopy was used to confirm internalization of STL. RESULTS: The binding and uptake of all three lectins in RCECs was time-dependent (reaching a plateau at 1-2 h period) and saturable at 1 h period. The rank order of affinity constants (km) was STL>TL>UEA-1 with values of 0.39>0.48>4.81 microM, respectively. However, maximal, specific binding/uptake potential was in the order UEA-1>STL>TL with values of 53.7, 52.3, and 15.0 nM/mg of cell protein, respectively. Lectins showed temperature dependence in their uptake, with STL exhibiting the highest endocytic capacity. Internalized STL was visualized by confocal microscopy to be localized to the cell membrane and cytoplasm. CONCLUSION: Based on favorable binding and uptake characteristics, potato lectin appears to be a useful candidate for further investigation as an ocular drug delivery system. PMID- 15290856 TI - Effect of preparation method on physical properties of amorphous trehalose. AB - PURPOSE: To determine the effect of preparation method on the physical properties of amorphous trehalose. METHODS: Amorphous anhydrous trehalose was prepared by four different methods. viz., freeze-drying, spray-drying, dehydration, and melt quenching. The glass transition temperature (Tg), enthalpic relaxation behavior, and crystallization were studied by differential scanning calorimetry, whereas X ray diffractometry was used for phase identification. The rate and extent of water uptake at different relative humidity values were also obtained. RESULTS: Though the enthalpic relaxation and crystallization behaviors were influenced by the method of preparation of amorphous trehalose, the Tg and fragility were not. The phase prepared by dehydration showed the highest enthalpic recovery at Tg, indicating that aging may have occurred during preparation. Among the four methods used, trehalose prepared by dehydration had the highest tendency to crystallize, whereas there was no crystallization in melt-quenched amorphous trehalose. The method of preparation influenced not only the rate and extent of water sorption but also the phase crystallized. Water vapor sorption removed the effects of structural history in the amorphous phase formed by dehydration. CONCLUSIONS: The method of preparation strongly influenced the pharmaceutically relevant properties of amorphous trehalose. The resistance to crystallization can be rank ordered as trehalose prepared by dehydration < freeze-dried approximately spray-dried < melt-quenched. The rate of water sorption can be rank ordered as trehalose prepared by dehydration < freeze-dried < spray-dried. PMID- 15290857 TI - Development of dividable one-step dry-coated tablets (dividable-OSDRC) and their evaluation as a new platform for controlled drug release. AB - PURPOSE: The purpose of this study is to develop novel dividable coated tablets that retain their characteristics even after they are divided. METHODS: We prepared dividable one-step dry-coated tablets (dividable-OSDRC) using our own manufacturing process with double structure punches. The release pattern of the dividable-OSDRC with hydroxypropyl methylcellulose (HPMC) or methacrylic acid copolymer LD (Eudragit) as an outer layer was investigated before and after the division, and dissolution profiles were statistically compared using difference factor f1 and similarity factor f2. RESULTS: The dividable-OSDRC with HPMC for sustained-release (compression pressure, 150 MPa; crashing strength, 6.1 N; friability, 0.05%; CV of divided tablet weight, 7.8%) showed statistically equivalent release patterns between the one-half and the whole (f1, 13.9; f2, 55.5) and between the one-half and the two-halves (5.5, 72.5). The surface area of the tablets affected the sustained-release profiles. Furthermore, the tablets made with Eudragit LD for timed-release (150 MPa. 12.8 N, 0.18%, 9.6%) also showed approximated release patterns before and after the division. CONCLUSIONS: We proved that dividable-OSDRC maintain their release characteristics after they are divided. We conclude that the dividable-OSDRC could be used as a new platform for the controlled release of drugs. PMID- 15290858 TI - Preparation and drug loading of poly(ethylene glycol)-block-poly(epsilon caprolactone) micelles through the evaporation of a cosolvent azeotrope. AB - PURPOSE: The aim of this work was to study the assembly, drug loading, and stability of poly(ethylene glycol)-block-poly(epsilon-caprolactone) (PEG-b-PCL) micelles. METHODS: Three PEG-b-PCL compositions with PCL number average molecular weights of 1000, 2500, and 4000 g/mol were used. The assembly of PEG-b-PCL micelles, induced by the addition of water to acetonitrile (ACN), was characterized with 1H nuclear magnetic resonance spectroscopy (1H-NMR) and dynamic light scattering (DLS) with and without the presence of fenofibrate, a poorly water-soluble drug. PEG-b-PCL micelles with encapsulated fenofibrate were prepared through the removal of a negative ACN-water azeotrope under reduced pressure. Fenofibrate content was measured using reverse-phase high-performance liquid chromatography (HPLC), whereas the kinetic stability of PEG-b-PCL micelles with and without encapsulated fenofibrate was evaluated using size exclusion chromatography (SEC). RESULTS: The critical water content (CWC), the water content at which amphiphilic block copolymer (ABC) micelle assembly begins, was determined using DLS and ranged from 10% to 30% water, depending on both PCL molecular weight and PEG-b-PCL concentration. As the water content was increased, the PEG-b-PCL unimers assembled into swollen structures with hydrodynamic diameters ranging from 200 to 800 nm. The 1H-NMR peaks associated with the PCL block exhibited line-broadening, following the addition of D2O, indicating that the PCL blocks reside in the core of the PEG-b-PCL micelle. With further addition of water, the PCL cores collapsed to form fairly monodisperse PEG-b-PCL micelles (20-60 nm). In the presence of fenofibrate, the CWC value was lowered, perhaps due to hydrophobic interactions of fenofibrate and the PCL block. Further addition of water and subsequent evaporation of the negative ACN-water azeotrope resulted in fenofibrate-loaded PEG-b-PCL micelles of under 50 nm. The extent of fenofibrate encapsulation was dependent on PCL block size. At a polymer concentration of 1.0 mg/ml, PEG-b-PCL (5000:4000) and (5000:2500) micelles could encapsulate more than 90% of the initial loading level of fenofibrate, whereas PEG-b-PCL (5000:1000) micelles encapsulate only 28%. SEC experiments revealed that PEG-b-PCL (5000:4000) and (5000:2500) micelles eluted intact, indicating kinetic stability, whereas PEG-b-PCL (5000:1000) micelles eluted primarily as unimers. CONCLUSIONS: PEG-b-PCL in ACN assembles with fenofibrate into drug loaded polymeric micelles with the addition of water and the subsequent removal of a negative ACN-water azeotrope. PMID- 15290859 TI - Rheological characterization of topical carbomer gels neutralized to different pH. AB - PURPOSE: The primary objective of this study is to perform detailed and extensive rheological characterization of rheology of carbomer (Carbopol) microgels formulated using a solvent system typically used in topical gel formulations. Solvents like glycerin and propylene glycol can alter rheology and drug delivery characteristics of topical gels owing to their different viscosities and due to the change in solvent-polymer and solvent-solvent interactions. METHODS: Aqueous gels with different pH were prepared by dissolving cross-linked Carbopol polymers in a co-solvent system comprising water, propylene glycol, and glycerol and subsequently neutralizing the carboxylic groups of the polymers with triethanolamine (TEA). Oscillatory, steady, and transient shear measurements were performed to measure viscoelastic properties, temperature dependency, yield strength, and thixotropy of carbomer pharmaceutical gels. RESULTS: The topical pharmaceutical gels exhibit remarkable temperature stability. Flow curves obtained at different temperatures indicate Carbopol microgels show much more pseudoplastic behavior (lower power law index) compared to Carbopol gels dissolved only in water. Substantial yield strength is required to break the microgel network of the topical gels. The gel samples exhibit modest thixotropy at higher deformation rates. CONCLUSIONS: The theological behavior of the Carbopol microgels do not change appreciably in the pH range 5.0-8.0, and the gels can be used as effective dermatological base for topical applications. PMID- 15290860 TI - Aerosol dispersion of respirable particles in narrow size distributions produced by jet-milling and spray-drying techniques. AB - PURPOSE: To examine the effect of particle size and morphology on aerosol dispersion using jet-milled and spray-dried mannitol particles in narrow size distributions within the respirable range. METHODS: Particle size and morphology were examined by laser diffraction and scanning electron microscopy, respectively. Aerosol dispersion was examined using a cascade impactor with a preseparator operating at a flow rate of 60 L/min, using two inhaler devices: Rotahaler (low-resistance device) and Inhalator (high-resistance device). Powder flow was examined using static and dynamic methods (Carr's compressibility index and vibrating spatula, respectively). RESULTS: Narrow size distributions of jet milled and spray-dried particles were produced (d50% = 1.4 to 10.3 microm, GSD = 1.8 to 2.1, and d50% = 1.6 to 7.5 microm; GSD = 1.5 to 1.9, respectively). All particles were highly crystalline. Differences in particle shape were observed between jet-milled and spray-dried particles. Higher fine particle fraction (FPF) and relative fine particle fraction (FPFrel) (greater aerosol dispersion) and lower geometric standard deviation (GSD) (less variation) were obtained using particles with d50% between 2 and 5 microm. Higher mass median aerodynamic diameter were obtained with larger d50%. Spray-dried particles produced greater aerosol dispersion compared with jet-milled particles. Greater aerosol dispersion was obtained using the Inhalator than the Rotahaler. CONCLUSIONS: Small changes in the particle size within the 1-10-microm range produced a major impact in the aerosol dispersion of jet-milled and spray-dried particles. Even in these narrow size ranges, aggregation plays an important role in aerosol dispersion. PMID- 15290861 TI - Aerosol dispersion of respirable particles in narrow size distributions using drug-alone and lactose-blend formulations. AB - PURPOSE: To examine the effect of formulation type on the aerosolization of respirable particles in narrow size distributions. METHODS: Aerosol dispersion of two formulation types (drug alone and 2% w/w drug-lactose blends) containing micronized or spray-dried fluticasone propionate (FP) particles (d50% = 1.3 to 9.6 microm, GSD = 1.8 to 2.2) were examined using cascade impaction at 60 l/min with low and high resistance inhaler devices: Rotahaler and Inhalator, respectively. RESULTS: The aerosol dispersion of FP particles was significantly affected by the particle size, particle type, inhaler device, and formulation type. Interactions were observed between all factors. Generally, greater powder entrainment was obtained with smaller d50%. Higher emitted doses were obtained from drug-alone formulations of spray-dried FP particles and lactose blends of micronized FP particles. Greater aerosol dispersion of spray-dried FP particles was obtained using lactose-blend formulations with d50% around 4 microm. Greater aerosol dispersion of micronized FP particles was obtained using formulations of drug alone. Larger d50% produced larger mass median aerodynamic diameters. CONCLUSIONS: Small changes in the particle size within the 1-10-microm range exerted a major influence on aerosol dispersion of jet-milled and spray-dried FP particles using drug-alone and lactose-blend formulations. PMID- 15290862 TI - Reverse iontophoresis as a noninvasive tool for lithium monitoring and pharmacokinetic profiling. AB - PURPOSE: Transdermal iontophoresis was investigated as a noninvasive tool for drug monitoring and pharmacokinetic profiling. Lithium, a frequently monitored drug, was used as a model. The objectives were a) to demonstrate the linear dependence of the iontophoretic extraction flux of lithium on the subdermal concentration of the drug, b) to evaluate the capacity of iontophoresis to monitor sudden changes in the subdermal level, c) to investigate the utility of reverse iontophoresis as a tool in pharmacokinetic studies, and d) to examine the validity of an internal standard calibration procedure to render the method completely noninvasive. METHODS: Transdermal, iontophoretic extraction was performed in vitro using dermatomed pig-ear skin. The subdermal solution consisted of a physiological buffer containing lithium chloride at concentrations in the therapeutic range and two putative internal standards, sodium and potassium, at fixed physiological levels. The subdermal concentration of lithium was changed either in a stepwise fashion or by simulating one of two pharmacokinetic profiles. RESULTS: Lithium was extracted via electromigration to the cathode. A excellent correlation between subdermal lithium concentration and iontophoretic extraction flux was observed. Iontophoresis tracked sudden concentration changes and followed kinetic profiles. In addition, the effective elimination rate constant could be directly, and noninvasively, estimated from the extraction flux data. CONCLUSIONS: Reverse iontophoresis is a potentially useful and noninvasive tool for lithium monitoring. PMID- 15290863 TI - Significant role of liver sinusoidal endothelial cells in hepatic uptake and degradation of naked plasmid DNA after intravenous injection. AB - PURPOSE: Uptake and degradation of naked plasmid DNA (pDNA) by liver sinusoidal endothelial cells (LSECs) were investigated. METHODS: Tissue distribution and intrahepatic localization were determined after an intravenous injection of 111In or 32P-labeled pDNA into rats. Cellular uptake and degradation of fluorescein- or 32P-labeled pDNA were evaluated using primary cultures of rat LSECs. RESULTS: Following intravenous injection, pDNA was rapidly eliminated from the circulation and taken up by the liver. Fractionation of liver-constituting cells by centrifugal elutriation revealed a major contribution of LSECs to the overall hepatic uptake of pDNA. Confocal microscopic study confirmed intracellular uptake of pDNA in cultured LSECs. Apparent cellular association of pDNA was similar at 37 degrees C and 4 degrees C. However, trichloroacetic acid (TCA) precipitation experiments showed the TCA-soluble radioactivity in the culture medium increased in an accumulative manner at 37 degrees C. Involvement of a specific mechanism was demonstrated, as the uptake of pDNA was significantly inhibited by excess unlabeled pDNA and some polyanions (polyinosinic acid, dextran sulfate, heparin) but not by others (polycytidylic acid, dextran). These inhibitors also reduced the amount of TCA-soluble radioactivity in the culture medium. CONCLUSION. These results suggest that LSECs efficiently ingested and rapidly degraded naked pDNA in vivo and in vitro and released the degradation products into the extracellular space. PMID- 15290864 TI - A new potent hFIX plasmid for hemophilia B gene therapy. AB - PURPOSE: The purpose of this work was to construct and characterize a new potent hFIX plasmid, p2SV-hFIX, which has two hFIX expression units containing the SV40 promoter/enhancer for hemophilia B gene therapy. METHODS: p1SV-hFIX was constructed by insertion of amplified hFIX cDNA at the ECORI and Xbal sites of pSI expression vector containing simian virus 40 (SV40) promoter/enhancer. To construct p2SV-hFIX, the hFIX expression cassette was isolated from p1SV-hFIX by digestion with restriction enzymes, and the purified expression cassette was inserted at the BglII site of another plSV-hFIX. The gene expression of p1SV hFIX, p2SV-hFIX, and a plasmid containing a liver-specific apoE enhancer and alpha antitrypsin promoter, pAAV-hAAT-hFIX. were evaluated in various cell lines using polyethylenimine (PEI) as a gene carrier in vitro. RESULTS: The construction of p1SV-hFIX and p2SV-hFIX were confirmed by restriction enzyme studies. The transfection efficiency of p2SV-hFIX was 3.83-fold and 7.16-fold higher than that of pAAV-hAAT-hFIX in C2C12 and NIH3T3 cells, respectively. p2SV hFIX also showed higher transfection efficiency than p1SV-hFIX in both cells. CONCLUSIONS: In accordance with these results, p2SV-hFIX is a new potent hFIX plasmid that can be transfected in various cells. Systemic delivery of p2SV-hFIX via intravenous or intramuscular injection is feasible for treatment of hemophilia B. PMID- 15290865 TI - Surface plasmon resonance studies of the direct interaction between a drug/intestinal brush border membrane. AB - PURPOSE: We describe here a new method to estimate the oral drug permeability from the small intestine using surface plasmon resonance (SPR) technology. The interaction between drugs and brush border membrane (BBM) surfaces immobilized on biosensor chip were evaluated by measuring the SPR response signal. METHODS: BBM vesicles, isolated from Sprague-Dawley (SD) rats, were immobilized onto the L1 chip composed of dextran derivatives with modified lipophilic residues. A SPR (BIAcore 3000) was used with L1 chip, and it was carried out in a running buffer, HEPES-buffered saline containing 0.1% DMSO. Fourteen drugs for the SPR experiments were flowed over the BBM immobilized L1 chip, and the response levels according to the BBM surfaces were evaluated directly in a continuous flow system. RESULTS: The immobilized BBM surface on L1 chip was very stable, and it was regenerated by injecting a new BBM vesicle solution. It was evident that drug binding events, using BBM surfaces, directly provides information that predicts the Fa value in human for transcellularly absorbed drugs. The throughput to assay each drug at a single concentration is 100 drugs for 24 h. CONCLUSIONS: The interaction between drugs and small intestinal surfaces was successfully assayed using SPR technology, and this SPR analysis exhibited advantages: lack of labeling requirements, the real-time acquirement of various results, and the repeated use for various drugs. PMID- 15290866 TI - Composition and surface charge of DNA-loaded microparticles determine maturation and cytokine secretion in human dendritic cells. AB - PURPOSE: Biodegradable microparticles prepared from poly(lactide) (PLA) and poly(lactide-co-glycolide) (PLGA) have been shown to be promising carrier systems for vaccine delivery. Here, we have investigated the capacity of different PLA and PLGA microparticle formulations to induce stimulation of human blood monocyte derived dendritic cells (DCs). METHODS: Stimulation of human derived dendritic cells by plain microparticles were compared with microparticles loaded with plasmid DNA or double-stranded salmon DNA either by encapsulation or adsorption to the surface of cationic microparticles. Stimulation of DCs was monitored by the up-regulation of surface maturation markers CD83 and CD86 and the secretion of IL-12 and TNF-alpha. RESULTS: Slowly degrading PLA microparticles did not induce any detectable stimulation or activation of DCs. In contrast, fast degrading PLGA microparticles were able to influence DC maturation and cytokine secretion dependent on their surface charge. Anionic PLGA microparticles induced an up-regulation of CD83 and high TNF-alpha secretion, which was further enhanced up to the level of the potent stimulator lipopolysaccharide (LPS) when plasmid DNA was encapsulated. Moreover, the secretion of significant amounts of IL-12 was observed. Cationic PLGA microparticles induced an up-regulation of CD86 and moderate TNF-alpha secretion, but no IL-12 secretion, with no additional effects in the presence of plasmid DNA. CONCLUSIONS: The data suggest that the composition and charge of biodegradable DNA-loaded microparticles profoundly influences maturation and cytokine secretion in DCs. Thus, the individual formulation of microparticles used as a vaccine carrier system might considerably influence the profile of the immune response. PMID- 15290867 TI - Cellular uptake but low permeation of human calcitonin-derived cell penetrating peptides and Tat(47-57) through well-differentiated epithelial models. AB - PURPOSE: To investigate whether cell penetrating peptides (CPP) derived from human calcitonin (hCT) possess, in addition to cellular uptake, the capacity to deliver their cargo through epithelial barriers. METHODS: Cellular uptake of hCT(9-32) and permeation of six hCT-derived peptides, namely, hCT(9-32), hCT(12 32), hCT(15-32), hCT(18-32), hCT(21-32), and a random sequence of hCT(9-32) were evaluated in fully organized confluent Madin-Darby canine kidney (MDCK), Calu-3, and TR146 cell culture models. For comparison, Tat(47-57) and penetratin(43-58) were investigated. The peptides were N-terminally labeled with carboxyfluorescein (CF). Uptake in the well-differentiated epithelial models was observed by confocal laser scanning microscopy (CLSM), whereas permeation through the models was analyzed by reversed-phase (RP)-HPLC. RESULTS: In MDCK epithelium hCT(9-32), Tat(47-57) and penetratin(43-58) demonstrated punctuated cytoplasmic distribution. In Calu-3, Tat(47-57) and penetratin(43-58) were simultaneously localized in a punctuated cytoplasmic and paracellular distribution, whereas hCT(9-32) showed strict paracellular distribution. By contrast, in TR146 cells, Tat(47-57) was located strictly paracellularily, whereas penetratin(43-58) showed a punctuated cytoplasmic pattern and hCT(9-32) both. The transepithelial permeability of all tested peptides and their cargo was lower than that of paracellular markers. CONCLUSIONS: The CPP uptake pattern depends on both the type of peptide and the cell culture model. In general, the investigated CPP have no apparent potential for systemic drug delivery across epithelia. Nevertheless, distinct patterns of cellular distribution may offer a potential for localized epithelial delivery. PMID- 15290868 TI - Uptake of the carborane derivative of cholesteryl ester by glioma cancer cells is mediated through LDL receptors. AB - PURPOSE: This study was to elucidate the mechanism of cellular uptake of cholesteryl 1,12-dicarba-closo-dodecaboranel-carboxylate (BCH), a new anti-cancer carborane derivative of cholesteryl ester, by glioma cancer cells. METHODS: BCH (solubilized with liposomal formulation) was incubated with SF-763 and SF-767 glioma cell lines in the presence of different amounts of monoclonal anti-LDL receptor antibody for cellular uptake studies. Various amounts of lipoprotein deficient serum (LPDS) were also used during the uptake. The effect of calcium ion and low temperature on BCH uptake were investigated. In addition, the transfer of BCH from liposomes to low-density lipoprotein (LDL) particles was determined through gradient ultracentrifugation. RESULTS: BCH uptake by these glioma cells was significantly inhibited by the monoclonal antibody. The uptake by both cell lines was reversely correlated with the amount of LPDS. The presence of calcium ion promoted the BCH uptake, whereas the low temperature decreased the BCH uptake. After 16 h incubation, about 46% of BCH was transferred from liposomes to LDL particles. CONCLUSIONS: These results strongly suggested that the cellular uptake of BCH (in liposomal formulation) by SF-763 and SF-767 glioma cell lines is mediated through LDL receptors. PMID- 15290869 TI - Combined effects of multiple flavonoids on breast cancer resistance protein (ABCG2)-mediated transport. AB - PURPOSE: The purpose of this study was to determine the dynamic parameter (EC50) of flavonoids apigenin, biochanin A, chrysin, genistein, kaempferol, hesperetin, naringenin, and silymarin for breast cancer resistance protein (BCRP) inhibition when used alone, and to evaluate their potential interactions (additive, synergistic, or antagonistic) with regards to BCRP inhibition when used in multiple-flavonoid combinations. METHODS: The effects of flavonoids on BCRP mediated transport were examined by evaluating their effects on mitoxantrone accumulation and cytotoxicity in MCF-7 MX100 cells overexpressing BCRP. The EC50 values of these flavonoids for increasing mitoxantrone accumulation were estimated using a Hill equation. The potential interactions among multiple flavonoids with regard to BCRP inhibition were assessed by isobologram and Berenbaum's interaction index methods. RESULTS: The EC50 values of these flavonoids for increasing mitoxantrone accumulation ranged from 0.39+/-0.13 microM to 33.7+/-2.78 microM. Quantitative analysis of the combined effects of multiple flavonoids on mitoxantrone accumulation indicated that these flavonoids act additively in inhibiting BCRP when given as 2-, 3-, 5-, or 8-flavonoid combinations with equimolar concentrations of all constituents. The results of the mitoxantrone cytotoxicity studies were consistent with these findings. CONCLUSIONS: The additive effects of multiple flavonoids for BCRP inhibition suggests that prediction of BCRP-mediated food (herbal product)-drug interactions should also take into consideration the presence of multiple flavonoids and provides a rationale for using "flavonoid cocktails" as a potential approach for multidrug resistance reversal in cancer treatment. PMID- 15290870 TI - Mishandling of the therapeutic peptide glucagon generates cytotoxic amyloidogenic fibrils. AB - PURPOSE: Some therapeutic peptides exhibit amyloidogenic properties that cause insolubility and cytotoxicity against neuronal cells in vitro. Here, we characterize the conformational change in monomeric therapeutic peptide to its fibrillar aggregate in order to prevent amyloidogenic formation during clinical application. METHODS: Therapeutic peptides including glucagon, porcine secretin, and salmon calcitonin were dissolved in acidic solution at concentrations ranging from 1 mg/ml to 80 mg/ml and then aged at 37 degrees C. Amyloidogenic properties were assessed by circular dichroism (CD), electron microscopy (EM), staining with beta-sheet-specific dyes, and size-exclusion chromatography (SEC). Cytotoxic characteristics were determined concomitantly. RESULTS: By aging at 2.5 mg/ml or higher for 24 h, monomeric glucagon was converted to fibrillar aggregates consisting of a beta-sheet-rich structure with multimeric states of glucagon. Although no aggregation was observed by aging at the clinical concentration of 1 mg/ml for 1 day, 30-day aging resulted in the generation of fibrillar aggregates. The addition of anti-glucagon serum significantly inhibited fibrillar conversion of monomeric glucagon. Glucagon fibrils induced significant cell death and activated an apoptotic enzyme, caspase-3, in PC12 cells and NIH-3T3 cells. Caspase inhibitors attenuated this toxicity in a dose-dependent manner, indicating the involvement of apoptotic signaling pathways in the fibrillar formation of glucagon. On the contrary to glucagon, salmon calcitonin exhibited aggregation at a much higher concentration of 40 mg/ml and secretin showed no aggregation at the concentration as high as 75 mg/ml. CONCLUSIONS: These results indicated that glucagon was self-associated by its beta-sheet-rich intermolecular structure during the aging process under concentrated conditions to induce fibrillar aggregates. Glucagon has the same amyloidogenic propensities as pathologically related peptides such as beta-amyloid (Abeta)1-42 and prion protein fragment (PrP)106-126 including conformational change to a beta-sheet rich structure and cytotoxic effects by activating caspases. These findings suggest that inappropriate preparation and application of therapeutic glucagon may cause undesirable insoluble products and side effects such as amyloidosis in clinical application. PMID- 15290871 TI - P-glycoprotein (P-gp/MDR1)-mediated efflux of sex-steroid hormones and modulation of P-gp expression in vitro. AB - PURPOSE: To determine whether female sex-steroid hormones and their metabolites can modulate P-glycoprotein (P-gp) expression and catalytic activity and to investigate P-gp mediated transport of these sex-steroids across MDR1-transfected Madine-Darby canine kidney (MDCK) cells. METHODS: Changes in P-gp protein and MDR1 mRNA expression levels were examined in the presence of various estrogens and progestins after a 72-h induction period in the LS180 human colon carcinoma cell line via Western blotting and semiquantitative Reverse-transcription polymerase chain reaction (RT-PCR), respectively. Concentration-dependent stimulation of vanadate-sensitive P-gp ATPase activity was measured in membranes of Sf9 insect cells infected with a recombinant baculovirus containing the human MDR1 cDNA used with appropriate control membranes. MDCK and MDR1-transfected MDCK cell lines were then used to measure bidirectional P-gp transport of various steroids in the presence and absence of the P-gp inhibitor, GG918. Samples obtained were quantified using LC/MS. RESULTS: Our findings show that P-gp protein levels are inducible by estrone (4-fold over control), estriol (2-fold), and ethynyl estradiol (3-fold). MDR1 mRNA expression levels were also inducible in a concentration-dependent manner from 25 nM to 10 microM. Bidirectional transport studies indicate that ethynyl estradiol, estrone, and estriol are all substrates for P-gp with respective efflux ratios of 10.3, 6.9, and 2.8. Norethindrone was not found to be a substrate for P-gp. Ethynyl estradiol and progesterone were able to significantly stimulate P-gp ATPase activity in a concentration-dependent manner. CONCLUSIONS: Our studies indicate that several sex-steroid hormones are substrates for P-gp-mediated transport and are also able to induce P-gp expression at both the protein and mRNA level in vitro. Stimulation of P-gp ATPase catalytic activity by steroid hormones was also observed, suggesting physical interactions and identifying a need for further investigations to understand the in vivo effects of endogenous and synthetic steroid hormones on the expression and function of P-gp. PMID- 15290874 TI - A sure start--but what about the good end? Lack of cohesion in older people's services needs swift action. PMID- 15290873 TI - Transport of amino acid esters and the amino-acid-based prodrug valganciclovir by the amino acid transporter ATB(0,+). AB - PURPOSE: The purpose of this study was to analyze the transport of amino acid esters and the amino-acid-based prodrug valganciclovir by the Na(+)/Cl(-)-coupled amino acid transporter ATB(0,+). METHODS: The interaction of amino acid esters and valganciclovir with the cloned rat ATB(0,+) was evaluated in a mammalian cell expression system and in the Xenopus oocyte expression system. RESULTS: In mammalian cells, expression of ATB(0,+) induced glycine uptake. This uptake was inhibited by valine and its methyl, butyl, and benzyl esters. The benzyl esters of other neutral amino acids were also effective inhibitors. Valganciclovir, the valyl ester of ganciclovir, was also found to inhibit ATB(0,+)-mediated glycine uptake competitively. Exposure of ATB(0,+)-expressing oocytes to glycine induced inward currents. Exposure to different valyl esters (methyl, butyl, and benzyl), benzyl esters of various neutral amino acids, and valganciclovir also induced inward currents in these oocytes. The current induced by valganciclovir was saturable with a K0.5 value of 3.1+/-0.7 mM and was obligatorily dependent on Na+ and Cl-. The Na+:Cl-:valganciclovir stoichiometry was 2 or 3:1:1. CONCLUSIONS: Amino acid esters and the amino-acid-based prodrug valganciclovir are transported by ATB(0,+). This shows that ATB(0,+) can serve as an effective delivery system for amino acid-based prodrugs. PMID- 15290872 TI - P-glycoprotein expression, localization, and function in sandwich-cultured primary rat and human hepatocytes: relevance to the hepatobiliary disposition of a model opioid peptide. AB - PURPOSE: The isolation of hepatocytes from intact liver involves collagenase digestion of the tissue, resulting in loss of cell polarization and functional vectorial excretion. These studies examined repolarization, localization of P glycoprotein (P-gp) to the canalicular domain of the hepatocyte, and re establishment of vectorial transport in sandwich-cultured (SC) rat and human primary hepatocytes. METHODS: Protein localization and expression were determined in SC hepatocytes by confocal microscopy and Western blotting, respectively. Transporter function was evaluated by measuring [D-penicillamine2,5]enkephalin (3H-DPDPE) and 5 (and 6)-carboxy-2',7'-dichlorofluorescein (CDF) biliary excretion in SC hepatocytes. RESULTS: P-gp and the canalicular marker protein dipeptidyl peptidase IV (DPPIV) co-localized by Day 3 and Day 6 in SC rat hepatocytes and SC human hepatocytes, respectively, consistent with canalicular network formation visualized by light microscopy. Co-localization of multidrug resistance associated protein 2 (MRP2) and P-gp in SC human hepatocytes was observed on Day 6 in culture. Expression levels of P-gp increased slightly in both species over days in culture; similar expression was observed for MRP2 in SC human hepatocytes. Oatp1a1 expression in SC rat hepatocytes was maintained over days in culture, whereas Oatp1a4 expression decreased. OATP1B1 expression decreased slightly on Day 3 in SC human hepatocytes. OATP1B3 expression was constant in SC human hepatocytes. In vitro biliary excretion of the opioid peptide 3H-DPDPE correlated with the proper localization of canalicular proteins in both species. Excretion of CDF in SC human hepatocytes confirmed network formation and MRP2 function. CONCLUSIONS: These studies indicate that SC hepatocytes repolarize and traffic functional canalicular transport proteins to the appropriate cellular domain. PMID- 15290875 TI - Catch them young. Public health community must begin in the classroom. PMID- 15290876 TI - Sea change. PMID- 15290877 TI - Hands off, that's our policy. PMID- 15290878 TI - Hip replacements. Community beds can have a significant impact on reducing length of stay in acute wards. PMID- 15290879 TI - Your money or their life. Drug to combat blood pressure may come at a price, but the wider health benefits should not be underestimated, says Nick Summerton. AB - New anti-hypertensive drugs, although more expensive than conventional treatments, have significant benefits in the management of cardiovascular disease. NICE analysis of cost-benefits don't consider their impact on other risk factors for cardiovascular disease, such as diabetes. A broader perspective is necessary when assessing the public health of anti-hypertensive drugs. PMID- 15290880 TI - Help from on high. Hospital trusts must now take steps to address their patients' religious needs. Some are already doing so, as Jeremy Davies reports. PMID- 15290881 TI - Out for the count. Simon Lawton-Smith askes whether community-based services will reduce acute admissions. PMID- 15290882 TI - Experts don't make exports. Is London overly reliant on overseas health. PMID- 15290883 TI - Acute effects of sildenafil on Humphrey visual field and intraocular pressure. AB - PURPOSE: To evaluate the effects of sildenafil on visual field and intraocular pressure in a group of healthy subjects. METHODS: Twenty-eight healthy male volunteers with normal eyes were included in the study. Visual field examinations were performed using FASTPAC 30-2 program (white-on-white and blue-on-yellow) with the Humphrey field analyzer before and one hour after receiving oral 50 and 100 mg sildenafil citrate. RESULTS: The mean age was 51.1 +/- 8.9 years. Mean deviation, pattern standard deviation, short-term fluctuation and corrected pattern standard deviation did not differ significantly among tests both in white on-white and blue-on-yellow visual field examinations. Changes in intraocular pressure were not statistically significant. CONCLUSIONS: No significant effect of sildenafil was seen on visual field and intraocular pressure in healthy subjects. PMID- 15290884 TI - Severe conjunctival and eyelid involvement in pemphigus vulgaris. AB - Pemphigus vulgaris (PV) is only rarely associated with severe ocular involvement. We present a case of PV with bilateral severe swelling, induration and thickening of eyelids, and multiple conjunctival and lid margin erosions, complicated by high intraocular pressure secondary to steroid treatment. After successful systemic immunosuppressive, steroid, and antiglaucomatous treatment, clinical features were resolved, and intraocular pressure returned to normal. Severe ocular diseases without dermatological involvement may be present in PV patients. PMID- 15290885 TI - The influence of the calcium channel blocker verapamil on experimental glaucoma. AB - PURPOSE: To study the influence of the calcium channel blocker verapamil on the development of glaucoma in the adrenalin-induced experimental model of glaucoma. METHODS: In the experimental model, glaucoma was induced in albino rabbits by repeated injections of small doses of adrenalin. The criteria for experimental glaucoma were (1) increased IOP, (2) decreased outflow and (3) decreased blood supply to the eye. One group with advanced glaucoma served as a control. In the other 3 groups, instillations of 0.25% verapamil were applied at different stages of the glaucomatous process. RESULTS: Verapamil blocked the development of experimental glaucoma when applied concurrently with the adrenalin injections, and instillations at the initial or advanced stages of the disease stabilized the progression of glaucoma. CONCLUSION: We demonstrate that mechanisms involving adrenalin and calcium are involved in glaucoma and that the use of calcium channel blockers is most effective in the early stages of the glaucomatous process. PMID- 15290886 TI - Cost efficiency and cost effectiveness of cataract surgery at the Malaysian Ministry of Health ophthalmic services. AB - PURPOSE: To determine the cost efficiency and to compare the cost effectiveness of conventional extracapsular cataract surgery (ECCE) and phacoemulsification at three hospitals of the Malaysian Ministry of Health (MOH). METHODS: Patient demography, pre-operative visual acuity, intra-operative complications, post operative complications and post-operative visual acuity were recorded for two hundred and forty seven of the 400 patients who underwent cataract surgery during a 2-week period. The cost of surgery, which included capital, staff and overhead, and patient care consumable costs were assessed prospectively in 8 randomly sampled patients over a 3-month period. Cost efficiency refers to cost per cataract surgery. Cost effectiveness refers to cost per successful cataract surgery. This is estimated by the ratio of cost efficiency to the proportion of successful cataract surgery. Successful surgery was defined as best-corrected visual acuity (BCVA) of better than 6/12 at 3 months post-operatively. RESULTS: Proportion of patients who had post-operative visual acuity of 6/12 or better was higher in phacoemulsification group (94%) than in the ECCE group (81%). Conventional extracapsular cataract surgery with intraocular lens implant costs RM3442 (USD 905.79) and phacoemulsification with intraocular lens implant costs RM4288 (USD 1128.42). DISCUSSION: There was no significant difference in cost effectiveness between ECCE and phacoemulsification. The cost of cataract surgery in the MOH hospital was found to be high due to the high overhead costs. PMID- 15290887 TI - Foveolar choroidal hemodynamics in proliferative diabetic retinopathy. AB - PURPOSE: We studied the choroidal circulation in patients with proliferative diabetic retinopathy (PDR) to assess whether choroidal hemodynamic abnormalities may exist in PDR. METHODS: Eighteen eyes of 18 patients with PDR and high-risk characteristics for visual loss were included in this study. Mean duration of diabetes was 20 +/- 9 years (mean +/- SD), hemoglobin A1c was 8.9 +/- 2.3%, and blood glucose at the time of blood flow measurement was 188 +/- 90 mg/dl. Choroidal circulatory parameters obtained in these patients were compared to those of 35 eyes of 35 age and blood pressure matched, healthy controls using a Wilcoxon rank sum test. Laser Doppler flowmetry (Oculix) was used to calculate relative choroidal blood velocity (Chvel), volume (Chvol), and flow (Chflow) in the center of the foveola. RESULTS: No significant differences in average age, mean blood pressure (BPm), or perfusion pressure (PP) were observed between diabetic patients and control subjects. In diabetic patients, Chvol was 0.29 +/- 0.08 (mean +/- 1 SD) arbitrary units (AU); this value was 15% lower than that of control subjects, 0.34 +/- 0.10 AU (p = 0.04). In contrast, average Chvel was not significantly different between subjects with PDR (0.39 +/- 0.07 AU) and control subjects (0.41 +/- 0.07 AU). The average Chflow in subjects with PDR (9.4 +/- 2.9 AU) was 27% lower than that of control subjects (12.8 +/- 4.2 AU; p = 0.003). No significant correlation was detected between the circulatory measurements and age, BPm, or PP. There was a statistically significant negative correlation between duration of diabetes and Chvel (R = -0.55; p = 0.03). CONCLUSIONS: Our results suggest that Chvol and Chflow are significantly reduced in patients with PDR. PMID- 15290888 TI - The effects of darkness on retrobulbar hemodynamics in patients with early stages retinitis pigmentosa. AB - PURPOSE: To investigate the retrobulbar hemodynamic changes occurring in light and darkness in patients with early stages retinitis pigmentosa (RP). METHODS: Eleven early stages RP patients were enrolled in the study. The peak systolic velocity (PSV), end-diastolic velocity (EDV), pulsatility index (PI) and resistivity index (RI) of the ophthalmic artery (OA) and central retinal artery (CRA) were measured by color Doppler imaging in light and darkness. The results were compared with that of 10 age-matched normal subjects. RESULTS: PSV and EDV of the OA did not show significant differences in both groups. However, PSV and EDV of the CRA in RP patients decreased significantly in darkness (p = 0.001, p < 0.001, respectively). In addition, we found a significant increase in RI of the CRA in RP patients in darkness (p = 0.003). On the contrary, PSV and EDV of the CRA in control subjects showed a significant increase (p = 0.002, p < 0.001, respectively) and RI showed a significant decrease in darkness (p = 0.004). CONCLUSIONS: Darkness causes a decreased blood flow in the CRA in early stages of RP. PMID- 15290889 TI - Value of the congenital hypertrophy of the retinal pigment epithelium in the diagnosis of familial adenomatous polyposis. AB - BACKGROUND: Several kinds of congenital hypertrophy of the retinal pigment epithelium (CHRPE) have been described in patients with familial adenomatous polyposis (FAP). This study aims to assess which properties of CHRPE better predict FAP and investigate whether a relationship exists between specific CHRPE characteristics and FAP variants. METHODS: We examined 286 subjects, Group I- patients with FAP plus individuals "at risk"; n = 173; Group II--controls n = 113. Retinal lesions were classified in five types (A-E) and different characteristics (distribution, number, shape, size, pigmentation and site) were evaluated. RESULTS: The most common lesions in affected subjects were types A-D (83.4%) whilst in the "at risk" and control groups were type E. Greater numbers of lesions and bilateral distribution occurred more frequently among affected subjects than in other participants (p < 0.001). Large lesions with mixed pigmentation were associated with polyposis (p > 0.5). Controls had solitary CHRPE lesions (3.5%) and types C and E lesions (23%). The cumulative sensitivities and specificities of CHRPE were 42 and 97%, respectively. CHRPE was most common among those with classical FAP, but no specific characteristic was associated with any particular FAP variant. CONCLUSIONS: Pigmented fundal lesions are highly pleomorphic and represent the variable expression of a common genetic defect of growth regulation. No association was found between CHRPE characteristics and specific FAP variants. PMID- 15290890 TI - Regression of optic nerve head neovascularization in proliferative diabetic retinopathy after intravitreal triamcinolone. Regression of diabetic optic disc neovascularization after intravitreal triamcinolone. AB - The aim of this study is to report the clinical outcome of a diabetic patient with optic nerve head neovascularization treated with an intravitreal injection of triamcinolone acetonide. A 52-year-old patient presented with clinically significant diffuse macular edema and optic nerve head neovascularization due to proliferative diabetic retinopathy in her right eye. Despite grid laser photocoagulation in the macular region macular edema progressed and visual acuity declined. The patient received a single intravitreal injection of 4 mg triamcinolone acetonide with topical anesthesia. After injection of triamcinolone acetonide visual acuity increased and macular edema decreased. Furthermore optic nerve head neovascularization had markedly regressed. No complication was observed during follow-up period. Intravitreal injection of triamcinolone acetonide may be useful for treatment of optic nerve head neovascularization in patients with proliferative diabetic retinopathy. PMID- 15290891 TI - Transpupillary thermotherapy in the treatment of circumscribed choroidal hemangiomas. AB - PURPOSE: To evaluate the efficacy of transpupillary thermotherapy (TTT) for the treatment of serous retinal detachment secondary to circumscribed choroidal hemangiomas (CCH). MATERIAL AND METHODS: Four eyes of four consecutive patients who presented decreased vision due to serous macular detachment secondary to CCH were enrolled in this study. After informed consent was obtained, the four eyes were treated with TTT. All the patients underwent pretreatment ocular examination, which included fluorescein angiography and ultrasonography. TTT was applied using a diode laser at 810 nm with a spot size of 4.3 mm. The diode laser was transmitted through a contact lens. The end-point of the treatment was a detectable light-gray appearance of the entire lesion. The patients were re examined monthly during the first 6 months, and regularly thereafter. RESULTS: Within 3 months of treatment all eyes had already demonstrated decreased exudation on clinical examination and on fluorescein angiography. Reduction in tumor prominence was observed in all eyes by A-B ultrasonography. Three patients showed an improvement in visual acuity (VA) over a period of 6 months. Case 2 from 20/60 to 20/25; case 3 from 20/400 to 20/50 and case 4 from 20/80 to 20/20. The VA in case 1 remained unchanged (counting fingers). No recurrences were observed within a mean follow-up of 14.5 months. CONCLUSIONS: TTT showed no deleterious side effects in treating serous macular detachment secondary to CCH, and must be regarded as a therapeutic alternative to manage selected cases. PMID- 15290892 TI - Corneal topographic changes after phacoemulsification through steep axis incision. AB - PURPOSE: To evaluate the effect of 5.5 mm sutured and 4.0 mm sutureless corneal incisions after phacoemulsification on corneal topograpy. MATERIAL AND METHODS: Twenty eyes of 20 patients that underwent phacoemulsification with IOL implantation were randomly divided into two. The first group received a PMMA IOL through a sutured 5.5 mm corneal incision. The second group received a foldable hydrophilic acrylic IOL through a sutureless 4.0 mm incision. Complete ocular examination and computerized corneal topography (Keratron Corneal Analyzer, Software version 3.2, Optikon 2000, Italy) were performed preoperatively, at 1 week, and 1 month postoperatively. The Maloney indices [best fit sphere (BFS), best fit cylinder (BFC) and topographic irregularity (TI)] were provided by the Keratron Corneal Analyzer. RESULTS: There was no statistically significant difference in the mean BFS between the groups preoperatively (p = 0.305), at postoperative week 1 (p = 0.362), and at postoperative month 1 (p = 0.160). The BFC was significantly higher in the 5.5 mm incision group than in the 4.0 mm incision group preoperatively (p = 0.025), however, there was no statistically significant difference between the groups at postoperative week 1 (p = 0.909), and at postoperative month 1 (p = 0.382). There was no statistically significant difference in the mean TI between the groups preoperatively (p = 0.494) and at postoperative week 1 (p = 0.271). However the mean TI at postoperative month 1 was significantly higher in the 5.5 mm incision group than the 4.0 mm incision group (p = 0.05). CONCLUSION: The 5.5 mm sutured incisions caused an increase in TI at 1 week and 1 month postoperatively. For a more rapid visual rehabilitation, 4.0 mm sutureless incisions are recommended. PMID- 15290893 TI - Trisomy 21 and the brain. AB - In fetuses with Down syndrome, neurons fail to show normal dendritic development, yielding a "tree in winter" appearance. This developmental failure is thought to result in mental retardation. In adults with Down syndrome, neuronal loss is dramatic and neurofibrillary and neuritic Abeta plaque pathologies are consistent with Alzheimer disease. These pathological changes are thought to underlie the neuropsychological and physiological changes in older individuals with Down syndrome. Two chromosome 21-based gene products, beta-amyloid precursor protein (betaAPP) and S100B, have been implicated in these neuronal and interstitial changes. Although not necessary for mental retardation and other features, betaAPP gene triplication is necessary, although perhaps not sufficient, for development of Alzheimer pathology. S100B is overexpressed throughout life in Down patients, and mice with extra copies of the S100B gene have dendritic abnormalities. S100B overexpression correlates with Alzheimer pathology in post adolescent Down syndrome patients and has been implicated in Abeta plaque pathogenesis. Interleukin-1 (IL-1) is a non-chromosome-21-based cytokine that is also overexpressed throughout life in Down syndrome. IL-1 upregulates betaAPP and S100B expression and drives numerous neurodegenerative and self-amplifying cascades that support a neuroinflammatory component in the pathogenesis of sporadic and Down syndrome-related Alzheimer disease. PMID- 15290894 TI - P2X7 receptor expression after ischemia in the cerebral cortex of rats. AB - Large amounts of adenosine 5'-triphosphate (ATP) released from cellular sources under pathological conditions such as ischemia may activate purinoceptors of the P2X and P2Y types. In the present study, the expression of the P2X7 receptor subtype in the brain cortex of spontaneously hypertensive rats was investigated using a permanent focal cerebral ischemia model. Immunocytochemistry with antibodies raised against the intracellular C-terminus of the P2X7 receptor showed a time-dependent upregulation of labeled cells in the peri-infarct region after right middle cerebral artery occlusion (MCAO) in comparison to controls. Double immunofluorescence visualized with confooal laser scanning microscopy indicated the localization of the P2X7 receptor after ischemia on microglial cells (after 1 and 4 days), on tubulin betaIII-labeled neurons (after 4 and 7 days), and on glial fibrillary acidic protein (GFAP)-positive astrocytes (after 4 days). In the following experiments, changes occurring 4 days after MCAO were investigated in detail. Western blot analysis of the cortical tissue around the area of necrosis indicated an increase in the P2X7 receptor protein. Immunoelectron microscopy revealed the receptor localization on synapses (presynaptically), on dendrites, as well as on the nuclear membrane of neurons (postsynaptically) and glial cells. Terminal deoxynucleotidyl transferase mediated deoxyuridine triphosphate-biotin nick end labeling in combination with P2X7 receptor immunocytochemistry indicated a co-expression on the apoptotic cells. Active caspase 3 was especially observed on GFAP-positive astrocytes. In conclusion, the present data demonstrate a postischemic, time-dependent upregulation of the P2X7 receptor-subtype on neurons and glial cells and suggest a role for this receptor in the pathophysiology of cerebral ischemia in vivo. PMID- 15290896 TI - A detrimental role for nitric oxide synthase-2 in the pathology resulting from acute cerebral injury. AB - Nitric oxide (NO) synthesized from the inducible isoform of nitric oxide synthase (NOS-2) has been suggested to play both beneficial and deleterious roles in various neuropathologies. To define the role of nitric oxide in traumatic brain injury, we subjected male mice lacking a functional NOS-2 gene (NOS-2-/-) and their wild-type littermates (NOS-2+/+) to mild or severe aseptic cryogenic cerebral injury. Expression of NOS-2 mRNA and protein was observed in NOS-2+/+ animals following injury. Lesion volume (as measured by histology and brain imaging) and neurological outcome (using motor and cognitive behavioral paradigms) were assessed at various times after injury. While magnetic resonance imaging revealed the extent of edema of the 2 genotypes to be similar, histology showed a reduced (32%) lesion volume in severely injured NOS-2-/- compared with NOS-2+/+ mice. In addition, NOS-2-/- mice showed significant improvements in both contralateral sensorimotor deficits (grid test: p = 0.011) and cognitive function (Morris water maze: p = 0.009) after severe injury compared to their wild-type littermates. This indicates that lesion volume is reduced and neurological recovery is improved after acute traumatic injury in mice lacking a functional NOS-2 gene, and strongly suggests that the post-trauma production of NO from this source contributes to neuropathology. PMID- 15290895 TI - Immunohistochemical detection of EGFRvIII in high malignancy grade astrocytomas and evaluation of prognostic significance. AB - The purpose of this study was to establish an accurate and accessible immunohistochemical (IHC) method for detecting vIII Egf receptor and to assess the prognostic significance of the method as applied to the detection of vIII in malignant astrocytomas. The accuracy of the method was determined by comparing vIII immunoreactivity in formalin-fixed and paraffin-embedded tumor sections versus RT-PCR results from the analysis of RNA extracted from corresponding frozen specimens. RT-PCR revealed vIII transcript in 18 of 44 cases in this series, and IHC analysis of matched formalin-fixed and paraffin-embedded sections showed EGFRvIII reactivity in each of these 18 tumors, as well as 1 additional tumor that was negative for vIII transcript. EGFR amplification was evident in all tumors expressing vIII; none of the 15 tumors lacking amplified EGFR were positive for vIII transcript or vIII protein. IHC analysis for vill expression was next applied to a large series of anaplastic astrocytomas (AAs) and glioblastoma multiforme (GBMs) from clinical trial patients with complete follow up and that had been previously examined by FISH for amplified EGFR. Among the GBMs, vIII detection by IHC was determined in 19 of 46 cases (41.3%) with EGFR amplification, and in only 3 of 59 tumors lacking amplified EGFR (5.1%). Among the AAs, vIII expression was observed in 3 of 14 cases with amplified EGFR (21.4%) and in 6 of 49 cases without EGFR amplification (12.2%). GBM and AA patient survival analysis as a function of vIII expression showed contrasting results, with vIII positivity having no association with survival among GBM patients (p = 0.84), but being highly associated with reduced survival among AA patients (p = 0.0016). This latter finding, though quite possibly a result of vIII's association with increasing AA patient age, suggests that vIII IHC will be useful for identifying and/or confirming the identity of malignant astrocytomas whose clinical behavior is consistent with that of GBM. PMID- 15290897 TI - Comparative clinico-pathological study of saposin-A-deficient (SAP-A-/-) and Twitcher mice. AB - The Twitcher mouse (twi/twi) has been widely used as an animal model of globoid cell leukodystrophy (GLD; Krabbe disease), a hereditary leukodystrophy due to genetic galactosylceramidase deficiency. Recently, we generated a new mouse model of late-onset, chronic GLD (SAP-A-/- mice) by introducing a mutation (C106F) in the saposin A domain of the sphingolipid activator protein gene. Comparative study of SAP-A-/- and twi/twi mice revealed delay in the onset of neurological symptoms in SAP-A-/- mice (90 days vs 20 to 25 days), milder symptoms, and prolonged average survival (134.4 +/- 29.1 days vs 47.5 +/- 3.9 days). However, in both, the earliest sites of demyelination and macrophage infiltration were in regions of the 8th nerve and the spinal tract of the 5th nerve and spinal-cord, where macrophages could be detected as early as day 30 in asymptomatic SAP-A-/- mice. Furthermore, spacio-temporal pattern of demyelination/macrophage infiltration and the extent of neuropathology at the terminal stage are closely similar in both. These results suggest that peripheral macrophages are readily accessible in these sites and participate in the demyelinating process in the central nervous system. PMID- 15290898 TI - Vascular smooth muscle actin is reduced in Alzheimer disease brain: a quantitative analysis. AB - We analyzed smooth muscle actin (SMA) immunoreactivity in brain blood vessels of 10 ApoE 4,4 Alzheimer disease (AD) patients and 10 ApoE 3,3 AD patients matched for age, sex, and duration of dementia. We also examined 10 cognitively and neuropathologically normal controls matched for age and sex. Vascular SMA immunoreactivity in the arachnoid, grey matter, and white matter was quantified by image analysis. There was less SMA immunoreactivity in blood vessels of all AD patients when compared to cognitively and neuropathologically normal controls (p < 0.001). In addition, arachnoidal vessels of ApoE 4,4 AD patients had less SMA immunoreactivity than ApoE 3,3 AD patients (p < 0.05). There is decreased vascular SMA density in arachnoid, grey matter, and white matter blood vessels in patients with AD when compared to age matched, cognitively and neuropathologically normal controls. The severity of the loss of SMA within the AD group may depend on ApoE type. PMID- 15290899 TI - Lewy body-related alpha-synucleinopathy in aging. AB - To clarify the significance of Lewy body (LB)-related alpha-synucleinopathy in aging, we investigated the incidence of LBs in 1,241 consecutive autopsy cases (663 males and 578 females). LB pathology was identified histologically in sections stained with hematoxylin and eosin and with anti-ubiquitin and anti alpha-synuclein antibodies. Cases without LBs were classified as LB stage 0 (987 cases). Cases with LBs were classified as follows: LB stage I = incidental LBs (149 cases); LB stage II = LB-related degeneration without attributable clinical symptoms (47 cases); LB stage III = Parkinson disease without dementia (10 cases); LB stage IV = dementia with Lewy bodies (DLB) transitional (limbic) form (25 cases); and LB stage V = DLB neocortical form (23 cases). The average age at death was greater for those cases with LBs. There were no gender differences in the LB pathology. G842A polymorphism in the paraoxonase I gene was associated with men in LB stage II or above and suggests a gender-specific risk factor. LB stage V had higher stages of neurofibrillary tangle and senile plaque involvement and also had a higher frequency of apolipoprotein E epsilon4. Our findings indicate that LBs are associated with cognitive decline, either independently or synergistically with neurofibrillary tangles and senile plaques. PMID- 15290900 TI - Meningoencephalitis and demyelination are pathologic manifestations of primary polyomavirus infection in immunosuppressed rhesus monkeys. AB - The human polyomavirus JC (JCV) is the etiologic agent of progressive multifocal leukoencephalopathy (PML), a demyelinating disease of the CNS that occurs in immunosuppressed individuals. Because polyomavirus-induced CNS pathology usually occurs as a result of the reactivation of latent virus, little is known about the disease manifestations of a primary polyomavirus-induced disease in man. To model such a primary infection, SV40-negative rhesus monkeys were immunosuppressed by infection with the virus SHIV-89.6P and then superinfected with the polyomavirus SV40. The animals developed CNS pathology characterized by both demyelination and meningoencephalitis. This observation suggests that a primary polyomavirus infection can be associated with an inflammatory CNS process. These data shed new light on the pathogenic mechanisms of primate polyomaviruses in the immunocompromised host. PMID- 15290901 TI - The G336S variant in the human neurofilament-M gene does not affect its assembly or distribution: importance of the functional analysis of neurofilament variants. AB - The human neurofilament medium (hNFM) subunit is one of the 3 neurofilament (NF) polypeptides, which are the most abundant intermediate filament (IF) proteins in post-mitotic neurons. The formation of neurofilamentous aggregates is a pathological hallmark of many neurodegenerative diseases, including the Lewy bodies found in Parkinson disease (PD). A Gly336Ser (G336S) variant in the rod domain of hNFM has recently been described in a patient with early-onset autosomal-dominant PD. In this study, we have generated a mammalian expression vector encoding the variant hNFM cDNA and characterized its effects on the formation of heteropolymeric IFs in heterologous cell lines. We have also investigated the distribution of the (G336S) hNFM variant protein in neuronal CAD cells, as well as the effects of the variant on the distribution of other cellular organelles and proteins. Our results demonstrate that the G336S variant does not affect the formation of IF networks nor the distribution of the variant hNFM protein. Our data suggest that if the G336S variant is involved in the development of PD, it does not appear to be due to defects in the assembly and distribution of NFs. PMID- 15290903 TI - Port maintenance. PMID- 15290902 TI - Targeting prion amyloid deposits in vivo. AB - The diagnosis of prion diseases in humans is challenging due to a lack of specific and sensitive non-invasive tests. Many forms of human prion disease including variant Creutzfeldt-Jakob disease (vCJD), Gerstmann-Straussler Scheinker (GSS) syndrome, and 10% of sporadic CJD cases are associated with amyloid deposition. Several positron emission tomography (PET) ligands have recently been developed to directly image beta-amyloid associated with Alzheimer disease. One of them, methoxy-X04, is a fluorescent derivative of Congo red with high binding affinity toward amyloid fibrils and good blood-brain barrier permeability. Using methoxy-X04, we investigated whether amyloid-targeting ligands can be also employed for direct imaging of amyloid deposits associated with some prion diseases. Such a method could potentially become a novel diagnostic approach for these conditions. Studies were performed on MB mice infected with the 87V mouse-adapted scrapie strain. Labeling of PrP amyloid plaques in brains of presymptomatic and symptomatic mice was demonstrated using in vivo transcranial two-photon microscopy after systemic administration of methoxy-X04. During real-time imaging, PrP amyloid deposits could be clearly distinguished 15 min after intravenous administration of methoxy-X04. The ligand showed rapid clearance from brain areas that did not contain amyloid deposits. PrP amyloid deposits could also be detected by direct application of methoxy-X04 on cerebellar sections from GSS patients. These results suggest that methoxy-X04 or similar derivatives could be used as PET imaging agents to improve the diagnosis of human prion diseases associated with amyloid deposition. PMID- 15290904 TI - The NCDB Public Benchmark and Survival Report: useful online data. PMID- 15290905 TI - Strategic planning: formulating strategies. PMID- 15290906 TI - Ten ways to improve patient relations. PMID- 15290907 TI - Strategic planning: five common problems with the implementation phase. AB - In spite of the extensive work that often goes into preparing a good strategic plan, most organizations experience delays, resistance, and other problems that adversely affect implementation. Knowing the most common implementation problems can help cancer program administrators avoid them during the planning phase. PMID- 15290908 TI - Final... Isn't all that! AB - The Centers for Medicare & Medicaid Services annually update their complicated set of reimbursement rules and final payment policies for fiscal intermediaries and carriers. "Final" rules are often "interim," requiring that providers and payers stay abreast of changing coverage and reimbursement requirements. PMID- 15290909 TI - Inflammatory bowel disease is linked to 19p13 and associated with ICAM-1. AB - Genome-wide scans have implicated several susceptibility loci, but linkage of 19p13 (IBD6) to Crohn's disease (CD) has not been fully replicated. We report a replication study of IBD6 in a UK Caucasian population. Two hundred eighty-four affected sibling pairs from 234 families were used for the linkage study. Linkage between IBD6 linkage and CD was replicated (LOD score = 1.59). Two candidate genes (DDXL and ICAM-1) within the IBD6 locus were examined in a case/control study with a total of 228 CD and 243 ulcerative colitis (UC) patients and 407 healthy controls. No association to either UC or CD was found in three novel intronic single nucleotide polymorphisms (SNPs) in DDXL. For ICAM-1, a significant association was found between K469 homozygosity and CD overall (39.9% vs 29.4%; Pc = 0.0096) and between E469 and fistulating disease (21.8% vs 10.0%, Pc = 0.030). In the UC group, limited disease extent was associated with homozygosity of the G241 allele (82.7% vs 64.7%, Pc = 0.0040). These data support linkage for CD at 19p13 and suggest that the amino acid polymorphisms in ICAM-1 may be associated with IBD. PMID- 15290910 TI - Therapeutic effects of a new lymphocyte homing reagent FTY720 in interleukin-10 gene-deficient mice with colitis. AB - BACKGROUND: FTY720 is a novel reagent that possesses potent immunosuppressive activity. The immunosuppression induced by FTY720 is mediated by completely different mechanisms from those of conventional immunosuppressants, that is, by altering the tissue distribution of lymphocytes rather than inhibiting activation. In this study, we examined the efficacy of FTY720 in the treatment of chronic colitis in an interleukin-10 gene-deficient (IL-0-/-) mouse model. METHODS: FTY720 was administered orally for 4 weeks to IL-10-/- mice with clinical signs of colitis. The gross and histologic appearance of the colon and the numbers, phenotype, cytokine production, and apoptosis of lymphocytes were compared with those characteristics in a control group. RESULTS: Single-dose administration of FTY720 resulted in the sequestration of circulating lymphocytes within the secondary lymphoid tissues. Four-week administration resulted in a significant reduction of the CD4+ T lymphocytes subpopulation in the colonic lamina propria and IFN-gamma production of the colonic lymphocytes, accompanied by a significant decrease in the severity of colitis. CONCLUSIONS: Treatment of established colitis in IL-10-/- mice with FTY720 ameliorated the colitis, probably as a result of decreasing the number of lymphocytes in the colonic mucosa and an associated reduction in IFN-gamma production. PMID- 15290912 TI - Clinical characteristics of familial versus sporadic Crohn's disease using the Vienna Classification. AB - BACKGROUND: The etiology of Crohn's disease, an illness protean in its manifestations, may be better resolved through studies involving more homogenous subgroups of patients. Because a strong genetic influence exists, family history of inflammatory bowel disease may be a useful variable for patient classification if patients with familial and sporadic Crohn's disease are clinically different. Our study attempted to define any possible differences. METHODS: The medical records of 552 patients were reviewed, and patients were classified according to guidelines of the Vienna Classification. Patients were then divided based on family history of inflammatory bowel disease, and the familial and sporadic groups were compared. RESULTS: Overall, 422 (78.9%) patients were diagnosed before age 40 years (A1) and 114 (21.1%) at age 40 years or older (A2). There were 141 (26.3%) patients with disease involving the terminal ileum only (L1), 211 (39.4%) in the colon only (L2), 117 (21.9%) in the terminal ileum and colon (L3), and 66 (12.3%) in the upper gastrointestinal tract (L4). Disease behavior, as determined at the time of last visit or telephone contact, was nonstricturing, nonpenetrating (B1) in 149 (27.9%) patients, stricturing (B2) in 50 (9.3%) patients, and penetrating (B3) in 336 (62.8%) patients. Comparisons among the groups of 53 patients with first-degree relatives only, the 96 patients with either first-, second-, or third-degree relatives (familial CD group), and the 439 patients with sporadic disease demonstrated no differences in sex, age at diagnosis, or disease location. There was a difference in disease behavior between the familial and sporadic groups (p = 0.048) that failed to exist when nonstricturing, nonpenetrating cases were excluded. No such difference was observed between the first-degree relatives only group and the sporadic group (p > 0.10). CONCLUSIONS: Using the Vienna Classification, familial and sporadic Crohn's disease differed only in disease behavior. However, this difference failed to exist after patients with nonstricturing, nonpenetrating disease were excluded. Therefore, familial and sporadic groups appear to be quite similar clinically, and family history does not appear to be a variable useful for disease subclassification. PMID- 15290911 TI - Circulating monocytes and plasma inflammatory biomarkers in active Crohn's disease: elevated oxidized low-density lipoprotein and the anti-inflammatory effect of atorvastatin. AB - We investigated inflammatory biomarkers in plasma and in circulating monocytes obtained from patients with Crohn's disease and healthy individuals. Additionally, we assessed the effects of atorvastatin, 10 microM, ex vivo on monocytes cultured for 18 hours from the same subjects. Plasma and blood monocytes from eight patients with active Crohn's disease and eight healthy individuals were analyzed by enzyme-linked immunosorbent and electrophoretic mobility assays. Patients with active Crohn's disease had increased plasma levels of tumor necrosis factor (TNF)-alpha (7.7-fold;p < 0.05), monocyte chemoattractant protein (MCP)-1 (1.3-fold; p < 0.05), and oxidized low density lipoprotein (oxLDL) (1.2-fold; p < 0.05). Monocytes from patients with Crohn's disease showed enhanced secretion of MCP-1 (4.8-fold; p < 0.05) and a markedly suppressed secretion of macrophage migration inhibitory factor (MIF) (93%; p < 0.001). Transcriptional activation of nuclear factor-kappaB did not differ between the groups. Treating monocytes with atorvastatin resulted in the suppression of MCP-1 (42%; p < 0.05) and TNF-alpha (45%; p < 0.05) secretion. These results show increased levels of certain proinflammatory biomarkers, including oxLDL, in plasma and indicate that peripheral blood monocytes in active Crohn's disease are sensitized to chemotaxis. Treatment with atorvastatin may be a potential strategy to reduce oxLDL and inhibit monocyte migration to inflamed tissue, thus attenuating the inflammatory response. PMID- 15290913 TI - Autoimmune disorders and extraintestinal manifestations in first-degree familial and sporadic inflammatory bowel disease: a case-control study. AB - BACKGROUND: Genetic factors may play a role in determining the development of extraintestinal manifestations (EIMs) and autoimmune diseases (AD) in patients with inflammatory bowel disease (IBD). We sought to determine the association between EIMs and AD in patients with first-degree familial IBD and sporadic IBD. METHODS: All patients evaluated in the IBD Clinic at the Mayo Clinic between January and September 1999 were offered enrollment. One clinic patient who was matched on age, gender, and geographic area of residence to each case served as controls. Information regarding EIMs and AD was obtained from a questionnaire completed by all IBD patients and controls. The adjusted odds ratios (95% CIs) for EIM as a function of first-degree familial IBD compared with sporadic IBD and AD as a function of first-degree familial IBD compared with sporadic IBD were estimated with a matched one-to-one conditional logistic regression model. RESULTS: Two hundred forty-three patients with IBD (47 first-degree familial IBD, 196 sporadic IBD) were enrolled. Forty percent of IBD patients had one or more EIMs compared with 14% matched controls [p < 0.001; OR = 3.1 (95% CI: 1.8 to 5.2)]. A total of 259 of the 1122 IBD patients and their first-degree family members indicated one or more EIM diagnoses (23%). The association between "familial versus sporadic" status and any EIM diagnosis was not significant [p = 0.59, the odds for an individual from a familial IBD family relative to an individual from a sporadic IBD family was 1.2 (95% CI: 0.8 to 1.7)]. Ten percent of IBD patients had one or more AD diagnoses compared with 19% matched controls [p = 0.04; OR = 0.4 (95% CI: 0.1 to 0.96)]. A total of 153 of the 1122 IBD patients and their first-degree family members indicated one or more AD diagnoses (14%). The association between disease status ("familial or sporadic") versus any AD diagnosis was not significant [p = 0.68, the odds for any AD in an individual from a familial IBD family relative to an individual from a sporadic IBD family was 1.4 (95% CI: 0.9 to 2.3)]. CONCLUSIONS: There was a positive association between IBD status (patient vs control) versus EIM, but not AD. A significant positive association between disease type (familial or sporadic) versus either EIM or AD was not detected. PMID- 15290914 TI - The clinical significance of right-sided colonic inflammation in patients with left-sided chronic ulcerative colitis. AB - BACKGROUND: Rarely, patchy right colonic inflammation has been observed in patients with left sided chronic ulcerative colitis (CUC), but the clinical significance of this finding is unknown. Therefore, the aim of this study was to evaluate the clinical and pathologic features and natural history of CUC patients with left-sided colitis combined with patchy right colonic inflammation and to compare the clinical course to a control group of patients with isolated left sided CUC. METHODS: Twelve patients with clinically and pathologically confirmed left-sided CUC, but also with patchy right colonic inflammation, were identified from a cohort of 352 consecutive patients with CUC who underwent colonoscopy at the Brigham and Women's Hospital between 1996 and 2000. In this cohort, 127 patients had left-sided colitis. As the first study to use controls in this setting, 35 consecutive patients with left-sided CUC, but without patchy right colonic inflammation, were selected and evaluated during the same time period. In all patients, the medical records were reviewed for a wide variety of clinical, endoscopic, and pathologic features. The mean follow-up time for the study and control groups was 105 +/- 128 and 112 +/- 80 months, respectively. RESULTS: Patients in the study group were significantly older than the control group at the time of diagnosis (47 +/- 17 years vs 35 +/- 14 years, p = 0.048), but the two groups had a similar gender distribution (25% male vs 40% male), prevalence of extraintestinal manifestations (25% vs 11%), frequency of nonsteroidal anti inflammatory drug use (75% vs 50%), family history of colitis (27% vs 15%), current tobacco use (8% vs 3%), history of appendectomy (8% vs 0%), and overall severity of disease (33% vs 46%). None of the patients in the study group, and only one control patient, had disease progression to pancolitis. One study patient developed high-grade dysplasia in the rectum that required a colectomy. None of the study or control patients developed clinical or pathologic features of Crohn's disease. CONCLUSIONS: Rarely patients with left-sided CUC may have patchy right colonic inflammation. The clinical features and natural history of patients with left-sided CUC and patchy right colonic inflammation is similar to patients with isolated left-sided CUC. PMID- 15290915 TI - Bones and Crohn's: risk factors associated with low bone mineral density in patients with Crohn's disease. AB - Previous studies have confirmed that the prevalence of decreased bone mineral density is elevated in patients with inflammatory bowel disease. The objective of the current study was to determine the prevalence of osteopenia and osteoporosis in a cross-sectional outpatient population of 242 adult patients with Crohn's disease and to determine which clinical characteristics and serum and urine biochemical factors might be predictive of bone loss. Thirty-seven percent had normal bone density, 50.0% were osteopenic, and 12.9% were osteoporotic. Among the sites used to diagnose low bone mineral density, the femoral neck demonstrated the highest prevalence of osteopenia and the ultra-distal radius the highest prevalence of osteoporosis. However, low bone mineral density at one site was always predictive of low bone mineral density at the other. Corticosteroid use during the year before assessment was found to be consistently predictive of low bone mineral density in males but not in females. In contrast, low body mass index and high platelet counts were consistently predictive of low bone mineral density in females but not in males. Disease location, smoking, and age were not predictive of changes in bone mineral density. PMID- 15290916 TI - Serum procalcitonin differentiates inflammatory bowel disease and self-limited colitis. AB - BACKGROUND: The distinction between idiopathic inflammatory bowel disease (IBD) and infectious, usually self-limited enterocolitis is still a diagnostic dilemma. Procalcitonin (PCT) is the prohormone of calcitonin and is considered a specific marker of bacterial infection. The aim of this prospective study was to determine the value of PCT in differentiating flares of IBD from self-limited colitis. In addition, because standard laboratory inflammatory parameters are poorly correlated with disease activity in IBD, the relation between PCT levels and disease activity was investigated. METHODS: A total of 76 patients (26 Crohn's disease, CD; 25 ulcerative colitis, UC; and 25 patients with self-limited enterocolitis) were enrolled. Serum levels of PCT were measured by a sandwich immunoluminometric assay. C-reactive protein (CRP) levels, white blood cell counts, and stool cultures were obtained from all patients. Disease activity was assessed by the Crohn's disease activity index (CDAI) and the Truelove index for CD and UC, respectively. RESULTS: Patients with self-limited enterocolitis showed significantly higher PCT levels when compared with IBD patients (0.36 ng/mL, range 0.18-1.7 vs 0.10 ng/mL, range 0.08 0.5, p < 0.001). For a PCT value of > or =0.4, the sensitivity for self-limited colitis was 92% and specifity 96%. The positive predictive value (PPV) for self-limited colitis was 96%, whereas the negative predictive value (NPV) was 93%. In IBD patients, PCT levels were in the normal range although significantly higher in active disease when compared with inactive disease (0.13 ng/mL, range 0.08-0.5 vs 0.09 ng/mL, range 0.08-0.15, p < 0.001). This difference was less pronounced for CD (0.11 ng/mL, range 0.08-0.2 vs 0.09 ng/mL, range 0.08-0.15, p < 0.05) than for UC (0.14 ng/mL, range 0.08-0.5 vs 0.09 ng/mL, range 0.08-0.11, p < 0.01). In CD, PCT levels correlated significantly 0.5, p < 0.01). with the CDAI (r =0.05, p <0.01). CONCLUSIONS: The measurement of PCT offers two diagnostic options in IBD. Supranormal levels indicate self-limited enterocolitis. Furthermore, although within the normal range in IBD, PCT levels may serve as a new serological marker of disease activity. PMID- 15290917 TI - Effects of disease activity on anti-Saccharomyces cerevisiae antibodies: implications for diagnosis and follow-up of children with Crohn's disease. AB - BACKGROUND: To determine diagnostic accuracy of anti-Saccharomyces cerevisiae antibodies (ASCA) in identifying children with inflammatory bowel disease (IBD) and to differentiate Crohn's disease (CD) from other IBD forms; and to determine the effect of medical or surgical treatment and of disease location and activity on ASCA titers. METHODS: Serum samples were obtained from 196 IBD children and 142 controls. ASCA IgA and IgG titers were measured by ELISA. Measurements were repeated during the follow up of CD children. RESULTS: ASCA titers were significantly higher in CD than in other IBD and in control patients. Combination of IgA and IgG ASCA positivity was highly specific for CD. Medical treatment and disease location did not influence assay results. Significantly lower ASCA titers were obtained in CD children with intestinal resection compared to CD-affected children who did not undergo surgical resection. ASCA titers correlated significantly with disease activity, and children with severe active disease showed higher ASCA values compared to those in remission. A significant reduction of ASCA was observed during the follow-up of CD children when clinical remission was achieved. CONCLUSIONS: The diagnostic accuracy of ASCA is influenced by disease activity and this suggests an additional use for the follow-up of CD children of this assay. PMID- 15290918 TI - Comparison of serological markers of inflammatory bowel disease with clinical diagnosis in children. AB - BACKGROUND: Our purpose was to study the concordance of serological tests for inflammatory bowel disease with clinical diagnosis established by traditional testing in children. METHODS: All children seen in our division who had IBD Diagnostic System (ie, pANCA, ASCA IgA, and ASCA IgG) performed over a 21-month period (June 1998 to February 2000) were identified. Their medical records were reviewed for basic demographics, test results (endoscopy, histology, and radiology), IBD Diagnostic System results, and patient symptoms/medications. Results of the IBD Diagnostic System were compared with several patient characteristics including age, sex, absence/presence of symptoms, medication use, disease activity and duration. RESULTS: One hundred seven patients were divided into 6 groups based on clinical diagnosis and IBD Diagnostic System results. The sensitivity, specificity and +/- predictive values of the IBD Diagnostic System for ulcerative colitis were 69.2, 95.1, 90.0, and 87.1%, respectively, and for Crohn's disease were 54.1, 96.8, 90.9, and 80.8%, respectively. Overall, the results of the IBD Diagnostic System were concordant with the clinical diagnosis in 76 of the 107 (71%) patients. CONCLUSIONS: In our experience, the specificity of IBD Diagnostic System is better than the sensitivity; the sensitivity is better for ulcerative colitis than Crohn's disease (69.2% vs 54.1%). The low sensitivity, especially for Crohn's disease, precludes the possibility that the IBD Diagnostic System can replace traditional studies when evaluating for inflammatory bowel disease. Though we do not exclude inflammatory bowel disease solely by IBD Diagnostic System results, it is reassuring to note that all patients without clinical evidence of inflammatory bowel disease also had negative IBD Diagnostic System results. PMID- 15290919 TI - The pathogenicity of cytomegalovirus in inflammatory bowel disease: a systematic review and evidence-based recommendations for future research. AB - During recent years, a clear association between complicated courses of ulcerative colitis and the presence of cytomegalovirus (CMV) has been established. The exact pathogenic role of CMV in these patients remains unclear despite a great number of published reports. Therefore, we undertook a systematic review to appraise critically all available evidence in the literature on the role of CMV during inflammatory bowel disease. We identified and analyzed more than 30 case reports and 9 case series. Based on these results, we propose a model for viral replication during inflammation and provide recommendations for future research. PMID- 15290920 TI - Granulocytapheresis is useful as an alternative therapy in patients with steroid refractory or -dependent ulcerative colitis. AB - BACKGROUND: Recently, granulocyte and monocyte adsorption apheresis (GCAP) has been shown to be safe and effective for active ulcerative colitis (UC). We analyzed the safety and efficacy of GCAP (G-1 Adacolumn) in patients with steroid refractory and -dependent UC. G-1 Adacolumn is filled with cellulose acetate carriers that selectively adsorb granulocytes and monocytes/macrophages. METHODS: Forty-four patients with UC were treated with GCAP. These patients received 5 apheresis sessions over 4 weeks. Twenty patients had steroid-refractory UC (group 1) and 10 had steroid-dependent UC (group 2). Fourteen patients who did not want readministration of steroids were treated with GCAP at the time of relapse, just after discontinuation of steroid therapy (group 3). RESULTS: Of 44 patients treated with GCAP, 24 (55%) obtained remission (CAI < or = 4), 9 (20%) showed a clinical response, and 11 (25%) remained unchanged. Only 2 of 10 patients (20%) with severe steroid-refractory UC (CAI > or = 12) achieved remission, whereas 7 of 10 patients (70%) with moderate steroid-refractory UC achieved remission (p < 0.05). The dose of corticosteroids was tapered in 9 of 10 (90%) patients with steroid-dependent UC after GCAP therapy. Twelve (86%) of 14 patients in group 3 showed an improvement in symptoms and could avoid re-administration of steroids after GCAP. No severe adverse effects occurred. CONCLUSIONS: The findings of this study suggest that GCAP may be a useful alternative therapy for patients with moderate steroid-refractory or -dependent UC, although cyclosporin A or colectomy is necessary in patients with severe UC. GCAP may also be useful for avoiding re administration of steroids at the time of relapse. Randomized, controlled clinical trials are needed to confirm these findings. PMID- 15290921 TI - Tolerance of 4-aminosalicylic acid enemas in patients with inflammatory bowel disease and 5-aminosalicylic-induced acute pancreatitis. AB - Derivatives of 5-aminosalicylic acid (5-ASA) used for the treatment of inflammatory bowel disease may induce acute pancreatitis of immunoallergic origin. 4-aminosalicylic acid (4-ASA) differs from its 5-ASA counterpart by the position of the NH2 group and has shown efficacy in ulcerative colitis. The risk of cross intolerance reaction between 5-ASA and 4-ASA has currently never been evaluated. We report three cases of 5-ASA-induced pancreatitis, with no recurrence of pancreatitis during subsequent treatment with 4-ASA enemas. We conclude that 4-ASA enemas are a safe and well-tolerated therapeutic alternative whenever 5-ASA-induced pancreatitis occurs. PMID- 15290922 TI - The inflammatory bowel disease questionnaire: a review of its national validation studies. AB - Health-related quality of life (HRQoL) is an important measure of illness perception on the part of the patient. The Inflammatory Bowel Disease Questionnaire (IBDQ) is a widely used questionnaire for HRQoL assessment in patients with inflammatory bowel diseases (IBDs). This questionnaire has been adapted and validated into several languages and cultural milieus. The aim of this study is to review the methods used by several adaptation studies for assessing the validity and reliability of the adapted IBDQ. A search was made of the Medline database for relevant articles since 1989. Standard validation criteria were used for including studies for further evaluation. The following aspects of the validation procedure were examined: translation, construct validity, reliability, sensitivity to change, and used statistical methods. Nine validation studies of the IBDQ, in England and in non English-speaking countries (Holland, Spain, Korea, Sweden, Greece, and China) were selected. All studies concluded that the adapted instrument was valid and reliable. Only few modifications were proposed. Two studies recommended the split of the four dimensions of the original questionnaire in five. Assessing HRQoL in patients with IBD is an ever-increasing practice, especially in clinical trials. IBDQ was proven to be valid and reliable in several cultural and linguistic milieus when appropriate validation procedures were applied. PMID- 15290923 TI - Scalloping of duodenal mucosa in Crohn's disease. AB - Scalloping of the duodenal mucosal folds is an endoscopic finding of small bowel mucosal pathology that is generally due to villous atrophy. Though it can be seen in many disease processes, it is most commonly associated with celiac disease. We report three patients with scalloping of duodenal folds and histologic confirmation of villous atrophy due to Crohn's disease. All patients had negative celiac serologies and two had positive markers for Crohn's disease (anti Saccharomyces cerevisiae antibodies). Patients had either ileitis or ileocolitis in addition to duodenal abnormalities. These cases illustrate that scalloping can occur in the duodenum in Crohn's disease. PMID- 15290924 TI - Diffuse alveolar hemorrhage after infliximab treatment of Crohn's disease. AB - Infliximab (Remicade), a chimeric monoclonal antibody to tumor necrosis factor alpha (anti-TNF-alpha), is being used with increasing frequency in the treatment of Crohn's disease. Infliximab's safety profile to date has been good with reported adverse events being mild to moderate. We report a case of diffuse alveolar hemorrhage after the second infliximab infusion in a patient with Crohn's disease. The mechanism by which infliximab may have caused the observed pulmonary insult remains unknown. Physicians should be aware of the possible association between infliximab treatment and the development of alveolar hemorrhage. Future cases should be reported. PMID- 15290925 TI - Video capsule endoscopy in inflammatory bowel disease: past, present, and future. AB - In this paper, we critically review the rationale, technical issues, and diagnostic findings, difficulties in interpretation, complications, potential clinical uses, and practical obstacles for capsule endoscopy in patients with inflammatory bowel disease (IBD). We will review the currently limited data on its use in IBD and discuss future areas of investigation required to evaluate critically its potential utility in these patients. PMID- 15290926 TI - Probiotics and the management of inflammatory bowel disease. AB - The demonstration that immune and epithelial cells can discriminate between different microbial species has extended our understanding of the actions of probiotics beyond simple barrier and antimicrobial concepts. Several probiotic mechanisms of action, relative to inflammatory bowel disease, have been elucidated: (1) competitive exclusion, whereby probiotics compete with microbial pathogens for a limited number of receptors present on the surface epithelium; (2) immunomodulation and/or stimulation of an immune response of gut-associated lymphoid and epithelial cells; (3) antimicrobial activity and suppression of pathogen growth; (4) enhancement of barrier function; and (5) induction of T cell apoptosis in the mucosal immune compartment. The unraveling of these mechanisms of action has led to new support for the use of probiotics in the management of clinical inflammatory bowel disease. Though level 1 evidence now supports the therapeutic use of probiotics in the treatment of postoperative pouchitis, only levels 2 and 3 evidence is currently available in support of the use of probiotics in the treatment of ulcerative colitis and Crohn's disease. Nevertheless, one significant and consistent finding has emerged during the course of research in the past year: not all probiotic bacteria have similar therapeutic effects. Rigorously designed, controlled clinical trials are vital to investigate the unresolved issues related to efficacy, dose, duration of use, single or multi-strain formulation, and the concomitant use of probiotics, synbiotics, or antibiotics. PMID- 15290927 TI - Inflammatory bowel disease gene hunting by linkage analysis: rationale, methodology, and present status of the field. AB - Observed inflammatory bowel disease (IBD) familial clustering and increased monozygotic twin concordance has led to the hypothesis that genetic loci containing IBD susceptibility genes can be identified by whole genome linkage mapping approaches. Methodology including collecting carefully phenotyped multiplex pedigrees, genotyping using highly informative microsatellite markers and linkage analysis by non-parametric allele sharing methods has been established. Eleven published genome wide screens (GWS) have studied more than 1,200 multiplex IBD pedigrees. Two-thirds of affected relative pairs were Crohn's disease (CD), 20% ulcerative colitis (UC) and the remaining were mixed. Seven loci (IBDI-7) on chromosomes 16q, 12, 6p, 14q, 5q, 19, and 1p have been identified with genome wide significant and independently replicated linkage. Risk alleles/haplotypes have been defined for the IBD1 (CARD15/NOD2), IBD3 (HLA) and IBD5 (5q cytokine cluster) loci. There has been evidence for a second chromosome 16 locus (IBD8) independent of NOD2 that overlaps IBD1 on the pericentromeric p-arm. Several other regions show great promise for containing additional IBD loci, particularly chromosome 3p with genome wide evidence in one study at 3p26 and more centromeric evidence in several other studies, and chromosomes 2q, 3q, 4q, 7, 11p, and Xp each with suggestive evidence of linkage in one and additional evidence in two or more studies. Single GWSs and fine mapping studies containing very large sets of pedigrees and in particular, more UC pedigrees, and the use of creative analytic and disease stratification schemes are required to identify, establish and refine weaker IBD loci. PMID- 15290928 TI - New approaches to gene hunting in IBD. AB - Without a doubt, one of the greatest promises of the human genome project has been the expectation that its completion will rapidly accelerate the discovery of the heritable components of complex disease. Such discoveries will educate us as to the underlying causes of disease and hold out great promise for medicine in the future by potentially enabling the prediction of individual risk, the selection of therapy based on genetic profile, and a more rational approach to designing drugs. Unfortunately, discovering these genetic components of complex disease has been much more difficult than initially hoped and, to date, very few positive results have been conclusively confirmed. We review here approaches to tackling the problem of gene finding in complex disease with an emphasis on new approaches that may complement and supplement existing methods that have been making only slow progress toward the elucidation of the genetic architecture of disease. Along the way we highlight the successful work by many groups in studying the genetics of IBD, where significant successes have arrived earlier than in most other complex human diseases. PMID- 15290929 TI - Antibiotics should be used as first-line therapy for Crohn's disease. AB - The etiology of Crohn's disease remains uncertain, and to date no therapy is curative. Recent experimental evidence suggests that an altered immune response to commensal enteric flora in a genetically susceptible host plays a key role in both the development and perpetuation of the intestinal inflammation of Crohn's disease. Thus, incorporation of antibiotics into the therapeutic armamentarium for Crohn's disease, either as first-line therapy or combined with immunomodulatory drugs, would seem to be a rational strategy. Indeed, most IBD clinicians would attest to the marked benefit of antibiotic therapy in individual patients. Skepticism surrounding this approach arises because evidenced-based analyses' show that the few clinical trials evaluating the efficacy of antibiotics for Crohn's disease have produced equivocal or negative results or have methodological deficiencies, including small number of patients and absence of a placebo group. However, by undertaking an analysis that integrates information from both basic and clinical spheres of study, the dichotomy between experimental and clinical observations tends to merge. This approach underscores certain key factors that determine an optimal response to antibiotics, emphasizes the requirement for assessment in well-defined subsets of patients, and leads to the conclusion that antibiotics do provide benefit for Crohn's disease. PMID- 15290930 TI - The role of antibiotics in the management of Crohn's disease. PMID- 15290931 TI - Antibiotics as a first-line therapy for Crohn's disease: is there any consensus? PMID- 15290932 TI - IBD in children: the more you look, the more you see.... PMID- 15290933 TI - Ileoanal pouch failure: can it be predicted? PMID- 15290934 TI - Seeing red in inflammatory bowel disease: the eosinophil at center stage? PMID- 15290935 TI - Limb-ited abnormalities in the offspring of ulcerative colitis patients. PMID- 15290936 TI - Multislice CT angiography of mesenteric vessels. AB - Due to several distinct advantages over conventional angiography (including minimal invasiveness, lower cost, and lower ionizing radiation exposure for patients and staff), computed tomography (CT) angiography has replaced diagnostic conventional angiography in several clinical situations. The recent introduction of multislice CT (MSCT) scanners has significantly improved CT angiographic applications, especially in the evaluation of the mesenteric vasculature. Thin slice collimation protocols associated with powerful postprocessing procedures allow the display of mesenteric circulation with excellent detail. The purposes of this presentation are (a) to illustrate the imaging technique that can be used to obtain state-of-the-art MSCT angiographic images of the mesenteric vasculature, (b) to review the normal anatomy and anatomic variants of mesenteric vessels, and (c) to illustrate some of the potential clinical applications of MSCT angiography of the mesenteric vessels. PMID- 15290937 TI - Isolated dissection of the superior mesenteric artery: CT findings in six cases. AB - Dissection of the superior mesenteric artery (SMA) not associated with aortic dissection is rare. The purpose of this study is to describe the computed tomographic (CT) findings of this condition. We studied the CT findings of six patients with isolated dissection of the SMA. CT demonstrated thrombosis of the false lumen or intramural hematoma (n = 4) and/or intimal flap (n = 4) in all six patients. Other CT findings were enlarged diameter of the SMA (n = 5), increased attenuation of the fat around the SMA (n = 5), and hematoma in the mesentery with hemorrhagic ascites (n = 1). CT is useful for the diagnosis of isolated dissection of the SMA, and increased attenuation of the fat around the artery is considered the key to the diagnosis when no definite findings are evident. PMID- 15290938 TI - Strangulated transmesosigmoid hernia: CT diagnosis. AB - We present a rare case of strangulated closed loop small bowel obstruction secondary to a transmesosigmoid hernia to emphasize the diagnostic role of computed tomography in patients with no history of previous surgery. The characteristic computed tomographic features showed a cluster of dilated, fluid filled, U- and C-shaped loops of small bowel entrapped the left posterior and lateral to the sigmoid colon through a defect in the mesosigmoid, which caused anterior and medial displacement of the sigmoid colon. PMID- 15290939 TI - Internal herniation through a defect of the broad ligament of the uterus. AB - Internal herniation through a defect of the uterine broad ligament is rarely encountered. We report a case of a 52-year-old woman in whom a closed bowel loop with approximation of its ends to the uterus was the definitive computed tomographic finding. This type of internal herniation should be considered in the differential diagnosis of female patients presenting with ileus. PMID- 15290940 TI - Internal hernia with volvulus and intussusception: case report. AB - An 82-year-old male presented to the hospital because of acute exacerbation of abdominal pain and biliary vomiting. Contrast-enhanced computed tomography of the abdomen was performed. A left paraduodenal hernia associated with volvulus, intussusception, and bowel wall ischemia were radiologically diagnosed. Surgery confirmed the diagnostic imaging findings. We present the first case of an association of these acute abdominal conditions. PMID- 15290941 TI - Dynamic MRI in indirect estimation of microvessel density, histologic grade, and prognosis in colorectal adenocarcinomas. AB - The relations of dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) parameters to microvessel density (MVD), histologic grade, and presence of metastasis were evaluated to establish new prognostic indicators in colorectal cancer (CRC). Fast-low angle shot DCE-MRI parameters (time-intensity curves, TICs; maximal relative enhancement within the first minute, E(max/1); maximal relative enhancement of the entire study, Emax; steepest slope of the contrast enhancement curve; and time to peak enhancement) of 21 CRCs (seven Duke stage B, 12 Duke stage C, and two Duke stage D) were retrospectively evaluated and correlated with corresponding postoperative MVD measurements, histologic grades, and presence of metastasis at 2 years. TICs were classified as type A in nine (43%), type B in seven (33%), and type C in five cases (24%). There was a significant difference between TIC types with regard to MVD (p < 0.05-0.001). Time to peak enhancement, steepest slope of TIC, and E(max/1) were strongly correlated with MVD (r = -0.765, p < 0.01; r = 0.681, p < 0.01; r = 0.634, p < 0.01; respectively). MVD, steepest slope of the enhancement curve, E(max/1), and Emax strongly correlated with histologic grade (r = 0.475, p < 0.05; r = 0.683, p < 0.01; r = 0.687, p < 0.01; r = 0.791, p < 0.01; respectively). There was a significant difference between groups of patients with and without metastasis with regard to histologic grade (p < 0.05) and two of the DCE-MRI parameters (p < 0.005 for E(max/1) and p < 0.05 for time to peak enhancement). Discriminant analysis correctly predicted the metastatic occurrence at 2 years in 90.5% of cases using E(max/1) (p < 0.001). Histologic grade resulted in lower rates of discrimination (66.7%; p < 0.05). DCE-MRI parameters may help in the prediction of MVD and histologic grade in CRC and may be used to predict therapeutic outcome. PMID- 15290942 TI - CT colonography: determination of optimal CT technique using a novel colon phantom. AB - The aim of this study was to determine the thickest slice at the lowest radiation dose for detection of colon polyps larger than 5mm in diameter at computed tomographic (CT) colonography. A colon phantom containing haustral folds, flexures, and straight segments was constructed of borosilicate. One hundred forty simulated polyps (5, 7, 10, and 12 mm) of various shapes (sessile, flat, and pedunculated) were attached at different colon locations (wall, base of fold, on the fold and fold tip). Polyps were positioned parallel, perpendicular, and oblique to the CT gantry. The air-filled phantom was scanned at different slice thicknesses (1.25-5 mm) and x-ray tube currents (5-308 mA). All polyps were identified in all data sets except one (1.25 mm slice thickness, 5 mA). In this acquisition, image noise reduced polyp visibility, and five of 140 (3%) polyps could not be identified. Unidentified polyps were 5 mm, flat or sessile in shape, located on the colon wall or base of the fold, and oblique or parallel to CT gantry. All tested CT techniques provided optimal polyp detection except settings at 1.25 mm and 5 mAs. Thin collimation (<5 mm) scans may not be necessary to detect clinically significant polyps. PMID- 15290944 TI - Ultrasound evaluation of liver transplants. PMID- 15290943 TI - Cardiac arrest: abdominal CT imaging features. AB - We report computed tomographic findings of two unusual cases of sudden cardiac arrest. The imaging features documented include reflux of contrast into the abdomen as indicated by opacification of renal veins, hepatic veins, inferior vena cava, and hepatic and renal parenchyma. The reflux of contrast into the portal vein in one patient has not been described in the literature. The thoracic findings were reflux of contrast into the coronary sinus, nonopacificaton of the left ventricle with intravenous contrast, and lack of cardiac motion artifact. PMID- 15290945 TI - Complications of orthotopic liver transplantation: imaging findings. AB - Orthotopic liver transplantation has become the major treatment for end-stage chronic liver disease and for severe acute liver failure. Despite the improvement in survival due to advances in organ preservation, improved immunosuppressive therapy agents, and refinement of surgical techniques, there are significant complications after liver transplantation. These complications mainly include biliary strictures, stones, and leakage; arterial and venous stenoses and thromboses; lymphoproliferative disorders; recurrent tumors; hepatitis virus C infection; liver abscesses; right adrenal gland hemorrhage; fluid collections; and hematomas. The diagnosis of acute rejection, one of the most serious complications after liver transplantation, is established with graft biopsy and histologic study. The role of imaging methods consists of excluding the other complications, which can have clinical signs and symptoms similar to those of acute rejection. This pictorial essay describes imaging findings of the various complications after liver transplantation and focuses on their radiologic diagnosis. Knowledge and early recognition of these complications with the most suitable imaging modality are crucial for graft and patient survival. PMID- 15290946 TI - Hepatocellular carcinoma in advanced liver cirrhosis: CT detection in transplant patients. AB - Computed tomography (CT) is being used as the standard pretransplantation imaging for recipients and donors in the evaluation of liver volume, liver reserve function, vascular anatomy, diagnosis of hepatocellular carcinoma and metastasis, and global information of the abdominal cavity. Whereas CT detection of hepatocellular carcinoma in non-cirrhotic patients is satisfactory, detection sensitivity in severely cirrhotic patients is limited, with a reported sensitivity of 53% to 68%. Tumors smaller than 2 cm are more difficult to detect. Innumerable regenerative nodules, localized or diffuse fibrosis, arterioportal shunts, nodular surface, and distorted anatomy in end-stage liver cirrhosis make it difficult to detect small hepatocellular carcinoma. Because of the shortage of cadavers and living donors, judicious use of CT is necessary in the selection of candidates and the decision of priority for liver transplantation in patients with advanced liver cirrhosis. PMID- 15290947 TI - Multiple infarcted regenerative nodules in liver cirrhosis after systemic hypotension due to septic shock: radiologic findings. AB - We describe a case of multiple infarcted regenerative nodules in a patient with advanced liver cirrhosis who had experienced an episode of septic shock. Sonography showed multiple hypoechoic or isoechoic nodules; contrast-enhanced computed tomography showed multiple, low-attenuating nodules with rim enhancement; and magnetic resonance imaging showed multiple nodules of low, iso-, or high signal intensity. Explanted liver showed coagulation necrosis of multiple regenerative nodules. Peribiliary cysts in chronic liver diseases, liver abscesses, spontaneous necrosis of hepatocellular carcinomas, and metastasis should be differentiated. PMID- 15290948 TI - Hepatic involvement in hereditary hemorrhagic telangiectasia: CT findings. AB - Hereditary hemorrhagic telangiectasia (HHT), also known as Rendu-Osler-Weber disease, is an autosomal-dominant vascular disease characterized by mucocutaneous or visceral angiodysplastic lesions (telangiectases and arteriovenous malformations) that may be widely distributed throughout the cardiovascular system. The recognition of mucocutaneous telangiectases, the occurrence of spontaneous and recurrent episodes of epistaxis, the presence of visceral involvement, and a family history of this disease are the clinical criteria that allow diagnosis. In comparison with skin, lungs, gastrointestinal tract, and brain involvement, hepatic involvement defined by clinical criteria alone has long been considered uncommon. Our experience with a large group of HHT patients, even those asymptomatic for liver involvement, demonstrates that it is more frequent than reported and is characterized by the presence of intrahepatic shunts, disseminated intraparenchymal telangiectases, and other vascular lesions. Congestive cardiac failure, portal hypertension, portosystemic encephalopathy, cholangitis, and atypical cirrhosis have been reported as possible serious complications related to this condition. Thus, a correct diagnosis is important, and diagnostic imaging has a fundamental role in detecting alterations involving the liver. The possibilities to perform a multiphasic study and to provide high quality multiplanar and angiographic reconstructions, gives multidetector row helical computed tomography the ability to detect and characterize the complex anatomopathologic alterations typical of this disease. PMID- 15290949 TI - Epithelioid hemangioendothelioma of the liver: radiologic-pathologic correlation. AB - Epithelioid hemangioendothelioma of the liver is a rare vascular tumor with intermediate malignant potential. On imaging studies, the lesion has a solid appearance and may mimic metastatic disease. We present a case in which the morphologic features (multifocal aspect, peripheral location, and capsular retraction) and the clinical history aided in including this entity in the differential diagnosis. PMID- 15290950 TI - Acute, complete splenic infarction in cancer patient is associated with a fatal outcome. AB - Splenic infarction frequently occurs in patients with myeloproliferative diseases, endocarditis, and sickle cell anemia. Various sonographic patterns of splenic infarction do exist. but little is known about tumor associated splenic infarction in cancer patients. Between January 1992 and December 2002, 66 patients were diagnosed with splenic infarction by color Doppler sonography (CDS). Ten patients had an underlying solid cancer. Clinical and sonographic data of cancer patients were evaluated retrospectively with regard to age, sex, frequency of thrombotic episodes, splenic size, echomorphology and vascularity of splenic lesions, and follow-up examination. The median age was 53 years (range, 16-73 years). Nine of 10 patients had abdominal metastases, four had evidence of a hypercoagulable state, five had a small spleen (< 7 x 3 cm), and seven had acute complete infarction of the spleen without hilar and parenchymal vessels on CDS. Survival of six patients with acute complete infarction ranged from 1 to 30 days. In cancer patients with splenic infarction, an acute complete infarction is the most common pattern. It is caused predominantly by a hypercoagulable state and is associated with an extremely short survival. PMID- 15290951 TI - Fatal, complete splenic infarction and hepatic infection due to disseminated Trichosporon beigelii infection: CT findings. AB - We report on a 56-year-old woman with acute lymphocytic leukemia who presented with right upper quadrant pain, fever, nausea, and vomiting. Laboratory studies confirmed fungemia with Trichosporum beigelii, and contrast-enhanced computed tomography of the abdomen demonstrated numerous low-attenuation liver lesions and a hypodense spleen with capsular enhancement suggestive of complete splenic infarction. Subsequent splenectomy confirmed that the spleen was completely infarcted and infiltrated with Trichosporum. The patient had a difficult postoperative course and died despite aggressive antifungal therapy. PMID- 15290952 TI - Use of thin-section, multidetector row helical CT images for coronal oblique reformations for optimal visualization of structures in the hepatoduodenal ligament. AB - Multidetector row computed tomography (CT) can acquire abdominal images of unprecedented thinness in a single breath-hold. This study investigated whether acquiring source axial images at 1.25 mm as opposed to 2.5 mm would result in a perceptible difference in image quality for coronal oblique reformations. Similarly, the hypothesis that a slice pitch of 3:1 would be superior to 6:1 was evaluated. Twenty-nine CT studies were retrospectively evaluated. The images were divided into four groups: 1.25-mm axial images, pitch 3:1; 2.5-mm axial images, pitch 3:1; 1.25-mm axial images, pitch 6:1; and 2.5-mm axial images, pitch 6:1. Three radiologists evaluated by consensus the coronal oblique reformations for overall image quality and image quality of structures in the hepatoduodenal ligament and of nodal groups. Use of 1.25-mm rather than of 2.5-mm source axial images resulted in statistically significant better scores for overall image quality and visualization of the hepatic artery, portal vein, pancreatic duct, and nodal groups. However, a pitch of 3:1 rather than of 6:1 did not result in significant differences in ratings of image quality. Use of 1.25-mm rather than of 2.5-mm source axial images improves image quality when creating coronal oblique reformations for abdominal anatomy. PMID- 15290953 TI - Vascular invasion in pancreatic cancer: value of multislice helical CT. AB - The evaluation of resectability in patients with pancreatic cancer remains a diagnostic challenge, especially regarding the preoperative assessment of vascular invasion. Thin-slice multiplanar reconstructions obtained with multislice helical computed tomography provide an exact depiction of the spatial relation between the tumor and the potentially invaded vessels and thus have the capability to improve the assessment of local resectability. PMID- 15290954 TI - Neuroendocrine pancreatic tumor: value of contrast enhanced ultrasonography. PMID- 15290955 TI - Arteriovenous malformation of the pancreas in a patient with gastrointestinal bleeding: helical CT findings. AB - Dynamic computed tomographic (CT) findings of arteriovenous malformation (AVM) of the pancreas include strong enhancement or conglomeration of small hypervascular spots in the pancreas and early contrast filling of the portal vein during the arterial phase. We describe a case with pancreatic AVM in which we identified enlarged arterial feeders and draining veins as supportive findings of the diagnosis and ulceration into the pancreatic duct as a possible cause of gastrointestinal bleeding at contrast-enhanced CT. PMID- 15290956 TI - Ureteral colic: US versus CT. PMID- 15290957 TI - Embolization of uterine fibroids. AB - Since the first description of uterine artery embolization for the treatment of symptomatic fibroids of the uterus in 1994, this minimally invasive procedure has been increasingly performed in many Western countries. The method is characterized by a high technical success rate of about 85%, a highly significant relief of symptoms, and a very low rate of complications that make this method an appealing alternative to classic treatment options of surgical or laparoscopic myomectomy or hysterectomy. These characteristics have made the procedure well accepted by affected women. Nevertheless, indications and potential contraindications have to be evaluated carefully, especially in patients of childbearing age whenever a considerable number of deliveries is reported after uterine fibroid embolization. This article discusses the clinical background, indications and contraindications, angiographic techniques, potential complications and side effects, and the mid-term results known at present. PMID- 15290958 TI - Plea for using the term n-7 fatty acids in place of C18:2 cis-9, trans-11, and C18:1 trans-11 or their trivial names rumenic acid and vaccenic acid rather than the generic term conjugated linoleic acids. PMID- 15290959 TI - Purification and characterization of a pregastric esterase from a hygienized kid rennet paste. AB - Rennet pastes obtained by maceration of gastric tissues from suckling kids are used traditionally to produce some artisanal cheeses in Spain. Besides milk clotting function, rennet pastes provide proteolytic activity and lipolytic system, essentially pregastric, necessary in the development of piquant flavor typical of these cheeses. A simple and reproducible procedure allows us to obtain a standardized rennet paste that posses the desired activity and is of good microbiological quality. Concomitantly, a kid pregastric esterase (KPGE) was purified to homogeneity. The purification procedure was based on an aqueous extract of hygienized rennet paste (HRP), which was chromatographed on DEAE Sepharose Fast Flow then adsorbed on phenyl superose followed by a re chromatography on the same column. The final enzymatic preparation, where the overall activity recovery was 3%, showed a molecular mass of 53 kDa. The highest activity was determined on p-nitrophenyl butyrate, but marked hydrolysis was also detected on beta-naphthyl caprylate. In contrast, low activity on tributyrin (substrate under emulsion form) was detected, thus confirming the esterase character of purified enzyme. PMID- 15290960 TI - Evaluation of salt whey as an ingredient in processed cheese. AB - The objective of this research was to determine whether salt whey, obtained from a traditional Cheddar cheese manufacturing process, could be used as an ingredient in processed cheese. Due to its high salinity level, salt whey is underutilized and leads to disposal costs. Consequently, alternative uses need to be pursued. The major components of salt whey (salt and water) are used as ingredients in processed cheese. Three replicates of pasteurized processed cheese (PC), pasteurized processed cheese food (PCF), and pasteurized processed cheese spread (PCS) were manufactured. Additionally, within each type of processed cheese, a control formula (CF) and a salt whey formula (SW) were produced. For SW, the salt and water in the CF were replaced with salt whey. The composition, functionality, and sensory properties of the CF and SW treatments were compared within each type of processed cheese. Mean melt diameter obtained for the CF and SW processed cheeses were 48.5 and 49.4 mm, respectively, for PC, and they were 61.6 and 63 mm, respectively, for PCF. Tube-melt results for PCS was 75.1 and 79.8 mm for CF and SW treatments, respectively. The mean texture profile analysis (TPA) hardness values obtained, respectively, for the CF and SW treatments were 126 N and 115 N for PC, 62 N and 60 N for PCF, and 12 N and 12 N for PCS. There were no significant differences in composition or functionality between the CF and SW within each variety of processed cheese. Consequently, salt whey can be used as an ingredient in PC without adversely affecting processed cheese quality. PMID- 15290961 TI - Predominant microflora of downgraded Danish bulk tank milk. AB - The microflora of downgraded Danish bulk tank milk was examined to identify the main causes of increased microbial counts. Seventy-five representative samples with a microbial count exceeding 3.0 x 10(4) cfu/mL were selected for a more detailed microbial examination. A total of 1237 isolates from these samples were identified. Gram-negative, oxidase-positive bacteria were found in 72% of the samples. Coliforms were found in 20% of the samples, and non-coliforms were found in 49% of the samples. Coryneforms, other gram-positive rods, Lactococcus spp., Micrococcus spp., and coagulase-negative Staphylococcus spp. were found in 28 to 53% of the samples. Bacillus spp., Enterococcus spp., Staphylococcus aureus, Streptococcus dysgalactiae, Streptococcus uberis, and yeasts were found in <25% of the samples. Additionally, the isolates were divided into 3 groups, based on the main cause of an elevated microbial count. Microorganisms primarily associated with poor hygiene dominated the microflora in 64% of the samples; bacteria also related to poor hygiene, but in addition associated with growth at low temperatures (psychrotrophic bacteria) dominated the microflora in 28% of the samples; and bacteria mainly associated with mastitis dominated the microflora in 8% of the samples. A bulk tank milk storage period of 48 h instead of 24 h did not affect the proportion of downgraded milk samples and could not be associated with a specific group of microorganisms. Further, no relationship was found between somatic cell counts and the presence of mastitis bacteria. PMID- 15290962 TI - Lactose/beta-lactoglobulin interaction during storage of model whey powders. AB - The objective of this study was to evaluate the presence or absence of interaction between lactose and beta-lactoglobulin during storage of model whey powders at different water activities (a(w)). Model whey powders were prepared by colyophilization of lactose with increasing quantities of beta-lactoglobulin. These colyophilized beta-lactoglobulin:lactose powders, assigned as BL powders, were stored from 0.11 to 0.95 a(w). The water sorption behavior of BL powders was studied gravimetrically, and the state of lactose was investigated using differential scanning calorimetry (DSC) and scanning electron microscopy (SEM). Before storage, BL powders were amorphous. After storage, a loss of water was observed on moisture sorption isotherms of BL powders. It was related to the formation of lactose crystals, detected by DSC and SEM analysis, and to the structural collapse of the powders. Water loss due to lactose crystallization was shifted to higher a(w) with increasing beta-lactoglobulin content in BL powders. Moreover, kinetics of moisture sorption demonstrated that beta-lactoglobulin was also responsible for a slower crystallization process in BL powders. Then, the water sorption behavior of BL powders was very different from the behavior of the 2 compounds mixed after separate lyophilization. All these results pointed out interaction between lactose and beta-lactoglobulin, which appeared during lyophilization and still occurred during storage. This lactose/beta-lactoglobulin interaction stabilized model whey powders against lactose crystallization. PMID- 15290963 TI - Gelation of casein-whey protein mixtures. AB - Heated milk consists of a mixture of whey protein-coated casein micelles and soluble whey protein aggregates. The acid-induced gelation properties of heated milk are consistently different from those of unheated milk--i.e., a shift in gelation pH, stronger gels, and a different microstructure of the gels. In this study we investigated the role of the different fractions of denatured whey proteins on the acid-induced gelation, the gel hardness, and the microstructure. Both whey protein fractions contribute to the observed shift in gelation pH, although by a different mechanism. Obtaining gels with high gel hardness occurs most effectively when all denatured whey proteins are present as whey protein aggregates. It was observed that disulfide bridge exchange reactions during the acid-induced gelation at ambient temperature play an important role for both whey protein fractions. Additionally, disulfide interactions seem to occur between the aggregates and the casein micelles during the gel state. In this study, we show the development of a new approach for confocal scanning laser microscopy measurements--i.e., separate staining of the proteins in milk. By using this method, we were able to determine that, although whey protein aggregates are not linked to the casein micelles, they nevertheless gel at the same moment. This work adds to a better understanding of the role of denatured whey proteins during acid-induced gelation and could improve the effective use of whey proteins. PMID- 15290964 TI - Impact of source and level of calcium fortification on the heat stability of reconstituted skim milk powder. AB - Calcium enrichment of food and dairy products has gained interest with the increased awareness about the importance of higher calcium intake. Calcium plays many important roles in the human body. Dairy products are an excellent source of dietary calcium, which can be further fortified with calcium salts to achieve higher calcium intake per serving. However, the addition of calcium salts can destabilize food systems unless conditions are carefully controlled. The effect of calcium fortification on the heat stability of reconstituted skim milk was evaluated, using reconstituted skim milks with 2 protein levels: 1.75 and 3.5% (wt/wt) prepared using low and high heat powders. Calcium carbonate, phosphate, lactate, and citrate were used for fortification at 0.15, 0.18, and 0.24% (wt/wt). Each sample was analyzed for solubility, heat stability, and pH. The addition of phosphate and lactate salts lowered the pH of milk, citrate did not have any major effect, and carbonate for the 1.75% protein samples increased the pH. In general, changes in solubility and heat stability were associated with changes in pH. Calcium addition decreased the solubility and heat stability. However, interestingly, the presence of carbonate salt greatly increased the heat stability for 1.75% protein samples. This is due to the neutralizing effect of calcium carbonate when it goes into solution. The results suggested that the heat stability of milk can be affected by the type of calcium salt used. This may be applied to the development of milk-based calcium enriched beverages. PMID- 15290965 TI - Changes in cisternal udder compartment induced by milking interval in dairy goats milked once or twice daily. AB - Fourteen Murciano-Granadina dairy goats were used to evaluate udder compartments (cisternal and alveolar) and cisternal recoil after an oxytocin (OT) challenge at different milking intervals (8, 16, and 24 h) during wk 7 of lactation. Goats were milked once (1x; n = 7) or twice (2x; n = 7) daily from wk 2 of lactation. Average milk yields for wk 4 and 8 were 1.76 and 2.24 L/d, for goats milked 1x and 2x, respectively. For each half udder, cisternal area was measured by ultrasonography and cisternal milk was measured by machine milking after i.v. injection of an OT receptor blocking agent. Alveolar milk was then obtained after i.v. injection of OT. Regardless of milking frequency, alveolar milk increased from 8 to 16 h after milking, but did not change thereafter. Cisternal area and cisternal milk increased linearly (R2 = 0.96 to 0.99) up to 24 h, indicating continuous milk storage in the cistern at any alveoli filling degree. Cisternal to alveolar ratio increased with milking interval (from 57:43 to 75:25), but differences between milking intervals were significant at 8 h only, at which time goats milked 2x showed a greater ratio (1x = 51:49; 2x = 62:38). Despite extended milking intervals, cisterns of goats milked 1x did not become larger than cisterns of goats milked 2x after 5 wk of treatment. The highest correlation between cisternal area and cisternal milk was detected at 8 h after milking (r = 0.74). Primiparous goats had smaller cisternal areas and less cisternal milk than multiparous goats at all milking intervals. Cisternal recoil was studied in a sample of multiparous goats milked 1x (n = 4) and 2x (n = 4) by scanning cisterns by ultrasonography at 0, 5, 15, and 30 min after an OT challenge for each milking interval. Cisternal area increased after OT injection for the 8- and 16-h milking intervals, but no differences were observed for the 24-h interval. Unlike cows, no changes in cisternal area were observed after OT injection, indicating the absence of cisternal recoil in goats. We conclude that goats show a large cisternal compartment that increases linearly after milking. Nevertheless, cisternal size did not increased after 1x milking, probably because of lesser milk yield. Multiparous goats had larger cisterns than primiparous goats and were able to store more milk in their cisterns at all milking intervals. Because of the high capacity of goat cisterns, no milk return from cistern to alveoli is expected if milking is delayed after milk letdown. PMID- 15290966 TI - Differential effects of steroids and retinoids on bovine myelopoiesis in vitro. AB - Pregnancy and parturition are associated with physiological changes caused by steroid hormones. Alterations in number, maturity, and function of polymorphonuclear leukocytes observed in dairy cows at parturition suggest a common causative relationship with steroid hormones. This study was designed to investigate the effects of progesterone, 17-beta-estradiol, and hydrocortisone on the proliferation of bovine progenitor cells. An in vitro culturing system was used, and colonies were scored after 7 d of incubation. At low concentrations, 17 beta-estradiol inhibited proliferation of granulocyte progenitor cells. Hydrocortisone reduced growth of granulocyte and monocyte colonies, whereas myelopoiesis was not altered by progesterone. Furthermore, we studied the effect of retinoids on colony formation of bovine bone marrow cells. All-trans- and 9 cis-retinoic acid stimulated growth of granulocyte colonies and inhibited proliferation of the monocyte lineage. The addition of the 13-cis-isomer also increased numbers of granulocyte colony-forming units. This study indicates that steroid hormones may be responsible for alterations in the bovine hematopoietic profiles observed in circulation during the postpartum period. White blood cells, especially polymorphonuclear leukocytes, which are derived from bone marrow, are an important first line defense against mastitis. Therefore, these effects of steroids might contribute to the increased susceptibility of dairy cows to Escherichia coli mastitis. We furthermore hypothesize that an important role might be attributed to retinoic acid in its regulation of bovine myelopoiesis. Modulation of myelopoiesis in favor of the granulocyte lineage during the acute phase reaction may be an adaptive mechanism designed to increase the capacity of first-line defense to intramammary infections. PMID- 15290967 TI - Use of conjugated linoleic acid (CLA) enrichments to examine the effects of trans 8, cis-10 CLA, and cis-11, trans-13 CLA on milk-fat synthesis. AB - Conjugated linoleic acid (CLA) supplements have typically been comprised of 4 isomers (trans-8, cis-10; cis-9, trans-11; trans-10, cis-12; and cis-11, trans-13 CLA). Abomasal infusion of pure isomers has shown that trans-10, cis-12 CLA is a potent inhibitor of milk-fat synthesis, whereas cis-9, trans-11 CLA has no effect. However, there appear to be additional fatty acids that inhibit milk-fat synthesis, and the objective of this study was to investigate the effects of additional CLA isomers present in CLA supplements. Four rumen fistulated Holstein cows (141+/-8 DIM, mean+/-SE) were randomly assigned in a 4 x 4 Latin square experiment. Treatments were abomasal infusion of (1) skim milk (negative control), (2) trans-10, cis-12 CLA supplement (positive control), (3) trans-8, cis-10 CLA supplement, and (4) cis-11, trans-13 CLA supplement. Treatments 2 through 4 were targeted to provide 4 g/d of the CLA isomer of interest. The trans 8, cis-10 CLA supplement had no effect on milk-fat yield, whereas the trans-10, cis-12 CLA supplement reduced milk-fat yield by 35%. The cis-11, trans-13 CLA supplement contained some trans-10, cis-12 CLA, and when data were compared to the positive control treatment group, it was obvious that cis-11, trans-13 CLA also had no effect on milk-fat synthesis. Milk-fat content of specific CLA isomers was significantly elevated within respective treatment groups. Milk yield, DMI, and milk protein yield were unaffected by treatment. Overall, trans 10, cis-12 CLA reduced milk-fat synthesis, whereas the other major isomers present in CLA supplements (trans-8, cis-10 CLA and cis-11, trans-13 CLA) had no effect. PMID- 15290968 TI - Effect of rubber flooring in front of the feed bunk on the time budgets of dairy cattle. AB - The objective of this experiment was to study the effect of rubber flooring in front of the feed bunk on the immediate behavioral response of dairy cattle. Four groups of 12 dairy cattle were alternately housed in sections of a free-stall barn with either 1.85 m of rubber flooring or grooved concrete in the area in front of the feed bunk. Rubber flooring did not affect time spent eating. However, animals showed a slight, but detectable, increase in time standing without eating on the rubber surface (5.5%) compared with concrete (4.8%). For reasons that are unclear, this increase in time spent standing was not limited to the area in front of the feed bunk; animals spent 11.0% of the available time standing elsewhere in the pen (outside of the free stall but not in front of the feed bunk) when they had access to the rubber flooring, compared with 9.0% when housed with access to only concrete floors. In addition, animals spent slightly less time lying in the free stall when they had access to rubber in front of the feed bunk (52.5 vs. 54.3%). Time spent engaged in behaviors such as standing elsewhere in the pen and eating were variable over time. For example, time spent eating declined from 23.1 to 17.4% over the 6-wk trial. In conclusion, dairy cattle with access to rubber flooring in front of the feeder showed small differences in where and how much time they spent standing, although the biological implications of these small changes are unclear. PMID- 15290969 TI - Free-stall dimensions: effects on preference and stall usage. AB - In 2 experiments, free-stall dimensions were examined to determine how they affected stall preference, usage, cleanliness, and milk production in Holstein dairy cattle. In experiment 1, stall width (112 or 132 cm) and stall length (229 and 274 cm from curb to wall) were compared in a 2 x 2 factorial arrangement of stall treatments using 15 individually housed, non-lactating animals. Cows showed no clear preference for stall size as measured by lying time. When animals had no choice between stalls, average lying time was higher in the wide stalls than in the narrow stalls (10.8 vs. 9.6+/-0.3 h/24 h). Both length and width affected time spent standing with only the front hooves in the stall; total stall area is best explained by the variation associated with this behavior. In experiment 2, 27 lactating dairy cattle were alternately housed with access to stalls of 106, 116, or 126 cm in width using a cross-over design with exposure to each treatment lasting 3 wk. Animals spent an additional 42 min/24 h lying in stalls measuring 126 cm in width compared with stalls with only 106 cm between partitions. Free stall width influenced the time spent standing with the front 2 hooves in the stall; animals averaged 58 min/24 h in the widest stalls and 85 min/24 h in the narrowest stalls. The amount of time spent standing with all 4 hooves in the stall tended to be longer in wider stalls, and these stalls were also most likely to become soiled with feces. Stall width did not affect the number of lying events or milk production. In conclusion, animals spent more time lying down, and less time was spent standing with only the front hooves in larger stalls, but larger stalls were also more likely to become soiled. PMID- 15290970 TI - Systematic clinical examinations for identification of latent udder health types in Danish dairy herds. AB - A cross-sectional study was conducted to explore the applicability of systematic clinical examinations of udders as an additional tool for the evaluation of udder health status on dairy farms. During 2000, each of the 16 dairy farms was visited 5 times; 20 cows per farm were chosen at random at each visit for clinical udder examination immediately after milking. The clinical examination included both pathological and morphological variables. One examination per cow was included in the analysis (n = 707 cows). Principal component analysis (PCA) was performed in 3 steps. First, 19 variables characterizing udder and teats were analyzed (PCA 1). Second, the variables parity and stage of lactation were included (PCA 2). Finally, somatic cell count (SCC) and milk yield (PCA 3) were included. The PCA resulted in 4 components that explained 30% of the variation of the data: 1) small udder, 2) distressed udder, 3) mastitis udder, and 4) soiled udder. Variables with high positive correlation to the "small udder" were small udder shape, short teats, and first parity. Impaired teat surface, hard udder texture, and a long udder shape were related to the "distressed udder." The "mastitis udder" was characterized by the clinical variables asymmetry between front quarters, asymmetry between hind quarters, knotty tissue, and acute clinical mastitis. Reduced milk yield and high SCC were related to the "mastitis udder," whereas low SCC was related to the "small udder." The "soiled udder" was related to early lactation. Including this information in the assessment of udder health may be of substantial value for data analysis in farms with suspected under reporting of clinical mastitis. PMID- 15290971 TI - Effect of dietary energy and somatotropin on components of the somatotropic axis in Holstein heifers. AB - The somatotropic axis, consisting of growth hormone (GH), GH receptor (GHR), insulin-like growth factor (IGF)-I, IGF binding proteins (IGFBP), and IGF receptors, controls growth and mammary development in heifers. Manipulation of the axis with recombinant bovine somatotropin (rbST) improves heifer growth and reduces age at first calving. The effects of rbST are influenced by dietary energy through partially understood mechanisms. The objective was to characterize the somatotropic axis in Holstein heifers fed a diet for either low or high rate of gain and treated with or without rbST. Heifers (120 d of age) were assigned to one of 2 diets to gain either 0.8 kg/d (low, n = 18) or 1.2 kg/d (high, n = 20). Within each diet, half of the heifers (n = 9 for low and n = 10 for high) received daily rbST injections (25 microg/kg of body weight). Treatments and diets continued until slaughter (2 mo after puberty). Blood was collected 2x per week, and a frequent sampling window was performed 1 d before slaughter. Liver was collected at slaughter. Feeding a high diet or treating with rbST increased serum IGF-I and decreased serum IGFBP-2. The observed changes in serum IGF-I and IGFBP-2 were reflected in their respective liver mRNA amounts. Feeding a high diet decreased serum GH concentrations after rbST injection, but the stimulatory effect of rbST on serum IGF-I was nonetheless greater in high-diet heifers. The differential IGF-I response may be explained by greater GHR 1A in the liver of high-diet heifers. We conclude that a high-gain diet modifies the somatotropic axis in rbST-treated heifers by decreasing serum GH but increasing serum IGF-I after rbST treatment. Greater IGF-I (indicative of an increased GH response) may be a consequence of greater GHR 1A expression in the liver. PMID- 15290972 TI - Effects of oxytocin administration on oxytocin release and milk ejection. AB - In experiment I, the effect of i.m. oxytocin (OT) injection (50 IU) on OT blood pattern was tested. Blood samples from 6 cows were collected for 2 h after OT injection either with or without milking. To test the effect of i.m. OT injection (50 IU) on milk ejection efficiency, intramammary pressure (IMP) was measured in 13 cows (experiment II). Milk ejection was induced by manual teat stimulation. After IMP increased and reached a plateau, OT was injected. In experiment III, the effect of chronic OT treatment on mammary gland sensitivity and endogenous OT release was tested. For 19 d, cows were i.m. injected at each milking with 50 IU OT (n = 13) or 5 mL of NaCl 0.9% (n = 14) 1 min before the start of udder preparation. To test mammary gland sensitivity, IMP recording was performed after a long (11 h, 7 OT cows, and 7 NaCl cows) and a short (3 h, 6 OT cows, and 7 NaCl cows) milking interval at d -1 and d 18. To test the effect of withdrawal of chronic i.m. treatment on OT release, blood sample collection was performed during evening milking at d 0 and d 19. Intramuscular oxytocin injection (experiment I) caused elevated OT blood levels observed at least for 2 h and showed an even more pronounced effect when milking was also performed. Intramuscular OT injection after teat stimulation (experiment II) caused additional milk ejection but only in 6 out of 13 cows. Withdrawal of chronic OT treatment (experiment III) did not reduce OT release during milking. However, ejection time was prolonged during OT infusion after a long milking interval. Ejection pressure tended to be lower after a short milking interval. It seems that the reduction of spontaneous milk removal after chronic OT treatment was due to reduced contractibility of myoepithelial cells in the mammary gland at a physiological range of OT concentrations. PMID- 15290973 TI - Relative acidifying activity of anionic salts commonly used to prevent milk fever. AB - High cation diets can cause milk fever in dairy cows as they induce a metabolic alkalosis reducing the ability of the cow to maintain calcium homeostasis at the onset of lactation. Adding anions to the diet can offset the effect of the high cation forages by inducing a mild metabolic acidosis, restoring the ability to maintain calcium homeostasis. The difference in mEq of dietary cations and anions (DCAD) is most often expressed as (Na(+) + K+) - (Cl- + S(--)). This equation implies that a mEq of chloride and a mEq of sulfate are equipotent in their ability to alter acid-base balance of the cow. Using blood and urine pH to monitor effects on acid-base balance, experiments were conducted to test the relative acidifying activity of various sulfate and chloride anion sources in nonpregnant, nonlactating Jersey cows. Across all experiments, chloride proved to have about 1.6 times the acidifying activity of sulfate. Calcium and magnesium, ignored by the common DCAD equation, had a small but significant alkalinizing effect when accompanying chloride or sulfate. The ranking of the anion sources tested at a dose of 2 Eq/d, from most to least potent urine acidifier, was hydrochloric acid, ammonium chloride, calcium chloride, calcium sulfate, magnesium sulfate, and sulfur. These data should allow more accurate prediction of the response of late gestation cows to dietary cation-anion manipulation. PMID- 15290974 TI - Somatic cell count distributions during lactation predict clinical mastitis. AB - This research investigated somatic cell count (SCC) records during lactation, with the purpose of identifying distribution characteristics (mean and measures of variation) that were most closely associated with clinical mastitis. Three separate data sets were used, one containing quarter SCC (n = 1444) and two containing cow SCC (n = 933 and 11,825). Clinical mastitis was defined as a binary outcome, present or absent, for each lactation, and SCC were log (base 10) transformed. A generalized linear mixed model within a Bayesian framework was used for analysis. Parameters were estimated using Markov Chain Monte Carlo with Gibbs sampling. Results from the 3 data sets were similar. Increased maximum and standard deviation log SCC during lactation, rather than increased geometric mean, were the best overall indicators of clinical mastitis. Distributions of SCC were also investigated separately for different mastitis pathogens. Increased maximum log SCC was associated with clinical mastitis caused by all pathogen types. Increased standard deviation log SCC was associated with Staphylococcus aureus, and Streptococcus uberis clinical mastitis and increased coefficient of variation log SCC (standard deviation divided by mean) was associated with Escherichia coli clinical mastitis. Increased geometric mean lactation SCC was associated with an increased risk of Staph. aureus clinical mastitis but a reduced risk of E. coli clinical mastitis. Our results suggest that using measures of variation and maximum cow SCC would enhance the accuracy of predicting clinical mastitis, compared with geometric mean SCC, and therefore improve genetic programs that aim to select for clinical mastitis resistance. The results are also consistent with low SCC increasing susceptibility to some mastitis pathogens. PMID- 15290975 TI - Partial replacement of corn grain by hydrogenated oil in grazing dairy cows in early lactation. AB - Thirty-six grazing dairy cows were used to determine milk production and composition, and dry matter and energy intake when corn grain was partially replaced by hydrogenated oil in the concentrate. Four additional cows, each fitted with a ruminal cannula, were used in a crossover design to evaluate effects of supplemental fat on rumen environment and pasture digestion. All cows grazed mixed pastures with an herbage allowance of 30 kg dry matter/cow per day. The control group was fed a concentrate containing corn grain (4.49 kg dry matter/cow per day) and fishmeal (0.37 kg dry matter/cow per day), whereas the other group (fat) received a concentrate containing corn grain (2.87 kg dry matter/cow per day), fishmeal (0.37 kg dry matter/cow per day) and fat (0.7 kg dry matter/cow per day). The fat was obtained by hydrogenation of vegetable oils (melting point 58 to 60 degrees C, 30.3% C16:0, 34.9% C18:0, 21.8% C18:1, 3.3% C18:2). Supplemental fat increased milk production (control = 23.7 vs. fat = 25.0 kg/cow per day), fat-corrected milk (control = 22.5 vs. fat = 24.5 kg/cow per day), milk fat content (control = 3.64% vs. fat = 3.86%) and yields of milk fat (control = 0.86 vs. fat = 0.97 kg/cow per day) and protein (control = 0.74 vs. fat = 0.78 kg/cow per day). Milk percentages of protein, lactose, casein, cholesterol, and urea nitrogen were not affected. Pasture DMI and total DMI of pasture and concentrate and estimated energy intake were unchanged. No differences in loss of body weight or body condition score were detected. Plasma concentrations of nonesterified fatty acids, somatotropin, insulin, and insulin like growth factor were not affected by supplemental fat. Concentrations of plasma triglyceride and total cholesterol were increased by supplemented fat, and no changes in plasma glucose and urea nitrogen were observed. The acetate-to propionate ratio was higher in rumen fluid of cows that consumed fat (fat = 3.39 vs. control = 3.27). In situ pasture NDF degradation was not affected. The partial replacement of corn grain with fat improved the productive performance of early-lactation cows grazing spring pastures. No negative effects of supplemental fat on ruminal fiber digestion were detected. PMID- 15290976 TI - Milk protein synthesis as a function of amino acid supply. AB - Most prediction schemes of milk protein secretion overestimate milk protein yield from dairy cows at high protein intakes, thereby overestimating milk protein yield response to protein supplementation. This study was conducted to determine factors contributing to such an overestimation. Using published studies, a database was constructed that was limited to amino acid (AA) infusion studies, as then only the digestible amino acid of dietary origin needed to be estimated, whereas the amount infused was known exactly, thereby reducing the dependence on estimated values. Although milk protein yield was positively related with total energy supply, and both digestible duodenal supply and infused AA, in this database there was no relationship between milk protein yield response above control treatments and the nutrient status of the cows (energy or protein). Total milk protein yield was defined as a function of individual AA supply, using a segmented-linear and a logistic model to obtain estimates of the efficiency of conversion of AA into milk protein. Except for Lys and Met supply, the segmented linear model yielded lower root mean square error and better correlation, but both models were similar in their reliability. For both models, the estimated efficiency of conversion of AA to milk differed among AA. Estimations of the ideal profile of AA for lactating dairy cows were similar between models, with requirements for Lys and Met in line with 2001 National Research Council recommendations. The major difference is that the segmented-linear model yields a constant efficiency of conversion of an AA until requirements are met, with zero efficiency beyond this point. The logistic model allows for an estimation of the decreasing marginal efficiency of conversion of AA as the supply approaches the requirements. The use of variable efficiency factors should improve our ability to predict protein yield in response to supplemental protein. PMID- 15290977 TI - Effect of corn hybrid and chop length of whole-plant corn silage on digestion and intake by dairy cows. AB - The objectives of this experiment were to study the effects of corn hybrid and chop length of whole-plant corn silage (WPCS) on intake, and to quantify ruminal digestive processes that could help to identify factors limiting dry matter intake (DMI). Eight lactating cows and 4 dry cows fitted with a ruminal cannula were randomly assigned to 4 treatments in a 4 x 4 Latin square design with replications for lactating cows and without for ruminally cannulated cows. Treatments were fed in a total mixed ration (TMR) containing 75% WPCS and 25% concentrate. The 4 WPCS differed in the characteristics of 2 conventional hybrids, less degradable vs. more degradable in the rumen and in the chop length, fine vs. coarse. The DMI was measured for all cows, and digestion measurements and chewing activities were recorded with the cannulated cows. With lactating cows, DMI and milk yield varied with corn hybrids but not with chop length. The less degradable hybrid in the rumen was the less ingested. Dry matter intake of dry ears followed the same trend, but the differences between hybrids were lower than that observed with the lactating cows and not significant. Dry matter digestibility in the total tract and rumen fill were not different between hybrids. Ruminal mean retention time was greater for the least degradable hybrid. The rumen fill capacity could explain intake differences between hybrids. Ingestive mastication strongly reduced particle size, and the efficiency of particle size reduction was more important with the coarsely chopped WPCS than the finely chopped ones. The small differences in particle size of material entering the rumen after mastication of WPCS during eating might explain the lack of response for decreasing chop length. Because the rumen fill decreased with the decrease in chop length, rumen fill could not be the only factor responsible for DMI control of WPCS. PMID- 15290978 TI - The effects of buffered propionic acid-based additives alone or combined with microbial inoculation on the fermentation of high moisture corn and whole-crop barley. AB - Buffered propionic acid-based additives (BP) alone or in combination with a microbial inoculant containing lactic acid bacteria (MI) were mixed with ground, high moisture corn or whole-crop barley and ensiled in triplicate laboratory silos to investigate their effects on silage fermentation and aerobic stability. The inoculant and chemicals were applied separately for treatments that included both additives. The addition of MI alone had no effect on DM recovery, fermentation end products, or aerobic stability of high moisture corn. However, treatments with 0.1 and 0.2% BP (alone and the combination) had more than 10- and 100-fold fewer yeasts, respectively, and they also had greater concentrations of propionic acid than did untreated corn. Corn treated with only 0.1 (161 h) and 0.2% (218 h) BP tended to be more stable when exposed to air than untreated corn (122 h). Treatment with MI + 0.2% BP markedly improved the aerobic stability (>400 h) of high moisture corn. With whole-crop barley, the addition of MI alone, BP alone, and combinations of MI and BP prevented the production of butyric acid that was found in untreated silage (0.48%). All barley silages that had MI in their treatments underwent a more efficient fermentation than treatments without MI, as evident by a greater ratio of lactic:acetic acid and more DM recovery than in untreated silage. Increasing levels (0.1 to 0.2%) of BP added together with MI improved the aerobic stability of barley (190 and 429 h) over the addition of MI alone (50 h). These data show that buffered propionic acid-based products are compatible with microbial inoculants and, in some circumstances when used together, they can improve the fermentation and aerobic stability of silages. PMID- 15290979 TI - Dietary preference of dairy cows grazing ryegrass and white clover. AB - The dietary preference of lactating dairy cows grazing perennial ryegrass (Lolium perenne L.) and white clover (Trifolium repens L.) was studied. Twelve groups of 2 lactating, Holstein-Friesian dairy cows grazed 1.2-ha plots containing conterminal monocultures of clover and grass. Half of the groups grazed a plot containing 75% clover and 25% grass (by ground area), with the remaining groups grazing a plot containing 25% clover and 75% grass. The intake rates of clover were higher than those of grass, and intake rates were higher in the evening than in the morning. During daylight hours, clover formed 63.2% of the diet of the groups offered 25% clover, which was higher than the 25% offered but lower than preference for 100% clover. This indicated that cows showed a partial preference for clover, with an overall value (i.e., the mean of the 75% and 25% clover groups) of 73.8%. There was a diurnal pattern to preference, with a stronger preference for clover in the morning and with the preference for grass increasing during the day. The basis for partial preference remains unclear and warrants further research. PMID- 15290980 TI - Bovine somatotropin increases hepatic phosphoenolpyruvate carboxykinase mRNA in lactating dairy cows. AB - Somatotropin (ST) increases milk production and through coordinated changes in hepatic glucose synthesis and amino acid metabolism in dairy cows. The objective of this study was to determine the effects of ST on hepatic mRNA expression for phosphoenolpyruvate carboxykinase (PEPCK) and pyruvate carboxylase (PC), enzymes that are critical to the synthesis of glucose in liver and hepatic mRNA expression for carbamylphosphate synthetase I (CPS-I), argininosuccinate synthetase (AS), and ornithine transcarbamylase (OTC), critical enzymes of the urea cycle. Eighteen cows were randomly allocated to 2 treatment groups and received either recombinant bovine ST (Posilac; Monsanto, St. Louis, MO) or saline injections at 14-d intervals during a 42-d period. Expression of mRNA was determined using Northern blot analysis. Nuclei, isolated from liver biopsy samples, were used to determine effects of ST on transcription rate of PEPCK. Milk production was increased with ST (37.3 vs. 35.1+/-0.6 kg/ d). Plasma NEFA was increased with ST (299 vs. 156+/-34 microM). There were no differences in the expression of CPS-I, AS, and OTC mRNA with ST. Expression of PEPCK and IGF-I mRNA were increased with ST but PC mRNA was unchanged. The data indicate increased PEPCK mRNA in cows given ST and indicates a greater capacity for gluconeogenesis from gluconeogenic precursors that form oxaloacetate. The effects of ST to elevate PEPCK mRNA expression require chronic administration and involve increased transcription of the PEPCK gene. PMID- 15290981 TI - Development and analysis of a rumen tissue sampling procedure. AB - A procedure for rumen tissue sampling was developed to determine treatment effects on rumen development and papillae growth in young calves and to improve repeatability in rumen tissue sampling techniques. Rumens were collected from 42 male Holstein calves from 4 separate experiments. Rumen sampling areas (n = 9) included the caudal dorsal blind sac, cranial dorsal sac, cranial ventral sac, and the caudal and ventral portions of the caudal ventral blind sac. Right and left sides of the rumen were sampled. Five 1-cm2 sections were removed from each area and measured for papillae length (n = 20/area), papillae width (n = 20/area), rumen wall thickness (n = 5/area), and number of papillae per cm2 (n = 5/area). Correlations between areas, samples, and measurements were obtained, and comparisons between experiments, areas, samples, and measurements were performed for all variables. In addition, power analyses were conducted for all variables to determine the efficacy of the procedure in detecting treatment differences. Results indicate that samples should be taken from the caudal and cranial sacs of the dorsal and ventral rumen to sufficiently represent papillae growth and development throughout the entire rumen. The procedure is capable of detecting treatment differences for papillae length and papillae width, has a decreased but acceptable capability of detecting treatment differences for rumen wall thickness, but appears limited in ability to detect treatment differences for papillae per square centimeter. In conclusion, rumen tissue sampling to determine extent of rumen development in calves can be performed in a nonbiased and repeatable manner utilizing a limited number of calves. PMID- 15290982 TI - Sources of variation in rates of in vitro ruminal protein degradation. AB - Rates and extents of ruminal protein degradation for casein, solvent soybean meal (SSBM), expeller soybean meal (ESBM), and alfalfa hay were estimated from net appearance of NH3 and total amino acids in in vitro media containing 1 mM hydrazine and 30 mg/L of chloramphenicol. Protein was added at 0.13 mg of N/mL of medium, and incubations were conducted for 4 to 6 h, usually with hourly sampling. Inocula were obtained from ruminally cannulated donor cows fed diets of grass silage or alfalfa and corn silages plus concentrates. Preincubation or dialysis of inocula was used to suppress background NH3 and total amino acids; however, preincubation yielded more rapid degradation rates for casein and SSBM and was used in subsequent incubations. Preincubation with added vitamins, VFA, hemin, or N did not alter protein degradation. Protein degradation rates estimated for SSBM, ESBM, and alfalfa were not different when computed from total N release or N release in NH3 plus total amino acids, regardless of whether amino acids were quantified using ninhydrin colorimetry or o-phthalaldehyde fluorescence. Accounting for the release of peptide-N also did not affect estimated degradation. However, casein degradation rates were more rapid when using total N release or accounting for peptide-N, indicating significant accumulation of small peptides during its breakdown. Rates also were more rapid with inocula from lactating cows versus nonlactating cows with lower feed intake. Protein degradation rates were different due to time after feeding: casein rate was more rapid, but SSBM and ESBM rates were slower with inocula obtained after feeding. Several characteristics of ruminal inoculum that influenced breakdown of the rapidly degraded protein casein did not appear to have direct effects on degradation of protein in soybean meal. PMID- 15290983 TI - Effect of inhibitor concentration and end-product accumulation on estimates of ruminal in vitro protein degradation. AB - Effects of varying the concentrations of hydrazine sulfate (HS) and chloramphenicol (CAP), inhibitors of microbial-N uptake and protein synthesis, on rates of protein degradation estimated from net appearance of NH3 and total amino acids (TAA) were studied in a ruminal in vitro fermentation system. Without inhibitors, recoveries of N added as NH3 and TAA were 4 and 6% after 4-h incubations, and apparent degradation rates estimated from release of NH3 and TAA for casein, solvent soybean meal (SSBM), and expeller soybean meal (ESBM) approached 0. Increasing inhibitor concentrations from the standard amounts of 1 mM HS plus 30 mg of CAP/L to 2 mM HS plus 90 mg of CAP/ L gave rise to numerically greater N recoveries and degradation rates, but these differences were not statistically significant. Compared with the standard inhibitor concentrations, use of 2 mM HS, without CAP, yielded similar recoveries and rates, but 30 or 90 mg of CAP/L, without HS, was not satisfactory. Versus that with 1 mM HS plus 30 mg of CAP/L, media containing 2 mM HS plus 90 mg of CAP/L gave increased TAA recoveries and higher rates for casein, but not SSBM, in the presence of added starch. Faster degradation rates were obtained for casein, but slower rates for SSBM and ESBM, in Sweden versus Wisconsin using inocula from cows fed different diets but with similar CP and energy contents. Differences in microbial catabolism of peptides may account for differences in degradation rates observed between Sweden and Wisconsin. Adding NH3 plus free and peptide-bound amino acids to the inoculum reduced apparent degradation rates, possibly via end product inhibition. Analysis of data from multiple time-point incubations indicated that casein degradation followed simple, first-order kinetics, while a biexponential model fitted degradation patterns for both SSBM and ESBM. PMID- 15290984 TI - Feeding behavior and performance of dairy cows fed pelleted nonroughage fiber byproducts. AB - The potential of pellets made of soy hulls (SH) and corn gluten feed (CGF) to replace starchy pelleted supplement in diets of lactating cows was measured in a feeding regime comparable to automatic milking systems. Twenty-four cows were divided into 2 equal groups and fed for 7 wk in individual feeders monitored by computer on one of the 2 experimental diets. Both diets contained 75% basic total mixed ration plus an additional 25% of pelleted supplement (17% CP), being either high starch pellets (HST) in treatment, or pellets made of SH + CGF (2:1) (SHCG) in treatment. In vitro dry matter digestibility was higher in the HST pellets, whereas neutral detergent fiber (NDF) digestibility was higher in the SHCG pellets. The NDF content was higher in the SHCG diet. Individual cow behavior at the feeding lane was analyzed during the experimental period. Average number of meals and daily eating duration of the SHCG cows were significantly greater, as compared with the HST group. However, intake per meal and rate of eating were greater in the HST cows, whereas meal duration was similar in both groups. Feeding behavior resulted in significantly higher daily dry matter and NDF intake by the SHCG cows (27.1 and 11.1 kg, respectively) as compared with the HST group (24.8 and 7.61 kg, respectively). Consequently, significantly higher milk fat content, milk fat yield, and 4% FCM yield were obtained in the SHCG cows. Milk and milk protein yields were similar in both treatments. Data suggest potential advantages of the SHCG pellets for herds using automatic milking systems. PMID- 15290985 TI - The effects of forage provision and group size on the behavior of calves. AB - The effects of providing alternative forages to individual or group-reared calves on their behavior were examined in 2 experiments. In experiment 1, 24 calves were reared in groups of 3 or individually in straw-bedded pens from age 1 to 7 wk. One-half of the calves in each treatment were provided with ad libitum cut perennial ryegrass herbage. Grass intakes and time spent eating grass were greater for grouped calves than for individual calves. Providing grass reduced concentrate intake of grouped calves and reduced the time that all calves, but particularly individual calves, spent eating straw bedding. Ruminating time was increased by offering grass to grouped calves compared with individual calves. Grass reduced the frequency of calves licking their buckets and their pen, vocalizing, and investigating their pen. Particularly for grouped calves provision of grass reduced all grooming. Group rearing reduced the frequency of calves licking their bucket, vocalizing, and investigating their pen, but had no effect on the frequency of pen licking. Calves were weaned at wk 7 and transferred to indoor silage feeding or grazing. Most effects of group rearing and grass provision were not maintained after weaning, but calves that had received grass ate for longer periods when turned out to pasture. In experiment 2, 72 calves were offered a mixture of straw, molasses, and pot ale syrup or grass hay. Calves offered the straw mixture ate more forage and concentrates and grew faster than calves offered hay. It was concluded that nontraditional forages, such as fresh grass and straw mixtures, could benefit the behavior and growth of calves compared with hay and straw. PMID- 15290986 TI - Effect of corn particle size on site and extent of starch digestion in lactating dairy cows. AB - Two experiments were conducted to determine the effects of corn particle size (CPS) on site and extent of starch digestion in lactating dairy cows. Animals were fitted with ruminal, duodenal, and ileal cannulas. Dry corn grain accounted for 36% of dry matter intake. In experiment 1, 6 cows were used in a duplicate 3 x 3 Latin square design. Semiflint corn was used. Corn processing methods were grinding, medium rolling, and coarse rolling. The mean particle size of the processed corn was 730, 1807, and 3668 microm, respectively. Rumen digestibility of starch linearly decreased from 59% with ground corn to 36% with coarsely rolled corn. Similarly, small intestine digestibility linearly decreased with increased CPS, and consequently, the amount of starch digested in the small intestine was not affected by corn processing. In experiment 2, 4 cows were used in a 2 x 2 crossover design. Dent corn was used. Corn processing methods were grinding and coarse rolling. The mean particle size of the processed corn was 568 and 3458 microm, respectively. Rumen digestibility of starch decreased from 70% with ground corn to 54% with coarsely rolled corn. Small intestine digestibility of starch was not significantly affected by CPS, and the amount of starch digested in the small intestine tended to be greater for rolled than for ground corn. In both experiments, starch total tract digestibility decreased with increased CPS. In conclusion, CPS is an efficient tool to manipulate rumen degradability of cornstarch. In midlactation cows, the decrease in the amount of starch digested in the rumen between grinding and coarse rolling is partly compensated for by an increase in the amount of starch digested in the small intestine with dent genotype, but with semiflint genotype postruminal digestion is not increased and rumen escape starch is not utilized by the animal. PMID- 15290987 TI - Tissue and external insulation estimates and their effects on prediction of energy requirements and of heat stress. AB - Published data were used to develop improved equations to predict tissue insulation (TI) and external insulation (EI) and their effects on maintenance requirements of Holstein cattle. These are used to calculate lower critical temperature (LCT), energy cost of exposure to temperatures below LCT, and excess heat accumulating in the body at temperatures above LCT. The National Research Council classifies TI by age groups and body condition score; and in the EI equation air velocity effects are linear and coat insulation values are derived from beef animals in cold climates. These lead to low LCT values, which are not compatible with known effects of environment on the performance of Holsteins in warm climates. Equations were developed to present TI as a function of body weight, improving prediction of TI for animals of similar age but differing in body weight. An equation was developed to predict rate of decrease of TI at ambient temperatures above LCT. Nonlinear equations were developed that account for wind effects as boundary layer insulation effects dependent on body weight and air velocity. Published data were used to develop adjustments for hair coat effects on EI in Holstein cows. While by NRC equations, wind has negligible effects on heat loss, the recalculated effects of air velocity on heat loss were consistent with published effects of forced ventilation on the responses of the Holstein cow. The derived LCT was higher by 10 to 20 degrees C than that calculated by NRC (2001) and accounted for known Holstein performance in temperate and warm climates. These equations pointed to tentative significant effects of cold (-10 degrees C) on energy requirements (7 Mcal/d) further increased by 1 m/s wind (15 Mcal/d), even in high-producing cows. Needs for increased heat dissipation and estimating heat stress development at ambient temperatures above the LCT are predicted. These equations can be used to revise NRC equations for heat exchange. PMID- 15290988 TI - Effects of fiber content and particle size of forage on the flow of microbial amino acids from continuous culture fermenters. AB - Eight dual-flow continuous culture fermenters (1320 mL) were used in two 10-d consecutive periods to study the effects of fiber content and particle size on rumen fermentation, nutrient flow, and the profile and flow of amino acids (AA) from microbial origin. Treatments were arranged in a 2 x 2 factorial; the main factors were fiber content (high fiber [HF] = 67% alfalfa hay, 33% concentrate; low fiber [LF] = 39% alfalfa hay, 61% concentrate) and forage stem particle size (ground not to pass a 3-mm sieve = large size [LS]; ground to pass a 1-mm sieve = small size [SS]). On the last day of the experiment, liquid- (LAB) and solid- (SAB) associated bacteria were isolated from each fermenter for chemical analysis. Microbial N and AA flows were estimated using LAB or SAB composition data. Total volatile fatty acid concentration was higher for LF (122 mM) than for HF (102 mM). The proportion of acetate was higher for HF (63.5%) than for LF (58.2%). When SS was fed instead of LS, there was a reduction in the acetate proportion (63.4% vs 58.3%, respectively) and in the acetate to propionate ratio (3.13 vs. 2.41, respectively). Bacterial N flow was higher, and dietary N flow was lower, in SS compared with LS when LAB were used for calculations. Efficiency of microbial protein synthesis (EMPS) was affected by particle size, when LAB were used for calculations, and by fiber content, when SAB were used for calculations. The AA profile of LAB differed from SAB in 4 of 16 AA. The fiber content and particle size had small effects on microbial AA profile. However, the bacterial AA flow was higher in 13 of 16 AA, when SAB instead of LAB were used for calculations. Estimates of bacterial N flow, EMPS, AA profile, and flows of microbial origin were dependent on the type of microbial population used for calculations. The representativeness of the microbial samples is essential for the accurate prediction of the AA flow from microbial origin. PMID- 15290989 TI - Short communication: Effects of feeding level on energy concentration in grass silage-based diets offered to dairy cattle. AB - Twelve grass silages were offered to sheep as a sole diet at maintenance and to lactating dairy cows ad libitum as mixed silage and concentrates diets (n = 13 diets). Fecal and urinary energy outputs were measured for silages and mixed diets. Digestible energy (DE) and metabolizable energy (ME) concentrations for mixed diets with sheep at maintenance were estimated based on the silage dry matter (DM) proportion obtained in the cattle trials, the silage energy utilization values (methane energy-predicted) determined using sheep, and tabulated concentrate values. A comparison of dietary mean data (n = 13) indicated that concentrations of ME (P < 0.01) and DE (P < 0.001) in mixed diets were significantly lower for cows at production feeding level than for sheep at maintenance. The reductions were proportionately 0.015 and 0.020 with each unit increase in feeding level above maintenance, respectively. These ME and DE data were also used to evaluate the feeding level correction factors previously proposed by Van Es (1975) (ME, 0.018) and Yan et al. (2002) (ME, 0.016; DE, 0.025) using the mean square prediction error technique. The ME correction factor proposed by Yan et al. (2002) had a greater prediction accuracy than that proposed by Van Es (1975) for the prediction of ME concentration in mixed diets offered to dairy cattle at production feeding level. PMID- 15290990 TI - Short commuunication: Effect of dietary protein depletion and repletion on skeletal muscle calpastatin during early lactation. AB - Sixteen multiparous Jersey cows were assigned at calving to one of 4 dietary treatments. An 18% crude protein (CP) diet was fed as a total mixed ration through 30 d in milk (DIM), and beginning at 31 DIM a 9, 12, 15, or 18% CP diet was fed through 58 DIM (depletion). All cows were then fed the 18% CP diet until 84 DIM (repletion). Muscle biopsies were taken under local anesthesia at 49 and 84 DIM from the semitendinosus muscle. Milk production, DMI, and milk component contents were measured. Calpain and calpastatin contents of muscle taken at biopsy were evaluated using Western blotting techniques. Milk production and milk protein content were reduced during the depletion period by decreasing dietary protein. Diet had no effect on milk fat content or DMI. During repletion, DMI was affected by dietary treatment. Western blots of muscle extracts indicated no differences in calpain content at any stage of the experiment or in calpastatin content of muscle at 49 DIM. However, at 84 DIM, calpastatin (135 kDa) was lower or undetectable in cows fed either the 9 or 12% CP diets from 31 to 59 DIM. Bands for a 110-kDa degradation product of calpastatin were present in some cows fed the 9, 12, and 15% CP diets during the depletion period. Results indicate a change in skeletal muscle calpain/calpastatin proteolytic system during protein repletion following depletion with diets of less than 15% CP during early to peak lactation in dairy cows. PMID- 15290991 TI - Effect of feeding space on the inter-cow distance, aggression, and feeding behavior of free-stall housed lactating dairy cows. AB - The objectives of this study were to determine whether doubling the amount of feeding space from 0.5 to 1.0 m per cow leads to increased spacing between cows at the feeder, fewer aggressive social interactions among cows, and ultimately increased feeding activity. Twenty-four lactating Holstein cows were provided with 0.5 and 1.0 m of feeding space per cow in a 2 x 2 crossover design replicated over time. Time-lapse video was used to quantify the inter-cow distance and incidence of aggressive displacements at the feed alley. An electronic feed alley monitoring system was used to monitor the feeding behavior of the cows. When animals had access to 1.0 m per cow, there was at least 60% more space between animals and 57% fewer aggressive interactions while feeding than with access to 0.5 m of feeding space. These changes in spacing and aggressive behavior in turn allowed cows to increase feeding activity throughout the day, especially during the 90 min after providing fresh feed (an increase of 24%). This increase in feeding activity was particularly evident for subordinate cows. These results indicate that increasing space allowance at the feeder increases feeding activity and reduces competition among lactating dairy cows. PMID- 15290992 TI - Higher intake of DK265 corn silage by dairy cattle. AB - In this experiment, intake of DK265 3-way corn hybrid by dairy cattle was compared specifically with intake 1) of its bm3 isogenic form, 2) of its 2 related single-way hybrids, and 3) of 2 controls that were registered hybrids of similar earliness. Both dry matter (DM) and lignin contents were similar in all hybrids except for the bm3 hybrid, which was less lignified. There was a tendency for lower starch content and, correlatively, higher neutral detergent fiber content in DK265 and in the 2 related single-way hybrids. Significant intake differences were observed between hybrids; the highest intake was recorded for the bm3 hybrid. Among normal hybrids, DK265 and one of its related single-way hybrids registered significantly higher intakes than other hybrids. Among normal hybrids, cell wall digestibility and/or lignin content did not explain all of the variations observed for intake, whereas the higher intake of DK265 bm3 could be related to its lower lignin content as compared with isogenic DK265. It was hypothesized that the higher intake observed for the DK265 hybrid was probably related to specific friability traits that are not relevantly measured through the usual tests used in corn breeding. PMID- 15290993 TI - Digestible energy values of diets with different fat supplements when fed to lactating dairy cows. AB - A total collection digestion trial using high producing lactating cows (average milk yield = 40.7 kg/d) was conducted to measure the effect of different fat supplements on dietary digestible energy (DE) concentrations and to calculate the DE value of the supplements. A diet with no supplemental fat, 2 diets with 1.75 or 3.5% (dry basis) Ca salts of palm fatty acids (Ca-PFA), and 2 diets with 1.6 or 3.2% hydrogenated triacylglycerides from palm oil (HPO) were fed in a 5 x 5 Latin square experiment with 28-d periods. Concentrations of supplemental long chain fatty acids in the diets were 1.7 and 3.4% for the 2 supplementation rates. Dry matter intake was reduced when cows were fed the high concentration of Ca PFA, but cows fed Ca-PFA produced more milk than cows fed the control diet or diets with HPO. The type or amount of fat supplementation did not affect measures of rumen fermentation or in situ fiber digestibility. Digestibility of energy, dry matter, and organic matter were higher for diets with Ca-PFA than for diets with HPO, mainly because of increased fatty acid digestibility. The dietary concentration of DE was similar between the control diet and diets with HPO (2.97 Mcal/kg), but it increased as the concentration of Ca-PFA increased (3.04 and 3.16 Mcal/kg for low and high supplementation rates). The calculated DE concentrations of the supplements averaged 7.3 and 3.1 Mcal/kg for Ca-PFA and HPO. The 2001 National Research Council dairy model accurately estimated DE concentrations in all diets (<1% difference). PMID- 15290994 TI - Short communication: effect of carbohydrate fermentation rate on estimates of mass fermented and milk response. AB - Current prediction equations were used to evaluate the effects of rates of fermentation of fiber or starch in individual feeds on amounts of carbohydrate fermented ruminally and milk yield responses. The small predicted increases in carbohydrate fermented and milk response associated with doubling the rates of fermentation suggest that current prediction equations are relatively insensitive to changes in rate of fermentation. PMID- 15290995 TI - Genetic correlations among production, body size, udder, and productive life traits over time in Holsteins. AB - Genetic correlations among milk, fat, and protein yields; body size composite (BSC); udder composite (UDC); and productive life (PL) in Holsteins were investigated over time. The data set contained 25,280 records of cows born in Wisconsin between 1979 and 1993. The multiple trait random regression (MT-RR) animal model included registration status, herd-year, age group, and stage of lactation as fixed effects; additive genetic effects with random regressions (RR) on year of birth using the first-order Legendre polynomial; and residual effects. Heterogeneous residual variances were considered in the model. Estimates of variance components and genetic correlations among traits from MT-RR were compared with those estimated with a multiple trait interval (MT-I) model, which assumed that every 3-yr interval was a separate trait and included the same effects as in the MT-RR model except for the RR. Genetic correlations estimated with MT-RR and MT-I models over time among all traits were compared with correlations among breeding values predicted with the single trait (ST) model without RR. Correlations among breeding values predicted with MT-RR, ST, and MT models were also calculated. Additive genetic and residual variances for all traits except PL increased over time; those for PL were constant. As a result, heritability estimates had no significant changes during the 15 yr. Genetic correlations of PL with milk, fat, protein, and BSC declined to zero or negative; those with UDC remained positive. Correlations among breeding values predicted with ST, MT, and MT-RR models were relatively high for all traits except PL. Genetic correlations between PL and other traits varied over time, with some correlations changing sign. For accurate indirect prediction of PL from other traits, the genetic correlations among the traits need to be re-estimated periodically. PMID- 15290996 TI - Comparison between a Weibull proportional hazards model and a linear model for predicting the genetic merit of US Jersey sires for daughter longevity. AB - Predicted transmitting abilities (PTA) of US Jersey sires for daughter longevity were calculated using a Weibull proportional hazards sire model and compared with predictions from a conventional linear animal model. Culling data from 268,008 Jersey cows with first calving from 1981 to 2000 were used. The proportional hazards model included time-dependent effects of herd-year-season contemporary group and parity by stage of lactation interaction, as well as time-independent effects of sire and age at first calving. Sire variances and parameters of the Weibull distribution were estimated, providing heritability estimates of 4.7% on the log scale and 18.0% on the original scale. The PTA of each sire was expressed as the expected risk of culling relative to daughters of an average sire. Risk ratios (RR) ranged from 0.7 to 1.3, indicating that the risk of culling for daughters of the best sires was 30% lower than for daughters of average sires and nearly 50% lower than than for daughters of the poorest sires. Sire PTA from the proportional hazards model were compared with PTA from a linear model similar to that used for routine national genetic evaluation of length of productive life (PL) using cross-validation in independent samples of herds. Models were compared using logistic regression of daughters' stayability to second, third, fourth, or fifth lactation on their sires' PTA values, with alternative approaches for weighting the contribution of each sire. Models were also compared using logistic regression of daughters' stayability to 36, 48, 60, 72, and 84 mo of life. The proportional hazards model generally yielded more accurate predictions according to these criteria, but differences in predictive ability between methods were smaller when using a Kullback-Leibler distance than with other approaches. Results of this study suggest that survival analysis methodology may provide more accurate predictions of genetic merit for longevity than conventional linear models. PMID- 15290997 TI - Undesired phenotypic and genetic trend for stillbirth in Danish Holsteins. AB - The primary aim of this study was to evaluate the phenotypic and genetic trends for stillbirth in Danish Holsteins. Trends of calving difficulty and calf size were also evaluated. The second aim was to compare predicted transmitting abilities (PTA) of sires for stillbirth using a linear and a threshold model. Direct and maternal genetic effects were modeled by fitting correlated additive genetic effects of the sire and the maternal grandsire (MGS). For both the calf and the dam, covariates of breed proportions of Holstein-Friesian (HF) and the heterozygosity between HF and the original Danish Black and White (ODBW) were included. Records from 1.8 million first-calving Danish Holstein cows calving from 1985 to 2002 were used. In this period, the overall frequency of stillbirth increased from 0.071 to 0.090. An unfavorable genetic trend of stillbirth was found for both the direct and maternal effect. The background for the genetic trends was an intense use of HF sires as sires of sons, which increased the proportion of HF genes to 94% in the Danish Holstein calves born in 2002. The effect of the imported HF genes was higher direct effects of calf size, calving difficulty, and stillbirth compared with the ODBW genes. The maternal effect of stillbirth was poorer for HF than for ODBW even though HF had a better maternal calving performance than ODBW. The threshold and the linear models showed almost similar predictions of transmitting abilities of sires. PMID- 15290998 TI - Genotype x environment interaction for yield and somatic cell score with automatic and conventional milking systems. AB - The objective of this study was to quantify genotype by environment interaction (G x E) between automatic milking systems (AMS) and conventional milking systems (CMS) for test-day milk, fat, and protein yield and for test-day somatic cell score (SCS) in The Netherlands. The G x E was studied in 2 ways: 1) between AMS farms and CMS farms in the same period and 2) within farms comparing the period before introduction of AMS with the period after introduction of AMS. For both sub-objectives, a separate data set was generated. Test-day records were used to be more flexible with respect to the introduction date of AMS. Multivariate, fixed regression, test-day sire models were used to estimate variance components. Genetic correlations between AMS farms and CMS farms in the same period were 0.93, >0.99, 0.98, and 0.79 for test-day milk yield, fat yield, protein yield, and SCS, respectively. Genetic correlations within farms between the period before and after introduction of AMS were lower for production traits and higher for SCS: 0.89, 0.91, 0.87, and >0.99, respectively, for test-day milk yield, fat yield, protein yield, and SCS. Heterogeneity of variance was observed between AMS and CMS in both data sets. Especially the residual variance increased with automatic milking. As a consequence, the heritability tended to be lower for automatic milking. It was concluded that effects of G x E are small between AMS and CMS. Therefore, AMS farms can select sires accurately based on national rankings. PMID- 15290999 TI - Genetic comparison of breeding schemes based on semen importation and local breeding schemes: framework and application to Costa Rica. AB - Local breeding schemes for Holstein cattle of Costa Rica were compared with the current practice based on continuous semen importation (SI) by deterministic simulation. Comparison was made on the basis of genetic response and correlation between breeding goals. A local breeding goal was defined on the basis of prevailing production circumstances and compared against a typical breeding goal for an exporting country. Differences in genetic response were <3%, and the correlation between breeding goals was 0.99. Therefore, difference between breeding objectives proved negligible. For the evaluation of genetic response, the current scheme based on SI was evaluated against a progeny testing (PT) scheme and a closed nucleus (CN) breeding scheme, both local. Selection intensities and accuracy of selection were defined according to current population size and reproduction efficiency parameters. When genotype x environment interaction (G x E) was ignored, SI was the strategy with the highest genetic response: 5.0% above the CN breeding scheme and 33.2% above PT. A correlation between breeding values in both countries lower than one was assumed to assess the effect of G x E. This resulted in permanent effects on the relative efficiencies of breeding strategies because of the reduction in the rate of genetic response when SI was used. When the genetic correlation was assumed equal to 0.75, the genetic response achieved with SI was reduced at the same level as local PT. When an initial difference in average genetic merit of the populations was assumed, this only had a temporal effect on the relative ranking of strategies, which is reverted after some years of selection because the rate of change in genetic responses remains unchanged. Given that the actual levels of genetic correlation between countries may be around 0.60, it was concluded that a local breeding scheme based on a nucleus herd could provide better results than the current strategy based on SI. PMID- 15291000 TI - Genetic parameters and trends in the Chilean multibreed dairy cattle population. AB - Estimates of additive and nonadditive multibreed co-variance components, genetic parameters, and predicted genetic values for first lactation 305-d mature equivalent (ME) milk yield, fat yield, and protein yield were computed using data from a sample of 3316 cows from the Chilean Holstein-other breeds multibreed population. Variances and covariances were estimated by 2-trait REML analyses using a Generalized Expectation-Maximization algorithm applied to multibreed populations. Multiple estimates of additive genetic, nonadditive genetic, and environmental variances from 2-trait analyses were averaged to yield a single variance estimate for each trait and effect. Heritabilities were moderate for all traits in Holstein, other, and Holstein x other crossbred groups. Interbreed interactibilities (ratio of nonadditive genetic to phenotypic variances) were all near zero. Multibreed additive, nonadditive, and total genetic trends were estimated using the complete dataset (56,277 cows). Upward trends between 1990 and 2000 existed for all traits, genetic effects, and breed groups, except for 305-d ME protein yield in 1/4 Holstein, indicating that Chilean dairy producers were successful in choosing progressively better semen and sires from imported and local sources over time. PMID- 15291001 TI - Genetic analysis of the Israeli Holstein dairy cattle population for production and nonproduction traits with a multitrait animal model. AB - Milk, fat, and protein production, somatic cell score (SCS), and female fertility in the Israeli Holstein dairy cattle population were analyzed using a multitrait animal model (AM) with parities 1 through 5 as separate traits. Female fertility was measured as the inverse of the number of inseminations to conception in percent. Variance components were estimated using both the repeatability AM and multitrait AM. The multitrait heritabilities for individual parities were greater than the heritabilities from the repeatability AM, and heritabilities decreased with an increase in parity number. Heritabilities were higher for production traits, lower for SCS, and lowest for female fertility. The genetic correlations were higher than the environmental correlations. Genetic correlations between parities decreased with an increase in the difference in parity number, but all were greater than 0.5. The environmental correlations were higher for production traits, lower for SCS, and close to zero for female fertility. In the analysis of the complete milk recorded population, genetic trends from the repeatability and multitrait models were very similar. The genetic trend for SCS was economically unfavorable until 1993, and favorable since then. The genetic trend for female fertility was close to zero, but the annual environmental trend was -0.2%. The multitrait lactation model is an attractive compromise between repeatability lactation models, which do not account for maturing trends across parities, and test-day models, which are much more demanding computationally. PMID- 15291002 TI - Efficiency of different selection criteria for persistency and lactation milk yield. AB - A conversion formula was developed to convert the genetic covariance matrices of daily yields and of random regression coefficients between 305-d and 335-d production periods under a random regression test day model. Five selection criteria were compared in terms of genetic improvement in persistency and lactation milk: 1) lactation estimated breeding value (EBVL), 2) P6 = 279Sigma(i=65) (D280 - Di), 3) ratio of daily estimated breeding value (EBV)(r280/65 = D280/D65), 4) ratio of partial lactation EBV (P280/65 = D66 approximately 28/D5 approximately 65), and 5) differential daily EBV (d65-280 = D65 - D280), where Di refers to EBV at days in milk (DIM) i. Fundamental differences among these 5 selection criteria were interpreted conceptually with a graph. Persistency, defined as k = (delta G65 - delta G280)/215, was the average daily rate of decline in selection gain from DIM 65 to 280, which is free from the effect of lactation milk on the rate of decline. Parameter k provides an objective measure of persistency, which increases when k < 0 and decreases when k > 0. Of the 5 selection criteria compared, d65-280 and P6 achieved greater persistency at the expense of genetic gain in lactation milk, whereas selection based on EBVL achieved the highest response in lactation milk, but was coupled with greatest decline in persistency. Selection on P280/65 or r280/65 improved both lactation milk and persistency and, thus, is recommended for simultaneous improvement of these 2 economically important traits. Further study of the relative economic values of persistency and lactation milk in order to combine both traits into an index for selection decision is warranted. PMID- 15291003 TI - The color of Brevibacterium linens depends on the yeast used for cheese deacidification. AB - The color of smear cheeses (Muenster) is traditionally thought to be due to the bacterial flora, e.g., Brevibacterium linens. This study was carried out to evaluate indirect effects of yeast on the color of B. linens. A 60% cheese medium was desacidified with Debaryomyces hansenii or Kluyveromyces marxianus until pH 5.8 was reached. After inactivation of the yeast and addition of agar-NaCl, B. linens was inoculated on the medium surface and incubated at 12 degrees C from d 2 to 28. For each bacterial biofilm, color was evaluated by L*C*h(degrees) (brightness, chroma, hue angle) spectrocolorimetry. After d 14 (D. hansenii deacidification) and d 21 (K marxianus desacidification), the color level (as a function of all 3 factors) of B. linens biofilms became maximal and remained so until d 28. Debaryomyces hansenii 304 (LGMPA) was less efficient for deacidification than K. marxianus Laf5. However, color intensity (function of chroma only) was higher when D. hansenii was used. The yeast used had an effect on the composition of the cheese medium in relation to production and consumption of metabolites during deacidification. The results concerning color are discussed with respect to this cheese medium composition. PMID- 15291004 TI - Comparison of volatile compounds produced in model cheese medium deacidified by Debaryomyces hansenii or Kluyveromyces marxianus. AB - The aroma of a deacidified cheese medium is the result of the overall perception of a large number of molecules belonging to different classes. The volatile compound composition of (60%) cheese medium (pH 5.8) deacidified by Debaryomyces hansenii (DCM(Dh)) was compared with the one deacidified by Kluyveromyces marxianus (DCM(Km)). It was determined by dynamic headspace extraction, followed by gas chromatography separation and quantification as well as by mass spectrometry identification. Whatever the media tested, a first class of volatile compounds can be represented by the ones not produced by any of the yeasts, but some of them are affected by K. marxianus or by D. hansenii. A second class of volatile compounds can be represented by the ones produced by K. marxianus, which were essentially esters. Their concentrations were generally higher than their thresholds, explaining the DCM(Km) global fruity odor. A third class can be represented by the ones generated by D. hansenii, which were essentially methyl ketones with fruity, floral (rose), moldy, cheesy, or wine odor plus 2 phenylethanol with a faded-rose odor. The impact of methyl ketones on the DCMDh global flavor was lower than the impact of 2-phenylethanol and even negligible. Therefore, the global faded-rose odor of D. hansenii DCM can be explained by a high concentration of 2-phenylethanol. PMID- 15291005 TI - A modified presynchronization protocol improves fertility to timed artificial insemination in lactating dairy cows. AB - To compare 2 hormonal protocols for submission of lactating dairy cows for timed artificial insemination (TAI), nonpregnant lactating Holstein cows (n = 269) >60 d in milk were randomly assigned to each of 2 treatments to receive TAI (TAI = d 0). Cows assigned to the first treatment (Ovsynch, n = 134) received 50 microg of GnRH (d -10), 25 mg of PGF2alpha (d -3), and 50 microg of GnRH (d -1) beginning at a random stage of the estrous cycle. Cows assigned to the second treatment (Presynch, n = 135) received Ovsynch but with the addition of 2 PGF2alpha (25 mg) injections administered 14 d apart beginning 28 d (d -38 and -24) before initiation of Ovsynch. All cows received TAI 16 to 18 h after the second GnRH injection. Ovulatory response after each GnRH injection for a subset of cows (n = 109) and pregnancy status 42 d after TAI for all cows were assessed using transrectal ultrasonography. Based on serum progesterone (P4) profiles determined for a subset of cows (n = 109), P4 concentrations decreased for Presynch cows after the first 2 PGF2alpha injections, and Presynch cows had greater P4 concentrations at the PGF2alpha injection on d -3 compared with Ovsynch cows. Although the proportion of cows ovulating after the first and second GnRH injections did not differ statistically between treatments (41.1 and 69.6% vs. 35.9 and 81.1% for Ovsynch vs. Presynch, respectively), pregnancy rate per artificial insemination (PR/AI) at 42 d post TAI was greater for Presynch than for Ovsynch cows (49.6 vs. 37.3%). Parity, DIM, and body condition score (BCS) at TAI did not affect PR/AI to TAI. These data support use of this presynchronization protocol to increase PR/ AI of lactating dairy cows receiving TAI compared with Ovsynch. PMID- 15291006 TI - On-farm management decisions to improve beef quality of market dairy cows. AB - A 3-phase study was conducted to assess on-farm management decisions to reduce antibiotic residue violations and improve carcass characteristics in market (cull) dairy cows. In Phase 1, questionnaires were mailed to dairy producers (n = 142) to determine current on-farm management strategies for reducing antibiotic residues in market dairy cattle. In Phase 2, Holstein market cows (n = 77) were assigned randomly to each of the 3 feeding treatments (0, 30, or 60 d). Average daily gain (ADG), body condition score (BCS), and carcass characteristics were assessed. Phase 3 determined the meat withdrawal time of Holstein cows (n = 62) administered procaine penicillin G. Eighty-six percent of dairy farms responding to the questionnaire had at least one cow condemned annually, and no producer had a designated feeding protocol for market cows prior to selling. In Phase 2, ADG was greater in cows fed for 30 d (1.4+/-0.6 kg/d) than in cows fed for 60 d (0.9+/-0.4 kg/d). Additional feeding did not influence the carcass characteristics studied with the exception of kidney, pelvic and heart fat, which was higher in cows fed for 60 d compared with those fed for 0 and 30 d. In Phase 3, 31% of cows treated with procaine penicillin G exceeded the 10-d label withdrawal recommendation by an average of 3.1+/-1.9 d. Feeding market cows may not influence carcass characteristics, but can increase ADG and may ensure that recommended meat withdrawal times for antibiotics are exceeded. PMID- 15291007 TI - An economic evaluation of recombinant bovine somatotropin approval in Japan. AB - A comprehensive econometric model was developed to evaluate potential impacts of recombinant bovine somatotropin (rbST) approval in Japan. Three novel features of the analyses include 1) investigation of impact of rbST on herd-size structure, 2) determination of economic feasibilities of rbST adoption by herd size, and 3) evaluation of policies to mitigate negative effects of rbST approval. Simulation analysis was conducted over a 10-yr projected period assuming rbST was approved in Japan in 2001. Nine hypothetical scenarios were simulated to examine sensitivity of simulation results. Simulation results indicate that rbST approval would accelerate structural change in Japan's dairy industry toward fewer, larger farms. Negative effects of rbST on farm income are projected to be more severe for smaller farms, because of higher costs, lower profit-earning ability, lower milk yields, and lower adoption rates of rbST. Larger farms benefit from rbST adoption if milk demand is maintained. However, if concerns about rbST induce significant milk demand decreases, even the largest farms' income and cow numbers will decrease. Thus, Japan's dairy industry could be caught in a double downward spiral of declining milk prices and production. Assuming rbST is approved, small farms would benefit by using the technology, but they fare best if rbST is not approved. Two policies could be effective in mitigating possible farm income losses. First, lost farm income can be offset if dairy cooperatives can exercise greater market power to control fluid milk marketings. Second, because generic milk advertising has positive effects on both milk demand and farm income, increasing check-off rates to fund more advertising could ease farm income losses. PMID- 15291008 TI - Sociodemographic profile of insomniac patients across national surveys. AB - INTRODUCTION: A survey of 2121 insomniac patients was conducted in France, Germany, Italy and the United Kingdom. DESIGN: The survey collected data on the sociodemographic and clinical features of the subjects by interview, as well as on healthcare resource utilization. A complementary analysis of physician attitudes to insomnia provided insights into the types of treatment prescribed and expectations of medical treatment. RESULTS: Slightly more women than men reported insomnia, but the difference was not significant. The age distribution of insomniac patients varied from country to country. Insomnia was more frequent in the unemployed and in those individuals living alone. The most frequently cited causes of insomnia were stress, loneliness and bereavement. Insomnia was usually reported as chronic, and frequently as episodic. Frequently cited symptoms were poor sleep quality, interrupted sleep, early awakening, difficulties in getting to sleep and daytime fatigue. CONCLUSIONS: Many individuals hesitate to consult their doctor about insomnia, whereas physicians question their patients about possible sleep problems relatively infrequently and systematically underestimate the severity of insomnia and the resulting functional impairment. Physicians frequently recommended lifestyle changes and sleep hygiene measures to patients complaining of insomnia. Hypnotic agents were the most frequently prescribed class of medication. Patient satisfaction with treatment was relatively high. PMID- 15291009 TI - The pharmacology and mechanisms of action of new generation, non-benzodiazepine hypnotic agents. AB - The new generation hypnotic drugs, zolpidem, zopiclone and zaleplon, are at least as efficacious in the clinic as benzodiazepines and may offer advantages in terms of safety. These drugs act through the BZ binding sites associated with GABAA receptors, but show some differences from benzodiazepines in pharmacological effects and mechanisms of action. Of particular interest is the finding that zolpidem shows a wide separation between doses producing sedative effects and those giving rise to other behavioural actions, and induces less tolerance and dependence than benzodiazepines. Zolpidem also demonstrates selectivity for GABAA receptors containing alpha1 subunits. Recent studies using genetically modified mice have confirmed that receptors containing alpha1 subunits play a particularly important role in mediating sedative activity, thus providing an explanation for the pharmacological profile of zolpidem. PMID- 15291010 TI - Overview of the therapeutic management of insomnia with zolpidem. AB - Benzodiazepine hypnotic agents were the mainstream pharmacotherapy for insomnia from the 1960s to the 1980s, but their safety profile proved to be not quite as perfect as originally expected with regard to daytime performance and cognition, and above all the risk of dependence. These risks are substantially diminished in the non-benzodiazepine hypnotic agents developed and marketed during the past two decades, but the fears engendered by certain benzodiazepines still greatly influence the attitude of both physicians and the general public to the treatment of insomnia. For this reason, as well as in the interests of matching the pharmacotherapy of insomnia more closely to the often fluctuating nature of this disorder, the possibility of the discontinuous or 'as needed' use of hypnotic drugs has attracted increasing attention in recent years. Current recommendations strongly favour the use of hypnotic drugs for a limited period of time. However, some insomniac patients need sleep medication for longer periods in spite of a non-pharmacological approach, whereas other patients become dependent on drugs as a result of rebound insomnia, withdrawal symptoms, or the recurrence of insomnia. The pharmacological properties of zolpidem make it feasible for non-nightly use. A double-blind, randomized, parallel-group study of continuous treatment with either zolpidem or estazolam, followed by an observation of the discontinuation of drug treatments combined with the non-pharmacological management of primary insomnia, showed a carry-over benefit for zolpidem treatment. PMID- 15291011 TI - Zolpidem 'as needed': methodological issues and clinical findings. AB - This article introduces 'as needed' hypnotic treatment as a possible option in the psychopharmacological treatment for chronic but non-daily occurring insomnia complaints. This treatment option is a well-structured regimen designed by both patient and treating physician. It aims at providing symptomatic relief for sleepless nights and preventing the development of possible dependence on the hypnotic agent. A significant amount of responsibility regarding the necessary accompanying lifestyle changes remains in the patient's hands. This paper describes three of the early experiments designed as a 'proof of concept'. All three studies demonstrated that zolpidem can be used effectively and safely on a non-daily basis in individuals with chronic insomnia. Although these studies have methodological limitations, they opened doors to well-designed clinical trials in both in and outpatients. The scientific validation of this therapeutic approach is still in progress. In addition to sleep laboratory studies, ecological studies are needed to investigate pill-taking behaviour and the risk of dependence over a long period of time in real life settings. PMID- 15291012 TI - Experience with zolpidem 'as needed' in primary care settings. AB - 'As needed' non-nightly intake of hypnotic agents by patients suffering from chronic insomnia is likely to offer benefits such as maintained efficacy while preventing unnecessary long-term nightly use associated with the risk of tolerance and dependence. To date, three studies have proven the 'as needed' approach with zolpidem in patients with primary insomnia treated in outpatient practice. In total, more than 3000 primary care patients, proved that flexible or semi-flexible schedules of zolpidem 'as needed' treatment are feasible. In all the studies, 'as needed' zolpidem treatment was both effective and well tolerated by the patients. Moreover, the results of these studies clearly indicated that chronic insomniac patients are capable of limiting their hypnotic intake, showing no tendency to increase intake over time and even a trend to decreasing use of medication. PMID- 15291013 TI - Selection of instrumentation and fusion levels for scoliosis: where to start and where to stop. Invited submission from the Joint Section Meeting on Disorders of the Spine and Peripheral Nerves, March 2004. AB - OBJECT: Although there are several papers in the literature regarding selection of fusion levels in the adolescent patient, fewer articles pertain to this in the adult patient. The author reviewed his experience and the literature and reports on the choice of fusion levels in the adolescent and adult patient. METHODS: After a review of available data, the author determined that the proximal and distal extent of the fusion should be based on defining curves as either major or minor in the adolescent patient. It is often possible to exclude minor curves from the fusion. Relative Cobb measurement, apical deviation from the plumb line, and apical rotation are the most useful means of distinguishing a major from a minor curve. Otherwise, the proximal and distal extent of a fusion should be performed in such a way that the proximal and distal vertebrae are both neutral and stable (bisected by the center sacral line) postoperatively. Additional segments may need to be included in the adult patient in whom extensive degenerative changes and subluxations are present. The decision of whether to terminate a long fusion at L-5 or the sacrum in an adult degenerative lumbar curve is complex and many factors have to be considered. CONCLUSIONS: Guidelines exist for fusion levels in both adolescent and adult patients. Not all curves require fusion. There are many coronal and sagittal considerations that have to be analyzed when making the final decision. PMID- 15291014 TI - Evaluation and treatment of adult spinal deformity. Invited submission from the Joint Section Meeting on Disorders of the Spine and Peripheral Nerves, March 2004. AB - Spinal deformity is a complex and dynamic process that occurs in both the sagittal and coronal planes of the thoracolumbar spine. Successful treatment is aimed at achieving satisfactory balance in both of these planes. The spinal curvatures in the adult differ greatly from those in adolescents. As a general rule the adult curves tend to be stiffer, whereas adolescent curves are more flexible. In addition to cosmetic concerns, adult patients frequently present with pain and neurological symptoms in contrast to adolescents who usually do not experience this degree of pain or neurological symptoms. The treatment of adult spinal deformity differs substantially from that of adolescent deformity. Surgeries in the former tend to be more complex procedures associated with higher rates of intra- and perioperative complications. The goals of surgery in the adult are to obtain a solid fusion with a balanced spine, to relieve pain, and to prevent further deformity. A secondary goal is to correct the curve, and, in so doing, to improve the cosmetic appearance. In this review the author addresses the basic principles of spinal corrective surgery in the adult and provides insight into the varied treatment options available. PMID- 15291015 TI - Contribution of recombinant human bone morphogenetic protein-2 to the rapid creation of interbody fusion when used in transforaminal lumbar interbody fusion: a preliminary report. Invited submission from the Joint Section Meeting on Disorders of the Spine and Peripheral Nerves, March 2004. AB - OBJECT: The authors compared fusion rates in transforaminal lumbar interbody fusion (TLIFs) when using either autograft or bone morphogenetic protein (BMP) placed in the interbody space. METHODS: Between September 2002 and December 2003, the authors performed 44 TLIF operations. Follow-up data were available for 40 patients. Of the 40 procedures, 19 involved cages filled with iliac crest autograft (Group 1) and 21 involved cages filled with a medium kit of recombinant human (rh) BMP-2 (Group 2). In all Group 2 patients, one BMP sponge was placed anterior to the cage and another was placed within the cage. In 12 of the Group 2 patients, iliac crest autograft was placed posterior to the BMP-filled cage (Group 2A). In the remaining nine Group 2 patients, only local autograft was placed posterior to the BMP-filled cage (Group 2B). Assessment of fusion was performed using dynamic radiography at 3-month intervals. Outcomes were assessed using the Prolo Scale, and iliac crest donor site pain was measured using a Visual Analog Scale (VAS). The mean follow-up period was 9 months (range 3-18 months). In Group 1 patients, one pseudarthrosis was detected. In Group 2 patients, dynamic radiography demonstrated solid fusion in all patients except one in Group 2B. Fifty-eight percent of patients in whom iliac crest autograft was used complained of donor site pain 6 months after surgery (5 of 10 points on the VAS). Symptomatic foraminal bone formation was not observed in any Group 2 patient. CONCLUSIONS: The use of rhBMP-2 is safe in TLIFs when the sponges are placed away from the dura mater, and BMP promotes a more rapid fusion than iliac crest autograft alone. The use of rhBMP-2 in combination with local autograft is an excellent option for promoting solid fusion with TLIF, and it eliminates the possibility of iliac donor site pain. PMID- 15291016 TI - Lumbar interbody fusion: state-of-the-art technical advances. Invited submission from the Joint Section Meeting on Disorders of the Spine and Peripheral Nerves, March 2004. AB - During the past few decades, three techniques have been used to achieve circumferential lumbar interbody fusion (LIF). They include posterior LIF, anterior LIF with supplemental posterior fixation, and transforaminal LIF. In this article, the authors describe the indications and contraindications for the use of interbody fusion. The advantages and disadvantages of each will be discussed in detail. Additionally, strategies for minimally invasive access and options for interbody spacer materials will be discussed. PMID- 15291017 TI - Lumbar stenosis: a personal record. Invited submission from the Joint Section Meeting on Disorders of the Spine and Peripheral Nerves, March 2004. AB - Although its management continues to evolve, lumbar stenosis remains a common societal problem. The present article is based on an invited lecture at the 2004 Annual Meeting of the Congress of Neurological Surgeons/American Association of Neurological Surgeons Joint Section on Disorders of the Spine and Peripheral Nerves. In it the authors provide a historical overview of lumbar stenosis and describe how the senior author's treatment of this condition has evolved over the past four decades. Within each era of treatment, the reasons for modification of treatment methods and relevant outcome measures are outlined. Additionally, specific subsets of patients with lumbar stenosis are also discussed to emphasize unique characteristics that affect treatment strategies. The authors' present technique for management of lumbar stenosis is also illustrated. PMID- 15291018 TI - Cervical magnetic resonance imaging abnormalities not predictive of cervical spine instability in traumatically injured patients. Invited submission from the Joint Section Meeting on Disorders of the Spine and Peripheral Nerves, March 2004. AB - OBJECT: Identifying instability of the cervical spine can be difficult in traumatically injured patients. The goal of this study was to determine whether cervical abnormalities demonstrated on magnetic resonance (MR) imaging are predictive of spinal instability. METHODS: Data in all patients admitted through the Level I trauma service at the authors' institution who had undergone cervical MR imaging were retrospectively reviewed. The reasons for MR imaging screening were neurological deficit, fracture, neck pain, and indeterminate clinical examination (for example, coma). Abnormal soft-tissue (prevertebral or paraspinal) findings on MR imaging were correlated with those revealed on computerized tomography (CT) scanning and plain and dynamic radiography to determine the presence/absence of cervical instability. Of 6328 patients admitted through the trauma service, 314 underwent MR imaging of the cervical spine. Of 166 patients in whom CT scanning or radiography demonstrated normal findings, 70 had undergone MR imaging that revealed abnormal findings. Of these 70 patients, 23 underwent dynamic imaging, the findings of which were normal. In each case of cervical instability (65 patients) CT, radiographic, and MR imaging studies demonstrated abnormalities. Furthermore, there were 143 patients with abnormal CT or radiographic study findings, in 13 of whom MR imaging revealed normal findings. Six of the latter underwent dynamic testing, which demonstrated normal results. CONCLUSIONS: Magnetic resonance imaging is sensitive to soft-tissue injuries of the cervical spine. When CT scanning and radiography detect no fractures or signs of instability, MR imaging does not help in determining cervical stability and may lead to unnecessary testing when not otherwise indicated. PMID- 15291019 TI - The impact of minimally invasive cervical spine surgery. Invited submission from the Joint Section Meeting on Disorders of the Spine and Peripheral Nerves, March 2004. AB - Since 1997, cervical endoscopic laminoforaminotomy (CELF) has been an effective and safe treatment option for unilateral cervical radiculopathy secondary to disc herniation or foraminal stenosis. The development of the surgical technique is reviewed and recent outcomes discussed. Its impact is addressed in relation to the patient and surgeon. PMID- 15291020 TI - Fusion rate: a time-to-event phenomenon. AB - OBJECT: The term "fusion rate" is generally denoted in the literature as the percentage of patients with successful fusion over a specific range of follow up. Because the time to fusion is a time-to-event phenomenon a more accurate method of representation may be made using the Kaplan-Meier method of estimation. METHODS: The current study was performed to illustrate that fusion rate is more accurately represented by median times as calculated using survival analysis. Patients undergoing a cervical decompressive corpectomy and reconstruction formed the basis of the primary analysis. A secondary analysis was made to evaluate the difference in the fusion times for one- compared with multilevel corpectomy cases. Data were collected at a tertiary care institution over a 5-year period with 6-month follow up after the last recruitment. Descriptive statistics of baseline patient characteristics, the extent of disease, and the surgical intervention were obtained. Fusion was the final outcome, and it was defined as the "event." The presence of any trabeculae bridging between the vertebral body and allograft signified the occurrence of an event. Postoperative static radiographs were evaluated by independent neuroradiologists to assess the presence of fusion. Fusion rate was determined using the Kaplan-Meier estimate. The median time to fusion was calculated, as were the 95% confidence intervals (CIs). These were stratified for patients who underwent one- and two-level vertebrectomy. The log-rank test was used to differentiate between one-level and multilevel corpectomy. Multivariate analysis was performed using Cox regression for further evaluation, by adjusting for covariates (age, sex, smoking history). Fifty-seven patients underwent single- or multilevel corpectomy and fusion. The male/female ratio was similar, with a median age of 53 years. Fourteen patients had a history of cigarette smoking. Thirty-six patients underwent a one-level corpectomy, 20 a two-level corpectomy, and one patient underwent a three-level corpectomy. The analysis was restricted to one- and two-level cases. The median time to fusion for the cephalad and caudad aspect of the graft-host interface was 88 days (95% CI 82-94 days) and 85 days (95% CI 77-93 days), respectively. As generally reported in the literature, this translates to a 92% (by 2.1 years) and 93% (by 1.5 years) fusion rate, for the cephalad and caudad, respectively. The median time to fusion for the cephalad aspect of the graft for one-level vertebrectomy was 87 days (95% CI 83-91 days), whereas for two-level vertebrectomy was 90 days (95% CI 59-121 days). The median time to fusion for the caudal aspect of the graft-host interface was 85 days (95% CI 80-90 days) for one level corpectomy and 90 days (95% CI 83-97 days) for the two-level cases. There was no statistically significant difference in the median time to fusion for one- and two-level corpectomy at either the superior or inferior aspect of the graft (p = 0.19 and 0.84, respectively). This held true even after adjusting for covariates. CONCLUSIONS: Fusion rate is a time-to-event phenomenon and is more accurately represented using the Kaplan-Meier method of estimation. PMID- 15291021 TI - Synovial cysts of the thoracic spine. AB - OBJECT: Thoracic synovial cysts (TSCs) are rare and are usually the subject of case reports. The authors studied the clinical manifestations, radiological aspects, and surgical treatment in a series of patients at their institution who harbored TSCs. They also review the literature to discuss the potential factors involved in the pathogenesis of this lesion. METHODS: A database search of 16,000 patients who underwent decompressive spine surgery at the Mayo Clinic (Rochester, MN) between 1976 and 2003 disclosed nine patients (0.06%) in whom a diagnosis of TSC had been made. All patients were men. The mean age at presentation was 73 +/- 5 years and mean duration of symptoms was 5 +/- 3 months. The mean duration of follow up was 4 +/- 3 years. The patients had no history of trauma or spine surgery. All patients had spastic paraparesis; two had urinary difficulties. Detailed neurological examination revealed myelopathy and radiculopathy with a sensory level of T10-L4. Magnetic resonance imaging revealed bilateral cysts in four patients and unilateral lesions in five. Three of the cysts were at the T-10 interspace, seven at the T-11 interspace, and three at the T-12 interspace. Seven cysts were on the right and six were on the left. Computerized tomography myelography performed in five patients revealed a gas bubble in the TSC in two patients. All patients underwent laminectomy/partial facetectomy, excision of the cyst, and decompression of the thecal sac and nerve root without any complications. None of these patients underwent a fusion. Eight patients (89%) experienced moderate to excellent relief of their preoperative signs and symptoms and one patient (11%) remained stable. There was no evidence of cyst recurrence at the site of surgery or other spinal segments at follow-up examination in any patient. CONCLUSIONS: When compared with their lumbar and cervical spine counterparts, TSCs are exceedingly rare. Their rarity may be explained by the decreased mobility of the thoracic spinal segments. The origin of TSCs is more likely degenerative rather than traumatic. Based on their experience and the follow-up duration, surgery provided durable relief from symptoms. PMID- 15291022 TI - Thoracic microendoscopic discectomy. AB - OBJECT: Various approaches exist for the treatment of thoracic disc herniation. Anterior approaches facilitate ventral exposure but place the intrathoracic contents at risk. Posterolateral approaches require extensive muscle dissection that adds to the risk of postoperative morbidity. The authors have developed a novel posterolateral, minimally invasive thoracic microendoscopic discectomy (TMED) technique that provides an approach to the thoracic spine which is associated with less morbidity. METHODS: Seven patients 23 to 54 years old with nine disc herniations underwent TMED. All lesions were soft lateral or midline thoracic disc herniations. Under fluoroscopic guidance with the patient positioned prone, the authors used a muscle dilation approach and the procedure was performed with endoscopic visualization through a tubular retractor. Based on a modified Prolo Scale, five patients experienced excellent results, one good, and one fair. No case required conversion to an open procedure. The mean operative time was 1.7 hours per level, and estimated blood loss was 111 ml per level. Hospital stays were short, and no complications occurred. CONCLUSIONS: The TMED is safe, effective, and provides a minimally invasive posterolateral alternative for treatment of thoracic disc herniation without the morbidity associated with traditional approaches. PMID- 15291023 TI - Surgical treatment of lumbosacral plexus injuries. AB - OBJECT: The purpose of this study was to analyze therapeutic possibilities and clinical outcomes in patients with lumbosacral plexus injuries to develop surgical concepts of treatment. METHODS: In a retrospective investigation 10 patients with injuries to the lumbosacral plexus were evaluated after surgery. The patients were assessed clinically, electrophysiologically, and based on the results of magnetic resonance imaging and computerized tomography myelography. In most patients a traction injury had occurred due to severe trauma that also caused pelvic fractures. In most cases the roots of the cauda equina of the lumbosacral plexus had ruptured. In cases of spinal root ruptures repair with nerve grafts were performed. In cases in which proximal stumps of the plexus could not be retrieved palliative nerve transfers by using lower intercostals nerves or fascicles from the femoral nerve were performed. CONCLUSIONS: Lesions of the proximal spinal nerves and cauda equina occur in the most serious lumbosacral plexus injuries. Patients with such injuries subjected to reconstruction of spinal nerves, repair of ventral roots in the cauda equina, and nerve transfers recovered basic lower-extremity functions such as unsupported standing and walking. PMID- 15291024 TI - Metabolic neuroimaging of the cervical spinal cord in patients with compressive myelopathy: a high-resolution positron emission tomography study. AB - OBJECT: The authors conducted a study to examine whether high-resolution [18F]fluorodeoxyglucose (FDG)-positron emission tomography (PET) could be used to visualize deterioration of cervical spinal cord function associated with various degrees of compression and to determine its potential usefulness during assessment of compressive myelopathy. METHODS: In 23 patients requiring decompressive surgery for myelopathy FDG-PET was performed. The preoperative findings of high-resolution FDG-PET were compared with the neurological scores and magnetic resonance (MR) imaging findings. The preoperative standardized uptake value (SUV) of FDG utilization rate of the cervical cord correlated with the pre- (r = 0.497, p = 0.016) and postoperative neurological scores (r = 0.595, p = 0.003), as well as with the rate of neurological improvement postoperatively (r = 0.538, p = 0.008). The FDG utilization rate did not correlate with the high signal intensity on T2-weighted MR images. CONCLUSIONS: Analysis of these results indicates that high-resolution FDG-PET imaging provides useful qualitative and quantitative estimates of impaired metabolic activity of the compromised cervical cord that correlate closely with the severity of neurological dysfunction. PMID- 15291025 TI - Suspended laminoplasty for wide posterior cervical decompression and intradural access: results, advantages, and complications. AB - OBJECT: Cervical laminoplasty is a recognized technique commonly used for multilevel posterior cervical decompression, and it is favored over laminectomy for maintaining spinal stability. Traditional hinge techniques, however, limit lateral exposure on one side and can limit dural exposure. The authors present their experience with a modified laminoplasty technique incorporating complete laminectomy and placement of titanium miniplate instrumentation. This method allows wide bilateral posterior decompression and unobscured dural access. METHODS: Twenty-eight patients (mean age 57 years) underwent cervical laminoplasty during a 4-year period. Twenty-seven patients presented with progressive cervical myelopathy. Seventeen patients (61%) had degenerative spondylotic stenosis; nine (32%) underwent resection of an intradural neoplasm. A mean of 3.5 levels were exposed and reconstructed. The follow-up period ranged from 4 months to 4 years (mean 15 months). The mean angular extension-flexion displacement measured between C-1 and C-7 was unchanged postoperatively, with preserved mobility across laminoplasty-treated segments in all patients. The anteroposterior diameter of the spinal canal increased 3.6 mm (27.2%) postoperatively (p = 0.004). In one patient an asymptomatic postoperative kyphosis developed. There were five cases of postoperative infection. One superficial infection resolved after intravenous antibiotic therapy alone, and four deep infections required surgical reexploration. CONCLUSIONS: The advantages of this technique over other laminoplasty methods include wide lateral spinal canal and intradural access, as well as preserved motion with partial restoration of the posterior tension band. PMID- 15291026 TI - Vulnerability of the subcostal nerve to injury during bone graft harvesting from the iliac crest. AB - OBJECT: Autologous bone graft harvesting from the iliac crest remains the gold standard for fusion surgery. One disadvantage of autologous bone harvesting is the patient's enduring postoperative pain at the donor site. Nerve injury is one of the postulated mechanisms that may account for this pain. The object of this study was to determine whether the lateral cutaneous branch of the subcostal nerve is vulnerable to injury in the process of obtaining grafts from the anterior iliac crest. METHODS: Anatomical dissections were performed on 10 cadaveric specimens to ascertain the size of the T-12 subcostal nerve and its position in relation to the iliac crest. CONCLUSIONS: The lateral cutaneous branch of the subcostal nerve may lie as close as 6 cm from the anterior superior iliac spine. This nerve is very vulnerable to injury when harvesting bone from the anterior iliac crest. Knowledge of the anatomy may decrease the risk of injury to this nerve. PMID- 15291027 TI - The use of bone morphogenetic protein-6 gene therapy for percutaneous spinal fusion in rabbits. AB - OBJECT: Fusion procedures in the lumbar spine have been performed in the US since 1911. Since that time, the indications and techniques for spinal fusion have evolved. Despite technical advancements, spinal fusion remains a major operation, and fusion nonunion rates of up to 35% are still reported. In this study, the authors were able to induce intertransverse process fusions in immune-competent New Zealand White rabbits by percutaneous administration of an adenoviral vector containing the bone morphogenetic protein (BMP-6) gene (Ad-BMP-6). The results represent an important step forward in finding new methods to increase the success and decrease the morbidity associated with spinal fusion. METHODS: Five New Zealand White rabbits were used. Injection of the adenoviral construct was performed at multiple levels (bilaterally) in each animal while using fluoroscopic guidance. Injection consisted of either Ad-BMP-6 or Ad-beta galactosidase (beta-gal) (control). Because multiple levels were injected, each animal served as an internal control. The animals underwent postinjection computerized tomography (CT) scanning at 7 and 14 weeks. After undergoing final CT scanning, the animals were killed and the spines were harvested. The fusion sites were analyzed by gross inspection, histopathological methods, and micro-CT studies. CONCLUSIONS: The results of this study show that an anatomically precise fusion can be accomplished by percutaneous administration of gene therapy. The next step in these studies will be extension of the technique to nonhuman primates and eventually to human clinical studies. PMID- 15291028 TI - Single-unit artificial intervertebral disc. AB - OBJECT: The authors describe a new type of artificial disc called a single-unit artificial disc (SUAD). It is a single-unit disc without components, and there is no fixation system with which to maintain it in the disc space. It is theorized that its shape, hardness, and surface consistency, together with the compressive force exerted by the body's axial load, should be adequate to maintain the position of the disc in the disc space. In this paper the authors present their results of the kinematic tests in which the stability and integrity of the SUADs was tested. METHODS: Panorobot was used for kinematic test fixture for fatigue testing for different types of SUAD. The test was performed after placing the disc between the C-5 and C-6 vertebral bodies (VBs) obtained from a cadaver. Eight pounds of weight was placed on the top of the container housing the C-5 VB to account for the weight of the head. The robot performed the following movements: 1) flexion-extension, 4.7 degrees each; 2) lateral left-right bending, +/- 2.1 degrees; and 3) coupled rotation, +/- 3.8 degrees. Two flat discs (FDs) of 85 durometer (D), four 30D FD, two recess-edged discs (REDs) of 85D, and four custom-designed discs (CDDs) (custom molded to the disc space) of 30D were tested. None of the discs showed cracks or breakage at the end of the study. After 1 million cycles of excursions all 85D and 30D FD, and both 85D REDs showed weight loss. The 30D CDDs showed minimal weight gain at 1 million cycles. One of the CDDs, tested up to 5 million cycles, did lose weight at 5 million cycles. One 85D FD was extruded after 0.2 million cycles and one 85D RED rotated within the disc space. All other discs maintained their position in the disc space. Dimensional changes were minimal. Scanning electron microscopy of particles collected from one 30D CDD sample after 1 million cycles showed rough irregular granular particles 1 to 600 microm in diameter. CONCLUSIONS: These findings show that a 30D SUAD can maintain its position in the disc space without any anchoring device. Furthermore, at 1 million cycles of testing of 30 D SUAD, FDs did as well as the CDDs. This may be because softer discs mold to changes in dimension of the disc space. In addition the softer discs tend to wear less than the harder ones. Further fatigue study of 10 million cycles is needed to determine long-term efficacy, and the effects of wear on particles surrounding the joints need to be studied. PMID- 15291029 TI - A biomechanical comparison of supplementary posterior translaminar facet and transfacetopedicular screw fixation after anterior lumbar interbody fusion. AB - OBJECT: Facet screw fixation is the lowest profile lumbar stabilization method. In this study the immediate biomechanical stability provided by the two different types of fixation are compared: translaminar facet screw (TLFS) and transfacetopedicular screw (TFPS) placement after anterior lumbar interbody fusion (ALIF) using a femoral ring allograft. Both facet screw fixation types were also compared with the gold standard, transpedicular screw and rod (TSR) fixation. METHODS: Twenty-four human lumbosacral spines were tested in the following sequence: intact state, after discectomy, after ALIF, and after TLFS, TFPS, or TSR fixation. Intervertebral motions were measured by a video-based motion capture system. The range of motion (ROM) and neutral zone (NZ) were compared for each loading to a maximum of 7.5 Nm. The ROMs for stand-alone ALIFs were less than but similar to those of the intact spine, but NZs were slightly increased in all modes. The ROMs for both TLFS and TFPS fixation were significantly decreased from those of the intact spine in all modes and those of the stand-alone ALIF in flexion and extension. The TLFS and TFPS fixations significantly reduced NZs to below that of the intact spine in all modes. Compared with NZs for ALIF, both types of fixation revealed significantly lower values, except for TLFS placement in lateral bending and TFPS fixation in lateral bending and rotation. There were no significant differences between TLFS and TFPS fixation. There were also no significant differences among both TLFS and TFPS and TSR fixations, except that TFPS was inferior to TSR in lateral bending. CONCLUSIONS: Stand-alone ALIF may not provide sufficient stability. Both facet fixations produced significant additional stability and both are comparable to TSR fixation. Although TFPS fixation revealed a slightly inferior result, TFPSs can be placed percutaneously with the assistance of fluoroscopic guidance and it makes the posterior facet fixation minimally invasive. Therefore, the TFPS fixation can be considered as a good alternative to TLFS fixation. PMID- 15291030 TI - A biomechanical comparison of three surgical approaches in bilateral subaxial cervical facet dislocation. AB - OBJECT: In bilateral cervical facet dislocation, biomechanical stabilities between anterior locking screw/plate fixation after anterior cervical discectomy and fusion (ACDFP) and posterior transpedicular screw/rod fixation after anterior cervical discectomy and fusion (ACDFTP) have not been compared using the human cadaver, although ACDFP has been performed frequently. In this study the stability of ACDFP, a posterior wiring procedure after ACDFP (ACDFPW), and ACDFTP for treatment of bilateral cervical facet dislocation were compared. METHODS: Spines (C3-T1) from 10 human cadavers were tested in the intact state, and then after ACDFP, ACDFPW, and ACDFTP were performed. Intervertebral motion was measured using a video-based motion capture system. The range of motion (ROM) and neutral zone (NZ) were compared for each loading mode to a maximum of 2 Nm. The ROM for spines treated with ACDFP was below that of the intact spine in all loading modes, with statistical significance in flexion and extension, but NZs were decreased in flexion and extension and slightly increased in bending and axial rotation; none of these showed statistical significance. The ACDFPW produced statistically significant additional stability in axial rotation ROM and in flexion NZ than ACDFP. The ACDFTP provided better stability than ACDFP in bending and axial rotation, and better stability than ACDFPW in bending for both ROM and NZ. There was no significant difference in extension with either ROM or NZ for the three fixation methods. CONCLUSIONS: The spines treated with ACDFTP demonstrated the most effective stabilization, followed by those treated with ACDFPW, and then ACDFP. The spines receiving ACDFP also revealed a higher stability than the intact spine in most loading modes; thus ACDFP can also provide a relatively effective stabilization in bilateral cervical facet dislocation, but with the aid of a brace. PMID- 15291031 TI - Biomechanical testing of anterior and posterior thoracolumbar instrumentation in the cadaveric spine. Invited submission from the Joint Section Meeting on Disorders of the Spine and Peripheral Nerves, March 2004. AB - OBJECT: Thoracolumbar burst fractures frequently require surgical intervention. Although the use of either anterior or posterior instrumentation has advantages and disadvantages, there have been few studies in which these two approaches have been compared biomechanically. METHODS: Ten human cadaveric spines were subjected to subtotal L-3 corpectomy. In five spines placement of L-3 wooden strut grafts with lateral L2-4 dual rod and screw instrumentation was performed. Five other spines underwent L1-5 pedicle screw fixation. The spines were fatigued between steps of the experiment. The spines were load tested with pure moments of 1.5, 3, 4.5, and 6 Nm in the intact state and after placement of instrumentation in six degrees of freedom (flexion, extension, right and left lateral bending, and right and left axial rotation). In axial rotation posterior instrumentation significantly increased spinal rigidity compared with that of the intact state, whereas anterior instrumentation did not. Combined anterior-posterior instrumentation did not significantly increase the rigidity of the spine when compared with anterior or posterior instrumentation alone. Posterior instrumentation alone provided a greater reduction in angular rotation compared with anterior instrumentation alone in all degrees of freedom; however, statistical significance was achieved only in extension at 6 Nm. CONCLUSIONS: The increased rigidity provided by pedicle screw instrumentation compared with the intact state or with anterior instrumentation is due to the longer construct spanning five levels and the three-column engagement of the pedicle screws. The decision to use anterior or posterior instrumentation should be based on the clinical necessity of canal decompression and correction of angulation. PMID- 15291032 TI - Mechanical properties and function of the spinal pia mater. AB - OBJECT: The pia mater has received little attention regarding its function in the deformation of the spinal cord under compression. In this study the mechanical properties and function of the spinal pia mater were investigated using three methods. METHODS: Spinal cord segments were excised from rabbits. The elastic modulus of the pia mater was measured by performing a tensile test using specimens with the pia mater intact and ones with the pia mater stripped off. The stiffness of the spinal cord was examined by performing a compression test with specimens containing an intact pia mater and ones with a pia mater that was incised at both sides. The cross-sectional area and circumference of the spinal cord were measured on axial views of magnetic resonance images in patients with cervical disc herniations before and after surgery. The pia mater had an elastic modulus of 2300 kPa, which was 460 times higher than that of spinal cord parenchyma. By covering the parenchyma, it tripled the overall elastic modulus of the spinal cord. The pia mater increased the stiffness of the spinal cord and enhanced its shape recovery after removal of the compression. The cross-sectional area of the spinal cord increased after surgery, whereas the circumference of the spinal cord changed little. CONCLUSIONS: The pia mater firmly covers the spinal cord and has a high elastic modulus; it therefore provides a constraint on the spinal cord surface. It prevents elongation of the circumference and produces a large strain energy that is responsible for shape restoration following decompression. PMID- 15291033 TI - Cervical spinal cord delivery of a rabies G protein pseudotyped lentiviral vector in the SOD-1 transgenic mouse. Invited submission from the Joint Section Meeting on Disorders of the Spine and Peripheral Nerves, March 2004. AB - OBJECT: Lentiviral vectors may constitute a vehicle for long-term therapeutic gene expression in the spinal cord. In amyotrophic lateral sclerosis, spinal cord sclerosis and altered axonal transport pose barriers to therapeutic gene distribution. In the present study the authors characterize gene expression distribution and the behavioral impact of the rabies G (RabG) protein pseudotyped lentiviral vector EIAV.LacZ through cervical spinal cord injection in control and Cu/Zn superoxide dismutase-1 (SOD-1) transgenic mice. METHODS: Seven-week-old SOD 1 transgenic mice and their wild-type littermates underwent exposure of the cervicomedullary junction and microinjection of RabG.EIAV.LacZ or vehicle. The Basso-Beattie-Bresnahan locomotor score, grip strength meter, and Rotarod assays were used to assess the effects of disease progression, spinal cord microinjection, and lentiviral gene expression. Spinal cords were removed when the mice were in the terminal stage of the disease. The distribution of LacZ gene expression was histologically evaluated and quantified. Direct cervical spinal cord microinjection of RabG.EIAV.LacZ results in extensive central nervous system uptake in SOD-1 transgenic mice; these findings were statistically similar to those in wild-type mice (p > 0.05). Gene expression lasts for the duration of the animal's survival (132 days). The SOD-1 mutation does not prevent retrograde axonal transport of the vector. Three behavioral assays were used to demonstrate that long-term gene expression does not alter sensorimotor function. In comparison with normative data, vector injection and transgene expression do not accelerate disease progression. CONCLUSIONS: Direct spinal cord injection of RabG.EIAV vectors represents a feasible method for delivering therapeutic genes to upper cervical spinal cord and brainstem motor neurons. Distribution is not affected by the SOD-1 mutation or disease phenotype. PMID- 15291034 TI - Anterior atlas fracture following suboccipital decompression for Chiari I malformation. Report of two cases. AB - Chiari I malformation, a congenital disorder involving downward displacement of the cerebellar tonsils through the foramen magnum, is often treated surgically by performing suboccipital craniectomy and C-1 laminectomy. The authors report two cases in which fracture of the anterior atlantal arch occurred during the postoperative period following Chiari I decompression and C-1 laminectomy causing significant neck pain. The findings indicate that interruption of the integrity of the posterior arch of C-1, iatrogenically or otherwise, confers increased risk of anterior arch fracture. A C-1 fracture should therefore be considered in the differential diagnosis of posterior cervical pain in patients who have previously undergone decompression for Chiari I malformation. PMID- 15291035 TI - Charcot spine: a complication of medullary arteriovenous malformation. Case illustration. PMID- 15291036 TI - Thoracic spinal nerve hemangioblastoma. Case illustration. PMID- 15291037 TI - [The heart's meanings in prehispanic Mexico]. PMID- 15291038 TI - [On the 25th anniversary of the I Chavez death. About the humanistic education of the physician]. PMID- 15291039 TI - [Study of Magnolia grandiflora extracts in guinea pigs cardiac muscle]. AB - Several extracts from diverse Magnolia grandiflora varieties were pharmacological evaluated in the cardiac muscle. From March to July, flowers and leaves from Magnolia grandiflora, native from the National Institute of Cardiology "Ignacio Chavez", from north, west, and orient zones from Mexico City, and from Puebla, Colima and Chiapas states were collected. They were separately processed and the extracts were obtained by maceration with ethanol-water (1:3 v/v) at 4 degrees C during two weeks. Qualitative analysis was accomplished with thin-layer, column and high-performance liquid chromatographies (HPLC). Functional and molecular analysis was made by specific chemical reactivity and by protonic magnetic resonance (RMN 1H). Pharmacological evaluation was completed in isolated and perfused male guinea pigs hearts. Extracts, fractions, and compounds were administrated by serial bolus in a gradual dose-response curves study in which left intraventricular pressure and coronary perfusion pressure were recorded, evaluating by such the positive inotropic and vasodilator effects of Magnolia grandiflora extracts. Vulgarenol and 2-p-hydroxyphenyl-2-hydroxy-ethylamine were isolated and identified, and the obtained results suggest that its positive inotropic and vasodilator effects are owed to these substances, being complemented by magnograndiolide and tyramine. PMID- 15291040 TI - [Evaluation of predisposing factors for mediastinal bleeding in myocardial revascularization surgery. Role of acetylsalicylic acid, other platelet adhesion inhibitors, and anticoagulants]. AB - OBJECTIVE: To determine the main factors for perioperative mediastinal bleeding during coronary artery by-pass grafting and to establish the role of acetylsalicylic acid, other inhibitors of platelet adhesion, and anticoagulants in its occurrence. METHODS: A multivariate analysis was performed to the data obtained from 251 patients subjected to coronary artery by-pass grafting in the year 2002. RESULTS: There were no significant differences for the occurrence of perioperative bleeding induced by the preoperative administration of acetylsalicylic acid. No significant increment in the need for blood, platelet, or cryoprecipitates transfusion existed. Patients receiving non-fractionated or low molecular weight heparin had a significantly greater (p < 0.001) transoperative bleeding than patients without this drug. Administration of the other studied drugs did not increase either mediastinal bleeding or the need for blood derivatives. CONCLUSIONS: No statistical evidence was found to suspend administration of inhibitors of platelet adhesion in patients with coronary syndromes, subjected to coronary artery by-pass grafting. However, data obtained suggest the convenience of suspending administration of low molecular weight or non-fractionated heparin to patients subjected electively to coronary artery by pass grafting. PMID- 15291041 TI - [Intracardiac echocardiography (ICE) in the operating room as a support measure in the evaluation of immediate surgical results in congenital cardiopathies]. AB - The utilization of intracardiac echocardiography (ICE) is a recent alternative in the control of procedures in the Cath Lab. In this paper we presents the results of a pilot study, in 10 patients submitted to total correction of its congenital heart disease, in who the analysis of the surgical results was done in base to the application of intracardiac echocardiography in operations room. In all the patients it was possible to obtain adequate images with ICE, there was a case false negative, a Total anomalous return Pulmonary Veins, in whom there not indentified a obstruction in right pulmonary veins flow. We conclude that ICE is a good alternative in the evaluation of surgical results. PMID- 15291042 TI - [Complex septal rupture due to an acute myocardial infarct. Echocardiographic study]. PMID- 15291043 TI - [Hypertension guidelines in Mexico]. PMID- 15291045 TI - Caring behaviors as perceived by nurse practitioners. AB - PURPOSE: To investigate nurse pratctitioners' (NPs') perceptions of their own caring behaviors and to examine NPs' demographics as a function of their caring behaviors. DATA SOURCES: Responses to the Caring Behaviors Inventory(CBI) and a demographic inquiry from 348 NPs in Louisiana. CONCLUSIONS: CBI mean scores and subscale scores were high for all 348 NPs. No statistically significant difference was found between male NPs' and female NPs' total mean CBI scores or between urban or rural total mean CBI scores. The interaction between nurse gender and area o practice was not statistically significant. IMPLICATIONS FOR PRACTICE: NPs often work in clinic situations where productivity is the most valued characteristic and where little time is afforded for identifying caring behaviors of the NP and/or establishing a caring relationship with the patient. NPs must be extremely conscious of the need not to "throw out the baby with the bathwater" and sacrifice characteristics that are inherent in nursing for those emphasized in primary care practice. As their responsibilities in the health care setting continue to expand, NPs must continually evaluate and validate their roles to ensure quality care that satisfies patients. PMID- 15291044 TI - Osteoporosis prevention starts in adolescence. AB - PURPOSE: To review the factors that influence the development of osteoporosis later in life, to relate those factors to adolescent health behaviors, and to provide nurse practitioners (NPs) with information on which to base evaluation and management decisions in primary care. DATA SOURCES: Review of selected scientific literature with a focus on adolescent health and osteoporosis. CONCLUSIONS: Osteoporosis is a major public health concern that affects more than 10 million people in the United States, resulting in annual costs to our health care system in excess of 13.5 billion dollars. Approximately 40% of women will experience a fracture due to osteoporosis. NPs should aim osteoporosis prevention efforts toward adolescents by encouraging habits that promote bone growth and development. IMPLICATIONS FOR PRACTICE: It is imperative that female adolescents achieve adult peak bone mass prior to the age of 30, and the bone-building years before the age of 18 are especially important. NPs are ideally positioned to provide the screening and counseling to ensure that adolescents have the best chance to develop the habits that will result in peak adult bone mass. PMID- 15291046 TI - Obesity and physical activity in college women: implications for clinical practice. AB - PURPOSE: To investigate the relationships between levels of physical activity, health attitudes and behaviors, and specific health indicators in women attending college. DATA SOURCES: A convenience sample of 116 college women, ages 18 to 24 years, participated in this research study at a moderate-sized midwestern university. The data were obtained through a self-administered questionnaire; trained technicians collected physiological measurements. CONCLUSIONS: The young women in this study had, on average, normal body mass indexes (BMIs) and reported activity levels consistent with or greater than the Centers for Disease Control and Prevention/American College of Sports Medicine guidelines. Items used to assign participants into the appropriate stage of the transtheoretical model of change were correlated with participants' perceived personal physical activity levels. Similarly, the participants, whose scores fell in the higher stages of the transtheoretical model, reported greater levels of physical activity; consumption of more fruits, vegetables, and water; and less consumption of high fat/high-calorie foods. IMPLICATIONS FOR PRACTICE: The years between ages 18 and 24 are a critical time in the lives of young women. During this period, they develop physical activity and nutrition habits that will affect their health across the life span. Because of the sometimes insidious development of major health problems, young women's current health status may not accurately reflect the possible long- term results of negative health habits. Nurse practitioners (NPs) have many opportunities to identify and address major factors that, if unattended, may threaten the life-long health status of women. Health teaching in the areas of physical activity and dietary habits may be useful even in young women who appear to be healthy, are of normal weight, and are physically active. Poor dietary habits, if unattended, may eventually contribute to the development of obesity and related illnesses. PMID- 15291047 TI - Characteristics of and problems with primary care interactions experienced by an ethnically diverse group of women. AB - PURPOSE: To reflect women's voices as they discussed characteristics of health care interactions and spontaneously identified problems in primary care interactions. DATA SOURCES: Five successive meetings with an ethnically diverse group of eight women were held to discuss primary care interactions with nurse practitioners. Field notes, seating charts, participant interaction notations, session transcripts, and audiotapes were repeatedly reviewed to identify significant statements. These were grouped into common categories to identify essential themes. Results were validated with participants. CONCLUSIONS: Components of primary care interactions included the process of making an appointment, access to the clinic, comfort of the waiting area and clinic rooms, and interactions with staff and clinicians. Problems were identified with interactions. The overarching issue was a lack of caring--a pervasive attitude demonstrated when clinicians failed to show concern, did not listen, were not trustworthy, or treated the women with disrespect or prejudice. IMPLICATIONS FOR PRACTICE: The women strongly valued caring clinicians. Caring, according to these women, is demonstrated when clinicians treat women as equals and show respect for their individual knowledge and life experiences. PMID- 15291048 TI - Severe exacerbations of asthma. AB - All asthmatics regardless of their perceived severity, are at risk of exacerbation, particularly if they are suboptimally treated in the outpatient arena. Fortunately most patients recover after administration of bronchodilators and anti-inflammatory medications, but preventable deaths continue to occur and refractory cases result in hospitalization and need for mechanical ventilation. We begin this article by reviewing the pathophysiology of acute exacerbations to build a foundation for the assessment of clinical status and to provide the rationale for a carefully contemplated and evidence-based therapeutic approach. We end this article with an in-depth examination of the particular problems that are encountered during mechanical ventilation and offer a strategy that helps minimize complications. In the final analysis, however, the greatest gains in the field of acute asthma will come not from its treatment but from its prevention by enhanced educational and environmental efforts and by the delivery of optimal medications at home. PMID- 15291049 TI - Community-acquired pneumonia: updates in assessment and management. AB - Community-acquired pneumonia (CAP) is a significant health condition. Knowledge of the clinical presentation and treatment of CAP are important for critical care nurses as up to 20% of patients with CAP require hospitalization and in-patient management. Patients with severe CAP requiring intensive care unit (ICU) treatment often require aggressive management including mechanical ventilation and multisystem organ support. This article presents an overview of CAP, including the presentation of typical and atypical CAP, clinical findings, and the essentials of management. Treatment differences between CAP and healthcare acquired pneumonia and nursing implications are also highlighted. PMID- 15291050 TI - Nosocomial pneumonia. AB - Nosocomial pneumonia (NP) is defined as pneumonia that develops within 48 hours or more of hospital admission and which was not developing at the time of admission. Nosocomial pneumonia, also known as hospital-acquired pneumonia (HAP), is the second most common hospital infection, while ventilator-associated pneumonia represents the most common intensive care unit (ICU) infection. Nosocomial pneumonia significantly contributes to morbidity, mortality, and escalating healthcare costs because of increases in antibiotic prescription and administration, length of ICU stay, and length of hospital stay. Aspiration and colonization of the upper respiratory tract seem to be the major pathogenetic mechanisms for the development of NP, either in intubated or spontaneously breathing patients. The microbiology of NP depends on the timing of onset. In early-onset NP, the responsible pathogens are generally endogenous community acquired pathogens. In late-onset NP, the responsible microbes include potentially multi-drug-resistant nosocomial organisms residing in oropharyngeal or gastric contents. Important risk factors for development of NP include coma, intubation, prolonged mechanical ventilation, repeated intubations, supine positioning, and long-term antibiotic use. The most significant preventive measures include routine hand washing and avoidance of (1) the supine position, (2) inappropriate antibiotics, and (3) overuse of H2-antagonists for stress ulcer prophylaxis. Accurate diagnosis of NP is difficult and controversial, warranting consideration for the application of invasive quantitative culture techniques over tracheal aspirates. Empiric antibiotic treatment should be prompt, starting on clinical suspicion, and based on local ICU pathogen epidemiology and antibiotic resistance patterns and on a deescalating antibiotic strategy. Innovative antibiotic strategies, such as antibiotic rotation, to help prevent the emergence of multi-drug-resistant pathogens and improve survival should be considered. PMID- 15291051 TI - Mechanical ventilation: what have we learned? AB - Mechanical ventilation is the second most frequently performed therapeutic intervention after treatment for cardiac arrhythmias in intensive care units today. Countless lives have been saved with its use despite being associated with a greater than 30% in-hospital mortality rate. As life expectancies increase and people with chronic illnesses survive longer, artificial support with mechanical ventilation is also expected to rise. In one survey, over half of senior internal medicine residents reported their training on mechanical ventilation as inadequate, whereas the majority of critical care nurses reported having received no formal education on its use. Technological advances resulting in the availability of sleeker ventilators with graphic waveform displays and new modes of ventilation have challenged the bedside clinicians to incorporate this new data along with evidenced-based research into their daily practice. A review of current thoughts on mechanical ventilation and weaning is presented. PMID- 15291052 TI - The evolution: Handwashing to hand hygiene guidance. AB - Handwashing is a fundamental principle and practice in the prevention, control, and reduction of healthcare-acquired infection. Advocated by Semmelweiss (Nursing, The Finest Art: An Illustrated History. St Louis: Mosby; 1985:204) from the 1800s to resolve an obstetric morbidity and mortality occurrence, the simple act of hand cleansing portrays the intuitive benefits to basic hygiene, health continuum, and, most important, disease prevention. According to recently published guidance (MMWR Morb Mortal Wkly Rep. October 25, 2002;51:32-34), the term handwashing is replaced by the new term hand hygiene, which includes hand cleansing, hand disinfecting, and surgical hand scrub. This article focuses on the published guidance, blending the salient aspects of hand hygiene practices from noted champions, reinforcing the aesthetics of meticulous cleansing, to guidance on its practice in healthcare settings. In healthcare, the principle of "clean hands are healing hands" bears value and demands compliance in order to prevent and control infectious processes while protecting the person from acquiring infectious diseases. PMID- 15291053 TI - Keystone aberration correction in overhead projectors. AB - Keystone distortion that occurs in overhead projectors when the projecting lens head is tilted upward to a high screen is commonly observed. Here we suggest a modification of the typical overhead projector to eliminate this distortion of the image. PMID- 15291054 TI - Optical fiber relative-humidity sensor with polyvinyl alcohol film. AB - We describe a fiber-optic relative-humidity (RH) sensor comprising a moisture sensitive overlay on a single-mode side-polished fiber. The hygroscopic polymeric material deposited was polyvinyl alcohol (PVA), which proved to have good adherence and stability. The film reached a fast equilibrium with atmospheric moisture (in less than 1 min), inducing changes in the output optical power of approximately 10 dB for the 70%-90% RH range. To yield a low-cost device, single mode standard communication fibers were used; therefore all the components of the sensor can be commercial, mass-produced telecommunication devices. The experimental results obtained are consistent with the expected behavior of the system; the output power decreases because of losses in the polished region of the fiber as the refractive index of its external medium approaches the fiber core value. Because the external medium is PVA film, its refractive index changes in response to its water content. PMID- 15291055 TI - Three-dimensional optical tomographic imaging of supersonic jets through inversion of phase data obtained through the transport-of-intensity equation. AB - We report experimental results of quantitative imaging in supersonic circular jets by using a monochromatic light probe. An expanding cone of light interrogates a three-dimensional volume of a supersonic steady-state flow from a circular jet. The distortion caused to the spherical wave by the presence of the jet is determined through our measuring normal intensity transport. A cone-beam tomographic algorithm is used to invert wave-front distortion to changes in refractive index introduced by the flow. The refractive index is converted into density whose cross sections reveal shock and other characteristics of the flow. PMID- 15291056 TI - Optical coherence tomography system with no high-precision scanning stage and stage controller. AB - Optical coherence tomography (OCT) is a novel technique for noninvasive imaging based on the use of a low-coherence interferometer. Conventionally, obtaining high-resolution images requires the use of high-precision sample and scanning stages and a stage controller for simultaneous measurement of the refractive index and the thickness of an optical sample. However, in this study a novel optical-fiber-type OCT system is developed that does not need both a high precision scanning stage and a stage controller. Additionally, two signal demodulation processes are described. Compared with that of conventional OCT systems, the current configuration eliminates the high-precision scanning stage and stage controller and is therefore cheaper and less complex. Also, this new technique could be applied to conventional OCTs in biotissue scanning. PMID- 15291057 TI - Design and processing of high-density single-mode fiber arrays for imaging and parallel interferometer applications. AB - The design and fabrication procedures for implementing a high-density (16-microm center spacing) single-mode fiber (SMF) array are described. The specific application for this array is a parallel optical coherence tomography system for endoscopic imaging. We obtained fiber elements by etching standard single-mode SMF-28 fibers to a diameter of 14-15 microm. We equalized 1-m lengths of fiber to within 1 mm by using a fiber interferometer setup, and we describe a method for packaging arrays with as many as 100 fibers. PMID- 15291058 TI - Fast scanning method for one-dimensional surface profile measurement by detecting angular deflection of a laser beam. AB - A fast scanning method for one-dimensional surface profile measurement is proposed. The profile is measured by integration of a slope distribution of the surface obtained from angular deflection of a scanning laser beam. A scanning optical system that consists principally of a spherical concave mirror and a rotating scanner mirror has reasonably low cost and is insensitive to mechanical vibration because of its high-speed scanning, of the order of milliseconds. A surface profile of a polygonal mirror along a 5-mm width was measured with the scanning method and with an interferometer. The root-mean-square difference between the two measured results is 0.98 nm. PMID- 15291059 TI - Instantaneous velocity displacement and contour measurement by use of shadow moire and temporal wavelet analysis. AB - A temporal wavelet analysis method is proposed for velocity, displacement, and three-dimensional surface-profile measurement of a continuously deforming object by use of the shadow moire technique. A grating is placed close to a deforming object, and its shadow is observed through the grating. The moire fringe patterns, generated by the interference of the grating lines and their shadows, are captured by a high-speed CCD camera with a telecentric gauging lens. Instantaneous frequency of gray-value variation is evaluated point by point with the continuous wavelet transform. From the instantaneous frequency of each point on the object, the velocity, displacement, and high-quality surface profile at different instants can be retrieved. In this application, two specimens are tested to demonstrate the validity of the proposed method: One is a small coin with a rigid body motion, and the other is a simply supported beam subjected to a central point load. The results are compared with those obtained from temporal Fourier-transform and mechanical stylus methods. PMID- 15291060 TI - In-plane dynamic speckle interferometry: comparison between a combined speckle interferometry/speckle correlation and an update of the reference image. AB - A common problem during study of, for instance, tensile tests with interferometers is that the sample moves too much so that the speckles decorrelate and no phase information is obtained. Two ways to overcome this problem are compared: a combination of speckle interferometry and speckle correlation and a method in which the reference image is updated during the experiment. The comparison shows that both techniques can be used to measure the deformation of an object even if it is exposed to rigid body motions. Both techniques are applied to measurements of microscale deformation fields of an adhesive joint in a carbon-fiber epoxy composite. PMID- 15291061 TI - Nonlinear Faraday rotation for optical limitation. AB - The possible use of the nonlinear Faraday effect for optical limitation of the laser power is investigated in a resonant Faraday medium placed between two crossed polarizers. The results are comparable with those obtained at strong magnetic fields as a result of the linear Faraday effect. Advantages of the method are the narrow bandwidth and the wide field of view. The investigations are interesting from the viewpoint of applications for optical sensor protection and automation of the experiment. All measurements are performed at the F(g) = 2 -> F(e) = 1 hyperfine structure transition of the 87Rb D1 line. PMID- 15291062 TI - Absolute linearity measurements on HgCdTe detectors in the infrared region. AB - The nonlinearity characteristics of photoconductive and photovoltaic HgCdTe detectors were experimentally investigated in the infrared wavelength region by use of the National Physical Laboratory detector linearity measurement facility. The nonlinearity of photoconductive HgCdTe detectors was shown to be a function of irradiance rather than the total radiant power incident on the detector. Photoconductive HgCdTe detectors supplied by different vendors were shown to have similar linearity characteristics for wavelengths around 10 microm. However, the nonlinearity of response of a photovoltaic HgCdTe detector was shown to be significantly lower than the corresponding value for photoconductive HgCdTe detectors at the same wavelength. PMID- 15291063 TI - Analysis of optical frequency-modulated continuous-wave interference. AB - I systematically analyze the theory of optical frequency-modulated continuous wave (FMCW) interference. There are three different versions of optical FMCW interference, discussed in detail: sawtooth-wave optical FMCW interference, triangular-wave optical FMCW interference, and sinusoidal-wave optical FMCW interference. The essential concepts and technical terms are clearly defined, the necessary simplifications are introduced according to the characteristics of optical waves, and the formulas used to calculate the signal intensities under two different situations (static and dynamic) are properly derived. Advantages and limitations of each version of optical FMCW interference are also discussed. PMID- 15291064 TI - Class of 4 + 1-phase algorithms with error compensation. AB - It is well known that phase-shifting interferometry suffers from inaccuracy in the presence of phase-shifting errors. We have proved the limitation of using a 4 phase algorithm to reduce the phase-measurement error in the presence of the phase-shifting error. A class of 4 + 1-phase error compensating algorithms is formulated. It is shown that the proposed algorithms can effectively minimize the effects of the constant phase-shifting error and possess a superior performance than existing error-compensating algorithms. The effectiveness of the proposed algorithm is demonstrated by computer simulations and experiments. PMID- 15291065 TI - Noninterferometric and nontomographic iterative method for field retrieval. AB - A new method of field recovery is proposed based on the fact that an arbitrary light beam can be expressed as a finite sum over orthogonal field distributions with unknown coefficients. If these field distributions are eigenmodes of a specific waveguide, the coefficients of the field decomposition in orthogonal eigenmodes can be determined iteratively when the unknown field is passed through a series of waveguides that support an increasing number of modes. This series of waveguides can be replaced by a single reconfigurable electro-optic waveguide, which one can use to recover the unknown field by performing only intensity measurements. PMID- 15291066 TI - Method for reduction of background artifacts of images in scanning holography with a Fresnel-zone-plate coded aperture. AB - Near-infrared scanning holography with a Fresnel zone plate (FZP) coded aperture has potential applications in imaging through turbid media. However, the nonnegative intensity-distribution function of the FZP coded aperture introduces the background artifacts into the reconstructed images, reducing the contrast and the signal-to-noise ratio (SNR) of the images. A novel method termed as the composite hologram is proposed to reduce the artifacts. The computer simulations showed that the contrast and the SNR of the reconstructed images had improvements of at least 50.2% and 5.58-dB, respectively, compared with the conventional method. The composite hologram of a metal ring with a 6.0-mm diameter made by a wire with a 0.4-mm diameter immersing in 1% intralipid solution was recorded, and the reconstruction was performed numerically. The experimental results demonstrated that the contrast and the SNR of the reconstructed image had improvements of at least 32.3% and 2.51-dB, respectively. PMID- 15291067 TI - Full characterization of holographic reflection gratings recorded on BB640 emulsions. AB - A complete characterization of unslanted holographic reflection gratings with high diffraction efficiency recorded on ultrafine grain emulsion BB640 has been achieved. By use of a wavelength-dependent absorption coefficient, the diffraction efficiency, the replay wavelength, and the spectral bandwidth of each recording has been obtained. Corresponding index modulation, absorption, and effective thickness have been obtained with high accuracy by fitting the experimental data with Kogelnik's theory. PMID- 15291068 TI - Perfectly matched layer in numerical wave propagation: factors that affect its performance. AB - The perfectly matched layer (PML) boundary condition is generally employed to prevent spurious reflections from numerical boundaries in wave propagation methods. However, PML requires additional computational resources. We have examined the performance of the PML by changing the distribution of sampling points and the PML's absorption profile with a view to optimizing the PML's efficiency. We used the collocation method in our study. We found that equally spaced field sampling points give better absorption of beams under both optimal and nonoptimal conditions for low PML widths. At high PML widths, unequally spaced basis points may be equally efficient. The efficiency of various PML absorption profiles, including new ones, has been studied, and we conclude that for better numerical efficiency it is important to choose an appropriate profile. PMID- 15291069 TI - Compact stacking of diode lasers for pulsed light sources of high brightness. AB - A compact stacking architecture for high-power diode-laser arrays is proposed and compared with traditional stacks. The objective of compact stacking is to achieve high brightness values without the use of microlenses. The calculated brightness for a compact stack is over 300 W mm(-2) sr(-1), which is approximately 40 times higher than that of a traditional stack made of similar laser emitters. Even higher brightness values of over 600 W mm(-2) sr(-1) were reached in practice. A laser head was manufactured in which the light from several compact laser stacks could be fiber coupled or the light could be transformed to a highly uniform beam. PMID- 15291070 TI - Study of absorption and reflection in solid-state random laser media. AB - Comparing the absorption spectrum of a single-crystalline material and the reflection spectrum of its powder, we have derived a simple empirical formula for the reflection coefficient of absorbing powder. The proposed equation, which slightly resembles the Lambert-Beer law for transmission, does not correspond to the equation derived in the approximation of a simple one-dimensional diffusion model. PMID- 15291071 TI - Three-dimensional analysis of a hybrid photonic crystal-conventional waveguide 90 degree bend. AB - We present a three-dimensional (3D) analysis of a hybrid photonic crystal conventional waveguide 90 degree bend proposed previously [Opt. Express 10, 1334 (2002)] as an ultracompact component for large-scale planar lightwave circuit integration. Both rigorous 3D finite-difference time-domain modeling and a simple perfect mirror model analysis were carried out for different Si post heights in the photonic crystal region. Results show that the bend efficiency increases rapidly with Si post height. For a post height of 6.5 microm, this structure yields a bend efficiency of 97.3% at a wavelength of 1.55 microm for 90 degree bends in 2 microm x 2 microm square channel conventional waveguides with a refractive index contrast of 3.55%, which is very close to the bend efficiency of 98.2% for the corresponding two-dimensional problem. Our 3D analysis permits the examination of issues such as out-of-plane scattering loss and the effects of finite Si post height that are not considered in two dimensions. PMID- 15291072 TI - Time evolution of reflective thermal lenses and measurement of thermal diffusivity in bulk solids. AB - A simple method for optically measuring the thermal diffusivity of solids is demonstrated. The thermal displacement created on a substrate by a focused laser beam is determined from the divergence that it induces in a weak probe beam. The dynamics of the surface lens and the amplitude of the probe beam's divergence are then used to determine the thermal diffusivity of the substrate. Several materials that span a wide range of thermal properties are studied. PMID- 15291073 TI - Above-water radiometry in shallow coastal waters. AB - Above- and in-water radiometric data were collected from two coastal platforms: a small boat and an oceanographic tower. The above-water data were processed with and without a correction for bidirectional effects (Q02 and S95, respectively). An intercomparison of water-leaving radiances over a wide range of environmental conditions showed (a) total uncertainties across the blue-green domain were to within 4%, (b) a convergence of the Q02 method with the in-water method (average Q02 intercomparisons were to within 4%), and (c) chlorophyll a concentrations derived from Q02 reflectances and the OC4V4 (Ocean Color 4 Version 4) algorithm agreed with independent high-performance liquid-chromatography determinations to within approximately 32%. PMID- 15291074 TI - Innoventions: cocktails of fantasy and science. PMID- 15291075 TI - Artificial aortic valves: an overview. AB - This review discusses strategies that may address some of the limitations associated with replacing diseased or dysfunctional aortic valves with mechanical or tissue valves. These limitations range from structural failure and thromboembolic complications associated with mechanical valves to a limited durability and calcification with tissue valves. In pediatric patients there is an issue with the inability of substitutes to grow with the recipient. The emerging science of tissue engineering potentially provides an attractive alternative by creating viable tissue structures based on a resorbable scaffold. Morphometrically precise, biodegradable polymer scaffolds may be fabricated from data obtained from scans of natural valves by rapid prototyping technologies such as fused deposition modelling. The scaffold provides a mechanical profile until seeded cells produce their own extra cellular matrix. The microstructure of the forming tissue may be aligned into predetermined spatial orientations via fluid transduction in a bioreactor. Although there are many technical obstacles that must be overcome before tissue engineered heart valves are introduced into routine surgical practice these valves have the potential to overcome many of the shortcomings of current heart valve substitutes. PMID- 15291076 TI - Short, thrice-weekly hemodialysis is inadequate regardless of small molecule clearance. AB - Chronic hemodialysis sessions, as developed in Seattle in the 1960s, were long procedures with minimal intra- and interdialytic symptoms. Over the next three decades, financial and logistical pressures related to the overwhelming number of patients requiring hemodialysis created an incentive to shorten dialysis time to four, three, and even two hours per session in a thrice weekly schedule. This method spread rapidly, particularly in the United States, after the National Cooperative Dialysis Study suggested that time of dialysis is of minor importance as long as urea clearance multiplied by dialysis time and scaled to total body water (Kt/Vurea) equals 0.95-1.0. This number was later increased to 1.3, but the assumption that hemodialysis time is of minimal importance, as long as it is compensated by increased urea clearance, remained unchanged. Patients accepted short dialysis as a godsend, believing that it would not be detrimental to their well being and longevity. However, Kt/Vurea measures only removal of low molecular weight substances and does not consider removal of larger molecules. Nor does it correlate with the other important function of hemodialysis, namely ultrafiltration. Whereas patients with substantial residual renal function may tolerate short dialysis sessions, patients with little or no urine output tolerate short dialyses poorly because at a given interdialytic weight gain the ultrafiltration rate is inversely proportional to dialysis time. Rapid ultrafiltration is associated with cramps, nausea, vomiting, headache, fatigue, hypotensive episodes during dialysis, and hangover after dialysis; patients remain fluid overloaded with subsequent poor blood pressure control leading to left ventricular hypertrophy, diastolic dysfunction, and high cardiovascular mortality. Short, high-efficiency dialysis requires high blood flow, which increases demands on blood access. The classic, wrist arteriovenous fistula, the access with the best longevity and lowest complication rates, provides "insufficient" blood flow and is replaced with an arteriovenous graft fistula or an intravenous catheter. Moreover, to achieve high blood flows, large diameter intravenous catheters are used; these fit veins "too tightly" and so predispose to central-vein thrombosis. Longer hemodialysis sessions (5-8 hours, thrice weekly), as practiced in some centers, are associated with lower complication rates and better outcomes. Frequent dialyses (four or more sessions per week) with total weekly dialysis time sufficient to allow gentle ultrafiltration rates provide the best clinical results, but are associated with increased costs which are not properly reimbursed in the USA at present. Therefore, it is my strong belief that before a more appropriate reimbursement is available, a wide acceptance of longer, gentler dialysis sessions, in the current thrice weekly schedule, would improve overall hemodialysis results, decrease access complications, hospitalizations and mortality, particularly in anuric patients. Kt/Vurea should be abandoned as a measure of dialysis quality. The formula suggests that it is possible to decrease t as long as K is proportionately increased, but this is not true. The use of rigid, quantitative guidelines (e.g., spKt/Vurea of 1.3 per dialysis) assumes that all patients behave identically in response to therapeutic maneuvers, like the mean of the group, but this is also not true. The individual, clinical approach assumes that there are differences among patients, which require adjustment of dialysis schedule for each patient. PMID- 15291077 TI - Hemoglobin normalization results in lower dialysis dose, despite high dialysate flow. Single needle offers inadequate dialysis. AB - Anemia correction by erythropoietin favorably affects dialysis outcome but may also reduce dialysis efficiency increasing morbidity and mortality. Single needle dialysis (SN) and high dialysate flow (DF) are dialysis variations. We studied the effect of hemoglobin (Hb) normalization on dialysis adequacy under high DF. We also compared double needle (DN) and SN dialysis efficiency. Seventeen stable anuric patients (13 M, 4 F), aged 62 (40-90), on hemodialysis for 48 months (8 204), were studied in two, 6 months apart, periods of low (A) and high Hb (B), during a midweek 4 h dialysis with DN and SN. DF was 500 in A and 800 ml/min in B. Rebound urea samples, 20 min post dialysis, were used for computer calculated double pool urea kinetics. Hb levels were 128 +/- 8 g/L (B) vs. 119 +/- 14 g/L (A), P < 0.03. Despite the use of higher DF less dialysis was delivered in B vs. A, under DN or SN (DN: URR 64.8 +/- 5.8 vs. 69.7 +/- 5.2%, Kt/Vequil. 1.09 +/- 0.19 vs. 1.26 +/- 0.21, nPCR 1.37 +/- 0.29 vs. 1.60 +/- 0.36g/kg/day, changes <0.001, SN: URR 49.7 +/- 7.5% vs. 52.6 +/- 8.8%, Kt/Vequil. 0.74 +/- 0.16 vs. 0.82 +/- 0.23, nPCR 1.05 +/- 0.33 vs. 1.20 +/- 0.31, changes NS). SN was found significantly (P < 0.001) less efficient than DN in A and B. Serum creatinine drop was significantly (P < 0.001) less in both periods with SN vs. DN. Hb (SN in B) correlated inversely to Kt/V (r = -0.5705, P < 0.02) and URR (r = -0.6432, P = 0.005). Hb correction to normality is associated with a decrease in dialysis efficiency. The use of high dialysate flow does not compensate for this loss. SN delivers inadequate dialysis independently of dialysate flow or hemoglobin concentration. PMID- 15291078 TI - Endotoxin removal from whole blood by a novel adsorption resin: efficiency and hemocompatibility. AB - The structural component of Gram- bacteria, endotoxin (ET), induces the release of endogenous mediators of sepsis. Attempts to remove these downstream molecules in vivo, have not improved survival. However, extracorporeal strategies such as continuous renal replacement therapy or therapeutic plasmapheresis have shown benefit. We are presenting an affinity-based extracorporeal technology for the removal of ET from whole blood. The small-scale device contains an adsorbent that removed 75% of ET present in whole blood. This affinity resin displayed good hemocompatibility regarding the coagulation pathway. Minimal platelet, neutrophil and complement activation were observed. There was also no evidence of consumption of coagulation factors or cell loss. In as much as ET participates in both the inflammatory and coagulation abnormalities in sepsis, this method represents an efficient and hemocompatible way to remove ET from whole blood, which, in an extracorporeal setting, may improve the outcome of sepsis. PMID- 15291079 TI - Analysis of red blood cell aggregation in cardio-pulmonary bypass (CPB) surgery. AB - Not much is known about red cell aggregation during cardio-pulmonary bypass surgery (CPB). Blood samples from 19 patients undergoing CPB were anticoagulated with EDTA. Hematocrit was adjusted to 40%. A red blood cell aggregometer (LORCA) measured changes in light reflection from each blood sample after cessation of the rotation, and calculated an aggregation index (AI). Reflection measurements were stored. Because LORCA software failed for 87 of 171 samples, we developed new software, and applied it to the stored reflection measurements. This software failed only in 7 out of 171 cases and showed that all LORCA failures occurred for AI < 40%. The new calculations revealed that the aggregation index significantly decreased from 46.6 +/- 10.1 (mean +/- standard deviation) baseline to 22.8 +/- 8.3 at the end of CPB and recovered to 37.1 +/- 13.5 at day 1. It is concluded that the new software can be used to study decreased red cell aggregation during CPB. PMID- 15291080 TI - Controversy over the benefits of pulsatile perfusion. PMID- 15291081 TI - Cascade filtration: a new filter for secondary filtration--a multicentric study. PMID- 15291082 TI - Acute renal controversies--the facts. PMID- 15291083 TI - Preventing HIV--a primary care imperative. PMID- 15291084 TI - How useful is the concept of 'failure to thrive' in care of the aged? PMID- 15291085 TI - Evaluating lymphadenopathy using lymph node FNA. PMID- 15291086 TI - Diagnosis of malaria should be considered by U.S. physicians. PMID- 15291087 TI - Therapy with ACE inhibitors and ARBs in heart failure. PMID- 15291089 TI - HIV infection--how to lower your risk. PMID- 15291088 TI - HIV counseling, testing, and referral. AB - Over the past decade, the annual number of new cases of human immunodeficiency virus (HIV) infection has been relatively stable but remains unacceptably high (an estimated 40,000 new cases per year). Furthermore, the demographics for HIV infection are changing. Rates of new infections are declining in newborns, older men who have sex with men, and whites. However, rates of new infections are rising in young persons, women, Hispanics, and blacks. In 2001, the Centers for Disease Control and Prevention issued revised guidelines for HIV counseling, testing, and referral. The guidelines focus on the reduction of barriers to testing, voluntary routine testing of high-risk populations and persons with risk factors, case management and partner tracing for infected persons, and universal testing of pregnant women. Effective strategies for reducing HIV infection include behavioral interventions, comprehensive school-based HIV and sex education, access to sterile drug equipment, screening of the blood supply, and postexposure prophylaxis for health care workers. PMID- 15291090 TI - Sarcoidosis. AB - Sarcoidosis is a systemic granulomatous disease of unknown cause affecting young and middle-aged adults. Patients commonly present with bilateral hilar lymphadenopathy, pulmonary infiltrates, and ocular and skin lesions. The heart, liver,spleen, salivary glands, muscles,bones, kidneys, and central nervous system also may be involved. Diagnosis is based on clinicoradiologic findings plus histologic evidence of noncaseating epithelioid granulomas, and exclusion of other granulomatous diseases. Prognosis correlates with mode of onset, host characteristics, initial clinical course, and extent of disease. The optimal management of sarcoidosis has not been well defined. Although corticosteroids remain the mainstay of treatment, there is little evidence for the optimal initiation, dosage, or duration of therapy. Topical steroids maybe considered for treatment of anterior uveitis and skin lesions. Systemic steroids are indicated for treatment of cardiac, nervous system, severe ocular, and symptomatic or progressive pulmonary involvement. There is little evidence for the efficacy of inhaled steroids. Cytotoxic agents and immunomodulators usually are reserved for treatment of complex or refractory disease. Of these agents, methotrexate is used more frequently because of its safety profile and possible steroid-sparing effects. Antimalarial agents are used frequently for skin lesions, and they have limited success in the treatment of pulmonary disease. Lung and cardiac transplantation is reserved for end-stage disease. Monitoring for symptoms of drug toxicity is essential, and prevention of osteoporosis must be addressed in patients taking long-term oral corticosteroids. It is not known if current therapy alters disease progression. PMID- 15291091 TI - Diagnosing heel pain in adults. AB - Heel pain is a common condition in adults that may cause significant discomfort and disability. A variety of soft tissue, osseous, and systemic disorders can cause heel pain. Narrowing the differential diagnosis begins with a history and physical examination of the lower extremity to pinpoint the anatomic origin of the heel pain. The most common cause of heel pain in adults is plantar fasciitis. Patients with plantar fasciitis report increased heel pain with their first steps in the morning or when they stand up after prolonged sitting. Tenderness at the calcaneal tuberosity usually is apparent on examination and is increased with passive dorsiflexion of the toes. Tendonitis also may cause heel pain. Achilles tendonitis is associated with posterior heel pain. Bursae adjacent to the Achilles tendon insertion may become inflamed and cause pain. Calcaneal stress fractures are more likely to occur in athletes who participate in sports that require running and jumping. Patients with plantar heel pain accompanied by tingling, burning, or numbness may have tarsal tunnel syndrome. Heel pad atrophy may present with diffuse plantar heel pain, especially in patients who are older and obese. Less common causes of heel pain, which should be considered when symptoms are prolonged or unexplained, include osteomyelitis, bony abnormalities (such as calcaneal stress fracture), or tumor. Heel pain rarely is a presenting symptom in patients with systemic illnesses, but the latter may be a factor in persons with bilateral heel pain, pain in other joints, or known inflammatory arthritis conditions. PMID- 15291092 TI - Geriatric failure to thrive. AB - In elderly patients, failure to thrive describes a state of decline that is multifactorial and may be caused by chronic concurrent diseases and functional impairments. Manifestations of this condition include weight loss, decreased appetite, poor nutrition, and inactivity. Four syndromes are prevalent and predictive of adverse outcomes in patients with failure to thrive: impaired physical function, malnutrition, depression, and cognitive impairment. Initial assessments should include information on physical and psychologic health, functional ability, socioenvironmental factors, and nutrition. Laboratory and radiologic evaluations initially are limited to a complete blood count, chemistry panel, thyroid-stimulating hormone level, urinalysis, and other studies that are appropriate for an individual patient. A medication review should ensure that side effects or drug interactions are not a contributing factor to failure to thrive. The impact of existing chronic diseases should be assessed. Interventions should be directed toward easily treatable causes of failure to thrive, with the goal of maintaining or improving overall functional status. Physicians should recognize the diagnosis of failure to thrive as a key decision point in the care of an elderly person. The diagnosis should prompt discussion of end-of-life care options to prevent needless interventions that may prolong suffering. PMID- 15291093 TI - Screening and behavioral counseling interventions in primary care to reduce alcohol misuse: recommendation statement. PMID- 15291094 TI - Tadalafil (Cialis) for erectile dysfunction. PMID- 15291095 TI - Rash and fever in an ill-appearing child. PMID- 15291096 TI - NAMS releases position statement on the treatment of vasomotor symptoms associated with menopause. AB - The NAMS recommends first considering lifestyle changes, alone or combined with a nonprescription remedy (such as dietary isoflavones, vitamin E, or black cohosh) for the relief of mild vasomotor symptoms. For moderate to severe menopause related hot flashes, prescription systemic estrogen-containing products are still the therapeutic standard. For women with concerns or contraindications to estrogen-containing products, possible treatment options include prescription progestogens, venlafaxine, paroxetine, fluoxetine, or gabapentin. PMID- 15291097 TI - [Preliminary screening of a reverse-subtracted cDNA library for differentially expressed genes in rat liver of prothrombotic state]. AB - OBJECTIVE: To identify differentially expressed cDNA sequences for rat liver of prothrombotic state (PTS) by screening a reverse-subtracted cDNA library. METHODS: Forward-subtracted cDNA probes and reverse-subtracted cDNA probes were prepared by nested PCR amplification, which were labeled with HRP. Positive clones were selected by differential screening in which forward-subtracted and reverse-subtracted cDNA probes were separately hybridized with the membranes slot blotted by plasmid DNAs amplified and isolated from the library. Inserts in the positive clones were submitted to DNA sequencing. Nucleic acid sequence homology search was performed against the GenBank DNA database (non-redundant, and non mouse and non-human EST entries) using the Standard nucleotide-nucleotide BLAST [blastn] program via a network connection to the National Center for Biotechnology information. RESULTS: Five positive clones were found by differential screening. For 3 of the 5 sequences, known genes of high homology (Score>225 bits) in the GenBank DNA database were found. For the other 2 sequences, only genes of low homology (Score=56) or intermediate homology (Score=151) were found. CONCLUSION: Five cDNA sequences were found PTS related. 3 of them may represent the genes that are either identical or nearly identical to previously reported genes The other 2 sequences are most probably new sequences not existing in GenBank DNA database, which have been registered in the GenBank DNA database as new sequences. PMID- 15291098 TI - [Construction of recombinant adenovirus for the angiogenic inhibitor, vasostatin, and its expression in vitro]. AB - OBJECTIVE: To construct a recombinant replication-deficient adenovirus for the angiogenic inhibitor, vasostatin and assay its expression in vitro. METHODS: The cDNA for vasostatin was got by PCR amplification, then it was cloned into T vector and confirmed by enzymatic analysis and direct sequencing. Subsequently the cDNA was subcloned into the shuttle plasmid, and co-transformed into 293 cells with backbone plasmid. The resulting recombinant andenovirus was confirmed by PCR and western blot. The activity was assayed by inhibition of HUVEC growth. RESULTS: The PCR product was about 590 bp in length, and sequencing result was identical to that reported. The construction of recombinant replication-deficient adenovirus was confirmed by PCR. Western blot analysis showed the expression of vasostatin in vitro. The supernatant from transduced HeLa cells inhibited the growth of HUVEC specifically. CONCLUSION: The recombinant adenovirus for vasostatin efficiently mediated the expression of the protein in vitro. PMID- 15291099 TI - [Adipose tissue-derived stromal cells as vector for gene therapy in central nervous system]. AB - OBJECTIVE: To investigate exogenous gene expressing ability of adipose tissue derived stromal cell (ADSCs) and cell distribution after they were transplanted into brains, and to get the genetically modified cells for autografting. METHODS: ADSCs were transfected by Ad5beta gal adenovirus containing a report gene, LacZ gene, then they were transplanted into the adult brain of rats, or ADSCs labeled by Hoechst33258 were transplanted into the adult brain of rats to investigate the migration and distribution of cells. RESULTS: ADSCs showed a good expression of LacZ with X-gal staining after transfecting and transplantation into adult brains, and they could incorporate into the host brain tissues and no disruption was observed. These cells showed good compatibility with the host brains. CONCLUSION: The results indicate that ADSCs could incorporate into host brains and express exogenous gene steadily when they were transplanted into adult brain tissues, no overproliferation and gliosis were identified, and ADSCs may be used as a therapeutic gene delivery vehicle in treating CNS disorders in humans. PMID- 15291100 TI - [The imprinting status and expression of insulin-like growth factor 2 gene in human hepatocellular carcinoma]. AB - OBJECTIVE: To study the imprinting status and expression level of insulin-like growth factor 2 (IGF2) gene in hepatocellular carcinoma and to provide a clue for elucidating the mechanism of carcinogenesis of hepatocellular carcinoma. METHODS: The heterozygote status of IGF2 gene was detected by restriction fragment length polymorphism. The imprinting status and expression level of IGF2 were evaluated by semi-quantitative reverse transcription-polymerase chain reaction. RESULTS: It was found that 10/18 (55.6%) of hepatocellular carcinoma showed the gain of imprinting (GOI), with 6/10 (60%) adjacent cirrhosis of liver tissues also displaying GOI of IGF2. Overexpression of IGF2 in cancer tissues was detected in 9/18 (50%) samples, but no significant difference was observed among each imprinting status. CONCLUSION: GOI of IGF2 may take part in human hepatocellular carcinogenesis. PMID- 15291101 TI - [Studies on the nucleotides sequences of extended-spectrum-beta-lactamases encoding genes of Escherichia coli and Klebsiella pneumoniae and the related molecular evolution]. AB - OBJECTIVE: To identify TEM-type and SHV-type ESBLs encoding genes of ESBLs producing Klebsiella pneumoniae and Escherichia coli isolated from clinical species in West China Hospital of Sichuan University and study the molecular evolution of the ESBLs. METHODS: The nucleotide sequences of TEM-type and SHV type ESBLs encoding genes amplified by PCR were detected by automatic sequencer, and the subtypes of the encoding genes were determined by Blastx searching. The molecular evolution of ESBLs was studied by means of bioinformatics. RESULTS: In this study, the subtypes of ESBLs were SHV-2 and TEM-19, the distribution of silent mutation in ten bla(SHV-2) was identical, and that of two bla(TEM-19) was the same; the distribution of silent mutation of bla(TEM-19) was the same as that of bla(TEM-1). The distribution of silent mutation of bla(SHV-2) observed here was different from that observed in other countries. CONCLUSION: SHV-2 was the main ESBLs in this study. The bla(SHV-2) and bla(TEM-19) in this study originated from the same transferable variants respectively. The prevalent SHV-2 in different countries resulted from convergent evolution. It seems possible that the bla(TEM-19) identified by this study might originate directly from the transferable bla(TEM-1) identified in this study. PMID- 15291102 TI - [Genetic polymorphisms of three STR loci on chromosome X and their forensic application in a Chinese Han population]. AB - OBJECTIVE: To understand the allele structure and genetic polymorphism at three STR loci on chromosome X in Chinese Han population and make an evalution of their forensic application. METHODS: EDTA-blood samples were collected from the unrelated individuals in Chengdu, China. After being extracted with Chelex method, the DNA samples were amplified by PCR technique. The PCR products were analyzed by PAG electrophoresis and the approach of automated fluorescence detection. Hardy-Weinberg equilibrium of females was tested and every forensic interested value was calculated. RESULTS: The polymorphisms of all 3 STR loci were obtained from 100 unrelated females and 120 unrelated males from Chinese Han ethnic group. Chi-square tests on the genotype frequencies in females did not reveal deviations from Hardy-Weinberg equilibrium. CONCLUSION: The obtained data are beneficial to understanding the population genetics of the three STR loci in Chinese Han population. For forensic genetics, the obtained data can be used to calculate the probabilities dealing with the paternity test and the individual identification. PMID- 15291103 TI - [The expression of ALK protein and Epstein-Barr virus gene products in anaplastic large cell lymphoma and its significance]. AB - OBJECTIVE: To investigate the characteristic of ALK protein expression in anaplastic large cell lymphoma (ALCL) and the relationship between ALCL and Epstein-Barr virus (EBV). METHODS: Histopathological method was used to study the morphological characteristic. Immunohistochemical staining was used to detect ALK protein and other antigen. EBV-encoded EBER was detected by in situ hybridization. RESULTS: All 2 cases of primary cutaneous ALCL were ALK protein negative. 20 cases of systemic ALCL were found to be ALK protein positive in 12 cases (60%), which included 8 cases of common variant, 3 cases of lymphohistiocytic variant and 1 case of small cell variant; the expression of ALK protein showed no significant difference between these 3 variants (P>0.05). The ALK protein expression pattern was characterized by the brown-stained nucleus and cytoplasm of the cells, but in 2 cases only the cytoplasm was stained. The patients with ALK protein expression were younger than those without ALK protein expression (P<0.05); all 22 cases were EBER negative. CONCLUSION: High incidence of the expression of ALK protein was seen in ALCL, especially in ALCL of younger patients. No relationship was observed between EBV and ALCL in this study. PMID- 15291104 TI - [Expression and variation of angiotensin II type 1 receptor on vascular smooth muscle cells of systemic and pulmonary circulations in experimental ventricular septal defect in pigs]. AB - OBJECTIVE: To investigate the expression of angiotensin II type 1 receptor (AT1R) on the vascular smooth muscle cells (VSMC) of both systemic and pulmonary circulations and their variation caused by intracardiac left to right shunting using an animal model of the ventricular septal defect (VSD). METHODS: Nineteen young pigs were divided into 3 groups: operation (experimental VSD), sham operation, and normal control. One month after operation, the pigs were catheterized and then put to death. Smooth muscle cells were taken from aorta, pulmonary artery, pulmonary arteriole, mesenteric arteriole and VSMC were isolated. Radioligand binding assay for AT1R was done to measure the Bmax and KD. RESULTS: There were no significant differences in Bmax and KD of AT1R between the sham-operation group and the control. Bmax of the aorta (112.11+/-35.77) fmol/10(6), main pulmonary artery (52.37+/-31.09) fmol/10(6) and mesenteric arteriole (106.98+/-100.48) fmol/10(6) in the operation group were remarkably elevated in comparison with the control (P<0.05). In the operation group, Bmax of the aorta VSMC was higher than that of main pulmonary artery VSMC (P<0.05), and Bmax of the mesenteric arteriole VSMC was higher than that of the small pulmonary arteriole VSMC (P<0.05). CONCLUSION: The findings in AT1R expression in experimental VSD were supportive to the previously proposed hypothesis that ATR expression might be stronger in systemic VSMC than in pulmonary VSMC in young animal, especially in the presence of intracardiac left to right shunt, which may provide a sound rationale for the pharmacological management of VSD infants with angiotensin converting enzyme inhibitor or AT1R antagonist. PMID- 15291105 TI - [Immunohistochemical detection and clinical significance of beta-catenin expression in human esophageal cancers]. AB - OBJECTIVE: To determine whether the expression level of beta-catenin correlates with the invasion and metastasis of human esophageal cancers. METHODS: Using immunohistochemical staining technique, we investigated the expression of beta catenin in different tissues from the specimens of the primary tumor and the nearby non-tumorous esophageal mucosa. RESULTS: Strong expression of beta-catenin in the nearby non-tumorous esophageal mucosa was observed. However, in the primary esophageal cancers, the expression of beta-catenin was frequently reduced and disorderly distributed. The level of beta-catenin expression was closely related with the growth pattern, cellular differentiation, depth of invasion, and lymphatic metastasis. CONCLUSION: The down-regulation of beta-catenin expression might be a predictor indicating the metastatic potential of esophageal cancers. PMID- 15291106 TI - [Expression of heat shock protein-70, estrogen receptor and progesterone receptor in ovarian carcinomas and the correlation between HSP70 and sex steroid receptor]. AB - OBJECTIVE: This study was aimed to determine the expression of HSP70, ER and PR in ovarian carcinomas and to explore the relationship between HSP70 and sex steroid receptor. METHODS: The immunohistochemical way SP was performed to estimate the expression of HSP70, ER and PR in 41 cases of ovarian carcinomas and in 11 cases of normal ovarian tissue. RESULTS: The positive staining rate of HSP70 was 68.29% (28/41), which was remarkably higher than that in normal ovarian tissue (18.18%) (P<0.05). Furthermore, the expression rate of HSP70 was much higher in poorly differentiated ovarian carcinomas than in well differentiated ovarian carcinomas (P<0.05). ER positive staining was observed in 19 cases (46.34%), and PR in 24 cases (58.54%). ER and PR positive staining occurred more frequently in the group of HSP70 negative staining than in the group of HSP70 positive staining. related with the expression of PR (P<0.05). CONCLUSION: The expression of HSP70 was negatively related with the expression of PR (P<0.05). PMID- 15291108 TI - [Expression of HSP70 induced by ketamine in the hippocampus of rat at different ages]. AB - OBJECTIVE: This experiment was designed to explore the possible mechanism of the brain injury caused by ketamine. METHODS: Thirty-five adult SD rats randomly divided into a control group and 6 experiment groups were given intrapertioneal injections of normal saline, 20.0, 40.0, 60.0, 80.0, 100.0, 120.0 mg/kg of ketamine respectively. Another 35 SD rats at 0-10, 11-20, 21-30, 31-45, 46-60, 61 90, 91-120 days old were allocated to 7 corresponding age groups and were given intrapertioneal injection of ketamine 80.0 mg/kg. Immunocytochemistry was used to detect HSP70 expression in rat hippocampus, and MIAS-2000 photography analytic software was chosen to analyze HSP70 expression. RESULTS: Ketamine induced HSP70 expression in adult rat hippocampus. Below the dose of 80.0 mg/kg, ketamine induced higher level of HSP70 expression as the dose increased; however, above the dose of 80.0 mg/kg, ketamine induced lower level of HSP70 expression as the dose increased. Ketamine did not induce HSP70 expression in the rats under 20 days of age; it induced the same HSP70 expression in rats of 90 days and over; and analyses of the rats of 20-90 days revealed that the elder the rat is, the higher the HSP70 expression will be. CONCLUSION: Ketamine injures the neurons of rat hippocampus and induces the expression of HSP70, and the higher the dose is given, the more serious the injury is seen; and ketamine induces higher level of HSP70 expression in adult rats than in infant rats. PMID- 15291107 TI - [Gene expression of heparanase in peripheral blood leukocytes from rats with nephrotic syndrome and its relationship to proteinuria]. AB - OBJECTIVE: The beta-D-endoglycosidase heparanase (Hpa) is HS-specific which leads to heparan sulfate (HS) degradation. An increased permeability of the glomerular basement membrane (GBM) for proteinuria is related with a decreased HS side chains of the GBM. However, whether an up-regulated expression of Hpa exists in nephrotic syndrome (NS) remains unknown. The purpose of this study was to evaluate the role of Hpa in the development of NS and the relevance to proteinuria. METHODS: NS was induced in SD rats by tail intravenous injection of 5 mg/kg Adriamycin. Rats were housed in the metabolic cages for 24 hours to collect urine prior to induction of disease and at day 7 and 14 following disease induction. By using the method of reverse transcriptase-PCR (RT-PCR), Hpa gene from the peripheral blood leukocytes of these rats was assayed. RESULTS: The resulting cDNA fragment was consistent with rat heparanase mRNA sequence in GenBank (accession number AF184963). The rats at day 7 and 14 showed significantly up-regulated Hpa mRNA expression in contrast to the normal rats. There was a significant correlation between the level of Hpa mRNA and the quantification of urinary protein (r=0.925, P<0.001). CONCLUSION: Up-regulated expression of Hpa mRNA may be an important contributor to loss of glomerular negative charge in GBM and hence may lead to proteinuria in nephrotic syndrome. PMID- 15291109 TI - [Heterogeneity of Na+/Ca2+ exchanger current across the left ventricular wall of rabbit]. AB - OBJECTIVE: To investigate the distribution characteristics of Na+/Ca2+ exchanger current (I(Na+/Ca2+)) across the left ventricular wall of rabbit and the relationship between I(Na+/Ca2+) and transmural depolarization heterogeneity. METHODS: By using whole-cell patch clamp techniques, action potentials (AP), I(Na+/Ca2+), and both rapid and slow components of delayed rectifier potassium current (I(Kr) and I(Ka)) were recorded in subendocardial (Endo), midmyocardial (M), and subepicardial (Epi) cells of the left ventricular wall of rabbit. RESULTS: AP duration in M cells was longer than that in Epi cells, P<0.01. At the test potential of +40 mV, outward I(Na+/Ca2+) in M cells was larger than that in Epi and Endo cells, P<0.01, P<0.05, respectively. At the test potential of -100 mV, inward I(Na+/Ca2+) in M cells was larger than that in Epi cells, P<0.05. At the test potential of +50 mV, the tail current density of I(Ka) in M cells was smaller than that in Epi cells, P<0.05, and there was no significant difference among the tail current densities of I(Kr) in Endo, M, and Epi cells. CONCLUSION: The distribution of I(Na+/Ca2+) and I(Ka) across the left ventricular wall of rabbit is unequal, which contributes to the transmural depolarization heterogeneity. PMID- 15291110 TI - [Relationship between the contents of serum HDL subclasses and the extent of coronary stenosis in coronary heart disease patients]. AB - OBJECTIVE: To explore the relationship between the contents of serum HDL subclasses and the extent of coronary stenosis in coronary heart disease (CHD) patients. METHODS: The contents of serum HDL subclasses in CHD patients (n = 51) and healthy controls (n = 56) were determined by two dimensional gel electrophoresis associated with immunodetection method. CHD patients were divided into three groups by average coronary severity score (CSS). Data were analyzed using linear regression and correlation method and multiple stepwise regression method. RESULTS: It was found that as the extent of coronary stenosis increases, the levels of pre beta1-HDL, HDL3b increase, meanwhile the level of HDL2b decreases (P < 0.001). There were significant positive correlations between CSS and pre beta1-HDL (P < 0.01), HDL3a (P < 0.05), HDL3b (P < 0.01), and significant negative correlation was observed between CSS and HDL2b (P < 0.01). apoB100, HDL2b, TG, HDL-C, TC/HDL-C, apoAI are all independent risk factors of CHD. CONCLUSION: The extent of coronary stenosis is highly correlated with the subclasses of HDL. PMID- 15291111 TI - [Function of actin in renal tubular epithelial cell of newborn swine during ATP deficiency]. AB - OBJECTIVE: To investigate the variations of actin of newborn porcine Renal Tubalar Epithelial (RTE) cell during ATP deficiency and shed light on the possible mechanisms of renal deficiency during newborn asphyxia. METHODS: After the establishment of ATP deficiency cell-model, the ATP content of the RTE cell of the control and hypoxic groups was determined. And the variations of actin in newborn porcine RTE cell during ATP deficiency were detected using flow cytometry. RESULTS: It was found that the ATP content of new-born porcine RTE cell decreased with the continuance of hypoxia, and the changes in G-actin and F actin contents of RTE cell both appeared at the time of one-minute ATP deficiency. The G-actin decreased first and then increased, and the F-actin decreased step by step. CONCLUSION: As the ATP deficiency time elongated, G-actin of the newborn porcine RTE cell decreased first and then increased, and the F actin decreased step by step. This may destruct the cell bone-skeleton of the newborn RTE cell and maybe one of the important mechanisms of renal deficiency during newborn asphyxia. PMID- 15291112 TI - [Epstein-Barr virus infection in lymphoepithelial carcinoma of salivary glands in Sichuan Chinese]. AB - OBJECTIVE: To determine the prevalence of Epstein-Barr virus (EBV) infection in lymphoepithelial carcinoma (LEC) of salivary glands in Sichuan Chinese and discuss the role of EBV infection in pathogenesis of LEC. METHODS: Paraffin sections of 16 cases of LEC and 4 cases of benign lymphoepithelial lesion (BLEL) of salivary glands, 4 cases of non-specific chronic sialadenitis were examined by in situ hybridization for EBV encoded small RNA-1 (EBER-1) using 30-base synthesized oligonucleotide probe. RESULTS: Strong signals for EBER-1 were obtained in most of tumor cell nuclei in all the specimens of LEC (16/16); in contrast, there were no significant signals observed in the examination of 4 cases of BLEL and 4 cases of non-specific chronic sialadenitis. CONCLUSION: The results indicate that EBV plays an important role in the pathogenesis of LEC of salivary glands in Sichuan Chinese. PMID- 15291113 TI - [Effects of nandrolone phenylpropionate on fibroblasts after injury in rats]. AB - OBJECTIVE: To explore the effects of nandrolone phenylpropionate (NP) on fibroblasts after injury in rats. METHODS: Thirty-two Wistar rats with a deep second-degree scald injury and 20% total body surface area were randomly divided into two groups to receive either 5 mg/kg NP (NP group) or normal saline as placebo (control group) every other day. Integrated optical density (IOD) of androgen receptor (AR) and mRNA expression level of alpha1 (I) procollagen were measured by immunohistochemistry and quantitive fluorescent RT-PCR individually on the post-burn days 4, 7, 14 and 21. Fibroblasts were isolated from granulation tissue of a Wistar rat and were cultured in RPMI1640 with the addition of NP at different concentrations. Cell viability of fibroblasts was measured by MTT test, and the proliferative index, by flow cytometry (FCM). RESULTS: (1) Compared with the A value of the concurrent control group, higher A values were seen in NP groups containing different concentrations of NP, and significant difference of proliferative index was observed in NP group, P<0.05. (2) The expression of alpha1 (I) procollagen mRNA in NP groups was much higher than that in control groups. A significant difference between the two groups was noted on the post burn days 7, 14 and 21, P<0.05, but no difference was seen on day 4. (3) The density of AR on fibroblasts in NP group was higher than that in control group at each time point. And a positive correlation between the expression of alpha1 (I) procollagen mRNA and the quantity of AR on fibroblasats was confirmed (r = 0.836). CONCLUSION: Nandrolone phenylpropionate could promote fibroblast replication, increase the mRNA level of alpha1 (I) procollagen and enhance the density of AR on fibroblasts. PMID- 15291114 TI - [Histopathologic and immunohistochemical studies on retina after laser photocoagulation]. AB - OBJECTIVE: To observe the histopathologic and immunohistochemical changes and reparable process of retina after laser photocoagulation in order to provide a theoretical basis of laser treatment for fudus diseases. METHODS: Normal retina was photocoagulated using I-, II-, III-grade photic spot after the signing of informed consent of 7 patients who were suffering from orbital malignant tumors and in need of exenteration. The retinae from removed eyeballs were fixed with formalin, imbedded in paraffin and stained by H. E. The SP immunohistochemical method was adopted using antibodies of S-100, GFAP, NSE,and NF label. RESULTS: By histopathologyical examination, 1 day after photocoagulation with I-grade photic spot, the retinal pigment epithelium (RPE) cells, photoreceptors and cells in outer nuclear layer were edematous and necrotic, and decreased in number; with II grade photic spot, many RPE cells, photoreceptors and cells in outer nuclear layer were lysed and destroyed; with III-grade photic spot, all layers of the retina were severely damaged. 3 days after photocoagulation, retinal edema was relieved. RPE and glial cells started proliferating. 7 days after photocoagulation, RPE and glial cells proliferated obviously to cover the destroyed areas. By immunohistochemistry, 3-7 days after photocoagulation, S-100 and GFAP were seen stained in the damage areas of photic spot. 1-7 days after photocoagulation, NSE and NF were found unstained in the photic spot. CONCLUSION: The above data indicated that different laser photic spots resulted in various degrees of damage to the retina. The destroyed regions were repaired by proliferative RPE and glial cells. Immunohistochemistry confirmed that proliferative gial cells repaired damaged retina, and neurons and nerve fibers could not regenerate. Laser could treat different retinal diseases. PMID- 15291115 TI - [Stress distribution of mandible under different loading and biting condition]. AB - OBJECTIVE: To study the stress distribution of intact mandible, especially that of the mandibular angle under different muscle loading and biting condition. METHODS: Develop a more accurate, more objective mandibular model, measure the different stress patterns on the outer surface of the mandible by strain gauges due to different biting sites (INC and ICP) and different loading methods (masseter, temporalis and four pairs of muscles). RESULTS: It was found that the strain in the zone of mandibular angle is more markedly under masseter loading; that the strain in the zone of anterior mandibular ramus is more markedly under temporalis loading; and that the stress in the zone of mentum becomes a tension because of the medial pterygoid under the load of four pairs of muscles. During anterior teeth biting, the stress of mandible angle is larger than that of the bilateral molar biting. When an occlusal load is on the ipsilateral molars, there is a reversal result of the stress direction in the upper line of the mandibular angle. CONCLUSION: Different muscular loading and biting condition can change the stress distribution of the mandible. It is important to develop a functional model of human mandible to study its complex biomechanics behavior. PMID- 15291116 TI - [Evaluation of the in vitro cariogenic potential of Streptococcus mutans (serotype C) strains isolated from caries-free and -active people: the ability of acidogenicity]. AB - OBJECTIVE: To investigate the ability of acidogenicity of S. mutans (serotype C) strains isolated from the people with different caries experience. METHODS: Same density solutions of all isolated S. mutans were made and cultured in different pH TPPE liquid for the same period of time. Terminal pH of the solutions was measured. The values of delta pH were compared. RESULTS: Significant difference of acidogenicity was observed between the strains of different genotypes isolated from the same person. The ratio of high acidogenicity isolates harbored in caries active people was greater than that of caries-free people; a significant difference was noted (P<0.05). CONCLUSION: The high cariogenicity of isolated S. mutans strains of caries-active people shows a close relationship with the high acidogenicity of the isolated S. mutans (serotype C) strains. PMID- 15291117 TI - [Animal test of nHA-PA66 used in root canal filling]. AB - OBJECTIVE: To assess the dogs' tissue reaction to nHA-PA66 which is prepared to be used as a new root canal filling sealer. METHODS: The experimental apical periodontitis of dogs was induced first, then nHA-PA66 was used in the test groups while CCQ was used in the control group. The reactions of periapical tissues were measured by histological means. RESULTS: One month after nHA-PA66 was used in root canal filling, the dogs' periapical tissues showed mild to moderate inflammatory reactions, the X-ray films showed decreased radiolucent area as compared with the previous film. Three months later, tissue repairs occurred, which exhibited even more decreased radiolucent area on X-ray films. nHA-PA66 caused weak inflammatory reaction while the reparative reaction happened early. CONCLUSION: nHA-PA66 has good tissue compatibility and is a potential sealer for root filling in clinical use. PMID- 15291118 TI - [Impact of traditional Chinese medicine WPY on ICAM-1 expression and MPO activity in pancreas and lungs of rats with acute necrotizing pancreatitis]. AB - OBJECTIVE: To explore the character of the ICAM-1 induced change of polymorphonuclear neutrophil (PMN) in the pancreas and lung of rats with acute necrotizing pancreatitis (ANP) and to evaluate the effects of Chinese medicine WPY on ANP. METHODS: ANP model was induced by retrograde injection of 3.5% sodium taurocholic acid into the biliopancreatic duct of Wistar rats. Expression of ICAM 1 at different time points in pancreas and lung was assessed by immunohistochemistry. The quantity of polymorphonuclear neutrophil (PMN) in tissues was measured by the level of myeloperoxidase activity (MPO). In addition, the indices above were also observed after the administration of WPY. RESULTS: In the ANP group, ICAM-1 expression and MPO activity were evidently up-regulated at 3 h in pancreas and at 6 h in lung, and both reached their highest level at 12 h. In the WPY treated group, ICAM-1 expression and MPO activity in pancreas and lung were significantly decreased at 12 h. CONCLUSION: The numerous PMNs infiltration mediated by ICAM-1 plays an important role in the damage to pancreas and lung caused by ANP. The traditional Chinese medicine WPY can attenuate ICAM-1 expression and MPO activity in pancreas and lung during ANP. PMID- 15291119 TI - [Therapeutic effect of "anti-hepatic-fibrosis 268" on hepatic fibrosis in rats]. AB - OBJECTIVE: To investigate the effects of the proprietary Chinese Medicine "anti hepatic-fibrosis 268" on hepatic fibrosis and the related mechanisms. METHODS: The model of CCl4-induced hepatic damage was established in SD rats. 54 male rats were divided into four groups, namely high dose and low dose "anti-hepatic fibrosis 268" groups, colchicine control group, and model control group. Using Masson stain and light microscope, the authors examined the rats' hepatic tissues and counted the hepatic fiber components, then examined and counted TGF-beta1, alpha-SMA, FN, Type I, III collagen by means of immunohistochemical technique. The groups were compared and the internal relationships of the data were analyzed. RESULTS: The levels of FN, LN, Type I and III collagen, TGF-beta1, and alpha-SMA of the CCl4 damaged rats increased (P<0.01). After 3 weeks of high dose "anti-hepatic-fibrosis 268" treatment, the levels of TGF-beta1, alpha-SMA, FN, LN, Type I and III collagen decreased (P<0.01) and the degree hepatic fibrosis took a favorable turn significantly (P<0.05) as compared with the model control. In the rats of the low dose group, the levels of TGF-beta1, alpha-SMA, FN, Type III collagen significantly decreased (P<0.05), the levels of LN, Type I collagen were not different from the model control; The hepatic fibrosis improved to a certain extent (P<0.05). CONCLUSION: The mechanism of reversing hepatic fibrosis by "anti-hepatic-fibrosis 268" in this experiment is that the medicine regulates TGF-beta1 and further affects alpha-SMA, thus resulting in the decline of FN, Type I, III collagen levels in liver extracellular matrix. PMID- 15291120 TI - [Pathological study of the therapeutic effect of wen-yang herbs on experimental liver cirrhosis]. AB - OBJECTIVE: To observe the pathological effect of wen-yang herbs on experimental hepatic fibrosis. METHODS: Thirty-two male Wistar rats were used in this study comprising four groups. To start with, 24 rats of three groups were given subcutaneous injection of CCl4 and drinking 10% alcohol so as to make the model of hepatic fibrosis. After the establishment of the pathologic model, the rats were divided into the model group, the pathological control group and the therapeutic group by randomization. The rats of the therapeutic group were given the herbal remedies via gastrogavage, q.d. x 30. The rats of the pathological control group were given normal saline via gastrogavage, q.d. x 30. Then liver tissue Hydroxyproline (Hyp) content was examined in these 3 groups and the normal group. Quantitative marks were done according to a modified semiquantitative scoring system (SSS). The results of SSS marks and Hyp contents were analysed using Pearson's coefficient of correlation. RESULTS: The Hyp content and SSS marks of the therapeutic group decreased remarkably as compared with those of the control group (P<0.01), and the SSS marks had a strong positive correlation with Hyp content (r=0.804). CONCLUSION: Wen-yang herbs can mitigate the rats' hepatic fibrosis and promote a recovery of their experimental illness. PMID- 15291121 TI - [The reliability and validity of adult attachment scale (AAS-1996 revised edition): a report on its application in China]. AB - OBJECTIVE: AAS is an adult attachment scale revised by Collins in 1996. This paper mainly reports on its use in our mental health center and on an evaluation of its internal consistency reliability, discriminatory validity and construct related validity. METHODS: The patients group (n=89) and the normal group (n=110) were studied with the use of AAS. RESULTS: Cronbach's alpha coefficients in normal group are higher than 0.7. The discriminatory validity in anxiety and close-dependence dimension is good. The score of construct-related validity is high. CONCLUSION: The results are consistent with the theoretic construct that AAS divides adult attachment into three factors. But this scale still needs revision of its one or two items on account of cultural difference. The application of AAS will accelerate the research into the relationship between adult attachment style and adult marriage, consanguinity. PMID- 15291122 TI - [An evaluation of effects of intervention on maternal and child health in the rural areas of China]. AB - OBJECTIVE: To evaluate the impact of training maternal and child health care providers in the rural areas of China on improvement of health care to pregnant and puerperal women. METHODS: The data originated from the Reproductive Health/Family Planning Project implemented by the State Family Planning Commission and the Ministry of Health from 1998 to 2002, which covered 32 counties in 22 provinces of China. A quasi-experimental design was used. 6 counties were selected from 32 project counties as the intervention group, while 6 non-project counties were taken as the control group with the condition similar to that of the selected project counties in respect to their number of population and economic level. The subjects of the study were mothers with child under 3 years. A total of 348 mothers were interviewed using a structured questionnaire by strictly trained surveyors. It was focused on prenatal care and postpartum follow-up in the survey. INTERVENTION: According to the plan of the project, all maternal and child health care providers at the grass-root level were given a 2 week theoretical training, and some of them were assigned to hospitals where they were gievn a 1-month clinical skills training. RESULTS: With regard to prenatal care, the mothers in intervention group received more prenatal care than those in control group (mean number of obstetric visits: 6.64 vs 5.64, P<0.05). The number of items of examination taken in intervention group was more than that in control group (6.71 versus 5.67, P<0.05). The proportion of the mothers in intervention group who were told that they must visit doctors if they felt uncomfortable in pregnant period, was higher than that in control group (P<0.05). 8 symptoms or signs that possibly occur in pregnant period were listed; in this connection, the mothers in intervention group knew more than those in control group (3.43 vs 2.09, P<0.05). In the postpartum follow-up, more mothers in the intervention group were examined by the doctors. The proportion of mothers who were informed of contraceptive methods was higher in intervention group than in control group (94% vs 78.5%, P<0.05). The descending rate of maternal mortality rate in the intervention areas was much higher than that in the control areas. CONCLUSION: The training of maternal and child health care providers had a significant impact on improving their service skills and quality; consequently, the women covered by their service could receive better maternal and child health care. This indicates that the Reproductive Health/Family Planning Project implemented in the rural areas of China is successful. PMID- 15291123 TI - [Study on the influence of medical abortion and surgical abortion on subsequent pregnancy]. AB - OBJECTIVE: To evaluate and compare medical abortion versus surgical abortion in respect of their influence on the safety of the mother and baby in the subsequent pregnancy and parturition. METHODS: Based on the principle of informed consent of subjects, 150 healthy pregnant women with a past history of having experienced medical abortion once were included in the study group (also called medical abortion group), and in the same period, 150 healthy pregnant women with a past history of having experienced surgical abortion once were enrolled into the comparison group (also called surgical abortion group). From then on, all the pregnant women in the two groups were followed up till a week after labor. The baseline data of the two groups were comparable (P>0.05). The rates of complications observed in these women during pregnancy and labor were evaluated. RESULTS: The incidence rates of miscarriage, placental abnormality, premature delivery and postpartum hemorrhage in the study group were significantly lower than those in the comparison group (P<0.05). No significant differences on other variables were observed between the two groups. CONCLUSION: Medical abortion is probably safer than surgical abortion in respect of their influence on subsequent pregnancy. So, provided there is less contraindication, medical abortion may be the choice for terminating unwanted pregnancy, especially for those women without a child. PMID- 15291124 TI - [Randomized controlled trial of hydrochlorothiazide in augmenting the dose of 131I absorbed by thyroid remnant]. AB - OBJECTIVE: Based on the rationale that the ablation of thyroid remnant can effectively reduce the risk for recurrence of differentiated thyroid carcinoma (DTC) and hence decrease the case fatality rate. This randomized controlled trial was designed to assess the value of hydrochlorothiazide in the ablation of thyroid remnant with 131I. METHODS: Thirty consecutive DTC patients with thyroid remnant after thyroid surgery were divided into two groups by randomization, the hydrochlorothiazide group received hydrochlorothiazide 25 mg tid for 4 days, the control group received placebo. Responses to treatment were evaluated by the increment of thyroid 131I uptake rate at 24 h and the augmentation of 131I absorbed dose. RESULTS: In the hydrochlorothiazide group, the 24 h 131I uptake rate was about (1.36+/-0.58) times larger than that before treatment, the absorbed dose was about (1.35+/-0.54) times larger than that before treatment. And in comparison with the control group, the 24 h 131I uptake rate of the hydrochlorothiazide group was significantly increased and the 131I absorbed dose was significantly augmented. CONCLUSION: Hydrochlorothiazide is effective for increasing 24 h 131I uptake rate and augmenting 131I absorbed dose of thyroid remnant. PMID- 15291125 TI - [The influence of abnormally higher WBC on hemoglobin determination and its redress]. AB - OBJECTIVE: To discuss the influence of abnormally higher white blood cell (WBC) on hemoglobin (Hb) determination and work for its redress. METHODS: High WBC concentration suspensions of 48 concentration gradients were prepared to be determined in the pre-dilution pattern of automated hematology analyzer while the changes of Hb concentration before and after removal of WBC being observed. At the same time, a regression equation was set up to correct errors and redress the Hb concentration. Then, the data from 14 whole blood samples with abnormally higher WBC were used in this regression equation to verify its utility. RESULTS: The Hb concentration of samples with abnormally higher WBC was higher than that of samples where WBC was removed (P<0.001). While WBC count > or = 40.0 x 10(9)/L, there was logarithmic linear correlation between Hb concentration difference and WBC count (r=0.9526). Using the regression equation Y (Hb difference) = -25.09 + 21.89 x lg[WBC (x 10(9)/L)], we found that there was no significant difference (P>0.05) between the "practically detected Hb difference" and the "theoretically adjusted Hb difference" in the whole blood samples with abnormally higher WBC. CONCLUSION: Abnormally higher WBC may lead to increase of error in the detection of Hb concentration; the ultimate result in close proximity to true Hb concentration of whole blood samples with abnormally higher WBC can be acquired through the linear regression equation. PMID- 15291126 TI - [Synthesis of N1-(aryl)alkyloxyacyl-5-fluorouracil derivatives]. AB - OBJECTIVE: To synthesize N1-(aryl)alkyloxyacyl-5-fluorouracil derivatives and investigate their antitumor activities. METHODS: Title compounds (I) were prepared by 5-fluorouracil which was trimethylsilylated by hexamethyl-di silazane, and then acylated. Hydrogenization of compounds (I) brought on compounds (II), which were esterified to produce title compounds (III) afterwards. RESULTS: Twelve title compounds were synthesized. CONCLUSION: These compounds were designed products confirmed by 1H-NMR, IR and MS spectral data. PMID- 15291127 TI - [Purification and characterization of sanguicin--a bacteriocin produced by Streptococcus sanguis]. AB - OBJECTIVE: Streptococcus sanguis plays an important Role in maintaining the periodontal microecological balance. Previous studies have demonstrated that sanguicin--a kind of bacteriocin produced by S. sanguis has a prominent function of inhibiting the growth of putative periodontopathic bacteria (PPB). The aim of this study was to purify sanguicin and investigate its characters. METHODS: The raw extract of sanguicin was obtained from cells of S. sanguis by ultrasonication, salting out and dialysis. Then the diethyl-aminoethyl-sepharose gradual washing was done in conjunction with glucosan gel filtration for in the purfication of sanguicin. The characters of purified sanguicin were determined and its inhibitiory effect on PPB was measured by agar diffusion test. RESULTS: The purifed sanguicin was obtained; it was heat labile and proteinaceous; and by SDS-PAGE, it showed a main band with the relative molecular mass of 65 x 10(3). The purifed sanguicin had a strong inhibitory effect on P. gingivalis, P. intermedia and F. nucleatum in vitro. CONCLUSION: Our methods are useful for the purification of sanguicin. Sanguicin has inhibitory effect on PPB obviously and will have a good prospect in periodontal ecological therapy. PMID- 15291128 TI - [Optimization of the separation conditions of fingerprint for Rhubarb by high performance liquid chromatography]. AB - OBJECTIVE: To optimize the separation conditions of fingerprint for Rhubarb by high performance liquid chromatography and lay the foundation of studies on its fingerprint by high performance liquid chromatography. METHODS: The reversed phase high performance liquid chromatographic method was adopted. The samples were extracted by ultrasound and separated by a linear gradient elution on Shim pack CLC-ODS column (6.0 mm i.d. x 150 mm, 5 microm), and then, with the methanol 1 ml/L phosphoric acid solution as mobile phase, were detected at UV 254 nm and column temperature 40 degrees C. During the linear gradient elution program, the volume fraction of methanol in mobile phase changed as follows: 0 min-5 min, 20%; 5 min-25 min, from 20% to 50%; 25 min-35 min, from 50% to 80%; 35 min-60 min, from 80% to 90%; 60 min-70 min, 90%. The flow rate was 0.8 ml/min from 0 min to 35 min, 0.6 ml/min from 35 min to 50 min and 1.0 ml/min after 50 min. RESULTS: The separation conditions described above achieved good result in the aspects of separable effect, stability, sensitivity, and reproducibility. CONCLUSION: The optimum separation conditions of fingerprint would be useful for establishing the method of fingerprint for Rhubarb by high performance liquid chromatography. PMID- 15291129 TI - [Application of M-RAPD technique to obtain the genomic fingerprints of various pathogenic microbials]. AB - OBJECTIVE: To obtain genomic fingerprints of different pathogenic microbials and make certain whether their patterns can be used in the identification of microbials by means of multiplex random amplified polymorphic DNA analysis (M RAPD). METHODS: Arbitrary primers of 10 oligonucleotides were randomly grouped, and various microbials chromosomal DNA were amplified with three combinatorial primers at a special higher annealing temperature. The products were detected by 15 g/L agarose electrophoresis and the patterns were analyzed by the software of Gelworks 1d Intermediate. RESULTS: Specific and resistant DNA fingerprints for different pathogenic microbials with combinatorial primers were gained. The profiles were clear, well-distributed and the number was great. The products of the three primers included most products of every two primers and would appear with no relation to their length, but small products had more opportunity; three primers could provide information contents half as many again as that two primers could provide for the same pathogenic microbials. There were obvious differences among different drug-resistant strains and between the drug-resistant strains and the corresponding reference strains, but different strains of the same microorganism had more similarity than discrepancy. The analytic data of the software of Gelworks 1d Intermediate also support our results. CONCLUSION: M-RAPD is a simple and rapid technique for the identification of different kinds of pathogenic microbials, and it can provide rich genetic information. PMID- 15291130 TI - Baumol's disease. PMID- 15291131 TI - Physician countersuits in medical malpractice cases. AB - The KMA House of Delegates passed Resolution 2003-35 in which the KMA was directed to draft and publish legal guidelines regarding when a suit may be brought against an attorney or plaintiff for filing a frivolous lawsuit against a physician. Thefollowing material, prepared by Peggy Appenfelder of the lawfirm of Stites & Harbison, discusses that issue. PMID- 15291132 TI - Muhlenberg vascular project: a rural vascular intervention project. PMID- 15291133 TI - Residency training in public psychiatry: a review of literature. AB - BACKGROUND: Training of psychiatry residents in a conmmunity setting has been emerging as a more rational and reasonable choice for psychiatry residency training, considering the shift of reliance in the recent years for psychiatric services from inpatient care in hospitals to outpatient care in the community. METHODS: A literature review regarding residency training in community psychiatry was performed using the Medline databases. Seven residency training programs' curricula were included in this review. The curriculum of each of these seven programs was also obtained from their respective Internet home pages. The authors describe the community psychiatry training curriculum of their affiliated program, the University of Alabama at Birmingham. RESULTS: A brief description of community training curricula of these eight programs is provided. These curricula, especially their unique characteristics, are then compared in a table. DISCUSSION: The psychiatry training programs differ in many noteworthy ways while providing their residents with "community psychiatry experience." The table provides a quick comparison and highlights the differences of these programs. The authors emphasize the need for a "model curriculum," taking into consideration various factors including the local resources available where the residents could rotate for their community psychiatry experience. CONCLUSION: The authors concluded that psychiatry residency training programs need to publish their curricula, so that other programs can modify their own curricula, if needed, and hence provide an excellent experience in community psychiatry to their residents and future psychiatrists. PMID- 15291134 TI - Internet use within a physician's practice in Kentucky. PMID- 15291135 TI - "A toast to my big brother, George, the richest man in town". PMID- 15291137 TI - Reflections on complementary and alternative medicine. PMID- 15291136 TI - The simple things in life. PMID- 15291138 TI - [Immunoglobulin light chain amyloidosis: recent molecular, clinical and therapeutic approach]. AB - AL amyloidosis is a rare disorder characterised by tissue deposition of a fibrillary proteinaceous material, formed from monoclonal immunoglobulin light (or exceptionally heavy) chains. Although it may complicate multiple myeloma or B cell lymphomas, AL amyloidosis is often associated with a low burden of clonal plasma cells ("primitive" AL amyloidosis). The mechanisms involved in the formation of AL amyloid deposits remain unclear, but are probably related to structural peculiarities of monoclonal immunoglobulin light chains. AL amyloidosis is usually a systemic disease, often revealed by renal involvement, the most common complication of the disease. The longterm prognosis of AL amyloidosis is poor, mainly related to amyloid restrictive cardiomyopathy leading to congestive heart failure. Oral melphalan and prednisone is considered the standard treatment for AL amyloidosis, but with limited increase in the median survival. High-dose intra-venous melphalan with autologous stem cell transplantation is an effective treatment, aimed at eliminating the clonaly expanded plasma cells, which has been shown to induce complete hematologic remissions and to prolong survival. However, the tolerability of such treatment is low, limiting its use to selected patients. The development of new drugs, able to interfere with amyloid fibril deposition, may provide a new therapeutic approach. PMID- 15291139 TI - [Immune dysfuntion of uremic patients: potential role for the soluble form of CD40]. AB - Immune deficiency is one of the numerous physiological alterations associated with chronic renal failure. Recurrent bacterial infections, diminished vaccinal response or beta2-microglobuline amyloidosis are some common features of clinically well known dysregulations of uraemic immune functions. During the last ten years, our knowledge concerning the molecular and cellular effectors involved in the immune response has considerably progressed. Recent data clearly demonstrated that despite their activated phenotype, the effector capacity of the immune cells are severely hampered. Of interest, those abnormalities are not corrected by haemodialysis which sometimes even accentuate them. However, the presence of uraemic toxins in the serum of chronic renal failure patients jeopardise the determination of the factors responsible for the alteration of immune response in those patients. Among those molecules, the soluble form of CD40 (sCD40), which seric levels are dramatically increased, seems to play a crucial role. A better understanding of the molecular and cellular actors involved in the immune disorders of uraemic patients should allows the emergence of new immuno-intervention strategies and improve haemodialysis traitement. PMID- 15291140 TI - [Sequential protocols with thymoglobuline and delayed introduction of calcineurin: effects on renal function in renal transplant recipients with non immunological risk and low risk of delayed graft function]. AB - Induction therapy with thymoglobuline significantly decreases the risk of acute rejection, particularly in high immunological risk patients, in combined transplantations and in pediatric renal transplantation. After the introduction of cyclosporin in the 1980's and the recognition of its potential nephrotoxicity, induction therapy with delayed introduction of cyclosporin (sequential protocols) have became very popular in many transplant centers and are now used even in low immunological risk patients. The aim of the present study was to assess the recovery of renal function in low-immunological risk patients receiving a sequential protocol versus those receiving a calcineurin inhibitor at day 1. Among patients receiving their first transplant in our center between January 1999 and June 2000, we selected 72 recipients with no immunological risk and no risk of delayed graft function (DGF). 35 patients (group I) have received a sequential protocol whereas 37 patients (group II) have received a calcineurin inhibitor (Neoral or prograf) at day 1. We analysed creatinine reduction ratio, 24-hour creatinine excretion on post-transplant day 2, and serum creatinine and creatinine clearance at day 15, 30, 45, 60 and 90 post-transplant. There was no difference between groups concerning demographic, immunological or surgical parameters. The percentage of patients with immediate graft function was similar between the two groups (10 versus 9). The number of patients requiring dialysis in the first post-transplant week was also similar (8/35 versus 6/37). The day serum creatinine reached 200 micromol/l was 15 +/- 11 versus 14 +/- 12 in groups I and II respectively. Serum creatinine and creatinine clearance were similar at all time intervals. CMV disease was significantly higher in the group 1 (42% versus 18.5%; p < 0.005). Our data suggest that in patients with low immunological risk and low-risk of DGF, introduction of calcineurin inhibitors as early as the post-transplant day 1 is not deleterious for renal function recovery. These data should be confirmed by a prospective randomised trial. PMID- 15291141 TI - [Surveillance of infections in chronic hemodialysis patients]. AB - OBJECTIVE: To confirm rates of infections from a previous survey in chronic hemodialysis patients; to get information about incidents and manipulations of vascular access-site, number and reasons of hospitalisation; to asses a relationship between the frequency of vascular access-site infections (VASI) and quality of care during the procedures of vascular access-site use. DESIGN: Prospective, multicenter survey performed from February 2000 to January 2001, including all patients underwent chronic hemodialysis in 5 participating centers. Standardized definitions used and different clinical and biological risk factors recorded. RESULTS: 429 patients for a total of 4273 dialysis months (DM) were enrolled. 245 infections in 164 infected patients were reported. The overall rate was 5.73 infections per 100 DM (18 VASI, 25 bacteraemia, 84 respiratory, 29 urinary tract, 1 endocarditis and 88 other infections). 50% of infections were microbiologically documented. 19 of 21 antibiotics resistant microorganisms were meticillin resistant Staphylococcus aureus. Compared to the incidence rate of fistula (0.05 per 1000 days of follow-up) or prosthesis related VASI (0.11), the incidence rate of catheter related VASI (0.65) was significantly higher. Poor hygiene and duration of catheter use were the significant risk factors for VASI showed by logistic analysis regression. VASI and bacteraemia occurred more frequently after incident or manipulation of the vascular access-site. The decrease of VASI between the 2 periods of survey was significantly higher in centers having reduced the catheter use and implemented written protocols. CONCLUSIONS: This second period of surveillance has confirmed the frequency of infections rate in chronic hemodialysis patients and particularly bacteraemia and VASI. This study has allowed to establish risk factors for infections and showed that VASI in hemodialysis are related to factors in part preventable. PMID- 15291142 TI - [Verocytotoxin positive haemolytic uremic syndrome and HIV-associated nephropathy]. AB - Haemolytic uremic syndrome (HUS) and HIV-associated nephropathy (HIVAN) are common renal diseases in the course of HIV-infected patients. CASE REPORT: We report the case of a 13-month-old Caucasian boy hospitalised for a verocytotoxin positive HUS associated with HIV infection. After the acute phase of HUS the creatinine level returned to normal values. Because of progressive renal failure with severe overload hypertension and glomerular proteinuria despite antiretroviral therapy and angiotensine converting enzyme inhibitor, the child required peritoneal dialysis 12 months later. Clinical and biological course together with pathological findings were consistent with both typical HUS and HIVAN. CONCLUSION: This is the first paediatric case of typical HUS revealing a HIVAN. The association of HUS and HIVAN may explain the progression to end-stage renal failure despite antiretroviral therapy associated with angiotensine converting enzyme inhibitor and a good control of HIV replication. HIVAN is rare in children and may occur in the early phase of HIV infection even not only in black patients. PMID- 15291143 TI - [Information and recommendations to patients to preserve vessels for vascular access]. PMID- 15291144 TI - [Information and recommendations to patients with arteriovenous fistula]. PMID- 15291145 TI - [Recommendations for prevention of contrast-media induced nepropathy]. PMID- 15291148 TI - [Reception of research in the natural sciences in middle Germany at the Padua University betwee 1770 and 1820]. AB - The German literature on natural sciences that was present in the public libraries in Padua between 1770 and 1820 is described. The citations of German authors in the publications of Paduan naturalists of that time and of the textbooks used in Padua University are outlined. German journals on natural sciences available in Venice and Padua and Italian translations of German monographs of that time are also documented. With the foundation of the Italian Empire by Napoleon, the organization of lectures and research in the University of Padua changed drastically. In consequence, the reception of chemistry and physics was exclusively directed to France. In the descriptive natural sciences the earlier German traditions prevailed. Therein, however, Paduan sciences adopted the earlier descriptive traditions that already existed at the end of the 18th century and did not respond to the new developments in German functional morphology and physiology. Jenensian naturalists, botanists and physicists who received attention in Padua around 1800 are described as part of the empiric tradition of Central Germany and not as followers of the speculative "Naturphilosophie". There is no explicit reference to romantic sciences. PMID- 15291149 TI - [The neurology department of the Lankwitz Hospital. A contribution to the history of emigration, psychotherapy and the Berlin hospital]. AB - At Lankwitz near Berlin the Jewish physicians James Fraenkel and Albert Oliven founded a private hospital in 1890. This hospital, which integrated seven departments, became one of the biggest private asylums in the Reich during the first decade of the 20th century. Parts of the hospital served the military during WWI. As most of the physicians at Lankwitz were Jewish, the year 1933 meant an immense moral and scientific decline, since these physicians were forced to leave and later threatened. We know of one physician killed in a concentration camp. Hardly any research about this hospital had been published until the 1990's. This contribution is the first portraying one department--that of neurology. From the early years of the hospital, therapists involved with psycho dynamic psychiatry and psychoanalysis were attracted to it. Lankwitz was a major experience for a number of later famous psychiatrists and psychoanalysts. This continued to be the case during the years of the Weimar Republic, after the hospital had been leased [verpachtet] to the insurance companies [gesetzliche Krankenkassen] of Greater Berlin. Revealing more about the history of Berlin's private clinics, this article also contributes to the city's history of neurology, psychiatry and psychotherapy. For the example of Lankwitz, the myth of a 'marginalized psychoanalysis', rejected by the contemorary medicine, cannot be corroborated. On the contrary, the Lankwitz physicians portrayed here seemed to have combined and integrated clinical work with psychoanalytic theory and practice. Germany's turn towards National Socialism however meant an immediate end for the Lankwitz clinic and the mode of therapy described. PMID- 15291150 TI - [German translation of the syphilis poem by Girolamo Fracastoro by the Essling wound physician Ernst Philipp Heinrich Spath (1809-1856). An unpublished hanwritten document of Dresden historical collection]. AB - In 1840 the Dresden physician and professor at the Chirurgisch-medicinischen Akademie Johann Ludwig Choulant (1791-1861)--at that time one of the most well known experts in history of medicine--received a German translation of the Fracastoro syphilis poem. This philological piece goes back to the physician and surgeon Ernst Philipp Heinrich Spath (1809-1856) who had already worked on the poem of Girolamo Fracastoro (1478-1553) and its translation in the 1830s. Spath referred to the 1830 Latin publication of Choulant's syphilis poem--which he later used as the literary basis for his adaptation. Spath had acquired his knowledge of philology at a theology seminar in Urach and during his theology studies inTubingen. However, he quit his theology studies to switch to medicine. In 1832 he took the state examination and received a doctorate of medicine and surgery. In the very same year he established a practice as general practitioner, surgeon and obstetrician in Esslingen to eventually be appointed chief surgeon at the Esslingen hospital. Besides his work as a physician Spath was also a publisher, e.g. he also became the editor of the first local Esslingen newspaper. Spath's translation, apparently finished almost completely already in 1837, was sent to Choulant to ask his opinion in 1840. This version seems to be not only one of the rarest but also earliest German adaptations. Although hardly any other medical poem was so often translated into modern languages as Fracastoro's "Syphilidis sive morbi gallici libri III", there are only three German publications mentioned in a 1935 bibliography with the first complete German translation being published in 1858. Spath's script, however, has never been published and has remained in the handwritten estate of Choulant in Dresden ever since. PMID- 15291151 TI - [Appendix to the "Darmstadt Johannes-Alexandrinus Fragment"]. AB - This appendix to "Das Darmstadter Johannes-Alexandrinus-Fragment" answers to the critical note by Klaus-Dietrich Fischer, and shows that these new-founded fragments are a testimony of an until now unknown abbreviated version of the Alexandrinian commentary on Galen's book 'De sectis'. PMID- 15291152 TI - [Treatment of coronary disease]. PMID- 15291153 TI - [From sirolimus to the Cypher stent: stages of the victory against restenosis]. PMID- 15291154 TI - [Sirolimus endoprosthesis: data from the RAVEL and the SIRIUS study]. PMID- 15291155 TI - [The E-SIRIUS and the new SIRIUS clinical trials]. PMID- 15291156 TI - [The e-Cypher registry: objectives and organization]. PMID- 15291157 TI - [The national registry EVASTENT: cost-effectiveness analysis of the sirolimus active stent in diabetic and non-diabetic patients]. PMID- 15291158 TI - [BRIDGE registry (safety and efficacy registry Bx Cyper stent in the RevascularIzation of patients with siGnificative risk of rEstenosis): protocol description and preliminary results]. PMID- 15291159 TI - [Sirolimus-releasing coronary stent: lessons from the Rotterdam experience]. PMID- 15291160 TI - [Cypher stent: the German registry. Results of the German prospective multicenter registry on Cypher]. PMID- 15291161 TI - [The Cypher stent: most recent news of the 2004 Congress of the American College of Cardiology]. PMID- 15291162 TI - [Cypher: ongoing studies]. PMID- 15291163 TI - [The Siro-ISR registry: treatment of intra-stent restenosis with the sirolimus stent in the real world]. PMID- 15291164 TI - [Cypher stent: upcoming clinical studies]. PMID- 15291166 TI - [Pizay days, Pizay events]. PMID- 15291165 TI - ["Active" endoprosthesis with in situ release of an antimitotic or cytostatic drug: a revolution that cannot do without thought]. PMID- 15291167 TI - [Efficacy and acceptability of lercanidipine are not age dependent in patients with essential hypertension: the AGATE study]. AB - Calcium channel blockers (CCB) are known to be more efficacious and better tolerated in elderly patients. Lercanidipine is a highly lipophilic CCB with a specific safety profile linked to its pharmacokinetics. OBJECTIVES: To evaluate and compare the efficacy and safety of lercanidipine according to age. METHODS: Two groups of hypertensive patients (G1: aged < 65, G2: aged > or = 65) entered an open study conducted over 56 days. All received lercanidipine 10 mg/d (monotherapy or add-on), titrated to 20 mg/d if blood pressure (BP) was not controlled at D28. BP was measured using a semi-automatic device at doctor's office (three measurements at 1-min intervals) and at home by the patient himself (three measurements in the morning and in the evening at 1-min intervals over the 7 days before D0 and D56). RESULTS: Seven hundred and fifty-six patients entered the study. Thirty-eight patients dropped out prematurely and 30 were excluded because they were normotensive; 691 patients (G1 n = 375, G2 n = 316) were kept for analysis. At the end of the study, 507 patients were treated with lercanidipine alone (10 mg/d n = 221, 20 mg/d n = 286) and 184 with a combination including lercanidipine (10 mg/d n = 91, 20 mg/d n = 93). Efficacy was not different between the groups excepted home pulse pressure which decreased more in G2. In the office, SBP decreased by 17 and 21 mmHg, respectively, for G1 and G2, and DBP by 9 and 10 mmHg. The prevalence of leg edema was not different between G1 and G2 and was particularly low in both groups (3%). CONCLUSION: Lercanidipine was as efficacious and well tolerated in younger patients as in elderly patients. PMID- 15291168 TI - [Drug prescription in the post-percutaneous coronary interventions period: results of the ECART study]. AB - Among 1000 French cardiologists, the ECART study investigated drug prescription in 1041 patients with coronary heart disease, before and after percutaneous coronary interventions. The baseline drug prescription rate CAD patients were the following: beta-blockers 96%, antiplatelets agents 85%, statines 56%, nitrates 36%, calcium blockers 26% and ACE inhibitors 8.7%. The main changes in patients having undergone PCI were: a significant increase in antiplatelets agents (to 97%), ACE inhibitors (to 29%) and statins (to 94%), a significant decrease in nitrates (to 23%). The calcium blockers rate remains unchanged at 26%. Those results are discussed in the field of evidence based medicine and are compared with data from previous drug prescription studies in post myocardial infarction or in secondary prevention. PMID- 15291169 TI - [Autonomic profile and cardiovascular symptoms]. AB - The autonomic nervous system controls all body functions. Dysregulation of this system, called dysautonomia, is responsible of polymorphic functional symptoms. Its exploration can reveal abnormalities which explains the clinical symptoms. The objective of this work is to try to find a connection between functional signs, mainly cardio-vascular and autonomic abnormalities. We also consider the autonomic profile of a group of patients with primary hypertension. METHODS: Patients with functional symptoms and normal clinical and paraclinical examinations underwent autonomic profile study. The following tests were performed: deep breathing, hand grip, echo stress, Valsalva maneuver and tilt test. RESULTS: Two hundred and eighty four patients were included in this study, females 70%, mean age 44 years (extremes 9 and 81). In this group of patients, the main autonomic syndromes found were: orthostatic hypotension, 88 patients in whom the alpha peripheral sympathetic activity is decreased and the vagal one is increased, postural orthostatic tachycardia syndrome (POTS), 123 patients who describe orthostatic symptoms and in whom beta sympathetic activity is high, the neurocardiogenic syncope, four patients in whom vagal activity is high, and the baro-receptor abnormality (115 patients) of variable degree. In patients with primary hypertension (137), the alpha sympathetic activity is high (mean increase of blood pressure of 38.5%), meanwhile the vagal one is low. Twelve patients having diabetes showed a low vagal activity, (mean EKG RR variation of 8%). The treatment, depending on the autonomic abnormality (hygienodietetic measures, ethylephrine, fludrocortisone, phenobarbital, clonidine, maproptyline, serotonine recapture inhibitor) showed an improvement in the functional state of the patients. CONCLUSION: Autonomic profile study helps to explain the causes of symptoms described by the patients whose clinical and conventional paraclinical examinations are normal. PMID- 15291170 TI - [Hypertension: the main causes revisited]. AB - Does "essential" mean "ignorance"? The historical demonstration made by Jerome Conn who was the first to describe primary hyperaldosteronism, back in 1954, shows how an outstanding diagnostic route has enlightened our knowledge. Using the examples of hyperaldosteronism and reno-vascular hypertension, a reflex ion is made to arouse our curiosity. In the end, the term "essential" is only the cover-up of what we yet have to learn. PMID- 15291171 TI - [Cardiac consequences of primary hyperaldosteronism]. AB - The activation of the renin-angiotensin system is associated with vascular and cardiac hypertrophy. But there are few data on the renal and cardiac consequences of the hypersecretion of aldosterone. In the experimental setting, hyperaldosteronism leads to an excess of fibrous interstitial tissue and cardiac hypertrophy. In man, these consequences are those of hyperaldosteronism. The aim of this study was to assess the cardiac consequences of hyperaldosteronism in a series of 31 patients with a documented Conn adenoma, in comparison with a matched population of 31 patients with primary hypertension. For the same level of blood pressure, cardiac hypertrophy is more prominent in hyperaldosteronism and there is a positive correlation between the level of plasma aldosterone and left ventricular wall thickness. Left ventricular hypertrophy is of the concentric type. In addition, an increase in myocardial fibrosis (that can now be quantified by echocardiography) is observed, with a positive correlation between plasma aldosterone and reflected ultrasound which might correspond to increased myocardial collagen. These anatomic modifications of myocardial structure result in diastolic dysfunction. Overall, Conn adenoma is associated in accelerated disease, which is partly independent of the level of blood pressure. PMID- 15291172 TI - [Renal impact of primary hyperaldosteronism]. AB - The impact of hyperaldosteronism on target organs, and particularly kidney function, is greater than that of essential hypertension. Hyperaldosteronism provokes a glomerular hyperfiltration and hypertension that may cause renal alterations. Those may explain why elevated blood pressure may persist, even after radical treatment of the cause of hyperaldosteronism. PMID- 15291173 TI - [Agonists and antagonists of mineralocorticoids. The relation between structure and activity]. AB - The mechanism of action of aldosterone and its links with the mineralocorticoids receptor (MR) are described. The physiologic importance of the MR structure is emphasized, in relation with the preferential activation of the receptor by aldosterone. PMID- 15291174 TI - [Surgical treatment in primary hyperaldosteronism]. AB - From 1994 to 2000, 32 patients (16 men, 16 women; mean age 53 years) underwent laparoscopic adrenalectomy at Hopital l'Antiquaille of Lyon. All but one had systemic arterial hypertension and hypokaliemia was noted in all patients. All patients had unilateral adrenalectomy, and only one required conversion to conventional laparotomy. There were no deaths and complications were rare. PMID- 15291175 TI - Me, and Walter Reed. PMID- 15291176 TI - Errors in temperature conversion from Celsius to Fahrenheit in the article "The Golden Hours" for Heatstroke Treatment. PMID- 15291177 TI - Wound Shock: A History of Its Study and Treatment by Military Surgeons. PMID- 15291178 TI - Trends in positive drug tests, United States Air Force, fiscal years 1997-1999. AB - We investigated the relationship between various demographic factors and the risk of testing positive for marijuana or cocaine use in the U.S. Air Force in fiscal years 1997 through 1999. Overall test positive rates for marijuana and cocaine were very low, at 0.24 and 0.07% of all tests, respectively. However, monthly test positive rates increased significantly during the study period while the number of tests conducted decreased by more than 50%. Gender, race/ethnicity, service component, military rank, education level, and assignment location each predicted the likelihood of testing positive for marijuana or cocaine use. These findings were consistent with annual surveys of self-reported drug use conducted in military and civilian populations in the United States. We conclude that overall testing percentages should be re-evaluated in light of these findings, but we do not recommend oversampling from population subgroups that demonstrated a higher likelihood of testing positive. PMID- 15291179 TI - Evaluation of an over-the-counter medication program. AB - Nonprescription medication (i.e., "over-the-counter") programs have historically been popular at many military treatment facilities. These programs were developed to provide a mechanism through which active duty members could obtain certain nonprescription medications without seeing a health care provider. The goal of such programs was to reduce demand on health care provider appointments and operational costs associated with such visits. Coast Guard military treatment facilities are encouraged to provide the service. The purpose of this paper was to evaluate whether the nonprescription medication program offered at the Coast Guard Alameda pharmacy reduced demand on health care provider appointments for self-limiting conditions and reduced costs associated with such visits. PMID- 15291180 TI - Dental emergency rates at two expeditionary medical support facilities supporting operations enduring and Iraqi Freedom. AB - This study reports dental emergency rates and distribution of causes of dental emergencies at two expeditionary medical support facilities supporting operations Enduring Freedom/ Iraqi Freedom. A retrospective cohort analysis of 9948 soldiers deployed to Prince Sultan Air Base, Kingdom of Saudi Arabia, and 1467 soldiers at Baghdad International Airport, Iraq, was accomplished from a phased deployment from January 2003 to September 2003. Procedures were divided into 11 categories: endodontic, extraction of teeth other than third molars, extraction of third molar teeth, restoration of teeth (caries), restoration of broken teeth (not caries), orthodontic bracket/wire problem, sensitive teeth, temperomandibular pain, periodontal, oral pathology, and prosthodontic. The dental emergency rates for Prince Sultan Air Base and Baghdad International Airport were 153 and 145 dental emergencies per 1000 soldiers per year, respectively. Most of the emergencies were because of dental caries. Pain from third molars was the second most common reason for visiting the dental clinic. PMID- 15291181 TI - Comparative measles incidence among exposed military and nonmilitary persons in Anchorage, Alaska. AB - An outbreak of measles that occurred in Anchorage, Alaska, in 1998 resulted in 33 diagnosed cases: 26 were laboratory confirmed and 7 were clinically confirmed. Twenty-nine (88%) of 33 cases occurred in individuals who had not been immunized with at least two measles-mumps-rubella (MMR) vaccinations; 25 (76%) of 33 occurred in school-age children, 0 to 19 years of age. This study identifies the difference in the incidence of measles between the civilian school-age population, who was not completely immunized (two MMR vaccinations given at least 30 days apart), and the military dependent population who had been completely immunized. All cases occurred among civilians, and most (25 of 33 confirmed cases) were associated with school attendance. The authors conclude that a two dose regimen of MMR vaccine is required to adequately protect individuals against measles. PMID- 15291182 TI - Laboratory-measured pregnancy rates and their determinants in a large, well described adult cohort. AB - OBJECTIVE: To measure the pregnancy rate directly, to describe the characteristics of women who become pregnant, and to identify the predictors of pregnancy. No recent studies have estimated population pregnancy rates using objective, laboratory-based criteria. Furthermore, none have characterized predictive factors of pregnancy. METHODS: Population-based prospective cohort study of 5578 women, ages 18 to 44, on active duty at Fort Lewis, Washington, from 1995 to 1997. Main outcome measures were standardized pregnancy incidence rate and predictive factors for pregnancy. RESULTS: In the cohort, 887 pregnancies and 597 births occurred during the study period. The age- and race standardized pregnancy rate was 108.1 per 1000 person-years. When compared with the 1995 U.S. population, the pregnancy rate ratio was 1.05 (95% confidence interval, 0.96-1.14). Factors that significantly affected the likelihood of pregnancy included age (=0.87/year), marital status (3.0, married versus single), race (1.2, African American versus Caucasian), Pap smear during study period (0.6), educational level (1.8, graduate training versus high school), and at least one prescription for oral contraceptives during the study period (0.8). CONCLUSIONS: Standardized pregnancy rates in the study population were statistically indistinguishable from United States estimates. Use of health care services was an important independent determinant of pregnancy occurrence. PMID- 15291183 TI - The effects of residency on physical fitness among military physicians. AB - PURPOSE: Demands of medical residency training would be expected to result in physical deconditioning. This study determined the effects of residency on physical fitness. METHODS: A prospective cohort study of the change in physical conditioning during residency using the Army Physical Fitness Test as a standardized measure of fitness was conducted. Ninety-four active duty U.S. Army physicians were followed during their medical residency training from July 1999 to June 2002. Army Physical Fitness Test results from interval tests performed throughout residency were compared to baseline results at matriculation. Changes in total score, number of push-ups and sit-ups completed in 2 minutes, 2-mile run time, and weight were recorded. Correlations between initial and final scores were evaluated. RESULTS: All measured parameters worsened with statistical significance noted in the weight (p < 0.01), push-ups (p < 0.05), and 2-mile run (p < 0.01). Statistically significant negative correlations between initial and subsequent tests were noted in push-ups (p = 0.004), sit-ups (p = 0.0003), and total score (p = 0.0192). CONCLUSIONS: Physical fitness declines during medical residency training. This effect is most notable in residents with higher levels of fitness at the start of medical education. Further studies are necessary to evaluate for a similar decline among civilian residents. PMID- 15291184 TI - Comparing elite soldiers' perceptions of psychological and physical demands during military training. AB - This research examined the impact of psychological and physical demands on a group of elite soldiers (N = 9) before, during and after training exercises involving routine and unrehearse tasks. Based on the psychological concept of toughening, we expected soldier responses to unrehearsed demands to be les resilient than to routine training demands. As hypothesized soldiers rated their perceptions of physical and psychologica demands higher during the unrehearsed training phase of the study. It was also hypothesized that soldiers would recover more from physical demands than from psychological de mands after the training exercise. Although a trend in the data marginally supported this hypothesis, a note of caution is warranted because of the small sample size. Limitations and implications of the study are discussed, with particular emphasis on the dual importance of psychological and physical training in preparing soldiers for mission demands. PMID- 15291185 TI - Liver injury after ischemia and reperfusion: the role of oxygen free radicals. AB - OBJECTIVE: This article describes conditions of ischemia and reperfusion in dog's liver. The aim of the study was to observe hepatic injury from oxygen free radicals and to evaluate mannitol and ascorbic acid as scavengers. METHODS: The criteria for evaluation were histopathological and biochemical. Twenty-one dogs were divided into three equal groups: group A with 90 minutes of ischemia and 10 minutes of reperfusion, and groups B and C with 120 minutes of ischemia and 10 minutes of reperfusion. In treatment group C, before the ischemia, mannitol and ascorbic acid were injected. RESULTS: Differences were significant for malondialdehyde in A and B group postischemia (p < 0.001). In group C, a significant difference (p < 0.091) was not observed. The histopathological examination revealed fatty degeneration, alterations of mitochondrial structure, and membranic cysts. In group C, the alterations were considerably milder. CONCLUSIONS: We can suggest the use of ascorbic acid and mannitol at the clinical level for scavenging and inhibition of oxygen free radicals in ischemic liver. PMID- 15291186 TI - Preventive health behaviors, health-risk behaviors, physical morbidity, and health-related role functioning impairment in veterans with post-traumatic stress disorder. AB - An examination of the relationships between health behaviors (preventive and risk related), physician-diagnosed medical problems, role-functioning impairment because of physical morbidity, and post-traumatic stress disorder was conducted on a large cohort of consecutive treatment-seeking cases (N = 826) presenting to an outpatient Veterans Affairs post-traumatic stress disorder clinic. Results revealed that the sample rates of several medical conditions were markedly elevated when compared with general population rates for men of comparable age. The rates of smoking and other behavioral risk variables were greater than rates among men in the general population. Moreover, the majority of the sample did not engage in preventive health behaviors such as exercise and medical screening at levels consistent with health care guidelines. Physical role functioning indices of the SF-36 reveal greater role-functioning impairment because of physical morbidity in this psychiatric sample relative to the age adjusted general population norms. The health care implications of these data are discussed, as are areas for future research. PMID- 15291187 TI - How leaders can influence the impact that stressors have on soldiers. AB - The present review addresses the importance of leader behaviors in influencing the extent to which various stressors soldiers experience (e.g., high workload and lack of sleep) are related to different types of strains (e.g., psychological health, poor job satisfaction, and low morale). Research conducted by the Walter Reed Army Institute of Research (WRAIR) in the area of leadership is reviewed. Researchers at the WRAIR have examined the role of leadership as a predictor of stress, as a buffer against the negative effects of stress, and as a variable that predicts or enables variables that have been found to decrease the adverse effects of stress (e.g., role clarity, self-efficacy, and job engagement). A key strength of the WRAIR program of research is the use of multilevel modeling to examine how perceptions of leadership at the unit level are related to unit and individual soldier well-being and motivation. PMID- 15291188 TI - A prospective trial of diaspirin cross-linked hemoglobin solution in patients after elective repair of abdominal aortic aneurysm. AB - We evaluated the safety, pharmacokinetics, and pharmacodynamics of diaspirin cross-linked hemoglobin (DCLHb) solution in patients after repair of abdominal aortic aneurysm. We performed a randomized, single-blind controlled study with 10 patients in the surgical intensive care unit of a tertiary care facility. Within 24 hours after repair of an abdominal aortic aneurysm, each patient received an infusion of DCLHb (50 mg/kg or 35 mL for a 70-kg patient) or an equal volume of hetastarch. Variables were measured before infusion, at 15 and 30 minutes postinfusion, and at hourly intervals up to 72 hours. Compared with controls, the experimental group had significantly greater mean pulmonary artery pressure at 30 minutes (mean +/- SD, 26.4 +/- 3.18 vs. 22.8 +/- 2.86 mm Hg), greater mean arterial pressure through 30 minutes (100.8 +/- 8.67 vs. 81.6 +/- 13.8 mm Hg), and greater plasma hemoglobin through 2 hours (69.3 +/- 6.08 vs. 1.8 +/- 0 g/dL). Cardiac output was significantly less in the DCLHb group at 2 hours (5.34 +/- 7.92 vs. 6.18 +/- 0.54 L/minute), levels of serum bilirubin were significantly less at 24 and 48 hours (94 +/- 0.26 vs. 1.56 +/- 0.73 mg/dL), and platelet counts were significantly greater at 24 hours (128 +/- 35.8 vs. 101 +/- 55.7 mg/dL). The two groups did not differ in oxygen delivery or consumption. One patient treated with DCLHb had a myocardial infarction 36 hours postinfusion. No patient had antibodies to DCLHb. At this dosage, DCLHb was well tolerated without severe organ dysfunction or toxicity. However, its use may lead to decreases in cardiac output because of increases in afterload, which may pose serious problems with left ventricular function. PMID- 15291189 TI - Sound attenuation of the indoor/outdoor range E-A-R plug. AB - The sound attenuation provided by the AOSafety Indoor/Outdoor Range E-A-R Plug was examined. This device, currently used in military operations in several countries, is comprised of two plugs that provide conventional ("indoor" plug) and level-dependent sound reduction (the "outdoor" plug), respectively. The effects of the user's gender and repeated fittings were explored. Eight men and eight women were tested on two separate occasions. Unoccluded and protected hearing thresholds were measured for each of nine one-third octave noise bands centered at 0.125 to 8 kHz. Attenuation was calculated as the difference between these two measures. The indoor plug provided 21 to 40 dB of sound reduction across the frequencies tested, closely matching the manufacturer's specification. The outdoor plug provided 5 to 22 dB of conventional attenuation, suggesting that it might serve as a safe means of conventional and level-dependant attenuation in hearing-impaired users. No differences were found in relation to gender or repeated fittings. PMID- 15291190 TI - Delayed traumatic occlusion of the extracranial internal carotid artery after earthquake: report of two cases. AB - We report on two patients with head and neck injuries, who remained asymptomatic after the earthquake for 1 and 2 years, respectively. They developed late transient ischemic neurological signs and eventually presented with complete occlusion of the internal carotid artery. They made a good recovery with medical treatment. Focal cerebral ischemic symptoms may develop months or even years after the head or neck trauma, and internal carotid artery occlusion should be considered in differential diagnosis as a late complication. PMID- 15291191 TI - Assessment of care and disease management of patients with asthma in an overseas U.S. Army health clinic. AB - The objective of this study was to evaluate care and management provided by family practitioners to patients with asthma. A retrospective chart review was conducted on patients treated for asthma at a military health clinic in Germany. Pertinent information from the National Asthma Education Program guidelines was gathered using a questionnaire. Eighty-five charts were reviewed and 44 charts were included in this study. Overall compliance was calculated using the 22 National Asthma Education Program points (12 questions) investigated by the questionnaire. Overall compliance was 64%. Components of four major categories of asthma care were examined. All categories except one were above or close to the overall compliance percentage--diagnosis (74%), treatment/control (90%), monitoring (61%), and patient education (25%). It appears from this study that family practitioners are providing adequate, but not optimal, care and management to patients with asthma. Although our findings were encouraging, this study has highlighted that there is opportunity for improvement. PMID- 15291193 TI - Foot drop due to cranial gunshot wound. AB - OBJECTIVE: We present a case of foot drop from hemorrhagic contusion after cranial gunshot, which has never been reported. METHODS: A 21-year-old man was admitted with inability of dorsiflexion 1 day after a tangential gunshot wound of the scalp. The scalp skin was cut by the rifle bullet. He had foot drop and his neurological examination was normal except for weakness at dorsiflexion of the right foot. Pathological reflexes and sensation failure were not detected. T1- and T2-weighted magnetic resonance images showed hyperintense contusion at the right superior frontal gyrus and mild subdural hemorrhage. Peripheral nervous system examination was electrophysiologically normal. Motor-evoked potentials showed the location of the lesion at the motor cortex because no electrical record was obtained from the right anterior tibial and extensor digitorum brevis muscles, and there was a normal record on the left. Six months later, the patient's neurological examination was uneventful. CONCLUSION: When a cranial gunshot wound injury victim presents with foot drop, the central causes should be included in the differential diagnosis list. PMID- 15291192 TI - Outcomes of Fort Jackson's Physical Training and Rehabilitation Program in army basic combat training: return to training, graduation, and 2-year retention. AB - Basic trainees at Fort Jackson, South Carolina, who were unable to continue basic combat training (BCT) because of a serious injury were assigned to the Physical Training and Rehabilitation Program (PTRP). Between January 3, 1998 and July 24, 2001, 4258 trainees were assigned to the PTRP. Using a retrospective cohort study design, return to training and BCT graduation rates were evaluated. PTRP graduates were compared with matched non-PTRP graduates for 2-year retention in the Army. More PTRP women than men were discharged from the PTRP (60% and 48%, respectively, p < 0.01). Of PTRP trainees returning to BCT, 10% and 12% of men and women, respectively, were discharged from the Army compared with overall Fort Jackson discharge rates of 9% and 15% for men and women, respectively. Comparing PTRP graduates to matched non-PTRP graduates, there were no differences in 2-year retention for men (14.9% and 14.7%, respectively; p = 0.93) or women (26.6% and 30.1%, respectively; p = 0.19). Despite the high discharge rate in the PTRP, the BCT discharge rate for trainees who successfully rehabilitated was similar to the overall discharge rate at Fort Jackson. The 2-year retention in service for PTRP trainees who graduated from BCT was similar to that of non-PTRP trainees. PMID- 15291194 TI - Fatigue, sore throat, and cough in a 24-year-old active duty man. AB - A 24-year-old active duty male smoker presented with 3 days of fatigue, rhinorrhea, and sore throat. The diagnosis and management of pharyngitis, including a field friendly approach, are reviewed. The impact of the discontinuation of the adenovirus vaccine to military recruits is highlighted. The effects of smoking among military personnel are discussed, and smoking cessation measures are reviewed. PMID- 15291195 TI - Stress fracture of the tarsal navicular. AB - Stress fractures of the lower extremity are common among military members and athletes at all levels of participation. They typically occur when an individual begins a new or different type of physical training or during periods of abrupt increase in the level of training. Stress fractures represent an incomplete remodeling of bone that occurs secondary to repetitive mechanical loading. In response to this increased loading, the osteoclastic resorption of lamellar bone outpaces the ability of the osteoblasts to create new lamellar bone, eventually leading to structural failure. The following case report reviews the typical clinical presentation, imaging findings, and treatment of the tarsal navicular stress fracture. PMID- 15291196 TI - Mental rotation and simulation of a reaching and grasping manual movement. AB - This paper deals with the kind of manual movement subjects mentally simulate when solving a left-right judgment task that requires rotating images of hands. 50 female students were asked to judge the laterality of drawings of rotated hands presented successively to the right and left visual hemifields by clicking on a mouse using either the right or left hand. Reaction times and accuracy of judgment were recorded. Analysis showed performances varied with the rotation angle at which the stimulus was presented, indicating that the subjects mentally simulated a rotation process. An interaction occurred between the visually presented hand and the responding hand, which suggests that the mental rotation process involved the simulation of a hand movement. Performance improved when the drawing of a hand was presented in the 'palm-up' position, and to the visual hemifield opposite with respect to the hand the subject moved mentally. The latter two findings suggest that the subjects performed a simulated reaching and grasping movement rather than a simulated positioning movement. PMID- 15291197 TI - Using computer-scored measures of emotion and style to discriminate among disputed and undisputed Pauline and non-Pauline epistles. AB - The Dictionary of Affect in Language that allows measurement of Pleasantness, Activation, and Imagery in texts and a computer program that provides several additional stylistic measures were used to score samples from Disputed (n = 22 samples) and Undisputed (n = 40) Pauline epistles and from Other New Testament epistles (n= 16). All samples came from an English translation. Several significant mean differences were noted between samples from Disputed and Undisputed epistles. A discriminant function predicting Disputed or Undisputed authorship limited to five predictors was 85% successful in assigning samples to either category. The majority of samples from Other epistles were classified as Disputed, i.e., less likely to have been written by Paul. Undisputed Pauline samples were predicted to be those with lower Imagery, shorter words, less frequent words, greater repetitiveness, and greater Pleasantness. There were significant differences in patterns of word use (vocabulary) between Disputed and Undisputed samples. PMID- 15291198 TI - Geomagnetic activity during the previous day is correlated with increased consumption of sucrose during subsequent days: is increased geomagnetic activity aversive? AB - In five separate blocks over a period of several months for 33 female rats the amount of geomagnetic activity during the day before ad libitum access to 10% sucrose or water was positively correlated with the volume of sucrose consumed per 24-hr. period. The strength of the correlation (.62 to .77) declined over the subsequent 10 days from between .12 to -.18 and resembled an extinction curve. In a subsequent experiment four rats exposed to 5 nT to 8 nT, 0.5-Hz magnetic fields that ceased for 30 min. once every 4 hr. for 4 days consumed 11% more sucrose than the four rats exposed to no field. We suggest that the initial consumption of 10% sucrose may have been reinforced because it diminished the aversive physiological effects associated with the increased geomagnetic activity. However, over the subsequent days, as geomagnetic activity decreased or habituation occurred, negative reinforcement did not maintain this behavior. PMID- 15291199 TI - Assessing potentially gifted students from lower socioeconomic status with nonverbal measures of intelligence. AB - The screening and identification of gifted students has historically been conducted using verbal measures of intelligence. However, the underrepresentation in gifted programs of culturally diverse children, who may have limited English proficiency or cultural values different from those measured in traditional intelligence tests, has prompted researchers to consider other measures. Nonverbal measures of intelligence have been utilized to increase the number of gifted children from diverse backgrounds. Researchers in the current study sought to increase the number of culturally diverse gifted students at a rural public school enrolling predominantly African-American students from low socioeconomic homes. 169 students in Grades 2 through 6 were assessed using three nonverbal measures of intelligence: the Culture-Fair Intelligence Test, the Naglieri Nonverbal Abilities Test, and the Raven Standard Progressive Matrices. The scores on these nonverbal measures indicated that the Culture-Fair Intelligence Test and the Raven Standard Progressive Matrices identified more students than the Naglieri Nonverbal Abilities Test. A discussion of the results and implications for research are presented. PMID- 15291200 TI - Elite athletes' differentiated action in trampolining: a qualitative and situated analysis of different levels of performance using retrospective interviews. AB - Using a situated cognition approach, this study analyzed elite athletes' actions, i.e., behaviors link to cognitions, during competitive trampoline performances, which are evaluated from a succession of 10 acrobatic movements characterized by flight time and fall risk. 27 exercises performed by 10 elite athletes were ranked poor, average, or good and analyzed. Self-confrontation interviews were conducted and transcribed in relation with behavioral descriptions derived from video recordings. Qualitative analysis was performed to identify units of meaningful action and their components. The succession of units describing the stream of actions was used to identify differentiated organization of trampolinists' performances. Three patterns, corresponding to performance levels, were distinguished by (a) an increasing number of meaningful actions occurring at the same time, (b) a reduction in actions of waiting, and (c) the emergence of new actions aimed at interaction with the situation. These results suggest that differentiation in performance level is linked with meaningful actions modified through interaction with the context. PMID- 15291201 TI - Random number generation in native and foreign languages. AB - The effects of different levels of language proficiency on random number generation were examined in this study. 16 healthy right-handed students (7 women, 9 men; aged 22 to 25 years, M=23.8, SD=.83) attempted to generate a random sequence of the digits 1 to 9 at pacing frequencies of 1, 1.5, and 2 Hz. Randomization was done in German (native language L1), English (first foreign language L2), and French (second foreign language L3). There was a pattern of redundancy and seriation tendencies, increasing with speed of generation for all languages (L1-L3). While using L2 and L3, responses slowed and the number of errors committed increased. Further, there was a peculiar pattern of dissociation in nonrandom performance with an increase of habitual counting in ones and a strong reduction of counting in twos. All effects were most pronounced when subjects used L3 and 2-Hz pacing rates. Slowing and nonrandomness was not correlated with self-assessment parameters regarding language proficiency. We suggest that in a task involving number activation in a nonnative language, lack of proficiency will interfere with random number generation, leading to interruptions and rule breaking, at least when reaching the limits of attentional capacity at higher pacing rates. PMID- 15291202 TI - Understanding Air Force members' intentions to participate in pro-environmental behaviors: an application of the theory of planned behavior. AB - At a single installation, a cross section of 307 active duty Air Force members completed questionnaires to assess whether the theory of planned behavior was useful in explaining the service members' intentions to participate in three environmentally protective behaviors-recycling, carpooling, and energy conservation. While the individual tenets of the theory of planned behavior, i.e., attitude toward the behavior, subjective norms, and perceived control, accounted for differing amounts of variance in intentions, the results indicated that the intentions of these Air Force members to recycle, conserve energy, and carpool were moderately explained by the tenets of the theory of planned behavior collectively when the results of a multiple regression were analyzed. PMID- 15291203 TI - Dynamic balance in high level athletes. AB - The purpose of this study was to investigate dynamic balance performance in highly skilled athletes. Participating athletes were currently competing at the collegiate Division I, professional, elite, or Olympic levels, or their individual coaches believed the athlete performed comparably to these levels. High level male and female gymnasts (n=57, M age=17.3 yr., SD=4.1), soccer players (n=58, M age= 19.8 yr., SD=1.6), swimmers (n=70, M age= 17.1 yr., SD=2.5), and individuals with no formal competitive sport experience (n=61, M age= 16.8 yr., SD= 2.0) volunteered. Dynamic balance performance was measured on a stabilometer, which requires participants to continuously adjust posture to maintain an unstable platform in the horizontal position for 30 sec. Each participant performed 3 practice trials followed by 7 test trials. Analysis indicated that athletes were superior to nonathletes in balance performance. Gymnasts performed better on the dynamic balance task than all other groups. Soccer players and swimmers performed similarly and were superior to the control subjects. There was no difference between the performance of men and women. Moderate to high negative correlations were found between dynamic balance performance and height and weight. PMID- 15291204 TI - Communicative responsibility and semantic task in the language of adults with dementia. AB - A probe technique requiring convergent and divergent semantic behavior and representing five levels of communicative responsibility served as the research tool. Stimuli were presented to adults identified as having Alzheimer disease or multi-infarct dementia. Within each group differences were observed on the semantic task (convergent and divergent) and on communicative responsibility. Group characteristics are compared with data previously published in 1994 on aphasic and schizophrenic adults responding to the same stimuli. PMID- 15291205 TI - Effects of expression and inhibition of negative emotions on health, mood states, and salivary secretory immunoglobulin A in Japanese mildly depressed undergraduates. AB - Previous studies have indicated that expression of negative emotions facilitates mental and physical health and inhibition of negative emotions increases susceptibility to illness. This study was conducted to examine whether those findings can be expanded to populations with non-Western cultural backgrounds. Specifically, we explored effects of expression and inhibition of negative emotions on health, mood states, and mucosal immune function in mildly depressed Japanese individuals. 16 depressed and 16 nondepressed female undergraduates were required either to write about their unpleasant experiences and superficial topics or to suppress any emotional responses and thoughts about them. Secretory immunoglobulin A (s-IgA) in saliva and psychological indices were measured at an experimental session and at a follow-up 1 wk. later. Beneficial effects of expression of emotions on subjective health were indicated in the nondepressed group, whereas harmful effects of inhibition on subjective health were shown in the depressed group. Emotional expression by writing improved mood states both in the depressed and nondepressed groups but induced elevation of salivary s-IgA only in the depressed group. PMID- 15291206 TI - Lateral difference and interhemispheric transfer on arm-positioning movement between right and left handers. AB - We investigated the transfer of an arm-positioning movement between the right and left arms of right and left handers. 30 male (15 strong right handers and 15 strong left handers) subjects were asked to perform a constrained criterion movement, 12 cm in length, with right or left arm and a test movement at estimated 6-, 12-, or 24-cm length with the contralateral arm. In the right handers, the constant error of the left arm test movement was near zero, and that of the right arm indicated overshooting. In the left handers, the constant errors of the left arm test movement were farther from zero than those of right arm test movement. Left handers as well as right handers showed manual asymmetry on positioning movement. A plausible explanation for the manual asymmetry on the arm positioning task is related to interhemispheric transfer of spatial information on positioning movement. PMID- 15291207 TI - Knowledge and information in prediction of intention to play badminton. AB - The aim of this preliminary study was to investigate the contributions of knowledge and information to the prediction of students' intention to play badminton. The sample, 121 students (53 men and 68 women) 18-25 years of age, were beginning students in a semester badminton course. A questionnaire was completed before and after the course (4 mo.). Hierarchical regression analyses showed strong association between the examined variables of the Planned Behavior Questionnaire, specifically signifying that knowledge, information, attitudes, subjective norms, perceived behavioral control, role identity, and attitude strength could account for anticipation of intention towards playing badminton. Overall, systematic access to information and knowledge appeared to accompany relatively greater intentions. PMID- 15291208 TI - Geophysical variables and behavior: C. Increased geomagnetic activity on days of commercial air crashes attributed to computer or pilot error but not mechanical failure. AB - Global geomagnetic activity (aa values) for the days of crashes of airplanes and for each of the three days before and after the crashes were compared for 373 events (years 1940 through 2002) attributed to unknown factors, mechanical errors, electronic/computer failures or pilot errors. Interactions between days and classifications of the crashes were due to the significantly greater geomagnetic activity on the days of crashes attributed to pilot or computer error but not to mechanical or unknown factors. Successive temporal analyses indicated that the elevated activity on the days of crashes attributed to pilot error have not changed over time, but there was an increase in those attributed to electronic errors after 1965. No more than 9% of the variance in geomagnetic activity on the days of the crashes was associated with the type of crash. These results are consistent with our hypothesis that some factor or factors associated with relative increases in geomagnetic activity may affect complex electronic systems composed of either silica (computer) or carbon (brain) aggregates. PMID- 15291209 TI - Eye-hand preference in schizophrenia: sex differences and significance for hand function. AB - Hand preference and eye dominance were investigated in 73 (30 women, 43 men) schizophrenic patients and 71 (30 women, 41 men) healthy controls. There were significantly more schizophrenic patients and normal controls who were significantly right-hand dominant. However, schizophrenic patients showed a significant excess of left-eye dominance relative to controls (65.8% vs 29.6%; Odds Ratio= 4.75, p< .001). In addition, female schizophrenic patients showed a higher rate of nonright (either left or inconsistent) eye dominance (80%) than male schizophrenic patients (55.8%) and controls (33.3%). Analysis of hand performance on the Purdue Pegboard Test indicated that schizophrenic patients who showed crossed eye-hand dominance scored higher than did patients without crossed eye-hand dominance. PMID- 15291210 TI - Conflicts, burnout, and bullying in a Finnish and a Polish company: a cross national comparison. AB - The prevalence of conflicts, burnout, and bullying among employees of two similar companies, one situated in Poland (n=66) and the other in Finland (n=330) was investigated with the Psychosocial Workplace Inventory of Bjorkqvist and Osterman. Both companies were of Finnish ownership and manufacturers in heavy industry. They were similar in most respects, such as organization, production and marketing. Significant differences were found between the two companies. Polish workers had higher scores on conflicts and self-experienced bullying, while Finnish workers reported higher burnout, both self-experienced and observed by others. PMID- 15291211 TI - Social support and salivary secretory immunoglobulin A response in women to stress of making a public speech. AB - Acute experimental stressors transiently increase volume of secretory immunoglobulin A (s-IgA) in saliva. The present study examined buffering effects of social support on response of s-IgA to a brief psychological stress (giving a public speech). 24 women were divided at random into three groups, an emotional support group, an informational support group and a no-support group (control). For each group, s-IgA measures were obtained from each person under baseline conditions, during preparation of a speech when social support or no support was given, immediately after the speech and during a 'recovery' period. Level of s IgA in the control group significantly elevated during preparation for the speech and just after the speech compared to baseline, suggesting that the speech task stimulated secretory immune function. On the other hand, the subjects in the emotional support group showed increased s-IgA during the preparation period but secretion of s-IgA rapidly returned to the baseline after the speech task. Secretion of s-IgA in the informational social support group was unchanged at any measurement point. These results suggest that social support attenuates the affect of a stressor on somatic state. PMID- 15291212 TI - Body-shape perceptions in older adults and motivations for exercise. AB - This study examined the relationships among age, sex, exercise and body-image dissatisfaction in older adults and evaluated the role of body-shape dissatisfaction as a motivation to exercise. A pencil-and-paper questionnaire was administered to 175 older adults (101 women and 74 men) ranging in age from 50 to 98 years (M=72 yr., SD=9) to obtain general information, information regarding exercise participation, motivations for exercise and body-shape perceptions. A body-shape dissatisfaction score was calculated using the difference between the participant's choice for current and ideal body shape from a nine-figure body silhouette scale. Present study findings suggested that both older adult men and women expressed a desire for a thinner body shape independent of age and current participation in exercise. In addition, the results indicated that body-shape dissatisfaction did not motivate this sample to engage in regular exercise; physical health and physical fitness emerged as the most important motivations to exercise. PMID- 15291213 TI - Handedness differences in widths of right and left craniofacial regions in healthy young adults. AB - In this work, handedness differences in the widths of right and left craniofacial regions were studied in a healthy sample of 39 male and 43 female students, 17 to 23 years old. Width of craniofacial regions was assessed by computerized tomography. Handedness was associated with the left face width especially for women. The left facial region was larger for right-handers than left-handers. The smaller measure for the left face of left-handers might be associated with an advantage of left ear sensitivity. PMID- 15291214 TI - Abstractness and emotionality values for 398 English words. AB - This study is aimed to replicate Vikis-Freibergs' classic study (1976) on the values of vividness for French words. Vividness resulted from the concreteness and the emotionality values of words, here defined, respectively, as referring to something that can be experienced through senses and that can arouse pleasant or unpleasant emotions. 398 English words were rated on two different scales, Abstractness and Emotionality, by a group of English native speakers and also by a group of Italian subjects who used English as a second language. Results show a low correlation between the concreteness and emotionality ratings in line with Vikis-Freibergs' previous study of French words (1976). A negative correlation between Abstractness and Emotionality was observed for British data but a slightly positive correlation for the Italian data. PMID- 15291215 TI - Role of strategies and prior exposure in mental rotation. AB - The purpose of these two studies was to examine sex differences in strategy use and the effect of prior exposure on the performance on Vandenberg and Kuse's 1978 Mental Rotation Test. A total of 152 participants completed the spatial task and self-reported their strategy use. Consistent with previous studies, men outperformed women. Strategy usage did not account for these differences, although guessing did. Previous exposure to the Mental Rotation Test, American College Test scores and frequent computer or video game play predicted performance on the test. These results suggest that prior exposure to spatial tasks may provide cues to improve participants' performance. PMID- 15291216 TI - Mastery of number conservation by a man with severe cognitive disabilities. AB - An autistic 21-yr.-old with a mental age of four years was taught number conservation. Mastery of this concept requires concrete operational thought and has not been thought to be possible for persons with severe disabilities. A learning set of 105 problems was used to promote generalization, and a "fade-out" procedure was used to make mastery of the problems as easy as possible. Combination of these techniques produced the first recorded success in teaching number conservation to a person with severe disabilities. This demonstration that one individual can perform at this cognitive level opens the door to research to determine the generality and limits of the potential of other individuals with severe cognitive disabilities. PMID- 15291217 TI - Anchoring procedures in reliability of ratings of perceived exertion during resistance exercise. AB - Although the validity of perceived exertion as a method of monitoring the intensity of resistance exercise has been established, little is known about the test-retest reliability of ratings of perceived exertion during resistance exercise. Specifically, it is unknown whether the use of different anchoring procedures influences the reliability of ratings of perceived exertion. 30 men were assigned to an Exercise, Memory, or combined Exercise and Memory anchoring group. Participants completed an assessment of maximal leg-extension strength and were introduced to the Borg 15-category rating of perceived exertion scale through anchoring procedures that varied across groups. During two sessions of resistance exercise, participants rated active muscle perceived exertion after performing one repetition of the leg-extension exercise at 40%, 50%, 60%, 70%, 80% and 90% of the one-repetition maximum. A three-factor (Group x Intensity x Session) analysis of variance was performed to examine the perceived exertion data. Perceived exertion increased significantly (p<.01) with increasing exercise intensity in all groups and in both sessions. Mean ratings did not differ significantly among groups. Reliability was assessed for each group. Intraclass correlation coefficients ranged from .07 to .80 and percent agreement ranged from 60% to 90%. The results indicate that the reliability of ratings of perceived exertion during resistance exercise is acceptable regardless of the type of anchoring procedures used. PMID- 15291218 TI - Effects of random outcomes on choice behavior. AB - Decision-making researchers have shown that making optimal decisions is aided by the detection of information salient to the task. When the task involves random events, humans tend to perceive these events as contingent. In this study, outcomes were grouped together with choices to identify some of the conditions under which random events are correctly perceived. Of the two groups (ns=40) only one was provided information regarding the relationship between choice and outcome. This provision did not improve the detection of the relationship between random events any more than direct contact with the underlying contingencies. Findings are discussed in terms of experiential contact with and sensitivity to underlying contingency. PMID- 15291219 TI - Sleep deprivation and hemispheric asymmetry for facial recognition reaction time and accuracy. AB - We investigated the processing of emotional stimuli during a non-sleep-deprived state and following sleep deprivation in 36 right-handed men. Using the visual half-field technique, cartoon line drawings of emotional facial expressions were flashed on a computer screen for 250 msec. The participants were instructed to remember the content of the picture seen and to recognize it among nine alternatives shown immediately after the display of a single picture. Compared to the nondeprived condition, response latencies increased and accuracy decreased in sleep deprivation. Moreover, response latencies indicated that the performance of the right hemisphere deteriorated more following sleep deprivation than did the performance of the left hemisphere. The results also showed that hemispheric preference (for response latencies and response accuracy) tended to favour the left hemisphere when the participants were tested during sleep deprivation. PMID- 15291220 TI - Cancel and rethink in the Wason selection task: further evidence for the heuristic-analytic dual process theory. AB - The reasoning process in the Wason selection task was examined by measuring card inspection times in the letter-number and drinking-age problems. 24 students were asked to solve the problems presented on a computer screen. Only the card touched with a mouse pointer was visible, and the total exposure time of each card was measured. Participants were allowed to cancel their previous selections at any time. Although rethinking was encouraged, the cards once selected were rarely cancelled (10% of the total selections). Moreover, most of the cancelled cards were reselected (89% of the total cancellations). Consistent with previous findings, inspection times were longer for selected cards than for nonselected cards. These results suggest that card selections are determined largely by initial heuristic processes and rarely reversed by subsequent analytic processes. The present study gives further support for the heuristic-analytic dual process theory. PMID- 15291221 TI - The fragmented self. AB - For a sample of 182 undergraduates analysis of responses to a 10-item scale to measure the plurality of the self yielded three orthogonal factors and only moderate reliability. PMID- 15291222 TI - A mobile device for measuring sensorimotor timing in synchronized tapping. AB - This paper describes the design of a mobile device for examining sensorimotor timing. Control software installed in this device has facilities for storing time series data of interstimulus onset intervals, intertap onset intervals, and response duration in a comma-delimited file of ASCII text format as well as for running an experiment on synchronization tapping. The device provides a highly convenient way to allow collecting such timing data even in real situations like a kindergarten or a day care center for elderly people, given its mobile property and ease of use. PMID- 15291223 TI - Differences in African-American and Euro-American athletes' perceptions of treatment by coaches. AB - Analysis of responses from 418 respondents from southern USA (198 African American, 220 Euro-American adults) in 53 different locations at 4 colleges and universities showed that African-American and Euro-American high school and college athletes differed significantly in agreement on 4 of 12 statements representing their treatment by coaches. African-American athletes rated their coaches significantly more negatively on these items. Implications for coaches lie in planning, design and evaluation of coaching behaviors by African-American athletes. PMID- 15291224 TI - Effects of strategy use on acquisition of a motor task during various stages of learning. AB - To specify the optimal point for introducing a learning strategy, 50 participants were randomly assigned into five groups based on the timing of strategy introduction while learning a badminton serve. Groups were instructed in the use of Singer's Five-step Strategy either prior to starting their acquisition trials (100% group) or following acquisition Trial Blocks 1 (83% group), 3 (50% group), 5 (17% group) or were assigned to a control (0% group) group). Participants were asked to complete six acquisition trial blocks of 10 serves each, followed by a break and then two retention trial blocks. Scores were obtained by hitting shuttles into a scoring grid, which served as the dependent measure. Data were analyzed using a mixed-model analysis of variance with a group x trial blocks design, which yielded significant main effects for both factors during acquisition. Introduction of a learning strategy may be more efficient once participants have become familiar with the task. No significant differences were observed between groups who received the strategy early and the control group. Thus, it appears that learning strategies should be introduced later in the learning process and may distract if provided too early. PMID- 15291225 TI - Students' perceptions of dangerousness to public safety of paraphrases from the Koran, New Testament, Book of Mormon, Tibetan Book of the Dead, and Egyptian Book of the Dead presented as patients' beliefs. AB - In one experiment 40 first-year psychology students were asked to judge dangerousness to society of 10 fictitious patients who professed beliefs about an "alien." The statements were actually paraphrases primarily concerning death and killing from the New Testament, the Koran, the Book of Mormon, the Egyptian Book of the Dead and the Tibetan Book of the Dead. In a second experiment 39 first year psychology students were asked to rate the dangerousness of the verbatim statements with their sources identified. In the first experiment, statements from the Koran, which involved accessing a positive afterlife by killing nonbelievers in the name of a deity, were ranked as more dangerous. The differences between the sources accommodated 33% of the variance in the rankings for dangerousness. The group of students who were given the original statements and their actual sources ranked the statements from the New Testament and the Koran as significantly less dangerous than those who were told the statements were from patients. These results suggest that statements about killing and death may be rated as less dangerous if the person believes the source was a "sacred text." PMID- 15291226 TI - Mock jurors' perceptions of facial hair on criminal offenders. AB - Two studies were conducted to measure whether mock jurors would stereotype criminal offenders as having facial hair. In Study 1, participants were asked which photograph belonged to a defendant in a rape case and which photograph belonged to a plaintiff in a head-injury case after they were "accidentally" dropped. The photographs were similar in appearance except one had facial hair. 78% of 63 participants (or 49) identified the photograph with facial hair as being involved in the rape case. In Study 2, 371 participants were asked to sketch the face of a criminal offender. 82% of the sketches (or 249) contained some form of facial hair. Results are consistent with the hypothesis that criminal defendants are perceived as having facial hair. PMID- 15291227 TI - Dementia and identification of words and sentences produced by native and nonnative English speakers. AB - The accurate identification of 30 words and 15 sentences spoken by native English, Taiwanese, and Spanish speakers was compared for 16 persons with and 16 persons without dementia. Statistically significant differences for words and sentences occurred between groups of listeners. PMID- 15291228 TI - Effects of automatically delivered stimulation on persons with multiple disabilities during their use of a stationary bicycle. AB - We assessed the effects of automatically delivered favorite stimulation on engagement and indices of happiness of two adults with multiple disabilities during their use of a stationary bicycle. The participants typically received four 5-min. sessions per day over a period of about 3.5 mo. Analysis showed that one participant had a significant increase in both those measures while the other participant had a significant increase in engagement during the intervention phases of the study (when the stimulation was present) as opposed to the baseline periods (when the stimulation did not occur). Implications of the findings are discussed. PMID- 15291229 TI - Velocity of a tennis serve and measurement of isokinetic muscular performance: brief review and comment. AB - Isokinetic strength testing provides objective and reliable muscular performance data on elite tennis players; however, such data are not highly correlated with performance on a multiple-joint kinetic chain activity such as the tennis serve. In this brief review, an overview of the muscular performance characteristics generated through isokinetic testing for elite tennis players is presented. Application of population specific isokinetic strength profiles in specific populations for rehabilitation and performance enhancement is recommended. PMID- 15291230 TI - The stroop strategy test: investigation of a computerised version of the Stroop test for classifying Swedish military servicemen. AB - A computerised test termed the Stroop Strategy Test, utilising the Stroop effect, is described. To assess the test's usefulness and discriminant power, it was given to three military groups adjudged on the basis of interview and a test of intelligence to differ in their qualifications (61 men of Level 1, 41 of Level 2, and 17 of Level 3, in descending order) and a group of 16 men imprisoned or put in detention school for violent behaviour. In a discriminant analysis in which the eight measures the test provided were included and the four groups were compared yielded a discriminant power of 52.6% for the group as a whole, highest (56%) for the military Level 1 group. The potential usefulness of the test is discussed. PMID- 15291231 TI - False recognition with the Deese-Roediger-McDermott-Reid-Solso procedure: a quantitative summary. AB - In the Deese-Roediger-McDermott-Read-Solso (DRMRS) procedure, participants study lists of words associated with central concepts (critical themes) that are not on the lists, then their memory is tested. Based on 224 estimates, the rate of False Recognition of the nonstudied critical themes was .59 (95% confidence interval of .56 to .61), which is smaller than the Hit rate of .75 for correct recognition of studied items (95% confidence interval of .73 to .77) but greater than various rates of False Alarms for other nonstudied items (ranging from .13 to .19). Ratings of subjective confidence were similar on Hits and on False Recognitions but higher than on False Alarms, confirming that false recognition was more like correct recognition than like other errors. The results from judgments of feeling of remembering or knowing, from the effects of intervening activities (particularly recall) between study and test, and from the effects of age suggest that False Recognition occurs because the critical theme is activated along with studied items during list presentation and perhaps also during recall. Invoking fuzzy trace theory, it is argued Hits are based on verbatim traces whereas False Recognition is based on gist traces and a failure of source memory. Proposals are made for research. PMID- 15291232 TI - Sailing experience and sex as correlates of spatial ability. AB - The relationship between sailing experience and men's and women's spatial ability was examined by assessing the sailing history and Mental Rotations Test scores of 230 participants. The 102 men and 128 women came from three groups: college sailors (n =65), members of the general student body (n= 110), and college crew team members (n=55). Participants completed the Vandenberg and Kuse Mental Rotations Test and Lawton's Way-finding Strategy Scale and Spatial Anxiety Scale. Demographic variables and sailing experience were also assessed. Men scored significantly higher on the Mental Rotations Test than did women, and sailing team members scored significantly higher on that test than did student body members and crew team members. Results are discussed in terms of current explanations for sex differences in spatial ability. PMID- 15291233 TI - Reliability and stability of a dream recall frequency scale. AB - Dream recall frequency varies widely between people as well as within individuals. To explore the relationship between dream recall frequency and trait variables such as personality dimensions, a measure of stable interindividual differences is necessary. In the present study (N = 198 patients with sleep disorders; 115 women, 83 men; M age = 45.8 +/- 15.3 yr.) a high retest reliability of the 7-point Dream Recall Frequency scale developed by Schredl in 2002a was found. If the participants' focus was not directed explicitly towards dream recall when the scale was presented within a general sleep questionnaire, the hitherto-reported increase of dream recall due to measuring dream recall frequency did not occur. In conclusion, the present scale is well suited for measuring interindividual differences in dream recall frequency reliably. PMID- 15291234 TI - Report order and identification of multidimensional stimuli: a study of event related brain potentials. AB - An experiment was conducted to investigate the effect of order of report on multidimensional stimulus identification. Subjects were required to identify each two-dimensional symbol by pushing corresponding buttons on the keypad on which there were two columns representing the two dimensions. Order of report was manipulated for the dimension represented by the left or right column. Both behavioral data and event-related potentials were recorded from 14 college students. Behavioral data analysis showed that order of report had a significant effect on response times. Such results were consistent with those of previous studies. Analysis of event-related brain potentials showed significant differences in peak amplitude and mean amplitude at time windows of 120-250 msec. at Fz, F3, and F4 and of 350-750 msec. at Fz, F3, F4, Cz, and Pz. Data provided neurophysiological evidence that reporting dimensional values according to natural language habits was appropriate and less cognitively demanding. PMID- 15291235 TI - Seasons in dreams. AB - Based on the continuity hypothesis of dreaming, whether season-related themes are more often found in five dreams collected over a 2-wk. period in particular seasons was examined for 376 women and 68 men. Whereas for winter-related themes the continuity hypothesis was supported, the percentage of summer-related themes did not differ between dream samples collected in the winter or summer months, respectively. Researchers should include the amount of time spent with season related activities, conversations, and films to test whether in dreams the relation is direct between waking life and dreaming about seasons. PMID- 15291236 TI - Auditory event-related potentials in Parkinson's disease in relation to cognitive ability. AB - Auditory event-related potentials were evaluated in 45 nondemented patients with mild to moderate Parkinson's disease and 40 matched normal controls. All patients were neuropsychologically assessed by means of the Raven Colored Progressive Matrices, four subtests of the Wechsler Memory Scale (Digit Span Forward, Logical Memory, Visual Memory, Associate Learning), and the Wisconsin Card-sorting Test. The P300 component of the auditory event-related potentials was significantly prolonged in the patients with Parkinson's disease. Correlations between P300 latency and neuropsychological measures showed significant associations with lower performance on the Raven Colored Progressive Matrices and the Wisconsin Card-sorting Test. Our results indicate that for patients with mild to moderate Parkinson's disease subtle changes in cognitive abilities may be reflected as P300 prolongation. PMID- 15291237 TI - Throwing accuracy during prism adaptation: male advantage for throwing accuracy is independent of prism adaptation rate. AB - Previous studies have found that men are more accurate at throwing an object at a target than are women, independent of experience. However, these studies' results are based on average scores from multiple trials. As such, it is unknown whether the male advantage results from superior throwing accuracy or from a superior ability to calibrate subsequent throws. This study examined whether men can calibrate repeated throws more quickly and accurately than women, 25 men and 30 women were required to throw velcro-covered balls at a carpet-covered target, both with and without 10-diopter prism lenses. Participants had multiple trials in both conditions. Analyses examined whether there was a sex difference in the rate of adaptation to the prism lenses (as indicated by calibration of subsequent throws), instead of simply averaging all throwing accuracy scores and looking for an overall sex difference. Men threw the balls significantly more accurately than women, both with and without the prism lenses. However, there was no significant sex difference found on the rate of prism adaptation, as measured by improvement across the trials, i.e., calibration. Although men were more accurate at throwing balls overall, there was no sex difference in calibration of subsequent throws in adapting to the prism lenses, therefore indicating that the male advantage in throwing accuracy does not result from superior ability to calibrate subsequent throws but rather from superior throwing accuracy overall. PMID- 15291238 TI - [Functional regulation of serotonin transporter, a molecule involved in the pathogenesis of anxiety disorder]. AB - The serotonin transporter (SET) is a member of the Na+/Cl(-)-dependent neurotransmitter transporter family and functions as a membrane protein which terminates the serotonergic neuronal transmission by re-uptaking serotonin into the pre-synaptic terminal. SET is thought to be involved in the pathogenesis of affective disorders, drug abuse and anxiety disorder. We have focused on SET regulation by phosphorylation/dephosphorylation since SET has many putative phosphorylation sites in its intracellular region. Our previous studies have revealed that phorbolesters, activators of PKC, decreased in SET uptake activity. Based on a mutagenesis analysis of PKC phosphorylation sites and an in vivo phosphorylation study of SET, we have concluded that PKC regulates SET activity via an indirect mechanism, probably via alternating actin cytoskeleton status. Recent reports and our investigation have demonstrated that the SET C-terminal region interacts with actin binding proteins, suggesting the crucial roles of this region in functional regulation of SET. PMID- 15291239 TI - [Effects of anxiolytic drugs on rewarding and aversive behaviors induced by intracranial stimulation]. AB - In considering characteristics of action of anxiolytic drugs and the mechanism of drug action in the brain, it may be necessary to study not only the behavioral pharmacology but also the brain site. In the present study, anxiolytic drugs have been examined in various kinds of behaviors induced by stimulating the brain areas with regard to emotional expression such as reward (pleasure) or aversion in rats. First, the low rate responding on lateral hypothalamic self-stimulation behavior was induced by schedules of low current brain stimulation, variable interval (VI) and differential reinforcement of low rate (DRL). Anxiolytic drugs such as benzodiazepines facilitated these low rate responses. The drug susceptibility was high in the order of the low current stimulation > VI > DRL schedules. Furthermore, it was found by the auto-titration method on intracranial self-stimulation behavior that anxiolytic drugs decreased the threshold of stimulation reward. Second, it was recognized using the decremental lever pressing (DLP) paradigm that anxiolytic drugs increased the threshold of aversive stimulation of mesencephalic dorsal central gray (DCG), and this increasing effect of the drug was antagonized by GABA receptor blockers such as biccuculline. Third, it has been examined whether the conflict situation is established by combining with brain stimulation reward and aversion such as foot shock or DCG stimulation. As a result, the conflict behavior was established by combining with not only the brain stimulation reward and foot shock aversion, but also the brain stimulation reward and DCG stimulation aversion. Further anxiolytic drugs exhibited the anti-conflict action to both situations. The susceptibility of anxiolytic drugs to the conflict behavior by intracranial reward and aversion was higher than the conventional method based on milk reward and foot shock aversion. Indeed, in the present brain stimulation behavioral study, anxiolytic drugs such as benzodiazepines increased the stimulation threshold in lateral hypothalamic self-stimulation and inhibited the DCG aversive stimulation, i.e. resulting in an anticonflict action of the drugs. Recently we have preliminarily established a new model for evaluating the drug which may facilitate the motivation contributing to result in various behaviors, using the priming stimulation paradigm in intracranial self-stimulation behavior. Diazepam, benzodiazepine, and nomifensine, an dopamine uptake inhibitor, exhibited a delay of the extinction process which included non-reinforcing stimulation and pretrial electric stimulation (priming stimulation) in the self-stimulation behavior. This action may be related to the brain-mechanism of motivation. PMID- 15291240 TI - [Functional involvement of endogenous anxiogenic neuropeptide in brains]. AB - Neuronal GABA(A)/benzodiazepine and monoamine receptors participate in anxiety. Diazepam binding inhibitor (DBI), an endogenous anxiogenic neuropeptide, significantly increases in brains only after treatment with psychological stress, and this increase is completely abolished by benzodiazepines. Therefore, it is through that DBI may be involved in anxiogenesis produced by psychological stress. Furthermore, increases in cerebral DBI are observed in patients with several disorders accompanying anxiety and fear, which suggest that cerebral DBI may be an essential factor for anxiogenesis, and that it may be, at least in part, a biological index to evalulate anxiety. PMID- 15291241 TI - [Biological basis of anxiety disorders and serotonergic anxiolytics]. AB - Selective serotonin reuptake inhibitors (SSRIs) have wide indications for the treatment of anxiety disorders, including panic disorder, generalized anxiety disorder, posttraumatic stress disorder, obsessive-compulsive disorder and social anxiety disorder in addition to depression. Until recently, no animal model has been available for screening the anxiolytic effect of SSRIs and studying its mechanism of action. We have investigated the relationship between serotonin neurotransmission and anxiety using conditioned fear stress (CFS), an animal model of anxiety. CFS increased serotonin neurotransmission in the medial prefrontal cortex and amygdala. In behavioral pharmacological studies, SSRIs, serotonin1A agonists and monoamine oxidase inhibitors, which are assumed to facilitate serotonin neurotransmission, decreased conditioned freezing, an index of anxiety or fear, in CFS. In vivo microdialysis studies showed that serotonin neurotransmission in the medial prefrontal cortex increased after recovery from the freezing behavior. Microinjection of SSRI to the basolateral nucleus of the amygdala reduced conditioned freezing, indicating that the amygdala is one of target brain sites of anxiolytic action of SSRIs. Furthermore, CFS-induced c-Fos expression in the basolateral nucleus of the amygdala was reduced by SSRI pretreatment. Taken together, recent studies indicate that facilitation of brain serotonin neurotransmission decreases anxiety in agreement with the clinical evidence. PMID- 15291242 TI - [Recent trends in pharmacotherapy for anxiety disorders]. AB - Since the introduction of SSRIs, pharmacotherapy for anxiety disorders has significantly changed. Although the SSRIs are considered to be a first-line treatment for the most of anxiety disorders benzodiazepines are still widely used in clinical practice despite the risk of dependence and strong recommendation for their use as a second-line. The SSRIs only replaced tricyclic antidepressants and the MAO inhibitors especially in the treatment of panic disorder, obsessive compulsive disorder and social phobia. Combination of the SSRIs and the benzodiazepines is widely used. Recently it has been suggested that the combination of SSRI and benzodiazepine is rational, because each drug has a different mechanism of action, the benzodiazepines enhancing GABAergic transmission, and the SSRIs stimulating the 5-HT1A receptor that may inhibit the postsynaptic neuronal excitability in the amygdala and the prefrontal cortex that comprise the brain circuit of fear and anxiety. Recent imaging studies suggested the hyperactivity of the amygdala in the patients with generalized social anxiety disorder and successful treatment with cognitive behavioral therapy or SSRI might significantly reduce the hyperactivity of the amygdala. It was suggested that the rational combination of SSRIs and benzodiazepines seems to be an effective and practical way of treatment for most anxiety disorders. PMID- 15291243 TI - [Genetic basis of drug dependence and comorbid behavioral traits]. AB - Drug dependence is characterized by symptoms causing uncontrollable use of a drug despite its negative consequences. Dependence occurs only in a small fraction of individuals who try an addictive drug, and there is a large variance in individual susceptibility to dependence. Individuals susceptible to dependence exhibit specific comorbid behavioral traits, such as sensation seeking, novelty seeking, and antisocial personality. Studies using genetically engineered mice have delineated the extent to which various genes contribute to both dependence susceptibility and comorbid behavioral traits. Evidence suggests that the genes for dopamine D4 receptor, phosphodiesterease1B, the AMPA receptor subunit GluR1, 5HT1B receptor, protein kinase C and the transcription factor FosB contribute to both dependence susceptibility and comorbid behavioral traits. However, MAO-B influences a behavioral response to novelty without affecting nicotine dependence susceptibility. The mechanisms by which genes influence dependence susceptibility and comorbid behavioral traits are likely to be complex. PMID- 15291244 TI - [Major depressive disorders and BDNF (brain-derived neurotrophic factor)]. AB - We reviewed the pre-clinical and clinical papers demonstrating that BDNF plays a role in the pathophysiology of major depressive disorders. Experimental studies suggest that BDNF expression is induced by chronic antidepressant treatments, and that BDNF itself have antidepressant activity in animal models of depression. Furthermore, BDNF may protect neurons against stress-induced damage, and affect neurogenesis in the adult hippocampus. Our clinical study showed that serum levels of BDNF in drug naive patients with depression were significantly decreased as compared with normal controls. These findings suggest that low BDNF levels may play a pivotal role in the pathophysiology of major depression. It is unclear whether low BDNF levels are primary or secondary in patients with depression. One hypothesis would be that reduced BDNF might reflect a genetic vulnerability in patients with depression. Another possible explanation would be that stress-induced BDNF reductions might cause neuronal damage, which would in turn lead to acquired biological vulnerability. In order to determine the precise mechanism underlying the relationship between reduced BDNF levels and the etiology of major depression under both genetic and environmental backgrounds, further detailed study will be necessary. PMID- 15291245 TI - [Stress sensitization induced by stressor and methamphetamine]. AB - Repetitive or acute treatment of methamphetamine (MAP) or amphetamine (AMP) induces sensitization to both subsequent challenge treatment of the drugs, and exposure to emotional and physiological stress. In addition, chronic treatment of AMP enhanced DA utilization/release in striatum. Similarly, repetitive exposure to footshock or tail shock stress induces sensitization of noradrenaline or 3 methoxy-4-hydroxyphenylglycol (MHPG) to subsequent mild stress and to small amounts of AMP or MAP injection. Striatum, nucleus accumbens and prefrontal dopaminergic systems have an important role in the development of this sensitization. Immediate early gene (IEG) expression in the hypothalamus, nucleus accumbens and striatum may be involved in this process. Neurobiological vulnerability to schizophrenia may be induced by the interaction of multiple gene disposition and environmental insult, and schizophrenia onset and/or relapse in response to mild, non-specific stress. Stress-sensitive systems therefore are postulated in the pathophysiology of schizophrenia. In this regard, mesolimbic DA systems may be involved in the pathophysiology of schizophrenia. In contrast to MAP- or AMP- and stress-induced sensitization, haloperidol and clozapine induce IEG expression in the caudate-putamen and amygdala. Collectively, MAP- or AMP induced sensitization may, in part, share an early functional process of neurobiological mechanisms. PMID- 15291246 TI - [Diabetes attenuates the antidepressant-like effect mediated by the activation of 5-HT1A receptors in the mouse tail suspension test]. AB - Several lines of evidence have indicated that the prevalence of psychiatric disorders in diabetic subjects is higher than that in the general population, however, little information is available on the effects of antidepressants in diabetes. In the present review, we summarized the effect of diabetes on the central serotonergic systems and the efficacy of serotonergic antidepressants. Streptozotocin-induced diabetic mice showed prolonged duration of immobility compared to non-diabetic mice in the tail suspension test. This behavioral change was unrelated to the transient increases in blood glucose concentrations or decreased body weights by diabetes. Fluoxetine, a selective serotonin (5-HT) reuptake inhibitor, reduced the duration of immobility in both non-diabetic and diabetic mice. However, a selective 5-HT1A receptor antagonist WAY-100635 reversed the antidepressant-like effect of fluoxetine only in non-diabetic mice. In addition, a 5-HT1A receptor agonist 8-OH-DPAT reduced the duration of immobility in non-diabetic mice, but not in diabetic mice. These results suggest a possibility that the antidepressant-like effect mediated by the activation of 5 HT1A receptors may be attenuated by diabetes. PMID- 15291247 TI - [Central nervous system involvement in connective tissue diseases]. AB - Neuropsychiatric manifestations are relatively common and serious complications in systemic lupus erythematosus (CNS lupus). Overall, in patients with CNS lupus, CSF IgM, IgA, IgG indexes (indicators of intrathecal Ig synthesis) as well as CSF IL-6 activities were significantly elevated. Of note, especially in patients lupus psychosis, but not in those with focal CNS lesions, anti-ribosomal P antibody (anti-P) in the sera as well as anti-neuronal antibody (anti-N) in the CSF was significantly elevated in relation to their CNS disease activities. These data indicate that the immune system activation within the CNS, possibly resulting in the elevation of CSF anti-N, plays an important role in the pathogenesis of CNS lupus, including lupus psychosis. CNS involvement in Behcet's disease, usually called neuro-Behcet's syndrome (NB), includes acute type and chronic progressive type. Acute NB is characterized by acute meningoencephalitis with focal lesions, presenting high intensity areas in T2-weightened images or Flare images on MRI scans, whereas chronic progressive NB is characterized by intractable slowly progressive dementia, ataxia and dysarthria with persistent elevation of CSF IL-6 activity. Chronic progressive NB is resistant to conventional treatment with steroid, cyclophosphamide, or azathioprine, but responds to low dose methotrexate. As for ANCA-related vasculitis, pachymeningitis has been found to be associated with P-ANCA as well as C-ANCA. PMID- 15291248 TI - [Collagen diseases with pulmonary involvement]. AB - Pulmonary involvement is a common feature in patients with various collagen diseases. Some types of the pulmonary involvement are resistant to currently available treatment regimens and thus considered as intractable conditions. These include acute/subacute interstitial pneumonia in dermatomyositis, pulmonary interstitial fibrosis in scleroderma, and diffuse alveolar hemorrhage. Acute/subacute interstitial pneumonia with the histology of diffuse alveolar damage (DAD) is mainly occurred in patients with amyopathic or hypomyopathic dermatomyositis who lack autoantibodies to aminoacyl tRNA synthetases. Intensive immunosuppressive treatment in the early phase of the disease may be effective for this intractable complication. Nearly one-third of patients with scleroderma have slowly progressive pulmonary interstitial fibrosis, leading to end-stage respiratory failure. Non-specific interstitial pneumonia (NSIP) with an excessive fibrotic change is a major histology of these patients. There are accumulating evidences showing the effectiveness of cyclophosphamide in patients with this intractable condition, especially those with active alveolitis. Diffuse alveolar hemorrhage is a fatal complication mainly occurred in patients with systemic lupus erythematosus and those with antineutrophil cytoplasmic antibody (ANCA) associated vasculitis, including microscopic polyangitis (MPA) and Wegener's granulomatosus. Immediate diagnosis and introduction of intensive treatment are necessary to save the patients with this complication. PMID- 15291249 TI - [Collagen diseases with pulmonary hypertension]. PMID- 15291250 TI - [Intractable renal vasculitis by lupus nephritis and ANCA associated vasculitis]. PMID- 15291251 TI - [Collagen diseases with gastrointestinal manifestations]. AB - Collagen vascular diseases are known to present with a diverse array of gastrointestinal manifestations. These can be classified as: 1) gastrointestinal damage due to the collagen vascular disease itself; 2) adverse events caused by pharmacotherapies; or 3) gastrointestinal infections following immunosuppression due to corticosteroid (CS) administration. The first group includes lupus enteritis and protein-losing gastroenteropathy in systemic lupus erythematosus (SLE), reflux esophagitis, chronic intestinal pseudo-obstruction, and pneumatosis cystoids intestinalis in systemic sclerosis, amyloidosis in rheumatoid arthritis, bowel ulcer and bleeding in rheumatoid vasculitis and microscopic polyangiitis, and ileocecal ulcer in Behcet disease. In particular, colonic ulcers associated with SLE represent refractory lesions resistant to CS. Analysis of reported cases showing colonic lesions with SLE (22 cases in Japan) revealed that mean duration of SLE was 9.9 years and 77% of colonic lesions were observed in the rectum and sigmoid colon. Half of the patients developed intestinal perforation or penetration, and 6 of the 11 patients with perforation died. The second group includes lesions in the small and large intestine due to nonsteroidal anti inflammatory drugs (NSAIDs) and CSs, in addition to peptic ulcers. As perforation in CS-treated patients displays relatively high incidence with poor prognosis, careful attention to such complications is needed. The third group includes candidal esophagitis and cytomegalovirus (CMV) enteritis. Prompt diagnosis is required to prevent colonic bleeding and perforation due to CMV. PMID- 15291252 TI - [Collagen diseases with opportunistic infections]. AB - Treatment of opportunistic infections emerging in collagen diseases is very important as well as the therapy of original diseases. Lung tuberculosis, Pneumocystis carinii and lung fungal infections are main opportunistic infections. There is effective prophylaxis against them, though the cases for their administration should be carefully chosen because of their adverse effects. We have administrated INH and ST (Sulphomethoxazole and Trimethoprim) more than 15 years as the prophylaxis against tuberculosis and P. carinii to the cases who are treated more than 60 mg of prednisolone per day as an initial dose until less than 30 mg per day and completely succeeded. Infliximab treatment has been reported that it often induces tuberculosis in abroad and lots of occurrence was anticipated in Japan where high incidence of tuberculosis is observed. So far, however, there are only few patients, maybe due to the selection of the patients and actively utilizing prophylaxis. PMID- 15291253 TI - [A case of Behcet's disease with esophageal ulcers complicated with systemic sclerosis, chronic hepatitis C, and pancytopenia]. AB - A 62-year-old man was admitted to our hospital because of retrosternal burning pain and high fever in May, 2002. In 1995 chronic hepatitis C was diagnosed. Five years before admission he had been suffering from recurrent oral aphthous ulcers and genital ulcers. Distal scleroderma developed and the diagnosis of systemic sclerosis was made by skin biopsy in 1999. Prednisolone therapy, 5-30 mg/day, had been administered since then. In May 2000, he was referred to our department, and diagnosis of incomplete-type Behcet's disease was made because he had erythema nodosa, oral aphthous ulcers and genital ulcers. Asymptomatic mild pancytopenia was also found. In November 2000, gastrofiberscopy revealed that he had esophageal and gastric ulcers resistant to regular treatment and was diagnosed as entero-Behcet's disease, a subtype of the disease. The activity of esophageal and gastric ulcers was resistant to the low dose glucocorticoid and more than a moderate dose (30 mg/day) of prednisolone was necessary to reduce the activity. His gastrointestinal symptoms fluctuated with low dose prednisolone. Gastrofiberscopy on admission revealed that he had four shallow active oval ulcers in the middle-lower esophagus and distinct blind-fistula in the lower esophagus. Prednisolone were increased to 30 mg/day for his active entero Behcet's disease, however, his burning retrosternal pain remained. He died on the 81st hospital day due to severe pneumonia. This is a rare case of Behcet's disease complicated with esophageal ulcers, systemic sclerosis, chronic hepatitis C, and pancytopenia. Of interest is the mechanism of coincidence of these diseases from the pathological point of view. PMID- 15291254 TI - [A case of rheumatoid arthritis/Sjogren's syndrome with acute renal failure due to hyperuricemia associated with mizoribine therapy]. AB - A 64-year-old woman was diagnosed as having rheumatoid arthritis in 1999 at a nearby hospital. She had been treated with etodolac, actarit, mizoribine (MZ) and prednisolone. On May 25, 2001, she noticed fever and nausea and was treated with diclofenac sodium and clindamycin. On May 31, a nasal bleeding, tarry stool, hyperuricemia, renal dysfunction and thrombocytopenia developed and she was admitted to our hospital. Administration of drugs except prednisolone was stopped and hemodialysis was carried out on June 1. Fever and nausea improved during several days. Hyperuricemia and renal dysfunction disappeared on June 11. The platelet count became normal after platelet transfusion and she was discharged from our hospital on July 2. She was also diagnosed as having Sjogren's syndrome. In our case, a delay in MZ discharge by transient renal dysfunction might have caused a hyperuricemia, following an aggravation of renal dysfunction. So, care should be taken about latent renal dysfunction during the use of MZ. Moreover, it may be necessary to consider a discontinuation of MZ and administration of hemodialysis in the case of transient renal dysfunction. PMID- 15291255 TI - [A case of gastric mucosal bridge with Behcet's disease]. AB - A 69-year-old man visiting our hospital with an epigastralgia and tarry stool was diagnosed as having Behcet's disease on the basis of repetitious aphthous stomatitis, erythema nodosum and arthralgia in 1991. The next year, he suffered from double active ulcers in the antrum of the stomach, and he had been operated on for intestinal perforation. In 1994, endoscopic examination revealed the gastric mucosal bridge between the double ulcers. The double ulcer healed after an eradication therapy of H. pylori, but the gastric mucosal bridge has remained there on the gastrointestinal endoscopy. The gastric mucosal bridge with Behcet's disease has not been reported in Japan, being considered to be very rare. PMID- 15291256 TI - [Encouragement of attending congresses of the International Union of physiological Sciences]. PMID- 15291257 TI - [Coronary circulation]. PMID- 15291258 TI - [My research activity in the Calcium Signals laboratory of the Johns Hopkins University]. PMID- 15291259 TI - [My research activity in the Waggoner Center for Alcohol and Addiction Research]. PMID- 15291260 TI - [Comment on comparative physiology]. PMID- 15291264 TI - [Gender difference in alcoholic liver injury]. AB - Gender differences of alcoholic liver injury have been described previously, but mechanisms have only partially characterized. For example, it is known that females develop alcoholic liver injury more rapidly and to a greater extent than males. It now appears that estrogen participates in several aspects of this phenomenon. On the other hand, attention has been directed towards the effect of ethanol ingestion on Kupffer cell function, which is stimulated by gut-derived endotoxins via mechanisms dependent on increased gut permeability and the possible relationship between Kupffer cell and alcohol-induced liver injury. PMID- 15291266 TI - [Actual conditions of heavy drinkers in a community and measures against alcoholic problems]. AB - We surveyed the health conditions of people in an area, as a baseline survey for a local plan of Healthy Japan 21. Among the 2,469 subjects, we selected 40 ones, as a drinker group, who drank alcohol more than 3 days a week (more than 60 g a day), and 80 ones, as a control group, whose gender, age and resident area were matched to the drinker group. More subjects took no breakfast in the drinker group than in the control group. More smoked and felt stress in the drinker group. Subjective health conditions were worse in the drinker group. Half in the drinker group took no annual health examination. Half in the drinker group wanted to reduce or quit alcohol, but felt that they could not do so and that the dissolution of stress was necessary to do so. The both groups had low understanding on alcoholism and little information on supporting organizations for alcoholic problems. These results suggest that measures against alcoholic problems in the area are encouraging people to take an annual health examination, giving them substantial guidance after a health examination for drinkers and intervention to raise the understanding on alcoholism. PMID- 15291265 TI - [GC-MS analysis of methamphetamine and amphetamine in hair of Thai drug addicts]. AB - Psycho-stimulant dependence in young individuals has become a serious problem in Thailand, where consumption of the so-called YaBa methamphetamine tablets has become a fashionable trend. Due to its easy availability in the form of a tablet, young individuals abuse methamphetamine. Methamphetamine tablets are known to be potently addictive and its difficulty in cessation of drug use and to be abstinent from the drug. We herein report the results obtained from GC-MS analysis of methamphetamine and amphetamine in 33 samples of urine and hair from patients with psycho-stimulant dependence. These samples were collected from patients registered at the outpatient clinic in the Department of Psychiatry, Chiang Mai University Hospital and were sent to Nippon Medical School, Department of Legal Medicine (Tokyo, Japan) for further analysis by Dr. Werawan Ruangyttikarn, Department of Forensic Medicine, Chiang Mai University. Sample preparation: Hairs samples were cropped near the hair root. After washing, they were cut into 1.2-cm sections and extracted with methanol/5N HCl (2:1) for an hour and then, solid-phase extraction was conducted using Bond-Elut Certify. Following extraction, GC-MS analysis was performed. Urine samples were subjected to GC-MS analysis after preparation with Bond Elut Certify. RESULTS AND DISCUSSION: In 6 samples, both urine and hair samples analyzed were negative for detection of the stimulant drugs. In those cases individuals might stop taking drug for about 5 months. In 18 samples, urine samples were negative whereas hair samples were positive. These results suggest that individuals might stop using drugs for a few days before they went to the hospital but they abuse drugs continuously. In 9 samples, both urine and hair samples were positive. These results show that individuals always abuse drugs. In order to treat drug dependence effectively it is necessary to obtain the patient history of drug use and to evaluate and determine short-term and long-term drug use with urinalysis and hair analysis, respectively. Our present data revealed that useful information concerned with the long term drug abuse can be obtained from hair analysis, and that this method of analysis is applicable not only to forensic cases but also for evaluating clinical cases. PMID- 15291267 TI - [Drinking practice and alcohol-related problems: the national representative sample survey for Healthy Japan 21]. AB - This article is the first survey report in Japan concerning the adult drinking behaviors and its overall alcohol-related problems based on a national representative sample. It deals with a wide range of alcohol topics such as frequency, volume, cultural aspects, social influence and its effect on individuals, both positive and negative as well as various other alcohol related problems. The report identified, for further implementation of Japan's national health program, "Healthy Japan 21", the following areas of focus; 1) 5% of males and 0.5% of females who are considered "Hard Drinker", the most prioritized target. Seemingly small portion of females, 0.5%, is far beyond the planned goal level. 2) The cultural and regional competency is highly expected for policy discussion and evaluation in order to decrease the hard drinker population by 20% off. 3) Clarify the definition of a "Hard drinker" and disseminate the information publicly for functional health monitoring activity by local governments. PMID- 15291269 TI - 2-year review of a novel vestibular rehabilitation program in Montreal and Laval, Quebec. AB - OBJECTIVE: To evaluate the benefits of a vestibular rehabilitation program (VRP) in the Montreal-Laval area. DESIGN: The VRP was conceptualized by a panel of experts including otolaryngologists, physiotherapists, and researchers from McGill University and its teaching hospitals. From February 1999 to December 2001, 117 patients were seen, and 88 of them completed the VRP. SETTING: The VRP has been established at the Jewish Rehabilitation Hospital in Laval, PQ, to provide specialized rehabilitation to clients suffering from vertigo, dizziness, and/or impaired balance owing to lesions or disorders of the vestibular system. MAIN OUTCOME MEASURES: Presence or absence of nystagmus or vertigo during the Dix Hallpike test, Dizziness Handicap Inventory (DHI), and Dynamic Gait Index (DGI). RESULTS: Thirty-five patients with benign paroxysmal positional vertigo of the posterior canal were treated with canalith repositioning manoeuvres. All of the patients (100%) had absence of nystagmus or vertigo after one to four treatment sessions. Forty-six patients with vestibular deficits or dizziness-disequilibrium completed the VRP, which consisted mainly of individualized eye-head and balance home exercise programs. At the end of the VRP, there was a significant decrease in DHI score (31 vs 57; p < .01) and a significant increase in DGI score (18.4 vs 22.6; p < .01). CONCLUSIONS: A range of modest to major improvements was shown by comparing initial and discharge scores of patients who had completed vestibular rehabilitation. The VRP appears to be beneficial for patients with a variety of vestibular disorders. Further research is needed to continue optimizing vestibular rehabilitation. PMID- 15291268 TI - Thyroid colloid nodules diagnosed by fine-needle aspiration: efficacy of suppression. AB - BACKGROUND: This study reviewed the accuracy of fine-needle aspiration biopsy (FNAB) and the efficacy of thyroid suppression for colloid nodules in our population to determine the utility of these two modalities on the decision to operate. METHODS AND MATERIALS: A retrospective chart review of patients with colloid nodules diagnosed by FNAB from January 1993 to July 1995 was conducted. A 52-patient cohort underwent surgical management, and their needle aspirate cytologies and final pathologies were reviewed. RESULTS: A 7.7% false-negative rate in the detection of thyroid malignancy by FNAB was obtained. This is in keeping with data reported in the literature. Virtually no efficacy of hormonal suppression in our population was found. CONCLUSION: When the literature is reviewed and compared with the results of this study, the use of FNAB as a decision tool to operate is valid. The decision to operate based on the outcome of hormonal suppression, however, is not valid based on our results. PMID- 15291270 TI - Selective irrigation of the sinuses in the management of chronic rhinosinusitis refractory to medical therapy: a promising start. AB - BACKGROUND: Although endoscopic sinus surgery has been widely used for the treatment of chronic rhinosinusitis, some patients fail to derive clinical benefit from this procedure. We evaluated the efficacy of a treatment regimen consisting of selective irrigation of diseased sinus mucosa with topical antibiotics and steroids in conjunction with oral antibiotics and steroids. METHODS: Twenty patients suffering from chronic rhinosinusitis and resistant to medical treatment (mean duration 3.4 years) underwent intubations of the affected maxillary and/or ethmoid sinuses for irrigation for a duration of 21 to 30 days. A computed tomographic (CT) scan of the paranasal sinus was taken both pre- and post-treatment and staged according to the Lund-MacKay system. Clinical symptoms were scored for rhinorrhea, facial pain, nasal congestion, and smell at least 2 months prior to treatment and approximately 18 months after the follow-up. RESULTS: The clinical experience with the technique of intubation and irrigation was well tolerated by all patients. We found an improvement in all symptom scores, including rhinorrhea, nasal congestion, smell (n = 20; p < .001), and facial pain (n = 20; p < .01). Similar improvements were seen on the CT scans, with reduced staging from 14.6 +/- 1.1 to 5.6 +/- 1.1 (p < .001). Only three patients did not respond to selective irrigation of the sinuses and needed further surgery. CONCLUSION: These results suggest that sinus irrigation could provide a reasonable and effective alternative to ethmoidectomy with drainage procedures and offer promise for the treatment of patients with chronic rhinosinusitis who are resistant to medical treatment. PMID- 15291271 TI - Using Swaystar to measure sway amplitude in an office setting. AB - OBJECTIVE: The study was conducted to determine if Swaystar (University Hospital, Basel), a device that measures trunk sway, could be used in the office to detect abnormalities in patients who had vestibular abnormalities. DESIGN: Prospective study design. SETTING: Patients were referred to our tertiary and quaternary care dizziness clinic. METHODS: We measured total sway amplitudes using Swaystar during tandem walking of 18 patients referred sequentially who were identified as abnormal by caloric testing. These patients were compared with age- and sex matched subjects from our normative Swaystar databank. MAIN OUTCOME MEASURES: We measured total sway amplitude in pitch and roll planes with Swaystar during tandem walking testing with eyes open and eyes closed. RESULTS: Patients with caloric abnormalities had significantly higher total sway than the age- and sex matched normals from our normative databank. CONCLUSIONS: Swaystar is sensitive to measuring increased sway amplitude in patients with balance system deficits (as documented by caloric abnormalities) whether the patient is unstable in the pitch plane, roll plane, or both. PMID- 15291272 TI - Image-guided sinus surgery. AB - Image-guided sinus surgery is a relatively new technology used to track the position of endoscopic instruments placed within the nose or sinus cavities. The different types of image-guided technologies are reviewed. A review of our experience with 55 patients in whom this technology was used over a 10-month period is presented. Our interpretation of the usefulness and limitations of this technology is described. PMID- 15291273 TI - Benefits of cochlear implantation in early-deafened adults: the Toronto experience. AB - OBJECTIVE: To present the results of a survey administered to a group of early deafened cochlear implants adults and to report the level of perceived benefit. DESIGN: Prospective. SETTING: Large tertiary referral centre. METHOD: A 47-item questionnaire designed to evaluate cochlear implant use and benefit was sent to 42 early-deafened adult cochlear implant users. The questionnaire can be divided into seven subcategories: time of use, associated symptoms, communication, employment status and function, socialization, perceived benefit, and the impact on quality of life. Responses from 30 patients were received. RESULTS: The majority of our patients use their cochlear implant all of their waking hours. The majority of patients continue to depend on lip-reading and hearing as their main mode of communication, although they reported improved lip-reading skills with their cochlear implant. Twenty-three patients (76.7%) were employed. Eleven patients had a change in employment subsequent to cochlear implantation, nine (81.8%) of whom attributed this to their cochlear implant. Our patients als reported greater independence, a greater sense of safety in their environment, and an improved social life. Twenty-nine patients (96.7%) said that they were satisfied with their implant, 28 (93.3%) said that they would go through the same process again, and 27 (90%) said that they would recommend it to a friend in a similar situation. Twenty-nine patients (96.7%) stated that the cochlear implant has had a positive effect on their quality of life. Family and peer support, prior auditory-verbal therapy, and a positive attitude were the most commonly cited factors in successful cochlear implant use. CONCLUSIONS: Early-deafened adult cochlear implant users perceive significant benefit from cochlear implantation. Importantly, family and peer support, prior auditory-verbal therapy, and a positive attitude are considered important factors in maximizing this benefit. PMID- 15291274 TI - Sinonasal undifferentiated carcinoma: case series and systematic review of the literature. AB - Sinonasal undifferentiated carcinoma is a rare, highly aggressive malignancy. A number of case series have been published in the literature. Most authors recommend aggressive management with a combination of surgery, radiotherapy, and chemotherapy, but the numbers in the individual studies are too small to produce a definitive opinion on the standard of care. In an attempt to determine the optimal treatment for this condition, we have undertaken a systematic review of the literature to evaluate all cases of sinonasal undifferentiated carcinoma that have been published since its initial description in 1986. Patient demographics, extent of the tumour at presentation, management, and outcomes were evaluated. We also present the experience from our institution. PMID- 15291275 TI - Foreign body aspiration in infants and toddlers: recent trends in British Columbia. AB - OBJECTIVES: The objectives of this study were to (1) examine recent trends in the demographics and presentation of children with foreign body aspiration at British Columbia's Children's Hospital and (2) develop safety guidelines regarding feeding nuts and other hard, crunchy foods to infants and toddlers. METHODS: The methods used were a retrospective chart review and a review of swallowing mechanics in early childhood. RESULTS: Between July 1997 and July 2001, 51 children under 3 years of age underwent rigid bronchoscopy for suspected foreign body aspiration. Of these patients, 27 (53%) were 18 months of age or younger. Of these 27 infants and toddlers, 24 (89%) had a witnessed choking event and 22 (81%) had an airway foreign body. Nuts, raw carrots, and popcorn kernels accounted for 14 (64%) of the foreign bodies aspirated by these infants and toddlers. Before 2 years of age, children are poorly equipped to grind and swallow hard, crunchy food because they lack second molars and are still adjusting to the descent of the larynx. CONCLUSIONS: Infants and toddlers in British Columbia have been aspirating foreign bodies at an alarmingly high rate. Most cases would have been prevented with better public awareness. Caregivers should be informed that children under 3 years of age should never be fed nuts or other hard, crunchy foods. A public awareness campaign is warranted. PMID- 15291276 TI - Pediatric laryngeal paralysis: a new proposed surgical therapy. AB - OBJECTIVES: The cricothyroid muscle (CTM) has a separate innervation from that of intrinsic laryngeal muscles; therefore, its action may contribute to airflow resistance in children with laryngeal paralysis (LP) secondary to recurrent laryngeal nerve (RLN) palsy. We proposed removal of the CTM as a means of indirectly widening the paralyzed neonatal glottis. METHODS: A prospective study was conducted using a piglet animal model to simulate LP and evaluate the proposed treatment's outcome. LP was induced via bilateral RLN sectioning in seven piglets. The CTMs were then removed. Animals acted as their own controls. Outcome measures consisted of serial inspiratory and expiratory airflow resistance measurements taken (1) with no intervention, (2) after RLN sectioning, and (3) after CTM removal. Several animals were awakened to assess their clinical responses to the interventions. The paired Student's t-test was used for statistical analysis. RESULTS: Inspiratory airflow resistance was significantly increased by RLN sectioning (p = .0062) and then significantly decreased after subsequent CTM removal (p = .0005). Clinical responses to the interventions mirrored the measured findings. CONCLUSIONS: Removal of the CTM significantly decreases inspiratory airway resistance in piglets with induced LP. This proposed surgical therapy for pediatric bilateral LP warrants further investigation. PMID- 15291277 TI - The use of middle meatal stents post-endoscopic sinus surgery. PMID- 15291278 TI - Tongue necrosis, temporal arteritis, and esophageal carcinoma: is there a connection? PMID- 15291279 TI - Capsaicin in burning mouth syndrome: titration strategies. PMID- 15291280 TI - Toxoplasma neck mass in a pregnant woman: diagnosis and management. PMID- 15291281 TI - Dysphonia as the initial symptom of myasthenia gravis. PMID- 15291282 TI - Giant congenital cholesteatoma of the middle ear as a cause of temporomandibular joint dysfunction. PMID- 15291283 TI - Cervical hematoma causing airway compromise after traumatic laceration of facial artery. PMID- 15291284 TI - Lung carcinoma metastatic to the masseter muscle. PMID- 15291285 TI - [Perspectives on obstetrics and special consideration on "midwife obstetrics"]. PMID- 15291286 TI - [Is a new ethics required for biomedical progress]. PMID- 15291287 TI - Challenges to potassium metabolism: internal distribution and external balance. AB - A complex pump-leak system involving both active and passive transport mechanisms is responsible for the appropriate distribution of potassium (K) between the intra- and extracellular fluid compartments. In addition, the kidneys, and to a lesser extent the colon, safeguard maintenance of the narrow range of low K concentrations in the extracellular fluid. Early renal clearance studies showed that K is normally both reabsorbed and secreted by renal tubules, and that regulated secretion is the major source of K excretion. Net K secretion occurs mainly in principal cells while K absorption takes place in intercalated cells. Studies on single tubules and principal and intercalated cells have defined the determinants of K secretion and reabsorption including the electrochemical driving forces, specific carriers, ATPases, and K channels. Recent studies on the properties and molecular identity of renal K channels have also contributed significantly to understanding the renal mechanisms that transport and regulate K excretion. PMID- 15291288 TI - Prevalence and management of anaemia in haematologic cancer patients receiving cyclic nonplatinum chemotherapy: results of a prospective national chart survey. AB - OBJECTIVES: Anaemia is common in patients with haematologic malignancies. In contrast to solid tumours there are only a few studies exploring anaemia in haematologic cancers. The aim of this study was to determine the prevalence of anaemia (haemoglobin [Hb] <12 g/dL) in patients with chronic lymphocytic leukemia (CLL), multiple myeloma (MM), non-Hodgkin's lymphoma (NHL), and Hodgkin's disease (HD) who were scheduled to receive cyclic chemotherapy. Predictive factors for anaemia development and anaemia treatment were also assessed. METHODS: This prospective chart survey was conducted at 35 oncology centers in Austria. A total of 273 patients were followed through four cycles of nonplatinum chemotherapy, and Hb-levels and anaemia therapy were documented. RESULTS: At baseline, prevalence of anaemia was greatest in patients with MM (77.4%). Prevalence of anaemia increased for all malignancies after cycle 4, with the largest increases noted for patients with NHL (from 35.1% at baseline to 73.7%) and HD (from 21.9% to 54.5%). Cyclic chemotherapy and prior anticancer treatment indicated an increased risk for developing anaemia. Notably, 27.5% of patients with Hb levels <10.5 g/dL remained untreated. Transfusions were most often given to patients with severe anaemia (Hb < 8 g/dL), and erythropoietin most often given to patients with mild or moderate anaemia. CONCLUSIONS: Our data confirm that anaemia prevalence in patients with haematologic malignancies is high and increases with chemotherapy. The current practice of anaemia management in these patients leaves room for improvement. PMID- 15291289 TI - The importance of preoperative localisation procedures in organic hyperinsulinism -experience in 67 patients. AB - BACKGROUND: Preoperative localisation of insulinomas has been regarded unnecessary, given the significantly higher detection rates of intraoperative ultrasonography and bidigital palpation. These are mandatory before endoscopic surgery. METHODS: 67 patients operated on for organic hyperinsulinism were retrospectively analysed regarding tumour localisation within the pancreas, tumour size, histological findings, sensitivities of preoperative imaging methods, and surgical techniques. RESULTS: 59 patients (88%) had solitary insulinomas, four patients (6%) multiple insulinomas and four adult patients (6%) nesidioblastosis. Well-differentiated neuroendocrine tumours with benign behaviour (including four patients with nesidioblastosis) were diagnosed in 53 patients (79%), tumours with uncertain behaviour in nine patients (13%) and well differentiated neuroendocrine carcinomas in five (8%). Tumours were evenly distributed throughout the pancreas. Endoscopic ultrasound localised tumours in 15 out of 21 patients (71%), conventional computed tomography (CT) in 7 out of 21 (33%), single-slice helical CT in 7 out of 12 (58%), multidetector CT in 5 out of 5 (100%), magnetic resonance imaging in 11 out of 13 (85%) and angiography in 15 out of 23 (65%). Various combinations of available methods achieved a sensitivity of 88% (49 patients true positive, 4 true negative, 7 false negative). Of 59 patients, solitary insulinomas were enucleated in 47 (80%), 11 patients underwent conventional open resection and one patient endoscopic distal pancreatic resection. Patients with nesidioblastosis or multiple tumours underwent pancreatic resections alone or in combination with enucleations. CONCLUSION: After biochemical diagnosis of organic hyperinsulinism, preoperative localisation is necessary for planning endoscopic pancreatic surgery, because of the possibility of multiple insulinomas, malignancy or nesidioblastosis in adults. PMID- 15291290 TI - Influence of the birth attendant on maternal and neonatal outcomes during normal vaginal delivery: a comparison between midwife and physician management. AB - BACKGROUND: The purpose of this study was to compare the obstetric outcome of low risk maternity patients attended by certified midwives with that of low-risk maternity patients attended by obstetricians. PATIENTS AND METHODS: Obstetric outcome of 1352 midwife patients was compared with that of 1352 age- and parity matched physician patients with normal spontaneous vaginal delivery at the Department of Obstetrics and Gynecology of the University Hospital Vienna during the period from January 1997 to July 2002. Our analysis was restricted to a sample of low-risk pregnant women. Women with medical or obstetric risk factors were excluded. RESULTS: A significant decrease in the use of oxytocin (p=0.0001) was observed in women who selected a midwife as their primary birth attendant compared with women in the physician group. In both groups most women gave birth in a supine position; however, significantly more alternative birth positions were used by midwife patients (p = 0.0001). Concerning perineal trauma, a significantly lower rate of episiotomies (p = 0.0001) and perineal tears of all degrees (p=0.006) were found in midwife patients. When analyzing severe postpartum hemorrhage and postpartum infections, there were no significant differences between the two groups (p > 0.05). Concerning neonatal outcome, there were no significant differences in APGAR score < 7 at 5 minutes (p > 0.05). Our data clearly show the ability of certified midwives to successfully provide prenatal care and delivery to low-risk maternity patients, with neonatal outcomes comparable to those of physician patients. The use of certified midwives supervised by obstetricians may provide the optimum model for perinatal care, particularly for those women who are low-risk maternity patients, leaving physicians free to attend to the high-risk elements of care. PMID- 15291291 TI - Increase of serum triiodothyronine concentration in soldiers with combat-related chronic post-traumatic stress disorder with or without alcohol dependence. AB - BACKGROUND: Post-traumatic stress disorder (PTSD) is a relatively new psychiatric disorder with three clusters of symptoms: trauma re-experiencing, avoidance, and increased arousal. The condition develops after a person sees, is involved in, or hears of an extreme traumatic stressor such as war, torture, natural catastrophe, assault, rape, or serious accident. PTSD is also often comorbid with other psychiatric disorders, especially with alcohol dependence. Several hormonal alterations have been reported in veterans with combat-related PTSD, including elevations in certain thyroid hormones, e.g., total T3; however, previous studies have not controlled for alcohol dependence, a common comorbid psychiatric disorder in this population. OBJECTIVE: The first aim of our study was to assess possible differences in basal serum levels of free triiodothyronine (FT3), total triiodothyronine (TT3), free thyroxine (FT4), total thyroxine (TT4), and thyroid stimulating hormone (TSH) in Croatian soldiers with combat-related chronic PTSD alone or comorbid with alcohol dependence and in healthy controls. The second purpose of the study was to determine any correlation between duration of combat activities, number of combat traumas, intensity and duration of PTSD symptoms, and serum levels of TT3, FT3, TT4, FT4, and TSH in this sample. METHOD: We analyzed basal serum FT3, TT3, FT4, TT4, and TSH concentrations in soldiers with combat-related chronic PTSD (N=43), combat-related chronic PTSD comorbid with alcohol dependence (N = 41), and in healthy controls (N = 39) using a luminoimmunochemical assay. RESULTS: Soldiers with chronic combat-related PTSD with or without comorbid alcohol addiction had significantly higher values of TT3 than the control group (F = 19.556, p<0.01). There was a significant correlation between TT3 levels and number of traumatic events in both the PTSD group (r=0.663, p<0.01) and those with PTSD comorbid with alcohol dependence (r=0.836, p<0.01). There was also a significant correlation between TT3 levels and symptoms of increased arousal in both PTSD (r=0.419, p<0.01) and PTSD comorbid with alcohol dependence (r=0.516, p<0.01). CONCLUSION: Elevated concentrations of serum TT3 are associated with combat-related PTSD, regardless of its comorbidity with alcohol dependence, and also with the number of traumatic events and symptoms of increased arousal. Given that current pharmacotherapy for PTSD is inadequate, reduction of TT3 may be a new strategy for pharmacologic intervention that could contribute to more effective treatment of this disorder. PMID- 15291292 TI - The anatomic variability of the coronary vasculature of the human heart--part I: functional vascular zones--a morphological view. AB - The myocardium may be divided into distinct zones which act as functional units of the coronary vascular network. The basic principle of this functional systematization of cardiac blood vessels is a constant relationship between the "distributing" and the "delivering" vessels. Six arterial zones of the myocardium are identified under this scheme: anterior right ventricular, lateral left ventricular, posterior left ventricular, interventricular septal, posterior right ventricular, and atrial. The distributing vessels of functional myocardial zones are most frequently both arterial and venous. However, between the lateral and posterior left ventricular zones there is sometimes only the left marginal vein (24%) or left marginal artery (20%). Between the atrial and posterior right ventricular zones there is sometimes only the right coronary artery (52%), or only the small cardiac vein (14%) as a distributing vessel. Between the left and the right posterior ventricular zones there is sometimes an arterial and sometimes a venous distributing vessel (10%). Between the posterior and anterior right ventricular zones there is either the artery (21%) or the right marginal vein (8%) as a distributing vessel. The permanence of the six functional myocardial zones is determined not only by the arterial but also by the venous distributing vessels, and in some cases the venous vessels are the only distributing vessels between certain zones. PMID- 15291293 TI - The anatomic variability of the coronary vasculature of the human heart--part II: some anatomical peculiarities of arteriovenous relations. AB - The arteriovenous relations in human heart are, in some instances, different from arteriovenous relations in other parts of the body. The specific relations between cardiac arteries and veins may enable diffusible substances carried through the system of juxta-arterial cardiac veins to influence the regulation of the lumen of the coronary arteries. Arteriovenous anastomoses (6% of our 150 cases) permit direct communication between the arteries and veins bypassing the capillary circulation; it is assumed that these anastomoses prevent coagulation of blood in small veins. In cases of arterial occlusion, the myocardium is supplied by veins that allow retrograde vascularization of the myocardium. In 33% of our cases the posterior atrial branches (0.5-1.0 mm in diameter) of the coronary arteries ran through the wall of the coronary venous sinus on their way from the parent vessel, which lay in the coronary sulcus, to the left atrium. In 11% of the cases, the arterial branch that ran through the distal portion of the wall of the coronary sinus was the interatrial branch. The blood flow through the parietal arteries of the venous coronary sinus probably depends on the condition of the muscular layer of the sinus during the phases of cardiac action, and this might be important in the course of certain cardiosurgical procedures. PMID- 15291294 TI - Epidemiology and outcome of pediatric trauma treated by an emergency-physician staffed advanced life-support unit. AB - BACKGROUND: The aim of this study was to describe the epidemiology and outcome of pediatric trauma in the setting of an emergency-physician-staffed mobile advanced life support (ALS) unit serving a predominantly urban area in Austria. METHODS: In this retrospective chart review, all pediatric trauma patients (0-14 years of age) who were treated by a physician-staffed ALS unit in Innsbruck within a 3 year period were analyzed. In addition, hospital charts were assessed to determine the clinical course and the outcome of these patients. RESULTS: 113 injured children were treated by the physician-staffed ALS unit (1.5% of all runs) during the study period; a frequency of three pediatric trauma patients per month. On average, injuries were of moderate severity (2.6 +/- 1.3 on the NACA severity scale). Thirteen children (11.5%) sustained severe to life-threatening injuries and two of whom underwent out-of-hospital resuscitation. The majority of the injuries were caused by vehicular accidents and sports/recreation-related trauma; head trauma was the most frequent injury. Violence-related trauma including weapon-inflicted injuries was uncommon. 40% of the children were hospitalized. The overall outcome was favorable: 78% of the hospitalized children had no impairment at the time of discharge. By comparing the prehospital trauma diagnosis with the final diagnosis, we found that the vast majority of emergency physician trauma diagnoses were accurate. CONCLUSION: Because the frequency of pediatric trauma is so low, ALS units may not gain adequate experience in the management of (severe) pediatric trauma, thus rendering regular training of paramount importance. PMID- 15291295 TI - Sternal osteomyelitis complicating percutaneous coronary artery stenting. AB - Hematogenous sternal osteomyelitis is a rare infection that has been associated with i.v drug abuse and blunt thoracic trauma, but iatrogenic infections have also been described following resuscitation and in conjunction with hemodialysis catheters. Staphylococcus aureus is the most common causative organism. Stenting is the preferred treatment for isolated stenosis of the coronary artery and is associated with a low complication rate and high patency rate. Such intravascular procedures are rarely complicated by infections. A 72-year-old man developed hematogenous sternal osteomyelitis following coronary artery stenting. Radiological diagnosis was made using CT scan and MRI, and blood cultures and aspiration fluid from the infected soft tissue were positive for S. aureus. Initial therapy consisted of i.v. second-generation cephalosporin followed by oral cephalexin and later linezolid. Prolonged antibiotic therapy without surgical intervention was successful in controlling this rare complication. The patient was well at the one-year follow up, with patent stent and no signs for recurrent osteomyelitis. Although transcutaneous stenting is a widely accepted strategy for treating stenosed arteries, this case highlights the possibility of hazardous infectious complications associated with such procedures. PMID- 15291296 TI - Anthroposophical medicine: a systematic review of randomised clinical trials. PMID- 15291297 TI - Redox photochemistry of thiouredopyrenetrisulfonate. AB - 1-Thiouredopyrene-3,6,8-trisulfonate (TUPS) has recently been used as a photoinduced covalent redox label capable of reducing various cofactors of proteins. A new reaction of this dye, whereby its excited triplet state oxidizes suitable electron donors, is now reported. The characteristic difference spectrum of the reduced radical of TUPS is determined. We also observe the self-exchange electron transfer between two TUPS molecules in their triplet excited states and determine the reaction scheme and the rate constants of the various pathways in the process of triplet depletion. The ability of photoexcited TUPS to withdraw an electron from reduced cytochrome-c is also observed. It is thus demonstrated that TUPS is an appropriate photoinduced covalent redox label for initiating both the oxidative and reductive phases of electron transfer processes in biological macromolecules. PMID- 15291299 TI - Matrix metalloproteinase-1 production observed after solar-simulated radiation exposure is assumed by dermal fibroblasts but involves a paracrine activation through epidermal keratinocytes. AB - Chronic exposure of human skin to solar UV radiation leads to serious dermal damages, a hallmark of photoaging. In vivo, acute UV radiation has been shown previously to induce various matrix-degrading proteases. Among them, matrix metalloproteinase-1 (MMP-1) has been suggested to be involved in skin photodamage. The purpose of this study was to investigate the effects of solar simulated radiation (SSR) on MMP-1 production in normal human skin cells. SSR exposure of human skin reconstructed in vitro comprising both a differentiated epidermis and a fibroblast-populated dermal equivalent led to an increase in MMP 1 production, which was abolished when epidermis was removed immediately after SSR exposure. In addition, SSR exposure of differentiated keratinocytes grown on an acellular collagen gel did not induce MMP-1 production. Experiments on cell cultures grown on plastic confirmed that keratinocytes failed, in contrast with fibroblasts, to produce MMP-1 in response to SSR exposure. However, when conditioned medium from SSR-exposed keratinocytes was added to human fibroblasts in culture, MMP-1 production was induced. Altogether, these data show that MMP-1 production observed after SSR exposure involved the release of soluble epidermal factors, which could modulate its production by dermal fibroblasts. PMID- 15291298 TI - Angiogenesis induced by photodynamic therapy in normal rat brains. AB - Angiogenesis promotes tumor growth and invasiveness in brain. Because brain injury often induces expression of angiogenic-promoting molecules, we hypothesize that oxidative insult induced by photodynamic therapy (PDT) could lead to an endogenous angiogenic response, possibly diminishing the efficacy of PDT treatment of tumors. Therefore, we sought to establish whether PDT induced an angiogenic response within the nontumored brain. PDT using Photofrin as a sensitizer at an optical dose of 140 J/cm2 was performed on normal rat brain (n = 30). Animals were sacrificed at 24 h, and 1, 2, 3 and 6 weeks after PDT treatment. Fluorescein isothiocyanatedextran perfusion was performed, and brains were fixed for immunohistological study. Immunostaining revealed that vascular endothelial growth factor (VEGF) expression increased within the PDT-treated hemisphere 1 week after treatment and remained elevated for 6 weeks. Three dimensional morphologic analysis of vasculature within PDT-treated and contralateral brain demonstrated PDT-induced angiogenesis, as indicated by a significant increase in vessel connectivity (P < 0.001) concomitant with decreased (P < 0.05) mean segment length compared with vessels within the contralateral hemisphere. Volumetric measurement of angiogenic regions indicate that neovascular expansion continued for 4 weeks after PDT. These data demonstrate that PDT induces VEGF expression and neovascularization within normal brain. Because angiogenesis promotes growth and invasiveness of tumor, antagonizing this endogenous angiogenic response to PDT may present a practical means to enhance the efficacy of PDT. PMID- 15291301 TI - Photovoltaic effects in hornets are correlated with the time of day. AB - This study deals with voltage values recorded off the cuticle of live specimens of the Oriental hornet Vespa orientalis (Hymenoptera, Vespidae). The relevant measurements were taken between the two tips of their bodies at various hours of the day and were made on a total of 90 worker hornets. Recorded voltage values varied within a range of 60-180 mV, with the lower values measured during the morning hours and the afternoon and the highest values during the noon hours. Measurements were made by direct contact of the electrodes with the hornet cuticle and did not prove lethal to the measured specimens. An additional 60 live hornets were measured in the same fashion but in the dark. The values recorded in the dark varied between 40 and 70 mV and displayed considerable fluctuations but were not found to be dependent on the time of measurement. The distribution of the voltage values in hornets measured at various hours in the daytime closely resembled that of the global radiations (in W/m2) on the same days the measurements were taken. PMID- 15291300 TI - Inhibition of DNA polymerization and antifungal specificity of furanocoumarins present in traditional medicines. AB - Antifungal activity is positively correlated to furanocoumarin content in extracts of the traditional phytomedicine northern prickly ash (Zanthoxylum americanum Mill. [Rutaceae]). The specificity of these furanocoumarins in inhibiting replication of DNA was investigated with reference to significant base composition differences between fungal and mammalian mitochondrial DNA. We developed a polymerase chain reaction-based assay to investigate whether (1) furanocoumarins inhibit DNA polymerization and (2) distinct furanocoumarins specifically inhibit DNA replication depending on base composition. Specific inhibition of DNA polymerization by 5-methoxypsoralen and psoralen through high adenine and thymine (AT) (84.3%) and low-AT (51.9%) DNA, respectively, suggests that furanocoumarins inhibit replicative functions of genomes or of regions within the genome that differ in base composition. Greater overall inhibition of DNA polymerization by Z. americanum husk extracts than with single or mixed furanocoumarins suggests that inhibitory compounds in addition to the major furanocoumarins are present in Z. americanum. PMID- 15291302 TI - A monochromatic action spectrum for the photoinduction of the UV-absorbing mycosporine-like amino acid shinorine in the red alga Chondrus crispus. AB - To determine the action spectrum for photoinduction of the ultraviolet (UV) absorbing mycosporine-like amino acid shinorine, specimens of the marine red alga Chondrus crispus were irradiated with monochromatic light of various wavelengths using the Okazaki large spectrograph at the National Institute for Basic Biology, Okazaki, Japan. Fluence response curves were determined for the wavelengths between 280 and 750 nm, by irradiating the algae with monochromatic light for 10 h, followed by 4 h of 25 micromol m(-2) s(-1) photosynthetically active radiation and 10 h darkness. Samples were taken after the second exposure interval. A linear correlation between fluence rate and accumulated shinorine concentration was detected for wavelengths between 350 and 490 nm in the fluence rate range of 20-30 micromol m(-2) s(-1), whereas there was no induction above 490 nm. Below 350 nm a decline in shinorine concentration could be observed at fluence rates above 30 micromol m(-2) s(-1), probably due to an inhibition of photosynthetic activity and a subsequent impairment of shinorine biosynthesis. The constructed action spectrum indicated that the photoreceptors mediating shinorine photoinduction might be an unidentified UV-A-type photoreceptor with absorption peaks at 320, 340 and 400 nm. PMID- 15291303 TI - Assessment of photosensitizer dosimetry and tissue damage assay for photodynamic therapy in advanced-stage tumors. AB - Photodynamic therapy (PDT) efficacy is a complex function of tissue sensitivity, photosensitizer (PS) uptake, tissue oxygen concentration, delivered light dose and some other parameters. To better understand the mechanisms and optimization of PDT treatment, we assessed two techniques for quantifying tissue PS concentration and two methods for quantifying pathological tumor damage. The two methods used to determine tissue PS concentration kinetic were in vivo fluorescence probe and ex vivo chemical extraction. Both methods show that the highest tumor to normal tissue PS uptake ratio appears 4 h after PS administration. Two different histopathologic techniques were used to quantify tumor and normal tissue damage. A planimetry assessment of regional tumor necrosis demonstrated a linear relationship with increasing light dose. However, in large murine tumors this finding was complicated by the presence of significant spontaneous necrosis. A second method (densitometry) assessed cell death by nuclear size and density. With some exceptions the densitometry method generally supported the planimetry results. Although the densitometry method is potentially more accurate, it has greater potential subjectivity. Finally, our research suggests that the tools or methods we are studying for quantifying PS levels and tissue damage are necessary for the understanding of PDT effect and therapeutic ratio in experimental in vivo tumor research. PMID- 15291304 TI - Cooperative effects in the photophysical properties of self-associated triguanosine diphosphates. AB - The present study deals with the photophysical properties of triguanosine diphosphate in aqueous solutions, which are compared with those of the 2' deoxyguanosine monophosphate. They are studied by steady-state absorption and fluorescence spectroscopy as well as by time-resolved fluorescence spectroscopy with femtosecond resolution. The temperature, salt and concentration dependence of the absorption and fluorescence spectra reveal that association of the trimers takes place. The resulting aggregates could correspond to a tetraplex structure. The aggregate fluorescence quantum yield is higher and the fluorescence lifetime much longer than those of the monomer. These results show the interaction between guanosine residues that may manifest itself via self-solvation, hydrogen bonding and/or delocalization of the excitation. PMID- 15291305 TI - Optical spectroscopy of hydrophobic sunscreen molecules adsorbed to dielectric nanospheres. AB - Fluorescence and absorption spectra of hydrophobic sunscreens, weakly fluorescent octyl methoxycinnamate, moderately fluorescent octyl salicylate and highly fluorescent 2-ethylhexyl-4-(dimethylamino)benzoate (padimate O) adsorbed to dielectric microspheres in aqueous suspension, have been compared with spectra in organic solution. The fluorescence of adsorbed salicylate and padimate is enhanced compared with fluorescence in methanol: about a factor of 6 and 30 in terms of fluorescence yield per molecule of salicylate and padimate, respectively. Cinnamate, which has a low fluorescence yield, does not show a comparable fluorescence enhancement. The fluorescence amplification is independent of sphere diameter from 30 to 1500 nm, at least for salicylate. The enhancement, as well as the location of absorption spectral peaks, is consistent with a low-dielectric constant environment of the molecules, in spite of the presumed location near the interface between polystyrene (epsilon = 2.4-3.8) and water (epsilon = 78). The adsorbed state of these sunscreens represents a proposed improved in vitro model for the environment of sunscreens in vivo, as well as a general model for chromophores in heterogeneous environments. PMID- 15291306 TI - Determination of the minimal erythema dose and colorimetric measurements as indicators of skin sensitivity to UV-B radiation. AB - There is a strong relation between chronic UV-B-induced sunburns and the development of skin cancer. Therefore, it is important to obtain a method that can be reproduced easily to detect individuals with similar skin color but different sensitiveness to sun exposure. The study evaluated 193 healthy volunteers (68% women; the average age was 38 years). They were divided into six groups of at least 30 subjects, according to skin type. The minimal erythema dose (MED) was assessed in two non-sun-exposed areas (thorax-infra-axillary area and on the buttocks), using a UV-B source (0.5 mW/cm2), with openings of 1 cm2, in increasing doses. The same areas were evaluated with a Minolta CR 300 Chromameter (L*a*b* system). The MED values ranged from 13 to 156 mJ/cm2; the coordinate L* (brightness) ranged from 75.96 to 30.15. The correlation between the MED and the brightness was negative in both areas (Pearson's correlation r = -0.91, P < 0.05). Color measurements, especially brightness, can be used to quickly assess skin sensibility. Considering the MED, there is a substantial overlapping of adjacent phototypes, but they could be separated into two groups: more sensitive individuals (Types I, II, III and IV) and less sensitive ones (Types V and VI). PMID- 15291307 TI - 5-aminolevulinic acid-based photodynamic therapy in leukemia cell HL60. AB - A study to explore the optimal experimental parameters and the photosensitization of 5-aminolevulinic acid (ALA)-based photodynamic therapy (PDT) in promyelocytic leukemia cell HL60 has been conducted, in which HL60 cells and their control groups, peripheral blood mononuclear cell (PBMC), first are incubated with different concentrations of ALA in dark for different periods of time and then followed by irradiating with different wavebands for different fluences. Fluorescence microscope and spectrofluorometer have been used to detect the fluorescence of protoporphyrin IX (PpIX) endogenously produced by ALA. The response of the cells to ALA-PDT was evaluated by the 3-(4,5-dimethyl-2 thiazolyl)-2-5-diphenyl-2H-tetrazolium bromide (MTT) assay (interval between irradiation and the MTT assay is 24 h) and by flow cytometry (the length of time between irradiation and the flow assay is 30 min). MTT results will reflect the relative number of metabolically active mitochondria in the population. Propidium iodide uptake in flow cytometry will test for membrane damage. The results of parameter experiments were obtained: 1 x 10(5)/mL HL60 cell was first incubated with 1 mmol/L ALA in dark for 4 h and the maximum fluorescence of PpIX level appeared; then irradiated with 410 nm (4 mW/cm2) for 14.4 J/cm2 and maximum photodamage to membrane and mitochondrial function of HL60 cell resulted. With the normal granulocytes, such response was not detected. Therefore a hypothetical idea can be brought forward that ALA-based PDT can be used for inactivation of leukemia cell HL60 and these optimal parameters may be useful for clinical application. PMID- 15291308 TI - Involvement of cyanobacterial phytochromes in growth under different light qualities and quantities. AB - Inactivation of the genes for the cyanobacterial phytochromes cph1 and cph2 in the unicellular cyanobacterium Synechocystis sp. strain PCC 6803 affected the growth of the cells under certain light conditions. Differences in growth were detected by recording growth curves and in competition experiments. Mutation of cph1 and cph2 resulted in different effects. The cph1-mutant strains exhibited a reduced growth rate under far-red light (FRL), whereas the growth of the cph2 mutant strains was inhibited by red light (RL). The growth rate of a cph1- / cph2 double mutant was reduced under both RL and FRL. Furthermore, cph1-, cph2- as well as double-mutant strains showed impaired growth under high-light (HL) conditions. Acclimation of the photosynthetic apparatus of the mutants to RL, FRL and HL, as determined by pigment analysis, was similar to that of the wild type. PMID- 15291309 TI - [Homeopathy does not bear up for scrutiny]. PMID- 15291310 TI - [Lightning injuries--a mixture of electrical, thermal and multiple trauma]. AB - There are several misconceptions even among hospital personnel regarding damages and injuries caused by lightning. Few health care providers have experience from lightning injuries as they are rare and different (DC) from the more common high voltage (AC) injuries. Furthermore, fatalities are uncommon. Burns do occur but are usually minor. Most lightning injuries occur in the summer season during outdoor leisure activities and in the vicinity of a tree or other large structures. In Sweden, on average, approximately seventeen persons per year are hospitalised and 0.2-0.8 persons per million inhabitants and year die due to lightning injuries. The primary treatment follows the general guidelines for other trauma, electrical, and burn injuries, i.e. as is described in the standardised ATLS, ABLS, or A-HLR programmes. However, there are some minor points that are different and may be stressed for a favourable outcome. In this paper these are addressed together with the epidemiology, effects and treatment of lightning injuries that are specific for Sweden. Unfortunately, little is known, apart from what is described in smaller case series, of the long time sequelae experienced by this patient population and further research is therefore particularly warranted in this respect. PMID- 15291311 TI - [Increasing incidence of ciprofloxacin resistant gonorrhea in Sweden. Choose a correct antibiotic and follow up the treatment!]. AB - The incidence of gonorrhoea has increased in Sweden and is now three times higher than in the middle of the 1990's. A remarkable increase of ciprofloxacin resistant gonorrhoea has been reported in Stockholm and other parts of Sweden during 2003. Among men attending a clinic for homosexual men in Stockholm the ciprofloxacin resistant cases have increased from a low level to over 50% during the last year. Most of the homosexual men are exposed in Stockholm and one serotype is dominant. Also in the county of Gavleborg there has been an outbreak of ciprofloxacin resistant gonorrhoea among young heterosexual men and women. No resistance to cefixime, ceftriaxone and spectinomycin has been noted and these antibiotics are then a better first choice of treatment in a Swedish context. PMID- 15291312 TI - [13 cases of cervical necrotizing fasciitis--all patients survived. Surgery, antibiotics and hyperbaric oxygenation give the best results]. AB - Cervical necrotizing fasciitis is a serious, rapidly progressive infection along fascia planes that sometimes involves skin, subcutaneous tissue and muscle (myositis). The condition, often of dental or pharyngeal origin, is associated with high morbidity and mortality. Thirteen consecutive cases of cervical necrotizing fasciitis treated with hyperbaric oxygen at the Karolinska Hospital during the period 1997-2003 were reviewed. Eight male and five female patients, 33 to 78 years old, were treated according to the Karolinska Hospital guidelines for severe soft tissue infections. All patients recovered. Eleven of thirteen patients required intensive care and eight inotropic drugs. Streptococcus milleri was the predominant pathogen found in initial cultures. Three case reports are presented. Our findings lend further support to the literature on the importance of a prompt multidisciplinary approach with aggressive surgical intervention, broad-spectrum antibiotic therapy and hyperbaric oxygen therapy. PMID- 15291313 TI - [Fetal programming of adult disease--heredity or environment?]. PMID- 15291314 TI - [Vaccinations--primary prevention with problems]. PMID- 15291315 TI - [The Lakartidningen 80 years ago: the medical ethics codex with impediment]. PMID- 15291316 TI - [The poisonous mercury a heritage from the god Mercurius]. PMID- 15291317 TI - [Views on health policy programs presented by the Medical Society: How do we solve the cooperation between different specialists in primary health care?]. PMID- 15291318 TI - [Individualized working hours schedule for physicians better than working three shifts]. PMID- 15291319 TI - [Anti-stigma campaigns: sympathetic but counterproductive]. PMID- 15291320 TI - [The SBU report on psychotherapy]. PMID- 15291321 TI - [The "primary health care crisis" and responsibility of hospital specialists]. PMID- 15291322 TI - [Isn't it good if one person out of 600 treated with statins will get better?]. PMID- 15291323 TI - [Should health personnel be engaged in life style diseases?]. PMID- 15291324 TI - [The framework WHO Convention on measures against smoking]. PMID- 15291325 TI - [The relationship between the population health and a nature of alcohol consumption]. AB - There is a category of individuals with a pronounced alcohol-addiction, which is known to the medical-facility system and in respect to which methods of specific and, possibly, active narcological correction must be taken. Motivations of a high death rate (in Russia) due to diseases related with blood circulation must comprise extra reasoning including an evaluation of easy access to modern cardiological medical care for broad strata of population. PMID- 15291326 TI - [Traumas and poisonings in Russia and abroad]. PMID- 15291327 TI - [Prognostication of the morbidity with temporary disability among the greenhouse female workers in Kuzbass]. AB - The paper carries data on the morbidity of greenhouse female workers in Kuzbass. Influence of the medical, social and occupational factors exerted on the occupational morbidity is evaluated; methodological approaches to prognosticating the morbidity with temporary disablement are defined. PMID- 15291328 TI - [The dynamics of the population age structure in the Far East Federal Okrug]. PMID- 15291329 TI - [Modern approaches to prognosticating the parameters of population health at the regional level]. PMID- 15291330 TI - [The social-and-hygienic aspects of andrologic morbidity among boys]. PMID- 15291331 TI - [Prevention within the context of the healthcare reform in Russia]. AB - Healthcare and promotion of preventive care rendered to population of the Russian Federation are the main components of the whole social sphere. Perfection of the legal basis for the high-quality primary prevention is a precondition of success of the Russian healthcare reform as a whole. Implementation of social and medical prevention measures must turn into a united purpose for the activities of medical personnel, enterprise management and public organizations. Prevention issues belong to the sphere of combined economic and social interests of public and healthcare. PMID- 15291332 TI - [The concept of screening medicine in oncology]. AB - Screening medicine is a basis of a suggested new concept; it comprises: an evaluation of the basic health fund within the terms of real time; continuity of the medical, ecological, epidemical, congenital and genetic information collected within the medical passport; as well as synthesis of the obtained data and monitoring of patients. The above concept enables a qualitatively new approach to coping with issues of individual health and to rendering a high-quality medical care in oncology. PMID- 15291333 TI - [On reforming the psychiatric care system in the Russian Federation]. AB - A need to amend the ideology of the psychiatric-care promotion in Russia is preconditioned by the medical-and-demographic, economic, legal, pharmacological as well as information-and-technological prerequisites. Main issues and tasks, arising in the reform before the bodies of state power, the psychiatric service and population were defined in the paper. Some priority trends of improving the psychiatric-care management in the Russian Federation were formulated within the context of a "new understanding" of the psychiatric-care protection by WHO, including staffing the system with high-skill ambulatory personnel; development of the chain of rehabilitation and of medical-and-social prevention; and a more comprehensive use of technologies substituting for inpatient facilities. PMID- 15291334 TI - [The social-and-psychological factors related to prevalence of breast feeding]. AB - A comprehensive social-and-hygienic study of families with newborns was undertaken for the purpose of defining the factors influencing the prevalence and duration of breastfeeding. A special questionnaire was worked out that was addressed to 1088 families in 3 cities, i.e. Moscow, Ulyanovsk and Cherkessk. The poor and worsening parameters of breast feeding were found to result, primarily, from the shaped practice of breast feeding present in families, when the mothers are insufficiently informed on such feeding by medical personnel, family members and family friends. An analysis of the obtained data provided for outlining the actual related ways and means on how to maintain, protect and promote the breastfeeding at the regional level. PMID- 15291335 TI - [Prevention of iodine deficit according to the results of neonatal screening for congenital hypothyroidism and medical check-ups]. PMID- 15291336 TI - [An automated system for assessing the efficiency in the treatment of patients with breast cancer]. PMID- 15291337 TI - [Emergence and development of standardization of public health in the Republic of Kazakhstan]. AB - The paper contains the results of an analysis of stages related with emergence of standardization of the healthcare system in the Republic of Kazakhstan. Attempts are described to standardize (unify) the medical services rendered to the country's population that lasted for as long as 10 years. The first medical standards of diagnosis and treatment comprised the clinical components alone, whereas the most recent standard embraces also the results of economic estimates, which show the cost of diagnostic and therapeutic measures as well as of surgical and anesthetic interventions. A lack of a unified methodological basis as well as of a normative-and-legal foundation and of analytical research is for today a problem for the promotion of standardization in Kazakhstan. At the same time, introduction of new economic relations into the healthcare system is impossible without evaluating the quality of medical care, which must be expressed both quantitatively and qualitatively and which is possible only if the actual measures and achieved results are compared with the defined norms and standards. PMID- 15291338 TI - [Outlooks for promotion of the medical-and-sanitary care rendered to the rural population in the Republic of Tajikistan]. PMID- 15291339 TI - [An outstanding Russian historian of medicine (on the occasion of 100th birthday anniversary of Professor B. D. Petrov)]. PMID- 15291340 TI - [Integration in medicine]. PMID- 15291341 TI - [Training of medical personnel in Byelorussia in the XIX-XX centuries]. PMID- 15291342 TI - [Asclepia in the antique Mediterranean]. PMID- 15291343 TI - Heterogeneity in the multiple myeloma tumor clone. AB - Multiple Myeloma (MM) is a plasma cell malignancy which is characterized by a very heterogeneous disease outcome. Heterogeneity in plasma cell characteristics, including morphology, maturation status, immunophenotype and genetic abnormalities partly account for the variable disease outcome. Although the plasma cell is the predominant cell type in MM, several studies have shown that less mature B cells, which are clonally related to the malignant plasma cells, are present in the bone marrow and peripheral blood of MM patients. The significance of these so-called myeloma clonotypic B cells in the disease process remains largely unknown. In this review the role of myeloma clonotypic B cells and myeloma tumor clone heterogeneity in relation to prognosis and clinical outcome are discussed. PMID- 15291344 TI - Adenovirus infections in stem cell transplant recipients: recent developments in understanding of pathogenesis, diagnosis and management. AB - Adenovirus is increasingly recognized as an important pathogen in stem cell transplant recipients, reflecting increased awareness about the virus, together with changes in transplant practice such as the performance of more high-risk transplants, and improvements in diagnostic methods. In retrospective studies, the reported incidence of adenovirus infections ranged between 4-20% with a similar variation in the proportion of patients developing invasive disease. In contrast, the incidence of adenovirus infection varies between 20-30% in recent prospective studies on T-cell depleted or mismatched allografts and about 30-40% of these patients develop invasive disease. These prospective studies have established a relationship between the risk of invasive adenovirus disease and a number of factors such as the extent of T-cell depletion, the intensity of immunosuppressive therapy and the kinetics of lymphocyte recovery post transplant. Polymerase chain reaction (PCR) assays to detect adenovirus DNA in peripheral blood have shown a strong correlation between viremia and the risk of disseminated adenovirus disease. These developments have led to the possibility of a preemptive antiviral treatment strategy for asymptomatic adenovirus infections. In addition, a better understanding of the interactions between adenovirus and host immune system in the post-transplant setting might enable development of effective immunotherapeutic strategies against adenovirus infections. PMID- 15291345 TI - Allelotyping in B-cell chronic lymphocytic leukemia (B-CLL). AB - B-cell chronic lymphocytic leukemia (B-CLL) is a heterogeneous malignant disease, both in terms of molecular abnormalities and clinical course. The most frequent chromosomal aberrations in B-CLL are deletions on 13q, 11q, and 17p, and trisomy 12, all of which are of prognostic significance. These aberrations can be detected by conventional cytogenetic analysis and fluorescence in situ hybridization (FISH), but cytogenetics are hampered by the low mitotic index of B CLL cells, and FISH depends on genetic information of candidate regions. Microsatellites are unique highly polymorphic and informative genetic markers dispersed in the human genome. They have become the most commonly used markers to trace loss of heterozygosity in tumors. Their detection by PCR is rapid and can be semi-automated with maximal robustness and reproducibility. In this review, we discuss the implications of a recent genome-wide analysis in B-CLL with 400 microsatellite markers. This analysis led to the detection of new aberrant loci in B-CLL which are not visible in the leukemic conventional karyotype. We conclude that microsatellite allelotyping provides a complementary comprehensive view of genetic alterations in B-CLL, and it may identify new loci with candidate genes relevant in the molecular biology of B-CLL. PMID- 15291346 TI - The potential of gene transfer into primary B-CLL cells using recombinant virus vectors. AB - Despite recent advances, chronic lymphocytic leukemia (CLL) as the most common leukemia remains a largely incurable disease. Modern treatment options include novel drugs like purine analogues, monoclonal antibodies and transplantation strategies. Moreover, gene transfer of immunostimulatory molecules is another, but still experimental approach that can be used to potentiate immune responses against leukemic cells. CD40 ligand (CD40L) was shown to be a promising molecule for immunotherapy of B-CLL playing a critical role in immune activation. However, CLL B cells are resistant to transduction with most currently available vector systems. Improving the efficiency and specificity of gene vectors is critical for the success of gene therapy in this area. Using replication defective adenovirus encoding CD40L (Ad-CD40L), immunologic and clinical responses were seen in CLL patients after infusion of autologous Ad-CD40L-CLL cells in a recent phase I trial. Due to the immunogenic nature of adenovirus vectors, alternative vector systems are currently explored. Recombinant adeno-associated virus (rAAV) was shown to enable efficient transduction of primary B-CLL cells. By use of a library of AAV clones with randomly modified capsids, receptor-targeting mutants with a tropism for CLL cells can be selected. Furthermore, helper-virus free Epstein-Barr virus (EBV)-based gene transfer vectors hold promise for development of CLL-targeted vaccines after remaining safety issues will be resolved. Herpes simplex virus (HSV)-based vectors, especially HSV amplicons, have favorable features for B-CLL gene transfer including high transduction efficiency, ability to infect postmitotic cells and a large packaging capacity. The challenge for the future will be to transfer these alternative vector systems into clinic and allow the detection of a CLL-specific immune response by use of defined tumor antigens. This will make it possible to establish the potential clinical role of gene therapy for CLL patients. PMID- 15291347 TI - Low dose total body irradiation followed by allogeneic lymphocyte infusion for refractory hematologic malignancy--an updated review. AB - Allogeneic stem cell transplantation is curative for certain cancers, but the high doses of chemotherapy and radiotherapy used in conventional myeloablative conditioning regimens may lead to severe toxicity. In our initial study, we treated 25 patients with refractory cancers with 100 cGy total body irradiation (TBI) followed by allogeneic, non mobilized peripheral blood cells. Eighteen patients received sibling and 7 patients received unrelated cord blood stem cells. None of the 13 patients with solid tumors achieved donor chimerism or had a sustained response. Twelve patients with hematologic malignancies were treated, 1 received a cord blood transplant and 11 received sibling donor cells. Nine of these 11 patients achieved donor chimerism, ranging from 5% to 100%. Four patients had sustained complete remission of their cancers, and 2 are long-term survivors. The development of chimerism correlated with total previous myelotoxic chemotherapy (p < 0.001). This technique is now being extended into the haploidentical setting. PMID- 15291348 TI - Bendamustine plus mitoxantrone--a new effective treatment for advanced chronic lymphocytic leukaemia: results of a phase I/II study. AB - The toxicity arid efficacy of the combination bendamustine plus mitoxantrone (BM) was evaluated in 22 patients with advanced chronic lymphocytic leukaemia (CLL). Twenty out of twenty-two patients had received at least one prior regimen. Median age was 71 years (range 53-86). Six out of twenty-two patients were Binet stage B and 16/22 Binet stage C. Bendamustine was given in escalating doses from 80 up to 240 mg/m2 divided in two to three doses per course, mitoxantrone was given as short infusion with 8 up to 10 mg/m2 per course. The number of courses was limited to six and chemotherapy stopped when a complete remission (CR) or partial remission (PR) according to NCI-Criteria was achieved. Haematotoxicity and infection were dose limiting with 4/6 patients suffering from grade 3 infectious complication in the bendamustine level with 240 mg/m2. Six out of twenty-two patients achieved a CR and 13/22 patients a PR resulting in an overall response rate of 86% (19/22 patients). Median time to progression was 10 months (range 4 22) and median survival 39 months (range 6-50). For further studies we recommend a bendamustine dose of 150 mg/m2 combined with 10 mg/m2 mitoxantrone. PMID- 15291349 TI - The incidence and survival of acute de novo leukaemias in Estonia and in a well defined region of Western Sweden during 1982-1996: a survey of patients aged 16 64 years. AB - In the present work the incidence and survival of acute de novo leukaemias in two neighbouring countries, were studied retrospectively over three 5-year periods, 1982-1996. The aim was to compare the above variables, particularly with respect to political/socio-economic and environmental factors, in a well defined area of Sweden, the so-called Western Swedish Health Care Region, with Estonia. Population-wise the Western Swedish Region and Estonia are very similar; area wise they are also well comparable. The present report covers only patients diagnosed between the ages of 16-64 years. The number of acute de novo leukaemias in the two regions was quite similar (Western Sweden n = 282 and Estonia n = 237). The age standardized incidence rate regarding total acute de novo leukaemias was slightly lower in Estonia than in Western Sweden (1.49/100,000 inhabitants/year for Estonia and 1.76 for Sweden, respectively), the difference being not statistically significant. However, the survival data for the two countries were highly different (P < 0.001). Thus, the relative survival for the total group of patients aged 16-64 years in Estonia at 1 year was 20.7% and at 5 years 3.6%, respectively. The corresponding figures for the Swedish patients were considerably higher, 65.2 and 29.4%, respectively. Further, the 5 year survival significantly (P < 0.05) increased for the Swedish patients over the 3 consecutive 5-year periods. No such improvement was recorded for the Estonian patients. PMID- 15291350 TI - T-cell and NK/T-cell lymphomas in southern Taiwan: a study of 72 cases in a single institute. AB - In an attempt to better understand the clinicopathologic features of T- and natural killer (NK)/T-cell lymphomas in Taiwan and the distribution and relative frequency of each subtype according to the new WHO classification, the pathology file of a medical center in southern Taiwan during 1989-2002 was retrospectively searched. The results of light microscopy, immunohistochemistry, in situ hybridization for Epstein-Barr virus (EBER), and T-cell receptor (TCR)-gamma chain gene rearrangement were correlated with clinical findings. A total of 72 cases were identified. They were peripheral T-cell lymphoma, unspecified (PTLu; n = 23, 31.9%), NK/T-cell lymphoma (n = 14, 19.4%), anaplastic large cell lymphoma (n = 13, 18.0%), angioimmunoblastic T-cell lymphoma (AITL; n = 9, 12.5%), precursor T-lymphoblastic lymphoma (n = 8, 11.1%), enteropathy-type intestinal T cell lymphoma (n = 2, 2.8%), adult T-cell leukemia/lymphoma (n = 2, 2.8%), and subcutaneous panniculitis-like T-cell lymphoma (n = 1, 1.4%). The male to female ratio was 1.5:1. Forty patients (55.6%) had extranodal presentation. Eleven cases including 9 of 14 (64.3%) NK/T-cell lymphomas expressed CD56. All 14 NK/T-cell lymphomas are EBER-positive. Seven of nine (77.8%) AITLs expressed CD10. The overall 5-year survival rate was 10.2%. In conclusion, we have characterized a large series of T- and NK/T-cell lymphomas in southern Taiwan, where there is male predominance and poor prognosis. CD56 is a specific but not very sensitive marker while EBER is most reliable for the diagnosis of NK/T-cell lymphoma. CD10 is a useful marker to differentiate AITL from PTLu. PMID- 15291351 TI - Clinical characteristics of familial vs. sporadic non-Hodgkin lymphoma in patients diagnosed at the Mayo Clinic (1986-2000). AB - There are few data on the clinical characteristics of familial vs. sporadic non Hodgkin lymphoma (NHL). Using the NHL registry at the Mayo Clinic, we compared age of diagnosis, gender, tumor site and histologic subtype between patients with sporadic and familial NHL. In 2001, we identified all new cases of adult NHL diagnosed between 1986 and 2000 in the Mayo Clinic NHL database (n = 2289) and mailed out a family history questionnaire to all living patients with a current address (n = 1043). Each NHL patient was categorized according to their self report of leukemia or lymphoma in first-degree (1 degree) relatives. We received complete FH information on 740 patients (71%). Age at diagnosis of NHL ranged from 18-88 years (mean = 59 years) and 53% of our cases were male. First-degree FH of lymphoma was reported by 43 patients (6%), 1 degree FH of leukemia by 27 patients (4%) and 1 degree FH of both in 4 (1%). There was a nonstatistically significant later age at diagnosis for cases with any family history of lymphoma or leukemia (mean age = 61.3 and 61.7 years, respectively) vs. no family history (59.0 years) (P = 0.58). The male to female ratio for those with a FH of leukemia (ratio = 2.9) was higher compared to those with FH lymphoma (0.95) or no FH (1.1) (P = 0.08). No differences were apparent between 1 degree FH and site of NHL (nodal vs. extranodal) (P = 0.53). Among recently diagnosed cases (since 1995), there was some suggestion of a greater proportion of aggressive tumors for those with any family history (69% and 55%) vs. none (50%) (P = 0.20). We found little evidence of large differences between familial and sporadic NHL with regard to age, gender, site or histologic subtype. PMID- 15291352 TI - Rituximab followed by cladribine in the treatment of heavily pretreated patients with indolent lymphoid malignancies. AB - The purpose of this study was to determine the efficacy and toxicity of combined therapy consisting of rituximab (RIT), an anti-CD20 monoclonal antibody, and cladribine (2-chlorodeoxyadenosine, 2-CdA) (RC regimen) in patients with refractory or relapsed indolent lymphoproliferative disorders. Twenty six CD20 antigen positive patients, 15 with B-cell chronic lymphocytic leukemia (B-CLL) and 11 with low grade non-Hodgin's lymphoma (LG-NHL) were enrolled to the study. Fourteen patients (53.8%) had refractory disease, the other 12 (46.2%) were recurrent after prior chemotherapy. RC regimen consisted of RIT at a dose of 375 mg/m2 in 6 h infusion on day 1 and 2-CdA at a dose of 0.12 mg/kg, in 2 h infusion, given on days 2-6. The RC courses were repeated at 4 week intervals or longer if severe myelosuppression occurred. Seventy eight cycles of RC with median of 3 cycles per patient were administered (range 1-5 cycles). Four patients (15.4%) (95% CI 1.5-29.3%), 1 with B-CLL and 3 with LG-NHL, achieved a complete response (CR). Fourteen patients (53,8%) (95%CI 34.6-72.9%), including 10 with B-CLL and 4 with LG-NHL, had a partial response (PR). Overall response rate (OR) was 69.2% (95%CI 51.4-86.9%) in the whole group, from 63.6% (95% CI 35.2 92.0%) in LG-NHL to 73.3% (95%CI 50.1-95.7%) in B-CLL patients. Twelve of 18 patients with CR/PR are still in remission, with the median follow up 10 (7-28 months). The median failure-free survival (FFS) of responders was 6.5 months. Hypersensitivity to RIT was the major toxicity of RC regimen, and occurred in 9 patients (34.6%), mostly only during the first infusion of RIT. Severe neutropenia (grade III) was seen in 3 patients (11.5%). Anemia and thrombocytopenia associated with RC treatment were observed in 5 (19.2%) and 2 patients (7.7%), respectively. Four episodes (15.4%) of grade III-IV infections were observed. There was no treatment related mortality. During the follow-up six patients (23.1%) died from the disease progression. In conclusion, the combination of RIT and 2-CdA is an effective and well tolerated treatment, even for heavily pre-treated patients, and the results seem to be better than in patients previously treated in our institution with 2-CdA alone. This regimen can be considered as an alternative treatment of CD-20 positive indolent lymphoproliferative disorders. PMID- 15291353 TI - Fludarabine, cyclophosphamide and mitoxantrone in relapsed or refractory chronic lymphocytic leukemia and low grade non-Hodgkin's lymphoma. AB - A regimen combining fludarabine, cyclophosphamide and mitoxantrone (FCM) was used to treat 29 patients with relapsed or refractory chronic lymphocytic leukemia (CLL, N = 24) and low-grade non-Hodgkin's lymphoma (NHL, N = 5) based on evidence suggesting synergism between the 3 drugs. Patients were treated with mitoxantrone 5mg/m2 i.v. day 1 only, fludarabine 25 mg/m2 i.v. for 3 days or 24 mg/m2 orally for 5 days, cyclophosphamide 250 mg/m2 i.v. for 3 days or 150 mg/m2 orally for 5 days inclusive. Eighteen patients had previously received fludarabine and most were heavily pretreated with 40% having >2 prior treatments. A median number of 4 FCM courses (range of 1-9) were given. The response rate was 78.5%: 32% complete remission, 25% nodular partial remission, 21.5%, partial remission. Median duration of response was 19 months and median survival was 42 months. Sixteen patients (57%) developed neutropenia to < 0.5 x 10(9)/l and 12 (43%) infectious complications. Four patients developed large cell lymphoma (Richter's syndrome) and 2 acute myeloid leukemia. FCM is a useful combination for relapsed or refractory CLL and low grade NHL with high response rates and long duration of response. The role of FCM as first line therapy deserves study as well as its combination with the monoclonal antibody Rituximab. PMID- 15291354 TI - Increased expression of the hPim-2 gene in human chronic lymphocytic leukemia and non-Hodgkin lymphoma. AB - Pim-1 and Pim-2 are murine proto-oncogenes implicated in lymphomagenesis. The aim of this study was to investigate whether the human Pim-2 (hPim-2) expression is altered in chronic lymphocytic leukemia (B-CLL) and non-Hodgkin's lymphomas (NHL). We analyzed hPim-2 expression in 48 patients with NHL and CLL by quantitative in-situ hybridization, quantitative RT-PCR and FACS analysis. In situ hybridization revealed a 5.5 +/- 2.2 times higher expression of hPim-2 in NHL over normal lymphocytes (P < 0.001). Similarly, with quantitative RT-PCR, expression in NHL was 1.5 to 2.6 times higher in involved splenic foci compared to nearby uninvolved regions (n = 3). hPim-2 mRNA was increased 3-folds in B-CLL over normal B-cells (P < 0.006). The increased hPim-2 levels correlated with lymphocyte doubling time (DT), in that mRNA levels were two times greater in patients with rapid DT (P < 0.006). Moreover, a significant correlation was found between hPim-2 expression and the Binet staging system of CLL (P < 0.022). The hPIM-2-protein expression was also upregulated in CLL, as assessed by FACS analysis. Therefore, this report provides direct evidence for a linkage of hPim-2 upregulation to NHL and CLL in man. This relationship between hPim-2 and NHL and CLL raises a number of novel mechanistic options for the genesis and/or progression of some types of human lymphomas. PMID- 15291356 TI - Detection of chromosome 13q deletions and IgH translocations in patients with multiple myeloma by FISH: comparison with karyotype analysis. AB - Multiple myeloma (MM) is a plasma cell dyscrasia characterized by frequent 13q deletions and IgH translocations that have clinical prognostic significance. We evaluated clonal plasma cells by interphase fluorescence in situ hybridization (FISH) and combined with immunofluorescence detection of cytoplasmic light chain (cIg-FISH) for the presence of 13q deletions and IgH translocations. The FISH results were compared with conventional cytogenetic analysis. Of the 25 bone marrow specimens from MM patients, 11 (44%) had 13q deletions. IgH translocations involving cyclin D1 (t(11;14)) and FGFR3 (t(4;14)) were found in 32 and 36%, respectively. P53 deletions were detected in 20% of the cases. One patient had coexistence of t(ll;14) and t(4;14), which has not been previously reported. Conventional cytogenetic analysis was performed in 15 cases and revealed complex numerical and structural changes in 7. Karyotype analysis failed to detect 3 of 6 cases with 13q deletions, and also missed most of the IgH translocations and p53 deletions detected by cIg-FISH. On the other hand, the complex numerical and structural changes shown by conventional cytogenetics were not demonstrated by interphase FISH. Since 13q deletions, IgH translocations and a hypodiploid karyotype are significant prognostic factors for MM, our study illustrates the importance of combining conventional cytogenetics with interphase FISH analysis in patients with MM. PMID- 15291355 TI - Polymorphisms of p53 Arg72Pro, p73 G4C14-to-A4T14 at exon 2 and p21 Ser31Arg and the risk of non-Hodgkin's lymphoma in Japanese. AB - We hypothesized that the polymorphisms in the two p53 family genes (p53 Arg72Pro and p73 G4C14-to-A4T14 at exon 2 (G4A)) and p21 Ser31Arg polymorphism might modulate the risk of non-Hodgkin's lymphoma, and conducted a hospital-based prevalent case control study at Aichi Cancer Center Hospital to clarify the association. Risk estimation for each genotype by the unconditional logistic model demonstrated the possible association between the p53 Pro72 allele and the risk of non-Hodgkin's lymphoma in Japanese population (OR = 1.59; 95% CI, 0.99 2.57, P = 0.057), although no other significant association was observed. The analyses of statistical interactions between these three polymorphisms (p73 G4A, p53 Arg72Pro and p21 Ser31Arg polymorphisms) revealed the marginally significant OR for interaction between p53 Arg72Pro and p73 G4A polymorphisms (OR = 2.54; 95% CI, 0.97 6.62, P = 0.057). When those without p53 Pro72 and p73 A4T14 alleles were defined as a reference, those with p53 Pro72 and p73 A4T14 alleles demonstrated a significantly higher OR (2.08; 95% CI, 1.11-3.90, P = 0.023). Further examination with a sufficiently larger population and other ethnicities are required to confirm our findings. PMID- 15291357 TI - Molecular disease eradication is a prerequisite for long-term remission in patients with t(8;21) positive acute myeloid leukemia: a single center study. AB - Association of long-term clinical remission and molecular disease-eradication is well established in acute myeloid leukemia (AML) patients with t(15;17) and inv(16). In patients with t(8;21) positive AML no consensus exists over the disappearance of the AML1/ETO fusion transcript during the course of disease and most studies reported reverse transcriptase polymerase chain reaction (RT-PCR) positivity as a common finding after consolidation chemotherapy, autologous and allogeneic stem cell transplantation (alloSCT). In our single center study, we performed RT-PCR monitoring in 14 patients with t(8;21) in CR1 (n = 13) and/or CR2 (n = 4). The median number of bone marrow (BM) and/or peripheral blood (PB) samples per patient was 18 (range, 2-43). In 5 out of 6 cases relapse occurred after persistence of minimal residual disease (MRD) in CR for 4-14 months. The sixth patient relapsed despite molecular remission (MR) in BM and PB for 3 months, molecular relapse preceded hematological relapse for 7 months. Eleven patients with a median follow-up of 7.8 (range, 1.5-15.4) years are in persistent CR and MR after consolidation chemotherapy (n = 7). mainly with repetitive cycles of high-dose Ara-C, autologous (n = 1) or myeloablative allogeneic (n = 3) stem cell transplantation. Molecular remission was attained immediately after alloSCT, but after 6-26 months in CR in patients with consolidation chemotherapy. In 7 patients, MRD was only studied in long-term remission. In conclusion, long-term CR was associated with persistent molecular disease-eradication. In our patients, molecular remission was a prerequisite but not a guarantee for long-term disease free survival. Hematological relapse never occurred without prior molecular relapse. Due to the slow kinetics of AML1/ETO after consolidation chemotherapy the value of qualitative RT-PCR to predict early relapse is limited. In this situation quantitative RT-PCR might help to define individual relapse risk and to improve as well as facilitate clinical decision-making. PMID- 15291359 TI - Phenylarsine oxide (PAO) more intensely induces apoptosis in acute promyelocytic leukemia and As2O3-resistant APL cell lines than As2O3 by activating the mitochondrial pathway. AB - We studied the cytotoxic effect of an organic arsenical compound, phenylarsine oxide (PAO) on an acute promyelocytic leukemia (APL) cell line (NB4) and an As2O3 resistant NB4 subline (NB4/As). Cell growth was inhibited by 50% (IC50) upon 2 day treatment with As2O3 or PAO at 0.54 and 0.06 microM, respectively in NB4 cells (P = 0.025), and 2.80 and 0.08 microM, respectively in NB4/As (P = 0.030). 0.1 microM PAO increased the proportion of hypodiploid cells (50.3%) by a greater degree than the same dose of As2O3 (3.8%) in NB4 cells. In NB4 cells, 0.1 microM PAO reduced the mitochondrial transmembrane potential (20.5% in a PI(negative) Rhodamine123(low) fraction) by a greater degree than 1 microM As2O3 (7.1%). Western blotting showed that 0.1 microM PAO downregulated the expression of both Bcl-2 and Bcl-X(L) proteins, whereas I microM As2O3 downregulated only Bcl-2 expression. These results suggest that the cytotoxic effect of PAO on an APL cell line and As2O3-resistant subline is significantly higher than that of As2O3. PAO induced apoptosis seems to be related to the activation of the mitochondrial pathway and downregulation of both Bcl-2 and Bcl-X(L). PAO is a considerable agent for relapsed/refractory APL and for purging APL cells following stem cell transplantation. PMID- 15291358 TI - Nuclear retinoid receptors are involved in N-(4-hydroxyphenyl) retinamide (Fenretinide)-induced gene expression and growth inhibition in HL-60 acute myeloid leukemia cells. AB - N-(4-hydroxyphenyl) retinamide (Fenretinide, 4-HPR) inhibits cell growth by inducing apoptosis in numerous tumor cell types including all-trans-retinoic acid (ATRA)-resistant tumor cells. However, the mechanism(s) by which 4-HPR mediates its anti-proliferative effects remains unclear. Here, we determined whether 4-HPR induced growth inhibition and gene expression involve retinoid receptors in human acute myeloid leukemia (AML) cells (HL-60). We treated HL-60 and ATRA-resistant HL-60 (HL-60R) cells that express mutated RARalpha and very low levels of RARbeta, RARgamma and RXRalpha with 4-HPR (2 microM) for 3 days. 4-HPR showed significant anti-proliferative effects against both cell types and induced growth inhibition (92.7%) in HL-60 cells. However, at the same dose, 4-HPR induced only 53.4% growth inhibition in HL-60R cells. Growth inhibition by 4-HPR was significantly enhanced in HL-60R cells that were retroviraly transduced to express human RARalpha, RARbeta or RXRalpha (95.6%, 97.1%, and 75.6%, respectively), in comparison to HL-60R cells (P < 0.05), but not in HL-60R cells expressing RARgamma. Although ATRA and 4-HPR induced expression of CYP26, an ATRA inducible gene encoding a cytochrome P450 enzyme, in HL-60 cells, both retinoids failed to induce CYP26 in HL-60R cells. However, induction of CYP26 mRNA by 4-HPR was restored in HL-60R cells expressing RARalpha and RARgamma, but not RARbeta and RXRalpha. In conclusion, our data suggest that nuclear retinoid receptors are involved in 4-HPR-induced growth inhibition and gene expression, and that 4-HPR can mediate its anti-proliferative effects through retinoid receptor-dependent mechanisms in HL-60 cells. PMID- 15291360 TI - Bryostatin induces protein kinase C modulation, Mcl-1 up-regulation and phosphorylation of Bcl-2 resulting in cellular differentiation and resistance to drug-induced apoptosis in B-cell chronic lymphocytic leukemia cells. AB - Bryostatin, a macrocyclic lactone and protein kinase C (PKC) modulator, has been shown to have differentiation and anti-tumor activity against several leukemia cell lines in vitro. In this study, we demonstrated Bryostatin-induced differentiation in B-cell chronic lymphocytic leukemia (B-CLL) cells, characterized by an increase in cell size and a marked up-regulation of CD11c expression. The specific inhibitors of the extracellular signal-regulated kinase (ERK) and protein kinase C pathways, PD98059 and GF 109203X respectively, each completely blocked Bryostatin-induced differentiation of B-CLL cells, suggesting that activation of the ERK pathway plays a direct role in this process in a PKC dependent manner. Furthermore, Bryostatin reduced both spontaneous and drug induced apoptosis with chlorambucil, fludarabine and 2-chloro-2'-deoxyadenosine (2-Cda) in B-CLL cells. This resistance was associated with an early up regulation of the anti-apoptotic protein Mcl-1 and post-translational phosphorylation of Bcl-2 at serine 70. The anti-apoptotic effects of Bryostatin were abrogated by GF 109203X, and to a lesser extent by the phosphatidylinositol 3-kinase (PI3-kinase) inhibitor, LY294002. Interestingly, the ERK inhibitor, PD98059 inhibited Mcl-1 expression but had little effect on Bryostatin-induced survival suggesting that the ERK pathway predominantly affects differentiation. Taken together these results present an explanation for Bryostatin-induced B-CLL cell survival in which modulation of the PKC pathway couples differentiation with an increase in antiapoptotic protein expression and calls into question the rationale for its use in the treatment of B-CLL. PMID- 15291361 TI - Signal transduction in anaplastic large cell lymphoma cells (ALCL) mediated by the tumor necrosis factor receptor CD30. AB - Expression of the cytokine receptor CD30 is a characteristic feature of anaplastic large cell lymphoma (ALCL). Reports regarding CD30-mediated signaling in ALCL cells are highly controversial, especially with respect to the regulation of cell survival. In this study, we stimulated 6 ALCL-derived cell lines with immobilized anti-CD30 antibody. CD30-induced cell death was investigated by Western blot and FACS analysis. CD30-dependent cell proliferation and activation was analyzed by applying the trypan blue exclusion method and a luciferase-based ATP assay. The expression of cell cycle relevant proteins and the activation of mitogen-activated protein (MAP) kinases were also examined. We demonstrated that activation of CD30 did not lead to the cleavage of pro-caspase-3. FACS analysis confirmed that in all examined cells cell death was not mediated by CD30. Cell growth was strongly inhibited in 2 of the 6 cell lines and restrained cell growth was accompanied by expression of the cell cycle inhibitor p21(WAF1/CIP1). Furthermore, stimulation of CD30 led to the activation of the p38 MAP kinase but not of the extracellular signal-regulated kinase (ERK) or the jun N-terminal kinase (JNK). Interestingly, activation of CD30 induced a strong synergistic reduction of cell activity, if the p38 MAP kinase activity was blocked by SB203580. The aim of the study was to elucidate CD30-induced signaling in different ALCL-cells. Our results suggest that CD30-mediated apoptosis is not a common feature in this cell type and that p38 MAP kinase is involved in CD30 mediated singal transduction. PMID- 15291362 TI - Granulocytic differentiation of leukemic cells with t(9;11)(p22;q23) induced by all-trans-retinoic acid. AB - Acute leukemia patients with MLL (mixed linage leukemia) rearrangements tend to respond poorly to conventional therapies. We examined differentiation of human myeloid leukemia cells displaying the MLL-AF9 gene, using several differentiation agents. When MOLM-14 cells were treated with all-trans retinoic acid (ATRA) or 1beta,25-dihydroxyvitamin D3, significant induced differentiation was observed. Trichostatin A (TSA), an inhibitor of histone deacetylase, demonstrated enhance effects with ATRA in regard to growth inhibition and differentiation induction in MOLM-14 cells. Pretreatment with TSA before exposure to ATRA displayed increased effect. Based on these findings, combined treatment with ATRA and TSA may be clinically useful in therapy for acute leukemia displaying MLL-AF9 fusion gene. PMID- 15291363 TI - Retinoic acid receptor antagonist inhibits CD38 antigen expression on human hematopoietic cells in vitro. AB - The CD34+ CD38- subset of human hematopoietic stem cells are crucial for long term ex-vivo expansion; conditions that decreased this specific sub-population reduced the self-renewal capacity and shortened the duration of the proliferative phase of the culture. Retinoids, such as all-trans retinoic acid (ATRA), have been shown to induce CD38 expression. ATRA present in serum may be responsible for the high CD38 of cells grown in serum-containing medium. In the present study we analyzed the effects of AGN 194310, a retinoic acid receptor pan-antagonist, on CD38 expression of human hematopoietic cells. Normal cells (cord blood derived CD34+ cells) and abnormal cells (myeloid leukemic lines) were studied when grown in either serum-containing or serum-free media. The results showed that both serum and ATRA enhanced differentiation and, thereby, reduced the proportion of CD34+ CD38- cells and total CD34+ cell expansion. AGN reversed these effects of serum and ATRA: it delayed differentiation and increased CD34+ CD38- cells. These results suggest that physiological ATRA levels in serum may prevent efficient cell expansion. AGN, by neutralizing ATRA, improves cell expansion in serum containing cultures, thus making AGN a useful agent for ex vivo expansion of stem cells and other specific sub-populations for research and clinical use. PMID- 15291364 TI - Sinus histiocytosis with massive lymphadenopathy (Rosai-Dorfman's disease) previously misdiagnosed as Toxoplasma Lymphadenitis. AB - Here we describe a case of Rosai-Dorfman Disease (RDD) in a 25-year-old female patient from Turkey who was previously misdiagnosed with Toxoplasma Lymphadenitis, and review the manifestations and treatment of this rare entity. To the best of our knowledge this is the third description of RDD [Sinus Histiocytosis with Massive Lymphadenopathy (SHML)], involving bilateral cervical lymphadenopathy and nephromegaly previously misdiagnosed as Toxoplasma Lymphadenitis. Representative clinical, radiographic and histological findings are presented. Its etiology, diagnosis and management are also reviewed. Sinus Histiocytosis with Massive Lymphadenopathy is a rare disorder of unknown etiology, usually associated with lymph node enlargement in various superficial or deep sites. The key histologic feature of SHML is the presence of various numbers of large, pale histiocytic cells that contain within their cellular borders apparently engulfed lymphocytes (emperipolesis); these distinctive large, pale cells are S-100 protein positive CD-68 positive and CD1a negative by immunostaining. According to the literature the most effective treatment found was surgical debulking. PMID- 15291365 TI - Epstein-Barr virus positive large B-cell lymphoma arising in a patient previously treated with Cladribine for hairy cell leukemia. AB - We describe the case of a patient treated with 2-chloro-2'-deoxyadenosine, CdA or Cladribine for hairy cell leukemia who subsequently developed an Epstein Barr virus (EBV)-positive polymorphous large B-cell lymphoma (p-LBCL). The time interval between Cladribine therapy and development of p-BCL was 11 months and morphologically resembled an EBV-positive post transplant lymphoproliferative disorder (PTLD). Molecular genetic studies for EBV-clonality by Southern blot hybridization showed a clonal population of infected cells, implying that this was an EBV induced lesion. The chronology of events suggest that Cladribine, a purine analog which has been previously described to induce long-lasting immunodeficiency, can, in some cases, weaken the host defense mechanism to a level at which an innocuous EBV infection may transform the normal lymphoid cells into an aggressive neoplasm. Unlike most methotrexate-related lymphoproliferative disorders (LPDs), which undergo spontaneous remission after discontinuation of therapy, LPDs secondary to purine analogs often fails to resolve after discontinuation of therapy and requires additional therapy. Our patient was treated with rituximab following the diagnosis of p-LBCL, with the goal of improving the pancytopenia to permit chemotherapy. However, the patient failed to show any dramatic improvements in counts, developed systemic symptoms and progressive ascites. He expired 3 weeks after a second dose of rituximab. Cladribine is a potent immunosuppressive agent and should be included with the list of immunosuppressive agents that may be associated with EBV-related B-cell lymphoproliferative disorders. PMID- 15291366 TI - Primary diffuse large cell lymphoma of the mandible. AB - Primary diffuse large cell lymphoma of the mandible is a rare form of extranodal non-Hodgkin's lymphoma (NHL). Herein we present 4 cases treated at our institution over a 5-year period and review 40 cases previously reported in the English-literature. The median age at presentation is 51 years with equal distribution between males and females. At presentation the lymphoma is usually limited to the jaw (stage IE) and the most common presenting symptoms include swelling of the jaw (58%), pain (53%), and mental dysesthesia or numbness (20%). Despite symptoms of numb chin syndrome, central nervous system (CNS) involvement at presentation has not been reported. The reported therapy of this rare diffuse large cell lymphoma presentation is very heterogeneous, however majority of patients were treated with combination of chemotherapy and radiotherapy with estimated 5-year overall survival of only 60%. Multi-center prospective clinical trials are needed to determine the optimal therapeutic approach to this rare diffuse large cell lymphoma presentation. PMID- 15291367 TI - Burkitt's lymphoma of the stomach: a case report with molecular cytogenetic analysis. AB - In 1974, a 28-year-old man presented with a 12 cm sized ulcerated tumor involving the middle third of the stomach, which was originally diagnosed as "lymphosarcoma". Clinical recurrence of the lymphoma resulted in rapidly progressing disease and the patient died 4 months after initial diagnosis. Retrospective work-up of the 29-year-old tumor blocks revealed the typical histologic appearance and phenotype (CD20, CD10, BCL-6 positive) of Burkitt's lymphoma (BL) with a proliferation rate of 95%. By fluorescence in situ hybridization (FISH). the tumor cells were shown to harbor an IGH-MYC fusion indicating the presence of the hallmark Burkitt-translocation t(8;14)(q24;q32). Considering the typical clinical features of BL requiring appropriate treatment regimes the case presented here highlights the importance of modern histopathologic and molecular cytogenetic techniques for the proper classification of such rare lymphomas presenting at atypical sites. PMID- 15291368 TI - Hemophagocytic, non-secretory multiple myeloma. AB - A 70-year-old woman presented with pancytopenia associated with plasma cell infiltration of her bone marrow. The plasma cells were often multinucleated and demonstrated phagocytosis of erythroid and granulocytic cells. Atypical immunophenotypic features included the expression of CD117 and CD13 and the lack of expression of CD56. Although kappa chains were demonstrable in the cytoplasm, no paraprotein was found in the serum or urine. Osteolytic lesions were absent. The pancytopenia of this unusual patient with non-secretory, hemophagocytic myeloma has improved on dexamethasone monotherapy, although her hemophagocytosis persists. PMID- 15291369 TI - Repeated clonal relapses in classical Hodgkin's lymphoma and the occurrence of a clonally unrelated diffuse large B cell non-Hodgkin lymphoma in the same patient. AB - Classical Hodgkin's lymphoma (cHL) and B-cell Non-Hodgkin Lymphoma (B-NHL) occasionally occur in the same patient. However, molecular studies that aim at determining the clonal relationship between both lymphomas are rare. In most instances, the lymphoma components appear to be derived from the same germinal center (GC) B cell precursor. We describe the molecular monitoring of repeated clonal relapses in a patient with cHL in whom diffuse large B cell NHL (DLBCL) developed concurrently. Intriguingly, both lymphomas were clonally unrelated. Thus, the derivation from a shared precursor likely is not a general rule in cHL and DLBCL arising in the same patient. PMID- 15291370 TI - An unusual case of composite lymphoma involving chronic lymphocytic leukemia follicular lymphoma and Hodgkin disease. AB - Composite lymphomas constitute the presence of two different types of non-Hodgkin lymphoma or Hodgkin and non-Hodgkin lymphoma at the same anatomic site. We report an unusual case of a 73-year-old woman who initially presented with a composite lymphoma of chronic lymphocytic leukemia (CLL) and follicular lymphoma. After 5 years of follow-up and intermittent treatment, she developed Hodgkin disease with diffuse liver involvement. Biopsy of the liver showed Reed-Sternberg cells with typical morphology and immunophenotype. While fluorescent in situ hybridization (FISH) analyses for t(14;18) were positive in the lymph node tissue with follicular lymphoma, we were unable to show the same in the liver biopsy specimen. Here, we describe the clinical, morphologic, immunophenotypic, and cytogenetic features of this unusual composite lymphoma case involving CLL and follicular lymphoma, with the subsequent development of a Hodgkin lymphoma. PMID- 15291371 TI - The incidence rate and clinical aspects of HGV infection in children with positive history of neoplasm. PMID- 15291372 TI - Brain abscess during induction in acute lymphoblastic leukemia. A case report. PMID- 15291373 TI - Changing prognostic role of preserved dendritic cell reticulum in Hodgkin's lymphoma. PMID- 15291374 TI - How important is optimal blood pressure control? AB - BACKGROUND: Evidence for the cardiovascular benefits of antihypertensive treatment is among the strongest in medicine. Randomized, prospective, unconfounded studies in thousands of people have shown that even small reductions in blood pressure for short periods substantially improve cardiovascular outcomes. Recent evidence has emphasized the importance of optimal blood pressure control, particularly in patients with high cardiovascular risk, such as those with type 2 diabetes mellitus. OBJECTIVE: This article discusses optimal target blood pressure goals, reviews the effects of antihypertensive treatment in high risk patients, presents current guidelines for blood pressure control, discusses the failure worldwide to achieve such control, and suggests approaches to improved treatment. CONCLUSIONS: In high-risk patients, small improvements in blood pressure control are associated with large reductions in cardiovascular risk. National and international guidelines for the management of hypertension therefore now recommend rigorous blood pressure targets. Despite extensive clinical evidence, the delivery of care for hypertension remains unsatisfactory. Hypertension is underdiagnosed and undertreated, and recommended target blood pressures are rarely achieved. Physicians appear reluctant to make changes to treatment, which would lead to more effective use of antihypertensive drugs. Overwhelming evidence supports the benefit of optimal blood pressure control in patients with hypertension. Several studies have shown that such control can be achieved, but most likely requires combination treatment. Combination treatment is likely to be successful only if drugs are well tolerated and patients are compliant. The angiotensin II receptor antagonists (sartans) have the therapeutic advantage of efficacy, excellent tolerability, and a good record of compliance. Blood pressure control can be more easily accomplished by using sartans early in treatment and by recognizing the benefits of even small reductions in blood pressure. PMID- 15291375 TI - Antagonizing the angiotensin II subtype 1 receptor: a focus on olmesartan medoxomil. AB - BACKGROUND: The orally active, nonpeptide antagonists of the angiotensin II subtype 1 (AT1) receptor represent a recent class of antihypertensive drugs that selectively block the renin-angiotensin system. Olmesartan medoxomil is the newest member of this class. OBJECTIVE: This article reviews the renin angiotensin system and how this system can be pharmacologically inhibited by the selective antagonists of the AT1 receptor, with a main focus on the AT1 receptor antagonist olmesartan. METHODS: Key studies were selected from previous work to illustrate the antihypertensive, cardioprotective, and renoprotective effects of olmesartan, and to compare class effects of AT1 receptor antagonists and angiotensin-converting enzyme (ACE) inhibitors. RESULTS: Olmesartan, the active metabolite of olmesartan medoxomil, is a highly potent antagonist of the AT1 receptor. It inhibits the contractile responses to angiotensin II in guinea pig aorta, inhibits the pressor responses to angiotensin II in rats and dogs, and exhibits dose-dependent antihypertensive effects in spontaneously hypertensive rats. In addition to its antihypertensive effects, olmesartan medoxomil provides protection against cardiac and renal damage in animal models. AT1 receptor antagonists are more specific inhibitors of the renin-angiotensin system compared with ACE inhibitors. They are well tolerated and have an excellent safety profile. Unlike angiotensin-converting enzyme inhibitors, AT1 receptor antagonists lack the nonangiotensin-related side effects such as cough and angioedema. CONCLUSIONS: AT1 receptor antagonists such as olmesartan represent a valid therapeutic option for the treatment of hypertension and other cardiovascular and renal diseases. PMID- 15291376 TI - Can the pharmacokinetic characteristics of olmesartan medoxomil contribute to the improvement of blood pressure control? AB - BACKGROUND: Poor compliance with treatment and early discontinuation negatively affect long-term blood pressure control. According to the Seventh Report of the Joint National Committee on Prevention, Detection, Evaluation, and Treatment of High Blood Pressure, poor compliance could contribute to the lack of adequate blood pressure control in approximately 70% of patients with hypertension, even after treatment with several antihypertensive drugs. OBJECTIVE: This article discusses blood pressure control and patient compliance with antihypertensive drugs. It then outlines the clinical pharmacokinetics and interaction profile of the novel antihypertensive olmesartan medoxomil, an angiotensin II receptor antagonist (AIIRA), and relates the findings to the recommended dosing regimen and tolerability of the drug. CONCLUSIONS: Many factors influence patient compliance, including forgetfulness, adverse effects, irregular lifestyle, and complexity of drug regimen, such as multiple drugs and, in particular, frequent dosing. Drugs that are well tolerated and easy to manage in a once-daily regimen improve compliance. In a study of different classes of antihypertensive drugs, patients taking AIIRAs were more likely to continue their treatment than those taking any other class of antihypertensive. Therefore, the right drug choice may contribute to patient compliance. The newest member of the AIIRAs, olmesartan medoxomil, is a long-acting antihypertensive drug with a favorable safety and tolerability profile. It is not metabolized by and does not interfere with the cytochrome P-450 enzyme system. These properties meet modern requirements for the treatment of hypertension. PMID- 15291377 TI - Clinical efficacy and tolerability of olmesartan. AB - BACKGROUND: Olmesartan medoxomil is an angiotensin II receptor antagonist that selectively and competitively inhibits the angiotensin II type 1 receptor. OBJECTIVE: This article reviews the results of some key studies that assessed the efficacy and tolerability of olmesartan in patients with hypertension. METHODS: Olmesartan has been investigated in several clinical studies. This article reports on data from 1 such study with a prospective, randomized, double-blind, placebo-controlled, parallel-group, dose-finding design in patients with mild to moderate hypertension (baseline mean sitting diastolic blood pressure, 100-114 mm Hg). The results from a meta-analysis of 7 randomized, double-blind, placebo controlled studies are also presented. RESULTS: In the dose-finding study, 792 patients were randomized to olmesartan (2.5-80 mg) or placebo once daily, and changes were recorded in trough mean sitting diastolic and systolic blood pressures from baseline to the end of a 12-week treatment period. For the meta analysis, 3055 patients were randomized to treatment; 2511 received olmesartan. In the dose-finding study as well as in the meta-analysis, olmesartan (2.5-80 mg once daily) produced a dose-dependent decrease in diastolic and systolic blood pressures, and at a dose of 10 to 80 mg showed significant superiority in reducing diastolic blood pressure over placebo (P < 0.05). The 20-mg dose was considered optimal, with a responder rate of 70%. Furthermore, in a 2-year study with 462 patients, olmesartan had a good safety profile and was well tolerated. The results of the clinical studies in >3000 patients receiving olmesartan showed that the frequency and profile of adverse events with olmesartan were generally similar to those with placebo; the frequency of adverse events was not dose related. Olmesartan, with or without hydrochlorothiazide, was well tolerated over 2 years of treatment. PMID- 15291378 TI - Olmesartan compared with other angiotensin II receptor antagonists: head-to-head trials. AB - BACKGROUND: Drugs that block the renin-angiotensin system represent one of the most significant therapeutic interventions available for the treatment of hypertension. The angiotensin II receptor antagonists (AIIRAs), also known as sartans, are one such class of drugs that block the effects of angiotensin II by antagonizing the angiotensin II type 1 receptor. Olmesartan is the newest member of this class. OBJECTIVE: The present article reviews 3 head-to-head trials directly comparing the antihypertensive efficacy of olmesartan with that of 4 other AIIRAs, at recommended maintenance doses, already in clinical use for the treatment of hypertension. RESULTS: In the first study, olmesartan 10 mg/d was compared with losartan 50 mg/d in 316 patients with mild to moderate hypertension (mean baseline diastolic blood pressure [DBP], 95-114 mm Hg). Dosage was doubled at week 4 and hydrochlorothiazide was added at week 12 if blood pressure response was inadequate. Olmesartan was significantly more effective than losartan with respect to the reduction in blood pressure at weeks 2, 4, and 12, and to the responder rate at weeks 2 and 4 (with the starting dose of the respective drug). In a second study, olmesartan 20 mg/d was shown to be significantly more effective than losartan 50 mg/d, valsartan 80 mg/d, and irbesartan 150 mg/d in 588 patients with mild to moderate hypertension (mean sitting baseline DBP, 100 115 mm Hg) (P < or = 0.05). At week 2, the reduction in blood pressure observed with olmesartan was significantly greater than that of the comparator treatments (P < or = 0.05). The superiority of olmesartan was maintained at week 8. The third study involved 643 evaluable patients with moderate to severe hypertension (mean DBP, 100-120 mm Hg; mean systolic blood pressure, >150 mm Hg). Olmesartan 20 mg/d was more effective than candesartan 8 mg/d in lowering 24-hour blood pressure at week 8 (P < or = 0.05). Most of this treatment effect was evident after only 1 or 2 weeks, with greater reductions in blood pressure compared with candesartan. CONCLUSIONS: These data indicate that, at the doses studied, olmesartan is more effective than other AIIRAs tested at their recommended doses, in terms of reduction of cuff or 24-hour ambulatory blood pressure, in patients with essential hypertension. These differences in blood pressure reduction between these agents may be clinically relevant and have important long-term implications. Additional studies will further define the role of olmesartan in the management of cardiovascular diseases, such as atherosclerosis. PMID- 15291379 TI - [We need to think before proposing a new test for detecting high risk HPV]. PMID- 15291380 TI - [What could epidemiology do against terrorism?]. PMID- 15291381 TI - [Scientific research in Italy: as before, worse than before]. PMID- 15291382 TI - [Lucio Luzzato's dismissal from Genoa Cancer Institute: an ignoble farce]. PMID- 15291383 TI - [New rules for the chemical industry in Europe]. PMID- 15291384 TI - [Environmental health surveillance system in urban area near incinerators: European INTERREG III C Project "Enhance health"]. PMID- 15291385 TI - [PCB contamination in Brescia: the results of the research performed by the local health unit]. PMID- 15291386 TI - [Lombard citizens, school medicine and EBP]. PMID- 15291387 TI - [A limit case]. PMID- 15291388 TI - [An unusual excess of mortality in a small Tuscan municipality and the "nursing home effect"]. AB - OBJECTIVE: In the last decades unusual mortality excesses were observed in the small area of Montaione, where the main activities are agriculture and tourism. The aim of this study was to evaluate if the observed excess mortality had to be attributed to the deaths occurred among the local large Nursing Home's guests which were half of the total deaths registered among residents. DESIGN: Empirical Bayesian Mortality Ratios (EBMR), applying the method of Clayton and Kaldor, were calculated either including or excluding the guests of the Nursing Home from the deaths and from the population. Only the population with age > or = 65 was included in the analysis. The expected deaths were calculated using the Tuscan population mortality rates by sex, age and specific cause of death (all causes, cardiovascular diseases, cerebrovascular diseases, digestive diseases and respiratory diseases). RESULTS: Excluding the guests of the Nursing Home from the analysis it was observed a strong decrease of the EBMRs for almost all causes considered, but those for cerebrovascular and respiratory diseases. CONCLUSION: The results obtained underline the necessity to take in consideration also a possible "Nursing Home effect" in evaluating mortality excesses in small areas. PMID- 15291389 TI - [Surveillance of congenital malformations in Italy: an investigation in the province of Siracusa]. AB - OBJECTIVE: The study describes briefly the current situation of the surveillance of congenital anomalies in Italy and gives an insight into the province of Siracusa in order to better characterise health status of populations residing in an area at high environmental risk. PARTICIPANTS: The authors, who coordinate the Italian registries of congenital malformations, have collaborated with the Eastern Sicily Registry of congenital malformations (ISMAC) and the registry of diseases of the Siracusa province. DESIGN AND SETTING: Data collected by the ISMAC Registry were used to calculate the prevalence of malformed newborns, resident in the municipalities of the province of Siracusa between 1991-2000. This prevalence was compared to that observed in the rest of the Siracusa province (RSP), in the whole area covered by the ISMAC Registry (ESR) and to the mean prevalence at birth of the North-East, Emilia Romagna, Toscana and Campania Registries (IR). Comparisons were made for all malformations and for groups of malformations (with the exclusion of groups with a 10 year frequence <10 cases in the province of Siracusa). In addition, heterogeneity among the municipalities of the province and temporal trends were statistically tested. MAIN OUTCOME MEASURES: Total, groups and specific congenital malformations. RESULTS: Results were statistically borderline considering all malformations when the Priolo Augusta-Melilli area was compared to IR and ESR (standardized morbidity ratio SMR(IR)=1.1, SMR(ESR)=1.2) and statistically significant when compared to RSP (prevalences ratio PR(RSP)=1.9). Significant excesses resulted in this area also for hypospadias (SMR(IR)=1.9, SMR(ESR)=2.4, PR(RSP)=2.5) and anomalies of the digestive system (SMR(IR)=2.1, SMR(ESR)=1.9, PR(RSP)=2.6). CONCLUSION: Following these results a case-control study on malformations observed in excess has been activated and a protocol for the surveillance of sensitive diseases in areas at environmental risk is being elaborated. PMID- 15291390 TI - [Environmental tobacco smoke exposure in public places in Florence, Italy]. AB - OBJECTIVES: Measurements of the environmental tobacco smoke (ETS) exposure in public places in Florence. This study was part of the first European multicenter project, intended to measure ETS exposure in public places in a number of European Cities (Florence, Barcelona, Paris, Oporto, Athens, Wien and Orebro). DESIGN: Nicotine vapour phase was measured using passive samplers, composed of a sodium bisulphate treated filter held in a plastic cassette with a windscreen on one side. The filters were analysed at the Laboratory of the Public Health Agency of Barcelona, Spain, by gas-chromatography/mass spectrometry (GC/MS). MAIN OUTCOME MEASURES: Nicotine concentration (in microg/m3) by public place, by smoking policy, and, for restaurants with separated areas, by smoking and non smoking section. SETTING: Nicotine measurements were conducted in 5 schools, 3 university departments, 5 hospitals, 1 railway station, 1 airport, 7 bars, 7 restaurants, and 4 discotheques in Florence. RESULTS: The average nicotine concentration in discotheques and restaurants were respectively 26.78 microg/m3 and 2.32 microg/m3. In the other public places the concentration was about 1 microg/m3. In smoke-free public places the average concentration was 0.85 microg/m3; in public places where smoking is allowed concentration was higher (11.53 microg/m3). In the smoking section and non-smoking section of restaurants with separated areas the average concentration was respectively 2.54 and 2.14 microg/m3. CONCLUSION: The highest nicotine concentrations were recorded in discos and restaurants. A smoke-free public place is effective in reducing ETS exposure. Smoking and non-smoking sections in restaurants without a separate ventilation system seem not to solve ETS exposure. PMID- 15291391 TI - [Temporal trends in AIDS incidence and mortality in Tuscany (1987-2000)]. AB - OBJECTIVE: To describe temporal trends in AIDS incidence and mortality in an Italian region. DESIGN: Descriptive study based on incidence and mortality registries. SETTING: Tuscany. MAIN OUTCOME MEASURES: AIDS incidence and mortality annual truncated rates, age-standardized; joinpoint analysis highlighted significant changes in the temporal trends. RESULTS: AIDS incidence rose until 1995 and then decreased by 30% every year; AIDS mortality rose in men until 1995 and in women until 1996, then decreased by 35.9% and 49.7% every year respectively. Differences in AIDS mortality have been found between the residents of the coastline municipalities and those of other municipalities. CONCLUSION: Differences in the temporal trends may mirror differences in risk group composition and in health-care access. PMID- 15291392 TI - [Survival analysis of malignant mesothelioma treated in Brescia, northern Italy, 1982-2000]. AB - OBJECTIVE: To study the survival for malignant mesothelioma on general population cases over the 1982-2000 period and to evaluate the effectiveness of the new therapeutic protocols (intrapleural immunotherapy and mulitmodality therapy) to improve the prognosis on the group of selected hospital patients treated from 1996 to 2000. DESIGN: Survival analysis of malignant mesothelioma on the general population cases and survival analysis for the 1996-2000 period on the selected group of hospital patients. SETTING: The Malignant Mesothelioma Register of the Brescia Province (northern Italy) and Pulmonology Dept. of the General hospital of Brescia. PARTICIPANTS: 353 mesothelioma cases observed in the province of Brescia from 1982 to 2000, 215 of which are residents in the province and 138 from other provinces, 324 are pleural and 29 peritoneal mesothelioma, 141 of all diagnosed between 1982 and 1995 and 212 between 1996 and 2000. MAIN OUTCOME MEASURES: Observed survival (%) at one, three, five-year and median survival by gender, site and residence on the general population cases treated with conventional therapy and on selected hospital patients group treated with intrapleural immunotherapy and mulimodality therapy. RESULTS: Median survival for pleural mesothelioma is of 233 days for the men and 291 days for the women in the group of incident cases; median survival is higher for cases from other provinces (388 and 496 days respectively). From 1996 the number of cases treated with new therapic protocols is steadily increasing, patients without therapy (or only talcaggio) passed from 87% in the period between 1982-1995 to 43% in the period 1996-2000. Nevertheless, multivariate analysis by the Cox model based on incident cases proved that histological subtype and age are the only most important prognostic factors (cases with fibrous morphology and older age are associated with lower survival). CONCLUSION: In the incident case group the increase of survival for pleural mesothelioma of cases treated with recent therapeutic protocols doesn't reach statistical significance. These results are similar to those of previous studies conducted to identify prognostic factors for mesothelioma survival; they don't permit confirmation of the efficacy of the recent therapy on the population base cases and only allow one to suppose them in the cases of the non resident population. The efficacy of the recent therapy has to be further investigated with regard to the stage of the disease. PMID- 15291393 TI - [Statistical approaches to test for spatial heterogeneity of relative risks of cause-specific mortality in Alto Vicentino Municipalities (years 1991-2000)]. AB - OBJECTIVE: To review the statistical approaches to test for spatial heterogeneity of relative risks. Three different statistical tests (Gail, Martuzzi-Hills and Potthoff-Whittinghill) are reviewed and applied--as motivating example--to the analysis of cause-specific mortality records (years: 1991-2000) of the Municipalities belonging to the Local Health Unit Alto Vicentino. METHODS: Spatial heterogeneity was found in 17 (Martuzzi-Hills) and 18 (Gail, Potthoff Whittinghill) among 70 selected causes of death. Cohens Kappa test was chosen to assess the agreement among the tests (k = 0.596; p < or = 0.001). CONCLUSION: Spatial heterogeneity must be interpreted with great caution, taking into account the available evidence about risk factors for specific causes of death and carefully evaluating available exposure data. PMID- 15291394 TI - [The proposal of a harmless alternative to female genital mutilation]. PMID- 15291395 TI - [Health education: a proposal for the schools]. PMID- 15291396 TI - Dietary modification of the intestinal microbiota. AB - Humans harbor a consortium of commensal bacteria in their gut that are thought to be crucial for normal health. However, the extent of microbial diversity in the gut and the physiologic functions of the microflora have not yet been fully characterized. Molecular tools are now available to characterize the associations between diet, microflora composition, and health in greater depth. New molecular studies have confirmed earlier culture-based observations that diet has a role in the regulation of microflora composition. In the near future, new insight into these associations should allow for the design of specific diets aimed at improving health by modulating microflora. PMID- 15291397 TI - Cre/loxP system for generating tissue-specific knockout mouse models. AB - Alteration of the mouse genome by conventional transgenic and gene-targeted approaches has greatly facilitated studies of gene function. However, a gene alteration expressed in the germ line may cause an embryonic lethal phenotype resulting in no viable mouse to study gene function. Similarly, a gene alteration may exert its effect in multiple different cell and tissue types, creating a complex phenotype in which it is difficult to distinguish direct function in a particular tissue from secondary effects resulting from altered gene function in other tissues. Therefore, methods have been developed to control conditions such as the timing, cell-type, and tissue specificity of gene activation or repression. This brief review provides an overview of the Cre/LoxP system for generating tissue-specific knockout mouse models. PMID- 15291398 TI - Exploiting micronutrient interaction to optimize biofortification programs: the case for inclusion of selenium and iodine in the HarvestPlus program. AB - Biofortification of staple food crops with micronutrients by either breeding for higher uptake efficiency or fertilization can be an effective strategy to address widespread dietary deficiency in human populations. Selenium and iodine deficiencies affect a large proportion of the population in countries targeted for biofortification of staple crops with Zn, Fe, and vitamin A, and inclusion of Se and I would be likely to enhance the success of these programs. Interactions between Se and I in the thyroid gland are well established. Moreover, Se appears to have a normalizing effect on certain nutrients in the body. For example, it increases the concentration of Zn and Fe at key sites such as erythrocytes when these elements are deficient, and reduces potentially harmful high Fe concentration in the liver during infection. An important mechanism in Se/Zn interaction is selenoenzyme regulation of Zn delivery from metallothionein to Zn enzymes. More research is needed to determine whether sufficient genetic variability exists within staple crops to enable selection for Se and I uptake efficiency. In addition, bioavailability trials with animals and humans are needed, using varying dietary concentrations of Se, I, Zn, Fe, and vitamin A to elucidate important interactions in order to optimize delivery in biofortification programs. PMID- 15291399 TI - Tissue-specific knockout defines peroxisome proliferator-activated receptor gamma function in muscle and liver. AB - Although the nuclear receptor peroxisome proliferator-activated receptor gamma (PPARgamma) is expressed mostly in adipose tissue, it also occurs in skeletal muscle and liver. Mice lacking a functional PPARgamma gene, specifically in muscle, showed increased adiposity with reduced caloric intake, resulting from shunting of lipid to the liver. This result suggested a function for muscle PPARgamma in the up-regulation of muscle lipid metabolism. A study in lipoatrophic mice--that were also defective in the liver PPARgamma gene- suggested that liver PPARgamma functions in the uptake and clearance of fat in that organ. However, PPARgamma in liver has an antidiabetic role, whereas muscle PPARgamma was found to have no part in opposing diabetes. PMID- 15291400 TI - Early milk intake, later bone health: results from using the milk history questionnaire. AB - Milk intake is considered an important determinant of peak bone mass; consequently, its intake early in life may decrease risk of osteoporotic fractures later. Using the milk history questionnaire, many investigators have conducted retrospective cross-sectional studies and have determined a positive effect of milk intake in childhood and adolescence on bone mineral density in adult white women. By contrast, a recent study indicates that early milk intake has no beneficial effect on bone mineral density in black women. The milk history questionnaire can assist in determining diet-bone relationships in various groups. PMID- 15291401 TI - Nutrition support research: it is our obligation to do better! PMID- 15291402 TI - Supplementation of total parenteral nutrition with butyrate acutely increases structural aspects of intestinal adaptation after an 80% jejunoileal resection in neonatal piglets. AB - BACKGROUND: Supplementation of total parenteral nutrition (TPN) with a mixture of short-chain fatty acids (SCFA) enhances intestinal adaptation in the adult rodent model. However, the ability and timing of SCFA to augment adaptation in the neonatal intestine is unknown. Furthermore, the specific SCFA inducing the intestinotrophic effects and underlying regulatory mechanism(s) are unclear. Therefore, we examined the effect of SCFA supplemented TPN on structural aspects of intestinal adaptation and hypothesized that butyrate is the SCFA responsible for these effects. METHODS: Piglets (n = 120) were randomized to (1) control TPN or TPN supplemented with (2) 60 mmol/L SCFA (36 mmol/L acetate, 15 mmol/L propionate and 9 mmol/L butyrate), (3) 9 mmol/L butyrate, or (4) 60 mmol/L butyrate. Within each group, piglets were further randomized to examine acute (4, 12, or 24 hours) and chronic (3 or 7 days) adaptations. Indices of intestinal adaptation, including crypt-villus architecture, proliferation and apoptosis, and concentration of the intestinotrophic peptide, glucagon-like pepide-2 (GLP-2), were measured. RESULTS: Villus height was increased (p < .029) within 4 hours by supplemented TPN treatments. Supplemented TPN treatments increased (p < .037) proliferating cell nuclear antigen expression along the entire intestine. Indicative of an antiapoptotic profile, jejunal Bax:Bcl-w abundance was decreased (p = .033) by both butyrate-supplemented TPN treatments, and ileal abundance was decreased (p = .0002) by all supplemented TPN treatments, regardless of time. Supplemented TPN treatments increased (p = .016) plasma GLP-2 concentration at all time points. CONCLUSIONS: Butyrate is the SCFA responsible for augmenting structural aspects of intestinal adaptations by increasing proliferation and decreasing apoptosis within 4 hours postresection. The intestinotrophic mechanism(s) underlying butyrate's effects may involve GLP-2. Ultimately, butyrate administration may enable an infant with short-bowel syndrome to successfully transition to enteral feedings by maximizing their absorptive area. PMID- 15291403 TI - Effects of L-arginine infusion during ischemia on gut blood perfusion, oxygen tension, and circulating myeloid cell activation in a murine gut ischemia/reperfusion model. AB - BACKGROUND: Gut hypoperfusion is considered to be a mechanism for early multiple organ failure after severe surgical insults. L-Arginine (ARG) may preserve gut microcirculation as a substrate of nitric oxide synthase, but simultaneously may enhance immune cell response. It remains unknown if ARG infusion during gut ischemia improves the outcome after gut ischemia-reperfusion (I/R). METHODS: Male Institute of Cancer Research mice were randomized to control and ARG groups. After i.v. cannulation, mice underwent 90 (Exp. 1) or 60 (Exp. 2 and 3) minutes of gut I/R. Control mice received normal saline infusion at 1 mL/h for 60 minutes during ischemia, whereas the ARG group was given 1% ARG hydrochloride solution. In Exp. 1, survival was observed for 72 hours (n = 35). In Exp. 2, blood perfusion and oxygen tension of the small intestine were measured (n = 9). In Exp. 3, peripheral blood was obtained at 2 or 4 hours after reperfusion (n = 22). Reactive oxygen intermediate (ROI) production by myeloid cells with or without phorbol myristate acetate (PMA) stimulation and expression of CD11a and CD11b on myeloid cells were examined using flow cytometry. RESULTS: Exp. 1: There was no significant difference in survival times (log rank test, p = .2). However, survival rates at 12 hours were 72% (13/18) for the control group and 35% (6/17) for the ARG group (p < .05 Fisher). Exp. 2: ARG infusion significantly improved gut blood perfusion ratio during ischemia but had no effect on oxygen tension. Exp. 3: In the ARG group, ROI production with PMA and CD11b expression at 4 hours were higher than those at 2 hours, whereas there were no significant changes in the control mice. CONCLUSIONS: ARG infusion improves intestinal blood perfusion during ischemia but primes and activates circulating myeloid cells excessively. Consequently, i.v. infusion of ARG during ischemia reduces survival rate. PMID- 15291404 TI - Synergistic anti-inflammatory activity of omega-3 lipid and rofecoxib pretreatment on macrophage proinflammatory cytokine production occurs via divergent NF-kappaB activation. AB - Omega-3 lipid pretreatment significantly decreases TNF-alpha production in LPS stimulated Mphis; however, this response is only a partial inhibition, suggesting that other nonsubstrate- (lipid) dependent mechanisms are involved. The cyclooxygenase (COX)-2 enzyme is principally responsible for lipid metabolism; thus, a selective COX-2 inhibitor (Rofecoxib) would clarify if it is an omega-3 lipid direct effect or a COX-2 enzyme-associated modulated reduction in TNF alpha. Moreover, potential synergy between omega-3 lipids and selective COX-2 inhibition is postulated. HYPOTHESIS: Through divergent regulatory mechanisms, omega-3 lipids in combination with Rofecoxib will synergistically decrease the LPS-stimulated Mphi inflammatory response. METHODS: RAW 264.7 cells were pretreated with omega-3 lipids, Rofecoxib, or combination treatment and then washed and exposed to LPS. Supernatants were collected for ELISA, total proteins were obtained to determine COX-2 protein expression by Western blot, and nuclear extracts were isolated to determine NF-kappaB activation by electromobility shift assay. RESULTS: TNF-alpha and PGE2 production was significantly decreased with omega-3 and Rofecoxib pretreatment, and with combination treatment a further decrease in TNF-alpha production was observed. COX-2 protein expression was demonstrated to increase in omega-3, Rofecoxib, and combination groups stimulated with LPS. No alteration in NF-kappaB activation was observed with Rofecoxib or combination pretreatment compared with LPS-stimulated control cells. Repletion of prostaglandin (PGE2) in the Mphi model significantly decreased TNF-alpha in all groups. CONCLUSIONS: Omega-3 lipids and Rofecoxib independently decrease TNF alpha and PGE2 production in LPS-stimulated Mphi, yet in combination a synergistic reduction in TNF-alpha production is observed. Although the anti inflammatory effects observed from omega-3 lipids are known to occur partially through decreasing NF-kappaB activation, we demonstrated that Rofecoxib or even a combination of omega-3 and Rofecoxib does not alter NF-kappaB activation, as seen with omega-3 lipids alone. These data support that combination treatment may result in decreased Mphi inflammation, yet this occurs via divergent mechanisms. PMID- 15291405 TI - In utero malnutrition influences wound healing of newborn rats as measured by tensile strength and collagen deposition. AB - BACKGROUND: Studies have investigated the consequences of intrauterine malnutrition on birth weight and overall survival but not on wound healing. This study aims to assess the influence of in utero malnutrition on wound healing of newborn rats. METHODS: Pregnant Wistar rats were divided into 2 groups. Study rats were given 50% of the food intake of controls throughout pregnancy in a pair fed manner. The body weight and length of the newborns were measured. Newborns were breast-fed until day 21, when a laparotomy was performed. The effect of the laparotomy was assessed by measure of the wound strength and collagen deposition at postoperative day (POD) 7 (n = 15) and POD 21 (n = 15). RESULTS: The body weight and length of newborns of malnourished mothers were significantly smaller at birth compared with controls (respectively, 4.5 +/- 0.1 g vs 5.8 +/- 0.1 g, p = .0003 and 4.6 +/- 0.1 cm vs 5.2 +/- 0.1 cm, p = .0003). Maximum, rupture, and tensile strength of malnourished newborns were smaller than controls on POD 7 (0.281 +/- 0.031 vs 0.470 +/- 0.031, p = .0061, 0.112 +/- 0.06 kgf vs 0.173 +/- 0.08 kgf, p = .0495 and 0.019 +/- 0.002 kgf/mm2 vs 0.024 +/- 0.003 kgf/mm2, p = .050, respectively). On POD 21, only tensile strength remained lower (0.044 +/- 0.003 kgf/mm2 vs 0.058 +/- 0.003 kgf/mm2, p = .0477). Type I collagen deposition of malnourished newborns was similar to controls on POD 7 (57.69 +/- 10.06 vs 48.34 +/- 15.65, p = .3187) and on POD 21 (75.6 +/- 7.21 vs 80.0 +/- 9.92, p = .4212). CONCLUSIONS: In utero malnutrition decreases the abdominal wound strength of newborn rats but not the collagen deposition, suggesting that breast-feeding nutrition is effective in recovering the collagen deposition but not overall wound strength. PMID- 15291406 TI - L-arginine-enriched parenteral nutrition affects lymphocyte phenotypes of gut associated lymphoid tissue. AB - BACKGROUND: Experimentally, total parenteral nutrition (TPN) diminishes gut associated lymphoid tissue (GALT) cell numbers and function. Although glutamine supplementation is known to reverse TPN-induced changes in GALT, effects of another conditionally essential amino acid, L-arginine (ARG), on GALT remain unclear. METHODS: Twenty-two male Institute of Cancer Research mice were randomized to standard TPN (0.3% arginine, STD-total parenteral nutrition) or 1% ARG-enriched TPN (ARG-total parenteral nutrition). After 5 days of feeding, lymphocytes were harvested from Peyer's patches (PP), the lamina propria, and intraepithelial (IE) spaces of the small intestine to determine cell yields. Lymphocyte phenotypes (alphabetaTCR, gammadeltaTCR, CD4, CD8, and B220 as a B cell marker) were determined using flow cytometry. IgA levels in washings of the small intestine, upper respiratory tract, and lungs were measured with ELISA. RESULTS: ARG-total parenteral nutrition did not affect lymphocyte yields. The percentages of CD4+ cells in PP and IE, and alphabetaTCR+ cells in PP, were significantly higher in the ARG-total parenteral nutrition than in the STD-total parenteral nutrition mice, without marked differences in other phenotypes examined. There were no significant differences in intestinal and respiratory tract IgA levels between the 2 groups of mice. CONCLUSIONS: One percent ARG supplementation of TPN does not improve GALT cell number or mucosal IgA level but benefits to increase CD4+ cell percentages in GALT. PMID- 15291407 TI - Does a multidisciplinary total parenteral nutrition team improve patient outcomes? A systematic review. AB - BACKGROUND: Total parenteral nutrition (TPN) is a specialized form of nutrition support. The complexity associated with the management of patients receiving TPN therapy has led to the development of multidisciplinary TPN teams. The purpose of this review was to critically analyze the literature and present the best available evidence that investigated the effectiveness of multidisciplinary TPN teams in the provision of TPN to adult hospitalized patients. METHODS: A systematic review of studies identified from the Cochrane Library (2001, Issue 4), CINAHL, Complete MEDLINE, Complete Biomedical Collection, Complete Nursing Collection, and EMBASE, published in any language. RESULTS: Eleven studies, 4 with concurrent controls and 7 with historical controls, were eligible for inclusion in the review. Results of the studies indicate that the incidence of total mechanical complications is reduced in patients managed by the TPN team. However, the benefit of the TPN team in the reduction of catheter-related sepsis remains inconclusive. Four of the 5 studies reported fewer total metabolic and electrolyte abnormalities in patients cared for by the team, and these patients were more likely to receive their optimal caloric intake. However it was unclear if the management of the patients by the TPN team prevented the inappropriate use of TPN therapy. Although only 2 studies (n = 356) investigated total costs associated with management of patients by the TPN teams, there was evidence that a team approach is a cost-effective strategy. CONCLUSIONS: Overall, the general effectiveness of the TPN team has not been conclusively demonstrated. There is evidence that patients managed by TPN teams have a reduced incidence of total mechanical complications; however, it is unclear if there is a reduction in catheter-related sepsis and metabolic and electrolyte complications. The available evidence, although limited, suggests financial benefits from the introduction of multidisciplinary TPN teams in the hospital setting. PMID- 15291408 TI - Validation of 2 approaches to predicting resting metabolic rate in critically ill patients. AB - BACKGROUND: Indirect calorimetry is the criterion method for determining resting metabolic rate for nutrition support in critically ill patients. However, calculation equations are more commonly used. In the current study we tested the validity of 2 such calculation systems. METHODS: Indirect calorimetry was performed with an open-circuit device in mechanically ventilated surgical, trauma, and medical patients at rest. Feedings were not stopped for the measurements. Two predictive equations by Ireton-Jones and 3 versions of a multivariate equation developed at our institution (referred to as Penn State equations) were then used to estimate resting metabolic rate. These estimates were compared on a percentage basis with the measured value of resting metabolic rate. Estimated resting metabolic rate within 10% of measured was considered accurate, whereas estimations >15% different from measured were considered large errors. RESULTS: Forty-seven subjects were measured. A larger percentage of subjects were estimated accurately by the Penn State equations (72% in the best equation) than by the Ireton-Jones equations (60% in the best equation; not significant). The incidence of errors >15% of measured was significantly lower in the Penn State equation (11% of subjects) compared with the Ireton-Jones equation (32% of subjects) (p < .05). CONCLUSIONS: The Penn State equation for resting metabolic rate in mechanically ventilated intensive care patients receiving nutrition support appears to be a valid clinical tool for determining energy goals in the absence of or as a supplement to indirect calorimetry. The Ireton Jones equation performed less well, especially in that a higher number of large errors occurred. PMID- 15291409 TI - Use of clonidine to decrease intestinal fluid losses in patients with high-output short-bowel syndrome. AB - BACKGROUND: Some patients who have short bowel syndrome (SBS) have high intestinal outputs, which increases the risk of dehydration and fluid-electrolyte abnormalities and impairs quality of life. METHODS: We evaluated the use of clonidine, an alpha2-adrenergic receptor agonist, in 2 patients with SBS and high intestinal outputs, who were refractory to conventional therapy with antidiarrheal and antisecretory agents (loperamide, belladonna, opiates, somatostatin, histamine2 receptor antagonists and proton pump inhibitors). The first patient (case 1) was a 29-year-old woman who had 175 cm of small bowel anastamosed to 8 cm of descending colon, ending in a colostomy. The second patient (case 2) was a 22-year-old man who had 30 cm of jejunum anastomosed to 30 cm of sigmoid colon and rectum. RESULTS: The addition of clonidine, 0.1 mg per os (PO) 2 times per day, to the treatment regimen of case 1 decreased her ostomy output from approximately 4 L/day to approximately 1 L/day, eliminating the need for parenteral nutrition and fluids and decreasing the need for opiate therapy. The addition of clonidine, 0.2 mg PO 2 times per day in case 2 resulted in decreased rectal outputs from approximately 4 L/day to approximately 1.5 L/day. CONCLUSIONS: These results demonstrate that clonidine can effectively reduce intestinal fluid and electrolyte losses and should be considered as an additional treatment option in patients with SBS and high intestinal outputs. PMID- 15291410 TI - Long-term parenteral nutrition, via the azygos system, in an adolescent with cystic fibrosis. AB - Venous access device is critically needed for long-term parenteral nutrition (PN), especially in children with chronic disease such as cystic fibrosis, short bowel syndrome, and permanent intestinal failure. When traditional sites are unavailable and venous access is very limited, alternative central routes are required. The access to the azygos system has been shown to be a safe, relatively easy solution in those special situations. We report the case of an adolescent who benefited from this central venous access for long-term PN and antibiotic treatment for >7 years with a limited number of complications. PMID- 15291411 TI - The ubiquitin-proteasome proteolysis pathway: potential target for disease intervention. PMID- 15291412 TI - Immunonutrition: back to science. PMID- 15291413 TI - Improving nutrition screening of hospitalized patients. PMID- 15291414 TI - Clinical dilemma: which energy expenditure equation to use? PMID- 15291415 TI - The proteasome. PMID- 15291416 TI - ['Hyperventilation syndrome': often an easy to treat panic disorder]. AB - Three patients, 2 men aged 35 and 26 years and 1 woman aged 41 years, had acutely occurring attacks, accompanied by diverse somatic complaints, and were diagnosed with hyperventilation syndrome. They recovered only when the complaints were recognised and treated as a panic disorder. Hyperventilation and the decrease of CO2 in the blood do not explain the symptoms and complaints in patients with panic disorder, a psychiatric disorder with a good prognosis. Treatment consists of cognitive behavioural therapy or a selective serotonin re-uptake inhibitor in the case of panic disorder and of a combination of those two treatments in the case of panic disorder with agoraphobia. Breathing exercises can form part of the behavioural therapy but not because the disorder is due to faulty breathing habits. PMID- 15291417 TI - [Mental competence in the context of deep brain stimulation]. AB - In a case of Parkinson's disease, the patient was treated with deep brain stimulation of the subthalamic nucleus (STN-DBS). STN-DBS affected the mental competence of the patient and ethical questions were raised about the decision as to the direction of further treatment. The patient was asked for his opinion on the therapeutic options during a phase of non-stimulation and chose to be stimulated and admitted to a psychiatric hospital because of mental incompetence rather than remaining unstimulated, mentally competent but bedridden. Developments in the neurosciences (including STN-DBS) raise a number of different fundamental (theoretical and philosophical) as well as practical questions. STN DBS can have various unintended (behavioural) effects. In the case presented, more weight was rightly given to the mental competence of the unstimulated patient, although comments can be made with regard to his decision making, as his choice was made in a phase of serious distress. Attention is paid to the relevance of a so-called self-binding directive. STN-DBS is not morally neutral and the case involves a tragic dilemma: a conflict between irreconcilable duties for the physician. The further development and proliferation of STN-DBS requires caution and moral deliberation. It remains important to search for alternative treatment strategies with less undesirable side effects. PMID- 15291418 TI - [Cellular microparticles and blood-vessel damage. I. Structure, detection and origin]. AB - In virtually all eukaryotic cells, activation and apoptosis lead to the formation of vesicles or microparticles by intracellular mechanisms which as yet are not completely understood. Flow cytometry, electron microscopy and ELISA techniques can be used to detect microparticles. Microparticles are a heterogeneous population and their numbers, cellular origin, composition and functional characteristics, both in vitro and in vivo, depend on the circumstances under which they were generated. Microparticles derived from various cells, primarily platelets but also lymphocytes, granulocytes, monocytes, erythrocytes and endothelial cells, are present in the circulation of healthy subjects. Elevated numbers of microparticles can be found in a wide variety of diseases, all of which are associated with hypercoagulability and blood-vessel damage, thus suggesting their role in the pathogenesis of vascular disease. PMID- 15291419 TI - [Cellular microparticles and blood-vessel damage. II. Functional characteristics and clinical significance]. AB - Cellular microparticles support coagulation by exposure of negatively charged phospholipids and sometimes tissue factor. They are involved in inflammatory processes, the transfer of membrane antigens and bioactive molecules, and in the modulation of endothelial functions. Patients with disturbances in membrane vesiculation, leading to decreased numbers of circulating microparticles, present clinically with an increased bleeding tendency. In contrast, elevated numbers of microparticles are found in a great variety of diseases involving blood-vessel damage and hypercoagulability. Microparticles are also a major component of human atherosclerotic plaques. In view of their functional properties, cellular microparticles may be a missing link between cellular and plasmatic processes underlying atherosclerotic blood-vessel damage. PMID- 15291420 TI - [Identification of cadavers in the Netherlands]. AB - The identification of cadavers of unknown persons is important from a humanitarian point of view but also for legal reasons and in connection with insurance. Various identification techniques are available today, such as fingerprinting, DNA profiling and the comparison of dental structures. Not all methods of identification are equally useful in practice and the ultimate identification is often made possible only by a combination of several techniques. Sometimes, the identification procedure cannot be completed successfully because one has no idea whatsoever who it might be and there is therefore no material available for comparison. In the Netherlands, identification is in principle a task of the police. In case of a mass disaster in the Netherlands, or abroad when Dutch citizens are involved, a special Disaster Identification Team is activated, consisting mainly of police officers supplemented by external experts. Major disasters in which such a team has been involved in the past are the air crash in Faro (Portugal) in 1992 (56 victims), the air crash in Eindhoven in 1996 (32 victims) and the explosion of the fireworks plant in Enschede in 2000 (21 victims). PMID- 15291421 TI - [Diagnostic image (197). A man who suddenly collapsed]. AB - A 40-year-old man suffered a witnessed collapse with brief loss of consciousness, due to electrical injury. PMID- 15291422 TI - [Surveillance of hepatitis A in the Netherlands 1993-2002]. AB - OBJECTIVE: To survey trends in data on hepatitis A using information from the notification system of the Municipal Medical and Health Services 1993-2002. DESIGN: Retrospective, descriptive. METHOD: Data collected from the notification system of the Dutch Municipal and Medical Services were analyzed. RESULTS: From 1993 to 2002, 64.4%, 6.5% and 11.5% of infections were contracted in the Netherlands, Turkey and Morocco, respectively. Although the absolute number of cases was highest for young people under the age of 16 infected in the Netherlands, incidence rates showed that the children of immigrants infected in Turkey or Morocco were at highest risk, followed by adult immigrants from Turkey and Morocco. In addition, a seasonal trend was observed starting with an increase in the number of notifications of young travellers infected in Turkey and Morocco during their summer holiday, followed by a steep increase in notifications of young people infected in the Netherlands in autumn. This was followed later in the year by a slight increase in notifications of adults who acquired the infection in the Netherlands. However, between 1993-2002 the total number of notifications was halved. This decrease was mainly restricted to infections acquired in the Netherlands. In recent years, fewer outbreaks have been reported in schools, households and families. In 2001, a peak of notifications from the homosexual scene was observed. There was a continued trend in the rise of the mean age of hepatitis-A onset. CONCLUSION: Risk of hepatitis A is highest for the children of immigrants travelling to Turkey or Morocco in the summer months. Secondary cases in the Netherlands are in strong decline, especially in schools and families. However, the almost stable incidence of infections among young people of Turkish or Moroccan extraction stresses the continued importance of immunization of this group before they travel. The continuing trend in the rise of the mean age of hepatitis-A onset deserves attention because of the increasing risk of mortality of the disease amongst older people. PMID- 15291423 TI - [Manipulation of mental competence: an ethical problem in case of electrical stimulation of the subthalamic nucleus for severe Parkinson's disease]. AB - Three years after the implantation of electrodes in the subthalamic nucleus (STN) and the start of deep brain stimulation (DBS) for advanced Parkinson's disease, a 62-year-old man was admitted because of a stimulation-related manic state that did not respond to treatment with a mood stabiliser and that led to chaotic behaviour, megalomania, serious financial debts and mental incompetence. Although adjustment of the stimulation parameters resulted in a normophoric state with a return of insight and capacity to judge, this was only at the cost of a serious exacerbation of his motor symptoms that left the patient bedridden. There was no therapeutic margin between the two states. Ultimately, there seemed to be only two alternatives: to admit the patient to a nursing home because of serious invalidity, but mentally in good condition, or to admit the patient to a chronic psychiatric ward because of a manic state, but with acceptable motor capacity and ADL functions. Thorough ethical evaluation followed. When not being stimulated, the patient was considered competent to decide about his own treatment; in this condition the patient chose for the second option. In accordance with his own wishes he was therefore legally committed to a chronic ward in the regional psychiatric hospital. The current ethical views on mental competence do not consider the potential influence of modern methods of treatment such as STN-DBS on this capacity. PMID- 15291424 TI - [Posttraumatic hypopituitarism]. AB - A 39-year-old woman was admitted with complaints of weight gain, a decreased sense of well-being and amenorrhoea. One and a half year prior to admission she had been involved in a serious road accident and had spent several days in coma due to an epidural haematoma. She was found to have hypopituitarism with deficient somatotropic and gonadotropic axes, as well as mild hyperprolactinaemia, probably due to a pituitary stalk lesion. All patients with severe trauma to the skull are at risk of developing posttraumatic hypopituitarism, so that pituitary function testing should be performed routinely, certainly in the presence of symptoms. PMID- 15291425 TI - [Prevention of cardiovascular complications after a stroke or TIA: hypotensive and hypocholesterolemic therapy]. PMID- 15291426 TI - [A cavernous haemangioma of the colon as the cause of rectal bleeding in childhood]. PMID- 15291428 TI - [Recognition and treatment of gamma hydroxybutyric acid poisoning]. PMID- 15291427 TI - [Recognition and treatment of gamma hydroxybutyric acid poisoning]. PMID- 15291429 TI - Lilian of Belgium, a patron for cardiology. PMID- 15291430 TI - Challenges of cardiology and cardiovascular surgery in 1958. AB - The Cardiology Foundation supported by the perspicacious Princess Lilian was created in 1958. As an introduction to this symposium, the challenges that the cardiologist and surgeon encountered in 1958, are recalled as well as the essential problems requiring solutions in that era of evolving cardiac surgery. PMID- 15291431 TI - Cardiac surgery in Belgium. State of the art in 2003. AB - Cardiac surgery in Belgium has reached since many years a high quality level and improves continuously. In mitral valve surgery, repair of the valve has been substituted to its replacement. Repair is now the standard operation for degenerative pathology. Long-term results are well established and indications standardised. In aortic valve surgery, prolapse of a leaflet, dilatation of the sinotubular junction, aneurysm of the sinus of Valsalva are now repaired with very good results. When the valve is unrepairable, a wide choice of available homograft, stentless valve or aortic root allows the most adequate operation. In coronary artery bypass graft (CABG), arterial grafts are increasingly used over the least years because of their higher long-term patency rates. Off-pump coronary bypass, that is CABG on the beating heart, is more and more frequently performed from 2.4% in 1998 to 17.9% of the cases in 2000. The development of mechanical circulatory support, also called "artificial heart" is encouraging but still far from perfect. During the 2 last years, 31 devices were implanted in Belgium. Twenty patients were either successfully transplanted or weaned from the pump and returned to their home. Cardiac surgery in Belgium is still developing and remains of high quality thanks to the continuous introduction of new therapeutic approaches and to the close collaboration between intensivists, anaesthesiologists, perfusionists and cardiologists. As a result, patients benefit today from the best therapeutic modalities. PMID- 15291432 TI - Congenital heart surgery in Belgium. State of the art in 2003. AB - Congenital heart surgery has evolved from its pioneering days to a multidisciplinary speciality with important scientific contributions from different fields. Progress became obvious in different fields. Surgical results improved considerably: immediate survival as well as long-term outcome. Interventional cardiology emerged and took care of a large number of easy surgical cases. It allowed the surgeon to address the more complex cardiac deformities, often in combination with interventional techniques. Therefore the daily surgical practice of the congenital heart surgeons has changed to more complex pathology and procedures. Very recent fields of evolution are the generation of knowledge in genetic predisposition and prenatal diagnosis. Both give additional information to improve parent information and to optimise treatment. PMID- 15291433 TI - New developments in cardiac surgery. AB - In cardiac surgery new developments in the current area are mainly directed towards minimally invasive surgery techniques. New anastomotic devices for CABG are clinically available and will facilitate partial and total endoscopic CABG techniques. In valve surgery percutaneous techniques are already available for AVR and PVR. Single percutaneous mitral valve repair is not a too far away procedure. Tissue engineering will offer biocompatible heart valves. New developments for heart failure therapy are booming. The major drawback for the achievements of this amazing progress is the financial cost. An open debate between all the parties concerned in healthcare is urgently needed in order to maintain the quality of surgical care for the decades to come. PMID- 15291434 TI - Cardiac rhythm disorders: problems of the present, challenges for the future. AB - Arrhythmology has become a well-confirmed subspecialty in cardiology. Many different fields of rhythm disorders are rapidly and constantly evolving. New and more specific oriented antiarrhythmic drugs are under evaluation, especially for the treatment of atrial fibrillation. Sudden cardiac death can be prevented by implantable defibrillators but precise selection of patients at risk and the cost remain still a major health problem. We also are unable at present to treat the substrate and to prevent the occurrence of the lethal arrhythmias which are responsible for sudden cardiac death. Pacemakers are widely used for the treatment of bradycardia but other indications are emerging like the treatment of advanced heart failure through resynchronization by biventricular pacing, as well as remote pressure monitoring in patients suffering from cardiac insufficiency. Cell therapy might one day replace pacemaker devices by correcting the failing automaticity or conduction of the heart. Atrial fibrillation is the most common arrhythmia for which ablation techniques might appear as a definitive treatment possibility but technical improvements are still awaited. PMID- 15291435 TI - Bringing the heart into focus: a perspective on non-invasive cardiac imaging. AB - Non-invasive cardiac imaging is already a cornerstone of cardiac diagnostics and will play an increasingly important role also in preventive and therapeutic cardiology. Several modalities can be used (echocardiography, nuclear, CT, CMR) and there is a need to study the respective role of these techniques in clinical practice and to anticipate on developments in the near future. PMID- 15291436 TI - Adventures in pharmacology, aspirin, prostacyclin and nitric oxide. PMID- 15291437 TI - Hartford Hospital surgeons share publicity with robot 'assistants.'. AB - Hartford Hospital, Hartford, Conn. finds some patients gravitate to Boston or New York City for high-tech procedures. Being the first Connecticut hospital with the daVinci Robotic Surgery System, it developed an advertising campaign emphasizing high-tech surgery with the human touch. Hartford also broadcast a robotic prostatectomy on its popular live webcast series. PMID- 15291438 TI - Healthcare marketers share vital role in patient satisfaction. PMID- 15291439 TI - ECHN creates a 'new look' for ads: series promotes new physicians, job opportunities and more. AB - Eastern Connecticut Health Network, (ECHN), Manchester, Conn., is using a "new look" for ads that announce the addition of new staff physicians. In view of the recent growth of the network, there was a need to build public awareness of ECHN's wide range of specialties. The new ads use informal photos of each new doctor with the tagline "A Welcome Choice," or a variation of the same phrase. PMID- 15291440 TI - Three cancer reports to think about: each approaches its objective in a unique manner. AB - Three cancer reports are profiled here. The 2002 Annual Report of INTEGRIS Oncology, Oklahoma City, Okla., was produced in customized versions for two regions. Ochsner Cancer Institute's Annual Report for 2002, New Orleans, focuses on cancer survivors, and the institute's multidisciplinary approach is featured throughout. And, the 2002 report for University Health Care System's Harry W. Jernigan Jr. Cancer Center, Augusta, Ga., uses a horizontal format, glossy paper and lots of white space to convey a "fresh" feeling not often seen in cancer reports. PMID- 15291441 TI - Pekin hospital says 'small is good': ongoing campaign emphasizes "Personal size. Personal care.". AB - Pekin Hospital, Pekin, Ill., launched a campaign in April 2003 to differentiate itself from the larger, big-city hospitals. Using non-healthcare images and emphasizing individualized care, the "Personal Size. Personal Care," campaign was well received and is expected to continue for some time. PMID- 15291442 TI - Anne Arundel opens world's first See and Treat Cancer Center. AB - Anne Arundel Medical Center (AAMC), Annapolis, Md., is promoting he world's first See and Treat Cancer Care Center of Excellence at The DeCesaris Cancer Institute. The system uses medical imaging equipment from GE Healthcare and radiation therapy technologies from Varian Medical Systems. PMID- 15291443 TI - Medical group forges information link with television station web site. AB - Methodist Medical Group, Peoria, Ill, has entered into an interesting partnership with the local WEEK-TV to provide timely informative health related information. Through the WEEK web site, members of the public have access to a variety of locally produced health related articles by clicking on the MedWatch tab at www.week.com. Viewers can request specific topics of interest by e-mail. PMID- 15291444 TI - [From the sources of public health to the Free University of Brussels]. PMID- 15291445 TI - [Subclinical hypothyroidism]. AB - Subclinical hypothyroidism is defined as normal serum free thyroxine and tri iodothyronine concentrations and a slightly eleveted serum thyrotropin (TSH) concentration. Only laboratory results can detect this disorder. The causes of this disease are the same as those of overt hypothyroidism. Most patients with subclinical hypothyroidism should be treated with thyroxine aiming to reduce the patient's serum TSH concentration to normal. Treatment will prevent progression to overt hypothyroidism and in some cases ameliorate non specific symptoms and lipid profile. PMID- 15291446 TI - [How to cope with tinea capitis?]. AB - In Brussels, as in the other European cities with a high rate of immigration, the cases of anthropophilic tinea capitis have risen. Their clinical aspect is more discrete than in zoophilic cases, and you could be easily fooled. This explains why, in 2001-2002, we had some epidemics in schools and creches. Griseofulvin, the gold treatment, was withdrawn from the Belgian market in 1997. The identification of the pathogen when using the new antifungals (fluconazole, itraconazole and terbinafine) is mandatory, dermatophytes showing a different sensitivity to these drugs. The management of an anthropophilic tinea capitis needs some good planning and the cooperation of the family and the school. We describe our procedures in this article. PMID- 15291447 TI - [New therapeutic approachs in melanoma]. AB - During the last decade, new insights in cellular and molecular biology have opened new avenues in cancer immunotherapy. Two distinct modalities have been developed: adoptive immunotherapy and anti-tumoral vaccination (active immunotherapy). We will first describe the main strategies of adoptive immunotherapy and then elaborate on the protocols of anti-tumoral vaccination against tumor associated antigens (TAA). In that context, we will pay peculiar attention on the pivotal role of dendritic cells (DC) as natural adjuvant. PMID- 15291448 TI - [Ten years follow-up of medically assisted procreation in Belgium]. AB - Since 1989, the data concerning medically assisted procreation (MAP) in Belgium have been registered through the BELRAP association (Belgian Register for Assisted Procreation). In 1999, the national College of Physicians in Reproductive Medicine has been installed and given the mission to carry out the quality controls and the registration of MAP activities in our country. In 2001, a new on line system of registration of all treatment cycles was started, the basic structure of the corresponding data base having undergone a series of changes along the years. Although the number of MAP trials has considerably increased in our country, mainly because of the introduction of new techniques such as intracytoplasmic injection of sperm (ICSI), the success rate per trial has remained almost unchanged, becoming stabilized around 18.8% of ongoing pregnancies per initiated cycle. The main modification that occurred during the analyzed period has been the reduction of the number of embryos transferred at each trial, being first reduced from 3 to 2 to evolve more recently towards the replacement of only one embryo per trial. Multiple pregnancies are indeed the chief complication of MAP and in order to avoid it Belgian centers have been among the first to reduce drastically the number of transferred embryos. In 2003, legal rules were enacted, fixing the number of replacable embryos in accordance to the patient's age and to the rank of trial, while also solving the problem of the long-awaited reimbursement of MAP by our national social security system. PMID- 15291449 TI - [Voriconazole: a new weapon against invasive fungal infections]. AB - Voriconazole is a fluoropyrimidine derivative of fluconazole with an extended spectrum of activity, non-linear pharmacokinetic characteristics, available intravenously and orally with an excellent bioavailability, and a good penetration into tissues including the brain. It is metabolized in the liver by the cytochrome P450 and less than 1% is eliminated in the urine. Voriconazole has been studied extensively in numerous randomized clinical trials of invasive fungal infections and became the therapy of choice of invasive aspergillosis, fusariosis and scedosporiosis. Voriconazole is an alternative for invasive candidiasis refractory or resistant to fluconazole. Voriconazole has a good tolerability and acceptable safety profile and has added a new weapon to our therapeutic armamentarium against fungi. PMID- 15291450 TI - [Oral verrucous carcinoma]. AB - Verrucous carcinoma is a rare, low-grade, well-differentiated form of squamous cell carcinoma seen on skin and mucosa. It is a slow-growing and locally aggressive tumor whose standard treatment is surgery. A case of strongly invasive oral verrucous carcinoma is presented with the medical history. The differential diagnosis of this neoplasm is difficult and requires clinic and pathologic data confrontation. In this paper, the importance of knowing the malignant potential of this lesion is stressed. Indeed, in 20% of verrucous carcinoma, foci of squamous cell carcinoma can be found. A review of literature and a differential diagnosis of verrucous carcinoma are presented. PMID- 15291451 TI - [Usual interstitial pneumonia]. AB - We report the case of a 49-year old woman with an idiopathic pulmonary fibrosis (IPF) initially diagnosed as a systemic lupus erythematosus. The IPF is an uncommon clinical entity with an estimated prevalence from 3 to 6 cases per 100,000 in the general population of the United States. This disease is characterised by an insidious onset, a pejorative course and poor survival prognosis (median survival: 2.8 years). The diagnosis is often difficult and depends on the exclusion of other diseases associated with interstitial lung injury. It is generally established only after collegial coordination between the clinician, the radiologist and the pathologist. New consensuses are now published to establish a clear and explicit classification of the IPF. Moreover, because of the poor results obtained with conventional immunosuppressive drugs, new treatments are proposed. PMID- 15291452 TI - [Gallstone ileus. Abdominal CT usefulness]. AB - The authors report the case of a 93-year old woman referred to the emergency department and presenting with an intestinal obstruction. Abdominal CT reveals a biliary ileus caused by the migration and the impaction of a 3 cm gallstone in the small bowel. Surgical treatment by enterolithotomy was successful. In over 90% of cases, gallstone ileus is a complication of cholelithiasis and accounts for 25% of intestinal obstruction in patients over 65 years. To reduce morbidity and mortality, early diagnosis and prompt treatment are essential. Abdominal CT scan is the gold standard technique. PMID- 15291453 TI - [Physician-assisted death. An essay of clarification]. PMID- 15291454 TI - [Interest in the treatment of depression. Proceedings of the European Congress "The physician in the presence of depression", Brussels, Apr 2-3, 2004]. PMID- 15291455 TI - A novel approach to the quantification of bovine milk in ovine cheeses using a duplex polymerase chain reaction method. AB - A duplex Polymerase Chain Reaction (PCR) method able to detect bovine milk in ovine cheeses was developed. This method is based on the mitochondrial 12S and 16S rRNA genes to generate fragments of different lengths. The proposed methodology presents an alternative DNA extraction procedure faster and more economical than the kits commercially available. A linear normalized calibration curve was obtained between the log of the ratio of the bovine band intensity and the sum of bovine and ovine band intensities versus the log of cow's milk percentage. The method was applied successfully to the detection and quantification of raw, pasteurized, and powdered bovine milk in different cheeses. The proposed duplex PCR provides a simple, sensitive, and accurate approach to detect as low as 0.1% bovine milk in cheeses and to quantify bovine milk in ovine cheeses in the range of 1-50%. PMID- 15291456 TI - Are molecular weights of proteins determined by superose 12 column chromatography correct? AB - Our research on several proteins indicates that accurate molecular weights cannot be determined by Superose 12 column chromatography. In support of this statement, we present data on molecular weights of purified red kidney bean alpha-amylase inhibitor (RKB alphaAI) and white kidney bean alpha-amylase inhibitor (WKB alphaAI) to document this problem. The molecular weight of purified RKB alphaAI determined by Sephadex G-100 gel filtration, polyacrylamide gel electrophoresis, Superose 12 gel filtration and cDNA were 49.0, 51.0, 22.9, and 49.805 kDa (not glycosylated), respectively. The molecular weights of WKB alphaAI by several methods were as follows: Sephadex G-100 gel filtration, 51.0 kDa; Superose 12 gel filtration in 0.2 M NaCl buffer, 23.1 kDa; polyacrylamide gel electrophoresis (PAGE), 51.0 kDa; sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS PAGE), 45.0 kDa; multiangle laser light scattering (MALLS), 49.940 kDa; laser assisted time-of-flight mass spectrometry (LATOFMS), 56.714 kDa; and cDNA sequence (with 12.2% carbohydrate), 55.9 kDa. The data indicate there is ionic interaction between proteins and the matrix of Superose 12 in low ionic strength buffers and hydrophobic interaction at higher ionic strength buffers. Researchers should be cautious when using Superose 12 columns for molecular weight determinations. PMID- 15291457 TI - Determination of ginsenosides Rb1, Rc, and Re in different dosage forms of ginseng by negative ion electrospray liquid chromatography-mass spectrometry. AB - A method based on high-performance liquid chromatography (HPLC) and negative ion electrospray mass spectrometry (LC-MS) has been used to determine ginsenosides Rb1, Rc, and Re in six different samples of ginseng. These included a liquid extract, capsules, tea bags, and an instant tea. It was found that four of the six samples had detectable levels of at least one of the ginsenosides. The liquid extract, capsules, instant tea, and tea bags labeled ginseng had ginsenosides. The labels on the two samples that did not have ginsenosides indicated that they were a mixture of green tea, licorice, and ginseng. Also, 13C NMR was used to identify the types of complex carbohydrates present in the samples. One of the samples of tea bags had none of the ginsenosides, but did have complex carbohydrates found in most of the other samples. The instant tea had all three ginsenosides, but had no complex carbohydrates, only sucrose. The amounts of ginsenosides found in standard doses from six different sources of ginseng varied considerably. It was found that steeping a tea bag for a longer time than that recommended on the label produced a larger recovery of ginsenosides and that reusing a tea bag produced even higher recoveries. PMID- 15291458 TI - Parent and harvest year effects on near-infrared reflectance spectroscopic analysis of olive (Olea europaea L.) fruit traits. AB - The influence of parent and harvest year on the determination of oil, moisture, oleic acid, and linoleic acid contents in intact olive fruit was studied by near infrared spectroscopy (NIRS). Spectral data from 400 to 1700 nm were recorded on 437 fruit samples collected in 1996 and 1997 from seedling plants derived from three different female parents. Partial least squares models were developed using samples for each year and for each female parent separately and were validated against the other groups. Calibration models were accurate enough to predict all constituents in new samples from a different female parent but were not transferable across years. However, a calibration equation of sufficient accuracy was obtained from the combined data set (r values of 0.94, 0.93, 0.84, and 0.88 and RMSECV values of 1.33, 1.88, 4.73, and 2.91 for oil, moisture, oleic acid, and linoleic acid contents, respectively). These results demonstrate the utility of NIRS as a selection tool in olive breeding programs. PMID- 15291459 TI - Glutamine in commercial liquid nutritional products. AB - Total glutamine concentrations in commercial nutritional products have been determined by enzymatic hydrolysis followed by HPLC quantification of free glutamine and free pyroglutamic acid. Hydrolysis was accomplished by a published three-enzyme (Pronase, leucine aminopeptidase, prolidase), 20-h/37 degrees C digestion. Glutamine was determined as its FMOC derivative by reverse phase HPLC fluorescence, and pyroglutamic acid was determined directly by organic acid HPLC UV. Approximately 4.11% of the released glutamine is converted to pyroglutamic acid during the 20-h digestion. Experimental ratios of enzyme hydrolysis glutamine to acid hydrolysis glutamic acid + glutamine + pyroglutamic acid (GLX) indicate that the method recovers >90% of the protein-bound glutamine. The nutritional products with casein dominant intact protein systems typically deliver >9 g of glutamine/100 g of protein, or approximately 40 g of glutamine/100 g of GLX. PMID- 15291460 TI - Nondestructive observation of bovine milk by NMR spectroscopy: analysis of existing States of compounds and detection of new compounds. AB - In this study were successfully observed the one- (1H, 13C) and two-dimensional (1H-13C, 1H-15N, 1H-31P) NMR spectra of milk directly without any pretreatment. The signals of each NMR spectrum were assigned, and their existing states were also analyzed. Lactose existed in a free state in milk. The signals due to the butyric acid chain can be assigned among the other fatty acid chains. Monounsaturated fatty acid (oleic acid chains) and polyunsaturated fatty acid chains (linoleic and linolenic acid) were assigned by their characteristic signals. The signals from citrate, N-acetylcarbohydrates, and lecithin could be observed directly in the 1H-13C HSQC NMR spectra; the assignment of their signals was made through the 1H-13C, 1H-15N, and 1H-31P HMBC spectra of extracted milk. Signals from creatine and N-acetylcarbohydrates were detected for the first time. PMID- 15291461 TI - Development of an indirect competitive ELISA for flumequine residues in raw milk using chicken egg yolk antibodies. AB - To detect flumequine in raw milk, an indirect competitive enzyme-linked immunosorbent assay (ELISA) was developed. By carbodiimide conjugation, flumequine was conjugated to cationized bovine serum albumin (cBSA-flumequine) and to cationized ovalbumin (cOVA-flumequine). For the immunization of chickens, cBSA-flumequine was used, which allowed the isolation of specific chicken egg yolk immunoglobulins (IgY) for flumequine. As the coating antigen in the immunoassay, cOVA-flumequine was used. In the indirect competitive assay, standard flumequine was incubated together with the anti-flumequine antibodies. The antibody by which the lowest concentration of free flumequine that gives 50% inhibition of binding (IC50) was found in aqueous dilution was further tested for the applicability to detect flumequine in raw milk. An IC50 level in milk was reached that was about 5 times lower than in aqueous solution. So flumequine can be detected directly in raw milk at maximum residue level (50 microg/kg). No cross-reactivity was noticed with various related quinolones. PMID- 15291462 TI - Magnetic resonance imaging of seeds by use of single point acquisition. AB - In general, magnetic resonance imaging (MRI) is used to obtain a spatial representation of the water distribution in an object. Water in soft materials (living matter) often shows a high degree of translational mobility, giving rise to relatively long magnetic relaxation times. This allows the use of conventional MRI techniques such as the spin-echo, to acquire an image. However, when hydration levels become low, water becomes less mobile, resulting in much shorter magnetic relaxation times and a corresponding signal loss. To avoid problems arising from rapid decaying signals, we investigated the use of single point imaging (SPI) in the study of seeds. We were able to obtain SPI images of nonimbibed and imbibed seeds. Using SPI with shaped gradients significantly reduced the acoustic noise level. PMID- 15291463 TI - Ethyl acetate/ethyl alcohol mixtures as an alternative to folch reagent for extracting animal lipids. AB - The lipids of fresh egg yolk, boiled yolk, yolk powder, and raw animal tissues including pork loin, belly pork, and pork fat were extracted with the mixed solvents composed of ethyl acetate (EtOAc) and ethyl alcohol (EtOH) at 2:1 and 1:1 volume ratios, and the results were compared with those obtained with Folch reagent, that is, a mixture of chloroform and methyl alcohol (2:1, v/v). Extraction yields, lipid profiles, and fatty acid compositions were determined by weighing, TLC-FID, and GC, respectively. Data of the extracts obtained with the mixtures of EtOAc and EtOH were not significantly different from those obtained with Folch reagent, implying that the mixed solvent composed of EtOAc and EtOH (1:1 to 2:1, v/v) may replace Folch reagent, which is considered to be toxic and mutagenetic due to its component of CHCl3, for lipid extraction. PMID- 15291464 TI - Flavonoid distribution during the development of leaves, flowers, stems, and roots of Rosmarinus officinalis. postulation of a biosynthetic pathway. AB - The distribution of seven flavonoids, eriocitrin, luteolin 3'-O-beta-d glucuronide, hesperidin, diosmin, isoscutellarein 7-O-glucoside, hispidulin 7-O glucoside, and genkwanin, has been studied in Rosmarinus officinalis leaves, flowers, stems, and roots during plant growth. The maximum level reached by luteolin 3'-O-beta-d-glucuronide in leaves during June-August suggests the existence of a delay between the activation of the enzymes involved in the flavanone and flavone biosynthesis. The presence of hesperidin and diosmin in the vascular system is significant, and hesperidin shows even higher levels than the phenolic diterpenes and rosmarinic acid. The distribution of flavonoids observed in R. officinalis suggests a functional and structural relationship between phytoregulators and flavonoids, where flavonoids would be "protectors" of the activity of phytoregulators. A hypothesis for the general pathway of biosynthesis of these compounds in plants of the family Labiatae is proposed. PMID- 15291465 TI - Antioxidant capacity and lipophilic content of seaweeds collected from the Qingdao coastline. AB - Lipophilic extracts from 16 species of seaweeds collected along the Qingdao coastline were screened and evaluated for their antioxidant activities (AA) using the beta-carotene-linoleate assay system. The diethyl ether soluble extracts of all selected seaweeds exhibited various degrees of antioxidative efficacy in each screen. The highest antioxidant capacities among the tested samples were observed for Rhodomela confervoides and Symphyocladia latiuscula and were comparable with that of the well-known antioxidant butylated hydroxytoluene and greater than that of propyl gallate. The lipophilic content of all 16 samples and the chemical composition of 4 selected seaweeds, R. confervoidesand S. latiuscula, which had higher AA, Laminaria japonica, which had intermediate AA, and Plocamium telfairiae, which had lower AA, were analyzed by gas chromatography and gas chromatography-mass spectrometry, respectively. Fatty acids and alkanes were found. The present data indicated an increase in antioxidative property with increasing content of unsaturated fatty acid. The result of this study suggests that seaweeds can be considered as a potential source for the extraction of lipophilic antioxidants, which might be used as dietary supplements or in production in the food industry. This is the first report on the antioxidant activities of lipophilic extracts from seaweeds. PMID- 15291466 TI - Hypolipidemic effects of modified psyllium preparations. AB - The hypolipidemic effects of two solid-state enzymatically modified psyllium preparations were compared to that of the original psyllium husks in hamsters. Hamsters were ad libitum fed 0.2 wt % cholesterol diets formulated to contain 12% cellulose or 5% cellulose plus 7% raw or enzymatically modified psyllium preparations. Psyllium additions to the diet did not significantly alter food consumption or the weekly mean hamster weight over the 5 weeks of feeding. However, the total weight gained over 35 days of feeding of modified psyllium Y 26-4, one of the modified psyllium preparations, was significantly lower, 48, 47, and 32% than that for the cellulose, raw psyllium, and modified psyllium Y-24-3 groups, respectively. At 35 days, psyllium feeding significantly reduced plasma total cholesterol, high-density lipoprotein cholesterol, and low-density lipoprotein cholesterol by 50-100% in comparison to cellulose feeding, with no significant differences between the psyllium preparations. Fecal dry weight was unaffected by dietary treatment. At days 29-31, fecal bile acid excretion was significantly increased by 30-70% with all three psyllium diets, with no significant differences between psyllium preparations. These results suggest that improving the functional properties of psyllium by solid-state enzymatic procedures, such that its incorporation into food products is feasible, does not alter psyllium-mediated hypolipidemic effects. PMID- 15291467 TI - Screening of free radical scavenging compounds in water extracts of Mentha samples using a postcolumn derivatization method. AB - An on-line high-performance liquid chromatography-1,1-diphenyl-2-picrylhydrazyl (HPLC-DPPH*) method has been improved for the detection of polar and nonpolar radical scavenging compounds in complex plant extracts. Nine water extracts were prepared from different Mentha species, varieties, hybrids, and cultivars. After the components within each extract had been separated by reverse phase chromatography using 10-100% methanol with 2% acetic acid as a mobile phase, analytes within the eluent capable of scavenging a citric acid-sodium citrate buffered methanol 1,1-diphenyl-2-picrylhydrazyl solution were detected by postcolumn derivatization at 517 nm. The HPLC-DPPH* on-line method was applied to the qualitative and quantitative analysis of Mentha extracts. There was a strong correlation between the scavenging (negative) peak area and the concentration of the radical scavenging reference substances used. The minimum detectable concentration (microg/mL) of the antioxidant compounds was determined. Caffeic acid, eriocitrin (eriodictyol-7-O-rutinoside), luteolin-7-O-glucoside, and rosmarinic acid were identified as the dominant radical scavengers in these extracts by this method. PMID- 15291468 TI - Molecular design of multifunctional food additives: antioxidative antifungal agents. AB - A series of alkyl 3,4-dihydroxybenzoates (protocatechuates) was synthesized, and their fungicidal activity against Saccharomyces cerevisiae was assayed using a 2 fold serial broth dilution method. Nonyl and octyl 3,4-dihydroxybenzoate were noted to be the most effective against this yeast with the minimum fungicidal concentration of 12.5 microg/mL each. The activity was found to correlate with the hydrophobic alkyl chain length. The time-kill curve study showed that nonyl 3,4-dihydroxybenzoate was fungicidal against S. cerevisiae at any growth stage and this activity was not influenced by pH values. The fungicidal activity of alkyl 3,4-dihydroxybenzoates was noted in combination with their ability to disrupt the native membrane-associated function nonspecifically as surface-active agents (surfactants) and to inhibit the respiratory electron transport. However, the primary fungicidal activity of nonyl 3,4-dihydroxybenzoate likely comes from its ability to act as a surfactant. PMID- 15291469 TI - Vasomodulating potential of mediterranean wild plant extracts. AB - The incidence of cardiovascular disease and endothelial dysfunction is low in the Mediterranean area, where the major proportion of daily calories comes from plant food, high in antioxidant polyphenols. It has been shown that a reduced production or enhanced inactivation of endothelium-derived nitric oxide (NO) is involved in the onset of endothelial dysfunction. We investigated the effects of Mediterranean wild plant, that is, wild artichoke and thyme, phenolic-rich extracts on NO release by porcine aortic endothelial cells (PAECs; by using indirect methods) and by cerebral cell membrane homogenates (by using direct NO detection). NO release by PAECs was significantly potentiated by 234% and 135% by wild artichoke and thyme extracts (10(-6) mol/L), respectively. Direct detection of NO release by brain membranes also showed significantly increased NO production after wild artichoke addition (+35.4%). Further, the release of another vasorelaxant factor by PAECs, that is, prostacyclin, was significantly increased by wild artichoke and thyme (10(-6) mol/L) (+269% and +190%, respectively). Investigation of the mechanism(s) of action of wild artichoke and thyme suggests maintenance of an intracellular reduced environment, as previously shown for ascorbate. Even though these data require in vivo confirmation, they suggest that regular intake of bioactive compounds from Mediterranean wild plants contributes to maintenance of proper vasomotion and to the low incidence of atherosclerosis and endothelial dysfunction recorded in the Mediterranean area. PMID- 15291470 TI - New mosquito larvicidal tetranortriterpenoids from Turraea wakefieldii and Turraea floribunda. AB - The crude methanol extracts of the root barks of Turraea wakefieldii and Turraea floribunda were found to show mosquito larvicidal activity against third-instar larvae of Anopheles gambiae sensu stricto. Four new limonoids comprising a vilasininoid 1 and three havanensinoids 2-4 were isolated from the chloroform fractions of the methanol extracts of T. wakefieldii and T. floribunda, respectively. The structures of the compounds were elucidated by NMR spectroscopy. Compounds 1, 2, and 4 had LD50 values of 7.1, 4.0, and 3.6 ppm, respectively, and were more potent than azadirachtin, which had an LD50 value of 57.1 ppm when tested against larvae of A. gambiae. PMID- 15291471 TI - Antioxidant activity of a flavonoid-rich extract of Hypericum perforatum L. in vitro. AB - A flavonoid-rich extract of Hypericum perforatum L. (FEHP) was prepared by adsorption on macroporous resin and desorption by ethanol. Total flavonoid content of FEHP was determined by a colorimetric method. The major constituents of FEHP, including rutin, hyperoside, isoquercitrin, avicularin, quercitrin, and quercetin, were determined by HPLC analysis and confirmed by LC-MS. Different antioxidant assays were utilized to evaluate free radical scavenging activity and antioxidant activity of FEHP. FEHP was an effective scavenger in quenching DPPH and superoxide radical with IC50 of 10.63 microg/mL and 54.3 microg/mL, respectively. A linear correlation between concentration of FEHP and reducing power was observed with a coefficient of r2 = 0.9991. Addition of 150 microg of FEHP obviously decreased the peroxidation of linoleic acid during 84 h incubation, but the amount of FEHP over 150 microg did not show statistically significant inhibitory effect of peroxidation of linoliec acid (p > 0.05). FEHP exhibited inhibitory effect of peroxidation of liposome induced both by hydroxyl radical generated with iron-ascorbic acid system and peroxyl radical and showed prominent inhibitory effect of deoxyribose degradation in a concentration dependent manner in site-specific assay but poor effect in non-site-specific assay, which suggested that chelation of metal ion was the main antioxidant action. According to the results obtained in the present study, the antioxidant mechanism of FEHP might be attributed to its free radical scavenging activity, metal-chelation activity, and reactive oxygen quenching activity. PMID- 15291472 TI - Properties of protein powders from arrowtooth flounder (Atheresthes stomias) and herring (Clupea harengus) byproducts. AB - Functional, nutritional, and thermal properties of freeze-dried protein powders (FPP) from whole herring (WHP), herring body (HBP), herring head (HHP), herring gonad (HGP), and arrowtooth flounder fillets (AFP) were evaluated. The FPP samples have desirable nutritional and functional properties and contained 63 81.4% protein. All FPP samples had desirable essential amino acid profiles and mineral contents. The emulsifying and fat adsorption capacities of all FPP samples were higher than those of soy protein concentrate. The emulsifying stability of WHP was lower than that of egg albumin but greater than that of soy protein concentrate. Thermal stability of the FPP samples is in the following order: HGP > HBP > WHP > HHP > AFP. PMID- 15291473 TI - Concise preparation of the (3E,5Z)-alkadienyl system. New approach to the synthesis of principal insect sex pheromone constituents. AB - A new rapid and low-cost preparation of the (3E,5Z)-3,5-alkadienyl system, encountered in several insect pheromone constituents, was developed. Knoevenagel condensation of (E)-2-alkenals with ethyl hydrogen malonate in dimethyl sulfoxide, in the presence of a catalytic amount of piperidinium acetate, led to a mixture of geometrical isomers of ethyl 3,5-alkadienoates and ethyl 2,4 alkadienoates, from which the (3E,5Z)-3,5-alkadienoate was conveniently separated, by the use of urea inclusion complex formation. The importance of this procedure has been illustrated by the preparation of the (3E,5Z)-3,5 tetradecadienoic acid (megatomoic acid) 1, the (3E,5Z)-3,5-dodecadienyl acetate 2, and the (3E,5Z)-3,5-tetradecadienyl acetate 3. These compounds are the main components of insect sex pheromones and constitute synthetic targets of considerable interest for the semiochemical community. PMID- 15291474 TI - Extracellular production of a functional soy cystatin by Bacillus subtilis. AB - A recombinant Bacillus subtilis producing soy cystatin was developed by subcloning with a soy cystatin gene cloned in Escherichia coli. An active form of cystatin against the cysteine protease from Pacific whiting fillets contaminated with Myxosporidia parasite was constitutively expressed and secreted extracelluarly into the medium. Two gene fragments of signal peptides from kerA and sacB were introduced and compared for secretion efficiency of cystatin. The secretion level of active cystatin improved with the signal peptide of kerA when compared to that of sacB. Inhibitor activity was reduced rapidly after peak expression of the target protein at 36 h of fermentation. The addition of 1% glucose, a suppressor of protease, into the medium sustained the increase of the cystatin activity during fermentation. This study introduced a potential new method for fermentation production of cystatin. PMID- 15291475 TI - Enzymatic and fungal treatments on sugarcane bagasse for the production of mechanical pulps. AB - Crude ligninolytic enzyme extracts from Phanerochaete chrysosporium fungi were applied to sugarcane bagasse, prior to thermomechanical (TMP) and chemithermomechanical pulping (CTMP), and their properties were compared with the normal TMP and CTMP and also with TMP and CTMP pretreated with Ceriporiopsis subvermispora and P. chrysosporium fungi. The sugarcane bagasse was impregnated with the crude enzyme extract containing lignin peroxidase (LiP), manganese peroxidase (MnP), and laccase (Lac). The results show that pretreatment with enzyme crude extract is an advantageous way to produce TMP and CTMP from sugarcane bagasse, as compared with only fungal pretreatment. Enzymatic pretreatments need only hours to enhance pulping and paper properties, compared with the weeks necessary for fungal treatments. Higher pulp yields were obtained compared with the fungal pretreatments. Enzymatic pretreatment reduced the energy consumption in a proportion similar to that of C. subvermispora fungal pretreatment and increased the pulp tensile index compared with the normal TMP and CTMP pulps, although the tensile strength was somewhat lower than that for pulps from C. subvermispora fungal pretreatment before CTMP processing. An advantage of enzymatic pretreatment is that brightness is increased compared with normal TMP and CTMP processes, whereas fungal pretreatments reduce the brightness. PMID- 15291476 TI - Proteome-level differences between auxinic-herbicide-susceptible and -resistant wild mustard (Sinapis arvensis L.). AB - To identify proteins that may be involved in mediating auxinic herbicide resistance (i.e., resistance to dicamba, picloram, 2,4-D), we compared the proteomes of an auxinic-herbicide-susceptible (S) and -resistant (R) wild mustard (Sinapis arvensis L.) biotype at different developmental stages. Using two dimensional electrophoresis and mass spectrometry, we identified 11 seedling and leaf proteins that showed reproducible differences in expression between the S and the R wild mustard biotype following application of dicamba. Our proteome level studies revealed the increased expression of the enzyme peptidylprolyl cis trans isomerase (PPIase), which has recently been implicated in auxin signal transduction. Juglone, an inhibitor of PPIase, interfered with the normal ability of R seeds to germinate in the presence of dicamba, whereas S seeds did not germinate in the presence of dicamba or dicamba plus juglone. When R and S plants (3-4 leaf stage) were treated with dicamba, S showed typical auxinic herbicide effects (e.g., epinasty) whereas R did not. However, the concomitant application of dicamba and juglone to greenhouse-grown R plants produced morphological changes that were consistent with known auxinic-herbicide-induced symptoms. This is the first report suggesting the potential involvement of differential expression of PPIase in mediating auxinic herbicide resistance. PMID- 15291477 TI - Heat-induced gelation of pea legumin: comparison with soybean glycinin. AB - Gel network formation of pea legumin (8.4% on a protein basis, pH 7.6) was monitored via dynamic rheological measurements. Gelation was performed in the absence and presence of the thiol-blocking reagent N-ethylmaleimide, at different rates of heating and cooling. Overall, it was shown that pea legumin gel formation was not effected by changes in the heating rate, and the two differently heated samples were unaffected by the addition of 20 mM NEM, which indicated that disulfide bonds were not essential within the network strands of these legumin gels. However, slowly cooling the legumin samples caused disulfide bonds to become involved within the network; this was observed by a large increase in gel strength that was then substantially reduced when repeating the sample in the presence of NEM. These experiments were repeated with soybean glycinin in order to determine whether a common model for gel formation of legumin-like proteins could be built, based upon molecular reasoning. The two proteins were affected in the same way by changes in the conditions used, but when applying a procedure of reheating and recooling the gel networks responded differently. Pea legumin gel networks were susceptible to rearrangements that caused the gels to become stronger after reheating/recooling, yet glycinin gel networks were not. It was concluded that the same physical and chemical forces drove the processes of denaturation, aggregation, and network formation. Each process can therefore be readily targeted for modification based upon molecular reasoning. Pea legumin and soybean glycinin gel networks had structurally different building blocks, however. A model of gelation aimed at texture control therefore requires additional information. PMID- 15291478 TI - Optimization of production of conjugated linoleic acid from soybean oil. AB - Linoleic acid from soybean oil was used to synthesize conjugated linoleic acid (CLA), and the response surface methodology (RSM) was applied to optimize the process. A temperature of -35 degrees C and a solvent to oil sample ratio of 8 were suggested for removal of saturated fatty acids by low-temperature crystallization. The ratio of oil sample/urea/methanol suggested was 1:2:5.5 (w/w/v) for removal of oleic acid by urea crystallization. A temperature of 150 degrees C and a time of 140 min were found to be the optimal conditions in the isomerization for the production of c-9,t-11 and t-10,c-12 CLA isomers. PMID- 15291479 TI - Decreased levels of tobacco-specific N-nitrosamines in moist snuff on the Swedish market. AB - Moist snuff, or snus, on the Swedish market in 2001 and 2002 was analyzed for tobacco-specific N-nitrosamines (TSNAs) using a recently developed LC-MS/MS method. All samples of moist snuff analyzed were found to contain detectable levels of N'-nitrosonornicotine (NNN), N'-nitrosoanatabine (NAT), N' nitrosoanabasine (NAB), and 4-(N-methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK). In the survey in 2001, all samples except for one were produced by Swedish Match (n = 14), which is the dominating manufacturer on the Swedish snuff market. In the survey in 2002, samples from both Swedish Match (n = 7) and seven smaller manufacturers (n = 20) were analyzed. Total TSNA levels of between 0.15 and 3.0 microg/g wet weight were found. In the survey in 2001 and 2002, the mean level of the total TSNA content in moist snuff was 1.1 microg/g (n = 14) and 1.0 microg/g (n = 27), respectively. The result of the survey shows that the level of TSNAs in moist snuff on the Swedish market has been greatly reduced since the middle of the 1980s. Clearly, efforts have been made by the manufacturers to reduce the level of TSNAs in snuff. PMID- 15291480 TI - Allium discoloration: precursors involved in onion pinking and garlic greening. AB - Precursors involved in the formation of pink and green-blue pigments generated during onion and garlic processing, respectively, have been studied. It has been confirmed that the formations of both pigments are of very similar natures, with (E)-S-(1-propenyl)cysteine sulfoxide (isoalliin) serving as the primary precursor. Upon disruption of the tissue, isoalliin and other S-alk(en)ylcysteine sulfoxides are enzymatically cleaved, yielding 1-propenyl-containing thiosulfinates [CH3CH=CHS(O)SR; R = methyl, allyl, propyl, 1-propenyl] among others. The latter compounds have been shown to subsequently react with amino acids to produce the pigments. Whereas the propyl, 1-propenyl, and methyl derivatives form pink, pink-red, and magenta compounds, those containing the allyl group give rise to blue products after reacting with glycine at pH 5.0. The role of other thiosulfinates [RS(O)SR'] (R, R' = methyl, allyl, propyl) and (Z) thiopropanal S-oxide (the onion lachrymatory principle) in the formation of the pigments is also discussed. PMID- 15291481 TI - Isolation and identification of novel pyranoanthocyanins from black carrot (Daucus carota L.) juice. AB - Six novel pyranoanthocyanins were identified by HPLC-ESI-MSn in black carrot (Daucus carota L. ssp. sativus var. atrorubens Alef.) juice. The two major compounds, namely, the vinylcatechol adducts of cyanidin 3-O-(6-O-feruloyl-beta-D glucopyranosyl)-(1-->6)-[beta-D-xylopyranosyl-(1-->2)]-beta-D-galactopyranoside and cyanidin 3-O-[beta-D-xylopyranosyl-(1-->2)]-beta-D-galactopyranoside, respectively, were isolated by a combination of high-speed countercurrent chromatography with semipreparative HPLC. Their structures were fully elucidated by means of one- and two-dimensional NMR spectroscopy and high-resolution mass spectrometry. The four remaining pigments were characterized as the vinylphenol and vinylguaiacol adducts of cyanidin 3-O-[beta-D-xylopyranosyl-(1-->2)]-beta-D galactopyranoside, the vinylguaiacol adduct of cyanidin 3-O-(6-O-feruloyl-beta-D glucopyranosyl)-(1-->6)-[beta-D-xylopyranosyl-(1-->2)]-beta-D-galactopyranoside, and the vinylcatechol adduct of cyanidin 3-O-(6-O-sinapoyl-beta-d-glucopyranosyl) (1-->6)-[beta-D-xylopyranosyl-(1-->2)]-beta-D-galactopyranoside. These compounds are formed during storage of the juice through the direct reaction of either caffeic, ferulic, or coumaric acid with the respective genuine anthocyanins. PMID- 15291482 TI - Model studies on the formation of monochloropropanediols in the presence of lipase. AB - The formation of chloropropanols was investigated using model systems comprised of lipase, vegetable oil or fat, water, and sodium chloride. The results showed that measurable levels of the foodborne carcinogen 3-chloro-1,2-propanediol (3 MCPD) are formed in the presence of commercially available lipases of mammalian, vegetable, and fungal origins, incubated at temperatures of 40 degrees C. The highest yield of 3-MCPD was obtained in reaction mixtures containing lipase from Rhizopus oryzae, and all the lipases studied exhibited a high hydrolytic activity toward triglycerides from palm and peanut oil. In contrast, hydrolysis over time and the yield of 3-MCPD in olive and sunflower oils were significantly lower (up to 10-fold), possibly linked to the relatively lower amount (<18%) of saturated fatty acids in these oils. We provide here for the first time evidence that lipases are able to induce the formation of chloropropanols under model system conditions. However, the key intermediates and precise mechanistic aspects governing the formation of 3-MCPD in the presence of lipase still need to be elucidated. PMID- 15291483 TI - Lettuce and chicory byproducts as a source of antioxidant phenolic extracts. AB - A process to obtain enriched antioxidant phenolic extracts from lettuce (baby, romaine, and iceberg cultivars) and chichory byproducts as a way to valorize these byproducts was developed. Two extraction protocols using water and methanol as solvent were used. Amberlite XAD-2 nonionic polymeric resin was used to purify the extracts. The extraction yield, phenolic content, and phenolic yield were evaluated as well as the antioxidant capacity of the extracts (DPPH, ABTS, and FRAP assays). Baby and romaine lettuce byproducts showed the highest water extract yields [27 and 26 g of freeze-dried extracts/kg of byproduct fresh weight (fw), respectively], whereas baby and iceberg lettuce showed highest methanol extract yields (31 and 23 g of freeze-dried extracts/kg of byproduct fw, respectively). Methanol extraction yielded a raw extract with a high phenolic content, the baby and chicory extracts being the richest with approximately 50 mg of phenolics/g of freeze-dried extract. Regarding the purified extracts, water extraction yielded a higher phenolic content, baby and chicory being also the highest with mean values of approximately 190 and 300 mg of phenolics/g of freeze dried extract, respectively. Both raw and purified extracts from baby and chicory showed the higher antioxidant contents (DPPH, ABTS, and FRAP assays). The antioxidant capacity was linearly correlated with the phenolic content. The results obtained indicate that lettuce byproducts could be, from the industrial point of view, an interesting and cheap source of antioxidant phenolic extracts to funcionalize foodstuffs. PMID- 15291484 TI - Effect of the maceration technique on the relationships between anthocyanin composition and objective color of Syrah wines. AB - The effects of two different vinification techniques, traditional fermentation and carbonic maceration, on the anthocyanin composition and color of young red wines, made with Syrah grapes grown in a warm climate, were compared. Tristimulus colorimetry was applied to study the color of wines during the vinification, and a high-performance liquid chromatography (HPLC) procedure was used for the analysis of anthocyanins. Carbonic maceration led to wines with lower anthocyanin content, mainly monoglucosides, and total phenols. This was related to lighter wines, less saturated, but more colorful (higher chroma C*ab values), and hues hab similar to those of the Syrah wines made by traditional vinification. Thus, the lightness L* had much more influence on the saturation s*uv of the wines obtained by carbonic maceration than the chroma (s*uv = C*uv/L*). From a study of the color-composition relationships using linear and multiple regression, better relationships were found for the wines from traditional vinification, where the chromatic parameters L*, hab, and s*uv could be predicted from the 3 monoglucosides of delphinidin, petunidin, peonidin, and malvidin concentrations (R > 0.9). However, a good prediction of the chroma C*ab from the anthocyanin composition was not possible. On the contrary, C*ab was the best predicted parameter from the anthocyanins monoglucosides (R > 0.9) in the carbonic maceration wines. PMID- 15291485 TI - Identification of trace volatile compounds in freshly distilled Calvados and Cognac using preparative separations coupled with gas chromatography-mass spectrometry. AB - Gas chromatography coupled with mass spectrometry (GC-MS) using both electron impact and chemical ionization detection modes led to the determination of the volatile composition of two samples of freshly distilled Cognac and two samples of freshly distilled Calvados. A total of 169 volatile compounds were directly identified in dichloromethane extracts obtained by liquid-liquid extraction. Trace compounds present in both spirits were characterized with the help of preparative separations. In a first step, groups of compounds were separated by preparative GC, and the fractions were analyzed on a polar stationary phase by GC MS. In a second step, silica gel fractionation was used to separate them by polarity. In this study, 331 compounds, of which 162 can be considered as trace compounds, were characterized in both freshly distilled Cognac and Calvados. Of these, 39 are common to both spirits; 30 are specific to Cognac with numerous hexenyl esters and norisoprenoidic derivatives, whereas 93 are specific to Calvados with compounds such as unsaturated alcohols, phenolic derivatives, and unsaturated aldehydes. PMID- 15291486 TI - Differentially enhanced insect resistance, at a cost, in Arabidopsis thaliana constitutively expressing a transcription factor of defensive metabolites. AB - A transgenic line of Arabidopsis thaliana constitutively expressing a conserved MYB transcription factor of phenylpropanoid biosynthesis resulting in solid purple leaves had significantly increased resistance to leaf feeding by first instar fall armyworms (Spodoptera frugiperda), but no enhanced resistance to cabbage looper (Trichoplusia ni) larvae, when compared to wild type plants. However, inflorescence and silique (seed pod) production were significantly reduced by 22 and 52%, respectively, in the transgenic line compared to wild type plants. Reduction in feeding by S. frugiperda was significantly positively correlated with reduction in weights of survivors, but both were negatively correlated with the concentration of anthocyanins. These results indicate that a single gene regulator can activate a defensive pathway sufficient to produce increased resistance to insects but that this activation confers a cost in plant productivity. PMID- 15291487 TI - Aminomethylphosphonic acid, a metabolite of glyphosate, causes injury in glyphosate-treated, glyphosate-resistant soybean. AB - Glyphosate-resistant (GR) soybean [Glycine max (L.) Merr.] was developed by stable integration of a foreign gene that codes insensitive enzyme 5 enolpyruvylshikimate-3-phosphate synthase, an enzyme in the shikimate pathway, the target pathway of glyphosate. Application of glyphosate to GR soybean results in injury under certain conditions. It was hypothesized that if GR soybean is completely resistant to the glyphosate, injury could be caused by a metabolite of glyphosate, aminomethylphosphonic acid (AMPA), a known phytotoxin. Glyphosate and AMPA effects on one- to two-trifoliolate leaf stage (16-18-days old) GR and non GR soybean were examined in the greenhouse. In GR soybean, a single application of glyphosate-isopropylammonium (1.12-13.44 kg/ha) with 0.5% Tween 20 did not significantly reduce the chlorophyll content of the second trifoliolate leaf at 7 days after treatment (DAT) or the shoot dry weight at 14 DAT compared with Tween 20 alone. A single application of AMPA (0.12-8.0 kg/ha) with 0.5% Tween 20 reduced the chlorophyll content of the second trifoliolate leaf by 0-52% at 4 DAT and reduced shoot fresh weight by 0-42% at 14 DAT in both GR and non-GR soybeans compared with Tween 20 alone. AMPA at 0.12 and 0.50 kg/ha produced injury in GR and non-GR soybean, respectively, similar to that caused by glyphosate isopropylammonium at 13.44 kg/ha in GR soybean. AMPA levels found in AMPA-treated soybean of both types and in glyphosate-treated GR soybean correlated similarly with phytotoxicity. These results suggest that soybean injury to GR soybean from glyphosate is due to AMPA formed from glyphosate degradation. PMID- 15291488 TI - Use of resistant ACCase mutants to screen for novel inhibitors against resistant and susceptible forms of ACCase from grass weeds. AB - The aryloxyphenoxypropionic acid (AOPP) and cyclohexanedione (CHD) herbicides inhibit the first committed enzyme in fatty acid biosynthesis, acetyl CoA carboxylase (ACCase). The frequent use of AOPP and CHD herbicides has resulted in the development of resistance to these herbicides in many grass weed species. New herbicides that inhibit both the susceptible and resistant forms of ACCase in grass weeds would have obvious commercial appeal. In the present study, an attempt was made to identify molecules that target both the herbicide-sensitive and -resistant forms of ACCase. Seven experimental compounds, either CHD-like or AOPP-CHD hybrids, were synthesized and assayed against previously characterized susceptible and resistant forms of ACCase. All seven compounds inhibited ACCase from sensitive biotypes of Setaria viridis and Eleusine indica (I50 values from 6.4 to >100 microM) but were not particularly potent compared to some commercialized herbicides (I50 values of 0.08-5.6 microM). In almost all cases, the I50 values for each compound assayed against the resistant ACCases were higher than those against the corresponding sensitive ACCase, indicating reduced binding to the resistant ACCases. One compound, a CHD analogue, was almost equally effective against the resistant and susceptible ACCases, although it was not a very potent ACCase inhibitor per se (I50 of 51 and 76 microM against susceptible ACCase from S. viridis and E. indica, respectively). The AOPP-CHD hybrid molecules also inhibited some of the resistant ACCases, with I50 values ranging from 6.4 to 50 microM. These compounds may be good leads for developing ACCase inhibitors that target a wider range of ACCase isoforms, including those found in AOPP- and CHD-resistant weed biotypes. PMID- 15291489 TI - Olive oil mill wastewater treatment using a chemical and biological approach. AB - Olive oil mill wastewaters (OMW) are recalcitrant to biodegradation for their toxicity due to high values of chemical oxygen demand (COD), biological oxygen demand (BOD), and phenolic compounds. In the present study OMW, collected in southern Italy, were subjected first to a chemical oxidative procedure with FeCl3 and then to a biological treatment. The latter was performed in a pilot plant where mixed commercial selected bacteria, suitable for polyphenols and lipid degradation, were inoculated. The effect of treatments was assessed through COD removal, reduction of total phenols, and decrease of toxicity using primary consumers of the aquatic food chain (the rotifer Brachionus calyciflorus and the crustacean Daphnia magna). The results showed that the chemical oxidation was efficacious in reducing all parameters analyzed. A further decrease was found by combining chemical and biological treatments. PMID- 15291490 TI - Aroma extract dilution analysis of cv. Meeker (Rubus idaeus L.) red raspberries from Oregon and Washington. AB - The aromas of cultivar Meeker red raspberry from Oregon and Washington were analyzed by aroma extract dilution analysis. Seventy-five aromas were identified [some tentatively (superscript T)] by mass spectrometry and gas chromatography retention index; 53 were common to both, and 22 have not been previously reported in red raspberry. Twenty-one compounds had an equivalent odor impact in both: 2,5 dimethyl-4-hydroxy-3-(2H)-furanone, hexanal, 4-(2,6,6-trimethyl-1-cyclohexen-1 yl)-3-buten-2-one, (E)-beta-3,7-dimethyl-1,3,6-octatrieneT, 6,6-dimethyl-2 methylenebicyclo[3.1.1]heptaneT, 1-(2,6,6-trimethyl-1,3-cyclohexadien-1-yl)-2 buten-1-one, ethanoic acid, (Z)-3-hexenalT, 3-methylmercaptopropionaldehyde, (Z) 3-hexenol, 2,6-dimethyl-2,7-octadien-6-ol, butanoic acid, ethyl 2 methylpropanoate, (E)-2-hexenal, hexyl formateT, 2,3-butanedione, heptanalT, thiacyclopentadieneT, cyclohexane carbaldehydeT, (E)-3,7-dimethyl-2,6-octadien-1 olT, and 4-(p-hydroxyphenyl)-2-butanone. Oregon Meeker had 14 odorants with higher flavor dilution (FD) factors than Washington Meeker: 4-(2,6,6-trimethyl-2 cyclohexen-1-yl)-3-buten-2-oneT, 1-octanol, 5-isopropyl-2-methylcyclohexa-1,3 dieneT, 7-methyl-3-methylene-1,6-octadieneT, ethyl hexanoate, 3-methylbutyl acetateT, ethyl propanoate, 4-(4-hydroxy-3-methoxyphenyl)-2-butanoneT, 2 methylbutanoic acid, 1-octen-3-ol, ethyl cyclohexane carboxylateT, 2 methylthiacyclopentadieneT, (Z)-3-hexenyl acetateT, and 4-(2,6,6-trimethyl-2 cyclohexen-1-yl)-3-buten-2-olT. Washington Meeker had 16 odorants with higher FD factors than Oregon Meeker: 5-ethyl-3-hydroxy-4-methyl-2-(5H)-furanoneT, dimethyl sulfideT, 2-ethyl-4-hydroxy-5-methyl-3-(2H)-furanoneT, 1-hexanolT, ethyl 2 methylbutanoate, 3,7-dimethyl-1,6-octadien-3-yl acetateT, methyl hexanoate, phenyl ethanoic acidT, neo-allo-3,7-dimethyl-1,3,6-octatrieneT, 2-nonanoneT, 2-(4 methylcyclohex-3-enyl)propan-2-olT, phenylmethanolT, 5-octanolideT, 2 phenylethanol, 1-isopropyl-4-methylenebicyclo[3.1.0]hexaneT, and 2-undecanone. PMID- 15291491 TI - Combination of supercritical CO2 and vacuum distillation for the fractionation of bergamot oil. AB - Supercritical CO2 can be used to separate oxygenated compounds from essential oils. This technique still cannot replace vacuum distillation as an industrial process because of low recoveries and inconsistent results. In the present work, a comparison between the two methods was made in terms of composition, recovery, and color. Vacuum distillation and supercritical CO2 are complementary processes for producing high quality oxygenated compounds with high recovery rates. The former is more suitable for removing monoterpenes at low fraction temperatures (< or =308 K), and the latter is more suitable for separating oxygenated compounds from pigments and waxes. Consequently, the two methods were combined. For supercritical CO2 fractionation, the parameters of pressure, temperature gradient, and the ratio of solvent to feed were investigated for the fractionation of oxygenated compounds with high recoveries (> or =85%) and without other macromolecules, such as pigments and waxes. PMID- 15291492 TI - Analysis of volatiles in porcine liver pates with added sage and rosemary essential oils by using SPME-GC-MS. AB - The effect of the addition of two natural antioxidant extracts (sage and rosemary essential oils) and one synthetic (BHT) on the generation of volatile compounds in liver pates from Iberian and white pigs was analyzed using SPME-GC-MS. Lipid derived volatiles such as aldehydes [hexanal, octanal, nonanal, hept-(Z)-4-enal, oct-(E)-2-enal, non-(Z)-2-enal, dec-(E)-2-enal, deca-(E,Z)-2,4-dienal] and alcohols (pentan-1-ol, hexan-1-ol, oct-1-en-3-ol) were the most abundant compounds in the headspace of porcine liver pates. Pates from different pig breeds presented different volatiles profiles due to their different oxidation susceptibilities as a probable result of their fatty acid profiles and vitamin E content. Regardless of the origin of the pates, the addition of BHT successfully reduced the amount of volatiles derived from PUFA oxidation. Added essential oils showed a different effect on the generation of volatiles whether they were added in pates from Iberian or white pigs because they inhibited lipid oxidation in the former and enhanced oxidative instability in the latter. SPME successfully allowed the isolation and analysis of 41 volatile terpenes from pates with added sage and rosemary essential oils including alpha-pinene, beta-myrcene, 1 limonene, (E)-caryophyllene, linalool, camphor, and 1,8-cineole, which might contribute to the aroma characteristics of liver pates. PMID- 15291493 TI - Optimization of extraction of apple aroma by dynamic headspace and influence of saliva on extraction of volatiles. AB - The dynamic headspace procedure of aroma extraction was optimized on Gala apples (Malus domestica). Two parameters affecting the extractability of compounds were studied: temperature and purge time. The influence of artificial saliva was also included. An increase in purge time and temperature caused an increase in the extraction of volatiles from the apple matrix. The optimum point of extraction was 40 degrees C and 70 min of purge. The study also showed that the addition of saliva influenced the extraction of volatile compounds, but this effect was different from one compound to another. To verify that the headspace extracts presented a global odor representativeness of fresh apple under these conditions of extraction, eight assessors compared the odor of extracts with fresh fruit odor for three different cultivars. With regard to the sensory profiles of extracts, the optimal conditions of extraction were suitable for extraction of volatile compounds, even if cooked apple odor appeared in some extracts. The similarity marks of extracts were low but acceptable. PMID- 15291494 TI - Total phenolics and antioxidant activities of fenugreek, green tea, black tea, grape seed, ginger, rosemary, gotu kola, and ginkgo extracts, vitamin E, and tert butylhydroquinone. AB - The total phenolics and antioxidant activities of fenugreek, green tea, black tea, grape seed, ginger, rosemary, gotu kola, and ginkgo extracts, vitamin E, and tert-butylhydroquinone, were determined. Grape seed and green tea were analyzed for their phenolic constituents using high-performance liquid chromatography. The total phenolics of the plant extracts, determined by the Folin-Ciocalteu method, ranged from 24.8 to 92.5 mg of chlorogenic acid equivalent/g dry material. The antioxidant activities of methanolic extracts determined by conjugated diene measurement of methyl linoleate were 3.4-86.3%. The antioxidant activity of the extracts using chicken fat by an oxidative stability instrument (4.6-10.2 h of induction time) followed a similar trend in antioxidant activity as determined by the Folin-Ciocalteu method. Seven phenolics in grape seed and green tea extracts were identified that ranged from 15.38 to 1158.49 and 18.3 to 1087.02 mg/100 g of extract, respectively. Plant extracts such as green tea and grape seed extracts can be used to retard lipid oxidation in a variety of food products. PMID- 15291495 TI - Selected nutrient contents, fatty acid composition, including conjugated linoleic acid, and retention values in separable lean from lamb rib loins as affected by external fat and cooking method. AB - Proximate composition and fatty acid profile, conjugated linoleic acid (CLA) isomers included, were determined in separable lean of raw and cooked lamb rib loins. The cooking methods compared, which were also investigated for cooking yields and true nutrient retention values, were dry heating of fat-on cuts and moist heating of fat-off cuts; the latter method was tested as a sort of dietetic approach against the more traditional former type. With significantly (P < 0.05) lower cooking losses, dry heating of fat-on rib-loins produced slightly (although only rarely significantly) higher retention values for all of the nutrients considered, including CLA isomers. On the basis of the retention values obtained, both techniques led to a minimum migration of lipids into the separable lean, which was higher (P < 0.05) in dry heating than in moist heating, and was characterized by the prevalence of saturated and monounsaturated fatty acids. On the whole, the response to cooking of the class of CLA isomers (including that of the nutritionally most important isomer cis-9,trans-11) was more similar to that of the monounsaturated than the polyunsaturated fatty acids. PMID- 15291496 TI - Antioxidant phenols in barley (Hordeum vulgare L.) flour: comparative spectrophotometric study among extraction methods of free and bound phenolic compounds. AB - Phenolic compounds are found in both free and bound forms in cereals. The majority is in the insoluble bound form, that is, bound to cell wall material, such as ferulic acid and its derivatives. The antioxidant properties of the phenolic compounds in grains are associated with the health benefits of grains and grain products. The extraction capacity of several solvent mixtures, for extracting free phenols from barley flours, and the possibility of employing a rapid automated solvent extraction method were studied. The extraction yield of each method was evaluated by correlating several spectrophotometric indices (absorption at 280, 320, and 370 nm and total phenolic compounds by the Folin Ciocalteu method) with the antioxidant activities of the barley extracts (scavenging activity by the 2,2-diphenyl-1-picrylhydrazyl method). Interesting results were obtained when ethanol and acetone-based extraction mixtures were employed to extract free phenols. A comparison was made between alkaline and acid hydrolysis. The extraction yield of bound phenolic compounds increased when the digestion time for alkaline hydrolysis was prolonged. PMID- 15291497 TI - Synthesis of antioxidants in wheat sprouts. AB - Aqueous and ethanolic extracts from wheat (Triticum aestivum) sprout powder were analyzed to determine its reduction and antioxidant activities. Mean and standard deviation of five independent samples were reported. The results showed that the micromoles of potassium ferricyanide reduced by aqueous and ethanolic extracts corresponding to 1 g of sprout powder (80.6 +/- 11.2 and 9.7 +/- 1.8, respectively) were higher than that reduced by 1 mg of other reducing compounds: ascorbic acid, rutin, quercetin, and reduced gluthatione (4.8 +/- 0.7, 3.8 +/- 1.2, 4.8 +/- 1.7, and 1.6 +/- 0.4, respectively). In addition, the oxygen superoxide scavenging activity performed by sprout extracts (from 1 g of powder) is comparable to that shown by 10 mg of antioxidant pure compounds (rutin and quercetin). Biochemical analysis of the sprout extracts shows that the antioxidant activity is mainly due to reducing glycoside and polyphenolic compounds. PMID- 15291498 TI - Comparison of volatile aldehydes present in the cooking fumes of extra virgin olive, olive, and canola oils. AB - Emissions of low molecular weight aldehydes (LMWAs) from deep-frying of extra virgin olive oil, olive oil, and canola oil (control) were investigated at two temperatures, 180 and 240 degrees C, for 15 and 7 h, respectively. The oil fumes were collected in Tedlar bags and then analyzed by gas chromatography-mass spectrometry. Seven alkanals (C2-C7 and C9), eight 2-alkenals (C3-C10), and 2,4 heptadienal were found in the fumes of all three cooking oils. The generation rates of these aldehydes were found to be dependent on heating temperature, showing significant increases with increases in temperature. The LMWA emissions from both kinds of olive oils were very similar and were lower than those observed from canola oil under similar conditions. These results suggest that frying in any type of olive oil, independent of its commercial category, will effectively decrease the generation of volatile aldehydes in the exhaust. This fact is important because less expensive refined olive oil is usually used for deep-frying operations, whereas extra virgin olive oil is usually used as salad dressing. PMID- 15291499 TI - Fresh israeli jaffa sweetie juice consumption improves lipid metabolism and increases antioxidant capacity in hypercholesterolemic patients suffering from coronary artery disease: studies in vitro and in humans and positive changes in albumin and fibrinogen fractions. AB - The contents of the bioactive compounds in the pummelo-grapefruit hybrid juice named Israeli Jaffa Sweetie and their influence on humans suffering from hypercholesterolemia were studied. It was found that Sweetie juice has a high content of bioactive compounds and a high antioxidant potential. Then 72 hypercholesterolemic patients, ages 43-71 years, after coronary bypass surgery recruited from the Institute pool of volunteers, were randomly divided into two experimental (EG1 and EG2) groups and one control (CG) group, each comprising 24 patients. The diets of EG1 and EG2 patients were daily supplemented with 100 or 200 mL of fresh Sweetie juice, respectively. Before and after diet consumption serum lipid levels, albumin and fibrinogen fractions, and their antioxidant capacities were determined. After 30 consecutive days of Sweetie juice supplemented diets, improvements in serum lipids levels were found in EG1 and EG2 versus CG: (a) total cholesterol, 7.34 versus 8.02 mmol/L, -9.5%, and 6.73 versus 8.02 mmol/L, -16.1%, respectively; (b) low-density lipoprotein cholesterol, 5.63 versus 6.37 mmol/L, -11.6%, and 5.03 versus 6.37 mmol/L, -21.0%, respectively; (c) total glycerides, 2.01 versus 2.27 mmol/L, -11.5%, and 1.71 versus 2.27 mmol/L, -24.7%, respectively. Serum albumin concentration was increased but not significantly in EG1 and EG2 versus CG: 47.5 versus 44.5 g/L, +6.7%, and 47.9 versus 44.5 g/L, +7.6%, respectively. A significant increase in the serum, albumin, and fibrinogen antioxidant capacities in EG2 and to a lesser degree in EG1 was observed. No changes in the CG were found. In conclusion, fresh Sweetie juice contains high quantities of bioactive compounds and has a high antioxidant potential. Diet supplemented with this juice positively influences serum lipid, albumin, and fibrinogen levels and their antioxidant capacities. Addition of fresh Sweetie juice to generally accepted diets may be beneficial for hypercholesterolemic patients. PMID- 15291500 TI - Detection and quantification of roundup ready soy in foods by conventional and real-time polymerase chain reaction. AB - Transgenic soybean line GTS-40-3-2, marketed under the trade name Roundup Ready (RR) soy, was developed by Monsanto (USA) to allow for the use of glyphosate, the active ingredient of the herbicide Roundup, as a weed control agent. RR soy was first approved in Canada for environmental release and for feed products in 1995 and later for food products in 1996 and is widely grown in Canada. Consumer concern issues have resulted in proposed labeling regulations in Canada for foods derived from genetically engineered crops. One requirement for labeling is the ability to detect and accurately quantify the amount of transgenic material present in foods. Two assays were evaluated. A conventional qualitative Polymerase Chain Reaction (PCR) assay to detect the presence of soy and RR soy and a real-time PCR to quantify the amount of RR soy present in samples that tested positive in the first assay. PCR controls consisted of certified RR soy reference material, single transgenic soybeans, and a processed food sample containing a known amount of RR soy. To test real-world applicability, a number of common grocery store food items that contain soy-based products were tested. For some samples, significant differences in amplification efficiencies during the quantitative PCR assays were observed compared to the controls, resulting in potentially large errors in quantification. A correction factor was used to try to compensate for these differences. PMID- 15291501 TI - Evaluation of vinification lees as a general medium for lactobacillus strains. AB - Lactobacillus species present high nutritional requirements, so it is necessary to find new low-cost nutrient components for fermentation media. This work compares the utilization of vinification lees (an important residue of wineries) from red and white winemaking technology, distilled or not. An amount of 20 g of lees/L was used as the unique nutrient to obtain lactic acid from glucose using Lactobacillus strains with different properties: L. plantarum CECT-221, L. pentosus CECT-4023, L. casei CECT-5275, and L. coryniformis subsp. torquens CECT 25600. Only L. casei using distilled lees showed values (Pmax = 92.1 g/L and Y(P/S) = 1.05 g/g) similar to those obtained with the MRS broth. The UV spectra of white and red lees, distilled or not, allowed an interpretation of the different phenolic compounds present and their influence on the fermentation. Their detoxification by extraction with organic compounds and fermentation with L. pentosus was also considered. Time courses of glucose and lactic acid were modeled according to reported models to obtain more information about the process. PMID- 15291502 TI - Aging of whiskey increases 1,1-diphenyl-2-picrylhydrazyl radical scavenging activity. AB - 1,1-Diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity of Japanese whiskey after various aging periods in oak barrels was measured to evaluate the antioxidative effects of whiskey. The activity of the whiskey increased with the aging period with high correlation. The activity of various types of whiskey was measured and shown to be correlated to the potentiation of the GABAA receptor response measured in a previous paper. However, the fragrant compounds in the whiskey which potentiated the GABAA receptor response had low DPPH radical scavenging activity, while phenol derivatives had high radical scavenging activity. The whiskey was extracted by pentane. The aqueous part showed the scavenging activity, whereas the pentane part did not. Thus, both the DPPH radical scavenging activity and the potentiation of the GABAA receptor response increased during whiskey aging in oak barrels, but were due to different components. The whiskey protected the H2O2-induced death of E. coli more than ethanol at the same concentration as that of the whiskey. The changes that occurred in the whiskey during aging may be the reason aged whiskies are so highly valued. PMID- 15291503 TI - Oleosins of Arabidopsis thaliana: expression in Escherichia coli, purification, and functional properties. AB - The interfacial behavior of oleosins, the most abundant proteins from seeds oil bodies, was investigated using the pendant drop method at water/oil interfaces and compared to the behavior of beta-casein and lysozyme, proteins with contrasted emulsifying properties. Recombined high (rS3) and low (rS4) molecular weight oleosins comprising N-terminal histidine tags were purified to electrophoretic homogeneity. rS3 decreased the interfacial tension at the oil/water interface better than rS4, oleosins being more efficient than beta casein. Oleosins formed aggregates when spread on noncompressed phospholipid (PL) films at the air/water interface as observed using a Langmuir-Blodgett balance equipped with a Brewster angle microscope. Oleosin spread at the surface of a compressed PL monolayer (5-20 mN/m) did not aggregate. Pressure increased immediately and proportionally to the amount of protein spread on the monolayer. The results stress the capacity of oleosins to be inserted in oil and in PL monolayers, which is of particular relevancy to their potential uses as water/oil emulsifiers. PMID- 15291504 TI - Nonenzymatic browning kinetics of a carbohydrate-based low-moisture food system at temperatures applicable to spray drying. AB - Effects of water contents on nonenzymatic browning (NEB) rates of amorphous, carbohydrate-based food model systems containing L-lysine and D-xylose as reactants were studied at different temperatures (40, 50, 60, 70, 80, and 90 degrees C) applicable to spray drying conditions. Water sorption was determined gravimetrically, and data were modeled using the Brunauer-Emmett-Teller and Guggenheim-Anderson-deBoer equations. Glass transition, Tg was measured by DSC. NEB was followed spectrophotometrically. The rate of browning increased with water content and temperature, but a lower T-Tg was needed for browning at decreasing water content. Water content seemed to affect the activation energy of NEB, and higher water contents decreased the temperature dependence of the NEB. At higher temperatures, the NEB became less water content dependent and enhanced browning in spray-drying. The temperature dependence of nonenzymatic browning could also be modeled using the Williams-Landel-Ferry (WLF) equation, but the WLF constants were dependent on the water content. PMID- 15291505 TI - Vicilin-like storage globulin from buckwheat (Fagopyrum esculentum Moench) seeds. AB - An 8S storage globulin from buckwheat seed, which resembles the structure and features common to the vicilin-like family of seed storage proteins, was analyzed for this paper. It was found that expression of the 8S globulin gene precedes that of the 13S globulin (the main buckwheat storage protein) and starts from an early stage of buckwheat seed development (9-11 days after flowering), continuing to accumulate throughout seed development to contribute approximately 7% of total seed proteins. This protein fraction might be more interesting for biotechnological application than the 13S buckwheat legumin consisting of 23-25 kDa subunits reported to be the major buckwheat allergen. A partial cDNA was also isolated, showing high homology with cDNAs coding for vicilin-like storage proteins from various plant species, and its expression profile throughout seed development as well as in different buckwheat tissues was analyzed. PMID- 15291506 TI - Assessment of the reactivity of selected isoflavones against proteins in comparison to quercetin. AB - Selected isoflavones (genistein, daidzein, formononetin, prunetin, biochanin A, and two synthetic isoflavones) were allowed to interact with soy and whey proteins. The reaction products were analyzed in terms of covalent binding at the nucleophilic side chains of proteins. Changes in molecular properties of the proteins derivatives were documented by SDS-PAGE, IEF, and SELDI-TOF-MS. The structural changes induced were studied using circular dichroism. The in vitro digestibility was assessed with trypsin. The results show that the occurrence of the catechol moiety, that is, the two adjacent (ortho) aromatic hydroxyl groups on ring B of the flavonoid structural skeleton appears to be prerequisite condition for covalent binding to proteins. The catechol moiety on ring A was less reactive. Its absence lead to a slight or no significant reaction, although noncovalent interactions may still be possible, even when lacking this structural element. A comparison of the data is also made with quercetin representing the flavonols. PMID- 15291507 TI - Antioxidant activity of a proanthocyanidin-rich extract from grape seed in whey protein isolate stabilized algae oil-in-water emulsions. AB - Algae oil-in-water emulsions stabilized with 0.2% whey protein isolate (WPI) at pH 3.0 and 7.0 were chosen to evaluate antioxidant activity of a proanthocyanidin rich extract from grape seed. In this emulsion system, (+)-catechin and ascorbic acid (620 microM) were found to be prooxidative at pH 3.0 and ineffective at pH 7.0. Grape seed extract was not able to effectively inhibit both lipid hydroperoxides and propanal formation when added to the emulsion at 124 microM. However, increasing the concentration of the grape seed extract to 620 microM resulted in inhibition of both lipid hydroperoxide and propanal formation at pH 3.0 and 7.0. None of the antioxidants tested had any effect on the physical stability of the WPI-stabilized emulsion. The superior antioxidant activity of the grape seed extract is likely due to the presence of oligomeric procyanidins which are better antioxidants compared to their monomeric counterparts. PMID- 15291508 TI - FT-Raman spectroscopy, fluorescent probe, and solvent accessibility study of egg and milk proteins. AB - Due to possible contribution of both electrostatic and hydrophobic interactions, use of anionic fluorescent probes such as 1-anilinonaphthalene-8-sulfonic acid (ANS) and cis-parinaric acid (CPA) for the measurement of protein surface hydrophobicity (S0) has been controversial. A neutral probe, 6-propionyl-2 (dimethylamino)-naphthalene (PRODAN), may circumvent this problem. To select the best indicator of S0, in this study, the data for nine model proteins in phosphate buffer, pH 7.5, measured using the above-mentioned probes, was compared to their FT-Raman spectra and calculated solvent accessibility values. Log S0 measured using CPA had the highest correlation (r = 0.874) with the intensities of Raman spectral signals at 760 cm(-1) and 2800-3100 cm(-1), which were combined using a mixture design based on the random-centroid optimization. The order of correlation of Raman spectral parameters with S0 values were CPA > PRODAN > ANS. FT-Raman spectroscopy, therefore, identified CPA, followed by PRODAN, as the fluorescent probe of choice for describing surface hydrophobicity. However, the amino acid surface accessibility calculated using the PredictProtein software was not useful in identifying the best fluorescent probe for the measurement of S0. PMID- 15291509 TI - Preventive effect of Thea sinensis melanin against acetaminophen-induced hepatic injury in mice. AB - The preventive effect of Thea sinensis melanin (TSM) against overdoses of N acetyl-p-aminophenol (NAPAP) was studied on ICR mice. Animals were given 400 mg/kg intraperitoneally (i.p.) of NAPAP, and TSM was injected i.p. in doses 10-40 mg/kg 2 h before intoxication. The protective effects were evidenced by a complete blockage of the NAPAP-induced elevation of plasma alanine aminotransferase (ALT) activity, decreased concentration of thiobarbituric acid reactive substances (TBARS) to the control level, and a partial prevention of reduced glutathione (GSH) depletion in the liver tissue. Preadministration of TSM also caused restoration of superoxide dismutase (SOD) activity and resumed content of coenzymes Q9 and Q10. TSM by itself, however, did not affect the hepatic functional parameters, including serum ALT, TBARS, GSH, SOD, or coenzymes Q in the liver. Administration of TSM caused a dose-dependent inhibition of N nitrosodimethylamine demethylase activity with ED50 of 15.8 mg/kg. Activities of ethoxyresorufin O-dealkylase and pentoxyresorufin O-alkylase isozymes were changed insignificantly. The immune suppressive effect of NAPAP on the in vivo antibody-forming cell responses was demonstrated using ICR-sensitized mice with sheep red blood cells. The joint effect of TSM and NAPAP indicated the capability of TSM to recover immunity of the animals to the level of intact mice. Results obtained demonstrate that TSM preadministration can prevent the multiple toxic effects of NAPAP. PMID- 15291510 TI - Effects of different cooking procedures on lipid quality and cholesterol oxidation of farmed salmon fish (Salmo salar). AB - Salmon fillets were steamed, or pan-fried without oil, with olive oil, with corn oil, or with partially hydrogenated plant oil. The exchange between the salmon and the pan-frying oils was marginal, but it was detectable as slight modifications in the fatty acid pattern and the tocopherol contents according to the oil used. Primary and secondary oxidation products were only slightly increased or remained unchanged, which indicated a slight lipid oxidation effect due to the heating procedures applied. The same was observed for tocopherol levels, which remained almost stable and were not affected by the oxidation process. The sum of cholesterol oxidation products (COPs) increased after the heating processes from 0.9 microg/g in the raw sample to 6.0, 4.0, 4.4, 3.3, and 9.9 microg/g extracted fat in pan-fried without oil, with olive oil, corn oil, partially hydrogenated plant oil, and steamed, respectively. A highly significant correlation was found between the fatty acid pattern and the total amount of COPs (r2 = 0.973, p < 0.001). No change has been determined in the n-3 fatty acids content and in the polyunsaturated/saturated-ratio of the cooked salmon fillets. Moderate pan-frying (6 min total) and steaming (12 min) of salmon did not accelerate lipid oxidation but significantly increased the content of COPs. The highest increase of COPs was found through steaming, mainly due to the longer heat exposure. The used frying oils did not influence the outcome; no significant difference between heat treatment with or without oil has been determined. PMID- 15291511 TI - Phenolics from commercialized grape extracts prevent early atherosclerotic lesions in hamsters by mechanisms other than antioxidant effect. AB - The aim of this study was to evaluate the antiatherosclerotic effect of commercially available phenolic-rich extracts from grape seeds (ExGrape seeds, EGS; grape seed extract, GSE) and marc (ExGrape total, EGT) in cholesterol-fed hamsters and to investigate possible operating mechanisms. These extracts fed at a moderate dose mimicking two glasses of red wine per meal reduced plasma cholesterol (-11% on average) but did not affect plasma antioxidant capacity of hamsters. The extracts prevented the development of aortic atherosclerosis by 68% (EGS), 63% (EGT), and 34% (GSE). Elsewhere, in an ex vivo experiment using rat aortic rings, EGS (7 microg/mL) induced 77% endothelium-dependent relaxation, whereas EGT and GSE (30 microg/mL) induced 84 and 72%, respectively. These results suggests that phenolic extracts from grape seeds and marc are beneficial in inhibiting atherosclerosis by indirect mechanism(s). PMID- 15291513 TI - Synthesis and properties of tadpole-shaped gold nanoparticles. AB - In this communication, gold nanoparticles with a tadpole shape were synthesized by a simple aqueous-phase chemical method. The unusual three-dimensional and crystallized structures were demonstrated by TEM, AFM, and HRTEM methods. The SEM and UV-visible absorption measurements and electrophoresis experiments revealed that the tadpoles had novel optical and electrical properties. These attractive structures and properties are expected to find uses in many fields such as electrics, optics, electrochemistry, and biological sensing. PMID- 15291512 TI - Helical beta-peptide inhibitors of the p53-hDM2 interaction. AB - hDM2 is recognized in vivo by a short alpha-helix within the p53 trans-activation domain (p53AD). Disruption of the p53.hDM2 interaction is an important goal for cancer therapy. A functional epitope comprised of three residues on one face of the p53AD helix (F19, W23, and L26) contributes heavily to the binding free energy. We hypothesized that the p53AD functional epitope would be recapitulated if the side chains of F19, W23, and L26 were presented at successive positions three residues apart on a stabilized beta3-peptide 14-helix. Here, we report a set of beta3-peptides that possess significant 14-helix structure in water; one recognizes a cleft on the surface of hDM2 with nanomolar affinity. The strategy for beta3-peptide design that we describe is general and may have advantages over one in which individual or multiple beta-amino acid substitutions are introduced into a functional alpha-peptide, because it is based on homology at the level of secondary structure, not primary sequence. PMID- 15291514 TI - Two types of intramolecular addition of an Al-N multiple-bonded monomer LAlNAr' arising from the reaction of LAl with N3Ar' (L = HC[(CMe)(NAr)]2, Ar' = 2,6 Ar2C6H3, Ar = 2,6-iPr2C6H3). AB - The reaction of beta-diketiminated aluminum(I) monomer LAl with a large bulky azide N3Ar' (L = HC(CMeNAr)2, Ar' = 2,6-Ar2C6H3, Ar = 2,6-iPr2C6H3) in the temperature range from -78 degrees C to room temperature affords two different isomers 2 and 3, which have been characterized by spectroscopic and X-ray structural analyses, as well as elemental analysis. The variable-temperature 1H NMR kinetic studies of this reaction indicate the existence of the monomer LAlNAr' (1) at low temperature and the thermal stability of the compounds increases in the order of 1 < 2 < 3. PMID- 15291515 TI - Highly conjugated (polypyridyl)metal-(porphinato)zinc(II) compounds: long-lived, high oscillator strength, excited-state absorbers having exceptional spectral coverage of the near-infrared. AB - Transient dynamical studies of ruthenium(II) [5-(4'-ethynyl-(2,2';6',2' ' terpyridinyl))-10,20-bis(2',6'-bis(3,3-dimethyl-1 butyloxy)phenyl)porphinato]zinc(II)-(2,2';6',2' '-terpyridine)2+ bis hexafluorophosphate (Ru-PZn), osmium(II) [5-(4'-ethynyl-(2,2';6',2' ' terpyridinyl))-10,20-bis(2',6'-bis(3,3-dimethyl-1 butyloxy)phenyl)porphinato]zinc(II)-(2,2';6',2' '-terpyridine)2+ bis hexafluorophosphate (Os-PZn), ruthenium(II) [5-(4'-ethynyl-(2,2';6',2' ' terpyridinyl))-15-(4'-nitrophenyl)ethynyl-10,20-bis(2',6'-bis(3,3-dimethyl-1 butyloxy)phenyl)porphinato]zinc(II)-(2,2';6',2' '-terpyridine)2+ bis hexafluorophosphate (Ru-PZn-A), osmium(II) [5-(4'-ethynyl-(2,2';6',2' ' terpyridinyl))-15-(4'-nitrophenyl)ethynyl-10,20-bis(2',6'-bis(3,3-dimethyl-1 butyloxy)phenyl)porphinato]zinc(II)-(2,2';6',2' '-terpyridine)2+ bis hexafluorophosphate (Os-PZn-A), and ruthenium(II) [5-(4'-ethynyl-(2,2';6',2' ' terpyridinyl))-ruthenium(II)-15-(4'-ethynyl-(2,2';6',2' '-terpyridinyl))-10,20 bis(2',6'-bis(3,3-dimethyl-1-butyloxy)phenyl)porphinato]zinc(II)-bis(2,2';6',2' ' terpyridine)4+ tetrakis-hexafluorophosphate (Ru-PZn-Ru), and ruthenium(II) [5-(4' ethynyl-(2,2';6',2' '-terpyridinyl))-osmium(II)-15-(4'-ethynyl-(2,2';6',2' ' terpyridinyl))-10,20-bis(2',6'-bis(3,3-dimethyl-1 butyloxy)phenyl)porphinato]zinc(II)-bis(2,2';6',2' '-terpyridine) tetrakis hexafluorophosphate (Ru-PZn-Os) show that these highly conjugated supermolecular chromophores feature electronically excited states that absorb over broad NIR spectral windows with considerable oscillator strength and manifest lifetimes (1 50 mus) that are extraordinarily long relative to those of classic low band-gap organic materials. The excited-state absorptive domains of these strongly coupled multipigment ensembles can be extensively modulated. For sequential one-photon absorptive processes, these compounds evince large sigmae, sigmae/sigmag, and sigmae - sigmag values. As the combination of all these properties within single chromophoric entities have heretofore lacked precedent within the NIR, these and closely related structures may find particular utility in a variety of technologically important optical-limiting applications. PMID- 15291516 TI - Rapid direct nanowriting of conductive polymer via electrochemical oxidative nanolithography. AB - Conductive polymer nanolines having widths as small as 45 nm were obtained on glass using a novel scanning probe lithographic (SPL) technique at writing speeds of >5 mum/s. Herein we demonstrate that our nanowriting is >1500 times faster than current SPL nanoscale writing of conductive polymers. The lack of a specific restriction on the choice of substrates and the ability to write within a polymer matrix to provide a continuous film that is either 2-D or 2.5-D provide tremendous potential for our SPL technique in nanotechnology and plastic electronics applications. PMID- 15291517 TI - Screening molecular associations with lipid membranes using natural abundance 13C cross-polarization magic-angle spinning NMR and principal component analysis. AB - We describe an NMR approach for detecting the interactions between phospholipid membranes and proteins, peptides, or small molecules. First, 1H-13C dipolar coupling profiles are obtained from hydrated lipid samples at natural isotope abundance using cross-polarization magic-angle spinning NMR methods. Principal component analysis of dipolar coupling profiles for synthetic lipid membranes in the presence of a range of biologically active additives reveals clusters that relate to different modes of interaction of the additives with the lipid bilayer. Finally, by representing profiles from multiple samples in the form of contour plots, it is possible to reveal statistically significant changes in dipolar couplings, which reflect perturbations in the lipid molecules at the membrane surface or within the hydrophobic interior. PMID- 15291518 TI - Carbon-nitrogen bond formation via the reaction of terminal alkynes with a dinuclear side-on dinitrogen complex. AB - The dinuclear dinitrogen complex ([P2N2]Zr)2(mu-eta2:eta2-N2) reacts with terminal aryl alkynes to generate a new species in which the dinitrogen unit has been functionalized. The products formed have the general formula ([P2N2]Zr)2(mu eta2:eta2-N2CCAr)(mu-CCAr) and display a styryl-hydrazido unit bridging the two Zr centers along with a bridging arylalkynide. The crystal structures of three of these products are reported. A mechanism is proposed for this process that involves cycloaddition of the alkyne to the side-on dinitrogen unit followed by protonation of the Zr-C bond by a second equivalent of terminal alkyne. A fluxional process is operative in solution that equilibrates the phosphorus nuclei at high temperature; in the slow exchange limit, the two [P2N2]Zr ends of complex are inequivalent as evidenced by four resonances in the 31P NMR spectrum for the inequivalent phosphorus donors. This C-N bond-forming reaction is unique in that an activated dinitrogen fragment undergoes a reaction with an alkyne. PMID- 15291519 TI - Synthesis and isolation of cobalt hexacyanoferrate/chromate metal coordination nanopolymers stabilized by alkylamino ligand with metal elemental control. AB - This Communication describes the novel isolation with metal elemental control of cobalt hexacyanoferrate/chromate metal coordination polymers, stabilized by stearylamine (Co-Fe/Cr-SA) as a protecting coordination ligand in a reverse micelle technique. Each Co-Fe/Cr-SA can be isolated with high uniformity of particle size and elemental composition, and the ratio of the metal component depends on the fundamental characteristics of Co-Fe/Cr-SA: the nanopolymer's shape, color, and magnetism. PMID- 15291520 TI - Designer ligands for beryllium. AB - We report the rational design of ligands that selectively bind beryllium. We selected two ligands to design Be based on binding polynulear species with a Be-O Be motif: 2-hydroxyisophthalic acid (HIPA) and 2,3-dihydroxybenzoic acid (DHBA). All previous work has focused on BeL or BeL2 species. The HIPA and DHBA have extremely high binding constants of 17.5 and 18.4, respectively. These ligands outcompete chromotropic acid, which has one the highest binding constants for Be reported in the literature for a simple BeL species. The binding of the second Be to form the Be-O-Be motif is so strong that polynuclear species predominates in solution down to micromolar concentrations. Both ligands show a fluorescence response in the presence of beryllium, making them promising candidates for fluorescence-based sensors. In the case of HIPA, there is a fluorescence shift, and in the case of DHBA, the presence of beryllium turns on the fluorescence by removing two OH bonds that otherwise lead to nonradiative decay. The most dramatic result is that DHBA selectively binds Be in the presence of a metal cocktail containing a 50-fold excess of Al, Fe, Cr, Cu, Zn, Cd, and Pb. This is the first time that such selectivity for beryllium has been demonstrated. PMID- 15291521 TI - Polythiophene containing thermally removable solubilizing groups enhances the interface and the performance of polymer-titania hybrid solar cells. AB - A polythiophene derivative containing thermally removable branched ester solubilizing groups has been prepared and tested as a processable organic semiconductor polymer with tunable electronic and chemical properties for hybrid polymer-inorganic solar cells. Thermal removal of the protecting group enhances the interface between the organic and inorganic components while also contributing to better light absorption, energy transfer, and overall cell performance. PMID- 15291522 TI - Intensely luminescent gold(I)-silver(I) cluster complexes with tunable structural features. AB - A new series of isostructural, brilliantly luminescent gold-silver complexes having the formula [Au3(mu3-E)Ag(PPh2py)3](BF4)2 where E = O, S, Se and Ph2Ppy = 2-diphenylphosphinopyridine has been synthesized and characterized. The structural core of these complexes is a Au3Ag metallophilically linked tetrahedron with a group-16 atom functioning as a mu3-ligand capping the three gold atoms. In the solid state, pairs of clusters are joined by two unsupported aurophilic interactions. The emission energy changes strikingly in going from O (blue) to S (yellow) and Se (orange). The luminescence from the E = O system is the first to be reported for a gold(I) oxo system. Additionally, the luminescent 4-methylpyridyl analogue with E = S has been prepared and structurally characterized. For E = S, Se, the change in emission energy with mu3-bridging atom provides a sound basis for an LMMCT assignment of the excited state while lifetime measurements support its spin-forbidden nature. Frozen glass measurements indicate the presence of a higher-energy emitting state for these systems, and for the E = O system, either LMMCT or metal-centered cluster-based emission can be proposed. PMID- 15291523 TI - Self-assembly of heterobimetallic d-f hybrid complexes: sensitization of lanthanide luminescence by d-block metal-to-ligand charge-transfer excited states. AB - Carboxylate-bearing d-transition metal bipyridyls form ternary complexes with seven-coordinate lanthanide centers. The neodymium,- ytterbium-, and erbium containing complexes exhibit lanthanide-centered emissions sensitized by the MLCT states of the d-block components. PMID- 15291524 TI - A minimum free energy reaction path for the E2 reaction between fluoro ethane and a fluoride ion. AB - The prototype binuclear elimination (E2) reaction illustrates the mechanism of a large number of biochemical and industrial applied processes but has received surprisingly little attention in theoretical studies compared to, for example, the substitution (SN2) reaction. This is due to its concerted mechanism, which requires an independent description of the three bonds that are being formed or broken. In this work, we have taken the advantage of a new and promising methodology to efficiently sample intrinsically multidimensional free-energy surfaces. We locate the lowest free-energy reaction path in the 3D configurational space and use this finite-temperature intrinsic reaction coordinate in an umbrella sampling scheme to access the temperature contributions to high accuracy. The small increase of the barrier and the decrease of the overall endothermicity for the E2 reaction due to entropic contributions is non trivial. Moreover, our strategy to efficiently handle multiple reaction coordinates could be a great benefit to many chemistry-related fields, such as enzyme catalysis, reactions in solution, and nucleation processes. PMID- 15291525 TI - On the Mossbauer spectra of isopenicillin N synthase and a model (FeNO)7 (S = 3/2) system. AB - We have carried out a series of density functional theory (DFT) calculations to predict the 57Fe Mossbauer quadrupole splittings (DeltaEQ) and isomer shifts (deltaFe) for the nitrosyl complex of isopenicillin N synthase with the substrate delta-(l-alpha-aminoadipoyl)-l-cysteinyl-d-valine (IPNS.ACV.NO) and an {FeNO}7 (S = 3/2) model system, FeL(NO)(N3)2 (L = N,N',N' '-trimethyl-1,4,7 triazacyclononane). B3LYP predictions on the model compound are in almost exact agreement with experiment. The same DFT methods did not enable the prediction of the experimental DeltaEQ and deltaFe results for IPNS.ACV.NO when using the experimental protein crystal structure but did permit good predictions of DeltaEQ, deltaFe, and the asymmetry parameter (eta) when using a fully optimized structure. This optimized structure also enabled good predictions of the Mossbauer spectra of the photodissociation product of IPNS.ACV.NO. Mulliken and natural bonding orbital (NBO) spin density analyses indicate an electronic configuration of FeII (S = 2) anti-ferromagnetically coupled to NO (S = 1/2) in the protein as well as in the model system and the geometry optimized structure helps explain part of the enzyme reaction. PMID- 15291526 TI - Specific band observed in VCD predicts the anomeric configuration of carbohydrates. AB - The vibrational circular dichroism (VCD) technique was applied to carbohydrates and produced a glycoside characteristic VCD band, which was named the "glycoside band." It confirmed that this vibration was related to anomeric configurations of carbohydrates in the case of the d-series by means of isotope substitution. This band is defined as a negative, sharp, and intense VCD band at around 1145 cm-1, exhibited by d-sugars with an axial alpha-glycosidic linkage in the 4C1 conformation. As applications of this band, the VCD studies of the maltose cellobiose mixture and the monitoring of the amyloglucosidase reaction were examined. PMID- 15291527 TI - Forced folding and structural analysis of metastable proteins. AB - A significant fraction of the proteins encoded by the human and other genomes appears to be significantly unfolded in vitro. This will undoubtedly hamper attempts to characterize their structure by classical crystallographic or solution NMR methods. Here we show that encapsulation of a metastable protein within the restricted volume a reverse micelle can be used to force fold the protein and allow its characterization by modern methods of NMR spectroscopy. This may have significant utility in the context of structural proteomics. In addition, variation of the inner volume of the reverse micelle can be used to probe the character of the manifold of unfolded states. PMID- 15291528 TI - Impact of protein flexibility on hydride-transfer parameters in thermophilic and psychrophilic alcohol dehydrogenases. AB - The hydride transfer catalyzed by thermophilic alcohol dehydrogenase (htADH) exhibits sharply different kinetic and activation parameters from that catalyzed by the more flexible psychrophilic alcohol dehydrogenase (psADH). In addition, the hydride transfer in htADH is affected by mutating two distal residues that are suggested to be responsible for the decreased local protein flexibility in htADH. These observations provide support for the view that protein dynamics is tightly coupled to the hydrogen-transfer processes in these enzymes. PMID- 15291529 TI - Ultrathin "bed-of-nails" membranes of single-wall carbon nanotubes. AB - Single-wall carbon nanotubes (SWNTs) were arranged in a membrane similar to a "bed-of-nails", in which a single layer of parallel SWNTs was densely packed and aligned along the normal to the membrane. The planar, free-standing, ultrathin SWNT membranes were prepared by milling a neat SWNT fiber with a gallium focused ion beam. The approach is readily applicable to cutting nanotubes to a desirable and precise length and enables further fabrication of devices using the "bed-of nails" membranes to test the transport properties of SWNTs. PMID- 15291530 TI - Solid-state NMR spectroscopic studies of an integral membrane protein inserted into aligned phospholipid bilayer nanotube arrays. AB - This communication demonstrates for the first time that solid-state NMR spectroscopic studies can be used to investigate aligned phospholipid bilayer nanotube arrays. Also, an integral membrane peptide can be successfully incorporated into the oriented phospholipid bilayer nanotube arrays and studied with 2H solid-state NMR spectroscopy. PMID- 15291531 TI - Role of membrane potential and hydrogen bonding in the mechanism of translocation of guanidinium-rich peptides into cells. AB - The results described herein support a mechanistic hypothesis for how guanidine rich transporters attached to small cargos (MW ca. <3000) can migrate across the lipid membrane of a cell and directly enter the cytosol. Arginine oligomers are found to partition almost completely into the aqueous layer of a water-octanol bilayer. However, when the same partitioning experiment is conducted in the presence of sodium laurate, a representative negatively charged membrane constituent, the arginine oligomer partitions almost completely (>95%) into the octanol layer. In contrast, ornithine oligomers partition almost exclusively into the water layer with and without added sodium laurate. The different partitioning between guanidinium-rich and ammonium-rich oligomers in the presence of sodium laurate is consistent with the ability of the former to form a bidentate hydrogen bonded ion pair. Mono- and dimethylated arginine oligomers, which like ornithine can only efficiently form monodentate hydrogen bonds, were prepared and found to exhibit poor cellular uptake. Ion pair formation converts a once water-soluble agent to a lipid-soluble agent, thereby reducing the energetic penalty for passage of guanidine-rich transporters through the lipid bilayer. Uptake of guanidine-rich transporters is known to be an energy-dependent process, and this requirement for cellular ATP is now rationalized by the inhibition of guanidine rich transporter uptake in the presence of agents that reduce the membrane potential. Specifically, incubation of cells in buffers with high potassium ion concentrations or pretreatment of cells with gramicidin A reduces the cellular uptake of Fl-aca-arg8-CONH2 by >90%. Furthermore, the reciprocal experiment of hyperpolarizing the cell with valinomycin increased uptake by >1.5 times. In summary, we propose that the water-soluble, positively charged guanidinium headgroups of the transporter form bidentate hydrogen bonds with H-bond acceptor functionality on the cell surface. The resultant ion pair complexes partition into the lipid bilayer and migrate across at a rate related to the membrane potential. The complex dissociates on the inner leaf of the membrane, and the transporter enters the cytosol. This hypothesis does not preclude uptake by other mechanisms, including endocytosis, which is likely to dominate with large cargos. PMID- 15291532 TI - Synthesis of isolable thiirane-2-thione (alpha-dithiolactone) from thioketene S oxide. AB - Reaction of di-tert-butyl thioketene S-oxide with Lawesson reagent gave 3,3-di tert-butylthiirane-2-thione in 88% yield. Oxidation of thiirane-2-thione with m chloroperbenzoic acid afforded 3,3-di-tert-butylthiirane-2-thione S-oxide. The reaction of thiirane-thione was described. PMID- 15291533 TI - Solvent control of spin-dependent charge recombination mechanisms within donor conjugated bridge-acceptor molecules. AB - We have shown recently that the oligomeric p-phenylene bridge within the PTZ (Ph)n-PDI (PTZ = phenothiazine, Ph = phenyl, and PDI = perylenediimide) donor bridge-acceptor system acts as a molecular wire in toluene, as shown by a change in the rate of radical ion pair (RP) recombination within PTZ+*-(Ph)n-PDI-* from an exponential distance dependence to a linear distance dependence as the bridge becomes longer. The population of the RP and its spin-selective recombination products are sensitive to the application of an external magnetic field, which can be used to directly measure the singlet-triplet splitting, 2J, within the RP. The value of 2J is a weighted sum of electronic coupling matrix elements that are to a good approximation directly proportional to VDA2, the effective coupling between the orbitals on the donor and acceptor sites. The dependence of RP population on magnetic field reveals the relative contributions of the singlet and triplet charge recombination (CR) pathways to overall RP decay. We have now observed an "inversion" of the MFE on the RP population within PTZ+*-(Ph)4-PDI-* and PTZ+*-(Ph)5-PDI-* upon a switch in solvent from toluene to 2 methyltetrahydrofuran (MTHF). We interpret the inversion of the MFE as a switch in the relative importance of the singlet and triplet charge recombination (CRS, CRT) pathways due to a stabilization of the RP state by more polar MTHF, making CRS more energetically favorable. This change in mechanism illustrates the sensitivity of molecular wire behavior to the surrounding environment. PMID- 15291534 TI - Hydrogel microparticles as dynamically tunable microlenses. AB - Tunable micro-optical elements were prepared by aqueous free-radical polymerization and electrostatic self-assembly techniques. Stimuli-responsive poly(N-isopropylacrylamide-co-acrylic acid) (pNIPAm-AAc) microgels were used as lenses to generate dynamically tunable optical elements. By using optical microscopy to investigate the micrometer-scale dynamics of the self-assembled microlenses, we demonstrate focal length tuning through modulation of the solution pH and/or temperature. PMID- 15291535 TI - Dissociation of acetaldehyde on beta-Mo2C to yield ethylidene and oxo surface groups: a possible pathway for active site formation in heterogeneous olefin metathesis. AB - The dissociative adsorption of acetaldehyde on beta-Mo2C was studied using reflection absorption infrared spectroscopy. In contrast to what is observed for all metals previously studied, acetaldehyde undergoes selective carbonyl bond scission on the carbide surface. By comparison to calculated spectra, the surface product is identified as an oxo-ethylidene species. The study thus provides the first extended-range infrared spectrum of a propene metathesis initiator or propagator alkylidene. Aldehydes may be formed in the presence of olefins during the induction period of supported metal oxide olefin metathesis catalysts. Hence, the observed dissociative chemisorption of acetaldehyde suggests a possible answer to the question of how initiator sites are formed in heterogeneous olefin metathesis. This question has never been satisfactorily answered. In the proposed mechanism, aldehydes formed during the induction period subsequently react with the catalyst surface to generate alkylidene sites. PMID- 15291536 TI - Activating the phosphate nucleophile at the catalytic site of purine nucleoside phosphorylase: a vibrational spectroscopic study. AB - Difference Raman and FTIR studies complemented by vibrational analysis based on ab initio calculations show that the dianionic phosphate in the PNP.ImmH.PO4 complex is forced into a unique bonding arrangement in which one of the PO bonds is greatly polarized by enzyme active site interactions, such that it resembles a PO bond that is about one-quarter of the way toward forming a bridging P-O-C single P-O bond. PMID- 15291537 TI - Conversion of alpha-haloaldehydes into acylating agents by an internal redox reaction catalyzed by nucleophilic carbenes. AB - Reactivity umpolung allows us to consider nontraditional bond disconnections. We report herein that treatment of an alpha-haloaldehyde with a nucleophile in the presence of catalytic amounts of nucleophilic carbenes results in an internal redox reaction giving rise to a dehalogenated acylating agent as an intermediate by a new reaction manifold. A brief illustration of the scope of this reaction is presented along with evidence supporting the direct intervention of the carbene in the acylation step. PMID- 15291538 TI - IR spectrum and structure of protonated ethanol dimer: implications for the mobility of excess protons in solution. AB - This Communication reports IR spectra and density functional calculations for the isolated protonated ethanol dimer and its N2-microsolvated complexes, (EtOH)2H+ (N2)n (n = 0-2) to investigate the degree of delocalization of the excess proton in this fundamental building block of an alcohol proton wire. The first spectroscopic characterization of isolated and microsolvated (EtOH)2H+ suggests that the excess proton is (nearly) equally shared between both EtOH units under symmetric solvation conditions (Zundel-type ion, n = 0 and 2), whereas it is largely localized on a single EtOH molecule for asymmetric solvation (Eigen-type ion, n = 1). PMID- 15291539 TI - Formation and utility of oxasilacyclopentenes derived from functionalized alkynes. AB - Oxasilacyclopentenes were shown to be synthetically useful masked allylic alcohols constructed in high yields and regioselectivities from terminal and internal alkynes. Several functional groups were shown to be tolerated utilizing silver-catalyzed silacyclopropenation of alkynes. In situ insertion of various carbonyl compounds into silacyclopropenes afforded regioselective formation of oxasilacyclopentenes. Elaboration of the oxasilacyclopentenes displayed the synthetic utility of these substrates. Both diastereoselective hydrogenation and Diels-Alder reactions utilizing the vinylsilane functionality demonstrated the reactivity and synthetic utility of oxasilacyclopentenes. PMID- 15291540 TI - Tandem cyclopropanation/ring-closing metathesis of dienynes. AB - Certain dienynes give cyclorearrangement by tandem cyclopropanation/ring-closing alkene metathesis, triggered by either a ruthenium carbene or noncarbene ruthenium(II) precatalyst. The process represents a variation of enyne metathesis where presumed cyclopropyl carbene intermediates undergo a consecutive ring closing metathesis. A mechanistic proposal is offered, and sequential use of catalysts provided a tandem ring-closing enyne/alkene metathesis product. PMID- 15291541 TI - Formation of dimers that contain unbridged W(IV)/W(IV) double bonds. AB - A tungsten neopentylidene complex has been found to decompose to yield a heterochiral dimer that contains a W=W double bond and no bridging ligands. Decompositions of related bisalkoxide complexes also yield compounds that contain an "unsupported" W=W double bond, while a sample of [Mo(NAr)(CH2-t-Bu)(OC6F5)]2 has been found to be a homochiral species related to [W(NAr)(CH2-t-Bu)(OC6F5)]2. PMID- 15291542 TI - Just add water: a new fluorous capping reagent for facile purification of peptides synthesized on the solid phase. AB - A novel fluorous capping reagent is introduced to facilitate purification during solid-phase peptide synthesis (SPPS). Reagent 1 is a trivalent iodonium salt that reacts vigorously with free amines to deliver a long-chain fluoroalkyl group. It has been used to tag all unreacted amines following the peptide coupling step in SPPS. The resulting fluoroalkylated amine is no longer able to couple in further peptide coupling steps and is also stable to standard peptide synthesis conditions. Deletion products are removed using flash fluorous chromatography to yield the pure, full-length peptide. PMID- 15291543 TI - Cyanogen bromide cleavage generates fragments suitable for expressed protein and glycoprotein ligation. AB - Herein cyanogen bromide is employed for the efficient production of N-terminal cysteine containing protein fragments for expressed protein ligation (EPL) from polyhistidine-tagged precursors. We provide three examples of efficient CNBr cleavage of fragments of the glycoprotein erythropoietin that can be ligated with peptides or glycopeptide mimetics potentially giving rise to semisynthetic glycoprotein therapeutics. PMID- 15291544 TI - 3-nitro-3-deaza-2'-deoxyadenosine as a versatile photocleavable 2'-deoxyadenosine mimic. AB - A new photocleavable 2'-deoxyadenosine mimic, 3-nitro-3-deaza-2'-deoxyadenosine (NidA), was prepared and introduced into DNA fragments via its 6-O trimethylphenyl precursor phophoramidite. Photocleavage of the resulting oligonucleotide is highly efficient in single and double strands. Hybridization properties of NidA are very similar to those of deoxyadenosine. PMID- 15291545 TI - Studying lateral diffusion in lipid bilayers by combining a magic angle spinning NMR probe with a microimaging gradient system. AB - An alternative technique for studying the lateral diffusion in lipid membranes using pulsed field gradients in combination with magic angle spinning is presented. It is shown that MAS probes inserted in a microimaging device that produce high field gradients can be used to monitor also slow diffusion processes. As an example, measurements of the lateral diffusion of lipids embedded in the bilayer of a cubic phase are presented. PMID- 15291546 TI - Platinum-catalyzed intramolecular hydroalkoxylation of gamma- and delta-hydroxy olefins to form cyclic ethers. AB - Reaction of 2,2-diphenyl-4-penten-1-ol with a catalytic mixture of [PtCl2(H2C=CH2)]2 (1 mol %) and P(4-C6H4CF3)3 (2 mol %) at 70 degrees C for 24 h led to the isolation of 2-methyl-4,4-diphenyltetrahydrofuran in 78% yield. The platinum-catalyzed hydroalkoxylation of gamma-hydroxy olefins tolerated substitution at the alpha, beta, and gamma-carbon atoms and at the internal and cis and trans terminal olefinic positions. Platinum-catalyzed hydroalkoxylation tolerated a number of functional groups including pivaloate and acetate esters, amides, silyl and benzyl ethers, and pendant hydroxyl and olefinic groups. Pt catalyzed olefin hydroalkylation was also applicable to the formation of fused- and spirobicyclic ethers and was effective for the hydroalkoxylation of delta hydroxy olefins to form tetrahydropyran derivatives. PMID- 15291547 TI - Human serum albumin-mediated stereodifferentiation in the triplet state behavior of (S)- and (R)-carprofen. AB - A remarkable stereodifferentiation has been observed in the interaction between the excited triplet state of carprofen (CP) and human serum albumin (HSA). Time resolved measurements using laser flash photolysis reveal the presence of two components with different lifetimes in triplet decay. This is explained by complexation of CP to the two possible HSA binding sites. The shorter-lived components are ascribed to the CP/HSA complexes in site I, where stereodifferentiation is more important (tauR/tauS ca. 4). This is correlated with formation of a dehalogenated photoproduct upon steady-state photolysis. PMID- 15291548 TI - Spermine participates in oxidative damage of guanosine and 8-oxoguanosine leading to deoxyribosylurea formation. AB - Oxidation of single- or double-stranded DNA containing a 7,8-dihydro-8 oxoguanosine lesion with the one-electron oxidant Na2IrCl6 in the presence of spermine led to formation of a covalent adduct that was analyzed by gel electrophoresis, HPLC, ESI-MS, and UV-vis. The adduct was labile to heat, exhibiting a t1/2 of 12 h at 37 degrees C, and the ultimate hydrolysis product was characterized as a deoxyribosylurea lesion. Data from model studies with 1,3 diaminopropane vs 1,4-diaminobutane are consistent with initial formation of a C5 spermine adduct from a dehydro-8-oxoguanosine intermediate, followed by rearrangement to a spiroaminal subject to slow hydrolysis at C4 of the purine. Spermine adducts could also be formed from oxidation of the analogous G containing oligomer from reaction with singlet oxygen, albeit in lower yield. These results are surprising in light of the traditional view that spermine is radioprotective against DNA oxidation. PMID- 15291549 TI - Palladium-catalyzed oxygenation of unactivated sp3 C-H bonds. AB - This communication describes a new palladium-catalyzed method for the oxygenation of unactivated sp3 C-H bonds. A wide variety of alkane substrates containing readily available oxime and/or pyridine directing groups are oxidized with extremely high levels of chemo-, regio-, and in some cases diastereoselectivity. The substrate scope of these reactions is discussed, and the high selectivities are rationalized on the basis of the requirements of putative palladium alkyl intermediates. PMID- 15291550 TI - Enantioselective Nazarov reactions through catalytic asymmetric proton transfer. AB - The development of catalytic asymmetric Nazarov reactions that require only 10 mol % of chiral Lewis acid and proceed with ee's between 72% and 97% is described. PMID- 15291551 TI - Synthesis of bistramide A. AB - We have developed an efficient and highly stereocontrolled synthesis of bistramide A, a selective activator of protein kinase C isotype delta. Our synthetic strategy featured a novel bidirectional approach for spiroketal construction based on the ring-opening/cross-metathesis sequence employing a highly strained cyclopropenone acetal. The synthesis afforded the final target with the longest linear sequence of 15 steps and provided unambiguous structural determination of bistramide A, including assignment of the previously unknown C(37) stereochemistry. PMID- 15291552 TI - Engineering human Fhit, a diadenosine triphosphate hydrolase, into an efficient dinucleoside polyphosphate synthase. AB - The putative human tumor suppressor gene FHIT encodes Fhit, the fragile histidine triad protein. Fhit is thought to participate in a signal transduction pathway involving dinucleoside polyphosphates. Fhit catalyzes the Mg2+-dependent hydrolysis of P1-5'-O-adenosine-P3-5'-O-adenosine triphosphate (Ap3A) to AMP and MgADP. Mutation of His96 to glycine disables Fhit as a catalyst for the hydrolysis of phosphoanhydrides such as Ap3A. However, the mutated enzyme H96G Fhit efficiently catalyzes the synthesis of phosphoanhydride bonds in reactions of nucleoside-5'-phosphimidazolides with nucleoside di- and triphosphates. H96G Fhit can be employed in the synthesis of a wide range of dinucleoside tri- and tetraphosphates. We here describe the use of H96G-Fhit to catalyze the synthesis of Ap3A, Ap3C, Ap3G, Ap3T, Ap3U, Cp3U, Tp3U, dAp3U, Ap4A, Ap4U, and the fluorescent Ap4etheno-C. PMID- 15291553 TI - Aerobic oxidation of 2,3,6-trimethylphenol to trimethyl-1,4-benzoquinone with copper(II) chloride as catalyst in ionic liquid and structure of the active species. AB - TMQ is an important precursor in industrial vitamin E synthesis. We report a "green chemistry approach" with respect to the highly selective and environmentally friendly oxidation of 2,3,6-trimethylphenol (TMP) to trimethyl 1,4-benzoquinone (TMQ) with molecular oxygen as oxidant and a copper catalyst immobilized in a molten salt. n-Butanol as co-solvent has a positive effect on the activity and selectivity. The structurally characterized catalyst, a 1-n butyl-3-methylimidazolium oxotetracuprat, is formed in situ via hydrolysis of CuCl2 in the presence of imidazolium chloride. We propose a mechanism of oxidative phenolate activation at a [Cu4(mu4-O)]6+ core as electronically coupled electron acceptor, formation of a copper-bound phenolate radical anion, spin delocalization into the aromatic ring, and attack by triplet oxygen at the para position. Attack of Cu(I) as reduction equivalent at the peroxy radical, proton mediated elimination of a copper(II)-hydroxo species, will either substitute a copper(I) site in the reduced oxo cluster or take up an electron from the reduced mixed valent cluster [Cu4(mu4-O)]6+ to regenerate the oxidized cluster as the active electron acceptor. PMID- 15291554 TI - The total synthesis of (+)-dragmacidin F. AB - The first total synthesis of (+)-dragmacidin F has been accomplished, establishing the absolute configuration of this biologically important, antiviral marine alkaloid. The convergent route described features a palladium-mediated oxidative pyrrole carbocylization reaction to construct the [3.3.1] bicycle, as well as a highly selective Suzuki coupling to build the carbon skeleton of the natural product. A late-stage Neber rearrangement allows for the facile installation of the aminoimidazole moiety to provide (+)-dragmacidin F. PMID- 15291555 TI - Highly regio- and stereoselective rearrangement of epoxides to aldehydes catalyzed by high-valent metalloporphyrin complex, Cr(TPP)OTf. AB - Chromium(III) tetraphenylporphyrin triflate, Cr(TPP)OTf, works as an efficient and characteristic Lewis acid catalyst in the regio- and stereoselective rearrangement of epoxides to aldehydes. This Cr(TPP)OTf-catalyzed reaction is an operationally simple and especially convenient method for synthesizing optically active beta-siloxy aldehydes from 2,3-epoxy silyl ethers which are readily available in enantiomerically enriched forms by the Sharpless epoxidation of allylic alcohols followed by silylation. PMID- 15291556 TI - Metal latticeworks formed by self-organization in oscillatory electrodeposition. AB - Strikingly well-ordered Sn latticeworks, standing perpendicular to the substrate, are formed spontaneously in oscillatory electrodeposition. Cooperation of various processes, such as needle formation by autocatalytic crystal growth, cuboid formation under a reaction-limited condition, and autocatalytic oxidation at closest-packed surfaces, enabled the self-organization of the latticeworks. The mechanism is generally applicable to deposition of other metals such as Zn, Pb, and Cu. The present work has opened a promising, unique field toward the formation of highly ordered 3-D micro- or nanostructures at solid surfaces. PMID- 15291557 TI - A new class of chiral pyrrolidine-pyridine conjugate base catalysts for use in asymmetric Michael addition reactions. AB - Direct catalytic asymmetric Michael addition reactions of ketones to nitroolefins using newly developed chiral pyrrolidine-pyridine conjugate bases as catalysts are described. The desired 1,4-adducts are obtained in excellent yields with high enantio- and diastereoselectivities. PMID- 15291558 TI - Toward fully synthetic carbohydrate-based HIV antigen design: on the critical role of bivalency. AB - Synthetic gp120331-335 glycopeptide fragments carrying hybrid and high-mannose type N-linked glycans were evaluated for binding to broadly neutralizing antibody 2G12 using surface plasmon resonance technology. None of the hybrid-type constructs demonstrated binding to 2G12. In the high-mannose series, the "Cys dimer" construct, presenting two undecasaccharide glycans, showed significantly higher binding than the Cys-protected monomer. The binding of the dimeric structure was further investigated in competition with recombinant gp120. The data suggest that gp120 and its designed synthetic epitope construct bind to the same site on 2G12. PMID- 15291559 TI - An organometallic intermediate during alkyne reduction by nitrogenase. AB - Nitrogenase is the metalloenzyme that catalyzes the nucleotide-dependent reduction of N(2), as well as reduction of a variety of other triply bonded substrates, including the alkyne, acetylene. Substitution of the alpha-70(Val) residue in the nitrogenase MoFe protein by alanine expands the range of substrates to include short-chain alkynes not reduced by the unaltered protein. Rapid freezing of the alpha-70(Ala) nitrogenase MoFe protein during reduction of the alkyne propargyl alcohol (HC triple bond CH(2)OH; PA) traps an S = (1)/(2) intermediate state of the active-site metal cluster, the FeMo-cofactor. We have combined CW and pulsed (13)C ENDOR (electron-nuclear double resonance) with two quantitative 35 GHz (1,2)H ENDOR techniques, Mims pulsed ENDOR and the newly devised "stochastic field-modulated" ENDOR, to study this intermediate prepared with isotopically substituted ((13)C, (1,2)H) propargyl alcohol in H(2)O and D(2)O buffers. These measurements allow the first description of a trapped nitrogenase reduction intermediate. The S = (1)/(2) turnover intermediate generated during the reduction of PA contains the 3-carbon chain of PA and exhibits resolved (1,2)H ENDOR signals from three protons, two strongly coupled (H(a)) and one weakly coupled (H(b)); H(a)(c) originates as the C3 proton of PA, while H(a)(s) and H(b) are solvent-derived. The two H(a) protons have identical hyperfine tensors, despite having different origins. The equality of the (H(a)(s), H(a)(c)) hyperfine tensors strongly constrains proposals for the structure of the cluster-bound reduced PA. Through consideration of model structures found in the Cambridge Structural Database, we propose that the intermediate contains a novel bio-organometallic complex in which a reduction product of propargyl alcohol binds as a metalla-cyclopropane ring to a single Fe atom of the Fe-S face of the FeMo-cofactor that is composed of Fe atoms 2, 3, 6, and 7. Of the two most attractive structures, one singly reduced at C3 (4), the other being the doubly reduced allyl alcohol product (6), we tentatively favor 6 because of the "natural" assignment it affords for H(b). PMID- 15291560 TI - Structural and energetic consequences of expanding a highly cooperative stable DNA hairpin loop. AB - Many hairpin loops are expanded versions of smaller, stable ones. Herein we investigate the extent to which the energetics and structure of d(cGNAg) hairpin loops will tolerate sequence variation. Changing the closing base pair from CG to GC was found to completely eliminate loop-loop interactions; in contrast, expanding the loop at the 3'-end resulted in similar energetics and nonadditivity parameters as the parent loop, suggesting that loop-loop interactions remain intact and highly coupled upon expansion. Together, these data suggest that the CG closing base pair forms an essential platform upon which a stable d(GNA) hairpin loop can fold and that this loop can undergo 3'-expansion with little effect to its structure or energetics. PMID- 15291561 TI - Selective derivatization and sequestration of ribose from a prebiotic mix. AB - Observations regarding the catalytic potential of RNA and the role of RNA in biology have formed the basis for the "RNA world" hypothesis, which suggests that a genetic system based on self-replicating polyribonucleotides preceded modern biology. However, attempts to devise a realistic prebiotic synthesis of nucleic acids from simple starting materials have been plagued by problems of poor chemical selectivity, lack of stereo- and regiospecificity, and similar rates of formation and degradation of some of the key intermediates. For example, ribose would have been only a small component of a highly complex mix of sugars resulting from the condensation of formaldehyde in a prebiotic world. In addition, ribose is more reactive and degrades more rapidly compared with most other monosaccharides. This study demonstrates an approach for the preferential sequestration of ribose relative to other sugars that takes advantage of its greater reactivity. Cyanamide reacts especially rapidly with ribose to form a stable bicyclic adduct. This product crystallizes spontaneously in aqueous solution, whereas the corresponding products derived from threose, galactose, glucose, mannose, and each of the other pentoses do not. Furthermore, when employing a racemic mixture of d- and l-ribose, enantiomerically twinned crystals are formed that contain discrete homochiral domains. PMID- 15291562 TI - Amino acid-type edited NMR experiments for methyl-methyl distance measurement in 13C-labeled proteins. AB - New NMR experiments are presented for the measurement of methyl-methyl distances in (13)C-labeled proteins from a series of amino acid-type separated 2D or 3D NOESY spectra. Hadamard amino acid-type encoding of the proximal methyl groups provides the high spectral resolution required for unambiguous methyl-methyl NOE assignment, which is particularly important for fast global fold determination of proteins. The experiments can be applied to a wide range of protein systems, as exemplified for two small proteins, ubiquitin and MerAa, and the 30 kDa BRP-Blm complex. PMID- 15291563 TI - Origin of enantioselectivity in the asymmetric Ru-catalyzed metathesis of olefins. AB - The mechanism of enantioselectivity in the asymmetric Ru-catalyzed metathesis of olefins is investigated with a theoretical approach. The models are based on the chiral N-heterocyclic (NHC)-based catalysts developed by Grubbs. Our analysis indicates that the origin of enantioselectivity in the ring-closing metathesis of achiral trienes is correlated to the chiral folding of the N-bonded aromatic groups, which is imposed by the Ph groups in positions 4 and 5 of the imidazole ring of the NHC ligand. This chiral folding of the catalyst imposes a chiral orientation around the Ru=C bond, which, in turn, selects between the two enantiofaces of the substrate. In the ring-closing transition state, the geometry in which additional groups on the forming ring are in pseudoequatorial positions is favored over transition states in which this additional group is in a pseudoaxial position. These combined effects rationalize the enantiomeric excesses experimentally obtained. PMID- 15291564 TI - 2,2,5,5-tetramethylpyrrolidin-3-one-1-sulfinyl group for 5'-hydroxyl protection of deoxyribonucleoside phosphoramidites in the solid-phase preparation of DNA oligonucleotides. AB - Several nitrogen-sulfur reagents have been investigated as potential 5'-hydroxyl protecting groups for deoxyribonucleoside phosphoramidites to improve the synthesis of oligonucleotides on glass microarrays. Out of the nitrogen-sulfur based protecting groups so far investigated, the 2,2,5,5-tetramethylpyrrolidin-3 one-1-sulfinyl group exhibited near optimal properties for 5'-hydroxyl protection by virtue of the mildness of its deprotection conditions. Specifically, the iterative cleavage of a terminal 5'-sulfamidite group in the synthesis of 5' d(ATCCGTAGCCAAGGTCATGT) on controlled-pore glass is efficiently accomplished by treatment with iodine in the presence of an acidic salt. Hydrolysis of the oligonucleotide to its 2'-deoxyribonucleosides upon exposure to snake venom phosphodiesterase and bacterial alkaline phosphatase did not reveal the formation of any nucleobase adducts or other modifications. These findings indicate that the 2,2,5,5-tetramethylpyrrolidin-3-one-1-sulfinyl group for 5'-hydroxyl protection of phosphoramidites, such as 10a-d, may lead to the production of oligonucleotide microarrays exhibiting enhanced specificity and sensitivity in the detection of nucleic acid targets. PMID- 15291565 TI - Silicate complexes of sugars in aqueous solution. AB - Certain sugars react readily with basic silicic acid to form soluble 2/1 (sugar/silicic acid) silicate complexes. Failure of monohydroxy compounds to give soluble products under these conditions indicates that the sugar silicates are chelates: five-membered diolato rings. Only furanose forms react. Pyranose sugars are stable under these conditions. Because all glycosides fail to react with silicic acid under these conditions, reaction appears to involve the anomeric position (C1 in aldoses, C2 in ketoses), which has a more acidic hydroxy group. Reaction is completed only when the anomeric hydroxy group is cis to an adjacent hydroxy group. The appropriate furanose form must have sufficient natural abundance and solubility to provide an observable product, as measured by (29)Si and (13)C NMR spectroscopy. These structural and practical constraints rationalize the successful reaction of the monosaccharides ribose, xylose, lyxose, talose, psicose, fructose, sorbose, and tagatose and the disaccharides lactulose, maltulose, and palatinose. Glucose, mannose, galactose, and sucrose, among others, failed to form complexes. This high selectivity for formation of sugar silicates may have ramifications in prebiotic chemistry. PMID- 15291566 TI - Isolable anion radical of blue disilene (tBu2MeSi)2Si=Si(SiMetBu2)2 formed upon one-electron reduction: synthesis and characterization. AB - The highly twisted tetrakis(di-tert-butylmethylsilyl)disilene 4 was prepared and reacted with (t)BuLi in THF, producing disilene anion radical 5 upon one-electron reduction. The anion radical 5 was isolated in the form of its lithium salt as extremely air- and moisture-sensitive red crystals. The molecular structure of 5 was established by X-ray crystallography, which showed a nearly orthogonal structure (twisting angle of 88 degrees ) along the central Si-Si bond, with a length of 2.341(5) A, which is 3.6% elongated relative to that of 4. The interesting feature of 5 is that one of the central Si atoms has radical character, whereas the other Si atom has silyl anion character. An electron spin resonance (ESR) study of the hyperfine coupling constants of the (29)Si nuclei indicates that rapid spin exchange occurs between these central Si atoms on the ESR time scale. PMID- 15291567 TI - Supramolecular control over donor-acceptor photoinduced charge separation. AB - A novel donor-bridge-acceptor system has been synthesized by covalently linking a p-phenylene vinylene oligomer (OPV) and a perylene diimid (PERY) at opposite ends of a m-phenylene ethynylene oligomer (FOLD) of twelve phenyl rings, containing nonpolar (S)-3,7-dimethyl-1-octanoxy side chains. For comparison, model compounds have been prepared in which either the donor or acceptor is absent. In chloroform, the oligomeric bridge is in a random coil conformation. Upon addition of an apolar solvent (heptane) the oligomeric bridge first folds into a helical stack and subsequently intermolecular self-assembly of the stacks into columnar architectures occurs. Photoexcitation in the random coil conformation, where the interaction between the donor and acceptor chromophores is small, results only in long-range intramolecular energy transfer in which the OPV singlet-excited state is transformed into the PERY singlet-excited state. In the folded conformation of the bridge, donor and acceptor are closer and their enhanced interaction favors the formation the OPV(*)(+)-FOLD-PERY(*)(-) charge-separated state upon photoexcitation. As a result, the extent of photoinduced charge separation depends on the degree of folding of the bridge between donor and acceptor and therefore on the apolar nature of the medium. As a consequence, and contrary to conventional photoinduced charge separation processes, the formation of the OPV(*)(+)-FOLD-PERY(*)(-) charge-separated state is more favored in apolar media. PMID- 15291568 TI - Catalytic regioselectivity control in ring-opening cycloisomerization of methylene- or alkylidenecyclopropyl ketones. AB - 2-Methylene- or alkylidenecyclopropanyl ketones were easily prepared by the regioselective cyclopropanation of allenes or the reaction of methylene /alkylidenecyclopropanyllithium with N,N-dimethyl carboxylic acid amides. Due to the presence of the exo-cyclic C=C bond and the strained cyclopropane, their highly selective ring-opening cycloisomerization using PdCl(2)(CH(3)CN)(2), NaI (or PdCl(2)(CH(3)CN)(2) + NaI), and Pd(PPh(3))(4) as catalysts provided five different products, i.e., 4H-pyrans, 2,3,4-trisubstituted furans (or 4,5 disubstituted-3-alkylidene-2,3-dihydrofurans), and 2,3,4,5-tetrasubtituted furans (or 2,4,5-trisubstituted-3-alkylidene-2,3-dihydrofurans) in good yields, respectively, depending on the nature of the catalyst and reaction conditions. The less-substituted C=C bonds in these products can be highly selectively hydrogenated or hydroborated to afford new heterocyclic products stereoselectively. These three types of different reactions may proceed through a highly regioselective cleavage of a carbon-carbon single bond in the cyclopropane ring triggered by regioselective halometalation of the C=C bond and beta decarbopalladation, halogen anion attack on the nonsubstituted carbon atom of the cyclopropane ring, or the direct oxidative addition of the distal carbon-carbon single bond of the cyclopropane ring with Pd(0). In some cases substituent effects were successfully applied to synthesize 2H-pyrans 8 and 3-alkylidene-2,3 dihydrofurans 5, which also provided some mechanistic information. PMID- 15291569 TI - Effect of meta electron-donating groups on the electronic structure of substituted phenyl nitrenium ions. AB - Density functional theory (UB3LYP/6-31G(d,p)) was used to determine substituent effects on the singlet-triplet-state energy gap for 21 meta-substituted phenylnitrenium ions. It was found that strongly electron-donating substituents stabilize the triplet state relative to the singlet state. With sufficiently strong meta electron donors (e.g., m,m'-diaminophenylnitrenium ion) the triplet is predicted to be the ground state. Analysis of equilibrium geometries, Kohn Sham orbital distributions, and Mulliken spin densities for the triplet states of this series of nitrenium ions leads to the conclusion that there are two spatially distinct types of low-energy triplet states. Simple arylnitrenium ions such as phenylnitrenium ions as well as those having electron-withdrawing or weakly donating meta substituents have lowest-energy triplet states that are n,pi in nature. That is, one singly occupied molecular orbital is orthogonal to the plane of the phenyl ring and one is coplanar. These n,pi triplets are generally characterized by large ArNH bond angles (ca. 130-132 degrees ) and an NH bond that is perpendicular to the plane of the phenyl ring. In contrast, meta donor arylnitrenium ions have a lowest-energy triplet state best described as pi,pi. That is, both singly occupied molecular orbitals are orthogonal to the aromatic ring. Such pi,pi states are characterized by NH bonds that are coplanar with the phenyl ring and have ArNH bond angles that are more acute (ca. 110-111 degrees ). These triplet nitrenium ions have electronic structures analogous to those of meta-benzoquinodimethane derivatives. PMID- 15291570 TI - Encapsulated guest molecules in the dimer of octahydroxypyridine[4]arene. AB - Recently a new type of calix[4]arenes has been synthesized via condensation of 2,6-dihydroxypyridine and a number of aldehydes. This type of pyridine[4]arenes forms capsules consisting of two single pyridine[4]arenes. These capsules can incorporate different guest molecules, like carboxylic acids and amides in this case. We proved that the guest acids really are incorporated inside the cavity of the capsules by electrospray mass spectrometry, NMR spectroscopy, and theoretical calculations. PMID- 15291571 TI - Assembling of amphiphilic highly branched molecules in supramolecular nanofibers. AB - We found that the amplification of weak multiple interactions between numerous peripheral branches of irregular, flexible, polydisperse, and highly branched molecules can facilitate their self-assembly into nanofibrillar micellar structures at solid surfaces and the formation of perfect long microfibers in the course of crystallization from solution. The core-shell architecture of the amphiphilic dendritic molecules provides exceptional stability of one-dimensional nanofibrillar structures. The critical condition for the formation of the nanofibrillar structures is the presence of both alkyl tails in the outer shell and amine groups in the core/inner shell. The multiple intermolecular hydrogen bonding and polar interactions between flexible cores stabilize these nanofibers and make them robust albeit flexible. This example demonstrates that one dimensional supramolecular assembling at different spatial scales (both nanofibers and microfibers) can be achieved without a tedious, multistep synthesis of shape-persistent molecules. PMID- 15291572 TI - Development of highly diastereo- and enantioselective direct asymmetric aldol reaction of a glycinate Schiff base with aldehydes catalyzed by chiral quaternary ammonium salts. AB - A highly efficient direct asymmetric aldol reaction of a glycinate Schiff base with aldehydes has been achieved under mild organic/aqueous biphasic conditions with excellent stereochemical control, using chiral quaternary ammonium salt 1b as a phase-transfer catalyst. The initially developed reaction conditions, using 2 equiv of aqueous base (1% NaOH (aq)), exhibited inexplicably limited general applicability in terms of aldehyde acceptors. The mechanistic investigation revealed the intervention of an unfavorable yet inevitable retro aldol process involving the chiral catalyst. On the basis of this information, a reliable procedure has been established by use of a catalytic amount of 1% NaOH (aq) and ammonium chloride, which tolerates a wide range of aldehydes to afford the corresponding anti-beta-hydroxy-alpha-amino esters almost exclusively in an essentially optically pure form. PMID- 15291573 TI - Antibody-catalyzed oxy-cope rearrangement: mechanism and origins of catalysis and stereoselectivity from DFT quantum mechanics and flexible docking. AB - Density functional theory using B3LYP and flexible ligand docking methods were used to investigate the complete potential surface for the uncatalyzed and the AZ28 antibody-catalyzed oxy-Cope reaction of 2,5-diaryl-1,5-hexadien-3-ol derivatives. The reaction mechanism is stepwise, involving a cyclohexane diyl intermediate. Theoretical deuterium isotope effects match well with those from experiment. Gas-phase transition structures show mixed preferences for the axial vs equatorial hydroxyl group, while solvation favors the axial isomer. Specific phenyl group conformations are shown to be critical to transition-state stabilization (by up to 15 kcal/mol), and the effective conformation is not that found in the hapten used to generate the germline and affinity-matured AZ28 catalytic antibodies. Docking studies support greater transition-state binding than reactant binding. Docking studies also predict the S stereoselectivity of mature AZ28 and suggest that binding modes quite different from those of the hapten might play a role in catalysis, with specific focus on ligand hydrogen bonding to Asp(H101). PMID- 15291574 TI - Intermolecular electron transfer from naphthalene derivatives in the higher triplet excited states. AB - Intermolecular electron transfer (ELT) from a series of naphthalene derivatives (NpD) in the higher triplet excited states (T(n)) to carbon tetrachloride (CCl(4)) in Ar-saturated acetonitrile was observed using the two-color two-laser flash photolysis method. The ELT efficiency depended on the driving force of ELT. Since the ELT from the T(n) state occurred competitively with the internal conversion (IC, T(n) --> T(1)) and the triplet energy transfer (ENT), the ELT became apparent only when sufficient free energy change of ELT was attained. On the other hand, ELT from the T(1) state was not observed, although ELT from the T(1) state with sufficiently long lifetime has a slightly exothermic driving force. The fast ELT from the T(n) state and lack of the reactivity of the T(1) state were explained well by the "sticky" dissociative electron-transfer model based on one-electron reductive attachment to CCl(4) leading to the C-Cl bond cleavage. PMID- 15291575 TI - A combined computational and experimental study of polynuclear Ru-TPPZ complexes: insight into the electronic and optical properties of coordination polymers. AB - We report a combined experimental and computational study of polynuclear [Ru(n)(TPPZ)(n)(+1)](2)(n)(+) complexes, of interest in the field of photoactive polymers. The complexes with n = 1, 2, 3 and n > 5 have been synthesized and spectroscopically characterized. A red-shift of the visible band maximum from 2.59 to 2.03 eV is observed going from the monomer to the longer oligomeric species (n > 5). To characterize the geometries, electronic structure, and excited states of these complexes, density functional theory (DFT) and time dependent DFT calculations on the [Ru(n)(TPPZ)(n)(+1)](2)(n)(+) series with n = 1 4 in solution have been performed. The agreement between experimental and calculated spectra is good, both in terms of absorption maximum energies and relative intensities for different values of n. For all the investigated complexes, we assign the main band in the visible region as a metal-to-metal plus ligand charge transfer (MMLCT) transition. The resulting excited states are delocalized throughout the entire complexes, as they originate from a superposition of pi(TPPZ)-t(2g)(Ru) states. The low-energy shoulder of the main visible absorption band, present in the experimental spectra for n > 1, is proposed to arise from spin-forbidden singlet-triplet transitions of similar MMLCT character, consistent with the observed enhancement of this feature in the spectra of the corresponding Os oligomers. PMID- 15291576 TI - Elucidating the significance of beta-hydride elimination and the dynamic role of acid/base chemistry in a palladium-catalyzed aerobic oxidation of alcohols. AB - The mechanistic details of aerobic alcohol oxidation with catalytic Pd(IiPr)(OAc)(2)(H(2)O) (IiPr = 1,3-bis(2,6-diisopropylphenyl)imidazol-2-ylidene) are disclosed. Under optimal conditions, beta-hydride elimination is rate limiting supported by kinetic studies including a high primary kinetic isotope effect (KIE) value of 5.5 +/- 0.1 and a Hammett rho value of -0.48 +/- 0.04. On the basis of these studies, a late transition state is proposed for beta-hydride elimination, which is further corroborated by theoretical calculations using density functional theory. Additive acetic acid modulates the rates of both the alcohol oxidation sequence and regeneration of the Pd catalyst. With no additive [HOAc], turnover-limiting reprotonation of intermediate palladium peroxo is kinetically competitive with beta-hydride elimination, allowing for reversible oxygenation and decomposition of Pd(0). With additive [HOAc] (>2 mol %), reprotonation of the palladium peroxo is fast and beta-hydride elimination is the single rate-controlling step. This proposal is supported by an apparent decomposition pathway modulated by [HOAc], a change in alcohol concentration dependence, a lack of [O(2)] dependence at high [HOAc], and significant changes in the KIE values at different HOAc concentrations. PMID- 15291578 TI - Adsorption of gases and vapors on nanoporous Ni2(4,4'-Bipyridine)3(NO3)4 metal organic framework materials templated with methanol and ethanol: structural effects in adsorption kinetics. AB - Desolvation of Ni(2)(4,4'-bipyridine)(3)(NO(3))(4).2CH(3)OH and Ni(2)(4,4' bipyridine)(3)(NO(3))(4).2C(2)H(5)OH give flexible metal-organic porous structures M and E, respectively, which have the same stoichiometry, but subtly different structures. This study combines measurements of the thermodynamics and kinetics of carbon dioxide, methanol, and ethanol sorption on adsorbents M and E over a range of temperatures with adsorbent structural characterization at different adsorbate (guest) loadings. The adsorption kinetics for methanol and ethanol adsorption on porous structure E obey a linear driving force (LDF) mass transfer model for adsorption at low surface coverage. The corresponding adsorption kinetics for porous structure M follow a double exponential (DE) model, which is consistent with two different barriers for diffusion through the windows and along the pores in the structure. The former is a high-energy barrier due to the opening of the windows in the structure, required to allow adsorption to occur, while the latter is a lower-energy barrier for diffusion in the pore cavities. X-ray diffraction studies at various methanol and ethanol loadings showed that the host porous structures E and M underwent different scissoring motions, leading to an increase in unit cell volume with the space group remaining unchanged during adsorption. The results are discussed in terms of reversible adsorbate/adsorbent (host/guest) structural changes and the adsorption mechanism involving hydrogen-bonding interactions with specific surface sites for methanol and ethanol adsorption in relation to pore size and extent of filling. This paper contains the first evidence for individual kinetic barriers to diffusion through windows and pore cavities in flexible porous coordination polymer frameworks. PMID- 15291577 TI - Spectroscopic and computational studies of Co2+corrinoids: spectral and electronic properties of the biologically relevant base-on and base-off forms of Co2+cobalamin. AB - Co(2+)cobalmain (Co(2+)Cbl) is implicated in the catalytic cycles of all adenosylcobalamin (AdoCbl)-dependent enzymes, as in each case catalysis is initiated through homolytic cleavage of the cofactor's Co-C bond. The rate of Co C bond homolysis, while slow for the free cofactor, is accelerated by 12 orders of magnitude when AdoCbl is bound to the protein active site, possibly through enzyme-mediated stabilization of the post-homolysis products. As an essential step toward the elucidation of the mechanism of enzymatic Co-C bond activation, we employed electronic absorption (Abs), magnetic circular dichroism (MCD), and resonance Raman spectroscopies to characterize the electronic excited states of Co(2+)Cbl and Co(2+)cobinamide (Co(2+)Cbi(+), a cobalamin derivative that lacks the nucleotide loop and 5,6-dimethylbenzimazole (DMB) base and instead binds a water molecule in the lower axial position). Although relatively modest differences exist between the Abs spectra of these two Co(2+)corrinoid species, MCD data reveal that substitution of the lower axial ligand gives rise to dramatic changes in the low-energy region where Co(2+)-centered ligand field transitions are expected to occur. Our quantitative analysis of these spectral changes within the framework of time-dependent density functional theory (TD-DFT) calculations indicates that corrin-based pi --> pi transitions, which dominate the Co(2+)corrinoid Abs spectra, are essentially insulated from perturbations of the lower ligand environment. Contrastingly, the Co(2+)-centered ligand field transitions, which are observed here for the first time using MCD spectroscopy, are extremely sensitive to alterations in the Co(2+) ligand environment and thus may serve as excellent reporters of enzyme-induced perturbations of the Co(2+) state. The power of this combined spectroscopic/computational methodology for studying Co(2+)corrinoid/enzyme active site interactions is demonstrated by the dramatic changes in the MCD spectrum as Co(2+)Cbi(+) binds to the adenosyltransferase CobA. PMID- 15291579 TI - Cooperative bimetallic reactivity: hydrogen activation in two-electron mixed valence compounds. AB - Reversible dihydrogen uptake by a two-electron mixed-valence di-iridium complex is examined with nonlocal density-functional calculations. Optimized metrics compare favorably with crystal structures of isolated species, and the calculated activation enthalpy of acetonitrile exchange is accurate within experimental error. Dihydrogen attacks the Ir(2) core at Ir(II); the Ir(0) center is electronically saturated and of incorrect orbital parity to interact with H(2). Isomeric eta(2)-H(2) complexes have been located, and harmonic frequency calculations confirm these to be potential energy minima. A transition state links one such complex with the final dihydride; calculated atomic charges suggest a heterolytic H(2) bond scission within the di-iridium coordination sphere. This investigation also establishes a ligand-design criterion for attaining cooperative bimetallic reactivity, namely, that the supporting ligand framework has sufficient mechanical flexibility so that the target complex can accommodate the nuclear reorganizations that accompany substrate activation. PMID- 15291580 TI - Chemical evaluation of hypothetical uninodal zeolites. AB - Optimized structural parameters, framework energies relative to alpha-quartz, and volumes accessible to sorption have been calculated for the systematically enumerated hypothetical uninodal zeolitic structures (structures in which all tetrahedral sites are equivalent). The structures were treated as silica polymorphs, and their energies were minimized using the GULP program with the Sanders-Catlow silica potential. Results are given for 164 structures, which include all 21 known uninodal zeolites, two known minerals (tridymite and cristobalite), and 78 unknown zeolite topologies. Twenty-three hypothetical structures were identified as chemically feasible. Complete structural information is provided, and several structures are discussed in detail. The results will assist in the design of new synthetic routes and in the identification of newly synthesized materials. PMID- 15291581 TI - Heterogeneous or homogeneous catalysis? Mechanistic studies of the rhodium catalyzed dehydrocoupling of amine-borane and phosphine-borane adducts. AB - In depth, comparative studies on the catalytic dehydrocoupling of the amine borane adduct Me(2)NH.BH(3) (to form [Me(2)N-BH(2)](2)) and the phosphine-borane adduct Ph(2)PH.BH(3) (to form Ph(2)PH-BH(2)-PPh(2)-BH(3)) with a variety of Rh (pre)catalysts such as [[Rh(1,5-cod)(micro-Cl)](2)], Rh/Al(2)O(3), Rh(colloid)/[Oct(4)N]Cl, and [Rh(1,5-cod)(2)]OTf have been performed in order to determine whether the dehydrocoupling proceeds by a homogeneous or heterogeneous mechanism. The results obtained suggest that the catalytic dehydrocoupling of Me(2)NH.BH(3) is heterogeneous in nature involving Rh(0) colloids, while that of Ph(2)PH.BH(3) proceeds by a homogeneous mechanism even when starting with Rh(0) precursors such as Rh/Al(2)O(3). The catalytic dehydrocoupling reactions are thought to proceed by different mechanisms due to a combination of factors such as (i) the greater reducing strength of amine-borane adducts, (ii) the increased ease of dissociation of phosphine-borane adducts, and (iii) phosphine ligation and/or poisoning of active catalytic sites on metal colloids. PMID- 15291582 TI - Mechanisms of water oxidation catalyzed by the cis,cis-[(bpy)2Ru(OH2)]2O4+ ion. AB - The cis,cis-[(bpy)(2)Ru(III)(OH(2))](2)O(4+) micro-oxo dimeric coordination complex is an efficient catalyst for water oxidation by strong oxidants that proceeds via intermediary formation of cis,cis-[(bpy)(2)Ru(V)(O)](2)O(4+) (hereafter, [5,5]). Repetitive mass spectrometric measurement of the isotopic distribution of O(2) formed in reactions catalyzed by (18)O-labeled catalyst established the existence of two reaction pathways characterized by products containing either one atom each from a ruthenyl O and solvent H(2)O or both O atoms from solvent molecules. The apparent activation parameters for micro-oxo ion-catalyzed water oxidation by Ce(4+) and for [5,5] decay were nearly identical, with DeltaH(++) = 7.6 (+/-1.2) kcal/mol, DeltaS() = -43 (+/-4) cal/deg mol (23 degrees C) and DeltaH(++) = 7.9 (+/-1.1) kcal/mol, DeltaS(++) = -44 (+/ 4) cal/deg mol, respectively, in 0.5 M CF(3)SO(3)H. An apparent solvent deuterium kinetic isotope effect (KIE) of 1.7 was measured for O(2) evolution at 23 degrees C; the corresponding KIE for [5,5] decay was 1.6. The (32)O(2)/(34)O(2) isotope distribution was also insensitive to solvent deuteration. On the basis of these results and previously established chemical properties of this class of compounds, mechanisms are proposed that feature as critical reaction steps H(2)O addition to the complex to form covalent hydrates. For the first pathway, the elements of H(2)O are added as OH and H to the adjacent terminal ruthenyl O atoms, and for the second pathway, OH is added to a bipyridine ring and H is added to one of the ruthenyl O atoms. PMID- 15291583 TI - Strong near-infrared luminescence in BaSnO3. AB - Powdered samples of the perovskite BaSnO(3) exhibit strong near-infrared (NIR) luminescence at room temperature, following band-gap excitation at 380 nm (3.26 eV). The emission spectrum is characterized by a broad band centered at 905 nm (1.4 eV), tailing on the high-energy side to approximately 760 nm. The Stokes shift is 1.9 eV, and measured lifetimes in the range 7-18 ms depend on preparative conditions. These extraordinary long values indicate that the luminescence involves a defect state(s). At low temperatures, both a sharp peak and a broad band appear in the visible portion of the luminescence spectrum at approximately 595 nm. Upon cooling, the intensity of the NIR emission decreases, while the integrated intensities of the visible emission features increase to approximately 40% of the NIR intensity at 77 K. Room-temperature photoluminescence (PL) is observed across the Ba(1-x)Sr(x)SnO(3) series. As the strontium content increases, the excitation maximum and band gap shift further into the UV, while the intensity of the NIR emission peak decreases and shifts further into the infrared. This combination leads to an unexpectedly large increase in the Stokes shift. The unusual NIR PL in BaSnO(3) may originate from recombination of a photogenerated valence-band hole and an occupied donor level, probably associated with a Sn(2+) ion situated roughly 1.4 eV above the valence band edge. PMID- 15291584 TI - Electron transfer in uranyl(VI)-uranyl(V) complexes in solution. AB - The rates and mechanisms of the electron self-exchange between U(V) and U(VI) in solution have been studied with quantum chemical methods. Both outer-sphere and inner-sphere mechanisms have been investigated; the former for the aqua ions, the latter for binuclear complexes containing hydroxide, fluoride, and carbonate as bridging ligand. The calculated rate constant for the self-exchange reaction UO(2)(+)(aq) + UO(2)(2+)(aq) <=>UO(2)(2+)(aq) + UO(2)(+)(aq), at 25 degrees C, is k = 26 M(-1) s(-1). The lower limit of the rate of electron transfer in the inner sphere complexes is estimated to be in the range 2 x 10(4) to 4 x 10(6) M(-1) s( 1), indicating that the rate for the overall exchange reaction may be determined by the rate of formation and dissociation of the binuclear complex. The activation energy for the outer-sphere model calculated from the Marcus model is nearly the same as that obtained by a direct calculation of the precursor- and transition-state energy. A simple model with one water ligand is shown to recover 60% of the reorganization energy. This finding is important because it indicates the possibility to carry out theoretical studies of electron-transfer reactions involving M(3+) and M(4+) actinide species that have eight or nine water ligands in the first coordination sphere. PMID- 15291585 TI - Complex mechanism of the gas phase reaction between formic acid and hydroxyl radical. Proton coupled electron transfer versus radical hydrogen abstraction mechanisms. AB - The gas phase reaction between formic acid and hydroxyl radical has been investigated with high level quantum mechanical calculations using DFT-B3LYP, MP2, CASSCF, QCISD, and CCSD(T) theoretical approaches in connection with the 6 311+G(2df,2p) and aug-cc-pVTZ basis sets. The reaction has a very complex mechanism involving several elementary processes, which begin with the formation of a reactant complex before the hydrogen abstraction by hydroxyl radical. The results obtained in this investigation explain the unexpected experimental fact that hydroxyl radical extracts predominantly the acidic hydrogen of formic acid. This is due to a mechanism involving a proton coupled electron-transfer process. The calculations show also that the abstraction of formyl hydrogen has an increased contribution at higher temperatures, which is due to a conventional hydrogen abstraction radical type mechanism. The overall rate constant computed at 298 K is 6.24 x 10(-13) cm3 molecules(-1) s(-1), and compares quite well with the range from 3.2 +/- 1 to 4.9 +/- 1.2 x 10(-13) cm3 molecules(-1) s(-1), reported experimentally. PMID- 15291586 TI - Simultaneous recording of spin-state-selective NMR spectra for different InS spin systems. AB - Heteronuclear magnetization transfer occurring during heteronuclear cross polarization mixing processes in liquid-state NMR experiments can be easily monitored as a function of the involved in-phase, antiphase, and multiple-quantum magnetization components. The theoretical background on the simultaneous detection of E.COSY-type, TROSY-type, or spin-edited multiplet patterns for different IS and I(2)S spin systems in the same solution-state NMR spectrum is described. The proposed pulse scheme preserves high sensitivity levels and shows good tolerance to the presence of undesired cross-talk artifacts for both NH and NH(2) multiplicities providing an interesting NMR tool for biomolecular applications. PMID- 15291587 TI - Stereospecific NMR assignments of prochiral methyls, rotameric states and dynamics of valine residues in malate synthase G. AB - Near complete stereospecific assignments of the prochiral methyl carbons of Leu and Val residues in malate synthase G, a 723 residue enzyme, are reported. Assignments were obtained on the basis of a 10% fractional (13)C-labeling strategy developed by Wuthrich and co-workers [Neri, D; Szyperski, T; Otting, G; Senn, H; Wuthrich, K. Biochemistry 1989, 28, 7510-7516] and, in the case of Val residues, supplemented with results from a series of new methyl-TROSY quantitative J experiments for measuring (3)J(C)(gamma)(N) and (3)J(C)(gamma)(C)' couplings. The measured (3)J couplings were also used to probe Val side chain dynamics. A strong correlation is observed between rotamer averaging established on the basis of the couplings and side chain millisecond time scale dynamics measured using methyl-TROSY based (1)H-(13)C multiple quantum relaxation dispersion experiments. PMID- 15291588 TI - Strong interfullerene electronic communication in a bisfullerene-hexarhodium sandwich complex. AB - Reaction of Rh(6)(CO)(12)(dppm)(2) (dppm = 1,2-bis(diphenylphosphino)methane) with 1.4 equiv. of C(60) in chlorobenzene at 120 degrees C affords a face-capping C(60) derivative Rh(6)(CO)(9)(dppm)(2)(micro(3)-eta(2),eta(2),eta(2)-C(60)) (1) in 73% yield. Treatment of 1 with excess CNR (10 equiv., R = CH(2)C(6)H(5)) at 80 degrees C provides a bisbenzylisocyanide-substituted compound Rh(6)(CO)(7)(dppm)(2)(CNR)(2)(micro(3)-eta(2),eta(2),eta(2)-C(60)) (2) in 59% yield. Reaction of 1 with excess C(60) (4 equiv.) in refluxing chlorobenzene followed by treatment with 1 equiv. of CNR at room temperature gives a bisfullerene sandwich complex Rh(6)(CO)(5)(dppm)(2)(CNR)(micro(3) eta(2),eta(2),eta(2)-C(60))(2) (3) in 31% yield. Compounds 1, 2, and 3 have been characterized by spectroscopic and microanalytical methods as well as by X-ray crystallographic studies. Electrochemical properties of 1, 2, and 3 have been examined by cyclic voltammetry. The cyclic voltammograms (CVs) of 1 and 2 show two reversible one-electron redox waves, a reversible one-step two-electron redox wave, and a reversible one-electron redox wave, respectively, within the solvent cutoff window. This observation suggests that compounds 1 and 2 undergo similar C(60)-localized electrochemical pathways up to 1(5)(-) and 2(5)(-). Each redox wave of 2 appears at more negative potentials compared to that of 1 because of the donor effect of the benzylisocyanide ligand. The CV of compound 3 reveals six reversible well-separated redox waves due to strong interfullerene electronic communication via the Rh(6) metal cluster bridge. The electrochemical properties of 1, 2, and 3 have been rationalized by molecular orbital calculations using the density functional theory (DFT) method. In particular, the molecular orbital (MO) calculation reveals significant contribution of the metal cluster center to the unoccupied molecular orbitals in 3, which is consistent with the experimental result of strong interfullerene electronic communication via the Rh(6) metal cluster spacer. PMID- 15291589 TI - High-efficiency, environment-friendly electroluminescent polymers with stable high work function metal as a cathode: green- and yellow-emitting conjugated polyfluorene polyelectrolytes and their neutral precursors. AB - A series of aminoalkyl-substituted polyfluorene copolymers with benzothiadiazole (BTDZ) of different content were synthesized by Suzuki coupling reaction, and their quaternized ammonium polyelectrolyte derivatives were obtained through a postpolymerization treatment on the terminal amino groups. Copolymers are soluble in environmentally friendlier solvents, such as alcohols. It was found that the efficient energy transfer occurs by exciton trapping on the narrow band gap BTDZ site under UV illumination. Only 1% of BTDZ content is needed to completely quench a fluorene emission for both the neutral and the quaternized copolymers in the neat film. Absolute PL efficiencies of copolymer films were greatly enhanced as a result of the suppression of excimer formation. Light-emitting devices fabricated from these copolymers show high external quantum efficiencies over 3% and 1% for the neutral precursor and the quaternized copolymers, respectively, with high work function metals such as Al as a cathode. To the best of our knowledge, this is the first report on an electroluminescent polymer which bears the high EL efficiency, the electron-injection ability from high work function metals, and the solubility in environment-friendly solvents at the same time. These features make them a promising candidate for the next generation of light emitting copolymers in PLED flat panel display application. PMID- 15291590 TI - Ab initio molecular dynamics simulations and g-tensor calculations of aqueous benzosemiquinone radical anion: effects of regular and "T-stacked" hydrogen bonds. AB - Car-Parrinello molecular dynamics (CP-MD) simulations of the benzosemiquinone radical anion in aqueous solution have been performed at ambient conditions. Analysis of the trajectory shows not only extensive hydrogen bonding to the carbonyl oxygen atoms (ca. 4-5.6 water molecules depending on distance criteria), but also relatively long-lived "T-stacked" hydrogen bonds to the semiquinone pi system. These results are discussed in the context of recent findings on semiquinone-protein interactions in photosynthetic reaction centers, and of EPR and vibration spectroscopical data for the aqueous system. Snapshots from the CP MD trajectory are used for the first quantum chemical analyses of dynamical effects on electronic g-tensors, using cluster models and a recently developed density functional method. In particular, the effects of intermolecular hydrogen bond dynamics on the g-tensor components are examined, in comparison with recent EPR and ENDOR studies. PMID- 15291591 TI - Cation-pi interactions of a thiocarbonyl group and a carbonyl group with a pyridinium nucleus. AB - Attractive interactions between a thiocarbonyl group and a pyridinium nucleus, and between a carbonyl group and a pyridinium nucleus have been proven by (1)H and (13)C NMR studies, UV-vis spectral analyses, and X-ray crystallographic analyses of nicotinic amides 1 and 3, and pyridinium salts 2 and 4. Comparison of the Deltadelta values, which are the differences in the chemical shifts with reference compounds 5 or 6, showed that the absolute Deltadelta values of 2 and 4 are much larger than those of 1 and 3. In the UV-vis spectra, the n-->pi absorption of the C=S group of 2a exhibited a significant blue shift in CHCl(3). X-ray crystallographic analysis of 1-4 clearly showed that the C=S group of 2a and the C=O group of 4 are very close to the pyridinium moiety compared to the case of 1 and 3. In addition, the X-ray crystal packing structure of 2a showed the C=S group is sandwiched between two pyridinium rings. These experimental results strongly suggested the existence of attractive (C=S)...Py(+) and (C=O)...Py(+) interactions in solution and in crystal. The optimized geometries of 1 and 2 calculated at the HF/6-311G level are in good agreement with their X ray geometries. MP2/6-311G calculations for the model systems of pyridinium salts 2 and 4 predicted that the electrostatic and induction energies are the major source of the attractive interactions. Since the larger contribution of electrostatic and induction interactions are characteristic features of cation-pi interactions, the (C=S)...Py(+) and (C=O)...Py(+) interactions would be classified as a cation-pi interaction. PMID- 15291594 TI - Optical excitation and absorption spectra of C50Cl10. AB - C50Cl10 [S. Y. Xie et al., Science 304, 699 (2004)] has been synthesized in large quantities enabling the capture of the labile fullerene C50. In this Communication, we report ab initio calculations on the optical excitation and absorption spectra of C50Cl10. We successfully explain and assign the measured UV visible absorption spectrum of C50Cl10. The first singlet excitation for C50Cl10 is optically forbidden, and its optical absorption gap is redshifted by 0.6 eV (110 nm) relative to that of C60. We demonstrate that passivating C50 with 10 hydrogen atoms and replacing one Cl in C50Cl10 by one methoxy group lead to 100 nm blueshift and 90 nm redshift of the optical gap predicted for C50Cl10, respectively, suggesting C50 derivatives are suitable for tunable optical applications. PMID- 15291595 TI - Theory of aging in structural glasses. AB - The random first-order transition theory of the dynamics of supercooled liquids is extended to treat aging phenomena in nonequilibrium structural glasses. A reformulation of the idea of "entropic droplets" in terms of libraries of local energy landscapes is introduced which treats in a uniform way the supercooled liquid (reproducing earlier results) and glassy regimes. The resulting microscopic theory of aging makes contact with the Nayaranaswamy-Moynihan-Tool nonlinear relaxation formalism and the Hodge-Scherer extrapolation of the Adam Gibbs formula, but deviations from both approaches are predicted and shown to be consistent with experiment. The nonlinearity of glassy relaxation is shown to quantitatively correlate with liquid fragility. The residual non-Arrhenius temperature dependence of relaxation observed in quenched glasses is explained. The broadening of relaxation spectra in the nonequilibrium glass with decreasing temperature is quantitatively predicted. The theory leads to the prediction of spatially fluctuating fictive temperatures in the long-aged glassy state, which have non-Gaussian statistics. This can give rise to "ultraslow" relaxations in systems after deep quenches. PMID- 15291596 TI - Multiresolution quantum chemistry in multiwavelet bases: Analytic derivatives for Hartree-Fock and density functional theory. AB - An efficient and accurate analytic gradient method is presented for Hartree-Fock and density functional calculations using multiresolution analysis in multiwavelet bases. The derivative is efficiently computed as an inner product between compressed forms of the density and the differentiated nuclear potential through the Hellmann-Feynman theorem. A smoothed nuclear potential is directly differentiated, and the smoothing parameter required for a given accuracy is empirically determined from calculations on six homonuclear diatomic molecules. The derivatives of N2 molecule are shown using multiresolution calculation for various accuracies with comparison to correlation consistent Gaussian-type basis sets. The optimized geometries of several molecules are presented using Hartree Fock and density functional theory. A highly precise Hartree-Fock optimization for the H2O molecule produced six digits for the geometric parameters. PMID- 15291597 TI - The quasi-independent curvilinear coordinate approximation for geometry optimization. AB - This paper presents an efficient alternative to well established algorithms for molecular geometry optimization. This approach exploits the approximate decoupling of molecular energetics in a curvilinear internal coordinate system, allowing separation of the 3N-dimensional optimization problem into an O(N) set of quasi-independent one-dimensional problems. Each uncoupled optimization is developed by a weighted least squares fit of energy gradients in the internal coordinate system followed by extrapolation. In construction of the weights, only an implicit dependence on topologically connected internal coordinates is present. This new approach is competitive with the best internal coordinate geometry optimization algorithms in the literature and works well for large biological problems with complicated hydrogen bond networks and ligand binding motifs. PMID- 15291598 TI - Revisiting infinite lattice sums with the periodic fast multipole method. AB - The evaluation of lattice sums as well as stress lattice sums encountered in the periodic fast multipole method is reinvestigated. Simple, accurate, and efficient recurrence expressions for such sums following the ideas of the renormalization method are derived. The first few nonzero lattice sum terms in a three dimensional cubic lattice are computed and given in Tables. The practical considerations accompanying the computation of the sums such as convergence and accuracy are discussed. PMID- 15291599 TI - Effective potential analytic continuation calculations of real time quantum correlation functions: asymmetric systems. AB - We apply the effective potential analytic continuation (EPAC) method to one dimensional asymmetric potential systems to obtain the real time quantum correlation functions at various temperatures. Comparing the EPAC results with the exact results, we find that for an asymmetric anharmonic oscillator the EPAC results are in very good agreement with the exact ones at low temperature, while this agreement becomes worse as the temperature increases. We also show that the EPAC calculation for a certain type of asymmetric potentials can be reduced to that for the corresponding symmetric potentials. PMID- 15291600 TI - Nonexponential statistics of fluorescence photobleaching. AB - In this paper, I consider theoretical models of the decay via photobleaching of a sample of surface-immobilized fluorescent molecules excited by a spatially varying laser intensity profile. I show that, with mild restrictions on the photobleaching mechanism, the fluorescence decay measured in a nonuniform excitation profile is always nonexponential. Under the same conditions, the fluorescence decay can always be approximated by a discrete sum of exponentials. A particular example is given in which a homogeneous population of fluorophores with a single (intensity-dependent) photobleaching lifetime, when illuminated by a Gaussian laser, exhibits power law fluorescence decay at long times. These results indicate that the observation of multiple exponentials in single molecule or ensemble photobleaching lifetime measurements can arise solely as an artifact of a spatially varying laser profile and is not necessarily indicative of heterogeneity in molecular internal states, conformation, or local environment. PMID- 15291601 TI - Overcoming free energy barriers using unconstrained molecular dynamics simulations. AB - Association of unconstrained molecular dynamics (MD) and the formalisms of thermodynamic integration and average force [Darve and Pohorille, J. Chem. Phys. 115, 9169 (2001)] have been employed to determine potentials of mean force. When implemented in a general MD code, the additional computational effort, compared to other standard, unconstrained simulations, is marginal. The force acting along a chosen reaction coordinate xi is estimated from the individual forces exerted on the chemical system and accumulated as the simulation progresses. The estimated free energy derivative computed for small intervals of xi is canceled by an adaptive bias to overcome the barriers of the free energy landscape. Evolution of the system along the reaction coordinate is, thus, limited by its sole self-diffusion properties. The illustrative examples of the reversible unfolding of deca-L-alanine, the association of acetate and guanidinium ions in water, the dimerization of methane in water, and its transfer across the water liquid-vapor interface are examined to probe the efficiency of the method. PMID- 15291602 TI - Linear-scaling formation of Kohn-Sham Hamiltonian: application to the calculation of excitation energies and polarizabilities of large molecular systems. AB - We present calculations of excitation energies and polarizabilities in large molecular systems at the local-density and generalized-gradient approximation levels of density-functional theory (DFT). Our results are obtained using a linear-scaling DFT implementation in the program system DALTON for the formation of the Kohn-Sham Hamiltonian. For the Coulomb contribution, we introduce a modification of the fast multipole method to calculations over Gaussian charge distributions. It affords a simpler implementation than the original continuous fast multipole method by partitioning the electrostatic Coulomb interactions into "classical" and "nonclassical" terms which are explicitly evaluated by linear scaling multipole techniques and a modified two-electron integral code, respectively. As an illustration of the code, we have studied the singlet and triplet excitation energies as well as the static and dynamic polarizabilities of polyethylenes, polyenes, polyynes, and graphite sheets with an emphasis on the trends observed with system size. PMID- 15291603 TI - Thermodynamic length in a two-dimensional thermodynamic state space. AB - The main goal of this paper is to reach an explicit formulation and possible interpretation of thermodynamic length in a thermodynamic state space with two degrees of freedom. Using the energy and entropy metric in a general form, we get explicit results about thermodynamic length along isotherms, its relation with work and with speed of sound. We also look at the relation between the determinants of the energy and entropy metrics and find that they differ by a factor of T4. PMID- 15291604 TI - Contracted basis Lanczos methods for computing numerically exact rovibrational levels of methane. AB - We present a numerically exact calculation of rovibrational levels of a five-atom molecule. Two contracted basis Lanczos strategies are proposed. The first and preferred strategy is a two-stage contraction. Products of eigenfunctions of a four-dimensional (4D) stretch problem and eigenfunctions of 5D bend-rotation problems, one for each K, are used as basis functions for computing eigenfunctions and eigenvalues (for each K) of the Hamiltonian without the Coriolis coupling term, denoted H0. Finally, energy levels of the full Hamiltonian are calculated in a basis of the eigenfunctions of H0. The second strategy is a one-stage contraction in which energy levels of the full Hamiltonian are computed in the product contracted basis (without first computing eigenfunctions of H0). The two-stage contraction strategy, albeit more complicated, has the crucial advantage that it is trivial to parallelize the calculation so that the CPU and memory costs are independent of J. For the one stage contraction strategy the CPU and memory costs of the difficult part of the calculation scale linearly with J. We use the polar coordinates associated with orthogonal Radau vectors and spherical harmonic type rovibrational basis functions. A parity-adapted rovibrational basis suitable for a five-atom molecule is proposed and employed to obtain bend-rotation eigenfunctions in the first step of both contraction methods. The effectiveness of the two methods is demonstrated by calculating a large number of converged J = 1 rovibrational levels of methane using a global potential energy surface. PMID- 15291605 TI - Incoherent noise and quantum information processing. AB - Incoherence in the controlled Hamiltonian is an important limitation on the precision of coherent control in quantum information processing. Incoherence can typically be modeled as a distribution of unitary processes arising from slowly varying experimental parameters. We show how it introduces artifacts in quantum process tomography and we explain how the resulting estimate of the superoperator may not be completely positive. We then go on to attack the inverse problem of extracting an effective distribution of unitaries that characterizes the incoherence via a perturbation theory analysis of the superoperator eigenvalue spectra. PMID- 15291606 TI - Ab initio calculations on low-lying electronic states of TeO2 and Franck-Condon simulation of the (1)1B2 <-- X1A1 TeO2 absorption spectrum including anharmonicity. AB - Ab initio calculations have been carried out on low-lying singlet and triplet states of TeO2 at different levels of theory with basis sets of up to the augmented-polarized valence-quintuple-zeta quality. Equilibrium geometrical parameters, harmonic vibrational frequencies, and relative electronic energies of the X1A1, 1B1, 1B2, 1A2, 3A1, 3B1, 3B2, and 3A2 states of TeO2 have been calculated. Potential energy functions (PEFs) of the X1A1 and the (1)1B2 states were computed at the complete-active-space self-consistent-field multireference configuration interaction level, with a basis set of augmented-polarized valence quadruple-zeta quality. Franck-Condon factors (FCFs) for the electronic transition between the X1A1 and (1)1B2 states of TeO2 were calculated with the above-mentioned ab initio PEFs. The (1)1B2 <-- X1A1 absorption spectrum of TeO2 was simulated employing the computed FCFs, which include Duschinsky rotation and anharmonicity, and compared with the recently published laser-induced fluorescence (LIF) spectrum of Hullah and Brown [J. Mol. Spectrosc. 200, 261 (2000)]. The ab initio results and spectral simulation reported here confirm the upper electronic state involved in the LIF spectrum to be the (1)1B2 state of TeO2 and also confirm the vibrational assignments of Hullah and Brown. However, our simulated spectrum suggests that the reported LIF spectrum from 345 to 406 nm represents only a portion of the full (1)1B2 <-- X1A1 absorption spectrum of TeO2, which extends from ca. 406 to 300 nm. Another dye other than the two used by Hullah and Brown is required to cover the 345-300 nm region of the LIF band. Ab initio calculations show strong configuration mixing of the (1)1B2 electronic surface with higher 1B2 states in a region of large TeO bond length (> or = 2.0 A) and OTeO bond angle (> or = 135.0 degrees). PMID- 15291607 TI - Raman spectra of (He)N-Br2(X) clusters: the role of boson/fermion statistics in a quantum solvent. AB - The aim of this paper is to elucidate the role played by the bosonic/fermionic character of N He atoms solvating a Br2(X) molecule. To this end, an adiabatic model in the molecular stretching coordinate is assumed and the ground energy levels of the complexes are searched by means of Hartree (or Hartree-Fock) Quantum Chemistry calculations for 4He (or 3He) solvent atoms. Simulations of vib rotational Raman spectra point at the spin multiplicity as the main feature responsible for the drastic difference in the rotational structures of molecules embedded in boson or fermion helium drops as already observed by the experiments of Grebenev et al. [S. Grebenev, J. P. Toennies, and A. F. Vilesov, Science 279 (1998) 2083]. PMID- 15291608 TI - Franck-Condon effects in collision-induced electronic energy transfer: I2(E; v = 1,2) + He, Ar. AB - Collisions of I2 in the E electronic state with rare gas atoms result in electronic energy transfer to the D, beta, and D' ion-pair electronic states. Rate constants for each of these channels have been measured when I2 is initially prepared in the J = 55, nu = 1 and 2 levels in the E state. The rate constants and effective hard sphere collision cross sections confirm the trends observed when nu = 0 in the E state is initially prepared: He collisions favor population of the D state, while Ar collisions favor population of the beta state. Final state vibrational level distributions are determined by spectral simulation and are found to be qualitatively consistent with the trends in the Franck-Condon factors. The experimental distributions are also compared to the recent quantum scattering calculations of Tscherbul and Buchachenko. PMID- 15291609 TI - Direct potential fit analysis of the X1sigma+ ground state of CO. AB - A collection of 21,559 highly precise spectroscopic line positions from pure rotational and vibration-rotational spectra for seven isotopomers of carbon monoxide in the X1sigma+ ground electronic state has been employed in direct least-squares fits of the rovibrational Hamiltonian operator obtained from Watson's work [J. Mol. Spectrosc. 80, 411 (1980)] and that obtained by Herman and Ogilvie [Adv. Chem. Phys 103, 187 (1998)]. Fully analytical models are used for the various functions, including the Born-Oppenheimer internuclear potential function, and an account is taken of breakdown of the Born-Oppenheimer approximation. The resulting representations are more compact than currently available traditional Ukl/deltakl extended Dunham descriptions, and they generate quantum-mechanical eigenvalues that reproduce reliably the spectroscopic line positions to within the associated measurement uncertainties. Rayleigh Schrodinger perturbation theory has been used to calculate highly accurate rotational and centrifugal distortion constants Bupsilon-Oupsilon for nine isotopomers of carbon monoxide. These constants are just as successful at reconstructing the observed spectroscopic information as the quantum-mechanical eigenvalues of the fitted Hamiltonian operators. PMID- 15291610 TI - Infrared spectrum and stability of a pi-type hydrogen-bonded complex between the OH and C2H2 reactants. AB - A hydrogen-bonded complex between the hydroxyl radical and acetylene has been stabilized in the reactant channel well leading to the addition reaction and characterized by infrared action spectroscopy in the OH overtone region. Analysis of the rotational band structure associated with the a-type transition observed at 6885.53(1) cm(-1) (origin) reveals a T-shaped structure with a 3.327(5) A separation between the centers of mass of the monomer constituents. The OH (v = 1) product states populated following vibrational predissociation show that dissociation proceeds by two mechanisms: intramolecular vibrational to rotational energy transfer and intermolecular vibrational energy transfer. The highest observed OH product state establishes an upper limit of 956 cm(-1) for the stability of the pi-type hydrogen-bonded complex. The experimental results are in good accord with the intermolecular distance and well depth at the T-shaped minimum energy configuration obtained from complementary ab initio calculations, which were carried out at the restricted coupled cluster singles, doubles, noniterative triples level of theory with extrapolation to the complete basis set limit. PMID- 15291611 TI - Spectroscopic implications of the coupling of unquenched angular momentum to rotation in OH-containing complexes. AB - A model is developed for the rotational energy levels and electric dipole transition intensities of nonlinear OH-containing complexes in which the OH is hydrogen bonded to its partner. Both the 2A' and 2A" electronic states arising from the lifting of the OH monomer electronic orbital degeneracy are explicitly included. Consequently, the model smoothly spans the entire range of the difference potential associated with the separation between these two states, and the model accounts for the partial quenching of the OH monomer electronic angular momentum in such complexes. The more familiar cases of completely unquenched and completely quenched electronic angular momentum are recovered in the limits of zero and very large difference potential, respectively. The sensitivity of rovibrational spectra to the value of the difference potential is investigated, and it is shown that spectra of reactant complexes reveal the extent of quenching, which must occur along the reaction coordinate as the system evolves from weakly interacting partners to addition product. The model is successfully applied to the analysis of the OH overtone spectrum of the OH-acetylene complex. PMID- 15291612 TI - Electron and nuclear dynamics of molecular clusters in ultraintense laser fields. III. Coulomb explosion of deuterium clusters. AB - In this paper we present a theoretical and computational study of the energetics and temporal dynamics of Coulomb explosion of molecular clusters of deuterium (D2)n/2 (n = 480 - 7.6 x 10(4), cluster radius R0 = 13.1 - 70 A) in ultraintense laser fields (laser peak intensity I = 10(15) - 10(20)W cm(-2)). The energetics of Coulomb explosion was inferred from the dependence of the maximal energy EM and the average energy Eav of the product D+ ions on the laser intensity, the laser pulse shape, the cluster radius, and the laser frequency. Electron dynamics of outer cluster ionization and nuclear dynamics of Coulomb explosion were investigated by molecular dynamics simulations. Several distinct laser pulse shape envelopes, involving a rectangular field, a Gaussian field, and a truncated Gaussian field, were employed to determine the validity range of the cluster vertical ionization (CVI) approximation. The CVI predicts that Eav, EM proportional to R0(2) and that the energy distribution is P(E) proportional to E1/2. For a rectangular laser pulse the CVI conditions are satisfied when complete outer ionization is obtained, with the outer ionization time toi being shorter than both the pulse width and the cluster radius doubling time tau2. By increasing toi, due to the increase of R0 or the decrease of I, we have shown that the deviation of Eav from the corresponding CVI value (Eav(CVI)) is (Eav(CVI) - Eav)/Eav(CVI) approximately (toi/2.91tau2)2. The Gaussian pulses trigger outer ionization induced by adiabatic following of the laser field and of the cluster size, providing a pseudo-CVI behavior at sufficiently large laser fields. The energetics manifest the existence of a finite range of CVI size dependence, with the validity range for the applicability of the CVI being R0 < or = (R0)I, with (R0)I representing an intensity dependent boundary radius. Relating electron dynamics of outer ionization to nuclear dynamics for Coulomb explosion induced by a Gaussian pulse, the boundary radius (R0)I and the corresponding ion average energy (Eav)I were inferred from simulations and described in terms of an electrostatic model. Two independent estimates of (R0)I, which involve the cluster size where the CVI relation breaks down and the cluster size for the attainment of complete outer ionization, are in good agreement with each other, as well as with the electrostatic model for cluster barrier suppression. The relation (Eav)I proportional to (R0)I(2) provides the validity range of the pseudo-CVI domain for the cluster sizes and laser intensities, where the energetics of D+ ions produced by Coulomb explosion of (D)n clusters is optimized. The currently available experimental data [Madison et al., Phys. Plasmas 11, 1 (2004)] for the energetics of Coulomb explosion of (D)n clusters (Eav = 5 - 7 keV at I = 2 x 10(18) W cm(-2)), together with our simulation data, lead to the estimates of R0 = 51 - 60 A, which exceed the experimental estimate of R0 = 45 A. The predicted anisotropy of the D+ ion energies in the Coulomb explosion at I = 10(18) W cm(-2) is in accord with experiment. We also explored the laser frequency dependence of the energetics of Coulomb explosion in the range nu = 0.1 - 2.1 fs(-1) (lambda = 3000 - 140 nm), which can be rationalized in terms of the electrostatic model. PMID- 15291613 TI - Kinetic temperature effects on 4He dimers in jets. AB - The formation of dimers in a free jet cryogenic expansion of 4He gas has been studied by measuring mole fractions as a function of source temperature and pressure using diffraction from a nanostructured transmission grating. The data sets are limited to low source pressures for which dimers and trimers are the only appreciable cluster populations in the beam. The final cluster mole fractions are corrected for residual gas attenuation in the source chamber by an extrapolation over several residual gas pressures. A set of rate equations used to model the cluster formation in a free jet expansion has been extended to include the departure of the ambient translational temperature from the isentropic-equilibrium values as the density decreases with increasing distance. The effect of collisions in restoring the equilibrium temperature is treated with a relaxation time approximation. There are distinct distance ranges where the dimer and trimer mole fractions and the ambient temperature near their asymptotic values. The present modeling reproduces the apparent threshold observed at low source pressures for the survival of dimers in the asymptotic beam. Except for these low source pressures, there are only small changes relative to results based on the isentropic temperature. PMID- 15291614 TI - Electric field effects on the shielding constants of noble gases: a four component relativistic Hartree-Fock study. AB - Second derivatives of nuclear shielding constants with respect to an electric field, i.e., shielding polarizabilities, have been calculated for the noble gas atoms from helium to xenon. The calculations have been carried out using the four component relativistic Hartree-Fock method. In order to assess the importance of the individual relativistic corrections, the shielding polarizabilities have also been calculated at the nonrelativistic Hartree-Fock level, with spin-orbit and scalar (Darwin and mass-velocity) effects having been established by perturbative methods. Electron correlation effects have been estimated using the second-order polarization propagator approach. The relativistic effects on the tensor components of the shielding polarizabilities are found to be larger and changing less regularly with the atomic number than for the shielding constant itself. However, there is a partial cancellation of the contributions to the parallel and perpendicular components of the shielding polarizability and as a consequence the mean shielding polarizability is far less affected than the individual components. PMID- 15291615 TI - Predissociation and autoionization of triplet Rydberg states in molecular hydrogen. AB - We present single-photon spectroscopy in molecular hydrogen starting from the metastable c3Piu- state to a number of triplet nd-Rydberg states (v = 0 - 4, n = 12 - 20). Using fast beam spectroscopy both the autoionization channel and the predissociation channel are quantified, field free, as well as with small electric fields. Coupling with the i3Pig state is assumed to be responsible for field-free predissociation of the v = 0 Rydberg levels. The stronger observed predissociation channel of the v = 1 Rydberg levels is due to the nonadiabatic interaction with the h3Sigmag+ state in combination with l mixing due to an external electric field. No direct evidence is found for possible electric field induced predissociation of the gerade Rydberg states by low lying ungerade states. The competition between autoionization and predissociation is discussed in terms of possible consequences for dissociative recombination involving low energy electron collisions with the H2+ molecular ion. PMID- 15291616 TI - Channel switching effect in photodissociating N2O+ ion at 312.5 nm. AB - A experimental observation is presented on the N2O+ photodissociation process, which exhibits a complete channel switching effect in a narrow energy range. The N2O+ ions, prepared at the X2Pi (000) state by (3+1) multiphoton ionization of neutral N2O molecules at 360.6 nm, were excited to different vibrational levels in the A2Sigma+ state in a wavelength range of 275-328 nm. Based on the estimates of total released kinetic energies from the time-of-flight mass spectrum, it was found that the dissociation pathway of N2O+ (A2Sigma+), NO+ (X1Sigma+) + N(4S) with lower dissociation limit, changes abruptly and completely to NO+ (X1Sigma+) + N(2D) with higher dissociation limit, in a excitation energy range of merely 250 cm(-1) at lambda approximately 312.5 nm. This phenomenon was explained by competition between the two dissociation pathways across the special excitation energy region. PMID- 15291617 TI - Collision-energy-resolved Penning ionization electron spectroscopy of bromomethanes (CH3Br, CH2Br2, and CHBr3) by collision with He*(2(3)S) metastable atoms. AB - Ionization of bromomethanes (CH3Br, CH2Br2, and CHBr3) upon collision with metastable He*(2(3)S) atoms has been studied by means of collision-energy resolved Penning ionization electron spectroscopy. Lone-pair (nBr) orbitals of Br4p characters have larger ionization cross sections than sigma(C-Br) orbitals. The collision-energy dependence of the partial ionization cross sections shows that the interaction potential between the molecule and the He*(2(3)S) atom is highly anisotropic around CH3Br or CH2Br2, while isotropic attractive interactions are found for CHBr3. Bands observed at electron energies of approximately 2 eV in the He*(2(3)S) Penning ionization electron spectra (PIES) of CH2Br2 and CHBr3 have no counterpart in ultraviolet (He I) photoionization spectra and theoretical (third-order algebraic diagrammatic construction) one electron and shake-up ionization spectra. Energy analysis of the processes involved demonstrates that these bands and further bands overlapping with sigma(C Br) or piCH2 levels are related to autoionization of dissociating (He+ - Br-) pairs. Similarly, a band at an electron energy of approximately 1 eV in the He*(2(3)S) PIES spectra of CH3Br has been ascribed to autoionizing Br** atoms released by dissociation of (unidentified) excited states of the target molecule. A further autoionization (S) band can be discerned at approximately 1 eV below the lone-pair nBr bands in the He*(2(3)S) PIES spectrum of CHBr3. This band has been ascribed to the decay of autoionizing Rydberg states of the target molecule (M**) into vibrationally excited states of the molecular ion. It was found that for this transition, the interaction potential that prevails in the entrance channel is merely attractive. PMID- 15291618 TI - Infrared spectra of seeded hydrogen clusters: (paraH2)N - OCS, (orthoH2)N - OCS, and (HD)N - OCS, N = 2 - 7. AB - Infrared spectra of hydrogen-carbonyl sulfide clusters containing paraH2, orthoH2, or HD have been studied in the 2060 cm(-1) region of the C-O stretching vibration. The clusters were formed in pulsed supersonic jet expansions and probed using a tunable infrared diode laser spectrometer. Simple symmetric rotor type spectra were observed and assigned for clusters containing up to N = 7 hydrogen molecules. There was no resolved K structure, and Q-branch features were present for orthoH2 and HD but absent for paraH2. These characteristics can be rationalized in terms of near symmetric rotor structures, very low effective rotational temperatures (0.15 to 0.6 K), and nuclear spin statistics. The observed vibrational shifts were compared with those from recent observations on the same clusters embedded in helium nanodroplets. The observed rotational constants for the paraH2 clusters are in good agreement with a recent quantum Monte Carlo simulation. Some mixed clusters were also observed, such as HD-HD-He OCS and paraH2 - orthoH2 - OCS. PMID- 15291619 TI - Assessment of recently developed exchange-correlation functionals for the description of torsion potentials in pi-conjugated molecules. AB - Newly developed exchange-correlation functionals in density functional theory (DFT) have been applied to describe conjugation effects in organic molecules. The performance of the various approaches is assessed through the calculation of torsion energy profiles and their critical comparison with available experimental data. Our results indicate that the OPTX-B95 exchange-correlation functional as well as its corresponding hybrid versions perform better than the well established BLYP or B3LYP schemes when dealing with pi-conjugated molecules. In contrast, the recently introduced VSXC functional is not as reliable as other DFT methods for the systems examined here. PMID- 15291620 TI - Evaporatively cooled M+ (H2O)Ar cluster ions: infrared spectroscopy and internal energy simulations. AB - Rotationally resolved IR spectra of M+ (H2O)Ar cluster ions for M=Na, K, and Cs in the O-H stretch region were measured in a triple-quadrupole mass spectrometer. Analysis of the spectra yields O-H stretch vibrational band origins and relative IR intensities of the symmetric and asymmetric modes. The effect of the alkali metal ions on these modes results in frequency shifts and intensity changes from the gas phase values of water. The A-rotational constants are also obtained from the rotational structure and are discussed. Experimentally, the temperatures of these species were deduced from the relative populations of the K-rotational states. The internal energies and temperatures of the cluster ions for Na and K were simulated using RRKM calculations and the evaporative ensemble formalism. With binding energies and vibrational frequencies obtained from ab initio calculations, the average predicted temperatures are qualitatively consistent with the experimental values and demonstrate the additional cooling resulting from argon evaporation. PMID- 15291621 TI - Insights into the structures, energetics, and vibrations of aqua-rubidium(I) complexes: ab initio study. AB - We have carried out ab initio and density functional theory calculations of hydrated rubidium cations. The calculations involve a detailed evaluation of the structures, thermodynamic properties, and IR spectra of several plausible conformers of Rb+ (H2O)(n=1-8) clusters. An extensive search was made to find out the most stable conformers. Since the water-water interactions are important in hydrated Rb+ complexes, we investigated the vibrational frequency shifts of the OH stretching modes depending on the number of water molecules and the presence/absence of outer-shell water molecules. The predicted harmonic and anharmonic vibrational frequencies of the aqua-Rb+ clusters reflect the H-bonding signature, and would be used in experimental identification of the hydrated structures of Rb+ cation. PMID- 15291622 TI - Ab initio potential energy surfaces of the ion-molecule reaction: C2H2 + O+. AB - High level ab initio calculations using complete active space self-consistent field and multi reference single and double excitation configuration interaction methods with cc-pVDZ (correlation consistent polarized valence double zeta) and cc-pVTZ (triple zeta) basis sets have been performed to elucidate the reaction mechanism of the ion-molecule reaction, C2H2(1Sigmag+) + O+(4S), for which collision experiment has been performed by Chiu et al. [J. Chem. Phys. 109, 5300 (1998)]. The minor low-energy process leading to the weak spin-forbidden product C2H2+ (2Piu) + O(1D) has been studied previously and will not be discussed here. The major pathways to form charge-transfer (CT) products, C2H2+ (2Piu) + O(3P) (CT1) and C2H2+ (4A2) + O(3P) (CT2), and the covalently bound intermediates are investigated. The approach of the oxygen atom cation to acetylene goes over an energy barrier TS1 of 29 kcal/mol (relative to the reactant) and adiabatically leads the CT2 product or a weakly bound intermediate Int1 between CT2 products. This transition state TS1 is caused by the avoided crossing between the reactant and CT2 electronic states. As the C-O distance becomes shorter beyond the above intermediate, the C1 reaction pathway is energetically more favorable than the Cs pathway and goes over the second transition state TS2 of a relative energy of 39 kcal/mol. Although this TS connects diabatically to the covalent intermediate Int2, there are many states that interact adiabatically with this diabatic state and these lead to the other charge-transfer product CT1 via either of several nonadiabatic transitions. These findings are consistent with the experiment, in which charge transfer and chemical reaction products are detected above 35 and 39 kcal/mol collision energies, respectively. PMID- 15291623 TI - Structures of HCN-Mgn (n=2-6) complexes from rotationally resolved vibrational spectroscopy and ab initio theory. AB - High-resolution infrared laser spectroscopy has been used to determine the structures of HCN-Mgn complexes formed in helium nanodroplets. The magnesium atoms are first added to the droplets to ensure that the magnesium complexes are preformed before the HCN molecule is added. The vibrational frequencies, structures, and dipole moments of these complexes are found to vary dramatically with cluster size, illustrating the nonadditive nature of the HCN-magnesium interactions. All of the complexes discussed here have the nitrogen end of the HCN pointing towards the magnesium clusters. For Mg3, the HCN binds to the "threefold" site, yielding a symmetric top spectrum. Although the HCN-Mg4 complex also has C3v symmetry, the HCN sits "on-top" of a single magnesium atom. These structures are confirmed by both ab initio calculations and measurements of the dipole moments. Significant charge transfer is observed in the case of HCN-Mg4, indicative of charge donation from the lone pair on the nitrogen of HCN into the lowest unoccupied molecular orbital of the Mg4. PMID- 15291624 TI - Solvation effects in near-critical binary mixtures. AB - A Ginzburg-Landau theory is presented to investigate solvation effects in near critical polar fluid binary mixtures. Concentration dependence of the dielectric constant gives rise to a shell region around a charged particle within which solvation occurs preferentially. As the critical point is approached, the concentration has a long-range Ornstein-Zernike tail representing strong critical electrostriction. If salt is added, strong coupling arises among the critical fluctuations and the ions. The structure factors of the critical fluctuations and the charge density are calculated and the phase transition behavior is discussed. PMID- 15291625 TI - Resonant enhancement of two-photon absorption in substituted fluorene molecules. AB - The degenerate and nondegenerate two-photon absorption (2PA) spectra for a symmetric and an asymmetric fluorene derivative were experimentally measured in order to determine the effect of intermediate state resonance enhancement (ISRE) on the 2PA cross section delta. The ability to tune the individual photon energies in the nondegenerate 2PA (ND-2PA) process afforded a quantitative study of the ISRE without modifying the chemical structure of the investigated chromophores. Both molecules exhibited resonant enhancement of the nonlinearity with the asymmetric compound showing as much as a twentyfold increase in delta. Furthermore, the possibility of achieving over a one order of magnitude enhancement of the nonlinearity reveals the potential benefits of utilizing ND 2PA for certain applications. To model ISRE, we have used correlated quantum chemical methods together with the perturbative sum-over-states (SOS) expression. We find strong qualitative and quantitative correlation between the experimental and theoretical results. Finally, using a simplified three-level model for the SOS expression, we provide intuitive insight into the process of ISRE for ND-2PA. PMID- 15291626 TI - The glass-to-liquid transition of the emulsified high-density amorphous ice made by pressure-induced amorphization. AB - Emulsified high-density amorphous ice, made by pressure-induced amorphization of emulsified ice Ih, was decompressed at about 160 K. The onset of an endothermic event was observed around 0.4 GPa during the decompression. This is consistent with existence of the glass transition to a liquid state, implying the close relationship between melting and amorphization. PMID- 15291627 TI - Characterization of heteronuclear decoupling through proton spin dynamics in solid-state nuclear magnetic resonance spectroscopy. AB - The work presented here aims at understanding the performance of phase modulated heteronuclear decoupling sequences such as Cosine Modulation or Two Pulse Phase Modulation. To that end we provide an analytical description of the intrinsic behavior of Cosine Modulation decoupling with respect to radio-frequency inhomogeneity and the proton-proton dipolar coupling network. We discover through a Modulation Frame average Hamiltonian analysis that best decoupling is obtained under conditions where the heteronuclear interactions are removed but notably where homonuclear couplings are recoupled at a homonuclear Rotary Resonance (HORROR) condition in the Modulation Frame. These conclusions are supported by extensive experimental investigations, and notably through the introduction of proton nutation experiments to characterize spin dynamics in solids under decoupling conditions. The theoretical framework presented in this paper allows the prediction of the optimum parameters for a given set of experimental conditions. PMID- 15291628 TI - Phase behavior and critical properties of size-asymmetric, primitive-model electrolytes. AB - The theory of J. Jiang et al. [J. Chem. Phys. 116, 7977 (2002)] for size symmetric electrolytes is extended to size-asymmetric electrolytes. When compared to molecular-simulation results, this extension gives the correct trend of critical properties with size asymmetry. PMID- 15291629 TI - The effect of the cation substitution on the structural and vibrational properties of Cs2NaGaxSc(1-x)F6 solid solution. AB - Raman scattering and x-ray diffration were used to characterize the structural and vibrational properties of the Cs2NaGaxSc(1-x)F6 solid solutions, for x ranging from 0.0 to 1.0. The Raman spectra, taken at room and low temperature, allow us to follow the phase evolution in detail and indicate the breaking of the local symmetry since low Ga concentration levels. Five compositions were studied by x-ray diffraction: x = 0.0, 0.2, 0.5, 0.8, and 1.0. A cubic space group, Fm3m, was found to x = 0.0 and x = 0.2 and a trigonal one was found to x = 0.5, 0.8, and 1.0. Details of both phases are presented and the correlation between x-ray diffraction and Raman scattering is discussed. PMID- 15291630 TI - A mean field analysis of the static dielectric behavior of linear lower alcohols. AB - The static dielectric responses of methanol, ethanol, and 1-propanol up to 1 hexanol are discussed in terms of a stiff-chain lattice model for the alcohol clusters. An analytical expression for the Kirkwood correlation factor gK is derived in terms of the canonical partition function associated to the configurational statistics of any of the dimers building up a chain. This allows for the estimate of the dipole moment mu0 of an alcohol molecule in the liquid phase from the temperature dependence of the dielectric constant. All alcohol species appear to be characterized by a dipole moment larger than in the vapor phase. The Kirkwood correlation factor is found to be an increasing function of the alkyl tail length. PMID- 15291631 TI - Modeling velocity autocorrelation functions for confined fluids using gamma distributions. AB - We propose a model for the short-time dynamics of fluids confined in slit-shaped pores. The model has been developed from the observation that the real lobe of the instantaneous normal mode density of states (INM DOS) closely follows a gamma distribution. By proposing that the density of states of the confined fluid can be represented by a gamma distribution, the resulting velocity autocorrelation function (VACF) is constructed such that it is accurate upto the fourth frequency moment. The proposed model results in an analytical expression for the VACF and relaxation times. The VACFs obtained from the model have been compared with the VACFs obtained from molecular dynamic simulations and INM analysis for fluids confined in slit-shaped pores over a wide range of confinement and temperatures. The model is seen to capture the short-time behavior of the VACF extremely accurately and in this region is superior to the predictions of the VACF obtained from the real lobe of the INM DOS. Although the model predicts a zero self diffusivity, the predicted relaxation times are in better agreement with the molecular dynamics results when compared with those obtained from the INM theory. PMID- 15291632 TI - A cellular automaton for the modeling of oscillations in a surface reaction. AB - The reaction of CO and O over a catalytic surface is studied with a cellular automata (CA) model. We extend the CA model proposed by Mai and von Niessen [Phys. Rev. A 44 R6165 (1991)] taking into account the variation of the temperature of the catalyst with the aim of analyzing the existence of oscillations in this reaction. The rate constants for different processes which govern the reaction are chosen in the Arrhenius form. Quasiperiodic, aperiodic, O poisoned, and CO-poisoned regimes are observed depending on the temperature relaxation parameter. The results from the CA model presented are in agreement with several oscillatory behaviors which the catalyzed oxidation of CO exhibits. PMID- 15291633 TI - Quasielastic and inelastic neutron scattering on hydrated calcium silicate pastes. AB - Using the inverse geometry spectrometer QENS at the Intense Pulsed Neutron Source of the Argonne National Laboratory, we collected quasielastic and inelastic neutron scattering spectra of hydrated tricalcium and dicalcium silicate, the main components of ordinary Portland cement. Data were obtained at different curing time, from a few hours to several months. Both the quasielastic and inelastic spectra have been analyzed at the same time according to the relaxing cage model, which is a model developed to describe the dynamics of water at supercooled temperatures. Short-time and long-time dynamics of hydration water in hydrated cement pastes as a function of the curing time have been simultaneously obtained. The results confirm the findings reported in previous experiments showing that it is possible to fit consistently the quasielastic and inelastic spectra giving insights on the effect of the curing time on the short-time vibrational dynamics of hydration water. PMID- 15291634 TI - Modulated hydrogen beam study of adsorption-induced desorption of deuterium from Si(100)-3x1:D surfaces. AB - We have studied the kinetic mechanism of the adsorption-induced-desorption (AID) reaction, H+D/Si(100) --> D2. Using a modulated atomic hydrogen beam, two different types of AID reaction are revealed: one is the fast AID reaction occurring only at the beam on-cycles and the other the slow AID reaction occurring even at the beam off-cycles. Both the fast and slow AID reactions show the different dependence on surface temperature Ts, suggesting that their kinetic mechanisms are different. The fast AID reaction overwhelms the slow one in the desorption yield for 300 K < or = Ts < or = 650 K. It proceeds along a first order kinetics with respect to the incident H flux. Based on the experimental results, both two AID reactions are suggested to occur only on the 3x1 dihydride phase accumulated during surface exposure to H atoms. Possible mechanisms for the AID reactions are discussed. PMID- 15291635 TI - Theoretical investigation of electromechanical effects for graphyne carbon nanotubes. AB - We present a theoretical study of the electronic and mechanical properties of graphyne-based nanotubes (GNTs). These semiconducting nanotubes result from the elongation of one-third of the covalent interconnections of graphite-based nanotubes by the introduction of yne groups. The effect of charge injection on the dimensions of GNTs was investigated using tight-binding calculations. Low amounts of electron injection are predicted to cause qualitatively different responses for armchair and zigzag graphyne nanotubes. Although the behavior is qualitatively similar to the usual carbon nanotubes, the charge-induced strains are predicted to be smaller for the GNTs than for ordinary single walled carbon nanotubes. PMID- 15291636 TI - Single molecule photon emission statistics for non-Markovian blinking models. AB - The statistics of photon emission from a single molecule under continuous wave excitation are considered. In particular, we study stochastic model systems where photon emission rates evolve in time with non-Markovian dynamics. Our calculations are based on the recently introduced generalized optical Bloch equation (GBE) formalism, but with numerical complications beyond those seen in previous Markovian stochastic models. A spectral representation is introduced to facilitate the numerical solution of the GBE equations for these more challenging cases. PMID- 15291637 TI - Grand canonical Monte Carlo simulation of a model colloid-polymer mixture: coexistence line, critical behavior, and interfacial tension. AB - Grand canonical Monte Carlo simulations are used to study phase separation in a simple colloid-polymer model, the so-called Asakura-Oosawa model. To overcome the problem of small acceptance rates of the grand-canonical moves, cluster moves are introduced. Successive umbrella sampling, recently introduced by Virnau and Muller [J. Chem. Phys. 120, 10925 (2004)], is used to access the phase-separated regime. The unmixing binodal and the interfacial tension are measured and compared to theoretical predictions. By means of finite-size scaling, the behavior close to the critical point is also investigated. Close to criticality, we observe substantial deviations from mean-field behavior. PMID- 15291638 TI - The role of molecular shape in bilayer elasticity and phase behavior. AB - A previously developed molecular level model for lipid bilayers [G. Brannigan and F. L. H. Brown, J. Chem. Phys. 120, 1059 (2004)] is extended to allow for variations in lipid length and simulations under constant surface tension conditions. The dependence of membrane elasticity on bilayer thickness is obtained by adjusting lipid length at constant temperature and surface tension. Additionally, bilayer fluidity at various lipid lengths is quantified by analysis of a length versus temperature phase diagram at vanishing tension. Regions of solid, gel-like (hexatic) and fluid bilayer behavior are established by identification of phase boundaries. The main melting transition is found to be density driven; the melting temperature scales inversely with lipid length since thermal expansion increases with lipid aspect ratio. PMID- 15291639 TI - A statistical-mechanical model of polymer liquid crystals subjected to external deformations. AB - We consider the nematiclike system of polymer liquid crystal (PLC) macromolecules represented by the Flory semiflexible chain model. Segments of the flexible (F) spacers are shorter than the LC hard-rod sequences. We investigate effects of imposition of external deformations. The behavior of LC sequences is largely governed by orienting interactions while for F spacers the short-range interactions determined by the chemical structure are the most important. The stress-strain relation is obtained in addition to the orientation-deformation relations. Orientational order phase transitions caused by the external deformations of the system are recognized and described. PMID- 15291640 TI - Hydrogen motions in the alpha-relaxation regime of poly(vinyl ethylene): a molecular dynamics simulation and neutron scattering study. AB - The hydrogen motion in poly(vinyl ethylene) (1,2-polybutadiene) in the alpha relaxation regime has been studied by combining neutron spin echo (NSE) measurements on a fully protonated sample and fully atomistic molecular dynamics simulations. The almost perfect agreement between experiment and simulation results validates the simulated cell. A crossover from Gaussian to non-Gaussian behavior is observed for the intermediate scattering function obtained from both NSE measurements and simulations. This crossover takes place at unusually low Q values, well below the first maximum of the static structure factor. Such anomalous deviation from Gaussian behavior can be explained by the intrinsic dynamic heterogeneity arising from the differences in the dynamics of the different protons in this system. Side group hydrogens show a markedly higher mobility than main chain protons. Taking advantage of the simulations we have investigated the dynamic features of all different types of hydrogens in the sample. Considering each kind of proton in an isolated way, deviations from Gaussian behavior are also found. These can be rationalized in the framework of a simple picture based on the existence of a distribution of discrete jumps underlying the atomic motions in the alpha process. PMID- 15291641 TI - Contributions of short-range and excluded-volume interactions to unperturbed polymer chain dimensions. AB - A Monte Carlo study is made of the mean-square radius of gyration for the freely rotating chain with such fictitious excluded-volume interactions that the Lennard-Jones 6-12 potentials at the Theta temperature act only between the fourth-through (3+Delta)th-neighbor beads (Delta > or = 1) along the chain. The behavior of the asymptotic value (/n)infinity of the ratio /n as a function of the number n of bonds in the chain in the limit of n --> infinity is examined as a function of Delta. It is shown that the approach of (/n)infinity to its value for the real unperturbed chain with Delta = infinity is so slow that the interactions between even up to about 100th-neighbor beads should be taken into account in order to reproduce nearly its dimension. The result implies that the unperturbed polymer chain dimension as experimentally observed at the Theta temperature depends not only on short-range interactions but also to a considerable extent on the long-range excluded-volume interactions, and that the asymptotic value Cinfinity of the characteristic ratio Cn for the rotational isomeric state model in the limit of n --> infinity, which is determined only by the very local conformational energy, cannot be directly compared with the corresponding experimental value. PMID- 15291642 TI - Anisotropic colloidal particles in critical fluids. AB - We consider anisotropic colloidal particles with dumbbell or lens shapes that are immersed in a critical binary fluid mixture. The orientation-dependent long ranged universal interactions mediated by the critical solvent between a particle and a wall or between two particles are investigated for mesoscopic particle sizes small compared to the correlation length and interparticle distances. Exact results are obtained using a "small particle operator expansion." The amplitudes of the isotropic and anisotropic operators in the expansion depend on the size and aspect ratio of the dumbbell or lens and are determined by density profiles in the Ising model at the critical point in a wedge geometry with symmetry breaking fixed-spin boundary conditions. Dumbbells and ellipsoids with a symmetry preserving surface are also considered. PMID- 15291643 TI - Comparison in fractal dimension between those obtained from structure factor and viscoelasticity of gel networks of 1,3:2,4-bis-O-(p-methylbenzylidene)-D-sorbitol in polystyrene melt at gel point. AB - We investigated time evolution of shear moduli in the physical gelation process of 1,3:2,4-bis-O-(p-methylbenzylidene)-D-sorbitol in polystyrene melt. At the gel point, storage and loss shear moduli, G' and G", were described by the power law of frequency omega, G' approximately G" approximately omegan, with the critical exponent n being nearly equal to 2/3, in agreement with the value predicted by the percolation theory. We also investigated the structure factor over two decades in length scale at gel point by using ultra-small-angle X-ray scattering, and small-angle X-ray scattering. We found the power-law behavior in low-q region, indicating that the gel network forms the self-similar structure with mass-fractal dimension. Comparison between the exponent of mass-fractal dimension from structure factor and that from viscoelasticity indicates that hydrodynamic interactions are completely screened out and the excluded volume effects are dominant in the gel. The gel strength was found to increase with the decrease in the lower limit length scale of fractality. PMID- 15291644 TI - Memory effects in collective dynamics of Brownian suspensions. AB - We obtain macroscopic equations for average suspension velocity and particle current in a Brownian suspension valid on long time scales for which the memory effects are important. The coefficients in these equations depend solely on local properties of the medium. This formalism allows one to obtain well-defined theoretical expressions for transport coefficients, free of the integrals diverging with the size of the system. As an example, the expression for long time collective diffusion coefficient is derived and the memory contribution to this coefficient is estimated. PMID- 15291645 TI - Comment on "Stiffness in stochastic chemically reacting systems: the implicit tau leaping method" [J. Chem. Phys. 119, 12784 (2003)]. PMID- 15291647 TI - Anticonvulsants as mood stabilizers and adjuncts to antipsychotics: valproate, lamotrigine, carbamazepine, and oxcarbazepine and actions at voltage-gated sodium channels. AB - Actions of certain anticonvulsants upon voltage-gated sodium channels may not only explain why they are effective mood stabilizers but may also explain why they could be useful adjuncts to antipsychotics for resistant psychosis. PMID- 15291648 TI - Medicine: the coming challenges and opportunities. PMID- 15291649 TI - Suicidality as a possible side effect of antidepressant treatment. PMID- 15291650 TI - Honoring our evidence base. PMID- 15291651 TI - The use of trazodone as a hypnotic: a critical review. AB - BACKGROUND: The last few years have seen a remarkable rise in the off-label use of trazodone for inducing sleep in nondepressed patients, to a degree that it is prescribed for this purpose as commonly as the leading hypnotic. In view of this widespread popularity, it seems prudent to review what is known of the safety and efficacy of trazodone when used in this context. DATA SOURCES AND SELECTION: A MEDLINE search of the literature published in English between 1975 and 2003 that included the keywords sleep, trazodone, Desyrel, depression, sleeping pill, and sedative-hypnotics was conducted. DATA SYNTHESIS: From this review, it is concluded that there are very few data to suggest that trazodone improves sleep in patients without mood disorder, though it does increase total sleep in patients with major depressive disorder. There are virtually no dose-response data for trazodone vis-a-vis sleep and, similarly, no available data on tolerance to its possible hypnotic effects. Areas of concern with its use include reports of significant dropout rates and induction of arrhythmias, primarily in patients with histories of cardiac disease, as well as the development of priapism. CONCLUSION: In summary, there are few data to support the use of trazodone in nondepressed subjects. When the risk-benefit ratio of trazodone is assessed, its side effect profile, which is much more significant than that of conventional hypnotics, should be considered. PMID- 15291652 TI - Relationship between costs and symptoms in schizophrenia patients treated with antipsychotic medication: a review. AB - BACKGROUND: The purpose of this review is to understand how changes in costs of illness are related to the effects of antipsychotic medications on symptoms in schizophrenia patients. METHOD: A search of the MEDLINE database was performed using the keywords costs, symptoms, and schizophrenia. Studies published between 1965 and 2003 in English, French, German, or Spanish that assessed costs, symptoms, and relationships between costs and symptoms were reviewed. RESULTS: Twenty studies were identified. Most of the reviewed clinical trials of antipsychotic medications reported a decrease in mean costs of illness and an improvement in symptoms. However, many of the studies did not examine the relationship between changes in costs and symptoms. CONCLUSION: There is little evidence that changes in costs of illness are directly related to the effects of antipsychotic medications on symptoms. This review emphasizes the need for standardizing the assessment of costs and clinical outcomes, looking more specifically at the relationship between types of costs and specific aspects of psychopathology and developing new statistical models relating changes in costs and clinical outcomes. PMID- 15291654 TI - No deterioration of cognitive performance in an aggressive unilateral and bilateral antidepressant rTMS add-on trial. AB - BACKGROUND: Cognitive functions were assessed before and following a course of repetitive transcranial magnetic stimulation (rTMS) in patients with depression participating in a sham-controlled, randomized trial of rTMS as adjunct to antidepressant treatment. METHOD: Forty-one medicated inpatients with a DSM-IV diagnosis of a depressive episode were consecutively randomly assigned to 1 of 3 groups comparing 2 active rTMS conditions with sham stimulation. The rTMS was applied either at high frequency over the left dorsolateral-prefrontal cortex (DLPFC) (10 sessions x 10 trains x 10 seconds 20 Hz at 100% motor threshold [MT], 90-second intertrain interval) or in a combined high- and low-frequency manner to the left and right DLPFC, respectively (10 sessions x 1 train x 10 minutes at 120% MT). Thirty-eight patients completed a neuropsychological test battery at baseline and following day 14. The cognitive assessment focused on motor skills, attention, executive functions, learning, and memory. Data were collected from November 1999 to August 2002. RESULTS: Active treatment groups did not differ with respect to assessed cognitive measures and thus were pooled. A comparison of short-term changes (baseline-day 14) in neuropsychological performance revealed a more favorable time course of the actively treated patients for encoding in the verbal memory test compared with the sham-stimulated patients. CONCLUSIONS: Unilateral rTMS as well as bilateral combined rTMS revealed no detrimental effects on cognition, as compared with the sham group. Moreover, neither the add on design nor the used aggressive parameters had a negative impact on cognitive measures in comparison with sham. Repetitive transcranial magnetic stimulation might have mild beneficial cognitive effects partly independent of its antidepressant efficacy. PMID- 15291653 TI - Adjunctive fluvoxamine inhibits clozapine-related weight gain and metabolic disturbances. AB - BACKGROUND: Adjunctive fluvoxamine inhibits clozapine metabolism and decreases plasma norclozapine (a toxic metabolite of clozapine) to clozapine ratios. This study aimed to demonstrate the effects of fluvoxamine on clozapine-related weight gain, hyperglycemia, and lipid abnormalities. METHOD: Sixty-eight treatment resistant inpatients with a DSM-IV diagnosis of schizophrenia were randomly assigned to 2 treatment groups for 12 weeks. The monotherapy group (N = 34) received clozapine (< or = 600 mg/day). The coadministration group (N = 34) received fluvoxamine (50 mg/day) plus low-dose clozapine (< or = 250 mg/day). The study was conducted from August 1999 to October 2002. RESULTS: The 2 groups were similar in demographic data; baseline body weight and body mass index (BMI); baseline serum glucose, triglyceride, and cholesterol levels; and steady-state plasma clozapine concentration. The monotherapy patients (but not the coadministration patients) had significantly higher (p < .05) body weight, BMI, and serum glucose and triglyceride levels after treatment than at baseline. At week 12, the monotherapy patients also had significantly higher glucose (p = .035), triglyceride (p = .041), and norclozapine (p = .009) (and numerically higher cholesterol) levels than the cotreatment patients. The changes in weight and serum glucose and triglyceride levels were significantly correlated (p = .026, p = .005, and p = .028, respectively) with the plasma concentration of norclozapine but not with plasma levels of clozapine. CONCLUSION: These results suggest that fluvoxamine cotreatment can attenuate weight gain and metabolic disturbances in clozapine-treated patients. Plasma levels of norclozapine, but not clozapine, are associated with increases in weight and serum glucose and triglyceride levels. Of note, coadministration of fluvoxamine could increase plasma clozapine levels markedly and carry the risk of adverse events. If this combined treatment is applied, conservative introduction with reduced clozapine dosage and careful therapeutic drug monitoring of clozapine concentration is recommended. PMID- 15291655 TI - Motor conversion disorders reviewed from a neuropsychiatric perspective. AB - BACKGROUND: Conversion disorder is a somatoform disorder defined by the presence of pseudoneurologic symptoms relating to voluntary sensory or motor function. The correct diagnosis of conversion disorder presenting with motor symptoms is complicated by the lack of gold-standard diagnostic tests and the absence of a universally accepted set of positive diagnostic criteria. This article reviews the epidemiology, pathophysiology, presentation, differential diagnosis, treatment, and prognosis of motor conversion, placing emphasis on diagnostic validity, reliability, and utility, while evaluating the empirical evidence supporting diagnostic and treatment strategies. DATA SOURCES AND STUDY SELECTION: Literature searches were carried out in PubMed using the keywords conversion disorder, motor conversion, dystonia, psychogenic, hysteria, somatization, motion disorder, movement disorder, and patho-physiology. Articles and book chapters in the author's personal collection were also utilized. CONCLUSIONS: Advances in neuropsychiatric research are leading to significant improvements in the diagnosis and understanding of motor conversion disorders. Positive, objective, and quantitative diagnostic criteria show significant promise for enhancing diagnostic accuracy. Current pathophysiologic research has begun to provide mechanistic explanations for conversion symptoms, thus blurring the distinction between psychogenic and organic motor disorders. PMID- 15291656 TI - The current understanding of lamotrigine as a mood stabilizer. AB - OBJECTIVE: To examine whether lamotrigine has a unique role in the treatment of bipolar disorder, we evaluated the results of recent clinical trials and molecular and cell biological studies on lamotrigine. DATA SOURCES: Using keywords such as bipolar disorder, lamotrigine, clinical trial, outcomes studies, and mechanisms, we conducted a search for English-language articles on MEDLINE and Index Medicus and also on abstracts presented in recent research conferences. DATA SYNTHESIS: Several studies have strongly suggested that lamotrigine is effective for the acute treatment of bipolar depression as well as for long-term maintenance treatment of bipolar disorder. Stevens-Johnson syndrome is a concern, but the incidence of this side effect may not be as high as previously believed, if dosing is slowly titrated. The action mechanisms underlying the mood stabilizing effects of lamotrigine are unknown at present but recent studies have produced interesting leads. Lamotrigine modulates various ion channels, altering neuronal excitability. The use-dependent inhibition of neuronal firing by lamotrigine is potentially important because it could result in attenuating supranormal neuronal activities that are possibly associated with bipolar disorder. Lamotrigine inhibits the release of glutamate, similarly to lithium, and its possible association with mood-stabilizing or antidepressant effects needs to be further examined. Unlike lithium or valproic acid, however, lamotrigine does not down-regulate the expression of protein kinase C or MARCKS, suggesting that lamotrigine employs different intracellular mechanisms for long term changes in neuro-biology from those of lithium or valproic acid. CONCLUSION: The efficacy of lamotrigine for bipolar depression may provide us with new options in the treatment of bipolar disorder. Examining the effects of lamotrigine on various molecular mechanisms in correlation with its unique efficacy on bipolar depression may enhance our understanding of action mechanisms of the mood stabilizers. PMID- 15291657 TI - Sex differences in emotional reactivity to daily life stress in psychosis. AB - BACKGROUND: The expression of schizophrenia has been reported to differ between the sexes. The current study investigates whether these sex differences in clinical expression are reflected in one underlying mechanism that may be causally related to psychosis, namely increases in stress sensitivity in daily life. METHOD: Forty-two participants (22 men, 20 women) with Research Diagnostic Criteria-defined psychotic disorder in a state of clinical remission were studied with the Experience Sampling Method (a structured diary technique assessing current context and mood in daily life) to assess (1) appraised subjective stress related to daily events and activities and (2) emotional reactivity conceptualized as changes in both negative affect and positive affect in relation to the subjective stress. Data were collected from January 1997 to May 1999. RESULTS: Multilevel regression analyses revealed that women reported a significantly (p < .05) increased emotional reactivity to daily life stress compared with men, reflected in both an increase in negative affect and a decrease in positive affect. CONCLUSION: These results suggest that gender differences may not be limited to the characteristics of psychosis but may also be reflected in underlying etiologic mechanisms. Furthermore, these results might strengthen the hypothesis that women are more susceptible than men to a schizoaffective expression of schizophrenia. PMID- 15291658 TI - Severity and comorbidity predict episode duration and recurrence of DSM-IV major depressive disorder. AB - BACKGROUND: Information on the naturalistic outcome of major depressive disorder (MDD) is important in developing rational clinical practices. The aim of this study was to determine the outcome of MDD in a modern secondary-level psychiatric setting and the influence of comorbidity plus psychosocial factors on the outcome of MDD. METHOD: The Vantaa Depression Study is a prospective, naturalistic cohort study of 269 secondary-level care psychiatric outpatients and inpatients diagnosed with a new episode of DSM-IV MDD. Patients were initially interviewed to determine the presence of MDD using the World Health Organization Schedule for Clinical Assessment in Neuropsychiatry and to assess Axis II diagnoses using the Structured Clinical Interview for DSM-III-R personality disorders between February 1, 1997, and May 31, 1998, and were interviewed again at 6 months and 18 months. The exact duration of the index episode and the timing of relapses/recurrences were examined using a life chart. RESULTS: The median length of time that patients met full criteria for a major depressive episode was 1.5 (95% CL = 1.3 to 1.7) months, and the median time to full remission was 8.1 (95% CL = 5.2 to 11.0) months after entry. During the follow-up, 38% of patients had a recurrence. Although numerous factors predict outcome of MDD to some extent, severity of depression and current comorbidity were the 2 most important predictors of longer episode duration and recurrence. CONCLUSION: The course of MDD in modern psychiatric settings remains unfavorable. Any estimates of duration of depressive episodes and rates of recurrence are likely to be dependent on the severity of depression and level of comorbidity. At least among a population of mostly outpatients with MDD in medium-term follow-up, severity of depression and comorbidity appear to be more useful predictors of recurrence than does the number of prior episodes. These factors should influence clinical decision-making regarding the need for maintenance therapy. PMID- 15291659 TI - Lifetime and 1-month prevalence rates of intermittent explosive disorder in a community sample. AB - OBJECTIVE: To determine the lifetime and 1-month prevalence of intermittent explosive disorder (IED) by both DSM-IV and research criteria in a community sample. METHOD: The final 253 (34.1%) of individuals who were entered into the Hopkins Epidemiology Study of Personality Disorder and sampled in the context of a follow-up study of participants from the Baltimore Epidemiologic Catchment Area (ECA) study completed a supplemental interview that allowed for the determination of IED by DSM-IV and/or research criteria. RESULTS: The mean +/- SE percentage of subjects who met inclusion criteria was 11.07% +/- 1.97%, and 6.32% +/- 1.53% met full criteria, for lifetime IED by either diagnostic criteria set; 2.37% +/- 0.96% met full criteria for IED within the previous 1 month. Adjusting the prevalence rates to account for differential sampling from the original ECA study did not substantially affect these results. Onset of problematic aggressive behavior in IED subjects (described as lifelong in most subjects) began as early as childhood, peaked in the third decade, and declined steadily after the fifth decade. While distress and/or impairment due to aggressive behavior was documented in 87.5% of IED subjects, only 12.5% of IED subjects reported seeking help for this problem. CONCLUSIONS: Intermittent problematic aggressive behavior in the community, as defined by IED, may be far more common than previously thought. Conservatively estimated, the number of individuals in the United States with IED, based on these data, may be no lower than 1.4 million for current IED or nearly 10 million for lifetime IED. PMID- 15291660 TI - Psychosis in mania: specificity of its role in severity and treatment response. AB - BACKGROUND: Psychosis is a prominent characteristic of manic episodes. We investigated relationships between the presence of psychotic features, the severity of the manic syndrome, and syndrome severity's response to treatment. METHOD: 179 subjects meeting Research Diagnostic Criteria for a manic episode of bipolar I disorder were hospitalized for acute manic episodes and treated in a randomized trial of lithium, divalproex sodium, or placebo. Factor and cluster analyses were carried out using the clinician-rated Schedule for Affective Disorders and Schizophrenia, Change version (SADS-C) and the nurse-rated Affective Disorder Rating Scale (ADRS). RESULTS: Subjects with psychotic features had significantly (p < .005) greater overall impairment (lower Global Assessment Scale [GAS] scores) but did not differ in severity of mania scores compared with those without psychotic features. Psychosis factor scores correlated significantly (p < .000001) with GAS scores but not with mania scores. Baseline psychosis factor scores did not correlate with subsequent treatment-associated change in mania scores, but change in mania scores during treatment correlated significantly (p < .000001) with change in the psychosis factor. Changes in psychosis factor scores correlated significantly with changes in mania rating scale scores regardless of treatment. CONCLUSIONS: Psychotic features as a component of manic episodes contribute substantially to overall impairment. Treatments that successfully treat mania also reduce psychosis scores. PMID- 15291661 TI - Serotonin function, personality-trait variations, and childhood abuse in women with bulimia-spectrum eating disorders. AB - BACKGROUND: Across populations, findings associate impulsivity, behavioral disinhibition, or hostility with reduced central serotonin (5-hydroxytryptamine: 5-HT) activity and increased likelihood of childhood abuse. Inconsistently, findings associate compulsivity, behavioral inhibition, or anxiousness with elevated 5-HT neurotransmission. We explored relationships among measures of 5-HT system functioning, behavioral inhibition/disinhibition, and childhood abuse in women with bulimia-spectrum eating syndromes. METHOD: In 73 bulimic (body mass index [kg/m2] under 30, binge eating at least once weekly) and 50 normal-eater control women, we obtained indices of platelet paroxetine binding and 5-HT agonist (m-CPP)-stimulated neuroendocrine responses. Cluster analysis was used to classify the bulimic women according to 5-HT "profiles." Resulting groups were then compared on symptom and trait measures. RESULTS: Measures of paroxetine binding density (Bmax) and affinity (Kd) contributed significantly (p < .001 and p < .02, respectively) to a classification of bulimic women into groups with "low density/high affinity" (N = 52) or "high density/low affinity" (N = 21) binding. The 5-HT based classification did not predict eating-symptom severity. However, the "high density" pattern was associated with increased perfectionism and compulsivity, reduced risk of childhood sexual abuse, and (to some extent) reduced probability of borderline personality disorder. DISCUSSION: In women with bulimic syndromes, serotonergic factors, personality-trait variations, and developmental typologies converge in principled fashion. Our findings corroborate (with neurobiological evidence) the concept of underregulated and overregulated subtypes within the bulimic population. PMID- 15291662 TI - Impact of gastrointestinal symptom severity on response to venlafaxine extended release in patients with generalized anxiety disorder. AB - BACKGROUND: This retrospective analysis evaluated the prevalence and severity of pre-treatment gastrointestinal (GI) symptoms in patients with generalized anxiety disorder (GAD), the impact of these GI symptoms on the efficacy and tolerability of venlafaxine extended-release (XR), and the effect of treatment on prestudy GI symptoms. METHOD: Data from 1932 nondepressed GAD patients were pooled from 5 randomized, double-blind, placebo-controlled studies of venlafaxine XR clinically conducted between May 1995 and December 1997. The GI symptom severity at baseline was estimated from item 11 on the Hamilton Rating Scale for Anxiety (HAM-A). Patients with a GI symptom severity score < or = 2 (moderate or less) and those with a GI symptom severity score > 2 (severe/very severe) were compared for baseline characteristics and short-term (8-week) and long-term (24-week) outcomes. RESULTS: At baseline, for all randomized patients with a HAM-A item 11 score, GI symptoms were rated moderate or lower in 82.8% of patients (GI-low) and severe/very severe in 17.2% (GI-high). The GI-high subgroup was statistically significantly (p < .05) younger, had a longer duration of GAD, and had higher mean HAM-A total scores than the GI-low subgroup. Compared with placebo, venlafaxine XR significantly reduced HAM-A total and psychic anxiety factor scores, regardless of baseline GI symptom severity. The incidence of adverse events, particularly nausea, was higher for the GI-high versus GI-low subgroup. CONCLUSION: Baseline severity of GI symptoms correlated with overall severity of GAD but had no impact on treatment outcome with venlafaxine XR. These data do not support the hypothesis that high baseline GI symptom severity has a negative effect on treatment with venlafaxine XR in GAD patients. PMID- 15291663 TI - Psychiatric complications of deep brain stimulation for Parkinson's disease. AB - BACKGROUND: The purpose of this article is to review the current literature regarding deep brain stimulation (DBS) of the subthalamic nucleus as a treatment for Parkinson's disease and to bring to the attention of the psychiatric community the possible psychiatric complications of this treatment. METHOD: A MEDLINE search of English-language publications was conducted using PubMed in July 2003. The search term used was deep brain stimulation. In addition, pertinent references were obtained from the retrieved articles. Reports and studies of psychiatric complications of DBS patients were reviewed and are discussed. A case report is presented of a man who developed hypomanic symptoms shortly after beginning DBS treatment for Parkinson's disease. RESULTS: There have been an increasing number of reports of postprocedure psychiatric complications, including depression, mania, aggression, and deficits in language. Improvement in symptoms of severe obsessive-compulsive disorder and depression has also been reported. CONCLUSION: As information continues to emerge, psychiatrists will play vital roles in the assessment and continuing care of patients who receive DBS. These findings may also provide the framework to determine which patients are at psychiatric risk from DBS. Symptoms of refractory obsessive-compulsive disorder have been noted to improve with DBS, which has led researchers to begin studying its effectiveness for this condition. PMID- 15291664 TI - Renal insufficiency in long-term lithium treatment. AB - OBJECTIVE: To compare long-term lithium patients who developed renal insufficiency (RI) with those who did not, and to examine what characterized these groups. METHOD: One hundred fourteen subjects with DSM-IV bipolar, major depressive, or schizoaffective disorder who had been taking lithium for 4 to 30 years from 1968 to 2000 were studied retrospectively. Subjects with blood creatinine levels > or = 1.5 mg/dL were defined as RI patients, and creatinine levels < 1.5 mg/dL indicated no renal insufficiency (NRI). Ninety-four unmedicated subjects, matched for sex and age, served as a comparison group and had 2 measures of creatinine with a mean interval of 11.88 years. RESULTS: Twenty four (21%) of the lithium-treated patients were defined as RI patients. These subjects exhibited the "creeping creatinine" phenomenon as their creatinine levels increased progressively. The NRI subjects showed no increase of creatinine levels in up to 30 years and remained comparable to the comparison group. RI was associated with episodes of lithium intoxication and diseases or medicines that could affect glomerular function, but not with sex, psychiatric diagnosis, age at onset of diagnosed disorder, duration of lithium therapy, serum lithium concentration, and cumulative lithium dose. CONCLUSIONS: Long-term lithium therapy did not influence glomerular function in an overwhelming majority of patients. However, about 20% of long-term lithium patients exhibited "creeping creatinine" and developed renal insufficiency. PMID- 15291666 TI - Atypical antipsychotics and weight gain in Chinese patients: a comparison of olanzapine and risperidone. AB - OBJECTIVE: To compare the effect of olanzapine with that of risperidone on weight change among Chinese patients in Hong Kong. METHOD: The body weight of subjects maintained on olanzapine or risperidone treatment was recorded at the outpatient clinic of a teaching hospital. Pretreatment weight of the subjects was retrieved from case records. Subjects on olanzapine treatment were matched in sex, age, and diagnosis with those on risperidone treatment, and demographic and clinical data were analyzed. The study was conducted in May and June 2002. RESULTS: Twenty eight olanzapine-risperidone matched pairs were studied. All were diagnosed with DSM-IV schizophrenia. In patients treated with olanzapine and risperidone, respectively, mean +/- SD duration of treatment with atypical neuroleptics was 103.5 +/- 47.4 weeks and 93.2 +/- 50.6 weeks (range, 21-255 weeks), and mean doses were 12.4 +/- 6.7 mg/day and 4.5 +/- 2.8 mg/day. The mean +/- SD weight gain of subjects on treatment with olanzapine and risperidone, respectively, was 8.34 +/- 5.97 kg (18.53 +/- 13.27 lb) and 2.74 +/- 8.09 kg (6.09 +/- 17.98 lb) with a statistically significant difference at p < .005. Lower baseline body weight and body mass index were associated with greater weight gain in both olanzapine- and risperidone-treated subjects. Gender, age, mean daily dose, and duration of treatment had no effect on weight change. CONCLUSION: Treatment with olanzapine was associated with significantly greater weight gain than treatment with risperidone in Chinese schizophrenia patients in Hong Kong. The effect of adjunctive anticonvulsant treatment on weight gain requires further study. PMID- 15291665 TI - A survey of reports of quetiapine-associated hyperglycemia and diabetes mellitus. AB - OBJECTIVE: To explore the clinical characteristics of hyperglycemia in patients treated with quetiapine. METHOD: A pharmacovigilance survey of spontaneously reported adverse events in quetiapine-treated patients was conducted using reports from the U.S. Food and Drug Administration MedWatch program (January 1, 1997, through July 31, 2002) and published cases using the search terms hyperglycemia, diabetes, acidosis, ketosis, and ketoacidosis. RESULTS: We identified 46 reports of quetiapine-associated hyperglycemia or diabetes and 9 additional reports of acidosis that occurred in the absence of hyperglycemia and were excluded from the immediate analyses. Of the reports of quetiapine associated hyperglycemia, 34 patients had newly diagnosed hyperglycemia, 8 had exacerbation of preexisting diabetes mellitus, and 4 could not be classified. The mean +/- SD age was 35.3 +/- 16.2 years (range, 5-76 years). New-onset patients (aged 31.2 +/- 14.8 years) tended to be younger than those with preexisting diabetes (43.5 +/- 16.4 years, p = .08). The overall male:female ratio was 1.9. Most cases appeared within 6 months of quetiapine initiation. The severity of cases ranged from mild glucose intolerance to diabetic ketoacidosis or hyperosmolar coma. There were 21 cases of ketoacidosis or ketosis. There were 11 deaths. CONCLUSION: Atypical antipsychotic use may unmask or precipitate hyperglycemia. UPDATE: An additional 23 cases were identified since August 1, 2002, the end of the first survey, by extending the search through November 30, 2003, bringing the total to 69. PMID- 15291667 TI - Depression and physical symptoms: the mind-body connection. PMID- 15291668 TI - Venlafaxine-induced increase in urinary frequency in 3 women. PMID- 15291669 TI - Transient hyperproinsulinemia during treatment with clozapine and amisulpride. PMID- 15291672 TI - Venlafaxine XR and chronic pelvic pain syndrome. PMID- 15291670 TI - Comments on cost analysis of risperidone versus olanzapine. PMID- 15291673 TI - Bupropion and breastfeeding: a case of a possible infant seizure. PMID- 15291674 TI - Mechanism of action of voltage sensitive sodium channel modulators. PMID- 15291675 TI - Stability in a time of uncertainty. PMID- 15291676 TI - Prepubertal bipolar I disorder and bipolar disorder NOS are separable from ADHD. PMID- 15291677 TI - A preliminary, randomized trial of fluoxetine, olanzapine, and the olanzapine fluoxetine combination in women with borderline personality disorder. AB - BACKGROUND: The intent of this study was to compare the efficacy and safety of fluoxetine, olanzapine, or the olanzapine-fluoxetine combination (OFC) in the treatment of women meeting criteria for borderline personality disorder (without concurrent major depressive disorder). METHOD: We conducted a randomized double blind study of these agents in female subjects meeting Revised Diagnostic Interview for Borderlines (DIB-R) and DSM-IV criteria for borderline personality disorder. Treatment duration was 8 weeks. Outcome measures were clinician-rated scales measuring depression (the Montgomery-Asberg Depression Rating Scale) and impulsive aggression (the Modified Overt Aggression Scale). Data were collected from August 2001 through March 2003. RESULTS: Fourteen subjects were randomized to fluoxetine; 16, to olanzapine; and 15, to OFC. Forty-two of these subjects (93.3%) completed all 8 weeks of the trial. Using random-effects regression modeling of panel data of change-from-baseline scores and controlling for time, olanzapine monotherapy and OFC were associated with a significantly greater rate of improvement over time than fluoxetine on both outcome measures. However, it should be noted that fluoxetine treatment led to a substantial reduction in impulsive aggression and severity of depression. Weight gain was relatively modest in all 3 groups but significantly greater in the olanzapine-treated group than in the groups treated with fluoxetine alone or OFC. CONCLUSION: All 3 compounds studied appear to be safe and effective agents in the treatment of women with borderline personality disorder, significantly ameliorating the chronic dysphoria and impulsive aggression common among borderline patients. However, olanzapine monotherapy and OFC seem to be superior to fluoxetine monotherapy in treating both of these dimensions of borderline psychopathology. PMID- 15291678 TI - Relationships of age at onset with clinical features and cognitive functions in a sample of schizophrenia patients. AB - BACKGROUND: A number of studies investigated the relationships of age at onset with clinical presentation and cognitive performance of schizophrenic patients. The aim of the present study was to assess demographic and clinical characteristics; psychopathologic, social functioning, and quality-of-life ratings; and neuropsychological measures in a sample of patients with stabilized schizophrenia and to identify which factors independently contributed to a multiple regression model with age at onset as the dependent variable. METHOD: Ninety-six consecutive outpatients with schizophrenia (DSM-IV-TR criteria) were included in the study. Assessment instruments were as follows: a semistructured interview, the Clinical Global Impressions scale, the Comprehensive Psychopathological Rating Scale, and the Positive and Negative Syndrome Scale (PANSS) for psycho-pathology of schizophrenia; the Calgary Depression Scale for Schizophrenia (CDSS) for depression; the Social and Occupational Functioning Assessment Scale and the Sheehan Disability Scale for social functioning; the Quality of Life Scale; and a neuro-psychological battery including the Wisconsin Card Sorting Test (WCST) and the Continuous Performance Test. Two models of multiple regression were tested: the first included clinical features and psychopathologic, social functioning, and quality-of-life scales; the second also considered neuro-psychological variables. Data were collected from October 2001 to November 2002. RESULTS: The first multiple regression showed that age at onset was significantly related to scores on the PANSS subscale for negative symptoms (p =.042) and the CDSS (p =.041); the second regression found a relation of age at onset with PANSS score for negative symptoms (p =.002) and 2 neuropsychological measures, number of preservative errors on the WCST and Continuous Performance Test reaction time (p =.0005 for both). CONCLUSION: Our data indicate that, when results of neuropsychological tests are considered, early age at onset of schizophrenia is associated with severity of negative symptoms and compromised cognitive measures of executive functioning and sustained attention. PMID- 15291679 TI - Paroxetine, other antidepressants, and youth suicide in New York City: 1993 through 1998. AB - BACKGROUND: Regulatory agencies in the United Kingdom and the United States have recently issued warnings about a possible link between suicidal ideation and attempts and the use of paroxetine in a pediatric patient population. The objective of this study was to determine the proportion of youth suicides that tested positive for paroxetine or other antidepressants in medical examiner toxicologic testing in New York City from 1993 through 1998, the first 6 years that paroxetine was available in the United States. METHOD: Subjects in this medical examiner surveillance study were suicides less than 18 years of age. Serum toxicology was examined for paroxetine and other antidepressants. RESULTS: There were 66 suicides among persons under 18 years of age in the years 1993 through 1998. Toxicology was tested in 58 (87.9%) of the 66 suicides, and 54 (81.8%) had injury-death intervals of 3 days or less. None of the victims had paroxetine detected in their blood obtained at the time of autopsy. Imipramine was detected in 2 victims and fluoxetine in another 2. CONCLUSION: Despite regulatory concerns, none of the autopsies of youth suicides in New York City detected paroxetine in the victims, although other antidepressants were detected in 4 victims. However, in the vast majority of the youth suicides, there was no evidence of anti-depressant use immediately prior to death. PMID- 15291680 TI - Neurosyphilis in newly admitted psychiatric patients: three case reports. AB - BACKGROUND: Neurosyphilis, also known as "general paresis of the insane," at one time accounted for a large portion of admissions to state psychiatric facilities. With the introduction of antibiotics, neurosyphilis is now considered very rare. METHOD: Chart review was performed on patients diagnosed with neurosyphilis who were admitted to a state psychiatric hospital in Raleigh, N.C., during 2002. RESULTS: We identified 3 cases of confirmed neurosyphilis, representing 0.1% of adult admissions, diagnosed in newly admitted psychiatric patients. None of the patients were immunocompromised. Response to antibiotic treatment was poor. CONCLUSIONS: Given the increase in primary and secondary syphilis reported in the 1980s and early 1990s, routine screening of psychiatric patients for the presence of syphilis should be considered. PMID- 15291681 TI - Cognitive-behavioral therapy for medication nonresponders with obsessive compulsive disorder: a wait-list-controlled open trial. AB - BACKGROUND: Cognitive-behavioral therapy (CBT) is generally recommended for obsessive-compulsive disorder (OCD) patients who have failed to respond to approved medications. However, few studies of the efficacy of CBT have selected patients who did not respond to medications. METHOD: We selected 20 adult OCD (DSM-IV criteria) patients with a history of inadequate response to adequate doses of multiple medications, as well as a high rate of comorbid disorders. After a 1-month wait-list period, patients received 15 sessions of outpatient CBT incorporating exposure and ritual prevention. RESULTS: OCD severity (as measured with the Yale-Brown Obsessive Compulsive Scale) decreased significantly (p <.05) after treatment, and gains appeared to have been maintained over a 6-month follow up period. Analysis of clinical significance indicated that 53% (8/15) of treatment completers met this criterion at posttreatment and 40% (6/15) met the criterion at 6-month follow-up. The sample was characterized as having generally poor insight and putting low effort into CBT; these factors significantly (p <.05) predicted degree of improvement. CONCLUSION: CBT is a useful treatment for OCD patients who have failed to respond adequately to multiple serotonin reuptake inhibitor medications. However, these results were attenuated compared with previous trials. Patients with a long history of poor response to medication may have poor insight and/or not put sufficient effort into treatment; these factors are likely to diminish treatment outcome. PMID- 15291682 TI - Quality of life in schizophrenia: the impact of psychopathology, attitude toward medication, and side effects. AB - BACKGROUND: Quality of life (QOL) is now seen as a key outcome variable in schizophrenia. Factors deemed relevant in this context include severity of symptoms, antipsychotic-induced side effects, sociodemographic variables, and patients' subjective response to medication. METHOD: In the current cross sectional study, 80 patients with a schizophrenic disorder according to ICD-10 criteria who had a duration of illness over 1 year and whose discharge from an inpatient unit had been at least 6 weeks earlier were investigated. Apart from the registration of demographic data, various rating scales were used: the Positive and Negative Syndrome Scale (PANSS), the St. Hans Rating Scale for Extrapy-ramidal Syndromes, the UKU Side Effect Rating Scale, the Drug Attitude Inventory, and the Lancashire Quality of Life Profile. RESULTS: More than half of all patients (47/80) indicated that they were satisfied with their life in general. The specific areas of subjective dissatisfaction that were most commonly noted in the present sample concerned partnership and mental health. The depression/anxiety component of the PANSS, parkinsonism, and a negative attitude toward antipsychotic medication negatively influenced the patients' QOL, while cognitive symptoms and employment status correlated with higher QOL. CONCLUSION: Our results highlight the importance of recognizing the complex nature of QOL in schizophrenia patients. They suggest that special attention should be paid to patients who experience anxiety and depressive symptoms or parkinsonism, to those who are unemployed, and to those with negative feelings and attitudes toward antipsychotics. PMID- 15291684 TI - Prevalence, correlates, and disability of personality disorders in the United States: results from the national epidemiologic survey on alcohol and related conditions. AB - OBJECTIVE: To present nationally representative data on the prevalence, sociodemographic correlates, and disability of 7 of the 10 DSM-IV personality disorders. METHOD: The data were derived from the 2001-2002 National Epidemiologic Survey on Alcohol and Related Conditions (N = 43,093). Diagnoses were made using the Alcohol Use Disorder and Associated Disabilities Interview Schedule-DSM-IV Version, and associations between personality disorders and sociodemographic correlates were determined. The relationship between personality disorders and 3 emotional disability scores (Short-Form 12, version 2) was also examined. RESULTS: Overall, 14.79% of adult Americans (95% CI = 14.08 to 15.50), or 30.8 million, had at least 1 personality disorder. The most prevalent personality disorder in the general population was obsessive-compulsive personality disorder, 7.88% (95% CI = 7.43 to 8.33), followed by paranoid personality disorder 4.41% (95% CI = 4.12 to 4.70), antisocial personality disorder 3.63% (95% CI = 3.34 to 3.92), schizoid personality disorder 3.13% (95% CI = 2.89 to 3.37), avoidant personality disorder 2.36% (95% CI = 2.14 to 2.58), histrionic personality disorder 1.84% (95% CI = 1.66 to 2.02), and dependent personality disorder 0.49% (95% CI = 0.40 to 0.58). The risk of avoidant, dependent, and paranoid personality disorders was significantly greater among women than men (p <.05); the risk of antisocial personality disorder was greater among men compared with women (p <.05); and no sex differences were observed in the risk of obsessive-compulsive, schizoid, or histrionic personality disorders. In general, risk factors for personality disorders included being Native American or black, being a young adult, having low socioeconomic status, and being divorced, separated, widowed, or never married. Avoidant, dependent, schizoid, paranoid, and antisocial personality disorders (p <.02 to p <.0001) were each statistically significant predictors of disability. Obsessive-compulsive personality disorder was inconsistently related to disability. In contrast, disability was not significantly different among individuals with histrionic personality disorder compared with those without the disorder. CONCLUSION: Personality disorders are prevalent in the general population and are generally highly associated with disability. This study highlights the need to develop more effective and targeted prevention and intervention initiatives for personality disorders. PMID- 15291683 TI - Changes in serum interleukin-2, -6, and -8 levels before and during treatment with risperidone and haloperidol: relationship to outcome in schizophrenia. AB - BACKGROUND: Many studies have indicated that immune cytokines may be involved in the pathophysiology of schizophrenia. Recently, there have been reports that typical and atypical antipsychotic drugs may influence the levels of cytokines or cytokine receptors. The aim of this study was to compare the effect of typical and atypical antipsychotic drugs on serum interleukin-2 (IL-2), interleukin-6 (IL 6), and interleukin-8 (IL-8) and to investigate the relationship between the changes in cytokines and the therapeutic outcome in schizophrenia. METHOD: From April 1996 to August 1997, seventy-eight inpatients with a diagnosis of chronic schizophrenia (DSM-III-R) were randomly assigned to 12 weeks of treatment with 6 mg/day of risperidone or 20 mg/day of haloperidol. Clinical efficacy was determined using the Positive and Negative Syndrome Scale. Serum IL-2 was assayed by radioimmunometric assay, and serum IL-6 and IL-8 concentrations were measured by quantitative enzyme-linked immunosorbent assay in patients and 30 sex- and age matched normal subjects. RESULTS: Both risperidone and haloperidol reduced the elevated serum IL-2 concentrations in schizophrenia, and no significant difference was noted in the reduction of serum IL-2 concentrations between risperidone and haloperidol treatment. Neither risperidone nor haloperidol showed significant influence on the higher serum IL-6 or IL-8 concentrations in schizophrenia. Correlations between serum IL-2 or IL-8 concentrations at baseline and the therapeutic outcome were observed, demonstrating that patients presenting with low concentrations of serum IL-2 or IL-8 at baseline showed greater improvement and patients presenting with higher serum IL-2 or IL-8 concentrations at baseline showed less improvement after treatment. CONCLUSIONS: Both typical and atypical anti-psychotic drugs may at least partially normalize abnormal immune alterations in schizophrenia. Some immune parameters at baseline may be useful for predicting the neuroleptic response of schizophrenic patients. PMID- 15291685 TI - Incidence and duration of side effects and those rated as bothersome with selective serotonin reuptake inhibitor treatment for depression: patient report versus physician estimate. AB - BACKGROUND: Selective serotonin reuptake inhibitors (SSRIs) are widely used as the first-line treatment for depression. Information regarding their side effects is mostly based on controlled clinical trials. METHOD: Patients who received an SSRI for a new or recurrent case of depression (ICD-9 code 296.2 or 311) between December 15, 1999, and May 31, 2000 were interviewed by telephone 75 to 105 days after initiation of SSRI therapy. Using closed-ended questions, investigators asked patients if they experienced any of 17 side effects commonly associated with SSRIs, how bothersome they were, and what their duration was. Prescribing physicians completed a written survey providing their estimates about frequency of side effects associated with SSRIs and how bothersome those side effects are. RESULTS: Of 401 patients who completed the phone interview, 344 patients (86%) reported at least 1 side effect, and 219 patients (55%) experienced 1 or more bothersome side effect(s). The most common bothersome side effects were sexual dysfunction and drowsiness (17% each). While most side effects first occurred within the first 2 weeks of treatment, the majority of patients were still experiencing the same side effects at the time of interview, most notably blurred vision (85%) and sexual dysfunction (83%). Overall, physicians (N = 137) significantly underestimated the occurrence of the 17 side effects explored, and they tended to underrate how bothersome those side effects were to their patients. CONCLUSION: Side effects associated with SSRIs are common and bothersome to patients. Treatment-emergent side effects tend to persist during the first 3 months of treatment. PMID- 15291686 TI - Are illness concepts a powerful predictor of adherence to prophylactic treatment in bipolar disorder? AB - BACKGROUND: Predicting and preventing premature discontinuation of medication would substantially improve prophylactic treatment of bipolar disorder. Patients' concepts regarding illness proved to have an impact on noncompliance in a retrospective study of patients with affective or schizoaffective illness treated with lithium. The present study is the first to prospectively investigate the influence of illness concepts on adherence of bipolar patients to different medications. METHOD: 171 bipolar patients (DSM-IV) were randomly assigned to receive either lithium (N = 86) or carbamazepine (N = 85) and observed for a maintenance period of 2.5 years (Multi-center Study of Affective and Schizoaffective Psychoses). The total score and 7 dimensions of illness concepts and treatment expectations of the Illness Concept Scale (ICS) were calculated for 141 patients with completed questionnaires and used to predict time to dropout (Cox regression). Analyses were corroborated by further multivariate analyses with sex, age, and premorbid personality as covariates. RESULTS: With lithium treatment, but not carbamazepine treatment, the total ICS score at study entry was associated with a longer time to study dropout (p =.001 and p =.224, respectively). The relevant ICS subscales affecting time to dropout in patients treated with lithium were trust in medication, trust in the treating physician, and absence of negative treatment expectations. Multivariate analyses suggested that the impact of these variables on adherence to lithium was largely independent of sociodemographic, clinical, and psychological variables. Our data indicate that the stronger impact of illness concepts with lithium as compared with carbamazepine treatment might be related to the drugs' different side effect profiles. CONCLUSION: As trust in drug treatment and trust in the treating physician had a clear impact on adherence to prophylactic lithium, patients' illness concepts and treatment expectations might be promising targets for psychoeducation and psychotherapy in the treatment of bipolar disorder. PMID- 15291687 TI - The effectiveness of olanzapine, risperidone, quetiapine, and ziprasidone as augmentation agents in treatment-resistant major depressive disorder. AB - BACKGROUND: Many questions remain regarding the use of atypical neuroleptics as antidepressant augmentation agents. To date, there have been no reports in the literature regarding the effectiveness of these drugs when trials of one or more of them have failed previously as antidepressant augmentation. METHOD: This retrospective chart review was conducted to determine the effectiveness of olanzapine, risperidone, quetiapine, and ziprasidone when given in a fee-for service setting as anti-depressant augmentation agents to patients with treatment resistant, nonpsychotic major depressive disorder (DSM-IV). Prospective (Global Assessment of Functioning [GAF]) along with retrospective (Clinical Global Impressions-Improvement [CGI-I] and -Severity of Illness scales) ratings were completed for each patient. Analyses were conducted in an attempt to identify factors that appeared to correlate with response, including order of administration and Thase-Rush staging of treatment resistance. RESULTS: In this study of 76 medication trials in 49 patients, the overall response rate based on the CGI-I ratings was 65% (32/49). Individual rates of response were 57% (21/37) for olanzapine, 50% (7/14) for risperidone, 33% (6/18) for quetiapine, and 10% (1/10) for ziprasidone. None of the differences between neuroleptics in rates of response were significant. The difference between baseline and final GAF scores was statistically significant only in the olanzapine (p <.001) and risperidone (p =.047) groups. Rates of discontinuation did not vary significantly between agents, though trends were present. Crossover trials from one atypical neuroleptic to another in the event of nonresponse appeared to be effective. CONCLUSIONS: Although limited by its design, this study suggests atypical neuroleptic augmentation of antidepressants may be a viable option in treatment resistant major depressive disorder. PMID- 15291688 TI - Naltrexone in the treatment of adolescent sexual offenders. AB - BACKGROUND: Naltrexone is a long-acting opioid used clinically in alcoholism, drug abuse, bulimia nervosa, obsessive-compulsive disorder, and impulse-control disorders. This study investigated whether naltrexone can decrease sexual arousal in legally adjudicated adolescent sexual offenders. METHOD: In an open-ended prospective study, naltrexone was given to 21 adolescents participating in an inpatient adolescent sexual offenders program who met any of the self-reported criteria of (1) masturbating 3 or more times per day, (2) feeling unable to control arousal, (3) spending more than 30% of awake time in sexual fantasies, or (4) having sexual fantasies or behavior that regularly intruded into and interfered with their functioning in the treatment program. After having been treated for more than 2 months, 13 patients had their naltrexone administratively stopped, thus providing a before, during, after, and resumption-of-treatment design. Behavioral changes were monitored daily with a fantasy-tracking log and a masturbation log. A positive result was recorded if there was more than a 30% decrease in any self-reported criterion that was applicable to each specific patient and this benefit lasted at least 4 months. Data were collected from July 2000 to December 2002. Leuprolide was given if naltrexone was not sufficiently helpful in controlling sexual impulses and arousal. RESULTS: Fifteen of 21 patients were considered to have a positive result and continued to respond for at least 4 months to an average dose of 160 mg per day with decreased sexual fantasies and masturbation. Dosages above 200 mg per day were not more helpful. Administrative discontinuation of naltrexone in a subset of 13 patients resulted in reoccurrence of symptoms that began when the dose taper reached 50 mg per day. There were no changes in clinical chemistries. Five of 6 patients who did not benefit from naltrexone responded favorably to leuprolide. CONCLUSIONS: Naltrexone at dosages of 100 to 200 mg per day provides a safe first step in treating adolescent sexual offenders. It is possible that the benefits observed here will generalize to the larger population of non-socially deviant hypersexual patients or "sexual addicts." PMID- 15291689 TI - Antidepressant-withdrawal mania:a critical review and synthesis of the literature. AB - BACKGROUND: Hypomania or mania have rarely been reported to develop shortly after the discontinuation of an antidepressant drug. The true incidence of this discontinuation syndrome is unknown because it may be underreported as a consequence of underrecognition or misattribution. This article examines the possible etiology, nosology, mechanisms, and other aspects of the syndrome. DATA SOURCES AND STUDY SELECTION: A PubMed search was conducted in May 2003 and repeated in January 2004 using the search terms antidepressant and mania. Relevant articles containing adequate descriptions for presentation were retrieved, and their reference lists were hand-searched for further pertinent material. Hand-searches of the indexes of leading psychiatry journals were also performed for the years 1998-2003. Twenty-three articles were identified for review. CONCLUSIONS: Antidepressant-withdrawal hypomania or mania may occur rarely with almost any antidepressant drug after sudden withdrawal, tapered discontinuation, or even merely a decrease in dose. The syndrome may be self limiting, may abate with the reinstitution of the antidepressant drug, or may require specific anti-manic treatments; mood stabilizers do not necessarily protect against the syndrome. The true incidence of the syndrome is unknown. Narrow and broad diagnostic criteria are proposed for the syndrome, and a synthesis of literature is provided. PMID- 15291690 TI - Obsessive-compulsive spectrum disorders in rheumatic fever with and without Sydenham's chorea. AB - BACKGROUND: Recent findings suggest that acute-phase rheumatic fever (RF) patients present with higher frequencies of obsessive-compulsive disorder (OCD) and tic disorders. Until now, there have been no such studies in RF in non-acute phases. OBJECTIVE: To verify whether patients with a history of RF with or without Sydenham's chorea (SC) present with higher rates of OCD, tic disorders, and other obsessive-compulsive (OC) spectrum disorders (such as body dysmorphic disorder [BDD]) than controls. METHOD: Between February 1999 and December 2002, 59 consecutive outpatients with non-acute RF (28 with and 31 without SC) from an RF clinic and 39 controls from an orthopedics clinic were blindly assessed for OC spectrum disorders using structured interviews to assign DSM-IV diagnosis. Data were analyzed with Fisher exact and chi(2) tests to compare frequencies of disorders, and Kaplan-Meier survival analyses were used to obtain age-corrected rates. RESULTS: The age-corrected rates of tic disorders were higher in patients with RF without SC (N = 3; 14.39%) (p =.003) when compared with controls. Age corrected rates for OC spectrum disorders (OCD, tic disorders, and BDD) combined were higher both in RF without SC (N = 4; 20.65%) and RF with SC (N = 5; 19.55%) groups than in controls (N = 1; 2.56%) (p =.048). CONCLUSIONS: RF, even in the non-acute phase, may increase the risk for some OC spectrum disorders, such as OCD, tic disorders, and BDD. These data, although preliminary, reinforce the idea that OC spectrum disorders may share common underlying pathophysiologic mechanisms and vulnerability factors with RF or that RF could trigger central nervous system late manifestations such as OC spectrum disorders. PMID- 15291692 TI - Sleepiness versus sleeplessness: shift work and sleep disorders. PMID- 15291691 TI - Alcohol use disorder comorbidity in eating disorders: a multicenter study. AB - BACKGROUND: Eating disorders and alcohol use disorders (AUDs) commonly co-occur, although the patterns of comorbidity differ by eating disorder subtype. Our aim was to explore the nature of the co-morbid relation between AUDs and eating disorders in a large and phenotypically well-characterized group of individuals. METHOD: We compared diagnostic and personality profiles of 97 women with lifetime anorexia nervosa only, 282 women with lifetime bulimia nervosa only, and 293 women with a lifetime history of both anorexia nervosa and bulimia nervosa or anorexia nervosa with binge eating (ANBN) (DSM-IV criteria). All individuals were participants in a multicenter study of the genetics of anorexia nervosa and bulimia nervosa. We explored pattern of onset, Axis I and II comorbidity, and personality characteristics of individuals with and without AUDs by eating disorder subtype. Personality characteristics were assessed with the Multidimensional Perfectionism Scale, the Temperament and Character Inventory, and the Barratt Impulsivity Scale. RESULTS: Alcohol use disorders were significantly more prevalent in women with ANBN and bulimia nervosa than in women with anorexia nervosa (p =.0001). The majority of individuals reported primary onset of the eating disorder, with only one third reporting the onset of the AUD first. After eating disorder subtype was controlled for, AUDs were associated with the presence of major depressive disorder, a range of anxiety disorders, and cluster B personality disorder symptoms. In addition, individuals with AUDs presented with personality profiles marked by impulsivity and perfectionism. CONCLUSIONS: Individuals with eating disorders and AUDs exhibit phenotypic profiles characterized by both anxious, perfectionistic traits and impulsive, dramatic dispositions. These traits mirror the pattern of control and dyscontrol seen in individuals with this comorbid profile and suggest that anxiety modulation may be related to alcohol use in this group. PMID- 15291693 TI - Schizophrenia and binge-eating disorders. PMID- 15291694 TI - The effects of newer antidepressants on low-density lipoprotein cholesterol levels. PMID- 15291696 TI - Duloxetine's role in the treatment of depression and associated painful physical symptoms. PMID- 15291697 TI - Metyrapone for delirium due to Cushing's syndrome induced by occult ectopic adrenocorticotropic hormone secretion. PMID- 15291698 TI - Beneficial effects of gonadotropin-releasing hormone analogue treatment on positive and negative symptoms of schizophrenia: a case report. PMID- 15291699 TI - Addressing immigration-related separations in Hispanic families with a behavior problem adolescent. AB - This article presents specialized family therapy intervention strategies for Hispanic families with behavior-problem adolescents who have experienced an immigration-related separation. Such specialized interventions correspond to a philosophy of customized treatment delivery for Hispanic families. Interactional and cognitive/affective features are presented, and guidelines for building therapeutic alliances, identifying core family processes/themes, and transforming interactions are offered. PMID- 15291700 TI - Self-concealment, social self-efficacy, acculturative stress, and depression in African, Asian, and Latin American international college students. AB - The primary purpose of this exploratory investigation was to examine self concealment behaviors and social self-efficacy skills as potential mediators in the relationship between acculturative stress and depression in a sample of 320 African, Asian, and Latin American international college students. The authors found several differences by demography with regard to the study's variables. After controlling for regional group membership, sex, and English language fluency, they found that self-concealment and social self-efficacy did not serve as mediators in the relationship between African, Asian, and Latin American international students' acculturative stress experiences and depressive symptomatology. Implications of the findings are discussed. PMID- 15291701 TI - Innovative ways to address the mental health and medical needs of marginalized patients: collaborations between family physicians, family therapists, and family psychologists. AB - This article describes an innovative program to meet the needs of homeless women, children, and families residing at a transitional living center in an urban setting. The program involves collaboration between medical and mental health professionals to address the multiple problems and unmet needs of this population. Recommendations for future work in expanding collaborative practice are discussed. ((c) 2004 APA, all rights reserved) PMID- 15291702 TI - Predicting children's reactions to terrorist attacks: the importance of self reports and preexisting characteristics. AB - Forty-eight mothers and their 11-year-old children, who were participants in a longitudinal study, were interviewed in their home after the terrorist attacks of September 11, 2001. Children's verbatim statements were analyzed for fear, separation anxiety, denial, rationalization, anger, and empathy. In the final model, preexisting child anxiety and maternal worry significantly explained 33% of the variance in children's self-reported fearful feelings. PMID- 15291703 TI - Teachers' psychological reactions 7 weeks after the 1995 Oklahoma City bombing. AB - This study assessed teachers' reactions to the Oklahoma City bombing. Peritraumatic reactions, the interaction of media exposure with stress from media coverage, feelings toward the perpetrators, and worry about safety predicted posttraumatic stress. Twenty percent reported difficulty handling demands; 5% sought counseling. Outreach efforts should assess and assist teachers. PMID- 15291704 TI - The extension of school-based inter- and intraracial children's friendships: influences on psychosocial well-being. AB - Children's (N=142) school friendships with same versus different race peers were coded for prevalence and the extent to which parents maintained social relationships with these friends (a proxy for extension of friendships beyond the school context). Membership in integrated versus nonintegrated social networks at school was unassociated with psychosocial well-being. Out-of-school extension of interracial friendships was linked with greater social competence among Black children. Black children whose friendships with both same and different race peers were extended beyond the school context reported higher levels of self esteem. PMID- 15291705 TI - PTSD and world assumptions following myocardial infarction: a longitudinal study. AB - The study aims to examine the association between exposure to trauma, posttraumatic stress disorder (PTSD), and world assumptions. The study included 3 groups of subjects: PTSD myocardial infarction (MI) patients, non-PTSD MI patients, and matched controls. World assumptions were examined twice: within a week of the patient's MI (Time 1) and 7 months later (Time 2). The findings indicate that world assumptions are not related to exposure to trauma but are associated with PTSD. PMID- 15291706 TI - Children's voices: the perceptions of children in foster care. AB - Scant research exists on how abused and neglected children view the foster care experience and how these perceptions vary by demographic characteristics and placement type. Data come from a national probability sample of children placed in child welfare supervised foster care for at least 1 year. These findings indicate that children generally feel positively toward their out-of-home care providers and maintain hope for reunification with their biological family. Differences are present between children in family foster care, group care, and kinship care placements. PMID- 15291707 TI - Youth problems among adoptees living in one-parent homes: a comparison with others from one-parent biological families. AB - Exploring how rising family dissolutions affect adopted children, the authors investigated 2 competing viewpoints: (a) a double jeopardy hypothesis, positing adoptees are susceptible to heightened risks of adjustment problems because of a compounding of parental losses, versus (b) a buffering hypothesis, suggesting early birth parent losses buffer an adoptee's ability to accept parental loss from divorce. With data from the 1994 National Longitudinal Study of Adolescent Health (Add Health, 2003), the authors compared adaptations of adolescent adoptees living with 1 adoptive parent (n=143) with those of children living in step- or single-parent biological families (n=7,457) in a nationally representative sample. Results were mixed, showing some support for both hypotheses and mostly nonsignificant differences on the largest number of behavioral comparisons made. PMID- 15291708 TI - Coping with "absence-presence": noncustodial fathers' parenting behaviors. AB - This article is based on the view that the nature of the divorced father's involvement with his children is affected by psychological processes that enable him to separate his parental from his spousal role and identity. It argues that the ability to cope with the simultaneous absence of the spousal role and identity and presence of the paternal role and identity is a key factor in shaping the divorced father's behavior toward his children. The article illustrates the claim in 3 case studies showing (a) parental functioning marred by ongoing conflict with the children's mother, (b) disengagement, and (c) stable and consistent parental functioning within the inevitable limitations of noncustodial fatherhood. PMID- 15291709 TI - Developing and tailoring mental health technologies for child welfare: the Comprehensive Assessment and Training Services (CATS) Project. AB - This article describes the development and implementation of the Comprehensive Assessment and Training Services Project, a multidisciplinary center designed to prevent children from lingering in the foster care system and to provide early interventions to prevent lifelong problems. This article outlines the conditions that led to the identified need for such a program; the conceptual model used to guide protocol development and refinement: the methodological approach to evaluation, intervention, and technology transfer; specific program components; and, finally, the challenges and barriers to success. PMID- 15291710 TI - Maternal and paternal stress in families with school-aged children with disabilities. AB - This study examined stress factors in families with a school-aged child with a disability. Path analyses revealed that children's demandingness and neediness for care was related more to maternal stress and that child's acceptability was related more to paternal stress. Professionals who serve families with children with disabilities may need to devise more specialized support programs to help fathers become emotionally close to their atypical children and may need to provide more respite services for mothers. To assist parents of school-aged children with disabilities, support services may also need to extend beyond the usual early childhood period. PMID- 15291711 TI - Supporting caregivers of frail older adults in an HMO setting. AB - The long-term effectiveness of a structured health education program (HEP) for spouses and frail older adults was evaluated in a staff model health maintenance organization (HMO). HEP is a multicomponent group program that includes emotion focused and problem-focused coping strategies, education, and support. For caregivers, HEP was more effective than usual care (UC) in reducing depression, increasing knowledge of community services and how to access them, and changing caregivers' feelings of competence and the way they respond to the caregiving situation. For care recipients, HEP was more effective than UC in preventing increases in somatic symptoms and symptoms of anxiety/insomnia. ((c) 2004 APA, all rights reserved) PMID- 15291713 TI - Use of patient and hospital variables in interpreting patient satisfaction data for performance improvement purposes. AB - Satisfaction scores of 349 patients being discharged from a state psychiatric hospital were examined in relation to available norms for the instrument used and selected patient and hospital variables. Mean item scores fell within the less than-satisfied category on both total and factor scores. Regression analyses indicated minimal effects of patient attributes. Two hospital factors (restraint rate on patient's unit and accessibility of psychosocial groups) significantly predicted satisfaction, with the former having an unexpected positive relationship to satisfaction. Clinicians were able to use the survey data to improve care, but patients' tendency toward undifferentiated positive or negative responding hindered the prioritizing of change efforts. PMID- 15291712 TI - Perceived reasons for substance misuse among persons with a psychiatric disorder. AB - The etiology of substance use among persons with severe mental illness remains unclear. This study investigates stated reasons for substance use among persons in recovery from co-occurring disorders of serious mental illness and substance abuse and dependence. The desire to fit in with peers played a key role in the initiation of substance use; boredom, loneliness, temptations to use, and stress were cited most as relapse triggers. The authors discuss the need for dually diagnosed persons to develop sobriety-supporting peer networks to help them learn adaptive strategies to deal with the stress of recovery; further, treatment programs should instill hope for recovery and provide opportunities for meaningful activities and relationships. PMID- 15291714 TI - The SMILES program: a group program for children with mentally ill parents or siblings. AB - The Simplifying Mental Illness + Life Enhancement Skills program, for children with a mentally ill parent or sibling, is a 3-day program that aims to increase children's knowledge of mental illness and to better equip them with life skills considered beneficial for coping in their family. Self-report data from 25 children who attended 3 of these programs, in Canada and Australia, indicate that these aims were achieved. Their parents also report benefits for their children. PMID- 15291715 TI - Galvanizing mental health research in low- and middle- income countries: the role of scientific journals. AB - The Department of Mental Health and Substance Abuse, World Health Organization, organized a meeting on Mental Health Research in Developing Countries: Role of Scientific Journals in Geneva on 20 and 21 November 2003 that was attended by twenty-five editors representing journals publishing mental health research. A number of other editors reviewed and contributed to the background and follow-up material. This statement is issued by all participants jointly (see Appendix B for the list of journals/organizations and their representatives). Research is needed to address the enormous unmet mental health needs of low- and middle income (LAMI) countries. Scientific journals play an important role in production and dissemination of research. However, at present, only a minute proportion of research published in widely accessible mental health and psychiatric journals is from or about these countries. Yet over 85% of the world's population lives in the 153 countries categorized as low and middle income, according to World Bank criteria. Even more worrying is the observation that the gap between these and high-income countries may be widening in terms of their number of publications. The meeting was aimed at finding ways of resolving this unsatisfactory situation. PMID- 15291718 TI - Long-term spatial memory: introduction and guide to the special section. AB - This article provides an introduction to the Special Section on Long-Term Spatial Memory in the journal Neuropsychology. It defines long-term spatial memory, explores how it has been considered historically, and provides brief descriptions of the principle cognitive mechanisms, with a guide to the terminology used in this field. PMID- 15291719 TI - The hippocampal role in spatial memory and the familiarity--recollection distinction: a case study. AB - Memory for object locations and for events (comprising the receipt of an object) was tested in a case of developmental amnesia with focal hippocampal damage. Tests used virtual reality environments and forced-choice recognition with foils chosen to equalize the performance of control participants across conditions. Memory for the objects received was unimpaired, but the context of their receipt was forgotten. Memory for short lists of object locations was unimpaired when tested from the same viewpoint as presentation but impaired when tested from a shifted viewpoint. Same-view performance was disrupted by changing the background scene. These results are consistent with Jon having preserved matching to fixed sensory-bound representations but impaired reconstructed or manipulable representations underlying shifted-viewpoint recognition and episodic recollection. PMID- 15291720 TI - Hippocampal function and spatial memory: evidence from functional neuroimaging in healthy participants and performance of patients with medial temporal lobe resections. AB - Several strategies can be used to find a destination in the environment. Using a virtual environment, the authors identified 2 strategies dependent on 2 different memory systems. A spatial strategy involved the use of multiple landmarks available in the environment, and a response strategy involved right and left turns from a given start position. Although a probe trial provided an objective measure of the strategy used, classification that was based on verbal reports was used in small groups to avoid risks of misclassification. The authors first demonstrated that the spatial strategy led to a significant activity of the hippocampus, whereas the response strategy led to a sustained activity in the caudate nucleus. Then, the authors administered the task to 15 patients with lesions to the medial temporal lobe, showing an impaired ability using the spatial strategy. Imaging and neuropsychological results are discussed to shed light on the human navigation system. PMID- 15291721 TI - Recalling spatial information as a component of recently and remotely acquired episodic or semantic memories: an fMRI study. AB - Activations produced by the recall of episodic and semantic memories differing in spatial content and age were examined. Recall of recent episodic memories with differing spatial content activated the medial temporal lobes and the retrosplenial-posterior cingulate cortex-precuneus complex more than recall of recent semantic memories with similarly differing spatial content. Some of these differences related to the amount of spatial information recalled because spatially richer recent memories, regardless of whether they were episodic or semantic, activated the right posterior parahippocampal cortex, precuneus, and posterior parietal cortex more. This spatial effect was found to be independent of memory age for semantic memories, although some episodic-semantic memory differences, including one in the left hippocampus, were not age independent. Episodic-semantic memory recall activation differences are therefore probably a function of the amount recalled, memory age, and what is recalled, particularly with respect to spatial information. PMID- 15291722 TI - Neural correlates of individual differences in spatial learning strategies. AB - Behavioral studies have shown that spatial skills, such as mental rotation, are correlated with preferences for certain types of spatial information. To be more specific, better mental rotation is associated with a preference for survey (maplike) spatial information relative to route (landmark or wayfinding) information. Functional MRI was used to investigate how individual differences in spatial skills (mental rotation) interact with encoding information from these 2 spatial perspectives. Despite similarities in performance across individuals for route and survey learning, differences between route and survey encoding activation increased with increased mental rotation ability in anterior cingulate, middle frontal gyrus, and postcentral gyrus. This correlation appeared to be due to decreasing activation during survey encoding and not activation changes during route learning. The results suggest that mental rotation skill contributes to survey or map learning but that alternative strategies can be used under the circumstances of this study to achieve equal performance. PMID- 15291723 TI - Allocentric spatial memory activation of the hippocampal formation measured with fMRI. AB - Hippocampal activation was investigated, comparing allocentric and egocentric spatial memory. Healthy participants were immersed in a virtual reality circular arena, with pattern-rendered walls. In a viewpoint-independent task, they moved toward a pole, which was then removed. They were relocated to another position and had to move to the prior location of the pole. For viewpoint-dependent memory, the participants were not moved to a new starting point, but the patterns were rotated to prevent them from indicating the final position. Hippocampal and parahippocampal activation were found in the viewpoint-independent memory encoding phase. Viewpoint-dependent memory did not result in such activation. These results suggest differential activation of the hippocampal formation during allocentric encoding, in partial support of the spatial mapping hypothesis as applied to humans. PMID- 15291724 TI - Allocentric versus egocentric spatial memory after unilateral temporal lobectomy in humans. AB - Thirty patients who had undergone either a right or left unilateral temporal lobectomy (14 RTL; 16 LTL) and 16 control participants were tested on a computerized human analogue of the Morris Water Maze. The procedure was designed to compare allocentric and egocentric spatial memory. In the allocentric condition, participants searched for a target location on the screen, guided by object cues. Between trials, participants had to walk around the screen, which disrupted egocentric memory representation. In the egocentric condition, participants remained in the same position, but the object cues were shifted between searches to prevent them from using allocentric memory. Only the RTL group was impaired on the allocentric condition, and neither the LTL nor RTL group was impaired on additional tests of spatial working memory or spatial manipulation. The results support the notion that the right anterior temporal lobe stores long-term allocentric spatial memories. PMID- 15291725 TI - The spatial brain. AB - Themes emerging from the collection of articles in the Special Section on Long Term Spatial Memory include the notion of multiple spatial systems, the relation between spatial representations and episodic memory, the role of context, and the neural systems involved in space. The authors conclude that distinguishing between egocentric and allocentric spatial systems makes sense of both behavioral and neurobiological data. The special role of the hippocampal system in allocentric space, and as a consequence, in context, suggests how a spatial system might end up central to the ability to remember episodes. PMID- 15291726 TI - Factors influencing Stroop performance in schizophrenia. AB - Recent studies suggest that individuals with schizophrenia show enhanced facilitation but similar interference in reaction times (RTs), compared with control participants, combined with increased error interference. This study examined the relationship between errors and RTs on the Stroop task among individuals with schizophrenia. The authors examined performance on a speeded Stroop task, designed to increase errors, in 29 individuals with schizophrenia and 29 nonpatient control participants. The authors also analyzed color-naming and word-reading estimates from process dissociation analyses. The findings suggest that a lack of increased RT interference among patients (compared with control participants) is not solely due to the increased number of errors they produce in the incongruent condition but is also influenced by a greater impact of the word even in the neutral condition. PMID- 15291727 TI - Neuropsychology of adults with attention-deficit/hyperactivity disorder: a meta analytic review. AB - A comprehensive, empirically based review of the published studies addressing neuropsychological performance in adults diagnosed with attention deficit/hyperactivity disorder (ADHD) was conducted to identify patterns of performance deficits. Findings from 33 published studies were submitted to a meta analytic procedure producing sample-size-weighted mean effect sizes across test measures. Results suggest that neuropsychological deficits are expressed in adults with ADHD across multiple domains of functioning, with notable impairments in attention, behavioral inhibition, and memory, whereas normal performance is noted in simple reaction time. Theoretical and developmental considerations are discussed, including the role of behavioral inhibition and working memory impairment. Future directions for research based on these findings are highlighted, including further exploration of specific impairments and an emphasis on particular tests and testing conditions. PMID- 15291728 TI - Is there a specific executive capacity for dual task coordination? Evidence from Alzheimer's disease. AB - Three experiments compared groups of Alzheimer's disease (AD) patients and healthy older and younger participants on visuospatial tracking and digit sequence recall, as single tasks and performed concurrently. In Experiment 1, tasks were performed concurrently with very low demand relative to span. Only the AD patients showed a dual task deficit. In Experiment 2, single task demand was manipulated on each task from below span to above span for each individual. All groups showed the same performance reductions with increasing demand. In Experiment 3, demand on 1 task was constant, whereas demand on the concurrent task was varied. AD patients showed a clear dual task deficit but were no more sensitive than control groups to varying demand. Results suggest an identifiable cognitive resource for dual task coordination within a multiple component working memory system. PMID- 15291729 TI - Contribution of frontal and temporal lobe function to memory interference from divided attention at retrieval. AB - On the basis of their scores on composite measures of frontal and temporal lobe function, derived from neuropsychological testing, seniors were divided preexperimentally into 4 groups. Participants studied a list of unrelated words under full attention and recalled them while concurrently performing an animacy decision task to words, an odd-digit identification task to numbers, or no distracting task. Large interference effects on memory were produced by the animacy but not by the odd-digit distracting task, and this pattern was not influenced by level of frontal or temporal lobe function. Results show associative retrieval is largely disrupted by competition for common representations, and it is not affected by a reduction in general processing resources, attentional capacity, or competition for memory structures in the temporal lobe. PMID- 15291730 TI - Phonological blocking during picture naming in dementia of the Alzheimer type. AB - Individuals with dementia of the Alzheimer type (DAT; n = 53, ages 55-91), healthy older adults (n = 75, ages 59-91), and younger adults (n = 24, ages 18 24) performed a word-primed picture-naming task. Word primes were neutral (ready), semantically or phonologically related, or unrelated to the correct picture name. AH groups produced equivalent unrelated-word interference and semantic priming effects in response latencies. However, analysis of errors revealed a DAT-related increase of phonological blocking. The results suggest that picture-naming errors in DAT are due, at least in part, to a breakdown in access to phonological representations of object names as a consequence of reduced inhibitory control over other highly active alternatives. PMID- 15291731 TI - Vernier threshold in patients with schizophrenia and in their unaffected siblings. AB - The aim of this study was to investigate magnocellular (M) and parvocellular (P) visual functions in nonmedicated patients with schizophrenia and in their unaffected siblings. Possible abnormalities in cortical integration of retinal receptive fields also were addressed. Twenty-two nonmedicated patients with schizophrenia, their unaffected siblings, and 20 age- and IQ-matched healthy control subjects received 4 vernier acuity tasks (blue-on-yellow, frequency doubling, achromatic low and high contrast conditions) in which they were asked to detect the spatial alignment of dots and gratings. Results revealed that the patients with schizophrenia and their unaffected siblings showed selective dysfunctions in the frequency-doubling and achromatic low contrast conditions, which were devoted to investigate M pathways. In the isoluminant blue-on-yellow and high contrast achromatic conditions, there were no significant differences between the experimental groups. These results suggest that the deficit of M pathway is an endophenotype of schizophrenia. PMID- 15291732 TI - Meta-analysis of intellectual and neuropsychological test performance in attention-deficit/hyperactivity disorder. AB - Cognitive measures are used frequently in the assessment and diagnosis of attention-deficit/hyperactivity disorder (ADHD). In this meta-analytic review, the authors sought to examine the magnitude of differences between ADHD and healthy participants on several commonly used intellectual and neuropsychological measures. Effect sizes for overall intellectual ability (Full Scale IQ; FSIQ) were significantly different between ADHD and healthy participants (weighted d =.61). Effect sizes for FSIQ were significantly smaller than those for spelling and arithmetic achievement tests and marginally significantly smaller than those for continuous performance tests but were comparable to effect sizes for all other measures. These findings indicate that overall cognitive ability is significantly lower among persons with ADHD and that FSIQ may show as large a difference between ADHD and control participants as most other measures. PMID- 15291733 TI - Associative recognition in Alzheimer's disease: evidence for impaired recall-to reject. AB - Patients with mild Alzheimer's disease (AD) were compared with age-matched control subjects on an associative recognition task. Subjects studied pairs of unrelated words and were later asked to distinguish between these same studied pairs (intact) and new pairs that contained either rearranged studied words (rearranged) or non-studied words (non-studied). Studied pairs were presented either once or 3 times. Repetition increased hits to intact pairs in both groups, but repetition increased false alarms to rearranged pairs only in patients. This latter pattern indicates that repetition increased familiarity of the rearranged pairs, but only the control subjects were able to counter this familiarity by recalling the originally studied pairs (a recall-to-reject process). AD impaired this recall-to-reject process, leading to more familiarity based false alarms. These data support the idea that recollection-based monitoring processes are impaired in mild AD. PMID- 15291735 TI - Social perception deficits after traumatic brain injury: interaction between emotion recognition, mentalizing ability, and social communication. AB - Thirty-four adults with severe traumatic brain injuries (TBI) and 34 matched control participants were asked to interpret videotaped conversational exchanges. Study participants were asked to judge the speakers' emotions, the speakers' beliefs (first-order theory of mind), what the speakers intended their conversational partners to believe (second-order theory of mind), and what they meant by remarks that were sincere or literally untrue (i.e., a lie or sarcastic retort). The TBI group had marked difficulty judging most facets of social information. They could recognize speaker beliefs only when this information was explicitly provided. In general, emotion recognition and first-order theory of mind judgments were not related to the ability to understand social (conversational) inference, whereas second-order theory of mind judgments were related to that ability. PMID- 15291734 TI - Effects of diffuse axonal injury on speed of information processing following severe traumatic brain injury. AB - To test the hypothesis that slowed information processing in traumatic brain injury is related to diffuse axonal injury (DAI), the authors compared 10 patients with predominant DAI (diffuse group) and minimal DAI (mixed injury group) on the Symbol Digit Modalities Test, simple and choice reaction time, Trail Making Tests A and B, and the Stroop Neuropsychological Screening Test. The diffuse group was slower than the mixed injury and control groups on basic speed of processing tasks. This difference was not apparent on complex speeded tasks once basic speed of processing was controlled for. The diffuse group's slower speed of processing was not accounted for by differences in injury severity, age, or time postinjury. The diffuse group showed greater recovery over time. PMID- 15291736 TI - Selective attention impairments in Alzheimer's disease: evidence for dissociable components. AB - Tasks emphasizing 3 different aspects of selective attention-inhibition, visuospatial selective attention, and decision making-were administered to subjects with mild Alzheimer's disease (AD) and to healthy elderly control (HEC) subjects to determine which components of selective attention were impaired in AD subjects and whether selective attention could be dissociated into different components. The tasks were administered with easy versus hard levels of difficulty to assess proportional slowing as the key variable across tasks. The results indicated that the inhibitory and visual search tasks showed greater proportional slowing in subjects with AD than in HEC subjects, and that the task involving inhibition was significantly more affected in subjects with AD. Furthermore, there were no significant intertask correlations, and the results cannot be explained simply in terms of generalized cognitive slowing. These results provide evidence that inhibition is the most strikingly affected aspect of selective attention that is observed to be impaired in early stages of AD. PMID- 15291737 TI - Recovery of short-term memory and psychomotor speed but not postural stability with long-term sobriety in alcoholic women. AB - The authors assessed effects of extended abstinence on cognitive and motor function deficits previously observed in a group of alcoholic women (n = 43) initially tested after 15 weeks of sobriety. Alcoholic women were retested 1 and 4 years later, and control women were retested 3 years later. At Year 1, 14 of 23 returners had maintained sobriety, but they did not perform significantly better than relapsers; the group as a whole continued to show deficits relative to age norms. By Year 4, 13 of 14 returners had maintained sobriety for more than 30 months; as a group, these women had returned to normal levels on tests of memory and psychomotor speed but remained impaired in standing balance. PMID- 15291738 TI - Microarray truths and consequences. AB - For many, analysis of a microarray experiment starts with a spreadsheet of expression levels. While great attention is duly paid to RNA extraction, preparation and hybridization, relatively little care is devoted to extraction of expression levels from the fluorescent image. By delegating this step to a click of the mouse the exact extraction process is masked and researchers may be unwittingly compromising their data. In this review, we describe the most common mistakes committed on the path from the image to the spreadsheet and their impact on data quality. Remedies are further proposed for most of the popular microarray platforms in use today. PMID- 15291739 TI - Galanin inhibits leptin expression and secretion in rat adipose tissue and 3T3-L1 adipocytes. AB - In addition to serving as a fat depot, adipose tissue is also considered as an important endocrine organ that synthesizes and secretes a number of factors. Leptin is an adipocyte-derived hormone that plays a vital role in energy balance. Expression of leptin is regulated by dietary status and hormones. In the present study, we report that galanin, an orexigenic peptide, inhibits leptin expression and secretion in rat adipose tissue and in 3T3-L1 adipocytes. Treatment with galanin (25 micro g/animal) induced approximately 46% down-regulation of leptin secretion at 15 min, followed by 40, 37 and 47% decreases in leptin secretion at 1, 2 and 4 h respectively. Although Northern blot analysis of adipose tissue from the same animals showed that leptin mRNA expression in adipose tissue was unaffected by galanin treatment for 2 h, galanin treatment for 4 h led to decline of leptin mRNA expression in a dose-dependent manner. Meanwhile, treating the rats with galanin had no effect on leptin mRNA expression in the hypothalamus. The inhibitory action of the galanin on leptin mRNA and protein levels was also observed in vitro. When incubated with 10 nM galanin for 48 h, leptin mRNA expression and protein secretion also decreased in 3T3-L1 adipocytes. On the other hand, galanin was found not only to express in rat adipose tissue, but also to increase about 8-fold after fasting. Based on these data, we speculate that increased galanin expression in rat adipose tissue after fasting may be involved in reducing leptin expression and secretion in fasting rats. PMID- 15291740 TI - Thyroid hormone regulation of prohormone convertase 1 (PC1): regional expression in rat brain and in vitro characterization of negative thyroid hormone response elements. AB - Most pro-neuropeptides are processed by the prohormone convertases, PC1 and PC2. We previously reported that changes in thyroid status altered anterior pituitary PC1 mRNA and this regulation was due to triiodothyronine (T(3))-dependent interaction of thyroid hormone receptor (TR) with negative thyroid hormone response elements (nTREs) contained in a large region of the human PC1 promoter. In this study, we demonstrated that hypothyroidism stimulated, while hyperthyroidism suppressed, PC1 mRNA levels in rat hypothalamus and cerebral cortex, but not in hippocampus. In situ hybridization was used to confirm real time PCR changes and localize the regulation within the hypothalamus and cortex. Using a human PC1 (hPC1) promoter construct (with and without deletions in two regions that each contain a negative TRE) transiently transfected into GH3 cells, we found that T(3) negatively regulated hPC1 promoter activity, and this regulation required both of these two regions. Electrophoretic mobility shift assays (EMSAs) using purified thyroid hormone receptor alpha1 (TRalpha1) and retinoid X receptor beta (RXRbeta) proteins demonstrated that RXR and TRalpha both bound the PC1 promoter. Addition of TRalpha1/RXRbeta to the wild-type PC1 probe demonstrated binding as both homodimers and a heterodimer. EMSAs with oligonucleotides containing deletion mutations of the putative nTREs demonstrated that the proximal nTRE binds more strongly to TR and RXR than the distal nTRE, but that both regions exhibit specific binding. We conclude that there are multiple novel TRE-like sequences in the hPC1 promoter and that these regions act in a unique manner to facilitate the negative effect of thyroid hormone on PC1. PMID- 15291741 TI - The expression of oestrogen receptor (ER)-beta and its variants, but not ERalpha, in adult human mammary fibroblasts. AB - Whilst oestrogen receptor (ER)-alpha and ERbeta have been shown to be important in the development of the mammary gland, the cell-specific expression pattern of these two receptors within the human breast is not clear. Although it is well established that in the developing rodent mammary gland stromal ERalpha mediates the secretion of growth factors which stimulate the proliferation of the ductal epithelium, the expression of ERalpha in human adult breast stromal fibroblasts is controversial, and the expression of ERbeta has not been properly defined. In the present study, we have evaluated the expression of ERalpha and ERbeta by immunohistochemistry in normal tissue samples, and in purified human breast fibroblasts by Western blotting, RT-PCR analysis and ligand-binding sucrose gradient assay. Our data clearly demonstrated that ERbeta variants, including ERbeta1, ERbeta2, ERbeta5, ERbetadelta and ERbetains, but not ERalpha, are expressed in human adult mammary fibroblasts. These results are supported by the findings that an ERbeta-selective ligand, BAG, but not the ERalpha high-affinity ligand oestradiol, can induce fibroblast growth factor-7 release and activate transcription from an oestrogen-responsive element promoter in these adult human mammary fibroblasts. Together, these observations revealed that, in the adult breast and in breast cancer, the proliferative signals derived from the stroma of adult mammary glands in response to oestrogen are not mediated by ERalpha and provide new insights into the nature of stromal-epithelial interactions in the adult mammary gland. In addition, the expression of these ERbeta variants in cells where there is no ERalpha suggested that these ERbeta splice forms may have functions other than that of modulating ERalpha activity. PMID- 15291742 TI - Functional characterisation of the CRE/TATA box unit of type 2 deiodinase gene promoter in a human choriocarcinoma cell line. AB - The regulation of expression of type II deiodinase (D2) is a critical mechanism to maintain appropriate intracellular concentrations of tri-iodothyronine in selected tissues. One of the major regulators of D2 concentrations is cAMP, which potently increases human type II deiodinase (hD2) gene transcription in some tissues via a conserved cAMP response element (CRE) located in the promoter region. In addition, the regulatory region of the hD2 gene contains several TATA box/transcription start site (TSS) units, suggesting the presence of different transcripts that might be characterised by different biological properties. However, it is still unclear whether one ore more TATA box/TSS units are needed in response to cAMP or to other signals able to modulate hD2 transcription. In this study we have analysed the ability of cAMP to regulate hD2 in JEG3 cells, a human choriocarcinoma cell line highly responsive to cAMP. Transient transfection assays of different hD2 gene promoter constructs revealed that cAMP induces transcription starting from the most 5' TSS, located about 80 nucleotides from the CRE. RT-PCR studies have revealed that cAMP activates the expression of a long-lived transcript in JEG3 cells. Site-directed mutagenesis and deletion analysis of promoter constructs have shown that a single CRE/TATA box/TSS unit is needed to confer responsiveness to cAMP. By using chromatin immunoprecipitation studies, we have also demonstrated that the response to cAMP involves the binding of transcription factor CRE binding protein (CREB) to the CRE located in the hD2 promoter. In summary, in JEG3 cells cAMP induces transcription of a long-lived hD2 RNA via CREB and a single CRE/TATA box/TSS unit. This study provides new insights to the regulation of expression of hD2 in placenta. PMID- 15291743 TI - The molecular characteristics of polycystic ovary syndrome (PCOS) ovary defined by human ovary cDNA microarray. AB - Polycystic ovary syndrome (PCOS) is one of the most common endocrine disorders; it is characterized by polycystic ovaries, hyperandrogenism and chronic anovulation. To obtain a global view of those genes that might be involved in the development of this complex clinical disorder, we used recently developed cDNA microarray technology to compare differential gene expressions between normal human ovary and ovaries from PCOS patients. A total of 9216 clones randomly selected from a commercial human ovary cDNA library were screened. Among them, 290 clones showed differential expressions, including 119 known genes and 100 known or unknown expressed sequence tags (ESTs). Among 119 known genes, 88 were upregulated and 31 downregulated in the PCOS ovary, as compared with normal human ovary. These differentially expressed genes are involved in various biologic functions, such as cell division/apoptosis, regulation of gene expression and metabolism, reflecting the complexity of clinical manifestations of PCOS. The molecular characteristics established from our study will further our understanding of the pathogenesis of PCOS and help us to identify new targets for further studies and for the development of new therapeutic interventions. PMID- 15291744 TI - Selective degradation of cyclin B1 mRNA in rat oocytes by RNA interference (RNAi). AB - Cyclic adenosine monophosphate (cAMP) keeps oocytes in meiotic arrest, thereby preventing activation of the key regulators of meiosis, p34cdc2/cyclin B1, (known as maturation-promoting factor (MPF)) and Erk 1 and 2, members of the mitogen activated protein kinase (MAPK) family. The activity of MAPK in oocytes is upregulated by Mos. We previously demonstrated that Mos translation in rat oocytes is negatively regulated by a PKA-mediated cAMP action, which inhibits c mos mRNA polyadenylation and is associated with the suppression of p34 cdc2 kinase. The goal of the present study was to provide definitive evidence that Mos translation is subjected to MPF regulation. In order to inhibit MPF activity, we employed the double-stranded (ds) RNA interference (RNAi) of gene expression. We demonstrated that the introduction of cyclin B1 dsRNA into rat oocytes selectively depleted the corresponding mRNA, further ablating its protein product. These oocytes, which exhibit low MPF activity, failed to elongate the c mos mRNA poly(A) tail, did not accumulate Mos and were unable to activate MAPK. We conclude that an active MPF in rat oocytes is necessary for c-mos mRNA polyadenylation and Mos translation. PMID- 15291745 TI - Expression of the orphan nuclear receptor ERRalpha is under circadian regulation in estrogen-responsive tissues. AB - Circadian gene expression has been demonstrated in many tissues and involves both positive and negative regulatory loops. The potential interferences of circadian rhythmicity with other well-known biologic rhythms, such as the ovarian cycle, at least in part controlled by estrogens, has not been questioned. The estrogen receptor-related receptor (ERR)alpha is an orphan nuclear receptor that is widely expressed in estrogen-responsive tissues such as liver, uterus and bone. In addition, expression of the ERRalpha gene has been proposed to be transcriptionally controlled by estrogens in the uterus. Here we show that the expression of ERRalpha displays a circadian rhythmicity in liver, bone and uterus. This is in contrast to other uterine estrogen-regulated genes. Analysis of clock/clock mutant mice shows that ERRalpha is an output gene of the circadian clock oscillator. The expression of clock-control genes, such as Bmal1 and Rev erbalpha, also displays diurnal oscillations in the uterus, but not in bone. In this tissue, however, Per2 displayed a rhythmic expression, altogether suggesting unconventional loops in the regulation of circadian rhythm in bone. PMID- 15291746 TI - YY1 binding within the human HSD3B2 gene intron 1 is required for maximal basal promoter activity: identification of YY1 as the 3beta1-A factor. AB - The oxidation and isomerization of 3beta-hydroxy-5-ene steroids into keto-4-ene steroids, a pivotal step in the synthesis of all hormonal steroids, is catalyzed by several isoforms of 3beta-hydroxysteroid dehydrogenase. In humans, two highly homologous isoforms exist, type I expressed by the HSD3B1 gene in peripheral tissues, and type II expressed by the HSD3B2 gene in steroidogenic organs. Previously, it was shown that the HSD3B1 gene 3beta1-A element, encompassing 24 nucleotides of intron 1 not perfectly conserved between the two genes and overlapping with a conserved TG box, contributes to maximal basal promoter activity by binding the ubiquitous and unidentified 3beta1-A transcription factor. In this study for the first time we report that similarly, the HSD3B2 gene intron 1 is required for maximal basal promoter activity in reporter gene analyses, as lack of intron 1 results in a 4- to 10-fold reduction in promoter activity. Mutational analysis in gel shift assays revealed that the 3beta1-A factor binds both the HSD3B2 and HSD3B1 gene intron 1 by requiring only seven nucleotides of a conserved segment within the 3beta1-A element. By competition analysis and use of anti-YY1 antibody in both gel shift and Western blot experiments, we identified the 3beta1-A protein as the ubiquitous transcription factor YY1. In addition, we have characterized another similar YY1 binding site differently located with respect to the 3beta1-A element in both genes. Deletion and mutational analysis in transient transfections experiments revealed that contrarily to as previously shown for the HSD3B1 gene, lack of YY1 binding to the type II 3beta1-A element only results in a marginal reduction of basal promoter activity. Instead, YY1 binding to the second site, placed 35 bp downstream from the 3beta1-A element, strongly activates the HSD3B2 gene basal promoter activity, as preventing YY1 binding to this region caused a 50% decrease of basal transcription. Complete abrogation of YY1 binding within type II intron 1 resulted in a gene reporter activity identical to a reporter construct lacking the whole intron 1. These results designate YY1 as the factor responsible for the intron 1-mediated boost of the HSD3B2 gene basal promoter activity. Similarities and dissimilarities between YY1 binding within the HSD3B1 and HSD3B2 gene intron 1 are discussed involving the conserved intron 1 TG box, that suggests different mechanisms are implicated in the YY1-mediated stimulation of these two genes basal promoter activity. PMID- 15291747 TI - Testosterone modulates mitochondrial aconitase in the full-length human androgen receptor-transfected PC-3 prostatic carcinoma cells. AB - In vitro studies indicated that dihydrotestosterone (DHT) stimulates the enzymatic activity of the mitochondrial aconitase (mACON) in androgen-sensitive prostatic carcinoma cells, LNCaP. Cell proliferation assay determined that DHT doubles the optimal proliferation response of LNCaP cells. The androgen insensitive human prostatic carcinoma cells, PC-3, were overexpressed in the human androgen receptor to assess the involvement of the native androgen receptor in the regulation by DHT of mACON gene expression. A stable-transfected clone that expresses the full-length androgen receptor was selected and termed PCAR9. The results revealed that DHT-treated PCAR9 cells paradoxically not only reduced the enzymatic activity of mACON but also blocked the biosynthesis of intracellular ATP attenuating cell proliferation. Transient gene expression assay indicated that DHT divergently regulates the promoter activity of the mACON gene in LNCaP and PCAR9 cells. This study suggested that DHT regulates mACON gene expression and the proliferation of cells in a receptor-dependent model through modulation by unidentified non-receptor factors. PMID- 15291748 TI - Fenofibrate increases the expression of high mobility group AT-hook 2 (HMGA2) gene and induces adipocyte differentiation of orbital fibroblasts from Graves' ophthalmopathy. AB - Expansion of adipose tissue in the orbits is a key feature of Graves' ophthalmopathy. Recent evidence shows that orbital fibroblasts are committed to differentiate into adipocytes under appropriate stimuli. Rosiglitazone, an agonist of the nuclear hormone receptor, peroxisome proliferator-activated receptor gamma (PPARgamma) is able to induce both differentiation of orbital fibroblasts into mature adipocytes and expression of the TSH receptor (TSHr) gene. Several studies have suggested an important role of the high mobility group AT-hook 2 (HMGA2) gene in adipocytic cell growth and development. To investigate further the association between adipogenesis-related genes and orbital fibroblasts, we treated fibroblasts from Graves' ophthalmopathy (FGOs) and from normal orbital tissues with fenofibrate, a specific agonist for PPARalpha. We then evaluated the expression of the PPARalpha, PPARgamma2, HMGA2, leptin and TSHr genes before and after 24 h of fenofibrate treatment, using semiquantitative and real-time PCR. For up to 96 h after exposure to fenofibrate, FGOs differentiated into adipocytes. PPARalpha and PPARgamma2 were expressed more in FGOs than in normal cultures, whereas TSHr mRNA was detected only in FGOs. Expression of HMGA2 mRNA and protein was significantly increased in FGOs from 6 to 24 h after fenofibrate, confirming its role in the early phase of adipocyte differentiation. Treatment with fenofibrate for 24 h significantly increased the expression of leptin and TSHr genes. Moreover, TSH treatment significantly increased the accumulation of cAMP, demonstrating that FGOs express functional TSHr. The high level of expression of PPARalpha other than PPARgamma2 transcripts and the stimulating effect of fenofibrate on adipogenesis and on HMGA2, leptin and TSHr genes also indicate that the PPARalpha pathway plays an important part in the adipocyte differentiation of FGOs. These findings suggest that novel drugs to antagonize PPARalpha, other than the PPARgamma signalling system, may also need to be considered in the treatment or prevention of Graves' ophthalmopathy. PMID- 15291749 TI - Effect of 17beta-estradiol on hypothalamic GnRH-II gene expression in the female rhesus macaque. AB - The hypothalamus of rhesus macaques expresses two molecular forms of gonadotropin releasing hormone (GnRH-I and GnRH-II). However, it is unclear whether these two neuropeptides play similar roles in the control of reproductive neuroendocrine function, especially in the context of positive and negative estrogen feedback. To address this issue, in situ hybridization histochemistry was used to compare the effect of 17beta-estradiol (E) on the expression of GnRH-I and GnRH-II mRNA in the medial basal hypothalamus (MBH) of adult female macaques. GnRH-I mRNA expression was found to be significantly (P<0.01) more abundant in ovariectomized (ovx) animals compared with ovariectomized E-treated (ovx+E) animals. In marked contrast, GnRH-II mRNA expression was found to be significantly (P<0.05) more abundant in ovx+E animals than in the ovx animals. To help elucidate how E exerts this stimulatory action on GnRH-II gene expression, hypothalamic sections were subsequently double labeled using a combination of immunohistochemisty for estrogen receptor (ER) -alpha or -beta and in situ hybridization histochemistry for GnRH-II. Approximately 50% of the GnRH-II positive cells in the MBH were found to express ERbeta, but none expressed ERalpha. Taken together, these data give credence to a novel pathway by which E may control the primate neuroendocrine reproductive axis, one that involves stimulation of GnRH-II release via an ERbeta-mediated mechanism. PMID- 15291750 TI - IGF-I inhibits apoptosis through the activation of the phosphatidylinositol 3 kinase/Akt pathway in pituitary cells. AB - In previous studies we demonstrated that IGF-I induces proliferation of pituitary lactotrophs. In addition to its mitotrophic actions, IGF-I is known to prevent apoptosis induced by diverse stimuli in several cell types. In this study, we investigated the action of IGF-I on pituitary cell survival and the intracellular signaling transduction pathway implicated in this effect. Treatment of cultured male rat pituitary cells with IGF-I (10(-7) M) for 24 h prevented pituitary cell death induced by serum deprivation. The protective effect of IGF-I was blocked by phosphoinositide 3-kinase (PI3-kinase) inhibitor, LY294002, but was unaffected by PD98059, which inhibits MAP/ERK kinase (MEK1). IGF-I activation of PI3-kinase induced the phosphorylation and activation of the serine/threonine kinase Akt. Moreover, IGF-I increased the phosphorylation of the pro-apoptotic factor Bad and the levels of the anti-apoptotic protein Bcl-2 through the PI3-kinase pathway in primary pituitary cells. PMID- 15291751 TI - Requirement for G proteins in insulin-like growth factor-I-induced growth of prostate cells. AB - Elevated levels of IGF-I in the circulation are associated with increased risk for the development of prostate cancer in men, and transgenic expression of human IGF-I in mouse epithelial prostate cells results in spontaneous prostate tumorigenesis. Little, however, is known about the mechanisms involved in the IGF I-regulated growth of prostate cells. Here, we have demonstrated that treatment with IGF-I induces the activation of the mitogenic extracellular signal-regulated kinase (ERK) pathway and the growth of human prostate cells. Stimulation with IGF I also promoted the tyrosine phosphorylation of epidermal growth factor receptor (EGFR). Signal relay from IGF-I to ERK requires heterotrimeric G proteins and EGFR; inhibition of Gi/o protein activation by pertussis toxin, or EGFR by tyrphostin AG1478 obliterated the ability of IGF-I to promote ERK activation. Further, treatment with pertussis toxin inhibited the IGF-I-mediated prostate cell growth. These data demonstrated the requirement of heterotrimeric G proteins in IGF-I-regulated prostate cell growth and suggest the potential utility of the G proteins as effective drug targets to combat this common cancer. PMID- 15291752 TI - Gene expression profiling of glucocorticoid-inhibited osteoblasts. AB - Glucocorticoid (GC) treatment for the management of autoimmune and inflammatory diseases is associated with decreased bone formation and increased risk for fracture. In MC3T3-E1 cell cultures, 0.1-1 microM dexamethasone (DEX) arrests development of the osteoblast phenotype when administration commences at a commitment stage around the time of confluency. To gain new insights into GC induced osteoporosis, we performed microarray-based gene expression analysis of GC-arrested MC3T3-E1 cultures, 2.5 days after the administration of DEX. Of the >12 000 transcripts interrogated, 74 were up-regulated and 17 were down-regulated by at least 2.5-fold (P < or = 0.05). Some of these genes, such as Mmp13, Serum/GC-regulated kinase and Tieg, have previously been reported as GC responsive. Others are shown here for the first time to respond to GCs. DEX strongly repressed Krox20/Egr2 at both the mRNA and the protein level. This is especially significant because mice lacking this transcription factor develop osteoporosis. The data also suggest that the bone morphogenetic protein (BMP) pathway, which is involved in regulating bone mass, and other pathways that influence BMP signaling, are abrogated by GCs: (i) DEX increased the mRNA levels of the BMP antagonists Follistatin and Dan; (ii) DEX increased the levels of p21 Rasgap3 and Ptpn16/MKP-1 mRNAs, negative regulators of the MAP kinase pathway; and (iii) DEX decreased Cox mRNA levels. DEX also increased thrombospondin mRNA levels, which negatively regulate bone mass in vivo, as well as the adipocytic marker Fkbp51. These and other observations disclose novel gene targets, whose regulation by GCs in osteoblasts may shed light on and provide new therapeutic approaches to osteoporosis. PMID- 15291753 TI - A quantitative RT-PCR study of the mRNA expression profile of the IGF axis during mammary gland development. AB - We have used quantitative RT-PCR to analyse the mRNA expression profile of the major components of the IGF axis in different stages of murine mammary gland development, including late pregnancy, lactation and involution. We have shown that all the genes studied, IGF-I, IGF-II, IGF receptor (IGFR) and IGF-binding protein (IGFBP)-1 to -6, were expressed in every stage, albeit at greatly differing levels and displaying unique expression profiles between developmental stages. IGF-I was always expressed at significantly higher levels than either IGF II or IGFR. This suggests that IGF-I may be the more important IGF during mammary morphogenesis. Overall, IGFBP-3 demonstrated the highest level of expression of any of the IGFBP genes throughout all the developmental stages studied. However, within developmental stages, by far the highest level of expression of any of the IGFBPs was that of IGFBP-5 at day 2 of involution; this was almost an order of magnitude higher than any of the other IGFBP levels recorded. This corroborated our previous findings that the levels of IGFBP-5 protein are highly elevated in the involuting mammary gland, and demonstrated that this up-regulation of IGFBP-5 operates at the level of transcriptional control or message stability. Comparison of the expression profile for these different genes would strongly suggest that they are likely to have differential functions throughout mammary gland development, and also highlights potential interactions and co-regulation between different members of this axis. In addition, our results have identified some similarities and differences in the expression of IGFBPs between the mouse mammary epithelial cell line, HC11, and the normal mammary gland which are worthy of study, most notably the differential regulation of IGFBP-2 and the site of expression of IGFBP-4 and -6. Overall, this study has demonstrated the importance and complexity of the IGF axis during mammary gland development and provides a valuable resource for future research in this area. PMID- 15291754 TI - Expression of two progesterone receptor isoforms in cumulus cells and their roles during meiotic resumption of porcine oocytes. AB - The present study aimed to investigate progesterone receptor (PR) gene expression in cumulus cells and their roles during meiotic resumption of porcine oocytes. The amount of PR-A or PR-B mRNA was analyzed by RT-PCR using primer sets for the PR-B region or the PR-A/B common region. The level of PR-B mRNA in cumulus cells was up-regulated by FSH and LH during the first 8 h of cultivation but the level significantly decreased at 12 h. However, a high level of total PR mRNA was maintained up to a cultivation period of 20 h. The level of PR-B protein in cumulus cells reached its maximum at 4 to 12 h, whereas PR-A predominated in cumulus cells of cumulus-oocyte complexes (COCs) at 20 h. Accompanying the shift in expression of PR isoforms, progesterone production in cumulus cells was significantly increased, and both the proliferative activity of cumulus cells during a 10- to 20-h cultivation period and the level of connexin-43, a major component of the gap junction, in cumulus cells significantly decreased. When COCs were cultured with FSH and LH for 10 h and then further cultured with additional RU486, there was a significant suppression in the shift in PR isoforms and in progesterone production, a loss of proliferative activity, and a decrease in connexin-43 mRNA in cumulus cells. Moreover, treatment with RU486 after 10-h cultivation of COCs inhibited the meiotic resumption of oocytes and cumulus cell expansion. These results suggest that the induction of PR isoforms in cumulus cells and their binding to progesterone appear to impact on proliferation and differentiation in a time-dependent manner, and the shift from PR-B to PR-A may help mediate certain events. PMID- 15291755 TI - Alternative splicing of parathyroid hormone-related protein mRNA: expression and stability. AB - Parathyroid hormone-related protein (PTHrP) is a multifunctional protein that is often dysregulated in cancer. The human PTHrP gene is alternatively spliced into three isoforms, each with a unique 3'-untranslated region (3'-UTR), encoding 139, 173 and 141 amino acid proteins. The regulation of PTHrP mRNA isoform expression has not been completely elucidated, but it may be affected by transforming growth factor-beta1 (TGF-beta1). In this study, we examined differences in the PTHrP mRNA isoform expression in two squamous carcinoma cell lines (SCC2/88 and HARA), an immortalized keratinocyte cell line (HaCaT), and spontaneous human lung cancer with adjacent normal tissue. In addition, the effect of TGF-beta1 on PTHrP mRNA isoform expression and stability was examined. Cell-type specific expression of PTHrP mRNA isoforms occurred between the various cell lines, normal human lung, and immortalized human keratinocytes (HaCaT). PTHrP isoform expression pattern was significantly altered between normal lung tissue and the adjacent lung cancer. In vitro studies revealed that TGF-beta1 differentially altered the mRNA steady-state levels and mRNA stability of the PTHrP isoforms. Protein-RNA binding studies identified different proteins binding to the 3'-UTR of the PTHrP isoforms (139) and (141), which may be important in the differential mRNA stability and response to cytokines between the PTHrP isoforms. The data demonstrate that there is cell-type specific expression of PTHrP mRNA isoforms, and disruption of the normal regulation during cancer progression may in part be associated with TGF beta1-induced changes in PTHrP mRNA isoform expression and stability. PMID- 15291756 TI - Tissue-specific estrogenic and non-estrogenic effects of a xenoestrogen, nonylphenol. AB - Alkylphenols perturb the endocrine system and are considered to have weak estrogenic activities. Although it is known that nonylphenol can bind weakly to the estrogen receptor, it is unclear whether all reported effects of nonylphenol are attributable to its estrogen receptor-binding activity. In order to examine whether alkylphenols have similar effects to the natural hormone, estradiol, we used a mouse model to examine the effects of nonylphenol on gene expression and compared it with estradiol. DNA microarray analysis revealed that, in the uterus, most of the genes activated by this alkylphenol at a high dose (50 mg/kg) were also activated by estradiol. At lower doses, nonylphenol (0.5 mg/kg and 5 mg/kg) had little effect on the genes that were activated by estradiol. Thus, we concluded that the effects of nonylphenol at a high dose (50 mg/kg) were very similar to estradiol in uterine tissue. Moreover, since evaluation of estrogenic activity by gene expression levels was comparable with the uterotrophic assay, it indicated that analysis of gene expression profiles can predict the estrogenic activities of chemicals. In contrast to the similar effects of nonylphenol and estradiol observed in the uterus, in the liver, gene expression was more markedly affected by nonylphenol than by estradiol. This indicated that, in the liver, nonylphenol could activate another set of genes that are distinct from estrogen responsive genes. These results indicated that nonylphenol has very similar effects to estradiol on gene expression in uterine but not in liver tissue, indicating that tissue-specific effects should be considered in order to elucidate the distinct effects of alkylphenols. PMID- 15291757 TI - Serine 124 completes the Tyr, Lys and Ser triad responsible for the catalysis of human type 1 3beta-hydroxysteroid dehydrogenase. AB - Human 3beta-hydroxysteroid dehydrogenase/isomerase (3beta-HSD) is a key steroidogenic enzyme that catalyzes the first step in the conversion of circulating dehydroepiandrosterone (DHEA), pregnenolone or 17alpha hydroxypregenolone to produce the appropriate, active steroid hormone(s): estradiol, testosterone, progesterone, aldosterone or cortisol respectively. Our mutagenesis studies have identified Tyr154 and Lys158 as catalytic residues for the 3beta-HSD reaction. Our three-dimensional homology model of 3beta-HSD shows that Tyr154 and Lys158 are oriented near the 3beta-hydroxyl group of the bound substrate steroid, and predicts that Ser123 or Ser124 completes a Tyr-Lys-Ser catalytic triad that operates in many other dehydrogenases. The S123A and S124A mutants of human type 1 3beta-hydroxysteroid dehydrogenase/isomerase (3beta-HSD1) were created by PCR-based mutagenesis, expressed in insect cells using baculovirus and purified to homogeneity. The S124A mutant exhibits no 3beta-HSD activity and has a K(m) value (83.6 microM) for the isomerase substrate that is threefold greater than that of wild-type 1 isomerase. In contrast, S123A has substantial 3beta-HSD activity (DHEA K(m)=11.2 microM; k(cat)=0.8 min(-1)) and utilizes isomerase substrate, 5-androstene-3,17-dione, with a K(m) value (27.6 microM) that is almost identical to wild-type. The K(m) value (4.3 microM) of S124A for NADH as an allosteric activator of isomerase is similar to that of the wild-type 1 enzyme, indicating that Ser124 is not involved in cofactor binding. S123A utilizes NAD as a cofactor for 3beta-HSD and NADH as the activator for isomerase with K(m) values that are similar to wild-type. The 3beta-HSD activities of S123A and wild-type 3beta-HSD increase by 2.7-fold when the pH is raised from 7.4 to the optimal pH 9.7, but S124A exhibits very low residual 3beta HSD activity that is pH-independent. These kinetic analyses strongly suggest that the Ser124 residue completes the catalytic triad for the 3beta-HSD activity. Since there are 29 Ser residues in the primary structure of human 3beta-HSD1, our homology model of the catalytic domain has been validated by this accurate prediction. A role for Ser124 in the binding of the isomerase substrate, which is the 3beta-HSD product-steroid of the bifunctional enzyme protein, is also suggested. These observations further characterize the structure/function relationships of human 3beta-HSD and bring us closer to the goal of selectively inhibiting the type 1 enzyme in placenta to control the timing of labor or in hormone-sensitive breast tumors to slow their growth. PMID- 15291758 TI - Insulin-mediated activation of activator protein-1 through the mitogen-activated protein kinase pathway stimulates collagenase-1 gene transcription in the MES 13 mesangial cell line. AB - The initial stages of diabetic nephropathy are characterized, in part, by expansion of the mesangial matrix and thickening of the glomerular basement membrane which are caused by increased extracellular matrix (ECM) protein synthesis and reduced degradation, a consequence of decreased matrix metalloproteinase (MMP) activity. These changes have been largely attributed to the effects of hyperglycemia such that the potential contribution of impaired insulin action to alterations in the ECM have not been studied in detail. We have shown here that insulin stimulates collagenase-1 fusion gene transcription in the MES 13 mesangial-derived cell line. Multiple collagenase-1 promoter elements are required for the full stimulatory effect of insulin but the action of insulin appears to be mediated through an activator protein-1 (AP-1) motif. Thus, mutation of this AP-1 motif abolishes insulin-stimulated collagenase fusion gene transcription and, in isolation, this AP-1 motif can mediate a stimulatory effect of insulin on the expression of a heterologous fusion gene. This suggested that the other collagenase-1 promoter elements that are required for the full stimulatory effect of insulin probably bind accessory factors that enhance the effect of insulin mediated through the AP-1 motif. In MES 13 cells, the AP-1 motif is bound by Fra-1, Fra-2, Jun B and Jun D. Stimulation of collagenase-1 fusion gene transcription by insulin requires activation of the mitogen-activated protein kinase (MEK) pathway since inhibition of MEK-1 and -2 blocks this effect. The potential significance of these observations with respect to a role for insulin in the pathophysiology of diabetic glomerulosclerosis is discussed. PMID- 15291759 TI - Expression of epiregulin and amphiregulin in the rat ovary. AB - We have previously reported that the epidermal growth factor (EGF) family growth factor, epiregulin, is expressed in rat ovarian granulosa cells by induction with pregnant mare serum gonadotropin (PMSG). In this study, we report that amphiregulin, another member of the EGF family, was also induced in the rat ovary by gonadotropin treatment. Northern blot analysis revealed that PMSG treatment induced the expression of both epiregulin and amphiregulin mRNA after 24 h, but the expression then decreased 48 h after treatment. Further treatment with human chorionic gonadotropin (hCG) rapidly induced the expression of both epiregulin and amphiregulin genes and maximal levels were reached 4 h after hCG treatment. A marginal increase in amphiregulin mRNA levels was also observed 6 h after PMSG treatment. In situ hybridization revealed that epiregulin and amphiregulin mRNAs were localized in the granulosa cells of large antral follicles. These spatio temporal expression patterns were similar to those of cyclo-oxygenase-2 (COX-2) and progesterone receptor (PR). In adult cycling rats, epiregulin and amphiregulin were strongly induced at 1800 and 2000 h on proestrus coinciding with the preovulatory LH surge. An in situ hybridization study also showed that epiregulin and amphiregulin mRNAs were detectable in the granulosa cells of preovulatory ovarian follicles at 2000 h on proestrus, where transcripts of COX-2 and PR were co-localized with those of epiregulin and amphiregulin. These observations suggested that the EGF family members, epiregulin and amphiregulin, may play a role in the ovulatory process of cycling rats as well as in the induction of ovulation in immature rats. PMID- 15291760 TI - Molecular cloning, pharmacological characterization, and histochemical distribution of frog vasotocin and mesotocin receptors. AB - The neurohypophysial nonapeptides vasotocin (VT) and mesotocin (MT) are the amphibian counterparts of arginine vasopressin (AVP) and oxytocin (OT). We have here reported the cloning and functional characterization of the receptors for vasotocin (VTR) and mesotocin (MTR) in two species of frog, Rana catesbeiana and Rana esculenta. The frog VTR and MTR cDNAs encode proteins of 419 and 384 amino acids respectively. Frog VTR exhibits a high degree of sequence identity with the mammalian AVP-1a (V1a) receptor while the frog MTR possesses a high degree of sequence identity with the mammalian OT receptor. Activation of VTR induced both c-fos promoter- and cAMP-responsive element (CRE)-driven transcriptional activities, while activation of MTR induced c-fos promoter-driven transcriptional activity but failed to evoke CRE-driven transcriptional activity, suggesting differential G protein coupling between VTR and MTR. The VTR exhibited the highest sensitivity for VT followed by OT>AVP approximately MT, whereas the MTR showed preferential ligand sensitivity for MT>OT>VT>AVP. A V1a agonist but not V2 and OT agonists substantially activated both VTR and MTR with a similar sensitivity. V1a, V2 and OT antagonists inhibited MT-induced MTR activation but not VT-induced VTR activation. In the frog brain, VTR and MTR mRNAs were found to be widely expressed in the telencephalon, diencephalon and mesencephalon, and exhibited very similar regional distribution. In the pituitary, VTR and MTR were expressed in the distal and intermediate lobes but were virtually absent in the neural lobe. Taken together, these data indicated that, although the distribution of VTR and MTR largely overlaps in the frog brain and pituitary, VT and MT may play distinct activities owing to the ligand selectivity and different signaling pathways activated by their receptors. PMID- 15291761 TI - Mutational analysis of histidine residues in human organic anion transporter 4 (hOAT4). AB - Human organic anion transporter 4 (hOAT4) belongs to a family of organic anion transporters which play critical roles in the body disposition of clinically important drugs, including anti-HIV therapeutics, antitumour drugs, antibiotics, anti-hypertensives and anti-inflammatories. hOAT4-mediated transport of the organic anion oestrone sulphate in COS-7 cells was inhibited by the histidine modifying reagent DEPC (diethyl pyrocarbonate). Therefore the role of histidine residues in the function of hOAT4 was examined by site-directed mutagenesis. All five histidine residues of hOAT4 were converted into alanine, singly or in combination. Single replacement of His-47, or simultaneous replacement of His 47/52/83 or His-47/52/83/305/469 (H-less) led to a 50-80% decrease in transport activity. The decreased transport activity of these mutants was correlated with a decreased amount of cell-surface expression, although the total cell expression of these mutants was similar to that of wild-type hOAT4. These results suggest that mutation at positions 47, 47/52/83 and 47/52/83/305/469 impaired membrane expression rather than function. We also showed that, although most of the histidine mutants of hOAT4 were sensitive to inhibition by DEPC, H469A (His-469- >Ala) was completely insensitive to inhibition by this reagent. Therefore modification of His-469 is responsible for the inhibition of hOAT4 by DEPC. PMID- 15291762 TI - New insights into the structure-function relationships of Rho-associated kinase: a thermodynamic and hydrodynamic study of the dimer-to-monomer transition and its kinetic implications. AB - The effect of the length of ROCK (Rho-associated kinase) on its oligomerization state has been investigated by analysing full-length protein and four truncated constructs using light-scattering and analytical ultracentrifugation methods. Changes in size correlate with the kinetic properties of the kinase. Sedimentation velocity, sedimentation equilibrium and light-scattering data analyses revealed that protein constructs of size Ser6-Arg415 and larger exist predominantly as dimers, while smaller constructs are predominantly monomeric. The amino acid segments comprising residues 379-415 and 47-78 are shown to be necessary to maintain the dimeric ROCK structure. kcat values ranged from 0.7 to 2.1 s(-1) and from 1.0 to 5.9 s(-1) using ROCK peptide (KKRNRTLSV) and the 20000 Da subunit of myosin light chain respectively as substrate, indicating that the effect of the ROCK oligomerization state on the kcat is minor. Values of ATP K(m) for monomeric constructs were increased by 50-80-fold relative to the dimeric constructs, and K(i) comparisons using the specific competitive ROCK inhibitor Y 27632 also showed increases of at least 120-fold, demonstrating significant perturbations in the ATP binding site. The corresponding K(m) values for the ROCK peptide and myosin light chain substrates increased in the range 1.4-16-fold, demonstrating that substrate binding is less sensitive to the ROCK oligomerization state. These results show that the oligomerization state of ROCK may influence both its kinase activity and its interactions with inhibitors, and suggest that the dimeric structure is essential for normal in vivo function. PMID- 15291764 TI - Sample trial to assess the level of understanding of the mechanisms of drug action. AB - The object of the present paper was to investigate whether or not the level of understanding of the mechanisms of drug action in nursing students is reflected in their descriptions of the factors in ineffective cases or adverse effects of prescribed medicines. The number of subcategories identified in the descriptions was closely connected with the test scores from the multiple-choice and short answer written examination on 'Pharmacology and Pharmaceutics', and those from questions about the mechanisms of drug action. These results suggest that the number is useful for an estimated assessment of the level of understanding of the mechanisms of drug action. PMID- 15291765 TI - Relationship between menopausal symptoms and menopausal status in Australian and Japanese women: preliminary analysis. AB - The main aim of the present study was to explore the midlife experience for women living in Australia and Japan. The specific objectives of the study included: (i) comparing menopausal symptoms between the two groups; and (ii) comparing the factor structure of symptoms and exploring their relationship to menopausal status. Postal questionnaires were distributed to two structured, random population based samples of midlife women aged 45-60 years; consisting of 712 women living in Australia and 1502 women living in Japan. Analysis showed significant differences in menopausal symptoms related to psychological symptoms (P < 0.001), including anxiety (P < 0.001) and depression (P < 0.001), somatic symptoms (P < 0.001), and vasomotor symptoms (P < 0.01). The analysis, which excluded hormone replacement therapy (HRT) users, found that there were significant differences seen across menopausal status in the following symptoms: difficulty in sleeping (P < 0.01), difficulty in concentrating (P < 0.01), feeling dizzy or faint (P < 0.001), loss of interest in most things (P < 0.01) and loss of feeling in hands or feet (P < 0.001). In the postmenopausal stage specifically, significant differences were seen in the areas of feeling tense or nervous (P < 0.01), feeling unhappy or depressed (P < 0.01), parts of body feeling numb or tingling (P < 0.05), headaches (P < 0.01), and sweating at night (P < 0.05). Our analysis revealed that the experience of menopause for women is different between Australian and Japanese women. PMID- 15291766 TI - Nationwide study of environmental assessment for people with schizophrenia in Japan. AB - The purpose of the present study was to develop an environmental assessment tool related to the living of the mentally disabled and to examine the health care professionals' perception of the current situations and its importance. The relationships between the current and ideal environmental situations were analyzed. The survey was conducted at 3310 municipalities nationwide by mailing self-report questionnaires, which consisted of 52 items developed by specialists for the mentally disabled and their families. The 52-item questionnaire was rated with a five-point scale for both current and important environmental situations. The main findings included statistically significant differences between the mean scores of the current and important environmental situations, items perceived as important at a similar degree, and those suggesting the largest gap between the current and important situations related to learning opportunities. These results could be useful for implementation in community health nursing. PMID- 15291767 TI - Comparative study of perceptions of work environment and moral sensitivity among Japanese and Norwegian nurses. AB - The aim of the present study was to explore the relationship between work environment and moral sensitivity among Japanese (n = 138) and Norwegian nurses (n = 71), and to compare the results from a sociocultural perspective using a descriptive-correlational design. Data were analyzed using descriptive and inferential statistics. The results point to a significant relationship between work environment and moral sensitivity for both groups of nurses. In comparison, the Japanese nurses were more focused on 'patient centered oriented care', reported 'work engagement', seeking 'meaning in difficult caring situations' and 'following rules'. In addition, they ranked the factor 'values in action of patient care' as significant and 'relation to superior and colleagues' and 'job stress and anxiety' ranked significant to 'moral conflicts'. The Norwegian nurses were more independent, which was correlated with moderate significance with 'job stress and anxiety'. A significant correlation was found between 'physical and mental symptoms' and 'moral conflicts' among Norwegian nurses. PMID- 15291769 TI - Smoking as a symbol of friendship: qualitative study of smoking behavior and initiation of a group of male nurse students in Iran. AB - The study of smoking initiation and behavior of nurses and nurse students is important when considering their professional health promotion roles. In the present qualitative study, 26 nursing students of all grades from two nursing schools in the capital city of Iran were surveyed to explore their reasons for smoking. Through in-depth and semistructured interviews, participants were encouraged to talk about their smoking behavior and explain initiating reasons freely. Majority of the participants began smoking before commencing nursing, while only a few initiated smoking after entering into nursing school. Reasons for smoking included belonging to a group and as a symbol of friendship and mutual trust. Indeed, peer group behaviors influenced most participates to commence smoking and social influences also emerged as a theme in this study. PMID- 15291768 TI - Development and validation study of the breast cancer risk appraisal for Korean women. AB - The purpose of the present study was to develop and validate breast cancer risk factors, which can be used to identify women who are at a high risk of developing breast cancer. Fourteen risk factors for breast cancer were identified from extensive literature reviews and in consultation with a panel of experts. A case control study was conducted, which aimed to validate these 14 risk factors. Cases were selected from four university hospitals in Korea and controls were selected from health screening centers in Korea. Interviews were used to obtain exposure history of each participant to the 14 risk factors. Group differences between the cases and controls were analyzed using a chi-squared test and a logistic regression method. Among the 14 risk factors, six were identified as significant: meat consumption more than once a week (odds ratio [OR] = 1.56, confidence interval [CI] = 1.32-1.85), breast disease experience (OR = 3.08, CI = 1.49 6.37), less than two children (OR = 1.61, CI = 1.09-2.35), family history of breast cancer (OR = 8.99, CI = 3.89-20.76), and no breast-feeding experience (OR = 3.08, CI = 1.41-3.85). Based on the OR, the risk scores were calculated by the absolute size of the OR having a total of 100 points. PMID- 15291770 TI - Moral conflict and collaborative mode as moral conflict resolution in health care. AB - Moral conflict as a complex moral issue in health care has emerged from several causes that are related to different values, beliefs and opinions. Moral conflict can occur when duties and obligations of health care providers or general guiding ethical principles are unclear. Health care providers and institutions or agencies need to resolve or initiate appropriate methods for professional staff so they can recognize, discuss and resolve moral conflicts in the health care delivery system. Collaborative mode is a useful method for moral conflict resolution, because patient care is a complex phenomenon that results from the integrated knowledge and work of individuals with different professional training. In the process of collaborative practice, all members need to respect each other's opinions, values and responsibilities regarding patient care. PMID- 15291771 TI - Consideration for legacies about diabetes and self-care for the family with a multigenerational occurrence of type 2 diabetes. AB - Type 2 diabetes mellitus is a familial disorder that is fast becoming epidemic in the USA. It is possible that nurses will care for entire families with diabetes in the near future. In multiple studies, family functioning, a family systems variable, has been correlated with self-management and glycemic outcomes. Most persons with diabetes live with family members who might facilitate or inhibit self-management tasks or skills. Multiple afflictions of family members with diabetes can further complicate matters relating to diabetes self-management. Diabetes collectively affects over 16 million people in the USA, incurring healthcare costs in excess of 92 billion dollars annually. Individuals engage in self-management behavior for diabetes care in the context of their family's environment. It is imperative that nurses consider factors that might inhibit or facilitate self-management efforts by individuals with type 2 diabetes mellitus. Assessment of family determinants of adaptation to diabetes with appropriate interventions can improve clinical outcomes, subsequently preventing complications. PMID- 15291772 TI - Development of web-based qualitative and quantitative data collection systems: study on daily symptoms and coping strategies among Japanese rheumatoid arthritis patients. AB - With the sharp rise in Internet access in recent years, the Internet is increasingly being used for research. We have developed two new systems using the Internet for both quantitative and qualitative data collection. One is a dynamic system for creating questionnaires and collecting responses and the other is an individual patient support follow-up system. These systems do not depend on the capacity of computers or servers and enable researchers to interact with participants privately and asynchronously. We have used these systems to collect data on daily symptoms and journal entries from 12 community-dwelling women with rheumatoid arthritis (RA) for a minimum of 1 month. The mean number of data submissions per month was 14.2 +/- 7.8, with the majority recording entries every day, and some submitting several times a day. The combination of two types of data elucidated the changes in coping and coping strategies in conjunction with changes in symptoms, even in a single day, and the coexistence of positive and negative coping. Research participants benefited from web-based symptom management and counseling resulting from 1 month of frequent interactions with the researcher. The use of the Internet for nursing research and interventions thus seems to show promising results. PMID- 15291774 TI - Methods and preliminary results. PMID- 15291775 TI - How children and mothers express gender essentialism. PMID- 15291776 TI - Talk about categories versus individuals (generics vs. non-generics). PMID- 15291777 TI - V. beyond the individual: discourse patterns and correlational analyses. PMID- 15291781 TI - Gendered language and sexist thought. PMID- 15291785 TI - Modes of transmission of Glugea plecoglossi (Microspora) via the skin and digestive tract in an experimental infection model using rainbow trout, Oncorhynchus mykiss (Walbaum). AB - Glugea plecoglossi (Microspora) is a significant cause of economic loss in cultured ayu, Plecoglossus altivelis, in Japan, due to the unsightly appearance of infected fish harbouring xenomas in the body cavity. Modes of transmission of G. plecoglossi via the skin and digestive tract were studied in an experimental infection model using rainbow trout, Oncorhynchus mykiss. Combined with Uvitex 2B and in situ hybridization (ISH) assays, the early development of G. plecoglossi was successfully traced. Following a bath exposure of fish Uvitex 2B-labelled G. plecoglossi spores were observed to attach to microscopic injuries (trypan blue positive sites) of fish skin, after which ISH-positive sporoplasms were found to invade the epidermis as early as 5 min post-infection (PI), migrating rapidly to the subdermis. It was also shown that G. plecoglossi entering via the skin does not spread into the internal organs but develops into subdermal xenomas. After rainbow trout were exposed to G. plecoglossi spores by oral intubation, spores germinated in the intestinal lumen, followed by penetration of sporoplasms into the gut mucosal epithelium 5 min PI. In vitro trials determining stimulation factors (fish mucus, changes in pH, digestive enzymes) for the extrusion of the polar tube were inconclusive. The present study indicates that skin wounds and the gut epithelium can be portals of entry of G. plecoglossi and that natural infection in fish seems to occur perorally rather than via the skin. PMID- 15291786 TI - The induction of laboratory-based amoebic gill disease revisited. AB - Previous work in our laboratory defined a method of inducing laboratory-based amoebic gill disease (AGD) in Atlantic salmon, Salmo salar L. Gills of AGD affected fish were scraped and the debris placed into fish-holding systems, eliciting AGD in naive Atlantic salmon. While this method is consistently successful in inducing AGD, variability in the kinetics and severity of infections has been observed. It is believed that the infections are influenced by inherently variable viability of post-harvest amoeba trophozoites. Here, a new method of experimental induction of AGD is presented that redefines the infection model including the minimum infective dose. Amoebae were partially purified from the gills of AGD-affected Atlantic salmon. Trophozoites were characterized by light microscopy and immunocytochemistry and designated Neoparamoeba sp., possibly Neoparamoeba pemaquidensis. Cells were placed into experimental infection systems ranging in concentration from 0 to 500 cells L(-1). AGD was detected by gross and histological examination in fish held in all systems inoculated with amoebae. The number of gross and histological AGD lesions per gill was proportional to the inoculating concentration of amoebae indicating that the severity of disease is a function of amoeba density in the water column. The implications of these observations are discussed in the context of the existing AGD literature base as well as Atlantic salmon farming in south-eastern Tasmania. PMID- 15291787 TI - Histopathological studies of bacterial haemorrhagic ascites of ayu, Plecoglossus altivelis (Temminck & Schlegel). AB - A histopathological study was carried out on ayu, Plecoglossus altivelis, with bacterial haemorrhagic ascites. The fish were obtained from culture ponds in Wakayama Prefecture in 2003. The causative agent was identified as Pseudomonas plecoglossicida by a slide agglutination test using anti-P. plecoglossicida FPC941 serum. Histopathological studies revealed lesions in spleen, kidney, liver, intestine, heart and gills. Lesions in the spleen and haematopoietic tissue were prominent and invaded by P. plecoglossicida. Necrotic lesions accompanied by haemorrhage, fibrin deposition and oedema occurred in the splenic pulp and sheathed tissue, and in the kidney. The liver also had necrotic lesions and abscess formations. However, the intestine, heart and gills were only slightly invaded by P. plecoglossicida. No lesions or bacteria were observed in the brain. PMID- 15291788 TI - Nodavirus infections in Israeli mariculture. AB - Viral encephalopathy and retinopathy (VER) infections were diagnosed in five fish species: Epinephelus aeneus, Dicentrarchus labrax, Sciaenops ocellatus, Lates calcarifer and Mugil cephalus cultured on both the Red Sea and Mediterranean coasts of Israel during 1998-2002. Spongiform vacuolation of nervous tissue was observed in histological sections of all examined species. With transmission electron microscopy, paracrystalline arrays and pieces of membrane-associated non enveloped virions measuring approximately 30 nm in diameter were observed in the brain and retina of all species. At the molecular level, the nodavirus was detected by using a primer set that amplified the T4 region of the coat protein gene. When the same set of primers was used to search for VER in an additional fish species, Sparus aurata, it was found to produce non-specific amplicons, giving rise to false-positive results. This problem was overcome by using a different primer set (F1/VR3), designed on a highly conserved region of the virus gene, which amplified a fragment of 254 bp, and confirmed that S. aurata was nodavirus-free. This set was validated on all five species of infected fish, as well as clinically healthy fish. Comparison of the coat protein genes from the Israeli isolated sequences indicated that more than one viral strain was involved. No strict host-specificity was evident. Red Sea and Mediterranean isolated sequences grouped in distinct clusters, together with several foreign isolates from the Mediterranean area and the Far East, as phylogenetically close to the Epinephelus akaara RGNNV type. PMID- 15291789 TI - Humoral response and susceptibility of five full-sib families of Atlantic salmon, Salmo salar L., to the haemoflagellate, Cryptobia salmositica. AB - Susceptibility and antibody production against pathogenic and vaccine strains of the haemoflagellate, Cryptobia salmositica were investigated in five full-sib families (A-E) of Atlantic salmon, Salmo salar. Humoral response and susceptibility of families were compared within three treatments: infection, vaccination and vaccination followed by challenge. Parasitaemias caused by the vaccine strain of C. salmositica were considerably lower than those caused by the pathogenic strain. All vaccinated families were protected when challenged with the pathogenic strain. Family B had significantly lower parasitaemias (with both strains) than the other families. When naive fish were infected with the pathogenic strain, this family had a significantly lower and earlier peak parasitaemia (4.3 +/-1.3 x 10(6) parasites mL(-1) blood at 3 weeks post infection; w.p.i.) than the other families. Family C had the highest peak (11.1 +/- 1.2 x 10(6) parasites mL(-1) blood), which occurred at 4 w.p.i. Antibodies against C. salmositica were detected earlier in Family B (3 w.p.i.) than in Family C (5 w.p.i.). This demonstrates an association of increased susceptibility with a delayed antibody response. Western immunoblot identified antibodies against 112, 181 and 200 kDa antigens earlier in more resistant fish (Family B). Antigenic stimulation leading to a stronger antibody response was shown with the vaccine strain and in the later stages of infection. PMID- 15291790 TI - Salmonid gill bacteria and their relationship to amoebic gill disease. AB - 16S ribosomal RNA gene analysis was used to assess the bacterial community associated with Atlantic salmon, Salmo salar L., gills which were either affected by amoebic gill disease (AGD) or were AGD-negative, in order to determine the role that bacteria may play in the development of AGD. AGD-positive specimens were either infected in the laboratory with Neoparamoeba pemaquidensis, the causative agent of AGD, or were obtained from commercial salmon cages. Samples from laboratory fish maintained in sea water possessed a marine-type community while field samples which had been treated by a series of freshwater baths possessed a more diverse community which included variable proportions of different bacterial ecotypes, including groups typically associated with soil, skin surfaces and faeces. Samples from fish infected with AGD in the laboratory and a sample from one of two salmon cage fish specimens were dominated by a phylotype belonging to the strictly marine bacterial genus Psychroserpens (family Flavobacteriaceae, phylum Bacteroidetes). The phylotype was not detected in any of the AGD-negative samples or in one of two AGD-positive samples obtained from fish subjected to temporary freshwater immersion. The possibility of certain Psychroserpens species as potential opportunistic pathogens associated with salmonid AGD is proposed. PMID- 15291791 TI - On cryptobiosis and infections by Cryptobia. PMID- 15291792 TI - Ventrally emigrating neural tube (VENT) cells: a second neural tube-derived cell population. AB - Two embryological fates for cells of the neural tube are well established. Cells from the dorsal part of the developing neural tube emigrate and become neural crest cells, which in turn contribute to the development of the peripheral nervous system and a variety of non-neural structures. Other neural tube cells form the neurons and glial cells of the central nervous system (CNS). This has led to the neural crest being treated as the sole neural tube-derived emigrating cell population, with the remaining neural tube cells assumed to be restricted to forming the CNS. However, this restriction has not been tested fully. Our investigations of chick, quail and duck embryos utilizing a variety of different labelling techniques (DiI, LacZ, GFP and quail chimera) demonstrate the existence of a second neural tube-derived emigrating cell population. These cells originate from the ventral part of the cranial neural tube, emigrate at the exit/entry site of the cranial nerves, migrate in association with the nerves and populate their target tissues. On the basis of its site of origin and route of migration we have named this cell population the ventrally emigrating neural tube (VENT) cells. VENT cells also differ from neural crest cells in that they emigrate considerably after the emigration of neural crest cells, and lack expression of the neural crest cell antigen HNK-1. VENT cells are multipotent, differentiating into cell types belonging to all four basic tissues in the body: the nerve, muscle, connective and epithelium. Thus, the neural tube provides at least two cell populations--neural crest and VENT cells--that contribute to the development of the peripheral nervous system and various non-neural structures. This review describes the origin of the idea of VENT cells, and discusses evidence for their existence and subsequent fates. PMID- 15291793 TI - Hand development and sequence of ossification in the forelimb of the European shrew Crocidura russula (Soricidae) and comparisons across therian mammals. AB - Hand development in the European shrew Crocidura russula is described, based on the examination of a cleared and double-stained ontogenetic series and histological sections of a c. 20-day-old embryo and a neonate. In the embryo all carpal elements are still mesenchymal condensations, and there are three more elements than in the adult stage: the 'lunatum', which fuses with the scaphoid around birth; a centrale, which either fuses with another carpal element or just disappears later in ontogeny; and the anlage of an element that later fuses with the radius. Carpal arrangement in the neonate and the adult is the same. In order to compare the relative timing of the onset of ossification in forelimb bones in C. russula with that of other therians, we built up two matrices of events based on two sets of data and used the event-pair method. In the first analysis, ossification of forelimb elements in general was examined, including that of the humerus, radius, ulna, the first carpal and metacarpal to ossify, and the phalanges of the third digit. The second analysis included each carpal, humerus, radius, ulna, the first metacarpal and the first phalanx to ossify. Some characters (= event-pairs) provide synapomorphies for some clades examined. There have been some shifts in the timing of ossification apparently not caused by ecological and/or environmental influences. In two species (Oryctolagus and Myotis), there is a tendency to start the ossification of the carpals relatively earlier than in all other species examined, the sauropsid outgroups included. PMID- 15291794 TI - Morphology of the long and short uveal nerves in the human eye. AB - The aim of this study was to examine the architecture of the uveal nerves in the sclera and suprachoroid of human eyes. Eyes from 17 adult human donors were investigated. The uveal nerves in different regions (retrobulbar, intrascleral, suprachoroidal, pars plana) were prepared and studied by light and electron microscopy. In addition, immunohistochemistry was performed for various neuronal markers. The long uveal nerves showed a characteristic suprachoroidal location with no branches supplying the choroid. It was found that typically they are composed of myelinated (75%) and non-myelinated (25%) nerve fibres. They mainly contain aminergic and sensory nerve fibres. A separate set of cholinergic non myelinated nerve fibre bundles runs parallel with these long uveal nerves. The short uveal nerves supply the suprachoroidal nerve plexus with approximately 13% of their nerve fibres. The nerves and the branches supplying the choroid appear as mixed nerves containing sympathetic, parasympathetic and sensory axons. This study therefore provides new information about the quantity, type and distribution of myelinated and non-myelinated nerve fibres in the posterior uvea of the human eye. PMID- 15291795 TI - Does the degree of laminarity correlate with site-specific differences in collagen fibre orientation in primary bone? An evaluation in the turkey ulna diaphysis. AB - de Margerie hypothesized that preferred orientations of primary vascular canals in avian primary cortical bone mediate important mechanical adaptations. Specifically, bones that receive habitual compression, tension or bending stresses typically have cortices with a low laminarity index (LI) (i.e. relatively lower cross-sectional areas of circularly (C) orientated primary vascular canals, and relatively higher areas of canals with radial (R), oblique (O) or longitudinal (L) orientations. By contrast, bones subject to habitual torsion have a high LI (i.e. relatively higher C-orientated canal area) [LI, based on percentage vascular canal area, = C/(C + R + O + L)]. Regional variations in predominant collagen fibre orientation (CFO) may be the adaptive characteristic mediated by LI. Using turkey ulnae, we tested the hypothesis that site-specific variations in predominant CFO and LI are strongly correlated. Mid diaphyseal cross-sections (100 +/- 5 micro m) from subadult and adult bones were evaluated for CFO and LI using circularly polarized light images of cortical octants. Results showing significant differences between mean LI of subadult (40.0% +/- 10.7%) and adult (50.9% +/- 10.4%) (P < 0.01) bones suggest that adult bones experience more prevalent/predominant torsion. Alternatively, this relationship may reflect differences in growth rates. High positive correlations between LI and predominant CFO (subadults: r = 0.735; adults: r = 0.866; P < 0.001) suggest that primary bone can exhibit potentially adaptive material variations that are independent of secondary osteon formation. PMID- 15291796 TI - Expression of amyloid precursor protein-like molecule in astroglial cells of the subventricular zone and rostral migratory stream of the adult rat forebrain. AB - In adult mammals, new neurons in the subventricular zone (SVZ) of the lateral ventricle (LV) migrate tangentially through the rostral migratory stream (RMS) to the olfactory bulb (OB), where they mature into local interneurons. Using a monoclonal antibody for the beta-amyloid precursor protein (APP) (mAb 22C11), which is specific for the amino-terminal region of the secreted form of APP and recognizes all APP isoforms and APP-related proteins, immunoreactivity was detected in specific subpopulations of cells in the SVZ and RMS of the adult rat forebrain. In the SVZ, APP-like immunoreactivity was detected in the ependymal cells lining the LV and some of the subependymal cells. The latter were regarded as astrocytes, because they were positive for the glial markers, S-100 protein (S 100) and glial fibrillary acidic protein (GFAP). APP-like immunoreactive astrocytes exhibited strong labelling of the perinuclear cytoplasm and often possessed a long, fine process similar to that found with radial glia. The process extended to an APP-like immunoreactive meshwork in the RMS that consisted of cytoplasmic processes of astrocytes forming 'glial tubes'. Double immunofluorescent labelling with a highly polysialylated neural cell adhesion molecule (PSA-NCAM) confirmed that the APP-like immunoreactive astrocytes in the SVZ and meshwork in the RMS made close contact with PSA-NCAM-immunopositive neuroblasts, suggesting an interaction between APP-containing cells and neuroblasts. This region of the adult brain is a useful in vivo model to investigate the role of APP in neurogenesis. PMID- 15291797 TI - Quantitative (stereological) study of the effects of vasectomy on spermatogenesis in rabbits. AB - Using stereological methods, especially the optical disector for unbiased estimation of nuclear number, our recent study demonstrated that long-term (6 or 12 months) vasectomy in the rhesus monkey had no significant effects on spermatogenesis (Peng et al. Reproduction 2002, 124, 847-856). This study aimed to determine the scenario in the rabbit using the same morphometric methodology. Three groups of normal male Japanese white rabbits (aged 4-5 months) were subjected to unilateral vasectomy; 10 days, 6 months and 12 months later both testes and epididymides were removed. Testicular and epididymal methacrylate embedded sections were obtained for stereology. Vasectomy-induced damage to spermatogenesis was observed, primarily sloughing of spermatogenic cells with a greater reduction in the number of advanced (adluminal) cells. The damage was most severe at 10 days, occurring in all the testes on the vasectomized side and involving sloughing of even type A spermatogonia, the number of which returned to normal at 6 and 12 months. Damage was less severe at 6 and 12 months, being found in half of the testes of the vasectomy side, in which the total numbers of later germ cell types were 24.0-59.1% (spermatocytes) and 0.3-11.6% (spermatids) of control at 6 months, and 20.1-22.1% (spermatocytes) and 0.4-12.0% (spermatids) of control at 12 months. By contrast, Sertoli cell number per testis was unchanged following vasectomy in any group. Epididymis on the vasectomy side, especially at 10 days and 6 months, appeared larger than on the contralateral side, but this difference was not statistically significant, and no sperm granuloma was seen in the epididymis. PMID- 15291801 TI - Expression of angiogenic factors in chronic myeloid leukaemia: role of the bcr/abl oncogene, biochemical mechanisms, and potential clinical implications. AB - Chronic myeloid leukaemia (CML) is a stem cell disease characterized by an increased production and accumulation of clonal BCR/ABL-positive cells in haematopoietic tissues. The chronic phase of CML is inevitably followed by an accelerated phase of the disease, with consecutive blast crisis. However, depending on genetic stability, epigenetic events, and several other factors, the clinical course and survival appear to vary among patients. Recent data suggest that angiogenic cytokines such as vascular endothelial growth factor (VEGF), are up-regulated in CML, and play a role in the pathogenesis of the disease. These factors appear to be produced and released in leukaemic cells in patients with CML. In line with this notion, increased serum-levels of angiogenic growth factors are measurable in CML patients. In this study we provide an overview of angiogenic growth factors expressed in CML cells, discuss the possible pathogenetic role of these cytokines, the biochemical basis of their production in leukaemic cells, and their potential clinical implications. PMID- 15291802 TI - The biological and clinical significance of MLL abnormalities in haematological malignancies. AB - The MLL (Mixed Lineage Leukaemia or Myeloid/Lymphoid Leukaemia) gene on chromosome 11q23 is frequently involved in chromosomal translocations associated with human acute leukaemias. These translocations lead to fusion genes generally resulting in novel chimeric proteins containing the amino terminus of MLL fused in-frame to one of about 30 distinct partner proteins. Abnormalities involving the MLL gene are observed in leukaemias of either lymphoid or myeloid lineage derivation, as well as in poorly differentiated or biphenotypic leukaemias. They are frequently seen in infant patients, and patients with therapy-related secondary AML following treatment with inhibitors of topoisomerase II (epipodophyllotoxins). In the majority of cases, abnormalities involving the MLL gene are associated with a very poor prognostic outcome. In this review, we will discuss some of the recent advances in MLL research resulting from biological as well as clinical studies. PMID- 15291803 TI - Novel molecular diagnostic and therapeutic targets in chronic lymphocytic leukaemia. AB - B-cell lymphocytic leukaemia (B-CLL) is an indolent non-Hodgkin's lymphoma and the most frequent leukaemia. However, after many years, the incurable disease CLL has again become an exciting subject for research. Recently, both serum and molecular markers have been identified which could be used to predict the outcome of patients in early stages. With the advent of microarray analysis, novel diagnostic and therapeutic targets have been discovered. Here we describe the molecular strategies for target identification and validation. An evaluation of some established, and the most promising novel factors, with their diagnostic and prognostic applications is given. Potential therapeutic target molecules and their inhibitors are reviewed. PMID- 15291804 TI - Human leukaemic stem cells: a novel target of therapy. AB - Acute myeloid leukaemia (AML) is a life-threatening haematopoietic disease that is characterized by clonal growth and the accumulation of myelopoietic progenitor cells. Although AML cells only have a limited potential to undergo differentiation and maturation, each AML clone is organized in a hierarchical manner similar to normal haematopoiesis. Recent data have shown that each AML clone consists of leukaemic stem cells and their progeny, and that AML stem cells differ from more mature cells in several aspects, including survival and target antigen profiles. Most importantly, AML stem cells, but not their progeny, have the capacity to repopulate haematopoietic tissues with leukaemias in NOD/SCID mice. Furthermore, AML stem cells are thought to be responsible for the infinite growth of leukaemias in patients with AML. The phenotypic properties of AML stem cells have also been described. In most cases, these cells are detectable within the CD34+, CD38-, Lin-, CD123+ subpopulation of AML cells. Because of their AML initiating and -renewing capacity and their unique phenotype, which includes several molecular targets of drug therapy, AML stem cells have recently been proposed as novel important target cell populations in the context of curative therapies. The present article gives an overview of our knowledge about AML stem cells, their phenotype, and their role as a 'therapy-target' in new concepts to treat and to cure patients with AML. PMID- 15291805 TI - On the way to targeted therapy of mast cell neoplasms: identification of molecular targets in neoplastic mast cells and evaluation of arising treatment concepts. AB - Several emerging treatment concepts for myeloid neoplasms are based on novel drugs targeting cell surface antigens, signalling pathways, or critical effector molecules. Systemic mastocytosis is a haematopoietic neoplasm that behaves as an indolent myeloproliferative disease in most patients, but can also present as aggressive disease or even as an acute leukaemia. In patients with aggressive disease or mast cell leukaemia, the response to conventional therapy is poor in most cases, and the prognosis is grave. Therefore, a number of attempts have been made to define novel treatment strategies for these patients. One promising approach may be to identify novel targets and to develop targeted drug therapies. In this article, we support the notion that neoplastic mast cells indeed express a number of potential molecular targets including immunoreactive CD antigens, the microphthalmia transcription factor (MITF), and members of the Bcl-2 family. In addition, the tyrosine kinase receptor KIT and downstream signalling pathways have been proposed as targets of a specific pharmacological intervention. A particular challenge is the disease-related D816V-mutated variant of KIT, which is resistant against diverse tyrosine kinase inhibitors including STI571, but may be sensitive to more recently developed targeted compounds. The therapeutic potential of target-specific approaches in malignant mast cell disorders should be evaluated in forthcoming clinical trials in the near future. PMID- 15291806 TI - Cold survival in freeze-intolerant insects: the structure and function of beta helical antifreeze proteins. AB - Antifreeze proteins (AFPs) designate a class of proteins that are able to bind to and inhibit the growth of macromolecular ice. These proteins have been characterized from a variety of organisms. Recently, the structures of AFPs from the spruce budworm (Choristoneura fumiferana) and the yellow mealworm (Tenebrio molitor) have been determined by NMR and X-ray crystallography. Despite nonhomologous sequences, both proteins were shown to consist of beta-helices. We review the structures and dynamics data of these two insect AFPs to bring insight into the structure-function relationship and explore their beta-helical architecture. For the spruce budworm protein, the fold is a left-handed beta helix with 15 residues per coil. The Tenebrio molitor protein consists of a right handed beta-helix with 12 residues per coil. Mutagenesis and structural studies show that the insect AFPs present a highly rigid array of threonine residues and bound water molecules that can effectively mimic the ice lattice. Comparisons of the newly determined ryegrass and carrot AFP sequences have led to models suggesting that they might also consist of beta-helices, and indicate that the beta-helix might be used as an AFP structural motif in nonfish organisms. PMID- 15291807 TI - Phylogenetic relationships in class I of the superfamily of bacterial, fungal, and plant peroxidases. AB - Molecular phylogeny among catalase-peroxidases, cytochrome c peroxidases, and ascorbate peroxidases was analysed. Sixty representative sequences covering all known subgroups of class I of the superfamily of bacterial, fungal, and plant heme peroxidases were selected. Each sequence analysed contained the typical peroxidase motifs evolved to bind effectively the prosthetic heme group, enabling peroxidatic activity. The N-terminal and C-terminal domains of catalase peroxidases matching the ancestral tandem gene duplication event were treated separately in the phylogenetic analysis to reveal their specific evolutionary history. The inferred unrooted phylogenetic tree obtained by three different methods revealed the existence of four clearly separated clades (C-terminal and N terminal domains of catalase-peroxidases, ascorbate peroxidases, and cytochrome c peroxidases) which were segregated early in the evolution of this superfamily. From the results, it is obvious that the duplication event in the gene for catalase-peroxidase occurred in the later phase of evolution, in which the individual specificities of the peroxidase families distinguished were already formed. Evidence is presented that class I of the heme peroxidase superfamily is spread among prokaryotes and eukaryotes, obeying the birth-and-death process of multigene family evolution. PMID- 15291808 TI - Platelet factor 4 disrupts the intracellular signalling cascade induced by vascular endothelial growth factor by both KDR dependent and independent mechanisms. AB - The mechanism by which the CXC chemokine platelet factor 4 (PF-4) inhibits endothelial cell proliferation is unclear. The heparin-binding domains of PF-4 have been reported to prevent vascular endothelial growth factor 165 (VEGF(165)) and fibroblast growth factor 2 (FGF2) from interacting with their receptors. However, other studies have suggested that PF-4 acts via heparin-binding independent interactions. Here, we compared the effects of PF-4 on the signalling events involved in the proliferation induced by VEGF(165), which binds heparin, and by VEGF(121), which does not. Activation of the VEGF receptor, KDR, and phospholipase Cgamma (PLCgamma) was unaffected in conditions in which PF-4 inhibited VEGF(121)-induced DNA synthesis. In contrast, VEGF(165)-induced phosphorylation of KDR and PLCgamma was partially inhibited by PF-4. These observations are consistent with PF-4 affecting the binding of VEGF(165), but not that of VEGF(121), to KDR. PF-4 also strongly inhibited the VEGF(165)- and VEGF(121)-induced mitogen-activated protein (MAP) kinase signalling pathways comprising Raf1, MEK1/2 and ERK1/2: for VEGF(165) it interacts directly or upstream from Raf1; for VEGF(121), it acts downstream from PLCgamma. Finally, the mechanism by which PF-4 may inhibit the endothelial cell proliferation induced by both VEGF(121) and VEGF(165), involving disruption of the MAP kinase signalling pathway downstream from KDR did not seem to involve CXCR3B activation. PMID- 15291809 TI - Relationships between structure, function and stability for pyridoxal 5' phosphate-dependent starch phosphorylase from Corynebacterium callunae as revealed by reversible cofactor dissociation studies. AB - Using 0.4 m imidazole citrate buffer (pH 7.5) containing 0.1 mm l-cysteine, homodimeric starch phosphorylase from Corynebacterium calluane (CcStP) was dissociated into native-like folded subunits concomitant with release of pyridoxal 5'-phosphate and loss of activity. The inactivation rate of CcStP under resolution conditions at 30 degrees C was, respectively, four- and threefold reduced in two mutants, Arg234-->Ala and Arg242-->Ala, previously shown to cause thermostabilization of CcStP [Griessler, R., Schwarz, A., Mucha, J. & Nidetzky, B. (2003) Eur. J. Biochem.270, 2126-2136]. The proportion of original enzyme activity restored upon the reconstitution of wild-type and mutant apo phosphorylases with pyridoxal 5'-phosphate was increased up to 4.5-fold by added phosphate. The effect on recovery of activity displayed a saturatable dependence on the phosphate concentration and results from interactions with the oxyanion that are specific to the quarternary state. Arg234-->Ala and Arg242-->Ala mutants showed, respectively, eight- and > 20-fold decreased apparent affinities for phosphate (K(app)), compared to the wild-type (K(app) approximately 6 mm). When reconstituted next to each other in solution, apo-protomers of CcStP and Escherichia coli maltodextrin phosphorylase did not detectably associate to hybrid dimers, indicating that structural complementarity among the different subunits was lacking. Pyridoxal-reconstituted CcStP was inactive but approximately 60% and 5% of wild-type activity could be rescued at pH 7.5 by phosphate (3 mm) and phosphite (5 mm), respectively. pH effects on catalytic rates were different for the native enzyme and pyridoxal-phosphorylase bound to phosphate and could reflect the differences in pK(a) values for the cofactor 5' phosphate and the exogenous oxyanion. PMID- 15291810 TI - Akt-dependent phosphorylation negatively regulates the transcriptional activity of dHAND by inhibiting the DNA binding activity. AB - HAND2/dHAND is a basic helix-loop-helix transcription factor expressed in the heart and neural crest derivatives during embryogenesis. Although dHAND is essential for branchial arch, cardiovascular and limb development, its target genes have not been identified. The regulatory mechanisms of dHAND function also remain relatively unknown. Here we report that Akt/PKB, a serine/threonine protein kinase involved in cell survival, growth and differentiation, phosphorylates dHAND and inhibits dHAND-mediated transcription. AU5-dHAND expressed in 293T cells became phosphorylated, possibly at its Akt phosphorylation motif, in the absence of kinase inhibitors, whereas the phosphatidylinositol 3-kinase inhibitor wortmannin and the Akt inhibitor NL-71 101, but not the p70 S6 kinase inhibitor rapamycin, significantly reduced dHAND phosphorylation. Coexpression of HA-Akt augmented dHAND phosphorylation at multiple serine and threonine residues mainly located in the bHLH domain and, as a result, decreased the transcriptional activity of dHAND. Consistently, alanine mutation mimicking the nonphosphorylation state abolished the inhibitory effect of Akt on dHAND, whereas aspartate mutation mimicking the phosphorylation state resulted in a loss of dHAND transcriptional activity. These changes in dHAND transcriptional activity were in parallel with changes in the DNA binding activity rather than in dimerization activity. These results suggest that Akt mediated signaling may regulate dHAND transcriptional activity through the modulation of its DNA binding activity during embryogenesis. PMID- 15291811 TI - Substrate specificity and transport mode of the proton-dependent amino acid transporter mPAT2. AB - The PAT2 transporter has been shown to act as an electrogenic proton/amino acid symporter. The PAT2 cDNA has been cloned from various human, mouse and rat tissues and belongs to a group of four genes (pat1 to pat4) with PAT3 and PAT4 still resembling orphan transporters. The first immunolocalization studies demonstrated that the PAT2 protein is found in the murine central nervous system in neuronal cells with a proposed role in the intra and/or intercellular amino acid transport. Here we provide a detailed analysis of the transport mode and substrate specificity of the murine PAT2 transporter after expression in Xenopus laevis oocytes, by electrophysiological techniques and flux studies. The structural requirements to the PAT2 substrates - when considering both low and high affinity type substrates - are similar to those reported for the PAT1 protein with the essential features of a free carboxy group and a small side chain. For high affinity binding, however, PAT2 requires the amino group to be located in an alpha-position, tolerates only one methyl function attached to the amino group and is highly selective for the L-enantiomers. Electrophysiological analysis revealed pronounced effects of membrane potential on proton binding affinity, but substrate affinities and maximal transport currents only modestly respond to changes in membrane voltage. Whereas substrate affinity is dependent on extracellular pH, proton binding affinity to PAT2 is substrate-independent, favouring a sequential binding of proton followed by substrate. Maximal transport currents are substrate-dependent which suggests that the translocation of the loaded carrier to the internal side is the rate-limiting step. PMID- 15291812 TI - A new framework for the estimation of control parameters in metabolic pathways using lin-log kinetics. AB - The control properties of biochemical pathways can be described by control coefficients and elasticities, as defined in the framework of metabolic control analysis. The determination of these parameters using the traditional metabolic control analysis relationships is, however, limited by experimental difficulties (e.g. realizing and measuring small changes in biological systems) and lack of appropriate mathematical procedures (e.g. when the more practical large changes are made). In this paper, the recently developed lin-log approach is proposed to avoid the above-mentioned problems and is applied to estimate control parameters from measurements obtained in steady state experiments. The lin-log approach employs approximative linear-logarithmic kinetics parameterized by elasticities and provides analytical solutions for fluxes and metabolite concentrations when large changes are made. Published flux and metabolite concentration data are used, obtained from a reconstructed section of glycolysis converting 3 phosphoglycerate to pyruvate [Giersch, C. (1995) Eur. J. Biochem. 227, 194-201]. With the lin-log approach, all data from different experiments can be combined to give realistic elasticity and flux control coefficient estimates by linear regression. Despite the large changes, a good agreement of fluxes and metabolite concentrations is obtained between the measured and calculated values according to the lin-log model. Furthermore, it is shown that the lin-log approach allows a rigorous statistical evaluation to identify the optimal reference state and the optimal model structure assumption. In conclusion, the lin-log approach addresses practical problems encountered in the traditional metabolic control analysis based methods by introducing suitable nonlinear kinetics, thus providing a novel framework with improved procedures for the estimation of elasticities and control parameters from large perturbation experiments. PMID- 15291813 TI - Neuroserpin Portland (Ser52Arg) is trapped as an inactive intermediate that rapidly forms polymers: implications for the epilepsy seen in the dementia FENIB. AB - The dementia familial encephalopathy with neuroserpin inclusion bodies (FENIB) is caused by point mutations in the neuroserpin gene. We have shown a correlation between the predicted effect of the mutation and the number of intracerebral inclusions, and an inverse relationship with the age of onset of disease. Our previous work has shown that the intraneuronal inclusions in FENIB result from the sequential interaction between the reactive centre loop of one neuroserpin molecule with beta-sheet A of the next. We show here that neuroserpin Portland (Ser52Arg), which causes a severe form of FENIB, also forms loop-sheet polymers but at a faster rate, in keeping with the more severe clinical phenotype. The Portland mutant has a normal unfolding transition in urea and a normal melting temperature but is inactive as a proteinase inhibitor. This results in part from the reactive loop being in a less accessible conformation to bind to the target enzyme, tissue plasminogen activator. These results, with those of the CD analysis, are in keeping with the reactive centre loop of neuroserpin Portland being partially inserted into beta-sheet A to adopt a conformation similar to an intermediate on the polymerization pathway. Our data provide an explanation for the number of inclusions and the severity of dementia in FENIB associated with neuroserpin Portland. Moreover the inactivity of the mutant may result in uncontrolled activity of tissue plasminogen activator, and so explain the epileptic seizures seen in individuals with more severe forms of the disease. PMID- 15291814 TI - Cdt1 and geminin are down-regulated upon cell cycle exit and are over-expressed in cancer-derived cell lines. AB - Licensing origins for replication upon completion of mitosis ensures genomic stability in cycling cells. Cdt1 was recently discovered as an essential licensing factor, which is inhibited by geminin. Over-expression of Cdt1 was shown to predispose cells for malignant transformation. We show here that Cdt1 is down-regulated at both the protein and RNA level when primary human fibroblasts exit the cell cycle into G0, and its expression is induced as cells re-enter the cell cycle, prior to S phase onset. Cdt1's inhibitor, geminin, is similarly down regulated upon cell cycle exit at both the protein and RNA level, and geminin protein accumulates with a 3-6 h delay over Cdt1, following serum re-addition. Similarly, mouse NIH3T3 cells down-regulate Cdt1 and geminin mRNA and protein when serum starved. Our data suggest a transcriptional control over Cdt1 and geminin at the transition from quiescence to proliferation. In situ hybridization and immunohistochemistry localize Cdt1 as well as geminin to the proliferative compartment of the developing mouse gut epithelium. Cdt1 and geminin levels were compared in primary cells vs. cancer-derived human cell lines. We show that Cdt1 is consistently over-expressed in cancer cell lines at both the protein and RNA level, and that the Cdt1 protein accumulates to higher levels in individual cancer cells. Geminin is similarly over-expressed in the majority of cancer cell lines tested. The relative ratios of Cdt1 and geminin differ significantly amongst cell lines. Our data establish that Cdt1 and geminin are regulated at cell cycle exit, and suggest that the mechanisms controlling Cdt1 and geminin levels may be altered in cancer cells. PMID- 15291815 TI - GlnK effects complex formation between NifA and NifL in Klebsiella pneumoniae. AB - In Klebsiella pneumoniae, the nif specific transcriptional activator NifA is inhibited by NifL in response to molecular oxygen and ammonium. Here, we demonstrate complex formation between NifL and NifA (approximately 1 : 1 ratio), when synthesized in the presence of oxygen and/or ammonium. Under simultaneous oxygen- and nitrogen-limitation, significant but fewer NifL-NifA complexes (approximately 1%) were formed in the cytoplasm as a majority of NifL was sequestered to the cytoplasmic membrane. These findings indicate that inhibition of NifA in the presence of oxygen and/or ammonium occurs via direct NifL interaction and formation of those inhibitory NifL-NifA complexes appears to be directly and exclusively dependent on the localization of NifL in the cytoplasm. We further observed evidence that the nitrogen sensory protein GlnK forms a trimeric complex with NifL and NifA under nitrogen limitation. Binding of GlnK to NifL-NifA was specific; however the amount of GlnK within these complexes was small. Finally, two lines of evidence were obtained that under anaerobic conditions but in the presence of ammonium additional NtrC-independent GlnK synthesis inhibited the formation of stable inhibitory NifL-NifA complexes. Thus, we propose that the NifL-NifA-GlnK complex reflects a transitional structure and hypothesize that under nitrogen-limitation, GlnK interacts with the inhibitory NifL-NifA complex, resulting in its dissociation. PMID- 15291816 TI - Differential interactions of decorin and decorin mutants with type I and type VI collagens. AB - The small leucine-rich proteoglycan decorin can bind via its core protein to different types of collagens such as type I and type VI. To test whether decorin can act as a bridging molecule between these collagens, the binding properties of wild-type decorin, two full-length decorin species with single amino acid substitutions (DCN E180K, DCN E180Q), which previously showed reduced binding to collagen type I fibrils, and a truncated form of decorin (DCN Q153) to the these collagens were investigated. In a solid phase assay dissociation constants for wild-type decorin bound to methylated, therefore monomeric, triple helical type I collagen were in the order of 10(-10) m, while dissociation constants for fibrillar type I collagen were approximately 10(-9) m. The dissociation constant for type VI was approximately 10(-7) m. Using real-time analysis for a more detailed investigation DCN E180Q and DCN E180K exhibited lower association and higher dissociation constants to type I collagen, compared to wild-type decorin, deviating by at least one order of magnitude. In contrast, the affinities of these mutants to type VI collagen were 10 times higher than the affinity of wild type decorin (K(D) approximately 10(-8) m). Further investigations verified that complexes of type VI collagen and decorin bound type I collagen and that the affinity of collagen type VI to type I was increased by the presence of decorin. These data show that decorin not only can regulate collagen fibril formation but that it also can act as an intermediary between type I and type VI collagen and that these two types of collagen interact via different binding sites. PMID- 15291817 TI - NMR and molecular dynamics studies of an autoimmune myelin basic protein peptide and its antagonist: structural implications for the MHC II (I-Au)-peptide complex from docking calculations. AB - Experimental autoimmune encephalomyelitis can be induced in susceptible animals by immunodominant determinants of myelin basic protein (MBP). To characterize the molecular features of antigenic sites important for designing experimental autoimmune encephalomyelitis suppressing molecules, we report structural studies, based on NMR experimental data in conjunction with molecular dynamic simulations, of the potent linear dodecapeptide epitope of guinea pig MBP, Gln74-Lys75-Ser76 Gln77-Arg78-Ser79-Gln80-Asp81-Glu82-Asn83-Pro84-Val85 [MBP(74-85)], and its antagonist analogue Ala81MBP(74-85). The two peptides were studied in both water and Me(2)SO in order to mimic solvent-dependent structural changes in MBP. The agonist MBP(74-85) adopts a compact conformation because of electrostatic interactions of Arg78 with the side chains of Asp81 and Glu82. Arg78 is 'locked' in a well-defined conformation, perpendicular to the peptide backbone which is practically solvent independent. These electrostatic interactions are, however, absent from the antagonist Ala81MBP(74-85), resulting in great flexibility of the side chain of Arg78. Sequence alignment of the two analogues with several species of MBP suggests a critical role for the positively charged residue Arg78, firstly, in the stabilization of the local microdomains (epitopes) of the integral protein, and secondly, in a number of post-translational modifications relevant to multiple sclerosis, such as the conversion of charged arginine residues to uncharged citrullines. Flexible docking calculations on the binding of the MBP(74-85) antigen to the MHC class II receptor site I-A(u) using haddock indicate that Gln74, Ser76 and Ser79 are MHC II anchor residues. Lys75, Arg78 and Asp81 are prominent, solvent-exposed residues and, thus, may be of importance in the formation of the trimolecular T-cell receptor-MBP(74-85)-MHC II complex. PMID- 15291818 TI - Val216 decides the substrate specificity of alpha-glucosidase in Saccharomyces cerevisiae. AB - Differences in the substrate specificity of alpha-glucosidases should be due to the differences in the substrate binding and the catalytic domains of the enzymes. To elucidate such differences of enzymes hydrolyzing alpha-1,4- and alpha-1,6-glucosidic linkages, two alpha-glucosidases, maltase and isomaltase, from Saccharomyces cerevisiae were cloned and analyzed. The cloned yeast isomaltase and maltase consisted of 589 and 584 amino acid residues, respectively. There was 72.1% sequence identity with 165 amino acid alterations between the two alpha-glucosidases. These two alpha-glucosidase genes were subcloned into the pKP1500 expression vector and expressed in Escherichia coli. The purified alpha-glucosidases showed the same substrate specificities as those of their parent native glucosidases. Chimeric enzymes constructed from isomaltase by exchanging with maltase fragments were characterized by their substrate specificities. When the consensus region II, which is one of the four regions conserved in family 13 (alpha-amylase family), is replaced with the maltase type, the chimeric enzymes alter to hydrolyze maltose. Three amino acid residues in consensus region II were different in the two alpha-glucosidases. Thus, we modified Val216, Gly217, and Ser218 of isomaltase to the maltase-type amino acids by site-directed mutagenesis. The Val216 mutant was altered to hydrolyze both maltose and isomaltose but neither the Gly217 nor the Ser218 mutant changed their substrate specificity, indicating that Val216 is an important residue discriminating the alpha-1,4- and 1,6-glucosidic linkages of substrates. PMID- 15291819 TI - How calcium inhibits the magnesium-dependent enzyme human phosphoserine phosphatase. AB - The structure of the Mg(2+)-dependent enzyme human phosphoserine phosphatase (HPSP) was exploited to examine the structural and functional role of the divalent cation in the active site of phosphatases. Most interesting is the biochemical observation that a Ca(2+) ion inhibits the activity of HPSP, even in the presence of added Mg(2+). The sixfold coordinated Mg(2+) ion present in the active site of HPSP under normal physiological conditions, was replaced by a Ca(2+) ion by using a crystallization condition with high concentration of CaCl(2) (0.7 m). The resulting HPSP structure now shows a sevenfold coordinated Ca(2+) ion in the active site that might explain the inhibitory effect of Ca(2+) on the enzyme. Indeed, the Ca(2+) ion in the active site captures both side-chain oxygen atoms of the catalytic Asp20 as a ligand, while a Mg(2+) ion ligates only one oxygen atom of this Asp residue. The bidentate character of Asp20 towards Ca(2+) hampers the nucleophilic attack of one of the Asp20 side chain oxygen atoms on the phosphorus atom of the substrate phosphoserine. PMID- 15291820 TI - Analysis of the transcarbamoylation-dehydration reaction catalyzed by the hydrogenase maturation proteins HypF and HypE. AB - The hydrogenase maturation proteins HypF and HypE catalyze the synthesis of the CN ligands of the active site iron of the NiFe-hydrogenases using carbamoylphosphate as a substrate. HypE protein from Escherichia coli was purified from a transformant overexpressing the hypE gene from a plasmid. Purified HypE in gel filtration experiments behaves predominantly as a monomer. It does not contain statistically significant amounts of metals or of cofactors absorbing in the UV and visible light range. The protein displays low intrinsic ATPase activity with ADP and phosphate as the products, the apparent K(m) being 25 micro m and the k(cat) 1.7 x 10(-3) s(-1). Removal of the C-terminal cysteine residue of HypE which accepts the carbamoyl moiety from HypF affected the K(m) (47 micro m) but not significantly the k(cat) (2.1 x 10(-3) s(-1)). During the carbamoyltransfer reaction, HypE and HypF enter a complex which is rather tight at stoichiometric ratios of the two proteins. A mutant HypE variant was generated by amino acid replacements in the nucleoside triphosphate binding region, which showed no intrinsic ATPase activity. The variant was active as an acceptor in the transcarbamoylation reaction but did not dehydrate the thiocarboxamide to the thiocyanate. The results obtained with the HypE variants and also with mutant HypF forms are integrated to explain the complex reaction pattern of protein HypF. PMID- 15291821 TI - Functional properties of the protein disulfide oxidoreductase from the archaeon Pyrococcus furiosus: a member of a novel protein family related to protein disulfide-isomerase. AB - Protein disulfide oxidoreductases are ubiquitous redox enzymes that catalyse dithiol-disulfide exchange reactions with a CXXC sequence motif at their active site. A disulfide oxidoreductase, a highly thermostable protein, was isolated from Pyrococcus furiosus (PfPDO), which is characterized by two redox sites (CXXC) and an unusual molecular mass. Its 3D structure at high resolution suggests that it may be related to the multidomain protein disulfide-isomerase (PDI), which is currently known only in eukaryotes. This work focuses on the functional characterization of PfPDO as well as its relation to the eukaryotic PDIs. Assays of oxidative, reductive, and isomerase activities of PfPDO were performed, which revealed that the archaeal protein not only has oxidative and reductive activity, but also isomerase activity. On the basis of structural data, two single mutants (C35S and C146S) and a double mutant (C35S/C146S) of PfPDO were constructed and analyzed to elucidate the specific roles of the two redox sites. The results indicate that the CPYC site in the C-terminal half of the protein is fundamental to reductive/oxidative activity, whereas isomerase activity requires both active sites. In comparison with PDI, the ATPase activity was tested for PfPDO, which was found to be cation-dependent with a basic pH optimum and an optimum temperature of 90 degrees C. These results and an investigation on genomic sequence databases indicate that PfPDO may be an ancestor of the eukaryotic PDI and belongs to a novel protein disulfide oxidoreductase family. PMID- 15291822 TI - Functional characterization of the evolutionarily divergent fern plastocyanin. AB - Plastocyanin (Pc) is a soluble copper protein that transfers electrons from cytochrome b(6)f to photosystem I (PSI), two protein complexes that are localized in the thylakoid membranes in chloroplasts. The surface electrostatic potential distribution of Pc plays a key role in complex formation with the membrane-bound partners. It is practically identical for Pcs from plants and green algae, but is quite different for Pc from ferns. Here we report on a laser flash kinetic analysis of PSI reduction by Pc from various eukaryotic and prokaryotic organisms. The reaction of fern Pc with fern PSI fits a two-step kinetic model, consisting of complex formation and electron transfer, whereas other plant systems exhibit a mechanism that requires an additional intracomplex rearrangement step. The fern Pc interacts inefficiently with spinach PSI, showing no detectable complex formation. This can be explained by assuming that the unusual surface charge distribution of fern Pc impairs the interaction. Fern PSI behaves in a similar way as spinach PSI in reaction with other Pcs. The reactivity of fern Pc towards several soluble c-type cytochromes, including cytochrome f, has been analysed by flavin-photosensitized laser flash photolysis, demonstrating that the specific surface motifs for the interaction with cytochrome f are conserved in fern Pc. PMID- 15291823 TI - Determination of skin irritation potential in the human 4-h patch test. AB - Recently adopted legislation in Europe has increased the focus that must be placed on the development of in vitro alternatives to the traditional toxicology tests employed to identify the human health hazards associated with chemicals. Included in these is the rabbit skin-irritation test which is used to discriminate those substances which possess significant acute skin irritation potential from those which are of more limited irritation potential. So far, the considerable efforts to replace this assay with in vitro alternatives have not been successful, which may in part be due to the relatively poor quality of the existing in vivo dataset. To help address this problem, we have elected to present our complete database of information on the skin irritation potential of some 65 substances, all of which have been tested in a standard human 4-h patch test. These provide a high quality dataset, generated in man (the goal of toxicologists' health protection-related activities). The data are presented in the context of results with a concurrent positive skin irritation control to ensure that results from individual experiments can be correlated. Consequently, in vitro or in silico alternatives which can identify the significant acute human skin irritants in this group may well represent suitable alternatives to the rabbit. PMID- 15291824 TI - Allergic contact dermatitis from hair dye and development of lichen simplex chronicus. AB - Those who dye their hair frequently manifest allergic contact dermatitis (ACD) from p-phenylenediamine (PPD)-containing hair dye. PPD is known to be the most frequent sensitizer in hair dye, but there has been no documentation of this sensitizer having a role in chronic dermatologic conditions. Our department experienced a case of a 62-year-old woman with lichen simplex chronicus (LSC), who complained of aggravation after hair dyeing and made such an association. We conducted a prospective and retrospective study to further evaluate the clinical associations between the two. In our prospective study, patch testing was performed in selected patients who regularly carried out hair dyeing and also had clinical manifestations of LSC. Also a retrospective examination was conducted in cases where patch testing had been performed with PPD in the past for suspected ACD and further selected cases with concurrent LSC and/or other neurodermatitis. 11 and 14 patients in our prospective and retrospective study, respectively, presented with both LSC and positive findings to PPD. 5 (45.5%) and 4 (28.6%) patients in our prospective and retrospective study showed clinical relevance from clinical improvement after stopping use and rechallenge. We report several cases of patients diagnosed as having LSC and/or prurigo nodularis who showed clinical improvement after discontinuing the use of hair dye. The suggestion can therefore be made that hair dye could be a possible aetiologic agent causing LSC in those using hair dyes. PMID- 15291825 TI - The validity of the Mathias criteria for establishing occupational causation and aggravation of contact dermatitis. AB - Mathias proposed 7 criteria for establishing occupational causation and aggravation of contact dermatitis (Mathias, J Am Acad Dermatol 1989, 20, 842 848). 4 of the 7 criteria must be positive to conclude occupational dermatitis. In order to evaluate the validity of these criteria, we re-examined 19 patients (17 male and 2 female) with contact dermatitis, who had given a positive answer to at least 4 of the criteria and had either exchanged their workplace or stopped working. We re-evaluated them, 2-5 years later, for the presence of contact dermatitis. 14 of 19 patients (74%) reported that their dermatitis had cleared after they had left their previous workplace. Only 5 patients still suffered from dermatitis, 2-5 years later. 3 of these 5 patients could have been exposed to the causative allergens in their new workplace. We conclude that the Mathias criteria are useful to assess occupational contact dermatitis. PMID- 15291826 TI - Allergic and non-allergic periorbital dermatitis: patch test results of the Information Network of the Departments of Dermatology during a 5-year period. AB - Periorbital dermatitis is common and can be due to the external use of ophthalmic drugs. We evaluated patch test results of the Information Network of the Departments of Dermatology. During a 5-year period (1995-99), of a total 49,256 patch-tested patients, 1053 (2.1%) were eventually diagnosed as allergic periorbital contact dermatitis (APD) and 588 (1.2%) as non-allergic periorbital dermatitis (NAPD). Patient characteristics between APD, NAPD and other cases (OCs) differed with respect to sex (19.7% male in both periorbital groups versus 36.3% in OCs), atopic dermatitis (10.4% in APD versus 60.2% in NAPD versus 16.9% in OCs) and age, APD being substantially more often (68.2%) aged 40 and above than NAPD (52.6%). Several of the top allergens in OCs [such as fragrance mix, Myroxylon pereirae resin (balsam of Peru), lanolin alcohol and potassium dichromate] caused significantly fewer positive test reactions in both periorbital groups. In contrast, thimerosal, phenylmercuric acetate, sodium disulfite, gentamicin sulfate, phenylephrine hydrochloride and benzalkonium chloride tested positively significantly more often in APD but not in NAPD, verifying them as true ophthalmic allergens. Finally, in 42 cases (4%) of APD patients, additional allergens were identified by testing of the patients' own substances (mostly beta-blockers, oxybuprocaine and dexpanthenol), supporting the necessity of testing with ophthalmic drugs as is where individual substances are not readily available. PMID- 15291827 TI - Immediate contact reactions to fragrance mix constituents and Myroxylon pereirae resin. AB - We have studied patients who have positive-patch test reactions to fragrance allergic screening substances fragrance mix (FM) or Myroxylon pereirae resin (balsam of Peru) for immediate contact reactions to the standard FM, the constituents of the FM and Myroxylon pereirae resin. In the fragrance-positive subjects (n = 60), there were positive immediate contact reactions to Myroxylon pereirae resin in 56.6% and to FM in 11.6%. In a control group (n = 50) of eczematous, patch test-negative patients there were positive immediate reactions to Myroxylon pereirae resin in 58.0% subjects and to FM in 12.0%. The absence of a significant difference between the fragrance-allergic group and control group is in keeping with a non-immunological basis for the majority of the immediate reactions seen. PMID- 15291828 TI - Environmental contact factors in eczema and the results of patch testing Chinese patients with a modified European standard series of allergens. AB - Environmental contact factors in eczema were investigated in China by clinical questionnaire and patch testing patients with a modified European standard series of allergens. 217 consecutive eczema patients were studied. Contact dermatitis (CD) was clinically diagnosed in 30% of the patients. Among the patients patch tested, 46 patients had clinically diagnosed allergic CD (ACD), 20 patients clinically had non-ACD (NACD) (including 16 cases of irritant contact dermatitis, 1 case of phototoxic contact reaction and 3 cases of asteatotic eczema) and 115 patients had clinically suspected ACD. 45 patients (98%) in the ACD group went on to have relevant patch test results. The most common ACD was from metals, fragrance materials, cosmetics and rubber materials. The most common contact allergens identified were nickel, fragrance mix, para-phenylenediamine (PPD), carba mix and thimerosal. No adverse reactions were observed to patch testing, except for pruritus in patch-test-positive patients. The positive rate of patch testing in ACD was much higher than that in NACD (98% versus 15%, P < 0.05, chi(2)-test). 60 (28%) patients had facial dermatitis (FD). Among these, 20 (33%) were confirmed as having ACD. 48 (22%) patients had hand dermatitis (HD). Among these, 7 (15%) were confirmed as having ACD. Fewer patients were confirmed as having ACD in the HD group than in the FD (15% versus 33%, P < 0.05, chi(2) test). Although the difference was not significant, the total positivity rate in the HD group (55%) was lower than in the other groups. 65 (30%) patients had unclassified endogenous eczema (UEE). The total positive rate of patch testing in the UEE group (56%) was no different from that in the FD or HD groups. However, the relevance of positive patch tests was hard to determine in UEE. These results indicate that CD is common in eczema; relatively more patients with FD have ACD, while other factors, such as irritation, may play more of a role in HD. The total positive rate of patch testing in the UEE group was no different from that in the FD or HD groups, suggesting that patch testing should be stressed in UEE and the relevance of positive patch test results in UEE should be studied further. It is effective and safe to patch eczema patients with a modified European standard series of allergens in China. PMID- 15291830 TI - The use of complementary medicine by patients referred to a contact dermatitis clinic. AB - Our aim was to quantify and qualify the use of complementary medicine (CM) by patients referred to our contact dermatitis clinic in Leicester, UK. A face-to face structured questionnaire study was made of 109 consecutive patients referred to the contact dermatitis clinic. 109 such questionnaires were completed. 21/109 (20%) of patients were Indo-Asian and 86/109 (79%) white Caucasian. 33/109 (30%) had tried some form of CM to treat their skin condition. This use was higher in the Indo-Asian group, where 13/21 (62%) had tried some form of CM. 21/33 (63%) who had used CM were happy to recommend it to other patients with skin disease, even though only 10/33 (30%) of these reported an improvement in their skin condition while using CM. The most frequently used CM treatments were herbal medicine [17/33 (51%)], Chinese herbal medicine [6/33 (18%)], traditional Indian medicine [4/33 (12%)] and aromatherapy [6/33 (18%)]. These proportions were similar in all ethnic groups. In a population of adults referred to a contact dermatitis clinic in a city-centre teaching-hospital dermatology department in Leicester, UK, 30% use, or intend to use, CM and this use is associated with ethnicity. PMID- 15291829 TI - Intra-individual variation of irritant threshold and relationship to transepidermal water loss measurement of skin irritation. AB - Irritant susceptibility studies have used either visual assessment or transepidermal water loss (TEWL) to determine subject response. We have developed a visual assessment method which determines subject irritant threshold. We examined the relationship between sodium lauryl sulfate (SLS) irritant threshold and TEWL measurements from normal skin and SLS patch tests. 19 subjects were recruited. The irritant threshold of each subject was measured and TEWL measurements made from the applied SLS patch tests. Individuals with a lower irritant threshold (easily irritated skin) had elevated TEWL levels compared to those with higher thresholds. The irritant threshold test had a low intraindividual variation. This study showed that the 2 methods grouped patients in a similar manner. The variation seen may reflect the different outcomes measured: irritant threshold visually assesses the skin inflammatory response while TEWL measures skin barrier modification. PMID- 15291831 TI - Occupational contact dermatitis with rhinoconjunctivitis due to Tilia cordata and colophonium exposure in a cosmetician. PMID- 15291832 TI - Allergic contact dermatitis due to methyl salicylate in a compress. PMID- 15291833 TI - Occupational nail fragility in a professional violist. PMID- 15291834 TI - Allergic contact dermatitis from gold on a hearing-aid mould. PMID- 15291835 TI - Severe irritant contact dermatitis from Cypress spurge. PMID- 15291836 TI - Curcumin allergy in relation to yellow chlorhexidine solution used for skin disinfection prior to surgery. PMID- 15291837 TI - Erythema multiforme associated with probable contact dermatitis from epichlorohydrin. PMID- 15291838 TI - Allergic contact dermatitis due to 1,3-butylene glycol in medicaments. PMID- 15291839 TI - Contact dermatitis from turpentine in a painter. PMID- 15291840 TI - Occupational protein contact dermatitis from milk proteins. PMID- 15291841 TI - Reproducibility of patch tests. PMID- 15291843 TI - Conversations. PMID- 15291844 TI - Reforming the clinical negligence system. PMID- 15291845 TI - Atrial fibrillation and the urologist. PMID- 15291846 TI - Virtual reality in urology. PMID- 15291847 TI - Smoking and erectile dysfunction: the role of NADPH oxidase. PMID- 15291848 TI - Laparoscopic nephroureterectomy for upper tract transitional cell carcinoma: a critical appraisal. PMID- 15291849 TI - Laparoscopic pyeloplasty: the first decade. PMID- 15291850 TI - State-of-the-art approaches to detecting early bone metastasis in prostate cancer. PMID- 15291851 TI - How is the lower urinary tract affected by gynaecological surgery? PMID- 15291852 TI - Diagnosis and management of ureteric injury: an evidence-based analysis. PMID- 15291853 TI - Laparoscopic cytoreductive nephrectomy for metastatic renal cell carcinoma. AB - OBJECTIVE: To critically analyse the results of laparoscopic cytoreductive surgery for renal cell carcinoma (RCC), as phase III evidence supports cytoreductive nephrectomy before immunotherapy, and there is an overall shift towards minimally invasive renal surgery for this disease. PATIENTS AND METHODS: Since October 2000, 22 patients were treated by laparoscopic cytoreductive nephrectomy for metastatic RCC (group 1). All patients had radiological evidence of metastatic disease, with biopsy confirmation in 10. To put the results into perspective, 25 consecutive contemporary patients with large organ-confined nonmetastatic RCC (>7 cm, clinical stage T2) undergoing laparoscopic radical nephrectomy (group 2) were compared retrospectively. The baseline demographics were comparable between the groups. RESULTS: The mean tumour size was 8 cm in group 1 and 9.6 cm in group 2 (P = 0.07). Variables during and after surgery were comparable between the groups, with a mean operative duration of 3.1 vs 3.2 h (P = 0.82), blood loss of 285 vs 308 mL (P = 0.79), complications in two vs eight (P = 0.08), morphine sulphate equivalent requirements of 51.7 vs 44.1 mg (P = 0.1) and a median length of hospital stay of 1.7 vs 1.6 days (P = 0.68). In group 1 the median (range) time to immunotherapy was 35 (13-136) days. CONCLUSIONS: Laparoscopic cytoreductive nephrectomy is safe and effective in selected patients. Currently the procedure is offered to candidates eligible for immunotherapy and with tumours of < or = 15 cm, and no evidence of adjacent organ invasion or inferior vena caval thrombus. Significant perihilar adenopathy and numerous parasitic vessels can increase the complexity of the surgery. Adequate laparoscopic experience is necessary. PMID- 15291854 TI - Wide variation of prostate-specific antigen doubling time of untreated, clinically localized, low-to-intermediate grade, prostate carcinoma. AB - OBJECTIVE: To assess the prostate specific antigen (PSA) doubling time of untreated, clinically localized, low-to-intermediate grade prostate carcinoma. PATIENTS AND METHODS: A prospective single-arm cohort study has been in progress since November 1995 to assess the feasibility of a watchful-observation protocol with selective delayed intervention for clinically localized, low-to-intermediate grade prostate adenocarcinoma. The PSA doubling time was estimated from a linear regression of ln(PSA) against time, assuming a simple exponential growth model. RESULTS: As of March 2003, 231 patients had at least 6 months of follow-up (median 45) and at least three PSA measurements (median 8, range 3-21). The distribution of the doubling time was: < 2 years, 26 patients; 2-5 years, 65; 5 10 years, 42; 10-20 years, 26; 20-50 years, 16; >50 years, 56. The median doubling time was 7.0 years; 42% of men had a doubling time of >10 years. CONCLUSIONS: The doubling time of untreated clinically localized, low-to intermediate grade prostate cancer varies widely. PMID- 15291855 TI - Radionuclide bone scintigraphy in patients with biochemical recurrence after radical prostatectomy: when is it indicated? AB - OBJECTIVE: To evaluate the use of radionuclide bone scintigraphy following biochemical recurrence after radical retropubic prostatectomy (RRP) for localized prostate cancer. PATIENTS AND METHODS: Of 1197 patients undergoing RRP we identified those with biochemical recurrence and who had also had a bone scan. Biochemical recurrence was defined as a prostate specific antigen (PSA) level of > or = 0.4 ng/mL. Patients with indeterminate bone scan findings and those in whom the interval between the PSA test and the bone scan was >3 months were excluded. Patient age, PSA level and other relevant pathological details were recorded. Clinical symptoms at the time of bone scan, androgen deprivation after RRP, bone scintigram details and time to recurrence were documented. RESULTS: Of the 1197 patients, 153 (12.8%) had a biochemical recurrence and 35 (23%) of these had a total of 44 bone scans taken over a mean (sd) follow-up of 70.4 (35.6) months; 34 (77%) bone scans were negative (group 1) and 10 (33%) positive (group 2). In group 1 the mean PSA at the bone scan was 5.2 ng/mL; 76% of the patients had a PSA of < 7 ng/mL. In group 2 the mean PSA at the bone scan was 30.7 ng/mL and all patients had a PSA of >7 ng/mL. The only significant difference between the groups was the PSA at the time of the bone scan (P < 0.001). CONCLUSION: Bone scintigraphy is a sensitive diagnostic tool for detecting prostate cancer metastases to bone. A bone scan in patients with a serum PSA of <7 ng/mL on biochemical recurrence after RRP is unlikely to be positive, whereas a PSA of > or = 20 ng/mL is. The presence of skeletal symptoms or a PSA level of >7 ng/mL should prompt the clinician to obtain a bone scintigram. PMID- 15291856 TI - The association between patient age and prostate cancer stage and grade at diagnosis. AB - OBJECTIVES: To investigate the association of age with prostate cancer stage and grade, as the latter factors at the time of diagnosis influence management and prognosis, with some studies suggesting that they may change as a function of patient age. PATIENTS AND METHODS: The charts were reviewed of an age-stratified (<60, 60-69, 70-79 and > or = 80 years) random sample of men with newly diagnosed, histologically confirmed prostate cancer in 1995/96 from three geographical areas in Ontario, Canada. Patients were identified using a comprehensive cancer registry, and the chi-square analysis used to examine the relationship between age and stage, logistic regression for the effect of age on clinically localized disease, and linear regression to assess the age and grade relationships. RESULTS: In all, 347 charts were reviewed; more men in the oldest group had T1 and metastatic disease than had younger men (P = 0.034). The proportion of patients with clinically localized disease (T1 and T2) did not change with age (P > 0.10). Tumour grade, as assessed by Gleason score, increased slightly with age (R(2) = 0.017, P = 0.011). Excluding those patients diagnosed by transurethral prostatectomy did not influence either the age/stage or age/grade relationship. Adjusting for prostate-specific antigen level attenuated the age/grade relationship. CONCLUSION: The stage and grade of prostate cancer at diagnosis changes only slightly with age, probably because of a lower intensity of screening and later diagnosis in older men, rather than any change in prostate cancer biology with age. PMID- 15291857 TI - Increasing prostate biopsy cores based on volume vs the sextant biopsy: a prospective randomized controlled clinical study on cancer detection rates and morbidity. AB - OBJECTIVE: To determine if a volume-adjusted increase in the number of biopsy cores could detect more prostate cancers than the standard sextant biopsy alone, without increasing morbidity, and to determine its applicability in Malaysian patients, as a standard sextant biopsy misses 20-25% of prostate malignancies. PATIENTS AND METHODS: In a prospective randomized study of patients undergoing transrectal ultrasonography (TRUS)-guided biopsy for a prostate-specific antigen (PSA) level of 4-20 ng/mL without abnormal digital rectal examination (DRE), the men were divided into five main groups (A-E) with prostate volumes of <20, 20-40, 40-60, 60-80 and >80 mL, respectively. Patients in groups B-E were randomized into sextant (B1 to E1) and increased biopsy-core subgroups, i.e. B2 (eight cores), C2 (10 cores), D2 (12 cores) and E2 (14 cores). The morbidity profile was also evaluated during and after TRUS biopsy, assessing a pain score, rectal bleeding, haematuria, haemospermia and development of fever. In all, 132 patients were recruited (mean age 67.8 years; mean PSA 9.41 ng/mL). RESULTS: The overall cancer detection rate was 24% (32 men). Taking more cores detected 65.5% of cancers, and the sextant biopsy 34.5% (P = 0.0025), but did not increase the overall morbidity. CONCLUSIONS: The volume-adjusted, increased-core regimen significantly increased the positive biopsy rate of TRUS-guided prostate biopsies with no added morbidity. PMID- 15291858 TI - Superficial papillary urothelial carcinomas in young and elderly patients: a comparative study. AB - OBJECTIVE: To compare the clinicopathological and immunohistochemical findings of superficial papillary transitional cell carcinomas in "young" and "elderly" patients, as the natural history and prognosis of bladder tumours in young patients remains a matter of debate. PATIENTS AND METHODS: Tumours from 50 patients with superficial urothelial tumours of the bladder diagnosed before 45 years old ("young" group, follow-up 25-119 months) were compared with 90 similar tumours developed in patients aged >55 years ("elderly", follow-up 24-102 months). All the patients had a transurethral resection with curative intent, and none had received any therapy before surgery. After surgery only patients diagnosed with pT1 tumours were treated by intravesical bacille Calmette-Guerin (BCG) instillations; all received intravesical BCG if there was a recurrence. The clinicopathological variables, recurrence and disease-free interval to recurrence were assessed. Proliferative activity (MIB-1) and expression of cell-cycle regulation proteins cyclin D1, p53 and p27(kip1) were detected by immunohistochemistry in the tumours of both groups. RESULTS There were statistically significant differences in tumour grade, stage and occurrence between the "young" and "elderly" groups. The 'young' group had a longer disease free interval to recurrence. Among the immunohistochemical markers analysed, only MIB-1 and cyclin D1 were associated with an increased risk of recurrence in the "young" group (P < 0.04 and <0.01, respectively) in a univariate analysis. CONCLUSIONS Superficial papillary urothelial tumours of the bladder in "young" patients had a better prognosis than those in the "elderly" group, showing a lower grade and stage at diagnosis, and a lower recurrence rate. Proliferative activity and cyclin D1 expression levels were of prognostic significance for the risk of recurrence in these patients. PMID- 15291859 TI - Early and large-dose intravesical instillation of epirubicin to prevent superficial bladder carcinoma recurrence after transurethral resection. AB - OBJECTIVE: To prospectively compare the prevention of tumour recurrence by four intravesical adjuvant administration protocols, and thus elucidate the efficacy of early and high total dose instillations of epirubicin to prevent superficial bladder tumour recurrence after transurethral resection of bladder tumour (TURBT). PATIENTS AND METHODS: In all, 69 patients with Ta/T1 bladder cancer were randomly assigned to four intravesical administration protocols: A, delayed instillation (first instillation 7 days after TURBT) and low-dose (30 mg once every 2 weeks, six times): B, early instillation (three instillations before 7 days after TURBT) and low-dose; C, delayed and high-dose (30 mg once weekly 12 times) instillation; D, early and high-dose. The influence of the instillation protocols and tumour characteristics on the probability of recurrence-free survival was examined using Kaplan-Meier analysis and a Cox regression hazard model. RESULTS: The early-instillation and high-dose groups had relatively lower recurrence rates after 6 months (A, 30%; B, 25%; C, one of 12; and D, none) and 1 year (50%, 35%, four of nine and one of eight, respectively). Patients who received 360 mg epirubicin (C and D) had a significantly better recurrence-free survival than those receiving 180 mg (A and B; P = 0.012). Preoperative urine cytology and tumour multiplicity were significantly associated with recurrence. However, multivariate analysis of the risk of recurrence using a Cox proportional hazard model showed that urine cytology (hazard ratio 3.11, 95% confidence interval 1.08-8.94, P = 0.04) and total dose (0.32, 0.11-0.92, P = 0.03) were independent prognostic factors for recurrence. CONCLUSION: Patients who received a high-dose epirubicin instillation had a significantly lower recurrence rate but the benefit of early instillation was not confirmed, as the study group was too small. PMID- 15291860 TI - Holmium:YAG laser vaporization of recurrent papillary tumours of the bladder under local anaesthesia. AB - OBJECTIVES: To evaluate holmium:YAG laser vaporization of papillary tumours of the bladder, focusing on surgical technique, patients' satisfaction, complications and cost-effectiveness when using the technique under local anaesthesia as an outpatient procedure. PATIENTS AND METHODS: In all, 52 consecutive patients with recurrent papillary tumours of the bladder were scheduled for holmium:YAG laser vaporization under local anaesthesia using a flexible cystoscope. The number of papillomas and total operative duration was recorded. Patients and surgeons were asked to complete a questionnaire about the procedure. RESULTS: In all, 197 papillomas were successfully vaporized in 88 operations, with a median operative duration of 15 min (5 min per papilloma) and no patient needed treatment under general anaesthesia. Most patients (86%) had no pain (as reported during standard cystoscopy) and none of the procedures was stopped because of pain. All patients would undergo the treatment again, compared with a standard transurethral resection of bladder tumour. The five surgeons rated the procedure as easy in most patients (78%) and difficult in a few (6%). The total cost for the outpatient procedure was less than that for standard treatment. CONCLUSION: This study clearly indicates that holmium:YAG laser vaporization of superficial bladder tumours is feasible, easy and fast, with a high degree of patient satisfaction, and it seems to be an attractive alternative to standard treatment. The procedure has some clear positive socio-economic perspectives in both the short- and long-term. PMID- 15291861 TI - Prevalence of lower urinary tract symptoms in men aged 45-79 years: a population based study of 40 000 Swedish men. AB - OBJECTIVE: To estimate the age-specific prevalence and severity of lower urinary tract symptoms (LUTS) among Swedish men, the intercorrelations between different symptoms, and to assess quality of life and health-seeking behaviour among men with LUTS. SUBJECTS AND METHODS: In 1997, an International Prostate Symptom Score (IPSS) questionnaire, together with other questions about lifestyle, was mailed to all men aged 45-79 years living in two counties in Sweden; the analyses included 39 928 men. RESULTS: Overall, 18.5% and 4.8% of the men were moderately and severely symptomatic; the prevalence of at least one symptom was 83%. LUTS were strongly age-dependent, with 1.8% of severe symptoms among men aged 45-49 years and increasing to 9.7% among those 75-79 years old. Frequent urination was the most common symptom among men aged < 70 years and nocturia among those aged >70 years. Symptoms like hesitancy, poor flow and intermittency were highly correlated with each other (Spearman coefficients 0.56-0.60). There was a high correlation between the IPSS and a poor score for quality of life resulting from the bothersomeness of LUTS (r = 0.70). Among symptomatic subjects, 36% reported a poor quality of life (fairly bad, very bad or terrible). Only 29% of symptomatic subjects (IPSS >7) reported that they had been diagnosed previously for their urinary problems, and only 11% received medication for that. CONCLUSION: Although the prevalence of LUTS in Sweden is high, the percentage of men whose quality of life is substantially affected is much lower. PMID- 15291862 TI - Natural history of lower urinary tract symptoms: preliminary report from a community-based Indian study. AB - OBJECTIVE: To describe the results of an analysis of baseline data from subjects included in the community-screening programme for lower urinary tract symptoms (LUTS) in the Anand and Kheda district in the Gujarat state of India. SUBJECTS AND METHODS: In all, 2406 men aged >40 years were screened in the community in 18 villages. All subjects were given an International Prostate Symptom Score (IPSS) to complete, had a detailed physical examination, uroflowmetry and urine analysis, and were assessed using transabdominal and transrectal ultrasonography. Spearman's correlation coefficient was used to assess the relationships between quality of life (QoL), age, IPSS, maximum flow rate (Q(max)) and prostate volume (PV); stepwise multiple regression was also used. RESULTS: The mean (sd) age of the men was 62.1 (9.5) years. The PV (mean 21.6 mL, sd 10.63) increased linearly with age, while Q(max) (mean 14.6 mL/s, sd 8.4) decreased linearly. The mean (sd) IPSS was 12.2 (8.6) and did not correlate with age but correlated strongly with QoL (coefficient 0.72). Nocturia was the commonest symptom but correlated least with the IPSS and QoL (0.56 and 0.44). The correlations between Q(max), IPSS and PV were weak to moderate. The subjects had a 56% higher risk of developing moderate to severe symptoms if their PV was >25 mL. CONCLUSION: These Indian men had smaller prostates with higher symptom scores than reported in the West. The IPSS was the strongest predictor of QoL. Overall, the correlation between Q(max), IPSS and PV was weak to moderate. PMID- 15291863 TI - Long-term results of sacral neuromodulation for women with urinary retention. AB - OBJECTIVE ; To review the long-term results of sacral nerve stimulation in the treatment of women with Fowler's syndrome, over a 6-year period at one tertiary referral centre. PATIENTS AND METHODS: Between 1996 and 2002, 26 women with urinary retention were treated by implanting a sacral nerve stimulator. Their case records were reviewed for follow-up, complications and revision procedures, and the most recent uroflowmetry results. RESULTS: There were 20 patients (77%) still voiding spontaneously at the time of review (with two having deactivated their stimulator because of pregnancy). Fourteen patients (54%) required revision surgery, and the most common complications included loss of efficacy, implant related discomfort and leg pain. The mean postvoid residual volume was 75 mL and mean maximum flow rate 20.8 mL/s. CONCLUSION: In young women with retention, for whom there is still no alternative to lifelong self-catheterization, sacral neuromodulation is effective for up to 5 years after implantation. However, there was a significant complication rate, in line with other reports, which may be improved by new technical developments. PMID- 15291864 TI - The role of a lipido-sterolic extract of Serenoa repens in the management of lower urinary tract symptoms associated with benign prostatic hyperplasia. AB - OBJECTIVE To examine the clinical profile of medication derived from a lipido sterolic extract of Serenoa repens (saw palmetto) for managing lower urinary tract symptoms (LUTS) associated with benign prostatic hyperplasia (BPH). METHODS We reviewed clinical trials involving extracts of S. repens, focusing on the benefit/risk ratio in patients with BPH. RESULTS S. repens extract significantly reduces the symptoms of BPH, increases urinary flow, improves the quality of life and is well tolerated. CONCLUSION Analysis of the overall clinical database indicates that extract of S. repens may be considered a viable first-line therapy for treating LUTS. PMID- 15291865 TI - Modified ureterosigmoidostomy (Mainz Pouch II) in different age groups and with different techniques of ureteric implantation. AB - OBJECTIVE: To examine the outcome of Mainz Pouch II urinary diversion in different age groups and with different techniques of ureteric implantation. PATIENTS AND METHODS: Between March 1995 and August 2002 a Mainz Pouch II was created in 41 patients (27 male and 14 female, median age 56.3 years, range 2-75) with 81 renal units (RU). For analysis, the patients were divided into 29 (70%) aged <65 years and 12 (30%) aged >65 years. Ureteric implantation with the Goodwin-Hohenfellner (GH) technique was used in 55 RU, with the Abol-Enein (AE) modification in 23 and Le-Duc procedure in three. The median (range) follow-up (available for 36 patients, 88%) was 19 (1-80) months. An unvalidated questionnaire was used to determine specific urinary diversion items. RESULTS: Early complications occurred in 7% of patients, none requiring surgical intervention. Pyelonephritis occurred in five of 36 patients and seven of 71 RU (14% of the patients, 10% of the RU); all patients were <65 years old. In five of seven RU pyelonephritis was caused by the development of upper urinary tract dilatation; none required surgical revision. Ureteric stenosis requiring reimplantation occurred in two RU (2%, one GH, one AE). All patients were continent in the daytime; all but one patient had to wake to urinate at night, with 36% having to do so more than six times. Of the patients, 63% were able to distinguish between stool and urine. Initially, alkalinizing drugs to prevent metabolic acidosis were taken by 30% of the patients. Of previously medicated patients with a follow-up of >1 year, 8% required oral alkalinizing medication. CONCLUSION: The Mainz Pouch II is a safe and reproducible urinary diversion, and serves as a satisfying alternative to other forms of continent urinary diversion in all age groups. The follow-up shows a low complication rate with good results in terms of continence. There were no significant differences in complication rates for the different ureteric implantation techniques. The long-term results remain to be evaluated. PMID- 15291866 TI - Modified S-pouch neobladder vs ileal conduit and a matched control population: a quality-of-life survey. AB - OBJECTIVES: To measure the quality-of-life (QoL) outcome and urinary and sexual function and bother after radical cystectomy and different types of urinary tract reconstruction (Bricker vs modified S-pouch neobladder), also assessing differences between them and a normal population. PATIENTS, SUBJECTS AND METHODS: Two groups of patients with bladder cancer were assessed; group 1 comprised 58 (mean age 65 years, mean follow-up 28 months) with an ileal conduit diversion, and group 2, 50 (mean age 61 years, mean follow-up 26 months) with a modified S pouch neobladder. All were disease-free. Group 3 comprised 54 healthy subjects (a control population) of similar age, gender and comorbidities other than bladder cancer. A QoL questionnaire was used to study changes in QoL, and a specific questionnaire for urinary and sexual function and bother was also constructed. RESULTS: There were no differences in the QoL scores among the three groups; group 3 (control) tended to have a better QoL for all domains except emotional functioning. Urinary function was seriously affected in group 1, with more daytime leakage than in groups 2 and 3 (37.8% vs 10%, P = 0.005, and 9.3%, P = 0.01), night loss of urine (39.5% vs 28%, P = 0.07, and 3.7%, P = 0.002) and urine odour (58.6% vs 4%, and 5.5%, both P = 0.001). Patients in group 2 differed from healthy individuals only in night loss of urine. Consequently urinary bother was more pronounced in group 1, as fewer were satisfied (68.9% vs 86% and 83.2%, both P = 0.03). Sexual function was seriously and similarly affected in groups 1 and 2; the erection rate was 28.9% for group 1, 35.5% for group 2 (P = 0.1) and 83.3% in group 3 (P = 0.003), while firm erections were present at 17.7%, 22.2% (P = 0.2) and 83.3% (P = 0.002). Women reported equivalent dysfunction in all three groups (15.4%, 20% and 16.6%, P = 0.3). Sexual desire was also equal in all groups (48.2%, 50% and 48.1). Patients in group 1 expressed more bother, while those in group 2 seemed more satisfied by their sexual life (84.4%, 68% and 68.5%, P = 0.04). CONCLUSIONS: Radical cystectomy does not affect QoL whichever urinary reconstruction is used, and this implies a determination by the patients to live and adjust to their new conditions. On the contrary, urinary and sexual function are affected and related to the method used to reconstruct the urinary system. PMID- 15291867 TI - Ileal orthotopic neobladder (modified Hautmann) via a shorter detubularized ileal segment: experience and results. AB - OBJECTIVE: To evaluate the clinical, urodynamic, functional, radiological and metabolic results of the ileal (modified Hautmann) orthotopic neobladder over 10 years of experience. PATIENTS AND METHODS: Between January 1992 and March 2002, 124 men (mean age 62.4 years, range 44-76) with advanced bladder cancer had a radical cystoprostatectomy and urinary diversion via an ileal orthotopic neobladder (modified Hautmann). Only 40 cm of small bowel (detubularized ileum) was used to construct the reservoir, as a modification of the method described by Hautmann. All patients were followed periodically and their data recorded. RESULTS: While no patients died during surgery six died (mortality rate was 5%) in the first 30 days afterward (two of them from causes unrelated to the urinary diversion surgery). The early reoperation rate was 14%; there were early complications not requiring surgery in 40 (34%) and later reoperation rate was required in 20.6%. The mean (range) maximum neobladder capacity was 550 (310-720) mL, the maximum intravesical pressure at maximum capacity 26.4 (11-48) cmH(2)O, and the minimum and maximum flow rates 25.2 (16-64) and 17.5 (11-30) mL/s, respectively. Day- and night-time continence rates were 92% and 90% after 4 years. While there was no electrolyte imbalance, there was mild to moderate metabolic acidosis in 58% of patients. There was no urethral tumour recurrence in any patient. CONCLUSION: Detubularization of ileum to form a neobladder gives a more favourable low-pressure and high-capacity reservoir. Therefore, a shorter ileal segment can be used for orthotopic urinary diversion, to avoid various metabolic dysfunctions when using detubularized bowel, but the surgery is not as free of complications as the original technique. PMID- 15291868 TI - Penile revascularization surgery for arteriogenic erectile dysfunction: the long term efficacy rate calculated by survival analysis. AB - OBJECTIVE: To determine the subjective and objective outcomes (by survival analysis) after penile revascularization surgery in patients with arteriogenic erectile dysfunction (ED), selected by established strict criteria. PATIENTS AND METHODS: The study included 51 patients diagnosed with arteriogenic ED caused by localised arterial lesions and who had microscopic penile revascularization surgery between January 1996 and March 2002. Before surgery, all patients had a full examination, including a medical and sexual history, laboratory testing, intracavernosal pharmacological tests, colour Doppler ultrasonography (CDU), pharmacodynamic infusion cavernosometry and cavernosography, and digital subtraction angiography (DSA). Penile revascularization surgery was indicated only in patients aged <50 years and with no history of diabetes mellitus, hypertension or hyperlipidaemia. When there were communicating branches between the dorsal and cavernosal arteries, Hauri's procedure was used; when there were none or there was no evidence for them on both CDU and DSA because of severe narrowing or obstruction in the proximal common penile artery, the Furlow-Fisher modification of the Virag V procedure (FFV5) was used. The patency of the neo arterial blood flow was assessed by CDU and effective rates calculated using the Kaplan-Meier method. The efficacy rate was recalculated whenever there was a recurrence. When occlusion of the neo-arterial blood was confirmed by CDU the date of occlusion was set as that midway between the last examination showing patency of the donor vessel and the latest examination indicating the occlusion. The patency period was the number of days from surgery to the date of occlusion. RESULTS: Of the 51 patients, 26 had Hauri's and 23 the FFV5 procedure (median age 32 years, range 21-49); in two patients with a previous pelvic fracture surgery was not possible because of scar formation in the dorsal area at the base of the penis. The mean (sd) subjectively estimated efficacy rate was 85.9 (6.3)% after 3 and 67.5 (10.7)% after 5 years of follow-up. The duration at 75% efficacy was 41.0 (5.6) months. The objectively estimated efficacy rate was 84.9 (7.3)% at 3 and 65.5 (13.5)% after 5 years of follow-up. The duration at 75% patency was 42.4 (9.5) months, and at 50% was 60.6 (19.4) months. There was no significant difference in subjective outcome between the FFV5 and Hauri procedures (P = 0.38, log rank test) and none objective outcome after surgery (P = 0.19, log rank test). Thirteen of the 18 patients in the Hauri group had venous dilatation in the deep dorsal, obturator, prostatic and the internal iliac veins. There were operative complications in four patients (hyperaemia of the glans in two, and one each with haemorrhage from the anastomosis site and scar contracture). CONCLUSIONS: The long-term efficacy rates (by the Kaplan-Meier method) of the Hauri and FFV5 procedures were both acceptable. The selection criteria gave acceptable outcomes from both procedures. Penile revascularization surgery is a treatment suitable only for young men and therefore attention must be given not only to the long-term outcome but also to long-term adverse events. PMID- 15291869 TI - Living-donor kidney retransplantation: risk factors and outcome. AB - OBJECTIVE: To review the results of kidney retransplantation at our centre. PATIENTS AND METHODS: Between March 1976 and January 2002, 1406 kidneys were transplanted; among these, 54 patients received a second graft (39 men, mean age 32.1 years, sd 8.6). The donors were 48 relatives (mean age 35.4 years, sd 10.1). RESULTS: The mean (sd, range) duration of the first graft was 49.1 (45.9, 1-192) months and the main cause of these grafts failing was immunological. The mean duration of graft failure was 17.3 (10.5, 5-62) months. The rate of histocompatibility leukocyte antigen (HLA)-A, -B >3 was 16.7% and of haplotype DR matching was 11%. The immunosuppression regimen was mainly based on cyclosporin (75%). There were 33 episodes of acute rejection in 23 patients. The major complications were hypertension (70%), infections (30%) and hepatitis (11%). The overall graft and patient survival was good; 15 grafts (27%) were lost during the follow-up of 1-17 years. Ten patients died, five with a functioning graft. Multivariate analysis showed that donor relationship, primary immunosuppression, duration of first graft and serum creatinine level at 1 year were predictors of graft survival. CONCLUSION: Renal retransplantation is the treatment of choice in patients who have lost their graft. The use of related living-donors and potent immunosuppression could help to improve the outcome. PMID- 15291870 TI - Anterior urethral valves and diverticula in children: a result of ruptured Cowper's duct cyst? AB - OBJECTIVE: To review a series of children with anterior urethral valves and diverticula, to elucidate the pathophysiology and optimal management of this entity. PATIENTS AND METHODS: Nine cases (all boys; 1963 to 2003) were reviewed retrospectively. RESULTS: Seven of nine boys had bulbar diverticula. Continuity between Cowper's duct and the diverticulum was noted endoscopically in two and confirmed radiographically in one. Initially, open surgery was curative but more recently endoscopic management has been the procedure of choice. CONCLUSION: This series indicates that the distal lip of a ruptured syringocele may function as a flap-valve, leading to anterior urethral obstruction. Advances in imaging and endoscopic instruments have altered the mode of presentation and management of this entity. PMID- 15291871 TI - Quo vadis? Ureteric reimplantation or ignoring reflux during augmentation cystoplasty. AB - OBJECTIVE: To decide whether antireflux surgery should be used in the presence of vesico-ureteric reflux (VUR) in children, in whom an augmentation procedure is needed, because secondary VUR in children with a neurogenic bladder, infravesical obstruction and primary VUR in the exstrophy-epispadias complex is expected to resolve after augmentation, which decreases the intravesical pressure and increases capacity. PATIENTS AND METHODS: Between 1987 and 2001, the bladder was augmented in 38 children, using no antireflux surgery in group 1 (15 patients) and antireflux surgery in group 2 (23 patients). RESULTS: VUR was detected in all patients on cysto-urethrography before surgery; reflux resolved after augmentation cystoplasty in 97% and 93% of refluxing units in groups 1 and 2, respectively. The increase in the expected bladder capacity was from 35% to 86% in group 1 and from 38% to 90% in group 2. No patient had any deterioration in renal function. CONCLUSIONS: We recommend using only augmentation in patients with low- or high-grade VUR and a neurogenic bladder, infravesical obstruction and exstrophy-epispadias. Combining antireflux surgery with cystoplasty has no significant effect on either the resolution of VUR or renal function. PMID- 15291872 TI - Long-term outcome of the endoscopic correction of vesico-ureteric reflux: a comparison of injected substances. AB - OBJECTIVE: To summarize the long-term outcome of endoscopic surgery to correct vesico-ureteric reflux (VUR) using different injected substances, i.e. autologous blood, hyaluronan/dextranomer copolymer (HDC), PTFE and glutaraldehyde cross linked bovine dermal (GAX) collagen. PATIENTS AND METHODS: Treatment results on 270 ureters of 185 patients followed for >5 years (mean 8.5) were summarized according to the injected substances. The substances were injected into the 6 o'clock position of the ureteric orifice endoscopically. "Success" was defined as the absence of VUR for >5 years after a single injection. RESULTS: The treatment was successful in two of 24 patients (8%) with autologous blood, 17 of 32 (53%) with HDC, 108 of 171 (63%) with PTFE and 24 of 43 (56%) with GAX collagen. The success rate was lower in patients with higher grades of VUR. CONCLUSIONS: Autologous blood is unsuitable for clinical application because of its poor durability. We will no longer use PTFE because its safety is not well established. The overall success rates of endoscopic surgery with GAX collagen and HDC were insufficient compared with surgical reimplantation, but it is advantageous that this procedure is less invasive and can be repeated. The cure rate could be improved by excluding high-grade VUR from the indications for endoscopic surgery. PMID- 15291873 TI - Ballooning of the foreskin and physiological phimosis: is there any objective evidence of obstructed voiding? AB - OBJECTIVES To determine whether physiological phimosis with or without ballooning of the prepuce is associated with noninvasive urodynamic or radiological evidence of bladder outlet obstruction. PATIENTS AND METHODS From August 2001 to October 2002 all boys with a foreskin problem and referred to one paediatric surgeon were assessed in special clinics. Those with physiological phimosis were recruited for the study and had upper tract and bladder ultrasonography (US), followed by uroflowmetry and US-determined postvoid residual urine volumes (PVR). Data were compared between boys with and with no ballooning of the prepuce. The project was approved by the local research ethics committee and informed consent was obtained from all study participants. RESULTS In all, 54 patients were referred for circumcision; 32 boys with physiological phimosis completed the uroflow and US investigations. Ballooning of the foreskin was present in 18 boys (mean age 6.8 years, range 3-12); 14 had physiological phimosis with no ballooning (mean age 6.5 years, range 4-11). Upper tract US and bladder wall thickness were normal in all boys. The mean maximum urinary flow rate (Q(max)) was not significantly different in boys with ballooning and those without (mean 15.3 mL/s, sd 4.4, range 9-24, vs 15.4, sd 2.9, range 10.7-20, P = 0.96). In addition, all Q(max) values were within the normal range when correlated with voided volume and compared with age-related nomograms. Most boys had flow rate patterns showing a normal bell-shaped curve; a few (9%) had subtle changes in the flow-rate profile, with either a plateau-type curve or slow initial increase in flow and prolonged time to achieve Q(max). The two groups had comparable mean PVRs (3.5 mL, sd 5.1, range 0-18 with ballooning vs 6.1, sd 10.7, range 0-38 without, P = 0.37). Only one patient had a marginally abnormal PVR. CONCLUSIONS Physiological phimosis with or without ballooning of the prepuce is not associated with noninvasive objective measures of obstructed voiding. Minor abnormalities in the flow-rate pattern in this patient group deserve further study. PMID- 15291874 TI - Gender assignment in female congenital adrenal hyperplasia: a difficult experience. AB - OBJECTIVE: To report experience of gender (re)assignment in genotypic female (46XX) patients with congenital adrenal hyperplasia (f-CAH), a difficult and stressful experience if complicated with delayed presentation and inadvertent assignment. PATIENTS AND METHODS: Between 1983 and 2002, 70 patients with f-CAH were counselled for gender assignment. The age at diagnosis and operation, the degree of virilization, parental consanguinity, the gender preference of the families, and the factors governing the decision-making process were determined. RESULTS: Forty-one (59%) patients presented after the neonatal period. All parents had already assumed or were advised of a gender for their children, based on the suggestive appearance of the external genitalia. Consequently, 49 patients were reared as female and 21 as "male". Only nine of these "males" could be reassigned as females (mean age at presentation 7.87 months, sd 10.42). Twelve children had to be reared as "male"(mean age at presentation 55.8 months, sd 32.42) in compliance with the parents' and the study group's decision, and appropriate masculinizing reconstructive surgery was undertaken. The difference in the mean age of those reassigned as female and those who remained "male" was significant (P < 0.001). The parental consanguinity rate among the families was especially high in the 'male' patients. CONCLUSIONS It is extremely difficult to correct the gender of patients with f-CAH when they present at >2.5 years old. Furthermore, the delay in diagnosis and the male bias in choice of gender in our population might be a result of strong social pressures on families, influenced by cultural, traditional and economic factors. PMID- 15291875 TI - The role of nitric oxide in bladder urothelial injury after bladder outlet obstruction. AB - OBJECTIVE: To investigate the role of nitric oxide (NO) in the pathogenesis of injury to the bladder mucosa after bladder outlet obstruction (BOO). MATERIALS AND METHODS: The response of bladder mucosa to BOO consists of thickening with fibrous connective tissue; NO is a multipurpose messenger important in blood vessel relaxation, neuronal communication and inflammatory activities of macrophages, and is synthesized by NO synthase (NOS), with three distinct isoforms; inducible (iNOS), endothelial (eNOS) and neuronal (nNOS). Fifteen male guinea pigs had a silk ligature placed around the bladder neck to induce BOO; controls included five sham-operated animals. The animals were killed at 2, 4, 6, 8 and 12 weeks after BOO. NADPH-diaphorase staining was used as a nonspecific method to determine NOS isoform distribution. Single- and double-label immunofluorescence histochemistry for iNOS, eNOS, nNOS and macrophage colony stimulating factor (M-CSF) were used, with laser scanning confocal microscopy to assess the results. The relative amount of iNOS mRNA was evaluated using reverse transcription-polymerase chain reaction (RT-PCR), and a standard method to determine the degree of apoptosis. RESULTS: Bladder mucosa had a markedly increased intensity of NADPH-diaphorase staining 2 weeks after BOO. iNOS immunoreactivity was markedly greater in the bladder mucosa 2 weeks after BOO than in controls. The immunolocalization of iNOS and M-CSF was identical in bladder mucosa after BOO. RT-PCR showed stronger iNOS mRNA expression in the obstructed bladder mucosa 2 weeks after BOO than in controls. eNOS and nNOS expression was detected only 8 weeks after BOO, with markedly more immunoreactivity 12 weeks after initiating BOO. Increased eNOS and nNOS immunoreactivity strongly correlated with the induction of apoptosis in the bladder mucosa. CONCLUSION: There was overexpression of iNOS by activated macrophages in the early stages of BOO. This overexpression did not induce apoptosis in the iNOS-expressing cells of bladder mucosa in the early stages. After long-term BOO bladder mucosa showed strong eNOS and nNOS immunoreactivity, correlating with apoptosis. Thus we suggest that eNOS and nNOS, by triggering cell death, may be important in eliminating hyperplastic urothelial cells, reflecting the plasticity of the bladder response to obstructive stimuli. PMID- 15291876 TI - Caffeic acid phenethyl ester-induced PC-3 cell apoptosis is caspase-dependent and mediated through the loss of inhibitors of apoptosis proteins. AB - OBJECTIVE: To investigate the effects of a novel agent, caffeic acid phethyl ester (CAPE) on nuclear factor (NF)-kappaB activation and apoptosis in the androgen-independent PC3 prostate cancer cell line. MATERIALS AND METHODS: PC-3 cells were assessed for NF-kappaB activation induced by paclitaxel and tumour necrosis factor-alpha (TNF-alpha), using a p65 enzyme-linked immunosorbent assay, with or without CAPE treatment. The corresponding apoptosis was assessed with propidium iodide DNA staining using flow cytometry. The pan-caspase inhibitor Z VAD-FMK was used to investigate the mechanism of apoptosis. Alterations in the expression of inhibitor of apoptosis proteins (IAP), cIAP-1, cIAP-2 and XIAP, were detected using western blot analysis. RESULTS: CAPE prevented paclitaxel and TNFalpha-mediated NF-kappaB activation. Its ability to induce apoptosis in a dose dependent manner was associated with the loss of cIAP-1, cIAP-2 and XIAP expression. Pretreatment with Z-VAD-FMK prevented CAPE-induced apoptosis and the loss of the IAPs. CONCLUSIONS: CAPE is an effective inhibitor of NF-kappaB activation in PC-3 cells, but the mechanism of apoptosis, and the corresponding loss of IAP expression, is caspase-dependent. PMID- 15291877 TI - Fluorescence in situ hybridization analysis of matched primary tumour and lymph node metastasis of D1 (pT2-3pN1M0) prostate cancer. AB - OBJECTIVE: To describe the chromosomal numerical changes present in primary prostate tumours and their matched lymph-node metastases, to identify a clonal cell migration process which could account for the metastatic behaviour. MATERIALS AND METHODS: Twenty-eight cases of unsuspected stage D1 (pT2-3pN1M0) prostate cancer were detected among patients who had a radical prostatectomy for clinically localized prostate cancer. Fluorescence in situ hybridization (FISH), using centromeric probes to enumerate chromosomes 7, 8, 10 and 12, was used to assess numerical chromosomal changes. FISH analysis was used on isolated nuclei obtained from matched primary tumours and their lymph node metastases. RESULTS: Of the 28 suitable cases it was possible to complete the study in 18 pairs of matched tissues; the remainder were excluded because of insufficient tissue or poor preservation of at least one of the tissues. There was cytogenetic change (aneuploidy) in 16 of the 18 primary tumours, the most common being monosomy 8, detected in 14, followed by trisomy 7, in 13 aneuploid tumours. All lymph node metastases were aneuploid by FISH. As in the primary tumours, monosomy 8 and trisomy 7 were the most common cytogenetic alterations, in 13 and 15 of the lymph node tissues. FISH analysis showed a high correlation (83%) in the cytogenetic pattern of changes between the primary tumours and their lymph node metastases. Moreover, a similar number of cells had the most common aneusomies when comparing prostate and the lymph node tissues. CONCLUSIONS: These results show a similar pattern of cytogenetic alteration in the primary tumour and its lymph node metastasis, characterized by the frequent presence of trisomy 7 and monosomy 8, suggesting that clonal cell selection is not involved in the metastatic process. PMID- 15291878 TI - Dendritic cell immunotherapy for urological cancers using cryopreserved allogeneic tumour lysate-pulsed cells: a phase I/II study. AB - OBJECTIVE: To assess the feasibility, toxicity and immunogenicity of dendritic cell (DC)-based immunotherapy in patients with advanced urological cancers. PATIENTS AND METHODS: Patients with hormone-refractory prostate cancer (11) and metastatic renal cell carcinoma (five) received 1-3 x 10(6) intradermal allogeneic tumour lystate-pulsed DCs fortnightly for six vaccinations then monthly until disease progression. Intradermal keyhole limpet haemocyanin was injected near the DCs as the adjuvant. DC vaccine was prepared from buffy coats, then lysate-pulsed, cryopreserved in aliquots, and tested for phenotypic expression and activity in an allogeneic mixed lymphocyte reaction before clinical use. RESULTS: There was no evidence of significant toxicity from vaccine or adjuvant. Delayed-type hypersensitivity skin testing and biopsy revealed a cellular infiltrate to intradermal re-challenge to tumour lysate and adjuvant in almost all patients. In addition, there was increased expression of T helper type 1 cytokines, interferon-gamma-expressing T cell by ELISPOT analysis, but also interleukin-10 in a few patients. Vaccination resulted in a reduction in the level of prostate-specific antigen (PSA) in one patient, a reduction in PSA velocity in a further man and an increased PSA doubling time in six. Two of five patients with renal cell carcinoma had stabilization of disease. CONCLUSION: The cryopreservation and repeated administration of DC vaccine was feasible and not toxic. There was evidence of induction of both humoral and cellular immunity to vaccine and adjuvant in most patients. The use of sequential aliquots of identical cryopreserved vaccine will ensure quality control and greatly facilitate future clinical studies in terms of consistency of vaccine administered and the provision of primed DCs for in vitro assessment of response. PMID- 15291879 TI - Mitochondrial metabolism in the rat during bladder regeneration induced by small intestinal submucosa. AB - OBJECTIVE: To assess mitochondrial metabolism of bladder tissue induced by small intestinal submucosa (SIS), by comparing the mitochondrial enzyme metabolism in this tissue with that in normal bladder tissue and thus evaluate intracellular normality. MATERIAL AND METHODS: In all, 70 rats were grouped into healthy controls (10), surgical controls with a simple bladder incision (15) and rats treated by partial cystectomy with replacement by the SIS graft (45). At 1, 3 and 6 months the rats were killed, the enzymes of mitochondrial respiratory chain complexes assayed, and the respiration of permeabilized bladder fibres assessed using polarographic analysis. RESULTS: The enzyme activities of control and treated rats at 3 months were identical. The results from the polarographic analysis of respiration were also similar to that in normal tissue apart from a decrease in the number of mitochondria. Histologically, there was complete regeneration at 6 months. CONCLUSION: After a phase of inflammation the bladder regenerates after a patch is placed. The new tissue has the same enzymatic and histological features as normal bladder tissue. PMID- 15291880 TI - Expression of transforming growth factor-beta1 in penile tissue from rats with bilateral cavernosal nerve ablation. AB - OBJECTIVE: To explore the expression of transforming growth factor-beta(1) (TGF beta1) in penile tissue from rats after bilateral cavernosal nerve (CN) ablation, mimicking patients who have had no nerve-sparing during prostatectomy. MATERIALS AND METHODS: Ten adult male rats (neurectomy group) had a bilateral CN resection aseptically under an operating microscope, with six sham-operated rats as controls. Fifteen weeks after surgery an apomorphine test was used in all rats to assess penile erection. The penile specimens were then collected and prepared for detecting the expression of TGF-beta1 by reverse transcription-polymerase chain reaction (RT-PCR), western blot and immunohistochemistry, and for quantitative analysis of the ratio of smooth muscle to collagen fibres in the corpus cavernosum with confocal microscopy. RESULTS: All rats in the sham-operated group but none after neurectomy had an erectile response after subcutaneous injection with apomorphine (100 micro g/kg). Immunohistochemistry, RT-PCR and western blot analyses showed a significantly higher expression of TGF-beta1 in the penile tissues after neurectomy than after sham surgery. Smooth muscle cells (fluorescing red) and collagen fibres (green autofluorescence) after paraformaldehyde fixation, were clearly identified by confocal microscopy. The fluorescence intensity expressed as the mean (sem) ratio of smooth muscle to collagen fibres in the corpus cavernosum after neurectomy was 0.265 (0.125), significantly lower than that in the sham-operated group, at 0.760 (0.196) (P < 0.01). CONCLUSION: An increased expression of TGF-beta1 in penile tissue which promotes the synthesis of collagen may be one of the important factors for the erectile dysfunction caused by bilateral CN ablation. Similar pathophysiological processes may occur in the corpus cavernosum of patients after radical prostatectomy. PMID- 15291881 TI - The protective effect of aminoguanidine on erectile function in diabetic rats is not related to the timing of treatment. AB - OBJECTIVE: To assess the accumulation of advanced glycation end products (AGEs) in streptozotocin (STZ)-induced diabetic rat cavernosal tissue, and to determine whether the protective effect of aminoguanidine (AG) on erectile function is related to the timing of treatment, as the accumulation of AGEs in the penis may be important in the pathogenesis of diabetes mellitus-induced erectile dysfunction, and prolonged treatment with AG (a selective AGE inhibitor), prevents erectile dysfunction in this situation. MATERIALS AND METHODS: Harlan Sprague-Dawley rats were divided into groups 1-4, i.e. age-matched controls; STZ diabetic rats (60 mg/kg intraperitoneal) given free access to water; STZ diabetic rats treated with AG (1 g/L per day in the drinking water) immediately after inducing diabetes; and STZ-diabetic rats treated with AG 1 month after inducing diabetes, respectively. Two months after inducing diabetes the intracavernosal pressure was measured after cavernosal nerve stimulation, and cavernosal AGE (5 hydroxy methyl furfural, 5-HMF) levels assessed. RESULTS: Cavernosal tissue 5-HMF levels from groups 2 and 4 were significantly higher than in group 1 (control). The expression of 5-HMF in group 3 was similar to that in group 1. Diabetic rats had significantly lower erectile function than controls, while groups 3 and 4 (treated with AG) had normal erectile function, as measured by cavernosal nerve stimulation. CONCLUSIONS: The effect of AG on AGE levels seems to be time dependent; that the 1-month treatment with AG improved erectile function with no change in AGEs suggests that AG has protective effects on the penile vasculature through alternative pathways. PMID- 15291882 TI - Fibrin glue for the suture-less correction of penile chordee: a pilot study in a rabbit model. AB - OBJECTIVE: To evaluate the use of fibrin glue as a scaffold for patching defects in the tunica albuginea in a rabbit model for a future application in correcting chordee. MATERIALS AND METHODS: Nine New Zealand white male rabbits were utilized. All had a 15 x 5-mm defect created in the ventral tunica albuginea. Fibrin glue (1 mL) was applied to cover the defect in tunica albuginea and the penile skin closed with a continuous 5/0 chromic catgut suture. Animals were killed in groups of three at 2, 6 and 12 weeks afterward. The evaluation included an artificial erection test with intracavernosal injection of prostaglandin E1 (5 microg), cavernosography and histopathological examination of sections of the penis stained with haematoxylin and eosin or Masson trichrome. RESULTS: None of the rabbits died during the procedure or developed bleeding or haematoma afterward. All animals had straight erections on testing with prostaglandin (5 microg). There was no evidence of corporal narrowing or venous leakage on cavernosography. Histopathological evaluation showed evidence of the fibrin sealant layer, with angiogenesis and a cell infiltrate at 2 weeks. At 6 and 12 weeks there was completely normal regeneration of the tunica albuginea. CONCLUSIONS: In this pilot study in a rabbit model the haemostatic effect of fibrin glue was confirmed on covering a defect in the tunica albuginea. Moreover, there was regeneration of normal tunica albuginea with no scarring at 6 weeks and maintained at 12 weeks. Further well-controlled studies are required before using fibrin glue for corporal body grafting to treat chordee. PMID- 15291883 TI - It's new and exciting--can it be AUA 2004? PMID- 15291884 TI - The use of contralateral native pelvis and ureter for total transplant ureteric reconstruction. PMID- 15291885 TI - A novel method to prevent retrograde displacement of ureteric calculi during intracorporeal lithotripsy. PMID- 15291886 TI - Ejaculatory dysfunction and alpha-adrenoceptor antagonists. PMID- 15291889 TI - Robotics in urology. PMID- 15291890 TI - Relaxation effects of adrenomedullin in isolated rabbit corpus cavernosum smooth muscle. PMID- 15291891 TI - Safe control of the renal vein during laparoscopic nephrectomy using the "loop around the vein" technique. PMID- 15291892 TI - The difficult urethral catheterization: use of a hydrophilic guidewire. PMID- 15291893 TI - Radical nerve-sparing laparoscopic prostatectomy. PMID- 15291895 TI - Breaking frontiers for better early allergy diagnosis. PMID- 15291896 TI - Genes of tolerance. AB - Activation-induced cell death, anergy and/or immune response modulation by T regulatory cells (T(Reg)) are essential mechanisms of peripheral T-cell tolerance. There is growing evidence that anergy, tolerance and active suppression are not entirely distinct, but rather, represent linked mechanisms possibly involving the same cells and multiple suppressor mechanisms. Skewing of allergen-specific effector T cells to T(Reg) cells appears as a crucial event in the control of healthy immune response to allergens and successful allergen specific immunotherapy. The T(Reg) cell response is characterized by abolished allergen-induced specific T-cell proliferation and suppressed T helper 1 (Th1)- and Th2-type cytokine secretion. In addition, mediators of allergic inflammation that trigger cAMP-associated G-protein coupled receptors, such as histamine receptor 2 may contribute to peripheral tolerance mechanisms. The increased levels of interleukin-10 (IL-10) and transforming growth factor-beta (TGF-beta) that are produced by T(Reg) cells potently suppress immunoglobulin E (IgE) production, while simultaneously increasing production of noninflammatory isotypes IgG4 and IgA, respectively. In addition, T(Reg) cells directly or indirectly suppress effector cells of allergic inflammation such as mast cells, basophils and eosinophils. In conclusion, peripheral tolerance to allergens is controlled by multiple active suppression mechanisms. It is associated with regulation of antibody isotypes and effector cells to the direction of a healthy immune response and opens a window for novel therapies of allergic diseases. PMID- 15291897 TI - Mouse models of global airway allergy: what have we learned and what should we do next? AB - Recent epidemiological and clinical data indicate that allergic rhinitis and asthma coexist and should be considered as one airway allergy syndrome. In spite of the importance of this new concept of global airway allergy, it has not fundamentally changed our daily diagnostic and therapeutic strategies so far because of the lack of essential clues to understand the correlation between allergic inflammation in upper and lower airways. Because of the resemblance of experimentally induced allergic airway inflammation in mice to inflamed airways of allergic patients, mouse models can enhance our insight into mechanisms underlying the global airway allergy syndrome. We here review data generated in mice that are relevant for understanding the development of airway allergy and provide new options for research on the so-called 'united airway disease'. PMID- 15291898 TI - Control and exacerbation of asthma: a survey of more than 3000 French physicians. AB - BACKGROUND: Asthma control and prevention of exacerbations are primary objectives for asthma care. However there is a lack of universal definition of these notions which may therefore have a different meaning according to the physician's practice. METHODS: In the present survey, 1805 general practitioners, 551 pulmonologists, 176 allergologists and 470 resuscitation/emergency care physicians were randomly selected in fall 2001 and asked to answer one question on asthma control and three questions on asthma exacerbation. RESULTS: Regarding asthma control, the presence of minimum symptoms was the primary criterion for asthma specialists (pulmonologists and allergologists), while a normal respiratory function was first considered by nonspecialist physicians (general practitioners and emergency care physicians). The first criterion of mild exacerbation for asthma specialists was the occurrence during at least 2 days of an increase in the frequency of dyspnoea and/or the use of short-acting bronchodilators. For nonspecialist physicians, dyspnoea affecting daily activities and/or sleeping was the preferred notion. Hospitalization was unanimously recognized as the first criterion for severe exacerbations. A decrease in peak expiratory flow of more than 30% below the baseline value on two consecutive days and an episode requiring systemic corticosteroids were the next criteria. CONCLUSIONS: This survey emphasizes the complexity of the notions of asthma control and exacerbation and provides novel informations to orient continuing education programmes. PMID- 15291899 TI - In vitro effects of flunisolide on MMP-9, TIMP-1, fibronectin, TGF-beta1 release and apoptosis in sputum cells freshly isolated from mild to moderate asthmatics. AB - BACKGROUND: Corticosteroids play an important role in inflammation and remodelling of airways and are considered an important therapeutic target in asthma. Inflammation in asthma is characterized by a dysregulation of eosinophil apoptosis and of markers of airways remodelling. We evaluated the ability of flunisolide to inhibit in vitro the release of metalloproteinases-9 (MMP-9), tissue inhibitor metalloproteinases-1 (TIMP-1), transforming growth factor (TGF beta) and fibronectin by sputum cells (SC) as well as to induce sputum eosinophil apoptosis. METHODS: The SC, isolated from induced sputum samples of 12 mild-to moderate asthmatics, were cultured for 24 h in the presence or absence of flunisolide (1, 10 and 100 microM). The release of mediators was assessed by enzyme-linked immunosorbent assay (ELISA) whereas apoptosis was studied by TUNEL technique. RESULTS: Flunisolide (10 microM) significantly reduced MMP-9 and TIMP 1 (P = 0.0011 and P < 0.0001 respectively) and increased MMP-9/TIMP-1 molar ratio (P = 0.004). In addition, flunisolide decreased TGF-beta and fibronectin release by SC (P = 0.006; and P < 0.0001 respectively) and increased eosinophil apoptosis (P < 0.001). CONCLUSIONS: These results demonstrate that flunisolide may play an important role in the inhibition of airway inflammation and remodelling, by promoting the resolution of eosinophilic inflammation and by inhibiting the release of MMP-9, TIMP-1, TGF-beta and fibronectin. PMID- 15291900 TI - Comparative immunophenotyping of monocytes from symptomatic and asymptomatic atopic individuals. AB - BACKGROUND: Allergy has at least two components - a genetic predisposition referred to as atopy and the progress from an atopic state to clinically apparent disease. Peripheral blood monocytes are circulating myeloid precursors of antigen presenting cells. The expression of cell surface proteins on monocytes may therefore witness the disease status and affect the development of allergic disease. METHODS: Monocytes were isolated from atopic individuals with seasonal allergic rhinitis (n = 10), from atopic individuals sensitized to aeroallergens but without any signs of acute disease (n = 11), and from healthy nonatopic donors (n = 21). Detailed comparative phenotypic analysis of CD14(+) and FcepsilonRI(+)CD14(+) monocytes was performed by flow cytometry. RESULTS: CD14(+) monocytes from symptomatic atopic donors showed a significant increase in the cell surface intensity of the integrin adhesion molecule CD11c over monocytes from asymptomatic atopic and nonatopic donors. Asymptomatic atopic individuals showed significantly enhanced expression of FcepsilonRI on the CD14(high)CD16(dim) monocyte subset compared with this subset from symptomatic atopic and nonatopic donors. CONCLUSION: The increase in monocyte surface intensity of the adhesion molecule CD11c may be involved in the manifestation of allergic disease. FcepsilonRI on CD14(high)CD16(dim) monocytes of asymptomatic atopic donors may be of functional importance for the maintenance of clinical unresponsiveness toward allergens. PMID- 15291901 TI - IgE level and Phadiatop in an elderly population from the PAQUID cohort: relationship to respiratory symptoms and smoking. AB - BACKGROUND: In the last decades in industrialized countries, the increase of life expectancy has resulted in an increase in the population of the elderly. However, little is known about the prevalence of allergies in the elderly population. The aim of the study was to investigate the specific relationship of serum IgE and Phadiatop, with asthma, rhinitis, and smoking in a sample of an elderly population. METHODS: Subjects from the Paquid cohort living in Gironde Department (age 65 years and over) in France were followed up since 1988 (PAQUID cohort). RESULTS: Among the randomized sample of 352 subjects, there were 158 men (45%) and 194 women (55%). The lowest levels of IgE were in subjects with chronic sputum; and in normal subjects (47.1 +/- 56.4 vs 56.2 +/- 73.9, NS). Multiple linear regression was performed with log(10) IgE values as a dependent variable and age, Phadiatop test, smoking, and respiratory symptoms independently in men and in women. In men, a significant relationship was observed between IgE values and Phadiatop test (P < 0.001), asthma history (P = 0.002), and smoking (P = 0.019). CONCLUSIONS: Respiratory allergy is present in persons >65 years of age. There is an association between smoking and IgE level independent of allergic reactivity to common allergens in the elderly. PMID- 15291902 TI - Exposure to house dust endotoxin and allergic sensitization in adults. AB - BACKGROUND: It has been suggested that exposure to elevated levels of endotoxin decreases the risk of allergic sensitization. OBJECTIVE: To examine the associations between current exposure to bacterial endotoxin in house dust and allergic sensitization in adults. METHODS: In 1995-1996, we conducted a nested case-control study following a cross-sectional study performed within the European Community Respiratory Health Survey (ECRHS). Data of 350 adults aged 25 50 years was analysed. Allergic sensitization was assessed by measurement of specific immunoglobulin E (IgE) against several inhalant allergens. Living room floor dust samples were taken. The endotoxin content was quantified using a chromogenic kinetic Limulus amoebocyte lysate test. RESULTS: Multiple logistic regression analysis showed a negative association between exposure to house dust endotoxin and severe allergic sensitization. Odds ratios (95% CI) adjusted for place of residence, gender, age, and 'caseness' were 0.80 (0.64-1.00) for sensitization to >/=1 allergen and 0.72 (0.56, 0.92) for sensitization to >/=2 allergens using 3.5 kU/l as a cut-off value for sensitization. With regard to single allergens, the protective effect of endotoxin was strongest for pollen sensitization [aOR (95% CI) = 0.74 (0.58, 0.93)]. CONCLUSION: Our results indicate that current exposure to higher levels of house dust endotoxin might be associated with a decreased odds of allergic sensitization in adults. PMID- 15291903 TI - Acupuncture and Chinese herbal medicine in the treatment of patients with seasonal allergic rhinitis: a randomized-controlled clinical trial. AB - BACKGROUND: Patients with allergic rhinitis (AR) increasingly use complementary medicine. The aim of this study was to determine whether traditional Chinese therapy is efficacious in patients suffering from seasonal AR. METHODS: Fifty-two patients between the ages of 20 and 58 who had typical symptoms of seasonal AR were assigned randomly and in a blinded fashion to (i) an active treatment group which received a semi-standardized treatment of acupuncture and Chinese herbal medicine, and (ii) a control group which received acupuncture applied to non acupuncture points in addition to a non-specific Chinese herbal formula. All patients received acupuncture treatment once per week and the respective Chinese herbal formula as a decoction three times daily for a total of 6 weeks. Assessments were performed before, during, and 1 week after treatment. The change in severity of hay fever symptoms was the primary outcome measured on a visual analogue scale (VAS). RESULTS: Compared with patients in the control group, patients in the active treatment group showed a significant after-treatment improvement on the VAS (P = 0.006) and Rhinitis Quality of Life Questionnaire (P = 0.015). Improvement on the Global Assessment of Change Scale was noted in 85% of active treatment group participants vs 40% in the control group (P = 0.048). No differences between the two groups could be detected with the Allergic Rhinitis Symptom Questionnaire. Both treatments were well-tolerated. CONCLUSIONS: The results of this study suggest that traditional Chinese therapy may be an efficacious and safe treatment option for patients with seasonal AR. PMID- 15291904 TI - Determinants of neonatal IgE level: parity, maternal age, birth season and perinatal essential fatty acid status in infants of atopic mothers. AB - OBJECTIVE: The hygiene hypothesis suggests that the protective 'siblings effect' against atopic diseases such as atopic dermatitis, allergic asthma and hay fever is a result of recurrent infections during early childhood. A recent study and review have indicated that this protective effect may already arise in utero. Lower n-3 essential fatty acid (EFA) status is associated with increased parity, and EFA status has also been related to atopy. The present study confirms the negative association between parity and neonatal immunoglobulin E (IgE) levels and further unravel the role of perinatal EFA status. METHODOLOGY: In a prospective cohort study in 184 atopic mothers and their neonates, we simultaneously measured serum total IgE and EFA levels in plasma phospholipids, both in the mother at 34-36 weeks of gestation and in the neonate at the age of 1 week. Linear regression analysis was used to estimate the effect of parity on maternal and neonatal IgE and EFA status, and the independent effects of parity and EFA status on IgE, controlling for confounding factors such as maternal age and birth season. RESULTS: Parity was associated with lower neonatal IgE level (P < 0.01), as well as with lower docosahexanoic acid (DHA, 22:6n-3) status of the mother (P = 0.01) but not of the neonate (P > 0.69). In the multivariate analysis, higher parity, higher maternal IgE, lower maternal age and birth in the first 3 months of the year were independently associated with neonatal IgE level. No association was detected between maternal or neonatal EFA status and neonatal IgE. CONCLUSIONS: As neonatal total serum IgE is predictive of later atopy, our results support the hypothesis that the sibling effect in atopy is already being programmed in utero. Our data also confirm earlier findings that DHA status is lower in multiparous women, but this did not confound the relation between parity and neonatal IgE. PMID- 15291905 TI - Efficacy in allergen control and air permeability of different materials used for bed encasement. AB - BACKGROUND: Different types of textile are used for the preparation of covers for bed encasement. The aim of the present study was to evaluate different fabrics employed for mattress covers regarding their efficacy in blocking Der p 1 and Fel d 1 as well as their air permeability. METHODS: Eleven different commercially available fabrics manufactured for allergen avoidance have been tested and compared with regular cotton. Dust samples titered for Der p 1 and Fel d 1 were pulled through the different fabrics using a modified Fussnecker dust trap and collected by a filter located downstream. Airflow through the dust trap was controlled by a vacuum pump operating for 5 min and measured at the beginning (T0) and at the end (T1) of the test. RESULTS: All the tightly woven and laminated materials were able to control mite allergen permeability allowing air passage but they significantly differed in Fel d 1 permeability. Laminated tissues and laminated tissue not tissued were effective in controlling both the allergens but they did not allow air permeability. CONCLUSIONS: Detailed knowledge about the actual properties of the products for bed encasement needs to be considered in order to optimize allergen avoidance, disease control and sleep comfort. PMID- 15291906 TI - The co-seasonal application of anti-IgE after preseasonal specific immunotherapy decreases ocular and nasal symptom scores and rescue medication use in grass pollen allergic children. AB - BACKGROUND: Specific immunotherapy (SIT) and treatment with anti-immunoglobulin (Ig)E antibody are complementary approaches to treat allergic rhinoconjunctivitis, which may be used for single or combined treatment. OBJECTIVE: A randomized, double-blind, placebo-controlled trial was conducted to compare the efficacy of single and combined treatment with SIT and anti-IgE (Omalizumab) in reducing symptom severity and rescue medication use. METHODS: A total of 221 subjects with birch and grass pollen allergic rhinoconjunctivitis aged 6-17 years were analysed during the grass pollen season. Group A (SITbirch + placebo) served as a reference group obtaining no effective treatment for grass pollen allergy. Group B received anti-IgE monotherapy during grass pollen season, group C SIT grass pollen monotherapy, and group D the combined treatment of SIT and Omalizumab. RESULTS: Preseasonal treatment with grass pollen SIT alone compared with SIT with the nonrelated allergen did not reduce symptoms or rescue medication use. Anti-IgE monotherapy significantly diminished rescue medication use and number of symptomatic days. The combined treatment with SIT and anti-IgE showed superior efficacy on symptom severity compared with anti-IgE alone. CONCLUSIONS: Co-seasonal Omalizumab therapy showed considerable effects in children with seasonal allergic rhinitis. The combination of SIT plus Omalizumab was clinically superior to each treatment alone during the first year of observation. PMID- 15291907 TI - A protocol for oral desensitization in children with IgE-mediated cow's milk allergy. AB - OBJECTIVES: To desensitize children with severe immunoglobulin (Ig)E-mediated cow's milk allergy in a period of 6 months by introducing increasing daily doses of cow's milk (CM) in order to enable the child to assume 200 ml of CM daily, or to induce tolerance of the highest possible CM dose. STUDY DESIGN: Twenty-one children at least 6 years old with severe IgE-mediated CM allergy were admitted to the study. A convincing history of IgE-mediated CM allergy or a positive double-blind placebo-controlled food challenge with CM confirmed the diagnosis. Oral desensitization was performed with increasing doses starting from 0.06 mg of CM proteins. RESULTS: Overall, 15 of 21 children (71.4%) achieved the daily intake of 200 ml during a 6-month period; three of 21 children (14.3%) tolerated 40-80 ml/day of undiluted CM; three of 21 children (14.3%) failed the desensitization because they presented allergic symptoms after ingesting minimal amounts of diluted CM. CONCLUSIONS: We successfully desensitized 15 of 21 children with severe IgE-mediated CM allergy in a period of 6 months. We stress the importance of the partial outcome in those three of 21 children who could not reach the maximum amount of 200 ml/day of whole CM, but were able to tolerate 40 80 ml/day of CM. In this way we dramatically reduced the risk of severe reactions after accidental or unnoticed introduction of low quantities of CM. We do not propose generalizing this method beyond trained staff. PMID- 15291908 TI - Birch pollen-related food as a provocation factor of allergic symptoms in children with atopic eczema/dermatitis syndrome. AB - BACKGROUND: Food allergy to cow's milk or hen's egg is a common problem in children with atopic eczema/dermatitis syndrome (AEDS) but the role of birch pollen-related food for the induction of allergic symptoms is still not clear. PATIENTS/METHODS: Twelve children (median age 5 years) with AEDS underwent an oral challenge with those birch pollen-related foods which were reported to induce no immediate symptoms, but were consumed on a regular basis. Total IgE and specific IgE to birch pollen, Bet v 1/2 and various birch pollen-related foods were determined. RESULTS: Seven of 12 children showed immediate and/or late eczematous reactions upon ingestion of birch pollen-related foodstuff. Four children showed a worsening of eczema 24 h upon oral challenge with a significant difference in SCORAD before and after challenge. There were no differences in terms of total IgE or birch pollen-specific IgE between children with a late eczematous response and non-reacting children. CONCLUSIONS: Birch pollen-related food may induce allergic symptoms in children with AEDS who exhibit a sensitization to birch pollen. Oral challenge tests should be performed in those children who suffer from severe AEDS and who are highly sensitized to birch pollen allergens even in the absence of a history suggestive of food allergy. PMID- 15291909 TI - Monoclonal antibody-based ELISA to quantify the major allergen of Artemisia vulgaris pollen, Art v 1. AB - BACKGROUND: Pollen of Artemisia vulgaris (mugwort) is a relevant cause of pollinosis in temperate and humid regions. Recently, the major allergen of this pollen, Art v 1, has been characterized. OBJECTIVE: To develop a monoclonal antibody (mAb)-based enzyme-linked immunosorbent assay (ELISA) to quantify Art v 1, and to assess the correlation of Art v 1 content with the biological activity of mugwort pollen extracts. METHODS: Art v 1-specific mAbs were obtained from a BALB/c mouse immunized with high-performance liquid chromatography (HPLC) purified Art v 1. One of these antibodies (Av 3.7), which recognizes the N terminal defensin-like domain of Art v 1, was used as the capture antibody in an ELISA method for allergen quantitation. An anti-A. vulgaris rabbit serum was used as the second antibody. Art v 1 was purified by immunoaffinity chromatography and used as the standard in the assay. RESULTS: The purity and identity of the affinity-purified Art v 1 was confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), mass spectrometry, amino acid composition, and N terminal amino acid sequencing. The prevalence of specific IgE against Art v 1, determined by radioallergosorbent test (RAST) in a population of 44 mugwort allergic patients, was 79%. The Art v 1-ELISA developed displays a detection limit of 0.1 ng/ml, and a practical working range of 0.2-10 ng/ml. The concentration of Art v 1 was measured in 10 A. vulgaris pollen extracts, and a good correlation was observed between the Art v 1 content and the allergenic activity of the extracts. CONCLUSIONS: The results prove the usefulness of the Art v 1-ELISA for the standardization of A. vulgaris pollen extracts intended for clinical use. PMID- 15291910 TI - Bee moth (Galleria mellonella) allergic reactions are caused by several thermolabile antigens. AB - BACKGROUND: Exposure and contact with bee moth (Galleria mellonella) larvae (Gm) can cause an allergic reaction both in anglers and breeders. We described the case of an amateur fisherman who experienced an allergic reaction using Gm but not using heat-treated Gm (h-Gm) (mummies). The aim of this study was to demonstrate by immunoblotting and radioallergosorbent test (RAST)-inhibition experiments the loss of allergenic epitopes in h-Gm extracts. METHODS: Galleria mellonella larvae and h-Gm were homogenized and extracted at 10% (w/v) in 0.5 M phosphate-buffered saline, pH 7.4 containing 0.5% NaN(3) for 16 h at 4 degrees C. Gm and h-Gm extracts were electrophoresed in a 10% polyacrylamide precast Nupage Bis-Tris gel at 180 mA for 1 h and the resolved proteins stained with 0.1% Coomassie brilliant blue and the molecular weight calculated. For the immunoblotting detection of allergenic components the resolved extracts were transferred onto a nitrocellulose membrane and incubated with the patient's serum. Bound specific-IgE was detected by peroxidase-conjugated anti-human IgE. RAST inhibition experiments were performed according to the Ceska method. RESULTS: The protein profile of Gm and h-Gm extracts resulted markedly different in number, intensity and the position of bands, indicating that heat-treatment modifies the chemical-physical characteristics of the protein contents. The Gm extract showed a strong-coloured band at 73 kDa and more than 20 components ranging from 12 to 133 kDa; h-Gm showed two main band at 77 and 38 kDa and about 15 faint bands between 20 and 133 kDa apparently without any correspondence to the bands present in the Gm extract. Immunoblotting with the patient's serum demonstrated several bands of reactivity with the Gm extract ranging from 20 to 100 kDa and no recognizable bands, but only a diffuse smear with h-Gm. When used in a RAST inhibition experiment the h-Gm extract demonstrated an inability to compete with the Gm one for the binding to patient's IgE serum. CONCLUSIONS: The h-Gm seems to lose the allergenic epitopes and has two advantages for anglers: to avoid new possible sensitizations as well as allergic symptoms in sensitized people, without interfering with their skills and satisfaction in their fishing performance. PMID- 15291911 TI - Sensitization to petrolatum: an unusual cause of false-positive drug patch-tests. AB - We report on an unexpected sensitization to petrolatum diagnosed with the occurrence of multiple nonrelevant and false-positive drug patch-tests performed while investigating a patient suffering from many cutaneous adverse drug reactions. All the positive drug patch-tests were prepared with GILBERT vaseline. This petrolatum reaction is positive as it was tested with five other brands of petrolatums a few months later. As the same petrolatums, but from different batches were tested, patch-tests with GILBERT petrolatum were doubtful, while other petrolatums were positive. White petrolatum is a mixture of semisolid hydrocarbons of the methane series. The sensitizing impurities of petrolatum are polycyclic aromatic hydrocarbons, e.g. phenanthrene derivatives. The purity of petrolatum depends on both the petroleum stock and on the production and packaging methods. Even if rare, contact sensitization to petrolatum can disturb the interpretation of drug patch-tests. It is necessary in the interpretation of drug patch-tests to test both in petrolatum and other vehicles and with all the different petrolatums used in preparing the material for drug patch-tests. So, it is essential to advise the patients sensitized to petrolatum to remove all the topical drugs, such as all the cosmetics, which contain petrolatum in their formulation. PMID- 15291912 TI - Is the first really the first? PMID- 15291913 TI - Basic immunology, allergen prediction, and bioinformatics. PMID- 15291914 TI - A clinical and serological follow-up study of health care workers allergic to natural rubber latex who had followed a latex avoidance programme. PMID- 15291915 TI - A latex-containing hepatitis-B vaccine administered in a severely latex allergic paediatric patient after specific sublingual immunotherapy: a case report. PMID- 15291916 TI - Pilot study assessing the impact of smoking on nasal-specific quality of life. PMID- 15291917 TI - Allergic rhinoconjunctivitis and asthma due to sensitization to Siberian hamster. PMID- 15291918 TI - Cow's milk allergy as a risk factor for forthcoming wheezing attacks. PMID- 15291919 TI - Toll-like receptor-4 genotype in children with respiratory infections. PMID- 15291920 TI - Anaphylaxis to boldo infusion, a herbal remedy. PMID- 15291921 TI - A case of urticaria due to sulpiride. PMID- 15291922 TI - Successful combined therapy for refractory chronic urticaria in a 10-year-old boy. PMID- 15291923 TI - Dendritic cells present neuromuscular blocking agent-related epitopes to T cells from allergic patients. PMID- 15291924 TI - Adverse reactions to vitamin B12 injections due to benzyl alcohol sensitivity: successful treatment with intranasal cyanocobalamin. PMID- 15291927 TI - Is more always better? PMID- 15291929 TI - The fluorescein angiography revolution: a breakthrough with sustained impact. PMID- 15291930 TI - How prostaglandins have changed the medical approach to glaucoma and its costs: an observational study of 2228 patients treated with glaucoma medications. AB - PURPOSE: The aim of the present study was to investigate how the medical treatment of glaucoma changed between 1997 and 2002, since the advent of prostaglandin derivatives, with regard to drug prescriptions and pharmaceutical costs. METHODS: A study was made of medical prescriptions for 2228 patients with glaucoma and/or ocular hypertension, in order to investigate the following: (i) the antiglaucoma drugs most commonly prescribed in 1997 and 2002, and any differences between the drugs prescribed in these 2 years; (ii) the number of drugs used per patient in 1997 and 2002, respectively, and (iii) any increase in the prescribing of antiglaucoma drugs and their relative costs from 1997 to 2002. RESULTS: From 1997 to 2002 there was a sharp drop in the prescribing of beta blockers (79% in 1997 and 55% in 2002). A marked increase in the use of prostaglandin derivatives (0% in 1997 and 18% in 2002) was registered and a marked increase in the prescribing of carbonic anhydrase inhibitors (5% in 1997 and 14% in 2002) was also noted. From 1997 to 2002 there was a trend towards drug addition rather than substitution, so that the number of drugs used per patient increased. The number of patients treated increased enormously (by 98%) from 1997 to 2002. The cost of medical therapy from 1997 to 2002 rose dramatically, with an increase of 148.9% per patient. CONCLUSION: The availability of prostaglandin derivatives has strongly influenced the medical approach to glaucoma. This class of drugs will soon become the type most commonly prescribed for patients with glaucoma and/or ocular hypertension. The increased number of treatments also suggests that the approach of ophthalmologists towards these diseases has changed. Ocular hypertension, as well as glaucoma, is now treated more aggressively. Given the increase in the prescription of prostaglandin derivatives, the pharmaceutical cost of treatment has risen dramatically. PMID- 15291931 TI - Comparison of survival of exfoliative glaucoma patients and primary open-angle glaucoma patients: impact of acetazolamide use. AB - PURPOSE: To compare the survival rates of patients with exfoliative glaucoma (XFG) and those with primary open-angle glaucoma (POAG), and to establish whether the use of acetazolamide has any influence on survival. METHODS: The survival data, including date and cause of death, for 1147 patients with XFG or POAG who were ultimately hospitalized at the Eye Department, National Hospital, Oslo, between 1961 and 1970, were analysed retrospectively. The Cox proportional hazard model was used in the survival analyses. RESULTS: No statistically significant differences in survival were found between patients with XFG and those with POAG (p = 0.85). As expected, female gender and younger age at diagnosis were associated with longer survival periods. Surprisingly, we found that patients with more recent birth dates had relatively lower survival rates than patients with earlier birth dates; when this was included in the analyses, the use of acetazolamide was found to be associated with reduced survival (n = 492, p = 0.02). PMID- 15291932 TI - Exfoliative glaucoma and primary open-angle glaucoma: associations with death causes and comorbidity. AB - PURPOSE: To investigate whether type of glaucoma or use of acetazolamide are associated with main cause of death and comorbidity. MATERIAL AND METHODS: The survival data, including date and cause of death, for 1147 patients with capsular or simple glaucoma who were ultimately hospitalized at the Eye Department, National Hospital, Oslo, between 1961 and 1970, were analysed. Binary logistic regression was carried out to investigate the patterns of death causes and comorbidity in subgroup analyses. RESULTS: Patients with exfoliative glaucoma (XFG) and those with primary open-angle glaucoma (POAG) showed no significant differences in rates of death caused by acute cerebrovascular diseases, cardiac diseases and cancer. Interestingly, we found that chronic cerebral diseases such as senile dementia, cerebral atrophy and chronic cerebral ischaemia (n = 81) were more common in patients with XFG than in those with POAG (p = 0.01) and in the group of acetazolamide users (p = 0.03). Patients with XFG had a higher probability of developing an acute cerebrovascular disease than patients with POAG (n = 228, p = 0.03). CONCLUSION: In this retrospective study, we found that comorbidity with acute cerebrovascular disease and chronic cerebral diseases (senile dementia, cerebral atrophy and chronic cerebral ischaemia) were more common in patients with XFG than in patients with POAG. Prospective data are needed in order to conclude upon the associations found in this study. PMID- 15291933 TI - Self-tonometry with the Ocuton S: evaluation of accuracy in glaucoma patients. AB - BACKGROUND: To compare two methods of measuring intraocular pressure (IOP) and to evaluate whether repeated measurements taken with the Ocuton S applanation self tonometer can improve reliability. METHODS: Ocuton S and Goldmann applanation tonometry (GAT), and corneal thickness measurements taken with the Orbscan topography system, were successfully performed in 64 of 68 glaucoma patients. RESULTS: The median IOPs were 15.5 mmHg using GAT, and 16 mmHg using the first self-taken Ocuton S measurement (n = 64). The differences between the median of the GAT measurements and the first Ocuton S measurement, and the medians of the three and six separate Ocuton S measurements were within 3 mmHg in 52%, 59% and 67% of cases, respectively. The mean corneal thickness of all evaluated eyes was 545.3 microm. There was no effect of corneal thickness on the accuracy of either of the two devices (p > 0.05). CONCLUSION: Repeated measurements can improve the reliability of the Ocuton S. However, even with repeated measurements only every second patient succeeds in obtaining reliable measurements. PMID- 15291934 TI - The nerve fibre layer symmetry test: computerized evaluation of human retinal nerve fibre layer thickness as measured by optical coherence tomography. AB - PURPOSE: To present and test a new interpretative concept, the nerve fibre layer symmetry test (NST), for computerized evaluation of retinal nerve fibre layer thickness (RNFLT) as measured by optical coherence tomography (OCT) in glaucoma. METHODS: The NST concept was constructed and tested in a pilot study. A total of 32 healthy and 40 age-matched glaucomatous eyes were included and examined by OCT, computerized perimetry, RNFL/disc photography, tonometry and a general ophthalmologic examination. RESULTS: The observed NST sensitivity and specificity were high, at 38/40 eyes (95%) and 32/32 eyes (100%), respectively, and 40/40 eyes (100%), and 31/32 eyes (97%), respectively, when correcting the OCT RNFLT measurement for the influence of variability in image signal/quality. The NST sensitivity was 8-10% higher than the single most sensitive traditional OCT RNFLT parameter; this difference was not statistically significant in this small sample. CONCLUSION: The NST showed high specificity and sensitivity for detection of RNFLT attenuation indicating early to severe glaucoma. Although promising, the NST needs to be further developed and validated in larger study samples and in patients with various stages of glaucomatous damage. PMID- 15291935 TI - The influence of age, sex, race, refractive error and optic disc parameters on the sensitivity and specificity of scanning laser polarimetry. AB - PURPOSE: To evaluate the influence of age, sex, race, refractive error and optic disc topography on the sensitivity and specificity of scanning laser polarimetry (SLP) in the diagnosis of glaucoma. METHODS: A total of 88 normal individuals and 95 glaucoma patients were included in this study. Glaucoma was defined on the basis of both optic nerve damage and visual field defects. Scanning laser polarimetry, optic disc topography, automated perimetry and refractometry were performed in all subjects. The sensitivity and specificity of SLP were assessed applying a previously calculated cut-off to a previously described linear discriminant function (LDF). RESULTS: The sensitivity and specificity of SLP in the study population were 82% and 83%, respectively. Sensitivity and specificity were not affected by age, sex, race, average disc diameter or disc area. The sensitivity of SLP tended to be higher in myopes (93%) than in emmetropes (80%) and hyperopes (71%) (p = 0.08). Sensitivities were higher in individuals with cup areas > 0.96 mm(2) (89%), rim areas < or = 1.36 mm(2) (92%), and cup area/disc area ratios > 0.45 (89%) (p < 0.05). Stepwise logistic regression analysis indicated that the presence of a cup area > 0.96 mm(2) and a rim area < 1.36 mm(2) significantly increased the sensitivity of the LDF, whereas a cup area/disc area ratio < or = 0.45 significantly increased the specificity of the LDF. CONCLUSION: The sensitivity and specificity of SLP may be influenced by refractive error and optic disc parameters that are affected by glaucomatous damage (cup area, rim area and cup area/disc area ratio). These parameters must be considered in studies evaluating the sensitivity and specificity of optic nerve/retinal nerve fibre layer imaging technologies. PMID- 15291936 TI - Comparison of colour Doppler imaging and retinal scanning laser fluorescein angiography in healthy volunteers and normal pressure glaucoma patients. AB - PURPOSE: To correlate retinal circulatory measurements using scanning laser fluorescein angiography and flow velocities of retrobulbar vessels measured by means of colour Doppler imaging. METHODS: Fifteen patients with normal pressure glaucoma (NPG) and 15 healthy volunteers underwent colour Doppler imaging and fluorescein angiographic studies. Peak systolic velocities (PSVs), end-diastolic velocities (EDVs) and resistive indices (RIs) of the ophthalmic artery (OA) and central retinal artery were obtained. In the fluorescein angiograms arteriovenous passage time (AVP) was quantified by means of digital dye dilution curve analysis. RESULTS: Arteriovenous passage time was significantly prolonged in NPG patients compared to healthy subjects (p = 0.0026). In the central retinal artery PSV (p = 0.023) and EDV (p < 0.0001) were significantly decreased and RI was increased (p < 0.0001) in patients with NPG. The EDV of the central retinal artery showed a significant correlation with AVP (EDV: r = - 0.53, p = 0.0023). The RI of the central retinal artery correlated significantly to AVP (RI: r = 0.63, p < 0.0001). The AVP did not correlate to EDV or PSV, nor to the RI measured in the ophthalmic artery. CONCLUSION: Arteriovenous passage time, which represents blood flow in a vascular segment of artery, capillary bed and corresponding vein, was found to be correlated to the EDV and the RI of the central retinal artery. The combination of different techniques allows further interpretation of ocular circulatory responses. PMID- 15291937 TI - Effects of moderate smoking on the central visual field. AB - PURPOSE: To investigate whether moderate cigarette smoking has any effects on the central visual field. METHODS: This study included 30 healthy, moderate cigarette smokers (10-20 cigarettes per day for at least the past 5 years) and 22 healthy non-smokers. After two training test sessions, all individuals underwent computerized visual field examinations (Humphrey 30-2 Full Threshold Test) with both white-on-white (W-W) perimetry and blue-on-yellow (B-Y) perimetry. One eye of each subject with reliable visual field test results was evaluated. The foveal threshold, mean deviation (MD), pattern standard deviation (PSD), short-term fluctuation (SF), corrected pattern standard deviation (CPSD), glaucoma hemifield test (GHT) and number of significantly depressed points deviating at p < 5%, p < 2%, p < 1% and p < 0.5% on the pattern deviation probability map of the smokers were compared with those of the non-smokers. RESULTS: When the results of W-W perimetry were analysed, the smokers were found to have significantly lower foveal thresholds (p = 0.001) and mean retinal sensitivity (p = 0.02), and higher PSD (p = 0.002) and CPSD (p = 0.01) than the non-smokers. Short-term fluctuation was similar in both groups (p = 0.55). The number of significantly depressed points deviating at p < 5%, p < 2% and p < 1% on the pattern deviation probability map was similar for both groups (p > 0.05). The number of depressed points deviating at p < 0.5% on the pattern deviation probability map was higher for the smokers than for the non-smokers (p = 0.03). The results of B-Y perimetry showed the smokers to have a significantly lower foveal threshold than the non smokers (p = 0.03). However, there were no significant differences in the global indices of the two groups (p > 0.05). The number of significantly depressed points deviating at p < 5%, p < 2%, p < 1% and p < 0.5% on the pattern deviation probability map was similar in both groups (p > 0.05). No significant difference in GHT was determined with either perimetry for the smokers compared with the non smokers (p > 0.05). CONCLUSION: This study suggests that moderate cigarette smoking is associated with both diffuse and localized reductions in retinal sensitivity with W-W perimetry. Only reduction in the foveal threshold was observed with B-Y perimetry, with no hints of diffuse and localized reductions. PMID- 15291938 TI - Colour blindness in everyday life and car driving. AB - PURPOSE: The aim of the present work was to ascertain, through the administration of a psychosocial questionnaire, the difficulties that subjects with defective colour vision experience in carrying out everyday tasks and work, including driving a car with a driver's licence held for no more than 3 years. METHODS: Subjects with defective colour vision (n = 151) and subjects with normal vision (n = 302) completed a psychosocial questionnaire regarding the difficulties associated with congenital colour vision deficiency in daily life, work and driving a car. Subjects were diagnosed as colour-blind using the Ishihara test. RESULTS: Statistically significant differences between the two samples were found for daily life activities. Subjects with defective colour vision preferred daytime driving. At night, subjects with defective colour vision had difficulty identifying reflectors on the road and the rear signal lights of cars ahead of them. CONCLUSION: Colour-blind Calabrian subjects admitted to experiencing colour related difficulties with a wide range of occupational tasks and leisure pursuits. In particular, colour-blind Calabrian subjects preferred daytime driving, and fewer drove regularly, compared to orthochromatics, who were indifferent to night or daytime driving. PMID- 15291939 TI - The prevalence of retinopathy in an unselected population of type 2 diabetes patients from Arhus County, Denmark. AB - PURPOSE: To determine the prevalence of diabetic retinopathy and the causes of visual impairment in an unselected population of type 2 diabetes patients, and to describe the risk factors for developing diabetic retinopathy in this population. METHODS: A total of 10 851 type 2 diabetes patients were identified in the county of Arhus. A representative sample of 378 patients underwent a routine ocular examination, including fundus photography. Blood pressure and serum haemoglobin A1c, total cholesterol, high density lipoprotein cholesterol, triglyceride and apolipoprotein a were measured. RESULTS: The prevalence of diabetic retinopathy in the type 2 diabetes population was 31.5%. In all, 2.9% had proliferative diabetic retinopathy and 5.3% had clinically significant macular oedema. Of the latter, 8/20 (40%) were newly identified and had not yet been laser-treated. There was a positive correlation between severity of retinopathy and duration of diabetes, HbA(1c), systolic blood pressure and treatment with insulin. None of the patients had social blindness (visual acuity < 0.1), but 15/378 (4.0%) had developed visual impairment (VA < 0.3). CONCLUSION: The prevalence of diabetic retinopathy and visual impairment in this unselected type 2 diabetes population was lower than anticipated from the existing literature, and causes other than diabetic retinopathy contributed significantly to the occurrence of visual loss. A substantial number of the patients with vision-threatening diabetic maculopathy had not been referred for timely photocoagulation treatment. PMID- 15291940 TI - Ocular refractive changes in patients receiving hyperbaric oxygen administered by oronasal mask or hood. AB - PURPOSE: The aim of this study was to quantify ocular refractive changes after a standard hyperbaric oxygen (HBO) treatment protocol and to characterize the time period of recovery. PATIENTS AND METHODS: Hyperbaric oxygen therapy was given for 90 min daily at a pressure of 240 kPa for 21 days. Oxygen was administered to 20 patients using an oronasal mask and to 12 patients using a hood. Follow-up examinations were carried out 2-4 days after treatment, and thereafter regularly for up to 10 weeks in both groups. Refraction was assessed automatically and by the monocular subjective refraction method. A subgroup of nine of the 20 patients to whom oxygen was administered by an oronasal mask underwent a separate eye examination, which included crystalline lens opacity measurements and LOCS III gradings. RESULTS: In the patients given oxygen by mask, there was a significant myopic shift in the mean spherical equivalent, which was largest 2-4 days after treatment. The shift was - 0.55 +/- 0.40 D in the right eye and - 0.53 +/- 0.42 D in the left eye. In the patients given oxygen by hood, the largest shift was observed after 12-16 days, and was - 1.06 +/- 0.52 D in the right eye and - 1.10 +/- 0.57 D in the left eye. The refractive changes returned to baseline 6 weeks and 10 weeks after HBO treatment, respectively. No significant changes in crystalline lens transparency were revealed. CONCLUSIONS: The myopic shift after HBO therapy recovers within 10 weeks and may be more pronounced when patients are given oxygen using a hood compared with using an oronasal mask. PMID- 15291941 TI - Pupil size and night vision disturbances after LASIK for myopia. AB - PURPOSE: To examine whether standardized, preoperative evaluation of pupil sizes can predict the risk of night vision visual disturbances after bilateral laser in situ keratomileusis (LASIK) for myopia. METHODS: A prospective study was carried out involving 46 patients who underwent bilateral LASIK for myopia. Pupil sizes were measured before surgery using an infrared pupillometer under standardized settings. Pre- and postoperative refraction and best spectacle-corrected visual acuity (BSCVA) were registered. At the 3-month follow-up visit, the patients completed a questionnaire regarding night vision pre- and postoperatively. RESULTS: The mean bilateral, spherical equivalent refraction (SE) was - 8.76 D (range 6.32 to - 12.0 D) preoperatively, and - 1.69 D (range 0 to - 4.38 D) postoperatively. The mean bilateral BSCVA was not changed by the operations. We found a significant correlation between large scotopic pupil sizes and the impression of worsened night vision (p < 0.01). A significant correlation between gender (males) and subjectively reduced night vision postoperatively was also found (p < 0.05). CONCLUSION: Large pupil size measured preoperatively is correlated with an increased frequency of subjectively experienced post-LASIK visual disturbances during scotopic conditions. We recommend preoperative evaluation of pupil size in all patients prior to LASIK surgery. PMID- 15291942 TI - Surgical treatment of complete anterior capsule contraction after cataract surgery. PMID- 15291943 TI - 16S rDNA PCR analysis of infectious keratitis: a case series. AB - PURPOSE: To discuss the value of polymerase chain reaction (PCR) in the management of bacterial infectious keratitis. METHODS: Corneal scrapings of four patients with severe infectious keratitis were analysed by culture and PCR of 16S ribosomal DNA (rDNA), followed by direct sequencing of the resulting amplicon. The medical history of the patients included laser-assisted in-situ keratomileusis (LASIK), penetrating keratoplasty (PKP) and trauma. RESULTS: Using PCR we were able to identify a possible pathogen in all four cases, while bacterial cultures were either negative or did not correspond to the clinical picture. The identified bacteria were a Pseudomonas species, Abiotrophia defectiva, Stenotrophomonas maltophilia and Porphyromonas gingivalis. CONCLUSIONS: Analysis of corneal scrapings by 16S rDNA PCR should be considered as a supplement to standard microbiological procedures. However, the results of this relatively new method have to be interpreted carefully. PMID- 15291944 TI - Successful primary culture and autologous transplantation of corneal limbal epithelial cells from minimal biopsy for unilateral severe ocular surface disease. AB - BACKGROUND: Patients with severe unilateral ocular surface disease require reconstruction of the damaged ocular surface. We succeeded in culturing primary corneal limbal epithelial cells taken from minimal biopsy and, once grown, transplanting them on denuded amniotic membrane (AM). METHODS: Autologous corneal limbal epithelial cells from a 3 mm(2) biopsy of the uninjured eye were grown for 3 weeks on a denuded AM carrier. The resultant sheet was then transplanted onto the unilateral severely chemically injured eye. RESULTS: Minimal biopsy showed the autologous cultivated corneal epithelial cells to have 4-5 layers of sufficient stratification and to be well differentiated. At 19 months post transplantation, the ocular surface epithelium was stable and there were no epithelial defects. CONCLUSION: We document that it is possible to produce sufficiently stratified, well differentiated, autologous cultivated corneal limbal epithelium on AM from a minimal biopsy of the donor eye and to transplant it onto the injured eye. PMID- 15291945 TI - Relapsing polychondritis: a rare disease with varying symptoms. AB - PURPOSE: Relapsing polychondritis (RPC) is a rare systemic disease affecting primarily cartilaginous and proteoglycan-rich structures. It is a potentially fatal disease with unknown aetiology. There are no specific tests for RPC. The diagnosis is dependent on clinical criteria, which include chondritis of both auricles, non-erosive inflammatory polyarthritis, nasal chondritis, ocular inflammation, respiratory tract chondritis and cochlear and/or vestibular damage. Ocular symptoms will occur in approximately 60% of RPC patients. As an example, a patient with signs of RPC is described. METHODS/RESULT: A 30-year-old woman was referred to our department for evaluation of a central corneal ulcer in the left eye. She had a history of recurrent pain in both her auricles and was also found to have a nasal septum perforation. Relapsing polychondritis was suspected. CONCLUSION: Non-healing corneal ulcers should alert the ophthalmologist to look for unusual reasons for this condition. RPC is one possible cause. PMID- 15291946 TI - Multifocal electroretinography and optical coherence tomography in two patients with solar retinopathy. AB - PURPOSE: Clinical investigation of central retinal dysfunction in two cases of solar retinopathy. METHODS: Two patients were examined for best corrected visual acuity (VA), fundus inspection, visual fields, multifocal electroretinography (mfERG) with a stimulus pattern of 241 hexagons and, at follow-up, also with optical coherence tomography (OCT). RESULTS: At the initial examination, mfERG revealed central retinal dysfunction, which had improved by the time of follow up. In Case 1, a foveal oedema regressed over time, although VA remained slightly reduced. In Case 2, OCT showed spots of increased reflectivity corresponding to the patient's symptoms. CONCLUSION: Central retinal dysfunction due to solar retinopathy may improve over time. However, structural and functional changes may persist. This report illustrates that mfERG and OCT are useful tools for objective documentation of the pathology in solar retinopathy. PMID- 15291949 TI - Promising visual improvement of cystoid macular oedema by hyperbaric oxygen therapy. PMID- 15291950 TI - Longterm refractive and visual outcomes in subjects with high hyperopia. PMID- 15291951 TI - Bilateral epiretinal membranes in nevoid basal cell carcinoma syndrome. PMID- 15291952 TI - Bilateral microphthalmia with cyst. PMID- 15291953 TI - Intraorbital wooden foreign body. PMID- 15291954 TI - Recurrent spontaneous hyphaema secondary to a melanocytic iris lesion. PMID- 15291957 TI - The significance of controlled conditions in lentiviral vector titration and in the use of multiplicity of infection (MOI) for predicting gene transfer events. AB - BACKGROUND: Although lentiviral vectors have been widely used for in vitro and in vivo gene therapy researches, there have been few studies systematically examining various conditions that may affect the determination of the number of viable vector particles in a vector preparation and the use of Multiplicity of Infection (MOI) as a parameter for the prediction of gene transfer events. METHODS: Lentiviral vectors encoding a marker gene were packaged and supernatants concentrated. The number of viable vector particles was determined by in vitro transduction and fluorescent microscopy and FACs analyses. Various factors that may affect the transduction process, such as vector inoculum volume, target cell number and type, vector decay, variable vector - target cell contact and adsorption periods were studied. MOI between 0-32 was assessed on commonly used cell lines as well as a new cell line. RESULTS: We demonstrated that the resulting values of lentiviral vector titre varied with changes of conditions in the transduction process, including inoculum volume of the vector, the type and number of target cells, vector stability and the length of period of the vector adsorption to target cells. Vector inoculum and the number of target cells determine the frequencies of gene transfer event, although not proportionally. Vector exposure time to target cells also influenced transduction results. Varying these parameters resulted in a greater than 50-fold differences in the vector titre from the same vector stock. Commonly used cell lines in vector titration were less sensitive to lentiviral vector-mediated gene transfer than a new cell line, FRL 19. Within 0-32 of MOI used transducing four different cell lines, the higher the MOI applied, the higher the efficiency of gene transfer obtained. CONCLUSION: Several variables in the transduction process affected in in vitro vector titration and resulted in vastly different values from the same vector stock, thus complicating the use of MOI for predicting gene transfer events. Commonly used target cell lines underestimated vector titre. However, within a certain range of MOI, it is possible that, if strictly controlled conditions are observed in the vector titration process, including the use of a sensitive cell line, such as FRL 19 for vector titration, lentivector-mediated gene transfer events could be predicted. PMID- 15291958 TI - The AMC Linear Disability Score project in a population requiring residential care: psychometric properties. AB - BACKGROUND: Currently there is a lot of interest in the flexible framework offered by item banks for measuring patient relevant outcomes, including functional status. However, there are few item banks, which have been developed to quantify functional status, as expressed by the ability to perform activities of daily life. METHOD: This paper examines the psychometric properties of the AMC Linear Disability Score (ALDS) project item bank using an item response theory model and full information factor analysis. Data were collected from 555 respondents on a total of 160 items. RESULTS: Following the analysis, 79 items remained in the item bank. The remaining 81 items were excluded because of: difficulties in presentation (1 item); low levels of variation in response pattern (28 items); significant differences in measurement characteristics for males and females or for respondents under or over 85 years old (26 items); or lack of model fit to the data at item level (26 items). CONCLUSIONS: It is conceivable that the item bank will have different measurement characteristics for other patient or demographic populations. However, these results indicate that the ALDS item bank has sound psychometric properties for respondents in residential care settings and could form a stable base for measuring functional status in a range of situations, including the implementation of computerised adaptive testing of functional status. PMID- 15291959 TI - The ECOS-16 questionnaire for the evaluation of health related quality of life in post-menopausal women with osteoporosis. AB - BACKGROUND: The aim of this study is to validate the questionnaire ECOS-16 (Assessment of health related quality of life in osteoporosis) for the evaluation of health related quality of life (HRQoL) in post-menopausal women with osteoporosis. METHODS: An observational, prospective and multi-centre study was carried out among post-menopausal women with osteoporosis in primary care centres and hospital outpatient clinics. All patients attended 2 visits: at baseline and at 6 months. In addition, the subgroup of outpatients attended another visit a month after the baseline to assess the test-retest reliability. The psychometric properties of the questionnaire were evaluated in terms of feasibility, validity (content validity and construct validity) and internal consistency in baseline, and in terms of test-retest reliability and responsiveness to change in visit at month and visit at 6 months, respectively. In all visits, ECOS-16, EUROQoL-5D (EQ 5D) and four 7-point items about health status (general health status, back pain, limitation in daily activities and emotional status) were administered, whereas only outpatients were given MINI-OQLQ (Mini Osteoporosis Quality of Life Questionnaire), besides all clinical variables; and sociodemographic variables at baseline. RESULTS: 316 women were consecutively included, 212 from primary care centres and 104 from hospital outpatient clinics. Feasibility: 94.3% of patients answered all items of the questionnaire. The mean administration time was 12.3 minutes. VALIDITY: factor analysis suggested that the questionnaire was unidimensional. In the multivariate analysis, patients with vertebral fractures, co-morbidity and a lower education level showed to have worse HRQoL. Moderate to high correlations were found between the ECOS-16 score and the other health status questionnaires (0.47-0.82). Reliability: internal consistency (Cronbach's alpha) was 0.92 and test-retest reliability (ICC) was 0.80. Responsiveness to change: ECOS-16 scores increased according to change perceived by the patient, as well as the effect size (ranges between 1.35 to 0.43), the greater the perception of change in patients' general health status, the greater the changes in patients' scores. The Minimal Clinically Important Difference (MCID) suggested a change of 0.5 points in the ECOS-16 score, representing the least improvement in general health status due to their osteoporosis: "slightly better". CONCLUSION: ECOS-16 has been proven preliminarily to have good psychometric properties, so that it can be potentially a useful tool to evaluate HRQoL of post-menopausal women with osteoporosis in research and routine clinical practice. PMID- 15291960 TI - The geographic distribution of breast cancer incidence in Massachusetts 1988 to 1997, adjusted for covariates. AB - BACKGROUND: The aims of this study were to determine whether observed geographic variations in breast cancer incidence are random or statistically significant, whether statistically significant excesses are temporary or time-persistent, and whether they can be explained by covariates such as socioeconomic status (SES) or urban/rural status? RESULTS: A purely spatial analysis found fourteen geographic areas that deviated significantly from randomness: ten with higher incidence rates than expected, four lower than expected. After covariate adjustment, three of the ten high areas remained statistically significant and one new high area emerged. The space-time analysis identified eleven geographic areas as statistically significant, seven high and four low. After covariate adjustment, four of the seven high areas remained statistically significant and a fifth high area also identified in the purely spatial analysis emerged. CONCLUSIONS: These analyses identify geographic areas with invasive breast cancer incidence higher or lower than expected, the times of their excess, and whether or not their status is affected when the model is adjusted for risk factors. These surveillance findings can be a sound starting point for the epidemiologist and has the potential of monitoring time trends for cancer control activities. PMID- 15291961 TI - Does preoperative abduction value affect functional outcome of combined muscle transfer and release procedures in obstetrical palsy patients with shoulder involvement? AB - BACKGROUND: Obstetric palsy is the injury of the brachial plexus during delivery. Although many infants with plexopathy recover with minor or no residual functional deficits, some children don't regain sufficient limb function because of functional limitations, bony deformities and joint contractures. Shoulder is the most frequently affected joint with internal rotation contracture causing limitation of abduction, external rotation. The treatment comprises muscle release procedures such as posterior subscapularis sliding or anterior subscapularis tendon lengthening and muscle transfers to restore the missing external rotation and abduction function. METHODS: We evaluated whether the preoperative abduction degree affects functional outcome. Between 1998 and 2002, 46 children were operated on to restore shoulder abduction and external rotation. The average age at surgery was 7.6 years and average follow up was 40.8 months. We compared the postoperative results of the patients who had preoperative abduction less than 90 degrees (Group I: n = 37) with the patients who had preoperative abduction greater than 90 degrees (Group II: n = 9), in terms of abduction and external rotation function with angle measurements and Mallet classification. We inquired whether patients in Group I needed another muscle transfer along with latissimus dorsi and teres major transfers. RESULTS: In Group I the average abduction improved from 62.5 degrees to 131.4 degrees (a 68.9 degrees +/- 22.9 degrees gain) and the average external rotation improved from 21.4 degrees to 82.6 degrees (a 61.1 degrees +/- 23 degrees gain). In Group II the average abduction improved from 99.4 degrees to 140 degrees (a 40.5 degrees +/- 16 degrees gain) and the average external rotation improved from 33.2 degrees to 82.7 degrees (a 49.5 degrees +/- 23.9 degrees gain). Although there was a significant difference between Group I and II for preoperative abduction (p = 0.000) and abduction gain in degrees (p = 0.001), the difference between postoperative values of both groups was not significant (p = 0.268). There was also no significant difference between the two groups in the preoperative external rotation, the external rotation gain and the postoperative external rotation (p = 0.163, p = 0.181 and p = 0.803, respectively). CONCLUSIONS: Obstetric palsy patients with shoulder sequela who had a preoperative abduction less than 90 degrees hadas good functional results using latissimus dorsi, teres major muscle transfer and subscapularis muscle release as the patients who hada preoperative abduction greater than 90 degrees. PMID- 15291963 TI - Corneal topographic changes following retinal surgery. AB - BACKGROUND: To study the effect of retinal/ vitreoretinal surgeries on corneal elevations. METHODS: Patients who underwent retinal/ vitreoretinal surgeries were divided into 3 groups. Scleral buckling was performed in 11 eyes (Group 1). In 8 (25%) eyes, vitreoretinal surgery was performed along with scleral buckling (Group 2). In 12 eyes, pars plana vitrectomy was performed for vitreous hemorrhage (Group 3). An encircling element was used in all the eyes. The parameters evaluated were best-corrected visual acuity (BCVA), change in axial length, and corneal topographic changes on Orbscan topography system II, preoperative and at 12 weeks following surgery. RESULTS: There was a statistically significant increase in anterior corneal elevation in all the three groups after surgery (p = 0.003, p = 0.008 & p = 0.003 respectively). The increase in posterior corneal elevation was highly significant in all the three groups after surgery (p = 0.0000, p = 0.0001 & p = 0.0001 respectively). The increase in the posterior corneal elevation was more than the increase in the anterior elevation and was significant statistically in all the three groups (group I: p = 0.02; group II: p = 0.01; group III: p = 0.008). CONCLUSIONS: Retinal/ vitreoretinal surgeries cause a significant increase in the corneal elevations and have a greater effect on the posterior corneal surface. PMID- 15291962 TI - Altered mRNA expression of genes related to nerve cell activity in the fracture callus of older rats: A randomized, controlled, microarray study. AB - BACKGROUND: The time required for radiographic union following femoral fracture increases with age in both humans and rats for unknown reasons. Since abnormalities in fracture innervation will slow skeletal healing, we explored whether abnormal mRNA expression of genes related to nerve cell activity in the older rats was associated with the slowing of skeletal repair. METHODS: Simple, transverse, mid-shaft, femoral fractures with intramedullary rod fixation were induced in anaesthetized female Sprague-Dawley rats at 6, 26, and 52 weeks of age. At 0, 0.4, 1, 2, 4, and 6 weeks after fracture, a bony segment, one-third the length of the femur, centered on the fracture site, including the external callus, cortical bone, and marrow elements, was harvested. cRNA was prepared and hybridized to 54 Affymetrix U34A microarrays (3/age/time point). RESULTS: The mRNA levels of 62 genes related to neural function were affected by fracture. Of the total, 38 genes were altered by fracture to a similar extent at the three ages. In contrast, eight neural genes showed prolonged down-regulation in the older rats compared to the more rapid return to pre-fracture levels in younger rats. Seven genes were up-regulated by fracture more in the younger rats than in the older rats, while nine genes were up-regulated more in the older rats than in the younger. CONCLUSIONS: mRNA of 24 nerve-related genes responded differently to fracture in older rats compared to young rats. This differential expression may reflect altered cell function at the fracture site that may be causally related to the slowing of fracture healing with age or may be an effect of the delayed healing. PMID- 15291964 TI - Elevated expression of MMP-13 and TIMP-1 in head and neck squamous cell carcinomas may reflect increased tumor invasiveness. AB - BACKGROUND: Matrix metalloproteinases [MMPs], which degrade the extracellular matrix, play an important role in the invasion and metastasis of squamous cell carcinomas. One MMP, MMP-13, is thought to play a central role in MMP activation. The purpose of this study was to investigate MMP-13 and TIMP-1 expression in squamous cell carcinomas of the head and neck and to relate these levels of expression to histologic patterns of invasion. METHODS: This study included T1 lesions obtained via biopsy from the larynx, tongue, and skin/mucosa of 78 patients with head and neck squamous cell carcinomas. The relationship between expression of MMP-13 and TIMP-1 and the mode of tumor invasion [MI] was evaluated immunohistochemically, using breast carcinoma tissue as a positive control. RESULTS: Increased expression was observed in highly invasive tumors, as reflected by the significant correlation between the degree of staining for MMP 13 or TIMP-1 and MI grade [p < 0.05]. There was no significant relationship between the degree of staining for MMP-13 or TIMP-1 and patient age, sex, tumor site, or tumor histologic grade. In addition, levels of staining for MMP-13 did not correlate with levels of staining for TIMP-1. CONCLUSION: The expression of MMP-13 and TIMP-1 appears to play an important role in determining the invasive capacity of squamous cell carcinomas of the head and neck. Whereas additional studies are needed to confirm these findings, evaluating expression of these MMPs in small biopsy samples may be useful in determining the invasive capacity of these tumors at an earlier stage. PMID- 15291965 TI - 'Perinatal outcome in preterm premature rupture of membranes with Amniotic fluid index < 5 (AFI < 5). AB - BACKGROUND: Our purpose was to determine whether AFI<5 cm after preterm premature rupture of the membranes (PPROM) is associated with an increased risk of perinatal morbidity. METHODS: We performed a prospective cohort study of 95 singleton pregnancies complicated by preterm premature rupture of the membranes (PPROM) with delivery between 26 and 34 weeks gestation.Patients were categorized in two groups on the basis of amniotic fluid index<5, (AFI<5 cm)(n = 26) or AFI >/= 5 cm (n = 69). Categorical data were tested for significance with the chi2 and Fisher exact tests. Continuous data were evaluated for normal distribution and tested for significance with the student t test.All 2-sided p values < 0.05 were considered significant. RESULTS: Both groups were similar with respect to selected demographics, gestational age at rupture of the membranes, gestational age at the delivery, birth weight. Both groups were similar with respect to selected variable, latency until delivery, early onset neonatal sepsis, RDS and neonatal death. Patients with AFI<5 cm demonstrated greater frequency of C/S delivery for non reassuring fetal tests (23%vs 2.8%) (p = 0.001). Our study demonstrated that patients in group I had a significant increase in the frequency of clinical chorioamnionitis (P < 0/001). Post partum infections were not seen in 2 groups. CONCLUSIONS: An AFI<5 cm after PPROM between 26 and 34 weeks gestation is associated with an increased risk of maternal infections and frequency of C/S. PMID- 15291966 TI - Candidate high myopia loci on chromosomes 18p and 12q do not play a major role in susceptibility to common myopia. AB - BACKGROUND: To determine whether previously reported loci predisposing to nonsyndromic high myopia show linkage to common myopia in pedigrees from two ethnic groups: Ashkenazi Jewish and Amish. We hypothesized that these high myopia loci might exhibit allelic heterogeneity and be responsible for moderate /mild or common myopia. METHODS: Cycloplegic and manifest refraction were performed on 38 Jewish and 40 Amish families. Individuals with at least -1.00 D in each meridian of both eyes were classified as myopic. Genomic DNA was genotyped with 12 markers on chromosomes 12q21-23 and 18p11.3. Parametric and nonparametric linkage analyses were conducted to determine whether susceptibility alleles at these loci are important in families with less severe, clinical forms of myopia. RESULTS: There was no strong evidence of linkage of common myopia to these candidate regions: all two-point and multipoint heterogeneity LOD scores were < 1.0 and non parametric linkage p-values were > 0.01. However, one Amish family showed slight evidence of linkage (LOD>1.0) on 12q; another 3 Amish families each gave LOD >1.0 on 18p; and 3 Jewish families each gave LOD >1.0 on 12q. CONCLUSIONS: Significant evidence of linkage (LOD> 3) of myopia was not found on chromosome 18p or 12q loci in these families. These results suggest that these loci do not play a major role in the causation of common myopia in our families studied. PMID- 15291967 TI - Arrhythmia-provoking factors and symptoms at the onset of paroxysmal atrial fibrillation: a study based on interviews with 100 patients seeking hospital assistance. AB - BACKGROUND: Surprisingly little information on symptoms of paroxysmal atrial fibrillation is available in scientific literature. Using questionnaires, we have analyzed the symptoms associated with arrhythmia attacks. METHODS: One hundred randomly-selected patients with idiopathic paroxysmal atrial fibrillation filled in a structured questionnaire. RESULTS: Psychic stress was the most common factor triggering arrhythmia (54%), followed by physical exertion (42%), tiredness (41%) coffee (25%) and infections (22%). Thirty-four patients cited alcohol, 26 in the form of red wine, 16 as white wine and 26 as spirits. Among these 34, red wine and spirits produced significantly more episodes of arrhythmia than white wine (p = 0.01 and 0.005 respectively). Symptoms during arrhythmia were palpitations while exerting (88%), reduced physical ability (87%), palpitations at rest (86%), shortage of breath during exertion (70%) and anxiety (59%). Significant differences between sexes were noted regarding swollen legs (women 21%, men 6%, p = 0.027), nausea (women 36%, men 13%, p = 0.012) and anxiety (females 79%, males 51%, p = 0.014). CONCLUSION: Psychic stress was the commonest triggering factor in hospitalized patients with paroxysmal atrial fibrillation. Red wine and spirits were more proarrhythmic than white wine. Symptoms in women in connection with attacks of arrhythmia vary somewhat from those in men. PMID- 15291968 TI - Short-term cytotoxic effects and long-term instability of RNAi delivered using lentiviral vectors. AB - BACKGROUND: RNA interference (RNAi) can potently reduce target gene expression in mammalian cells and is in wide use for loss-of-function studies. Several recent reports have demonstrated that short double-stranded RNAs (dsRNAs), used to mediate RNAi, can also induce an interferon-based response resulting in changes in the expression of many interferon-responsive genes. Off-target gene silencing has also been described, bringing into question the validity of certain RNAi based approaches for studying gene function. We have targeted the plasminogen activator inhibitor-2 (PAI-2 or SERPINB2) mRNA using lentiviral vectors for delivery of U6 promoter-driven PAI-2-targeted short hairpin RNA (shRNA) expression. PAI-2 is reported to have anti-apoptotic activity, thus reduction of endogenous expression may be expected to make cells more sensitive to programmed cell death. RESULTS: As expected, we encountered a cytotoxic phenotype when targeting the PAI-2 mRNA with vector-derived shRNA. However, this predicted phenotype was a potent non-specific effect of shRNA expression, as functional overexpression of the target protein failed to rescue the phenotype. By decreasing the shRNA length or modifying its sequence we maintained PAI-2 silencing and reduced, but did not eliminate, cytotoxicity. ShRNA of 21 complementary nucleotides (21 mers) or more increased expression of the oligoadenylate synthase-1 (OAS1) interferon-responsive gene. 19 mer shRNA had no effect on OAS1 expression but long-term selective pressure on cell growth was observed. By lowering lentiviral vector titre we were able to reduce both expression of shRNA and induction of OAS1, without a major impact on the efficacy of gene silencing. CONCLUSIONS: Our data demonstrate a rapid cytotoxic effect of shRNAs expressed in human tumor cell lines. There appears to be a cut-off of 21 complementary nucleotides below which there is no interferon response while target gene silencing is maintained. Cytotoxicity or OAS1 induction could be reduced by changing shRNA sequence or vector titre, but stable gene silencing could not be maintained in extended cell culture despite persistent marker gene expression from the RNAi-inducing transgene cassette. These results underscore the necessity of careful controls for immediate and long-term RNAi use in mammalian cell systems. PMID- 15291969 TI - P2 receptor mRNA expression profiles in human lymphocytes, monocytes and CD34+ stem and progenitor cells. AB - BACKGROUND: Extracellular nucleotides (ATP, ADP, UTP and UDP) exert a wide range of biological effects in blood cells mediated by multiple ionotropic P2X receptors and G protein-coupled P2Y receptors. Although pharmacological experiments have suggested the presence of several P2 receptor subtypes on monocytes and lymphocytes, some results are contradictory. Few physiological functions have been firmly established to a specific receptor subtype, partly because of a lack of truly selective agonists and antagonists. This stimulated us to investigate the expression of P2X and P2Y receptors in human lymphocytes and monocytes with a newly established quantitative mRNA assay for P2 receptors. In addition, we describe for the first time the expression of P2 receptors in CD34+ stem and progenitor cells implicating a potential role of P2 receptors in hematopoietic lineage and progenitor/stem cell function. RESULTS: Using a quantitative mRNA assay, we assessed the hypothesis that there are specific P2 receptor profiles in inflammatory cells. The P2X4 receptor had the highest expression in lymphocytes and monocytes. Among the P2Y receptors, P2Y12 and P2Y2 had highest expression in lymphocytes, while the P2Y2 and P2Y13 had highest expression in monocytes. Several P2 receptors were expressed (P2Y2, P2Y1, P2Y12, P2Y13, P2Y11, P2X1, P2X4) in CD34+ stem and progenitor cells. CONCLUSIONS: The most interesting findings were the high mRNA expression of P2Y12 receptors in lymphocytes potentially explaining the anti-inflammatory effects of clopidogrel, P2Y13 receptors in monocytes and a previously unrecognised expression of P2X4 in lymphocytes and monocytes. In addition, for the first time P2 receptor mRNA expression patterns was studied in CD34+ stem and progenitor cells. Several P2 receptors were expressed (P2Y2, P2Y1, P2Y12, P2Y13, P2Y11, P2X1, P2X4), indicating a role in differentiation and proliferation. Thus, it is possible that specific antibodies to P2 receptors could be used to identify progenitors for monocytes, lymphocytes and megakaryocytes. PMID- 15291970 TI - Comparison of the accuracy of methods of computational haplotype inference using a large empirical dataset. AB - BACKGROUND: Analyses of genetic data at the level of haplotypes provide increased accuracy and power to infer genotype-phenotype correlations and evolutionary history of a locus. However, empirical determination of haplotypes is expensive and laborious. Therefore, several methods of inferring haplotypes from unphased genotypic data have been proposed, but it is unclear how accurate each of the methods is or which methods are superior. The accuracy of some of the leading methods of computational haplotype inference (PL-EM, Phase, SNPHAP, Haplotyper) are compared using a large set of 308 empirically determined haplotypes based on 15 SNPs, among which 36 haplotypes were observed to occur. This study presents several advantages over many previous comparisons of haplotype inference methods: a large number of subjects are included, the number of known haplotypes is much smaller than the number of chromosomes surveyed, a range in values of linkage disequilibrium, presence of rare SNP alleles, and considerable dispersion in the frequencies of haplotypes. RESULTS: In contrast to some previous comparisons of haplotype inference methods, there was very little difference in the accuracy of the various methods in terms of either assignment of haplotypes to individuals or estimation of haplotype frequencies. Although none of the methods inferred all of the known haplotypes, the assignment of haplotypes to subjects was about 90% correct for individuals heterozygous for up to three SNPs and was about 80% correct for up to five heterozygous sites. All of the methods identified every haplotype with a frequency above 1%, and none assigned a frequency above 1% to an incorrect haplotype. CONCLUSIONS: All of the methods of haplotype inference have high accuracy and one can have confidence in inferences made by any one of the methods. The ability to identify even rare (>/= 1%) haplotypes is reassuring for efforts to identify haplotypes that contribute to disease in a significant proportion of a population. Assignment of haplotypes is relatively accurate among subjects heterozygous for up to 5 sites, and this might be the largest number of SNPs for which one should define haplotype blocks or have confidence in haplotype assignments. PMID- 15291971 TI - Organization of the mitochondrial genomes of whiteflies, aphids, and psyllids (Hemiptera, Sternorrhyncha). AB - BACKGROUND: With some exceptions, mitochondria within the class Insecta have the same gene content, and generally, a similar gene order allowing the proposal of an ancestral gene order. The principal exceptions are several orders within the Hemipteroid assemblage including the order Thysanoptera, a sister group of the order Hemiptera. Within the Hemiptera, there are available a number of completely sequenced mitochondrial genomes that have a gene order similar to that of the proposed ancestor. None, however, are available from the suborder Sternorryncha that includes whiteflies, psyllids and aphids. RESULTS: We have determined the complete nucleotide sequence of the mitochondrial genomes of six species of whiteflies, one psyllid and one aphid. Two species of whiteflies, one psyllid and one aphid have mitochondrial genomes with a gene order very similar to that of the proposed insect ancestor. The remaining four species of whiteflies had variations in the gene order. In all cases, there was the excision of a DNA fragment encoding for cytochrome oxidase subunit III(COIII)-tRNAgly-NADH dehydrogenase subunit 3(ND3)-tRNAala-tRNAarg-tRNAasn from the ancestral position between genes for ATP synthase subunit 6 and NADH dehydrogenase subunit 5. Based on the position in which all or part of this fragment was inserted, the mitochondria could be subdivided into four different gene arrangement types. PCR amplification spanning from COIII to genes outside the inserted region and sequence determination of the resulting fragments, indicated that different whitefly species could be placed into one of these arrangement types. A phylogenetic analysis of 19 whitefly species based on genes for mitochondrial cytochrome b, NADH dehydrogenase subunit 1, and 16S ribosomal DNA as well as cospeciating endosymbiont 16S and 23S ribosomal DNA indicated a clustering of species that corresponded to the gene arrangement types. CONCLUSIONS: In whiteflies, the region of the mitochondrial genome consisting of genes encoding for COIII-tRNAgly-ND3-tRNAala-tRNAarg-tRNAasn can be transposed from its ancestral position to four different locations on the mitochondrial genome. Related species within clusters established by phylogenetic analysis of host and endosymbiont genes have the same mitochondrial gene arrangement indicating a transposition in the ancestor of these clusters. PMID- 15291972 TI - A steady state analysis indicates that negative feedback regulation of PTP1B by Akt elicits bistability in insulin-stimulated GLUT4 translocation. AB - BACKGROUND: The phenomenon of switch-like response to graded input signal is the theme involved in various signaling pathways in living systems. Positive feedback loops or double negative feedback loops embedded with nonlinearity exhibit these switch-like bistable responses. Such feedback regulations exist in insulin signaling pathway as well. METHODS: In the current manuscript, a steady state analysis of the metabolic insulin-signaling pathway is presented. The threshold concentration of insulin required for glucose transporter GLUT4 translocation was studied with variation in system parameters and component concentrations. The dose response curves of GLUT4 translocation at various concentration of insulin obtained by steady state analysis were quantified in-terms of half saturation constant. RESULTS: We show that, insulin-stimulated GLUT4 translocation can operate as a bistable switch, which ensures that GLUT4 settles between two discrete, but mutually exclusive stable steady states. The threshold concentration of insulin required for GLUT4 translocation changes with variation in system parameters and component concentrations, thus providing insights into possible pathological conditions. CONCLUSION: A steady state analysis indicates that negative feedback regulation of phosphatase PTP1B by Akt elicits bistability in insulin-stimulated GLUT4 translocation. The threshold concentration of insulin required for GLUT4 translocation and the corresponding bistable response at different system parameters and component concentrations was compared with reported experimental observations on specific defects in regulation of the system. PMID- 15291973 TI - What is the clinically relevant endpoint for cancer prevention trials? PMID- 15291974 TI - Novel approaches to immunotherapy for B-cell malignancies. AB - Immunotherapy for cancer refers to a wide array of novel therapeutic interventions that harness the immune system to target and eradicate malignant cells in the host. Advances in the understanding of how tumor cells evade host immune detection, coupled with improved gene transduction technologies, have enabled investigators to propose and test novel immune-based therapies for B-cell malignancies. As a result, more immunogenic vaccination strategies, able to elicit immune responses to otherwise poorly immunogenic tumor antigens, are being tested in early clinical trials. Furthermore, with the development of efficient T cell transduction methodologies, investigators are able to generate autologous antitumor T-cell responses through the introduction of chimeric antigen receptors able to target tumor antigens. However, whether the promising preclinical and phase I clinical data presented here will ultimately translate into improved survival of patients with B-cell malignancies remains largely unknown. PMID- 15291975 TI - Advances in the understanding of biology and prognosis in chronic lymphocytic leukemia. AB - Chronic lymphocytic leukemia (CLL) has a heterogeneous clinical course and outcome. Recent advances in the diagnosis and molecular characterization of CLL permit improved prediction of disease prognosis, which could result in better management. We review the biology and diagnosis of CLL and the role of analysis of immunoglobulin heavy-chain variable region mutation status, molecular cytogenetics (interphase fluorescence in situ hybridization), and the expression of CD38 and ZAP-70 in the evaluation and management of patients with CLL. PMID- 15291976 TI - Variations on the CHOP regimen: II. Prospective study in young patients. PMID- 15291977 TI - Variations on the CHOP regimen: I. Prospective study in the elderly with aggressive non-Hodgkin's lymphomas. PMID- 15291979 TI - New molecular targets for treatment of lymphoma. AB - In recent years, several molecular mechanisms involved in promoting cancer cell survival and growth have been identified. These discoveries helped in designing and testing novel drugs that target specific cellular pathways. In this review, we focus on new molecular targets that are being explored for the treatment of non-Hodgkin's lymphoma and Hodgkin's disease. PMID- 15291980 TI - Molecular diagnosis in lymphoma. AB - The evolution of our ability to diagnose and classify lymphomas in an increasingly refined manner has paralleled the development of novel technologic approaches, with contemporary practice dependent upon the harnessing of a plethora of data that include microscopic, immunophenotypic, and genetic information. Although each of these components is currently indispensable, there is a purported progressive improvement in biologic objectivity as one maneuvers through these respective technologies. Accordingly, and in particular given the rapid pace at which key insights into lymphoma biology are emerging with microarray and other cutting-edge technologies, the role of molecular genetic testing is assuming even greater relevance. The ability to diagnose and classify lymphomas more accurately, precisely, and rationally by incorporating molecular data ought to lead to the development of more appropriate directed therapies. PMID- 15291981 TI - The genetics of familial lymphomas. AB - Whereas familial clustering of malignant lymphoma is well documented, the molecular changes underlying familial lymphoma syndromes remain unclear. An understanding of the hereditary basis of lymphoma may lead to the identification of new molecular markers for disease or novel therapeutic targets. This paper reviews the genetics of familial lymphoma, focusing on germline susceptibilities to lymphoma as well as germline susceptibilities to environmental exposures that have been linked to lymphoma. PMID- 15291983 TI - Treatment of advanced melanoma with temozolomide: I. PMID- 15291982 TI - Primary central nervous system lymphoma. AB - Primary central nervous system lymphoma (PCNSL) is a rare variant of non Hodgkin's lymphoma that is increasing in incidence. Methotrexate-based chemotherapy in combination with whole-brain radiotherapy (WBRT) has dramatically improved the outcome of patients. However, treatment-related neurotoxicity is a significant complication, especially after radiotherapy in the elderly. Despite advances in therapy, several important questions remain regarding optimal methotrexate dose, dosing frequency, adjunct chemotherapy, and the impact of deferring WBRT. Advances in biologic therapy and strategies to intensify the delivery of chemotherapy may help to limit the use of radiotherapy, thus lessening potential neurotoxicity. Studies looking at oncogenic proteins as potential prognostic markers for PCNSL may help us to develop risk-adapted therapies. PMID- 15291984 TI - Treatment of advanced melanoma with temozolomide: II. PMID- 15291985 TI - Melanoma genomics. AB - Sophisticated molecular techniques that have recently become available permit analysis of genome-wide changes in gene copy number (DNA) or expression (RNA). These genomic strategies have the potential to transform our view of cancer biology and pathogenesis. This review surveys the studies performed using genome wide analyses of melanoma, primarily in patient specimens but also in selected experimental models of melanoma. The insights gained and the challenges and potential of these approaches are discussed with respect to their ability to dissect out the molecular events that are responsible for melanoma progression. PMID- 15291986 TI - Chemoprevention of melanoma. AB - The United States is experiencing a surge in the incidence of cutaneous malignant melanoma. Because melanoma is typically refractory to available anticancer therapy, exploration of preventive strategies has become a priority. In this review, the rationale for chemoprevention, a new and potentially powerful approach to controlling melanoma, is discussed. Chemoprevention success is based on the principles that ultraviolet-induced melanoma is a multistep process, and that molecular events and pathways associated with these steps can be targeted. Early studies using genetically engineered mice have begun to identify a number of relevant molecular pathways in melanoma. For example, Ras signaling pathways comprise all melanoma-related alterations in N-Ras, B-RAF, MAPK/ERK, and Rho proteins, and thus provide a host of potential molecular targets for melanoma chemoprevention. Among the available prospects, the statins, which inhibit Ras and Rho, have shown much promise as chemoprevention agents. However, thorough evaluation of chemoprevention candidates will require the identification of surrogate biomarkers for risk and molecular targets for intervention, as well as high-risk groups in which to focus clinical studies. We anticipate that melanoma chemoprevention research will progress in step with advances in genomics, proteomics, and preclinical mouse modeling, and ultimately provide us with powerful weapons in our struggle to control this escalating, often fatal disease. PMID- 15291989 TI - Emergency contraception: politics trumps science at the U.S. Food and Drug Administration. PMID- 15291987 TI - Overview of melanoma vaccines and promising approaches. AB - It is difficult to envision anything better than melanoma vaccines to exemplify the effectiveness of modern biotechnology in developing biologically rational therapeutics. Melanoma vaccines can reproducibly induce cytotoxic T lymphocyte (CTL) responses better than any other anticancer therapy. Anticancer vaccines have been labeled by some as ineffective for the simple reason that they only rarely lead to cancer regression. This oxymoron stems from the naive expectation that CTLs are all that is needed to reject cancer. Little is known about requirements for CTL localization and effector function within the tumor microenvironment. In the future, more attention should be given to events downstream of immunization (afferent arm of immune response) to identify combination therapies likely to facilitate localization and activation of CTL at the receiving end (efferent arm). PMID- 15291991 TI - Intraventricular hemorrhage and periventricular leukomalacia in preterm infants. AB - OBJECTIVE: To evaluate whether intraventricular hemorrhage and periventricular leukomalacia are characterized by different risk factors. METHODS: In a cohort of 653 consecutive singleton neonates born after preterm membrane rupture, spontaneous preterm labor, or indicated preterm delivery at 24 to 33 weeks of gestation from January 1, 1993, to December 31, 2002, we evaluated the obstetric and histopathologic placental variables in reference to the development of intraventricular hemorrhage (n = 44), periventricular leukomalacia (n = 19), or no ultrasonographic cerebral lesion (n = 589). Excluded were stillbirths and congenital anomalies. Statistical analysis included Fisher exact test, Student t test, and stepwise logistic regression analysis with a 2-tailed P <.05 considered significant. RESULTS: Multivariate analysis showed that occurrence of neonatal intraventricular hemorrhage and periventricular leukomalacia were associated only with spontaneous prematurity (odds ratio = 1.9; 95% confidence interval 1.1-3.4) and gestational age at delivery in weeks (odds ratio = 0.8; 95% confidence interval 0.7-0.9). Neonates with intraventricular hemorrhage did not differ from those with periventricular leukomalacia in any obstetric or neonatal variable, but there was a higher risk of neurodevelopmental delay associated with periventricular leukomalacia. CONCLUSION: Among premature infants born at less than 34.0 weeks of gestation, intraventricular hemorrhage and periventricular leukomalacia share common clinical characteristics, with spontaneous preterm delivery and gestational age at delivery as the only independent antenatal predictors. PMID- 15291992 TI - Multiple pregnancy: knowledge and practice patterns of obstetricians and gynecologists. AB - OBJECTIVE: We sought to assess knowledge and practices of obstetricians regarding multiple gestation. METHODS: A questionnaire investigating knowledge and practice patterns pertaining to multiple gestation was mailed to randomly selected American College of Obstetricians and Gynecologists (ACOG) Fellows and Junior Fellows in January 2003. Fifty-one percent (589/1,146) of the surveys were returned. RESULTS: Statistical analysis included the responses from 430 practicing obstetricians. More than 60% rated their training regarding management of multiples as adequate. Men (56.5%) were older and had been in practice longer than females. Sixty-two percent of general obstetrician-gynecologists managed twins without input from a maternal-fetal medicine specialist. An understanding of chorionicity was less than anticipated. The majority of practitioners managed twins according to recent ACOG educational materials. They did not use prophylactic cerclage, home uterine-activity monitoring, or tocolytics. Fort-six percent encouraged prophylactic bed rest. The management of breech second twins varied. CONCLUSION: Most obstetricians manage multiples according to current ACOG educational materials independent of maternal-fetal medicine specialists. This survey identified knowledge gaps, specifically in chorionicity, indicating the need to develop educational strategies addressing these insufficiencies. PMID- 15291993 TI - Does human immunodeficiency virus infection protect against preeclampsia eclampsia? AB - OBJECTIVE: In view of recent suggestions that human immunodeficiency virus (HIV) infection may protect against preeclampsia, this study was done to evaluate whether untreated HIV-positive pregnant women have a lower rate of preeclampsia eclampsia than HIV-negative women. METHODS: Subjects for this study were pregnant women from Soweto, South Africa, who gave birth from March to December 2002 at midwife-run clinics or at the Chris Hani Baragwanath Hospital and in whom the HIV status was known. A sample size calculation indicated that 2,588 subjects would be required to show statistical significance at P <.05 with a power of 80% for a reduction in the rate of preeclampsia from 8% to 5% with HIV seropositivity, assuming an HIV seroprevalence rate of 30%. Data collection was by record review from randomly selected patient files and birth registers. RESULTS: In the total sample of 2,600 women, 1,797 gave birth at the hospital and 803 at the midwife run clinics. The HIV seroprevalence rate was 27.1%. Hypertension was found in 17.3% of women, with 5.3% having preeclampsia-eclampsia. The rates of preeclampsia-eclampsia were 5.2% in HIV-negative and 5.7% in HIV-positive women (P =.61). CD4 count results were available for only 13 women (0.5%). CONCLUSION: Human immunodeficiency virus seropositivity was not associated with any reduction in the risk of developing preeclampsia-eclampsia. PMID- 15291994 TI - Bupivacaine plus epinephrine for laparoscopic myomectomy: a randomized placebo controlled trial. AB - OBJECTIVE: To evaluate the effectiveness of the injection of bupivacaine plus epinephrine before laparoscopic myomectomy. METHODS: Sixty premenopausal women with uterine leiomyomata were enrolled in a randomized controlled design and intraoperatively treated with injection of bupivacaine plus epinephrine (group A) or saline solution (group B) during laparoscopic myomectomy. Uterine size and volume, number of leiomyomata, hematological parameters, total operative time, enucleation time of each myoma, suturing time of the myomectomy, blood loss, degree of surgical difficulty, and postoperative pain were evaluated. Just before and after the injection of vasoconstrictive or saline solution, systolic and diastolic blood pressure and heart rate were recorded in each subject. RESULTS: Blood loss, total operative and enucleation time, and degree of surgical difficulty was significantly (P <.05) lower in group A than in group B. No difference was observed between groups in suturing time of the myomectomy. The number of vials of pain medication used postoperatively was significantly (P <.05) lower in group A than in group B. No differences in systolic and diastolic blood pressure or heart rate was recorded between the 2 groups. CONCLUSION: The injection of bupivacaine plus epinephrine during laparoscopic myomectomy is effective in reducing blood loss, total operative and enucleation time, degree of surgical difficulty, and postoperative pain. PMID- 15291995 TI - The clinical significance of a negative loop electrosurgical cone biopsy for high grade dysplasia. AB - OBJECTIVE: We sought to estimate the incidence and clinical significance of a negative therapeutic loop electrosurgical excision procedure (LEEP) and to evaluate patient specimens for limiting histologic features associated with a negative LEEP. METHODS: We identified 674 patients with biopsy-confirmed high grade cervical dysplasia who were treated with LEEP from 1991 through 2001. The results of these LEEP procedures were reviewed for the absence of dysplasia or the presence of cervical intraepithelial neoplasia stages 1-3. Computerized pathology files of patients were then reviewed through July 2002 to determine whether dysplasia recurred. Slides of negative LEEP specimens were reviewed to confirm the absence of dysplasia and to search for histologic features that may have limited our interpretation of the specimen. RESULTS: Ninety-three (14%) of LEEP specimens reviewed were completely negative for dysplasia. Clinical follow up was available on 75 of the 93 patients, with a median follow-up time of 2 years. Eighteen (24%) patients had subsequent positive follow-up, including carcinoma (n = 2), high-grade squamous intraepithelial lesions (n = 8), low-grade squamous intraepithelial lesion (n = 6), and atypical squamous cells of undetermined significance (n = 2). Patients with negative LEEPs had a recurrence rate similar to patients with positive LEEPs (24% versus 27%). Limiting histologic features were more commonly identified in negative LEEPs as compared with LEEPs containing dysplasia (16% versus 5%, P <.001). CONCLUSION: A negative LEEP is not an uncommon finding, occurring in 14% (95% confidence interval 11 17%) of specimens at our institution. Negative LEEPs are more likely to contain histologic features that limit pathology interpretation. A negative LEEP is not a reassuring finding and was associated with a recurrence rate similar to those of a positive LEEP. Both negative and positive populations should be carefully followed. PMID- 15291996 TI - Does tamoxifen use affect prognosis in breast cancer patients who develop endometrial cancer? AB - OBJECTIVE: The use of tamoxifen to prevent breast cancer and decrease recurrence is not controversial. However, the effect that tamoxifen may have in women with a history of breast cancer in whom endometrial cancer develops is unclear. The purpose of this study was to estimate whether a history of tamoxifen use is a prognostic factor for such patients. METHODS: Between 1990 and 2002, patients seen at The University of Texas M. D. Anderson Cancer Center with a history of breast cancer who developed endometrial cancer were identified. Medical records were reviewed to identify clinical, pathologic, and outcome information. RESULTS: Eighty-nine patients with a history of breast cancer in whom endometrial carcinoma developed were identified. Fifty-two percent (46/89) had a history of tamoxifen use (median duration 48 months; range 2-120 months). There were no significant differences in the clinical or pathologic features between tamoxifen users and nonusers. A history of tamoxifen use was associated with a shorter interval from breast cancer to endometrial cancer diagnosis (77.2 versus 121.3 months for nonusers; P =.01). There was no significant difference in overall survival between tamoxifen users and nonusers (39.2 months versus 48.3 months, P =.27), and there was no difference in endometrial cancer-specific survival duration between tamoxifen users and nonusers (55.7 versus 51.0 months, P =.92). CONCLUSION: Among tamoxifen users, the interval from breast cancer to endometrial cancer diagnosis was significantly shorter than that in nonusers. In this cohort, a history of tamoxifen use was not associated with a worse overall or disease specific survival. PMID- 15291997 TI - Surgical staging of ovarian low malignant potential tumors. AB - OBJECTIVE: Women diagnosed with ovarian tumors of low malignant potential have an excellent prognosis. Because few will receive adjuvant therapy, the benefit of surgical staging has recently been challenged. The purpose of this study was to compare the outcome of surgically staged patients with low malignant potential tumors with those who were not staged. METHODS: Between 1984 and 2003, all women with ovarian low malignant potential tumors were identified at 3 institutions. Data were extracted from clinical records. RESULTS: One hundred eighty-three (74%) of 248 women were surgically staged. Forty of 183 staged patients had clinically obvious extraovarian disease. Forty (28%) of the remaining 143 women with disease apparently confined to the ovary were upstaged. Cytologic washings were positive in 28 cases, 10 had microscopic implants detected by peritoneal or omental biopsy, and 2 were upstaged to stage IIIC solely on the basis of nodal metastases. One hundred eighteen women underwent pelvic node dissection (median: 5 nodes), and 86 underwent para-aortic node dissection (median: 2 nodes). Overall, 9 (1%) metastases were detected in 832 submitted pelvic nodes. All 314 para-aortic nodes were negative. Intraoperative blood loss (P <.001) and length of hospital stay (P <.001) were increased in women without gross disease who were surgically staged. Eight (3%) of 248 patients received adjuvant platinum-based chemotherapy, but neither of the women upstaged to IIIC based on the results of their nodal dissection were treated. Fifteen (6%) recurrences developed and 1 (0.4%) death occurred after a median follow-up of 28 (range, 1-208) months. CONCLUSION: Routine pelvic and para-aortic lymph node dissection is not necessary in the majority of women with ovarian low malignant potential tumors. PMID- 15291998 TI - Vulvovaginal symptoms in women with bacterial vaginosis. AB - OBJECTIVE: A substantial, but highly variable, percentage of women with bacterial vaginosis are said to be asymptomatic. The purpose of this study was to estimate the prevalence of symptoms among women with bacterial vaginosis compared with women without bacterial vaginosis by direct, explicit, and detailed questioning of these women. METHODS: Women presenting for a routine health care visit at 12 health department clinics in Birmingham, Alabama, were recruited to participate in a longitudinal study of vaginal flora. At the first visit, they underwent a pelvic examination, lower genital tract microbiological evaluation, and an interview that included detailed questions regarding lower genital tract symptoms. The prevalence of symptoms among women with and without bacterial vaginosis (Gram stain score 7 or higher) was compared. RESULTS: Among 2,888 women without gonorrhea, Chlamydia, or trichomonas, 75% of women with and 82% of women without bacterial vaginosis never noted any vaginal odor in the past 6 months (P <.001). The corresponding values were 63% and 65% for never noting vaginal "wetness" (P =.02); 58% and 57% for vaginal discharge (P =.65); 91% and 86% for irritation (P =.004); 88% and 85% for itching (P =.64); and 96% and 94% for dysuria (P =.002), respectively. Cumulatively, 58% of women with bacterial vaginosis noted odor, discharge, and/or wetness in the past 6 months compared with 57% of women without bacterial vaginosis (P =.70). CONCLUSION: The 2 classic symptoms of bacterial vaginosis discharge and odor are each reported by a minority of women with bacterial vaginosis and are only slightly more prevalent than among women without bacterial vaginosis. PMID- 15291999 TI - Effect of prior vaginal delivery or prior vaginal birth after cesarean delivery on obstetric outcomes in women undergoing trial of labor. AB - OBJECTIVE: We sought to study the effects of prior vaginal delivery or prior vaginal birth after cesarean delivery (VBAC) on the success of a trial of labor after a cesarean delivery. METHODS: An observational study of patients who underwent a trial of labor after a single low-transverse cesarean delivery. Patients with a previous cesarean delivery and no vaginal birth were compared with patients with a single vaginal delivery before or after the previous cesarean delivery. The rates of successful VBAC, uterine rupture, and scar dehiscence were analyzed. Multivariable regression was performed to adjust for confounding variables. RESULTS: Of 2,204 patients, 1,685 (76.4%) had a previous cesarean delivery and no vaginal delivery, 198 (9.0%) had a vaginal delivery before the cesarean delivery, and 321 (14.6%) had a prior VBAC. The rate of successful trial of labor was 70.1%, 81.8%, and 93.1%, respectively (P <.001). A prior VBAC was associated with fewer third- and fourth-degree lacerations (8.5% versus 2.5% versus 3.7%, P <.001) and fewer operative vaginal deliveries (14.7% versus 5.6% versus 1.9%, P <.001) but not with uterine rupture (1.5% versus 0.5% versus 0.3%, P =.12). Patients with a prior VBAC had, in addition, a higher rate of uterine scar dehiscence (21.8%) compared with patients with a previous cesarean delivery and no vaginal delivery (5.3%; P =.001). CONCLUSION: A prior vaginal delivery and, particularly, a prior VBAC are associated with a higher rate of successful trial of labor compared with patients with no prior vaginal delivery. In addition, prior VBAC is associated with an increased rate of uterine scar dehiscence. PMID- 15292000 TI - Twin deliveries in the United States over three decades: an age-period-cohort analysis. AB - OBJECTIVE: Time is an important variable in understanding the recent increase in twin deliveries in the United States. Therefore, this study was designed to estimate the influences of maternal age, period (year) of delivery, and maternal birth-year cohort on trends in rates of twin deliveries. METHODS: United States natality data were used to assess trends in twin pregnancies resulting in live births. This age-period-cohort analysis included 7, 5-year maternal-age groups (15-19 through 45-49 years), 6 twin delivery periods (1975, 1980, 1985, 1990, 1995, and 2000), and 12, 5-year maternal birth cohorts (1926-1930 through 1981 1985). The independent effects of maternal age, twin delivery period, and maternal birth cohort on twin delivery rates for blacks and whites were modeled using Poisson regression techniques. RESULTS: Our study assessed 95,042 blacks and 401,989 whites with twin deliveries. Twin deliveries increased by 46% for blacks and 62% for whites from 1975 to 2000, with the largest increase occurring in the year 2000. For blacks, maternal age had the strongest impact on the increasing twin delivery rates, followed by period of delivery. For whites, the greatest effect was due to period of delivery, followed by maternal birth year cohort and, lastly, maternal age. CONCLUSION: Our data confirm the importance of nature's biologic contribution of maternal aging to twin delivery rates, but suggest that recent changes in the environment surrounding pregnancy (nurture) also influence twin delivery rates. The relative contributions of biologic versus environmental influences appear to differ among blacks and whites. PMID- 15292001 TI - Body mass index change between pregnancies and small for gestational age births. AB - OBJECTIVE: To estimate whether maternal weight changes between pregnancies influence the risk for small for gestational age (SGA) births. METHODS: SGA cases (n = 8,062) below the tenth percentile birth weight for gestational age were selected from liveborn singletons born of Missouri residents during 1989-1997. Normal weight controls (n = 8,062) were selected according to birth year. The risk of SGA from interpregnancy body mass index (BMI) change and other maternal factors was estimated using logistic regression analysis. RESULTS: An increase in BMI between pregnancies decreased SGA risk (adjusted odds ratio = 0.8; 95% confidence interval 0.7, 1.0). Other risk factors were prior SGA (4.4; 4.0, 4.8), preeclampsia/eclampsia (2.6; 2.1, 3.2), maternal cardiac disease (1.8; 1.1, 2.9), inadequate weight gain (1.9; 1.8, 2.2), and cigarette smoking (1.9; 1.7, 2.3 for 1-9 cigarettes per day; 2.5; 2.2, 2.8 for 10-19/d; and 2.8; 2.5, 3.3 for 20/d or more). CONCLUSION: Increase in interpregnancy BMI lowers SGA risk, but adequate weight gain during pregnancy is more effective. PMID- 15292002 TI - Ethnic differences of polymorphisms in cytokine and innate immune system genes in pregnant women. AB - OBJECTIVE: Investigations of the possible role of polymorphic genes in pregnancy outcome may be influenced by ethnic variations in genotype or allele frequencies. Differences in allelic carriage of immune system-related genes among white, black, and Hispanic pregnant women living in New York City and Boston were evaluated. METHODS: DNA was extracted from buccal or vaginal epithelial cells collected from 198 white, 75 black, and 114 Hispanic pregnant women who delivered at term and who had no history of a preterm birth. Genetic polymorphisms in the immunoregulatory genes encoding interleukin (IL)-1beta, tumor necrosis factor alpha, IL-4, IL-10, IL-1 receptor antagonist (IL-1ra), mannose-binding lectin, toll-like receptor-4, and the 70-kDa heat shock protein were determined. RESULTS: Allele 2 of the IL-1ra gene (IL1RN*2) and IL-4 -590C homozygosity were 4-fold less common in blacks than in whites or Hispanics (P <.001). The IL-4 -590T allele was almost 2-fold more common in Hispanics than in whites (P <.001). The frequency of the 70-kDa heat shock protein 1267G allele was at least 1.4 times greater in blacks compared with whites (P <.001) or Hispanics (P =.002), whereas the homozygous mannose-binding lectin codon 54G allele was observed at least 4.5 times more often in Hispanics compared with whites (P =.007) or blacks (P =.02). CONCLUSION: Investigations of the role of genetic factors affecting pregnancy outcome must be cognizant of ethnic variations when enrolling case and control subjects for studies on allele and genotype frequencies. PMID- 15292003 TI - Urinary incontinence in elderly women: findings from the Health, Aging, and Body Composition Study. AB - OBJECTIVE: To estimate the prevalence of and risk factors for stress and urge incontinence in a biracial sample of well-functioning older women. METHODS: We performed a cross-sectional analysis of 1,584 white and black women, aged 70-79 years, enrolled in a longitudinal cohort study. Participants were asked about incontinence, medical problems, and demographic and reproductive characteristics and underwent physical measurements. Using multivariable logistic regression, we compared women reporting at least weekly incontinence with those without incontinence. RESULTS: Overall, 21% reported incontinence at least weekly. Of these, 42% reported predominantly urge incontinence, and 40% reported stress. Nearly twice as many white women as black women reported weekly incontinence (27% versus 14%, P <.001). Factors associated with urge incontinence included white race (odds ratio [OR] 3.1, 95% confidence interval [CI] 2.0-4.8), diabetes treated with insulin (OR 3.5, 95% CI 1.6-7.9), depressive symptoms (OR 2.7, 95% CI 1.4-5.3), current oral estrogen use (OR 1.7, 95% CI 1.1-2.6), arthritis (OR 1.7, 95% CI 1.1-2.6), and decreased physical performance (OR 1.6 per point on 0-4 scale, 95% CI 1.1-2.3). Factors associated with stress incontinence were chronic obstructive pulmonary disease (OR 5.6, 95% CI 1.3-23.2), white race (OR 4.1, 95% CI 2.5-6.7), current oral estrogen use (OR 2.0, 95% CI 1.3-3.1), arthritis (OR 1.6, 95% CI 1.0-2.4), and high body mass index (OR 1.3 per 5 kg/m2, 95% CI 1.1 1.6). CONCLUSION: Urinary incontinence is highly prevalent, even in well functioning older women, whites in particular. Many risk factors differ for stress and urge incontinence, suggesting differing etiologies and prevention strategies. PMID- 15292004 TI - Learning laparoscopic-assisted hysterectomy. AB - OBJECTIVE: The aim of this study was to evaluate the factors considered for proficiency and to estimate the number of procedures needed to achieve competence in laparoscopic-assisted vaginal hysterectomy in a teaching hospital. METHODS: The length of the learning curve, duration of surgery, change of hemoglobin (in grams per liter), conversion rate, and intra- and postoperative complications were evaluated. Cases were analyzed according to the order for the individual surgeon. RESULTS: Thirty-three surgeons performed 929 laparoscopic-assisted vaginal hysterectomies during the study period. Analyzing the duration of surgery and rate of complications, we decided on a cutoff of 30 cases. Eight surgeons with more than 30 cases performed 668 laparoscopic-assisted vaginal hysterectomies. Their initial 30 cases (group A, the first 30 cases) were compared with their subsequent cases (group B, cases 31 and after). Patient age, body mass index, and uterine weight did not differ between the groups. The intraoperative complication rate dropped from 4.2% to 0.5% (P =.001), hemoglobin drop decreased from -0.8 +/- 0.9 g/L to -0.5 +/- 1.0 g/L (P =.002), and postoperative complications dropped from 12.9% to 7.0% (P =.017). The duration of surgery was also shorter (148.8 +/- 45.4 minutes versus 125.1 +/- 46.5 minutes), but this difference was taken from the results of 1 surgeon. CONCLUSION: A learning experience of 30 laparoscopic-assisted vaginal hysterectomies was necessary in our institution to reach a low level of complications. Duration of the surgical procedure was not an adequate study endpoint to assess a learning effect. PMID- 15292005 TI - Reproductive consequences of contraceptive failure in 19 developing countries. AB - OBJECTIVE: To ascertain the contribution of contraceptive failure to unintended births and fetal loss in developing countries. METHODS: Nationally representative survey data from married women in 19 developing countries were analyzed. All surveys contained retrospective monthly calendars of contraceptive use and pregnancies for a 5-year period preceding each survey. Information on the intendedness of live births, ascertained earlier in the interview, were linked to the calendar data. Single-decrement life table analysis was applied to episodes of use to estimate failure probabilities. The reproductive consequences of failure were established by simple tabulation. Logistic regression was used to explore the determinants of fetal loss. RESULTS: Reported contraceptive failure rates were similar to those derived from studies conducted mainly in the United States. About three fourths of pregnancies resulting from contraceptive failure were carried to term, and all but 16% of those were classified by the mother as unwanted or mistimed. Just over one tenth ended in fetal loss, either induced or spontaneous. Analysis of determinants of fetal loss suggested that a large proportion were induced. The median contribution of failure to all unintended births for all 19 surveys was about 15%, and the contribution to fetal loss was 12%. CONCLUSION: The contribution of contraceptive failure in developing countries is much lower than the estimate of 50% in the United States. Despite the substantial increases in contraceptive practice that have occurred in Asia, Latin America, the Middle East, and to a lesser extent, Africa, the level of use is still below the 75% mark achieved in most industrialized countries. Nonuse of contraception remains the dominant direct cause of unintended births, and family planning promotion should remain a public health priority. PMID- 15292006 TI - Renal blood flow alteration after paracentesis in women with ovarian hyperstimulation. AB - OBJECTIVE: To evaluate renal arterial resistance to flow by Doppler indices concurrently with ascites drainage in women with severe ovarian hyperstimulation syndrome. METHODS: We conducted an interventional clinical study of 19 women with severe ovarian hyperstimulation syndrome, manifested by free peritoneal fluid. The subjects were evaluated before and after therapeutic paracentesis by measuring urine output, blood urea nitrogen, intra-abdominal pressure, and renal artery flow measures by Doppler ultrasonography (systolic/diastolic ratio [S/D] and resistance index). RESULTS: An average of 3,340 mL of ascitic fluid was drained, and the intra-abdominal pressure decreased from 17.5 +/- 1.24 cm H2O to 10 +/- 1.22 cm H2O. Urine output was increased (by 65%, from 925 +/- 248 mL/d before paracentesis to 1,523 +/- 526 mL/d on the day after paracentesis, P <.001). The mean renal arterial S/D decreased from 3 +/- 0.15 to 2.29 +/- 0.13 (P =.001). Most of the decrease in intra-abdominal pressure as well as in renal vasculature resistance was apparent after an initial drainage of 2,000 mL. Additional fluid drainage had only negligible effect on intra-abdominal pressure and renal flow. CONCLUSION: Paracentesis lowered intra-abdominal pressure and decreased renal arterial resistance (lowered S/D and resistance index), ultimately resulting in increased urine production. It is plausible that the beneficial effects of paracentesis on urine output in ovarian hyperstimulation syndrome are due to improved renal blood flow from a direct decompression effect. PMID- 15292007 TI - Quantitative electromyography of the anal sphincter after uncomplicated vaginal delivery. AB - OBJECTIVE: Fecal incontinence in women is thought to be associated with sphincter laceration or pudendal nerve damage. A prolonged pudendal nerve terminal motor latency is evidence of profound nerve damage, but pudendal nerve terminal motor latency can be normal even when nerve injury has been sustained. We performed quantitative electromyography (EMG) to compare multiple motor unit action potential parameters between recently postpartum women and nulliparous women. METHODS: Standardized examinations were prospectively performed on 2 groups: 1) healthy nulliparous women without pelvic floor disorders (n = 28) and 2) asymptomatic women who were postpartum following vaginal delivery of their first child (n = 23). The examinations included pelvic organ prolapse quantification measurements, endoanal ultrasonography, pudendal nerve terminal motor latency, sacral reflexes, and concentric needle EMG using multiple motor unit action potential analysis. RESULTS: A mean of 11.5 (standard deviation [sd] 1.1) weeks had elapsed since first vaginal deliveries in the postpartum group. The mean fetal weight at delivery was 3,495 (sd 458) grams. There were no sphincter defects seen by ultrasonography. Compared with the nulliparous women, pudendal nerve terminal motor latency and sacral reflexes (clitoral-anal reflex, urethral anal reflex) were not increased in the postpartum group. Each of the quantitative parameters (duration, amplitude, area, turns, and phases), measured from motor unit action potentials in the postpartum group, were larger than in the nulliparous group (P < or =.004, nested analysis of variance [ANOVA]). CONCLUSION: Quantitative EMG using multiple motor unit action potential analysis can detect the presence after vaginal childbirth of subtle nerve injury not demonstrable by pudendal nerve terminal motor latency. Even asymptomatic women show evidence of pelvic floor nerve injury after uncomplicated deliveries. PMID- 15292008 TI - Hyperhomocysteinemia, pregnancy complications, and the timing of investigation. AB - OBJECTIVE: To assess associations between vitamin-dependent homocysteine metabolism and vascular-related pregnancy complications by considering interval between delivery and postpartum investigation and maternal age. METHODS: Case control study performed at the University Medical Center Nijmegen in the Netherlands. Patients had experienced pregnancy-induced hypertension (n = 37), preeclampsia (n = 144), hemolysis, elevated liver enzymes, low platelets (HELLP) syndrome (n = 104), recurrent early pregnancy loss (n = 544), abruptio placentae (n = 135), intrauterine growth restriction (n = 144), or intrauterine fetal death (n = 104). Controls comprised 176 women with uncomplicated obstetric histories. Oral methionine loading tests and fasting vitamin profiles were performed more than 6 weeks after delivery. Odds ratios and 95% confidence intervals were calculated after logistic regression analysis. RESULTS: Hyperhomocysteinemia was associated with an approximately 2-fold to 3-fold increased risk for pregnancy induced hypertension, abruptio placentae, and intrauterine growth restriction. Cobalamin deficiency was associated with HELLP syndrome, abruptio placentae, intrauterine growth restriction, and intrauterine fetal death. Pyridoxal 5 phosphate deficiency increased the risk for pregnancy-induced hypertension 4 fold. These associations lost their significance after adjustment for time interval and maternal age. High red cell folate was associated with a decreased risk for abruptio placentae and intrauterine growth restriction. An increased creatinine concentration was associated with pregnancy-induced hypertension, preeclampsia, HELLP syndrome, and abruptio placentae. CONCLUSION: Hyperhomocysteinemia and vitamin deficiencies are largely determined by the interval between delivery and postpartum investigation and by maternal age. Time interval and maternal age should be considered in the risk estimation for vascular-related pregnancy complications. PMID- 15292009 TI - Maternal and neonatal outcomes in pregnancies complicated by bone and soft-tissue tumors. AB - OBJECTIVE: Primary bone and soft-tissue tumors occur rarely in pregnancy. The objective of this study was to describe the outcome of a large cohort of pregnant patients with these rare tumors. METHODS: Pregnant women diagnosed with bone or soft-tissue tumors during pregnancy or within 3 months after delivery were identified retrospectively for the years 1983-2003 in the University Health Network database, University of Toronto. Relevant maternal and neonatal data were collected on a standardized data form. RESULTS: In more than 60,000 deliveries during the study period, 17 patients were identified. Gestational age at diagnosis ranged from 11 weeks to 2 months postpartum. Eight cases involved the lower extremity and 6 involved the upper extremity. Osteosarcoma, chondrosarcoma, and giant-cell tumors were the most common histological types. Metastases occurred in 7 cases. Nine cases were treated surgically during the course of pregnancy. The majority of patients were delivered at term. Chemotherapy was deferred until the postpartum period. One patient elected for early termination of pregnancy. Three patients were delivered before 37 weeks of gestation to proceed with therapy. One neonate delivered at 34 weeks developed respiratory distress syndrome and required intubation. Three patients died, all as the result of metastatic disease. There were no perinatal or infant deaths. CONCLUSION: Most cases of soft-tissue and bone tumors during pregnancy can be successfully managed with surgery during gestation. Therapies with fetal toxicity were more likely to be deferred to the postpartum period. PMID- 15292010 TI - Small-group discussion versus lecture format for third-year students in obstetrics and gynecology. AB - OBJECTIVE: To compare lecture and small-group discussion for third-year medical students in obstetrics and gynecology. METHODS: Over a 2-year period, 91 third year medical students in the obstetrics and gynecology clerkships were given educational sessions on diabetes and hypertension in pregnancy by a single instructor, either in a traditional lecture format or in a small-group discussion. After the instructional sessions, students anonymously completed a 20 question multiple-choice examination on the covered topics. They also completed an evaluation form on the instructional format, using a 5-point Likert scale, ranging from 1 (strongly disagree) to 5 (strongly agree). At the completion of each clerkship, students repeated the same multiple-choice examination. RESULTS: There was a significantly higher level of enjoyment (median value 5 versus 4, P <.001) and sense of educational stimulation (median value 5 versus 4, P <.001) in the discussion group, and students in the discussion group were less desirous of the alternate instructional format than those in the lecture group (median value 2 versus 3, P <.001). However, there were no differences in the test scores, either immediately after the instructional sessions or at the end of the clerkships. There was a 90% power to detect a 15% difference in postinstructional test scores. CONCLUSION: Third-year medical students learning about hypertension and diabetes in pregnancy during their obstetrics and gynecology clerkship strongly preferred small-group discussions over traditional lectures. However, this preference did not lead to improved test scores on these subjects. PMID- 15292011 TI - Effects of low-molecular-weight and unfractionated heparin on trophoblast function. AB - OBJECTIVE: Unfractionated and low-molecular-weight heparin and low-dose aspirin are used for the prevention of pregnancy loss in pregnant women with thrombophilia. We investigated the effect of these drugs on in vitro models of human extravillous trophoblast motility and differentiation. METHODS: Chorion from term placentas was digested and extravillous trophoblast isolated. Extravillous trophoblast formed giant multinuclear cells that were counted after 24, 36, and 48 hours of culture. This model was then used to investigate the effect of unfractionated, low-molecular-weight heparin and aspirin on in vitro extravillous trophoblast differentiation at both therapeutic and supratherapeutic doses. In addition, the effect of unfractionated and low-molecular-weight heparin on hepatocyte growth factor-stimulated SGHPL-4 cell (extravillous trophoblast cell line) motility was determined by time-lapse microscopy. RESULTS: At therapeutic doses unfractionated heparin promoted extravillous trophoblast differentiation. However, low-molecular-weight heparin inhibited giant multinuclear cells formation. At supratherapeutic doses, both low-molecular weight and unfractionated heparin promoted extravillous trophoblast differentiation. Low-dose aspirin had minimal effects on the extravillous trophoblast differentiation. Both unfractionated and low-molecular-weight heparin inhibited hepatocyte growth factor-stimulated extravillous trophoblast motility at supratherapeutic doses. At a therapeutic dose of 0.25 IU/mL, only unfractionated heparin inhibited hepatocyte growth factor-stimulated motility, whereas low-molecular-weight heparin had no effect. CONCLUSION: Our data suggest that unfractionated and low-molecular-weight heparin have differing effects on trophoblast differentiation and motility at therapeutic doses. This finding may be one of many factors that contribute to the clinical scenario. PMID- 15292012 TI - Noninvasive ultrasound assessment of maternal vascular reactivity during pregnancy: a longitudinal study. AB - OBJECTIVE: To estimate the pattern of maternal vascular reactivity in normal and high-risk pregnancies using postocclusion brachial artery diameter. METHODS: Prospective, longitudinal study of 44 low-risk singleton pregnancies and 28 high risk pregnancies, defined as pregestational diabetes (n = 7), chronic hypertension (n = 4), twin gestation (n = 6), and a previous history of preeclampsia, fetal growth restriction, or vascular disease (n = 11). During each trimester, the brachial artery was ultrasonographically imaged above the antecubital crease. Brachial artery diameter was measured and then occluded for 5 minutes using an inflated blood pressure cuff. Changes in brachial artery diameter at 1 minute after occlusion were expressed as percent change from baseline and were compared across trimesters for both low-risk and high-risk groups, adjusting for potential confounders. RESULTS: Brachial artery diameters were increased after occlusion in every trimester for all groups. For low-risk women, the degree of postocclusion brachial artery dilatation was similar in the first and second trimesters, but was lower in the third trimester. In the first trimester, low-risk women had significantly greater brachial artery diameter increases at 1 minute compared with high-risk singleton pregnancies (19% compared with 12%; P <.001). Compared with low-risk women, pregnancies complicated by pregestational diabetes or chronic hypertension had significantly smaller 1 minute brachial artery diameter changes in the first trimester (7.0 +/- 0.5%, P <.001), whereas twin gestations had greater brachial artery responses (22.9 +/- 6.0%, P <.001). Women with previous preeclampsia or vascular disease had responses similar to low-risk women. CONCLUSION: Maternal vascular reactivity as assessed by postocclusion brachial artery dilatation decreases in the third trimester in both low-risk and high-risk women. In addition, singleton pregnancies at high risk for preeclampsia display decreased brachial artery reactivity compared with low-risk women. PMID- 15292013 TI - Random urine protein-creatinine ratio to predict proteinuria in new-onset mild hypertension in late pregnancy. AB - OBJECTIVE: The aim of this study was to evaluate the diagnostic accuracy of random urine protein-creatinine ratio for prediction of significant proteinuria (> or = 300 mg/24 h) in patients with new-onset mild hypertension in late pregnancy. METHODS: Medical records of 185 consecutive pregnant patients with new onset of mild hypertension in late pregnancy were reviewed. Random urine samples were taken before 24-hour urine collection. The predictive values of the random urine protein-creatinine ratio for diagnosis of significant proteinuria were estimated by using at least a 300-mg protein level within the collected 24-hour urine as the gold standard. RESULTS: Thirty-nine patients (21%) had significant proteinuria. There was a significant association between 24-hour protein excretion and the random urine protein-creatinine ratio (rs = 0.56, P <.01). With a cutoff protein-creatinine ratio greater than 0.19 as a predictor of significant proteinuria, sensitivity and specificity were 85% and 73%, respectively. Positive and negative predictive values of the test were 46% and 95%, respectively. CONCLUSION: The random urine protein-creatinine ratio was a poor predictor for significant proteinuria in patients with new-onset mild hypertension in late pregnancy. PMID- 15292014 TI - Nucleated red blood cells in uncomplicated prolonged pregnancy. AB - OBJECTIVE: Elevated counts of nucleated red blood cells (NRBCs), as well as prolongation of pregnancy, have been suggested as predictors of adverse perinatal outcome. However, the association between these 2 variables has received only minimal attention. We sought to evaluate fetal NRBCs in prolonged pregnancies. METHODS: Umbilical cord blood was prospectively collected at delivery from 75 prolonged (at or beyond 287 days) pregnancies. One hundred and fifty term deliveries (260-286 days) served as controls. All pregnancies were accurately dated with the use of first-trimester sonography. Fetal biophysical profile testing was initiated at 40 weeks of gestation. Patients were delivered if they were in spontaneous labor or the biophysical profile was nonreassuring or by 42 weeks of gestation. Nucleated red blood cell counts were expressed per 100 white blood cells (WBC). Umbilical artery pH studies, as well as other demographic and clinical variables, were obtained. RESULTS: Prolonged pregnancy was associated with a significantly increased incidence of induction of labor and a greater birth weight. There were no other differences between the study group and controls. The median NRBCs per 100 WBCs in prolonged pregnancy was not significantly elevated over the term values (median 3, range 0-35 versus median 3, range 0-34, respectively; P =.25). Neonatal outcome was also comparable between groups. The univariate regression analysis demonstrated a significant association between elevated NRBC counts and low arterial cord blood pH (P <.008, R = 0.175), elevated base excess (P =.02, R = 0.149), low platelet counts (P =.046, R = 0.134), and male gender (P =.028). Stepwise regression analysis revealed that low arterial cord blood pH and male gender were the only independent variables predicting elevated NRBC counts at birth. CONCLUSION: The findings of this study suggest that elevated NRBC counts are associated with specific pregnancy complications rather than uncomplicated prolonged pregnancies in general. PMID- 15292015 TI - The flaming funis. AB - The authors observed a Nicaraguan traditional birth attendant burn the fetal end of the umbilical cord with camphor. They review this practice and reflect on the role of foreign medical volunteers in the developing world. There is a long history to the use of camphor in rituals and medicine. No print references to burning the umbilical cord with camphor, its effectiveness, or its safety could be identified. Interviews with Nicaraguan traditional birth attendants revealed that the practice is passed from generation to generation and that it is believed to decrease infections through the medicinal properties of camphor as well as the flame it produces. It is continued in modern times because it is easy and inexpensive and because there are no clearly better and sustainable alternatives available. Gradual and culturally sensitive modernization to improve the health for mothers and babies is appropriate, but it will be a slow process. Health care volunteers in the developing world struggle with doing the best they can despite the limited resources and sometimes the local traditions. Volunteering as a medical worker in the developing world provides inspiring rewards, teaches powerful lessons, and exposes challenging conflicts. PMID- 15292016 TI - Testing for von Willebrand disease in women with menorrhagia: a systematic review. AB - OBJECTIVE: To review the evidence supporting screening of adult women with menorrhagia for von Willebrand disease. DATA SOURCES: MEDLINE search from January 1,1990, to December 31, 2003, for articles in English, using keywords "menorrhagia," "von Willebrand disease," "diagnosis," and "screening," with a hand-search of bibliographies of identified articles, review of published abstracts, and discussion with experts. METHODS OF STUDY SELECTION: One hundred seven articles meeting search criteria were reviewed. Articles included in the study were those that provided primary data on the prevalence of von Willebrand disease in adult women with menorrhagia, quality of life, surgical complications, and the effectiveness of medical therapy in women with menorrhagia and von Willebrand disease and test characteristics of screening tests for von Willebrand disease. TABULATION, INTEGRATION, AND RESULTS: The reported prevalence of von Willebrand disease in women with menorrhagia ranged from 5-20% in 5 published studies. Comparison of results was limited by small sample sizes and large confidence intervals, as well as differences in the definitions of menorrhagia and von Willebrand disease used in the studies. Although menorrhagia in women with known von Willebrand disease has a substantial impact on quality of life, there are no data suggesting that this impact is substantially greater than that of menorrhagia in women without von Willebrand disease. Data on the risk of surgical bleeding in women with von Willebrand disease are limited, with only 3 studies with a total of 29 patients identified. Data on the effectiveness of specific therapies are also limited; only one controlled trial was identified. Of single tests for screening, one study of the ristocetin cofactor assay had a sensitivity of 79% and specificity of 90%. Studies of a test of platelet adhesion and aggregation resulted in pooled sensitivities of 83-94% and specificities of 80-88%; however, significant heterogeneity was present. CONCLUSION: There are inadequate data to justify routine testing for von Willebrand disease in adult women with menorrhagia outside of the research setting. PMID- 15292017 TI - Reflections on being a spouse in academic medicine. PMID- 15292018 TI - Uterine myomas: an overview of development, clinical features, and management. AB - OBJECTIVE: To review the biology and the pathophysiology of uterine myomas, focus on options for management, and emphasize principles that will render the decision making process as logical as possible. DESIGN: Literature review and synthesis of the authors' experience and philosophy. RESULTS: Uterine myomas are the most common solid pelvic tumors in women. There is increasing evidence that they have a genetic basis and that their growth is related to genetic predisposition, hormonal influences, and various growth factors. Treatment choices are wide and include pharmacologic, surgical, and radiographically directed intervention. Most myomas can be followed serially with surveillance for development of symptoms or progressive growth. CONCLUSION: The past century has witnessed development of highly sophisticated diagnostic and therapeutic technology for myomas. The tools currently at our disposal permit greater management flexibility with safe options, which must be tailored to the individual clinical situation. PMID- 15292020 TI - Perinatal outcomes in singletons following in vitro fertilization: a meta analysis. PMID- 15292021 TI - Very-early-onset discordant growth in monochorionic twin pregnancy. PMID- 15292026 TI - ACOG Committee Opinion. Number 297, August 2004. Nonmedical use of obstetric ultrasonography. AB - The American College of Obstetricians and Gynecologists (ACOG) has endorsed the "Prudent Use" statement from the American Institute of Ultrasound in Medicine (AIUM) discouraging the use of obstetric ultrasonography for nonmedical purposes (eg, solely to create keepsake photographs or videos). The ACOG Committee on Ethics provides reasons in addition to those offered by AIUM for discouraging this practice. PMID- 15292027 TI - ACOG committee opinion. Number 298, August 2004. Prenatal and preconceptional carrier screening for genetic diseases in individuals of Eastern European Jewish descent. AB - Certain autosomal recessive disease conditions are more prevalent in individuals of Eastern European Jewish (Ashkenazi) descent. Previously, the American College of Obstetricians and Gynecologists recommended that individuals of Eastern European Jewish ancestry be offered carrier screening for Tay-Sachs disease, Canavan disease, and cystic fibrosis as part of routine obstetric care. Based on the criteria used to justify offering carrier screening for Tay-Sachs disease, Canavan disease, and cystic fibrosis, the American College of Obstetricians and Gynecologists' Committee on Genetics recommends that couples of Ashkenazi Jewish ancestry also should be offered carrier screening for familial dysautonomia. Individuals of Ashkenazi Jewish descent may inquire about the availability of carrier screening for other disorders. Carrier screening is available for mucolipidosis IV Niemann-Pick disease type A, Fanconi anemia group C, Bloom syndrome, and Gaucher's disease. PMID- 15292028 TI - Endurance training increases LKB1 and MO25 protein but not AMP-activated protein kinase kinase activity in skeletal muscle. AB - LKB1 complexed with MO25 and STRAD has been identified as an AMP-activated protein kinase kinase (AMPKK). We measured relative LKB1 protein abundance and AMPKK activity in liver (LV), heart (HT), soleus (SO), red quadriceps (RQ), and white quadriceps (WQ) from sedentary and endurance-trained rats. We examined trained RQ for altered levels of MO25 protein and LKB1, STRAD, and MO25 mRNA. LKB1 protein levels normalized to HT (1 +/- 0.03) were LV (0.50 +/- 0.03), SO (0.28 +/- 0.02), RQ (0.32 +/- 0.01), and WQ (0.12 +/- 0.03). AMPKK activities in nanomoles per gram per minute were HT (79 +/- 6), LV (220 +/- 9), SO (22 +/- 2), RQ (29 +/- 2), and WQ (42 +/- 4). Training increased LKB1 protein in SO, RQ, and WQ (P < 0.05). LKB1 protein levels after training (%controls) were SO (158 +/- 17), RQ (316 +/- 17), WQ (191 +/- 27), HT (106 +/- 2), and LV (104 +/- 7). MO25 protein after training (%controls) was 595 +/- 71. Training did not affect AMPKK activity. MO25 but not LKB1 or STRAD mRNA increased with training (P < 0.05). Trained values (%controls) were MO25 (164 +/- 22), LKB1 (120 +/- 16), and STRAD (112 +/- 17). LKB1 protein content strongly correlated (r = 0.93) with citrate synthase activity in skeletal muscle (P < 0.05). In conclusion, endurance training markedly increased skeletal muscle LKB1 and MO25 protein without increasing AMPKK activity. LKB1 may be playing multiple roles in skeletal muscle adaptation to endurance training. PMID- 15292029 TI - Diurnal variations in the responsiveness of cardiac and skeletal muscle to fatty acids. AB - Cardiac and skeletal muscle both respond to elevated fatty acid availability by increasing fatty acid oxidation, an effect mediated in large part by peroxisome proliferator-activated receptor-alpha (PPAR alpha). We hypothesized that cardiac and skeletal muscle alter their responsiveness to fatty acids over the course of the day, allowing optimal adaptation when availability of this substrate increases. In the current study, pyruvate dehydrogenase kinase 4 (pdk4) was utilized as a representative PPAR alpha-regulated gene. Opposing diurnal variations in pdk4 expression were observed in cardiac and skeletal muscle isolated from the ad libitum-fed rat; pdk4 expression peaked in the middle of the dark and light phases, respectively. Elevation of circulating fatty acid levels by high-fat feeding, fasting, and streptozotocin-induced diabetes increased pdk4 expression in both heart and soleus muscle. Highest levels of induction were observed during the dark phase, regardless of muscle type or intervention. Specific activation of PPAR alpha with WY-14643 rapidly induced pdk4 expression in heart and soleus muscle. Highest levels of induction were again observed during the dark phase. The same pattern of induction was observed for the PPAR alpha-regulated genes malonyl-CoA decarboxylase and uncoupling protein 3. Investigation into the potential mechanism(s) for these observations exposed a coordinated upregulation of transcriptional activators of the PPAR alpha system during the night, with a concomitant downregulation of transcriptional repressors in both muscle types. In conclusion, responsiveness of cardiac and skeletal muscle to fatty acids exhibits a marked diurnal variation. These observations have important physiological and pathophysiological implications, ranging from experimental design to pharmacological treatment of patients. PMID- 15292030 TI - Evidence for mitochondrial thioesterase 1 as a peroxisome proliferator-activated receptor-alpha-regulated gene in cardiac and skeletal muscle. AB - The physiological role of mitochondrial thioesterase 1 (MTE1) is unknown. It was proposed that MTE1 promotes fatty acid (FA) oxidation (FAO) by acting in concert with uncoupling protein (UCP)3. We previously showed that ucp3 is a peroxisome proliferator-activated receptor-alpha (PPAR alpha)-regulated gene, allowing induction when FA availability increases. On the assumption that UCP3 and MTE1 act in partnership to increase FAO, we hypothesized that mte1 is also a PPAR alpha-regulated gene in cardiac and skeletal muscle. Using real-time RT-PCR, we characterized mte1 gene expression in rat heart and soleus muscles. Messenger RNA encoding for mte1 was 3.2-fold higher in heart than in soleus muscle. Cardiac mte1 mRNA exhibited modest diurnal variation, with 1.4-fold higher levels during dark phase. In contrast, skeletal muscle mte1 mRNA remained relatively constant over the course of the day. High-fat feeding, fasting, and streptozotocin-induced diabetes, interventions that increase FA availability, muscle PPAR alpha activity, and muscle FAO rates, increased mte1 mRNA in heart and soleus muscle. Conversely, pressure overload and hypoxia, interventions that decrease cardiac PPAR alpha activity and FAO rates, repressed cardiac mte1 expression. Specific activation of PPAR alpha in vivo through WY-14643 administration rapidly induced mte1 mRNA in cardiac and skeletal muscle. WY-14643 also induced mte1 mRNA in isolated adult rat cardiomyocytes dose dependently. Expression of mte1 was markedly lower in hearts and soleus muscles isolated from PPAR alpha-null mice. Alterations in cardiac and skeletal muscle ucp3 expression mirrored that of mte1 in all models investigated. In conclusion, mte1, like ucp3, is a PPAR alpha regulated gene in cardiac and skeletal muscle. PMID- 15292031 TI - Attenuation of vasopressin-induced antidiuresis in poorly controlled type 2 diabetes. AB - Renal resistance to vasopressin has been demonstrated in type 1 diabetes and in type 2 diabetes with nephropathy. However, renal response to vasopressin in type 2 diabetes without nephropathy has not been studied. We studied 10 subjects with poorly controlled type 2 diabetes (PCDS; Hb A(1c) >9%), 10 subjects with well controlled type 2 diabetes (WCDS; Hb A(1c) <7%), and 10 matched nondiabetic control subjects (NDCS) during a euglycemic 8-h water deprivation test. None of the subjects had nephropathy. Water deprivation caused similar rises in plasma vasopressin concentrations in all three groups, but the rise in urine osmolality in PCDS (280.3 +/- 49.7 to 594.4 +/- 88.5 mosmol/kgH(2)O) was lower than in WCDS (360.7 +/- 142.8 to 794.1 +/- 77.3 mosmol/kgH(2)O, P < 0.001) or NDCS (336.0 +/- 123.3 to 786.5 +/- 63.3 mosmol/kgH(2)O, P = 0.019). Total urine output was higher in the PCDS than in WCDS and NDCS (P < 0.05). Linear regression analysis showed that, in PCDS, the osmotic thresholds for thirst (291.9 +/- 4.6 mosmol/kgH(2)O) and vasopressin release (291.1 +/- 2.9 mosmol/kgH(2)O) were higher compared with WCDS (286.6 +/- 1.8 and 286.0 +/- 3.6 mosmol/kgH(2)O, respectively) and NDCS (286.0 +/- 2.4 and 284.1 +/- 4.7 mosmol/kgH(2)O, respectively) (between groups P < 0.001 for both variables). Under conditions of euglycemia, PCDS have impaired renal response to vasopressin and elevated osmotic threshold for thirst and vasopressin release in response to dehydration. Under conditions of chronic hyperglycemia, these abnormalities may significantly contribute to the development of dehydration in PCDS. PMID- 15292032 TI - Short-term intermittent exposure to diazoxide improves functional performance of beta-cells in a high-glucose environment. AB - Prolonged periods of "beta-cell rest" exert beneficial effects on insulin secretion from pancreatic islets subjected to a high-glucose environment. Here, we tested for effects of short-term intermittent rest achieved by diazoxide. Rat islets were cultured for 48 h with 27 mmol/l glucose alone, with diazoxide present for 2 h every 12 h or with continuous 48-h presence of diazoxide. Both protocols with diazoxide enhanced the postculture insulin response to 27 mmol/l glucose, to 200 mumol/l tolbutamide, and to 20 mmol/l KCl. Intermittent diazoxide did not affect islet insulin content and enhanced only K(ATP)-dependent secretion, whereas continuous diazoxide increased islet insulin contents and enhanced both K(ATP)-dependent and -independent secretory effects of glucose. Intermittent and continuous diazoxide alike increased postculture ATP-to-ADP ratios, failed to affect [(14)C]glucose oxidation, but decreased oxidation of [(14)C]oleate. Neither of the two protocols affected gene expression of the ion channel-associated proteins Kir6.2, sulfonylurea receptor 1, voltage-dependent calcium channel-alpha1, or Kv2.1. Continuous, but not intermittent, diazoxide decreased significantly mRNA for uncoupling protein-2. A 2-h exposure to 20 mmol/l KCl or 10 mumol/l cycloheximide abrogated the postculture effects of intermittent, but not of continuous, diazoxide. Intermittent diazoxide decreased islet levels of the SNARE protein SNAP-25, and KCl antagonized this effect. Thus short-term intermittent diazoxide treatment has beneficial functional effects that encompass some but not all characteristics of continuous diazoxide treatment. The results support the soundness of intermittent beta-cell rest as a treatment strategy in type 2 diabetes. PMID- 15292033 TI - Exposure to pressure stimulus enhances succinate dehydrogenase activity in L6 myoblasts. AB - Contraction of skeletal muscle generates pressure stimuli to intramuscular tissues. However, the effects of pressure stimuli, other than those created by electricity or nerve impulse, on physiological and biochemical responses in skeletal muscles are unknown. The purpose of this study is to examine the effects of a pure pressure stimulus on metabolic responses in a skeletal muscle cell line. Atmospheric pressure was applied to L6 myoblasts using an original apparatus. Succinate dehydrogenase (SDH) activity was evaluated by colorimetric assay using tetrazolium monosodium salt. The amounts of 2-deoxy-[(3)H]glucose uptake and lactate release were measured. SDH activity was 2.6- to 2.9-fold higher in pressurized L6 cells than in nonpressurized L6 cells (P < 0.01), and 2 deoxy-[(3)H]glucose uptake was 2.2-fold higher (P < 0.001). In addition, the amount of released lactate decreased from 6.8 to 3.7 mumol/dish when pressure was applied (P < 0.001). In contrast, the intracellular lactate contents of the pressurized cells were higher than those of nonpressurized cells (P < 0.01). However, the total amount of released lactate and intracellular lactate was lower in the pressurized cells than in nonpressurized cells. These findings demonstrate that a pure pressure stimulus enhances aerobic metabolism in L6 skeletal muscle cells and raise the possibility that elevated intramuscular pressure during muscle activity may be an important factor in stimulating oxidative metabolic responses in skeletal muscles. PMID- 15292034 TI - The ageing of the population: implications for multidisciplinary care in hospital. AB - BACKGROUND: Comprehensive geriatric assessment and multidisciplinary intervention are of proven benefit in the care of older people. OBJECTIVE: To determine whether patients' multidisciplinary needs in hospital can be met by current service provision. DESIGN: A comprehensive census assessing the multidisciplinary needs of an entire inpatient population compared to available multidisciplinary therapy time. SETTING: A large teaching hospital Trust, comprising six hospital sites. METHODS: On census day, the age, Barthel Index score and multidisciplinary needs of all adult inpatients were documented. Each therapist completed a questionnaire regarding their direct patient contact time on census day. RESULTS: 889 of 1,324 eligible patients (69%) had multidisciplinary needs on census day. These patients were scattered throughout all 46 acute wards, 14 rehabilitation and 4 continuing care settings. Mean age was 65.3 years in acute wards, 73.5 in rehabilitation wards and 80.8 in continuing care. Age correlated inversely with Barthel Index score (r -0.255, P <0.01). The percentage of patients with multidisciplinary need increased with increasing age. The calculated number of minutes of therapy time per day available to each patient varied between therapies and across sites. Mean physiotherapy time available per patient needing physiotherapy on census day ranged from 17 minutes 41 seconds in acute wards to 26 minutes 24 seconds in rehabilitation wards. CONCLUSIONS: A high proportion of inpatients, particularly older patients, across all care settings have multidisciplinary needs. This needs to be expressly considered in the planning of future health services if multidisciplinary needs of older people in hospital are to be met. PMID- 15292035 TI - The effects of age, glucose ingestion and gluco-regulatory control on episodic memory. AB - BACKGROUND: Previous research has been inconclusive regarding the impact of glucose ingestion and gluco-regulatory control on cognitive performance in healthy older adults. The aim of this research was to determine whether glucose specifically enhanced episodic memory in an older population. In addition, the link between individual differences in glucose regulation and the magnitude of the enhancement effect was examined. DESIGN AND SUBJECTS: A within subjects, counterbalanced, crossover design was used with 20 participants (60-80 year olds), each serving as his/her control. METHODS: Episodic memory was tested by presenting unrelated paired associates followed by immediate and delayed cued recall, and delayed recognition, under single and dual task conditions. In addition, a battery of cognitive tests was administered, including tests of semantic memory, working memory and speed of processing. RESULTS: Glucose ingestion was found to largely facilitate performance of episodic memory. Furthermore, subsidiary analyses found that gluco-regulatory efficiency predicted episodic memory performance in both control and glucose conditions. CONCLUSIONS: A boost in performance after glucose ingestion was particularly seen in the episodic memory domain. Notably, strong evidence was provided for the utility of gluco-regulatory control measures as indicators of cognitive decline in the elderly. PMID- 15292036 TI - Is tube feeding associated with altered arterial oxygen saturation in stroke patients? AB - BACKGROUND: Reduced arterial oxygen saturation (SaO(2)) during swallowing, oral feeding and feeding tube placement has been demonstrated in stroke patients. It is not known if tube feeding causes similar episodes of arterial desaturation and whether there is a case for routine pulse oximetry during tube feeding. OBJECTIVE: To determine if tube feeding in stroke patients is associated with hypoxia. METHODS: We compared ischaemic or haemorrhagic stroke patients who were NG or PEG fed with a control group of age matched non-dysphagic stroke patients who were orally fed. We excluded people already on supplemental oxygen. Pulse oximetry was performed before, during a meal (for 20 min) and for 10 min after and changes from baseline readings determined. RESULTS: Data were collected for 20 controls and 18 tube-fed patients. Mean age was 75 years and median time to assessment 14.5 days. The two groups were reasonably matched for age, sex, type of stroke and time to assessment, but differed significantly in the Oxfordshire Community Stroke Project (OCSP) classification and Rankin score. The mean baseline SaO(2) of controls was 96.5% (SD 1.47) and that of the tube-fed group 96.0% (SD 1.46). Reduction in SaO(2) from baseline during and after feeding ranged from 0.35% to 0.78% with no statistically or clinically significant differences between the two groups. CONCLUSIONS: No clinically significant reduction in SaO(2) was found in our tube-fed patients as compared to controls. Our study suggests that routine pulse oximetry during tube feeding is not necessary. PMID- 15292037 TI - Urinary 1-hydroxypyrene and polycyclic aromatic hydrocarbon exposure among asphalt paving workers. AB - OBJECTIVES: Using urinary 1-hydroxypyrene (1-OHP) as a measure of total absorbed dose, the primary objective of this study was to evaluate the total effect of inhalation and dermal PAH exposures while considering other factors such as age, body mass index and smoking that may also have a significant effect on urinary 1 OHP. METHODS: The study population included two groups of highway construction workers: 20 paving workers and 6 milling workers. During multiple consecutive workshifts, personal air and dermal samples were collected from each worker and analyzed for pyrene. During the same work week, urine samples were collected pre shift, post-shift and at bedtime each day and analyzed for 1-OHP. Distributed lag models were used to evaluate the independent effect of inhalation and dermal exposures that occurred at each of several preceding exposure periods and were used to identify the relevant period of influence for each pathway. RESULTS: The paving workers had inhalation (mean 0.3 micro g/m(3)) and dermal (5.7 ng/cm(2)) exposures to pyrene that were significantly higher than the milling workers. At pre-shift on Monday morning, following a weekend away from work, the pavers and millers had the same mean baseline urinary 1-OHP level of 0.4 micro g/g creatinine. The mean urinary 1-OHP levels among pavers increased significantly from pre-shift to post-shift during each work day, while the mean urinary 1-OHP levels among millers varied little and remained near the baseline level throughout the study period. Among pavers there was a clear increase in the pre shift data during the work week, such that the average pre-shift level on day 4 (1.4 micro g/g creatinine) was 3.5 times higher than the average pre-shift results on day 1 (0.4 micro g/g creatinine). The results of the distributed lag model indicated that the impact of dermal exposure was approximately eight times the impact of inhalation exposure. Furthermore, dermal exposure that occurred during the preceding 32 h had a statistically significant effect on urinary 1 OHP, while the effect of inhalation exposure was not significant. CONCLUSIONS: We found that distributed lag models are a valuable tool for analyzing longitudinal biomarker data and our results indicate that dermal contact is the primary route of exposure to PAHs among asphalt paving workers. An exposure assessment of PAHs that does not consider dermal exposure may considerably underestimate cumulative exposure and control strategies aimed at reducing occupational exposure to asphalt-related PAHs should include an effort to reduce dermal exposure. PMID- 15292038 TI - Determination of triglycidyl isocyanurate from powder coatings in occupational hygiene samples by gas chromatography with mass spectrometric detection. AB - A method for the determination of triglycidyl isocyanurate (TGIC) from powder coatings using gas chromatography with mass spectrometric (GC/MS) detection is presented. The new method is considerably easier to use than the existing methods and has superior performance. It has been novelly applied to the determination of TGIC in a variety of occupational hygiene samples (cotton swabs, gloves and whole suits). Using the GC/MS method, the following percentage recoveries were found: gloves 114 +/- 1.9; swabs 73 +/- 6.5; whole suit 125 +/- 16.3 and 108 +/- 6.8 (two powder coatings); filter 79 +/- 8. The estimated limit of detection was 0.002 micro g/ml. PMID- 15292039 TI - Evaluating exposures to complex mixtures of chemicals during a new production process in the plastics industry. AB - The goal of this study was to monitor emission of chemicals at a factory where plastics products were fabricated by a new robotic (impregnated tape winding) production process. Stationary and personal air measurements were taken to determine which chemicals were released and at what concentrations. Principal component analyses (PCA) and linear regression were used to determine the emission sources of different chemicals found in the air samples. We showed that complex mixtures of chemicals were released, but most concentrations were below Dutch exposure limits. Based on the results of the principal component analyses, the chemicals found were divided into three groups. The first group consisted of short chain aliphatic hydrocarbons (C2-C6). The second group included larger hydrocarbons (C9-C11) and some cyclic hydrocarbons. The third group contained all aromatic and two aliphatic hydrocarbons. Regression analyses showed that emission of the first group of chemicals was associated with cleaning activities and the use of epoxy resins. The second and third group showed strong association with the type of tape used in the new tape winding process. High levels of CO and HCN (above exposure limits) were measured on one occasion when a different brand of impregnated polypropylene sulphide tape was used in the tape winding process. Plans exist to drastically increase production with the new tape winding process. This will cause exposure levels to rise and therefore further control measures should be installed to reduce release of these chemicals. PMID- 15292040 TI - Genome size variation in Central European species of Cirsium (Compositae) and their natural hybrids. AB - BACKGROUND AND AIMS: Nuclear DNA amounts of 12 diploid and one tetraploid taxa and 12 natural interspecific hybrids of Cirsium from 102 populations in the Czech Republic, Austria, Slovakia and Hungary were estimated. METHODS: DAPI and PI flow cytometry were used. KEY RESULTS: 2C-values of diploid (2n = 34) species varied from 2.14 pg in C. heterophyllum to 3.60 pg in C. eriophorum (1.68-fold difference); the 2C value for the tetraploid C. vulgare was estimated at 5.54 pg. The DNA contents of hybrids were located between the values of their putative parents, although usually closer to the species with the smaller genome. Biennial species of Cirsium possessed larger nuclear DNA amounts than their perennial relatives. Genome size was negatively correlated with Ellenberg's indicator values for continentality and moisture and with eastern limits of distribution. A negative relationship was also detected between the genome size and the tendency to form natural interspecific hybrids. On the contrary, C-values positively corresponded with the spinyness (degree of spinosity). AT frequency ranged from 48.38 % in C. eriophorum to 51.75 % in C. arvense. Significant intraspecific DNA content variation in DAPI sessions was detected in C. acaule (probably due to the presence of B-chromosomes), and in tetraploid C. vulgare. Only the diploid level was confirmed for the Pannonian C. brachycephalum, generally considered to be tetraploid. In addition, triploidy was discovered for the first time in C. rivulare. CONCLUSIONS: Considerable differences in nuclear DNA content exist among Central European species of Cirsium on the diploid level. Perennial soft spiny Cirsium species of wet habitats and continental distributions generally have smaller genomes. The hybrids of diploid species remain diploid, and their DNA content is smaller than the mean of the parents. Species with smaller genomes produce interspecific hybrids more frequently. PMID- 15292041 TI - The use of a principal axis model to examine individual plant harvest index in four grain legumes. AB - BACKGROUND AND AIMS: A principal axis model (PAM) has been proposed to enable the selection of crop ideotypes. The PAM enables plant-to-plant variability within crops to be quantified and compared. The aim of this paper is to validate the PAM for four grain legumes. METHODS: Four grain legumes (Cicer arietinum, Lens culinaris, Lupinus angustifolius, Pisum sativum) were used to quantify the influence of plant-to-plant variability on crop yields. To create variability, populations of 10, 100 and 400 plants m(-2) were established 'on-the-square' with sowing depths of 2, 5 and 10 cm. Further, a central plant was treated with nitrogen and the impact of this on its four neighbouring plants was examined. Seeds were sown and plants harvested individually by hand. KEY RESULTS: Mean individual plant seed weight (SWT) and plant weight (PWT) decreased as plant population increased but there was a consistent and strong (R2 > 0.90) linear relationship between SWT and PWT, with a negative SWT-axis intercept in all species. These components form the basis of the principal axis model (PAM). The PAM was used to summarize the performance of individual plants within a crop and quantify the variability caused by N treatment and the lowest and highest yielding individual plants. A negative SWT-axis intercept indicated that a minimum plant weight (MPW) was required for seed production and therefore the relationship between plant harvest index (PHI) and PWT was asymptotic. The heaviest MPW was calculated for plants grown at the lowest plant population and it was species-dependent, being higher in the larger seeded species. CONCLUSIONS: Agronomic or physiological characteristics that lead to variability in PWT within a population will decrease PHI, and crop yield. The PAM may be useful in breeding programmes to identify plant phenotypes that minimize this plant-to-plant variability. PMID- 15292042 TI - Genetic responses to phosphorus deficiency. AB - BACKGROUND: Phosphorus (P) is an essential macronutrient for plants. Plants take up P as phosphate (Pi) from the soil solution. Since little Pi is available in most soils, P fertilizers are applied to crops. However, the use of P fertilizers is unsustainable and may cause pollution. Consequently, there is a need to develop more P-use-efficient (PUE) crops and precise methods to monitor crop P status. SCOPE: Manipulating the expression of genes to improve the PUE of crops could reduce their P fertilizer requirement. This has stimulated research towards the identification of genes and signalling cascades involved in plant responses to P deficiency. Genes that respond to P deficiency can be grouped into 'early' genes that respond rapidly and often non-specifically to P deficiency, or 'late' genes that impact on the morphology, physiology or metabolism of plants upon prolonged P deficiency. SUMMARY: The use of micro-array technology has allowed researchers to catalogue the genetic responses of plants to P deficiency. Genes whose expression is altered by P deficiency include various transcription factors, which are thought to coordinate plant responses to P deficiency, and other genes involved in P acquisition and tissue P economy. Several common cis regulatory elements have been identified in the promoters of these genes, suggesting that their expression might be coordinated. It is suggested that knowledge of the genes whose expression changes in response to P deficiency might allow the development of crops with improved PUE, and could be used in diagnostic techniques to monitor P deficiency in crops either directly using 'smart' indicator plants or indirectly through transcript profiling. The development of crops with improved PUE and the adoption of diagnostic technology could reduce production costs, minimize the use of a non-renewable resource, reduce pollution and enhance biodiversity. PMID- 15292043 TI - Lipoprotein lipase in the kidney: activity varies widely among animal species. AB - Much evidence points to a relationship among kidney disease, lipoprotein metabolism, and the enzyme lipoprotein lipase (LPL), but there is little information on LPL in the kidney. The range of LPL activity in the kidney in five species differed by >500-fold. The highest activity was in mink, followed by mice, Chinese hamsters, and rats, whereas the activity was low in guinea pigs. In contrast, the ranges for LPL activities in heart and adipose tissue were less than six- and fourfold, respectively. The activity in the kidney (in mice) decreased by >50% on food deprivation for 6 h without corresponding changes in mRNA or mass. This decrease in LPL activity did not occur when transcription was blocked with actinomycin D. Immunostaining for kidney LPL in mice and mink indicated that the enzyme is produced in tubular epithelial cells. To explore the previously suggested possibility that the negatively charged glomerular filter picks up LPL from the blood, bovine LPL was injected into rats and mice. This resulted in decoration of the glomerular capillary network with LPL. This study shows that in some species LPL is produced in the kidney and is subject to nutritional regulation by a posttranscriptional mechanism. In addition, LPL can be picked up from blood in the glomerulus. PMID- 15292044 TI - Alterations in cell-adhesive and migratory properties of proximal tubule and collecting duct cells from bcl-2 -/- mice. AB - Bcl-2 protects cells from apoptosis initiated by a variety of stimuli including loss of cell adhesion. Bcl-2 -/- mice develop renal hypoplastic/cystic dysplasia with renal cyst formation coinciding with renal maturation in normal mice. To gain a better understanding of the role cell-adhesive mechanisms play during renal maturation, we generated proximal tubule and collecting duct cell lines from postnatal day 10 (P10) and P20 bcl-2 +/+ and bcl-2 -/- mice. Very little is known about the role cell-adhesive and migratory mechanisms play during renal maturation. We observed that modulation of cell-adhesive properties, which normally occur in a nephron segment-specific manner during renal maturation, and cell migration were altered in cells from bcl-2 -/- mice. Enhanced migration of bcl-2 -/- proximal tubule cells in a scratch wound assay was completely inhibited by incubation with PP1 (Src inhibitor) and moderately affected by incubation with SB-203580 (p38 inhibitor). These cells expressed increased levels of fibronectin and had numerous central focal adhesions. P20 bcl-2 -/- proximal tubule cells adhered to fibronectin but adhered poorly to collagen, vitronectin, or laminin. Collecting duct cells, similar to proximal tubule cells from bcl-2 -/- mice, demonstrated enhanced migration in a scratch wound assay that was inhibited by incubation with PP1. Migration of these cells was moderately affected by incubation with PD-98059 (MEK inhibitor) or LY-294002 (PI3 kinase inhibitor), whereas incubation with SB-203580 had no effect. P10 bcl-2 -/- collecting duct cells also expressed increased levels of fibronectin but decreased levels of thrombospondin-1 and demonstrated precocious binding to fibronectin and vitronectin compared with bcl-2 +/+ cells. The ability of P20 bcl-2 +/+ collecting duct cells to adhere to fibronectin and vitronectin corresponded with a decline in thrombospondin-1 expression. Therefore, alterations in cell-adhesive and migratory characteristics may be an early indicator of aberrant renal epithelial cell differentiation. PMID- 15292045 TI - Involvement of cytosolic Cl- in osmoregulation of alpha-ENaC gene expression. AB - Hypotonicity stimulates transepithelial Na(+) reabsorption in renal A6 cells, but the mechanism for this stimulation is not fully understood. In the present study, we found that hypotonicity stimulated Na(+) reabsorption through increases in mRNA expression of the alpha-subunit of the epithelial Na(+) channel (alpha ENaC). Hypotonicity decreases cytosolic Cl(-) concentration; therefore, we hypothesized that hypotonicity-induced decreases in cytosolic Cl(-) concentration could act as a signal to regulate Na(+) reabsorption through changes in alpha ENaC mRNA expression. Treatment with the flavone apigenin, which activates the Na(+)-K(+)-2Cl(-) cotransporter and increases cytosolic Cl(-) concentration, markedly suppressed the hypotonicity-induced increase in alpha-ENaC mRNA expression. On the other hand, blockade of the Na(+)-K(+)-2Cl(-) cotransporter decreases cytosolic Cl(-) concentration and increased alpha-ENaC mRNA expression and Na(+) reabsorption. Blocking Cl(-) channels with 5-nitro-2-(3 phenylpropylamino)-benzoic acid (NPPB) inhibited the hypotonicity-induced decrease in cytosolic Cl(-) concentration and suppressed the hypotonicity-induced increase in alpha-ENaC mRNA expression. Coapplication of NPPB and apigenin synergistically suppressed alpha-ENaC mRNA expression. Thus, in every case, changes in cytosolic Cl(-) concentration were associated with changes in alpha ENaC mRNA expression and changes in Na(+) reabsorption: decreases in cytosolic Cl(-) concentration increased alpha-ENaC mRNA and increased Na(+) reabsorption, whereas increases in cytosolic Cl(-) concentration decreased alpha-ENaC mRNA and decreased Na(+) reabsorption. These findings support the hypothesis that changes in cytosolic Cl(-) concentration are an important and novel signal in hypotonicity-induced regulation of alpha-ENaC expression and Na(+) reabsorption. PMID- 15292046 TI - Protection of transplant-induced renal ischemia-reperfusion injury with carbon monoxide. AB - Carbon monoxide (CO), a product of heme metabolism by heme oxygenases, is known to impart protection against oxidative stress. We hypothesized that CO would protect ischemia-reperfusion (I/R) injury of transplanted organs, and the efficacy of CO was studied in the rat kidney transplantation model. A Lewis rat kidney graft, preserved in University of Wisconsin solution at 4 degrees C for 24 h, was orthotopically transplanted into syngeneic rats. Recipients were maintained in room air or exposed to CO (250 ppm) in air for 1 h before and 24 h after transplantation. Animals were killed 1, 3, 6, and 24 h after transplantation to assess efficacy of inhaled CO. Rapid upregulation of mRNA for IL-6, IL-1beta, TNF-alpha, ICAM-1, heme oxygenase-1, and inducible nitric oxide synthase was observed within 3 h after transplantation in the control grafts of air-exposed recipients, associating with histopathological evidences of acute tubular necrosis, interstitial hemorrhage, and edema. In contrast, the increase of inflammatory mediators was markedly inhibited in kidney grafts of CO-treated recipients, which correlated with improved renal cortical blood flow. Further detailed morphological analyses revealed that CO preserved the glomerular vascular architecture and podocyte viability with less apoptosis of tubular epithelial cells and less ED1(+) macrophage infiltration. CO inhalation resulted in improved serum creatinine levels and clearance, and animal survival was significantly improved with CO to 60.5 from 25 days in untreated controls. The study demonstrates that exposure of kidney graft recipients to CO at a low concentration can impart significant protective effects against renal I/R injury and improve function of renal grafts. PMID- 15292047 TI - Transepithelial HCO3- absorption is defective in renal thick ascending limbs from Na+/H+ exchanger NHE1 null mutant mice. AB - In the medullary thick ascending limb (MTAL) of rat kidney, inhibiting basolateral Na(+)/H(+) exchange with either amiloride or nerve growth factor (NGF) results secondarily in inhibition of apical Na(+)/H(+) exchange, thereby decreasing transepithelial HCO(3)(-) absorption. To assess the possible role of the Na(+)/H(+) exchanger NHE1 in this regulatory process, MTALs from wild-type and NHE1 knockout (NHE1(-/-)) mice were studied using in vitro microperfusion. The rate of HCO(3)(-) absorption was decreased 60% in NHE1(-/-) MTALs (15.4 +/- 0.5 pmol.min(-1).mm(-1) wild-type vs. 6.0 +/- 0.5 pmol.min(-1).mm(-1) NHE1(-/-)). Transepithelial voltage, an index of the NaCl absorption rate, did not differ in wild-type and NHE1(-/-) MTALs. Basolateral addition of 10 microM amiloride or 0.7 nM NGF decreased HCO(3)(-) absorption by 45-49% in wild-type MTALs but had no effect on HCO(3)(-) absorption in NHE1(-/-) MTALs. Inhibition of HCO(3)(-) absorption by vasopressin and stimulation by hyposmolality, both of which regulate MTAL HCO(3)(-) absorption through primary effects on apical Na(+)/H(+) exchange, were similar in wild-type and NHE1(-/-) MTALs. Thus the regulatory defect in NHE1(-/-) MTALs is specific for factors (bath amiloride and NGF) shown previously to inhibit HCO(3)(-) absorption through primary effects on basolateral Na(+)/H(+) exchange. These findings demonstrate a novel role for NHE1 in transepithelial HCO(3)(-) absorption in the MTAL, in which basolateral NHE1 controls the activity of apical NHE3. Paradoxically, a reduction in NHE1-mediated H(+) extrusion across the basolateral membrane leads to a decrease in apical Na(+)/H(+) exchange activity that reduces HCO(3)(-) absorption. PMID- 15292048 TI - Effect of endogenous angiotensin II on the frequency response of the renal vasculature. AB - The renal vasculature functions as an efficient low-pass filter of the multiple frequencies contained within renal sympathetic nerve activity. This study examined the effect of angiotensin II on the frequency response of the renal vasculature. Physiological changes in the activity of the endogenous renin angiotensin system were produced by alterations in dietary sodium intake. The frequency response of the renal vasculature was evaluated using pseudorandom binary sequence renal nerve stimulation, and the role of angiotensin II was evaluated by the administration of the angiotensin II AT(1)-receptor antagonist losartan. In low-sodium-diet rats with increased renin-angiotensin system activity, losartan steepened the renal vascular frequency response (i.e., greater attenuation); this was not seen in normal- or high-sodium-diet rats with normal or decreased renin-angiotensin system activity. Analysis of the transfer function from arterial pressure to renal blood flow, i.e., dynamic autoregulation, showed that the tubuloglomerular feedback but not the myogenic component was enhanced in low- and normal- but not in high-sodium-diet rats and that this was reversed by losartan administration. Thus physiological increases in endogenous renin angiotensin activity inhibit the renal vascular frequency response to renal nerve stimulation while selectively enhancing the tubuloglomerular feedback component of dynamic autoregulation of renal blood flow. PMID- 15292049 TI - PDZ binding motif-dependent localization of K+ channel on the basolateral side in distal tubules. AB - Kir5.1, a nonfunctional inwardly rectifying K(+) channel by itself, can form functional channels by assembling with other proteins. We previously showed that Kir5.1 assembled with Kir4.1 and functioned as an acid-base regulator in the kidney. In this study, we examined the intrarenal distribution of Kir5.1 by RT PCR analysis on dissected nephron segments and immunohistochemical analysis with the specific anti-Kir5.1 antibody. Strong expression of Kir5.1 was detected in distal convoluted tubules, and weak expression was also detected in thick ascending limb of Henle's loop. Colocalization of Kir5.1 with Kir4.1 indicated expression of Kir5.1/Kir4.1 heteromer in these nephron segments. In a renal epithelial cell line, Madin-Darby canine kidney cells, heteromer formation with Kir4.1 changed the localization of Kir5.1 from intracellular components to the cell surface. The COOH-terminal cytoplasmic portion that includes the PDZ binding motif of Kir4.1 was responsible for this intracellular localization. These data suggest the signals on the COOH terminus of Kir4.1, including PDZ binding motif, determine the intracellular localization of Kir5.1/Kir4.1 heteromer in distal tubules. PMID- 15292050 TI - The Cl-/HCO3- exchanger pendrin in the rat kidney is regulated in response to chronic alterations in chloride balance. AB - Pendrin (Pds; Slc26A4) is a new anion exchanger that is believed to mediate apical Cl(-)/HCO(3)(-) exchange in type B and non-A-non-B intercalated cells of the connecting tubule and cortical collecting duct. Recently, it has been proposed that this transporter may be involved in NaCl balance and blood pressure regulation in addition to its participation in the regulation of acid-base status. The purpose of our study was to determine the regulation of Pds protein abundance during chronic changes in chloride balance. Rats were subjected to either NaCl, NH(4)Cl, NaHCO(3), KCl, or KHCO(3) loading for 6 days or to a low NaCl diet or chronic furosemide administration. Pds protein abundance was estimated by semiquantitative immunoblotting in renal membrane fractions isolated from the cortex of treated and control rats. We observed a consistent inverse relationship between Pds expression and diet-induced changes in chloride excretion independent of the administered cation. Conversely, NaCl depletion induced by furosemide was associated with increased Pds expression. We conclude that Pds expression is specifically regulated in response to changes in chloride balance. PMID- 15292051 TI - Activation of EP4 receptors contributes to prostaglandin E2-mediated stimulation of renal sensory nerves. AB - Induction of cyclooxygenase-2 (COX-2) in the renal pelvic wall increases prostaglandin E(2) (PGE(2)) leading to stimulation of cAMP production, which results in substance P (SP) release and activation of renal mechanosensory nerves. The subtype of PGE receptors involved, EP2 and/or EP4, was studied by immunohistochemistry and renal pelvic administration of agonists and antagonists of EP2 and EP4 receptors. EP4 receptor-like immunoreactivity (LI) was colocalized with calcitonin gene-related peptide (CGRP)-LI in dorsal root ganglia (DRGs) at Th(9)-L(1) and in nerve terminals in the renal pelvic wall. Th(9)-L(1) DRG neurons also contained EP3 receptor-LI and COX-2-LI, each of which was colocalized with CGRP-LI in some neurons. No renal pelvic nerves contained EP3 receptor-LI and only very few nerves COX-2-LI. The EP1/EP2 receptor antagonist AH 6809 (20 microM) had no effect on SP release produced by PGE(2) (0.14 microM) from an isolated rat renal pelvic wall preparation. However, the EP4 receptor antagonist L-161,982 (10 microM) blocked the SP release produced by the EP2/EP4 receptor agonist butaprost (10 microM) 12 +/- 2 vs. 2 +/- 1 and PGE(2), 9 +/- 1 vs. 1 +/- 0 pg/min. The SP release by butaprost and PGE(2) was similarly blocked by the EP4 receptor antagonist AH-23848 (30 microM). In anesthetized rats, the afferent renal nerve activity (ARNA) responses to butaprost 700 +/- 100 and PGE(2).780 +/- 100%.s (area under the curve of ARNA vs. time) were unaffected by renal pelvic perfusion with AH-6809. However, 1 microM L-161,982 and 10 microM AH 23848 blocked the ARNA responses to butaprost by 94 +/- 5 and 78 +/- 10%, respectively, and to PGE(2) by 74 +/- 16 and 74 +/- 11%, respectively. L-161,982 also blocked the ARNA response to increasing renal pelvic pressure 10 mmHg, 85 +/ 5%. In conclusion, PGE(2) increases renal pelvic release of SP and ARNA by activating EP4 receptors on renal sensory nerve fibers. PMID- 15292052 TI - Cell density-dependent expression of EDG family receptors and mesangial cell proliferation: role in lysophosphatidic acid-mediated cell growth. AB - Lysophosphatidic acid (LPA), a major member of the bioactive lysophospholipids in serum, possesses diverse physiological activities including cell proliferation. Recently, three endothelial differentiation gene (EDG) family receptors, including EDG-2 (LPA1), EDG-4 (LPA2), and EDG-7 (LPA3), have been identified as LPA receptors. The role of LPA and their receptors in mesangial cell physiology is not clearly understood. This study examined the expression profile of EDG receptors as a function of cell density and the participation of EDG receptors in human mesangial cell proliferation by LPA. We showed that mesangial cells express all three EDG family LPA receptors in a cell density-dependent manner. EDG-7 maximally expressed at sparse cell density and minimally expressed in dense cell population. The EDG-2 expression pattern was opposite to the EDG-7. No changes in EDG-4 expression as a function of cell density were noted. DNA synthetic rate was greater in sparse cell density compared with dense cell population and followed a similar pattern with EDG-7 expression. Comparative studies in sparse and dense cell density indicated that EDG-7 was positively associated, whereas EDG-2 was negatively associated with cell proliferation rate. LPA induced mesangial cell proliferation by 1.5- to 3.5-fold. Dioctanoylglycerol pyrophosphate, an antagonist for EDG-7, almost completely inhibited mesangial cell proliferation induced by LPA. We suggest that EDG-7 regulates LPA-mediated mesangial cell proliferation. Additionally, these data suggest that EDG-7 and EDG-2 LPA receptors play a diverse role as proliferative and antiproliferative, respectively, in mesangial cells. Regulation of EDG family receptors may be importantly linked to mesangial cell-proliferative processes. PMID- 15292053 TI - Behavioural disorders induced by external globus pallidus dysfunction in primates: I. Behavioural study. AB - The current model of basal ganglia organization postulates the existence of a functional partitioning into sensorimotor, associative and limbic territories, implicated in motor, cognitive and emotional aspects of behaviour, respectively. This organization was proposed initially on the basis of the cortico-striatal projections and was extended to the various structures of the basal ganglia. While there is a considerable body of experimental evidence in support of an involvement of the basal ganglia sensorimotor territory in basic control of movements, evidence for the functional relevance of the non-motor territories has had to be based on a growing number of clinical observations due to the paucity of relevant animal studies. Previous studies in monkeys have, however, shown that a reversible and focal dysfunction induced by microinjections of bicuculline in the sensorimotor territory of the external globus pallidus (GPe) can generate abnormal movements. We therefore hypothesized that the same approach applied to the associative and limbic territories of the GPe would induce behavioural disorders rather than abnormal movements. To address this hypothesis, we performed microinjections of bicuculline, using the same concentration in each of the sensorimotor, associative and limbic territories of the GPe, as defined by striato-pallidal projections. Spontaneous behaviour and performance of a simple food-retrieving task during the effects of these microinjections were compared with data obtained in control conditions in the same monkeys. We found that bicuculline microinjections induced stereotypy when performed in the limbic part of the GPe, and attention deficit and/or hyperactivity when performed in the associative part. No movement disorders were observed during these behavioural disturbances. As previously described, abnormal movements were observed when bicuculline was injected into the sensorimotor territory of the GPe. The relationship between the localization of microinjection sites and the type of behavioural effect was similar for the three monkeys. Control microinjections of bicuculline into surrounding structures (striatum and internal globus pallidus) and saline injections into the GPe failed to induce any observable effect. These results support the hypotheses of functional diversity and territorial specificity in the GPe, in agreement with the parallel circuits organizational model of the basal ganglia. Furthermore, the behavioural effects shared similar features with symptoms observed in Tourette's syndrome, attention deficit/hyperactivity and compulsive disorders. Thus, our study provides experimental evidence for the involvement of the associative and limbic parts of the basal ganglia in these pathologies. These results may provide the basis for a primate model of these disorders. PMID- 15292054 TI - Behavioural disorders induced by external globus pallidus dysfunction in primates II. Anatomical study. AB - The anatomical organization of the basal ganglia supports their involvement in movement and behavioural disorders. Thus dyskinesia, attention deficit with or without hyperactivity, and stereotyped behaviour can be induced by microinjections of bicuculline, a GABAergic antagonist, into different parts of the external globus pallidus (GPe) in monkeys. The aim of the present study was to determine the anatomo-functional circuits inside the basal ganglia which are specifically related to each of these behavioural changes. For that, axonal tracers were injected in the same pallidal sites where abnormal behaviours have previously been obtained by bicuculline microinjections. The labelling was mapped in the different basal ganglia and matched with the topography of the cortico striato-pallidal projections already reported in the literature and with the distribution of calbindin immunoreactivity. Our results first show that the pallidal sites related to dyskinesia, attention deficit with or without hyperactivity, and stereotyped behaviour, were respectively in motor, associative and limbic territories, defined as weak, moderate and intensive calbindin immunoreactivity. The same relationship was observed between the distribution of the labelling in the different basal ganglia after tracer injections performed in these different pallidal sites and the anatomo-functional territories. Thus regarding the origin of the circuits within the striatum, tracer injections performed in the dyskinesia site labelled neurons located in the posterior sensorimotor putamen, those performed in the hyperactivity and/or attention deficit labelled neurons in the laterodorsal putamen and caudate nucleus, regions corresponding to associative and anterior motor territories, while those performed in the stereotyped behaviour site labelled neurons in the ventral limbic striatum. Regarding the GPe output on the basal ganglia, the different circuits also appeared underlined by different anatomo-functional territories, even if a partial overlap exists. Each of these anatomical circuits systematically involves both the internal globus pallidus (GPi) and the substantia nigra pars reticulata (SNr) but, whereas movement circuit is mainly related to the GPi, stereotyped behaviour is mainly related to the SNr. Additionally, subregions of the subthalamic nucleus were also systematically involved, depending on the movement or behavioural disorder produced. These results demonstrate that distinct circuits involving different anatomo-functional territories of the basal ganglia, with partial overlap, participate in different behavioural disorders in monkeys. It seems likely that these neuronal circuits are involved in pathologies like Tourette's syndrome, attention deficit/hyperactivity disorders and obsessional compulsive troubles. This study provides the basis for further researches with a therapeutical viewpoint. PMID- 15292055 TI - An exploration of the role of the superior temporal gyrus in visual search and spatial perception using TMS. AB - This study sought to investigate the recent claim by H.-O. Karnath and his colleagues that the crucial locus of neurological damage in neglect patients lies in the right superior temporal gyrus (STG), and not in the right posterior parietal cortex (PPC) as conventionally thought. Using transcranial magnetic stimulation (TMS), we first tested the involvement of the right STG in a task commonly used in the diagnosis of neglect, the landmark task. No evidence was found for a critical involvement of the right STG in the processing of this task, though evidence was found for the involvement of the right PPC. In contrast, however, when we examined the effects of TMS on exploratory search, a double dissociation between right STG and right PPC was found. When the processing of conjunction items was required, involvement of the right PPC (and not STG) was found in accordance with our previous research. When a difficult exploratory search through feature items was required, however, the right STG (not PPC) was found to be involved. A hitherto unknown role for right STG in visual search tasks was thus uncovered. These data suggest that conclusions about the area of brain damage resulting in neglect-like symptoms are highly dependent on the task used to diagnose them, with lesions in right PPC leading to deficits on the landmark task and conjunction visual search, and lesions in right STG resulting in deficits in feature based serial exploratory search tasks. PMID- 15292056 TI - Effects of reduction in heroin supply on injecting drug use: analysis of data from needle and syringe programmes. PMID- 15292057 TI - L-Selectin(hi) but not the L-selectin(lo) CD4+25+ T-regulatory cells are potent inhibitors of GVHD and BM graft rejection. AB - Graft-versus-host disease (GVHD) is a major cause of morbidity and mortality after bone marrow transplantation (BMT). CD4(+)CD25(+) immune regulatory T cells (Tregs), long recognized for their critical role in induction and maintenance of self-tolerance and prevention of autoimmunity, are also important in the regulation of immune responses in allogeneic bone marrow (BM) and solid organ transplantation. Published data indicate that ex vivo activated and expanded donor Tregs result in significant inhibition of lethal GVHD. This study provides a direct comparison of LSel(hi) and LSel(lo) Tregs for GVHD inhibition and for the promotion of allogeneic BM engraftment. Imaging of green fluorescent protein positive effectors in GVHD control mice and LSel(hi) and LSel(lo) Treg-treated mice vividly illustrate the multisystemic nature of GVHD and the profound inhibition of GVHD by LSel(hi) Tregs. Data indicate that LSel(hi) Tregs interfere with the activation and expansion of GVHD effector T cells in secondary lymphoid organs early after BMT. Either donor- or host-type LSel(hi), but not LSel(lo), Tregs potently increased donor BM engraftment in sublethally irradiated mice, an event occurring independently of transforming growth factor beta signaling of host T cells. These data indicate that Treg cellular therapy warrants clinical consideration for the inhibition of GVHD and the promotion of alloengraftment. PMID- 15292058 TI - Cytotoxic activity of the maytansinoid immunoconjugate B-B4-DM1 against CD138+ multiple myeloma cells. AB - We tested the in vitro and in vivo antitumor activity of the maytansinoid DM1 (N(2')-deacetyl-N(2')-(3-mercapto-1-oxopropyl)-maytansine), a potent antimicrotubule agent, covalently linked to the murine monoclonal antibody (mAb) B-B4 targeting syndecan-1 (CD138). We evaluated the in vitro activity of B-B4-DM1 against a panel of CD138(+) and CD138(-) cell lines, as well as CD138(+) patient multiple myeloma (MM) cells. Treatment with B-B4-DM1 selectively decreased growth and survival of MM cell lines, patient MM cells, and MM cells adherent to bone marrow stromal cells. We further examined the activity of B-B4-DM1 in 3 human MM models in mice: (1) severe combined immunodeficient (SCID) mice bearing subcutaneous xenografts; (2) SCID mice bearing green fluorescent protein-positive (GFP(+)) xenografts; and (3) SCID mice implanted with human fetal bone (SCID-hu) and subsequently injected with patient MM cells. Tumor regression and inhibition of tumor growth, improvement in overall survival, and reduction in levels of circulating human paraprotein were observed in mice treated with B-B4-DM1. Although immunohistochemical analysis demonstrates restricted CD138 expression in human tissues, the lack of B-B4 reactivity with mouse tissues precludes evaluation of its toxicity in these models. In conclusion, B-B4-DM1 is a potent anti-MM agent that kills cells in an antigen-dependent manner in vitro and mediates in vivo antitumor activity at doses that are well tolerated, providing the rationale for clinical trials of this immunoconjugate in MM. PMID- 15292059 TI - Enhanced tumorigenesis in HTLV-1 tax-transgenic mice deficient in interferon gamma. AB - The oncoprotein Tax of human T-cell leukemia virus type I (HTLV-1) is the major mediator of viral pathogenesis in infected individuals. Expression of Tax under the regulation of the human granzyme B promoter in mice results in a lymphoproliferative disorder resembling adult T-cell leukemia/lymphoma (ATL). Tax expression is associated with the production of high levels interferon-gamma (IFN gamma) in HTLV-1-infected CD4(+) cells and Tax-transgenic tumors. We examined the role of IFN-gamma in tumorigenesis, by mating Tax-transgenic mice with a gene specific knockout for IFN-gamma. IFN-gamma(-/-) Tax(+)-transgenic mice show accelerated tumor onset (median, 4 versus 6 months), dissemination (median, 5 versus 7 months), and death (median, 7 versus 10 months), compared with IFN gamma(+/-) or IFN-gamma(+/+) Tax(+) mice. Pathologic and immunophenotypic characteristics of tumors from all genotypes are indistinguishable, except for enhanced interleukin 2 receptor-beta (IL-2Rbeta) and suppressed intercellular adhesion molecule-1 (ICAM-1) expression on tumors from IFN-gamma(-/-) Tax(+) transgenic mice. IFN-gamma(-/-) tumors demonstrate enhanced CD31 (platelet endothelial CAM-1 [PECAM-1]) staining compared with those from IFN-gamma(+/-) or IFN-gamma(+/+) Tax(+) mice. Angiogenesis-specific cDNA microarray analysis identified 4 mediators of angiogenic growth differentially expressed in tumors from Tax(+)IFN-gamma(-/-) mice compared with Tax(+)IFN-gamma(+/+) littermates. As confirmed by reverse transcription-polymerase chain reaction (RT-PCR), loss of IFN-gamma results in down-regulation of tumor necrosis factor-alpha (TNF-alpha) and tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) while up-regulating expression of vascular endothelial growth factor (VEGF) and tenascin C. These results provide insight into a possible mechanism by which IFN-gamma contributes to host resistance against HTLV-induced tumors through an angiostatic effect. PMID- 15292060 TI - Duration of immunosuppressive treatment for chronic graft-versus-host disease. AB - Chronic graft-versus-host disease (GVHD) requires long-term immunosuppressive therapy after hematopoietic cell transplantation. We retrospectively analyzed a cohort of 751 patients with chronic GVHD to identify characteristics associated with the duration of immunosuppressive treatment. Among the 274 patients who discontinued immunosuppressive therapy after resolution of chronic GVHD before recurrent malignancy or death, the median duration of treatment was 23 months. Results of a multivariable model showed that treatment was prolonged in patients who received peripheral blood cells, in male patients with female donors, in those with graft-versus-host HLA mismatching, and in those with hyperbilirubinemia or multiple sites affected by chronic GHVD at the onset of the disease. Nonrelapse mortality was increased among patients with HLA mismatching or hyperbilirubinemia but not among those with other risk factors associated with prolonged treatment for chronic GVHD. Nonrelapse mortality was also increased in older patients and those with older donors, in patients with platelet counts less than 100 000/microL or progressive onset of chronic GVHD from acute GVHD, and in those receiving higher doses of prednisone immediately before the diagnosis of chronic GVHD. After the dose of prednisone was taken into account, progressive onset was not associated with an increased risk of nonrelapse mortality. PMID- 15292061 TI - Overexpression of the NOTCH ligand JAG2 in malignant plasma cells from multiple myeloma patients and cell lines. AB - The NOTCH ligand, JAG2, was found to be overexpressed in malignant plasma cells from multiple myeloma (MM) patients and cell lines but not in nonmalignant plasma cells from tonsils, bone marrow from healthy individuals, or patients with other malignancies. In addition, JAG2 overexpression was detected in 5 of 5 patients with monoclonal gammopathy of undetermined significance (MGUS), an early phase of myeloma disease progression. This overexpression appears to be a consequence of hypomethylation of the JAG2 promoter in malignant plasma cells. An in vitro coculture assay was used to demonstrate that JAG2 induced the secretion of interleukin-6 (IL-6), vascular endothelial growth factor (VEGF), and insulin-like growth factor-1 (IGF-1) in stromal cells. Further, the induction of IL-6 secretion was blocked in vitro by interference with anti-Notch-1 monoclonal antibodies raised against the binding sequence of Notch-1 with JAG2. Taken together, these results indicate that JAG2 overexpression may be an early event in the pathogenesis of multiple myeloma involving IL-6 production. PMID- 15292062 TI - Low-dose thalidomide and donor lymphocyte infusion as adoptive immunotherapy after allogeneic stem cell transplantation in patients with multiple myeloma. AB - To improve the antimyeloma effect of donor lymphocyte infusion (DLI) after allogeneic stem cell transplantation in multiple myeloma, we investigated in a phase 1/2 study the effect of low-dose thalidomide (100 mg) followed by DLI in 18 patients with progressive disease or residual disease and prior ineffective DLI after allografting. The overall response rate was 67%, including 22% complete remission. Major toxicity of thalidomide was weakness grade I/II (68%) and peripheral neuropathy grade I/II (28%). Only 2 patients experienced mild grade I acute graft versus host disease (aGvHD) of the skin, while no grades II to IV aGvHD was seen. De novo limited chronic GvHD (cGvHD) was seen in 2 patients (11%). The 2-year estimated overall and progression-free survival were 100% and 84%, respectively. Adoptive immunotherapy with low-dose thalidomide and DLI induces a strong antimyeloma effect with low incidence of graft versus host disease. PMID- 15292063 TI - All-trans retinoic acid and anthracycline monochemotherapy for the treatment of elderly patients with acute promyelocytic leukemia. AB - Therapeutic results in elderly patients with acute promyelocytic leukemia (APL) have been generally reported as less effective than for younger patients. Patients 60 years or older with APL who were enrolled in 2 successive multicenter PETHEMA studies received induction therapy with all-trans retinoic acid (ATRA) and idarubicin, consolidation with 3 anthracycline monochemotherapy courses with or without ATRA, and maintenance with ATRA and low-dose chemotherapy. Eighty seven of 104 patients achieved complete remission (84%). Eighty-six proceeded to consolidation therapy (2 withdrew after the first and second courses). Deaths in remission occurred during consolidation and maintenance therapy in 3 and 4 patients, respectively. One patient showed molecular persistence after consolidation and 5 had a relapse. The 6-year cumulative incidence of relapse, leukemia-free survival, and disease-free survival were 8.5%, 91%, and 79%, respectively. A significantly higher incidence of low-risk patients found among the elderly, as compared to younger patients, may partially account for the low relapse rate observed. This study confirms the high antileukemic efficacy, low toxicity, and high degree of compliance of protocols using ATRA and anthracycline monochemotherapy for induction and consolidation therapy in elderly patients. PMID- 15292064 TI - Successful correction of the human beta-thalassemia major phenotype using a lentiviral vector. AB - beta-thalassemias are the most common single gene disorders and are potentially amenable to gene therapy. However, retroviral vectors carrying the human beta globin cassette have been notoriously unstable. Recently, considerable progress has been made using lentiviral vectors, which stably transmit the beta-globin expression cassette. Thus far, mouse studies have shown correction of the beta thalassemia intermedia phenotype and a partial, variable correction of beta thalassemia major phenotype. We tested a lentiviral vector carrying the human beta-globin expression cassette flanked by a chromatin insulator in transfusion dependent human thalassemia major, where it would be ultimately relevant. We demonstrated that the vector expressed normal amounts of human beta-globin in erythroid cells produced in in vitro cultures for unilineage erythroid differentiation. There was restoration of effective erythropoiesis and reversal of the abnormally elevated apoptosis that characterizes beta-thalassemia. The gene-corrected human beta-thalassemia progenitor cells were transplanted into immune-deficient mice, where they underwent normal erythroid differentiation, expressed normal levels of human beta-globin, and displayed normal effective erythropoiesis 3 to 4 months after xenotransplantation. Variability of beta globin expression in erythroid colonies derived in vitro or from xenograft bone marrow was similar to that seen in normal controls. Our results show genetic modification of primitive progenitor cells with correction of the human thalassemia major phenotype. PMID- 15292065 TI - Inhibition of thrombin generation by protein S at low procoagulant stimuli: implications for maintenance of the hemostatic balance. AB - The activated protein C (APC)-independent anticoagulant activity of protein S on tissue factor-induced thrombin generation was quantified in plasma. In absence of APC, protein S significantly decreased the endogenous thrombin potential (ETP) in a concentration-dependent manner. The APC-independent anticoagulant activity of protein S in plasma was not affected by phospholipid concentrations but strongly depended on tissue factor concentrations: protein S inhibited the ETP from 6% at 140 pM tissue factor to 74% at 1.4 pM tissue factor. Plasma with both 60% protein S and 140% prothrombin showed an ETP of 240% compared to normal plasma, suggesting an APC-independent protective role of protein S in the development of thrombosis as a result of protein S deficiency and the prothrombin-G20210A mutation. At high tissue-factor concentrations, protein S hardly expressed APC independent anticoagulant activity but exerted potent APC-cofactor activity when thrombomodulin or APC were added to plasma. Neutralization of protein S under these conditions resulted in a 20-fold reduction of the anticoagulant activity of APC. The present study shows that protein S effectively regulates coagulation at 2 levels: at low procoagulant stimuli, protein S maintains the hemostatic balance by directly inhibiting thrombin formation, and at high procoagulant stimuli, protein S restores the hemostatic balance via its APC-cofactor activity. PMID- 15292066 TI - IL-8 responsiveness defines a subset of CD8 T cells poised to kill. AB - CD8 T cells play a key role in host defense against intracellular pathogens. Efficient migration of these cells into sites of infection is therefore intimately linked to their effector function. The molecular mechanisms that control CD8 T-cell trafficking into sites of infection and inflammation are not well understood, but the chemokine/chemokine receptor system is thought to orchestrate this process. Here we systematically examined the chemokine receptor profile expressed on human CD8 T cells. Surprisingly, we found that CXC chemokine receptor 1 (CXCR1), the predominant neutrophil chemokine receptor, defined a novel interleukin-8/CXC ligand 8 (IL-8/CXCL8)-responsive CD8 T-cell subset that was enriched in perforin, granzyme B, and interferon-gamma (IFNgamma), and had high cytotoxic potential. CXCR1 expression was down-regulated by antigen stimulation both in vitro and in vivo, suggesting antigen-dependent shaping of the migratory characteristics of CD8 T cells. On virus-specific CD8 T cells from persons with a history of Epstein-Barr virus (EBV) and influenza infection, CXCR1 expression was restricted to terminally differentiated effector memory cells. In HIV-1 infection, CXCR1-expressing HIV-1-specific CD8 T cells were present only in persons who were able to control HIV-1 replication during structured treatment interruptions. Thus, CXCR1 identifies a subset of CD8 T cells poised for immediate cytotoxicity and early recruitment into sites of innate immune system activation. PMID- 15292067 TI - Immunomodulatory derivative of thalidomide (IMiD CC-4047) induces a shift in lineage commitment by suppressing erythropoiesis and promoting myelopoiesis. AB - Immunomodulatory derivative (IMiD) CC-4047, a new analog of thalidomide, directly inhibits growth of B-cell malignancies in vivo and in vitro and exhibits stronger antiangiogenic activity than thalidomide. However, there is little information on whether CC-4047 affects normal hematopoiesis. Here we investigated the effect of CC-4047 on lineage commitment and differentiation of hematopoietic stem cells. We found that CC-4047 effectively inhibits erythroid cell colony formation from CD34+ cells and increases the frequency of myeloid colonies. We also demonstrate that development of both erythropoietin-independent and erythropoietin-dependent red cell progenitors was strongly inhibited by CC-4047, while terminal red cell differentiation was unaffected. DNA microarray analysis revealed that red cell transcription factors, including GATA-1, GATA-2, erythroid Kruppel-like factor (EKLF), and growth factor independence-1B (Gfi-1b), were down-regulated in CC 4047-treated CD34+ cells, while myeloid transcription factors such as CCAAT/enhancer binding protein-alpha (C/EBPalpha), C/EBPdelta, and C/EBPepsilon were induced. Analysis of cytokine secretion indicated that CC-4047 induced secretion of cytokines that enhance myelopoiesis and inhibit erythropoiesis. In conclusion, these data indicate that CC-4047 might directly influence lineage commitment of hematopoietic cells by increasing the propensity of stem and/or progenitor cells to undergo myeloid cell development and concomitantly inhibiting red cell development. Therefore, CC-4047 provides a valuable tool to study the mechanisms underlying lineage commitment. PMID- 15292068 TI - Increased and pathologic emperipolesis of neutrophils within megakaryocytes associated with marrow fibrosis in GATA-1(low) mice. AB - Deletion of megakaryocytic-specific regulatory sequences of GATA-1 (Gata1(tm2Sho) or GATA-1(low) mutation) results in severe thrombocytopenia, because of defective thrombocytopoiesis, and myelofibrosis. As documented here, the GATA-1(low) mutation blocks megakaryocytic maturation between stage I and II, resulting in accumulation of defective megakaryocytes (MKs) in the tissues of GATA-1(low) mice. The block in maturation includes failure to properly organize alpha granules because von Willebrand factor is barely detectable in mutant MKs, and P selectin, although normally expressed, is found frequently associated with the demarcation membrane system (DMS) instead of within granules. Conversely, both von Willebrand factor and P-selectin are barely detectable in GATA-1(low) platelets. Mutant MKs are surrounded by numerous myeloperoxidase-positive neutrophils, some of which appear in the process to establish contact with MKs by fusing their membrane with those of the DMS. As a result, 16% (in spleen) to 34% (in marrow) of GATA-1(low) MKs contain 1 to 3 neutrophils embedded in a vacuolated cytoplasm. The neutrophil-embedded GATA-1(low) MKs have morphologic features (high electron density and negativity to TUNEL staining) compatible with those of cells dying from para-apoptosis. We suggest that such an increased and pathologic neutrophil emperipolesis may represent one of the mechanisms leading to myelofibrosis by releasing fibrogenic MK cytokines and neutrophil proteases in the microenvironment. PMID- 15292069 TI - Flowing cells through pulsed electric fields efficiently purges stem cell preparations of contaminating myeloma cells while preserving stem cell function. AB - Autologous stem cell transplantation, in the setting of hematologic malignancies such as lymphoma, improves disease-free survival if the graft has undergone tumor purging. Here we show that flowing hematopoietic cells through pulsed electric fields (PEFs) effectively purges myeloma cells without sacrificing functional stem cells. Electric fields can induce irreversible cell membrane pores in direct relation to cell diameter, an effect we exploit in a flowing system appropriate for clinical scale. Multiple myeloma (MM) cell lines admixed with human bone marrow (BM) or peripheral blood (PB) cells were passed through PEFs at 1.35 kV/cm to 1.4 kV/cm, resulting in 3- to 4-log tumor cell depletion by flow cytometry and 4.5- to 6-log depletion by tumor regrowth cultures. Samples from patients with MM gave similar results by cytometry. Stem cell engraftment into nonobese diabetic severe combined immunodeficient (NOD/SCID)/beta2m-/- mice was unperturbed by PEFs. Flowing cells through PEFs is a promising technology for rapid tumor cell purging of clinical progenitor cell preparations. PMID- 15292070 TI - Direct multiplex assay of lysosomal enzymes in dried blood spots for newborn screening. AB - BACKGROUND: Newborn screening for deficiency in the lysosomal enzymes that cause Fabry, Gaucher, Krabbe, Niemann-Pick A/B, and Pompe diseases is warranted because treatment for these syndromes is now available or anticipated in the near feature. We describe a multiplex screening method for all five lysosomal enzymes that uses newborn-screening cards containing dried blood spots as the enzyme source. METHODS: We used a cassette of substrates and internal standards to directly quantify the enzymatic activities, and tandem mass spectrometry for enzymatic product detection. Rehydrated dried blood spots were incubated with the enzyme substrates. We used liquid-liquid extraction followed by solid-phase extraction with silica gel to remove buffer components. Acarbose served as inhibitor of an interfering acid alpha-glucosidase present in neutrophils, which allowed the lysosomal enzyme implicated in Pompe disease to be selectively analyzed. RESULTS: We analyzed dried blood spots from 5 patients with Gaucher, 5 with Niemann-Pick A/B, 11 with Pompe, 5 with Fabry, and 12 with Krabbe disease, and in all cases the enzyme activities were below the minimum activities measured in a collection of heterozygous carriers and healthy noncarrier individuals. The enzyme activities measured in 5-9 heterozygous carriers were approximately one half those measured with 15-32 healthy individuals, but there was partial overlap of each condition between the data sets for carriers and healthy individuals. CONCLUSION: For all five diseases, the affected individuals were detected. The assay can be readily automated, and the anticipated reagent and supply costs are well within the budget limits of newborn-screening centers. PMID- 15292071 TI - False-negative result in the detection of an IgM monoclonal protein by capillary zone electrophoresis. PMID- 15292072 TI - A piece of my mind. Soul (re)searching. PMID- 15292073 TI - Reclaiming child soldiers' lost lives. PMID- 15292074 TI - Efforts grow to keep mentally ill out of jails. PMID- 15292075 TI - Disarming violent domestic abusers is key to saving lives, say experts. PMID- 15292079 TI - Rates of domestic violence in southern Iraq. PMID- 15292080 TI - Posttraumatic stress among survivors of bioterrorism. PMID- 15292081 TI - A community-based tuberculosis program in Cambodia. PMID- 15292082 TI - Mortality in a cohort of street youth in Montreal. AB - CONTEXT: Many studies have shown a high prevalence of sexually transmitted diseases, human immunodeficiency virus (HIV) infection, viral hepatitis, drug dependence, and mental health problems among street youth. However, data on mortality among these youth are sparse. OBJECTIVES: To estimate mortality rate among street youth in Montreal and to identify causes of death and factors increasing the risk of death. DESIGN, SETTING, AND POPULATION: From January 1995 to September 2000, 1013 street youth 14 to 25 years of age were recruited in a prospective cohort with semi-annual follow-ups. Original study objectives were to determine the incidence and risk factors for HIV infection in that population; however, several participants died during the first months of follow-up, prompting investigators to add mortality to the study objectives. Mortality data were obtained from the coroner's office and the Institut de la Statistique du Quebec. MAIN OUTCOME MEASURES: Mortality rate among participants and factors increasing the risk of death. RESULTS: Twenty-six youth died during follow-up for a mortality rate of 921 per 100 000 person-years (95% confidence interval [CI], 602-1350); this represented a standardized mortality ratio of 11.4. The observed causes of death were as follows: suicide (13), overdose (8), unintentional injury (2), fulminant hepatitis A (1), heart disease (1); 1 was unidentified. In multivariate Cox regression analyses, HIV infection (adjusted hazard ratio [AHR] = 5.6; 95% CI, 1.9-16.8), daily alcohol use in the last month (AHR = 3.2; 95% CI, 1.3-7.7), homelessness in the last 6 months (AHR = 3.0; 95% CI, 1.1-7.6), drug injection in the last 6 months (AHR = 2.7; 95% CI, 1.2-6.2), and male sex (AHR = 2.6; 95% CI, 0.9-7.7) were identified as independent predictors of mortality. CONCLUSIONS: Current heavy substance use and homelessness were factors associated with death among street youth. HIV infection was also identified as an important predictor of mortality; however, its role remains to be clarified. These findings should be taken into account when developing interventions to prevent mortality among street youth. PMID- 15292083 TI - Mental health, social functioning, and disability in postwar Afghanistan. AB - CONTEXT: More than 2 decades of conflict have led to widespread human suffering and population displacement in Afghanistan. In 2002, the Centers for Disease Control and Prevention and other collaborating partners performed a national population-based mental health survey in Afghanistan. OBJECTIVE: To provide national estimates of mental health status of the disabled (any restriction or lack of ability to perform an activity in the manner considered normal for a human being) and nondisabled Afghan population aged at least 15 years. DESIGN, SETTING, AND PARTICIPANTS: A national multistage, cluster, population-based mental health survey of 799 adult household members (699 nondisabled and 100 disabled respondents) aged 15 years or older conducted from July to September 2002. Fifty district-level clusters were selected based on probability proportional to size sampling. One village was randomly selected in each cluster and 15 households were randomly selected in each village, yielding 750 households. MAIN OUTCOME MEASURES: Demographics, social functioning as measured by selected questions from the Medical Outcomes Study 36-Item Short-Form Health Survey, depressive symptoms measured by the Hopkins Symptoms Checklist-25, trauma events and symptoms of posttraumatic stress disorder (PTSD) measured by the Harvard Trauma Questionnaire, and culture-specific symptoms of mental illness and coping mechanisms. RESULTS: A total of 407 respondents (62.0%) reported experiencing at least 4 trauma events during the previous 10 years. The most common trauma events experienced by the respondents were lack of food and water (56.1%) for nondisabled persons and lack of shelter (69.7%) for disabled persons. The prevalence of respondents with symptoms of depression was 67.7% (95% confidence interval [CI], 54.6%-80.7%) and 71.7% (95% CI, 65.0%-78.4%), and symptoms of anxiety 72.2% (95% CI, 63.8%-80.7%) and 84.6% (95% CI, 74.1%-95.0%) for nondisabled and disabled respondents, respectively. The prevalence of symptoms of PTSD was similar for both groups (nondisabled, 42.1%; 95% CI, 34.2% 50.1%; and disabled, 42.2%; 95% CI, 29.2%-55.2%). Women had significantly poorer mental health status than men did. Respondents who were disabled had significantly lower social functioning and poorer mental health status than those who were nondisabled. Feelings of hatred were high (84% of nondisabled and 81% of disabled respondents). Coping mechanisms included religious and spiritual practices; focusing on basic needs, such as higher income, better housing, and more food; and seeking medical assistance. CONCLUSIONS: In this nationally representative survey of Afghans, prevalence rates of symptoms of depression, anxiety, and PTSD were high. These data underscore the need for donors and health care planners to address the current lack of mental health care resources, facilities, and trained mental health care professionals in Afghanistan. PMID- 15292084 TI - Mental health symptoms following war and repression in eastern Afghanistan. AB - CONTEXT: Decades of armed conflict, suppression, and displacement resulted in a high prevalence of mental health symptoms throughout Afghanistan. Its Eastern province of Nangarhar is part of the region that originated the Taliban movement. This may have had a distinct impact on the living circumstances and mental health condition of the province's population. OBJECTIVES: To determine the rate of exposure to traumatic events; estimate prevalence rates of symptoms of posttraumatic stress disorder (PTSD), depression, and anxiety; identify resources used for emotional support and risk factors for mental health symptoms; and assess the present coverage of basic needs in Nangarhar province, Afghanistan. DESIGN, SETTING, AND PARTICIPANTS: A cross-sectional multicluster sample survey of 1011 respondents aged 15 years or older, conducted in Nangarhar province during January and March 2003; 362 households were represented with a mean of 2.8 respondents per household (72% participation rate). MAIN OUTCOME MEASURES: Posttraumatic stress disorder symptoms and traumatic events using the Harvard Trauma Questionnaire; depression and general anxiety symptoms using the Hopkins Symptom Checklist; and resources for emotional support through a locally informed questionnaire. RESULTS: During the past 10 years, 432 respondents (43.7%) experienced between 8 and 10 traumatic events; 141 respondents (14.1%) experienced 11 or more. High rates of symptoms of depression were reported by 391 respondents (38.5%); anxiety, 524 (51.8%); and PTSD, 207 (20.4%). Symptoms were more prevalent in women than in men (depression: odds ratio [OR], 7.3 [95% confidence interval [CI], 5.4-9.8]; anxiety: OR, 12.8 [95% CI, 9.0-18.1]; PTSD: OR, 5.8 [95% CI, 3.8-8.9]). Higher rates of symptoms were associated with higher numbers of traumas experienced. The main resources for emotional support were religion and family. Medical care was reported to be insufficient by 228 respondents (22.6%). CONCLUSIONS: In this survey of inhabitants of Nangarhar province, Afghanistan, prevalence rates of having experienced multiple traumatic events and having symptoms of anxiety, depression, and PTSD were high. These findings suggest that mental health symptoms in this region should be addressed at the population and primary health care level. PMID- 15292085 TI - Association between youth-focused firearm laws and youth suicides. AB - CONTEXT: Firearms are used in approximately half of all youth suicides. Many state and federal laws include age-specific restrictions on the purchase, possession, or storage of firearms; however, the association between these laws and suicides among youth has not been carefully examined. OBJECTIVE: To evaluate the association between youth-focused firearm laws and suicides among youth. DESIGN, SETTING, AND PARTICIPANTS: Quasi-experimental design with annual state level data on suicide rates among US youth aged 14 through 20 years, for the period 1976-2001. Negative binomial regression models were used to estimate the association between state and federal youth-focused firearm laws mandating a minimum age for the purchase or possession of handguns and state child access prevention (CAP) laws requiring safe storage of firearms on suicide rates among youth. MAIN OUTCOME MEASURES: Association between youth-focused state and federal firearm laws and rates of firearm, nonfirearm, and total suicides among US youth aged 14 to 17 and 18 through 20 years. RESULTS: There were 63 954 suicides among youth aged 14 through 20 years during the 1976-2001 study period, 39 655 (62%) of which were committed with firearms. Minimum purchase-age and possession-age laws were not associated with statistically significant reductions in suicide rates among youth aged 14 through 20 years. State CAP laws were associated with an 8.3% decrease (rate ratio [RR], 0.92; 95% confidence interval [CI], 0.86-0.98) in suicide rates among 14- to 17-year-olds. The annual rate of suicide in this age group in states with CAP laws was 5.97 per 100 000 population rather than the projected 6.51. This association was also statistically significant for firearm suicides (RR, 0.89; 95% CI, 0.83-0.96) but not for nonfirearm suicides (RR, 1.00; 95% CI, 0.91-1.10). CAP laws were also associated with a significant reduction in suicides among youth aged 18 through 20 years (RR, 0.89; 95% CI, 0.85-0.93); however, the association was similar for firearm suicides (RR, 0.87; 95% CI, 0.82 0.92) and nonfirearm suicides (RR, 0.91; 95% CI, 0.85-0.98). CONCLUSIONS: There is evidence that CAP laws are associated with a modest reduction in suicide rates among youth aged 14 to 17 years. As currently implemented, minimum age restrictions for the purchase and possession of firearms do not appear to reduce overall rates of suicide among youth. PMID- 15292086 TI - Trauma and PTSD symptoms in Rwanda: implications for attitudes toward justice and reconciliation. AB - CONTEXT: The 1994 genocide in Rwanda led to the loss of at least 10% of the country's 7.7 million inhabitants, the destruction of much of the country's infrastructure, and the displacement of nearly 4 million people. In seeking to rebuild societies such as Rwanda, it is important to understand how traumatic experience may shape the ability of individuals and groups to respond to judicial and other reconciliation initiatives. OBJECTIVES: To assess the level of trauma exposure and the prevalence of posttraumatic stress disorder (PTSD) symptoms and their predictors among Rwandans and to determine how trauma exposure and PTSD symptoms are associated with Rwandans' attitudes toward justice and reconciliation. DESIGN, SETTING, AND PARTICIPANTS: Multistage, stratified cluster random survey of 2091 eligible adults in selected households in 4 communes in Rwanda in February 2002. MAIN OUTCOME MEASURES: Rates of exposure to trauma and symptom criteria for PTSD using the PTSD Checklist-Civilian Version; attitudes toward judicial responses (Rwandan national and gacaca local trials and International Criminal Tribunal for Rwanda [ICTR]) and reconciliation (belief in community, nonviolence, social justice, and interdependence with other ethnic groups). RESULTS: Of 2074 respondents with data on exposure to trauma, 1563 (75.4%) were forced to flee their homes, 1526 (73.0%) had a close member of their family killed, and 1472 (70.9%) had property destroyed or lost. Among the 2091 total participants, 518 (24.8%) met symptom criteria for PTSD. The adjusted odds ratio (OR) of meeting PTSD symptom criteria for each additional traumatic event was 1.43 (95% CI, 1.33-1.55). More respondents supported the local judicial responses (90.8% supported gacaca trials and 67.8% the Rwanda national trials) than the ICTR (42.1% in support). Respondents who met PTSD symptom criteria were less likely to have positive attitudes toward the Rwandan national trials (OR, 0.77; 95% CI, 0.61-0.98), belief in community (OR, 0.76; 95% CI, 0.60-0.97), and interdependence with other ethnic groups (OR, 0.71; 95% CI, 0.56-0.90). Respondents with exposure to multiple trauma events were more likely to have positive attitudes toward the ICTR (OR, 1.10; 95% CI, 1.04-1.17) and less likely to support the Rwandan national trials (OR, 0.90; 95% CI, 0.84-0.96), the local gacaca trials (OR, 0.80; 95% CI, 0.72-0.89), and 3 factors of openness to reconciliation: belief in nonviolence (OR, 0.92; 95% CI, 0.87-0.97), belief in community (OR, 0.92; 95% CI, 0.87-0.98), and interdependence with other ethnic groups (OR, 0.86; 95% CI, 0.81-0.92). Other variables that were associated with attitudes toward judicial processes and openness to reconciliation were educational level, ethnicity, perception of change in poverty level and access to security compared with 1994, and ethnic distance. CONCLUSIONS: This study demonstrates that traumatic exposure, PTSD symptoms, and other factors are associated with attitudes toward justice and reconciliation. Societal interventions following mass violence should consider the effects of trauma if reconciliation is to be realized. PMID- 15292087 TI - Quality of malnutrition assessment surveys conducted during famine in Ethiopia. AB - CONTEXT: During 1999 and 2000, approximately 10 million people were affected by famine in Ethiopia. Results of nutrition assessments and surveys conducted by humanitarian organizations were used by donors and government agencies to determine needs for food aid and to make other decisions on geographic allocation of limited resources; however, accurate results might have been hampered by methodological errors. OBJECTIVES: To identify common methodological errors in nutrition assessments and surveys and to provide practical recommendations for improvement. DESIGN AND SETTING: Nutrition assessments and surveys (n = 125) conducted by 14 nongovernmental organizations (NGOs) in 54 woredas (districts) in Ethiopia from May 1, 1999, through July 31, 2000. Surveys were ranked as valid and precise according to 5 criteria: use of population proportional to size sampling, sample size, number of clusters, number of children per cluster, and use of weight-for-height index. MAIN OUTCOME MEASURES: Number and proportion of surveys that used standard, internationally accepted methods and reported valid and precise results. RESULTS: Fifty-eight of the 125 surveys (46%) were not intended to be standard 30 x 30 cluster surveys. Of the remaining 67 surveys, 6 (9%) met predetermined criteria for validity and precision. All 67 used the anthropometric index of weight-for-height, with 58 (87%) reporting z scores. Fifty-four (81%) used nonrandom sampling without consideration of population size and 6 (9%) had sample sizes of fewer than 500 persons. CONCLUSIONS: Major methodological errors were identified among 30 x 30 cluster surveys designed to measure acute malnutrition prevalence in Ethiopia during the famine of 1999-2000. Donor agencies and NGOs should be educated about the need for improved quality of nutrition assessments and their essential role in directing allocation of scarce food resources. PMID- 15292088 TI - Attention-deficit/hyperactivity disorder in adults. PMID- 15292089 TI - Street youth mortality: leaning with intent to fall. PMID- 15292090 TI - Mental health in postwar afghanistan. PMID- 15292091 TI - JAMA patient page. Malnutrition in children. PMID- 15292092 TI - Prolactin in the male: 25 years later. PMID- 15292093 TI - Ethical offspring? PMID- 15292094 TI - Why the WHO recommendations for eosin-nigrosin staining techniques for human sperm vitality assessment must change. PMID- 15292095 TI - The clinical conundrum of complete asthenospermia. PMID- 15292097 TI - Control of penile erection by the melanocortinergic system: experimental evidences and therapeutic perspectives. PMID- 15292098 TI - Expression of the p63 and Notch signaling systems in rat testes during postnatal development: comparison with their expression levels in the epididymis and vas deferens. AB - The role of tubular structures that contribute to the passage of spermatozoa is not solely passive; these structures actively contribute to their own functions, although these tubules and ducts are contiguous and collaborate in the development of the male gamete along their lengths. The testis has the specific function to generate spermatozoa and spermatozoa undergo numerous changes as they pass through the epididymis. A member of the p53 family of genes, p63, is highly expressed in the basal layers of epithelial tissues and plays a key role in maintaining their cell populations, whereas Notch 1 and its ligand Jagged 2 have an important role in the differentiation of germ cells and Jagged 2 is up regulated by TAp63, one of the p63 isoforms, which transactivates p53 target genes and induces apoptosis. Although the presence of p63 in most epithelia is established, the role of p63 and its possible relationship with the Notch system in the seminiferous epithelium have not been examined. Therefore, we investigated the expression of p63, Jagged 2, and Notch 1 in the testis during postnatal development in comparison with their expression levels in the vaso-epididymal epithelium. In the testis, the expression of TAp63 mRNA increased at day 14 after birth and the expressions of Jagged 2 and Notch 1 mRNA increased at day 16 after birth, suggesting that TAp63-mediated Jagged 2 induction activates the Notch signaling system. On the other hand, the strong signal of DeltaNp63 mRNA was already recognized in the vas deferens at day 0 after birth and advanced chronologically along the duct to the caput epididymis and p63 protein was expressed in basal cells in their epithelium, whereas the mRNAs of Jagged 2 and Notch 1 were maintained at a low level. Consequently, examination of our data raises the probability that TAp63 has an important role for maintenance of germ cell numbers, triggering or balancing the development, differentiation, and apoptosis of germ cells in the testis, which is completely different from the role of DeltaNp63 in other epithelial tissues. PMID- 15292099 TI - Dominant prostasome immunogens for sperm-agglutinating autoantibodies of infertile men. AB - The presence of naturally occurring anti-sperm antibodies (ASA) is a well-known cause of infertility in men and women, but the antigens for these antibodies are poorly characterized. We have previously shown that prostasomes adhere to sperm cells and that prostasomes are major targets for ASA associated with infertility. These autoantigens have not been characterized. We used 2-dimensional electrophoresis, immunoblotting, and mass-spectrometry to identify the prostasome antigens for these autoantibodies. By these techniques, we revealed that prolactin-inducible protein (PIP) and clusterin were dominant prostasome immunogens for sperm-agglutinating autoantibodies of 20 patients with immunological infertility. PIP was identified by 19 of 20 (95%) patient sera and clusterin by 17 of 20 (85%). In addition, 10 sporadically occurring prostasomal antigens were identified in this context, viz alcohol dehydrogenase [NADP+], annexin I, annexin III, BRCA1-associated ring domain protein 1, heat shock 27-kd protein, isocitrate dehydrogenase, lactoylglutathione lyase, NG,NG dimethylarginine dimethylaminohydrolase 1, peroxiredoxin 2, and syntenin 1. PMID- 15292100 TI - Insulin-dependent diabetes affects testicular function by FSH- and LH-linked mechanisms. AB - A study was conducted to form a unified hypothesis regarding the gonadotropin related mechanisms that underlie alterations in the male reproductive system in individuals with diabetes. Streptozotocin-induced diabetes resulted in reduced fertility, prolificacy, and libido. Testes showed a marked decrease in the number and function of Leydig cells, the latter manifested as changes in the expression of biochemical markers, including the GLUT-3 hexose transporter, c-kit, insulin like growth factor I (IGF-I), androgen receptors, and overall tyrosine phosphorylation, as assessed by Western blot and immunocytochemical analyses. The expression of c-kit, IGF-I, insulin, and follicle-stimulating hormone (FSH) receptors in the seminiferous tubules was also affected. Serum levels of luteinizing hormone (LH), FSH, and testosterone significantly decreased. There was a significant (P <.05) correlation between the serum levels of insulin and FSH. No significant correlation was found between the serum levels of insulin or glucose and LH. On the basis of our results, we conclude that, in insulin dependent diabetes, 1) Leydig cell function and testosterone production decrease because of the absence of the stimulatory effect of insulin on these cells and an insulin-dependent decrease in FSH, which, in turn, reduces LH levels; and 2) sperm output and fertility are reduced because of a decrease in FSH caused by a reduction in insulin. PMID- 15292101 TI - Male hormonal contraception: suppression of spermatogenesis by injectable testosterone undecanoate alone or with levonorgestrel implants in chinese men. AB - Monthly injections of testosterone undecanoate (TU) act as a male contraceptive by reversibly suppressing spermatogenesis to azoospermia or severe oligoazoospermia in 95% of Chinese men. In 5% of Chinese men, however, monthly TU administered alone fails to suppress spermatogenesis into contraceptive ranges, or sperm "rebound," leading to occurrences of pregnancy during treatment. Since combinations of progestins and androgens are associated with greater degrees of sperm suppression in white men, we hypothesized that the combination of TU and the progestin levonorgestrel (LNG) would result in improved spermatogenic suppression in Chinese men. Sixty-two healthy Chinese men were randomly assigned to one of the following 3 regimens: group I (n = 21) received 4 LNG rods (75 mg each), which were followed 4 weeks later by 500 mg of TU by intra-muscular (IM) injection every 8 weeks for 24 weeks; group II (n = 20) received 4 LNG implants, which were followed 4 weeks later by 1000 mg of TU by IM injection every 8 weeks for 24 weeks; and group III (n = 21) received TU 1000 mg by IM injection every 8 weeks for 24 weeks. Sperm counts, serum testosterone (T), luteinizing hormone, follicle-stimulating hormone, and LNG were measured every 2 weeks before, during, and after treatment. During treatment, group II demonstrated a trend toward a greater attainment of azoospermia than groups I and III (90% vs 62% [group I] vs 67% [group III]; P =.09). Attainments of either azoospermia or oligozoospermia (sperm density, <3 x 10(6)/mL) were 95%, 100%, and 86% for groups I, II, and III, respectively (P >.05 for comparisons between groups). Spermatogenesis in all subjects returned to the normal range after the implants were removed. No serious adverse events and no significant changes in serum chemistry occurred during the study. These results demonstrate that the combination of IM injections of high dose TU every 2 months and LNG implants is associated with marked suppression of spermatogenesis in Chinese men. The combination of high-dose TU every 2 months and LNG implants is a promising candidate for future large-scale efficacy studies of hormonal male contraception in Chinese men. PMID- 15292102 TI - Glu298Asp endothelial nitric oxide synthase polymorphism is a risk factor for erectile dysfunction in the Mexican Mestizo population. AB - Penile erection depends on the balanced action between antagonist vasoactive molecules such as nitric oxide (NO) and angiotensin. Endothelial nitric oxide synthase (eNOS) and angiotensin-converting enzyme (ACE) polymorphisms have been associated with endothelial dysfunction, which is described as a cause of erectile dysfunction (ED). Endothelial NOS and ACE are both regulators of vascular and corporal smooth muscle tone, which are connected by interaction between the NO-cyclic guanosine monophosphate pathway and the renin-angiotensin system. We analyzed the frequencies of 894 G/T (Glu298Asp) eNOS and ACE I/D polymorphisms in Mexican patients with ED (n=53) and in an age-matched control group (n=62). The populations analyzed were in Hardy Weinberg equilibrium. We found significant differences in allelic (chi2=4.42; P=.03) and genotypic frequencies (chi2=3.96; P=.04) between patients and controls for the 894 G/T eNOS polymorphism. Presence of the 894T allele in carriers increased the risk of ED (odds ratio [TT + GT versus GG] = 2.37; 95% confidence interval, 1.08 to 5.21; P=.02). Multiple logistic regression analysis showed that the Glu298Asp polymorphism was an independent factor for ED, as was diabetes mellitus, hypertension, cardiac disease, and cigarette smoking. No association was found between ACE I/D polymorphism and ED in the population studied. Therefore, our results suggest that Glu298Asp eNOS polymorphism plays a role as a genetic susceptibility factor for ED. PMID- 15292103 TI - Enkephalin-degrading enzymes in normal and subfertile human semen. AB - Opioid peptides have been reported to have important functions in human reproduction. Indeed, very high concentrations of enkephalins and their degrading enzymes have been reported in human semen. In the present paper, we compare the activity of two enkephalin-degrading enzymes, aminopeptidase N and neutral endopeptidase 24.11, in different fractions of semen from normozoospermic, fertile men and from subfertile patients with different abnormalities revealed by spermiogram analysis (asthenozoospermia, necrozoospermia, and teratozoospermia). High levels of activity of aminopeptidase N were found in the soluble and particulate sperm fractions of semen from patients presenting asthenozoospermia with necrozoospermia. In contrast, lower aminopeptidase N activity was measured in the soluble sperm fraction of asthenozoospermic semen. The percentage of dead spermatozoa was positively correlated with aminopeptidase N activity in both soluble and particulate sperm fractions. In contrast, the percentage of immobile spermatozoa was negatively correlated with aminopeptidase activity in soluble and particulate sperm, and in prostasome fractions. Levels of activity of neutral endopeptidase were found to be unaltered among the different conditions. In summary, the results of the present study indicate that alterations in the activity of aminopeptidase N may be one of the molecular components that contribute to male human subfertility. PMID- 15292104 TI - Preservation of testicular arteries during subinguinal microsurgical varicocelectomy: clinical considerations. AB - Microsurgical varicocelectomy with intentional preservation of the testicular artery(ies) is regarded as the gold standard approach to varicocele repair. We sought to determine whether the number of testicular arteries preserved at the time of micro-surgical varicocelectomy predicts improvement in postoperative semen parameters. We analyzed the records of 334 infertile men who underwent varicocelectomy performed by a single surgeon using a subinguinal microsurgical technique between July 1996 and January 2003. We examined the association between the number of testicular arteries preserved at the time of varicocelectomy and serum follicle-stimulating hormone (FSH), luteinizing hormone (LH), varicocele grade, testicular volume, and postoperative improvement in semen parameters. Unilateral, left-sided varicocelectomy was performed in 194 men, while bilateral varicocelectomy was performed in 140 men. Mean (+/-SE) sperm concentration (20.1 +/- 1.5 x 10(6)/mL to 26.7 +/- 1.9 x 10(6)/mL, P =.001), percent motility (24.7 +/- 1.0% to 30.9 +/- 1.2%, P =.001), and percent normal morphology (35.8 +/- 1.4% to 37.7 +/- 1.5%, P =.046) improved significantly following varicocelectomy. The mean number of preserved testicular arteries was 1.5 on the left (range, 1-4) and 1.5 on the right (range, 1-4). The number of testicular arteries preserved at the time of varicocelectomy did not correlate significantly with preoperative assessment of serum FSH, LH, varicocele grade, and testicular volume or with postoperative improvement in semen parameters. Our data indicate that preoperative parameters are not predictive of the number of testicular arteries identified at the time of microsurgery. These data also suggest that the number of arteries identified and preserved with meticulous spermatic cord dissection does not correlate with improvement in semen parameters. PMID- 15292105 TI - Sperm membrane dynamics assessed by changes in lectin fluorescence before and after capacitation. AB - Sperm capacitation is correlated with acquisition of fertilizing ability, and the molecular events underlying this process are only beginning to be understood. A number of membrane changes associated with capacitation have been documented. In this study we used lectin probes to identify changes in glycoprotein localization as a result of capacitation of mouse sperm. Eight lectins (LEA, PSA, PNA, AAA, UEA-1, WGA, STA, and TPA) stained regions of the mouse sperm head, tail, or both. No changes in tail staining patterns were detected when sperm were incubated under capacitating conditions. In contrast, 7 of 8 lectins tested showed clear shifts in staining patterns in the sperm head as a result of incubation under capacitating conditions. When staining patterns were quantified, a distinct heterogeneity within the sperm population was observed. Each lectin displayed 3 distinct staining patterns in both uncapacitated and capacitated sperm samples. The least common pattern represented the acrosome-reacted (AR) pattern, as independently assessed by lectin staining of ionophoretreated sperm that were >95% AR as judged by Coomassie staining. However, a reciprocal shift in the two predominant staining patterns was correlated with capacitation and suggests that changes in distribution of cell surface proteins during capacitation constitute part of the molecular changes which result in changes in sperm function acquired during this process. PMID- 15292106 TI - Calcification of the epididymis and the tunica albuginea of the corpora cavernosa in patients on maintenance hemodialysis. AB - The aims of this study were to determine the incidence rates of genital calcification in male hemodialysis patients based on ultrasonography findings and to identify risk factors for this condition. Twenty-three male end-stage renal disease (ESRD) patients (mean age, 51.4 +/- 12.1 years) who were on maintenance hemodialysis underwent penile and scrotal ultrasonography. For each case, we recorded the underlying renal disease and measured serum levels of phosphorus, intact parathormone, and calcium x phosphorus product. Patients were also questioned about erectile dysfunction. The control group consisted of 22 consecutive patients (mean age, 51 years) with type 2 diabetes mellitus with normal renal function who underwent penile and scrotal ultrasonography for various reasons. In the ESRD group, ultrasound revealed calcification of the tunica albuginea of the corpora cavernosa in 15 patients (65%) and calcification of the epididymis in 16 patients (70%; 14 bilateral and 2 unilateral cases). Twenty patients (87%) showed calcification of the epididymis and/or the tunica, and 10 (43%) showed calcification of both these tissues. The rates of epididymal and penile calcification in the ESRD patients and the controls were significantly different (P <.001 for both). There were no significant differences between patients with and without penile and epididymal calcification with respect to age, hemodialysis duration, frequencies of elevated serum phosphorus, elevated serum intact parathormone, elevated calcium x phosphorus product, and frequency of erectile dysfunction (ED) (P >.05 for all). Ultrasonography revealed high rates of penile (tunica albuginea of the corpora cavernosa) and epididymal calcification (65% and 70%, respectively) in the ESRD patients studied, but no association was found between risk factors such as age, underlying renal disease, hemodialysis duration, frequencies of elevated serum phosphorus, elevated serum intact parathormone, and elevated calcium x phosphorus product. PMID- 15292107 TI - Detection of a short CCR5 messenger RNA isoform in human spermatozoa. AB - It has recently been reported that the Regulated upon Activation of Normal T cells Expressed and Secreted (RANTES) chemokine may exhibit a chemotactic effect on sperm. The RANTES chemokine acts on target cells by binding to the CCR5 receptor, which is present on the surface of various cells. Spermatozoa contain a complex repertoire of messenger RNAs (mRNAs) that may provide an insight into past events of spermatogenesis. The type and amount of CCR5 chemokine receptor transcript were investigated in spermatozoa that were isolated by the swim-up method from semen samples of men with normozoospermia. Using reverse transcription and real-time quantitative polymerase chain reaction (RQ-PCR) analysis, we found that the CCR5 mRNA isoform in human spermatozoa consists of exons 3 and 4, and is shorter than the transcript in leukocytes. This CCR5 transcript may represent a more stable mRNA isoform; one that is used to biosynthesize the CCR5 receptor in spermatogenesis or the early stages of embryonic development. PMID- 15292108 TI - Cinnoxicam and L-carnitine/acetyl-L-carnitine treatment for idiopathic and varicocele-associated oligoasthenospermia. AB - The objective of this study was to detect a therapy for idiopathic and varicocele associated oligoasthenospermia (OAT). Idiopathic and varicocele OAT patients were randomized into 3 groups. Each group was composed of varying degrees of left varicoceles (graded into 5 grades with echo-color Doppler) and of idiopathic OATs. Group 1 used a placebo, group 2 used oral L-carnitine (2 g/d) + acetyl-L carnitine (1 g/d), group 3 used L-carnitine/acetyl-L-carnitine + 1 x 30-mg cinnoxicam suppository every 4 days. Drugs were administered for 6 months. The groups were composed as follows: group 1, 71 varicoceles and 47 idiopathic OATs; group 2, 62 varicoceles and 39 idiopathic OATs; group 3, 62 varicoceles and 44 idiopathic OATs. Sperm concentration, motility, and morphology before during and after treatments were assessed. Pregnancy rates and side effects were recorded. Group 1 did not have modified sperm patterns during treatment. Group 2 had significantly increased sperm patterns at 3 and 6 months into therapy in idiopathic patients and in patients with grades I, II, and III varicocele, but not in grades IV and V. Group 3 had significantly increased sperm parameters in all patients, with the exception of grade V varicocele. Group 3 sperm patterns proved significantly higher during therapy than group 2. All sperm patterns fell to baseline after therapy suspension. Minor side effects occurred. Pregnancy rates were 1.7% (group 1), 21.8% (group 2), and 38.0% (group 3) (P <.01). L carnitine/acetyl-L-carnitine + cinnoxicam suppositories proved a reliable treatment for low-grade varicoceles and idiopathic OATs. PMID- 15292110 TI - Concentration of glutathione and expression of glutathione peroxidases 1 and 4 in fresh sperm provide a forecast of the outcome of cryopreservation of human spermatozoa. AB - Oxidative stress imbalance potentially leads to damage of the structure of the cell and macromolecules such as plasma membrane components, proteins, and DNA. The plasma membrane of the sperm cell, which has high levels of polyunsaturated fatty acids, renders it particularly sensitive to free radical-mediated attacks. The freezing and subsequent thawing of sperm is a physically stressful process carried out during routine procedures in assisted reproduction techniques, which results in a highly variable and unpredictable reduction in the number of motile sperm cells. Subsequently, oxidative status can positively or negatively affect the motility, viability, and fertilizing capacity of thawed sperm. These effects are counteracted by various oxidative defense enzymes and anti-oxidants such as glutathione peroxidase isoforms GPx1 and GPx4, glutathione reductase (GR), and cellular glutathione (reduced) (GSH). In this way, oxidative status could represent a predictive marker of sperm quality following the freeze-thaw process. This study was based on 56 human sperm samples. We observed direct positive and negative relationships between the postthaw motile sperm recovery rate and GPx1 and GPx4 expression and activity, on the one hand, and GSH concentrations, on the other. No correlation was found between this recovery rate and GR or basic semen parameters. Predictive values clearly demonstrate that, among the molecules analyzed, the most accurate diagnoses result when analyses are conducted for GPx1 and GPx1 messenger RNA expression, GPx1 and GPx4 enzymatic activity, and GSH concentration. In conclusion, a reserve of glutathione, together with GPx expression, is necessary to eliminate free radicals using GSH or a like structural protein and seems to be essential for a good postthaw recovery. These molecules can be employed as indicators of postthaw sperm quality. PMID- 15292111 TI - Males with subnormal hypo-osmotic swelling test scores have lower pregnancy rates than those with normal scores when ovulation induction and timed intercourse is used as a treatment for mild problems with sperm count, motility, or morphology. AB - This study was designed to evaluate the effectiveness and clinical usefulness of the hypo-osmotic swelling (HOS) test in predicting successful conception in couples in which men with mild male-factor infertility criteria were undergoing a timed vaginal inter-course protocol. One hundred couples, in which mild male infertility was the only abnormality, were included in the study. Semen was analyzed according to standard World Health Organization (WHO) criteria and subjected to the HOS test. Patients were divided into 2 groups: group 1 (n=39) with normal HOS test and group 2 (n=61) with abnormal HOS test. All women underwent three consecutive cycles of follicular growth ultrasound monitoring and timed intercourse. Ten couples were exclude from the study. Ten clinical pregnancies were achieved in group 1 with a pregnancy rate per patient and per cycle of 28.5% and 9.5%, respectively. In group 2, 6 pregnancies were achieved, with a pregnancy rate per patient and per cycle of 10.9% and 3.6%, respectively. Both pregnancy rates per patients and per cycle was significantly higher (P <.05) in group 1 than in group 2. The HOS test may be considered an easy and reliable test in identifying among subfertile men those who have a greater possibility of causing pregnancy. PMID- 15292112 TI - Vasectomy-dependent dysregulation of a local renin-angiotensin system in the epididymis of the cynomolgus monkey (Macaca fascicularis). AB - The mammalian epididymis is a fundamental organ for sperm cell maturation; it allows mammals to acquire their fertilizing ability. We have previously shown that during obstruction in cases of vasectomy, gene expression profiles were modified in human and cynomolgus monkey epididymides. Paracrine factors thus appear to be key elements in local gene expression along the epididymis. Local renin-angiotensin systems (RAS) have been described in many other organs as paracrine regulators of gene expression. This work demonstrates the presence of a local RAS in the epididymis of the cynomolgus monkey and investigates the vasectomy-dependent changes occurring in this system. After unilateral vasectomy in 4 monkeys (two for 3 days and two others for 7 days), the presence of two major components of the RAS (ie, angiotensinogen [ANG] and the type 1 receptor to angiotensin II [AT-I]) was evaluated in the vasectomized and the normal controlateral epididymides of each monkey. We also show by in situ hybridization that the principal cells of the epididymis express ANG and AT-I mRNAs and immunohistochemistry permitted to verify the co-localization of the AT-I protein and mRNA. Quantitative comparisons of individual variations in the mRNA and protein profiles for ANG and AT-I revealed that vasectomy altered the RAS expression profiles in an individual manner, thus confirming its role as a local system. This study provides a good basis for further investigation of the possible implications of the RAS in the physiology of the epididymis. Furthermore, the individual dependent modifications are in accordance with the very fluctuating results obtained in the fertility status of human patients undergoing a vasectomy reversal. The variations observed in the RAS expression profiles may be a good model to study the causes of the overall epididymal gene expression dysregulation that follows vasectomy and potentially affects fertility. PMID- 15292113 TI - AnnexinV binding and merocyanine staining fail to detect human sperm capacitation. AB - The signaling pathways that characterize the process of capacitation of human spermatozoa are still largely unknown. Modifications in the lipid architecture of the sperm plasma membrane have been described in spermatozoa from different species, including translocation of phosphatidylserine (PS) from the inner to the outer leaflet and increased phospholipid disorder in the membrane. In human spermatozoa, however, results of PS exposure are controversial. In the present study, we used flow cytometry to investigate both membrane PS exposure by Annexin V (Ann V) binding and lipid disorder by merocyanine 540 (M540) staining, in swimup-selected live spermatozoa after incubation in conditions leading to capacitation. Our results indicate that neither probe is able to detect capacitation-related membrane modifications. Investigation of the nature of PS exposure and M540-positive live cells was then carried out. We found that M540 stains elements devoid of nuclei are present in seminal plasma. Live PS-exposing cells were mainly represented by damaged spermatozoa as revealed by the occurrence of a negative correlation between PS exposure and normal morphology and motility in unselected samples. The same cells were also positive for M540. These results demonstrate that Ann V and M540 binding in human sperm samples mainly detects cells with early membrane degeneration as well as dead cells, which is in agreement with findings obtained for somatic cells in which the two probes recognize cells with a damaged membrane due to the apoptotic process. PMID- 15292114 TI - In vivo effects of histone-deacetylase inhibitor trichostatin-A on murine spermatogenesis. AB - The acetylation state of core histones is controlled by two classes of enzymes, histone acetyl transferases (HATs) and histone deacetylases (HDACs). HDAC inhibitors, such as trichostatin-A (TSA), are able to induce cell cycle arrest by stimulating transcription of genes that negatively regulate cell growth and survival. However, little is known about the effect of HDAC inhibitors on spermatogenesis. TSA treatment of cultured murine germ cells from whole testes resulted in an increase of histone H4 acetylation in round spermatids, suggesting that a hypoacetylated state of these cells is important for their normal differentiation. In the present study, the in vivo effects of TSA on murine spermatogenesis were investigated. Subcutaneously applied TSA resulted in a dose dependent decrease in relative testis weight due to impaired spermatogenesis. No obvious toxic effects of TSA treatment could be found. A second animal experiment confirmed that male mice receiving TSA under the same conditions as in the first experiment became infertile. This phenomenon was completely reversible. No evidence of histone H4 hyperacetylation in round spermatids could be found; however, the number of spermatids significantly decreased with increasing TSA concentrations. Additionally, a dramatic loss of pachytene-diplotene spermatocytes due to increased apoptosis was observed. This suggests that TSA was mainly effective at the level of meiosis. The other male reproductive organs showed no morphological changes compared to controls, suggesting that TSA action on the testis was not mediated by sex hormones. PMID- 15292115 TI - Phenotypic characteristics of male subfertility and its familial occurrence. AB - Genetic factors can attribute to male subfertility. A case-control study was carried out to investigate familial occurrence of male subfertility and the phenotypic characteristics of familial male subfertility. The medical data and family histories of 253 severely subfertile men who were candidates for intracytoplasmic sperm injection were compared to the data from 243 randomly selected men. The prevalence of male fertility problems among brothers and maternal uncles of subfertile men was significantly higher than among controls (brothers 10.4% vs 0.5% and maternal uncles 1.7% vs 0.2%). The phenotypes of subfertile men with a positive family history more often showed normal levels of follicle-stimulating hormone (FSH) and luteinizing hormone (LH) compared to the phenotypes of subfertile men with a negative family history. In addition, subfertile men with a positive family history had a lower percentage of motile sperm. Genetic aberrations, including a chromosomal abnormality or a microdeletion of the Y chromosome, were present in 13.8% of the severely subfertile men. Male subfertility appears to have a familial occurrence, especially among brothers and maternal uncles. Furthermore, examination of the data suggests that subfertile men with a familial occurrence of male subfertility more often have normal levels of FSH and LH and a lower percentage of motile sperm. PMID- 15292116 TI - Electrophoretic characterization of the human sperm-specific enolase at different stages of maturation. AB - The presence of a sperm-specific enolase isoform (ENO-S) in human ejaculated spermatozoa was previously demonstrated. The objective of this study was to characterize this ENO-S in spermatozoa at different steps of maturation. Sperm ENO-S was characterized in testicular, epididymal, and ejaculated spermatozoa to determine whether any change occurred in the isoform patterns of this enzyme during epididymal maturation. In testicular sperm, ENO-S was present under 2 main bands named S1 and S3. In epididymal sperm, S1 and S3 bands and a prominent additional S2 band, with the same electrophoretic properties as the S isoform of ejaculated sperm, were visualized. In the testicular extracts obtained from testes in which no spermatozoa were visualized by histologic analysis, none of the 3 ENO-S bands was found. ENO-S exists as different isoforms (electrophoretic variants) in the different stages of sperm maturation. Passage through the epididymis seems to play a major role in the maturational process of this sperm specific enolase. PMID- 15292117 TI - Oxidative stress and antioxidant therapy: their impact in diabetes-associated erectile dysfunction. AB - Oxidative stress is believed to affect the development of diabetic-associated vasculopathy, endothelial dysfunction, and neuropathy within erectile tissue. Our hypothesis is that, given adequate concentrations of the oxygen free radical scavenger vitamin E, enhanced levels of circulating nitric oxide (NO) should improve erectile function with the potential for a synergistic effect with a phosphodiesterase type 5 (PDE5) inhibitor. Twenty adult male Sprague-Dawley streptozotocin-induced (60 mg/kg intraperitoneally) diabetic rats were placed in 4 therapeutic groups (n = 5 per group) as follows: 1) peanut oil only (diabetic control), 2) 20 IU of vitamin E per day, 3) 5 mg/kg of sildenafil per day, and 4) vitamin E plus sildenafil using oral gavage for 3 weeks. In addition, 5 age matched rats served as normal nondiabetic controls (normal). Erectile function was assessed by measuring the rise in intracavernous pressure (ICP) following cavernous nerve electrostimulation. Penile tissue was evaluated for neuronal NO synthase (nNOS), smooth muscle alpha-actin, nitrotyrosine, and endothelial cell integrity. Urine nitrite and nitrate (NOx) concentration was quantified, and electrolytes were tested by a serum biochemistry panel. A significant decrease in ICP was recorded in the diabetic animals, with improvement measured in the animals receiving PDE5 inhibitors either with or without vitamin E; the controls had a pressure of 54.8 +/- 5.3 cm H2O, the vitamin E group had a pressure of 73.5 +/- 6.6 cm H2O, the sildenafil group had a pressure of 78.4 +/- 10.77 cm H2O, and the vitamin E plus sildenafil group had a pressure of 87.9 +/- 5.5 cm H2O (P <.05), compared with the normal cohorts at 103.0 +/- 4.8 cm H2O. Histoexaminations showed improved nNOS, endothelial cell, and smooth muscle cell staining in the vitamin E plus sildenafil group compared to the control animals. Urine NOx increased significantly in all the diabetic groups but was blunted in the vitamin E and vitamin E plus sildenafil groups. A significant increase in positive staining for nitrotyrosine was observed in the vitamin E plus sildenafil group. Vitamin E enhanced the therapeutic effect of the PDE5 inhibitor in this study, supporting the potential use of oxygen free radical scavengers in salvaging erectile function in diabetic patients. PMID- 15292118 TI - Identification of a key step in the biosynthetic pathway of bacteriochlorophyll c and its implications for other known and unknown green sulfur bacteria. PMID- 15292120 TI - Phosphate control of the biosynthesis of antibiotics and other secondary metabolites is mediated by the PhoR-PhoP system: an unfinished story. PMID- 15292119 TI - Biphenyl dioxygenases: functional versatilities and directed evolution. PMID- 15292121 TI - An active type IV secretion system encoded by the F plasmid sensitizes Escherichia coli to bile salts. AB - F(+) strains of Escherichia coli infected with donor-specific bacteriophage such as M13 are sensitive to bile salts. We show here that this sensitivity has two components. The first derives from secretion of bacteriophage particles through the cell envelope, but the second can be attributed to expression of the F genes required for the formation of conjugative (F) pili. The latter component was manifested as reduced or no growth of an F(+) strain in liquid medium containing bile salts at concentrations that had little or no effect on the isogenic F(-) strain or as a reduced plating efficiency of the F(+) strain on solid media; at 2% bile salts, plating efficiency was reduced 10(4)-fold. Strains with F or F like R factors were consistently more sensitive to bile salts than isogenic, plasmid-free strains, but the quantitative effect of bile salts depended on both the plasmid and the strain. Sensitivity also depended on the bile salt, with conjugated bile salts (glycocholate and taurocholate) being less active than unconjugated bile salts (deoxycholate and cholate). F(+) cells were also more sensitive to sodium dodecyl sulfate than otherwise isogenic F(-) cells, suggesting a selectivity for amphipathic anions. A mutation in any but one F tra gene required for the assembly of F pili, including the traA gene encoding F pilin, substantially restored bile salt resistance, suggesting that bile salt sensitivity requires an active system for F pilin secretion. The exception was traW. A traW mutant was 100-fold more sensitive to cholate than the tra(+) strain but only marginally more sensitive to taurocholate or glycocholate. Bile salt sensitivity could not be attributed to a generalized change in the surface permeability of F(+) cells, as judged by the effects of hydrophilic and hydrophobic antibiotics and by leakage of periplasmic beta-lactamase into the medium. PMID- 15292122 TI - Genetic manipulation of carotenoid biosynthesis in the green sulfur bacterium Chlorobium tepidum. AB - The green sulfur bacterium Chlorobium tepidum is a strict anaerobe and an obligate photoautotroph. On the basis of sequence similarity with known enzymes or sequence motifs, nine open reading frames encoding putative enzymes of carotenoid biosynthesis were identified in the genome sequence of C. tepidum, and all nine genes were inactivated. Analysis of the carotenoid composition in the resulting mutants allowed the genes encoding the following six enzymes to be identified: phytoene synthase (crtB/CT1386), phytoene desaturase (crtP/CT0807), zeta-carotene desaturase (crtQ/CT1414), gamma-carotene desaturase (crtU/CT0323), carotenoid 1',2'-hydratase (crtC/CT0301), and carotenoid cis-trans isomerase (crtH/CT0649). Three mutants (CT0180, CT1357, and CT1416 mutants) did not exhibit a discernible phenotype. The carotenoid biosynthetic pathway in C. tepidum is similar to that in cyanobacteria and plants by converting phytoene into lycopene using two plant-like desaturases (CrtP and CrtQ) and a plant-like cis-trans isomerase (CrtH) and thus differs from the pathway known in all other bacteria. In contrast to the situation in cyanobacteria and plants, the construction of a crtB mutant completely lacking carotenoids demonstrates that carotenoids are not essential for photosynthetic growth of green sulfur bacteria. However, the bacteriochlorophyll a contents of mutants lacking colored carotenoids (crtB, crtP, and crtQ mutants) were decreased from that of the wild type, and these mutants exhibited a significant growth rate defect under all light intensities tested. Therefore, colored carotenoids may have both structural and photoprotection roles in green sulfur bacteria. The ability to manipulate the carotenoid composition so dramatically in C. tepidum offers excellent possibilities for studying the roles of carotenoids in the light-harvesting chlorosome antenna and iron-sulfur-type (photosystem I-like) reaction center. The phylogeny of carotenogenic enzymes in green sulfur bacteria and green filamentous bacteria is also discussed. PMID- 15292123 TI - The CcpA protein is necessary for efficient sporulation and enterotoxin gene (cpe) regulation in Clostridium perfringens. AB - Clostridium perfringens is the cause of several human diseases, including gas gangrene (clostridial myonecrosis), enteritis necroticans, antibiotic-associated diarrhea, and acute food poisoning. The symptoms of antibiotic-associated diarrhea and acute food poisoning are due to sporulation-dependent production of C. perfringens enterotoxin encoded by the cpe gene. Glucose is a catabolite repressor of sporulation by C. perfringens. In order to identify the mechanism of catabolite repression by glucose, a mutation was introduced into the ccpA gene of C. perfringens by conjugational transfer of a nonreplicating plasmid into C. perfringens, which led to inactivation of the ccpA gene by homologous recombination. CcpA is a transcriptional regulator known to mediate catabolite repression in a number of low-G+C-content gram-positive bacteria, of which C. perfringens is a member. The ccpA mutant strain sporulated at a 60-fold lower efficiency than the wild-type strain in the absence of glucose. In the presence of 5 mM glucose, sporulation was repressed about 2,000-fold in the wild-type strain and 800-fold in the ccpA mutant strain compared to sporulation levels for the same strains grown in the absence of glucose. Therefore, while CcpA is necessary for efficient sporulation in C. perfringens, glucose-mediated catabolite repression of sporulation is not due to the activity of CcpA. Transcription of the cpe gene was measured in the wild-type and ccpA mutant strains grown in sporulation medium by using a cpe-gusA fusion (gusA is an Escherichia coli gene encoding the enzyme beta-glucuronidase). In the exponential growth phase, cpe transcription was two times higher in the ccpA mutant strain than in the wild-type strain. Transcription of cpe was highly induced during the entry into stationary phase in wild-type cells but was not induced in the ccpA mutant strain. Glucose repressed cpe transcription in both the wild-type and ccpA mutant strain. Therefore, CcpA appears to act as a repressor of cpe transcription in exponential growth but is required for efficient sporulation and cpe transcription upon entry into stationary phase. CcpA was also required for maximum synthesis of collagenase (kappa toxin) and acted as a repressor of polysaccharide capsule synthesis in the presence of glucose, but it did not regulate synthesis of the phospholipase PLC (alpha toxin). PMID- 15292124 TI - Differences in enzymatic properties allow SodCI but not SodCII to contribute to virulence in Salmonella enterica serovar Typhimurium strain 14028. AB - Salmonella enterica serovar Typhimurium produces two Cu/Zn cofactored periplasmic superoxide dismutases, SodCI and SodCII. While mutations in sodCI attenuate virulence eightfold, loss of SodCII does not confer a virulence phenotype, nor does it enhance the defect observed in a sodCI background. Despite this in vivo phenotype, SodCI and SodCII are expressed at similar levels in vitro during the stationary phase of growth. By exchanging the open reading frames of sodCI and sodCII, we found that SodCI contributes to virulence when placed under the control of the sodCII promoter. In contrast, SodCII does not contribute to virulence even when expressed from the sodCI promoter. Thus, the disparity in virulence phenotypes is due primarily to some physical difference between the two enzymes. In an attempt to identify the unique property of SodCI, we have tested factors that might affect enzyme activity inside a phagosome. We found no significant difference between SodCI and SodCII in their resistance to acid, resistance to hydrogen peroxide, or ability to obtain copper in a copper-limiting environment. Both enzymes are synthesized as apoenzymes in the absence of copper and can be fully remetallated when copper is added. The one striking difference that we noted is that, whereas SodCII is released normally by an osmotic shock, SodCI is "tethered" within the periplasm by an apparently noncovalent interaction. We propose that this novel property of SodCI is crucial to its ability to contribute to virulence in serovar Typhimurium. PMID- 15292125 TI - Comparative genetic diversity of Pseudomonas stutzeri genomovars, clonal structure, and phylogeny of the species. AB - A combined phylogenetic and multilocus DNA sequence analysis of 26 Pseudomonas stutzeri strains distributed within the 9 genomovars of the species has been performed. Type strains of the two most closely related species (P. balearica, former genomovar 6, and P. mendocina), together with P. aeruginosa, as the type species of the genus, have been included in the study. The extremely high genetic diversity and the clonal structure of the species were confirmed by the sequence analysis. Clustering of strains in the consensus phylogeny inferred from the analysis of seven nucleotide sequences (16S ribosomal DNA, internally transcribed spacer region 1, gyrB, rpoD, nosZ, catA, and nahH) confirmed the monophyletic origin of the genomovars within the Pseudomonas branch and is in good agreement with earlier DNA-DNA similarity analysis, indicating that the selected genes are representative of the whole genome in members of the species. PMID- 15292126 TI - The Escherichia coli GTPase CgtAE cofractionates with the 50S ribosomal subunit and interacts with SpoT, a ppGpp synthetase/hydrolase. AB - CgtA(E)/Obg(E)/YhbZ is an Escherichia coli guanine nucleotide binding protein of the Obg/GTP1 subfamily whose members have been implicated in a number of cellular functions including GTP-GDP sensing, sporulation initiation, and translation. Here we describe a kinetic analysis of CgtA(E) with guanine nucleotides and show that its properties are similar to those of the Caulobacter crescentus homolog CgtA(C). CgtA(E) binds both GTP and GDP with moderate affinity, shows high guanine nucleotide exchange rate constants for both nucleotides, and has a relatively low GTP hydrolysis rate. We show that CgtA(E) is associated predominantly with the 50S ribosomal subunit. Interestingly, CgtA(E) copurifies with SpoT, a ribosome-associated ppGpp hydrolase/synthetase involved in the stress response. The interaction between CgtA(E) and SpoT was confirmed by reciprocal coprecipitation experiments and by two-hybrid assays. These studies raise the possibility that the ribosome-associated CgtA(E) is involved in the SpoT-mediated stress response. PMID- 15292127 TI - Control of virulence by the two-component system CiaR/H is mediated via HtrA, a major virulence factor of Streptococcus pneumoniae. AB - The CiaR/H two-component system is involved in regulating virulence and competence in Streptococcus pneumoniae. The system is known to regulate many genes, including that for high-temperature requirement A (HtrA). This gene has been implicated in the ability of the pneumococcus to colonize the nasopharynx of infant rats. We reported previously that deletion of the gene for HtrA made the pneumococcal strains much less virulent in mouse models, less able to grow at higher temperatures, and more sensitive to oxidative stress. In this report, we show that the growth phenotype as well as sensitivity to oxidative stress of Delta ciaR mutant was very similar to that of a Delta htrA mutant and that the expression of the HtrA protein was reduced in a ciaR-null mutant. Both the in vitro phenotype and the reduced virulence of Delta ciaR mutant could be restored by increasing the expression of HtrA. PMID- 15292128 TI - Identification of sarV (SA2062), a new transcriptional regulator, is repressed by SarA and MgrA (SA0641) and involved in the regulation of autolysis in Staphylococcus aureus. AB - The expression of genes involved in the pathogenesis of Staphylococcus aureus is known to be controlled by global regulatory loci, including agr, sarA, sae, arlRS, lytSR, and sarA-like genes. Here we described a novel transcriptional regulator called sarV of the SarA protein family. The transcription of sarV is low or undetectable under in vitro conditions but is significantly augmented in sarA and mgrA (norR or rat) (SA0641) mutants. The sarA and mgrA genes act as repressors of sarV expression, as confirmed by transcriptional fusion and Northern analysis data. Purified SarA and MgrA proteins bound specifically to separate regions of the sarV promoter as determined by gel shift and DNase I footprinting assays. The expression of 19 potential target genes involved in autolysis and virulence, phenotypes affected by sarA and mgrA, was evaluated in an isogenic sarV mutant pair. Our data indicated that the sarV gene product played a role regulating some virulence genes and more genes involved in autolysis. The sarV mutant was more resistant to Triton X-100 and penicillin induced lysis compared to the wild type and the sarA mutant, whereas hyperexpression of sarV in the parental strain or the sarV mutant rendered the resultant strain highly susceptible to lysis. Zymographic analysis of murein hydrolase activity revealed that inactivation of the sarV gene results in decreased extracellular murein hydrolase activity compared to that of wild-type S. aureus. We propose that sarV may be part of the common pathway by which mgrA and sarA gene products control autolysis in S. aureus. PMID- 15292129 TI - Interaction of PomB with the third transmembrane segment of PomA in the Na+ driven polar flagellum of Vibrio alginolyticus. AB - The marine bacterium Vibrio alginolyticus has four motor components, PomA, PomB, MotX, and MotY, responsible for its Na(+)-driven flagellar rotation. PomA and PomB are integral inner membrane proteins having four and one transmembrane segments (TMs), respectively, which are thought to form an ion channel complex. First, site-directed Cys mutagenesis was systematically performed from Asp-24 to Glu-41 of PomB, and the resulting mutant proteins were examined for susceptibility to a sulfhydryl reagent. Secondly, the Cys substitutions at the periplasmic boundaries of the PomB TM (Ser-38) and PomA TMs (Gly-23, Ser-34, Asp 170, and Ala-178) were combined. Cross-linked products were detected for the combination of PomB-S38C and PomA-D170C mutant proteins. The Cys substitutions in the periplasmic boundaries of PomA TM3 (from Met-169 to Asp-171) and the PomB TM (from Leu-37 to Ser-40) were combined to construct a series of double mutants. Most double mutations reduced the motility, whereas each single Cys substitution slightly affected it. Although the motility of the strain carrying PomA-D170C and PomB-S38C was significantly inhibited, it was recovered by reducing reagent. The strain with this combination showed a lower affinity for Na(+) than the wild-type combination. PomA-D148C and PomB-P16C, which are located at the cytoplasmic boundaries of PomA TM3 and the PomB TM, also formed the cross-linked product. From these lines of evidence, we infer that TM3 of PomA and the TM of PomB are in close proximity over their entire length and that cooperation between these two TMs is required for coupling of Na(+) conduction to flagellar rotation. PMID- 15292130 TI - Biochemical characterization of StyAB from Pseudomonas sp. strain VLB120 as a two component flavin-diffusible monooxygenase. AB - Pseudomonas sp. VLB120 uses styrene as a sole source of carbon and energy. The first step in this metabolic pathway is catalyzed by an oxygenase (StyA) and a NADH-flavin oxidoreductase (StyB). Both components have been isolated from wild type Pseudomonas strain VLB120 as well as from recombinant Escherichia coli. StyA from both sources is a dimer, with a subunit size of 47 kDa, and catalyzes the enantioselective epoxidation of CC double bonds. Styrene is exclusively converted to S-styrene oxide with a specific activity of 2.1 U mg(-1) (k(cat) = 1.6 s(-1)) and K(m) values for styrene of 0.45 +/- 0.05 mM (wild type) and 0.38 +/- 0.09 mM (recombinant). The epoxidation reaction depends on the presence of a NADH-flavin adenine dinucleotide (NADH-FAD) oxidoreductase for the supply of reduced FAD. StyB is a dimer with a molecular mass of 18 kDa and a NADH oxidation activity of 200 U mg(-1) (k(cat) [NADH] = 60 s(-1)). Steady-state kinetics determined for StyB indicate a mechanism of sequential binding of NADH and flavin to StyB. This enzyme reduces FAD as well as flavin mononucleotide and riboflavin. The NADH oxidation activity does not depend on the presence of StyA. During the epoxidation reaction, no formation of a complex of StyA and StyB has been observed, suggesting that electron transport between reductase and oxygenase occurs via a diffusing flavin. PMID- 15292131 TI - Defined inactive FecA derivatives mutated in functional domains of the outer membrane transport and signaling protein of Escherichia coli K-12. AB - The FecA outer membrane protein of Escherichia coli functions as a transporter of ferric citrate and as a signal receiver and signal transmitter for transcription initiation of the fec transport genes. Three FecA regions for which functional roles have been predicted from the crystal structures were mutagenized: (i) loops 7 and 8, which move upon binding of ferric citrate and close the entrance to the ferric citrate binding site; (ii) the dinuclear ferric citrate binding site; and (iii) the interface between the globular domain and the beta-barrel. Deletion of loops 7 and 8 abolished FecA transport and induction activities. Deletion of loops 3 and 11 also inactivated FecA, whereas deletion of loops 9 and 10 largely retained FecA activities. The replacement of arginine residue R365 or R380 and glutamine Q570, which are predicted to serve as binding sites for the negatively charged dinuclear ferric citrate, with alanine resulted in inactive FecA, whereas the binding site mutant R438A retained approximately 50% of the FecA induction and transport activities. Residues R150, E541, and E587, conserved among energy coupled outer membrane transporters, are predicted to form salt bridges between the globular domain and the beta-barrel and to contribute to the fixation of the globular domain inside the beta-barrel. Mutations E541A and E541R affected FecA induction and transport activity slightly, whereas mutations E587A and E587R more strongly reduced FecA activity. The double mutations R150A E541R and R150A E587R nearly abolished FecA activity. Apparently, the salt bridges are less important than the individual functions these residues seem to have for FecA activity. Comparison of the properties of the FecA, FhuA, FepA, and BtuB transporters indicates that although they have very similar crystal structures, the details of their functional mechanisms differ. PMID- 15292132 TI - Molecular analysis of cytolysin A (ClyA) in pathogenic Escherichia coli strains. AB - Cytolysin A (ClyA) of Escherichia coli is a pore-forming hemolytic protein encoded by the clyA (hlyE, sheA) gene that was first identified in E. coli K-12. In this study we examined various clinical E. coli isolates with regard to the presence and integrity of clyA. PCR and DNA sequence analyses demonstrated that 19 of 23 tested Shiga toxin-producing E. coli (STEC) strains, all 7 tested enteroinvasive E. coli (EIEC) strains, 6 of 8 enteroaggregative E. coli (EAEC) strains, and 4 of 7 tested enterotoxigenic E. coli (ETEC) strains possess a complete clyA gene. The remaining STEC, EAEC, and ETEC strains and 9 of the 17 tested enteropathogenic E. coli (EPEC) strains were shown to harbor mutant clyA derivatives containing 1-bp frameshift mutations that cause premature termination of the coding sequence. The other eight EPEC strains and all tested uropathogenic and new-born meningitis-associated E. coli strains (n = 14 and 3, respectively) carried only nonfunctional clyA fragments due to the deletion of two sequences of 493 bp and 204 or 217 bp at the clyA locus. Expression of clyA from clinical E. coli isolates proved to be positively controlled by the transcriptional regulator SlyA. Several tested E. coli strains harboring a functional clyA gene produced basal amounts of ClyA when grown under standard laboratory conditions, but most of them showed a clyA-dependent hemolytic phenotype only when SlyA was overexpressed. The presented data indicate that cytolysin A can play a role only for some of the pathogenic E. coli strains. PMID- 15292133 TI - The usher N terminus is the initial targeting site for chaperone-subunit complexes and participates in subsequent pilus biogenesis events. AB - Pilus biogenesis on the surface of uropathogenic Escherichia coli requires the chaperone/usher pathway, a terminal branch of the general secretory pathway. In this pathway, periplasmic chaperone-subunit complexes target an outer membrane (OM) usher for subunit assembly into pili and secretion to the cell surface. The molecular mechanisms of protein secretion across the OM are not well understood. Mutagenesis of the P pilus usher PapC and the type 1 pilus usher FimD was undertaken to elucidate the initial stages of pilus biogenesis at the OM. Deletion of residues 2 to 11 of the mature PapC N terminus abolished the targeting of the usher by chaperone-subunit complexes and rendered PapC nonfunctional for pilus biogenesis. Similarly, an intact FimD N terminus was required for chaperone-subunit binding and pilus biogenesis. Analysis of PapC FimD chimeras and N-terminal fragments of PapC localized the chaperone-subunit targeting domain to the first 124 residues of PapC. Single alanine substitution mutations were made in this domain that blocked pilus biogenesis but did not affect targeting of chaperone-subunit complexes. Thus, the usher N terminus does not function simply as a static binding site for chaperone-subunit complexes but also participates in subsequent pilus assembly events. PMID- 15292134 TI - Role of an inducible single-domain hemoglobin in mediating resistance to nitric oxide and nitrosative stress in Campylobacter jejuni and Campylobacter coli. AB - Campylobacter jejuni expresses two hemoglobins, each of which exhibits a heme pocket and structural signatures in common with vertebrate and plant globins. One of these, designated Cgb, is homologous to Vgb from Vitreoscilla stercoraria and does not possess the reductase domain seen in the flavohemoglobins. A Cgb deficient mutant of C. jejuni was hypersensitive to nitrosating agents (S nitrosoglutathione [GSNO] or sodium nitroprusside) and a nitric oxide-releasing compound (spermine NONOate). The sensitivity of the Cgb-deficient mutant to methyl viologen, hydrogen peroxide, and organic peroxides, however, was the same as for the wild type. Consistent with the protective role of Cgb against NO related stress, cgb expression was minimal in standard laboratory media but strongly and specifically induced after exposure to nitrosative stress. In contrast, the expression of Cgb was independent of aeration and the presence of superoxide. In the absence of preinduction by exposure to nitrosative stress, no difference was seen in the degree of respiratory inhibition by NO or the half life of the NO signal when cells of the wild type and the cgb mutant were compared. However, cells expressing GSNO-upregulated levels of Cgb exhibited robust NO consumption and respiration that was relatively NO insensitive compared to the respiration of the cgb mutant. Based on similar studies in Campylobacter coli, we also propose an identical role for Cgb in this closely related species. We conclude that, unlike the archetypal single-domain globin Vgb, Cgb forms a specific and inducible defense against NO and nitrosating agents. PMID- 15292135 TI - Penetration of membrane-containing double-stranded-DNA bacteriophage PM2 into Pseudoalteromonas hosts. AB - The icosahedral bacteriophage PM2 has a circular double-stranded DNA (dsDNA) genome and an internal lipid membrane. It is the only representative of the Corticoviridae family. How the circular supercoiled genome residing inside the viral membrane is translocated into the gram-negative marine Pseudoalteromonas host has been an intriguing question. Here we demonstrate that after binding of the virus to an abundant cell surface receptor, the protein coat is most probably dissociated. During the infection process, the host cell outer membrane becomes transiently permeable to lipophilic gramicidin D molecules proposing fusion with the viral membrane. One of the components of the internal viral lipid core particle is the integral membrane protein P7, with muralytic activity that apparently aids the process of peptidoglycan penetration. Entry of the virion also causes a limited depolarization of the cytoplasmic membrane. These phenomena differ considerably from those observed in the entry process of bacteriophage PRD1, a dsDNA virus, which uses its internal membrane to make a cell envelope penetrating tubular structure. PMID- 15292137 TI - Role of the pilot protein YscW in the biogenesis of the YscC secretin in Yersinia enterocolitica. AB - The YscC secretin is a major component of the type III protein secretion system of Yersinia enterocolitica and forms an oligomeric structure in the outer membrane. In a mutant lacking the outer membrane lipoprotein YscW, secretion is strongly reduced, and it has been proposed that YscW plays a role in the biogenesis of the secretin. To study the interaction between the secretin and this putative pilot protein, YscC and YscW were produced in trans in a Y. enterocolitica strain lacking all other components of the secretion machinery. YscW expression increased the yield of oligomeric YscC and was required for its outer membrane localization, confirming the function of YscW as a pilot protein. Whereas the pilot-binding site of other members of the secretin family has been identified in the C terminus, a truncated YscC derivative lacking the C-terminal 96 amino acid residues was functional and stabilized by YscW. Pulse-chase experiments revealed that approximately 30 min were required before YscC oligomerization was completed. In the absence of YscW, oligomerization was delayed and the yield of YscC oligomers was strongly reduced. An unlipidated form of the YscW protein was not functional, although it still interacted with the secretin and caused mislocalization of YscC even in the presence of wild-type YscW. Hence, YscW interacts with the unassembled YscC protein and facilitates efficient oligomerization, likely at the outer membrane. PMID- 15292136 TI - Differential and cross-transcriptional control of duplicated genes encoding alternative sigma factors in Streptomyces ambofaciens. AB - The duplicated hasR and hasL genes of Streptomyces ambofaciens encode alternative sigma factors (named sigma(B(R)) and sigma(B(L))) belonging to the sigma(B) general stress response family in Bacillus subtilis. The duplication appears to be the result of a recent event that occurred specifically in S. ambofaciens. The two genes are 98% identical, and their deduced protein products exhibit 97% identity at the amino acid level. In contrast with the coding sequences, their genetic environments and their transcriptional control are strongly divergent. While hasL is monocistronic, hasR is arranged in a polycistronic unit with two upstream open reading frames, arsR and prsR, that encode putative anti-anti-sigma and anti-sigma factors, respectively. Transcription of each has gene is initiated from two promoters. In each case, one promoter was shown to be developmentally controlled and to be similar to those recognized by the B. subtilis general stress response sigma factor sigma(B). Expression from this type of promoter for each of the has genes dramatically increases during the course of growth in liquid or on solid media and following oxidative and osmotic stresses. Reverse transcription-PCR measurements indicate that hasR is 100 times more strongly expressed than hasL from the sigma(B)-like promoter. Transcription from the second promoter of each gene (located upstream of arsR in the case of the hasR locus) appears to be constitutive and weak. Quantitative transcriptional analysis in single and double has mutant strains revealed that sigma(B(R)) and sigma(B(L)) direct their own transcription as well as that of their duplicates. Only a slight sensitivity in response to oxidative conditions could be assigned to either single or double mutants, revealing the probable redundancy of the sigma factors implied in stress response in Streptomyces. PMID- 15292138 TI - Bacillus subtilis YhcR, a high-molecular-weight, nonspecific endonuclease with a unique domain structure. AB - In a continuing effort to identify ribonucleases that may be involved in mRNA decay in Bacillus subtilis, fractionation of a protein extract from a triple mutant strain that was missing three previously characterized 3'-to-5' exoribonucleases (polynucleotide phosphorylase [PNPase], RNase R, and YhaM) was undertaken. These experiments revealed the presence of a high-molecular-weight nuclease encoded by the yhcR gene that was active in the presence of Ca(2+) and Mn(2+). YhcR is a sugar-nonspecific nuclease that cleaves endonucleolytically to yield nucleotide 3'-monophosphate products, similar to the well-characterized micrococcal nuclease of Staphylococcus aureus. YhcR appears to be located principally in the cell wall and is likely to be a substrate for a B. subtilis sortase. Zymogram analysis suggests that YhcR is the major Ca(2+)-activated nuclease of B. subtilis. In addition to having a unique overall domain structure, YhcR contains a hitherto unknown structural domain that we have named "NYD," for "new YhcR domain." PMID- 15292139 TI - Indole-3-acetic acid biosynthesis is deficient in Gluconacetobacter diazotrophicus strains with mutations in cytochrome c biogenesis genes. AB - Gluconacetobacter diazotrophicus is an endophyte of sugarcane frequently found in plants grown in agricultural areas where nitrogen fertilizer input is low. Recent results from this laboratory, using mutant strains of G. diazotrophicus unable to fix nitrogen, suggested that there are two beneficial effects of G. diazotrophicus on sugarcane growth: one dependent and one not dependent on nitrogen fixation. A plant growth-promoting substance, such as indole-3-acetic acid (IAA), known to be produced by G. diazotrophicus, could be a nitrogen fixation-independent factor. One strain, MAd10, isolated by screening a library of Tn5 mutants, released only approximately 6% of the amount of IAA excreted by the parent strain in liquid culture. The mutation causing the IAA(-) phenotype was not linked to Tn5. A pLAFR3 cosmid clone that complemented the IAA deficiency was isolated. Sequence analysis of a complementing subclone indicated the presence of genes involved in cytochrome c biogenesis (ccm, for cytochrome c maturation). The G. diazotrophicus ccm operon was sequenced; the individual ccm gene products were 37 to 52% identical to ccm gene products of Escherichia coli and equivalent cyc genes of Bradyrhizobium japonicum. Although several ccm mutant phenotypes have been described in the literature, there are no reports of ccm gene products being involved in IAA production. Spectral analysis, heme associated peroxidase activities, and respiratory activities of the cell membranes revealed that the ccm genes of G. diazotrophicus are involved in cytochrome c biogenesis. PMID- 15292140 TI - Kinetic analysis of tRNA-directed transcription antitermination of the Bacillus subtilis glyQS gene in vitro. AB - Binding of uncharged tRNA to the nascent transcript promotes readthrough of a leader region transcription termination signal in genes regulated by the T box transcription antitermination mechanism. Each gene in the T box family responds independently to its cognate tRNA, with specificity determined by base pairing of the tRNA to the leader at the anticodon and acceptor ends of the tRNA. tRNA binding stabilizes an antiterminator element in the transcript that sequesters sequences that participate in formation of the terminator helix. tRNA(Gly) dependent antitermination of the Bacillus subtilis glyQS leader was previously demonstrated in a purified in vitro assay system. This assay system was used to investigate the kinetics of transcription through the glyQS leader and the effect of tRNA and transcription elongation factors NusA and NusG on transcriptional pausing and antitermination. Several pause sites, including a major site in the loop of stem III of the leader, were identified, and the effect of modulation of pausing on antitermination efficiency was analyzed. We found that addition of tRNA(Gly) can promote antitermination as long as the tRNA is added before the majority of the transcription complexes reach the termination site, and variations in pausing affect the requirements for timing of tRNA addition. PMID- 15292141 TI - Isoleucine biosynthesis in Leptospira interrogans serotype lai strain 56601 proceeds via a threonine-independent pathway. AB - Three leuA-like protein-coding sequences were identified in Leptospira interrogans. One of these, the cimA gene, was shown to encode citramalate synthase (EC 4.1.3.-). The other two encoded alpha-isopropylmalate synthase (EC 4.1.3.12). Expressed in Escherichia coli, the citramalate synthase was purified and characterized. Although its activity was relatively low, it was strictly specific for pyruvate as the keto acid substrate. Unlike the citramalate synthase of the thermophile Methanococcus jannaschii, the L. interrogans enzyme is temperature sensitive but exhibits a much lower K(m) (0.04 mM) for pyruvate. The reaction product was characterized as (R)-citramalate, and the proposed beta methyl-d-malate pathway was further confirmed by demonstrating that citraconate was the substrate for the following reaction. This alternative pathway for isoleucine biosynthesis from pyruvate was analyzed both in vitro by assays of leptospiral isopropylmalate isomerase (EC 4.2.1.33) and beta-isopropylmalate dehydrogenase (EC 1.1.1.85) in E. coli extracts bearing the corresponding clones and in vivo by complementation of E. coli ilvA, leuC/D, and leuB mutants. Thus, the existence of a leucine-like pathway for isoleucine biosynthesis in L. interrogans under physiological conditions was unequivocally proven. Significant variations in either the enzymatic activities or mRNA levels of the cimA and leuA genes were detected in L. interrogans grown on minimal medium supplemented with different levels of the corresponding amino acids or in cells grown on serum containing rich medium. The similarity of this metabolic pathway in leptospires and archaea is consistent with the evolutionarily primitive status of the eubacterial spirochetes. PMID- 15292142 TI - Molecular characterization of the eis promoter of Mycobacterium tuberculosis. AB - To further understand Mycobacterium tuberculosis pathogenesis, the regulation of potential virulence genes needs to be investigated. The eis gene of M. tuberculosis H37Rv enhances the intracellular survival of Mycobacterium smegmatis, which does not contain eis, within macrophages (J. Wei, J. L. Dahl, J. W. Moulder, E. A. Roberts, P. O'Gaora, D. B. Young, and R. L. Friedman, J. Bacteriol. 182:377-384, 2000). Experiments were done to characterize the eis promoter in M. smegmatis and M. tuberculosis H37Ra. The putative -10 and -35 regions matched the Escherichia coli sigma(70) consensus 67 and 83%, respectively, making it a group A/SigA-like mycobacterial promoter. Expression of site-directed variants of the core promoter region, determined by flow cytometry using gfp as a reporter, showed that the putative -10 region is essential for eis expression. In addition, site-directed alteration of the eis promoter to the consensus E. coli sigma(70) promoter elements increased gfp transcription to levels similar to that driven by the heat shock promoter, phsp60, of Mycobacterium bovis BCG. Upstream promoter deletion analysis showed that a 200- and 412-bp region of the promoter was necessary for maximum expression of gfp in M. smegmatis and M. tuberculosis H37Ra, respectively. Random mutagenesis of the 412-bp eis promoter, using a catechol 2,3-dioxygenase screen and activity assay, defined nucleotides upstream of the core promoter region that are essential to eis expression in both M. smegmatis and M. tuberculosis H37Ra, including a region homologous to a DinR cis element. PMID- 15292143 TI - Differential regulation of soluble and membrane-bound inorganic pyrophosphatases in the photosynthetic bacterium Rhodospirillum rubrum provides insights into pyrophosphate-based stress bioenergetics. AB - Soluble and membrane-bound inorganic pyrophosphatases (sPPase and H(+)-PPase, respectively) of the purple nonsulfur bacterium Rhodospirillum rubrum are differentially regulated by environmental growth conditions. Both proteins and their transcripts were found in cells of anaerobic phototrophic batch cultures along all growth phases, although they displayed different time patterns. However, in aerobic cells that grow in the dark, which exhibited the highest growth rates, Northern and Western blot analyses as well as activity assays demonstrated high sPPase levels but no H(+)-PPase. It is noteworthy that H(+) PPase is highly expressed in aerobic cells under acute salt stress (1 M NaCl). H(+)-PPase was also present in anaerobic cells growing at reduced rates in the dark under either fermentative or anaerobic respiratory conditions. Since H(+) PPase was detected not only under all anaerobic growth conditions but also under salt stress in aerobiosis, the corresponding gene is not invariably repressed by oxygen. Primer extension analyses showed that, under all anaerobic conditions tested, the R. rubrum H(+)-PPase gene utilizes two activator-dependent tandem promoters, one with an FNR-like sequence motif and the other with a RegA motif, whereas in aerobiosis under salt stress, the H(+)-PPase gene is transcribed from two further tandem promoters involving other transcription factors. These results demonstrate a tight transcriptional regulation of the H(+)-PPase gene, which appears to be induced in response to a variety of environmental conditions, all of which constrain cell energetics. PMID- 15292144 TI - Requirements for nitric oxide generation from isoniazid activation in vitro and inhibition of mycobacterial respiration in vivo. AB - Isoniazid (INH), a front-line antituberculosis agent, is activated by mycobacterial catalase-peroxidase KatG, converting INH into bactericidal reactive species. Here we investigated the requirements and the pathway of nitric oxide (NO*) generation during oxidative activation of INH by Mycobacterium tuberculosis KatG in vitro. We also provide in vivo evidence that INH-derived NO* can inhibit key mycobacterial respiratory enzymes, which may contribute to the overall antimycobacterial action of INH. PMID- 15292145 TI - Analysis of the genome structure of the nonpathogenic probiotic Escherichia coli strain Nissle 1917. AB - Nonpathogenic Escherichia coli strain Nissle 1917 (O6:K5:H1) is used as a probiotic agent in medicine, mainly for the treatment of various gastroenterological diseases. To gain insight on the genetic level into its properties of colonization and commensalism, this strain's genome structure has been analyzed by three approaches: (i) sequence context screening of tRNA genes as a potential indication of chromosomal integration of horizontally acquired DNA, (ii) sequence analysis of 280 kb of genomic islands (GEIs) coding for important fitness factors, and (iii) comparison of Nissle 1917 genome content with that of other E. coli strains by DNA-DNA hybridization. PCR-based screening of 324 nonpathogenic and pathogenic E. coli isolates of different origins revealed that some chromosomal regions are frequently detectable in nonpathogenic E. coli and also among extraintestinal and intestinal pathogenic strains. Many known fitness factor determinants of strain Nissle 1917 are localized on four GEIs which have been partially sequenced and analyzed. Comparison of these data with the available knowledge of the genome structure of E. coli K-12 strain MG1655 and of uropathogenic E. coli O6 strains CFT073 and 536 revealed structural similarities on the genomic level, especially between the E. coli O6 strains. The lack of defined virulence factors (i.e., alpha-hemolysin, P-fimbrial adhesins, and the semirough lipopolysaccharide phenotype) combined with the expression of fitness factors such as microcins, different iron uptake systems, adhesins, and proteases, which may support its survival and successful colonization of the human gut, most likely contributes to the probiotic character of E. coli strain Nissle 1917. PMID- 15292146 TI - DNA microarray-based genome comparison of a pathogenic and a nonpathogenic strain of Xylella fastidiosa delineates genes important for bacterial virulence. AB - Xylella fastidiosa is a phytopathogenic bacterium that causes serious diseases in a wide range of economically important crops. Despite extensive comparative analyses of genome sequences of Xylella pathogenic strains from different plant hosts, nonpathogenic strains have not been studied. In this report, we show that X. fastidiosa strain J1a12, associated with citrus variegated chlorosis (CVC), is nonpathogenic when injected into citrus and tobacco plants. Furthermore, a DNA microarray-based comparison of J1a12 with 9a5c, a CVC strain that is highly pathogenic and had its genome completely sequenced, revealed that 14 coding sequences of strain 9a5c are absent or highly divergent in strain J1a12. Among them, we found an arginase and a fimbrial adhesin precursor of type III pilus, which were confirmed to be absent in the nonpathogenic strain by PCR and DNA sequencing. The absence of arginase can be correlated to the inability of J1a12 to multiply in host plants. This enzyme has been recently shown to act as a bacterial survival mechanism by down-regulating host nitric oxide production. The lack of the adhesin precursor gene is in accordance with the less aggregated phenotype observed for J1a12 cells growing in vitro. Thus, the absence of both genes can be associated with the failure of the J1a12 strain to establish and spread in citrus and tobacco plants. These results provide the first detailed comparison between a nonpathogenic strain and a pathogenic strain of X. fastidiosa, constituting an important step towards understanding the molecular basis of the disease. PMID- 15292147 TI - spoIVH (ykvV), a requisite cortex formation gene, is expressed in both sporulating compartments of Bacillus subtilis. AB - It is well known that the ykvU-ykvV operon is under the regulation of the sigma(E)-associated RNA polymerase (Esigma(E)). In our study, we observed that ykvV is transcribed together with the upstream ykvU gene by Esigma(E) in the mother cell and monocistronically under Esigma(G) control in the forespore. Interestingly, alternatively expressed ykvV in either the forespore or the mother cell increased the sporulation efficiency in the ykvV background. Studies show that the YkvV protein is a member of the thioredoxin superfamily and also contains a putative Sec-type secretion signal at the N terminus. We observed efficient sporulation in a mutant strain obtained by replacing the putative signal peptide of YkvV with the secretion signal sequence of SleB, indicating that the putative signal sequence is essential for spore formation. These results suggest that YkvV is capable of being transported by the putative Sec-type signal sequence into the space between the double membranes surrounding the forespore. The ability of ykvV expression in either compartment to complement is indeed intriguing and further introduces a new dimension to the genetics of B. subtilis spore formation. Furthermore, electron microscopic observation revealed a defective cortex in the ykvV disruptant. In addition, the expression levels of sigma(K)-directed genes significantly decreased despite normal sigma(G) activity in the ykvV mutant. However, immunoblotting with the anti-sigma(K) antibody showed that pro-sigma(K) was normally processed in the ykvV mutant, indicating that YkvV plays an important role in cortex formation, consistent with recent reports. We therefore propose that ykvV should be renamed spoIVH. PMID- 15292148 TI - The LuxR homolog ExpR, in combination with the Sin quorum sensing system, plays a central role in Sinorhizobium meliloti gene expression. AB - Quorum sensing, a population density-dependent mechanism for bacterial communication and gene regulation, plays a crucial role in the symbiosis between alfalfa and its symbiont Sinorhizobium meliloti. The Sin system, one of three quorum sensing systems present in S. meliloti, controls the production of the symbiotically active exopolysaccharide EPS II. Based on DNA microarray data, the Sin system also seems to regulate a multitude of S. meliloti genes, including genes that participate in low-molecular-weight succinoglycan production, motility, and chemotaxis, as well as other cellular processes. Most of the regulation by the Sin system is dependent on the presence of the ExpR regulator, a LuxR homolog. Gene expression profiling data indicate that ExpR participates in additional cellular processes that include nitrogen fixation, metabolism, and metal transport. Based on our microarray analysis we propose a model for the regulation of gene expression by the Sin/ExpR quorum sensing system and another possible quorum sensing system(s) in S. meliloti. PMID- 15292149 TI - Comparative whole-genome analysis of virulent and avirulent strains of Porphyromonas gingivalis. AB - We used Porphyromonas gingivalis gene microarrays to compare the total gene contents of the virulent strain W83 and the avirulent type strain, ATCC 33277. Signal ratios and scatter plots indicated that the chromosomes were very similar, with approximately 93% of the predicted genes in common, while at least 7% of them showed very low or no signals in ATCC 33277. Verification of the array results by PCR indicated that several of the disparate genes were either absent from or variant in ATCC 33277. Divergent features included already reported insertion sequences and ragB, as well as additional hypothetical and functionally assigned genes. Several of the latter were organized in a putative operon in W83 and encoded enzymes involved in capsular polysaccharide synthesis. Another cluster was associated with two paralogous regions of the chromosome with a low G+C content, at 41%, compared to that of the whole genome, at 48%. These regions also contained conserved and species-specific hypothetical genes, transposons, insertion sequences, and integrases and were located adjacent to tRNA genes; thus, they had several characteristics of pathogenicity islands. While this global comparative analysis showed the close relationship between W83 and ATCC 33277, the clustering of genes that are present in W83 but divergent in or absent from ATCC 33277 is suggestive of chromosomal islands that may have been acquired by lateral gene transfer. PMID- 15292150 TI - Tra proteins characteristic of F-like type IV secretion systems constitute an interaction group by yeast two-hybrid analysis. AB - Using yeast two-hybrid screens, we have defined an interaction group of six Tra proteins encoded by the F plasmid and required by F(+) cells to elaborate F pili. The six proteins are TraH, TraF, TraW, TraU, TrbI, and TrbB. Except for TrbI, these proteins were all identified as hallmarks of F-like type IV secretion systems (TFSSs), with no homologues among TFSS genes of P-type or I-type systems (T. Lawley, W. Klimke, M. Gubbins, and L. Frost, FEMS Microbiol. Lett. 224:1-15, 2003). Also with the exception of TrbI, which is an inner membrane protein, the remaining proteins are or are predicted to be periplasmic. TrbI consists of one membrane-spanning segment near its N terminus and an 88-residue, hydrophilic domain that extends into the periplasm. Hence, the proteins of this group probably form a periplasmic cluster in Escherichia coli. The interaction network identifies TraH as the most highly connected node, with two-hybrid links to TrbI, TraU, and TraF. As measured by transcriptional activation of lacZ, the TrbI-TraH interaction in Saccharomyces cerevisiae requires the TraH amino acid segment from residues 193 to 225. The TraU and TraF interactions are localized to C-terminal segments of TraH (amino acids 315 to 458 for TraF and amino acids 341 to 458 for TraU). The TrbI-TraH interaction with full-length (less the signal peptide) TraH is weak but increases 40-fold with N-terminal TraH deletions; the first 50 amino acids appear to be critical for inhibiting TrbI binding in yeast. Previous studies by others have shown that, with the exception of trbB mutations, which do not affect the elaboration or function of F pili under laboratory conditions, a mutation in any of the other genes in this interaction group alters the number or length distribution of F pili. We propose a model whereby one function of the TraH interaction group is to control F-pilus extension and retraction. PMID- 15292151 TI - Identification of the secretion and translocation domain of the enteropathogenic and enterohemorrhagic Escherichia coli effector Cif, using TEM-1 beta-lactamase as a new fluorescence-based reporter. AB - Enteropathogenic and enterohemorrhagic Escherichia coli (EPEC and EHEC) strains are human and animal pathogens that inject effector proteins into host cells via a type III secretion system (TTSS). Cif is an effector protein which induces host cell cycle arrest and reorganization of the actin cytoskeleton. Cif is encoded by a lambdoid prophage present in most of the EPEC and EHEC strains. In this study, we analyzed the domain that targets Cif to the TTSS by using a new reporter system based on a translational fusion of the effector proteins with mature TEM-1 beta-lactamase. Translocation was detected directly in living host cells by using the fluorescent beta-lactamase substrate CCF2/AM. We show that the first 16 amino acids (aa) of Cif were necessary and sufficient to mediate translocation into the host cells. Similarly, the first 20 aa of the effector proteins Map, EspF, and Tir, which are encoded in the same region as the TTSS, mediated secretion and translocation in a type III-dependent but chaperone-independent manner. A truncated form of Cif lacking its first 20 aa was no longer secreted and translocated, but fusion with the first 20 aa of Tir, Map, or EspF restored both secretion and translocation. In addition, the chimeric proteins were fully able to trigger host cell cycle arrest and stress fiber formation. In conclusion, our results demonstrate that Cif is composed of a C-terminal effector domain and an exchangeable N-terminal translocation signal and that the TEM-1 reporter system is a convenient tool for the study of the translocation of toxins or effector proteins into host cells. PMID- 15292152 TI - Multiple-locus variable-number tandem repeat analysis of Dutch Bordetella pertussis strains reveals rapid genetic changes with clonal expansion during the late 1990s. AB - Bordetella pertussis, the causative agent of whooping cough, has remained endemic in The Netherlands despite extensive nationwide vaccination since 1953. In the 1990s, several epidemic periods have resulted in many cases of pertussis. We have proposed that strain variation has played a major role in the upsurges of this disease in The Netherlands. Therefore, molecular characterization of strains is important in identifying the causes of pertussis epidemiology. For this reason, we have developed a multiple-locus variable-number tandem repeat analysis (MLVA) typing system for B. pertussis. By combining the MLVA profile with the allelic profile based on multiple-antigen sequence typing, we were able to further differentiate strains. The relationships between the various genotypes were visualized by constructing a minimum spanning tree. MLVA of Dutch strains of B. pertussis revealed that the genotypes of the strains isolated in the prevaccination period were diverse and clearly distinct from the strains isolated in the 1990s. Furthermore, there was a decrease in diversity in the strains from the late 1990s, with a remarkable clonal expansion that coincided with the epidemic periods. Using this genotyping, we have been able to show that B. pertussis is much more dynamic than expected. PMID- 15292153 TI - Escherichia coli serogroup O107/O117 lipopolysaccharide binds and neutralizes Shiga toxin 2. AB - The AB(5) toxin Shiga toxin 2 (Stx2) has been implicated as a major virulence factor of Escherichia coli O157:H7 and other Shiga toxin-producing E. coli strains in the progression of intestinal disease to more severe systemic complications. Here, we demonstrate that supernatant from a normal E. coli isolate, FI-29, neutralizes the effect of Stx2, but not the related Stx1, on Vero cells. Biochemical characterization of the neutralizing activity identified the lipopolysaccharide (LPS) of FI-29, a serogroup O107/O117 strain, as the toxin neutralizing component. LPSs from FI-29 as well as from type strains E. coli O107 and E. coli O117 were able bind Stx2 but not Stx1, indicating that the mechanism of toxin neutralization may involve inhibition of the interaction between Stx2 and the Gb(3) receptor on Vero cells. PMID- 15292154 TI - Novel archaeal alanine:glyoxylate aminotransferase from Thermococcus litoralis. AB - A novel alanine:glyoxylate aminotransferase was found in a hyperthermophilic archaeon, Thermococcus litoralis. The amino acid sequence of the enzyme did not show a similarity to any alanine:glyoxylate aminotransferases reported so far. Homologues of the enzyme appear to be present in almost all hyperthermophilic archaea whose whole genomes have been sequenced. PMID- 15292155 TI - Amino acid substitutions in putative selectivity filter regions III and IV in KdpA alter ion selectivity of the KdpFABC complex from Escherichia coli. AB - When grown under conditions of potassium limitation or high osmolality, Escherichia coli synthesizes the K(+)-translocating KdpFABC complex. The KdpA subunit, which has sequence homology to potassium channels of the KcsA type, has been shown to be important for potassium binding and transport. Replacement of the glycine residues in KdpA at positions 345 and 470, members of putative selectivity filter regions III and IV, alters the ion selectivity of the KdpFABC complex. PMID- 15292156 TI - Complete genomic nucleotide sequence of the temperate bacteriophage Aa Phi 23 of Actinobacillus actinomycetemcomitans. AB - The entire double-stranded DNA genome of the Actinobacillus actinomycetemcomitans bacteriophage Aa Phi 23 was sequenced. Linear DNA contained in the phage particles is circularly permuted and terminally redundant. Therefore, the physical map of the phage genome is circular. Its size is 43,033 bp with an overall molar G+C content of 42.5 mol%. Sixty-six potential open reading frames (ORFs) were identified, including an ORF resulting from a translational frameshift. A putative function could be assigned to 23 of them. Twenty-three other ORFs share homologies only with hypothetical proteins present in several bacteria or bacteriophages, and 20 ORFs seem to be specific for phage Aa Phi 23. The organization of the phage genome and several genetic functions share extensive similarities to that of the lambdoid phages. However, Aa Phi 23 encodes a DNA adenine methylase, and the DNA packaging strategy is more closely related to the P22 system. The attachment sites of Aa Phi 23 (attP) and several A. actinomycetemcomitans hosts (attB) are 49 bp long. PMID- 15292157 TI - Characterization of lipoteichoic acids as Lactobacillus delbrueckii phage receptor components. AB - Lipoteichoic acids (LTAs) were purified from Lactobacillus delbrueckii subsp. lactis ATCC 15808 and its LL-H adsorption-resistant mutant, Ads-5, by hydrophobic interaction chromatography. L. delbrueckii phages (LL-H, the LL-H host range mutant, and JCL1032) were inactivated by these poly(glycerophosphate) type of LTAs in vitro in accordance to their adsorption to intact ATCC 15808 and Ads-5 cells. PMID- 15292158 TI - Evidence of bar minigene expression and tRNA2Ile sequestration as peptidyl tRNA2Ile during lambda bacteriophage development. AB - Lambda bacteriophage development is impaired in Escherichia coli cells defective for peptidyl (pep)-tRNA hydrolase (Pth). Single-base-pair mutations (bar(-)) that affect translatable two-codon open reading frames named bar minigenes (barI or barII) in the lambda phage genome promote the development of this phage in Pth defective cells (rap cells). When the barI minigene is cloned and overexpressed from a plasmid, it inhibits protein synthesis and cell growth in rap cells by sequestering tRNA(2)(Ile) as pep-tRNA(2)(Ile). Either tRNA(2)(Ile) or Pth may reverse these effects. In this paper we present evidence that both barI and barII minigenes are translatable elements that sequester tRNA(2)(Ile) as pep tRNA(2)(Ile). In addition, overexpression of the barI minigene impairs the development even of bar(-) phages in rap cells. Interestingly, tRNA or Pth may reestablish lambda phage development. These results suggest that lambda bar minigenes are expressed and tRNA(2)(Ile) is sequestered as pep-tRNA(2)(Ile) during lambda phage development. PMID- 15292159 TI - Polymorphic mutation frequencies in Escherichia coli: emergence of weak mutators in clinical isolates. AB - Polymorphisms in the rifampin resistance mutation frequency (f) were studied in 696 Escherichia coli strains from Spain, Sweden, and Denmark. Of the 696 strains, 23% were weakly hypermutable (4 x 10(-8) < or = f < 4 x 10(-7)), and 0.7% were strongly hypermutable (f > or = 4 x 10(-7)). Weak mutators were apparently more frequent in southern Europe and in blood isolates (38%) than in urinary tract isolates (25%) and feces of healthy volunteers (11%). PMID- 15292160 TI - The RpoS sigma factor in the dissimilatory Fe(III)-reducing bacterium Geobacter sulfurreducens. AB - Geobacter sulfurreducens RpoS sigma factor was shown to contribute to survival in stationary phase and upon oxygen exposure. Furthermore, a mutation in rpoS decreased the rate of reduction of insoluble Fe(III) but not of soluble forms of iron. This study suggests that RpoS plays a role in regulating metabolism of Geobacter under suboptimal conditions in subsurface environments. PMID- 15292161 TI - Erwinia chrysanthemi O antigen is required for betaine osmoprotection in high salt media. AB - Cellular components necessary for osmoprotection are poorly known. In this study we show that O antigen is specifically required for the effectiveness of betaines as osmoprotectants for Erwinia chrysanthemi in saline media. The phenotype is correlated with the inability of rfb mutant strains to maintain a high accumulation level of betaines in hypersaline media. PMID- 15292162 TI - Excision of the Shigella resistance locus pathogenicity island in Shigella flexneri is stimulated by a member of a new subgroup of recombination directionality factors. AB - Pathogenicity islands are capable of excision and insertion within bacterial chromosomes. We describe a protein, Rox, that stimulates excision of the Shigella resistance locus pathogenicity island in Shigella flexneri. Sequence analysis suggests that Rox belongs to a new subfamily of recombination directionality factors, which includes proteins from P4, enterohemorrhagic Escherichia coli, and Yersinia pestis. PMID- 15292163 TI - Phosphodiesterase IV inhibition by piclamilast potentiates the cytodifferentiating action of retinoids in myeloid leukemia cells. Cross-talk between the cAMP and the retinoic acid signaling pathways. AB - Inhibition of phosphodiesterase IV by N-(3,5-dichloropyrid-4-yl)-3-cyclopentyloxy 4-methoxybenzamide (piclamilast) enhances the myeloid differentiation induced by all-trans-retinoic acid (ATRA), retinoic acid receptor alpha (RARalpha), or retinoic acid receptor X agonists in NB4 and other retinoid-sensitive myeloid leukemia cell types. ATRA-resistant NB4.R2 cells are also partially responsive to the action of piclamilast and retinoic acid receptor X agonists. Treatment of NB4 cells with piclamilast or ATRA results in activation of the cAMP signaling pathway and nuclear translocation of cAMP-dependent protein kinase. This causes a transitory increase in cAMP-responsive element-binding protein phosphorylation, which is followed by down-modulation of the system. ATRA + piclamilast have no additive effects on the modulation of the cAMP pathway, and the combination has complex effects on cAMP-regulated genes. Piclamilast potentiates the ligand dependent transactivation and degradation of RARalpha through a cAMP-dependent protein kinase-dependent phosphorylation. Enhanced transactivation is also observed in the case of PML-RARalpha. In NB4 cells, increased transactivation is likely to be at the basis of enhanced myeloid maturation and enhanced expression of many retinoid-dependent genes. Piclamilast and/or ATRA exert major effects on the expression of cEBP and STAT1, two types of transcription factors involved in myeloid maturation. Induction and activation of STAT1 correlates directly with enhanced cytodifferentiation. Finally, ERK and the cAMP target protein, Epac, do not participate in the maturation program activated by ATRA + piclamilast. Initial in vivo studies conducted in severe combined immunodeficiency mice transplanted with NB4 leukemia cells indicate that the enhancing effect of piclamilast on ATRA-induced myeloid maturation translates into a therapeutic benefit. PMID- 15292164 TI - Enhanced toxicity and cellular binding of a modified amyloid beta peptide with a methionine to valine substitution. AB - The amyloid beta peptide (Abeta) is toxic to neuronal cells, and it is probable that this toxicity is responsible for the progressive cognitive decline associated with Alzheimer's disease. However, the nature of the toxic Abeta species and its precise mechanism of action remain to be determined. It has been reported that the methionine residue at position 35 has a pivotal role to play in the toxicity of Abeta. We examined the effect of mutating the methionine to valine in Abeta42 (AbetaM35V). The neurotoxic activity of AbetaM35V on primary mouse neuronal cortical cells was enhanced, and this diminished cell viability occurred at an accelerated rate compared with Abeta42. AbetaM35V binds Cu2+ and produces similar amounts of H2O2 as Abeta42 in vitro, and the neurotoxic activity was attenuated by the H2O2 scavenger catalase. The increased toxicity of AbetaM35V was associated with increased binding of this mutated peptide to cortical cells. The M35V mutation altered the interaction between Abeta and copper in a lipid environment as shown by EPR analysis, which indicated that the valine substitution made the peptide less rigid in the bilayer region with a resulting higher affinity for the bilayer. Circular dichroism spectroscopy showed that both Abeta42 and AbetaM35V displayed a mixture of alpha-helical and beta sheet conformations. These findings provide further evidence that the toxicity of Abeta is regulated by binding to neuronal cells. PMID- 15292165 TI - Hairpin structure-forming propensity of the (CCTG.CAGG) tetranucleotide repeats contributes to the genetic instability associated with myotonic dystrophy type 2. AB - The genetic instabilities of (CCTG.CAGG)(n) tetranucleotide repeats were investigated to evaluate the molecular mechanisms responsible for the massive expansions found in myotonic dystrophy type 2 (DM2) patients. DM2 is caused by an expansion of the repeat from the normal allele of 26 to as many as 11,000 repeats. Genetic expansions and deletions were monitored in an African green monkey kidney cell culture system (COS-7 cells) as a function of the length (30, 114, or 200 repeats), orientation, or proximity of the repeat tracts to the origin (SV40) of replication. As found for CTG.CAG repeats related to DM1, the instabilities were greater for the longer tetranucleotide repeat tracts. Also, the expansions and deletions predominated when cloned in orientation II (CAGG on the leading strand template) rather than I and when cloned proximal rather than distal to the replication origin. Biochemical studies on synthetic d(CAGG)(26) and d(CCTG)(26) as models of unpaired regions of the replication fork revealed that d(CAGG)(26) has a marked propensity to adopt a defined base paired hairpin structure, whereas the complementary d(CCTG)(26) lacks this capacity. The effect of orientation described above differs from all previous results with three triplet repeat sequences (including CTG.CAG), which are also involved in the etiologies of other hereditary neurological diseases. However, similar to the triplet repeat sequences, the ability of one of the two strands to form a more stable folded structure, in our case the CAGG strand, explains this unorthodox "reversed" behavior. PMID- 15292166 TI - Binding of the Golgi sorting receptor muclin to pancreatic zymogens through sulfated O-linked oligosaccharides. AB - Sorting and packaging of regulated secretory proteins involves protein aggregation in the trans-Golgi network and secretory granules. In this work, we characterized the pH-dependent interactions of pancreatic acinar cell-regulated secretory proteins (zymogens) with Muclin, a putative Golgi cargo receptor. In solution, purified Muclin co-aggregated with isolated zymogens at mildly acidic pH. In an overlay assay, [35S]sulfate biosynthetically labeled Muclin bound directly at mildly acidic pH to the zymogen granule content proteins amylase, prolipase, pro-carboxypeptidase A1, pro-elastase II, chymotrypsinogen B, and Reg1. Denaturation of Muclin with reducing agents to break the numerous intrachain disulfide bonds in Muclin's scavenger receptor cysteine-rich and CUB domains did not interfere with binding. Non-sulfated [35S]Met/Cys-labeled Muclin showed decreased binding in the overlay assay. Extensive Pronase E digestion of unlabeled Muclin was used to produce glycopeptides, which competed for binding of [35S]sulfate-labeled Muclin to zymogens. The results demonstrate that the sulfated, O-glycosylated groups are responsible for the pH-dependent interactions of Muclin with the zymogens. The behavior of Muclin fulfils the requirement of a Golgi cargo receptor to bind to regulated secretory proteins under the mildly acidic pH conditions that exist in the trans-Golgi network. PMID- 15292167 TI - Amyloidogenicity and cytotoxicity of recombinant mature human islet amyloid polypeptide (rhIAPP). AB - Pancreatic amyloid plaques formed by the pancreatic islet amyloid polypeptide (IAPP) are present in more than 95% of type II diabetes mellitus patients, and their abundance correlates with the severity of the disease. IAPP is currently considered the most amyloidogenic peptide known, but the molecular bases of its aggregation are still incompletely understood. Detailed characterization of the mechanisms of amyloid formation requires large quantities of pure material. Thus, availability of recombinant IAPP in sufficient amounts for such studies constitutes an important step toward elucidation of the mechanisms of amyloidogenicity. Here, we report, for the first time, the successful expression, purification and characterization of the amyloidogenicity and cytotoxicity of recombinant human mature IAPP. This approach is likely to be useful for the production of other amyloidogenic peptides or proteins that are difficult to obtain by chemical synthesis. PMID- 15292168 TI - A unique region in bacteriophage t7 DNA polymerase important for exonucleolytic hydrolysis of DNA. AB - A crystal structure of the bacteriophage T7 gene 5 protein/Escherichia coli thioredoxin complex reveals a region in the exonuclease domain (residues 144-157) that is not present in other members of the E. coli DNA polymerase I family. To examine the role of this region, a genetically altered enzyme that lacked residues 144-157 (T7 polymerase (pol) Delta144-157) was purified and characterized biochemically. The polymerase activity and processivity of T7 pol Delta144-157 on primed M13 DNA are similar to that of wild-type T7 DNA polymerase implying that these residues are not important for DNA synthesis. The ability of T7 pol Delta144-157 to catalyze the hydrolysis of a phosphodiester bond, as judged from the rate of hydrolysis of a p-nitrophenyl ester of thymidine monophosphate, also remains unaffected. However, the 3'-5' exonuclease activity on polynucleotide substrates is drastically reduced; exonuclease activity on single-stranded DNA is 10-fold lower and that on double-stranded DNA is 20-fold lower as compared with wild-type T7 DNA polymerase. Taken together, our results suggest that residues 144-157 of gene 5 protein, although not crucial for polymerase activity, are important for DNA binding during hydrolysis of polynucleotides. PMID- 15292169 TI - Conformational changes of murine polyomavirus capsid proteins induced by sialic acid binding. AB - Murine polyomavirus (Py) infection initiates by the recognition of cell membrane molecules containing terminal sialic acid (SA) residues through specific binding pockets formed at the major capsid protein VP1 surface. VP1 Pockets 1, 2, and 3 bind terminal SA, Gal, and second branched SA, respectively. The consequence of recognition on viral cell entry remains elusive. In this work, we show that preincubation of Py with soluble compounds within Pocket 1 (N-acetyl or N glycolyl neuraminic acids) increases Py cell binding and infectivity in murine 3T6 fibroblasts. In contrast, Gal does not significantly alter Py binding nor infectivity, whereas sialyllactose, in Pockets 1 and 2, decreases cell binding and infectivity. Binding experiments with Py virus-like particles confirmed the direct involvement of VP1 in this effect. To determine whether such results could reflect VP1 conformational changes induced by SA binding, protease digestion assays were performed after pretreatment of Py or virus-like particles with soluble receptor fragments. Binding of SA with the VP1 Pocket 1, but not of compounds interacting with Pocket 2, was associated with a transition of this protein from a protease-sensitive to a protease-resistant state. This effect was transmitted to the minor capsid proteins VP2 and VP3 in virus particles. Attachment of Py to cell monolayers similarly led to a VP1 trypsin-resistant pattern. Taken together, these data present evidence that initial binding of Py to terminal SA induces conformational changes in the viral capsid, which may influence subsequent virus cell entry steps. PMID- 15292171 TI - Functional characterization of the yeast Ppz1 phosphatase inhibitory subunit Hal3: a mutagenesis study. AB - Saccharomyces cerevisiae Hal3 is a conserved protein that binds the carboxyl terminal catalytic domain of the PP1c (protein phosphatase 1)-related phosphatase Ppz1 and potently inhibits its activity, thus modulating all of the characterized functions so far of the phosphatase. It is unknown how Hal3 binds to Ppz1 and inhibits its activity. Although it contains a putative protein phosphatase 1c binding-like sequence (263KLHVLF268), mutagenesis analysis suggests that this motif is not required for Ppz1 binding and inhibition. The mutation of the conserved His378 (possibly involved in dehydrogenase catalytic activity) did not impair Hal3 functions or Ppz1 binding. Random mutagenesis of the 228 residue conserved central region of Hal3 followed by a loss-of-function screen allowed the identification of nine residues important for Ppz1-related Hal3 functions. Seven of these residues cluster in a relatively small region spanning from amino acid 446 to 480. Several mutations affected Ppz1 binding and inhibition in vitro, whereas changes in Glu460 and Val462 did not alter binding but resulted in Hal3 versions unable to inhibit the phosphatase. Therefore, there are independent Hal3 structural elements required for Ppz1 binding and inhibition. S. cerevisiae encodes a protein (Vhs3) structurally related to Hal3. Recent evidence suggests that both mutations are synthetically lethal. Surprisingly, versions of Hal3 carrying mutations that strongly affected Ppz1 binding or inhibitory capacity were able to complement lethality. In contrast, the mutation of His378 did not. This finding suggests that Hal3 may have both Ppz1-dependent and independent functions involving different structural elements. PMID- 15292170 TI - Structure of the conserved core of the yeast Dot1p, a nucleosomal histone H3 lysine 79 methyltransferase. AB - Methylation of Lys79 on histone H3 by Dot1p is important for gene silencing. The elongated structure of the conserved core of yeast Dot1p contains an N-terminal helical domain and a seven-stranded catalytic domain that harbors the binding site for the methyl-donor and an active site pocket sided with conserved hydrophobic residues. The S-adenosyl-L-homocysteine exhibits an extended conformation distinct from the folded conformation observed in structures of SET domain histone lysine methyltransferases. A catalytic asparagine (Asn479), located at the bottom of the active site pocket, suggests a mechanism similar to that employed for amino methylation in DNA and protein glutamine methylation. The acidic, concave cleft between the two domains contains two basic residue binding pockets that could accommodate the outwardly protruding basic side chains around Lys79 of histone H3 on the disk-like nucleosome surface. Biochemical studies suggest that recombinant Dot1 proteins are active on recombinant nucleosomes, free of any modifications. PMID- 15292172 TI - Functional analysis of SoxR residues affecting transduction of oxidative stress signals into gene expression. AB - SoxR protein, a member of the MerR family of transcriptional activators, mediates a global oxidative stress response in Escherichia coli. Upon oxidation or nitrosylation of its [2Fe-2S] centers SoxR activates its target gene, soxS, by mediating a structural transition in the promoter DNA that stimulates initiation by RNA polymerase. We explored the molecular basis of this signal transduction by analyzing mutant SoxR proteins defective in responding to oxidative stress signals in vivo.We have confirmed that the DNA binding domain of SoxR is highly conserved compared with other MerR family proteins and functions in a similar manner to activate transcription. Several mutations in the dimerization domain of SoxR disrupted intersubunit communication, and the resulting proteins were unable to propagate redox signals to the soxS promoter. Mutations scattered throughout the polypeptide yielded proteins that were under-responsive to in vivo redox signals, which indicates that the redox properties of the [2Fe-2S] centers are influenced by global protein structure. These findings indicate that SoxR functions as a redox-responsive molecular switch in which subunit interactions transduce a subtle alteration in oxidation state into a profound change in DNA structure. PMID- 15292173 TI - Structure and orientation of pardaxin determined by NMR experiments in model membranes. AB - Pardaxins are a class of ichthyotoxic peptides isolated from fish mucous glands. Pardaxins physically interact with cell membranes by forming pores or voltage gated ion channels that disrupt cellular functions. Here we report the high resolution structure of synthetic pardaxin Pa4 in sodium dodecylphosphocholine micelles, as determined by (1)H solution NMR spectroscopy. The peptide adopts a bend-helix-bend-helix motif with an angle between the two structure helices of 122 +/- 9 degrees , making this structure substantially different from the one previously determined in organic solvents. In addition, paramagnetic solution NMR experiments on Pa4 in micelles reveal that except for the C terminus, the peptide is not solvent-exposed. These results are complemented by solid-state NMR experiments on Pa4 in lipid bilayers. In particular, (13)C-(15)N rotational echo double-resonance experiments in multilamellar vesicles support the helical conformation of the C-terminal segment, whereas (2)H NMR experiments show that the peptide induces considerable disorder in both the head-groups and the hydrophobic core of the bilayers. These solid-state NMR studies indicate that the C-terminal helix has a transmembrane orientation in DMPC bilayers, whereas in POPC bilayers, this domain is heterogeneously oriented on the lipid surface and undergoes slow motion on the NMR time scale. These new data help explain how the non-covalent interactions of Pa4 with lipid membranes induce a stable secondary structure and provide an atomic view of the membrane insertion process of Pa4. PMID- 15292174 TI - Heparan sulfate structure in mice with genetically modified heparan sulfate production. AB - Using a high throughput heparan sulfate (HS) isolation and characterization protocol, we have analyzed HS structure in several tissues from mice/mouse embryos deficient in HS biosynthesis enzymes (N-deacetylase/N-sulfotransferase (NDST)-1, NDST-2, and C5-epimerase, respectively) and in mice lacking syndecan-1. The results have given us new information regarding HS biosynthesis with implications on the role of HS in embryonic development. Our main conclusions are as follows. 1) The HS content, disaccharide composition, and the overall degree of N- and O-sulfation as well as domain organization are characteristic for each individual mouse tissue. 2) Removal of a key biosynthesis enzyme (NDST-1 or C5 epimerase) results in similar structural alterations in all of the tissues analyzed. 3) Essentially no variation in HS tissue structure is detected when individuals of the same genotype are compared. 4) NDST-2, although generally expressed, does not contribute significantly to tissue-specific HS structures. 5) No change in HS structure could be detected in syndecan-1-deficient mice. PMID- 15292175 TI - The mechanism of potent GTP cyclohydrolase I inhibition by 2,4-diamino-6 hydroxypyrimidine: requirement of the GTP cyclohydrolase I feedback regulatory protein. AB - Inhibition of GTP cyclohydrolase I (GTPCH) has been used as a selective tool to assess the role of de novo synthesis of (6R)-5,6,7,8-tetrahydro-L-biopterin (BH4) in a biological system. Toward this end, 2,4-diamino-6-hydroxypyrimidine (DAHP) has been used as the prototypical GTPCH inhibitor. Using a novel real-time kinetic microplate assay for GTPCH activity and purified prokaryote-expressed recombinant proteins, we show that potent inhibition by DAHP is not the result of a direct interaction with GTPCH. Rather, inhibition by DAHP in phosphate buffer occurs via an indirect mechanism that requires the presence of GTPCH feedback regulatory protein (GFRP). Notably, GFRP was previously discovered as the essential factor that reconstitutes inhibition of pure recombinant GTPCH by the pathway end product BH4. Thus, DAHP inhibits GTPCH by engaging the endogenous feedback inhibitory system. We further demonstrate that L-Phe fully reverses the inhibition of GTPCH by DAHP/GFRP, which is also a feature in common with inhibition by BH4/GFRP. These findings suggest that DAHP is not an indiscriminate inhibitor of GTPCH in biological systems; instead, it is predicted to preferentially attenuate GTPCH activity in cells that most abundantly express GFRP and/or contain the lowest levels of L-Phe. PMID- 15292176 TI - Oxidants inhibit ERK/MAPK and prevent its ability to delay neutrophil apoptosis downstream of mitochondrial changes and at the level of XIAP. AB - Normal spontaneous apoptosis in neutrophils is enhanced by "stress" stimuli such as tumor necrosis factor-alpha, Fas ligand, and oxidants, and this effect is inhibited by anti-apoptotic stimuli including granulocyte-macrophage colony stimulating factor, lipopolysaccharide, and formylmethionine-leucine phenylalanine. In this report we demonstrate that anti-apoptotic stimuli protect neutrophils from stress-induced apoptosis via activation of the ERK/MAPK pathway. The protection occurs downstream of mitochondrial alterations assessed as a decrease in membrane potential concomitant with enhanced cytochrome c release. ERK activation was shown to inhibit apoptosis by maintaining levels of XIAP, which is normally decreased in the presence of the pro-apoptotic/stress stimuli. This report also demonstrates that potent intra- and extracellular oxidants inhibit the protective effect of ERK. Oxidant-dependent inhibition of ERK was because of activation of p38 MAPK and activation of the protein phosphatases PP1 and PP2A. Our data suggest that ERK suppresses stress-induced apoptosis downstream of mitochondrial alterations by maintaining XIAP levels and that oxidants block this effect through activation of p38 and protein phosphatases. PMID- 15292177 TI - Establishment and characterization of cultured epithelial cells lacking expression of ZO-1. AB - In well polarized epithelial cells, closely related ZO-1 and ZO-2 are thought to function as scaffold proteins at tight junctions (TJs). In epithelial cells at the initial phase of polarization, these proteins are recruited to cadherin-based spotlike adherens junctions (AJs). As a first step to clarify the function of ZO 1, we successfully generated mouse epithelial cell clones lacking ZO-1 expression (ZO-1-/- cells) by homologous recombination. Unexpectedly, in confluent cultures, ZO-1-/- cells were highly polarized with well organized AJs/TJs, which were indistinguishable from those in ZO-1+/+ cells by electron microscopy. In good agreement, by immunofluorescence microscopy, most TJ proteins including claudins and occludin appeared to be normally concentrated at TJs of ZO-1-/- cells with the exception that a ZO-1 deficiency significantly up- or down-regulated the recruitment of ZO-2 and cingulin, another TJ scaffold protein, respectively, to TJs. When the polarization of ZO-1-/- cells was initiated by a Ca2+ switch, the initial AJ formation did not appear to be affected; however, the subsequent TJ formation (recruitment of claudins/occludin to junctions and barrier establishment) was markedly retarded. This retardation as well as the disappearance of cingulin were rescued completely by exogenous ZO-1 but not by ZO 2 expression. Quantitative evaluation of ZO-1/ZO-2 expression levels led to the conclusion that ZO-1 and ZO-2 would function redundantly to some extent in junction formation/epithelial polarization but that they are not functionally identical. Finally, we discussed advantageous aspects of the gene knock-out system with cultured epithelial cells in epithelial cell biology. PMID- 15292178 TI - Re-expression of detachment-inducible chloride channel mCLCA5 suppresses growth of metastatic breast cancer cells. AB - The calcium-activated chloride channel hCLCA2 has been identified as a candidate tumor suppressor in human breast cancer. It is greatly down-regulated in breast cancer, and its re-expression suppresses tumorigenesis by an unknown mechanism. To establish a mouse model, we identified the mouse ortholog of hCLCA2, termed mCLCA5, and investigated its behavior in mammary epithelial cell lines and tissues. Expression in the immortalized cell line HC11 correlated with slow or arrested growth. Although rapidly dividing, sparsely plated cells had low levels of expression, mCLCA5 was induced by 10-fold when cells became confluent and 30 fold when cells were deprived of growth factors or anchorage. The apoptosis effector Bax was induced in parallel. Like hCLCA2, mCLCA5 was down-regulated in metastatic mammary tumor cell lines such as 4T1 and CSML-100. Ectopic re expression in 4T1 cells caused a 20-fold reduction in colony survival relative to vector control. High mCLCA5 expression in stable clones inhibited proliferation and enhanced sensitivity to detachment. Moreover, mCLCA5 was induced in lactating and involuting mammary gland, correlating with differentiation and onset of apoptosis. Together, these results establish mCLCA5 as the mouse ortholog of hCLCA2, demonstrate that mCLCA5 is a detachment-sensitive growth inhibitor, and suggest a mechanism whereby these channels may antagonize mammary tumor progression. PMID- 15292179 TI - NADPH oxidase and ERK signaling regulates hyperoxia-induced Nrf2-ARE transcriptional response in pulmonary epithelial cells. AB - Oxidative stress plays a major role in hyperoxia-induced acute lung injury. We have shown previously that mice lacking the Nrf2 are more susceptible to hyperoxia than are wild-type mice. Nrf2 activates antioxidant response element (ARE)-mediated gene expression involved in cellular protection against toxic insults. The present study was designed to investigate the mechanisms that control the activation of Nrf2 by hyperoxia using a non-malignant murine alveolar epithelial cell line, C10. No significant alteration in the levels of Nrf2 mRNA and protein was found following exposure to hyperoxia. In contrast, hyperoxia caused the translocation of Nrf2 from the cytoplasm to the nucleus within 30-60 min of exposure. Consistent with these observations, gel shift and reporter analyses demonstrated a correlation between the hyperoxia-enhanced ARE DNA binding activity of Nrf2 and an up-regulation of ARE-driven transcription. Inhibition of NADPH oxidase with diphenyleneiodonium (DPI) blocked both Nrf2 translocation and ARE-mediated transcription. Inhibition of the MEK/ERK pathway caused a similar effect. Consistent with this finding, hyperoxia stimulated ERK-1 and ERK-2 phosphorylation, whereas DPI or N-acetyl-l-cysteine blocked such activation. Hyperoxia stimulated the phosphorylation of endogenous Nrf2, but not in the presence of U0126, suggesting a critical role for ERK signaling in the activation of Nrf2. Consistent with this notion, hyperoxia did not stimulate the phosphorylation of Nrf2 in fibroblasts lacking the ERK-1. Collectively, our findings suggest that hyperoxia-induced, ARE-driven, Nrf2-dependent transcription is controlled by NADPH oxidase and ERK-1 signaling. PMID- 15292180 TI - Altered expression of genes of the Bmp/Smad and Wnt/calcium signaling pathways in the cone-only Nrl-/- mouse retina, revealed by gene profiling using custom cDNA microarrays. AB - Many mammalian retinas are rod-dominant, and hence our knowledge of cone photoreceptor biology is relatively limited. To gain insights into the molecular differences between rods and cones, we compared the gene expression profile of the rod-dominated retina of wild type mouse with that of the cone-only retina of Nrl(-/-) (Neural retina leucine zipper knockout) mouse. Our analysis, using custom microarrays of eye-expressed genes, provided equivalent data using either direct or reference-based experimental designs, confirmed differential expression of rod- and cone-specific genes in the Nrl(-/-) retina and identified novel genes that could serve as candidates for retinopathies or for functional studies. In addition, we detected altered expression of several genes that encode cell signaling or structural proteins. Prompted by these findings, additional real time PCR analysis revealed that genes belonging to the Bmp/Smad and Wnt/Ca(2+) signaling pathways are expressed in the mature wild type retina and that their expression is significantly altered in the Nrl(-/-) retina. Chromatin immunoprecipitation analysis of adult retina identified Bmp4 and Smad4, which are down-regulated in the Nrl(-/-) retina, as possible direct transcriptional targets of Nrl. Consistent with these studies, Bmp4 and Smad4 are expressed in the mature rod photoreceptors of mouse retina. Modulation of Bmp4 and/or Smad4 by Nrl may provide a mechanism for integrating diverse cell signaling networks in rods. We hypothesize that Bmp/Smad and Wnt/Ca(2+) pathways participate in cell-cell communication in the mature retina, and expression changes observed in the Nrl(-/ ) retina reflect their biased utilization in rod versus cone homeostasis. PMID- 15292181 TI - Proliferation of neointimal smooth muscle cells after arterial injury. Dependence on interactions between fibroblast growth factor receptor-2 and fibroblast growth factor-9. AB - The growth factor signaling mechanisms responsible for neointimal smooth muscle cell (SMC) proliferation and accumulation, a characteristic feature of many vascular pathologies that can lead to restenosis after angioplasty, remain to be identified. Here, we examined the contribution of fibroblast growth factor receptors (FGFRs) 2 and 3 as well as novel fibroblast growth factors (FGFs) to such proliferation. Balloon catheter injury to the rat carotid artery stimulated the expression of two distinctly spliced FGFR-2 isoforms, differing only by the presence or absence of the acidic box, and two distinctly spliced FGFR-3 isoforms containing the acidic box and differing only by the presence of either the IIIb or IIIc exon. Post-injury arterial administration of recombinant adenoviruses expressing dominant negative mutant forms of these FGFRs were used to assess the roles of the endogenous FGFR isoforms in neointimal SMC proliferation. Dominant negative FGFR-2 containing the acidic box inhibited such proliferation by 40%, whereas the dominant negative FGFR-3 forms had little effect. Expression of FGF 9, known to be capable of binding to all four neointimal FGFR-2/-3 isoforms, was abundant within the neointima. FGF-9 markedly stimulated both the proliferation of neointimal SMCs and the activation of extracellular signal-related kinases 1/2, effects which were abrogated by the administration of antisense FGF-9 oligonucleotides to injured arteries and the expression of the dominant negative FGFR-2 adenovirus in cultured neointimal SMCs. These studies demonstrate that, although multiple FGFRs are induced in neointimal SMCs following arterial injury, specific interactions between distinctly spliced FGFR-2 isoforms and FGF-9 contribute to the proliferation of these SMCs. PMID- 15292182 TI - Intectin, a novel small intestine-specific glycosylphosphatidylinositol-anchored protein, accelerates apoptosis of intestinal epithelial cells. AB - Intestinal epithelial cells undergo rapid turnover and exfoliation especially at the villus tips. This process is modulated by various nutrients especially fat. Apoptosis is one of the important regulatory mechanisms of this turnover. Therefore, identification of the factors that control epithelial cell apoptosis should help us understand the mechanism of intestinal mucosal turnover. Here, we report the identification of a novel small intestine-specific member of the Ly-6 family, intectin, by signal sequence trap method. Intectin mRNA expression was exclusively identified in the intestine and localized at the villus tips of intestinal mucosa, which is known to undergo apoptosis. Intectin mRNA expression was modulated by nutrition. Intestinal epithelial cells expressing intectin were more sensitive to palmitate-induced apoptosis, compared with control intestinal epithelial cells, and such effect was accompanied by increased activity of caspase-3. Intectin expression also reduced cell-cell adhesion of intestinal epithelial cells. PMID- 15292183 TI - A proteome-wide approach identifies sumoylated substrate proteins in yeast. AB - The ubiquitin-related protein SUMO-1 is covalently attached to proteins by SUMO-1 ligases. We have performed a proteome-wide analysis of sumoylated substrate proteins in yeast. Employing the powerful affinity purification of Protein A-Smt3 (Smt3 is the yeast homologue of SUMO-1) from yeast lysates in combination with tandem liquid chromatography mass spectrometry, we have isolated potential Smt3 carrying substrate proteins involved in DNA replication and repair, chromatin remodeling, transcription activation, Pol-I, Pol-II, and Pol-III transcription, 5' pre-mRNA capping, 3' pre-mRNA processing, proteasome function, and tubulin folding. Employing tandem affinity purifications or a rapid biochemical assay referred to as "SUMO fingerprint," we showed that several subunits of RNA polymerases I, II, and III, members of the transcription repression and chromatin remodeling machineries previously not known to be sumoylated, are modified by SUMO-1. Thus, the identification of a broad range of SUMO-1 substrate proteins is expected to lead to further insight into the regulatory aspects of sumoylation. PMID- 15292184 TI - Hepatitis C virus core selectively suppresses interleukin-12 synthesis in human macrophages by interfering with AP-1 activation. AB - Hepatitis C virus (HCV) is remarkably efficient at establishing persistent infection, suggesting that it has evolved one or more strategies aimed at evading the host immune response. T cell responses, including interferon-gamma production, are severely suppressed in chronic HCV patients. The HCV core protein has been previously shown to circulate in the bloodstream of HCV-infected patients and inhibit host immunity through an interaction with gC1qR. To determine the role of the HCV core-gC1qR interaction in modulation of inflammatory cytokine production, we examined interleukin (IL)-12 production, which is critical for the induction of interferon-gamma synthesis, in lipopolysaccharide-stimulated human monocyte/macrophages. We found that core protein binds the gC1qR displayed on the cell surface of monocyte/macrophages and inhibits the production of IL-12p70 upon lipopolysaccharide stimulation. This inhibition was found to be selective in that HCV core failed to affect the production of IL-6, IL-8, IL-1beta, and tumor necrosis factor alpha. In addition, suppression of IL-12 production by core protein occurred at the transcriptional level by inhibition of IL-12p40 mRNA synthesis. Importantly, core-induced inhibition of IL-12p40 mRNA synthesis resulted from impaired activation of AP-1 rather than enhanced IL-10 production. These results suggest that the HCV core gC1qR interaction may play a pivotal role in establishing persistent infection by dampening TH1 responses. PMID- 15292185 TI - Mechanism of CD47-induced alpha4beta1 integrin activation and adhesion in sickle reticulocytes. AB - We recently reported that CD47 (integrin-associated protein) on sickle red blood cells (SS RBCs) activates G-protein-dependent signaling, which promotes cell adhesion to immobilized thrombospondin (TSP) under relevant shear stress. These data suggested that signal transduction in SS RBCs may contribute to the vaso occlusive pathology observed in sickle cell disease. However, the CD47-activated SS RBC adhesion receptor(s) that mediated adhesion to immobilized TSP remained unknown. Here we demonstrate that the alpha4beta1 integrin (VLA-4) is the receptor that mediates CD47-stimulated SS RBC adhesion to immobilized TSP. This adhesion requires both the N-terminal heparin-binding domain and the RGD site of TSP. CD47 signaling induces an "inside-out" activation of alpha4beta1 on SS RBCs as indicated by an RGD-dependent interaction of this integrin with soluble, plasma fibronectin. However, CD47 engagement also induces an alpha4beta1 mediated, RGD-independent adhesion of SS RBCs to immobilized vascular cell adhesion molecule-1 (VCAM-1). CD47 signaling in SS RBCs appears to be independent of large scale changes in cAMP formation but nonetheless promotes alpha4beta1 mediated adhesion via a protein kinase A-dependent, serine phosphorylation of the alpha4 cytoplasmic domain. CD47-activated SS RBC adhesion absolutely requires the Src family tyrosine kinases and is also enhanced by treatment of SS RBCs with low concentrations of cytochalasin D, which may release alpha4beta1 from cytoskeletal restraints. In addition, CD47 co-immunoprecipitates with alpha4beta1 in a sickle reticulocyte-enriched fraction of SS RBCs. These studies therefore identify the alpha4beta1 integrin on SS RBCs as a CD47-activated receptor for TSP, VCAM-1, and plasma fibronectin, revealing novel binding characteristics of this integrin. PMID- 15292186 TI - The three-dimensional structure of the ZAP-70 kinase domain in complex with staurosporine: implications for the design of selective inhibitors. AB - The ZAP-70 tyrosine kinase plays a critical role in T cell activation and the immune response and therefore is a logical target for immunomodulatory therapies. Although the crystal structure of the tandem Src homology-2 domains of human ZAP 70 in complex with a peptide derived from the zeta subunit of the T cell receptor has been reported (Hatada, M. H., Lu, X., Laird, E. R., Green, J., Morgenstern, J. P., Lou, M., Marr, C. S., Phillips, T. B., Ram, M. K., Theriault, K., Zoller, M. J., and Karas, J. L. (1995) Nature 377, 32-38), the structure of the kinase domain has been elusive to date. We crystallized and determined the three dimensional structure of the catalytic subunit of ZAP-70 as a complex with staurosporine to 2.3 A resolution, utilizing an active kinase domain containing residues 327-606 identified by systematic N- and C-terminal truncations. The crystal structure shows that this ZAP-70 kinase domain is in an active-like conformation despite the lack of tyrosine phosphorylation in the activation loop. The unique features of the ATP-binding site, identified by structural and sequence comparison with other kinases, will be useful in the design of ZAP-70 selective inhibitors. PMID- 15292187 TI - Small interfering RNA-mediated down-regulation of caveolin-1 differentially modulates signaling pathways in endothelial cells. AB - Caveolin-1 is a scaffolding/regulatory protein that interacts with diverse signaling molecules in endothelial cells. To explore the role of this protein in receptor-modulated signaling pathways, we transfected bovine aortic endothelial cells (BAEC) with small interfering RNA (siRNA) duplexes to down-regulate caveolin-1 expression. Transfection of BAEC with duplex siRNA targeted against caveolin-1 mRNA selectively "knocked-down" the expression of caveolin-1 by approximately 90%, as demonstrated by immunoblot analyses of BAEC lysates. We used discontinuous sucrose gradients to purify caveolin-containing lipid rafts from siRNA-treated endothelial cells. Despite the near-total down-regulation of caveolin-1 expression, the lipid raft targeting of diverse signaling proteins (including the endothelial isoform of nitric-oxide synthase, Src-family tyrosine kinases, Galphaq and the insulin receptor) was unchanged. We explored the consequences of caveolin-1 knockdown on kinase pathways modulated by the agonists sphingosine-1 phosphate (S1P) and vascular endothelial growth factor (VEGF). siRNA-mediated caveolin-1 knockdown enhanced basal as well as S1P- and VEGF induced phosphorylation of the protein kinase Akt and did not modify the basal or agonist-induced phosphorylation of extracellular signal-regulated kinases 1/2. Caveolin-1 knock-down also significantly enhanced the basal and agonist-induced activity of the small GTPase Rac. We used siRNA to down-regulate Rac expression in BAEC, and we observed that Rac knockdown significantly reduced basal, S1P-, and VEGF-induced Akt phosphorylation, suggesting a role for Rac activation in the caveolin siRNA-mediated increase in Akt phosphorylation. By using siRNA to knockdown caveolin-1 and Rac expression in cultured endothelial cells, we have found that caveolin-1 does not seem to be required for the targeting of signaling molecules to caveolae/lipid rafts and that caveolin-1 differentially modulates specific kinase pathways in endothelial cells. PMID- 15292188 TI - The PDZ domain of the SpoIVB transmembrane signaling protein enables cis-trans interactions involving multiple partners leading to the activation of the pro sigmaK processing complex in Bacillus subtilis. AB - In sporulating cells of Bacillus subtilis, the serine peptidase SpoIVB is the essential component of a transmembrane signaling cascade between the two intracellular compartments (forespore and mother cell) that leads to activation of the sigmaK transcription factor in the mother cell chamber. This regulatory process, referred to as the sigmaK checkpoint, is essential for ensuring proper development of the spore and introduces an appropriate level of fidelity to the developmental process. This work unravels the signaling process and establishes how SpoIVB interacts with other protein partners in the sigmaK checkpoint. SpoIVB is synthesized as a zymogen that is autoproteolytically activated and carries a PDZ domain that is responsible for at least three distinct binding reactions, a phenomenon not previously demonstrated for an individual PDZ domain. First, binding to the SpoIVB NH2 terminus to maintain the protein in its zymogen form. Second, following secretion across a spore membrane, binding in trans to the COOH terminus of another SpoIVB molecule. Binding in trans facilitates the first cleavage event of SpoIVB near the NH2 terminus releasing it from the inner forespore membrane. We show that at least two further cis cleavage events occur at specific sites near the NH2 terminus after which the PDZ domain targets SpoIVB to the pro-sigmaK processing complex in the outer forespore membrane. Specifically, SpoIVB binds to the COOH terminus of BofA. In turn, this allows SpoIVB to cleave the COOH terminus of SpoIVFA an event pivotal to activating the SpoIVFB zinc metalloprotease by disruption of the heteroligomeric pro-sigmaK complex. PMID- 15292189 TI - A novel venom peptide from an endoparasitoid wasp is required for expression of polydnavirus genes in host hemocytes. AB - Maternal factors introduced into host insects by endoparasitoid wasps are usually essential for successful parasitism. This includes polydnaviruses (PDVs) that are produced in the reproductive organ of female hymenopteran endoparasitoids and are injected, together with venom proteins, into the host hemocoel at oviposition. Inside the host, PDVs enter various tissue cells and hemocytes where viral genes are expressed, leading to developmental and physiological alterations in the host, including the suppression of the host immune system. Although several studies have shown that some PDVs are only effective when accompanied by venom proteins, there is no report of an active venom ingredient(s) facilitating PDV infection and/or gene expression. In this study, we describe a novel peptide (Vn1.5) isolated from Cotesia rubecula venom that is required for the expression of C. rubecula bracoviruses (CrBVs) in host hemocytes (Pieris rapae), although it is not essential for CrBV entry into host cells. The peptide consists of 14 amino acids with a molecular mass of 1598 Da. In the absence of Vn1.5 or total venom proteins, CrBV genes are not expressed in host cells and did not cause inactivation of host hemocytes. PMID- 15292190 TI - Peptidoglycan amidase MepA is a LAS metallopeptidase. AB - LAS enzymes are a group of metallopeptidases that share an active site architecture and a core folding motif and have been named according to the group members lysostaphin, D-Ala-D-Ala carboxypeptidase and sonic hedgehog. Escherichia coli MepA is a periplasmic, penicillin-insensitive murein endopeptidase that cleaves the D-alanyl-meso-2,6-diamino-pimelyl amide bond in E. coli peptidoglycan. The enzyme lacks sequence similarity with other peptidases, and is currently classified as a peptidase of unknown fold and catalytic class in all major data bases. Here, we build on our observation that two motifs, characteristic of the newly described LAS group of metallopeptidases, are conserved in MepA-type sequences. We demonstrate that recombinant E. coli MepA is sensitive to metal chelators and that mutations in the predicted Zn2+ ligands His 113, Asp-120, and His-211 inactivate the enzyme. Moreover, we present the crystal structure of MepA. The active site of the enzyme is most similar to the active sites of lysostaphin and D-Ala-D-Ala carboxypeptidase, and the fold is most closely related to the N-domain of sonic hedgehog. We conclude that MepA-type peptidases are LAS enzymes. PMID- 15292191 TI - A CDC6-like factor from the archaea Sulfolobus solfataricus promotes binding of the mini-chromosome maintenance complex to DNA. AB - The archaeal replication apparatus appears to be a simplified version of the eukaryotic one with fewer polypeptides and simpler protein complexes. Herein, we report evidence that a Cdc6-like factor from the hyperthermophilic crenarchaea Sulfolobus solfataricus stimulates binding of the homohexameric MCM-like complex to bubble- and fork-containing DNA oligonucleotides that mimic early replication intermediates. This function does not require the Cdc6 ATP and DNA binding activities. These findings may provide important clues to understanding how the DNA replication initiation process has evolved in the more complex eukaryotic organisms. PMID- 15292192 TI - Bone morphogenetic protein-1/tolloid-related metalloproteinases process osteoglycin and enhance its ability to regulate collagen fibrillogenesis. AB - The mammalian bone morphogenetic protein-1 (BMP-1)/Tolloid-related metalloproteinases play key roles in regulating formation of the extracellular matrix (ECM) via biosynthetic processing of various precursor proteins into mature functional enzymes, structural proteins, and proteins involved in initiating the mineralization of hard tissue ECMs. They also have been shown to activate several members of the transforming growth factor-beta superfamily, and may serve to coordinate such activation with formation of the ECM in morphogenetic events. Osteoglycin (OGN), a small leucine-rich proteoglycan with unclear functions, is found in cornea, bone, and other tissues, and appears to undergo proteolytic processing in vivo. Here we have successfully generated recombinant OGN and have employed it to demonstrate that a pro-form of OGN is processed to varying extents by all four mammalian BMP-1/Tolloid-like proteinases, to generate a 27-kDa species that corresponds to the major form of OGN found in cornea. Moreover, whereas wild-type mouse embryo fibroblasts (MEFs) produce primarily the processed, mature form of OGN, MEFs homozygous null for genes encoding three of the four mammalian BMP-1/Tolloid-related proteinases produce only unprocessed pro-OGN. Thus, all detectable pro-OGN processing activity in MEFs is accounted for by products of these genes. We also demonstrate that both pro- and mature OGN can regulate type I collagen fibrillogenesis, and that processing of the prodomain by BMP-1 potentiates the ability of OGN to modulate the formation of collagen fibrils. PMID- 15292193 TI - Identification of nine sucrose nonfermenting 1-related protein kinases 2 activated by hyperosmotic and saline stresses in Arabidopsis thaliana. AB - Several calcium-independent protein kinases were activated by hyperosmotic and saline stresses in Arabidopsis cell suspension. Similar activation profiles were also observed in seedlings exposed to hyperosmotic stress. One of them was identified to AtMPK6 but the others remained to be identified. They were assumed to belong to the SNF1 (sucrose nonfermenting 1)-related protein kinase 2 (SnRK2) family, which constitutes a plant-specific kinase group. The 10 Arabidopsis SnRK2 were expressed both in cells and seedlings, making the whole SnRK2 family a suitable candidate. Using a family-specific antibody raised against the 10 SnRK2, we demonstrated that these non-MAPK protein kinases activated by hyperosmolarity in cell suspension were SnRK2 proteins. Then, the molecular identification of the involved SnRK2 was investigated by transient expression assays. Nine of the 10 SnRK2 were activated by hyperosmolarity induced by mannitol, as well as NaCl, indicating an important role of the SnRK2 family in osmotic signaling. In contrast, none of the SnRK2 were activated by cold treatment, whereas abscisic acid only activated five of the nine SnRK2. The probable involvement of the different Arabidopsis SnRK2 in several abiotic transduction pathways is discussed. PMID- 15292194 TI - Crystal structure of arachidonic acid bound to a mutant of prostaglandin endoperoxide H synthase-1 that forms predominantly 11-hydroperoxyeicosatetraenoic acid. AB - Kinetic studies and analysis of the products formed by native and mutant forms of ovine prostaglandin endoperoxide H synthase-1 (oPGHS-1) have suggested that arachidonic acid (AA) can exist in the cyclooxygenase active site of the enzyme in three different, catalytically competent conformations that lead to prostaglandin G2 (PGG2), 11R-hydroperoxyeicosatetraenoic acid (HPETE), and 15R,S HPETE, respectively. We have identified an oPGHS-1 mutant (V349A/W387F) that forms predominantly 11R-HPETE. Thus, the preferred catalytically competent arrangement of AA in the cyclooxygenase site of this double mutant must be one that leads to 11-HPETE. The crystal structure of Co3+-protoporphyrin IX V349A/W387F oPGHS-1 in a complex with AA was determined to 3.1 A. Significant differences are observed in the positions of atoms C-3, C-4, C-5, C-6, C-10, C 11, and C-12 of bound AA between native and V349A/W387F oPGHS-1; in comparison, the positions of the side chains of cyclooxygenase active site residues are unchanged. The structure of the double mutant presented here provides structural insight as to how Val349 and Trp387 help position C-9 and C-11 of AA so that the incipient 11-peroxyl radical intermediate is able to add to C-9 to form the 9,11 endoperoxide group of PGG2. In the V349A/W387F oPGHS-1.AA complex the locations of C-9 and C-11 of AA with respect to one another make it difficult to form the endoperoxide group from the 11-hydroperoxyl radical. Therefore, the reaction apparently aborts yielding 11R-HPETE instead of PGG2. In addition, the observed differences in the positions of carbon atoms of AA bound to this mutant provides indirect support for the concept that the conformer of AA shown previously to be bound within the cyclooxygenase active site of native oPGHS-1 is the one that leads to PGG2. PMID- 15292195 TI - Involvement of phosphoinositide 3-kinase gamma, Rac, and PAK signaling in chemokine-induced macrophage migration. AB - In macrophages, chemotactic stimuli cause the activation of Rac and PAK, but little is known about the signaling pathways involved and their role in chemotactic gradient sensing. Herein, we report that in macrophages, the chemokine RANTES (regulated on activation normal T cell expressed and secreted)/CCL5 activates the small GTPase Rac and its downstream target PAK2 within seconds. This response depends on Gi activation and largely on the subsequent triggering of phosphoinositide 3-kinase gamma (PI3Kgamma) and Rac. Retroviral transduction of tagged Rac1 and -2 indicates that RANTES/CCL5-mediated activation of PI3Kgamma triggers Rac1 but not Rac2. In agreement, silencing of Rac1 by shRNA blocks PAK2 activity and inhibits RANTES/CCL5-induced macrophage polarization and directional migration. On the other hand, the tyrosine kinase receptor agonist CSF-1 activates PAK2 independently of PI3Kgamma and Rac. Our results thus demonstrate a chemokine-specific signaling pathway in which Gi and PI3Kgamma coordinate to drive Rac1 and PAK2 activation that eventually controls the chemotactic response. PMID- 15292197 TI - JACOP, a novel plaque protein localizing at the apical junctional complex with sequence similarity to cingulin. AB - The apical junctional complex is composed of various cell adhesion molecules and cytoplasmic plaque proteins. Using a monoclonal antibody that recognizes a chicken 155-kDa cytoplasmic antigen (p155) localizing at the apical junctional complex, we have cloned a cDNA of its mouse homologue. The full-length cDNA of mouse p155 encoded a 148-kDa polypeptide containing a coiled-coil domain with sequence similarity to cingulin, a tight junction (TJ)-associated plaque protein. We designated this protein JACOP (junction-associated coiled-coil protein). Immunofluorescence staining showed that JACOP was concentrated in the junctional complex in various types of epithelial and endothelial cells. Furthermore, in the liver and kidney, JACOP was also distributed along non-junctional actin filaments. Upon immunoelectron microscopy, JACOP was found to be localized to the undercoat of TJs in the liver, but in some tissues, its distribution was not restricted to TJs but extended to the area of adherens junctions. Overexpression studies have revealed that JACOP was recruited to the junctional complex in epithelial cells and to cell-cell contacts and stress fibers in fibroblasts. These findings suggest that JACOP is involved in anchoring the apical junctional complex, especially TJs, to actin-based cytoskeletons. PMID- 15292196 TI - The role of interleukin 1 receptor-associated kinase-4 (IRAK-4) kinase activity in IRAK-4-mediated signaling. AB - Interleukin 1 receptor (IL-1R)-associated kinase-4 (IRAK-4) is required for various responses induced by IL-1R and Toll-like receptor signals. However, the molecular mechanism of IRAK-4 signaling and the role of its kinase activity have remained elusive. In this report, we demonstrate that IRAK-4 is recruited to the IL-1R complex upon IL-1 stimulation and is required for the recruitment of IRAK-1 and its subsequent activation/degradation. By reconstituting IRAK-4-deficient cells with wild type or kinase-inactive IRAK-4, we show that the kinase activity of IRAK-4 is required for the optimal transduction of IL-1-induced signals, including the activation of IRAK-1, NF-kappaB, and JNK, and the maximal induction of inflammatory cytokines. Interestingly, we also discover that the IRAK-4 kinase inactive mutant is still capable of mediating some signals. These results suggest that IRAK-4 is an integral part of the IL-1R signaling cascade and is capable of transmitting signals both dependent on and independent of its kinase activity. PMID- 15292198 TI - Cardiolipin biosynthesis and mitochondrial respiratory chain function are interdependent. AB - Cardiolipin (CL) is an acidic phospholipid present almost exclusively in membranes harboring respiratory chain complexes. We have previously shown that, in Saccharomyces cerevisiae, CL provides stability to respiratory chain supercomplexes and CL synthase enzyme activity is reduced in several respiratory complex assembly mutants. In the current study, we investigated the interdependence of the mitochondrial respiratory chain and CL biosynthesis. Pulse labeling experiments showed that in vivo CL biosynthesis was reduced in respiratory complexes III (ubiquinol:cytochrome c oxidoreductase) and IV (cytochrome c oxidase) and oxidative phosphorylation complex V (ATP synthase) assembly mutants. CL synthesis was decreased in the presence of CCCP, an inhibitor of oxidative phosphorylation that reduces the pH gradient but not by valinomycin or oligomycin, both of which reduce the membrane potential and inhibit ATP synthase, respectively. The inhibitors had no effect on phosphatidylglycerol biosynthesis or CRD1 gene expression. These results are consistent with the hypothesis that in vivo CL biosynthesis is regulated at the level of CL synthase activity by the DeltapH component of the proton-motive force generated by the functional electron transport chain. This is the first report of regulation of phospholipid biosynthesis by alteration of subcellular compartment pH. PMID- 15292199 TI - Regulation of the Cell Type-specific dentin sialophosphoprotein gene expression in mouse odontoblasts by a novel transcription repressor and an activator CCAAT binding factor. AB - Dentin sialophosphoprotein (DSPP) is an extracellular matrix protein that is cleaved into dentin sialoprotein (DSP) and dentin phosphoprotein (DPP) with a highly restricted expression pattern in tooth and bone. Mutations of the DSPP gene are associated with dentin genetic diseases. Regulation of tissue-specific DSPP expression has not been described. To define the molecular basis of this cell-specific expression, we characterized the promoter responsible for the cell specific expression of the DSPP gene in odontoblasts. Within this region, DNase I footprinting and electrophoretic mobility shift assays delineated one element that contains an inverted CCAAT-binding factor site and a protein-DNA binding site using nuclear extracts from odontoblasts. A series of competitive electrophoretic mobility shift assay analyses showed that the protein-DNA binding core sequence, ACCCCCA, is a novel site sufficient for protein binding. These two protein-DNA binding sequences are conserved at the same proximal position in the mouse, rat, and human DSPP gene promoters and are ubiquitously present in the promoters of other tooth/bone genes. Mutations of the CCAAT-binding factor binding site resulted in a 5-fold decrease in promoter activity, whereas abolishment of the novel protein-DNA binding site increased promoter activity by about 4.6-fold. In contrast to DSPP, expression levels of the novel protein were significantly reduced during odontoblastic differentiation and dentin mineralization. The novel protein was shown to have a molecular mass of 72 kDa. This study shows that expression of the cell type-specific DSPP gene is mediated by the combination of inhibitory and activating mechanisms. PMID- 15292200 TI - Nuclear envelope breakdown requires overcoming the mechanical integrity of the nuclear lamina. AB - In prophase cells, lamin B1 is the major component of the nuclear lamina, a filamentous network underlying the nucleoplasmic side of the nuclear membrane, whereas lamin A/C is dissociated from the scaffold. In vivo fluorescence microscopy studies have shown that, during the G2/M transition, the first gap in the nuclear envelope (NE) appears before lamin B1 disassembly and is caused by early spindle microtubules impinging on the NE. This result suggests that the mechanical tearing of the NE by microtubules plays a central role to the progression of mitosis. To investigate whether this microtubule-induced NE deformation is sufficient for NE breakdown, we assess the mechanical resilience of a reconstituted lamin B1 network. Quantitative rheological methods demonstrate that human lamin B1 filaments form stiff networks that can resist much greater deformations than those caused by microtubules impinging on the NE. Moreover, lamin B1 networks possess an elastic stiffness, which increases under tension, and an exceptional resilience against shear deformations. These results demonstrate that both mechanical tearing of the lamina and biochemical modification of lamin B1 filaments are required for NE breakdown. PMID- 15292201 TI - Sec15 is an effector for the Rab11 GTPase in mammalian cells. AB - Rab/Ypt GTPases play key roles in the regulation of vesicular trafficking. They perform most of their functions in a GTP-bound form by interacting with specific downstream effectors. The exocyst is a complex of eight polypeptides involved in constitutive secretion and functions as an effector for multiple Ras-related small GTPases, including the Rab protein Sec4p in yeast. In this study, we have examined the localization and function of the Sec15 exocyst subunit in mammalian cells. Overexpressed Sec15 associated with clusters of tubular/vesicular elements that were concentrated in the perinuclear region. The tubular/vesicular clusters were dispersed throughout the cytoplasm upon treatment with the microtubule depolymerizing agent nocodazole and were accessible to endocytosed transferrin, but not exocytic cargo (vesicular stomatitis virus glycoprotein). Consistent with these observations, Sec15 colocalized selectively with the recycling endosome marker Rab11 and exhibited a GTP-dependent interaction with the Rab11 GTPase, but not with Rab4, Rab6, or Rab7. These findings provide the first evidence that the exocyst functions as a Rab effector complex in mammalian cells. PMID- 15292202 TI - Heparanase uptake is mediated by cell membrane heparan sulfate proteoglycans. AB - Heparanase is a mammalian endoglycosidase that degrades heparan sulfate (HS) at specific intrachain sites, an activity that is strongly implicated in cell dissemination associated with metastasis and inflammation. In addition to its structural role in extracellular matrix assembly and integrity, HS sequesters a multitude of polypeptides that reside in the extracellular matrix as a reservoir. A variety of growth factors, cytokines, chemokines, and enzymes can be released by heparanase activity and profoundly affect cell and tissue function. Thus, heparanase bioavailability, accessibility, and activity should be kept tightly regulated. We provide evidence that HS is not only a substrate for, but also a regulator of, heparanase. Addition of heparin or xylosides to cell cultures resulted in a pronounced accumulation of, heparanase in the culture medium, whereas sodium chlorate had no such effect. Moreover, cellular uptake of heparanase was markedly reduced in HS-deficient CHO-745 mutant cells, heparan sulfate proteoglycan-deficient HT-29 colon cancer cells, and heparinase-treated cells. We also studied the heparanase biosynthetic route and found that the half life of the active enzyme is approximately 30 h. This and previous localization studies suggest that heparanase resides in the endosomal/lysosomal compartment for a relatively long period of time and is likely to play a role in the normal turnover of HS. Co-localization studies and cell fractionation following heparanase addition have identified syndecan family members as candidate molecules responsible for heparanase uptake, providing an efficient mechanism that limits extracellular accumulation and function of heparanase. PMID- 15292203 TI - Heat shock induces preferential translation of ERGIC-53 and affects its recycling pathway. AB - ERGIC-53 is a lectin-like transport receptor protein, which recirculates between the ER and the Golgi complex and is required for the intracellular transport of a restricted number of glycoproteins. We show in this article that ERGIC-53 accumulates during the heat shock response. However, at variance with the unfolded protein response, which results in enhanced transcription of ERGIC-53 mRNA, heat shock leads only to enhanced translation of ERGIC-53 mRNA. In addition, the half-life of the protein does not change during heat shock. Therefore, distinct signal pathways of the cell stress response modulate the ERGIC-53 protein level. Heat shock also affects the recycling pathway of ERGIC 53. The protein rapidly redistributes in a more peripheral area of the cell, in a vesicular compartment that has a lighter sedimentation density on sucrose gradient in comparison to the compartment that contains the majority of ERGIC-53 at 37 degrees C. This effect is specific, as no apparent reorganization of the endoplasmic reticulum, intermediate compartment and Golgi complex is morphologically detectable in the cells exposed to heat shock. Moreover, the anterograde transport of two unrelated reporter proteins is not affected. Interestingly, MCFD2, which interacts with ERGIC-53 to form a complex required for the ER-to-Golgi transport of specific proteins, is regulated similarly to ERGIC-53 in response to cell stress. These results support the view that ERGIC-53 alone, or in association with MCFD2, plays important functions during cellular response to stress conditions. PMID- 15292204 TI - BBK32, a fibronectin binding MSCRAMM from Borrelia burgdorferi, contains a disordered region that undergoes a conformational change on ligand binding. AB - BBK32 is a fibronectin-binding lipoprotein on Borrelia burgdorferi, the causative agent of Lyme disease. Analysis using secondary structure prediction programs suggested that BBK32 is composed of two domains, an N-terminal segment lacking well defined secondary structure and a C-terminal segment composed largely of alpha-helices. Analysis of purified recombinant forms of the two domains by circular dichroism spectroscopy, gel permeation chromatography, and intrinsic viscosity determination were consistent with an N-terminal-extended, unstructured segment and a C-terminal globular domain in BBK32. Solid phase binding experiments suggest that the unstructured N-terminal domain binds fibronectin. Analysis of changes in circular dichroism spectra of the N-terminal segment of BBK32 upon binding of the N-terminal domain of fibronectin revealed an increase in beta-sheet content in the complex. Hence, BBK32, which belongs to a different family of proteins and shows no overall sequence similarity with the fibronectin binding MSCRAMMs (microbial surface components recognizing adhesive matrix molecules) of Gram-positive bacteria, binds fibronectin by a mechanism that is reminiscent of the "tandem beta-zipper" previously demonstrated for the fibronectin binding of streptococcal adhesins. PMID- 15292205 TI - Interaction of the pacemaker channel HCN1 with filamin A. AB - Pacemaker channels are encoded by the HCN gene family and are responsible for a variety of cellular functions including control of spontaneous activity in cardiac myocytes and control of excitability in different types of neurons. Some of these functions require specific membrane localization. Although several voltage-gated channels are known to interact with intracellular proteins exerting auxiliary functions, no cytoplasmic proteins have been found so far to modulate HCN channels. Through the use of a yeast two-hybrid technique, here we showed that filamin A interacts with HCN1, an HCN isoform widely expressed in the brain, but not with HCN2 or HCN4. Filamin A is a cytoplasmic scaffold protein with actin binding domains whose main function is to link transmembrane proteins to the actin cytoskeleton. Using several HCN1 C-terminal constructs, we identified a filamin A-interacting region of 22 amino acids located downstream from the cyclic nucleotide-binding domain; this region is not conserved in HCN2, HCN3, or HCN4. We also verified by immunoprecipitation from bovine brain that the filamin A-HCN1 interaction is functional in vivo. In filamin A-expressing cells (filamin+), HCN1 (but not HCN4) channels were expressed in hot spots, whereas they were evenly distributed on the membrane of cells lacking filamin A (filamin-) indicating that interaction with filamin A affects membrane localization. Also, in filamin- cells the gating kinetics of HCN1 were strongly accelerated relative to filamin+ cells. The interaction with filamin A may contribute to localizing HCN1 channels to specific neuronal areas and to modulating channel activity. PMID- 15292206 TI - Mammary gland remodeling depends on gp130 signaling through Stat3 and MAPK. AB - The interleukin-6 (IL6) family of cytokines signals through the common receptor subunit gp130, and subsequently activates Stat3, MAPK, and PI3K. Stat3 controls cell death and tissue remodeling in the mouse mammary gland during involution, which is partially induced by IL6 and LIF. However, it is not clear whether Stat3 activation is mediated solely through the gp130 pathway or also through other receptors. This question was explored in mice carrying two distinct mutations in the gp130 gene; one that resulted in the complete ablation of gp130 and one that led to the loss of Stat3 binding sites (gp130Delta/Delta). Deletion of gp130 specifically from mammary epithelium resulted in a complete loss of Stat3 activity and resistance to tissue remodeling comparable to that seen in the absence of Stat3. A less profound delay of mammary tissue remodeling was observed in gp130Delta/Delta mice. Stat3 tyrosine and serine phosphorylation was still detected in these mice suggesting that Stat3 activation could be the result of gp130 interfacing with other receptors. Experiments in primary mammary epithelial cells and transfected COS-7 cells revealed a p44/42 MAPK and EGFR-dependent Stat3 activation. Moreover, the gp130-dependent EGFR activation was independent of EGF ligands, suggesting a cytoplasmic interaction and cross-talk between these two receptors. These experiments establish that two distinct Stat3 signaling pathways emanating from gp130 are utilized in mammary tissue. PMID- 15292207 TI - Bim is an apoptosis sensor that responds to loss of survival signals delivered by epidermal growth factor but not those provided by integrins. AB - Anoikis is a rapid apoptosis response that is initiated within a few minutes after inhibition of integrin signaling. In mammary epithelia, anoikis is mediated by subcellular translocation of Bax from the cytosol to mitochondria where it activates the intrinsic apoptosis pathway. The Bcl-2 homology 3 domain-only protein, Bim, has been proposed to have a key role in the apoptosis response of an epithelial cell line with reduced sensitivity to loss of integrin signaling, which undergoes apoptosis over a period of several days in suspension culture. Here we tested the involvement of Bim in the rapid anoikis response of mouse mammary epithelial cells and discovered that Bim does not have a role in detecting integrin-mediated signals. Instead Bim senses the loss of survival cues mediated by epidermal growth factor. Cell lines selected over many passages in culture have lost much of their sensitivity to anoikis signals arising from an altered cellular microenvironment and may undergo apoptosis through acquired mechanisms. PMID- 15292208 TI - Agonist-independent activation of Src tyrosine kinase by a cholecystokinin-2 (CCK2) receptor splice variant. AB - Src activity is elevated in a majority of colonic and pancreatic cancers and is associated with late stage aggressive cancers. However, the mechanisms leading to its increased activity remain largely undefined. Agonist binding to the cholecystokinin-2 (CCK2)/gastrin receptor (CCK2R), a G-protein-coupled receptor, increases Src activity in a variety of normal and neoplastic cell lines. Recently, we and others (Hellmich, M. R., Rui, X. L., Hellmich, H. L., Fleming, R. Y., Evers, B. M., and Townsend, C. M., Jr. (2000) J. Biol. Chem. 275, 32122 32128; Ding, W. Q., Kuntz, S. M., and Miller, L. J. (2002) Cancer Res. 62, 947 952; Smith, J. P., Verderame, M. F., McLaughlin, P., Martenis, M., Ballard, E., and Zagon, I. S. (2002) Int. J. Mol. Med. 10, 689-694) have identified a splice variant of CCK2R, called CCK2i4svR, that is expressed in human colorectal and pancreatic cancers but not by cells of the adjacent nonmalignant tissue. Compared with CCK2R, CCK2i4svR contains an additional 69 amino acids within its third intracellular loop (3il) domain. Because CCK2i4svR is the only splice variant expressed in some human colon and pancreatic cancers, we questioned whether CCK2i4svR could regulate Src activity. Stably transfected HEK293 cells were used because, unlike many cancer-derived cells, they have a low level of basal Src activity. We report that, in contrast to CCK2R, CCK2i4svR activates Src kinase in the absence of agonist stimulation. In vitro kinase assay of immunoprecipitated receptor protein showed a 6-8-fold increase in Src kinase activity associated with CCK2i4svR compared with CCK2R. Expression of the 3il domain of the CCK2i4svR alone was sufficient to partially activate Src kinase. Together, these data support the hypothesis that the increased Src activity observed in some pancreatic and colorectal cancers is due, in part, to the co-expression of CCK2i4svR. PMID- 15292209 TI - Loss of autoinhibition of the plasma membrane Ca(2+) pump by substitution of aspartic 170 by asparagin. A ctivation of plasma membrane calcium ATPase 4 without disruption of the interaction between the catalytic core and the C terminal regulatory domain. AB - The plasma membrane calcium ATPase (PMCA) actively transports Ca(2+) from the cytosol to the extra cellular space. The C-terminal segment of the PMCA functions as an inhibitory domain by interacting with the catalytic core. Ca(2+)-calmodulin binds to the C-terminal segment and stops inhibition. Here we showed that residue Asp(170), in the putative "A" domain of human PMCA isoform 4xb, plays a critical role in autoinhibition. In the absence of calmodulin a PMCA containing a site specific mutation of D170N had 80% of the maximum activity of the calmodulin activated PMCA and a similar high affinity for Ca(2+). The mutation did not change the activation of the PMCA by ATP. Deletion of the C-terminal segment further downstream of the calmodulin-binding site led to an additional increase in the maximal activity of the mutant, which suggests that the mutation did not affect the inhibition because of this portion of the C-terminal segment. The calmodulin-activated PMCA was more sensitive to vanadate inhibition than the autoinhibited enzyme. In contrast, inhibition of the D170N mutant required higher concentrations of vanadate and was not affected by calmodulin. Despite its higher basal activity, the mutant had an apparent affinity for calmodulin similar to that of the wild type enzyme, and its rate of proteolysis at the C-terminal segment was still calmodulin-dependent. Altogether these results suggest that activation by mutation D170N does not involve the displacement of the calmodulin binding autoinhibitory domain from the catalytic core and may arise directly from changes in the accessibility to the calcium-binding residues of the pump. PMID- 15292210 TI - Human Rad51C deficiency destabilizes XRCC3, impairs recombination, and radiosensitizes S/G2-phase cells. AB - The highly conserved Rad51 protein plays an essential role in repairing DNA damage through homologous recombination. In vertebrates, five Rad51 paralogs (Rad51B, Rad51C, Rad51D, XRCC2, and XRCC3) are expressed in mitotically growing cells and are thought to play mediating roles in homologous recombination, although their precise functions remain unclear. Among the five paralogs, Rad51C was found to be a central component present in two complexes, Rad51C-XRCC3 and Rad51B-Rad51C-Rad51D-XRCC2. We have shown previously that the human Rad51C protein exhibits three biochemical activities, including DNA binding, ATPase, and DNA duplex separation. Here we report the use of RNA interference to deplete expression of Rad51C protein in human HT1080 and HeLa cells. In HT1080 cells, depletion of Rad51C by small interfering RNA caused a significant reduction of frequency in homologous recombination. The level of XRCC3 protein was also sharply reduced in Rad51C-depleted HeLa cells, suggesting that XRCC3 is dependent for its stability upon heterodimerization with Rad51C. In addition, Rad51C depleted HeLa cells showed hypersensitivity to the DNA-cross-linking agent mitomycin C and moderately increased sensitivity to ionizing radiation. Importantly, the radiosensitivity of Rad51C-deficient HeLa cells was evident in S and G(2)/M phases of the cell cycle but not in G(1) phase. Together, these results provide direct cellular evidence for the function of human Rad51C in homologous recombinational repair. PMID- 15292211 TI - Functional characterization of sonic hedgehog mutations associated with holoprosencephaly. AB - Mutations of the developmental gene Sonic hedgehog (SHH) and alterations of SHH signaling have been associated with holoprosencephaly (HPE), a rare disorder characterized by a large spectrum of brain and craniofacial anomalies. Based on the crystal structure of mouse N-terminal and Drosophila C-terminal hedgehog proteins, we have developed three-dimensional models of the corresponding human proteins (SHH-N, SHH-C) that have allowed us to identify within these two domains crucial regions associated with HPE missense mutations. We have further characterized the functional consequences linked to 11 of these mutations. In transfected HEK293 cells, the production of the active SHH-N fragment was dramatically impaired for eight mutants (W117R, W117G, H140P, T150R, C183F, L271P, I354T, A383T). The supernatants from these cell cultures showed no significant SHH-signaling activity in a reporter cell-based assay. Two mutants (G31R, D222N) were associated with a lower production of SHH-N and signaling activity. Finally, one mutant harboring the A226T mutation displays an activity comparable with the wild-type protein. This work demonstrates that most of the HPE-associated SHH mutations analyzed have a deleterious effect on the availability of SHH-N and its biological activity. However, because of the lack of correlation between genotype and phenotype for SHH-associated mutations, our study suggests that other factors intervene in the development of the spectrum of HPE anomalies. PMID- 15292212 TI - Imperfect CAG repeats form diverse structures in SCA1 transcripts. AB - The expanded CAG repeat in the coding sequence of the spinocerebellar ataxia type 1 (SCA1) gene is responsible for SCA1, one of the hereditary human neurodegenerative diseases. In the normal SCA1 alleles usually 1-3 CAT triplets break the continuity of the CAG repeat tracts. Here we show what is the structural role of the CAU interruptions in the SCA1 transcripts. Depending on their number and localization within the repeat tract the interruptions either enlarge the terminal loop of the hairpin formed by the repeats, nucleate the internal loops in its stem structure, or force the repeats to fold into two smaller hairpins. Thus, the interruptions destabilize the CAG repeat hairpin, which is likely to decrease its ability to participate in the putative RNA pathogenesis mechanism driven by the long CAG repeat hairpins. PMID- 15292213 TI - The zinc finger motif of Escherichia coli RecQ is implicated in both DNA binding and protein folding. AB - The RecQ family of DNA helicases has been shown to be important for the maintenance of genomic integrity. Mutations in human RecQ genes lead to genomic instability and cancer. Several RecQ family of helicases contain a putative zinc finger motif of the C4 type at the C terminus that has been identified in the crystalline structure of RecQ helicase from Escherichia coli. To better understand the role of this motif in helicase from E. coli, we constructed a series of single mutations altering the conserved cysteines as well as other highly conserved residues. All of the resulting mutant proteins exhibited a high level of susceptibility to degradation, making functional analysis impossible. In contrast, a double mutant protein in which both cysteine residues Cys397 and Cys400 in the zinc finger motif were replaced by asparagine residues was purified to homogeneity. Slight local conformational changes were detected, but the rest of the mutant protein has a well defined tertiary structure. Furthermore, the mutant enzyme displayed ATP binding affinity similar to the wild-type enzyme but was severely impaired in DNA binding and in subsequent ATPase and helicase activities. These results revealed that the zinc finger binding motif is involved in maintaining the integrity of the whole protein as well as DNA binding. We also showed that the zinc atom is not essential to enzymatic activity. PMID- 15292214 TI - A new type of NADH dehydrogenase specific for nitrate respiration in the extreme thermophile Thermus thermophilus. AB - A four-gene operon (nrcDEFN) was identified within a conjugative element that allows Thermus thermophilus to use nitrate as an electron acceptor. Three of them encode homologues to components of bacterial respiratory chains: NrcD to ferredoxins; NrcF to iron-sulfur-containing subunits of succinate-quinone oxidoreductase (SQR); and NrcN to type-II NADH dehydrogenases (NDHs). The fourth gene, nrcE, encodes a membrane protein with no homologues in the protein data bank. Nitrate reduction with NADH was catalyzed by membrane fractions of the wild type strain, but was severely impaired in nrc::kat insertion mutants. A fusion to a thermophilic reporter gene was used for the first time in Thermus spp. to show that expression of nrc required the presence of nitrate and anoxic conditions. Therefore, a role for the nrc products as a new type of membrane NDH specific for nitrate respiration was deduced. Consistent with this, nrc::kat mutants grew more slowly than the wild type strain under anaerobic conditions, but not in the presence of oxygen. The oligomeric structure of this Nrc-NDH was deduced from the analysis of insertion mutants and a two-hybrid bacterial system. Attachment to the membrane of NrcD, NrcF, and NrcN was dependent on NrcE, whose cytoplasmic C terminus interacts with the three proteins. Interactions were also detected between NrcN and NrcF. Inactivation of nrcF produced solubilization of NrcN, but not of NrcD. These data lead us to conclude that the Nrc proteins form a distinct third type of bacterial respiratory NDH. PMID- 15292215 TI - A novel NADPH thioredoxin reductase, localized in the chloroplast, which deficiency causes hypersensitivity to abiotic stress in Arabidopsis thaliana. AB - Plants contain three thioredoxin systems. Chloroplast thioredoxins are reduced by ferredoxin-thioredoxin reductase, whereas the cytosolic and mitochondrial thioredoxins are reduced by NADPH thioredoxin reductase (NTR). There is high similarity among NTRs from plants, lower eukaryotes, and bacteria, which are different from mammal NTR. Here we describe the OsNTRC gene from rice encoding a novel NTR with a thioredoxin-like domain at the C terminus, hence, a putative NTR/thioredoxin system in a single polypeptide. Orthologous genes were found in other plants and cyanobacteria, but not in bacteria, yeast, or mammals. Full length OsNTRC and constructs with truncated NTR and thioredoxin domains were expressed in Escherichia coli as His-tagged polypeptides, and a polyclonal antibody specifically cross-reacting with the OsNTRC enzyme was raised. An in vitro activity assay showed that OsNTRC is a bifunctional enzyme with both NTR and thioredoxin activity but is not an NTR/thioredoxin system. Although the OsNTRC gene was expressed in roots and shoots of rice seedlings, the protein was exclusively found in shoots and mature leaves. Moreover, fractionation experiments showed that OsNTRC is localized to the chloroplast. An Arabidopsis NTRC knock-out mutant showed growth inhibition and hypersensitivity to methyl viologen, drought, and salt stress. These results suggest that the NTRC gene is involved in plant protection against oxidative stress. PMID- 15292216 TI - Role of ATP on the interaction of alpha-crystallin with its substrates and its implications for the molecular chaperone function. AB - ATP plays a significant role in the function of molecular chaperones of the large heat shock protein families. However, its role in the functions of chaperones of the small heat shock protein families is not understood very well. We report here a study on the role of ATP on the structure and function of the major eye lens chaperone alpha-crystallin. Our in vitro study shows that at physiological temperature, ATP induces the association of alpha-crystallin with substrate proteins. The association process is reversible and low affinity in nature with unit binding stoichiometry. 4,4'-Dianilino-1,1'-binaphthyl-5,5-disulfonic acid, dipotassium salt, binding studies show that ATP induces the exposure of additional hydrophobic sites on alpha-crystallin, but no appreciable enhancement of the same was observed for the substrate protein gamma-crystallin or carbonic anhydrase. An equilibrium unfolding study reveals that ATP at 3 mgm concentration stabilizes the alpha-crystallin structure by 4.5 kJ/mol. The compactness induced by ATP makes it more resistant to tryptic cleavage. ATP-induced association of chaperone alpha-crystallin with substrate enhanced its aggregation prevention ability and also enhanced the refolding yield of lactate dehydrogenase from the unfolded state. Our results suggest that the binding of ATP to alpha-crystallin and not its hydrolysis is required for all these effects, as replacement of ATP by its nonhydrolyzable analogue adenosine-5'-O-(3-thiotriphosphate), tetralithium salt, reproduced all the results faithfully. The implication of the ATP-induced reversible protein-protein association at physiological temperatures on the functional role of alpha-crystallin in vivo is discussed. PMID- 15292217 TI - A genetic screen for the identification of thiamin metabolic genes. AB - A genetic screen was developed for the identification of genes related to thiamin biosynthesis and degradation. Genes conferring resistance to bacimethrin or 4 amino-2-trifluoromethyl-5-hydroxymethylpyrimidine were selected from Escherichia coli and Bacillus subtilis genomic libraries. Hits from the selection included the known thiamin biosynthetic genes thiC, thiE, and dxs as well as five genes of previously unknown function (E. coli yjjX, yajO, ymfB, and cof and B. subtilis yveN). The gene products YmfB and Cof catalyze the hydrolysis of 4-amino-2-methyl 5-hydroxymethylpyrimidine pyrophosphate to 4-amino-2-methyl-5 hydroxymethylpyrimidine phosphate. YmfB also converts thiamin pyrophosphate into thiamin phosphate. PMID- 15292218 TI - Involvement of tumor necrosis factor receptor-associated protein 1 (TRAP1) in apoptosis induced by beta-hydroxyisovalerylshikonin. AB - beta-Hydroxyisovalerylshikonin (beta-HIVS), a compound isolated from the traditional oriental medicinal herb Lithospermum radix, is an ATP non-competitive inhibitor of protein-tyrosine kinases, such as v-Src and EGFR, and it induces apoptosis in various lines of human tumor cells. However, the way in which beta HIVS induces apoptosis remains to be clarified. In this study, we performed cDNA array analysis and found that beta-HIVS suppressed the expression of the gene for tumor necrosis factor receptor-associated protein 1 (TRAP1), which is a member of the heat-shock family of proteins. When human leukemia HL60 cells and human lung cancer DMS114 cells were treated with beta-HIVS, the amount of TRAP1 in mitochondria decreased in a time-dependent manner during apoptosis. A similar reduction in the level of TRAP1 was also observed upon exposure of cells to VP16. Treatment of DMS114 cells with TRAP1-specific siRNA sensitized the cells to beta HIVS-induced apoptosis. Moreover, the reduction in the level of expression of TRAP1 by TRAP1-specific siRNA enhanced the release of cytochrome c from mitochondria when DMS114 cells were treated with either beta-HIVS or VP16. The suppression of the level of TRAP1 by either beta-HIVS or VP16 was blocked by N acetyl-cysteine, indicating the involvement of reactive oxygen species (ROS) in the regulation of the expression of TRAP1. These results suggest that suppression of the expression of TRAP1 in mitochondria might play an important role in the induction of apoptosis caused via formation of ROS. PMID- 15292219 TI - The murine gastrin promoter is synergistically activated by transforming growth factor-beta/Smad and Wnt signaling pathways. AB - The transforming growth factor-beta (TGF-beta) and Wnt/wingless pathways play critical roles in the specification of cell fate during development and also contribute to cancer formation and progression. Whereas Wnt signaling is clearly pro-oncogenic, TGF-beta signaling is cell- and context-dependent, manifesting both inhibitory and proliferative effects. The growth factor, gastrin, has previously been shown to be a downstream target of the Wnt pathway and a promoter of gastrointestinal cancer. In this study, we show that the mouse gastrin promoter is regulated synergistically by TGF-beta/Smads and beta-catenin/T-cell factor (TCF). Co-transfection of Smad3/Smad4 and beta-catenin expression constructs synergistically activated mouse gastrin promoter activity 30-60-fold in AGS cells with minimal effect seen with either construct alone. This activation was further potentiated by TGF-beta1 treatment. Mutating either the TCF binding site or the Smad-binding element (SBE) diminished the activation of gastrin expression by Smad3/Smad4 and beta-catenin and led to a loss of gastrin promoter responsiveness to TGF-beta1 treatment. Wnt and TGF-beta regulated endogenous gastrin mRNA levels in AGS cells in a similar fashion, as revealed by small interference RNA studies or overexpression of Smads and TCF4/beta-catenin. Electrophoretic mobility shift assays and DNA affinity precipitation assays showed that the putative SBE and T-cell factor (TCF) sites were able to bind a complex containing Smads and beta-catenin/TCF4. In addition, the synergy between Smads and beta-catenin/TCF4 was dependent on CREB-binding protein (CBP)/P300, as demonstrated by overexpression of CBP or E1A. Moreover, by using a heterogeneous promoter reporter system, we showed that this complex containing Smads/TCF4/beta catenin complex was able to up-regulate transcription at isolated SBE or TCF sites. Thus, the Wnt signaling pathway is able to activate some target genes through its actions as a co-activator at non-TCF sites and has the potential to profoundly alter transcriptional responses to TGF-beta signaling. PMID- 15292220 TI - Ikappab kinase-beta (ikkbeta) modulation of epithelial sodium channel activity. AB - Using the yeast two-hybrid system, we identified a number of proteins that interacted with the carboxyl termini of murine epithelial sodium channel (ENaC) subunits. Initial screens indicated an interaction between the carboxyl terminus of beta-ENaC and IkappaB kinase-beta (IKKbeta), the kinase that phosphorylates Ikappabeta and results in nuclear targeting of NF-kappaB. A true two-hybrid reaction employing full-length IKKbeta and the carboxyl termini of all three subunits confirmed a strong interaction with beta-ENaC, a weak interaction with gamma-ENaC, and no interaction with alpha-ENaC. Co-immunoprecipitation studies for IKKbeta were performed in a murine cortical collecting duct cell line that endogenously expresses ENaC. Immunoprecipitation with beta-ENaC, but not gamma ENaC, resulted in co-immunoprecipitation of IKKbeta. To examine the direct effects of IKKbeta on ENaC activity, co-expression studies were performed using the two-electrode voltage clamp technique in Xenopus oocytes. Oocytes were injected with cRNAs for alphabetagamma-ENaC with or without cRNA for IKKbeta. Co injection of IKKbeta significantly increased the amiloride-sensitive current above controls. Using cell surface ENaC labeling, we determined that an increase of ENaC in the plasma membrane accounted for the increase in current. The injection of kinase-dead IKKbeta (K44A) in ENaC-expressing oocytes resulted in a significant decrease in current. Treatment of mpkCCD(c14) cells with aldosterone increased whole cell amounts of IKKbeta. Because this result suggested that aldosterone might activate NF-kappaB, mpkCCD(c14) cells were transiently transfected with a luciferase reporter gene responsive to NF-kappaB activation. Both aldosterone and tumor necrosis factor-alpha (TNFalpha) stimulation caused a similar and significant increase in luciferase activity as compared with controls. We conclude that IKKbeta interacts with ENaC by up-regulating ENaC at the plasma membrane and that the presence of IKKbeta is at very least permissive to ENaC function. These studies also suggest a previously unexpected interaction between the NF-kappaB transcription pathway and steroid regulatory pathways in epithelial cells. PMID- 15292221 TI - Isoforms of the polarity protein par6 have distinct functions. AB - PAR-6 is essential for asymmetric division of the Caenorhabditis elegans zygote. It is also critical for cell polarization in many other contexts throughout the Metazoa. The Par6 protein contains a PDZ domain and a partial CRIB (Cdc42/Rac interactive binding) domain, which mediate interactions with other polarity proteins such as Par3, Cdc42, Pals1, and Lgl. A family of mammalian Par6 isoforms (Par6A-D) has been described, but the significance of this diversification has been unclear. Here we demonstrate that Par6 family members localize differently when expressed in Madin-Darby canine kidney epithelial cells and have distinct effects on tight junction (TJ) assembly. Par6B localizes to the cytosol and inhibits TJ formation, but Par6A co-localizes predominantly with the TJ marker ZO 1 at cell-cell contacts and does not affect junctions. These functional differences correlate with differences in Pals1 binding; Par6B interacts strongly with Pals1, whereas Par6A binds weakly to Pals1 even in the presence of active Cdc42. Pals1 has a low affinity for the isolated CRIB-PDZ domain of Par6A, but analysis of chimeras showed that in addition Pals1 binding is blocked by an inhibitory property of the N terminus of Par6A. Unexpectedly, the localization of Par6A to cell-cell contacts is Cdc42-independent. PMID- 15292222 TI - Identification and characterization of zipper-interacting protein kinase as the unique vascular smooth muscle myosin phosphatase-associated kinase. AB - Excitation-contraction coupling in smooth muscle involves activation of myosin light chain (MLC) phosphorylation, which increases activity of the myosin actin activated ATPase, resulting in contraction. Phosphorylation of MLC phosphatase (SMPP-1M) by Rho-associated kinase or endogenous SMPP-1M-associated kinase inhibits SMPP-1M, enhancing MLC phosphorylation and contraction. However, the precise identity of SMPP-1M-associated kinase remains unclear. Biochemical evidence strongly supports the idea that SMPP-1M-associated kinase is related to the human serine/threonine leucine zipper-interacting protein kinase (hZIPK), which is important in cell apoptosis, and the SMPP-1M-associated kinase has therefore been called ZIP-like kinase (MacDonald, J. A., Borman, M. A., Murani, A., Somlyo, A. V., Hartshorne, D. J., and Haystead, T. A. J. (2001) Proc. Natl. Acad. Sci. U. S. A. 98, 2419-2424). Whether the vascular smooth muscle SMPP-1M associated kinase is a truncated version of hZIPK, native hZIPK, or a unique homologue of hZIPK is unclear. Here we show that only native hZIPK mRNA and protein are detectable in human vascular smooth muscle cells (VSMCs). High stringency screening of a human aortic cDNA library for the SMPP-1M-associated kinase identified 18 positive clones, all of which proved to be clones of hZIPK. PCR-based studies of VSMC RNA revealed native hZIPK transcripts but no evidence for splice variants of hZIPK or a ZIP-like kinase. Northern blotting studies of multiple vascular and non-vascular tissue RNAs, including human bladder RNA, showed only 2.3 kb of mRNA predicted for full-length hZIPK. Immunoblotting showed native full-length 52-kDa hZIPK expression in VSMCs. Full-length and N-terminal hZIPK bound the C-terminal domain (amino acids 681-847) of the myosin binding subunit (MBS) of SMPP-1M. hZIPK immunoprecipitated with the MBS of SMPP-1M and dominant negative RhoA inhibited the hZIPK-MBS interaction. These data identify hZIPK as the unique SMPP-1-associated kinase expressed in human vesicular smooth muscle and support a role for Rho in promoting the hZIPK-MBS interaction. PMID- 15292223 TI - The testicular form of hormone-sensitive lipase HSLtes confers rescue of male infertility in HSL-deficient mice. AB - Inactivation of the hormone-sensitive lipase gene (HSL) confers male sterility with a major defect in spermatogenesis. Several forms of HSL are expressed in testis. HSLtes mRNA and protein are found in early and elongated spermatids, respectively. The other forms are expressed in diploid germ cells and interstitial cells of the testis. To determine whether the absence of the testis specific form of HSL, HSLtes, was responsible for the infertility in HSL-null mice, we generated transgenic mice expressing HSLtes under the control of its own promoter. The transgenic animals were crossed with HSL-null mice to produce mice deficient in HSL in nongonadal tissues but expressing HSLtes in haploid germ cells. Cholesteryl ester hydrolase activity was almost completely blunted in HSL deficient testis. Mice with one allele of the transgene showed an increase in enzymatic activity and a small elevation in the production of spermatozoa. The few fertile hemizygous male mice produced litters of very small to small size. The presence of the two alleles led to a doubling in cholesteryl ester hydrolase activity, which represented 25% of the wild type values associated with a qualitatively normal spermatogenesis and a partial restoration of sperm reserves. The fertility of these mice was totally restored with normal litter sizes. In line with the importance of the esterase activity, HSLtes transgene expression reversed the cholesteryl ester accumulation observed in HSL-null mice. Therefore, expression of HSLtes and cognate cholesteryl ester hydrolase activity leads to a rescue of the infertility observed in HSL-deficient male mice. PMID- 15292224 TI - ExoS Rho GTPase-activating protein activity stimulates reorganization of the actin cytoskeleton through Rho GTPase guanine nucleotide disassociation inhibitor. AB - ExoS is a bifunctional Type III cytotoxin of Pseudomonas aeruginosa with N terminal Rho GTPase-activating protein (RhoGAP) and C-terminal ADP ribosyltransferase domains. Although the ExoS RhoGAP inactivates Cdc42, Rac, and RhoA in vivo, the relationship between ExoS RhoGAP and the eukaryotic regulators of Rho GTPases is not clear. The present study investigated the roles of Rho GTPase guanine nucleotide disassociation inhibitor (RhoGDI) in the reorganization of actin cytoskeleton mediated by ExoS RhoGAP. A green fluorescent protein-RhoGDI fusion protein was engineered and found to elicit actin reorganization through the inactivation of Rho GTPases. Green fluorescent protein-RhoGDI and ExoS RhoGAP cooperatively stimulated actin reorganization and translocation of Cdc42 from membrane to cytosol, and a RhoGDI mutant, RhoGDI(I177D), that is defective in extracting Rho GTPases off the membrane inhibited the actions of RhoGDI and ExoS RhoGAP on the translocation of Cdc42 from membrane to cytosol. A human RhoGDI small interfering RNA was transfected into HeLa cells to knock down 90% of the endogenous RhoGDI expression. HeLa cells with knockdown RhoGDI were resistant to the reorganization of the actin cytoskeleton elicited by type III-delivered ExoS RhoGAP. This indicates that ExoS RhoGAP and RhoGDI function in series to inactivate Rho GTPases, in which RhoGDI extracting GDP-bound Rho GTPases off the membrane and sequestering them in cytosol is the rate-limiting step in Rho GTPase inactivation. A eukaryotic GTPase-activating protein, p50RhoGAP, showed a similar cooperativity with RhoGDI on actin reorganization, suggesting that ExoS RhoGAP functions as a molecular mimic of eukaryotic RhoGAPs to inactivate Rho GTPases through RhoGDI. PMID- 15292225 TI - Inhibition of glucocorticoid receptor-mediated transcriptional activation by p38 mitogen-activated protein (MAP) kinase. AB - Tumor necrosis factor (TNF) promotes certain immune and inflammatory responses, whereas glucocorticoids exert immunosuppressive and anti-inflammatory actions. We show that TNF treatment produced a modest inhibition of glucocorticoid receptor (GR)-mediated transcriptional activation of a mouse mammary tumor virus (MMTV) promoter-driven luciferase construct in HeLa cells. The mitogen-activated protein (MAP) kinases, p38 and c-Jun N-terminal kinase (JNK), are important mediators of target gene activation by TNF, and JNK activation was earlier shown to inhibit GR mediated transcriptional activation by direct phosphorylation of GR at Ser-246. Transfection of HeLa cells with MKK6b(E), a constitutively active specific upstream activator of p38, led to a potent inhibition of GR activation of the MMTV promoter-driven luciferase construct. A similar inhibition of activation of the MMTV promoter-driven luciferase construct was seen in HeLa cells transfected with MKK7(D), a constitutively functional activator of JNK. Data from "domain swap" experiments using GR chimeras indicated that the main target of the p38 mediated (but not JNK-mediated) inhibition is the ligand-binding domain of GR (spanning amino acids 525-795), whereas the constitutively active N-terminal AF-1 region (spanning amino acids 106-237) is dispensable for the inhibitory effect of p38. We also demonstrate that activated p38 targets the GR ligand-binding domain indirectly. Suppression of GR function by activated p38 and JNK MAP kinases may be physiologically important as a mechanism of resistance to glucocorticoids seen in many patients with chronic inflammatory conditions. PMID- 15292226 TI - Ultraviolet A-induced modulation of Bcl-XL by p38 MAPK in human keratinocytes: post-transcriptional regulation through the 3'-untranslated region. AB - We examined the effect of inhibiting p38 MAPK on UVA-irradiated HaCaT cells, a spontaneously immortalized human keratinocyte cell line. Recent work from our laboratory has shown that UVA (250 kJ/m2) induces a rapid phosphorylation of p38 MAPK in the HaCaT cell line. Inhibition of p38 MAPK activity through the use of a specific inhibitor, SB202190, in combination with UVA treatment induced a rapid cleavage of caspase-9, caspase-8, and caspase-3, whereas UVA irradiation alone had no effect. Similarly, cleavage of the caspase substrate poly(ADP-ribose) polymerase was observed in UVA-irradiated HaCaT cells treated with SB202190 or in cells expressing a dominant-negative p38 MAPK. No effect of p38 MAPK inhibition upon caspase cleavage was observed in mock-irradiated HaCaT cells. In addition, increases in apoptosis were observed in UVA-irradiated cells treated with SB202190 by morphological analysis with no significant apoptosis occurring from UVA irradiation alone. Similar results were obtained by using normal human epidermal keratinocytes. UVA induced expression of the anti-apoptotic Bcl-2 family member, Bcl-XL, with abrogation of expression by using the p38 MAPK inhibitor SB202190. Overexpression of Bcl-XL prevented poly(ADP-ribose) polymerase cleavage induced by the combination of UVA and p38 MAPK inhibition. UVA enhanced the stability of Bcl-XL mRNA through increases in p38 MAPK activity. We determined that increases in UVA-induced expression of Bcl-XL occur through a posttranscriptional mechanism mediated by the 3'-untranslated region (UTR). We used Bcl-XL 3'-UTR luciferase constructs to determine the mechanism by which UVA increased Bcl-XL mRNA stability. Additionally, RNA binding studies indicate that UVA increases the binding of RNA-binding proteins to Bcl-XL 3'-UTR mRNA, which can be decreased by using SB202190. In conclusion, p38 MAPK and Bcl-XL expression play critical roles in the survival of UVA-irradiated HaCaT cells. PMID- 15292227 TI - Molecular mapping of the thrombin-heparin cofactor II complex. AB - We used 55 Ala-scanned recombinant thrombin molecules to define residues important for inhibition by the serine protease inhibitor (serpin) heparin cofactor II (HCII) in the absence and presence of glycosaminoglycans. We verified the importance of numerous basic residues in anion-binding exosite-1 (exosite-1) and found 4 additional residues, Gln24, Lys65, His66, and Tyr71 (using the thrombin numbering system), that were resistant to HCII inhibition with and without glycosaminoglycans. Inhibition rate constants for these exosite-1 (Q24A, K65A, H66A, Y71A) thrombin mutants (0.02-0.38 x 10(8) m(-1) min(-1) for HCII heparin when compared with 2.36 x 10(8) m(-1) min(-1) with wild-type thrombin and 0.03-0.53 x 10(8) m(-1) min(-1) for HCII-dermatan sulfate when compared with 5.23 x 10(8) m(-1) min(-1) with wild-type thrombin) confirmed that the structural integrity of thrombin exosite-1 is critical for optimal HCII-thrombin interactions in the presence of glycosaminoglycans. However, our results are also consistent for HCII-glycosaminoglycan-thrombin ternary complex formation. Ten residues surrounding the active site of thrombin were implicated in HCII interactions. Four mutants (Asp51, Lys52, Lys145/Thr147/Trp148, Asp234) showed normal increased rates of inhibition by HCII-glycosaminoglycans, whereas four mutants (Trp50, Glu202, Glu229, Arg233) remained resistant to inhibition by HCII with glycosaminoglycans. Using 11 exosite-2 thrombin mutants with 20 different mutated residues, we saw no major perturbations of HCII-glycosaminoglycan inhibition reactions. Collectively, our results support a "double bridge" mechanism for HCII inhibition of thrombin in the presence of glycosaminoglycans, which relies in part on ternary complex formation but is primarily dominated by an allosteric process involving contact of the "hirudin-like" domain of HCII with thrombin exosite-1. PMID- 15292228 TI - Human atherosclerotic intima and blood of patients with established coronary artery disease contain high density lipoprotein damaged by reactive nitrogen species. AB - High density lipoprotein (HDL) is the major carrier of lipid hydroperoxides in plasma, but it is not yet established whether HDL proteins are damaged by reactive nitrogen species in the circulation or artery wall. One pathway that generates such species involves myeloperoxidase (MPO), a major constituent of artery wall macrophages. Another pathway involves peroxynitrite, a potent oxidant generated in the reaction of nitric oxide with superoxide. Both MPO and peroxynitrite produce 3-nitrotyrosine in vitro. To investigate the involvement of reactive nitrogen species in atherogenesis, we quantified 3-nitrotyrosine levels in HDL in vivo. The mean level of 3-nitrotyrosine in HDL isolated from human aortic atherosclerotic intima was 6-fold higher (619 +/- 178 micromol/mol Tyr) than that in circulating HDL (104 +/- 11 micromol/mol Tyr; p < 0.01). Immunohistochemical studies demonstrated striking colocalization of MPO with epitopes reactive with an antibody to 3-nitrotyrosine. However, there was no significant correlation between the levels of 3-chlorotyrosine, a specific product of MPO, and those of 3-nitrotyrosine in lesion HDL. We also detected 3 nitrotyrosine in circulating HDL, and linear regression analysis demonstrated a strong correlation between the levels of 3-chlorotyrosine and levels of 3 nitrotyrosine. These observations suggest that MPO promotes the formation of 3 chlorotyrosine and 3-nitrotyrosine in circulating HDL but that other pathways also produce 3-nitrotyrosine in atherosclerotic tissue. Levels of HDL isolated from plasma of patients with established coronary artery disease contained twice as much 3-nitrotyrosine as HDL from plasma of healthy subjects, suggesting that nitrated HDL might be a marker for clinically significant vascular disease. The detection of 3-nitrotyrosine in HDL raises the possibility that reactive nitrogen species derived from nitric oxide might promote atherogenesis. Thus, nitrated HDL might represent a previously unsuspected link between nitrosative stress, atherosclerosis, and inflammation. PMID- 15292229 TI - Building the stator of the yeast vacuolar-ATPase: specific interaction between subunits E and G. AB - The vacuolar (H+)-ATPase (or V-ATPase) is a membrane protein complex that is structurally related to F1 and F0 ATP synthases. The V-ATPase is composed of an integral domain (V0) and a peripheral domain (V1) connected by a central stalk and up to three peripheral stalks. The number of peripheral stalks and the proteins that comprise them remain controversial. We have expressed subunits E and G in Escherichia coli as maltose binding protein fusion proteins and detected a specific interaction between these two subunits. This interaction was specific for subunits E and G and was confirmed by co-expression of the subunits from a bicistronic vector. The EG complex was characterized using size exclusion chromatography, cross-linking with short length chemical cross-linkers, circular dichroism spectroscopy, and electron microscopy. The results indicate a tight interaction between subunits E and G and revealed interacting helices in the EG complex with a length of about 220 angstroms. We propose that the V-ATPase EG complex forms one of the peripheral stators similar to the one formed by the two copies of subunit b in F-ATPase. PMID- 15292230 TI - Characterization of the distinct collagen binding, helicase and cleavage mechanisms of matrix metalloproteinase 2 and 14 (gelatinase A and MT1-MMP): the differential roles of the MMP hemopexin c domains and the MMP-2 fibronectin type II modules in collagen triple helicase activities. AB - Matrix metalloproteinase-2 (MMP-2, gelatinase A) and membrane type (MT)1-MMP (MMP 14) are cooperative dynamic components of a cell surface proteolytic axis involved in regulating the cellular signaling environment and pericellular collagen homeostasis. Although MT1-MMP exhibits type I collagenolytic but poor gelatinolytic activities, MMP-2 is a potent gelatinase with weak type I collagenolytic behavior. Recombinant linker/hemopexin C domain (LCD) of MT1-MMP binds native type I collagen, blocks MT1-MMP collagenolytic activity in trans, and by circular dichroism spectroscopy, induces localized structural perturbation in the collagen. These changes were reflected by enhanced cleavage of the MT1-LCD bound collagen by the collagenases MMP-1 and MMP-8 but not by trypsin or MMP-7. Thus, the MT1-LCD alone can initiate triple helicase activity. In contrast, the native and denatured collagen binding properties of MMP-2 reside in the fibronectin type II modules, accordingly termed the collagen binding domain (CBD). Recombinant CBD (but not the MMP-2 LCD) also changed the circular dichroism spectra leading to increased MMP-1 and -8 cleavage of native collagen. However, recombinant CBD reduced gelatin and collagen cleavage by MMP-2 in trans as did CBD23, which comprises the second and third fibronectin type II modules, but not the CBD23 mutant W316A/W374A, which neither binds gelatin nor collagen. This indicates that MMP-2 and MT1-MMP bind collagen at a different site than MMP 1 and MMP-8. Thus, MMP-2 utilizes the CBD in cis for collagen binding and triple helicase activity, which compensates for the lack of collagen binding by the MMP 2 LCD. Hence, the MMP family has evolved two distinct mechanisms for collagen triple helicase activity using two structurally distinct domains, with triple helicase activity occurring independent of alpha-chain hydrolysis. PMID- 15292231 TI - Regulation of Swi6/HP1-dependent heterochromatin assembly by cooperation of components of the mitogen-activated protein kinase pathway and a histone deacetylase Clr6. AB - A study of gene silencing within the mating-type region of fission yeast defines two distinct pathways responsible for the establishment of heterochromatin assembly. One is RNA interference-dependent and acts on centromere-homologous repeats (cenH). The other is a stochastic Swi6 (the fission yeast HP1 homolog) dependent mechanism that is not fully understood. Here we find that activating transcription factor (Atf1) and Pcr1, the fission yeast bZIP transcription factors homologous to human ATF-2, are crucial for proper histone deacetylation of both H3 and H4. This deacetylation is a prerequisite for subsequent H3 lysine 9 methylation and Swi6-dependent heterochromatin assembly across the rest of the silent mating-type (mat) region lacking the RNA interference-dependent cenH repeat. Moreover, Atf1 and Pcr1 can form complexes with both a histone deacetylase, Clr6, and Swi6, and clr6 mutations affected the H3/H4 acetylation patterns, similar to the atf1 and pcr1 deletion mutant phenotypes at the endogenous mat loci and at the ctt1+ promoter region surrounding ATF/CRE-binding site. These data suggest that Atf1 and Pcr1 participate in an early step essential for heterochromatin assembly at the mat locus and silencing of transcriptional targets of Atf1. Furthermore, a phosphorylation event catalyzed by the conserved mitogen-activated protein kinase pathway is important for regulation of heterochromatin silencing by Atf1 and Pcr1. These findings suggest a role for the mitogen-activated protein kinase pathway and histone deacetylase in Swi6-based heterochromatin assembly. PMID- 15292232 TI - Dopamine receptor-mediated Ca(2+) signaling in striatal medium spiny neurons. AB - Inositol 1,4,5-trisphosphate (InsP(3)) and cAMP are the two second messengers that play an important role in neuronal signaling. Here, we investigated the interactions of InsP(3)- and cAMP-mediated signaling pathways activated by dopamine in striatal medium spiny neurons (MSN). We found that in approximately 40% of the MSN, application of dopamine elicited robust repetitive Ca(2+) transients (oscillations). In pharmacological experiments with specific agonists and antagonists, we found that the observed Ca(2+) oscillations were triggered by activation of D1 class dopamine receptors (DARs). We further demonstrated that activation of phospholipase C was required for induction of dopamine-induced Ca(2+) oscillations and that maintenance of dopamine-evoked Ca(2+) oscillations required both Ca(2+) influx and Ca(2+) mobilization from internal Ca(2+) stores. In "priming" experiments with a type 2 5-hydroxytryptamine receptor agonist, we have shown a likely role for calcyon in coupling D1 class DARs with Ca(2+) oscillations in MSN. In experiments with the DAR-specific agonist SKF83959, we discovered that phospholipase C activation alone could not account for dopamine induced Ca(2+) oscillations. We further demonstrated that direct activation of protein kinase A by 8-bromo-cAMP or inhibition of protein phosphatase-1 (PP1) or calcineurin (PP2B) resulted in elevation of basal Ca(2+) levels in MSN, but not in Ca(2+) oscillations. In experiments with competitive peptides, we have shown an importance of type 1 InsP(3) receptor association with PP1alpha and with AKAP9.protein kinase A for dopamine-induced Ca(2+) oscillations. In experiments with MSN from DARPP-32 knock-out mice, we demonstrated a regulatory role of DARPP 32 in dopamine-induced Ca(2+) oscillations. Our results indicate that, following D1 class DAR activation, InsP(3) and cAMP signaling pathways converge on the type 1 InsP(3) receptor, resulting in Ca(2+) oscillations in MSN. PMID- 15292233 TI - The POU factor Oct-25 regulates the Xvent-2B gene and counteracts terminal differentiation in Xenopus embryos. AB - The Xvent-2B promoter is regulated by a BMP-2/4-induced transcription complex comprising Smad signal transducers and specific transcription factors. Using a yeast one-hybrid screen we have found that Oct-25, a Xenopus POU domain protein related to mammalian Oct-3/4, binds as an additional factor to the Xvent-2B promoter. This interaction was further confirmed by both in vitro and in vivo analyses. The Oct-25 gene is mainly transcribed during blastula and gastrula stages in the newly forming ectodermal and mesodermal germ layers. Luciferase reporter gene assay demonstrated that Oct-25 stimulates transcription of the Xvent-2B gene. This stimulation depends on the Oct-25 binding site and the bone morphogenetic protein-responsive element. Furthermore, Oct-25 interacts in vitro with components of the Xvent-2B transcription complex, like Smad1/4 and Xvent-2. Overexpression of Oct-25 results in anterior/posterior truncations and lack of differentiation for neuroectoderm- and mesoderm-derived tissues including blood cells. This effect is consistent with an evolutionarily conserved role of class V POU factors in the maintenance of an undifferentiated cell state. In Xenopus, the molecular mechanism underlying this process might be coupled to the expression of Xvent proteins. PMID- 15292234 TI - The cblD defect causes either isolated or combined deficiency of methylcobalamin and adenosylcobalamin synthesis. AB - Intracellular cobalamin is converted to adenosylcobalamin, coenzyme for methylmalonyl-CoA mutase and to methylcobalamin, coenzyme for methionine synthase, in an incompletely understood sequence of reactions. Genetic defects of these steps are defined as cbl complementation groups of which cblC, cblD (described in only two siblings), and cblF are associated with combined homocystinuria and methylmalonic aciduria. Here we describe three unrelated patients belonging to the cblD complementation group but with distinct biochemical phenotypes different from that described in the original cblD siblings. Two patients presented with isolated homocystinuria and reduced formation of methionine and methylcobalamin in cultured fibroblasts, defined as cblD-variant 1, and one patient with isolated methylmalonic aciduria and deficient adenosylcobalamin synthesis in fibroblasts, defined as cblD-variant 2. Cell lines from the cblD-variant 1 patients clearly complemented reference lines with the same biochemical phenotype, i.e. cblE and cblG, and the cblD-variant 2 cell line complemented cells from the mutant classes with isolated deficiency of adenosylcobalamin synthesis, i.e. cblA and cblB. Also, no pathogenic sequence changes in the coding regions of genes associated with the respective biochemical phenotypes were found. These findings indicate heterogeneity within the previously defined cblD mutant class and point to further complexity of intracellular cobalamin metabolism. PMID- 15292235 TI - Severe hypoalphalipoproteinemia in mice expressing human hepatic lipase deficient in binding to heparan sulfate proteoglycan. AB - Unlike human hepatic lipase (hHL) that is mainly cell surface-anchored via binding to heparan sulfate proteoglycans (HSPG), mouse HL (mHL) has a low affinity to HSPG and thus is largely blood-borne. The reduced HSPG binding of mHL is attributable to the C-terminal amino acids. To determine the functions of HSPG binding of hHL in vivo, we created adenovirus vectors encoding hHL or a chimeric protein (designated hHLmt) in which the C-terminal HSPG-binding sequences were replaced with the corresponding mouse sequences. Injecting hHLmt-expressing virus into C57BL/6J mice (1.8 x 10(10) virus particles/mouse) resulted in a 3-fold increase in pre-heparin HL activity, whereas infection with an identical dose of hHL virus did not change pre-heparin HL activity. In hHLmt-expressing mice, the concentration of total cholesterol and phospholipids was inversely related to the hHL activity in pre-heparin plasma in a dose- and time-dependent manner, and the decrease was mainly attributable to high density lipoproteins (HDL) cholesterol and HDL phospholipids. The expression of hHL exhibited no change in plasma total cholesterol or phospholipid levels as compared with control mice infected with luciferase or injected with saline. The reduced HDL lipids in the hHLmt expressing mice were accompanied by markedly decreased plasma and hepatic apolipoprotein (apo) A-I. In primary hepatocytes isolated from hHLmt-expressing mice, the concentration of cell-associated and secreted apoA-I was decreased by 2 3-fold as compared with hepatocytes isolated from control mice, whereas the levels of apoB and apoE were unaltered. Infection of primary hepatocytes with hHLmt virus ex vivo also resulted in reduced apoA-I secretion but had no effect on cell-associated apoA-I. These results suggest that expression of HSPG binding deficient hHL has a profound HDL-lowering effect. PMID- 15292236 TI - Hsp105alpha suppresses Hsc70 chaperone activity by inhibiting Hsc70 ATPase activity. AB - Hsp105alpha is a mammalian member of the HSP105/110 family, a diverged subgroup of the HSP70 family. Hsp105alpha associates with Hsp70/Hsc70 as complexes in vivo and regulates the chaperone activity of Hsp70/Hsc70 negatively in vitro and in vivo. In this study, we examined the mechanisms by which Hsp105alpha regulates Hsc70 chaperone activity. Using a series of deletion mutants of Hsp105alpha and Hsc70, we found that the interaction between Hsp105alpha and Hsc70 was necessary for the suppression of Hsc70 chaperone activity by Hsp105alpha. Furthermore, Hsp105alpha and deletion mutants of Hsp105alpha that interacted with Hsc70 suppressed the ATPase activity of Hsc70, with the concomitant appearance of ATPase activity of Hsp105alpha. As the ATPase activity of Hsp70/Hsc70 is essential for the efficient folding of nonnative protein substrates, Hsp105alpha is suggested to regulate the substrate binding cycle of Hsp70/Hsc70 by inhibiting the ATPase activity of Hsp70/Hsc70, thereby functioning as a negative regulator of the Hsp70/Hsc70 chaperone system. PMID- 15292237 TI - Dual engagement regulation of protein interactions with the AP-2 adaptor alpha appendage. AB - Clathrin-mediated endocytosis depends upon the coordinated assembly of a large number of discrete clathrin coat components to couple cargo selection with rapid internalization from the cell surface. Accordingly, the heterotetrameric AP-2 adaptor complex binds not only to clathrin and select cargo molecules, but also to an extensive family of endocytic accessory factors and alternate sorting adaptors. Physical associations between accessory proteins and AP-2 occur primarily through DP(F/W) or FXDXF motifs, which engage an interaction surface positioned on the C-terminal platform subdomain of the independently folded alpha subunit appendage. Here, we find that the WXX(F/W)X(D/E) interaction motif found in several endocytic proteins, including synaptojanin 1, stonin 2, AAK1, GAK, and NECAP1, binds a second interaction site on the bilobal alpha appendage, located on the N-terminal beta sandwich subdomain. Both alpha appendage binding sites can be engaged synchronously, and our data reveal that varied assemblies of interaction motifs with different affinities for two sites upon the alpha appendage can provide a mechanism for temporal ordering of endocytic accessory proteins during clathrin-mediated endocytosis. PMID- 15292238 TI - Role of regulator of G protein signaling 2 (RGS2) in Ca(2+) oscillations and adaptation of Ca(2+) signaling to reduce excitability of RGS2-/- cells. AB - Regulators of G protein signaling (RGS) proteins accelerate the GTPase activity of Galpha subunits to determine the duration of the stimulated state and control G protein-coupled receptor-mediated cell signaling. RGS2 is an RGS protein that shows preference toward Galpha(q). To better understand the role of RGS2 in Ca(2+) signaling and Ca(2+) oscillations, we characterized Ca(2+) signaling in cells derived from RGS2(-/-) mice. Deletion of RGS2 modified the kinetic of inositol 1,4,5-trisphosphate (IP(3)) production without affecting the peak level of IP(3), but rather increased the steady-state level of IP(3) at all agonist concentrations. The increased steady-state level of IP(3) led to an increased frequency of [Ca(2+)](i) oscillations. The cells were adapted to deletion of RGS2 by reducing Ca(2+) signaling excitability. Reduced excitability was achieved by adaptation of all transporters to reduce Ca(2+) influx into the cytosol. Thus, IP(3) receptor 1 was down-regulated and IP(3) receptor 3 was up-regulated in RGS2(-/-) cells to reduce the sensitivity for IP(3) to release Ca(2+) from the endoplasmic reticulum to the cytosol. Sarco/endoplasmic reticulum Ca(2+) ATPase 2b was up-regulated to more rapidly remove Ca(2+) from the cytosol of RGS2(-/-) cells. Agonist-stimulated Ca(2+) influx was reduced, and Ca(2+) efflux by plasma membrane Ca(2+) was up-regulated in RGS2(-/-) cells. The result of these adaptive mechanisms was the reduced excitability of Ca(2+) signaling, as reflected by the markedly reduced response of RGS2(-/-) cells to changes in the endoplasmic reticulum Ca(2+) load and to an increase in extracellular Ca(2+). These findings highlight the central role of RGS proteins in [Ca(2+)](i) oscillations and reveal a prominent plasticity and adaptability of the Ca(2+) signaling apparatus. PMID- 15292239 TI - Defects in cell adhesion and the visceral endoderm following ablation of nonmuscle myosin heavy chain II-A in mice. AB - Previous work has shown that ablation or mutation of nonmuscle myosin heavy chain II-B (NMHC II-B) in mice results in defects in the heart and brain with death occurring between embryonic day 14.5 (E14.5) and birth (Tullio, A. N., Accili, D., Ferrans, V. J., Yu, Z. X., Takeda, K., Grinberg, A., Westphal, H., Preston, Y. A., and Adelstein, R. S. (1997) Proc. Natl. Acad. Sci. U. S. A. 94, 12407 12412). Here we show that mice ablated for NMHC II-A fail to develop a normal patterned embryo with a polarized visceral endoderm by E6.5 and die by E7.5. Moreover, A(-)/A(-) embryoid bodies grown in suspension culture constantly shed cells. These defects in cell adhesion and tissue organization are explained by loss of E-cadherin and beta-catenin localization to cell adhesion sites in both cell culture and in the intact embryos. The defects can be reproduced by introducing siRNA directed against NMHC II-A into wild-type embryonic stem cells. Our results suggest an essential role for a single, specific nonmuscle myosin isoform in maintaining cell-cell adhesions in the early mammalian embryo. PMID- 15292240 TI - Regulation of the GTPase cycle in post-translational signal recognition particle based protein targeting involves cpSRP43. AB - The chloroplast signal recognition particle consists of a conserved 54-kDa GTPase and a novel 43-kDa chromodomain protein (cpSRP43) that together bind light harvesting chlorophyll a/b-binding protein (LHCP) to form a soluble targeting complex that is subsequently directed to the thylakoid membrane. Homology-based modeling of cpSRP43 indicates the presence of two previously identified chromodomains along with a third N-terminal chromodomain. Chromodomain deletion constructs were used to examine the role of each chromodomain in mediating distinct steps in the LHCP localization mechanism. The C-terminal chromodomain is completely dispensable for LHCP targeting/integration in vitro. The central chromodomain is essential for both targeting complex formation and integration because of its role in binding the M domain of cpSRP54. The N-terminal chromodomain (CD1) is unnecessary for targeting complex formation but is required for integration. This correlates with the ability of CD1 along with the ankyrin repeat region of cpSRP43 to regulate the GTPase cycle of the cpSRP-receptor complex. PMID- 15292241 TI - Calcium/calmodulin up-regulates a cytoplasmic receptor-like kinase in plants. AB - Calcium/calmodulin-dependent kinases play an important role in protein phosphorylation in eukaryotes. However, not much is known about calcium/calmodulin-dependent protein phosphorylation and its role in signal transduction in plants. By using a protein-protein interaction-based approach, we have isolated a novel plant-specific calmodulin-binding receptor-like cytoplasmic kinase (CRCK1) from Arabidopsis thaliana, as well as its ortholog from Medicago sativa (alfalfa). CRCK1 does not show high homology to calcium/calmodulin dependent protein kinases in animals. In contrast, it shows high homology in the kinase domain to serine/threonine receptor-like kinases in plants. However, it contains neither a transmembrane domain nor an extracellular domain. Calmodulin binds to CRCK1 in a calcium-dependent manner with an affinity of approximately 20.5 nm. The calmodulin-binding site in CRCK1 is located in amino acids 160-183, which overlap subdomain II of the kinase domain. CRCK1 undergoes autophosphorylation in the presence of Mg2+ at the threonine residue(s). The Km and Vmax values of CRCK1 for ATP are 1 microm and 33.6 pmol/mg/min, respectively. Calcium/calmodulin stimulates the kinase activity of CRCK1, which increases the Vmax of CRCK1 approximately 9-fold. The expression of CRCK1 is increased in response to stresses such as cold and salt and stress molecules such as abscisic acid and hydrogen peroxide. These results indicate the presence of a calcium/calmodulin-regulated receptor-like cytoplasmic kinase in plants. Furthermore, these results also suggest that calcium/calmodulin-regulated protein phosphorylation involving CRCK1 plays a role in stress signal transduction in plants. PMID- 15292242 TI - Formation of D-tyrosyl-tRNATyr accounts for the toxicity of D-tyrosine toward Escherichia coli. AB - D-Tyr-tRNATyr deacylase cleaves the ester bond between a tRNA molecule and a D amino acid. In Escherichia coli, inactivation of the gene (dtd) encoding this deacylase increases the toxicity of several D-amino acids including D-tyrosine, D tryptophan, and D-aspartic acid. Here, we demonstrate that, in a Deltadtd cell grown in the presence of 2.4 mm D-tyrosine, approximately 40% of the total tRNATyr pool is converted into D-Tyr-tRNATyr. No D-Tyr-tRNATyr is observed in dtd+ cells. In addition, we observe that overproduction of tRNATyr, tRNATrp, or tRNAAsp protects a Deltadtd mutant strain against the toxic effect of D-tyrosine, D-tryptophan, or D-aspartic acid, respectively. In the case of D-tyrosine, we show that the protection is accounted for by an increase in the concentration of L-Tyr-tRNATyr proportional to that of overproduced tRNATyr. Altogether, these results indicate that, by accumulating in vivo, high amounts of D-Tyr-tRNATyr cause a starvation for L-Tyr-tRNATyr. The deacylase prevents the starvation by hydrolyzing D-Tyr-tRNATyr. Overproduction of tRNATyr also relieves the starvation by increasing the amount of cellular L-Tyr-tRNATyr available for translation. PMID- 15292243 TI - Evaluation of the contribution of different ADAMs to tumor necrosis factor alpha (TNFalpha) shedding and of the function of the TNFalpha ectodomain in ensuring selective stimulated shedding by the TNFalpha convertase (TACE/ADAM17). AB - Tumor necrosis factor-alpha (TNFalpha), a potent pro-inflammatory cytokine, is released from cells by proteolytic cleavage of a membrane-anchored precursor. The TNF-alpha converting enzyme (TACE; a disintegrin and metalloprotease17; ADAM17) is known to have a key role in the ectodomain shedding of TNFalpha in several cell types. However, because purified ADAMs 9, 10, and 19 can also cleave a peptide corresponding to the TNFalpha cleavage site in vitro, these enzymes are considered to be candidate TNFalpha sheddases as well. In this study we used cells lacking ADAMs 9, 10, 17 (TACE), or 19 to address the relative contribution of these ADAMs to TNFalpha shedding in cell-based assays. Our results corroborate that ADAM17, but not ADAM9, -10, or -19, is critical for phorbol ester- and pervanadate-stimulated release of TNFalpha in mouse embryonic fibroblasts. However, overexpression of ADAM19 increased the constitutive release of TNFalpha, whereas overexpression of ADAM9 or ADAM10 did not. This suggests that ADAM19 may contribute to TNFalpha shedding, especially in cells or tissues where it is highly expressed. Furthermore, we used mutagenesis of TNFalpha to explore which domains are important for its stimulated processing by ADAM17. We found that the cleavage site of TNFalpha is necessary and sufficient for cleavage by ADAM17. In addition, the ectodomain of TNFalpha makes an unexpected contribution to the selective cleavage of TNFalpha by ADAM17: it prevents one or more other enzymes from cleaving TNFalpha following PMA stimulation. Thus, selective stimulated processing of TNFalpha by ADAM17 in cells depends on the presence of an appropriate cleavage site as well as the inhibitory role of the TNF ectodomain toward other enzymes that can process this site. PMID- 15292244 TI - Detection and mapping of widespread intermolecular protein disulfide formation during cardiac oxidative stress using proteomics with diagonal electrophoresis. AB - Regulation of protein function by reversible cysteine-targeted oxidation can be achieved by multiple mechanisms, such as S-glutathiolation, S-nitrosylation, sulfenic acid, sulfinic acid, and sulfenyl amide formation, as well as intramolecular disulfide bonding of vicinal thiols. Another cysteine oxidation state with regulatory potential involves the formation of intermolecular protein disulfides. We utilized two-dimensional sequential non-reducing/reducing SDS-PAGE (diagonal electrophoresis) to investigate intermolecular protein disulfide formation in adult cardiac myocytes subjected to a series of interventions (hydrogen peroxide, S-nitroso-N-acetylpenicillamine, doxorubicin, simulated ischemia, or metabolic inhibition) that alter the redox status of the cell. More detailed experiments were undertaken with the thiol-specific oxidant diamide (5 mm), a concentration that induces a mild non-injurious oxidative stress. This increase in cellular oxidation potential caused global intermolecular protein disulfide formation in cytosolic, membrane, and myofilament/cytoskeletal compartments. A large number of proteins that undergo these associations were identified using liquid chromatography-mass spectrometry/mass spectrometry. These associations, which involve metabolic and antioxidant enzymes, structural proteins, signaling molecules, and molecular chaperones, were confirmed by assessing "shifts" on non-reducing immunoblots. The observation of widespread protein-protein disulfides indicates that these oxidative associations are likely to be fundamental in how cells respond to redox changes. PMID- 15292245 TI - Characterization of C3a and C5a receptors in rat cerebellar granule neurons during maturation. Neuroprotective effect of C5a against apoptotic cell death. AB - There is now clear evidence that the Complement anaphylatoxin C3a and C5a receptors (C3aR and C5aR) are expressed in glial cells, notably in astrocytes and microglia. In contrast, very few data are available concerning the possible expression of these receptors in neurons. Here, we show that transient expression of C3aR and C5aR occurs in cerebellar granule neurons in vivo with a maximal density in 12-day-old rat, suggesting a role of these receptors during development of the cerebellum. Expression of C3aR and C5aR mRNAs and proteins was also observed in vitro in cultured cerebellar granule cells. Quantification of the mRNAs by real-time reverse transcription-PCR showed a peak of expression at day 2 in vitro (DIV 2); the C3aR and C5aR proteins were detected by Western blot analysis at DIV 4 and by flow cytometry and immunocytochemistry in differentiating neurons with a maximum density at DIV 4-9. Apoptosis of granule cells plays a crucial role for the harmonious development of the cerebellar cortex. We found that, in cultured granule neurons in which apoptosis was induced by serum deprivation and low potassium concentration, a C5aR agonist promoted cell survival and inhibited caspase-3 activation and DNA fragmentation. The neuroprotective effect of the C5aR agonist was associated with a marked inhibition of caspase-9 activity and partial restoration of mitochondrial integrity. Our results provide the first evidence that C3aR and C5aR are both expressed in cerebellar granule cells during development and that C5a, but not C3a, is a potent inhibitor of apoptotic cell death in cultured granule neurons. PMID- 15292246 TI - Structural features of microRNA (miRNA) precursors and their relevance to miRNA biogenesis and small interfering RNA/short hairpin RNA design. AB - We have established the structures of 10 human microRNA (miRNA) precursors using biochemical methods. Eight of these structures turned out to be different from those that were computer-predicted. The differences localized in the terminal loop region and at the opposite side of the precursor hairpin stem. We have analyzed the features of these structures from the perspectives of miRNA biogenesis and active strand selection. We demonstrated the different thermodynamic stability profiles for pre-miRNA hairpins harboring miRNAs at their 5'- and 3'-sides and discussed their functional implications. Our results showed that miRNA prediction based on predicted precursor structures may give ambiguous results, and the success rate is significantly higher for the experimentally determined structures. On the other hand, the differences between the predicted and experimentally determined structures did not affect the stability of termini produced through "conceptual dicing." This result confirms the value of thermodynamic analysis based on mfold as a predictor of strand section by RNAi induced silencing complex (RISC). PMID- 15292247 TI - MiRP1 modulates HCN2 channel expression and gating in cardiac myocytes. AB - MinK-related protein (MiRP1 or KCNE2) interacts with the hyperpolarization activated, cyclic nucleotide-gated (HCN) family of pacemaker channels to alter channel gating in heterologous expression systems. Given the high expression levels of MiRP1 and HCN subunits in the cardiac sinoatrial node and the contribution of pacemaker channel function to impulse initiation in that tissue, such an interaction could be of considerable physiological significance. However, the functional evidence for MiRP1/HCN interactions in heterologous expression studies has been accompanied by inconsistencies between studies in terms of the specific effects on channel function. To evaluate the effect of MiRP1 on HCN expression and function in a physiological context, we used an adenovirus approach to overexpress a hemagglutinin (HA)-tagged MiRP1 (HAMiRP1) and HCN2 in neonatal rat ventricular myocytes, a cell type that expresses both MiRP1 and HCN2 message at low levels. HA-MiRP1 co-expression with HCN2 resulted in a 4-fold increase in maximal conductance of pacemaker currents compared with HCN2 expression alone. HCN2 activation and deactivation kinetics also changed, being significantly more rapid for voltages between -60 and -95 mV when HA-MiRP1 was co expressed with HCN2. However, the voltage dependence of activation was not affected. Co-immunoprecipitation experiments demonstrated that expressed HA-MiRP1 and HCN2, as well as endogenous MiRP1 and HCN2, co-assemble in ventricular myocytes. The results indicate that MiRP1 acts as a beta subunit for HCN2 pacemaker channel subunits and alters channel gating at physiologically relevant voltages in cardiac cells. PMID- 15292248 TI - Early loss of E-cadherin from cell-cell contacts is involved in the onset of Anoikis in enterocytes. AB - Anoikis, i.e. apoptosis induced by detachment from the extracellular matrix, is thought to be involved in the shedding of enterocytes at the tip of intestinal villi. Mechanisms controlling enterocyte survival are poorly understood. We investigated the role of E-cadherin, a key protein of cell-cell adhesion, in the control of anoikis of normal intestinal epithelial cells, by detaching murine villus epithelial cells from the underlying basement membrane while preserving cell-cell interactions. We show that upon the loss of anchorage, normal enterocytes execute a program of apoptosis within minutes, via a Bcl-2-regulated and caspase-9-dependent pathway. E-cadherin is lost early from cell-cell contacts. This process precedes the execution phase of detachment-induced apoptosis as it is only weakly modulated by Bcl-2 overexpression or caspase inhibition. E-cadherin loss, however, is efficiently prevented by lysosome and proteasome inhibitors. We also found that a blocking anti-E-cadherin antibody increases the rate of anoikis, whereas the activation of E-cadherin using E cadherin-Fc chimera proteins reduces anoikis. In conclusion, our results stress the striking sensitivity of normal enterocytes to the loss of anchorage and the contribution of E-cadherin to the control of their survival/apoptosis balance. They open new perspectives on the key role of this protein, which is dysregulated in the intestinal epithelium in both inflammatory bowel disease and cancer. PMID- 15292249 TI - In rat hepatocytes glucagon increases mammalian target of rapamycin phosphorylation on serine 2448 but antagonizes the phosphorylation of its downstream targets induced by insulin and amino acids. AB - The major function of mammalian target of rapamycin (mTOR) is the control of cell growth. Insulin and amino acids regulate the mTOR pathway, and both are needed to promote its maximal activation. To further understand mTOR regulation by insulin and amino acids, we have studied the enzyme in primary cultures of hepatocytes. We show that insulin increases mTOR phosphorylation on Ser2448, a consensus phosphorylation site for protein kinase B (PKB). Ser2448 phosphorylation is also increased by amino acids, although they do not activate PKB. Furthermore, insulin and amino acids have an additive effect, indicating that they act through distinct pathways. We also show that phosphorylation of Ser2448 does not seem to modulate in vitro phosphorylation of eukaryotic initiation factor 4E-binding protein 1 by mTOR. However, stimulation of hepatocytes with insulin and amino acids leads to an increase in mTOR kinase activity. Rapamycin has no effect on insulin-, glucagon-, and 8-(4-chlorophenylthio)adenosine-cAMP-induced amino acid transport. Surprisingly, glucagon and 8-(4-chlorophenylthio)adenosine-cAMP, which do not activate PKB, stimulate the phosphorylation on Ser2448 of mTOR. However, glucagon inhibits amino acid- and insulin-induced activation of ribosomal S6 protein kinase 1 and phosphorylation of the translational repressor eukaryotic initiation factor 4E-binding protein 1. Our results demonstrate that glucagon, which is not able to activate but rather inhibits the mTOR pathways, stimulates the phosphorylation of mTOR on Ser2448. This finding suggests that phosphorylation of this site might not be sufficient for mTOR kinase activity but is likely to be involved in other functions. PMID- 15292250 TI - Disruption of hepatic C/EBPalpha results in impaired glucose tolerance and age dependent hepatosteatosis. AB - C/EBPalpha is highly expressed in liver and regulates many genes that are preferentially expressed in liver. Because C/EBPalpha-null mice die soon after birth, it is impossible to analyze the function of C/EBPalpha in the adult with this model. To address the function of C/EBPalpha in adult hepatocytes, liver specific C/EBPalpha-null mice were produced using a floxed C/EBPalpha allele and the albumin-Cre transgene. Unlike whole body C/EBPalpha-null mice, mice lacking hepatic C/EBPalpha expression did not exhibit hypoglycemia, nor did they show reduced hepatic glycogen in adult. Expression of liver glycogen synthase, phosphoenolpyruvate carboxykinase, and glucose-6-phosphatase remained at normal levels. However, these mice exhibited impaired glucose tolerance due in part to reduced expression of hepatic glucokinase, and hyperammonemia from reduced expression of hepatic carbamoyl phosphate synthase-I. These mice also had reduced serum cholesterol and steatotic livers that was exacerbated with aging. This phenotype could be explained by increased expression of hepatic lipoprotein lipase and reduced expression of microsomal triglyceride transfer protein, apolipoproteins B100, and A-IV. These data demonstrate that hepatic C/EBPalpha is critical for ammonia detoxification and glucose and lipid homeostasis in adult mice. PMID- 15292251 TI - The kinase activity of fibroblast growth factor receptor 3 with activation loop mutations affects receptor trafficking and signaling. AB - Amino acid substitutions at the Lys-650 codon within the activation loop kinase domain of fibroblast growth factor receptor 3 (FGFR3) result in graded constitutive phosphorylation of the receptor. Accordingly, the Lys-650 mutants are associated with dwarfisms with graded clinical severity. To assess the importance of the phosphorylation level on FGFR3 maturation along the secretory pathway, hemagglutinin A-tagged derivatives were studied. The highly activated SADDAN (severe achondroplasia with developmental delay and acanthosis nigricans) mutant accumulates in its immature and phosphorylated form in the endoplasmic reticulum (ER), which fails to be degraded. Furthermore, the Janus kinase (Jak)/STAT pathway is activated from the ER by direct recruitment of Jak1. Abolishing the autocatalytic property of the mutated FGFR3 by replacing the critical Tyr-718 reestablishes the receptor full maturation and inhibits signaling. Differently, the low activated hypochondroplasia mutant is present as a mature phosphorylated form on the plasma membrane, although with a delayed transition in the ER, and is completely processed. Signaling does not occur in the presence of brefeldin A; instead, STAT1 is activated when protein secretion is blocked with monensin, suggesting that the hypochondroplasia receptor signals at the exit from the ER. Our results suggest that kinase activity affects FGFR3 trafficking and determines the spatial segregation of signaling pathways. Consequently, the defect in down-regulation of the highly activated receptors results in the increased signaling capacity from the intracellular compartments, and this may determine the severity of the diseases. PMID- 15292252 TI - p38 MAPK mediates gamma-irradiation-induced endothelial cell apoptosis, and vascular endothelial growth factor protects endothelial cells through the phosphoinositide 3-kinase-Akt-Bcl-2 pathway. AB - Therapeutic radiation is widely used in cancer treatments. The success of radiation therapy depends not only on the radiosensitivity of tumor cells but also on the radiosensitivity of endothelial cells lining the tumor vasculature. Vascular endothelial growth factor (VEGF) plays a critical role in protecting endothelial cells against a number of antitumor agents including ionizing radiation. Strategies designed to overcome the survival advantage afforded to endothelial cells by VEGF might aid in enhancing the efficacy of radiation therapy. In this report we examined the signaling cascade(s) involved in VEGF mediated protection of endothelial cells against gamma-irradiation. gamma Irradiation-induced apoptosis of human dermal microvascular endothelial cells (HDMECs) was predominantly mediated through the p38 MAPK pathway as an inhibitor of p38 MAPK (PD169316), and dominant negative mutants of p38 MAPK could significantly enhance HDMEC survival against gamma-irradiation. Inhibition of the PI3K and MAPK pathways markedly up-regulated gamma-irradiation-mediated p38 MAPK activation resulting in enhanced HDMEC apoptosis. In contrast, VEGF-treated HDMECs were protected from gamma-irradiation-induced apoptosis predominantly through the PI3K/Akt pathway. Bcl-2 expression was markedly elevated in VEGF treated HDMECs, and it was significantly inhibited by the PI3K inhibitor LY294002. HDMECs exposed to irradiation showed a significant decrease in Bcl-2 expression. In contrast, VEGF-stimulated HDMECs, when irradiated, maintained higher levels of Bcl-2 expression. Taken together our results suggest that gamma irradiation induces endothelial cell apoptosis predominantly via the activation of p38 MAPK, and VEGF protects endothelial cells against gamma-irradiation predominantly via the PI3K-Akt-Bcl-2 signaling pathway. PMID- 15292253 TI - The interactions of hepatocyte growth factor/scatter factor and its NK1 and NK2 variants with glycosaminoglycans using a modified gel mobility shift assay. Elucidation of the minimal size of binding and activatory oligosaccharides. AB - Full-length hepatocyte growth factor/scatter factor interacts with both heparan and dermatan sulfates and is critically dependent upon them as cofactors for activation of the tyrosine kinase receptor Met. Two C-terminally truncated variants (NK1 and NK2) of this growth factor also occur naturally. Their glycosaminoglycan binding properties are not clear. We have undertaken a comparative study of the heparan/dermatan sulfate binding characteristics of all three proteins. This has entailed the development of a modified gel mobility shift assay, utilizing fluorescence end-tagged oligosaccharides, that is also widely applicable to the analysis of many glycosaminoglycan-protein interactions. Using this we have shown that all three hepatocyte growth factor/scatter factor variants share identical heparan/dermatan sulfate binding properties and that both glycosaminoglycans occupy the same binding site. The minimal size of the oligosaccharide that binds with high affinity in all cases is a tetrasaccharide from heparan sulfate but a hexasaccharide from dermatan sulfate. These findings demonstrate that functional glycosaminoglycan binding is restricted to a binding site situated solely within the small N-terminal domain. The same minimal size fractions are also able to promote hepatocyte growth factor/scatter factor mediated activation of Met and consequent downstream signaling in the glycosaminoglycan-deficient Chinese hamster ovary pgsA-745 cells. A covalent complex of heparan sulfate tetrasaccharide with monovalent growth factor is also active. The binding and activity of tetrasaccharides put constraints upon the possible interactions and molecular geometry within the ternary signaling complex. PMID- 15292254 TI - Staphylococcus aureus 3-hydroxy-3-methylglutaryl-CoA synthase: crystal structure and mechanism. AB - 3-Hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) synthase, a member of the family of acyl-condensing enzymes, catalyzes the first committed step in the mevalonate pathway and is a potential target for novel antibiotics and cholesterol-lowering agents. The Staphylococcus aureus mvaS gene product (43.2 kDa) was overexpressed in Escherichia coli, purified to homogeneity, and shown biochemically to be an HMG-CoA synthase. The crystal structure of the full-length enzyme was determined at 2.0-A resolution, representing the first structure of an HMG-CoA synthase from any organism. HMG-CoA synthase forms a homodimer. The monomer, however, contains an important core structure of two similar alpha/beta motifs, a fold that is completely conserved among acyl-condensing enzymes. This common fold provides a scaffold for a catalytic triad made up of Cys, His, and Asn required by these enzymes. In addition, a crystal structure of HMG-CoA synthase with acetoacetyl CoA was determined at 2.5-A resolution. Together, these structures provide the structural basis for an understanding of the mechanism of HMG-CoA synthase. PMID- 15292255 TI - Perilipin A mediates the reversible binding of CGI-58 to lipid droplets in 3T3-L1 adipocytes. AB - Perilipins, the major structural proteins coating the surfaces of mature lipid droplets of adipocytes, play an important role in the regulation of triacylglycerol storage and hydrolysis. We have used proteomic analysis to identify CGI-58, a member of the alpha/beta-hydrolase fold family of enzymes, as a component of lipid droplets of 3T3-L1 adipocytes. CGI-58 mRNA is highly expressed in adipose tissue and testes, tissues that also express perilipins, and at lower levels in liver, skin, kidney, and heart. Both endogenous CGI-58 and an ectopic CGI-58-GFP chimera show diffuse cytoplasmic localization in 3T3-L1 preadipocytes, but localize almost exclusively to the surfaces of lipid droplets in differentiated 3T3-L1 adipocytes. The localization of endogenous CGI-58 was investigated in 3T3-L1 cells stably expressing mutated forms of perilipin using microscopy. CGI-58 binds to lipid droplets coated with perilipin A or mutated forms of perilipin with an intact C-terminal sequence from amino acid 382 to 429, but not to lipid droplets coated with perilipin B or mutated perilipin A lacking this sequence. Immunoprecipitation studies confirmed these findings, but also showed co-precipitation of perilipin B and CGI-58. Remarkably, activation of cAMP dependent protein kinase by the incubation of 3T3-L1 adipocytes with isoproterenol and isobutylmethylxanthine disperses CGI-58 from the surfaces of lipid droplets to a cytoplasmic distribution. This shift in subcellular localization can be reversed by the addition of propanolol to the culture medium. Thus, CGI-58 binds to perilipin A-coated lipid droplets in a manner that is dependent upon the metabolic status of the adipocyte and the activity of cAMP dependent protein kinase. PMID- 15292256 TI - Proteomic analysis of articular cartilage shows increased type II collagen synthesis in osteoarthritis and expression of inhibin betaA (activin A), a regulatory molecule for chondrocytes. AB - We show that proteomic analysis can be applied to study cartilage pathophysiology. Proteins secreted by articular cartilage were analyzed by two dimensional SDS-PAGE and mass spectrometry. Cartilage explants were cultured in medium containing [35S]methionine/cysteine to radiolabel newly synthesized proteins. To resolve the cartilage proteins by two-dimensional electrophoresis, it was necessary to remove the proteoglycan aggrecan by precipitation with cetylpyridinium chloride. 50-100 radiolabeled protein spots were detected on two dimensional gels of human cartilage cultures. Of 170 silver-stained proteins identified, 19 were radiolabeled, representing newly synthesized gene products. Most of these were known cartilage constituents. Several nonradiolabeled cartilage proteins were also detected. The secreted protein pattern of explants from 12 osteoarthritic joints (knee, hip, and shoulder) and 14 nonosteoarthritic adult joints were compared. The synthesis of type II collagen was strongly up regulated in osteoarthritic cartilage. Normal adult cartilage synthesized little or no type II collagen in contrast to infant and juvenile cartilage. Potential regulatory molecules novel to cartilage were identified; pro-inhibin betaA and processed inhibin betaA (which dimerizes to activin A) were produced by all the osteoarthritic samples and half of the normals. Connective tissue growth factor and cytokine-like protein C17 (previously only identified as an mRNA) were also found. Activin induced the tissue inhibitor for metalloproteinases-1 in human chondrocytes. Its expression was induced in isolated chondrocytes by growth factors or interleukin-1. We conclude that type II collagen synthesis in articular cartilage is down-regulated at skeletal maturity and reactivated in osteoarthritis in attempted repair and that activin A may be an anabolic factor in cartilage. PMID- 15292257 TI - Differential modulation of human melanoma cell metalloproteinase expression by alpha2beta1 integrin and CD44 triple-helical ligands derived from type IV collagen. AB - Tumor cell binding to components of the basement membrane is well known to trigger intracellular signaling pathways. Signaling ultimately results in the modulation of gene expression, facilitating metastasis. Type IV collagen is the major structural component of the basement membrane and is known to be a polyvalent ligand, possessing sequences bound by the alpha1beta1, alpha2beta1, and alpha3beta1 integrins, as well as cell surface proteoglycan receptors, such as CD44/chondroitin sulfate proteoglycan (CSPG). The role of alpha2beta1 integrin and CD44/CSPG receptor binding on human melanoma cell activation has been evaluated herein using triple-helical peptide ligands incorporating the alpha1(IV)382-393 and alpha1(IV)1263-1277 sequences, respectively. Gene expression and protein production of matrix metalloproteinases-1 (MMP-1), -2, -3, -13, and -14 were modulated with the alpha2beta1-specific sequence, whereas the CD44-specific sequence yielded significant stimulation of MMP-8 and lower levels of modulation of MMP-1, -2, -13, and -14. Analysis of enzyme activity confirmed different melanoma cell proteolytic potentials based on engagement of either the alpha2beta1 integrin or CD44/CSPG. These results are indicative of specific activation events that tumor cells undergo upon binding to select regions of basement membrane collagen. Based on the present study, triple-helical peptide ligands provide a general approach for monitoring the regulation of proteolysis in cellular systems. PMID- 15292258 TI - Heparan sulfate/heparin oligosaccharides protect stromal cell-derived factor-1 (SDF-1)/CXCL12 against proteolysis induced by CD26/dipeptidyl peptidase IV. AB - Stromal cell-derived factor-1 (SDF-1) is a CXC chemokine that is constitutively expressed in most tissues and displayed on the cell surface in association with heparan sulfate (HS). Its numerous biological effects are mediated by a specific G protein-coupled receptor, CXCR4. A number of cells inactivate SDF-1 by specific processing of the N-terminal domain of the chemokine. In particular, CD26/dipeptidyl peptidase IV (DPP IV), a serine protease that co-distributes with CXCR4 at the cell surface, mediates the selective removal of the N-terminal dipeptide of SDF-1. We report here that heparin and HS specifically prevent the processing of SDF-1 by DPP IV expressed by Caco-2 cells. The level of processing increases with the level of differentiation of these cells, which correlates with an increase of DPP IV activity. A mutant SDF-1 that does not interact with HS is readily cleaved by DPP IV, a process that is not inhibited by HS, demonstrating that a productive interaction between HS and SDF-1 is required for the protection to take place. Moreover, we found that protection depends on the degree of polymerization of the HS sulfated S-domains. Finally a structural model of SDF-1, in complex with HS oligosaccharides of defined length, rationalizes the experimental data. The mechanisms by which HS regulates SDF-1 may thus include, in addition to its ability to locally concentrate the chemokine at the cell surface, a control of selective protease cleavage events that directly affect the chemokine activity. PMID- 15292259 TI - Ligand-induced conformational shift in the N-terminal domain of GRP94, an Hsp90 chaperone. AB - GRP94 is the endoplasmic reticulum paralog of cytoplasmic Hsp90. Models of Hsp90 action posit an ATP-dependent conformational switch in the N-terminal ligand regulatory domain of the chaperone. However, crystal structures of the isolated N domain of Hsp90 in complex with a variety of ligands have yet to demonstrate such a conformational change. We have determined the structure of the N-domain of GRP94 in complex with ATP, ADP, and AMP. Compared with the N ethylcarboxamidoadenosine and radicicol-bound forms, these structures reveal a large conformational rearrangement in the protein. The nucleotide-bound form exposes new surfaces that interact to form a biochemically plausible dimer that is reminiscent of those seen in structures of MutL and DNA gyrase. Weak ATP binding and a conformational change in response to ligand identity are distinctive mechanistic features of GRP94 and suggest a model for how GRP94 functions in the absence of co-chaperones and ATP hydrolysis. PMID- 15292260 TI - Histone deacetylase 3 interacts with runx2 to repress the osteocalcin promoter and regulate osteoblast differentiation. AB - The Runt domain transcription factor Runx2 (AML-3, and Cbfa1) is essential for osteoblast development, differentiation, and bone formation. Runx2 positively or negatively regulates osteoblast gene expression by interacting with a variety of transcription cofactor complexes. In this study, we identified a trichostatin A sensitive autonomous repression domain in the amino terminus of Runx2. Using a candidate approach, we found that histone deacetylase (HDAC) 3 interacts with the amino terminus of Runx2. In transient transfection assays, HDAC3 repressed Runx2 mediated activation of the osteocalcin promoter. HDAC inhibitors and HDAC3 specific short hairpin RNAs reversed this repression. In vivo, Runx2 and HDAC3 associated with the osteocalcin promoter. These data indicate that HDAC3 regulates Runx2-mediated transcription of osteoblast genes. Suppression of HDAC3 in MC3T3 preosteoblasts by RNA interference accelerated the expression of Runx2 target genes, osteocalcin, osteopontin, and bone sialoprotein but did not significantly alter Runx2 levels. Matrix mineralization also occurred earlier in HDAC3-suppressed cells, but alkaline phosphatase expression was not affected. Thus, HDAC3 regulates osteoblast differentiation and bone formation. Although HDAC3 is likely to affect the activity of multiple proteins in osteoblasts, our data show that it actively regulates the transcriptional activity of the osteoblast master protein, Runx2. PMID- 15292261 TI - The NS3 protein of bluetongue virus exhibits viroporin-like properties. AB - Viroporins compose a group of small hydrophobic transmembrane proteins that can form hydrophilic pores through lipid bilayers. Viroporins have been implicated in promoting virus release from infected cells and in affecting cellular functions including protein trafficking and membrane permeability. Nonstructural protein 3 (NS3) of bluetongue virus has been shown previously to be important for efficient release of newly made virions from infected cells. In this report, we demonstrate that NS3 possesses properties commonly associated with viroporins. Our findings indicate that: (i) NS3 localizes to the Golgi apparatus and plasma membrane in transfected cells, (ii) NS3 can homo-oligomerize in transfected cells, (iii) targeting of NS3 to the Golgi apparatus and plasma membrane correlates with the enhanced permeability of cells to the translation inhibitor hygromycin B (hyg-B), (iv) amino acids 118-148 comprising transmembrane region 1 (TM1) of NS3 are critical for Golgi targeting and hyg-B permeability, and (v) deletion of amino acids 156-181 comprising transmembrane region 2 (TM2) of NS3 has little to no affect on Golgi targeting and hyg-B permeability. These viroporin-like properties may contribute to the role of NS3 in virus release and may have important implications for pathogenicity of bluetongue virus infections. PMID- 15292262 TI - Negative regulation of T cell receptor signaling by Siglec-7 (p70/AIRM) and Siglec-9. AB - Siglec-7 (p70/AIRM) and Siglec-9 are "CD33"-related siglecs expressed on natural killer (NK) cells and subsets of peripheral T cells. Like other inhibitory NK cell receptors, they contain immunoglobulin receptor family tyrosine-based inhibitory motifs in their cytoplasmic domains, and Siglec-7 has been demonstrated to negatively regulate NK cell activation. Based on reports of the presence of these siglecs on T cells, we sought to determine if they are capable of modulating T cell receptor (TCR) signaling using Jurkat T cells stably and transiently transfected with Siglec-7 or Siglec-9. Following either pervanadate stimulation or TCR engagement, both Siglecs exhibited increased tyrosine phosphorylation and recruitment of SHP-1. Effects of Siglec-7 and -9 were also evident in downstream events in the signaling pathway. Both siglecs reduced phosphorylation of Tyr319 on ZAP-70, known to play a pivotal role in up regulation of gene transcription following TCR stimulation. There was also a corresponding decreased transcriptional activity of nuclear factor of activated T cells (NFAT) as determined using a luciferase reporter gene. Like all siglecs, Siglec-7 and -9 recognize sialic acid-containing glycans of glycoproteins and glycolipids as ligands. Mutation of the conserved Arg in the ligand binding site of Siglec-7 (Arg124) or Siglec-9 (Arg120) resulted in reduced inhibitory function in the NFAT/luciferase transcription assay, suggesting that ligand binding is required for optimal inhibition of TCR signaling. The combined results demonstrate that both Siglec-7 and Siglec-9 are capable of negative regulation of TCR signaling and that ligand binding is required for optimal activity. PMID- 15292263 TI - Fourier transform infrared spectroscopy on the Rap.RapGAP reaction, GTPase activation without an arginine finger. AB - GTPase activating proteins (GAPs) down-regulate Ras-like proteins by stimulating their GTP hydrolysis, and a malfunction of this reaction leads to disease formation. In most cases, the molecular mechanism of activation involves stabilization of a catalytic Gln and insertion of a catalytic Arg into the active site by GAP. Rap1 neither possesses a Gln nor does its cognate Rap-GAP employ an Arg. Recently it was proposed that RapGAP provides a catalytic Asn, which substitutes for the Gln found in all other Ras-like proteins (Daumke, O., Weyand, M., Chakrabarti, P. P., Vetter, I. R., and Wittinghofer, A. (2004) Nature 429, 197-201). Here, RapGAP-mediated activation has been investigated by time-resolved Fourier transform infrared spectroscopy. Although the intrinsic hydrolysis reactions of Rap and Ras are very similar, the GAP-catalyzed reaction shows unique features. RapGAP binding induces a GTP(*) conformation in which the three phosphate groups are oriented such that they are vibrationally coupled to each other, in contrast to what was seen in the intrinsic and the Ras.RasGAP reactions. However, the charge shift toward beta-phosphate observed with RasGAP was also observed for RapGAP. A GDP.P(i) intermediate accumulates in the GAP catalyzed reaction, because the release of P(i) is eight times slower than the cleavage reaction, and significant GTP synthesis from GDP.P(i) was observed. Partial steps of the cleavage reaction are correlated with structural changes of protein side groups and backbone. Thus, the Rap.RapGAP catalytic machinery compensates for the absence of a cis-Gln by a trans-Asn and for the catalytic Arg by inducing a different GTP conformation that is more prone to be attacked by a water molecule. PMID- 15292264 TI - TIN2 mediates functions of TRF2 at human telomeres. AB - Telomeres are protective structures at chromosome ends and are crucial for genomic stability. Mammalian TRF1 and TRF2 bind the double-stranded telomeric repeat sequence and in turn are bound by TIN2, TANK1, TANK2, and hRAP1. TRF1 is a negative regulator of telomere length in telomerase-positive cells, whereas TRF2 is important for telomere capping. TIN2 was identified as a TRF1-interacting protein that mediates TRF1 function. We show here that TIN2 also interacts with TRF2 in vitro and in yeast and mammalian cells. TIN2 mutants defective in binding of TRF1 or TRF2 induce a DNA damage response and destabilize TRF1 and TRF2 at telomeres in human cells. Our findings suggest that the functions of TRF1 and TRF2 are linked by TIN2. PMID- 15292265 TI - Molecular dissection of the role of two methyltransferases in the biosynthesis of phenolglycolipids and phthiocerol dimycoserosate in the Mycobacterium tuberculosis complex. AB - A few mycobacterial species, most of which are pathogenic for humans, produce dimycocerosates of phthiocerol (DIM) and of glycosylated phenolphthiocerol, also called phenolglycolipid (PGL), two groups of molecules shown to be important virulence factors. The biosynthesis of these molecules is a very complex pathway that involves more than 15 enzymatic steps and has just begun to be elucidated. Most of the genes known to be involved in these pathways are clustered on the chromosome of M. tuberculosis. Based on their amino acid sequences, we hypothesized that the proteins encoded by Rv2952 and Rv2959c, two open reading frames of this locus, are involved in the transfer of methyl groups onto various hydroxyl functions during the biosynthesis of DIM, PGL, and related p hydroxybenzoic acid derivatives (p-HBAD). Using allelic exchange and site specific recombination, we produced three recombinant strains of Mycobacterium tuberculosis carrying insertions in Rv2952 or Rv2959c. Analysis of these mutants revealed that (i) the protein encoded by Rv2952 is a methyltransferase catalyzing the transfer of a methyl group onto the lipid moiety of phthiotriol and glycosylated phenolphthiotriol dimycocerosates to form DIM and PGL, respectively, (ii) Rv2959c is part of an operon including the newly characterized Rv2958c gene that encodes a glycosyltransferase also involved in PGL and p-HBAD biosynthesis, and (iii) the enzyme encoded by Rv2959c catalyzes the O-methylation of the hydroxyl group located on carbon 2 of the rhamnosyl residue linked to the phenolic group of PGL and p-HBAD produced by M. tuberculosis. These data further extend our understanding of the biosynthesis of important mycobacterial virulence factors and provide additional tools to decipher the molecular mechanisms of action of these molecules during the pathogenesis of tuberculosis. PMID- 15292266 TI - Sphingosine 1-phosphate stimulates smooth muscle cell differentiation and proliferation by activating separate serum response factor co-factors. AB - Sphingosine 1-phosphate (S1P) is a lipid agonist that regulates smooth muscle cell (SMC) and endothelial cell functions by activating several members of the S1P subfamily of G-protein-coupled Edg receptors. We have shown previously that SMC differentiation is regulated by RhoA-dependent activation of serum response factor (SRF). Because S1P is a strong activator of RhoA, we hypothesized that S1P would stimulate SMC differentiation. Treatment of primary rat aortic SMC cells with S1P activated RhoA as measured by precipitation with a glutathione S transferase-rhotekin fusion protein. In SMC and 10T1/2 cells, S1P treatment up regulated the activities of several transiently transfected SMC-specific promoters, and these effects were inhibited by the Rho-kinase inhibitor, Y-27632. S1P also increased smooth muscle alpha-actin protein levels in SMC but had no effect on SRF binding to the smooth muscle alpha-actin CArG B element. Quantitative reverse transcriptase-PCR showed that S1P treatment of SMC or 10T1/2 cells did not increase the mRNA level of either of the recently identified SRF co factors, myocardin or myocardin-related transcription factor-A (MRTF-A). MRTF-A protein was expressed highly in SMC and 10T1/2 cultures, and importantly the effects of S1P were inhibited by a dominant negative form of MRTF-A indicating that S1P may regulate the transcriptional activity of MRTF-A. Indeed, S1P treatment increased the nuclear localization of FLAG-MRTF-A, and the effect of MRTF-A overexpression on smooth muscle alpha-actin promoter activity was inhibited by dominant negative RhoA. S1P also stimulated SMC growth by activating the early growth response gene, c-fos. This effect was not attenuated by Y-27632 but could be inhibited by the MEK inhibitor, UO126. S1P enhanced SMC growth through ERK-mediated phosphorylation of the SRF co-factor, Elk-1, as measured by gel shift and Elk-1 activation assays. Taken together these results demonstrate that S1P activates multiple signaling pathways in SMC and regulates proliferation by ERK-dependent activation of Elk-1 and differentiation by RhoA-dependent activation of MRTF-A. PMID- 15292267 TI - Functional characterization of a P2X receptor from Schistosoma mansoni. AB - The cloning and characterization of a P2X receptor (schP2X) from the parasitic blood fluke Schistosoma mansoni provides the first example of a non-vertebrate ATP-gated ion channel. A number of functionally important amino acid residues conserved throughout vertebrate P2X receptors, including 10 extracellular cysteines, aromatic and positively charged residues involved in ATP recognition, and a consensus protein kinase C site in the amino-terminal tail, are also present in schP2X. Overall, the amino acid sequence identity of schP2X with human P2X(1-7) receptors ranges from 25.8 to 36.6%. ATP evoked concentration-dependent currents at schP2X channels expressed in Xenopus oocytes with an EC(50) of 22.1 microM. 2',3'-O-(4-Benzoylbenzoyl)adenosine 5'-triphosphate (Bz-ATP) was a partial agonist (maximum response 75.4 +/- 4.4% that of ATP) with a higher potency (EC(50) of 3.6 microM) than ATP. Suramin and pyridoxal-phosphate-6 azophenyl-2',4'-disulfonic acid blocked schP2X responses to 100 microm ATP with IC(50) values of 9.6 and 0.5 microM, respectively. Ivermectin (10 microM) potentiated currents to both ATP and Bz-ATP by approximately 60% with a minimal effect on potency (EC(50) of 18.2 and 1.6 microM, respectively). The relative permeability of schP2X expressed in HEK293 cells to various cations was determined under bi-ionic conditions. schP2X has a relatively high calcium permeability (P(Ca)/P(Na) = 3.80 +/- 0.29) and an estimated minimum pore diameter similar to that of vertebrate P2X receptors. SchP2X provides a useful comparative model for the better understanding of human P2X receptor function and may also provide an alternative drug target for treatment of schistosomiasis. PMID- 15292268 TI - Molecular structure of alpha-D-glucose-1-phosphate cytidylyltransferase from Salmonella typhi. AB - Dideoxysugars, which display biological activities ranging from mediating cell cell interactions to serving as components in some antibiotics, are synthesized in various organisms via complex biochemical pathways that begin with the attachment of alpha-D-glucose 1-phosphate to either CTP or dTTP. Here we describe the three-dimensional structure of the alpha-D-glucose-1-phosphate cytidylyltransferase from Salmonella typhi, which catalyzes the first step in the production of CDP-tyvelose. For this investigation, the enzyme was crystallized in the presence of its product, CDP-glucose. In contrast to previous reports, the enzyme exists as a fully integrated hexamer with 32-point group symmetry. Each subunit displays a "bird-like" appearance with the "body" composed predominantly of a seven-stranded mixed beta-sheet and the two "wings" formed by beta-hairpin motifs. These two wings mediate subunit-subunit interactions along the 3-fold and 2-fold rotational axes, respectively. The six active sites of the hexamer are situated between the subunits related by the 2-fold rotational axes. CDP-glucose is anchored to the protein primarily by hydrogen bonds with backbone carbonyl oxygens and peptidic NH groups. The side chains of Arg111 and Asn188 from one subunit and Glu178 and Lys179 from the second subunit are also involved in hydrogen bonding with the ligand. The topology of the main core domain bears striking similarity to that observed for glucose-1-phosphate thymidylyltransferase and 4-diphosphocytidyl-2-C-methylerythritol synthetase. PMID- 15292269 TI - Pinpoint mapping of recognition residues on the cohesin surface by progressive homologue swapping. AB - The high affinity cohesin-dockerin interaction dictates the suprastructural assembly of the multienzyme cellulosome complex. The connection between affinity and species specificity was studied by exploring the recognition properties of two structurally related cohesin species of divergent specificity. The cohesins were examined by progressive rounds of swapping, in which corresponding homologous stretches were interchanged. The specificity of binding of the resultant chimeric cohesins was determined by enzyme-linked affinity assay and complementary protein microarray. In succeeding rounds, swapped segments were systematically contracted, according to the binding behavior of previously generated chimeras. In the fourth and final round we discerned three residues, reputedly involved in interspecies binding specificity. By replacing only these three residues, we were able to convert the specificity of the resultant mutated cohesin, which bound preferentially to the rival dockerin with approximately 20% capacity of the wild-type interaction. These residues represent but 3 of the 16 contact residues that participate in the cohesin-dockerin interaction. This approach allowed us to differentiate, in a structure-independent fashion, between residues critical for interspecies recognition and binding residues per se. PMID- 15292270 TI - A role for caveolae/lipid rafts in the uptake and recycling of the endogenous cannabinoid anandamide. AB - The mechanisms responsible for the uptake and cellular processing of the endogenous cannabinoid anandamide are not well understood. We propose that anandamide uptake may occur via a caveola/lipid raft-related endocytic process in RBL-2H3 cells. Inhibitors of caveola-related (clathrin-independent) endocytosis reduced anandamide transport by approximately 50% compared with the control. Fluorescein derived from fluorescently labeled anandamide colocalized with protein markers of caveolae at early time points following transport. In this study, we have also identified a yet unrecognized process involved in trafficking events affecting anandamide following its uptake. Following uptake of [(3)H]anandamide by RBL-2H3 cells, we found an accumulation of tritium in the caveolin-rich membranes. Inhibitors of both anandamide uptake and metabolism blocked the observed enrichment of tritium in the caveolin-rich membranes. Mass spectrometry of subcellular membrane fractions revealed that the tritium accumulation observed in the caveolin-rich membrane fraction was not representative of intact anandamide, suggesting that following metabolism by the enzyme fatty acid amide hydrolase (FAAH), anandamide metabolites are rapidly enriched in caveolae. Furthermore, HeLa cells, which do not express high levels of FAAH, showed an accumulation of tritium in the caveolin-rich membrane fraction only when transfected with FAAH cDNA. Western blot and immunocytochemistry analyses of RBL-2H3 cells revealed that FAAH was localized in intracellular compartments distinct from caveolin-1 localization. Together, these data suggest that following uptake via caveola/lipid raft-related endocytosis, anandamide is rapidly metabolized by FAAH, with the metabolites efficiently recycled to caveolin-rich membrane domains. PMID- 15292271 TI - Identification of novel beta1 integrin binding sites in the type 1 and type 2 repeats of thrombospondin-1. AB - In addition to the three known beta(1) integrin recognition sites in the N-module of thrombospondin-1 (TSP1), we found that beta(1) integrins mediate cell adhesion to the type 1 and type 2 repeats. The type 1 repeats of TSP1 differ from typical integrin ligands in that recognition is pan-beta(1)-specific. Adhesion of cells that express one dominant beta(1) integrin on immobilized type 1 repeats is specifically inhibited by antagonists of that integrin, whereas adhesion of cells that express several beta(1) integrins is partially inhibited by each alpha subunit-specific antagonist and completely inhibited by combining the antagonists. beta(1) integrins recognize both the second and third type 1 repeats, and each type 1 repeat shows pan-beta(1) specificity and divalent cation dependence for promoting cell adhesion. Adhesion to the type 2 repeats is less sensitive to alpha-subunit antagonists, but a beta(1) blocking antibody and two disintegrins inhibit adhesion to immobilized type 2 repeats. beta(1) integrin expression is necessary for cell adhesion to the type 1 or type 2 repeats, and beta(1) integrins bind in a divalent cation-dependent manner to a type 1 repeat affinity column. The widely used TSP1 function blocking antibody A4.1 binds to a site in the third type 2 repeat. A4.1 proximally inhibits beta(1) integrin dependent adhesion to the type 2 repeats and indirectly inhibits integrin dependent adhesion mediated by the TSP1 type 1 repeats. Although antibody A4.1 is also an antagonist of CD36 binding to TSP1, these data suggest that some biological activities of A4.1 result from antagonism of these novel beta(1) integrin binding sites. PMID- 15292272 TI - Characterization of three glycosyltransferases involved in the biosynthesis of the phenolic glycolipid antigens from the Mycobacterium tuberculosis complex. AB - Mycobacterium tuberculosis and Mycobacterium leprae, the two main mycobacterial pathogens in humans, produce highly specific long chain beta-diols, the dimycocerosates of phthiocerol, and structurally related phenolic glycolipid (PGL) antigens, which are important virulence factors. In addition, M. tuberculosis also secretes glycosylated p-hydroxybenzoic acid methyl esters (p HBAD) that contain the same carbohydrate moiety as the species-specific PGL of M. tuberculosis (PGL-tb). The genes involved in the biosynthesis of these compounds in M. tuberculosis are grouped on a 70-kilobase chromosomal fragment containing three genes encoding putative glycosyltransferases: Rv2957, Rv2958c, and Rv2962c. To determine the functions of these genes, three recombinant M. tuberculosis strains, in which these genes were individually inactivated, were constructed and biochemically characterized. Our results demonstrated that (i) the biosynthesis of PGL-tb and p-HBAD involves common enzymatic steps, (ii) the Rv2957, Rv2958c, and Rv2962c genes are involved in the formation of the glycosyl moiety of the two classes of molecules, and (iii) the product of Rv2962c catalyzes the transfer of a rhamnosyl residue onto p-hydroxybenzoic acid ethyl ester or phenolphthiocerol dimycocerosates, whereas the products of Rv2958c and Rv2957 add a second rhamnosyl unit and a fucosyl residue to form the species-specific triglycosyl appendage of PGL-tb and p-HBAD. The recombinant strains produced provide the tools to study the role of the carbohydrate domain of PGL-tb and p-HBAD in M. tuberculosis pathogenesis. PMID- 15292273 TI - Crystal structure of a family 54 alpha-L-arabinofuranosidase reveals a novel carbohydrate-binding module that can bind arabinose. AB - As the first known structures of a glycoside hydrolase family 54 (GH54) enzyme, we determined the crystal structures of free and arabinose-complex forms of Aspergillus kawachii IFO4308 alpha-l-arabinofuranosidase (AkAbfB). AkAbfB comprises two domains: a catalytic domain and an arabinose-binding domain (ABD). The catalytic domain has a beta-sandwich fold similar to those of clan-B glycoside hydrolases. ABD has a beta-trefoil fold similar to that of carbohydrate binding module (CBM) family 13. However, ABD shows a number of characteristics distinctive from those of CBM family 13, suggesting that it could be classified into a new CBM family. In the arabinose-complex structure, one of three arabinofuranose molecules is bound to the catalytic domain through many interactions. Interestingly, a disulfide bond formed between two adjacent cysteine residues recognized the arabinofuranose molecule in the active site. From the location of this arabinofuranose and the results of a mutational study, the nucleophile and acid/base residues were determined to be Glu(221) and Asp(297), respectively. The other two arabinofuranose molecules are bound to ABD. The O-1 atoms of the two arabinofuranose molecules bound at ABD are both pointed toward the solvent, indicating that these sites can both accommodate an arabinofuranose side-chain moiety linked to decorated arabinoxylans. PMID- 15292274 TI - Molecular cross-talk between MEK1/2 and mTOR signaling during recovery of 293 cells from hypertonic stress. AB - To investigate the role for initiation factor phosphorylation in de novo translation, we have studied the recovery of human kidney cells from hypertonic stress. Previously, we have demonstrated that hypertonic shock causes a rapid inhibition of protein synthesis, the disaggregation of polysomes, the dephosphorylation of eukaryotic translation initiation factor (eIF)4E, 4E-BP1, and ribosomal protein S6, and increased association of 4E-BP1 with eIF4E. The return of cells to isotonic medium promotes a transient activation of Erk1/2 and the phosphorylation of initiation factors, promoting an increase in protein synthesis that is independent of a requirement for eIF4E phosphorylation. As de novo translation is associated with the phosphorylation of 4E-BP1, we have investigated the role of the signaling pathways required for this event by the use of cell-permeable inhibitors. Surprisingly, although rapamycin, RAD001, wortmannin, and LY294002 inhibited the phosphorylation of 4E-BP1 and its release from eIF4E, they did not prevent the recovery of translation rates. These data suggest that only a small proportion of the available eIF4F complex is required for maximal translation rates under these conditions. Similarly, prevention of Erk1/2 activity alone with low concentrations of PD184352 did not impinge upon de novo translation until later times of recovery from salt shock. However, U0126, which prevented the phosphorylation of Erk1/2, ribosomal protein S6, TSC2, and 4E BP1, attenuated de novo protein synthesis in recovering cells. These results indicate that the phosphorylation of 4E-BP1 is mediated by both phosphatidylinositol 3-kinase-dependent rapamycin-sensitive and Erk1/2-dependent signaling pathways and that activation of either pathway in isolation is sufficient to promote de novo translation. PMID- 15292275 TI - Markers of macrophage differentiation in experimental silicosis. AB - Macrophages are characterized by a marked phenotypic heterogeneity depending on their microenvironmental stimulation. Beside classical activation (M1), it has been shown that macrophages could follow a different activation pathway after stimulation with interleukin (IL)-4 or IL-13 (M2). Recently, it has been postulated that those "alternatively activated" macrophages may be critical in the control of fibrogenesis. In an experimental model of silicosis, where pulmonary macrophages play a central role, we addressed the question of whether lung fibrosis development would be associated with alternative macrophage activation. As available markers for alternative macrophage activation, type-1 arginase (Arg-1), Fizz1, Ym1/2, and mannose receptor expression were evaluated at the mRNA and/or protein levels at different stages of the disease. Nitric oxide synthase-2 (NOS-2) expression was also examined to investigate the classical counterpart. We found that the expression of Arg-1, Fizz1, and NOS-2 in adherent bronchoalveolar lavage cells was highly up-regulated 3 days after silica administration but returned to control levels during the fibrotic stage of the disease (60 days). By comparing the early response to silica in C57BL/6 and BALB/c mice, we observed that the amplitude of Arg-1 mRNA up-regulation was not associated with the severity of lung fibrosis. Using a model of manganese dioxide particles (resolutive alveolitis), we showed that this early Arg-1 mRNA was not specific to a fibrogenic lung response. Our data indicate that the modifications of M1/M2 marker expression are limited to the early inflammatory stage of silicosis and that the establishment of a fibrotic process is not necessarily associated with M2 polarization. PMID- 15292276 TI - Antimicrobial proteins and peptides: anti-infective molecules of mammalian leukocytes. AB - Phagocytic leukocytes are a central cellular element of innate-immune defense in mammals. Over the past few decades, substantial progress has been made in defining the means by which phagocytes kill and dispose of microbes. In addition to the generation of toxic oxygen radicals and nitric oxide, leukocytes deploy a broad array of antimicrobial proteins and peptides (APP). The majority of APP includes cationic, granule-associated (poly)peptides with affinity for components of the negatively charged microbial cell wall. Over the past few years, the range of cells expressing APP and the potential roles of these agents have further expanded. Recent advances include the discovery of two novel families of mammalian APP (peptidoglycan recognition proteins and neutrophil gelatinase associated lipocalin), that the oxygen-dependent and oxygen-independent systems are inextricably linked, that APP can be deployed in the context of novel subcellular organelles, and APP and the Toll-like receptor system interact. From a clinical perspective, congeners of several of the APP have been developed as potential therapeutic agents and have entered clinical trials with some evidence of benefit. PMID- 15292277 TI - The orphan nuclear receptor SHP is involved in monocytic differentiation, and its expression is increased by c-Jun. AB - Small heterodimer partner (SHP) is an atypical member of nuclear receptor superfamily that lacks a DNA binding domain. Here, we show that SHP expression increases during monocytic differentiaton with exposure HL-60 leukemia cells to a 12-O-tetradecanoylphorbol-13-acetate (TPA) response element, whose treatment induced the SHP promoter activity dependent on c-Jun expression, which is well known to be involved in the commitment step in the TPA-induced differentiation of HL-60 leukemia cells. We also show that overexpression and activation signaling of c-Jun increase the SHP promoter activity, suggesting that the level of SHP expression is normally limiting for c-Jun-dependent monocytic differentiation. Electrophoretic mobility shift assays using oligonucleotides derived from the SHP promoter reveal that c-Jun exhibit TPA-induced DNA binding, providing a mechanism for the transcriptional activation of SHP gene expression. It was also found that overexpression of SHP and c-Jun greatly facilitated monocytic differentiation by TPA and surprisingly, that expression of SHP or c-Jun alone was sufficient to make cells differentiate into functionally mature monocytes, but silencing of SHP and c-Jun by RNA interference diminished the TPA-induced monocytic differentiation. Taken together, these works suggest that c-Jun works to activate the expression of SHP genes associated with the cascade regulation of monocytic differentiation. PMID- 15292278 TI - Expression and regulation of NFAT (nuclear factors of activated T cells) in human CD34+ cells: down-regulation upon myeloid differentiation. AB - The calcineurin-dependent, cyclosporin A (CsA)-sensitive transcription factor nuclear factor of activated T cells (NFAT) represents a group of proteins, which is well-characterized as a central regulatory element of cytokine expression in activated T cells. In contrast, little is known about the expression or function of NFAT family members in myeloid cells; moreover, it is unclear whether they are expressed by hematopoietic stem/progenitor cells. Here, we show that NFATc2 (NFAT1) is expressed at high levels in CD34+ cells and megakaryocytes but not in cells committed to the neutrophilic, monocytic, or erythroid lineages. Cytokine induced in vitro differentiation of CD34+ cells into neutrophil granulocytes results in the rapid suppression of NFATc2 RNA and protein. NFATc2 dephosphorylation/rephosphorylation as well as nuclear/cytoplasmic translocation in CD34+ cells follow the same calcineurin-dependent pattern as in T lymphocytes, suggesting that NFATc2 activation in these cells is equally sensitive to inhibition with CsA. Finally, in vitro proliferation, but not differentiation, of CD34+ cells cultured in the presence of fms-like tyrosine kinase 3 ligand (FLT3L), stem cell factor, granulocyte macrophage-colony stimulating factor (GM CSF), interleukin-3, and G-CSF is profoundly inhibited by treatment with CsA in a dose-dependent manner. These results suggest a novel and unexpected role for members of the NFAT transcription factor family in the hematopoietic system. PMID- 15292279 TI - Islet amyloid: a critical entity in the pathogenesis of type 2 diabetes. AB - Islet amyloid deposition is a pathogenic feature of type 2 diabetes, and these deposits contain the unique amyloidogenic peptide islet amyloid polypeptide. Autopsy studies in humans have demonstrated that islet amyloid is associated with loss of beta-cell mass, but a direct role for amyloid in the pathogenesis of type 2 diabetes cannot be inferred from such studies. Animal studies in both spontaneous and transgenic models of islet amyloid formation have shown that amyloid forms in islets before fasting hyperglycemia and therefore does not arise merely as a result of the diabetic state. Furthermore, the extent of amyloid deposition is associated with both loss of beta-cell mass and impairment in insulin secretion and glucose metabolism, suggesting a causative role for islet amyloid in the islet lesion of type 2 diabetes. These animal studies have also shown that beta-cell dysfunction seems to be an important prerequisite for islet amyloid formation, with increased secretory demand from obesity and/or insulin resistance acting to further increase islet amyloid deposition. Recent in vitro studies suggest that the cytotoxic species responsible for islet amyloid-induced beta-cell death are formed during the very early stages of islet amyloid formation, when islet amyloid polypeptide aggregation commences. Interventions to prevent islet amyloid formation are emerging, with peptide and small molecule inhibitors being developed. These agents could thus lead to a preservation of beta-cell mass and amelioration of the islet lesion in type 2 diabetes. PMID- 15292280 TI - Clinical characteristics of 104 children referred for evaluation of precocious puberty. AB - There is controversy over the age of onset of puberty in normal children and the risk of missing pathology if the recently proposed revised age guidelines for referring patients are followed. However, there is little recently published information on the frequency of different diagnoses in children referred for signs of early puberty. The purposes of this study were 1) to analyze the spectrum of diagnoses made in a consecutive group of children referred to a single clinician for signs of early puberty; 2) to see whether certain patient groups were more likely to be obese; and 3) to estimate the incidence of endocrine pathology in this sample. The charts of all children referred to the author for evaluation of signs of early puberty between October 1999 and October 2002 were reviewed. Criteria were developed to assign patients to one of seven diagnostic categories based on age, growth, and clinical findings, and differences from the population mean for height and percentage of ideal body weight in the different groups were determined. Most of the patients referred (87%) were female, and the two most common diagnoses made were premature adrenarche (46%) and premature thelarche (18%). Only 9% (all girls) were thought to have true precocious puberty. Two conditions not well described in the literature, pubic hair of infancy and premature menses, were found in 8 and 5%, respectively. Patients with premature adrenarche were significantly taller and more overweight than the general population; a subgroup had evidence of accelerated growth and bone maturation but no worrisome endocrine findings. Acanthosis nigricans was found in 13% of the girls in this study, but the incidence of true endocrine pathology was very low. The majority of children being referred for precocious puberty have benign normal variants, with a very low incidence of endocrine pathology. Most girls presenting with minimal breast or pubic hair development and normal growth velocity may be managed with observation and without a full endocrine evaluation. PMID- 15292281 TI - Official positions of the international society for clinical densitometry. AB - The International Society for Clinical Densitometry (ISCD) periodically holds Position Development Conferences for the purpose of establishing standards and guidelines for indications, acquisition, and interpretation of bone density tests. Topics are selected for consideration by the ISCD Scientific Advisory Committee, reviewed by scientific working groups, and presented to an international panel of experts. Topic categories addressed to date include indications for bone density testing, selection of reference databases for T scores and Z-scores, clinical applications for central and peripheral bone densitometry, serial bone density testing, instrument precision assessment, phantom scanning and calibration testing, requirements for a bone density report, nomenclature, and diagnosis of osteoporosis in postmenopausal women, premenopausal women, men, and children. After an open session for public comment and discussion, the panel convenes for consideration of each topic and makes recommendations for positions to the ISCD Board of Directors. Recommendations that are accepted become the Official Positions of the ISCD. This Special Report summarizes the methodology of the ISCD Position Development Conferences and presents selected Official Positions of general interest. PMID- 15292282 TI - Clinical review 169: Interferon-alpha-related thyroid disease: pathophysiological, epidemiological, and clinical aspects. PMID- 15292283 TI - A randomized trial of remnant ablation--in search of an impossible dream? PMID- 15292284 TI - Patients with differentiated thyroid carcinoma benefit from radioiodine remnant ablation. PMID- 15292285 TI - Clinical review 170: A systematic review and metaanalysis of the effectiveness of radioactive iodine remnant ablation for well-differentiated thyroid cancer. AB - Radioactive iodine remnant ablation destroys residual thyroid tissue after surgical resection of papillary or follicular thyroid cancer. We systematically reviewed 1543 English references to determine whether remnant ablation decreases the risk of thyroid cancer-related death or recurrence after bilateral thyroidectomy for papillary or follicular thyroid cancer. In 13 cohort studies in which the analysis of thyroid cancer-related outcomes was statistically adjusted to a variable degree for prognostic factors or cointerventions, rates of recurrences of thyroid cancer-related outcomes were significantly decreased in the following: one of seven studies examining thyroid cancer-related mortality, three of six studies examining any tumor recurrence, three of three studies examining locoregional recurrence, and two of three studies examining distant metastases. Thyroid hormone suppressive therapy was not adjusted for in the majority of these analyses. In 18 cohort studies not adjusted for prognostic factors or interventions, the benefit of radioactive iodine ablation in decreasing the thyroid cancer-related mortality and any recurrence at 10 yr was inconsistent among centers. However, pooled analyses were suggestive of a statistically significant treatment effect of ablation for the following 10-yr outcomes: locoregional recurrence (relative risk of 0.31, 95% confidence interval, 0.2, 0.49) and distant metastases (absolute decrease in risk 3%, 95% confidence interval, risk decreases 1-4%). In conclusion, radioactive iodine ablation may be beneficial in decreasing recurrence of well-differentiated thyroid cancer; however, results are inconsistent among centers for some outcomes, and the incremental benefit of remnant ablation in low-risk patients treated with bilateral thyroidectomy and thyroid hormone suppressive therapy is unclear. PMID- 15292286 TI - Adrenal replacement therapy: time for an inward look to the medulla? PMID- 15292288 TI - Improving neonatal screening for congenital adrenal hyperplasia. PMID- 15292287 TI - Stress dose of hydrocortisone is not beneficial in patients with classic congenital adrenal hyperplasia undergoing short-term, high-intensity exercise. AB - Classic congenital adrenal hyperplasia (CAH) is associated with impaired function of the adrenal cortex and medulla leading to decreased production of cortisol and epinephrine. As a result, the normal exercise-induced rise in blood glucose is markedly blunted in such individuals. We examined whether an extra dose of hydrocortisone, similar to that given during other forms of physical stress such as intercurrent illness, would normalize blood glucose levels during exercise in patients with CAH. We studied hormonal, metabolic, and cardiorespiratory parameters in response to a standardized high-intensity exercise protocol in nine adolescent patients with classic CAH. Patients were assigned to receive either an additional morning dose of hydrocortisone or placebo, in addition to their usual glucocorticoid and mineralocorticoid replacement in a randomized, double-blind, crossover design 1 h before exercising. Although plasma cortisol levels approximately doubled after administration of the additional hydrocortisone dose compared with the usual single dose, fasting and exercise-induced blood glucose levels did not differ. In addition, no differences were observed in the serum concentrations of the glucose-modulating hormones epinephrine, insulin, glucagon, and GH and of the metabolic parameters lactate and free fatty acids. Although maximal heart rate was slightly higher after stress dosing (193 +/- 3 vs. 191 +/- 3 beats/min, mean +/- sem, P < 0.05), this did not affect exercise performance or perceived exertion. We conclude that patients with classic CAH do not benefit from additional hydrocortisone during short-term, high-intensity exercise. Although this has not been tested with long-term exercise, a high degree of caution should be used when considering the frequent use of additional hydrocortisone administration with exercise, given the adverse side effects of glucocorticoid excess. PMID- 15292289 TI - Steroid profiling by tandem mass spectrometry improves the positive predictive value of newborn screening for congenital adrenal hyperplasia. AB - Congenital adrenal hyperplasia (CAH) is primarily caused by 21-hydroxylase deficiency and leads to an accumulation of 17-hydroxyprogesterone and reduced cortisol levels. Newborn screening for CAH is traditionally based on measuring 17 hydroxyprogesterone by different immunoassays. Despite attempts to adjust cutoff levels for birth weight, gestational age, and stress factors, the positive predictive value for CAH screening remains less than 1%. To improve this situation, we developed a method using liquid chromatography-tandem mass spectrometry to measure 17-hydroxyprogesterone, androstenedione, and cortisol simultaneously in blood spots. A total of 1222 leftover blood spots from six different screening programs using different immunoassays (fluorescent immunoassay and ELISA) were reanalyzed in a blinded fashion by liquid chromatography-tandem mass spectrometry. Thirty-one samples were from babies with CAH, 190 had yielded false-positive results by immunoassay, and the remaining 1001 samples were from babies with normal screening results. Steroid profiling allowed for an elimination of 169 (89%) of the false-positive results and for an improvement of the positive predictive value from the reported 0.5 to 4.7%. Although this method is not suitable for mass screening due to the length of the analysis (12 min), it can be used as a second-tier test of blood spots with positive results for CAH by the conventional methods. This would prevent unnecessary blood draws, medical evaluations, and stress to families. PMID- 15292290 TI - Cardiovascular risk in PCOS. PMID- 15292291 TI - The cardiovascular risk of young women with polycystic ovary syndrome: an observational, analytical, prospective case-control study. AB - To evaluate the cardiovascular risk of polycystic ovary syndrome (PCOS), we investigated lipid profile, metabolic pattern, and echocardiography in 30 young women with PCOS and 30 healthy age- and body mass index (BMI)-matched women. PCOS women had higher fasting glucose and insulin levels, homeostasis model assessment score of insulin sensitivity, total cholesterol (TC) and low density lipoprotein cholesterol (LDL-C) levels, and TC/high density lipoprotein cholesterol (HDL-C) ratio and lower HDL-C levels than controls. Additionally, PCOS women had higher left atrium size (32.0 +/- 4.9 vs. 27.4 +/- 2.1 mm; P < 0.0001) and left ventricular mass index (80.5 +/- 18.1 vs. 56.1 +/- 5.4 g/m(2); P < 0.0001) and lower left ventricular ejection fraction (64.4 +/- 4.1 vs. 67.1 +/- 2.6%; P = 0.003) and early to late mitral flow velocity ratio (1.6 +/- 0.4 vs. 2.1 +/- 0.2; P < 0.0001) than controls. When patients and controls were grouped according to BMI [normal weight (BMI, >18 and <25 kg/m(2)), overweight (BMI, 25.1-30 kg/m(2)), and obese (BMI, >30 kg/m(2))], the differences between PCOS women and controls were maintained in overweight and obese women. In normal weight PCOS women, a significant increase in left ventricular mass index and a decrease in diastolic filling were observed, notwithstanding no change in TC, LDL-C, HDL-C, TC/HDL-C ratio, and TG compared with controls. In conclusion, our data show the detrimental effect of PCOS on the cardiovascular system even in young women asymptomatic for cardiac disease. PMID- 15292292 TI - Challenges of serum thyroglobulin (Tg) measurement in the presence of Tg autoantibodies. PMID- 15292293 TI - Detection of thyrotropin-receptor messenger ribonucleic acid (mRNA) and thyroglobulin mRNA transcripts in peripheral blood of patients with thyroid disease: sensitive and specific markers for thyroid cancer. AB - Because thyroid cancer cells express functional TSH receptors (TSHR), TSHR-mRNA in peripheral blood might serve as a tissue-/cancer-specific marker. We measured circulating TSHR-mRNA by RT-PCR in 51 normal controls, 27 patients with benign thyroid disease, 67 patients with treated differentiated thyroid cancer (DTC), and eight patients with newly diagnosed DTC, preoperatively. Results were compared with thyroglobulin (Tg) mRNA and serum Tg levels. TSHR-mRNA signals were not detected in normal controls and in 24 of 27 (89%) patients with benign thyroid disease. All 19 patients with treated DTC with evidence of distant or local disease tested positive for TSHR-mRNA (sensitivity 100%). Among patients with no evidence of disease, TSHR-mRNA was detected in 1 in 48 (specificity 98%). Six of the eight newly diagnosed DTC patients tested preoperatively were positive for TSHR-mRNA. The concordance between TSHR-mRNA and Tg-mRNA and between TSHR mRNA and serum Tg was 95%. Fourteen patients with DTC (21%) had Tg antibodies, three with local disease (all positive for TSHR-mRNA), and 11 with no evidence of disease (all negative for TSHR-mRNA). Our results indicate that TSHR-mRNA and/or Tg-mRNA in peripheral blood are both equally sensitive and specific markers for monitoring thyroid cancer patients. Their principal value resides in the Tg antibody-positive patients in whom a positive or a negative mRNA value might have indicated or obviated the need for a whole-body scan. Furthermore, the high specificity combined with their ability to predict thyroid cancer preoperatively suggests a potential role in detecting thyroid cancer in patients with thyroid nodules. PMID- 15292294 TI - Papillary thyroid microcarcinoma outcomes and implications for treatment. PMID- 15292295 TI - Clinical behavior and outcome of papillary thyroid cancers smaller than 1.5 cm in diameter: study of 299 cases. AB - To investigate predictors of relapse in small (35%) in peripheral ACTH levels after CRH treatment, even though the IPS:P ratio did not reach the threshold. There were two patients in whom the IPS:P ratio reached threshold criteria, and ectopic tumors were demonstrated as the source of ACTH overproduction (false positives). The sensitivity after CRH stimulation was 90% (95% confidence interval, 80.8 95.5%) with a specificity of 67% (95% confidence interval, 11.4-94.5%). The positive and negative predictive values were 99 and 20%, respectively. Our data show that patients with an IPS:P ratio suggestive of a nonpituitary source of ACTH overproduction may still have Cushing's disease. Analyzing the CRH stimulated peripheral ACTH levels in addition to the standard IPS:P ratios may provide improved diagnostic accuracy. Transsphenoidal exploration should be considered in all cases of unsuccessful sampling and in those cases for which no ectopic source can be identified after further body imaging, even if the IPSS is negative, and especially if peripheral ACTH levels rise significantly with CRH stimulation. PMID- 15292302 TI - Osteoprotegerin gene polymorphisms in men with coronary artery disease. AB - Osteoprotegerin (OPG) antagonizes receptor activator of nuclear factor-kappaB ligand (RANKL), the principal regulator of osteoclasts. Of note, OPG-deficient mice display osteoporosis and arterial calcification. Recently, OPG gene polymorphisms have been associated with osteoporosis and early predictors of cardiovascular disease. In this study, we examined OPG gene polymorphisms in 468 men who had absence of coronary artery disease (CAD) or single-, double-, or triple-vessel disease on coronary angiography. Denaturing gradient gel electrophoresis followed by DNA sequencing revealed nucleotide substitutions 149 T-->C, 163 A-->G, 209 G-->A, 245 T-->G, 950 T-->C (all promoter), 1181 G-->C (exon 1), and 6890 A-->C (intron 4), respectively. Although single polymorphisms were not associated with CAD, linkage of polymorphisms 950 and 1181 revealed that haplotypes were overrepresented in men with CAD (chi(2) = 17.05; P = 0.03) with an increased risk of CAD in carriers of genotypes 950 TC/1181 GC and 950 CC/1181 CC (odds ratio, 1.67; 95% confidence interval, 1.02-2.72; P = 0.04). Furthermore, serum OPG levels were correlated with the presence of a C allele at position 950 (P = 0.02). In summary, linkage of genetic variations of the OPG gene at positions 950 and 1181 may confer an increased risk of CAD in Caucasian men. PMID- 15292303 TI - Ovarian age-related responsiveness to human chorionic gonadotropin in women with polycystic ovary syndrome. AB - Ovarian steroid secretion capacity starts to decline as early as around the age of 30 yr. Whether an age-related decrease in androgen secretion, as in normal women, also occurs in women with polycystic ovary syndrome (PCOS) and whether the enhanced androgen production in PCOS remains throughout the fertile period of life are not known. The aim of this study was to determine the age-related serum basal and gonadotropin-stimulated androgen levels in women with PCOS and to compare the results with those obtained from our previous study in healthy women with normal ovaries. Human chorionic gonadotropin (hCG) stimulation tests were carried out among 42 women with PCOS (age, 16-44 yr; body mass index, 31.02 +/- 1.1 kg/m(2)). An im injection of 5000 IU hCG was given 2-4 d after spontaneous or progestin-induced menstrual bleeding, and blood samples for LH, FSH, inhibin B, 17-hydroxyprogesterone, androstenedione (A), testosterone (T), and estradiol assays were collected at 0, 24, 48, and 96 h. In women with PCOS, basal serum T and A levels were about 50% higher than in healthy women. The responses of A and T to hCG [area under the curve (AUC), 96 h)] were significantly higher in women with PCOS than in normal women [A, 1183.6 +/- 60 (+/-se) vs. 814.4 +/- 39 (P or=0.5 vs. <0.5). Adiponectin was negatively correlated to percentage body fat (r = -0.44; P = 0.002), insulin resistance (r = -0.33; P = 0.016), and age (r = -0.41; P = 0.003). Adiponectin levels were significantly (P = 0.017) higher in pubertal girls compared with boys, but there was no significant difference in prepubertal children in respect to gender (P = 0.833). Adiponectin was significantly (P < 0.001) lower in pubertal compared with prepubertal children. The significant weight loss in 16 children was associated with a significant increase in adiponectin (P = 0.010) and a decrease in insulin resistance (P = 0.013), whereas there were no changes in the 26 children without significant weight loss. Adiponectin levels in obese children were negatively correlated to age, body fat, and insulin resistance and were decreased in puberty. Significant weight loss led to an increase in adiponectin levels and an improvement of insulin resistance. PMID- 15292307 TI - Jugular venous sampling: an alternative to petrosal sinus sampling for the diagnostic evaluation of adrenocorticotropic hormone-dependent Cushing's syndrome. AB - Bilateral sampling of the inferior petrosal sinuses (IPSS) to distinguish Cushing's disease from the ectopic ACTH syndrome is accurate but risky and technically difficult. Bilateral sampling of the internal jugular vein (JVS) is simpler and presumably safer. To compare jugular and petrosal sinus venous sampling for distinguishing Cushing's disease from ectopic ACTH syndrome, we studied 74 patients with surgically proven Cushing's disease, 11 with surgically confirmed, and three with occult ectopic ACTH secretion. Patients underwent JVS and IPSS with administration of CRH on separate days. Ratios of central-to peripheral ACTH in venous samples were calculated. At 100% specificity, IPSS correctly identified 61 of 65 patients with Cushing's disease [sensitivity, 94%; confidence interval (CI), 84-98%]. When patients with abnormal venous drainage were excluded, sensitivity was 98% (CI, 90-100%). JVS had a sensitivity of 83% (CI, 71-91%) at 100% specificity. Receiver operated characteristics plot areas under the curve were similar (0.968 +/- 0.020 and 0.974 +/- 0.016, area under the curve +/- se, JVS vs. IPSS). Although petrosal sampling had better diagnostic accuracy, CIs overlapped (95% CI, 90-100% vs. 86% CI, 78-94%). Centers with limited sampling experience may choose to use the simpler JVS and refer patients for IPSS when the results are negative. PMID- 15292308 TI - Lowering dietary protein to U.S. Recommended dietary allowance levels reduces urinary calcium excretion and bone resorption in young women. AB - High-protein diets increase calciuria. No previous studies have examined the ad libitum U.S. diet's effect on calciuria or bone resorption.Thirty-nine healthy, premenopausal women consuming ad libitum diets [mean, 1.1 g/kg protein, 819 mg (20.5 mmol) Ca, 1152 mg (37 mmol) P, 129 mmol Na] were switched to isocaloric diets containing the U.S. recommended dietary allowance (RDA) of protein (0.8 g/kg) and similar amounts of calcium, phosphorus, and sodium. Bone resorption and related endpoints were assessed before and 1 wk after the switch. As dietary protein changed from ad libitum to RDA levels, mean urine nitrogen decreased 26% (2.4 g/d; P < 0.001) and mean blood urea nitrogen decreased 15% (1.9 mg/dl; P < 0.001). Mean urine pH increased from 6.3 to 6.8 (P < 0.001), and net renal acid excretion (NRAE = urine ammonium plus titratable acids minus bicarbonate) decreased 68% (21.4 mEq/d; P < 0.001). Mean urinary calcium decreased 32% [42 mg (1 mmol)/d; P < 0.001], and bone resorption urine N-telopeptides) decreased 17% (74 micromol bovine collagen equivalents/d; P < 0.001). Mean serum calcium, PTH, and 1,25 dihydroxy vitamin D remained unchanged. In this 2-wk study, decreasing dietary protein from ad libitum to RDA levels decreased NRAE, calciuria and estimates of bone resorption, suggesting that decreased U.S. protein consumption might reduce bone loss. Inasmuch as other dietary modifications, such as increasing vegetable and fruit intake, can result in sustained reductions in NRAE without reducing protein intake, the advisability of reducing protein intake for skeletal protection from acid attack requires further investigation. PMID- 15292309 TI - Impact of ovarian hyperstimulation on thyroid function in women with and without thyroid autoimmunity. AB - Pregnancy is accompanied by changes in thyroid function, but limited data are available on these changes in the very first weeks of pregnancy. Yet, T(4) plays a major role in implantation and early fetal development. We sought to determine thyroid function during this period and during the first trimester, in pregnancies achieved by assisted reproductive technology. Furthermore, the thyroid hormone profile was compared between euthyroid women with (TAI+) and without (TAI-) thyroid autoimmunity. We prospectively analyzed data from 35 women who received ovarian hyperstimulation (OH) and presented clinical pregnancies. The mean age of the women was 32 +/- 5 yr. Thyroid function tests [serum TSH and free T(4) (FT(4))] and thyroid antibody status were determined before OH (baseline values) and every 20 d after ovulation induction during the first trimester of pregnancy. Serum TSH and FT(4) increased significantly at d 20, compared with baseline values (3.3 +/- 2.4 vs. 1.8 +/- 0.9 mU/liter; P < 0.0001 and 13.2 +/- 1.7 vs. 12.4 +/- 1.9 ng/liter; P = 0.005). During the first trimester of pregnancy, there was a significant change over time for TSH and FT(4) (P < 0.001 and P = 0.005, respectively). Nine women (27%) were TAI+. The TSH curve among these TAI+ women was significantly higher compared with TAI- women (P = 0.010). The opposite was observed for the FT(4) curve (P = 0.020). In conclusion, the present study showed a significant increase of serum TSH and FT(4) levels after OH in the very first period of pregnancy compared with pre-OH levels and a significant impact of TAI on the thyroid hormone profile during the first trimester. This provides evidence for an altered thyroid function in euthyroid TAI+ patients. PMID- 15292310 TI - Blood testosterone threshold for androgen deficiency symptoms. AB - There are few systematic studies of the relationship between blood testosterone concentrations and the symptoms of overt androgen deficiency. Because most testosterone preparations are relatively short-term, the rapid changes in blood testosterone concentrations they cause make it difficult to define any testosterone threshold. By contrast, subdermal testosterone implants provide stable blood testosterone concentrations over days to weeks, while gradually declining to baseline over 5-7 months. Hence, this provides an opportunity to define a blood testosterone threshold for androgen deficiency symptoms by observing androgen-deficient men as their familiar androgen deficiency symptoms return as testosterone pellets slowly dissolve. Among 52 androgen-deficient men who underwent 260 implantations over 5 yr, at the time of return of androgen deficiency symptoms the blood total and free testosterone concentrations were highly reproducible within individuals (F = 0.8, P = 0.49 and F = 1.4, 0.24, respectively) but varied markedly between men (F = 167 and F = 138, both P < 0.001), indicating that each person had a consistent testosterone threshold for androgen deficiency symptoms that differed markedly between individuals. The most reported symptoms of androgen deficiency were lack of energy, lack of motivation, and reduced libido. The symptomatic threshold was significantly lower in men with secondary hypogonadism compared with men with primary or mixed hypogonadism (total, 9.7 +/- 0.5 nmol/liter vs. 11.7 +/- 0.4 nmol/liter and 10.2 +/- 0.3 nmol/liter, P = 0.006; free, 146 +/- 10 pmol/liter vs. 165 +/- 6 pmol/liter and 211 +/- 18 pmol/liter, P = 0.002) but was not affected by the underlying cause of hypogonadism or by specific symptoms of any severity. Despite a wide range in individual thresholds for androgen deficiency symptoms, the mean blood testosterone threshold corresponded to the lower end of the eugonadal reference range for young men. The implications of these observations for the development of more specific quality-of-life measures, as well as for other potential androgen deficiency states such as chronic diseases and aging, remain to be determined. PMID- 15292311 TI - Prepubertal Cushing's disease is more common in males, but there is no increase in severity at diagnosis. AB - Sex distribution and severity of biochemical indices at the diagnosis of Cushing's disease (CD) were analyzed in 50 patients (21 males and 29 females; aged 0.1). In conclusion, lower cord ghrelin levels are associated with slower weight gain from birth to 3 months of age. PMID- 15292317 TI - Intelligence quotient and iodine intake: a cross-sectional study in children. AB - The association between iodine deficiency and poor mental and psychomotor development is known. However, most studies were undertaken in areas of very low iodine intake. We investigated whether a similar association is found in schoolchildren from southern Europe with a median urinary iodine output of 90 microg/liter. Urinary iodine levels were measured in 1221 children who also completed a questionnaire about their usual dietary habits. Intelligence quotient (IQ) was measured by Cattell's g factor test. IQ was significantly higher in children with urinary iodine levels above 100 microg/liter. The risk of having an IQ below the 25th percentile was significantly related to the intake of noniodized salt and drinking milk less than once a day. As expected, the risk of having an IQ below 70 was greater in children with urinary iodine levels less than 100 microg/liter. In conclusion, this study demonstrates that the IQ of schoolchildren in a developed country can be influenced by iodine intake. The results support the possibility of improving the IQ of many children from areas with mild iodine deficiency by ensuring an iodine intake sufficient to achieve a urinary iodine concentration greater than 100 microg/liter. PMID- 15292318 TI - The -514 C->T hepatic lipase promoter region polymorphism and plasma lipids: a meta-analysis. AB - Investigations of the -514 C-->T single nucleotide polymorphism (SNP) in the hepatic lipase (HL) gene promoter region (LIPC) have yielded contradictory results regarding its association with changes in plasma lipids. The current study is a meta-analysis of 25 publications on this SNP, comprising over 24,000 individuals, and its relationship with total cholesterol, low-density lipoprotein cholesterol, high-density lipoprotein cholesterol (HDL), triglycerides, and HL activity. Significant decreases were observed in HL activity for both the CT and TT genotypes compared with the CC genotype [weighted mean difference (WMD), -5.83 mmol/liter.h (95% confidence interval, -8.48, -3.17) and -11.05 mmol/liter.h (95% confidence interval, -14.74, -7.36), respectively]. Moreover, significant increases in HDL were found; the CT to CC comparison showed an increase in WMD of 0.04 mmol/liter (95% confidence interval, 0.02, 0.05) mmol/liter, and the increase in the TT vs. CC difference was WMD of 0.09 mmol/liter (95% confidence interval, 0.07, 0.12). These changes appear to be stepwise, implying an allele dosage effect. All P values for these associations were less than 0.001. This meta-analysis demonstrates the importance of the -514C-->T SNP in determining HL activity and plasma HDL concentration and helps quantify the role that hepatic lipase plays in the metabolism of HDL. PMID- 15292319 TI - Intramuscular and liver triglycerides are increased in the elderly. AB - Magnetic resonance spectroscopy studies have shown that intramyocellular lipids (IMCL) and liver fat (LFAT) levels vary with insulin sensitivity and obesity, which are common in the elderly. Thus, magnetic resonance spectroscopy was used to investigate the hypothesis that IMCL and LFAT are increased in the elderly. IMCL and LFAT in young (aged 20-32 yr) and elderly (aged 65-74 yr) were measured fasted, and glucose, insulin, total free fatty acids levels, and free fatty acids profiles were measured during a 2-h oral glucose tolerance test. Body fat percentage was determined with dual x-ray absorptiometry. The elderly had significantly greater IMCL (0.12 +/- 0.01 vs. 0.08 +/- 0.01, mean +/- sem; P = 0.01) and LFAT (0.28 +/- 0.06 vs. 0.08 +/- 0.01; P = 0.004; expressed as ratios to Intralipid standard) than the young. The elderly had increased insulin resistance as calculated by the Matsuda model compared with the young (5.1 +/- 0.9 vs. 9.9 +/- 1.4; P = 0.02). Regression analysis of all subjects indicated that the increases in IMCL and LFAT were correlated with insulin sensitivity, glycosylated hemoglobin, plasma lipids, and body fat. Furthermore, the correlation between insulin sensitivity and IMCL and LFAT remained significant, after accounting for the effect of body fat. Increases of IMCL and LFAT occur in elderly individuals and may be related to insulin resistance. PMID- 15292321 TI - The thyroid epidemiology, audit, and research study: thyroid dysfunction in the general population. AB - The objective of this study was to define the level of treated thyroid dysfunction in a complete and representative population base in an area of sufficient dietary iodine intake. We used record-linkage technology to retrospectively identify subjects treated for hyperthyroidism or hypothyroidism in the general population of Tayside, Scotland from 1 January 1993 to 30 April 1997. Thyroid status was ascertained by record linkage of patient-level datasets containing details of treatments for hyperthyroidism and hypothyroidism. We identified 620 incident cases of hyperthyroidism, an incidence rate of 0.77/1000 x yr [95% confidence interval (CI), 0.70-0.84] in females and 0.14/1000 x yr (95% CI, 0.12-0.18) in males. There were 3,486 incident cases of diagnosed primary hypothyroidism, an incidence rate of 4.98/1000 x yr (95% CI, 4.81-5.17) in females and 0.88/1000 x yr (95% CI, 0.80-0.96) in males. For both hyperthyroidism and hypothyroidism, the incidence increased with age, and females were affected two to eight times more than males across the age range. The midyear point prevalence of all-cause hypothyroidism rose from 2.2% in 1993 to 3.0% in 1996. The level of thyroid dysfunction in Tayside, Scotland is higher than previously reported, and it increased from 1993 to 1996. PMID- 15292320 TI - Plasma leptin levels are associated with coronary atherosclerosis in type 2 diabetes. AB - Leptin signaling may promote atherothrombosis and lead to cardiovascular disease. However, whether leptin is associated with human atherosclerosis, distinct from thrombosis, is unknown. We determined the association of plasma leptin levels with coronary artery calcification (CAC), a measure of coronary atherosclerosis, in a cross-sectional study of type 2 diabetes. Leptin levels were associated with CAC after adjusting for established risk factors [odds ratio (95% confidence interval) for 5 ng/ml leptin increase: 1.31 (1.10-1.55); P = 0.002]. Leptin remained associated with CAC after further controlling for body mass index (BMI) [1.29 (1.07-1.55); P = 0.008], waist circumference [1.30 (1.09-1.57); P = 0.003], C-reactive protein (CRP) levels [1.28 (1.07-1.55); P = 0.008], and subclinical vascular disease [1.30 (1.08-1.57); P = 0.006]. Addition of BMI (P = 0.97), waist (P = 0.55), or CRP (P = 0.39) to a model with leptin failed to improve the model's explanatory power, whereas addition of leptin to a model with BMI (P = 0.029), waist (P = 0.006), or CRP (P = 0.005) improved the model significantly. Plasma leptin levels were associated with CAC in type 2 diabetes after controlling adiposity and CRP. Whether leptin signaling promotes atherosclerosis directly or represents a therapeutic target for the prevention of atherosclerotic cardiovascular disease remains to be explored. PMID- 15292322 TI - Correlation between fasting plasma ghrelin levels and age, body mass index (BMI), BMI percentiles, and 24-hour plasma ghrelin profiles in Prader-Willi syndrome. AB - Ghrelin is a GH-releasing acylated peptide found in the stomach and a centrally acting food intake stimulator. Prader-Willi syndrome (PWS) is a genetic disorder characterized by a voracious appetite and increased fasting ghrelin levels. In this report we describe 24-h ghrelin profiles in PWS children (n = 5) and compare these with age, sex, and body mass index (BMI)-matched controls (n = 5). A 3- to 4-fold increase in ghrelin levels was found in PWS over a 24-h period, compared with controls (P < 0.001). Interestingly, there was a greater tendency for the up regulation of ghrelin level in lean PWS than in obese PWS. To confirm this finding, we measured fasting ghrelin levels in 39 patients with PWS. Inverse correlations were found between plasma ghrelin levels and the following: age (r = -0.408, P = 0.005), BMI (r = -0.341, P = 0.017), percentage of the ideal weight for age (r = -0.382, P = 0.008), and BMI percentile (r = -0.311, P = 0.027). Our data show that there may be a suppressive (or up-regulating) controlling mechanism of ghrelin secretion in obese (or lean) PWS children. We hope that our data may further explain the mechanisms underlying the insatiable appetite and obesity characteristic of PWS. PMID- 15292323 TI - Comparison of continuation or cessation of growth hormone (GH) therapy on body composition and metabolic status in adolescents with severe GH deficiency at completion of linear growth. AB - Although GH replacement improves the features of GH deficiency (GHD) in adults, it has yet to be established whether cessation of GH at completion of childhood growth results in adverse consequences for the adolescent with GHD. Effects of continuation or cessation of GH on body composition, insulin sensitivity, and lipid levels were studied in 24 adolescents (13 males, 11 females, aged 17.0 +/- 0.3, yr, mean +/- se, puberty stage 4 or 5) in whom height velocity was less than 2 cm/yr. Provocative testing confirmed severe GHD [peak GH < 9 mU/liter (3 microg/liter)] in all cases and was followed by a lead-in period of 3 months during which the pediatric dose of GH continued unchanged. Baseline investigations were then performed using dual-energy x-ray absorptiometry (body composition), lipid measurements, and assessment of insulin sensitivity by both homeostasis model assessment and a short insulin tolerance test. Twelve patients remained on GH (0.35 U/kg.wk), and 12 patients ceased GH treatment. The groups were followed up in parallel with repeat observations made after 6 and 12 months. No endocrine differences were evident between the groups at baseline. GH cessation resulted in a reduction of serum IGF-I Z score [-1.62 +/- 0.29, baseline vs. -2.52 +/- 0.12, 6 months (P < 0.05) vs. -2.52 +/- 0.10, 12 months (P < 0.01)] but values remained unchanged in those continuing GH replacement. Lean body mass increased by 2.5 +/- 0.5 kg ( approximately 6%) over 12 months in those receiving GH but was unchanged after GH discontinuation. Cessation of GH resulted in increased insulin sensitivity [short insulin tolerance test, 153 +/- 22 micromol/liter.min, baseline vs. 187 +/- 20, 6 months (P < 0.05) vs. 204 +/- 14, 12 months (P = 0.05)], but no significant change was seen during 12 months of GH continuation. Lipid levels remained unaltered in both groups. Continuation of GH at completion of linear growth resulted in ongoing accrual of lean body mass (LBM), whereas skeletal muscle mass remained static after GH cessation in these adolescents with GHD. This divergence of gain in LBM is of potential importance because increases in LBM occur as a feature of healthy late adolescent development. GH is a major mediator of insulin sensitivity, independent of body composition in adolescents. Further studies are required to determine whether discontinuation of GH in the adolescent with severe GHD once linear growth is complete results in long-term irreversible adverse physical and metabolic consequences and to determine conclusively the benefits of continuing GH therapy. PMID- 15292324 TI - Prediction of autoantibody positivity and progression to type 1 diabetes: Diabetes Autoimmunity Study in the Young (DAISY). AB - Diabetes Autoimmunity Study in the Young (DAISY) has followed 1972 children for islet autoimmunity and diabetes: 837 first-degree relatives of persons with type 1 diabetes and 1135 general population newborns identified through human leukocyte antigen (HLA) screening. During follow-up of 4.06 yr (range, 0.17-9 yr), serial determination of autoantibodies to glutamic acid decarboxylase, protein tyrosine phosphatase IA2, and insulin has generated approximately 20,000 results. Among 162 children with at least one positive autoantibody, in 31% the test was false positive (autoantibodies were negative twice on blinded duplicate aliquots), in 31% it was transiently positive (confirmed on blinded duplicate aliquots but negative on follow-up), and in 36% it was persistently positive. Using proportional hazards modeling, the HLA-DR3/4 DQ8 genotype, another positive autoantibody at the first positive visit, and level of autoantibody were predictive of persistent positivity. Only HLA-DR3/4 DQ8 genotype was predictive of progression to diabetes in proportional hazards modeling. This prospective study reveals that cross-sectional determination of islet autoantibodies in a population with relatively low previous probability of autoimmunity identifies as "positive" a large number of individuals who are either false or transiently positive. Predictive value of autoantibodies increases with blinded duplicate and independent sample retesting and incorporation of the level of autoantibody in the predictive algorithm. PMID- 15292325 TI - Effects of risedronate on bone density in anorexia nervosa. AB - Anorexia nervosa (AN), a psychiatric disease characterized by chronic starvation, is complicated by severe bone loss (1), for which there is no effective, available therapy. Although bone resorption is markedly increased in these patients, estrogen is an ineffective anti-resorptive therapy in the setting of undernutrition. We hypothesized bisphosphonate administration would result in a decrease in bone resorption and an increase in bone density in women with AN and bone loss, despite undernutrition. We therefore administered risedronate 5 mg daily for nine months to 10 women with AN, all of whom had osteopenia (mean AP spine T score: -2.7 +/- 2) and compared NTX and bone density with baseline values and with those from available control data prospectively followed for the same time period. Bone density increased significantly in patients who received risedronate compared with controls and compared with baseline, despite lack of significant weight gain, for an increase of AP spine bone density of 4.1 +/- 1.6% at six months and 4.9 +/- 1.0% at nine months. Bone resorption, as measured by NTX, decreased 23.8% at one month and 29.6% at three months, from the high-normal to mid-normal range of young women. Our data suggest that risedronate 5 mg daily administered to women with AN and osteopenia may increase in bone density at the AP spine despite low weight. This is the first study to demonstrate marked increases in bone density in women with AN. Because of the lack of data regarding the safety of such medications in women of reproductive age, bisphosphonates are not approved in the U.S. for premenopausal women other than those receiving glucocorticoids. Further studies are needed to establish the efficacy and safety of bisphosphonate therapy in this population. PMID- 15292326 TI - Impaired endothelial function in young women with premature ovarian failure: normalization with hormone therapy. AB - Normal menopause is associated with vascular endothelial dysfunction, an early stage of atherosclerosis. The effect of premature ovarian failure (or premature menopause) on endothelial function in young women is unknown. Endothelial function was assessed in 18 women with premature ovarian failure before and after 6 months of hormone therapy and was compared with the endothelial function of 20 age- and body mass index-matched premenopausal women. Brachial artery diameter was measured both during hyperemia (an index of endothelium-dependent vasodilation) and in response to glyceryl trinitrate (an index of endothelium independent vaso-dilation). Flow-mediated dilation was significantly lower in women with premature ovarian failure at baseline (increase in brachial artery diameter during hyperemia by 3.06 +/- 4.33%) than in control women (increase by 8.84 +/- 2.15%; P < 0.0005). Glyceryl trinitrate-induced vasodilation did not differ between the groups. After hormone therapy for 6 months, flow-mediated dilation was improved in women with premature ovarian failure, increasing by more than 2-fold (7.41 +/- 3.86%; P < 0.005 compared with pretreatment) and reaching normal values (P not significant compared with control women). Glyceryl trinitrate-induced vasodilation did not change after treatment in women with premature ovarian failure. Young women with premature ovarian failure have significant vascular endothelial dysfunction. Early onset of endothelial dysfunction associated with sex steroid deficiency may contribute to the increased risk of cardiovascular disease and mortality in young women with premature ovarian failure. Hormone therapy restores endothelial function within 6 months of treatment. PMID- 15292327 TI - Separate contribution of diabetes, total fat mass, and fat topography to glucose production, gluconeogenesis, and glycogenolysis. AB - The contribution of increased gluconeogenesis (GNG) to the excessive rate of endogenous glucose production (EGP) in type 2 diabetes (T2DM) is well established. However, the separate effects of obesity (total body fat), visceral adiposity, and T2DM have not been investigated. We measured GNG (by the (2)H(2)O technique) and EGP (with 3-(3)H-glucose) after an overnight fast in 44 type 2 diabetic and 29 gender/ethnic-matched controls. Subjects were classified as obese (body mass index 30 kg/m(2) or greater) or nonobese (body mass index < 30 kg/m(2)); diabetic subjects were further subdivided according to the severity of fasting hyperglycemia [fasting plasma glucose (FPG) < 9 mm or >or= 9 mm]. EGP was similar in nondiabetic controls and T2DM with FPG less than 9 mm but was increased in T2DM with FPG >or= 9 mm (P < 0.001). Within the diabetic groups, obesity had an independent effect to further increase basal EGP (P < 0.01). In both nonobese diabetic groups, both the percent GNG and gluconeogenic flux were increased, compared with nonobese nondiabetic controls. In both diabetic groups, obesity further increased both percent GNG and gluconeogenic flux. In obese and nonobese T2DM, the increase in gluconeogenic flux was not accompanied by a reciprocal decrease in glycogenolysis, indicating a loss of hepatic autoregulation. By multivariate analysis, gluconeogenic flux was positively correlated with percent body fat, visceral fat, and the fasting plasma free fatty acid and glucose concentrations (all P 0.05). On the contrary, the ghrelin concentration was significantly reduced in poorly controlled patients (353 +/- 23 ng/liter; P < 0.0001). Ghrelin correlated negatively with Phe in all three groups, whereas it correlated positively with catecholamine levels and energy intake and negatively with BMI only in diet-controlled patients and controls. We conclude that ghrelin secretion may receive positive direct or indirect input from catecholamines. The absence of a correlation between ghrelin and catecholamines, energy intake, or BMI in PKU patients on an inadequate diet may be due to dysregulation of their neuroendocrine system and might be affected by high Phe levels in the stomach and/or central nervous system. PMID- 15292338 TI - Effects of estrogen and recombinant human insulin-like growth factor-I on ghrelin secretion in severe undernutrition. AB - Ghrelin is a nutritionally regulated gut peptide that increases with fasting and chronic undernutrition and decreases with food intake. Sex steroid levels change in chronic undernutrition and might signal changes in ghrelin. At the same time, chronic undernutrition is characterized by low IGF-I that might also influence ghrelin, either directly or through changes in the GH axis. Little is known regarding sex steroid regulation of ghrelin and the effects of IGF-I on ghrelin in severe undernutrition. We investigated the effects of sex steroids and IGF-I on ghrelin in 78 female subjects with anorexia nervosa simultaneously randomized to receive estrogen (Ovcon 35, 35 microg ethinyl estradiol, and 0.4 mg norethindrone) as well as recombinant human (rh)IGF-I (30 microg/kg sc twice a day) in a two-by-two factorial model, in which the individual effects of estrogen (E) and rhIGF-I on ghrelin could be determined. Subjects were 24.9 +/- 0.7 (mean +/- sem) yr of age and had low weight (body mass index, 16.7 +/- 0.2 kg/m(2)). At baseline, ghrelin was inversely correlated with body mass index (r = -0.39, P = 0.0005) and IGF-I (r = -0.30, P = 0.01). IGF-I increased significantly more in subjects receiving rhIGF-I alone (Delta 23.0 +/- 5.8 nmol/liter) and rhIGF-I and E (Delta 34.9 +/- 6.3 nmol/liter) compared with subjects receiving E alone (Delta -3.2 +/- 1.9 nmol/liter) or control (C; rhIGF-I placebo and no E) (Delta 0.4 +/- 2.0 nmol/liter) (overall P < 0.0001 by multivariate analysis of variance, P < 0.0001 for rhIGF-I vs. C, P < 0.0001 for rhIGF-I and E vs. C). Ghrelin increased significantly more over 6 months in response to E alone (Delta 150 +/- 86 pg/ml), rhIGF-I alone (Delta 198 +/- 116 pg/ml), and the combination (E and rhIGF-I) (Delta 441 +/- 214 pg/ml) compared with C (Delta -39 +/- 48 pg/ml) (overall P = 0.02 by multivariate analysis of variance, P = 0.01 for E vs. C, P = 0.04 for rhIGF-I vs. C, and P = 0.001 for rhIGF-I and E vs. C). Weight, caloric intake, and morning GH levels did not change significantly between the groups, but the change in ghrelin was inversely related to the change in GH among all subjects (r = -0.27, P = 0.03).Our data demonstrate that, in a model of severe undernutrition, rhIGF-I and E individually increase ghrelin levels. The mechanisms of these effects are unknown and may relate to direct effects on ghrelin or changes in GH. Further studies are needed to determine the mechanisms by which rhIGF-I and E increase ghrelin in human physiology. PMID- 15292339 TI - Impaired activation of protein kinase C-zeta by insulin and phosphatidylinositol 3,4,5-(PO4)3 in cultured preadipocyte-derived adipocytes and myotubes of obese subjects. AB - Insulin resistance in obesity is partly due to diminished glucose transport in myocytes and adipocytes, but underlying mechanisms are uncertain. Insulin stimulated glucose transport requires activation of phosphatidylinositol (PI) 3 kinase (3K), operating downstream of insulin receptor substrate-1. PI3K stimulates glucose transport through increases in PI-3,4,5-(PO(4))(3) (PIP(3)), which activates atypical protein kinase C (aPKC) and protein kinase B (PKB/Akt). However, previous studies suggest that activation of aPKC, but not PKB, is impaired in intact muscles and cultured myocytes of obese subjects. Presently, we examined insulin activation of glucose transport and signaling factors in cultured adipocytes derived from preadipocytes harvested during elective liposuction in lean and obese women. Relative to adipocytes of lean women, insulin-stimulated [(3)H]2-deoxyglucose uptake and activation of insulin receptor substrate-1/PI3K and aPKCs, but not PKB, were diminished in adipocytes of obese women. Additionally, the direct activation of aPKCs by PIP(3) in vitro was diminished in aPKCs isolated from adipocytes of obese women. Similar impairment in aPKC activation by PIP(3) was observed in cultured myocytes of obese glucose intolerant subjects. These findings suggest the presence of defects in PI3K and aPKC activation that persist in cultured cells and limit insulin-stimulated glucose transport in adipocytes and myocytes of obese subjects. PMID- 15292340 TI - Circulating glucocorticoid bioactivity in the preterm newborn after antenatal betamethasone treatment. AB - Antenatal glucocorticoid treatment of mothers at risk of premature delivery is highly cost-effective in reducing neonatal mortality and morbidity. However, there is only limited information on the actual glucocorticoid bioactivity (GBA) reaching the fetus. By employing a recently developed recombinant cell bioassay, we studied circulating GBA in preterm newborns exposed to the standard antenatal betamethasone regimen (12 mg betamethasone twice, 24-h interval, for the mother; repeated in 7-10 d if required). Plasma GBA and cortisol concentrations were measured in cord blood of 71 infants (mean gestational age, 28.9 wk; range, 24.6 32 wk; mean birth weight, 1208 g; range, 480-2010 g). The median time between the last administered betamethasone dose and birth was 2.0 d. Cord GBA ranged from less than 15.6 to 170 nmol/liter cortisol equivalents. The level was highly dependent on the time between the last betamethasone dose and birth, i.e. infants born shortly (<12 h) after the last steroid dose displayed on average 4-fold higher GBA than that in the reference group (infants with >7 d since the last betamethasone dose before birth or without treatment; 74 vs. 21 nmol/liter cortisol equivalents; P < 0.0001). By contrast, if more than 72 h had elapsed between the last steroid dose and birth, circulating GBA was strongly dependent on cord cortisol (r = 0.85; P < 0.0001; n = 30). In multiple regression analysis adjusted for cord cortisol concentration and the time since the last steroid dose, increased umbilical artery resistance, a sign of severe fetal distress, was associated with lower cord GBA (P = 0.01). In conclusion, antenatal exposure of preterm fetuses to betamethasone causes a sizeable, but brief, peak of supraphysiological GBA, and approximately 3 d after the last betamethasone dose, circulating GBA derives from cord cortisol concentration. PMID- 15292341 TI - The ER22/23EK polymorphism in the glucocorticoid receptor gene is associated with a beneficial body composition and muscle strength in young adults. AB - Glucocorticoids play an important role in determining body composition. A polymorphism of the glucocorticoid receptor gene (in codons 22 and 23) has previously been found to be associated with relative glucocorticoid resistance, low cholesterol levels, and increased insulin sensitivity. In this study, we investigated whether this ER22/23EK polymorphism is associated with differences in body composition and muscle strength. We studied a cohort of 350 subjects who were followed from age 13 until 36 yr. We compared noncarriers and carriers of the ER22/23EK variant in anthropometric parameters, body composition, and muscle strength, as measured by arm pull tests and high jump from standing. We identified 27 (8.0%) heterozygous ER22/23EK carriers. In males at 36 yr of age, we found that ER22/23EK carriers were taller, had more lean body mass, greater thigh circumference, and more muscle strength in arms and legs. We observed no differences in body mass index or fat mass. In females, waist and hip circumferences tended to be smaller in ER22/23EK carriers at the age of 36 yr, but no differences in body mass index were found. Thus, the ER22/23EK polymorphism is associated with a sex-specific, beneficial body composition at young-adult age, as well as greater muscle strength in males. PMID- 15292342 TI - Adiponectin concentrations are influenced by renal function and diabetes duration in Pima Indians with type 2 diabetes. AB - Adiponectin is produced exclusively by adipocytes, and its serum concentration is inversely associated with adiposity. This study examines the relationship among diabetes, renal function, and serum adiponectin in Pima Indians. Serum adiponectin was measured in 1069 people in whom glycemia and renal function had been measured. Serum adiponectin, adjusted for age, sex, and body mass index, was lowest in those with impaired glucose regulation or diabetes of less than 10 yr duration and highest in those with normal glucose tolerance or diabetes of duration of at least 10 yr. Both urinary albumin to creatinine ratio (ACR) and serum creatinine were positively correlated with adiponectin (Spearman's r = 0.43; P < 0.0001, and r = 0.37; P < 0.0001, respectively) in diabetic subjects. After stratification by albuminuria (normoalbuminuria ACR < 30 mg/g, microalbuminuria ACR = 30-299 mg/g, and macroalbuminuria ACR >or= 300 mg/g), the highest adiponectin concentration was in the macroalbuminuria group (geometric mean = 9.6 microg/ml) and the lowest was in the normoalbuminuric group (geometric mean = 5.6 microg/ml). After adjustment for age, sex, body mass index, and diabetes duration, the serum adiponectin concentration in the macroalbuminuria group was significantly higher than in both other groups (P < 0.0001). Serum adiponectin is lowest in the presence of impaired glucose regulation and early diabetes. In the presence of diabetes, serum adiponectin is positively associated with abnormal renal function and diabetes duration. PMID- 15292343 TI - Estrogen regulates expression of tumor necrosis factor receptors in breast adipose fibroblasts. AB - In breast cancer, a dense layer of undifferentiated fibroblasts is formed around malignant breast epithelial cells and referred to as desmoplastic reaction. These cells provide structural and functional support for tumor growth. Aromatase, the key enzyme in the biosynthesis of estrogen, is overexpressed in these undifferentiated fibroblasts, producing large quantities of estrogen, which in turn influences the growth and progression of malignant epithelial cells. We previously demonstrated that malignant epithelial cells produce large amounts of TNFalpha, which inhibit the differentiation of breast fibroblasts. TNF action is mediated by its two receptors (TNFRs), TNFR1, which mediates inhibition of adipocyte differentiation, and TNFR2, which was linked to the proliferation of thymocytes. We present evidence here that estrogen modulates the synthesis of receptors for TNF in human adipose fibroblasts (HAFs) from breast tissue in a paracrine fashion, which may serve as a mechanism for the inhibition of adipocyte differentiation in breast cancer. Estradiol (E(2)) treatment increased TNFR1 mRNA and protein levels in primary HAFs in a dose- and time-dependent manner, which could be reversed by the estrogen antagonist ICI182,780. Interestingly, higher concentration of E(2) inhibited whereas lower concentrations stimulated TNFR2 mRNA levels in HAFs. To investigate the specific roles of TNFRs in adipocyte differentiation, we incubated breast HAFs with receptor selective muteins of TNF. TNFR1-selective mutein decreased mRNA levels of aP2, a marker for adipogenic differentiation. This antiadipogenic effect was enhanced by cotreatment with E(2). We conclude that high levels of estrogen found in breast tumors promote the antiadipogenic action of TNF on breast adipose fibroblasts by selectively up regulating TNFR1, which may be a critical mechanism for desmoplastic reaction. PMID- 15292344 TI - Hypercalciuria in familial hyperkalemia and hypertension accompanies hyperkalemia and precedes hypertension: description of a large family with the Q565E WNK4 mutation. AB - Familial hyperkalemia and hypertension (FHH; pseudohypoaldosteronism type II) is an autosomal dominant disorder characterized by hyperkalemia, hypertension, and low renin. WNK1 kinase overexpression and WNK4 kinase inactivating missense mutations cause FHH. When expressed in frog oocyte, WNK4 inhibits Na-Cl cotransporter surface expression, and WNK1 relieves this inhibition. We have reported hypercalciuria in subjects with the WNK4 Q565E mutation. In contrast, in subjects with WNK1 overexpression, normocalciuria was found. Here we report a major extension of our previously described kindred that contains 34 subjects, 18 of them affected by the mutation. Hypertension was diagnosed in 13 affected subjects at the age of 31 +/- 12 yr. Five of the affected or obligatory affected subjects had stroke, in four at the age of 50-62 yr. Seven subjects with FHH were diagnosed 27 yr previously. All four subjects who were normotensive at diagnosis became hypertensive during follow-up. The mean time between detection of hyperkalemia and appearance of hypertension was 13 yr. In the extended kindred, compared with the unaffected subjects, affected subjects had hyperkalemia, low transtubular potassium gradient, hyperchloremia, low bicarbonate, higher aldosterone, and marked suppression of renin. Urinary calcium levels in affected and unaffected subjects were 0.85 +/- 0.27 and 0.28 +/- 0.12 mmol/mmol creatinine, respectively (P < 0.0001). Hypercalciuria was accompanied by lower serum calcium levels [9.44 +/- 0.15 vs. 9.81 +/- 0.31 mg/dl (2.36 +/- 0.04 vs. 2.45 +/- 0.08 mmol/liter); P = 0.01], supporting a mechanism of renal calcium leak. The six affected, currently normotensive subjects had the same degree of hyperkalemia, hypercalciuria, and low renin as the affected hypertensive subjects. We conclude that in FHH with WNK4 mutations, with time all affected subjects will apparently develop hypertension. Hypercalciuria accompanies hyperkalemia, and both precede hypertension. Based on the recent findings that WNK4 regulates the renal outer medullary potassium channel as well as epithelial Cl(-)/base exchanger and the Na(+)-K(+)-2Cl(-) cotransporter, we suggest that WNK4 interacts with a calcium channel or transporter. PMID- 15292345 TI - Cord plasma adiponectin: a 20-fold rise between 24 weeks gestation and term. AB - Adiponectin is an adipocyte-derived hormone with profound insulin-sensitizing, antiinflammatory, and antiatherogenic effects. Apart from its obvious potential as a mediator of adult metabolic syndrome, adiponectin could have a significant role in regulating fetal growth.We measured plasma adiponectin concentrations by ELISA in cord vein of 197 infants. Of them, 122 were born preterm (gestational age, 22-32 wk), and 75 at term (49 from a healthy and 26 from a diabetic pregnancy, with similar findings, and thus all data from term infants pooled).Mean adiponectin concentrations increased from less than 1 microg/ml at 24 wk gestation to approximately 20 microg/ml at term. One week increase in gestational age corresponded in preterm infants to 43% increase (95% confidence interval 34-53%; P < 0.0001) in adiponectin and term infants to 21% increase (12 31%; P < 0.0001). In preterm infants, one unit increase in birth weight sd score corresponded to 42% increase (22-66%; P = 0.0001) in adiponectin, and females had 57% higher adiponectin concentrations (0-146%; P = 0.05) than males. These differences were not seen in term infants. Adiponectin levels were lower in preterm infants with recent (<12 h) exposure to maternal betamethasone but were unrelated to mode of delivery, preeclampsia, or impaired umbilical artery flow. In conclusion, adiponectin concentrations in fetal circulation show a 20-fold rise between 24 wk gestation and term and, in preterm infants are associated with birth weight sd score, sex, and glucocorticoid exposure. Adiponectin may play an important role in regulating fetal growth and explaining its links to the metabolic syndrome and its consequences during adult life. PMID- 15292346 TI - Absolute risk of childhood-onset type 1 diabetes defined by human leukocyte antigen class II genotype: a population-based study in the United Kingdom. AB - The autoimmune disease process leading to childhood-onset type 1 diabetes appears to start in infancy, and decisions on treatment to prevent initiation of autoimmunity will need to be based on genetic susceptibility alone. We set out to quantify the absolute risk associated with human leukocyte antigen (HLA) DRB1 DQA1-DQB1 class II genotypes and to develop strategies for recruitment into primary prevention trials. HLA class II haplotype- and genotype-specific risks were derived from 753 United Kingdom families from the Bart's-Oxford population based study of type 1 diabetes and combined with incidence data from the region to calculate the absolute risk of development of diabetes. A hierarchy of susceptibility was established for both HLA class II haplotypes and genotypes, and the sensitivity and specificity of each genotype was established relative to age at disease onset. Highest risk was conferred by the genotype DRB1*03 DQA1*0501-DQB1*0201/DRB1*0401-DQA1*0301-DQB1*0302 (5% absolute risk of diabetes by age 15 yr), although sensitivity was only 22.6%. Combining the six highest risk genotypes conferred similar risk but increased sensitivity to 36.6% and was most sensitive for diagnosis of diabetes before age 5 yr (48.4%), whereas inclusion of 11 genotypes achieved the same sensitivity for diagnosis for ages 10 14 yr. Analysis of genotype-specific risk should form the basis for design of future primary prevention trials in the general population. PMID- 15292347 TI - Genome-wide linkage analysis reveals evidence for four new susceptibility loci for familial euthyroid goiter. AB - Euthyroid goiter is characterized by diffuse or nodular enlargement of the thyroid gland. Iodine deficiency and cigarette smoking have been identified as important environmental factors. However, family and twin pair studies suggest a strong genetic predisposition. Therefore, we performed the first extended genome wide scan to identify susceptibility loci that predispose for euthyroid goiter using 450 microsatellite markers in 18 extended Danish, German, and Slovakian families. Parametric and nonparametric multipoint linkage analyses were performed. The highest nonparametric LOD scores were obtained for chromosomes 2q and 3p with values of 2.54 at D2S1363 and 2.25 at D3S3038, respectively. Assuming heterogeneity and dominant inheritance, heterogeneity LOD scores (HLOD) of 2.71 and 1.94 were calculated for 2q and 3p, respectively. Furthermore, nonparametric LOD scores of 1.87 (HLOD 1.39) at D7S1808 on 7q and 1.79 (HLOD 1.80) at D8S264 on 8p were obtained. Haplotyping of families contributing to the linkage signals revealed four families compatible with a putative locus on 3p and one family each showing strict cosegregation with the loci on 2q, 7q, and 8p. The four novel candidate loci corroborate the assumed heterogeneity in the etiology of euthyroid familial goiter. For the first time, a more prevalent putative locus, present in 20% of the families investigated, was identified. PMID- 15292348 TI - Gender differences of adiponectin levels develop during the progression of puberty and are related to serum androgen levels. AB - Adiponectin is an adipocytokine with profound antidiabetic and antiatherogenic effects that is decreased in obesity. With the increasing prevalence of obesity and the emergence of related disorders, including type 2 diabetes in children, the regulation of adiponectin and its relationship to childhood obesity is of great interest. In this study we aimed to elucidate the impact of gender, pubertal development, and obesity on adiponectin levels in children. We investigated two phenotypically characterized cohorts of 200 normal weight and 135 obese children and adolescents covering a wide range of age (3.4-17.9 yr) and body mass index (-2.1 to +4.8 sd score). In healthy lean boys, adiponectin levels significantly declined in parallel with physical and pubertal development, subsequently leading to significantly reduced adiponectin levels in adolescent boys compared with girls (5.6 +/- 0.5 vs. 7.1 +/- 0.5 mg/liter; P = 0.03). This decline was inversely related to testosterone (r = -0.42; P < 0.0001) and dehydroepiandrosterone sulfate (r = -0.20; P = 0.0068) serum concentrations and may account for the gender differences seen in adults. Using a stepwise forward multiple regression model, pubertal stage was the strongest independent predictor of adiponectin (r(2) = 0.206; P < 0.0001), with additional influences of body mass index sd score and testosterone. Adiponectin levels were decreased in obese children and adolescents compared with lean peers of corresponding age and pubertal stage (5.18 vs. 7.13 mg/liter; P = 0.015). In obese children, adiponectin levels were closely associated with parameters related to the metabolic syndrome, such as insulin resistance, hyperinsulinemia, blood pressure, and uric acid, in univariate and multivariate analyses, with the insulin sensitivity index being the strongest independent parameter identified by stepwise forward multiple regression (r(2) = 0.226; P < 0.0001). Hence, there is a strong association of adiponectin serum concentrations with obesity, pubertal development, and metabolic parameters in children indicating epidemiological and pathophysiological relevance already in childhood. PMID- 15292349 TI - Macrophage inhibitory factor, plasminogen activator inhibitor-1, other acute phase proteins, and inflammatory mediators normalize as a result of weight loss in morbidly obese subjects treated with gastric restrictive surgery. AB - Obesity is demonstrated to be associated with an enhanced inflammatory state, which is suggested to be a cause for the development of obesity-related morbidity. It was hypothesized that a decrease in body weight in morbid obese subjects would lead to a reduction of the inflammatory state in these subjects. Weight loss was achieved by gastric restrictive surgery in 27 morbidly obese patients. Preoperative as well as 3-, 6-, 12-, and 24-month postoperative plasma concentrations of inflammatory mediators macrophage inhibitory factor, plasminogen activator inhibitor-1, lipopolysaccharide binding protein, alpha-1 acid glycoprotein, C-reactive protein, soluble TNFalpha receptors 55 and 75, and leptin were measured. Macrophage inhibitory factor levels remained low normal for 6 months, during weight loss, after which they significantly increased to normal levels at 24 months postoperatively. The other inflammatory mediators remained elevated up to minimally 3 months postoperatively; thereafter they decreased significantly. Both TNFalpha receptors remained elevated up to at least 12 months postoperatively to decrease significantly at 2 yr postoperatively. This study demonstrates that during weight loss, after gastric restrictive surgery, inflammatory mediators remain elevated for at least 3 months postoperatively, suggesting initially an ongoing inflammatory state. However, 2 yr after surgery, the inflammatory mediators reach near normal values.These findings may be an explanation for the reduced comorbidity seen in morbidly obese patients after gastric restrictive surgery. PMID- 15292350 TI - 3,5,3'-Triiodothyronine down-regulates Fas and Fas ligand expression and suppresses caspase-3 and poly (adenosine 5'-diphosphate-ribose) polymerase cleavage and apoptosis in early placental extravillous trophoblasts in vitro. AB - The present study was conducted to determine whether T(3) receptor exists in early placental extravillous trophoblasts (EVTs) and evaluate the influence of T(3) on Fas/Fas ligand expression, caspase-3, and poly (ADP-ribose) polymerase (PARP) cleavage and apoptosis in cultured early placental EVTs. EVTs with invasive phenotype, isolated from normal placental explants from early pregnancy through preincubation on human fibronectin-coated dishes and exhibited cytokeratin 7 and human placental lactogen immunopositive staining, were cultured in the absence or presence of T(3) (10(-7) to 10(-9) m). The presence of T(3) receptor in cultured EVTs was examined by immunocytochemistry, RT-PCR, and Southern blot analysis. Fas sensitivity was determined by treating the cells with an agonistic Fas antibody. Apoptosis was assessed by terminal deoxynucleotidyl transferase deoxyuridine triphosphate nick end labeling, flow cytometry, and Hoechst nuclear staining. Fas and Fas ligand expression and caspase-3 and PARP cleavage were evaluated by immunocytochemistry. Early placental EVTs expressed a 212-bp c-erb Abeta1 transcript and the T(3) receptor protein and exhibited significant levels of apoptosis in culture. Treatment with T(3) reduced the expression of Fas and Fas ligand as well as cleavage of caspase-3 and PARP and suppressed apoptosis in cultured EVTs. Although addition of agonistic Fas antibody increased apoptosis in these cells, this response was markedly attenuated by the presence of T(3). These results demonstrate that T(3) receptor is present in early placental EVTs and that T(3) suppresses apoptosis by down regulating the expression of Fas and Fas ligand. These findings are consistent with the hypothesis that T(3) promotes EVT invasion to the decidua by suppressing apoptosis in early pregnancy. PMID- 15292351 TI - Human endocrine gland-derived vascular endothelial growth factor: expression early in development and in Leydig cell tumors suggests roles in normal and pathological testis angiogenesis. AB - Angiogenesis is essential for tumor growth and metastasis. A new human angiogenic mitogen, endocrine gland-derived vascular endothelial growth factor (EG-VEGF), has been recently identified; its expression pattern is restricted to endocrine glands, with the highest expression in testis. We used in situ hybridization and newly generated monoclonal antibodies to investigate the expression of EG-VEGF in normal human prenatal and adult testis and in 48 human testicular tumors of different subtypes. We found that EG-VEGF was expressed from 14 wk until birth in human fetal testis. In the adult testis, EG-VEGF was strongly expressed only in Leydig cells. In testicular tumors, EG-VEGF was expressed specifically in Leydig cell tumors, whereas germ cell-derived neoplasms, including carcinoma in situ, seminoma, and nonseminomatous germ cell tumors, were negative for this antigen. In contrast, VEGF, another powerful angiogenic factor, was expressed in seminoma, but very weakly in Leydig cell tumors. Interestingly, we found that Leydig cell tumors presented vessel surface density 3.2-fold higher than seminoma. These findings argue that human EG-VEGF may play a role in angiogenesis both during the early endocrine development of testis and in the adult testis as well as in Leydig cell tumor growth. PMID- 15292352 TI - Evidence for the presence of glucose transporter 4 in the endometrium and its regulation in polycystic ovary syndrome patients. AB - Glucose transporter 4 (GLUT4) seems to be involved in the mechanism of insulin resistance in polycystic ovary syndrome (PCOS) patients (PCOSs) in both muscular and adipose tissue. The observation that insulin stimulates glucose oxidation in endometrial cells led us to investigate the presence of GLUT4 in this tissue and whether a defect of GLUT4 is present at the endometrial level in PCOSs. We also investigated whether body weight influences GLUT4 expression in this syndrome. GLUT4 mRNA content was examined by real-time quantitative RT-PCR and immunostaining reaction in the endometrial tissue of nine normal subjects, nine lean and eight obese hyperinsulinemic (h-INS), and eight lean and 10 obese normoinsulinemic (n-INS) PCOSs. GLUT4 mRNA and its positive immunostaining reaction were present in epithelial cell level in the endometrium of both normal and PCOS subjects. Significantly higher levels of GLUT4 were observed in normal and lean n-INS PCOSs in comparison with other groups. In both n-INS and h-INS obese PCOSs, GLUT4 was significantly lower than in lean subjects. However, obese n-INS and lean h-INS PCOSs showed a similar low GLUT4 expression, whereas obese h INS PCOSs showed the lowest expression when compared with other groups. In conclusion, our data demonstrate that GLUT4 is present in the endometrium of normal and PCOS subjects and that hyperinsulinism and obesity seem to have a negative effect on endometrial GLUT4 expression in PCOS. PMID- 15292353 TI - Human fetal and cord serum thyroid hormones: developmental trends and interrelationships. AB - Thyroid hormone is essential for fetal and neonatal development in particular of the brain, but little is known about regulation of fetal thyroid hormone levels throughout human gestation. The purpose of this study was to clarify developmental trends and interrelationships among T(4), free T(4) (FT4), thyroxine-binding globulin (TBG), TSH, T(3), rT(3), and T(4) sulfate (T4S) levels in cord and fetal blood sera (n = 639, 15-42 wk gestation) and correlate infant levels (23-42 wk gestation) to maternal values (n = 428, 16-45 yr) and those of nonpregnant women (n = 233, 16-46 yr). In cord and fetal serum, T(4), T(3), and TBG levels increase with gestation until term; TSH, FT4, T4S, and rT(3) levels increase and peak in the late second/early third trimester and then decline to term; T(4)/TBG ratios increase until late second trimester and plateau to term. Term cord sera TSH, TBG, and all iodothyronine levels, except T(3), are higher than nonpregnant women. In the third trimester, cord serum FT4, TSH, rT(3), and T4S levels are also higher than corresponding maternal levels, but T(4), T(3), and TBG levels are lower than maternal values. The late second/early third trimester is a critical transition period in fetal thyroid hormone metabolism, which may be interrupted by preterm birth and contribute to postnatal thyroid dysfunction. PMID- 15292354 TI - Localization of osteoprotegerin, tumor necrosis factor-related apoptosis-inducing ligand, and receptor activator of nuclear factor-kappaB ligand in Monckeberg's sclerosis and atherosclerosis. AB - Vascular calcification may occur at different areas of the vessel wall, including the intima in atherosclerosis and the media in Monckeberg's sclerosis. Medial calcification of arteries is common in patients with diabetes mellitus or chronic renal failure. Osteoprotegerin (OPG) and receptor activator of nuclear factor kappaB ligand are essential modulators of bone homeostasis and may be involved in the process of vascular calcification. In this study we investigated arteries from patients with Monckeberg's sclerosis and atherosclerosis. Apoptosis, which precedes vascular calcification in vitro, was assessed by an in situ ligation assay and was localized to the medial layer of arteries (Monckeberg's sclerosis) and the neointima (atherosclerosis). Immunohistochemistry and in situ hybridization revealed OPG immunoreactivity and mRNA expression surrounding calcified areas in the medial layer (Monckeberg's sclerosis), whereas OPG was mainly expressed adjacent to calcified neointimal lesions (atherosclerosis). Receptor activator of nuclear factor-kappaB ligand protein and mRNA were barely or not detectable. Of note, TNF-related apoptosis-inducing ligand, an inducer of apoptosis that is also blocked by OPG, displayed a similar spatial distribution as OPG. In summary, we demonstrate enhanced apoptosis adjacent to vascular calcification, and the concurrent expression of regulators of apoptosis and osteoclastic differentiation, TNF-related apoptosis-inducing ligand and OPG, suggesting their involvement in the pathogenesis of vascular calcification. PMID- 15292355 TI - Nur-related factor 1 and nerve growth factor-induced clone B in human adrenal cortex and its disorders. AB - Nerve growth factor-induced clone B (NGFI-B; NR4A1) and Nur-related factor 1 (Nurr1; NR4A2) are members of NGFI-B family of orphan receptors. We recently demonstrated induction of CYP11B2 (aldosterone synthase) by Nurr1 and NGFI-B, suggesting possible important roles of these transcriptional factors in the regulation of adrenocortical steroidogenesis. Therefore, we immunolocalized Nurr1 and NGFI-B in various human adrenal specimens to study their biological significance. In nonpathological adrenal glands (n = 25), Nurr1 and NGFI-B immunoreactivities were detected at high levels in the fetal definitive zone or postnatal zona glomerulosa. NGFI-B immunoreactivity was increased according to development in the zona fasciculata, reaching a level similar to that in the zona glomerulosa in adult adrenal cortex. In adrenocortical neoplasms (n = 44), Nurr1 immunoreactivity was higher in aldosteronoma than in Cushing's adenoma or adrenocortical carcinoma. NGFI-B immunoreactivity was also higher in aldosteronoma than in adrenocortical carcinoma, but was not significantly different among the types of adenoma. Both Nurr1 and NGFI-B mRNA expressions were correlated with their immunoreactivities in adrenocortical neoplasms (n = 23), and mRNA expression of Nurr1 was significantly (P < 0.0001) associated with that of CYP11B2. These results suggest that the expression of Nurr1 and NGFI-B plays an important role in human adrenal cortex and its neoplasms, including possible regulation of steroidogenesis. PMID- 15292356 TI - Presence of luteinizing hormone/human chorionic gonadotropin receptors in male breast tissues. AB - Receptors for LH/human chorionic gonadotropin (hCG) have been found in a variety of nongonadal tissues including the female breast. Using in situ hybridization and immunohistochemistry, we demonstrated the presence of LH/hCG receptor mRNA and protein in normal male breast tissue obtained at autopsy (n = 4) and archival samples of benign gynecomastia (n = 14) and male breast carcinoma (n = 5). Although the function of these receptors remains to be determined, the findings suggest the possibility that LH and hCG may play a role in the pathogenesis of male breast disorders. PMID- 15292357 TI - Novel intronic mutation of MEN1 gene causing familial isolated primary hyperparathyroidism. AB - Primary hyperparathyroidism may occur as part of hereditary syndromes, including multiple endocrine neoplasia types 1 and 2A (MEN1 and MEN2A), hyperparathyroidism jaw tumor syndrome, and the familial isolated hyperparathyroidism (FIHP). It is unclear whether FIHP corresponds to a different genetic entity or a variant of MEN1 (or hyperparathyroidism-jaw tumor syndrome). We report a patient and 11 family members with FIHP in whom we identified a heterozygous G-to-A mutation at nucleotide 7361 of tumor suppressor MEN1 gene. This mutation is located in the first base of intron 9 (IVS9 + 1 G>A). All the family members with hyperparathyroidism were heterozygous for the intronic mutation. In vitro studies were performed in COS cells transfected with minigenes carrying the coding regions spanning exon-intron 9 and 10 with the mutant and the wild-type sequences. RT-PCR analyses showed an abnormal mRNA of greater size (829 bp) in the mutated MEN1 gene than the normal transcript (629 bp). The longer PCR product includes the exon 9, the unspliced intron 9, and part of exon 10. RT-PCR of MEN1 mRNA from patient's blood confirmed the existence of unspliced intron 9 in mature mRNA. In summary, we report a case of FIHP associated with a new intronic heterozygous germline mutation (IVS9 + 1 G>A) of MEN1 gene. This mutation produces an aberrant splicing of mRNA that could lead to a truncated protein, without activity, explaining the clinical picture of this patient and his family. PMID- 15292358 TI - Expression of SHOX in human fetal and childhood growth plate. AB - Abnormalities in the growth plate may lead to short stature and skeletal deformity including Leri Weil syndrome, which has been shown to result from deletions or mutations in the SHOX gene, a homeobox gene located at the pseudoautosomal region of the X and Y chromosome. We studied the expression of SHOX protein, by immunohistochemistry, in human fetal and childhood growth plates and mRNA by in situ hybridization in childhood normal and Leri Weil growth plate. SHOX protein was found in reserve, proliferative, and hypertrophic zones of fetal growth plate from 12 wk to term and childhood control and Leri Weil growth plates. The pattern of immunostaining in the proliferative zone of childhood growth plate was patchy, with more intense uniform immunostaining in the hypertrophic zone. In situ hybridization studies of childhood growth plate demonstrated SHOX mRNA expression throughout the growth plate. No difference in the pattern of SHOX protein or mRNA expression was seen between the control and Leri Weil growth plate. These findings suggest that SHOX plays a role in chondrocyte function in the growth plate. PMID- 15292359 TI - Four new cases of congenital secondary hypothyroidism due to a splice site mutation in the thyrotropin-beta gene: phenotypic variability and founder effect. AB - Isolated TSH deficiency is a rare cause of congenital hypothyroidism. We here report four children from two consanguineous Turkish families with isolated TSH deficiency. Affected children who were screened at newborn age had an unremarkable TSH result and a low serum TSH level at diagnosis. Age at diagnosis and clinical phenotype were variable. All affected children carried an identical homozygous splice site mutation (IVS2 + 5 G--> A) in the TSHbeta gene. This mutation leads to skipping of exon 2 and a loss of the translational start codon without ability to produce a TSH-like protein. However, using specific monoclonal antibodies, we detected a very low concentration of authentic, heterodimeric TSH in serum, indicating the production of a small amount of correctly spliced TSH mRNA. By genotyping all family members with polymorphic markers at the TSHbeta locus, we show that the mutation arose on a common ancestral haplotype in three unrelated Turkish families indicating a founder mutation in the Turkish population. These results suggest that this TSHbeta mutation is among the more common TSHbeta gene mutations and stress the need for a biochemical and molecular genetic workup in children with symptoms suggestive of congenital hypothyroidism, even when the neonatal TSH screening is normal. PMID- 15292360 TI - Pheochromocytoma and medullary thyroid carcinoma: a new genotype-phenotype correlation of the RET protooncogene 891 germline mutation. AB - Prior experience in kindreds with a codon 891 RET protooncogene mutation indicates that carriers of this mutation develop only hereditary medullary thyroid carcinoma without evidence of other manifestations of multiple endocrine neoplasia type 2. In this paper, we report the first documented case in which medullary thyroid carcinoma and pheochromocytoma were clinically expressed in members of a family affected by the codon 891 RET mutation. Genetic analysis of the RET protooncogene in this family revealed an exon 15 missense mutation at codon 891 that resulted in a serine to alanine amino acid substitution. These findings indicate that patients with this mutation should be screened for pheochromocytoma. PMID- 15292361 TI - X-linked sex-determining region Y box 3 (SOX3) gene mutations are uncommon in men with idiopathic oligoazoospermic infertility. AB - The X-linked sex-determining region Y box 3 (SOX3) gene is expressed in the developing gonads and brain. Sox3-null mice developed according to genetic sex, but the hemizygous null males were hypogonadal, with extensive Sertoli cell vacuo lization, loss of germ cells, and reduced sperm count. We hypothesized that SOX3 mutations might occur in a subset of infertility patients. Genomic DNA samples from 56 infertile men with idiopathic oligo-azoospermia were screened for SOX3 mutations. Three nucleotide substitutions (609 T-->C, 732 A-->C, and 978 G-->A) were identified, none of which altered the amino acid sequence, suggesting that they are polymorphic variants. The 609 T-->C substitution was in the HMG box, and the two other substitutions were identified within the polyalanine repeat regions. Three patients had 609 T-->C, 2 patients had 609 T-->C and 732 A-->C, and one had 978 G-->A. These data indicate that mutations in the SOX3 gene are not a common cause of male infertility. PMID- 15292362 TI - Paradoxical proinflammatory actions of interleukin-10 in human amnion: potential roles in term and preterm labour. AB - IL-10 is regarded predominantly as an inhibitor of cell-mediated inflammatory reactions. As such, it has been suggested that IL-10 could have therapeutic potential, including the treatment of preterm labor. Using explant cultures of gestational membranes we have found that IL-10 does indeed exert anti inflammatory properties in choriodecidua, but that in the adjacent amnion it has remarkable pro-inflammatory actions. Amnion prostaglandin (PG) E(2) production was significantly increased following 24-h treatment with IL-10 (5-100 ng/mL). Production of IL-8 also showed a significant stimulation at 100 ng/mL IL-10. In contrast, choriodecidual production of IL-8 and tumor necrosis factor (TNF)-alpha was dramatically inhibited by IL-10, confirming the ability of this tissue to exhibit a classical IL-10 response. IL-10 retained its stimulatory actions on amnion in the presence of IL-1beta and TNF-alpha stimulation. These findings suggest that the fetal membranes can exhibit opposing responses to IL-10, depending on whether the inflammatory insult occurs at the maternal or fetal face. While inflammatory reactions are negatively regulated by IL-10 in choriodecidua, if the pathogen reaches the amnion and threatens the fetus, pro inflammatory reactions may predominate to ensure successful labor to spare and protect the fetus. PMID- 15292363 TI - Increased expression of regulator of G protein signaling-2 (RGS-2) in Bartter's/Gitelman's syndrome. A role in the control of vascular tone and implication for hypertension. AB - Regulator of G protein signaling-2 (RGS-2) plays a key role in the G protein coupled receptor (GPCR) angiotensin II (Ang II) signaling. NO and cGMP exert a vasodilating action also through activation and binding to RGS-2 of cGMP dependent protein kinase 1-alpha, which phosphorylates RGS-2 and dephosphorylates myosin light chain. In Bartter's/Gitelman's patients (BS/GS) Ang II related signaling and vasomotor tone are blunted. Experiments were planned to explore whether RGS-2 may play a role in BS/GS vascular hyporeactivity. NO metabolites and cGMP urinary excretion were also measured. Mononuclear cells (PBM) from six BS/GS patients and six healthy controls were used. PBM RGS-2 mRNA and RGS-2 protein were increased in BS/GS: 0.47 +/- 0.06 d.u. vs 0.32 +/- 0.04, (p < 0.006) (RGS-2 mRNA), and 0.692 +/- 0.02 vs 0.363 +/- 0.06 (p < 0.0001) (RGS2 protein). Incubation of PBM with Ang II increased RGS-2 protein in controls (from 0.363 +/- 0.06 d.u. to 0.602 +/- 0.05; p < 0.0001) but not in BS/GS (from 0.692 +/- 0.02 to 0.711 +/- 0.02). NO(2)(-)/NO(3)(-) and cGMP urinary excretion were increased in BS/GS (0.46 +/- 0.13 vs 0.26 +/- 0.05 micromol/micromol of urinary creatinine, p < 0.005, and 0.060 +/- 0.030 vs 0.020 +/- 0.01 p < 0.009, respectively). These results demonstrate that RGS-2 is increased and maximally stimulated in BS/GS and human RGS-2 system reacts as predicted by knockout mice experiments. This is the first report of RGS-2 level in a human clinical condition characterized by altered vascular tone, underlines the importance of RGS-2 as a key regulator element for Ang II signaling and provides insight into the links between BS/GS genetic abnormalities and abnormal vascular tone regulation. PMID- 15292364 TI - Serum levels of matrix extracellular phosphoglycoprotein (MEPE) in normal humans correlate with serum phosphorus, parathyroid hormone and bone mineral density. AB - Matrix extracellular phosphoglycoprotein (MEPE), a member of the Small Integrin Binding Ligand N-linked Glycoprotein (SIBLING) family, is primarily expressed in normal bone and has been proposed as a phosphaturic factor because of high expression and secretion in oncogenic hypophosphatemic osteomalacia tumors. In order to begin to address the role of MEPE in normal human physiology, we developed a competitive ELISA to measure serum levels of MEPE. The ELISA was used to characterize the distribution pattern in a population consisting of 114 normal adult subjects. The mean value of MEPE was 476 +/- 247 ng/ml and levels decreased significantly with increasing age. MEPE levels were also significantly correlated with serum phosphorus and parathyroid hormone (PTH). In addition, MEPE levels correlated significantly with measures of bone mineral density in the femoral neck and total hip in a subset of 50 elderly subjects. The results are consistent with MEPE being involved in phosphate and bone metabolism in a normal population. PMID- 15292366 TI - Timing of pituitary stalk assessment in Langerhans cell histiocytosis: "when" is sometimes more important than "what". PMID- 15292365 TI - Somatostatin infusion lowers plasma ghrelin without reducing appetite in adults with Prader-Willi syndrome. AB - Prader-Willi syndrome (PWS) is characterized by life-threatening childhood-onset hyperphagia, obesity and, uniquely, high plasma levels of ghrelin, the orexigenic gastric hormone. Somatostatin suppresses ghrelin secretion in normal subjects. We therefore examined the effect of somatostatin on plasma ghrelin and appetite in four male PWS adults fasted overnight in a double-blind, placebo-controlled, randomized cross-over study. Subjects received an intravenous infusion of somatostatin (250 microg/hr) or saline for 300 min, and had blood samples taken every 30 min for measurement of plasma ghrelin and PYY3-36 (anorexigenic intestinal hormone) by radio-immunoassay, and glucose. Appetite was measured by counting sandwiches eaten over a 60 min free food access period from +120 min. Despite somatostatin lowering fasting plasma ghrelin by 60 +/- 2% (P = 0.04) to levels seen in non-PWS men, there was no associated reduction in food intake (105 +/- 9% of food intake during saline infusion, P = 0.6). Somatostatin also lowered plasma PYY levels by 45 +/- 16% (P = 0.04), and produced post-prandial hyperglycemia (P = 0.04). We conclude that either hyperghrelinemia may not contribute to hyperphagia in PWS adults, or perhaps concomitant reductions in anorexigenic gastrointestinal hormones by somatostatin counteracted any anorexigenic effect of lowering orexigenic ghrelin. Somatostatin analogues may therefore not be an effective therapy for obesity in PWS. Larger chronic studies with long-acting somatostatin analogues will be needed to determine their benefits and risks in treating PWS obesity. PMID- 15292367 TI - Revised nomenclature for the mammalian long-chain acyl-CoA synthetase gene family. AB - By consensus, the acyl-CoA synthetase (ACS) community, with the advice of the human and mouse genome nomenclature committees, has revised the nomenclature for the mammalian long-chain acyl-CoA synthetases. ACS is the family root name, and the human and mouse genes for the long-chain ACSs are termed ACSL1,3-6 and Acsl1,3-6, respectively. Splice variants of ACSL3, -4, -5, and -6 are cataloged. Suggestions for naming other family members and for the nonmammalian acyl-CoA synthetases are made. PMID- 15292368 TI - Tissue-specific autoregulation of the LXRalpha gene facilitates induction of apoE in mouse adipose tissue. AB - The functions of the liver X receptors (LXRs) are not well documented in adipose tissue. We demonstrate here that expression of the LXRalpha gene is highly induced in vivo and in vitro in mouse and human adipocytes in the presence of the synthetic LXR agonist T0901317. This autoregulation is caused by an identified LXR-responsive element motif in the mouse LXRalpha promoter, which is conserved in the human LXRalpha promoter. Using different LXR-deficient mice, we demonstrate that the basal expression level of LXRalpha is increased in LXRbeta( /-) mice, whereas the basal expression level of LXRbeta is unchanged in LXRalpha( /-) mice. The two LXRs can compensate for each other in mediating ligand activated regulation of LXR target genes involved in lipid homeostasis in adipose tissue. Sterol regulatory element binding protein-1 (SREBP-1), ATP binding cassette transporter A1 (ABCA1), ABCG1, as well as apolipoprotein E (apoE) are induced in vivo by T0901317 in wild-type, LXRalpha(-/-) or LXRbeta(-/-) mice but not in LXRalpha(-/-)beta(-/-) mice. Although SREBP-1 and ABCG1 are induced in liver, muscle, and adipose tissue, the apoE, glucose transporter-4 (GLUT4), and LXRalpha genes are specifically induced only in adipose tissue. We suggest that an important aspect of LXRalpha autoregulation in adipose tissue may be to increase the level of LXRalpha over a threshold level necessary to induce the expression of certain target genes. PMID- 15292369 TI - Oligomerization state-dependent hyperlipidemic effect of angiopoietin-like protein 4. AB - Angiopoietin-like protein 4 (Angptl4) is the second member of the angiopoietin like family of proteins previously shown to increase plasma triglyceride (TG) levels in vivo. We recently reported that Angptl4 is a variable-sized oligomer formed by intermolecular disulfide bonds and undergoes regulated proteolytic processing upon secretion. We now show that adenoviral overexpression of Angptl4 potently increases plasma TG levels by a mechanism independent of food intake or hepatic VLDL secretion. We determined that cysteine residues at positions 76 and 80 of Angptl4, conserved among mouse, rat, and human, are required to form higher order structures. By generating adenoviral expression vectors of Angptl4 containing different epitope tags at both N and C termini, we show that loss of oligomerization results in decreased stability of the N-terminal coiled-coil domain of Angptl4 as well as decreased ability to increase plasma TG levels, suggesting that intermolecular disulfide bond formation plays important roles in determining the magnitude of the hyperlipidemic effect of Angptl4. Because Angptl4 is more potent than Angptl3 in increasing plasma TG levels in mice, inappropriate oligomerization of Angptl4 could be associated with disorders of lipid metabolism in vivo. PMID- 15292370 TI - Polymorphisms in the gene encoding lipoprotein lipase in men with low HDL-C and coronary heart disease: the Veterans Affairs HDL Intervention Trial. AB - Our goal was to further define the role of LPL gene polymorphisms in coronary heart disease (CHD) risk. We determined the frequencies of three LPL polymorphisms (D9N, N291S, and S447X) in 899 men from the Veterans Affairs HDL Intervention Trial (VA-HIT), a study that examined the potential benefits of increasing HDL with gemfibrozil in men with established CHD and low high density lipoprotein cholesterol (HDL-C; < or =40 mg/dl), and compared them with those of men without CHD from the Framingham Offspring Study (FOS). In VA-HIT, genotype frequencies for LPL D9N, N291S, and S447X were 5.3, 4.5, and 13.0%, respectively. These values differed from those for men in FOS having an HDL-C of >40, who had corresponding values of 3.2% (P = 0.06), 1.5% (P < 0.01), and 18.2% (P < 0.01). On gemfibrozil, carriers of the LPL N9 allele in VA-HIT had lower levels of large LDL (-32%; P < 0.01) but higher levels of small, dense LDL (+59%; P < 0.003) than did noncarriers. Consequently, mean LDL particle diameter was smaller in LPL N9 carriers than in noncarriers (20.14 +/- 0.87 vs. 20.63 +/- 0.80 nm; P < 0.003). In men with low HDL-C and CHD: 1) the LPL N9 and S291 alleles are more frequent than in CHD-free men with normal HDL-C, whereas the X447 allele is less frequent, and 2) the LPL N9 allele is associated with the LDL subclass response to gemfibrozil. PMID- 15292371 TI - Novel fatty acid elongases and their use for the reconstitution of docosahexaenoic acid biosynthesis. AB - In algae, the biosynthesis of docosahexaenoic acid (22:6omega3; DHA) proceeds via the elongation of eicosapentaenoic acid (20:5omega3; EPA) to 22:5omega3, which is required as a substrate for the final Delta4 desaturation. To isolate the elongase specific for this step, we searched expressed sequence tag and genomic databases from the algae Ostreococcus tauri and Thalassiosira pseudonana, from the fish Oncorhynchus mykiss, from the frog Xenopus laevis, and from the sea squirt Ciona intestinalis using as a query the elongase sequence PpPSE1 from the moss Physcomitrella patens. The open reading frames of the identified elongase candidates were expressed in yeast for functional characterization. By this, we identified two types of elongases from O. tauri and T. pseudonana: one specific for the elongation of (Delta6-)C18-PUFAs and one specific for (Delta5-)C20-PUFAs, showing highest activity with EPA. The clones isolated from O. mykiss, X. laevis, and C. intestinalis accepted both C18- and C20-PUFAs. By coexpression of the Delta6- and Delta5-elongases from T. pseudonana and O. tauri, respectively, with the Delta5- and Delta4-desaturases from two other algae we successfully implemented DHA synthesis in stearidonic acid-fed yeast. This may be considered an encouraging first step in future efforts to implement this biosynthetic sequence into transgenic oilseed crops. PMID- 15292372 TI - Genetic variants of the lipoprotein lipase gene and myocardial infarction in the Central Valley of Costa Rica. AB - To assess common variants of the LPL gene that could influence susceptibility to myocardial infarction (MI), we assessed three functional single-nucleotide polymorphisms (SNPs), D9N, N291S, and S447X, in 1,321 survivors of a first acute MI and 1,321 population-based controls, matched for age, gender, and area of residence, all living in the Central Valley of Costa Rica. Conditional logistic regression was used to estimate odds ratio (OR) and 95% confidence interval (CI). The frequency of the X447 mutant allele was significantly lower in cases than in controls (6.2% vs. 7.6%; P < 0.01), whereas no association with MI was found for D9N or N291S. The OR (95% CI) for carriers vs. noncarriers of the X447 allele was 0.80 (0.63-1.01); when considering the haplotype that contained X447 and normal alleles of D9N and N291S, the OR (95% CI) was 0.66 (0.48-0.91). Twelve other SNPs were assessed in a subgroup of the population, of which the four functional SNPs were found to be monomorphic, and no correlation with MI was observed for the other eight neutral SNPs. The X447 mutant allele of the LPL gene may protect from MI risk, although this effect is small. PMID- 15292373 TI - Model class A and class L peptides increase the production of apoA-I-containing lipoproteins in HepG2 cells. AB - Class A peptides inhibit atherosclerosis and protect cells from class L peptide mediated lysis. Because the cytolytic process is concentration dependent, we hypothesized that at certain concentrations both classes of peptides exert similar effect(s) on cells. To test this hypothesis, we studied the effects of a class L peptide (18L = GIKKFLGSIWKFIKAFVG) and a class A peptide, 18A-Pro-18A (18A = DWLKAFYDKVAEKLKEAF) (37pA), on apolipoprotein and lipoprotein production in HepG2 cells. Secretion of (35)S-labeled apolipoprotein A-I (apoA-I) was stimulated by both 18L (110%) and 37pA (135%) at 10 and 20 nM of peptides, respectively. Both peptides enhanced the secretion of (3)H-labeled phospholipids by 140% and (14)C-labeled HDL-cholesterol (HDL-C) by 35% but had no significant effect on the total cholesterol mass or secretion. These results indicate that class L and class A peptides cause redistribution of cholesterol among lipoproteins in favor of HDL-C. Both peptides remodeled apoA-I-containing particles forming prebeta- as well as alpha-HDL. This study suggests that increased secretion of phospholipids and apoA-I and the formation of prebeta-HDL particles might contribute to the antiatherogenic properties of these peptides. PMID- 15292374 TI - Gene-selective modulation by a synthetic oxysterol ligand of the liver X receptor. AB - Liver X receptors (LXRs) play key roles in the regulation of cholesterol homeostasis by limiting cholesterol accumulation in macrophages within arterial wall lesion sites by a mechanism that includes the upregulation of ATP binding cassette transporters. These atheroprotective properties distinguish LXRs as potential targets for pharmaceutical intervention in cardiovascular disease. Their associated activity for promoting lipogenesis and triglyceride accretion through the activation of sterol-response element binding protein 1c (SREBP-1c) expression, however, represents a potential proatherogenic liability. A newly characterized synthetic oxysterol, N,N-dimethyl-3beta-hydroxycholenamide (DMHCA), represents a gene-selective LXR modulator that mediates potent transcriptional activation of ABCA1 gene expression while exhibiting minimal effects on SREBP-1c both in vitro and in vivo in mice. DMHCA has the potential to stimulate cholesterol transport through the upregulation of LXR target genes, including ABCA1, in liver, small intestine, and peritoneal macrophages. Compared with known nonsteroidal LXR agonists, however, DMHCA exhibits only limited activity for increasing hepatic SREBP-1c mRNA and does not alter circulating plasma triglycerides. Cell-based studies also indicate that DMHCA enhances cholesterol efflux in macrophages and suggest a mechanism whereby this selective modulator can potentially inhibit cholesterol accumulation. DMHCA and related gene selective ligands of LXR may have application to the study and treatment of atherosclerosis. PMID- 15292375 TI - Intracellular cholesterol mobilization involved in the ABCA1/apolipoprotein mediated assembly of high density lipoprotein in fibroblasts. AB - Differential regulation has been suggested for cellular cholesterol and phospholipid release mediated by apolipoprotein A-I (apoA-I)/ABCA1. We investigated various factors involved in cholesterol mobilization related to this pathway. ApoA-I induced a rapid decrease of the cellular cholesterol compartment that is in equilibrium with the ACAT-accessible pool in cells that generate cholesterol-rich HDL. Pharmacological and genetic inactivation of ACAT enhanced the apoA-I-mediated cholesterol release through upregulation of ABCA1 and through cholesterol enrichment in the HDL generated. Pharmacological activation of protein kinase C (PKC) also decreased the ACAT-accessible cholesterol pool, not only in the cells that produce cholesterol-rich HDL by apoA-I (i.e., human fibroblast WI-38 cells) but also in the cells that generate cholesterol-poor HDL (mouse fibroblast L929 cells). In L929 cells, the PKC activation caused an increase in apoA-I-mediated cholesterol release without detectable change in phospholipid release and in ABCA1 expression. These results indicate that apoA-I mobilizes intracellular cholesterol for the ABCA1-mediated release from the compartment that is under the control of ACAT. The cholesterol mobilization process is presumably related to PKC activation by apoA-I. PMID- 15292376 TI - Thematic review series: The pathogenesis of atherosclerosis. Toward a biological network for atherosclerosis. AB - The goal of systems biology is to define all of the elements present in a given system and to create an interaction network between these components so that the behavior of the system, as a whole and in parts, can be explained under specified conditions. The elements constituting the network that influences the development of atherosclerosis could be genes, pathways, transcript levels, proteins, or physiologic traits. In this review, we discuss how the integration of genetics and technologies such as transcriptomics and proteomics, combined with mathematical modeling, may lead to an understanding of such networks. PMID- 15292377 TI - Adjuvant therapy for colon cancer: a historical perspective. PMID- 15292378 TI - Predicting sensitivity of non-small-cell lung cancer to gefitinib: is there a role for P-Akt? PMID- 15292379 TI - Environmental protection: studies highlight importance of tumor microenvironment. PMID- 15292380 TI - Cell of origin: mouse model offers insights into process of malignancy. PMID- 15292381 TI - Results of cancer care quality study lead to cautious optimism, caveats. PMID- 15292382 TI - Stat bite: Health insurance coverage in the United States by age group, 2003. PMID- 15292383 TI - Project creates repository for microarray datasets. PMID- 15292384 TI - Randomized trial of adjuvant therapy in colon carcinoma: 10-year results of NSABP protocol C-01. AB - BACKGROUND: The National Surgical Adjuvant Breast and Bowel Project C-01 trial reported in 1988 that, for patients with adenocarcinoma of the colon, compared with surgery alone, 1) postoperative chemotherapy with 1-(2-chloroethyl)-3-(4 trans-methylcyclohexyl)-1-nitrosourea (i.e., MeCCNU or semustine), vincristine, and 5-fluorouracil was associated with better 5-year disease-free and overall survival and 2) postoperative immunotherapy with bacillus Calmette-Guerin was associated with better 5-year overall, but not disease-free, survival. We now provide a 10-year update of this trial. METHODS: Between November 11, 1977, and February 28, 1983, 1166 patients with resected Dukes' stage B and C adenocarcinoma of the colon were stratified by Dukes' stage, sex, and age (<65 years or > or =65 years) and then randomly assigned to receive no further treatment (surgery alone; 394 patients), adjuvant chemotherapy (379 patients), or adjuvant immunotherapy (393 patients). Those eligible for follow-up included 375 (95.2%) patients in the surgery-alone group, 349 (92.1%) patients in the adjuvant chemotherapy group, and 372 (94.7%) patients in the adjuvant-immunotherapy group. All statistical tests were two-sided. RESULTS: No difference was observed between patients in the chemotherapy group and those in the surgery-alone group in 10 year disease-free survival (hazard ratio [HR] = 1.14, 95% confidence interval [CI] = 0.94 to 1.39;P =.17) or overall survival (HR = 1.12, 95% CI = 0.91 to 1.38; P=.27). Immunotherapy did not appear to prevent tumor relapse after 10 years (for surgery alone versus immunotherapy, relative risk [RR] = 0.99, 95% CI = 0.78 to 1.25; P =.93) but had a beneficial effect on 10-year overall survival (for surgery alone versus immunotherapy, RR = 1.27, 95% CI = 1.03 to 1.56; P =.02) that apparently results from a reduction in deaths associated with comorbidities in the immunotherapy group. CONCLUSION: The disease-free and overall survival benefit associated with chemotherapy in this patient population is of limited duration, disappearing after 10 years. PMID- 15292385 TI - Akt phosphorylation and gefitinib efficacy in patients with advanced non-small cell lung cancer. AB - BACKGROUND: Gefitinib, a specific epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor, has activity against approximately 10% of unselected non-small-cell lung cancer (NSCLC) patients. Phosphatidylinositol 3'-kinase (PI3K)/Akt and Ras/Raf/mitogen-activated protein kinase (MAPK), the two main EGFR signaling pathways, mediate EGFR effects on proliferation and survival. Because activation of these pathways is dependent on the phosphorylation status of the components, we evaluated the association between phosphorylation status of Akt (P Akt) and MAPK (P-MAPK) and gefitinib activity in patients with advanced NSCLC. METHODS: Consecutive patients (n = 106) with NSCLC who had progressed or relapsed on standard therapy received gefitinib (250 mg/day) until disease progression, unacceptable toxicity, or patient refusal. P-Akt and P-MAPK positivity was determined with immunohistochemistry using tumor tissues obtained before any anticancer treatment. Association of P-Akt and time to progression was determined by univariable and multivariable analyses. All statistical tests were two-sided. RESULTS: Of the 103 evaluable patients, 51 (49.5%) had tumors that were positive for P-Akt, and 23 (22.3%) had tumors that were positive for P-MAPK. P-Akt positivity status was statistically significantly associated with being female (P<.001), with never-smoking history (P =.004), and with bronchioloalveolar carcinoma histology (P =.034). Compared with patients whose tumors were negative for P-Akt, patients whose tumors were positive for P-Akt had a better response rate (26.1% versus 3.9%; P =.003), disease control rate (60.9% versus 23.5%; P<.001), and time to progression (5.5 versus 2.8 months; P =.004). Response rate, disease control rate, and time to progression did not differ according to P-MAPK status. The multivariable analysis showed that P-Akt positivity was associated with a reduced risk of disease progression (hazard ratio = 0.58, 95% confidence interval = 0.35 to 0.94). CONCLUSIONS: Patients with P-Akt-positive tumors who received gefitinib had a better response rate, disease control rate, and time to progression than patients with P-Akt-negative tumors, suggesting that gefitinib may be most effective in patients with basal Akt activation. PMID- 15292386 TI - Her-2/neu gene amplification and response to paclitaxel in patients with metastatic breast cancer. AB - BACKGROUND: HER-2/neu overexpression appears to be associated with improved response to anthracycline-based chemotherapy, but its association with response to taxane-based chemotherapy is unclear. In this retrospective subset analysis of patients with metastatic breast cancer enrolled in a randomized treatment trial, we investigated the response of patients with known HER-2/neu status to treatment with taxane-based epirubicin-paclitaxel (ET) chemotherapy compared with treatment with epirubicin-cyclophosphamide (EC) chemotherapy. METHODS: HER-2/neu status (positive [i.e., HER-2/neu amplification] or negative [i.e., no HER-2/neu amplification]) of archival specimens of primary tumors from 297 patients with metastatic breast cancer was determined by use of fluorescence in situ hybridization. Associations between HER-2/neu status and the efficacy of randomly assigned chemotherapy (ET versus EC) were investigated. All statistical tests were two-sided. RESULTS: Patients with HER-2/neu-positive tumors had a statistically significantly greater objective response rate than patients with HER-2/neu-negative tumors to treatment with ET (76% versus 50%, respectively; P =.005) but not to treatment with EC (46% versus 33%; P =.130). The objective response rate associated with ET was greater than that associated with EC for both HER-2/neu-positive tumors (76% versus 46%; P =.004) and HER-2/neu-negative tumors (50% versus 33%; P =.002). However, the improvement in the objective response rate associated with ET, compared with that associated with EC, was greater for patients with HER-2/neu-positive tumors (adjusted odds ratio [OR] = 3.64, 95% confidence interval [CI] = 1.48 to 8.92; P=.005) than for patients with HER-2/neu-negative tumors (adjusted OR = 1.92, 95% CI = 1.01 to 3.64; P=.046). Among patients with HER-2/neu-positive tumors, those who received ET had better progression-free survival and overall survival than those who received EC (for progression-free survival, adjusted relative risk [RR] = 0.65, 95% CI = 0.42 to 1.02; P=.062; for overall survival, adjusted RR = 0.60, 95% CI = 0.36 to 1.02; P=.059). However, among patients with HER-2/neu-negative tumors, those who received ET and those who received EC had similar progression-free survival and overall survival. CONCLUSIONS: HER-2/neu amplification does not adversely influence response to first-line chemotherapy with either ET or EC. Furthermore, a taxane-containing regimen such as ET may provide a preferential benefit to patients with HER-2/neu-positive tumors. PMID- 15292387 TI - Serum high-density lipoprotein cholesterol, metabolic profile, and breast cancer risk. AB - BACKGROUND: The prevalence of metabolic syndrome (obesity, glucose intolerance, low serum high-density lipoprotein cholesterol [HDL-C], high serum triglycerides, hypertension) is high and increasing in parallel with an increasing breast cancer incidence worldwide. HDL-C represents an important aspect of the syndrome, yet its role in breast cancer is still undefined. METHODS: In two population-based screening surveys during 1977-1983 and 1985-1987, serum HDL-C was assayed enzymatically among 38,823 Norwegian women aged 17-54 years at entry. Height, weight, blood pressure, serum lipids, fat and energy intake, physical activity, parity, oral contraceptive use, hormone therapy use, alcohol intake, and tobacco use were also assessed. We used Cox proportional hazards modeling to estimate the relative risk (RR) of breast cancer associated with serum HDL-C levels and to adjust for potential confounding variables. We performed stratified analyses to evaluate effect modification by body mass index (BMI) and menopausal status. All statistical tests were two-sided. RESULTS: During a median follow-up of 17.2 years, we identified 708 cases of invasive breast cancer. In multivariable analysis, the risk of postmenopausal breast cancer was inversely related to quartile of HDL-C (P(trend) =.02). Among women with HDL-C above 1.64 mmol/L (highest quartile) versus below 1.20 mmol/L (lowest quartile), the relative risk was 0.75 (95% confidence interval [CI] = 0.58 to 0.97). The HDL-C association was confined to women in the heavier subgroup (BMI > or =25 kg/m2), for whom the relative risk of postmenopausal breast cancer in those with HDL-C above 1.64 mmol/L versus below 1.20 mmol/L was 0.43 (95% CI = 0.28 to 0.67; P(trend)<.001; P(interaction) =.001). CONCLUSION: Low HDL-C, as part of the metabolic syndrome, is associated with increased postmenopausal breast cancer risk. PMID- 15292388 TI - Associations among beta-TrCP, an E3 ubiquitin ligase receptor, beta-catenin, and NF-kappaB in colorectal cancer. AB - BACKGROUND: The ubiquitin-proteasome pathway is important in regulating protein signaling pathways that are involved in tumorigenesis. beta-transducin repeat containing proteins (beta-TrCP) are components of the ubiquitin ligase complex targeting beta-catenin and IkappaBalpha for proteasomal degradation and are thus a negative regulator of Wnt/beta-catenin signaling and a positive regulator of NF kappaB signaling. We analyzed expression of beta-TrCP in colorectal cancers and its association with types of beta-catenin subcellular localization, an indirect measure of activation. METHODS: Levels of beta-TrCP1 mRNA and protein were measured by quantitative reverse transcription-polymerase chain reaction and immunoblotting, respectively, in samples of tumor and normal tissues from 45 patients with colorectal cancer. Types of beta-catenin activation (diffuse or invasion edge) and NF-kappaB activation were examined by immunohistochemistry. Apoptosis was determined by the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick-end labeling (TUNEL) assay. All statistical tests were two-sided. RESULTS: Compared with the beta-TrCP1 levels in normal tissues, 25 (56%) of 45 tumors had increased beta-TrCP1 mRNA and protein levels. Of the 22 (49%) tumors with beta-catenin activation, 12 had the diffuse type (i.e., nuclear accumulation throughout the tumor) and 10 had the invasion edge type (i.e., nuclear accumulation predominantly in the tumor cells that formed the invasion edge). Increased beta-TrCP1 levels were statistically significantly associated with beta-catenin activation (P =.023) and decreased apoptosis (P =.035). beta-TrCP accumulated in the nuclei of tumor cells that contained increased levels of beta-TrCP1 mRNA and the active form of NF-kappaB. Higher levels of beta-TrCP1 mRNA were detected in primary tumors of patients who had metastases (0.960 arbitrary units, 95% confidence interval = 0.878 to 1.042) than in the tumors of patients who did not (0.722 arbitrary units, 95% confidence interval = 0.600 to 0.844; P =.016). CONCLUSION: In colorectal cancer, increased expression of beta-TrCP1 is associated with activation of both beta-catenin and NF-kappaB, suggesting that the integration of these signaling pathways by increased beta-TrCP expression may contribute to an inhibition of apoptosis and tumor metastasis. PMID- 15292389 TI - Immune cell-mediated antitumor activities of GD2-targeted liposomal c-myb antisense oligonucleotides containing CpG motifs. AB - BACKGROUND: Expression of the c-myb proto-oncogene in neuroblastoma, the most common extracranial solid tumor of infancy, is linked with cell proliferation and differentiation. Neuroblastoma can be selectively targeted via monoclonal antibodies against the disialoganglioside (GD2) tumor-associated antigen. Liposomes coated with anti-GD2 antibodies (targeted liposomes) and entrapping a c myb antisense oligonucleotide have antitumor activity. Because antisense oligonucleotides containing CpG motifs can stimulate immune responses, we evaluated the effect of CpG-containing c-myb antisense oligonucleotides encapsulated within targeted liposomes. METHODS: Antisense (myb-as) and scrambled (myb-scr) control oligonucleotides with CpG motifs were encapsulated within GD2 targeted and non-targeted liposomes. Two murine (nude and SCID-bg) xenograft models of neuroblastoma were established. Mice (groups of 10) were injected intravenously with various oligonucleotide and liposome formulations, and life span, long-term survival, immune cell activation, and cytokine release were measured over time. RESULTS: Tumor-bearing mice injected with targeted liposome CpG-myb-as or targeted liposome-CpG-myb-scr lived longer than mice in any other group, although long-term survival (i.e., more than 120 days) was obtained only in mice injected with targeted liposome-CpG-myb-as. Splenocytes isolated from mice injected with targeted liposome-CpG-myb-as contained activated macrophages, B cells, and natural killer (NK) cells, but only activated NK cells were associated with antitumor cytotoxic activity. In vivo immune cell activation was accompanied by the time-dependent increases in plasma levels of the cytokines interleukin 12 (IL-12; maximum level reached by 2 hours) and interferon gamma (IFN-gamma; maximum level reached by 18 hours) and was dependent on the oligonucleotide CpG motif. Ablation of macrophages or NK cells resulted in a loss of in vivo antitumor activity. CONCLUSION: Immune cell activation, involving the time-dependent activation of macrophages and NK cells, contributes to the antitumor activity of targeted liposome-CpG-myb-as against neuroblastoma and could improve the effectiveness of antitumor targeted liposomes. PMID- 15292390 TI - Re: Human papillomavirus in oral exfoliated cells and risk of head and neck cancer. PMID- 15292391 TI - Re: A Randomized, placebo-controlled trial of zoledronic acid in patients with hormone-refractory metastatic prostate carcinoma. PMID- 15292392 TI - Re: BRCA1 and BRCA2 founder mutations and the risk of colorectal cancer. PMID- 15292393 TI - Re: Decreasing women's anxieties after abnormal mammograms: a controlled trial. PMID- 15292394 TI - Biannual report of the Cochrane Haematological Malignancies Group. PMID- 15292395 TI - AtSGP1, AtSGP2 and MAP4K alpha are nucleolar plant proteins that can complement fission yeast mutants lacking a functional SIN pathway. AB - In the fission yeast Schizosaccharomyces pombe, the onset of septum formation is signalled via the septation initiation network (SIN) involving several protein kinases and a GTPase. Arabidopsis thaliana and Brassica napus proteins homologous to fission yeast spg1p (AtSGP1, AtSGP2), cdc7p (AtMAP3K epsilon 1, AtMAP3K epsilon 2, BnMAP3K epsilon 1) and sid1p (AtMAP4K alpha 1, AtMAP4K alpha 2, BnMAP4K alpha 2) exhibit a significant similarity. The plant proteins AtSGP1/2 and BnMAP4K alpha 2 are able to complement the S. pombe mutant proteins spg1-B8 and sid1-239, respectively and to induce mutisepta when overexpressed in wild type yeast. Yeast two-hybrid assays demonstrated interactions both between plant proteins and between plant and yeast proteins of the SIN pathway. However, the primary structure of the proteins as well as the partial complementation of yeast mutants indicates that plant homologous proteins and their yeast counterparts have diverged during evolution. Real-time RT-PCR studies demonstrated plant SIN related gene expression in all organs tested and a co-expression pattern during the cell cycle, with a higher accumulation at G(2)-M. During interphase, the plant SIN-related proteins were found to co-localise predominantly in the nucleolus of the plant cells, as shown by fusions to green fluorescent protein. These data suggest the existence of a plant SIN-related pathway. PMID- 15292396 TI - Sorting nexin 16 regulates EGF receptor trafficking by phosphatidylinositol-3 phosphate interaction with the Phox domain. AB - Sorting nexins (SNXs) containing the Phox (PX) domain are implicated in the regulation of membrane trafficking and sorting processes of epithelial growth factor receptor (EGFR). In this study, we investigated whether SNX16 regulates EGF-induced cell signaling by regulating EGFR trafficking. SNX16 is localized in early and recycling endosomes via its PX domain. Mutation of the PX domain disrupted the association between SNX16 and phosphatidylinositol 3-phosphate [PtdIns(3)P]. Treatment with wortmannin, a PtdIns 3-kinase inhibitor, abolished the endosomal localization of SNX16, suggesting that the intracellular localization of SNX16 is regulated by PtdIns 3-kinase activity. SNX16 was found to associate with EGFR after stimulation with EGF in COS-7 cells. Moreover, overexpression of SNX16 increased the rate of EGF-induced EGFR degradation and inhibited the EGF-induced up-regulation of ERK and serum response element (SRE). In addition, mutation in the PX domain significantly blocked the inhibitory effect of SNX16 on EGF-induced activation of ERK and SRE. From these results, we suggest that SNX16 directs the sorting of EGFR to the endosomal compartment and thus regulates EGF-induced cell signaling. PMID- 15292397 TI - Neurogenesis of Rhesus adipose stromal cells. AB - In this study, we isolated and characterized a population of non-human primate adipose tissue stromal cells (pATSCs) containing multipotent progenitor cells. We show that these pATSCs can differentiate into several mesodermal lineages, as well as neural lineage cells. For neural induction of pATSCs and non-human primate bone marrow stromal cells (pBMSCs), the cells were cultured in Neurobasal (NB) media supplemented with B27, basic fibroblast growth factor (bFGF), brain derived neurotrophic factor (BDNF) and epidermal growth factor (EGF). After 4 days in culture, the pATSCs form compact, spheroid bodies that ultimately become neurospheres (NS). Free-floating neurospheres undergo extensive differentiation when cultured on PDL-laminin. Our data suggest that the neurogenic potential of pATSCs is markedly higher than that of pBMSCs. We have also performed microarray analysis and characterized the gene expression patterns in undifferentiated pATSCs. The direct comparison of gene expression profiles in undifferentiated pATSCs and pATSC-NS, and delineated specific members of important growth factor, signaling, cell adhesion and transcription factors families. Our data indicate that adipose tissue may be an alternative source of stem cells for therapy of central nervous system (CNS) defects. PMID- 15292398 TI - Sialic acid residues on astrocytes regulate neuritogenesis by controlling the assembly of laminin matrices. AB - In the developing nervous system migrating neurons and growing axons are guided by diffusible and/or substrate-bound cues, such as extracellular matrix associated laminin. In a previous work we demonstrated that laminin molecules could self-assemble in two different manners, giving rise to matrices that could favor either neuritogenesis or proliferation of cortical precursor cells. We investigated whether the ability of astrocytes to promote neuritogenesis of co cultivated neurons was modulated by the assembling mode of the laminin matrix secreted by them. We compared the morphologies and neuritogenic potentials of laminin deposited by in vitro-differentiated astrocytes obtained from embryonic or neonatal rat brain cortices. We showed that, while permissive astrocytes derived from embryonic brain produced a flat laminin matrix that remained associated to the cell surface, astrocytes derived from newborn brain secreted a laminin matrix resembling a fibrillar web that protruded from the cell plane. The average neurite lengths obtained for E16 neurons cultured on each astrocyte layer were 198+/-22 and 123+/-13 microm, respectively. Analyses of surface-associated electrostatic potentials revealed that embryonic astrocytes presented a pI of 2.8, while in newborn cells this value was -3.8. Removal of the sialic acid groups on the embryonic monolayer by neuraminidase treatment led to the immediate release of matrix-associated laminin. Interestingly, laminin reassembled 1 hour after neuraminidase removal converted to the features of the newborn matrix. Alternatively, treatment of astrocytes with the cholesterol-solubilizing detergent methyl-beta-cyclodextrin also resulted in release of the extracellular laminin. To test the hypothesis that sialic-acid-containing lipids localized at cholesterol-rich membrane domains could affect the process of laminin assembly, we devised a cell-free assay where laminin polymerization was carried out over artificial lipid films. Films of either a mixture of gangliosides or pure ganglioside GT1b induced formation of matrices of morpho-functional features similar to the matrices deposited by embryonic astrocytes. Conversely, films of phosphatidylcholine or ganglioside GM1 led to the formation of bulky laminin aggregates that lacked a defined structure. We propose that the expression of negative lipids on astrocytes can control the extracellular polymerization of laminin and, consequently, the permissivity to neuritogenesis of astrocytes during development. PMID- 15292399 TI - Simulation of calcium waves in ascidian eggs: insights into the origin of the pacemaker sites and the possible nature of the sperm factor. AB - Fertilization triggers repetitive waves of cytosolic Ca(2+) in the egg of many species. The mechanism involved in the generation of Ca(2+) waves has been studied in much detail in mature ascidian eggs, by raising artificially the level of inositol 1,4,5-trisphosphate [Ins(1,4,5)P(3)] or of its poorly metabolizable analogue, glycero-myo-phosphatidylinositol 4,5-bisphosphate [gPtdIns(4,5)P(2)]. Here, we use this strategy and the experimental results it provides to develop a realistic theoretical model for repetitive Ca(2+) wave generation and propagation in mature eggs. The model takes into account the heterogeneous spatial distribution of the endoplasmic reticulum. Our results corroborate the hypothesis that Ca(2+) wave pacemakers are associated with cortical accumulations of endoplasmic reticulum. The model is first tested and validated by the adequate match between its theoretical predictions and the observed effects of localized injections of massive amounts of Ins(1,4,5)P(3) analogues. In a second step, we use the model to make some propositions about the possible characteristics of the sperm factor. We find that to account for the spatial characteristics of the first series of Ca(2+) waves seen at fertilization in ascidian eggs, it has to be assumed that, if the sperm factor is a phospholipase C, it is Ca(2+)-sensitive and highly diffusible. Although the actual state of knowledge does not allow us to explain the observed relocalization of the Ca(2+) wave pacemaker site, the model corroborates the assumption that PtdIns(4,5)P(2), the substrate for phospholipase C is distributed over the entire egg. We also predict that the dose of sperm factor injected into the egg should modulate the temporal characteristics of the first, long-lasting fertilization wave. PMID- 15292400 TI - Alfy, a novel FYVE-domain-containing protein associated with protein granules and autophagic membranes. AB - Phosphatidylinositol-3-phosphate [PtdIns(3)P] regulates endocytic and autophagic membrane traffic. In order to understand the downstream effects of PtdIns(3)P in these processes, it is important to identify PtdIns(3)P-binding proteins, many of which contain FYVE zinc-finger domains. Here, we describe a novel giant FYVE domain-containing protein, named autophagy-linked FYVE protein (Alfy). Alfy is ubiquitously expressed, shares sequence similarity with the Chediak-Higashi syndrome protein and has putative homologues in flies, nematodes and fission yeast. Alfy binds PtdIns(3)P in vitro and partially colocalizes with PtdIns(3)P in vivo. Unlike most other FYVE-domain proteins, Alfy is not found on endosomes but instead localizes mainly to the nuclear envelope. When HeLa cells are starved or treated with a proteasome inhibitor, Alfy relocalizes to characteristic filamentous cytoplasmic structures located close to autophagic membranes and ubiquitin-containing protein aggregates. By electron microscopy, similar structures can be found within autophagosomes. We propose that Alfy might target cytosolic protein aggregates for autophagic degradation. PMID- 15292401 TI - Identification of the Drosophila interband-specific protein Z4 as a DNA-binding zinc-finger protein determining chromosomal structure. AB - The subdivision of polytene chromosomes into bands and interbands suggests a structural chromatin organization that is related to the formation of functional domains of gene expression. We made use of the antibody Z4 to gain insight into this level of chromosomal structure, as the Z4 antibody mirrors this patterning by binding to an antigen that is present in most interbands. The Z4 gene encodes a protein with seven zinc fingers, it is essential for fly development and acts in a dose-dependent manner on the development of several tissues. Z4 mutants have a dose-sensitive effect on w(m4) position effect variegation with a haplo suppressor and triplo-enhancer phenotype, suggesting Z4 to be involved in chromatin compaction. This assumption is further supported by the phenotype of Z4 mutant chromosomes, which show a loss of the band/interband pattern and are subject to an overall decompaction of chromosomal material. By co immunoprecipitations we identified a novel chromo domain protein, which we named Chriz (Chromo domain protein interacting with Z4) as an interaction partner of Z4. Chriz localizes to interbands in a pattern that is identical to the Z4 pattern. These findings together with the result that Z4 binds directly to DNA in vitro strongly suggest that Z4 in conjunction with Chriz is intimately involved in the higher-order structuring of chromosomes. PMID- 15292402 TI - Trichostatin A-induced histone acetylation causes decondensation of interphase chromatin. AB - The effect of trichostatin A (TSA)-induced histone acetylation on the interphase chromatin structure was visualized in vivo with a HeLa cell line stably expressing histone H2A, which was fused to enhanced yellow fluorescent protein. The globally increased histone acetylation caused a reversible decondensation of dense chromatin regions and led to a more homogeneous distribution. These structural changes were quantified by image correlation spectroscopy and by spatially resolved scaling analysis. The image analysis revealed that a chromatin reorganization on a length scale from 200 nm to >1 microm was induced consistent with the opening of condensed chromatin domains containing several Mb of DNA. The observed conformation changes could be assigned to the folding of chromatin during G1 phase by characterizing the effect of TSA on cell cycle progression and developing a protocol that allowed the identification of G1 phase cells on microscope coverslips. An analysis by flow cytometry showed that the addition of TSA led to a significant arrest of cells in S phase and induced apoptosis. The concentration dependence of both processes was studied. PMID- 15292403 TI - Cpc1, a Chlamydomonas central pair protein with an adenylate kinase domain. AB - Mutations at CPC1 disrupt assembly of a central pair microtubule-associated complex and alter flagellar beat frequency in Chlamydomonas. Sequences of wild type genomic clones that complement cpc1, and of corresponding cDNAs, reveal the gene product to be a 205 kDa protein with two predicted functional domains, a single EF hand motif near the C-terminus and an unusual centrally located adenylate kinase domain. Homologs are expressed in mammals (testis and tracheal cilia) as well as ciliated lower eukaryotes. Western blots confirm that Cpc1 is one of six subunits in a 16S central pair-associated complex. Motility defects associated with cpc1 alleles in vivo are partially rescued in vitro by reactivation of axonemes or cell models in saturating concentrations of ATP; thus the Cpc1 complex is essential for maintaining normal ATP concentrations in the flagellum. PMID- 15292404 TI - p120 catenin is required for morphogenetic movements involved in the formation of the eyes and the craniofacial skeleton in Xenopus. AB - During Xenopus development, p120 transcripts are enriched in highly morphogenetic tissues. We addressed the developmental function of p120 by knockdown experiments and by expressing E-cadherin mutants unable to bind p120. This resulted in defective eye formation and provoked malformations in the craniofacial cartilage structures, derivatives of the cranial neural crest cells. Closer inspection showed that p120 depletion impaired evagination of the optic vesicles and migration of cranial neural crest cells from the neural tube into the branchial arches. These morphogenetic processes were also affected by p120-uncoupled cadherins or E-cadherin containing a deletion of the juxtamembrane domain. Irrespective of the manipulation that caused the malformations, coexpression of dominant-negative forms of either Rac1 or LIM kinase rescued the phenotypes. Wild type RhoA and constitutively active Rho kinase caused partial rescue. Our results indicate that, in contrast to invertebrates, p120 is an essential factor for vertebrate development and an adequate balance between cadherin activity and cytoskeletal condition is critical for correct morphogenetic movements. PMID- 15292405 TI - Decrease in neuromuscular control about the knee with maturation in female athletes. AB - BACKGROUND: Compared with male athletes, female athletes demonstrate increased dynamic valgus angulation of the knee during landing from a jump, although prior to maturation male and female athletes have similar forces and motions about the knee when they land from a jump. Our hypothesis was that musculoskeletal changes that accompany maturation result in poor neuromuscular control of the knee joint in female athletes. METHODS: One hundred and eighty-one middle-school and high school soccer and basketball players-100 girls and eighty-one boys-participated in the study. Dynamic control of the knee joint was measured kinematically by assessing medial knee motion and the lower-extremity valgus angle and was measured kinetically by assessing knee joint torques; the values were then compared between female and male athletes according to maturational stage. Lower extremity bone length was measured with three-dimensional kinematic analysis. RESULTS: Following the onset of maturation, the female athletes landed with greater total medial motion of the knees and a greater maximum lower-extremity valgus angle than did the male athletes. The girls also demonstrated decreased flexor torques compared with the boys as well as a significant difference between the maximum valgus angles of their dominant and nondominant lower extremities after maturation. CONCLUSIONS: After girls mature, they land from a jump differently than do boys, as measured kinematically and kinetically. PMID- 15292406 TI - The outcome of total knee arthroplasty in obese patients. AB - BACKGROUND: Evidence linking increased body weight to osteoarthritis of the knee and the high prevalence of obesity underscore the importance of defining the outcome of total knee arthroplasty in obese patients. The purpose of this study was to compare the clinical and radiographic results of total knee arthroplasties performed in obese patients with those of total knee arthroplasties performed in nonobese patients. METHODS: Clinical and radiographic data on seventy-eight total knee arthroplasties in sixty-eight obese patients were compared with data on a matched group of nonobese patients. The analysis was also performed after stratification of the obese group for the degree of obesity. All patients had the same prosthesis. The clinical data that were analyzed included the Knee Society objective and functional scores, patellofemoral symptoms, activity level, and complications. RESULTS: The percentage of knees with a Knee Society score of > or =80 points at an average of eighty months was 88% in the obese group, which was significantly lower than the 99% rate in the nonobese group at the same time. The morbidly obese subgroup had a significantly higher revision rate than did the nonobese group (p = 0.02). CONCLUSIONS: The results of the present study suggest that any degree of obesity, defined as a body mass index of > or =30, has a negative effect on the outcome of total knee replacement. PMID- 15292407 TI - Talar neck fractures: results and outcomes. AB - BACKGROUND: Talar neck fractures occur infrequently and have been associated with high complication rates. The purposes of the present study were to evaluate the rates of early and late complications after operative treatment of talar neck fractures, to ascertain the effect of surgical delay on the development of osteonecrosis, and to determine the functional outcomes after operative treatment of such fractures. METHODS: We retrospectively reviewed the records of 100 patients with 102 fractures of the talar neck who had been managed at a level-1 trauma center. All fractures had been treated with open reduction and internal fixation. Sixty fractures were evaluated at an average of thirty-six months (range, twelve to seventy-four months) after surgery. Complications and secondary procedures were reviewed, and radiographic evidence of osteonecrosis and posttraumatic arthritis was evaluated. The Foot Function Index and Musculoskeletal Function Assessment questionnaires were administered. RESULTS: Radiographic evidence of osteonecrosis was seen in nineteen (49%) of the thirty nine patients with complete radiographic data. However, seven (37%) of these nineteen patients demonstrated revascularization of the talar dome without collapse. Overall, osteonecrosis with collapse of the dome occurred in twelve (31%) of thirty-nine patients. Osteonecrosis was seen in association with nine (39%) of twenty-three Hawkins group-II fractures and nine (64%) of fourteen Hawkins group-III fractures. The mean time to fixation was 3.4 days for patients who had development of osteonecrosis, compared with 5.0 days for patients who did not have development of osteonecrosis. With the numbers available, no correlation could be identified between surgical delay and the development of osteonecrosis. Osteonecrosis was associated with comminution of the talar neck (p < 0.03) and open fracture (p < 0.05). Twenty-one (54%) of thirty-nine patients had development of posttraumatic arthritis, which was more common after comminuted fractures (p < 0.07) and open fractures (p = 0.09). Patients with comminuted fractures also had worse functional outcome scores. CONCLUSIONS: Fractures of the talar neck are associated with high rates of morbidity and complications. Although the numbers in the present series were small, no correlation was found between the timing of fixation and the development of osteonecrosis. Osteonecrosis was associated with talar neck comminution and open fractures, confirming that higher-energy injuries are associated with more complications and a worse prognosis. This finding was strengthened by the poor Foot Function Index and Musculoskeletal Function Assessment scores in these patients. We recommend urgent reduction of dislocations and treatment of open injuries. Proceeding with definitive rigid internal fixation of talar neck fractures after soft-tissue swelling has subsided may minimize soft-tissue complications. PMID- 15292408 TI - Anatomic location of the peroneal nerve at the level of the proximal aspect of the tibia: Gerdy's safe zone. AB - BACKGROUND: Injury to the peroneal nerve is one of the most serious complications in orthopaedic surgery. Because percutaneous procedures at the level of the proximal aspect of the tibia are becoming increasingly popular, it is critical to have a thorough knowledge of the trajectory of the peroneal nerve and its main branches at the level of the proximal aspect of the tibia. This anatomic study was conducted in an attempt to (1) define the anatomy of the common peroneal nerve and its branches in a three-dimensional fashion and (2) identify an anatomic landmark on the surface to help define a safe area that is void of the main nerve and its branches. METHODS: Thirty-one adult unembalmed cadaveric legs were dissected. The peroneal nerve was identified at the level of the posterior aspect of the lateral femoral condyle and was dissected distally to the level of its intramuscular branches. The relationship between the peroneal nerve and Gerdy's tubercle was explored, and the distances from the nerve and its branches to the tubercle were measured and recorded in millimeters. The average distances and standard deviations from Gerdy's tubercle to the neural structures were calculated. RESULTS: The course of the common peroneal nerve trunk and its anterior recurrent branch defined an arc with a circumference having an average radius of 45 mm. This circumferential trajectory was seen to be centered at the most prominent aspect of Gerdy's tubercle. CONCLUSIONS AND CLINICAL RELEVANCE: The path of the common peroneal nerve and its proximal branch were notable in two regards: their circular nature and their consistent relationship to the most prominent aspect of Gerdy's tubercle. With Gerdy's tubercle used as a landmark, the trajectory of the peroneal nerve can be easily defined at the level of the proximal aspect of the tibia and marked prior to the placement of devices and instrumentation, thereby avoiding damage to the peroneal nerve and its branches. PMID- 15292409 TI - Validation of a clinical prediction rule for the differentiation between septic arthritis and transient synovitis of the hip in children. AB - BACKGROUND: The differentiation between septic arthritis and transient synovitis of the hip in children can be difficult. The purpose of the present study was to validate a previously published clinical prediction rule for this differentiation in a new patient population. METHODS: We prospectively studied children who presented to a major children's hospital between 1997 and 2002 with an acutely irritable hip. As in the previous study, diagnoses of septic arthritis (fifty-one patients) and transient synovitis (103 patients) were operationally defined on the basis of the white blood-cell count in the joint fluid, the results of cultures of joint fluid and blood, and the clinical course. Univariate analysis and multiple logistic regression were used to compare the two groups. The predicted probability of septic arthritis of the hip from the prediction rule was compared with actual distributions in the current patient population. The area under the receiver operating characteristic curve was determined. RESULTS: The same four independent predictors of septic arthritis of the hip (a history of fever, non-weight-bearing, an erythrocyte sedimentation rate of 40 mm/hr, and a serum white blood-cell count of >12,000 cells/mm(3) (>12.0 x 10(9)/L)) were identified in the current patient population. The predicted probability of septic arthritis of the hip from the prediction rule was similar to the actual distributions in the current patient population. The area under the receiver operating characteristic curve for the current patient population was 0.86, compared with 0.96 in the original population. CONCLUSIONS: Clinical prediction rules typically demonstrate diminished performance in a new patient population because they are optimally modeled to the original data set. The previously published clinical prediction rule for the differentiation between septic arthritis and transient synovitis of the hip in children demonstrated diminished, but nevertheless very good, diagnostic performance in a new patient population. PMID- 15292410 TI - Functional outcomes following trauma-related lower-extremity amputation. AB - BACKGROUND: The principal aims of this study were to examine functional outcomes following trauma-related lower-extremity amputation and to compare outcomes according to the amputation levels. We hypothesized that above-the-knee amputations would result in less favorable outcomes than would through-the-knee or below-the-knee amputations. A secondary aim was to examine the factors, in addition to amputation level, that influence outcome, including the type of soft tissue coverage, selected patient characteristics, and the technological sophistication of the prosthetic device. METHODS: A cohort of 161 patients who had undergone an above-the-ankle amputation at a trauma center within three months following the injury was followed prospectively at three, six, twelve, and twenty-four months after the injury. The Sickness Impact Profile, a self-reported measure of functional status, was used as the principal measure of outcome. Secondary outcomes included pain; degree of independence in transfers, walking, and climbing stairs; self-selected walking speed; and the physician's satisfaction with the clinical, functional, and cosmetic recovery of the limb. Longitudinal multivariate regression techniques were used to determine whether outcomes differed according to the level of amputation after we controlled for covariates. RESULTS: There was no significant difference in the scores on the Sickness Impact Profile between the patients treated with above-the-knee and those treated with below-the-knee amputation. However, patients with a below-the knee amputation performed better than did patients with an above-the-knee amputation on the timed test for walking speed (p = 0.04). Patients with a through-the-knee amputation had worse regression-adjusted Sickness Impact Profile scores (p = 0.05) and slower self-selected walking speeds (p = 0.004) than did patients with either a below-the-knee or an above-the-knee amputation. Differences according to the level of amputation were most pronounced for physical function. In general, physicians were less satisfied with the clinical, cosmetic, and functional recovery of the patients with a through-the-knee amputation. Except for problems encountered with insufficient gastrocnemius coverage of the stump in many patients with a through-the-knee amputation, neither the soft-tissue coverage nor the technological sophistication of the prosthesis correlated with outcome. CONCLUSIONS: Severe disability accompanies above-the-ankle lower-extremity amputation following trauma, regardless of the level of amputation. Clinicians should critically evaluate the need for a through the-knee amputation in patients with a traumatic injury. The results of this study also underscore the need for controlled studies that examine the relationship between the type and fit of prosthetic devices and functional outcomes. PMID- 15292411 TI - Combined dorsal and volar plate fixation of complex fractures of the distal part of the radius. AB - BACKGROUND: Fractures of the distal part of the radius that are associated with complex comminution of both the articular surface and the metaphysis (subgroup C3.2 according to the Comprehensive Classification of Fractures) are a challenge for surgeons using standard operative techniques. METHODS: Twenty-five patients with subgroup-C3.2 fractures that had been treated with combined dorsal and volar plate fixation were evaluated at an average of twenty-six months after the injury. Subsequent procedures included implant removal in twenty-one patients and reconstruction of a ruptured tendon in two patients. RESULTS: An average of 54 degrees of extension, 51 degrees of flexion, 79 degrees of pronation, and 74 degrees of supination were achieved. The grip strength in the involved limb was an average of 78% of that in the contralateral limb. The average radiographic measurements were 2 degrees of dorsal angulation, 21 degrees of ulnar inclination, 0.8 mm of positive ulnar variance, and 0.7 mm of articular incongruity. Seven patients had radiographic signs of arthrosis during the follow up period. A good or excellent functional result was achieved for twenty-four patients (96%) according to the rating system of Gartland and Werley and for ten patients (40%) according to the more stringent modified system of Green and O'Brien. CONCLUSIONS: Combined dorsal and volar plate fixation of the distal part of the radius can achieve a stable, mobile wrist in patients with very complex fractures. The results are limited by the severity of the injury and may deteriorate with longer follow-up. A second operation for implant removal is common, and there is a small risk of tendon-related complications. PMID- 15292412 TI - Development and evaluation of an integrated musculoskeletal disease course for medical students. AB - BACKGROUND: A recent survey of medical and surgical residents in the United States suggested that our current training of physicians may be inadequate to meet the increasing demand for diagnosis and treatment of musculoskeletal disorders. In response, we developed an integrated, multidisciplinary course to teach knowledge and skills related to musculoskeletal disease to second-year medical students. A three-year prospective outcomes study was conducted to evaluate the new course. METHODS: The primary outcomes that were studied during the first year of the new course were the gains in knowledge, changes in levels of confidence, and long-term retention of skills. Secondary outcomes consisted of student and faculty satisfaction. Ten-item pre-tests and post-tests covering core course concepts were administered to students. A matched-pairs t test was used to evaluate the difference between pre-test and post-test scores. Students were also asked to rate, on a 10-point scale, how much confidence they had in their ability to perform the musculoskeletal physical examination before and after the institution of the new curriculum. A general linear model analysis with post hoc pairwise comparisons (F test) was used to evaluate the changes in the confidence levels of the students. Also, a knee examination station was organized to compare students' scores before and after revision of the course. At the conclusion of the course, students rated each aspect of it on a scale of 1 to 5. Instructors were asked to rate the effectiveness of all elements of the course on the same scale. RESULTS: On the basis of student satisfaction and confidence and faculty satisfaction, the most effective changes in the curriculum were the introduction of a physical examination workshop and simulated clinical situations. Students' knowledge increased significantly (p < 0.001), and their level of confidence increased significantly in thirteen specifically targeted areas (p < 0.0001). On the end-of-the-year examination assessing retention of physical examination skills, the scores for the skills emphasized in the revised course increased significantly whereas the scores for a skill not emphasized in the course remained the same. Revisions made in the second and third years after implementation of the course expanded the more successful elements and further improved student ratings. CONCLUSIONS: Integration of the three clinical disciplines related to musculoskeletal disease--orthopaedics, rheumatology, and physical medicine and rehabilitation--resulted in a highly effective introductory course for second-year medical students. The heuristic strategy of introducing core content through lectures and workshops followed by small-group teaching sessions for practice with the new knowledge effectively increased students' knowledge, confidence, and satisfaction. PMID- 15292414 TI - Initial tension and anterior load-displacement behavior of high-stiffness anterior cruciate ligament graft constructs. AB - BACKGROUND: Because the tension that exists in an anterior cruciate ligament graft when the knee is unloaded (the initial tension) affects the surgical outcome and because high initial tension has a number of adverse consequences, the primary purpose of this study was to determine quantitatively how much less initial tension was required for a high-stiffness construct than for a low stiffness construct. A secondary purpose was to determine how the stiffness of the graft construct affects the anterior load-displacement behavior of the knee from 0 degrees to 90 degrees of flexion. METHODS: Anterior-posterior load displacement was measured in each of ten intact cadaveric knee specimens, the anterior cruciate ligament was excised, and the anterior cruciate ligament was reconstructed with a double-loop bovine tendon graft. Graft constructs of different stiffness were created with use of six springs, ranging in stiffness from 25 to 275 N/mm to simulate the fixation stiffness. After adjusting the initial tension of the graft so that the anterior-posterior laxity of the reconstructed knee matched that of the intact knee, the 0-N posterior limit and the 225-N anterior limit were measured at 0 degrees, 30 degrees, 60 degrees, and 90 degrees of flexion. RESULTS: The highest stiffness fixation (275 N/mm) required an average of 73 N of initial tension, which was more than three times less than the average of 242 N of initial tension required by the lowest stiffness fixation (25 N/mm). The 225-N anterior limit was overconstrained an average of 1.0 mm with the highest stiffness fixation (275 N/mm), which was 3.6 mm less than the overconstraint with the lowest stiffness fixation (25 N/mm). Likewise, the posterior limit was overconstrained an average of 2.6 mm with the highest stiffness fixation (275 N/mm), which was 3.8 mm less than the overconstraint with the lowest stiffness fixation (25 N/mm). CONCLUSIONS: The initial tension for a high-stiffness graft construct is more than three times less than that for a low-stiffness construct. The initial tension for a high stiffness graft construct better restores both the 225-N anterior limit and the 0 N posterior limit to normal than the initial tension for a low-stiffness graft construct over the range of flexion from 0 degrees to 90 degrees. PMID- 15292413 TI - The effect of opening wedge thoracostomy on thoracic insufficiency syndrome associated with fused ribs and congenital scoliosis. AB - BACKGROUND: Thoracic insufficiency syndrome is the inability of the thorax to support normal respiration or lung growth and is seen in patients who have severe congenital scoliosis with fused ribs. Traditional spinal surgery does not directly address this syndrome. METHODS: Twenty-seven patients with congenital scoliosis associated with fused ribs of the concave hemithorax had an opening wedge thoracostomy with primary longitudinal lengthening with use of a chest-wall distractor known as a vertical, expandable prosthetic titanium rib. Repeat lengthenings of the prosthesis were performed at intervals of four to six months. Radiographs were analyzed with respect to correction of the spinal deformity, as indicated by a change in the Cobb angle, and lateral deviation of the spine, as indicated by the interpedicular line ratio. Spinal growth was assessed by measuring the change in the length of the spine. Correction of the thoracic deformity and thoracic growth were assessed on the basis of the increase in the height of the concave hemithorax compared with the height of the convex hemithorax (the space available for the lung), the increase in the thoracic spinal height, and the increase in the thoracic depth and width. The thoracic deformity in the transverse plane was measured with computed tomography, and the scans were analyzed for spinal rotation, thoracic rotation, and the posterior hemithoracic symmetry ratio. Clinically, the patients were assessed on the basis of the relative heights of the shoulders and of head and thorax compensation. Pulmonary status was evaluated on the basis of the respiratory rate, capillary blood gas levels, and pulmonary function studies. RESULTS: The mean age at the time of the surgery was 3.2 years (range, 0.6 to 12.5 years), and the mean duration of follow-up was 5.7 years. All patients had progressive congenital scoliosis, with a mean increase of 15 degrees /yr before the operation. The scoliosis decreased from a mean of 74 degrees preoperatively to a mean of 49 degrees at the time of the last follow-up. Both the mean interpedicular line ratio and the space available for the lung ratio improved significantly. The height of the thoracic spine increased by a mean of 0.71 cm/yr. At the time of the last follow-up, the mean percentage of the predicted normal vital capacity was 58% for patients younger than two years of age at the time of the surgery, 44% for those older than two years of age (p < 0.001), and 36% for those older than two years of age who had had prior spine surgery. In a group of patients who had sequential testing, all increases in the volume of vital capacity were significant (p < 0.0001), but the changes in the percentages of the predicted normal vital capacity were not. There was a total of fifty-two complications in twenty-two patients, with the most common being asymptomatic proximal migration of the device through the ribs in seven patients. CONCLUSIONS: Opening wedge thoracostomy with use of a chest-wall distractor directly treats segmental hypoplasia of the hemithorax resulting from fused ribs associated with congenital scoliosis. The operation addresses thoracic insufficiency syndrome by lengthening and expanding the constricted hemithorax and allowing growth of the thoracic spine and the rib cage. The procedure corrects most components of chest-wall deformity and indirectly corrects congenital scoliosis, without the need for spine fusion. The technique requires special training and should be performed by a multispecialty team. PMID- 15292415 TI - Patient survival after hip arthroplasty for metastatic disease of the hip. AB - BACKGROUND: The hip joint is a common location for metastatic disease. Actual as well as impending fractures at this site are frequently due to mechanical instability after tumor invasion and are usually treated surgically with hip arthroplasty. The objective of this study was to analyze survival and influences on survival after hip arthroplasty for metastatic hip disease. METHODS: Two hundred and ninety-nine patients who had undergone a total of 306 hemiarthroplasty or total hip arthroplasty procedures for treatment of a pathologic or an impending pathologic hip fracture between 1969 and 1996 at our institution were included in this study. Data that had been acquired prospectively within the total joint registry of our institution were reviewed retrospectively. RESULTS: The median duration of survival after the arthroplasty was 8.6 months. The duration of survival was significantly associated with the site of the fracture, location of the primary tumor, and time from the diagnosis of the primary tumor to the surgery for the fracture (p < or = 0.05). The time from the diagnosis to the arthroplasty was a significant independent predictor of survival. CONCLUSIONS: Patients undergoing hip arthroplasty for metastatic disease have a limited life expectancy, with only 40% (120) of the 299 patients in our series still alive at one year after the surgery. By identifying prognostic factors regarding life expectancy, this study provides surgeons and oncologists with information with which to weigh risks and benefits of hip arthroplasty for individual patients preoperatively. PMID- 15292417 TI - Clinical and radiographic results of expansive lumbar laminoplasty in patients with spinal stenosis. AB - BACKGROUND: In 1981, we developed a technique of expansive lumbar laminoplasty to alleviate the problems of conventional laminectomy in the treatment of spinal stenosis. The purposes of this study were to assess the long-term outcome following expansive lumbar laminoplasty and to investigate the postoperative problems. METHODS: Fifty-four patients underwent expansive lumbar laminoplasty for the treatment of spinal stenosis. There were forty-three men and eleven women with a mean age of 52.6 years. The average length of follow-up was 5.5 years. Preoperatively, twenty-five patients had degenerative stenosis; thirteen, stenosis due to spondylolisthesis; twelve, combined stenosis (disc herniation and stenosis); and six, hyperostotic stenosis. (Two patients with hyperostotic stenosis and spondylolisthesis were included in both groups.) The clinical results were assessed with use of the Japanese Orthopaedic Association score, and the rate of recovery was calculated. Radiographic findings were analyzed on the basis of the cross-sectional area of the spinal canal, kyphosis, range of motion of the lumbar spine, and the rate of interlaminar fusion. RESULTS: The average recovery rate at the time of the last follow-up was 69.2% for patients with degenerative stenosis, 66.5% for patients with combined stenosis, 65.2% for those with hyperostotic stenosis, and 54.7% for those with spondylolisthesis. The factors resulting in a poor recovery were an older age and insufficient decompression of the lateral stenosis. During the follow-up period, the Japanese Orthopaedic Association score became worse for seven patients, six patients had lesions develop at the level adjacent to the laminoplasty, and five patients had spondylolisthesis develop. Interlaminar fusion was observed in twenty-two patients (41%). CONCLUSIONS: The satisfactory results of expansive lumbar laminoplasty were maintained at an average of 5.5 years after surgery. The best indications for the lumbar laminoplasty procedure were young and active patients with central spinal stenosis. PMID- 15292416 TI - Isolated acetabular revision with use of the Harris-Galante Cementless Component. Study with intermediate-term follow-up. AB - BACKGROUND: Isolated revision of an acetabular total hip component is associated with special problems related to the retention of the femoral component. We reviewed the intermediate-term results of a series of such operations with use of the Harris-Galante Porous acetabular component. METHODS: We retrospectively studied the results of ninety-five isolated acetabular revisions, specifically focusing on sixty-three that had been followed for a minimum of sixty months (average, 130 months). Evaluation measures included the Harris hip score, radiographic analysis, complications, and prosthetic survival. Follow-up information was obtained with self-administered questionnaires, telephone contact, and/or clinical examination. The effects of a femoral component with a modular neck-head junction and of trochanteric osteotomy on the dislocation rate were evaluated. RESULTS: Nine shells were rerevised: four because of recurrent dislocation, four because of aseptic loosening, and one because of dissociation of the liner. The survival rate with rerevision of the shell as the end point was 90.5% at 120 months. Pelvic osteolysis occurred in 4% of the cases. The dislocation rate for the sixty-three hips was 8%. When femoral component modularity was accounted for, the analysis of the dislocations revealed a significantly higher dislocation rate for the hips without a trochanteric osteotomy (p = 0.04). Eight arthroplasties were complicated by nerve palsies, seven of which resolved fully or nearly so. CONCLUSIONS: Isolated acetabular revision with use of the Harris-Galante Porous acetabular component was associated with a low rate of loosening, lysis, and rerevision of the shell at the time of intermediate-term follow-up. However, there was a high rate of complications, including trochanteric nonunion, dislocation, and nerve palsy. The performance of a trochanteric osteotomy was associated with a decreased rate of dislocation. PMID- 15292418 TI - Intraspinal anomalies associated with isolated congenital hemivertebra: the role of routine magnetic resonance imaging. AB - BACKGROUND: Previous studies have demonstrated high rates of intraspinal anomalies in patients with congenital scoliosis; however, various authors have not considered the presence of an isolated hemivertebra to be sufficient reason for further evaluation with magnetic resonance imaging. Consequently, the rate of magnetic resonance imaging-detected intraspinal anomalies and subsequent neurosurgical intervention in patients with a single hemivertebra is unknown. Therefore, we studied all patients with a hemivertebra, after eliminating patients with a myelomeningocele, to compare those who had a single hemivertebra with those who had a complex hemivertebral pattern. METHODS: A retrospective review of the history, physical examination findings, and magnetic resonance imaging findings for patients who had presented with at least one hemivertebra, excluding those who had a myelomeningocele, was conducted to identify the prevalence of intraspinal anomalies as seen on magnetic resonance imaging and the rate of subsequent neurosurgical intervention. Additionally, the diagnostic value of the history and the physical examination in predicting the presence of intraspinal anomalies was determined. RESULTS: One hundred and sixteen patients with congenital scoliosis and a curve that included at least one hemivertebra were identified. Seventy-six of these patients had had magnetic resonance imaging and were included in the present study. The mean age of these patients at the time of presentation was 4.9 years, and the mean duration of follow-up was 7.7 years. Twenty-nine patients had an isolated hemivertebra, and forty-seven patients had a complex hemivertebral pattern. Eight (28%) of the twenty-nine patients with an isolated hemivertebra and ten (21%) of the forty-seven patients with a complex hemivertebral pattern had an intraspinal anomaly that was detected with magnetic resonance imaging. Overall, an abnormal finding on the history or physical examination demonstrated an accuracy of 71%, a sensitivity of 56%, a specificity of 76%, a positive predictive value of 42%, and a negative predictive value of 85% for the diagnosis of an intraspinal anomaly. Three patients with an isolated hemivertebra and five patients with a complex hemivertebral pattern underwent neurosurgical intervention. All eight patients who underwent neurosurgical intervention had had detection of an intraspinal anomaly with magnetic resonance imaging, whereas only four of these patients (two of whom had an isolated hemivertebra and two of whom had a complex hemivertebral pattern) had had an abnormal finding on either the history or the physical examination. CONCLUSIONS: Patients who have an isolated hemivertebra and those who have a complex hemivertebral pattern have similar rates of intraspinal anomalies that are detected with magnetic resonance imaging and similar rates of subsequent neurosurgical intervention. The history and physical examination findings are not predictive of intraspinal anomalies. Therefore, a magnetic resonance imaging evaluation of the entire spine should be considered for all patients with congenital scoliosis, including those with an isolated hemivertebra. PMID- 15292419 TI - Operative treatment of femoral neck fractures in patients between the ages of fifteen and fifty years. AB - BACKGROUND: There is a paucity of data on the treatment of femoral neck fractures in young patients. The purpose of the present study was to review the results and complications associated with the treatment of femoral neck fractures with internal fixation in a large consecutive series of young patients. METHODS: Between 1975 and 2000, eighty-three femoral neck fractures in eighty-two consecutive patients who were between fifteen and fifty years old were treated with internal fixation at our institution. Two patients died, and eight were lost to follow-up. Seventy-three fractures were followed until union, until conversion to hip arthroplasty, or for a minimum of two years; the mean duration of follow up was 6.6 years. Fifty-one of the seventy-three fractures were displaced, and twenty-two were nondisplaced. The results and complications of treatment were retrospectively reviewed, and the effects of fracture displacement, reduction quality, and capsular decompression on outcome were evaluated. Function was assessed by evaluating pain, walking capacity, and the need for gait aids. The mean duration of follow-up for the fifty-seven patients (fifty-eight fractures) who had not undergone early conversion to arthroplasty was 8.1 years. RESULTS: Fifty-three (73%) of the seventy-three fractures healed after one operation and were associated with no evidence of osteonecrosis of the femoral head. Osteonecrosis developed in association with seventeen fractures (23%), and a nonunion developed in association with six (8%). Four of the six nonunions later healed after a secondary procedure. At the time of the final follow-up, thirteen patients had had a conversion to a total hip arthroplasty because of osteonecrosis (eleven), nonunion (one), or both (one). Five (9.8%) of the fifty one displaced fractures were associated with the development of nonunion, and fourteen (27%) were associated with the development of osteonecrosis. Three (14%) of the twenty-two nondisplaced fractures were associated with the development of osteonecrosis, and one (4.5%) was associated with the development of nonunion. Eleven (24%) of the forty-six displaced fractures with a good to excellent reduction were associated with the development of osteonecrosis, and two (4%) were associated with the development of nonunion. Four of the five displaced fractures with a fair or poor reduction were associated with the development of osteonecrosis, nonunion, or both. CONCLUSIONS: The ten-year survival rate of the native femoral head free of conversion to total hip arthroplasty was 85%. Osteonecrosis was the main reason for conversion to total hip arthroplasty, but not all patients with osteonecrosis required further surgery. The results of treatment were influenced by fracture displacement and the quality of reduction. PMID- 15292420 TI - Interobserver agreement in the application of levels of evidence to scientific papers in the American volume of the Journal of Bone and Joint Surgery. AB - BACKGROUND: Since January 2003, all clinical scientific articles published in the American volume of The Journal of Bone and Joint Surgery (JBJS-A) have included a level-of-evidence rating. The aim of the current study was to evaluate the interobserver agreement among reviewers, with varying levels of epidemiology training, in categorizing the levels of evidence of these clinical studies. METHODS: Fifty-one consecutive clinical papers published in the American volume of JBJS were identified by a computerized search of the table of contents from January 2003 through June 2003. Each paper was blinded so that only the title, abstract (without the level of evidence designated), and methods section were provided to the reviewers. The papers were coded and were randomly organized in a binder. Six surgeons graded each blinded paper for (1) the type of study (therapeutic, prognostic, diagnostic test, or economic or decision analysis), (2) the level of evidence (on a scale of I through V), and (3) the subcategory within the particular level of evidence. Three surgeons were members of JBJS American Editorial Board, two surgeons were reviewers for JBJS-A, and one surgeon was an active researcher not formally associated with JBJS-A. The reviewers did not receive any formal training in the application of the classification system, but each was provided with a detailed description of the classification system used by JBJS-A. Intraclass correlation coefficients with 95% confidence intervals were determined for the reviewers' agreement regarding the type of study, level of evidence, and subcategory within the level of evidence. RESULTS: The majority (69%) of the fifty-one included articles were studies of therapy, and 57% of the studies constituted Level-IV evidence. The intraclass correlation coefficients for the agreement among all reviewers with regard to the study type, level of evidence, and subcategory within the level of evidence ranged from 0.61 to 0.75. Reviewers trained in epidemiology demonstrated greater agreement (range in intraclass correlation coefficients, 0.99 to 1.0), across all aspects of the classification system, than did reviewers who were not trained in epidemiology (range in intraclass correlation coefficients, 0.60 to 0.75). CONCLUSIONS: These findings suggest that epidemiology and non-epidemiology-trained reviewers can apply the levels-of-evidence guide to published studies with acceptable interobserver agreement. The validity of this system remains a question for future research. PMID- 15292421 TI - Knee kinematics with a high-flexion posterior stabilized total knee prosthesis: an in vitro robotic experimental investigation. AB - BACKGROUND: An analysis of contemporary total knee arthroplasty reveals that, on the average, patients rarely flex the knee beyond 120 degrees. The biomechanical mechanisms that inhibit further flexion after total knee arthroplasty are unknown. The objective of the present study was to investigate the capability of a single design of a fixed-bearing, high-flexion posterior stabilized total knee arthroplasty system (LPS-Flex) to restore the range of flexion to that of the intact knee. METHODS: Thirteen cadaveric human knees were tested, with use of a robotic testing system, before and after total knee arthroplasty with the LPS Flex prosthesis. The passive path and the kinematics under an isolated quadriceps force of 400 N, under an isolated hamstring force of 200 N, and with these forces combined were determined. Posterior femoral translation of the lateral and medial femoral condyles and tibial rotation were recorded from 0 degrees to 150 degrees of flexion. RESULTS: The medial and lateral condyles of the intact knee translated posteriorly from full extension to 150 degrees, reaching a mean peak (and standard deviation) of 22.9 +/- 11.3 mm and 31.9 +/- 12.5 mm, respectively, under the combined muscle forces. Following total knee arthroplasty, the amount of posterior femoral translation was lower than that observed in the intact knee. At 150 degrees, approximately 90% of the intact posterior femoral translation was recovered by the total knee replacement. Internal tibial rotation was observed for all knees throughout the range of motion. The cam-spine mechanism engaged at approximately 80 degrees and disengaged at 135 degrees. Despite the absence of cam-spine engagement, further posterior femoral translation occurred from 135 degrees to 150 degrees. CONCLUSIONS: The tibiofemoral articular geometry of the intact knee and the knee after total knee arthroplasty with use of the LPS-Flex design demonstrated similar kinematics at high flexion angles. The cam-spine mechanism enhanced posterior femoral translation only at the mid-range of flexion. The femoral component geometry of the LPS-Flex total knee prosthesis may improve posterior tibiofemoral articulation contact in high flexion angles. PMID- 15292422 TI - The effect of radial head excision and arthroplasty on elbow kinematics and stability. AB - BACKGROUND: Radial head fractures are common injuries. Comminuted radial head fractures often are treated with radial head excision with or without radial head arthroplasty. The purpose of the present study was to determine the effect of radial head excision and arthroplasty on the kinematics and stability of elbows with intact and disrupted ligaments. We hypothesized that elbow kinematics and stability would be (1) altered after radial head excision in elbows with intact and disrupted ligaments, (2) restored after radial head arthroplasty in elbows with intact ligaments, and (3) partially restored after radial head arthroplasty in elbows with disrupted ligaments. METHODS: Eight cadaveric upper extremities were studied in an in vitro elbow simulator that employed computer-controlled actuators to govern tendon-loading. Testing was performed in stable, medial collateral ligament-deficient, and lateral collateral ligament-deficient elbows with the radial head intact, with the radial head excised, and after radial head arthroplasty. Valgus angulation and rotational kinematics were determined during passive and simulated active motion with the arm dependent. Maximum varus-valgus laxity was measured with the arm in a gravity-loaded position. RESULTS: In specimens with intact ligaments, elbow kinematics were altered and varus-valgus laxity was increased after radial head excision and both were corrected after radial head arthroplasty. In specimens with disrupted ligaments, elbow kinematics were altered after radial head excision and were similar to those observed in specimens with a native radial head after radial head arthroplasty. Varus-valgus laxity was increased after ligament disruption and was further increased after radial head excision. Varus-valgus laxity was corrected after radial head arthroplasty and ligament repair; however, it was not corrected after radial head arthroplasty without ligament repair. CONCLUSIONS: Radial head excision causes altered elbow kinematics and increased laxity. The kinematics and laxity of stable elbows after radial head arthroplasty are similar to those of elbows with a native radial head. However, radial head arthroplasty alone may be insufficient for the treatment of complex fractures that are associated with damage to the collateral ligaments as arthroplasty alone does not restore stability to elbows with ligament injuries. PMID- 15292423 TI - Langerhans cell histiocytosis of the spine in children. Long-term follow-up. AB - BACKGROUND: Langerhans cell histiocytosis causes destructive lesions in a child's spine. Few large, long-term studies have evaluated the clinical and radiographic presentation, natural history, outcomes of modern treatment approaches, and maintenance of normal spinal growth and stability after the diagnosis of this disease in children. METHODS: Twenty-six children with biopsy-proven Langerhans cell histiocytosis involving the spine were treated at our institution between 1970 and 2003. They had a total of forty-four involved vertebrae (twenty cervical, fourteen thoracic, and ten lumbar). Vertebral body collapse was measured on radiographs and classified as grade I (0% to 50% collapse) or grade II (51% to 100% collapse) and subclassified as A (symmetric collapse) or B (asymmetric collapse). Lesions of the posterior elements of the spine were classified as grade III. Twenty-three children were followed for two years or more (mean, 9.4 years), and the analyses of treatment and long-term outcomes were performed in that group of patients. RESULTS: There was a predominance of lesions in the cervical spine (p pirenzepine > 11-[[2 [(diethylamino)methyl]-1-piperidinyl]acetyl]-5,11-dihydro-6H-pyrido[2,3 b][1,4]benzodiazepin-6-one, a pharmacological profile identical to that obtained for M3 mAChR-stimulated phosphoinositide hydrolysis. Agonist-stimulated efflux of both osmolytes could be inhibited by inclusion of either anion channel blockers known to inhibit the volume-sensitive organic anion channel (VSOAC) or by a tyrosine kinase inhibitor alpha-cyano-(3,4-dihydroxy)cinnamonitrile. The results indicate that the activation of M3 mAChRs on SH-SY5Y neuroblastoma facilitates the ability of these cells to respond to very limited reductions in osmolarity via a release of osmolytes. mAChR-stimulated osmolyte efflux is mediated via a VSOAC and seems to require the activity of a tyrosine kinase. PMID- 15292462 TI - N-glucuronidation of carbamazepine in human tissues is mediated by UGT2B7. AB - Carbamazepine (CBZ) is one of the most widely prescribed anticonvulsants despite a high incidence of idiosyncratic side effects. Metabolism of CBZ is complex, and of the more than 30 metabolites identified, one of the most abundant is CBZ N glucuronide. To date the uridine diphosphate glucuronosyltransferase (UGT) isoform responsible for the N-glucuronidation of CBZ has not been identified. We have developed a sensitive liquid chromatography/mass spectrometry assay to quantify CBZ glucuronidation, and we report that CBZ is specifically glucuronidated by human UGT2B7. Kinetics of CBZ glucuronidation in human liver, kidney, and intestine microsomes were consistent with those of recombinant UGT2B7, which displayed a Km value of 214 microM and Vmax value of 0.79 pmol/mg/min. In addition to revealing the isoform responsible for CBZ glucuronidation, this is the first example of primary amine glucuronidation by UGT2B7. PMID- 15292463 TI - Oxidative stress induced by iron released from transferrin in low pH peritoneal dialysis solution. AB - BACKGROUND: Transferrin binds extracellular iron and protects tissues from iron induced oxidative stress. The binding of iron and transferrin is pH dependent and conventional peritoneal dialysis (PD) solutions have unphysiologically low pH values. Herein, we investigated whether conventional PD solution releases iron from transferrin and if the released iron causes oxidative stress. METHODS: Effects of PD solutions on iron binding to transferrin were examined with purified human transferrin and transferrin in dialysates drained from PD patients. Oxidative stress induced by iron released from transferrin was evaluated in terms of the formation of thiobarbituric acid reactive substance (TBARS) and protein carbonylation in the human red blood cell (RBC) membrane. The iron deposition in peritoneal tissue from PD patients was evaluated by Perls' staining with diaminobenzidine intensification. RESULTS: Low pH PD solution released iron from transferrin. This iron release occurred within 1 min. Iron release was not observed in neutralized PD solution. Iron released from transferrin in low pH PD solution increased TBARS formation and protein carbonylation in the human RBC membrane. Iron deposition, which is prominent in the fibrotic area facing the peritoneal cavity, was observed in the peritoneum of PD patients. CONCLUSIONS: Iron released from transferrin in low pH PD solution can produce oxidative stress in the peritoneum of a PD patient. Neutralizing PD solution can avoid this problem. Iron deposition in the peritoneum may participate in the pathogenesis of peritoneal fibrosis in PD patients. PMID- 15292464 TI - Monitoring the progress of BK virus associated nephropathy in renal transplant recipients. AB - BACKGROUND: Nephropathy associated with BK virus (BKVAN) has recently emerged as an important cause of allograft failure following renal transplantation. The aim of this study was to evaluate the effectiveness of laboratory markers in the follow-up of patients with BKVAN. METHODS: Serial samples from seven renal transplant recipients with biopsy proven BKVAN were studied. The median follow-up time from diagnosis was 76 weeks. Intervention after the diagnosis of BKVAN included immunosuppression dose reduction, alternative immunosuppressive agents and/or antiviral therapy with cidofovir. Serial urine samples (n = 127) were collected for electron microscopy (EM), decoy cell detection and quantitative urine BK viral load using real-time polymerase chain reaction. Serum BK viral load was also measured serially (n = 72). RESULTS: All patients showed a reduction in serum and urine viral load during the period of follow-up co incident with the loss of decoy cells and negative urine EM. Urine samples that were negative for decoy cells or polyomavirus by EM had a urine viral load <10(6) copies/ml and a corresponding serum viral load <10(3) copies/ml. In paired serum/urine samples, there was a proportional relationship between serum and urine viral load with each urine viral load approximately 1000-fold higher than the corresponding serum level. Serum and urine viral loads that decreased to <200 and < 10(6) copies/ml, respectively, correlated with histological improvement. CONCLUSION: Negative EM and absence of decoy cells could be used as broad indicators of a response to intervention. However, measurement of BK virus DNA level provided a wider dynamic range and could be a better choice for determining the extent of viral control. PMID- 15292465 TI - Different impact of normo- and hypotensive brain death on renal macro- and microperfusion--an experimental evaluation in a porcine model. AB - BACKGROUND: Despite the growing use of kidneys from living donors, organs harvested from brain dead donors are the dominant graft types used in renal transplantation. It is accepted that brain death (BD) has a damaging effect on the renal allograft, with a lower graft survival. Amongst various causes, changes in renal microperfusion could be responsible. Renocortical microperfusion was assessed during BD using thermal diffusion in a porcine model. METHODS: Two types of BD were induced in two groups of pigs [hypotension (Hypo-BD): n = 11; normotension (Normo-BD): n = 10] and compared to controls (n = 5) over a period of 210 min. We analysed systemic parameters [heart rate (HR), mean arterial blood pressure (MAP)], aortic blood flow (ABF) and renal perfusion [renal artery blood flow (RABF) and renocortical blood flow (RCBF)]. RESULTS: Following the two distinct forms of BD induction, a stable normo- or hypotension was observed. Haemodynamic parameters were only slightly changed (control group: MAP, 62+/-2 mmHg; HR, 95+/-3/min; Normo-BD: MAP, 56+/-4 mmHg; HR, 104+/-8/min; Hypo-BD: MAP, 43+/-3 mmHg; HR, 112+/-7/min). Solely dependent on systemic haemodynamics, RABF and RCBF decreased in the Hypo-BD (RABF: 142+/-19 to 94+/-9 ml/100 g/min; RCBF: 80+/-4 to 52+/-2 ml/100 g/min), while in Normo-BD group RABF mildly changed (158+/-13 ml/100 g/min) and RCBF decreased slightly from 76+/-3 to 70+/-6 ml/100 g/min. As opposed to the Normo-BD group, animals with Hypo-BD showed a significant decrease in RABF (reduction of 34%) and RCBF (reduction of 35%) with a sharp drop of MAP (reduction of 25%), however ABF remained relatively constant. CONCLUSIONS: In this model, a reduction of renocortical microperfusion in brain dead pigs was only found during haemodynamic instability (hypotension) and could not be attributed to BD as such. Our findings would support intensive cardiocirculatory stabilization for potential BD donors in order to minimize kidney preservation damage. PMID- 15292466 TI - The IL-6 gene G-174C polymorphism related to health indices in Greek primary school children. AB - Interleukin-6 (IL-6) is a pleiotropic cytokine expressed in many tissues. A polymorphism in the IL-6 gene, associated with differences in IL-6 transcription rate, has been recently described. Subjects with the -174GG genotype are prone to lipid abnormalities. We investigated the effect of the G-174C IL-6 polymorphism on health indices and lipid values of 184 Greek primary school children. The genotype distribution of the polymorphism was 37.5% for GG and 52.2% and 10.3% for GC and CC, respectively. No differences were observed between genotype distribution and gender (p = 0.37). Boys homozygous for the G allele showed higher triglyceride levels than boys carrying the C allele (86 +/- 28 vs. 74 +/- 20 mg/dL, p = 0.02) and lower mid-upper arm muscle circumference (17.46 +/- 1.86 vs. 18.91 +/- 2.53 cm, p = 0.013). In addition, girls homozygous for the G allele had higher values for suprailiac skinfolds compared with those bearing the C allele (21.28 +/- 12.56 vs. 17.09 +/- 13.36 mm, p = 0.06). These findings were confirmed by multiple linear regression analysis, after controlling for age, sex, BMI, energy and total fat intake, and weekly physical activity. From the results of the present study, we concluded that individuals homozygous for G allele on the IL-6 gene have higher values in some parameters associated with obesity. PMID- 15292467 TI - Body fat and fat-free mass and all-cause mortality. AB - OBJECTIVE: To investigate whether the association between BMI and all-cause mortality could be disentangled into opposite effects of body fat and fat-free mass (FFM). RESEARCH METHODS AND PROCEDURES: All-cause mortality was studied in the Danish follow-up study "Diet, Cancer and Health" with 27,178 men and 29,875 women 50 to 64 years old recruited from 1993 to 1997. By the end of year 2001, the median follow-up was 5.8 years, and 1851 had died. Body composition was assessed by bioelectrical impedance. Cox regression models were used to estimate the relationships among body fat mass index (body fat mass divided by height squared), FFM index (FFM divided by height squared), and mortality. All analyses were adjusted for smoking habits. RESULTS: Men and women showed similar associations. J-shaped associations were found between body fat mass index and mortality adjusted for FFM and smoking. The mortality rate ratios in the upper part of body fat mass were 1.12 per kg/m2 (95% confidence interval: 1.07, 1.18) in men and 1.06 per kg/m2 (95% confidence interval: 1.02, 1.10) in women. Reversed J-shaped associations were found between FFM index and mortality with a tendency to level off for high values of FFM. DISCUSSION: Our findings suggest that BMI represents joint but opposite associations of body fat and FFM with mortality. Both high body fat and low FFM are independent predictors of all-cause mortality. PMID- 15292468 TI - Mediators of weight loss in a family-based intervention presented over the internet. AB - OBJECTIVE: To assess the process variables involved in a weight loss program for African-American adolescent girls. Several process variables have been identified as affecting success in in vivo weight loss programs for adults and children, including program adherence, self-efficacy, and social support. The current study sought to broaden the understanding of these process variables as they pertain to an intervention program that is presented using the Internet. It was hypothesized that variables such as program adherence, dietary self-efficacy, psychological factors, and family environment factors would mediate the effect of the experimental condition on weight loss. RESEARCH METHODS AND PROCEDURES: Participants were 57 adolescent African-American girls who joined the program with one obese parent; family pairs were randomized to either a behavioral or control condition in an Internet-based weight loss program. Outcome data (weight loss) are reported for the first 6 months of the intervention. RESULTS: Results partially supported the hypotheses. For weight loss among adolescents, parent variables pertaining to life and family satisfaction were the strongest mediating variables. For parental weight loss, changes in dietary practices over the course of 6 months were the strongest mediators. DISCUSSION: The identification of factors that enhance or impede weight loss for adolescents is an important step in improving weight loss programs for this group. The current findings suggest that family/parental variables exert a strong influence on weight loss efforts for adolescents and should be considered in developing future programs. PMID- 15292469 TI - The Val103Ile polymorphism of melanocortin-4 receptor regulates energy expenditure and weight gain. AB - OBJECTIVE: The melanocortin-4 receptor (MC4R) regulates energy intake. On the basis of animal studies, it may also regulate energy expenditure. RESEARCH METHODS AND PROCEDURES: The effect of the Val103Ile polymorphism of the MC4R gene on energy metabolism was studied in 229 middle-aged nondiabetic subjects (Group 1, age 51.2 +/- 9.8 years, BMI 26.8 +/- 4.5 kg/m2) and on weight gain in 1013 elderly subjects (Group 2, age 69.9 +/- 2.9 years, BMI 27.4 +/- 4.1 kg/m2) during a 3.5-year follow-up study. In Group 1, insulin sensitivity, energy expenditure, and substrate oxidation were measured with the hyperinsulinemic euglycemic clamp combined with indirect calorimetry. RESULTS: In Group 1, the Val103Ile genotype was associated with high rates of energy expenditure (63.42 +/- 13.40 in eight subjects with the Val103Ile genotype vs. 59.86 +/- 7.33 J/kg per minute in 221 subjects with the Val103Val genotype, p = 0.007), high rates of glucose oxidation (8.90 +/- 6.15 vs. 6.07 +/- 4.38 micromol/kg per minute, p = 0.020), and low levels of free fatty acids (0.45 +/- 0.18 vs. 0.56 +/- 0.23 mM, p = 0.029) in the fasting state, and with high rates of glucose oxidation during the clamp (18.88 +/- 4.63 vs. 17.60 +/- 3.24 micromol/kg per minute, p = 0.031). In Group 2, the 103Ile allele was associated with an increase in weight gain during the follow-up (0.78 +/- 3.98 vs. -0.82 +/- 3.98 kg, p = 0.038). DISCUSSION: The Val103Ile polymorphism of the MC4R gene is associated with energy expenditure in humans. Furthermore, it may associate with glucose oxidation, free fatty acid levels, and weight gain. PMID- 15292470 TI - Effects of diet and time of the day on serum and CSF leptin levels in Osborne Mendel and S5B/Pl rats. AB - OBJECTIVE: To characterize the effects of dietary fat on the diurnal variation in serum and cerebrospinal fluid (CSF) leptin levels in Osborne-Mendel (OM) and S5B/Pl rats and quantitate the dose response to lower doses of leptin administered into the third cerebral ventricle. RESEARCH METHODS AND PROCEDURES: Rats were fitted with implanted vascular ports or third ventricular cannulas and fed either laboratory chow or one of two semipurified high-fat or low-fat diets. Leptin and insulin were measured by immunoassay. RESULTS: Serum leptin and insulin levels were positively correlated and had similar patterns of diurnal change. CSF leptin and insulin also had diurnal rhythms, with a peak at 7:00 am, but the diurnal oscillations of leptin and insulin were significantly lower in the S5B/Pl rats than the OM rats. Thus, the ratio of CSF to serum leptin was significantly higher in the S5B/Pl rats than in the OM rats. Dietary fat had no effect on these diurnal patterns. There was a right shift in the dose response to leptin in the OM rats compared with the S5B/P1 rats. S5B/P1 rats treated with leptin had higher signal transduction and translation (STAT-3) mRNA levels compared with pair-fed or saline injected S5B/P1 rats. Hypothalamic suppressors of cytokine signaling mRNA levels were not statistically different between the groups. DISCUSSION: The higher CSF-to-serum leptin ratio in the S5B/P1 rats, the enhanced suppression of food intake and body weight with leptin injections, and the higher STAT-3 activity in these animals suggest that S5B/P1 rats are more sensitive to leptin than OM rats. PMID- 15292471 TI - Taurine alters respiratory gas exchange and nutrient metabolism in type 2 diabetic rats. AB - OBJECTIVE: To assess the effect of taurine supplementation on respiratory gas exchange, which might reflect the improved metabolism of glucose and/or lipid in the type 2 diabetic Otsuka Long-Evans Tokushima Fatty (OLETF) rats. RESEARCH METHODS AND PROCEDURES: Male OLETF rats (16 weeks of age) were randomly divided into two groups: unsupplemented group and taurine-supplemented (3% in drinking water) group. After 9 weeks of treatment, indirect calorimetry and insulin tolerance tests were conducted. The amounts of visceral fat pads, tissue glycogen, the blood concentrations of glucose, triacylglycerol, taurine, and electrolytes, and the level of hematocrit were compared between groups. A nondiabetic rat strain (Long-Evans Tokushima Otsuka) was used as the age-matched normal control. RESULTS: The indirect calorimetry showed that the treatment of OLETF rats with taurine could reduce a part of postprandial glucose oxidation possibly responsible for the increase of triacylglycerol synthesis in the body. Taurine supplementation also improved hyperglycemia and insulin resistance and increased muscle glycogen content in the OLETF rats. Supplementation with taurine increased the blood concentration of taurine and electrolyte and fluid volume, all of which were considered to be related to the improvement of metabolic disturbance in OLETF rats. DISCUSSION: Taurine supplementation may be an effective treatment for glucose intolerance and fat/lipid accumulation observed in type 2 diabetes associated with obesity. These metabolic changes might be ascribed, in part, to the alteration of circulating blood profiles, where the improved hyperglycemia and/or the blood accumulation of taurine itself would play roles. PMID- 15292472 TI - Lean and weight stable: behavioral predictors and psychological correlates. AB - OBJECTIVE: To examine behavioral characteristics associated with being lean and weight stable during adulthood. RESEARCH METHODS AND PROCEDURES: Participants included 165 white married couples who were part of a larger longitudinal study. Participants' height and weight were measured on three occasions at 2-year intervals. Men and women were classified as being lean and weight stable (the target group) if they had a BMI < 25 at baseline and maintained their weight within 5% of baseline at 2nd and 4th year follow-up. Individuals not fulfilling these criteria were included in the comparison group. Group differences in background characteristics including childhood weight status, BMI at each occasion, dieting history, and mental and physical health history were examined. In addition, multiple measures of dietary intake and physical activity were obtained and used to predict the likelihood of being in the target group. RESULTS: Men (N = 22) and women (N = 36) in the target group had lower mean BMI scores at each occasion, were less overweight during childhood, were less likely to have dieted in the past year, and rated themselves as being more healthy than men and women in the comparison group. No differences were identified in mental health. Relative to the comparison group, women in the target group reported higher levels of physical activity and higher levels of physical activity among their spouses, and men in the target group reported healthier dietary patterns. DISCUSSION: Results from this study suggest that being lean and weight stable in adulthood is linked to childhood weight status in combination with dietary and activity patterns during adulthood. PMID- 15292473 TI - Development of health-related waist circumference thresholds within BMI categories. AB - OBJECTIVE: To develop and cross-validate waist circumference (WC) thresholds within BMI categories. The utility of the derived values was compared with the single WC thresholds (women, 88 cm; men, 102 cm) recommended by NIH and Health Canada. RESEARCH METHODS AND PROCEDURES: The sample included adults classified as normal weight (BMI = 18.5 to 24.9), overweight (BMI = 25 to 29.9), obese I (BMI = 30 to 34.9), and obese II+ (BMI > or = 35) from the Third U.S. National Health and Nutrition Examination Survey (NHANES III; n = 11,968) and the Canadian Heart Health Surveys (CHHS; n = 6286). Receiver operating characteristic curves were used to determine the optimal WC thresholds that predicted high risk of coronary events (top quintile of Framingham scores) within BMI categories using the NHANES III. The BMI-specific WC thresholds were cross-validated using the CHHS. RESULTS: The optimal WC thresholds increased across BMI categories from 87 to 124 cm in men and from 79 to 115 cm in women. The validation study indicated improved sensitivity and specificity with the BMI-specific WC thresholds compared with the single thresholds. DISCUSSION: Compared with the recommended WC thresholds, the BMI-specific values improved the identification of health risk. In normal weight, overweight, obese I, and obese II+ patients, WC cut-offs of 90, 100, 110, and 125 cm in men and 80, 90, 105, and 115 cm in women, respectively, can be used to identify those at increased risk. PMID- 15292474 TI - Pelvic floor dysfunction in morbidly obese women: pilot study. AB - OBJECTIVE: The objective of this study was to evaluate the impact of obesity on pelvic floor function in women. RESEARCH METHODS AND PROCEDURES: This was a prospective controlled study of 20 morbidly obese female patients planning to undergo gastric bypass surgery and 20 age-matched female controls. Subjects completed symptom and impact questionnaires, including the Incontinence Impact Questionnaire (IIQ-7), Urogenital Distress Inventory (UDI), the Kobashi Prolapse Symptom Inventory and Quality-of-Life Questionnaire (PSI-QOL), and Index of Female Sexual Function. Data were analyzed with Wilcoxon or ratio chi2 tests. RESULTS: Mean weight was 295.7 +/- 87.9 lbs in the study group and 144.79 +/- 33.07 lbs in the control group. Mean BMI was 52.65 +/-14.49 kg/m2 in the study group and 25.11 +/- 5.27 kg/m2 in the control group. According to the IIQ-7, urinary incontinence significantly affected lifestyle in the study group. The total IIQ-7 score was also significantly affected in the study group (p = 0.03). The UDI indicated more urinary leakage with activity (p = 0.04) and more incidents of small amounts of leakage (p = 0.02) in the study group. According to the PSI-QOL, women in the study group experienced constipation more often because of difficulty in emptying the rectum (p = 0.04). The PSI-QOL score was higher in the study group (6.75 +/- 6.84) than in the control group (2.65 +/- 3.03; p = 0.04). There were no significant differences between groups regarding sexual function. DISCUSSION: Morbid obesity is associated with a significant negative impact on urogenital health. Sexual function did not seem to be affected in women who are morbidly obese. PMID- 15292475 TI - Short-term effects of gastric bypass surgery on circulating ghrelin levels. AB - OBJECTIVE: To prospectively evaluate the short-term effects of Roux-en-Y gastric bypass (RYGBP) on ghrelin secretion and its relevance on food intake and body weight changes. RESEARCH METHODS AND PROCEDURES: Ghrelin response to a standardized test meal was evaluated in eight obese patients (BMI, 43.5 to 59.1 kg/m2) before and 6 weeks after RYGBP. Ghrelin response was compared with that of an age-matched group of six normal weight individuals (BMI, 19.6 to 24.9 kg/m2). RESULTS: Fasting serum ghrelin levels were lower in obese subjects compared with controls (p < 0.05). Meal ingestion significantly suppressed ghrelin concentration in controls (p < 0.05) and obese subjects (p < 0.05), albeit to a lesser degree in the latter group (p < 0.05). Despite a 10.3 +/- 1.5% weight loss, fasting serum ghrelin levels were paradoxically further decreased in obese subjects 6 weeks after RYGBP (p < 0.05). Moreover, at this time-point, food intake did not elicit a significant ghrelin suppression. The changes in ghrelin secretion after RYGBP correlated with changes in insulin sensitivity (p < 0.05) and caloric intake (p < 0.05). DISCUSSION: This study showed that the adaptive response of ghrelin to body weight loss was already impaired 6 weeks after RYGBP. Our study provides circumstantial evidence for the potential role of ghrelin in the negative energy balance in RYGBP-operated patients. PMID- 15292476 TI - Prevalence of obesity and associated risk factors in a Turkish population (trabzon city, Turkey). AB - OBJECTIVE: To estimate the prevalence of overweight and obesity (general and central) in the Trabzon Region and its associations with demographic factors (age, sex, marital status, reproductive history in women, and level of education), socioeconomic factors (household income and occupation), family history of selected medical conditions (diabetes, hypertension, and obesity), lifestyle factors (smoking habits, physical activity, and alcohol consumption), and hypertension in the adult population. RESEARCH METHODS AND PROCEDURES: A sample of households was systematically selected from the central province of Trabzon and its five towns, namely, Surmene, Vakfikebir, Macka, Hayrat, and Tonya. A total of 5016 subjects (2728 women and 2288 men) were included in the study. Individuals more than 20 years old were selected from their family health cards. Demographic factors, socioeconomic factors, family history of selected medical conditions, and lifestyle factors were obtained for all participants. Systolic blood pressure and diastolic blood pressure levels were measured for all subjects. Study procedures were carried out in the local health centers in each town over an 8-month period. Obesity was defined as BMI > or = 30 kg/m2 and overweight as BMI = 25.0 to 29.9 kg/m2. RESULTS: The prevalence of obesity was 23.5%: 29.4% in women and 16.5% in men. The combined prevalence of both overweight and obesity was 60.3%. The prevalence of abdominal obesity was 29.4%: 38.9% among women and 18.1% among men. The prevalence of obesity increased with age, being highest in the 60- to 69-year-old age group (40.8%) but lower again in the 70+ age group. Obesity was associated positively with marital status, parity, cessation of cigarette smoking, alcohol consumption, and household income and inversely with level of education, cigarette use, and physical activity. Also, obesity was associated positively with hypertension. DISCUSSION: In the Trabzon Region, 60.3% of the adult population presents with some excess weight. Obesity is a major public health problem that requires generalized interventions to prevent it among the adult population. PMID- 15292477 TI - Effects of dinner composition on postprandial macronutrient oxidation in prepubertal girls. AB - OBJECTIVE: To see whether a fat-rich (50%) evening meal promoted fat oxidation and a different spontaneous food intake on the following day at breakfast than a meal with a lower fat content (20%) in 10 prepubertal obese girls. RESEARCH METHODS AND PROCEDURES: The postabsorptive and postprandial (10.5 hours) energy expenditure after a low-fat (LF) (20% fat, 68% carbohydrate, 12% protein) and an isocaloric (2.1 MJ) and isoproteic high-fat (HF; 50% fat, 38% carbohydrate, 12% protein) meal were measured by indirect calorimetry. RESULTS: Fat oxidation was not significantly different after the two meals [LF, 31 +/- 9 vs. HF, 35 +/- 9 g/10.5 hours, p = not significant (NS)]. The girls oxidized 1.8 +/- 0.9 times more fat than that ingested (11.1 grams) with the LF meal vs. 0.3 +/- 0.3 times more fat than that ingested (27.1 grams) with the HF meal (p < 0.001). Carbohydrate oxidation was significantly higher after an LF than an HF meal (39 +/- 12 vs. 29 +/- 9 g/10.5 hours, p < 0,05). At breakfast, the girls spontaneously ingested a similar amount of energy (1.5 +/- 0.7 vs. 1.5 +/- 0.6 MJ, p = NS) and macronutrient proportions (fat, 23% vs. 26%, p = NS; protein, 9% vs. 10%; carbohydrate, 68% vs. 64%,) independently of their having eaten an HF or an LF dinner. DISCUSSION: An HF dinner did not stimulate fat oxidation, and no compensatory effect in spontaneous food intake was observed during breakfast the following morning. Cumulated total fat oxidation after dinner was higher than total fat ingested at dinner, but a much larger negative fat balance was observed after the LF meal. Spontaneous energy and nutrient intakes at breakfast were similar after LF and HF isocaloric, isoproteic dinners. This study points out the lack of sensitivity of short-term fat balance to subsequently readjust fat intake and emphasizes the importance of an LF meal to avoid transient positive fat imbalance. PMID- 15292478 TI - Race differences in accuracy of self-reported childhood body size among white and black women. AB - OBJECTIVE: To assess the relation of self-reported current and recalled preadolescent body size to measured BMI (kilograms per meter squared) and interviewer's assessment of body size. RESEARCH METHODS AND PROCEDURES: This was a prospective cohort study of 1890 white and black women who were 9 to 10 years old at time of enrollment and were followed up 10 to 13 years later. At baseline, subjects had their weight and height measured and were asked to indicate their current body size from a series of nine pictograms. A sample of the subjects also had their body size evaluated by interviewers. At the young-adult follow-up visit, subjects were asked to recall their body size at 9 and 10 years old and to indicate their current weight, height, body size, and level of concern with weight. RESULTS: Among the women with interviewer assessments, 84% of the white women and 67% of black women recalled a body size that was within one body size of the interviewer's assessment. Independent of weight status in childhood or at follow-up, black women were 3 times more likely than white women to recall a body figure that was more than one figure leaner than the shape they reported at baseline (odds ratio = 3.5, 95% confidence interval 2.8 to 4.5) or than the interviewer's rating at baseline (odds ratio = 3.4, 95% confidence interval 2.4 to 4.9). DISCUSSION: The results suggest that the use of body figures to recall childhood size are best suited for ranking subjects in terms of BMI. The higher rate of underestimation of size by black women suggests that body figure ratings work best for white women. PMID- 15292479 TI - Adult obesity and the burden of disability throughout life. AB - OBJECTIVE: To analyze the prevalence of disability throughout life and life expectancy free of disability, associated with obesity at ages 30 to 49 years. RESEARCH METHODS AND PROCEDURES: We used 46 and 20 years of mortality follow-up, respectively, for 3521 Original and 3013 Offspring Framingham Heart Study participants 30 to 49 years and classified as normal weight, overweight, or obese at baseline. Disability measures were available between 36 and 46 years of follow up for 1352 Original participants and at 20 years of follow-up for 2268 Offspring participants. We measured the odds of disability in the Original cohort after 46 years follow-up, and we estimated life expectancy with and without disability from age 50. Two disability measures were used, one representing limitations with mobility only and the second representing limitations with activities of daily living (ADL). RESULTS: Obesity at ages 30 to 49 years was associated with a 2.01 fold increase in the odds of ADL limitations 46 years later. Nonsmoking adults who were obese between 30 and 49 years lived 5.70 (95% confidence interval, 4.11 to 7.35) (men) and 5.02 (95% confidence interval, 3.36 to 6.61) (women) fewer years free of ADL limitations from age 50 than their normal-weight counterparts. There was no significant difference in the total number of years lived with disability throughout life between those obese or normal weight, due to both higher disability prevalence and higher mortality in the obese population. DISCUSSION: Obesity in adulthood is associated with an increased risk of disability throughout life and a reduction in the length of time spent free of disability, but no substantial change in the length of time spent with disability. PMID- 15292480 TI - Effect of a dietary herbal supplement containing caffeine and ephedra on weight, metabolic rate, and body composition. AB - OBJECTIVE: To evaluate the effect of a dietary supplement containing herbal caffeine (70 mg/dose) and ephedra (24 mg/dose; C&E) on metabolic rate, weight loss, body composition, and safety parameters. RESEARCH METHODS AND PROCEDURES: In phase I, 12 healthy subjects with a BMI of 25 to 35 kg/m2 had resting metabolic rate (RMR) measured for 2 hours after ingesting C&E or a placebo on two occasions 1 week apart, followed by a 1-week washout before phase II. In phase II, these 12 and 28 additional subjects were randomized to a 12-week, double blind trial comparing C&E (3 times/day) to placebo. In phase III, the C&E group was given open-label C&E for 3 months, and the placebo group was given C&E for 6 months. RESULTS: In phase I, C&E gave an average 8 +/- 0.1% (SE) rise in RMR over 2 hours compared with placebo (p < 0.01). In phase II, weight loss at 12 weeks was 3.5 +/- 0.6 kg with C&E compared with 0.8 +/- 0.5 kg with placebo (p < 0.02). The percentage fat lost, shown by DXA, was 7.9 +/- 2.9% with C&E and 1.9 +/- 1.1% with placebo (p < 0.05). Pulse decreased more in the placebo group that in the C&E group (p < 0.03). There were no differences in lipid levels or blood pressure. In phase III, there was a 6-month loss of 7.3% and 7.8% of initial body weight for the groups on placebo and C&E during phase II, respectively. There were no serious adverse events. DISCUSSION: C&E increased RMR significantly by 8% compared with placebo, promoted more weight and fat loss than placebo, and was well tolerated. PMID- 15292481 TI - Postexercise muscle triacylglycerol and glycogen metabolism in obese insulin resistant zucker rats. AB - OBJECTIVE: To determine the impact of insulin resistance and obesity on muscle triacylglycerol (IMTG) and glycogen metabolism during and after prolonged exercise. RESEARCH METHODS AND PROCEDURES: Female lean (fa/?; N = 40, ZL) and obese insulin-resistant (fa/fa; N = 40, ZO) Zucker rats performed an acute bout of swimming exercise (8 times for 30 minutes) followed by 6 hours of carbohydrate supplementation (CHO) or fasting (FAST). IMTG and glycogen were measured in the extensor digitorum longus (EDL) and red vastus lateralis (RVL) muscles. RESULTS: Despite resting IMTG content being 4-fold higher in ZO compared with ZL rats, IMTG levels were unchanged in either EDL or RVL muscles immediately after exercise. Resting glycogen concentration in EDL and RVL muscles was similar between genotypes, with exercise resulting in glycogen use in both muscles from ZL rats (approximately 85%, p < 0.05). However, in ZO rats, there was a much smaller decrease in postexercise glycogen content in both EDL and RVL muscles (approximately 30%). During postexercise recovery, there was a decrease in EDL muscle levels of IMTG in ZL rats supplemented with CHO after 30 and 360 minutes (p < 0.05). In contrast, IMTG content was increased above resting levels in RVL muscles of ZO rats fasted for 360 minutes. Six hours of CHO refeeding restored glycogen content to resting levels in both muscles in ZL rats. However, after 6 hours of FAST in ZO animals, RVL muscle glycogen content was still lower than resting levels (p < 0.05). At this time, IMTG levels were elevated above basal (p < 0.05). DISCUSSION: In both healthy and insulin-resistant skeletal muscle, there was negligible net IMTG degradation after a single bout of prolonged exercise. However, during postexercise recovery, there was differential metabolism of IMTG between phenotypes. PMID- 15292482 TI - Long-term weight development in women: a 15-year follow-up of the effects of pregnancy. AB - OBJECTIVE: The aim of this study was to evaluate how well prepregnancy BMI, gestational weight gain, and postpartum weight retention predict retention of weight 15 years later among parous women. RESEARCH METHODS AND PROCEDURES: The Stockholm Pregnancy and Women's Nutrition (SPAWN) study is a long-term follow-up study of women who delivered children in 1984 to 1985 (n = 2342). The participants initially filled out questionnaires about their eating and exercise habits, social circumstances, etc. before, during, and at 1 year after pregnancy. Anthropometric data were also sampled. Fifteen years later, these women were invited to take part in the follow-up study. Anthropometric measurements were collected, and similar questions were asked. Five hundred sixty-three women participated in the SPAWN 15-year follow-up study. The sample was divided into groups to examine three presumably critical time periods: 1) overweight and normal weight before pregnancy; 2) low, intermediate, and high weight gainers during pregnancy; and 3) low, intermediate, and high weight retainers at 1 year after pregnancy. RESULTS: The overweight women did not gain more weight during pregnancy or retain more weight at 1 year follow-up. High weight gainers during pregnancy retained more weight at the 1-year and the 15-year follow-ups. High weight retainers had gained more during pregnancy and retained it at the 15-year follow-up. Fifty-six percent of the high weight gainers during pregnancy ended up in the high weight retainers group. DISCUSSION: Women who are overweight before pregnancy do not have a higher risk of postpartum weight retention than normal weight women. Thus, it is not necessarily the initially overweight woman who should be the target or focus of weight control programs during or after pregnancy. Both high weight gainers and high weight retainers had higher BMI at the 15-year follow-up, although only 56% of the high weight gainers during pregnancy were also classified as high weight retainers at the 1-year follow-up. Weight retention at the end of the postpartum year predicts future overweight 15 years later. PMID- 15292483 TI - Effect of voluntary exercise on genetically obese Cpefat/fat mice: quantitative proteomics of serum. AB - OBJECTIVE: To compare the effect of voluntary exercise on body weight, food consumption, and levels of serum proteins between wild-type and carboxypeptidase E-deficient (Cpefat/fat) mice. RESEARCH METHODS AND PROCEDURES: Study 1 consisted of three groups of female mice: Cpefat/fat mice with continuous access to exercise wheels for 3 weeks (n = 4); wild-type C57BKS mice with access to exercise wheels for 3 weeks (n = 4); and sedentary Cpefat/fat mice (n = 3). Activity, body weight, and food consumption were monitored for this period and a subsequent 9-week period without exercise wheels. Study 2 consisted of four groups of male mice (n = 6 to 7 each): Cpefat/fat mice with exercise wheels, wild type mice with exercise wheels, and Cpefat/fat and wild-type mice without exercise wheels. Body weight and food consumption were measured over 4 weeks. Sera were collected, and the protein profile was determined by 2-dimensional gel electrophoresis and mass spectrometry. RESULTS: Cpefat/fat mice were moderately hyperphagic but lost weight during the initial exercise period because of greater energy expenditure. The effect of exercise was temporary, and the mice gained weight after the second week. Several serum proteins were found to be altered by exercise: haptoglobin was decreased by exercise in Cpefat/fat mice, and several kallikreins were increased by exercise in wild-type mice. DISCUSSION: The access to exercise wheels provided an initial weight loss in Cpefat/fat mice, but this effect was offset by elevated food consumption. The serum proteomics results indicated that Cpefat/fat and wild-type mice differed in their response to exercise. PMID- 15292484 TI - The skinny on COI analysis. PMID- 15292486 TI - Microarray analysis of gene expression after transverse aortic constriction in mice. AB - Cardiac hypertrophy is a compensatory response initially beneficial to heart function but can ultimately lead to cardiac decompensation. It is an integrated process involving multiple cellular signaling pathways and their cross talk. Microarray GeneChip technology is a powerful new tool to identify gene expression profiles of cardiac hypertrophy. To identify well-characterized as well as novel adaptive mechanisms, we utilized a murine model of compensated pressure overload hypertrophy (transverse aortic constriction, TAC). At 48 h, 10 days, and 3 wk, hearts were harvested and total RNA hybridized to Affymetrix U74Av2 GeneChips, which contain a 12,488-gene/EST probe set. Verification of gene expression was performed by SYBR quantitative real-time RT-PCR (QRT-PCR) for selected genes. A rigorous evaluation of the adequacy of the control condition was also performed. For statistical analysis we generated a four-step filtering criteria. Our results show an upregulation of 38 genes (48 h), 269 genes (10 days), and 203 genes (3 wk) and downregulation of 15 genes (48 h), 160 genes (10 days), and 124 genes (3 wk). Transcripts differentially expressed after TAC were categorized into 12 functional groups and revealed the presence of several intriguing transcripts, e.g., cell proliferation-related Ki-67 and several apoptosis-related genes. Overall changes in QRT-PCR were in accordance with GeneChip data, with the highest correlation for genes with the largest up- or downregulation with TAC. Thus TAC results in altered expression of genes in several pathways regulating both cardiac structure and function. However, for in vivo gene microarray experiments, it is critical to define adequate controls, perform rigorous statistical analysis, and provide validation by alternative methods. PMID- 15292487 TI - Hyperthermia-enhanced splenic cytokine gene expression is mediated by the sympathetic nervous system. AB - Whole body hyperthermia (WBH) has been used in experimental settings as an adjunct to radiochemotherapy for the treatment of various malignant diseases. The therapeutic effect of WBH has been hypothesized to involve activation of the immune system, although the effect of hyperthermia-induced activation of sympathetic nerve discharge (SND) on splenic immune function is not known. We tested the hypothesis that heating-induced splenic sympathoexcitation would alter splenic cytokine gene expression as determined using gene array and real-time RT PCR analyses. Experiments were performed in splenic-intact and splenic-denervated anesthetized Sprague-Dawley rats (n=32). Splenic SND was increased during heating (internal temperature increased from 38 degrees to 41 degrees C) in splenic intact rats but remained unchanged in nonheated splenic-intact rats. Splenic interleukin-1beta (IL-1beta), interleukin-6 (IL-6), and growth-regulated oncogene 1 (GRO 1) mRNA expression was higher in heated than in nonheated splenic-intact rats. Splenic IL-1beta, IL-6, and GRO 1 mRNA expression was reduced in heated splenic-denervated compared with heated splenic-intact rats, but did not differ between heated splenic-denervated and nonheated splenic-intact rats. These results support the hypothesis that hyperthermia-induced activation of splenic SND enhances splenic cytokine gene expression. PMID- 15292488 TI - Trappin ovine molecule (TOM), the ovine ortholog of elafin, is an acute phase reactant in the lung. AB - As large animal models continue to play an important role in translating lung directed therapeutic strategies from laboratory animals to humans, there is an increasing interest in the analysis of endogenous regulators of inflammation at both a genomic and a therapeutic level. To this end, we have sought to characterize the ovine ortholog of elafin, an important regulator of inflammation in humans. We have isolated both the elafin cDNA and gene, which have a similar structure to other species' orthologs. Interestingly, we have isolated two alleles for ovine elafin, which contain a very high number of transglutamination repeats, thought to be important in binding elafin to the interstitium. The mainly mucosal mRNA distribution for ovine elafin suggests that ovine elafin may, like its human ortholog, have functions in innate immunity. This is supported by analysis of elafin and the related protein secretory leukocyte protease inhibitor (SLPI) in ovine bronchoalveolar fluid in response to locally administered lipopolysaccharide and confirmation of them acting as "alarm" antiproteases. We have also cloned the ovine elafin cDNA into an adenoviral vector and have demonstrated correct processing of the secreted protein as well as biological activity. Overexpression of ovine elafin in a lung-derived epithelial cell line has a protective effect against the enzymes human neutrophil and porcine pancreatic elastase. The identification of the ovine elafin gene and its translated protein are important in developing practical strategies aimed at regulating inflammation in the large mammalian lung. PMID- 15292489 TI - Dermal papilla-induced hair differentiation of adult epithelial stem cells from human skin. AB - The epithelial-mesenchymal interactions between keratinocyte stem cells and dermal papilla (DP) cells are crucial for normal development of the hair follicle as well as during hair cycling. During the cyclical regrowth of a new lower follicle, the multipotent hair follicle stem cells are stimulated to proliferate and differentiate through interactions with the underlying mesenchymal DP cells. To characterize the events occurring during the process of epithelial stem cell fate determination, we utilized a coculture system by incubating human hair follicle keratinocyte stem cells with DP cells. Using GeneChip microarrays, we analyzed changes in gene expression within the stem cells upon coculture with the DP over a 5-day time course. A number of important signaling pathways and growth factors were regulated. The hair-specific keratin 6hf (K6hf) gene proved a particularly good marker of hair differentiation, with a 7.9-fold increase in mRNA and resulting increased protein levels. The high expression of K6hf was unique to DP-induced keratinocyte differentiation, since expression of K6hf was not induced by high calcium. Since the beta-catenin signaling pathway has been implicated in hair follicle development, we examined the role of beta-catenin in our system and demonstrated that beta-catenin/lef-1 signaling is required for DP induced hair differentiation. PMID- 15292490 TI - Possible interaction between sevoflurane and Aloe vera. AB - OBJECTIVE: To describe a patient with massive intraoperative bleeding after oral consumption of Aloe vera tablets. CASE SUMMARY: A 35-year-old woman lost 5 L of blood during surgery as a result of a possible herb-drug interaction between Aloe vera and sevoflurane. DISCUSSION: Aloe vera is a common herb used for antiinflammatory and antiarthritic activity, as well as antibacterial, hypoglycemic, and lipid-lowering effects. Compounds contained within Aloe vera can cause a reduction in prostaglandin synthesis, which may inhibit secondary aggregation of platelets. Sevoflurane inhibits thromboxane A(2) formation by suppression of cyclooxygenase activity, impairs platelet aggregation, and prolongs bleeding. Although the vascularity and size of the hemangioma were the most important factors for the massive intraoperative blood loss, concomitant use of sevoflurane and Aloe vera played a contributory role. An objective causality assessment revealed that this adverse event was possible as a result of the sevoflurane and Aloe vera interaction. CONCLUSIONS: There is a potential herb drug interaction between Aloe vera and sevoflurane based on the antiplatelet effects of these 2 agents. Herbal medications with antiplatelet potential should be discontinued before anesthesia and surgery. PMID- 15292496 TI - Neutropenia due to clopidogrel in a patient with end-stage renal disease. PMID- 15292497 TI - Use of single-dose rasburicase in an obese female. AB - OBJECTIVE: To report the use of single-dose rasburicase in an obese patient. CASE SUMMARY: A 53-year-old obese African American woman weighing 136 kg (ideal body weight [IBW] 55 kg) with new-onset chronic myelomonocytic leukemia in leukocytic blast crisis was treated with hydroxyurea 5 g daily. In addition, she received allopurinol 300 mg daily for prevention of tumor lysis syndrome (TLS). The following day, allopurinol was discontinued and rasburicase was administered at a dose of 0.2 mg/kg of IBW for a serum uric acid level of 11.9 mg/dL. The patient's serum uric acid level decreased to 1.9 mg/dL 48 hours after a single dose. DISCUSSION: Rasburicase is indicated for the initial management of elevated plasma uric acid levels in patients with hematologic and solid tumor malignancies who are at risk for TLS. This case is unique because the patient received one dose of rasburicase followed by allopurinol rather than 5 daily doses of rasburicase. Additionally, the dose was based on IBW rather than actual body weight. Efficacy of this approach is apparent from the uric acid levels and the lack of hemodialysis requirements. CONCLUSIONS: A single dose of rasburicase (based on IBW) followed by allopurinol can effectively prevent TLS based on serum uric acid concentration. This approach resulted in a substantial cost savings. PMID- 15292498 TI - Nonestrogen treatment modalities for vasomotor symptoms associated with menopause. AB - OBJECTIVE: To systematically review the literature regarding the efficacy and safety of nonestrogen treatments for menopause-associated vasomotor symptoms not due to cancer or chemotherapy. DATA SOURCES: Pertinent literature and clinical studies were identified by searching MEDLINE (1966-February 2004) and EMBASE (1959-February 2004) using the key search terms vasomotor symptoms, hot flashes, and menopause. Bibliographies of relevant articles were reviewed for additional references. STUDY SELECTION AND DATA EXTRACTION: English-language articles reporting efficacy and safety of nonestrogen treatment modalities for perimenopausal and postmenopausal vasomotor symptoms were evaluated. All articles identified from the data sources were evaluated, and all information deemed relevant was included. Emphasis was placed on randomized, double-blind, placebo controlled clinical trials, as these provide the best efficacy and safety data. Studies evaluating treatment of vasomotor symptoms from other causes, such as cancer or chemotherapy, were excluded. DATA SYNTHESIS: Prescription medications reviewed for efficacy and safety in postmenopausal vasomotor symptoms include clonidine hydrochloride, danazol, gabapentin, methyldopa, mirtazapine, progestins, propranolol hydrochloride, selective serotonin-reuptake inhibitors (SSRIs), and venlafaxine. Nonprescription therapies reviewed include black cohosh, dong quai, evening primrose oil, physical activity, phytoestrogens, and red clover. CONCLUSIONS: According to this systematic literature review, postmenopausal vasomotor treatments that have been shown to be safe and effective in short-term use include black cohosh, exercise, gabapentin, medroxyprogesterone acetate, SSRIs (ie, paroxetine hydrochloride), and soy protein. Initial, small reports are suggestive for efficacy in menopausal vasomotor symptoms with megestrol acetate and venlafaxine. PMID- 15292499 TI - Intentional topiramate ingestion in an adolescent female. AB - OBJECTIVE: To describe an intentional topiramate ingestion by an adolescent and warn of the potential for topiramate abuse. CASE SUMMARY: A 17-year-old female intentionally ingested approximately eight 100-mg topiramate tablets for the purpose of "getting high." Soon after ingestion, she was found at school obtunded and nonresponsive. Upon transfer to the emergency department, she became combative and aggressive with evolving neurologic abnormalities including incoherence, confusion, disorientation, and significant speech impairments including echolalia. Approximately 24 hours after ingestion, the patient had completely recovered without requiring specific treatment or experiencing sequelae. DISCUSSION: The clinical effects following acute topiramate intoxication appear consistent with the drug's known pharmacologic properties. There are few other reports of topiramate ingestions and most cases have had mild outcomes. CONCLUSIONS: Due to the multifactorial effects topiramate may have upon the central nervous system and its anorectic effect, abuse of this drug by adolescents should be considered upon presentation of an adolescent with mental status changes. PMID- 15292500 TI - Treating functional impairment of autism with selective serotonin-reuptake inhibitors. AB - OBJECTIVE: To review literature describing use of selective serotonin-reuptake inhibitors (SSRIs) in the management of functional impairments associated with autistic disorder. DATA SOURCES: EMBASE (1980-3rd quarter of 2003), International Pharmaceutical Abstracts (1970-August 2003), and MEDLINE (1966-August 2003) were searched. Search terms included autism, autistic disorder, citalopram, fluoxetine, fluvoxamine, paroxetine, selective serotonin-reuptake inhibitors, and sertraline. DATA SYNTHESIS: Studies and case reports evaluating treatment outcomes associated with the use of SSRIs in managing impairments of autism were reviewed. Multiple SSRI dosing ranges were evaluated in autistic patients of different ages with various functional impairments. No specific SSRI or dose range has been shown to improve a specific autistic symptom although some patients have demonstrated improvements. CONCLUSIONS: Benefits with SSRIs in treating functional impairments in autism have been observed. Response to therapy and adverse effects are individualized. Current evidence does not support selection of one SSRI over another for any impairment associated with autism. PMID- 15292501 TI - Comment: drug-related problem classification systems. PMID- 15292502 TI - Comment: drug-related problem classification systems. PMID- 15292503 TI - Comment: drug-related problem classification systems. PMID- 15292504 TI - DNA vaccines to attack cancer. AB - Delivery of antigens by injection of the encoding DNA allows access to multiple antigen-presenting pathways. Knowledge of immunological processes can therefore be used to modify construct design to induce selected effector functions. Expression can be directed to specific intracellular sites, and additional genes can be fused or codelivered to amplify responses. Therapeutic vaccination against cancer adds a requirement to overcome tolerance and to activate a weakened immune repertoire. Induction of CD4(+) T helper cells is critical for both antibody and T cell effector responses. To activate immunity against tumor antigens, we fused the tumor-derived sequences to genes encoding microbial proteins. This strategy engages T helper cells from the large antimicrobial repertoire for linked help for inducing antibody against cell-surface tumor antigens. The principle of linked T cell help also holds for induction of epitope-specific antitumor CD8(+) T cells, but the microbial sequence has to be minimized to avoid competition with tumor antigens. Epitope-specific DNA vaccination leads to powerful antitumor attack and can activate immunity from a profoundly tolerized repertoire. Vaccine designs validated in preclinical models are now in clinical trial with immune responses detected against both tumor antigens and fused microbial antigens. DNA priming is highly efficient, but boosting may benefit from increased antigen expression. Physical methods including electroporation provide increased expression without introducing additional competing antigens. A wide range of cancers can be targeted, and objective assays of response will determine efficacy. PMID- 15292505 TI - Humoral immune response to native eukaryotic prion protein correlates with anti prion protection. AB - Prion diseases are characterized by the deposition of an abnormal form (termed PrP(Sc)) of the cellular prion protein (PrP(C)). Because antibodies to PrP(C) can antagonize deposition of PrP(Sc) in cultured cells and mice, they may be useful for anti-prion therapy. However, induction of protective anti-prion immune responses in WT animals may be hindered by host tolerance. Here, we studied the cellular and molecular basis of tolerance to PrP(C). Immunization of Prnp(o/o) mice with bacterially expressed PrP (PrP(REC)) resulted in vigorous humoral immune responses to PrP(REC) and native cell-surface PrP(C). Instead, WT mice yielded antibodies that failed to recognize native PrP(C) despite immunoreactivity with PrP(REC), even after immunization with PrP-PrP polyprotein and/or upon administration of anti-OX40 antibodies. Consequently, immunized WT mice experienced insignificantly delayed prion pathogenesis upon peripheral prion challenge. Anti-PrP immune responses in Prnp(o/o) mice were completely abrogated by transgenic expression of PrP(C) in B cells, T cells, neurons, or hepatocytes, but only moderately reduced by expression in myelinating cells, despite additional thymic Prnp transcription in each case. We conclude that tolerance to PrP(C) can coexist with immunoreactivity to PrP(REC) and does not depend on thymic PrP(C) expression. Its circumvention might represent an important step toward the development of effective anti-prion immunotherapy. PMID- 15292506 TI - The in and out of monocytes in atherosclerotic plaques: Balancing inflammation through migration. PMID- 15292507 TI - De novo design of catalytic proteins. AB - The de novo design of catalytic proteins provides a stringent test of our understanding of enzyme function, while simultaneously laying the groundwork for the design of novel catalysts. Here we describe the design of an O(2)-dependent phenol oxidase whose structure, sequence, and activity are designed from first principles. The protein catalyzes the two-electron oxidation of 4-aminophenol (k(cat)/K(M) = 1,500 M(-1).min(-1)) to the corresponding quinone monoimine by using a diiron cofactor. The catalytic efficiency is sensitive to changes of the size of a methyl group in the protein, illustrating the specificity of the design. PMID- 15292508 TI - Single-molecule study of RuvAB-mediated Holliday-junction migration. AB - Branch migration of Holliday junctions is an important step of genetic recombination and DNA repair. In Escherichia coli, this process is driven by the RuvAB complex acting as a molecular motor. Using magnetic tweezers, we studied the RuvAB-directed migration of individual Holliday junctions formed between two approximately 6-kb DNA molecules of identical sequence, and we measured the migration rate at 37 degrees C and 1 mM ATP. We directly demonstrate that RuvAB is a highly processive DNA motor protein that is able to drive continuous and unidirectional branch migration of Holliday junctions at a well defined average speed over several kilobases through homologous sequences. We observed directional inversions of the migration at the DNA molecule boundaries leading to forth-and-back migration of the branch point and allowing us to measure the migration rate in the presence of negative or positive loads. The average migration rate at zero load was found to be approximately 43 bp/sec. Furthermore, the load dependence of the migration rate is small, within the force range of 3.4 pN (hindering force) to +3.4 pN (assisting force). PMID- 15292509 TI - Direct observation of RuvAB-catalyzed branch migration of single Holliday junctions. AB - Holliday junctions form during DNA repair and homologous recombination processes. These processes entail branch migration, whereby the length of two arms of a cruciform increases at the expense of the two others. Branch migration is carried out in prokaryotic cells by the RuvAB motor complex. We study RuvAB-catalyzed branch migration by following the motion of a small paramagnetic bead tethered to a surface by two opposing arms of a single cruciform. The bead, pulled under the action of magnetic tweezers, exerts tension on the cruciform, which in turn transmits the force to a single RuvAB complex bound at the crossover point. This setup provides a unique means of measuring several kinetic parameters of interest such as the translocation rate, the processivity, and the force on the substrate against which the RuvAB complex cannot effect translocation. RuvAB-catalyzed branch migration proceeds with a small, discrete number of rates, supporting the view that the monomers comprising the RuvB hexameric rings are not functionally homogeneous and that dimers or trimers constitute the active subunits. The most frequently encountered rate, 98 +/- 3 bp/sec, is approximately five times faster than previously estimated. The apparent processivity of branch migration between pauses of inactivity is approximately 7,000 bp. Branch migration persists against opposing forces up to 23 pN. PMID- 15292510 TI - Nitrated lipids: a class of cell-signaling molecules. PMID- 15292512 TI - Bayesian Markov chain Monte Carlo sequence analysis reveals varying neutral substitution patterns in mammalian evolution. AB - We describe a model of neutral DNA evolution that allows substitution rates at a site to depend on the two flanking nucleotides ("context"), the branch of the phylogenetic tree, and position within the sequence and implement it by using a flexible and computationally efficient Bayesian Markov chain Monte Carlo approach. We then apply this approach to characterize phylogenetic variation in context-dependent substitution patterns in a 1.7-megabase genomic region in 19 mammalian species. In contrast to other substitution types, CpG transition substitutions have accumulated in a relatively clock-like fashion. More broadly, our results support the notion that context-dependent DNA replication errors, cytosine deamination, and biased gene conversion are major sources of naturally occurring mutations whose relative contributions have varied in mammalian evolution as a result of changes in generation times, effective population sizes, and recombination rates. PMID- 15292511 TI - Tetrodotoxin-sensitive Na+ channels and muscarinic and purinergic receptors identified in human erythroid progenitor cells and red blood cell ghosts. AB - This article concerns the identification of different types of voltage-gated Na(+) channels and of muscarinic and purinergic receptors that are expressed in human erythroid precursor cells and red cell ghosts. We analyzed, by RT-PCR, RNA that was extracted from purified and synchronously growing human erythroid progenitor cells, differentiating from erythroblasts to reticulocytes in 7 to 14 days. These extracts were free of white cell and platelet contamination. Two types of voltage-gated, tetrodotoxin-sensitive Na(+) channels were found. These were Na(v)1.4 and Na(v)1.7, the former known to be present in skeletal muscle and the latter in peripheral nerve. By using a pan Na(+) channel antibody and Western blotting, an immunoreactive channel was detected in ghosts of human red blood cells, consistent with the expression of these two channels. The transcripts for four of the five known subtypes of muscarinic receptors were also identified, including subtypes M2, M3, M4, and M5, whereas subtype M1 was not found. Expression was also detected for the purinergic type receptors P2X(1), P2X(4), P2X(7), and P2Y(1) whereas types P2Y(2), P2Y(4), and P2Y(6) were not found. We also searched for but did not find transcripts for hBNP-1, a type 1b human brain sodium phosphate cotransporter, and cystic fibrosis transmembrane conductance regulator (CFTR). Implications regarding the presence of these different types of channels and receptors in human red blood cells and their functional significance are discussed. PMID- 15292513 TI - Modified amino acid copolymers suppress myelin basic protein 85-99-induced encephalomyelitis in humanized mice through different effects on T cells. AB - A humanized mouse bearing the HLA-DR2 (DRA/DRB1*1501) protein associated with multiple sclerosis (MS) and the myelin basic protein (MBP) 85-99-specific HLA-DR2 restricted T cell receptor from an MS patient has been used to examine the effectiveness of modified amino acid copolymers poly(F,Y,A,K)n and poly (V,W,A,K)n in therapy of MBP 85-99-induced experimental autoimmune encephalomyelitis (EAE) in comparison to Copolymer 1 [Copaxone, poly(Y,E,A,K)n]. The copolymers were designed to optimize binding to HLA-DR2. Vaccination, prevention, and treatment of MBP-induced EAE in the humanized mice with copolymers FYAK and VWAK ameliorated EAE more effectively than Copolymer 1, reduced the number of pathological lesions, and prevented the up-regulation of human HLA-DR on CNS microglia. Moreover, VWAK inhibited MBP 85-99-specific T cell proliferation more efficiently than either FYAK or Copolymer 1 and induced anergy of HLA-DR2-restricted transgenic T cells as its principle mechanism. In contrast, FYAK induced proliferation and a pronounced production of the antiinflammatory T helper 2 cytokines IL-4 and IL-10 from nontransgenic T cells as its principle mechanism of immunosuppression. Thus, copolymers generated by using different amino acids inhibited disease using different mechanisms to regulate T cell responses. PMID- 15292514 TI - Amelioration of proteolipid protein 139-151-induced encephalomyelitis in SJL mice by modified amino acid copolymers and their mechanisms. AB - Copolymer 1 [Cop1, glatiramer acetate, Copaxone, poly(Y,E,A,K)n] is widely used in the treatment of relapsing/remitting multiple sclerosis in which it reduces the frequency of relapses by approximately 30%. In the present study, copolymers with modified amino acid compositions (based on the binding motif of myelin basic protein 85-99 to HLA-DR2) have been developed with the aim of suppressing multiple sclerosis more effectively. The enhanced efficacy of these copolymers in experimental autoimmune encephalomyelitis (EAE) induced in SJL/J mice with proteolipid protein 139-151 was demonstrated by using three protocols: (i) simultaneous administration of autoantigen and copolymer (termed prevention), (ii) pretreatment with copolymers (vaccination), or (iii) administration of copolymers after disease onset (treatment). Strikingly, in the treatment protocol administration of soluble VWAK and FYAK after onset of disease led to stasis of its progression and suppression of histopathological evidence of EAE. The mechanisms by which these effects are achieved have been examined in several types of assays: binding of copolymers to I-A(s) in competition with proteolipid protein 139-151 (blocking), cytokine production by T cells (T helper 2 polarization), and transfer of protection by CD3(+) splenocytes or, notably, by copolymer-specific T cell lines (induction of regulatory T cells). The generation of these copolymer-specific regulatory T cells that secrete IL-4 and IL-10 and are independent of the immunizing autoantigen is very prominent among the multiple mechanisms that account for the observed suppressive effect of copolymers in EAE. PMID- 15292515 TI - A eukaryotic BLUF domain mediates light-dependent gene expression in the purple bacterium Rhodobacter sphaeroides 2.4.1. AB - The flavin-binding BLUF domain functions as a blue-light receptor in eukaryotes and bacteria. In the photoreceptor protein photo-activated adenylyl cyclase (PAC) from the flagellate Euglena gracilis, the BLUF domain is linked to an adenylyl cyclase domain. The PAC protein mediates a photophobic response. In the AppA protein of Rhodobacter sphaeroides, the BLUF domain is linked to a downstream domain without similarity to known proteins. AppA functions as a transcriptional antirepressor, controlling photosynthesis gene expression in the purple bacterium R. sphaeroides in response to light and oxygen. We fused the PACalpha1-BLUF domain from Euglena to the C terminus of AppA. Our results show that the hybrid protein is fully functional in light-dependent gene repression in R. sphaeroides, despite only approximately 30% identity between the eukaryotic and the bacterial BLUF domains. Furthermore, the bacterial BLUF domain and the C terminus of AppA can transmit the light signal even when expressed as separated domains. This finding implies that the BLUF domain is fully modular and can relay signals to completely different output domains. PMID- 15292516 TI - Detection of the mycophenolate-inhibited form of IMP dehydrogenase in vivo. AB - IMP dehydrogenase (IMPDH) is the rate-limiting enzyme for de novo GMP synthesis. Its activity is correlated with cell growth, and it is the target of a number of proven and experimental drug therapies including mycophenolic acid (MPA). MPA inhibits the enzyme by trapping a covalent nucleotide-enzyme intermediate. Saccharomyces cerevisiae has four IMPDH genes called IMD1-IMD4. IMD2 is transcriptionally regulated and is the only one that enables yeast to grow in the presence of MPA. We show here that de novo synthesis of the IMD2-encoded protein is strongly induced upon MPA treatment. We also monitor the in vivo formation of a covalent nucleotide-enzyme intermediate for Imd2, Imd3, and Imd4 that accumulates in the presence of MPA. Complete formation of the Imd2 intermediate requires drug concentrations manyfold higher than that required to quantitatively trap the Imd3- or Imd4-nucleotide adducts. Purification of the tagged IMD gene products reveals that the family of polypeptides coassemble to form heteromeric IMPDH complexes, suggesting that they form mixed tetramers. These data demonstrate that S. cerevisiae harbor multiple IMPDH enzymes with varying drug sensitivities and offer an assay to monitor the inhibition of IMPDH in living cells. They also suggest that mixed inhibition profiles may result from heteromeric complexes in cell types that contain multiple IMPDH gene products. The mobility shift assay could serve as a tool for the detection of drug inactivated IMPDH in the cells of patients receiving MPA therapy. PMID- 15292517 TI - Ca2+/calmodulin-dependent protein kinase II potentiates ATP responses by promoting trafficking of P2X receptors. AB - To elucidate the functional link between Ca(2+)/calmodulin protein kinase II (CaMKII) and P2X receptor activation, we studied the effects of electrical stimulation, such as occurs in injurious conditions, on P2X receptor-mediated ATP responses in primary sensory dorsal root ganglion neurons. We found that endogenously active CaMKII up-regulates basal P2X3 receptor activity in dorsal root ganglion neurons. Electrical stimulation causes prolonged increases in ATP currents that lasts up to approximately 45 min. In addition, the total and phosphorylated CaMKII are also up-regulated. The enhancement of ATP currents depends on Ca(2+) and calmodulin and is completely blocked by the CaMKII inhibitor, 2-[N-(2-hydroxyethyl)]-N-(4-methoxybenzenesulfonyl)]amino-N-(4 chlorocinnamyl)-N-methylbenzylamine). Western analyses indicate that electrical stimulation enhances the expression of P2X3 receptors in the membrane and that the enhancement is blocked by the inhibitor. These results suggest that CaMKII up regulated by electrical stimulation enhances ATP responses by promoting trafficking of P2X receptors to the membrane and may play a key role in the sensitization of P2X receptors under injurious conditions. PMID- 15292518 TI - The pyruvate formate lyase family: sequences, structures and activation. AB - We cloned and expressed in Escherichia coli the Archaeglobus fulgidus gene that encodes pyruvate formate lyase 2 (PFL2). PFL2, despite its homology to the other glycyl radical enzymes, differs from them by exhibiting a completely different oligomerization. The most abundant form of PFL2 when expressed in E.coli is a trimer. The closest homologue of PFL2 with a known structure is E. coli PFL, which is a dimer. Sequence comparisons allowed us to reclassify PFL-like enzymes and the consensus sequences allowed us to propose an activation route for PFL like glycyl radical enzymes. Surprisingly, most of the conserved residues in PFL like enzymes appear to be involved in preserving the structure, rather than forming the active site. PMID- 15292519 TI - A system based on specific protein-RNA interactions for analysis of target protein-protein interactions in vitro: successful selection of membrane-bound Bak Bcl-xL proteins in vitro. AB - Ribosome display systems are very effective and powerful tools for in vitro screening of transcribed mRNAs that encode proteins (or peptides) with specific (known or unknown) functions. We have modified such a system by exploiting the interaction between a tandemly fused MS2 coat-protein (MSp) dimer and the RNA sequence of the corresponding specific binding motif, C-variant (or Cv). We placed the MSp dimer at the N-terminus of a nascent protein and the Cv binding motif was attached to the 5' end of the protein's mRNA. This configuration enhanced the stability of the ribosome-mRNA complex. We demonstrate here that this improved ribosome display system provides an effective method for identifying the gene for a protein that binds to a protein of interest. We visualized the formation of polysome complexes in this advanced polysome display by atomic force microscopy (AFM) and found that the AFM images of polysomes in our system were different from those observed in the case of conventional ribosome display systems. Our results suggest that our technology might usefully complement yeast two-hybrid assays. PMID- 15292520 TI - Designer androgens in sport: when too much is never enough. AB - The recent identification of tetrahydrogestrinone (THG), the first true "designer androgen," as a sports doping agent reflects both an alarmingly sophisticated illicit manufacturing facility and an underground network of androgen abusers in elite sports, as well as the still untapped potential for designer androgens in medicine. Never marketed, THG was apparently developed as a potent androgen that was undetectable by conventional International Olympic Committee-mandated urinary sports doping tests. As a potent androgen and progestin with unspecified contaminants, its distribution for use at high doses without any prior biological or toxicological evaluation poses significant health risks. Yet this diversion of science also highlights the prospect of designer androgens for use in human medicine. Designer androgens also offer the possibility of tissue-specific effects enhancing the beneficial effects of androgens while mitigating the undesirable ones. Further developments require better understanding of the postreceptor tissue selectivity of androgens, comparable to the mechanism underlying that of partial estrogen agonists (SERMs). This experience highlights the ongoing need for vigilance to detect novel drug doping strategies in order to maintain fairness and safety in elite sports. This will require the deployment of generic catch-all tests, such as sensitive and specific in vitro androgen bioassays, coupled with the development of mass spectrometry-based tests for specific doping agents. PMID- 15292521 TI - Molecular and cellular determinants of skeletal muscle atrophy and hypertrophy. AB - The maintenance of adult skeletal muscle mass is ensured by physical exercise. Accordingly, physiological and pathological situations characterized by either impaired motor neuron activity, reduced gravity (microgravity during space flights), or reduced physical activity result in loss of muscle mass. Furthermore, a plethora of clinical conditions, including cancer, sepsis, diabetes, and AIDS, are associated with varying degrees of muscle atrophy. The cellular and molecular pathways responsible for maintaining the skeletal muscle mass are not well defined. Nonetheless, studies aimed at the understanding of the mechanisms underlying either muscular atrophy or hypertrophy have begun to identify the physiological determinants and clarify the molecular pathways responsible for the maintenance of muscle mass. PMID- 15292522 TI - A re-evaluation of the occupancy factors for effective dose estimate in tropical environment. AB - In the estimation of the effective dose to the public, outdoor and indoor occupancy factors have been an important parameter. These factors vary, depending on the prevailing environmental condition in a particular location. The factors have been estimated for the rural and urban areas in Nigeria. An outdoor factor of 0.3 and 0.22 have been estimated for rural and urban dwellers, respectively. The rural outdoor factor is 50% above the value recommended as the world average by the UNSCEAR. The urban outdoor factor is 10% higher than this value. The total outdoor gamma dose rate in the air due to (40)K, (238)U and (232)Th in the soil for some rural population in the southern part of Nigeria is 29.50 +/- 3.80 nGy h(-1) and the average outdoor effective dose has been estimated to be 54.28 +/- 6.95 microSv y(-1) using the present occupancy factor. PMID- 15292523 TI - Comparison of observed lung retention and urinary excretion of thorium workers and members of the public in India with the values predicted by the ICRP biokinetic model. AB - The daily intake of natural Th and its contents in lungs, skeleton and liver of an Indian adult population group were estimated using radiochemical neutron activation analysis (RNAA) technique. These data on daily intake (through inhalation and ingestion) were used to compute Th contents in lungs and other systemic organs such as skeleton and liver using the new human respiratory tract model (HRTM) and the new biokinetic model of Th. The theoretically computed Th contents in lungs, skeleton and liver of an average Indian adult are 2.56, 4.00 and 0.17 microg, respectively which are comparable with the corresponding experimentally measured values of 4.31, 3.45 and 0.14 microg in an urban population group living in Mumbai. The measured lung contents of Th in a group of five occupational workers were used to compute their total body Th contents and the corresponding daily urinary excretions. The computed total body contents and daily urinary excretions of Th in the five subjects compared favourably with their measured values. These studies, thus, validate the new biokinetic model of Th in natural as well as in occupational exposures in Indian conditions. PMID- 15292524 TI - Radiation dose measurements to the interventional cardiologist using an electronic personal dosemeter. AB - The aim of this study was to investigate the use of an electronic personal dosemeter (EPD) worn by a senior cardiologist in an Interventional Cardiology (IC) Laboratory of a busy cardiac centre and how the results could help in the evaluation of radiation protection equipment used. Patient samples consist of 28 patients (10 coronary angiographies (CAs) and 18 percutaneous transluminal coronary angioplasties (PTCAs)). Patient dose was measured with a dose-area product (DAP) meter. Cardiologist radiation dose value written on the EPD as well as the protective equipment used was collected. Between patient and cardiologist dose, a significant correlation was found in CA and a moderate correlation in PTCA. Mean cardiologist effective dose E per procedure was found to be 0.2 microSv in CA and 0.3 microSv in PTCA. EPD proved to be an easy, direct and straightforward way to measure the radiation dose that the cardiologist receives in an IC laboratory. PMID- 15292525 TI - Radioactivity level in Chinese building ceramic tile. AB - The activity concentrations of (226)Ra, (232)Th and (40)K have been determined by gamma ray spectrometry. The concentrations of (226)Ra, (232)Th and (40)K range from 158.3 to 1087.6, 91.7 to 1218.4, and 473.8 to 1031.3 Bq kg(-1) for glaze, and from 63.5 to 131.4, 55.4 to 106.5, and 386.7 to 866.8 Bq kg(-1) for ceramic tile, respectively. The measured activity concentrations for these radionuclides were compared with the reported data of other countries and with the typical world values. The radium equivalent activities (Ra(eq)), external hazard index (H(ex)) and internal hazard index (H(in)) associated with the radionuclides were calculated. The Ra(eq) values of all ceramic tiles are lower than the limit of 370 Bq kg(-1). The values of H(ex) and H(in) calculated according to the Chinese criterion for ceramic tiles are less than unity. The Ra(eq) value for the glaze of glazed tile collected from some areas are >370 Bq kg(-1). PMID- 15292526 TI - Rheumatological prescribing in athletes: a review of the new World Anti-Doping Agency guidelines. AB - Rheumatologists, with their musculoskeletal background, often care for athletes. The effect of a positive anti-doping test, whether through illegitimate use or accidental prescribing of banned drugs, is devastating to an athlete's career. It is therefore incumbent upon rheumatologists to be aware of issues relating to drugs in sport. This involves both therapeutic drugs and doping. It is vital to ensure that any substance prescribed should be approved for use and should not adversely affect (or benefit) the athlete's performance. In March 2004, 5 months prior to the 2004 Olympic Games in Athens, the joint World Anti-Doping Agency/International Olympic Committee published the revised list of banned substances in athletes. This article aims to provide an overview of the current status of medications commonly prescribed in rheumatological practice. PMID- 15292527 TI - Osteoarthritis, angiogenesis and inflammation. AB - Angiogenesis and inflammation are closely integrated processes in osteoarthritis (OA) and may affect disease progression and pain. Inflammation can stimulate angiogenesis, and angiogenesis can facilitate inflammation. Angiogenesis can also promote chondrocyte hypertrophy and endochondral ossification, contributing to radiographic changes in the joint. Inflammation sensitizes nerves, leading to increased pain. Innervation can also accompany vascularization of the articular cartilage, where compressive forces and hypoxia may stimulate these new nerves, causing pain even after inflammation has subsided. Inhibition of inflammation and angiogenesis may provide effective therapeutics for the treatment of OA by improving symptoms and retarding joint damage. This review aims to summarize (i) the evidence that angiogenesis and inflammation play an important role in the pathophysiology of OA and (ii) possible directions for future research into therapeutics that could effectively treat this disease. PMID- 15292528 TI - Detection of differentially expressed genes in synovial fibroblasts by restriction fragment differential display. AB - OBJECTIVE: To identify differentially expressed genes in synovial fibroblasts and examine the effect on gene expression of exposure to TNF-alpha and IL-1beta. METHODS: Restriction fragment differential display was used to isolate genes using degenerate primers complementary to the lysophosphatidic acid acyl transferase gene family. Differential gene expression was confirmed by reverse transcription-polymerase chain reaction and immunohistochemistry using a variety of synovial fibroblasts, including cells from patients with osteoarthritis and self-limiting parvovirus arthritis. RESULTS: Irrespective of disease process, synovial fibroblasts constitutively produced higher levels of IL-6 and monocyte chemoattractant protein 1 (MCP-1) (CCL2) than skin fibroblasts. Seven genes were differentially expressed in synovial fibroblasts compared with skin fibroblasts. Of these genes, four [tissue factor pathway inhibitor 2 (TFPI2), growth regulatory oncogene beta (GRObeta), manganese superoxide dismutase (MnSOD) and granulocyte chemotactic protein 2 (GCP-2)] were all found to be constitutively overexpressed in synoviocytes derived from patients with osteoarthritis. These four genes were only weakly expressed in other synovial fibroblasts (rheumatoid and self-limiting parvovirus infection). However, expression in all types of fibroblasts was increased after stimulation with TNF-alpha and IL-1beta. Three other genes (aggrecan, biglycan and caldesmon) were expressed at higher levels in all types of synovial fibroblasts compared with skin fibroblasts even after stimulation with TNF-alpha and IL-1. CONCLUSIONS: Seven genes have been identified with differential expression patterns in terms of disease process (osteoarthritis vs rheumatoid arthritis), state of activation (resting vs cytokine activation) and anatomical location (synovium vs skin). Four of these genes, TFPI2, GRObeta (CXCL2), MnSOD and GCP-2 (CXCL6), were selectively overexpressed in osteoarthritis fibroblasts rather than rheumatoid fibroblasts. While these differences may represent differential behaviour of synovial fibroblasts in in vitro culture, these observations suggest that TFPI2, GRObeta (CXCL2), MnSOD and GCP-2 (CXCL6) may represent new targets for treatments specifically tailored to osteoarthritis. PMID- 15292529 TI - Protein oxidation status in patients with ankylosing spondylitis. AB - OBJECTIVES: Ankylosing spondylitis (AS) is a chronic inflammatory disease with unknown aetiology. Since various functions of neutrophils are increased in AS, neutrophil activation-mediated oxidative stress may have an important role in the pathogenesis of AS. Therefore, the importance of neutrophil activation as the main source of oxidative stress was investigated in patients with AS. METHODS: Forty-one patients with AS, divided into active and inactive groups according to their CRP and ESR, and 30 healthy volunteers were entered into the study. The Bath Ankylosing Spondylitis Disease Activity Index (BASDAI) and the visual analogue scale (VAS) were also used in all patients before the study. In addition, the patients were evaluated according to spinal involvement and peripheral involvement. Plasma myeloperoxidase (MPO) activity, representing neutrophil activation, and oxidative stress biomarkers, such as advanced oxidation protein products (AOPP) and thiol levels, were measured. RESULTS: There were significant increases in plasma MPO activity and AOPP but decreases in thiol levels in the total AS patient group and in the subgroups, compared with controls. All parameters, except thiol, were higher in active and peripheral involvement groups than in the inactive and spinal patients, respectively; but the highest levels were observed in the peripheral subgroup. In addition, AOPP was found to be positively correlated with ESR, CRP and BASDAI in the total patient group and with white blood cell and neutrophil count in the peripheral involvement subgroup. BASDAI was positively correlated with VAS, ESR and CRP; but MPO was negatively correlated with thiol/albumin ratio, both in total and active AS patients. Negative correlations were also observed between albumin and ESR, and between CRP and neutrophil counts in the peripheral subgroup. CONCLUSIONS: This is the first study suggesting neutrophil-MPO-hypochlorous acid-mediated AOPP formation in AS. Therefore, active neutrophils and chlorinated oxidants of neutrophil origin may be considered to be important factors in the pathogenesis of AS which are related to oxidative stress, notably protein oxidation. PMID- 15292530 TI - Rheumatoid cachexia: metabolic abnormalities, mechanisms and interventions. AB - We have previously identified the phrase 'rheumatoid cachexia' to describe the loss of body cell mass (BCM) that may occur among patients with rheumatoid arthritis (RA). Specifically, rheumatoid cachexia is characterized by altered energy and protein metabolism (reduced total energy expenditure, increased resting energy expenditure and increased whole-body protein catabolism) and increased inflammatory cytokine production (interleukin-1beta and tumour necrosis factor-alpha). Patients with rheumatoid cachexia consistently have a diet that appears adequate in protein and calories (based on US Dietary Reference Intakes), but with reduced physical activity. These phenomena are similar to some of the metabolic abnormalities that occur with normal ageing, but the aetiology appears to be different in RA. This review will focus on describing the metabolic abnormalities observed in rheumatoid cachexia, identifying potential mechanisms for loss of BCM and discussing strategies for intervention. PMID- 15292531 TI - The impact of low family income on self-reported health outcomes in patients with rheumatoid arthritis within a publicly funded health-care environment. AB - OBJECTIVE: Self-rated health (SRH) is an independent, strong predictor of morbidity and mortality. Socio-economic status (SES) is strongly associated with SRH. This study investigated the relationship between SES and SRH outcomes in a sample of patients with rheumatoid arthritis (RA) in Canada. METHODS: Both generic preference-based [Health Utilities Index Mark 3 (HUI3) and Short Form 6D (SF-6D)] and non-preference-based [disease-specific (Rheumatoid Arthritis Quality of Life, RAQoL) and a functional status (Health Assessment Questionnaire, HAQ)] SRH questionnaires were administered to 313 RA patients. Both proximate (education and annual household income) and contextual (neighbourhood income, education and unemployment) measures of SES were captured. Ordinary least squares (OLS) regression was used to adjust for RA severity while assessing the relationship between SRH and SES measures. Two-stage least-squares (TSLS) regression was used to determine if there was an inter-relationship between SES and SRH measures. RESULTS: The sample was well distributed across RA severity and SES measures. Contextual and proximate measures of SES were poorly correlated. Lower levels of proximate SES measures (but not contextual) were associated with poorer SRH outcomes. The OLS regressions showed significant associations between the HUI3 and the SF-6D overall scores and the HAQ for self-reported income. The RAQoL did not differ significantly across SES. TSLS regression confirmed the finding that self-reported income was similarly associated with the SRH measures. CONCLUSIONS: Even in a country with universal access to health-care, the impact of a chronic disease such as RA on SRH is associated with self-reported income. The finding that preference-based measures vary with income independently of RA severity could bias economic evaluation. PMID- 15292532 TI - Ten days matter. PMID- 15292533 TI - Personal accounts: help-seeking preferences of high school students: the impact of personal narratives. PMID- 15292534 TI - Datapoints: acceptability of asking parents about their children's traumatic symptoms. PMID- 15292535 TI - Growth of mental health services in state adult correctional facilities, 1988 to 2000. AB - OBJECTIVE: This study examined trends in the availability and use of mental health services in state adult correctional facilities. METHODS: Results from the 1988 Inventory of Mental Health Services in State Adult Correctional Facilities of the Center for Mental Health Services were compared with those from the 2000 Census of State and Federal Adult Correctional Facilities survey of the Bureau of Justice Statistics. The two surveys were chosen because they occurred more than a decade apart, had a reasonable amount of data, and could be made comparable. RESULTS: This analysis used data from 757 state adult correctional facilities that were sampled in 1988. The number of such facilities increased to 1,097 in 2000, a 44.9 percent increase. A dramatic increase was also seen in the prison population, from 505,712 in 1988 to 1,084,625 in 2000, a 114.5 percent increase. Mental health services were offered in significantly more facilities in 2000 than in 1988. However, the relative percentage of facilities that offered mental health services decreased overall. Simultaneously, the percentage of inmates who used these services increased overall. CONCLUSIONS: The growth in prison facilities and the growth in prisoner populations are outstripping the more meager growth in mental health services. These results suggest that mental health services are becoming less available to the prison population, and service populations are becoming more concentrated in the facilities that do offer such services. PMID- 15292536 TI - Relationship between race and ethnicity and forensic clinical triage dispositions. AB - OBJECTIVE: Racial and ethnic disparities in the criminal justice system have been widely reported, as have racial and ethnic disparities in diagnoses and certain aspects of clinical management. This study examined the association between race and ethnicity and dispositions for pretrial defendants who were referred for forensic mental health evaluations. METHODS: Available data were reviewed for all defendants in Massachusetts who were referred to a Massachusetts court clinic from 1994 to 2001 for a screening evaluation of their competence to stand trial, their criminal responsibility, or both. Logistic regression models were developed to assess the relationship between defendants' race and ethnicity and the likelihood that they would be referred for inpatient evaluation and the likelihood that they would be evaluated within a strict-security facility. Race or ethnicity of the pretrial defendants was identified by clinicians. RESULTS: Blacks, but not Hispanics, were significantly more likely than whites to be referred for an inpatient evaluation after an outpatient forensic screening evaluation. Among male defendants, both Hispanics and blacks were more likely than whites to be referred for an inpatient evaluation in a strict-security facility, regardless of diagnoses and the level of severity of the criminal charges. CONCLUSIONS: Racial and ethnic disparities in disposition decisions exist within the forensic mental health system. These disparities, however, likely reflect numerous clinician and nonclinician variables. PMID- 15292537 TI - A computerized clinical decision support system as a means of implementing depression guidelines. AB - The authors describe the history and current use of computerized systems for implementing treatment guidelines in general medicine as well as the development, testing, and early use of a computerized decision support system for depression treatment among "real-world" clinical settings in Texas. In 1999 health care experts from Europe and the United States met to confront the well-documented challenges of implementing treatment guidelines and to identify strategies for improvement. They suggested the integration of guidelines into computer systems that is incorporated into clinical workflow. Several studies have demonstrated improvements in physicians' adherence to guidelines when such guidelines are provided in a computerized format. Although computerized decision support systems are being used in many areas of medicine and have demonstrated improved patient outcomes, their use in psychiatric illness is limited. The authors designed and developed a computerized decision support system for the treatment of major depressive disorder by using evidence-based guidelines, transferring the knowledge gained from the Texas Medication Algorithm Project (TMAP). This computerized decision support system (CompTMAP) provides support in diagnosis, treatment, follow-up, and preventive care and can be incorporated into the clinical setting. CompTMAP has gone through extensive testing to ensure accuracy and reliability. Physician surveys have indicated a positive response to CompTMAP, although the sample was insufficient for statistical testing. CompTMAP is part of a new era of comprehensive computerized decision support systems that take advantage of advances in automation and provide more complete clinical support to physicians in clinical practice. PMID- 15292538 TI - Partial compliance and risk of rehospitalization among California Medicaid patients with schizophrenia. AB - OBJECTIVE: The objective of this study was to evaluate the relationship between compliance with an antipsychotic medication regimen and risk of hospitalization in a cohort of California Medicaid patients with schizophrenia. METHODS: Compliance behavior was estimated by using a retrospective review of California Medicaid pharmacy refill and medical claims for 4,325 outpatients for whom antipsychotics were prescribed for treatment of schizophrenia from 1999 to 2001. Compliance behavior was estimated by using four different definitions: gaps in medication therapy, medication consistency and persistence, and a medication possession ratio. Patients were followed for one year and had an average of 19.1 dispensing events. Logistic regression models using each compliance estimate were used to determine the odds of hospitalization. RESULTS: Risk of hospitalization was significantly correlated with compliance. With all definitions, lower compliance was associated with a greater risk of hospitalization over and above any other risk factors for hospitalization. For example, the presence of any gap in medication coverage was associated with increased risk of hospitalization, including gaps as small as one to ten days (odds ratio [OR]=1.98). A gap of 11 to 30 days was associated with an OR of 2.81, and a gap of more than 30 days was associated with an OR of 3.96. CONCLUSIONS: This study showed a direct correlation between estimated partial compliance and hospitalization risk among patients with schizophrenia across a continuum of compliance behavior. PMID- 15292539 TI - A comparison of type 2 diabetes outcomes among persons with and without severe mental illnesses. AB - OBJECTIVE: Type 2 diabetes is an important comorbid medical condition associated with schizophrenia. The objective of this study was to compare glycosylated hemoglobin (HbA(1c)) levels of patients who had type 2 diabetes and schizophrenia with those of patients who had type 2 diabetes and major mood disorders and those who had type 2 diabetes but who did not have severe mental illness. METHODS: A sample of 300 patients with type 2 diabetes was recruited from community mental health centers in the greater Baltimore region and nearby primary care clinics. Of these, 100 had schizophrenia, 101 had a major mood disorder, and 99 had no identified severe mental illness. HbA(1c), the main outcome measure, was compared between the group with schizophrenia and the other two groups. RESULTS: All three groups had HbA(1c) values above recommended levels. HbA(1c) levels were significantly lower among patients with schizophrenia than among patients who did not have severe mental illness but were not significantly different from those of patients who had major mood disorders. Patients for whom olanzapine was prescribed had higher HbA(1c) levels than those for whom other antipsychotic agents were prescribed. CONCLUSIONS: All three groups of patients require improved diabetes treatment to achieve acceptable HbA(1c) levels. There may be previously unrecognized benefits for diabetes management among persons with severe mental illnesses who are receiving regular mental heath care, but these individuals may also have risk factors that can influence diabetes outcomes and HbA(1c) levels. PMID- 15292540 TI - Review of literature on aftercare services among children and adolescents. AB - OBJECTIVE: Psychiatric hospital lengths of stay have decreased for children and adolescents, in part because of the presumption that aftercare services in the community are effective and accessible. This review critically examines the literature that pertains to the rates of aftercare service use, the effectiveness of aftercare services, and predictors of aftercare service use. METHODS: Studies were selected on the basis of MEDLINE and PsychINFO computer searches, covering the period between January 1992 and August 2003. Reports that were selected (N=21) included data on outpatient aftercare service use among youths who were aged 18 years and younger and who were discharged from child and adolescent inpatient facilities. RESULTS AND DISCUSSION: A majority of youths received aftercare services after hospitalization, but many youths and families were not fully compliant with aftercare recommendations. Many youths and families continued to receive services up to three months after hospitalization. The literature documents only a small amount of evidence about the effectiveness of aftercare services, but the evidence suggested that aftercare services for youths with substance use problems may have beneficial effects. Few studies examined predictors of aftercare service use and discontinuation, but previous recent mental health service use and decreased family dysfunction appeared to be related to aftercare service use. PMID- 15292542 TI - Converting cultural capital among teen refugees and their families from Bosnia Herzegovina. AB - OBJECTIVES: The objective of this study was to identify the processes by which teen refugees adapt and apply cultural capital in conditions of refuge in order to develop preventive interventions for refugee youths. METHODS: The study was a multisite ethnographic study in Chicago that involved observation of Bosnian participants in schools, community sites, service organizations, and households as well as in-depth interviews with a subsample of 30 Bosnian adolescents and their families. Field notes and interview data were subjected to thematic analysis. RESULTS: The concept of converting cultural capital emerged as a useful construct for representing the cultural resources that Bosnian teen refugees and their families bring to the refugee trauma experience. Conversion of cultural capital refers to processes of adapting and applying the meanings, knowledge, customs, achievements, and outlooks that teen refugees and their families bring to new environments in order to enhance teens' cultural vitality and social incorporation. Nine mechanisms of converting cultural capital were identified, labeled, and defined in emic terms: using our language, obliging family, sticking together, returning to religion, going ghetto, building a future, taking pride in tradition, critiquing America, and seeking freedom. These mechanisms represent cultural strategies by which teen refugees attempt to manage enormous historical, social, cultural, economic, familial, and psychological changes associated with refugee trauma. CONCLUSIONS: Ethnography is an important methodologic tool in mental health services research, and the concept of converting cultural capital is useful in designing preventive interventions for teen refugees and their families. PMID- 15292541 TI - The development and implementation of case management for substance use disorders in North America and Europe. AB - Because of the multifaceted, chronic, and relapsing nature of substance use disorders, case management has been adapted to work with persons who have these disorders. Deliberate implementation has been identified as a powerful determinant of successful case management. This article focuses on six key questions about implementation of case management services on the basis of a comparison of experiences from the United States, the Netherlands, and Belgium. It was found that case management has been applied in various populations with substance use disorders, and distinct models have been associated with positive effects, such as increased treatment participation and retention, greater use of services, and beneficial drug-related outcomes. Program fidelity, robust implementation, extensive training and supervision, administrative support, a team approach, integration in a comprehensive network of services, and minimal continuity have all been linked to successful implementation. PMID- 15292543 TI - The cost of schizophrenia treatment in Taiwan. AB - The costs associated with mental illness in Taiwan have been the subject of discussion and concern in Taiwan's Bureau of National Health Insurance. The authors report the first estimates of these costs on the basis of national data for 52,432 patients treated in 1999. Total schizophrenia-related health care expenditure was estimated at 112.4 million dollars, which constituted 1.2 percent of national health care expenditures that year. The cost per outpatient visit was 57 dollars, the cost per admission was 1,123 dollars, and the annual average direct cost of treating a person with schizophrenia was 2,144 dollars. PMID- 15292544 TI - Concurrent psychiatric diagnoses by age and race among persons with bipolar disorder. AB - The authors characterized concurrent psychiatric diagnoses among patients with a diagnosis of bipolar disorder who were in routine care by using administrative data from a Department of Veterans Affairs facility. Of 813 patients who had a diagnosis of bipolar disorder in 2000, 21 percent were older (>/=60 years) whites, and 2 percent were older African Americans. Older African Americans were the most likely to have a diagnosis of schizophrenia documented in the medical record compared with younger African Americans, older whites, and younger whites (67 percent, 34 percent, 38 percent, and 27 percent, respectively). The results suggest that older African-American patients with bipolar disorder are more likely to receive diagnoses of mutually exclusive conditions, such as schizophrenia, and thus appear to have an elevated risk of their illness being underrecognized or misdiagnosed and receiving inappropriate treatment. PMID- 15292545 TI - Using conjoint analysis to assess depression treatment preferences among low income Latinos. AB - The authors examined the feasibility of conjoint analysis for measuring the depression treatment preferences of low-income, low-literacy Latino primary care patients. Forty-two patients with depression (58 percent of those eligible for the study) completed a survey about preferences for treatment and strategies to reduce barriers to care. They preferred combined counseling and medication to either approach alone and preferred individual over group treatment but did not show a significant preference for treatment setting. The odds of treatment acceptance were increased by the availability of telephone appointments, bus passes, and help with making appointments. Although further validation is required, conjoint analysis appears to be feasible for assessing preferences regarding depression treatment in this underserved population. PMID- 15292546 TI - Emergency department visits for depression in the United States. AB - The authors used data from the 1997-2000 National Hospital Ambulatory Care Surveys to quantify and characterize visits to emergency departments for depression. Each year there were 580,000 visits associated with a primary diagnosis of depression. More than half of these visits resulted in admission to a hospital or another facility. Twelve percent of visits involved a self inflicted injury. Antidepressants were offered during 18 percent of visits. Mental status examinations were given during 52 percent of visits. Overall, 81 percent of visits were by patients who had health insurance coverage. PMID- 15292548 TI - Community psychiatry education through homeless outreach. PMID- 15292547 TI - Use of topical application of lidocaine-prilocaine cream to reduce injection-site pain of depot antipsychotics. AB - This study took place in Israel and examined the use of a local topical anesthetic cream to ameliorate the pain at the injection site caused by depot antipsychotic medications. Fifteen consecutive outpatients who had schizophrenia and who were under treatment with depot antipsychotic medications were enrolled in this randomized, double-blind, placebo-controlled crossover study. The patients received either lidocaine-prilocaine cream or a placebo one hour before the injection. The degree of pain at the injection site was quantified by the patients' use of a visual analogue scale five minutes after the injection. The application of the lidocaine-prilocaine cream led to a significant reduction of pain compared with the placebo. PMID- 15292549 TI - A specialist early intervention for first-episode psychosis. PMID- 15292550 TI - Drug company representatives and psychiatrists. PMID- 15292551 TI - Articles should reflect psychiatry's contributions. PMID- 15292552 TI - Satisfaction with eGroups among persons with psychiatric disorders. PMID- 15292553 TI - Patient perspective on collaborative treatment. PMID- 15292554 TI - Choroid plexus cyst and echogenic intracardiac focus in women at low risk for chromosomal anomalies: the obligation to inform the mother. PMID- 15292555 TI - The history of breast ultrasound. AB - What began as a laboratory-based spin-off of military technology has matured over the past 50 years into an integral part of the breast imaging armamentarium. It has revolutionized the evaluation of breast abnormalities and has provided a rapid, cost-effective, and accurate guidance method for a wide range of interventional techniques. Subsequent improvements in technology will only serve to further enhance its pivotal clinical role. PMID- 15292556 TI - Are intracardiac echogenic foci markers of congenital heart disease in the fetus with chromosomal abnormalities? AB - OBJECTIVE: To determine whether intracardiac echogenic foci (ICEF) are markers of congenital heart disease (CHD) in fetuses with chromosomal abnormalities. METHODS: We identified all fetuses with chromosomal abnormalities undergoing targeted sonography at 17 weeks' to 21 weeks 6 days' gestation in a single perinatal center from January 1, 1994, to June 30, 2003. Offspring with and without CHD were compared for the presence or absence of ICEF. RESULTS: Two (8%) of 25 fetuses with ICEF had CHD versus 38 (33.3%) of 114 fetuses without ICEF (P = .006). Similarly, 1 (5.5%) of 18 fetuses with trisomy 21 and ICEF had CHD compared with 16 (37.2%) of 43 fetuses with trisomy 21 without ICEF (P = .009). CONCLUSIONS: Intracardiac echogenic foci in fetuses with chromosomal abnormalities, including those with trisomy 21, are not useful markers for CHD. PMID- 15292557 TI - Karyotyping of fetuses with isolated choroid plexus cysts is not justified in an unselected population. AB - OBJECTIVE: The main objective of this study was to determine whether isolated choroid plexus cysts are a risk factor for trisomy 18. METHODS: A total of 12,672 unselected patients were examined, and the outcome of fetuses with choroid plexus cysts was assessed. The cases with choroid plexus cysts were selected from the 12,672 patients and further divided into cases with minor markers of aneuploidy and cases with associated structural anomalies. Previous similar work was reviewed, analyzed, and, where possible, compared with the results of this study. RESULTS: The findings revealed 366 fetuses with choroid plexus cysts (2.9%). Thirty-three percent of fetuses with trisomy 18 had choroid plexus cysts, and, in every case, structural anomalies were also present. From the 12 cases with choroid plexus cysts in addition to major associated anomalies, amniocentesis revealed 2 cases of trisomy 18. Forty-three patients who had choroid plexus cysts and minor anomalies within our population had normal outcomes. CONCLUSIONS: These results indicate that the risk of amniocentesis is not acceptable if isolated choroid plexus cysts are isolated findings. More data are needed to establish whether choroid plexus cysts and other soft signs independently increase the risk of aneuploidy. PMID- 15292558 TI - Second-trimester sonographic comparison of the lower uterine segment in pregnant women with and without a previous cesarean delivery. AB - OBJECTIVE: To compare measurements of the lower uterine segment during a second trimester sonographic examination in women with and without a previous cesarean delivery. METHODS: Women undergoing second-trimester sonographic examination, 24 with a history of cesarean delivery and 30 control subjects with no history of cesarean delivery, were recruited for transvaginal sonographic evaluation of the lower uterine segment with a high-frequency probe. The uterine niche or previous cesarean scar site was defined as a small triangular anechoic defect in the anterior wall of the uterus. The uterine wall thickness was measured successively at the level where the bladder dome meets the lower uterine segment. Measurements were obtained with cursors at the interface of the urine-bladder and the amniotic fluid-decidua. The study was approved by the Institutional Review Board, and P < .05 was considered significant. RESULTS: The uterine niche was identified in 14 (58%) of 24 women with a previous cesarean delivery. The lower uterine segment was significantly thinner in women with a previous cesarean delivery compared with control subjects (mean +/- SD, 4.7 +/- 1.1 versus 6.6 +/- 2.0 mm; P < .001). In the previous cesarean group, the mean lower uterine segment thickness was similar in the 5 women with 2 cesarean deliveries when compared with those with 1 cesarean delivery (4.6 +/- 1.0 versus 4.7 +/- 1.4 mm; P = .91). In a linear regression model, the only variable retaining significance in the prediction of uterine wall thickness was previous cesarean delivery (P= .002). Maternal age, parity, number of previous cesarean deliveries, and gestational age did not attain significance in the model. CONCLUSIONS: The lower uterine segment during a second-trimester sonographic examination is significantly thinner in women with a previous cesarean delivery. Identification of the scar niche is possible in most of these women. PMID- 15292559 TI - Sonography in the diagnosis of cervical recurrence in patients with differentiated thyroid carcinoma. AB - OBJECTIVE: To determine the sensitivity of thyroglobulin (Tg), iodine scanning, and sonography in the diagnosis of cervical recurrence of thyroid cancer. METHODS: This prospective study assessed 81 patients with cervical metastases or extrathyroid invasion at first appearance who underwent clinical examination, scanning, measurement of Tg after thyroxine withdrawal, and sonography about 8 months after thyroidectomy followed by radioiodine treatment. Only patients without distant metastases and without anti-Tg antibodies were included. RESULTS: Fifty patients showed persistence of the disease in the cervical region, with only 16% of them having had a suspicion on clinical examination, 33 with Tg levels of 2 ng/mL or greater (66% sensitivity), and 29 with positive scan findings (58% sensitivity). A combination of the 2 methods detected disease in 40 (80%) of 50 patients but failed to show 20% of cases that were identified by sonography and confirmed by fine-needle aspiration. Sonography had sensitivity of 96%. Specificity values for Tg, iodine scanning, and sonography were 80.6%, 90.3%, and 87%, respectively. CONCLUSIONS: Classic follow-up methods may not detect cervical disease in some patients with differentiated thyroid carcinoma, and sonography is necessary even in patients apparently free of the disease. PMID- 15292560 TI - In vitro sonographic evaluation of sentinel lymph nodes for detecting metastasis in breast cancer: comparison with histopathologic results. AB - OBJECTIVE: To assess the accuracy and the effectiveness of in vitro sonography of sentinel lymph nodes (LNs) for differentiating benignity from metastasis during breast cancer operations. METHODS: Two hundred twenty-two fresh sentinel LNs obtained during surgery were examined by in vitro sonography to detect metastasis in 47 patients with breast cancer. Sonographic criteria of malignant LNs were defined as uneven cortical thickness of greater than 3 mm in depth, round hypoechoic nodes, and the absence of an echogenic hilum. Histopathologic results were correlated with those obtained by the sonographic study. Statistical analysis was performed with a kappa value to evaluate the accuracy of in vitro sonography of sentinel LNs. RESULTS: On sonographic examination, 190 (85.6%) of the 222 sentinel LNs were benign. Among the 190 benign LNs, in only 8 (4.2%) was the presence of metastasis pathologically proved, which indicated focal micrometastasis in the cortex. Thirty-two sentinel LNs were sonographically positive. In 19 (59.4%) of the 32 sentinel LNs, metastasis was pathologically confirmed. The remaining 13 sonographically positive LNs (40.6%) did not reveal any histologically malignant cells. The sensitivity, specificity, positive predictive value, negative predictive value, and accuracy were 70.4%, 93.3%, 59.4%, 95.7%, and 90.5%, respectively. CONCLUSIONS: The in vitro sonographic evaluation of the sentinel LNs during breast cancer operations may be a reliable method of predicting nodal metastasis and may be helpful in deciding the extent of LN dissection. PMID- 15292561 TI - Doppler sonography of renal obstruction: value of venous impedance index measurements. AB - OBJECTIVE: Arterial resistive index values have poor sensitivity and specificity for alterations in renal perfusion induced by collecting system obstruction. We aimed to determine whether the intrarenal venous impedance index values could be useful in evaluating renal parenchymal compliance in cases of obstruction and in differentiating acute obstruction from chronic cases. METHODS: Fifteen patients with acute renal colic having unilateral stone disease and another 15 patients having unilateral chronic obstruction due to various causes were evaluated sonographically. The diagnosis was confirmed either by computed tomography or intravenous urography in all cases. Fifteen subjects with normal kidneys were investigated as a control group. All patients were examined prospectively by conventional and Doppler sonography. The impedance indices and peak flow signals of the interlobar arteries and veins of both kidneys were calculated from spectral Doppler waveforms in all 3 groups. RESULTS: The mean venous impedance index on the acutely obstructed side was lower than the index on the unobstructed side: 0.25 +/- 0.07 and 0.53 +/- 0.3 (mean +/- SD), respectively (P = .005). The mean venous impedance index on the acutely obstructed side was less than the indices both on the chronically obstructed side and in the control subjects (P > .0001). In acute cases, the mean arterial resistive index on the obstructed side was higher than the index on the unobstructed side: 0.62 +/- 0.06 and 0.57 +/- 0.06, respectively (P = .001). No statistically significant difference was detected between other parameters evaluated for the test and control groups. CONCLUSIONS: Renal venous impedance index values may be helpful in evaluating renal hemodynamics in obstruction and in differentiating acute obstruction from chronic cases when used in conjunction with the arterial resistive index. PMID- 15292562 TI - Sonographic and color doppler findings in aortoarteritis (Takayasu arteritis). AB - OBJECTIVE: To review the sonographic appearance of aortoarteritis. METHODS: A pictorial review of cases is presented. RESULTS: Sonography in conjunction with color and pulsed Doppler imaging is a valuable tool in the evaluation of aortoarteritis. We can accurately diagnose, grade, and follow the progress of the disease. The response to treatment can also be assessed. This presentation reviews the sonographic findings in aortoarteritis. CONCLUSIONS: Color-coded Doppler sonography can facilitate an accurate diagnosis of Takayasu arteritis by the characteristic appearance. Associated organ involvement can also be assessed. PMID- 15292563 TI - Prenatal diagnosis of isolated tricuspid valve atresia: report of 4 cases and review of the literature. AB - OBJECTIVE: To describe the prenatal features of fetal tricuspid atresia. METHODS: Four cases of fetal tricuspid atresia were prenatally diagnosed, sonographically described, and followed. RESULTS: On the basis of this small series, the key findings for diagnosis included the demonstration of no patent tricuspid valve on the 4-chamber view, no flow across the tricuspid valve on pulsed or color Doppler flow mapping, small right ventricles, and associated interventricular septal defects. Increased nuchal translucency thickness may give the first clue leading to follow-up scans, resulting in a definite diagnosis. CONCLUSIONS: Tricuspid atresia can be readily diagnosed prenatally. The key findings and differential diagnoses are provided. PMID- 15292564 TI - Polyorchidism: report of 3 cases and review of the literature. AB - OBJECTIVE: To describe the embryologic, clinical, sonographic, and magnetic resonance imaging features of polyorchidism and to review the literature on similar cases. METHODS: Over a 5-year period, we encountered 3 patients who were found to have polyorchidism on scrotal sonography. All 3 patients had a painless scrotal mass. Two patients also had magnetic resonance imaging of the scrotum, and the results were correlated with the sonograms. We also performed a literature search for other reports of polyorchidism. RESULTS: One patient had 2 right testicles and a single left testicle. The second patient had 3 left testicles and 1 right testicle. In the third patient, who had 2 left testicles and 1 right testicle, microlithiasis was found in all 3 testes. The supernumerary testes were within the scrotum in all cases. All testicles were identified by sonography. Magnetic resonance imaging in 2 cases provided confirmatory data regarding the presence of an extra testicle but did not add other relevant information. Conservative treatment was chosen in all cases. CONCLUSIONS: Polyorchidism is a rare congenital anomaly. There are characteristic sonographic features of polyorchidism, and the diagnosis is often made on the basis of sonography. Magnetic resonance imaging can be used for confirmation but may be more helpful in cases complicated by cryptorchism or neoplasia. Conservative treatment is advised in uncomplicated cases. PMID- 15292565 TI - Leydig cell tumors of the testis: gray scale and color Doppler sonographic appearance. AB - OBJECTIVE: To determine the gray scale and color Doppler sonographic features of Leydig cell tumors of the testis in a series of patients. METHODS: We retrospectively analyzed the sonographic appearance of 10 proven Leydig cell tumors in 9 patients aged 26 to 47 years. Sonographic features that were reviewed included the size and echogenicity of the tumors, presence of cystic areas or calcifications, and distribution pattern of detectable blood flow on color or power Doppler imaging. RESULTS: The tumors ranged from 0.4 to 3.0 cm in diameter, but most were less than 1.0 cm in diameter. In 1 testis, 2 discrete Leydig cell tumors were found. Nine (90%) of the 10 tumors were homogeneously hypoechoic. Only 1 tumor was isoechoic with the testis. None of the tumors contained calcifications. Of 8 tumors with color Doppler imaging, 7 (88%) showed a characteristic pattern of increased peripheral blood flow, which was either circumferential or punctate. Only 1 tumor was found with internal hypervascularity. CONCLUSIONS: Peripheral hypervascularity in a hypoechoic testicular tumor that has little or no internal color Doppler flow should suggest the possibility of a Leydig cell tumor, and consideration should be given to testicle-sparing surgery. PMID- 15292566 TI - Postnephrectomy renal arteriovenous fistula. PMID- 15292567 TI - Sonographic diagnosis of a glass foreign body in the urinary bladder. PMID- 15292568 TI - Sonographic features of spermatic cord leiomyosarcoma. PMID- 15292569 TI - Sonographic appearance of testicular adrenal rest tissue in congenital adrenal hyperplasia. PMID- 15292570 TI - Holt-Oram syndrome: contribution of prenatal 3-dimensional sonography in an index case. PMID- 15292571 TI - Gastric outlet obstruction secondary to linitis plastica of the stomach as shown on transabdominal sonography. PMID- 15292572 TI - Hemorrhagic cholecystitis simulating gallbladder carcinoma. PMID- 15292573 TI - Gastroduodenal artery pseudoaneurysm secondary to pancreatic head biopsy. PMID- 15292574 TI - Ultrasonographic appearance of idiopathic radial nerve constriction proximal to the elbow. PMID- 15292575 TI - An isolated echogenic heart focus is a low-level risk marker for Down syndrome. PMID- 15292576 TI - Proteomics: The Next Frontier-A Biotech Perspective. PMID- 15292577 TI - The Developing Country Reactions to Biomedical Techniques and Plant Biotechnology: The Tunisian Experience. AB - In the present study we present the conditions offered to biotechnology development in Tunisia and we compare three main biotechnology applications which raise ethical and health problems: organ transplant, assisted reproductive techniques, and genetically modified organisms. We try to identify factors that have allowed success of the first two applications and failure of the latter. Conditions offered to biotechnology in other African countries are also discussed. PMID- 15292578 TI - A Gene Encoding Sialic-Acid-Specific 9-O-Acetylesterase Found in Human Adult Testis. AB - Using differential display RT-PCR, we identified a gene of 2750 bp from human adult testis, named H-Lse, which encoded a putative protein of 523 amino acids and molecular weight of 58 kd with structural characteristics similar to that of mouse lysosome sialic-acid-specific 9-O-acetylesterase. Northern blot analysis showed a widespread distribution of H-Lse in various human tissues with high expression in the testis, prostate, and colon. In situ hybridization results showed that while H-Lse was not detected in embryonic testis, positive signals were found in spermatocytes but not spermatogonia in adult testis of human. The subcellular localization of H-Lse was visualized by green fluorescent protein (GFP) fused to the amino terminus of H-Lse, showing compartmentalization of H-Lse in large dense-core vesicles, presumably lysosomes, in the cytoplasm. The developmentally regulated and spermatogenic stage-specific expression of H-Lse suggests its possible involvement in the development of the testis and/or differentiation of germ cells. PMID- 15292579 TI - $3,3',4,4',5$ -Pentachlorobiphenyl Inhibits Drug Efflux Through P-Glycoprotein in KB-3 Cells Expressing Mutant Human P-Glycoprotein. AB - The effects on the drug efflux of $3,3',4,4',5$ -pentachlorobiphenyl (PCB-126), the most toxic of all coplanar polychlorinated biphenyls (Co-PCBs), were examined in KB-3 cells expressing human wild-type and mutant P-glycoprotein in which the 61st amino acid was substituted for serine or phenylalanine ( ${?text{KB3 - Phe}};{61} $ ). In the cells expressing P-glycoproteins, accumulations of vinblastine and colchicine decreased form 85% to 92% and from 62% to 91%, respectively, and the drug tolerances for these chemicals were increased. In ${?text{KB3 - Phe}};{61} $, the decreases in drug accumulation were inhibited by adding PCB-126 in a way similar to that with cyclosporine A: by adding 1 $?mu$ M PCB-126, the accumulations of vinblastine and colchicine increased up to 3.3- and 2.3-fold, respectively. It is suggested that PCB-126 decreased the drug efflux by inhibiting the P-glycoprotein in ${?text{KB3 - Phe}};{61} $. Since there were various P-glycoproteins and many congeners of Co-PCBs, this inhibition has to be considered a new cause of the toxic effects of Co-PCBs. PMID- 15292580 TI - ELISA for Determination of Human Growth Hormone: Recognition of Helix 4 Epitopes. AB - Human growth hormone (hGH) signal transduction initiates with a receptor dimerization in which one molecule binds to the receptor through sites 1 and 2. A sandwich enzyme-linked immunosorbent assay was developed for quantifying hGH molecules that present helix 4 from binding site 1. For this, horse anti-rhGH antibodies were eluted by an immunoaffinity column constituted by sepharose-rhGH. These antibodies were purified through a second column with synthetic peptide correspondent to hGH helix 4, immobilized to sepharose, and used as capture antibodies. Those that did not recognize synthetic peptide were used as a marker antibody. The working range was of 1.95 to 31.25 ng/mL of hGH. The intra-assay coefficient of variation (CV) was between 4.53% and 6.33%, while the interassay CV was between 6.00% and 8.27%. The recovery range was between 96.0% to 103.8%. There was no cross-reactivity with human prolactin. These features show that our assay is an efficient method for the determination of hGH. PMID- 15292581 TI - A Simple Chemical Method for Rendering Wild-Type Yeast Permeable to Brefeldin A That Does Not Require the Presence of an erg6 Mutation. AB - The present work aims to develop a growth medium to render a wild-type strain of Saccharomyces cerevisiae permeable to the antifungal drug Brefeldin A. In the current study, a synthetic medium containing 0.1% L-proline and supplemented with $3.0?times 10;{-3}$ % SDS is employed. When Brefeldin A is added to this medium, a wild-type strain shows increased growth sensitivity and a diminished transport of the amino acid L-leucine. Since Brefeldin A exerts its effect on the endoplasmic reticulum and the Golgi apparatus, the medium permits the study of the drug effect on the intracellular traffic of L-leucine permeases. PMID- 15292582 TI - Role of Peroxisome Proliferator-Activated Receptors in Inflammation Control. AB - Peroxisome proliferator-activated receptors (PPARs) were discovered over a decade ago, and were classified as orphan members of the nuclear receptor superfamily. To date, three PPAR subtypes have been discovered and characterized (PPAR $?alpha$, $?beta/?delta$, $?gamma$ ). Different PPAR subtypes have been shown to play crucial roles in important diseases and conditions such as obesity, diabetes, atherosclerosis, cancer, and fertility. Among the most studied roles of PPARs is their involvement in inflammatory processes. Numerous studies have revealed that agonists of PPAR $?alpha$ and PPAR $?gamma$ exert anti-inflammatory effects both in vitro and in vivo. Using the carrageenan-induced paw edema model of inflammation, a recent study in our laboratories showed that these agonists hinder the initiation phase, but not the late phase of the inflammatory process. Furthermore, in the same experimental model, we recently also observed that activation of PPAR $?delta$ exerted an anti-inflammatory effect. Despite the fact that exclusive dependence of these effects on PPARs has been questioned, the bulk of evidence suggests that all three PPAR subtypes, PPAR $?alpha, ?delta, ?gamma$, play a significant role in controlling inflammatory responses. Whether these subtypes act via a common mechanism or are independent of each other remains to be elucidated. However, due to the intensity of research efforts in this area, it is anticipated that these efforts will result in the development of PPAR ligands as therapeutic agents for the treatment of inflammatory diseases. PMID- 15292583 TI - Host Cell Phenotypic Variability Induced by Trypanosomatid-Parasite-Released Immunomodulatory Factors: Physiopathological Implications. AB - The parasitic protozoa Trypanosoma cruzi and Leishmania sp release a variety of molecules into their mammalian hosts (ESA: excretory-secretory products). The effects of these ESA on the host cell function may participate in the establishment of a successful infection, in which the parasite persists for a sufficient period of time to complete its life cycle. A number of regulatory components or processes originating from the parasite that control or regulate the metabolism and the growth of host cell have been identified. The purpose of the present review is to analyze some of the current data related to the parasite ESA that interfere with the host cell physiology. Special attention is given to members of conserved protein families demonstrating remarkable diversity and plasticity of function (ie, glutathione S-transferases and related molecules; members of the trans-sialidase and mucin family; and members of the ribosomal protein family). The identification of parasite target molecules and the elucidation of their mode of action toward the host cell represents a step forward in efforts aimed at an immunotherapeutic or pharmacological control of parasitic infection. PMID- 15292584 TI - The B chromosome database. AB - The database is compiled from the world literature on B chromosomes published between 1906 and 1994, and has 3,484 records. A brief description is given of the history and structure of the database, which runs in Microsoft ACCESS. A downloadable version is available at http://www.bchromosomes.org/bdb/. PMID- 15292585 TI - The distribution of B chromosomes across species. AB - In this review we look at the broad picture of how B chromosomes are distributed across a wide range of species. We review recent studies of the factors associated with the presence of Bs across species, and provide new analyses with updated data and additional variables. The major obstacle facing comparative studies of B chromosome distribution is variation among species in the intensity of cytogenetic study. Because Bs are, by definition, not present in all individuals of a species, they may often be overlooked in species that are rarely studied. We give examples of corrections for differences in study effort, and show that after a variety of such corrections, strong correlations remain. Several major biological factors are associated with the presence of B chromosomes. Among flowering plants, Bs are more likely to occur in outcrossing than in inbred species, and their presence is also positively correlated with genome size and negatively with chromosome number. They are no more frequent in polyploids than in diploids, nor in species with multiple ploidies. Among mammals, Bs are more likely to occur in species with karyotypes consisting of mostly acrocentric chromosomes. We find no evidence for an association with chromosome number or genome size in mammals, although the sample for genome size is small. The associations with breeding system and acrocentric chromosomes were both predicted in advance, but those with genome size and chromosome number were discovered empirically and we can offer only tentative explanations for the very strong associations we have uncovered. Our understanding of why B chromosomes are present in some species and absent in others is still in its infancy, and we suggest several potential avenues for future research. PMID- 15292586 TI - Are the dot-like chromosomes in Trinomys iheringi (Rodentia, Echimyidae) B chromosomes? AB - In this article we review the existing cytogenetic information on the polymorphic dot-like chromosomes in Trinomys iheringi, the only species in the family Echimyidae harboring them, and provide new data on the frequency, banding properties, meiotic behavior and DNA composition of these minute chromosomes. Since no individuals lacking these chromosomes have hitherto been found, one of the main properties of B chromosomes, i.e. dispensability, has not yet been tested, so that some reasonable doubt might exist on whether they are true B chromosomes. The dot-like chromosomes were also present in the twelve new individuals analyzed, showed intraindividual variation in number, most likely due to mitotic instability during development, failed to show C-bands, showed late replication, paired among them in meiosis, but not with the large chromosomes, and appeared to be mainly composed of telomeric DNA. These results suggest that these dot-like chromosomes might actually be mitotically unstable micro B chromosomes showing very high frequency in the natural populations thus far analyzed. But, to be confident of this conclusion, individuals lacking the dot like chromosomes should actively be searched in future research to test their dispensability. PMID- 15292587 TI - Human supernumeraries: are they B chromosomes? AB - In humans, the presence of supernumerary chromosomes is an unusual phenomenon, which is often associated with developmental abnormalities and malformations. In contrast to most animal and plant species, the extensive knowledge of the human genome and the ample set of molecular and cytogenetic tools available have permitted to ascertain not only that most human supernumerary chromosomes (HSCs) derive from the A chromosome set, but also the specific A chromosome from which most of them arose. These extra chromosomes are classified into six types on the basis of morphology and size. There are both heterochromatic and euchromatic HSCs, the latter being more detrimental. Most are mitotically stable, except some producing individual mosaicism. No information is available on the HSC transmission rate since extensive familial studies are not usually performed generally because of death of the relatives or lack of cooperation. The main B chromosome property failing in HSCs seems to be their population spread as polymorphisms, since most HSCs seem to correspond to extra A chromosomes or centric fragments spontaneously arisen in the analysed individual or one of his/her parents. However, we cannot rule out at this moment, that more intensive studies on population distribution and frequency of those HSCs most closely resembling B chromosomes (i.e. those heterochromatic and thus less detrimental) would reveal possible HSCs polymorphisms. Although HSCs cannot be considered B chromosomes, some of them might be a source for future B chromosomes. The best candidates would be heterochromatic HSCs, which might manage to drive in either sex. To ascertain this possibility, research on inheritance and population studies would be very helpful in combination with the powerful cytogenetic and molecular tools available for our species. PMID- 15292588 TI - Is the aneuploid chromosome in an apomictic Boechera holboellii a genuine B chromosome? AB - The Boechera holboellii complex comprises B. holboellii and B. drummondii, both of which can reproduce through sex or apomixis. Sexuality is associated with diploidy, whereas apomictic individuals can either be diploid, aneuploid or triploid. Aneuploid individuals are found in geographically and genetically distinct populations and contain a single extra chromosome. It is unknown whether the supernumerary chromosomes are shared by common descent (single origin) or have originated via introgressive hybridizations associated with the repeated transition from diploidy to triploidy. Diploid plants containing the extra chromosome(s) reproduce apomictically, suggesting that the supernumerary elements are associated with apomixis. In this study we compared flow cytometry data, chromosome morphology, and DNA sequences of sexual diploid and apomictic aneuploids in order to establish whether the extra chromosome fits the classical concept of a B chromosome. Karyotype analyses revealed that the supernumerary chromosome in the metaphase complement is heterochromatic and often smaller than the A chromosomes, and differs in length between apomictic plants from different populations. DNA sequence analyses furthermore demonstrated elevated levels of non-synonymous substitutions in one of the alleles, likely that on the aneuploid chromosome. Although the extra chromosome in apomictic Boechera does not go through normal reductional meiosis, in which it may get eliminated or accumulated by a B-chromosome-specific process, its variable size and heterochromatic nature does meet the remaining criteria for a genuine B chromosome in other species. Its prevalence and conserved genetic composition nonetheless implies that this chromosome, if truly a B, may be atypical with respect to its influence on its carriers. PMID- 15292589 TI - The occurrence of different Bs in Cestrum intermedium and C. strigilatum (Solanaceae) evidenced by chromosome banding. AB - In this study, we examine the morphology, mitotic stability, meiotic behavior and the composition of heterochromatin of B chromosomes in Cestrum intermedium and C. strigilatum. The results showed that B chromosome number shows intraindividual variation in the root meristem, which seems to lead to a slight rate of B elimination in this somatic tissue. B chromosomes in both species were similar in size and shape, but differed with regard to the type, size and distribution of heterochromatin. Possible evolutionary pathways for B chromosome origin in Cestrum are discussed. PMID- 15292590 TI - Distribution and stability of supernumerary microchromosomes in natural populations of the Amazon molly, Poecilia formosa. AB - In animals, supernumerary chromosomes and their evolution have mostly been studied in sexual reproducing species. In the present study, for the first time, the natural distribution and stability of supernumerary microchromosomes were investigated in the unisexual fish species Poecilia formosa. Natural habitats throughout the range of P. formosa were screened for the presence of microchromosomes over several years. A high frequency of microchromosomes was found in the Rio Purificacion river system. Evidence points to the presence of the same microchromosome lineage over many generations. No supernumerary chromosomes were found elsewhere than in the Rio Purificacion representing a significant difference in the distribution of microchromosome-bearing individuals between the Rio Purificacion and all other collection sites. PMID- 15292591 TI - B chromosomes in Amazonian cichlid species. AB - B chromosomes are reported in three different Amazonian cichlid species. One to three supernumerary microchromosomes were detected in the peacock bass Cichla monoculus (4 out of 28 specimens) and Cichla sp. (4 out of 13 specimens), and pike cichlids Crenicichla reticulata (2 out of 5 specimens), with no similar standard chromosomal morphology. C-banding revealed that B chromosomes are totally heterochromatic. We suggest two scenarios for the origin of these B chromosomes either by chromosomal breakdowns due to mutagenic action of methyl mercury present in the aquatic environment or by interspecific origin due to hybridization events. PMID- 15292592 TI - The B chromosomes in Brachycome. AB - This review presents a historical account of studies of B chromosomes in the genus Brachycome Cass. (synonym: Brachyscome) from the earliest cytological investigations carried out in the late 1960s though to the most recent molecular analyses. Molecular analyses provide insights into the origin and evolution of the B chromosomes (Bs) of Brachycome dichromosomatica, a species which has Bs of two different sizes. The larger Bs are somatically stable whereas the smaller, or micro, Bs are somatically unstable. Both B types contain clusters of ribosomal RNA genes that have been shown unequivocally to be inactive in the case of the larger Bs. The large Bs carry a family of tandem repeat sequences (Bd49) that are located mainly at the centromere. Multiple copies of sequences related to this repeat are present on the A chromosomes (As) of related species, whereas only a few copies exist in the A chromosomes of B. dichromosomatica. The micro Bs share DNA sequences with the As and the larger Bs, and they also have B-specific repeats (Bdm29 and Bdm54). In some cases repeat sequences on the micro Bs have been shown to occur as clusters on the A chromosomes in a proportion of individuals within a population. It is clear that none of these B types originated by simple excision of segments from the A chromosomes. PMID- 15292593 TI - B chromosomes in Sternorrhyncha (Hemiptera, Insecta). AB - In the hemipteroid insects of the suborder Sternorrhyncha, B chromosomes are relatively common in comparison with other suborders of Hemiptera. However, the occurrence of supernumerary chromosomes is restricted, in most cases, to several genera or closely related species. At least in some species of Psylloidea with the XY sex determination system, a mitotically stable B chromosome integrated into an achiasmatic segregation system with the X, and became fixed as a Y chromosome. In some Aphidoidea with a multiple X system of sex determination, B chromosomes appear to be in fact non-functional X chromosomes. Supernumerary chromosomes thus probably play an important role in the evolution of sex determination systems in Sternorrhyncha. PMID- 15292594 TI - B chromosomes in Crustacea Decapoda. AB - Among crustacean Decapoda numerical chromosome variability is frequent, and it has been hypothesized that the presence of supernumerary chromosomes accounts for this variability. Thanks to the improvement of cytogenetic analysis by chromosomal banding techniques, supernumerary B chromosomes (Bs) have been demonstrated in Nephrops norvegicus, Homarus americanus,Palinurus elephas and P. mauritanicus, belonging to different crustacean families. In all four species Bs were variable in number, mainly heterochromatic and undigested by various endonucleases, and in meiosis they showed non-Mendelian segregation. Compared to the other chromosomes of the complement, the Bs are very small in almost all species, but some of them were very large in N. norvegicus. PMID- 15292595 TI - Current knowledge on B chromosomes in natural populations of helminth parasites: a review. AB - Helminths, traditionally classified into three phyla Platyhelminthes, Nemathelminthes and Acanthocephala, are a phylogenetically broadly diversified group of invertebrates, characterised by a parasitic life style. Current estimates of the helminth species diversity are at least 23-40,000 platyhelminthes, 10-26,000 nematodes and 1,200 acanthocephalans. Recent information on helminth karyotypes is fragmentary, and basic karyological data are known from approximately 1.1% of known species. Supernumerary chromosomes have been reported in selected populations of only 11 digenean flukes (Platyhelminthes), 1 thorny-headed worm (Acanthocephala) and 4 roundworms (Nematoda), which represent 3.6, 7.7 and 1.3% of the total number of species cytogenetically analysed to date within respective helminth groups. B chromosome presence was not generally associated with heteromorphic sex chromosomes as they occurred both in hermaphroditic flukes and dioecious helminths, and in species having male or female heterogametic sex chromosomes (ZW of schistosomes, XO of acanthocephalans and XY of nematodes). Numbers of B chromosomes varied from 1 to 10. Most often, Bs represented one or two of the smallest elements of the complement but they could be much bigger in some digenean flukes. B chromosomes showed a diverse morphology, including telocentric to metacentric structure. There is no detailed banding or ultrastructural study of Bs in the majority of helminth carriers. Assumptions on the possible relation between the occurrence of Bs in endoparasitic helminths and extreme environments are discussed. PMID- 15292596 TI - B chromosomes in the fish Astyanax scabripinnis (Characidae, Tetragonopterinae): an overview in natural populations. AB - Astyanax scabripinnis, a small neotropical freshwater fish, is a headwater species living in small tributaries of many Brazilian rivers, where they form isolated populations. This species harbors a B chromosome system in several populations. Among the several kinds of Bs reported in this species, the B(M) variant, a large metacentric of a similar size to the largest A chromosome, is the most widespread in natural populations. It probably corresponds to the ancestral B type in this species and a very similar B chromosome is also found in other Astyanax species. Strong evidence suggests that this B is an isochromosome showing structural and functional homology between its two arms, as shown by satellite DNA localization and the formation of a ring B univalent during meiosis. The B(SM) and B(m) variants, a large submetacentric and a small metacentric, respectively, represent rare variants and may be derived from structural rearrangements of the B(M) chromosome. In addition, B microchromosomes (B(micro)) were found in some populations. Frequency analyses in mountain populations have shown that B chromosomes are found in populations located at high altitude, but are absent in populations at low altitude, which is consistent with their parasitic nature, given the ecological peculiarities of both kinds of populations. PMID- 15292597 TI - Structure and evolution of B chromosomes in amphibians. AB - B chromosomes are known from 26 species of salamanders and frogs, equivalent to about 2% of amphibian species that have been karyotyped. In most cases, the structure of amphibian B chromosomes has not been extensively investigated. The exceptions are the B chromosomes of Hochstetter's frog, Leiopelma hochstetteri, from New Zealand, and the Coastal Giant salamander, Dicamptodon tenebrosus, from North America. Dicamptodon tenebrosus carries from 0 to 10 non-heterochromatic, telocentric B chromosomes per individual, averaging 0 to 3.4 B chromosomes per individual in populations throughout its extensive range. The B chromosomes of L. hochstetteri occur in frequencies averaging from 0 to 11.4 per individual in different populations, with a known maximum of 15 B chromosomes. Amphibian B chromosomes vary in size, heterochromatin, occurrence and frequency. They are commensurate in size and structure with the rest of the A set of chromosomes of the same species in which they occur. The B chromosomes are at least partly composed of repetitive DNA sequences which exist in numerous copies throughout the autosomes, in conformity to an hypothesis of intraspecific B chromosome origins. PMID- 15292598 TI - Occurrence of B chromosomes in lizards: a review. AB - Although B chromosomes have been reported in many species of plants and animals, few studies have revealed the presence of these extra chromosomes in lizards. B chromosomes of lizards show different morphologies and sizes, from microchromosomes to macrochromosomes, or elements of intermediate size between smaller and larger A chromosomes, and number variability at intra- and inter individual levels. In most cases, they are late-replicating and show either heterochromatic or no distinctive patterns after C-banding. The great majority of the publications about supernumerary chromosomes in this group have been based on conventional staining analyses, and there is no study designed to address questions related to their composition and structure or origin and evolution. PMID- 15292599 TI - B chromosomes in populations of mammals. AB - B chromosomes (Bs) have been found in 55 out of 4629 living species of mammals. The summarized data show great variability in types of mammalian Bs, including differences in size, shape and molecular composition. This variability extends to the origin, mode of transmission and population dynamics. In general, B chromosomes in mammals do not differ from Bs found in other animal or plant species, but some peculiarities do exist. Most species in which Bs are found are widespread. Some data support the view that Bs may contribute to the successful expansion of some of these species, but it is possible that Bs are just more easily scored in them due to their frequent occurrence. Most of these species are also characterized by cycling fluctuations of abundance and characteristic social organization that produce conditions favorable for Bs to spread. All areas of research on Bs in mammals suffer from lack of data, emphasizing the necessity for intensified research on the molecular structure and ways of maintenance of Bs in populations. PMID- 15292600 TI - B chromosomes in Brazilian rodents. AB - B chromosomes are now known in eight Brazilian rodent species: Akodon montensis, Holochilus brasiliensis, Nectomys rattus, N. squamipes, Oligoryzomys flavescens, Oryzomys angouya, Proechimys sp. 2 and Trinomys iheringi. Typically these chromosomes are heterogeneous relative to size, morphology, banding patterns, presence/absence of NORs, and presence/absence of interstitial telomeric signals after FISH. In most cases, Bs are heterochromatic and late replicating. Active NORs were detected in two species: Akodon montensis and Oryzomys angouya. As a rule, Bs behave as uni or bivalents in meiosis, there is no pairing between Bs and autosomes or sex chromosomes and also their synaptonemal complexes are isopycnotic with those in A chromosomes. PMID- 15292601 TI - The mammalian model for population studies of B chromosomes: the wood mouse (Apodemus). AB - The presence of B chromosomes was reported in six species of the genus Apodemus (A. peninsulae, A. agrarius, A. sylvaticus, A. flavicollis, A. mystacinus, A. argenteus). High frequencies of Bs were recorded particularly in A. peninsulae and A. flavicollis. The origin of Bs in Apodemus seems to be rather ancient, and it is possible that the supernumerary elements, and/or a tendency for their appearance, were inherited from the common ancestor of the extant species. We have not found any correlated changes between frequencies of Bs and the level of protein polymorphism and/or heterozygosity assessed in electrophoretic studies. No measurable effect of Bs on overall genetic variability was thus revealed in studied populations. The pattern of evolutionary dynamics of Bs can be distinctly different between geographical populations, and both the parasitic and the heterotic models can be applied to explain the maintenance of Bs in different populations. Further studies are desirable to improve our understanding of the complicated evolutionary dynamics of Bs in the Apodemus species. An essential condition for success in this respect is much more detailed information on inheritance and the molecular structure of Bs. PMID- 15292602 TI - A complex B chromosome system in the Korean field mouse, Apodemus peninsulae. AB - Information on B chromosomes of six subspecies of A. peninsulae Thomas, 1906, from 79 local populations of Russia (Siberia, Altai, Buryatia and the Far East), Mongolia, China, Korea and Japan (Hokkaido) is reviewed. The frequency of animals with B chromosomes is higher in this taxon than in other mammals and ranges from 0.4 up to 1.0, excluding two insular populations (Sakhalin Island and Stenin Island, Primorye) where Bs were not found. The B chromosome polymorphism shows four levels of variation in number (intraindividual mosaicism, intrapopulational and interpopulational), as well as variability in size, morphology and differential staining. Geographic variation was found among populations for these cytogenetic characteristics and, in some cases, it coincided with subspecies distribution. Comparative chromosome banding of micro and macro Bs illuminates possible pathways for their origin. PMID- 15292603 TI - A RAPD marker associated with B chromosomes in Partamona helleri (Hymenoptera, Apidae). AB - The hymenopteran Partamona helleri is found in southwestern Brazil in the Mata Atlantica from the north of the state of Santa Catarina until the south of Bahia. This work shows that P. helleri can carry up to four B chromosomes per individual. In order to obtain more information about P. helleri B chromosomes, the RAPD technique was used to detect DNA fragments associated with these chromosomes. The results showed that the RAPD technique is useful to detect specific sequences associated with B chromosomes. One RAPD marker was identified, cloned and used as probe in a DNA blot analysis. This RAPD marker hybridized with sequences present only in individuals containing B chromosomes. PMID- 15292604 TI - Comparative FISH analysis of distribution of B chromosome repetitive DNA in A an d B chromosomes in two subspecies of Podisma sapporensis (Orthoptera, Acrididae). AB - FISH analysis of B chromosome repetitive DNA distribution in A and B chromosomes of two subspecies of Podisma sapporensis (P. s. sapporensis and P. s. krylonensis) was performed. In the B chromosomes, C-positive regions contained homologous DNA repeats present also in some C-positive A chromosome regions. Most C-negative regions contained DNA repeats characteristic of A chromosome euchromatic regions. The two subspecies analyzed differed in the location of A chromosome regions enriched with repeats homologous to repeats of B chromosomes. The only common region enriched with these B chromosome repeats in both subspecies was the X chromosome pericentromeric region. The origin of B chromosomes in P. sapporensis is discussed. PMID- 15292605 TI - Comparative analysis of micro and macro B chromosomes in the Korean field mouse Apodemus peninsulae (Rodentia, Murinae) performed by chromosome microdissection and FISH. AB - Comparative analysis of micro B and macro B chromosomes of the Korean field mouse Apodemus peninsulae, collected in populations from Siberia and the Russian Far East, was performed with Giemsa, DAPI, Ag-NOR staining and chromosome painting with whole and partial chromosome probes generated by microdissection and DOP PCR. DNA composition of micro B chromosomes was different from that of macro B chromosomes. All analyzed micro B chromosomes contained clusters of DNA repeats associated with regions characterized by an uncondensed state in mitosis. Giemsa and DAPI staining did not reveal these regions. Their presence in micro B chromosomes led to their special morphology and underestimation in size. DNA repeat clusters homologous to DNA of micro B chromosome arms were also revealed in telomeric regions of some macro B chromosomes of specimens captured in Siberian regions. Neither active NORs nor clusters of ribosomal DNA were found in the uncondensed regions of micro B chromosomes. Possible evolutionary pathways for the origin of macro and micro B chromosomes are discussed. PMID- 15292606 TI - FISH detection of ribosomal cistrons and assortment-distortion for X and B chromosomes in Dichroplus pratensis (Acrididae). AB - Assortment-distortion with respect to the X and NOR activity of a rare mitotically stable B chromosome (B(N)), was examined in 16 males of Dichroplus pratensis (Acrididae: Melanoplinae) from Argentine populations. In 1B individuals, the X and B associate preferentially during prophase I reaching a maximum level of association at zygotene. Frequency of X/B association remains relatively high up to diplotene-diakinesis and decreases steeply towards metaphase I. The percent X/B association at each stage is positively influenced by association at the previous stage, and interindividual variability in X/B association decreases as the frequency of association increases. Both chromosomes tended to preferentially orientate toward the same pole at MI (mean ratio of 16 individuals, 1.50:1) which determined an excess of XB and 00 second spermatocytes over X0 and 0B ones (1.39:1). No significant differences occurred between the MI, AI and MII assortment ratios. Fluorescent in situ hybridisation (FISH) confirmed that the B chromosome carries ribosomal genes and helped to establish that, during spermiogenesis, both the B and the normal NOR-bearing chromosome (S8) are clustered near the centriole adjunct region of spermatids. However, FISH failed to reveal the existence of inactive ribosomal cistrons in the X chromosome, as previously suggested, thus providing no support to a simple origin of the B from the X. PMID- 15292607 TI - X and B chromosomes display similar meiotic characteristics in male grasshoppers. AB - We have analysed the chromosome organisation and the location and temporal appearance of different proteins in X and B chromosomes in the grasshopper Eyprepocnemis plorans throughout the first meiotic prophase. We have used adult males that carry a B chromosome collected in natural Spanish populations. The scaffold organisation has been analysed by means of silver stained chromatid cores. In addition, we have detected by immunolabelling the presence of phosphoepitopes, the ensemble of cohesin axes, the location of histone gamma H2AX, and recombinase Rad51. Our observations demonstrate that X and B chromosomes share similarities in chromatin organisation and in the expression of the tested proteins, which strongly differ from those of the autosomes. These results could be interpreted either as a support to the hypothesis that the Bs analysed here originated from the X chromosome, and/or that their chromatin composition and precocious condensation could determine their meiotic behaviour. PMID- 15292608 TI - An asymptotic determination of minimum centromere size for the maize B chromosome. AB - The maize B chromosome is a dispensable chromosome and therefore serves as a model system to study centromere function. The B centromere region is estimated to be approximately 9,000 kb in size and contains a 1.4 kb repeat that is specific to this centromere. When maintained as a univalent, the B chromosome occasionally undergoes centric misdivision. Consecutive misdivision analysis of the maize B chromosome centromere has generated a collection of functional centromeres that are greatly reduced in complexity. These small centromeres are often correlated with strongly reduced meiotic transmission. Molecular analyses of the misdivision collection have revealed that the smallest functional maize B centromere is a minimum of 110 kb in size. Considering the collection as a whole, meiotic transmission becomes severely compromised when the estimated centromere size is reduced to a few hundred kilobases. PMID- 15292609 TI - B chromosomes in hybrids of temperate cereals and grasses. AB - B chromosomes are considered to be genetically inert, yet often have pronounced and surprising effects upon the A chromosome behaviour at meiosis in inter generic and inter-specific hybrids. We review here our current knowledge of these effects in a number of different hybrids of the temperate cereals and grasses. Through hybridisation, many effects comparable to the pairing control system of wheat are uncovered, together with complex interactions of B chromosomes with hybrid host genotypes. We discuss the genetic and physical basis of the effects and try to make sense of them in terms of what we know about the origin and evolution of B chromosomes in plants. PMID- 15292610 TI - Different numbers of rye B chromosomes induce identical compaction changes in distinct A chromosome domains. AB - In rye each B chromosome (B) represents 5.5% of the diploid A genome. Rye Bs have several nuclear to whole plant effects although they seem to bear no genes except for the ones that lead to their maintenance within a population. In this context, and considering that rye Bs are enriched in repetitive non-coding regions that build up heterochromatin (het), we investigated the influence of Bs on the organization of two chromatin fractions, namely the ribosomal DNA (facultative het) and satellite (non-het) domain of rye chromosome 1 by silver staining on root tip metaphase cells. The results show that rye Bs cause condensation both in the NOR and in the chromosome 1 satellite domain. Since the silver staining technique used is indicative of the transcriptional activity of the NORs, the condensation observed at those loci demonstrates that the rRNA gene arrays are down-regulated in the presence of Bs, regardless of their number per individual. Furthermore, the organizational changes of metaphase NORs find parallel with the interphase organization of ribosomal chromatin, since the frequency of cells with intranucleolar condensed rDNA regions increases drastically and nuclear matrix attachment pattern is altered in the presence of the Bs. Our results show an identical effect of the Bs on the organization of two distinct chromosome domains displaying a presence/absence dichotomy. PMID- 15292611 TI - The odd-even effect in mitotically unstable B chromosomes in grasshoppers. AB - The odd-even effect, by which B chromosomes are more detrimental in odd numbers, has been reported in plants and animals. In grasshoppers, there are only a few reports of this effect and all were referred to as traits related to the formation of aberrant meiotic products (AMPs). Here we review the existing information about B chromosome effects on AMPs, chiasma frequency and the number of active nucleolus organizer regions (NORs) per cell. Polysomy for A chromosomes and B chromosomes are two kinds of chromosome polymorphism frequently found in grasshoppers. In some aspects, e.g. meiotic behaviour and mitotic instability leading to individual mosaicism (in the case of mitotically unstable Bs), polysomic As show similar characteristics to B chromosomes. In fact, polysomy is regarded as one of the main mechanisms for B chromosome origin. Here we review some features of meiotic behaviour in known cases of polysomy and mitotically unstable Bs in grasshoppers, in looking for possible causes for the odd-even effect. In all these traits, the odd-even effect was apparent, although its appearance was not universal in any case, with variation among species or populations within the same species. The equational division and lagging of the extra chromosomes, when univalents, could favour the appearance of abnormal meiotic products, and the formation of bivalents, when there are two or more extra chromosomes, inhibits this process. Therefore, the odd-even effect might be a consequence of the concomitant operation of both aspects of extra chromosome meiotic behaviour. The possibility that the odd-even effect might result from an increase in cell stress generated by odd numbers is suggested. PMID- 15292612 TI - The B chromosome polymorphism of the grasshopper Eyprepocnemis plorans in North Africa. IV. Transmission of rare B chromosome variants. AB - In addition to the principal B chromosome (B(1)) in Moroccan populations of the grasshopper Eyprepocnemis plorans, nine B chromosome variants appeared at low frequency. The transmission of five of these rare B chromosome variants through females was analysed in three natural populations. Sixteen controlled crosses provided useful information on the transmission of B(M2), B(M6) and B(M7) in Smir, B(M3) and B(M6) in SO.DE.A. (Societe de Developpement Agricole lands near Ksar-el-Kebir city), and B(M2) and B(M10) in Mechra, all located in Morocco. Since six female parents carried two different B variants, a total of 22 progeny analyses could be studied. Intraindividual variation in B transmission rate (k(B)) was observed among the successive egg pods in 26.7 % of the females, but this variation did not show a consistent temporal pattern. Only the B(M2) and B(M6) variants in Smir showed net drive, although variation was high among crosses, especially for B(M2). These two variants are thus good candidates for future regenerations (the replacement of a neutralized B, B(1) in this case, by a new driving variant, B(M2) or B(M6)) in Smir, the northern population where the B polymorphism is presumably older. The analysis of all crosses performed in the three populations, including those reported previously for the analysis of B(1) transmission, showed that the largest variance in k(B) among crosses stands at the individual level, and not at population or type of B levels. The implications of these findings for the occurrence of possible regeneration processes in Moroccan populations are discussed. PMID- 15292614 TI - Transmission analysis of B chromosomes in Rattus rattus from Northern Africa. AB - Traditionally, B chromosomes have been classified as parasitic or heterotic, depending of whether or not they show selfish behaviour. Nevertheless, experimental evidence has been found supporting the idea that supernumerary chromosomes may evolve from parasitism to neutrality. In this work, B chromosome transmission in Rattus rattus has been analysed by performing several crosses between individuals carrying different numbers of supernumerary chromosomes. Our results demonstrated a Mendelian transmission rate through males, but slight accumulation of the Bs through females. This parasitic behaviour is shared in populations as distant as Asia and Africa, and even in a related species in Australia, suggesting the possibility of an ancient origin of these supernumerary chromosomes. PMID- 15292613 TI - Rapid suppression of drive for a parasitic B chromosome. AB - The persistence of parasitic B chromosomes in natural populations depends on both B ability to drive and host response to counteracting it. In the grasshopper Eyprepocnemis plorans, the B24 chromosome is the most widespread B chromosome variant in the Torrox area (Malaga, Spain). Its evolutionary success, replacing its ancestral neutralized B variant, B2, was based on meiotic drive in females, as we showed in a sample caught in 1992. In females collected six years later, mean B24 transmission ratio (k(B)) was 0.523, implying a very rapid decrease from the 0.696 observed in 1992. This shows that B24 neutralization is running very fast and suggests that it might most likely be based on a single gene of major effect. PMID- 15292615 TI - B chromosomes and Robertsonian fusions of Dichroplus pratensis (Acrididae): intraspecific support for the centromeric drive theory. AB - We tested the centromeric drive theory of karyotypic evolution in the grasshopper Dichroplus pratensis, which is simultaneously polymorphic for eight Robertsonian fusions and two classes of B chromosomes. A logistic regression analysis performed on 53 natural populations from Argentina revealed that B chromosomes are more probably found in populations with a higher proportion of acrocentric chromosomes, as the theory predicts. Furthermore, frequencies of B-carrying individuals are significantly negatively correlated with the mean frequency of different Robertsonian fusions per individual. No significant correlations between presence/absence or frequency of Bs, and latitude or altitude of the sampled populations, were found. We thus provide the first intraspecific evidence supporting the centromeric drive theory in relation to the establishment of B chromosomes in natural populations. PMID- 15292616 TI - Cytogeography and the evolutionary significance of B chromosomes in relation to inverted rearrangements in a grasshopper species. AB - The analysis of geographic distribution of polymorphic cytological markers, briefly termed as cytogeography, can be considered an important tool to be applied when studying the evolutionary significance of chromosome variability within a species, either to unravel the adaptive significance of chromosome polymorphisms or to investigate the parasitic nature of some genomic elements. In this article we review cytogeographical studies in Trimerotropis pallidipennis, a grasshopper species whose South American populations display geographical patterns of distribution of inversion and B chromosome polymorphisms. Several lines of evidence that issue from the analysis of the geographic distribution of polymorphic markers suggest that inverted chromosomes are special sequences that are maintained by deterministic forces. On the other hand, the pattern of distribution of B chromosome polymorphism clearly demonstrates its selfish nature, being more frequent in those populations in central environments. We also present the analysis of 272 individuals of T. pallidipennis from Uspallata, and demonstrate that Bs in this population have some influence on body size, enlarging many of the morphometric characters of individuals and we propose it could be the consequence of its genotypic disequilibrium with one inversion. These investigations are finally discussed with regard to the models proposed for the maintenance of B chromosomes in natural populations and in relation to the possible interactions with chromosome inversions. PMID- 15292617 TI - Mitotically unstable B chromosome polymorphism in the grasshopper Dichroplus elongatus. AB - Dichroplus elongatus, a widespread South American phytophagous grasshopper, exhibits polymorphisms for supernumerary chromosomes and segments (SS) in natural populations in Argentina. In this paper we review the available information on B chromosome polymorphism in D. elongatus related to geographic distribution, patterns of chromosome variation and influence on sperm formation. In D. elongatus the different forms of supernumerary variants are not independent. The proportion of B-carrying individuals (B prevalence) is negatively correlated with SS10 and positively with SS6 frequencies. The analysis of population structure considering the different supernumerary variants would suggest that the patterns of chromosome variation can not be explained only by random factors. Geographic distribution was analyzed scoring the prevalence of B chromosomes in 13 natural populations collected in three different biogeographical provinces from Northwest (Las Yungas province) and East (Espinal and Pampeana provinces) of Argentina. The detected heterogeneity may be explained by significant differentiation between Northwest and East regions and among populations within Las Yungas and Pampeana provinces. Correlation analysis suggested that B chromosome prevalence is associated with maximum temperature and with latitude. Additional information about the nature of the patterns of B chromosome variation was obtained comparing them with those obtained at the mitochondrial DNA level. The hierarchical analysis of molecular differentiation revealed discrepancy with respect to chromosome differentiation and also suggested that the pattern of B chromosomes may not be explained by historical factors. We also discussed the probable influence on fertility of carriers considering the production of abnormal sperm formation (macro and microspermatids) in relation to the number of Bs per follicle. PMID- 15292618 TI - Geographic and seasonal variations of the number of B chromosomes and external morphology in Psathyropus tenuipes (Arachnida: Opiliones). AB - Psathyropus tenuipes (= Metagagrella tenuipes) is a harvestman that harbors B chromosomes with extremely high frequency (individuals without Bs are only 1% of the total number of specimens so far examined) and high numbers (mean number of Bs per individual is about 4). Geographic variations of the number of Bs and external morphology of the species and the relationship between them were studied. A northward increase in the number of Bs was detected throughout the Japanese Islands, though the number also varied considerably locally. Latitudinal gradients were also found in some external characters, while there were no correlations between those external morphologies and the number of Bs. Principal component analysis using eight morphological data for 21 populations revealed four geographical groups that reflect actual location of the populations. Populations along the Seto Inland Sea were characterized by a lower number of Bs than those in other areas. Seasonal change was also found in a population (Yatsukami in western Honshu) in both 1994 and 1995 for the number of Bs, though the number in the same population was stable at least throughout later postembryonic stages in both 1997 and 1998. Embryos contained fewer numbers of Bs than adults, suggesting that females of the species tend to lay eggs with fewer numbers of Bs. PMID- 15292619 TI - Spatio-temporal dynamics of a neutralized B chromosome in the grasshopper Eyprepocnemis plorans. AB - Spatial and temporal patterns of frequency variation for a neutralized B chromosome in the grasshopper Eyprepocnemis plorans were analyzed along six transects in the east of Spain to explore possible factors affecting the population dynamics of this polymorphism. Three parameters were employed to quantify B frequency: prevalence, load and mean frequency. Of them, load seemed to be the less sensitive parameter, probably due to its small range of variation. Prevalence, however, shows ample variation, but the mean frequency of B chromosomes per individual is the best parameter to characterize B frequency. Only river transects revealed significant differences among populations, and the use of two geographic explicit approaches (Mantel test and distograms) revealed significant isolation by distance (IBD), especially at the Segura River mouth, presumably due to low gene flow and drift. No temporal trend was found in the Segura River transects, which is consistent with the slow changes in B frequency expected during the random walk for neutralized B chromosomes. But these transects showed a clear spatial pattern, with B1 showing lower frequency in the upper course of this river. The present results provide the first empirical evidence of IBD in the evolution of a neutralized B chromosome, and support the notion that B dynamics at this evolutionary stage is best explained by a metapopulation approach. PMID- 15292620 TI - The parasitic effects of rye B chromosomes might be beneficial in the long term. AB - Rye B chromosomes (Bs) have strong parasitic effects on fertility. B carrying plants are less fertile than 0B ones, whereas the Bs have no significant effects on plant vigour. On the other hand, it has been reported that B transmission is under genetic control in such a way that H line plants transmit the Bs at high frequency, whereas the Bs in the low B transmission rate line (L) fail to pair at metaphase I and are frequently lost. In the present work we analyse variables affecting vigour and fertility considering not only the number of Bs of each plant, but also its H or L status and the B number of its maternal parent. Our results show that the Bs not only decrease female fertility of the B carrier, but the fertility of its progeny, with the exception of 0B plants coming from a 4B mother, which are the most fertile. In this way B chromosomes can be considered as a selective factor. Pollen abortion was higher in B carriers, in the progeny of B carriers and in H plants, but 4B plants coming from B carrying mothers produce less aborted pollen, indicating that a high B number is more deleterious if it is transmitted in the pollen grains. A similar result was obtained for endosperm quality estimated as grain weight, because it is negatively influenced by the Bs in 4B plants coming from a 0B mother. H plants were always less fertile than L ones, indicating that alleles increasing the loss of Bs in the L line will be probably selected as a defence of the A genome against the invasive Bs of the H line. Flower number is not affected by the Bs. PMID- 15292621 TI - Interaction of B chromosomes with A or B chromosomes in segregation in insects. AB - Additional or B chromosomes not belonging to the regular karyotype of a species are found in many animal and plant groups. They form a highly heterogeneous group with respect to their morphology and behaviour both in mitosis and meiosis. Achiasmatic mechanisms that ensure the segregation of a B chromosome from another B chromosome or from an A chromosome are reviewed. An achiasmatic mechanism characterized by the "distance pairing" of segregating univalents at metaphase I was found to be responsible for the preferential segregation of B chromosome univalents in Hemerobius marginatus L. (Neuroptera), and a mechanism characterized by the "touch and go pairing" of segregating univalents was responsible for the highly regular segregation of a B chromosome and the X chromosome in Rhinocola aceris (L.) (Psylloidea, Homoptera). The latter mechanism resulted in the integration of a B chromosome to the A chromosome set as a Y chromosome in a psyllid species Cacopsylla peregrina (Frst.). Furthermore, B chromosomes can disturb the regular segregation of the achiasmatic X and Y chromosomes resulting in the formation of X0/XY polymorphism in a population, which might precede the loss of the Y chromosome. The absence of observations on accurately functioning achiasmatic segregation mechanisms in grasshoppers (Orthoptera) was attributed to the X and B chromosomes, which re-orient one or several times during metaphase I. Apparently, these re-orientations mask any achiasmatic segregation mechanism that might operate during meiotic prophase in these insects. PMID- 15292622 TI - Imitate to integrate: reviewing the pathway for B chromosome integration in Trypoxylon (Trypargilum) albitarse (Hymenoptera, Sphecidae). AB - B chromosomes are genomic "intruders" normally characterized by their total dispensability counteracted by a variety of drive mechanisms, which assures their presence regardless of their harmful effects on the host genome. From an evolutionary standpoint, the relationship between standard (A) and B chromosomes can go through different pathways, from an everlasting arms race to a cordial B integration. Examples underlying the first situation are fairly common; B integration, however, has been more a theoretical than a practical possibility. The B chromosome in the haplodiploid solitary wasp Trypoxylon albitarse is probably the first example of a "mimetic" B, which is being integrated into the A genome by limiting itself to one B per haploid genome, the same dosage as the A chromosomes. Here we review some of the findings underlying this hypothesis and discuss the T. albitarse B strategy as a possible mechanism for B chromosome integration as a regular member of the chromosome complement in haplodiploid organisms. PMID- 15292623 TI - B chromosomes: the troubles of integration. AB - Starting with a spontaneous B-A centric fusion found in a natural population of the grasshopper Eyprepocnemis plorans, we have obtained different strains carrying the rearrangement in various conditions and doses. Using this material, we have analyzed the meiotic behavior of the translocated chromosome in living cultured spermatocytes, simulating the successive steps of a hypothetical process of integration of a B chromosome into the standard genome via B-A centric fusion. Remarkably, the behavior of fusion heterozygotes, the initial step of the integration process, is much more regular than that of any other configuration, including homozygotes. The reasons for the failure of B chromosome integration into the normal complement by translocation are discussed. PMID- 15292624 TI - Fibromyalgia: a stress disorder? Piecing the biopsychosocial puzzle together. AB - Fibromyalgia (FM) is a controversial syndrome, characterised by persistent widespread pain, abnormal pain sensitivity and additional symptoms such as fatigue and sleep disturbance. The syndrome largely overlaps with other functional somatic disorders, particularly chronic fatigue syndrome (CFS). Although the exact aetiology and pathogenesis of FM are still unknown, it has been suggested that stress may play a key role in the syndrome. This article first reviews the function of the stress response system, placing special emphasis on the relationships between adverse life experiences, stress regulation and pain-processing mechanisms, and summarising the evidence for a possible aetiopathogenetic role of stress in FM. Finally, an integrative biopsychosocial model that conceptualizes FM as a stress disorder is proposed, and the clinical and research implications of the model are discussed. PMID- 15292625 TI - Psychosocial intervention for women with primary, non-metastatic breast cancer: a comparison between participants and non-participants. AB - BACKGROUND: Despite improvements in the medical treatment of breast cancer, resulting in better prognoses, women diagnosed with the illness often experience psychosocial problems. As a result, many psychosocial intervention programs have been developed, usually with positive results. The aim of this study was to examine the effects of two 12-week psychosocial intervention programs for women with a primary, non-metastatic breast cancer diagnosis in comparison to women who were on a waiting list for these interventions for 3 months. METHODS: Sixty-nine women with primary, non-metastatic breast cancer, but otherwise without psychosocial problems, were randomized to a group of patients treated with the intervention program (group psychotherapy or social support group) or a control group (on a waiting list). Differences between both groups in psychosocial adjustment, social support and coping at the short-term follow-up were described in this study. RESULTS: Women who participated in the group intervention programs did not differ from women in the control group regarding psychosocial adjustment at the end of the study. Women who participated in the social support groups reported to receive more social support from others not very close to them. They also used more palliative coping than women from the psychotherapy group. CONCLUSIONS: Apparently, women who are being diagnosed with breast cancer, but otherwise have no psychosocial adjustment problems following the diagnosis, do not especially benefit from a short-term intervention program compared to women in the control group. PMID- 15292626 TI - Reduced anxiety level by therapeutic interventions and cell-mediated immunity in panic disorder patients. AB - OBJECTIVE: This study examined the relationship between reduced anxiety level by therapeutic interventions and cell-mediated immunity (CMI) in patients with panic disorder. METHODS: The subjects consisted of 42 patients with panic disorder and 42 normal gender- and age-matched controls. Among the patients, 21 were randomly assigned to a combined treatment of cognitive-behavioral therapy and the benzodiazepine antianxiety agent ethyl loflazepate (2 mg daily), and 21 were assigned to the antianxiety agent only. The treatment lasted for 6 weeks. Cell mediated immune function was measured by the lymphocyte proliferative response to phytohemagglutinin (PHA) and interleukin-2 (IL-2) production. The anxiety level was assessed by the Hamilton Rating Scale for Anxiety and the anxiety subscale of the Symptom Checklist-90 Revised. RESULTS: Prior to treatment, the panic disorder patients had significantly lower IL-2 production and blastogenic response to PHA than the normal controls. However, no significant differences in CMI were found between the pretreatment and posttreatment period in either the patient group receiving medication only or the combined treatment group, though after treatment, patients were significantly less anxious than before treatment in both intervention groups. The delta change (posttreatment value minus pretreatment value) in the self-reported anxiety level was significantly associated with the delta change in the blastogenic response in the combined treatment group. CONCLUSION: These findings suggest that panic disorder may be associated with decreased CMI, and the reduced level of self-reported anxiety in the patients who underwent combined therapeutic intervention is likely to increase the blastogenic response. Further studies are needed to evaluate the long-term effects of treatment on immune function. PMID- 15292627 TI - Predicting long-term outcome in group treatment of atopic dermatitis. AB - BACKGROUND: Psychological group treatments are effective adjuncts to dermatological treatments in improving dermatological symptoms and quality of life. However, no data are available on variables that may predict which patients benefit from such group treatments. METHODS: On the basis of data from the Marburg group treatment study, we explored which variables are associated with successful treatment. Treatment outcome was defined by a marked reduction of dermatological symptoms at 1-year follow-up. RESULTS: Regardless of the treatment condition, a high pretreatment level of scratching, a low serum IgE level, low scores on the internal Health Locus of Control Scale and a high frequency of coping-related cognitions regarding itching predicted good outcome at 1 year. Logistic regression analyses indicated that the predictive power of locus of control and itch-related cognitions differed between different treatment conditions. CONCLUSION: In patients with atopic dermatitis, indication for psychological treatment may be based on the assessment of psychological target problems including excessive scratching and dysfunctional cognitions. PMID- 15292628 TI - Study on effects of life review activities on the quality of life of the elderly: a randomized controlled trial. AB - BACKGROUND: The objective of this study was to evaluate the mid-term efficacy of life review activities on the quality of life (QOL) of the elderly by conducting a randomized controlled trial, and to identify the factors that should be taken into consideration when conducting life review activities. METHODS: Written consent was obtained from 80 of the 97 eligible elderly persons. After randomly assigning them to two groups, an intervention group and a control group, group life review activities were conducted in the intervention group and discussion activities about health were conducted in the control group. In both the intervention group and the control group, life satisfaction, self-esteem, depression, and hopelessness were evaluated using self-rating scales at three points: at baseline, immediately after completion of the 8 weeks of sessions, and 3 months after completion of the intervention. RESULTS: Repeated measures analysis of covariance showed significant differences between the two groups in the changes in scores for depression (p = 0.04) and hopelessness (p = 0.04). Regarding the factors that were associated with depression and hopelessness, 3 months after completion of the intervention, depression and hopelessness of a more severe nature at baseline and having greater unresolved conflicts in the past were extracted by multiple regression analysis. CONCLUSIONS: The results suggested that group life review activities have a role in assisting the developmental stage of old age and supporting mental health, and have mid- to long-term effectiveness in maintaining and improving the QOL of the elderly. PMID- 15292629 TI - Impact of a psychoeducational family intervention on caregivers of stabilized bipolar patients. AB - BACKGROUND: Environmental stress has an important role in the course of bipolar disorder. Some findings have shown that family beliefs about the illness could predict family burden, and this burden could influence the outcome of bipolar disorder. To the best of our knowledge, there is scant information about the effects of family intervention on the caregiver's burden in bipolar disorder. The aim of this study was to assess the effects of psychoeducational family intervention on bipolar patients' caregivers, including the assessment of the caregiver's burden. METHODS: 45 medicated euthymic bipolar outpatients were randomized into an experimental and a control group. Relatives of patients from the experimental group received 12 psychoeducational, 90-min sessions about bipolar disorder and coping skills. The caregivers' knowledge of bipolar disorder, the relationship subscales of the Family Environment Scale, and the family burden subscales from an adapted version of the Social Behavior Assessment Schedule were assessed for both caregiver groups before and after the intervention. RESULTS: Psycho-educated caregivers significantly improved their knowledge of bipolar disorder and reduced both the subjective burden and the caregiver's belief about the link between the objective burden and the patient. No significant differences were found in the objective burden nor in the family relationship subscales. CONCLUSIONS: These preliminary results suggest that psychoeducational intervention on caregivers of bipolar patients may improve the caregiver's knowledge of the illness, reduce their distress or subjective burden and alter their beliefs about the link between the disruptions in their life and the patient's illness. PMID- 15292630 TI - Language acquisition in relation to cumulative posttraumatic stress disorder symptom load over time in a sample of re-settled refugees. AB - OBJECTIVE: To study the effects of symptoms of posttraumatic stress disorder (PTSD), depression and dissociation as well as cumulative symptom load on language learning during the introduction phase in re-settled refugees. METHOD: Participants were re-settled refugees of Iraqi origin. They were assessed by means of a structured interview for PTSD at baseline as well as self-rating questionnaires. Language acquisition was studied by means of register data from the school system. Five levels of language proficiency were recorded. Self reported symptom scores for PTSD, depression and dissociation (Impact of Events Scale-22, Hopkins Symptom Checklist-25, Dissociative Experiences Scale) were measured at four time points during 9 months immediately after resettlement. In 49 participants in a longitudinal study, data regarding progress in language studies were accessible. RESULTS: The results of the study indicate that the speed of language acquisition - the number of levels taken during the study, adjusted to hours of school presence - is related to the cumulative PTSD symptom load over time (Events Scale-22), but is not related neither to the symptom load of depression and dissociation, nor to the number of previous school years. CONCLUSION: The study shows that the symptom load of PTSD during the follow-up period is significantly inversely related to the speed of language acquisition in refugees. This implies that treatment as well as preventive measures against worsening of PTSD symptoms are important in order to minimise harmful post migration stress for the facilitation of integration. PMID- 15292631 TI - QT interval and QT dispersion in eating disorders. AB - BACKGROUND: Eating disorders are thought to be risk factors for cardiac sudden death secondary to arrhythmia. Results in previous studies on QT interval and QT dispersion, markers of fatal arrhythmia, have been inconsistent. METHODS: We prospectively examined 179 female eating disorder patients, being over 18 years old and diagnosed according to the DSM-IV criteria between January 1995 and December 2002, and 52 healthy women. Patients with abnormal plasma electrolytes or taking medications that might influence the electrocardiogram (ECG) were excluded from the study. QT intervals were corrected for heart rate using Bazett's formula and the nomogram method, which is more reliable at extremely low heart rates than Bazett's formula. QT dispersion was measured as the difference between the longest and shortest QT intervals. QT intervals and QT dispersion in each patient group were compared with those in the control group. RESULTS: The 164 eligible patients consisted of 43 patients with anorexia nervosa restricting type, 35 with anorexia nervosa binge eating/purging type, 63 with bulimia nervosa purging type, and 23 with bulimia nervosa non-purging type. There was no significant difference in age between eating disorder patients and controls. QT interval and QT dispersion were significantly longer in all eating disorder subtypes than in the control group. QT interval and QT dispersion were significantly correlated with the rate of body weight loss in bulimia nervosa. CONCLUSIONS: QT interval and QT dispersion were prolonged in both anorexia nervosa and bulimia nervosa. Examination of ECG in eating disorder patients without extremely low body weight also appears to be clinically important. PMID- 15292632 TI - Occult tight filum terminale syndrome: results of surgical untethering. AB - The entity of an occult tight filum terminale syndrome, characterized by clinical findings consistent with a tethered cord syndrome, but with the conus ending in a normal position, has been recognized recently. The indications for sectioning the filum terminale in this situation are not well characterized and are controversial. We report a retrospective review of a consecutive series of 60 children (ages 3-18 years) with a diagnosis of occult tight filum terminale syndrome who underwent section of the filum and were followed for more than 6 months (mean 13.9 months). The criteria for surgical intervention were (1) spina bifida occulta, (2) progressive bladder instability unresponsive to conservative measures, (3) urological/nephrological evaluation to confirm or rule out nonneurogenic etiology, and (4) two or more of the following: (a) bowel involvement (fecal incontinence or chronic constipation), (b) lower extremity weakness, (c) gait changes, (d) reflex/tone abnormalities, (e) sensory disturbances, (f) back/leg pain, (g) orthopedic abnormalities/limb length discrepancy, (h) scoliosis/lordosis, (i) recurrent urinary tract infections, (j) abnormal voiding cystourethrogram/ultrasound, (k) syringomyelia, and (l) neurocutaneous stigmata. Postoperatively, urinary incontinence/retention showed complete resolution in 52%, marked improvement (>95% resolution) in 35%, moderate improvement (>75%) in 6%, minimal improvement (> 50%) in 6%, and no improvement (<50%) in 2%. Fecal incontinence completely resolved in 56%, improved in 41%, and was unchanged in 3%. Weakness, sensory abnormalities, and pain improved or resolved in all patients. PMID- 15292634 TI - Endoscopic cyst fenestration outcomes in children one year of age or less. AB - The use of endoscopic fenestration (EF) is becoming an increasingly common treatment for symptomatic intracranial cysts. Very little data exist regarding outcomes for this procedure in children 1 year of age or younger. We retrospectively reviewed the clinical outcomes of 8 children 1 year of age or less treated at our institution with endoscopic cyst fenestration. The mean follow-up was roughly 2.5 years. These data were combined with 17 other cases obtained from the published literature. EF was successful in rendering patients shunt-free or minimizing the number of ventricular catheters in 18 of 26 operations. There were 8 outright failures -- two in 1 patient. Given the risks and complications of cerebrospinal fluid shunting in children less than 1 year of age, we advocate the consideration of EF as initial treatment of symptomatic intracranial cysts. PMID- 15292635 TI - In vitro bacterial adherence to ventriculoperitoneal shunts. AB - Bacterial adherence to medical devices has been recognized as an important initial step in the infectious process, but it has not been fully elucidated regarding ventriculoperitoneal (VP) shunts. The aim of the present study was to quantitatively determine the adherence in vitro of bacteria known to cause VP shunt infections and to identify factors affecting the process. Clinical isolates studied included Staphylococcus epidermidis, Staphylococcus aureus, Streptococcus pneumoniae, and Escherichia coli. Adherence was examined quantitatively per square centimeter, visualized by electron microscopy and related to slime production and hydrophobicity. Although all four strains adhered to VP shunts, there were marked differences, with S. epidermidis and S. aureus showing the highest adherence (67.0 x 10(3) and 15.2 x 10(3) bacteria/cm(2), respectively). Factors affecting adherence included incubation time and temperature, bacterial concentration, device material (lower for silicone than Teflon), slime production and hydrophobicity. These data might be helpful for devising novel strategies to reduce VP shunt infections. PMID- 15292636 TI - Spinal cord compression caused by extradural arachnoid cysts. Clinical examples and review. AB - Most spinal arachnoid cysts are asymptomatic and detected incidentally during magnetic resonance imaging or myelography. The etiology of intraspinal arachnoid cyst is not yet clear. We present two children with three spinal extradural arachnoid cysts and each cyst protruded from a separate dura defect. In both patients, plain radiographs demonstrated widening of the interpedicular distance, which suggested progressive widening of the spinal bony canal. Limited laminectomy was performed to remove the intraspinal cysts. Separate dura defects, the apparent predisposing factors, were also found and repaired. The patients completely recovered neurologically. Radical cyst removal and dura defect closure are the surgical intervention of choice in patients with symptomatic extradural arachnoid cyst. PMID- 15292637 TI - Thoracic meningocele, meningomyelocele or myelocystocele? Diagnostic difficulties, consequent implications and treatment. AB - Spina bifida cystica is a closing disorder of the neural tube which infrequently occurs in the thoracic region. A rare lesion called myelocystocele is a variant of spina bifida cystica and is associated with syringomyelia, Chiari type 2 malformation and hydrocephalus. Usually the patient has no neurological deficit, but future deterioration can occur due to posterior tethering of the spinal cord by adhesions. The prenatal diagnosis by ultrasound study can be misleading and in order to attain the correct diagnosis, especially if abortion is considered, a prenatal MRI scan should be done before the parents are counselled, and should be repeated prior to operative treatment. Surgical correction of myelocystocele is not only for cosmetic reasons, but also to untether the spinal cord prophylactically to prevent future neurological deterioration. In this case report, we present a child born with a thoracic myelocystocele, the diagnostic difficulties, consequent implications and surgical treatment. PMID- 15292638 TI - Split cord malformation as a cause of tethered cord syndrome in a 78-Year-old female. AB - A 78-year-old woman presented for evaluation of back pain, urinary dysfunction, leg weakness and progressive equinovarus foot deformity. She reported that shortly after her birth in 1924, she underwent resection of a subcutaneous 'cyst' in the lower lumbar area. Seven years prior to evaluation at our institution, she had undergone bilateral total knee arthroplasty for osteoarthritis. After the procedure, she began to experience severe low back pain that radiated into her legs. Weakness of the foot inverters, urinary dysfunction and worsening bilateral equinovarus foot deformity developed in the years following the surgery. MRI revealed a split cord malformation with a tethered spinal cord. Because of the patient's age and poor medical condition, her symptoms were managed conservatively. This case demonstrates symptomatic deterioration in an elderly patient with a tethered spinal cord after many years of clinical stability. PMID- 15292639 TI - Expanding cava septi pellucidi with hydrocephalus in infancy. PMID- 15292640 TI - Untreated parieto-occipital encephalomeningohydrocele. PMID- 15292641 TI - Blood vessel mimicking a dermal sinus tract. PMID- 15292642 TI - Rare complications of shunt infection. PMID- 15292643 TI - Ventral tethering of the spinal cord. PMID- 15292644 TI - Meroacrania. PMID- 15292645 TI - Currarino triad as an anterior sacral meningocele. PMID- 15292646 TI - Surgical outcome following resection of contrast-enhanced pediatric brainstem gliomas. PMID- 15292648 TI - Efficacy of donepezil treatment in Alzheimer patients with and without subcortical vascular lesions. AB - In a pilot study designed as a case control study the efficacy of donepezil treatment was investigated in patients with Alzheimer's disease (AD). Patients were stratified according to radiological criteria into patients without (AD group) and with subcortical vascular lesions (AD+SVD group). Changes in cognition were assessed as the primary outcome measurement after 6 and 18 months of treatment by the Mini-Mental Status Examination (MMSE) and the Consortium to Establish a Registry for Alzheimer's Disease (CERAD) test battery. After 6 months, patients had improved from baseline by 0.7 points in MMSE score in the AD group and by 1.8 in the AD+SVD group. After 18 months of treatment, the AD+SVD group performed significantly worse in one CERAD subscore, whereas a deterioration in two subscores was observed in the AD group. A comparison between the 2 groups revealed that treatment did not lead to statistically significant differences between the AD and AD+SVD groups in any of CERAD parameters following 6 or 18 months of treatment. These data support previous observations that donepezil therapy is effective in AD patients with and without subcortical vascular lesions. PMID- 15292649 TI - Roxithromycin suppresses mucin gene expression in epithelial cells. AB - Macrolide antibiotics are believed to inhibit mucus secretion but the mechanism of action is unclear. This study was designed to investigate an effect of roxithromycin on MUC2 gene expression in cultured intestinal epithelial cells (HM3-MUC2 cells). A reporter gene assay was used for analysis. Roxithromycin suppressed MUC2 gene transcriptional activity in a dose-dependent manner in HM3 MUC2 cells. Phorbol 12-myristate 13-acetate (PMA), lipoteichoic acid (LTA), lipopolysaccharide (LPS) and leukotriene D4 (LTD4) significantly increased MUC2 luciferase activities in the following order: PMA > LTA > LTD4 > LPS. Roxithromycin also decreased MUC2 gene transcriptional activity induced by PMA in a dose-dependent manner. NF-kappaB activation, but not AP-1 activation, was significantly suppressed by roxithromycin in HM3-MUC2 cells. A suppression of NF kappaB activation was also observed in NCI-H292 cells. These results suggest that roxithromycin suppresses MUC2 gene expression in epithelial cells and that this suppression is probably via inhibition of NF-kappaB activation. PMID- 15292650 TI - Thiopentone and methohexitone enantiomers do not act stereoselectively on the oxidative response in human neutrophils in vitro. AB - To elucidate potential stereoselective effects of single barbiturate isomers, we compared the inhibitory potency of single thiopentone enantiomers, two isomer enriched mixtures of methohexitone and racemic mixtures of both barbiturates on the fMLP-induced neutrophil oxidative response. A suppression of the response to 50% compared to control required a 100-fold therapeutic concentration of methohexitone, while therapeutic concentrations of the thiopentone racemate led to a significant inhibition (relative fluorescence of neutrophils 0.46 +/- 0.03 compared to fMLP controls). The racemate of thiopentone produced significantly greater inhibition than the single enantiomers. Stereoselectivity in favor of one isomer could not be shown for both barbiturates. The greater inhibition by the thiopentone racemate might suggest two separate binding sites for the enantiomers which are positively coupled. PMID- 15292651 TI - Central endothelin-B receptor stimulation does not affect morphine analgesia in rats. AB - Several neurotransmitter mechanisms have been proposed to play a role in the actions of morphine. We reported that centrally administered endothelin A (ETA) receptor antagonists potentiate morphine analgesia in rats. It has also been reported that ETB agonist, IRL1620, has antinociceptive action mediated through opiate receptors in the periphery. The present study was conducted to determine if central ETB receptors are involved in analgesic actions of morphine. The effect of intracerebroventricular (i.c.v.) administration of ETB receptor agonist, IRL1620, on morphine-induced analgesia and hyperthermia was determined in the rat. Morphine (4 mg/kg, s.c.) produced a significant increase (84%) in tail-flick latency compared to the control group and the analgesic response lasted for 4 h. IRL1620 (30 microg, i.c.v.) did not produce any increase (16%) in tail-flick latency over the 5-hour observation period in vehicle-treated rats. Pretreatment with IRL1620 (3, 10, and 30 microg, i.c.v.) did not have any significant effect on the intensity and duration of morphine (4 mg/kg, s.c.) induced analgesia. Morphine (4 mg/kg, s.c.) administration produced an increase in body temperature compared to the control group. In vehicle-pretreated rats, IRL1620 (30 microg, i.c.v.) did not produce any change in body temperature. The morphine-induced hyperthermic effect was not altered in IRL1620-pretreated rats. These studies demonstrate that IRL1620, a specific ETB receptor agonist, did not affect the morphine-induced analgesic and hyperthermic effect in rats. It can be concluded that central ETB receptors are not involved in modulation of pharmacological actions of morphine. PMID- 15292652 TI - Effects of pranidipine, a novel calcium channel antagonist, on the progression of left ventricular dysfunction and remodeling in rats with heart failure. AB - The cardioprotective properties of pranidipine were studied in a rat model of heart failure after autoimmune myocarditis. Twenty-eight days after immunization the surviving rats were divided into three groups and received oral treatment of 0.03 mg/kg/day (group 0.03) or 0.3 mg/kg/day (group 0.3) of pranidipine or vehicle (group V) for 1 month. High-dose pranidipine (group 0.3) improved the survival rate, and significantly reduced heart weight, heart weight to body weight ratio, myocardial fibrosis, central venous pressure and left ventricular end-diastolic pressure than low-dose pranidipine (group 0.03) and group V. Pranidipine at high dose also decreased the left ventricular systolic and diastolic dimensions, and increased fractional shortening compared with group V. The increase in level of TGF-beta1 and collagen-III mRNA were suppressed by pranidipine in a dose-dependent manner. Our results indicated that pranidipine has cardioprotective effects on heart failure, and that the beneficial effect can be partly explained by attenuation of fibrotic response through suppression of TGF-beta1 and collagen-III mRNA expression, and regression of myocyte hypertrophy. PMID- 15292653 TI - Radioligand binding and functional characterization of recombinant human NmU1 and NmU2 receptors stably expressed in clonal human embryonic kidney-293 cells. AB - Neuromedin U (NmU) is a smooth muscle contracting peptide. Recently, two G protein-coupled receptors for NmU (NmU1R and NmU2R) have been cloned having approximately 50% homology. They have distinct patterns of expression suggesting they may have different biological functions. This study provides a comprehensive characterization of both NmU receptors expressed in human embryonic kidney 293 cells. [125I]hNmU binding to the recombinant NmU receptors was rapid, saturable, of high affinity and to a single population of binding sites. Exposure of these cells to NmU isopeptides resulted in an increase in intracellular [Ca2+]i release (EC50 value of 0.50 +/- 0.10 nmol/l) and inositol phosphate formation (EC50 1.6 +/- 0.2 and 1.50 +/- 0.4 nmol/l for NmU1R and NmU2R respectively). Furthermore, hNmU inhibited forskolin (3 micromol/l)-stimulated accumulation of cAMP in intact HEK-293 cells expressing either NmU1R or NmU2R. The inhibitory effect was significant for the cells expressing NmU2R with IC50 value of 0.80 +/- 0.21 nmol/l. In summary, both NmU1R and NmU2R in HEK-293 cells have similar signaling capability. PMID- 15292654 TI - Nordihydroguairetic acid, a lignin, prevents oxidative stress and the development of diabetic nephropathy in rats. AB - Recent evidences indicate a pivotal role of reactive oxygen species in etiology of diabetic nephropathy, an important microvascular complication of diabetes mellitus. Moreover, oxidative stress leads to an increased production of lipoxygenase derivatives which also play a role in diabetic nephropathy. The present study was thus designed to examine the effect of an antioxidant and a lipoxygenase inhibitor, nordihydroguairetic acid (NDGA), on renal function and oxidative stress in streptozotocin (STZ)-induced diabetic rats. Diabetes was induced by a single intraperitoneal injection of streptozotocin (65 mg/kg) in rats. After the 4th week of STZ injection, NDGA (5 and 10 mg/kg) was given subcutaneously (s.c.) for another 4 weeks to both control and diabetic rats. At the end of the 8th week, diabetic rats exhibited renal dysfunction as evidenced by reduced creatinine and urea clearance along with enhanced albumin excretion rate as compared with control rats. Biochemical analysis of kidneys revealed a marked increase in oxidative stress demonstrated by increased lipid peroxidation and decreased activities of key antioxidant enzymes, glutathione (GSH), superoxide dismutase (SOD) and catalase in diabetic rats. Chronic treatment with NDGA in diabetic rats significantly prevented both renal dysfunction and oxidative stress as compared with vehicle-treated diabetic rats. The kidneys of diabetic rats showed morphological changes such as hyaline casts, glomerular thickening and moderate interstitial fibrosis and arteriolopathy, whereas NDGA administration in diabetic rats markedly prevented renal morphological alterations. These results emphasize the role of oxidative stress in the pathophysiology of diabetic nephropathy and point towards the potential of NDGA as a complementary therapy for the prevention/treatment of diabetic nephropathy. PMID- 15292655 TI - Itch-scratch responses induced by lysophosphatidic acid in mice. AB - The present investigation was conducted in order to determine whether lysophosphatidic acid (LPA) induces itch-scratch responses (ISRs) in mice. Intradermal administration of LPA induces ISRs; furthermore, the time course for LPA-induced ISRs was similar to that for histamine-induced responses. Comparative study of the pruritogenic activity revealed that histamine possessed a potent effect characterized by a dose-response relationship; however, prostaglandin D2 failed to induce this response. Pretreatment with ketotifen, a histamine H1 receptor antagonist, and capsaicin inhibited LPA-induced ISRs. Additionally, LPA induced ISRs were abolished by Y-27632, an inhibitor of Rho-associated protein kinase (ROCK). These findings suggest that LPA-induced ISRs are attributable to histamine- and substance-P-mediated pathways. Moreover, the Rho/ROCK-mediated pathway may be involved. PMID- 15292656 TI - Lachrymal determinations: methods and updates on biopharmaceutical and clinical applications. AB - This article displays different procedures used to collect lachrymal fluid and describes some of its applications. Sampling tears represents the main difficulty to produce precise and reproducible results. The direct sampling procedure consists in collecting tears with capillary tubes and has the drawback of demanding previous stimulation that induces major dilution. The indirect method does not require preliminary stimulation but has been held responsible for altering epithelium and promoting leakage from plasma. Schirmer strips and sponges are classically required. Applications are numerous in biopharmaceutical and clinical fields. The determination of endogenous components has great potentiality as a diagnostic tool, but the use of tear as a substitute of plasma does not present clinical relevance. Levels of drugs like immunosuppressive or antibiotic agents are determined in tears to verify that pharmacological concentrations are reached and frequency of administration is deduced from kinetic fitting. PMID- 15292657 TI - Transplantation of the autologous submandibular gland to the lacrimal basin in rats. AB - PURPOSE: In this study, we designed an animal experiment in which we transferred a part of the autologous submandibular gland without performing a microvascular anastomosis. We studied histological changes and functional effects in the transfer glands. METHOD: Thirty male adult Sprague-Dawley rats were divided into three groups. In each animal of the three groups, the right eye was the control eye and the left eye the experimental eye. Surgical removal of the extraorbital lacrimal gland from the control eyes was performed to create a condition simulating keratoconjunctivitis sicca. In the experimental eyes of group 1, in addition to the removal of the extraorbital lacrimal gland, a part of the autologous submandibular gland was transferred to the orbit and fixed to the intraorbital lacrimal gland. In the experimental eyes of group 2, in addition to the removal of the extraorbital lacrimal gland, an aseptic silicon rubber was transferred to the orbit and fixed to the intraorbital lacrimal gland. In the experimental eyes of group 3, there was no removal of the extraorbital lacrimal gland but instead a sham operation was performed. The histological changes and innervation pattern in the transferred submandibular gland of group 1 were observed. Tear secretion of each group was measured to study the functional effect. RESULTS: Three months after the transplantation, the transferred submandibular glands were similar to the unoperated submandibular glands both in histology and innervation pattern. In group 1, the tear secretion in the experimental eyes was significantly greater than that of the control eyes at 2 and 3 months following transfer. In group 2, no significant difference was noted between the experimental eyes and control eyes. In group 3, the difference in tear secretion between the experimental eyes and control eyes was significant. CONCLUSION: The result of this study revealed that although the procedure did not involve vascular anastomosis, the transfer gland showed a normal histological appearance and good reinnervation, even 3 months after the transfer. These transferred submandibular glands continued to secrete tears. This secretion continued to increase up until the end of the experimental period. PMID- 15292658 TI - Retinal pigment epithelial phagocytosis and metabolism differ from those of macrophages. AB - The purpose of this study is to compare primary human retinal pigment epithelium (RPE) cells with respect to particle uptake and further processing steps with immunological phagocytes for a better understanding of the possible role of RPE cells in triggering autoimmune diseases in the eye. We investigated the similarities of human RPE and monocytes/macrophages studying the uptake of fluorescein- and europium-labeled synthetic microparticles and microbial pathogens by human and bovine RPE cultures and a permanent RPE cell line (CRL). The uptake was monitored by laser scanning microscopy, flow cytometry and time resolved fluorescence analysis; for comparison, macrophages and a macrophage-like cell line (MonoMac6) were used. A size-dependent uptake was seen in primary RPE cultures as well as in CRL, showing a preferential uptake of smaller beads followed by Staphylococcus aureus and Escherichia coli. Opsonization with serum caused a modest increase in bacteria uptake, but in contrast to macrophages, the classical complement receptors were not found on RPE cells. Living bacteria were also ingested in a time-dependent manner, but, as no intracellular overgrowth was observed, we further investigated the oxidative ability of RPE as a possible mechanism for microbial suppression. Unlike macrophages/granulocytes, no respiratory burst was detected in RPE cells, but, comparable to MonoMac6, IFN gamma induced neopterin in the human RPE. Interestingly a diurnal rhythm of phagocytosis was observed which was influenced by light exposure suggesting that RPE cells maintain their circadian rhythm also in cell culture to a certain extent. This study further demonstrates that in addition to similar phagocytic properties the RPE still shows substantial metabolic differences in comparison to blood-derived phagocytes. PMID- 15292659 TI - Glycosaminoglycans in human trabecular meshwork: age-related changes. AB - Glycosaminoglycans play a central role in maintaining the normal outflow resistance in the human trabecular meshwork. We evaluated the possible morphological, histochemical and morphometrical age-related changes in glycosaminoglycans of the trabecular meshwork. Small human samples were drawn from 24 eyes after exitus from young and old humans. Samples were harvested from the anterior chamber of the eye, without any aesthetic damage for the face. They were divided into three fragments, each used for morphological, histochemical and ultrastructural staining. Quantitative analysis of images was performed to evaluate morphometrical data that were statistically analysed. Our findings demonstrate the following age-related changes: (1) deposition of fibrous granular material in the trabecular meshwork; (2) increased electron density of the structures; (3) strong decrease in the hyaluronic acid content, and (4) increase in sulphated proteoglycans. Glycosaminoglycans of human trabecular meshwork undergo age-related changes, as demonstrated by our morphological, histochemical and morphometrical results. PMID- 15292661 TI - Expression of c-Fos and c-Jun in developing mouse lens. AB - We carried out immunohistochemistry in embryonic eye-specific antibodies in the mouse to determine the spatial/temporal expression pattern of c-Fos and c-Jun proteins--the main components of the AP-1 transcription factor, in lens epithelial cells during mouse lens morphogenesis. c-Fos protein expression was detected in the equatorial epithelium at E14.5-P25, with a peak at P1-P8. c-Jun protein expression was detected in equatorial lens epithelial cells at E18.5-P8, with a peak at P1. During these intervals, the anterior epithelium lacked expression of c-Fos and c-Jun. They were not detected in an adult mouse lens. The expression pattern of the AP-1 transcription factor in equatorial epithelium may suggest its role in fiber differentiation. PMID- 15292660 TI - Heat treatment enhances healing process of experimental pseudomonas corneal ulcer. AB - We investigated the effects of hyperthermia on the healing process of experimental Pseudomonas corneal ulceration (PCU). Hartley guinea pigs were used to develop animal models of PCU. As a heat source, disposable chemical pocket warmers were applied. The healing process of PCU was compared between the heat treated corneas and the control corneas. The severity of infection and the degree of angiogenesis were classified by a clinical scoring system. The animals were euthanized 14 days after infection and the corneas were submitted for histopathological examination. The expression of vascular endothelial growth factor (VEGF) was examined immunohistochemically. Comparative reverse transcription polymerase chain reaction was performed to measure the expression level of VEGF in the cornea. Hyperthermia significantly promoted corneal epithelization and neovascularization in the PCU model. Heat treatment significantly decreased the number of viable Pseudomonas organisms present in PCU. On immunohistochemistry, the heated cornea demonstrated more intense staining for VEGF. Comparative reverse transcription polymerase chain reaction showed upregulation of the expression level of VEGF mRNA in the heat-treated cornea. Hyperthermia accelerated the healing process of PCU with increased corneal neovascularization. Angiogenesis may play an important role in the PCU healing process, which is enhanced by the heat treatment. PMID- 15292662 TI - An alternative method of steroid-induced lens opacification in brown norway rat eyes applying systemic pulse administration. AB - PURPOSE: To confirm whether long-term administration of prednisolone sodium succinate (prednisolone) alone is able to induce cataract in rat eyes. MATERIALS AND METHODS: A 1% solution of prednisolone was administered topically as eye drops to Brown Norway rat eyes, and a systemic pulse administration of 10 mg/kg/day was given via the tail vein. Both administration methods were applied in different combinations. Eighty-three 6-week-old male rats were divided into 8 groups: group 1 = control; group 2 = topical instillation every day; group 3 = single pulse; group 4 = single pulse + eye drops; group 5 = 3 times pulse; group 6 = 3 times pulse + eye drops; group 7 = 3 times pulse per 2 months; group 8 = 3 times pulse per 2 months + eye drops. Observations for changes of lens transparency were made by slitlamp microscopy and documented by an anterior eye segment analysis system (Nidek EAS-1000) from the onset of drug administration to a maximum period of 16 months. RESULTS: Lens opacity in the shallow anterior and posterior lens layers developed from the tenth month following commencement of prednisolone administration. The incidence of anterior and/or posterior cortical cataract at the sixteenth month was 15% in group 2, 12.5% in group 5, 25% in group 6, 17.9% in group 7 and 35.3% in group 8. The lenses of groups 1, 3 and 4 maintained their transparency throughout the observation period. Light scattering intensity in groups 8 and 7 was the highest, followed by groups 6 and 5, then groups 2, 4, 3 and 1. CONCLUSION: Cortical cataract was successfully induced in Brown Norway rat eyes by sustained administration of prednisolone succinate alone applied as systemic pulse. PMID- 15292663 TI - Fluphenazine and its toxic maculopathy. AB - Ten years ago a 45-year-old female presented with bilateral maculopathy with visual acuity of 6/18 on the right and 6/24 on the left. She had been on fluphenazine for the past 10 years for schizophrenia. Investigation including fluorescein angiogram, colour vision assessment and electrophysiological testing confirmed the nature of the retinal damages. Her maculopathy deteriorated further despite discontinuation of the medication. A previous publication has reported maculopathy with fluphenazine in association with the welding arc injury, but that particular patient had not been exposed to welding flash or other extreme photochemical sources. We believe this is the first reported incidence that fluphenazine has directly caused a maculopathy secondary to its accumulation at the retinal pigment epithelium and its toxic effect. PMID- 15292664 TI - Subtle disruption of the middle cerebellar peduncles in patients with schizophrenia. AB - Diffusion tensor imaging (DTI) was used to investigate subtle disruption in the middle cerebellar peduncles in patients with schizophrenia. Fractional anisotropy (FA) was measured in 25 patients with schizophrenia and 21 healthy subjects using DTI. The FA of the right and left middle cerebellar peduncles was significantly lower in the schizophrenic patients compared to healthy subjects. FA in the left middle cerebellar peduncles was significantly correlated with the dosage of neuroleptics in patients with schizophrenia. There were no significant differences of mean diffusivity in the right and left middle cerebellar peduncles between patients with schizophrenia and healthy subjects. The findings of the study suggest that antipsychotics may improve the subtle disruption in the middle cerebellar peduncles in patients with schizophrenia. PMID- 15292665 TI - Linkage of M5 muscarinic and alpha7-nicotinic receptor genes on 15q13 to schizophrenia. AB - Most antipsychotic drugs act on the forebrain by blocking dopamine receptors. In rodents, the M5 muscarinic receptor (CHRM5) is important for prolonged dopamine release. We typed polymorphisms in CHRM5 and alpha7-nicotinic receptor (CHRNA7) genes on 15q13 in 82 Canadian families having at least 1 schizophrenic patient. Using the Family-Based Association Test, we performed haplotype analysis of the 2 loci and found biased transmission in schizophrenia (z = -2.651, p = 0.008). In the families tested, the 2 cholinergic genes interacted to affect schizophrenia in combination, while neither was sufficiently alone to confer susceptibility. Our present study provided the first line of direct evidence suggesting that the CHRM5 gene combined with the CHRNA7 gene may be linked to schizophrenia. PMID- 15292666 TI - Effects of Hypericum extract (LI160) on the change of auditory evoked potentials by cortisol administration. AB - Target symptoms treated with Hypericum extract, i.e. somatisation, fatigue and depression could be related to an increased activity of glucocorticoids in the brain. One potential mechanism is the increased permeability of the blood-brain barrier for glucocorticoids. Hypericum extract LI160 reduces intracerebral glucocorticoid concentration possibly by its action to induce the expression of the transport protein P-glycoprotein (P-gp). To test this hypothesis directly, we performed a randomised double-blind crossover study to examine the effect of intravenously administered cortisol on auditory evoked potentials (AEPs) and salivary cortisol concentration. Nineteen healthy subjects were treated for 2 weeks with 300 mg LI160 twice a day or placebo. On the 14th day, AEPs were recorded every 30 min, at times -60, -30 and 0 min before the start of the infusion and at +30, +60 and +90 min after starting the infusion. The rate of infusion was 20 mg cortisol/h. No changes in the AEP, especially the N1-P2 component, could be observed under cortisol infusion and consequently no modification with the treatment of Hypericum extract. The salivary concentration of cortisol under cortisol infusion was slightly but significantly decreased in the Hypericum condition compared to placebo. The results of the present study are therefore inconclusive with respect to the influence of LI160 treatment on the expected cortisol-induced AEP changes, but support the concept of an action of Hypericum on P-gp function by the observed changes in salivary cortisol. PMID- 15292667 TI - Consensus on the use of substituted benzamides in psychiatric patients. AB - The class of substituted benzamides includes compounds able to modulate dopaminergic neurons selectively and specifically. The first synthetic substituted benzamide was sulpiride, which has been replaced in the clinic by the more modern amisulpride. The compound is very selective for mesolimbic D2 and D3 receptors and, therefore, has a dual mechanism of action, which is associated with two different indications. At low doses (50 mg), amisulpride preferentially blocks presynaptic autoreceptors, producing an increase in dopamine release, and therefore acting as a dopaminergic compound able to resolve the dopaminergic hypoactivity that characterizes depression. At higher doses (400-1,200 mg), the drug exerts its activity on postsynaptic D3/D2 receptors located in the limbic region and prefrontal areas, producing selective dopaminergic inhibition, eliciting antipsychotic effects. In the present review, the clinical use of amisulpride in depressive syndromes is discussed, in particular in dysthymia and in schizophrenia. Based on experimental data, amisulpride is a treatment of choice for dopaminergic transmission disorders, both in depression and in schizophrenia. PMID- 15292668 TI - Should major depression with 'high normal' thyroid-stimulating hormone be treated preferentially with tricyclics? AB - In a prospective and naturalistic setting, two samples representing 209 depressed inpatients were assessed for thyroid functioning at admission before antidepressant treatment, and for depression before and after 1 month of antidepressant treatment. We hypothesized that serum thyroid-stimulating hormone (TSH) elevation > or = upper 25th percentile of the normal reference range is associated with poorer response to antidepressant therapy and differences between tricyclic antidepressants (TCA) and other antidepressants. Screening for mild thyroid failure defined as serum TSH concentrations > or = upper 25th percentile of the normal range may provide clues to the clinician. Such patients have a more severe form of depression and a slower or impaired response to antidepressant therapy. It is also possible that they would benefit preferentially from TCA rather than other antidepressants. PMID- 15292669 TI - Sleep in young adults with Asperger syndrome. AB - Asperger syndrome (AS) is a neurodevelopmental disorder belonging to autism spectrum disorders. Both children and adults with AS have subjective impairment in the initiation and continuity of sleep, and studies using objective assessment are sparse. Twenty young AS adults with frequent complaints of low sleep quality were compared to 10 age-, gender- and education-matched controls without sleep complaints using polysomnography and spectral power analysis of slow-wave sleep. AS subjects displayed a similar polysomnographic profile as compared with controls. In spectral power analysis, a statistically nonsignificant trend towards decreased relative delta power and increased theta power in slow-wave sleep was found in the AS group. It seems that nonorganic insomnia, due to anxiety inherent in AS, is responsible for the low sleep quality in these subjects. PMID- 15292670 TI - Association study of catechol-O-methyltransferase gene and dopamine D4 receptor gene polymorphisms and personality traits in healthy young chinese females. AB - Human personality traits, which are substantially heritable, may be modulated by monoamine neurotransmitters. It has been demonstrated that the catechol-O methyltransferase (COMT) Val158Met genetic polymorphism, a functional polymorphism that may affect monoamine metabolism, is possibly associated with specific personality traits. In addition, a polymorphism in the dopamine D4 receptor (DRD4) gene exon 3 has been associated in some, but not all, studies with the novelty seeking personality trait, as evaluated by the Tridimensional Personality Questionnaire (TPQ). In this study, associations between these two polymorphisms and TPQ personality traits were investigated in a sample population of 120 healthy young Chinese females. The results of this analysis reveal that the COMT Val158Met polymorphism was significantly associated with novelty seeking (p = 0.017) and reward dependence scores (p = 0.015) in our sample. However, no significant differences were demonstrated comparing TPQ-specific scores for subjects bearing different DRD4 genotypes. The present study suggests that the functional COMT Val158Met genetic polymorphism contributes to individual differences in the personality traits novelty seeking and reward dependence. Similar to the results of a recent meta-analytic review, however, no association was demonstrated between this DRD4 polymorphism and novelty seeking in our young Chinese female sample population. PMID- 15292671 TI - Low platelet-poor plasma levels of serotonin in adult autistic patients. AB - BACKGROUND: Hyperserotonemia has been reported in about a third of autistic patients. However, most studies have examined whole blood levels of serotonin (5 HT), the vast majority of which is found in platelets. The aim of this study was to determine 5-HT levels in platelet-poor plasma (PPP) in a group of adult patients with autism. METHODS: Levels of PPP 5-HT were compared between 10 adult drug-free autistic patients and 12 healthy controls. The Ritvo-Freeman Real-Life Rating Scale and the Overt Aggression Scale (OAS) were administered to the autistic group as a measure of symptom severity. RESULTS: Significantly lower PPP 5-HT levels were observed in the autistic group as compared to the controls (p = 0.03). In addition, PPP 5-HT levels were inversely correlated with OAS scores among subjects with autism (r = -0.64, p < 0.05). CONCLUSION: PPP 5-HT ('free') levels appear to be low in autistic patients and may play a role in the pathophysiology and symptomatology of the disorder. PMID- 15292672 TI - Cocaine addicts with conduct disorder are typified by decreased cortisol responsivity and high plasma levels of DHEA-S. AB - There is evidence that children with antisocial behaviors have increased plasma levels of the adrenal androgen dehydroepiandrosterone sulfate (DHEA-S) and either a decreased level of another adrenal steroid, cortisol, or a decreased cortisol responsivity to stress. Low levels of cortisol have also been reported in antisocial adults but their levels of DHEA-S have not been studied. The present study was designed to perform in adult cocaine addicts simultaneous assessments of DHEA-S and cortisol as a function of a diagnosis of antisocial personality disorder (adult symptoms) and of a retrospective diagnosis of conduct disorder (CD). Basal cortisol and DHEA-S were determined in the plasma samples of 40 hospitalized men. The patients' cortisol responsivity was also assessed while they were being exposed to a stressful situation. Patients who had a retrospective CD diagnosis had significantly increased DHEA-S levels and secreted less cortisol when stressed. Comparisons between patients who did and did not meet the antisocial personality disorder adult criteria did not reveal any significant difference in DHEA-S or in cortisol responsivity. This could be attributed to the nature of the criteria used to define the adult disorder, which focus mostly on a failure to conform to social norms, whereas a number of CD criteria involve displays of some degree of violence. In conclusion, adults who retrospectively qualified for a CD diagnosis had increased DHEA-S levels and a decreased cortisol reactivity, confirming observations made in children and indicating that mechanisms underlying adrenal steroid alterations in childhood could still be at play in adulthood. PMID- 15292673 TI - P300 event-related potential amplitude and impulsivity in cocaine-dependent subjects. AB - Previous studies report reduced amplitude of the P300 event-related potential in cocaine-dependent individuals. Cocaine dependence is also associated with increased impulsivity, possibly due to deficits in cognitive function that are associated with reduced P300 amplitude. In the current study, the relationship between cocaine dependence, impulsivity, and P300 amplitude were examined. An auditory oddball event-related potential task along with self-report (Barratt Impulsiveness Scale version 11) and behavioral laboratory (Immediate and Delayed Memory Task) measures of impulsivity were assessed in healthy controls (n = 14) and subjects who met DSM-IV criteria for current cocaine dependence (n = 17). P300 amplitude was reduced and self-reported and behavioral laboratory impulsivity scores were elevated among the cocaine-dependent group compared to controls. There was a positive correlation between the questionnaire and behavioral laboratory measures of impulsivity, and a negative correlation between impulsivity measures and P300 amplitude. The correlation between self-reported impulsivity scores and P300 amplitude remained after taking into account the number of childhood conduct disorder symptoms. This study supports the hypothesis that the basic neurophysiology responsible for the P300 amplitude in cocaine dependent individuals is associated with impulsivity independent of a history of childhood conduct disorder symptoms. PMID- 15292674 TI - Polymorphisms in the serotonin transporter and monoamine oxidase A genes and their relationship to personality traits measured by the Temperament and Character Inventory and NEO Five-Factor Inventory in healthy volunteers. AB - The associations between 5-HTT-linked polymorphic region (5-HTT-LPR), monoamine oxidase A (MAOA)-LPR and the dimensions of temperament evaluated using the Temperament and Character Inventory (TCI) and NEO Five-Factor Inventory (NEO-FFI) were studied. One hundred healthy volunteers (without psychiatric disorders) were recruited to represent a cross-section of the population of Szczecin (Poland) in terms of sex, age and education. No associations between 5-HTT-LPR and the TCI harm avoidance dimension and between 5-HTT-LPR and the NEO-FFI neuroticism dimension were found. Males carrying the 3-VNTR MAOA gene variant (209 bp) had significantly lower values on the NEO-FFI openness dimension (p = 0.039) and obtained higher scores on the subdimension 3 of the TCI reward dependence (RD3), i.e. attachment vs. detachment (p = 0.005). Individuals carrying the 'short' variant of 5-HTT-LPR had lower values on the reward dependence dimension and the RD4 subdimension (dependence vs. independence) than individuals not carrying the 'short' variant (p = 0.039 and p = 0.011, respectively). Females carrying the 'short' variant had lower values on NS1 (exploratory excitability vs. stoic rigidity) and RD4 (dependence vs. independence) than those not carrying the variant (p = 0.042 and 0.043, respectively). The obtained level of significance with respect to the observed associations between 5-HTT-LPR and the reward dependence scales and subscales and between 5-HTT-LPR and the NS1 subscale are too weak for further interpretation. Our results do not confirm the hypothesis that there is a simple correlation between single gene polymorphisms and a personality trait measured by the TCI and NEO-FFI scales. PMID- 15292675 TI - Long-term olanzapine treatment and p300 parameters in schizophrenia. AB - The well-known amplitude reduction of the P300 appears to be unaffected by the treatment with classical antipsychotics in schizophrenia, whereas the effects of atypical neuroleptics on this event-related potential are less understood. The study of these changes could help in deciding whether the P300 amplitude reduction in schizophrenia is a trait or state marker of that illness and in better describing the effect of atypical antipsychotics on altered cognitive functions. We present a prospective longitudinal study of P300 amplitude and latency before and after 6 months' treatment with olanzapine in 11 patients with schizophrenia. A healthy control group (n = 30) was also studied. Overall, no significant changes, either in amplitude or in latency as measured at Pz and Fz electrodes, were found when comparing the pre- and postolanzapine conditions, despite the overall improvement in positive and negative symptoms. Nevertheless a direct specific association was observed between a P300 amplitude increase with olanzapine and the improvement in negative symptoms. These data would suggest that P300 amplitude reduction in schizophrenia may be relatively independent from clinical state and treatment, thus constituting a trait marker of schizophrenia. Our data also suggest that, in addition to this, some further changes in P300 amplitude might depend on the clinical state of the patients. PMID- 15292676 TI - Individual trait anxiety levels characterizing the properties of zen meditation. AB - Meditation is a specific consciousness state in which deep relaxation and increased internalized attention coexist. There have been various neurophysiological studies on meditation. However, the personal predispositions/traits that characterize the properties of meditation have not been adequately studied. We analyzed changes in neurophysiological parameters [EEG coherence and autonomic nervous activity using heart rate variability (HRV) as an index] during Zen meditation, and evaluated the results in association with trait anxiety (assessed by Spielberger's State-Trait Anxiety Inventory) in 22 healthy adults who had not previously practiced any form of meditation. During meditation, in terms of mean values in all subjects, an increase in slow alpha interhemispheric EEG coherence in the frontal region, an increase in high frequency (HF) power (as a parasympathetic index of HRV), and a decrease in the ratio of low-frequency to HF power (as a sympathetic index of HRV) were observed. Further evaluation of these changes in individuals showed a negative correlation between the percent change (with the control condition as the baseline) in slow alpha interhemispheric coherence reflecting internalized attention and the percent change in HF reflecting relaxation. The trait anxiety score was negatively correlated with the percent change in slow alpha interhemispheric coherence in the frontal region and was positively correlated with the percent change in HF. These results suggest that lower trait anxiety more readily induces meditation with a predominance of internalized attention, while higher trait anxiety more readily induces meditation with a predominance of relaxation. PMID- 15292677 TI - Urine of patients with nephrotic syndrome contains the plasma type of PAF acetylhydrolase associated with lipoproteins. AB - BACKGROUND: Platelet-activating factor (PAF) is a proinflammatory phospholipid mediator involved in the pathogenesis of glomerulonephritis (GN). In plasma, PAF is hydrolyzed and inactivated by PAF-acetylhydrolase (PAF-AH), an enzyme associated with lipoproteins, mainly with the low-density lipoprotein. PAF-AH activity has been found in urine of patients with primary GN, however the source and type of urinary PAF-AH remain unknown. We characterized the type of PAF-AH excreted in the urine of patients with primary GN and studied the possible relationship of this enzyme with the lipiduria and proteinuria observed in these patients. METHODS: Eighteen patients with primary GN (8 with nephrotic syndrome (NS) and 10 with non-nephrotic range proteinuria (NNRP)) and 20 normolipidemic age- and sex-matched controls participated in the study. PAF-AH activity in plasma, in urine and in individual lipoprotein particles was determined by the trichloroacetic acid precipitation procedure, whereas the PAF-AH protein was detected by Western blotting analysis. Plasma and urine lipoproteins were fractionated by gradient ultracentrifugation and characterized by Western blotting analysis. RESULTS: Plasma PAF-AH activity was higher in NS patients compared with NNRP patients and controls, whereas the enzyme activity associated with high-density lipoprotein was significantly lower in both patient groups compared with controls. PAF-AH was detected only in the urine of NS patients. It was the plasma type of PAF-AH and was associated with lipoprotein particles. Enzyme activity was also positively correlated with urine cholesterol levels. CONCLUSION: Urine of NS patients contains the plasma type of PAF-AH, which is related to the extent of lipiduria and is associated with urine lipoproteins. PMID- 15292678 TI - Role of isoprenoids in cytoskeleton integrity and albumin endocytosis by opossum kidney cells. AB - BACKGROUND: The actin cytoskeleton has been increasingly implicated in endocytic events that are involved in the reabsorption of filtered protein by the proximal tubule. Isoprenylated small G proteins have emerged as key regulators of the actin cytoskeleton. This study examines the role of isoprenoid intermediates in organization of the cytoskeleton, and the effect of modification of the cytoskeleton on albumin endocytosis. METHODS: The effect of lovastatin on cytoskeleton morphology of opossum kidney cells (OK cells) was determined by staining with fluorescent isothiocyanate (FITC)-phalloidin followed by examination using confocal microscopy. Quantitative effects on albumin binding and uptake were determined using a well-established method. RESULTS: Inhibition of isoprenoid synthesis by lovastatin led to morphological disruption of the cytoskeleton with a concentration-dependent decrease in albumin uptake into OK cells. Addition of mevalonate, but not cholesterol, ameliorated the effects of lovastatin on the cytoskeleton and albumin uptake. Selective inhibition of isoprenoid intermediates with a farnesyltransferase inhibitor suggests that geranylgeranylated proteins mediate cytoskeleton organization. CONCLUSION: These results confirm the importance of cytoskeleton integrity in albumin endocytosis in renal proximal tubules. While the present data suggest that synthesis of isoprenoids via the mevalonate pathway is a critical step in maintenance of the cytoskeleton, the role of individual small G proteins in control of the cytoskeleton and other endocytic events is yet to be defined. PMID- 15292679 TI - Reliability of t7-based mRNA linear amplification validated by gene expression analysis of human kidney cells using cDNA microarrays. AB - Genome wide gene expression analysis by cDNA microarrays is often limited by minute amounts of starting RNA. We therefore tested an optimized linear RNA amplification protocol using the RiboAmp amplification kit in the setting of cDNA microarrays. We isolated mRNA from a human kidney cell line (HK-2; ATCC) and from Universal Human Reference RNA (STR; Stratagene). After performing one and two rounds of linear RNA amplification, respectively, the amplified RNAs were co hybridized to cDNA microarrays. Linearity and reproducibility of the individual experiments were then assessed by calculating the Pearson correlation. The intra amplification consistency showed a correlation of 0.968 for the first round, 0.907 for the second round and 0.912 for two successive rounds of amplification. If the first round was compared to unamplified material, r was 0.925. The second round amplification yielded a correlation of 0.897 if compared to unamplified mRNA. Two rounds of amplification starting from 200 pg of mRNA compared to unamplified material resulted in a correlation of 0.868. These results indicate that linear amplification using RiboAmp kit yields amplified RNA with a high degree of linearity and reproducibility. PMID- 15292680 TI - Expression of osteopontin in cisplatin-induced tubular injury. AB - Osteopontin (OPN) is considered as a key protein in cell regeneration. OPN is thought to have many functions in cell-cell binding and cell-matrix binding via OPN receptors in various organs. But there is little information on the precise role of OPN. To clarify the functional role of OPN in tubular injury, we performed in situ hybridization and immunohistochemical analysis of the expression of OPN in a renal cortical necrosis model induced by cisplatin from the acute injury to late recovery phases. In the acute injury phase, both mRNA and protein of OPN were markedly induced in damaged tubular lumens with cell debris. In the late recovery phase, on the other hand, OPN protein and mRNA were observed in dilated and flattened tubular epithelial cells showing a regenerative appearance. Most of these cells were also immunostained with CD44, a receptor of OPN. PCNA staining was also co-localized with these expressions. In light of the CD44 function regulating cell proliferation, these findings suggest that OPN may contribute to regeneration of tubular epithelial cells during the acute to late recovery phases of cortical tubular damage induced by cisplatin. PMID- 15292681 TI - Expression of nuclear pore complex oxalate binding protein p62 in experimental hyperoxaluria. AB - Proteins are thought to play a major role in stone formation. Oxalate binding protein plays a vital role in the transport of oxalate. This study was aimed at determining whether hyperoxaluria induces the expression of nuclear pore complex oxalate binding protein p62 which has the transport function. Hyperoxaluria was induced in male Wistar rats by feeding 0.75% ethylene glycol in water. The oxalate binding activity of the nuclear pore complex protein increased markedly during experimental hyperoxaluria, whereas nuclear lamina had no binding at all. There was an alteration in the elution profile of the nuclear pore complex oxalate binding protein during the hyperoxaluric condition. The protein was purified and had a molecular weight of 62 kDa (data not shown). The purified protein showed cross-reactivity with the monoclonal antibody (MAb 414) and it showed homogeneity. The expression of this protein (p62) during the hyperoxaluric condition was determined by ELISA and a 3-fold increase was observed when compared to control rats. The increased expression is further confirmed by Western blotting and immunohistochemistry. The increase in p62 protein expression may be either due to increased expression of certain genes or degradation of the cell membrane by oxalate-induced cell injury. Thus, the present study suggests that the increased expression of this protein (p62) may be due to the oxalate induction. PMID- 15292682 TI - Leptin and anorexia in renal insufficiency. PMID- 15292683 TI - Anorexia and serum leptin levels in hemodialysis patients. AB - BACKGROUND AND AIMS: Hyperleptinemia is a common feature in hemodialysis (HD) patients. However, the role of increased serum leptin levels in the pathogenesis of HD-related anorexia is still controversial. The purpose of the present prospective study was to ascertain whether hyperleptinemia is causally implicated in the pathogenesis of HD-related anorexia. METHODS: We measured the serum leptin levels and the serum leptin/body mass index (BMI) ratio in 24 healthy subjects and in 49 end-stage renal disease patients on maintenance HD. HD patients were subdivided into anorexic (14/49, 28.5%) and non-anorexic (35/49, 71.5%) according to a questionnaire discriminating for the presence of anorexia-related symptoms. RESULTS: Calorie (kcal/kg/day) and protein (g/ kg/day) intakes were significantly lower in anorexic than in non-anorexic patients (20.1 +/- 1.1 vs. 27.9 +/- 1.3, p = 0.004, and 0.82 +/- 0.05 vs. 1.19 +/- 0.05, p = 0.001, respectively). Accordingly, serum albumin, total lymphocyte count, mid-arm muscle circumference, and the protein equivalence of nitrogen appearance (PNA) were significantly lower in anorexic patients. The serum leptin concentration (ng/ml) was significantly higher in HD patients than in controls, in males (15.33 +/- 3.4 vs. 3.7 +/- 0.3, p = 0.003) and in females (42.3 +/- 7.2 vs. 10.5 +/- 1.3, p = 0.03). Similarly, serum leptin/BMI ratio was significantly higher in HD patients than in controls, in males (0.56 +/- 0.1 vs. 0.16 +/- 0.02, p = 0.0028) and in females (1.8 +/- 0.2 vs. 0.4 +/- 0.04, p < 0.0001). However, serum leptin levels were similar in anorexic and in non-anorexic patients, in males (15.3 +/- 5.6 vs. 16.9 +/- 4.2, p = 0.85) and in females (46.6 +/- 12.9 vs. 47.4 +/- 9.4, p = 0.96). No differences were observed between the 2 groups in the serum leptin/BMI ratio, in males (0.59 +/- 0.2 vs. 0.58 +/- 0.14, p = 0.92) and in females (1.5 +/- 0.4 vs. 1.8 +/- 0.3, p = 0.94). Similarly, no statistically significant differences in terms of serum leptin levels and leptin/BMI ratio were observed between patients with dietary energy intake of <30 or > or =30 kcal/kg/day and between those with a dietary protein intake of <1.2 or > or =1.2 g/kg/day. No significant correlations were found between serum leptin levels and PNA, albumin, cholesterol, total lymphocytes number, weight change, C-reactive protein, fibrinogen, ferritin, and complement. CONCLUSION: The present results indicate that mechanisms other than increases in serum leptin levels might be involved in the pathogenesis of HD related anorexia. PMID- 15292684 TI - Pharmacokinetics of bupropion and its metabolites in haemodialysis patients who smoke. A single dose study. AB - To date, no study has investigated the effects of bupropion (BP) in renal impaired humans. This study aims to identify the pharmacokinetics of BP and metabolites in haemodialysis patients who smoke, determine whether haemodialysis affects BP and metabolite clearance, and suggest the BP dose in haemodialysis. The pharmacokinetics of BP and two of its major metabolites, hydroxybupropion (HB) and threohydrobupropion (TB) were studied in 8 smokers with ESRD receiving haemodialysis. Following a single oral dose of 150 mg bupropion hydrochloride sustained-release, blood samples were taken over 7 days, which were assayed using HPLC-mass spectrometry. Pharmacokinetic analysis was undertaken by non-linear regression using MWPharm. The BP results were similar to those for individuals with normal renal function. The metabolites demonstrated increased areas under the curve, indicating accumulation. Dialysis clearance of HB is unlikely. The results suggest significant accumulation of the metabolites in renal failure. Clarification of the clinical importance of the metabolites and toxic plasma levels is required. The effects of haemodialysis on BP and metabolites require further study. A dose of 150 mg bupropion every 3 days in patients receiving haemodialysis is more appropriate than the current manufacturer's recommendation (in renal impaired patients) of 150 mg daily. A multi-dose study is required. PMID- 15292685 TI - Enhancement of quality of life with adjustment of dry weight by echocardiographic measurement of inferior vena cava diameter in patients undergoing chronic hemodialysis. AB - BACKGROUND/AIMS: Ideal dry weight (DW) can serve as a marker of good quality of life (QOL) in patients receiving chronic hemodialysis. The size of the inferior vena cava (IVC) reflects the intravascular fluid status, and the diameter of IVC correlates indirectly with DW in these patients. Adjusting DW using echocardiographic measurement of the diameter of the IVC thus may be useful in maintenance of a better QOL in patients receiving chronic hemodialysis. METHODS: This study included 119 patients with ages ranging between 27 and 90 years (mean +/- standard deviation of 58.3 +/- 12.8). All of the patients received the IVC diameter (IVCD) measurement by echocardiography every 2 months for 1 year. The study group included 68 patients in whom the DW were adjusted by echocardiographic measurement of the IVCD, while the control group included 51 patients in whom the DW was adjusted by the conventional method. QOL was evaluated using the short form 36 questionnaire (SF-36) at the beginning and at the end of the study. Besides, the Kt/V(urea) value, a parameter of total urea clearance, was measured at the beginning and at the end of the study in patients of both groups. RESULTS: The scores of physical functioning, physical role functioning, general health and physical component summary showed a prominent improvement in the study group but not in the control group. The impact of periodic echocardiographic evaluation also demonstrated a significant change in the scores of physical functioning, physical role functioning and physical component summary in the study group. Furthermore, the Kt/V(urea) value, a parameter of total urea clearance, also increased significantly in the study group. CONCLUSION: Ideal DW is better adjusted by periodic echocardiographic measurement of the IVCD in patients undergoing chronic hemodialysis. Maintenance of a better DW leads to improve hemodialysis quality and QOL in these patients. PMID- 15292686 TI - Measuring blood pressure in stable renal transplant recipients: what you measure depends on what you use. AB - BACKGROUND: Blood pressure levels have a major impact on cardiovascular and renal transplant outcomes after renal transplantation. But there are significant challenges to accurately measure blood pressure levels in stable healthy renal transplant outpatients. We aimed to test whether there are differences in BP measurements taken using either automated oscillometric machines or a random zero sphygmomanometer. METHODS: Blood pressure was recorded twice in a random order with each of three BP measuring devices (DINAMAP BP8800; OMRON HEM 713 and a Random-Zero Hawskley sphygmomanometer). Results were analysed to determine observer bias, cardiovascular artefacts, and intra- and inter-machine BP variation. RESULTS AND CONCLUSIONS: There was no significant observer bias or cardiovascular artefacts. Intra-machine variability was small. BP measurement using DINAMAP and OMRON could lead to a difference of up to 30 mm Hg higher or 15 mm Hg lower than Hawskley random zero BP readings. Though widely used for 'convenience', oscillometric measures of BP in the renal transplant clinic are not optimal. PMID- 15292687 TI - Proatrial natriuretic peptide (1-98), but not cystatin C, is predictive for occurrence of acute renal insufficiency in critically ill septic patients. AB - INTRODUCTION: N-terminal prohormone of atrial natriuretic peptide ((proANP(1-98)) has been extensively analyzed in patients with chronic renal failure. It has been found to be closely related to the renal function and to interdialytic hydration status. The clinical relevance of proANP(1-98) and cystatin C, a novel marker of glomerular filtration, has not been investigated in the subgroup of critically ill septic patients with no history of chronic renal impairment. METHODS: We measured plasma level ofproANP(1-98) and cystatin C in 29 critically ill septic patients on admittance to the surgical intensive care unit and correlated it with the occurrence of acute renal failure. RESULTS: The proANP(1-98) plasma level was significantly higher in the group of patients who developed renal failure (12,722 +/- 12,421 vs. 2,801+/- 2,023 fmol/ml, p < 0.05). Multiple regression analysis shows that proANP(1-98) on the first day in the intensive care unit has a superior predictive value for the occurrence of renal failure to diuresis, calculated creatinine clearance or cystatin C (r = 0.42, p < 0.039). proANP(1-98) is also higher in non-survivors (9,303.8 +/- 11,053 vs. 2,448.5 +/- 1,803 fmol/ml, p < 0.018). CONCLUSION: proANP(1-98) is possibly a better predictor of acute renal failure to calculated creatinine clearance or diuresis among critically ill septic patients. Cystatin C was not correlated with occurrence of acute renal failure in this subgroup of patients. PMID- 15292688 TI - Elevated epsilon-(gamma-glutamyl)lysine in human diabetic nephropathy results from increased expression and cellular release of tissue transglutaminase. AB - INTRODUCTION: Diabetic nephropathy (DN) is the leading cause of chronic kidney failure, however the mechanisms underlying the characteristic expansion of the extracellular matrix (ECM) in diabetic kidneys remain controversial and unclear. In non-diabetic kidney scarring the protein crosslinking enzyme tissue transglutaminase (tTg) has been implicated in this process by the formation of increased epsilon-(gamma-glutamyl)lysine bonds between ECM components in both experimental and human disease. Studies in db+/db+ diabetic mice and in streptozotocin-treated rats have suggested a similar mechanism, although the relevance of this to human disease has not been addressed. METHODS: We have undertaken a retrospective analysis of renal biopsies from 16 DN patients with type 2 diabetes mellitus using an immunohistochemical and immunofluorescence approach, with tTg and epsilon-(gamma-glutamyl)lysine crosslink quantified by confocal microscopy. RESULTS: Immunofluorescent analysis of human biopsies (confocal microscopy) showed increases in levels of tTg (+1,266%, p < 0.001) and epsilon-(gamma-glutamyl)lysine (+486%, p < 0.001) in kidneys with DN compared to normal. Changes were predominantly in the extracellular periglomerular and peritubular areas. tTg staining correlated with epsilon-(gamma-glutamyl)lysine (r = 0.615, p < 0.01) and renal scarring (Masson's trichrome, r = 0.728, p < 0.001). Significant changes in epsilon-(gamma-glutamyl)lysine were also noted intracellularly in some (< or =5%) tubular epithelial cells. This is consistent with cells undergoing a novel transglutaminase-mediated cell death process in response to Ca2+ influx and subsequent activation of intracellular tTg. CONCLUSION: Changes in tTg and epsilon-(gamma-glutamyl)lysine occur in human DN. Cellular export of tTg may therefore be a factor in the perpetuation of DN by crosslinking and stabilisation of the ECM, while intracellular activation may lead to cell death contributing towards tubular atrophy. PMID- 15292689 TI - Renal functional reserve evolution in children with a previous episode of hemolytic uremic syndrome. AB - INTRODUCTION: Glomerular filtration rate (GFR) is the most widely used indicator of kidney function in patients with renal disease, although it does not invariably reflect functional status after renal injury. The concept of renal functional reserve (RFR) as the ability of the kidney to increase GFR following a protein load was introduced in the 1980s. In this study we evaluated the RFR test in 26 children who had developed hemolytic-uremic syndrome (HUS) at least 2 years before the first evaluation, then 8 years later. At the beginning of the study they had no signs of proteinuria, hypertension or renal insufficiency. RFR was also evaluated in 15 healthy control children. METHODS: Proteinuria and creatinine in serum and urine were tested. Functional reserve index (FRI) was defined in order to evaluate RFR. Patients with FRI level >1.36 were considered as responders (R) and with FRI <1.36 as non-responders (NR). RESULTS: R and NR groups failed to show any significant differences when basal creatinine clearance (C(Cr)) was evaluated. The NR group presented a significant low initial FRI that persisted unchanged at the end of the study. These patients developed proteinuria and a renal protector treatment with protein restriction was indicated. Although the proteinuria diminished, it remained within pathological range. The lack of RFR response in the NR group was significantly related to the presence of oliguria lasting longer than 8 days during the acute phase of disease. CONCLUSIONS: Those patients with a previous history of HUS with normal basal C(Cr) should be evaluated by the RFR test to detect those at risk of developing glomerular hyperfiltration. PMID- 15292690 TI - CT in the differentiation of pancreatic neoplasms--progress report. AB - Today, computed tomography (CT) is the most commonly used imaging method in the assessment of pancreatic tumors. The sensitivity of CT in detection of pancreatic tumors is more than 90% when direct and indirect signs are used for diagnosis. However, the potential to differentiate exocrine (non-endocrine) tumors of the pancreas is limited. CT is used in these lesions to perform an adequate staging, especially for surgical purposes. The operative resectability, primarily in regard to vessels, lymph node metastasis and hepatic metastasis, has to be assessed. Keeping in mind the limitations of this macromorphological imaging procedure, CT has the best reproducibility and overall accuracy of all imaging methods. Using multislice CT it is possible to perform non-axial reconstructions with high resolution. In functional endocrine tumors, multislice spiral CT will enhance the diagnostic capabilities, since the whole organ can be examined in thin slices, with high resolution during the rather short arterial phase of the contrast medium. Since some endocrine tumors are hypovascular, a scan during the portovenous phase is recommended too. The diagnosis of benign pancreatic tumors, like serous cystadenoma and pancreatic lipomas, is addressed. The most important pseudotumors of the pancreas are discussed. PMID- 15292691 TI - ERCP and MRCP in the differentiation of pancreatic tumors. AB - The introduction of endoscopic retrograde cholangiopancreatography (ERCP) in the early 1970s provided gastroenterologists with a number of diagnostic as well as therapeutic possibilities for examining biliopancreatic systems. In the meantime, magnetic resonance cholangiopancreatography presents a non-invasive alternative to diagnostic ECRP providing the advantage of a lower rate of possible complications. This article addresses the two methods presently available for differentiating pancreatic tumors. The objective of this article is to describe the advantages and disadvantages as well as the possibilities inherent in both methods. PMID- 15292692 TI - Role of endoscopic ultrasound in the diagnosis of patients with solid pancreatic masses. AB - Endoscopic ultrasound (EUS) is the most sensitive imaging procedure for the detection of small solid pancreatic masses and is accurate in determining vascular invasion of the portal venous system. Even compared to the new CT techniques, EUS provides excellent results in preoperative staging of solid pancreatic tumors. Compared to helical CT techniques, EUS is less accurate in detecting tumor involvement of the superior mesenteric artery. EUS staging and EUS-guided FNA can be performed in a single-step procedure, to establish the diagnosis of cancer. There is no known negative impact of tumor cell seeding due to EUS-guided fine needle aspiration (FNA). Without FNA, EUS and additional methods are not able to reliably distinguish between inflammatory and malignant masses. PMID- 15292693 TI - Echo-enhanced sonography--an increasingly used procedure for the differentiation of pancreatic tumors. AB - Echo-enhanced sonography is increasingly being used for differential diagnosis of pancreatic tumors. Ductal carcinomas are often hypovascularized compared with the surrounding tissue. On the other hand, neuroendocrine tumors are hypervascularized lesions. Tumors associated with pancreatitis have a different vascularization pattern depending on inflammation and necrosis. Cystadenomas frequently show many vessels along the fibrotic strands. Pancreatic tumors have a different vascularization pattern in echo-enhanced sonography. These characteristics can be used for differential diagnosis. However, histology is the standard of reference. PMID- 15292694 TI - CT Scan and MRI in the Differentiation of Liver Tumors. AB - Computed tomography (CT) and magnetic resonance imaging (MRI) are useful for detection and characterization of liver tumors, and widely used in clinical practice. By using dynamic CT and MRI with extracellular contrast material or MRI with liver-specific contrast material, we can investigate the morphological, hemodynamical and functional nature of focal hepatic lesions. Tumors often show characteristic findings on CT and MRI. Thus, we can correctly establish a diagnosis of liver tumors based on those findings. In this article, we describe the role of CT scan and MRI in the differentiation of liver tumors as well as presenting some typical CT and MR images. PMID- 15292695 TI - Imaging of inflammatory bowel disease: CT and MR. AB - Cross-sectional imaging has come to play a central role in the imaging of the abdomen. Concurrent to this, the role of CT and MRI in the imaging of inflammatory bowel disease has also increased in importance. These modalities offer numerous advantages over more traditional methods of radiologic diagnosis, and provide essential information not only for initial diagnosis, but for management, follow-up and detection of potential complications. On the horizon are several derivative techniques involving CT and MRI, potentially in combination with PET imaging; these may further improve the specificity and sensitivity of imaging modalities for diagnosis of inflammatory bowel disease. PMID- 15292696 TI - Doppler sonography in the diagnosis of inflammatory bowel disease. AB - Over the past few years, thanks to its ability to reveal neovascularization and inflammatory hyperemia, Doppler sonography has proved to be a valuable method for the assessment of disease activity in inflammatory bowel disease. Hypervascularization has been detected by Doppler imaging both in splanchnic vessels, in terms of flow volume and velocity, or resistance and pulsatility index on spectral analysis, and in small vessels of the affected bowel wall in terms of vessel density. In particular, power Doppler has been shown to be a highly sensitive method for evaluating the presence of flow in vessels that are poorly imaged by conventional color Doppler, and in detecting internal fistulas complicating Crohn's disease. Recently, the use of ultrasound contrast agents, such as Levovist, has been shown to improve the image quality of color Doppler by increasing the backscattered echoes from the desired regions, thus making it possible to better monitor the response to treatment and discriminate between active inflammatory and fibrotic bowel wall thickness in Crohn's disease. Additionally, Levovist-enhanced power Doppler sonography has proved to be highly sensitive and specific in the detection of inflammatory abdominal masses associated with Crohn's disease. In clinical practice, used in combination with second harmonic imaging and new generations of stable contrast agents, Doppler sonography appears to be a non-invasive and effective diagnostic tool in the diagnosis and follow-up of Crohn's disease and ulcerative colitis. PMID- 15292697 TI - Evaluation of criteria for the activity of Crohn's disease by power Doppler sonography. AB - BACKGROUND/AIM: In recent years, power Doppler sonography has been proposed as a method to assess disease activity in patients with Crohn's disease. The aim of this prospective study was to evaluate diagnostic criteria for power Doppler sonography by blinded comparison with ileocolonoscopy. METHODS: Twenty-two patients with confirmed Crohn's disease were prospectively investigated with B mode and power Doppler sonography (HDI 5000, Philips Ultrasound) as well as ileocolonoscopy. Sonography was performed within 3 days before endoscopy. All procedures were performed by experienced examiners who were blinded to the clinical data and other results. Defined ultrasound parameters (bowel wall thickness, vascularization pattern) were used to determine a sonographic score of the activity. The degree of activity was scored from 1 (none) to 4 (high) by both ultrasound and ileocolonoscopy (pattern, extent of typical lesions). For each patient all segments of the colon and the terminal ileum were evaluated by both ultrasound and endoscopy. The weighted kappa test was used (StatXact software) for statistical analysis. RESULTS: In total, 126 bowel segments were evaluated by both ultrasound and endoscopy. The study showed a high concordance of power Doppler sonography and ileocolonoscopy (weighted kappa by region: sigmoid colon: 0.81; transverse colon: 0.78; ascending colon: 0.75; cecum: 0.84; terminal ileum: 0.82). Highest concordance was found in the descending colon (weighted kappa: 0.91; 95% CI: 0.83-0.98). CONCLUSIONS: Combination of B-mode and power Doppler sonography has a high accuracy in the determination of disease activity in Crohn's disease when compared to ileocolonoscopy. The diagnostic criteria established in this study can be useful for the evaluation of inflammatory bowel diseases by ultrasound. PMID- 15292698 TI - Differential diagnosis of focal liver lesions in signal-enhanced ultrasound using BR 1, a second-generation ultrasound signal enhancer. AB - AIM: The aim was to evaluate the diagnostic value of contrast-enhanced ultrasound in the differential diagnosis of focal liver lesions, in a blinded experiment. In clinical routine the examiner can generally be influenced by the patient's history. METHOD: 62 patients with focal liver lesions, which could not be clearly differentiated and diagnosed by conventional ultrasound, were examined with contrast-enhanced (BR1, SonoVue, Bracco) ultrasound and included in a blinded prospective and randomized study. The examinations performed on a Sequoia 512 (Acuson) in a coherent contrast imaging method were recorded by an S-VHS recorder and afterwards analyzed by an examiner who did not know the patient's history. The basis of the diagnosis was the dynamic appearance and enhancement of the ultrasound contrast enhancer in different phases of liver perfusion. The conformation of the diagnoses was made by corresponding reference methods, as computer tomography, magnetic resonance imaging, biopsy and clinical follow-up. RESULTS: The following diagnoses were confirmed by reference methods: 18 patients with metastases, 4 hepatocellular carcinomas, 19 haemangiomas, 6 focal nodular hyperplasias, 13 patients with focal fatty infiltration and 2 patients with focal fatty sparing. 59 out of 62 patients with one or more liver lesions were correctly diagnosed by contrast-enhanced ultrasound. CONCLUSION: Second generation ultrasound contrast enhancers improve the differential diagnosis of benign and malignant liver lesions considerably, especially in a blinded study. PMID- 15292699 TI - Evaluation of diagnostic criteria for liver metastases of adenocarcinomas and neuroendocrine tumours at conventional ultrasound, unenhanced power Doppler sonography and echo-enhanced ultrasound. AB - PURPOSE: In order to improve the differential diagnosis between liver metastases of neuroendocrine tumours and adenocarcinomas, criteria for the masses at conventional ultrasound, unenhanced power Doppler sonography and echo-enhanced ultrasound were evaluated. METHODS: Seventy-three patients with histologically proven liver metastases of a neuroendocrine tumour (n = 26) or an adenocarcinoma (n = 47) were investigated by conventional ultrasound as well as unenhanced power Doppler sonography and echo-enhanced ultrasound focusing on specific properties of the lesions. RESULTS: Liver metastases of neuroendocrine tumours and adenocarcinomas showed a different contrast behaviour with echo-enhanced sonography. A hypervascularisation at the arterial and capillary phase were found in 85% of the neuroendocrine metastases, and in 17% of the masses of adenocarcinomas, respectively (p < 0.05). CONCLUSIONS: The successful treatment of liver metastases requires a highly sensitive and specific diagnostic procedure for their differentiation. A hypervascularisation of the lesions during the arterial and capillary phase at echo-enhanced ultrasound may point to a neuroendocrine primary tumour. However, histology is the only standard of reference for the differentiation of liver metastases, and is necessary for optimal therapy. PMID- 15292700 TI - A community-based epidemiologic study on gallstone disease among type 2 diabetics in Kinmen, Taiwan. AB - BACKGROUND: This study was conducted to assess the prevalence and associated risk factors of gallstone disease (GSD) among type 2 diabetics in Kinmen, Taiwan. METHODS: Based on a total of 858 type 2 diabetics ascertained in 1991-1993, an ultrasound sonography screening was performed by a panel of specialists in 2001. A total of 440 (51.3%) subjects were examined. RESULTS: Sixty-three out of 440 type 2 diabetics were diagnosed with GSD. The overall prevalence of GSD was 14.4%, including single stone 8.0% (n = 35), multiple stones 3.2% (n = 14), and cholecystectomy 3.2% (n = 14). The significant risk factors of GSD based on multiple logistic regression analysis were age (OR = 1.06, 95% CI: 1.02-1.10) and BMI (OR = 1.11, 95% CI: 1.01-1.22). CONCLUSIONS: Our results found that older age and higher BMI may increase the risk of developing GSD in type 2 diabetics. PMID- 15292701 TI - Spoke-like vascular pattern--a characteristic sign of focal nodular hyperplasias in the liver. PMID- 15292702 TI - Taking care of business: self-help and sleep medicine in american corporate culture. AB - This article argues that corporate management in the United States has expanded its scope beyond office walls and encompasses many aspects of workers' daily lives. One new element of corporate training is the micromanagement of sleep; self-help books, newspaper reports, magazine articles, and consulting firms currently advise workers and supervisors on optimizing productivity by cultivating certain sleep habits. Although consultants and self-help books make specific recommendations about sleep, most medical research is inconclusive about sleep's benefits for human performance. Using the ideas of Michel Foucault and Gilles Deleuze as a philosophical backdrop, this article examines the complex and often contradictory links between self-help, medicine, and corporate governance. PMID- 15292703 TI - Disorders of desire: addiction and problems of intimacy. AB - This essay investigates the tensions produced by the categorization of different forms of excessive desire under the singular model of addiction, and it challenges the increasing acceptance of addiction as an all-purpose explanation for unruly desires through a comparison of the different forms of disordered desire in sex addiction and alcoholism. Moreover, it argues for a broad understanding of addictive processes to undermine the normative and moralizing assumptions of addiction discourses. Refiguring addiction as a kind of intimacy is one way of making sense of the intense relationships people can develop with substances and with activities. PMID- 15292704 TI - Of rats and women: fetal sexuality and hybrid agency. AB - This paper investigates the way in which the sexuality of women has been posited in relation to rats as experimental subjects, exploring the stakes of a scientific debate that takes the social world of female sexuality as its focus and as a political problem. Studies that purport to understand female sexuality by investigating rat behavior rely on problematic assumptions about sovereign agents motivating sexual behavior. Such studies also aim to do away with so called deviant sexual behaviors and, as a consequence, gay people. Theories of agential realism and hybridity serve as counterforces to these inherently repressive perspectives by insisting on the multiple determinations of sexuality and subjectivity among women. PMID- 15292705 TI - Detached, as in a theater, I watch. PMID- 15292706 TI - Preliminary evaluation in vitro of the inhibition of cell proliferation, cytotoxicity and induction of apoptosis by 1,4-bis(1-naphthyl)-2,3-dinitro-1,3 butadiene. AB - The pattern of inhibition of cell proliferation and cytotoxicity in vitro by 1,4 bis(1-naphthyl)-2,3-dinitro-1,3-butadiene (Naph-DNB) was evaluated by the 3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and the trypan blue (TB) dye exclusion assays in nine murine and human cell lines of different histologic origin. In our culture conditions Naph-DNB showed a good inhibiting activity against all cell lines tested, with IC(50)s varying within a narrow micromolar range of concentrations (2.0 +/- 0.2-14.3 +/- 2.3 microM). In particular, murine P388 (leukemia), human Jurkat (leukemia), A2780, PA-1 (ovarian carcinoma) and Saos-2 (osteosarcoma) cells showed the highest sensitivity to the inhibiting potential of Naph-DNB, while human A549 (non small cell lung cancer, NSCLC), MDA-MB-231 (breast cancer), HGC-27 (gastric cancer) and HCT-8 (colon carcinoma) were the least sensitive cell lines. Moreover, the analysis of cytotoxicity of Naph-DNB evaluated by the TB test showed that this compound was able to kill cells with IC(50)s ranging from 1.7 to 39.2 microM. The study of the induction of apoptosis was carried out by 4'-6-diamidine-2'-phenylindole (DAPI) staining of segmented nuclei, western blot of p53 protein and TdT-mediated dUTP biotin nick end labeling (TUNEL) method, while the interaction with DNA was evaluated through the analysis of interstrand cross-link (ISCL) formation. Our data show that in all cell lines tested Naph-DNB was able to form ISCLs, to upregulate p53 oncosuppressor-protein and to induce apoptosis. Moreover, TUNEL analysis also suggested that Naph-DNB, similarly to other anticancer drugs, was able to block cells in the G (0)/ G (1) phase of the cell cycle. In conclusion our data suggest that Naph-DNB may be an effective novel lead molecule for the design of new anticancer compounds. PMID- 15292707 TI - Antineoplastic potency of arylchloroethylurea derivatives in murine colon carcinoma. AB - In a search for new antineoplastic agents the lead compound N-(4-tert butylphenyl)-N'-(2-chloroethyl)urea (CEU-22) of a series of 1-aryl-3-(2 chloroethyl)ureas and its iodinated bioisostere CEU-98, were previously selected on the basis of their cytotoxicity and the potent tropism for the intestinal tract (evidenced for CEU-22). In this study, we investigated the antitumour profile of these two drugs for the indication of colon cancer. In vitro, we found that micromolar concentrations of both CEU-22 and CEU-98 inhibited proliferation of DLD-1, Caco-2, HT-29, SW-948 and CT-26 lines. In vivo, a high inhibition of tumour growth and a life span increase were observed when BALB/c mice grafted subcutaneously with CT-26 cells received 5 daily intratumoural injections of each drug. When administered by the intraperitoneal route according to an intermittent schedule starting Day 1 or Day 7 post-implant, only CEU-98 demonstrated antitumour activity ( T / C = 29% for the Day-1,5,9-treatment versus 40% for the Day-7,11,15-treatment) and a life span increase around 40% for the two protocols. These results make CEU-98 a candidate for further investigations with a view to developing an efficacious treatment of colorectal cancer. PMID- 15292708 TI - Growth inhibition, G(1)-arrest, and apoptosis in MCF-7 human breast cancer cells by novel highly lipophilic 5-fluorouracil derivatives. AB - In this study we evaluate the antitumour activity, the cell cycle arrest and apoptotic properties of novel lipophilic benzene-fused seven-membered 5 fluorouracil (5-FU) analogs in comparison to 5-FU on MCF-7 human breast cancer cells. The lipophilicities of ESB-786B, ESB-252A and ESB-928A were predicted by using the CDR option of the PALLAS 2.0 program. Cytotoxic assays were evaluated in MCF-7 cells treated with the sulforhodamine B colorimetric method. Cell cycle perturbations were studied by flow cytometry. Apoptosis was determined by both DNA fragmentation and annexin V-FITC and propidium iodide staining. The novel derivatives were more lipophilic than 5-FU and induced a marked growth inhibition, in a dose-dependent manner. After treatment with IC(50) value (ranged from 2.5 to 22 microM) for each compound, light microscopy observation showed modifications in the morphology of MCF-7 cells. In addition, the 5-FU analogs arrest cells in the G(0)/G(1) phase of the cell cycle whereas 5-FU induced arrest in S-phase. Moreover, induction of apoptosis was demonstrated by the annexin-V based assay and confirmed using DNA fragmentation analysis on MCF-7 cells, a cell line in which the induction of DNA laddering is very difficult. The novel benzannelated seven-membered 5-FU analogs can be considered as specific apoptotic inducers. These experimental findings provide support for the use of these novel compounds as new weapons in the fight against breast cancer. PMID- 15292709 TI - Inhibitory effect of polypeptide from Chlamys farreri on UVA-induced apoptosis in human keratinocytes. AB - Polypeptide from Chlamys farreri (PCF, Mr = 879) is a novel marine active product isolated from gonochoric Chinese scallop Chlamys farreri which has been served as sea food for several thousand years. As an octapeptide, PCF consists of 8 amino acids, namely, Pro, Asn, Ser, Thr, Arg, Hyl, Cys, and Gly. PCF had been identified as a marine chemopreventive drug that protected hairless mice's epidermis against UV-induced damage in our previous study. However, the molecular mechanisms that underlie the effect of PCF on ultraviolet A-induced apoptosis in ketatinocytes are not well understood yet. In the present study, PCF was investigated as a potential inhibitory agent for UVA-induced apoptosis in a human keratinocyte cell line, HaCaT. The effects of PCF on UVA-induced generation of ROS and MDA, DNA damage, apoptosis rate were examined. We also investigated whether PCF could inhibit UVA-induced decreasing of mitochondrial membrane potential and the changing of morphology of the cells. We found that, compared with UVA only group, PCF attenuated UVA-induced generation of ROS and MDA, increased the mitochondrial membrane potential, and decreased the apoptosis rate. These results indicate that PCF may protect HaCaT keratinocytes against UVA induced apoptosis. PMID- 15292710 TI - Antiproliferative effects of ZD0473 (AMD473) in combination with 5-fluorouracil or SN38 in human colorectal cancer cell lines. AB - PURPOSE: ZD0473 (AMD473) [cis-amminedichloro(2-methylpyridine) platinum(II)] is a novel platinum agent of proven activity in vitro against a variety of human tumor derived cell lines even with intrinsic or acquired resistance to CDDP. The aim of this study is to provide the basis for a rational design of ZD0473-based combination in colon cancer. EXPERIMENTAL DESIGN: We evaluated the cytotoxic effect of ZD0473 administered alone or in combination with 5-Fluorouracil (5FU) or SN38 in a panel of sensitive and 5FU-resistant colorectal cell lines (HT29/HT29-5FUR and LoVo/LoVo-5FUR). We analyzed four sequential schedules of administration: ZD0473 --> 5FU, 5FU --> ZD0473, ZD0473 --> SN38 and SN38 --> ZD0473. MTT-assay and isobologram analyses were performed to determine the synergism/antagonism. RESULTS: The pattern of response towards ZD0473, administered as single agent, was similar in all cases and independent of the 5FU resistance phenotype (IC50 from 48.1 to 76.6 microM) and/or p53 status. No differences in sensitivity to ZD0473 alone or in combination were observed between DNA-mismatch repair-proficient (HT29/HT29-5FUR) and -deficient (LoVo/LoVo 5FUR) cells. ZD0473 administered prior to 5FU leads to synergistic/additive effect in all cell lines, while the 5FU --> ZD047 schedule was only synergistic in HT29 cells. Exposure to ZD0473 prior to SN38 leads to a synergistic/additive schedule in LoVo/LoVo-5FUR cells, while SN38 --> ZD0473 schedule was only synergistic in parental cell lines. CONCLUSIONS: The combinations of ZD0473 and 5FU or SN38 have shown to be active in sensitive and 5FU-resistant colorectal cell lines when a correct schedule of administration is applied. These results may be further exploited to promote new schedules of administration for advanced colorectal cancer treatment. PMID- 15292711 TI - Antitumor efficacy and acute toxicity of the novel dipeptide melphalanyl-p-L fluorophenylalanine ethyl ester (J1) in vivo. AB - The novel alkylating dipeptide melphalanyl-p-L-fluorophenylalanine ethyl ester (J1) was evaluated for acute toxicity and antitumor activity in mice, with melphalan as a reference. To determine a safe and tolerable dose for efficacy studies the acute toxicity following intravenous injection in the tail vein was monitored using a 14-day schedule with up to four doses. The highest tested dose, 25 micromoles/kg, was considered close to this level, with minor effects on body weight gain but significant effects on hematological parameters. Melphalan and J1 appeared equitoxic with no statistically significant differences. Subsequently a mouse hollow fiber model was employed with subcutaneous implantation of fibers containing human tumor cells. Three different human tumor cell lines as well as two samples of primary human tumor cells (ovarian carcinoma and chronic lymphatic leukemia) were used as tumor models. At the dose level tested there was a marked and statistically significant decrease in both T-cell leukemia CCRF-CEM and small cell lung cancer NCI-H69 tumor cell growth and viability in response to J1 as compared with both placebo and melphalan treated groups. In primary ovarian carcinoma cells only J1 treatment resulted in significant tumor regression (net cell kill). In summary the results indicate that, despite an expected short half time in the blood circulation, the promising in vitro data from the previous studies of J1 seems translatable into the in vivo situation. At equal doses of alkylating units J1, compared to melphalan, was more active in the mouse hollow fiber model, but showed similar general toxicity. PMID- 15292712 TI - Phase I trial of continuous infusion recombinant human interleukin-4 in patients with cancer. AB - BACKGROUND: A phase I study using recombinant human interleukin-4 (rhuIL-4) administered as a continuous intravenous infusion was conducted in patients with advanced cancer to study the toxicity profile and to determine the maximum tolerated dose (MTD) of this cytokine. METHODS: Twenty-six patients with non hematologic malignancies were treated with escalating doses of rhuIL-4 administered as 24-hour continuous intravenous infusion on days 1-5 and 15-19 every 28 days. The dose levels of rhuIL-4 were: dose level I-0.25 microg/kg/day (3 patients), dose level II-0.5 microg/kg/day (5 patients), dose level III-1.0 microg/kg/day (3 patients), dose level IV-2.0 microg/kg/day (10 patients) and dose level V-4.0 microg/kg/day (5 patients). RESULTS: Dose limiting toxicity of continuous infusion rhuIL-4 occurred at 4.0 microg/kg/day D1-5 and 15-19, in three of five patients and consisted of hematologic (thrombocytopenia and prolongation of PT) and neurologic (headache and neurocortical toxicity) toxicity. A mild flu-like syndrome characterized by fever, chills, fatigue, headache, anorexia, arthralgias and myalgias was seen almost universally, occurred more commonly and with increasing severity with higher dose levels and resolved completely on discontinuing therapy with rhuIL-4. None of the enrolled patients had an objective response to treatment with continuous infusion rhuIL-4. CONCLUSIONS: A five-day continuous infusion of rhuIL-4 given biweekly is well tolerated with a MTD of 2.0 microg/kg/day. PMID- 15292713 TI - Phase I/pharmacokinetic study of CCI-779 in patients with recurrent malignant glioma on enzyme-inducing antiepileptic drugs. AB - OBJECTIVES: CCI-779 is an ester of the immunosuppressive agent sirolimus (rapamycin) that causes cell-cycle arrest at G1 via inhibition of key signaling pathways resulting in inhibition of RNA translation. Antitumor activity has been demonstrated using cell lines and animal models of malignant glioma. Patients receiving enzyme-inducing anti-epileptic drugs (EIAEDs) can have altered metabolism of drugs like CCI-779 that are metabolized through the hepatic cytochrome P450 enzyme system. The objectives of this study were to determine the pharmacokinetic profile and the maximum tolerated dose of CCI-779 in patients with recurrent malignant gliioma taking EIAEDs. STUDY DESIGN: The starting dose of CCI-779 was 250 mg intravenously (IV) administered weekly on a continuous basis. Standard dose escalation was performed until the maximum tolerated dose was established. Toxicity was assessed using the National Cancer Institute common toxicity criteria. RESULTS: Two of 6 patients treated at the second dose level of 330 mg sustained a dose-limiting toxicity: grade III stomatitis, grade 3 hypercholesterolemia, or grade 4 hypertriglyceridemia. The maximum tolerated dose was reached at 250 mg IV. Pharmacokinetic profiles were similar to those previously described, but the area under the whole blood concentration-time curve of rapamycin was 1.6 fold lower for patients on EIAEDs. CONCLUSIONS: The recommended phase II dose of CCI 779 for patients on enzyme-inducing antiepileptic drugs is 250 mg IV weekly. A phase II study is ongoing to determine the efficacy of this agent. PMID- 15292714 TI - A phase I and pharmacokinetic study of the nonpolyglutamatable thymidylate synthase inhibitor ZD9331 plus docetaxel in patients with advanced solid malignancies. AB - PURPOSE: To assess the feasibility of administering ZD9331, a thymidylate synthase (TS) inhibitor that does not undergo polyglutamation and has broad antitumor activity, in combination with docetaxel in patients with advanced solid malignancies. The study also sought to determine the principal toxicities of the regimen and recommend appropriate doses for phase II studies, characterize the pharmacokinetics of the agents, evaluate the possibility of major drug-drug interactions, and seek preliminary evidence of anti-cancer activity. PATIENTS AND METHODS: Patients with advanced solid malignancies were treated with escalating doses of docetaxel as a 60-minute intravenous (IV) infusion followed 30 minutes later by ZD9331 as a 30-minute IV infusion every 3 weeks. At least three patients were treated at each dose level, and the maximum tolerated dose level was defined as the highest dose level that was not associated with an unacceptably high incidence of severe toxicity. The pharmacokinetics of both ZD9331 and docetaxel were also characterized. RESULTS: Nineteen patients were treated with 71 cycles of ZD9331 and docetaxel (ZD9331/docetaxel) at dose levels that encompassed dosing iterations of ZD9331 ranging from 65 to 260 mg/m(2) and docetaxel doses in the range of 50 to 75 mg/m(2). Neutropenia was the principal toxicity of the ZD9331/docetaxel regimen. Since five of six patients treated at the ZD9331/docetaxel dose-level of 260/60 mg/m(2) had grade 4 neutropenia that was brief and uncomplicated in the first course, a rigorous exploration of higher dose levels was not undertaken. Nonhematologic toxicities, consisting of malaise, diarrhea, rash, nausea, and vomiting, were also observed, but these effects were rarely severe. No major antitumor responses were observed. The pharmacokinetics of both ZD9331 and docetaxel were similar to those reported in previous studies of each agent administered alone, suggesting the lack of major drug-drug interactions. CONCLUSION: The combination regimen, consisting of ZD9331 and docetaxel, is feasible and well tolerated at single-agent doses that are clinically-relevant. This ZD9331/docetaxel regimen does not appear to be associated with either major pharmacokinetic or toxicologic drug-drug interactions. A ZD9331/docetaxel dose level of 260/60 mg/m(2) is recommended as an initial dose level in disease-directed studies of the regimen, with further dose escalation of docetaxel to 75 mg/m(2) if the initial treatment is well tolerated. Further studies with this regimen are warranted in tumor types that have demonstrated sensitivity to both agents. PMID- 15292715 TI - Phase I clinical trial of CEP-2563 dihydrochloride, a receptor tyrosine kinase inhibitor, in patients with refractory solid tumors. AB - CEP-2563 dihydrochloride (CEP-2563) is a soluble lysinyl-beta-alanyl ester of CEP 751, a potent inhibitor of the trk family of receptor tyrosine kinases and the platelet-derived growth factor (PDGF) receptor tyrosine kinase. CEP-2563 was developed because of the limited aqueous solubility of CEP-751. Preclinical models have demonstrated that both CEP-751 and CEP-2563 have antitumor activity in a variety of tumors. A Phase I clinical trial involving 18 patients was conducted to determine the toxicity profile, maximum tolerated dose (MTD), toxicity profile, and pharmacokinetics of CEP-2563 in patients with advanced solid tumors refractory to standard therapy. CEP-2563 was administered over 1 hour via a central venous catheter once daily for five consecutive days every three weeks. A rapid dose titration strategy with initial single patient cohorts and 100% dose escalations was used. With the appearance of drug-related toxicity, escalations were decreased to 50% or 25% and cohorts were expanded to 3 or 6 patients until establishment of the MTD. Dose escalation rapidly proceeded to 320 mg/m(2)/d. The dose limiting toxicities (DLTs) observed were grade 3 hypotension and grade 2 allergic reaction. Other toxicities included anemia, thrombocytopenia, anorexia, asthenia, diarrhea, fatigue, headache, nausea, vomiting, and rash. Pharmacokinetic analysis showed that CEP-2563 is reliably converted to CEP-751. This study demonstrated that single agent CEP-2563 therapy is feasible with acceptable toxicities. The recommended phase II dose is 256 mg/m(2)/d. Rapid dose escalation with single patient cohorts was a safe and efficient method of conducting this phase I trial. PMID- 15292716 TI - A phase I surrogate endpoint study of SU6668 in patients with solid tumors. AB - PURPOSE: To evaluate the biologic effects of SU6668 in patients with solid tumors using comprehensive measures of pharmacokinetics (PK), functional imaging, and tissue correlative studies. EXPERIMENTAL DESIGN: Eligible patients with tumors accessible for core needle biopsy were treated with SU6668 at doses of 200 or 400 mg/m(2)/day. Functional computed tomography (CT) scan and dynamic contrast enhanced magnetic resonance imaging (DCE-MRI) were performed at baseline and repeated 4 weeks and 12 weeks after treatment for analysis of tumor angiogenesis. The PK was analyzed using a high-performance liquid chromatography assay. Tumor specimens obtained via core needle biopsy at baseline and 4 weeks later were analyzed for the biologic effects of SU6668. RESULTS: Six of a total of seven patients received treatment for at least 3 months and underwent comprehensive correlative studies, including PK, imaging, and tissue biopsy. Functional CT showed that five of six patients had decreased blood flow in tumors in response to treatment, and DCE-MRI results indicated significant change of area under the signal intensity vs. time curve (AUC) and/or maximum slope (maximum rate of signal intensity change) in two of four patients evaluated with this technique. PK studies showed that the mean apparent oral clearance (Cl(oral)) measured on day 1 was 6.3 +/- 2.7 L/hr/m(2), yielding a mean AUC of 16.6 +/- 4.3 mg/L.hr. By day 22, the Cl(oral) was 40% more than that observed on day 1. CONCLUSION: It is feasible to evaluate the biologic effects of antiangiogenic agents using comprehensive surrogate measures. PMID- 15292717 TI - A phase II pilot study of high-dose 24-hour continuous infusion of 5-FU and leucovorin and low-dose PALA for patients with colorectal cancer: a Southwest Oncology Group study. AB - PURPOSE: The purpose of this phase II multi-institutional study was to define the efficacy and toxicity of infusional 5-FU in combination with PALA and leucovorin in patients with advanced colorectal cancer. PATIENTS AND METHODS: Patients were required to have histologically confirmed colorectal cancer with distant metastases. The treatment regimen consisted of 5-FU 2600 mg/m(2) as a 24-hours continuous infusion given once a week, concurrently with leucovorin (LV) at 500 mg/m(2) as a 24-hour continuous infusion. PALA was administered 24 hours prior to 5-FU/LV at a dose of 250 mg/m(2) iv over 15 minutes weekly. Patients were continued on the assigned treatment regimen until progression of disease, unacceptable toxicity, or the patient declined further therapy. RESULTS: This study accrued 28 patients and all were eligible and evaluable for toxicity. Four patients had inadequate assessment of response and are considered non-responders. There was one complete response and five partial responses for an overall response rate of 6/28 or 21% (95% confidence interval 8-41%). Estimated median survival was 17.4 months (95% confidence interval 13.3-20.5 months). One patient died of a treatment related infection. This patient also had grade 4 diarrhea and vomiting. CONCLUSION: The combination of 5-FU, leucovorin, and PALA in the doses and schedule used here, produces a response rate similar to other modulated schedules of 5-FU with similar survival and toxicity profiles. PMID- 15292718 TI - Gemcitabine and cisplatin chemotherapy is an active combination in the treatment of platinum-resistant ovarian and peritoneal carcinoma. AB - The treatment of recurrent ovarian cancer with the combination of gemcitabine and cisplatin chemotherapy has recently been shown to be an active regimen. But the majority of positive responses have been observed in patients considered either platinum-sensitive or who have had extended platinum-free intervals. The purpose of our study was to review our experience with this regimen in women with platinum-resistant ovarian and peritoneal carcinoma with more recent exposure to platinum. We studied twenty-two patients who had relapsed within six months of their most recent platinum-based regimen and were treated with gemcitabine (450 600 mg/m(2)) and cisplatin (30 mg/m(2)) on days 1 and 8 of a 21-day cycle. The overall response rate was 64% (95% C.I. 42-85%) with seven (32%) complete and seven (32%) partial responses. The median progression-free interval was 6.7 months for responding patients and 3.9 months for the entire study group. Median survival for responders was 15.8 months compared to 8.8 months for non responders. Overall survival was 11.4 months. Grade 3 or 4 toxicity was encountered in 59% of treatments. We conclude from this limited review that the combination of gemcitabine and cisplatin chemotherapy is an active regimen in platinum-resistant ovarian and peritoneal carcinoma and warrants consideration in the management of patients with recurrent disease. PMID- 15292719 TI - Docetaxel plus fractionated cisplatin is a safe and active schedule as first-line treatment of patients with advanced non-small cell lung cancer: results of a phase II study. AB - This phase II trial evaluated the efficacy and safety of docetaxel 85 mg/m(2) (day 1) and cisplatin 80 mg/m(2) (administered as 40 mg/m(2) doses each on days 1 and 2) every 3 weeks as first-line treatment of patients with advanced non-small cell lung cancer (NSCLC). Forty-two NSCLC patients were enrolled, most of them with stage IV disease (74%). A total of 195 chemotherapy cycles were administered (median 6, range 1-6). All patients were considered evaluable for efficacy and toxicity in an intention-to-treat (ITT) analysis. The overall response rate was 48% (95% CI, 33-64), including one CR (3%) and 19 PRs (45%). Stable disease was found in 6 patients (14%). The median time to disease progression was 4.9 months (95% CI, 4.0-5.7) and the median overall survival was 10.5 months (95% CI, 5.1 16.0). The survival rates at 1 and 2 years were 36.0% (95% CI, 19.9-52.0) and 18.0% (95% CI, 3.9-32.1), respectively. Overall, the combination showed an excellent safety profile. Severe hematological toxicities were uncommon: neutropenia (5% of patients, 1% of cycles) and febrile neutropenia (2% of patients, 0.5% of cycles). Asthenia (12%) was the only grade 3/4 non hematological toxicity that affected more than 10% of patients. There were no toxic deaths. In conclusion, docetaxel plus fractionated cisplatin as first-line treatment of advanced NSCLC patients showed similar efficacy as the same combination with higher doses of docetaxel but where cisplatin was administered in a single dose. This new schedule shows promise in its excellent hematological and non-hematological toxicity profile. A randomized phase III trial is needed to confirm these results. PMID- 15292720 TI - Decision tree model of the treatment-seeking behaviors among Korean cancer patients. AB - The purpose of this study was to develop and test a decision tree model of the treatment-seeking behaviors among Korean cancer patients. The study used methodological triangulation, applying the cognitive ethnographic decision tree modeling approach. The model was developed based on qualitative data collected from in-depth interviews with 29 cancer patients. The model was tested using qualitative and quantitative data collected from interviews and a structured questionnaire involving 165 cancer patients. The predictability of the decision tree model was quantified as the proportion of participants who followed the pathway predicted by the model. Two models were developed, the first for decision making about when to visit a doctor after detecting symptoms, and the second for decision making about treatment type following the diagnosis. Decision outcomes for the first model were categorized into immediate visit and delayed visit. The first model was influenced by the perceived seriousness of symptoms, the experiences of visiting a doctor previously with similar symptoms, social-group influences on visiting a doctor, and barriers to visiting a doctor. Decision outcomes for the second model were hospital treatment only, and a mixture of hospital treatment and alternative therapies. The second model was influenced by curability, social-group influences on alternative therapies, and confidence in alternative therapies. The predictabilities of the 2 models were 90.3% and 94.5%, respectively. This study result can help nurses understand the treatment-seeking behaviors of cancer patients, and hence develop nursing intervention strategies. PMID- 15292721 TI - Effect of acupressure on nausea and vomiting during chemotherapy cycle for Korean postoperative stomach cancer patients. AB - Despite the development of effective antiemetic drugs, nausea and vomiting remain the main side effects associated with cancer chemotherapy. The purpose of this study was to examine the effect of acupressure on emesis control in postoperative gastric cancer patients undergoing chemotherapy. Forty postoperative gastric cancer patients receiving the first cycle of chemotherapy with cisplatin and 5 Fluorouracil were divided into control and intervention groups (n = 20 each). Both groups received regular antiemesis medication; however, the intervention group received acupressure training and was instructed to perform the finger acupressure maneuver for 5 minutes on P6 (Nei-Guan) point located at 3-finger widths up from the first palmar crease, between palmaris longus and flexor carpi radialis tendons point, at least 3 times a day before chemotherapy and mealtimes or based on their needs. Both groups received equally frequent nursing visits and consultations, and reported nausea and vomiting using Rhode's Index of Nausea, Vomiting and Retching. We found significant differences between intervention and control groups in the severity of nausea and vomiting, the duration of nausea, and frequency of vomiting. This study suggests that acupressure on P6 point appears to be an effective adjunct maneuver in the course of emesis control. PMID- 15292722 TI - Dimensions of neutropenia in adult cancer patients: expanding conceptualizations beyond the numerical value of the absolute neutrophil count. AB - Neutropenia is a common and dangerous toxicity of cancer therapy that profoundly affects patients' lives. Neutropenia is typically defined by the numerical value of the absolute neutrophil count. However, considering neutropenia exclusively as the numerical value of the absolute neutrophil count limits its conceptualizations to physiologically related aspects, minimizes its complexities, and neglects dimensions of human response and the patient experience. This article offers a dimensional analysis of neutropenia derived from 42 research and clinical articles. Schatzman's dimensional analysis methods were applied to the literature to identify aspects of this phenomenon lying beyond its numerical boundaries. Dimensions of neutropenia that emerged were sorted into categories of perspective, context, conditions, processes, and consequences. The presence of the same dimension in more than 1 category and the circuitous relationships among categories begin to explicate the complexity and gravity of neutropenia. Articulation of these dimensions is necessary to assemble the beginnings of a theoretical understanding of neutropenia, which is crucial for the development and application of knowledge to research and practice. Limitations evident in the literature illuminate the urgent need for research into the psychosocial as well as physiologic dimensions of neutropenia. PMID- 15292724 TI - Reliability and validity of the breast cancer screening belief scale among Turkish women. AB - Breast cancer is the second leading cause of cancer deaths in Turkish women, and the use of breast self-examination (BSE) and mammography remains low in Turkey. Therefore, we need to identify the beliefs, influencing BSE and mammography, and a valid and reliable tool to measure constructs. The Champion's health belief model scale (CHBMS) is a valid and reliable tool to measure beliefs about breast cancer, BSE, and mammography in an English culture. The purpose of this study was to assess the psychometric characteristics of a Turkish version of the CHBMS related to breast cancer, BSE, and mammography. A convenience sample of 656 women was recruited from 3 health centers and 2 maternal and child health centers in Istanbul. The CHBMS was translated to Turkish, validated by professional judges, back translated, and tested. Factor analysis yielded 7 factors for BSE: confidence, seriousness, barriers-BSE, health motivation 1 and 2, susceptibility, and benefits-BSE. For mammography scale, 6 factors were identified: seriousness, benefits-mammography, barriers-mammography, health motivation 1 and 2, and susceptibility. All items on each factor were from the same construct. Cronbach alpha reliability coefficients ranged from.75 to.87 for the subscales. The Turkish version of the CHBMS showed adequate reliability and validity for use in Turkish women. It could easily be used to evaluate the health beliefs about breast cancer, BSE, and mammography. Further refinement is required to study Turkish women's health beliefs and breast cancer screening behaviors in various settings. PMID- 15292725 TI - African American women's breast memories, cancer beliefs, and screening behaviors. AB - African American women experience higher breast cancer mortality and lower survival rates compared with white women of comparable age and cancer stage. The literature is lacking in studies that address the influence of past events on current health behaviors among women of diverse cultural groups. This qualitative exploratory study used participant narratives to examine associations between women's memories and feelings concerning their breasts and current breast cancer screening behaviors. Twelve professional African American women, aged 42 to 64 years, shared stories about memories and feelings regarding their breasts. Codes grouped together with related patterns and recurrences revealed categories that encompassed the language and culture of the participants. The categories identified were Seasons of Breast Awareness, Womanhood, Self-Portraits, Breast Cancer and Cancer Beliefs, Breast Cancer Screening Experiences, and Participants' Advice for Change. These categories provide direction for further exploration of barriers to health promotion practices among African American women and women in general. PMID- 15292726 TI - Lived experience of survivors of leukemia or malignant lymphoma. AB - Individuals (n = 18) in remission from acute leukemia or highly malignant lymphoma were asked to narrate their lived experience of falling ill, of being under treatment, and life following this event. The transcribed texts were analyzed from a phenomenological-hermeneutic perspective, expanded by their medical and social history as related in interviews. The analysis revealed 3 themes: (I) Believed in life, fought for it and came through stronger; (II) Life went on, adapted and found a balance in the new life; (III) Life was over, felt out of control and lost belief in life. Participants in the first 2 groups viewed their quality of life as improved and stated that the struggle had been meaningful and that the experience had made them grow, as a person, related to the experience of gaining new insight or strength. The third group of survivors viewed their quality of life as worse. They found no meaning in their experience and evaluated the situation with bitterness. Thus the core of living through having acute leukemia or highly malignant lymphoma seemed to be to find meaning with it and the profound crisis it meant to them. To help people retell their experiences may be one way of processing this life-threatening disease and treatment and may be one way to developing a sense of meaning and to regain balance in life. PMID- 15292727 TI - End-of-life challenges: honoring autonomy. AB - Patients' end-of-life decisions challenge nurses to improve palliative care, symptom management, and patient advocacy, and examine ethical issues. When terminally ill patients take charge of the last stages of life, they may challenge nurses to reexamine attitudes about lifesaving technology and autonomy and values about preserving life. Staff members can become benevolent and believe that they know what is best despite the patient's independent decisions. When patients unsuccessfully decline continued aggressive, life prolonging strategies, they may decide to hasten dying rather than accept a natural death. Researchers (Breitbart WS et al. JAMA. 2000;284:2907-2911) defined desire for hastened death as a unifying construct underlying requests for assisted suicide, euthanasia, and withdrawal of food and fluids. When a terminally ill patient considers a hastened death, the nurse needs to examine the patient's mental health, symptom management, advance directives, and decision making. Medical and psychological symptoms and spiritual distress often trigger thoughts of hastening death even when pain and symptoms have been treated (Breitbart WS et al. JAMA. 2000;284:2907 2911). Ethical issues and guidelines for management of patients and evaluation of rationality are presented. PMID- 15292728 TI - Hope and coping in patients with cancer diagnoses. AB - This descriptive correlational study explored hope and coping in patients with various cancer diagnoses. Four groups of patients with gastrointestinal/ genitourinary, breast, head and neck, or hematologic malignancies completed the Herth Hope Scale, the Jalowiec Coping Scale, and a basic demographic form. Subjects were recruited from the oncology outpatient clinic of a large mid Atlantic teaching institution. Information about tumor site and stage was obtained from a chart review. Fifteen different malignancies were represented. Seventy-one percent of the 183 participants had metastatic or recurrent disease. No significant differences were found in the levels of hope or coping style use and coping effectiveness by type of cancer. The level of hope was relatively high, even in those patients who knew that their disease was in an advanced stage. A positive relationship was found between hope and coping style use (P =.013) and coping effectiveness (P <.001) in all 4 groups. The findings demonstrate that the level of hope was high and was positively related to coping in patients with cancer, regardless of gender, age, marital status, education, or site of malignancy. These findings support the need for nurses to continue to practice hope-inspiring behaviors, to implement hope-fostering interventions, and to avoid hope-hindering practices among their patients. PMID- 15292729 TI - Adolescents with cancer: experience of life and how it could be made easier. AB - This study is about what adolescents with cancer think about their life situation, the support they get, and the information they receive about their illness. The data for this qualitative and descriptive study were collected in 3 focus group interviews with 20 adolescents aged 13 to 18 years attending a cancer adjustment camp. Interpretation was based on the method of inductive content analysis. The adolescents' experiences of their current situation were analyzed into 5 categories: views on life here and now, negative experiences of self because of the illness, resources recognized in self, difficulties caused by the illness in relation to life around them, and resources identified in the world around. They made very little, if any, conscious effort to plan ahead for the future. The information received by the adolescents concerned their illness and its treatment here and now, various practical matters, as well as the future impacts of the illness and its treatments. Most of this information focused on the here and now, whereas the adolescents' information needs were mainly oriented to the future. As for the adolescents' chances to take part in making decisions about their care and life, the analysis yielded 6 categories: joint decision making, inadequate chances for decision making, independent decision making, illusion of decision making, reluctant to make decisions, and excluded from decision making. Finally, the adolescents' hopes for improvement were focused on staff activities, physical care facilities, chances to discuss and work through their experiences of the illness, and the support received from society. PMID- 15292731 TI - Heart rate variability of recently concussed athletes at rest and exercise. AB - PURPOSE: The objective of this study was to assess the neuroautonomic cardiovascular regulation in recently concussed athletes at rest and in response to low-moderate steady-state exercise, using heart rate variability (HRV). METHODS: A 5-min ECG sample was taken at rest from the 14 concussed athletes at 1.8 (+/- 0.2) days postinjury and again at 5 d later. Once asymptomatic at rest, the concussed athletes and their matched controls (N = 14) participated in an exercise protocol. The protocol consisted of a 2-min warm-up with a pedaling frequency between 50 and 60 rpm against a load of 40 W. After the warm-up, the athletes engaged in a low-moderate intensity steady state 10-min exercise bout where the pedaling frequency and load increased to 80-90 rpm and 1.5 W x kg(-1) body weight, respectively. The protocol was repeated 5 d later. A 5-min ECG sample from minutes 4 to 9 of the low-moderate intensity steady state exercise bout was used to assess HRV during exercise. Mixed model ANOVA were used to analyze the data. RESULTS: No difference at rest was detected between the concussed athletes and their matched controls in any of the HRV variables measured. However, across both exercise tests, the concussed group demonstrated a significant decrease in the mean RR interval, and low- and high-frequency power (P < 0.05) in relation to their matched controls. CONCLUSION: Low-moderate steady state exercise elicits a neuroautonomic cardiovascular dysfunction in concussed athletes that is not present in a rested state. This dysfunction alludes to an exercise induced uncoupling between the autonomic and cardiovascular systems. PMID- 15292732 TI - DAQIHF: methodology and validation of a daily activity questionnaire in heart failure. AB - PURPOSE: To assess the validity, reliability, and sensitivity of a new self administered physical activity questionnaire estimating daily energy expenditure (DEE) in patients with congestive heart failure (CHF). There is a need to develop a low cost, practical, and accurate tool increasing the knowledge of the type and dose of physical activity in patients with CHF for clinical and epidemiological aims. METHODS: One hundred five participants with stable CHF performed an incremental symptom-limited VO2(peak) test and completed the questionnaire. For DEE calculation, time spent in each activity was multiplied by its energy cost corrected for weight, age, sex, autonomy, and the total was calculated over 24 h. Reproducibility and sensitivity of the questionnaire as well as interrater reliability were tested. Concurrent validity was assessed against VO2(peak), anthropometric characteristics and data from the literature. RESULTS: Test-retest correlation coefficients used to measure reproducibility ranged from 0.82 for activities ranging from 3 to 5 METs to 0.98 for DEE and a paired Student's t-test didn't reach statistical significance for any activity score studied. Interrater reliability was high with an error in DEE estimation of 1.37% (t value = -1.064; P = NS). Sensitivity (changes in VO2(peak) concurrent to changes in DEE) was high (r = 0.88, P < 0.0001). DEE was in line with the literature in patients with CHF and relationships between DEE and VO2(peak) (r = 0.71, P < 0.0001), and DEE and anthropometric characteristics (<0.0001) were significant. Activity level above 3 METs was the best intensity criteria related to VO2(peak) (r = 0.62, P < 0.0001) and DEE (r = 0.80, P < 0.0001). CONCLUSION: The questionnaire seems reliable, sensitive and valid for the estimation of DEE. VO2(peak) appears related to global DEE and more particularly to activities above 3 METs in patients with CHF. PMID- 15292733 TI - Metabolic and cardiovascular parameters in type 1 diabetes at extreme altitude. AB - PURPOSE: The American Diabetes Association states that physical activity can be performed by individuals with Type 1 diabetes. Nevertheless, extreme altitude mountaineering represents a demanding challenge. We present the metabolic and cardiovascular parameters found in individuals with Type 1 diabetes during the ascent to Cho Oyu located at a height of 8201 m. METHODS: Six individuals with Type 1 diabetes and 10 matched controls participated in the expedition. Both groups were evaluated before and after 4 h of trekking for vital indices, blood gases, acute mountain sickness, and metabolic control at 0, 3700, and 5800 m. RESULTS: No difference between the groups was observed in acute mountain sickness scores. There was a progressive elevation in basal heart rates in both groups at increasing altitude while no changes were observed in mean blood pressures. After the 3 h of trekking, a significant increase in heart rate was observed in the controls at 0 m whereas a significant decrease in blood pressure was observed at higher altitude only in controls. HbA1c levels were worse after the expedition in both groups. A progressive increase in insulin requirement was observed in subjects with Type 1 diabetes (38 +/- 6 U x d(-1) at 0 m to 51 +/- 6 at 4200 m, P < 0.001). At an altitude of 5800 m, there was a significant increase in blood lactate concentration, independently of the activity level in the two groups. CONCLUSIONS: At extreme altitude, highly motivated trekkers with Type 1 diabetes but free from long-term complications present metabolic and cardiovascular parameters comparable with those of control subjects despite a worsening in metabolic control. This type of physical activity must be accompanied by careful glucose monitoring. PMID- 15292734 TI - Gender differences in viral infection after repeated exercise stress. AB - Fatiguing exercise can increase susceptibility to respiratory infection after intranasal inoculation with herpes simplex virus-1 (HSV-1) in male mice. Although gender differences in susceptibility to certain pathogens do exist, it is unknown whether female mice will respond differently than males in response to strenuous exercise and HSV-1 infection. PURPOSE: To test the effects of gender on susceptibility to HSV-1 respiratory infection after repeated exhaustive exercise. METHODS: Male (N = 86) and female (N = 89) CD-1 mice (approximately 60 d old) were randomly assigned to exercise (Ex) or control (C) groups. Exercise consisted of 3 d of treadmill running at 36 m x min(-1) at 8% grade until volitional fatigue (135 +/- 5min). Fifteen minutes after the last bout of exercise, Ex and C mice were inoculated intranasally with a standard dose (LD30) of HSV-1. Mice were monitored for 21 d for morbidity (time to sickness and symptom severity) and mortality. RESULTS: Run time to fatigue was significantly longer in females than males (P = 0.027). Significant gender differences in susceptibility to infection were found after exercise stress. In males, exercise stress resulted in increased morbidity (66%, P < 0.05) and mortality (30%, P < 0.05) whereas in females, exercise stress only resulted in increased morbidity (66%, P < 0.05). CONCLUSION: Results suggest that although males and females have similar morbidity rates after infection and exercise stress, females recover to a greater extent and are ultimately better protected from death. PMID- 15292735 TI - Trunk muscle strength and disability level of low back pain in collegiate wrestlers. AB - INTRODUCTION/PURPOSE: Low back pain (LBP) is a frequent injury in athletes. This study examined the relationship between isokinetic trunk muscle strength and the functional disability level of chronic LBP. We particularly focused on the existence of radiological abnormalities (RA) in the lumbar region. METHODS: Subjects were 53 collegiate wrestlers. Trunk extensor and flexor muscle strength was measured at three angular velocities (60, 90, and 120degrees x s(-1)). The examined parameters for trunk muscle strength were peak torque, work, average torque, and average power. The disability level of LBP was estimated by using two questionnaires. Based on the RA evaluation with x-ray and MRI, all 53 wrestlers were assigned to two groups as the RA group (N = 35, 66%) and the non-RA group (N = 18, 34%). Correlations between trunk muscle strength and the disability level of LBP in each group were analyzed with Spearman's rank test. RESULTS: Without considering the disability level, there were 14 subjects with LBP (40%) in the RA group and 8 (44%) in the non-RA. Significantly correlated parameters with the disability level of LBP could be observed only when the subjects were restricted to the non-RA group. The correlated parameters with the two questionnaires were peak torque at 120degrees x s(-1), work at 60degrees x s(-1) and 90degrees x s( 1), and average torque at 90degrees x s(-1) and 120degrees x s(-1). There were no significantly correlated parameters in the RA group. None of the trunk flexor parameters were significantly correlated with the disability level of LBP. CONCLUSION: The relatively low strength of trunk extensors may be one of the factors related to nonspecific chronic low back pain in collegiate wrestlers. PMID- 15292736 TI - Associations of muscle strength and fitness with metabolic syndrome in men. AB - PURPOSE: To examine the associations for muscular strength and cardiorespiratory fitness with the prevalence of metabolic syndrome among men. METHODS: Participants were 8570 men (20-75 yr) for whom an age-specific muscular strength score was computed by combining the body weight adjusted one-repetition maximum measures for the leg press and the bench press. Cardiorespiratory fitness was quantified by age-specific maximal treadmill exercise test time. RESULTS: Separate age and smoking adjusted logistic regression models revealed a graded inverse association for metabolic syndrome prevalence with muscular strength (beta = -0.37, P < 0.0001) and cardiorespiratory fitness (beta = -1.2, P < 0.0001). The association between strength and metabolic syndrome was attenuated (beta = -0.08, P < 0.01) when further adjusted for cardiorespiratory fitness. The association between cardiorespiratory fitness and metabolic syndrome was unchanged (beta = -1.2, P < 0.0001) after adjusting for strength. Muscular strength added to the protective effect of fitness among men with low (P trend = 0.0002) and moderate (P trend < 0.0001) fitness levels. Among normal weight (BMI < 25), overweight (BMI 25-30), and obese (BMI >or= 30) men, respectively, being strong and fit was associated with lower odds (73%, 69%, and 62% respectively, P < 0.0001) of having prevalent metabolic syndrome. CONCLUSIONS: Muscular strength and cardiorespiratory fitness have independent and joint inverse associations with metabolic syndrome prevalence. PMID- 15292737 TI - Effects of massage on limb and skin blood flow after quadriceps exercise. AB - PURPOSE: At present, there is little scientific evidence that postexercise manual massage has any effect on the factors associated with the recovery process. The purpose of this study was to compare the effects of massage against a resting control condition upon femoral artery blood flow (FABF), skin blood flow (SKBF), skin (SKT), and muscle (MT) temperature after dynamic quadriceps exercise. METHODS: Thirteen male volunteers participated in 3 x 2-min bouts of concentric quadriceps exercise followed by 2 x 6-min bouts of deep effleurage and petrissage massage or a control (rest) period of similar duration in a counterbalanced fashion. Measures of FABF, SKBF, SKT, MT, blood lactate concentration (BLa), heart rate (HR), and blood pressure (BP) were taken at baseline, immediately after exercise, as well as at the midpoint and end of the massage/rest periods. Data were analyzed by two-way ANOVA. RESULTS: Significant main effects were found for all variables over time due to effects of exercise. Massage to the quadriceps did not significantly elevate FABF (end-massage 760 +/- 256 vs end-control 733 +/ 161 mL x min(-1)), MT, BL, HR, and BP over control values (P < 0.05). SKBF (end massage 150 +/- 49 vs end control 6 +/- 4 au) SKT (end-massage 32.2 +/- 0.9 vs end-control 31.1 +/- 1.3degreesC) were elevated after the application of massage compared with the control trial (P < 0.05). CONCLUSION: From these data it is proposed that without an increase in arterial blood flow, any increase in SKBF is potentially diverting flow away from recovering muscle. Such a response would question the efficacy of massage as an aid to recovery in postexercise settings. PMID- 15292738 TI - The ACE gene and endurance performance during the South African Ironman Triathlons. AB - PURPOSE: Several studies have suggested that the insertion (I) variant rather than the deletion (D) variant of the human angiotensin-converting enzyme (ACE) gene is associated with elite endurance performance. The aim of this study was to determine whether the ID polymorphism is associated with the performance of the fastest finishers of the South African Ironman Triathlons. METHODS: A total of 447 Caucasian male triathletes of a variety of nationalities and athletic ability who completed either the 2000 or 2001 South African Ironman Triathlons and 199 Caucasian male control subjects were genotyped for the ACE ID polymorphism. RESULTS: There was a significantly higher frequency of the I allele in the fastest 100 South African-born finishers (103 I, 51.5% and 97 D, 48.5%) compared with the 166 South African-born control subjects (140 I, 42.2% and 192 D, 57.8%) (P = 0.036). There was also a significant linear trend for the allele distribution among the fastest 100 finishers (I allele = 51.5%), slowest 100 finishers (I allele = 47.5%), and control (I allele = 42.2%) South African-born subjects (P = 0.033). There was, however, no significant difference in the ACE genotype or allele frequencies when athletes born outside South Africa were analyzed. CONCLUSION: To our knowledge this is the first study that has examined the effect of an athlete's ACE genotype on their actual performance during an ultra-endurance race. The I allele of the ACE gene was associated with the endurance performance of the fastest 100 South African-born finishers in these triathlons. PMID- 15292739 TI - Effects of oat beta-glucan on innate immunity and infection after exercise stress. AB - PURPOSE: To test the effects of oat beta-glucan (ObetaG) on respiratory infection, macrophage antiviral resistance, and NK cytotoxicity. METHODS: Mice were randomly assigned to one of four groups: Ex-H2O, Ex-ObetaG, Con-H2O, or Con ObetaG. ObetaG was fed in the drinking water for 10 d before intranasal inoculation of HSV-1 or sacrifice. Exercise consisted of treadmill running to volitional fatigue (approximately 140 min) for three consecutive days. Fifteen minutes after the last bout of exercise or rest, mice (N = 24) were intranasally inoculated with a standardized dose of HSV-1. Mice were monitored twice daily for morbidity and mortality. Additional mice were sacrificed after exercise, peritoneal macrophages were obtained via i.p. lavage and assayed for antiviral resistance to HSV-1 (N = 18), and spleens were harvested and assayed for NK cell cytotoxicity (N = 12). RESULTS: Exercise stress was associated with a 28% increase in morbidity (P = 0.036) and 18% increase in mortality (P = 0.15). Ingestion of ObetaG before infection prevented this increase in morbidity (P = 0.048) and mortality (P = 0.05). Exercise stress was associated with a decrease in macrophage antiviral resistance (P = 0.007), which was blocked by ingestion of ObetaG (P < 0.001). There were no effects of exercise or ObetaG on NK cytotoxicity. CONCLUSION: These data suggest that daily ingestion of ObetaG may offset the increased risk of URTI associated with exercise stress, which may be mediated, at least in part, by an increase in macrophage antiviral resistance. PMID- 15292740 TI - Vitamin E and immunity after the Kona Triathlon World Championship. AB - PURPOSE: To measure the influence of vitamin E ingestion on oxidative stress and immune changes in response to the Triathlon World Championship in Kona, Hawaii. METHODS: Thirty-eight triathletes received vitamin E (VitE) (800 IU x d(-1) alpha tocopherol) or placebo (Pla) capsules in randomized, double-blind fashion for 2 months before the race event. Blood, urine, and saliva samples were collected the day before the race, 5-10 min postrace, and 1.5 h postrace. RESULTS: Race times did not differ between VitE (N = 19, 721 +/- 24 min) and Pla groups (N = 17, 719 +/- 27 min, P = 0.959), and both groups maintained an intensity of approximately 80% maximum heart rate during the bike and run portions. Plasma alpha-tocopherol was approximately 75% higher in the VitE versus Pla group prerace (24.1 +/- 1.1 and 13.8 +/- 1.1 micromol x L(-1), P < 0.001, respectively) and postrace. Plasma F2-isoprostanes increased 181% versus 97% postrace in the VitE versus Pla groups (P = 0.044). IL-6 was 89% higher (166 +/- 28 and 88 +/- 13 pg x mL(-1), respectively, P = 0.016), IL-1ra was 107% higher (4848 +/- 1203 and 2341 +/- 790 pg x mL(-1), respectively, P = 0.057), and IL-8 was 41% higher postrace in the VitE versus Pla groups (26.0 +/- 3.6 and 18.4 +/- 2.4 pg x mL(-1), respectively, P = 0.094). CONCLUSION: These data indicate that vitamin E (800 IU x d(-1) for 2 months) compared with placebo ingestion before a competitive triathlon race event promotes lipid peroxidation and inflammation during exercise. PMID- 15292741 TI - Effects of nandrolone decanoate on VO2max, running economy, and endurance in rats. AB - PURPOSE: The aim of the present study was to determine the effects of treatment with an anabolic androgenic steroid (AAS), nandrolone decanoate, on the submaximal running endurance (SRE), maximum oxygen consumption (VO2max), running economy (VO2submax), and blood oxygen carrying capacity of endurance trained rats. METHODS: Forty male Wistar rats were randomly allocated into two groups: a sedentary group and an exercising group training on treadmill for 8 wk. Half of the trained and half of the sedentary rats received weekly either nandrolone decanoate (10 mg x kg(-1)) or placebo (Pl) for the last 6 wk of experiment. SRE and VO2max tests were performed several times for all four groups (N = 10 each).Red blood cells parameters were measured at the end of the experiment. RESULTS: The trained rats had increased their SRE compared with sedentary rats throughout the experiment. At the end of the trial, the trained rats receiving nandrolone decanoate ran 46% longer than trained rats receiving Pl during the SRE test (P < 0.05). At the end of the experiment, trained rats had greater maximal time to exhaustion and higher VO2max than those of the sedentary rats but there were no differences in VO2max, VO2submax, and red blood cells parameters between the trained rats receiving nandrolone decanoate and those receiving Pl. CONCLUSIONS: Nandrolone decanoate has no effect on the SRE, VO2max and VO2submax of untrained rats. AAS treatment combined with submaximal training enhances SRE more than training alone but exerts no additive effects on VO2max, running economy, and oxygen carrying capacity of blood. The results suggest that this improvement in SRE of trained rats is due to the impact of AAS on other factors involved in exercise adaptation. PMID- 15292742 TI - T-wave and heart rate variability changes to assess training in world-class athletes. AB - PURPOSE: A decrease of electrocardiographic T-wave voltage with increasing training loads has been reported in elite endurance athletes and ascribed to training-related adaptation in sympathetic activity to the ventricles. A switch from vagal to sympathetic predominance in sino-atrial node regulation on going from low to peak training load has been reported in world-class rowers. In this study on world-class endurance athletes, we tested the hypothesis that training induced variations in T-wave amplitude at higher training loads are paralleled by changes in HR spectral profile. METHODS: We studied eight male rowers of the Italian national team in the season culminating with the Rowing World Championship. Athletes were evaluated at 50 and 100% of training load, approximately 20 d before the World Championship, and during the World Championship, when the intensity was markedly reduced. We assessed T-wave maximum amplitude in chest lead V6 and cardiac autonomic regulation by power spectral analysis of R-R interval variability. RESULTS: The increase in training load from 50 to 100% was accompanied by a significant decrease in high frequency and a significant increase in low-frequency R-R variability (in normalized units) with a concomitant significant decrease in T-wave amplitude (microV). Reduction in training load during the World Championship resulted in a return of spectral profile to the level observed at 50% training load and in a partial recovery of T wave amplitude. HR did not change significantly. CONCLUSIONS: In high-performance world-class athletes, training load simultaneously affects both ventricular repolarization and HR variability patterns possibly through variations in cardiac sympathetic modulation to the ventricles and the sino-atrial node. Information on concomitant changes in ventricular repolarization and autonomic cardiac regulation might be employed to tailor training protocols of elite athletes. PMID- 15292743 TI - Time course of neuromuscular alterations during a prolonged running exercise. AB - PURPOSE: This study investigated the time course of contractile and neural alterations of knee extensor (KE) muscles during a long-duration running exercise. METHODS: Nine well-trained triathletes and endurance runners sustained 55% of their maximal aerobic velocity (MAV) on a motorized treadmill for a period of 5 h. Maximal voluntary contraction (MVC), maximal voluntary activation level (%VA), and electrically evoked contractions (single and tetanic stimulations) of KE muscles were evaluated before, after each hour of exercise during short (10 min) interruptions, and at the end of the 5-h period. Oxygen uptake was also measured at regular intervals during the exercise. RESULTS: Reductions of MVC and %VA were significant after the 4th hour of exercise and reached -28% (P < 0.001) and -16% (P < 0.01) respectively at the end of the exercise. The reduction in MVC was highly correlated with the decline of %VA (r = 0.98, P < 0.001). M-wave was also altered after the fourth hour of exercise (P < 0.05) in both vastus lateralis and rectus femoris muscles. Peak twitch was potentiated at the end of the exercise (+18%, P = 0.01); 20- and 80-Hz maximal tetanic forces were not altered by the exercise. Oxygen uptake increased linearly during the running period (+18% at 5 h, P < 0.001). CONCLUSION: These findings suggest that KE maximal voluntary force generating capability is depressed in the final stages of a 5-h running exercise. Central activation failure and alterations in muscle action potential transmission were important mechanisms contributing to the impairment of the neuromuscular function during prolonged running. PMID- 15292744 TI - Effect of exercise intensity on relationship between VO2max and cardiac output. AB - PURPOSE: The purpose of this study was to determine whether the maximal oxygen uptake (VO2max) is attained with the same central and peripheral factors according to the exercise intensity. METHODS: Nine well-trained males performed an incremental exercise test on a cycle ergometer to determine the maximal power associated with VO2max (pVO2max) and maximal cardiac output (Qmax). Two days later, they performed two continuous cycling exercises at 100% (tlim100 = 5 min 12 s +/- 2 min 25 s) and at an intermediate work rate between the lactate threshold and pVO2max (tlimDelta50 +/- 12 min 6 s +/- 3 min 5 s). Heart rate and stroke volume (SV) were measured (by impedance) continuously during all tests. Cardiac output (Q) and arterial-venous O2 difference (a-vO2 diff) were calculated using standard equations. RESULTS: Repeated measures ANOVA indicated that: 1) maximal heart rate, VE, blood lactate, and VO2 (VO2max) were not different between the three exercises but Q was lower in tlimDelta50 than in the incremental test (24.4 +/- 3.6 L x min(-1) vs 28.4 +/- 4.1 L x min(-1); P < 0.05) due to a lower SV (143 +/- 27 mL x beat(-1) vs 179 +/- 34 mL x beat(-1); P < 0.05), and 2) maximal values of a-vO2 diff were not significantly different between all the exercise protocols but reduced later in tlimDelta50 compared with tlim100 (6 min 58 s +/- 4 min 29 s vs 3 min 6 s +/- 1 min 3 s, P = 0.05). This reduction in a-vO2 diff was correlated with the arterial oxygen desaturation (SaO2 = -15.3 +/- 3.9%) in tlimDelta50 (r = -0.74, P = 0.05). CONCLUSION: VO2max was not attained with the same central and peripheral factors in exhaustive exercises, and tlimDelta50 did not elicit the maximal Q. This might be taken into account if the training aim is to enhance the central factors of VO2max using exercise intensities eliciting VO2max but not necessarily Qmax. PMID- 15292745 TI - Single sessions of intermittent and continuous exercise and postprandial lipemia. AB - PURPOSE: This study compared the effects of continuous (CON-EX) and intermittent (INT-EX) exercise on postprandial lipemia (PPL). METHODS: Subjects were 18 inactive males (N = 7) and females (N = 11), aged 25 +/- 1.8 yr (mean +/- SE), VO2max 38.4 +/- 1.5 (mL x kg(-1)x min(-1)), and BMI 23.2 +/- 0.8 (kg x m(-2)). After 48-h activity and 24-h dietary control periods, subjects consumed a high fat meal (HFM) containing 1.5 g fat (88% of calories), 0.05 g protein, and 0.4 g carbohydrate per kilogram body weight for three trials: no exercise (NOEX), CON EX, and INT-EX. Both exercise trials consisted of 30 min of treadmill running at 60% VO2max. INT-EX was conducted in a single session of three bouts, each lasting 10 min and separated by a 20-min rest period. Blood was collected before the HFM (0 h) and at 2, 4, 6, and 8 h post-HFM. Exercise trials were completed 12 h before the HFM. Trials were separated by 7-10 d and were performed in random order. RESULTS: Plasma analysis indicated TG incremental area under the curve (AUCI) and TG incremental peak (PeakI) were significantly lower in INT-EX compared with NOEX, but CON-EX was not different from INT-EX or NOEX. Compared with females, males had significantly higher AUCI and PeakI in both exercise trials, but genders were not different in the NOEX trial. No difference was discovered among trials in high density lipoprotein (HDL)Total-C, HDL2-C, and HDL3-C, or fasting total cholesterol (TC) or fasting TC:HDL ratio. Females had higher fasting HDLTotal-C, HDL2-C, and HDL3-C compared with males. No gender or trial difference was found for fasting TC or TC:HDL ratio. CONCLUSIONS: Our data suggest that a single bout of INT-EX is more effective than CON-EX for lowering PPL as compared with NOEX in inactive, normolipidemic individuals. PMID- 15292746 TI - Energy balance, bone turnover, and skeletal health in physically active individuals. AB - Over the past 20 years, there have been a growing number of reports of low bone mineral density (BMD) or premature bone loss in individuals with a high physical activity level. These skeletal problems have been documented mainly in underweight women with amenorrhea and have often been linked to a sex hormone deficiency. However, sex hormone treatment has been shown to have limited efficacy for the prevention or treatment of low BMD in such women. Studies of bone turnover in women with sustained exercise-associated amenorrhea using metabolic markers of osteoblast activities and collagen synthesis have demonstrated an apparent reduction of bone formation that is associated with a low body mass index (BMI) and with endocrine disturbances that are characteristic of energy deficit. Comparable metabolic and endocrine disturbances have been observed in anorexic women that reverse with weight gain. Furthermore, increases of BMD accompany weight gain in both groups of women, even without a return of menses. Collectively, these observations suggest an important link between energy balance and the balance of bone turnover in women with exercise and/or diet associated amenorrhea. Although there have been few studies that have explored relations between bone turnover, BMD, and energy balance in physically active men, there is evidence for a link between reduced bone formation and an abrupt, short-term energy deficit. Interestingly, the presence of low BMD in physically active men has not been associated with a sex hormone deficiency. This review evaluates the evidence that underlies the hypothesis that an energy deficit is instrumental in the disturbance of bone turnover that has been observed in physically active individuals. PMID- 15292747 TI - Evaluation of a two-year middle-school physical education intervention: M-SPAN. AB - PURPOSE: School physical education (PE) is highly recommended as a means of promoting physical activity, and randomized studies of health-related PE interventions in middle schools have not been reported. We developed, implemented, and assessed an intervention to increase physical activity during middle-school PE classes. METHODS: Twenty-four middle schools (approximately 25,000 students, 45% nonwhite) in Southern California participated in a randomized trial. Schools were assigned to intervention (N = 12) or control (N = 12) conditions, and school was the unit of analysis. A major component of the intervention was a 2-yr PE program, which consisted of curricular materials, staff development, and on-site follow-up. Control schools continued usual programs. Student activity and lesson context were observed in 1849 PE lessons using a validated instrument during baseline and intervention years 1 and 2. RESULTS: The intervention significantly (P = 0.02) improved student moderate to vigorous physical activity (MVPA) in PE, by approximately 3 min per lesson. Effects were cumulative; by year 2 intervention schools increased MVPA by 18%. Effect sizes were greater for boys (d = 0.98; large) than girls (d = 0.68; medium). CONCLUSIONS: A standardized program increased MVPA in middle schools without requiring an increase in frequency or duration of PE lessons. Program components were well received by teachers and have the potential for generalization to other schools. Additional strategies may be needed for girls. PMID- 15292748 TI - An acute bout of static stretching: effects on force and jumping performance. AB - INTRODUCTION/PURPOSE: The objectives of this study were to examine whether a static stretching (SS) routine decreased isometric force, muscle activation, and jump power while improving range of motion (ROM). Second, the study attempted to compare the duration of the dependent variable changes with the duration of the change in ROM. METHODS: Twelve participants were tested pre- and post- (POST, 30, 60, 90, and 120 min) SS of the quadriceps and plantar flexors (PF) or a similar period of no stretch (control). Measurements during isometric contractions included maximal voluntary force (MVC), evoked contractile properties (peak twitch and tetanus), surface integrated electromyographic (iEMG) activity of the agonist and antagonistic muscle groups, and muscle inactivation as measured by the interpolated twitch technique (ITT). Vertical jump (VJ) measurements included unilateral concentric-only (no countermovement) jump height as well as drop jump height and contact time. ROM associated with seated hip flexion, prone hip extension, and plantar flexion-dorsiflexion was also recorded. RESULTS: After SS, there were significant overall 9.5% and 5.4% decrements in the torque or force of the quadriceps for MVC and ITT, respectively. Force remained significantly decreased for 120 min (10.4%), paralleling significant percentage increases (6%) in sit and reach ROM (120 min). After SS, there were no significant changes in jump performance or PF measures. CONCLUSION: The parallel duration of changes in ROM and quadriceps isometric force might suggest an association between stretch induced changes in muscle compliance and isometric force output. PMID- 15292749 TI - Effect of acute static stretching on force, balance, reaction time, and movement time. AB - PURPOSE: The purpose of the study was to investigate the effect of an acute bout of lower limb static stretching on balance, proprioception, reaction, and movement time. METHODS: Sixteen subjects were tested before and after both a static stretching of the quadriceps, hamstrings, and plantar flexors or a similar duration control condition. The stretching protocol involved a 5-min cycle warm up followed by three stretches to the point of discomfort of 45 s each with 15-s rest periods for each muscle group. Measurements included maximal voluntary isometric contraction (MVC) force of the leg extensors, static balance using a computerized wobble board, reaction and movement time of the dominant lower limb, and the ability to match 30% and 50% MVC forces with and without visual feedback. RESULTS: There were no significant differences in the decrease in MVC between the stretch and control conditions or in the ability to match submaximal forces. However, there was a significant (P < 0.009) decrease in balance scores with the stretch (decreasing 9.2%) compared with the control (increasing 17.3%) condition. Similarly, decreases in reaction (5.8%) and movement (5.7%) time with the control condition differed significantly (P < 0.01) from the stretch-induced increases of 4.0% and 1.9%, respectively. CONCLUSION: In conclusion, it appears that an acute bout of stretching impaired the warm-up effect achieved under control conditions with balance and reaction/movement time. PMID- 15292750 TI - Mechanisms of compensating for anterior cruciate ligament deficiency during gait. AB - INTRODUCTION: The quadriceps avoidance gait pattern may not be as common in ACL deficient (ACLd) gait as previously described. PURPOSE: The purpose of this study was to investigate the existence of the quadriceps avoidance pattern in ACL deficient patients and to further identify gait compensations that may exist in this subject pool. METHODS: In the present study, hip, knee, and ankle gait kinematics, and kinetics and thigh EMG profiles were recorded and compared for 16 ACLd and 8 control subjects. RESULTS: The quadriceps avoidance gait pattern was not observed for any of the subjects. Hip, knee, and ankle kinematics and kinetics were not different between groups. However, nine ACLd subjects (group A) demonstrated a normal biphasic knee moment pattern, whereas seven (group B) demonstrated an all knee extensor pattern. This indicates different adaptive mechanisms may be present in ACLd gait. Group A exhibited a hip strategy that increased hip extensor output, decreased knee extensor output, and allowed normal knee kinematics. Group B demonstrated a knee strategy that increased the stiffness of the joint and utilized a flexed knee gait. CONCLUSION: The prevalence of multiple adaptive strategies to compensate for ACL deficiency has several important ramifications. First, an ACLd subject pool with mixed compensating strategies may deter the identification of specific coping mechanisms and account for the confounding results in the literature. Second, the importance of the hip extensors should not be overlooked when studying this population. PMID- 15292751 TI - Transferability of strength gains from limited to full range of motion. AB - PURPOSE: This study was aimed at exploring the transferability of short range of motion (RoM) isokinetic conditioning on quadriceps performance inside and outside the trained range. METHODS: Fifty-five women were randomly assigned to one of four groups: G1 (N = 14) and G2 (N = 14) trained concentrically at 30 and 90degrees x s(-1), respectively, whereas G3 (N = 13) and G4 (N = 14) trained similarly but using the eccentric mode. All four groups trained within 30 60degrees of knee flexion. The training paradigm consisted of 4 sets of 10 maximal repetitions, 3x wk(-1) for a total period of 6 wk. Before the training period and 2 d after its termination, the isokinetic work output (Wisk) was assessed within three angular RoM: 85-60degrees (R1), 60-30degrees (R2), and 30 5degrees (R3). Isometric peak extension moment (PM) and rate of force development (RFD) were evaluated at 10degrees, 45degrees and 80degrees. RESULTS: Significant increases were observed in the isometric output (at all three angles), Wisk (in R1 and R2), and the RFD (45degrees). The PM increased significantly more in G3 and G4 compared with G1 and G2, evidencing specificity of contraction mode. CONCLUSION: These findings point out to the potential benefits of short RoM conditioning, particularly in those cases where, during specific phases of rehabilitation, a wider RoM may be contraindicative. PMID- 15292752 TI - Validation of a new method for estimating VO2max based on VO2 reserve. AB - PURPOSE: The American College of Sports Medicine's (ACSM) preferred method for estimating maximal oxygen consumption (VO2max) has been shown to overestimate VO2max, possibly due to the short length of the cycle ergometry stages. This study validates a new method that uses a final 6-min stage and that estimates VO2max from the relationship between heart rate reserve (HRR) and VO2 reserve. METHODS: A cycle ergometry protocol was designed to elicit 65-75% HRR in the fifth and sixth minutes of the final stage. Maximal workload was estimated by dividing the workload of the final stage by %HRR. VO2max was then estimated using the ACSM metabolic equation for cycling. After the 6-min stage was completed, an incremental test to maximal effort was used to measure actual VO2max. Forty-nine subjects completed a pilot study using one protocol to reach the 6-min stage, and 50 additional subjects completed a modified protocol. RESULTS: The pilot study obtained a valid estimate of VO2max (r = 0.91, SEE = 3.4 mL x min(-1) x kg-1) with no over- or underestimation (mean estimated VO2max = 35.3 mL x min(-1) x kg( 1), mean measured VO2max = 36.1 mL x min(-1) x kg(-1)), but the average %HRR achieved in the 6-min stage was 78%, with several subjects attaining heart rates considered too high for submaximal fitness testing. The second study also obtained a valid estimate of VO2max (r = 0.89, SEE = 4.0 mL x min(-1) x kg(-1)) with no over- or underestimation (mean estimated VO2max = 36.7 mL x min(-1) x kg( 1), mean measured VO2max = 36.9 mL x min(-1) x kg(-1), and the average %HRR achieved in the 6-min stage was 64%. CONCLUSIONS: A new method for estimating VO2max from submaximal cycling based on VO2 reserve has been found to be valid and more accurate than previous methods. PMID- 15292753 TI - Validity of VO2max equations for aerobically trained males and females. AB - PURPOSE: The purpose of this investigation was to cross-validate existing VO2max prediction equations on samples of aerobically trained males and females. METHODS: A total of 142 aerobically trained males (mean +/- SD; 39.0 +/- 11.1 yr, N = 93) and females (39.7 +/- 10.1 yr, N = 49) performed a maximal incremental test to determine actual VO2max on a cycle ergometer. The predicted VO2max values from 18 equations (nine for each gender) were compared with actual VO2max by examining the constant error (CE), standard error of estimate (SEE), correlation coefficient (r), and total error (TE). RESULTS: The results of this investigation indicated that all of the equations resulted in significant (P < 0.006) CE values ranging from -216 to 1415 mL x min(-1) for the males and 132 to 1037 mL x min(-1) for the females. In addition the SEE, r, and TE values ranged from 266 to 609 mL x min(-1), 0.36 to 0.88, and 317 to 1535 mL x min(-1), respectively. Furthermore, the lowest TE values for the males and females represented 10% and 12% of the mean actual VO2max values, respectively. CONCLUSIONS: The results of the analysis indicated that the two equations using age, body weight, and the power output achieved at VO2 as predictor variables had the lowest SEE (7.7-9.8% of actual VO2max) and TE (10-12% of actual VO2max) values and are recommended for estimating VO2max in aerobically trained males and females. The magnitude of the TE values (>or= 20% of actual VO2max) associated with the remaining 16 equations, however, were too large to be of practical value for estimating VO2max. PMID- 15292754 TI - Accuracy of polar S410 heart rate monitor to estimate energy cost of exercise. AB - PURPOSE: The purpose of this study was to examine the accuracy of the Polar S410 for estimating gross energy expenditure (EE) during exercise when using both predicted and measured VO2max and HRmax versus indirect calorimetry (IC). METHODS: Ten males and 10 females initially had their VO2max and HRmax predicted by the S410, and then performed a maximal treadmill test to determine their actual values. The participants then performed three submaximal exercise tests at RPE of 3, 5, and 7 on a treadmill, cycle, and rowing ergometer for a total of nine submaximal bouts. For all submaximal testing, the participant had two S410 heart rate monitors simultaneously collecting data: one heart rate monitor (PHRM) utilized their predicted VO2max and HRmax, and one heart rate monitor (AHRM) used their actual values. Simultaneously, EE was measured by IC. RESULTS: In males, there were no differences in EE among the mean values for the AHRM, PHRM, and IC for any exercise mode (P > 0.05). In females, the PHRM significantly overestimated mean EE on the treadmill (by 2.4 kcal x min(-1)), cycle (by 2.9 kcal x min(-1)), and rower (by 1.9 kcal x min(-1)) (all P < 0.05). The AHRM for females significantly improved the estimation of mean EE for all exercise modes, but it still overestimated mean EE on the treadmill (by 0.6 kcal x min(-1)) and cycle (by 1.2 kcal x min(-1)) (P < 0.05). CONCLUSION: When the predicted values of VO2max and HRmax are used, the Polar S410 HRM provides a rough estimate of EE during running, rowing, and cycling. Using the actual values for VO2max and HRmax reduced the individual error scores for both genders, but in females the mean EE was still overestimated by 12%. PMID- 15292755 TI - Difference in mechanical and energy cost between highly, well, and nontrained runners. AB - INTRODUCTION: Recently it has been shown that endurance training decreases the variability in stride rate. This decrease would lead to a reduction in the mechanical and the energy cost of running. PURPOSE: This study therefore aimed to compare the mechanical and the energy cost of running according to the training status of the runner (highly, well, and nontrained endurance runners). METHODS: The kinetic, potential, and internal mechanical costs (Cke, Cpe, and Cint) were measured with a 3D motion analysis system (ANIMAN3D). The energy cost of running (C) was measured from pulmonary gas exchange using a breath-by-breath portable gas analyser (Cosmed K4b2, Rome, Italy). All the parameters were measured on track, for a speed of 4.84 +/- 0.36 m x s(-1). RESULTS: Highly trained runners did not exhibit significantly lower C compared with well or nontrained runners (4.46 +/- 0.38; 4.33 +/- 0.32; 4.46 +/- 0.46 J x kg(-1) x m(-1), respectively; P = 0.75). However, Cpe was significantly lower in highly and well-trained runners compared with nontrained runners (0.43 +/- 0.07; 0.45 +/- 0.05; 0.54 +/- 0.08 J x kg(-1) x m(-1), respectively; P < 0.05). In contrast, Cint was significantly higher in highly trained runners compared with well and nontrained runners (respectively, 0.80 +/- 0.12; 0.60 +/- 0.09; 0.59 +/- 0.10 J x kg(-1) x m(-1); P < 0.05). CONCLUSION: Although there is a significant difference in Cpe and in Cint between runners of various training status, there is no difference in C. Differences in Cpe and Cint may be associated with the same self-optimizing mechanism that contributes to a reduction in the impact loads during the initial portion of the support phase of the stride. PMID- 15292756 TI - BMI in the Old Order Amish. PMID- 15292758 TI - Positioning technology to serve global heart health: the Fifth International Heart Health Conference. PMID- 15292759 TI - Fish oils--adjuvant therapy in chronic heart failure? AB - Despite advances in medical management and device therapy, chronic heart failure (CHF) remains a condition of high mortality and poor quality of life. Patients with CHF endure frequent admissions to hospital, with exacerbations of breathlessness or recurrent acute myocardial infarction and have a high incidence of sudden death. A high intake of marine polyunsaturated fatty acids (PUFAs) is associated with lower cardiovascular mortality in the general population, and diabetics, and can reduce cardiovascular deaths post-infarction. Many of the effects of PUFAs could be of benefit in CHF patients. They can improve endothelial function, reduce vascular tone, reduce platelet aggregability, improve myocardial relaxation, stabilize myocardial cells by prolonging the refractory period, and lead to increased appetite and weight gain. They also have potentially important immune-modulating effects, reducing cytokine production and release and altering prostaglandin metabolism. In this review article we discuss the potential benefits of PUFA supplementation in CHF patients using data from clinical trials and in vitro experiments. PMID- 15292760 TI - Cardiovascular disease risk factor levels and their relations to CVD rates in China--results of Sino-MONICA project. AB - BACKGROUND: The Sino-MONICA project is a 7-year study monitoring trends and determinants of cardiovascular disease (CVD) in geographically defined populations in different parts of China. This report focuses on risk factor levels and their relations to CVD rates. DESIGN: Successive surveys on smoking habits, blood pressure (BP), serum total cholesterol (TC), weight and height were conducted in independent random samples of the same populations early (1987-1988) and late (1992-1993) in the study period, by the methodology and criteria of the WHO MONICA project. Associations between risk levels and CVD rates were also assessed by correlation analysis. RESULTS: In general, the mean level of BP in the populations studied was high by international standards. Serum TC and body mass index (BMI) were low compared with the world average. There were significant geographic variations in CVD risk levels, being higher in the north and lower in the south, which correlated with the north-south difference of CVD event rates. CONCLUSIONS: The Sino-MONICA study has established the feasibility of long-term monitoring of CVD events and risk factors with international standardized methods in Chinese communities. The results will have significance in curbing the CVD epidemic not only in China, but also internationally. PMID- 15292761 TI - Physical activity, leisure habits and obesity in first-grade children. AB - BACKGROUND: Overweight and obesity are already on the rise in early childhood years. The relationships between genetic factors, malnutrition and physical inactivity are the underlying mechanisms. In this study, we examine the association between body indices, motor abilities and active (sport) and passive (television/computer) leisure time activities in a cohort of first-grade pupils. METHODS: The study group consisted of 344 children (51.5% male, 48.5% female). They were 6.8+/-0.4 years old, height was 123.9+/-4.9 cm, weight 24.8+/-5.0 kg, body mass index (BMI) 16.1+/-2.3 kg/m. After determination of the anthropometric data, a fitness test was performed in order to determine motor abilities. Parents were questioned about their children's leisure time activities, using a standardized questionnaire. Differences between BMI groups were evaluated using multivariate ANCOVA, adjusted for gender and age. RESULTS: Based on German BMI references, overweight and obesity were found in 12% of the children. They had poorer results with respect to endurance (P<0.001), leg strength (P=0.002), co ordination and balance (P=0.045) and spent more leisure time in watching television and at the computer (each P<0.001). No differences were found between their active leisure habits such as club sports. DISCUSSION: Our examinations with first-grade children show no differences in active leisure habits between obese children and their counterparts, although the former had poorer results in motor abilities, but they spend more time on sedentary leisure habits like audiovisual media. A possible explanation is their fewer regular daily activities. PMID- 15292762 TI - Environmental factors associated with body mass index in a population of Southern France. AB - STUDY OBJECTIVE: Environmental-factor changes may largely be accountable for the dramatic increase of obesity prevalence in industrialized countries. This study investigated the relationships between body mass index (BMI) and various socio economic, clinical, behavioural and reproductive factors in a population from Southern France. METHODS: Using a cross-sectional study, a sample of 3127 current and former salaried workers (1658 men and 1469 women) completed a questionnaire on personal and medical histories, and had a clinical examination including height and weight measurements. Age-adjusted and multiple linear regression analyses were performed. RESULTS: The overall prevalence of obesity (BMI> or =30 kg/m) was 9.8% and was higher in men than in women (11.1 versus 8.3%). Multivariate analyses showed that in both sexes, low educational level, television watching, low physical activity and ex-smoking habits, were independently associated with a higher BMI. Furthermore, in women, we found independent and positive associations between BMI and the number of naps per week, short sleep duration, daily alcohol consumption, the number of pregnancies, early age at menarche or the non-use of oral contraceptives. CONCLUSIONS: Our results reveal the complexity that exists between BMI and environmental factors and stress the need to analyse and to handle these factors simultaneously. PMID- 15292763 TI - The association between physical activity and the development of acute coronary syndromes in diabetic subjects (the CARDIO2000 II study). AB - BACKGROUND: The prevalence of type-2 diabetes is increasing dramatically, primarily being driven by environmental factors, like dietary and exercise habits. In this study we investigated the association of physical activity and acute coronary events in diabetic patients, an issue that has not been adequately studied so far. DESIGN: Cross-sectional, case-control study. METHODS: We studied demographic, lifestyle, dietary and clinical information in 216 hospitalized diabetic patients (171 men, 45 women) with a first event of an acute coronary syndrome and 196 frequency matched (by age and sex) diabetic controls (154 men, 42 women) without any evidence of coronary heart disease. Physical activity was evaluated according to the kcal/min expended and the weekly frequency of exercise. Physically active were considered those who reported non-occupational physical activity >once/week (at least 30 min/time). RESULTS: Seventy-eight (36%) of 216 patients and 110 (56%) of 196 controls were classified as physically active (P<0.001). Multivariate conditional logistic regression analysis revealed that the odds ratio for developing an acute coronary event in diabetic subjects who reported moderate levels of physical activity was 0.22 [95% confidence interval (CI): 0.12-0.47], while in those who reported vigorous physical activity it was 0.33 (95% CI: 0.21-0.59), after adjusting for age, sex, and the conventional cardiovascular risk factors. CONCLUSIONS: Physical activity (moderate and vigorous) seems to be associated with a lower prevalence of acute coronary events in the investigated group of diabetic subjects. Light physical activity does not seem to have any significant association with the development of acute coronary events. PMID- 15292764 TI - Determinants of the effects of physical training and of the complications requiring resuscitation during exercise in patients with cardiovascular disease. AB - BACKGROUND: Benefits of cardiac rehabilitation with exercise therapy are well established, although individual reactions are heterogeneous. The identification of determinants of training effects is useful from a prognostic point of view, but data regarding this are scarce. Furthermore, limited data exist on the determinants of complications during exercise in cardiac patients. This study aimed to investigate the determinants (1) of training effects in cardiac rehabilitation and (2) of complications requiring resuscitation during exercise activities at the hospital and during continued exercise at a sports club for cardiac patients. DESIGN: Clinical association study. METHODS: Determinants of changes in peak oxygen uptake (VO2) after 3 months of cardiac rehabilitation were determined by multiple regression analysis (n=1909). Determinants of events requiring resuscitation (n=21) were assessed by logistic regression analysis. RESULTS: Improvements in peak VO2 and exercise duration averaged 26%. Eighteen per cent of the variance in absolute improvements of peak VO2 was explained, with age and training characteristics as the strongest determinants. Twenty-one per cent of the variation in relative improvements was explained, with baseline exercise performance and training characteristics being the strongest determinants. The intake of anti-arrhythmics (odds ratio=5.5; P<0.001) and the presence of ST-segment depression (> or =1 mm) at baseline exercise testing (odds ratio=1.6; P<0.001) were predictive for serious complications. The occurrence of events requiring resuscitation was higher at the sports club (1/16,533 versus 1/29,214 patient-hours). CONCLUSIONS: Age, baseline exercise performance and training characteristics were predictive for training effects in cardiac rehabilitation. Anti-arrhythmics and ST-segment depression at baseline exercise testing were predictive for complications. PMID- 15292765 TI - Differences in sustainability of exercise and health-related quality of life outcomes following home or hospital-based cardiac rehabilitation. AB - BACKGROUND: Home-based cardiac rehabilitation (CR) has been demonstrated to be as effective as institution-based CR in post-coronary artery bypass graft surgery (CABG) patients in terms of short-term physical and psychosocial outcomes. The sustainability of these effects is less well studied. The aim of this study was to examine the sustainability of observed changes in physical, quality of life (HRQL), and social support (SS) outcomes in patients 12 months after discharge from a randomized controlled trial (RCT) of 6 months of monitored home-based versus supervised hospital-based CR. DESIGN: Two-hundred and twenty-two (n=222) patients were followed-up 12 months after discharge from a RCT of 6 months of monitored 'Home' versus supervised 'Hospital' CR after CABG. METHODS: At discharge from the 6-month RCT, participants who consented to the 12-month follow up study, were given individualized guidelines for ongoing exercise, and were not contacted for 1 year. The primary outcome was peak oxygen uptake (VO2). Secondary outcomes were: HRQL, SS and habitual physical activity. RESULTS: One hundred and ninety-eight patients (89.2%), 102 'Hospital' and 96 'Home', returned for follow up 12-months after discharge from CR. Both groups had similar medical and socio demographic characteristics. Peak VO2 declined in 'Hospital' but was sustained in 'Home' patients 12 months after discharge from CR (P=0.002). Physical HRQL was higher in the 'Home' group at the 12-month follow-up (P<0.01). Mental HRQL showed general, minor deterioration over time in both groups (P=0.019). Twelve months after discharge from CR, physical and mental HRQL remained higher than at entry to CR in both groups. 'Home' patients had higher habitual physical activity scores compared to 'Hospital' patients. CONCLUSIONS: This follow-up study suggests that low-risk patients whose CR is initiated in the home environment may be more likely to sustain positive physical and psychosocial changes over time than patients whose program is initially institution-based. PMID- 15292766 TI - Symptom-limited exercise testing, ST depressions and long-term coronary heart disease mortality in apparently healthy middle-aged men. AB - BACKGROUND: Previous studies have shown that ST depressions > or =1.0 mm during or post-exercise increase long-term risk of dying from coronary heart disease (CHD), the need for coronary artery bypass grafting (CABG) or the development of acute myocardial infarction (AMI) in healthy men. In the present prospective cohort study we investigate whether less marked ST depressions may influence CHD mortality, incidence of AMI, the need for a CABG or having a non-fatal stroke. METHODS: During 1972-75, 2014 men aged 40-59 years, free from somatic diseases and not using any drugs, underwent an examination programme including case history, clinical examination, various blood tests and a symptom-limited exercise ECG-test. ECG was registered during exercise and at 30 s, 1, 2, 3 and 5 min post exercise. The possible prognostic impact of ST-changes of 0.50-0.99 mm and > or =1.00 mm compared with normal ST-segments were studied separately and combined. Horizontal, down-sloping and slowly up-sloping ST-segment patterns were combined. RESULTS: After adjustment for age, smoking, blood pressure, cholesterol, maximal heart rate, left ventricular hypertrophy and physical fitness ST depressions > or =0.50 mm--during and/or post-exercise--were associated with a 1.47-fold [95% confidence interval (CI) 1.10-1.95], and 1.54-fold (95% CI of 1.17-2.04) increased 26 years risk of CHD-mortality, respectively. The same ST-changes also increased 22 years risk of developing non-fatal AMI or needing CABG but not developing non-fatal stroke. CONCLUSIONS: Even an ST depression > or =0.50 mm during and/or after exercise increases the long-term risk of CHD-death, developing an AMI or needing CABG. No association was found between ST-changes and incidence of non-fatal strokes. PMID- 15292767 TI - Cardiac rehabilitation programmes: predictors of non-attendance and drop-out. AB - BACKGROUND: Despite evidence of its benefits, attendance at cardiac rehabilitation (CR) programmes is poor. Past studies to identify predictors of non-attendance have been limited by their small sample size, particularly for female patients. The present study was designed to identify socio-demographic and clinical predictors of non-attendance and drop-out separately for men and women automatically referred to CR programmes. METHOD AND SUBJECTS: Prospective study of CR programme attendance amongst 808 patients consecutively admitted over an 11 month period to one of two hospitals in Melbourne, Australia, after acute myocardial infarction (AMI), or to undergo coronary artery bypass graft surgery (CABGS) or percutaneous coronary intervention (PCI). RESULTS: Of the 652 eligible patients, 573 (88%) were successfully tracked at 4 months. Of these, 284 (49.6%) had attended a CR programme, while 272 (47.5%) had not. Using logistic regression, the significant predictors of programme non-attendance among men were having had a PCI, being a non-driver, and being aged 70 or more. The only factor predictive of non-attendance for women was being aged 70 or more. Amongst attenders, 67 (23.6%) patients discontinued the programme. Being a smoker, having diabetes and being unemployed at the time of hospital admission were predictive of programme drop-out by men. Being physically inactive at admission was predictive of programme drop-out by women. CONCLUSIONS: The present study demonstrated a relatively high rate of CR programme attendance. Special attention needs to be directed towards males who are older, PCI patients, smokers, unemployed or non-drivers, and females who are older or inactive. PMID- 15292768 TI - Alcohol consumption and cardiovascular disease: differential effects in France and Northern Ireland. The PRIME study. AB - BACKGROUND: The effects of wine and other alcoholic beverages on coronary heart disease (CHD) have seldom been studied in several countries using a common methodology. DESIGN: Five-year prospective study conducted among 9750 men (7352 in France and 2398 in Northern Ireland) free of CHD at entry. Outcomes were angina pectoris, myocardial infarction or CHD death. RESULTS: In all, 90% of subjects in France reported drinking at least once per week, versus 61% in Northern Ireland. In France, after adjusting for other CHD risk factors, subjects in the highest quartile of alcohol consumption had a significantly lower risk of developing angina pectoris relative to non-drinkers. For myocardial infarction and all CHD events, the risk also decreased from the first to the fourth quartile (P for trend=0.02). Conversely, in Northern Ireland, no significant relationship was found between alcohol consumption and the incidence of angina pectoris or all CHD events, although alcohol consumption appeared to decrease the risk for myocardial infarction. Similar findings were obtained when the 5% higher alcohol consumers were excluded from the analysis. Finally, splitting the alcohol consumption into wine, beer and spirits did not improve the relationships, the three types of beverage exerting comparable effects on CHD events. CONCLUSIONS: Alcohol consumption patterns exert differential effects on CHD risk in middle aged men from France and Northern Ireland. Further, the amount of alcohol consumption, rather than the type of alcoholic beverage, is related to both angina pectoris and myocardial infarction in France, whereas no relationship was found in Northern Ireland. PMID- 15292769 TI - Promoter polymorphism of the gene for CD14 receptor is not associated with sub clinical carotid atherosclerosis in a community population. AB - BACKGROUND: The monocyte receptor CD14 is an important mediator of the inflammatory response to bacterial endotoxin. Recently, a functional polymorphism in the promoter of the CD14 gene (CD14-260C>T) was found to be associated with coronary heart disease. We examined if this polymorphism was associated with sub clinical carotid atherosclerosis in a community population. DESIGN AND METHODS: A randomly selected community population (557 men and 553 women; aged 27-77 years) underwent conventional risk factor assessment and ultrasound evaluation of the common carotid intima-medial wall thickness (IMT) and carotid plaque formation. CD14-260C>T genotypes were examined by restriction fragment length polymorphism analysis. Chlamydia pneumoniae-specific immunoglobulin G (IgG) and immunoglobulin A (IgA) antibody titres were determined by micro-immunofluorescence. RESULTS: The carrier frequency of the T allele and TT genotype was 0.48 and 0.22 respectively. Genotype frequencies met Hardy-Weinberg expectation. There was no significant association of -260C>T genotypes with traditional risk factors. On multivariate analysis, there was no independent association of genotypes with common carotid IMT in men and women or with prevalence of carotid plaque in women. Contrary to expectation, men who were TT homozygotes relative to CC wild-type had a lower adjusted risk of carotid plaque formation (odds ratio 0.34, 95% confidence interval 0.17-0.69; P=0.003). There was no evidence that smoking or C. pneumoniae infection modified the association of genotypes with carotid IMT or plaque formation. CONCLUSION: The CD14-260C>T gene polymorphism was not associated with an increased risk of sub-clinical carotid atherosclerosis in a community population. PMID- 15292770 TI - Homocysteine and cardiovascular disease: a 17-year follow-up study in Busselton. AB - STUDY OBJECTIVE: Prospective assessment of serum homocysteine level in relation to risk of coronary heart disease (CHD) and stroke. DESIGN: Case-cohort study with 17 years follow up. METHODS: Homocysteine was measured from stored serum. Proportional hazards regression models were used to obtain adjusted hazard ratios. RESULTS: There was no significant overall relationship between homocysteine and cardiovascular disease after controlling for known confounders. For women, removal of creatinine from the multivariate model resulted in a significant relationship. CONCLUSIONS: These results provide little support for a significant independent relationship between level of homocysteine and risk of CHD or stroke in men and women with no evidence of pre-existing cardiovascular disease. PMID- 15292771 TI - Recommendations for resistance exercise in cardiac rehabilitation. Recommendations of the German Federation for Cardiovascular Prevention and Rehabilitation. AB - Aerobic endurance training has been an integral component of the international recommendations for cardiac rehabilitation for more than 30 years. Notwithstanding, only in recent years have recommendations for a dynamic resistance-training program been cautiously put forward. The perceived increased risk of cardiovascular complications related to blood pressure elevations are the primary concern with resistance training in cardiac patients; recent studies however have demonstrated that this need not be a contraindication in all cardiac patients. While blood pressure certainly may rise excessively during resistance training, the actual rise depends on a variety of controllable factors including magnitude of the isometric component, the load intensity, the amount of muscle mass involved as well as the number of repetitions and/or the load duration. Intra-arterial blood pressure measurements in cardiac patients have demonstrated that that during low-intensity resistance training [40-60% maximum voluntary contraction (MVC)] with 15-20 repetitions, only modest elevations in blood pressure are revealed, similar to those seen during moderate endurance training. When properly implemented by an experienced exercise therapist, in specific patient groups an individually tailored, medically supervised dynamic resistance training program carries no inherent higher risk for the patient than aerobic endurance training. As an adjunct to endurance training, in selected patients, resistance training can increase muscle strength and endurance, as well as positively influence cardiovascular risk factors, metabolism, cardiovascular function, psychosocial well-being and quality of life. According to present data, resistance training is however not recommended for all patient groups. The appropriate training method and correct performance are highly dependent on each patient's clinical status, cardiac stress tolerance and possible comorbidities. Most studies have used middle-aged men of average normal aerobic performance capacity and with good left-ventricular (LV) function. Data are lacking for high risk groups, women and older patients. With the current knowledge it is reasonable to include resistance training without any restraints as part of cardiac rehabilitation programs for coronary artery disease (CAD) patients with good cardiac performance capacity (i.e., revascularised and with good myocardial function). As patients with myocardial ischaemia and/or poor left ventricular function may develop wall motion disturbances and/or severe ventricular arrhythmias during resistance exercise, the following criteria are suggested for resistance training: moderate-to-good LV function, good cardiac performance capacity [>5-6 metabolic equivalents of oxygen consumption (METS)=1.4 watt/kg body weight], no symptoms of angina pectoris or ST segment depression under continued maintenance of the medical therapy. Based on available data, this article presents recommendations for risk stratification in cardiac rehabilitation programs with respect to the implementation of dynamic resistance training. Additional recommendations for specific patient groups and detailed directions showing how to structure and implement such therapy programs are presented as well. PMID- 15292774 TI - Criteria of candidacy for unilateral cochlear implantation in postlingually deafened adults I: theory and measures of effectiveness. AB - OBJECTIVES: The objectives of this study were to distinguish the equivalent effectiveness, health-economic, and actuarial approaches to specifying criteria of candidacy for medical interventions; to apply the equivalent-effectiveness approach to unilateral cochlear implantation for postlingually deafened adults; and to determine whether the criterion should take age at implantation and duration of profound deafness into account. DESIGN: The study was designed as a prospective cohort study in 13 hospitals with four groups of severely-profoundly hearing-impaired subjects distinguished by their preoperative ability to identify words in sentences when aided acoustically. The groups represent a progressive relaxation of criteria of candidacy: Group I (N=134) scored 0% correct without lipreading and did not improve their lipreading score significantly when aided; group II (N=93) scored 0% without lipreading but did improve their lipreading score significantly when aided; group III (N=53) scored 0% without lipreading when the to-be-implanted ear was aided but between 1% and approximately 50% when the other ear was aided. Group IV (N=31) scored between 1% and approximately 50% without lipreading when the to-be-implanted ear was aided. Measures of speech intelligibility, health utility, and otologically relevant quality of life were obtained before surgery and 9 mo after surgery from each subject. Measures of effectiveness were calculated as the difference between 9-mo and preoperative scores. RESULTS: Effectiveness differed only slightly between groups. Effectiveness was not strongly associated with age at the time of implantation. Greater effectiveness was associated with implantation in the ear with the shorter duration of profound deafness. Cochlear implantation was least effective when the preoperative audiological status of the better-hearing ear was good and the duration of profound deafness of the implanted ear was long. As a result, effectiveness was not significant for the subsets of groups III and IV, who were given implants in ears that had been profoundly deaf for more than 30 yr. CONCLUSIONS: The effectiveness of cochlear implantation differs little between groups of candidates who score zero with acoustic hearing aids before surgery and groups who score up to approximately 50% correct, thereby justifying a relaxation of the criterion of candidacy to embrace some members of the latter groups. The criterion should be based not only on preoperative speech intelligibility but also on duration of profound deafness in the to-be-implanted ear. PMID- 15292775 TI - Criteria of candidacy for unilateral cochlear implantation in postlingually deafened adults II: cost-effectiveness analysis. AB - OBJECTIVES: The objectives of this study were to estimate the cost-effectiveness of unilateral cochlear implantation for postlingually deafened adults; to study the impact on cost-effectiveness of relaxing criteria of candidacy to include patients who benefit from acoustic hearing aids; and to study the further impact of age at implantation and duration of profound deafness before implantation. DESIGN: This prospective cohort study was carried out in 13 hospitals with four groups of severely to profoundly hearing-impaired subjects distinguished by their preoperative ability to identify words in prerecorded sentences when aided acoustically. The groups represent a progressive relaxation of criteria of candidacy: Group I (N=134) scored 0% correct without lipreading and did not improve their lipreading score significantly when aided; group II (N=93) scored 0% without lipreading but did improve their lipreading score significantly when aided; group III (N=53) scored 0% without lipreading when the ear to be given an implant was aided but between 1% and approximately 50% when the other ear was aided; and group IV (N=31) scored between 1% and approximately 50% without lipreading when the ear to be given an implant was aided. Lifetime costs to the UK National Health Service of providing and maintaining a cochlear implant were estimated for each subject. The gain in health utility from cochlear implantation was estimated with the Mark III Health Utilities Index and was combined with life expectancy to estimate the number of quality-adjusted life-years (QALYs) that would be gained from cochlear implantation. Cost/QALY ratios were calculated by means of the Net Benefit technique and were compared with an upper limit of acceptability of 50,000 euros/QALY. RESULTS: Averaged over the whole cohort, the cost of gaining a QALY was 27,142 euros (95% confidence interval, 24,532 euros to 30,323 euros); 203 of 311 (67%) of the cohort displayed cost/QALY ratios more favorable than 50,000 euros/QALY. The average cost of gaining a QALY increased from group I (24,032 euros) to groups II (27,062 euros) and IV (27,092 euros) to group III (39,009 euros). Cost/QALY varied with age at implantation from 19,223 euros for subjects who were younger than 30 yr of age to 45,411 euros for subjects who were older than 70 yr of age. Cost/QALY was unacceptable because of minimal gain in health utility for the subset of groups I and II, who were given implants in ears that had been profoundly deaf for more then 40 yr and for the subset of groups III and IV, who were given implants in ears that had been profoundly deaf for more than 30 yr. CONCLUSIONS: Cochlear implantation was a cost-effective intervention for the majority of subjects, including the group given implants when older than 70 yr of age. Relaxation of criteria of candidacy for cochlear implantation reduces cost-effectiveness. Prioritization of the provision of cochlear implantation should take duration of profound deafness in the ear to be given an implant into account, as well as preoperative word recognition performance. PMID- 15292776 TI - Criteria of candidacy for unilateral cochlear implantation in postlingually deafened adults III: prospective evaluation of an actuarial approach to defining a criterion. AB - OBJECTIVE: Outcomes from unilateral cochlear implantation in postlingually deafened adults are variable and difficult to predict precisely from data gathered before surgery. The objective was to derive and validate a method for specifying criteria of candidacy for implantation that takes this variability into account. DESIGN: Accuracy of identifying words in prerecorded sentences without lipreading was measured in 480 users of unilateral multichannel cochlear implants. These patients had all scored zero before surgery on prerecorded open set tests of word recognition in sentences with acoustic hearing aids. Statistical models were derived that calculated the odds that a patient would score higher with an implant than a criterion score, given knowledge of the duration of profound deafness in the implanted ear. The accuracy of the models was evaluated prospectively with two new groups of patients who scored between 1% and approximately 50% correct before surgery in one or both ears with acoustic hearing aids. Group I (N=53) was implanted in an ear that scored zero. Group II (N=31) was implanted in an ear that scored above zero. Benefits from implantation, measured as changes in word recognition performance and in health utility, were compared with the odds calculated by the statistical models. RESULTS: The preferred model was based on data from 376 subjects. It made accurate predictions of the proportion of patients in group I, and, disregarding minor exceptions, accurate predictions of the proportion of patients in group II, who improved on their preoperative word recognition score. Benefit from implantation was low for patients implanted with odds less favorable than 4:1 (4 chances out of 5). CONCLUSIONS: Adoption of odds of 4:1 as the criterion of candidacy for unilateral cochlear implantation would be likely to maintain acceptable benefit and cost-effectiveness while being explicit and informative for patients, clinicians, and commissioners of health care. PMID- 15292777 TI - Recognition of speech presented at soft to loud levels by adult cochlear implant recipients of three cochlear implant systems. AB - OBJECTIVE: The purpose of this study was to conduct a large-scale investigation with adult recipients of the Clarion, Med-El, and Nucleus cochlear implant systems to (1) determine average scores and ranges of performance for word and sentence stimuli presented at three intensity levels (70, 60, and 50 dB SPL); (2) provide information on the variability of scores for each subject by obtaining test-retest measures for all test conditions; and (3) further evaluate the potential use of lower speech presentation levels (i.e., 60 and/or 50 dB SPL) in cochlear implant candidacy assessment. DESIGN: Seventy-eight adult cochlear implant recipients, 26 with each of the three cochlear implant systems, participated in the study. To ensure that the data collected reflect the range of performance of adult recipients using recent technology for the three implant systems (Clarion HiFocus I or II, Med-El Combi 40+, Nucleus 24M or 24R), a composite range and distribution of consonant-nucleus-consonant (CNC) monosyllabic word scores was determined. Subjects using each device were selected to closely represent this range and distribution of CNC performance. During test sessions, subjects were administered the Hearing in Noise Test (HINT) sentence test and the CNC word test at three presentation levels (70, 60, and 50 dB SPL). HINT sentences also were administered at 60 dB SPL with a signal-to-noise ratio (SNR) of +8 dB. Warble tones were used to determine sound-field threshold levels from 250 to 4000 Hz. Test-retest measures were obtained for each of the speech recognition tests as well as for warble-tone sound-field thresholds. RESULTS: Cochlear implant recipients using the Clarion, Med-El, or Nucleus devices performed on average equally as well at 60 compared with 70 dB SPL when listening for words and sentences. Additionally, subjects had substantial open-set speech perception performance at the softer level of 50 dB SPL for the same stimuli; however, subjects' ability to understand speech was poorer when listening in noise to signals of greater intensity (60 dB SPL + 8 SNR) than when listening to signals presented at a soft presentation level (50 dB SPL) in quiet. A significant correlation was found between sound-field thresholds and speech recognition scores for presentation levels below 70 dB SPL. The results demonstrated a high test-retest reliability with cochlear implant users for these presentation levels and stimuli. Average sound-field thresholds were between 24 and 29 dB HL for frequencies of 250 to 4000 Hz, and results across sessions were essentially the same. CONCLUSIONS: Speech perception measures used with cochlear implant candidates and recipients should reflect the listening challenges that individuals encounter in natural communication situations. These data provide the basis for recommending new candidacy criteria based on speech recognition tests presented at 60 and/or 50 dB SPL, intensity levels that reflect real-life listening, rather than 70 dB SPL. PMID- 15292778 TI - Hearing loss associated with ear infections in Nord-Trondelag, Norway. AB - OBJECTIVE: We have assessed the effect of recurrent childhood ear infections on adult hearing. We also examined whether adult hearing is poorer with an early age of onset of ear infections compared with later onset. DESIGN: A population-based cohort of 50,398 subjects, 20 yr of age or older, were examined with air conduction, pure-tone audiometry and reported if they had had recurrent ear infections (EI) in childhood (or subsequently) with the age of onset. RESULTS: There were poorer hearing thresholds associated with recurrent EI for all frequency ranges from 0.25 kHz to 8 kHz, regardless of age or gender. The effect increased with age from approximately 2 dB among younger subjects (20 to 44 yr old) to approximately 5 to 6 dB among older subjects (65 yr or older). Among younger subjects, the effect of EI was somewhat stronger for men, whereas among older subjects the effect was somewhat stronger for women. Early age of onset for EI was associated with poorer hearing thresholds than late onset. The mean loss was close to 9 to 10 dB for all frequency ranges among older subjects reporting onset of EI before 2 yr of age. The mean loss was only about 4 dB among older subjects reporting onset of EI after 7 yr of age. CONCLUSIONS: Reported EIs are associated with similar consequences in terms of reduced hearing across frequencies 0.25 to 8 kHz for male and female subjects. The observed effects are stronger among older than among younger subjects, perhaps because effects increase with age or possibly because EI affected the hearing more before 1940 than during subsequent decades. Reported early age of onset of EI increases the risk of a substantially reduced hearing level later in life. PMID- 15292779 TI - Mean and median hearing thresholds among children 6 to 19 years of age: the Third National Health And Nutrition Examination Survey, 1988 to 1994, United States. AB - OBJECTIVE: The objective of this study was to provide the first national representative values for mean and median hearing thresholds among US children 6 to 19 yrs of age. METHODS: Hearing thresholds were obtained from 6166 children in the Third National Health and Nutrition Examination Survey (1988 to 1994), a national, population-based cross-sectional survey with household interview and audiometric testing at 0.5 to 8 kHz. Means, medians, and standard errors of the mean were obtained and reported by ear, frequency, sex, and age. RESULTS: The mean and median thresholds ranged from 3.0 to 11.8 dB HL and -1.0 to 10.8 dB HL, respectively. The highest (poorest) thresholds were obtained at test frequencies above 4000 Hz. Similar mean and median thresholds were found between boys and girls at all frequencies. CONCLUSIONS: These data indicate that the mean thresholds fall below the standard screening guidelines recommended by the American Speech-Language-Hearing Association (< or =20 dB HL for the frequencies from 1000< or =20 dB HL for the frequencies from 2000, and 4000 Hz). The results of this study suggest the need to include the test frequency of 6000 Hz in screening protocols for children. PMID- 15292781 TI - Editorial comment. The aging epidemic. PMID- 15292782 TI - The limits of medicine. PMID- 15292783 TI - Biomechanical considerations of fracture treatment and bone quality maintenance in elderly patients and patients with osteoporosis. AB - Osteoporosis is a major public health problem that is characterized by low bone mass and structural deterioration of bone tissue, leading to bone fragility and an increased susceptibility to fractures of the hip, spine, and wrist. Poor bone quality in patients with osteoporosis presents the surgeon with difficult treatment decisions. Bone fracture repair has more pathways with combinations of bone formation mechanisms, which depend on the type of fracture fixation to be applied to achieve the desirable immobilization. There only may be one remodeling principle and in less than ideal conditions, mechanical and biophysical stimuli may provide effective augmentation of fracture healing in elderly patients. A different stimulus may limit its association to a specific healing mechanism. However, no matter which fixation method is used, an accurate reduction is a requisite for bone healing. Failure to realign the fracture site would result in delayed union, malunion, or nonunion. Therefore, a basic understanding of the biomechanics of osteoporotic bone and its treatment is necessary for clinicians to establish appropriate clinical treatment principles to minimize complications and enhance the patient's quality of life. We describe the biomechanical considerations of osteoporosis and fracture treatment from various aspects. First, bone structure and strength characterization are discussed using a hierarchical approach, followed by an innovative knowledge-based approach for fracture reduction planning and execution, which particularly is beneficial to osteoporotic fracture. Finally, a brief review of the results of several experimental animal models under different fracture types, gap morphologic features, rigidity of fixation devices, subsequent loading conditions, and biophysical stimulation is given to elucidate adverse mechanical conditions associated with different bone immobilization techniques that can compromise normal bone fracture healing significantly. PMID- 15292784 TI - Systemic pharmacologic postoperative pain management in the geriatric orthopaedic patient. AB - Although older adults have surgical procedures more frequently than any other group, they also experience the worst postoperative pain management. Among patients with orthopaedic disorders, this undertreatment of pain impacts postsurgical functional recovery and clinical outcomes. Recently adopted evidence based pain management guidelines have improved care, but there still is significant room for improvement. We review standards for pain assessment in cognitively intact and impaired older adults, provide detailed guidelines for the pharmacologic treatment of postoperative pain in the orthopaedic geriatric patient, and review the stepwise approach to effective side-effect management in this population. PMID- 15292785 TI - Orthogeriatric care in patients with fractures of the proximal femur. AB - How are we to cope with the complex problems that now surround elderly patients with fractures? What are these problems and what systems should be in place to address them? No matter how successful we may be in treatment of osteoporosis and prevention of falls, it is inevitable that a massive worldwide increase in incidence of fragility fractures will occur in the next 50 years. It is crucial that we upgrade our systems to cope with this, if severe disruption of health services is to be avoided. This review considers the topics of falls, osteoporosis, and impact modification. We review the major medical issues that should be addressed in delivering hospital care that is efficient and of high quality. Various models for organizing the necessary liaison between fracture staff and medical or geriatric staff are examined and the current literature regarding orthogeriatric care is reviewed. We propose that the model of continuous orthogeriatric care, as practiced in Belfast, may be the way forward in addressing the complex problems associated with elderly patients with fractures. A system of ongoing audit of treatment outcomes and the journey of care for elderly patients with fractures of the proximal femur is necessary as a monitoring tool in developing the optimal system for orthogeriatric liaison. PMID- 15292786 TI - The older orthopaedic patient: general considerations. AB - People older than 65 years are more likely to need elective and emergent orthopaedic surgery compared with younger persons. They also experience significant benefits. Although age-related changes increase the risk of perioperative complications, understanding those changes allows prevention or at least early recognition and treatment when problems arise. Because of comorbidities, older persons take more medications that need to be managed in the perioperative period. Care could be simplified if patients were to bring their medications to the preoperative evaluation. Central nervous system sensitivity to certain pain medications (meperidine and propoxyphene) means that these drugs are best avoided as good alternatives exist (morphine and oxycodone). Adverse reactions to drugs are an important cause of acute confusion (delirium) that often complicates orthopaedic care. Early mobilization after surgery, avoiding certain drugs, avoiding restraints (including Foley catheters), attending to hydration, promoting normal sleep, compensating for sensory disorders, and stimulating daytime activities can prevent delirium. Patients with dementia are more likely to have delirium develop and, like many older people, will present special challenges in communication and decision making. Including family members in discussions may be helpful in ensuring truly informed consent. PMID- 15292787 TI - Locking compression plates for osteoporotic nonunions of the diaphyseal humerus. AB - Poor bone quality increases the technical difficulty and complications of operative treatment of nonunions and delayed unions of the diaphyseal humerus in older patients. Plates with screws that lock to the plate (transforming each screw into a fixed blade) are intended to improve the fixation of poor quality bone. Twenty-four patients (20 women, four men) with an average age of 72 years (range, 52-86 years) were followed up for a minimum of 12 months after locking compression plate fixation of an osteopenic delayed union (nine patients) or nonunion (15 patients) of the diaphyseal humerus. Twelve patients had iliac crest cancellous bone grafts, two patients had local graft, and 13 patients had demineralized bone applied to the fracture site. All the fractures eventually healed; two healed after a second procedure for autogenous bone grafting in patients who initially received demineralized bone. Using a modification of the Constant and Murley shoulder score, the results were good or excellent in 22 patients, and fair in two patients. Locking compression plates provide stable fixation of poor quality bone in patients with delayed union or nonunion of the humerus. Successful union and restoration of function are achieved in most patients. We no longer consider osteoporosis a contraindication to operative fixation of an ununited fracture of the humeral diaphysis. PMID- 15292788 TI - Treatment of distal humerus fractures in the elderly. AB - Geriatric patients with osteopenic bone present unique challenges in the treatment of fractures of the distal humerus, and require different strategies from the traditional treatment philosophies. Fracture union, rather than motion, is the first priority, because motion can be restored reliably by subsequent contracture release, if necessary, as long as the fracture heals. Modifications in the surgical technique, combined with newer implants incorporating distal, transcondylar screws into the plate to improve distal fixation, may improve outcomes. The use of massive, tricortical autogenous bone grafts to replace very comminuted segments of the medial and lateral columns also is helpful. Finally, modification of olecranon osteotomy fixation will minimize healing and hardware problems at this site. For fractures that are judged intraoperatively not to be stable enough to commence early motion, the implementation of a short period of immobilization followed by early soft tissue release will avoid exposing the patient to the risk of nonunion, and result in a more predictable functional outcome. PMID- 15292790 TI - Clinical pathway for hip fractures in the elderly: the Hospital for Joint Diseases experience. AB - Hip fractures are common injuries in the elderly and are associated with considerable morbidity and mortality. Although technical advances in the treatment of the elderly have resulted in improved fracture fixation and surgical outcomes, clinical pathways have been developed to further improve patient outcome while shortening hospital length of stay after hip fracture. We describe the clinical pathway used since 1990 at the Hospital for Joint Diseases. The outcomes of 747 patients treated before 1990 were compared with outcomes of 318 patients treated at our hospital after initiation of the clinical pathway. Use of the clinical pathway was associated with significant decreases in the acute care hospital length of stay, in-hospital mortality, and 1-year mortality. PMID- 15292791 TI - The lateral trochanteric wall: a key element in the reconstruction of unstable pertrochanteric hip fractures. AB - Pertrochanteric hip fractures still are a major orthopaedic challenge, and those that are unstable have the poorest prognosis. Fracture collapse is one of the postoperative complications reported in association with these fractures. My objective was to evaluate possible causes for pertrochanteric hip fracture collapse. Twenty-four patients with documented postoperative fracture collapse were evaluated retrospectively. The findings showed unequivocally that in all patients, this complication followed fracture of the lateral wall and resulted in protracted period of disability until fracture healing. The importance of the integrity of the lateral wall for event-free fracture healing clearly is indicated, and fracture of the lateral wall should be avoided in any fixation procedure. The presence of the lateral wall on the preoperative radiograph should be a major factor in determining the internal fixation device used for fracture stabilization. In unstable pertrochanteric hip fractures, the traditional description of the posteromedial fracture part as the most important prognostic factor should be revised to include the structural description of the lateral wall. Special caution should be taken when drilling at the base of this often delicate structure. PMID- 15292789 TI - Functional outcomes and mortality vary among different types of hip fractures: a function of patient characteristics. AB - A review of prospectively collected data was done to compare functional outcomes and mortality among patients with different hip fracture types. Five hundred thirty-seven elderly patients who sustained a hip fracture were followed up prospectively. Orthopaedists blinded to treatment and outcome radiographically classified the fractures as either: (1) nondisplaced or impacted femoral neck; (2) displaced femoral neck; (3) stable intertrochanteric; or (4) unstable intertrochanteric fracture. Functional independence measure scores were calculated for preinjury function and at 2- and 6- month follow-ups. Comorbidities, operative details, postoperative complications, and deaths were recorded. Six-month mortality was lowest for patients with nondisplaced femoral neck fractures (5.7%) and highest for patients with displaced femoral neck fractures (15.8%), but multivariate analysis only identified preinjury function as an independent predictor of mortality. All preinjury and followup functional independence measure scores were greatest for patients with nondisplaced femoral neck fractures and least for patients with unstable intertrochanteric fractures. However, multivariate analysis identified only patient age and preinjury functional independence measure scores as independent predictors of functional outcome. These data show differences in mortality and functional outcomes among fracture types that can be attributed to differences in functional status before injury. PMID- 15292792 TI - HA-coated screws decrease the incidence of fixation failure in osteoporotic trochanteric fractures. AB - This study was done to determine if elderly patients with trochanteric fractures and with osteoporosis could benefit from treatment with a dynamic hip screw fixed with HA-coated AO/ASIF screws. One hundred twenty patients with AO, A1, or A2 trochanteric fractures were selected. Patients were divided into two groups and randomized to receive a 135 degree-four-hole dynamic hip screw fixed with either standard lag and cortical AO/ASIF screws (Group A), or HA-coated lag and cortical AO/ASIF screws (Group B). Lag screw cutout occurred in four patients in Group A, but not in any patients in Group B. In Group A, the femoral neck shaft angle was 134 degrees +/- 5 degrees postoperatively and 127 degrees +/- 12 degrees at 6 months. In Group B, the femoral neck shaft angle was 134 degrees +/- 7 degrees postoperatively and 133 degrees +/- 7 degrees at 6 months. The Harris hip score at 6 months was 60 +/- 25 in Group A and 71 +/- 18 in Group B. The superior results of Group B can be attributed to the increased screw fixation provided by the HA-coated screws. We recommend lag screws coated with HA for dynamic hip screw fixation, especially in osteoporotic bone. PMID- 15292793 TI - Use of the talon hip compression screw in intertrochanteric fractures of the hip. AB - A retrospective analysis of a compression hip screw with four reversibly deployable talons was done. Fifty-four patients had sufficient radiographs to be included in this analysis. One-year mortality was 17% and increased to 41% by 2 years. No lag screws cut out, and postoperative slide was reduced compared with that in many published series. Three patients had revision of a failed alternate type hip pin with the Talon hip compression screw. Previous studies showed the talons provide the definitive difference in allowing enhanced compression at the time of surgery, preventing cut-out by enhanced rotational stability, and allowing immediate postoperative weightbearing without excessive limb shortening. The failure mode of the Talon compression hip screw seems to be side-plate loosening rather than varus deformity and lag screw cut-out. The Talon compression hip screw especially is effective with weak, osteoporotic bone and in unstable, three-part and four-part fractures. A previous study showed that Talon deployment notably improved interfragment compression and torsional strength, and that engagement or penetration into or through the cortical bone at the base of the femoral head-neck junction in the inferior lag screw position was the critical technical step to maximize the talon lag screw purchase. PMID- 15292794 TI - Primary total knee arthroplasty for complex distal femur fractures in elderly patients. AB - Fractures of the distal femur in the geriatric population are associated with a high incidence of postoperative complications and poor results. Nonunion, loss of fixation, and malunion of these fractures occur with all types of treatment. The postoperative treatment of these patients demands a lengthy period of limited weightbearing that can increase the rate of medical complications. Our experience with these challenging fractures caused us to consider the use of a primary distal femur replacement total knee arthroplasty with the goals of elimination of fracture healing issues, early mobilization, and immediate weightbearing. Twenty four distal femoral replacement knee arthroplasties were done from July 1998 to January 1999. Reviewed with a mean followup of 11 months, 17 patients (71%) resumed their preoperative level of ambulation. Knee range of motion averaged 1 degree - 103 degrees. No major surgical or significant medical complications were experienced by these patients. Our experiences with this small number of patients have shown that an immediate arthroplasty offers many advantages over open reduction and internal fixation for geriatric patients with poor bone quality, preexisting degenerative joint disease, and medical problems. PMID- 15292795 TI - Long-term results of total joint arthroplasty in elderly patients who are frail. AB - Many studies of joint replacement in the aged population include a wide spectrum of geriatric patients ranging from relative healthy and otherwise vigorous younger-elderly to small numbers of much older patients with many comorbid problems. To clarify the latter patient subset we assessed the results of total hip and knee arthroplasties done on a group of frail elderly patients. We retrospectively reviewed the preoperative and postoperative charts and radiographs of 130 patients who were at least 80 years when they had a total joint arthroplasty. One hundred arthroplasties (70 hips, 30 knees) were done. On a subjective grading scale, 95% of the patients were very satisfied, 5% reported satisfactory outcomes and no patients considered their results poor. After surgery, 90% of the patients became community walkers without assistance. The level of independent living was maintained in 97% of patients at long-term followup. Causes of morbidity included two infections, one dislocation, and one leg-length discrepancy. This study specifically addresses the outcomes of total joint replacement in elderly patients who are frail. We show that satisfactory and cost-effective health outcomes can be anticipated after total joint arthroplasties in this age group. PMID- 15292796 TI - Spinal disorders in the elderly. AB - As life expectancies increase, the geriatric population will increase, and the treatment of spinal diseases in the elderly will become even more commonplace. Treatment of spinal disorders in the geriatric patient population is a difficult challenge and involves numerous surgical, medical, and social issues. This review will provide an overview of the various spinal disorders particular to the geriatric patient population and will highlight certain concepts critical in the treatment of the spine in the geriatric population. Multiple factors, including poor tolerance of immobilization, medical comorbidities, use of multiple chronic medications, poor nutritional status, inadequate bone stock, and poor bone quality limit rigorous adherence to one treatment algorithm. These issues should be taken into consideration when formulating an individualized treatment plan that emphasizes early mobilization and functional rehabilitation. Goals, expectations, and surgical indications should be realistic and often will differ from those for a younger, healthier population. The use of a multidisciplinary approach will increase the likelihood of a successful treatment outcome and decrease the likelihood of potential complications. PMID- 15292797 TI - Osteoporosis: a review. AB - Osteoporosis is the most common metabolic bone disorder and remains an increasingly significant problem, affecting 200 million individuals worldwide. Osteoporosis often is undertreated and underrecognized, in part because it is a clinically silent disease until it manifests in the form of fracture. Sufficient recognition of the disease and its appropriate medical and nonmedical treatment are essential. Treatments including calcium and vitamin D, the bisphosphonates, estrogen, selective estrogen receptor modulators, calcitonin, parathyroid hormone, balance and exercise training programs, and the minimally invasive spine procedures vertebroplasty and kyphoplasty comprise a comprehensive multidisciplinary approach in the treatment of osteoporosis. The data suggest that medical treatment of osteoporosis is increasing each year as physician awareness is heightened. Nonmedical treatment of osteoporosis complements the appropriate pharmacologic treatment, and these treatments should be used together to maximize outcomes for patients with osteoporosis. Fracture data for the intravenous biphosphonates and the long-term effects of the minimally-invasive spine procedures vertebroplasty and kyphoplasty have yet to be reported in the literature, but the effects on bone mineral density, and short-term results of these procedures, respectively, have been promising. PMID- 15292798 TI - High-dose alendronate uncouples osteoclast and osteoblast function: a study in a rat spine pseudarthrosis model. AB - The effect of alendronate on osteoclast and osteoblast function was studied in a novel spine pseudarthrosis model in rats. Sixty-three Sprague-Dawley rats were divided into three groups: control group (saline), therapeutic dose group (1 microg/kg/week), and one-log overdose group (10 microg/kg/week). Animals had L4 L5 posterior intertransverse process fusion with limited bone graft and were sacrificed at 2, 4, and 6 weeks. Manual palpation showed no notable differences among groups. Treatment group radiographic scores were equal to or better than control group scores and were higher than the overdose group at 2 and 6 weeks. Qualitatively, limited histologic remodeling and poor osteoclastic and osteoblastic function were noted in the alendronate treated groups. Quantitative histologic analysis showed fewer osteoclasts in the therapeutic and high-dose groups (p < 0.001). The percent osteoblasts per bone surface area was lower in the high-dose group (p < 0.05). The results suggest that the effect of alendronate was dose dependent and animal model dependent and that supranormal doses of alendronate had a deleterious effect on osteoclastic and osteoblastic function in this model. PMID- 15292799 TI - Osteoporosis: a disease in men. AB - Osteoporosis, a metabolic bone disease characterized by low bone mass, microarchitectural deterioration of bone tissue, and increased susceptibility to fracture, now is recognized to be a disease of men and women. Based on bone mineral density measurements established by the World Health Organization, approximately 2 million men in the United States currently are affected. Information about the pathogenesis, diagnosis, and treatment of osteoporosis in men is accumulating. Estrogen now is known to be necessary for normal skeletal maturation and for maintenance of adult bone mass in men. As we will discuss, the age-related decline in bone mineral density in men seems to result from an age related decline in the levels of bioavailable estrogen. Osteoporosis candidate genes have been discovered, and their polymorphisms may predispose men to a weakened, osteoporotic phenotype. Determining a bone mineral density standard to diagnose men with osteoporosis has proven challenging, often leaving diagnosis until a fragility fracture has occurred. General prevention strategies for osteoporosis in men are targeted at men most likely to sustain a fragility fracture. Specific medical therapies for osteoporosis in men, including bisphosphonates and intermittent parathyroid hormone, are proving promising. PMID- 15292800 TI - Dorsal wrist ligament insertions stabilize the scapholunate interval: cadaver study. AB - This study examined sequential arthroscopic sectioning of volar, interosseous, and dorsal ligaments about the scapholunate complex in cadaver wrists. We attempted to clarify the contributions of the dorsal ligamentous complex to scapholunate instability and carpal collapse. We found that after sequential sectioning of volar ligaments and the scapholunate interosseous ligament, no scapholunate diastasis or excessive scaphoid flexion occurred. After dividing the dorsal intercarpal ligament, scapholunate instability occurred without carpal collapse. With sectioning of the dorsal radiocarpal ligament from the lunate, a dorsal intercalated scapholunate instability deformity ensued. This information may be of value in comprehending the pathogenesis of scapholunate instability and carpal collapse and in devising the rationales for conservative measures and surgical intervention. PMID- 15292801 TI - Does pain predict outcome in hips with osteonecrosis? AB - It generally is accepted that without specific treatment 70-80% of hips with clinically diagnosed osteonecrosis will progress to collapse. However, there are conflicting reports regarding the relationship between pain and outcome before femoral head collapse. Some surgeons are reluctant to operate on patients with asymptomatic or minimally symptomatic hips, assuming that these patients have a better prognosis than patients with pain. This study reviewed the outcome of 328 hips in 235 patients with nontraumatic osteonecrosis, all treated with core decompression and grafting. The preoperative stage, the extent of necrosis, and the Harris pain scores were correlated with the clinical and radiographic outcomes. Mean followup was 46 months. Patients with hips treated surgically did better as a group than patients with hips treated without surgery. A direct correlation was found between outcome and the stage and size of the necrotic lesion. Hips that had femoral head collapse were more painful than hips that did not have collapse and had a poorer outcome. Before collapse, outcome was correlated with the size of the necrotic lesion but there was no correlation with the preoperative pain level. These findings, although limited to patients with hips which had core decompression and grafting, support the observations of investigators who reported that most asymptomatic hips with osteonecrosis would progress without specific treatment. They also may apply to hips which have other forms of prophylactic treatment. Although several factors must be considered in determining the optimum treatment of hips with early stages of osteonecrosis, prophylactic treatment should not be withheld specifically because of the absence or paucity of pain. PMID- 15292802 TI - Disassembly of bipolar cup with self-centering system: a report of seven cases. AB - Disassembly of bipolar cups with a self-centering system occurred in six patients (seven hips; five women, one man) and the cause of the failure was evaluated. The mean age of the patients at the time of arthroplasty was 49.7 years (range, 27-85 years), and mean weight was 48.4 kg (range, 37-65 kg). The mean time to failure was 7.5 years (range, 4.8-9.2 years). Before failure, all implants functioned well and none of the patients had sustained trauma. Retrieval study showed that the cause of failure of the locking mechanism with a self-centering system was severe polyethylene abrasion at the rim attributable to impingement and deformity of the locking ring. After the locking ring detached, the inner head dislocated from the outer head resulting in a varus position of the outer head in the acetabulum. If the deformed locking ring did not detach, the inner head could dislocate, with the varus outer head remaining in the acetabulum. The incidence of this failure was 11%. This disassembly is not a rare occurrence numerous years after a well-functioning bipolar hemiarthroplasty, even with the self-centering system. PMID- 15292803 TI - Factor V leiden and prothrombin gene mutation: risk factors for osteonecrosis of the femoral head in adults. AB - The purpose of the current study was to determine whether factor V Leiden and the prothrombin 20210A gene mutation are risk factors for osteonecrosis of the femoral head in different etiologic groups of osteonecrosis in adults and whether patients with idiopathic osteonecrosis of the femoral head have a higher frequency of thromboembolic events compared with the general population. We investigated 63 adult patients with nontraumatic osteonecrosis of the femoral head for etiologic factors, such as corticosteroid medication and alcohol abuse, and the occurrence of factor V Leiden and the prothrombin 20210A gene mutation. In 35 patients, the disease was considered idiopathic and 10 of these patients (29%) had factor V Leiden or the prothrombin 20210A gene mutation or both. Mutations in factor V or the prothrombin 20210A gene were significantly more frequent in patients with idiopathic osteonecrosis than in a population of healthy control subjects (odds ratio, 2.7; 95% confidence interval range, 1.2 5.8) and in patients with osteonecrosis caused by corticosteroid medication or alcohol abuse (odds ratio, 10.8; 95% confidence interval range, 1.4-84). 36% of patients with a gene mutation had had a thromboembolic event compared with 8% of patients without a gene mutation. Thromboembolic events were more common among patients with idiopathic osteonecrosis (17%) compared with the general population (4%) and with patients with osteonecrosis caused by corticosteroid medication or alcohol abuse (7%). PMID- 15292804 TI - Intraoperative complications of revision hip arthroplasty using a porous-coated, distally slotted, fluted femoral stem. AB - Intraoperative complications of 175 cementless revision total hip arthroplasties done at four institutions using a porous-coated, uncemented, distally slotted, fluted femoral stem were reviewed. Three types of complications were recorded: eccentric reaming, femoral perforation, and femoral fracture. Intraoperative complications occurred in 16 patients (9.1%). There was no statistically significant association between complication rate and type of surgical approach, stem length, stem diameter, or host bone quality. This complication rate is comparable to or lower than that reported with the use of similar uncemented long femoral revision stems. PMID- 15292805 TI - Incisional hernia after periacetabular osteotomy. AB - The incidence of incisional abdominal hernias, an unreported complication after a Bernese periacetabular osteotomy, was evaluated. Two cases of an incisional hernia above the iliac crest were detected in a series of 950 cases since 1984. Although the incidence has been small, risk factors may be obesity, weak abdominal muscle strength, or increased abdominal pressure attributable to chronic coughing or obstipation. The surgeon should recognize the importance of restoring continuity of the abdominal fascia in patients with such factors. PMID- 15292806 TI - Use of the Cusum technique for evaluation of a CT-based navigation system for total knee replacement. AB - Most of the early failures of total knee replacements are related to technical flaws. Conventional ancillary devices achieve good alignment in the frontal plane in only an average of 75% of total knee replacements. Computer-assisted surgery may improve the technical quality of implantation surgery. The aim of our study was to evaluate the use of computer-assisted surgery using a quality control process. Seventy-eight total knee arthroplasties were done with a CT-based computer-assisted surgery system. The outcomes studied were alignment of the lower limb, implant positioning, and operative time. The target for alignment was 180 degrees +/- 3 degrees. Cusum analysis showed that the three outcomes were controlled during the study. The cusum test identified any existing outliers. Because few data were available at the beginning of this study regarding computer assisted surgery for total knee replacement, a randomized study was not relevant. However, a control of the procedure was mandatory. The cusum technique allowed continuous evaluation of the performance of the new procedure, and is a useful tool in assessing new technology. The results of this study showed that it is possible to do a randomized study to determine if computer-assisted surgery can improve the technical result of total knee replacement. PMID- 15292807 TI - Neuromuscular dynamic restraint in women with anterior cruciate ligament injuries. AB - The purpose of this study was to identify neuromuscular characteristics related to dynamic restraint in the knee. Observing compensatory changes to these characteristics in women with anterior cruciate ligament injuries provides important information for understanding functional knee stability, injury prevention, and performance. Twelve female subjects with anterior cruciate ligament injuries and 17 female control subjects participated in this study to assess electromyographic activity during landing from a hop and knee perturbation; hamstring muscle stiffness and flexibility; and isokinetic strength. Females with anterior cruciate ligament deficiencies had significantly increased preparatory muscle activity in the lateral hamstring before landing, but no differences in reactive muscle activity during landing or reflex latency after joint perturbation. Females with anterior cruciate ligament deficiencies had significantly less hamstring muscle stiffness and flexibility, but also had greater peak torque and torque development for knee flexion. Lower Lysholm scores were observed in females with anterior cruciate ligament deficiencies but no difference was found in functional performance of the single leg hop test. These neuromuscular characteristics provide a foundation for future research investigating injury prevention and rehabilitation techniques that maximize dynamic restraint through stiffness regulation and the timing of specific muscle activation strategies. PMID- 15292808 TI - Reoperations after 3200 revision TKAs: rates, etiology, and lessons learned. AB - The purpose of this study was to evaluate the prevalence, etiology, and evolution with time of reoperations done after index revision total knee arthroplasties. After exclusion criteria were met, 1814 index knee revisions in 1627 patients were included in the final cohort with more than 50% of the revisions done for loosening and extensor mechanism problems and more than 80% of the revisions involving the femoral, tibial, or both components. Three hundred seventy-three knees subsequently have been reoperated on one or more times. The average time from index revision total knee arthroplasty to the first reoperation was 3.5 years (range, 1 day-19 years). Of the 1814 index revision total knee arthroplasties, 373 (20%) had 593 reoperations in 336 patients. The cumulative risks of first reoperation at 5, 10, and 15 years were 16.1% (95% CI, 14.2, 17.9), 26% (95% CI, 23.4, 28.6), and 31.4% (95% CI, 30.2, 39), respectively. There was no difference in risk to first reoperation when comparing the decades in which the index revisions were done (1970-1980, 1981-1990, and 1991-2000). There was a trend toward a higher cumulative risk of first time reoperations for deep infection, loosening, and instability in the last decade, but with the numbers available this was not statistically significant. The prevalence of reoperations in this large series of index revision total knee arthroplasties done for aseptic reasons was surprisingly high. Despite substantial improvements during the past 3 decades in component design, surgical technique, and prevention of infection, patients who have a revision total knee arthroplasty are at substantial risk of having one or more subsequent problems that result in a reoperation. PMID- 15292809 TI - Dislocation of the rotating platform after low contact stress total knee arthroplasty. AB - From a one-surgeon series of 2485 patients, we report on 10 patients with rotating platform dislocation after primary Low Contact Stress total knee arthroplasty. All dislocations occurred within 2 years of the index procedure. Of the 10 patients, nine required open reduction. Five of these patients also had exchange of the original insert. One patient was treated by closed reduction. All knees were immobilized in a cast for 8 weeks. Eight of the 10 patients had no additional dislocation and at followup (average, 35 months; range, 12 months-5 years), had a stable functional joint. Two patients had recurrent spinout of the rotating platform develop. One patient had arthrodesis whereas the other patient had the insert cemented to the tibial tray as a salvage procedure. Increasing age, a preoperative valgus deformity, and prior patellectomy were significantly associated with rotating platform spinout. Surgical experience and an improved understanding of the soft tissue constraints, particularly in the valgus knee, are important in minimizing this complication. PMID- 15292810 TI - Malignant tumors of the pelvis: an outcome study. AB - This is a report of outcomes after a review of the demographic, diagnostic, therapeutic, and survival data for patients with pelvic primary and secondary tumors treated during the past 28 years. Using a computerized system it has been possible to assess the results for 206 patients with bone and soft tissue sarcomas and metastatic carcinomas to define the variation in outcome and the factors which statistically show an effect on survival. The data were compared with data for other anatomic sites. Based on our study, it is apparent that the outcome for all the tumors was approximately 50% survival with only soft tissue sarcomas having a poorer result. There were only minimal to moderate differences in outcome on the basis of gender, age, type of surgery, or adjunctive therapy. Patients who had intralesional surgery did less well as did patients with higher Musculoskeletal Tumor Society stages. Comparing the results for these patients with results for patients with the same stage and diagnosis but with tumors located in other sites showed significant differences. Results for patients with pelvic allograft compared with results of patients who had femoral allografts for the same diagnoses showed a poorer outcome for the patients who had pelvic allografts. Several possible explanations are provided for these variations in results. PMID- 15292811 TI - Malignant fibrous histiocytoma at the site of a total hip arthroplasty. AB - Malignant fibrous histiocytoma is the most frequent sarcoma in adults. Predisposing factors for malignant fibrous histiocytoma are Paget's disease, bone infarcts, malignant disorders of the hematopoetic system, or prolonged intake of corticosteroids. Malignant fibrous histiocytoma has been described as occurring with increasing frequency after endoprosthetic therapy and has been attributed to the implants or to their alloy constituents. Malignant fibrous histiocytoma at the site of an endoprosthesis of the hip constitutes a distinct rarity. To our knowledge, only 13 cases have been described to date. In this report, we present the case of a 66-year-old woman with rheumatoid joint disease. Eight years after primary endoprosthetic surgery, loosening of the implant with severe osteolysis of the surrounding bone required replacement surgery. Histopathologic evaluation of resected tissue revealed scar and granulation tissue and Grade 3 malignant fibrous histiocytoma. The patient died 1 year after revision arthroplasty because of diffuse pulmonary and cerebral metastases. In patients with loosening of a total hip endoprosthesis in combination with severe periprosthetic osteolysis an accompanying malignancy should be in the differential diagnosis. The histopathologic examination of the resected tissue should be obligatory. PMID- 15292812 TI - Clinical spectrum of acute compartment syndrome of the thigh and its relation to associated injuries. AB - The reason for the described clinical variability of acute compartment syndrome of the thigh, with high morbidity and mortality in some patients and an uncomplicated clinical course in others, is not known. To better define the clinical spectrum and factors determining the clinical course of this rare clinical entity, we did a retrospective multicenter study of 28 patients with 29 thigh compartment syndromes. The leading cause of acute thigh compartment syndrome was blunt trauma from motor vehicle accidents (46%) or contusion (39%). Pain with passive motion was present in all patients who were conscious, followed by paresthesia (60%), and paralysis (42%). The anterior compartment was involved most frequently with mean compartment pressure of 58 +/- 3 mm Hg. Myonecrosis, sepsis, and need for skin grafting were observed more frequently in patients with ipsilateral femur fracture. Only 7% of patients with isolated thigh compartment syndromes had short-term complications compared with 57% of patients with ipsilateral femur fractures. The incidence of complications correlated with the time to fasciotomy. Mortality was limited to patients with high injury severity scores. The clinical spectrum of thigh compartment syndrome is comparable with that of other compartment syndromes and its clinical course is determined by its associated injuries. PMID- 15292813 TI - Protease expression in interface tissues around loose arthroplasties. AB - To determine whether cathepsins and matrix metalloproteinase-1 are involved in accelerating tissue destruction, we examined, immunohistochemically, the expression of matrix metalloproteinase-1 and cathepsins B, D, L, and X in periprosthetic synovial-like interface tissues from 14 patients with failed prosthetic hips and in the synovial membranes of hips from 18 patients with rheumatoid arthritis and 25 patients with primary osteoarthritis. The expression levels of all these proteases in the interface tissue were higher than in the synovial membrane of osteoarthritis. The expression levels of cathepsins B and X in the interface tissue were higher than in the rheumatoid synovium. The results show similarities in the expression patterns of cathepsins D and L and matrix metalloproteinase-1 between aseptic prosthetic loosening and rheumatoid arthritis. In addition, these data suggest that the impact of cathepsins B and X on tissue degradation is more pronounced in aseptic prosthetic loosening than in rheumatoid arthritis. PMID- 15292814 TI - Feasibility of percutaneous gene transfer to an atrophic nonunion in a rabbit. AB - Treatment of atrophic nonunions is a challenge to orthopaedic surgeons. Growth factors potentially are valuable factors for improvement of tissue healing. The use of growth factors, however, is limited by their short half-lives. Gene therapy has the potential to improve the treatment. This study aimed to establish and validate an atrophic nonunion model in a rabbit for the use of a percutaneous in vivo gene therapy protocol. An atrophic tibial nonunion was established in 24 New Zealand White rabbits. Radiologic and histologic followup was for 64 weeks. The rabbit tibias showed no radiologic or histologic signs of healing. In addition, an adenoviral vector carrying a marker gene was injected percutaneously into the nonunion site in 12 rabbits. Expression of the marker gene was assessed for as many as 4 weeks. The percutaneous gene delivery resulted in transgene expression in the nonunion site for as many as 4 weeks. The described model reliably leads to an atrophic tibial nonunion in rabbits. Adenoviral percutaneous gene delivery into the nonunion site is feasible and leads to transgene expression locally for at least 1 month. This study provides investigators with a reliable and reproducible model of an atrophic nonunion. PMID- 15292815 TI - A simple murine model for immobilization osteopenia. AB - Reduction of loading force to bone induces osteopenia. Although the tail suspension model is the frequently used osteopenia model, this model burdens the animals with nonphysiologic blood distribution and systemic stress. We developed a new simple animal model for osteopenia under reduced loading. Both hind legs of male ICR mice (8 weeks old) in the experimental group were inserted into plastic tubes, which then were connected with wires of three sizes. This apparatus completely immobilized both femurs while allowing the tibias to move. Animals were pair-fed and sacrificed on Days 3, 7, 10, and 14 after immobilization. Bone mineral density measurement with dual energy xray absorptiometry revealed bone loss in the immobilized femurs. Histomorphometric analysis showed increased bone resorption and decreased bone formation, which started from Day 7 and continued until Day 14, resulting in structural disorders in the cancellous bone. Osteoclast population increase before osteoblast population decrease revealed that osteoclasts initially affect the process of this type of osteopenia. Our immobilization model is simple, easy to use, well-tolerated by the animal, and has potential for evaluating the therapeutic effects of drugs to treat osteopenia caused by reduced loading. PMID- 15292816 TI - Iliolumbar veins have a high frequency of variations. AB - The spectrum of individual anatomic variations of the vascular structures are broad, however, the exact incidence of variations of the lumbosacral vein is obscure. In the current study, 38 iliolumbar veins were dissected from 19 formaldehyde-preserved male cadavers. The drainage pattern of the iliolumbar vein was determined. The diameter and the length of the iliolumbar vein were measured, and the relationships of the iliolumbar vein with the lumbosacral trunk, obturator nerve, and iliolumbar artery were ascertained. Means and standard deviations were used as descriptive measures to define variations among the cases. The iliolumbar vein or veins were detected in both sides of all 19 cadavers. Five drainage patterns were seen between the iliolumbar vein and the lumbosacral major veins. In only five cadavers, symmetric drainage patterns were seen on the left and the right sides. In our study, two drainage patterns were seen that were not previously reported. Anatomic variations of the iliolumbar vein are numerous and should be considered to avoid complications when doing surgery. PMID- 15292817 TI - MD-PhD students in a major training program show strong interest in becoming surgeon-scientists. AB - A wide spectrum of individuals have discussed the importance of promoting research in orthopaedics and of developing clinician-scientists (physicians who also do significant research) in the field. Although orthopaedic research may benefit from recruitment of MD-PhD students as clinician-scientists, it is unclear to what extent MD-PhD students are interested in pursuing research and surgical specialties concurrently. To better understand their professional goals, all MD-PhD students enrolled in our institution's training program were invited to complete an online questionnaire concerning training satisfaction and future career goals. Twenty-four percent of respondents (57.5% response rate of 167 recruits) reported a primary clinical interest in a surgical field (3% interest in orthopaedics); interest was strongest late in training. The majority of surgical MD-PhD students, like nonsurgical students, were planning to make research a significant part of their careers. In addition, students identified the importance of factors such as family issues and faculty role models in determining their clinical interests. The study data indicate that MD-PhD students have strong interests in becoming surgical clinician-scientists. They also suggested that active recruitment (especially early in training) that is responsive to the personal and professional needs of students has the potential to increase the number of clinician-scientists in orthopaedics. PMID- 15292818 TI - Modulation of distraction osteogenesis in the aged rat by fibroblast growth factor. AB - This study explores a series of hypotheses related to modulation of bone formation using the distraction model. The tibial lengthening model was scaled down from dog to rat to use immunohistochemistry, in situ hybridization, and reverse transcription-polymerase chain reaction to evaluate cellular events during in vivo bone formation. Different delivery systems such as oral, intragastric, intravenous, subcutaneous, and local diffusion by either extraperiosteal or intramedullary routes, were developed and standardized. Systemic modulators, including diet (total enteral nutrition, calcium, phosphate, soy, whey, casein, lead, and alcohol) and hormones (estrogen, testosterone, growth hormone, and gonadectomy), were tested. To investigate the effects of aging on bone formation, rats of different age groups had tibial lengthening. The aging effect could be distinguished by a reproducible deficiency of endosteal bone formation, consistent with similar deficits in older adult patients having distraction osteogenesis or in patients with senile osteoporosis. Expression of endogenous fibroblast growth factor-2 at the cellular level during the coupled osteogenesis and angiogenesis in young rats was dramatically diminished in old rats. Exogenous fibroblast growth factor-2 reversed the endosteal deficits found in old rats having distraction osteogenesis. PMID- 15292819 TI - Knee pain in a 12-year-old girl. PMID- 15292820 TI - Femoral head resurfacing for the treatment of osteonecrosis in the young patient. PMID- 15292822 TI - [New antifungal agents and bronchopulmonary mycoses]. AB - The frequency of respiratory mycosal infections has increased over recent Years. Diagnosis has been improved by recent epidemiological data and advances in radiological and mycological diagnostic methods. Two new antifungal agents have recently received marketing approval: voriconazole and caspofungine. Voriconazole belongs to the echinocandin family of antifungals. Sites of action of antifungals have become more diversified: amphotericins act on ergosterol directly, azolated agents act on the synthesis of ergosterol, flucytosine affects synthesis of nucleic acids, and echinocandins alter the fungal wall. Synergetic or additive combinations, such as amphotericin-caspofungine, or voriconazole-caspofungine, can be proposed for advanced disease. Thus both first intention and secondary treatments, particularly for systemic candidiasis and aspergillosis, have been modified. These new protocols take into consideration the severity of the mycosal infection, co-morbidity, and drug combinations as well as cost. PMID- 15292823 TI - [Imaging pulmonary embolism]. AB - The diagnostic performance of computed tomography images of pulmonary embolism is directly related to the acquisition parameters. Any physician evaluating these scans must have proper knowledge of the acquisition, injection, reconstruction, and radiation parameters. Cardiac gating and morphological and functional image processing should be understood since they are now routine techniques particularly important for preoperative assessment of chronic thromboembolism. Elementary knowledge of the imaging techniques reduces the risk of diagnostic limitations. Understanding these techniques does not require any particularly advanced knowledge of physics, data processing or technology, but is necessary to chose the appropriate technical facilities and equipment adapted for diagnostic purposes. While specific training is not a prerequisite, interpretation of an angioscan of the pulmonary vessels does require precise knowledge of the pulmonary anatomy in addition to the technical knowledge mentioned above. Proper analysis may reach the 4th and 5th generation vessels. Different analysis methods have been developed which take into account the technical parameters and avoid the need for serial images. Each slice can then be analyzed within an acquisition Volume. Differential diagnosis is also very technique-dependent, minimally operator-dependent but highly machine-dependent. Differential diagnosis becomes less and less a problem with advancing equipment. Sufficient knowledge of the physiological and pathogenic basis is relatively easy to retain. PMID- 15292824 TI - [Reactive airway dysfunction syndrome: more flexible application of diagnostic criteria are important for occupational accident victims]. AB - Reactive airway dysfunction syndrome (RADS), or Brooks syndrome, is a complication observed after inhalation of caustic or highly irritating substances. The diagnosis is based on a group of criteria which include the absence of prior respiratory disease. Strict application of these criteria could have a prejudicial effect for certain victims. Three serious cases of RADS were observed in workers who were exposed to massive inhalation of caustic substances. The products implicated (phosphoric oxychloride, titanium tetrachloride, and trichloroacetyl chloride) hydrolyze to hydrochloric acid when they come in contact with the airway mucosa. After an initial period of acute respiratory distress, the patients encountered serious difficulties in achieving an appropriate diagnosis, and in having their sequellae recognized as resulting from an occupational accident. The problem was that these patients had a history of cured allergic asthma or smoking-related COPD. The presence of prior respiratory disorders must not exclude the diagnosis of RADS. A prior respiratory disorder cannot be used as an argument to exclude such victims from indemnities for occupational accident sequelae. PMID- 15292825 TI - [The "Compli'Asthme" therapeutic observation survey on good use of inhaled drugs for asthma: perception by general practitioners]. AB - A cross sectional survey was conducted in 2000 in coordination with the CHIESI medical representatives among 1758 French physicians caring for patients with persistent asthma (80% general practitioners, 20% specialists). This "Compli'Asthme" survey was based on a self-administered questionnaire designed to learn more about the physicians' experience with good use of inhaled drugs and to collect information on therapeutic observance, corticophobia, and use of prescribed inhalers. Poor observance was noted as an important problem by 58-85% of the participants. Most of the problems were related to inability to use the inhaler properly (children, elderly subjects) or to patients forgetting to take their medications (adults, parents). For 58% of the participating physicians, corticophobia is frequent. The patients are worried about the anabolizing effect, secondary effects, and dependence. When there is a potential problem with corticophobia, physicians generally question the patients and provide explanations to achieve good observance. Patient preference is taken into consideration by 86% of the physicians prescribing inhalation devices; 90% demonstrate use of the device at the first prescription and 68% make repeated demonstrations at subsequent consultations. For 56-87% of the physicians, poor therapeutic observance, corticophobia, and poor use of the inhaler can be detected and corrected. Patient education is an important element for 77% of the physicians for improving observance and achieving good use of the inhaler. When poor observance and poor use of the inhaler occur, the physicians responding to this questionnaire applied the currently recommended guidelines. PMID- 15292826 TI - [An unusual cause of acute respiratory distress: obstructive bronchial aspergillosis]. AB - We report the case of a 77-Year-old immunocompetent woman who required intensive care for acute dyspnea revealing complete atelectasia of the left lung related to an aspergillus mycelium plug blocking the principal bronchus. The clinical course was favorable after deobstruction by thermocoagulation and oral itraconazole given for six Months. The patient was free of parenchymatous or endobronchial sequelae. Adjuvant oral corticoid therapy was given temporarily during the second Month of treatment when signs of transition towards allergic aspergillosis developed. Four Months after discontinuing the antifungal treatment, the patient developed a new episode of acute dyspnea caused by atelectasia limited to the right lower lobe. Treatment by itraconazole was resumed and continued as long term therapy. No recurrence has been observed for eighteen Months. The diagnostic and therapeutic problems raised by Aspergillus fumigatus are well known in the immunocompromised subject, but can also be encountered in the immunocompetent subject. PMID- 15292827 TI - [Fibrous tumor of the pleura]. AB - Pleural fibroma, or fibrous tumor of the pleura, is an uncommon entity which is characterized by slow proliferation of undifferentiated, intermediary or mature fibroblasts associated with collagen fibers forming a tumor stroma. We report a case in a 49-Year-old man who developed exercise-induced dyspnea and right chest pain. The thoracic CT scan revealed the presence of a mass in the right lung base composed of heterogeneous encapsulated tIssue. Tumor resection was performed leading to the histological diagnosis of pleural fibroma. Immunohistochemistry tests revealed positive vimetin and CD34, and negative cytokeratin uptake. These immunohistochemistry data contributed to the differential diagnosis with malignant pleural mesothelium. Pleural fibroma is a benign tumor in 80% of the cases. Prognosis is excellent. Local recurrence is exceptional and generally occurs after incomplete resection. Radial surgical treatment determines the prognosis and is required to prevent local recurrence. Other criteria of malignancy are not correlated with the clinical course of this type of tumor. PMID- 15292828 TI - [Coagulation disorders and arterial thromboembolic events in non-small-cell lung cancer. A case report]. AB - Certain coagulation disorders can occur in patients with cancer and thromboembolic complications are frequent. We report the case of a 53-Year-old patient with metastatic adenocarcinoma of the lung treated with chemotherapy who presented several cerebral arterial thromboembolic events leading to death a few weeks after the initial diagnosis of cancer. This case illustrates the important role of certain satellite disorders related to coagulation activation: non bacterial thrombotic endocarditis, disseminated intravascular coagulation, anti phospholipid antibody syndrome. The role of anticancer chemotherapy as a favoring factor for thromboembolic events is also emphasized in patients with non-small cell lung cancer. PMID- 15292829 TI - [Cerebral metastasis(es) of non-small-cell lung cancer. Contribution of stereotactic cerebral radiotherapy]. PMID- 15292830 TI - [Therapeutic education of the asthmatic child]. PMID- 15292833 TI - [Concertation is needed to apply health programs against chronic respiratory diseases in Africa]. PMID- 15292834 TI - [Is the therapeutic index a key to therapeutic strategy? From theory to practice]. PMID- 15292835 TI - [The respiratory physician, the lung, and the liver]. PMID- 15292836 TI - [Home mechanical ventilation in France]. PMID- 15292837 TI - [Palliative care of dyspnea at the end of life]. PMID- 15292838 TI - [The new anticoagulants are here!]. PMID- 15292839 TI - [Respiratory physicians unite to organise the fight against tuberculosis]. PMID- 15292840 TI - [Application of the WHO chronic respiratory diseases programme in Sub-Saharian Africa: problems in Senegal]. AB - INTRODUCTION: To prevent and control chronic respiratory diseases, the WHO has outlined a programme to be applied in all the countries. In Sub-Saharan countries like Senegal, its application faces many barriers. METHODS: This work is a detailed analysis of the main areas of the programme based on the socio-economic, cultural and medical realities present in Sub-Saharan Africa countries, and particularly in Senegal. RESULTS: There is a lack of political engagement resulting from the precarious socio-economical situation. The demand for care is impaired because of the prevalence of illiteracy and the practice of traditional healing methods which is the first recourse for patients. The inaccessibility and lack of adequate health structures, technical equipment and qualified medical staff, and the high cost of medications for the majority of the population is the cause of this weakness of the healthcare system. These international recommendations are both not compatible with the local realities and not known to the majority of health providers. CONCLUSIONS: The mobilisation of international and national human and financial resources, the involvement of community-based structures, the adaptation and revision of the recommendations, the introduction of medications advocated by the Bamako Initiative, the training of health providers, the adaptation of regional and continental initiatives against tobacco and the creation of smoking cessation centres are needed if this chronic respiratory diseases programme is to be efficiently implemented. PMID- 15292841 TI - [The costs of asthma in France: an economic analysis by a Markov model]. AB - INTRODUCTION: In France about 3.5 million people suffer from asthma, with an estimated annual cost of 1.5 billion euros. Few data on the components of the cost are available. To develop a medico-economic model to identify the mean annual cost of a patient with asthma together with the components of the cost and the consequences of different modalities of management (out-patient and in patient). METHODS: A first degree homogeneous Markov model was constructed, based on published data, to identify the different states of health of asthmatic patients, their development and probabilities of change from one to another. RESULTS: The mean annual cost of asthma is 631 +/- 299 euros; 298 euros for stable patients, 1,052 for those who have at least one acute attack and 3,811 for those hospitalised. Actions leading to better control of the disease in the community (prevention and management of attacks) would produce a larger reduction in mean costs than actions undertaken in patients seen as an emergency or admitted to hospital. CONCLUSIONS: From the point of view of society better control of asthma in the community is the most effective strategy to reduce expenditure. PMID- 15292842 TI - [The cost components of the management strategies for lung cancer in France]. AB - INTRODUCTION: To evaluate the average cost of therapeutic strategies for the management of lung cancer in relation to histological type and diagnostic staging and of the individual components of the management strategy. METHODS: Samples were taken between 1 September 1998 and 30 June 1999 from centres with sufficient numbers of lung cancer (LC) cases. All events over an 18 Month period were collected from a retrospective analysis of the records. A Markov model was constructed based on decision branches for localised and diffuse small cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC). Several components of management were identified: first line treatment, second line treatment, observation, terminal care and death. RESULTS: The average cost of LC was 22,006 Euro (10,631-36,296) for one year and 25,643 Euro (10,631-46,191) for two years. For SCLC the average annual costs were 22,420 Euro for diffuse disease and 27,098 for localised disease. For NSCLC the totals ranged from 19,543 Euro for inoperable stage I and II tumours to 39,424 for operable tumours. The cost for stage IV tumours was 24,383 Euro. The cost components over two Years varied according to the tumour type. The cost of diagnosis ranged from 6-14%, the cost of management and of terminal care from 33-45% of the total. Analyses of sensitivity confirmed that whatever the histological type or diagnostic staging the percentage of patients initially treated actively (that is to say not by palliative care) had the greatest effect on the total cost, greater than the costs of terminal care and of two courses of chemotherapy. CONCLUSIONS: This model has allowed for the first time the calculation of the contributions of the different therapeutic components to the total cost of the management of lung cancer in France. In the future it will allow analysis of the economic impact of new methods of treatment. PMID- 15292843 TI - [The therapeutic index in asthma: how should it be defined?]. AB - INTRODUCTION: The therapeutic index (efficacy/tolerance or benefit/risk ratio) is a major determinant of treatment decisions in asthma. METHODS: For the numerator, the therapeutic index depends on efficacy (maximal effect) and not potency (dose response relationship). With regard to the denominator, several pharmacological factors influence the occurrence of side-effects, the acceptability of which also has to be considered. RESULTS: In asthma, some strategies have a more favourable therapeutic index than others;e.g additional treatment (long acting beta2 agonists, leukotriene receptor antagonists, theophylline) to inhaled corticosteroids instead of doubling the dose of the latter. Conversely, it is extremely difficult to compare the therapeutic indices of different molecules of inhaled corticosteroids. CONCLUSIONS: The potential risk of systemic side effects with long-term administration of high doses of inhaled corticosteroids suggests the need to seek the minimal effective dose. PMID- 15292844 TI - [Molecular basis of the primary ciliary dyskinesias]. AB - INTRODUCTION: The primary ciliary dyskinesias (PCD) are rare diseases characterised by infection of the airways due to impaired muco-ciliary clearance. Half the patients have situs inversus making up Kartagener's syndrome. STATE OF THE ART: Primary cilia play a role in development. In the adult ciliated cells occur mainly in the airways and the genital tract. The axoneme, the internal structure of the cilia, is made up of a central pair of microtubules surrounded by peripheral doublets carrying the inner and outer dynein arms. These multiprotein complexes are composed of chains of dynein whose ATPase activity is the basis of ciliary movement. Structural and functional abnormalities of the respiratory ciliated cells are the cause of PCD, diseases that are heterogeneous at both the genetic and ultrastructural levels. PERSPECTIVES: There are more than two hundred axonemal proteins. The synthesis and assembly of these proteins are controlled by transcription factors and intraflagellar transport molecules respectively. The genes that code for these proteins are as numerous as candidate genes for PCD. CONCLUSIONS: To date only two dynein genes, DNA11 and DNAH5, have been implicated and only in individuals suffering from PCD with absence of outer dynein arms. PMID- 15292845 TI - [Targeted therapies in thoracic oncology]. AB - INTRODUCTION: Better understanding of the alterations of cellular physiology during carcinogenesis has resulted in the development of new anticancer agents called biological targeted therapies. These therapies will probably complement the traditional treatments (surgery, radiotherapy and chemotherapy). STATE OF THE ART: In thoracic oncology, targeted therapies may interfere with signal transduction by interaction with growth factors receptors. This is the case with monoclonal antibodies directed against the extracellular part of the receptors and with small molecules inhibiting the intracellular part of the receptors (for example, EGF-R). PERSPECTIVES: Other strategies include the use of farnesyl transferase inhibitors or of antisense oligonucleotides; the new therapies may also inhibit angiogenesis by targeting either the VEGF receptor or the matrix metalloproteases. Inhibitors of metalloproteases were the first targeted agents tested. However, all published studies on metalloproteases inhibitors have been negative so far. CONCLUSIONS: Currently, the agents most advanced in clinical development are the EGFR inhibitors (either monoclonal antibodies or small molecules inhibiting tyrosine-kinases) which, in those that have clinical activity, may produce a sustained response at a cost of a degree of toxicity. PMID- 15292846 TI - [Tobacco smoke and risk of bacterial infection]. AB - INTRODUCTION: Tobacco smoke is a proven risk factor for bacterial infection. STATE OF THE ART: In adults without COPD, smoking is associated with a significant increase in the relative risk (RR) of pneumonia (RR=2.97; 95% CI 1.52 5.81), S pneumoniae pneumonia (RR=2.50; 95% IC 1.50-5.10), Legionella infection (RR=3.75; 95% CI 2.17-6.17). Smoking has clearly been shown to be associated with an increased risk of tuberculosis (RR=2.60; 95% CI 2,20-3,20), and also with increased incidence of post-operative infections. In young children whose parents smoke, passive exposure to tobacco smoke is associated with an increased relative risk of seasonal infections (RR=1.7; CI 95% 1.55-1.91) and recurrent otitis media (RR=1.48; 95% CI 1.08-2.04). Passive smoking also increases risk of pneumonia in adults (RR=2.5; CI 95% 1.2-5.1). Plausible explanations of the increased risk of infection in active or passive smokers include increased bacterial adherence, decrease of lung and nasal clearance, and changes in the immune response. CONCLUSIONS: Exposure to tobacco smoke approximately doubles the risk of infection. This increased burden of infection has significant healthcare cost implications. Each infectious episode in an individual should prompt an attempt at smoking cessation. PMID- 15292847 TI - [Involvement of dendritic cells in allergic airway diseases]. AB - INTRODUCTION: Dendritic cells (DCs) are able to present antigen to T lymphocytes and to orientate the immune response towards a Th1 or a Th2 type. STATE OF THE ART: The aim of this report is to present different studies comparing DCs from allergic patients with those from healthy subjects using in vivo and in vitro experimental conditions. These studies have demonstrated that cellules dentritiquess from house dust mite allergic patients:i) take up house dust mite allergen more efficiently, ii) after stimulation by house dust mite allergen, secrete a restricted panel of cytokines and chemokines, and express characteristic co-stimulatory molecules, favoring a Th2 profile, iii) after stimulation by house dust mite allergen, induce Th2 cytokines secretion by T lymphocytes, and iv) favor an allergen-specific Th2 airway inflammatory response in an in vivo model of humanized mice. PERSPECTIVES: Functional modulation of DC could be a new therapeutic concept in allergic airway diseases. CONCLUSIONS: These results show phenotypic and functional specificities of DC from house dust mite allergic patients, and suggest a key-role for cellules dentritiques in the pathogenesis of allergen-dependent airway inflammatory response. PMID- 15292848 TI - [Practical and technical aspects of noninvasive ventilation]. AB - INTRODUCTION: Noninvasive ventilation refers to the delivery of positive pressure ventilation via a mask or "interface" rather than via an invasive conduit. Until recently, equipment for noninvasive ventilation was frequently custom-made to meet the needs of an individual patient. During the past 15 years, there have been significant advances in the equipment available for noninvasive ventilation. STATE OF THE ART: Interfaces that have been designed specifically for noninvasive ventilation are now commercially available from several manufacturers. Commonly used interfaces include nasal and full face masks, and mouthpieces. The main characteristics, and potential advantages and disadvantages of each interface are described. Portable volume-limited or pressure-limited ventilators are available for home noninvasive ventilation. As with critical care ventilators, home mechanical ventilators are capable of delivering a variety of modes of ventilation. Furthermore, they are lightweight and economical. Technical aspects of ventilator circuits are also discussed here and some practical considerations about selection and maintenance of materials are proposed. CONCLUSIONS: Although major technical advances have been made, optimal delivery of noninvasive ventilation requires knowledge of, and experience with, the application of the equipment used. PMID- 15292849 TI - [Lung volume reduction surgery for emphysema: a unilateral or bilateral approach?]. AB - INTRODUCTION: Lung Volume reduction surgery (LVRS) is a recognized therapeutic option for patients presenting with severe and disabling pulmonary emphysema. Case selection is based upon clinical, morphological and functional criteria. STATE OF THE ART: LVRS has shown promising results, with improvements in exercise capacity, pulmonary function and quality of life, in selected patients with severe and disabling emphysema. A variety of surgical techniques have been described. The procedure may be unilateral or bilateral, through a sternotomy or by a video-assisted thoracoscopic (VATS) technique. The controversial aspects of the surgical technique will be analysed and discussed in the following review. PERSPECTIVES: A bilateral approach clearly offers a better functional improvement when compared to a unilateral procedure, however, the postoperative functional decline appears greater and more rapid after a bilateral procedure. A unilateral approach, with often less postoperative morbidity, allows the option to perform a future contra-lateral procedure in the event of further clinical or functional deterioration. CONCLUSIONS: In selected cases LVRS is an effective treatment for severe pulmonary emphysema. Different surgical techniques have been described. Nowadays VATS is considered to be the technique of choice, with the option to carry out a future unilateral or bilateral procedure. PMID- 15292851 TI - [Contingency tables and the chi2]. PMID- 15292850 TI - [Effects of home pulmonary rehabilitation in patients with chronic respiratory failure and nutritional depletion]. AB - STATE OF THE ART: The IRAD2 trial is evaluating a 3-month home intervention which includes education, oral supplements, exercise and androgenic steroids in undernourished patients with chronic respiratory failure. The main objective is to increase the six-minute walking distance by more than 50 m with an improvement in health-related quality-of-life. Secondary end-points include a reduction in exacerbation rates by 25%, a reduction in health-related costs and an increase in survival during the year following intervention. MATERIAL AND METHODS: This interventional, multi-centre, prospective, two-armed parallel, controlled trial is being conducted in 200 patients. In both groups, "Control" and "Rehabilitation", 7 home visits are scheduled during the 3-month intervention for education purpose. In the "Rehabilitation" group, patients will receive 160 mg/d of oral testosterone undecanoate in men, 80 mg/d in women, oral dietary supplements (563 kcal/d) and exercises on an ergometric bicycle 3 to 5 times a week. EXPECTED RESULTS: In the event of significant responses to intervention, this trial would validate a comprehensive and global home-care for undernourished patients with chronic respiratory failure combining therapeutic education, oral supplements, androgenic substitution and physical activity. PMID- 15292852 TI - [Isolated pleural amebiasis: about one case in France]. AB - INTRODUCTION: Isolated pleural amebiasis is exceptional in the world and specially in France. CASE REPORT: We report a case of acute isolated pleural amebiasis in a 56 year old man who did not travelled in endemic countries for the 20 past years. Chocolate-colored pus from the pleural puncture suggested the diagnosis. Positive amebic serology and above all, trophozoites (Entamoeba histolytica) in the pleural liquid confirmed the diagnosis of pleural amebiasis. Drainage was required and metronidazole was introduced. Other antibiotics were necessary to treat bacterial co-infection, which is frequent. Of interest, a nurse developed an acute intestinal amebiasis probably infected by pleural pus, an indirect confirmation of diagnosis. CONCLUSION: This diagnosis should be suspected even in low endemic areas in case of infectious pleurisy resistant to traditional antimicrobial regimens. PMID- 15292853 TI - [Actinomycosis presenting as a chronic excavated pneumonia in a young woman]. AB - INTRODUCTION: Pulmonary actinomycosis is an anaerobic bacterial infection occurring primarily in debilated patients with poor oral hygiene. Before the penicillin era, thoracic actinomycosis looked like tuberculosis or neoplasia with chest wall invasion and fistula formation. OBSERVATION: We report the case of a 39 years old woman presenting with a chronic lung abscess of the left upper lobe hospitalised after several unsuccessful courses of antibiotics. The diagnosis was made after thoracic surgery. Three years after lobectomy, which had been followed by three months of amoxycillin and multiple dental extractions, there was no sign of relapse of the infection. CONCLUSIONS: We review the role of thoracic surgery, antibiotic treatment and diagnosis in pulmonary actinomycosis. PMID- 15292854 TI - [Disordered higher functions in a smoker]. AB - INTRODUCTION: Paraneoplastic syndromes sometimes lead to the discovery of an intrathoracic tumour, notably small cell lung cancer (SCLC). MATERIALS AND METHODS: We report the case of a patient presenting with a paraneoplastic syndrome characterised by disordered higher functions and convulsions, representing a paraneoplastic encephalo-myelitis (PEM). This PEM led to the diagnosis of SCLC. The diagnostic features and progress of the PEM are discussed. CONCLUSION: Recognition of PEM allows the diagnosis and early treatment of the underlying cancer, strongly influencing the prognosis, particularly in SCLC. PMID- 15292855 TI - [Bronchial presentation of stage IVB Hodgkins disease]. AB - INTRODUCTION: Bronchial localisation of Hodgkins disease has been described only rarely and it is exceptional as the presenting feature. More frequently the clinical presentation is extrinsic bronchial compression. OBSERVATION: Mr B.C., 34 years old, had a past history of cured alcoholism and abuse of cannabis and buprenorphine. In February 2001 he presented with generalised pruritus, weight loss of 8 kg in one year, a 1 cm node above the right clavicle, hypereosinophilia, and CT evidence of non-obstructive mediastinal adenopathy and interstitial lesions in the right lower lobe. Five months later he was admitted to hospital with a febrile illness. The right supraclavicular node had disappeared but there was gross mediastinal adenopathy, associated with a right perihilar soft tIssue mass encircling the upper, middle and lower lobe bronchi and multiple peripheral parenchymal nodules. Bronchoscopy revealed complete stenosis of the right upper lobe bronchus and bronchial histology confirmed Hodgkins disease. CONCLUSIONS: Bronchoscopy is an effective investigation for the detection of endobronchial Hodgkins disease, often overlooked because parenchymal abnormalities are attributed to extrinsic compression. In order to determine the disease stage more precisely we advise immediate bronchoscopy when there is suspicion that Hodgkins disease may be the cause of clinical. PMID- 15292856 TI - [Pneumothorax complicating legionellosis]. PMID- 15292857 TI - [Palliative care at the end of life in respiratory medicine: we should anticipate]. PMID- 15292858 TI - [Complications of portal hypertension: a consensus]. PMID- 15292859 TI - [Consensus conference on the management of hepatopulmonary syndrome. Complications of portal hypertension: how to treat pleuropulmonary complications]. PMID- 15292860 TI - [Management of hepatic hydrothorax. Complications of portal hypertension: how to treat pleuro-pulmonary complications?]. PMID- 15292861 TI - [Therapeutic management of porto-pulmonary hypertension. Complication of portal hypertension: how to treat pleuro-pulmonary complications?]. PMID- 15292862 TI - [Complications of portal hypertension in adults (Paris, December 4-5, 2003]. PMID- 15292863 TI - [Mechanical and functional properties of the thorax]. PMID- 15292864 TI - Protocol for the clinical management of dry mouth. AB - Dry mouth is a very common condition in dental practice, and the causes underlying this alteration in salivary secretion are diverse. The problem is particularly common in polymedicated elderly people. Treatment should aim to eliminate the background cause; however, when this is not possible, management should focus on the stimulation of salivation or the provision of a saliva substitute. The present study provides a systematic account of the management protocol for patients with dry mouth. PMID- 15292865 TI - Orofacial infections of odontogenic origin. AB - The polymicrobial nature of the odontogenic infections as well as the variety of associated conditions are a consequence of the diversity of the buccal microbiota and the anatomical and functional complexity of the oral cavity. In addition to this, all these processes can give way to multiple complications which range from the local to the systemic level. The appropriate choice of antibiotic and posology is crucial in the successful management of these infections. Pharmacodynamics provides those parameters that make it possible to assess how antibiotics activity varies in time. As a general rule, the first step in the initial management of orofacial infections in adults, included odontogenic infections, will be the administration of 875 mg of amoxicillin and 125 mg of clavulanic every 8 hours. Therapeutic compliance is paramount to avoid resistance, therefore patient acceptance must be sought. In this sense, it has been proved that Augmentine Plus (2000/125) every twelve hours both as profylaxis and as treatment significantly decreases the rate of infective complications associated to extraction of the third molar. PMID- 15292866 TI - DNA microarrays in oral cancer. AB - One of the principal aims of modern cancer research is to identify markers allowing individual prediction of prognosis or response to treatment. In this connection, the number of genes thought to be involved in the different stages of different types of oral cancer increases apace. DNA microarrays allow simultaneous evaluation of the expression of hundreds of genes in a single assay. The parallel format of microassay slides is designed to allow rapid comparison of gene expression between two samples, for example tumor cells and healthy cells. This article reviews studies that have aimed to identify genes related to oral cancer, and to classify these genes into groups that are commonly co-expressed. These studies suggest that DNA microarrays are set to become routine tools in the detection, diagnosis, characterization and treatment of oral cancers. PMID- 15292867 TI - Orofacial pain management: an update. AB - New treatments for orofacial pain have been developed in recent years. In the case of cluster headache, new drugs are now administered via the intranasal route, while in patients with chewing pain the topical application of capsaicin and the use of oral splints in combination with jaw movement exercises are the most widely used management approaches. In the case of neurogenic pain new anticonvulsivants have been introduced, with fewer side effects than carbamazepine. The latest pharmacological advances involve the use of nonsteroidal antiinflammatory drugs and opioids via the topical route, and the combination of different analgesics. The present review discusses the latest advances in the treatment of orofacial pain. PMID- 15292868 TI - Considerations on the diagnosis of oral psoriasis: a case report. AB - This paper discusses the difficulties in making a definitive diagnosis of oral psoriasis based upon clinical and histological evidence only. A young black male presented with multiple lesions showing erosions, fissures, and yellowish scales on the vermilion borders of both lips. He also had erythematous-erosive areas on the gingivae, a fissured tongue showing greyish areas on its ventral surface, whitish lesions and longitudinal sulci in the hard palate with lacelike lesions on the soft palate. Biopsies from the lower lip, gingiva and soft palate showed hyperkeratosis, spongiosis, acanthosis, and elongation of rete ridges. In addition, collections intraepithelial micro-abscesses of Munro were observed. These findings are consistent with oral psoriasis. Typical cutaneous lesions and a family history of psoriasis were absent. PMID- 15292869 TI - Necrotizing sialometaplasia: report of five cases. AB - Necrotizing sialometaplasia (NS) is a self-limiting inflammatory disease, that involves salivary glands, more frequently the minor ones. Although its etiopathogenesis remains still unknown some authors suggest that a physico chemical or biological injury on the blood vessels may produce ischemic changes, leading to infarction of the gland and its further necrosis. Its clinical and histologic feature resemble malignancy. Clinically it may appear like an ulcer with slightly elevated irregular borders and necrotic base. Histologic features are squamous metaplasia of ducts and acini and a pseudoepitheliomatous hyperplasia of the overlying mucosa. These characteristics may induce to an inapropiated diagnosis of malignant neoplasia. A correct diagnosis to avoid mutilant surgical treatments is essential, considering that it is a self-limiting disease. In this report we describe five cases of NS in females, located in minor glands of the palate. PMID- 15292870 TI - Evaluation of medical risk in dental practice through using the EMRRH questionnaire. AB - OBJECT: Due to the fact that the population is getting older and to new medical and dental techniques, the number of medical complications during treatment is tending to increase. In order to avoid these complications a correct clinical history should be obtained of all these patients. The search for a suitable questionnaire which would be able to take into account all these factors is therefore necessary. MATERIAL AND METHOD: In this study we have used the questionnaire EMRRH which has been proposed by a group of European investigators, in order to study the prevalence of past medical problems in a population that attends a dental clinic for treatment. RESULTS: A total number of 716 patients were registered. Of these, 219 had a medical history that was of some interest to us. There were significant differences between the average age of the population with or without former medical problems (p<0.0005). Secondly, out of the 30.6% of the population with medical problems (N=219), we separated into groups those patients who would have a low, medium or high risk of complications when submitted to dental treatment. 17.31% of the patients were classified as risk ASA II; 9.49% as ASA III, and 2.51% as ASA IV. Among the diverse pathologies the highest percentage was hypertension (13.8%), followed by allergies to different drugs (8.37%), palpitations (7.82%), respiratory pathologies (5.16%) and diabetes (4.3%). CONCLUSION: We can see the obvious need for a detailed medical history to be taken because of the existing prevalence of these above pathologies taking into account that without one we could seriously harm the patient health with our dental treatment. PMID- 15292871 TI - Distraction osteogenesis of the alveolar ridge: a review of the literature. AB - One of the principal problems in dental implantation is the lack of sufficient bone height or width. In the case of the alveolar ridge, a very effective technique for resolving this problem is distraction osteogenesis, introduced in this context about a decade ago. This technique is based on the gradual separation of a mobile but fully vascularized bone segment from the basal bone, leading to the formation of an intervening soft callus which gradually transforms to mature bone. A key researcher in the development of this technique was the Russian traumatologist Ilizarov. The present article reviews alveolar ridge distraction procedures and their clinical application. Alveolar ridge distraction may often be preferable to bone grafting or guided bone regeneration for increasing ridge height and width prior to implantation. PMID- 15292872 TI - Chronic Sialadenitis revealing hepatitis C: a case report. AB - One 53-year-old male was referred with a history of sensitive peripheral neuropathy and Raynaud disease leading to suspect a Sjogren syndrome (SS). Labial salivary gland biopsy shown the classical features of chronic lymphocytic sialadenitis. As clinical and immunologic tests were positive, we conclude to a chronic lymphocytic sialadenitis simulating SS. Enzyme immunoassay and recombinant immunoblot were positive to HCV. Circulating HCV-RNA was detected by PCR. Liver biopsy revealed chronic persistent hepatitis. In one second biopsy, RNA was extracted. PCR amplification and southern blot hybridization for HCV-RNA were performed. After a 8-month treatment by interferon alpha, HCV-RNA was no longer detected in the serum. There was an objective improvement of the Schirmer test. The detection of HCV-RNA in the salivary glands of a Sjogren like syndrome patient suggests that a direct infection of the salivary glands by HCV could play an important role in the pathogenesis of HCV related sialadenitis. PMID- 15292873 TI - Ossifying fibroma of the upper jaw: report of a case and review of the literature. AB - A number of processes generically referred to as benign fibroosseous lesions comprise different disorders such as fibrous dysplasia, sclerotizing osteomyelitis and ossifying or cementing fibroma. These processes are all characterized by the existence of a vascular fibroblastic stroma, with the production of a calcified matrix ranging from bone to cementum. Ossifying fibroma involves slow-evolving growth with deforming swelling generally arising in the mandible, with possible early dental displacement. From the radiological perspective the disorder generally manifests as a well defined and delimited, unilocular radiotransparency, as a radiotransparent image with central opacifications, or as multilocular transparencies. The lesions exceptionally can be radiopaque. We present the case of a 22-year-old male presenting for evaluation of a three-month, asymptomatic tumor mass in the anterior sector of the upper jaw. Radiologically, the lesion appeared as a radiotransparent zone surrounded by a poorly delimited sclerotic halo. The definitive diagnosis following surgical resection of the lesion was ossifying fibroma. The case is discussed, and a review is provided of the literature on the subject. PMID- 15292874 TI - Odontoameloblastoma: a case report and a review of the literature. AB - Odontoameloblastoma (OA) is an extremely rare mixed odontogenic tumor appearing within the maxillary bone, with both epithelial and mesenchymal components. The term odontoameloblastoma (OA) was included in the 1971 WHO classification. Only 23 well-documented cases have been reported in the medical literature. Because of their rarity, controversy exists in the treatment of this tumor. We present a new case of OA involving the mandible mimicking a compound odontoma and a brief review of the related literature. PMID- 15292875 TI - Hepatocellular carcinoma metastatic to the mandible: a case involving severe hemorrhage. AB - Hepatocellular carcinoma (HCC) is quite uncommon in Europe and USA, although in this last country more than 13,000 new cases are diagnosed every year. Mandibular metastases are unusual, with only 50 cases documented. In this article, we present a 54 year-old male patient with a metastasis of HCC in right mandibular body, one year after treating the primary tumor. Curettage and biopsy of the lesion was performed, followed by a profuse hemorrhage. The patient expired six months after the biopsy, with multiple metastases. We describe the different paths for hematogenic dissemination of the process. PMID- 15292876 TI - Aggressive glomus tumor of the tongue: report of a case. AB - A glomus tumor or glomangioma is a benign neoplasm originating from the modified smooth muscle cells of the vascular glomus Glomangiomas of the head and/or neck are extremely rare, with an incidence of 0.6%. We present an observational case report of a recurrent glomic tumor originally located in the mobile tongue, with an aggressive clinical course that complicated the therapeutic options of the case. PMID- 15292877 TI - Applications of exfoliative cytology in the diagnosis of oral cancer. AB - Exfoliative cytology is a simple non-aggressive technique that is well accepted by the patient, and that is therefore an attractive option for the early diagnosis of oral cancer, including epithelial atypias and especially squamous cell carcinoma. However, traditional exfoliative cytology methods show low sensitivity (i.e. a high proportion of false negatives) in the diagnosis of these pathologies. This low sensitivity is attributable to various factors, including inadequate sampling, procedural errors, and the need for subjective interpretation of the findings. More recently, the continuing development of automated cytomorphometric methods, DNA content determination, tumour marker detection, and diverse molecular-level analyses has contributed to renewed interest in exfoliative cytology procedures for the diagnosis of oral cancer. The present study briefly reviews developments in these areas. PMID- 15292878 TI - Granular cell tumour. PMID- 15292880 TI - Cerebral palsy: A reconceptualization of the spectrum. AB - Approximately 50 years ago, interest in cerebral palsy increased, and the current definitions and classification were developed. The interplay among the dimensions of significant impairment, nonprogressive lesions, and persistence defines a group of children who were of interest to the researchers who developed the definition. Cerebral palsy as a definition does not attend to the broader issues of neurodevelopmental dysfunction. It isolates a portion of the spectrum of motor dysfunction and creates a category whose bounds are defined by a range of motor capability. The classifications of cerebral palsy that require revision are discussed. Some classifications should be discarded. Others should be brought in line with current knowledge and approaches. Still others should be modified to encompass the broader views of function and therapy that reflect the current expectations for persons with disabilities. PMID- 15292881 TI - Strategies for the early diagnosis of cerebral palsy. AB - Abstract Strategies for the early detection and diagnosis of cerebral palsy include multiple measures of the underlying brain abnormalities and their neurodevelopmental consequences. These measures can be grouped into the categories of pathogenesis, impairment, and functional limitation. Neuroimaging techniques are the most predictive measures of pathogenesis of cerebral palsy in both the preterm and term infant. Measures of neurological impairment focusing on muscle tone, reflexes, and other features of the neurological examination are poorly predictive in the first months of life. Detection of functional limitations manifested by motor developmental delay is sensitive and specific for later cerebral palsy, but not until well into the second 6 months of life. Abnormal spontaneous general movements in the infant 16 to 20 weeks postterm and earlier reflect functional limitations in the first months of life and have been shown to predict later cerebral palsy. Recognition of abnormal spontaneous general movements may improve early detection and diagnosis of cerebral palsy if these techniques can be successfully incorporated into organized follow-up programs and developmental surveillance. PMID- 15292882 TI - General movements: A window for early identification of children at high risk for developmental disorders. AB - Detection of children with a developmental disorder, such as cerebral palsy, at an early age is notoriously difficult. Recently, a new form of neuromotor assessment of young infants was developed, based on the assessment of the quality of general movements (GMs). GMs are movements of the fetus and young infant in which all parts of the body participate. The technique of GM assessment is presented and the features of normal, mildly abnormal, and definitely abnormal GMs discussed. Essential to GM assessment is the Gestalt evaluation of movement complexity and variation. The quality of GMs at 2 to 4 months postterm (so-called fidgety GM age) has been found to have the highest predictive value. The presence of definitely abnormal GMs at this age--that is, GMs devoid of complexity and variation--puts a child at very high risk for cerebral palsy. This implies that definitely abnormal GMs at fidgety age are an indication for early physiotherapeutic intervention. PMID- 15292883 TI - Neuroimaging in cerebral palsy. AB - Parents and clinicians concerned about high-risk infants and children with motor delay or cerebral palsy seek information on cause, treatment, prognosis, and recurrence risk. Used in combination with history and examination, neuroimaging studies can improve diagnosis and management. In premature infants, cranial ultrasound is a reliable, noninvasive diagnostic modality. Nuclear magnetic resonance techniques including magnetic resonance imaging and diffusion weighted imaging can be used effectively in neonatal encephalopathies. In children with motor delay and cerebral palsy syndromes including spastic diplegia, quadriplegia, hemiplegia, and extrapyramidal movement disorders, conventional magnetic resonance imaging has become an important determinant of diagnosis and management. The aim of this article is to help clinicians select and interpret imaging studies of benefit in clinical care. PMID- 15292884 TI - Feeding method and health outcomes of children with cerebral palsy. AB - Disorders of feeding and swallowing are common in children with cerebral palsy. Feeding and swallowing disorders have significant implications for development, growth and nutrition, respiratory health, gastrointestinal function, parent-child interaction, and overall family life. Assessments need to be comprehensive in scope and centered around the medical home. Oral feeding interventions for children with cerebral palsy may be effective in promoting oral motor function, but have not been shown to be effective in promoting feeding efficiency or weight gain. Feeding gastrostomy tubes are a reasonable alternative for children with severe feeding and swallowing problems who have had poor weight gain. PMID- 15292885 TI - The emerging role of therapeutic botulinum toxin in the treatment of cerebral palsy. AB - Therapeutic injections of botulinum toxin have been used for the past decade to decrease muscle tone in children with cerebral palsy. Although this unique intervention has been shown to provide safe localized spasticity reduction, functional benefits have been more difficult to demonstrate. Appropriate selection of patients and clearly defined goals are key factors in the success of a treatment program. The therapeutic approach to treating children with cerebral palsy should include a variety of complementary interventions that address the effect of abnormal muscle tone on the abilities of these children as they grow and develop. PMID- 15292886 TI - Health and social outcomes of children with cerebral palsy. AB - Health and social outcomes in children who have cerebral palsy (CP) depend on several factors, including the severity of the CP, medical interventions, and the child's environment. One of the hallmarks of cerebral palsy is its variability. Several formal methods of classifying persons with CP, such as the Gross Motor Function Classification System, have been developed to standardize that variability. Children and adults with CP must contend with increased mortality as well as many secondary conditions that can affect activities and participation in society. Predicting outcomes for children with CP, establishing realistic goals, and determining the best interventions to improve functioning are facilitated by applying research results to individual decisions. Many more such studies are needed--especially those performed with large representative samples. In addition, the childhood factors that affect outcomes in adults with CP still need to be elucidated. PMID- 15292887 TI - The treatment of cerebral palsy: What we know, what we don't know. AB - The treatment of cerebral palsy is directed at repair of the injured brain and at the management of the impairments and disabilities resulting from developmental brain injury. Currently, there are no clinically meaningful interventions that can successfully repair existing damage to the brain areas that control muscle coordination and movement. However, several interventions are available to diminish the degree of impairment (eg, muscle spasticity) and to increase participation in activities of daily living. Data on treatment compatible with evidence-based medicine are now being collected. PMID- 15292889 TI - The effect of moving to a new hospital facility on the prevalence of methicillin resistant Staphylococcus aureus. AB - BACKGROUND: The influence of hospital design on nosocomial transmission of methicillin-resistant Staphylococcus aureus (MRSA) is unknown. Our hospital's relocation to a new building with radically different ward design allowed us to study this question. Our old hospital facility had open bay wards and intensive care units, and few poorly located sinks for handwashing (bed:sink ratio 4:1). Our new hospital facility had optimized hand-washing geography and distribution of ward beds into mostly single or double rooms (bed:sink ratio 1.3:1). METHODS: We compared the prevalence of MRSA in the 2 institutions by obtaining nasal swabs from all patients on 8 selected wards and intensive care units at 2 time points both before and after the move. In addition, passive surveillance rates of MRSA for all hospitalized patients for 2 years both before and after the move were compared. Hand hygiene practices, although unrelated to the study periods, were directly observed. RESULTS: Eight of 123 patients cultured before the move were MRSA positive, compared to 5 of 138 patients cultured after the move (P=NS). MRSA prevalence determined by passive surveillance of all hospitalized patients before and after the move was also unchanged. An insignificant increase in the frequency of hand-hygiene performance following the move (20% to 23%) was observed. CONCLUSION: Radical facility design changes, which would be permissive of optimal infection control practices, were not sufficient, by themselves, to reduce the nosocomial spread of MRSA in our institution. PMID- 15292888 TI - Assessing the status of infection control programs in small rural hospitals in the western United States. AB - BACKGROUND: Organized infection control (IC) interventions have been successful in reducing the acquisition of hospital-associated infections. Rural community hospitals, although contributing significantly to the US health care system, have rarely been assessed regarding the nature and quality of their IC programs. METHODS: A sample of 77 small rural hospitals in Idaho, Nevada, Utah, and eastern Washington completed a written survey in 2000 regarding IC staffing, infrastructure support, surveillance of nosocomial infections, and IC policies and practices. RESULTS: Almost all hospitals (65 of 67, 97%) had one infection control practitioner (ICP), and 29 of 61 hospitals (47.5%) reported a designated physician with IC oversight. Most ICPs (62 of 64, 96.9%) were also employed for other activities outside of IC. The median number of ICP hours per week for IC activities was 10 (1-40), equating to a median of 1.56 (0.30-21.9) full-time ICPs per 250 hospital beds. Most hospitals performed total house surveillance for nosocomial infections (66 of 73, 90.4%) utilizing Centers for Disease Control and Prevention (CDC) definitions (69 of 74, 93.2%). Most also monitored employee bloodborne exposures (69 of 73, 94.5%). All hospitals had a written bloodborne pathogen exposure plan and isolation policies. CDC guidelines were typically followed when developing IC policies. Access to medical literature and online resources appeared to be limited for many ICPs. CONCLUSIONS: Most rural hospitals surveyed have expended reasonable resources to develop IC programs that are patterned after those seen in larger hospitals and conform to recommendations of consensus expert panels. Given these hospitals' small patient census, short length of stay, and low infection rates, further studies are needed to evaluate necessary components of effective IC programs in these settings that efficiently utilize limited resources without compromising patient care. PMID- 15292890 TI - Failure rates in nonlatex surgical gloves. AB - BACKGROUND: The purpose of this study was to compare the frequency of glove defects for nonlatex surgical gloves while surgeons performed routine surgery and to evaluate surgeons' satisfaction with nonlatex sterile gloves. METHODS: Two brands of latex gloves and 6 brands of nonlatex gloves were tested. Gloves were collected at the end of each surgical procedure and tested for visual defects and barrier integrity using an automated calibrated water test machine consistent with FDA's recommended standards. A total of 6386 gloves used by 101 surgeons and 164 residents representing 15 surgical services were included in the analysis. RESULTS: Higher after-use defect rates occurred in nonlatex surgical gloves than in latex gloves. Higher times of use were related to higher defect rates for some surgical specialties, and both surgeons and residents were less satisfied with nonlatex surgical gloves. CONCLUSION: Intact latex and nonlatex surgical gloves provide adequate barrier protection. Nonlatex surgical gloves have higher failure rates and lower user satisfaction than latex gloves do. Both nonlatex and latex gloves should be changed after 2 to 3 hours of use because the barrier of either type of glove becomes compromised with extended use. PMID- 15292891 TI - Is there an association between shigellosis incidence and socioeconomic status in metropolitan Haifa? AB - BACKGROUND: Shigellosis incidence rates in Israel have declined continuously over the past 50 years, but they remain 20 times greater than those in the United States. Socioeconomic factors may influence shigellosis morbidity, but this may be difficult to demonstrate in the absence of data for individual patients and when using composite rates for large geographic areas. Use of census tract data for small, relatively homogeneous geographic areas may lessen the effects of the "ecological fallacy." The present study analyzes the effect of socioeconomic status (SES) on shigellosis morbidity in the Haifa metropolitan region. METHODS: The study population consisted of the 7 cities in the Haifa subdistrict that constitute the greater metropolitan region. Cases of shigellosis reported during the years 2000 and 2001 were mapped, and age-standardized rates were calculated for the census tract areas. The incidence rates were then compared with the SES category of the census tract using the Kruskal-Wallis test. RESULTS: No association was found between incidence rates of shigellosis and SES category of the census tract areas in the Haifa metropolitan area for the years 2000 and 2001 (Kruskal-Wallis chi(2)=0.440; P=.803). CONCLUSION: We found no association between shigellosis morbidity and socioeconomic status. This finding is probably real and not the result of reporting bias. Analysis of morbidity using small geographical units such as census tracts is more accurate than analysis using large geographical areas such as cities. PMID- 15292892 TI - Interhospital transfer of pan-resistant Acinetobacter strains in Johannesburg, South Africa. AB - BACKGROUND: Acinetobacter species infections are increasingly found to cause nosocomial infections, particularly in intensive care units. These pathogens are difficult to eliminate from the hospital environment, and the emergence of multiple-drug-resistant strains complicates patient treatment. In this retrospective study, several strains were analyzed to study the possible spread of pan-resistant strains. METHODS: Macrorestriction analysis was performed on isolates collected in July 2001 from Johannesburg Hospital and strains collected from a number of hospitals in Johannesburg a year later. RESULTS: A strain endemic to Johannesburg Hospital that was cefepime and ceftazidime sensitive in 2001 developed resistance to these antibiotics within 1 year. This and other resistant strains were found to have spread among academic and private hospitals in the area by July 2002. CONCLUSIONS: The development of resistance is believed to be a response to antibiotic pressure and the spread of resistant strains a result of health care worker and/or patient transfer among hospitals. This snapshot epidemiologic study highlights the need to institute stricter infection control measures to limit the spread of organisms such as Acinetobacter among hospitals. PMID- 15292893 TI - Infection control and its application to the administration of intravenous medications during gastrointestinal endoscopy. AB - Several infection control practices and procedures crucial to the prevention of disease transmission in the health care setting are reviewed and discussed. Emphasis is placed on the importance of infection control to gastrointestinal endoscopy. Recommendations that minimize the risk of nosocomial infection during the preparation, handling, and administration of intravenous medications, particularly propofol, are provided. These recommendations include the labeling of predrawn syringes; use of sterile single-use syringes, needles, and administration sets for each patient; and, whenever feasible, administration of intravenous medications promptly after opening their prefilled syringes or after opening their ampoules or vials and filling the sterile syringes. PMID- 15292894 TI - Toys in a pediatric hospital: are they a bacterial source? AB - BACKGROUND: In children's hospitals, children are commonly provided with toys. Measures to guarantee the safety of these toys are usually not taken. This study was conducted to determine whether toys were contaminated with potentially pathogenic bacteria when they arrived in the hospital, and whether they were contaminated in the hospital. METHODS: The study was conducted during a 3-month period. Children who were hospitalized for at least 3 days were chosen as study subjects. Once these children were identified, cultures from their toys were obtained within the first 48 hours of admission. After this first culture, toys were cleaned with 4% chlorhexidine and water and were immediately re-cultured. Following cultures were collected on days 5 to 7, 10 to 15, and every week thereafter until the owner-patient was discharged. Specimens were collected in a standardized manner with moistened swabs and placed in transport media. They were later inoculated onto trypticase soy agar with 5% sheep blood and brain heart infusion agar, incubated at 37 degrees C for 48 hours and examined for colony growth at 24 to 48 hours. RESULTS: Seventy children's toys were included in this study. Patients' median age was 26 months (range: 1 day to 9 years). Respiratory infections (43%) and diarrhea (26%) were the most common causes of hospitalization. Fifty-three (76%) toys were made of plastic, 8 (11%) metallic, and 9 (13%) other materials. Twenty-nine (41%) were brought from home, 38 (55%) were purchased from roving vendors, and 3 (4%) were purchased from toy stores. All first cultures were positive for at least 1 pathogenic microorganism: 55 (78%) coagulase-negative Staphylococcus (CNS); 26 (37%) Bacillus spp; 13 (18%) Staphylococcus aureus; 8 (11%) alpha-hemolytic Streptococcus; 5 (9%), Pseudomonas spp; 2 (3%) Stenotrophomonas maltophilia, and 6 (11%) other gram-negative organisms. After toys were cleaned, subsequent cultures showed significant decreases in bacterial growth rates (P <.05). Because some patients were discharged, additional cultures were obtained for only 31 toys. CONCLUSIONS: Toys entering a hospital can be contaminated with potentially dangerous bacteria and may provide unnecessary risks for nosocomial infection. Effective measures must be implemented to prevent the spread of infections via toys. PMID- 15292895 TI - Prevention of catheter-related bloodstream infection in critically ill patients using a disinfectable, needle-free connector: a randomized controlled trial. AB - OBJECTIVE: The aim of this study was to assess the efficacy of a disinfectable, needle-free connector in the prophylaxis of catheter-related bloodstream infection. METHODS: A randomized controlled trial was performed in a polyvalent intensive care unit. Patients who needed multilumen central venous catheters were randomly assigned to a study or a control group. All catheters were inserted and manipulated according to the Centers for Disease Control and Prevention (CDC) recommendations. Study group patients were equipped with catheters with disinfectable, needle-free connectors whereas control group patients were equipped with catheters with 3-way stopcocks. Two peripheral blood cultures and a semiquantitative culture of the catheter tip were performed on removal of the catheter. RESULTS: The study included 243 patients, with a total of 278 central venous catheters. The catheters' mean insertion duration was 9.9 days. Both groups were comparable regarding patient and catheter characteristics. Incidence rate of catheter-related bloodstream infection was 0.7 per 1000 days of catheter use in the study group, compared with 5.0 per 1000 days of catheter use in the control group (P=.03). CONCLUSIONS: To add a disinfectable, needle-free connector to the CDC recommendations reduces the incidence of catheter-related bloodstream infection in critically ill patients with central venous catheters. PMID- 15292896 TI - Experimental study on the safety of a new connecting device. AB - BACKGROUND: The tested device is a new connecting tool for infusion systems that has been designed to replace conventional single-use stopcocks. Because outbreaks of bloodstream infections have been observed during the use of similar connectors in the United States, we examined the microbiological safety of the connecting device after artificial contamination in the laboratory setting and during routine clinical use. METHODS: In the first part of the study, the new device was tested in 3 types of in vitro experiments. In the second part of the study, surgical intensive care patients had their entry ports capped with novel devices (n=27) or with conventional stopcocks (n=32), and samples of infusion fluids and swabs from entry ports were taken after completion of infusion periods. RESULTS: The new device did not perpetuate bacterial contaminations in spite of high artificial inocula in the in vitro experiments. Microbial contamination rates after 96 hours of infusion therapy for the novel connecting tool versus conventional stopcock groups were as follows: swabs from 3-way ports, 6/129 versus 1/111; rest fluid from infusion lines, 0/20 versus 1/22; rest fluid from infusion bottles, 2/196 versus 2/208; rest fluid from perfusor syringes, 7/180 versus 6/142 (all differences not significant). CONCLUSION: The novel connecting device was microbiologically safe and did not increase microbial contamination rates of intravenous infusion systems. PMID- 15292897 TI - Reducing percutaneous injuries at an academic health center: a 5-year review. AB - BACKGROUND: The University of Connecticut Health Center Employee Health Service collected and used National Surveillance System for Hospital Health Care Workers (NaSH) data to (1) improve surveillance of health care worker blood and body fluid exposures (BBFEs) and (2) target specific interventions for higher-risk groups (nursing staff, medical and dental students, and residents). METHODS: All 870 BBFE incidents were abstracted from the NaSH database from the 1997 through 2002 academic years. Incidence rates per 100 full-time-equivalent workers were determined for each targeted occupation group with 95% confidence intervals. RESULTS: The number of percutaneous injuries declined among medical/dental students and nursing staff, and to a lesser degree for residents. The incidence rates decreased from 7.9% in 2000 to 2001 to 2.6% in 2001 to 2002 for students and from 9.2% in 1997 to 1998 to 2.7% in 2001 to 2002 for nursing staff. CONCLUSIONS: Data from a surveillance database provided guidance for administrative, educational, and engineering control interventions. Active surveillance and periodic review of interventions are important aspects to reduce BBFEs in targeted high-risk occupational groups, especially when the workforce has a high turnover, as is typical in academic health centers. PMID- 15292898 TI - Practical limitations of disinfection of body fluid spills with 10,000 ppm sodium hypochlorite (NaOCl). AB - The purpose of this study was to monitor disinfection with 10,000 ppm sodium hypochlorite for decontamination of common hospital spills. Simulated spills deliberately contaminated with 10(8) bacterial challenges were used for the study. Results showed greater than 5 log reduction in the challenge bacteria for all spills (serum, pus, sputum, csf, ascitic fluid, urine, and stool) except blood. Disinfection was satisfactory for blood contaminated with gram-negative bacteria, but not for staphylococci. As a practical procedure, surfaces contaminated from gross spillage of human body fluids should first be contained with absorbent materials, then disinfected with hypochlorite. PMID- 15292899 TI - Medical cases who would benefit from treatment on a spinal injury unit. PMID- 15292900 TI - Do spinal cord injury patients always get the best treatment for neuropathic bladder after discharge from regional spinal injuries centre? AB - OBJECTIVE: To draw attention to inadequate care received by some spinal cord injury patients after discharge from the regional spinal injury center. SETTING: Regional Spinal Injuries Centre, Southport, UK. METHODS: Presence of the urethral stricture was not recognised in a 69-year-old male with T-3 paraplegia, who attended a health-care facility with a urinary infection. A Foley catheter was inserted into the urethra only half-way and the catheter balloon was then inflated in the urethra distal to the stricture. In a 68-year-old male with T-8 paraplegia, a long-term indwelling catheter was eroding the urethra and he developed a severe degree of hypospadias while being managed in the community. A 49-year-old male with C-4 tetraplegia developed recurrent urine infections. He received several courses of antibiotics, which were prescribed by community health professionals. But he continued to be unwell. Subsequently, the patient was admitted to a district general hospital, where he was diagnosed to have mild chest infection and was about to be sent home. However, his wife was not happy, and then ultrasound of abdomen was taken, which revealed pyonephrosis. He was then transferred to a spinal unit. RESULTS: : These patients were not seen promptly in a regional spinal injury centre when they developed medical problems. The complications, which ensued, might have been prevented if expert medical treatment had been provided without delay. CONCLUSION: In order to meet the needs of a growing population of persons living in the community with spinal cord injury, more beds are required in spinal units. Provision of day surgery wards within spinal units, out-reach clinics and home visits by spinal cord clinicians may reduce the demand for admission in a spinal unit. Education of community health professionals on delayed complications of spinal cord injury, and good communication between spinal cord clinicians, patients, carers, and community health professionals by telephone, e-mail or conventional postal system are likely to improve the care of spinal cord injury patients after discharge from spinal injury centres. Spinal cord clinicians should adopt a patient-centred care instead of the traditional, paternalistic, doctor-centred care. PMID- 15292901 TI - Iraqi National Spinal Cord Injuries Centre (INSCIC) twice destroyed since last war but we refunctioned it again. AB - STUDY DESIGN: Description of spinal cord injuries in a war situation. SETTING: Baghdad, Iraq. PMID- 15292903 TI - Individual differences in cocaine- and amphetamine-induced activation of male Sprague-Dawley rats: contribution of the dopamine transporter. AB - Previously we found that outbred male Sprague-Dawley rats can be classified as either low or high cocaine responders (LCRs or HCRs, respectively), based on their open-field locomotor response to acute cocaine (COC; 10 mg/kg, i.p.). Here, we extended this analysis to amphetamine (AMPH; 0.5, 1, and 5 mg/kg, i.p.) and found that the individual differences in behavioral activation were not as pronounced as with COC. This was confirmed with observational analysis of behaviors. Differences in drug-induced activation could involve differential dopamine transporter (DAT) function/trafficking. To address this possibility, we measured [3H]DA uptake into dorsal striatal synaptosomes prepared from rats injected 30 min earlier with saline, COC, or AMPH to determine DAT activity, and radioligand binding to determine the total number of DATs. Striatal [3H]DA uptake in COC-treated HCRs was significantly higher than in LCRs. Furthermore, regardless of LCR/HCR classification, uptake in individual COC-treated rats was significantly correlated with their locomotor behavior in the 30 min after drug administration. In contrast, AMPH-treated rats did not differ in uptake, nor were uptake and locomotor activity correlated. DAT number did not differ between LCRs or HCRs, or between AMPH-treated rats. In addition, when individual differences in COC-induced behavior were no longer detected in LCRs and HCRs 1 week after initial classification, uptake was also similar. Together, these results suggest that a difference in expression of functional DATs on the cell surface contributes to the individual differences observed in COC-induced, but not AMPH induced, behavioral activation of rats. PMID- 15292904 TI - Serotonin and dopamine transporter availabilities correlate with the loudness dependence of auditory evoked potentials in patients with obsessive-compulsive disorder. AB - Brain monoaminergic function is involved in the pathophysiology of psychiatric disorders. The loudness dependence (LD) of the N1/P2 component of auditory evoked potentials has been proposed as a noninvasive indicator of central serotonergic function, whereas single photon emission computed tomography (SPECT) and [123I]beta-CIT can be used to visualize both serotonin (SERT) and dopamine transporters (DAT). The aim of the study was to correlate LD and SPECT measures in patients with obsessive-compulsive disorder, a condition with evidence for a serotonergic dysfunction. A total of 10 subjects received both neurophysiological and imaging investigations. Evoked potentials were recorded following the application of acoustic stimuli with increasing intensities. The LD of the relevant subcomponents (tangential dipoles) was investigated using dipole source analysis. SPECT was performed 20-24 h after injection of a mean 140 MBq [123I]beta-CIT. As a measure of brain SERT and DAT availabilities, a ratio of specific to nonspecific [123I]beta-CIT binding for the midbrain . pons region (SERT) and the striatum (DAT) was used. The LD of the right tangential dipole correlated significantly with both SERT and DAT availabilities (Pearson's correlations: rho = 0.69, p < 0.05, and rho = 0.80, p < 0.01, respectively). The correlations remained significant after controlling for the effects of age, gender, and severity of clinical symptoms. Associations between LD and both SERT and DAT availabilities further validate the use of neurophysiological approaches as noninvasive indirect measures of neurochemical brain function and point at a hypothesized interconnection of central monoaminergic systems. PMID- 15292905 TI - Nicotine-associated cues maintain nicotine-seeking behavior in rats several weeks after nicotine withdrawal: reversal by the cannabinoid (CB1) receptor antagonist, rimonabant (SR141716). AB - Conditioned stimuli are important for nicotine dependence and may trigger craving and relapse after prolonged nicotine abstinence. However, little is known about the pharmacology of this process. Among the systems that have been shown to play a role in drug-seeking behavior is the endocannabinoid transmission. Therefore, the present study examined the resistance to extinction of drug-seeking behavior elicited by nicotine-associated environmental stimuli and the effects of the selective CB1 cannabinoid antagonist rimonabant (SR141716) on the reinforcing effects of nicotine-related stimuli. Rats were trained to self-administer nicotine (0.03 mg/kg/injection, i.v.) under conditions in which responding was reinforced jointly by response-contingent nicotine injections and stimuli (light and tone). After self-administration acquisition, nicotine was withdrawn and lever pressing was only reinforced by contingent presentation of the audiovisual stimuli. Under such a condition, responding persisted for 3 months, following which nonpresentation of the cues produced a progressive extinction of responding. As expected, rats trained to lever-press for saline injections paired with the audiovisual stimuli did not acquire the self-administration. These findings indicate that the cues required learned association with nicotine to acquire reinforcing properties and to function as conditioned reinforcers. When administered 1 month following nicotine withdrawal, rimonabant (1 mg/kg, i.p.) decreased conditioned behavior. These results showing the persistence of a nicotine-conditioned behavior are congruent with the role of nicotine-related environmental stimuli in nicotine craving in abstinent smokers. Rimonabant, which has been shown previously to reduce nicotine self-administration, may be effective not only as an aid for smoking cessation but also in the maintenance of abstinence. PMID- 15292906 TI - A review and evaluation of intraurban air pollution exposure models. AB - The development of models to assess air pollution exposures within cities for assignment to subjects in health studies has been identified as a priority area for future research. This paper reviews models for assessing intraurban exposure under six classes, including: (i) proximity-based assessments, (ii) statistical interpolation, (iii) land use regression models, (iv) line dispersion models, (v) integrated emission-meteorological models, and (vi) hybrid models combining personal or household exposure monitoring with one of the preceding methods. We enrich this review of the modelling procedures and results with applied examples from Hamilton, Canada. In addition, we qualitatively evaluate the models based on key criteria important to health effects assessment research. Hybrid models appear well suited to overcoming the problem of achieving population representative samples while understanding the role of exposure variation at the individual level. Remote sensing and activity-space analysis will complement refinements in pre-existing methods, and with expected advances, the field of exposure assessment may help to reduce scientific uncertainties that now impede policy intervention aimed at protecting public health. PMID- 15292907 TI - Communicating exposure and health effects results to study subjects, the community and the public: strategies and challenges. AB - The Mickey Leland National Urban Air Toxics Research Center sponsored a Symposium in August 2002 that focused on the communication of health effects results from community studies involving exposure to hazardous substances in the environment. Some of the audiences identified for presentation of study results were the study subjects, the community, and the general public. Principles and approaches to communicating findings were discussed, as were the challenges that may confront researchers in developing and implementing a communication plan. The Symposium included four sessions. The first was an overview session where Timothy McDaniels (University of British Columbia) described risk communication as a decision aiding process. In the second session, case studies were presented by Timothy Buckley (Johns Hopkins University), Jane Hoppin (National Institute of Environmental Health Sciences), and Anne-Marie Nicol (University of British Columbia). Approaches and strategies used by different stakeholders to communicate study results was the topic for a panel discussion at the third session. Panelists included: James Collins (The Dow Chemical Company), Mary White (Agency for Toxic Substances and Disease Registry), Richard Clapp (Boston University), Valerie Zartarian (Environmental Protection Agency), Pamela Williams (Chemrisk), and Tina Bahadori (American Chemistry Council). The final session was a summary presentation on lessons learned given by Rebecca Parkin of George Washington University, in which she synthesized the preceding presentations and formulated guidelines for effective risk communication in community research studies. PMID- 15292908 TI - Children's mouthing and food-handling behavior in an agricultural community on the US/Mexico border. AB - Children's mouthing and food-handling activities were measured during a study of nondietary ingestion of pesticides in a south Texas community. Mouthing data on 52 children, ranging in age from 7 to 53 months, were collected using questionnaires and videotaping. Data on children's play and hand-washing habits were also collected. Children were grouped into four age categories: infants (7 12 months), 1-year-olds (13-24 months), 2-year-olds (25-36 months) and preschoolers (37-53 months). The frequency and type of events prompting hand washing did not vary by age category except for hand washing after using the bathroom; this increased with increasing age category. Reported contact with grass and dirt also increased with increasing age category. The median hourly hand-to-mouth frequency for the four age groups ranged from 9.9 to 19.4, with 2 year-olds having the lowest frequency and preschoolers having the highest. The median hourly object to mouth frequency ranged from 5.5 to 18.1 across the four age categories; the frequency decreased as age increased (adjusted R(2)=0.179; P=0.003). The median hourly hand-to-food frequency for the four age groups ranged from 10.0 to 16.1, with the highest frequency being observed in the 1-year-olds. Hand-to-mouth frequency was associated with food contact frequency, particularly for children over 12 months of age (adjusted R(2)=0.291; P=0.002). The frequency and duration of hand-to-mouth, object-to-mouth and food-handling behaviors were all greater indoors than outdoors. Infants were more likely to remain indoors than children in other age groups. The time children spent playing on the floor decreased with increasing age (adjusted R(2)=0.096; P=0.031). Parental assessment was correlated with hand-to-mouth activity but not with object-to-mouth activity. The highest combined (hand and object) mouthing rates were observed among infants, suggesting that this age group has the greatest potential for exposure to environmental toxins. PMID- 15292909 TI - Differential chromosome pairing affinities at meiosis in polyploid sugarcane revealed by molecular markers. AB - Chromosome pairing at meiosis is an essential feature in cell biology, which determines trait inheritance and species evolution. Complex polyploids may display diverse pairing affinities and offer favorable situations for studying meiosis. The genus Saccharum encompasses diverse forms of polyploids with predominantly bivalent pairing. We have focused on a modern cultivar of sugarcane, R570, and taken advantage of a particular single copy probe (BNL 12.06) revealing 11 alleles by restriction fragment length polymorphism (RFLP). As for other cultivars, R570 is highly polyploid (2n=ca. 115) and indirectly derived from interspecific hybridization between Saccharum officinarum (2n=80, x=10) and S. spontaneum (2n=40-128, x=8). Here we determined the doses of the various BNL12.06 RFLP alleles among 282 progeny of R570 and estimated the mutual pairing frequencies among the corresponding homo- or homoeologous chromosomes using a maximum likelihood method. The result is an atypical picture, with pairing frequencies ranging from 0 to 40% and differential affinities leading to the identification of several chromosome subsets. This example illustrates the unsystematic meiotic behavior in a complex polyploid. It highlights a continuous range of pairing affinities between chromosomes and pinpoints a strong role of individual chromosome features, partly related to their ancestral origin, in the determination of these affinities. PMID- 15292910 TI - Patterns of pollen dispersal in a small population of Pinus sylvestris L. revealed by total-exclusion paternity analysis. AB - Patterns of pollen dispersal were investigated in a small, isolated, relict population of Pinus sylvestris L., consisting of 36 trees. A total-exclusion battery comprising four chloroplast and two nuclear microsatellites (theoretical paternity exclusion probability EP=0.996) was used to assign paternity to 813 seeds, collected from 34 trees in the stand. Long-distance pollen immigration accounted for 4.3% of observed matings. Self-fertilization rate was very high (0.25), compared with typical values in more widespread populations of the species. The average effective pollen dispersal distance within the stand was 48 m (or 83 m excluding selfs). Half of effective pollen was dispersed within 11 m, and 7% beyond 200 m. A strong correlation was found between the distance to the closest tree and the mean mating-distance calculated for single-tree progenies. The effective pollen dispersal distribution showed a leptokurtic shape, with a large and significant departure from that expected under uniform dispersal. A maximum-likelihood procedure was used to fit an individual pollen dispersal distance probability density function (dispersal kernel). The estimated kernel indicated fairly leptokurtic dispersal (shape parameter b=0.67), with an average pollen dispersal distance of 135 m, and 50% of pollen dispersed beyond 30 m. A marked directionality pattern of pollen dispersal was found, mainly caused by the uneven distribution of trees, coupled with restricted dispersal and unequal male success. Overall, results show that the number and distribution of potential pollen donors in small populations may strongly influence the patterns of effective pollen dispersal. PMID- 15292911 TI - Microsatellite null alleles in parentage analysis. AB - Highly polymorphic microsatellite markers are widely employed in population genetic analyses (eg, of biological parentage and mating systems), but one potential drawback is the presence of null alleles that fail to amplify to detected levels in the PCR assays. Here we examine 233 published articles in which authors reported the suspected presence of one or more microsatellite null alleles, and we review how these purported nulls were detected and handled in the data analyses. We also employ computer simulations and analytical treatments to determine how microsatellite null alleles might impact molecular parentage analyses. The results indicate that whereas null alleles in frequencies typically reported in the literature introduce rather inconsequential biases on average exclusion probabilities, they can introduce substantial errors into empirical assessments of specific mating events by leading to high frequencies of false parentage exclusions. PMID- 15292912 TI - Mating system, sex ratio, and persistence of females in the gynodioecious shrub Daphne laureola L. (Thymelaeaceae). AB - Although in gynodioecious populations male steriles require a fecundity advantage to compensate for their gametic disadvantage, southern Spanish populations of the long-lived shrub Daphne laureola do not show any fecundity advantage over hermaphrodites in terms of seed production and early seedling establishment. By using allozyme markers, we assess the mating system of this species in five populations differing in sex ratio, and infer levels of inbreeding depression over the whole life cycle by comparing the inbreeding coefficients at the seed and adult plant stages. Extremely low outcrossing rates (0.001/=5 standard deviations above the mean optical density obtained for studied virgins at enrollment (n=573). Seroconnversion (n=409), persistence (n=675), and clearance (n=541) were defined based on enrollment and follow-up serology measurements. Age specific distributions revealed that HPV-16 seroconversion was highest among 18- to 24-year-old women, steadily declining with age; HPV-16 seropersistence was lowest in women 65+ years. In age-adjusted multivariate logistic regression models, a 10-fold risk increase for HPV-16 seroconversion was associated with HPV 16 DNA detection at enrollment and follow-up; two-fold risk of seroconversion to HPV-16 was associated with increased numbers of lifetime and recent sexual partners and smoking status. Determinants of HPV-16 seropersistence included a 1.5-fold risk increase associated with having one sexual partner during follow up, former oral contraceptive use, and a 3-fold risk increase associated with HPV 16 DNA detection at both enrollment and follow-up. Higher HPV-16 viral load at enrollment was associated with seroconversion, and higher antibody titres at enrollment were associated with seropersistence. PMID- 15292930 TI - Educational attainment among survivors of childhood cancer: a population-based cohort study in Denmark. AB - We identified 2384 patients in the Danish Cancer Register in whom cancer had been diagnosed in 1960-1996 before they reached the age of 20 and compared them with 53 143 sex- and age-matched controls identified from the Register of Population Statistics. Complete education records and demographic and socioeconomic information for the period 1980-2000 were obtained for both cohorts from Statistics Denmark. The rate ratio (RR) for educational attainment was estimated by discrete-time Cox regression analyses. An overall reduction in attaining basic education was found (RR, 0.90; 95% confidence interval, 0.83-0.96). Female survivors of central nervous system (CNS) tumours showed the largest educational deficit (RR, 0.55; 95% confidence interval, 0.37-0.82). Non-CNS tumour survivors attained education as controls at most levels. When the analyses were conditioned on completion of youth education, further educational attainment was not reduced for any group of survivors. These findings confirm that only survivors of CNS tumours in childhood experience significant educational deficits. The deficit was mainly seen among persons whose tumour was diagnosed before they reached the level of secondary education. PMID- 15292931 TI - Radiation-induced malignancies following radiotherapy for breast cancer. AB - With advances in diagnosis and treatment, breast cancer is becoming an increasingly survivable disease resulting in a large population of long-term survivors. Factors affecting the quality of life of such patients include the consequences of breast cancer treatment, which may have involved radiotherapy. In this study, we compare the incidence of second primary cancers in women who received breast radiotherapy with that in those who did not (non-radiotherapy). All women studied received surgery for their first breast cancer. Second cancers of the lung, colon, oesophagus and thyroid gland, malignant melanomas, myeloid leukaemias and second primary breast cancers were studied. Comparing radiotherapy and non-radiotherapy cohorts, elevated relative risks (RR) were observed for lung cancer at 10-14 years and 15 or more (15+) years after initial breast cancer diagnosis (RR 1.62, 95% confidence interval [CI] 1.05-2.54 and RR 1.49, 95% CI 1.05-2.14, respectively), and for myeloid leukaemia at 1-5 years (RR 2.99, 95% CI 1.13-9.33), for second breast cancer at 5-10 years (RR 1.34, 95% CI 1.10-1.63) and 15+ years (RR 1.26, 95% CI 1.00-1.59) and oesophageal cancer at 15+ years (RR 2.19, 95% CI 1.10-4.62). PMID- 15292932 TI - No topoisomerase I alteration in a neuroblastoma model with in vivo acquired resistance to irinotecan. AB - CPT-11 (irinotecan) is a DNA-topoisomerase I inhibitor with preclinical activity against neuroblastoma (NB) xenografts. The aim was to establish in vivo an NB xenograft resistant to CPT-11 in order to study the resistance mechanisms acquired in a therapeutic setting. IGR-NB8 is an immature NB xenograft with MYCN amplification and 1p deletion, which is sensitive to CPT-11. Athymic mice bearing advanced-stage subcutaneous tumours were treated with CPT-11 (27 mg kg(-1) day( 1) x 5) every 21 days (1 cycle) for a maximum of four cycles. After tumour regrowth, a new in vivo passage was performed and the CPT-11 treatment was repeated. After the third passage, a resistant xenograft was obtained (IGRNB8-R). The tumour growth delay (TGD) was reduced from 115 at passage 1 to 40 at passage 4 and no complete or partial regression was observed. After further exposure to the drug, up to 28 passages, the resistant xenograft was definitively established with a TGD from 17 at passage 28. Resistant tumours reverted to sensitive tumours after 15 passages without treatment. IGR-NB8-R remained sensitive to cyclophosphamide and cisplatin and cross-resistance was observed with the topoisomerase I inhibitor topotecan. No quantitative or qualitative topoisomerase I modifications were observed. The level of expression of multidrug resistance 1 (MDR1), MDR-associated protein 1 (MRP1) and, breast cancer resistance protein, three members of the ATP-binding cassette transporter family was not modified over passages. Our results suggest a novel resistance mechanism, probably not involving the mechanisms usually observed in vitro. PMID- 15292933 TI - The cumulative risk of lung cancer among current, ex- and never-smokers in European men. AB - Recent analyses based on UK data indicate that people who stop smoking, even well into middle age, avoid most of their subsequent risk of lung cancer. We investigated whether similar absolute risks of lung cancer in men are found in other European countries with different smoking patterns and at different stages of their lung cancer epidemic. Using data for men from a multicentre case-control study of lung cancer in the UK, Germany, Italy and Sweden, and including 6523 lung cancer cases and 9468 controls, we combined odds ratio estimates with estimates of national lung cancer incidence rates to calculate the cumulative risk of lung cancer among men by age 75. Lung cancer cumulative risks by age 75 among continuing smokers were similar for the UK, Germany and Italy at 15.7, 14.3 and 13.8% respectively, whereas the cumulative risk among Swedish male smokers was 6.6%. The proportion of the risk of lung cancer avoided by quitting smoking before the age of 40 was comparable between the four countries, at 80% in Italy and 91% in the UK, Germany and Sweden. Similarly, the proportion of the excess risk avoided by quitting before the age of 50 ranged from 57% in Italy to 69% in Germany. Our results support the important conclusion that for long-term smokers, giving up smoking in middle age avoids most of the subsequent risk of lung cancer, and that lung cancer mortality in European men over the next three decades will be determined by the extent to which current smokers can successfully quit smoking. PMID- 15292934 TI - Reassurance and the anxious cancer patient. AB - Many cancer patients are anxious even when disease is in remission. Anxiety about health, 'health anxiety', has distinct features, notably seeking medical reassurance about symptoms. Doctors may then communicate that these symptoms are not due to serious illness, a process known as 'reassurance'. However, reassurance may inadvertently perpetuate some patients' anxiety. We aimed to observe the relation between symptoms, anxiety and reassurance in consultations with cancer patients. A total of 95 outpatients, with breast or testicular cancers in remission, completed questionnaires measuring health anxiety at study entry, then general anxiety - before a consultation, immediately afterwards, 1 week later, and before their next consultation. We examined symptoms reported and reassurance by oncologists from audio recordings of consultations, and the outcome of subjects' anxiety. The results showed that substantial health anxiety was reported by one-third of the patients. Patients with higher levels of health anxiety reported more symptoms during consultations. Reassurance was ubiquitous, but not followed by an enduring improvement in anxiety. Certain forms of reassurance predicted increased anxiety over time, particularly for subjects who were most anxious. In conclusion, health anxiety can be a problem after cancer. Reassurance may not reduce patients' anxiety. Some reassurance was counterproductive for the most anxious patients. Oncologists may need to use reassurance as a procedure, balancing risk, and benefits, and patient selection and to manage cancer patients in remission. PMID- 15292935 TI - Phase I-II study of irinotecan (CPT-11) plus nedaplatin (254-S) with recombinant human granulocyte colony-stimulating factor support in patients with advanced or recurrent cervical cancer. AB - Combination chemotherapy with irinotecan (CPT-11) and platinum compounds is effective for treating cervical cancer. Nedaplatin (254-S) is a new cisplatin analogue that achieves a high response rate (53%) in patients with primary cervical cancer. We performed a phase I-II study of combination chemotherapy with CPT-11 plus 254-S for advanced or recurrent cervical cancer. The inclusion criteria were stage IV disease or recurrence. CPT-11 and 254-S were administered intravenously on day 1, while rhG-CSF (50 microg) was given on days 3-12. This regimen was repeated after 4 weeks. Dose escalation was carried out in tandem (CPT-11/254-S: 50/70, 50/80, and 60/80 mg m(-2)). A total of 27 patients (stage IV=seven, recurrence=20) were enrolled. The phase I study enrolled eight patients. At dose levels 1 and 2, no dose-limiting toxicities were observed. At dose level 3, the first two patients developed DLTs. The maximum tolerated dose of CPT-11 and 254-S was 60 and 80 mg m(-2), respectively, and the recommended doses were 50 and 80 mg m(-2). Grade 3/4 haematologic toxicity occurred in 67% in phase II study, but there were no grade 3 non-haematologic toxicities except for nausea or lethargy. In all 27 patients, there were two complete responses (7%) and 14 Partial responses (52%), for an overall response rate of 59% (95% confidence interval: 39-78%). Among the 12 responders with recurrent disease, the median time to progression and median survival were 161 days (range: 61-711 days) and 415 days (range: 74-801 days). This new regimen is promising for cervical cancer. PMID- 15292936 TI - Decreased rates of advanced breast cancer due to mammography screening in The Netherlands. AB - The effect of the implementation of the Dutch breast cancer screening programme during 1990-1997 on the incidence rates of breast cancer, particularly advanced breast cancer, was analysed according to stage at diagnosis in seven regions, where no screening took place before 1990. The Netherlands Cancer Registry provided detailed data on breast cancer incidence in 1989-1997 by tumour stage, age and region. Annual age-adjusted incidence rates of all breast cancers and advanced cancers, defined as large tumours T2+ with lymph node and/or distant metastases, were compared with rates in 1989. In general, breast cancer incidence rose strongly in the early 1990s, especially in the age category 50-69 years (estimated annual percentage change (EAPC) 4.25; 95% CI 1.70, 6.86). The increase was mainly due to the increase in small T1 cancers and ductal carcinoma in situ. However, in women aged 50-69, advanced cancer incidence rates showed a significant decline by 12.1% in 1997 compared with 1989 (EAPC -2.14, 95% CI 3.47, -0.80), followed by a breast cancer mortality reduction of similar size after approximately 2 years. We confirm that breast cancer screening initially leads to a temporary strong increase in the breast cancer incidence, which is followed by a significant decrease in advanced diseases in the women invited for screening. It is evident that breast cancer screening contributes to a reduction in advanced breast cancers and breast cancer mortality. PMID- 15292937 TI - Allelic imbalance at 1p36 may predict prognosis of chemoradiation therapy for bladder preservation in patients with invasive bladder cancer. AB - Invasive bladder cancers have been treated by irradiation combined with cis platinum (CDDP) as a bladder preservative option. The aim of this study was to find a marker for predicting patient outcome as well as clinical response after chemoradiation therapy (CRT) by investigating allelic loss of apoptosis-related genes. A total of 67 transitional cell carcinomas of the bladder treated by CRT (median dose: 32.4 Gy of radiation and 232 mg of CDDP) were studied. We investigated allelic imbalances at 14 loci on chromosomes 17p13 and 1p36 including the p53 and p73 gene regions by fluorescent multiplex PCR based on DNA from paraffin-embedded tumour specimens and peripheral blood. The response to CRT was clinical response (CR) in 21 patients (31%), partial response (PR) in 31 (46%), and no change(NC) in 15 (22%). There was no statistical correlation between treatment response and clinical parameters, such as tumour grade, stage, radiation dose, or CDDP dose. The frequencies of allelic imbalance for TP53 and TP73 were 21 and 56%, respectively; neither was correlated with clinical treatment response and tumour stage or grade. There was no statistical correlation between treatment response and allelic imbalance at the other 12 loci. We found a significant correlation between cancer-specific survival and an imbalance of D1S243 (P=0.0482) or TP73 (P=0.0013) using a Log-rank test, although other loci including TP53 did not correlate with survival (P=0.4529 Multivariate analysis showed performance status (P=0.0047), recurrence (P=0.0017), and radiation doses (P=0.0468) were independent predictive factors for cancer specific survival. However, an allelic imbalance of TP73 was the most remarkable independent predictive factor of poor patient survival (P=0.0002, risk ratio: 3382). Our results suggest that the allelic loss of the p73 gene predicts a clinical outcome of locally advanced bladder cancer when treated by CRT. PMID- 15292938 TI - Upregulation of p16(INK4A) and Bax in p53 wild/p53-overexpressing crypts in ulcerative colitis-associated tumours. AB - In ulcerative colitis (UC)-associated tumours, p53 gene mutations and p53 protein overexpression are frequently found in early stages, but the two types of alteration do not always coincide. To clarify this discrepancy, p53 mutations and expression of p53-associated molecules were analysed in UC-associated dysplasias by a combination of microdissection, polymerase chain reaction-direct sequencing and immunohistochemistry at the single crypt level. Mismatch of p53 protein overexpression (+)/mutation (-) or p53 overexpression (-)/gene mutation (+) was found in nine crypts in regenerative mucosa (19 crypts), in 27 in low-grade dysplasia (41), in one in high-grade dysplasia (5) and in 12 in invasive carcinomas (17). Regarding these mismatched crypts of the first type, significant increase in p16(INK4A) and Bax expression was found. The Ki-67 labelling index was depressed in such p53-diffusely positive lesions with the wild-type p53 gene, compared to their p53-diffusely positive and mutant type counterparts. p16(INK4A) was upregulated indirectly as part of the negative feedback, and increase in Bax, directly controlled by wild-type p53, indicates upregulation of apoptosis. No significant relation with p53-related gene products was detected with the p53 protein overexpression (-)/p53 mutation (+) mismatch. Therefore, a tumorigenesis pathway independent of p53 dysfunction appears to exist in association with ulcerative colitis. PMID- 15292939 TI - Cervical HPV infection and neoplasia in a large population-based prospective study: the Manchester cohort. AB - Cytology and histology records and cervical samples for HPV assay were obtained from a prospective cohort of 49 655 women attending clinics for routine cervical cytology in or near Manchester between 1988 and 1993. The women were followed up for cytological abnormality and neoplasia through the cytology laboratory's records. HPV at entry was assayed in an age- and period-stratified random sample of 7278 women and in prevalent and incident CIN3 cases. The prevalence of newly diagnosed CIN3 increased with time since last normal smear, indicating that most cases persist for several years. CIN3 prevalence did not increase further for screening intervals exceeding 5 years, however, suggesting that CIN3 eventually regresses cytologically. CIN2 prevalence increased less steeply with screening interval, while the prevalence of lesser abnormality was almost independent of screening interval. The prevalence of oncogenic HPV at entry declined from 19% among women aged under 25 to less than 3% at age 40 or above. Oncogenic HPV infection was strongly predictive of subsequent CIN3 (OR 17.2, 95% CI 10.4-28.4), but only weakly related to CIN2 (OR 2.3, 95% CI 0.5-10.7) and lesser abnormality (OR 1.4, 95% CI 0.8-2.5). At current incidence rates, the lifetime risk of developing CIN3 will be 9% in this population. The cumulative risk of CIN3 diagnosis among cytologically normal women with oncogenic HPV detected at entry was 28% (CI 18-43%) after 14 years. Persistence of oncogenic HPV may be more sensitive and specific than cytology for early detection of CIN3 and invasive cancer. PMID- 15292940 TI - Correlation of in vitro infiltration with glioma histological type in organotypic brain slices. AB - Diffuse invasion of the brain, an intrinsic property of gliomas, renders these tumours incurable, and is a principal determinant of their spatial and temporal growth. Knowledge of the invasive potential of gliomas is highly desired in order to understand their behaviour in vivo. Comprehensive ex vivo invasion studies including tumours of different histological types and grades are however lacking, mostly because reliable physiological invasion assays have been difficult to establish. Using an organotypic rodent brain slice assay, we evaluated the invasiveness of 42 grade II-IV glioma biopsy specimens, and correlated it with the histological phenotype, the absence or presence of deletions on chromosomes 1p and 19q assessed by fluorescent in situ hybridisation, and proliferation and apoptosis indices assessed by immunocytochemistry. Oligodendroglial tumours with 1p/19q loss were less invasive than astrocytic tumours of similar tumour grade. Correlation analysis of invasiveness cell proliferation and apoptosis further suggested that grade II-III oligodendroglial tumours with 1p/19q loss grow in situ as relatively circumscribed compact masses in contrast to the more infiltrative and more diffuse astrocytomas. Lower invasiveness may be an important characteristic of oligodendroglial tumours, adding to our understanding of their more indolent clinical evolution and responsiveness to therapy. PMID- 15292941 TI - Coordinate hypermethylation at specific genes in prostate carcinoma precedes LINE 1 hypomethylation. AB - In prostate carcinoma (PCa) increased DNA methylation ('hypermethylation') occurs at specific genes such as GSTP1. Nevertheless, overall methylation can be decreased ('hypomethylation') because methylation of repetitive sequences like LINE-1 retrotransposons is diminished. We analysed DNA from 113 PCa and 36 noncancerous prostate tissues for LINE-1 hypomethylation by a sensitive Southern technique and for hypermethylation at eight loci by methylation-specific PCR. Hypermethylation frequencies for GSTP1, RARB2, RASSF1A, and APC in carcinoma tissues were each >70%, strongly correlating with each other (P<10(-6)). Hypermethylation at each locus was significantly different between tumour and normal tissues (10(-11)82% of PCas. PCa may fall into three classes, that is, with few DNA methylation changes, with frequent hypermethylation, or with additional LINE-1 hypomethylation. PMID- 15292942 TI - Allelic losses on chromosome 3p are accumulated in relation to morphological changes of lung adenocarcinoma. AB - We performed allelotyping analysis at nine regions on chromosome 3p using 56 microdissected samples from 23 primary lung adenocarcinomas to examine the process of progression within individual lung adenocarcinoma with various grades of differentiation. Identical allelic patterns among various grades of differentiation were found in eight cases. Accumulation of allelic losses from high to lower differentiated portions was found in seven cases and accumulation of allelic losses from low to higher differentiated portions was found in five cases. Various allelic patterns among various grades of differentiation were found in three cases. These results suggested that allelic losses on 3p play an important role in morphological changes of lung adenocarcinomas. We also investigated the relationship between allelic losses on 3p and histological subtypes of lung adenocarcinoma. The frequencies of allelic losses at 3p14.2 and telomeric region of 3p21.3 were higher in papillary type tumour (nine out of 14, 64% and 11 out of 15, 73%) than in bronchioloalveolar carcinoma-type tumour (one out of 8, 13%; P=0.031 and four out of 12, 33%; P = 0.057). These results indicated that allelic losses at 3p14.2 and telomeric region of 3p21.3 are related to pattern of the proliferation of lung adenocarcinoma. PMID- 15292943 TI - Frequent downregulation of 14-3-3 sigma protein and hypermethylation of 14-3-3 sigma gene in salivary gland adenoid cystic carcinoma. AB - 14-3-3 sigma:, a target gene of the p53 tumour suppressor protein, has been shown to regulate the cell cycle at the G2/M checkpoint. Recent studies have demonstrated that 14-3-3 sigma is downregulated by hypermethylation of the CpG island in several types of cancer. In this study, we investigated the expression and methylation status of 14-3-3 sigma in human salivary gland adenoid cystic carcinoma (ACC) and mucoepidermoid carcinoma (MEC). Immunohistochemical analysis revealed that the positive expression rate of 14-3-3 sigma in ACC (one out of 14) was markedly lower than that in MEC (ten out of 10). Since most of the ACCs carried the wild-type p53 protein, downregulation of 14-3-3 sigma in ACC may not be due to the dysfunction of p53 pathway. Microdissection-methylation-specific PCR revealed that frequent hypermethylation of the 14-3-3 sigma gene was observed in ACC when compared to that in MEC. In cultured-ACC cells, we confirmed the downregulation of 14-3-3 sigma via hemimethylation of the gene by sequencing analysis after sodium bisulphite treatment. Furthermore, re-expression of 14-3-3 sigma in the ACC cells was induced by the treatment with DNA demethylating agent, 5-aza-2'-deoxycytidine. Irradiation apparently induced the enhanced expression of 14-3-3 sigma and G2/M arrest in normal salivary gland cells; however, in the ACC cells, neither induction of 14-3-3 sigma nor G2/M arrest was induced by irradiation. These results suggest that downregulation of 14-3-3 sigma might play critical roles in the neoplastic development and radiosensitivity of ACC. PMID- 15292944 TI - Timed flat infusion of 5-fluorouracil increases the tolerability of 5 fluorouracil/docetaxel regimen in metastatic breast cancer: a dose-finding study. AB - A dose-finding study was undertaken to determine the maximum-tolerated dose, and the recommended dose of docetaxel in combination with 12-h timed (22:00-10:00) flat infusion of 5-fluorouracil (5-FU) in metastatic breast cancer patients. This schedule seems to reduce the occurrence of stomatitis of the docetaxel and infusional 5-FU regimen. PMID- 15292947 TI - Electrochemical sensors for environmental monitoring: design, development and applications. AB - The advancement in miniaturization and microfabrication technology has led to the development of sensitive and selective electrochemical devices for field-based and in situ environmental monitoring. Electrochemical sensing devices have a major impact upon the monitoring of priority pollutants by allowing the instrument to be taken to the sample (rather than the traditional way of bringing the sample to the laboratory). Such devices can perform automated chemical analyses in complex matrices and provide rapid, reliable and inexpensive measurements of a variety of inorganic and organic pollutants. Although not exhaustive due to the vast amounts of new and exciting electrochemical research, this review addresses many important advances in electrochemical sensor design and development for environmental monitoring purposes. Critical design factors and development issues including analytical improvements (e.g. detection limits), microfabrication and remote communication are presented. In addition, modern environmental applications will be discussed and future perspectives considered. PMID- 15292948 TI - Baseline survey of concentrations of toxaphene congeners in fish from European waters. AB - The European Union project "Investigation into the monitoring, analysis and toxicity of toxaphene"(MATT) began in 1997 involving participants from the Netherlands, Ireland, Norway and Germany. Concentration information, analytical methodology and statistical interpretation of 207 samples covering 23 different fish species from European waters are presented for three toxaphene indicator congeners: CHBs 26, 50 and 62 (CHB = chlorobornane). Concentrations for the Sigma3CHBs were more elevated in fish from more northern latitudes, such as the Barents and Norwegian Sea, compared to fish from Irish, Dutch and German waters. Concentrations were lowest in shellfish and in fish species having low lipid content and were highest in medium/high lipid species. Females from a number of fish species were shown to contain significantly higher concentrations than those observed in male fish. Overall no samples were shown to exceed existing German MRL legislation, with only one Greenland halibut sample shown to exceed Canadian TDI recommendations. PMID- 15292949 TI - Measurement of chemical oxygen demand (COD) in natural water samples by flow injection ozonation chemiluminescence (FI-CL) technique. AB - COD determination based on ozone oxidation of alpha-naphthol combined with UV radiation (UV-O3) has been studied in the present work. Utilizing the phenomenon that luminol can be oxidized by the dissolved ozone to produce luminescence, we have established a new method of utilizing aqueous chemiluminescence to determine COD. The kinetics and mechanism of the ozonation reaction of alpha-naphthol have been investigated in order to gain a better understanding of the general applicability and limitation of the technique. Real world samples were analyzed and the results show that the relative error of COD(FI-CL) measurement for water samples was < 10%. Compared with the results of the conventional potassium permanganate method, the COD values of the FI-CL method are consistently higher (0-20% relative). The higher COD values suggest that the ozone-UV system is a more effective oxidation technique. PMID- 15292950 TI - Trace metal speciation and contamination in an intertidal estuary. AB - An analysis of the geochemical distribution of selected trace metals among various geochemical phases of the sediments in the Tees estuary was carried out using a sequential extraction technique and Differential Stripping Voltammetry. The sediments of the estuary are mainly organic rich clay silts and metal concentrations exceed those in the water column. Speciation results show that contamination of the estuary is mainly from anthropogenic sources. Pb and Zn are associated with the reducible, residual and oxidisable fractions. The speciation pattern of Cd was similar to those of lead and zinc. However there were also some exchangeable and bound to carbonate fractions although these were less significant. Cu is largely associated with the oxidisable and residual fractions, with insignificant bound to carbonate, exchangeable and reducible fractions. The most bioavailable forms of the metals are the free inorganic ions. Total metal concentrations in the estuary display a downward trend since the 1970s. PMID- 15292951 TI - Ecological interpretation of metal contents and contaminant source characterisation of sediments from a megatidal estuary. AB - Sediments and the associated biota constitute an important compartment in the biogeochemical cycle of trace metals in soft substrate megatidal estuaries. The relationship between physicochemical, ecological properties, metal concentrations determined in megatidal estuary sediments from the French coast of the English Channel, the Baie des Veys and the macrobenthic organisms, are analysed, interpreted and reported. Total concentrations of zinc, cadmium, lead and copper were measured in mudflats and saltmarsh sediments using Differential Pulse Voltammetry. Sediment characteristics were obtained by measurement of particle size, water content, total organic carbon and carbonate content, using AFNOR standards. A semi-quantitative scale was used for assessing the density of the macrobenthic flora and fauna at each sampling site. Analysis of data obtained from this study showed a correlation between the concentrations of monitored trace metals and species of the macrobenthic fauna. The results of this study show that the physico-chemical characteristics of the sediments affect the retention of metals in the sediment and this in turn affects the biota. PMID- 15292952 TI - Mercury in chemical fractions of recent pelagic sediments of the sea of Japan. AB - The distribution of mercury (Hg) in chemical fractions (H2O, 0.05 M Na2-EDTA pH 3, 1 M HCl, humic and fulvic acids, and non-hydrolysing residue) of recent pelagic sediment cores of the Sea of Japan (East Sea) was studied. Total Hg content in the sediments was rather low: 83 +/- 30 (21-173) etag g(-1), indicating the absence of substantial specific sources of the element in the deep part of the sea. Hg content within the sediment core varied by a factor of 1.3 1.8, showing peaks that coincide with the near-surface and buried sediment slices of light brown and brown "oxidized" colours and evidencing Hg redox-sensitive diagenetic redistribution. Hg exerted its maximum mobility in the near-surface sediment strata as a component of water-soluble organic matter. Despite the predominance of fulvic acids in extracted humus fractions, humic acids were a much more efficient concentrator for Hg (0-79 vs. 188-233 microg Hg g(-1) C(org), respectively). Nevertheless, the most refractory non-hydrolyzing residue (humin) fraction contained the principal Hg pool in the sediments. Hg content in all the extracted fractions decreased with core depth, thus indicating Hg immobilization as a principal tendency in Hg fate during post-depositional diagenesis. PMID- 15292953 TI - Mercury fractionation in contaminated soils from the Idrija mercury mine region. AB - Mercury (Hg) fractionation was investigated in contaminated soil in the Idrija Hg mine region, Slovenia. The main aim of this study was to test and apply sequential extraction and quantification of different Hg phases in order to estimate the mobility and potential bioavailability of Hg in contaminated soils. Separation of Hg phases was performed by means of a selective sequential extraction procedure complemented by volatilization of elemental mercury (Hg0). The influence of temperature, moisture and storage on Hg0 volatilization was also investigated. The total Hg concentrations varied between 8.4 and 415 mg kg(-1) and were up to 40-fold higher than the maximum permissible set by Slovenian legislation. Fractionation measurements indicated cinnabar as the predominant Hg fraction, followed by Hg0. Accumulation of cinnabar predominantly occurred in coarse grained flood plain sediments, where on average it constituted more than 80% of total Hg. In contrast non-cinnabar fractions were found to be enriched in areas where fine grained material was deposited, reaching up to 60% of total Hg. The strong positive correlation (R2 = 0.71-0.99) among non-cinnabar fractions suggested that these fractions predominantly control the mobility and potential bioavailability of Hg. Sample pretreatment before fractionation influenced the partition of Hg between different fractions, and therefore fractionation in fresh, nontreated samples is suggested. In addition, the specificity of the extraction steps needs further attention, as it was shown that some extraction steps, such as the organo-chelating Hg fraction, do not provide meaningful results. This further suggests that protocols for mercury fractionation need further harmonization in order to improve the comparability of the results and their use in risk assessment. Volatile mercury fluxes averaged between 0.04 and 6.5 ng g(-1) h(-1). Good agreement (R2 = 0.81-0.95) was found between the non cinnabar fractions and evaporation of Hg0. Both the temperature and sample moisture had significant effects on mercury volatilization. The results in this study were obtained at 70 degrees C, which may be somewhat high, in particular for bacterial activity which may also play an important role in Hg volatilization. Therefore it is strongly suggested that further optimisation of the protocol to assess Hg volatilization from soil is required. PMID- 15292954 TI - Predictive models of copper runoff from external structures. AB - A general model for annual runoff rate predictions of total copper from naturally patinated copper on buildings at specific urban or rural sites of low chloride influence has been deduced from laboratory and field data. All parameters within the model have a physical meaning and include the average annual rain acidity (pH), the annual rain quantity and the geometry of a building in terms of surface inclination. In 70% of all reported annual runoff rates, the predicted values are within 30% from the observed values. The individual and interactive effect of rain composition in terms of pH, sulfate, chloride and nitrate concentration was investigated in immersion experiments in artificial rain water representative of urban and rural sites of Europe. The results show pH to have a dominating effect on patina dissolution, nitrate to have a small inhibiting effect, whereas no significant effect was seen for chloride and sulfate. In case pH data are not available, a model has been statistically deduced from field data by considering SO2 as influencing parameter, rather than pH. The predictability with the SO2 model is not as good as with the pH model i.e. the pH model should preferentially be used since it is a better predictor and all parameters within the model can be physically explained. PMID- 15292955 TI - Characterisation of weathered hydrocarbon wastes at contaminated sites by GC simulated distillation and nitrous oxide chemical ionisation GC-MS, with implications for bioremediation. AB - An extended analytical characterisation of weathered hydrocarbons isolated from historically contaminated sites in Alberta is presented. The characterisation of soil extracts, chromatographically separated into component classes, by GC simulated distillation and nitrous oxide (N2O) chemical ionisation (CI) GC-MS provides new insights into the composition of the residual oil at these sites, the principal partition medium for risk critical components. The combined polar and asphaltene content of representative soil extracts ranged from 40% w/w to 70% w/w of the oils encountered. An abundance of C14-C22 2-4 ring alicyclics, alkylbenzenes and benzocycloparaffins is prevalent within the saturate fractions of site soils. Implications for the partitioning of risk critical compounds at sites with weathered hydrocarbons and the practical application of bioremediation technologies are discussed. PMID- 15292956 TI - [Do fixed-angle T-plates offer advantages for distal radius fractures in elderly patients?]. AB - In the treatment of distal radius fractures, plate osteosynthesis using fixed angle T-plates has become more common. Higher stability often allows functional aftertreatment in metaphyseal and articular fractures. So far it remains unclear whether these advantages also apply to elderly patients who commonly suffer from osteoporosis and reduced cooperativeness. Therefore, we evaluated the radiological loss of correction during fracture consolidation in patients aged more than 70 years. Fixed-angle plates were used in 44 patients (mean age: 79.4 years) while conventional T-plates were used in 30 patients (mean age: 78.2 years). Postoperative immobilization for 6 weeks by plaster or external fixator was performed in all patients. The loss of correction was significantly lower for fixed-angle plates (4.6% vs 40.0%). As a result of this study, we have gradually reduced immobilization in favor of early functional treatment. In a recent study fixed-angle plates so far seemed to permit stable fracture fixation. Our results underline the advantage of stable fixation in displaced fractures of the distal radius even in osteoporotic bone of elderly patients. PMID- 15292957 TI - [Bronchial rupture combined with luxation fracture of the thoracic spine following direct trauma]. AB - Tracheobronchial injuries in blunt thoracic trauma are very rare (incidence: under 1%), with potentially devastating consequences. Appropriate pre-, intra-, and postoperative management is mandatory to ensure the patient's survival and maintain lung function. We report the case of a 62-year-old male patient hit by a tree over the chest while cutting down trees, suffering a rupture of the right bronchus and a tear of the trachea combined with a luxation fracture of the thoracic spine between Th2 and Th3 (without neurological deficit). With immediate suture of the torn bronchus and trachea and stabilization of the spine fracture on the following day, we achieved a successful outcome in this patient. To our knowledge, this is the first description in the literature of the combination of both injuries. PMID- 15292958 TI - [Anterior cruciate ligament reconstruction with hamstring tendons in the young]. AB - Regarding the controversial discussion about how and when to operate a patient with an ACL lesion and still open physes,we routinely perform ACL reconstruction in those patients. We evaluated 30 patients with a mean age of 14.2 years at the time of operation (range: 10-18) and a mean follow-up of 30.8 months (range: 13 77). The ACL reconstruction was performed using a four-strand hamstring graft. Fixation was strictly extracortical using an endobutton and a suture washer. The placement of the graft was transepiphyseal. Using the IKDC score, 86.7% were classified as normal or nearly normal. In three cases an ACL insufficiency recurred during the first 12 months postoperatively. There was no growth disturbance. Expecting a poor outcome when treating an ACL lesion conservatively during the growth period and carefully performing the operation, we were able to show that the method provided satisfactory results and should be considered an operative method of choice. PMID- 15292959 TI - [Diagnostic apparatus in the shock trauma room]. AB - Opinions vary with regard to the equipment and structural furnishings required for adequate management of the trauma patient in the dedicated shock suite. In order to assess the current situation in Germany, we conducted a survey of the 76 centers participating in the Polytrauma Registry of the DGU. Fifty-one questionnaires were returned by centers representing all levels of care. Responses revealed, for example, that not all centers possess capabilities for conventional radiography in the shock suite (7/51). Only 20 centers had a fixed table; the remaining 24 hospitals used either an image converter or a mobile X ray unit. A dedicated ultrasound scanner was provided for the shock suite in 39 of 51 centers responding. Dedicated computed tomography scanners were provided for the shock suite in only eight centers (one dedicated trauma center, three level 3 centers, four university hospitals). All eight scanners use helical CT technology; at least three of the units are 8- or 16-slice. Of 51 shock suites, 12 are air-conditioned in compliance with sterile criteria (and are officially designated as surgical suites), while the remaining 39 are not. In acute cases, emergency surgeries can be performed in the shock suite in 37 centers, but not in the remaining 14 shock suites. According to the survey, slightly less than half of the hospitals responding are un-satisfied with the shock suite infrastructure ( n=24) and, of these, 13 centers are actively planning changes (the necessary financial resources have been guaranteed in 10 centers). Fourteen centers desire changes but do not currently have the required money. Information provided by Philips and Siemens suggests that the cost of furnishing a new shock suite ranges between 1.4 and 1.7 million euros. Responses to our survey show that a large gap remains between wishes and reality in the technical infrastructure in many shock suites in Germany. PMID- 15292960 TI - [Treatment of dislocated 3- and 4-part fractures of the proximal humerus with an angle-stabilizing fixation plate]. AB - In the presented prospective study 35 consecutive patients with displaced 3- and 4-part fractures of the proximal humerus, including fracture dislocations, were treated with a fixator plate comprising angular stability between May 2001 and December 2002. After 18.5 (8-29) months 31 (89%) fractures were available for follow-up. Good and very good results were obtained in 64%. A poor result was documented in 23%. 64% of the patients had no or mild pain, 71% were able to abduct the arm over 90 degrees . Fracture classification according to Neer and AO had no influence on the outcome, with a mean Constant Score of 76 points. Partial avascular necrosis (AVN) of the humeral head was seen in 16% of all cases representing 4% of the fractures without dislocation and 80% of the fracture dislocations. Fracture dislocation (p=0.02) and AVN (p=0.005) had a negative effect on the Constant Score, with AVN being a predictor for a high level of pain (p=0.04). Secondary dislocation of the greater tuberosity was seen in two patients, loosening of screws in one patient and a fracture below the plate in another one. Secondary dislocation or loss of reduction of the head was not recorded. Angle stable plate fixation with tension band wiring of the tuberosities is an effective and safe option to treat this difficult fractures, also in elderly patients with osteoporotic bone. Because 40% of the 4-part fractures with fracture dislocation yielded a satisfactory or better result, the plate fixator with angular stability may be an alternative to prosthetic replacement in selected cases. PMID- 15292961 TI - The zinc finger transcription factor Egr-1 is upregulated in arsenite-treated human keratinocytes. AB - Arsenite is a human carcinogen that may induce cancer in skin, liver, kidney, bladder or lung. Arsenite executes its toxic effects by the induction of signaling cascades. In particular, the activation of the stress-induced protein kinase c-Jun N-terminal protein kinase and p38 and the phosphorylation and activation of the transcription factor c-Jun have been linked to the biological effects of arsenite. We analyzed whether arsenite has an impact on the biosynthesis of the zinc finger transcription factor Egr-1. Egr-1 transcription is upregulated following treatment of cells with hormones, cytokines or toxic chemicals, and thus Egr-1 integrates many signaling cascades with changes in gene expression patterns. Here, we show by Western blot experiments that arsenite induces a transient synthesis of Egr-1 in human HaCaT keratinocytes. Egr-1 biosynthesis was activated by arsenite concentrations insufficient for the induction of c-Jun biosynthesis. This arsenite-triggered Egr-1 biosynthesis was completely inhibited by the mitogen-activated protein kinase kinase inhibitor PD98059 and by AG1487, an epidermal growth factor (EGF) receptor-specific tyrosine kinase inhibitor. These results indicate that activation of the EGF receptor as well as stimulation of the mitogen activated/extracellular signal regulated protein kinase is essential for arsenite-induced upregulation of Egr-1. Moreover, we detected an elevated transcriptional activation potential of the ternary complex factor Elk1, a key transcriptional regulator of serum response element-driven gene transcription. The Egr-1 5'-flanking region contains five serum response elements. Accordingly, we observed an increase in Egr-1 promoter activity as a result of arsenite treatment. The fact that low concentrations of arsenite are sufficient to induce Egr-1 biosynthesis suggests that Egr-1 may be an integral part of arsenite-triggered signaling cascades leading to tumor formation or cell death via alterations of the cellular genetic program. PMID- 15292962 TI - [Ethics and dermatology]. PMID- 15292963 TI - [Highlights in dermato-endocrinology]. PMID- 15292964 TI - [Mice and mitochondria]. PMID- 15292965 TI - ["Relaxant" awake but still relaxed]. AB - We report on a 23-year-old female patient who underwent removal of the implants after maxillary surgery. At the end of surgery the administration of anaesthetic agents was discontinued. During the following 30 min several attempts were made to wake the patient, but she did not respond to verbal or pain stimuli. No changes in heart rate, blood pressure vegetative reactions such as sweating, lacrimation, or mydriasis were noted. Protective reflexes like coughing could not be elicited. After 30 min neuromuscular monitoring was applied and indicated residual muscle paralysis after the use of mivacurium. The patient was again sedated and transferred to the ICU, where she was mechanically ventilated for an additional 9 h. An atypical cholinesterase was determined as the underlying reason for the prolonged action of mivacurium. Retrospectively, the patient remembered the attempted wake-up period in detail. However, she reported no feelings of fear or helplessness because she had faith in the anaesthesiologist, a close friend of the patient's family for many years, who kept her calm and comfortable by talking to her during the entire period. Several months after the incident, the patient reported having neither increased fear of surgery nor any negative psychological effects on her life following this incident of awareness. PMID- 15292966 TI - [Venous thromboembolism prophylaxis]. AB - Venous thromboembolism is a common and frequent complication of hospitalized patients. Some venous thromboembolisms may be subclinical, while others present as symptomatic deep vein thrombosis and/or pulmonary embolism. Venous thromboembolism and pulmonary embolism contribute significantly to inhospital morbidity and mortality. The risk of venous thromboembolism is aggravated by dispositional and/or expositional risk factors. In patients at intermediate or high risk of venous thromboembolism, additional pharmacological thromboembolism prophylaxis becomes mandatory. PMID- 15292967 TI - [On past practices and future directions of informed consent in (radiation) oncology]. AB - BACKGROUND AND PURPOSE: Informed consent, especially in oncology, is predominantly seen from a legal point of view. Such a limited perspective runs the risk of reducing informed consent to some tiresome formalism. The present article highlights how the relationship between patient and physician might be enriched by a comprehensive historicocultural understanding of informed consent. The authors show in which future directions the practice of informed consent might develop. MATERIAL AND METHODS: Analysis of historical and forensic literature regarding informed consent. RESULTS: With the terms "information" and "consent" the last 2500 years of medical history can schematically be divided in three epochs: the first epoch started around 500 years BC, lasted until the 19th century AC and was dominated by the principle of "salus aegroti suprema lex". The patient's benefit was exclusively defined by the treating physician. Formal consent was not required in those times. The era of enlightment brought new ideas to Europe, especially the principle of individual autonomy. In 1894, the Supreme Court of the German Reich decided that any medical intervention without the patient's consent was regarded as physical injury and was thus illegal. Systematic requirements regarding patient information on planned medical interventions were not known. The beginning of the third epoch is marked by the introduction of the term "informed consent" in modern medicine in 1957. Since then, a comprehensive information of the patient is seen as a prerequisite for consent. The patient's right of self-determination is attributed a higher legal and moral value than the physician's concept of the proposed treatment. Nowadays, the debate regarding informed consent is dominated by the continuing differentiation of modern medicine, the development of medical practice as part of the service sector, and the changing ways how patients see themselves. CONCLUSIONS: Social and legal developments have strongly influenced medical practice in the past. The importance of informed consent will continue to rise in the future, while the emphasis of the physician's task will shift from information to counseling. Informed consent will be increasingly established as independent service. PMID- 15292968 TI - Incidence, therapy and prognosis of colorectal cancer in different age groups. A population-based cohort study of the Rostock Cancer Registry. AB - PURPOSE: Determination of frequency, treatment modalities used and prognoses of colorectal cancer in a population-specific analysis in relation to age. MATERIAL AND METHODS: In 1999 and 2000, 644/6,016 patients were documented as having colorectal carcinomas in the Cancer Registry of Rostock. 39 patients were excluded (16 cases: "in situ" carcinomas; 23 cases: insufficient data). Three age groups were formed: < 60 years, 60-74 years; > or = 75 years. RESULTS: The relative percentage of colorectal cancer increases with advanced age (< 60 years 7%; 60-74 years 12%, > or = 75 years 15%; p < 0.001). In older patients with stage III carcinomas, adjuvant treatment was done less frequently in accordance with the treatment recommendations (< 60 years 83-89%; 60-74 years 67-77%; > or = 75 years 29-36% according to stage and tumor localization); in stage IV, the use of chemotherapy was reduced (< 60 years 87.5-100%; 60-74 years 38-47%; > or = 75 years 33-37%). In the univariate analysis, age > or = 75 years (4-year survival rates: < 60 years 68 +/- 4.1%; 60-74 years 58 +/- 2.8%; > or = 75 years 38 +/- 3.7%), UICC stage and surgical treatment had a significant effect on prognosis. Adjuvant treatment had no significant effect on the whole population but on patients with UICC stage III and IV. In the multivariate analysis, however, the only independent prognostic parameters were age > or = 75 years (p = 0.001), performance of chemotherapy (colon cancer) or radiochemotherapy (rectal cancer; p = 0.004-0.001), and tumor stage (p = 0.045-0.001). Sex (p = 0.063) and age between 60 and 74 years (p = 0.067) had a borderline influence. CONCLUSION: With increasing age, there is a departure in daily practice from the treatment recommendations. The patient's prognosis is dependent upon age (especially > or = 75 years), tumor stage, and therapy. PMID- 15292969 TI - Survival of patients in clinical stages I-IIIb of non-small-cell lung cancer treated with radiation therapy alone. Results of a population-based study in Southern Saxony-Anhalt. AB - BACKGROUND AND PURPOSE: Up to now, evidence about survival of patients with non small-cell lung cancer treated with radiation therapy alone is only available from clinical studies. The authors analyzed survival experience depending on several prognostic factors from a population-based cancer registry and compared this to survival data from the literature. PATIENTS AND METHODS: Between April 1996 and September 1999, 1,696 patients with lung cancer were recruited by the Halle Lung Cancer (HALLUCA) Study. 1,183 patients were diagnosed as having non small-cell lung cancer, and 188 in clinical stages I-IIIb (15.9%) were treated with radiation therapy alone. RESULTS: The median survival time of all patients was 10.2 months, the 2-year overall survival rate amounted to 15.8%. Besides tumor stage, radiation dose was found to be a statistically significant prognostic factor for survival in univariate analysis. The median survival time was 4.2 months for 66 patients treated with < 50 Gy, 10.7 months for 80 patients treated with 50 to < 60 Gy, and 18.9 months for 42 patients treated with >/= 60 Gy; the corresponding 2-year overall survival rates were 8.7%, 13.4%, und 35.2%. The significant influence of dose persisted even after adjustment for different confounders in a Cox regression model. CONCLUSION: Patients treated with 50 to < 60 Gy under a potentially curative therapeutic regimen had a significantly lower survival, compared to patients treated with >/= 60 Gy. In terms of quality assurance, the large proportion of patients treated with radiation doses below the curative range of >/= 60 Gy was unexpected. PMID- 15292970 TI - The influence of CA 125 and CEA levels on the results of (18)F-deoxyglucose positron emission tomography in suspected recurrence of epithelial ovarian cancer. AB - BACKGROUND AND PURPOSE: The follow-up of epithelial ovarian cancer (OCA) consists of clinical investigation, sonography, and tumor markers (TMs), especially CA 125. If tumor recurrence is suspected, other imaging modalities including positron emission tomography (PET) with (18)F-deoxyglucose (FDG) are often used. While there is still no consensus about the method of choice and the timing of its application, this study aims to find a TM threshold at which a PET would be appropriate. MATERIAL AND METHODS: A total of 90 PET studies and the associated CA 125 values (normal value < 35 U/ml) were available in 71 patients during the follow-up after primary therapy for OCA. In 48 studies a CEA value (normal value < 3 ng/ml) was also available. The results of PET imaging were related to the level of TM increase. RESULTS: In 23/90 studies the PET scan was normal. These patients had a median CA 125 of 13.3 U/ml (range 4.2-168 U/ml). In 67/90 studies the PET indicated a potential recurrence of OCA and the median CA 125 was 166.7 U/ml (range 13.3-4,060 U/ml). The TM levels were significantly different (p < 0.001, U-test). With one exception, there were no normal PET scans above CA 125 levels of 30 U/ml; between 20 and 30 U/ml PET was positive in 4/7 studies. CONCLUSION: In suspected recurrence of OCA, if imaging modalities are to be used, an FDG PET may be considered. Since the costs of this investigation are high, it should be restricted to clinical situations where it is likely to be most effective. In this study a PET indication is worthwhile at CA 125 levels of approximately 30 U/ml. PMID- 15292971 TI - Effectiveness and prognostic factors of radiotherapy for painful plantar heel spurs. AB - BACKGROUND AND PURPOSE: The efficacy of radiation treatment (RT) for plantar heel pain has been reported repeatedly. Yet, the results referring to the pain relief rate, to long-term effects and prognostic factors are not consistent. In this paper, the effectiveness (pain relief rate and long-term results) and prognostic factors of RT for plantar heel pain have been investigated. PATIENTS AND METHODS: From January 2000 to October 2000, 62 patients (73 heels) with painful plantar heel spurs and a minimum pain history of 3 months were treated and evaluated in a prospective study. Mean age was 54 years (range 28-84 years). All patients were treated with a total dose of 5 Gy in seven fractions (= one series), given twice a week at a single-dose sequence of 0.25-0.25-0.5-1.0-1.0-1.0-1.0 Gy (10-MV photons, source-skin distance [SSD] 100 cm, direct portal, field size 12 x 17 cm). The mean duration of heel pain before RT was 26 weeks (= 6.5 months; range 3 120 months). By means of a visual analog scale (VAS) the patients had to self assess the quantity of their heel pain once before, three times during and four times after RT at a longterm median follow-up of 28 and 40 months. Additionally, the patients had to assess their mechanical heel stress extent during RT. Effectiveness was estimated according to the patients' judgment of pain reduction. RESULTS: A significant reduction of heel pain extent measured by VAS has been observed already during the RT series (before RT: 6.3 +/- 1.5 vs. 3.8 +/ 2.1 at the end of RT; p < 0.001). 6 weeks after RT (FU 1) pain reduction (> 20%) was achieved in 60 heels (82.3%; n = 73), in 64 heels (91.4%; n = 70) after a mean follow-up of 28 months (FU 2), and in 61 heels (89.7%; n = 68) after a mean follow-up of 40 months (FU 3), respectively. Sufficient pain relief (> 80% compared to initial extent) was observed in 18/73 heels (24.6%) at FU 1 (FU 2: 42/70; 60.0%; FU 3: 37/68; 54.4%), including 13/73 heels (17.8%) with complete pain relief (FU 2: 39/70; 55.7%; FU 3: 36/68; 52.9%). Partial improvement (50-80% pain reduction) was observed in 27/73 heels (37.0%) at FU 1 (FU 2: 14/70; 20.0%; FU 3: 15/68; 22.1%), and minor partial improvement (20-50% pain reduction) in 15/73 heels (20.5%) at FU 1 (FU 2: 8/70; 11.4%; FU 3: 9/68; 13.2%), respectively. No change was seen in 13/73 heels (17.8%) at FU 1 (FU 2: 6/70; 8.6%; FU 3: 7/68; 10.3%). Older patients (p = 0.04) and patients who avoided heel stress during the period of RT (p < 0.01) demonstrated a better short-term response (FU 1); both effects were lost 28 and 40 months after RT. Moreover, significant differences in the extent of heel pain reduction by RT were observed in dependence on previous pain duration (at FU 2-3). CONCLUSION: The results confirm the high efficacy of RT in painful plantar spur and add new aspects to formerly published data concerning the time course of changes in heel pain reduction. Pain relief can be expected during and shortly after RT. In addition, the initial success can be transformed into effective long-term results > 2 years after RT; however, further improvement is not to be expected. As a new prognostic factor, the reduction of mechanical heel stress during RT may ameliorate the short-term results, whereas short heel pain history improves the long-term results. Especially for older patients, RT should be taken into consideration as primary treatment. PMID- 15292972 TI - Prospective study on exclusive, nonsurgical strontium-/yttrium-90 irradiation of pterygia. AB - PURPOSE: Prospective study to evaluate consecutive treatment results and to demonstrate safety and efficacy of nonsurgical, exclusive strontium-/yttrium-90 beta-irradiation of nonoperated pterygia. PATIENTS AND METHODS: Between November 1999 and March 2002, 20 patients with 21 primary pterygia and six patients with recurrent pterygia after former surgery were treated with exclusive strontium /yttrium-90 irradiation up to a total dose of 3,600 cGy (six fractions) and 4,800 cGy (eight fractions), respectively. All patients were referred from a single institution. The mean follow-up is 35.6 +/- 7.3 months (range 24-48 months). RESULTS: Prior to irradiation the mean horizontal diameter of all pterygia was 2.6 mm and shrank to a mean diameter of 1.6 mm after treatment (p = 0.0011, Student's t-test). The treatment led to a reduction in size of all 21 primary and all six recurrent pterygia. Visual acuity reached a value of 0.73 before and 0.82 after treatment. This improvement was not significant in Student's t-test (p = 0.12). The visual acuity did not decrease in any patient, complications were not observed, and in none of the 27 pterygia a recurrence developed CONCLUSION: Exclusive strontium-/yttrium-90 irradiation of the early and moderately advanced pterygium is a very efficient and very well-tolerated method of treatment. As to the therapeutic management, it is suggested to apply beta-irradiation prior to the development of an astigmatism-relevant pterygium, which requires excision. PMID- 15292973 TI - Radioprotective effects of amifostine in vitro and in vivo measured with the comet assay. AB - PURPOSE: The authors investigated whether a potential radioprotective effect of amifostine (WR-2721) after in vitro or in vivo administration can be detected with the comet assay. Moreover, it was determined whether radioprotection by WR 2721 is dependent on the concentration of amifostine or alkaline phosphatase (AP, the enzyme which activates the prodrug). Furthermore, the authors tried to detect possible interindividual differences in radioprotection by amifostine. MATERIAL AND METHODS: In vitro administration of amifostine: Freshly isolated lymphocytes from two healthy volunteers were incubated with different concentrations of AP (0 210 U/ml) and amifostine (0-5,000 microg/ml). IN VIVO ADMINISTRATION OF AMIFOSTINE: Blood samples were collected from six postoperative rectal cancer patients before and after intravenous administration of amifostine 500 mg (no pretreatment with radio- or chemotherapy). Leukocytes and lymphocytes were irradiated and repaired in vitro and investigated with the alkaline comet assay. The radioprotective effect was evaluated by calculating dose-modifying factors (DMFs) and the paired t-test. RESULTS: Amifostine alone did not alter the radiation-induced DNA damage in vitro. The addition of at least 0.5-1 U/ml AP was required. A significant radioprotective effect (p < 0.05) was seen after administration of amifostine in vitro for all concentrations investigated (250 5,000 microg/ml, initial DNA damage). A comparable radioprotective effect after in vivo administration of 500 mg amifostine was measured with a mean DMF of 0.87. Interindividual differences were present in vivo and in vitro. CONCLUSION: Amifostine 500 mg intravenously yields an adequate radioprotective concentration. The effect was only marginally improved by extreme concentrations of amifostine in in vitro experiments. The comet assay is capable of detecting small changes in radiosensitivity by amifostine. PMID- 15292974 TI - Exogenous modulation of TGF-beta(1) influences TGF-betaR-III-associated vascularization during wound healing in irradiated tissue. AB - BACKGROUND AND PURPOSE: Following preoperative radiotherapy prior to ablative surgery of squamous epithelial cell carcinomas of the head and neck region, wound healing disorders occur. Previous experimental studies showed altered expression of transforming growth factor-(TGF-)beta isoforms following surgery in irradiated graft beds. Altered levels of TGF-beta(1) are reported to promote fibrosis and to suppress vascularization during wound healing, whereas expression of TGF-beta receptor-III (TGF-betaR-III) is associated with vascularization. The aim of the study was to analyze the influence of anti-TGF-beta(1) treatment on TGF-betaR-III associated vascularization in the transition area between irradiated graft bed and graft. MATERIAL AND METHODS: Wistar rats (male, weight 300-500 g) underwent preoperative irradiation of the head and neck region with 40 Gy (four fractions of 10 Gy each; n = 16 animals). A free myocutaneous gracilis flap taken from the groin was then transplanted to the neck in all rats. The time interval between operation and transplantation was 4 weeks. Eight animals received 1 micro g anti TGF-beta(1) into the graft bed by intradermal injection on days 1-7 after surgery. On days 3, 7, 14, 28, 56, and 120, skin samples were taken from the transition area between transplant and graft bed and from the graft bed itself. Immunohistochemistry was performed using the ABC-POX method to analyze the TGF betaR-III and E-selectin expression. Histomorphometry was performed to analyze the percentage and the area of positively stained vessels. RESULTS: A significantly higher expression of TGF-betaR-III was seen in the irradiated and anti-TGF-beta(1)-treated graft bed in comparison to the group receiving preoperative irradiation followed by transplantation alone. The percentage of TGF betaR-III positively staining capillaries from the total amount of capillaries in the anti-TGF-beta(1)-treated graft bed was higher than in the group irradiated only. The total area of capillaries was also higher in the TGF-beta(1)-treated group. CONCLUSION: Neutralizing of TGF-beta(1) activity in irradiated tissue undergoing surgery leads to a higher expression of TGF-betaR-III and increased vascularization. TGF-betaR-III seems to be associated with newly formed blood vessels during neovascularization in wound healing. PMID- 15292975 TI - Experimental disentangling of spatial-compatibility and interhemispheric-relay effects in simple reaction time (Poffenberger paradigm). AB - Spatial-compatibility effects can be obtained in simple reaction time (SRT) provided that spatially distinct responses are frequently required. Since this effect is limited to trials with relatively long reaction times (RTs), Hommel (1996b) proposed that if the response does not occur shortly after stimulus detection, then the spatial code of the stimulus can interfere with that of the response. A series of experiments is reported showing that (a) spatial compatibility in SRT to lateralized stimuli is not an alternative, but rather a complementary, explanation to interhemispheric transfer time (contrary to what Hommel surmised), and (b) the spatial compatibility component is essentially limited to the first trial after shifting response preparation from one-half of the visual fields to the other, suggesting a mechanism akin to an orienting response. PMID- 15292976 TI - Mercury speciation in thawed out and refrozen fish samples by gas chromatography coupled to inductively coupled plasma mass spectrometry and atomic fluorescence spectroscopy. AB - Different sub-sampling procedures were applied for the determination of mercury species (as total mercury Hg, methylmercury MeHg+ and inorganic mercury Hg2+) in frozen fish meat. Analyses were carried out by two different techniques. After the sample material was pre-treated by microwave digestion, atomic fluorescence spectroscopy (AFS) was used for the determination of total Hg. Speciation analysis was performed according to the following procedure: dissolution of sample material in tetramethylammonium hydroxide (TMAH), derivatisation with sodium tetraethylborate (NaBEt4), extraction into isooctane and measurement with gas chromatography inductively coupled plasma mass spectrometry (GC-ICPMS) for the identification and quantification of methylmercury (MeHg+) and inorganic mercury (Hg2+). The concentration range of total Hg measured in the shark fillets is between 0.9 and 3.6 microg g(-1) thawed out shark fillet. Speciation analysis leads to > or =94% Hg present as MeHg+. Homogeneity, storage conditions and stability of analytical species and sample materials have great influence on analytical results. Sub-sampling of half-frozen/partly thawed out fish and analysis lead to significantly different concentrations, which are on average a factor of two lower. PMID- 15292977 TI - Transforming growth factor beta1 is not involved in 2,3,7,8-tetrachlorodibenzo- p dioxin-dependent release from contact-inhibition in WB-F344 cells. AB - TCDD (2,3,7,8-tetrachlorodibenzo- p-dioxin) is the most potent tumor promoter ever tested in rodents. Although it is known that most of the effects of TCDD are mediated by binding to the aryl hydrocarbon receptor (AhR), the mechanisms leading to tumor promotion still remain to be elucidated. Loss of contact inhibition is one characteristic hallmark in tumorigenesis. In WB-F344 cells, TCDD induces a release from contact-inhibition, which is manifested by a two- to three-fold increase in DNA-synthesis when TCDD (1 nM) is given to confluent cells. Since proliferation of epithelial cells is known to be inhibited by transforming growth factor beta (TGF-beta) we investigated whether decreased TGF beta expression mediates TCDD-dependent release from contact-inhibition in WB F344 cells. Expression of TGF-beta (type II) receptor in WB-F344 cells was shown by Western blot analysis. Exposure of exponentially growing WB-F344 cells to 0.1 ng/ml TGF-beta1 resulted in a 40% decrease in DNA synthesis, which could be blocked by pre-incubation with a neutralizing anti-TGF-beta1 antibody indicating that the TGF-beta receptor in WB-F344 cells is functionally active. Pre incubation of confluent, G1-arrested cultures with the neutralizing anti-TGF beta1 antibody did not lead to an increase in DNA synthesis, ruling out an involvement of TGF-beta1 in mediating contact-inhibition in WB-F344 cells. In accordance with this, Western blot analysis revealed that protein expression of TGF-beta1 was neither upregulated in confluent cultures nor decreased after TCDD treatment. We therefore conclude that TGF-beta1 is not involved in contact inhibition nor in TCDD-dependent release from contact-inhibition in WB-F344 cells. PMID- 15292978 TI - The burden of osteoporosis in Latin America. AB - Osteoporosis causes considerable morbidity, mortality and resource utilization in industrialized nations. Its burden is relatively well known in United States and Canada, but poorly studied in the rest of America. This study aimed to discover the burden of osteoporosis in Latin America through a review of literature and publicly available information. In this transversal and descriptive study, information from 20 countries in Latin American region was collected from diverse published and electronic sources. Rheumatologists and bone specialists were asked for additional information through a questionnaire created by consensus. In the year 2000, the population of the Latin America and Caribbean region was 524 million from diverse ethnic origins. On average, 5.5% of the population is 65 years and older. However, with life expectancy higher than 70 years in most countries, a significant growth in the elderly population is anticipated. Studies using World Health Organization's criteria for osteoporosis report 12-18% of women 50 years and older with vertebral osteoporosis and 8-22% with proximal femur osteoporosis. Community based studies in Argentina reported between 263 and 331 hip fractures per 100,000 people 50 years and older. Hospital based studies in Colombia, Chile, Brazil, Mexico, Panama, Peru and Venezuela reported between 40 and 362 hip fractures per 100,000 persons aged 50 and more. Between 17 and 37% of hip fracture sufferers die in the year following fracture. Prevalence of vertebral fractures in community-dwelling women aged 50 and more in Mexico is 19.35%. Data on other fractures are rare. Direct costs of a hip fracture ranged from $4500 to $6000. National gross income per capita in the region ranges from $410 to $7550. The burden of osteoporosis varies across countries with differences in populations and health resources. Considerable support for research is required, since numerous gaps in knowledge need to be filled to face the anticipated explosive growth in osteoporotic fractures. PMID- 15292980 TI - Comorbidities affect the impact of urinary incontinence as measured by disease specific quality of life instruments. AB - The aim of this study was to determine if the impact of urinary incontinence on activities of daily living, as measured by a disease-specific quality of life instrument, is dependent on comorbid conditions. Incontinent kidney transplant recipients participated in a survey to determine the impact of urinary incontinence on activities of daily living using the Incontinence Impact Questionnaire (IIQ-7). Similar information was collected from the charts of nontransplant incontinent women. Participants were matched for age, incontinence severity, pads per day, and leaks per day noted in a 3-day 24-h bladder diary. IIQ-7 scores from participants were compared using Mann-Whitney U tests. Nontransplant incontinent women reported a 200% greater affect of incontinence on activities of daily living than incontinent renal transplant recipients (35.5+/ 26.5 vs 12.9+/-15.4, p<0.0001) despite similarities in incontinence severity measures. Urinary incontinence has less of an impact on activities of daily living for renal transplant recipients than nontransplant incontinent women with similar incontinence severity measures because the disease-specific quality of life instrument used in this study was sensitive to their comorbid condition (transplant status). PMID- 15292984 TI - Does intubation really equal death in cirrhotic patients? Factors influencing outcome in patients with liver cirrhosis requiring mechanical ventilation. AB - OBJECTIVE: It is not known whether the poor outcome of ventilated cirrhotic patients is related to the severity of the underlying liver disease or to the severity of the acute illness for which ICU care is required. This study examines parameters both of chronic liver disease and of acute illness with regard to their influence on outcome in mechanically ventilated cirrhotic patients. DESIGN AND SETTING: Retrospective observational case series in a 9-bed medical ICU in an academic tertiary care center. PATIENTS AND MEASUREMENTS: Seventy-six consecutive cirrhotic patients who received mechanical ventilation were identified. Clinical and laboratory parameters were compared between ICU survivors and ICU deaths. RESULTS: There were 45/76 (59%) patients who died during their ICU stay. By univariate analysis, the Child-Pugh score, its components (serum bilirubin, prothrombin time), ALT, creatinine concentration, a clinical suspicion of infection, and the APACHE II score, but not the acute physiology score (APS), differed significantly between ICU survivors and ICU non-survivors. The Child Pugh score was highly correlated to ICU mortality both in logistic regression analysis and receiver-operating characteristics analysis. Conversely, markers of acute illness, in particular the APS component of the APACHE II score, did not predict ICU survival. CONCLUSIONS: Markers of advanced chronic liver disease but not of the severity of acute illness are correlated to ICU outcome in ventilated cirrhotic patients. The outcome of advanced cases (Child-Pugh score of 12 and above) is poor. PMID- 15292983 TI - Year in review in intensive care medicine-2003. Part 3: intensive care unit organization, scoring, quality of life, ethics, neonatal and pediatrics, and experimental. PMID- 15292985 TI - New regulations for the care of the critically ill patients in Italy. PMID- 15292986 TI - Dyskalemia and cerebral insult: therapeutic barbiturate coma, endogenous catecholamine release or both? PMID- 15292987 TI - WY-14643 and 9- cis-retinoic acid induce IRS-2/PI 3-kinase signalling pathway and increase glucose transport in human skeletal muscle cells: differential effect in myotubes from healthy subjects and Type 2 diabetic patients. AB - AIMS/HYPOTHESIS: To determine the effects of peroxisome proliferator-activated receptor alpha (PPARalpha) and retinoid X receptor (RXR) agonists on insulin action, we investigated the effects of Wy-14643 and 9- cis-retinoic acid (9- cis RA) on insulin signalling and glucose uptake in human myotubes. METHODS: Primary cultures of differentiated human skeletal muscle cells, established from healthy subjects and Type 2 diabetic patients, were used to study the effects of Wy-14643 and 9- cis-RA on the expression and activity of proteins involved in the insulin signalling cascade. Glucose transport was assessed by measuring the rate of [(3)H]2-deoxyglucose uptake. RESULTS: Wy-14643 and 9- cis-RA increased IRS-2 and p85alpha phosphatidylinositol 3-kinase (PI 3-kinase) mRNA and protein expression in myotubes from non-diabetic and Type 2 diabetic subjects. This resulted in increased insulin stimulation of protein kinase B phosphorylation and increased glucose uptake in cells from control subjects. Myotubes from diabetic patients displayed marked alterations in the stimulation by insulin of the IRS-1/PI 3 kinase pathway. These alterations were associated with blunted stimulation of glucose transport. Treatment with Wy-14643 and 9- cis-RA did not restore these defects but increased the basal rate of glucose uptake. CONCLUSIONS/INTERPRETATION: These results demonstrate that PPARalpha and RXR agonists can directly affect insulin signalling in human muscle cells. They also indicate that an increase in the IRS-2/PI 3-kinase pathway does not overcome the impaired stimulation of the IRS-1-dependent pathway and does not restore insulin stimulated glucose uptake in myotubes from Type 2 diabetic patients. PMID- 15292989 TI - A genetic linkage map of grape, utilizing Vitis rupestris and Vitis arizonica. AB - A genetic linkage map of grape was constructed, utilizing 116 progeny derived from a cross of two Vitis rupestris x V. arizonica interspecific hybrids, using the pseudo-testcross strategy. A total of 475 DNA markers-410 amplified fragment length polymorphism, 24 inter-simple sequence repeat, 32 random amplified polymorphic DNA, and nine simple sequence repeat markers-were used to construct the parental maps. Markers segregating 1:1 were used to construct parental framework maps with confidence levels >90% with the Plant Genome Research Initiative mapping program. In the maternal (D8909-15) map, 105 framework markers and 55 accessory markers were ordered in 17 linkage groups (756 cM). The paternal (F8909-17) map had 111 framework markers and 33 accessory markers ordered in 19 linkage groups (1,082 cM). One hundred eighty-one markers segregating 3:1 were used to connect the two parental maps' parents. This moderately dense map will be useful for the initial mapping of genes and/or QTL for resistance to the dagger nematode, Xiphinema index, and Xylella fastidiosa, the bacterial causal agent of Pierce's disease. PMID- 15292990 TI - Molecular mapping of resistance to Pyrenophora tritici-repentis race 5 and sensitivity to Ptr ToxB in wheat. AB - Tan spot, caused by Pyrenophora tritici-repentis (Ptr), is an economically important foliar disease in the major wheat growing areas of the world. Multiple races of the pathogen have been characterized based on their ability to cause necrosis and/or chlorosis in differential wheat lines. Isolates of race 5 cause chlorosis only, and they produce a host-selective toxin designated Ptr ToxB that induces chlorosis when infiltrated into sensitive genotypes. The international Triticeae mapping initiative (ITMI) mapping population was used to identify genomic regions harboring QTLs for resistance to fungal inoculations of Ptr race 5 and to determine the chromosomal location of the gene conditioning sensitivity to Ptr ToxB. The toxin-insensitivity gene, which we are designating tsc2, mapped to the distal tip of the short arm of chromosome 2B. This gene was responsible for the effects of a major QTL associated with resistance to the race 5 fungus and accounted for 69% of the phenotypic variation. Additional minor QTLs were identified on the short arm of 2A, the long arm of 4A, and on the long arm of chromosome 2B. Together, the major QTL on 2BS identified by tsc2 and the QTL on 4AL explained 73% of the total phenotypic variation for resistance to Ptr race 5. The results of this research indicate that Ptr ToxB is a major virulence factor, and the markers closely linked to tsc2 and the 4A QTL should be useful for introgression of resistance into adapted germplasm. PMID- 15292991 TI - The development of ISSR-derived SCAR markers around the SEASONAL FLOWERING LOCUS (SFL) in Fragaria vesca. AB - Fragaria vesca is a short-lived perennial with a seasonal-flowering habit. Seasonality of flowering is widespread in the Rosaceae and is also found in the majority of temperate polycarpic perennials. Genetic analysis has shown that seasonal flowering is controlled by a single gene in F. vesca, the SEASONAL FLOWERING LOCUS ( SFL). Here, we report progress towards the marker-assisted selection and positional cloning of SFL, in which three ISSR markers linked to SFL were converted to locus-specific sequence-characterized amplified region (SCAR1-SCAR3) markers to allow large-scale screening of mapping progenies. We believe this is the first study describing the development of SCAR markers from ISSR profiles. The work also provides useful insight into the nature of polymorphisms generated by the ISSR marker system. Our results indicate that the ISSR polymorphisms originally detected were probably caused by point mutations in the positions targeted by primer anchors (causing differential PCR failure), by indels within the amplicon (leading to variation in amplicon size) and by internal sequence differences (leading to variation in DNA folding and so in band mobility). The cause of the original ISSR polymorphism was important in the selection of appropriate strategies for SCAR-marker development. The SCAR markers produced were mapped using a F. vesca f. vesca x F. vesca f. semperflorens testcross population. Marker SCAR2 was inseparable from the SFL, whereas SCAR1 mapped 3.0 cM to the north of the gene and SCAR3 1.7 cM to its south. PMID- 15292992 TI - Evaluating the genetic basis of multiple-locule fruit in a broad cross section of tomato cultivars. AB - Lycopersicon esculentum accessions bearing fasciated (multiloculed) fruit were characterized based on their flower organ and locule number phenotypes. Greenhouse and field evaluations indicate that increases in locule number are associated with increases in the number of other floral organs (e.g., sepals, petals, stamens) in all stocks. F1 complementation, F2 segregation analysis, and genetic mapping indicate that at least four loci account for increases in the number of carpels/locules in these stocks. The most significant of these map to the bottoms of chromosomes 2 and 11 and correspond to the locule number and fasciated loci. All stocks tested were fixed for mutations at the fasciated locus, which maps to the 0.5-cM interval between the markers T302 and cLET24J2A and occurs in at least three allelic forms (wild type and two mutants). One of the fasciated mutant alleles is associated with nonfused carpels and repressed recombination and may be due to a small inversion or deletion. The other two loci controlling locule number correspond to the lcn1.1 and lcn2.2 loci located on chromosomes 1 and 2, respectively. PMID- 15292993 TI - [The proof of paternity. An andrological-forensic challenge in historical perspective]. AB - For centuries, difficulties have occurred in determining unresolved paternities. In addition to the modern standard methods, such as the examination of DNA or serological proof, expert opinion on fertility once played an important role. The andrological difference between incapability to fertilise and the inability to participate in sexual intercourse was also distinguished historically. Of special significance was the discovery of spermatozoa by the medical student Johan Ham in 1677 and their further investigation by Antoni van Leeuwenhoek.Recently, modern DNA methods have also been applied for historical investigations. Illustrious examples are the DNA analysis in the case of Kaspar Hauser of Ansbach and the dispute about Thomas Jefferson, President of the U.S., fathering a child by one of his slaves. In this discourse, a medicinal-forensic review of the development of expert opinion, illustrated with historical case studies, is given. PMID- 15292995 TI - [Biofeedback for urinary bladder dysfunctions in childhood. Indications, practice and the results of therapy]. AB - In children, abnormal behavior during micturition, i.e. detrusor/sphincter dyscoordination, causes persistent voiding problems, urinary incontinence and/or recurrent urinary tract infections in up to 15% of cases. Contractions of the external urethral sphincter during micturition lead to functional subvesical obstruction. Nowadays, biofeedback training is the most suitable therapy. Biofeedback training for children is based on the assumption that relaxation and contraction of the urinary external sphincter is a habitual phenomenon and can be restored. With specially developed, computer-assisted biofeedback programs, sphincter contraction and relaxation can be transformed into acoustic or visual signals. Acoustic or optical feedback indicates relaxation and contraction control to the patient. The residual urine volume should subsequently be assessed. The results should be reviewed after each micturition. Poor compliance sometimes makes biofeedback training impossible. Further biofeedback training at home is a reasonable suggestion. Good results-a response rate of up to 90% demonstrates that biofeedback training is successful in the treatment of detrusor sphincter dyscoordination. After effective therapy, associated urinary tract infections and vesicoureterorenal reflux may disappear. PMID- 15292996 TI - [Vulvar carcinoma. Diagnosis and therapy]. AB - Vulvar cancer is a rare entity. It appears mostly in older women aged 70-79 years with a slow tendency to younger age. More than 90% of the tumors show a squamous differentiation. The correspondent preneoplasia is VIN 3. This lesion occurs in women mostly younger than 35 years. Experts assume vulvar cancer to appear in two different types:HPV-induced type in younger women and non-HPV-dependent type in older women. The preneoplasia VIN 3 already should be treated by resection or destruction. Invasive carcinomas stage I or II can be treated by wide local excision. The inguinofemoral lymph nodes should be resected if invasion exceeds 1 mm in depth. In larger primary tumors, vulvectomy with bilateral inguinofemoral node dissection is indicated. In advanced tumor stages, multimodal concepts are applied: primary radiotherapy or radiochemotherapy may precede a salvage operation. PMID- 15293001 TI - [Multimodal diagnosis of multiple and heterogeneous liver lesions in a young patient]. AB - The classification of liver lesions is often problematic in particular if they are multiple and show an heterogeneous shape. Here we report of a young patient with multiple liver lesions of up to 3 cm size. Using ultrasound, the lesions were hyper-, hypoechogen or mixed. In serial contrast enhanced CT scans some of the lesions showed the typical enhancement pattern of hemangiomas, however, the diagnosis could still not be faithfully determined for all lesions. Therefore, the patient was conducted to contrast enhanced MRI (Gd-DTPA and MnDPDP). While with Gd-DTPA some of the lesions showed a strong enhancement, they remained hypointense after administration of MnDPDP. Finally to exclude a metastatic disease a (99m)Tc-erythrocyte SPECT was performed confirming the diagnosis of hemangiomas for most of the lesions. Diagnosis was not assessed by biopsy because this would only clarify the diagnosis for one or few of the lesions. The patient was subsequently followed up for 3 years and all lesions remained unchanged. This case clearly illustrates the difficulty to get a certain diagnosis of multiple liver lesions with heterogeneous appearance despite the multimodal diagnostic conduct. PMID- 15293003 TI - [Pulmonary function testing before ablative methods]. AB - Laser-induced thermotherapy (LITT) and radiofrequency thermoablation (RFTA) are increasingly used for pulmonary interventions. Primarily patients with severe functional limitations precluding a surgical approach are selected for these procedures. In this patient group a valid preinterventional risk assessment is of paramount importance. The occurrence of a pneumothorax is one of the most important complications associated with these procedures. Therefore, the functional capacity and pulmonary reserve of these patients should allow for at least short periods of lung collapse. The periinterventional risk of these patients can be estimated from basic lung function studies when certain comorbidities are excluded. PMID- 15293004 TI - [Organopathy. About interdisciplinary terminology from the radiology viewpoint]. PMID- 15293007 TI - [Synaptic vesicle proteins and psychiatric disorders]. AB - Synaptic vesicle proteins modulate the release of neurotransmitters in the synaptic cleft via regulation of vesicle transport, membrane fusion and exocytosis. Due to their relevance for neural and synaptic plasticity, they represent an important object of molecular psychiatric research. There is increasing evidence that they play a significant role in the pathophysiology of several psychiatric disorders. Furthermore, psychotropic drugs differentially modify the expression of synaptic vesicle proteins; thus, this group of molecules is also of considerable interest from a therapeutic point of view. PMID- 15293008 TI - [Dual diagnosis of psychosis and addiction. From principles to practice]. AB - The prevalence of comorbidity of psychosis and substance abuse/addiction has been on the rise during the last 10-20 years. Meanwhile, dual diagnosis patients (DD patients) represent a large core group among patients with schizophrenia, and they are difficult to treat. Biological, psychological, and social factors may account for the comorbidity, and the single factors may interact with each other. DD patients tend to have poor compliance and unfavorable outcomes with frequent psychotic relapses and hospitalizations. Efficient treatment models integrate traditional psychiatric therapy and therapy of addiction and modify or adjust the two components to each other. The most successful programs offer integrated treatment for both disorders in one setting. These programs focus on outpatient treatment, they offer pharmacotherapy, motivation therapy, psychoeducation, cognitive-behavioral therapy, and family interventions, and they can achieve significant improvements of social adjustment and decreases of substance use. PMID- 15293009 TI - [Wilhelm Strohmayer (1874-1936)]. PMID- 15293010 TI - [Alfred Adler (1870-1937)]. PMID- 15293013 TI - Consequences of symbiosis for food web dynamics. AB - Basic Lotka-Volterra type models in which mutualism (a type of symbiosis where the two populations benefit both) is taken into account, may give unbounded solutions. We exclude such behaviour using explicit mass balances and study the consequences of symbiosis for the long-term dynamic behaviour of a three species system, two prey and one predator species in the chemostat. We compose a theoretical food web where a predator feeds on two prey species that have a symbiotic relationships. In addition to a species-specific resource, the two prey populations consume the products of the partner population as well. In turn, a common predator forages on these prey populations. The temporal change in the biomass and the nutrient densities in the reactor is described by ordinary differential equations (ODE). Since products are recycled, the dynamics of these abiotic materials must be taken into account as well, and they are described by odes in a similar way as the abiotic nutrients. We use numerical bifurcation analysis to assess the long-term dynamic behaviour for varying degrees of symbiosis. Attractors can be equilibria, limit cycles and chaotic attractors depending on the control parameters of the chemostat reactor. These control parameters that can be experimentally manipulated are the nutrient density of the inflow medium and the dilution rate. Bifurcation diagrams for the three species web with a facultative symbiotic association between the two prey populations, are similar to that of a bi-trophic food chain; nutrient enrichment leads to oscillatory behaviour. Predation combined with obligatory symbiotic prey interactions has a stabilizing effect, that is, there is stable coexistence in a larger part of the parameter space than for a bi-trophic food chain. However, combined with a large growth rate of the predator, the food web can persist only in a relatively small region of the parameter space. Then, two zero-pair bifurcation points are the organizing centers. In each of these points, in addition to a tangent, transcritical and Hopf bifurcation a global heteroclinic bifurcation is emanating. This heteroclinic cycle connects two saddle equilibria where the predator is absent. Under parameter variation the period of the stable limit cycle goes to infinity and the cycle tends to the heteroclinic cycle. At this global bifurcation point this cycle breaks and the boundary of the basin of attraction disappears abruptly because the separatrix disappears together with the cycle. As a result, it becomes possible that a stable two-nutrient-two-prey population system becomes unstable by invasion of a predator and eventually the predator goes extinct together with the two prey populations, that is, the complete food web is destroyed. This is a form of over-exploitation by the predator population of the two symbiotic prey populations. When obligatory symbiotic prey-interactions are modelled with Liebig's minimum law, where growth is limited by the most limiting resource, more complicated types of bifurcations are found. This results from the fact that the Jacobian matrix changes discontinuously with respect to a varying parameter when another resource becomes most limiting. PMID- 15293014 TI - Refining the measurement of rate constants in the BIAcore. AB - When estimating rate constants using the BIAcore surface plasmon resonance (SPR) biosensor, one must have an accurate mathematical model to interpret sensogram data. Several models of differing complexity are discussed, including the effective rate constant (ERC) approach. This model can be shown formally to be good within O(Da) in the limit of small Damkohler number Da, which is the ratio of the reaction rate to the rate of transport to the surface. Numerical results are presented that show that except for very slow reactions, parameter estimates from the ERC model are very close to those estimated using a more complicated model. The BIAcore measures the behavior of an evanescent wave whose signal strength decays as it penetrates into the device. It is shown that this decay does not appreciably affect the sensogram readout at low Da, but at moderate Da can lead to situations where two vastly different rate constants can produce the same short-time sensogram data. PMID- 15293015 TI - Intermingled basins in a two species system. AB - We present simple examples of (competitive) two species systems with complicated dynamic behaviour. From almost all initial conditions one of the two species dies out. But the survivor is unpredictable: The basins of the two chaotic one-species attractors are everywhere dense and intermingled. PMID- 15293016 TI - Traveling wave solutions for a one-dimensional crawling nematode sperm cell model. AB - In this paper, we proved that the one-dimensional crawling nematode sperm cell model proposed by Mogilner and Verzi (2003) supports traveling wave solutions if there is no disassembly of unbundled filaments in the cell. Uniqueness of traveling wave is established under additional assumptions and numerical examples are also given in the paper. Mathematical methods used include dynamical system techniques, implicit function theorem and global bifurcation theory. PMID- 15293018 TI - A stage structured predator-prey model and its dependence on maturation delay and death rate. AB - Many of the existing models on stage structured populations are single species models or models which assume a constant resource supply. In reality, growth is a combined result of birth and death processes, both of which are closely linked to the resource supply which is dynamic in nature. From this basic standpoint, we formulate a general and robust predator-prey model with stage structure with constant maturation time delay (through-stage time delay) and perform a systematic mathematical and computational study. Our work indicates that if the juvenile death rate (through-stage death rate) is nonzero, then for small and large values of maturation time delays, the population dynamics takes the simple form of a globally attractive steady state. Our linear stability work shows that if the resource is dynamic, as in nature, there is a window in maturation time delay parameter that generates sustainable oscillatory dynamics. PMID- 15293019 TI - Unequal crossover dynamics in discrete and continuous time. AB - We analyze a class of models for unequal crossover (UC) of sequences containing sections with repeated units that may differ in length. In these, the probability of an 'imperfect' alignment, in which the shorter sequence has d units without a partner in the longer one, scales like qd as compared to 'perfect' alignments where all these copies are paired. The class is parameterized by this penalty factor q. An effectively infinite population size and thus deterministic dynamics is assumed. For the extreme cases q = 0 and q = 1, and any initial distribution whose moments satisfy certain conditions, we prove the convergence to one of the known fixed points, uniquely determined by the mean copy number, in both discrete and continuous time. For the intermediate parameter values, the existence of fixed points is shown. PMID- 15293021 TI - Abstracts of the Annual Congress of the European Association of Nuclear Medicine (EANM). Helsinki, Finland, September 5-8, 2004. PMID- 15293029 TI - Evaluation of chemical pretreatment of contaminated soil for improved PAH bioremediation. AB - The efficiency of several chemical treatments as potential enhancers of the biodegradation of polycyclic aromatic hydrocarbons (PAHs) in contaminated soil was evaluated by analyzing the mineralization of 14C-labeled phenanthrene, pyrene, and benzo(a)pyrene. The effect of nonionic surfactants with Fenton oxidation and combinations of surfactants with the Fenton oxidation was evaluated in a microtiter plate assay. The surfactants selected for the study were Tween 80, Brij 35, Tergitol NP-10, and Triton X-100. The addition of Fenton's reagent significantly enhanced the mineralization of pyrene at the two concentrations studied: 2.8 M H2O2 with 0.1 M FeSO4 and 0.7 M H2O2 with 0.025 M FeSO4. Phenanthrene mineralization was also positively induced by the Fenton treatments. However, none of the treatments had a significant effect on benzo(a)pyrene mineralization. Surfactant additions at concentrations of 20% and 80% of the aqueous critical micelle concentration did not significantly affect the mineralization rates. When surfactant addition was combined with the Fenton oxidation, reduced mineralization rates were obtained when compared with mineralization after Fenton's treatment alone. The results indicate that the addition of Fenton's reagent may enhance the mineralization of PAHs in contaminated soil, whereas the addition of surfactants has no significant beneficial effect. The efficiency of the Fenton oxidation may decrease when surfactants are added simultaneously with Fenton's reagent to contaminated soil. PMID- 15293031 TI - Folate requirements of the 2-keto-L-gulonic acid-producing strain Ketogulonigenium vulgare LMP P-20356 in L-sorbose/CSL medium. AB - In this study, the requirements for growth factors of Ketogulonigenium vulgare LMP P-20356, a 2-keto-L-gulonic acid-producing strain of particular interest for the manufacture of vitamin C, were assessed. Various growth factors were studied in order to obtain improved growth of the strain when cultured in an L sorbose/corn steep liquor medium. Cultures grown in the presence of reduced mono- and polyglutamated folate derivatives showed a 15- to 20-fold higher biomass content than control cultures lacking these supplements, indicating that the strain has a requirement for folate. Although most folate derivatives used in this study promoted growth, the amplitude of the response varied depending on the compound used. Dihydrofolic acid was found to be the most active form, followed by 5-formyltetrahydrofolic acid, 5-methyltetrahydrofolic acid and tetrahydrofolic acid. Folic acid had no effect. The effectiveness of polyglutamated derivatives was inversely proportional to the polyglutamated chain-length of the derivative used. Our results suggest that the rate-limiting step in the utilisation of monoglutamated folates is most probably related to their transport and/or their intracellular interconversion rather than their polymerisation into polyglutamated forms (physiological forms). The industrial production of 2-keto-L gulonic acid by K. vulgare LMP P-20356 could be improved by using media in which low-molecular-weight reduced folates are present. PMID- 15293017 TI - Mathematical modeling of tumor-induced angiogenesis. PMID- 15293032 TI - A cloned Bacillus halodurans multicopper oxidase exhibiting alkaline laccase activity. AB - The gene product of open reading frame bh2082 from Bacillus halodurans C-125 was identified as a multicopper oxidase with potential laccase activity. A homologue of this gene, lbh1, was obtained from a B. halodurans isolate from our culture collection. The encoded gene product was expressed in Escherichia coli and showed laccase-like activity, oxidising 2,2'-azino-bis(3-ethylbenz-thiazoline-6-sulfonic acid), 2,6-dimethoxyphenol and syringaldazine (SGZ). The pH optimum of Lbh1 with SGZ is 7.5-8 (at 45 degrees C) and the laccase activity is stimulated rather than inhibited by chloride. These unusual properties make Lbh1 an interesting biocatalyst in applications for which classical laccases are unsuited, such as biobleaching of kraft pulp for paper production. PMID- 15293034 TI - Practical approach to prenatal posterior fossa abnormalities using MRI. AB - This review focuses on the optimum use of fetal MRI as an additional imaging tool to sonographic data in posterior fossa (PF) abnormalities in the second and third trimesters of gestation. We have chosen three particular situations to demonstrate the value of MRI as a complementary investigation to US: (1) the pattern of increased fluid-filled space of the PF, (2) decreased cerebellar sonographic biometry and (3) the diagnosis of focal echogenic lesions of the cerebellum. For increased fluid-filled space of the PF and decreased cerebellar sonographic biometry, a practical approach is proposed, largely based on prenatal imaging, especially MRI. PMID- 15293035 TI - 3-D CT angiographic demonstration of a neonatal ductus arteriosus aneurysm with development of ductal calcification: are the "ductus bump", ductus arteriosus aneurysm, and ductal calcification related? AB - A "ductus bump" was noted as an incidental finding on a chest radiograph in a newborn infant with congenital cystic adenomatoid malformation. A chest CT performed on the first day of life demonstrated this to be a ductus arteriosus aneurysm (DAA), which subsequently thrombosed. Ductal calcification was noted on follow-up imaging. We propose that the "ductus bump" may actually be a DAA, which resolves presumably by thrombus formation. Ductal calcification may also be related to the regression of the thrombus. PMID- 15293038 TI - Dietary fiber in conservative management of chronic renal failure. AB - Conservative management of chronic renal failure has included a variety of dietary manipulations. Supplementation of dietary fiber to reduce adverse symptoms is a novel approach, but whether it can make an effective clinical impact is yet undetermined. This commentary accompanies a brief report of improved well-being during fiber supplementation. It explains some of the background thinking and early trials of fiber supplementation in chronic renal failure patients. PMID- 15293040 TI - Pathophysiology, diagnosis, and treatment of methylmalonic aciduria-recent advances and new challenges. AB - Classical methylmalonic aciduria is a relatively rare inborn error of branched chain amino acid metabolism, occurring in 1:50,000 to 1:80,000 newborns. Three decades after its recognition, major progress has been made in survival and prevention of neurological sequelae in affected children, if the diagnosis is made early and treatment and follow-up care are meticulous. Therapy consists of a specially formulated protein diet, carnitine supplementation, and vigorous emergency treatment during intercurrent illnesses aimed at preventing the development of catabolism. Recently the clinician has been challenged by partially unexpected long-term complications. These include chronic neurological symptoms, specifically an extrapyramidal movement disorder caused by progressive destruction of the basal ganglia, which are similar to those observed in other organic acid disorders, such as propionic aciduria or glutaric aciduria type I. Unexpected and unique is the development of chronic renal failure in a major subset of patients. As the pathophysiological basis of renal failure is still obscure, no causative treatment is available and hemodialysis may become necessary. Experience with transplantation of liver, kidney, or kidney and liver is very limited and allows as yet no conclusions. Interdisciplinary research efforts in this field should reveal new pathophysiological links and hopefully provide additional therapeutic approaches. PMID- 15293039 TI - Acacia gum supplementation of a low-protein diet in children with end-stage renal disease. AB - Patients with end-stage renal disease (ESRD) die in the absence of renal replacement therapy (RRT). In developing countries RRT is not uniformly available and treatment often relies on conservative management and intermittent peritoneal dialysis (IPD). This study investigates the possibility of using acacia gum supplementation to improve the quality of life and provide children with ESRD with a dialysis-free period. Three patients referred to our hospital with ESRD during a 3-month period were enrolled in a therapeutic trial to investigate the efficacy of acacia gum (1 g/kg per day in divided doses) as a complementary conservative measure aimed at improving the quality of life. Inclusion criteria included a pre-dialysis creatinine clearance of <5 ml/min, current dietary restrictions and supplementation, at least one dialysis session to control uremic symptoms, absence of life-threatening complications, and sufficient motivation to ensure compliance with the study protocol. One patient complied with the protocol for only 10 days and died after 6 months, despite IPD. Two patients completed the study. Both reported improved well-being. Neither became acidotic or uremic, and neither required dialysis during the study period. Both patients maintained urinary creatinine and urea levels not previously achieved without dialysis. In conclusion, dietary supplementation with acacia gum may be an alternative to renal replacement therapy to improve the quality of life and reduce or eliminate the need for dialysis in children with ESRD in some developing countries. PMID- 15293041 TI - On-line estimation of biomass concentration using a neural network and information about metabolic state. AB - This paper deals with the design of a neural network-based biomass concentration estimation system. This system is enhanced by the incorporation of information about the actual metabolism of the microorganism cultivated, which is taken from an on-line knowledge-based system. Two different design approaches have been investigated using the fed-batch cultivation of baker's yeast as the model process. In the first, metabolic state (MS) data were passed as additional input to the neural network; in the second, these data were used to select a neural network suitable for the specific MS. Two neural network types--feed-forward (Levenberg-Marquardt) and cascade correlation--were applied to this system and tested, and the performances of these neural networks were compared. PMID- 15293042 TI - A transient, positive effect of habitat fragmentation on insect population densities. AB - We conducted an experimental landscape study to test the hypotheses that: (1) habitat removal results in short-term increases in population density in the remaining habitat patches (the crowding effect); (2) following habitat removal, density is higher in landscapes with more, smaller patches and more habitat edge (i.e., a higher level of habitat fragmentation per se) than in less fragmented landscapes, for the same total amount of habitat on the landscapes; (3) this positive effect of fragmentation per se on density is larger in landscapes with smaller inter-patch distances; and (4) these last two effects should be reduced or disappear over time following habitat removal. Our results did not support the first hypothesis, but they provided some support for the other three hypotheses, for two of the four Coccinellid species studied. As in other empirical studies of fragmentation per se on population density, the effects of fragmentation per se were weak and positive (when they did occur). This is the first study to document a transient effect of fragmentation per se on population density, and to show that this effect depends on inter-patch distances. We suggest that fragmentation per se increased the rate of immigration to patches, resulting in higher population densities in more fragmented landscapes. PMID- 15293043 TI - What limits the Serengeti zebra population? AB - The populations of the ecologically dominant ungulates in the Serengeti ecosystem (zebra, wildebeest and buffalo) have shown markedly different trends since the 1960s: the two ruminants both irrupted after the elimination of rinderpest in 1960, while the zebras have remained stable. The ruminants are resource limited (though parts of the buffalo population have been limited by poaching since the 1980s). The zebras' resource acquisition tactics should allow them to outcompete the ruminants, but their greater spatial dispersion makes them more available to predators, and it has been suggested that this population is limited by predation. To investigate the mechanisms involved in the population dynamics of Serengeti zebra, we compared population dynamics among the three species using demographic models based on age-class-specific survival and fecundity. The only major difference between zebra and the two ruminants occurred in the first-year survival. We show that wildebeest have a higher reproductive potential than zebra (younger age at first breeding and shorter generation time). Nevertheless, these differences in reproduction cannot account for the observed differences in the population trends between the zebra and the ruminants. On the other hand, among species differences in first-year survival are great enough to account for the constancy of zebra population size. We conclude that the very low first-year survival of zebra limits this population. We provide new data on predation in the Serengeti and show that, as in other ecosystems, predation rates on zebras are high, so predation could hold the population in a "predator pit". However, lion and hyena feed principally on adult zebras, and further work is required to discover the process involved in the high mortality of foals. PMID- 15293044 TI - Mechanism of adhesion and detachment at the anterior end of Neoheterocotyle rhinobatidis and Troglocephalus rhinobatidis (Monogenea: Monopisthocotylea: Monocotylidae). AB - The anterior adhesion and detachment mechanisms observed for Neoheterocotyle rhinobatidis and Troglocephalus rhinobatidis (Monogenea: Monocotylidae) appear similar to those observed for the two other monopisthocotylean monogenean species with anterior apertures for which published data are available. This supports the theory that monogeneans with apertures may utilise a common mechanism. Adult anterior apertures can open and close and duct endings can evert during the adhesion phase and retract during detachment and searching behaviour. The adhesive is comprised of two secretory types, rod-shaped and spheroidal bodies, found within anterior apertures. These exit together and undergo mixing to produce the adhesive matrix in which elongate membranes from rod-shaped bodies are seen intermixed with a granular electrondense matrix. The morphology of the adhesive matrix differs from that found for some other monogenean taxa. Anterior detachment by these monocotylids appears to involve a depletion of rod-shaped bodies in ducts and mechanical withdrawal of the anterior end. PMID- 15293045 TI - The distribution of Entamoeba histolytica and Entamoeba dispar in northern, central, and southern Iran. AB - The present study was carried out from August 1999 through February 2002 in order to determine the prevalence of Entamoeba histolytica and Entamoeba dispar in three different climatic regions of Iran by using a PCR-RFLP method. A total of 16,592 stool samples were randomly collected from different age-groups in central, northern, and southern Iran in both urban and rural areas. The samples were examined by direct and formalin-ether concentration methods. A total of 226 samples were positive for E. histolytica/E. dispar cysts. Of these, 101 isolates were cultured and maintained successfully in Robinson's medium and were identified by the PCR-RFLP method. The study showed that 92.1% of isolates were E. dispar and 7.9% were E. histolytica or mixed infections. The ratio of E. histolytica to E. dispar was higher in southern regions (tropical and subtropical) than in the other two regions. This study demonstrated that E. dispar is the predominant species found among "cyst passers" in Iran. PMID- 15293046 TI - Efficacy of ivermectin in calves against a resistant Cooperia oncophora field isolate. AB - Since 1999, two Cooperia oncophora isolates, originally obtained from the United Kingdom, have been maintained by regular passage through calves at the Macdonald Campus, McGill University farm. One isolate, IVS, was originally susceptible to ivermectin, while the IVR isolate was originally resistant to ivermectin. These two isolates have been used to study the mechanisms of ivermectin resistance. To confirm the susceptible/resistant status after 4 years of passaging through calves, a controlled study was performed in which two worm-free calves were experimentally infected with IVS and another two worm-free calves with the IVR infective larvae. The calves were treated with ivermectin (0.2 mg/kg) subcutaneously (Ivomec Injection) 21 days after infection. Ivermectin at the recommended dose was 100% effective at eliminating the IVS isolate, since no eggs were found in feces, and no adult worms were found in the small intestine of the treated IVS-infected calf. In contrast, the IVR-infected calf continued to pass eggs in feces even after treatment with ivermectin, and adult worms (250) were found in the small intestine at necropsy. The untreated calves had 1,330 and 848 adult worms, respectively, for the IVS and IVR infected animals. PMID- 15293047 TI - Prenatal morphogenesis of the human incisive canal. AB - The literature describing the formation of the incisive canal is very bizarre. The fusion of the primary and secondary palatal processes leads to formation of a triangular seam, which erroneously has been taken for the future incisive canal. If so, the nasopalatine (incisive) nerve and its accompanying vessels were to run through the primary oronasal cavity, which is not compatible with our biological experience. This study was undertaken to shed light on this region of fusion. We focus on the formation of the incisive canal; the neighboring nasopalatine ducts, which are a transient formation, are mentioned where present. A series of seven horizontal cross-sections of human embryos and fetuses from the 7th to the 24th week of pregnancy (between 25 and 225 mm CRL, crown-rump-length) were examined histologically and partly reconstructed in 3D applying the software analySIS (Soft Imaging Systems, Munster, Germany). The incisive canal did not develop at the junction of the primary and the secondary palate, but within the primary palate rostral to that location. The nasopalatine nerve and the nasopalatine artery are structures that exist before ossification starts in the area of the future incisive canal. The neighboring nasopalatine ducts were found in regions laterally and anterolaterally of the nasopalatine nerve, and it was mostly separated from it by bone. In advanced stages of development, the nasopalatine duct only existed as single epithelial remnants and was prone to obliteration. PMID- 15293048 TI - Evolution of the larval peripheral nervous system in Drosophila species has involved a change in sensory cell lineage. AB - A key challenge in evolutionary biology is to identify developmental events responsible for morphological changes. To determine the cellular basis that underlies changes in the larval peripheral nervous system (PNS) of flies, we first described the PNS pattern of the abdominal segments A1-A7 in late embryos of several fly species using antibody staining. In contrast to the many variations reported previously for the adult PNS pattern, we found that the larval PNS pattern has remained very stable during evolution. Indeed, our observation that most of the analysed Drosophilinae species exhibit exactly the same pattern as Drosophila melanogaster reveals that the pattern observed in D. melanogaster embryos has remained constant for at least 40 million years. Furthermore, we observed that the PNS pattern in more distantly related flies (Calliphoridae and Phoridae) is only slightly different from the one in D. melanogaster. A single difference relative to D. melanogaster was identified in the PNS pattern of the Drosophilinae fly D. busckii, the absence of a specific external sensory organ. Our analysis of sensory organ development in D. busckii suggests that this specific loss resulted from a transformation in cell lineage, from a multidendritic-neuron-external-sensory-organ lineage to a multidendritic neuron-solo lineage. PMID- 15293049 TI - Characterization of five RALF-like genes from Solanum chacoense provides support for a developmental role in plants. AB - Five RALF (rapid alkalinization factor)-like genes, named ScRALF1 to 5, were isolated from fertilized ovule and ovary cDNA libraries of Solanum chacoense. They showed high sequence similarities with the RALF protein sequence from Nicotiana tabacum, and exhibited the characteristic architecture of RALF polypeptides. All ScRALFs were moderately to highly expressed at some stage of fruit maturation. ScRALF1 and ScRALF3 were predominantly expressed in ovaries and larger fruits, while ScRALF2, ScRALF4, and ScRALF5 were also expressed in other tissues, indicating that while some RALFs may be involved in fruit maturation, others could be involved in other developmental processes. Wounding or treatment of plants with growth regulators involved in plant defense responses had no significant impact on the mRNA level of any of these genes. These results suggest and support previous data showing that RALF peptides are more likely to act as a small peptide involved in plant development than in defense responses. PMID- 15293050 TI - Inappropriate annotation of a key defence marker in Arabidopsis: will the real PR 1 please stand up? AB - PR-1 has been extensively used as a marker for salicylic acid (SA)-mediated defence and systemic and local acquired resistance. The Arabidopsis Genome Project annotates At2g19990 as PR-1. This gene is also identified as PR-1 in two "full genome" Arabidopsis microarrays, and TAIR cites approximately 60 articles to describe its patterns of expression. However, most of these citations are incorrect; the probes used were not At2g19990, but a homologous gene At2g14610, which is annotated as "PR-1-like". Because of the potential for confusion, we analyzed the expression of both genes in Arabidopsis thaliana (L.) Heynh. At2g14610 (PR-1-like) showed the archetypal patterns of SA-responsive expression: mRNA levels increased following SA-treatment, inoculation with an avirulent (but not a virulent) strain of Pseudomonas syringae, and in wild-type (but not NahG) Arabidopsis infected with cauliflower mosaic virus (CaMV). In cpr5 mutants it was expressed constitutively. In contrast, expression of At2g19990 (annotated as PR 1) was detectable in neither SA-treated Col-0 nor in cpr5. Infection by virulent and avirulent isolates of P. syringae up-regulated expression, but to a similar level, and infection by CaMV induced a modest increase in expression in both the wild type and NahG. At2g19990, although pathogen responsive, does not show the SA dependent patterns of expression expected from a member of the PR-1 regulon, and its annotation as " PR-1" is inappropriate. The annotations should identify At2g14610 as the authentic PR-1. PMID- 15293051 TI - Cell-volume-dependent vascular smooth muscle contraction: role of Na+, K+, 2Cl- cotransport, intracellular Cl- and L-type Ca2+ channels. AB - This study elucidates the role of cell volume in contractions of endothelium denuded vascular smooth muscle rings (VSMR) from the rat aorta. We observed that hyposmotic swelling as well as hyper- and isosmotic shrinkage led to VSMR contractions. Swelling-induced contractions were accompanied by activation of Ca2+ influx and were abolished by nifedipine and verapamil. In contrast, contractions of shrunken cells were insensitive to the presence of L-type channel inhibitors and occurred in the absence of Ca2+ o. Thirty minutes preincubation with bumetanide, a potent Na+, K+, CI- cotransport (NKCC) inhibitor, decreased Cl(-)i content, nifedipine-sensitive 45Ca uptake and contractions triggered by modest depolarization ([K+]o = 36 mM). Elevation of [K+]o to 66 mM completely abolished the effect of bumetanide on these parameters. Bumetanide almost completely abrogated phenylephrine-induced contraction, partially suppressed contractions triggered by hyperosmotic shrinkage, but potentiated contractions of isosmotically shrunken VSMR. Our results suggest that bumetanide suppresses contraction of modestly depolarized cells via NKCC inhibition and Cl(-)i-mediated membrane hyperpolarization, whereas augmented contraction of isosmotically shrunken VSMR by bumetanide is a consequence of suppression of NKCC-mediated regulatory volume increase. The mechanism of bumetanide inhibition of contraction of phenylephrine-treated and hyperosmotically shrunken VSMR should be examined further. PMID- 15293052 TI - Leptin, catecholamines and free fatty acids related to reduced recovery delays after training. AB - The aim of this study was to observe the effects of exercise on plasma free fatty acids (FFA), the catecholamines (adrenaline, noradrenaline, dopamine) and leptin levels, before and after training, to determine their possible influence on the improvement of the regulation of energy homeostasis. Eleven trained rowers performed two exercise sessions (S1, S2) of 90 min each (70-75% V(.)O(2peak)), separated by a 36-week period of intense endurance training. Leptin, FFA and catecholamine plasma concentrations were measured both at the beginning and at the end of S1 and S2, and after recovery periods of 2 and 24 h. Training modified leptin levels in S2 as opposed to S1 ( P<0.001); in S1 leptin levels remained lower after a 24 h recovery whereas they returned to pre-exercise levels in S2. The respiratory exchange ratio was significantly reduced in S2 compared to S1 ( P=0.018). The FFA and leptin levels were correlated after a 24-h recovery in S1 ( r=0.87; P=0.0082), and after a 2-h recovery in S2 ( r=0.66; P=0.021). All data were expressed as the means and standard errors of the mean. In the two sessions, an immediate exercise effect was observed on the levels of the catecholamines, which did not persist after recovery. This training effect was apparent for all catecholamines in response to exercise, particularly on noradrenaline ( P=0.0006). The noradrenaline and leptin levels were correlated after a 2-h recovery in S2 ( r=-0.74; P=0.0042). We conclude that the effect of training on the response of noradrenaline to exercise seems to be involved in the delay in the normalization of leptin levels. We suggest that the amplitude of the noradrenaline response to exercise induced an increase in fat use and a rapid leptin recovery after exercise. The sensitivity of leptin to changes in the fat stores may be improved after training. Both training effects seemed to be involved in the reduction of the recovery time observed for the leptin levels. PMID- 15293053 TI - Effect of concurrent aerobic and resistance circuit exercise training on fitness in older adults. AB - The purpose of this study was to determine the physiological effects of a programmed accommodating circuit exercise (PACE) program consisting of aerobic exercise and hydraulic-resistance exercise (HRE) on fitness in older adults. Thirty-five volunteers were randomly divided into two groups [PACE group (PG) 8 men and 10 women, 68.3 (4.9) years, and non-exercise control group (CG) 7 men and 10 women, 68.0 (3.4) years). The PG participated in a 12-week, 3 days per week supervised program consisting of 10 min warm-up and 30 min of PACE (moderate intensity HRE and aerobic movements at 70% of peak heart rate) followed by 10 min cool-down exercise. PACE increased ( P<0.05) oxygen uptake ( V(.)O(2)) at lactate threshold [PG, pre 0.79 (0.20) l min(-1), post 1.02 (0.22) l min(-1), 29%; CG, pre 0.87 (0.14) l min(-1), post 0.85 (0.15) l min(-1), -2%] and at peak V(.)O(2) [PG, pre 1.36 (0.24) l min(-1), post 1.56 (0.28) l min(-1), 15%; CG, pre 1.32 (0.29) l min(-1), post 1.37 (0.37) l min(-1), 4%] in PG measured using an incremental cycle ergometer. Muscular strength evaluated by a HRE machine increased at low to high resistance dial settings for knee extension (9-52%), knee flexion (14-76%), back extension (18-92%) and flexion (50-70%), chest pull (6-28%) and press (3-17%), shoulder press (18-31%) and pull (26-85%), and leg press (21%). Body fat (sum of three skinfolds) decreased (16%), and high-density lipoprotein cholesterol (HDLC) increased (10.9 mg dl(-1)) for PG. There were no changes in any variables for CG. These results indicate that PACE training incorporating aerobic exercise and HRE elicits significant improvements in cardiorespiratory fitness, muscular strength, body composition, and HDLC for older adults. Therefore, PACE training is an effective well-rounded exercise program that can be utilized as a means to improve health-related components of fitness in older adults. PMID- 15293054 TI - Changes in the angle-force curve of human elbow flexors following eccentric and isometric exercise. AB - The aim of this study was to explore and compare the magnitude and time-course of the shift in the angle-force curves obtained from maximal voluntary contractions of the elbow flexors, both before and 4 consecutive days after eccentric and isometric exercise. The maximal isometric force of the elbow flexors of fourteen young male volunteers was measured at five different elbow angles between 50 degrees and 160 degrees . Subjects were then divided into two groups: the eccentric group (ECC, n=7) and the isometric group (ISO, n=7). Subjects in the ECC group performed 50 maximal voluntary eccentric contractions of the elbow flexors on an isokinetic dynamometer (30 degrees x s(-1)), while subjects in the ISO group performed 50 maximal voluntary isometric muscle contractions with the elbow flexors at a lengthened position. Following the ECC and ISO exercise protocols, maximal isometric force at the five angles, muscle soreness, and the relaxed (RANG) and flexed (FANG) elbow angles were measured at 24 h intervals for 4 days. All results were presented as the mean and standard error, and a quadratic curve was used to model the maximal isometric force data obtained at the five elbow angles. This approach not only allowed us to mathematically describe the angle-force curves and estimate the peak force and optimum angle for peak force generation, but also enabled us to statistically compare the shift of the angle-force curves between and within groups. A large and persistent shift of the angle-force curve towards longer muscle lengths was observed 1 day after eccentric exercise ( P<0.01). This resulted in a approximately 16 degrees shift of the optimum angle for force generation, which remained unchanged for the whole observation period. A smaller but also persistent shift of the angle-force curve was seen after isometric exercise at long muscle length ( P<0.05; shift in optimum angle approximately 5 degrees ). ECC exercise caused more muscle damage than ISO exercise, as indicated by the greater changes in RANG and ratings of muscle soreness ( P<0.05). It was suggested that the shift in the angle-force curve was proportional to the degree of muscle damage and may be explained by the presence of overstretched sarcomeres that increased in series compliance of the muscle. PMID- 15293056 TI - Red blood cell membrane mechanical fluctuations in non-proliferative and proliferate diabetic retinopathy. AB - PURPOSE: To study whether cell membrane mechanical fluctuation (CMF) of red blood cells (RBCs) are attenuated in non-proliferative diabetic retinopathy (NPDR) and proliferative diabetic retinopathy (PDR). PATIENTS AND METHODS: Point dark-field microscopy-based recordings of local membrane displacements (frequency 0.3-25 Hz) were compared between type 2 diabetes patients with mild-to-moderate and severe NPDR and type 2 diabetes patients with PDR. The matched control group, corresponding to each stage of diabetic retinopathy, was based on non-diabetic patients who were evaluated in our clinic due to cataract. RESULTS: The average mean values of the maximal CMF amplitude did not differ between RBCs of NPDR patients (n=20) and controls (n=20) (19.5+/-1.5% and 19.6+/-1.7%, respectively). A statistically significant decrease in CMF amplitudes was observed in patients with PDR compared with patients with a non-proliferative disease (NPDR -20%, PDR 90%). CONCLUSION: This new rheological characteristic demonstrates differences in the mechanical properties of RBCs in different stages of diabetic retinopathy. The significant reduction in CMF in patients with PDR may shed more light on the possible mechanism modulating retinal ischemia and leading to angiogenesis in these patients. Larger-scale studies are needed to evaluate these findings and the possible correlation between significantly lower CMF values and the progression of diabetic retinopathy. PMID- 15293060 TI - UVA1-induced decrease in dermal neuron-specific enolase (NSE) in acrosclerosis. AB - Besides its role in small-cell carcinoma of the lung, elevated serum levels of neuron-specific enolase (NSE) have recently been reported to be associated with autoimmune rheumatic disorders such as systemic sclerosis. Serum NSE seems to correlate with disease activity as well as Rodnan skin score. The aim of the study was to assess the neuromodulatory effects of conventional UVA1 phototherapy on acrosclerosis as an additional mechanism besides an assumed T cell apoptosis, collagenase induction and angiogenesis. Punch skin biopsies of acrosclerotic skin lesions taken before and after treatment from four patients were evaluated immunohistochemically for the presence of NSE, S100 and neurofilament. Immunolabeling revealed a UVA-induced decrease in dermal NSE expression. In contrast, no alteration in neurofilament+ cells could be detected. In line with the findings of a previous investigation, a high number of S100+ cells were detected in most specimens. We demonstrated a UVA1-induced reduction in dermal NSE levels correlating with a softening of former sclerotic lesions. Even though the origin and the functional mechanisms remain obscure, NSE might be relevant directly within sclerotic skin lesions and may possibly be used as a diagnostic marker at least in SSc-associated acrosclerotic skin. PMID- 15293057 TI - Neuropeptide Y-evoked proliferation of retinal glial (Muller) cells. AB - BACKGROUND: Glial cells in human retinas and in fibrocellular membranes from patients with proliferative vitreoretinopathy (PVR) have been described to upregulate their expression of Y1 receptors for neuropeptide Y (NPY) (Soler et al.: Glia 39:320, 2002). However, it is unknown whether Y1 receptor activation causes proliferation of retinal glial cells. We investigated whether NPY exerts a proliferation-stimulating effect on retinal glial cells, and compared the NPY evoked signaling with the signaling of purinergic P2Y receptors. METHODS: Proliferation assays using bromodeoxyuridine were carried out on primarily cultured Muller glial cells of the guinea pig, in the absence and presence of blockers of Y1 receptors, of receptor tyrosine kinases (RTKs), of mitogen activated protein kinases (MAPKs) and of phosphatidylinositol-3 kinase (PI3K). RESULTS: NPY exerted a biphasic effect on Muller cell proliferation. At low concentrations (0.1 ng/ml and 1 ng/ml) it decreased the proliferation rate of the cells, while at higher concentration (100 ng/ml) it increased Muller cell proliferation. The NPY-evoked proliferation was mediated by Y1 receptor stimulation and by activation of the p44/p42 MAPKs and partially of the p38 MAPK. Moreover, Y1 receptor-induced activation of PI3K as well as transactivations of the platelet-derived and the epidermal growth factor RTKs were necessary for full mitogenic effect of NPY. Y1 and P2Y receptors share partially common signal transduction pathways in Muller cells. CONCLUSION: It is suggested that NPY may be involved in stimulation of retinal glial cell proliferation during PVR when it is released at higher amounts into the injured retina. PMID- 15293055 TI - Assembly of the phagocyte NADPH oxidase. AB - Stimulated phagocytes undergo a burst in respiration whereby molecular oxygen is converted to superoxide anion through the action of an NADPH-dependent oxidase. The multicomponent phagocyte oxidase is unassembled and inactive in resting cells but assembles at the plasma or phagosomal membrane upon phagocyte activation. Oxidase components include flavocytochrome b558, an integral membrane heterodimer comprised of gp91phox and p22phox, and three cytosolic proteins, p47phox, p67phox, and Rac1 or Rac2, depending on the species and phagocytic cell. In a sense, the phagocyte oxidase is spatially regulated, with critical elements segregated in the membrane and cytosol but ready to undergo nearly immediate assembly and activation in response to stimulation. To achieve such spatial regulation, the individual components in the resting phagocyte adopt conformations that mask potentially interactive structural domains that might mediate productive intermolecular associations and oxidase assembly. In response to stimulation, post-translational modifications of the oxidase components release these constraints and thereby render potential interfaces accessible and interactive, resulting in translocation of the cytosolic elements to the membrane where the functional oxidase is assembled and active. This review summarizes data on the structural features of the phagocyte oxidase components and on the agonist dependent conformational rearrangements that contribute to oxidase assembly and activation. PMID- 15293062 TI - The absence of early diarrhea with atropine premedication during irinotecan therapy in metastatic colorectal patients. PMID- 15293061 TI - The "strip" patch test: results of a multicentre study towards a standardization. AB - BACKGROUND: The "strip" patch test (SPT) is a variant of patch testing which is used for substances with a poor percutaneous penetration. Penetration of the substances is enhanced by repeated applications of adhesive tape prior to their application to the skin. However, no guidelines exist for standardized performance of the SPT. OBJECTIVES: The aim of this multicentre study was to obtain a first practical approach towards a standardized SPT procedure. METHODS: Intact noninflamed skin of the upper back of 83 healthy volunteers was tape stripped. For sequential strips, a 25-mm diameter 3M Blenderm surgical tape was vertically applied and gently pressed downward using the fingertips for about 2 s. The tape was removed in one quick movement at an angle of 45 degrees in the direction of adherence. Each strip was performed with a new piece of tape on exactly the same skin area. RESULTS. In each subject, we first determined the number of strips (A) until the skin surface started to glisten and calculated the median number of strips (A) in the sample (A=26 strips). We then ascertained the median number of strips (a) in the sample that was necessary to achieve a statistically significant and twofold increase in TEWL (a=11 strips), revealing a "critical" stratum corneum strip depth. The unknown number of strips (a) for each subject was finally calculated from the formula a/A=a/A, i.e. the individual number of strips (A) until the skin surface started to glisten was multiplied by a derived tape-specific correction factor (cf=a/A=11/26=0.4). The increase in percutaneous penetration in strip patch testing by performing "a" strips versus conventional patch testing was shown by scoring of clinical and subjective SLS irritant reactions. CONCLUSIONS: The present multicentre study outlines an experimentally derived approach for a uniform SPT procedure, which does not require the use of complex technical equipment. This first approach now requires validation by a study involving the application of allergens to obtain evidence of enhancement in the sensitivity of patch testing. PMID- 15293063 TI - Does the incidence of anal canal cancers differ in Moslem societies? PMID- 15293064 TI - Management of the pelvic recurrence of rectal cancer with radiofrequency thermoablation: a case report and review of the literature. AB - INTRODUCTION: The results of rectal cancer surgery are limited by the development of local recurrence (LR) that represents a great challenge to the surgeon. In the presence of unfavourable conditions for performing a curative operation, various forms of palliative treatment are indicated to control the patient's symptoms and the disease's complications. Recently, radiofrequency thermoablation (RFTA) has become a complimentary alternative therapy for malignant inoperable liver tumours. The present paper reports the use of RFTA in the management of pelvic recurrence of rectal adenocarcinoma. CASE REPORT: Fourteen months after abdominoperineal resection, a 32-year-old woman began to complain of progressive pelvic and lumbar pain. A large pelvic mass was found and serum CEA was elevated (66.4 ng/ml) at that time. Due to the dimensions of the presacral tumour (8 x 5 x 4 cm3) and the associated refractory pain, the patient underwent RFTA of the recurrent disease. Under epidural anaesthesia, a computed tomography-guided percutaneous needle electrode was introduced into the tumour. Although the procedure provided immediate pain control, the patient developed an intestinal obstruction 3 months later. This complication required surgical treatment to release adherences from the necrosed tumour. CONCLUSION: Apart from this complication, RFTA allowed prolonged relief of the pelvic pain and improved quality of life. Faced with an unresectable pelvic recurrence, RFTA proved to be a viable option for controlling pain, although a relatively high cost and eventual complications may limit its use. PMID- 15293065 TI - Palliative portal vein stent placement for lymphatic recurrence of gastric cancer. AB - INTRODUCTION: Percutaneous transhepatic stenting of the main portal vein is a rare intervention. CASE REPORT: In the current patient, percutaneous angioplasty and stenting of a main portal vein stenosis due to lymphatic recurrence of gastric cancer ameliorated the progressing therapeutic restriction. The wall stent achieved portal venous patency that enabled ongoing chemotherapy. The stent remained patent for the entire subsequent survival period. PMID- 15293066 TI - Dynamic graciloplasty for total anorectal reconstruction after abdominoperineal resection for rectal tumour. AB - AIM: To review the results of dynamic graciloplasty for total anorectal reconstruction after abdominoperineal resection (APR) for rectal cancer. PATIENTS AND METHODS: Chart reviews were done on 17 patients who had dynamic graciloplasty following abdominoperineal resection and details of post-operative complications, bowel functions and recurrences were obtained. RESULTS: Seventeen patients (12 males) had dynamic graciloplasty after APR for low rectal tumours. The median age was 58.5 years (range 33-78). Three patients from overseas were lost to follow up, and three still have not had the defunctioning stoma closed. Only 11 patients were available for evaluation of function. The median time from graciloplasty to continence to solids and liquids is 15.7 months (range 0.4-21.9 months). Six patients had defecatory problems, requiring daily irrigation to evacuate. Nine patients were continent without need for gracilis stimulation. Only two patients needed gracilis stimulation to maintain continence. Fifty percent of rectal carcinoma patients had developed a recurrence. CONCLUSION: Dynamic graciloplasty had a high morbidity and did not always bring about normal defecatory function. Gracilis stimulation was not needed to achieve continence in all cases. Conversely, dynamic graciloplasty may lead to defecatory difficulties in a large number of patients. Graciloplasty should only be considered three years after the initial APR to avoid performing the procedure in a patient who may develop recurrence as well as to select patients who are psychologically prepared for the surgery and its complications. PMID- 15293067 TI - Metabolic rates during rest and activity in differently tracheated spiders (Arachnida, Araneae): Pardosa lugubris (Lycosidae) and Marpissa muscosa (Salticidae). AB - With flow-through respirometry under video tracking, the CO(2) release of adult male and female Pardosa lugubris (wolf spider) and Marpissa muscosa (jumping spider) was measured during rest and activity. Activity metabolism was measured in phases in which the animals were spontaneously active and during forced exercise. Standard metabolic rates (V(CO2)/ t) were 1.43 nmol s(-1) g(-1) in M. muscosa and 1.7-1.8 nmol s(-1) g(-1) in P. lugubris. Egg production caused higher resting rates in females compared with the males in P. lugubris. Maximum mass specific CO(2) release, the additional amount of CO(2 )released after activity and the factorial aerobic scope were higher in M. muscosa. Additionally, half time recovery and the lag between end of activity and maximum CO(2) release were lower in the jumping spider. The results are consistent with the hypothesis that the well-developed tracheal system in jumping spiders increases the efficiency of the respiratory system in comparison with wolf spiders, which possess similarly developed lungs but only a simple tracheal system that is restricted to the opisthosoma. PMID- 15293068 TI - Endobronchial lipomatous hamartoma: CT and MR imaging features (2004:5b). PMID- 15293069 TI - A case of headache and concentration disorders (2004:8a). PMID- 15293072 TI - Prosthetic joint infections-towards an improved outcome. PMID- 15293073 TI - Open, randomized study to compare the immunogenicity and reactogenicity of an influenza split vaccine with an MF59-adjuvanted subunit vaccine and a virosome based subunit vaccine in elderly. AB - BACKGROUND: An open, randomized, multicenter study was carried out in elderly to compare the immunogenicity and reactogenicity of a conventional influenza split vaccine (SpV) with an MF59-adjuvanted subunit vaccine (aSuV) and a virosome-based subunit vaccine (vSuV) since earlier studies reported better immunogenicity for adjuvanted and virosome-based vaccines. PATIENTS AND METHODS: A total of 840 subjects, aged 60 years or more, who had not been vaccinated or diagnosed with influenza in the preceding season were investigated. Hemagglutination-inhibition antibody titers were measured, and signs and symptoms recorded. RESULTS: The three vaccines exceeded EU efficacy requirements for subjects aged older than 60 years and seroprotective levels (titers > 1:40) were equally maintained with the three vaccines during 8 months post vaccination. SpV was as immunogenic as aSuV for the A/H3N2 strain (p < 0.0001) and significantly more immunogenic than aSuV for A/H1N1 strain (p = 0.0006). SpV was as immunogenic as vSuV for all three strains and significantly more immunogenic than vSuV for the A/H1N1 strain (p < 0.0001). In terms of reactogenicity, aSuV showed a higher rate of solicited local signs and symptoms than SpV (p = 0.021) and vSuV (p = 0.046), respectively. Incidence of solicited general symptoms was comparable on all treatments. No serious adverse event related to vaccination was reported. CONCLUSION: These findings suggest that all three vaccines are highly immunogenic with an acceptable reactogenicity profile and that they are appropriate for use in elderly. PMID- 15293074 TI - Influenza-associated myositis in children. AB - BACKGROUND: Influenza-associated myositis (IAM) is an infrequent and poorly known complication of influenza virus infection in children. The aim of this study was to describe five cases of IAM and to review the literature on IAM in children. PATIENTS AND METHODS: We conducted a retrospective analysis of cases of IAM diagnosed at two university children's hospitals in Switzerland during two consecutive influenza seasons. Findings were compared with 39 individual case reports and five publications summarizing an additional 272 cases identified by a medical online library (MEDLINE) search. RESULTS: Overall, 316 cases were analyzed. IAM typically occurred in school-aged children with a 2:1 male predominance. Influenza B and A viruses were identified in 76% and 24% of cases, respectively. The median interval between onset of influenza and onset of IAM was 3 days (range 0-18). The calf muscles were involved alone or together with other muscle groups in 69% and 31% of cases, respectively. Blood creatine phosphokinase (CPK) concentration was invariably elevated. Median duration to clinical recovery was 3 days (range 1-30). Rhabdomyolysis occurred in ten of 316 patients (3%), was more common in girls (80%), more often associated with influenza A (86%), and led to renal failure in eight patients (80%). CONCLUSION: Clinical and laboratory findings of IAM are highly characteristic and allow a rapid diagnosis during the influenza season. PMID- 15293075 TI - A prediction model for community-acquired Chlamydia pneumoniae pneumonia in hospitalized patients. AB - BACKGROUND: The objective of the study was to identify factors that help to predict community-acquired Chlamydia pneumoniae pneumonia in hospitalized patients. PATIENTS AND METHODS: Clinical data of 83 patients with serologically confirmed C. pneumoniae pneumonia were compared with the data obtained from 72 patients with bacterial pneumonia. The criteria of bacterial pneumonia included positive blood and/or sputum cultures and negative serology for Mycoplasma pneumoniae, Chlamydia psittaci, Chlamydia pneumoniae and Coxiella burnetii. The data collected included demographics, chronic diseases, pre- and post hospitalization course of pneumonia, clinical data on admission and laboratory findings. Descriptive statistical analysis, involving numerous variables, was followed by univariate and multivariate logistic regression analysis. RESULTS: Two different situations, one including demographic data and information on the pre-hospitalization course of pneumonia, and another based on clinical information on admission and on laboratory results, were modeled using multivariate logistic regression. Several variables selected from these two models were incorporated into the third model, and the following four variables were found to have the highest predictive value of C. pneumoniae pneumonia: nursing home residence (odds ratio [OD] 3.73, 95% confidence interval [CI] 1.39 10.06), low c-reactive protein (CRP) levels (OD 5.99, 95% CI 1.82-19.67), nonproductive cough (OD 0.32, 95% CI 0.14-0.73), and a normal urinalysis (OD 0.38, 95% CI 0.17-0.83). CONCLUSION: Our findings seem to allow for a more reliable differentiation between C. pneumoniae pneumonia and other bacterial pneumonias, but further investigations will be needed to validate the proposed model. PMID- 15293076 TI - Outcomes of ventilated COPD patients with nosocomial tracheobronchitis: a case control study. AB - BACKGROUND: The aim of this study was to determine the impact of nosocomial tracheobronchitis (NTB) related to new bacteria on the outcome in patients with chronic obstructive pulmonary disease (COPD). PATIENTS AND METHODS: A prospective observational case-control study was conducted in medical COPD patients requiring intubation and mechanical ventilation for more than 48 hours. Patients with nosocomial pneumonia were excluded. Six matching criteria were used, including the duration of mechanical ventilation before NTB occurrence. RESULTS: 81 matched case-control pairs were studied. Although the mortality rate was similar (40% vs 34%; p = 0.48), median duration of mechanical ventilation (20 vs 12 days; p = 0.015) and intensive care unit (ICU) stay (25 vs 18 days; p = 0.022) were higher in cases than in controls. NTB was independently associated with a longer than median period of mechanical ventilation among case and control patients (OR = 4.7 [95%CI = 2-10.9]; p < 0.001). In cases with appropriate antibiotic treatment compared with those who did not receive antibiotics, a shorter median duration of mechanical ventilation (12 vs 23 days; p = 0.006) and ICU stay (16 vs 29 days; p = 0.029) were observed. CONCLUSION: NTB is associated with an increased duration of mechanical ventilation and ICU stays. Further studies are required to determine whether antibiotics could improve the outcome of patients with NTB. PMID- 15293077 TI - Major hypertriglyceridemia in HIV-infected patients on antiretroviral therapy: a role of the personal and family history. AB - BACKGROUND: Our aim was to identify factors predisposing HIV-infected patients on long-term antiretroviral therapy (ART) to major hypertriglyceridemia (HTG). PATIENTS AND METHODS: We conducted a retrospective, case-control study involving 76 HIV-infected patients with HTG, defined by 12-hour fasting plasma triglyceride (TG) > 4.5 mmol/l on at least one occasion, and 150 HIV-infected matched control patients with TG consistently below 1.8 mmol/l. RESULTS: Patients coinfected by the hepatitis C virus appeared to be protected from HTG. In addition to known predisposing factors for HTG in HIV-infected patients (ART and immune/viral status), patients with a history of excess body weight were twice as likely to have HTG (odds ratio [OR] 2.8, 95% confidence interval [CI]: 1.1-6.9); HTG was also more frequent in patients who had a first-degree relative with cardiovascular disease (CVD) or a major risk factor for CVD (OR = 3.6, CI: 1.3 9.9). CONCLUSION: By identifying subgroups of highly predisposed patients, appropriate lifestyle and dietary measures could be recommended on ART initiation. PMID- 15293078 TI - Management of infection associated with total hip arthroplasty according to a treatment algorithm. AB - BACKGROUND: An algorithm for the management of hip arthroplasty-associated infections was validated in a cohort study. PATIENTS: 60 patients with 63 episodes of total hip arthroplasty-associated infections observed from 1985 to 2001 were included. The treatment algorithm was based on the time of manifestation, pathogenesis, and condition of implant and soft tissue. Three treatment options were proposed, namely debridement with retention, one-stage and two-stage replacement. RESULTS: The median patients' age was 72 years, the median follow-up 28 months; 29% were early, 41% delayed, and 30% late infections, 57% of the infections were exogenously and 43% hematogenously acquired. The overall success rate for the first treatment attempt was 83% (52/63). Patients treated according to the algorithm had a better outcome than the others (44/50 = 88% vs 8/13 = 62%, Relative risk (RR) 0.31, 95% confidence interval (CI): 0.11-0.86, p < 0.03); those treated with adequate antimicrobial therapy had a better success rate (87% vs. 50%, p < 0.01). CONCLUSION: The proposed algorithm defines a rational surgical/antibiotic treatment strategy. PMID- 15293079 TI - Lipid lowering therapy with fluvastatin and pravastatin in patients with HIV infection and antiretroviral therapy: comparison of efficacy and interaction with indinavir. AB - BACKGROUND: Lipoprotein disorders in HIV-positive patients receiving highly active antiretroviral therapy (HAART) are becoming a major concern in HIV treatment, since there is growing evidence for an association between HAART induced hyperlipidemia and increased cardiovascular risk. Yet relatively few data are available on the possible interactions of HAART and treatment with statins. PATIENTS AND METHODS: In this prospective study, 25 HIV-positive, treatment experienced patients (five female, 20 male, all Caucasian) were treated with either fluvastatin or pravastatin. Total cholesterol, low density lipoprotein (LDL) and high density lipoprotein (HDL) levels, and serum triglycerides were determined at regular intervals, as well as therapeutic drug monitoring to assess possible drug interactions. RESULTS: In 13 pravastatin-treated patients, a decrease in total cholesterol levels (from 7.12 mmol/l to 6.29 mmol/l) after 12 weeks of therapy was seen. In 12 patients treated with fluvastatin, a permanent reduction of total cholesterol (from 6.46 mmol/l to 5.31 mmol/l) after 12 weeks was observed. The reduction of LDL levels was 30.2% in the fluvastatin group and 14.4% in the pravastatin group. In eight patients receiving an indinavir containing HAART, indinavir plasma levels were not significantly influenced. No effect on triglycerides or HDL was observed. CONCLUSION: Fluvastatin and pravastatin are efficient in lowering total and LDL cholesterol levels in HIV positive patients receiving HAART. Furthermore, no influence on indinavir plasma levels could be observed. Therefore, both compounds seem to be a viable treatment option in HAART-induced hypercholesterolemia. PMID- 15293080 TI - CAPNETZ-community-acquired pneumonia competence network. PMID- 15293081 TI - Chronic necrotizing pulmonary aspergillosis complicated by pneumothorax. AB - A 61-year-old man presented with left-sided pneumothorax. On the chest computed tomograghy (CT), severe bilateral emphysema and left-sided pleural thickening were seen. His pneumothorax was drained with a chest tube. Because of a persistent air leakage, video-thoracoscopic wedge-resection of the suspected fistula and muscle-sparing minithoracotomy with extensive wedge resections of the left upper lobe were performed. Biopsy specimens showed micronodular mycetomas with septate hyphae highly suggestive of Aspergillus. The fungus destructed the lung tissue without vessel invasion. The patient had not been taking immunosuppressant drugs and had no prior opportunistic infections. Itraconazole was begun, the lung was expanded and the patient recovered. We propose that extensive resection of affected lung tissue in combination with long-term antifungal therapy with itraconazole is a valuable therapeutic option in patients with a complicated course of chronic necrotizing pulmonary aspergillosis (CNPA). PMID- 15293082 TI - Mixed lung infection by Legionella pneumophila and Legionella gormanii detected by fluorescent in situ hybridization. AB - A mixed infection by Legionella pneumophila and a nonpneumophila Legionella species was detected in a lung biopsy specimen obtained from a patient with atypical pneumonia by fluorescent in situ hybridization (FISH). This result was confirmed by polymerase chain reaction (PCR). Sequencing of PCR products confirmed mixed infection by L. pneumophila and L. gormanii. Culture for Legionella spp. was negative and serology showed a rise only in IgG anti- Legionella pneumophila titer. To our knowledge, this is the first report of a mixed infection by L. pneumophila and a non-pneumophila Legionella species detected by FISH. Because FISH is a rapid and culture independent method that detects specific microorganisms in biopsy specimens it is recommended, in particular, for the detection of fastidious bacteria. PMID- 15293083 TI - Failure of ceftriaxone in an intravenous drug user with invasive infection due to Ralstonia pickettii. AB - We report a case of septic arthritis due to Ralstonia pickettii in an intravenous drug user with unfavorable clinical course under antibiotic therapy with ceftriaxone despite in vitro susceptibility to the drug. The treatment failure may have been due to a discrepancy between in vitro and in vivo susceptibility of R. pickettii, or to resistance development mediated by a recently described inducible beta-lactamase. PMID- 15293085 TI - What's new in HIV/AIDS. Protective immunity in HIV infection: where do we stand? AB - In the face of a global epidemic of HIV/AIDS with the majority of individuals living in the developing countries, a protective vaccine seem the foremost goal. As this is not within imminent reach, the other option would be immunotherapy that prolongs the asymptomatic phase of infection. The basis for such a therapy is to understand the correlates of protective immunity against HIV. Here we present the most recent advances regarding selected parts of the immune response towards HIV. PMID- 15293086 TI - Fibrodysplasia ossificans progressiva in a Malian boy of Bamako. AB - Fibrodysplasia ossificans progressiva (FOP) is an extremely rare hereditary connective tissue disorder characterized by congenital malformation of the great toes and by progressive heterotopic ossification of striated muscles and soft tissues. We report a case of FOP in a Malian boy and review the clinical and radiographic manifestations of this disorder. The body TDM showed ossifications and calcifications in the muscles of the large rhomboid, the erector muscles of the rachis and the trapezius muscles. Radiography of the dorsolumbar rachis showed paravertebral soft tissue calcification adjacent to intact lumbar vertebrae. The diagnosis is based on clinical and radiological findings and demonstration of skeletal malformations. The differential diagnosis of this rare condition from other muscle and joint disease is discussed. There is no effective prevention or treatment. There is a need for a wider knowledge of this condition. PMID- 15293087 TI - Contrast-enhanced power Doppler sonography of knee synovitis in rheumatoid arthritis: assessment of therapeutic response. AB - The aim of this study was to evaluate the ability of power Doppler sonography (PDS) with ultrasound contrast agent to assess the synovial perfusion changes induced by intra-articular steroid injection therapy in the knee joints of patients with rheumatoid arthritis (RA). Eighteen RA patients (16 women, 2 men) with a history and signs of active knee synovitis were studied. Tenderness was evaluated using Thompson's modified index of synovitis activity. All patients underwent joint aspiration followed by intra-articular injection of 40 mg of triamcinolone hexacetonide. Gray-scale ultrasonography and PDS with an intravenous ultrasound contrast agent (Levovist) examinations were carried out before and 3 weeks after the intra-articular steroid injection. The calculation of the time--intensity curves provided a quantitative estimation of the synovial perfusion. The median values of the index of synovitis activity decreased significantly from 7.0 (95% confidence interval (CI) 6.0-8.0) to 3.0 (95% CI 2.0 4.0) ( p<0.01) 3 weeks after the intra-articular steroid injection. All patients showed a reduction of PDS signal after intra-articular steroid therapy and the baseline and follow up median values of the area underlying time-intensity curves were 7.48 (95% CI 5.79-8.73) and 2.45 (95% CI 1.92-3.61), respectively. The comparison between baseline and follow-up median values of the area under the curves showed a statistically significant reduction of PDS findings ( p<0.01). At follow-up examinations the changes in the index score of the synovitis activity were significantly correlated to the changes in the values of the area underlying time-intensity curves ( r=0.785; p<0.01). A significant correlation was also observed between baseline values of the area underlying time-intensity curves and C-reactive protein (CRP) ( r=0.548; p=0.023). In conclusion, PDS with an intravenous ultrasound contrast agent has been shown to be able to detect changes in synovial perfusion after intra-articular steroid injection and may be an additional useful method in the evaluation of synovial inflammation and in the assessment of the therapeutic response. PMID- 15293088 TI - Systematic review of primary total elbow prostheses used for the rheumatoid elbow. AB - Total elbow prosthesis (TEP) has been shown to be a viable option for treatment of the rheumatoid elbow. Many types of TEP have been studied, but the heterogeneity of the studies makes most conclusions subject to discussion. The aim of this systematic review is to show the differences between the most commonly used TEP for the destroyed rheumatoid elbow. After a search in Pubmed (NLM, Bethesda, USA) the senior author selected eight frequently used TEP: the Capitellocondylar, Coonrad-Morrey, GSB III, Kudo, Liverpool, Norway, Roper-Tuke and Souter-Strathclyde. For inclusion studies we arbitrarily formulated nine criteria, after which clearly adverse events were defined for comparison purposes. The Capitellocondylar and Souter-Strathclyde prostheses are the most studied treatments for replacing the rheumatoid elbow. In contrast to the Capitellocondylar, the Souter-Strathclyde prosthesis showed higher loosening rates but implemented modifications of the design have reduced these rates in recent studies. Nevertheless, in relation to most other joint replacements in rheumatoid patients, all TEP still have higher complication rates. For this reason an elbow prosthesis may just be warranted in seriously disabled patients. PMID- 15293089 TI - Associations between walking time, quadriceps muscle strength and cardiovascular capacity in patients with rheumatoid arthritis and ankylosing spondylitis. AB - The aim of this study was to examine whether there are any associations between walking time, quadriceps muscle strength and cardiovascular capacity in patients with rheumatoid arthritis (RA) and ankylosing spondylitis (AS). Thirty-one patients with RA and 26 patients with AS belonging to Steinbrocker's functional class I-II were examined. Cardiovascular capacity was calculated from the expired air during a bicycle test and quadriceps muscle strength by the peak torque from an isokinetic dynamometer test. Walking time was the time it took to walk a distance of 160 m on a flat floor and to climb up and down a staircase. In patients with RA, flat floor walking and stair climbing times correlated inversely with quadriceps muscle strength and cardiovascular capacity. Similar results were seen in patients with AS, although the association between cardiovascular capacity and stair-climbing time was not statistically significant. Multiple regression analysis was performed for all patients with quadriceps muscle strength and cardiovascular capacity applied as independent variables in two separate models. Cardiovascular capacity explained 32% and quadriceps muscle strength 21% of the variance in flat floor walking time. Quadriceps muscle strength, together with diagnosis and age, explained 38% of the variance in stair-climbing time, and cardiovascular capacity together with age and pain explained 36% of the variance. In conclusion, in spite of cardiovascular capacity and quadriceps muscle strength being associated with walking times, the findings suggest that they play only a modest role in explaining rapid walking on flat floor and in stairs. PMID- 15293090 TI - Experience with azathioprine in systemic sclerosis associated with interstitial lung disease. AB - The aim of this study was to evaluate the safety and efficacy of azathioprine in the treatment of interstitial lung disease (ILD) associated with systemic sclerosis (SSc). The records of patients with SSc with ILD who were treated with azathioprine were reviewed. Patients were treated with azathioprine and low-dose prednisone if they had progressive pulmonary symptoms (deterioration in the dyspnea score) or poor or deteriorating lung function. Response was classified as improved if the FVC increased more than 10% from baseline, and stable if it remained within 10% of baseline. Serial dyspnea scores were recorded. Eleven patients were treated with azathioprine, three of whom received treatment for 6 months or less owing to adverse effects (nausea, leukopenia and pulmonary tuberculosis in one patient each). The remaining eight patients received at least 12 months' treatment and the results suggested an improvement in the mean percent predicted FVC from a baseline value of 54.25+/-3.53 to 63.38+/-6.15 after 12 months ( p=0.101). Overall, five patients improved and three remained stable. The mean dyspnea score ( n=8) improved from a baseline of 1.55+/-0.19 to 0.50+/-0.19 at 12 months ( p=0.011). This is the first case series of patients with SSc associated ILD treated with azathioprine. Our results suggest that azathioprine may have a role in stabilizing lung function and improving symptoms in SSc, although this needs confirmation by a randomized controlled trial. PMID- 15293091 TI - Long-term follow-up of vertebral osteoporotic fractures treated by percutaneous vertebroplasty. AB - The aim of this study was: to assess the long-term efficacy and safety of percutaneous vertebroplasty (PVP) for treating painful vertebral osteoporotic fractures, and to estimate the risk of vertebral fracture in the vicinity of a cemented vertebra. A prospective open study was conducted. PVP were carried out between July 1995 and September 2000 for 16 patients with symptomatic osteoporotic vertebral fracture that had not responded to extensive conservative medical therapy. All the patients were followed-up for more than 1 year. The efficacy of the PVP was assessed by the changes over time in pain on Huskisson's visual analog scale (VAS) and on the McGill-Melzack scoring system (MGM). The efficacy of the procedure was also assessed by measuring the changes over time in quality of life assessed by the Nottingham Health Profile (NHP instrument): twenty-one vertebrae treated by PVP in 16 patients were evaluated. The mean duration of follow-up was 35 months. Pain assessed by the VAS significantly decreased from a mean of 71.4 mm+/-13 before PVP to 36 mm+/-30 after 6 months, and to 39 mm+/-33 at the time of maximal follow-up ( p<0.05 for both comparisons). The results were also significant for the MGM: 3.00+/-0.57 before PVP to 1.6+/-1.4 at the long-term follow-up ( p<0.05). The solely statistically significant decrease for quality of life was noted for pain. A slight but not significant improvement was noted for 3/6 dimensions of the NHP scores. A slight but significant increase in social isolation was also found. No severe complication occurred immediately after PVP. At the long term follow-up (35 months) there was a slight but not significantly increased risk of vertebral fracture in the vicinity of a cemented vertebra: odds ratio 3.18 (95% confidence interval (CI) 0.51-19.64). The odds ratio of a vertebral fracture in the vicinity of an uncemented fractured vertebra was 2.14 (95% CI: 0.17-26.31). In conclusion, PVP appears to be safe and effective for treating persistent painful osteoporotic fractures. Controlled studies with long-term follow-up are needed to evaluate the risk of vertebral fractures in the vicinity of a cemented vertebra. PMID- 15293092 TI - Retrospective analysis of the renal outcome of pediatric lupus nephritis. AB - The aim of this study was to analyze the renal outcome of pediatric lupus nephritis in the past two decades. We retrospectively reviewed the medical records of patients who fulfilled the 1987 American Rheumatism Association revised criteria for systemic lupus erythematosus who were followed up at the National Taiwan University Hospital between 1980 and 2001. All new patients who were under 18 years of age at the time of diagnosis were enrolled and were followed up until death, loss to follow-up, or till the end of 2002. The response to the treatment and renal outcome were analyzed. Seventy-two children (64 girls and 8 boys) were enrolled in the study. The mean age at diagnosis was 13.93+/ 0.35 years (mean +/- SEM). The mean duration of follow-up was 7.12+/-0.51 years. The 5-year renal survival rate (survival without dialysis or transplantation) was 63.13% and the 10-year survival rate was 53.54%. It was significantly better in patients receiving cyclophosphamide (CYC) pulse therapy. The 5-year survival rate for these patients was 87.82% and the 10-year survival rate was 81.06%. The renal survival curve was better in the CYC pulse therapy group than in the no CYC pulse therapy group, with p=0.0022. The duration between the diagnosis of lupus nephritis and end-stage renal disease (ESRD) was significantly longer in the CYC group (9.66+/-1.32 yrs) than in the no CYC group (3.24+/-0.94 yers), p=0.036. In the multivariate analysis, risk factors of developing ESRD were failure to achieve complete remission, higher serum creatinine at the initiation of treatment, and not receiving CYC pulse therapy. The renal survival was significantly better in the CYC pulse therapy group. The CYC pulse therapy was recommended in pediatric lupus nephritis patients and every effort should be made to achieve complete remission. PMID- 15293094 TI - Hypertrophic pulmonary osteoarthropathy: control of pain and symptoms with pamidronate. AB - This case presents a patient with hypertrophic osteoarthropathy of the lower extremities that developed secondary to congenital cyanotic heart disease. The major clinical manifestation was severe bilateral leg pain. The pain that was debilitating in nature completely resolved following a single administration of 60 mg pamidronate. Hypertrophic osteoarthropathy (HOA) is an acquired, uncommon disorder of obscure etiology. It has been described mainly in association with chronic suppurative pulmonary diseases, bronchogenic carcinoma and lung metastases, cystic fibrosis, and cyanotic congenital malformations of the heart. PMID- 15293093 TI - Bone status in rheumatoid arthritis assessed at peripheral sites by three different quantitative ultrasound devices. AB - Rheumatoid arthritis (RA) is characterized by periarticular and generalized loss of bone mass. Quantitative ultrasound (QUS) has been introduced as a method for the assessment of bone status and fracture risk. In this cross-sectional study bone status was assessed by QUS at different peripheral sites in 27 women with RA (mean disease duration 15 years) and in 36 healthy women matched for age, height and weight. Speed of sound (SOS, m/s), broadband ultrasound attenuation (BUA, dB/MHz) and stiffness of the calcaneus were assessed by a Lunar Achilles device. Amplitude-dependent SOS (Ad-SOS, m/s) of the second to fifth phalanx was measured by a DBM Sonic 1200, and SOS of the distal forearm and third phalanx was measured by a Omnisense multisite scanner. Bone mass (g/cm2 or g) of the hip, spine, distal forearm and total body was measured by dual-energy X-ray absorptiometry. QUS values were significantly reduced in RA at most sites ( p<0.005-0.001), but between-group differences were small, and large overlaps between the groups were noticed. After correction for bone mass, the observed differences remained statistically significant for the calcaneus and distal radius ( p<0.05). Independent associations between ultrasound measures and markers of disease activity were not demonstrated. In conclusion, bone status as assessed by QUS was compromised in RA, but whether ultrasound transmission may serve as a marker of disease progression and fracture risk in the individual patient remains to be clarified in prospective studies. PMID- 15293095 TI - Continuity of cytokine activation in patients with familial Mediterranean fever. AB - Familial Mediterranean fever (FMF) is a recessively inherited inflammatory disorder, characterized by recurrent attacks of fever and polyserositis. It has been considered that miscellaneous cytokines take part in the pathogenesis of the disease. The aim of this study was to investigate serum levels of soluble interleukin-2 receptor (sIL-2R), interleukin-6 (IL-6), and interleukin-10 (IL-10) in patients with FMF. The study included 42 patients with FMF (3 females, 39 males, mean age: 24.43 years) and 20 healthy volunteers as the control group (18 males, 2 females, mean age: 23.2 years). The patients were chosen according to Eliakim criteria. After recording their history and performing an examination, leukocyte counts, erythrocyte sedimentation rates (ESR), C-reactive protein (CRP), fibrinogen, sIL-2R, IL-6, and IL-10 levels were measured before and during attacks. A significant increase was found in leukocyte ( p<0.001), ESR ( p<0.001), CRP ( p<0.001), and fibrinogen ( p<0.001) levels of the patient group in the attack period compared to those in the quiescent state. sIL-2R ( p=0.019) and IL-6 ( p<0.001) levels showed significant increases during attacks compared to the levels before an attack. There was no significant difference between IL-10 levels. The levels of the three cytokines were significantly high both before and during the attacks compared to the control group. As a result, the elevation of sIL-2R and IL-6 levels both before and during the attacks compared to control group suggests the existence of continuous cytokine activation in the patients. No significant increase in the IL-10 levels in spite of the significant rise of sIL-2R and IL-6 during attacks supports the notion of inflammation and also reveals that compensation by anti-inflammatory IL-10 does not seem to occur. PMID- 15293096 TI - Mutational analysis of serotonin receptor genes: HTR3A and HTR3B in fibromyalgia patients. AB - The neurotransmitter serotonin (5-hydroxytryptamine, 5-HT) has been implicated in numerous human disorders. Dysfunction of serotonergic neurotransmission is thought to play a major role in the pathophysiology of the fibromyalgia syndrome (FMS) which is characterised by non-restorative sleep and severe pain. In our study, both serotonin receptor subunit genes, HTR3A and HTR3B, have been investigated for sequence variations in FMS patients in order to reveal a possible involvement in the aetiology of FMS. We examined DNA samples from 48 patients with FMS representing sporadic cases by single-strand conformation polymorphism (SSCP) and denaturing high-performance liquid chromatography (dHPLC) analysis, sequenced samples with conspicuous patterns and performed statistical calculations. HTR3A mutational analysis revealed one novel as well as five known sequence variations. Investigating HTR3B, we detected seven formerly described mutations and one novel sequence variant. Statistical computation rated all variants as probably non-disease-related polymorphisms. Nevertheless, one might speculate about an effect of the respective sequence variants on the severity of the disease. Sequence variants of the serotonin receptor subunit genes HTR3A and HTR3B indicate no obvious significance in the aetiology of fibromyalgia, yet they represent the basis for future studies on their pharmacogenetic relevance. PMID- 15293097 TI - Improvement of skin sclerosis after occurrence of anticentromere antibody in a patient with diffuse cutaneous systemic sclerosis. AB - A 38-year-old woman had noticed sclerodactylia and Raynaud's phenomenon 10 months before consultation. She was diagnosed as having systemic sclerosis (SSc) based on the skin sclerosis of her arms, chest, and face. Antinuclear antibody (ANA) level was 1:1280 with a speckled pattern, but specific autoantibodies were negative. Following the treatment with oral prednisolone and D-penicillamine, her skin sclerosis gradually improved. Three months after initiation of prednisolone, she presented Pneumocystis carinii pneumonia. About 1 year after the first admission, the pattern of indirect immunofluorescence staining changed from the speckled pattern to the discrete speckled pattern, and simultaneously anticentromere antibody (ACA) was detected by enzyme-linked immunosorbent assay. Her skin sclerosis rapidly and remarkably improved after appearance of ACA. It is generally considered that once certain SSc-specific autoantibody occurs, it does not disappear and change into other specificity of autoantibody thereafter. This case suggests that the presence of ACA closely correlates with clinical features and also suggests that clinical features may change during the clinical course with the appearance of another specific ANA. This case is very rare because such a case was not reported previously. PMID- 15293098 TI - Mucosa-associated lymphoid tissue lymphoma (MALT) of salivary glands and scleroderma: a case report. AB - The connection between scleroderma and lymphoma is uncommon and its pathogenic relationship is a much debated subject. We describe the case of a patient with mucosa-associated lymphoid tissue lymphoma (MALT) of both parotid glands without clinical signs of Sjogren's syndrome who simultaneously developed scleroderma. Independently of the pathogenic mechanism of these two diseases, it seems very important to emphasize that scleroderma may be the first manifestation of lymphoma. PMID- 15293099 TI - Systemic lupus erythematosus associated with recurrent lupus enteritis and peritonitis. AB - We describe the case of a 41-year-old woman with systemic lupus erythematosus (SLE) who suffered from repeated reversible lupus enteritis characterized by marked edematous thickening of the small intestine. Ultrasonography (US) and computed tomography (CT) manifested as an 'accordion-like appearance' and a 'target-like appearance', respectively. Resolution of gastrointestinal tract wall thickening was observed on follow-up US performed a week after the increase in predinosolone (PSL). We conclude that careful evaluation of sonographic and radiographic findings helps to establish the diagnosis of lupus enteritis. PMID- 15293100 TI - A case of anti-Jo1 myositis with pleural effusions and pericardial tamponade developing after exposure to a fermented Kombucha beverage. AB - The pathogenesis of the idiopathic inflammatory myopathies has been postulated to be an environmental trigger causing the expression of the disease in a genetically predisposed patient. We report a case of anti-Jo1 antibody-positive myositis which was associated with pleural effusions, pericardial effusion with tamponade, and 'mechanic's hands', probably related to the consumption of a fermented Kombucha beverage. Kombucha 'mushroom', a symbiosis of yeast and bacteria, is postulated to be the trigger for our patient's disease owing to the proximity of his symptoms to the consumption of the Kombucha beverage. PMID- 15293101 TI - Giant geode at the olecranon in the rheumatoid elbow--two case reports. AB - A single giant geode at the olecranon in a patient with rheumatoid arthritis (RA) is relatively rare, and may cause diagnostic difficulties or cause a spontaneous pathological fracture owing to weakness of the cortical bone associated with osteoporosis. We report two cases of patients presenting with single giant geodes at the olecranon. In one case we performed an open reduction and internal fixation with bone grafting for a pathological fracture due to the geode. In the other case we performed curettage of the geode with bone grafting to prevent a pathological fracture, and a synovectomy of the elbow. We suggest that the presence of a giant geode at the olecranon may necessitate surgical intervention to prevent the occurrence of a spontaneous pathological fracture. PMID- 15293102 TI - Lupus anticoagulant, factor V Leiden, and methylenetetrahydrofolate reductase gene mutation in a lupus patient with cerebral venous thrombosis. AB - We describe the case of a young Lebanese woman with systemic lupus erythematosus (SLE) and a positive lupus anticoagulant (LAC) who developed right internal jugular vein and sigmoid sinus thrombosis. Coagulation studies showed that in addition to the LAC the patient was heterozygous for the factor V (FV) Leiden mutation, and C677T mutation of the methylenetetrahydrofolate reductase gene. The high prevalence of FV Leiden in the eastern Mediterranean region suggests that we should probably screen our SLE patients in this area, especially those with anticardiolipin antibodies and/or LAC who have no history of thrombosis, for this and other thrombophilia markers. The detection of such abnormalities may have major practical consequences for the long-term management of these patients to prevent further thrombotic episodes. PMID- 15293103 TI - Neuro-Behcet's disease with chorea after remission of intestinal Behcet's disease. AB - Neuro-Behcet's disease shows various neuropsychiatric symptoms, but chorea has rarely been reported. We report a case of neuro-Behcet's disease in a 67-year-old woman with depression and chorea that occurred 22 years after the onset of intestinal Behcet's disease. Brain magnetic resonance imaging (MRI) using a fluid attenuated inversion-recovery (FLAIR) sequence demonstrated lesions more clearly than did T2-weighted MRI. Some of the lesions appeared as small ring-like foci, i.e. low-intensity spots rimmed with remarkable hyperintense signals, in the periventricular white matter and basal ganglia. A review of the literature revealed that the onset of chorea in cases of Behcet's disease varied from the time of onset of Behcet's disease to 31 years after onset of the disease. Psychiatric manifestations have often been associated with neuro-Behcet's disease. In the present patient, treatment with prednisolone resolved the chorea, suggesting that the chorea was caused by an autoimmune mechanism. It seems likely that the long-term development of vasculitis in patients with Behcet's disease results in the formation of these particular brain lesions on FLAIR MR images. Chorea should be taken into consideration as one of the manifestations of Behcet's disease, even many years after remission of the disease. PMID- 15293104 TI - Pure red cell aplasia and adult-onset Still's disease. AB - Pure red cell aplasia (PRCA) associated with adult-onset Still's disease (AOSD) is very rare. In this report a 28-year-old woman was admitted with fever, skin rash, jaundice and anemia. She was diagnosed as having AOSD with PRCA by bone marrow examination. Treatment with high-dose prednisolone and intravenous immunoglobulin resulted in remission of the PRCA and a good response of the AOSD. PMID- 15293105 TI - Antiphospholipid and antisynthetase syndrome in a patient with polymyositis rheumatoid arthritis overlap. AB - Idiopathic inflammatory myopathies, mainly polymyositis (PM) may occur in the course of several autoimmune diseases. The overlapping forms of myositis, when the patient also meets the criteria for rheumatoid arthritis (RA), affect 3%-5% of myositis patients [1]. To the best of our knowledge this is the first report on the overlapping form of RA, antiphospholipid syndrome (APS) and the serological subgroup of PM, the antisynthetase syndrome (ASS). PMID- 15293106 TI - Depigmentation--a rare side effect to intra-articular glucocorticoid treatment. AB - Intra-articular glucocorticoid injections are often used in patients with rheumatoid arthritis, either as bridging therapy or in periods with increased disease activity. We present a case of local skin depigmentation that occurred at the site of injection in a dark-skinned patient. Depigmentation is a rare complication of such therapy but may have important implications for dark-skinned patients. PMID- 15293107 TI - Yellow nail syndrome in rheumatoid arthritis: a drug-induced disease? PMID- 15293108 TI - Mycobacterium bovis and septic glenohumeral arthritis. PMID- 15293109 TI - Monostotic Paget's disease of the tibia in Korea. AB - The incidence of Paget's disease has been estimated to be about 3%, but it is extremely rare in Asia, especially in Korea. In addition, monostotic involvement seems to be far less frequent. In this report, we describe a case of monostotic Paget's disease localized in the right tibia. PMID- 15293111 TI - Inguinal herniation in the adult, defect or disease: a surgeon's odyssey. PMID- 15293112 TI - Herniosis: a biological approach. PMID- 15293113 TI - The unified theory of hernia formation. AB - The perusal of surgical journals suggests that the etiology and the treatment of hernias are still based on the understanding of a simple mechanical defect, an idiopathic happenstance requiring a reliable hernia repair, preferably with a prosthetic mesh or device. The need for additional elucidation does not constitute an aim that is pervasive in the surgical community or with the corporate manufacturers of surgical implements. This may well be because surgeons are not trained scientists and laboratory workers. Fortunately, several disciplines are injecting a healthy dose of curiosity matched by ingenuity. Among these contributors, we can count anatomists, electron microscopists, biochemists, organic chemists, pathologists, geneticists, and molecular biologists, who have looked at collagen, enzymes, tobacco smoke, congenital diseases, and chromosomal defects. Every aspect of the researchers' work has identified and converged onto a final common organ: collagen. It is the pathological changes in collagen that set the stage for the development of a hernia. The multiple theories on mechanisms of hernia formation have, at last, melded into one single Unified Theory of hernia formation. PMID- 15293114 TI - Parailiac hernia repair. PMID- 15293115 TI - The concept of bimaxillary transverse osteodistraction: a paradigm shift? AB - Severe crowding due to narrow upper and lower apical bases can be corrected by the extraction of four premolars, or by bimaxillary transverse osteodistraction. The first strategy is prone to unaesthetic changes in lip posture, nasolabial angle and buccal corridors. Life-long retention is necessary because of the known correlation between increased intercanine distance and relapse of crowding. The second strategy involves surgery and the final outcome regarding stability is not yet known. Theoretically, because the canines have not been moved outside of the skeletal envelope, and because the functional matrix positively influences the dental arches, relapse of crowding should be less. Facial appearance is improved because of the reduction of the buccal corridors and the fullness of the mouth both at rest, and upon smiling. PMID- 15293116 TI - [Recurrent tumors in the oral and maxillofacial region. Results and treatment strategies in 20 years]. AB - SUBJECT MATTER: Following clinical diagnosis of a recurrent tumor, curative treatment is seldom available. Depending on the size of the recurrent tumor and the patient's general health condition, extensive surgical resections and reconstructions are avoided in favor of nonsurgical treatment modalities with palliative intent. According to the literature, location of the tumor, tumor size and R-1 and R-2 resection rates are the most frequent reasons for the development of recurrent tumors. PATIENTS AND METHODS: In a retrospective evaluation, a population of 1000 patients who had been treated for primary head and neck cancer during the period from 1979 to 1996 were analyzed descriptively. Survival probabilities of patients with recurrent tumors were calculated according to the Kaplan-Meier product-limit method, and different treatment concepts were compared and analyzed with the log-rank test for significant differences. RESULTS: The largest proportion of primary tumors involved the floor of the mouth (n=369, 36.9%). A total of 198 patients (19.8%) developed recurrent cancer; 79.8% of patients experienced recurrent cancer within 2 years following primary treatment. Within the group of T1/T2 tumors the incidence of recurrent tumors was 28.9%, whereas the incidence in the T3/T4 group was 44.6%. Tumor infiltration of the resection margins was detected in 12.9%. CONCLUSIONS: In line with the literature, tumor infiltration of the resection margins is a relevant prognostic factor; therefore, intraoperative frozen section must be recommended. Treatment with curative intention, in particular extensive surgical resections, is seldom possible and always requires a very intensive discussion with the patient. PMID- 15293117 TI - [Chorioallantoic membrane of fertilized avian eggs as a substrate for assessment of cancerous invasiveness]. AB - PURPOSE: Invasiveness is a characteristic feature of malignant tumors considerably determining the prognosis of affected patients. For assessment, apart from in vitro procedures with limited validity, tests on animal models have been established which certainly should be replaced by alternative methods whenever possible. The chorioallantoic membrane (CAM) of fertilized avian eggs represents an epithelial-lined membrane composed of all three blastodermic germ layers. In an "in ovo" assay cancer cells can be applied to this membrane after sinking (CAM assay). Tumor growth and invasiveness should be monitored in succession. MATERIAL AND METHODS: Hybrid chorionic carcinoma trophoblast cells were expanded in cell culture and spread over the CAM of hen's eggs after sinking followed by further incubation at 37 degrees C. The growth and development of the tumors were assessed macroscopically and finally (immuno-)histologically. Additionally, cytokeratin 19 was determined by enzyme-linked immunosorbent assay following homogenization of the tumor cells. RESULTS. Macroscopically, development of solid tumors was evident. Histological and immunohistochemical analysis revealed initial intraepithelial followed by cone-shaped infiltration of the CAM by the tumor cells. Tumor growth could be correlated with quantitative cytokeratin 19 measurements. CONCLUSIONS: Histomorphological appearance of the tumors was comparable with those results achieved in an immunodeficient mouse model. In addition, the CAM assay can be used for qualitative assessment of invasiveness of malignant tumors and yields quantitative results regarding growth kinetics. In contrast to conventional animal models, there is no need for official approval. Finally, this method is economical and facilitates processing many cases within a short time. PMID- 15293118 TI - [Noninvasive brush biopsy as an innovative tool for early detection of oral carcinomas]. AB - PURPOSE: The aim of this prospective study was to investigate the diagnostic accuracy of DNA image cytometry in combination with non-invasive brush biopsies taken from suspicious oral lesions. MATERIAL AND METHODS: Cytological diagnoses obtained from 1328 exfoliative smears of 332 different lesions were compared with histology and/or clinical follow-ups of the respective patients. Additionally, nuclear DNA contents were measured after Feulgen restaining using a TV image analysis system. DNA aneuploidy was assumed if abnormal DNA stemlines or cells with DNA content greater than 9c were observed. RESULTS; The sensitivity of our cytological diagnosis in addition to DNA image cytometry on oral smears for the detection of cancer cells was 97.8%, specificity 100%, positive predictive value 100%, and negative predictive value 98.1%. CONCLUSION: The application of DNA image cytometry with DNA aneuploidy as a marker for neoplastic transformation in oral smears secures cytologic diagnosis of carcinomas. Smears from brushings of all visible oral lesions are an easily practicable, cheap, noninvasive, painless, and safe screening method for detection of oral precancerous lesions and squamous cell carcinoma in all stages. We conclude that DNA image cytometry is a very sensitive and highly specific, objective, and reproducible adjuvant tool for identification of neoplastic cells in oral smears. PMID- 15293119 TI - [Stress tests of reconstruction plates for bridging mandibular angle defects]. AB - AIM: The aim of the present study was to evaluate the mechanical stress in reconstruction plates used for bridging mandibular angle defects and in the screw plate-bone interface with the finite element method. Additionally, the influence of reconstruction plate geometry, screw configuration, and screw diameter upon the mechanical stress distribution was determined. Suggestions for design improvements of the plate were derived from the results. MATERIAL AND METHODS: Based on the geometrical data of a human mandible, an angle defect bridged by a titanium reconstruction plate was generated and exposed to chewing force. The reconstruction plate was securely fixed by M 2.7 titanium screws. A variation of plate design, screw configuration, and screw diameter was carried out. The mechanical stress was calculated following the von Mises stress hypothesis. RESULTS: Using the standard plate the mechanical stress in all components exceeded by far the ultimate tensile strength. Possible clinical consequences could be a fatigue fracture of the plate, loosening of the screw, and irreversible damage of the bone leading to infection. Increasing the screw diameter by 50% would lead to a decrease of the mechanical stress by far more than 50%. An increase of the interface area between bone and plate and a triangular screw configuration diminishes the mechanical stress further, which may consequently allow a reduction of plate thickness with better adaptation to the actual jaw geometry. CONCLUSION: As a preliminary result the reconstruction plate could be thinned out in areas subject to less mechanical load. PMID- 15293120 TI - [Sonography as a training tool for screening of dubious midfacial fractures]. AB - BACKGROUND: In uncertain midfacial fractures, sonography is an alternative first line imaging modality to conventional radiographs. Patients with sonographically confirmed fractures can then be directly admitted to three-dimensional imaging, resulting in decreased radiation exposure since the conventional radiographs are omitted. MATERIAL AND METHODS: Using a high-frequency linear and curved array scanner in a healthy proband, images of the zygomatic arch, anterior maxillary sinus wall, infraorbital rim, and lateral orbital wall were obtained. For identification and anatomical allocation corresponding navigated ultrasound images of a reference skull were generated and fused with a segmented CT data set. Navigated sonography was reproduced in a patient with orbitozygomatical fracture of the left side. Therefore, the CT data set, performed during preoperative diagnostics, was fused with the ultrasound images. RESULTS: Because of different coupling shapes, the high-frequency linear array scanner was subjectively found to be more suitable for sonography in the field of the zygomatic arch, anterior maxillary sinus wall, and infraorbital rim, and the curved array scanner was better suited for transbulbar sonography of the orbital walls. After coupling sonography with the navigation system and referencing the scanner, it was possible to verify ultrasound findings objectively by navigation of the scanner and fusion with the CT data set. Using the reference skull, ultrasound images corresponding to normal findings were obtained and with the fused CT data, providing colored segmentation of the facial bones, an anatomically correct identification was possible. Clinical application of this tool is described in a patient with left-sided orbitozygomatical fracture. CONCLUSION: By fusion of ultrasound images and corresponding CT data with the help of a navigation system, a sonographic training tool for preliminary evaluation of midfacial fractures is available. PMID- 15293121 TI - [Experimental pilot study on surface activation of implants with liposomal vectors]. AB - BACKGROUND: Surface coating with mitogenic or morphogenic proteins can improve the healing of bone adjacent to implants and increase the bone-implant interface. Clinical surveys have shown liposome-mediated gene transfer to be a promising and safe new therapeutic method. The aim of our study was to evaluate an experimental model of new approaches for topical treatment of the implant surface and of periimplant defects by using DNA liposomes encoding for BMP-2 (bone morphogenetic protein). MATERIAL AND METHODS: A total of 27 implants (3.5 x 14 mm) were placed in critically sized defects of the frontal skull bone of adult pigs (n=3). The bottom of the implant was placed in the base of the defect which guaranteed primary stability, whereas the superior part of the implant (10 mm) represented an implant in a defect area. Liposomes containing DNA encoding for BMP-2 and GFP (green fluorescence protein) were used. In a first trial GFP-DNA liposomes on a collagen matrix were directly applied to the periimplant defect. In a second stage, the surface of the implants was encoded with BMP-2 DNA liposomes. Subsequently, these implants were inserted in the manner described. The resulting bone samples were prepared for immunohistochemical staining. Staining for GFP was performed in the area of the defect and for BMP-2 on the bone-implant interface. RESULTS: Immunohistochemical staining on day 3 postoperatively revealed an increased GFP expression in the periimplant defect. Therefore, the effectiveness of the liposomal vector was verified for the chosen animal model. On the surface of the implants encoded with BMP-2 DNA liposomes an increased BMP-2 expression was found. Thus, the liposomal vector system was validated also for BMP-2 DNA transfer in the chosen animal model. Further, the established system allows a sustainable and delayed release of BMP-2 in the area of the bone-implant interface. CONCLUSIONS: As a result of the study we were able to collect data concerning the influence of implant surface conditioning on the bone-implant interface and on therapeutically relevant options for the treatment of periimplant defects. These approaches are currently being evaluated in a long term study. PMID- 15293122 TI - [The use of a resorbable, synthetic membrane for the elevation of the sinus floor]. AB - The elevation of the sinus floor is a routine procedure in pre-implantological surgery. We report about our experience utilizing a resorbable, synthetic membrane (Ethisorb-Patch) for stabilization of the new generated floor of the sinus and for covering minor perforations of the sinus mucosa. Due to the easy handling and the good clinical results the use of the Ethisorb-Patch during sinus floor augmentation can be recommended. PMID- 15293123 TI - [Shammah-associated oral leukoplakia-like lesions]. AB - CASE REPORT: Shammah is a chewing tobacco, commonly used in Northern Africa. Leukoplakia-like lesions and oral cancer may be induced. In a 73-year-old male patient from Algeria leukoplakia-like lesions were observed in the anterior mandibular vestibulum and lower lip. The patient has been using shammah for 39 years. During the day three portions of shammah wrapped in a piece of paper tissue are prepared and rest in situ for 4-5 h. Due to its high alkalinity, shammah induces lesions resembling a burn. FINDINGS: Clinically, a white homogeneous lesion was seen in the vestibulum and mucosa of the lower lip. The white lesions could not be wiped off. Gingival recessions were seen in the lower front teeth. Root surfaces showed black-brown discoloration. A brush biopsy did not reveal epithelial atypia. The oral cavity showed signs of denture stomatitis and erythematous candidiasis. Microbiologically, Candida albicans was demonstrated. CONCLUSION: Due to the possibility of oral cancer in association with this habit, leukoplakia-like lesions have to be followed up diligently. Since chewing tobacco is a rare habit in Germany, changes as observed in the present case report may only rarely be observed. Due to migration it seems likely that in the future oral habits may be observed which are atypical for Western Europe. This phenomenon has also been addressed as transcultural dentistry. PMID- 15293129 TI - Clinical significance of the minor duodenal papilla and accessory pancreatic duct. AB - The accessory pancreatic duct (APD) is the main drainage duct of the dorsal pancreatic bud in the embryo, entering the duodenum at the minor duodenal papilla (MIP). As development progresses, the duct of the dorsal bud undergoes varying degrees of atrophy at the duodenal end. In cases of patent APD, smooth-muscle fiber bundles derived from the duodenal proper muscular tunics surround the APD. The APD shows long and short patterns on pancreatography, and ductal fusion in the two types appears to differ embryologically. Patency of the APD in control cases, as determined by dye-injection endoscopic retrograde pancreatography, was 43%. Patency of the APD may depend on duct caliber, course, and terminal shape of the APD. A patent APD may prevent acute pancreatitis by reducing the pressure in the main pancreatic duct. Pancreas divisum is a common anatomical anomaly in which the ventral and dorsal pancreatic ducts do not unite embryologically. As the majority of exocrine flow is routed through the MIP in individuals with pancreas divisum, interrelationships between poor function of the MIP and increased flow of pancreatic juice caused by alcohol or diet may increase dorsal pancreatic duct pressure and lead to the development of pancreatitis. Wire-guided minor sphincterotomy, followed by dorsal duct stenting, is recommended for acute recurrent pancreatitis associated with pancreas divisum. PMID- 15293130 TI - The minimum pressure of the lower esophageal sphincter, determined by the rapid pull-through method, is an index of severe reflux esophagitis. AB - BACKGROUND: Because of the wide overlap between the groups, patients with reflux esophagitis (RE) cannot be distinguished from healthy subjects by basal lower esophageal sphincter (LES) pressure. The LES has radial asymmetry, with maximum pressure on the left side. It has been noted that one directional LES pressure, determined using the rapid pull-through (RPT) method, in most patients with severe RE is very low, compared with that in healthy subjects. The aim of this study was to examine whether or not the minimum value of four directional LES pressures could become an index of RE by the RPT method. METHODS: Thirty patients with severe RE (grade C or D of the Los Angeles classification) were compared with 30 patients with mild RE (grade A or B) and 30 healthy subjects of comparable age and sex. LES pressure was measured by the RPT method, with the subject in the supine position. The catheter was withdrawn manually at a rate of 10 mm/s during suspended respiration at the end-expiratory phase after 5-min rest accommodation. LES pressure, with reference to the intragastric fundic pressure, was recorded by three consecutive RPTs, and 12 LES pressures were obtained for each subject. RESULTS: Minimum LES pressure, mean LES pressure, and maximum LES pressure were calculated from the 12 LES pressures. Wide overlaps in the mean and maximum LES pressures between healthy subjects and patients with mild or severe RE occurred; however, the overlap of minimum LES pressure between these groups was clearly less than that for the mean and maximum LES pressures. If it is accepted that more than the tenth percentile of the minimum LES pressure in healthy subjects is the cutoff value for healthy subjects, the sensitivity was 93.3% and the specificity for severe RE was 93.3%. CONCLUSIONS: This finding would suggest that the minimum LES pressure determined by the RPT method is an index of severe RE. PMID- 15293131 TI - Comparison between a new 13C-urea breath test, using a film-coated tablet, and the conventional 13C-urea breath test for the detection of Helicobacter pylori infection. AB - BACKGROUND: In Japan, urea breath-testing includes mouth rinsing with water immediately after the ingestion of (13)C-urea solution, to prevent false-positive results that are caused by oral bacteria with urease activity. Our objective was to evaluate the diagnostic performance of a urea breath test using a film-coated (13)C-urea tablet and omitting mouth rinsing. METHODS: The study was a multicenter trial comparing the solution- and tablet-based urea breath tests (UBTs). Helicobacter pylori status was determined by histology, culture, and rapid urease testing. RESULTS: Of the 255 subjects who completed the study, evaluation of the tablet-based UBT was possible in 254, and comparison of the tablet-based UBT and the solution-based UBT was possible in 250 patients. When the assessment achieved by a combination of biopsy-based methods was used as a reference standard, the sensitivity, specificity, and accuracy of the tablet based method were determined to be 97.7%, 98.4%, and 98.0%, respectively. When the results of the solution-based UBT were used as a reference standard, the sensitivity, specificity, and accuracy of the tablet-based UBT were determined to be 96.9%, 97.6%, and 97.2%, respectively. CONCLUSIONS: The (13)C-urea tablet based method proved to be a simple and accurate test for the diagnosis of H. Pylori infection. Mouth rinsing was not required. PMID- 15293132 TI - 13C-Urea breath test, using a new compact nondispersive isotope-selective infrared spectrophotometer: comparison with mass spectrometry. AB - BACKGROUND: The (13)C-urea breath test ((13)C-UBT) is the most commonly used noninvasive method of detecting Helicobacter pylori infection. The isotope ratio mass spectrometer (IRMS) is the most commonly used device for this test, but the UBiT-IR300 infrared spectrophotometer, which, by comparison, is a more compact, less expensive, and easier to use analytical device, has now become widely used in the clinical setting in Japan. The objective of this study was to examine the diagnostic performance of the (13)C-UBT, using the UBiT-IR300. METHODS: A multicenter open-label study was performed, in which the (13)C-UBT was conducted using 100 mg of (13)C-urea. Analysis of (13)CO(2) in the expired breath was performed by infrared spectroscopy and mass spectrometry, and assessment of H. pylori infection was performed by culture, histological examination, and rapid urease test. RESULTS: In 255 cases of H. pylori infection diagnosed by biopsy methods, the (13)C-UBTs, performed with two different (13)C-ruea formulations, and using infrared spectroscopy for evaluation, showed a sensitivity of 97.7%, specificity of 98.0%, and accuracy of 97.8% (total number of evaluable cases, n = 505). The rate of agreement in the assessment of H. pylori infection between infrared spectroscopy and mass spectrometry was 100% ( n = 505). The regression equation for infrared spectroscopy to mass spectrometry was y = 0.9822x - 0.0809 ( n = 2542), with a correlation coefficient of r = 0.99989 ( P = 0.0001). CONCLUSIONS: Diagnosis of H. pylori infection can be performed using infrared spectroscopy as well as mass spectrometry. PMID- 15293133 TI - Evaluation of tumor development from the viewpoint of the three-dimensional configuration of isolated crypts in colorectal adenomas with a type IIIL pit pattern. AB - BACKGROUND: The relationship between the three-dimensional configurations of superficial and protruded-type colon tumors with a type IIIL pit pattern was examined to evaluate their morphological development. METHODS: Forty-five isolated crypts of superficial-type adenomas and 65 isolated crypts of protruded type adenomas were evaluated, all derived from segments of 12 colorectal lesions removed by surgery or endoscopic resection. For isolation of the crypts, an HCl digestion method was used. Morphological features of the isolated crypts were observed by scanning electron microscopy (SEM). We investigated the configuration, length, and width of the isolated crypts in a comparison of the superficial and protruded-type tumors. The configuration and surface structure of the crypts were classified into three types (scores 1 to 3) by the shape and degree of branching and protuberances on their surface. RESULTS: The average lengths of the isolated crypts of the superficial and protruded types were 310.9 +/- 122.9 micro m ( n = 45) and 434.4 +/- 163.3 micro m ( n = 65), respectively ( P < 0.001), and the average widths of the superficial and protruded types were 140.4 +/- 68.2 micro m and 171.5 +/- 73.3 micro m, respectively ( P = 0.02). The average scores for the degree of branching and protruberances in the superficial and protruded types were 1.178 +/- 0.442 and 1.569 +/- 0.529, respectively ( P < 0.0001). In terms of configuration, in the isolated crypts, the superficial-type tumors exhibited a two-layer structure and the protruded type had the shape of a spindle with protuberances. CONCLUSIONS: In colorectal tumors with a type IIIL pit pattern, the isolated crypts of protruded-type tumors displayed more roughness on their surface structure and were longer than those of superficial type tumors. This suggested that the three-dimensional configuration of the isolated crypt constitutes an important factor in the formation of the gross configuration of colorectal tumors. Colorectal superficial and protruded-type tumors may develop with different growth patterns. PMID- 15293135 TI - Peripheral CD8+/CD25+ lymphocytes may be implicated in hepatocellular injuries in patients with acute-onset autoimmune hepatitis. AB - BACKGROUND: The mechanism of liver injuries in autoimmune hepatitis (AIH) is not fully understood, especially because the onset is insidious and the clinical courses fluctuate with spontaneous exacerbation and improvement even without immunosuppressive therapies. METHODS: Eleven patients with acute-onset AIH, some of whom were without hypergammaglobulinemia or anti-nuclear antibodies, and 41 patients with chronic AIH were compared serologically, biochemically, and histologically to determine differences in liver injuries between patients with acute-onset and chronic AIH. All patients fulfilled the diagnostic criteria according to a scoring system proposed in 1999. RESULTS: Lymphocytes with CD8+/CD25+ markers in pretreatment blood were significantly more prevalent in acute-onset than chronic AIH patients (24% vs 14%; P < 0.05). After treatment, however, CD8+/CD25+ lymphocytes were fewer in patients with acute-onset than in those with chronic AIH at 1 and 2 weeks ( P = 0.0001). No other differences were noted in clinical characteristics or immunological parameters between patients with acute-onset and those with chronic AIH. In a patient with typical acute onset AIH, CD8+/CD25+ lymphocytes increased and decreased in parallel with the activity of liver disease. CONCLUSIONS: Activated CD8+ T lymphocytes with CD25 markers may be implicated in the development of acute-onset AIH. PMID- 15293134 TI - Epidemiological and clinical study of sporadic acute hepatitis E caused by indigenous strains of hepatitis E virus in Japan compared with acute hepatitis A. AB - BACKGROUND: We compared acute hepatitis E (AH-E) and acute hepatitis A (AH-A) to investigate the epidemiology, clinical features, and prognosis of AH-E caused by an indigenous hepatitis E virus (HEV) in Japan. METHODS: We enrolled 58 patients diagnosed with AH-A or AH-E (32 men and 26 women; age, 20-72 years) from December 1997 to October 2002. Phylogenetic analysis of the partial 412-nucleotide sequence of open reading frame (ORF) 2 was performed in patients with AH-E. RESULTS: Regarding the geographic distribution of the HEV genotype, genotype III was principally distributed in Honshu Island, and genotype IV in Hokkaido Island ( P = 0.0034). The phylogenetic analysis of the ORF2 region revealed that there were significant geographic differences in the distribution of the HEV strains in Japan, with some strains being widespread and some, localized. In comparison with AH-A patients, those with AH-E were older (56.1 +/- 10.6 vs 45.9 +/- 10.8 years; P = 0.0017). The proportion of males among patients with AH-E was significantly higher ( P = 0.0001). Pyrexia was often observed in AH-A, and malaise in AH-E. Laboratory data indicate that AH-E induces a weak immunological reaction, whereas jaundice appears earlier in AH-E than in AH-A. One patient with AH-E died of acute hepatic failure, but none of those with AH-A died during the study period. CONCLUSIONS: Our results suggest that there are geographical differences between HEV strains in Japan, and that AH-E is more common in males and older patients than AH-A. Laboratory data indicate a weak immunological reaction and early appearance of jaundice in AH-E. PMID- 15293136 TI - Clinical characteristics and prevalence of GB virus C, SEN virus, and HFE gene mutation in Japanese patients with nonalcoholic steatohepatitis. AB - BACKGROUND: This study was carried out to clarify differences in clinical characteristics between fatty liver and nonalcoholic steatohepatitis in a Japanese population, and to assess the significance of GB virus C (GBV-C) infection, SEN virus (SENV) infection, and HFE gene mutation in the pathophysiology of these conditions. METHODS: Twenty patients with nonalcoholic steatohepatitis and 18 patients with simple steatosis were enrolled, and their clinical characteristics and histological findings were compared. Detection of GBV-C RNA and SENV DNA was performed by polymerase chain reaction (PCR). Mutational analysis of the HFE gene was performed by PCR-restriction fragment length polymorphism (RFLP). RESULTS: Serum aspartate aminotransferase (AST) and ferritin were significantly higher ( P < 0.05, for both) in NASH than in simple steatosis, and serum total cholesterol (T-Chol) was significantly lower ( P < 0.05) in NASH than in simple steatosis. While GBV-C was detectable in the serum of only one patient with NASH, SENV was detected in 50% (15/30) of the patients whose sera were tested for this virus, but the prevalence was not significantly different between the two groups (42% [8/19] in simple steatosis and 64% [7/11] in NASH). The sex ratio, body mass index (BMI), and age were not significantly different between the two groups, and mutation in the HFE gene was not detected in any patient. CONCLUSIONS: Higher serum AST and ferritin, and lower serum T Chol are distinctive features in NASH when compared with simple steatosis. GBV-C infection, SENV infection, and HFE gene mutation were not considered to influence the development of NASH from simple fatty liver. PMID- 15293137 TI - Portal and splenic hemodynamics in cirrhotic patients: relationship between esophageal variceal bleeding and the severity of hepatic failure. AB - BACKGROUND: The relationship between portal and splenic vein hemodynamics, liver function, and esophageal variceal bleeding in patients with cirrhosis remains unclear. The aim of the present study was to investigate quantitative Doppler parameters of splanchnic hemodynamics in cirrhotic patients and to determine the value of the Doppler parameters in predicting esophageal variceal bleeding. METHODS: With the help of pulsed Doppler ultrasonography, we investigated portal and splenic hemodynamics in 18 healthy controls and in 45 patients with liver cirrhosis, in whom the relationship of splenic hemodynamics with esophageal variceal bleeding and the grade of cirrhosis was examined. RESULTS: Portal flow velocity was decreased in cirrhotic patients with Child's C cirrhosis, as compared to those with Child's A cirrhosis ( P < 0.001). The portal blood flow volume in Child's C cirrhosis were also significantly low compared to patients with Child's A and Child's B cirrhosis ( P < 0.001 and P < 0.05, respectively). There was a significant increase in the portal vein congestion index and splenic vein congestion index in patients with Child's C cirrhosis as compared to patients with Child's A cirrhosis ( P < 0.001). Among cirrhotic patients, the group with esophageal variceal bleeding had significantly greater splenic blood flow volume and splenic vein congestion index ( P < 0.001). Patients with ascites had significantly lower portal flow velocity ( P < 0.001) and higher portal vein congestion index and splenic vein congestion index ( P = 0.003 and P = 0.05, respectively) as compared to those without ascites. CONCLUSIONS: In this report we have shown that the decrease in blood flow and increased congestion indexes in the portal vein and splenic vein are related to the impairment of liver function in cirrhotic patients; these indexes may be valuable factors for predicting esophageal variceal bleeding. PMID- 15293138 TI - Factors contributing to ribavirin dose reduction due to anemia during interferon alfa2b and ribavirin combination therapy for chronic hepatitis C. AB - BACKGROUND: Recent studies indicate that combination therapy with ribavirin and interferon alfa2b (IFNAlpha2b) is effective for chronic hepatitis C virus (HCV) infection. However, reversible hemolytic anemia is a common side effect of this therapy. METHODS: We determined those factors that contribute to ribavirin dose reduction due to anemia during this treatment by using multiple logistic regression analysis in Japanese patients. The study included 123 patients with chronic hepatitis C (85 male, 38 female; mean age, 50 years; range, 20-70 years), who received 24-week combination therapy. All patients were treated with IFNAlpha2b daily for 2 weeks, followed by three times weekly dosing for 22 weeks, with oral ribavirin twice daily, at a total daily dose of 600 or 800 mg. RESULTS: Of the 123 patients, 34 patients required dose reduction of ribavirin, and 78 patients required no dose reduction. Overall, 20 patients discontinued. On univariate analysis, reduction of the ribavirin dose correlated significantly with pretreatment hemoglobin (Hb) levels of less than 14 g/dl, female sex, and patient age 55 years or older. On multivariate analysis, pretreatment Hb of less than 14 g/dl level and age 55 years or older were significantly associated with ribavirin dose reduction. The hazard ratios were 3.56 (95% confidence interval [CI], 1.48-8.53) for pretreatment Hb levels of less than 14 g/dl, and 2.50 (95% CI, 1.05-5.94) for age 55 years or more. CONCLUSIONS: Because patient age of 55 years or more, and Hb levels of less than 14 g/dl are significant factors that influence ribavirin-induced hemolytic anemia, more careful monitoring is necessary during combination therapy for patients with these risk factors. PMID- 15293139 TI - Types of human leukocyte antigen and decrease in HCV core antigen in serum for predicting efficacy of interferon-Alpha in patients with chronic hepatitis C: analysis by a prospective study. AB - BACKGROUND: A prospective study was conducted to evaluate the influence of host factors, including human leukocyte antigen (HLA), and viral factors, including hepatitis C virus (HCV) core antigen, on the response to interferon (IFN)-Alpha. METHODS: Natural IFN-Alpha was given to 66 patients with chronic hepatitis C at a dose of 9 million units per day for 2 weeks, followed by 9 million units three times a week for 22 weeks. RESULTS: Sustained virological response without detectable HCV RNA in serum 24 weeks after the end of IFN therapy was achieved in 21 patients, while it was not in 32 patients; the remaining 13 patients were not evaluated. HCV core antigen and HCV RNA started to decrease 1 and 4 weeks, respectively, after the commencement of IFN in responders ( P = 0.02 and P = 0.05, respectively). On univariate analysis, age of 50 years or less ( P < 0.001); lack of HLA DR6 ( P = 0.018) or DR52 ( P < 0.041); platelets more than 14 x 10(4)/mm(3) ( P = 0.031); HCV core antigen 500 fmol/l or less ( P = 0.001); and HCV RNA 100 KIU/ml or less were predictive of response. On multivariate analysis, age 50 years or less (odds ratio [OR], 4.009; P = 0.039); lack of HLA DR6 (OR, 8.130; P = 0.027); IFN-naive (OR, 11.63; P = 0.016); HCV core antigen 500 fmol/l or less (OR, 10.61; P = 0.007); and genotypes other than 1b (OR, 8.929; P = 0.010) were predictive of response. CONCLUSIONS: Lack of HLA DR6 determined the response to IFN. HCV core antigen was useful in predicting and monitoring the response to IFN. PMID- 15293140 TI - Recurrence of hepatocellular carcinoma 102 months after successful eradication and removal of membranous obstruction of the inferior vena cava. AB - We report a 54-year-old Japanese woman who developed liver tumors 102 months after hepatic resection for hepatocellular carcinoma (HCC) and percutaneous transluminal angioplasty (PTA) for membranous obstruction of the inferior vena cava (MOVC), which is one form of Budd-Chiari syndrome. In the present admission workup showed no evidence of co-infection with hepatitis B and C viruses. Dynamic computed tomography (CT) and magnetic resonance imaging showed an enhanced lesion, 1.5 cm in diameter, in segment 3 of the liver, and no obstruction of the inferior vena cava after PTA. CT during both arterial portography and hepatic arteriography revealed another lesion, showing different hemodynamics, in segment 2. The patient therefore underwent hepatic resection, and the tumors were diagnosed histologically as HCC. The two tumors differed in their morphological features, one containing abundant fibrous stroma, whereas the other did not. The nontumorous liver tissue showed central zonal fibrosis, i.e., reversed lobulation, and partial expansion of nodule-like formations, indicating lack of progression since the situation seen at the initial hepatectomy. The presence of nontumorous liver tissue showing the above features suggests that, even after successful treatment for relief of congestion, patients who have had MOVC should be followed closely for as long as possible because of the risk of HCC recurrence. This is the first reported case of HCC recurrence after successful treatment of MOVC. PMID- 15293141 TI - Hypoplasia of the right hepatic lobe associated with portal hypertension and idiopathic retroperitoneal fibrosis. AB - A 22-year-old man was referred to our hospital because of thrombocytopenia. Abdominal computed tomography (CT) revealed hypoplasia of the right hepatic lobe, the development of porto-systemic collateral vessels, splenomegaly and a periaortic soft-tissue mass. Laboratory tests and needle liver biopsy indicated no evidence of liver cirrhosis. Consequently, a diagnosis of hypoplasia of the right hepatic lobe associated with portal hypertension and idiopathic retroperitoneal fibrosis was established. Portal hypertension and hypersplenism was thought to be the cause of the thrombocytopenia. CT arterioportography revealed that anomalies of the portal venous system could have resulted in the hypoplasia of the right hepatic lobe. This is the first report describing hypoplasia of the right hepatic lobe accompanied by supervening idiopathic retroperitoneal fibrosis. PMID- 15293142 TI - Primary non-Hodgkin's lymphoma of the common bile duct presenting as obstructive jaundice. AB - Primary non-Hodgkin's lymphoma of the extrahepatic bile duct presenting as obstructive jaundice is an extremely rare disease. At this writing, a review of the medical literature disclosed 17 reported cases of primary non-Hodgkin's lymphoma arising from the extrahepatic bile duct. We, herein, report an additional case of obstructive jaundice caused by primary non-Hodgkin's lymphoma of the common bile duct, in a 21-year-old woman. Our patient showed clinical evidence of obstructive jaundice, and endoscopic retrograde cholangiopancreatography and abdominal magnetic resonance imaging demonstrated a long strictured segment of the common bile duct with proximal bile duct dilatation. These clinical and radiological findings resembled those of cholangiocarcinoma. Resection of the common bile duct tumor, cholecystectomy, lymph node dissection, and Roux-en-Y hepaticojejunostomy were carried out. Histology and immunohistochemistry of the resected specimen confirmed a diffuse large B-cell-type malignant lymphoma involving the common bile duct. She received four courses of combination chemotherapy, including cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP), and 3060 cGy external irradiation. She has been well, without evidence of tumor recurrence, 17 months after the surgery. In summary, first, primary non-Hodgkin's lymphoma of the extrahepatic bile duct, despite its rarity, should be considered in the differential diagnosis of causes of obstructive jaundice. Second, an accurate histopathologic diagnosis and surgical resection, if feasible, combined with chemotherapy with or without radiotherapy may be the approach to offer a chance for cure. PMID- 15293143 TI - Rectal carcinoid arising in ulcerative colitis associated with rectal adenocarcinoma. PMID- 15293144 TI - Taiwan's contribution to clinical research in gastroenterology and hepatology. PMID- 15293146 TI - A new index of severe reflux esophagitis: what does the minimum LES pressure represent? PMID- 15293147 TI - Epidemiological and clinical features of hepatitis E in Japan. PMID- 15293148 TI - Ribavirin-induced hemolytic anemia in chronic hepatitis C patients. PMID- 15293149 TI - Budd-Chiari syndrome and hepatocellular carcinoma. PMID- 15293154 TI - Minimally invasive thoracic surgery for diagnostic assessment and palliative treatment in recurrent neoplastic pleural effusion. AB - BACKGROUND: We evaluated the effectiveness of VATS in the diagnosis and palliative treatment of recurrent neoplastic pleural effusions. METHODS: From 1987 to 2001, we performed 325 VATS chemical pleurodesis for malignant pleural effusions. We used talc in 253 subjects (78 %) and alcohol in 72 (22 %) as the sclerosant agent. In 226 patients (68 %) we performed biopsies because the histology was unknown. RESULTS: Mean operating time was 33.38 +/- 9.77 minutes (median: 32; range: 19 - 58), and the mean duration of chest intubation was 3.78 +/- 1.33 days (median: 4; range 2 - 8). Complications occurred in 2 % of patients. Thirty-day mortality was 2 %. Mean postoperative in hospital stay was 5.53 +/- 1.90 days (median 6; range: 2 - 11). We obtained 264 (81 %) therapeutic successes (no effusion recurrence within 4 months), and 55 relapses of which 32 had talc insufflation (13 % of talc group) and 23 alcohol instillation (32 % of alcohol group). CONCLUSIONS: VATS chemical pleurodesis is a safe, useful, versatile procedure for oncological pleural effusion management. The use of talc rather than alcohol significantly increased the therapeutic success rate. VATS should be considered the treatment of choice in patients with advanced neoplasm to obtain good palliation and a better quality of life. PMID- 15293155 TI - Clinicopathological analysis of clinical N0 peripheral lung cancers with a diameter of 1 cm or less. AB - BACKGROUND: The appropriateness of limited resection for small-sized lung cancer continues to be debated. It is not yet clear whether tumor size alone is a reliable indicator for limited resection. METHODS: From 1980 to 2002, 27 patients with clinical N0 peripheral lung cancers having diameters of 1 cm or less underwent pulmonary resection. Clinicopathological features of these cases were reviewed retrospectively. RESULTS: Histological analysis showed that there were 23 cases of adenocarcinoma (81.5 %), two large cell carcinomas (7.4 %), and two carcinoid tumors. Twenty-two cases were classified as pathological stage 1, one was stage II, and four were stage III. Four patients (14.8 %) had lymphatic vessel invasion, three (11.1 %) had lymph node metastasis, and two (7.4 %) had intrapulmonary metastasis. The 5-year disease-free survival rate was 76.6 %. CONCLUSIONS: It is impossible to predict the effectiveness of an intentional limited resection for lung cancer using only tumor size. PMID- 15293156 TI - Hemi-clamshell approach for advanced primary lung cancer. AB - BACKGROUND: The hemi-clamshell approach provides a wide anterior view of the mediastinum, apical dome, and cervicothoracic area. However, only a few reports have been made regarding this technique. METHODS: The hemi-clamshell approach was used in 24 patients, of whom 5 had a Pancoast tumor, 15 had mediastinal involvement, and 4 underwent mediastinal lymphadenopathy for left-sided lung cancer. Twenty-one of the patients received preoperative therapy. RESULTS: Twenty one operations were complete resections. In addition, 12 patients received cardio vascular reconstruction and 5 a first rib resection. Postoperative major morbidity was 21 % (6/24) and mortality was 4.2 % (1/24). Nine patients died of systemic tumor relapse and 14 patients were alive after a median follow-up of 24 months (range 3 - 68 months) following the initial therapy. The 5-year survival rate of patients with mediastinal involvement was 37 % and that of 13 patients with postoperative stage I or II was 35 %. CONCLUSIONS: The hemi-clamshell approach provides a wide exposure allowing a safe and complete removal of lung cancer that involves the mediastinum and apical thoracic dome, leading to a better long-term survival rate for patients with this disease. PMID- 15293157 TI - Outcome of surgery for small cell lung cancer -- response to induction chemotherapy predicts survival. AB - BACKGROUND: The role of surgery for local control of small cell lung cancer (SCLC) is controversial. METHODS: Sixty-nine consecutive patients who underwent complete resection of SCLC in our hospital were reviewed. The patients included 62 men and 7 women. Clinical stage at the time of diagnosis was c-stages IA and B in 29, c-stages IIA and B in 12, c-stage IIIA in 21, and c-stage IIIB in 7. RESULTS: Thirty-two patients received induction chemotherapy, and 37 patients underwent initial surgery. The overall response rate to induction chemotherapy was 71.9 %. The survival rate stratified by clinical stage at the time of diagnosis was 48.9 % for c-stage I, 33.3 % for c-stage II, 20.2 % for c-stage IIIA, and 0 % for c-stage IIIB. Downstaging after induction chemotherapy conferred a survival benefit. Survival after lobectomy or bilobectomy was better than after pneumonectomy. Patients who received adjuvant chemotherapy survived longer than patients who did not. CONCLUSIONS: Surgery combined with chemotherapy is a therapeutic option in selected patients with SCLC. Pathologic nodal status and response to induction chemotherapy are predictors of survival. PMID- 15293153 TI - [Pain management in non-juvenile, aseptic osteonecrosis]. AB - BACKGROUND: Adult aseptic osteonecrosis (ON) represents a clinical picture with unexplained etiology. Since curative treatment of this disease often succeeds only in the early stage, pain therapy plays an important role in the treatment process. METHOD: We compared established and novel treatment options for ON as well as our own results after i.v. administration of the prostacyclin analogue iloprost with corresponding studies in the literature. RESULTS: In addition to treatment with nonsteroidal antirheumatic agents and opioids, surgical "core decompression," vasoactive medications, and hyperbaric oxygenation are effective. Treatment with iloprost for 5 days resulted in highly significant pain reduction. CONCLUSION: Symptomatic treatment is indicated in all stages of ON and curative treatment in stage I and early stage II. In cases of disease progression in the large joints, early endoprosthetic replacement is indicated to avoid secondary damage. In addition to employing vasoactive substances, a further curative treatment approach could be the use of mesenchymal stem cells. PMID- 15293158 TI - Home monitoring of patients after prosthetic valve surgery -- experimental background and first clinical attempts. AB - PURPOSE: The purpose of this study was to investigate whether: 1. sound phenomena may be used to detect prosthetic valve dysfunction; 2. clinical and experimental data permit conclusions about alterations in the functional state of mechanical valves; 3. patients can record and pass on signals via Internet. METHODS: 1. We implanted bi-leaflet valves in pigs. By gradually influencing the motion of the tilting discs prosthetic dysfunction could be generated. 2. Thrombosis and lysis of bi-leaflet valves was studied in sheep. This process was documented using echocardiography and acoustically by the Fast Fourier Transformation. 3. Thirty devices were set up and handed out to patients following mechanical valve replacement. All patients regularly sent data to the hospital via Internet, regardless of their location at the time. The data were evaluated by comparing them with the reference file. RESULTS: Animal experiments proved that changes in prosthetic function led to a significant change in sound phenomena. In contrast to echocardiography alterations at an early stage (onset of thrombosis) could be reliably verified. The sensitivity was greater than in echo-control analysis. All patients regularly recorded and passed on their signals. Surveys revealed high acceptance and easy handling of the devices. CONCLUSIONS: Online registration of sound phenomena seems to be suitable for the detection of changes in prosthetic function. This led to the development of the first hand-held device for home monitoring of valve function. Registration of flow, frequency spectrum, and ECG envisaged at the next level opens up potential applications for Internet-based, remote monitoring of cardiac patients. PMID- 15293159 TI - Autologous fibrin glue reinforced by platelets in surgery of ascending aorta. AB - OBJECTIVES: Fibrin sealants are popular for the improvement of perioperative hemostasis and reducing blood transfusion needs. Biological glues prepared from pooled human donor plasma have an inherent risk of transmission of blood-borne disease and are quite expensive to use. A system for the production of autologous fibrin sealant re-enforced by platelets has been developed. Its efficacy, safety and economic benefits have been evaluated in a prospective, randomized study. MATERIAL: 20 consecutive patients undergoing replacement of the ascending aorta by the same surgical team had local application of either Tissucol (Group A) or autologous fibrin glue (Group B) for hemostasis. RESULTS: No adverse effects of either glue were recorded. The volume of produced autologous fibrin glue was 25 cc PRP. Platelet yield was 72 %. The two groups were comparable. Efficacy was group comparable. Average cost for sealants in Group A was 470 +/- 100 Euros compared to 273 Euros in Group B, p = 0.004. CONCLUSIONS: Autologous fibrin glue re-enforced by autologous platelets can be safely produced in the operating room in a large volume, with an comparable efficacy at a lower cost than commercial sealants. PMID- 15293160 TI - Surgical treatment of a large left-main coronary artery aneurysm. AB - Coronary artery aneurysms are a rare condition with left-main trunk aneurysms occurring in only about 0.1 % of the population. We report on a giant left-main coronary artery aneurysm in a young male status post two previous open-heart operations. The aneurysm was successfully treated by patch occlusion of the ostial orifice and coronary revascularization of the left anterior descending and circumflex arteries. PMID- 15293161 TI - Successful long-term bridge to transplant in a 5-year-old boy with the EXCOR left ventricular assist device. AB - We report on a 5-year-old boy who presented with postcardiotomy failure after aortic valve replacement and had to undergo implantation of a Berlin Heart-Excor system since treatment with ECMO did not improve myocardial pump function. After a stormy postoperative course with delayed sternal closure after 9 days, the young boy finally recovered and could be fully mobilized. Until successful heart transplantation after a support interval of 77 days, he experienced no device related infectious or thromboembolic complications. PMID- 15293162 TI - Massive hemothorax caused by intercostal artery bleeding: selective embolization may be an alternative to thoracotomy in selected patients. AB - Massive hemothorax is an indication for thoracotomy. We report a case of an 85 year-old debilitated patient, in whom massive hemorrhage from an actively bleeding intercostal artery was controlled by angiographic embolization. Angiographic embolization proved to be an effective alternative to thoracotomy in this patient, thus avoiding numerous postoperative complications and high mortality. Massive bleeding from an intercostal artery should be considered an indication for angiographic embolization in selected patients. PMID- 15293163 TI - Biological bypass in cardiovascular surgery. AB - Protein and gene therapy offer a tremendous opportunity to improve the care of critically ill patients with ischemic heart and peripheral artery occlusion disease. With the availability of purified growth factors such as vascular endothelial and fibroblast growth factors (FGF), several experimental and clinical studies provided data, that the growth of capillaries (angiogenesis) and of collateral arteries (arteriogenesis) is not limited to its natural time course. When applied in experimental models and in conjunction with coronary artery bypass operations, FGF in particular, led to a significant increase in endogenous rerouting of blood flow by collateral vessels inside the tissue itself. Thus, the proliferation of preexisting bypassing arterioles could be enhanced therapeutically (biological bypass). The purpose of this review is to discuss the physiological importance of different kinds of cytokines which are able to induce angio- and arteriogenesis in ischemic limbs or the heart. It is outlined that a combination of a sufficient amount of large arterioles and a capillary network are needed to compensate perfusion deficits. Each patient, who has an ischemic area and cannot be conventionally revascularized, is a potential candidate for the biological bypass. PMID- 15293164 TI - ICD implantation -- between necessity and tragedy. AB - We report on a 42-year-old patient with known dilative cardiomyopathy who underwent placement of a multiple transvenous pacemaker and ICD electrodes, and required removal of all leads via median sternotomy, followed by placement of epicardial electrodes. This experience has led us to question the necessity of the current implantation policies for these systems. PMID- 15293165 TI - The right of self-determination -- why not valid for Jehovah's Witnesses? PMID- 15293166 TI - [Therapeutic dose of omalizumab = 20 x [IgE]: is the equation correct or is enough too little?]. PMID- 15293167 TI - [Seasonal variations of serum-IgE and potential impact on dose-calculation of omalizumab (rhuMab-E25, anti-IgE)]. AB - BACKGROUND: The anti-IgE antibody Omalizumab has been approved for the treatment of perennial allergic asthma. Dosing of omalizumab is adjusted according to total IgE levels and body weight, and a near complete suppression of free IgE is deemed to be necessary for optimal efficacy. Many asthmatics, however, have additional sensitisations against seasonal allergens, e. g. pollen, and seasonal exposure may increase total and specific IgE levels ("boost"), thus leading to an imbalance between IgE levels and omalizumab with possible impact on therapeutic outcome. METHODS: We studied serum total and specific (timothy grass) IgE levels in 17 patients with seasonal allergic rhinitis and/or asthma prior to and during the grass pollen season. Based on total IgE levels, we then calculated hypothetical doses of omalizumab required for treatment at each time point. RESULTS: During the pollen season, total IgE increased significantly from a pre seasonal mean of 89 (50 - 178) kU/l (geom. mean with 95 % confidence interval) to 126 (63 - 251) kU/l (p = 0.0006). Accordingly, specific IgE increased from 11 (6.3 - 19) kU/l to 15.1 (8.3 - 29) kU/l (p = 0.0013). Calculated doses of omalizumab based on pre-seasonal IgE levels were: no dosing (IgE < 30 kU/l): n = 2; 150 mg 4-weekly: n = 7; 300 mg 4-weekly: n = 2; 225 mg 2-weekly: n = 4; 300 mg 2-weekly: n = 1; 375 mg 2-weekly: n = 1. Based on seasonal IgE levels, doses of omalizumab would have changed in 5/17 patients requiring the next possible dosing step. Of these, two patients would have fallen out of the current dosing scheme. CONCLUSIONS: A seasonal increase of serum total and specific IgE can be observed in patients with pollen allergy, although this increase would have no impact on omalizumab doses in the majority of patient. Individual variations, however, can be large and necessitate a dose correction. Therefore, therapeutic monitoring of free IgE levels during anti-IgE treatment appears as desirable tool. PMID- 15293168 TI - [Pneumopathy of drug addicts]. PMID- 15293169 TI - [Cardiopulmonary exercise tests -- proposals for standardization and interpretation]. AB - I give some recommendations concerning methodology and interpretation of cardiopulmonary exercise tests. The recommendations are based on our comprehensive data bank of exercise tests (282 tests and about 200 single parameters assessed during each test). When I expect an exercise capacity lower than 100 W I perform a ramp test; concerning expected higher exercise capacity steps of 25 W every 2 min are preferred. In order to achieve an optimal assessment of exercise capacity an exhaustion or symptom limited test should be performed. The achieved maximum oxygen consumption does not allow differing between cardiac or pulmonary causes of exercise limitation. It is only a marker of cardiopulmonary exercise capacity. A lot of algorithms to assess the maximum oxygen consumption are available, yet the results of calculating oxygen consumption with these algorithms differ considerably. Therefore it is mandatory to mention the used algorithm when referring to a calculated predicted oxygen consumption value. There are also several methods to assess the ventilatory and metabolic anaerobic threshold. For clinical purposes assessing lactate values is not necessary. The so called 4 mmol x l(-1) threshold accords primarily to the threshold assessed with the V-slope method. The Hf-slope may be used as an index for classification of heart failure stages analogous to the NYHA classification. Changes in dead space ventilation are mainly an expression of changed ventilation perfusion relationships and do not give evidence for any specific cardiac or pulmonary disorder. The slope of the equivalent for CO(2) is a relevant parameter of prognosis in cardiac failure. The value of the breathing reserve is not indicative of pathologic ventilatory limitation of exercise. You may find a reduced breathing reserve of about 0 also in healthy volunteers who are driven to exhaustion limited exercise. The value of the breathing reserve depends strongly on the kind of calculation or measuring mode and depending on the mode you can get normal or extremely reduced values in the same test person. The analysis of the flow volume curve during exercise provides some criteria of ventilatory exercise limitation. Pulse oxymetry is relevant only as a safety parameter. Because of its inaccuracy it should not be used to prove desaturation during exercise. The assessment of the alveolar-arterial pO (2) difference is of diagnostic relevance. The Borg scale, the course of the oxygen equivalent of O(2), the respiratory exchange ratio, and the aerobic capacity are of no major relevance for differential diagnosis. PMID- 15293170 TI - [Multicenter study on "non-invasive ventilation in patients with severe chronic obstructive pulmonary disease and emphysema(COPD)"]. AB - Non-invasive ventilation is applied with increasing frequency in patients with chronic hypercapnic COPD and insufficiency of the ventilatory pump. In the few existing clinical trials on long-term use of NIV, no significant improvement on survival could be proven, mainly due to methodical reasons. The "National Task Force for Non-invasive ventilation and weaning" plans to study patients with severe COPD and hypercapnic ventilatory pump insufficiency in a prospective, randomised, multicentre clinical trial over one year. In the intervention group, NIV will be applied for at least six hours per day in addition to standard COPD treatment. The target of mechanical ventilation is a reduction of PCO (2) during spontaneous breathing by at least 20 %, or into the normal range. The main outcome parameter is all-cause mortality, secondary outcome parameters are course of the disease, exercise capacity, quality of life and consumption of medical resources. The sample size is estimated on 300 patients (150 control group, 150 intervention group). The whole study will take approximately three years. PMID- 15293173 TI - Guidelines for preoperative administration of patients' home medications. AB - Currently, there are no agreed upon guidelines for the administration of preoperative medications. Institutional guidelines were formulated after a review of the literature, recommendations by experts, and a consensus among anesthesiologists, surgeons, pharmacists, and nursing educators. These guidelines were then provided to the preadmission staff to instruct patients regarding preoperative medications. These guidelines will have to be reassessed periodically as new medications and medical evidence emerge. PMID- 15293171 TI - [Guideline of the German respiratory society for diagnosis and treatment of patients suffering from acute or chronic cough]. PMID- 15293174 TI - Telephone follow-up for day surgery patients: patient perceptions and nurses' experiences. AB - This article is an analysis of qualitative data collected from telephone interviews by a nurse researcher with patients recovering from day surgery. The nurse researcher used a standard protocol to telephone 238 recovering day surgery patients. While answering their questions and providing advice, the researcher found that patients held many biases and misconceptions about pain and pain management. Many of these misconceptions were not apparent preoperatively nor at discharge because patients are anxious, still recovering from the surgical experience, and not always able to absorb information or anticipate future issues. This article discusses those misconceptions and the necessity that follow up occurs over a time period, as the patient's need for advice and support changes throughout the recovery process. PMID- 15293175 TI - A PACU crisis: a case study on the development and management of methemoglobinemia. AB - The development of methemoglobinemia requires rapid recognition, confirmation, and treatment. This case study describes the development, diagnosis, and management of a 63-year-old male scheduled for a laparoscopic cholecystectomy with an intraoperative cholangiogram who developed methemoglobinemia after benzocaine was given for intubation. PMID- 15293176 TI - "Do not use" these abbreviations! PMID- 15293178 TI - Pain assessment for the PACU nurse: science or art? PMID- 15293180 TI - Hand washing: essential in the PACU. PMID- 15293181 TI - Too busy to wash your hands in PACU? PMID- 15293211 TI - The future of nursing: Are we "too posh to wash?". PMID- 15293212 TI - Ovine ooplasm directs initial nucleolar assembly in embryos cloned from ovine, bovine, and porcine cells. AB - Here we present ultrastructural and immunocytochemical evidence that ovine ooplasm is directing the initial assembly of the nucleolus independent of the species of the nuclear donor. Intergeneric porcine-ovine somatic cell nuclear transfer (SCNT) and intrageneric ovine-ovine SCNT embryos were constructed and the nucleolus ultrastructure and nucleolus associated rRNA synthesis examined in 1-, 2-, 4-, early 8-, late 8-, and 16-cell embryos using transmission electron microscopy (TEM) and light microscopical autoradiography. In addition, immunocytochemical localization by confocal microscopy of nucleolin, a key protein involved in processing rRNA transcripts, was performed on early 8-, late 8-, and 16-cell embryos for both groups of SCNT embryos. Intergeneric porcine ovine SCNT embryos exhibited nucleolar precursor bodies (NPBs) of an ovine (ruminant) ultrastructure, but no active rRNA producing fibrillo-granular nucleoli at any of the stages. Unusually, cytoplasmic organelles were located inside the nucleus of two porcine-ovine SCNT embryos. The ovine-ovine SCNT embryos, on the other hand, revealed fibrillo-granular nucleoli in 16-cell embryos. In parallel, autoradiographic labeling over the nucleoplasm, and in particular, the nulcleoli was detected. Bovine-ovine SCNT embryos at the eight cell stage were examined for nucleolar morphology and exhibited ruminant-type NPBs as well as structures that appeared as fibrillar material surrounded by a rim of electron dense granules, perhaps formerly of nucleolar origin. Nucleolin was localized throughout the nucleoplasm and with particular intensity around the presumptive nucleolar compartments for all developmental stages examined in porcine-ovine and ovine-ovine SCNT embryos. In conclusion, this study suggests that factors within the ovine ooplasm are playing a role in the initial assembly of the embryonic nucleolus in intrageneric SCNT embryos. PMID- 15293213 TI - Origin of the murine implantation serine proteinase subfamily. AB - The S1 serine protease family is one of the largest gene families known. Within this family there are several subfamilies that have been grouped together as a result of sequence comparisons and substrate identification. The grouping of related genes allows for the speculation of function for newly found members by comparison and for novel subfamilies by contrast. Analysis of the evolutionary patterns of genes indicates whether or not orthologs are likely to be identified in other species as well as potentially indicating that hypothesized orthologs are in fact not. Looking at subtle differences between subfamily members can reveal intricacies about function and expression. Previously, we have described genes encoding two novel serine proteinases, ISP1 and ISP2, which are most closely related to tryptases. The ISP1 gene encodes the embryo-derived enzyme strypsin, which is necessary for blastocyst hatching and invasion in vitro. Additionally both ISP1 and ISP2 are co-expressed in the endometrial gland during the time of hatching, suggesting that they may also both participate in zona lysis from within the uterine lumen. Here, we demonstrate that the ISPs are tandemly linked within the tryptase cluster on 17A3.3. We suggest that remarkable similarities within the 5'-untranslated and first intron regions of ISP1 and ISP2 may explain their intimate co-regulation in uterus. We also suggest that ISP genes have evolved through gene duplication and that the ISP1 gene has also begun to adopt an additional new function in the murine preimplantation embryo. PMID- 15293214 TI - P26h and dicarbonyl/L-xylulose reductase are two distinct proteins present in the hamster epididymis. AB - We have previously identified a 34 kDa protein (P34H) on the human sperm surface covering the acrosome. Using the hamster, we have also described a sperm protein, P26h, which is acquired by spermatozoa during epididymal transit. Both P34H and P26h belong to the carbonyl reductase family. Using molecular tools derived from P34H, we searched in the hamster epididymis for another protein related to the human sperm protein. Cloning and sequencing of P31h cDNA revealed 100% homology with the kidney DCXR (Dicarbonyl/L-Xylulose reductase). Northern Blot experiments revealed a single mRNA that was more expressed in the caput than in the corpus and cauda segment of adult epididymides. In situ hybridization was performed on sexually mature hamsters showing that the mRNA was localized in the principal cells throughout the epididymis. Using an anti-P34H antibody we have identified a P34H related protein named P31h (for 31 kDa). This protein showed 2D electrophoretic behavior different from P26h and was detectable all along the epididymis (caput, corpus, and cauda) by Western Blot analysis. Immunohistochemistry techniques showed that P31h was localized in the perinuclear region of the principal cells of the epididymal epithelium within the three sections, both in sexually mature and immature animals. Results are discussed with regards to the potential function of DCXR in the epididymis. PMID- 15293215 TI - Temporospatial expression of placental lactogen and prolactin-related protein-1 genes in the bovine placenta and uterus during pregnancy. AB - The anatomical location of binucleate cells (BNC) influences protein expression but not steroid synthesis in ruminants. In order to determine if BNC in disparate locations differentially express bovine placental lactogen (bPL) and prolactin related protein-1 (bPRP-1), we quantitated bPL and bPRP-1 transcripts in placentomal (cotyledonary, caruncular) and interplacentomal (intercotyledonary, intercaruncular) tissues throughout pregnancy in the bovine using real-time reverse transcription PCR (RT-PCR) and in situ hybridization. Levels of both bPL and bPRP-1 transcripts at peri-implantation were significantly higher (P < 0.01) in the fetal membrane than in caruncular and intercaruncular tissues. Thereafter, mRNA for these related proteins demonstrated different spatial as well as temporal patterns of expression. Levels of bPRP-1 transcripts peaked at day 60 of pregnancy. Between day 60 and 100, bPRP-1 transcripts fell by approximately sevenfold (P < 0.01) in cotyledonary and intercotyledonary tissues, and fourfold in caruncular (P < 0.01) tissue. Levels of bPRP-1 transcripts remained low in the cotyledonary, intercotyledonary, and caruncular tissues until peripartum. In contrast, bPL expression in placentomes increased with progression of gestation (P < 0.01), but decreased in interplacentomal tissue around peripartum. To conclude, disparate patterns of bPRP-1 and bPL genes are transcribed in the placentomal and interplacentomal tissues during gestation in the bovine, suggesting that these prolactin-like hormones serve distinct functions and are regulated differently in the uteroplacental unit in this species. PMID- 15293216 TI - Production of transgenic rats by ooplasmic injection of spermatogenic cells exposed to exogenous DNA: a preliminary study. AB - The aim of the present study was to investigate the efficiencies of producing transgenic rats by the ooplasmic injection of sperm heads (intracytoplasmic sperm injection: ICSI) and elongating spermatids (elongating spermatid injection: ELSI) exposed to the EGFP DNA solution. A slightly lower proportion of ICSI oocytes using sperm heads exposed to a concentration of 0.5 microg/ml DNA solution for 1 min developed into offspring (13.3%, 48/361) when compared to that of oocytes injected with nontreated sperm heads (19.4%, 32/165). Eight ICSI offspring were found to be EGFP-carrying transgenic rats (16.7% per offspring; 2.2% per embryo). After a 1-min exposure of the elongating spermatids to 5 microg/ml of DNA solution, 8.8% (45/511) of the ELSI oocytes developed into offspring while 12.7% (22/173) of the ELSI oocytes using nontreated spermatids developed. Six ELSI offspring carried the EGFP DNA (13.3% per offspring; 1.2% per embryo). The conventional pronuclear microinjection of 5 microg/ml of DNA solution resulted in the higher production of offspring (29.7%, 104/350) and the birth of three transgenic rats (2.9% per offspring; 0.9% per embryo). Thus, sperm heads and elongating spermatids were practically useful as the vector of exogenous DNA if the DNA-exposed spermatogenic cells were microinseminated into rat oocytes. PMID- 15293217 TI - Effects of bone morphogenetic protein-7 (BMP-7) on primordial follicular growth in the mouse ovary. AB - Previously, bone morphogenetic protein-7 (BMP-7) was suggested as a factor that may act to facilitate the transition of follicles from primordial stage to the pool of developed primary, preantral, and antral follicles (Lee et al. 2001: Biol Reprod 65:994-999.). Thus, aim of the present study was to evaluate effect(s) of BMP-7 on the primordial-primary follicle transition. Neonatal mouse ovaries were cultured in the presence or absence of 100 mIU/ml FSH with various doses of BMP-7 (0, 10, and 100 ng/ml). After 4-day culture period, number of follicles was counted and the expression of transcripts for FSH receptor (FSHR), kit ligand (KL), and c-kit was measured by RT-PCR. BMP-7 alone at 100 ng/ml concentration stimulated follicle development with concurrent increase of mRNA for FSHR. BMP-7 alone down-regulated KL expression however, the ratio between KL1 and KL2 was increased. There was no change in the c-kit mRNA expression. Results of the present study suggest that the BMP-7 is one of the factors involved in primordial primary follicle transition in the mouse ovary and it may play a role in expression of FSHR for further follicular development. PMID- 15293218 TI - Isolation and characterization of a bovine trophectoderm cell line derived from a parthenogenetic blastocyst. AB - A bovine trophectoderm cell line was established from a parthenogenetic in vitro produced blastocyst. To initiate the cell line, 8-day parthenogenetic blastocysts were attached to a feeder layer of STO fibroblasts and primary outgrowths occurred that consisted of trophectoderm, endoderm, and very occasionally epiblast tissue. Any endoderm and epiblast outgrowths were removed from the primary cultures within the first 10 days of culture by dissection. One of the primary trophectoderm cell cultures was chosen for further propagation and was passaged by physical dissociation and replating on STO feeder cells. The cell culture, designated BPT-1, was maintained in T25 flasks and passaged at a 1:3 split ratio for the first 15 passages approximately once every 2 weeks. Thereafter, the cell culture was passaged at 1:10-1:40 split ratios. Transmission electron microscopic examination showed the cells to be a polarized epithelium with apical microvilli, a thin basal lamina, and lateral junctions consisting of tight junctions and desmosomes. Lipid vacuoles and digestive vacuoles were also prominent features of the BPT-1 cells. Metaphase spread analysis at passage 59 indicated a near diploid cell population (2n = 60) with a mode and median of 60 and a mean of 64. BPT-1 cells secreted interferon-tau into the medium as measured by anti-viral assay and Western blot analysis. The cell line provides an in vitro model of parthenogenote trophectoderm whose biological characteristics can be compared to trophectoderm cell lines derived from bovine embryos produced by normal fertilization or nuclear transfer. PMID- 15293219 TI - Studies on Ca2+-channel distribution in maturation arrested mouse oocyte. AB - The present study was carried out to identify the existence of voltage-dependent Ca2+-channels (P/Q-, N-, and L-type) and their distributional differences in germinal vesicle (GV) and GV breakdown (GVBD)-arrested mouse oocytes which includes GVBD to telophase I of meiosis I and matured oocytes (MII, metaphase of meiosis II) by using the immunocytochemical method and a confocal laser scanning microscope. (1) Comparison between follicular oocytes (GV) and GV-arrested oocytes after 17 hr of in vitro culture. In follicular oocytes, P/Q-, N-, L (anti alpha1C anti-alpha1D)-type Ca2+-channels showed both localized and uniform staining. In contrast, GV-arrested oocytes, after in vitro culture for 17 hr, showed no presence of Ca2+-channels in most oocytes. (2) Comparison between GVBD oocytes after culture in vitro for 3 hr and GVBD-arrested oocytes after culture in vitro for 17 hr. In GVBD oocytes, P/Q-, N-, L (anti-1C, anti-alpha1D)-type Ca2+-channels showed both localized and uniform staining. In contrast, in GVBD arrested oocytes, none of the three types of Ca2+-channels were identified in 72 86% of oocytes. The present study demonstrates that in most GVBD-arrested oocytes that do not mature to MII, there is no Ca2+-channel identified. Therefore, most of the GVBD-arrested oocytes seem to have defects in Ca2+-channel expression/translation. Also, distributional changes of Ca2+-channels take place depending on the maturation progress in GV oocytes and MII stage oocytes (ovulated and 17 hr cultured MII stage oocytes). In addition, we found evidence that a functional voltage-dependent Ca2+-channel (L-type) exists in mouse oocytes (ovulated and cultured MII staged oocytes by a confocal laser scanning microscope). PMID- 15293220 TI - Isolation and characterization of a haploid germ cell specific sperm associated antigen 9 (SPAG9) from the baboon. AB - Previously, we cloned and sequenced a sperm specific antigen, designated as HSS (EMBL nomenclature human sperm associated antigen 9: hSPAG9) from human testis (Shankar et al.: Biochem Biophys Res Commun 243:561-565, 1998). The present study was conducted to isolate baboon proteomic homologue in order to find out whether the baboon can provide a suitable model for examining its immunocontraception effects. Baboon SPAG9 (bSPAG9) was cloned and sequenced from the baboon testis cDNA library. The baboon cDNA contained open reading frame encoding 760 amino acids. A 90.6 and 96.8% homology between baboon and human SPAG9 was found at protein and DNA levels. Analysis for tissue specificity by Northern blot procedure using various baboon tissues indicated that bSPAG9 was specifically expressed only in the baboon testis. Further, cell type expression analysis by in situ hybridization in baboon testis demonstrated the expression of bSPAG9 mRNA transcript only in the round spermatid suggesting haploid germ cell expression. Anti-human SPAG9 antibodies recognized the acrosomal compartment region of baboon sperm in indirect immunofluorescence (IIF). Flow cytometry analysis showed surface localization of bSPAG9 in live baboon sperm. The amino acid sequence data for nonhuman primate SPAG9 suggest that antibodies generated by vaccinating baboon with hSPAG9 will recognize nonhuman primate SPAG9, supporting the testing of SPAG9 contraceptive vaccine based on hSPAG9 in the nonhuman primate model. PMID- 15293221 TI - A unique mechanism for cyclic adenosine 3',5'-monophosphate-induced increase of 32-kDa tyrosine-phosphorylated protein in boar spermatozoa. AB - A cAMP-induced increase of tyrosine-phosphorylated proteins is involved in the expression of fertilizing ability in mammalian spermatozoa. We (Harayama, 2003: J Androl 24:831-842) reported that incubation of boar spermatozoa with a cell permeable cAMP analog (cBiMPS) increased a 32-kDa tyrosine-phosphorylated protein (TyrP32). The purpose of this study is to characterize the signaling cascades that regulate the cAMP-induced increase of TyrP32. We examined effects of tyrosine kinase inhibitor (lavendustin A), tyrosine phosphatase inhibitor (Na3VO4), cell-permeable calcium chelator (BAPTA-AM), and cholesterol acceptor (methyl-beta-cyclodextrin: MBC) on the increase of TyrP32 and the change and loss of acrosomes in boar spermatozoa. The spermatozoa were used for detection of tyrosine-phosphorylated proteins by Western blotting and indirect immunofluorescence and for examination of acrosomal integrity by Giemsa staining. At least eight tyrosine-phosphorylated proteins including TyrP32 exhibited the cAMP-dependent increase during incubation with cBiMPS. In many proteins of them, this increase was reduced by lavendustin A but was enhanced by Na3VO4. In contrast, the cAMP-induced increase of TyrP32 was abolished by Na3VO4 but was hardly affected by lavendustin A. Giemsa staining showed that the increase of spermatozoa with weakly Giemsa-stained acrosomes (severely damaged acrosomes) or without acrosomes was correlative to the cAMP-induced increase of TyrP32. Moreover, the lack of calcium chloride in the incubation medium or pretreatment of spermatozoa with BAPTA-AM blocked the change and loss of acrosomes and the increase of TyrP32, suggesting these events are dependent on the extracellular and intracellular calcium. On the other hand, incubation of spermatozoa with MBC in the absence of cBiMPS could mimic the change and loss of acrosomes and increase of TyrP32 without increase of other tyrosine-phosphorylated proteins. Based on these results, we conclude that the cAMP-induced increase of TyrP32 is regulated by a unique mechanism that may be linked to the calcium-dependent change and loss of acrosomes. PMID- 15293222 TI - Thyroglobulin type-1 domain protease inhibitors exhibit specific expression in the cortical ooplasm of vitellogenic rainbow trout oocytes. AB - The synthesis, uptake, and processing of yolk proteins remain poorly described aspects of oviparous reproductive development. In this study, we report the identification and characterization of two protease inhibitors in rainbow trout ovary whose expression and distribution are directly associated with yolk protein uptake in vitellogenic oocytes. The first transcript, termed "oocyte protease inhibitor-1" (OPI-1), is predicted to encode a 9.1 kDa, 87 amino acid protein containing a single thyroglobulin type-1 (TY) domain, identifying it as a putative TY domain inhibitor. The second transcript, termed OPI-2, is predicted to encode an 18.3 kDa, 173 amino acid protein with two similar, but not identical, TY domains. Messenger RNA expression of both genes was first detected in ovarian tissues at the onset of vitellogenesis, and persisted throughout the vitellogenic growth phase. We did not detect expression of either gene in previtellogenic ovaries, nor in any somatic tissues examined. Expression of OPI-1 mRNA was significantly reduced in atretic follicles as compared to healthy vitellogenic follicles, suggesting a downregulation of inhibitor expression during oocyte atresia. Western immunoblot analyses of whole yolk from vitellogenic oocytes revealed the presence of two immunoreactive proteins that corresponded to the predicted sizes of OPI-1 and OPI-2. We detected strong crossreactivity of this antiserum with specific vesicles in the cortical ooplasm of vitellogenic oocytes, in regions directly associated with vitellogenin processing. The identification of OPI-1 and OPI-2 provides new evidence for the expression of multiple TY domain protease inhibitors likely involved in the regulation of yolk processing during oocyte growth in salmonids. PMID- 15293223 TI - Maternal chromatin remodeling during maturation and after fertilization in mouse oocytes. AB - Immunofluorescence staining with antibodies against acetylated histone H4 and 5 methylcytosine was carried out to investigate female chromatin remodeling throughout oocyte maturation and chromatin rearrangement involving both male and female genomes after fertilization. Oocyte cytoplasm remodels female chromatin in preparation of the fertilizing event and the subsequent chromatin rearrangement. Histone H4 are in fact progressively deacetylated whereas demethylating enzymes do not seem to be active over this period. The acetylase/deacetylase balance seems to be cell cycle dependent as female chromatin is deacetylated during maturation and reacetylated at telophase II stage both after fertilization and activation. On the contrary, DNA demethylation seems to be strictly selective. It is in fact confined to the remodeling of paternal genome after fertilization of mature oocytes as the ooplasm is not effective in demethylating either paternal chromatin in germinal vesicle breakdown (GVBD) fertilized oocytes or maternal genome of partenogenetically activated oocytes. Surprisingly, we induced maternal chromatin demethylation after fertilization by treating oocytes with a combination of a methyltransferase inhibitor, 5-azacytidine (5-AzaC), and a reversible and specific inhibitor of histone deacetylase, trichostatin A (TSA). This treatment likely induces a hyperacetylation of histones (thus favoring the access to demethylating enzymes by opening female chromatin structure) associated with a block of reparative methylation by inhibiting methytransferases. This manipulation of chromatin remodeling may have applications regarding the biological significance of aberrant DNA methylation. PMID- 15293224 TI - Regulation of histone acetylation during meiotic maturation in mouse oocytes. AB - Histone acetylation is an important epigenetic modification implicated in the regulation of chromatin structure and, subsequently, gene expression. Global histone deacetylation was reported in mouse oocytes during meiosis but not mitosis. The regulation of this meiosis-specific deacetylation has not been elucidated. Here, we demonstrate that p34(cdc2) kinase activity and protein synthesis are responsible for the activation of histone deacetylases and the inhibition of histone acetyltransferases (HATs), respectively, resulting in deacetylation of histone H4 at lysine-12 (H4K12) during mouse oocyte meiosis. Temporal changes in the acetylation state of H4K12 were examined immunocytochemically during meiotic maturation using an antibody specific for acetylated H4K12. H4K12 was deacetylated during the first meiosis, temporarily acetylated around the time of the first polar body (PB1) extrusion, and then deacetylated again during the second meiosis. Because these changes coincided with the known oscillation pattern of p34(cdc2) kinase activity, we investigated the involvement of the kinase in H4K12 deacetylation. Roscovitine, an inhibitor of cyclin-dependent kinase activity, prevented H4K12 deacetylation during both the first and second meiosis, suggesting that p34(cdc2) kinase activity is required for deacetylation during meiosis. In addition, cycloheximide, a protein synthesis inhibitor, also prevented deacetylation. After PB1 extrusion, at which time H4K12 had been deacetylated, H4K12 was re-acetylated in the condensed chromosomes by treatment with cycloheximide but not with roscovitine. These results demonstrate that HATs are present but inactivated by newly synthesized protein(s) that is (are) not involved in p34(cdc2) kinase activity. Our results suggest that p34(cdc2) kinase activity induces the deacetylation of H4K12 and that the deacetylated state is maintained by newly synthesized protein(s) that inhibits HAT activity during meiosis. PMID- 15293225 TI - Changes in germinal vesicle (GV) chromatin configurations during growth and maturation of porcine oocytes. AB - Changes in germinal vesicle (GV) chromatin configurations during growth and maturation of porcine oocytes were studied using a new method that allows a clearer visualization of both nucleolus and chromatin after Hoechst staining. The GV chromatin of porcine oocytes was classified into five configurations, based on the degree of chromatin condensation, and on nucleolus and nuclear membrane disappearance. While the GV1 to 4 configurations were similar to those reported by previous studies, the GV0 configuration was distinct by the diffuse, filamentous pattern of chromatin in the whole nuclear area. Most of the oocytes were at the GV0 stage in the <1 and 1-1.9 mm follicles, but the GV0 pattern disappeared completely in the 2-2.9 and 3-6 mm follicles. As follicles grew, the number of oocytes with GV1 configurations increased and reached a maximum in the preovulatory follicles 4 hr post-hCG injection. During maturation in vivo, the number of GV1 oocytes decreased while oocytes undergoing GVBD increased. The percentage of oocytes with GV3 and GV4 configurations was constant during oocyte growth except at the 2-2.9 mm follicle stage, but these configurations disappeared completely after hCG injection. On the contrary, the in vitro maturing oocytes showed a large proportion of GV3 and GV4 configurations. There was no significant difference in distribution of chromatin configurations between the nonatretic and atretic follicles, and between oocytes with more than two layers of cumulus cells and those with less than one layer or no cumulus cells. Overall, our results suggested that (i) the GV0 configuration in porcine oocytes corresponded to the "nonsurrounded nucleolus" pattern in mice and other species; (ii) all the oocytes were synchronized at the GV1 stage before GVBD and this pattern might, therefore, represent a nonatretic state; (iii) the GV3 and GV4 configurations might represent stages toward atresia, or transient events prior to GVBD that could be switched toward either ovulation or atresia, depending upon circumstances; (iv) the in vitro systems currently used were not favorable for oocytes to switch toward ovulation (or final maturation); (v) the number of cumulus cells was not correlated with the chromatin configuration of oocytes, indicating that the beneficial effect of cumulus cells on oocyte maturation and development may simply be attributed to their presence during in vitro culture. PMID- 15293226 TI - Expression of gp20, a human sperm antigen of epididymal origin, is reduced in spermatozoa from subfertile men. AB - gp20, a sialylglycoprotein of human sperm homologous to CD52, is present everywhere on the surface of the freshly ejaculated sperm but is prevalently localized in the equatorial region of the head of capacitated sperm. In the present study, we confirmed this feature on large scale and correlated equatorial exposure of the antigen to the presence of serum albumin (SA) in the capacitation medium. Furthermore, we analyzed the relationship between the presence of the antigen and its equatorial exposure after capacitation and fertility, by comparing immunostaining for gp20 in the motile fraction of spermatozoa from fertile and subfertile men. A significantly higher percentage of nonimmunostained spermatozoa before capacitation (38.5% +/- 23 vs. 12% +/- 7, P < 0.0001) and a lower increase in the percentage of sperm with equatorial localization after capacitation (19.3% +/- 25 vs. 34.6% +/- 22, P = 0.039) were observed in subfertile men (n = 60) compared to fertile men (n = 15). In the whole study group, a positive correlation was also found between the percentage of spermatozoa exhibiting equatorial localization in capacitated samples and normal head forms (R = 0.50; P < 0.0001). PMID- 15293228 TI - Nestin expression in hippocampal astrocytes after injury depends on the age of the hippocampus. AB - Analysis of the expression of nestin in reactive astrocytes facilitates quantification of the extent of activation of astrocytes after injury in the mature CNS. We hypothesize that the capability of astrocytes for re-expressing nestin in response to CNS injury diminishes as a function of age. We quantified astrocytes positive for S-100beta protein, glial fibrillary acidic protein (GFAP) and nestin in the hippocampus of young adult, middle-aged, and aged Fischer 344 rats after an intracerebroventricular kainic acid (KA) administration. In all age groups, KA administration induced degeneration of CA3 pyramidal neurons, which led to a significant deafferentation in the CA1 region. The KA-induced neurodegeneration and deafferentation resulted in an increased population of astrocytes positive for S-100beta and glial fibrillary acidic protein (GFAP) in all age groups. Interestingly, these increases were highly comparable across the three age groups. However, in areas of both neurodegeneration and deafferentation, the overall numerical density of nestin-positive reactive astrocytes varied depending on the age at the time of injury with noticeably decreased numerical density in the injured middle-aged and aged hippocampus. In contrast, nestin-immunoreactive radial glia framework after lesion is not impaired with aging in the ependymal lining of the CA3 region. PMID- 15293229 TI - Differential generation of oligodendrocytes from human and rodent embryonic spinal cord neural precursors. AB - Human neural precursors are considered to have widespread therapeutic possibilities on account of their ability to provide large numbers of cells whilst retaining multipotentiality. Application to human demyelinating diseases requires improved understanding of the signalling requirements underlying the generation of human oligodendrocytes from immature cell populations. In this study, we compare and contrast the capacity of neural precursors derived from the developing human and rodent spinal cord to generate oligodendrocytes. We show that the developing human spinal cord (6-12 weeks of gestation) displays a comparable ventrodorsal gradient of oligodendrocyte differentiation potential to the embryonic rodent spinal cord. In contrast, fibroblast growth factor 2 (FGF-2) expanded human neural precursors derived from both isolated ventral or dorsal cultures show a reduced capacity to generate oligodendrocytes, whereas comparable rodent cultures demonstrate a marked increase in oligodendrocyte formation following FGF-2 treatment. In addition, we provide evidence that candidate growth factors suggested from rodent studies, including FGF-2 and platelet-derived growth factor (PDGF) do not stimulate proliferation of human oligodendrocyte lineage cells. Finally, we show that the in vivo environment of the acutely demyelinating adult rat spinal cord is insufficient to stimulate the differentiation of immature human spinal cord cells to oligodendrocytes. These results provide further evidence for inter-species difference in the capacity of neural precursors to generate oligodendrocytes. PMID- 15293230 TI - N-myc downstream-regulated gene 1 expression in injured sciatic nerves. AB - N-myc downstream-regulated gene 1 (NDRG1)/RTP/Drg1/Cap43/rit42/TDD5/Ndr1 is expressed ubiquitously and has been proposed to play a role in growth arrest and cell differentiation. A recent study showed that mutation of this gene is responsible for hereditary motor and sensory neuropathy-Lom. However, the role of this gene in the peripheral nervous system is not fully understood. In our study, rabbit polyclonal antibodies were raised against this gene product and were used to examine changes in its expression over the time course of Wallerian degeneration and ensuing regeneration after crush injury of mouse sciatic nerves. Fluorescent immunohistochemistry showed that NDRG1 was expressed over the intact nerve fibers. Double labeling with a Schwann cell (SC) marker, S-100 protein (S 100), revealed that NDRG1 was localized in the cytoplasm of S-100-positive Schwann cells (SCs). NDRG1 expression was maintained in the early stage of myelin degradation but was then markedly depleted at the end stage of myelin degradation when frequent occurrence of BrdU-labeled SCs was observed (at 7-9 days). The depletion of NDRG1 at this time point was also confirmed by Western blotting analysis. NDRG1 expression finally recovered at the stage of remyelination, with immunoreactivity stronger than that in intact nerves. These findings suggest that NDRG1 may play an important role in the terminal differentiation of SCs during nerve regeneration. PMID- 15293231 TI - CD44 overexpression by oligodendrocytes: a novel mouse model of inflammation independent demyelination and dysmyelination. AB - The CD44 transmembrane glycoprotein family has been implicated in cell-cell adhesion and cell signaling in response to components of the extracellular matrix but its role in the nervous system is not understood. CD44 proteins are elevated in Schwann cells and oligodendrocytes following nervous system insults, in inflammatory demyelinating lesions, and in tumors. Here, we tested the hypothesis that elevated CD44 expression influences Schwann cell and oligodendrocyte functions by generating transgenic mice that express CD44 under the control of the 2',3'-cyclic nucleotide-3'-phosphodiesterase (CNPase) promoter. These mice failed to develop peripheral nerve or CNS tumors. However, they did develop severe tremors that were associated with CNS dysmyelination and progressive demyelination. Loss of CNS myelin was not due to alterations in early oligodendrocyte precursor differentiation, proliferation, or survival. Myelination in the PNS appeared normal. In no instance was there any evidence of an inflammatory response that could account for the loss of CNS myelin. These findings suggest that CNPase-CD44 mice are a novel model for noninflammatory progressive demyelinating disease and support a potential role for CD44 proteins expressed by glial cells in promoting demyelination. PMID- 15293232 TI - Unique distributions of the gap junction proteins connexin29, connexin32, and connexin47 in oligodendrocytes. AB - Oligodendrocytes of adult rodents express three different connexins: connexin29 (Cx29), Cx32, and Cx47. In this study, we show that Cx29 is localized to the inner membrane of small myelin sheaths, whereas Cx32 is localized on the outer membrane of large myelin sheaths; Cx29 does not colocalize with Cx32 in gap junction plaques. All oligodendrocytes appear to express Cx47, which is largely restricted to their perikarya. Cx32 and Cx47 are colocalized in many gap junction plaques on oligodendrocyte somata, particularly in gray matter. Cx45 is detected in the cerebral vasculature, but not in oligodendrocytes or myelin sheaths. This diversity of connexins in oligodendrocytes (in different populations of cells and in different subcellular compartments) likely reflects functional differences between these connexins and perhaps the oligodendrocytes themselves. PMID- 15293233 TI - Interleukin-10 attenuates production of HSV-induced inflammatory mediators by human microglia. AB - Infection of the central nervous system (CNS) with herpes simplex virus (HSV)-1 initiates a rapidly progressive, necrotizing, and fatal encephalitis in humans. Even with the advent of antiviral therapy, effective treatments for HSV-1 brain infection are limited because the cause of the resulting neuropathogenesis is not completely understood. We previously reported that human microglial cells, while nonproductively infected, respond to HSV-1 by producing robust amounts of pro inflammatory mediators, such as tumor necrosis factor(TNF), interleukin (IL) 1beta, CCL5 (RANTES), and CXCL10 (IP-10). Although initiation of immune responses by glial cells is an important protective mechanism in the CNS, unrestrained inflammatory responses may result in irreparable brain damage. To elucidate the potential immunomodulatory role of anti-inflammatory cytokines, we investigated the effects of IL-4, IL-10, and transforming growth factor (TGF)-beta on microglial cell cytokine and chemokine production in response to HSV-1. Results from these studies demonstrated a consistent IL-10-mediated suppression of TNF alpha (60% +/- 2%), IL-1beta (68% +/- 3%), CCL5 (62 +/- 4%), but not CXCL10 production by HSV-1-infected microglial cells. This inhibition was associated with decreased HSV-1-induced activation of NF-kappaB. These results suggest that IL-10 has the ability to regulate microglial cell production of immune mediators and thereby, dampen the pro-inflammatory response to HSV-1. PMID- 15293234 TI - Activation of P2 nucleotide receptors stimulates acid efflux from astrocytes. AB - Acidification of the extracellular fluid modulates neurotransmission and ischemic injury in brain. The purpose of the present study was to investigate the effects of purine and pyrimidine transmitters on acid efflux from brain astrocytes. Using RT-PCR, we detected transcripts for the following nucleotide receptors in rat primary astrocyte cultures: P2X1, P2X2, P2X3, P2X4, P2X6, P2X7, P2Y1, P2Y2, P2Y4, and P2Y6. Adenosine 5'-triphosphate (ATP) was found to rapidly induce a biphasic increase in acid efflux, monitored by microphysiometry, consisting of an initial transient and a sustained plateau. Compared with ATP, the P2Y agonist uridine 5' triphosphate (UTP) induced a much smaller initial response but an equal plateau. The poorly hydrolyzable ATP analogue ATPgammaS caused the same initial response as did ATP, but a much smaller plateau, suggesting that the latter phase was due to extracellular degradation of nucleotides. The P2 receptor antagonist, suramin, blocked stimulation of acid efflux by ATP. Removal of extracellular glucose or elevation of extracellular K+ decreased the basal rate of acid efflux but not the stimulation induced by ATP. Inhibition of Na+/H+ exchange by cariporide suppressed the initial phase of ATP-stimulated acid efflux. The intracellular Ca2+ chelator bisaminophenoxyethane- tetraacetic acid (BAPTA) lowered basal acid efflux and abolished the initial phase of the response to ATP. In conclusion, ATP acts through P2 nucleotide receptors on astrocytes to stimulate the Ca2+ dependent efflux of protons, mediated in part by activation of Na+/H+ exchange. The resulting acidification of the extracellular fluid may serve as an intercellular signal in brain. PMID- 15293235 TI - Lesional RhoA+ cell numbers are suppressed by anti-inflammatory, cyclooxygenase inhibiting treatment following subacute spinal cord injury. AB - Inhibition of the small GTPase RhoA or its downstream target Rho-associated coiled kinase (ROCK) has been shown to promote axon regeneration and to improve functional recovery following spinal cord injury (SCI) in the adult rat. RhoA has also been implicated in delayed secondary injury pathophysiology, such as free radical formation and loss of endothelial integrity leading to edema formation. In the present report, we have analyzed the effect of the central nervous system (CNS) permissive, putatively neuroprotective, anti-inflammatory cyclooxygenase-1/ 2 (COX-1/-2) inhibitor indomethacin in CNS effective dosage (2 mg/kg/day) on lesional RhoA expression following subacute spinal cord injury. In control rats receiving vehicle alone, RhoA+ cells accumulate at the lesion site (Th8). At day 3 following SCI, the RhoA+ cellular composition is composed prevailingly of microglia/macrophages and polymononuclear granulocytes, but few reactive astrocytes. In contrast, in the verum group, lesional numbers of RhoA cells were reduced by indomethacin treatment by more than 60% (P < 0.0001). Inflammation dependent RhoA expression accessible by cyclooxygenase inhibition proposes an immune-related mechanism. Our results identify COX blockers as candidates for a safe, synergistic, adjuvant treatment option in combination with cell-specific approaches to Rho inactivation, effectively minimizing the pool of RhoA+ cells at the lesion site following SCI. PMID- 15293239 TI - Ozone in arteriosclerotic plaques: searching for the "smoking gun". PMID- 15293240 TI - The Bellus-Claisen rearrangement. AB - Among the reactions available to synthetic chemists for the construction of new C -C bonds, the Claisen rearrangement is one of the most powerful, elegant, and well-characterized methods. A genuinely new variant, the Bellus-Claisen rearrangement came to light a quarter of a century ago: The reaction of an allylic ether, thioether, or amine with a ketene leads through a [3,3] sigmatropic bond reorganization of a zwitterionic intermediate to an E unsaturated ester, thioester, or amide. When applied to cyclic allylic substrates, a ring-enlargement by four carbon atoms in one step provides medium ring unsaturated E-configured lactones, thiolactones, and lactams. The scope of the Bellus-Claisen rearrangement and the optimum reaction conditions will be discussed in this Minireview. PMID- 15293241 TI - Sulfotransferases: structure, mechanism, biological activity, inhibition, and synthetic utility. AB - The sulfonation (also known as sulfurylation) of biomolecules has long been known to take place in a variety of organisms, from prokaryotes to multicellular species, and new biological functions continue to be uncovered in connection with this important transformation. Early studies of sulfotransferases (STs), the enzymes that catalyze sulfonation, focused primarily on the cytosolic STs, which are involved in detoxification, hormone regulation, and drug metabolism. Although known to exist, the membrane-associated STs were not studied as extensively until more recently. Involved in the sulfonation of complex carbohydrates and proteins, they have emerged as central players in a number of molecular-recognition events and biochemical signaling pathways. STs have also been implicated in many pathophysiological processes. As a result, much interest in the complex roles of STs and in their targeting for therapeutic intervention has been generated. Progress in the elucidation of the structures and mechanisms of sulfotransferases, as well as their biological activity, inhibition, and synthetic utility, are discussed in this Review. PMID- 15293242 TI - A DNA-based machine that can cyclically bind and release thrombin. PMID- 15293243 TI - An autonomous DNA nanomotor powered by a DNA enzyme. PMID- 15293244 TI - Multivalency in the gas phase: the study of dendritic aggregates by mass spectrometry. PMID- 15293245 TI - New oxidation states and defect chemistry in the pyrochlore structure. PMID- 15293246 TI - Exploiting the joint action of chemical shielding and heteronuclear dipolar interactions to probe the geometries of strongly hydrogen-bonded silanols. PMID- 15293247 TI - The pristine oil/water interface: surfactant-free hydroxide-charged emulsions. PMID- 15293248 TI - An unusual cyclization in a bis(cysteinyl-S) diiron complex related to the active site of Fe-only hydrogenases. PMID- 15293249 TI - Highly active, binary catalyst systems for the alternating copolymerization of CO2 and epoxides under mild conditions. PMID- 15293250 TI - A dodecameric water cluster built around a cyclic quasiplanar hexameric core in an organic supramolecular complex of a cryptand. PMID- 15293251 TI - A straightforward and mild synthesis of functionalized 3-alkynoates. PMID- 15293252 TI - Crystal structure of a Dewar benzene derivative formed from fluoro(triisopropylsilyl)acetylene. PMID- 15293253 TI - Cyclometalated ruthenium(II) complexes as highly active transfer hydrogenation catalysts. PMID- 15293254 TI - An unusually fast nucleophilic aromatic displacement reaction: the gas-phase reaction of fluoride ions with nitrobenzene. PMID- 15293255 TI - Design of a sequence-specific DNA bisintercalator. PMID- 15293256 TI - Preparation and reactivity of 1,3,5,7-tetrakis[4 (diacetoxyiodo)phenyl]adamantane, a recyclable hypervalent iodine(III) reagent. PMID- 15293257 TI - A novel tin-free procedure for alkyl radical reactions. PMID- 15293258 TI - Total synthesis of the salicylate enamide macrolide oximidine III: application of relay ring-closing metathesis. PMID- 15293259 TI - Palladium-catalyzed synthesis of N-aryl pyrrolidines from gamma-(N-Arylamino) alkenes: evidence for chemoselective alkene insertion into Pd--N bonds. PMID- 15293261 TI - Novel analogues of sydnone: synthesis, characterization and antibacterial evaluation. AB - New sydnone derivatives bearing a substituted phenyl ring at the 3-position have been synthesized. Two separate series of 3-(carboxyphenyl)sydnone derivatives have been prepared by cyclization of the corresponding N-nitroso-N (carboxyphenyl)-glycine 3. The obtained 3-(carboxyphenyl)sydnones 4 were subjected to a series of different chemical reactions on the carboxylic acid group. Compound 5, the potassium salt of 4a, was reacted with alpha chloroacetanilide derivatives 6 to give the corresponding esters 7. On the other hand, the acid hydrazide 9 was condensed with different aromatic aldehydes to give the corresponding arylidene derivatives 10. The synthesized compounds were tested for their antibacterial activities against both gram-positive and gram negative organisms. Some of the test compounds exhibited high activity; among them, 10d is considered to be a lead compound possessing high broad-spectrum antibacterial activity. PMID- 15293262 TI - Synthesis and cytostatic properties of some 6H-Indolo[2, 3-b][1, 8]naphthyridine derivatives. AB - The new and efficient synthesis of the title heterocyclic ring system is described starting from suitable 2-chloro-1, 8-naphthyridines. The synthesized 6H indolo[2, 3-b][1, 8]naphthyridine derivatives were tested in vitro on 55 tumor cell lines for their anticancer properties. The presence of the acetylamino moiety at position 3 in the main ring system proved to be crucial for the cytostatic activity of this class of compounds. PMID- 15293263 TI - QSAR analysis of some fused pyrazoles as selective cyclooxygenase-2 inhibitors: a Hansch approach. AB - Quantitative structure activity relationships (QSAR) for two unique series of centrally fused pyrazole ring systems have been studied for selective cyclooxygenase-2 inhibitory activity. Several statistically significant QSAR models were developed and suggest that hydrophobicity of entire molecules and a fluorine atom substitution at position 8 of the non benzene sulphonyl ring fused with central pyrazole core of series 1 compounds is crucial for improved COX-2 selectivity. Various structural and physicochemical stipulations to improve the inhibitory activities of the enzymes among individual series of compounds are also discussed. The conclusions derived may serve as an example to advance the design of new selective COX-2 inhibitors. PMID- 15293264 TI - Synthesis and analgesic and antiinflammatory activity of methyl 6-substituted 3(2h)-pyridazinone-2-ylacetate derivatives. AB - A series of methyl 6-substituted-3(2H)-pyridazinone-2-ylacetates 9 were synthesized and their analgesic and anti-inflammatory effects were evaluated in the phenylbenzoquinone-induced writhing test (PBQ test) and carrageenan-induced paw edema method, respectively. Side effects of the compounds were examined on gastric mucosa. None of the compounds showed gastric ulcerogenic effect compared with reference nonsteroidal anti-inflammatory drugs. Methyl 6-(4-(4 fluorophenyl)piperazine)-3(2H)-pyridazinone-2-ylacetate 9e was found to be more active than acetylsalicylic acid (ASA). Methyl 6-(4-(2-ethoxyphenyl)piperazine) 3(2H)-pyridazinone-2-ylacetate 9c has shown an anti-inflammatory activity as compared to the standard compound indometacin at the carrageenan-induced paw edema method.A significant dependence of the anti-inflammatory effect on the substituents has been observed. The pharmacological study of these compounds confirms that modification of the chemical group at the position 6 of the 3(2H) pyridazinone system influences analgesic and anti-inflammatory activities. The structures of these new pyridazinone derivatives were confirmed by their IR and (1)H-NMR spectra and elemental analysis. PMID- 15293265 TI - Chiral resolution of some piperidine 2, 6-dione drugs by high performance liquid chromatography on Kromasil CHI-DMB column. AB - Kromasil CHI-DMB chiral stationary phase (CSP) is used to resolve some piperidine 2, 6-diones under normal mobile phase mode. Base-line separation was achieved using hexane and dioxane (90 : 10 v/v) for aminoglutethimide (AG), pyridoglutethimide (PG), cyclohexylaminoglutethimide (ChAG), thioglutethimide (TG), and N-acetyl aminoglutethimide (AAG). The chromatographic parameters are reported. The chiral recognition mechanisms between the analytes and the CSP depend primarily on hydrogen bonding and/or (pi-pi) interactions. PMID- 15293266 TI - Synthesis and antiviral activities of novel 2', 4'- or 3', 4'-doubly branched carbocyclic nucleosides as potential antiviral agents. AB - In this study, a series of 2', 4-' or 3', 4'-doubly branched carbocyclic nucleosides (11, 12, 19, and 20) were synthesized from simple acyclic ketone derivatives as starting materials. The installation of the 4'-quaternary carbon needed was carried out using a [3, 3]-sigmatropic rearrangement. In addition, the introduction of a methyl group in the 2'- or 3'-position was accomplished by either Grignard reaction or Horner-Wadsworth-Emmons reaction with triethyl-2 phosphonopropionate, respectively. Bis-vinyl was successfully cyclized using a Grubbs' catalyst II. The natural bases (adenine, cytosine) were coupled efficiently using a Pd(0) catalyst. Although all the synthesized compounds were assayed against several viruses, only the cytosine analogue showed moderate antiviral activity against the human cytomegalovirus. PMID- 15293267 TI - The preclinical detection of Parkinson's disease: ready for prime time? PMID- 15293268 TI - Poison and antidote: a novel model to study pathogenesis and therapy of LHON. PMID- 15293269 TI - Idiopathic hyposmia as a preclinical sign of Parkinson's disease. AB - Olfactory dysfunction is an early and common symptom in Parkinson's disease (PD). In an effort to determine whether otherwise unexplained (idiopathic) olfactory dysfunction is associated with an increased risk of developing PD, we designed a prospective study in a cohort of 361 asymptomatic relatives (parents, siblings, or children) of PD patients. A combination of olfactory detection, identification, and discrimination tasks was used to select groups of hyposmic (n = 40) and normosmic (n = 38) individuals for a 2-year clinical follow-up evaluation and sequential single-photon emission computed tomography (SPECT), using [123I]beta-CIT as a dopamine transporter ligand, to assess nigrostriatal dopaminergic function at baseline and 2 years from baseline. A validated questionnaire, sensitive to the presence of parkinsonism, was used in the follow up of the remaining 283 relatives. Two years from baseline, 10% of the individuals with idiopathic hyposmia, who also had strongly reduced [123I]beta CIT binding at baseline, had developed clinical PD as opposed to none of the other relatives in the cohort. In the remaining nonparkinsonian hyposmic relatives, the average rate of decline in dopamine transporter binding was significantly higher than in the normosmic relatives. These results indicate that idiopathic olfactory dysfunction is associated with an increased risk of developing PD of at least 10%. PMID- 15293270 TI - SOD2 gene transfer protects against optic neuropathy induced by deficiency of complex I. AB - Mutations in genes encoding the NADH ubiquinone oxidoreductase, complex I of the respiratory chain, cause a diverse group of diseases. They include Leber hereditary optic neuropathy, Leigh syndrome, and mitochondrial encephalomyopathy with lactic acidosis and stroke-like episodes. There is no effective treatment for these or any other mitochondrial disorder. Using a unique animal model of severe complex I deficiency induced by ribozymes targeted against a critical complex I subunit gene (NDUFA1), we attempted rescue of the optic nerve degeneration associated with Leber hereditary optic neuropathy. We used adenoassociated virus to deliver the human gene for SOD2 to the visual system of disease-induced mice. Relative to mock infection, SOD2 reduced apoptosis of retinal ganglion cells and degeneration of optic nerve fibers, the hallmarks of this disease. Rescue of this animal model supports a critical role for oxidative injury in disorders with complex I deficiency and shows that a respiratory deficit may be effectively treated in mammals, thus offering hope to patients. PMID- 15293271 TI - Interferon-beta stabilizes barrier characteristics of brain endothelial cells in vitro. AB - Multiple sclerosis (MS) is accompanied by a breakdown of the blood-brain barrier (BBB) leading to edema formation and aggravation of the disease. Interferon-beta (IFN-beta) has been approved for the treatment of MS and besides its immunomodulatory effects has been demonstrated to lead to a stabilization of BBB integrity in vivo. To investigate whether human recombinant IFN-beta exerts direct effects on the BBB, we used an in vitro BBB model in which brain endothelial cells in coculture with astrocytes form a tight permeability barrier for 3H-inulin and 14C-sucrose. Removal of the astrocytes from the coculture or alternatively addition of histamine resulted in an increased paracellular permeability for small tracers across the brain endothelial cell monolayer. Strikingly, in the presence of IFN-beta, permeability increase under both conditions was inhibited. Permeability changes were accompanied by minor changes in the staining for tight junction-associated proteins in brain endothelial cell monolayers. Taken together, our data demonstrate a direct stabilizing effect of IFN-beta on BBB cerebral endothelial cells in vitro that might significantly contribute to the beneficial effects of IFN-beta treatment in MS in vivo. PMID- 15293272 TI - Influence of somatosensory input on motor function in patients with chronic stroke. AB - In healthy volunteers, reduction of somatosensory input from one hand leads to rapid performance improvements in the other hand. Thus, it is possible that reduction of somatosensory input from the healthy hand can influence motor function in the paretic hand of chronic stroke patients with unilateral hand weakness. To test this hypothesis, we had 13 chronic stroke patients perform motor tasks with the paretic hand and arm during cutaneous anesthesia of the healthy hand and healthy foot in separate sessions. Performance of a finger tapping task, but not a wrist flexion task, improved significantly with anesthesia of the hand, but not the foot. This effect progressed with the duration of anesthesia and correlated with baseline motor function. We conclude that cutaneous anesthesia of the healthy hand elicits transient site-specific improvements in motor performance of the moderately paretic hand in patients with chronic stroke, consistent with interhemispheric competition models of sensorimotor processing. PMID- 15293273 TI - Functional implications of a novel EA2 mutation in the P/Q-type calcium channel. AB - Episodic ataxia type 2 (EA2) is an autosomal dominant condition characterized by paroxysmal attacks of ataxia, vertigo, and nausea, typically lasting minutes to days in duration. These symptoms can be prevented or significantly attenuated by the oral administration of acetazolamide; however, the mechanism by which acetazolamide ameliorates EA2 symptoms is unknown. EA2 typically results from nonsense mutations in the CACNA1A gene that encodes the alpha1A (Cav2.1) subunit of the P/Q-type calcium (Ca2+) channel. We have identified a novel H1736L missense mutation in the CACNA1A gene associated with the EA2 phenotype. This mutation is localized near the pore-forming region of the P/Q-type Ca2+ channel. Functional analysis of P/Q-type channels containing the mutation show that the H1736L alteration affects several channel properties, including reduced current density, increased rate of inactivation, and a shift in the voltage dependence of activation to more positive values. Although these findings are consistent with an overall loss of P/Q-type channel function, the mutation also caused some biophysical changes consistent with a gain of function. We also tested the direct effect of acetazolamide on both wild-type and H1736L mutated P/Q-type channels and did not observe any direct action on channel properties of this pharmacological agent used to treat EA2 patients. PMID- 15293274 TI - Folic acid supplementation enhances repair of the adult central nervous system. AB - Folic acid supplementation has proved to be extremely effective in reducing the occurrence of neural tube defects (NTDs) and other congenital abnormalities in humans, suggesting that folic acid can modulate key mechanisms for growth and differentiation in the central nervous system (CNS). To prevent NTDs, however, supplemental folate must be provided early in gestation. This suggests that the ability of folic acid to activate growth and differentiation mechanisms may be confined to the early embryonic period. Here, we show that folic acid can enhance growth and repair mechanisms even in the adult CNS. Using lesion models of CNS injury, we found that intraperitoneal treatment of adult rats with folic acid significantly improves the regrowth of sensory spinal axons into a grafted segment of peripheral nerve in vivo. Regrowth of retinal ganglion cell (RGC) axons into a similar graft also was enhanced, although to a smaller extent than spinal axons. Furthermore, folic acid supplementation enhances neurological recovery from a spinal cord contusion injury, showing its potential clinical impact. The results show that the effects of folic acid supplementation on CNS growth processes are not restricted to the embryonic period, but can also be effective for enhancing growth, repair, and recovery in the injured adult CNS. PMID- 15293275 TI - An anti-ganglioside antibody-secreting hybridoma induces neuropathy in mice. AB - Immune responses against gangliosides are strongly implicated in the pathogenesis of some variants of Guillain-Barre syndrome (GBS). For example, IgG antibodies against GM1, GD1a, and related gangliosides are frequently present in patients with post-Campylobacter acute motor axonal neuropathy (AMAN) variant of GBS, and immunization of rabbits with GM1 has produced a model of AMAN. However, the role of anti-ganglioside antibodies in GBS continues to be debated because of lack of a passive transfer model. We recently have raised several monoclonal IgG anti ganglioside antibodies. We passively transfer these antibodies by intraperitoneal hybridoma implantation and by systemic administration of purified anti ganglioside antibodies in mice. Approximately half the animals implanted with an intraperitoneal clone of anti-ganglioside antibody-secreting hybridoma developed a patchy, predominantly axonal neuropathy affecting a small proportion of nerve fibers. In contrast to hybridoma implantation, passive transfer with systemically administered anti-ganglioside antibodies did not cause nerve fiber degeneration despite high titre circulating antibodies. Blood-nerve barrier studies indicate that animals implanted with hybridoma had leaky blood-nerve barrier compared to mice that received systemically administered anti-ganglioside antibodies. Our findings suggest that in addition to circulating antibodies, factors such as antibody accessibility and nerve fiber resistance to antibody-mediated injury play a role in the development of neuropathy. PMID- 15293276 TI - PGE2 receptors rescue motor neurons in a model of amyotrophic lateral sclerosis. AB - Recent studies suggest that the inducible isoform of cyclooxygenase, COX-2, promotes motor neuron loss in rodent models of ALS. We investigated the effects of PGE2, a principal downstream prostaglandin product of COX-2 activity, on motor neuron survival in an organotypic culture model of ALS. We find that PGE2 paradoxically protects motor neurons at physiological concentrations in this model. PGE2 exerts its downstream effects by signaling through a class of four distinct G-protein-coupled E-prostanoid receptors (EP1-EP4) that have divergent effects on cAMP. EP2 and EP3 are dominantly expressed in ventral spinal cord in neurons and astrocytes, and activation of these receptor subtypes individually or in combination also rescued motor neurons. The EP2 receptor is positively coupled to cAMP, and its neuroprotection was mimicked by application of forskolin and blocked by inhibition of PKA, suggesting that its protective effect is mediated by downstream effects of cAMP. Conversely, the EP3 receptor is negatively coupled to cAMP, and its neuroprotective effect was blocked by pertussis toxin, suggesting that its protective effect is dependent on Gi-coupled heterotrimeric signaling. Taken together, these data demonstrate an unexpected neuroprotective effect mediated by PGE2, in which activation of its EP2 and EP3 receptors protected motor neurons from chronic glutamate toxicity. PMID- 15293277 TI - Novel haplotypes in 17q21 are associated with progressive supranuclear palsy. AB - Progressive supranuclear palsy (PSP) and corticobasal degeneration (CBD) are sporadic neurodegenerative diseases presenting as atypical parkinsonian disorders, characterized by the presence of tau-positive neurofibrillary tangles. Recently, an extended haplotype (H1E) of 787.6 kb that comprises several genes including MAPT showed increased association with PSP. The objective of this study was to determine the size of the H1E haplotype associated with PSP and CBD in different populations and to identify specific subhaplotypes in the background of H1E haplotype. Nineteen single nucleotide polymorphisms (SNPs) in the 17q21 region were genotyped in two case-control samples. The SNPs that were associated with higher risk for the disease in the homozygous state delimit a region of more that 1 Mb. Haplotype analyses in the Spanish sample showed that the most frequent haplotype found among the patients (H1E'), which extends 1.04 Mb and contains several genes such as MAPT, CRHR1, IMP5, Saitohin, WTN3, and NSF. A specific subhaplotype (H1E'A) was present in 16% of PSP patients but was not observed in the controls. Furthermore, the H2E'A haplotype, was rarely present in the disease group suggesting that it plays a protective role. The identification of these specific subhaplotypes that modify risk for PSP/CBD supports the hypothesis that a pathogenic allele exists in a subgroup of PSP patients. PMID- 15293278 TI - Adaptation of the attention network in human immunodeficiency virus brain injury. AB - Human immunodeficiency virus (HIV)-positive patients commonly have attention and concentration problems. However, it remains unclear how HIV infection affects the attention network. Therefore, blood oxygenation level dependent functional magnetic resonance imaging (BOLD-fMRI) was performed in 36 subjects (18 HIV and 18 seronegative [SN] controls) during a set of visual attention tasks with increasing levels of attentional load. Compared with SN controls, HIV subjects showed similar task performance (accuracies and reaction times) but decreased activation in the normal visual attention network (dorsal parietal, bilateral prefrontal, and cerebellar regions) and increased activation in adjacent or contralateral brain regions. Cognitive performance (assessed with NPZ-8), CD4, and viral load all correlated with activated BOLD signals in brain regions that activated more in HIV subjects. Furthermore, HIV subjects activated more than SN controls in brain regions that showed load-dependent increase in activation (right prefrontal and right parietal regions) but less in regions that showed a saturation effect with increasing load. These findings suggest that HIV associated brain injury leads to reduced efficiency in the normal attention network, thus requiring reorganization and increased usage of neural reserves to maintain performance during attention-requiring tasks. Exceeding the brain reserve capacity may lead to attention deficits and cognitive impairment in HIV patients. PMID- 15293279 TI - Role of magnetic resonance imaging within diagnostic criteria for multiple sclerosis. AB - The diagnosis of multiple sclerosis (MS) has been improved in recent decades with the incorporation of paraclinical investigations in diagnostic workup. In the last 15 years, magnetic resonance imaging (MRI) has become an especially valuable tool for supporting MS diagnosis, and specific imaging criteria became fundamental to the guidelines for the diagnosis of MS published in 2001 by an international panel (IP). The new IP criteria include MRI evidence of dissemination in space and time, making it possible to diagnose MS after a single clinical episode. This review considers current evidence concerning the reliability of the new IP criteria for the diagnosis of relapsing-onset MS, discusses strengths and weaknesses of the criteria, and outlines areas which may need modification or should be the focus of future research directed toward improving diagnostic accuracy. It also makes practical recommendations when using MRI and the IP criteria in MS diagnosis, especially in patients with clinically isolated syndromes or atypical presentations. The IP criteria are timely and concrete and introduce an important concept to MS diagnosis. Future modifications, based on emerging evidence, should further facilitate their implementation and improve their accuracy. PMID- 15293280 TI - Fuel utilization in patients with very long-chain acyl-coa dehydrogenase deficiency. AB - Fuel utilization in two adult patients with the myopathic form of very long-chain acyl-CoA dehydrogenase (VLCAD) deficiency and five healthy subjects was investigated with stable isotopes during exercise at 50% of VO2max. The findings indicate that residual VLCAD activity in the patients is sufficient to maintain normal oxidation of fat at rest, but that fat oxidation rate cannot increase above basal levels during exercise. This can cause an energy deficit and intramuscular accumulation of fat intermediates that may induce the exercise induced symptoms. PMID- 15293281 TI - Microstructural white matter changes in carriers of the DYT1 gene mutation. AB - We tested the hypothesis that the DYT1 genotype is associated with a disorder of anatomical connectivity involving primarily the sensorimotor cortex. We used diffusion tensor magnetic resonance imaging (DTI) to assess the microstructure of white matter pathways in mutation carriers and control subjects. Fractional anisotropy (FA), a measure of axonal integrity and coherence, was reduced (p < 0.005) in the subgyral white matter of the sensorimotor cortex of DYT1 carriers. Abnormal anatomical connectivity of the supplementary motor area may contribute to the susceptibility of DYT1 carriers to develop clinical manifestations of dystonia. PMID- 15293282 TI - The factor V G1691A mutation is a risk for porencephaly: A case-control study. AB - This study was initiated to investigate prothrombotic risk factors in children with porencephaly. 76 porencephalic and 76 healthy infants were investigated for factor V (FV) G1691A mutation, factor II G20210A variant, methylenetetrahydrofolate reductase (MTHFR) C677T genotype, lipoprotein (a), protein C, protein S, and antithrombin. Only the FV mutation (p = 0.005) and combinations of two or three different risk factors (p = 0.003) were significantly associated with porencephaly. These data give evidence that the FV G1691A mutation and a combination of prothromboic factors play a major role in the development of childhood porencephaly. PMID- 15293283 TI - Bilateral globus pallidus stimulation for Huntington's disease. AB - Bilateral globus pallidus internus (GPi) deep brain stimulation (DBS) was performed in a patient with Huntington's disease (HD) with severe chorea. Stimulation at 40 and 130 Hz improved chorea. Stimulation at 130 Hz slightly worsened bradykinesia overall, whereas 40 Hz had little effect. A [15O] H2O positron emission tomography showed increased regional cerebral blood flow in motor decision making and execution areas more evident at 40 Hz. Adjustment of stimulation parameters in GPi DBS may have the potential to optimize the motor response in HD, improving chorea without aggravating bradykinesia. PMID- 15293284 TI - Functional significance of S6K overexpression in meningioma progression. AB - One common genetic change in anaplastic meningiomas is amplification of chromosome 17q23 containing the S6 kinase (S6K) gene. We show, for the first time to our knowledge, increased S6K mRNA expression in anaplastic meningiomas compared with benign tumors. To evaluate S6K as a candidate meningioma progression gene, we generated IOMM-Lee human meningioma cell lines overexpressing S6K. Whereas no effect of S6K overexpression on meningioma cell growth, motility, or adhesion was observed in vitro, S6K overexpression resulted in increased tumor size in vivo. Collectively, these results suggest that S6K is functionally important for meningioma progression and may represent a target for future meningioma therapy. PMID- 15293285 TI - Chorein detection for the diagnosis of chorea-acanthocytosis. AB - Chorea-acanthocytosis (ChAc) is a severe, neurodegenerative disorder that shares clinical features with Huntington's disease and McLeod syndrome. It is caused by mutations in VPS13A, which encodes a large protein called chorein. Using antichorein antisera, we found expression of chorein in all human cells analyzed. However, chorein expression was absent or noticeably reduced in ChAc patient cells, but not McLeod syndrome and Huntington's disease cells. This suggests that loss of chorein expression is a diagnostic feature of ChAc. PMID- 15293286 TI - Clinical implications of benign multiple sclerosis: a 20-year population-based follow-up study. AB - In 2001, we followed up all patients from the 1991 Olmsted County Multiple Sclerosis (MS) prevalence cohort. We found that the longer the duration of MS and the lower the disability, the more likely a patient is to remain stable and not progress. This is particularly powerful for patients with benign MS with Expanded Disability Status Scale score of 2 or lower for 10 years or longer who have a greater than 90% chance of remaining stable. This is important because these patients represent 17% of the entire prevalence cohort. These data should assist in the shared therapeutic decision-making process of whether to start immunomodulatory medications. PMID- 15293287 TI - Combined antiretroviral therapy and the incidence of acquired immunodeficiency syndrome-related central nervous system diseases. PMID- 15293289 TI - Evidence for pathogenic heterogeneity in multiple sclerosis. PMID- 15293291 TI - Deep brain stimulation for Parkinson's disease: Potential risk of tissue damage associated with external stimulation--erratum. PMID- 15293292 TI - Risk of tissue damage and deep brain stimulation with external devices: A technical note. PMID- 15293297 TI - Color Doppler imaging of fibroadenomas of the breast with histopathologic correlation. AB - PURPOSE: This study was conducted to characterize the spatial distribution of blood vessels in breast fibroadenomas. METHODS: We performed a prospective study to map the anatomic distribution of the vessels in 29 fibroadenomas of the breast using color Doppler sonography. We categorized the detected vessels according to their location in or on the fibroadenoma, counted the different types of vessels, and tested for correlations between vessel distributions or numbers and histopathologic findings. RESULTS: Blood flow was demonstrated in 24/29 (83%) of fibroadenomas. We found 3 vessel types: feeding vessels, which are prominent vessels leading from the surrounding breast tissue into the fibroadenoma; capsular vessels, which are located within the tissue capsule; and segmental vessels, which are located within the fibrous septa of the fibroadenoma. Capsular and segmental vessels were demonstrated in 23/24 (96%) and 24/24 (100%) of the cases, respectively. Feeding vessels were seen in 12/24 (50%) of the cases. Histopathologic analysis revealed the same location and distribution of the vessels as color Doppler imaging. However, there was no correlation between numbers of vessels counted on sonograms and on histopathologic specimens. CONCLUSIONS: Examination of the vascularity demonstrated on color Doppler imaging helps in the diagnosis of benign breast neoplasms such as fibroadenomas. PMID- 15293298 TI - Role of sonography in diagnosing and staging invasive lobular carcinoma. AB - PURPOSE: The goal of this study was to compare the sensitivity of sonography with that of mammography in the detection of invasive lobular carcinoma (ILC), to identify ILC's typical imaging characteristics, and to further show the important role of ultrasound in the staging and treatment planning of this elusive tumor. METHODS: We identified all patients with ILC seen at our institution from 1998 through 2001; 62 had pathologically proven pure ILC. We retrospectively reviewed and analyzed the sonographic appearances in correlation with mammographic, pathologic, and clinical findings. We reviewed the results of sonographic examinations of the nodal basins and fine-needle aspiration (FNA) of suspicious nodes and correlated them with initial clinical and final pathologic staging. We noted all cases of multicentricity or multifocality and analyzed the relative sensitivity of mammography and sonography according to tumor size. RESULTS: Sonography had a sensitivity of 98% versus 65% for mammography. The most common mammographic pattern was a spiculated mass or architectural distortion (39%). On sonography the most common pattern was a hypoechoic mass with (58%) or without (27%) shadowing. An infiltrative pattern was observed in 13% of the cases. Ultrasound-guided FNA confirmed disease was present in the axillary lymph nodes in 21% of the patients, and sonographic examination of the nodal basins resulted in a change of clinical staging from N0 to N1 in 75% and from N1 to N0 in 30% of the cases. Multicentricity/multifocality was identified sonographically and proved by FNA in 21% of patients. CONCLUSIONS: Sonography has a much higher sensitivity than mammography in detecting ILC and therefore is an important adjunctive tool in the diagnosis of this form of cancer. Routine examination of node-bearing areas in patients with ILC proved useful in refining the disease staging. PMID- 15293299 TI - Sonographic guidance for electron boost planning after breast-conserving surgery. AB - PURPOSE: This study was conducted to determine the feasibility of using sonography for electron boost planning in breast cancer treatment and to define the factors that influence the accuracy and reproducibility of this technique. PATIENTS AND METHODS: Seventy-seven patients underwent 102 sonographic examinations after breast-conserving surgery and before and after radiotherapy. The size of the electron boost field was defined by measuring the postoperative cavity. Reproducibility of the sonographic findings was investigated in 25 of the patients who were examined before and after radiotherapy (at a total dose of 46 50.4 Gy). Depth (distance from the skin surface to the posterior aspect of the postoperative cavity) was measured, and sonographic appearance of the postoperative cavity was evaluated. Type of surgical procedure, time elapsed since surgery, use of systemic therapy, menopausal status, breast size, and radiation dose were investigated for their influence on sonographic appearance and visualization of the postoperative cavity. RESULTS: The postoperative cavity was well visualized in 78% of patients and visualized with some difficulty in 22%. In all but 5 patients, it was hypoechoic and inhomogeneous. The mean depth of the postoperative cavity after radiotherapy was 27 +/- 4 mm. Smaller breast (p < 0.001) and younger age (p < 0.05) were associated with decreased visibility of the postoperative cavity. Sonographic appearance was the same before and after radiotherapy, but the mean difference in postoperative cavity depth between the 2 measurements was 2 mm (range, 0-4 mm). In 43/77 (56%) of the patients, changes in electron energy or in field size were required after sonographic measurement. CONCLUSION: Sonography is a useful and reproducible means of electron boost planning, helping to avoid underdosage of the postoperative cavity and overdosage of normal tissue. PMID- 15293300 TI - Sonoanatomy of the Achilles tendon insertion in children. AB - PURPOSE: The aim of this study was to describe typical age-related sonographic features of the Achilles tendon and calcaneal apophysis in children, providing a reference for the assessment of heel pathologies during the growth period. METHODS: The calcaneal apophysis and Achilles tendon insertion of 100 children 2 months to 18 years old were examined by high-frequency gray-scale and color Doppler sonography along both the longitudinal and transverse planes. The thicknesses of the apophyseal cartilage at the calcaneal tuberosity and of the Achilles tendon were measured. Also, the sonographic appearance of the bone cartilage interface was studied. RESULTS: In children 2 months to 3 years old, the cartilage of the calcaneal tuberosity apophysis was anechoic, with small scattered echoes. In 19 of these 25 children (76%), the echogenic areas contained at least 1 small vessel, visualized on color Doppler sonography. In 15 of 25 children (60%) 4-6 years old, a wavy interface was noted at the junction of the calcaneus and the apophyseal cartilage. CONCLUSIONS: High-frequency sonography can yield reliable information about the bone-cartilage interface and the Achilles tendon insertion site at the calcaneal tuberosity in children. The sonographic features of the normal heel described here may contribute to improved assessment of pathologies in this anatomic region. PMID- 15293301 TI - Sonographic assessment of the liver in children with psoriasis. AB - PURPOSE: The aim of this study was to compare the size and echotexture of the liver in psoriatic and healthy children. METHODS: In 70 psoriatic and 43 healthy children, longitudinal sonograms of the liver were obtained along standardized section planes defined by the anterior axillary line, medioclavicular line, and midline. The livers' size and echotexture were examined and compared between the study groups. RESULTS: The measurements of the liver along the 3 section planes were not significantly different between psoriatic and healthy children. Parenchymal liver echogenicity in psoriatic children was neither decreased nor increased. CONCLUSIONS: No abnormality in size or echotexture of the liver was found in the psoriatic children. PMID- 15293302 TI - Value of sonography in the diagnosis of abnormal vaginal bleeding. AB - Abnormal vaginal bleeding is one of the most common presenting complaints in women of any age seeking gynecologic health care. Two of the most frequently used diagnostic tests to investigate the cause of the bleeding are endometrial biopsy and transvaginal sonography. The most worrisome cause of abnormal bleeding is endometrial carcinoma, yet benign etiologies are far more prevalent, including fibroids, polyps, and endometrial atrophy. Endometrial biopsy and transvaginal sonography have equal sensitivities for carcinoma, but sonography is far more effective in diagnosing benign disease. This article reviews the state-of-the-art in the diagnostic evaluation of abnormal vaginal bleeding and analyzes the data, with emphasis on the prevalence of benign and malignant disease as the basis for determining whether sonography or biopsy is more cost-effective in evaluating women with abnormal vaginal bleeding. PMID- 15293303 TI - Sonographically guided injection of anesthetic for iliopsoas tendinopathy after total hip arthroplasty. AB - We report 2 patients who developed pain in the region of the iliopsoas tendon after undergoing total hip arthroplasty. The pain was temporarily relieved by sonographically guided injection of steroid and anesthetic and was subsequently treated by surgical release of the tendon. PMID- 15293304 TI - Primary hydatid cyst in the anterior thigh: Sonographic findings. AB - Hydatid cysts rarely involve the musculoskeletal system. We present the case of a 23-year-old man with a primary hydatid cyst between the femur and the quadriceps muscle in his left thigh. No cysts were located in the adjacent femur or quadriceps muscle. Cyst resection with sparing of the surrounding muscles, combined with anthelmintic therapy, was curative. In regions where hydatidosis is endemic, hydatid cysts should be included in the differential diagnosis of any unusual soft-tissue swelling. PMID- 15293305 TI - Primary peripheral T-cell lymphoma in subcutaneous tissue: sonographic findings. AB - We describe the sonographic findings in a case of Lennert's lymphoma, a rare type of peripheral T-cell lymphoma, involving the subcutaneous tissues of the arm. The sonographic appearance was thought to be more helpful than MRI to establish the diagnosis. PMID- 15293306 TI - Enlargement and hypervascularity of both the epididymis and testis do not exclude involvement with lymphoma or leukemia. AB - We present 3 cases of diffuse infiltration of the testes and epididymides by malignant lymphoma and leukemia. Gray-scale and color Doppler sonograms showed diffuse hypoechoic enlargement and hypervascularity of the involved testes and epididymides. The authors emphasize that enlargement and hypervascularity of both the epididymis and testis can be caused by lymphomatous/leukemic involvement and is not always indicative of epididymo-orchitis. PMID- 15293307 TI - Bilateral epidermoid cysts of the testis: sonographic and MRI findings. AB - Epidermoid cysts of the testis are rare benign lesions, without malignant potential, that can be managed conservatively with cyst enucleation and testis sparing surgery. Bilateral epidermoid cysts in the testes have been reported very infrequently. We report the sonographic and MRI findings in a patient who presented with a palpable nodule in only 1 testis, but in whom sonography showed bilateral lesions. On sonography, the cysts had an internal "onion-ring" structure; color Doppler signals were absent. On MRI, the cysts had a laminated appearance, with alternating low- and high-signal-intensity areas on T2-weighted images. These imaging findings and the negative results of laboratory tests for tumor markers suggested the correct diagnosis, and testis-sparing surgery could be performed. PMID- 15293308 TI - Resistance index in veins? PMID- 15293309 TI - Microrough implant surface topographies increase osteogenesis by reducing osteoclast formation and activity. AB - Titanium implant surfaces with rough microtopographies exhibit increased pullout strength in vivo suggesting increased bone-to-implant contact. This is supported by in vitro studies showing that as surface microroughness increases, osteoblast proliferation decreases whereas differentiation increases. Differentiation is further enhanced on microrough surfaces by factors stimulating osteogenesis including 1alpha,25(OH)2D3. Levels of PGE2 and TGF-beta1 are increased in cultures grown on rough microtopographies; this surface effect is enhanced synergistically by 1alpha,25(OH)2D3-treatment. PGE2 and TGF-beta1 regulate osteoclasts as well as osteoblasts, suggesting that surface microtopography may modulate release of other factors from osteoblasts that regulate osteoclasts. To test this hypothesis, we examined the effects of substrate microarchitecture on production of osteoprotegerin (OPG) and receptor activator of nuclear factor kappa B ligand (RANKL), which have been identified as a key regulatory system of bone remodeling. We also examined the production of 1alpha,25(OH)2D3, which regulates osteoblast differentiation and osteoclastogenesis. MG63 osteoblast-like cells were grown on either tissue culture plastic or titanium disks of different surface microtopographies: PT (Ra < 0.2 microm), SLA (Ra = 4 microm), and TPS (Ra = 5 microm). At confluence, cultures were treated for 24 h with 0, 10(-8) M or 10(-7) M 1alpha,25(OH)2D3. RANKL and OPG were determined at the transcriptional level by RT-PCR and real time PCR and soluble RANKL, OPG and 1alpha,25(OH)2D3 in the conditioned media were measured using immunoassay kits. Cell number was reduced on SLA and TPS surfaces and 1alpha,25(OH)2D3 caused further decreases. OPG mRNA levels increased on rougher surfaces and 1alpha,25(OH)2D3 treatment caused a further synergistic increase. While the cells expressed RANKL mRNA, levels were low and independent of surface microtopography. OPG protein was greater when cells were grown on SLA and TPS. 1alpha,25(OH)2D3 increased OPG by 50% on the smooth Ti surface but on SLA, 10(-8) M 1alpha,25(OH)2D3 caused a 100% increase and 10(-7) M 1alpha,25(OH)2D3 increased OPG by 200%. On TPS 10(-7) M 1alpha,25(OH)2D3 increased OPG 350%. Soluble RANKL was not detected in the conditioned media of any of the cultures. 1alpha,25(OH)2D3 was produced endogenously and levels were positively correlated with surface roughness. Thus, on surfaces with rough microtopographies, osteoblasts secrete factors that enhance osteoblast differentiation while decreasing osteoclast formation and activity. PMID- 15293310 TI - Development of multinuclear giant cells during the degradation of Bioglass particles in rabbits. AB - Bioglass particles of the compositions 45s5, 52s, and 55s were implanted in the distal femoral epiphysis of rabbits. Animals were sacrificed at 7, 28, and 84 days postoperatively and specimens investigated using electron microscopy and electron dispersive X-ray analysis. The intention was to correlate the finding of different types of multinuclear giant cells (MNGC) in the center of the implantation bed with earlier hypothesized accumulated particle eluates and changed particle compositions. The distribution of Si, Na, Ca, P, O, S, and Cl throughout the implantation bed was analyzed. Bioglass particles degraded either in Si-rich remnants or in CaP-shells. MNGC of foreign body giant cell type in high numbers as well as of osteoclast-like type at later time intervals in small numbers were found on the surface of Si-rich as well as on Ca- and P-rich particle remnants. Osteoclast-like cells were detected on the particles after transformation in CaP-shells. It is concluded that the formation of different types of MNGC is determined by the composition of the substrate, that is, osteoclast-like cells develop exclusively on resorbable substrates. The absolute number of MNGC depended on the time after implantation and the solubility of the implant. Bone bonding, however, only occurred on Ca- and P-rich surfaces. PMID- 15293311 TI - Macrophage responses to vascular stent coatings. AB - Diamond-like carbon (DLC) films have been proposed as potential coatings for blood-contacting devices. In this study, tetrahedral amorphous carbon (ta-C) films deposited by filtered cathodic vacuum arc system (FCVA) were compared with commercially deposited polyurethane coatings (PU) and uncoated stainless steel samples. X-ray reflectivity (XRR) measurements were performed to check density and thickness of the ta-C coatings, and contact angles measurements were used to assess surface wettability. J774 macrophages were used to assess the cell responses to the materials. Cell number, metabolic activity, and hydrogen peroxide production were measured by using biochemical assays, and the cell attachment and morphology were determined by using scanning electron microscopy and confocal laser scanning microscopy. Wettability measurements showed that of the materials, the stainless steel was the most hydrophilic, and the ta-C coatings were the most hydrophobic. Although the initial attachment and morphology did not appear to be dependent on the wettability, the cell numbers did increase with increasing wettability. Macrophages on the stainless steel samples were the most active in producing hydrogen peroxide. These data show that ta-C samples performed as well as commercial PU-coated samples in blocking cell reactions to the substrate and may prove to be effective coatings for blood contacting materials. PMID- 15293312 TI - Patterns of gene expression in rat bone marrow stromal cells cultured on titanium alloy discs of different roughness. AB - Rat bone marrow stromal cells were cultured on either Ra (0.14 microm) or Ra (5.8 microm) Ti6Al4V discs for 24 or 48 h. Cells on the Ra (0.14 microm) surface showed typical fibroblastic morphology, whereas cells on the Ra (5.8 microm) surface were in clusters with a more epithelial appearance. RNA was extracted from the cells at both time points, and gene expression was analyzed by using a rat gene microarray. At 24 and 48 h, a similar number of genes were both up- and down-regulated at least twofold on the Ra (5.8 microm) surface compared to the Ra (0.14 microm) surface. We analyzed the relative level of specific groups of genes related to bone and cartilage development, cell adhesion and extracellular matrix proteins, transcription factors, bone morphogenetic proteins, phospholipases, and protein kinases. Roughness did not appear to be a specific stimulator of osteogenesis because genes of both the bone and cartilage lineage were up regulated on the Ra (5.8 microm) surface. The most prominent change among transcription factors was up-regulation of Hox 1.4 on the Ra (5.8 microm) surface. Up-regulation of phospholipase A2 and SMAD 4 indicate these genes are also involved in the response of cells to an Ra (5.8 microm) surface. Our data show surface roughness alters the expression of a large number of genes in marrow stromal cells, which are related to multiple pathways of mesenchymal cell differentiation. PMID- 15293313 TI - Transforming growth factor-beta3-loaded microtextured membranes for skin regeneration in dermal wounds. AB - Adverse effects of wound healing, such as excessive scar tissue formation, wound contraction, or nonhealing wounds represent a major clinical issue in today's healthcare. Transforming growth factor (TGF)-beta3 has specifically been implicated in wound healing. Our hypothesis was that local administration of TGF beta3 to excisional dermal wounds would diminish wound contraction and scar formation. Microtextured wound covers, containing different concentrations of TGF beta3, were placed onto full-thickness excisional skin wounds in guinea pigs. Tattooed reference marks were used to quantify wound contraction. Sixty-four male guinea pigs in four study groups (5 ng TGF-beta3, 50 ng TGF-beta3, no growth factor, sham wound) were followed for up to 6 weeks. We analyzed 19 different parameters of wound healing. Results showed that, in some instances, the 50-ng TGF-beta3 group gave less contraction, whereas the 5-ng TGF-beta3 group gave more contraction. These differences confirm that TGF-beta3 has an optimum working concentration, and suggest this concentration to be closer to 50 ng than to 5 ng TGF-beta3. However, only very few significant differences occurred, and thus we conclude that the clinical relevance of our findings is negligible. Earlier studies, reporting clinically improved wound healing by TGF-beta3, could therefore not be confirmed by this study. PMID- 15293314 TI - Injectable microparticle-gel system for prolonged and localized lidocaine release. I. In vitro characterization. AB - Current treatment protocol for postoperative pain is to infuse anesthetic solution around nerves or into the epidural space. This clinical practice is beset by the short duration of the anesthetic effect unless the infusion is continuous. Continuous infusion, however, requires hospitalization of the patients, thereby increasing medical costs. In addition, it also causes systemic accumulation of the drug. We reported herein a novel treatment for the postoperative pain by applying to the surgical site a biodegradable microsphere gel system for prolonged and localized release of encapsulated anesthetic drugs. This lidocaine-containing biodegradable poly(D,L-lactic acid) (PLA) microsphere system, although being established previously by other investigators, was hindered by a burst release and a followed rapid release of the drug within several hours in vitro. In this article, we demonstrated that by a step-by-step modification of the formulation, prolonged release of lidocaine, up to several days in vitro, could be achieved. Differential scanning calorimetry revealed a lower glass transition temperature for these lidocaine-loaded microspheres comparing to that of lidocaine-free microspheres. This decreased Tg explained for the tendency of the lidocaine-loaded microspheres to physically fuse at higher temperatures. In vitro studies showed that microspheres, when loaded with 35% lidocaine, yielded a threefold increase in the degradation rate. The molecular weight of PLA of the drug-loaded microspheres was reduced by 50% within a period of 1 month. Based on the results (of prolonged lidocaine release and rapid PLA microsphere degradation), this lidocaine-loaded PLA microsphere system could offer a simple solution to the treatment of postoperative pain. PMID- 15293315 TI - Bovine serum albumin conformational changes upon adsorption on titania and on hydroxyapatite and their relation with biomineralization. AB - The biocompatibility of implant materials used for substitution of bone tissue depends on its ability to induce the deposition of a hydroxyapatite layer when in contact with body fluids. In previous work, some of the authors found that bovine serum albumin (BSA) promotes calcium phosphate deposition if preadsorbed on hydroxyapatite and retards precipitation if preadsorbed on titania. In the present study, we investigated the adsorption of BSA upon particles of titania and hydroxyapatite in order to understand the different role played by the protein on the mineralization of both biomaterials. The adsorption isotherms were determined and the structural changes induced by adsorption at different surface coverages were investigated by circular dichroism spectroscopy and differential scanning microcalorimetry. At low surface coverages, the adsorbed BSA molecules lost part of their alpha-helix content. However, at high surface coverages, corresponding to the plateau values of the adsorption isotherms, the BSA molecules did not undergo structural rearrangements upon adsorption. In the latter circumstances, the availability of BSA calcium binding sites, which should be responsible for inducing mineralization, depends on the electrostatic interactions between BSA and the sorbent surface. A possible explanation for the different mineralization behavior of hydroxyapatite and titania is advanced. PMID- 15293316 TI - Ultrastructural study of mineralization of a strontium-containing hydroxyapatite (Sr-HA) cement in vivo. AB - The purpose of this study was to investigate the mineralization leading to osseointegration of strontium-containing hydroxyapatite (Sr-HA) bioactive bone cement injected into cancellous bone in vivo. Sr-HA cement was injected into the ilium of rabbits for 1, 3, and 6 months. The bone mineralization area was found to be largest at 3 months, then at 1 month, and smallest at 6 months (p < 0.01) measured with tetracycline labeling. Osseointegration of Sr-HA cement was achieved at 3 months as observed by scanning electron microscopy. A high calcium and phosphorus area was observed at the interface of bone-Sr-HA cement determined by energy-dispersive X-ray analysis. Transmission electron microscopy gave evidence of the mechanism of bone formation. Dissolution of Sr-HA into debris by the bone remodeling process was thought to increase the concentration of calcium and phosphorus at the interface of bone-Sr-HA cement and stimulate bone formation. Crystalline Sr-HA formed an amorphous layer and dissolved into the surrounding solution, then apatite crystallites were precipitated and formed new bone at 3 months. This young bone then becomes mature bone, which bonds tightly to the Sr-HA cement with collagen fibers inserted perpendicularly after 6 months. PMID- 15293317 TI - Electrostatic interactions as a predictor for osteoblast attachment to biomaterials. AB - The present study utilizes zeta (zeta)-potential analysis as an indicator of bonding of osteoblasts and whole bone to various biomaterials. Common metal alloys (316L stainless steel, CoCrMo, and Ti6Al4V) and bioceramics (hydroxyapatite and beta-tricalcium phosphate) used in orthopedic applications were suspended in particulate form in physiologic saline, both as-received and supplemented with bovine serum albumin (BSA). Metal alloys were also treated with NaOH washing to study the effect of such a surface treatment on the zeta potential. The NaOH wash was found to increase the zeta-potential for CoCrMo and Ti6Al4V, but there was a decrease in the magnitude of the zeta-potential for 316L stainless steel. When the metal alloy powders were suspended in BSA-supplemented physiologic saline, the zeta-potential as a function of pH increased, thereby increasing the electronegativity gap and increasing the propensity for bonding between each of the metal alloys and bone. This increase is likely due to matrix proteins in the BSA, which adsorb onto the metal alloy surfaces, promoting bone growth. With the addition of BSA to each bioceramic system, a uniform decrease in zeta-potential was observed. However, the electronegativity gap remained large in each case, maintaining the anticipation of bonding. zeta-Potential analysis is an effective predictor of biomaterial attraction to osteoblasts and bone, providing a useful in vitro method for predicting such interactions. PMID- 15293318 TI - Synthesis and evaluation of novel bioactive composite starch/bioactive glass microparticles. AB - The aim of the development of composite materials is to combine the most desired properties of two or more materials. In this work, the biodegradable character, good controlled-release properties, and natural origin of starch-based biomaterials are combined with the bioactive and bone-bonding properties of bioactive glass (BG). Novel, bioactive composite starch-BG microparticles were synthesized starting from a blend of starch and polylactic acid (50%/50% wt) with BG 45S5 powder using a simple emulsion method. Morphological and chemical characterization showed that these particles exhibited a spherical morphology with sizes up to 350 microm and that BG 45S5 was incorporated successfully into the composite particles. Upon immersion in a solution simulating body fluids, for periods up to 3 weeks, their bioactive nature was confirmed, as a calcium phosphate layer resembling biological apatite was formed onto their surface. The short-term cytotoxicity of these materials was also tested by placing 24-h leachables of the materials extracted in culture medium in contact with a fibroblastic cell line (L929) up to 72 h. At this time period, two biochemical tests--MTT and total protein quantification--were performed. The results showed that these materials are not cytotoxic. These results constitute the basis of future encapsulation studies using bone-acting therapeutic agents such as bone morphogenetic proteins or other bone-relevant factors. The particles developed here may be very useful for applications in which controlled release, degradability, and bone-bonding ability are the main requirements. PMID- 15293319 TI - A new bone-inducing biodegradable porous beta-tricalcium phosphate. AB - A new type of degradable biomaterial with bone-inducing capacity was made by combining porous beta-tricalcium phosphate (beta-TCP) with a delivery system for recombinant human bone morphogenetic protein-2 (rhBMP-2). The BMP delivery system consisted of a block copolymer composed of poly-D,L-lactic acid with random insertion of p-dioxanone and polyethylene glycol (PLA-DX-PEG), a known biocompatible and biodegradable material. The efficacy of this biomaterial in terms of its bone-inducing capacity was examined by ectopic bone formation in the dorsal muscles of the mouse. In the beta-TCP implants coated with the PLA-DX-PEG polymer containing more than 0.0025% (w/w) of rhBMP-2, new ectopic bone tissues with marrow were consistently found on the surface of implants. The radiographic density of beta-TCP was diminished in a time-dependent manner. On histological examination, numerous multinucleated osteoclasts with positive tartrate-resistant acid-phosphatase (TRAP) staining were noted on the surface of the beta-TCP. These experimental results indicate that beta-TCP implants coated with synthetic rhBMP 2 delivery system might provide effective artificial bone-graft substitutes with osteoinductive capacity and biodegradable properties. In addition, this type of biomaterial may require less rhBMP-2 to induce significant new bone mass. PMID- 15293320 TI - Injectable microparticle-gel system for prolonged and localized lidocaine release. II. In vivo anesthetic effects. AB - Current treatment protocols for postoperative pain are beset by either the short duration of the anesthetic effect or requirement of hospitalization of the patients. We reported herein a novel treatment by applying to the surgical site a biodegradable microparticle-gel system for prolonged and localized release of encapsulated anesthetic drugs. In a previous publication, lidocaine-loaded poly(D,L-lactic acid) microspheres were fabricated and their formulations were optimized. In vitro characterization of these lidocaine-loaded microspheres, however, revealed a shortcoming of this system; that is, microspheres tend to fuse physically. Fusion of the microspheres could hinder their clinical applications, as it would clog the needle. In this article, we demonstrated that fabricating microspheres with high molecular weight (approximately 60 KDa) poly(lactic-co-glycolic acid) would increase the glass transition temperature of the microspheres after lidocaine loading, thereby increasing their mechanical stability and eliminating their fusion during storage. Such microspheres containing 31% (w/w) lidocaine in the presence or absence of 25% (w/v) poloxamer 407 gel were then evaluated in vivo by monitoring the sensory and motor functions of the rats after sciatic nerve block, using the previously established hot-plate and weight-bearing testing methods. Results showed that microspheres formulated with poloxamer 407 gel yielded the longest duration of sensory and motor block for a period of approximately 8.5 h, compared to 5 h by microspheres in saline, 5 h by lidocaine in poloxamer 407 gel, and 2 h by lidocaine in saline. This study suggests that the microsphere-gel system containing lidocaine could potentially be applied clinically to the treatment of postoperative pain. PMID- 15293321 TI - Effect of biphasic calcium phosphates on drug release and biological and mechanical properties of poly(epsilon-caprolactone) composite membranes. AB - Poly(epsilon-caprolactone) (PCL) and biphasic calcium phosphate (CaP) composite membranes were prepared for use in tissue regeneration by a novel solvent casting pressing method. An antibiotic drug, tetracycline hydrochloride (TCH), was entrapped within the membranes to investigate the efficacy of the material as a drug delivery system. The CaP powders were varied in amount (0-50 wt %) and in powder characteristics by heat treating at different temperatures, and their effects on the mechanical and biological properties and drug release of the membranes were examined. With CaP addition up to 30 wt %, the elastic modulus of the membranes was enhanced much due to the rigidity of CaP. While the tensile strength and elongation rate decreased gradually with CaP addition because the CaP powders acted as a failure source. The osteoblast-like cells cultured on the CaP-PCL composite membranes exhibited significant improvements in proliferation and alkaline phosphatase (ALP) activity compared to pure PCL and culture plastic control, indicating excellent cell viability and functional activity. The TCH drugs were released from the PCL and CaP-PCL membranes in a similar fashion; an initial burst followed by a reduced release rate. The initial burst effect diminished much by the addition of CaP powders. The CaP addition increased the drug release rate after an initial period, and this was attributed to the high water uptake capacity and dissolution of the CaP containing membranes. Compared to the composite membranes containing heat-treated CaP powders, those with as precipitated ones had higher dissolution and drug releases. These observations on mechanical properties and cellular responses as well as on drug release profiles suggested that the CaP-PCL composite membranes are potentially applicable to tissue regeneration and drug delivery system. PMID- 15293322 TI - Local properties of a functionally graded interphase between cementum and dentin. AB - The study of natural interfaces may provide information necessary to engineer functionally graded biomaterials for bioengineering applications. In this study, the mechanical, structural, and chemical composition variations associated with a region between cementum and dentin were studied with the use of nanoindentation, microindentation, optical microscopy, and Raman microspectroscopy techniques. Three-millimeter-thick transverse sections (N = 5) were obtained from the apical one-third of the roots of sterilized human molars. The samples were ultrasectioned at room temperature with the use of a diamond knife and an ultramicrotome. Longitudinal ground sections of 100 microm thickness were prepared and stained with von Kossa stain to determine the mineralized regions within the molar roots. Raman microspectroscopy was used to determine the relative inorganic content, mainly apatite (PO4(3-)nu1 mode at 960 cm(-1)) and organic content, mainly collagen (C--H stretch at 2940 cm(-1)) between cementum and dentin bulk tissues. The microindentation and nanoindentation results indicated a gradual transition in hardness from cementum to dentin over a width ranging from 100 to 200 microm. However, the variation in hardness data for cementum and dentin by nanoindentation was larger (0.62 +/- 0.21, 0.77 +/- 0.14 GPa) than from microindentation (0.49 +/- 0.03, 0.69 +/- 0.07 GPa). Within the 100 to 200 microm region there was a 10 to 50 microm fibrillar hydrophilic cementum-dentin junction (CDJ) with mechanical properties significantly lower than either the cementum or the dentin side of CDJ. Light microscopy revealed a 100 to 200 microm translucent region between cementum and dentin. Raman microspectroscopy results showed a variation in organic and inorganic composition 80 to 140 microm wide. It was concluded that a morphologically and biomechanically different CDJ lies within a wider cementum-dentin interphase. Hence, cementum, dentin, and the interphase can be classified as a functionally graded dental tissue within the root of a tooth. PMID- 15293323 TI - Cyclic mechanical strain alters tissue-factor activity in rat osteosarcoma cells cultured on a titanium substrate. AB - Tissue factor (TF), a transmembrane glycoprotein, plays a role in the initiation of blood coagulation at sites of vascular injury. Activated products of coagulation may then enhance inflammatory responses. The present investigation assesses the ability of rat osteosarcoma (UMR-106) cells cultured on titanium alloy (Ti6Al4V) to express differential surface TF activity in response to cyclic mechanical strain. Strains ranged from -2000 micro-strain to +2000 micro-strain, and durations from 5, 10, and 20 min per day over 5 days to 24 h continuous stimulation. ROS cells exhibited significant TF activity as demonstrated by the conversion of Factor X to Factor Xa. Strains of +2000 micro-strain with 5-20-min duration exhibited decreased TF activity with duration from 1.4E-04 nM/cell to 8.7E-05 nM/cell. Additionally, ROS cells stimulated with calcium ionophore (A23187) exhibited at least twice the activity of nonstimulated cells. Strains of +1340 micro-strain with 5-20-min duration exhibited an increasing trend with 4.15E-05 nM/cell to 7.38E-05 nM/cell. Strain direction had no significant effect on TF activity. Thus, both mechanical and chemical stimuli induce differential expression of TF activity by ROS cells cultured on Ti6Al4V, a phenomenon that may potentiate or regulate the inflammatory responses associated with the implantation of orthopedic biomaterials. PMID- 15293324 TI - Effect of the difference of bone turnover on peri-titanium implant osteogenesis in ovariectomized rats. AB - High and low bone turnover situations, both of which are typically observed as postmenopausal and senile osteoporosis, were created by ovariectomy (OVX), and then an investigation of whether or not the difference of bone turnover affected peri-titanium (Ti) implant osteogenesis in rats was conducted. Female rats were divided into four groups. The experimental and control groups underwent OVX or sham operations at 15 or 27 weeks of age, as high or low bone turnover groups, respectively. Ti implants were inserted into the tibiae at 30 weeks, then fluorochromes were injected 10 or 20 days after the implantation for histometry. The implants were retained for 30 days and then ground sections were prepared. Afterward, the cortical bone growth rate, bone contact ratio (BCR) of the implant in both the cortical bone area and medullary canal area, and the average trabecular bone thickness around the implant were evaluated. Biochemical markers of bone turnover were also measured. Biochemical measurements indicated both increasing osteocalcin production in OVX rats and decreasing tartrate-resistant acid phosphatase activity in the low-turnover group. Histometrical measurements showed decreasing cortical growth and low BCR in the medullary canal of the low turnover group. The high-turnover group demonstrated BCR as high as that of the control group. There was no significant difference in the average trabecular bone thickness around the implant among the groups. As a result, two types of osteoporotic situations were confirmed and it was shown that the difference of bone turnover was clearly due to the diverse osteogenesis around the Ti implant. PMID- 15293325 TI - Solvent effects on the microstructure and properties of 75/25 poly(D,L-lactide-co glycolide) tissue scaffolds. AB - Poly(lactide-co-glycolide) (PLGA) is used in many biomedical applications because it is biodegradable, biocompatible, and FDA approved. PLGA can also be processed into porous tissue scaffolds, often through the use of organic solvents. A static light scattering experiment showed that 75/25 PLGA is well solvated in acetone and methylene chloride, but forms aggregates in chloroform. This led to an investigation of whether the mechanical properties of the scaffolds were affected by solvent choice. Porous 75/25 PLGA scaffolds were created with the use of the solvent casting/particulate leaching technique with three different solvents: acetone, chloroform, and methylene chloride. Compression testing resulted in stiffness values of 21.7 +/- 4.8 N/mm for acetone, 18.9 +/- 4.2 N/mm for chloroform, and 30.2 +/- 9.6 N/mm for methylene chloride. Permeability testing found values of 3.9 +/- 1.9 x 10(-12) m2 for acetone, 3.6 +/- 1.3 x 10(-12) m2 for chloroform, and 2.4 +/- 1.0 x 10(-12) m2 for methylene chloride. Additional work was conducted to uncouple polymer/solvent interactions from evaporation dynamics, both of which may affect the scaffold properties. The results suggest that solvent choice creates small but significant differences in scaffold properties, and that the rate of evaporation is more important in affecting scaffold microstructure than polymer/solvent interactions. PMID- 15293326 TI - Fracture toughness of steel-fiber-reinforced bone cement. AB - Fractures in the bone-cement mantle (polymethyl methacrylate) have been linked to the failure of cemented total joint prostheses. The heat generated by the curing bone cement has also been implicated in the necrosis of surrounding bone tissue, leading to loosening of the implants. The addition of reinforcements may improve the fracture properties of bone cement and decrease the peak temperatures during curing. This study investigates the changes in the fracture properties and the temperatures generated in the ASTM F451 tests by the addition of 316L stainless steel fibers to bone cement. The influence of filler volume fraction (5-15% by volume) and aspect ratios (19, 46, 57) on the fracture toughness of the acrylic bone cement was assessed. Increasing the volume fraction of the steel fibers resulted in significant increases in the fracture toughness of the steel-fiber reinforced composite. Fracture-toughness increases of up to 2.63 times the control values were obtained with the use of steel-fiber reinforcements. No clear trend in the fracture toughness was discerned for increasing aspect ratios of the reinforcements. There is a decrease in the peak temperatures reached during the curing of the steel-fiber-reinforced bone cement, though the decrease is too small to be clinically relevant. Large increases in the fatigue life of acrylic bone cement were also obtained by the addition of steel fibers. These results indicate that the use of steel fibers may enhance the durability of cemented joint prostheses. PMID- 15293327 TI - Role of carnivores in the accumulation of the Sterkfontein Member 4 hominid assemblage: a taphonomic reassessment of the complete hominid fossil sample (1936 1999). AB - New taphonomic data on the Sterkfontein Member 4 (South Africa) fossil hominid assemblage are presented. The previous estimate of hominid individuals represented in the deposit (45) is increased to 87. New minimum numbers of hominid skeletal elements are provided, and incidences of bone surface damage inflicted by prehistoric biological agents are summarized. The hominid sample from Member 4 is composed predominately of gnathic remains and has a paucity of postcrania. This dearth of postcrania limits, to some extent, inferences about the formation of the Sterkfontein assemblage. However, carnivore tooth marks on some fossil specimens and an overall broad similarity in patterns of skeletal part representation between Sterkfontein and primate bone assemblages created by extant carnivores suggest that carnivores did have some involvement in the accumulation of the fossil hominid assemblage. Thus, this study provides support for the "carnivore-collecting hypothesis" of Brain (Brain [1981] The Hunters or the Hunted? Chicago: University of Chicago Press), which implicates large carnivores as prominent collecting agents of hominid body parts in Sterkfontein Member 4. Evidence of bone surface damage is, however, too scant to make confident inferences about specific carnivore taxon/taxa involved in hominid bone collection at the site. PMID- 15293328 TI - Bony ponticles of the atlas (C1) over the groove for the vertebral artery in humans and primates: polymorphism and evolutionary trends. AB - The aim of this study was to ascertain the distribution in primates of the three possible bony ponticles over the groove for the vertebral artery (ventral, lateral, and dorsal ponticles), in order to attempt to understand the variants observed in humans and to ascertain possible evolutionary trends in primates. The material consisted of 393 atlases of extant nonhuman primates representative of 41 genera, and of 500 human atlases (dried bones of adults). For each atlas, we studied the existence and morphology of the ponticles, and the type of association of these three ponticles on a given side, which are theoretically of eight in number (types A-H). The occurrence of these ponticles varied from complete absence to constant presence, according to the genera and taxa of primates. The presence of each of these ponticles in primates can be interpreted as a primitive or plesiomorphic character, and their absence as a derived or apomorphic character. The strepsirhines-platyrrhines-cercopithecines group, presenting a predominant primitive pattern (type A), appeared to be separated from the colobines-hominoids group, presenting predominant derived patterns (type C in colobines, Pongo pygmaeus, and Pan troglodytes, and the more derived type D in Hylobates, Gorilla gorilla, and Homo sapiens). The last derived stage, corresponding to the disappearance of the three atlantal ponticles (type H), was only observed in some individuals in hominoids. A marked intraspecific polymorphism characterized the hominoids. The presence of lateral and dorsal ponticles in humans appeared to correspond to their persistence within the progressive disappearance of the atlantal ponticles, constituting an evolutionary tendency characteristic of primates and particularly of hominoid evolution. PMID- 15293329 TI - Grasping behavior in tufted capuchin monkeys (Cebus apella): grip types and manual laterality for picking up a small food item. AB - This study investigates prehension in 20 tufted capuchins (Cebus apella) in a reaching task requiring individuals to grasp a small food item fixed to a tray. The aim was twofold: 1) to describe capuchins' grasping techniques in detail, focusing on digit movements and on different areas of contact between the grasping fingers; and 2) to assess the relationship between grip types and manual laterality in this species. Capuchins picked up small food items using a wide variety of grips. In particular, 16 precision grip variants and 4 power grip variants were identified. The most frequently used precision grip involved the distal lateral areas of the thumb and the index finger, while the most preferred kind of power grip involved the thumb and the palm, with the thumb being enclosed by the other fingers. Immature capuchins picked up small food items using power grips more often than precision grips, while adult individuals exhibited no significant preference for either grip type. The analysis performed on the time capuchins took to grasp the food and withdraw it from the tray hole revealed that 1) precision grips were as efficient as power grips; 2) for precision grips, the left hand was faster than the right hand; and 3) for power grips, both hands were equally quick. Hand preference analysis, based on the frequency for the use of either hand for grasping actions, revealed no significant hand bias at group level. Likewise, there was no significant relationship between grip type and hand preference. PMID- 15293330 TI - Compliant walking in primates: elbow and knee yield in primates compared to other mammals. AB - It has been suggested that primates utilize a compliant gait to help reduce peak locomotor stresses on their limbs (Schmitt [1994] J. Hum. Evol. 26:441-458; Schmitt [ 1998] Primate Locomotion, p. 175-200; Schmitt [ 1999] J. Zool. Lond. 248:149-160). However, the components of such a gait, i.e., increased step length, prolonged contact time, and substantial limb yield, have only been documented on a handful of primate species. In order to explore the generality of this claim, elbow and knee angles during walking were documented at touchdown, midstance, and liftoff in a sample of primates, carnivores, marsupials, rodents, and artiodactyls, all under 25 kg. Limb yield was calculated as the change in angle from touchdown to midstance, and re-extension as the change in angle from midstance to liftoff for both forelimbs and hind limbs. Use of a compliant gait (as reflected in significant limb yield) in primates was confirmed for both forelimbs and hind limbs. However, there was variability within primates in the degree of either elbow or knee yield. Surprisingly, marsupials were found to exhibit almost as much elbow yield and even greater knee yield than primates. Carnivores and rodents display a modest amount of limb yield during walking, while artiodactyls appear to display a relatively stiff gait. These data are consistent with the suggestion that the use of a compliant gait to attenuate peak substrate reaction forces may have facilitated the primate invasion of a small branch niche. However, limb compliance (as reflected by elbow or knee yield) does not appear to be exclusive to the primate order. PMID- 15293331 TI - Molar size and shape variations among Asian colobines. AB - Despite decades of research, little is known about morphometric differences within the dentition of Asian colobines. Although some differences, such as the M3 hypoconulid, are often cited as distinct among genera, no comprehensive assessment has been made. The objectives of this study were to document size and shape differences in third molars in eight Asian colobine genera, including Kasi, Nasalis, Presbytis, Pygathrix, Rhinopithecus, Semnopithecus, Simias, and Trachypithecus, and to quantify length differences for all molars in order to understand the potential impact of an occlusal surface area that includes the hypoconulid. To achieve these objectives, the most extensive survey yet published of Asian colobine dentition was conducted. Differences within and among genera for continuous and categorical variables were analyzed using descriptive statistics, chi-square, t-tests, and post hoc comparisons. Results indicate that species of Presbytis differ from all other Asian colobines in both the size and shape of the maxillary and mandibular third molars. Specifically, M3 of Presbytis is relatively shorter in length and less likely to exhibit a hypoconulid optimally positioned for occlusion with M3. Moreover, Presbytis expresses concomitant changes in the maxillary third molar such as absence of a distal shelf and/or a distal cusp with separated crests. We conclude that shape and size changes impact the molar size sequence patterns of Presbytis, setting this genus apart from all other Asian colobine taxa. These differences may reflect variation in overall body size, dietary adaptation, and phylogeny. PMID- 15293332 TI - Demography, life history, and social structure in Propithecus diadema edwardsi from 1986-2000 in Ranomafana National Park, Madagascar. AB - Prosimian lemurs differ fundamentally from anthropoid primates in many traits related to social structure. By exploring the demography of Milne-Edwards' sifakas (Propithecus diadema edwardsi), and comparing it to other well-studied primates, we explore the effect of demographic and life-history factors on social structure. Specifically, we compare lemur survivorship and fertility patterns to two published composite models: one created for New World and another created for Old World monkeys. Using longitudinal data collected on individual Propithecus diadema edwardsi from four study groups from 1986-2000 in Ranomafana National Park, Madagascar, we quantify 1) group composition, 2) birth seasonality, 3) interbirth interval, 4) life-table values, and 5) population growth estimates. The mortality, survivorship, and life-expectancy schedules indicate high infant and juvenile mortality. Fertility remains high until death. The intrinsic rate of increase and net reproductive rate indicate a shrinking population. We suggest that high mortality rather than low fertility causes the observed population decline. While sifaka survivorship closely resembles New World patterns, fertility resembles Old World patterns, i.e., like New World monkeys, few sifakas survive to reproductive age, and those that do, reproduce at a slow rate resembling the Old World pattern. This necessarily impacts social structure. An adult sifaka at the end of her lifespan will have one only daughter who survives to reproductive age, compared to 3.4 for New World or 2.7 for Old World monkeys. Demography limits the formation of large kin-based groups for sifakas, and survivorship and fertility patterns do not easily permit sifakas to form large same-sex family groups. PMID- 15293333 TI - Social and seasonal influences on the reproductive cycle in female mandrills (Mandrillus sphinx). AB - We present 12 years of perineal swelling data for a semifree-ranging colony of mandrills (Mandrillus sphinx), and evaluate the influence of rank, parity, and seasonality on reproductive parameters. Female sexual swellings showed a seasonal pattern, with August the median month of ovulation. Overlapping periovulatory periods did not decrease the likelihood of conception. Females showed their first genital swelling at age 3.6 years (n = 28; range, 3.2-4.6 years), and higher ranking females experienced their first swelling earlier than low-ranking females. Median postpartum amenorrhea (PPA) duration was 208 days (n = 92; range, 74-538 days). PPA was longer in primiparous females than in multiparous females, but PPA duration was unrelated to female rank. Median follicular phase duration was 24 days for the first cycle after parturition (n = 84; range, 12-40 days), shortening to 17 days in subsequent cycles (n = 55; range, 6-39 days). The follicular phase was longer in nulliparous females than in parous females, but was unrelated to female rank. Median cycle length (from one sexual swelling breakdown to the next) was 38 days (n = 57; range, 18-108 days). Eighty-seven percent of conceptions occurred within two cycles, and half of the nulliparous females conceived during their first swelling cycle. Lower-ranking females were more likely to require more cycles to conceive than higher-ranking females. The cycling phase was significantly longer in nulliparous females than in parous females, and was also significantly longer in lower-ranking females than in higher-ranking females. We discuss the influence of provisioning on female reproductive parameters, the influence of parity and rank on the different phases of the interbirth interval, and the evolution of long and variable follicular phases in mandrills. PMID- 15293334 TI - DYS19 and DYS199 loci in a Chilean population of mixed ancestry. AB - The current Chilean population originated from admixture between aboriginal populations (Amerindians) and Spanish conquerors of European origin. Consequently, the unions that gave rise to the Chilean population were chiefly between Spanish males and aboriginal females, and not the converse. To test the hypothesis that the Y chromosome of the Chilean population is mainly of Spanish origin, while the other chromosomes are from mixed (European and aboriginal) origin, we studied the DYS19 and DYS199 loci in two samples. One sample was obtained from a high socioeconomic stratum, while a second sample was from a low stratum. We studied male blood donors (N = 187) from Santiago, the capital of the country. Subjects were typed for the autosomal ABO and Rh (locus D) blood groups, and for the Y-linked DYS19 and the DYS199 loci, reported as Y-chromosome haplotypes. The aboriginal admixture was estimated for each genetic marker. The percentage of aboriginal admixture was 38.17% for the ABO system and 31.28% for the Rh system in the low socioeconomic stratum and 19.22% and 22.5%, respectively, in the high stratum. Y-chromosome haplotype frequencies constructed from the DYS19 and DYS199 loci demonstrated that the main haplotypes were DYS19*14/DYS199 C, as is often the case with many European populations, and DYS19*13/DYS199 C. The aboriginal admixture from Y-haplotype frequencies was estimated to be 15.83% in the low socioeconomic stratum and 6.91% in the high stratum. These values are lower than the values found using autosomal genetic markers, and are consistent with the historical background of the population studied. This study highlights the population genetic consequences of the asymmetric pattern of genome admixture between two ancestral populations (European and Amerindian). PMID- 15293335 TI - The longer you stay, the bigger you get: length of time and language use in the U.S. are associated with obesity in Puerto Rican women. AB - This cross-sectional study examined whether length of time in the U.S., language use, and birthplace (proxy measures of acculturation) were associated with body mass index (BMI) and obesity in a sample of 174 low-income Puerto Rican women from Hartford, Connecticut. The mean BMI for the total sample (N = 174) was 27.39 (S.D. = 5.07), and nearly 34% of the sample was considered obese (BMI > or = 30). There was a statistically significant increase in BMI with length of time in the U.S. (P = 0.012) and these differences were even greater among women born in Puerto Rico (P = 0.003). Moreover, obesity prevalence was highest among women who had been in the U.S. for 10 years or more (40%), as compared to those who had been in the U.S. less than 1 year (29%; P = 0.045). There were no statistically significant associations between language and BMI for the total sample. However, among bilingual speakers born in Puerto Rico, there were significant differences in BMI according to their level of English fluency. Those who spoke fluent or very good English had a significantly higher BMI (mean = 29.72; SD = 4.12) than women whose English was good to not-so-good (mean = 26.8; SD = 5.24; P = 0.016). The findings from this study point to the need for more research on the acculturation process and obesity, in order to design culturally tailored obesity prevention programs. PMID- 15293336 TI - FI catalysts: new olefin polymerization catalysts for the creation of value-added polymers. AB - This contribution reports the discovery and application of phenoxy-imine-based catalysts for olefin polymerization. Ligand-oriented catalyst design research has led to the discovery of remarkably active ethylene polymerization catalysts (FI Catalysts), which are based on electronically flexible phenoxy-imine chelate ligands combined with early transition metals. Upon activation with appropriate cocatalysts, FI Catalysts can exhibit unique polymerization catalysis (e.g., precise control of product molecular weights, highly isospecific and syndiospecific propylene polymerization, regio-irregular polymerization of higher alpha-olefins, highly controlled living polymerization of both ethylene and propylene at elevated temperatures, and precise control over polymer morphology) and thus provide extraordinary opportunities for the syntheses of value-added polymers with distinctive architectural characteristics. Many of the polymers that are available via the use of FI Catalysts were previously inaccessible through other means of polymerization. For example, FI Catalysts can form vinyl terminated low molecular weight polyethylenes, ultra-high molecular weight amorphous ethylene-propylene copolymers and atactic polypropylenes, highly isotactic and syndiotactic polypropylenes with exceptionally high peak melting temperatures, well-defined and controlled multimodal polyethylenes, and high molecular weight regio-irregular poly(higher alpha-olefin)s. In addition, FI Catalysts combined with MgCl(2)-based compounds can produce polymers that exhibit desirable morphological features (e.g., very high bulk density polyethylenes and highly controlled particle-size polyethylenes) that are difficult to obtain with conventionally supported catalysts. In addition, FI Catalysts are capable of creating a large variety of living-polymerization-based polymers, including terminally functionalized polymers and block copolymers from ethylene, propylene, and higher alpha-olefins. Furthermore, some of the FI Catalysts can furnish living-polymerization-based polymers catalytically by combination with appropriate chain transfer agents. Therefore, the development of FI Catalysts has enabled some crucial advances in the fields of polymerization catalysis and polymer syntheses. PMID- 15293337 TI - Metal-catalyzed living radical polymerization: discovery and developments. AB - Control of radical polymerization has been one of the most challenging frontiers in polymerization chemistry. This review presents the discovery of metal catalyzed living radical polymerization and recent developments in the evolution of catalysts in terms of versatility and activity, scope of monomers, controlled polymerization in water, catalyst removal, and precision synthesis of well controlled polymers such as random, block, end-functionalized, and star polymers. PMID- 15293338 TI - Solvent-free low-dimensional polymer electrolytes for lithium-polymer batteries. AB - The development of solvent-free low-dimensional polymer electrolytes intended for use in solvent-free lithium batteries operating at ambient or sub-ambient temperatures is described. The synthetic routes to the amphiphilic polymers I having 5-alkoxy-3,4-phenylene units connected with oligoethoxy segments via polyester-ether or pure polyether links (abbrev. CmOn, m = 12, 16, 18, n = 1-5) and to the copolymers CmO1-CmOn are described. The structures, thermal properties and SAXS long spacings of their complexes with lithium salts (type A) and with long chain n-alkane or alkyl side chain intercalation (type B) are discussed. However, high ambient conductivities (10(-4)-10(-3) S cm(-1)) are observed in type C systems when a second copolymer based on polytetramethylene oxide segments (II) is incorporated as a microphase between the lamellae of I and serving as an ion bridge or "glue". DC polarization between Li electrodes also gives ambient conductivities >/=ca.10(-3) S cm(-1). In type D systems the I/II interface is stabilized by including a copolymer III, promoting high reproducibility in performance. Copolymers I of CmO1-CmO5 having CmO1 in excess give optimum conductivities with low temperature-dependence. This, together with molecular modeling, suggests uncoupled ion mobilities by hopping between small aggregates in the interlamellar spaces. PMID- 15293339 TI - A novel electrochemical approach to the characterization of oxidoreductase reactions. AB - Electrochemical methods based on enzyme-electrochemical reactions have been developed for studying oxidoreductase reactions. The methods measure a current resulting from an oxidoreductase reaction with an electrode serving as a final electron acceptor (or donor) in the reaction. A theoretical equation for the enzyme-electrochemical reaction, called bioelectrocatalysis, is derived, which enables kinetic analysis of the reaction. In combination with spectrophotometry, the electrochemical method provides a method for determining the redox potentials of proteins and enzymes. An alternative method based on bulk electrolysis in a quartz cell for UV-vis spectroscopy has been developed for the measurements of protein redox potentials on a conventional spectrophotometer. The electrochemical methods are applied to kinetic and thermodynamic analyses for the reactions of a variety of enzymes including a newly discovered enzyme, quinohemoprotein amine dehydrogenase (QH-AmDH), and bilirubin oxidase (BOD) [EC 1.3.3.5, from Myrothecium verrucaria], a copper-containing enzyme useful for bioelectrocatalytic O(2) reduction in biofuel cells. The electrochemical method for kinetic analysis has been successfully applied to the analysis of oxidoreductase reactions in vivo, as demonstrated by the reaction of glucose dehydrogenase in Escherichia coli. The advantages of the electrochemical methods are discussed. PMID- 15293340 TI - Electron transport in bipyridinium films. AB - Bipyridinium dications are versatile building blocks for the assembly of functional materials. In particular, their reliable electrochemical response has encouraged the design of electroactive films. Diverse and elegant experimental strategies to coat metallic and semiconducting electrodes with bipyridinium compounds have, in fact, emerged over the past two decades. The resulting interfacial assemblies span from a few nanometers to several micrometers in thickness. They incorporate from a single molecular layer to large collections of entangled polymer chains. They transport electrons efficiently from the electrode surface to the film/solution interface and vice versa. Electron self-exchange between and the physical diffusion of the bipyridinium building blocks conspire in defining the charge transport properties of these fascinating electroactive assemblies. Often, the matrix of electron-deficient bipyridinium dications can be exploited to entrap electron-rich analytes. Electrostatic interactions promote the supramolecular association of the guests with the surface-confined host matrix. Furthermore, chromophoric sites can be coupled to the bipyridinium dications to produce photosensitive arrays capable of harvesting light and generating current. Thus, thorough investigations on the fundamental properties of these functional molecule-based materials can lead to promising applications in electroanalysis and solar energy conversion, while contributing to advances in the basic understanding of electron transport in interfacial assemblies. PMID- 15293342 TI - Can the life span of human marrow stromal cells be prolonged by bmi-1, E6, E7, and/or telomerase without affecting cardiomyogenic differentiation? AB - BACKGROUND: Cell transplantation has recently been challenged to improve cardiac function of severe heart failure. Human mesenchymal stem cells (hMSCs) are multipotent cells that can be isolated from adult marrow stroma, but because of their limited life span, it is difficult to study them further. To overcome this problem, we attempted to prolong the life span of hMSCs and investigate whether the hMSCs modified with cell-cycle-associated genes can differentiate into cardiomyocytes in vitro. METHODS: We attempted to prolong the life span of hMSCs by infecting retrovirus encoding bmi-1, human papillomavirus E6 and E7, and/or human telomerase reverse transcriptase genes. To determine whether the hMSCs with an extended life span could differentiate into cardiomyocytes, 5-azacytidine treated hMSCs were co-cultured with fetal cardiomyocytes in vitro. RESULT: The established hMSCs proliferated over 150 population doublings. On day 3 of co cultivation, the hMSCs became elongated, like myotubes, began spontaneously beating, and acquired automaticity. Their rhythm clearly differed from that of the surrounding fetal mouse cardiomyocytes. The number of beating cardiomyocytes increased until 3 weeks. hMSCs clearly exhibited differentiated cardiomyocyte phenotypes in vitro as revealed by immunocytochemistry, RT-PCR, and action potential recording. CONCLUSIONS: The life span of hMSCs was prolonged without interfering with cardiomyogenic differentiation. hMSCs with an extended life span can be used to produce a good experimental model of cardiac cell transplantation and may serve as a highly useful cell source for cardiomyocytic transplantation. PMID- 15293343 TI - Regeneration of a well-differentiated human airway surface epithelium by spheroid and lentivirus vector-transduced airway cells. AB - BACKGROUND: Following injury to the airway epithelium, rapid regeneration of a functional epithelium is necessary in order to restore the epithelial barrier integrity. In the perspective of airway gene/cell therapy, we analyzed the capacity of human airway epithelial cells cultured as three-dimensional (3-D) spheroid structures to be efficiently transduced on long term by a pseudotyped lentiviral vector. The capacity of the 3-D spheroid structures to repopulate a denuded tracheal basement membrane and regenerate a well-differentiated airway epithelium was also analyzed. METHODS: An HIV-1-derived VSV-G pseudotyped lentiviral vector encoding the enhanced green fluorescent protein (eGFP) was used. Airway epithelial cells were isolated from mature human fetal tracheas and airway xenografts, cultured as 3-D spheroid structures, and either transduced at multiplicity of infection (MOI) 10 and 100 or assayed in an ex vivo and in vivo model to evaluate their regeneration capacity. RESULTS: An in vivo repopulation assay in SCID-hu mice with transduced isolated fetal airway epithelial cells shows that lentiviral transduction does not alter the airway reconstitution. Transduction of the 3-D spheroid structures shows that 12% of cells were eGFP positive for up to 80 days. In ex vivo and in vivo assays (NUDE-hu mice), the 3-D spheroid structures are able to repopulate denuded basement membrane and reconstitute a well-differentiated human airway surface epithelium. CONCLUSIONS: The efficient and long-term lentiviral transduction of 3-D spheroid structures together with their capacity to regenerate a well-differentiated mucociliary epithelium demonstrate the potential relevance of these 3-D structures in human airway gene/cell therapy. PMID- 15293344 TI - Combined alpha tumor necrosis factor gene therapy and engineered dendritic cell vaccine in combating well-established tumors. AB - BACKGROUND: Although current immunotherapeutic strategies including adenovirus (AdV)-mediated gene therapy and dendritic cell (DC) vaccine can all stimulate antitumor cytotoxic T lymphocyte (CLT) responses, their therapeutic efficiency has still been limited to generation of prophylactic antitumor immunity against re-challenge with the parental tumor cells or growth inhibition of small tumors in vivo. However, it is the well-established tumors in animal models that mimic clinical patients with existing tumor burdens. Alpha tumor necrosis factor (TNF alpha) is a multifunctional and immunoregulatory cytokine that induces antitumor activity and activates immune cells such as DCs and T cells. We hypothesized that a combined immunotherapy including gene therapy and DC vaccine would have some advantages over each modality administered as a monotherapy. METHODS: We investigated the antitumor immunotherapeutic efficiency of gene therapy by intratumoral injection of AdVTNF-alpha and DC vaccine using subcutaneous injection of TNF-alpha-gene-engineered DC(TNF-alpha) cells, and further developed a combined AdV-mediated TNF-alpha-gene therapy and TNF-alpha-gene-engineered DC(TNF-alpha) vaccine in combating well-established MO4 tumors expressing the ovalbumin (OVA) gene in an animal model. RESULTS: Our data show that vaccination of DC(TNF-alpha) cells pulsed with the OVA I peptide can (i) stimulate type 1 immune response with enhanced antitumor CTL activities, (ii) induce protective immunity against challenge of 5 x 10(5) MO4 tumor cells, and (iii) reduce growth of the small (3-4 mm in diameter), but not large, established MO4 tumors (6-8 mm in diameter). Our data also show that AdVTNF-alpha-mediated gene therapy can completely eradicate small tumors in 6 out of 8 (75%) mice due to the extensive tumor necrosis formation, but not the large tumors (0%). Interestingly, a combined AdVTNF-alpha-mediated gene therapy and TNF-alpha-gene-engineered DC(TNF alpha) vaccine is able to cure 3 out of 8 (38%) mice bearing large MO4 tumors, indicating that the combined immunotherapy strategy is much more efficient in combating well-established tumors than monotherapy of either gene therapy or DC vaccine alone. CONCLUSIONS: This novel combined immunotherapy may become a tool of considerable conceptual interest in the implementation of future clinical objectives. PMID- 15293345 TI - Adenoviral-mediated HGF expression inhibits germ cell apoptosis in rats with cryptorchidism. AB - BACKGROUND: Prior studies have shown that the hepatocyte growth factor (HGF), as known for its multiple biological effects, possibly regulates spermatogenesis or tubulogenesis in the testis. To clarify the effect of HGF on restoration of spermatogenesis, or testicular weight, we transferred the HGF gene into the testis of the rat experimental cryptorchid model. METHODS: Replication-deficient recombinant adenoviral vectors containing the CAG promoter driving rat HGF (pAxCAHGF) and LacZ (pAxCALacZ) were constructed. Sprague-Dawley rats surgically induced with unilateral cryptorchidism and subsequent orchidopexy were divided into three groups: control (PBS), pAxCALacZ and pAxCAHGF by intratesticular injection. At 2 and 4 weeks after subsequent orchidopexy, testes were removed and weighed. These specimens were analyzed histopathologically, and examined for cell apoptosis. HGF expression in these specimens associated with c-Met receptor mediated signal molecules was examined by reverse transcription-polymerase chain reaction (RT-PCR), Western blot or immunohistochemical study. RESULTS: Adenovirus mediated HGF gene transfer induced overexpression of HGF in some seminiferous epithelial cells and interstitial cells, increased the phosphorylation of ERK and Akt, and decreased numbers of apoptotic cells of germ cells. HGF transduction also significantly increased the numbers of germ cells and testicular weight by 4 weeks compared with the other control groups. CONCLUSIONS: Adenoviral-mediated HGF gene transfer into the testis in the cryptorchidism rats inhibited germ cell apoptosis and restored spermatogenesis. PMID- 15293346 TI - Hydrodynamics-based transfection of the liver: entrance into hepatocytes of DNA that causes expression takes place very early after injection. AB - BACKGROUND: The mechanism of gene transfer into hepatocytes by the hydrodynamics based transfection procedure is not clearly understood. It has been shown that, after a hydrodynamic injection, a large proportion of plasmid DNA remains intact in the liver where it is bound to plasma membrane and suggested that this DNA could be responsible for the efficiency of the transfection. METHODS: We have investigated the problem by giving mice a hydrodynamic injection of isotonic NaCl, followed at different time intervals by a conventional injection of DNA, cold or labelled with (35)S, with cDNA of luciferase as a reporter gene. Then, we determined the consequences of that dual injection on luciferase expression and on DNA uptake by the liver and its intracellular fate. By such experiments, it is possible to establish the time dependency of the induction of liver changes caused by a hydrodynamic injection on the one hand and the expression and DNA uptake and fate on the other. Moreover, some experiments have been performed on primary cultures of hepatocytes isolated after a hydrodynamic injection of DNA. RESULTS: When DNA is given to mice by a conventional injection a few seconds after an hydrodynamic injection of isotonic NaCl, luciferase expression in the liver is considerably lower than that observed after a single hydrodynamic injection of the plasmid. On the other hand, as assessed by the rate of DNA degradation and by centrifugation results obtained after injection of (35)S-DNA, the uptake and the intracellular fate of the bulk of DNA are similar whether DNA is administered by a single hydrodynamic injection or by a conventional injection given up to at least 2 h after a hydrodynamic injection of isotonic NaCl. Hepatocytes isolated a few minutes after a hydrodynamic injection exhibit a maximal expression that does not depend on the large amount of DNA that remains bound to the plasma membrane for a relatively long time. CONCLUSIONS: Our results show that the efficiency of hydrodynamics-based transfection depends on a process that takes place very quickly after injection and is not linked to a delay of DNA degradation and the persistence of a large proportion of DNA bound to hepatocytes of the plasma membrane, strongly suggesting that expression after a hydrodynamic injection is caused by a small proportion of DNA molecules that rapidly enter the cytosol probably by plasma membrane pores generated by the hydrodynamic pressure. PMID- 15293347 TI - Enhanced gene transfer and cell death following p53 gene transfer using photochemical internalisation of glucosylated PEI-DNA complexes. AB - BACKGROUND: p53 is frequently mutated in many cancers including human head and neck squamous cell carcinoma and pancreatic cancer. In tumor models, wild-type (wt) p53 gene transfer induces apoptosis and tumor regression in vivo, justifying the extensive clinical investigation of p53 gene therapy. METHODS: p53 nonviral mediated gene transfer was achieved using glucosylated polyethylenimine (PEI) in conjunction with photochemical internalisation (PCI). Experimental conditions were optimised using the green fluorescent protein (GFP) as a reporter. p53 gene transfer was then evaluated using semi-quantitative RT-PCR in p53-deleted PANC3 and p53-mutated FaDu cell lines. Following gene transfer, induction of apoptosis was investigated using phosphatidylserine externalisation and nuclear fragmentation assays. Induction of long-term cell death was analysed using colony forming assays. RESULTS: PCI was found to enhance GFP gene transfer after 48 h in both cell lines. Whether using glucosylated-PEI alone or associated with PCI, p53 gene transfer was achieved with subsequent recovery of p53 mRNA expression in PANC3 cells and a significant 4-fold increase in p53 mRNA expression in FaDu cells. PCI was found to further enhance p53 mRNA expression by 2.3-fold in PANC3 cells. Spontaneous induction of apoptosis following wt-p53 gene transfer was achieved in both cell lines. PCI was found to enhance apoptosis up to levels similar to those achieved with chemotherapy. As a consequence, long-term cell death was significantly enhanced after wt-p53 gene transfer when PCI was used in both cell lines, yielding up to 60% cell death. CONCLUSIONS: PCI increases glucosylated-PEI-mediated p53 gene transfer, apoptosis as well as cell death in mutant p53 human cancer cells. PMID- 15293348 TI - The role of dextran conjugation in transfection mediated by dextran-grafted polyethylenimine. AB - BACKGROUND: Conjugation through primary amines is one of the most commonly used methods to modify polycationic vectors for gene delivery. A better understanding of the effect of the conjugation on the mechanisms of transgene expression can help design efficient polycationic vectors. METHODS: Dextran with a molecular weight of 1500 was grafted onto polyethylenimine (PEI) to produce various degrees of grafting in an effort to investigate how the conjugation affected the mechanisms of transgene expression. Flow cytometry was employed to quantitate the cellular entry of plasmid and the level of transgene expression, which were measured using ethidium monoazide labeled plasmid and green fluorescent protein (GFP), respectively. The buffering capacity of the grafted PEI was determined by titration, and the integrity of the DNA-polymer complexes were examined by exposure to heparin. RESULTS: Grafting of dextran onto PEI was found to significantly diminish the cytotoxicity, buffering capacity, cellular entry, and the integrity of the DNA-polymer complexes. The reductions enlarged as the degree of grafting increased from 0 to 1.84%; however, at an optimal degree of grafting, the dextran-grafted PEI enhanced the percentages of GFP-positive cells to a level 3 times and 1.3 times of those mediated by unmodified PEI for CHO and MDA-MB-231 cells, respectively. CONCLUSIONS: These results demonstrated that the conjugation of dextran onto the primary amines of PEI inhibited the entry of plasmid across the cell membrane, but the change in the structures of the DNA-polymer complexes was able to promote transgene expression when the degrees of conjugation fell below 0.64%. PMID- 15293349 TI - Organ-specific expression of the lacZ gene controlled by the opsin promoter after intravenous gene administration in adult mice. AB - BACKGROUND: The tissue-specific expression of an exogenous gene, under the influence of a tissue-specific promoter, has been examined in the past with pro nuclear injections of the transgene and the development of transgenic mouse models. 'Adult transgenics' is possible with the acute expression of an exogenous gene that is administered to adult animals, providing the transgene can be effectively delivered to distant sites following an intravenous administration. METHODS: The organ specificity of exogenous gene expression in adult mice was examined with a bacterial beta-galactosidase (LacZ) expression plasmid under the influence of the bovine rhodopsin gene promoter. The 8-kb plasmid DNA was delivered to organs following an intravenous administration with the pegylated immunoliposome (PIL) non-viral gene transfer technology. The PIL carrying the gene was targeted to organs with the rat 8D3 monoclonal antibody (MAb) to the mouse transferrin receptor (TfR). RESULTS: The rhodopsin/beta-galactosidase gene was expressed widely in both the eye and the brain of adult mice, but was not expressed in peripheral tissues, including liver, spleen, lung, or heart. Ocular expression included the retinal-pigmented epithelium, the iris, and ciliary body, and brain expression was observed in neuronal structures throughout the cerebrum and cerebellum. CONCLUSIONS: The expression of trans-genes in adult animals is possible with the PIL non-viral gene transfer method. The opsin promoter enables tissue-specific gene expression in the eye, as well as the brain of adult mice, whereas gene expression in peripheral tissues, such as liver or spleen, is not observed. PMID- 15293350 TI - Gene delivery to respiratory epithelial cells by magnetofection. AB - BACKGROUND: For the topical application of DNA vector complexes to the airways, specific extracellular barriers play a major role. In particular, short contact time of complexes with the cell surface caused by the mucociliary clearance hinders cellular uptake of complexes. The aim of this study was to evaluate the ability of magnetofection, a technique based on the principle of magnetic drug targeting, to overcome these barriers in comparison with conventional nonviral gene transfer methods such as lipofection and polyfection. METHODS: Experiments were carried out on permanent (16HBE14o-) and primary airway epithelial cells (porcine and human), and native porcine airway epithelium ex vivo. Transfection efficiency and dose-response relationship of magnetofection were examined by luciferase reporter gene expression. Sedimentation patterns and uptake of gene transfer complexes were characterized by fluorescence and electron microscopy, respectively. RESULTS: We show that (i) application of a magnetic field allows the magnetofectins to sediment and to enrich at the cell surface within a few minutes, (ii) magnetofection bears an improved dose-response relationship, (iii) magnetofection enhances transfection efficiency in both, permanent and primary airway epithelial cells, and (iv) magnetofection leads to significant transgene expression at very short incubation times in an ex vivo airway epithelium organ model. CONCLUSIONS: Magnetofection provides a potential novel method, which may overcome fundamental limitations of nonviral gene transfer to the airways. Due to the accelerated enrichment at the cell surface it may be of major interest for in vivo applications, where long-term incubation times at the target tissue are hardly achievable. PMID- 15293351 TI - Insights into the mechanism of magnetofection using PEI-based magnetofectins for gene transfer. AB - BACKGROUND: Gene delivery by the use of magnetic forces, so-called magnetofection, has been shown to enhance transfection efficiency of viral and non-viral systems up to several-hundred-fold. For this purpose gene carriers, such as polyethylenimine (PEI), are associated with superparamagnetic nanoparticles and complexed with plasmid DNA. Gene delivery is targeted by the application of a magnetic field. METHODS: To investigate the underlying mechanism, we studied the impact of the applied magnetic field on the transfection process of PEI-coated superparamagnetic iron oxide gene vectors (magnetofectins) using various cell lines. In particular, we addressed the question whether accelerated sedimentation of magnetofectins is the driving force or if the magnetic field itself directly influences the endocytic processing of the magnetofectins. The cellular uptake mechanism of magnetofectins was studied by electron microscopy and transfection experiments in the presence of various inhibitors that operate at different steps of endocytosis. RESULTS: In this study we could show that cellular uptake of magnetofectins proceeds obviously by endocytosis. Cellular uptake of magnetofectins behaves almost analogously as compared with PEI polyplexes. Besides unspecific endocytosis, apparently clathrin dependent as well as caveolae-mediated endocytic uptake is involved. CONCLUSIONS: The magnetic field itself does not alter the uptake mechanism of magnetofectins. Obviously, the magnetic forces lead to an accelerated sedimentation of magnetofectins on the cell surface and do not directly affect the endocytic uptake mechanism. So further improvement of magnetic field application could lead to efficient targeting of gene expression into the desired organ and tissue in vivo. PMID- 15293352 TI - An efficient targeted radiotherapy/gene therapy strategy utilising human telomerase promoters and radioastatine and harnessing radiation-mediated bystander effects. AB - BACKGROUND: Targeted radiotherapy achieves malignant cell-specific concentration of radiation dosage by tumour-affinic molecules conjugated to radioactive atoms. Combining gene therapy with targeted radiotherapy is attractive because the associated cross-fire irradiation of the latter induces biological bystander effects upon neighbouring cells overcoming low gene transfer efficiency. METHODS: We sought to maximise the tumour specificity and efficacy of noradrenaline transporter (NAT) gene transfer combined with treatment using the radiopharmaceutical meta-[(131)I]iodobenzylguanidine ([(131)I]MIBG). Cell-kill was achieved by treatment with the beta-decay particle emitter [(131)I]MIBG or the alpha-particle emitter [(211)At]MABG. We utilised our novel transfected mosaic spheroid model (TMS) to determine whether this treatment strategy could result in sterilisation of spheroids containing only a small proportion of NAT expressing cells. RESULTS: The concentrations of [(131)I]MIBG and [(211)At]MABG required to reduce to 0.1% the survival of clonogens derived from the TMS composed of 100% of NAT gene-transfected cells were 1.5 and 0.004 MBq/ml (RSV promoter), 8.5 and 0.0075 MBq/ml (hTR promoter), and 9.0 and 0.008 MBq/ml (hTERT promoter), respectively. The concentrations of radiopharmaceutical required to reduce to 0.1% the survival of clonogens derived from 5% RSV/NAT and 5% hTERT/NAT TMS were 14 and 23 MBq/ml, respectively, for treatment with [(131)I]MIBG and 0.018 and 0.028 MBq/ml, respectively, for treatment with [(211)At]MABG. CONCLUSIONS: These results indicate that the telomerase promoters have the capacity to drive the expression of the NAT. The potency of [(211)At]MABG is approximately three orders of magnitude greater than that of [(131)I]MIBG. Spheroids composed of only 5% of cells expressing NAT under the control of the RSV or hTERT promoter were sterilised by radiopharmaceutical treatment. This observation is indicative of bystander cell-kill. PMID- 15293354 TI - Elimination of linezolid by an in vitro extracorporeal circuit model. AB - Linezolid is an oxazolidinone antibiotic with activity against important grampositive aerobic bacteria, including nosocomial pathogens. It is not known whether dosage adjustments are necessary in patients treated with continuous renal replacement therapies. This in vitro study was conducted to investigate the elimination of linezolid in an in vitro continuous hemo(dia)filtration model using different filter materials (polysulfone, polyacrylonitrile, polyamide), surface areas, and different modes of renal replacement therapies. Linezolid was measured using HPLC with UV-detection. No adsorption of linezolid to any of the tested membranes was detected. Recovery of linezolid in the ultrafiltrate was 98.2 +/- 10.5% in the filtration mode. During dialysis, recovery was significantly less (87.6 +/- 16.1%; p = 0.02). Linezolid elimination was not altered by filter size, when polysulfone filters with surface areas of 0.7 m2 and 1.3 m2 were tested. In conclusion, the dosage recommendations for linezolid are independent of the filter materials. However, the elimination is significantly higher during hemofiltration compared to dialysis. PMID- 15293355 TI - A preclinical cadaver fitting study of implantable biventricular assist device-- AnyHeart. AB - A multifunctional, Korean-made artificial heart (AnyHeart) was developed, and prior to its clinical application, a cadaver-fitting study was performed. The study proposed to determine the optimal cannulation approach, implantation technique and route of the cannula to minimize the organ compression of AnyHeart. The anatomical feasibility and a variety of surgical techniques were evaluated using ten preserved, human cadavers. Implanting AnyHeart with ease is possible using various approaches, including a median sternotomy, and a right or left lateral thoracotomy. The lateral thoracotomy approach is shown to be safe and reproducible, especially in patients who have already undergone an operation that used a median sternotomy. The results of this study will guide improvements in the designs of cannulae and AnyHeart for future clinical applications. PMID- 15293356 TI - Tissue-engineered acellularized valve xenografts: a comparative animal study between plain acellularized xenografts and autologous endothelial cell seeded acellularized xenografts. AB - BACKGROUND: Acellularized valve xenografts are considered a promising way of overcoming the inherent limitations of current prosthetic valves. The aim of this study was to compare the biological responses of an autologous endothelial cell seeded acellularized xenograft (AAX) and a plain acellularized xenograft (PAX) implanted in the pulmonary valve leaflet in the same animal. METHODS: Endothelial cells were isolated and cultured from the jugular vein of goats. Porcine valve leaflets were acellularized with Nacl-SDS, and for AAX, leaflets were then seeded with autologous endothelial cells. A PAX and an AAX were implanted as double pulmonary valve leaflet replacement in the same animal in a goat model (n = 6). After sacrifice, the implanted valve leaflet tissues were retrieved and analyzed visually and under a light microscope. RESULTS AND CONCLUSIONS: Six animals were sacrificed as scheduled during the short-term (6 and 24 hours), mid-term (1 week and 1 month) and long-term (3 and 6 months). Gross and ultrasonographic examinations revealed good valve function with no thrombosis but with slight thickening. Microscopic analysis of the leaflets showed abundant cellular ingrowth into the acellularized leaflets over time. The role of endothelial cell seeding remains controversial. This animal experiment demonstrates the practical feasibility of using acellularized valve xenografts. PMID- 15293357 TI - Contemporary issues in women's health. PMID- 15293358 TI - ACOG committee opinion: patient safety in obstetrics and gynecology. Number 286, October 2003: Committee on Quality Improvement and Patient Safety. AB - Emphasis on patient safety has increased in the past few years mostly in response to the Institute of Medicine report To Err Is Human:Building a Safer Health System. Obstetrician-gynecologists should incorporate elements of patient safety into their practices and also encourage others to use these practices. PMID- 15293359 TI - ACOG committee opinion: ethical considerations in research involving women. Number 290, November 2003: Committee on Ethics. AB - In this Committee Opinion, the American College of Obstetricians and Gynecologists Committee on Ethics affirms both the need for women to serve as research participants and the obligation for researchers, institutional review boards, and others reviewing clinical research to evaluate the potential effect of proposed research on women of childbearing potential, pregnant women, and the developing fetus. PMID- 15293360 TI - Nuclear medicine technologist. PMID- 15293362 TI - Bibliograpy of Toxinology. PMID- 15293361 TI - Computer-enhanced minimally invasive mitral valve surgery. PMID- 15293363 TI - Proceedings of the VII International Potsdam Symposium on Tick-Borne Diseases (IPS VII). March 13-14, 2003. Berlin, Germany. PMID- 15293364 TI - Current awareness on yeast. PMID- 15293366 TI - Bibliography current world literature. Hypertension. PMID- 15293365 TI - Assessment of quality of life and symptom improvement in lung cancer clinical trials. AB - The majority of lung cancers are diagnosed at advanced stages when treatment options are limited and mainly palliative. In advanced lung cancer, quality-of life (QOL) issues have become an integral part of making decisions about various treatment options. Recent clinical trials in patients with lung cancer have assessed symptom improvement and QOL as important endpoints. There are several valid and reliable QOL assessment instruments that specifically evaluate symptoms of lung cancer. These questionnaires evaluate a variety of factors related to emotional, physical, and social well-being. Several key factors, including age, gender, comorbidities, and quality of supportive care may affect symptoms and QOL in patients with lung cancer. Overall, QOL is important for patients with advanced lung cancer; therefore, symptom and QOL assessments are becoming vital in evaluating the efficacy of emerging cancer treatments. PMID- 15293367 TI - Bibliography current world literature. HDL cholesterol. PMID- 15293368 TI - The Nightingale experience. PMID- 15293369 TI - Reactive oxygen species and cardiac preconditioning: many questions remain. PMID- 15293370 TI - Do pharmacological differences among beta-blockers affect their clinical efficacy in heart failure? PMID- 15293371 TI - [The role of risk factors of ischemic stroke in patients with pathology of the extracranial segment of the carotid arteries]. PMID- 15293372 TI - [Role of the lymphatic system in potentiation of the clinical stages of lower extremity chronic venous insufficiency]. PMID- 15293373 TI - [A follow-up study of lung function among ex-asbestos workers with and without pleural plaques]. AB - BACKGROUND: Pulmonary function testing is widely accepted as an integral part of medical surveillance of occupational lung diseases. There are several cross sectional studies evaluating lung function among asbestos-exposed workers, but only few longitudinal surveys have been performed. OBJECTIVES: To evaluate, over a mean follow-up period of 3.7 (SD 1.8) years, the loss of lung function in a group of 103 workers with previous exposure to asbestos (mainly ship building/repairing), according to the presence or absence of pleural plaques at radiological examination. METHODS: Chest radiographic examination was used to ascertain the presence/absence of pleural plaques. If chest X-ray films were positive for pleural plaques, HRCT (High Resolution Computed Tomography) was used to exclude any parenchymal disease. The assessment of lung function over time included repeated measurement of vital capacity (VC), forced expiratory volume in one second (FEV1) and total lung capacity (TLC). Smoking was assessed in terms of pack-years. A Generalized Estimating Equations (GEE) approach to repeated spirometric measurement was used to investigate the relationship between the loss of pulmonary function and (i) presence/absence of pleural plaques, (ii) smoking status, and (iii) work seniority in workplaces with exposure to asbestos. RESULTS: In the ex-asbestos workers, mean age at the first examination was 49 (SD 6) years and work seniority averaged 25 (SD 7) years; 36% were non-smokers, 27% smoked < 15 pack-years, and 37% smoked > or = 15 pack-years. Thirty-six workers showed pleural plaques at radiological examination. Overall, 236 measurements of VC and FEV1, and 234 determinations of TLC were available. Multivariate GEE approach to age- and height-adjusted spirometric data showed that pleural plaques were not associated with a significant loss of pulmonary function during the follow-up. When compared with non-smokers, heavy smokers (> or = 15 pack-years) showed on average a significant loss of VC (-5.3%, IC 95%: -9.4 - -1.2%), FEV1 ( 8.4%, IC 95%: -13.2 - -3.5%), and TLC (-4.0%, IC 95%: -7.4 - -0.5%). An occupational history of previous exposure to asbestos was significantly associated with an 10-year decrease in VC (-3.1%, IC 95%: -5.9 - -0.3%) and FEV1 (-4.9%, IC 95%: -8.3 - -1.5%). CONCLUSIONS: The results of this follow-up study showed that smoking and previous asbestos exposure were associated with a mild, but statistically significant, loss of lung function. Radiological findings of pleural plaques were not related to deterioration of lung function over the follow-up period. PMID- 15293374 TI - [An important variability factor in determination of urinary activity of N-acetyl beta-D-glucosaminidase: contamination of urine with semen]. AB - BACKGROUND: Urinary N-acetyl-beta-D-glucosaminidase (U-NAG) and its isoenzyme B (U-NAG-B) have been demonstrated useful and early markers of renal damage, although they are present in many other tissues and organs. OBJECTIVES: To evaluate semen contamination of the urine and its role in variability of U-NAG. METHODS: To assess control group values beta2-microglobulin, retinol-binding protein and U-NAG were measured in the urine of 30 healthy, non-smoking and non metal-exposed adults (19 females and 11 males). RESULTS: In four urine samples U NAG was higher than the method reference value (5 U/g creatinine), without increases in other functional markers. Microscope examination revealed the presence of sperm in these samples. U-NAG variability decreased after the exclusion of these four values. The role of contamination was confirmed by adding semen to urine: when semen to urine ratio was 1:1000, enzyme activity was more than twice the basal level. CONCLUSIONS: U-NAG variability is strongly increased by contamination with semen, where enzyme concentration (especially NAG-B) is very high. Increased excretion of U-NAG and of its iso-form (U-NAG-B) in males, not correlated with other renal alterations or with exposure to heavy metals or other renal toxic substances, should be carefully evaluated and microscopic observation is advisable to detect the presence of sperm in urine. PMID- 15293375 TI - [Results of an interlaboratory fibre count comparison program concerning the SEM technique]. AB - BACKGROUND: The necessity of quality control for laboratories carrying out airborne asbestos fibre monitoring is recognized in Italian law concerning the worker and the environmental protection against asbestos. The scanning electron microscopy technique is prescribed to control the airborne fibre concentration after asbestos removal from buildings. OBJECTIVES: In 2002 the Italian laboratories carrying out SEM analysis on asbestos were requested by the Istituto Superiore di Sanita to take part in a fibre count comparison program. Thirty-nine agreed to participate. METHODS: Fifteen samples were exchanged among the laboratories participating in the program. Seven samples consisted of polycarbonate membranes and eight of cellulose ester membranes. The sample loads did not exceed 50 fibres/mm2. The fibre counting results were evaluated with reference to the mean value of each sample calculated from the results provided by all the participants. RESULTS: Only 19% of the fibre count results were considered "insufficient", and the results falling within the 95% confidence interval were the 65% of the total. University and public laboratories obtained on the whole the best results. PMID- 15293376 TI - [Assessment of occupational noise exposure: methodology, measurement error, evaluation criteria]. AB - BACKGROUND: Occupational noise exposure can be monitored directly by personal sampling or indirectly, by area sampling. Personal sampling is performed using an integrating sound level meter, worn by the worker while performing his/her job. When area sampling is used, measurements need to be made in all locations where a typical worker stays while performing his/her tasks; the respective partial lengths of exposure need to be accurately monitored, and the time-weighted average sound level of the measured noise levels must be calculated. OBJECTIVES: Current regulations identify three different thresholds, corresponding to different types of action, but they do not propose any standard criteria to decide whether a threshold has been exceeded. Defining standard procedures to assess occupational noise exposure and identifying such thresholds is crucial. METHODS: Using empirical data collected in the field, the effects are illustrated of the number of sampling locations and of the partial lengths of exposure on area sampling measurements, and the effects of duration of noise exposure on both area and personal samplings. RESULTS: When dealing with area samplings, an accurate definition of both sampling locations and partial lengths of exposure is crucial. When arbitrary decisions are taken in selecting sampling locations and/or establishing partial lengths of exposure, spatial changes in noise level and operator' displacements while performing his/her tasks may affect results. Sampling for less than the duration of noise exposure is the major contributor to measurement error, particularly under conditions of unpredictable variation in noise level. In fact, as noise level in the non-monitored time fraction is unknown, measurement error cannot be determined. We estimate that, even under the most favorable circumstances, sampling should last not less than 40% of the duration of a given noise-generating occurrence, for repeated measurements to be dispersed within a range not wider than that generated by the instrumental error. Inter-daily variability is another important aspect in personal noise exposure evaluation. This is a general occurrence, whose effects cannot be controlled by simply considering weekly instead of daily exposures. Results of an investigation, covering about 60 different jobs within a primary aluminum plant, show an inter-daily variability in noise exposure greater than 5 dBA in about 75% of cases. CONCLUSIONS: Personal sampling, when correctly performed and covering the total duration of exposure, provides the most reliable result as it integrates noise over all locations where the worker actually stays while performing his/her tasks, and over the total length of time spent in each task. We propose extending personal sampling to the total duration of actual noise exposure as the standard procedure for monitoring daily personal noise exposure, and valid for the majority of work places. When the range of daily noise exposure includes one regulatory threshold, corresponding to a given type of action, we propose as a standard decision criterion to refer prudentially to the upper 95% confidence limit of the LEP,d arithmetic mean. Such criterion would allow to standardize procedures and decision methods, with the prospect of further improvements in the assessment of exposure to noise. PMID- 15293377 TI - [Job as a risk factor for obesity... and the contrary]. AB - BACKGROUND: Obesity constitutes a risk for several vascular, metabolic and neoplastic diseases. In industrialised countries, but more and more in developing countries too, the prevalence of obesity is increasing. Body Mass Index and circumference of the abdomen are the two simplest and most utilized methods of measuring the degree of obesity in an individual and of comparing selected groups with different ethnic, social, cultural and occupational features. OBJECTIVES: The main aim of this article is to initiate a discussion on the possible contribution that the Occupational Health Physician can make to solving the problem of obesity, which is becoming more and more alarming in social terms. The working conditions favouring an increase in body weight and the negative effects that obesity has on various types of work are reported. METHODS AND RESULTS: A critical review of the literature on obesity and overweight stresses that a low educational level, a low socio-economic status, lack of physical activity in leisure time and certain working conditions, together with the ready availability of food, are the main factors favouring increased prevalence of obesity. Certain jobs also contribute significantly to this problem. Automation and the use of machines/robots for very heavy work in industrialised countries have the "collateral effect" of favouring body weight increase due to low energy consumption. Jobs that are a source of stress, such as work on three rotating shifts, can cause metabolic disorders leading to an increased prevalence of obesity. Contrariwise, obesity renders the individual unfit for some jobs, in fact, an increased incidence of industrial accidents has been related to obesity. CONCLUSIONS: The occupational health physicians engaged in surveillance of workers' health conditions can make a positive contribution to alleviating this problem by focusing their activity on the primary prevention of obesity and advising workers on how to maintain the right weight; otherwise, obese workers should be referred to appropriate medical centres. PMID- 15293378 TI - [Hypermobile syndrome: functional and aesthetic damages. Observations on 50 cases]. AB - BACKGROUND: Type III Ehlers-Danlos syndrome (EDS III) is a collagen defect and manifests as hypermobile joints, which cause dislocations, and fragile skin, which causes scars. There are three clinical-ultrasound (US) stages. OBJECTIVES: Diagnostic techniques are described that permit assessment of cutaneous and phalangeal impairment, and job fitness of patients with EDS III. METHODS: The Beighton scoring system (Bss) and the ultrasound test (US) of the skin and phalanxes (with Sonora Logic 400 MD and 7.5-10 MHz probes) were used in the diagnosis of 50 patients. Dislocations and scars were considered as end-point surrogates and were correlated (in a graph) to clinical-US stages and phalangeal mobility, in order to assess job fitness. RESULTS: Bss and US results agreed with the literature and differentiated the EDS III into 3 clinical stages. Dislocations involved the wrist-hand (52%), shoulder-arm (16%) and foot-ankle (32%) joints. Spearman test for correlation between joint space and scars, dislocations and phalangeal mobility showed high significance (p<0.0001%). CONCLUSION: Dislocations and scars are useful end-point surrogates in the assessment of job fitness. The correlation between Bss and US findings offers a qualitative clinical evaluation (3 clinical stages), while the graph expresses a quantitative evaluation of the biological impairment. The increase in joint mobility produces positive physiological effects (increase in agility) and negative effects (dislocations, scars) that recommend avoiding excessive load on the joints. PMID- 15293379 TI - [Results of risk and impairment assessment in groups of workers exposed to repetitive strain and movement of the upper limbs in various sectors of industry]. AB - BACKGROUND: This presents study the results of a number of investigations regarding risks associated with biomechanical overload of the upper limbs and the consequent health effects (UL-WMSDs) in a large sample of workers in various different jobs. METHODS: Risk assessment regarded 15 different working environments in which 4044 subjects were employed. Most were metalworking factories in which the workers performed assembly tasks (3015 workers). Some made motors for electrical appliances (714 workers), others assembled miniature components (shock absorbers and remote controls: 259 workers), while others handled larger sized parts such as components of large domestic appliances (refrigerators, freezers, ovens: 2037 workers). The sample also included workers in the meat processing industry (chicken and turkey, 969 workers) and hotel room cleaners (60). Exposure assessment was performed using the OCRA checklist for quantifying the risk attributable intrinsically to each individual workstation, as if used for the entire shift. The values thus obtained were entered into a special software program that, for each working area, produced mean weighted values for the results of the checklist and their percentage distribution over four categories: no risk (green), low risk (yellow), moderate risk (red) and high risk (purple). In 11 of the 15 working environments considered, a total of 3511 workers (2221 women and 1290 men) underwent a complete and standardized clinical examination of the upper limbs. Comparisons of the results of exposure evaluation and of the clinical surveys were made between the different types of jobs and between males and females. PMID- 15293380 TI - [On "moderate risk" and health surveillance]. PMID- 15293381 TI - Mental well-being in settings of 'complex emergency': an overview. AB - The mental state of people affected by war and other disasters has been a subject of special interest to academic researchers and practitioners in humanitarian assistance and public health for over two decades. The last decade in particular has seen a rise in the number of papers published in scholarly journals around the Post-Traumatic Stress Disorder (PTSD) debate. Anthropologists have rarely engaged in this debate. Nevertheless, some of the most illuminating contributions have come from socio-medical anthropology (Last, 2000). This volume brings together a wide range of disciplines in the human sciences to address some of the key questions that bear upon the mental health and well-being of populations affected by war and displacement, with contributions from applied biosocial and medical anthropology (Almedom; Lewando-Hundt et al.); applied psychology/public health and social psychiatry (Carballo et al.; Snider et al.; Fullilove et al.); social work (Ahearn & Noble); and political sciences (Pupavac). The four themes that run through this set of papers (outlined below) remain topical areas of contention in contemporary humanitarianism. Scholars and practitioners in the biosocial sciences may wish to engage in the empirical study of human (if not humanitarian) responses to disaster focusing on questions as yet unanswered. PMID- 15293382 TI - Psychosocial assessment for victims of violence in Peru: the importance of local participation. AB - This paper describes a pilot study assessing the psychosocial impact of political violence in the Peruvian Andes, utilizing a collaborative approach with local professionals and communities. The study team prioritized dialogue and information exchange with the local professional community and villagers participating in the assessment in order to raise awareness of psychosocial issues and provide education and support. Participation in the pilot study had positive therapeutic effects for villagers, and inspired ongoing discussion groups to address psychosocial problems in communities. This paper also describes a psychosocial assessment strategy utilizing qualitative methods and an adaptation of the Harvard Trauma Questionnaire in collaboration with Andean villagers. Usefulness and limitations of the data will be reviewed, in terms of cultural and context relevance, usefulness for informing interventions, and comparisons with ethnographic methodologies and other survey instruments. PMID- 15293383 TI - Post-civil war adaptation and need in Managua, Nicaragua. AB - Within seven years after the end of the Nicaraguan civil war in 1990, forced migrants, whose lives had been most disrupted by the conflict, were self-settled in a squatter community in the capital city of Managua and lived in extreme poverty with minimal health, education, security and social service supports. Compared with voluntary migrant neighbours, whose lives had been less affected by the conflict, forced migrants exhibited equal clinically significant symptoms of physical and mental health and psychosocial maladaptation. These findings run counter to generally held theory and assumptions about the negative long-lasting effects of the trauma and stress of war, forced migration and resettlement. Explanations are offered to explain the discrepancies between theory and the study findings as well as the dominance of poverty and socioeconomic status. Implications are also drawn for increasing social support and other durable forms of assistance that emerge from the study as important to meeting the needs of equally poor and unhealthy forced and voluntary migrants in proliferating squatter communities throughout the Third World. PMID- 15293384 TI - Advocating multi-disciplinarity in studying complex emergencies: the limitations of a psychological approach to understanding how young people cope with prolonged conflict in Gaza. AB - The paper looks at the limitations and strengths of using the A-cope questionnaire for measuring strategies for coping with prolonged conflict by Palestinian young people in Gaza. The scale was administered to young people between the ages of 8 and 17. The results show some gender differences in coping strategies. However, some items on the subscales are not relevant for Muslim societies or societies in situations of prolonged conflict. The authors suggest that combining an anthropological contextual perspective and qualitative data with psychological instruments is an effective way of addressing the limitations of using a single quantitative method of assessment in non-Western complex social and cultural settings. PMID- 15293386 TI - Factors that mitigate war-induced anxiety and mental distress. AB - The effects of war-induced anxiety and mental distress on individuals and groups can either be mitigated or exacerbated by 'humanitarian action'. This paper focuses on two key factors that protect the mental well-being of war-affected populations: organized displacement or assisted relocation; and coordinated humanitarian aid operations that are responsive to local needs. Qualitative data from two internally displaced person (IDP) camps in Eritrea are presented. Analysis of these data serves to substantiate and refine a working hypothesis: that social support of the right type, provided at the right time and level, can mitigate the worst effects of war and displacement on victims/survivors. An integrated model of psychosocial transition is suggested. The implications of this approach for humanitarian policy and practice are discussed in the wider context of current debates and lamentations of the 'humanitarian idea'. PMID- 15293385 TI - The social diagnostics of stroke-like symptoms: healers, doctors and prophets in Agincourt, Limpopo Province, South Africa. AB - This paper focuses on the clinical and social diagnostics of stroke-like symptoms in Limpopo Province, South Africa. The research questions addressed here are: what are the lay understandings of stroke-like symptoms and what are the health seeking behaviours of Tsongan Mozambican refugees and South Africans in this area? The study site is ten villages in the Agincourt sub-district of Limpopo Province which are within the health surveillance area of the Agincourt Health and Population Unit (AHPU) of the University of Witwatersrand. The population are Tsongan who speak Shangaan and comprise self-settled Mozambican refugees who fled to this area during the 1980s across the nearby border and displaced South African citizens. The latter were forcibly displaced from their villages to make way for game reserves or agricultural development and moved to this area when it was the former 'homeland' of Gazankulu. The team collected data using rapid ethnographic assessment and household interviews as part of the Southern Africa Stroke Prevention Initiative (SASPI). The main findings are that stroke-like symptoms are considered to be both a physical and social condition, and in consequence plural healing using clinical and social diagnostics is sought to address both these dimensions. People with stroke-like symptoms maintain their physical, mental and social well-being and deal with this affliction and misfortune by visiting doctors, healers, prophets and churches. PMID- 15293387 TI - Mental health and coping in a war situation: the case of Bosnia and Herzegovina. AB - The latter part of the twentieth century has seen an increased concern for the implications of war for civilian populations, and more attention has been given to psychosocial impacts of uprooting and displacement. 'Loss of place', acute and chronic trauma, family disruption and problems of family reunification have become issues of concern. The war in Bosnia was characterized by massive displacement, disruption and loss of life, relatives and property. Health and psychosocial well-being were affected in a number of ways. There was an overwhelming loss of perceived power and self-esteem. Over 25%, of displaced people, for example, said they no longer felt they were able to play a useful role; even in non-displaced populations approximately 11% of those interviewed said that they had lost a sense of worth. Widespread depression and feelings of fatigue and listlessness were common and may have prevented people from taking steps to improve their situation. Almost a quarter of internally displaced people had a high startle capacity and said they were constantly nervous. Most adverse psychosocial responses increased with age and in a population that includes many elderly people this poses serious problems. The findings point to major challenges with respect to repatriation and reconstruction. They highlight the importance of family reunification and the facilitating of decision-making by affected people themselves. The findings also shed light on potential problems associated with over-dependence on external assistance and hence the need for people to be given the means of using their skills and knowledge to control their day-to-day lives. PMID- 15293388 TI - Promoting collective recovery through organizational mobilization: the post-9/11 disaster relief work of NYC RECOVERS. AB - NYC RECOVERS, an alliance of organizations concerned with New York City's social and emotional recovery post-9/11, was formed to meet the need to rebuild social bonds strained or ruptured by the trauma to the regional system caused by the destruction of the Twin Towers. NYC RECOVERS, with minimal funding, was able to create a network of 1000 organizations spanning the five boroughs, carrying out recovery events throughout the 'Year of Recovery', September 2001 to December 2002. This paper describes the concepts, techniques and accomplishments of NYC RECOVERS, and discusses potentials of the model, as well as obstacles to its implementation. PMID- 15293389 TI - Psychosocial interventions and the demoralization of humanitarianism. AB - This paper critically analyses from a political sociology standpoint the international conceptualization of war-affected populations as traumatized and in need of therapeutic interventions. It argues for the importance of looking beyond the epidemiological literature to understand trauma responses globally. The paper explores how the imperative for international psychosocial programmes lies in developments within donor countries and debates in their humanitarian sectors over the efficacy of traditional aid responses. The aim of the paper is threefold. First, it discusses the emotional norms of donor states, highlighting the psychologizing of social issues and the cultural expectations of individual vulnerability. Second it examines the demoralization of humanitarianism in the 1990s and how this facilitated the rise of international psychosocial work and the psychologizing of war. Third, it draws attention to the limitations of a mental health model in Croatia, a country which has been receptive to international psychosocial programmes. Finally it concludes that the prevalent trauma approaches may inhibit recovery and argues for the need to re-moralize resilience. PMID- 15293390 TI - Colorimetric recognition of different enzymology-concerning transition metals based on a hybrid cluster complex. AB - A hybrid cluster complex, formed by chelating a chromogenic ligand to a [2Fe-2S] cluster, sensitively exhibited differential colorimetric responses towards Hg2+, Cd2+, Cr3+, Pb2, Sn2+, Cu2+, Zn2+, Fe3+ and Co2+ in water at physiological pH. Speciation of some of these metal elements, such as Cr(III) and Sn(IV), was also studied by UV/Vis absorption. PMID- 15293391 TI - Fluorescent-based single-strand conformation polymorphism/heteroduplex capillary electrophoretic mutation analysis of the P53 gene. AB - Fluorescent-based single-strand conformation polymorphism (F-SSCP) analysis with capillary electrophoresis (CE) is the most common method for the detection of mutation because of its high sensitivity and resolution. In this study, we prepared an inexpensive linear polyacrylamide (LPA), and successfully applied it to CE-SSCP analysis and tandem CE-SSCP/heteroduplex analysis (HA) of the P53 gene on an ABI capillary genetic analyzer. A comparison of the sieving capabilities of a homemade LPA and commercial polydimethylacrylamide (PDMA) demonstrates that the homemade LPA has a higher resolution, a shorter analysis time, and is more suitable for tandem SSCP/HA than commercial PDMA. To show the usefulness, mutations of P53 gene exon 7 - 8 in 37 tumor samples were investigated by using homemade LPA. The results indicate that 10 mutations were found in 9 of 37 cases; the majority of P53 mutations were missense mutations, and 70% were located in exon 7, which plays an important role in neoplastic progression in human tumorigenesis. PMID- 15293392 TI - ppt level detection of samarium(III) with a coated graphite sensor based on an antibiotic. AB - N-[2-[4-[[[(Cyclohexylamino)carbonyl]amino]sulfonyl]phenyl]ethyl]-5-methyl pyrazine carboxamide (glipizid) was explored as an electro-active material for preparing a polymeric membrane-based sensor selective to samarium ions. The membrane incorporated 30% poly(vinyl chloride) (PVC), 53% benzyl acetate (BA), 11% glipizid and 6% sodium tetraphenyl borate. When coated on the surface of a graphite electrode, it exhibits Nernstian responses in the concentration range of 1.0 x 10(-5) to 1.0 x 10(-10) M, with a detection limit of 8.0 x 10(-11)M samarium. The electrode shows high selectivity towards samarium over several cations (alkali, alkaline earth, transition and heavy metal ions), and specially lanthanide ions. The proposed sensor has a very short response time (< 15 s), and can be used in a wide pH range for at least ten weeks. It was used as an indicator electrode in potentiometric titration of Sm(III) ions with an EDTA solution, and for determination of samarium in binary and ternary mixtures. PMID- 15293393 TI - Direct fluorescence quantification of chromium(VI) in wastewater with organic nanoparticles sensor. AB - Under ultrasonic irradiation, organic fluorescence nanoparticles have been prepared by a reprecipitation method. Compared with single organic fluorophores, these nanoparticles are brighter, more stable against photobleaching and more water-soluble. They also have high room-temperature fluorescence quantum yields (approximately 20%) and a long fluorescence lifetime (approximately 0.2 micros). Based on the fluorescence quenching of nanoparticles by chromium(VI), a method for the selective determination of chromium(VI) without the separation of chromium(III) in water was developed. Under the optimal conditions, the linear range of the calibration curve was 7.0 x 10(-6) - 1.0 x 10(-4) mol L(-1). The detection limit was 2.8 x 10(-6) mol L(-1). The method is characterized by a short reaction time, stable fluorescence signals, simplicity and high selectivity. The present assay has been applied to the selective quantification of Cr(VI) in wastewater with satisfactory results. PMID- 15293394 TI - A new method for atmospheric nitrogen dioxide measurements using the combination of a stripping coil and fluorescence detection. AB - A new stripping coil for the collection of nitrogen dioxide (NO2) has been developed to increase its versatility and efficiency. Nitrogen dioxide measurements based on quantitative collection through a reaction coil into an alkaline solution has been examined. Nitrogen dioxide is collected in a 0.1 N NaOH solution. This collection system has an efficiency of nearly 100%. The absorbed nitrogen dioxide has been measured by fluorescence detection with sub ppbv detection limits. The excitation wavelength at 360 nm and the produced emission wavelength at 405 nm were suitable for nitrite ion measurements. PMID- 15293395 TI - Determination of trace levels of iron in a seawater sample using isotope dilution/inductively coupled plasma mass spectrometry. AB - An analytical method for trace levels of iron in a seawater sample using isotope dilution ICP-MS was developed. Preconcentration of iron and the removal of major elements in seawater such as alkali and alkaline-earth elements can be carried out quickly using a chelating resin disk by adjusting the sample pH to 3. The collision cell option of the ICP-MS instrument method was used to improve the performance of the instrument for iron measurements since ArO and ArN interferences could be reduced using this analytical method. About 4 ml min(-1) helium, as the collision gas, were introduced into the cell. 40Ar14N and 40Ar16O which interfere with 54Fe and 56Fe in water had their amounts decreased by 5 orders of magnitude. Then, the isotope dilution method was used for iron determination below ng g(-1) level of trace iron in four environmental reference materials (river water standard sample JAC-0031 (Japan Soc. for Analytical Chemistry), estuarine standard sample SLEW-2 (NRC Canada) and seawater standard samples CASS-3 and NASS-5 (NRC Canada)) were measured. Good agreement between analytical results and certified values of reference materials was obtained, which confirmed the effectiveness of this method. PMID- 15293397 TI - An enrichment method of molybdenum on a cellulose nitrate resin for the determination by electrothermal atomic absorption spectrometry coupled with the resin suspension injection. AB - Cellulose derivative resin (CDR) suspensions containing resin particles of cellulose nitrate (CDR(CN)), cellulose acetate (CDR(CA)), or cellulose triacetate (CDR(CTA)) were prepared as the sorbent for resin suspension injection (RSI) electrothermal atomic absorption spectrometry (ETAAS), in which fine resin particles holding a desired trace element were directly injected into the graphite tube as the suspension. To compare the sorption ability, the distribution ratios of the Mo(VI)-phenylfluorone complex were measured between the aqueous phase and the sorbents, including the CDRs mentioned above and the finely divided anion exchange resin (AR). The results showed that the sorption ability decreases in the following order: CDR(CN) > or = AR > CDR(CA) > or = CDR(CTA). It was concluded that CDR(CN) was able to extract more than 98% of Mo(VI), and was suitable for RSI-ETAAS as well as AR. CDR(CN) was used for the determination of Mo in NIES certified reference materials, No. 10 Rice flour unpolished; the results showed fairly good agreements between the analytical values and the certified values. PMID- 15293396 TI - Determination of cadmium in river water samples by flame AAS after on-line preconcentration in mini-column packed with 2-aminothiazole-modified silica gel. AB - A rapid and sensitive method was developed to determine trace levels of Cd2+ ions in an aqueous medium by flame atomic absorption spectrometry, using on-line preconcentration in a mini-column packed with 100 mg of 2-aminothiazol modified silica gel (SiAT). The Cd2+ ions were sorbed at pH 5.0. The preconcentrated Cd2+ ions were directly eluted from the column to the spectrometer's nebulizer-burner system using 100 microL of 2 mol L(-1) hydrochloric acid. A retention efficiency of over 95% was achieved. The enrichment factor (calculated as the ratio of slopes of the calibration graphs) obtained with preconcentrations in a mini column packed with SiAT (A = -1.3 x 10(-3) + 1.8 x 10(-3)[Cd2+]) and without preconcentrations (A = 4 x 10(-5) + 3.5 x 10(-5)[Cd2+]), was 51 and the detection limit calculated was 0.38 microg L(-1). The preconcentration procedure was applied to determine trace levels of Cd in river water samples. The optimum preconcentration conditions are discussed herein. PMID- 15293398 TI - A novel voltammetric method for the direct determination of copper in complex environmental samples. AB - A novel voltammetry with a modified gold electrode for the direct determination of copper in environmental samples, without any pretreatment, is proposed in this paper. A porous disorganized monolayer was formed on the surface of the gold electrode by the self-assembly of mercaptoacetic acid (MAA), which could selectively permeate small molecules. Subtractive square wave anodic stripping voltammetry (SASV) was applied to determine copper, in which the underpotential deposition (UPD) of copper was used as the deposition step. The linear range was from 8 x 10(-7) to 1 x l0(-5) mol l(-1) by the modified electrode in the presence of human serum albumin, and the determination was not interfered with common metal ions. Copper in a real environmental sample was successfully detected. PMID- 15293399 TI - New plastic membrane and carbon paste ion selective electrodes for the determination of triprolidine. AB - Triprolidine (Trip) ion selective electrodes of three types: the conventional polymer membrane (I), graphite coated electrode (II) and carbon paste electrode (III), have been prepared, based on the ion pair of triprolidine hydrochloride with sodium tetraphenylborate. The electrodes exhibit a linear response with a mean calibration graph slope of 56.12, 55.00 and 54.32 mV decade(-1) at 25 degrees C for I, II and III, respectively, within the concentration ranges 1.96 x 10(-5) - 1.00 x 10(-2) M for I and 3.84 x 10(-5) - 1.00 x 10(-2) M for II and III. The detection limits are 1.13+/-0.13 x 10(-5), 1.70+/-0.06 x 10(-5) and 1.78+/-0.05 x 10(-5) M for the three electrodes, respectively. The change of pH within the ranges 4.85 - 8.75 and 4.70 - 8.50 for I and III, respectively, did not affect the electrode performance. The standard electrode potentials were determined at different temperatures and were used to calculate the isothermal coefficient of the electrode. The electrodes showed a very good selectivity for Trip with respect to a large number of inorganic cations and compounds. The standard addition method was applied to the determination of TripCl in pure solution, pharmaceutical preparations, and urine samples. PMID- 15293400 TI - Enhanced reduction and determination of trace thyroxine at carbon paste electrode in the presence of trace cetyltrimethylammonium bromide. AB - The electrochemical response of thyroxine (T4) at a carbon paste electrode (CPE) in the presence of cetyltrimethylammonium bromide (CTAB) was investigated. It gave a well-defined oxidation peak at 0.80 V and a sensitive reduction peak at 0.40 V. Compared with the indiscernible signal in the absence of CTAB, the reduction current of T4 at CPE was greatly enhanced in the presence of CTAB, due to the interactions between T4, CTAB and the hydrophobic electrode surface. The electrode process of T4 was explored by cyclic voltammetry and chronocoulometry. The effect of surfactants on the reduction reaction proved that bromide ions (Br( )) in CTAB might form special ion complexes with T4 via a special interaction with the iodine atoms on T4, which would activate the reduction of T4. The sensitive and selective reduction of T4 in this system was applied to the determination of T4 in drugs; a detection limit of 6.5 x 10(-9) M was obtained (sigma= 3). PMID- 15293401 TI - A chitosan-multiwall carbon nanotube modified electrode for simultaneous detection of dopamine and ascorbic acid. AB - A chemically modified electrode based on a chitosan-multiwall carbon nanotube (MWNT) coated glassy carbon electrode (GCE) is described, which exhibits an attractive ability to determine dopamine (DA) and ascorbic acid (AA) simultaneously. The modified electrode exhibited a high differential pulse voltammetry (DPV) current response to DA at 0.144 V and AA at -0.029 V (vs. SCE) in a 0.1 mol l(-1) phosphate buffer solution (pH = 7.2). The properties and behaviors of the chitosan-multiwall carbon nanotube modified electrode (MC/GCE) were characterized using cyclic voltammetry (CV) and DPV methods. The mechanism for the discrimination of dopamine from ascorbic acid at MC/GCE is discussed. The linear calibration range for DA and AA were 5 x 10(-7) mol l(-1) to 1 x 10(-4) mol l(-1) (r = 0.997), and 5 x 10(-6) mol l(-1) to 1 x 10(-3) mol l(-1) (r = 0.996), respectively. The MC/GCE showed good sensitivity, selectivity and stability. PMID- 15293402 TI - Preparation of a new type of fiber adsorbent attached with silica microparticles. AB - A new type of fiber adsorbent attached with silica microparticles was prepared. The silica microparticles were formed by the polymerization of silica oligomers on glass fibers, which were woven into a glass filter. The surface of the silica microparticles was chemically modified by bonding C18-ligands. SEM images indicated that the diameter of the uniform and spherical silica microparticles fixed on glass fibers was on the order of micrometers. It was confirmed that the glass filter adsorbent was effective for the adsorptive removal of toluene of low concentrations. PMID- 15293403 TI - Sensitive determination of domoic acid using high-performance liquid chromatography with electrogenerated tris(2,2'-bipyridine)ruthenium(III) chemiluminescence detection. AB - A new sensitive determination method of domoic acid based on the chemiluminescent reaction of tris(2,2'-bipyridine)ruthenium(III) has been developed. The method exhibited good reproducibility. The relative standard deviation of six replicate measurements was 1.6% for 10 ng ml(-1). A calibration graph, based on a standard domoic acid solution, was linear over the range of 1 - 500 ng ml(-1) (coefficient of correlation, r2 = 0.9995) and the detection limit was 8 pg (signal-to-noise ratio = 3) without any preconcentration and derivatization steps. This new method was successfully applied to a real sample of blue mussels spiked with 2 microg g( 1) domoic acid. PMID- 15293404 TI - Direct detection of a phase change in PdO/CeO2 supported on chi-Al2O3 by means of in situ high-temperature measurements of XRD and FTIR. AB - Phase changes between PdO and Pd metal can be directly detected in PdO/CeO2 catalysts supported on chi-Al2O3 by means of in situ high-temperature measurements of X-ray diffraction and FT-IR in relation to the catalytic activity for the methane oxidation of microcrystalline PdO. Reversible changes in the solid phases are observed from PdO to Pd and Pd to PdO under O2-deficient and O2 excess atmospheres, respectively. Nanosizes of PdO and Pd crystallites, the distorted PdO crystal structure along the (110) plane, and also a distorted Pd metal crystal structure along the (200) plane as well as the large surface area elucidate the high catalytic activity for the methane oxidation of PdO/CeO2 catalysts prepared with an atomic ratio of Pd:Ce = 1:1. PMID- 15293405 TI - Terbium-sensitized fluorescence method for the determination of pazufloxacin mesilate and its application. AB - A simple, rapid and highly sensitive fluorometric method for the determination of pazufloxacin mesilate (PZFX) is described. It is based on the formation of the complex [Tb(PZFX)2](3+), which shows the intensive characteristic bands of Tb3+. Optimum conditions for the determination were investigated. Under the optimum experimental condition, the fluorescence intensity responds linearly to the PZFX concentration in the range of 2.0 x 10(-8) - 5.0 x 10(-6) mol/l with a detection limit of 6.2 x 10(-9) mol/l. The method has been successfully applied to the determination of PZFX in urine and serum samples. PMID- 15293406 TI - Simultaneous XRD-DSC measurements of water-2-propanol at sub-zero temperatures. AB - A simultaneous low-temperature X-ray powder-diffractometric (XRD)-DSC technique was applied to the solid state and melting process of frozen aqueous solutions of 2-propanol. 1H NMR spectra were also obtained at low temperatures. The chemical shifts of the CH3 proton and the CH proton can be classified into four temperature regions: higher than -20 degrees C, around -20 degrees C, -50 to -20 degrees C, and lower than -50 degrees C. In the XRD data, five small diffraction peaks for 2 theta at 21.0 degrees, 25.2 degrees, 27.8 degrees, 31.1 degrees and 32.1 degrees can be attributed to the peritectic, while five diffraction peaks at 22.5 degrees, 24 degrees, 25.6 degrees, 33.4 degrees and 39.8 degrees can be attributed to ice; these peaks are due to the hexagonal form of ice, which disappears upon melting. However, the diffraction peak at 33.4 degrees showed a different pattern than the other peaks due to hexagonal ice. These results indicate that the temperature dependence of the diffraction peak at 33.4 degrees for 2 theta is related to the formation of hydrogen bonds between 2-propanol and water. The simultaneous XRD-DSC technique was effective for investigating this water-alcohol mixture at low temperatures. PMID- 15293407 TI - DNA network structures on various solid substrates investigated by atomic force microscopy. AB - We have fabricated DNA network structures on glass and sapphire substrates. As a comparison, we also formed the network structure on mica substrate. For titanate strontium substrate, however, DNA network can not be obtained even if it is wet treated by Na2HPO4 solution to make it hydrophilic. We also discuss the factors that affect the DNA networks formed on various substrates. PMID- 15293408 TI - Aging of concrete buildings and determining the pH value on the surface of concrete by using a handy semi-conductive pH meter. AB - A new method was devised for measuring the pH of a concrete surface by pHBOY-P2 with a piece of filter paper by extracting the pH value from concrete. This is a simple and inexpensive method that does not damage the concrete building, and is easy to apply on concrete samples for monitoring. By using the method mentioned above, a drastic decrease of the pH value of concrete bridges and buildings has investigated. The method is environmentally friendly to detect the pH value change of concrete as an environmental sample investigation. PMID- 15293409 TI - Synergistic cloud point extraction behavior of aluminum(III) with 2-methyl-8 quinolinol and 3,5-dichlorophenol. AB - The cloud point extraction behavior of aluminum(III) with 8-quinolinol (HQ) or 2 methyl-8-quinolinol (HMQ) and Triton X-100 was investigated in the absence and presence of 3,5-dichlorophenol (Hdcp). Aluminum(III) was almost extracted with HQ and 4(v/v)% Triton X-100 above pH 5.0, but was not extracted with HMQ-Triton X 100. However, in the presence of Hdcp, it was almost quantitatively extracted with HMQ-Triton X-100. The synergistic effect of Hdcp on the extraction of aluminum(III) with HMQ and Triton X-100 may be caused by the formation of a mixed ligand complex, Al(dcp)(MQ)2. PMID- 15293410 TI - H-point standard addition method for the selective simultaneous determination of nickel and copper using 1-(2-pyridylazo)-2-naphthol in Tween 80 micellar media. AB - The H-point standard addition method (HPSAM) has been applied for the simultaneous determination of nickel and copper in trace levels, using 1-(2 pyridylazo)-2-naphthol (PAN) as a chromogenic reagent in aqueous Tween 80 micellar media. Under the optimum condition, the simultaneous determinations of nickel and copper by HPSAM were performed. The absorbances at one pair of wavelengths, 548 and 579 nm, were monitored with the addition of standard solutions of copper. The method is able to accurately determine copper-to-nickel ratios of 15:1 to 1:10 (Wt/Wt). The effects of diverse ions on the determination of nickel and copper to investigate the selectivity of the method were also studied. The recommended procedure was successfully applied to some water and alloy samples. PMID- 15293411 TI - Ion-exchange selectivity of anion exchange resin modified with polystyrenesulfonic acid. AB - In order to change the ion-exchange selectivity of anion-exchange resin, the surface of a gel-type anion exchange resin was modified with anionic polyelectrolyte, polystyrenesulfonic acid. Using this modified resin, the ion exchange rate of nitrate was little decreased, but that of sulfate was evidently decreased. It is considered that the ion-exchange reaction of the multivalent anion is suppressed by the greater electrostatic repulsive force against the modification layer than that against the monovalent anion. Thus, this modified resin may be suitable for the selective separation of monovalent anions. The influence of the modified condition on the ion-exchange rate was examined. Furthermore, this modified resin was used to separate nitrate ions from sulfate ions in the aqueous solution. PMID- 15293412 TI - Structural analysis of selected characteristic flavones by electrospray tandem mass spectrometry. AB - Seven structure analogical flavonoid aglycones have been analyzed using electrospray ionization tandem mass spectrometry (ESI-MSn) in the negative-ion mode. The spectra obtained ESI-MSn allowed us to propose plausible schemes for their fragmentation mechanism. By analyzing the product ions spectra of deprotonated molecule ions [M-H](-), some neutral diagnostic losses and specific retro Diels-Alder fragments were obtained. By using all of these characteristic fragment ions we can specially differentiate the flavone isomer. PMID- 15293413 TI - Patients accessing Web-based medical records. AB - A handful of provider organizations are allowing patients to access parts of their electronic medical records. Most of them see it as an expansion of their patient-physician messaging service: It can improve on simple messaging by providing historical data, making it easier for patients to find health content related to their conditions, and allowing them to correct errors in their charts. PMID- 15293414 TI - Evaluating patient access to electronic health records. PMID- 15293415 TI - Opening your site to people with disabilities. PMID- 15293416 TI - Identification of collagen encapsulation at the dentin/adhesive interface. AB - PURPOSE: The purpose of this study was to investigate the dentin/adhesive interfacial characteristics of three current commercial adhesives with different relative hydrophilic/hydrophobic composition, using a nondestructive staining technique. MATERIALS AND METHODS: Dentin surfaces of 18 unerupted human third molars were randomly selected for treatment with one of three commercial dentin bonding agents according to manufacturers' instructions for the "wet" bonding technique. The adhesives were ranked based on hydrophilic/hydrophobic component ratios (ie, ability to dissolve in water), highest to lowest, as follows: Uno (Pulpdent) > Prime&Bond NT (PBNT, Dentsply Caulk) > Single Bond (SB, 3M ESPE). Dentin/adhesive (d/a) interface sections were stained with Goldner's trichrome, a classical bone stain, and examined using light microscopy. RESULTS: The extent and degree to which the adhesive encapsulates the demineralized dentin matrix is reflected in the color differences in the stained sections. The depth of demineralization appeared comparable among these bonding systems, but adhesive infiltration varied from highest to lowest as follows: Uno > PBNT > SB. CONCLUSIONS: The differential staining technique provided a clear representation of the depth of dentin demineralization and extent/degree of adhesive encapsulation of the exposed collagen at the d/a interface. This technique provides a mechanism for readily identifying vulnerable sites at the d/a interface. The composition of the one-bottle adhesive systems has a substantial effect on the interfacial structure of the d/a bond. PMID- 15293417 TI - Effect of bur-cut dentin on bond strength using two all-in-one and one two-step adhesive systems. AB - PURPOSE: To compare the microtensile bond strength (MTBS) of two all-in-one adhesive systems and one experimental two-step self-etching adhesive system to two types of bur-cut dentin. MATERIALS AND METHODS: Using one of the three adhesives, Xeno CF Bond (Xeno), Prompt L-Pop (PL), or the experimental two-step system ABF (ABF), resin composite was bonded to flat buccal and root dentin surfaces of eight extracted human premolars. These surfaces were produced using either regular-grit or superfine-grit diamond burs. After storage overnight in 37 degrees C water, the bonded specimens were sectioned into six or seven slices approximately 0.7 mm thick perpendicular to the bonded surface. They were then subjected to microtensile testing. The surfaces of the fractured specimens were observed microscopically to determine the failure mode. In addition, to observe the effect of conditioning, the two types of bur-cut dentin surfaces were conditioned with the adhesives, rinsed with acetone, and observed with SEM. RESULTS: When Xeno and PL were bonded to dentin cut with a regular-grit diamond bur, MTBS values were lower than to superfine bur-cut dentin, and failures occurred adhesively at the interface, whereas the experimental two-step adhesive showed no significant difference in microtensile bond strength between two differently cut surfaces. CONCLUSION: The all-in-one adhesives tested here improved bond strengths when bonded to superfine bur-cut dentin as a substrate, whereas the experimental two-step adhesive system showed unchanged bonding to both regular and superfine bur-cut dentin surfaces. PMID- 15293418 TI - Evaluation of etching time on dentin bond strength using single bottle bonding systems. AB - PURPOSE: Incomplete infiltration of the demineralized collagen network may result in a weak zone within the hybrid layer and between the hybrid layer and dentin. The current study evaluates whether reducing the etching time to 5 s from the recommended 15 s or increasing it to 30 s has an effect on dentin bonding. MATERIALS AND METHODS: 108 extracted molars were assigned to 3 bonding agent groups (n = 36): (a) Single Bond (SB), (b) One-Step (OS), and (c) Syntac Single Component (SSC). Each group was further divided into three subgroups (n = 12) of different etching times: 5, 15, and 30 s. All groups were bonded with Z100 composite resin according to the manufacturer's instructions. All specimens were thermocycled 300 times between +/-5 degrees C and +/-55 degrees C, and shear bond strength testing and mode of failure analysis were performed. RESULTS: The bond strength of SB (5 s: 15.5 MPa +/- 4.4; 15 s: 16.5 MPa +/- 3.1; 30 s: 16.8 MPa +/- 3.2) and OS (5 s: 13.7 MPa +/- 1.8; 15 s: 12.4 MPa +/- 3.8; 30 s: 10.6 MPa +/- 3.8) showed no significant differences (p < 0.05) for the different etching times. For SSC, different etching times showed significant differences (5 s: 10.9 MPa +/- 1.8; 15 s: 7.5 MPa +/- 2.5; 30 s: 6.4 MPa +/- 2.1). The mode of failure for SB and OS was adhesive or mixed adhesive/cohesive. For SSC, all failures were adhesive. CONCLUSION: Etching times of less than 15 s do not seem to adversely affect bonding to dentin. PMID- 15293419 TI - Penetration of a bonding agent into De- and remineralized enamel in vitro. AB - PURPOSE: To evaluate bonding penetration into different enamel substrates. MATERIALS AND METHODS: Ten extracted human molars were mesiodistally sectioned. Buccal and lingual enamel surfaces were divided into four equal areas using sticky wax. The central two areas of each tooth (n = 20) were demineralized for 12 weeks using an acidic gel (pH 4.8). The lateral areas served as controls. After demineralization, ten specimens were remineralized in a saliva substitute for three weeks. An amine fluoride solution (Elmex Fluid) was applied on one half of each specimen before acid etching. After etching for 120 s, an enamel-bonding agent (Heliobond) containing 0.1% rhodamine was applied onto test and control areas, and was light cured for 60 s. Subsequently, the specimens were sectioned and tag length was determined using a confocal laser scanning microscope (CLSM). Results were statistically compared with ANOVA followed by Scheffe's and Bonferroni/Dunn post hoc tests. RESULTS: With a mean penetration depth of 68 +/- 22 microm, tags in demineralized enamel were significantly longer than in other groups (p < or = 0.01). Penetration decreased significantly in remineralized areas or when fluoride was used (p < or = 0.01), but was still significantly deeper than in control sites (p < or = 0.01). CONCLUSION: Penetration of an unfilled resin into enamel was considerably influenced by the degree of dental hard tissue mineralization. Penetration was increased in demineralized enamel; however, remineralized enamel also allowed good penetration of the bonding agent. PMID- 15293420 TI - Comparison of microtensile bond strength to enamel and dentin of human, bovine, and porcine teeth. AB - PURPOSE: To determine the bond strengths promoted by an adhesive system to human, bovine, and porcine enamel and dentin, and compare their etched micromorphology by scanning electron microscopy. MATERIALS AND METHODS: Thirty sound freshly extracted teeth were used in this study: ten human third molars, ten bovine incisors, and ten porcine molars. The crowns of human (H), bovine (B), and porcine (P) teeth were ground with 600-grit SiC paper to expose either enamel (E) or mid-depth dentin (D) surfaces. After application of the adhesive resin, composite crowns approximately 8 mm high were built up with TPH Spectrum composite. After 24 h of water storage, specimens were serially sectioned in the buccal-lingual direction to obtain 0.8 mm slabs, which were trimmed to an hourglass shape of approximately 0.8 mm2 at the bonded interface. Specimens were tested in tension in a universal testing machine (0.5 mm/min). Results were statistically analyzed with ANOVA and Tukey's test at the 95% confidence level. RESULTS: Tukey's test showed significant differences between bond strengths obtained on enamel and dentin (p < 0.05). However, there were no statistically significant differences in microTBS between human, bovine, and porcine teeth. SEM observations revealed a similar dentinal morphology for the three species. However, porcine enamel specimens presented a very different distribution of enamel prisms. CONCLUSION: Bovine teeth proved to be possible substitutes for human teeth in either dentin or enamel bond testing. However, even though porcine teeth provided enamel and dentin bond strengths similar to human and bovine teeth, enamel morphology presented a very different configuration. PMID- 15293421 TI - The effect of sodium hypochlorite on microleakage of composite resin restorations using three adhesive systems. AB - PURPOSE: The aim of this study was to evaluate the effect of three different adhesive systems on microleakage of Class V restorations after the use of sodium hypochlorite. MATERIALS AND METHODS: One-hundred eighty bovine incisors were selected and randomly divided into 9 groups (n = 20): G1: Single Bond (SB); G2: 10% NaOCl solution (NS) + SB; G3: 10% NaOCl gel (NG) + SB; G4: Prime & Bond NT (PB); G5: NS + PB; G6: NG + PB; G7: Gluma One Bond (GOB); G8: NS + GOB; G9: NG + GOB. Standardized Class V cavities were prepared. All teeth were etched with 37% phosphoric acid for 15 s. In groups 2, 5, and 8, a 10% NaOCl solution was applied for 60 s to the dentin, and in groups 3, 6, and 9, a 10% NaOCl gel was applied to dentin for 60 s. All cavities were restored with composite resin Definite. The specimens were thermocycled for 1000 cycles (5 degrees C to 55 degrees C) and then immersed in 2% buffered solution of methylene blue for 4 h. The specimens were sectioned and analyzed according to a ranking score (0 to 4). Kruskal-Wallis and Mann-Whitney non-parametric tests (p < or = 0.05) were used for statistical analysis. RESULTS: The NaOCl treatment significantly increased microleakage at the dentin margin (p = 0.0129) as shown by the following sums of ranks: G1 = 1008.0a; G4 = 1301.5ab; G3 = 1687.0ab; G7 = 1744.0bc; G2 = 1802.0c; G9 = 1880.0c; G5 = 1889.0c; G8 = 1950.0c; G6 = 1963.0c (different superscripts indicate significant differences). For enamel, there were no statistically significant differences among the groups (p > 0.05). CONCLUSION: Depending on the adhesive system used, the application of NaOCl increased microleakage along dentin margins. PMID- 15293423 TI - Microleakage of a microhybrid composite resin using three different adhesive placement techniques. AB - PURPOSE: To evaluate the efficacy of two adhesive systems in reducing microleakage when applied with three different adhesive placement techniques. MATERIALS AND METHODS: Sixty freshly extracted caries-free human premolars and molars were used. MO/DO Class II standardized preparations were performed with the gingival margin placed 1 mm above the CEJ. Teeth were randomly divided into 2 groups (group I: Prime& Bond NT, Dentsply/Caulk; group II: Single Bond, 3M Espe). Each group was divided into 3 subgroups: (A) application of 2 coats and one cure: IA-IIA; (B) 2 coats and 2 cures of each adhesive system: IB-IIB; and (C) one coat of each adhesive along with the manufacturers' B1 flowable resin (0.5-mm thick layer) cured together at once: IC-IIC. Each coat was cured for 20 s at 800 mW/cm2 using a quartz-tungsten halogen light (Elipar Trilight, 3M ESPE). Teeth were then restored using 2-mm increments of an A2 microhybrid composite (Esthet-X, Dentsply/Caulk). All teeth were stored in distilled water at 37 degrees C for 24 h, thermocycled (500x, 5 degrees to 55 degrees C, 30 s dwell) and then placed in a 0.5% methylene blue dye solution for 24 h at 37 degrees C. Samples were sectioned longitudinally and evaluated for microleakage at the gingival margin under a stereomicroscope at 20x magnification. Dye penetration was scored using an ordinal scoring system, where 0: no penetration; 1: enamel penetration; 2: gingival dentin penetration; 3: axial dentin penetration. Kruskal-Wallis and Mann Whitney tests were used. RESULTS: A Mann-Whitney U-Test revealed no statistically significant difference between subgroups. Although not statistically significant, P&B NT (two coats and one cure) revealed the lowest microleakage scores. CONCLUSION: In the experimental model adopted for this study, microleakage was not affected either by the adhesive or its placement technique. PMID- 15293422 TI - Microtensile bond strength to root canal vs pulp chamber dentin: effect of bonding strategies. AB - PURPOSE: The aim of this in vitro study was to compare the microtensile bond strength (MTBS) between root canal and pulp chamber dentin with two bonding strategies (self-etching primer and total-etch technique). MATERIALS AND METHODS: The pulp chamber of four human canines was accessed and the pulp chamber and root canal prepared with spiral drills. The teeth were cut into halves parallel to the long axis of the tooth and randomly assigned to two groups: Clearfil SE Bond + AP X(SE) (Kuraray) and SingleBond + Filtek Z250(SB) (3M ESPE). After 24 h in water storage, the specimens were cut perpendicular to the root into 1.0-mm slices with a low-speed diamond saw. Specimens were trimmed to obtain hourglass shapes with a bond area of 1.0 mm2 (n = 12). Three specimens were obtained from the cervical root canal dentin (R) and from the coronal pulp chamber dentin (C). The MTBS was measured in a Bencor device with an Instron machine at a crosshead speed of 0.5 mm/min. The data was analyzed with two-way ANOVA and Tukey LSD post hoc (p < 0.05). RESULTS: MTBS in MPa (mean +/- SD) were: SB.C = 25.3 (+/-6.5)a; SB.R = 16.9 (+/-6.0)b; SE.C = 16.9 (+/-3.4)b; SE.R = 16.8 (+/-5.3)b. Means with the same letter are not statistically different at p < 0.05. When data were pooled for "dentin region", coronal pulp chamber dentin resulted in statistically higher bond strengths than root canal dentin (p < 0.013). CONCLUSION: Bonding to pulp chamber dentin seems to be more predictable than to root canal dentin. In the former region, the total-etch technique may result in a higher bond strength. PMID- 15293424 TI - Depth of cure of LED vs QTH light-curing devices at a distance of 7 mm. AB - PURPOSE: To determine the depth of cure of 5 blue LED curing devices compared to that obtained with 3 QTH curing devices. MATERIALS AND METHODS: The LED curing devices tested were 1) e-Light: 40 s; 2) Elipar FreeLight: 40 s; 3) Elipar FreeLight 2: 20 s and 40 s; 4) Ultra-Lume LED 2: 20 s and 40 s; 5) LEDemetron 1: 20 s and 40 s. The QTH curing devices tested were 1) Optilux 501: standard light guide 20 s and 40 s, turbo light guide 20 s; 2) Elipar TriLight: 40 s; 3) Astralis 10: 20 s. Surface hardness was measured (Zwick Z2.5/TS1S) 10 min after exposure on the top and bottom surface of resin samples (Tetric Ceram A3, 1 to 5 mm; 0.5 mm increment, diameter 5 mm, n = 9) which were cured at a distance of 7 mm from the bottom of the sample to the light-guide tip to simulate a Class II curing situation. A reference sample was cured under direct contact with the light guide. The reference sample with the greatest top surface hardness of all devices measured served as the overall control. A bottom/top surface hardness ratio of > or = 80% of the reference sample cured at zero distance was defined as clinically acceptable for safe curing. A descriptive statistical analysis was carried out. RESULTS: With QTH lamps, the mean maximum resin composite sample thickness which cured sufficiently (relative surface ratio > or = 80%) was: 3 mm for Optilux 501, standard light guide, 40 s; 2.5 mm for Trilight, 40 s; and 1.5 mm for Astralis 10, 20 s. The first-generation LED curing devices FreeLight and GC e-Light, both applied for 40 s, and the Optilux 501 operated for 20 s with the standard and the turbo light guide could not sufficiently cure a 1-mm-thick sample at a distance of 7 mm. The new FreeLight 2 and the Ultra-Lume LED 2 cured resin samples up to 2.5 mm thick in 40 s with a relative surface ratio > or = 80%, while no sufficient depth of cure was found after 20 s exposure time for the FreeLight 2. However, a 1.5-mm depth of cure with the Ultra-Lume LED 2 and the LEDemetron 1 with the 13/11 mm light guide was obtained after 20 s. The LEDemetron 1 equipped with a 13/8 mm light guide reached a depth of cure of 2.0 mm. No significant difference was found between the Elipar FreeLight 2, Ultra Lume LED 2, and LEDemetron 1 in their overall curing potential (linear statistical model, 5% level, Bonferroni-correction) given 40 s or 20 s of exposure time. CONCLUSION: Application of the first-generation LED curing devices FreeLight and e-Light did not ensure clinically sufficient depths of cure, while the new high-power LED curing devices FreeLight 2, Ultra-Lume LED 2, and LEDemetron 1 showed a curing potential equal to the Optilux 501, given 40 s of exposure time. PMID- 15293425 TI - In vivo study of the marginal integrity of composite resin buildups after full crown preparation. AB - PURPOSE: The purpose of this in vivo study was to evaluate the marginal integrity of composite resin buildups after full crown preparation, and to identify possible changes during the provisional phase of the manufacturing process of the final restorations. MATERIALS AND METHODS: Two test groups were determined for the use of an autocuring (Clearfil New Bond, Kuraray) and a light-curing (Optibond FL, Kerr) dentin adhesive. After rubber-dam application, both dentin adhesives were used with the total-etch technique. Then, an autocuring composite resin was applied for the core buildup. Impressions of the built-up teeth were taken directly after preparation and before cementation of the final restorations. During the provisional phase, which lasted from 7 to 28 days, the prepared teeth were restored with cemented provisional crowns. Replicas of the built-up teeth were manufactured and examined with a scanning electron microscope. RESULTS: For the autocuring dentin adhesive, only one buildup showed a gap 10 microm wide and 100 microm long. For the light-curing dentin adhesive, two samples revealed gaps that were 10 microm wide and had lengths of 100 microm and 75 microm, respectively. No change could be perceived when comparing the state of the composite/tooth interface after preparation and before final cementation. CONCLUSION: The results show that when using an autocuring composite resin in combination with dentin adhesives, nearly gap-free margins of composite buildups can be achieved and therefore might serve as a sound basis for the final restoration. PMID- 15293426 TI - One-year clinical evaluation of two polyacid-modified resin composites (compomers) in posterior permanent teeth. AB - PURPOSE: The aim of this study was to evaluate the clinical performance of two polyacid-modified resin composites (Dyract AP and F2000) in posterior teeth after 1 year of clinical service. MATERIALS AND METHODS: Seventy-two Class I restorations were performed in 33 patients (average age 25 years) by one operator. Eighty-two per cent of the restorations were located in molars. Before the proceedings, patients were informed about the aim of the study and they gave written consent to participate. At least one restoration of each material was placed in each individual. The materials were applied according manufacturer's instructions. Following finishing and polishing, one examiner performed the clinical baseline examination using the adapted USPHS system. To be included in the clinical trial, a restoration had to be rated "Alpha". After one year, 25 patients were recalled and 56 restorations were evaluated using the adapted USPHS system. RESULTS: All restorations were classified as clinically satisfactory (Alpha or Bravo). However, there was a decrease in restoration quality compared to baseline. Statistical analysis (chi2 and Fisher's exact test) demonstrated differences only in relation to superficial roughness, with exhibiting F2000 more surface roughness than Dyract AP (p < 0.01). CONCLUSION: Based on the methodology employed, all restorations were satisfactory after one year and the two materials performed similarly, except for the surface roughness criteria. PMID- 15293427 TI - The study of values: an unconventional approach to cross-cultural and social psychiatry. PMID- 15293428 TI - Cross-national study of attitudes towards seeking professional help: Jordan, United Arab Emirates (UAE) and Arabs in Israel. AB - BACKGROUND: Help-seeking processes provide critical links between the onset of mental health problems and the provision of professional care. But little is known about these processes in the Arab world, and still less in transnational, comparative terms. This is the first study to compare help-seeking processes among Muslim Arab female students in Jordan, the United Arab Emirates and Israel. AIMS: The present study compares the attitudes of Arab Muslim female students from Israel, Jordan and the United Arab Emirates (UAE) towards mental health treatment. METHOD: A convenience sample of 262 female Muslim-Arab undergraduate university students from Jordan, United Arab Emirates (UAE) and Arab students in Israel completed a modified Orientation for Seeking Professional Help (OSPH) Questionnaire. RESULTS: Data revealed that nationality was not statistically significant as a variable in a positive attitude towards seeking professional help; year of study, marital status and age were found to be significant predictors of a positive attitude towards seeking help. High proportions of respondents among the nationalities referred to God through prayer during times of psychological distress. CONCLUSIONS: The discussion considers implications for professional service delivery and programme development. Future research could extrapolate findings to other Arab countries and to Arab peoples living in the non-Arab world. PMID- 15293429 TI - The therapeutic relationship in the treatment of severe mental illness: a review of methods and findings. AB - AIMS: To review the methods and findings from studies of the therapeutic relationship (TR) in the treatment of severe mental illness. METHOD: A literature search was conducted to identify all studies that used an operationalised measurement of the TR in the treatment of severe mental illness. RESULTS: Fifteen scales--the majority of which were developed for psychotherapy--and the expressed emotion index have been used. Most scales have acceptable internal, inter-rater and test-retest reliability. As none of the scales has been used in more than five studies, no single scale is widely established in psychiatric research. A more positive relationship consistently predicts a better short- and long-term outcome. It appears that a large global factor accounts for the greatest proportion of the variance in the therapeutic relationship. CONCLUSIONS: The therapeutic relationship is a reliable predictor of patient outcome in mainstream psychiatric care. Valid assessments may need to take account of different, specific aspects of the relationship in psychiatric settings such as greater heterogeneity of treatment components and goals, increased variability of setting and the statutory responsibility of the clinician. Methodological progress may require conceptual work to ensure valid assessments of this central element of treatment. PMID- 15293430 TI - Sex roles and marital adjustment in Indian couples. AB - BACKGROUND: Marital theorists suggest a link between sex role differences and close relationships for men and women. Marriage is often a context for the activation and expression of sex roles. As marital adjustment is influenced by complementarity of roles between husband and wife, the same could hold true for sex roles as well. AIM: To study the relationship between sex roles and marital adjustment in Indian couples. METHODS: The sample consisted of 20 distressed and 20 non-distressed couples from a marital and family therapy centre in the city of Bangalore, India. The measures used included a sociodemographic data sheet, the Dyadic Adjustment Scale, the Bem Sex Role Inventory and a semi-structured interview schedule for gendered experiences. Means, percentages and ANOVAS were used to analyse statistically the data. Content analysis was applied on material from the semi-structured interview schedule. RESULTS: The study revealed that: (a) the group as a whole showed greater femininity than masculinity; (b) more non distressed individuals show high androgyny; (c) androgynous dyads show better marital adjustment; and (d) qualitative analysis suggests a trend for couples to move towards more gender-neutral constructions of marriage. CONCLUSIONS: The results indicate a link between androgyny and marital adjustment. The results also suggest the type of match between dyads. PMID- 15293431 TI - Factors associated with hopelessness: a population study. AB - BACKGROUND: Hopelessness is associated with depression and suicidality in clinical as well as in non-clinical populations. However, data on the prevalence of hopelessness and the associated factors in general population are exiguous. AIMS: To assess the prevalence and the associated factors of hopelessness in a general population sample. METHODS: The random population sample consisted of 1722 subjects. The study questionnaires included the Beck Hopelessness Scale (HS), Beck Depression Inventory (BDI), Toronto Alexithymia Scale (TAS-20) and Life Satisfaction Scale (LS). RESULTS: Eleven percent of the subjects reported at least moderate hopelessness. A poor financial situation (OR 3.64), poor subjective health (OR 2.87) and reduced working ability (OR 2.67) independently associated with hopelessness. Moreover, the likelihood of moderate or severe hopelessness was significantly increased in subjects dissatisfied with life (OR 5.99), with depression (OR 4.86), with alexithymia (OR 2.37) and with suicidal ideation (OR 1.85). CONCLUSIONS: This study demonstrated a moderately high prevalence of hopelessness at the population level. Hopelessness appears to be an important indicator of low subjective well-being in the general population that health care personnel should pay attention to. PMID- 15293432 TI - The stigma of mental illness: patients' anticipations and experiences. AB - BACKGROUND: There are studies that either deal with the stigmatization patients anticipate or with patients' concrete stigmatization experiences. Up until now, however, research is short of studies that investigate both aspects of subjective stigmatization simultaneously. AIMS: This study aims at investigating to what extent patients with schizophrenia or depression anticipate and experience stigmatization and how this is influenced by the type of mental disorder and the social environment. METHOD: A total of 210 patients with schizophrenia or a depressive episode were interviewed, one half living in a city and the other in a small town. RESULTS: Most of the patients expect negative reactions from the environment, particularly as concerns the access to work. Concrete stigmatization experiences were most frequently reported in the domain of interpersonal interaction. Even though schizophrenia patients and patients with depression anticipated stigmatization similarly frequently, the former reported concrete stigmatization experiences more frequently than the latter. Conversely, patients living in a small town anticipated stigmatization more frequently than patients from the city, even though both had actually experienced stigmatization at a similar rate. CONCLUSION: The results underline the necessity to differentiate between anticipated and experienced stigmatization. This is highly relevant for planning interventions aimed at reducing the stigma of mental disorder. PMID- 15293433 TI - Differences between indigenous Samis and Norwegians in mental disorders and background factors: patients listed as under treatment at the DPS-Lakselv (County District Psychiatric Centre), Finnmark, Spring 2000. AB - BACKGROUND: Earlier comparisons of adult Samis and Norwegians living within the same area are--thus far--unknown concerning mental well being/health and possible background factors. AIM: To compare patients from the two communities in search of differences which may be useful in planning health services. METHOD: Retrospective and anonymised data collected from journals. Comprises 48 'Samis' and 70 'Norwegians' in 108 'demographic cells' according to ethnicity, sex, municipality, education and age. RESULTS: 'Samis' rated higher in schizophrenia/psychosis (0.05 < p < 0.1) and crises (0.1 < p < 0.25), 'Norwegians' rated higher in anxiety disorders (0.001 < p < 0.005), low self esteem (0.005 < p < 0.01), illegal drugs (0.01 < p < 0.025) and in identity/existential problems (0.1 < p < 0.25). 'Samis' appeared to have suffered more at provincial boarding schools (0.005 < p < 0.01), while 'Norwegians' presented more mental illnesses in their families (0.025 < p < 0.05), more family break-ups (0.05 < p < 0.1) and earlier alcohol/drug abuse (0.05 < p < 0.1). CONCLUSION: Significant differences were found. 'In the black box' results will raise interesting questions for further research, but will also be useful for planning health services. PMID- 15293434 TI - Household survey of psychiatric morbidity in Cambodia. AB - AIMS: To estimate the prevalence of psychiatric symptoms in the Kampong Cham province and to determine the association between these symptoms and an impaired social functioning. METHODS: Cross-sectional cluster sample survey conducted among adults randomly selected within 50 clusters distributed over the province. METHODS: Of the respondents, 42.4% reported symptoms that met the Diagnostic and Statistical Manual of Mental Disorders, 4th edition criteria for depression, 53% displayed high anxiety symptoms and 7.3% met posttraumatic stress disorder (PTSD) criteria. Posttraumatic symptoms of intrusion and avoidance were present in 47.8% and 45.4% respectively. When reviewing comorbidities, 29.2% had depression and anxiety symptoms, 16.5% anxiety symptoms, 6.1% depression and 7.1% had triple comorbidity (PTSD, depression and anxiety). Regarding social functioning, 25.3% reported being socially impaired. Respondents with comorbid symptoms for depression, anxiety and PTSD were associated with an increased risk for social impairment compared with others. Being over 65 years and having experienced violent events were other factors associated with social impairment. CONCLUSION: Five years after the return of a more stable context in Cambodia, the prevalence of psychiatric symptoms in the community remains high. In addition, these symptoms are strongly associated with social impairment. This suggests that beyond psychosocial programs, the implementation of adapted clinical psychiatric care should be considered as a priority. PMID- 15293435 TI - Oral health of psychiatric in-patients in Hong Kong. AB - BACKGROUND: Poor oral health has been reported among various psychiatric populations. Little is known regarding the oral health among psychiatric patients in Asia. AIMS: To examine the oral health status of a group of Chinese psychiatric in-patients in a long-term rehabilitation facility. METHODS: A dental survey using the WHO standardised dental evaluation form was conducted in adult psychiatric patients in a rehabilitation programme. A qualified dentist examined all consenting patients. RESULTS: Ninety-one patients (64.8% male; mean age: 44.7 +/- 9.9 years; mean length of illness: 20.3 +/- 11.5 years) were included in the study, the majority (80.2%) diagnosed with schizophrenia. Malocclusion was found in 79.1% of patients. The mean number of missing teeth was 9.5 +/- 8.9. Bleeding on probing, calculus, shallow and deep pockets were found in 7.1%, 71.8%, 72.9% and 28.2% of patients, respectively. Dental caries were found in 75.3% of dentate patients. The mean number of caries per patient was 5.5 +/- 6.1. Fifty-four per cent of patients needed dental extraction and 78.8% required conservative dental treatment. Older age and length of illness were significantly associated with poor dental health. CONCLUSIONS: Oral health status of chronic psychiatric patients seems to be considerably worse than that of the general population. Mental health professionals should pay more attention to preventive oral health habits of psychiatric patients. PMID- 15293436 TI - [From triumphant medicine to humility]. PMID- 15293437 TI - [Heart arrest: which reality and which training for the practitioner?]. AB - Sudden death related to out-of hospital cardiac arrest is an important cause of mortality, which is mainly caused by ventricular fibrillation, a potentially reversible condition. The prognosis of out-of-hospital cardiac arrest remains dismal despite well developed emergency medical services. Witnessed arrest, ventricular fibrillation as the initial arrhythmia, cardiopulmonary resuscitation and early defibrillation are systematically associated with better survival. Key interventions must therefore be enforced to improve survival from out-of-hospital cardiac, introducing the concept of a "chain of survivals". The aim of the present article, which is illustrated by local results, is to review this important public health issue, to emphasize the role of the general practitioner in the chain of survival, and to promote education and training of basic and advanced life support. PMID- 15293438 TI - [Pre-hospital management of acute myocardial infarction: weaknesses]. AB - The overall mortality of acute myocardial infarction (AMI) is still high, half of deaths occurring in the prehospital setting. The American Heart Association (AHA) recommendations strongly emphasize the rapid use of "Emergency Medical Services". Despite this guideline, we frequently observe patients with AMI arriving at the hospital without an emergency ambulance service (SMUR). We undertook a retrospective study to quantify this problem with special interest in the mean of transport, in the role of the primary care practitioner and in the influence of the SMUR on the in-hospital delay. We had 125 AMI in 2000 and 2001: 42 reached the hospital by self-transportation, 57 by ambulance with SMUR and 26 by ambulance without SMUR. An out-hospital doctor was first warned by half of the patients 38% of which arrived by ambulance with SMUR, 37% by ambulance without SMUR and 25% by their own means. A thrombolysis was applied in one third of the AMI: the median prehospital and in-hospital delays were 120 and 40 minutes respectively. The in-hospital delay was significantly shorter when the emergency ambulance service was used. Our results confirm a suboptimal utilisation of the emergency ambulance service for AMI, due to patient and out-hospital doctors. This encourages us to go on providing specific information to the population but also to the primary care practitioners. PMID- 15293440 TI - [Severe acute asthma]. AB - Acute severe asthma is defined by the occurrence of an acute exacerbation resistant to the initial medical treatment, complicated by life-threatening respiratory distress due to severe lung hyperinflation. The conventional therapeutic approach is based on oxygen therapy and on the combined treatment of inhaled beta2-agonists at repeated doses and systemic corticosteroids. Inhaled or systemic magnesium sulfate is also recommended. The unresponsiveness to the initial bronchodilating therapy and the development of respiratory distress requiring intubation significantly increases mortality, due to the complications induced by mechanical ventilation. In these situations, a ventilatory strategy, including controlled hypoventilation with permissive hypercapnia, aiming at preventing lung hyperinflation, is indicated. Non-invasive ventilation may be successful in certain patients and represents an effective alternative to intubation. In ventilated patients, helium-oxygen mixtures can be considered as adjunctive therapies. After having reviewed the basic pathophysiological principles, this article will focus on the current medical treatment and of the modalities of mechanical ventilation in acute severe asthma. PMID- 15293439 TI - [New therapeutic strategies in severe sepsis and septic shock]. AB - Severe sepsis and septic shock are an important cause of mortality. Until recently, in spite of major progresses in our understanding of the pathogenic mechanisms of this syndrome, clinicians had only a limited therapeutic arsenal. Considerable efforts have been made in the past few years to develop novel therapeutic interventions to reduce mortality in sepsis. So far, five specific therapies have proven their efficacy to achieve such goal in large randomised controlled trials: early goal-directed therapy, recombinant activated protein C, moderate doses of steroids, low tidal volume ventilation in acute respiratory distress syndrome and intensive insulin therapy to control hyperglycemia. This review will focus on these recently acquired therapeutic modalities, that are presently available to clinicians for the treatment of severe sepsis and septic shock. PMID- 15293441 TI - [Value of non-invasive ventilation in intensive care]. AB - Noninvasive ventilation (NIV) refers to the delivery of mechanical ventilation using a nasal or facial mask. Compared to mechanical ventilation with endotracheal intubation, the occurence of complications, mainly infectious, is reduced by NIV. Reduction of respiratory workload and improvement of gas exchange are achieved with the use of NIV. In patients with exacerbations of chronic obstructive pulmonary disease (COPD), NIV reduces the need for endotracheal intubation, the length of ICU stay and the mortality. It is equally effective in acute cardiogenic pulmonary edema and for ventilatory weaning of patients with COPD. In selected groups of patients with acute hypoxemic respiratory failure, NIV diminishes the rate of endotracheal intubation and improves survival. The purpose of this article is to review the mechanisms, the technical aspects and the indications of NIV for the treatment of acute respiratory failure. PMID- 15293442 TI - [Infective endocarditis: update]. AB - The pathogenesis of infective endocarditis (IE) is being dissected at the molecular level, which should help redefine new preventive and therapeutic strategies against IE. In spite of improving health care, the incidence of IE has not decreased over the last decades. While classical predisposing conditions such as rheumatic heart disease were being eradicated, new features of IE have emerged. These include IE in intravenous drug users, IE in elderly patients with sclerotic valve disease, prosthetic valve IE and nosocomial IE. The epidemiology, pathogenesis, diagnosis, prevention and treatment of IE are being reviewed in this article. PMID- 15293443 TI - [Management of upper digestive hemorrhage in resuscitation: contribution of interventional radiology]. AB - Blood loss due to acute gastro-intestinal bleeding is a frequent cause of admission in critical care units. Rapid diagnosis and effective treatment are the determinant prognostic factors. The initial therapeutic approach in the acute settings consists of combined medical and endoscopic treatment. The recurrence of haemorrhage and the development of haemorrhagic shock significantly increase morbidity and mortality, and depend on the extent of the lesion and on patient co morbidities. Until recently, recurrent or refractory haemorrhage required urgent surgical therapy, which was associated with significant mortality. In the last years, the development of interventional radiology techniques has offered to clinicians an alternative, effective and less invasive therapeutic approach for the treatment of acute blood loss. In this article, we will show the outstanding role of interventional radiology in the treatment of acute gastro-intestinal bleeding. PMID- 15293444 TI - [Epidemiology, diagnosis, and treatment of pulmonary embolism]. AB - Pulmonary embolism (PE) is a frequent pathology, the mortality of which remains elevated despite the efficacy of anticoagulation. This is mainly due to diagnostic difficulty, resulting from the low sensibility and specificity of clinical signs and routine exams in PE. The best diagnostic approach relies on decision making algorithms based on the determination of a clinical probability, which in turn dictates the choice of specialized exams (C-scan/scintigraphy, venous duplex, angiography and D-dimers). Risk stratification, which is based on the presence of hemodynamic alterations (right ventricular dysfunction, shock) is essential to guide therapy. In case of shock (massive embolism), treatment relies on systemic thrombolysis. In the presence of right ventricular dysfunction without shock (sub-massive embolism), the advantage of thrombolysis over anticoagulation alone has not been clearly demonstrated. In all other cases, as long as no contraindication exists, therapeutic anticoagulation with heparin must be initiated as soon as PE is suspected, since the mortality of this condition is highest in the two first hours following presentation. PMID- 15293445 TI - [Cervical nodules: diagnosis and management]. AB - Family physicians frequently encounter patients with neck lumps. The causes are numerous but in the adult the origin is most often a lymph node, the majority of which are malignant. Inappropriate management may often lead to a very poor outcome. Relevant investigations must therefore be correctly chosen. The risk for a neck lump to be malignant depends mainly on age, male sex, and alcohol and tobacco consumption and to a lesser extent on a family history for head and neck malignancy. Careful medical history looks for symptoms such as dysphagia, pain, dysphonia, otalgia, or weight loss. On physical examination, the location, size, consistency and mobility of the mass is described. A careful inspection of the scalp, skin of the face and mucosal surface of the upper aerodigestive tract is performed followed by palpation. If no inflammatory or tumoral lesion is identified, the next step is to perform a fine needle aspiration biopsy of the neck mass which will most often lead to a definite diagnosis. When this is not the case, an otolaryngology consultation and excisional biopsy should be obtained. PMID- 15293446 TI - [Unilateral laryngeal paralysis]. AB - Investigations, diagnosis and treatment of laryngeal paralysis depend upon history taking and examination of phonation, swallowing and of the pharyngo larynx. In unilateral paralysis, the main symptom is dysphonia. Dysphagia lasting more than 10 days may indicate a proximal vagus nerve lesion. Voice and swallowing therapy may be undertaken. If this remains insufficient after one month, a temporary or definitive vocal fold medialisation may be considered. Paralysis is considered to be definitive if lasting for more than 12 months. A minimal of one-year follow-up is indicated in case of idiopathic paralysis. PMID- 15293447 TI - [Vertigo]. PMID- 15293448 TI - Zentralblatt fur bakteriologie--100 years ago: streptococcal mastitis of cows- long-lasting dilemmas in diagnosis and nomenclature. PMID- 15293449 TI - The omptin family of enterobacterial surface proteases/adhesins: from housekeeping in Escherichia coli to systemic spread of Yersinia pestis. AB - The omptins are a family of enterobacterial surface proteases/adhesins that share high sequence identity and a conserved beta-barrel fold in the outer membrane. The omptins are multifunctional, and the individual omptins exhibit differing virulence-associated functions. The Pla plasminogen activator of Yersinia pestis contributes by several mechanisms to bacterial invasiveness and the systemic, uncontrolled proteolysis in plague. Pla proteolytically activates the human proenzyme plasminogen and inactivates the antiprotease alpha2-antiplasmin, and its binding to laminin localizes the uncontrolled plasmin activity onto basement membranes. These properties enhance bacterial migration through tissue barriers. Pla also degrades circulating complement proteins and functions in bacterial invasion into human epithelial cells. PgtE of Salmonella enterica and OmpT of Escherichia coli have been shown to degrade cationic antimicrobial peptides from epithelial cells or macrophages. PgtE and SopA of Shigella flexneri appear important in the intracellular phases of salmonellosis and shigellosis, whereas functions of OmpT have mainly been associated with protein degradation in E. coli cells. The differing virulence roles and functions have been attributed to minor sequence variations at the surface-exposed regions important for substrate recognition, to the dependence of omptin functions on lipopolysaccharide, and to the different regulation of omptin expression. PMID- 15293451 TI - Intracellular survival of persistent group A streptococci in cultured epithelial cells. AB - Group A streptococcus (GAS) is the principle etiologic agent of bacterial pharyngotonsillitis and a wide range of other diseases. Failure to eradicate GAS from patients has been documented in 5-30% of patients with pharyngotonsillitis, in spite of the continued sensitivity of GAS to penicillin and other beta lactams. It was recently proposed that eradication failure might be attributed to the ability of GAS to maintain an intracellular reservoir during antibiotic treatment. We have previously shown that strains derived from patients with bacterial eradication failure, despite antibiotic treatment (persistent strains), adhered to and were internalized by cultured epithelial cells more efficiently than strains that were successfully eradicated. Since, penicillin and other beta lactams do not penetrate well into mammalian cells, intracellular survival of GAS is crucial in order to persist during prolonged antibiotic treatment. In this study, we compared the survival of GAS strains from cases of eradication failure and eradication success, using an epithelial cell culture model. We found that persistent strains show significantly increased intracellular survival, compared to the 'eradication success' strains. This finding supports the idea that an intracellular reservoir of GAS plays a role in the etiology of antibiotic eradication failure. PMID- 15293450 TI - Reduced expression of the global regulator protein CsrA in Legionella pneumophila affects virulence-associated regulators and growth in Acanthamoeba castellanii. AB - Legionella bacteria have a developmental cycle in which they go from existing in the aquatic environment to replicating inside eukaryotic host cells. The adaptation to the new environment requires an efficient regulatory system. Overexpression of CsrA, a global regulatory protein found in a variety of gram negative bacteria has been shown to suppress virulence-associated traits in Legionella pneumophila. Since evidence resulting only from overproduction may not be sufficient to validate the role of a regulatory protein, a csrA mutant strain, CsrA(-), with a drastically reduced production of CsrA, was created. Using RNA slot blots and Western blotting it was shown that fliA and flaA, genes which contribute to flagellation, were expressed early in the mutant. Additionally, in CsrA(-) the levels of the stationary-phase sigma factor, RpoS, and a recently described regulator of virulence traits, LetE, were increased. Growth curves of CsrA(-) bacteria were delayed with pigment production occurring at the same OD578 but at reduced levels in the mutant. Replication ability of the CsrA(-) mutant in amoebae was also affected. Based on these results, we could show that CsrA is involved in the regulation of the bacterial switch from the replicative to the transmissible form. PMID- 15293452 TI - Protection of mice against challenge with Bacillus anthracis STI spores after DNA vaccination. AB - Immune responses against the protective antigen (PA) of Bacillus anthracis are known to confer immunity against anthrax. We evaluated the efficacy of genetic vaccination with plasmid vectors encoding PA, in protecting mice from a lethal challenge with B. anthracis STI spores. BALB/c and A/J mice were immunized via gene gun inoculation, using eukaryotic expression vectors with different cellular targeting signals for the encoded antigen. The vector pSecTag PA83, encoding the full-length PA protein, has a signal sequence for secretion of the expressed protein. The plasmids pCMV/ER PA83 and pCMV/ER PA63, encoding the full-length and the physiologically active form of PA, respectively, target and retain the expressed antigen in the endoplasmic reticulum of transfected cells. All three plasmids induced PA-specific humoral immune responses, predominantly IgG1 antibodies, in mice. Spleen cells collected from plasmid-vaccinated BALB/c mice produced PA-specific interleukin-4, interleukin-5, and interferon-gamma in vitro. Vaccination with either pSecTag PA83 or pCMV/ER PA83 showed significant protection of A/J mice against infection with B. anthracis STI spores. PMID- 15293453 TI - PCR-enzyme immunoassay of rDNA in the diagnosis of candidemia and comparison with amplicon detection by agarose gel electrophoresis. AB - We have developed a semi-nested PCR-enzyme immunoassay (snPCR-EIA) for the detection of Candida species in serum specimens, and the sensitivity of amplicon detection was compared with the detection of amplified product by agarose gel electrophoresis (AGE). The universal outer primers amplified the 3' end of 5.8S and the 5' end of 28S rDNA including the internally transcribed spacer 2 (ITS2) in PCR with genomic DNA as template from all the tested Candida species. The biotin-labeled species-specific primers derived from ITS2 from the four commonly encountered Candida species, viz. C. albicans, C. tropicalis, C. parapsilosis and C. glabrata, together with digoxigenin-labeled reverse primer amplified species specific DNA in the reamplification step of the snPCR. The snPCR-EIA was positive for genomic DNA recovered from 0.06 Candida cells in culture and one organism/ml in spiked serum specimens. Evaluation of snPCR-EIA and snPCR-AGE for specific identification of Candida species with 26 clinical Candida isolates showed 100% concordant results with Vitek and ID32C yeast identification systems. Further evaluation of snPCR-EIA and snPCR-AGE for detection of Candida species in serum samples from culture proven (n = 6) and suspected (n = 10) patients showed concordance with the corresponding species isolated in culture. The serum samples from none of the healthy volunteers (n = 10) were positive for the presence of Candida DNA by snPCR-EIA or snPCR-AGE. Our results show that the snPCR-EIA has the same sensitivity as snPCR-AGE, however, it offers additional advantages of simultaneous testing of a large number of serum samples and avoids the use of ethidium bromide, a potent mutagen. The snPCR-EIA could, therefore, be a method of choice for the diagnosis of candidemia. PMID- 15293454 TI - Growth factor production in human endothelial cells after Chlamydia pneumoniae infection. AB - Seroepidemiological and histopathological studies have suggested a link of atherosclerosis with chronic Chlamydia pneumoniae infection. The present study was designed to examine the effect of C. pneumoniae on expression of basic fibroblast growth factor (bFGF) and platelet-derived growth factor (PDGF) in human endothelial cells. Using reverse transcription-polymerase chain reaction, immunoblotting and enzyme-linked immunosorbent assay we found stimulation of bFGF expression depending on the number of infecting bacteria and the time of infection as well. This stimulatory effect was diminished by heat and UV light treatment of the chlamydial inoculum, suggesting that viable bacteria but not bacterial LPS may be essential for eliciting this growth factor. In contrast, the expression of both PDGF-A and PDGF-B was not increased following C. pneumoniae infection. This study demonstrates that C. pneumoniae activates endothelial cells to produce bFGF, a growth factor which is linked to the development of atherosclerotic plaques. PMID- 15293455 TI - ST64B is a defective bacteriophage in Salmonella enterica serovar Typhimurium DT64 that encodes a functional immunity region capable of mediating phage-type conversion. AB - The Salmonella enterica subsp. enterica serovar Typhimurium (S. Typhimurium) defective bacteriophage ST64B has a putative immunity (immC) region consisting of cI, cro and cII-like genes. Since ST64B is widespread in S. Typhimurium, studies were undertaken to determine whether this region might be functional and influence phage typing results. Cloning of ST64B immC-like genes and their subsequent expression in S. Typhimurium DTs showed that this region is able to mediate phage-type conversion in DTs 41 and 44. This confirms the functionality of the immC region and the patterns of lysis produced by phage typing are consistent with the predicted mechanism of action of the encoded protein products. PMID- 15293456 TI - [Separation of Bacteroides forsythus ATCC43037 proteins by horizontal two dimensional gel electrophoresis]. AB - OBJECTIVE: To set up a rapid, efficient, reliable and accurate method for separation of Bacteriodes forsythus proteins. METHODS: Bacteroides forsythus ATCC43037 cells were harvested by centrifugation, washed in Tris buffer to remove excess medium, and lysed by sonication. The sonicated lysis proteins were extracted step by step with ReadyPrep Sequential Extraction Kit (Bio-Rad). The supernatant of B. forsythus proteins were used for two-dimensional gel electrophoresis. The first dimension IEF (isoelectric focusing) was run with Immobiline DryStrip (pH 3-10) and the second dimension SDS-PAGE was run with Excelgel SDS, gradient 8-18 precast gel and buffer strips. The separated proteins were stained with Coomassie Brilliant Blue and silver staining kit (Plusone Silver Staining Kit, Protein, Pharmacia Biotech). RESULTS: 1. The protein spots were clear when sample cups were used in the middle of IPG strip during IEF. 2. B. forsythus proteins were separated clearly by horizontal two-dimensional electrophoresis and most of B. forsythus whole-cell proteins were acidic proteins (P13-7). CONCLUSION: Horizontal two-dimension electrophoresis is a useful method for separating B. forsythus proteins. PMID- 15293457 TI - [Expression of proliferating cell nuclear antigen/bcl-2 in gray gingival tissue]. AB - OBJECTIVE: To study the expression of PCNA/bcl-2 protein in gray and normal gingival tissue, and to investigate the biological effect of procelain-fused-to metals (PFM). METHODS: Gray gingival tissue animal model was established by PFM prosthesis and immunohistochemistry S-P method was used to detect the PCNA/bcl-2 protein expression in gingival tissue after 3 months, 6 months in PFM groups and control group. RESULTS: The expression of PCNA had significant difference between the 3th month group and the control group (P < 0.05); no significant difference was found between the 6th month group and the control group (P > 0.05). The expression of bcl-2 had no significant difference between the 6th month group and the control group (P > 0.05). CONCLUSION: There were a correlation between the expression of PCNA and inflammation of the gray gingival tissue; The expression of PCNA and bcl-2 protein does not indicate that gray gingival tissue had dysplasia change. PMID- 15293459 TI - [Recombinant human BMP-2 accelerates bone formation of mandibular distraction osteogenesis in rabbits]. AB - OBJECTIVE: To study the effects of rhBMP-2 on bone formation of mandibular distraction osteogenesis in rabbits. METHODS: Bilateral mandibular osteotomies were performed in 12 mature rabbits. 5 mg rhBMP-2 with the collagen carrier was implanted in the osteotomy site of one side of the mandibles. Only the collagen sponge was placed in the contra-lateral side as control. The mandibles of 8 rabbits were lengthened by 6mm using a custom-made distractor. At 4 weeks after the end of distraction, all animals were killed and the distracted calluses were harvested and processed for histological, scanning electron microscopic, as well as Ca/P ratio analysis. RESULTS: The regenerated bone was found in the distraction gap after mandibular lengthening. The mandibular side treated with rhBMP-2 had greater amounts of new bone formation and earlier mineralization than contra-lateral side (non-rhBMP-2 treated). CONCLUSION: Recombinant human BMP-2 appears to be able to accelerate bone formation of mandibular distraction osteogenesis in rabbits. PMID- 15293458 TI - [Preparation of lymphatic targeting dosage form: pingyangmycin absorbed on activated carbon nanoparticles]. AB - OBJECTIVE: To prepare anticancer nanoparticles for targeting therapy for oral cancer lymph node metastasis. METHODS: The activated carbon nanoparticles (CH-NP) were prepared for drug carrier. Pingyangmycin (PYM), a high sensitive anticancer drug for oral squamous cell carcinoma, were selected as model drug. The activated carbon nanoparticles and PYM were mixed with saline and shaken 20 minutes so that PYM was absorbed on activated carbon enough, resulting in a new formulation of PYM (PYM-CH-NP). The absorbency of PYM on activated carbon nanoparticles was evaluated. RESULTS: The diameter distribution for CH-NP ranged form 136 nm, to 540 nm, the average diameter was 176 nm. The proportion of CH-NP to PYM was increased and more absorbency of PYM on activated carbon nanoparticles was achieved. CONCLUSION: The activated carbon nanoparticles has high absorbency of PYM. The new formulation PYM-CH-NP can be used as targeting therapy of cervical lymph node metastasis by peri-cancer submucosal injection. PMID- 15293460 TI - [Effect of fas gene transfection on tumourigenicity and proliferation of transplanted tumor of oral squamous cell carcinoma cell line Tca8113]. AB - OBJECTIVE: To investigate the effect of fas gene transfection and monoclonal anti fas antibody on tumorigenicity and proliferation of transplanted tumor of Tca8113 cell. METHODS: Plasmid including fas gene was transfected into Tca8113 cell by lipofectamine kit. Some transfected cells were treated by monoclonal anti-fas antibody after 48 hours since transfection. Untransfected cell (control), fas tansfected cell and fas-transfected cell treated with antibody were transplanted to nude mice subcutaneously. Growth of transplanted tumor was observed and recorded regularly. Animals were sacrificed and tumor samples were harvested at the end of experiment. Fas expression in each neoplasm was assessed by RT-PCR. Apoptosis, proliferation and expression of fas protein in tumor tissue were measured by flow cytometry (FCM). RESULTS: Tumor occurred much later in fas transfected group and fas-transfected plus antibody treated group. Growth arrest was found in them. RT-PCR and FCM suggested that fas-transfection up-regulated the expression of fas mRNA and protein, increased apoptosis index (AI). But no effect on proliferation index (PI) was observed. Monoclonal anti-fas antibody did not effect the expression of fas mRNA and protein, but increased AI and decreased PI. CONCLUSION: Fas-transfection suppressing tumorigenesis of Tca8113 cell transplanted in nude mice might be caused by up-regulation of expression of fas gene and enhancement of apoptosis. However, anti-fas antibody suppressing tumorigenesis might be associated with activation of apoptosis and repression of proliferation. PMID- 15293461 TI - [FEM study on displacement, position of rotation center and stress distribution of PDL under various loading force systems]. AB - OBJECTIVE: To investigate the displacement, the position of rotation center and the stress distribution of PDL under different loading force system (Moment/Force, M/F) by simulating clinical loading force system. METHODS: A three dimensional finite element model of upper central incisor, which consists of 945 isoparametric elements and 1,245 nodes was developed. The displacement, the position of the rotation center and the stress distribution of PDL were analyzed under 13 types of loading force system. RESULTS: 1. Different force system led to different types of tooth movement. When M/F= -9.15:1, -10.30 - -10.50:1 and 10.90:1, it brought the result of controlled tipping movement, the bodily movement and the root movement; 2. The graph of the center rotation was a hyperbolic asymptotic line: Mx/Fy = -10.50 (horizontal axis) and L = 6.75 (vertical axis). Moreover, a little change of M/F between -9.15 and -10.90 led to apparent change of the position of rotation center. 3. The maximum strain and stress during the tipping movement were 1.47 x 10(-2) MPa and -2.81 x 10(-2) MPa, and during the bodily movement the results were 1.10 x 10(-2) MPa and -1.86 x 10( 2) MPa, while during the root movement were 0.96 x 10(-2) MPa and -2.58 x 10(-2) MPa. CONCLUSION: 1 . Different force system (M/F) leads to different types of tooth movement. 2. It is necessary to adjust the force system accurately to obtain prescient tooth movement, especially when M/F changes between -9.15:1 and 10.90:1. 3. This study suggested that the tooth movement style and the force system (M/F) should be controlled to protect the periodontal tissue. PMID- 15293462 TI - [The effect of left bacteria in the root canal on prognosis of the root canal therapy]. AB - OBJECTIVE: To study the effect of the left bacteria on the root canal therapy. METHODS: 50 single-rooted teeth with chronic apical periodontitis were divided into two groups, one was instrumented with step-back technique and 2.5%NaOCl ultrasonic irrigation for 3 min, then filled with Thermafil. Samples were taken after instrumentation to culture. The other was treated with traditional RCT at three visits. RESULTS: In 24 months the apical radiolucency were greatly reduced in all cases. There weren't significant relationship among the postoperative pain and the left bacteria, the degree of the obturation or the pre-operative symptoms (P > 0.05). CONCLUSION: The effect of left bacteria in root canal filled with Thermafil wasn't observed. PMID- 15293463 TI - [Compositive treatment of square face]. AB - OBJECTIVE: To investigate the treatment of square face with compositive methods. METHODS A total of 71 patients with square-shape face were treated. According to the different face skeleton and desire of patient, mandible angle curved osteotomy and mentoplasty were used as main methods and zygomatoplasty, buccal fat pad resecting and other aesthetic methods as assistant methods to recontour the whole face skeleton. RESULTS: The face skeleton of all patients was improved with satisfaction. The following-up period was 6 months to 2 years. In this patients group, massive haemorrhage was occurred during operation in one patient, mental nerve of of one side was injured in two patients, the lip mucosa was wounded in five patients. CONCLUSION: Mandible angle curve-osteotomy and mentoplasty combined with other assistant aesthetic operations were ideal methods to recontour square face. The result was satisfactory. PMID- 15293464 TI - [A retrospective study on 615 cases of minor salivary gland tumors]. AB - OBJECTIVE: To analyse the clinical and pathological characteristics of minor salivary gland tumors. METHODS: A retrospective analysis on 615 cases of intraoral minor salivary gland tumors from 1990-2002 with a confirmed pathologic diagnosis was carried out. RESULTS: In 615 cases of minor salivary gland tumors, 265 cases were benign, 350 cases malignant. Pleomorphic adenoma was the most common entity and accounted for 81.1% of all benign tumors. Adenoid cystic carcinoma comprised 32.9% of the malignant sample and was the most frequent malignant tumor. The principal location was the palate. Female was a little more of the benign and a male prevalence was observed on the malignant tumor. The mean age of patients with benign and malignant tumors were 40.9 and 49.1 years old, respectively (P < 0.05). CONCLUSION: We consider extraordinary manifestation as histopathology, tumor, primary location, age and sex, presenting the results of a review of our experience with those minor salivary gland tumors. PMID- 15293465 TI - [Application of facial canal dissection for recovery of facial nerve after operation of parotid carcinoma]. AB - OBJECTIVE: To study the recovery method of facial nerve function and to compare the clinical effect after operation of parotid carcinoma, which invades stylomastoid foramen and peripheral bone in deep lobe. METHODS: Three operation methods were taken: (1) The tumor, parotid, invasive facial nerve and bone around the tumor were resected with transplantation of facial nerve. (2) Tumor parotid and facial nerve were resected without transplantation of facial nerve. (3) Tumor and parotid were dislocated from facial nerve, keeping the continuity of facial nerve. RESULTS: For the first method, facial nerve function of 68.2% patients came back to the patients without facial paralysis before operation, while facial nerve function of 16.7% patients came back to the patients with facial paralysis before operation. There was obvious difference between them (P < 0.05). To the patients with facial paralysis before operation, the first and the second method were taken. The ratio of local control was 33.3% and 10.0% respectively. And survival for 5 years were 25.0% and 10.0% respectively. There was no obvious difference between the two methods (P > 0.05). To the patients without facial paralysis before operation, three methods of operation were taken. The ratio of local control was 77.3%, 48.0% and 33.3% respectively. And survival for 5 years were 86.4%, 52.0% and 41.7% respectively. There was obvious difference between the first method and the other two (P < 0.05). There was no obvious difference between the second and the third methods (P > 0.05). CONCLUSION: Facial canal dissection in operation of parotid carcinoma with recovery of facial nerve can not only resect tumor completely, but also fit for development of functional surgery. It is an ideal method for surgery operation. PMID- 15293466 TI - [Treatment of Bell's palsy with combination of traditional Chinese medicine and western medicine]. AB - OBJECTIVE: To evaluation the clinical effect of combination of traditional Chinese medicine and western medicine on Bell's palsy. METHODS: 83 patients with Bell's palsy were randomly divided into two groups (trail group 54 cases and control group 29 cases). Patients in two groups were treated with medicine, acupuncture, physiotherapy, while patients in the trail group were treated with massage and functional exercise as the same time. The results of both groups were evaluated according to Portmann's Simple Scale. RESULTS: The score before treatment of trail group was 2.907 +/- 1.794, while control group was 2.931 +/- 2.034. And the score after treatment of trail group was 18.593 +/- 1.743, while control group was 9.862 +/- 3.091. Score of the function of facial muscles obtained from trail group was distinctly higher than that was from the control group (P < 0.01), as well as the improvement index (P < 0.01, trail group: 0.844 +/- 0.095, control group: 0.712 +/- 0.129). CONCLUSION: There is significant curative effect and suitability in the treatment of Bell's palsy with combination of traditional Chinese medicine and western medicine. The improvement of facial muscles' motive function pre- and post-treatment and quantitative evaluation of curative effect can be objectively obtained by evaluation of facial muscles' function. PMID- 15293467 TI - [Clinical study on the effect of Vitapex paste in apexification]. AB - OBJECTIVE: The purpose of this article was to observe the effect of Vitapex Paste in apexilication. METHODS: 64 younger permanent teeth with underdeveloped apex and necrotic pulp were randomly selected. After root canals were prepared and sterilized, Vitapex paste was used in the apexification. All the teeth were observed for three years. RESULTS: 24 teeth (37.5%) were successful, 37 teeth (57.81%) were progressive, 3 teeth (4.69%) were failed. Altogether 61 teeth were effective, the effective rate was 95.31%. CONCLUSION: Vitapex paste was effective for the younger permanent teeth in the apexification. PMID- 15293468 TI - [Research of cranio-occlusional change of skeletal class III malocclusion in permanent dentition treated by the multiloop edgewise arch wire technique]. AB - OBJECTIVE: To analyze the mechanics in correction of skeletal class III malocclusion with Multiloop Edgewise Arch Wire (MEAW). METHODS: 15 patients with skeletal class III malocclusion were treated with MEAW technique. Cephalometric analysis was performed with pre-treatment and post-treatment cephalograms. Paired t-test was conducted to assess the treatment effects. RESULTS: L6-XI decreased by 2.87 mm, L6/MP increased by 8.60 degrees, L1-XI decreased by 2.60 mm, OP/MP increased by 2.33 degrees. Skeleton changed a little. There was no significant change in the soft tissue. CONCLUSION: (1) Dento-alveolar compensation is the main change after the treatment by MEAW technique; (2) The improvement in molar relationship and overjet is achieved with upright and distal movement of the lower posterior teeth; (3) The lower anterior teeth moved lingually and protracted. Occlusal plane is flattened. PMID- 15293469 TI - [Clinic significance of morphometric study on squamous cell carcinoma of floor of mouth]. AB - OBJECTIVE: The purpose of this study was to investigate the role of cell morphometric factor (FC) and the average density of light in squamous cell carcinomas at the floor of mouth by immunohistochemical method (SABC). METHODS: The slices made from the paraffin specimens of 28 patients with squamous cell carcinomas at the floor of mouth were stained by the immunohistochemical technique with monoclonal antibody CK10 and CK14. In these stained slices the FC parameters and the average density of light were measured by using an image analysis system. RESULTS: In primitive tumors, the FC parameters and the average density of light were significantly different between positive and negative lymph nodes metastasis or between different pathological grades (P < 0.05 or < 0.001). The significance was not existed among clinical stages (P > 0.05). The difference of FC expressed in positive and negative lymph nodes metastasis were significant (P < 0.05 or < 0.001). CONCLUSION: The cell morphometric factor (FC) parameters and the average density of light of CK10 and CK14 could be used as a biological guideline to evaluate the clinically infiltrative and metastasic potential of squamous cell carcinoma at the floor of mouth. PMID- 15293470 TI - [The computer-aided design and manufacture of unilateral orbital defect restoration]. AB - OBJECTIVE: By reverse engineering and rapid prototyping techniques to found a new design method of maxillofacial restoration. METHODS: By laser scanning apparatus the plaster face model was scanned and the primitive face point data were acquired. With the reverse engineering software, the point data were reconstructed to one smooth face surface image and the defect orbital tissue shape data was obtained from the normal contralateral tissue data in the software. The model designed the three-dimensional data of defect part and the rapid prototyping technique made the resin orbital restoration. RESULTS: The laser scanning apparatus acquired the distinct and precise model data of the plaster face-model. The Digisurface retrograde engineer software succeeded to fulfill the unilateral orbital defect computer-aided design. The orbital restoration inosculated the plaster model tightly and symmetrically. CONCLUSION: The reverse engineering software and rapid prototyping technique could finish the computer-aided design and manufacture of the unilateral orbital defect restoration smoothly and satisfactorily. PMID- 15293471 TI - [Restoring ceramo-metal tooth by using self-made crown post-core in order to re utilize the broken crown of the base tooth]. AB - OBJECTIVE: To explore a new method and its clinical significance of restorative ceramo-metal by using a self-made crown post-core to reutilize the broken crown. METHODS: 18 teeth were restored with ceramo-metal tooth by using self-made crown post-core in order to preserve the old broken base tooth. The patients' own feelings and clinical effect were observed. RESULTS: Followed up for 1-3 years, 83.3% (15 teeth) of the patients were successful. Among the group one was failed because of bad condition of post and caused split of tooth root. The second failure was caused by uneven force by coherent paste uneven. The third failure was caused by tight biting force and caused the tooth broken. CONCLUSION: Restoring ceramo-metal tooth by using self-made crown post-core in order to re utilize the broken crown of the base tooth is convenient and low cost and is suitable for basic hospitals. PMID- 15293472 TI - [In vitro culture and biological characteristics of cranial neural crest stem cell]. AB - OBJECTIVE: According to development biology, cranial neural crest stem cell (CNCSC) can differentiate into precursor cells of tooth, jaw and peripheral nerve system, but in vitro study is less reported. In the present study, CNCSC were dissociated and cultured in vitro, and the biological characteristics of CNCSC was investigated. METHODS: Cranial neural tubes, dissected from mouse E9d, were explanted onto fibronectin-coated dishes. CNCSCs emigrated from the explanted neural tubes, cultured in a serum-free medium containing modified DMEM/F12. Biological characteristics of CNCSC were detected by morphology, nuclear labeled with BurdU and immunocytochemistry. RESULTS: Fibroblast-like CNCSCs demonstrated the characteristics of stem cell, such as clonality, self-renewal and multipotentiality. The result of immunocytochemical stain showed that CNCSC expressed HNK-1 antigen. CONCLUSION: CNCSC were cultured successfully, providing a experimental basis for study on tooth/jaw-like differentiation of CNCSC in vitro, especially providing a cell source for investigating tooth/jaw regeneration. PMID- 15293473 TI - [Establishment of a culture system of chick embryo for mouse tooth germ development]. AB - OBJECTIVE: To establish a new culture system for mouse tooth germs in chick embryo. METHODS: The mandibular first molar germ fragments of 15 embryonic days' Kunming mouse embryo were implanted into the lateral mesenchyme of 4-5 days' chick embryo wing buds in ove. Eggs were reincubated and implanted tissues were examined by histochemistry. RESULTS: The cultured tooth germ development continued from cap stage to latest bell stage. The ameloblast and the odontoblast all differentiated maturely and secreted matrix. CONCLUSION: 4-5 days' wing buds chick embryo could serve as developing the mouse tooth germs and demonstrate well physiological process of differentiation and morphogenesis. PMID- 15293474 TI - [Investigation of the expression of calcitonin receptor mRNA in human osteoclasts on deciduous teeth]. AB - OBJECTIVE: To investigate the expression of calcitonin receptor mRNA in the osteoclasts of the resorbing deciduous teeth. METHODS: After fixing the collected deciduous teeth, toluidine blue was performed and tartrate-resistant acid phosphatase (TRAP) staining was used to identify the osteoclasts on the resorbing surface of human deciduous teeth and in situ hybridization of calcitonin receptor mRNA to show its existence. RESULTS: There were a number of TRAP positive osteoclasts on the root surface which showed the expression of calcitonin receptor mRNA. CONCLUSION: On the resorbing surface of human deciduous teeth there are osteoclasts that express calcitonin receptor mRNA, so it is feasible to use this kind of osteoclast to test the effect of external factors on the expression of CTR mRNA. PMID- 15293475 TI - [Centrosome hyperamplificationin oral precancerous lesions and squamous cell carcinomas]. AB - OBJECTIVE: It is currently considered that the defect of mitotic spindle caused by centrosome abnormalities may be one of the reasons for the development of aneuploidy in tumors. This study attempted to elucidate the possible role of centrosome defects in the development and progression of OSCC by investigating the frequency of centrosome amplification in oral precancerous lesions and OSCC. METHODS: Formalin-fixed, paraffin-embedded tissues of 12 cases of normal oral epithelium, 22 case of dysplasia with different degree epithelium dysplasia and 32 cases of OSCC with different differentiation were investigated for centrosome status by using indirect immunofluorescence double staining with antibodies to centrosome protein gamma-tubulin and cytokeratin. The differences and the change trend of centrosome status in these groups were statistically analyzed by SPSS10.0. RESULTS: Normal oral epithelium showed normal centrosomes in epithelium cells, while 16 of 22 cases (72.73%) of dysplasia (DYS) and 27 of 32 cases (84.38%) of OSCC showed the evidence of centrosome amplification and morphological abnormalities characterized by huge size, clump or supernumerary centrosomes in a fraction of epithelium or tumor cells. The percentage of cells with abnormal centrosomes increased gradually from mild-dysplasia epithelium to poorly differentiated OSCC, which positively correlated with the histologicalcytologic grade of oral precancerous lesions and OSCC (P < 0.01). CONCLUSION: Centrosome amplification was an early event and that might play a role in the establishment and perhaps the progression of OSCC. There might be some direct relationship between centrosome defects and the cellular morphological phenotype characteristics of dysplasia and OSCC. Centrosome amplification could be served as an alternative diagnostic indicator of dysplasia and the intervention of centrosome cycle might serve as a particular way for the prevention and treatment of OSCC in the future. PMID- 15293476 TI - [The morphological study of bone-implant interfaces in vivo]. AB - OBJECTIVE: To evaluate the bone-implant interfaces of two kinds of implants with different surfaces in different time in vivo. METHODS: CDIC and ITI-TPS solid screw cylinder pure titanium implants were selected and implanted in the regions of posterior molars of rhesus monkeys. 1 month, 2 months, 3 months and 1 year after surgery, the bone-implant interfaces were evaluated respectively through oral examination, X-ray inspection, light microscope and scanning electron microscope (SEM) observation. RESULTS: None of the implants was loose. Soft tissue around implants appeared no inflammation. There were no apparent transparent shadow around the implants interfaces in X-ray photos except little angle-shaped absorption was showed in neck region of CDIC implants of one-month. New bone was observed around implants of one-month through light microscope and SEM. More bone growing around ITI implants were seen than that around CDIC implants except the interfaces of one-year. CONCLUSION: The osseointegration of ITI implants are better than that of CDIC implants during three months after implanting without loading. The bone formation at the interfaces of ITI and CDIC implants has no significant difference after one year without loading. PMID- 15293477 TI - [The effect of intraperitoneal heparin on distribution of ACC-M in lung and liver]. AB - OBJECTIVE: To investigate the effect of intraperitoneal heparin on the adhesion of highly lung metastasic adenoid cystic carcinoma cell line (ACC-M) in lung. METHODS: 3HTdR labeled ACC-M cells were injected by intravenous infusion after intraperitoneal injection with 200 units heparin. 4 mice of each group were killed at 2 h, 6 h and 18 h after infusion. The relative radioactivity (CPM) in lung and liver was detected. RESULTS: 3H-activity per gram in lung of heparin group was lower than control at the same time. There was significant difference between the two groups (P < 0.001). There was no difference between the two groups in liver (P > 0.05). CONCLUSION: The results of this study suggest that the highly lung metastasis characteristic of ACC-M may be partially inhabited by the use of intraperitoneal heparin. PMID- 15293478 TI - [Experimental study of periosteal osteoblasts adhesion to artificial bone scaffolds based on rapid prototype]. AB - OBJECTIVE: To study the biocompatibility of bone engineering scaffolds designed and fabricated by CAD and Rapid Prototyping techniques. METHODS: Infant rat calvarias osteoblasts were isolated and expanded in vitro and the cells (2nd passage) were seeded onto scaffolds with porosity 80%, 90%, 95% at a density of 2.06 x 10(9)/L. Cell adhesion number and morphology were measured with SEM after 4 days, 10 days co-culture. RESULTS: (1) The osteoblasts' adhesion amounts increased with culture time in three porosity group (P < 0.05), but the increase were different among three groups, 80% group was 0.35 x 10(5), 90% group was 2.84 x 10(5); (2) Through SEM observations, it showed that osteoblasts adhered to all scaffolds well. CONCLUSION: The scaffolds designed and fabricated by CAD and rapid prototyping own a good cellular biocompatibility. The results suggest the feasibility of using such scaffold fabricating method for bone tissue engineering research and clinical therapy. PMID- 15293479 TI - [The effect of cryotreat on tensile properties of medium melting-point and high melting-point castable alloy]. AB - OBJECTIVE: To introduce cryotreat technique into prosthetic dentistry by testing the tensile properties of CW-H Co-Cr-Mo cast alloy before and after cryotreat and to observe the image changes by SEM to study the mechanism that changes the tensile properties of the alloy. METHODS: 15 CW-H cast alloy were divided into 3 groups, i.e. control group (G1), cryotreated group (G2) and cryotreated plus post cryogenic treated group (G3). The gauge test technique was employed to test the modulus of elasticity. Then the strength and percentage elongation (PE) were tested. SEM images were used to analyze the mechanism that improved the tension properties of the alloy. RESULTS: For CW-H alloy the strength and the modulus of elasticity of both G2 and G3 were effectively increased but PE effectively decreased than G1. There was no effectively difference between G2 and G3, but G3 was larger than G2. SEM images of G2 and G3 showed that secondary-carbonide separated out all over the alloy. CONCLUSION: The results obtained above suggest that cryotreat is an effective method in enhancing tensile properties of CW-H Co Cr-Mo cast alloy. PMID- 15293480 TI - [Designing dental manpower index to evaluate dental manpower resources]. AB - OBJECTIVE: The purpose of this investigation is to find out a method to evaluate dental manpower resources. METHODS: We selected population, GDP, number of dentist and number of different oral health professionals from certain internet stations, published books and journals from 1996 to 2000 as our investigating data. RESULTS: Data was collected from 100 countries. Our investigation found that the design of dental manpower index to evaluate dental manpower resources was effective and convenient. CONCLUSION: Dental manpower index is a good method to evaluate dental manpower resources. PMID- 15293481 TI - Rehabilitation for Parkinson's disease: a systematic review of available evidence. AB - OBJECTIVE: To systematically review the available evidence on the effectiveness of nonpharmacological rehabilitation interventions for people with Parkinson's disease, and identify future research needs. DESIGN: Electronic searches of four databases (CINAHL, Cochrane Library, MEDLINE, PsychLIT) 1980-2002; examination of reference lists of relevant papers. Controlled trials and observational studies were included. Data extraction and quality assessment of papers by two independent reviewers. A narrative review. SETTING: Rehabilitation interventions delivered either in subjects' own homes, or in clinical settings as outpatients. SUBJECTS: Community-living adults with Parkinson's disease. INTERVENTIONS: Physiotherapy, occupational therapy, speech and language therapy, psychological counselling and support, and education. MAIN MEASURES: A range of outcomes: mobility, functional status, speech, swallowing, psychological well-being, as determined by the studies included in the review. RESULTS: Forty-four different studies (reported in 51 papers) were included (25 physiotherapy, 4 occupational therapy, 10 speech and language therapy, 3 psychological counselling, 1 educational, 1 multidisciplinary). All studies, except one, reported improvements on at least one outcome measure. CONCLUSIONS: Findings may reflect publication bias, but suggest interventions can affect patients' lives for the better in a variety of ways. It is difficult to interpret the clinical importance of statistically significant improvements reported in most studies. There is a need for methodologically more robust research with meaningful follow-up periods, designed in a manner that separates specific and nonspecific effects. Cost effectiveness evidence is required to provide clear guidance on service extensions. PMID- 15293482 TI - Animal-assisted therapy for middle-aged schizophrenic patients living in a social institution. A pilot study. AB - OBJECTIVES: To determine whether animal-assisted therapy is effective in the rehabilitation of middle-aged schizophrenic patients living in a social institution. DESIGN: A before and after study with nine-month treatment period. SETTING: Social institute for psychiatric patients. SUBJECTS: Seven schizophrenic patients living in the social institute. INTERVENTIONS: Weekly sessions of animal assisted therapy for a nine-month period, each therapeutic session lasting for 50 minutes. MEASURES USED: The Independent Living Skills Survey assessed by an independent rater. RESULTS: After the completion of the therapy significant improvement in the domestic and health activities occurred. CONCLUSIONS: Animal assisted therapy seems to be helpful in the rehabilitation of schizophrenic patients living in a social institution. PMID- 15293483 TI - Would the addition of TENS to exercise training produce better physical performance outcomes in people with knee osteoarthritis than either intervention alone? AB - OBJECTIVE: To examine if the addition of transcutaneous electrical nerve stimulation (TENS) to exercise training would produce better physical outcomes than TENS or exercise alone in people with knee osteoarthritis. DESIGN: Sixty-two subjects were randomly allocated to four groups. INTERVENTIONS: Patients received either (1) TENS, (2) placebo stimulation, (3) exercise training, or (4) TENS and exercise training five days a week for four weeks. MAIN OUTCOME MEASURES: The isometric peak torque, spatiotemporal gait parameters and range of knee movement were assessed in treatment session1, session10 and session20 and the four-week follow-up. RESULTS: By session20, the TENS and exercise group demonstrated an average of 26.6% cumulative gain in the knee extensor peak torque for the different knee positions (all p < 0.05). Although the between-group difference was short of being statistically significant, the gain found in the TENS and exercise group was greater than that found in the other three groups. The TENS and exercise group also tended to show greater cumulative increase in stride length (12.6%, p = 0.006), walking cadence (9.3%, p = 0.098) and gait velocity (22.4%, p = 0.034) than the other groups. By session20, it was the only group that produced a significant increase in the range of knee motion during walking (12.0%, p = 0.000). The TENS group and the exercise group both demonstrated some improvements in the above physical outcomes, but negligible change was found in the group receiving placebo stimulation (all p > 0.05). CONCLUSION: No significant difference was found among the four treatment protocols, but the addition of TENS to exercise training tended to produce the best overall improvement in physical outcomes in people with knee osteoarthritis. PMID- 15293484 TI - Home training with and without additional group training in physically frail old people living at home: effect on health-related quality of life and ambulation. AB - OBJECTIVE: To test the effect of two exercise regimes on health-related quality of life (HRQoL) and ambulatory capacity. DESIGN: Randomized controlled trial. SUBJECTS: Seventy-seven community-dwelling physically frail people over 75 years of age (mean = 81, SD = 4.5). INTERVENTIONS: Home training (HT, N= 38) comprised twice daily functional balance and strength exercises and three group meetings. Combined training (CT, N= 39) included group training twice weekly and the same home exercises. Interventions lasted 12 weeks. Physiotherapists ran both programmes. Home exercises were recorded daily. MAIN MEASURES: HRQoL was assessed by SF-36, and ambulatory capacity by walking speed and frequency and duration of outdoor walks. RESULTS: Following intervention, CT improved the SF-36 mental health index significantly more than HT (p = 0.01). The SF-36 physical health index (p = 0.002) and walking speed (p = 0.02) demonstrated improvements, but no group differences. Six months after cessation of intervention there was still overall improvements on the mental health index (p = 0.032), borderline overall improvements on the physical health index (p = 0.057), higher weekly number of outdoor walks for the CT group than for the HT group (p = 0.027) and an improved habitual walking speed in the CT group only (p = 0.022). CONCLUSIONS: HT improved HRQoL and walking speed, but additional group training gave larger benefits on mental health. Group training away from home may be beneficial for mental health and ambulatory capacity. PMID- 15293485 TI - A task-orientated intervention enhances walking distance and speed in the first year post stroke: a randomized controlled trial. AB - OBJECTIVE: To evaluate the efficacy of a task-orientated intervention in enhancing competence in walking in people with stroke. DESIGN: Two-centre observer-blinded stratified block-randomized controlled trial. SETTING: General community. SUBJECTS: Between May 2000 and February 2003, 91 individuals with a residual walking deficit within one year of a first or recurrent stroke consented to participate. INTERVENTIONS: The experimental intervention comprised 10 functional tasks designed to strengthen the lower extremities and enhance walking balance, speed and distance. The control intervention involved the practice of upper extremity activities. Subjects in both groups attended sessions three times a week for six weeks. MAIN MEASURES: Six-minute walk test (SMWT), 5-m walk (comfortable and maximum pace), Berg Balance Scale, timed 'up and go'. RESULTS: At baseline, subjects in the experimental (n = 44) and control (n = 47) groups walked an average distance of 209 m (SD = 126) and 204 m (SD =131), respectively, on the SMWT. Mean improvements of 40 m (SD =72), and 5 m (SD =66) were observed following the experimental and control interventions, respectively. The between group difference was 35 m (95% confidence interval (CI) 7, 64). Significant between-group effects of 0.21 m/s (95% CI 0.12, 0.30) and of 0.11 m/s (95% CI 0.03, 0.19) in maximum and comfortable walking speed, respectively, were observed. People with a mild, moderate or severe walking deficit at baseline improved an average of 36 (SD =96), 55 (SD = 56) and 18 m (SD = 23), respectively, in SMWT performance following the experimental intervention. CONCLUSIONS: Study findings support the efficacy of a task-orientated intervention in enhancing walking distance and speed in the first year post stroke, particularly in people with moderate walking deficits. PMID- 15293486 TI - The long-term outcomes from a randomized controlled trial of an educational behavioural joint protection programme for people with rheumatoid arthritis. AB - OBJECTIVE: To evaluate the long-term effects of joint protection on health status of people with early rheumatoid arthritis (RA). DESIGN: A four-year follow-up of a randomized, controlled, assessor-blinded trial was conducted. SETTING: Two rheumatology outpatient departments. PARTICIPANTS: People with rheumatoid arthritis less than five years since diagnosis. INTERVENTIONS: Two 8-hour interventions were originally compared: a standard arthritis education programme, including 2(1/2) hours of joint protection based on typical UK occupational therapy practice (plus 5(1/2) hours on RA, exercise, pain management, diet and foot care); and a joint protection programme, using educational-behavioural training. MAIN MEASURES: Adherence to joint protection, pain, hand pain on activity, Arthritis Impact Measurement Scales 2 and Arthritis Self-efficacy were recorded at 0 and 4 years. RESULTS: Sixty-five people attended the joint protection and 62 the standard programmes. Groups at entry were similar in age (51 years; 49 years), disease duration (21 months: 17.5 months) and use of nonsteroidal anti-inflammatory and disease-modifying drugs. At four years, the joint protection group continued to have significantly better: joint protection adherence (p=0.001); early morning stiffness (p=0.01); AIMS2 activities of daily living (ADL) scores (p=0.04) compared with the standard group. The joint protection group also had significantly fewer hand deformities: metacarpophalangeal (MCP) (p =0.02) and wrist joints (p=0.04). CONCLUSION: Attending an educational-behavioural joint protection programme significantly improves joint protection adherence and maintains functional ability long term. This approach is more effective than standard methods of training and should be more widely adopted. PMID- 15293487 TI - Can augmented physiotherapy input enhance recovery of mobility after stroke? A randomized controlled trial. AB - OBJECTIVE: To discover if the provision of additional inpatient physiotherapy after stroke speeds the recovery of mobility. DESIGN: A multisite single-blind randomized controlled trial (RCT) comparing the effects of augmented physiotherapy input with normal input on the recovery of mobility after stroke. SETTING: Three rehabilitation hospitals in North Glasgow, Scotland. SUBJECTS: Patients admitted to hospital with a clinical diagnosis of stroke, who were able to tolerate and benefit from mobility rehabilitation. INTERVENTION: We aimed to provide double the amount of physiotherapy to the augmented group. MAIN MEASURES: Primary outcomes were mobility milestones (ability to stand, step and walk), Rivermead Mobility Index (RMI) and walking speed. RESULTS: Seventy patients were recruited. The augmented therapy group received more direct contact with a physiotherapist (62 versus 35 minutes per weekday) and were more active (8.0% versus 4.8% time standing or walking) than normal therapy controls. The augmented group tended to achieve independent walking earlier (hazard ratio 1.48, 95% confidence interval 0.90-2.43; p=0.12) and had higher Rivermead Mobility Index scores at three months (mean difference 1.6; -0.1 to 3.3; p=0.068) but these differences did not reach statistical significance. There was no significant difference in any other outcome. CONCLUSIONS: A modest augmented physiotherapy programme resulted in patients having more direct physiotherapy time and being more active. The inability to show statistically significant changes in outcome measures could indicate either that this intervention is ineffective or that our study could not detect modest changes. PMID- 15293488 TI - Does motor imagery training improve hand function in chronic stroke patients? A pilot study. AB - OBJECTIVE: To assess the efficacy of motor imagery training for arm function in chronic stroke patients. The relation between mental processes such as attentional and perceived personal control over recovery, and motor imagery was additionally investigated. DESIGN AND SUBJECTS: Twenty patients with long-term motor impairments (mean two years post stroke), were assessed before and after four weeks of training. Ten patients mentally rehearsed movements with their affected arm. Their recovery was compared with patients who performed nonmotor imagery (n =5), or who were not engaged in mental rehearsal (n=5). SETTING: Patients were recruited from the stroke database of Ninewells Hospital, Dundee. Assessment and training were performed at the patients' home. INTERVENTIONS: The motor imagery group was asked to practise daily imagining moving tokens with their affected arm. The nonmotor imagery group rehearsed visual imagery of previously seen pictures. All patients practised physically moving the tokens. MAIN MEASURES: The following variables were assessed before and after training: motor function (training task, pegboard and dynamometer), perceived locus of control, attention control and ADL independence. RESULTS: All patient groups improved on all motor tasks except the dynamometer. Improvement was greater for the motor imagery group on the training task only (average of 14% versus 6%). No effect of motor imagery training was found on perceived or attentional control. CONCLUSIONS: Motor imagery training without supervision at home may improve performance on the trained task only. The relation between movement imagery, attention and perceived personal control over recovery remained unclear. PMID- 15293489 TI - The effect of an ankle-foot orthosis on walking ability in chronic stroke patients: a randomized controlled trial. AB - OBJECTIVE: Regaining walking ability is a major goal during the rehabilitation of stroke patients. To support this process an ankle-foot orthosis (AFO) is often prescribed. The aim of this study is to investigate the effect of an AFO on walking ability in chronic stroke patients. DESIGN: Cross-over design with randomization for the interventions. METHODS: Twenty chronic stroke patients, wearing an AFO for at least six months, were included. Walking ability was operationalized as comfortable walking speed, scores on the timed up and go (TUG) test and stairs test. Patients were measured with and without their AFO, the sequence of which was randomized. Additionally, subjective impressions of self confidence and difficulty of the tasks were scored. Clinically relevant differences based on literature were defined for walking speed (20 cm/s), the TUG test (10 s). Gathered data were statistically analysed using a paired t-test. RESULTS: The mean difference in favour of the AFO in walking speed was 4.8 cm/s (95% CI 0.85-8.7), in the TUG test 3.6 s (95% CI 2.4-4.8) and in the stairs test 8.6 s (95% CI 3.1-14.1). Sixty-five per cent of the patients experienced less difficulty and 70% of the patients felt more self-confident while wearing the AFO. CONCLUSIONS: The effect of an AFO on walking ability is statistically significant, but compared with the a priori defined differences it is too small to be clinically relevant. The effect on self-confidence suggests that other factors might play an important role in the motivation to use an AFO. PMID- 15293490 TI - Effects of specific rehabilitation for dizziness among patients in primary health care. A randomized controlled trial. AB - OBJECTIVE: To investigate whether specific rehabilitation for patients with dizziness has any effect on clinical balance measures and/or the apprehension of dizziness measured with a visual analogue scale (VAS). DESIGN: Randomized controlled trial. SUBJECTS: Forty-two patients, 50 years or older with dizziness of central or age-related origin, identified in primary health care. METHOD: The patients were randomized to either an intervention or a control group. The intervention included balance training and vestibular rehabilitation in group sessions twice a week for six weeks. All patients were assessed at baseline, after six weeks and after three months with five different balance measures and visual analogue scale. RESULTS: Statistically significant differences were found between the two groups comparing results at baseline and after six weeks regarding standing one leg eyes closed (SOLEC) on right foot (p =0.011). Results of SOLEC right foot after three months differed significantly between the groups (p=0.033) as did SOLEC left foot (p=0.035). No difference between the groups were found in the Romberg test, figure of eight, walking heel to toe, 'stops walking when talking', standing one leg eyes open or estimating the experience of dizziness measured with visual analogue scale. CONCLUSIONS: Balance training and vestibular rehabilitation improved the ability to stand on one leg with eyes closed in persons with dizziness aged 50 years or over. PMID- 15293491 TI - Differences in outcome of a multidisciplinary treatment between subgroups of chronic low back pain patients defined using two multiaxial assessment instruments: the multidimensional pain inventory and lumbar dynamometry. AB - OBJECTIVE: To investigate the effects of a multidisciplinary back school programme (Roessingh Back Rehabilitation Programme, RRP) compared with usual care, as well as differences in treatment outcome between subgroups defined using two multiaxial assessment instruments: the Multidimensional Pain Inventory (MPI DLV) and lumbar dynamometry. DESIGN: Randomized controlled trial. SETTING: Rehabilitation. SUBJECTS: One hundred and sixty-three patients with chronic, aspecific low back pain. INTERVENTION: All subjects were randomly assigned either to a multidisciplinary, physically oriented group treatment or to their usual care. MAIN OUTCOME MEASURES: The Roland Disability Questionnaire and health related quality of life (EuroQol, EQ5-D) were measured as primary outcomes before randomization and after eight weeks and six months follow-up. RESULT: Only 30-50% of the patients in the RRP group showed improvement and this number is not significantly different from the control group. Subgroup analyses give some first indications that multiaxial measurement instruments can be used to identify subgroups with differences in treatment effects. CONCLUSION: The overall effect of a multidisciplinary treatment is disappointing, however multiaxial assessment before admission might be valuable in clinical practice, resulting in more effective treatments for patients with chronic low back pain. PMID- 15293492 TI - Acute stroke unit care combined with early supported discharge. Long-term effects on quality of life. A randomized controlled trial. AB - OBJECTIVES: The aim of the present trial was to compare the effects of an extended stroke unit service (ESUS) with the effects of an ordinary stroke unit service (OSUS) on long-term quality of life (QoL). DESIGN: One year follow-up of a randomized controlled trial with 320 acute stroke patients allocated either to OSUS (160 patients) or ESUS (160 patients) with early supported discharge and follow-up by a mobile team. The intervention was a mobile team and close co operation with the primary health care service. All assessments were blinded. MAIN OUTCOME MEASURE: Primary outcome of QoL in this paper was measured by the Nottingham Health Profile (NHP) at 52 weeks. Secondary outcomes measured at 52 weeks were differences between the groups measured by the Frenchay Activity Index, Montgomery-Asberg Depression Scale, Mini-Mental State Score and the Caregivers Strain Index. RESULTS: The ESUS group had a significantly better QoL (mean score 78.9) assessed by global NHP after one year than the OSUS group (mean score 75.2) (p =0.048). There were no significant differences between the groups in the secondary outcomes, but a trend in favour of ESUS. Caregivers Strain Index showed a mean score of 23.3 in the ESUS group and 22.6 in the OSUS group (p=0.089). CONCLUSION: It seems that stroke unit treatment combined with early supported discharge in addition to reducing the length of hospital stay can improve long-term QoL. However, similar trials are necessary to confirm the benefit of this type of service. PMID- 15293494 TI - [Tomography reconstruction of optic parameters of biological objects based on the proportional medium approximation]. AB - Described in the paper is a reconstruction algorithm for special distributions of optical parameters of biological objects based on the non-stationary axial model of radiation transport in case of proportionality of the absorption and scattering coefficients. The proportionality coefficient, used in reconstruction, is shown to influence a precision of images being restored. The appropriate recommendations are defined of how to diminish the occurring distortions. PMID- 15293493 TI - [Integration of fundamental and applied medical and technical research made at the department of the biomedical systems, Moscow State Institute of Electronic Engineering]. AB - The integration results of fundamental and applied medical-and-technical research made at the chair of biomedical systems, Moscow state institute of electronic engineering (technical university--MSIEE), are described in the paper. The chair is guided in its research activity by the traditions of higher education in Russia in the field of biomedical electronics and biomedical engineering. Its activities are based on the extrapolation of methods of electronic tools, computer technologies, physics, biology and medicine with due respect being paid to the requirements of practical medicine and to topical issues of research and design. PMID- 15293495 TI - [Recovery of medical ultrasound images based on an effective deconvolution of scanning data]. AB - Described in the paper is a method of data deconvolution of ultrasound scanning (solution of the deconvolution equation in the linear metric) that ensures a higher precision of images being restored versus the traditional Wiener filtering. Simplex algorithm with an originally modified efficiency function was used to solve the task. The results of experiments testifying to the efficiency of algorithms in the ultrasound image processing of the mammary gland of a patient are presented. PMID- 15293496 TI - [Passive propagation of the transmembrane potential induced by the capacities of the myocardium bioelectrolytes in its electric defibrillation]. AB - The mechanism of passive propagation of transmembrane potential (TMP), caused by the availability of their own electric capacities in bioelectrolytes, is elucidated within the framework of the bi-domain description of the myocardium electrophysiological properties. The above mechanism is related with the possibility of the external and internal bioelectrolytes and has, in respect to each other, electric charges unbalanced by their absolute capacity; it is explained by the ability of such charges to diffuse rapidly enough through bioelectrolytes and, owing to their electric induction, to intensify the TMP. PMID- 15293497 TI - [A choice and realization of bipolar pulses for external electric defibrillators]. AB - There is a comparison of parameters of different-form bipolar pulses for external electric defibrillators described in the paper. The electric schemes of generators used in pulse shaping are elucidated; the appropriate recommendations concerning their choice are defined. PMID- 15293499 TI - [Broad-band analogue-to-digital converter of electrocardiograms]. AB - The use of a broad-band analogue-to-digital converter (ADC) with a reading rate of up to several kilohertz and with a subsequent digital filtration and decimation provides for obtaining the amplitude-and-frequency characteristics with the parameters, which are virtually unachievable for the ordinary analogue scheme technologies or digital filters inbuilt into the digital-filter integral ADC. Multi-digit ADC and digital signal processors (DSP) are used for the purpose. Additionally, with DSP it is possible to process algorithms of the broad band ECG, like isolation of the impulse of the cardiac pacemaker. PMID- 15293498 TI - [Comparison of the efficiency and noise-immunity of the algorithms detecting the shock cardiac rhythms]. AB - Comparative assessments of the efficiency and noise immunity of the algorithms detecting the shock cardiac rhythms (SCR) are described in the paper. It is suggested to use the rejection filter of ventricular fibrillation as the principle algorithm for the SCR detection in an automated external defibrillator. The use of the method combining the after-threshold interval and the method of spectral analysis is demonstrated to be preferable for a portable external defibrillator. PMID- 15293500 TI - [A telemedicine electrocardiography system based on the component-architecture soft]. AB - The paper deals with a universal component-oriented architecture for creating the telemedicine applications. The worked-out system ensures the ECG reading, pressure measurements and pulsometry. The system design comprises a central database server and a client telemedicine module. Data can be transmitted via different interfaces--from an ordinary local network to digital satellite phones. The data protection is guaranteed by microchip charts that were used to realize the authentication 3DES algorithm. PMID- 15293501 TI - [An integral chip for the multiphase pulse-duration modulation used for voltage changer in biomedical microprocessor systems]. AB - An integral chip (IC) was designed for controlling the step-down pulse voltage converter, which is based on the multiphase pulse-duration modulation, for use in biomedical microprocessor systems. The CMOS technology was an optimal basis for the IC designing. An additional feedback circuit diminishes the output voltage dispersion at dynamically changing loads. PMID- 15293502 TI - [A semiconductor photostimulator for electroencephalography with USB control]. AB - The paper contains a description of electroencephalography (EEG) photostimulator for examining the induced cerebral potentials. The device was designed to function within the EEG computer complexes; a high-intensity irradiation LED matrix is used as the light source. The 1.1 USB control bus serves as the communication interface; there is an extra external synchronization bus. It is possible to preset the frequency, brightness, on-off time ratio and pattern of light stimuli. A library of application programmer is supplied to monitor the photostimulation. PMID- 15293503 TI - [A signal generator for testing the electroencephalography and electrocardiography equipment]. AB - The paper contains a description of the "Neurotest7A" signal generator designed for testing the electroencephalography and electrocardiography equipment, including its functional and technical characteristics, structural scheme and operational algorithm. PMID- 15293504 TI - [Advantages and disadvantages of inactivated and live influenza vaccine]. AB - Published data related with comparison studies of safety, efficiency and some other properties of cold-adapted live influenza vaccine (LIV) and of inactivated influenza vaccine (IIV) are analyzed. LIV and IIV do not differ by systemic reactions after administration; however, it is not ruled out that there can be unfavorable reactions in vaccination of persons with allergy to the chicken embryo proteins as well as in cases of persistence/reversion of cold-adapted strain observed in vaccination of persons with primary impairments of the immune system. There are no convincing data, up to now, on that LIV is superior to IIV in coping with influenza pandemics. The efficiency of LIV and IIV for children aged 3 years and more and for healthy adults is virtually identical. Additional controllable field comparative studies of LIV and IIV efficiency in immunization of elderly persons are needed. Limited data on LIV efficiency for children aged 2 months and more were obtained. The need in a 2-stage vaccination of all age group with the aim of ensuring responses to all 3 LIV components is, certainly, a LIV disadvantage. In case of IIV, the 2-stage vaccination is needed only for persons who were not ill with influenza. The intranasal LIV administration has, from the practical and psychological standpoints, an advantage before the IIV administration by syringe. The ability of LIV to protect from the drift influenza virus variations could be its advantage before IIV; still, more research is needed to verify it. Transplantable cell lines meeting the WHO requirements could be an optimal substrate for the production of LIV and IIV. Children are the optimal age group for influenza prevention by cold-adapted LIV, whereas, IIV fits better for vaccination of adults and elderly persons. PMID- 15293505 TI - [Genetic characterisation of wild measles strains circulating in European part of Russia in 1998-2002]. AB - The primary structure of the N-gene COOH terminus of measles virus isolated, 1998 2000, in Russia's European part was investigated. The general analysis as well as an analysis of the primary gene structure showed the two group's isolates as belonging to the D4 genotype. A subsequent analysis of the primary structure of the N-gene COOH-terminus of Moscow/2002/61 isolated during the 2002 measles outbreak in Moscow also showed it as belonging to the D4 genotype. The obtained data are indicative of that the wild measles strains belonging to the D4 genotype have been recently circulating in Russia's territory. PMID- 15293506 TI - [Analysis of changes in the gp120 V3 region as observed in some patients with HIV infected from a common infection source]. AB - HIV-1 genome regions encoding the gp120 V3 part were sequenced in samples isolated from persons belonging to the category of those infected in the Rostov Elista outbreak and having the common infection source. Samples were obtained from 5 patients in 1992 and in 2001. A total of 27 sequences obtained in 1992 and 35 sequences obtained in 2001, 2 to 8 sequences for each patient, were analyzed. The diversity level of V3 sequences made, in some patients, 2.2% in 1992 and went up to 4.2% in 2001 samples (p < 0.07). The ratio between the synonymous and non synonymous substitutions was determined for the gp120 V3 region. The mean ratio value made 0.12 in 1992 samples and 0.23 in 2001 samples. The obtained data confirm the assumption, made previously in a population analysis, on the evolution of the gp120 V3 epitope towards substitution of the Lg initial structure in positions 14 and 15 (through intermediate stages represented by the IG and FG structures) for the FA structure. PMID- 15293507 TI - [The Siberian and Far-Eastern subtypes of tick-borne encephalitis virus registered in Russia's Asian regions: genetic and antigen characteristics of the strains]. AB - Agar gel precipitation test with cross-adsorbed immune sera was used for the antigenic differentiation of strains of tick-borne encephalitis virus (TBEV). Fifty strains of the Far East TBEV serotype and 46 strains of the Siberian (Aina) TBEV serotype were isolated from Ixodes persulcatus, which is the main vector of the above TBEV subtypes in the Asian and European parts of Russia. The fragment of the envelope protein gene was sequenced for TBEV strains. Sequences of new group strains of the Siberian subtypes isolated from 3 patients with chronic TBE and from brain tissues of 4 deceased patients were determined. Lethal TBE outcomes were registered in Siberia (Irkutsk Region and Krasnoyarsk Territory) and in Russia's European part (Yaroslavl Region). PMID- 15293508 TI - [The age-related specificity of site-oriented immunity in respiratory-syncytial virus infection]. AB - ELISA test systems were designed on the basis of synthetic peptides (SP) simulating the primary structure of functionally significant epitopes of the respiratory and cyncyntial virus (RCV) F-protein for the purpose of investigating the structure and age-related peculiarities of humoral immunity in respect to separate epitopes of RCV F-protein. One of them (221-232) simulates a part of RCV "virus-neutralizing domain" and another one (479-491) is highly important for the fusion mechanisms. New SP-based ELISA were used to examine pair sera in 159 patients with documented RCV infection including children, aged up to 3 years and 3 to 15, and adults. The activity of anti-RCV antibodies to SP was found to be significantly lower in children aged up to 3 years versus the older children and especially versus the adults. The virus neutralizing and, to a greater extent, fusion-inhibiting activities of antibodies were increasing with age, which collated with the results of detecting the antibodies to SP by immune-enzyme assay. The results testify to synchronism of formation of antibodies to different epitopes of the RCV F-protein. The shaping-up of antibodies with the above SP could denote the protective properties of humoral immunity, which justifies the use of the SP-based ELISA in its analysis, especially, in babies as well as in different-type immunodeficiency and immunopathology conditions. PMID- 15293509 TI - [HIV-1 serotyping of V3-mimicking peptides circulating in the Republic of Tajikistan among intravenous drug users]. AB - The paper summarizes the results of HIV-1 serotyping by using 56 positive sera collected in the territory of the Republic of Tajikistan (RT) from intravenous drug users (IVDU). It was made by solid-phase ELISA based on synthetic peptides, mimicking different variants of the apical epitope of the HIV-1 gp120 V3-loop. Two types of conjugates, those specific to human IgG and IgA, were used to detect the immune complexes. Serotypes, as determined according to IgG and IgA-based ELISA, coincided, however, the latter were proven to be more suitable for serotyping. There is a high level of HIV-1 serotype heterogeneity among IVDU in RT; altogether, 4 serotypes were identified, i.e. B (10%), B+A/C (18%), A/C (20%) and A/C+B (52%). The modern serotype HIV-1 diversity in RT resembles the epidemiologic situation in the territory of the former USSR as observed in the late 80-ies-early 90-ies of the last century. PMID- 15293510 TI - [Spectrum of cytokines produced by blood leukocytes in infection of herpes simplex virus, type 1]. AB - Three types of reaction of human blood leucocytes to herpes simplex virus 1 (HSV 1) were detected. The first reaction type, i.e. production of IFN-alpha, IFN gamma, IL-1beta, IL-6 and TNFalpha but not of IL-2 or IL-4, denotes the primary body reaction to an infectious agent. The second reaction type is related with infection of activated HSV-1 leucocytes and is accompanied by an inhibited production of IFN-gamma, IL-6 and IL-8, which is targeted at suppressing the antivirus cell mechanisms. The third reaction type is associated with production of IFN-alpha, IFN-gamma, IL-beta, IL-6 and TNFalpha by blood leucocytes affected by HSV-1-infected leucocytes. PMID- 15293511 TI - [Inactivation of viruses of different taxonomic groups by cuprous sulphate]. AB - Study results of inactivated effects exerted by cuprous sulphate on viruses of different taxonomy groups are summarized in the paper. Cuprous sulphate is a simple and reliable agent in inactivation of viruses of classical porcine fever, Aujeszky's disease and bovine infectious rhinotracheitis. Its inactivation action is based on the ability to reduce the viral genome to low-molecular fragment. Apart from inactivation of the virus material, a decreased level of protective antibody determinants is observed when cuprous sulphate is used in case of sheep catarrhal fever. PMID- 15293512 TI - [An enhanced viral safety of blood preparations]. AB - Inactivation or elimination of (possibly) contaminated viruses from a pool of prepared several hundreds or thousands of donor-blood samples are an obligatory stage in the donor-blood preparation process. Virus-inactivation is verified through contaminating the basic material with viruses. The quality control of blood preparations, according to the Russian compulsory regulations, must include the testing of ready blood-based drugs for a lack of antibodies to HIV, hepatitis C virus and hepatitis B virus by using the test systems, which could not be exactly designed for the above purpose. Therefore, the below tasks are vital for the Russian Blood Service: 1) cancellation of the norm (belonging to the regulations of the quality control of blood preparations) to test the blood preparations for a lack of antibodies to HIV, hepatitis C virus and to the surface antigen of hepatitis B virus because it is biologically inexpedient and has no analogues in the world practice; 2) introduction of the virus-inactivation methods into the practice of plasma processing; 3) establishment of a special center that would evaluate the efficiency of the virus-inactivation methods used by producers of blood-based preparations; and 4) introduction of the methods of genetic testing of HIV, hepatitis B and C viruses into monitoring the quality of donor-sera pools that are later used in preparations' manufacturing. PMID- 15293513 TI - Relevant bounds on hierarchical levels in the description of mechanisms. AB - Mechanisms pervade the sciences, and a significant portion of scientific research is concerned with their discovery and description. This paper is concerned with the latter, categorizing the components that should be included in the description of a mechanism. In describing mechanisms, the question of relevant level of description often arises; the question is where, if anywhere, one should halt a reductionist approach. In this paper, I propose a framework for mechanistic description that identifies the 'relevant' hierarchical levels of a mechanism based on their contributions to the functionality of the mechanism's endproduct. Mechanisms culminate in an endproduct, be it an activity or an entity, and that endproduct has qualities that allow it to participate in certain subsequent (or higher level) mechanisms. The proposed framework takes into account the level of description necessary to result in a functional endproduct; that is, an endproduct that exhibits the enabling properties required for it to fulfill its known roles in other mechanisms. Adhering to this framework results in a mechanistic description containing only the components that directly contribute to endproduct functionality. By constraining the hierarchical level of the description, the mechanistic sequence is clarified and made practical for many research applications where inadvertent focus on irrelevant details can prove costly. PMID- 15293514 TI - Cytology and mendelism: early connection between Michael F. Guyer's contribution. AB - This paper examines the contribution of the PhD dissertation of the American cytologist Michael F. Guyer (1874-1959) to the early establishment (in 1902-1903) of the parallel relationship between cytological chromosome behaviour in meiosis and Mendel's laws. Guyer's suggestions were among the first, which attempted to relate the variation observed in the offspring in hybridisation studies by a coherent cytological chromosome mechanism to meiosis before the rediscovery of Mendel's principles. This suggested for the first time that the chromosome mechanism involved a conjugation of maternal and paternal chromosomes during the synapsis followed by a segregation of parental chromosomes in the final germ cells and a random union of the final germ cells in the fertilization. It shows that this early suggestion was similar to William Austin Cannon's later chromosome proposal attempting to explain Mendel's principles and had some influence on Walter Sutton's cytological suggestion explaining correctly the behaviour of Mendel's particle by 1903. PMID- 15293515 TI - Evolutionary ethics from Darwin to Moore. AB - Evolutionary ethics has a long history, dating all the way back to Charles Darwin. Almost immediately after the publication of the Origin, an immense interest arose in the moral implications of Darwinism and whether the truth of Darwinism would undermine traditional ethics. Though the biological thesis was certainly exciting, nobody suspected that the impact of the Origin would be confined to the scientific arena. As one historian wrote, 'whether or not ancient populations of armadillos were transformed into the species that currently inhabit the new world was certainly a topic about which zoologists could disagree. But it was in discussing the broader implications of the theory...that tempers flared and statements were made which could transform what otherwise would have been a quiet scholarly meeting into a social scandal' (Farber 1994, 22). Some resistance to the biological thesis of Darwinism sprung from the thought that it was incompatible with traditional morality and, since one of them had to go, many thought that Darwinism should be rejected. However, some people did realize that a secular ethics was possible so, even if Darwinism did undermine traditional religious beliefs, it need not have any effects on moral thought. Before I begin my discussion of evolutionary ethics from Darwin to Moore, I would like to make some more general remarks about its development. There are three key events during this history of evolutionary ethics. First, Charles Darwin published On the Origin of the Species (Darwin 1859). Since one did not have a fully developed theory of evolution until 1859, there exists little work on evolutionary ethics until then. Shortly thereafter, Herbert Spencer (1898) penned the first systematic theory of evolutionary ethics, which was promptly attacked by T.H. Huxley (Huxley 1894). Second, at about the turn of the century, moral philosophers entered the fray and attempted to demonstrate logical errors in Spencer's work; such errors were alluded to but never fully brought to the fore by Huxley. These philosophers were the well known moralists from Cambridge: Henry Sidgwick (Sidgwick 1902, 1907) and G.E. Moore (Moore 1903), though their ideas hearkened back to David Hume (Hume 1960). These criticisms were so strong that the industry of evolutionary ethics was largely abandoned (though with some exceptions) for many years. Third, E.O. Wilson, a Harvard entomologist, published Sociobiology: The New Synthesis in 1975 (Wilson E.O. 1975), which sparked renewed interest in evolutionary ethics and offered new directions of investigation. These events suggest the following stages for the history of evolutionary ethics: development, criticism and abandonment, revival. In this paper, I shall focus on the first two stages, since those are the ones on which the philosophical merits have already been largely decided. The revival stage is still in progress and we shall eventually find out whether it was a success. PMID- 15293516 TI - Human cloning: mission futile, dangerous or impossible? PMID- 15293517 TI - Leptin and bone metabolism. AB - Leptin is a hormone involved with satiety and energy balance and proposed to be an anti-obesity factor. Much effort has been dedicated to the relationship between leptin and bone. This interest stems from the knowledge that body weight is a major determinant of bone density. It is known that obese persons have stronger bones and lose bone tissue at a slower pace. Therefore, attention has been given to leptin as a mediator of increased osteogenesis. Leptin has been shown to play a role on bone both in vitro and in vivo. The administration of leptin in vitro induced the expression of leptin receptors on stromal cells, the differentiation to osteoblasts and inhibition of differentiation into the adipocyte phenotype. In addition, leptin was able to inhibit osteoclastogenesis of peripheral blood mononuclear cells. Therefore, there is in vitro and experimental evidence that leptin is able both to stimulate osteoblasts and inhibit osteoclast differentiation. This would be in line with the hypothesis that the correlation between obesity and increased BMD is linked to leptin activity. However, experimental results are indicative of a role of CNS in mediating the effect of leptin on bone metabolism. These effects are opposite to the direct effects on bone cells and lead to bone loss. To solve the problem, it has been suggested that obese individuals have a resistance of nervous structures to leptin. In chronic renal failure serum leptin levels are markedly increased. An inverse correlation between histomorphometric parameters of bone turnover and serum leptin levels and between leptin and PTH have been reported. Therefore, the hypothesis has been raised that leptin lowers bone turnover in chronic renal failure. Since leptin has a direct stimulatory effect on bone and an indirect opposite effect via the CNS, it has been suggested that in CRF a resistance of nervous structures to leptin, like in obesity, may be present. By now, coherent findings suggest that the prevailing effect of leptin on bone in ESRD is that of reducing bone turnover. PMID- 15293518 TI - The fate of bone after renal transplantation. AB - Post-transplantation bone disease is a multifactorial, complex condition. It derives in a significant part from pre-existing renal osteodystrophy, but it is aggravated by factors emerging after renal transplantation. Among the latter factors, the key pathophysiological contributor to bone disease is immunosuppressive agent application (especially glucocorticoids (GC)). Post transplantation bone disease is detectable even years after renal transplantation in the vast majority of patients, and potentially it never resolves completely. Due to post-transplantation bone disease, a rapid reduction of bone mineral density (BMD) develops that can exceed 10% in the first 12 months. Subsequently, the bone loss slows down or even a secondary increase occurs. Post transplantation bone disease results in a significantly elevated fracture risk, which largely contributes to the increased morbidity in transplant patients. Currently, vitamin D metabolites and bisphosphonates are the most extensively tested therapeutic agents against this accelerated bone loss. Both substances have proven effective. However, it is yet unproven that they reduce the fracture risk. In patients with adynamic bone disease, bisphosphonate usage cannot be recommended, since this group of drugs could oversuppress bone metabolism. PMID- 15293519 TI - Determinants of coronary vascular calcification in patients with chronic kidney disease and end-stage renal disease: a systematic review. AB - BACKGROUND: Vascular calcification (VC) is a recognized process involved in senescence and atherosclerosis. Chronic kidney disease (CKD) and end-stage renal disease (ESRD) are conditions associated with metabolic disorders related to soft tissue calcification. METHODS: We performed a systematic review of the literature confined to patients with CKD or ESRD with clinical observations of VC. Case reports of calciphylaxis were excluded. We identified 30 studies over 20 years: 11 prospective cohort, 7 cross-sectional, 11 case-control, and 1 retrospective cohort; n = 2918 subjects, mean age 51 years, 59% men and 41% women. Imaging methods used included: x-ray 43%, computed tomography 30%, ultrasound 17%, and other methods 10%. RESULTS: The most consistent determinants of VC were older age and dialysis vintage. Eight analyses determined a relationship between VC and measures of calcium-phosphate balance while 20 analyses specifically did not find such a relationship. Three studies suggested the degree of calcium loading, treatment with phosphate binders, or treatment with vitamin D analogues were related to VC. When taken into consideration, the lipid profile (primarily low high-density lipoprotein cholesterol, elevated triglycerides, elevated low density lipoprotein, and elevated total cholesterol) were predictive factors in four analyses. CONCLUSIONS: VC is a common observation in CKD and ESRD and is mainly related to age, length of time on dialysis therapy, and possibly dyslipidemia. The calcium-phosphorus balance and its related treatments are likely not related to this unique form of vascular calcification. Further research into the determinants and potential treatments for vascular calcification is warranted. PMID- 15293520 TI - A primer on survival analysis. AB - Survival analysis is the statistical method for studying the time between entry to a study and a subsequent event. Survival is often the most important outcome in both observational and intervention studies. The analysis of survival, however, is not simple because of a number of factors. The time between entry to a study and the end of follow-up varies during the study period, as recruitment time for each subject is different. In addition, most studies have a finite duration during which not all subjects would have experienced the outcome of interest, while some patients might leave the study or are lost to follow-up. Survival analysis is not just concerned with time from treatment to death. The outcome could be any defined event such as time to doubling of serum creatinine concentration or first rejection episode following transplantation. An understanding of the principles involved in the statistical analyses of survival is essential for critical appraisal of studies, which employ these methods, and for designing clinical studies. This brief review describes the principles of survival analysis and its application to studies in nephrology. PMID- 15293521 TI - Access site-related infection in dialysis: the AStRID project: a multicenter prospective Italian study. AB - A multicenter prospective study has been planned, in a large sample of Italian end-stage renal disease (ESRD) patients, aiming to assess the vascular access (VA) site-related infection rates and to identify variables associated with them. All ESRD patients undergoing chronic hemodialysis (HD) in the participating centers will be enrolled in the study. Participating centers were selected on a voluntary basis. Patients will be enrolled within an 18-month recruitment period. Primary study end points are the overall incidence rate of VA-related infections in ESRD patients on chronic HD (defined as infection episodes/100 patient months), and the incidence rate of different types of VA-related infections (exit site, tunnel and bacteraemia/sepsis). All VA types in use will be evaluated: fistula, graft, tunneled (permanent) central venous catheter (CVC) and temporary CVC. PMID- 15293522 TI - Infective endocarditis in maintenance hemodialysis patients: fifteen years' experience in one medical center. AB - BACKGROUND: Infective endocarditis (IE) is a serious infectious condition, with high morbidity and mortality in hemodialysis (HD) patients. This study was undertaken to determine the IE risk factors in maintenance HD patients, and the mortality risk factors. METHODS: We retrospectively reviewed all IE cases of maintenance HD patients at our center over the past 15 yrs (the study group). Regular HD patients without IE in the same period were used as the control group. The basic data of the two groups were analyzed to determine IE risk factors in HD patients. The in-hospital parameters of survival and mortality in the study group patients were used for mortality risk factors analysis. RESULTS: There were 18 definite, and two possible, IE diagnoses in the study group and no cases in the 268 controls. There was no significant difference in age, sex, diabetes, hypertension, underlying malignancy, previous cerebral vascular accident (CVA) history, and calcium multiplied by phosphate product. There was a significant difference between the two groups (study group vs. controls) in pacemaker implant history (15 vs. 1.1%, p<0.01), previous heart surgery history (15 vs. 0.4%, p<0.01), congestive heart failure (CHF) (50 vs. 10.4%, p<0.05), duration on maintenance HD (12.9+/-19.1 vs. 57.9+/-42.3 months, p<0.001), serum albumin at the time of admission (2.91+/-0.40 vs. 3.96+/-0.52 g/dL, p<0.001). There were more patients dialyzed via non-cuffed dual-lumen catheters in the study group (55 vs. 0%, p<0.001), and fewer patients dialyzed via arteriovenous fistula (AVF) (25 vs. 87.7%, p<0.001). The mortality in HD patients with IE was high (60%), especially in patients with methicillin-resistant Staphylococcus aureus (MRSA) endocarditis (100%). The most common pathogen was S. aureus (n=12). MRSA was more common than methicillin-susceptible S. aureus (MSSA) (67 vs. 33%). Univariant analysis of in-hospital clinical parameters for mortality revealed no significant difference in age, diabetes, dual-lumen catheter implantation, serum albumin, time to diagnosis, and time to antibiotic use. Borderline statistical significance was noted in serum C-reactive protein (CRP) (p=0.051), and blood glucose level (p=0.056). There were more IE cases due to MRSA in the mortality group than in the survival group (8 vs. 0 cases, p=0.013), but fewer cases due to MSSA (0 vs. 4 cases, p=0.050). CONCLUSIONS: IE should be considered in HD patients with the following risk factors, which include previous heart surgery or pacemaker implantation, shorter HD duration, and especially for patients dialyzed via dual-lumen catheters. The in-hospital clinical parameters including CRP and blood sugar level can offer information concerning prognosis. Since MRSA has increased in recent years and is associated with high mortality, strategies for prevention and treatment require development. PMID- 15293523 TI - Bacterial meningitis in hemodialyzed patients. AB - BACKGROUND: To analyze the clinical features, causative pathogens and therapeutic outcomes of bacterial meningitis in hemodialyzed patients. PATIENTS AND METHODS: Two hundred and sixty-seven patients, > or = 16 yrs, were identified with culture proven bacterial meningitis. In addition, the causative pathogens and therapeutic outcomes between uremic and non-uremic patients with adult bacterial meningitis were analyzed. RESULTS: Nine uremic patients with bacterial meningitis, accounting for 3% (9/267) of our adult patients with culture-proven bacterial meningitis had fever, disturbed consciousness and seizures. These were the three most common manifestations in our patients. The interval between the onset of symptoms and therapy start was 5-11 days (mean: 9 days). No patients were initially diagnosed with bacterial meningitis, two patients were initially suspected of having infection of unknown origin. In the non-uremic patient group, klebsiella pneumoniae, Staphylococcus aureus, and Streptococcus pneumoniae were the most frequently detected pathogens, while pseudomonas aeruginosa and coagulase-negative Staphylococcus were the most prevalent in the uremic patients group. The overall mortality rates for the non-uremic and uremic patient groups were 33 and 78% respectively. CONCLUSION: The mortality rate for bacterial meningitis in the uremic patients group remained high. Due to non-specific manifestations and slow evolution, bacterial meningitis was commonly misdiagnosed as uremic encephalopathy. Therefore, effective treatment was usually delayed. To avoid treatment failure, early diagnosis, careful monitoring of clinical condition and appropriate antibiotic choices are necessary. PMID- 15293524 TI - Decrease in infections with the introduction of mupirocin cream at the peritoneal dialysis catheter exit site. AB - Staphylococcus aureus associated peritonitis and catheter exit site infections (ESI) are an important cause of hospitalization and catheter loss in the patients undergoing chronic peritoneal dialysis (PD). We aimed to determine the potential effectiveness of the application of mupirocin cream at the catheter exit site in preventing exit site infection and peritonitis. METHODS: This prospective historically controlled study was done in a total of 86 patients who entered our PD program from April 1999 to January 2001. They were instructed to apply Mupirocin cream 2% to the exit site daily or on alternate days. The patients were not screened to determine whether they were staphylococcus aureus carriers. One hundred and thirteen patients on PD prior to April 1999 acted as historical controls. Both groups were followed prospectively for a period of 22 months. RESULTS: In the study group application of mupirocin lead to a significant reduction in the incidence rate of both exit site infections overall (0.43 vs. 0.09; p<0.0001) and ESI due staphylococcus aureus (0.14 vs. 0.02; p=0.004) amounting to a relative reduction of 79% and 85% respectively. Although the overall incidence of peritonitis did not change (0.28 vs. 0.26; p=0.7) there was a significant reduction in peritonitis caused by staphylococcus aureus (0.07 vs. 0; p=0.01) Although only one catheter required removal in the mupirocin group as against 5 in the control group, this was not statistically significant. CONCLUSIONS: Mupirocin application at the exit site significantly lowers the incidence of ESI and peritonitis caused by staphylococcus aureus without any significant side effects. PMID- 15293525 TI - Total lymphocyte count in peripheral blood of peritoneal dialysis patients: relationship to clinical parameters and outcome. AB - BACKGROUND: Total lymphocyte count (TLC) is used as a nutritional status measurement. The impact of TLC on mortality in peritoneal dialysis (PD) patients is controversial. This study aimed at evaluating the effect of TLC on mortality, and assessing the relationship between TLC and nutritional status, anemia and erythropoietin (EPO) response, acute-phase response, dialysis adequacy and volume status in PD patients. METHODS: Seventy-three PD patients were monitored for 3 yrs from the beginning of the treatment. Data recorded for each patient included demographic features, comorbidity, TLC, blood biochemistry, systolic and diastolic blood pressures (BP), indices of dialysis adequacy and nutritional status, total fluid removal and mortality. Adjusted mortality risk for TLC was estimated using the Cox's regression models composed by TLC and one covariate having a value p<0.05 in univariate analysis. RESULTS: The 3-yr patient survival rates were significantly different among the TLC quartiles. The adjusted TLC was found, generally, to be a significant predictor of death in reduced Cox's models, except in models composed of TLC and total fluid removal or serum albumin. The receiver operating characteristics (ROC) analysis demonstrated that TLC provided a significant prediction of mortality. TLC correlated positively to total fluid removal, serum albumin, triglyceride and hematocrit, and negatively correlated to BP, high peritoneal transport and EPO-need. It did not correlate to other measures of nutritional status, dialysis adequacy and acute-phase response. Fluid removal and serum triglyceride were independent predictors of TLC in multivariate analysis. CONCLUSIONS: Our findings suggest that TLC can be used as a simple prognostic tool in PD patients; however, the association between TLC and mortality is confounded by other prognostic factors. Volume status could be a more important factor affecting the TLC than nutritional status. PMID- 15293526 TI - Leptin and biochemical markers of bone turnover in dialysis patients. AB - BACKGROUND AND OBJECTIVE: The adipose tissue cytokine leptin is suggested to interfere with bone turnover mechanisms because, in rats with leptin deficiency, intra-cerebroventricular administration of this cytokine causes a reduction in bone mass. We studied the relationship between plasma leptin and biochemical bone turnover indicators in 161 hemodialysis (HD) patients. RESULTS: Plasma leptin was sex-dependent, being significantly higher (p<0.001 ) in female dialysis patients than in male dialysis patients, and it related directly to body mass index (BMI). In males, plasma leptin related inversely to serum intact parathyroid (PTH) (partial r= -0.34), serum PTH(1-84) (r= -0.36), carboxyterminal PTH (C-PTH) fragment (r= -0.31) and serum PTH(1-84)/C-PTH fragment ratio (r= -0.22), while no such relationships were found in females. Of 93 male dialysis patients, 44 had a serum intact PTH <100 pg/mL and 14 had a serum PTH(1-84)/C-PTH fragment ratio <1. In a multiple logistic regression analysis in males, for each 1 ng/mL increase in plasma leptin there was an 11% excess risk of serum intact PTH <100 pg/mL (odds ratio (OR) 1.11, 95% confidence interval (95% CI): 1.02-1.20, p=0.01) and a similar OR was found when low bone turnover was defined based on a serum PTH(1 84)/C-PTH fragment ratio <1 (p=0.01). In addition, plasma leptin related inversely to skeletal alkaline phosphatase and again this relationship was found in male but not in female dialysis patients. CONCLUSIONS: Our data support the theory that leptin reduces bone turnover in male dialysis patients. Whether this link underlies a noxious or a protective mechanism, i.e. if it can serve to limit high bone turnover due to hyperparathyroidism, remains to be established in prospective studies based on solid outcome measures like the risk of fractures. PMID- 15293527 TI - Efficacy and tolerability of manidipine in the treatment of hypertension in patients with non-diabetic chronic kidney disease without glomerular disease. Prospective, randomized, double-blind study of parallel groups in comparison with enalapril. AB - BACKGROUND: Calcium channel blockers (CCBs) are effective blood pressure lowering agents, giving rise to a prevalent dilation of the afferent arteriole. Manidipine, a long-lasting dihydropyridine CCB, demonstrates its action not only on the afferent arteriole, but also on the efferent one. This suggests theoretically a renoprotective effect in patients with chronic kidney diseases (CKD). METHODS: This was a multicenter, prospective, randomized, double-blind, parallel group study, to evaluate the efficacy and tolerability of manidipine (M; 10-20 mg/day), in comparison with enalapril (E; 10-20 mg/day) in the treatment of hypertension in 136 patients with CKD secondary to primary renoparenchymal disease. Changes in blood pressure values from baseline were considered as the primary outcome of the study. Proteinuria changes and the rate of renal function decline were also evaluated. RESULTS: During a 48-week follow-up, mean SBP decreased from 155+/-11.7 to 138.7+/-13.9 mmHg in M and from 157.3 +/-11.8 to 134.2+/-13.9 mmHg in E; mean DBP decreased from 100.3+/-4.2 to 86.1+/-6.5 mmHg in M and from 100.3+/-4.2 to 84.7+/-6.3 mmHg in E. Proteinuria remained unchanged in M (from 1.6+/-1.59 to 1.62+/-1.79 g/24h), and decreased significantly in E (from 1.37+/-1.45 g/24h to 1+/-1.55 g/24h). No significant difference was observed in the rate of renal function decline in the two groups. CONCLUSIONS: Manidipine was safe and effective, obtaining a significant reduction in SBP and DBP from baseline. Although patients treated with enalapril showed a better antiproteinuric response, the two treatments were equally effective in reducing the rate of CRF progression in patients without glomerular disease. PMID- 15293528 TI - Cardiac cyclic variation of integrated backscatter in hypertension and dialysis patients. AB - BACKGROUND: Cyclic variation of myocardial-integrated backscatter (CV-IB) offers a non-invasive myocardial contractile performance assessment. There is limited data concerning CV-IB in end-stage renal disease (ESRD) patients. METHODS: Forty essential hypertensive (EH) patients (mean age 51+/-8 yrs) and 24 ESRD patients (mean age 49+/-14 yrs) were compared to 10 healthy controls (mean age 45+/-10 yrs). A 2D-Doppler echocardiography with digitized imaging was performed to characterize myocardial ultrasonic tissue by CV-IB between systole and diastole at the interventricular septum (IVS) and left ventricular (LV) posterior wall (PW). RESULTS: There was no significant difference between age and sex among groups. Systolic and diastolic blood pressures (BP) were both higher in EH patients (157/96 mmHg in EH, 129/81 mmHg in ESRD and 115/77 mmHg in controls, p<0.001). Left ventricular mass index (LVMI) was higher in EH and ESRD patients than in controls (respectively, 119+/-37, 130+/-46, 87+/-12 g/m2, p<0.05), while there was no significant difference found between EH and ESRD patients. EH patient CV-IB values were significantly lower than in ESRD patients and controls (respectively, 6.9+/-1.6, 8.6+/-0.7, 10.6+/-1.1 dB, p<0.001 for IVS, 7.7+/-1.3, 8.7+/-0.8, 10.4+/-1.1 dB, p<0.001 for PW). CV-IB for PW and IVS were significantly lower in ESRD patients than in controls (p<0.001). CONCLUSIONS: CV IB can offer useful parameters for myocardial structure in EH and ESRD patients. Further studies are needed to clarify CV-IB in ESRD patients. PMID- 15293529 TI - Referral of type 1 diabetic patients to a nephrology unit: will pre-emptive transplantation change our life? AB - BACKGROUND: Type 1 diabetic patients are a small but challenging subset of chronic kidney disease. The new frontiers of pancreas-kidney transplantation may enhance the need for early referral. OBJECTIVE: To analyze the referral pattern of type 1 diabetics to a specialized Nephrology Unit, and to quantify the indications for pancreas or pre-emptive pancreas-kidney transplantation at referral in this population. PATIENTS AND METHODS: Setting of study was a Nephrology Outpatient Unit, dedicated to diabetics, active since 1986; period of study 1991--2002. The main biochemical and clinical parameters were analyzed at referral. Indications for transplantation were put at: serum creatinine (sCr)> or =2 mg/dL or > or =3 mg/dL and/or nephrotic syndrome. Pancreas: lesser degrees of functional impairment without worsening after FK-506 challenge. RESULTS: 90 type 1 diabetics were referred: 48 males, 42 females; median age: 38 (18-65) years; median diabetological follow-up 20 (3-37) years; sCr 1.2 (0.6-7) mg/dL, proteinuria 0.9 (0-12.3) g/day; creatinine clearance: 58 (6-234) ml/min; Hbalc: 8.8% (5.9-14), diastolic blood pressure: 80 (55-100) mmHg, systolic blood pressure: 137.5 (70-180) mmHg. 85.6% had signs of end-organ damage due to diabetes. 67% of the patients had diabetic nephropathy, 20.7% hypertensive with or without diabetic nephropathy. According to the chosen criteria, 30.6% had indications for pancreas-kidney graft (sCr > or = 2 mg/dL), 25.9% considering sCr > or = 3 mg/dL; 28.2% further patients could be considered for isolated pancreas graft. CONCLUSIONS: At referral to the nephrologist, over 50% of type 1 diabetics may have indications for pancreas-kidney or pancreas graft; an earlier multidisciplinary work-up is needed to optimize an early pre-emptive transplant approach. PMID- 15293530 TI - Neoral dose adjustment after conversion from C0 to C2 monitoring in stable renal transplant recipients: a prospective single center study. AB - To determine the clinical impact of conversion from C0 to C2 Neoral monitoring, we conducted a 6-month prospective study in 62 stable renal transplant recipients. Neoral was given alone (19%), with steroids (31%), combined with azathioprine (Aza) or mycophenolate mofetil (MMF) (50%). C0 and C2 target ranges were, respectively, 130-190 and 700-900 ng/mL. Neoral dosages were adjusted according to the C2 range. At baseline, mean C0 and C2 were 157 and 762 ng/mL. After 6 months C0 was 173 ng/mL (p<0.02) and C2 was 804 ng/mL (ns). Although the mean Neoral dose at 6 months was unchanged from baseline, the dose was reduced in 24 patients from 3.6+/-1.2 to 3.0+/-0.9 mg/kg/day, with a mean reduction in serum creatinine (Cr) from 1.4+/-0.4 to 1.3+/-0.3 mg/dL (p<0.001), stable in 8 patients and increased in 30 patients from 3.3+/-1.0 to 3.8+/-1.2 mg/kg/day with no change in serum Cr. Serum transaminases and blood pressure (BP) were unchanged in the three groups. C0 and C2 showed a positive correlation, but with a large dispersion of values (r2=0.14, p<0.001). Overall concordance between the C0 and C2 ranges was 49%. Therefore, in stable transplant patients C0 cannot be considered a C2 surrogate. The conversion from C0 to C2 led to a Neoral dose reduction in approximately 40% of patients with significant improvement in renal function. Most of the remaining patients required an increased dose; however, without an increased incidence of cyclosporin-induced side-effects. PMID- 15293531 TI - Erectile dysfunction in kidney transplanted patients: efficacy of sildenafil. AB - BACKGROUND: Erectile dysfunction (ED) is the inability to achieve and/or maintain an erection for satisfactory sexual performance. The effects of kidney transplantation on pre-existing ED are poorly understood, as well as the onset of new ED cases after kidney transplantation. This study aimed to evaluate the effects of kidney transplantation on pre-existing ED, to assess the onset of new ED cases after renal transplantation and to assess both the efficacy and safety of sildenafil. METHODS: Erectile function was assessed using the self administered International Index of Erectile Function (IIEF) to kidney transplanted patients. A 50 mg dose of sildenafil was prescribed. Sildenafil efficacy was assessed by re-administering the questionnaire after 4 weeks of therapy. Blood chemistry and serum cyclosporine concentration were evaluated at the beginning of the study and, in patients treated with sildenafil, after 4 weeks of treatment. RESULTS: One hundred and thirteen patients (87.5%) completed the questionnaire. Fifty-three patients (47%) did not complain of ED, the remaining 60 patients (53%) reported ED. ED was already present during dialysis in 40 patients; it appeared ex novo in 20 patients after transplantation. Pre existing ED disappeared in 8 patients (20%), ameliorated in 13 patients (32.5%), worsened in 2 patients (5%), and remained unchanged in 17 patients (42.5%) after transplantation. The IIEF score significantly improved in sildenafil-treated patients (n=20); there were no observed changes in blood chemistry, blood pressure (BP), renal function and cyclosporine concentration. The side-effects were minimal. CONCLUSIONS: ED was still present in a large cohort of kidney transplanted men. Renal transplantation cures few ED cases. ED can appear ex novo after transplantation. Sildenafil is an effective ED treatment in kidney transplanted men. PMID- 15293532 TI - Leishmaniasis in patients with chronic renal failure: a diagnostic and therapeutic challenge for the clinician. AB - BACKGROUND: The role of leishmaniasis in dialyzed or transplanted patients for chronic renal failure is generally neglected. In this study, the authors present a series of three cases of leishmaniasis (one visceral, one mucous and one muco visceral) in patients with end-stage renal failure characterized by an atypical presentation and/or resistance to therapy. CASE DESCRIPTION: Two patients had an atypical infection: the first patient demonstrated a mucosal form, while the second had visceral and mucosal involvement. These two presentations are very rare and, to the best of our knowledge, other autoctonous disease cases have never been described in Italy. In the first patient, a cycle of oral itraconazole was scarcely effective and poorly tolerated, while treatment with 15% topical paromomycin sulfate was successful. Patients two and three failed to respond to meglumine antimonate and amphotericin B lipid complex. A second cycle with liposomal amphotericin B was effective in both cases. In addition, a superior safety profile for liposomal amphotericin B in comparison with the lipid complex amphotericin B was observed. CONCLUSIONS: These three cases highlight the problem of leishmaniasis in both renal transplanted and dialyzed patients and suggest that this infection could be far from infrequent in addition to being resistant to therapies. Leishmaniasis should be considered in the differential diagnosis of fevers of unknown origin and mucosal lesions in these patients, even in countries not at risk for mucosal leishmaniasis. PMID- 15293533 TI - End-stage renal disease in leprosy. AB - BACKGROUND: Leprosy or Hansen's disease (HAD) undoubtedly remains an emergency in certain countries. It is an ancient deforming disease caused by Mycobacterium leprae. The countries with the highest endemic leprosy rate in 2000 were Brazil, India and Madagascar. In Italy, the old epidemic has been defeated and there are approximately 400 patients under constant monitoring with three to four new cases per year involving Italian residents. The kidney is one of the target organs during the splanchnic localization of leprosy. The histopathological renal lesion spectrum includes glomerulonephritis (GN), renal amyloidosis (RA) and interstitial nephritis (IN). Both proteinuria and chronic renal failure are the main clinical expressions of renal damage in leprosy. To the best of our knowledge, very little is reported concerning end-stage renal disease (ESRD) in leprosy patients both in the most important national and international renal registries and in the available literature. This study aimed to report the long term experience of our department in this field. METHODS: To achieve this, we analyzed retrospectively the HAD Center (Gioia del Colle) database at ourhospital. RESULTS: Eight leprosy patients were dialyzed from 1980 to June 2003 (six males and two females), with a mean age of 61.0+/-8.9 SD yrs (range: 51-76) and a mean HAD duration of 36.1+/-5.1 yrs. The first clinical nephropathymanifestations were non-nephrotic proteinuria associated with chronic renal failure in four patients, and nephrotic proteinuria in four patients. Kidney biopsies performed in three patients showed two had RA, and one had IN. Two patients were treated initially by peritoneal dialysis; they were then switched to hemodialysis (HD) after 3 and 10 months because of recurrent peritonitis. HD treatment lasted 40.6+/-31.4 months (range: 9-101). Six patients died, one due to hyperkalemia, one because of a technical dialysis accident, and the remainder due to causes unrelated to the dialysis treatment. Two patients are still alive, treated with HD for 17 and 44 months. CONCLUSIONS: Uremia represents a late complication of leprosy and has a multifactorial genesis, although RA is among the most frequent causes, conventional bicarbonate HD appears to offer good results in the treatment of uremia in leprosy patients. PMID- 15293534 TI - Juvenile renal cell carcinoma as first manifestation of von Hippel-Lindau disease. AB - Von Hippel-Lindau (VHL) disease is an autosomal dominant syndrome characterized by germline mutations in the VHL tumor suppressor gene located at chromosome 3p25 26 and pleomorphic clinical picture. The major clinical manifestations include retinal angiomas, central nervous system hemangioblastomas, pheopleochromocytoma, pancreatic cysts, epididymal cystoadenomas and renal lesions. Recently, we observed a 58-year-old male patient with macrohematuria and a history of nephrectomy due to renal cell carcinoma (RCC). The patient showed retinal angiomatosis, cerebellar hemangioblastomas, multiple pancreatic cysts, right kidney with polycystic features plus two RCC. The patient's offspring, two females and one male, showed VHL lesions, such as retinal angiomatosis, cerebellar hemangioblastomas and polycystic kidney disease (PKD). The affected family members were screened for mutations in the VHL gene. Data suggested the presence of a deletion encompassing exon 1 of the VHL gene. Early diagnosis of VHL disease in patients and their relatives is important for clinical and geneticreasons. VHL disease patients have an increased incidence of malignant carcinomas and the syndrome can mimic the presentation of other cystic kidney diseases. Early diagnosis and molecular genetic testing of family members is essential to improve the clinical management of patients and to allow an accurate risk assessment in asymptomatic individuals. In conclusion, nephrologists and urologists must carefully evaluate patients with PKD and RCC to confirm or exclude VHL disease, and physicians must play a crucial role in the clinical process of therapeutical decisions and choices for VHL patients. PMID- 15293535 TI - Bilateral renal infarctions and lower limbs artery thrombosis in a patient with nephrotic syndrome. AB - Thromboembolism is a well-recognized complication in patients with nephrotic syndrome owing to their hypercoagulable status. Usually, the venous system is affected, whereas the very rare occurrence of arterial thrombosis is mainly restricted to pediatric patients. This complication often results in high rates of mortality and limb loss. We report the case of an adult female patient with histologically diagnosed minimal change disease and nephrotic syndrome associated with malignant thymoma, who eventually developed concurrent bilateral kidney and lower limb thrombosis. Conservative systemic anticoagulation was administered and she recovered ample kidney function. In addition, although she underwent an emergent thrombectomy of the left popliteal artery, an amputation was necessary. To the best of our knowledge, this is the first reported arterial thrombosis case involving bilateral kidney and lower limb simultaneously in nephrotic patients. Our experience indicates that arterial thrombosis is a serious complication in nephrotic patients, and early detection and aggressive management are crucial in these patients to improve their outcome. PMID- 15293536 TI - History of home hemodialysis. PMID- 15293537 TI - Galeno's De Pulsis et Urinis from the Casinensis Codex 97. AB - Between the end of the 9th and the first decade of the 10th century, two medical codes were produced in the Cassino area: The Pluteo 73.41 manuscript of the Laurenziana library in Florence and the Casinensis Codex 97 kept in the archives of the abbey. They are written in Beneventano-Cassinese language and are enriched with drawings of plants and animals. They were most certainly used for the teaching and learning of medicine. On pages 26-31, there is Galeno's De Pulsis et Urinis, that is presented in the form of "Epistola", containing 30 chapters in which the characteristics and variations of pulse, fever and urine are described in different illnesses, thus influencing the prognosis. Galeno's manuscript De Pulsis et Urinis, as well as his other manuscripts contained in Code 97, have an important role in emphasizing the writings of one of the greatest and prolific doctors of ancient times who laid the basis for medicine in the East and in the West. The source is most likely Sergio's which belong to the medical school of Costantinople (his death is known to be in 536); he translated all of Galeno's known writings in the 6th century. PMID- 15293538 TI - Kidney and urinary therapeutics in early medieval monastic medicine. AB - This study will explore a few recipes in relation to what monastic medicine considered kidney disorders. Technical terms, such as strangury, cause us difficulties in interpreting early medieval monastic medicine. The action described can vary considerably. For example, he has action for urination problems that center in the verb: (in translation) "dries out", "quietens", "expels", "moves", "provoking", and treating "retention of urine". Sometimes the term "diuretic" is used. The Lorsch monastic book of medical recipes, written around 800, employed the words "deuritica" and "diureticon" but most of monastic accounts simply say something similar to that in Reichenau Monastery's recipe book, 9th or 10th century: "urinam movet/moves the urine"; "urinam provocat/stimulates urination"; "ad difficultate urine/difficulty in urination". In order to examine the relationship between diagnosis and therapy, let us turn to a disease that is difficult even for modern medicine, diabetes. An examination of several early medieval monastic accounts reveals that they could have effectively treated diabetes. PMID- 15293539 TI - Pietro Anzolino da Eboli and the thermal therapy of renal pathologies. AB - In the Campi Flegrei area of Campania, a region of Southern Italy, some thermal baths still exist, known for the therapeutical properties of their waters. Utilized since remote ages for the treatment of a variety of pathologies. In "De Balneis Terrae Laboris", an Italian poet of the Middle Ages and scientist, Pietro Anzolino da Eboli, reports every detail about more than thirty different thermal sites in the Campi Flegrei, including all therapeutic effects of their waters. Based on Pietro's manuscript, we report in the current paper all the sites whose waters were - among other indications - also recommended for the treatment of some urinary diseases. PMID- 15293540 TI - Monastic nephrology in the School of Salerno and in an unpublished treatise in middlelatin and Italian volgare of the manuscript 2Qq C63 in the public library of Palermo. AB - The relevance of the Salerno Medical School is examined through the contributions of Magister Maurus, Alfano I, Giovanni Plateario, Giovanni Afflacio, Bartolomeo and Urso of Calabria. An unpublished treatise on De Urinis in middlelatin and Italian volgare is introduced here for the first time by discussing the lapis of the ancient mediaeval urologic and uroscopic culture. PMID- 15293541 TI - Licorice from antiquity to the end of the 19th century: applications in medical therapy. PMID- 15293542 TI - The diuretic use of Scilla from Dioscorides to the end of the 18th century. AB - Ancient texts contain an extremely large and readily accessible body of information on traditional medicine describing a range of plants and other substances that have been recently investigated systematically. However, prospecting for drugs from herbals raises problems with philology and plant identification. We combined our expertise to re-examine Squill an ancient medicinal plant which deserves modern scientific investigation. For this, invaluable help has come from new computer technologies which allow access to the most important libraries of the History of Medicine. PMID- 15293543 TI - 2-D graphical representation of proteins based on virtual genetic code. AB - We consider a novel 2-D graphical representation of proteins in which individual nucleic acids are represented as "spots" within a square frame distributed according to specific construction rules. The resulting "images" of proteins can not only serve to facilitate visual comparison of similarities and dissimilarities between lengthy protein sequences, but also offer a way for mathematical characterization of protein sequences, analogous to similar considerations for lengthy DNA sequences. Basically the approach is based on the concept of virtual genetic code, which is a hypothetical string of RNA nucleic acid bases, A, C, U and G, which generates reported protein sequences, without the knowledge of the actual genetic code that produces the protein. PMID- 15293544 TI - Molecular structural characteristics governing biocatalytic chlorination of PAHs by chloroperoxidase from Caldariomyces fumago. AB - Based on some fundamental quantum chemical descriptors computed by PM3 Hamiltonian, a quantitative structure-property relationship (QSPR) for specific activity of 17 polycyclic aromatic hydrocarbons (PAHs) of biocatalytic chlorination by chloroperoxidase (CPO) from Caldariomyces fumago was developed using partial least squares (PLS) regression. The model can be used to estimate biocatalytic chlorination reaction rates of PAHs. The main factors affecting specific activity of PAHs of biocatalytic chlorination by CPO from Caldariomyces fumago are absolute hardness, dipole moment, absolute electronegativity, and molecular bulkness of the PAH molecules. The biocatalytic chlorination reaction rates of PAHs with large values of absolute hardness, absolute electronegativity, and molecular bulkness tend to be slow. Increasing dipole moment of PAHs leads to increase the specific activity. PMID- 15293545 TI - QSARS for toxicity to the bacterium Sinorhizobium meliloti. AB - In the present study, structure-activity relationship (QSAR) models for the prediction of the toxicity to the bacterium Sinorhizobium meliloti have been developed, based on a data set of 140 compounds. The data set is highly heterogeneous both in terms of chemistry and mechanisms of toxic action. For deriving QSARs, chemicals were divided into groups according to mechanism of action and chemical structure. The QSARs derived are considered to be of moderate statistical quality. A baseline effect (relationship between the toxicity and logP), which can be related to non-polar narcosis, was observed. To explain toxicity greater than the baseline toxicity, other structural descriptors were used. The development of models for non-polar and polar narcosis had some success. It appeared that the toxicity of compounds acting by more specific mechanisms of toxic action is difficult to predict. A global QSAR was also developed, which had square of the correlation coefficient r2 = 0.53. A QSAR with reasonable statistical parameters was developed for the aliphatic compounds in the data set (r2 = 0.83). QSARs could not be obtained for the aromatic compounds as a group. PMID- 15293546 TI - Highly compact 2D graphical representation of DNA sequences. AB - Most 2D graphical representations of primary DNA sequences, while offering visual geometrical patterns for depicting sequences, do require considerable space if enough details of such representations are to be visible. In this contribution, we consider a highly compact graphical representation of DNA, which allows visual inspection and numerical characterization of DNA sequences having a large number of nucleic acid bases. The approach is illustrated on the DNA sequences of the first exon of human beta-globin. The same graphical approach not only allows one to depict differences in composition within a single DNA, but makes possible graphical representation of protein sequences, which have hitherto evaded similar 2D visual representations. PMID- 15293547 TI - QSAR-based toxicity classification and prediction for single and mixed aromatic compounds. AB - Quantitative structure-activity relationships (QSARs) based on the octanol/water partition coefficient were employed to predict acute toxicities of 36 substituted aromatic compounds and their mixtures. In this study, the model developed by Verhaar et al. was modified and used to calculate octano/water partition coefficients of chemical mixtures. To validate the model, acute toxicities of these chemicals were measured to Vibrio fischeri in terms of EC50. The results indicated that the obtained QSAR models could be used to predict toxicities of samples consi sting of these substituted aromatic compounds, individually or in combinations. The obtained equations were proved to be robust enough by using the leave-one-out test method. By classifying these chemicals into two groups, polar and non-polar, the toxicities of chemical mixtures within each group can be predicted accurately from their calculated partition coefficients. PMID- 15293548 TI - CISOC-PSCT: a predictive system for carcinogenic toxicity. AB - A SAR based carcinogenic toxicity prediction system, CISOC-PSCT, was developed. It consisted of two principal phases: the construction of relationships between structural descriptors and carcinogenic toxicity indices, and prediction of the toxicity from the SAR model. The training set included 2738 carcinogenic and 4130 non-carcinogenic compounds. Three predefined topological types of substructures termed Star, Path and Ring were used to generate the descriptors for each structure in the training set. In this system, the defined carcinogenic toxicity index (CTI) was obtained from the probability of a structural descriptor to either belong to the carcinogenic or non-carcinogenic compounds. Based on these structural descriptors and their CTI, a SAR model was derived. Then the carcinogenic possibility (CP) and the carcinogenic impossibility (CIP) of compounds were predicted. The model was tested from a testing set of 304 carcinogenic compounds (MDL toxicity database), 460 non-carcinogenic compounds (CMC database) and 94 compounds extracted from two traditional Chinese medicine herbs. PMID- 15293549 TI - The defense mechanisms in mammalian cells against oxidative damage in nucleic acids and their involvement in the suppression of mutagenesis and cell death. AB - To counteract oxidative damage in nucleic acids, mammalian cells are equipped with several defense mechanisms. We herein review that MTH1, MUTYH and OGG1 play important roles in mammalian cells avoiding an accumulation of oxidative DNA damage, both in the nuclear and mitochondrial genomes, thereby suppressing carcinogenesis and cell death. MTH1 efficiently hydrolyzes oxidized purine nucleoside triphosphates, such as 8-oxo-dGTP, 8-oxo-dATP and 2-hydroxy (OH)-dATP, to the monophosphates, thus avoiding the incorporation of such oxidized nucleotides into the nuclear and mitochondrial genomes. OGG1 excises 8-oxoG in DNA as a DNA glycosylase and thus minimizes the accumulation of 8-oxoG in the cellular genomes. MUTYH excises adenine opposite 8-oxoG, and thus suppresses 8 oxoG-induced mutagenesis. MUTYH also possesses a 2-OH-A DNA glycosylase activity for excising 2-OH-A incorporated into the cellular genomes. Increased susceptibilities to spontaneous carcinogenesis of the liver, lung or intestine were observed in MTH1-, OGG1- and MUTYH-null mice, respectively. The increased occurrence of lung tumors in OGG1-null mice was abolished by the concomitant disruption of the Mth1 gene, indicating that an increased accumulation of 8-oxoG and/or 2-OH-A might cause cell death. Furthermore, these defense mechanisms also likely play an important role in neuroprotection. PMID- 15293550 TI - The neurotoxic effect of sickle cell hemoglobin. AB - A growing body of experimental evidence suggests that the oxidative neurotoxicity of hemoglobin A may contribute to neuronal loss after CNS hemorrhage. Several hemoglobin variants, including hemoglobin S, are more potent oxidants in cell free systems. However, despite the increased incidence of hemorrhagic stroke associated with sickle cell disease, little is known of the effect of hemoglobin S on cells of neural origin. In the present study, its toxicity was quantified and directly compared with that of hemoglobin A in murine cortical cell cultures. Reactive oxygen species production, as assessed by cellular fluorescence after treatment with dihydrorhodamine 123, was significantly increased by exposure to 10 microM hemoglobin S for 2-4 h. Neuronal death, as measured by propidium iodide staining and lactate dehydrogenase release, commenced at 4 h; for a 20-h exposure, the EC50 was approximately 0.71 microm. Glial cells were not injured. Cell death was completely blocked by iron chelation with deferoxamine or phenanthroline. Direct comparison of sister cultures exposed to either hemoglobin A or hemoglobin S revealed a similar amount of cell injury in both groups. A significant difference was consistently observed only after treatment with 1 microM hemoglobin for 20 h, which resulted in death of approximately one third more neurons with hemoglobin S than with hemoglobin A. The results of this study suggest that sickle cell hemoglobin is neurotoxic at physiologically relevant concentrations. This toxicity is iron-dependent, oxidative, and quantitatively similar to that produced by hemoglobin A. PMID- 15293551 TI - Photo-irradiated titanium dioxide catalyzes site specific DNA damage via generation of hydrogen peroxide. AB - Titanium dioxide (TiO2) is a potential photosensitizer for photodynamic therapy. In this study, the mechanism of DNA damage catalyzed by photo-irradiated TiO2 was examined using [32P]-5'-end-labeled DNA fragments obtained from human genes. Photo-irradiated TiO2 (anatase and rutile) caused DNA cleavage frequently at the guanine residue in the presence of Cu(II) after E. coli formamidopyrimidine-DNA glycosylase treatment, and the thymine residue was also cleaved after piperidine treatment. Catalase, SOD and bathocuproine, a chelator of Cu(I), inhibited the DNA damage, suggesting the involvement of hydrogen peroxide, superoxide and Cu(I). The photocatalytic generation of Cu(I) from Cu(II) was decreased by the addition of SOD. These findings suggest that the inhibitory effect of SOD on DNA damage is due to the inhibition of the reduction of Cu(II) by superoxide. We also measured the formation of 8-oxo-7,8-dihydro-2'-deoxyguanosine, an indicator of oxidative DNA damage, and showed that anatase is more active than rutile. On the other hand, high concentration of anatase caused DNA damage in the absence of Cu(II). Typical free hydroxyl radical scavengers, such as ethanol, mannnitol, sodium formate and DMSO, inhibited the copper-independent DNA photodamage by anatase. In conclusion, photo-irradiated TiO2 particles catalyze the copper mediated site-specific DNA damage via the formation of hydrogen peroxide rather than that of a free hydroxyl radical. This DNA-damaging mechanism may participate in the phototoxicity of TiO2. PMID- 15293552 TI - Protective effect of the xanthate, D609, on Alzheimer's amyloid beta-peptide (1 42)-induced oxidative stress in primary neuronal cells. AB - Tricyclodecan-9-yl-xanthogenate (D609) is an inhibitor of phosphatidylcholine specific phospholipase C, and this agent also has been reported to protect rodents against oxidative damage induced by ionizing radiation. Previously, we showed that D609 mimics glutathione (GSH) functions and that a disulfide is formed upon oxidation of D609 and the resulting dixanthate is a substrate for GSH reductase, regenerating D609. Considerable attention has been focused on increasing the intracellular GSH levels in many diseases, including Alzheimer's disease (AD). Amyloid beta-peptide [Abeta(1-42)], elevated in AD brain, is associated with oxidative stress and toxicity. The present study aimed to investigate the protective effects of D609 on Abeta(1-42)-induced oxidative cell toxicity in cultured neurons. Decreased cell survival in neuronal cultures treated with Abeta(1-42) correlated with increased free radical production measured by dichlorofluorescein fluorescence and an increase in protein oxidation (protein carbonyl, 3-nitrotyrosine) and lipid peroxidation (4-hydroxy-2-nonenal) formation. Pretreatment of primary hippocampal cultures with D609 significantly attenuated Abeta(1-42)-induced cytotoxicity, intracellular ROS accumulation, protein oxidation, lipid peroxidation and apoptosis. Methylated D609, with the thiol functionality no longer able to form the disulfide upon oxidation, did not protect neuronal cells against Abeta(1-42)-induced oxidative stress. Our results suggest that D609 exerts protective effects against Abeta(1-42) toxicity by modulating oxidative stress. These results may be of importance for the treatment of AD and other oxidative stress-related diseases. PMID- 15293553 TI - 2-substituted-3H-indol-3-one-1-oxides: preparation and radical trapping properties. AB - A series of 2-alkyl and 2-aryl substituted-3H-indol-3-one-1-oxides was prepared and evaluated for its radical trapping properties. Spin trapping and electron paramagnetic resonance experiments demonstrate the ability of these indolone-1 oxides to trap hetero- and carbon-centered radicals. The most stable spin adducts (lifetime of several hours) are obtained with 2-alkyl substituted nitrones, the 2 ethyl-5,6-dioxolo-3H-indolone-1-oxide, 5e and the 2-secbutyl-3H-indolone-1-oxide, 5f. These two nitrones are also sensitive to redox reactions in solution. Therefore this indolone-1-oxide series lacking a beta-hydrogen atom gives rise to highly stable adducts with free radicals. PMID- 15293554 TI - Effects of antioxidant and nitric oxide on chemokine production in TNF-alpha stimulated human dermal microvascular endothelial cells. AB - Chemokines have been implicated convincingly in the driving of leukocyte emigration in different inflammatory reactions. Multiple signaling mechanisms are reported to be involved in intracellular activation of chemokine expression in vascular endothelial cells by various stimuli. Nevertheless, redox-regulated mechanisms of chemokine expression in human dermal microvascular endothelial cells (HDMEC) remain unclear. This study examined the effects of pyrrolidine dithiocarbamate (PDTC, 0.1 mM) and spermine NONOate (Sper-NO, 1 mM) on the secretion and gene expression of chemokines, interleukin (IL)-8, monocyte chemotactic protein (MCP)-1, regulated upon activation normal T cell expressed and secreted (RANTES), and eotaxin. This study also addresses PDTC and Sper-NO effects on activation of nuclear factor kappa B (NF-kappaB) induced by TNF-alpha (10 ng/ml). Treatment with TNF-alpha for 8 h significantly increased secretion of IL-8, MCP-1, and RANTES, but not of eotaxin, in cultured HDMEC. Up-regulation of these chemokines was suppressed significantly by pretreatment with PDTC or Sper NO for 1 h, but not by 1 mM 8-bromo-cyclic GMP. The mRNA accumulation of IL-8, MCP-1, RANTES, and eotaxin, and activation of NF-kappaB were induced by TNF-alpha for 2 h; all were suppressed significantly by the above two pretreatments. These findings indicate that both secretion and mRNA accumulation of IL-8, MCP-1, and RANTES in HDMEC induced by TNF-alpha are inhibited significantly by pretreatment with PDTC or Sper-NO, possibly via blocking redox-regulated NF-kappaB activation. These results suggest that restoration of the redox balance using antioxidant agents or nitric oxide pathway modulators may offer new opportunities for therapeutic interventions in inflammatory skin diseases. PMID- 15293555 TI - The semi-quantitative comparison of oxidative stress mediated DNA single and double strand breaks using terminal deoxynucleotidyl transferase mediated end labeling combined with a slot blot technique. AB - The accumulation of DNA damages by environmental stresses is represented by the steady state level of single strand breaks (SSBs) and double strand breaks (DSBs). Terminal deoxynucleotidyl transferase (TdT) mediated end labeling is suitable in detecting DSBs, but is unsuitable for SSBs due to its catalyzing characteristics. However, the sensitivity of TdT to detect SSBs may be significantly improved by first denaturing the double strands and expose all the DNA nicks as potential substrates for TdT. By coupling DNA denaturation to slot blot southern hybridization, the authors demonstrate the sensitive detection of SSBs as well as DSBs in 20 ng DNA samples derived from a retinal pigment epithelial cell line treated with tert-butyl hydroperoxide. The signal intensity of denatured and TdT-treated DNA in slot blot hybridization correlated to the amount of SSBs calculated in an S1 nuclease digestion assay. The signal ratio between denatured and non-denatured DNA likely approximates the SSBs/DSBs ratio in genomic DNA. The combination of DNA denaturing, TdT treatment and slot blot hybridization could be a useful method to assess oxidative stress-induced DNA strand damages. PMID- 15293556 TI - Mechanism of hydroxyl radical scavenging by rebamipide: identification of mono hydroxylated rebamipide as a major reaction product. AB - Rebamipide, an antiulcer agent, is known as a potent hydroxyl radical (*OH) scavenger. In the present study, we further characterized the scavenging effect of rebamipide against *OH generated by ultraviolet (UV) irradiation of hydrogen peroxide (H2O2), and identified the reaction products to elucidate the mechanism of the reaction. Scavenging effect of rebamipide was accessed by ESR using DMPO as a *OH-trapping agent after UVB exposure (305 nm) to H2O2 for 1 min in the presence of rebamipide. The signal intensity of *OH adduct of DMPO (DMPO-OH) was markedly reduced by rebamipide in a concentration-dependent fashion as well as by dimethyl sulfoxide and glutathione as reference radical scavengers. Their second order rate constant values were 5.62 x 10(10), 8.16 x 10(9) and 1.65 x 10(10) M( 1) s(-1), respectively. As the rebamipide absorption spectrum disappeared during the reaction, a new spectrum grew due to generation of rather specific reaction product. The reaction product was characterized by LC-MS/MS and NMR measurements. Finally, a hydroxylated rebamipide at the 3-position of the 2(1H)-quinolinone nucleus was newly identified as the major product exclusively formed in the reaction between rebamipide and the *OH generated by UVB/H2O2. Specific formation of this product explained the molecular characteristics of rebamipide as a potential *OH scavenger. PMID- 15293557 TI - Monosialoganglioside increases catalase activity in cerebral cortex of rats. AB - Monosialoganglioside (GM1) is a neuroprotective agent that has been reported to scavenge free radicals generated during reperfusion and to protect receptors and enzymes from oxidative damage. However, only a few studies have attempted to investigate the effects of GM1 on enzymatic antioxidant defenses of the brain. In the present study, we evaluate the effects of the systemic administration of GM1 on the activity of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px), and on spontaneous chemiluminescence and total radical trapping potential (TRAP) in cerebral cortex of rats ex vivo. The effects of GM1 on CAT activity and spontaneous chemiluminescence in vitro were also determined. Animals received two injections of GM1 (50 mg/kg, i.p.) or saline (0.85% NaCl, i.p.) spaced 24 h apart. Thirty minutes after the second injection the animals were sacrificed and enzyme activities and spontaneous chemiluminescence and TRAP were measured in cell-free homogenates. GM1 administration reduced spontaneous chemiluminescence and increased catalase activity ex vivo, but had no effect on TRAP, SOD or GSH-Px activities. GM1, at high concentrations, reduced CATactivity in vitro. We suggest that the antioxidant activity of GM1 ganglioside in the cerebral cortex may be due to an increased catalase activity. PMID- 15293559 TI - Effect of 4-hydroxynonenal on antioxidant capacity and apoptosis induction in Jurkat T cells. AB - 4-Hydroxynonenal (HNE) is one of the major end products of lipid peroxidation and may have either physiological or pathological significance regulating cell proliferation. We studied some biochemical effects of HNE, at various concentrations (0.1-100 microM), on Jurkat T cells incubated thereafter for 24, 48 and 72 h. HNE at low concentrations significantly enhanced the proliferation index, whereas at higher concentrations progressively blocked cell proliferation. Caspase 3 activity increased significantly at HNE concentrations between 1 and 10 microM and decreased at higher concentrations. Superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and glutathione reductase (GSH-Rd) increased progressively with HNE concentrations, particularly GSH-Px. Glucose-6-phosphate dehydrogenase (G6PDH) showed a different pattern, increasing at low HNE (1-5 microM) concentrations and rapidly declined thereafter. These results show that HNE may induce growth inhibition of Jurkat T cells and regulate the activity of typical antioxidant enzymes. Furthermore, the protective effect of doubling the foetal calf serum still points out the risk that cultured cells undergo oxidative stress during incubation. PMID- 15293558 TI - Antioxidants inhibit angiogenesis in vivo through down-regulation of nitric oxide synthase expression and activity. AB - Although reactive oxygen species (ROS) participate in many cellular mechanisms, only few data exist concerning their involvement in physiological angiogenesis. The aim of the present work was to elucidate possible mechanisms through which ROS affect angiogenesis in vivo, using the model of the chicken embryo chorioallantoic membrane (CAM). Superoxide dismutase (SOD) and its membrane permeable mimetic tempol, dose dependently decreased angiogenesis and down regulated inducible nitric oxide synthase (iNOS) expression and nitric oxide (NO) production. The NADPH oxidase inhibitors, 4-(2-aminoethyl)-benzenesulfonyl fluoride (AEBSF) and apocynin, but not allopurinol, also had a dose dependent inhibitory effect on angiogenesis and NO production in vivo. Catalase and the intracellular hydrogen peroxide (H2O2) scavenger sodium pyruvate decreased, while H2O2 increased in a dose-dependent manner the number of CAM blood vessels, as well as the expression and activity of iNOS. Dexamethasone, which down-regulated NO production by iNOS and L-NAME, but not D-NAME, dose dependently decreased angiogenesis in vivo. These data suggest that antioxidants affect physiological angiogenesis in vivo, through regulation of NOS expression and activity. PMID- 15293560 TI - Oxidative stress and myocardial damage during elective percutaneous coronary interventions and coronary angiography. A comparison of blood-borne isoprostane and troponin release. AB - The role of oxidative stress in clinical cardiology is still controversial. The aims of the present study were to examine if minor ischaemic episodes as may occur during elective percutaneous coronary intervention (PCI) induce oxidative stress and, eventually, if oxygen stress correlates with myocardial injury. Thirty eight and nine patients underwent PCI and diagnostic coronary angiography, respectively. Peripheral blood was sampled at different time points for plasma analyses of: 8-iso-PGF2alpha (free radical-mediated oxidative stress); 15-keto dihydro-PGF2alpha (cyclooxygenase-mediated inflammation); troponin-T (myocardial injury); hsCRP, vitamin A and vitamin E; and, total antioxidants status (TAS). In both groups 8-iso-PGF2alpha increased transiently by approximately 80% (p < 0.001) during the procedure. There was a minor troponin-T release (p < 0.001) after PCI, but no correlation with 8-iso-PGF2alpha. Troponin-T did not increase after angiography. 15-keto-dihydro-PGF2alpha decreased by 50% after ended procedure, but increased by 100% after 24 h compared to baseline. hsCRP increased significantly (p < 0.001) from baseline to the next day in the PCI-group, but not in the angiography group. Vitamins and TAS decreased slightly after the procedures. It is concluded that a moderate oxidative stress was induced by both elective PCI and coronary angiography but that no correlation was found between oxidative stress and myocardial injury in this setting. This indicates that other mechanisms than ischaemia-reperfusion episodes caused an elevation in plasma isoprostane such like the injury at a vascular site mutual for both procedures. A secondary finding from the study was elevated markers of early inflammatory response, not only after PCI, but also after angiography. PMID- 15293561 TI - Combination of ascorbate and alpha-tocopherol as a preventive therapy against structural and functional defects of erythrocytes in visceral leishmaniasis. AB - The redox unbalance in erythrocytes has been found to contribute significantly in the development of anemia in visceral leishmaniasis (VL). The present study revealed enhanced production of reactive oxygen species (ROS) and gradual depletion of alpha-tocopherol and ascorbate in the erythrocytes of infected animals. The response of erythrocytes to chronic treatment with antioxidants was studied in hamsters during leishmanial infection. Treatment with a combination of alpha-tocopherol and ascorbate proved to be the most effective preventive for the proteolytic degradation of erythrocyte membrane. Erythrocytes from infected animals were thermally more sensitive compared to the control ones. Combination of both antioxidants was most successful in resisting heat induced structural defects in the cells. Cross-linking of membrane proteins subsequent to oxidative damage in the red cells was accompanied by the formation of high molecular weight protein band at the top of the resolving gel in the presence of the cross-linking agent dimethyladepimidate (DMA). Marked inhibition of cross-linking was observed with combination of both antioxidants. Treatment with alpha-tocopherol and ascorbate together could withstand osmotic lysis of erythrocytes in the infected animals very efficiently. Decreased hemoglobin (Hb) level was successfully replenished and was coupled with significant increase in the life span of red cells after treating the animals with both antioxidants. Results indicate better efficacy of the combination therapy with alpha-tocopherol and ascorbate in protecting the erythrocytes from structural and functional damages during leishmanial infection. PMID- 15293562 TI - Differentiation and diversification of vascular cells from embryonic stem cells. AB - Pluripotent embryonic stem (ES) cells are potent materials for both regenerative therapeutic approaches and developmental research. Recently, a novel ES cell differentiation system combined with 2-dimensional culture and flow cytometry assisted cell sorting has been developed. In this system, endothelial, mural, and blood cells can be systematically induced from common progenitor vascular endothelial growth factor receptor-2 (Flk1)-expressing cells. This system is amenable for the in vitro observation of multiple steps of the vascular developmental process, such as vascular cell differentiation and diversification from progenitors, endothelial cell maturation and differentiation into arterial, venous, and lymphatic endothelium, and vascular formation. This constructive in vitro approach provides novel possibilities for elucidating the cellular and molecular mechanisms of vascular development. Vascular cell induction from primate ES cells reveals primate-specific vascular developmental mechanisms. ES cell research in developmental biology would be indispensable, especially in the human species for which a knock-out animal model is not available. ES cells should also contribute to regenerative medicine, not only as a cellular source for transplantation but also for the discovery of novel genes and drugs for regeneration. In this review, the significance of ES cell study in basic science and clinical medicine in the vascular field is discussed. PMID- 15293564 TI - Angiopoietin-related/angiopoietin-like proteins regulate angiogenesis. AB - A general understanding of the molecular mechanisms underlying angiogenesis is emerging from the analysis of targeted mutations in vasculature-related genes. These analyses reveal that angiopoietin signaling through the TIE2 receptor is involved in regulating angiogenesis. Recently, we and several other groups have independently identified several molecules containing a coiled-coil domain and a fibrinogen-like domain, both of which are structurally conserved in angiopoietins. Because these molecules do not bind to the angiopoietin-specific receptor,TIE2, they have been named angiopoietin-related proteins (ARPs) or angiopoietin-like proteins (Angptls). ARPs/Angptls, which are all currently orphan ligands, also have potent activity for regulating angiogenesis as proangiogenic or antiangiogenic factors, suggesting that their receptors may be expressed on endothelial cells. In addition, ARPs/Angptls show pleiotropic effects not only on vascular cells but also on cells of other lineages, such as skin and chondrocyte cells. More recent studies have proposed that ARPs/Angptls are involved in various pathologies, such as tumor angiogenesis and metabolic diseases. To summarize the current findings relating to these proteins, we focus in this review on the functions of ARPs/Angptls as new angiogenic modulating factors in the vascular system and discuss the pleiotropic functions of ARPs/Angptls in nonvascular cell lineages. PMID- 15293563 TI - Vascular endothelial growth factor and other signaling pathways in developmental and pathologic angiogenesis. AB - The field of angiogenesis received a huge boost in 2003 with the announcement of positive results in a phase III clinical trial using a vascular endothelial growth factor (VEGF)-blocking antibody for the treatment of cancer. Although the VEGF pathway has emerged as a central signaling pathway in normal and pathologic angiogenesis, several other pathways are also now recognized as playing essential roles. This review focuses on 2 specific areas. First, we summarize some of the work on newly discovered angiogenic signaling pathways by primarily describing the molecular biology of the pathways and the evidence for their involvement in vascular development. Second, we describe progress in therapeutic antiangiogenesis in cancer, particularly with agents that block the VEGF pathway. PMID- 15293565 TI - Molecular mechanisms of lymphangiogenesis. AB - Although the process of vascular development has been well documented, little is understood about lymphatic vasculature formation, despite its importance in normal and pathologic conditions. The dysfunction or abnormal growth of lymphatic vessels is associated with lymphedema and cancer metastasis. The recent discovery of lymphangiogenic growth factors vascular endothelial growth factor (VEGF)-C and VEGF-D and of their receptor, VEGFR-3, on lymphatic endothelial cells has started to provide an understanding of the molecular mechanisms of lymphangiogenesis. In addition, other genes that participate in the specification of lymphatic endothelial cells and the modulation of lymphatic vascular development have been identified. The capacity to induce or inhibit lymphangiogenesis by the manipulation of such molecules offers new opportunities to understand the function of the lymphatic system and to develop novel treatments for lymphatic disorders. This review describes the main players in lymphangiogenesis that have been identified so far and the attempts to shed some light on the mysteries surrounding this process. PMID- 15293566 TI - Response to cyclosporine therapy in patients with myelodysplastic syndrome: a clinical study of 12 cases and literature review. AB - Cyclosporine (CyA) was administered to 12 patients with myelodysplastic syndrome (MDS), and a response (major erythroid response, according to International Working Group criteria) was observed in 7 patients (58.3%). The median duration of response was 18 months (range, 3-22 months). Two patients are still responding and continuing to take CyA. Three patients stopped because of malignancy complications. To identify variables associated with responsiveness to CyA therapy, we analyzed the treatments of 72 MDS patients, comprising the 12 new patients and 60 patients previously described in the literature. Responses were observed in 44 of the 72 patients (61.1%). Univariate analyses revealed that higher daily dose of CyA (P for trend test, .007) and shorter disease duration (median, 5 months versus 17.5 months, P = .04) were factors significantly associated with response. No significant associations were observed between response and bone marrow features such as erythroid hypoplasia or hypoplastic marrow. Multivariate analysis also demonstrated that high CyA dose (>5 mg/kg per day) was significantly associated with response (P = .02). The present study showed that CyA therapy is useful for MDS patients with any marrow cellularity. Shorter disease duration is a pretreatment variable correlated with response, and a higher CyA dose results in a higher response rate. PMID- 15293567 TI - Two entities of precursor T-cell lymphoblastic leukemia/lymphoma based on radiologic and immunophenotypic findings. AB - Precursor T-cell lymphoblastic leukemia/lymphoma (T-ALL/LBL) presents a mediastinal mass in one half of cases. Although the immunophenotypic features of T-ALL/LBL have been analyzed in several studies, few studies have been focused on the relationship between the anatomic distribution of lesions and immunophenotypic findings. We analyzed the clinicopathologic findings for 17 patients with T-ALL/LBL diagnosed since 1993 and whose radiologic findings were available. Data on 14 men and 3 women with a median age of 26 years (range, 10-61 years) were analyzed. On the basis of radiologic findings, the cases were divided into thymic type (n = 8) and nonthymic type (n = 9). Patients with the thymic type of T-ALL/LBL had a large mediastinal mass and minimal systemic lymphadenopathy only in the supradiaphragmatic region. Those with the non-thymic type had predominantly systemic lymphadenopathy that included infradiaphragmatic lesions. Expression of CD8 (6/7 versus 0/9) was more frequently found in the thymic type (P < .001), whereas expression of CD56 (0/7 versus 5/9) was more frequent in the nonthymic type (P = .034). In conclusion, T-ALL/LBL was divided into 2 entities, thymic type and nonthymic type, on the basis of radiologic findings and immunophenotypic features. Analysis of the expression of CD8 and CD56 would be useful for biologically classifying T-ALL/LBL into the 2 types. This study was performed in a single institution, was retrospective, and had a limited number of patients; multicenter confirmatory studies are warranted. PMID- 15293568 TI - Clinical relevance of survivin as a biomarker in neoplasms, especially in adult T cell leukemias and acute leukemias. AB - Survivin has been identified as one of the top 4 transcripts among 3.5 million human transcriptomes uniformly up-regulated in cancer tissues but not in normal tissues. Therefore, we quantitatively determined the messenger RNA (mRNA) expression profile for survivin by a real-time reverse transcriptase-polymerase chain reaction (RT-PCR) technique in 113 patients with leukemias, such as adult T cell leukemia (ATL), acute lymphoid leukemia (ALL), acute myeloid leukemia (AML), chronic myeloid leukemia in crisis, and chronic lymphocytic leukemia (CLL), and in 25 cell lines, including 7 ATL cell lines and 15 solid-tumor cell lines. Furthermore, we examined whether the plasma level of survivin protein as measured by enzyme-linked immunosorbent assay (ELISA) substituted for mRNA expression by PCR quantification. Gene expression was quantitatively confirmed to be up regulated in approximately 90% of ATL and acute leukemia cases and in all of the cell lines tested, whereas it was down-regulated in almost all cases of CLL. Furthermore, with respect to the interpretation of the gene expression findings, attention was paid to standardization with a housekeeping gene, glyceraldehyde-3 phosphate dehydrogenase (GAPDH), in the real-time PCR quantification, because the variability in GAPDH expression among the different cell types was significant. GAPDH expression was relatively low in ATL cells and high in ALL and AML cells. The rates of increase in the levels of survivin protein in the plasma of ATL patients and in the supernatants from in vitro cultures of solid-tumor cell lines were low compared with rates of increase of the mRNA and protein level in the cells, suggesting that the protein levels in plasma do not always reflect survivin expression in tumor cells. Our findings indicate the potential clinical relevance of survivin quantified by real-time PCR but not for the protein level in plasma as determined by ELISA, especially in cases of ATL and acute leukemias. PMID- 15293569 TI - Exaggerated cutaneous response to mosquito bites in a patient with chronic lymphocytic leukemia. AB - Although various cutaneous manifestations can occur in patients with chronic lymphocytic leukemia (CLL), bullous lesions due to mosquito bites are rare. We describe a patient with a typical case of CLL complicated by exaggerated cutaneous response to mosquito bites. The symptoms of this patient were self limited and not severe, but it has been reported that the treatment of such symptoms is sometimes difficult. Because exaggerated cutaneous hypersensitivity to mosquito bites can be observed even before the diagnosis of hematological malignancies such as CLL, careful examinations are recommended to find underlying diseases in cases in which this type of cutaneous reaction occurs. PMID- 15293570 TI - Existence of leukemic clones resistant to both imatinib mesylate and rituximab before drug therapies in a patient with Philadelphia chromosome-positive acute lymphocytic leukemia. AB - Imatinib mesylate and rituximab are molecularly targeted drugs against the BCR ABL fusion protein and the CD20 antigen, respectively. Although these drugs have excellent anticancer effects, a major concern is drug resistance. We have investigated the case of a patient with Philadelphia chromosome-positive and CD20+ acute lymphocytic leukemia who acquired resistance to imatinib and rituximab. Imatinib therapy resulted in prompt cytogenetic remission, but resistance developed shortly thereafter. Sequencing of the kinase domain of the ABL gene and allele-specific polymerase chain reaction analysis revealed a point mutation resulting in an E255V substitution that was present before the therapy. After the patient received mild chemotherapy followed by rituximab administration, hematologic and cytogenetic remission was sustained for 5.5 months. The recurrent leukemic cells after the rituximab therapy showed not only the E255V mutation in the ABL gene but also loss of the CD20 antigen due to impaired transcription of the CD20 gene. The results of 2-color flow cytometry analysis showed that a small population of CD20(-) leukemic cells existed before the imatinib therapy. These results suggest that leukemic subclones carrying a genetic perturbation of the targeted molecules for both imatinib and rituximab were present before the therapies. The preexistence of primary resistant clones suggests the inability of combination therapy with 2 molecularly targeted drugs to overcome drug resistance in leukemia. PMID- 15293571 TI - Sustained cytogenetic remission induced by imatinib mesylate in a chronic myeloid leukemia patient who had a relapse into lymphoid crisis after allogeneic hematopoietic stem cell transplantation. AB - We report on the response to imatinib mesylate in a chronic myeloid leukemia patient who, after undergoing allogeneic hematopoietic stem cell transplantation (alloHSCT) for treatment of lymphoid blastic crisis, had a relapse into blastic crisis despite the presence of chronic and grade II acute graft-versus-host disease (GVHD). Complete hematologic response and the disappearance of the Bcr Abl fusion signal on fluorescence in situ hybridization analysis were achieved after 10 weeks of imatinib therapy and were maintained for 26 months with no adverse effects, including recurrence of GVHD. This case highlights the ability of imatinib to induce sustained hematologic and cytogenetic remission in some patients who have had relapses into advanced-stage chronic myeloid leukemia after alloHSCT. PMID- 15293572 TI - T(11;18)-bearing pulmonary mucosa-associated lymphoid tissue lymphoma responding to cladribine. AB - Mucosa-associated lymphoid tissue (MALT) lymphoma is a low-grade B-cell lymphoma and is usually slow to disseminate. The t(11;18)(q21;q21) translocation has been identified in a subset of MALT lymphoma cases, and AP12 and MLT1/MALT1 genes have been implicated in this translocation. However, the clinicopathologic features of t(11;18)-bearing MALT lymphoma have not been fully elucidated, and the optimal therapy for patients with disseminating disease is unknown. We report an outstanding case of MALT lymphoma that showed massive pulmonary infiltration and leukemic transformation during a prolonged course of more than 16 years. Reverse transcriptase-polymerase chain reaction and dual-color fluorescence in situ hybridization analyses confirmed the existence of the AP12/MLT1 fusion gene in the lymphoma cells. Peripheral blood lymphoma cells disappeared after glucocorticoid therapy. Although the pulmonary lymphoma was slowly progressive and caused severe hypoxemia despite the continuation of glucocorticoid therapy, treatment with cladribine induced a marked reduction of pulmonary lymphomatous lesions and an improvement of the hypoxemia. These findings show the unique clinical features of this particular indolent B-cell lymphoma with t(11;18) translocation and suggest the potential therapeutic usefulness of glucocorticoid and cladribine. PMID- 15293573 TI - Successful treatment of idiopathic hypereosinophilic syndrome with imatinib mesylate: a case report. AB - Patients with idiopathic hypereosinophilic syndrome (HES) show persistent hypereosinophilia of unknown etiology that is associated with end-organ damage. Different treatments, including the use of corticosteroids and cytotoxics, have been investigated for HES with modest success. We describe a patient with HES who had significant end-organ damage from hypereosinophilia and remained refractory to conventional therapy. Therapy with imatinib mesylate, a selective tyrosine kinase inhibitor that is highly effective in treating patients with BCR-ABL positive chronic myeloid leukemia, was tried with the patient. The result was impressive, with hematologic remission achieved after 12 days of administration. Our finding concurs with recent reports that imatinib mesylate may be a promising agent in the treatment of some cases of HES. PMID- 15293574 TI - Unmanipulated HLA-haploidentical bone marrow transplantation for the treatment of fatal, nonmalignant diseases in children and adolescents. AB - Fetomaternal microchimerism has been demonstrated, and immunologic tolerance to unshared HLA antigens between mother and offspring may be suggested. We used T cell-repleted bone marrow transplantation (BMT) from their HLA-haploidentical mothers to treat 6 patients with fatal nonmalignant diseases. The number of mismatched HLA loci in the graft-versus-host disease (GVHD) direction was 3 in 4 patients and 2 in 2 patients. The number in the host-versus-graft direction was 3 in 4 patients, 2 in 1 patient, and 1 in 1 patient. Microchimerism of inherited paternal antigens was demonstrated in 5 donors, and microchimerism of noninherited maternal antigens was detected in 3 recipients. GVHD prophylaxis consisted of short-course methotrexate, tacrolimus, and mycophenolate mofetil (3 patients) or short-course methotrexate, tacrolimus, and methylprednisolone (1 patient). Engraftment was achieved in 5 patients who had received preconditioning, and T-cell engraftment was confirmed in 1 patient with severe combined immunodeficiency. Acute GVHD developed in 3 patients: grade 1 in 2 patients and grade 2 in 1 patient. Chronic GVHD was observed in 5 patients: localized type in 3 patients and extended type in 2 patients. Five patients were alive 11 to 30 months after BMT and 1 patient died of chronic GVHD. Unmanipulated haploidentical BMT from a maternal donor may be the treatment of choice of poor prognosis nonmalignant diseases. PMID- 15293575 TI - Glanzmann thrombasthenia associated with a 21-amino acid deletion (Leu817-Gln837) in glycoprotein IIb due to abnormal splicing in exon 25. AB - We report a novel genetic defect in a Japanese patient with type I Glanzmann thrombasthenia. The glycoprotein (GP) Ilb complementary DNA (cDNA) from platelet messenger RNA had a 63-base pair deletion in the 5' boundary of exon 25, resulting in an in-frame deletion of 21 amino acid residues (Leu817-Gln837) in the calf-2 domain. The deleted region was present in the genomic DNA, but the splice acceptor site (AG) of exon 25 was mutated to AC, leading to the use of an AG sequence in the middle of exon 25 as an abnormal cryptic splice acceptor site. The effect of this deletion on protein synthesis was further analyzed. Mutant GPIIb-IIIa complexes were not detected on the surfaces of cells cotransfected with cDNAs of mutant GPIIb and normal GPIIIa. Mutant pro-GPIIb was detected in cell lysates and was coimmunoprecipitated with an anti-GPIIb-IIIa complex antibody. Immunostaining demonstrated that the mutant pro-GPIIb colocalized with an endoplasmic reticulum protein, calnexin, within the cells. These results indicate that complex formation was not completely prevented and that impairment of the subsequent transport was the major reason for the defect in cell surface expression. The data suggest that the GPIIb calf-2 domain is important for intracellular transport of GPIIIb-IIIa complexes. PMID- 15293576 TI - The effects of desflurane on human platelet aggregation in vitro. AB - In view of the possible antiplatelet effects of general anesthetics, we investigated the in vitro effects of desflurane, a new inhalation agent, on platelet aggregation. For 15 patients who underwent elective operations, blood was sampled with desflurane induction before and after anesthesia but prior to surgery so that platelet aggregation in the drawn blood could be tested before desflurane anesthesia and again after exposure to the anesthetic. Platelet aggregation was measured with a whole-blood aggregometer. Adenosine diphosphate (ADP), collagen, and ristocetin were used as aggregating agents. Our results showed that aggregation in response to ADP, collagen, or ristocetin was not inhibited in patients who received desflurane anesthesia. This study with an in vitro model showed that desflurane had no influence on platelets in clinically relevant doses. PMID- 15293577 TI - Treatment of resistant thrombotic thrombocytopenic purpura with rituximab and cyclophosphamide. AB - Thrombotic thrombocytopenic purpura (TTP) is an uncommon acquired disease in adults, especially young women, characterized by fever, neurologic manifestations, microangiopathic hemolytic anemia, thrombocytopenia, and renal dysfunction. Treatment with plasmapheresis has increased the survival rate from 10% to greater than 90%. Still, a subset of patients with resistant TTP fail to respond to plasmapheresis or remain dependent on this procedure. We report such a patient who was successfully treated with rituximab and cyclophosphamide. She has now been disease free for more than 6 months. This novel treatment modality for TTP has been described for only a few patients. A well-controlled clinical trial is warranted to determine the role and place of this therapeutic approach in the management of TTP. PMID- 15293578 TI - False-positive plasma (1-->3)-beta-D-glucan test following immunoglobulin product replacement in an adult bone marrow recipient. PMID- 15293579 TI - Procrastination is the thief of time: surviving guidelines. PMID- 15293580 TI - Arginine immunonutrition in critically ill patients. PMID- 15293581 TI - Appropriately timed analgesics control pain due to chest tube removal. AB - BACKGROUND: Pain during chest tube removal can be moderately to severely intense and distressful to patients. Little evidence-based research has guided clinicians in attempts to alleviate such pain. OBJECTIVE: To test pharmacological and nonpharmacological interventions to alleviate pain during chest tube removal in cardiac surgery patients. METHODS: Four interventions were tested in 74 patients in a randomized, double-blind study: (1) 4 mg intravenous morphine and procedural information; (2) 30 mg intravenous ketorolac and procedural information; (3) 4 mg intravenous morphine plus procedural and sensory information; and (4) 30 mg intravenous ketorolac plus procedural and sensory information. Analgesics were administered to correspond to peak effect, and scripted information was provided. Pain intensity and pain distress were measured before analgesic administration, immediately after chest tube removal, and 20 minutes later Pain quality was measured immediately after chest tube removal. Level of sedation was measured before and 20 minutes after chest tube removal. Repeated-measures analyses of variance were used to test differences among groups over time. RESULTS: Pain intensity, pain distress, and sedation levels did not differ significantly among groups. However, procedural pain intensity (mean 3.26, SD 3.00) and pain distress (mean 2.98, SD 3.18) scores for all were low. Patients remained alert, regardless of which analgesic was administered. CONCLUSIONS: If used correctly, either an opioid (morphine) or a nonsteroidal anti-inflammatory (ketorolac) can substantially reduce pain during chest tube removal without causing adverse sedative effects. Thus, clinicians may choose among several safe and effective analgesic interventions during chest tube removal. PMID- 15293582 TI - Strategies for behavior change in patients with heart failure. AB - Appropriate management of chronic heart failure and its signs and symptoms requires a considerable amount of participation by patients. Behavioral changes that prevent or minimize signs and symptoms and disease progression are just as important as the medications prescribed to treat the heart failure. The most difficult lifestyle changes include smoking cessation, weight loss, and restriction of dietary sodium. The Transtheoretical Model is a framework for assessing and addressing the concept of readiness for behavior change, which occurs in a 6-step process. The model consists of 3 dimensions: the stages of change, the processes of change on which interventions are based, and the action criteria for actual behavior. The stages of change are discussed, and interventions are presented to assist patients with heart failure in progressing through those stages toward maintenance of changed lifestyle behaviors. Methods for measuring the level of readiness for change of patients with heart failure are also presented, because correct staging is required before appropriate interventions matched to a patient's stage can be delivered. PMID- 15293583 TI - Cannulation injuries of the radial artery. PMID- 15293584 TI - Cannulation injury of the radial artery: diagnosis and treatment algorithm. AB - Cannulation of the radial artery can result in complications ranging from arterial thrombosis, arterial aneurysm, compartment syndrome, infection, nerve injury and skin necrosis to possible thumb or even hand necrosis if not recognized and treated early. The anatomy of the radial artery, the diagnosis of injury, and a treatment algorithm are presented so that potential devastating hand complications can be avoided. PMID- 15293585 TI - Critical care research: weaving a body-mind-spirit tapestry. AB - Master weavers historically characterize the weaving of a tapestry as a calling, a transformation, a healing or sacred work. Tapestries are created by the collective efforts of many and are configured by the weavers' consciousness and spirit. A holistic framework used to weave a body-mind-spirit tapestry for guiding holistic clinical practice and research is described. Various research studies that document the effects of holistic interventions on patients' outcomes are examined. Implications for clinical practice are explored. PMID- 15293586 TI - Documentation on withdrawal of life support in adult patients in the intensive care unit. AB - BACKGROUND: Patients' charts have been a source of data for retrospective studies of the quality of end-of-life care. In the intensive care unit, most patients die after withdrawal of life support. Chart reviews of this process could be used not only to assess the quality of documentation but also to provide information for quality improvement and research. OBJECTIVE: To assess the documentation of end of-life care of patients and their families by care providers in the intensive care unit. METHOD: Charts of 50 adult patients who died in the intensive care unit at a large midwestern hospital after initiation of withdrawal of life support (primarily mechanical ventilation) were reviewed. A form developed for the study was used for data collection. RESULTS: The initiation of the decision making for withdrawal was documented in all 50 charts. Sixteen charts (32%) had no information on advance directives. Eight charts (16%) had no documentation on resuscitation status. About two thirds of the charts documented nurses' participation during the withdrawal process; only one tenth documented physicians' participation. A total of 13 charts (26%) had no information on the time of initiation of the withdrawal process, and 11 (22%) had no documentation of medications administered for withdrawal. Thirty-seven charts (74%) had information on whether the patient was or was not extubated during withdrawal. CONCLUSION: Comprehensive documentation of end-of-life care is lacking. PMID- 15293587 TI - Randomized trial of an intensive care unit-based early discharge planning intervention for critically ill elderly patients. AB - BACKGROUND: Few investigators have targeted elderly patients and monitored outcomes of care in studies on discharge planning interventions after critical illness. OBJECTIVES: To pilot test an intensive care unit-based nursing screening intervention to assist in determining the discharge needs and outcomes of critically ill elderly patients. METHOD: A randomized clinical trial with in hospital and mailed questionnaires was used. Patients 65 years and older who were hospitalized in 1 of 2 intensive care units at 2 midwestern university-affiliated medical centers were recruited for the study. Control patients (n = 53) received usual discharge planning, experimental patients (n = 47) were screened in the intensive care unit by using the Discharge Planning Questionnaire. Both groups were assessed for readiness for discharge when discharged from the hospital and were followed up 2 weeks later with a survey completed at home. RESULTS: One hundred patients 65 to 90 years old (mean 73, SD 5.78) completed the study. Sixty six percent were men. The 2 groups did not differ with regard to age, race, sex, severity of illness, lengths of stay in the intensive care unit or hospital, education level, or income. Patients in the experimental group were more ready than patients in the control group for discharge (P =.06). Patients in the experimental group were also more likely to report they had adequate information, had less concern about managing their care at home, knew their medicines, and knew danger signals indicating potential complications. CONCLUSION: Intensive care unit-based early discharge planning can affect elderly patients' preparation for discharge. PMID- 15293588 TI - Relational ethics of comfort, touch, and solace-endangered arts? PMID- 15293589 TI - Prinzmetal's angina. AB - Prinzmetal's angina, often referred to as "variant" angina, is a temporary increase in coronary vascular tone (vasospasm) causing a marked, but transient reduction in luminal diameter. This coronary vasospastic state is usually focal at a single site and can occur in either a normal or diseased vessel. Patients are predominantly younger women who may not have the classical cardiovascular risk factors (except for cigarette use). PVA has been associated with vasospastic disorders such as Raynaud's phenomenon and migraine headaches. Arrhythmias are common and may be life threatening especially when the effects of vasospasm are seen in those ECG leads that reflect the potential variations of the epicardial surface of the left ventricle. Endothelial dysfunction has been considered as primarily responsible for PVA. The diagnosis is made by observing transient ST segment elevation during the attack of angina. Since PVA is not a "demand"- induced symptom, but rather a supply (vasospastic) abnormality, exercise treadmill stress testing is of no value in the diagnosis of PVA. The most sensitive and specific test for PVA is the administration of ergonovine intravenously. Fifty micrograms at 5-minute intervals is given until a positive result or a maximum dose of 400 microg has been administered. When positive, the symptoms and associated ST-segment elevation should be present. Nitroglycerin rapidly reverses the effects of ergonovine if refractory spasm occurs. Medical therapy classically employs vasodilator drugs, which include nitrates and calcium channel blockers. The prognosis is good when there is no significant coronary artery stenosis. Treatment of associated coronary atherosclerosis in elderly patients with PVA is advised. When PVA is associated with coronary atherosclerosis, the prognosis is determined by the severity of the underlying disease. beta-Blockers and large doses of aspirin are contraindicated in PVA. PMID- 15293590 TI - Wide QRS duration. PMID- 15293591 TI - Yes, Dr Still, an MD won the 2004 Northup Writing Award! PMID- 15293592 TI - Prevalence of prevention and treatment modalities used in populations at risk of osteoporosis. AB - STUDY OBJECTIVE: Determine prevalence of osteoporosis screening and prevention and modes of treatment in women older than 65 years at risk of osteoporosis. METHODS: Retrospective chart review of older female patients seeking osteoporosis screening in the community setting. RESULTS: 399 women at risk of low bone mineral density (BMD) underwent dual-energy x-ray absorptiometry scanning. Among participants younger than 65 years (n=52), low BMD was diagnosed in 44.2%; among participants older than 65 years (n=347), low BMD was diagnosed in 70.0%, a statistically significant difference (P=.001). CONCLUSION: From a community-level perspective, the authors have shown that osteoporosis screening at local senior centers, living facilities, and health fairs is an effective tool for identifying low BMD in women at high risk of osteopenia and osteoporosis. PMID- 15293593 TI - Levalbuterol in the treatment of patients with asthma and chronic obstructive lung disease. AB - Effective asthma control requires long-term (anti-inflammatory) controller medications for patients with mild-persistent to severe-persistent disease, and quick-relief bronchodilator medication for all patients with asthma to control intermittent symptoms of cough, wheeze, and bronchoconstriction, as well as acute exacerbations. For patients with chronic obstructive pulmonary disease, quick relief and long-acting bronchodilators are primarily used in the maintenance and treatment of associated symptoms, including shortness of breath. For many years, the most widely used bronchodilator has been racemic (R, S)-albuterol, a short acting beta2-adrenergic agonist, commonly dispensed as an inhaled aerosol or solution. Until the introduction of levalbuterol inhalation solution (Xopenex) in 1999, all marketed forms of albuterol (including Ventolin and Proventil brands) were racemic mixtures composed of a 1:1 ratio of (R)- and (S)-stereoisomers. Administered as a proportionally equivalent nebulized dose, levalbuterol [(R) albuterol] provides greater bronchodilation than racemic albuterol and, in the appropriate clinical setting, offers the possibility for improving clinical outcomes in patients with asthma and other obstructive airway diseases. Additionally, levalbuterol can be given at lower doses than racemic albuterol to provide comparable bronchodilation, with the potential for reduced beta-mediated adverse effects in adults and children. Only since the past decade has the technology to separate stereoisomers become available, and thus the biologic activities of the albuterol stereoisomers had not been established. Binding studies have demonstrated that (R)-albuterol binds to the beta2-adrenergic receptor with a high affinity, whereas (S)-albuterol binds with 100-fold less affinity than (R)-albuterol. Other evaluations have suggested that (R)-albuterol possesses the bronchodilatory, bronchoprotective, and ciliary-stimulatory properties of racemic albuterol, while (S)-albuterol does not contribute beneficially to the therapeutic effects of the racemate and was originally assumed to be inert. However, preclinical evaluations have shown that (S) albuterol has effects that work in opposition to (R)-albuterol and may diminish the therapeutic effects of (R)-albuterol. PMID- 15293594 TI - Performance on the American Osteopathic Board of Internal Medicine certifying examination 1986-2002 of various demographic groups and the impact of AOA reentry resolutions on allopathic-trained candidates taking the examination. AB - The authors report the performance levels and pass rates of various candidate demographic groups and the effect on performance of delaying taking the certifying examination. They also report on the effect American Osteopathic Association reentry resolutions have on allopathic-trained candidates entering the osteopathic certification process in internal medicine. Included in the study were all candidates for the American Osteopathic Board of Internal Medicine certifying examination for the period between 1986 and 2002. Investigators performed group analysis based on type of residency track leading to board eligibility, as well as on the number of retake candidates, candidates reestablishing board eligibility 6 or more years after completion of residency training, and allopathic-trained candidates. Results indicate that medicine-track candidates performed better than any other study demographic group, including allopathic-trained candidates. A delay in taking the certifying examination after completion of residency results in lower candidate performance and pass rates. Various AOA reentry resolutions have not been successful in the repatriating of allopathic internal medicine-trained candidates into the certification process. Candidates in larger training programs have similar mean performance levels and pass rates as candidates in smaller programs. PMID- 15293595 TI - Is there a place for anti-remodelling drugs in asthma which may not display immediate clinical efficacy? PMID- 15293596 TI - Autoimmunity, T-cells and STAT-4 in the pathogenesis of chronic obstructive pulmonary disease. PMID- 15293597 TI - Limitation to muscular activity in chronic obstructive pulmonary disease. PMID- 15293598 TI - Fixed obstruction in severe asthma: not just a matter of time. PMID- 15293599 TI - Tuberculosis in HIV-infected persons in the context of wide availability of highly active antiretroviral therapy. AB - Highly active antiretroviral therapy (HAART) greatly reduces the risk of developing tuberculosis for HIV-infected persons. Nonetheless, HIV-associated tuberculosis continues to occur in countries where HAART is widely used. To identify the characteristics of HIV-infected persons who develop tuberculosis in the context of the availability of HAART, the current authors analysed data taken from 271 patients diagnosed, in Italy, during 1999-2000. These patients represent 0.7% of the 40,413 HIV-infected patients cared for in the clinical units participating in this current study. From the data it was observed that 20 patients (7.4%) had a previous episode of tuberculosis whose treatment was not completed. Eighty-one patients (29.9%) were diagnosed with HIV at tuberculosis diagnosis, 108 (39.8%) were aware of their HIV status but were not on antiretroviral treatment and 82 (30.3%) were on antiretroviral treatment. Patients on antiretroviral treatment were significantly less immunosuppressed than patients with HIV diagnosed concurrently with tuberculosis, or other patients not on antiretrovirals (median CD4 lymphocytes count: 220 cells x mm(-3) versus 100 cells x mm(-3), and 109 cells x mm(-3), respectively). No significant differences in clinical presentation of tuberculosis according to antiretroviral therapy status were recorded. Failure of tuberculosis control interventions (e.g. noncompletion of treatment) and of HIV care (delayed diagnosis of HIV infection and suboptimal uptake of therapy) may contribute to continuing occurrence of HIV associated tuberculosis in a country where highly active antiretroviral therapy is largely available. However, a significant proportion of cases occur in patients who are on antiretroviral treatment. PMID- 15293600 TI - PPAR-gamma agonists as therapy for diseases involving airway neutrophilia. AB - Peroxisome proliferator-activated receptors (PPARs) are a family of ligand activated nuclear hormone receptors belonging to the steroid receptor super family. Previously, the present authors have shown that PPAR-gamma agonists inhibit the release of inflammatory cell survival factors and induce apoptosis in vitro. The aim of this study was to determine the effect of two structurally different PPAR agonists in an in vivo model of lipopolysaccharide (LPS)-induced airway inflammation. Mice were treated with PPAR agonists, rosiglitazone or SB 219994, prior to exposure to aerosolised LPS, and the extent of airway inflammation was assessed 3 h later. In these experiments, the PPAR ligands inhibited LPS-induced airway neutrophilia and associated chemoattractants/survival factors (keratinocyte-derived chemokine and granulocyte colony stimulating factor) in the mouse lung. The present authors postulate that if a peroxisome proliferator-activated receptor agonist has the same effect in man, and neutrophils are important in the progression of respiratory diseases, such as chronic obstructive pulmonary disease, then this class of compounds could be a potential therapy. Furthermore, several peroxisome proliferator-activated receptor-gamma agonists have been shown to be clinically effective for the treatment of type II diabetes, suggesting that any benefit of peroxisome proliferator-activated receptor-gamma ligands in the progression of respiratory diseases, which may involve airway neutrophilia, could be explored relatively quickly. PMID- 15293601 TI - Clinical prognostic indicators of high-grade pre-invasive bronchial lesions. AB - Lung cancer arises from multistep genetic damage of bronchial epithelium, driving multifocal progressive dysplastic lesions. However, the risk of progression of high-grade pre-invasive bronchial lesions to cancer is poorly assessed. The purpose of this study was to better define the parameters that predict the outcome of these lesions. The current authors prospectively studied 27 patients with 31 histologically proven severe dysplasia (SD) and carcinoma in situ (CIS), with repeated bronchoscopy and endobronchial treatment. The influence of respiratory-cancer history, histopathological classification, tobacco consumption, and number of biopsies on the progression rate into cancer was studied. The actuarial progression rate to cancer was 17% at 1 yr and 63% at 3 yrs. A total of 11 cases of CIS progressed to invasive cancer, 17 were stable or regressed during the study, two with SD regressed and one progressed to invasive cancer. Progression of CIS appeared more frequent in lesions diagnosed as "questionable CIS". Persistence of smoking did not influence high-grade lesion outcome. The existence of synchronous lung cancer did not seem to impact on progression. The number of biopsies did not influence the outcome. In conclusion, the current study suggests that the outcome of high-grade pre-invasive lesions is not modified by the number of biopsies performed on these lesions. Careful pathological examination of these lesions and pathological revision seem necessary, since questionable cases have the worse progression rate. PMID- 15293603 TI - Proteasome inhibitors modulate chemokine production in lung epithelial and monocytic cells. AB - Proteasome inhibition has become a target for antitumour and anti-inflammatory therapy. The present study investigated the influence of cysteine proteinase and proteasome inhibitors on chemokine production in lung epithelial cells and monocytic cells. The lung carcinoma cell lines A549, SK-MES, NCI-H727, virus transformed bronchial epithelial cell line BEAS-2B, primary lung epithelial cells, and the acute monocytic leukaemia cell lines Mono-Mac-6 and THP-1 were incubated with proteasome (N-acetyl-L-leucyl-L-leucyl-L-norleucinal (ALLN), beta lactone) or cysteine proteinase inhibitor (L-trans-Epoxysuccinyl-Leu-3 methylbutylamide-ethyl ester) and the influence on chemokine production (interleukin-8: IL-8, monocyte chemoattractant protein-1, RANTES) was quantified at protein and mRNA levels. Inhibition of proteasome activity by ALLN and beta lactone resulted in significantly increased IL-8 secretion (5- to 22-fold). Cysteine proteinase inhibitors did not influence chemokine production. The simultaneous rise in IL-8 mRNA was caused by an increased half-life of mRNA and increased RNA synthesis. Moreover, analysis of transcription factor activation revealed induction of activator protein-1 (c-Jun) activity by proteasome inhibition, whereas nuclear factor-kappaB (p50 and p65) was not activated. The significant increase in IL-8 production after proteasome inhibition was also observed in primary lung epithelial cells and in monocytic cells. In addition, the secreted IL-8 was biologically active as shown by the neutrophil chemotaxis assay. In conclusion, it was shown that proteasome inhibitors stimulate interleukin-8 secretion in lung epithelial cells and monocytic cells, thus recruiting neutrophils. PMID- 15293602 TI - Interstitial lung disease in a baby with a de novo mutation in the SFTPC gene. AB - Mutations in the surfactant protein C gene (SFTPC) were recently reported in patients with interstitial lung disease. In a 13-month-old infant with severe respiratory insufficiency, a lung biopsy elicited combined histological patterns of nonspecific interstitial pneumonia and pulmonary alveolar proteinosis. Immunohistochemical and biochemical analyses showed an intra-alveolar accumulation of surfactant protein (SP)-A, precursors of SP-B, mature SP-B, aberrantly processed proSP-C, as well as mono- and dimeric SP-C. Sequencing of genomic DNA detected a de novo heterozygous missense mutation of the SFTPC gene (g.1286T>C) resulting in a substitution of threonine for isoleucine (173T) in the C-terminal propeptide. At the ultrastructural level, abnormal transport vesicles were detected in type-II pneumocytes. Fusion proteins, consisting of enhanced green fluorescent protein and wild-type or mutant proSP-C, were used to evaluate protein trafficking in vitro. In contrast to wild-type proSP-C, mutant proSP-C was routed to early endosomes when transfected into A549 epithelial cells. In contrast to previously reported mutations, the 173T represents a new class of surfactant protein C gene mutations, which is marked by a distinct trafficking, processing, palmitoylation, and secretion of the mutant and wild-type surfactant protein C. This report heralds the emerging diversity of phenotypes associated with the expression of mutant surfactant C proteins. PMID- 15293604 TI - CCL22 and CCL17 in rat radiation pneumonitis and in human idiopathic pulmonary fibrosis. AB - Pulmonary fibrosis is caused by various known and unknown aetiologies, but the key pathogenic mechanisms are still ill-defined. Chemokines are a large family of chemotactic cytokines that play pivotal roles in various inflammatory diseases. In the present study, the roles of chemokines in a rat model of radiation pneumonitis/ pulmonary fibrosis were examined. Accumulation of inflammatory cells and pneumonitis were observed on day 28, and diffuse alveolar wall thickening with extensive fibrosis was observed on day 56. In addition to the previously reported CCL2 (macrophage chemoattractant protein-1) induction, selective upregulation of CCL22 (macrophage-derived chemokine) and CCL17 (thymus and activation-regulated chemokine) were demonstrated for the first time in the irradiated lung tissues. Immunohistochemically, it was demonstrated that CCL22 and CCL17 were localised primarily to alveolar macrophages, whereas their receptor CC chemokine receptor 4 (CCR4) was detected on alveolar lymphocytes and macrophages. On further analysis of bronchoalveolar lavage fluid from patients with idiopathic pulmonary fibrosis and sarcoidosis, elevated levels of CCL22, but not of CCL17, were observed in the idiopathic pulmonary fibrosis patients. Since these two chemokines play pivotal roles in various type-2 T-helper cell-dominant diseases, it was speculated that CCL22, and probably CCL17, are involved in the pathophysiology of radiation pneumonitis/pulmonary fibrosis and idiopathic pulmonary fibrosis through the recruitment of CC chemokine receptor 4-positive type-2 T-helper cells and alveolar macrophages. PMID- 15293605 TI - Pirfenidone attenuates expression of HSP47 in murine bleomycin-induced pulmonary fibrosis. AB - Heat shock protein (HSP) 47, a collagen-specific molecular chaperone, is involved in the processing and/or secretion of procollagen. The present study was undertaken to investigate whether treatment with the antifibrotic drug pirfenidone attenuates the bleomycin (BL)-induced overexpression of HSP47 in the lungs. Male ICR mice were intravenously injected with BL or saline (SA). Pirfenidone or control drug (CD) was administered 14 days after commencement of BL or SA, and continued throughout the course of the experiment. The mice were randomly divided into three experimental groups: 1) SA-treated with CD (SA group); 2) BL-treated with CD (BL group); and 3) BL-treated with pirfenidone (pirfenidone group). Lungs of the pirfenidone group showed a marked reduction of fibrotic lesions compared with the corresponding BL group. Immunohistochemical studies showed that BL treatment significantly increased the number of macrophages, myofibroblasts, HSP47-positive type II pneumocytes and HSP47 positive interstitial spindle-shaped cells. Treatment with pirfenidone significantly reduced the number of these cells compared with the corresponding BL group. Furthermore, treatment with pirfenidone significantly suppressed the BL induced increase of the positive ratio of HSP47 and alpha-smooth muscle actin to interstitial spindle-shaped cells. The present study results showed that pirfenidone inhibited heat shock protein 47-positive cells and myofibroblasts, the principal cells responsible for the accumulation and deposition of extracellular matrix seen in pulmonary fibrosis. PMID- 15293606 TI - Laser-assisted uvulopalatoplasty for snoring: does it meet the expectations? AB - The high prevalence of habitual snoring (35% of the general population) and the increasing demand for an effective treatment have led, in the last decade, to the generalisation of laser-assisted uvulopalatoplasty (LAUP). However, acceptable studies on its effectiveness are lacking. The present randomised, placebo controlled study included 25 nonapnoeic and mild obstructive sleep apnoea snorers to evaluate LAUP effectiveness for snoring. Group I received a one-stage LAUP treatment and group II a placebo (simulated snore surgery followed by an oral placebo). Before each treatment and 3 months after, the variables and procedures assessed were: body weight; sleepiness (Epworth sleepiness scale); quality of life (SF-36); subjective snoring intensity (0-10 analogue scale); objective snoring intensity (average decibel intensity); snoring index (number of snores per hour); and apnoea/hypopnea index. No differences were observed in body weight, sleepiness, quality of life, subjective and objective intensity, and frequency of snoring, and apnoea/hypopnea index between the groups before and 3 months after treatment. In conclusion, this study provides evidence of the lack of effectiveness of one-stage laser-assisted uvulopalatoplasty for snoring in nonapnoeic and mild obstructive sleep apnoea patients, with the result that it does not meet the expectations generated by the procedure. PMID- 15293607 TI - Older individuals have increased oro-nasal breathing during sleep. AB - Breathing route during sleep has been studied very little, however, it has potential importance in the pathophysiology of sleep disordered breathing. Using overnight polysomnography, with separate nasal and oral thermocouple probes, data were obtained from 41 subjects (snorers and nonsnorers; 25 male and 16 female; aged 20-66 yrs). Awake, upright, inspiratory nasal resistance (Rn) was measured using posterior rhinomanometry. Each 30-s sleep epoch (not affected by apnoeas/hypopnoeas) was scored for presence of nasal and/or oral breathing. Overnight, seven subjects breathed nasally, one subject oro-nasally and the remainder switched between nasal and oro-nasal breathing. Oral-only breathing rarely occurred. Nasal breathing epochs were 55.79 (69.78) per cent of total sleep epochs (%TSE; median (interquartile range)), a value not significantly different to that for oro-nasal (TSE: 44.21 (68.66)%). Oro-nasal breathing was not related to snoring, sleep stage, posture, body mass index, height, weight, Rn (2.19 (1.77) cm H2O x L(-1) x sec(-1)) or sex, but was positively associated with age. Subjects > or = 40 yrs were approximately six times more likely than younger subjects to spend >50% of sleep epochs utilising oro-nasal breathing. Ageing is associated with an increasing occurrence of oro-nasal breathing during sleep. PMID- 15293608 TI - STAT4 activation in smokers and patients with chronic obstructive pulmonary disease. AB - Activation of the transcription factor signal transducer and activator of transcription (STAT)-4 is critical for the differentiation of T-helper 1 cells/type-1 cytotoxic T-cells and the production of interferon (IFN)-gamma. Expression of STAT4, phospho-STAT4, IFN-gamma and T-box expressed in T-cells (T bet) proteins in bronchial biopsies and bronchoalveolar lavage (BAL)-derived lymphocytes, obtained from 12 smokers with mild/moderate chronic obstructive pulmonary disease (COPD) (forced expiratory volume in one second (FEV1) 59 +/- 16% predicted), 14 smokers with normal lung function (FEV1 106 +/- 12% pred) and 12 nonsmoking subjects (FEV1 111 +/- 14% pred), was examined by immunohistochemistry and immunocytochemistry. In bronchial biopsies of COPD patients, the number of submucosal phospho-STAT4+ cells was increased (240 (22 406) versus 125 (0-492) versus 29 (0-511) cells mm(-2)) when compared with both healthy smokers and control nonsmokers, respectively. In smokers, phospho-STAT4+ cells correlated with the degree of airflow obstruction and the number of IFN gamma+ cells. Similar results were seen in BAL (2.8 (0.2-5.9) versus 1.03 (0.09 1.6) versus 0.69 (0-2.3) lymphocytes x mL(-1) x 10(3)). In all smokers who underwent lavage, phospho-STAT4+ lymphocytes correlated with airflow obstruction and the number of IFNgamma+ lymphocytes. T-bet expression was not altered in bronchial biopsies and BAL-derived lymphocytes between the three groups. In conclusion, this study suggests that stable mild/moderate chronic obstructive pulmonary disease is associated with an active T-helper 1 cell/type-1 cytotoxic T cell inflammatory process involving activation of signal transducer and activator of transcription 4 and interferon-gamma production. PMID- 15293609 TI - Effect of salmeterol on the ventilatory response to exercise in chronic obstructive pulmonary disease. AB - This study examined the effects of bronchodilator-induced reductions in lung hyperinflation on breathing pattern, ventilation and dyspnoea during exercise in chronic obstructive pulmonary disease (COPD). Quantitative tidal flow/volume loop analysis was used to evaluate abnormalities in dynamic ventilatory mechanics and their manipulation by a bronchodilator. In a randomised double-blind crossover study, 23 patients with COPD (mean +/- SEM forced expiratory volume in one second 42 +/- 3% of the predicted value) inhaled salmeterol 50 microg or placebo twice daily for 2 weeks each. After each treatment period, 2 h after dose, patients performed pulmonary function tests and symptom-limited cycle exercise at 75% of their maximal work-rate. After salmeterol versus placebo at rest, volume corrected maximal expiratory flow rates increased by 175 +/- 52%, inspiratory capacity (IC) increased by 11 +/- 2% pred and functional residual capacity decreased by 11 +/- 3% pred. At a standardised time during exercise, salmeterol increased IC, tidal volume (VT), mean inspiratory and expiratory flows, ventilation, oxygen uptake (VO2) and carbon dioxide output. Salmeterol increased peak exercise endurance, VO2 and ventilation by 58 +/- 19, 8 +/- 3 and 12 +/- 3%, respectively. Improvements in peak VO2 correlated best with increases in peak VT; increases in peak VT and resting IC were interrelated. The reduction in dyspnoea ratings at a standardised time correlated with the increased VT. Mechanical factors play an important role in shaping the ventilatory response to exercise in chronic obstructive pulmonary disease. Bronchodilator-induced lung deflation reduced mechanical restriction, increased ventilatory capacity and decreased respiratory discomfort, thereby increasing exercise endurance. PMID- 15293610 TI - Normal nasal mucociliary clearance in CF children: evidence against a CFTR related defect. AB - Studies on mucociliary clearance (MCC) in cystic fibrosis (CF) have produced conflicting results. This study aimed to differentiate primary (ion transport related) from secondary (inflammatory) causes of delayed MCC in CF. Nasal MCC was measured in 50 children (CF, primary ciliary dyskinesia (PCD) and no respiratory disease). Nasal lavage fluid was analysed for interleukin (IL)-8 and tumour necrosis factor-alpha. Similar measurements were obtained in adult CF patients with and without chronic sinusitis (CS). Children with CF had neither delayed MCC nor increased levels of cytokines. Conversely, children with PCD had prolonged MCC times (all >30 min) and significantly raised levels of IL-8. CS-positive CF adults had significantly slower MCC than CS-negative subjects, but IL-8 levels were low and similar in both groups. Decreased airway surface liquid and delayed mucociliary clearance are the postulated primary mechanisms in cystic fibrosis. However, the current study reports that cystic fibrosis children have normal nasal mucociliary clearance. Abnormalities appeared in cystic fibrosis adults with symptoms of chronic sinus disease, suggesting a secondary rather than primary phenomenon. Studies to explore this mechanism in the distal, more sparsely-ciliated airways could aid an understanding of pathogenesis and the development of new treatments. PMID- 15293611 TI - Clonal strains of Pseudomonas aeruginosa in paediatric and adult cystic fibrosis units. AB - Despite recent reports of clonal strains of Pseudomonas aeruginosa in cystic fibrosis (CF) units, the need for routine microbiological surveillance remains contentious. Sputum was collected prospectively from productive patients attending the regional paediatric and adult CF units in Brisbane, Australia. All P. aeruginosa isolates were typed using pulsed-field gel electrophoresis. Spirometry, anthropometrics, hospitalisations and antibiotic sensitivity data were recorded. The first 100 sputum samples (first 50 patients at each clinic) harboured 163 isolates of P. aeruginosa. A total of 39 patients shared a common strain (pulsotype 2), 20 patients shared a strain with at least one other patient and 41 patients harboured unique strains. Eight patients shared a strain identical to a previously reported Australian transmissible strain (pulsotype 1). Compared with the unique strain group, patients harbouring pulsotype 2 were younger and had poorer lung function. Treatment requirements were similar in these two groups, as were the rates of multiresistance. In conclusion, 59% of patients harboured a clonal strain, supporting the need for routine microbiological surveillance. In contrast to previously described clonal strains, the dominant pulsotype was indistinguishable from nonclonal strains with respect to both colonial morphology and multiresistance. The clinical significance of clonal strains remains uncertain and requires longitudinal study. PMID- 15293612 TI - Increase in laminin expression in allergic airway remodelling and decrease by dexamethasone. AB - Lung expression of the extracellular matrix protein, laminin, and its receptor, laminin-1 receptor, were examined in a mouse model of asthma with airway remodelling. Ovalbumin (OVA) was administered to BALB/c mice, intraperitoneally on days 0 and 14, and intranasally periodically between days 14 and 75. The mice developed airway eosinophil and mononuclear inflammatory cell infiltration and fibrosis. On day 76, a marked increase in total laminin was seen in the airways of OVA-treated mice compared to controls by Western blot analysis. The increased laminin expression was detected immunocytochemically in the thickened subepithelial basement membrane and around airways and blood vessels. The OVA treated mice showed increased expression of the alpha1, beta1, and gamma1 chains of the laminin-1 isoform in monocytes, macrophages and eosinophils infiltrating the airways. Laminin-1 receptor expression was increased in inflammatory and endothelial cells in the lungs of OVA-treated mice compared to controls. Treatment of OVA-sensitised/challenged mice with dexamethasone reduced airway expression of laminin and laminin-1 receptor in OVA-treated mice but not airway hyperresponsiveness to methacholine. Laminin deposition may be an important component of the airway remodelling observed in chronic allergic lung inflammation and is a process modulated by corticosteroids. PMID- 15293613 TI - Obesity and nocturnal gastro-oesophageal reflux are related to onset of asthma and respiratory symptoms. AB - Several studies have identified obesity as a risk factor for asthma in both children and adults. An increased prevalence of asthma in subjects with gastro oesophageal reflux (GOR) and obstructive sleep apnoea syndrome has also been reported. The aim of this investigation was to study obesity, nocturnal GOR and snoring as independent risk factors for onset of asthma and respiratory symptoms in a Nordic population. In a 5-10 yr follow-up study of the European Community Respiratory Health Survey in Iceland, Norway, Denmark, Sweden and Estonia, a postal questionnaire was sent to previous respondents. A total of 16,191 participants responded to the questionnaire. Reported onset of asthma, wheeze and night-time symptoms as well as nocturnal GOR and habitual snoring increased in prevalence along with the increase in body mass index (BMI). After adjusting for nocturnal GOR, habitual snoring and other confounders, obesity (BMI >30) remained significantly related to the onset of asthma, wheeze and night-time symptoms. Nocturnal GOR was independently related to the onset of asthma and in addition, both nocturnal GOR and habitual snoring were independently related to onset of wheeze and night-time symptoms. This study adds evidence to an independent relationship between obesity, nocturnal gastro-oesophageal reflux and habitual snoring and the onset of asthma and respiratory symptoms in adults. PMID- 15293614 TI - Parameters associated with persistent airflow obstruction in chronic severe asthma. AB - The significance of severe airflow obstruction in severe asthma is unclear. The current study determined whether severe airflow obstruction is related to inflammatory or structural changes in the airways. Patients with severe asthma from a tertiary referral clinic were divided into two groups according to their postbronchodilator forced expiratory volume in one second (FEV1): severe persistent airflow limitation (FEV1 <50% predicted; group S; n=37) and no obstruction (FEV1 >80% pred; group N; n=29). Smoking history, atopic status, lung function tests, exhaled NO, blood eosinophil count, quality of life scores using St George's Respiratory Questionnaire and high resolution computed tomography (HRCT) of the lungs were assessed. Patients from group S were older and had longer disease duration. There was no difference in smoking history, atopic status, hospital admissions, quality of life scores and amount of treatment with inhaled or oral corticosteroids. Exhaled NO and peripheral blood eosinophils were higher in group S (21.0 +/- 2.4 versus 12.8 +/- 2.3 ppb; 0.41 +/- 0.06 versus 0.15 +/- 0.03 x 10(9) cells x L(-1) respectively). HRCT scores for bronchial wall thickening and dilatation were higher in group S with no differences in air trapping. Peripheral blood eosinophilia and bronchial wall thickening on HRCT scan were the only parameters significantly and independently associated with persistent airflow obstruction. Patients with severe asthma and irreversible airflow obstruction had longer disease duration, a greater inflammatory process and more high resolution computed tomography airway abnormalities suggestive of airway remodelling, despite being on similar treatments and experiencing equivalent impairment in quality of life. PMID- 15293615 TI - Relevance of assessing quadriceps endurance in patients with COPD. AB - The aims of this study were to investigate whether the impairment in endurance of limb muscles is a general finding in chronic obstructive pulmonary disease (COPD) patients, affecting even those with mild-to-moderate disease or relatively normal physical activity. In addition, this study aimed to determine the physiopathology of exhaustion in local endurance tests and whether the reduction in quadriceps endurance can be predicted from muscle strength measurements. A total of 75 volunteers were assigned to one of two groups according to pulmonary function tests: COPD patients or healthy age-matched controls. Functional assessment included both quadriceps strength (maximum voluntary contraction (QMVC)), and quadriceps endurance (contractions against a load equivalent to 10% QMVC until task failure or for up to a limiting time of 30 min (QTlim)). COPD patients showed a decrease of approximately 43%, in QMVC and approximately 77% in QTlim compared with controls. Task failure occurred only in COPD patients and was due to muscle fatigue, since limiting symptoms were associated with a decrease in the median frequency of quadriceps electromyographical signal and a reversible decrease in QMVC. The impairment in skeletal muscle endurance was present even in patients with mild-to-moderate airflow obstruction and individuals with relatively normal physical activity, and was irrespective of lung function variables, anthropometrical data or quadriceps strength. Peripheral muscle endurance was impaired in chronic obstructive pulmonary disease patients, even in those with relatively normal physical activity and mild-to-moderate airflow obstruction. This impairment associated with an early onset of muscle fatigue and could not be predicted from the severity of the disease or the reduction in quadriceps strength. PMID- 15293616 TI - Acute effect of oral steroids on muscle function in chronic obstructive pulmonary disease. AB - Prospective data to support the hypothesis that corticosteroids are a significant cause of muscle weakness in patients with chronic obstructive pulmonary disease (COPD) are lacking. The authors studied respiratory and quadriceps muscle function, using both volitional techniques and magnetic nerve stimulation, as well as measuring metabolic parameters during incremental cycle ergometry, in 25 stable COPD patients. The forced expiratory volume in one second was 37.6 +/- 21.4% predicted, before and after a 2-week course of o.d. prednisolone 30 mg. Quadriceps strength was also assessed in 15 control patients on two occasions. Only two patients met the British Thoracic Society definition of steroid responsiveness. There was no change either in sniff transdiaphragmatic pressure (pre: 96.8 +/- 19.7 cmH2O; post: 98.6 +/- 22.4 cmH2O) or in twitch transdiaphragmatic pressure elicited by bilateral anterolateral magnetic phrenic nerve stimulation (pre: 16.8 +/- 9.1 cmH2O; post: 17.9 +/- 10 cmH2O). Quadriceps twitch force did not change significantly either in the steroid group (pre: 9.5 +/- 3.1 kg; post: 8.9 +/- 3.7 kg) or in the control patients (pre: 8.1 +/- 2.7 kg; post: 7.9 +/- 2.2 kg). There were no changes in either peak or isotime ventilatory and metabolic parameters during exercise. In conclusion, in stable patients with chronic obstructive pulmonary disease, a 2-week course of 30 mg prednisolone daily does not cause significant skeletal muscle dysfunction or alter metabolic parameters during exercise. PMID- 15293617 TI - Immunokinetics in severe pneumonia due to influenza virus and bacteria coinfection in mice. AB - Coinfections of bacteria and influenza are a major cause of excessive mortality during influenza epidemics. However, the mechanism of the synergy between influenza virus and bacteria are poorly understood. In this study, mice were inoculated with influenza virus, followed 2 days later by inoculation with Streptococcus pneumoniae. The kinetics of viral titres, bacterial numbers and the immune response (cytokine and chemokine production) were also analysed. Short term survival correlated with pathological changes in the lungs of infected mice. Influenza virus or S. pneumoniae infection alone induced moderate pneumonia; however, severe bronchopneumonia with massive haemorrhage in coinfected mice, which caused death of these mice approximately 2 days after inoculation with S. pneumoniae, was noted. Intrapulmonary levels of inflammatory cytokines/chemokines, type-1 T-helper cell cytokines and Toll-like receptors, and the related mitogen-activated protein kinase signalling molecules (phosphorylated extracellular signal-regulated kinase -1 and - 2, p38 and c-Jun N-terminal kinase), were increased in coinfected mice. These results suggest that immune mediators, including cytokines and chemokines, through Toll-like receptors/mitogen-activated protein kinase pathways, play important roles in the pathology of coinfection caused by influenza virus and Streptococcus pneumoniae. PMID- 15293618 TI - Comparison of 99mTc-DTPA and urea for measuring cefepime concentrations in epithelial lining fluid. AB - The efficacy of antimicrobial agents against pulmonary infections depends on their local concentrations in the lung. The aims of the present study were to: 1) compare technetium-99m diethylenetriaminepenta-acetic acid (99mTc-DTPA) and urea as markers of epithelial lining fluid (ELF) dilution for measuring ELF concentrations of pharmaceuticals; 2) quantify ELF cefepime concentrations in normal and injured lung; and 3) measure the increase in permeability to cefepime following oleic acid-induced acute lung injury. A modified bronchoalveolar lavage technique, based on equilibration of infused 99mTc-DTPA, was used to measure ELF volume. Cefepime was administered intravenously at steady plasma levels. Six serial bronchoalveolar lavages were performed 5 h after the beginning of infusion. ELF to plasma cefepime concentration ratios were 95 +/- 17 and 100 +/- 14.5% in normal and injured lung respectively. When urea was used as marker, cefepime concentration ratios were underestimated at 16.4 +/- 2.7 and 73.9 +/- 8.4% respectively. Cefepime blood/ airspace clearance increased from 3.8 +/- 0.7 micro x min(-1) in controls to 39.8 +/- 4.9 microL x min(-1) in acute lung injury. It was concluded that: 1) cefepime concentrations in epithelial lining fluid were in equilibrium with those in plasma in both normal and injured lung after 5 h at steady plasma concentrations; 2) epithelial lining fluid cefepime concentration by the urea method was much less underestimated in injured versus normal lung; and 3) acute lung injury induces a 10-fold elevation of cefepime blood/airspace clearance. PMID- 15293619 TI - Area deprivation predicts lung function independently of education and social class. AB - The cross-sectional association between socioeconomic status (at both the individual and area-based level) and lung function, as measured by forced expiratory volume in one second, in a large population-based cohort was investigated. The study population consisted of 22,675 males and females aged 39 79 yrs. They were recruited from the general community in Norfolk, UK using general practice age/sex registers, as part of the European Prospective Investigation into Cancer (EPIC-Norfolk). It was found that being in a manual occupational social class, having no educational qualifications and living in a deprived area all independently predicted significantly lower lung function, even after controlling for smoking habit. The influence of area-deprivation on lung function, independent of individual socioeconomic status and of individual smoking habit, suggests that apart from targeting individuals who are at high risk, such as smokers, environmental determinants also need to be examined when considering measures to improve respiratory health. PMID- 15293620 TI - Idiopathic pulmonary haemosiderosis revisited. AB - Idiopathic pulmonary haemosiderosis is a rare cause of diffuse alveolar haemorrhage of unknown aetiology. It occurs most frequently in children, has a variable natural history with repetitive episodes of diffuse alveolar haemorrhage, and has been reported to have a high mortality. Many patients develop iron deficiency anaemia secondary to deposition of haemosiderin iron in the alveoli. Examination of sputum and bronchoalveolar lavage fluid can disclose haemosiderin-laden alveolar macrophages (siderophages), and the lung biopsy shows numerous siderophages in the alveoli, without any evidence of pulmonary vasculitis, nonspecific/granulomatous inflammation, or deposition of immunoglobulins. Contrary to earlier reports, corticosteroids alone or in combination with other immunosuppressive agents may be effective for either exacerbations or maintenance therapy of idiopathic pulmonary haemosiderosis. PMID- 15293621 TI - Atypical pathogens and respiratory tract infections. AB - The atypical respiratory pathogens Chlamydia pneumoniae, Mycoplasma pneumoniae and Legionella pneumophila are now recognised as a significant cause of acute respiratory-tract infections, implicated in community-acquired pneumonia, acute exacerbations of chronic bronchitis, asthma, and less frequently, upper respiratory-tract infections. Chronic infection with C. pneumoniae is common among patients with chronic obstructive pulmonary disease and may also play a role in the natural history of asthma, including exacerbations. The lack of a gold standard for diagnosis of these pathogens still handicaps the current understanding of their true prevalence and role in the pathogenesis of acute and chronic respiratory infections. While molecular diagnostic techniques, such as polymerase chain reaction, offer improvements in sensitivity, specificity and rapidity over culture and serology, the need remains for a consistent and reproducible diagnostic technique, available to all microbiology laboratories. Current treatment guidelines for community-acquired pneumonia recognise the importance of atypical respiratory pathogens in its aetiology, for which macrolides are considered suitable first-line agents. The value of atypical coverage in antibiotic therapy for acute exacerbations of chronic bronchitis and exacerbations of asthma is less clear, while there is no evidence to suggest that atypical pathogens should be covered in antibiotic treatment of upper respiratory tract infections. PMID- 15293622 TI - Cavitary lung masses in SLE patients: an unusual manifestation of CMV infection. AB - The typical radiographical findings of cytomegalovirus pneumonitis are bilateral interstitial infiltrates. In this study, the current authors describe two patients on corticosteroid treatment for systemic lupus erythematosus, complicated by histologically confirmed cytomegalovirus pneumonitis, presenting as cavitary masses. This rare presentation of cytomegalovirus pneumonitis broadens the differential diagnosis of cavitary lesions to include cytomegalovirus infection in immunocompromised individuals. PMID- 15293623 TI - A 13-yr-old male with fever, malaise, body pains and dry cough. PMID- 15293624 TI - Ireland needs healthier airways and lungs: the evidence (INHALE). PMID- 15293625 TI - Exhaled breath condensate: a space odessey, where no one has gone before.. PMID- 15293626 TI - Inhaled steroids and mortality in COPD: bias from unaccounted immortal time. PMID- 15293627 TI - Two-sample statistics for testing the equality of survival functions against improper semi-parametric accelerated failure time alternatives: an application to the analysis of a breast cancer clinical trial. AB - This paper presents two-sample statistics suited for testing equality of survival functions against improper semi-parametric accelerated failure time alternatives. These tests are designed for comparing either the short- or the long-term effect of a prognostic factor, or both. These statistics are obtained as partial likelihood score statistics from a time-dependent Cox model. As a consequence, the proposed tests can be very easily implemented using widely available software. A breast cancer clinical trial is presented as an example to demonstrate the utility of the proposed tests. PMID- 15293628 TI - Marginal means/rates models for multiple type recurrent event data. AB - Recurrent events are frequently observed in biomedical studies, and often more than one type of event is of interest. Follow-up time may be censored due to loss to follow-up or administrative censoring. We propose a class of semi-parametric marginal means/rates models, with a general relative risk form, for assessing the effect of covariates on the censored event processes of interest. We formulate estimating equations for the model parameters, and examine asymptotic properties of the parameter estimators. Finite sample properties of the regression coefficients are examined through simulations. The proposed methods are applied to a retrospective cohort study of risk factors for preschool asthma. PMID- 15293629 TI - Tests for the proportional intensity assumption based on the score process. AB - Proportional intensity models are widely used for describing the relationship between the intensity of a counting process and associated covariates. A basic assumption in this model is the proportionality, that each covariate has a multiplicative effect on the intensity. We present and study tests for this assumption based on a score process which is equivalent to cumulative sums of the Schoenfeld residuals. Tests within principle power against any type of departure from proportionality can be constructed based on this score process. Among the tests studied, in particular an Anderson-Darling type test turns out to be very useful by having good power properties against general alternatives. A simulation study comparing various tests for proportionality indicates that this test seems to be a good choice for an omnibus test for proportionality. PMID- 15293630 TI - Bayesian estimation of survival functions under stochastic precedence. AB - When estimating the distributions of two random variables, X and Y, investigators often have prior information that Y tends to be bigger than X. To formalize this prior belief, one could potentially assume stochastic ordering between X and Y, which implies Pr(X < or = z) > or = Pr(Y < or = z) for all z in the domain of X and Y. Stochastic ordering is quite restrictive, though, and this article focuses instead on Bayesian estimation of the distribution functions of X and Y under the weaker stochastic precedence constraint, Pr(X < or = Y) > or = 0.5. We consider the case where both X and Y are categorical variables with common support and develop a Gibbs sampling algorithm for posterior computation. The method is then generalized to the case where X and Y are survival times. The proposed approach is illustrated using data on survival after tumor removal for patients with malignant melanoma. PMID- 15293631 TI - Estimating marginal effects in accelerated failure time models for serial sojourn times among repeated events. AB - Recurrent event data are commonly encountered in longitudinal studies when events occur repeatedly over time for each study subject. An accelerated failure time (AFT) model on the sojourn time between recurrent events is considered in this article. This model assumes that the covariate effect and the subject-specific frailty are additive on the logarithm of sojourn time, and the covariate effect maintains the same over distinct episodes, while the distributions of the frailty and the random error in the model are unspecified. With the ordinal nature of recurrent events, two scale transformations of the sojourn times are derived to construct semiparametric methods of log-rank type for estimating the marginal covariate effects in the model. The proposed estimation approaches/inference procedures also can be extended to the bivariate events, which alternate themselves over time. Examples and comparisons are presented to illustrate the performance of the proposed methods. PMID- 15293632 TI - Inference from accelerated degradation and failure data based on Gaussian process models. AB - An important problem in reliability and survival analysis is that of modeling degradation together with any observed failures in a life test. Here, based on a continuous cumulative damage approach with a Gaussian process describing degradation, a general accelerated test model is presented in which failure times and degradation measures can be combined for inference about system lifetime. Some specific models when the drift of the Gaussian process depends on the acceleration variable are discussed in detail. Illustrative examples using simulated data as well as degradation data observed in carbon-film resistors are presented. PMID- 15293633 TI - Evaluation of a vocal mand assessment and vocal mand training procedures. AB - A common deficiency in the verbal repertoires of individuals with autism and related disorders is the absence of socially appropriate vocal mands. The vocal mand repertoires of these individuals may be lacking in several respects: (a) The individual might engage in no mands whatsoever, (b) the mand might be topographically dissimilar to an appropriate response, (c) the mand might be only partially topographically similar to an appropriate response, and (d) the mand might occur only after prompting. Depending on specific deficiencies in an individual's repertoire, different procedures for establishing appropriate mands may be needed. The purpose of Study 1 was to evaluate an assessment prior to teaching vocal mands for 3 individuals with developmental disabilities. The assessment showed that 1 individual displayed partial utterances of mands, 1 displayed vocal mands after mands had been reinforced, and 1 displayed vocal mands when prompted. Thus, in Study 2, a different teaching strategy was tested for each individual. Results showed that the assessment information could be linked directly to mand training for all 3 participants. PMID- 15293634 TI - An assessment of antecedent events influencing noncompliance in an outpatient clinic. AB - Several studies have shown that various factors can influence noncompliance, including task novelty, rate of presentation, and task preference. This study examined the impact of selected antecedent variables on noncompliance in an outpatient clinic setting. In two experiments involving 6 typically developing children, the consequences for noncompliance remained constant. During Experiment 1, demands that included noncontingent access to adult attention were contrasted with the same demands that did not include attention within a multielement design. In Experiment 2, demands were altered by decreasing the difficulty or amount of work or providing access to attention. In both experiments, results indicated idiosyncratic responses to the manipulated variables, with decreases in noncompliance observed following introduction of one or more antecedent variables with 5 of the 6 participants. These results suggested that noncompliance can be reduced via changes in antecedent variables, including adding potential positive reinforcers to the task situation, and that it is possible to probe variables that alter noncompliance in an outpatient clinic setting. PMID- 15293635 TI - Increasing variety of foods consumed by blending nonpreferred foods into preferred foods. AB - A treatment with differential or noncontingent reinforcement and nonremoval of the spoon increased the acceptance of one or two of 16 foods for 2 participants with severe food refusal. These differential levels of acceptance were demonstrated empirically in an ABAB design in which A was the presentation of the accepted (preferred) foods and B was the presentation of foods the participants refused (nonpreferred foods). Subsequently, we implemented a blending treatment that consisted of mixing (blending) nonpreferred foods into preferred foods in various ratios (e.g., 10% nonpreferred/90% preferred, 20% nonpreferred/80% preferred). We then presented nonpreferred foods that had been exposed to blending to determine if consumption of nonpreferred foods would increase following the blending treatment. We also conducted periodic reversals in which we presented nonpreferred foods that had not been exposed to the blending treatment. Following initial implementation of the blending treatment, consumption was high for nonpreferred foods that had been blended and low for nonpreferred foods that had not been blended. Consumption increased for all foods (i.e., foods that had been exposed to blending and foods that had not been exposed to blending) after seven or eight foods had been exposed to the blending treatment. Thus, the variety of foods consumed by the participants increased from one or two to 16. These results are discussed in terms of stimulus fading, conditioned food preferences, and escape extinction. DESCRIPTORS: conditioned food preferences, food refusal, negative reinforcement, stimulus fading PMID- 15293636 TI - Enhancing the effects of extinction on attention-maintained behavior through noncontingent delivery of attention or stimuli identified via a competing stimulus assessment. AB - Recent research has shown that the noncontingent delivery of competing stimuli can effectively reduce rates of destructive behavior maintained by social positive reinforcement, even when the contingency for destructive behavior remains intact. It may be useful, therefore, to have a systematic means for predicting which reinforcers do and do not compete successfully with the reinforcer that is maintaining destructive behavior. In the present study, we conducted a brief competing stimulus assessment in which noncontingent access to a variety of tangible stimuli (one toy per trial) was superimposed on a fixed ratio 1 schedule of attention for destructive behavior for individuals whose behavior was found to be reinforced by attention during a functional analysis. Tangible stimuli that resulted in the lowest rates of destructive behavior and highest percentages of engagement during the competing stimulus assessment were subsequently used in a noncontingent tangible items plus extinction treatment package and were compared to noncontingent attention plus extinction and extinction alone. Results indicated that both treatments resulted in greater reductions in the target behavior than did extinction alone and suggested that the competing stimulus assessment may be helpful in predicting stimuli that can enhance the effects of extinction when noncontingent attention is unavailable. DESCRIPTORS: Attention-maintained problem behavior, competing stimuli, extinction, functional analysis, noncontingent reinforcement PMID- 15293637 TI - Promoting response variability and stimulus generalization in martial arts training. AB - The effects of reinforcement and extinction on response variability and stimulus generalization in the punching and kicking techniques of 2 martial arts students were evaluated across drill and sparring conditions. During both conditions, the students were asked to demonstrate different techniques in response to an instructor's punching attack. During baseline, the students received no feedback on their responses in either condition. During the intervention phase, the students received differential reinforcement in the form of instructor feedback for each different punching or kicking technique they performed during a session of the drill condition, but no reinforcement was provided for techniques in the sparring condition. Results showed that both students increased the number of different techniques they performed when reinforcement and extinction procedures were conducted during the drill condition, and that this increase in response variability generalized to the sparring condition. PMID- 15293638 TI - Increasing pretend toy play of toddlers with disabilities in an inclusive setting. AB - We evaluated a program for increasing pretend toy play of 2-year-old children with disabilities in an inclusive classroom. Classroom personnel implemented the program, which involved choices of classroom centers containing toys that tend to occasion pretend play in toddlers without disabilities, along with prompting and praise. Increases occurred in independent pretend-play rates among all 5 participating toddlers. Results are discussed regarding the importance of promoting toy play of very young children with disabilities that is similar to the type of play of their nondisabled peers, and the need to identify critical program components that are applicable in inclusive settings. DESCRIPTORS: pretend play, young children with disabilities PMID- 15293639 TI - Using paired-choice assessment to identify variables maintaining sleep problems in a child with severe disabilities. AB - In this study, we used a paired-choice assessment protocol to identify the relative reinforcing value of stimuli and activities for a child with severe disabilities when she failed to settle to sleep at night. The results of this assessment indicated that the child preferred the mother's attention relative to other activities presented. Assessment results were incorporated into an intervention, that produced a reduction in sleep disturbance that was maintained at a 12-month follow up. PMID- 15293640 TI - An alternative method of thinning reinforcer delivery during differential reinforcement. AB - Differential reinforcement of alternative behavior (DRA) may result in rates of reinforcement that are impractical for caregivers to implement; therefore, recent research has examined methods for thinning reinforcer delivery during DRA. In this study, reinforcer delivery was thinned during DRA by restricting access to the participant's alternative response materials. PMID- 15293641 TI - Use of response cards with a group of students with learning disabilities including those for whom English is a second language. AB - The current study compared the effects of hand raising and response cards during a writing instruction class in a middle-school resource classroom with students who were learning English as their second language. Response cards increased the rate and accuracy of academic responding, increased weekly quiz scores, and had mixed effects on off-task behavior, but most students reported that they preferred hand raising. PMID- 15293642 TI - Effects of delayed constructed-response identity matching on spelling of dictated words. AB - We studied the effects of delayed constructed-response identity matching on spelling with 6 first graders with histories of school failure. After training, the children learned to spell words to dictation and their cursive writing improved. These results replicate studies showing that delayed constructed response matching establishes spelling. For 2 children, spelling of generalization words--words formed by recombining the syllables of training words -also improved. These results extend studies that have shown recombinative generalization in reading and spelling. PMID- 15293643 TI - Functional analysis and treatment of elopement for a child with attention deficit hyperactivity disorder. AB - We conducted a functional analysis of elopement in an outdoor setting for a child with a diagnosis of attention deficit hyperactivity disorder. A subsequent treatment consisting of noncontingent attention and time-out was demonstrated to be effective in eliminating elopement. Modifications of functional analysis procedures associated with the occurrence of elopement in a natural setting are demonstrated. PMID- 15293644 TI - An evaluation of in vivo desensitization and video modeling to increase compliance with dental procedures in persons with mental retardation. AB - Fear of dental procedures deters many individuals with mental retardation from accepting dental treatment. This study was conducted to assess the effectiveness of two procedures, in vivo desensitization and video modeling, for increasing compliance with dental procedures in participants with severe or profound mental retardation. Desensitization increased compliance for all 5 participants, whereas video modeling increased compliance for only 1 of 3 participants. PMID- 15293645 TI - Obtained versus programmed reinforcement practical considerations in the treatment of escape-reinforced aggression. AB - This investigation provides a preliminary examination of the difference between programmed and obtained reinforcement rates and its potential influence during treatment of aggression in a natural setting. Following a functional analysis that suggested that the aggression of a boy with autism was negatively reinforced, intervention was implemented by the boy's mother. Concurrent fixed ratio (FR) 1 FR 1 schedules of escape were arranged for manding and aggression. When mands failed to compete effectively with aggression, obtained reinforcement ratios were calculated; these indicated that obtained reinforcement varied from the programmed schedule for aggression but not for mands. Increasing the rate of prompts for mands resulted in an increase in mands and a decrease in aggression to near-zero levels. PMID- 15293646 TI - Developing culturally effective family-based research programs: implications for family therapists. AB - Recently, some family scholars have developed greater sensitivity to the relative neglect of families of color in clinical and empirical research. Consequently, a proliferation of research elucidating many nuances of ethnic families has come to the forefront, containing a wealth of knowledge with useful implications for family therapists and other mental health providers. The findings of these studies hold enormously important implications for how family therapists can better engage and accommodate families of color in therapy. In this article we discuss some of the etiological and methodological issues associated with planning, conducting, and disseminating family-based prevention and intervention research programs with ethnic minority families. PMID- 15293647 TI - Partnering with community stakeholders: engaging rural African American families in basic research and The Strong African American Families Preventive Intervention Program. AB - The Center for Family Research has implemented the first family-community preventive intervention program designed specifically for rural African American families and youths. Basic information garnered during a decade of research in rural African American communities formed the theoretical and empirical foundations for the program, which focuses on delaying the onset of sexual activity and discouraging substance use among youths. The Center's researchers have formulated future directions for engaging rural families in basic research and preventive intervention programs. PMID- 15293648 TI - A program of research with Hispanic and African American families: three decades of intervention development and testing influenced by the changing cultural context of Miami. AB - In this article we summarize work with poor, inner-city Hispanic and African American families conducted at the University of Miami Center for Family Studies. We elucidate ways in which this research program has paralleled the treatment development paradigm and has been responsive to changes in local demographics. Specific cultural issues pertaining to Hispanics and African Americans are discussed in light of treatment development and implementation. Future directions and challenges for working with poor, inner-city minority families are addressed. PMID- 15293649 TI - Treating intimate partner violence within intact couple relationships: outcomes of multi-couple versus individual couple therapy. AB - An experimental design was used to determine outcomes of a domestic violence focused treatment program for couples that choose to stay together after mild-to moderate violence has occurred. Forty-two couples were randomly assigned to either individual couple or multi-couple group treatment. Nine couples served as the comparison group. Male violence recidivism rates 6 months after treatment were significantly lower for the multi-couple group (25%) than for the comparison group (66%). In contrast, men in the individual couple condition were not significantly less likely to recidivate (43%) than those in the comparison group. In addition, marital satisfaction increased significantly, and both marital aggression and acceptance of wife battering decreased significantly among individuals who participated in multi-couple group therapy, but not among those who participated in individual couple therapy or the comparison group. PMID- 15293650 TI - Enacting relationships in marriage and family therapy: a conceptual and operational definition of an enactment. AB - Enactments are a potential common clinical process factor contributing to positive outcomes in many relational therapies. Enactments provide therapists a medium for mediating relationships through simultaneous experiential intervention and change at multiple levels of relationships--including specific relationship disagreements and problems, interaction process surrounding these issues, and underlying emotions and attachment issues confounded with those problems. We propose a model of enactments in marriage and family therapy, consisting of three components--initiation operations, intervention operations, and evaluation operations. We offer a conceptual framework to help clinicians know when and to what purpose to use this model of enactments. We provide an operational description of each component of an enactment, exemplifying them using a hypothetical clinical vignette. Directions for future research are suggested. PMID- 15293651 TI - Moving beyond the blame game: toward a discursive approach to negotiating conflict within couple relationships. AB - The concept of discourse is an important tool in negotiating conflict and facilitating conversations within therapy. This article offers a useful framework for negotiating conflict in a couple relationship by highlighting the manner in which individual's expectations are mutually emergent from particular discursive positions. Specific discursive practices and approaches that make more visible the cultural production of conflicts are presented via a case illustration. These practices provide more freedom to couples in relationships to explore conflicts with less totalizing descriptions of the other as blameworthy. In addition, a discursive analysis of conflict invites therapists to be more intentional, reflexive, and socially responsible in their work. PMID- 15293652 TI - Solution-focused premarital counseling: helping couples build a vision for their marriage. AB - This article outlines a solution-focused approach to premarital counseling. Solution-focused premarital counseling is a strength-based approach that focuses on a couple's resources to develop a shared vision for the marriage. Background information about premarital counseling and solution-focused therapy provide the framework for the development of intervention strategies that are grounded in the solution-focused approach. Solution-oriented interventions include solution oriented questions, providing feedback, and the Couple's Resource Map, an original intervention that is described in this article. PMID- 15293653 TI - Do general treatment guidelines for Asian American families have applications to specific ethnic groups? The case of culturally-competent therapy with Korean Americans. AB - To serve Korean American families effectively, marriage and family therapists need to develop a level of cultural competence. This content analysis of the relevant treatment literature was conducted to discover the most common expert recommendations for family therapy with Asian Americans and to examine their application to Korean Americans. Eleven specific guidelines were generated: Assess support systems, assess immigration history establish professional credibility, provide role induction, facilitate "saving face," accept somatic complaints, be present/problem focused, be directive, respect family structure, be nonconfrontational, and provide positive reframes. Empirical support (clinical and nonclinical research) and conceptual support for each guideline are discussed, and conclusions are reached regarding culturally competent therapy with Korean American families. PMID- 15293654 TI - Student-faculty perceptions of multicultural training in accredited marriage and family therapy programs in relation to students' self-reported competence. AB - Although the marriage and family therapy field's recent attention to multicultural issues is laudable, there appears to be little clarity on what constitutes an effective multicultural training program and the impact of the effects of such training on trainee multicultural competence. The field continues to be challenged at different levels-training, practice, research, the setting of the standards and the work of the Commission on Accreditation for Marriage and Family Therapy Education, and the goals and strategic plan of the American Association for Marriage and Family Therapy Board. This study focused on assessing the extent of multicultural integration at different levels of training and the relationship between such training and students' perception of their own multicultural competence. PMID- 15293655 TI - Process drift: preventing the adulteration of management methods in clinical practices. AB - This article discusses methods of mapping and standardizing processes in physician offices and ambulatory settings. It reviews the phenomenon of process shift: how a process, once established, changes over time and loses its effectiveness. The article presents the importance of this shift and how it leads to poor performance, along with methods to prevent it from occurring. The author details the development of process measures and internal benchmarking as a way of ensuring the ongoing integrity of office operations and maintenance of a high performing office. The author also reviews the need to link job descriptions to process flowcharts. PMID- 15293656 TI - Physician joint ventures: new opportunities and risks. AB - Joint ventures involving physicians and institutions or lay investors had fallen out of favor in recent years because of concerns about transgressing government regulations. These regulations have now been clarified leading to a resurgent interest in these arrangements. This article outlines the business principles, control issues, legal setting, and the various modalities for joint venturing. PMID- 15293657 TI - Internet protection basics for the medical office. AB - The increasingly frequent invasion of new computer viruses--some 500 new ones are discovered each month--has wreaked havoc in many large network systems. Medical office computers are vulnerable as well to significant corruption of files and interruption of access to important data. This article outlines basic steps that practices can follow to assure some minimum level of security. These include the importation and continuing use of scanning technologies and adherence to staff in office protocols. PMID- 15293658 TI - Computer briefs: wireless networking-security. PMID- 15293659 TI - The seven deadly sins of yellow-page advertising...and why most health-care providers commit them! AB - Yellow-page advertising is thought to be necessary for many practices. Purchasing such advertising is a relatively complex and expensive undertaking. This article outlines some of the pitfalls managers should consider when making such purchases. They should not be left to a neophyte. They should be considered within the general context of the practice's ability to market to patients directly, and careful attention must be given to the type, pricing, and placement of the advertising. PMID- 15293660 TI - What do employees really want? AB - Each person who works in your medical practice is unique and has different goals, values, dreams, and perceptions. This uniqueness means that you can't make assumptions about what your employees want. By learning what makes the people working in your practice tick, you will be able to develop meaningful rewards and incentives and eliminate some nagging practice management problems once and for all. This article suggests a simple exercise that enables staff members to rank 10 job features in order of importance. It shares the results of original research indicating common complaints medical office employees have about their doctor-bosses. It cautions readers not to solicit employee opinions unless they can live with the truth and keep from becoming bitter, defensive, or retaliatory. The article also offers two hands-on tools for helping practice managers find out what their employees really want: a ready-to-use staff morale survey and an employee suggestion program. PMID- 15293661 TI - Restrictions on workplace romance and consensual relationship policies. AB - The workplace is frequently the site of romantic and consensual relationships. These relations may sometimes be deemed by employers to infringe on the proper conduct of business and prompt them to adopt rules that limit such interactions. This article explores some of the rights of employees and the possible policies that employers may realistically adopt within the confines of federal and state constitutional statutes, in light of court precedent and in keeping with possible union contract. PMID- 15293662 TI - Disclosing unexpected outcomes and medical error. AB - Health-care professionals have a responsibility to communicate honestly and empathetically with patients and families when there has been a disappointing outcome in care. They need to understand how all parties think about, feel, and react to the situation. It is too easy for health-care providers and patients and their families to feel that their interests are no longer aligned and to have expectations go unmet. Based on the authors' clinical experience, this article outlines a suggested educational approach for organizations interested in developing their clinicians' skills in the art of disclosure when an unexpected outcome or medical error occurs. PMID- 15293663 TI - Seven steps to reduce your malpractice risk. AB - More than ever, malpractice is one of the biggest concerns in the medical community. High premiums have caused providers to reduce services and even close shop in some areas or specialties, where premiums have increased by 100 to 200 percent. At this writing, the federal government is stymied in its efforts to provide relief. So what is a health-care organization to do? This article presents seven steps that may not lower premiums, but will help an organization decrease the likelihood of being entangled in a long, drawn-out lawsuit that has little or no medical basis. PMID- 15293664 TI - Analysis of California's Medical Injury Compensation Review Act (MICRA). PMID- 15293665 TI - Accelerated depreciation for capital expenditures under the 2003 tax law: is 2004 the year to buy? PMID- 15293666 TI - Hospital price increases continue. PMID- 15293667 TI - Trends in the consumer price index. PMID- 15293668 TI - The development and professionalization of health-care business consulting. AB - Health-care business consulting is a relatively new profession. Evolving from uncertified individuals who responded to the business needs of medical providers, professionals have formed associations and set standards for certification to ensure quality and ethical practices. This article describes the history of health-care business consulting and the evolution of its professional organizations and credentialing process. PMID- 15293669 TI - Benign cystic peritoneal mesothelioma. PMID- 15293670 TI - Thymic carcinoid tumor. PMID- 15293671 TI - Mucinous cystadenoma of the ovary. PMID- 15293672 TI - Recurrent Spigelian hernia and incidental mesenteric carcinoid. PMID- 15293673 TI - Peritoneal actinomycosis. PMID- 15293674 TI - Small bowel obstruction due to foreign body. PMID- 15293675 TI - Pneumothorax complicating tracheobronchomegaly. PMID- 15293676 TI - Pulmonary sequestration. PMID- 15293677 TI - Patellar tendon-lateral femoral condyle friction syndrome. PMID- 15293678 TI - Maxillary sinus ossifying fibroma. PMID- 15293679 TI - Ureterocele in duplicated ureteral tract. PMID- 15293680 TI - Lateral tibial rim fracture. PMID- 15293681 TI - Osteoblastoma of thoracic vertebra. PMID- 15293682 TI - Mayer-Rokitansky-Kuster-Hauser syndrome. PMID- 15293683 TI - Plexiform neurofibroma of the neck. PMID- 15293684 TI - Adult Moyamoya disease. PMID- 15293685 TI - Congenital cystic adenomatoid malformation of the lung. PMID- 15293686 TI - Duodenal adenocarcinoma. PMID- 15293687 TI - Mediastinal extramedullary hematopoiesis in hemolytic anemia. PMID- 15293688 TI - Pulmonary lymphangioleiomyomatosis and rounded atelectasis. PMID- 15293689 TI - Popliteal artery entrapment syndrome. PMID- 15293690 TI - Midgut volvulus with malrotation in a 14-year-old child. PMID- 15293691 TI - Heterotaxy syndrome. PMID- 15293692 TI - Epidermoid cyst in the prepontine cistern and pituitary macroadenoma. PMID- 15293693 TI - Stromal tumor of the small intestine. PMID- 15293694 TI - What's eating you? Cheyletiella mites. PMID- 15293695 TI - What is your diagnosis? Cicatricial pemphigoid. PMID- 15293696 TI - A diagnostic pearl: the school chair sign. AB - Allergic contact dermatitis (ACD) reactions to nickel have pleomorphic presentations owing to the ubiquity of this allergen in the environment. In children, ACD to nickel classically occurs on the earlobes, neck, wrists, and infraumbilical area. We describe a newly recognized presentation for this common childhood ACD--on the posterior thighs--which we have termed the school chair sign. PMID- 15293697 TI - Therapeutic options for herpes labialis, I: Oral agents. AB - Given the prevalence of herpes labialis, effective therapy has the potential to affect the lives of many and presents a challenge for clinicians. Over the last several years, most of the focus of herpes research has been on the treatment of genital herpes. Recently, however, several studies have been published examining the efficacy of therapies specifically for herpes labialis. Several therapeutic agents, both prescription and over-the-counter, are available for controlling and managing the disease. In this series of articles, we review oral and topical therapeutic agents that are available in the treatment of herpes labialis and its associated symptoms. This article will review oral treatment options. PMID- 15293698 TI - Therapeutic options for herpes labialis, II: Topical agents. PMID- 15293699 TI - Dermatology research within the Department of Defense. AB - Dermatologists in the US Department of Defense have made numerous research contributions over the past several decades. This article focuses on research performed during the past few years. Space does not permit a complete discussion of all research activities of the numerous Department of Defense investigators, and this review concentrates on the work of a few physicians who have made an impact in 4 areas of dermatologic research. PMID- 15293700 TI - Over-the-counter topical skin products--a common component of skin disease management. AB - Over-the-counter (OTC) products are widely recommended by physicians and utilized by the public for the treatment and prevention of disease. The use of OTC drugs has been studied extensively, but the patterns of physician recommendations for OTC topical skin products and the characteristics associated with patients receiving such recommendations remain unclear. We aimed to look at patterns of OTC topical skin product recommendations by physician specialty, patient demographics, geographical region, diagnosis, and metropolitan status to determine whether there are differences in the utilization of these products in the treatment of dermatologic conditions. We analyzed office-based physician visits for OTC topical skin product recommendations recorded in the 1995 to 2000 National Ambulatory Medical Care Survey (NAMCS). From 1995 to 2000, there were an estimated 36 million physician recommendations for OTC topical skin products. Although dermatologists were responsible for 53.8% of recommendations, pediatricians had the largest proportion of recommendations per prescription recommendation (OTC/Rx=0.58). Women patients, white patients, patients younger than 20 years, urban residents, and those living in the Southern United States received greater numbers of OTC topical skin product recommendations. Of the leading products recommended, hydrocortisone (27.6%), anti-infectives (23.4%), and moisturizers (13.4%) were the most common. OTC topical skin product recommendations by US physicians are substantial, particularly among dermatologists and primary care physicians. Physician specialty, gender, race, and age appear to be factors associated with those recommendations. PMID- 15293701 TI - Prevention and management of osteoporosis. AB - Bone is hard tissue that is in a constant state of flux, being built up by bone forming cells called osteoblasts while also being broken down or resorbed by cells known as osteoclasts. During childhood and adolescence, bone formation is dominant; bone length and girth increase with age, ending at early adulthood when peak bone mass is attained. Males generally exhibit a longer growth period, resulting in bones of greater size and overall strength. In males after the age of 20, bone resorbtion becomes predominant, and bone mineral content declines about 4% per decade. Females tend to maintain peak mineral content until menopause, after which time it declines about 15% per decade. Osteoporosis is a disease characterized by low bone mass and structural deterioration of bone tissue, leading to bone fragility and an increased susceptibility to fractures, especially of the hip, spine, and wrist. Osteoporosis occurs primarily as a result of normal ageing, but can arise as a result of impaired development of peak bone mass (e.g. due to delayed puberty or undernutrition) or excessive bone loss during adulthood (e.g. due to estrogen deficiency in women, undernutrition, or corticosteroid use). Osteoporosis-induced fractures cause a great burden to society. Hip fractures are the most serious, as they nearly always result in hospitalization, are fatal about 20% of the time, and produce permanent disability about half the time. Fracture rates increase rapidly with age and the lifetime risk of fracture in 50 year-old women is about 40%, similar to that for coronary heart disease. In 1990, there were 1.7 million hip fractures alone worldwide; with changes in population demographics, this figure is expected to rise to 6 million by 2050. To help describe the nature and consequences of osteoporosis, as well as strategies for its prevention and management, a WHO Scientific Group meeting of international experts was held in Geneva, which resulted in this technical report. This monograph describes in detail normal bone development and the causes and risk factors for developing osteoporosis. The burden of osteoporosis is characterized in terms of mortality, morbidity, and economic costs. Methods for its prevention and treatment are discussed in detail for both pharmacological and non-pharmacological approaches. For each approach, the strength of the scientific evidence is presented. The report also provides cost-analysis information for potential interventions, and discusses important aspects of developing national policies to deal with osteoporosis. Recommendations are made to the general population, care providers, health administrators, and researchers. Lastly, national organizations and support groups are listed by country. PMID- 15293702 TI - [Can you recognize traumatic brain injury?]. PMID- 15293703 TI - [Home hemodialysis]. PMID- 15293704 TI - [Values of the society and well-being of an individual member]. PMID- 15293705 TI - [Prognostic factors and improving treatment of cutaneous melanoma]. PMID- 15293706 TI - [Endocannabinoids multifunctional messenger system in control of pleasure and eating habit]. PMID- 15293707 TI - [Birth control during lactation and breast feeding]. PMID- 15293708 TI - [Tooth implant--a spare part becoming commonly used]. PMID- 15293709 TI - [Topically used antiinflammatory drugs]. PMID- 15293710 TI - [Solitary fibrous tumors of the pleura]. PMID- 15293711 TI - [Permanent changes in insulin resistance by changes in life style among subjects with impaired glucose tolerance]. PMID- 15293712 TI - [Young women suffering from chronic bleeding in the genital area]. PMID- 15293713 TI - [Schizophrenia]. PMID- 15293714 TI - [Missing details from the neurologist's report regarding a case report in Duodecim 3/2004]. PMID- 15293715 TI - [Food allergy among children]. PMID- 15293716 TI - Philip Barrough, Elizabethan physician with the first English book on medicine. AB - Philip Barrough was an Elizabethan surgeon and physician who, in 1587, published the first book on medicine in the English language. In addition to a comprehensive presentation of the overall knowledge of medicine of the time, Barrough devoted several chapters to the diagnosis and management of the diseases of the teeth and diseases of the mouth and oropharynx. Very little is known about the life of Barrough, such as the dates of his birth and death and where he practiced medicine. It is curious that despite his major contribution to medicine, and the fact that his book went into seven editions, his name is not even mentioned in the major histories of medicine, such as those of Garrison, Castiglioni, Major, and Mettler. This paper attempts to place Barrough in his proper place in the history of medicine, and to emphasize his overall knowledge of medicine as well as his concern about diseases of the mouth and teeth. PMID- 15293717 TI - Root canal irrigation--a historical perspective. AB - The concept of the germ theory of disease combined with the development of dentistry during the latter half of the 19th century had a direct effect on the practice of endodontics. The significance of root canal irrigation to endodontics strengthened in the period between 1859 when Taft recommended frequent syringing of the root canal to remove "irritants" until the mid-1940s when endodontics became a special field within dentistry and the American Endodontic Society was established. This paper reviews the role of irrigants and irrigation in root canal treatment during this period. A variety of recommendations on the use of solutions to clean root canals appeared in the dental literature, often innovative and at times entrepreneurial, but invariably empirically based. While it was widely assumed that by wiping the root canal with disinfectants sterilization would be achieved, many of the principles associated with cleaning the root canal published during this period, in particular by Willoughby Dayton Miller in the 1890s and Louis Grossman in the 1940s, remain equally relevant in the 21st century. PMID- 15293718 TI - Dentistry on stamps. PMID- 15293719 TI - Dr. Byron McKeeby's contribution to Grant Wood's "American Gothic". AB - Grant Wood (1891-1942) was a major American regionalist artist from Iowa who produced his world famous icon, American Gothic in 1930. Along with Mona Lisa and Whistler's Mother, this work has become one of the most recognized paintings in the world. While it has been greatly admired by most of its viewers, it has been criticized by others. Currently on display at the Art Institute of Chicago, this oil painting portrays a grim faced farmer and his somber daughter who are resolutely standing in front of their rural, Gothic Revival farmhouse. The tall, gaunt farmer, looking straight ahead, firmly holds a pitchfork whose tines point upward, while his daughter, with her eyes averted appears to be showing disapproval. In American art, it is unusual to find the hard, cold realism and honest, direct and earthy qualities that Wood captures in American Gothic. Wood's actual intention was to present the work as a subtle and witty commentary on midwestern, rural life. To achieve his goal, Wood emulated the meticulous style and technique of the 16th century Flemish masters. His sister, Nan Wood Graham (1900-1990), and his 62-year-old dentist, Dr. Byron H. McKeeby (1867-1950), posed for the painting. This article describes how Grant developed this artistic style which is reflected in American Gothic. Additionally, it examines the strained interactions between Grant Wood, the artist, and Dr. Byron McKeeby, the model. PMID- 15293720 TI - Ralph L. Huber, DMD: forgotten inventor of the "Tuohy" needle. AB - The hypodermic needle popularized by Edward Tuohy, MD for use in his method of continuous spinal anesthesia has been incorrectly called the "Tuohy needle" since he first popularized it in 1945. It was recently determined that Ralph L. Huber, a Seattle dentist, was the inventor of this widely used needle. However, very little is known about Huber. There is no mention of him in the medical literature, and until recently, he was forgotten. Through the location of various primary sources, including the original needle point invented by Dr. Huber, this essay introduces the fascinating life of Dr. Huber, and also his personal notes about his inventions. Not only was he a prominent Seattle dentist, but also a prolific inventor. PMID- 15293721 TI - The Boston Massacre: the dental connection. PMID- 15293722 TI - Gleanings about dentistry from the world of literature (thirty-three in a series). PMID- 15293723 TI - [Clinical implications of pharmacokinetics--factors affecting on the serum drug concentration]. PMID- 15293724 TI - [Transabdominal ultrasound of gastrointestinal diseases]. PMID- 15293725 TI - [Endosonographic diagnosis of the depth of cancer invasion of the alimentary tube]. PMID- 15293726 TI - [A case of enhanced FDG uptake in esophageal GIST (gastrointestinal stromal tumor)]. PMID- 15293727 TI - [Four cases of obstructive colitis diagnosed preoperatively]. PMID- 15293728 TI - [A case of pseudolymphoma of the liver diagnosed before operation]. PMID- 15293729 TI - [A case of rapid onset autoimmune hepatitis resembling drug-induced liver disease in a Japanese young woman]. PMID- 15293730 TI - [A case of communicating accessory bile duct]. PMID- 15293731 TI - [A case of pancreaticobiliary malfunction complicated with particular accessory pancreatic duct detected by three dimensional-CT after ERCP]. PMID- 15293732 TI - [Intractable stomatitis with C type liver cirrhosis and genital lichen planus]. PMID- 15293734 TI - [Extracorporeal shockwave lithotripsy in the treatment of distal ureteral stones larger than 10 mm in diameter]. AB - Optimal treatment for distal ureteral stones remains controversial. During a period of 10 years, from December 1992 to December 2002, 103 distal ureteral stones larger than 10 mm in diameter were treated at our institution with extracorporeal shock wave lithotripsy (ESWL) using the Siemens Lithostar. Only 2 patients had a ureteral stent in place at the time of treatment. The overall stone-free rate was 98% with 1-12 session and 3-month stone-free rate was 95.1%. These data reveal that a high success rate was achieved in multisession ESWL. Therefore, ESWL is considered to be acceptable as first-line therapy for fragmentation of distal ureteral stones larger than 10 mm in diameter. PMID- 15293735 TI - Partial nephrectomy for small localized renal cell carcinoma. AB - We retrospectively reviewed the records of 54 patients with RCC who underwent partial nephrectomy for the primary lesion between 1992 and 2001. The indications for partial nephrectomy were elective in 43 and imperative in 11 patients. We selected 51 patients with clinical stage T1a who underwent open radical nephrectomy for localized RCC for comparison during the same period. We evaluated the peri- and postoperative complications, disease-free survival rates and changes of renal function in the partial nephrectomy (PN) group, compared to the radical nephrectomy (RN) group. There was no significant difference with regard to pathological findings and clinical outcomes between two groups, except for the amount of intraoperative bleeding. Three patients in the PN group developed postoperative complications, consisting of urine leakage in 2 patients and renal hypertension in 1 patient. The 5-year disease-free survival rates in the PN and RN groups were 90% and 97%, respectively. Local recurrence from the resected area of the renal parenchyma was not found in patients in the PN group. All patients in the PN group maintained satisfactory and stable renal function. In the RN group, renal function slowly deteriorated in 2 patients. Therefore, partial nephrectomy offers cancer control and an acceptable low mortality rate, comparable to those of radical nephrectomy. PMID- 15293736 TI - [Bladder neck contraction after radical prostatectomy: morbidity and risk factors]. AB - We retrospectively analyzed the morbidity and risk factors of bladder neck contraction (BNC) after retropubic prostatectomy. Of 99 consecutive patients who underwent radical retropubic prostatectomy between 1995 and 2001, 10 (10%) developed anastomotic stricture after surgery. BNC was diagnosed 7 months on average after the surgery. Nine patients were successfully treated by cold-knife incision, but 5 required additional incision against the recurrence of BNC. None of them revealed newly diagnosed stricture or incontinence. Thirteen potential risk factors including age, stage, PSA, neoadjuvant treatment, pathology, intraoperative blood loss, operation time, operator, nerve sparing, extravasation of urine, marginal status, the year of operation, and the method of ureterovesical anastomosis were evaluated using univariate and multivariate analysis. The BNC rate was increased in patients with longer (more than 4 hrs) operations and excessive bleeding (more than 1,000 ml) (p=0.0027, respectively). The year of operation (before 1998, p=0.0015), the method of ureterovesical anastomosis (p=0.0017), operator experience (less than 5 cases, p=0.0056), and neoadjuvant treatment (p=0.09) were also risk factors. In multivariate analysis, the year of operation (p=0.03) and operator experience(p=0.04) were strong predictors of BNC. The skill levels of surgeons and institutes are expected to decrease BNC. PMID- 15293738 TI - Solitary fibrous tumor of the prostate. AB - Solitary fibrous tumor of the prostate is extremely rare. Only five cases have been reported to the present. A 36-year-old man presented to our hospital complaining of difficulty in urination. Retrograde urethrography and urethroscopy demonstrated intraurethral protrusion of the left prostatic lobe and complete obstruction of prostatic urethra. Magnetic resonance imaging demonstrated the prostatic tumor with prominent intravesical protrusion. Transrectal echo-guided biopsy was performed on the supposition of leiomyosarcoma of prostate. However, the tumor was diagnosed as benign fibrous tumor, so the patient underwent transurethral resection of left prostatic lobe for the purpose of improving urinary condition, avoiding retrograde ejaculation. Histologic examination revealed a dense proliferation of spindle cells with no nuclear atypia. No mitotic figure was seen. The pathologic diagnosis was benign solitary fibrous tumor of the prostate, because the tumor was positive for CD34 and negative for alpha-smooth muscle actin and desmin by immunohistochemical study. The patient remained well without regrowth of the tumor during the last six years. PMID- 15293737 TI - [Laparoscopic adrenalectomy: Akita University experience]. AB - We performed 52 laparoscopic adrenalectomies between January 1997 and March 2003. In 51 patients, adrenal tumors were successfully removed laparoscopically. In one patient, the laparoscopic procedure was converted to open surgery because of insufflator's trouble and hemorrhage. Perioperative blood transfusion was required in two patients; one for intraoperative and another for postoperative bleeding. We compared laparoscopic adrenalectomy with conventional open surgery which had been performed in our clinic before the laparoscopic adrenalectomy era. The mean operative time of the laparoscopic adrenalectomy (187 +/- 59 min) was not significantly longer than that of the open surgery (193 +/- 49 min). The mean estimated blood loss of laparoscopic adrenalectomy (75 +/- 145 g) was significantly less than that of the open surgery (438 +/- 447 g). Time to oral intake and ambulation for laparoscopic adrenalectomy were significantly earlier than those of the open surgery. Operation time was significantly shortened as the experience of this surgery was accumulated. The results of our experience support the concept that laparoscopic adrenalectomy is safe and is a standard treatment for surgical treatment of adrenal tumors. PMID- 15293739 TI - [Intraurethral condyloma acuminatum: a case report]. AB - A 47-year-old man presented with meatal bleeding. Visual inspection revealed papillary meatal tumors at 5 and 7 o'clock without other genital warts. These lesions were completely resected, and the histological diagnosis was condyloma acuminata. After 2 months, recurrence of warts was not observed macroscopically in the genital area, including the urethral meatus. However, urethroscopy revealed a papillary tumor in the navicular fossa. This lesion was removed and pathological examination revealed that it was also a condyloma acuminatum. PMID- 15293740 TI - [A case of ex vivo renal artery reconstruction and autotransplantation for renal artery aneurysm]. AB - A 74-year male patient was admitted to our department with a left renal artery aneurysm (RAA). It was detected by a computed tomography (CT) scan while performing an examination for hypertension. The diameter of the aneurysm was 25 mm. There was no evidence of calcification. Selective left renal angiography and a 3-dimensional (3D)-CT image revealed a saccular renal aneurysm in the left main renal artery. Because of the risk of rupture, autotransplantation of the left kidney to the left iliac fossa was performed after resecting the aneurysm and reconstructing the left artery under bench surgery. Postoperative 3D-CT revealed no stenosis. This ex-vivo technique and autotransplantation into ipsilateral iliac fossa are both effective and safe for the treatment of RAA. PMID- 15293741 TI - [Retroperitoneal malignant peripheral nerve sheath tumor (MPNST) complicated with von Recklinghausen's disease: a case report]. AB - A retroperitoneal malignant peripheral nerve sheath tumor (MPNST) in a patient with von Recklinghausen's disease is reported. A 55-year-old woman was admitted with a left side abdominal mass. Physical examination showed numerous cafe-au lait-spots, subcutaneous masses, scoliosis, and a baby's head-sized fixed mass in the left abdomen. Computed tomography (CT) and magnetic resonance imaging (MRI) revealed a 9 x 9 cm retroperitoneal mass. Two other tumors were also found. One on the left side of the T2-T3 thoracic spine, and the other posterior to the right hip joint. The retroperitoneal tumor was resected en bloc. The tumor was a solid yellow mass. Macroscopically it has a pseudocapsule of fibrous tissue, weighed 1,120 g and measured 9 x 9 x 15 cm. The histopathological diagnosis was malignant peripheral nerve sheath tumor (MPNST). Since the responsiveness of these tumors to chemotherapy and radiation therapy is poor, we did not administer adjuvant therapy. The patient is alive with no evidence of recurrence more than 6 months after surgey. PMID- 15293742 TI - [A case of malignant peripheral nerve sheath tumor of the kidney]. AB - We report a case of malignant peripheral nerve sheath tumor (MPNST) of the kidney. A 50-year-old man was referred to our hospital because of left back pain. Computed tomographic scan and magnetic resonance imaging revealed a huge left retroperitoneal tumor (30 x 13 x 11 cm), which consisted of a solid tumor with calcification surrounding the cystic component. He underwent resection of the retroperitoneal tumor and additional descending colectomy. Histopathological examination revealed MPNST arising from severely atrophic renal parenchyma of giant hydronephrosis. He died of tumor metastasis to the lungs 3 months after the operation. To our knowledge, this is the fourth case of MPNST of the kidney, moreover the third case arising from the renal parenchyma to be reported in Japan. PMID- 15293743 TI - [Bellini duct carcinoma: a case report]. AB - A 53-year-old man was admitted to our hospital for the extensive examination and treatment of suspicioun of right renal pelvic tumor. Retrograde pyelography (RP), computed tomography (CT) and magnetic resonance imaging (MRI) showed a space occupying lesion, about 2 cm in diameter, spread from the renal parenchyma to the renal pelvis. Right nephroureterectomy was performed because transitional cell carcinoma was suspected from the histopothology of the frozen section. The gross examination revealed a white tumor in the upper pole, protruding into the renal pelvis with hemorrhagic necrosis. Histological examination showed Bellini duct carcinoma of the papillary type. He received adjuvant combination chemotherapy with M-VAC (Methotrexate, vinblastine, doxorubicin, cisplatin). He has been alive without evidence of recurrence since the surgery. PMID- 15293744 TI - A case of quadruple cancer including urinary bladder, oral cavity, stomach and lung. AB - A 67-year-old man, who had smoked heavily for many years, was found in 1997 to have bladder tumors, and transurethral resection of the bladder tumor (TUR-Bt) was performed. Histopathological diagnosis was urothelial carcinoma (G2>G3, pTa, N0, M0, ly0, v0). In December, 1998, he noticed an oral cavity tumor. After preoperative radiation therapy (total 40 Gy, 17 times), surgical treatment was undertaken. Histopathological diagnosis was well differentiated squamous cell carcinoma (pT2, pN2b, M0). In February, 2000, gastric tumor was detected by endoscopic examination, and subtotal gastorectomy and Roux en Y operation were performed. Histopathological diagnosis was well differentiated adenocarcinoma (pT2, pN0, M0, P0, CY0). A chest computed tomographic (CT) scan revealed a solitary lung tumor in April, 2000. Partial peumonectomy was performed, and histopathological diagnosis was poorly differentiated adenocarcinoma (pT1, N0, M0, P0). In April, 2000, multiple lesions of bladder cancer in the neck of the urinary bladder and posterior urethra were found and radical cystoprostatourethrectomy combined with lymph node dissection and bilateral cutaneous ureterostomy were performed (urothelial carcinoma, G3, pT4a, pN2, M0, pL2, pV0, pR0). Since then, the patient has been followed carefully. PMID- 15293745 TI - [Right ectopic ureter with ipsilateral renal agenesis presenting with infertility: a case report]. AB - A 30-year-old man was referred to our hospital with a complaint of male infertility, presenting abnormal semen analysis (semen volume 0.1 ml, sperm concentration 1.2 x 10(6)/ml, sperm motility 0%). Radiological examinations demonstrated right renal agenesis and a cystic mass extending from the prostate to the posterior of the bladder. Our final diagnosis was obstruction of the ejaculatory duct secondary to right ectopic ureter associated with ipsilateral renal agenesis. The patient was treated by transurethral unroofing of the cyst. Three months after the surgery, the cystic mass disappeared and the seminal findings showed marked improvement. PMID- 15293746 TI - Renal pelvic urothelial carcinoma in horseshoe kidney. AB - A 40-year-old man with asymptomatic gross hematuria visited our hospital. He had been followed up on the horseshoe kidney and left ureteral stone. Cystoscopy revealed a flow of gross hematuria from the left orifice. Drip infusion pyelography, retrograde pyelography, abdominal computerized tomography, magnetic resonance imaging revealed a renal pelvic mass in the upper pole of left kidney. Left nephroureterectomy and isthmusectomy and partial cystectomy were done. A microwave tissue coagulator (Microtaze AZM-520, AZWELL) was used to divide the isthmus. There was very little bleeding and leakage of urine from the divided isthmus. The postoperative course was favorable. PMID- 15293747 TI - [A case of vesico-urethral foreign body with urinary fistula]. AB - We report a rare case of vesico-urethral foreign body with urinary fistula. On March 12, 2002, a 53-year-old single men inserted a 3 m vinyl tube 6 mm in diameter into his urethra by himself for the purpose of masturbation, but he could not remove the tube. He cut his urethra by himself and tried to remove the tube. The next day he was admitted to our hospital, with complaints of urinary fistula and fever. Open surgical removal of foreign body, cystostomy and repair of bulbous portion of the urethra were performed. At 24 days postoperatively the cystostomy and urethral catheter were removed and urination became smooth. PMID- 15293748 TI - [A case of schwannoma in the renal hilum]. AB - A 58-year-old female presented with a well-encapsulated tumor in the left renal hilum on computed tomography (CT). On magnetic resonance imaging (MRI), the tumor showed low intensity on the T1-weighted image, high intensity on the T2-weighted image. Laparoscopic radical nephrectomy was performed because we could not exclude the possibility of malignancy such as renal cell carcinoma. Histopathological examination revealed that the tumor was a benign Schwannoma. Although renal hilum is a common site of renal schwannoma, it is difficult to differentiate benign tumors from malignant ones, and then nephrectomy is usually performed. PMID- 15293749 TI - [Pulmonary nontuberculous mycobacteriosis in patients with lung cancer]. AB - OBJECTIVES: To clarify the clinical features of the coexisting lung cancer and nontuberculous mycobacteriosis of the lung. MATERIALS AND METHODS: We analyzed clinical data on 11 admitted cases of coexisting lung cancer and pulmonary non tuberculous mycobacteriosis at National Organization Tokyo Hospital during the period from 1997 to 2002. RESULTS: There were 10 men and 1 woman, with a mean age of 66 years. Five of 11 patients had underlying pulmonary disorders, such as healed tuberculosis and lung cyst. Histological types of lung cancer were squamous cell carcinoma in 4, adenocarcinoma and small cell carcinoma in 3 each, and 8 out of 11 cases were in stages III to IV. We classified the 11 cases into 2 groups: (1) lung cancer concurrently detected with mycobacteriosis (8 cases) and (2) lung cancer sequentially detected during the follow-up of mycobacteriosis (3 cases). Lung cancers in the latter group were in relatively early stages and all patients of this group received resection of the cancer, while most of lung cancers in the concurrent group were in far-advanced, and palliative and/or supportive treatment for lung cancer were frequently selected. The strains of mycobacteria were as follows: M. avium complex (6 cases) and M. kansasii (5 cases). The incidence of lung cancer among patients with nontuberculous mycobacteriosis was 2.5 percent (2 percent of M. avium complex diseases patients and 8.2 percent of M. kansasii disease patients), while the incidence of nontuberculous mycobacteriosis in untreated lung cancer patients was 1.4 percent. Analysis of anatomical relationship between lung cancer and non-tuberculous mycobacteriosis revealed that the two diseases located in the same lung in 8 cases, and also in the same lobe in 4 out of the 8 cases. Outcome of treatment for nontuberculous mycobacteriosis was good especially in patients with M. kansasii disease, and it seemed that coexisting nontuberculous mycobacteriosis did not influence on the prognosis of lung cancer patients. CONCLUSION: In the management of lung cancer, physicians should consider the possibility of coexisting pulmonary non-tuberculous mycobacteriosis, as well as coexisting pulmonary tuberculosis. PMID- 15293750 TI - [Current status of the criteria for discharging patients with pulmonary tuberculosis--results of questionnaire survey on the criteria for discharging patients from tuberculosis wards of the hospitals in Kanto area]. AB - INTRODUCTION: Emphasis in treating patients with infectious pulmonary tuberculosis has come to be laid on the execution of reliable standard chemotherapy. As a result, hospitalization for a prolonged period has become unnecessary any more. However, few attempts have been made so far on the determination of discharging criteria. METHODS: We performed a questionnaire survey to hospitals with wards for tuberculosis in Kanto area and asked questions on the current status of discharging criteria. RESULTS: The effective response rate to the survey was 63.0 %. Sputum smear examination carried out mainly by Ziehl-Neelsen method and fluorescence method in 17.2% and 72.4 % of the hospitals, respectively. Sputum culture examination was carried out using mainly Ogawa medium and a liquid medium in 62.1% and 27.6% of the hospitals, respectively. Discharging criteria were standardized in 79.3% of hospitals. Negative sputum smear was used as the criterion for determining discharge in 11 sets of criteria. Negative sputum culture was used as the criterion for determining discharge in 17 sets of criteria. In the remaining one hospital, patients were to be discharged after 2-month treatment. There was no consistency in the procedure and the frequency of sputum examinations, how many negative results are needed to confirm negative status and the criteria for judgment. CONCLUSION: These results suggest that further evaluation must be made on the treatment outcome at each hospital, and the standard discharging criteria should be worked out taking into account the capacity of each hospital and the care situation of local community. PMID- 15293751 TI - [Tuberculin skin test reaction of health-care workers exposed to tuberculosis infection]. AB - OBJECTIVE: The objective of this study was to describe tuberculin skin test (TST) of health-care workers who had had contacts with a tuberculosis patient and to investigate whether the distributions of the size of TST were different by sputum smear status of index cases and medical procedures done to the patient. SUBJECTS: Health-care workers who were exposed to tuberculosis infection through contact with patients before diagnosis of tuberculosis were included in this study. Index cases had pulmonary tuberculosis with positive sputum smear and were registered at Sapporo Public Health Office from April 2001 to March 2002. Subjects without past history of BCG (bacilli Calmette-Guerin) vaccinacion were excluded, and final study subjects were 415 (52 male and 363 female, mean age 29.1 years). METHOD: Characteristics of index cases and contact status were obtained from the registration cards of tuberculosis cases at Sapporo Public Health Office. Subjects were divided into two or more categories by the characteristics of index cases and the contact status. Distributions of TST of the subjects in different categories were compared. RESULTS: Contacts with patients received tracheal aspiration showed significantly larger TST reaction than contacts with patients not receiving tracheal aspiration. Among contacts with patients showing minimum to moderately positive sputum-smear, TST reaction was significantly larger in contacts with patients received tracheal aspiration (mean diameter of erythema 35.8 mm) than those not receiving tracheal aspiration (24.8 mm). In contrast, among contacts with patients of severely positive sputum-smear, TST reaction was not significantly different between contacts of patients received tracheal aspiration (35.3 mm) and not receiving tracheal aspiration (33.1 mm). CONCLUSION: TST was significantly stronger in contacts with a tuberculosis patient received tracheal aspiration, which indicates medical procedures such as tracheal aspiration increases the risk of tuberculosis infection in health-care workers. PMID- 15293752 TI - [Multi-drug resistant lung tuberculosis due to double infection of MDR strain]. AB - The case is 47-year-old, homeless man. He was diagnosed as cavitary, sputum smear positive pan-sensitive lung tuberculosis, and admitted to TB ward of our hospital. At the age of 4-year-old he had tuberculous hilar lymphadenopathy and took medicine. He had no other associated disease, and HIV test was negative. He started standard chemotherapy and 2 months later his sputum culture was converted to negative. His adherence to medicine was thought to be good. But about 2 weeks after the sputum conversion, his sputum culture was re-converted to positive for Mycobacterium tuberculosis. Thereafter, during and after the completion of standard chemotherapy, his sputum culture had been intermittently positive. The drug sensitivity tests of the strain after re-conversion showed multi-drug resistance. RFLP analysis revealed that the strain before conversion was totally different strain from the strain after re-conversion to positive. The case was considered to be caused by the double infection of MDR strain of Mycobacterium tuberculosis during the course of treatment for tuberculosis due to a sensitive strain of Mycobacterium tuberculosis. PMID- 15293753 TI - [CT imaging findings in congenital tuberculosis, Part I: Usefulness of periportal hypodensity in the diagnosis of congenital tuberculosis]. AB - We treated three newborns and early infants with congenital tuberculosis between 1996 and 2001. We reported imaging presentations of the three cases born in and after 1996 when new CT equipment (CT-HAS-SGS, GE-Yokokawa, Tokyo) was introduced in our institute. These three cases of congenital tuberculosis showed periportal hypodensity, in addition to pulmonary infiltrate, mediastinal and abdominal lymphadenopathy on CT images. Early diagnosis of congenital tuberculosis is urgently needed, however, it is very difficult. Our findings suggested that clinical suspicion supplemented by careful imaging examinations may facilitate the early diagnosis of congenital tuberculosis, and the detection of periportal hypodensity may offer a new additive diagnostic option to congenital tuberculosis. PMID- 15293754 TI - [A case of pulmonary Mycobacterium kansasii infection complicated with pleural effusion]. AB - A 60-year-old man was admitted to our hospital complaining of general malaise for a few months, discomfort of the right shoulder and fever in May 2003. Chest X-ray film showed an infiltrative shadow in the right lung field and chest CT scan revealed right pleural effusion. Pulmonary tuberculosis complicated with pleurisy was first suspected from the findings of high ADA level of the effusion and positive result of PPD skin test. But, microscopic examination of the specimens obtained by transbronchial lung biopsy disclosed granulomatous lesions and Mycobacterium kansasii was identified from all specimens; sputum, fluids of brushing and bronchial washing. The patient was diagnosed as pulmonary Mycobacterium kansasii infection and treated with anti-tuberculous drugs including RFP. His clinical course was good and no recurrence of pleural effusion was seen. This case was a rare case of pulmonary Mycobacterium kansasii infection complicated with pleural effusion. PMID- 15293755 TI - [Clinical characteristics of cerebral infarction in China and Japan]. AB - In order to elucidate the clinical characteristics of cerebral infarction in a northeastern district of China, 1,353 patients with first-ever cerebral infarction in Shenyang, China, were compared with 2,929 patients registered in Japan. Using the identical database (Japan Standard Stroke Registry Study), we prospectively collected clinical data on acute ischemic stroke patients who were admitted to two main hospitals in Shenyang, China, and 50 hospitals in Japan. The mean age was 67.3 years in China and 71.3 years in Japan. Of the patients in China, 78% were classified as atherothrombotic infarction, 18% as lacunae, and 1% as cardioembolism. By contrast, 30% of the patients in Japan were classified as atherothrombotic infarction, 32% as lacunae, and 31% as cardioembolism. Regarding the risk factors, the incidence of hypertension was 76% in China and 62% in Japan. Diabetes mellitus was 10% in China and 26% in Japan. Hyperlipidemia was 10% in China and 22% in Japan. Atrial fibrillation was 3% in China and 21% in Japan. As the diagnostic criteria for hyperlipidemia differed, we could not compare this risk factor between the two countries statistically; however, cardioembolism, lacunae, and atrial fibrillation were significantly less common in China than in Japan. Our data suggest that hypertension is an important risk factor for ischemic stroke, and the incidence of cardioembolism and atrial fibrillation is very low in Shenyang, China. PMID- 15293756 TI - [Brain hemorrhage with medical treatment: prediction of early outcome and production of critical paths]. AB - The first purpose of this study was to detect clinical and radiological factors on admission which predict early outcome of patients in brain hemorrhage with medical treatment. For 50 consecutive patients in our Cerebrovascular Center, NIH Stroke Scale score was a useful indicator for prediction of independent daily life, discharge to home, and death in the acute stage; the score < or = 4, < or = 7, and > or = 23 were the most appropriate cut-off values for the above events, respectively. The second purpose of this study was to produce critical paths of medical management for brain hemorrhage based on the above results. We prepared three courses of paths according to clinical severity. As inclusion criteria for each course, we used the above three cut-off values and hematoma volume. Duration of hospitalization of the three courses was 16, 20, and 28 days. When we applied the paths to 200 patients with brain hemorrhage who were enrolled in the research grant supported by the Japanese ministry of health, labor and welfare (12C-10), duration of hospitalization for the majority of the patients were 5 days or more than the planned duration in the paths. It is indispensable to manage acute stroke patients according to critical paths, because standard and efficient strategies of clinical medicine have been stressed these years. We will immediately apply the new paths in this study to patients in our center, and renew them at short intervals. We think that we can contribute to new evidences for standard medical management of brain hemorrhage by our approach to the critical paths. PMID- 15293757 TI - [Relapsing polychondritis with an intracranial granuloma: a case report]. AB - We report a case of relapsing polychondritis (RP) with an intracranial granuloma. A 67-year-old man developed progressive disorientation during the course of RP with left auricular chondritis and episcleritis. He had history of sinusitis and rupture of an aneurysm in middle cerebral artery. Laboratory examinations revealed high erythrocyte sedimentation rate and positive C-reactive protein. Head CT and MRI with contrast enhancement showed a mass adjacent to the falx cerebri and lesions in the frontal skull base. The mass was surrounded by extensive perifocal edema that spread mainly into the frontal white matter on both sides. Histologically, the mass displayed an inflammatory granuloma. By removal of the mass, edema decreased around the granuloma, and his disorientation improved markedly. Surgical findings revealed the granuloma was separated from sinusitis. There are a few reports on RP with an intracranial granuloma. PMID- 15293758 TI - [A case of acute encephalopathy due to datura seed poisoning]. AB - A 83 year-old-man presented with a stuporous state. He ingested datura seeds approximately 1.5 hour before the onset of the symptom. On arrival at our emergency clinic 2 hours and 40 minutes after the ingestion, he was semi-comatose with intermittent tonic convulsive seizures. The pupils were fully-dilated and unreactive, and salivation was decreased. He showed hypohidrosis and exaggerated deep tendon reflexes with positive Babinski's sign. His body temperature was 37.6 degrees C, and blood pressure was 167/99 mmHg. After gastrolavage and administration of charcoal and cathartics, his pupil became reactive to light, and he became alert gradually. His consciousness became clear 24 hours after seed ingestion and discharged with no residual neurological signs. PMID- 15293759 TI - [Bulbovascular compression by megadolichobasilar artery manifested as neurogenic and refractory hypertension]. AB - A 37-year-old man with juvenile and refractory hypertension was admitted to our hospital for progressive left hearing loss, vertigo, and dizziness. Neurological examination revealed left hearing loss and exaggerated deep tendon reflexes and vestibular dysfunction. MRI and cerebral angiography disclosed megadolichobasilar artery (MDBA). Moreover, modified MR cisternography at the medulla disclosed marked compression and deformity of the left rostral ventrolateral medulla (RVLM) by the dolichoectatic right vertebral artery. In the literatures, bulbovascular compression has been reported in 4 among 9 patients with MDBA (including the present patient), for whom MRI of the medulla was presented. All 4 patients suffered from hypertension, and at least 3 of them showed juvenile and refractory hypertension. Ipsilateral pyramidal tract disturbance (Opalski syndrome) was observed in 3 patients. Considering the recent concept that the cardiovascular center can be localized at the RVLM, juvenile and refractory hypertension, and possibly Opalski syndrome in the present patient can be attributed to bulbovascular compression by MDBA. In the patients with MDBA and hypertension or Opalski syndrome, MR cisternography of the medulla is warranted to evaluate compression by MDBA. PMID- 15293760 TI - [Late-onset of idiopathic paroxysmal kinesigenic choreoathetosis: a case report]. AB - A case of late-onset idiopathic paroxysmal kinesigenic choreoathetosis (PKC) is described herein. A 50-year-old right-handed Japanese man with no family history of neurological disease or consanguinity was referred to our neurological unit because of paroxysmal involuntary movement of his right extremities. Physical examination findings were normal. Neurological examination during the interictal period revealed no abnormality. The attacks were triggered by abrupt initiation of voluntary movement and occurred about 30 times a day, lasting 30 to 60 seconds with no disturbance of consciousness. No metabolic abnormalities were found, and brain magnetic resonance imaging showed no abnormality including lacunar infarction. Interictal single photon emission computed tomography (SPECT) showed hypoperfusion of left basal ganglia. Interictal electroencephalography was normal without epileptic discharges. Because the patient did not have any condition that could cause secondary PKC, he was diagnosed as having idiopathic PKC. Treatment with carbamazepine 200 mg once a day resulted in control of the attacks, and 500 mg a day resulted complete resolution. To the best of our knowledge, this case may represent the oldest age reported for idiopathic PKC onset. Although the etiology of PKC remains unknown, late-onset idiopathic PKC is rare. Our SPECT finding leads us to suppose that dysfunction of the basal ganglia is likely involved in the pathogenesis of PKC. PMID- 15293761 TI - [A case of Crow-Fukase syndrome showing improvement following excision and irradiation of bone lesions]. AB - A 57-year-old woman suffering from pleural and pericardial effusion, pulmonary hypertention, lymphadenopathy, hepatosplenomegaly, edema, hypertrichosis, small hemangioma and polyneuropathy was diagnosed as Crow-Fukase syndrome. Osteoctomy of the left second rib and irradiation of this rib and the left iliac bone were performed. Serum vascular endothelial growth factor (VEGF) level decreased to less than one-half the level before the operation (from 5,180 to 2,150 pg/ml). Immediately after the operation, pleural and pericardial effusions due to hyperpenetration improved, and polyneuropathy and hypertrichosis due to hypervasularity also gradually improved. The resected lesion was histopathologically found to be of a plasmacytoma of the IgG lambda type. Since the level of VEGF in the tissue specimen was much lower (116 pg/ml) than that in the serum, VEGF could not have been produced by the plasmacytoma. PMID- 15293762 TI - [A case of cerebral putaminal hemorrhage complicating a brain abscess proved by craniotomy]. AB - A 40-year-old man was admitted to our hospital because of consciousness disturbance, dysarthria and numbness in his right hand. Computed tomography of the head showed a cerebral hemorrhage of the left putamen. The patient was judged to have an indication of operation, and frontal craniotomy to evacuate hematoma was performed. During the operation, massive whitish pus flowed out at the time of surgery. Therefore, hemorrhage into a brain abscess was diagnosed. We reported this unique and interesting case whose brain abscess could not be differentiated from an ordinary hypertensive putaminal hemorrhage based on clinical findings and CT image. This diagnosis was not made until the patient was operated on through a craniotomy. PMID- 15293763 TI - [A patient with limb girdle muscular dystrophy type 2B (LGMD2B) manifesting cardiomyopathy]. AB - A 57-year-old woman first noticed difficulty in walking at the age of 34 years, and since then muscle wasting and weakness in the lower limbs and proximal portion of the upper limbs had progressed slowly. Serum CK was elevated. Immunohistochemical study of the biceps brachii muscle showed deficiency of dysferlin in sarcolemma, and the dysferlin gene analysis disclosed 3370 G-->T missense mutation. These findings led us to diagnose her as LGMD2B. Moreover echocardiogram revealed ventricular enlargement and diffuse hypokinesia suggesting secondary cardiomyopathy atributable to muscular dystrophy. Careful cardiac monitoring should be carried out in dysferlinopathy patients. PMID- 15293764 TI - [A case of hair loss induced by carbamazepine]. AB - We report a 52 year-old man presenting with an acute considerable hair loss induced by carbamazepine (CBZ). The remarkable scalp hair loss started within a week after CBZ administration. There was no evidence of dermatitis or allergic reaction, or other cause for the hair loss. The serum concentration of CBZ was 8.6 microg/ml (therapeutic range 8-12 microg/ml). CBZ was discontinued, and the hair loss stopped within several days with new hair growth. Medication-induced hair loss is an occasional adverse effect of many drugs used for neuropsychological diseases. CBZ also induces hair loss and its frequency was reported below 2%. Only a limited number of detailed case reports describing CBZ induced hair loss were available, and we found these cases could divide into two groups with regard to a delay in starting hair loss after administration of CBZ. In one group, the hair loss started within a week suggesting anagen effluvium and in another it started after two or three months suggesting telogen effluvium. This finding suggests the causative mechanism of CBZ-induced hair loss is not unitary. PMID- 15293765 TI - Depression: underdiagnosed, undertreated, underappreciated. AB - Major depressive disorder is significantly underdiagnosed and undertreated, particularly in the primary care environment. Although more patients are seeking help for depression and the utilization of antidepressants is on the rise, the level of treatment is inadequate. Rectifying this will involve patients, providers, payers, employers, accrediting agencies, and even governmental entities. PMID- 15293766 TI - Prevalence and economic effects of depression. AB - The lifetime risk of major depression among Americans is 17 percent, with as many as 10 percent suffering from depression in any 1-year period. The author reviews the epidemiology of depression, costs of treatment and nontreatment, and its economic impact on quality of life and daily function. This review also examines ways to improve value for money spent relative to this disease. PMID- 15293767 TI - Clinical practice guidelines for treating depression in primary care. AB - Clinical practice guidelines for depression offer the best available treatment options as evidenced by randomized controlled clinical studies. Although the use of evidence-based guidelines has been shown to improve treatment success, there are few data to suggest the extent to which they are used or their effect on treatment outcomes. Keeping guidelines current and increasing the probability of use in daily clinical practice are critical. PMID- 15293768 TI - An overview of SSRI and SNRI therapies for depression. AB - Selective serotonin reuptake inhibitors (SSRIs) and serotonin norepinephrine re uptake inhibitors (SNRIs) are used widely to treat mood and anxiety disorders. Indications, pharmacologic characteristics, and dosing and administration are outlined. Because more patients receive SSRIs in general medical versus psychiatric settings, this chapter includes information relevant to both. PMID- 15293769 TI - Pharmacoeconomic evaluation of antidepressant therapies. AB - With today's emphasis in the managed care environment on evidentiary literature, new antidepressants must provide economic evidence of their value compared with previously available agents. Various pharmacoeconomic models have shown newer antidepressant agents to be cost-effective relative to older agents. The authors review methodologies and published results. PMID- 15293770 TI - Adherence with antidepressant medication. AB - Nonadherence with antidepressant medications is a well-documented factor in treatment failure. This article uses the literature to build a collaborative counseling model to increase antidepressant adherence. Studies may be warranted to determine whether specific antidepressants, when used as part of a collaborative strategy, can improve long-term adherence rates. PMID- 15293771 TI - A review of HEDIS measures and performance for mental disorders. AB - Performance on mental health HEDIS measures has been modest, with only minimal improvements in recent years. Performance on mental health measures has been consistently worse than that for other medical conditions. A critical step toward improving performance is to understand where care is provided and then to identify clinicians who are responsible for ensuring that care is delivered appropriately. PMID- 15293772 TI - Overview and treatment of social anxiety disorder. AB - Social anxiety disorder is a highly prevalent disorder that is frequently comorbid with major depressive disorder. Pharmacotherapies, including SSRIs and benzodiazepines, as well as cognitive-behavioral interventions, are effective for many patients, helping them to attain a higher quality of life. PMID- 15293773 TI - Where have all the teachers gone? PMID- 15293774 TI - Dr. Melis' comments on Lerman's article in the January 2004 issue of CRANIO. PMID- 15293775 TI - Muscle contractions and auditory perception in tinnitus patients and nonclinical subjects. AB - Evidence has been accumulating linking subjective tinnitus to the somatosensory system. Most subjective tinnitus patients can change the psychoacoustic attributes of their tinnitus with forceful head and neck contractions. This study assessed the significance of such somatic modulation of tinnitus by testing nonclinical subjects. Like tinnitus patients, about 80% of nonclinical subjects, who had ongoing tinnitus at the time of testing (whether or not they were previously aware of it), could modulate their tinnitus with head and neck contractions. Over half of those with no tinnitus at the time of testing could elicit a tinnitus-like auditory perception with head and neck contractions. The finding that forceful head and neck contractions, as well as loud sound exposure, were significantly more likely to modulate ongoing auditory perception in people with tinnitus than in those without tinnitus supports the concept of a neural threshold for tinnitus. Somatic influences upon auditory perception are not limited to tinnitus sufferers but appear to be a fundamental property of the auditory system. PMID- 15293776 TI - Assessing orthotic normalization of pharyngeal dynamics. AB - Airway orthotic therapy, considered mainstream in the treatment of sleep disordered breathing, has been demonstrated to normalize both structure and function of the pathological airway through manipulation of mandibular posture. Although effective, the literature reports a variable rate of success and no validated candidacy selection protocol. Acoustic reflection has been used to evaluate and document the upper airway and its dynamics with and without an orthotic in place. This paper will discuss the use of acoustic reflection to assess the level of airway normalization resulting from protrusive and vertical repositioning of the mandible and its utility to establish orthotic candidacy, construction, titration, and maintenance parameters. This protocol has potential for use in both medical and dental facilities that treat patients with sleep disordered breathing. PMID- 15293777 TI - Treatment of joint pain and joint noises associated with a recent TMJ internal derangement: a comparison of an anterior repositioning splint, a full-arch maxillary stabilization splint, and an untreated control group. AB - Pain and joint noises associated with temporomandibular joint (TMJ) internal derangement are often treated by using an intra-oral splint. This study evaluated whether an anterior repositioning splint (AR splint) could be more effective in the treatment of these symptoms than a full-arch maxillary stabilization splint (FAMS splint), because of its capability to re-establish immediately the normal condyle/disk relationship. The authors treated 40 patients (average age 16.8; range 8.0-24.0) with confirmed internal derangement, joint pain, and joint noises in at least one TMJ for at least two months, with AR splint (20 subjects) or FAMS splint (20 subjects); 10 untreated patients comprised the control group. Joint noise, joint pain, and the intensity of pain were assessed using a visual analogic scale (VAS), and the pain was characterized (i.e., constant or chewing/biting pain) and evaluated monthly for eight months. Significantly fewer AR splint patients experienced pain after four months of treatment. A significantly lower intensity of pain was experienced by the AR splint patients after two months of treatment. Significantly fewer AR splint patients experienced chewing/biting pain after eight months of treatment. The frequency of joint noises decreased over time, with no significant differences between the groups. In conclusion, the AR splint seems to be more effective in decreasing pain, but it seems to make no difference in the treatment of joint noises. PMID- 15293778 TI - Parameters of an optimal physiological state of the masticatory system: the results of a survey of practitioners using computerized measurement devices. AB - While bioelectronic instruments have been available for nearly 30 years to assist dentists in day-to-day evaluations of patients' masticatory systems, little guidance has been published to support physiological norms or ideals. An electronic questionnaire was developed and administered to an international group of dentists familiar with the use of bioelectronic instrumentation. Respondents were asked to provide feedback on the norms or ideal parameters of jaw movement, masticatory muscle function with electromyography, and joint sounds through electrosonography that they use in guiding evaluation and treatment of patients with temporomandibular disorders, neuromuscular occlusion, and orthodontics. Surveys were collated to determine areas of consensus. Out of 150 surveys, 55 responses were received from dentists representing nine different countries. Sixty percent of the respondents reported treating more than 150 cases in the past five years using bioelectronic testing. While experience ranged from 2-30 years with different types of devices, average experience was longer with mandibular/jaw tracking (mean 15.3 years) and electromyography (mean 14.1 years) than with electrosonography (mean 7.0 years). Parameters proposed as norms or ideals for electromyographic rest and clench values, and mandibular tracking (velocity, freeway space, and trajectory to closure) were very consistent. Although a smaller number of respondents reported utilization of electrosonography, their criteria for data significance and tissue-type genesis of joint sounds were consistent. While the intra-patient variability may limit the diagnostic use of bioelectronic instruments, the current study demonstrates that through decades of experience, dentists have independently arrived at very consistent definitions of an ideal physiology that can be used to guide treatment. PMID- 15293779 TI - Mood spectrum in patients with different painful temporomandibular disorders. AB - The purpose of this study was to investigate for difference in the prevalence of mood disorders between patients with different painful temporomandibular disorders (TMD). After a sample size necessary for the study was calculated, 60 patients with a painful TMD were selected and divided into the following groups: myofascial pain (n=20), temporomandibular joint (TMJ) pain (n=18), combined myofascial and TMJ pain (n=22). Two distinct comparison groups were selected: subjects with a nonpainful TMD (n=25) and TMD-free subjects (n=29). All participants filled out a self-report validated instrument (MOODS-SR) to evaluate psychopathological symptoms related to mood disturbances. A one-way analysis of variance (ANOVA) with Bonferroni's post hoc test for multiple comparisons was performed to investigate for significant differences among the groups. The three groups of patients with painful TMD scored significantly higher than comparison groups in all MOODS-SR domains investigating depression, but no difference was shown between subjects with myofascial pain and those with TMJ pain. No significant differences among the groups emerged for the presence of manic symptoms, indicating that depressive disorders associated with TMD are not an expression of a more complex manic depressive illness. The study concluded that the presence of depressive symptoms in TMD patients seems to be related to the presence of a painful condition and seems to be unrelated to the location of pain. Furthermore, depressive disturbances in painful TMD patients affect the whole spectrum of depressive psychopathology. PMID- 15293780 TI - A new method for semiautomatic analysis of surface EMG in patients with oral parafunctions. AB - Sleep-related phenomena or disorders, including snoring and tooth grinding, can be investigated using polysomnography. This method, however, generates large amounts of synchronically recorded data that are often analyzed visually with subjective interpretation. The purpose of this study was to minimize the need for subjective evaluation by developing a computer program for analysis of EMG data linked with polysomnographic recordings in a standardized and semi-automatic way. The selected algorithm differs from the Root Mean Square (RMS) method by being based on the theory of "differentiated EMG" (DIFEMG), which relies on two principles. The first says that the activation of a larger number of motor units results in a greater force production. The second principle says that the force production will continue for some time after the muscle is no longer stimulated. After a visual check for artifacts in the basic EMG recordings, the computer program is used to analyze the corrected basic EMG signal. The results were that both methods yield identical results as far as the detected number of events is concerned. There is, however, a significant difference when the duration of the events is considered, because the start and end of an event can be more accurately determined with the new method presented here. The computer program described will make comparison of data from different studies easier. PMID- 15293781 TI - Reproducibility of visual analog scale (VAS) pain scores to mechanical pressure. AB - This study tested the reproducibility of visual analog scale (VAS) pain scores to measure changes in masseter muscle pain evoked by maximally tolerable mechanical stimulation over a short time period in healthy subjects. This study also evaluated gender differences in reproducibility of VAS scores to mechanical stimulation. Ten healthy female and eight healthy male individuals participated in this study. The recordings of VAS pain scores to an identical mechanical pressure on the masseter muscle were performed at three different sessions (T1, T2, and T3). The subjects rated their pain on a VAS to a maximally tolerable stimulus that was recorded on an algometer at the first session. The algometer pressure reading was recorded for each subject and then used to duplicate the same identical mechanical stimulus at each of the three sessions. This identical pressure was repeated in the same marked spot at six minutes and after 30 minutes. The subjects rated the pain on a VAS to this identical stimulus at each session. There was no significant difference in VAS pain scores of all subjects at T1, T2, and T3. There was no significant difference in reproducibility of VAS pain scores in females compared to males. Intraclass correlation coefficients were 0.811 on the right masseter and 0.844 on the left masseter. VAS pain scores to mechanical stimulation were reproducible over a short time period. Gender did not affect the reproducibility. This previously unreported method of measuring pain to repeated identical mechanical stimulation appears to have potential for both clinical and research application. PMID- 15293783 TI - Expression of a transgene encoded on a non-viral episomal vector is not subject to epigenetic silencing by cytosine methylation. AB - Currently available vectors for mammalian cells suffer from a number of limitations which make them only partially useful for genetic modification of eukaryotic cells and organisms and for gene therapy. While integration of a vector can lead to unpredictable interactions with the host genome and silencing of the integrated transgene, most non-integrating vectors mediate only transient expression of a transgene. All available vector types can lead to transformation of the recipient cell and many of them can cause serious immunological side effects in the organism. The ideal vector has to be free of these side effects and should allow long-term expression of a transgene in the absence of selection. In this report we describe a novel non-viral episomal expression system fulfilling these criteria. The gene encoding the truncated rat NGF-receptor gene under the control of the CMV-promoter was inserted into a vector construct containing a scaffold/matrix attached region (S/MAR). This vector was then transfected into CHO cells and human HaCat cells. We show that this vector replicates episomally in these cells and is mitotically stable in the abscence of selection over more than 100 generations. Moreover, we provide the first experimental data that the CMV-promoter in an episome is not subject to silencing by cytosine methylation, thus allowing long-term expression of the transgene in the absence of selection. PMID- 15293782 TI - Caveolae: biochemical analysis. AB - Caveolae appear in a multitude of processes encompassing growth regulation and trafficking. We demonstrate the abundant presence of ESA/reggie-1/flotillin-2, ATP synthase beta subunit and annexin V in endothelial caveolae by immunopurification of caveolae from vascular endothelial membrane. Five proteins are abundant in a caveolin-1 protein complex, analyzed by sucrose gradient velocity sedimentation following octyl-beta-D-glucopyranoside extraction. Caveolin-1 alpha interacts with caveolin-1beta, caveolin-2, actin, the microsomal form of NADH cytochrome B5 reductase and ESA/reggie-1/flotillin-2 as shown by co immunoprecipitation. We propose the concept that ATP biosynthesis in caveolae regulates mechanosignaling and is induced by membrane depolarization and a proton gradient. Pressure stimuli and metabolic changes may trigger gene regulation in endothelial cells, involving a nuclear conformer of caveolin-1, shown here with an epitope-specific caveolin-1 antibody, and immediate response of ion channel activity, regulated by ESA/reggie-1/flotillin-2. PMID- 15293784 TI - Amplification and overexpression of oncogene Mdm2 and orphan receptor gene Nr1h4 in immortal PRKDC knockout cells. AB - DNA-dependent protein kinase (DNA-PK) is required for the repair of double strand DNA breaks by nonhomologous DNA end joining. The catalytic subunit of DNA-PK, PRKDC, may also be involved in repair-related or separate cell signaling pathways. To learn more about the cellular function of DNA-PK under normal physiological conditions, we identified genes that are differentially expressed between an immortalized wild-type mouse fibroblast cell line and its DNA-PK deficient counterpart (Prkdc -/-). The proto-oncogene Mdm2 and the farnesoid X receptor gene Nrlh4 were overexpressed in the DNA-PK-deficient cell line. We show that in the DNA-PK-deficient cell line the genes for both Mdm2 and Nrlh4 are amplified to a degree that could account for most, if not all, of their increased expression. Other genes were strongly downregulated in the DNA-PK-deficient cell line, but this opposite expression pattern was not due to gene amplification in the wild-type cells. None of these genes was differentially expressed in DNA-PK containing and DNA-PK-deficient primary mouse embryo fibroblasts. Our results suggest a model in which DNA-PK indirectly affects the cellular gene expression profile through its caretaker role and by preventing gene amplification. PMID- 15293785 TI - Refining the DFNB17 interval in consanguineous Indian families. AB - We previously mapped the DFNB17 locus to a 3-4 cM interval on human chromosome 7q31 in a large consanguineous Indian family with congenital profound sensorineural hearing loss. To further refine this interval, 30 new highly polymorphic markers and 8 SNPs were analyzed against the pedigree. Re-analysis in the original DFNB 17 family and additional data from a second unrelated consanguineous family with congenital deafness found to map to the interval, limited the area of shared homozygosity-by-descent (HBD) to approximately 4 megabase (Mb) between markers D7S2453 and D7S525. Nineteen known genes and over 20 other cDNAs have been identified in the refined DFNB 17 interval, including the SLC26A4 gene. We have analyzed 4 other cochlear-expressed genes that map to the DFNB17 interval as candidate genes. Analysis of coding and splice site regions of these cochlear expressed genes did not reveal any disease causing mutations. Further study of other candidate genes is currently underway. PMID- 15293786 TI - Inverted Gcg/CGC trinucleotide microsatellites in the 5'-region of Mus IDS mRNA: recurrent induction of aberrant reverse transcripts. AB - An investigation was initiated to explore previously published results indicating that approximately 80 bp of the 5'-end of the iduronate sulfatase (IDS) cDNA sequence (Accession No L07291) are 100% homologous with the 3'-UTR of isoform I of the sodium hydrogen exchanger (Acc. No. U51112). 5'-RACE carried out on IDS mRNA demonstrated the apparent homology to be a cloning artifact. A sequence comparison of the IDS 5'-RACE product with a mouse BAC clone covering the region, and with various IDS ESTs, suggested that the region is highly susceptible to cloning artifacts, a common one of which is template switching by reverse transcriptase. The nucleotide sequence flanking the translation start site is unusual in containing two inverted repeats composed of the complementary trinucleotide microsatellites, (GCG)9 and (CGC)6. These likely form a highly stable stem of 20-21 nt, through which reverse transcription is compromised. Such a stem could be involved in the regulation of IDS expression by directly affecting translation, message turnover, or serving as a substrate for siRNA production. Though such mRNA features are relatively rare, they may be more abundant but overlooked due to difficulties in their reverse transcription. PMID- 15293787 TI - Chitinase induced by jasmonic acid, methyl jasmonate, ethylene and protein phosphatase inhibitors in rice. AB - Chitinase is a pathogenesis-related protein that hydrolyzes chitin, a major component of fungal cell walls. Two-week-old rice seedling leaf, leaf sheath and root tissues responded to an exogenous treatment by jasmonic acid (JA) with induction of the chitinases as determined by immunoblot analysis using an anti endochitinase antibody. Induced accumulation of these chitinases was observed within 24 to 48 h in the leaf sheaths, leaves and roots. Besides, ethylene generator ethephon and abiotic stressor copper could also induce chitinases accumulation among various plant hormones and stress agents examined. Cycloheximide effectively blocked their accumulation by JA, suggesting that de novo protein synthesis is required. Partial blockage of the induced accumulation of chitinases by NADPH oxidase inhibitor and free radical scavengers suggested involvement of reactive oxygen species. Moreover, induced accumulation of these chitinases also by methyl jasmonate and certain protein phosphatase inhibitors indicated their potential importance and wider role in rice seedlings. PMID- 15293788 TI - Biodistribution of filamentous phage peptide libraries in mice. AB - In vivo phage display is a new approach to acquire peptide molecules that bind stably to a given target. Phage peptide display libraries have been selected in mice and humans and numerous vasculature-targeting peptides have been reported. However, in vivo phage display has not typically produced molecules that extravasate to target specific organ or tumor antigens. Phage selections in animals have been performed for very short times without optimization for biodistribution or clearance rates to a particular organ. It is hypothesized that peptides that home to a desired antigen/organ can be obtained from in vivo phage experiments by optimization of incubation times, phage extraction and propagation procedures. To accomplish this goal, one must first gain a better understanding of the in vivo biodistribution and rate of clearance of engineered phage peptide display libraries. While the fate of wild type phage in rodents has been reported, the in vivo biodistribution of the commonly used engineered fd-tet M13 phage peptide display libraries (such as in the fUSE5 vector system) have not been well established. Here we report the biodistribution and clearance properties of fd-tet fifteen amino acid random peptide display libraries in fUSE5 phage in three common mouse models employed for drug discovery - CF-1, nude, and SCID mice. PMID- 15293789 TI - V-erba homodimers mediate the potent dominant negative activity of v-erba on everted repeats. AB - The oncoprotein v-erbA is a mutated form of TRalpha1 that is unable to bind thyroid hormone (T3). V-erbA homodimerizes or heterodimerizes with retinoid X receptor (RXR) on core motifs arranged as direct, everted, or inverted repeats (DRs, ERs, or IRs). We created a series of v-erbA mutants in order to obtain a better understanding of the role of v-erbA homodimers versus v-erbA-RXR heterodimers in the dominant negative activity of v-erbA on ERs (the most potent v-erbA response elements). We found that one of these mutants, v-erbA mutant E325A, is able to homodimerize but unable to heterodimerize with RXR on ERs. Our data also suggest that v-erbA homodimers interact preferentially with the corepressor NCoR over SMRT and that the interaction with corepressors is stronger with v-erbA homodimers over v-erbA-RXR heterodimers. Furthermore, functional studies showed that v-erbA homodimers rather than v-erbA-RXR heterodimers mediate the dominant negative activity of v-erbA on ERs. PMID- 15293790 TI - Bsu2413I and Bfi2411I, two new thermophilic type II restriction endonucleases from Bacillus subtilis and Bacillus firmus: isolation and partial purification. Thermophilic endonucleases from two Bacillus species. AB - Two new thermophilic type II restriction endonucleases, which we designated as Bsu2413I and Bfi2411I, have been isolated from gram-positive thermophilic bacteria Bacillus subtilis strain 2413 and Bacillus firmus strain 2411 respectively and partially purified. The restriction endonucleases were extracted from cell extracts and purified using single step purification through phosphocellulose column chromatography. SDS-PAGE profile showed denatured molecular weights of 33 and 67 kDa for the Bsu2413I and 39 and 67 kDa for the Bfi2411I. The partially purified Bsu2413I enzyme restricted pBR322 DNA into two fragments of 3250 and 1100 bp whereas Bfi2411I enzyme restricted pBR322 DNA into two fragments of 3500 and 800 bp. The activity of both endonucleases was assayed at 55 degrees C and they required Mg+2 as cofactor like other type II restriction endonucleases. PMID- 15293791 TI - [Atherogenesis as an immunoinflammatory process]. AB - The paper contains findings, obtained at the laboratory of atherosclerosis of Anichkov's Research Institute of Experimental Medicine, Russia's Academy of Medical Sciences, during the last decade with the above research results being compared with published data. Atherogenesis is discussed from the standpoint of the development of immune inflammation in the arterial wall. The mechanisms and factors triggering the chain of immunoinflammatory reactions, which provoke and keep up the inflammatory process, are also under discussion. The results are indicative of that the atherosclerosis pathogenesis is equally related both with mLDL and with the reaction developing in the vascular wall. PMID- 15293792 TI - [Social psychiatry as a priority trend in psychiatric science and practice]. PMID- 15293793 TI - [Phototherapy and its place in modern medicine]. AB - The notion of "phototherapy" is defined in the paper; the evolution stages of the above promising medicine trend are described; variants and methods of its application as well as action mechanisms of different optic-irradiation ranges on human body are elucidated. Medical fields, in which phototherapy can be used, are specified. PMID- 15293794 TI - [Oxidative stress in the pathogenesis of anti-phospholipid syndrome]. AB - Summarized in the paper are the data describing the significance of the oxidative stress within the pathogenesis of anti-phospholipid syndrome (APS). The mechanisms related with formation of active oxygen metabolites in human body, triggers of the oxidative stress as well as its involvement in the APS pathogenesis (i.e. in the aPL synthesis), in the endothelial malfunction development and in the development of pro-coagulation impairments within the hemostasis system leading to thrombosis are in the focus of attention. Approaches to treatment schemes of vascular disorders in APS comprising the oxidative stress correction are suggested. PMID- 15293795 TI - [Penetration of lipoproteins and apolipoproteins of very low density into the myocardium and changes of its structure in rat heart perfusion]. AB - The method of electron microscopy, involving extra-low density lipoproteins (ELDLP) marked by colloid gold as well as their protein components (i.e. apolipoproteins--apoELDLP), was made use of to show that they are captured by capillary endotheliocytes of the isolated perfusing rat heart. Receptor-mediated endocytosis is the main mechanism of penetration. Within 30 minutes, ELDLP interact with the capillary wall, pass through the endothelial barrier and, from the intersticium, they are captured by tissue microphages, which induces their activity. They have, on the whole, a positive effect on the intactness of muscle cells of the beating heart. During the same time span, apoELDLP remain in endotheliocytes, however, they exert a pronounced negative impact on the myocardium. The capillary endothelium, in whose cells the lysosomal apparatus activates itself and clasmatosis sets on, is affected morphologically most of all. The impairment of the capillary endothelium, development of the perivascular edema and a reduced coronary flow trigger a sequence of events leading to enhanced cytolytic processes in the myocardium due to an activated lysosomal apparatus of cells. PMID- 15293796 TI - [Physiological reasoning and results of using the functional electrostimulation of muscles in pathological walking]. AB - The ideology of a method of muscle electrostimulation (MES) in walking up and down the stairs is under discussion in the paper. It is pointed out that the above locomotor acts, which are based on the central innervation program, are principally similar; periods and zones of muscle excitation and inhibition during the locomotor cycle are specified; MES is shown to be a variety of phase muscle electrostimulation. A hypothesis is put forward on the MES significance in normalizing the locomotor skills. The main MES function are described, i.e. therapy, diagnostics and prognostication. The key indication for MES administration in pathological walking is defined, i.e. a deficit of the muscle function (DMF). A new method is suggested for DMF determination through examining the muscle-force moments estimated by mathematic modeling. The MES technique is shown to comprise 5 main stages: choice of corrected movements and muscle stimulation as well as setting-up of the amplitude and temporal MES parameters, of electrode parameters and of a stimulation regime in walking. Medical contraindications to MES stages of patients' rehabilitation are defined. The results of a successful treatment of above 6000 patients with different pathological conditions of the support-motor apparatus are summarized. PMID- 15293797 TI - [Mechanisms of psychophysiologic regulation of the protein composition in the mixed human saliva]. AB - As everybody knows, human saliva contains a huge variety of different proteins. The protein composition of the mixed saliva was found to depend to a certain extent on a psycho-emotional condition. The main elements of mechanisms of the psychophysiologic regulation of the protein composition of the mixed saliva are under discussion in the paper. A hypothesis is put forward, which envisages a possible different-type joint activity of the vegetative nervous centers controlling the saliva glands as triggered by emotions. Besides, issues related with the fundamental research as well as outlooks for using the analysis of the protein composition of the mixed saliva for the purpose of investigating the human psychophysiology and of coping with practical medical tasks are also under discussion. PMID- 15293798 TI - [The pathogenetic role of the infection component in children with lymphadenopathy under modern conditions]. AB - Apart from lymphadenopathy types observed in well-known diseases (oncology and hematology pathologies, systemic lesions of connective tissues, tuberculosis, brucellosis, infection mononucleosis, toxoplasmosis, AIDS and other specific processes), there is an extensive group of lymphadenopathy (up to 70% of all disease cases), whose cause cannot be diagnosed by using the routine examination schemes. This was reflected in the diagnosis of "lymphadenopathy of unverified etiology" according to the MKB-1 now in force. Investigations of the role (in etiopathogenesis) of unverified lymphadenopathy types as observed in children with the infection component as well as optimization of their diagnosis and treatment schemes are elucidated in the paper. It is substantiated, on the basis of published data and authors' independent research, that some of lymphadenopathy types of unverified etiology are related with such opportunistic infections, like clamidiosis, mycoplasmosis, herpes simplex and cytomegalovirus infection. The above stated is, primarily, compatible with the tropicity of such infections' agents to the lymphoid tissue and/or with their possible lymphogenic spread as well as with polymorphism of clinical signs conditioned by the discussed infection agents. PMID- 15293799 TI - [Some approaches to elaborating vaccines against the HIV infection]. AB - A variety of approaches to creating vaccines against the HIV-infection are discussed in the paper. It is demonstrated that vaccines can be used both for treatment and prevention. They can be synthetic, DNA-vaccines and vector-type ones based on the attenuated recombinant strain of Salmonella. Apart from gp120, reverse transcriptase and tat-gene products can be used as the antigen. PMID- 15293800 TI - [Synchronism of a many-year dynamics of morbidity]. AB - Synchronism of many-year morbidity fluctuations is discussed for different territories. The methods of mathematic modeling revealed that the exchange of the low-intensity infections, in populations with identical conditions results in a virtual complete morbidity dynamics, whereas, the effect from synchronism is by far less pronounced for territories with differing activity values of the transfer mechanism. PMID- 15293801 TI - Cellular and molecular mechanisms of regeneration in Xenopus. AB - We have employed transgenic methods combined with embryonic grafting to analyse the mechanisms of regeneration in Xenopus tadpoles. The Xenopus tadpole tail contains a spinal cord, notochord and segmented muscles, and all tissues are replaced when the tail regenerates after amputation. We show that there is a refractory period of very low regenerative ability in the early tadpole stage. Tracing of cell lineage with the use of single tissue transgenic grafts labelled with green fluorescent protein (GFP) shows that there is no de-differentiation and no metaplasia during regeneration. The spinal cord, notochord and muscle all regenerate from the corresponding tissue in the stump; in the case of the muscle the satellite cells provide the material for regeneration. By using constitutive or dominant negative gene products, induced under the control of a heat shock promoter, we show that the bone morphogenetic protein (BMP) and Notch signalling pathways are both essential for regeneration. BMP is upstream of Notch and has an independent effect on regeneration of muscle. The Xenopus limb bud will regenerate completely at the early stages but regenerative ability falls during digit differentiation. We have developed a procedure for making tadpoles in which one hindlimb is transgenic and the remainder wild-type. This has been used to introduce various gene products expected to prolong the period of regenerative capacity, but none has so far been successful. PMID- 15293802 TI - Fibroblast growth factors in epithelial repair and cytoprotection. AB - Growth factors are polypeptides that stimulate the division of certain cell types at low concentrations. Fibroblast growth factor (FGF) 7 (FGF-7) and its homologue FGF-10 act specifically on various types of epithelial cells including keratinocytes of the skin, intestinal epithelial cells and hepatocytes. In addition, FGF-7 and FGF-10 have been shown to be more than growth factors: they can protect epithelial cells from damaging effects induced, for example, by radiation and oxidative stress. Therefore, they are currently in clinical trials for the treatment of oral mucositis, a severe side-effect of cancer therapy characterized by painful inflammation and ulceration of the oral epithelium. To gain insight into the mechanisms of FGF-7/FGF-10 action in epithelial cells, we searched for genes that are regulated by these growth factors. Indeed, we identified genes that help us to explain the mechanisms that underlie the effects of FGF-7. Most interestingly, several genes were identified that are likely to mediate the cytoprotective effect of FGF-7 for epithelial cells in vitro and possibly also in injured and diseased tissues in vivo. PMID- 15293803 TI - Regeneration and the need for simpler model organisms. AB - The problem of regeneration is fundamentally a problem of tissue homeostasis involving the replacement of cells lost to normal 'wear and tear' (cell turnover), and/or injury. This attribute is of particular significance to organisms possessing relatively long lifespans, as maintenance of all body parts and their functional integration is essential for their survival. Because tissue replacement is broadly distributed among multicellular life-forms, and the molecules and mechanisms controlling cellular differentiation are considered ancient evolutionary inventions, it should be possible to gain key molecular insights about regenerative processes through the study of simpler animals. We have chosen to study and develop the freshwater planarian Schmidtea mediterranea as a model system because it is one of the simplest metazoans possessing tissue homeostasis and regeneration, and because it has become relatively easy to molecularly manipulate this organism. The developmental plasticity and longevity of S. mediterranea is in marked contrast to its better-characterized invertebrate cohorts: the fruitfly Drosophila melanogaster and the roundworm Caenorhabditis elegans, both of which have short lifespans and are poor at regenerating tissues. Therefore, planarians present us with new, experimentally accessible contexts in which to study the molecular actions guiding cell fate restriction, differentiation and patterning, each of which is crucial not only for regeneration to occur, but also for the survival and perpetuation of all multicellular organisms. PMID- 15293804 TI - A critical role for thrombin in vertebrate lens regeneration. AB - Lens regeneration in urodele amphibians such as the newt proceeds from the dorsal margin of the iris where pigment epithelial cells (PEC) re-enter the cell cycle and transdifferentiate into lens. A general problem in regeneration research is to understand how the events of tissue injury or removal are coupled to the activation of plasticity in residual differentiated cells or stem cells. Thrombin, a pivotal regulator of the injury response, has been implicated as a regulator of cell cycle re-entry in newt myotubes, and also in newt iris PEC. After removal of the lens, thrombin was activated on the dorsal margin for 5-7 days. Inactivation of thrombin by either of two different inhibitors essentially blocked S-phase re-entry by PEC at this location. The axolotl, a related species which can regenerate its limb but not its lens, can activate thrombin after amputation but not after lens removal. These data support the hypothesis that thrombin is a critical signal linking injury to regeneration, and offer a new perspective on the evolutionary and phylogenetic questions about regeneration. PMID- 15293805 TI - Wound healing and inflammation: embryos reveal the way to perfect repair. AB - Tissue repair in embryos is rapid, efficient and perfect and does not leave a scar, an ability that is lost as development proceeds. Whereas adult wound keratinocytes crawl forwards over the exposed substratum to close the gap, a wound in the embryonic epidermis is closed by contraction of a rapidly assembled actin purse string. Blocking assembly of this cable in chick and mouse embryos, by drugs or by inactivation of the small GTPase Rho, severely hinders the re epithelialization process. Live studies of epithelial repair in GFP-actin expressing Drosophila embryos reveal actin-rich filopodia associated with the cable, and although these protrusions from leading edge cells appear to play little role in epithelial migration, they are essential for final zippering of the wound edges together-inactivation of Cdc42 prevents their assembly and blocks the final adhesion step. This wound re-epithelialization machinery appears to recapitulate that used during naturally occurring morphogenetic episodes as typified by Drosophila dorsal closure. One key difference between embryonic and adult repair, which may explain why one heals perfectly and the other scars, is the presence of an inflammatory response at sites of adult repair where there is none in the embryo. Our studies of repair in the PU. 1 null mouse, which is genetically incapable of raising an inflammatory response, show that inflammation may indeed be partly responsible for scarring, and our genetic studies of inflammation in zebrafish (Danio rerio) larvae suggest routes to identifying gene targets for therapeutically modulating the recruitment of inflammatory cells and thus improving adult healing. PMID- 15293806 TI - The scarless heart and the MRL mouse. AB - The ability to regenerate tissues and limbs in its most robust form is seen in many non-mammalian species. The serendipitous discovery that the MRL mouse has a profound capacity for regeneration in some ways rivalling the classic newt and axolotl species raises the possibility that humans, too, may have an innate regenerative ability. The adult MRL mouse regrows cartilage, skin, hair follicles and myocardium with near perfect fidelity and without scarring. This is seen in the ability to close through-and-through ear holes, which are generally used for lifelong identification of mice, and the anatomic and functional recovery of myocardium after a severe cryo-injury. We present histological, biochemical and genetic data indicating that the enhanced breakdown of scar-like tissue may be an underlying factor in the MRL regenerative response. Studies as to the source of the cells in the regenerating MRL tissue are discussed. Such studies appear to support multiple mechanisms for cell replacement. PMID- 15293807 TI - Genetic approaches to disease and regeneration. AB - Cardiovascular disease is largely a consequence of coronary artery blockage through excessive proliferation of smooth muscle cells. It in turn leads to myocardial infarction and permanent and functionally devastating tissue damage to the heart wall. Our studies have revealed that elastin is a primary player in maintaining vascular smooth muscle cells in their dormant state and thus may be a useful therapeutic in vascular disease. By studying zebrafish, which unlike humans, can repair damage to heart muscle, we have begun to uncover some of the genes that seem necessary to undertake the de-differentiation steps that currently fail and prevent the formation of new proliferating cardiomyocytes at the site of damage in a mammalian heart. PMID- 15293808 TI - Retinoic acid in alveolar development, maintenance and regeneration. AB - Recent data suggest that exogenous retinoic acid (RA), the biologically active derivative of vitamin A, can induce alveolar regeneration in a rat model of experimental emphysema. Here, we describe a mouse model of disrupted alveolar development using dexamethasone administered postnatally. We show that the effects of dexamethasone are concentration dependent, dose dependent, long lasting and result in a severe loss of alveolar surface area. When RA is administered to these animals as adults, lung architecture and the surface area per unit of body weight are completely restored to normal. This remarkable effect may be because RA is required during normal alveolar development and administering RA re-awakens gene cascades used during development. We provide evidence that RA is required during alveologenesis in the mouse by showing that the levels of the retinoid binding proteins, the RA receptors and two RA synthesizing enzymes peak postnatally. Furthermore, an inhibitor of RA synthesis, disulphiram, disrupts alveologenesis. We also show that RA is required throughout life for the maintenance of lung alveoli because when rats are deprived of dietary retinol they lose alveoli and show the features of emphysema. Alveolar regeneration with RA may therefore be an important novel therapeutic approach to the treatment of respiratory diseases characterized by a reduced gas-exchanging surface area such as bronchopulmonary dysplasia and emphysema for which there are currently no treatments. PMID- 15293809 TI - Exploring the mechanisms regulating regeneration of deer antlers. AB - Deer antlers are the only mammalian appendages capable of repeated rounds of regeneration; every year they are shed and regrow from a blastema into large branched structures of cartilage and bone that are used for fighting and display. Longitudinal growth is by a process of modified endochondral ossification and in some species this can exceed 2 cm per day, representing the fastest rate of organ growth in the animal kingdom. However, despite their value as a unique model of mammalian regeneration the underlying mechanisms remain poorly understood. We review what is currently known about the local and systemic regulation of antler regeneration and some of the many unsolved questions of antler physiology are discussed. Molecules that we have identified as having potentially important local roles in antlers include parathyroid hormone-related peptide and retinoic acid (RA). Both are present in the blastema and in the rapidly growing antler where they regulate the differentiation of chondrocytes, osteoblasts and osteoclasts in vitro. Recent studies have shown that blockade of RA signalling can alter cellular differentiation in the blastema in vivo. The trigger that regulates the expression of these local signals is likely to be changing levels of sex steroids because the process of antler regeneration is linked to the reproductive cycle. The natural assumption has been that the most important hormone is testosterone, however, at a cellular level oestrogen may be a more significant regulator. Our data suggest that exogenous oestrogen acts as a 'brake', inhibiting the proliferation of progenitor cells in the antler tip while stimulating their differentiation, thus inhibiting continued growth. Deciphering the mechanism(s) by which sex steroids regulate cell-cycle progression and cellular differentiation in antlers may help to address why regeneration is limited in other mammalian tissues. PMID- 15293810 TI - Stem cells: cross-talk and developmental programs. AB - The thesis advanced in this essay is that stem cells-particularly those in the nervous system-are components in a series of inborn 'programs' that not only ensure normal development, but persist throughout life so as to maintain homeostasis in the face of perturbations-both small and great. These programs encode what has come to be called 'plasticity'. The stem cell is one of the repositories of this plasticity. This review examines the evidence that interaction between the neural stem cell (as a prototypical somatic stem cell) and the developing or injured brain is a dynamic, complex, ongoing reciprocal set of interactions where both entities are constantly in flux. We suggest that this interaction can be viewed almost from a 'systems biology' vantage point. We further advance the notion that clones of exogenous stem cells in transplantation paradigms may not only be viewed for their therapeutic potential, but also as biological tools for 'interrogating' the normal or abnormal central nervous system environment, indicating what salient cues (among the many present) are actually guiding the expression of these 'programs'; in other words, using the stem cell as a 'reporter cell'. Based on this type of analysis, we suggest some of the relevant molecular pathways responsible for this 'cross-talk' which, in turn, lead to proliferation, migration, cell genesis, trophic support, protection, guidance, detoxification, rescue, etc. This type of developmental insight, we propose, is required for the development of therapeutic strategies for neurodegenerative disease and other nervous system afflictions in humans. Understanding the relevant molecular pathways of stem cell repair phenotype should be a priority, in our view, for the entire stem cell field. PMID- 15293811 TI - Scar-free healing: from embryonic mechanisms to adult therapeutic intervention. AB - In man and domestic animals, scarring in the skin after trauma, surgery, burn or sports injury is a major medical problem, often resulting in adverse aesthetics, loss of function, restriction of tissue movement and/or growth and adverse psychological effects. Current treatments are empirical, unreliable and unpredictable: there are no prescription drugs for the prevention or treatment of dermal scarring. Skin wounds on early mammalian embryos heal perfectly with no scars whereas wounds to adult mammals scar. We investigated the cellular and molecular differences between scar-free healing in embryonic wounds and scar forming healing in adult wounds. Important differences include the inflammatory response, which in embryonic wounds consists of lower numbers of less differentiated inflammatory cells. This, together with high levels of morphogenetic molecules involved in skin growth and morphogenesis, means that the growth factor profile in a healing embryonic wound is very different from that in an adult wound. Thus, embryonic wounds that heal without a scar have low levels of TGFbeta1 and TGFbeta2, low levels of platelet-derived growth factor and high levels of TGFbeta3. We have experimentally manipulated healing adult wounds in mice, rats and pigs to mimic the scar-free embryonic profile, e.g. neutralizing PDGF, neutralizing TGFbeta1 and TGFbeta2 or adding exogenous TGFbeta3. These experiments result in scar-free wound healing in the adult. Such experiments have allowed the identification of therapeutic targets to which we have developed novel pharmaceutical molecules, which markedly improve or completely prevent scarring during adult wound healing in experimental animals. Some of these new drugs have successfully completed safety and other studies, such that they have entered human clinical trials with approval from the appropriate regulatory authorities. Initial trials involve application of the drug or placebo in a double-blind randomized design, to experimental incision or punch biopsy wounds under the arms of human volunteers. Based on encouraging results from such human volunteer studies, the lead drugs have now entered human patient-based trials e.g. in skin graft donor sites. We consider the evolutionary context of wound healing, scarring and regeneration. We hypothesize that evolutionary pressures have been exerted on intermediate sized, widespread, dirty wounds with considerable tissue damage e.g. bites, bruises and contusions. Modem wounds (e.g. resulting from trauma or surgery) caused by sharp objects and healing in a clean or sterile environment with close tissue apposition are new occurrences, not previously encountered in nature and to which the evolutionary selected wound healing responses are somewhat inappropriate. We also demonstrate that both repair with scarring and regeneration can occur within the same animal, including man, and indeed within the same tissue, thereby suggesting that they share similar mechanisms and regulators. Consequently, by subtly altering the ratio of growth factors present during adult wound healing, we can induce adult wounds to heal perfectly with no scars, with accelerated healing and with no adverse effects, e.g. on wound strength or wound infection rates. This means that scarring may no longer be an inevitable consequence of modem injury or surgery and that a completely new pharmaceutical approach to the prevention of human scarring is now possible. Scarring after injury occurs in many tissues in addition to the skin. Thus scar-improving drugs could have widespread benefits and prevent complications in several tissues, e.g. prevention of blindness after scarring due to eye injury, facilitation of neuronal reconnections in the central and peripheral nervous system by the elimination of glial scarring, restitution of normal gut and reproductive function by preventing strictures and adhesions after injury to the gastrointestinal or reproductive systems, and restoration of locomotor function by preventing scarring in tendons and ligaments. PMID- 15293813 TI - The Cochrane Oral Health Group. PMID- 15293812 TI - Stem cell biology and neurodegenerative disease. AB - The fundamental basis of our work is that organs are generated by multipotent stem cells, whose properties we must understand to control tissue assembly or repair. Central nervous system (CNS) stem cells are now recognized as a well defined population of precursors that differentiate into cells that are indisputably neurons and glial cells. Work from our group played an important role in defining stem cells of the CNS. Embryonic stem (ES) cells also differentiate to specific neuron and glial types through defined intermediates that are similar to the cellular precursors that normally occur in brain development. There is convincing evidence that the differentiated progeny of ES cells and CNS stem cells show expected functions of neurons and glia. Recent progress has been made on three fundamental developmental processes: (i) cell cycle control; (ii) the control of cell fate; and (iii) early steps in neural differentiation. In addition, our work on CNS stem cells has developed to a stage where there are clinical implications for Parkinson's and other degenerative disorders. These advances establish that stem cell biology contributes to our understanding of brain development and has great clinical promise. PMID- 15293814 TI - Ceramic inlays for restoring posterior teeth. PMID- 15293815 TI - An historical perspective on early progress of Queensland water fluoridation 1945 1954: sheep, climate and sugar. AB - BACKGROUND: Queensland's virtual rejection of artificial water fluoridation sets it apart from other Australian states, yet the early fluoride environs has been scantily recorded. METHODS: This paper used archives, literature review, personal interview and the traditional historic method. RESULTS: The connection between Queensland artesian bore water and caries resistance was postulated as early as 1912. Four decades later, two Queensland-specific factors influenced the planning to fluoridate community water supplies. The first (1945-1950) was confusion between the high levels of fluoride in artesian water supplying the pastoral industry and the scientific concept of artificial water fluoridation of communal supplies. The second (1952-1954) involved further scientific investigation involving water consumption patterns, occupational dehydration and fluid homeostasis within a sub-tropical climate. The role of the Australian Dental Association Queensland Branch (ADAQ) in early fluoride politics was minimal. Four early protagonists are identified--two dentists, an engineer and the sugar industry. CONCLUSIONS: Queensland had its advocates for artificial water fluoridation of communal supply as a means of caries prevention. Interest came from the dental, medical and engineering professions, and from the sugar industry. However, these efforts met with indifference based on confused extrapolation of the artesian experience (1945-1952) and hesitancy (1952-1954) due to contemporaneous concerns about human fluid homeostasis in Queensland's sub tropical climate. PMID- 15293816 TI - Diagnostic comparison of three groups of examiners using visual and laser fluorescence methods to detect occlusal caries in vitro. AB - BACKGROUND: To evaluate the accuracy of the DIAGNOdent laser device (DD) for detecting occlusal fissure caries when used by three groups of examiners. METHODS: Three final-year dental students (S), three General Dental Practitioners (G), and three Academic Clinicians (A) individually examined the non-cavitated occlusal surfaces of 25 extracted permanent molars using visual inspection (VI) then DD assessments. The presence of caries was confirmed following tooth sectioning. A cut-off limit of 30 was used for the DD to avoid over-treatment in a low caries-risk situation. RESULTS: For VI, individual examiner sensitivity (caries correctly diagnosed) ranged from 53 to 86 per cent, and specificity (sound teeth correctly diagnosed) ranged from 76 to 95 per cent, with low Kappa agreements. Group S achieved the highest sensitivity (80 per cent) and Groups G and A achieved the highest specificities (88 per cent). For DD, individual examiner sensitivity ranged from 19DD, individual examiner sensificity ranged from 19 to 77 per cent, and specificity from 71 to 97 per cent, with generally moderate Kappa agreements. Group A achieved the highest (67 per cent) and Group G the lowest (44 per cent) sensitivities, and Group G achieved the highest specificity (94 per cent). CONCLUSIONS: There were similar widely varying results for the two diagnostic methods and for the three groups of examiners. However, the relatively high sensitivities found with VI and specificities found with DD should avoid over-treatment in low caries-risk populations. PMID- 15293817 TI - Morphological analysis of subgingival biofilm formation on synthetic carbonate apatite inserted into human periodontal pockets. AB - BACKGROUND: Details of the development of human subgingival biofilm are unknown due to the difficulties in conducting experiments and especially in obtaining undisturbed materials. METHODS: This study was performed using deposits on carbonate apatite that had been inserted into human periodontal pockets for up to three weeks. Scanning electron microscopy using the vertically sectioned method and transmission electron microscopy using the freeze-substitution method were adopted. RESULTS: The development of subgingival biofilm occurred in five sequential phases: pellicle formation, microbial adherence, initial colonization, microbial organization, and establishment. Certain species in each of the initial, secondary and tertiary colonizers were considered to have a predilection for biofilm formation. Gram-positive, bacillary initial colonizers and gram negative, filamentous secondary colonizers organized one stable structure that served as the framework for biofilm formation, and gram-negative, rod-shaped tertiary colonizers with cell-surface vesicles showed multigeneric coaggregation. The microbiota in the tertiary colonizers underwent repeated microflora alteration. CONCLUSIONS: Subgingival biofilm is constituted by initial, secondary and tertiary colonizers. Microflora alteration which is suggested to be related to periodontal disease, frequently occurred in the tertiary colonizers. PMID- 15293818 TI - The effects of a tea tree oil-containing gel on plaque and chronic gingivitis. AB - BACKGROUND: [corrected] This clinical study assessed the effects of topically applied tea tree oil (TTO)-containing gel on dental plaque and chronic gingivitis. METHODS: This was a double-blind, longitudinal, non-crossover study in 49 medically fit non-smokers (24 males and 25 females) aged 18-60 years with severe chronic gingivitis. Subjects were randomly assigned to three groups and given either TTO-gel (2.5 per cent), chlorhexidine (CHX) gel (0.2 per cent), or a placebo gel to apply with a toothbrush twice daily. Treatment effects were assessed using the Gingival Index (GI), Papillary Bleeding Index (PBI) and plaque staining score (PSS) at four and eight weeks. RESULTS: No adverse reactions to any of the gels were reported. The data were separated into subsets by tooth (anterior and posterior) and tooth surface (buccal and lingual). The TTO group had significant reduction in PBI and GI scores. However,,TTO did not reduce plaque scores, which tended to increase over the latter weeks of the study period. CONCLUSION: Although further studies are required, the anti-inflammatory properties of TTO-containing gel applied topically to inflamed gingival tissues may prove to be a useful non-toxic adjunct to chemotherapeutic periodontal therapy. PMID- 15293819 TI - Psychic and occlusal factors in bruxers. AB - BACKGROUND: The aim of this study was to investigate the existence of associations between bruxism and psychic and occlusal factors. METHODS: Participants in this study (n=85) were recruited from the Section of Odontostomatology, Deparent of Neuroscience, University of Pisa, Italy. They were split into two groups, bruxers (n=34) and non-bruxers (n=51), on the basis of the presence of both clinical and anamnestical indicators of bruxim. All participants were administered two self-reported validated questionnaires to evaluate (MOODS SR) and panic-agoraphobic (PAS-SR) spetra. A number of occlusal variables (deep bite: cross-bite, open-bite, mediotrusive and laterotrusive interferences, slide RCP-ICP, laterotrutsive guides, canine and molar classes) were also recorded. RESULTS: With regards to occlusal factors, the only association was revealed between bruxism mediotrusive interferences (p < 0.05). As for psychiatric investigation, significant differences between bruxers and controls emerged for the presence of both depressive (p < 0.01) and manic symptoms (p < 0.01) in MOODS SR, and for stress sensitivity (p < 0.01), anxious expectation (p < 0.05), and reassurance sensitivity symptoms (p < 0.05) in PAS-SR. In particular, both mood (p < 0.01) and panic-agoraphobic (p < 0.05) spectra symptoms seem to differentiate bruxers from controls in males, while in females strong differences emerged for stress sensitivity symptoms (p < 0.05). CONCLUSIONS: It can be confirmed that certain psychic traits are present in bruxers, while occlusal factors are not useful parameters to discern bruxers from non-bruxers. PMID- 15293820 TI - A rare presentation of dens invaginatus in a mandibular lateral incisor occurring concurrently with bilateral maxillary dens invaginatus: case report and review of literature. AB - BACKGROUND: Dens Invaginatus (DI), commonly known as dens in dente, is a developmental malformation of tooth that most commonly affects permanent maxillary lateral incisor teeth. Deciduous teeth are infrequently affected. Presence of DI in mandibular permanent teeth is extremely rare. Further, the presence of DI bilaterally in the maxillary lateral incisors of the same patient is even more unusual. METHODS: In this article, an unusual case of DI affecting a mandibular lateral incisor tooth is described. This malformation was uncovered after a full mouth radiographic examination when the patient presented for dental treatment unrelated to this finding. In addition, the various radiographic appearances of DI as they present within the maxillary and mandibular teeth are described. Essential clinical considerations and treatment options are presented. A review of the pertinent literature is undertaken and a table summarizing previous published findings of mandibular DI is presented. RESULTS: A review of the literature indicates that DI in mandibular teeth is extremely rare with only 10 other cases involving 13 teeth reported previously. CONCLUSIONS: Although this is an extremely rare case, DI is an anomaly that should be familiar to all practising dentists due to the clinical implications of potential sequelae of pulpal involvement. PMID- 15293821 TI - Histoplasmosis in Australia: a report of a case with a review of the literature. AB - Histoplasmosis is a rare but serious fungal infection commonly presenting as mucosal ulceration of the oral cavity. It is increasingly recognized in Australia but the source of infection remains obscure and it is likely to be under diagnosed. We report a case of chronic mucosal ulceration which failed to fully respond to periodontal therapy. Histology and culture of a gingival biopsy was consistent with histoplasmosis, and the patient responded favourably to treatment with oral itraconazole. Histoplasmosis may present to general dental practitioners as chronic mucosal ulceration and should be considered in the differential diagnosis of such lesions. Diagnosis is best made by culture and histology of biopsy specimens. PMID- 15293822 TI - Changes in South Australian children's caries experience: is caries re-surfacing? PMID- 15293823 TI - Status epilepticus--a review article. AB - BACKGROUND: Status epilepticus is a neurological emergency associated with significant morbidity and mortality and outcome that are inversely related to duration of the condition. Any type of epileptic seizure can develop into status epilepticus (SE) but some types evolve into SE more commonly than others. METHODS: This article reviews both convulsive and non-convulsive types of SE with emphasis on generalised convulsive type the most commonly seen. The aetiology, pathophysiology, complications and drugs used in the management of SE are also discussed. RESULTS: To prevent complications resulting from the repeated seizures, prompt and aggressive management is essential and should proceed along four fronts: (a) Treatment of the SE (b) Prevention of recurrence (c) Management of potential precipitating causes and (d) Management of complication and underlying conditions. The morbidity and mortality associated with SE are often due to: (1) Injury from repetitive electrical discharge (2) Systemic stress from repeated generalised tonic-clonic seizure and (3) The damage to the central nervous system (CNS) caused by the acute insult precipitating the SE. CONCLUSION: The sequelae resulting from SE can be life threatening. Therefore prompt treatment is very crucial with the attending physician having a low threshold for intensive care management. PMID- 15293824 TI - Neurological manifestations of uraemia and chronic dialysis. AB - BACKGROUND: The management of uraemic neurological manifestations is a major target of the treatment of the uraemic syndrome. Chronic dialysis is associated with novel neurological manifestations. OBJECTIVE: To describe the clinical characteristics, pathogenesis and management of the main neurological syndromes encountered in uraemia and chronic dialysis. METHODS: Review of the pertinent literature. Selected references, which have been critical in the understanding of the topic, were included in this review. RESULTS: The main neurological manifestations of uraemia include encephalopathy, neuropathy that can affect cranial, peripheral and autonomic nerves, and proximal myopathy. Retention of uraemic toxins is the main putative cause of uraemic encephalopathy and neuropathy. Arrest or prevention of uraemic encephalopathy and neuropathy are main targets of the dialytic treatment and constitute major criteria of its adequacy. The main cause of uraemic myopathy is secondary hyperparathyroidism and parathyroidectomy is its main treatment. Chronic dialysis is associated with three main neurological syndromes, the disequilibrium syndrome, seen usually in the first few haemodialysis sessions and prevented by starting dialysis with a low dose and progressively increasing the dialysis dose in subsequent dialysis sessions, dialysis dementia, which results from aluminium overloading and is prevented by reducing exposure of the dialysis patients to aluminium, and nerve entrapment, particularly carpal tunnel syndrome, which is caused by beta2 microglobulin amyloidosis and may be prevented by the use of high-flux dialysers which provide relatively high clearance for beta2-microglobulin or by daily haemodialysis. CONCLUSIONS: Specific neurological manifestations are part of the uraemic syndrome and may complicate chronic dialysis. The diagnosis of these manifestations, their differentiation from other neurological syndromes that can complicate the course of renal failure or dialysis, and their specific treatment require clinical acumen and represent a major challenge for physicians treating patients with chronic renal failure or undergoing chronic dialysis. PMID- 15293825 TI - Outcome of term breech by intended mode of delivery. AB - BACKGROUND: The optimal mode of delivery for the term breech fetus is undetermined. METHODS: Over a 10-year period, the outcome of 157 planned caesarean sections and 265 trials of vaginal delivery for the matured, selected, singleton breech presentation were compared. RESULTS: The perinatal mortality among the planned vaginal group was not statistically different from the planned caesarean group (3.0% vs 0.6%, p = 0.102), so was the incidence of birth asphyxia (9.4% vs 4.5%, p = 0.06), admission into neonatal intensive care unit (13.6 vs 7.6%, p = 0.063) and neonatal birth injury (3.4% vs 0.6%, p = 0.072). The overall maternal morbidity was 28% in the planned caesarean group, and 9% in the planned vaginal group, p = 0.025. CONCLUSION: Given appropriate selection criteria and management protocol, the outcome from elective caesarean section might not be better than that from planned vaginal delivery. PMID- 15293826 TI - Cancers of the uterine cervix in Port Harcourt, Rivers State--a 13-year clinico pathological review. AB - BACKGROUND: Carcinoma of the uterine cervix is still the commonest gynaecological malignancy among women in the developing world. This study aims at assessing the prevalence of carcinoma of the cervix, the most common presenting complaints and histological types as seen in Port Harcourt, Rivers State. METHODS: All the tissue slides of specimen (cervix and hysterectomy) diagnosed as cervical carcinoma over a period of 13 years were retrieved and re-evaluated for confirmation of diagnosis and tumour typing in the Anatomical Pathology Department of the University of Port Harcourt Teaching Hospital, Port Harcourt, Rivers State. Clinical data were obtained from histopathology registers, request forms as well as case notes of patients from the records department. RESULTS: There were 2,236 malignancies diagnosed; 302 (13.5%) were malignancies of the genital tract while 188 (8.4%) were carcinoma of the cervix constituting 62.3% of female genital malignancies. Cervical cancer was commonest between the ages of 50 69 years. There was no patient with carcinoma of the cervix below the age of 20 years. Abnormal vaginal bleeding (84.1%) in pre and post-menopausal periods was the commonest presenting complaint while squamous cell carcinoma (70.2%) and adenocarcinoma (14.9%) of the uterine cervix were the main histological types. CONCLUSION: Post menopausal women are mostly affected by cancers of the uterine cervix while abnormal vaginal bleeding is the most common presenting complaint. Squamous cell and adenocarcinoma are the most frequent histological types. PMID- 15293827 TI - Prevalence of HIV antibodies amongst apparently healthy pregnant women in Mongomo, Guinea Equatoria. AB - BACKGROUND: HIV infection among pregnant women is common and continues to grow. This study was undertaken to evaluate the seroprevalence of HIV antibody in apparently healthy pregnant women. METHOD: Sera of three hundred and twenty-four consecutive pregnant women who had booked for antenatal care between February and July 1997 at the 'Provincial Hospital de Mongomo', Mongomo, Guinea Equatoria were screened for the presence of HIV antibody. Only four (1.23%) pregnant women tested positive using the Enzyme Linked Immunosorbent Assay (ELISA) technique (Wellcome HIV recombinant EIA Kit, Wellcome Diagnostics, UK). This was later confirmed using the Western Blot Technique (Dupont, USA). RESULTS: The study shows that the HIV seroprevalence rate amongst pregnant women in Mongomo, Guinea Equatoria is low compared with the neighbouring countries in the Central African Region. This result, however, is higher than the National rate; this is not surprising since Mongomo has been shown to have a seroprevalence rate higher than the national average in the general population. CONCLUSIONS: Routine HIV screening for all antenatal care patients, provision of anti-retroviral drugs to the sero-positive pregnant women at the subsidised rates, improved socio-economic status especially of women and certain cultural behavioural changes which empower the women are suggested as some of the ways to curb the menace of HIV infection amongst pregnant women and by extension reduce the vertical transmission of HIV to the baby. PMID- 15293828 TI - Missing IUD strings: diagnosis and management at Ilorin. AB - BACKGROUND: Intrauterine contraceptive device (IUD) is a widely accepted method of contraception. One of the complications reported with IUD use is missing IUD. METHODS: This retrospective study was carried out on a total of forty-two women who had missing IUDs over a three-year period (January 1992-December 1994). Socio demographic data, type of previous contraception, timing of insertion, service provider of IUD, diagnostic and management methods of these cases were reviewed. Epiinfo statistical software package was used for analysis and the level of significance was at p<0.001. RESULTS: The total missing IUDs for the years under review accounted for 0.25% of total IUD continuous users or 0.89% of total new IUD acceptors. There was a slight decline over the study period in the percentage of missing IUDs. Majority (85.71%) presented more than 6 months postpartum for insertion while insertions during menses was common (71.43%). Expulsions of IUDs was recorded in 21 (50%) clients. Pelvic examination with uterine sound and abdominopelvic ultrasound were the diagnostic methods commonly used. Trainees at the family planning unit inserted most of the missing IUDs while most clients (71.43%) missed their IUDs especially within the first three months of use. Retrieval loop alone (64.29%) and with dilatation (11.91%) and uterine sounding alone (16.67%) were commonly employed in the management of missing IUDs. About a third had no associated complications. CONCLUSIONS: Prevalence of missing IUDs is low in this centre occurring mostly within 3 months of insertion. Good selection of women using IUDs will result in less reported cases of missing IUDs. Counselling will motivate the IUD user to present early when any complication arise. The trainees need closer supervision and should be taught appropriate insertion techniques. PMID- 15293829 TI - Genital prolapse in Port Harcourt, Nigeria. AB - BACKGROUND: Damage to the genitourinary supports from repeated pregnancies and labour is the most important predisposing aetiological factor in genital prolapse, a chronic gynaecological disorder that causes distressing morbidity (stress urinary incontinence, micturitional difficulties and problems of defecation). OBJECTIVE: To determine the pattern of presentation and management of genital prolapse at the University of Port Harcourt Teaching Hospital, and compare with experience elsewhere. METHODS: Retrospective review of all cases of genital prolapse admitted and operated upon during the period January 1990 to December 1999. Data collected and analyzed by simple percentages included sociodemographic characteristics, aspects of clinical presentation, management modality and outcome. RESULTS: Genital prolapse (118 cases) accounted for 37.5 per 1,000 gynaecological admissions during the study period. Seventy percent of the study subjects were above age 45 years, while seventy-three percent were grandmultiparous. Thirty percent were of reproductive age. The commonest symptom was "something coming down the vagina" (95% of the study subjects). Only 4% had stress incontinence. Second degree prolapse was the commonest finding on presentation (68.6%) while vaginal hysterectomy with pelvic floor repair was the main operative procedure (undertaken in 73.7% of the subjects). Postoperative deaths occurred in two patients, both diabetics (case fatality ratio 1:59) from septicaemia and intestinal obstruction respectively following vaginal hysterectomy. Postoperative morbidity following vaginal hysterectomy was also higher than that following Manchester Repair. CONCLUSION: This study's results are similar to previous findings in Nigeria and other developing countries. Measures to reduce grandmultiparity, difficult deliveries and postoperative morbidity and mortality are advocated. PMID- 15293830 TI - Current aetiology of neonatal sepsis in Jos University Teaching Hospital. AB - BACKGROUND: Periodic bacteriologic surveillance in neonatal units is a necessity. OBJECTIVE: To determine the currently prevalent pathogens of neonatal sepsis in the Special Care Baby Unit of Jos University Teaching Hospital, and their antibiotic susceptibility profiles. METHODOLOGY: One hundred and twenty two neonates with clinical suspicion of sepsis underwent bacteriologic screening over a 4 month period. RESULTS: One hundred and fourteen bacteria were isolated from 66 (54.1%) of the infants. Forty two (36.8%) isolates were gram positive (predominantly Staphylococcus aureus), while 72 (63.2%) were gram negative (predominantly Escherichia coli). Gentamicin was most useful antibiotic, though effective against only 67% of both gram positive and gram negative bacteria. The susceptibilities of both gram positive and gram negative bacteria to the third generation cephalosporins were particularly poor (less than 10%). CONCLUSION: Our data show a change in the predominant gram negative bacterial pathogen compared with an earlier report from our unit, and an alarming overall decline in the susceptibilities of both gram positive and gram negative pathogens to the commonly used antibiotics. PMID- 15293831 TI - Obstructive uropathy in childhood, as seen in University of Port Harcourt Teaching Hospital, Nigeria. AB - BACKGROUND: Obstructive uropathy is a cause of morbidity and mortality in children. In Port Harcourt, data on causes of obstructive uropathies are not readily available. This study was carried out to identify the causes and outcome of obstructive uropathies seen in our children's ward. METHODS: The case records of children with the diagnosis of obstructive uropathies who were treated at the University of Port Harcourt Teaching Hospital (UPTH) between October 1997 and October 2002 were reviewed. RESULTS: A total of 20 patients, all males were seen. The ages ranged from 4 weeks to 13 years with a mean of 2.3+/-2.8 years. The causes of obstructive uropathy were posterior urethral valves (PUV) 16 (80%), bladder calculi 2 (10%), bladder rhabdomyosarcoma and urethral stenosis 1 (5%) each. Poor stream of urine and dysuria were the commonest presentation. The duration of symptoms ranged from 2 days to 13 years. None of the patients with PUV was diagnosed prenatally. Hypertension and urinary tract infection each were found in 50% of the patients while 6 (30%) presented with features of renal failure. Seven patients died, giving a mortality rate of 35%. Age at presentation less than one year and duration of symptoms longer than one month was associated with higher mortality although it was not statistically significant. CONCLUSION: PUV is the commonest cause of obstructive uropathy seen in UPTH. Earlier diagnosis during pre-natal period or when this is not possible, diagnosis within the first week of life should be encouraged. Parents, nurses and attending doctors should ensure they observe the urinary stream of every male child before discharge from the hospital for early detection and management of PUV. PMID- 15293832 TI - Paediatric accidental deaths in Port Harcourt, Nigeria: a 10-year retrospective study. AB - BACKGROUND: Some studies on medicolegal autopsies have been conducted in Nigeria. This study was carried out to highlight the causes, peculiarities and possible factors responsible for paediatric accidental deaths in Port Harcourt, Nigeria. METHODS: All 3058 medicolegal autopsies referred to the coroner and performed in the department of Anatomical Pathology University of Port Harcourt Teaching Hospital, over a 10 year period covering January 1986 to December 1995 were analyzed. The accidental death subset was specially analyzed with respect to the paediatric age group of 0-16 years. RESULTS: A total of 3084 coroners autopsies were carried out during the study period. Of these 356 (11.5%) were paediatric accidental deaths. The commonest cause of death was road traffic accidents constituting 84.3% (n=300). The male:female ratio was about the same. The head was the commonest body region affected (n=90). Cars and buses were the commonest vehicles involved. Pedestrians were most at risk constituting 66 percent. Bulk of the cases occurred over the weekends. Drowning, electrocution and burns accounted for 8.7%, 2.8% and 4.2% respectively. CONCLUSION: Most of the deaths are avoidable. Parental monitoring, control of movements, legislation and general concern of the adult population for children's welfare will reduce these largely preventable deaths. PMID- 15293833 TI - Epidemiology of childhood burns in Maiduguri north-eastern Nigeria. AB - BACKGROUND: Burns is a global problem and has its toll especially in a developing region like ours where poverty and ignorance are still rife. Previous studies in the sub-region have lumped children and adults together. We retrospectively studied the factors that lead to burns in children and the peculiarities in managing them. METHODS: All case notes of burns injury in children managed at the University of Maiduguri Teaching Hospital between 1991-2000 were retrospectively studied. RESULTS: A total of 219 children were studied. Children of age below 5 years were affected more than children above 5 years (168 vs. 51) with toddlers 1 2 years constituting a significant proportion of those below 5 years (71 vs. 168). The male to female ratio was 1.6:1 with a preponderance of male children below 10 years and the females between 11-15 years of age. The commonest cause of burns was scald (64.4%) in the household, which is usually accidental, but 3 were suicide attempts by teenage pregnant females 11-15 years protesting forced marriages, a cultural problem in our environment. Flame burns ranked second (27.4%) and results mainly from careless storage, adulteration and hawking of petroleum products. More than 50% of the patients sustained major burns resulting in high morbidity and mortality rate of 16%. CONCLUSION: Burn is a major public health problem and will require public/school health education campaign on childhood household safety. Appropriate legislation and enforcement on the sale of petroleum products would help to reduce the scourge. PMID- 15293834 TI - Informed consent: how informed are patients? AB - BACKGROUND: By casual interaction with patients our impression was that patients lacked adequate knowledge about their ailments and surgical treatment. There is now increasing educational exposure and legal awareness in the society. AIM: To reassess the extent of the practice of "informed consent" expose failures if any, and make appropriate recommendation. METHODS: A written questionnaire, randomly distributed to patients, sought to find out to what extent patients admitted into the University of Port Harcourt Teaching Hospital (UPTH) were adequately informed about their ailments and the implications of surgical treatment. RESULTS: One hundred and fifty patients were randomly recruited into the study and all responded to the questionnaire. Majority of the patients (75%) knew their diagnoses. Most patients (63%) did not know the problems that could be associated with their surgery while 75% did not know the complications of anaesthesia. CONCLUSION: There is need for a closer interaction in communication between patients and their surgeons. This will facilitate the participation of patients in decisions on treatment options and consequently help to maintain a satisfactory standard of practice and reduce litigation following an unfavourable outcome of surgical treatment. PMID- 15293835 TI - Hoarseness in adult Nigerians: a University College Hospital Ibadan experience. AB - BACKGROUND: Hoarseness is a major symptom of laryngeal disease. Persistent hoarseness may be an early warning of such sinister lesions as cancer of the larynx and nasopharynx, hence the need for increased awareness of this symptom and its causes. METHODS: This is a retrospective study of all hoarse adult patients seen in the Otorhinolaryngology clinics of the University College Hospital Ibadan over an 8-year period (1995-2002). RESULTS: The total study population of 124 patients consisted of 72 (58.06%) males and 52 (41.94%) females with an overall mean age of 46.98 years and age range 16-84 years. The mean duration of hoarseness before presentation was 23.29 months; 56 (45.2%) patients presented within 6 months of onset of hoarseness. Chronic non-specific laryngitis including vocal cord nodules was the most common cause of hoarseness (55.6%). Chronic non-specific laryngitis patients had a mean age of 45.05 years. Fourteen point forty-nine percent (14.49%) of patients with chronic non-specific laryngitis smoked cigarette and drank alcohol, 60.88% were professional voice users. The other causes of hoarseness included laryngeal cancer (24.2%), recurrent laryngeal nerve palsy (8.1%) and laryngeal papilloma (6.5%). The patients with laryngeal cancer had a mean age of 57.63 years. The two male patients with laryngeal tuberculosis were all secondary to pulmonary phthisis. CONCLUSION: The causes of hoarseness are varied and late presentation may worsen the prognosis. Persistent hoarseness of more than three weeks should have detailed Otolaryngological evaluation. PMID- 15293836 TI - Cataract blindness and barriers to cataract surgical intervention in three rural communities of Oyo State, Nigeria. AB - BACKGROUND: The study was a community based field survey that used a rapid assessment method to determine the prevalence of cataract blindness in people aged 50 years and above in 3 rural communities. OBJECTIVE: The purpose of the study was to determine the prevalence of cataract blindness and barriers to cataract surgical intervention in an area served by a health facility managed by a tertiary institution. METHOD: Abedo, Akinyele and Ketepe Villages in Akinyele Local Government Area of Oyo State Nigeria were selected based on their nearness to the primary health care centre in Abedo. RESULT: A total of 477 persons aged 50 years and above were seen being 73.3 percent of expected. Those with visual acuity of less than 3/60 in an eye and those with visual acuity of less than 3/60 in the better eye were examined in more details using a pen torch, an ophthalmoscope and tonometer to determine the cause of blindness. All persons who have had surgery were examined. The prevalence of blindness in persons aged 50 years and above was 1.47% and that of cataract blindness in the same age group in the villages was 0.84% constituting 57.14% of blindness. The main barriers to hospital presentation were cost of surgery (52.8%) and distance to hospital (33.8%). The constraints and limitations encountered during the study included rural-urban migration and population discrepancies between what obtained at the villages and those supplied by the Local Government Population Commission. CONCLUSION: It is hoped that this study will serve as a preliminary survey and a base line for further studies and the initiation of a blindness prevention programme in the area. PMID- 15293837 TI - Traumatic spinal cord injuries (SCI): a study of 104 cases. AB - BACKGROUND: Management of spinal cord injured patients is an integral part of trauma care. There is need to congregate these patients in spinal units where dedicated experts and facilities exist for better outcome of treatment and rehabilitation. The objective of this study therefore, is to promote improved quality of care in the group of patients by highlighting the deficiencies in our setting. METHODS: This is a retrospective study of all traumatic spinal cord injured (SCI) seen at the National Orthopaedic Hospital, Enugu over a six-year period. Information about the demographics, aetiology, level of injury, associated injuries, time of presentation, referral hospital, state of injury, duration of hospitalization, outcome of treatment were obtained from patients' records. Data were analyzed using commercially available SPSS for windows 9. Descriptive statistics are given. RESULTS: There were 104 patients, 88 males and 16 females, giving a ratio of 5.5:1. Road traffic accident, 60 (57.7%), was the commonest aetiological factor and the cervical spine, 70 (67.3%), was most often involved. Head and neck injuries were common associated injuries, 19 (18.3%) in patients with cervical spine injury. Six (5.8%) of our patients were obtunded. Most patients, 69 (66.4%), were received from private hospitals after a mean duration of 7 days. Hospitalization ranged from 1 week-120 weeks with a mean of 11 weeks. Pressure sore was the commonest complication, 30 (28.8%). Thirty six (34.4%) of patients died mainly from respiratory failure. Most of the discharged wheel bound patients (97%) were lost to follow up. CONCLUSION: For better outcome we re-emphasize the need to establish and congregate these patients into regional spinal units. We also suggest a social legislation, which will be fundamental to social re-integration of these SCI and other severely disabled patients. PMID- 15293838 TI - Attitudes to, knowledge and practice of breast self-examination (BSE) in Port Harcourt. AB - BACKGROUND: Many of the cases of carcinoma of the breast we find in our clinic are advanced, sometimes with fungation. We believe that if women in Port Harcourt are knowledgeable about breast self-examination (BSE) and practise it, this scenario may not be so. AIM: The study is intended to highlight the extent of their knowledge vis-a-vis their practice of BSE. METHOD: A written questionnaire was distributed to 200 women from different walks of life in Port Harcourt to assess their attitudes to, knowledge and practice of BSE. Their responses were then collated and analyzed. RESULTS: Ninety-eight percent of the respondents had formal education, majority having obtained tertiary education. Eighty-five point five percent of them had heard of BSE but 39.0% practised BSE only occasionally, while 24.0% did not practise it at all. Among 76 health workers who participated in the study, 60.0% of doctors and 53.7% of nurses practised BSE only occasionally. Only one doctor could describe how to perform BSE correctly. The news media, nurses and physicians were the commonest sources of information on BSE. CONCLUSION: Most women in Port Harcourt, though aware of BSE and its usefulness never practise it. Those who care to practise it are ignorant of how to correctly do it. There is need for a vigorous health education programme on this subject for our women. It is hoped that this will help to reduce the morbidity and mortality associated with carcinoma of the breast. PMID- 15293839 TI - Aortic arch diameter in frontal chest radiographs of a normal Nigerian population. AB - BACKGROUND: The objective of our study was to establish age, sex, weight and height specific nomogram of radiographic aortic arch diameter in a normal adult Nigerian population. METHODS: The aortic arch diameter of 303 normotensive males and females aged between 20 and 93 years were measured in normal posteroanterior chest radiographs. RESULTS: The mean values for the 20-39, 40-59 and the > or = 60 year-old males were 5.3, 5.5 and 6.3 cm respectively while the corresponding values for females were 4.9, 5.3 and 5.9 cm. The aortic arch diameter increased progressively with age in both sexes and was larger in males than in females. It correlated better with body mass index than with weight, height or body surface area. The proportion of males and females below the age of sixty that had aortic arch diameter of > or = 6.0 cm ranged from 0.9 to 9.7%. For the > or = 60 year old, the proportion was 30% for females and 70% for males. CONCLUSION: The study showed that whereas radiographic aortic arch diameter seemed to be a good index of systemic hypertension for the under sixty it is not so for the > or = 60 year old. PMID- 15293840 TI - The prevalence of hepatitis B surface antigenaemia in HIV positive patients in the Niger Delta Nigeria. AB - BACKGROUND: Both hepatitis-B virus (HBV) and human immunodeficiency virus (HIV) infections are common in Nigeria and are a significant cause of mortality and morbidity. This study was undertaken to determine the prevalence of hepatitis B surface antigen (HBsAg) in HIV infected patients and to highlight the need to pay attention to the recognition of potentially severe concurrent illness that may increase morbidity and mortality of HIV infected patients. METHODS: Three hundred and forty-two HIV positive patients recruited into the antiretroviral therapy pilot project at the University of Port Harcourt Teaching Hospital comprising 192 males and 150 females between June 1995 to February 2003 constituted subjects for this study. HIV status of subjects was confirmed using the WHO approved Immunocomb (Organics, Israel) and Recombigen HIV I and II kits (Cambridge diagnostics, Ireland). HBsAg was assayed using the commercially available Clinotech HBsAg kits based on the chromatographic immunoassay technique (Clinotech diagnostics, Canada). RESULTS: HBsAg was detected in 33 (9.7%) of patients infected with HIV. Co-infection rate was highest in the 33-39 years age group. Single/unmarried patients constituted the highest proportion of those with HIV/HBV co-infection 21/195 (10.8%) followed by widowed/separated 4/47 (8.5%) and married (8.0%). Commercial sex workers had the highest prevalence among the occupational groups 4/30 (13.3%) followed by applicants 8/75 (10.7%) and drivers 4/40 (10.0%) while the lowest occupational prevalence occurred among farmers. Chi square analysis indicated that age was an independent risk factor for HBV co infection in HIV infected patients (P<0.05). CONCLUSIONS: This study demonstrates a high prevalence (9.7%) of HBsAg in HIV infected patients. This calls for a more intensive prevention and surveillance measures to control the HIV/AIDS scourge and co-infection with HBV. PMID- 15293841 TI - Rising trends in caesarean section rates: an issue of major concern in Nigeria. AB - BACKGROUND: Caesarean section rates have been on the increase for the past decades. Reasons for this increase and the danger it portends, especially in developing countries, are evaluated. METHOD: Literature on caesarean sections performed in various centres in Nigeria and Western countries were retrieved through manual library search. RESULTS: There has been a progressive rise in caesarean section rates from the 1970s up to 2002, from 9.4% to 34.6%. Measures to reduce this rising rates were discussed. CONCLUSION: There is great need to reduce the caesarean section rate in Nigeria. There should be a clear and unquestionable indication for caesarean section in any patient. PMID- 15293842 TI - Health and safety in clinical laboratories in developing countries: safety considerations. AB - BACKGROUND: Clinical laboratories are potentially hazardous work areas. Health and safety in clinical laboratories is becoming an increasingly important subject as a result of the emergence of highly infectious diseases such as hepatitis and HIV. This is even more so in developing countries where health and safety have traditionally been regarded as low priority issues, considering the more important health problems confronting the health authorities in these countries. METHODS: We conducted a literature search using the medical subheadings titles on the INTERNET over a period of twenty years and summarized our findings. RESULTS: This article identifies hazards in the laboratories and highlights measures to make the laboratory a safer work place. It also emphasizes the mandatory obligations of employers and employees towards the attainment of acceptable safety standards in clinical laboratories in Third World countries in the face of the current HIV/AIDS epidemic in many of these developing countries especially in the sub-Saharan Africa while accommodating the increasing work load in these laboratories. CONCLUSION: Both the employer and the employee have major roles to play in the maintenance of a safe working environment. This can be achieved if measures discussed are incorporated into everyday laboratory practice. PMID- 15293843 TI - Sigmund Freud: smoking habit, oral cancer and euthanasia. AB - BACKGROUND: Sigmund Freud, the father of modern psychoanalysis had a well-known love of the cigar. The natural progression of this vice was the development of oral cancer for which he underwent a lengthy ordeal. An account is given in this article of Sigmund Freud's illness and care following the diagnosis of his oral cancer. The role of euthanasia and physician assisted suicide is also discussed. METHODS: A review of relevant literature on Sigmund Freud's illness, risk factors for oral cancer and euthanasia was undertaken. RESULTS: Sigmund Freud was a heavy smoker with a 20-cigar/day habit. In 1923, a diagnosis of squamous cell carcinoma of the palate was made, for which he underwent a lengthy ordeal which span a total of 16 years. During this period, he bluntly refused to quit smoking. Freud consulted many specialists (otolaryngologists, oral and maxillofacial surgeons, prosthodontists and general surgeons), during the course of his ordeal with oral cancer. He underwent 34 surgical procedures before his eventual death in 1939 through euthanasia. CONCLUSION: Continued indulgence in smoking and procrastination on the part of Freud, as well as mediocrity, negligence and incompetence on the part of the first surgeon that operated on Freud, could partly be responsible for his lengthy ordeal. PMID- 15293844 TI - Ruptured advanced tubal ectopic pregnancy simulating uterine rupture: a case report. AB - BACKGROUND: Ectopic pregnancy is one of the most critical and life-threatening emergencies in gynaecological practice and poses a diagnostic dilemma in advanced cases. This report highlights a case of ruptured advanced tubal ectopic pregnancy simulating uterine rupture. METHOD: Case-note of a patient managed for ruptured advanced tubal ectopic pregnancy was used with a review of the relevant literature. RESULT: A 24-year old primigravida who presented at 23 weeks gestation with signs and symptoms suggestive of ectopic pregnancy is presented. The advanced nature of the pregnancy posed a diagnostic dilemma as ruptured uterus shares the same characteristic dramatic presentation, especially in this patient with previous myomectomy. Prompt resuscitation and immediate laparotomy produced a good outcome. CONCLUSION: High index of suspicion is important in the diagnosis of ectopic pregnancy. Even when diagnosis is in doubt, exploratory laparotomy may be life saving. PMID- 15293845 TI - Severe diarrhoea complicating metformin therapy: an unusual presentation. AB - BACKGROUND: Gastro-intestinal symptoms especially diarrhoea could complicate metformin therapy. METHODS: A 57-year old Nigerian man who had severe diarrhoea as a result of metformin therapy is presented. CONCLUSION: Clinicians should be wary of this complication and be on the watch out for it. PMID- 15293847 TI - The best of times, the worst of times: the wealth and poverty of heart failure pharmacotherapies: is there a role for vesnarinone? PMID- 15293848 TI - Clinical characteristics of vesnarinone. AB - Congestive heart failure is a common condition with a poor prognosis. Its high rates of morbidity and mortality produce a huge societal burden. Current pharmacological treatment approaches are based on angiotensin-converting enzyme inhibitors, diuretics and digoxin, but up to 5% of patients may have refractory disease with persistent symptoms at rest. Such patients with advanced-stage disease may be candidates for treatment with the novel agent vesnarinone, a mixed phosphodiesterase inhibitor and ion-channel modifier that has modest, dose dependent, positive inotropic activity, but minimal negative chronotropic activity. Vesnarinone improves ventricular performance most in patients with the worst degree of heart failure. However, before the initiation of vesnarinone therapy, risk-benefit profiles in individual patients should be considered, because in two large-scale studies [i.e. of the high dosage used in the Vesnarinone Study Group Trial (VSGT), and of both dosages used in the Vesnarinone Trial (VEST)] a dose-dependent increase in mortality was identified for vesnarinone 30-120 mg/day. The two studies also found significant vesnarinone induced, short-term improvements in quality of life (QOL) in patients with refractory end-stage heart failure. Such patients are the most willing to trade off a slightly increased risk of mortality for improved QOL. It is thus in these patients with refractory end-stage heart failure that vesnarinone may ultimately establish an important treatment role. However, detailed further investigation of the overall place of vesnarinone in heart failure management, with particular reference to the clinical potential of vesnarinone plus beta-blocker combination therapy, for example, is certainly warranted. PMID- 15293849 TI - Precautions for use and adverse effects of vesnarinone: potential mechanisms and future therapies. AB - This article reviews the precautions and adverse effects associated with vesnarinone use, and the potential mechanisms responsible for these complications as well as suggested treatment strategies. Vesnarinone, a quinolinone derivative, improves the haemodynamics and quality of life in patients with congestive heart failure (CHF); however, it is associated with the adverse effects of increased sudden cardiac death and neutropenia. These adverse effects have limited the application of vesnarinone to the general population but perhaps with continued research into vesnarinone-induced neutropenia and advances in arrhythmia management, the risk/ benefit ratio of vesnarinone may become favourable. For now, the use of vesnarinone should be limited to patients with CHF who have demonstrated a poor response to other cardiac medications and devices. These patients should be closely monitored for both cardiac and non-cardiac adverse effects. PMID- 15293850 TI - Optimising outcomes in end-stage heart failure: differences in therapeutic responses between diverse ethnic groups. AB - Clinical and pathophysiological differences between Japanese and Caucasian patients are observed in many aspects of heart disease. Indeed, data derived from studies in one population cannot be automatically extrapolated to the other. The therapeutic goal of heart failure has recently been aimed at improving mortality in Western societies. The long-term use of an inotropic agent in the energy starved failing heart has been expected to increase myocardial energy use and accelerate the disease process. However, this may not be the case in the Japanese population in whom mortality is relatively low. Therefore, vesnarinone therapy could be justified, since it allows optimal care in terms of an improved quality of life. Nevertheless, re-analysis of the findings of the Vesnarinone Trial (VEST) emphasised again the reasons for the precautions relating to vesnarinone use: (i) vesnarinone was associated with increased death, usually occurring within 7 months of initiation of the drug; (ii) the mortality rate was higher in patients receiving concomitant digoxin, which necessitated close monitoring of renal function; (iii) the mortality rate also increased in patients with severe bradycardia, indicating the importance of regular ECG monitoring; and (iv) improvements in cardiac function and symptoms by the drug may result in sudden death, particularly in patients with severe heart failure. Such patients should be closely monitored by a physician. PMID- 15293851 TI - Sex causes altruism. Altruism causes sex. Maybe. AB - This study presents a mathematical model in which the fitness of an individual depends on the individual's genotype (individual effects) and on the genotypes of other members of the individual's local group (group effects). The findings suggest that, if phenotypes are a result of complex interactions between genes at different loci, then fitness-enhancing group effects may become common in sexual populations. The spread of fitness-enhancing group effects is facilitated when environmental conditions sometimes deteriorate temporarily. This is so even if the genotypes with the highest group effects also tend to have relatively low individual effects. In this sense, the process described here can lead to the evolution of altruism. By contrast, when populations are asexual it appears that group effects are much less important in determining the outcome of evolution. Thus, in nature, asexual populations may tend to be characterized by more antagonistic interactions than those that typically prevail when reproduction is sexual. This might help to explain why asexual lineages are prone to rapid extinction. PMID- 15293852 TI - Occasional sex in an 'asexual' polyploid hermaphrodite. AB - Asexual populations are usually considered evolutionary dead-ends because they lack the mechanisms to generate and maintain sufficient genetic diversity. Yet, some asexual forms are remarkably widespread and genetically diverse. This raises the question whether asexual systems are always truly clonal or whether they have cryptic forms of sexuality that enhance their viability. In the planarian flatworm Schmidtea polychroa parthenogens are functional hermaphrodites (as are their sexual conspecifics), copulate and exchange sperm. Sperm is required for initiation of embryogenesis but usually does not contribute genetically to the offspring (sperm-dependent parthenogenesis). Using karyology and genotyping of parents and offspring, we show that in a purely parthenogenetic population an estimated 12% of all offspring are the result of partial genetic exchange. Several processes of chromosome addition and loss are involved. Some of these result in an alternation between a common triploid and a rare tetraploid state. We conclude that genetic recombination does not necessarily require segregation and fusion within the same generation, as is the case in most sexual species. These occasional sexual processes help to explain the geographical dominance of parthenogens in our study species. PMID- 15293853 TI - Multiple mating and reproductive skew in Trinidadian guppies. AB - Male offspring production in promiscuously mating species is typically more skewed than female offspring production. It is therefore advantageous for males to seek as many mating partners as possible. However, given the documented benefits of polyandry we expect females, as well as males, to mate multiply. We tested these ideas using Trinidadian guppies, Poecilia reticulata. Fishes were collected from the wild, housed in groups of 10 males and 10 females and allowed to reproduce freely over a period of three months. We used hypervariable microsatellite loci to identify the parents of 840 offspring and to quantify the variance in mating success. As anticipated, and in line with the Bateman gradient, there was greater skew in the number of progeny produced by males. By contrast, we found no sex difference in mating partner number over the duration of the experiment. A median of two males fathered each brood and there was marked turnover in the identities of the sires of successive broods. Female partner turnover was, however, less than expected under random mating. We suggest that partner switching over time, as well as polyandry within broods, could contribute to the maintenance of genetic diversity in guppy populations. PMID- 15293854 TI - Breathing with a mouth full of eggs: respiratory consequences of mouthbrooding in cardinalfish. AB - Mouthbrooding occurs among several groups of fishes. Although a mouth full of eggs can be expected to pose a considerable respiratory problem, to our knowledge no study has examined respiratory consequences of mouthbrooding in fishes, or how hypoxia or strenuous swimming may affect the success of this reproductive strategy. In two species of cardinalfish (Apogon fragilis and Apogon leptacanthus), from the reef at Lizard Island (Great Barrier Reef), we found that mouthbrooding significantly reduced the ability to take up oxygen at low ambient oxygen levels. While the direct energetic cost of mouthbrooding appeared insignificant at rest in well-oxygenated water, mouthbrooding significantly reduced the respiratory scope of the fishes and their capacity for sustained aerobic swimming. The males spat out their eggs in hypoxia. Interestingly, the species with the larger brood, A. fragilis, spat out the brood at a higher water [O2] than did A. leptacanthus, which had a smaller mean brood mass. Moreover, in contrast to mouthbrooding A. leptacanthus, mouthbrooding A. fragilis was unable to increase its ventilatory frequency in response to hypoxia. This suggests a trade-off situation between hypoxia tolerance and brood size. Apparently, A. fragilis has sacrificed hypoxia tolerance in favour of a large brood size to a greater extent than has A. leptacanthus. PMID- 15293855 TI - Testosterone, cuckoldry risk and extra-pair opportunities in the Seychelles warbler. AB - In male birds, testosterone (T) plays an important role in aggressive and mate attraction behaviour. In the cooperatively breeding Seychelles warbler, Acrocephalus sechellensis, extra-group copulations (EGCs) occur frequently, but are not accompanied by sexual courtship displays as in within-pair copulations. Paternity is nearly always gained by primary males. We investigated whether T levels and sperm storage capability (cloacal protuberance (CP)) in adult primary and subordinate males were related to timing of egg laying, levels of cuckoldry and extra-group paternity (EGP) opportunities. During the sexually active period before egg laying, T levels and CP were only elevated or enlarged (respectively) in primary males, and some suggestion was found that subordinate males do not invest in elevated T levels. The peak in T occurred during the fertile period of the female partner and corresponded to the peak period of male sexual displays and mate guarding, but was independent of cuckoldry risk (density of neighbouring primary males). CP was also enhanced during this period; however, CP but not T remained elevated after egg laying by their mates, and CP but not T was positively related to EGP opportunities (density of neighbouring fertile females). We conclude that T is involved in sexual courtship displays and mate guarding, but not in gaining EGCs. These findings contrast with those in other species where EGP involves elaborate sexual displays. PMID- 15293856 TI - Male red-winged blackbirds distrust unreliable and sexually attractive neighbours. AB - In many species, territorial neighbours fight to establish their mutual border and then develop a truce, known as the dear-enemy phenomenon, characterized by reduced vigilance and aggression along the border. We present evidence that among male red-winged blackbirds (Agelaius phoeniceus) the dear-enemy relationship is a form of reciprocal conditional cooperation that is stabilized, at least in part, by retaliation against cheaters. Simulated intrusions by randomly chosen neighbours were punished by a targeted increase in vigilance and aggression that persists for days. We interpret this increase in vigilance towards trespassers as a manifestation of distrust. The conditional decrease in vigilance and aggression is tempered by each neighbour's probability of cuckolding the focal male. Male red-winged blackbirds maintained greater vigilance and aggression towards sexually attractive neighbours that were more successful at extra-pair fertilizations (EPFs). It is unlikely that males directly observed neighbours copulating with their mates. They were more likely to assess a neighbour's ability to achieve extra-pair copulations using surrogate cues that correlate with success at EPFs, including body size. Our results suggest that red-winged blackbirds use rules that incorporate their neighbour's behaviour and quality in their territorial interactions with one another. Our results expand our understanding of cooperation for animals and for humans as well. PMID- 15293857 TI - Reduced flocking by birds on islands with relaxed predation. AB - Adaptive hypotheses for the evolution of flocking in birds have usually focused on predation avoidance or foraging enhancement. It still remains unclear to what extent each factor has contributed to the evolution of flocking. If predation avoidance were the sole factor involved, flocking should not be prevalent when predation is relaxed. I examined flocking tendencies along with mean and maximum flock size in species living on islands where predation risk is either absent or negligible and then compared these results with matched counterparts on the mainland. The dataset consisted of 46 pairs of species from 22 different islands across the world. The tendency to flock was retained on islands in most species, but in pairs with dissimilar flocking tendencies, island species were less likely to flock. Mean and maximum flock size were smaller on islands than on the mainland. Potential confounding factors such as population density, nest predation, habitat type, food type and body mass failed to account for the results. The results suggest that predation is a significant factor in the evolution of flocking in birds. Nevertheless, predation and other factors, such as foraging enhancement, probably act together to maintain the trait in most species. PMID- 15293858 TI - Signature-whistle production in undisturbed free-ranging bottlenose dolphins (Tursiops truncatus). AB - Data from behavioural observations and acoustic recordings of free-ranging bottlenose dolphins (Tursiops truncatus) were analysed to determine whether signature whistles are produced by wild undisturbed dolphins, and how whistle production varies with activity and group size. The study animals were part of a resident community of bottlenose dolphins near Sarasota, Florida, USA. This community of dolphins provides a unique opportunity for the study of signature whistle production, since most animals have been recorded during capture-release events since 1975. Three mother-calf pairs and their associates were recorded for a total of 141.25 h between May and August of 1994 and 1995. Whistles of undisturbed dolphins were compared with those recorded from the same individuals during capture-release events. Whistles were conservatively classified into one of four categories: signature, probable signature, upsweep or other. For statistical analyses, signature and probable signature whistles were combined into a 'signature' category; upsweep and other whistles were combined into a 'non signature' category. Both 'signature' and 'non-signature' whistle frequencies significantly increased as group size increased. There were significant differences in whistle frequencies across activity types: both 'signature' and 'non-signature' whistles were most likely to occur during socializing and least likely to occur during travelling. There were no significant interactions between group size and activity type. Signature and probable signature whistles made up ca. 52% of all whistles produced by these free-ranging bottlenose dolphins. PMID- 15293859 TI - Acoustic monitoring on a humpback whale (Megaptera novaeangliae) feeding ground shows continual singing into late Spring. AB - Singing by males is a major feature of the mating system of humpback whales, Megaptera novaeangliae (Borowski). Although a few songs have been opportunistically recorded on the whales' high-latitude feeding grounds, singing in these regions was thought to be only sporadic. We report results from the first continuous acoustic monitoring of a humpback whale feeding ground (off Cape Cod, MA, USA) in spring. Using autonomous sea-floor recording systems, we found singing on a daily basis over the entire 25 day monitoring period, from 14 May to 7 June 2000. For much of the period, song was recorded 24 h per day. These results, combined with evidence for aseasonal conceptions in whaling catch data, suggest that the humpback whale breeding season should no longer be considered as confined to lower-latitude regions in winter. Rather, we suggest breeding extends geographically and temporally onto feeding grounds into at least spring and early summer. Singing at these times represents either low-cost opportunistic advertising by (perhaps relatively few) males to court females that failed to conceive during the winter, and/or possibly an intrasexual display. PMID- 15293860 TI - Red environmental noise and the appearance of delayed density dependence in age structured populations. AB - Previous work suggests that red environmental noise can lead to the spurious appearance of delayed density dependence (DDD) in unstructured populations regulated only by direct density dependence. We analysed the effect of noise reddening on the pattern of spurious DDD in several variants of the density dependent age-structured population model. We found patterns of spurious DDD in structured populations with either density-dependent fertility or density dependent survival of the first age class, inconsistent with predictions from unstructured population models. Moreover, we found that nonspurious negative DDD always emerges in populations with deterministic chaotic dynamics, regardless of population structure or the type of environmental noise. The effect of noise reddening in generating spurious DDD is often negligible in the chaotic region of population deterministic dynamics. Our findings suggest that differences in species' life histories may exhibit different patterns of spurious DDD (owing to noise reddening) than predicted by unstructured models. PMID- 15293861 TI - Evolution of the human ABO polymorphism by two complementary selective pressures. AB - The best-known example of terminal-glycan variation is the ABO histo-blood group polymorphism in humans. We model two selective forces acting on histo-blood group antigens that may account for this polymorphism. The first is generated by the invasion of opportunistic bacterial or other pathogens that interact with the epithelial-mucosal surfaces. The bacteria adapt to the microenvironments of common host phenotypes and so create frequency-dependent selection for rarer host alleles. The second is generated by intracellular viruses, and accounts for the observed differentials between the ABO-phenotype frequencies. It is thought that viruses acquire histo-blood group structures as part of their envelope from their previous host. The presence of host antigens on the viral envelope causes differential transmission of the virus between host types owing to the asymmetric action of ABO natural antibodies. Our model simulations show that these two forces acting together can account for the major features of the ABO polymorphism in humans. PMID- 15293862 TI - Host heterogeneity is a determinant of competitive exclusion or coexistence in genetically diverse malaria infections. AB - During an infection, malaria parasites compete for limited amounts of food and enemy-free space. Competition affects parasite growth rate, transmission and virulence, and is thus important for parasite evolution. Much evolutionary theory assumes that virulent clones outgrow avirulent ones, favouring the evolution of higher virulence. We infected laboratory mice with a mixture of two Plasmodium chabaudi clones: one virulent, the other avirulent. Using real-time quantitative PCR to track the two parasite clones over the course of the infection, we found that the virulent clone overgrew the avirulent clone. However, host genotype had a major effect on the outcome of competition. In a relatively resistant mouse genotype (C57B1/6J), the avirulent clone was suppressed below detectable levels after 10 days, and apparently lost from the infection. By contrast, in more susceptible mice (CBA/Ca), the avirulent clone was initially suppressed, but it persisted, and during the chronic phase of infection it did better than it did in single infections. Thus, the qualitative outcome of competition depended on host genotype. We suggest that these differences may be explained by different immune responses in the two mouse strains. Host genotype and resistance could therefore play a key role in the outcome of within-host competition between parasite clones and in the evolution of parasite virulence. PMID- 15293863 TI - The evolution of phylogenetic differences in the efficiency of digestion in ruminants. AB - This study investigates, for the first time (to our knowledge) for any animal group, the evolution of phylogenetic differences in fibre digestibility across a wide range of feeds that differ in potential fibre digestibility (fibre to lignin ratio) in ruminants. Data, collated from the literature, were analysed using a linear mixed model that allows for different sources of random variability, covariates and fixed effects, as well as controlling for phylogenetic relatedness. This approach overcomes the problem of defining boundaries to separate different ruminant feeding styles (browsers, mixed feeders and grazers) by using two covariates that describe the browser-grazer continuum (proportion of grass and proportion of browse in the natural diet of a species). The results indicate that closely related species are more likely to have similar values of fibre digestibility than species that are more distant in the phylogenetic tree. Body mass did not have any significant effect on fibre digestibility. Fibre digestibility is estimated to increase with the proportion of grass and to decrease with the proportion of browse in the natural diet that characterizes the species. We applied an evolutionary model to infer rates of evolution and ancestral states of fibre digestibility; the model indicates that the rate of evolution of fibre digestibility accelerated across time. We suggest that this could be caused by a combination of increasing competition among ruminant species and adaptation to diets rich in fibre, both related to climatically driven environmental changes in the past few million years. PMID- 15293864 TI - Gait selection in the ostrich: mechanical and metabolic characteristics of walking and running with and without an aerial phase. AB - It has been argued that minimization of metabolic-energy costs is a primary determinant of gait selection in terrestrial animals. This view is based predominantly on data from humans and horses, which have been shown to choose the most economical gait (walking, running, galloping) for any given speed. It is not certain whether a minimization of metabolic costs is associated with the selection of other prevalent forms of terrestrial gaits, such as grounded running (a widespread gait in birds). Using biomechanical and metabolic measurements of four ostriches moving on a treadmill over a range of speeds from 0.8 to 6.7 m s( 1), we reveal here that the selection of walking or grounded running at intermediate speeds also favours a reduction in the metabolic cost of locomotion. This gait transition is characterized by a shift in locomotor kinetics from an inverted-pendulum gait to a bouncing gait that lacks an aerial phase. By contrast, when the ostrich adopts an aerial-running gait at faster speeds, there are no abrupt transitions in mechanical parameters or in the metabolic cost of locomotion. These data suggest a continuum between grounded and aerial running, indicating that they belong to the same locomotor paradigm. PMID- 15293865 TI - Improvement of cutaneous microcirculation and oxygen supply in patients with chronic venous insufficiency by orally administered extract of red vine leaves AS 195: a randomised, double-blind, placebo-controlled, crossover study. AB - OBJECTIVE: To investigate the effect of the red vine leaf extract AS 195 on cutaneous microvascular blood flow, transcutaneous oxygen pressure (tcpO2), and leg oedema in patients with chronic venous insufficiency (CVI). DESIGN AND PATIENTS: The study was a randomised, double-blind, placebo-controlled, crossover trial for which 129 men and women, aged > or =18 years, with CVI stage I or II were screened. Seventy-one fulfilled the inclusion criteria and were randomised. INTERVENTIONS: A total of 71 patients were divided into two groups. The first group (n = 36) received AS 195 360mg once daily during a first 6-week treatment period, followed by a 4-week placebo washout period and then placebo during the second 6-week treatment period. The second group (n = 35) started with placebo and received AS 195 360mg after the placebo washout. The cutaneous microvascular blood flow in the malleolar region was measured using a newly developed laser Doppler device. TcpO2 was measured using a solid-state electrode. RESULTS: After 6 weeks, patients in the AS 195 group had increased microvascular blood flow values (+241.8 +/- 18.7 arbitrary units [AU] versus a decrease of -41.0 +/- 18.7AU in the placebo group; p < 0.0001). Oxygen increased to 1.35 +/- 0.97mm Hg (placebo: decrease of -7.27 +/- 0.97mm Hg; p < 0.0001). After 6 weeks of treatment the leg circumference was decreased (ankle level: by -0.39 +/- 0.09cm versus +0.29 +/- 0.09cm; p < 0.0001; calf level: by -0.54 +/- 0.05cm versus +0.14 +/- 0.05cm; p < 0.0001). CONCLUSION: The administration of AS 195 improved objective symptoms of CVI and may prevent CVI deterioration. PMID- 15293866 TI - Solifenacin demonstrates high absolute bioavailability in healthy men. AB - OBJECTIVE: Solifenacin succinate (YM905; Vesicare) is a promising new bladder selective muscarinic receptor antagonist under investigation for the treatment of overactive bladder. This study was designed to assess the absolute bioavailability of a single oral dose of solifenacin 10 mg, which is twice the suggested starting dose. STUDY DESIGN: Single-centre, open-label, randomised, two period, crossover, single-dose study. METHODS: Solifenacin was administered orally as a 10 mg dose and intravenously as a 5 mg dose. Oral and intravenous (IV) doses were divided by a washout period of > or =14 days. STUDY PARTICIPANTS: The study group consisted of 12 healthy young men, aged 20-45 years, nine of whom completed the study. RESULTS: The pharmacokinetic analysis comprised nine subjects. A single oral dose of solifenacin 10 mg had a high absolute bioavailability of 88.0% (95% confidence interval 75.8, 102.1), low clearance (9.39 L/h [SD 2.68]), and an extensive mean volume of distribution at steady state (599L [SD 86]). Only 7% of solifenacin was excreted intact in the urine. Single oral and IV administration of solifenacin was well tolerated in this study. The most common adverse events related to drug treatment were headache and somnolence. CONCLUSIONS: Pharmacokinetic analyses of single oral and IV doses of solifenacin demonstrated that the drug has a high absolute oral availability of 88%. This finding suggests that solifenacin may have a higher and less variable bioavailability than other antimuscarinic agents. PMID- 15293867 TI - AE 941. AB - AE 941 [Arthrovas, Neoretna, Psovascar] is shark cartilage extract that inhibits angiogenesis. AE 941 acts by blocking the two main pathways that contribute to the process of angiogenesis, matrix metalloproteases and the vascular endothelial growth factor signalling pathway. When initial development of AE 941 was being conducted, AEterna assigned the various indications different trademarks. Neovastat was used for oncology, Psovascar was used for dermatology, Neoretna was used for ophthalmology and Arthrovas was used for rheumatology. However, it is unclear if these trademarks will be used in the future and AEterna appears to only be using the Neovastat trademark in its current publications regardless of the indication. AEterna Laboratories signed commercialisation agreements with Grupo Ferrer Internacional SA of Spain and Medac GmbH of Germany in February 2001. Under the terms of the agreement, AEterna has granted exclusive commercialisation and distribution rights to AE 941 in oncology to Grupo Ferrer Internacional for the Southern European countries of France, Belgium, Spain, Greece, Portugal and Italy. It also has rights in Central and South America. Medac GmbH will have marketing rights in Germany, the UK, Scandinavia, Switzerland, Austria, Ireland, the Netherlands and Eastern Europe. In October 2002, AEterna Laboratories announced that it had signed an agreement with Australian healthcare products and services company Mayne Group for marketing AE 941 (as Neovastat) in Australia, New Zealand, Canada and Mexico. In March 2003, AEterna Laboratories announced it has signed an agreement with Korean based LG Life Sciences Ltd for marketing AE 941 (as Neovastat) in South Korea. The agreement provides AEterna with upfront and milestone payments, as well as a return on manufacturing and sales of AE 941. AEterna Laboratories had granted Alcon Laboratories an exclusive worldwide licence for AE 941 for ophthalmic products. However, this licence has been terminated. In 1999, AEterna secured funding for AE 941, part of which is from Technology Partnerships Canada (TPC), a research support programme run by Canada's federal government. Industry Canada will contribute $Can 1 for every $Can3 spent by AEterna on the development of AE 941, up to a total figure of $Can29.4 million. AEterna will reimburse TPC upon commercialisation of AE 941-derived products as a royalty on income generated. In January 2004 AEterna announced that development of AE 941 would be focusing on non-small cell lung cancer and that development for renal cell carcinoma would be discontinued. AEterna had previously announced in January 2003, following its acquisition of Zentaris, that development of AE 941 would be "strictly focused" on renal and non-small cell lung cancer, suggesting that development for all other indications has been discontinued, at least for the foreseeable future. PMID- 15293868 TI - Calcitonin intranasal--unigene: Salcatonin intranasal--unigene. AB - An intranasal spray formulation of recombinant salmon calcitonin [salcatonin] is in development with Unigene Laboratories as therapy for postmenopausal osteoporosis. Calcitonin is an endogenous polypeptide hormone that regulates calcium and bone metabolism. It is produced by the parafollicular cells of the thyroid gland in humans and other species. Calcitonin inhibits bone loss through the suppression of osteoclast activity. Salmon calcitonin is approximately 40-50 times more potent than natural human calcitonin at inhibiting osteoclast function. It can be obtained naturally from salmon or can be synthesised with the same chemical structure. Calcitonin was originally available only as an injectable formulation, but in recent years more convenient formulations have become available. Unigene is actively seeking to license its intranasal calcitonin product in Europe and other territories outside the US. nigene licensed its intranasal calcitonin product to Upsher-Smith Laboratories in December 2002, under a $US10 million exclusive US licensing agreement. Under the terms of the agreement, Unigene received an upfront payment of $US3 million from Upsher-Smith and will be eligible to receive milestone payments and royalty payments on product sales. Unigene will be responsible for manufacturing the product at its Boonton facility in New Jersey, USA, and will sell finished calcitonin product to Upsher-Smith. Upsher-Smith will package, market and distribute the product nationwide. Unigene granted an exclusive license to Faran Laboratories in September 2003 for its intranasal calcitonin osteoporosis product in Greece. Unigene will sell the finished product to Faran, who will promote and market it throughout the country after Unigene obtains European regulatory approval and local pricing approval. Unigene will receive an upfront payment and is eligible to receive milestone payments prior to product launch. Faran will pay Unigene a fixed price for each unit of product received. Qingdao General Pharmaceutical Company was a licensee for Unigene's injectable and intranasal calcitonin products in the People's Republic of China, and Unigene had received initial payments from Qingdao General Pharmaceutical in 1996. However, in June 2000, Unigene announced that it has entered into a joint venture with Shijiazhuang Pharmaceutical Group Company for the manufacture and distribution of injectable and intranasal calcitonin for the treatment of osteoporosis in China. Unigene initially will be responsible for supplying bulk calcitonin manufactured in its production facility in New Jersey and the joint venture will be responsible for filling, packaging, promoting and marketing the products. Unigene owns 45% of the contractual joint venture. Unigene is also developing oral and injectable formulations of calcitonin. In January 2004, Unigene announced it had received an approvable letter from the US FDA to market its calcitonin intranasal spray for the treatment of osteoporosis. The letter indicates that the FDA will approve the NDA upon finalisation of the labelling and resolution of specific remaining issues, including the submission of additional information and clinical data. Unigene filed the NDA in March 2003. Using the results from a pilot study completed in the UK, Unigene filed an IND with the FDA and began clinical testing of this intranasal calcitonin in the US. Clinical studies were successfully completed by year end 2001, demonstrating significant bone marker activity and similar serum concentrations between this product and that of an existing nasal calcitonin product. The European Union's regulatory authority, the Committee for Proprietary Medicinal Products (CPMP), has confirmed the efficacy of calcitonin formulations for the treatment of postmenopausal osteoporosis and other bone disorders. The CPMP has recommended revisions to and harmonisation within the European Union of the authorised indications for calcitonin formulations. The CPMP has determined that authorised indications for intranasal calcitonin will be approved for "treatment of established post-menopausal osteoporosis in order to reduce the risk of vertebral fractures", and new prescribing information will clarify that intranasal calcitonin does not appear to reduce the number of hip fractures. These recommendations will be implemented in the near future and will eliminate discrepancies between countries and between formulations. The Chinese regulatory authorities have granted Unigene an import license for calcitonin, and Unigene and Shijiazhuang have submitted an NDA in China. If this NDA is approved, the joint venture will have up to 6 years' market exclusivity for intranasal and injectable calcitonin. Unigene was granted a US patent for the intranasal formulation of calcitonin in 2002. PMID- 15293869 TI - Conivaptan: YM 087. AB - Conivaptan [YM 087], a benzazepine derivative, belongs to a series of highly potent, orally active arginine vasopressin V1 and V2 receptor antagonists that are being developed by Yamanouchi. Yamanouchi licensed conivaptan to Warner Lambert for co-development and marketing in the Americas, Europe and Africa. In return, Yamanouchi has rights to market atorvastatin in Japan. In June 2000, Warner-Lambert merged with Pfizer. The resulting company retained the Pfizer name. However, Yamanouchi and Pfizer discontinued the co-development and marketing agreement for conivaptan. Yamanouchi is continuing the independent development of conivaptan in the US and Europe. Yamanouchi is developing an oral drug delivery formulation of conivaptan for administration in patients with chronic heart failure. The company has initiated the ADVANCE (A Dose evaluation of a Vasopressin ANtagonist in CHF patients undergoing Exercise) trial, a double blind, multicentre trial in which 345 patients with heart failure will receive placebo or one of three doses of conivaptan for 12 weeks and their functional capacity will be assessed. Conivaptan demonstrated a potent diuretic effect in animal studies. PMID- 15293870 TI - Mitiglinide: KAD 1229, S 21403. AB - Mitiglinide [KAD 1229, S 21403], a derivative of benzylsuccinic acid, is a potassium channel antagonist undergoing development with Kissei for the treatment of type 2 diabetes mellitus. It has potent oral hypoglycaemic activity and is structurally different from the sulphonylureas, although it stimulates calcium influx by binding to the suphonylurea receptor on pancreatic beta-cells and closing K+ATP channels. Mitiglinide belongs to a family of meglitinide analogues that also includes repaglinide and nateglinide. Mitiglinide is licensed to Servier for Europe, where it is undergoing phase III development, and for Russia, the Commonwealth of Independent States, the Baltic Republics, the Middle East, Oceania, China (including Hong Kong) and Taiwan. Kissei exclusively licensed mitiglinide to Choongwae Pharma for South Korea in March 2003. In August 2002, Kissei and Takeda entered into a co-marketing agreement for mitiglinide in Japan. The companies will co-market the agent under a single brand name. Mitiglinide was licensed to Purdue Pharma for the US, Canada, Mexico and Central and South America. However, Kissei and Purdue Pharma terminated their agreement in February 2001 following Purdue Pharma's decision to concentrate on core areas such as oncology and analgesics. Kissei's US subsidiary, Kissei Pharma US, is currently carrying on the ongoing phase II clinical development in the US. However, in its Annual Report 2003, Kissei announced that it is considering outlicensing mitiglinide for development in marketing in North America. Mitiglinide has been recommended for approval in Japan for the management of postprandial hyperglycaemia in patients with type 2 diabetes. Kissei is also conducting phase II/III clinical trials with a combination of mitiglinide and an alpha-glucosidase inhibitor (additional indication) in patients with type 2 diabetes in Japan. In the US, the agent is being evaluated in phase II clinical trials with Kissei Pharma USA. Mitiglinide is also undergoing a phase-III, 12-month, multicentre, randomised, double-blind study in a total of 710 patients in comparison with repaglinide for the treatment of type 2 diabetes. This study will be followed by a 12-month open-label treatment with mitiglinide alone or in combination therapy. Servier (Australia) conducted a randomised, double-blind, multicentre phase III study in Australia comparing mitiglinide with metformin or a combination of the two for the treatment of type 2 diabetes. PMID- 15293871 TI - Natalizumab: AN 100226, anti-4alpha integrin monoclonal antibody. AB - Natalizumab [AN 100226, anti-alpha4 integrin monoclonal antibody, Antegren] is a humanised monoclonal antibody that blocks alpha4beta1 integrin-mediated leukocyte migration. Natalizumab is in phase III trials for the treatment of multiple sclerosis in North America and the UK, and for the treatment of Crohn's disease also in the UK. It may have potential in the treatment of other immune-related inflammatory disease. Elan Corporation intends to examine the potential of natalizumab in rheumatoid arthritis and ulcerative colitis. 4beta1 integrin on circulating leukocytes binds to vascular cell adhesion molecule-1, which is expressed at high levels in the blood vessels in the CNS during exacerbations of multiple sclerosis. This allows leukocytes expressing alpha4beta1 integrin (very late antigen-4) to move from the peripheral blood into the CNS. Inflammatory proteins and other factors released from lymphocytes in the brain lead to the progression of symptoms. A limitation of natalizumab is that it must be injected and cannot be administered orally. Scientists have transformed the large anti alpha4 monoclonal antibody into much smaller, drug-like molecules suitable for oral administration. Protein Design Labs has granted a worldwide nonexclusive licence under its antibody humanisation patents to Elan Pharmaceuticals for natalizumab. Biogen Inc. has entered into an agreement with Elan for a worldwide exclusive collaboration to develop, manufacture and commercialise natalizumab for multiple sclerosis and Crohn's disease and rheumatoid arthritis. Development of natalizumab is also being funded, in part, by Axogen (acquired by Elan in 1999). In November 2003, Biogen and IDEC Pharmaceuticals merged to form Biogen Idec. Elan repurchased royalty rights on a package of products, including natalizumab, from Autoimmune Disease Research Company. Elan and Genzyme Transgenics Corporation signed an agreement to produce natalizumab in GTC's genetically engineered goats, which will express the compound in their milk. Genzyme Transgenics Corporation changed its name to GTC Biotherapeutics in June 2002; it is no longer a subsidiary of Genzyme Corporation. Following discussions with the US FDA, Elan completed enrolment in a second phase III trial, involving approximately 420 patients with Crohn's disease. This Evaluation of Natalizumab as Continuous Therapy-2 (ENACT-2) trial evaluated the effect of natalizumab on duration of response and remission in patients with Crohn's disease. In January 2004, Elan Corporation and Biogen Idec announced that the phase III, ENACT-2 maintenance trial of natalizumab in Crohn's disease met the primary endpoint of maintenance of response. Elan and Biogen Idec will discuss these data with regulatory authorities in both the US and Europe and determine the appropriate path forward for natalizumab in Crohn's disease. An NDA for Antegren in Crohn's disease was expected to be filed at the end of 2003; however, due to failing to meet the primary endpoint in the induction trial, Elan is unable to predict when and if a regulatory filing will be made. Earlier, on 23 January 2001, the Wall Street Journal reported that the Biogen CEO expects Antegren to become a blockbuster drug, with sales of at least $US1 billion. He also predicted that Antegren could be on the market as early as 2003 for the indication of Crohn's disease and in 2004 for multiple sclerosis. The Journal stated that Biogen is under pressure to develop new drugs since its flagship product Avonex will be losing its US Orphan Drug Act protection in 2003. Antegren has a different mechanism to that of Avonex and could be used either alone or as a combination therapy. PMID- 15293872 TI - Procaine oral stabilised--Samaritan pharmaceuticals: SP-01, SP001. AB - Samaritan Pharmaceuticals (formerly Steroidogenesis Inhibitors) is developing a high-dose oral stabilised formulation of procaine hydrochloride [AnticortSP 01, SP001, Virucort] as an anticortisol drug for the treatment of HIV and AIDS. Preclinical investigations for its potential in the treatment of patients with Alzheimer's disease are also underway with Samaritan. The levels of cortisol, a hormone produced by the adrenal gland, are increased in many diseases, including AIDS. Serum cortisol levels are elevated during all stages of HIV infection, particularly in AIDS patients, whereas dehydroepiandrosterone (DHEA) levels are higher in the early stages of disease rather than in the advanced stages. Anticortisol drugs have been shown to reduce both the infectivity and the production of HIV in vitro. Anticort is available for licensing. Altachem Pharma had an exclusive licence to manufacture, use, distribute and sell oral procaine in Canada with an option to purchase the same rights for other Commonwealth countries. However, this agreement has been terminated. Cato Research in the US has been enlisted to implement a long-range clinical and business development plan to facilitate US FDA approval and manufacturing of the procaine formulation worldwide. Altachem Pharma received approval to begin a phase I trial of Anticort in Canada in HIV-positive patients receiving anti-HIV treatment. In January 2004, Cortisol Research International-Romania filed for orphan drug designation with the US FDA for Virucort 50 for the treatment of depression in children with HIV. A study examining the efficacy of Anticort in children infected with HIV has been conducted in Romania. One aspect of this study will investigate the possibility of reducing the dosage of zidovudine required if it is used in combination with Anticort. Following early indications that children treated with placebo in this trial have deteriorated while some children under the Anticort have shown improvements, especially in the 150-200mg group, Romanian clinicians have asked the Romanian FDA Commissioner for approval to switch children receiving placebo to receive Anticort. PMID- 15293873 TI - Recombinant human antithrombin III: rhATIII. AB - GTC Biotherapeutics (formerly Genzyme Transgenics Corporation) is developing a transgenic form of antithrombin III known as recombinant human antithrombin III [rhATIII]. It is produced by inserting human DNA into the cells of goats so that the targeted protein is excreted in the milk of the female offspring. The transgenic goats have been cloned in collaboration with the Louisiana State University Agriculture Center. GTC Biotherapeutics is conducting clinical trials of rhATIII in coagulation disorders. rhATIII is believed to be both safer and more cost-effective than the currently available plasma-derived product. rhATIII is also being investigated in cancer and acute lung injury. Genzyme Transgenics Corporation, originally a subsidiary of Genzyme Corporation, changed its name to GTC Biotherapeutics in June 2002; it is no longer a subsidiary of Genzyme Corporation. GTC Biotherapeutics is seeking partners for the commercialisation of rhATIII. Restructuring of GTC Biotherapeutics to support its commercialisation programmes was announced in February 2004. Genzyme Transgenics Corporation was developing rhATIII in association with Genzyme General (Genzyme Corporation) in the ATIII LLC joint venture, but in November 2000 a letter of intent was signed for the reacquisition of the rights by Genzyme Transgenics Corporation. It was announced in February 2001 that this reacquisition was not going to be completed and that the development of rhATIII was to continue with ATIII LLC. However, in July 2001, Genzyme Transgenics Corporation reacquired all the rights in the transgenic antithrombin III programme. SMI Genzyme Ltd, a joint venture between Sumitomo Metal Industries, Japan, and Genzyme Transgenics Corporation, USA, was set up to fund development of transgenic antithrombin III in Asia. However, in October 2000, Genzyme Transgenics Corporation reacquired, from Sumitomo Metal Industries, the rights to its technology for production of medicines from milk in 18 Asian countries, including Japan. The 10-year-old joint venture, SMI Genzyme Ltd, was dissolved. In June 2002, GTC Biotherapeutics estimated the current market size for plasma ATIII products to be approximately $US250 million - of which sales in Europe amounted to $US110 million, Japan $US130 million and the US $US10 million. PMID- 15293874 TI - Rotavirus vaccine--AVANT immunotherapeutics/GlaxoSmithKline: 89-12, RIX4414. AB - GlaxoSmithKline is developing a two-dose, oral rotavirus vaccine [Rotarix] under a licensing agreement with AVANT Immunotherapeutics. Rotarix is a live attenuated human rotavirus strain, which was developed by AVANT in collaboration with the JN Gamble Institute of Medical Research in the US. Originally, the research partner for the JN Gamble Institute was the Virus Research Institute, but the latter company merged with T Cell Sciences in August 1998 to form AVANT Immunotherapeutics. In 1997, AVANT signed an agreement with SmithKline Beecham (now GSK) for collaboration in the development of Rotarix. In December 2000 SmithKline Beecham merged with Glaxo Wellcome to form GlaxoSmithKline (GSK). The agreement grants GlaxoSmithKline exclusive worldwide marketing rights to the vaccine in exchange for royalties, licence fees and milestone payments to AVANT. Under the agreement, GlaxoSmithKline assumed all responsibilities for the development and manufacture of the vaccine following the successful completion of the US phase II study of Rotarix, for which AVANT received a milestone payment from GlaxoSmithKline. PMID- 15293875 TI - TLK 286. AB - TLK 286 [TELCYTA] is an antitumour agent in clinical development with Telik. It was developed through the application of Telik's proprietary TRAP chemogenomics technology. TLK 286 works by targeting tumours that overexpress glutathione S transferase (GST) P1-1, an enzyme that has been implicated in drug resistance and poor prognosis, and is elevated in solid tumours such as head and neck, breast, gastrointestinal, lung and ovarian tumours. TLK 286 is activated by GST P1-1 and, subsequently, initiates apoptosis in targeted tumour cells. Telik owns worldwide rights to TLK 286 and intends to commercialise it in the North American market. The company plans to select a collaborator in other territories with capabilities in manufacturing, sales and marketing. Telik was previously collaborating with Taiho, Japan, on development of TLK 286, but this agreement appears to have been terminated. Telik announced in October 2002 that it had begun the first of a series of planned clinical trials of TLK 286 in combination with docetaxel in patients with non-small cell lung cancer. A phase II trial was initiated in May 2001 in patients with ovarian cancer. Two additional combination trials were initated in ovarian cancer patients in December 2002. One of the trials will evaluate the combination of TLK 286 with Doxil in patients who have failed platinum-based chemotherapy. The second trial will evaluate TLK 286 in combination with carboplatin in patients who have recurrent, platinum-sensitive ovarian cancer. Telik held a successful phase III meeting with the US FDA to discuss plans for the first registration trial of TLK 286, and in March 2003 it was announced that a phase III registration trial of TLK 286 as monotherapy had been initiated in platinum-refractory ovarian cancer patients. The multinational trial has been designated the ASSIST-1 (Assessment of Survival in Solid Tumours 1) trial. Telik plans to enrol approximately 440 women who will be assigned to either TLK 286 treatment or a control (doxorubicin liposomal or topotecan). PMID- 15293876 TI - Vincristine liposomal--INEX: lipid-encapsulated vincristine, onco TCS, transmembrane carrier system--vincristine, vincacine, vincristine sulfate liposomes for injection, VSLI. AB - INEX Pharmaceuticals is developing a liposomal formulation of vincristine [Onco TCS, vincacine, VSLI, Vincristine sulfate liposomes for injection] for the treatment of relapsed aggressive non-Hodgkin's lymphoma (NHL) and other cancers. It is being developed using INEX's proprietary drug-delivery technology platform called the transmembrane carrier systems (TCS), which enables the targeted intracellular delivery of various therapeutic agents. Liposomal vincristine is expected to have certain advantages over the existing standard preparation of vincristine because the use of TCS technology enables the vincristine to circulate in the blood for longer, accumulate in the tumour, and be released over an extended period of time at the tumour site. The application of TCS technology to any agent, including vincristine, has the potential to increase the efficacy and decrease the side effects of the agent. INEX decided in 1998 to focus on gaining approval for liposomal vincristine in the treatment of relapsed aggressive NHL because no standard therapy was approved for this indication. In 1999, liposomal vincristine was granted accelerated development status by the US FDA, which enables the FDA to approve it based on the surrogate endpoint of a single clinical trial. In addition, the FDA granted liposomal vincristine fast track status in August 2000. In April 2001, INEX and Elan Corporation formed a joint venture for the development and commercialisation of liposomal vincristine, with both companies contributing assets to the venture including worldwide rights to the product and intellectual property rights. The joint venture was called IE Oncology. However, in June 2002, Elan announced that it was going to focus its business strategy on three specific areas, which would not include cancer therapies. INEX announced it had regained 100% ownership of liposomal vincristine in April 2003, by reacquiring the 19.9% equity interest held by Elan and in addition retaining a fully paid-up licence to Elan's intellectual property pertaining to liposomal vincristine. All obligations to Elan under the agreement will be met through three milestone payments totalling $8 million. Some of the milestones may be paid in shares valued at the then current market price. In January 2004, INEX and Enzon Pharmaceuticals formed a strategic partnership to develop and commercialise liposomal vincristine. Under the terms of the agreement, Enzon receives the exclusive North American commercialisation rights for liposomal vincristine for all indications. INEX will receive upfront and milestone payments as well as a percentage of commercial sales. Additionally, the formation of this partnership triggered a US$3 million payment from INEX to the former joint venture partner, Elan Corporation. Nine clinical trials of liposomal vincristine are currently being conducted, including one phase I/II trial and eight phase II trials. A phase II/III trial was completed in December 2002. In September 2003, Inex commenced a 'rolling submission' for liposomal vincristine by submitting the first of three major sections of the NDA to the FDA. The second major section was submitted to the FDA in December 2003. INEX expects to complete the filing with the submission of the clinical section of the NDA in the first quarter of 2004. Dow Jones Newswires reported on 1 October 2001 that the CEO of INEX expects Onco TCS to achieve sales of between $US100 and $US400 million annually for the company. FDA approval was then predicted for late 2002 or early 2003. PMID- 15293877 TI - Influence of innate immune responses on autoimmunity. PMID- 15293878 TI - Murine lupus autoantibodies identify distinct subsets of apoptotic bodies. AB - The specific modification of autoantigens and their redistribution into blebs at the surface of apoptotic cells contribute to the induction of autoimmune responses. Blebs containing fragments of the apoptotic nucleus separate from the remainder of the cell to form membrane-bound sub-cellular particles (SCPs), otherwise known as apoptotic bodies. To determine whether apoptotic bodies containing nuclear antigens represent a defined subset of SCPs, we examined the heterogeneity of particles generated by Jurkat cells following synchronization of the cell cycle by serum withdrawal and inhibition of topoisomerase I by camptothecin. Particles were purified by filtration, incubated in the presence of antinucleosome or anti-cardiolipin autoantibodies, annexin V, and Sytox Orange and analyzed by flow cytometry and confocal microscopy. We demonstrate that nuclear autoantigens are associated with one clearly defined subset of SCPs that can be distinguished from other products of late apoptosis. Our experiments represent an important step towards characterizing the heterogeneity of SCPs that are generated in late apoptosis and identifying their contributions to tolerance and autoimmunity. PMID- 15293879 TI - Complement activation by apoptotic cells occurs predominantly via IgM and is limited to late apoptotic (secondary necrotic) cells. AB - Apoptotic cells activate complement via various molecular mechanisms. It is not known which of these mechanisms predominate in a physiological environment. Using Jurkat cells as a model, we investigated complement deposition on vital, early and late apoptotic (secondary necrotic) cells in a physiological medium, human plasma, and established the main molecular mechanism involved in this activation. Upon incubation with recalcified plasma, binding of C3 and C4 to early apoptotic cells was similar to background binding on vital cells. In contrast, late apoptotic (secondary necrotic) cells consistently displayed substantial binding of C4 and C3 and low, but detectable, binding of C1q. Binding of C3 and C4 to the apoptotic cells was abolished by EDTA or Mg-EGTA, and also by C1-inhibitor or a monoclonal antibody that inhibits C1q binding, indicating that complement fixation by the apoptotic cells was mainly dependent on the classical pathway. Late apoptotic cells also consistently bound IgM, in which binding significantly correlated with that of C4 and C3. Depletion of plasma for IgM abolished most of the complement fixation by apoptotic cells, which was restored by supplementation with purified IgM. We conclude that complement binding by apoptotic cells in normal human plasma occurs mainly to late apoptotic, secondary necrotic cells, and that the dominant mechanism involves classical pathway activation by IgM. PMID- 15293880 TI - Accelerated autoimmune disease in MRL/MpJ-Fas(lpr) but not in MRL/MpJ following immunization with high load of syngeneic late apoptotic cells. AB - Numerous studies have shown that autoantigens may be clustered in the blebs of apoptotic cells. However, it is not yet clear in what circumstances apoptotic cells could be immunogenic rather than tolerogenic when interacting with macrophages, dendritic cells, and B cells. In order to further study this question we compared immunization of high load of syngeneic late apoptotic cells in two genetically close pro-autoimmune mice strains: MRL/MpJ and MRL/MpJ Fas(lpr). We show that high apoptotic load could accelerate the generation of anti-dsDNA and anticardiolipin, and the extent of kidney disease, in MRL/MpJ Fas(lpr) but could not generate autoimmunity in MRL/MpJ. Thus, in this model, a high load of apoptotic cells could augment the autoimmune response in established autoimmunity, but did not generate de novo autoimmune response in pro-autoimmune mice. Taken together with previous observations, apoptotic cell load may modify autoimmune disease generating either immune inhibition and down regulation of autoimmunity or immune stimulation and acceleration of an autoimmune disease. PMID- 15293882 TI - Role of surfactant protein A in the innate host defense and autoimmunity. AB - Surfactant protein A (SP-A) is a lung collectin with diverse immunoregulatory activities. SP-A regulates the innate host defense by enhancing phagocytosis of pathogens and modulating the production of nitric oxide and cytokines by immune cells. Additionally, SP-A also modulates the phenotypic and functional properties of the cells of adaptive immune response such as dendritic cells (DCs) and lymphocytes. Bone marrow-derived DCs generated in the presence of SP-A fail to increase lipopolysaccharide-induced upregulation of major histocompatibility complex (MHC) class II and CD86 costimulatory molecule on DCs surface and behaves like "tolerogenic DCs". SP-A may also induce tolerance by suppressing the proliferation of activated T lymphocytes. Thus, based on immunoregulatory properties of SP-A, it may be employed as a therapeutic agent for the treatment of autoimmune disease and organ transplantation. PMID- 15293881 TI - CR1/CR2 deficiency alters IgG3 autoantibody production and IgA glomerular deposition in the MRL/lpr model of SLE. AB - CR1 and CR2 expression is decreased by approximately 50% on B cells of patients with systemic lupus erythematosus (SLE). Expression is also decreased in the MRL/lpr murine model of SLE prior to the development of clinical disease, suggesting that this alteration may play a role in pathogenesis. To determine whether the decrease in receptor levels affects the development of SLE, we analyzed MRL/lpr mice in which CR1/CR2 expression was altered by gene targeting. Mice from each cohort (Cr2+/+, Cr2+/-, and Cr2-/-) were analyzed biweekly for the development of proteinuria and autoantibodies. Kidneys were examined at 12 and 16 weeks for evidence of immune complex deposition and renal disease. Deficiency of CR1/CR2 did not affect survival or development of renal disease as measured by proteinuria. Mice deficient in CR1/CR2 had significantly lower levels of IgG3 rheumatoid factor (RF) and total serum IgG3, suggesting a specific defect in production of IgG3 in response to endogenous autoantigens. Since IgG3 RF has been associated with the development of vasculitis in this model, we examined the mice for alterations in development of this clinical manifestation. Although there was no difference in the development of ear necrosis among the three groups, renal arteritis was not identified in any of the Cr2+/- mice, whereas it was present in 20% of the Cr2+/- and 40% of the Cr2+/+ mice. Finally, significantly higher levels of IgA were seen in the glomeruli of Cr2+/- mice compared to Cr2+/- or Cr2+/+ mice, suggesting that CR1/CR2 are involved in either the regulation of IgA production or the clearance of IgA immune complexes. Together these data support the concept that alterations in CR1/CR2 expression or function affect the regulation of autoantibody production and/or clearance and may have clinical consequences. PMID- 15293883 TI - Mast cells and innate cytokines are associated with susceptibility to autoimmune heart disease following coxsackievirus B3 infection. AB - The development of autoimmune disease involves a combination of genetic and environmental factors. Many autoimmune diseases are believed to be triggered by viral infections. Since the early, natural immune response to infection can determine the later development of the adaptive immune response, innate immunity likely influences the progression from viral immunity to autoimmunity. To investigate the role of the innate immune response on susceptibility to autoimmune disease, we compared the early cytokine response of mice susceptible or resistant to the development of autoimmune heart disease following viral infection. We found that susceptible BALB/c mice produced elevated levels of TNF alpha, IL-1beta, and IL-4 within hours of Coxsackievirus B3 (CB3) infection. These cytokines are known to be critical for the development of autoimmune heart disease, and are also rapidly produced from activated mast cells (MC). Degranulating MC were observed as early as 6 h following CB3 infection in the heart, and significantly higher numbers of MC were found in the spleen of susceptible BALB/c mice at this time. Thus, susceptibility to autoimmune heart disease can be determined as early as 6 h following viral infection in susceptible strains of mice. PMID- 15293884 TI - The role of NK cells and NK cell receptors in autoimmune disease. AB - Natural killer (NK) cells are the first line of defense against infection and transformation. Additionally, NK cells can play seemingly opposite roles in autoimmune disease. Here, we summarize the functions of NK cells as both regulators and inducers of autoimmune disease. The role NK cells play depends on which cells become targets for NK cell attack. The activity of NK cells is controlled by inhibitory receptors specific for MHC Class I molecules, and by activating receptors with diverse specificities. The ligands for both activating and inhibitory receptors are present on potential target cells. It is the balance in expression of these different ligands that determines NK cell activation and therefore whether the cell becomes a target for NK cell-mediated killing. We further discuss the roles of NK cell receptors and their ligands in autoimmune disease. PMID- 15293885 TI - The IL-12 role in donor cell engraftment in a murine model of semiallogenic GVH disease with signs of autoimmune disease. AB - We analyzed the IL-12 effect in an autoimmune disease induced in a semiallogenic murine model of graft-vs-host disease (GVHD) Balb/c semiallogenic lymphoid cells i.v. infected in hybrid mice (Balb/c x A/J) F1 (CAF). IL-12 was administered 1 h before cell transplantation following two different protocols: (a) injecting 2 microg of mrIL-12 (murine recombinant IL-12) per mouse before the first semiallogenic cell injection; or (b) injecting the 2 microg of mrIL-12 fractionated in 5 days. ATh1 response was produced but an acute GVHD did not appear although differences in class I and II major histocompatibility complex (MHC) antigens were present. Four days after the semiallogenic cell transfer, IL 12 treated mice showed a marked reduction in the percent of spleen B cells compared with CAF1 control and CAF1 + Balb/c GVHD mice. After 5-6 months of follow-up, the donor cell chimerism increased significantly in spleen (70 +/- 31 vs. 43 +/- 31%) and in thymus. Flow cytometry of spleen lymphocytes demonstrated that donor chimerism was made up of TCD4, TCD8 and B lymphocytes and was higher in animals injected with IL-12. Moreover, CD8 T lymphocytes were 100% donor origin in the IL-12-injected group of GVHD animals and 50% origin in the IL-12 non-injected CAF1 + Balb/c group of animals. This paper shows that: (1) IL- 12 may play a role in the mechanisms of donor cell engraftment, probably produced by a CTL donor anti-host mechanism; (2) no acute GVHD was induced in spite of class I and II MHC differences; (3) IL-12 did not show any effect on the AR-like clinical signs of disease developed in this model of GVHD although histological subclinical signs were less frequent, and no glomerulonephritis was detected in the IL-12-treated GVHD mice. PMID- 15293886 TI - Diagnostic accuracy of ELISA methods as an alternative screening test to indirect immunofluorescence for the detection of antinuclear antibodies. Evaluation of five commercial kits. AB - Detection of antinuclear antibodies (ANA) is a fundamental laboratory test for diagnosing systemic autoimmune diseases. Currently, the method of choice is indirect immunofluorescence (IIF) on a HEp-2 cell substrate. The goal of this study was to evaluate the diagnostic accuracy of five commercially available enzyme immunoassay (EIA) kits for ANA detection and to verify the possibility of using them as an alternative to the IIF method. The study involved 1513 patients, 315 of whom were diagnosed with a systemic autoimmune disease and 1198 in whom an autoimmune disorder was excluded. For all sera, ANA detection was performed via IIF and with five different EIA kits. The results were evaluated in relation to clinical diagnosis and the presence of possible specific autoantibodies (anti-ENA or anti-dsDNA); lastly, they were compared with the results obtained using ANA IIF as the method of reference. The positive rate of the ANA-IIF test in subjects with systemic autoimmune diseases was 92%, whereas in the five ANA-EIA kits there was broad diversity in terms of response, with positive rates ranging from 74 to 94%. All the EIA kits correctly detected the presence of antibodies (anti-dsDNA, anti-RNP, anti-Ro/SSA) responsible for homogeneous and speckled fluorescence pattern, but at the same time they showed substantial inaccuracy with the nucleolar pattern, with a mean sensitivity of approximately 50% in this case. Instead, there was a large kit-to-kit difference in terms of identification of anti-Scl70 and centromere patterns, for which sensitivities ranged between 45 and 91%, and between 49 and 100%, respectively. The results of the study demonstrate that the commercially available ANA-EIA kits show different levels of sensitivity and specificity. Some of them have a diagnostic accuracy that is comparable and, in some cases, even higher than the IIF method. Consequently, these could be used as an alternative screening test to IIE. However, others do not ensure acceptable results. Therefore, careful evaluation of the various kits on the market is advisable before including any of these methods in the clinical and diagnostic testing. PMID- 15293887 TI - Estrogens and women's health: a scary or a fairy tale? PMID- 15293888 TI - Serum levels of prostate-specific antigen and androgens after nasal administration of a gonadotropin releasing hormone-agonist in hirsute women. AB - We aimed to determine whether ovarian suppression affects the production rate of prostate-specific antigen (PSA) in hirsute women. A total of 34 hirsute women who had a modified Ferriman-Gallwey (FG) score of > or = 7 and 14, non-hirsute women as the control group were recruited for this prospective controlled study. Serum samples for evaluation of basal hormones and PSA concentration were collected and were analyzed by commercial kits and chemiluminescent enzyme immunoassay. The hirsute women were given 400 microg/day nafarelin acetate for 3 months. Basal hormones, PSA levels and FG scores were then assessed. ANOVA and Tukey test were used to compare differences in means between the hirsute and the non-hirsute group at the beginning of the study. Student's t test, Tukey test and repeated measures variance analysis were used to evaluate differences in the study group and between the women with polycystic ovary syndrome (PCOS) and idiopathic hirsutism after gonadotropin releasing hormone (GnRH)-agonist administration. Statistical significance was assumed with a value of p < 0.05. PCOS and idiopathic hirsutism were diagnosed in 58.8% and 41.2% of 34 hirsute women, respectively. Age and body mass index (BMI) were similar in the hirsute and the control group (p > 0.05). FG scores in the PCOS group (20.3 +/- 1.7) were statistically similar to those of the group with idiopathic hirsutism (17.6 +/- 1.7) (p > 0.05). The non-hirsute women had significantly lower serum PSA concentrations than the hirsute group (p < 0.001). The basal mean level of PSA was 0.095 +/- 0.001 in the PCOS, 0.0061 +/- 0.009 in the idiopathic hirsute and 0.0040 +/- 0.004 ng/ml in the control group. No significant difference in the mean PSA levels was noted between the PCOS and the idiopathic hirsute subgroups before and after GnRH agonist treatment (0.0096 +/- 0.01 and 0.0051 +/- 0.032 ng/ml, respectively) (p > 0.05). FG scores, testosterone, 17alpha hydroxyprogesterone and dehydroepiandrosterone sulfate levels were significantly decreased in the hirsute group following treatment (p < 0.001). PSA levels in hirsute women were higher than in non-hirsute women and independent of BMI, age and androgen deprivation. PSA concentration may be mediated through extragonadal sites and possibly through a long-standing hyperandrogenemic environment such as in PCOS and idiopathic hirsutism. Further investigation as to the significance of PSA in women with hirsutism and whether antiandrogens directly act to inhibit biosynthesis of PSA is warranted. PMID- 15293889 TI - Effect of nitric oxide on contractions of uterine and cervical tissues from pregnant rats. AB - Our objective was to evaluate the role of the nitric oxide (NO)-cyclic guanosine monophosphate (cGMP) pathway in rat uterine and cervical contractility at mid- and late gestation. Rings of uterus and cervix from Sprague Dawley rats on day 14 of pregnancy (mid-) and day 21 of pregnancy (late) were equilibrated at 2 g passive tension in organ chambers filled with Krebs-Henseleit solution and bubbled with 5% CO2 in air (37 degrees C, pH approximately 7.4). Rings were treated with an inhibitor of outward potassium current, tetraethylammonium, to activate phasic contractions, and the concentration-response relationships to diethylamine/NO and 8-bromo-cGMP (8-br-cGMP) were assessed. Baseline spontaneous activity was significantly higher at term gestation in both uterine and cervical rings compared with mid-gestation. Spontaneous contractile activity was not apparent in cervical rings from rats in mid-gestation, but was persistent after treatment with tetraethylammonium. Oxyhemoglobin did not affect NO-induced inhibition of activation by tetraethylammonium contractile activity in either cervical or uterine tissues in mid- or late gestation. The 8-br-cGMP concentration-dependently inhibited tetraethylammonium-activated contractions that were more pronounced in uterine tissues compared with cervical tissues in both mid- and late gestation. We concluded that activation of the NO-cGMP pathway inhibits both uterine and cervical smooth muscle contractility. Both tissues demonstrated refractoriness to NO at term. PMID- 15293890 TI - Veralipride administered in combination with raloxifene decreases hot flushes and improves bone density in early postmenopausal women. AB - We evaluated the administration of raloxifene and veralipride in postmenopausal women with high osteoporosis risk and hot flushes in whom hormone replacement therapy (HRT) was contraindicated. A group of early postmenopausal women (n = 29) (mean age 51.8 +/- 4.1), complaining of severe vasomotor symptoms and with a bone mineral density (BMD) T-score between -1.5 and -2.5 were evaluated. They were randomly assigned to two treatment groups: raloxfene (60 mg/day) continuously in association with veralipride (100 mg/day) on alternate days (n = 17); or on alternate months (n = 12). BMD, serum prolactin concentration and endometrial thickness were assessed at baseline and after 6 months of therapy. Kupperman Index and hot flushes were assessed before and after 3 and 6 months of therapy. BMD was significantly higher at the end of therapy with an increase of 1.1%. Kupperman Index was significantly reduced after 3 months and a further decrease at 6 months was observed with both protocols. Both treatments led to a significant reduction of hot flushes after 3 and 6 months. No signifcant changes of prolactin levels were observed in either protocol. We found that the combined raloxifene-veralipride treatment, both every other day and every other month, led to a significant improvement in bone density and was effective in hot flushes and other menopause-associated symptoms. These protocols could represent a new way to administer raloxifene in early postmenopausal women at high osteoporosis risk with HRT contraindication. PMID- 15293891 TI - Do lipid profiles of postmenopausal women under oral hormone replacement therapy remain stable or reveal a multiphasic course in time? AB - The object of this study was to compare the effects of oral conjugated estrogen (CEE) alone, CEE plus medroxyprogesterone acetate (MPA) and tibolone on lipid profiles, and investigate whether these effects change in time. Plasma lipid levels were studied for CEE (n = 49), CEE + MPA (n = 50) and tibolone (n = 51). Mean per cent changes at certain intervals were compared with their previous intervals for each therapy. Paired t-test was used for statistical analysis. CEE alone had raised high-density lipoprotein (HDL) and triglyceride levels and lowered total cholesterol and low-density lipoprotein (LDL) levels at the end of the 2-year study period. Addition of MPA to the CEE regimen weakened the effect on HDL and triglyceride, augmented the decrease in total cholesterol and did not affect LDL. The tibolone group revealed similar but more prominent effects in total cholesterol and LDL levels. HDL and triglyceride levels were significantly below baseline in the first 6 months, but HDL changes vanished and triglyceride levels remained decreased at the end of 2 years. These data did not support a correlation between lipid levels and the biphasic incidence of cardiac events that were observed in the Heart and Estrogen/progestin Replacement Study (HERS), but revealed period-dependent changes in the tibolone group. PMID- 15293892 TI - Psychological well-being and the dyadic relationship of Chinese menopausal women (and their spouses) attending hormone replacement clinics. AB - This survey examined the general health and the marital relationship of 122 Chinese menopausal women and their spouses attending hormone replacement clinics. Climacteric symptoms of the participants were assessed by the modified Greene Climacteric Scale (GCS). The psychological well-being of the participants and their spouses was assessed by the 12-item Chinese General Health Questionnaire (GHQ), and their marital quality was assessed by the Chinese Dyadic Adjustment Scale (DAS). The GCS scores of our cohort were significantly higher than that of a community-based sample of menopausal Chinese women. About one-third of the participants and one-fifth of their spouses suffered from reduced psychological well-being. Although the DAS total scores of the participants and their spouses were comparable to those of the adjusted couples in a younger population, the affectional DAS subscores were significantly lower. The GCS scores of the menopausal women were significantly positively correlated with their GHQ scores but negatively correlated with their DAS scores. In summary, the menopausal women attending the hormonal replacement clinics, especially those with more dimacteric symptoms, suffered from significant psychiatric morbidity and marital maladjustment. The psychological dimension of the menopause should never be neglected. PMID- 15293893 TI - Fluctuations of fatty acid amide hydrolase and anandamide levels during the human ovulatory cycle. AB - Implantation is possible within a defined period of the menstrual cycle, referred to as the 'implantation window'. It is during this critical period that proper dialog can be established between the blastocyst and a receptive endometrium. If for any reason this dialog is not established or is altered, the embryo is aborted. The factors responsible for the interaction between the embryo and the mother at the moment of implantation remain poorly understood. Recent studies indicate that endocannabinoids may contribute to the development of an adequate milieu at the implantation site. Here we show that the levels of anandamide and of its degrading enzyme, the fatty acid amide hydrolase, in peripheral lymphocytes undergo specific variations during the various phases of the human ovulatory cycle. In particular, we found the highest levels of fatty acid amide hydrolase activity and protein content, paralleled by the lowest anandamide concentrations, in the period that temporally coincides with the putative window of implantation in humans. On the other hand, the anandamide-synthesizing phospholipase D, the anandamide membrane transporter and the anandamide-binding cannabinoid receptors of lymphocytes did not change during the menstrual cycle. This study indicates that high fatty acid amide hydrolase activity and low anandamide levels may be among the factors that co-operate in the success of implantation. This would add to our understanding of the pathophysiological and therapeutic implications of the endocannabinoid system in human fertility. PMID- 15293894 TI - Comparative analytical evaluation of thyroid hormone levels in pregnancy and in women taking oral contraceptives: a study from an iodine deficient area. AB - Increase of serum thyroxine binding globulin (TBG) resulting from estrogen action may lead to problems in thyroid diagnostics. The aim of the present study was to define the most diagnostically reliable thyroid parameters in women exposed to differentially elevated estrogens. Sera of three groups of healthy women were analyzed: women taking no medicine (controls), those taking oral contraceptives and pregnant women (in weeks 16 or 32 of gestation). All women involved in the study lived in a moderately iodine-deficient geographical area. Thyroid stimulating hormone (TSH), TBG, total thyroxine (T4), total tri-iodothyronine (T3) and free T3 were determined and free T4 indices (total T4 x T3 uptake; total T4/thyroxine binding capacity (TBC); total T4/TBG) were calculated. Free T4 was measured simultaneously with a one-step T4-analog enzyme-linked immunosorbent assay (ELISA), a labeled T4 antibody radioimmunoassay (RIA), and a two-step microparticle enzyme immunoassay (MEIA). Estrogen-dependent differences were found in all investigated parameters; however, they remained in the reference interval for TSH, total T4 x T3 uptake, total T4/TBC,free T3 and free T4 MEIA. It was concluded that simultaneous estimations of free T4 and free T3 should follow a primary TSH measurement. The necessity of a distinct reference range has emerged for free thyroid hormones in midterm and late pregnancy as well as in the use of oral contraceptives, especially in iodine-deficient areas. PMID- 15293895 TI - The influence of 5-year therapy with tibolone on the lipid profile in postmenopausal women with mild hypercholesterolemia. AB - Our objective was to investigate the effects of 5-year therapy with tibolone on the lipid profile in postmenopausal women with mild hypercholesterolemia (total cholesterol, 241 +/- 7 mg/dl; LDL cholesterol, 153 +/- 9 mg/dl). Eighty-two patients were divided into two groups. Group A (53 women) received 2.5 mg of tibolone per day. Group B (29 women) received no tibolone. Total, low- and high density lipoprotein cholesterol and lipoprotein(a) were found to be decreased in the tibolone group, by 17.7%, 32%, 15.5% and 12%, respectively (p < 0.01) throughout the 5-year treatment, while triglycerides showed no significant change. The lipid profile in the control group remained at its initial values. Menopausal symptoms disappeared in the treatment group within the first 5 months, whereas they deteriorated in the control group during the first 2 years. Although a few unwanted side-effects on hormone-dependent tissues were observed (including vaginal spotting in 11.3% and febrile hemorrhagic cystic mastopathy in 3.8%) long term therapy with tibolone seemed to be well tolerated, and appeared to have a beneficial effect on the levels of serum lipids. PMID- 15293896 TI - [The epidemiology of nasopharyngeal colonisation of Streptococcus pneumoniae in children with respiratory tract infection]. AB - Nasopharyngeal swab specimens from 324 children with respiratory tract infections were evaluated to detect the rate of Streptococcus pneumoniae colonisation. S. pneumoniae was detected in 92 (28%) of the subjects. Forty three (46.7%) of the isolates were serotyped by the capsular swelling tests. The most common serotypes were 19F, 6B, 3 and 23F. The 7, 9 and 11 valent conjugate pneumococcal vaccines covered 35.8%, 40% and 46.7% of all the S. pneumoniae isolates, respectively. Thirty two (34.8%) isolates exhibited penicillin MIC values between 0.1 and 1 microg/ml, only 1 isolate had MIC > or = 2 microg/ml. Penicillin resistant pneumococcal colonisation was most frequently detected in children with viral upper respiratory tract infections (12.5%). Resistance rates of trimetoprim sulfamethoxazole, erythromycin, chloramphenicol, clindamycin, ceftriaxone, rifampin were 31.5%, 9.8%, 6.5%, 4.3%, 1%, 0%, respectively. Being a children of a family with low income was the only risk factor for colonisation with S. pneumoniae, whereas having a sibling attending to a day care center, antibiotic use in the last three months and use of more than one antibiotic were significant risk factors for colonisation with penicillin resistant S. pneumoniae (p < 0.05). PMID- 15293897 TI - [Molecular epidemiology of Stenotrophomonas maltophilia strains isolated from paediatric patients]. AB - As alternative regimens in therapy of Stenotrophomonas maltophilia infections are limited, epidemiological investigation of S. maltophilia is necessary for developing new strategies in prophylaxis of these infections. "Enterobacterial repetetive intergenic consensus-polymerase chain reaction (ERIC-PCR)", is a simple and rapid method to detect the genomic polymorphism at the strain level in nosocomial outbreaks due to S. maltophilia. The aim of this study was to investigate the molecular epidemiology of S. maltophilia strains isolated from paediatric patients in a university hospital. A total of 40 clinical isolates one from each of 35 patients, two pairs from two patients and one from an environmental source, were collected between 1998-2001 period. Susceptibility to eleven antimicrobials was studied by agar dilution test according to NCCLS criteria. Resistance rates of the tested isolates against piperacillin, piperacillin-tazobactam, ceftriaxone, ceftazidime, cefepime, ciprofloxacin, imipenem, meropenem, amikacin, gentamicin and trimethoprim-sulfamethoxazole were 57.5%, 25%, 85%, 37.5%, 47.5%, 20%, 100%, 95%, 62.5%, 62.5% and 5%, respectively. Trimethoprim-sulfamethoxazole was the most active agent against the tested isolates. The isolates were analyzed by ERIC-PCR by using each of ERIC-I and ERIC II primers. Genotypic analysis by ERIC-PCR identified 24 different major and 4 minor profiles. Three isolates from each of four different groups, two isolates from each of other four different groups were unique. The results of this study, revealing a great genomic diversity and low clonality within the strains of S. maltophilia strains suggest that patients may be colonized with the naturally occuring isolates in the hospital environment and then get infected with these strains under the selective antibiotic pressure, thus supporting the view that clonal spread of S. maltophilia strains between patients is of rather low possibility. PMID- 15293898 TI - [Investigation of the effects of efflux pump inhibitors on ciprofloxacin MIC values of high level fluoroquinolone resistant Escherichia coli clinical isolates]. AB - In this study, effects of efflux pump inhibitors, phe-arg-beta-naphthylamide (MC 207,110), verapamil, omeprazole and lansoprazole, on ciprofloxacin minimal inhibitory concentrations (MIC) against Escherichia coli clinical isolates with high level fluoroquinolone (FQ) resistance, were evaluated. Fourteen FQ resistant clinical isolates of E. coli isolated from urine samples and identified by BD BBL CRYSTAL Identification Systems Enteric/Nonfermenter ID kit (Becton Dickinson and Company Sparks, USA) were tested. In order to investigate the effects of inhibitory agents, MIC values were determined by broth microdilution method in the absence or presence of 20 microg/ml MC-207,110, verapamil, omeprazole and lansoprazole. All 14 clinical isolates were resistant to ciprofloxacin (MIC range; 16-512 microg/ml). No difference was observed on ciprofloxacin MIC values in the presence of 20 microg/ml omeprazole, whereas MIC values were decreased 2 folds in 2 isolates in the presence of verapamil and lansoprazole, 2 folds in 6 isolates and 4 folds in 2 isolates in the presence of MC-207,110. In conclusion, we observed that there were no effects of the compounds used in our study, on ciprofloxacin resistance of E. coli clinical isolates. PMID- 15293899 TI - [Determination of serologic markers against bacterial atypical pneumonia agents in pneumonia patients]. AB - Approximately one third of all community acquired pneumonia cases are caused by Legionella pneumophila, Mycoplasma pneumoniae and Chlamydophila pneumoniae (previously, Chlamydia pneumoniae) which are known as bacterial atypical pneumonia agents. Serological tests are used commonly for laboratory diagnosis of these agents. The aim of this study was to evaluate the causative role of bacterial atypical pneumonia agents in clinically diagnosed pneumonia patients. Acute and convalescent serum samples were collected from a total of 65 clinically diagnosed adult pneumonia patients in order to evaluate IgM and IgG positivities against L. pneumophila, M. pneumoniae and C. pneumoniae. IgM and IgG were evaluated by enzyme immunoassay (ELISA) for L. pneumophila and M. pneumoniae, and by indirect fluorescent antibody (IFA) method for C. pneumoniae. In acute serum samples, 4 (6.2%) M. pneumoniae IgM positivity in addition to 3 (4.6%) L. pneumophila IgG, 3 (4.6%) M. pneumoniae IgG and 62 (95.4%) C. pneumoniae IgG positivity were detected. In convelescent serum samples, 3 (4.6%) L. pneumophila, 1 (1.5%) M. pneumoniae, 3 (4.6%) C. pneumoniae IgM positivity and 4 (6.2%) L. pneumophila with 1 (1.5%) M. pneumoniae IgG positivity were detected in addition to acute sample positivities. According to these serological data, totally 16 (24.6%) of the patients were infected by bacterial atypical pneumonia agents. These results show that bacterial atypical pneumonia agents are important etiological factors for community acquired pneumonia. PMID- 15293900 TI - [Epidemiological evaluation of a possible outbreak in and nearby Tokat province]. AB - Between the dates of May 4th-August 6th 2002, 46 cases were detected with abdominal pain nausea, vomiting, arthralgia/myalgia, headache, fever, diarrhea and rash, in the middle Blacksea and north inner Anatolia regions. Their laboratory findings yielded elevated levels of liver enzymes (AST, ALT, LDH), leucopenia and thrombocytopenia. As the infection was treated easily with tetracyclines, clinical diagnosis was considered to be rickettsiosis or ehrlichiosis. Serum and blood samples obtained from some of the patients were tested against Rickettsia, Ehrlichia, Leptospira and Coxiella, in the national and international laboratories. Samples from 19 patients were sent to National Reference Centre and WHO Collaborating Centre for Rickettsial Reference and Research Laboratory, France, and 7 of them were reported as acute Q fever while 8 of them were reported as passed Q fever (QF) cases. In May 2003, new cases with similar symptoms have been reported from the same regions, with different epidemiologic and serologic findings (tick exposure history was higher, response to tetracycline was lower, C. burnetii antibodies were negative), indicating a viral etiology. The samples of these patients have been sent to National Reference Centre and WHO Collaborating Centre for Arboviruses and Viral Heamorrhagic Fevers, France, and the initial reports were marked as Crimean Congo hemorrhagic fever virus (CCHFV). Then the serum samples of previous 26 patients which were stored in National Serum Bank have been retrospectively investigated for viral aetiology in the same center, and 17 of them have been found positive for CCHFV IgM antibodies. Four of these patients were diagnosed as acute QF in 2002, one was passed QF, 2 were negative for QF and 10 were patients not investigated for QF. As a result, the detection of the both infections together in the same area shows the essential need for further epidemiological investigations. PMID- 15293901 TI - [Six years evaluation of Clostridium difficile associated diarrhea]. AB - This study was aimed to detect the presence of Clostridium difficile toxin in the stool samples of patients with antibiotic-associated diarrhea or pseudomembranous colitis, and to relate its presence with the clinical findings of the patients. Between January 1997-April 2003, a total of 726 stool samples were investigated for C. difficile toxin A and/or B by enzyme immunoassay. Of them, 68 (9.4%) were found positive for C. difficile toxin (62 were toxin A, 6 were toxin B). C. difficile associated diarrhea were found to be related mostly with the use of beta-lactam/beta-lactamase inhibitor combinations (32/68), followed by aminoglycosides (12/68), and cephalosporins (8/68). The ages of the patients were between 1-86 years old (mean: 43.3 years), and 36 (52.9%) of them had an underlying conditions. Chronic obstructive pulmonary disease and chronic renal failure were the underlying disease in 18, malignancy in 11, and others (diabetes, hepatitis, transplantation, multiple sclerosis) in 7 of the patients. In conclusion, toxin detection and knowledge of the risk factors are the beneficial guidelines for the diagnosis of C. difficile associated diarrhea in the routine setting. PMID- 15293902 TI - [Herpes simplex virus type 1 in the samples of patients with clinically prediagnosed as herpetic keratitis or keratoconjunctivitis]. AB - The aim of this study was to investigate the presence of Herpes simplex virus type 1, in the specimens from patients clinically prediagnosed as herpetic keratitis or keratoconjunctivitis, by virus isolation and direct immunoperoxidase methods. The samples obtained from a total of 33 patients included ulcerated corneal epithelium scrapings and, swabs from ulcer, corneal epithelium, and lower conjunctivas. For virus isolation, both scraping and swab samples for each patient, were inoculated into monolayered Vero cell lines which have previously cultivated on 24-well plates, and examined for the presence of cytopathic effects typical for HSV-1. At the end of 5-days incubation period, the culture media were discarded, the cells were fixed and stained with peroxidase labeled specific HSV 1 antibodies (direct immunoperoxidase--DIP--method). Simultaneously, the smears were prepared from the samples which were sufficient in amount, and stained with DIP method. As a result, cytopathic effects on cell cultures were observed in 4 (12.1%) of the samples, of which 2 were also positive with DIP method. Following DIP staining of smears prepared from 15 samples, HSV-1 antigen positivity was detected in only 1 of the samples. This sample was negative in cell culture. In contrast, one of the 2 cell culture positive samples yielded negative result in smear examination, while the other could not be examined since the sample was not enough. In conclusion, HSV-1 has been shown as the etiologic agent in 3 (9.1%) of our patients clinically prediagnosed as herpetic keratitis or keratoconjunctivitis, and lesion scrapings were determined as the most appropriate samples for virus isolation. PMID- 15293903 TI - [TT virus and hepatitis G virus in different risk groups in Afyon]. AB - The main transmission route of TT virus (Transfusion-Transmitted Virus, TTV) and hepatitis G virus (HGV) is by parenteral route of blood and blood-products. Since they form the same risk group, some of TTV or HGV positive patients may also be infected by hepatitis B virus (HBV) and/or hepatitis C virus (HCV). In this study, the presence of TTV and HGV has been investigated by reverse transcriptase polymerase chain reaction (RT-PCR, GeneAmp 5700 Sequence Detection System, AB), in 40 hepatitis B, 30 hepatitis C and 5 hepatitis B and C co-infected patients, 50 HBV and HCV negative hemodialysis patients, and 50 randomly selected healthy blood donors. As a result, 37 (21.1%) TTV and 11 (6.3%) HGV positivity were detected, out of a total 175 cases. The positivity rates for TTV and HGV were found as 40% and 5% in HBV-positive, 23.3% and 20% in HCV-positive, 20% and 20% in HBV+HCV co-infected patients, 20% and 4% in hemodialysis patients, and 6% and 0% in healthy blood donors, respectively. In conclusion, as the positivity rates were not low for these viruses, their role on the hepatitis pathogenesis should be further investigated by detailed studies. PMID- 15293904 TI - [Varicella-zoster virus seroprevalence in children between 0-15 years old]. AB - The incidence of chicken pox infections in our country is not clear since it is not an obligatory reported disease, and there is not enough seroepidemiologic studies on this subject. The aim of the study was to determine the seroprevalence of infections caused by Varicella-Zoster virus (VZV), and to determine the relation of prevalence with some factors. For this purpose, 885 children ages between 0-15 years old, were investigated for the presence of VZV-IgG antibodies. Specific IgG antibodies were screened qualitatively by enzyme immunoassay (ELISA). As a result, it was found that after the declination of maternal antibodies, seropositivity rates were low up to the end of the first year, and then showed a gradual increase. The seropositivity rates were found 41.2% for 4-5 years old group, whereas it was 80% for 10-11 years old group and 85% for 13-15 years old group. There was no statistically important difference between seropositivity and sexes of children (p>0.05), but the seropositivity rates showed statistically important differences between increasing age and the number of siblings. In conclusion, as the majority of varicella infections occur in early childhood, the best way to prevent the circulation of wild type VZV, is the vaccination of children against chicken pox. PMID- 15293905 TI - [Comparison of manual and automated (MagNA Pure) nucleic acid isolation methods in molecular diagnosis of HIV infections]. AB - Rapid quantitative molecular methods are very important for the diagnosis of human immunodeficiency virus (HIV) infections, assessment of prognosis and follow up. The purpose of this study was to compare and evaluate the performances of conventional manual extraction method and automated MagNA Pure system, for the nucleic acid isolation step which is the first and most important step in molecular diagnosis of HIV infections. Plasma samples of 35 patients in which anti-HIV antibodies were found as positive by microparticule enzyme immunoassay and confirmed by immunoblotting method, were included in the study. The nucleic acids obtained simultaneously by manual isolation kit (Cobas Amplicor, HIV-1 Monitor Test, version 1.5, Roche Diagnostics) and automated system (MagNA Pure LC Total Nucleic Acid Isolation Kit, Roche Diagnostics), were amplified and detected in Cobas Amplicor (Roche Diagnostics) instrument. Twenty three of 35 samples (65.7%) were found to be positive, and 9 (25.7%) were negative by both of the methods. The agreement between the methods were detected as 91.4%, for qualitative results. Viral RNA copies detected by manual and MagNA Pure isolation methods were found between 76.0-7.590.000 (mean: 487.143) and 113.0-20.300.0000 (mean: 2.174.097) copies/ml, respectively. When both of the overall and individual results were evaluated, the number of RNA copies obtained with automatized system, were found higher than the manual method (p<0.05). Three samples which had low numbers of nucleic acids (113, 773, 857, respectively) with MagNA Pure, yielded negative results with manual method. In conclusion, the automatized MagNA Pure system was found to be a reliable, rapid and practical method for the isolation of HIV-RNA. PMID- 15293906 TI - [Comparison of indirect immunofluorescence and enzyme immunoassay methods for the determination of antinuclear antibodies]. AB - Antinuclear antibodies (ANA) are widely used for screening and monitoring of connective tissue diseases (CTD). Indirect immunofluorescence assay (IFA) is the standard method which is more often preferred for detecting these antibodies. Another method is enzyme immunoassay (ELISA) which includes extractable nuclear antigens (ENA). The aim of this study was to compare two different methods in view of their performances in the detection of ANA. A total of 27 sera from patients prediagnosed as different types of CTD, were screened by ANA-IFA (Zeus Scientific Inc, USA) and ELISA (Zeus Scientific Inc, ENA Profile-6, USA) methods. In addition, specific staining patterns of ANA on HEp-2 cells as a substrate, were enrolled with IFA. As a result, ANA positivity was detected in all of the 27 samples (100%) by IFA, and only in 7 (25.9%) by ELISA. The concordance rate between two different assays was estimated as 38.6%, and a statistically significant difference was found between the methods in the detection of ANA (x2=20, p<0.001). In conclusion, ANA-IFA method is still a reliable routine screening method for the laboratory diagnosis of CTD, while ENA Profile-6 ELISA may give false negative results, because of its limited antigen content, and should be supported with additional antigens. PMID- 15293907 TI - [Biofilm production and antifungal susceptibility patterns of Candida species]. AB - In this study, biofilm production and antifungal susceptibility of various Candida species were examined and compared. A total number of 156 Candida species (94 C. albicans, 21 C. tropicalis, 18 C. glabrata, 12 C. parapsilosis, 9 C. krusei, 1 C. guilliermondii and 1 C. kefyr) isolated from different clinical specimens were included in the study. The biofilm production of the strains was searched by modified tube adherence and microplate methods. Their antifungal susceptibilities against fluconazole and amphotericin B were determined by microdilution method, according to NCCLS M27-A2 standards. Forty three (27.6%) and 26 (16.7%) of the strains were found to be slime producing by tube adherence and microplate methods, respectively. The agreement between the two methods were detected as 65 percent. The rate of biofilm formation by different species ranged between 17% and 55% by tube adherence test and 0 and 48% by microplate method. No significant difference was found between the biofilm production of C. albicans and non-albicans species by tube adherence test (p=0.29). However; a statistically important difference was found when microplate method was considered (p=0.04). MIC50 and MIC90 values for fluconazole ranged between 4-64 microg/ml and 32-->64 microg/ml for different Candida species while these values changed between 0.25-1 microg/ml and 0.5-2 microg/ml for amphotericin B, respectively. Forty-five (28.8%) and 23 (14.7%) of the isolates were found to be dose dependent susceptible and resistant to fluconazole, respectively. Eleven (6.7%) of the strains had MIC values >1 microg/ml for amphotericin B. When the relation between the biofilm production and the susceptibility categories of the strains for amphotericin B were searched, no statistical differences were found by any of the two methods (p=0.12 and p=0.50). A statistically important difference (p=0.03) was determined by tube adherence test and no important difference (p=0.11) was detected by microplate method when fluconazole susceptibility categories were considered. As a conclusion, it has been determined that biofilm production which is a potential virulence factor for Candida species seems to be in agreement with the antifungal susceptibility categories of the strains especially for amphotericin B when the planktonic cells were used for the susceptibility testing. PMID- 15293908 TI - [Esterase activities of Candida species]. AB - The aim of this study was to determine the esterase activity of clinically important Candida species and to evaluate tween 80 opacity test medium as an additional identification method. A total of 118 Candida isolates (86 C. albicans, 12 C. tropicalis, 5 C. glabrata, 4 C. krusei, 3 C. guilliermondii, 3 C. kefyr, 3 C. parapsilosis, 1 C. famata, 1 C. dubliniensis) were examined for their response to tween 80 opacity test. Our results showed that, 83 of 86 (96.5%) Candida albicans, and all of the C. tropicalis and C. guilliermondii strains (100%) yielded a halo around their colonies. The remaining Candida species did not produce a positive test response after 13 days of incubation. It can be concluded that, tween 80 opacity test is a simple and economical test that can be used as a supplementary test for the identification of Candida species. PMID- 15293909 TI - [Comparison of the methenamine silver staining, direct fluorescent antibody and nested-polymerase chain reaction methods in the diagnosis of Pneumocystis carinii pneumonia]. AB - Pneumocystis carinii is one of the most common causative agents of pneumonia in immunocompromised patients, but the problems in the laboratory diagnosis of the disease frequently leads to diagnosis according to the response to medical treatment. In this study, the presence of P. carinii was investigated in immunocompromised patients who were presenting with the clinical symptoms of atypical pneumonia, by Gomori methenamine silver staining (GMS), direct fluorescent antibody (DFA) test and nested-polymerase chain reaction (nPCR) methods. Fifty-three samples of 49 patients were included in the study. Twelve of the samples (22.6%) were found to be positive by nPCR, 6 of them (11.3%) were found to be positive by DFA, while only one of them (1.8%) was positive by GMS staining method. As a result, for the appropriate treatment and prophylaxis of P. carinii infections, PCR which is a rapid and reliable diagnostic test should be used for diagnosis. PMID- 15293910 TI - [Characteristics of malaria cases diagnosed in Edirne province between 1994 2002]. AB - In this study, the epidemiological characteristics of malaria cases in Edirne province were investigated. Between the years of 1994-2002, a total of 317,087 blood samples were collected from soldiers in the province with selective active surveillance and from the resident population with active or passive surveillance methods, by the medical staff of Malaria Control Department and Health Centers, to search the presence of Plasmodium. In 281 of them Plasmodium spp. were detected, and the characteristics of malaria cases were investigated. Of the cases, 238 (84.7%) were detected in the first three years and mostly in September. While the indigenous cases were detected in the districts where rice planted intensely, the imported cases were detected in the districts heavily populated by military staff. Of the imported cases, 62% originated from Diyarbakir, Batman and Sanliurfa provinces (Southeast part of Turkey). P. vivax was detected as the causative agent in all blood samples except one P. ovale. This latter case has been the only one in Turkey so far and he was a student from Afghanistan. Attaching importance to fight off mosquitoes in intensely rice planted districts and strictly surveying the military staff, particularly from the region of Southern-East Anatolia, have led to successful control of the malaria cases in Edirne region. PMID- 15293911 TI - [Investigation of intestinal parasites in AIDS patients]. AB - In this study, enteric parasites were investigated in the stool samples of 38 AIDS patients (23 with chronic diarrhea and 15 without diarrhea) prospectively. At least three stool samples from each patient were investigated microscopically for ova or trophozoites. The samples were concentrated with formol-ether method and wet preparations stained with lugol were examined. In addition, the concentrated samples were stained with modified asid-fast (Kinyoun's), rhodamine auramine, modified trichrom and calcoflor methods. Enteric parasites were detected in 18 (47%) of the 38 patients, 16 patients harbored a single parasite, and 2 patients were found to be infected with more than one parasite. Only one (7%) of 15 AIDS patients without diarrhea, were found to be infected with Giardia lamblia. On the other hand, 17 (74%) of 23 AIDS patients with chronic diarrhea were found to be infected with various enteric parasites. Cryptosporidium spp. was detected in 9 (39%) of these 23 patients, and in 2 of them Microsporidium spp. accompanied Cryptosporidium. In 2 (9%) of these 23 patients G. lamblia were detected, while Isospora belli, Blastocystis hominis, Entamoeba histolytica, Strongyloides stercoralis and Trichuris trichiura were detected in one patient each. As a result, the detection rate of emerging parasites, including Cryptosporidium spp, Microsporidium spp, I. belli, B. hominis, and S. stercoralis was significantly higher than conventional parasites (39% versus 13%; z=2.34, p=0.01), and CD4 T cell counts were found to be significantly lower among AIDS patients with chronic diarrhea than those without diarrhea (x2=34.33, p<0.001). PMID- 15293912 TI - [Comparison of two different enzyme immunoassays in the diagnosis of Fasciola hepatica infections]. AB - In this study, the serum samples of 37 patients who were diagnosed as fascioliasis (=fasciolosis) by parasitologic, pathologic and/or radiographic methods, 51 patients who were previously infected with parasites other than Fasciola hepatica, and 31 healthy subjects, have been investigated for the presence of F. hepatica specific IgG antibodies with two different enzyme immunoassay (ELISA) methods. For this purpose, in-house DOT-ELISA and excretory secretory (ES)-ELISA techniques have been performed by using F. hepatica ES antigens. All of the 37 fasciolosis patients found to be seropositive with ES ELISA, while 36 were found positive with DOT-ELISA method. ES-ELISA method yielded 3 (5.9%) positive results for the patients with other parasitosis (2 toxocariasis, 1 hydatic cyst), and DOT-ELISA method gave positive results for 2 (3.9%) toxocariasis patients. In 31 healthy subjects, the seropositivity rates were detected as 3.2% and 16.1% for ES-ELISA and DOT-ELISA methods, respectively. As a result, the sensitivity and specificity of DOT-ELISA method were found as 97.3% and 93.3%, respectively, while these rates were 100% and 97.0% for ES ELISA, respectively. The highest positive predictive value was detected for ES ELISA (90.2%), while both of the methods showed high negative predictive values (100% for ES-ELISA, 99.1% for DOT-ELISA). In conclusion, ELISA methods would be useful especially for the laboratory diagnosis of fasciolosis, in the clinically suspected patients where the etiologic agent could not be identified. PMID- 15293913 TI - [Comparison of antibacterial effects of different antiseptics after hand washing]. AB - The aim of this study was to compare 4% chlorhexidine gluconate, 7.5% povidone iodine and liquid soap, which are used as hand washing solutions for the immediate, cumulative and residual effects in bacterial growth. For this purpose, 18 volunteers washed their hands with 7.5% povidone iodine, 4% chlorhexidine gluconate and liquid soap by applying standard hygienic hand washing technique. In order to find out the bacterial amount in the hands of study group, glove liquid test was used. For the evaluation of immediate effects, samples were collected just after the washing procedure, for the residual effects samples were collected from gloved hands after 3 hours, and for the cumulative effects, samples were collected after 5 days with daily hand washings. It is found that the immediate effects of 4% chlorhexidine gluconate was superior than others, and 7.5% povidone iodine was superior to liquid soap. There was no difference between 4% chlorhexidine gluconate and 7.5% povidone iodine for residual effects. The cumulative effects were observed for 4% chlorhexidine gluconate and 7.5% povidone iodine, while there was no cumulative effect for liquid soap. According to these results, it can be suggested that 4% chlorhexidine gluconate can be used as a reliable antiseptic agent in the hospitals, especially for laboratories, intensive care units and operating rooms, owing to its favourable immediate, residual and cumulative effects. PMID- 15293914 TI - [Case report: Otitis due to Vibrio alginolyticus]. AB - Infections caused by Vibrio alginolyticus is generally disregarded because of the fact that it is an unusual pathogen for humans. In this report, a 57 years old male patient with otitis media has been presented. Intense and pure isolation of V. alginolyticus was achieved from the ear discharge sample. According to the history, it was detected that he was on vacation at Mediterranean coast three weeks ago, thus the transmission of the bacteria is possibly via seawater. The isolate was found sensitive to all of the tested antibiotics, and the patient was treated successfully with the combination of ciprofloxacin and amoxicillin clavulanate. This case was presented to withdraw attention to V. alginolyticus infections which are very rarely seen, but should be taken into consideration in related cases. PMID- 15293915 TI - [Case report: A brucellosis case with subacute thyroiditis]. AB - Brucellosis is a systemic infectious disease, and multi-organ involvement is commonly seen. However, thyroid gland involment is a rare complication of brucellosis. In this report, a 34 years old male patient who had been followed up with the complaints of malaise, pain in front of the neck, throat pain, joint pains and fever, was presented. This case was diagnosed as brucella subacute thyroiditis by serological and histological findings. The patient was succesfully treated with rifampicin (300 mgr/day) plus doxycycline (200 mgr/day) therapy for eight weeks. The aim of this presentation was to discuss subacute thyroiditis, as a rare complication of brucellosis, and to withdraw the attentions of the clinicians on this subject. PMID- 15293916 TI - [Mycobacterium tuberculosis virulence factors and its immune evasion mechanisms]. AB - One-third of the world population is infected with Mycobacterium tuberculosis. When tuberculosis develops, the disease localization, severity, and outcome are highly variable between different individuals. The various manifestations of infection with M. tuberculosis reflect the balance between the bacilli and host defense mechanisms, in which the quality of host defense determines the outcome. Recent studies have identified several mycobacterial cell wall components that may be involved in the key steps of pathogenicity. M. tuberculosis is successful as a pathogen because of its ability to persist in an immunocompetent host. This bacterium lives within the macrophages. Hosts infected with M. tuberculosis mount a strong immune response. This response is usually sufficient to prevent progression to active disease. The strong immune response can control, but not eliminate the bacilli, indicating that M. tuberculosis has evolved mechanisms to modulate or avoid detection by the host immune response. Recent advances have improved our understanding of how M. tuberculosis evades two major antimicrobial mechanisms of macrophages; phagolysosome fusion and the production of toxic reactive nitrogen intermediates (RNI). In this review, the recent evidence of M. tuberculosis evasion from phagolysosome fusion and RNI toxicity, as well as prevention of the recognition of infected macrophages by CD4+ T lymphocytes by inhibiting MHC class II presentation, were discussed. PMID- 15293917 TI - [Little flaps in hand surgery]. AB - Cutaneous flaps, with spontaneous healing and skin grafts, are parts of the armaentarium of plastic surgeons. Many publications have emphasized the use of island flaps or distant flaps for soft tissue reconstruction of the hand. We will only focus on random flaps in this work. We will describe the different techniques and indications. PMID- 15293918 TI - [Should we divide Osborn's ligament during epicondylectomy and in situ decompression of the ulnar nerve?]. AB - INTRODUCTION: Two groups of patients with cubital tunnel syndrome were treated by neurolysis and medial epicondylectomy. In the first group, the operative procedure consisted solely of dividing Osborn's ligament and fascia but in the second group Osborn's ligament was reinserted after epicondylectomy to avoid dislocation of the nerve. The aim of this retrospective study was to compare the level of complete recovery after surgery and the frequency of dislocation of the nerve. MATERIAL AND METHOD: Group one: Nineteen patients, with a mean age of 47.7 (15-65), and 52% female, with the dominant hand involved in 63% cases, were treated. According to Mac Gowan's criteria, 32% of the elbows were classified preoperatively as grade I, 52% as grade II and 16% as grade III. Sensory nerve conduction velocity across the elbow was less than 40 m/s in 40% of cases. The mean duration of the disease was longer than 3 years in 16% of cases. Group two: Twenty three patients, with a mean age of 54.1 (33-75), and 56% female, with the dominant hand involved in 56% cases, were treated. According to Mac Gowan's criteria, three 17% of the elbows were classified preoperatively as grade I, 47% as grade II and 34% as grade III. Sensory nerve conduction velocity across the elbow was less than 40 m/s in 60% of cases. The mean duration of the disease was longer than 3 years in 4% of cases. Both groups were evaluated by a surgeon not involved in the treatment by clinical examination and DASH scoring. RESULTS: DASH scoring is correlated with functional recovery, grip strength and Mac Gowan preoperative scoring. In group one, (divided and reinserted ligament) with younger patients, half the incidence of Mac Gowan stage II and a shorter follow up, there were no dislocations, but less complete resolution of preoperative symptoms (68%/82%) and a higher DASH scoring (30.6/24.9). In group two (resected ligament), dislocation of the nerve was noted in 17% of cases. In both groups, pain at the epicondylectomy site was noted in 20% of cases. The chance of complete recovery was inversely related to the age (>50), and to the duration of the disease (>1 year). DISCUSSION: Surgical treatment of ulnar nerve entrapment at the elbow remains controversial. None of the presently advocated procedures (simple decompression of the ulnar nerve, medial epicondylectomy or transposition of the ulnar nerve) has proven optimal regarding long-term results. In both groups in this study, neurolysis of ulnar nerve by section of Osborn's ligament and fascia together with medial epicondylectomy proved to be an effective surgical procedure for treating grade I to II ulnar neuropathy. Section of Osborn's ligament without its reattachment is followed by more cases of complete recovery as well as more dislocation of the nerve although the latter elicited no subjective complaints from the patients. DASH scoring is effective in evaluating the recovery. PMID- 15293919 TI - [Use of a bilobed flap for the treatment of mucous cysts]. AB - INTRODUCTION: In the case of mucous cysts with attenuated skin, the authors suggest radical excision of the cyst together with the overlying skin. The skin defect is repaired with a bilobed flap whose donor site is left to heal by secondary intention. This surgical procedure also allows exploration of other areas of mucoid degeneration and repair of the proximal nail fold when necessary. METHOD: Twenty-six patients with an average age of 59 years (27 cysts), were operated with this procedure. Nail bed deformities were present in 55% of the cases. The cyst and the overlying skin were radically excised in conjunction with a dorsal capsulectomy; the use of the bilobed flap made the dissection easier, and flap translation allowed cover of the capsulectomy area and simultaneous repair of the nail fold in eight cases. RESULTS: Patients were reviewed with an average follow-up of 13.7 months. Seventy percent of the patients had no pain, and in 85% of the cases there was no loss of motion. Cosmetic appearance was satisfactory, and nail bed deformities disappeared or clearly subsided in 86% of the cases. One patient developed recurrence. DISCUSSION: Many surgical procedures have been described for mucous cysts treatment. This simple procedure allows radical excision of the cyst and the attenuated skin with low risk for the germinal matrix, precise location of cyst origin, repair of the nail fold and good skin cover in the capsulectomy area. PMID- 15293920 TI - [The protected tendinous graft in zone 2 or fibroblast trap]. AB - INTRODUCTION: We present a simple technique for preventing adhesions to tendon grafts in zone 2. A silicone tube is cut longitudinally and the third of its diameter is removed. It is then introduced under the pulley with the slot towards the skeleton. The tendon graft is introduced into this tube which will protect it from adhesions except at the slot level where fibroblasts create a new vasculature. MATERIALS AND METHODS: Between April 2002 and April 2003, we have used this technique in three patients, two in their thumbs and one in the long finger. In the Boyes classification, one thumb lesion corresponded to stage 1 and the other two cases to stage 4. The main pulleys were preserved. The tube measured 12 cm in length, and extended from the proximal juncture to the distal one (Pulvertaft technique). After 5 weeks of strict immobilization, the tube was withdrawn and the patients began active mobilization. RESULTS: After 8 weeks, comparison of the results with the opposite hand showed: Normal extension in all cases. Flexion deficit of 15 degrees in the thumb interphalangeal joints. Flexion deficit of 20 degrees in the proximal and distal interphalangeal joints of the long finger. There were no reported complications. With a mean follow-up of 1 year, these results were stable. CONCLUSION: These results support the contention that a good blood supply to the tendon and a good gliding surface have been developed with this technique. The tube seems to work as a fibroblast trap and blocks harmful adhesions. PMID- 15293921 TI - [Results of a five-year series of 44 trapeziectomies associated with ligamentoplasty and interposition arthroplasty]. AB - BACKGROUND: Surgical management of trapeziometacarpal joint osteoarthritis remains controversial. There have been few long term studies of trapeziectomy combined with ligamentoplasty and interposition arthroplasty (TLIA). Our results are based on a five year minimum follow-up study. METHODS: We carried out a study of 44 TLIA in 39 consecutive patients. A physical and radiological assessment was undertaken after on average of 6.9 years by a independent observer. RESULTS: A durable physical improvement was obtained in 18 cases in less than six months and in five cases after more than one year. Thereafter there was no secondary deterioration. A standard pain measurement gave an average result of 1.4 on a ten point scale. Pain was independent of displacement of the first metacarpal bone but had a tendency to be greater where associated with scaphotrapezoidal joint osteoarthritis. Strength was improved in 36 cases. The patients were satisfied and considered their grip to be normal in 41 cases. These variables did not change over time. DISCUSSION: TLIA give an excellent result in more than 90% of cases. This remains unchanged seven years after surgery. As opposed to prostheses, there is no secondary deterioration once healing is achieved. Algodystrophy is the main drawback. CONCLUSION: In our opinion, TLIA remains the best available surgical treatment of trapeziometacarpal joint osteoarthristis. PMID- 15293922 TI - [Soft-tissue chondroma of the hand. A new case]. AB - The authors report the case of a multilocular tumour of the soft tissues of the hand and wrist which was removed by excision biopsy and which proved to be a chondroma arising from the soft tissues. Soft tissue chondromas consist of islands of heterotopic cartilaginous tissue and can equally affect the viscera as well as the limbs. The hypothesis that microtrauma is involved in the aetiology of this condition has yet to find any factual support. Simple excision-biopsy should suffice to treat the condition but care should be taken to make the excision complete if recurrences are to be avoided. PMID- 15293923 TI - [Coronal fractures of scaphoid]. AB - Two cases of frontal fracture of the scaphoid proximal pole after a high energy trauma are reported. Diagnosis was delayed in both cases and was only possible with a CT-scan. One fracture was slightly displaced: the patient was treated conservatively and followed during 11 years with an excellent result. The other patient was operated on because of persistent pain and malunion. Only during surgery was the correct diagnosis made and due to scaphoid malunion, a proximal row carpectomy was performed. At a post operative assessment, in both cases, X rays showed a double contour of the proximal pole of the scaphoid. We believe than an arthroCT-scan or MRI is necessary to assess the fracture displacement and search for an associated ligamentous injury. PMID- 15293924 TI - [Work-related musculoskeletal disorders of the upper limbs: Presentation of the "Health Passport"]. PMID- 15293925 TI - American Journal of Health Promotion presents the recipient of the 2004 Robert F. Allen Symbol of H.O.P.E. Award. Harold P. Freeman, MD. PMID- 15293926 TI - Which population-based interventions would motivate smokers to think seriously about stopping smoking? AB - OBJECTIVE: To describe what smokers say about the impact of different population based interventions to motivate them to think seriously about stopping smoking. METHODS: A random-digit dialed cross-sectional telephone survey of adult current cigarette smokers was conducted in Erie and Niagara counties, New York, in October through November 2002. A total of 815 smokers were asked which of eight interventions would motivate them to think seriously about stopping smoking in the next 6 months. RESULTS: The offer of free nicotine patches/gum (53%) and cash incentives (49%) were the most frequently mentioned interventions that smokers said would get them to think seriously about stopping smoking. The degree of motivation to stop smoking was the most consistent and strongest predictor of how respondents answered the question about the influence of the various intervention options. CONCLUSION: Communities need to offer a wide array of interventions that are likely to appeal to different subgroups of smokers in order to have a population-wide impact on smoking behavior. PMID- 15293927 TI - Best practice in group-based smoking cessation: results of a literature review applying effectiveness, plausibility, and practicality criteria. AB - OBJECTIVE: Apply a "best practices" model to evidence regarding group smoking cessation to inform organizational decisions about adopting such programs. The best-practices model attempts to integrate rigorous review of evidence with context and practical considerations important to organizations contemplating adoption. DATA SOURCES: First, we identified effective practices by systematic literature review with two blinded reviewers to (1) search databases (99.8% agreement), (2) hand search journals with five or more papers selected in first step (99.9% agreement), (3) search reference lists of included papers (99.4% agreement), and (4) contact published experts. Second, model programs, theory, and expert opinion suggested plausible practices. Finally, a practitioner researcher advisory group suggested practical considerations affecting adoption decisions. STUDY SELECTION: All 67 studies included in the review met six requirements: (1) peer reviewed, (2) primary studies, (3) using experimental or quasi-experimental design, (4) compared one or more smoking-cessation interventions that involved two or more group sessions, (5) studied persons 18+ years old, and (6) reported > or =6-month point prevalence or continuous abstinence outcomes. DATA EXTRACTION: Two independent raters assessed study quality (89.5% agreement). Effective practices consistently exhibited a statistically significant effect. Plausible practices showed consistency across three types of evidence. An advisory group based practicality criteria on critical review and experience. DATA SYNTHESIS: Two practices were rated effective: multicomponent behavioral intervention and nicotine replacement therapy. Five practices received plausible ratings: components of behavioral skills, information about smoking, self-monitoring, social support, and four or more sessions of 60 to 90 minutes. The Advisory Group identified 11 practicality questions to assist organizations to make adoption decisions regarding effective and plausible practices. CONCLUSIONS: No research evidence guides potential smoking-cessation program adopters regarding program participants, providers, settings, or quality assurance. Reviews to influence practice must consider science and practice (context) to facilitate adoption of best practices. PMID- 15293928 TI - Effect of community coalition structure and preparation on the subsequent implementation of cancer control activities. AB - PURPOSE: The purpose of this paper is to identify factors within a three-phase community coalition development model that influence short-term success in cancer control coalitions based on the cumulative number of educational and screening activities conducted by the coalitions. DESIGN: This study was a nonrandomized community intervention trial involving four autonomous, independently funded multistate projects aimed at using coalitions to increase cancer screening and early detection. SETTING: The study was conducted in medically underserved, rural counties of Appalachia. SUBJECTS: Sixty-three coalitions involved 1725 members representing 71 counties within 10 states. Intervention. A network of local and regional community cancer control coalitions throughout rural Appalachia delivered culturally sensitive cancer control messages to women, with the long term goal of increasing the early detection of breast cancer ANALYSIS: County level coalitions were the unit of analysis for this study. Multiple linear regression was used to determine if three classes of variables corresponding to the developmental history of coalitions--formation, preparation for implementation, and implementation--were associated with the number of educational and screening activities conducted by the coalitions. RESULTS: The presence of a paid coordinator and formal organizational structure were both strongly associated with the number of activities conducted, accounting for 71% of the variability in coalition activities. Other variables positively associated with number of activities were the preparation of written community assessments and the modification of implementation plans. The same factors (structure, written plans) were associated with activities in coalitions without paid organizers (r2 = .57), as was the number of meetings. However, such coalitions produced an average of only 2.2 activities vs. 21.7 activities in coalitions with paid coordinators. CONCLUSION: The ALIC study would seem to indicate that community-based coalitions with paid coordinators and formal structures are capable of generating significantly higher levels of activity than those without either a paid coordinator or formal structure. PMID- 15293929 TI - Concepts guiding the study of the impact of the built environment on physical activity for older adults: a review of the literature. AB - OBJECTIVE: To identify theoretical models and key concepts used to predict the association between built environment and seniors' physical activity on the basis of a comprehensive review of the published literature. DATA SOURCE: Computer searches of Medline (1966-2002), PubMed (1966-2002), and Academic Search Elite (1966-2002) were conducted, and 27 English-language articles were found. Search terms included built environment, physical activity, exercise, walking, neighborhood, urban design, seniors, aging, aging in place, and physical environment. STUDY INCLUSION AND EXCLUSION CRITERIA: The primary inclusion criterion included the relation between the built environment and the physical activity among seniors living in neighborhoods. Studies assessing physical activity or overall health of a community-based population were included if underlying theoretical models and concepts were applicable to a senior population. Studies solely assessing social or psychosocial characteristics of place were excluded, as were review articles. DATA EXTRACTION: Extracted data included theoretical model, aspect of built environment studied, methods, and outcomes. DATA SYNTHESIS: Tables present key definitions and summarize information from empirical studies. RESULTS: Twenty-seven articles that focused on the environment-behavior relation in neighborhoods, six specific to seniors, were found. This area of research is in its infancy, and inconsistent findings reflect difficulties in measurement of the built environment. CONCLUSIONS: The relation between the built environment and the physical activity among seniors has been the subject of a limited number of studies. The choice of theoretical model drives the selection of concepts and variables considered. Safety, microscale urban design elements, aesthetics, and convenience of facilities are consistently studied across models. Few validated instruments have been developed and tested to measure neighborhood built environment. PMID- 15293930 TI - Measuring cues for healthy choices on restaurant menus: development and testing of a measurement instrument. AB - PURPOSE: To develop and test the Menu Checklist, an instrument to be used by community members to assess cues for healthy choices in restaurants. DESIGN: Menus from 14 restaurants were coded independently by two trained community reviewers to test the interrater reliability of the instrument. SETTING: A low income, urban, African-American community in Los Angeles, California. SUBJECTS: Restaurants were selected based on community perceptions of their potential to be included in a nutrition education and advocacy program to improve the availability of healthy foods. MEASURES: The Menu Checklist was adapted from previously tested measurement tools developed by the Prevention Research Center at Saint Louis University. Intraclass correlation coefficients (ICCs), kappa statistics, and percent agreements were calculated to assess interrater reliability. Descriptive statistics were calculated to show the availability of cues for healthy foods. RESULTS: The interrater reliability coefficients for the majority of items were high (.93-1.0). Labeling on restaurant menus was rare, as were low-fat choices. Fruits and vegetables were readily available: 31% of all entrees included one serving and 39% of all appetizers were primarily fruits and vegetables. CONCLUSIONS: The Menu Checklist is a reliable, low-cost means for community members to collect data on influences on food choices in restaurants. PMID- 15293931 TI - Expert opinions on "best practices" in worksite health promotion (WHP). AB - Best practices for worksite health promotion are highlighted in this edition of the Art of Health Promotion. Twelve academic experts and 12 practitioner experts were asked to rank almost 100 program management, behavior change, recruitment and participation, and ongoing communication strategies for use in worksite health promotion. Their similarities and differences are described along with their identification of newer, promising yet largely unproven strategies. These expert opinion-derived program strategies are helpful adjuncts to peer-review studies and articles. PMID- 15293932 TI - Have your medical staff bylaws reviewed by independent legal counsel. PMID- 15293933 TI - Despite advantages, perks fewer physicians work as employees. PMID- 15293934 TI - Plan now for retirement--MSMS symposium offers help. PMID- 15293935 TI - Disease management: the new frontier in community-based pain medicine. PMID- 15293936 TI - Help uninsured patients connect with needed programs. PMID- 15293937 TI - Dedicated doctor leads drive to push politicians. PMID- 15293938 TI - Grassroots action for the busy physician: how to effect change when you're short on time. PMID- 15293939 TI - A challenge to spouses: expand your 'comfort zone'. PMID- 15293940 TI - Meet your lawmakers. Interview by Jennifer Higgins. PMID- 15293941 TI - Leadership, business, politics: an alternative to victimhood. PMID- 15293942 TI - Two-triplet CGA repeats impede DNA replication in bacteriophage M13 in Escherichia coli. AB - Expansion and contraction instabilities associated with CAG, CGG, GAA and CGA (GAC) repeats propagation cause more than a dozen human genetic diseases and cancers. In this work, the propagation behavior of a bacteriophage M13 carrying a calf prochymosin cDNA fragment with a (CGA)2 repeat in a small hairpin forming region is reported. Such a M13 derivative when propagated in Escherichia coli, produces small plaques by decreasing phage yield and also mitigates the inhibition on host cell growth, compared to those control bacteriophages either containing a "CTGCTA" sequence or wildtype, suggesting that CGA2 repeat impedes DNA replication in vivo. Moreover, an increased internal free energy is found associated with (CGA)2 sequence compared to those "CTGCTA" and wildtype, which ruled out a possibility of CGA2 repeat effects on propagation is through influencing the hairpin structure formation. PMID- 15293943 TI - Additions to the hyphomycete genus Veronaea as phytoparasitic species. AB - Three new species of Veronaea, V. ficina on Ficus hispida L. (Moraceae), V. grewiicola on Grewia asiatica L. (Tiliaceae), and V. hippocratiae on Hippocratia arborea Willd. (Celastraceae), collected from forests of Nepal and the Terai belt of North-Eastern Uttar Pradesh, India, are described, illustrated and compared with related taxa. PMID- 15293944 TI - Assessment of toxigenic fungi on Argentinean medicinal herbs. AB - This work was performed to determine the incidence of toxigenic fungi and their mycotoxins on 152 dried medicinal and aromatic herbs, belonging to 56 species, which are used as raw material for drugs. International methodologies for fungal enumeration and identification were applied as well as TLC and HPLC techniques for toxins detection. The 52% out of 152 samples were contaminated with species from Aspergillus genus, 27% belonging to the Flavi section and 25% to the Circumdati section. The 16% of the total samples was contaminated with species from Fusarium genus. Aspergillus flavus and A. parasiticus (Flavi section), were the predominant species isolated, 50% out of 40 isolates were toxigenic. Aflatoxin concentrations ranged from 10 to 2000 ng/g. Only 26% of isolates from the Circumdati section (A. alliaceus, A. ochraceus and A. sclerotiorum) produced ochratoxin A in low concentrations between 0.12 and 9 ng/g. From a total of 29 strains of Fusarium spp., 27.5% were Fusarium verticillioides and F. proliferatum, which produced fumonisin Bland fumonisin B2 ranged from 20 to 22000 microg/g and from 5 to 3000 microg/g respectively. The remaining species, F. equiseti, F. oxysporum, F. semitectum, F. compactum, F. sombucinum and F. solani were able to produce neither group A and B trichothecenes nor zearalenone. The incidence of A. ochraceus and Fusarium spp. and their toxigenic capacities on medicinal herbs were studied for the first time in Argentina. It would be important to look for natural contamination to define acceptability Limits which allow the control of sanitary quality of medicinal herbs used as phytotherapic medicines in several countries. PMID- 15293945 TI - Effectivity of host-Rhizobium leguminosarum symbiosis in soils receiving sewage water containing heavy metals. AB - Disposal of sewage water in cultivated soils often containing considerable amount of potentially toxic metals such as Cu, Zn, Ni, Cd, Pb and Cr can be beneficial or harmful to plant growth, rhizobial survival, nodulation and nitrogen fixation. Soil samples from 14 such locations were collected. Symbiotic effectivity of host Rhizobium leguminosarum symbiosis in these soils was assessed. The total metal contents of Cd, Cu, Zn and Ni in all the 14 samples collected from farmer's fields receiving sewage water ranged between 1.3 and 6.7, 55.8-353.2, 356.0 1028.0 and 90.0-199.7 mg kg(-1) of soil, respectively. In Rohtak 1 soil, levels of Cd, Cu and Zn were highest while Ni was highest in Sonipat 2 soil. The content of available Cd, Cu, Zn and Ni in these soils ranged from 1.0-29.3; 6.2-47.0; 2.4 13.5, respectively, and was 2-9 percent of their total metal contents. All the N2 fixing parameters in pea and Egyptian clover were adversely affected by the presence of heavy metals. Available Cd and Cu contents significantly affected the N contents of pea and Egyptian clover plants, whereas Ni contents were negatively correlated with the plant biomass of pea and Egyptian clover. PMID- 15293946 TI - Bme585 I [5'-CCCGC(4/6)-3'], a new isoschizomer of restriction endonuclease Fau I, isolated from a strain of Bacillus mesentericus. AB - Bme585 I is a new member of the restriction endonuclease type IIS family. It was partially purified from the heterothrophic, mesophilic bacterial strain Bacillus mesentericus 585 by ammonium sulphate precipitation and phosphocellulose column chromatography. Bme585 I is a monomeric protein with a molecular mass of 62 kD. The enzyme is active over a broad pH range from 7.0 to 8.8, has a temperature optimum of 37 degrees C and tolerance of NaCl in reaction buffer from 0 to 400 mM. Bme585 I recognizes the asymmetric sequence 5'-CCCGC(4/6)-3' and is therefore an isoschizomer of restriction endonuclease Fau I. PMID- 15293947 TI - Laundry detergent compatibility of the alkaline protease from Bacillus cereus. AB - The endogenous protease activity in various commercially available laundry detergents of international companies was studied. The maximum protease activity was found at 50 degrees C in pH range 10.5-11.0 in all the tested laundry detergents. The endogenous protease activity in the tested detergents retained up to 70% on incubation at 40 degrees C for 1 h, whereas less than 30% activity was only found on incubation at 50 degrees C for 1 h. The alkaline protease from an alkalophilic strain of Bacillus cereus was studied for its compatibility in commercial detergents. The cell free fermented broth from shake flask culture of the organism showed maximum activity at pH 10.5 and 50 degrees C. The protease from B. cereus showed much higher residual activity (more than 80%) on incubation with laundry detergents at 50 degrees C for 1 h or longer. The protease enzyme from B. cereus was found to be superior over the endogenous proteases present in the tested commercial laundry detergents in comparison to the enzyme stability during the washing at higher temperature, e.g., 40-50 degrees C. PMID- 15293948 TI - Microbial diversity of soil from two hot springs in Uttaranchal Himalaya. AB - Soil samples collected from two hot springs, Soldhar and Ringigad, both located in the Garhwal region of Uttaranchal Himalaya were analysed for their physical, chemical and microbial components. The alkaline pH, total absence of carbon and nitrogen, and high temperature were features common to soil samples from both sites. The Soldhar samples contained higher amounts of Cu, Fe and Mn. Ringigad soil was devoid of Cu, but had much higher phosphate. While the optimum incubation temperature for isolating the maximum microbial counts from soil samples from the two sites was 50 degrees C, microbial growth in broth was also observed when incubated at 80 degrees C. Microscopic examination revealed three types of microbial populations, i.e., bacteria, yeast and filamentous organisms. The soil samples were found to be dominated by spore forming rods. Out of 58 aerobic isolates, 53 were gram positive bacilli. Gram positive anaerobic oval rods were also observed up to 60 degrees C. Soil dilution plates revealed the presence of antagonistic and phosphate solubilizing populations. PMID- 15293949 TI - Optimization of expression of an annexin V-hirudin chimeric protein in Escherichia coli. AB - A human Annexin V-Hirudin chimeric protein, Annexin V-Hirudin C, was expressed in Escherichia coli. A broad range of parameters such as plasmid stability during propogation and expression, expression capacity stability, the culture media, the growth time before induction and the induction duration were examined and optimized. Recombinant Annexin V-Hirudin C was purified from the cell lysate supernatants by ethanol precipitation, DEAE-cellulose chromatography and Sephadex G-75 chromatography, and the purified protein showed dose-dependent thrombin inhibitory activity. The overall production of purified Annexin V-Hirudin C protein is 10 mg/l/OD600. PMID- 15293950 TI - Extracellular proteases from eight psychrotolerant Antarctic strains. AB - Extracellular proteases from 8 Antarctic psychrotolerant Pseudomonas sp. strains were purified and characterised. All of them are neutral metalloproteases, have an apparent molecular mass of 45kDa, optimal activity at 40 degrees C and pH 7-9, retaining significant activity at pH 5-11. With the exception of P96-18, which is less stable, all retain more than 50% activity after 3 h of incubation at pH 5-9 and show low thermal stability (their half-life times range from 20 to 60 min at 40 degrees C and less than 5 min at 50 degrees C). These proteases can be used in commercial processes carried out at neutral pH and moderate temperatures, and are of special interest for their application in mixtures of enzymes where final thermal selective inactivation is needed. Results also highlight the relevance of Antarctic biotopes for the isolation of protease-producing enzymes active at low temperatures. PMID- 15293951 TI - Hypochlorite inactivation kinetics of Listeria monocytogenes in phosphate buffer. AB - The inactivation kinetics of Listeria monocytogenes in a phosphate buffer (PB) was determined at different hypochlorite concentrations, pH values and temperatures. D-values, using a linear regression, of L. monocytogenes in PB (pH 6.5) were 23.54, 17.40, 14.24 and 12.00s at 5, 10, 50 and 100 mg l(-1) hypochlorite, respectively, at 30 degrees C. The k-values ranged from 0.098 to 0.192s(-1) and 0.007 to 0.018s(-1) for hypochlorite concentrations (from 5 to 100 mg l(-1)) in PB (pH 6.5) and PB containing 0.1% peptone (pH 6.5), respectively, at 30 degrees C. D-values of L. monocytogenes exposed to hypochlorite were decreased with decreasing pH of PB (pH from 8.5 to 4.5). Hypochlorite showed higher antimicrobial activity at higher temperature. Not only the effect of hypochlorite concentration on the inactivation of L. monocytogenes but also other parameters like temperature, pH and suspending solutions effect the inactivation rates. PMID- 15293952 TI - [Health services economic evaluation: the investigation before decision making]. PMID- 15293953 TI - [Uniformity requirements for writing and editing biomedical papers. November 2003 actualization]. PMID- 15293954 TI - [New initiatives to improve medication safety in hospitals]. AB - Medication errors constitute a significant public health problem and are recognised as such nowadays among healthcare professionals, societies, authorities and international organizations. This has led to seeking and implementing effective practices focused on improving medication use safety. This article briefly describes some of the most recent initiatives promoted to prevent medication errors in the hospital setting. These safety improvement initiatives are based upon progressively developing an institutional culture of safety and on establishing practices designed to reduce errors or detect them in time, thus avoiding adverse effects to patients. Among these recent initiatives are the safety practices approved by the National Quality Forum, and the National Patient Safety Goals that the Joint Commission on Healthcare Accreditation has required since 2003. Also mentioned are several strategies that have been offered to facilitate the application of these practices, among which are the Pathways to Medication Safely, the development of collaborative projects among hospitals and organizations of experts, and the inclusion of a medication safety specialist in hospitals as a support figure overseeing the application of safety measures. Finally, the challenges inherent in putting these preventive measures into real patient's care are discussed. The barriers confronting this step must obviously be faced if improvements in patient safety are truly to be achieved. PMID- 15293955 TI - [Cost-effectiveness of a cervical cancer screening programme in the Algarve region, Portugal]. AB - BACKGROUND: Economic evaluation of health care is an instrument of support to decision-making in the allocation of resources between different options. The current study was conducted with a view to implement an organised mass-screening programme. The objective was to evaluate the cost-effectiveness of three options: two programmes to be implemented that are called "Pap screenings" and "Thin-prep screening", and the strategy currently in place called "Spontaneous screening". METHODS: The analysis was undertaken from the Health Care System perspective. The analytic horizon was 10 years. Direct medical costs were estimated and discounted at a rate of 5%. Effectiveness was estimated as number of preinvasive carcinomas detected and life years gained. The cost-effectiveness ratio was estimated for the three options and incremental cost-effectiveness was estimated by comparison of the options to be implemented with the current strategy. A sensitivity analysis was conducted on the key variables. RESULTS: The average cost per carcinoma detected was 1,199 euros with "Pap screening", 3,148 euros with "spontaneous screening" and 4,619 euros with "Thin-prep screening". The average cost per life year gained was 29 euros with "Pap screening", 77 euros with "Spontaneous screening" and 114 euros with "Thin-prep screening". "Pap screening" had an additional cost of 623 euros per additional carcinoma detected and 15 euros per additional life year gained. "Thin-prep screening" had an additional cost of 6,350 euros per additional carcinoma detected and 156 euros per additional life year gained. CONCLUSIONS: "Pap screening" had the best cost effectiveness relation and the lowest additional cost-effectiveness. PMID- 15293956 TI - [Functional gain and length of hospital stay at a medium-stay geriatric care unit at the Central Red Cross Hospital in Madrid, Spain]. AB - BACKGROUND: The medium-stay or convalescent care geriatric units were defined by the Spanish National Health Institute in 1996 as being the level of geriatric hospital care aimed at recovering those functions, activities or sequelae having undergone changes as a result of different prior processes. This study is aimed at evaluating the characteristics of patients related to functional gain and stay in medium-stay geriatric units. METHODS: A study was made of all those patients admitted throughout the May 2000-December 2001 period. The weekly and overall functional gain was evaluated using the Barthel Index (BI), the hospital stay and the effectiveness (BI at discharge-BI at admission/during stay) having been evaluated. An improvement in the weekly gain of BI>5 points was set at the effectiveness threshold. RESULTS: A total of 459 patients averaging age 80.56 (+/ 7.45) admitted for functional recovery from sequelae of ictus (48.4%), orthopedic disorders (26.3%) and immobility due to other ailments (23.5%) were evaluated. The total functional gain was 29.71 (+/-16.75) Barthel Index points, entailing an average stay of 24.93 (+/-12.94) days and a 1.44 (+/-1.02) effectiveness. The weekly functional gain was above the threshold set during the first three weeks, independently of the age and disorder for which admitted. In the multivariate regression analysis, the age, admission due to ictus, functional impairment prior to admission, cognitive impairment at admission, comorbility and delay in admission were related to a lesser functional gain. Admission due to ictus and a better functional condition prior to admission and better cognitive condition at admission were related to a longer stay. CONCLUSIONS: Hospital stays in medium stay geriatric units is adequate, at least during the first three weeks. A comparison of the results among units should be adjusted by age, the disorder for which admitted, comorbility and functional and cognitive condition of the patients. PMID- 15293957 TI - [Accessibility to healthcare services in the recent cervical cytology performed in an urban area in Colombia]. AB - BACKGROUND: Cervical cancer is a public health problem in developing countries. Cytology of the uterine cervix has been promoted to change this situation, the coverage thereof not being adequate for many different reasons. This study is aimed at delving into the relationship between the recent cervical cytology performed and that performed three years previous to the survey, insurance and access to healthcare services. METHODS: An analysis was made of 1,021 records of women within the 18-64 age range from a cross-sectional study conducted in an urban area of Bogota (Colombia). Research was conducted regarding the date of the last cervical cytology undergone, access to healthcare services, health insurance affiliation and socio-demographic variables. Once the data had been gathered, a descriptive analysis was conducted and a logic regression model was then constructed. RESULTS: A 97.8% response was achieved. A total of 38 of the 1,021 records were ruled out due to their being records of women having had hysterectomies. A total of 733 (69.7%) of the other 983 records reported a recent cervical cytology, which was related to being over 30 years of age, being from a middle-class socioeconomic stratum, not being single, having a major degree of schooling, health insurance affiliation and having a health institution to which to go for care when needed. CONCLUSIONS: Short-range and medium-range strategies aimed at increasing healthcare insurance and access to healthcare services are fundamental for increasing the coverage of performing cervical cytologies in developing countries. PMID- 15293958 TI - [The use of anxiolytic and hypnotic drugs in Spain (1995-2002)]. AB - BACKGROUND: Recent studies have revealed a rise in the use of anxiolytic and hypnotic drugs as well as the improper use thereof in Western countries. This study is aimed at ascertaining the pattern of use of anxiolytic and hypnotic drugs in Spain within the 1995-2002 period. METHODS: The data related to the use of medications was taken from the Ministry of Health and Consumer Affairs' ECOM (Medicinal Products Consumption) database, which includes information on the use of medications delivered through the community pharmacies and reimbursed by the National Health System. The data are expressed in Defined Daily Doses per 1,000 inhabitants per day. RESULTS: The use of anxiolytic and hypnotic drugs rose from 39.71 Defined Daily Doses per 1,000 inhabitants per day in 1995 to 62.02 in 2002. Throughout the period under study, benzodiazepines having a medium-range half life (8-24 h.) were those most used, especially lorazepam, alprazolam and lormetazepam. The active ingredient having shown the greatest drop in use was flunitrazepam. CONCLUSIONS: Although the use of anxiolytic and hypnotic drugs has undergone a considerable rise in recent years in Spain, the pattern of use has shown no major changes. PMID- 15293959 TI - [Emerging zoonoses linked to pets in the autonomous community of Madrid: design of a method for setting public health priorities, Spain]. AB - BACKGROUND: The emergence and re-emergence of infectious diseases are a major concern in Public Health. The unique socio-demographic aspects of the Autonomous Community of Madrid make it necessary for a greater deal of attention to be paid to pet-transmitted zoonoses. This study is aimed as heightening the knowledge of the emerging and re-emerging pet-transmitted zoonoses and the design of a method for prioritizing the same. METHODS: Based on the diseases obtained from official sources, laboratories and a review of published studies, a quantification and weighting method designed by the working group and adapted to the specificity of this study was applied. RESULTS: Based on the analysis of 137 diseases, 24 met the admission requirements. The weighting method is provided in table form, including eleven scoring criteria, the criteria categories and the coefficients employed. Salmonellosis was the top-ranked disease, followed by Q Fever, Tularemia and Hantavirus Infection. CONCLUSIONS: A method for specifically evaluating emerging and re-emerging diseases was designed, affording the possibility of setting priorities in the Public Health planning field. This study provides a listing of 24 zoonoses ranked in order of importance, based on which new strategy-related lines must be set out for the research and/or control thereof. PMID- 15293960 TI - [Labor satisfaction of dentists working for the Galicia health service]. PMID- 15293961 TI - Alzheimer's patient with broken hip languishes untreated for 8 hours. PMID- 15293962 TI - Nurse entitled to workers' comp. 18 years after leaving work: case on point: Graham v. Workers' Comp. Appeal Board, no. 320 CD 2003 A.2d -PA (2004). PMID- 15293963 TI - CA: male nurse sexually assaults male patient: was act performed "in course of employment?". PMID- 15293964 TI - PA: employer terminates nurse's comp.: nurse's attempt to revitalize her claim fails. PMID- 15293965 TI - Workers' comp. benefit payments start 7 years after injury case on point: Miller v. Christus St. Patrick Hospital, 2004 WL 840076 So.2d -LA. PMID- 15293966 TI - Spontaneous bacterial peritonitis. PMID- 15293967 TI - Pattern of transitional cell carcinoma of the urinary bladder as seen at Kenyatta National Hospital, Nairobi. AB - OBJECTIVE: To highlight the pattern of patients with transitional cell carcinoma of the urinary bladder with regards to age, sex, ethnic origin and histopathological classification. DESIGN: A ten year retrospective study. SETTING: Kenyatta National Hospital, Nairobi, Kenya. SUBJECTS: Fifty two patients who presented at Kenyatta National Hospital over the ten year period with histologically proven transitional cell carcinoma of the urinary bladder. There were 41 males and 11 females aged 27 to 84 years. The mean age was 57 years. RESULTS: The peak incidence was in the 60-69 years age group. The male to female ratio was 4:1. The regional (provincial) distribution revealed Central and Eastern had 77%, Rift valley had 6%, Nairobi, North Eastern, Western and Coast provinces had 2% each. In the ethnic distribution; Kikuyus, Kambas and Merus were 77% while others were 17.3%. Transitional cell carcinoma was found in 67% of the patients, 60% had advanced disease. Twenty nine percent were smokers while 25% consumed alcohol. The main occupation was farming in 65%. The most Common clinical presentations were haematuria 98% and lower abdominal pains in 71%. A total of 99,028 patients were admitted to the surgical wards,transitional cell carcinoma patients represented only 0.6%. CONCLUSION: Transitional cell carcinoma is a rare disease. At Kenyatta National Hospital it only represented 0.6% of all surgical admissions during the study period. It accounted for 67% of all bladder tumours an increase in incidence compared to previous studies. It is common in males more than females, with a peak in the seventh decade. Majority of the patients were from central Kenya. Alcohol, smoking and farming were the most important risk factors. Haematuria was the most important presenting clinical feature. Poor record keeping may have contributed to the low number of patients enrolled into the study. There is need for a thorough prospective study to find out the actual prevalence of bladder tumours. PMID- 15293968 TI - Kaposis sarcoma in a Nairobi hospital. AB - BACKGROUND: Kaposi's sarcoma (KS) is associated epidemiologically with HIV infection and a number of countries have reported a dramatic increase in the incidence of KS with the advent of AIDS. Although AIDS is prevalent in Kenya, no studies on the impact of AIDS on the pattern of KS has been carried out. OBJECTIVE: To determine any changes in the pattern of KS that might have occurred since the advent of AIDS in the country. DESIGN: Retrospective descriptive study. SETTING: Kenyatta National Hospital (KNH). METHOD: Pathology records of cases of KS diagnosed at KNH from 1968 to 1997 were analysed with respect to relative frequency, age, sex and site distribution; and trend. The period was divided into the pre and post AIDS era from 1983, which is the time the first AIDS patient was reported in the country. RESULT: A total of 1108 cases of KS consisting of 911 males and 197 females were recorded. The relative frequency of KS ranged between 2% to 5% of the total malignancies. There was a gradual decline in the male to female ratio from about 10:1 in the sixties to about 2:1 in 1997. There was no dramatic difference in the age distribution in the pre and post AIDS era, although a large number of cases were recorded as adults without age specification in the post AIDS era. Site distribution was characteristic of the disease with most of the cases having the lesions occurring in the lower limbs and involving the skin. CONCLUSION: Although these findings do not demonstrate a dramatic alteration in the pattern of KS in the post AIDS era there were indications that such changes may have been obscured by under-reporting. The fall in the male:female ratio is a strong indication of a rise in KS among female patients. A further study is necessary to elucidate the true impact of AIDS on the pattern of KS in the country. PMID- 15293969 TI - Missed opportunities and inappropriately given vaccines reduce immunisation coverage in facilities that serve slum areas of Nairobi. AB - OBJECTIVES: To quantify missed opportunities for immunisation, document reasons for their occurrence and evaluate the extent of inappropriately given vaccine doses. DESIGN: A cross sectional study of children under two years of age attending health facilities. SETTING: Six health facilities predominantly serving the slums of Nairobi. METHODOLOGY: Information on vaccination was extracted from child immunisation cards as well as from mothers or guardians of children. RESULTS: Effective immunisation coverage for Bacille-Callmette Guerin (BCG) was 91%. Coverage for the birth dose, first, second, and third doses of oral polio vaccine (OPV0, OPVI, OPV2, and OPV3) was 44%, 83%, 79% and 75% respectively. Effective coverage for first, second and third doses of diphtheria-pertusis tetanus (DPTI, DPT2 and DPT3) vaccine was 88%, 87% and 85% respectively. Measles coverage was 80%. Immunisation coverage for all antigens except OPV0 and OPV3 would have been increased to over 90% had missed immunisation opportunities and inappropriately administered vaccination been avoided. There would have been an 11% increase in OPV3 coverage to 86%. Increases in coverage for OPVI and OPV2 would have been 16% and 18% respectively. Coverage would have increased by 10% for diphtheria pertusistetanus (DPT) doses DPTI and DPT2, and 7% for DPT3. Measles immunisation coverage would have increased by 19% had missed immunisation opportunities and inappropriately administered vaccinations been avoided. The overall missed opportunities rate was 3%. The proportions of missed opportunities were higher for the OPV series than DPT series. CONCLUSION: Missed immunisation opportunities among clinic attendees in Nairobi occur and routine supervision should be strengthened in these health facilities in order to minimise such missed opportunities and inappropriately administered vaccines. PMID- 15293970 TI - Morphological study of the uncommon rectus sterni muscle in German cadavers. AB - OBJECTIVE: The present investigation has been designed to study the incidence of the rectus stern muscle in German human cadavers dissected in the Kingdom of Saudi Arabia, trying to find a postulation for the development of such muscle when present. DESIGN: Gross dissection of 130 cadavers, of both sexes, was performed throughout a period of 10 years. SETTING: Department of Anatomy College of Medicine, King Faisal University, Dammam, Saudi Arabia. INTERVENTION: Investigation of the origin and insertion of the rectus sterni and measurements of its length and width. RESULTS: Two adult cadavers, one of each sex, had shown well-developed bilateral rectus stern muscles. All muscles identified were parasternal in position, being superficial to the medial portion of the pectoralis major muscle. Minor morphological differences were observed among the four muscle masses concerning their length, breadth, origin and insertion. CONCLUSION: The current study has determined the incidence of the rectus sterni muscle, in German cadavers to be 1.54% per bodies examined compared to 4% in cadavers from Saudis. Such a frequency is compared to that reported in different geographic populations. The rectus sterni muscle is innervated by the anterior cutaneous branches of the intercostal nerves. The description of the rectus sterni muscle and its incidence determined in the present study, might be of a great help for clinicians radiographing or tackling the pectoral region. PMID- 15293971 TI - Comparative efficacy of albendazole and three brands of mebendazole in the treatment of ascariasis and trichuriasis. AB - OBJECTIVE: To evaluate the comparative efficacy of 400 mg albendazole (Smith Kline Beecham) as a single dose and three brands of mebendazole (Janssen, Unibios and East African Pharmaceuticals) at doses of 100 mg twice a day for three consecutive days in the treatment of single or mixed infections with Ascaris lumbricoides and or Trichuris trichiura in four treatment groups of school children. DESIGN: Randomized trial. SETTING: Wondo-Genet, southern Ethiopia. SUBJECTS: School children, aged six to nineteen years. RESULTS: The percentage cure rate and egg reduction rate obtained with albendazole and mebendazole from the three brands were not significantly different in the treatment of ascariasis. However, significant differences were found among the percentage cure rates and egg reduction rates of the four treatment groups in the treatment of trichuriasis. Comparatively, high cure rate (89.8%) and egg reduction rate (99.1%) were observed in vermox (Janssen) treated group followed by Unibios (India) treated group (53.3% and 96.53% cure and egg reduction rates, respectively), whereas low cure rate (17.1%) and egg reduction rate (69.8%) were seen in the albendazole treated group. CONCLUSION: The results of this study suggest that in areas of single or mixed infections with Trichuris trichiura and/or Ascaris lumbricoides are common public health problems and where laboratory facilities are not available to make parasite identification, mebendazole (particularly vermox, a product of Janssen laboratory) would be the drug of choice to treat trichuriasis and ascariasis. However, either mebendazole from the different brands or albendazole is effective in the treatment of ascariasis in areas where trichuriasis is not prevalent. PMID- 15293972 TI - Obstetric performance of women aged over forty years. AB - BACKGROUND: Advanced age and parity constitute two major factors in the outcome of pregnancy and labour management both in the developed and developing countries. OBJECTIVE: To examine pregnancy outcomes in women aged 40 years and above with the view of proffering solution to some of the problems encountered. DESIGN: A case control retrospective study. SETTING: Obafemi Awolowo University Teaching Hospital, Ile-Ife, Nigeria from 1st January, 1995 to 31st December, 1999. SUBJECTS: Three hundred and three women who delivered at 40 years of age or above. The control group comprised of 303 women who delivered between 20 and 29 years during the five years period. MAIN OUTCOME MEASURES: Gestational age at delivery, birth weight, mode and type of delivery, pregnancy and birth outcome. RESULTS: This showed a significant increase in prematurity, low birth weight, medical complications, operative deliveries (Caesarean section, vacuum and forceps), birth asphyxia and perinatal deaths all at P < 0.05. CONCLUSION: There is a poor pregnancy outcome at fourty years and above. Patients need to be counselled for care in a specialised centre. PMID- 15293973 TI - The first six month growth and illness of exclusively and non-exclusively breast fed infants in Nigeria. AB - OBJECTIVE: To compare the growth and illness pattern of infants who were exclusively breast fed for six months with those of infants commenced on complementary feeding before the age of six months and ascertain reasons for the early introduction of complementary feeding. DESIGN: A comparative prospective study. SETTING: Urban Comprehensive Health Centre (UCHC), Obafemi Awolowo University Teaching Hospitals Complex, Ile-Ife. SUBJECTS: Three hundred and fifty two mothers and their normal birth weight babies, weighing 2.500kg or more, and aged less than 14 days were serially recruited into the study. MAIN OUTCOME MEASURES: Mean/median monthly weights in the first six months of life, history/outpatient presentation for illnesses. RESULTS: Of the 352 mother-infant pairs recruited into the study, 345 (98%) were successfully followed up for the first six months of life. At six months, 264 (76.5%) were exclusively breast-fed, 45 (13.1%) were started on complementary feeding, between the ages of four and six months while 36 (10.4%) commenced complementary feeding before the age of four months. Infants who were exclusively breast-fed for six months had median weights above the 50th percentiles of the WHO/NCHS reference that is currently used in the national "road to health" (growth monitoring) cards. Furthermore, the mean weight of these babies at age six months was above those of babies who started complementary foods before six months. They also reported fewer symptoms and had fewer illness episodes (0.1 episodes per child) compared to those who started complementary feeding before six months. Infants who commenced complementary feeding before four months reported more symptoms and had more illness episodes (1.4 episodes per child) compared to those that commenced complementary feeding between four and six months (1.2 episodes per child). Common symptoms/illnesses seen or reported during the study among the groups were fever, diarrhoea and cough. Reasons given for early introduction of complementary foods include insufficient breast milk, thirst and convenience. CONCLUSION: It is concluded that exclusive breast-feeding supported adequate growth during the first six months of life for most of the infants studied. Early introduction of complementary foods did not provide any advantages in terms of weight gain in our environment, it was frequently associated with illness episodes and growth faltering. Many mothers however require support, encouragement and access to health care providers to breastfeed exclusively for the first six months of life. PMID- 15293974 TI - Comparison of calculated and direct low density lipoprotein cholesterol determinations in a routine laboratory. AB - BACKGROUND: Low density lipoprotein cholesterol (LDL-C) concentrations form the basis for treatment guidelines established for hyperlipidaemic patients. LDL-C concentrations are commonly calculated using the Friedwald formula (FF) which has several limitations. Recently, direct methods for LDL-C estimation have been developed which are suitable for routine laboratories. OBJECTIVE: To compare serum LDL-C concentrations determined by a direct assay and the Friedwald formula. DESIGN: Cross-sectional study. SETTING: Mater Hospital Laboratory, Nairobi, Kenya. METHODS: The clinical performance of the two methods was evaluated by analysing 211 fresh plasma samples from fasting adult patients. The samples were divided into four groups-normolipidaemic; and Types IIa, IIb and IV hyperlipidaemias. RESULTS: The Friedwald formula (FF) correlated best with the direct assay in the normolipidaemic samples (r = 0.879; y= 0.468 + 0.852x). Direct LDL-C values were significantly lower than the FF in the Type IIa hyperlipidaemia samples (paired differences 0.38 +/- 0.62). There was only 65% agreement between the two methods in the borderline high LDL-C group of the National Cholesterol Education Program (NCEP) classification (LDL-C 3.36-4.14 mmol/L). CONCLUSION: There is lack of agreement between the FF and the Abbott direct LDL-C assay. If the two methods are used interchangeably, there may be confusion in the classification and control of lipid lowering medication for patients with hyperlipidaemia. PMID- 15293975 TI - Salicylate poisoning in children: report of three cases. AB - To raise clinicians' awareness of chronic (therapeutic) salicylate poisoning as a common cause of admission in paediatric patients presenting to hospital with respiratory distress (a clinical manifestation of metabolic acidosis) and a history of 'over the counter' treatment with salicylate (Aspirin). We present two complex cases and provide a review of the literature on pathogenesis, clinical presentation and management of salicylate poisoning. A complete history of the illness, including questions on drug use, is vital in assessing the cause of metabolic acidosis in children. Due to the limited options available in managing such patients in many developing countries, emphasis should be placed on prevention of poisoning by educating the community and health care providers. PMID- 15293976 TI - Re: laparoscopic appendicectomy. PMID- 15293978 TI - Novel "second-generation" approaches for the control of type 2 diabetes. PMID- 15293979 TI - Diabetes mellitus: pathogenesis and treatment strategies. AB - This article introduces the disease state of diabetes mellitus and provides a background of the impact of the disease on the population, its biology and pathophysiology, and the current treatment strategies for treating diabetes. PMID- 15293980 TI - Peroxisome proliferator-activated receptor alpha/gamma dual agonists for the treatment of type 2 diabetes. PMID- 15293981 TI - Glucagon-like peptide-1: the basis of a new class of treatment for type 2 diabetes. PMID- 15293982 TI - Dipeptidyl peptidase IV inhibitors for the treatment of diabetes. PMID- 15293983 TI - Prospects for inhibitors of protein tyrosine phosphatase 1B as antidiabetic drugs. PMID- 15293984 TI - Structure-based design, synthesis, and evaluation of conformationally constrained mimetics of the second mitochondria-derived activator of caspase that target the X-linked inhibitor of apoptosis protein/caspase-9 interaction site. AB - A successful structure-based design of conformationally constrained second mitochondria-derived activator of caspase (Smac) mimetics that target the XIAP/caspase-9 interaction site is described. The most potent Smac mimetic 12d has a Ki of 350 nM for binding to the XIAP BIR3 domain protein. 12d is found to be effective in enhancing apoptosis induced by cisplatin in PC-3 human prostate cancer cells. PMID- 15293985 TI - Rational approaches to discovery of orally active and brain-penetrable quinazolinone inhibitors of poly(ADP-ribose)polymerase. AB - A novel class of quinazolinone derivatives as potent poly(ADP-ribose)polymerase-1 (PARP-1) inhibitors has been discovered. Key to success was application of a rational discovery strategy involving structure-based design, combinatorial chemistry, and classical SAR for improvement of potency and bioavailability. The new inhibitors were shown to bind to the nicotinamide-ribose binding site (NI site) and the adenosine-ribose binding site (AD site) of NAD+. PMID- 15293986 TI - Dopamine/serotonin receptor ligands. 9. Oxygen-containing midsized heterocyclic ring systems and nonrigidized analogues. A step toward dopamine D5 receptor selectivity. AB - Eleven-membered heterocycles (dibenz[g,j]-1-oxa-4-azacycloundecenes) and open chain analogues were synthesized and investigated for affinities to human dopamine receptor subtypes. The moderately rigidized rings displayed nanomolar and subnanomolar Ki values at D1-like receptors with a significant D1 to D2 and a slight D5 to D1 selectivity. The open-chain analogues showed lower affinities but significant D1 to D2 selectivities. Compound 3 (Ki(D5) = 0.57 nmol) showed antagonistic or inverse agonistic binding characteristics in a functional Ca assay. PMID- 15293987 TI - Bulky nonproteinogenic amino acids permit the design of very small and effective cationic antibacterial peptides. AB - The rate of multidrug-resistant infections is rapidly rising. Cationic antibacterial peptides are active against resistant pathogens and have low propensity for resistance development, but because of their unfavorable medicinal properties, cationic antibacterial peptides have been a limited clinical success. We have found that introduction of nongenetically coded amino acids and other lipophilic modifications opens the opportunity for development of extremely short and highly active antibacterial peptides with improved medicinal properties. PMID- 15293988 TI - A nonpeptidic sulfonamide inhibits the p53-mdm2 interaction and activates p53 dependent transcription in mdm2-overexpressing cells. AB - The evaluation of a sulfonamide inhibitor of the p53-mdm2 interaction is presented. The compound was identified using 3D database searching techniques based on a computationally derived pharmacophore model of mdm2 binding. It inhibits the physical interaction of recombinant p53 and mdm2 in vitro and increases p53-dependent transcription in an mdm2-overexpressing cell line. PMID- 15293989 TI - Human integrin alphavbeta5: homology modeling and ligand binding. AB - The recently reported crystal structures of the extracellular domains of the alphavbeta3 integrin in its unligated state and in complex with the peptide cyclo(-RGDf[NMe]V-) have dramatically increased our understanding of ligand binding to integrins. Nonetheless, ligand selectivity toward different integrin subtypes is still a challenging problem complicated by the fact that 3D structures of most of the integrin subtypes remain unknown. In this study, a three-dimensional model for the human alphavbeta5 integrin was obtained using homology modeling based on the crystal coordinates of alphavbeta3 in its bound conformation as template. The modeled receptor was refined using energy minimization and molecular dynamics simulations in explicit solvent. The refined alphavbeta5 model was used to explore the interactions between this integrin and alphavbeta3/alphavbeta5 dual and alphavbeta3-selective ligands in the attempt to provide a preliminary rationalization, at the molecular level, of ligand selectivity toward the two alphav integrins. It was found that, in the RGD binding site of the alphavbeta5 receptor, a partial "roof" composed mainly of the SDL residues Tyr179 and Lys180 is present and hampers the binding of compounds containing bulky substituents in the proximity of the carboxylate group. This study provides a testable hypothesis for alphav integrins subtype ligand binding selectivity, in line with both mutagenesis data and SARs studies. PMID- 15293990 TI - Combining pharmacophore search, automated docking, and molecular dynamics simulations as a novel strategy for flexible docking. Proof of concept: docking of arginine-glycine-aspartic acid-like compounds into the alphavbeta3 binding site. AB - A novel and highly efficient flexible docking approach is presented where the conformations (internal degrees of freedom) and orientations (external degrees of freedom) of the ligands are successively considered. This hybrid method takes advantage of the synergistic effects of structure-based and ligand-based drug design techniques. Preliminary antagonist-derived pharmacophore determination provides the postulated bioactive conformation. Subsequent docking of this pharmacophore to the receptor crystal structure results in a postulated pharmacophore/receptor binding mode. Pharmacophore-oriented docking of antagonists is subsequently achieved by matching ligand interacting groups with pharmacophore points. Molecular dynamics in water refines the proposed complexes. To validate the method, arginine-glycine-aspartic acid (RGD) containing peptides, pseudopeptides, and RGD-like antagonists were docked to the crystal structure of alphavbeta3 holoprotein and apoprotein. The proposed directed docking was found to be more accurate, faster, and less biased with respect to the protein structure (holo and apoprotein) than DOCK, Autodock, and FlexX docking methods. The successful docking of an antagonist recently cocrystallized with the receptor to both apo and holoprotein is particularly appealing. The results summarized in this report illustrated the efficiency of our light CoMFA/rigid body docking hybrid method. PMID- 15293991 TI - Validation of automated docking programs for docking and database screening against RNA drug targets. AB - The increasing awareness of the essential role of RNA in controlling viral replication and in bacterial protein synthesis emphasizes the potential of ribonucleoproteins as targets for developing new antibacterial and antiviral drugs. RNA forms well defined three-dimensional structures with clefts and binding pockets reminiscent of the active sites of proteins. Furthermore, it precedes proteins in the translation pathway; inhibiting the function of a single RNA molecule would result in inhibition of multiple proteins. Thus, small molecules that bind RNA specifically would combine the advantages of antisense and RNAi strategies with the much more favorable medicinal chemistry of small molecule therapeutics. The discovery of small-molecule inhibitors of RNA with attractive pharmacological potential would be facilitated if we had available effective computational tools of structure-based drug design. Here, we systematically test automated docking tools developed for proteins using existing three-dimensional structures of RNA-small molecule complexes. The results show that the native structures can generally be reproduced to within 2.5 angstroms more than 50-60% of the time. For more than half of the test complexes, the native ligand ranked among the top 10% compounds in a database-scoring test. Through this work, we provide parameters for the validated application of automated docking tools to the discovery of new inhibitors of RNA function. PMID- 15293992 TI - Tricyclic alkylamides as melatonin receptor ligands with antagonist or inverse agonist activity. AB - This work reports the design and synthesis of novel alkylamides, characterized by a dibenzo[a,d]cycloheptene nucleus, as melatonin (MLT) receptor ligands. The tricyclic scaffold was chosen on the basis of previous quantitative structure activity studies on MT1 and MT2 antagonists, relating selective MT2 antagonism to the presence of an aromatic substituent out of the plane of the MLT indole ring. Some dibenzo seven-membered structures were thus selected because of the noncoplanar arrangement of their benzene rings, and an alkylamide chain was introduced to fit the requirements for MLT receptor binding, namely, dibenzocycloheptenes with an acylaminoalkyl side chain at position 10 and dibenzoazepines with this side chain originating from the nitrogen atom bridging the two phenyl rings. Binding affinity at human cloned MT1 and MT2 receptors was measured by 2-[125I]iodomelatonin displacement assay and intrinsic activity by the GTPgammaS test. The majority of the compounds were characterized by higher affinity at the MT2 than at the MT1 receptor and by very low intrinsic activity values, thus confirming the importance of the noncoplanar arrangement of the two aromatic rings for selective MT2 antagonism. Dibenzocycloheptenes generally displayed higher MT1 and MT 2affinity than dibenzoazepines. N-(8-Methoxy-10,11 dihydro-5H-dibenzo[a,d]cyclohepten-10-ylmethyl)propionamide (4c) and -butyramide (4d) were the most selective MT2 receptor antagonists of the series, with MT2 receptor affinity comparable to that of melatonin and as such among the highest reported in the literature for MLT receptor antagonists. The acetamide derivative 4b produced a noticeable reduction of GTPgammaS binding at MT2 receptor, thus being among the few inverse agonists described. PMID- 15293993 TI - Liver-selective glucocorticoid antagonists: a novel treatment for type 2 diabetes. AB - Hepatic blockade of glucocorticoid receptors (GR) suppresses glucose production and thus decreases circulating glucose levels, but systemic glucocorticoid antagonism can produce adrenal insufficiency and other undesirable side effects. These hepatic and systemic responses might be dissected, leading to liver selective pharmacology, when a GR antagonist is linked to a bile acid in an appropriate manner. Bile acid conjugation can be accomplished with a minimal loss of binding affinity for GR. The resultant conjugates remain potent in cell-based functional assays. A novel in vivo assay has been developed to simultaneously evaluate both hepatic and systemic GR blockade; this assay has been used to optimize the nature and site of the linker functionality, as well as the choice of the GR antagonist and the bile acid. This optimization led to the identification of A-348441, which reduces glucose levels and improves lipid profiles in an animal model of diabetes. PMID- 15293994 TI - Solution structure of amyloid beta-peptide (25-35) in different media. AB - The design of molecules able to interact with the amyloid peptides either as inhibitors of fibril formation or as inhibitors of amyloid membrane pore formation represents one of the most relevant approaches in the development of anti-Alzheimer therapies. Abeta-(25-35), sequence GSNKGAIIGLM, is a highly toxic synthetic derivative of amyloid beta-peptides (Abeta-peptides), which forms fibrillary aggregates. Here, we report the NMR and CD investigation of Abeta-(25 35) in a membrane-mimicking environment and in isotropic mixtures of water and fluoro-alcohols to scan its conformational properties as a function of the medium. The analysis of the 3D structures in the mentioned conditions indicates a propensity of the peptide to behave as a typical transmembrane helix in the lipidic environment. In media characterized by different polarity, it loses the structural regularity at specific points of the sequence as a function of the environment. Furthermore, a comparison with the solution structure of full-length amyloid peptides suggests a role for the 25-27 kink region, which appears to be a general feature of all peptides under the solution conditions explored. PMID- 15293995 TI - Design, synthesis, and biological evaluation of new cyclic disulfide decapeptides that inhibit the binding of AP-1 to DNA. AB - The transcription factor activator protein-1 (AP-1) is an attractive target for the treatment of immunoinflammatory diseases, such as rheumatoid arthritis. Using the three-dimensional (3D) X-ray crystallographic structure of the DNA-bound basic region leucine zipper (bZIP) domains of AP-1, new cyclic disulfide decapeptides were designed and synthesized that demonstrated AP-1 inhibitory activities. The most potent inhibition was exhibited by Ac-c[Cys-Gly-Gln-Leu-Asp Leu-Ala-Asp-Gly-Cys]-NH2 (peptide 2) (IC50 = 8 microM), which was largely due to the side chains of residues 3-6 and 8 of the peptide, as shown by an alanine scan. To provide structural information about the biologically active conformation of peptide 2, the structures of peptide 2 derived from molecular dynamics simulation of the bZIP-peptide 2 complex with explicit water molecules were superimposed on the solution structures derived from NMR measurements of peptide 2 in water. These showed a strong structural similarity in the backbones of residues 3-7 and enabled the construction of a 3D pharmacophore model of AP-1 binding compounds, based on the chemical and structural features of the amino acid side chains of residues 3-7 in peptide 2. PMID- 15293997 TI - Three-dimensional quantitative structure-activity relationship analysis of a set of Plasmodium falciparum dihydrofolate reductase inhibitors using a pharmacophore generation approach. AB - A 3D pharmacophore model able to quantitatively predict inhibition constants was derived for a series of inhibitors of Plasmodium falciparum dihydrofolate reductase (PfDHFR), a validated target for antimalarial therapy. The data set included 52 inhibitors, with 23 of these comprising the training set and 29 an external test set. The activity range, expressed as Ki, of the training set molecules was from 0.3 to 11 300 nM. The 3D pharmacophore, generated with the HypoGen module of Catalyst 4.7, consisted of two hydrogen bond donors, one positive ionizable feature, one hydrophobic aliphatic feature, and one hydrophobic aromatic feature and provided a 3D-QSAR model with a correlation coefficient of 0.954. Importantly, the type and spatial location of the chemical features encoded in the pharmacophore were in full agreement with the key binding interactions of PfDHFR inhibitors as previously established by molecular modeling and crystallography of enzyme-inhibitor complexes. The model was validated using several techniques, namely, Fisher's randomization test using CatScramble, leave one-out test to ensure that the QSAR model is not strictly dependent on one particular compound of the training set, and activity prediction in an external test set of compounds. In addition, the pharmacophore was able to correctly classify as active and inactive the dihydrofolate reductase and aldose reductase inhibitors extracted from the MDDR database, respectively. This test was performed in order to challenge the predictive ability of the pharmacophore with two classes of inhibitors that target very different binding sites. Molecular diversity of the data sets was finally estimated by means of the Tanimoto approach. The results obtained provide confidence for the utility of the pharmacophore in the virtual screening of libraries and databases of compounds to discover novel PfDHFR inhibitors. PMID- 15293996 TI - Concise synthesis and structure-activity relationships of combretastatin A-4 analogues, 1-aroylindoles and 3-aroylindoles, as novel classes of potent antitubulin agents. AB - The synthesis and study of the structure-activity relationships of two new classes of synthetic antitubulin compounds based on 1-aroylindole and 3 aroylindole skeletons are described. Lead compounds 3, 10, and 14 displayed potent cytotoxicities with IC50 = 0.9-26 nM against human NUGC3 stomach, MKN45 stomach, MESSA uterine, A549 lung, and MCF-7 breast carcinoma cell lines. The inhibition of proliferation correlated with in vitro polymerization inhibitory activities. Structure-activity relationships revealed that 6-methoxy substitution of 3-aroylindoles and 5-methoxy substitution of 1-aroylindoles contribute to a significant extent for maximal activity by mimicking the para substitution of the methoxy group to the carbonyl group in the case of aminobenzophenones. Addition of a methyl group at the C-2 position on the indole ring exerts an increased potency. The 3,4,5-trimethoxybenzoyl moiety was necessary for better activity but not essential and can be replaced by 3,5-dimethoxybenzoyl and 3,4,5 trimethoxybenzyl moieties. We conclude that 1- and 3-aroylindoles constitute an interesting new class of antitubulin agents with the potential to be clinically developed for cancer treatment. PMID- 15293998 TI - 2-(1-adamantyl)-4-(thio)chromenone-6-carboxylic acids: potent reversible inhibitors of human steroid sulfatase. AB - Steroid sulfatase (STS) is an attractive target for the potential therapy of a number of estrogen- and androgen-dependent disorders. Most potent STS inhibitors known so far act as irreversible enzyme blockers and feature an aryl sulfamate moiety; even minor modifications at the sulfamate group result in drastically decreased activity. On the basis of a recently reported subclass of highly potent STS inhibitors, i.e., chromenone sulfamates, we now extended the investigation of structure-activity relationships to hitherto unstudied sulfamate replacements. Thereby, we discovered 2-(1-adamantyl)-4-(thio)chromenone-6-carboxylic acids (5d and 5j) as potent, reversible inhibitors of STS. In a cell-free system using purified human STS, both new inhibitors show similar Ki values (0.50 microM and 0.53 microM, respectively). However, the thio analogue 5j is superior to 5d (IC50 = 0.18 microM versus 9.4 microM) in a cellular assay system using CHO cells overexpressing STS. Compound 5j is an example of a reversible STS inhibitor with potent activity toward the target enzyme in a cellular test system. Moreover, 5d,j are stable and have no estrogenic potential. PMID- 15293999 TI - Phenoxyphenyl pyridines as novel state-dependent, high-potency sodium channel inhibitors. AB - In the search for more efficacious drugs to treat neuropathic pain states, a series of phenoxyphenyl pyridines was designed based on 4-(4 flurophenoxy)benzaldehyde semicarbazone. Through variation of the substituents on the pyridine ring, several potent state-dependent sodium channel inhibitors were identified. From these compounds, 23 dose dependently reversed tactile allodynia in the Chung model of neuropathic pain. Administered orally at 10 mg/kg the level of reversal was ca. 50%, comparable to the effect of carbamazepine administered orally at 100 mg/kg. PMID- 15294000 TI - Identification of structurally diverse growth hormone secretagogue agonists by virtual screening and structure-activity relationship analysis of 2 formylaminoacetamide derivatives. AB - Two molecules with known growth hormone secretagogue (GHS) agonist activity were used as templates to computationally screen approximately 80000 compounds. A total of 108 candidate compounds were selected, and five of them were found to be active in the low-micromolar range in both cell-based and direct binding assays. These compounds were structurally diverse and significantly differed from known GHS agonists. The most active compound was subjected to SAR evaluation, which slightly increased its potency and identified molecular regions important for specific GHS agonist activity. PMID- 15294001 TI - Piperazine derivatives of [1,2,4]triazolo[1,5-a][1,3,5]triazine as potent and selective adenosine A2a receptor antagonists. AB - The [1,2,4]triazolo[1,5-a]triazine derivative 3, more commonly known in the field of adenosine research as ZM-241385, has previously been demonstrated to be a potent and selective adenosine A2a receptor antagonist, although with limited oral bioavailability. This [1,2,4]triazolo[1,5-a]triazine core structure has now been improved by incorporating various piperazine derivatives. With some preliminary optimization, the A2a binding affinity of some of the best piperazine derivatives is almost as good as that of compound 3. The selectivity level over the adenosine A1 receptor subtype for some of the more active analogues is also fairly high, > 400-fold in some cases. Many compounds within this piperazine series of [1,2,4]triazolo[1,5-a]triazine have now been shown to have good oral bioavailability in the rat, with some as high as 89% (compound 35). More significantly, some piperazines derivatives of [1,2,4]triazolo[1,5-a]triazine also possessed good oral efficacy in rodent models of Parkinson's disease. For instance, compound 34 was orally active in the rat catalepsy model at 3 mg/kg. In the 6-hydroxydopamine-lesioned rat model, this compound was also quite effective, with a minimum effective dose of 3 mg/kg po. PMID- 15294002 TI - Fluorescent pirenzepine derivatives as potential bitopic ligands of the human M1 muscarinic receptor. AB - Following a recent description of fluorescence resonance energy transfer between enhanced green fluorescent protein (EGFP)-fused human muscarinic M1 receptors and Bodipy-labeled pirenzepine, we synthesized seven fluorescent derivatives of this antagonist in order to further characterize ligand-receptor interactions. These compounds carry Bodipy [558/568], Rhodamine Red-X [560/580], or Fluorolink Cy3 [550/570] fluorophores connected to pirenzepine through various linkers. All molecules reversibly bind with high affinity to M1 receptors (radioligand and energy transfer binding experiments) provided that the linker contains more than six atoms. The energy transfer efficiency exhibits modest variations among ligands, indicating that the distance separating EGFP from the fluorophores remains almost constant. This also supports the notion that the fluorophores may bind to the receptor protein. Kinetic analyses reveal that the dissociation of two Bodipy derivatives (10 or 12 atom long linkers) is sensitive to the presence of the allosteric modulator brucine, while that of all other molecules (15-24 atom long linkers) is not. The data favor the idea that these analogues might interact with both the acetylcholine and the brucine binding domains. PMID- 15294003 TI - Tetramethylcyclopropyl analogue of a leading antiepileptic drug, valproic acid. Synthesis and evaluation of anticonvulsant activity of its amide derivatives. AB - Although valproic acid (VPA) is an extensively used antiepileptic drug for treatment of various kinds of epilepsies, it has been proven to possess two life threatening side effects: hepatotoxicity and teratogenicity. Amide and urea derivatives of 2,2,3,3-tetramethylcyclopropanecarboxylic acid (TMCA) were prepared to discover lead compounds with clinical potential. In the amide and alkylamide series of TMCA derivatives, N-methoxy-2,2,3,3 tetramethylcyclopropanecarboxamide (21) was one of the most active compounds, having the subcutaneous metrazol test (scMet) ED50 values of 35 mg/kg in rats and 74 mg/kg in mice. In the maximal electroshock-induced seizure test (MES), this compound had ED50 values of 108 mg/kg in rats and 115 mg/kg in mice. Compound 21 was 18.5 and 4.5 times more potent than VPA in the corresponding rat tests. The most active compound in the series of urea derivatives was 2,2,3,3 tetramethylcyclopropanecarbonylurea (25), possessing MES ED50 values of 29 mg/kg in rats and 90 mg/kg in mice. In the scMet test this compound had ED50 values of 92 mg/kg in rats and 125 mg/kg in mice. The median toxic dose (TD50) in rats was 538 mg/kg, providing compound 25 with a wide safety margin and a protective index (TD50/ED50) of 18.5 in the MES test, which is about 12 times greater than that of VPA. Compounds 21 and 25 have the potential for development as novel potent and safe central nervous system active drugs with a broad spectrum of antiepileptic activity. PMID- 15294004 TI - Zinc ion dependent B-cell epitope, associated with primary Sjogren's syndrome, resides within the putative zinc finger domain of Ro60kD autoantigen: physical and immunologic properties. AB - The Ro/La ribonucleoprotein (RNP) complex is composed of the proteins Ro60kD, Ro52kD, and La48kD that are in association with one small cytoplasmic RNA (YRNA). Specific protein-RNA and protein-protein interactions are thought to occur through the RNP and zinc-finger secondary structure elements of the Ro60kD protein. The aim of our study was to investigate the antigenic properties of the zinc finger domain of the Ro60KD autoantigen and its contribution to the formation of Ro/La RNP complex. It was found that the peptide VSLVCEKLCNEKLLKKARIHPFHILIA (Zif-1), which corresponds to the natural sequence of the zinc finger domain (301-327), and the peptide C(Acm)NEKLLKKARIC(Acm), analogous to the intermediate loop 310-319 (Zif-3) of the same domain of Ro60KD, are recognized by the majority of anti-Ro/SSA and anti-La/SSB positive sera (82.6% and 77.1%, respectively) in the absence of zinc ions. The same sera failed to react with Zif-1 peptide in the presence of Zn2+. In contrast, the addition of zinc ions was necessary for the binding of Zif-1 to recombinant Ro52KD as shown by direct binding experiments of the recombinant protein with synthetic peptides. Our data suggest the zinc finger domain of Ro60kD contains a B-cell epitope with high specificity for primary Sjogren's syndrome. Furthermore, depending on the presence of zinc ions, the zinc finger domain of the Ro60KD protein can exist in two different conformational states favoring either an interaction with the Ro52KD protein or binding with autoantibodies. PMID- 15294005 TI - O-alkoxyamidine prodrugs of furamidine: in vitro transport and microsomal metabolism as indicators of in vivo efficacy in a mouse model of Trypanosoma brucei rhodesiense infection. AB - Five O-alkoxyamidine analogues of the prodrug 2,5-bis[4 methoxyamidinophenyl]furan were synthesized and evaluated against Trypanosoma brucei rhodesiense in the STIB900 mouse model by oral administration. The observed in vivo activity of these prodrugs demonstrates that compounds with an O methoxyamidine or O-ethoxyamidine group effectively cured all trypanosome infected mice, whereas prodrugs with larger side-chains did not completely cure the mice. Permeability across Caco-2 cell monolayers and microsomal metabolism were used to identify the underlying mechanisms of prodrug efficacy. PMID- 15294006 TI - Exercise immunology: the current state of man and mouse. AB - The mechanisms governing the body's response to physical exercise have been investigated from various perspectives including metabolism, nutrition, age and sex. Increased attention to the immune system during recent decades is reflected by a rapidly growing number of publications in the field. This article highlights the most recent findings and only briefly summarises more basic concepts already reviewed by others. Topics include Th1/Th2 cytokine balance, inoculation time, age and immune compensation. Some less investigated areas are discussed including studies in children, the environment and dendritic cells. Because physical exercise enhances some aspects and suppresses other aspects of immunity, the biological significance of alterations in the immune system are unknown. So far, no link between immunological alterations and infection rate has been established and infection after strenuous physical exercise is equally likely to be the result of exercising with an already established rather than a new infection. If there is an increased risk for infections with increased exercise duration and intensity, why do overtrained athletes not display the greatest risk for upper respiratory tract infections? Increased knowledge on immune system modulations with physical exercise is relevant both from a public health and elite athlete's point of view. PMID- 15294007 TI - Sex differences in respiratory exercise physiology. AB - Respiratory exercise physiology research has historically focused on male subjects. In the last 20 years, important physiological and functional differences have been noted between the male and female response to dynamic exercise where sex differences have been reported for most of the major determinants of exercise capacity. Female participation in competitive and recreational sport is growing worldwide and it is universally accepted that participation in regular physical activity is of health benefit for both sexes. Understanding sex differences is of potential importance to both the clinician scientist and the exercise physiologist since differences could impact upon exercise rehabilitation programmes for patient populations, exercise prescription for disease prevention in healthy individuals and training strategies for competitive athletes. Sex differences have been shown in resting pulmonary function, which may impact on the respiratory response to exercise. Women typically have smaller lung volumes and maximal expiratory flow rates even when corrected for height relative to men. Differences in resting and exercising ventilation across the menstrual cycle and relative to men have also been reported, although the functional significance remains unclear. Expiratory flow limitation and a high work of breathing are seen in women. Pulmonary system limitations, in particular exercise-induced arterial hypoxia, have been reported in both men and women; however, the prevalence in women is not yet known. From the available literature, it appears that there are sex differences in some areas of respiratory exercise physiology. However, detailed sex comparisons are difficult because the number of subjects studied to date has been woefully small. PMID- 15294008 TI - Public health and clinical recommendations for physical activity and physical fitness: special focus on overweight youth. AB - Numerous physical activity and physical fitness recommendations exist for youth. To date, however, no investigator has systematically reviewed these public health and clinical guidelines to determine whether the recommendations address overweight youth. This review examines youth-oriented physical activity and physical fitness recommendations for both the public health community and the clinical community, and assesses how overweight youth are specifically targeted by each of these two groups. Our review determined the extent to which the recommendations assessed four components of physical activity (i.e. frequency, intensity, duration and type) and four components of physical fitness (i.e. cardiorespiratory capacity, strength, flexibility and body composition). We further reviewed clinical recommendations to determine how they included two facets of the physician-patient encounter: assessment and counselling. After identifying all current physical activity and physical fitness recommendations for youth, we evaluated whether public health (n = 13) and clinical recommendations (n = 12) addressed physical activity and physical fitness for overweight youth. Findings revealed inconsistent, yet explicit, recommendations for the public health community where most organisations (12 of 13, 92%) included > or =3 physical activity components. In addition, organisations encouraged volumes of daily moderate- to vigorous-intensity physical activity for youth ranging from 30-60 or more minutes. Recommendations for the clinical community generally did not provide explicit physical activity and fitness recommendations to advise physicians on the assessment and counselling of patients and their families. Overweight youth were addressed within some recommendations (6 of 12, 50%) for the clinical community, but within few recommendations (2 of 13, 15%) for the public health community. To best inform public health and clinical communities, organisations developing future recommendations should include information fully documenting the decision-making processes used to develop the recommendations. In cases where mutual goals exist, public health and clinical communities should consider collaborating across agencies to develop joint recommendations. PMID- 15294010 TI - Incorporation of statistical uncertainty in health economic modelling studies using second-order Monte Carlo simulations. AB - Health economic modelling studies are of interest to many parties with different responsibilities and diverging interests. Therefore, it is obvious that recognising the relevance of statistical uncertainty and dealing with it appropriately are required to obtain unbiased results from health economic modelling studies, especially when those data are being used for reimbursement decisions. In this manuscript we explore the relevance of the incorporation of statistical uncertainty in a health economic model and identify various types of statistical uncertainty. The concepts were applied to a hypothetical Markov model for a hypothetical antiparkinsonian (AP) product. The method was based on the incorporation of probability distributions in the input variables using a second order Monte Carlo simulation and the definition of minimum relevant differences for clinical and economic input variables and outcomes. Our paper shows that the outcomes of a health economic model might be severely biased when statistical uncertainty is not taken into account, which justifies the need for the incorporation of statistical uncertainty in a health economic model. PMID- 15294011 TI - Assessing the use of retrospective databases in conducting economic evaluations of drugs: the case of asthma. AB - When evaluating drug substances, the traditional clinical study setting does not allow scope for observing real-life behaviour since all alternative actions are determined beforehand. However, a study based on prospective or retrospective databases containing real-life data can examine how patients and physicians behave in a real-world setting and can investigate the relationship between the introduction of a drug and the amount of healthcare used in actual practice. We reviewed the quality and potential policy application of published retrospective database studies in which an economic evaluation of the use of drugs in asthma was conducted. A search in literature databases found 16 such studies, which were reviewed and evaluated according to a published checklist. No article fulfilled all the criteria for a 'good' economic evaluation. The results of many of the evaluations may be informative, but not transparent enough to deliver policy conclusions. This may limit the use of the currently published retrospective database studies as a base for policy decision, compared with randomised controlled trials, despite the additional value of these database analyses when well conducted. A greater transparency when presenting material and results is therefore called for, to increase the usefulness of database studies. PMID- 15294013 TI - Cost effectiveness of fluticasone propionate plus salmeterol versus fluticasone propionate plus montelukast in the treatment of persistent asthma. AB - BACKGROUND: Asthma is a chronic disease, the two main components of which are inflammation and bronchoconstriction. Fluticasone propionate (FP) and salmeterol, a strategy that treats both main components of asthma, has been recently compared with FP plus montelukast in a randomised clinical trial. The present study reports economic evaluation of these two strategies. OBJECTIVE: To determine the relative cost effectiveness when persistent asthma is treated with FP/salmeterol 100/50 microg twice daily administered via a single Diskus inhaler device versus treatment with FP 100 microg twice daily via a Diskus inhaler plus oral montelukast 10mg once daily. STUDY DESIGN: A cost-effectiveness analysis was performed by applying cost unit data to resource utilisation data collected prospectively during a US randomised, double-blind, 12-week trial of FP/salmeterol (n = 222) versus FP + montelukast (n = 225). Patients were > or =15 years of age and were symptomatic despite inhaled corticosteroid (ICS) therapy. PATIENTS AND METHODS: Efficacy measurements in this analysis included improvement in forced expiratory volume in 1 second (FEV(1)) and symptom-free days. Direct costs included those related to study drugs, emergency room department visits, unscheduled physician visits, treatment of drug-related adverse events (oral candidiasis), and rescue medication (salbutamol [albuterol]). The study assumed a US third-party payer's perspective with costs in 2001 US dollars. RESULTS: Treatment with FP/salmeterol resulted in a significantly higher proportion (p < 0.001) of patients who achieved a > or =12% increase in FEV(1) than treatment with FP + montelukast (54% [95% CI 47%, 61%] vs 32% [95% CI 26%, 38%]). Lower daily costs and greater efficacy of FP/salmeterol resulted in a cost effectiveness ratio of US6.77 dollars (95% CI US5.99 dollars, US7.66 dollars) per successfully treated patient in the FP/salmeterol group compared with US14.59 dollars (95% CI US12.12 dollars, US17.77 dollars) for FP + montelukast. In addition, FP/salmeterol achieved similar efficacy in terms of symptom-free days compared with FP + montelukast (31% [95% CI 26%, 35%] vs 27% [95% CI 23%, 32%]), but at a significantly lower daily per-patient cost (US3.64 dollars [95% CI US3.60, US3.68 dollars] vs US4.64 dollars [95% CI US4.56 dollars, US4.73 dollars]). Sensitivity analyses demonstrated the stability of the results over a range of assumptions. CONCLUSION: From a US third-party payer's perspective, these findings suggest that treating the two main components of asthma (inflammation and bronchoconstriction) with FP/salmeterol may not only be a more cost-effective strategy but may actually lead to cost savings compared with the addition of montelukast to low-dose FP in patients with persistent asthma. The results were found to be robust over a range of assumptions. PMID- 15294012 TI - Economic aspects of severe sepsis: a review of intensive care unit costs, cost of illness and cost effectiveness of therapy. AB - Severe sepsis remains both an important clinical challenge and an economic burden in intensive care. An estimated 750,000 cases occur each year in the US alone (300 cases per 100,000 population). Lower numbers are estimated for most European countries (e.g. Germany and Austria: 54-116 cases per year per 100,000). Sepsis patients are generally treated in intensive care units (ICUs) where close supervision and intensive care treatment by a competent team with adequate equipment can be provided. Staffing costs represent from 40% to >60% of the total ICU budget. Because of the high proportion of fixed costs in ICU treatment, the total cost of ICU care is mainly dependent on the length of ICU stay (ICU-LOS). The average total cost per ICU day is estimated at approximately 1200 Euro for countries with a highly developed healthcare system (based on various studies conducted between 1989 and 2001 and converted at 2003 currency rates). Patients with infections and severe sepsis require a prolonged ICU-LOS, resulting in higher costs of treatment compared with other ICU patients. US cost-of-illness studies focusing on direct costs per sepsis patient have yielded estimates of 34,000 Euro, whereas European studies have given lower cost estimates, ranging from 23,000 Euro to 29,000 Euro. Direct costs, however, make up only about 20-30% of the cost of illness of severe sepsis. Indirect costs associated with severe sepsis account for 70-80% of costs and arise mainly from productivity losses due to mortality. Because of increasing healthcare cost pressures worldwide, economic issues have become important for the introduction of new innovations. This is evident when introducing new biotechnology products, such as drotrecogin-alpha (activated protein C), into specific therapy for severe sepsis. Data so far suggest that when drotrecogin-alpha treatment is targeted to those patients most likely to achieve the greatest benefit, the drug is cost effective by the standards of other well accepted life-saving interventions. PMID- 15294009 TI - The cardiovascular effects of chronic hypoestrogenism in amenorrhoeic athletes: a critical review. AB - In premenopausal women, the most severe menstrual dysfunction is amenorrhoea, which is associated with chronic hypoestrogenism. In postmenopausal women, hypoestrogenism is associated with a number of clinical sequelae related to cardiovascular health. A cardioprotective effect of endogenous oestrogen is widely supported, yet recent studies demonstrate a deleterious effect of hormone replacement therapy for cardiovascular health. What remain less clear are the implications of persistently low oestrogen levels in much younger amenorrhoeic athletes. The incidence of amenorrhoea among athletes is much greater than that observed among sedentary women. Recent data in amenorrhoeic athletes demonstrate impaired endothelial function, elevated low- and high-density lipoprotein levels, reduced circulating nitrates and nitrites, and increased susceptibility to lipid peroxidation. Predictive serum markers of cardiovascular health, such as homocysteine and C-reactive protein, have not yet been assessed in amenorrhoeic athletes, but are reportedly elevated in postmenopausal women. The independent and combined effects of chronic hypoestrogenism and exercise, together with subclinical dietary behaviours typically observed in amenorrhoeic athletes, warrants closer examination. Although no longitudinal studies exist, the altered vascular health outcomes reported in amenorrhoeic athletes are suggestive of increased risk for premature cardiovascular disease. Future research should focus on the presentation and progression of these adverse cardiovascular parameters in physically active women and athletes with hypoestrogenism to determine their effects on long-term health. PMID- 15294014 TI - The SARS coronavirus nucleocapsid protein induces actin reorganization and apoptosis in COS-1 cells in the absence of growth factors. AB - In March 2003, a novel coronavirus was isolated from patients exhibiting atypical pneumonia, and was subsequently proven to be the causative agent of the disease now referred to as SARS (severe acute respiratory syndrome). The complete genome of the SARS-CoV (SARS coronavirus) has since been sequenced. The SARS-CoV nucleocapsid (SARS-CoV N) protein shares little homology with other members of the coronavirus family. In the present paper, we show that SARS-CoV N is capable of inducing apoptosis of COS-1 monkey kidney cells in the absence of growth factors by down-regulating ERK (extracellular-signal-regulated kinase), up regulating JNK (c-Jun N-terminal kinase) and p38 MAPK (mitogen-activated protein kinase) pathways, and affecting their downstream effectors. SARS-CoV N expression also down-regulated phospho-Akt and Bcl-2 levels, and activated caspases 3 and 7. However, apoptosis was independent of the p53 and Fas signalling pathways. Furthermore, activation of the p38 MAPK pathway was found to induce actin reorganization in cells devoid of growth factors. At the cytoskeletal level, SARS CoV N down-regulated FAK (focal adhesion kinase) activity and also down-regulated fibronectin expression. This is the first report showing the ability of the N protein of SARS-CoV to induce apoptosis and actin reorganization in mammalian cells under stressed conditions. PMID- 15294015 TI - Studies of SpoIIAB mutant proteins elucidate the mechanisms that regulate the developmental transcription factor sigmaF in Bacillus subtilis. AB - SigmaF, the first compartment-specific sigma factor of sporulation, is regulated by an anti-sigma factor, SpoIIAB (AB) and its antagonist SpoIIAA (AA). AB can bind to sigmaF in the presence of ATP or to AA in the presence of ADP; in addition, AB can phosphorylate AA. The ability of AB to switch between its two binding partners regulates sigmaF. Early in sporulation, AA activates sigmaF by releasing it from its complex with AB. We have previously proposed a reaction scheme for the phosphorylation of AA by AB which accounts for AA's regulatory role. A crucial feature of this scheme is a conformational change in AB that accompanies its switch in binding partner. In the present study, we have studied three AB mutants, all of which have amino-acid replacements in the nucleotide binding region; AB-E104K (Glu104-->Lys) and AB-T49K (Thr49-->Lys) fail to activate sigmaF, and AB-R105A (Arg105-->Ala) activates it prematurely. We used techniques of enzymology, surface plasmon resonance and fluorescence spectroscopy to analyse the defects in each mutant. AB-E104K was deficient in binding to AA, AB-T49K was deficient in binding to ADP and AB-R105A bound ADP exceptionally strongly. Although the release of sigmaF from all three mutant proteins was impaired, and all three failed to undergo the wild-type conformational change when switching binding partners, the phenotypes of the mutant cells were best accounted for by the properties of the respective AB species in forming complexes with AA and ADP. The behaviour of the mutants enables us to propose convincing mechanisms for the regulation of sigmaF in wild-type bacteria. PMID- 15294016 TI - Octameric mitochondrial creatine kinase induces and stabilizes contact sites between the inner and outer membrane. AB - We have investigated the role of the protein ubiquitous mitochondrial creatine kinase (uMtCK) in the formation and stabilization of inner and outer membrane contact sites. Using liver mitochondria isolated from transgenic mice, which, unlike control animals, express uMtCK in the liver, we found that the enzyme was associated with the mitochondrial membranes and, in addition, was located in membrane-coated matrix inclusions. In mitochondria isolated from uMtCK transgenic mice, the number of contact sites increased 3-fold compared with that observed in control mitochondria. Furthermore, uMtCK-containing mitochondria were more resistant to detergent-induced lysis than wild-type mitochondria. We conclude that octameric uMtCK induces the formation of mitochondrial contact sites, leading to membrane cross-linking and to an increased stability of the mitochondrial membrane architecture. PMID- 15294017 TI - Targeting the A site RNA of the Escherichia coli ribosomal 30 S subunit by 2'-O methyl oligoribonucleotides: a quantitative equilibrium dialysis binding assay and differential effects of aminoglycoside antibiotics. AB - The bacterial ribosome comprises 30 S and 50 S ribonucleoprotein subunits, contains a number of binding sites for known antibiotics and is an attractive target for selection of novel antibacterial agents. On the 30 S subunit, for example, the A site (aminoacyl site) close to the 3'-end of 16 S rRNA is highly important in the decoding process. Binding by some aminoglycoside antibiotics to the A site leads to erroneous protein synthesis and is lethal for bacteria. We targeted the A site on purified 30 S ribosomal subunits from Escherichia coli with a set of overlapping, complementary OMe (2'-O-methyl) 10-mer oligoribonucleotides. An equilibrium dialysis technique was applied to measure dissociation constants of these oligonucleotides. We show that there is a single high-affinity region, spanning from A1493 to C1510 (Kd, 29-130 nM), flanked by two lower-affinity regions, within a span from U1485 to G1516 (Kd, 310-4300 nM). Unexpectedly, addition of the aminoglycoside antibiotic paromomycin (but not hygromycin B) caused a dose-dependent increase of up to 7.5-fold in the binding of the highest affinity 10-mer 1493 to 30 S subunits. Oligonucleotides containing residues complementary to A1492 and/or A1493 showed particularly marked stimulation of binding by paromomycin. The results are consistent with high resolution structures of antibiotic binding to the A site and with greater accessibility of residues of A1492 and A1493 upon paromomycin binding. 10-mer 1493 binding is thus a probe of the conformational switch to the 'closed' conformation triggered by paromomycin that is implicated in the discrimination by 30 S subunits of cognate from non-cognate tRNA and the translational misreading caused by paromomycin. Finally, we show that OMe oligonucleotides targeted to the A site are moderately good inhibitors of in vitro translation and that there is a limited correlation of inhibition activity with binding strength to the A site. PMID- 15294018 TI - A dual function fusion protein of Herpes simplex virus type 1 thymidine kinase and firefly luciferase for noninvasive in vivo imaging of gene therapy in malignant glioma. AB - BACKGROUND: Suicide gene therapy employing the prodrug activating system Herpes simplex virus type 1 thymidine kinase (HSV-TK)/ ganciclovir (GCV) has proven to be effective in killing experimental brain tumors. In contrast, glioma patients treated with HSV-TK/ GCV did not show significant treatment benefit, most likely due to insufficient transgene delivery to tumor cells. Therefore, this study aimed at developing a strategy for real-time noninvasive in vivo monitoring of the activity of a therapeutic gene in brain tumor cells. METHODS: The HSV-TK gene was fused to the firefly luciferase (Luc) gene and the fusion construct HSV-TK Luc was expressed in U87MG human malignant glioma cells. Nude mice with subcutaneous gliomas stably expressing HSV-TK-Luc were subjected to GCV treatment and tumor response to therapy was monitored in vivo by serial bioluminescence imaging. Bioluminescent signals over time were compared with tumor volumes determined by caliper. RESULTS: Transient and stable expression of the HSV-TK-Luc fusion protein in U87MG glioma cells demonstrated close correlation of both enzyme activities. Serial optical imaging of tumor bearing mice detected in all cases GCV induced death of tumor cells expressing the fusion protein and proved that bioluminescence can be reliably used for repetitive and noninvasive quantification of HSV-TK/ GCV mediated cell kill in vivo. CONCLUSION: This approach may represent a valuable tool for the in vivo evaluation of gene therapy strategies for treatment of malignant disease. PMID- 15294019 TI - Expression and importance of matrix metalloproteinase 2 and 9 (MMP-2 and -9) in human trophoblast invasion. AB - BACKGROUND: The aim of this study was to examine the invasiveness of first trimester trophoblasts according to the secretion profile of MMP-2 and -9 at different gestational stages, and to test the similarity between primary trophoblast cell-culture and the JAR choriocarcinoma cell-line. METHODS: First trimester trophoblasts were divided into two groups: 6-8 weeks (early) and 9-12 w (late) of gestation. The two trophoblast groups and JAR cells were cultured in medium, with various concentrations of forskolin and Epidermal Growth Factor (EGF). Proteolytic activity was detected by zymography and invasiveness was assessed by Matrigel invasion assay. Student's T-test was used for statistical analysis. RESULTS: In 6-8 w trophoblast, proMMP-2 was only slightly dominant over proMMP-9 (53.2% vs. 46.8% respectively), whereas in 9-12 w, proMMP-9 was clearly dominant over proMMP-2 (61.7% vs.38.3% respectively). In JAR cells proMMP-2 was strongly dominant (90.2% vs.9.8% respectively). In JAR cells forskolin significantly increased proMMP-2 and -9 secretion (128.5% +/- 12 and 183.2% +/- 27.9 of control, respectively). EGF had a dual effect on JAR cells: at 8 ng/ml both proMMP-2 and -9 were increased (133.5% +/-15 and 223.9% +/- 32.4 of control, respectively) while at 80 ng/ml both proMMP-2 and -9 were decreased (65.1% +/- 18.3 and 66.6% +/- 37 of control, respectively). Forskolin significantly increased both proMMP-2 and -9 secretion in 6-8 w and 9-12 w trophoblasts (125.9% +/- 6.3,128.4% +/- 6.4; 169.7% +/- 20.3, 120.3% +/- 4.5 of control, respectively). EGF also significantly increased both proMMP-2 and -9 secretion in 6-8 w and 9-12 w trophoblasts (141.22% +/- 14.8, 138.8% +/- 10.3; 168.3% +/- 18.2, 117.3 +/- 3.8 of control, respectively). Both forskolin and EGF increased trophoblast cells invasiveness in all groups. The invasive ability of trophoblast cells, induced by forskolin, was reduced by MMP-2 antibody in: JAR cells, 6-8 w and 9-12 w trophoblasts. Likewise trophoblast invasion induced by EGF was reduced by MMP-2 antibody in all groups. However the invasive ability induced by forskolin or EGF was inhibited by MMP-9 antibody only in trophoblasts from 9-12 w. CONCLUSIONS: First trimester trophoblasts express differential gelatinase secretion profile according to the gestational week. In JAR and early trophoblasts (6-8 w) MMP-2 is the main gelatinase and the key enzyme in trophoblast invasion. Thereafter in late first trimester trophoblasts (9-12 w), both MMP-2 and -9 participate in trophoblast invasion. PMID- 15294020 TI - Construction of a questionnaire measuring outpatients' opinion of quality of hospital consultation departments. AB - BACKGROUND: Few questionnaires on outpatients' satisfaction with hospital exist. All have been constructed without giving enough room for the patient's point of view in the validation procedure. The main objective was to develop, according to psychometric standards, a self-administered generic outpatient questionnaire exploring opinion on quality of hospital care. METHOD: First, a qualitative phase was conducted to generate items and identify domains using critical analysis incident technique and literature review. A list of easily comprehensible non redundant items was defined using Delphi technique and a pilot study on outpatients. This phase involved outpatients, patient association representatives and experts. The second step was a quantitative validation phase comprised a multicenter study in 3 hospitals, 10 departments and 1007 outpatients. It was designed to select items, identify dimensions, measure reliability, internal and concurrent validity. Patients were randomized according to the place of questionnaire completion (hospital v. home) (participation rate = 65%). Third, a mail-back study on 2 departments and 248 outpatients was conducted to replicate the validation (participation rate = 57%). RESULTS: A 27-item questionnaire comprising 4 subscales (appointment making, reception facilities, waiting time and consultation with the doctor). The factorial structure was satisfactory (loading >0.50 on each subscale for all items, except one item). Interscale correlations ranged from 0.42 to 0.59, Cronbach alpha coefficients ranged from 0.79 to 0.94. All Item-scale correlations were higher than 0.40. Test-retest intraclass coefficients ranged from 0.69 to 0.85. A unidimensional 9-item version was produced by selection of one third of the items within each subscale with the strongest loading on the principal component and the best item-scale correlation corrected for overlap. Factors related to satisfaction level independent from departments were age, previous consultations in the department and satisfaction with life. Completion at hospital immediately after consultation led to an overestimation of satisfaction. No satisfaction score differences existed between spontaneous respondents and patients responding after reminder(s). CONCLUSION: Good estimation of patient opinion on hospital consultation performance was obtained with these questionnaires. When comparing performances between departments or the same department over time scores need to be adjusted on 3 variables that influence satisfaction independently from department. Completion of the questionnaire at home is preferable to completion in the consultation facility and reminders are not necessary to produce non-biased data. PMID- 15294021 TI - The effect of Fucus vesiculosus, an edible brown seaweed, upon menstrual cycle length and hormonal status in three pre-menopausal women: a case report. AB - BACKGROUND: Rates of estrogen-dependent cancers are among the highest in Western countries and lower in the East. These variations may be attributable to differences in dietary exposures such as higher seaweed consumption among Asian populations. The edible brown kelp, Fucus vesiculosus (bladderwrack), as well as other brown kelp species, lower plasma cholesterol levels. Since cholesterol is a precursor to sex hormone biosynthesis, kelp consumption may alter circulating sex hormone levels and menstrual cycling patterns. In particular, dietary kelp may be beneficial to women with or at high risk for estrogen-dependent diseases. To test this, bladderwrack was administered to three pre-menopausal women with abnormal menstrual cycling patterns and/or menstrual-related disease histories. CASE PRESENTATION: Intake of bladderwrack was associated with significant increases in menstrual cycle lengths, ranging from an increase of 5.5 to 14 days. In addition, hormone measurements ascertained for one woman revealed significant anti estrogenic and progestagenic effects following kelp administration. Mean baseline 17beta-estradiol levels were reduced from 626 +/- 91 to 164 +/- 30 pg/ml (P = 0.04) following 700 mg/d, which decreased further to 92.5.0 +/- 3.5pg/ml (P = 0.03) with the 1.4 g/d dose. Mean baseline progesterone levels rose from 0.58 +/- 0.14 to 8.4 +/- 2.6 ng/ml with the 700 mg/d dose (P = 0.1), which increased further to 16.8 +/- 0.7 ng/ml with the 1.4 g/d dose (P = 0.002). CONCLUSIONS: These pilot data suggest that dietary bladderwrack may prolong the length of the menstrual cycle and exert anti-estrogenic effects in pre-menopausal women. Further, these studies also suggest that seaweed may be another important dietary component apart from soy that is responsible for the reduced risk of estrogen related cancers observed in Japanese populations. However, these studies will need to be performed in well-controlled clinical trials to confirm these preliminary findings. PMID- 15294022 TI - DLQI scores in vitiligo: reliability and validity of the Persian version. AB - BACKGROUND: The objective of this study was to translate and to test the reliability and validity of the 10-item Dermatology Life Quality Index (DLQI) questionnaire in Iranian patients with vitiligo. METHODS: Using a standard "forward-backward" translation procedure, the English language version of the questionnaire was translated into Persian (the Iranian official language) by two bilinguals. Seventy patients with vitiligo attending the Department of Dermatology, Saadi Hospital, Shiraz, Iran, were enrolled in this study. The reliability and internal consistency of the questionnaire were assessed by Cronbach's alpha coefficient and Spearman's correlation, respectively. Validity was performed using convergent validity. RESULTS: In all, seventy people entered into the study. The mean age of respondents was 28.3 (SD = 11.09) years. Scores on the DLQI ranged from 0 to 24 (mean +/- SD, 7.05 +/- 5.13). Reliability analysis showed satisfactory result (Cronbach's alpha coefficient = 0.77). There were no statistically significant differences between daily activity (DA) and personal relationship (PR) scale mean scores in generalized versus focal segmental involvement in sufferers (P = 0.056, P = 0.053, respectively). There were also strong differences between the mean scores of the PR (personal relationship) scale with the involvement of covered only and covered/uncovered areas (P= 0.016) that was statistically significant in the second group. CONCLUSIONS: The study findings showed that the Persian version of the DLQI questionnaire has a good structural characteristic and is a reliable and valid instrument that can be used for measuring the effects of vitiligo on quality of life. PMID- 15294023 TI - Estimating the beginning of the waterpipe epidemic in Syria. AB - BACKGROUND: Waterpipe smoking is becoming a global public health problem, especially in the Eastern Mediterranean region (EMR). METHODS: We try in this study, which is a cross sectional survey among a representative sample of waterpipe smokers in cafes/restaurants in Aleppo-Syria, to assess the time period for the beginning of this new smoking hype. We recruited 268 waterpipe smokers (161 men, 107 women; mean age +/- standard deviation (SD) 30.1 +/- 10.2, response rate 95.3%). Participants were divided into 4 birth cohorts (1980) and year of initiation of waterpipe smoking and daily cigarette smoking were plotted according to these birth cohorts. RESULTS: Data indicate that unlike initiation of cigarette smoking, which shows a clear age-related pattern, the nineties was the starting point for most of waterpipe smoking implicating this time period for the beginning of the waterpipe epidemic in Syria. CONCLUSION: The introduction of new flavored and aromatic waterpipe tobacco (Maassel), and the proliferation of satellite and electronic media during the nineties may have helped spread the new hype all over the Arab World. PMID- 15294024 TI - Hedgehog-interacting protein is highly expressed in endothelial cells but down regulated during angiogenesis and in several human tumors. AB - BACKGROUND: The Hedgehog (Hh) signaling pathway regulates a variety of developmental processes, including vasculogenesis, and can also induce the expression of pro-angiogenic factors in fibroblasts postnatally. Misregulation of the Hh pathway has been implicated in a variety of different types of cancer, including pancreatic and small-cell lung cancer. Recently a putative antagonist of the pathway, Hedgehog-interacting protein (HIP), was identified as a Hh binding protein that is also a target of Hh signaling. We sought to clarify possible roles for HIP in angiogenesis and cancer. METHODS: Inhibition of Hh signaling by HIP was assayed by measuring the induction of Ptc-1 mRNA in TM3 cells treated with conditioned medium containing Sonic hedgehog (Shh). Angiogenesis was assayed in vitro by EC tube formation on Matrigel. Expression of HIP mRNA was assayed in cells and tissues by Q-RT-PCR and Western blot. HIP expression in human tumors or mouse xenograft tumors compared to normal tissues was assayed by Q-RT-PCR or hybridization of RNA probes to a cancer profiling array. RESULTS: We show that Hedgehog-interacting protein (HIP) is abundantly expressed in vascular endothelial cells (EC) but at low or undetectable levels in other cell types. Expression of HIP in mouse epithelial cells attenuated their response to Shh, demonstrating that HIP can antagonize Hh signaling when expressed in the responding cell, and supporting the hypothesis that HIP blocks Hh signaling in EC. HIP expression was significantly reduced in tissues undergoing angiogenesis, including PC3 human prostate cancer and A549 human lung cancer xenograft tumors, as well as in EC undergoing tube formation on Matrigel. HIP expression was also decreased in several human tumors of the liver, lung, stomach, colon and rectum when compared to the corresponding normal tissue. CONCLUSION: These results suggest that reduced expression of HIP, a naturally occurring Hh pathway antagonist, in tumor neo-vasculature may contribute to increased Hh signaling within the tumor and possibly promote angiogenesis. PMID- 15294025 TI - Liver, spleen, pancreas and kidney involvement by human fascioliasis: imaging findings. AB - BACKGROUND: Fasciola hepatica primarily involves the liver, however in some exceptional situations other organs have been reported to be involved. The ectopic involvement is either a result of Parasite migration or perhaps eosinophilic reaction. CASE PRESENTATION: Here we report a known case of multiple myeloma who was under treatment with prednisolone and melphalan. He was infected by Fasciola hepatica, which involved many organs and the lesions were mistaken with metastatic ones. DISCUSSION: Presented here is a very unusual case of the disease, likely the first case involving the pancreas, spleen, and kidney, as well as the liver. PMID- 15294026 TI - A study comparing the actions of gabapentin and pregabalin on the electrophysiological properties of cultured DRG neurones from neonatal rats. AB - BACKGROUND: Gabapentin and pregabalin have wide-ranging therapeutic actions, and are structurally related to the inhibitory neurotransmitter GABA. Gabapentin, pregablin and GABA can all modulate voltage-activated Ca2+ channels. In this study we have used whole cell patch clamp recording and fura-2 Ca2+ imaging to characterise the actions of pregabalin on the electrophysiological properties of cultured dorsal root ganglion (DRG) neurones from neonatal rats. The aims of this study were to determine whether pregabalin and gabapentin had additive inhibitory effects on high voltage-activated Ca2+ channels, evaluate whether the actions of pregabalin were dependent on GABA receptors and characterise the actions of pregabalin on voltage-activated potassium currents. RESULTS: Pregabalin (25 nM - 2.5 microM) inhibited 20-30% of the high voltage-activated Ca2+ current in cultured DRG neurones. The residual Ca2+ current recorded in the presence of pregabalin was sensitive to the L-type Ca2+ channel modulator, Bay K8644. Saturating concentrations of gabapentin failed to have additive effects when applied with pregabalin, indicating that these two compounds act on the same type(s) of voltage-activated Ca2+ channels but the majority of Ca2+ current was resistant to both drugs. The continual application of GABA, the GABAB receptor antagonist CGP52432, or intracellular photorelease of GTP-gamma-S had no effect on pregabalin-induced inhibition of Ca2+ currents. Although clear inhibition of Ca2+ influx was produced by pregabalin in a population of small neurones, a significant population of larger neurones showed enhanced Ca2+ influx in response to pregabalin. The enhanced Ca2+ influx evoked by pregabalin was mimicked by partial block of K+ conductances with tetraethylammonium. Pregabalin produced biphasic effects on voltage-activated K+ currents, the inhibitory effect of pregabalin was prevented with apamin. The delayed enhancement of K+ currents was attenuated by pertussis toxin and by intracellular application of a (Rp)-analogue of cAMP. CONCLUSIONS: Pregabalin reduces excitatory properties of cultured DRG neurones by modulating voltage-activated Ca2+ and K+ channels. The pharmacological activity of pregabalin is similar but not identical to that of gabapentin. The actions of pregabalin may involve both extracellular and intracellular drug target sites and modulation of a variety of neuronal conductances, by direct interactions, and through intracellular signalling involving protein kinase A. PMID- 15294027 TI - Immobilized probe and glass surface chemistry as variables in microarray fabrication. AB - BACKGROUND: Global gene expression studies with microarrays can offer biological insights never before possible. However, the technology possesses many sources of technical variability that are an obstacle to obtaining high quality data sets. Since spotted microarrays offer design/content flexibility and potential cost savings over commercial systems, we have developed prehybridization quality control strategies for spotted cDNA and oligonucleotide arrays. These approaches utilize a third fluorescent dye (fluorescein) to monitor key fabrication variables, such as print/spot morphology, DNA retention, and background arising from probe redistributed during blocking. Here, our labeled cDNA array platform is used to study, 1) compression of array data using known input ratios of Arabidopsis in vitro transcripts and arrayed serial dilutions of homologous probes; 2) how curing time of in-house poly-L-lysine coated slides impacts probe retention capacity; and 3) the retention characteristics of 13 commercially available surfaces. RESULTS: When array element fluorescein intensity drops below 5,000 RFU/pixel, gene expression measurements become increasingly compressed, thereby validating this value as a prehybridization quality control threshold. We observe that the DNA retention capacity of in-house poly-L-lysine slides decreases rapidly over time (~50% reduction between 3 and 12 weeks post-coating; p < 0.0002) and that there are considerable differences in retention characteristics among commercially available poly-L-lysine and amino silane coated slides. CONCLUSIONS: High DNA retention rates are necessary for accurate gene expression measurements. Therefore, an understanding of the characteristics and optimization of protocols to an array surface are prerequisites to fabrication of high quality arrays. PMID- 15294028 TI - Design, implementation and evaluation of a practical pseudoknot folding algorithm based on thermodynamics. AB - BACKGROUND: The general problem of RNA secondary structure prediction under the widely used thermodynamic model is known to be NP-complete when the structures considered include arbitrary pseudoknots. For restricted classes of pseudoknots, several polynomial time algorithms have been designed, where the O(n6)time and O(n4) space algorithm by Rivas and Eddy is currently the best available program. RESULTS: We introduce the class of canonical simple recursive pseudoknots and present an algorithm that requires O(n4) time and O(n2) space to predict the energetically optimal structure of an RNA sequence, possible containing such pseudoknots. Evaluation against a large collection of known pseudoknotted structures shows the adequacy of the canonization approach and our algorithm. CONCLUSIONS: RNA pseudoknots of medium size can now be predicted reliably as well as efficiently by the new algorithm. PMID- 15294029 TI - Physical activity and health, novel concepts and new targets: report from the 12th Conference of the International Research Group on the Biochemistry of Exercise. AB - The present paper is the introductory paper to a series of brief reviews representing the proceedings of a recent conference on 'The biochemical basis for the health effects of exercise' organized by the International Research Group on the Biochemistry of Exercise in conjunction with the Nutrition Society. Here the aim is to briefly review and highlight the main innovations presented during this meeting. The following topics were covered during the meeting: exercise signalling pathways controlling fuel oxidation during and after exercise; the fatty acid transporters of skeletal muscle; mechanisms involved in exercise induced mitochondrial biogenesis in skeletal muscle; new methodologies and insights in the regulation of fat metabolism during exercise; muscle hypertrophy: the signals of insulin, amino acids and exercise; adipose tissue-liver-muscle interactions leading to insulin resistance. In these symposia state-of-the-art knowledge on how physical exercise exerts its effects on health was presented. The fast-growing number of identified pathways and processes involved in the health effects of physical exercise, which were discussed during the meeting, will help to develop tailored physical-activity regimens in the prevention of inactivity-induced deterioration of health. PMID- 15294030 TI - The biochemical basis of the health effects of exercise: an integrative view. AB - Physical inactivity-gene interactions result in changes in gene expression, leading to phenotypic changes in the skeletal muscle cell. A subpopulation of those genes that show changes in expression during physical inactivity are candidates for the environment-gene interactions that cross a threshold of biological significance such that overt clinical disease occurs. AMP kinase, GLUT4 and myosin heavy chain IIx are proposed as candidates for physical inactivity-modulated genes that have an altered function that may trigger a crossing of a threshold to disease. Future experiments will be needed to test the validity of the ideas presented. PMID- 15294031 TI - Regulation of glucose transport by the AMP-activated protein kinase. AB - The AMP-activated protein kinase (AMPK) is an energy-sensing enzyme that is activated during exercise and muscle contraction as a result of acute decreases in ATP:AMP and phosphocreatine:creatine. Physical exercise increases muscle glucose uptake, enhances insulin sensitivity and leads to fatty acid oxidation in muscle. An important issue in muscle biology is to understand whether AMPK plays a role in mediating these metabolic processes. AMPK has also been implicated in regulating gene transcription and, therefore, may function in some of the cellular adaptations to training exercise. Recent studies have shown that the magnitude of AMPK activation and associated metabolic responses are affected by factors such as glycogen content, exercise training and fibre type. There have also been conflicting reports as to whether AMPK activity is necessary for contraction-stimulated glucose transport. Thus, during the next several years considerably more research will be necessary in order to fully understand the role of AMPK in regulating glucose transport in skeletal muscle. PMID- 15294032 TI - Exercise signalling to glucose transport in skeletal muscle. AB - Contraction-induced glucose uptake in skeletal muscle is mediated by an insulin independent mechanism that leads to translocation of the GLUT4 glucose transporter to the muscle surface membrane from an intracellular storage site. Although the signalling events that increase glucose transport in response to muscle contraction are not fully elucidated, the aim of the present review is to briefly present the current understanding of the molecular signalling mechanisms involved. Glucose uptake may be regulated by Ca(2+)-sensitive contraction-related mechanisms, possibly involving Ca(2+)/calmodulin-dependent protein kinase II and some isoforms of protein kinase C. In addition, glucose transport may be regulated by mechanisms that reflect the metabolic status of the muscle, probably involving the 5'AMP-activated protein kinase. Furthermore, the p38 mitogen activated protein kinase may be involved in activating the GLUT4 translocated to the surface membrane. Nevertheless, the picture is incomplete, and fibre type differences also seem to be involved. PMID- 15294033 TI - Muscle glycogen and metabolic regulation. AB - Muscle glycogen is an important fuel for contracting skeletal muscle during prolonged strenuous exercise, and glycogen depletion has been implicated in muscle fatigue. It is also apparent that glycogen availability can exert important effects on a range of metabolic and cellular processes. These processes include carbohydrate, fat and protein metabolism during exercise, post-exercise glycogen resynthesis, excitation-contraction coupling, insulin action and gene transcription. For example, low muscle glycogen is associated with reduced muscle glycogenolysis, increased glucose and NEFA uptake and protein degradation, accelerated glycogen resynthesis, impaired excitation-contraction coupling, enhanced insulin action and potentiation of the exercise-induced increases in transcription of metabolic genes. Future studies should identify the mechanisms underlying, and the functional importance of, the association between glycogen availability and these processes. PMID- 15294034 TI - Transcriptional regulation of pyruvate dehydrogenase kinase 4 in skeletal muscle during and after exercise. AB - The pyruvate dehydrogenase complex (PDC) has a key position in skeletal muscle metabolism as it represents the entry of carbohydrate-derived fuel into the mitochondria for oxidation. PDC is regulated by a phosphorylation dephosphorylation cycle, in which the pyruvate dehydrogenase kinase (PDK) phosphorylates and inactivates the complex. PDK exists in four isoforms, of which the PDK4 isoform is predominantly expressed in skeletal and heart muscle. PDK4 transcription and PDK4 mRNA are markedly increased in human skeletal muscle during prolonged exercise and after both short-term high-intensity and prolonged low-intensity exercise. The exercise-induced transcriptional response of PDK4 is enhanced when muscle glycogen is lowered before the exercise, and intake of a low carbohydrate high-fat diet during recovery from exercise results in increased transcription and mRNA content of PDK4 when compared with intake of a high carbohydrate diet. The activity of pyruvate dehydrogenase (PDH) is increased during the first 2 h of low-intensity exercise, followed by a decrease towards resting levels, which is in line with the possibility that the increased PDK4 expressed influences the PDH activity already during prolonged exercise. PDK4 expression is also increased in response to fasting and a high-fat diet. Thus, increased PDK4 expression when carbohydrate availability is low seems to contribute to the sparing of carbohydrates by preventing carbohydrate oxidation. The impact of substrate availability on PDK4 expression during recovery from exercise also underlines the high metabolic priority given to replenishing muscle glycogen stores and re-establishing intracellular homeostasis after exercise. PMID- 15294035 TI - Exercise-induced mitogen-activated protein kinase signalling in skeletal muscle. AB - Exercise training improves glucose homeostasis through enhanced insulin sensitivity in skeletal muscle. Muscle contraction through physical exercise is a physiological stimulus that elicits multiple biochemical and biophysical responses and therefore requires an appropriate control network. Mitogen activated protein kinase (MAPK) signalling pathways constitute a network of phosphorylation cascades that link cellular stress to changes in transcriptional activity. MAPK cascades are divided into four major subfamilies, including extracellular signal-regulated kinases 1 and 2, p38 MAPK, c-Jun NH2-terminal kinase and extracellular signal-regulated kinase 5. The present review will present the current understanding of parallel MAPK signalling in human skeletal muscle in response to exercise and muscle contraction, with an emphasis on identifying potential signalling mechanisms responsible for changes in gene expression. PMID- 15294036 TI - Regulation of glycogen synthase activity and phosphorylation by exercise. AB - Glycogen synthase (GS) catalyses the rate-limiting step of UDP-glucose incorporation into glycogen. Exercise is a potent regulator of GS activity, leading to activation of GS immediately after exercise promoting glycogen repletion by mechanisms independent of insulin. The incorporation of UDP-glucose is energy demanding, and during intense exercise GS is deactivated, diminishing energy utilization but also increasing the potential for glycogen breakdown. An apparent activation of GS is observed during moderate exercise, which could be considered as a potential waste of energy, although the cellular capacity for glycogen breakdown is considerably higher than that for glycogen synthesis. The understanding of this complex regulation of GS activity in response to exercise is just at its beginning. In the present review potential mechanisms by which exercise regulates GS activity are described, factors that may promote GS activation and factors that may deactivate GS are discussed, pointing to the view that GS activity during exercise is the result of the relative strength of these opposing factors. PMID- 15294037 TI - Studies of plasma membrane fatty acid-binding protein and other lipid-binding proteins in human skeletal muscle. AB - The first putative fatty acid transporter identified was plasma membrane fatty acid-binding protein (FABPpm). Later it was demonstrated that this protein is identical to the mitochondrial isoform of the enzyme aspartate aminotransferase. In recent years data from several cell types have emerged, indicating that FABPpm plays a role in the transport of long-chain saturated and unsaturated fatty acids. In the limited number of studies in human skeletal muscle it has been demonstrated that dietary composition and exercise training can influence the content of FABPpm. Ingestion of a fat-rich diet induces an increase in FABPpm protein content in human skeletal muscle in contrast to the decrease seen during consumption of a carbohydrate-rich diet. A similar effect of a fat-rich diet is also observed for cytosolic fatty acid-binding protein and fatty acid translocase/CD36 protein expression. Exercise training up regulates FABPpm protein content in skeletal muscle, but only in male subjects; no significant differences were observed in muscle FABPpm content in a cross-sectional study of female volunteers of varying training status, even though muscle FABPpm content did not depend on gender in the untrained state. A higher utilization of plasma long-chain fatty acids during exercise in males compared with females could explain the gender-dependent influence of exercise training on FABPpm. The mechanisms involved in the regulation of the function and expression of FABPpm protein remain to be clarified. PMID- 15294038 TI - Regulation of fatty acid transport by fatty acid translocase/CD36. AB - Fatty acid (FA) translocase (FAT)/CD36 is a key protein involved in regulating the uptake of FA across the plasma membrane in heart and skeletal muscle. A null mutation of FAT/CD36 reduces FA uptake rates and metabolism, while its overexpression increases FA uptake rates and metabolism. FA uptake into the myocyte may be regulated (a) by altering the expression of FAT/CD36, thereby increasing the plasmalemmal content of this protein (i.e. streptozotocin-induced diabetes, chronic muscle stimulation), or (b) by relocating this protein to the plasma membrane, without altering its expression (i.e. obese Zucker rats). By repressing FAT/CD36 expression, and thereby lowering the plasmalemmal FAT/CD36 (i.e. leptin-treated animals), the rate of FA transport is reduced. Within minutes of beginning muscle contraction or being exposed to insulin FA transport is increased. This increase is a result of the contraction- and insulin-induced translocation of FAT/CD36 from an intracellular depot to the cell surface. Neither PPAR alpha nor PPAR gamma activation alter FAT/CD36 expression in muscle, despite the fact that PPAR alpha activation increases FAT/CD36 by 80% in liver. A novel observation is that FAT/CD36 also appears to be involved in mitochondrial FA oxidation, as this protein is located on the mitochondrial membrane and seems to be required to participate in moving FA across the mitochondrial membrane. Clearly, FAT/CD36 has an important role in FA homeostasis in skeletal muscle and the heart. PMID- 15294039 TI - Signalling components involved in contraction-inducible substrate uptake into cardiac myocytes. AB - Glucose and long-chain fatty acids (LCFA) are two major substrates used by heart and skeletal muscle to support contractile activity. In quiescent cardiac myocytes a substantial portion of the glucose transporter GLUT4 and the putative LCFA transporter fatty acid translocase (FAT)/CD36 are stored in intracellular compartments. Induction of cellular contraction by electrical stimulation results in enhanced uptake of both glucose and LCFA through translocation of GLUT4 and FAT/CD36 respectively to the sarcolemma. The involvement of protein kinase A, AMP activated protein kinase (AMPK), protein kinase C (PKC) isoforms and the extracellular signal-regulated kinases was evaluated in cardiac myocytes as candidate signalling enzymes involved in recruiting these transporters in response to contraction. The collected evidence excluded the involvement of PKA and implicated an important role for AMPK and for one (or more) PKC isoform(s) in contraction-induced translocation of both GLUT4 and FAT/CD36. The unravelling of further components along this contraction pathway can provide valuable information on the coordinated regulation of the uptake of glucose and of LCFA by an increase in mechanical activity of heart and skeletal muscle. PMID- 15294040 TI - New concepts of cellular fatty acid uptake: role of fatty acid transport proteins and of caveolae. AB - Efficient uptake and channelling of long-chain fatty acids (LCFA) are critical cell functions. Evidence is emerging that proteins are important mediators of LCFA-trafficking into cells and various proteins have been suggested to be involved in this process. Amongst these proteins is a family of membrane associated proteins termed fatty acid transport proteins (FATP). So far six members of this family, designated FATP 1-6, have been characterized. FATP 1, 2 and 6 show a highly-conserved AMP-binding region that participates in the activation of very-long-chain fatty acids (VLCFA) to form their acyl-CoA derivatives. The mechanisms by which FATP mediate LCFA uptake are not well understood, but several studies provide evidence that uptake of LCFA across cellular membranes is closely linked to acyl-CoA synthetase activity. It is proposed that FATP indirectly enhance LCFA uptake by activating VLCFA to their CoA esters, which are required to maintain the typical structure of lipid rafts in cellular membranes. Recent work has shown that the structural integrity of lipid rafts is essential for cellular LCFA uptake. This effect might be exerted by proteins, e.g. caveolin-1 and FAT/CD36, that use lipid rafts as platforms and bind or transport LCFA. The proposed molecular mechanisms await further experimental investigation. PMID- 15294041 TI - The metabolic role of IL-6 produced during exercise: is IL-6 an exercise factor? AB - For most of the last century, researchers have searched for a muscle contraction induced factor that mediates some of the exercise effects in other tissues such as the liver and the adipose tissue. It has been called the 'work stimulus', the 'work factor' or the 'exercise factor'. In the search for such a factor, a cytokine, IL-6, was found to be produced by contracting muscles and released into the blood. It has been demonstrated that IL-6 has many biological roles such as: (1) induction of lipolysis; (2) suppression of TNF production; (3) stimulation of cortisol production. The IL-6 gene is rapidly activated during exercise, and the activation of this gene is further enhanced when muscle glycogen content is low. In addition, carbohydrate supplementation during exercise has been shown to inhibit the release of IL-6 from contracting muscle. Thus, it is suggested that muscle-derived IL-6 fulfils the criteria of an exercise factor and that such classes of cytokines could be termed 'myokines'. PMID- 15294042 TI - Involvement of PPAR gamma co-activator-1, nuclear respiratory factors 1 and 2, and PPAR alpha in the adaptive response to endurance exercise. AB - Endurance exercise training induces an increase in the respiratory capacity of muscle, resulting in an increased capacity to generate ATP as well as improved efficiency of muscle contraction. Such adaptations are largely the result of a coordinated genetic response that increases mitochondrial proteins, fatty acid oxidation enzymes and the exercise- and insulin-stimulated glucose transporter GLUT4, and shifts the contractile and regulatory proteins to their more efficient isoforms. In recent years a number of the transcriptional regulators involved in this genetic response have been identified and these factors can be classified into two different groups. The first group comprises transcription factors such as nuclear respiratory factors (NRF) 1 and 2 and PPAR alpha that bind DNA in a sequence-specific manner. The second group, referred to as transcriptional co activators, alter transcription without directly binding to DNA. The PPAR gamma co-activator (PGC) family of proteins have been identified as the central family of transcriptional co-activators for induction of mitochondrial biogenesis. PGC-1 alpha is activated by exercise, and is sufficient to produce the endurance phenotype through direct interactions with NRF-1 and PPAR alpha, and potentially NRF-2. Furthering the understanding of the activation of PGC proteins following exercise has implications beyond improving athletic performance, including the possibility of providing targets for the treatment of frailty in the elderly, obesity and diseases such as mitochondrial myopathies and diabetes. PMID- 15294043 TI - Role of calcium and AMP kinase in the regulation of mitochondrial biogenesis and GLUT4 levels in muscle. AB - Contractile activity induces mitochondrial biogenesis and increases glucose transport capacity in muscle. There has been much research on the mechanisms responsible for these adaptations. The present paper reviews the evidence, which indicates that the decrease in the levels of high-energy phosphates, leading to activation of AMP kinase (AMPK), and the increase in cytosolic Ca(2+), which activates Ca(2+)/calmodulin-dependent protein kinase (CAMK), are signals that initiate these adaptative responses. Although the events downstream of AMPK and CAMK have not been well characterized, these events lead to activation of various transcription factors, including: nuclear respiratory factors (NRF) 1 and 2, which cause increased expression of proteins of the respiratory chain; PPAR alpha, which up regulates the levels of enzymes of beta oxidation; mitochondrial transcription factor A, which activates expression of the mitochondrial genome; myocyte-enhancing factor 2A, the transcription factor that regulates GLUT4 expression. The well-orchestrated expression of the multitude of proteins involved in these adaptations is mediated by the rapid activation of PPAR gamma co-activator (PGC) 1, a protein that binds to various transcription factors to maximize transcriptional activity. Activating AMPK using 5-aminoimidizole-4 carboxamide-1-beta-D-riboside (AICAR) and increasing cytoplasmic Ca(2+) using caffeine, W7 or ionomycin in L6 myotubes increases the concentration of mitochondrial enzymes and GLUT4 and enhances the binding of NRF-1 and NRF-2 to DNA. AICAR and Ca-releasing agents also increase the levels of PGC-1, mitochondrial transcription factor A and myocyte-enhancing factors 2A and 2D. These results are similar to the responses seen in muscle during the adaptation to endurance exercise and show that L6 myotubes are a suitable model for studying the mechanisms by which exercise causes the adaptive responses in muscle mitochondria and glucose transport. PMID- 15294044 TI - The role of calcium and calcium/calmodulin-dependent kinases in skeletal muscle plasticity and mitochondrial biogenesis. AB - Intracellular Ca(2+) plays an important role in skeletal muscle excitation contraction coupling and also in excitation-transcription coupling. Activity dependent alterations in muscle gene expression as a result of increased load (i.e. resistance or endurance training) or decreased activity (i.e. immobilization or injury) are tightly linked to the level of muscle excitation. Differential expression of genes in slow- and fast-twitch fibres is also dependent on fibre activation. Both these biological phenomena are, therefore, tightly linked to the amplitude and duration of the Ca(2+) transient, a signal decoded downstream by Ca(2+)-dependent transcriptional pathways. Evidence is mounting that the calcineurin-nuclear factor of activated T-cells pathway and the Ca(2+)/calmodulin-dependent kinases (CaMK) II and IV play important roles in regulating oxidative enzyme expression, mitochondrial biogenesis and expression of fibre-type specific myofibrillar proteins. CaMKII is known to decode frequency dependent information and is activated during hypertrophic growth and endurance adaptations. Thus, it was hypothesized that CaMKII, and possibly CaMKIV, are down regulated during muscle atrophy and levels of expression of CaMKII alpha, -II beta, -II gamma and -IV were assessed in skeletal muscles from young, aged and denervated rats. The results indicate that CaMKII gamma, but not CaMKIIalpha or beta, is up regulated in aged and denervated soleus muscle and that CaMKIV is absent in skeletal but not cardiac muscle. Whether CaMKII gamma up-regulation is part of the pathology of wasting or a result of some adaptational response to atrophy is not known. Future studies will be important in determining whether insights from the adaptational response of muscle to increased loads will provide pharmacological approaches for increasing muscle strength or endurance to counter muscle wasting. PMID- 15294045 TI - The role of uncoupling protein 3 in fatty acid metabolism: protection against lipotoxicity? AB - The physiological function of the mitochondrial uncoupling protein (UCP), UCP3, is still under debate. There is, however, ample evidence to indicate that, in contrast to UCP1, the primary function of UCP3 is not the dissipation of energy. Rather, several lines of evidence suggest that UCP3 is associated with cellular fatty acid metabolism. The highest levels of expression of UCP3 have been found in type 2 glycolytic muscle fibres, and fasting and high-fat diets up regulate UCP3. This up-regulation is most pronounced in muscle with a low fat oxidative capacity. Acute exercise also up regulates UCP3, and this effect has been shown to be a result of the exercise-induced increase in plasma fatty acid levels. In contrast, regular physical activity, which increases fat oxidative capacity, reduces UCP3 content. Based on these data it has been postulated that UCP3 functions to export those fatty acids that cannot be oxidized from the mitochondrial matrix, in order to prevent fatty acid accumulation inside the matrix. Several experiments have been conducted to test this hypothesis. Blocking carnitine palmitoyltransferase 1, thereby reducing fat oxidative capacity, rapidly induces UCP3. High-fat diets, which increase the mitochondrial supply of fatty acids, also up regulate UCP. However, feeding a similar amount of medium chain fatty acids, which can be oxidized inside the mitochondrial matrix and therefore does not need to be exported from the matrix, does not affect UCP3 protein levels. In addition, UCP3 is increased in patients with defective beta oxidation and is reduced after restoring oxidative capacity. In conclusion, it is suggested that UCP3 has an important physiological function in facilitating outward transport from the mitochondrial matrix of fatty acid anions that cannot be oxidized, thereby protecting against lipid-induced mitochondrial damage. PMID- 15294046 TI - Mitochondrial assembly: protein import. AB - The protein import process of mitochondria is vital for the assembly of the hundreds of nuclear-derived proteins into an expanding organelle reticulum. Most of our knowledge of this complex multisubunit network comes from studies of yeast and fungal systems, with little information known about the protein import process in mammalian cells, particularly skeletal muscle. However, growing evidence indicates that the protein import machinery can respond to changes in the energy status of the cell. In particular, contractile activity, a powerful inducer of mitochondrial biogenesis, has been shown to alter the stoichiometry of the protein import apparatus via changes in several protein import machinery components. These adaptations include the induction of cytosolic molecular chaperones that transport precursors to the matrix, the up-regulation of outer membrane import receptors, and the increase in matrix chaperonins that facilitate the import and proper folding of the protein for subsequent compartmentation in the matrix or inner membrane. The physiological importance of these changes is an increased capacity for import into the organelle at any given precursor concentration. Defects in the protein import machinery components have been associated with mitochondrial disorders. Thus, contractile activity may serve as a possible mechanism for up-regulation of mitochondrial protein import and compensation for mitochondrial phenotype alterations observed in diseased muscle. PMID- 15294047 TI - Intramyocellular triacylglycerol as a substrate source during exercise. AB - The role of intramyocellular triacylglycerol (IMTG) as a substrate source during exercise has recently regained much attention as a result of the proposed functional relationship between IMTG accumulation and the development of insulin resistance. It has been speculated that elevated NEFA delivery and/or impaired fatty acid (FA) oxidation result in intramyocellular accumulation of triacylglycerol and FA metabolites, which are likely to induce defects in the insulin signalling cascade, causing insulin resistance. The progressive accumulation of IMTG in sedentary patients and patients who are obese and/or have type 2 diabetes should therefore form a major therapeutic target, and efforts should be made to develop interventions that prevent excess IMTG accretion by stimulating their rate of oxidation. Although regular exercise is likely to represent such an effective means, there is much controversy about the actual contribution of the IMTG pool as a substrate source during exercise. The apparent discrepancy in the published literature might be explained by differences in the applied research protocol and the selected subject population, but most of all by the techniques that have been employed to estimate IMTG use during exercise. Data obtained in trained-endurance athletes indicate that athletes can substantially reduce their IMTG pool following a single exercise session. With the growing awareness that skeletal muscle has a tremendous potential to oxidise IMTG during prolonged moderate-intensity exercise, more research is warranted to develop combined exercise, nutritional and/or pharmacological interventions that can stimulate IMTG oxidation in sedentary patients and patients who are obese and/or have type 2 diabetes. PMID- 15294048 TI - Regulation and role of hormone-sensitive lipase in rat skeletal muscle. AB - Intramyocellular triacylglycerol (TG) is an important energy store, and the energy content of this depot is higher than the energy content of the muscle glycogen depot. It has recently been shown that the mobilization of fatty acids from this TG pool may be regulated by the neutral lipase hormone-sensitive lipase (HSL). This enzyme is known to be rate limiting for intracellular TG hydrolysis in adipose tissue. The presence of HSL has been demonstrated in all muscle fibre types by Western blotting of muscle fibres isolated by collagenase treatment or after freeze-drying. The content of HSL varies between fibre types, being higher in oxidative fibres than in glycolytic fibres. When analysed under conditions optimal for HSL, neutral lipase activity in muscle can be stimulated by adrenaline as well as by contractions. These increases are abolished by the presence of anti-HSL antibody during analysis. Moreover, immunoprecipitation with affinity-purified anti-HSL antibody causes similar reductions in muscle HSL protein concentration and in measured neutral lipase responses to contractions. The immunoreactive HSL in muscle is stimulated by adrenaline via beta-adrenergic activation of cAMP-dependent protein kinase (PKA). From findings in adipocytes it is likely that PKA phosphorylates HSL at residues Ser(563), Ser(659) and Ser(660). Contraction probably also enhances muscle HSL activity by phosphorylation, because the contraction-induced increase in HSL activity is elevated by the protein phosphatase inhibitor okadaic acid and reversed by alkaline phosphatase. A novel signalling pathway in muscle by which HSL activity may be stimulated by protein kinase C (PKC) via extracellular signal-regulated kinase (ERK) has been demonstrated. In contrast to previous findings in adipocytes, in muscle the activation of ERK is not necessary for stimulation of HSL by adrenaline. However, contraction-induced HSL activation is mediated by PKC, at least partly via the ERK pathway. In fat cells ERK is known to phosphorylate HSL at Ser(600). Hence, phosphorylation of different sites may explain the finding that in muscle the effects of contractions and adrenaline on HSL activity are partially additive. In line with the view that the two stimuli act by different mechanisms, training increases contraction-mediated HSL activation but diminishes adrenaline-mediated HSL activation in muscle. In conclusion, HSL is present in skeletal muscle and can be activated by phosphorylation in response to both adrenaline and muscle contractions. Training increases contraction-mediated HSL activation, but decreases adrenaline-mediated HSL activation in muscle. PMID- 15294049 TI - Regulation and role of hormone-sensitive lipase activity in human skeletal muscle. AB - Hormone-sensitive lipase (HSL) is believed to play a regulatory role in initiating the degradation of intramuscular triacylglycerol (IMTG) in skeletal muscle. A series of studies designed to characterise the response of HSL to three stimuli: exercise of varying intensities and durations; adrenaline infusions; altered fuel supply have recently been conducted in human skeletal muscle. In an attempt to understand the regulation of HSL activity the changes in the putative intramuscular and hormonal regulators of the enzyme have also been measured. In human skeletal muscle at rest there is a high constitutive level of HSL activity, which is not a function of biopsy freezing. The combination of low adrenaline and Ca(2+) levels and resting levels of insulin appear to dictate the level of HSL activity at rest. During the initial minute of low and moderate aerobic exercise HSL is activated by contractions in the apparent absence of increases in circulating adrenaline. During intense aerobic exercise, adrenaline may contribute to the early activation of HSL. The contraction-induced activation may be related to increased Ca(2+) and/or other unknown intramuscular activators. As low- and moderate-intensity exercise continues beyond a few minutes, activation by adrenaline through the cAMP cascade may also occur. With prolonged moderate intensity exercise beyond 1-2 h and sustained high-intensity exercise, HSL activity decreases despite continuing increases in adrenaline, possibly as a result of increasing accumulations of free AMP, activation of AMP kinase and phosphorylation of inhibitory sites on HSL. The existing work in human skeletal muscle also suggests that there are numerous levels of control involved in the regulation of IMTG degradation, with control points downstream from HSL also being important. For example, it must be remembered that the actual flux (IMTG degradation) through HSL may be allosterically inhibited during prolonged exercise as a result of the accumulation of long-chain fatty acyl-CoA. PMID- 15294050 TI - Basic disturbances in skeletal muscle fatty acid metabolism in obesity and type 2 diabetes mellitus. AB - The present article addresses the hypothesis that disturbances in skeletal muscle fatty acid handling in abdominal obesity and type 2 diabetes mellitus may play a role in the aetiology of increased adipose tissue stores, increased triacylglycerol storage in skeletal muscle and skeletal muscle insulin resistance. The uptake and/or oxidation of fatty acids have been shown to be impaired during post-absorptive conditions in abdominally-obese subjects and/or subjects with type 2 diabetes. Also, human studies have shown that muscle of subjects that are (abdominally) obese and/or have type 2 diabetes is characterized by an inability to increase fatty acid uptake and/or fatty acid oxidation during beta-adrenergic stimulation and exercise. This disturbance in fat oxidation may promote, on one hand, the development of increased adipose tissue stores and obesity. On the other hand, fatty acids that are taken up by muscle and not oxidized may increase triacylglycerol storage in muscle, which has been associated with skeletal-muscle insulin resistance. Disturbances in the capacity to increase fat oxidation during post-absorptive conditions, beta adrenergic stimulation and exercise in subjects who are obese and/or have type 2 diabetes persist after weight reduction, indicating that the diminished fat oxidation may be a primary factor leading to the obese and/or insulin-resistant state rather than an adaptational response. Clearly, the precise sequence of events leading to an increased adiposity and insulin resistance in obesity and type 2 diabetes mellitus is not yet fully understood. PMID- 15294051 TI - Mechanotransduction and the regulation of protein synthesis in skeletal muscle. AB - Repeated bouts of resistance exercise produce an increase in skeletal muscle mass. The accumulation of protein associated with the growth process results from a net increase in protein synthesis relative to breakdown. While the effect of resistance exercise on muscle mass has long been recognized, the mechanisms underlying the link between high-resistance contractions and the regulation of protein synthesis and breakdown are, to date, poorly understood. In the present paper skeletal muscle will be viewed as a mechanosensitive cell type and the possible mechanisms through which mechanically-induced signalling events lead to changes in rates of protein synthesis will be examined. PMID- 15294052 TI - Insulin-like growth factor 1 and muscle growth: implication for satellite cell proliferation. AB - Insulin-like growth factor 1 (IGF-1) has been shown to rescue the aging-related or inactivity-induced loss of muscle mass through the activation of satellite cells. However, the signalling pathways and the mechanism by which IGF-1 affects satellite cells have not been not completely identified. The purpose of the present review is to provide current understanding of the cellular and molecular events underlying IGF-1 induced proliferation of satellite cells. PMID- 15294053 TI - Calcineurin and skeletal muscle growth. AB - Recruitment determines the profile of fibre-type-specific genes expressed across the range of muscle fibres associated with slow, fast fatigue-resistant and fast fatiguable motor units. Downstream signalling pathways activated by neural signalling and mechanical load have been the focus of intensive research in past years. It is now known that Ca(2+)-dependent calcineurin-nuclear factor of activated T cells and insulin-like growth factor 1 pathways and their downstream mediators contribute to these adaptive responses. These pathways regulate gene expression through muscle-specific (myocyte-enhancing factor 2, myoblast determination protein) and non-specific (nuclear factor of activated T cell 2, GATA-2) transcription factors. Transcriptional signals activated with increased contractile activity result in altered expression of fibre-type specific genes, including the myosin heavy chain isoforms and oxidative and glycolytic enzymes and a net change in muscle fibre-type composition. In contrast, transcriptional signals activated by increased load bearing result in hypertrophy or a growth response, a component of which involves satellite cell recruitment and fusion with existing adult myofibres. Calcineurin has been identified as a key mediator in the hypertrophic response, and the current challenge has been to determine the downstream target genes of this pathway. Exciting new data have emerged, showing that myostatin, a negative regulator of muscle growth, and utrophin, a cytoskeletal protein important in maintaining membrane integrity, are downstream targets of calcineurin signalling. Increased understanding of these mediators of muscle growth may provide strategies for the development of effective therapeutics to counter muscle weakness and muscular dystrophy. PMID- 15294054 TI - Regulation of protein synthesis associated with skeletal muscle hypertrophy by insulin-, amino acid- and exercise-induced signalling. AB - Although insulin, amino acids and exercise individually activate multiple signal transduction pathways in skeletal muscle, one pathway, the phosphatidylinositol 3 kinase (PI3K)-mammalian target of rapamycin (mTOR) signalling pathway, is a target of all three. Activation of the PI3K-mTOR signal transduction pathway results in both acute (i.e. occurring in minutes to hours) and long-term (i.e. occurring in hours to days) up-regulation of protein synthesis through modulation of multiple steps involved in mediating the initiation of mRNA translation and ribosome biogenesis respectively. In addition, changes in gene expression through altered patterns of mRNA translation promote cell growth, which in turn promotes muscle hypertrophy. The focus of the present discussion is to review current knowledge concerning the mechanism(s) through which insulin, amino acids and resistance exercise act to activate the PI3K-mTOR signal transduction pathway and thereby enhance the rate of protein synthesis in muscle. PMID- 15294055 TI - The role of ubiquitin-proteasome-dependent proteolysis in the remodelling of skeletal muscle. AB - In skeletal muscle, as in any mammalian tissue, protein levels are dictated by relative rates of protein synthesis and breakdown. Recent studies have shown that the ubiquitin-proteasome-dependent proteolytic pathway is mainly responsible for the breakdown of myofibrillar proteins. In this pathway proteins that are to be degraded are first tagged with a polyubiquitin degradation signal. Ubiquitination is performed by the ubiquitin-activating enzyme, ubiquitin-conjugating enzymes and ubiquitin-protein ligases, which are responsible for the recognition of specific substrates. Polyubiquitinated protein substrates are then specifically recognised and degraded by the 26S proteasome. The present review focuses on: (1) the mechanisms of ubiquitination-deubiquitination that make the system highly selective; (2) the mechanisms of proteolysis in skeletal muscle. In particular, the role of the system in the remodelling of skeletal muscle during exercise and disuse and in recovery or regeneration that prevails during post-atrophic conditions is reviewed. PMID- 15294056 TI - Metabolic flexibility. AB - Human physiology needs to be well adapted to cope with major discontinuities in both the supply of and demand for energy. This adaptability requires 'a clear capacity to utilize lipid and carbohydrate fuels and to transition between them' (Kelley et al. 2002b). Such capacities characterize the healthy state and can be termed 'metabolic flexibility'. However, increasing evidence points to metabolic inflexibility as a key dysfunction of the cluster of disease states encompassed by the term 'metabolic syndrome'. In obese and diabetic individuals this inflexibility is manifest in a range of metabolic pathways and tissues including: (1) failure of cephalic-phase insulin secretion (impaired early-phase prandial insulin secretion concomitant with failure to suppress hepatic glucose production and NEFA efflux from adipose tissue); (2) failure of skeletal muscle to appropriately move between use of lipid in the fasting state and use of carbohydrate in the insulin-stimulated prandial state; (3) impaired transition from fatty acid efflux to storage in response to a meal. Finally, it is increasingly clear that reduced capacity for fuel usage in, for example, skeletal muscle, as indicated by reduced mitochondrial size and density, is characteristic of the metabolic syndrome state and a fundamental component of metabolic inflexibility. Key questions that remain are how metabolic flexibility is lost in obese and diabetic individuals and by what means it may be regained. PMID- 15294057 TI - Physiological regulation of NEFA availability: lipolysis pathway. AB - Plasma NEFA are an important energy substrate and, furthermore, play a key role in the induction of insulin resistance in the body. The availability of NEFA is determined predominantly by their mobilization from adipose tissue triacylglycerol stores by the process of lipolysis. Adipose tissue lipolysis in man is regulated by a number of hormonal and paracrine and/or autocrine signals. The main hormonal signals may be represented by catecholamines, insulin, growth hormone, natriuretic peptides and some adipocytokines. The absolute levels and relative importance and contribution of these signals vary in different physiological situations, with diet and physical exercise being the main physiological variables that affect the hormonal signalling. Thus, modulations in hormonal signals induce an increase in NEFA mobilization in the post-absorptive state and during an acute bout of exercise, and suppress NEFA mobilization in the postprandial state. In addition, hormonal regulation is modified by long-term interventions in energy balance, such as dietary restriction and/or physical training, and is disturbed in some pathological states, such as obesity or diabetes. The question that remains is whether disturbances in lipolysis regulation in obese and diabetic subjects may be 'corrected' by the long-term interventions in diet and physical activity. PMID- 15294058 TI - The molecular mechanism linking muscle fat accumulation to insulin resistance. AB - Skeletal muscle insulin resistance is a co-morbidity of obesity and a risk factor for the development of type 2 diabetes mellitus. Insulin resistance is associated with the accumulation of intramyocellular lipids. Intramyocellular triacylglycerols do not appear to be the cause of insulin resistance but are more likely to be a marker of other lipid intermediates such as fatty acyl-CoA, ceramides or diacylglycerols. Fatty acyl-CoA, ceramides and diacylglycerols are known to directly alter various aspects of the insulin signalling cascade. Insulin signalling is inhibited by the phosphorylation of serine and threonine residues at the levels of the insulin receptor and insulin receptor substrate 1. Protein kinase C is responsible for the phosphorylation of the serine and threonine residues. Fatty acyl-CoA and diacylglycerols are known to activate protein kinase C. The cause of the intramyocellular accumulation of fatty acyl CoA and diacylglycerols is unclear at this time. Reduced fatty acid oxidation does not appear to be responsible, as fatty acyl-CoA accumulates in skeletal muscle with a normal fatty acid oxidative capacity. Other potential mechanisms include oversupply of lipids to muscle and/or up regulated fatty acid transport. PMID- 15294059 TI - Metabolic and hormonal interactions between muscle and adipose tissue. AB - From the perspective of a muscle physiologist, adipose tissue has long been perceived predominantly as a fuel reservoir that provides muscle and other tissues with NEFA when exogenous nutrients are insufficient for their energy needs. Recently, studies have established that adipose tissue is also an endocrine organ. Among the hormones it releases are adiponectin and leptin, both of which can activate AMP-activated protein kinase and increase fatty acid oxidation in skeletal muscle and probably other tissues. Deficiencies of leptin or leptin receptor, adiponectin and IL-6 are associated with obesity, insulin resistance and a propensity to type 2 diabetes. In addition, a lack of adiponectin has been linked to atherosclerosis. Whether this pathology reflects a deficient activation of AMP-activated protein kinase in peripheral tissues remains to be determined. Finally, recent studies have suggested that skeletal muscle may also function as an endocrine organ when it releases the cytokine IL-6 into the circulation during sustained exercise. Interestingly, one of the apparent effects of IL-6 is to stimulate lipolysis, causing the release of NEFA from the adipocyte. Thus, hormonal communications exist between the adipocyte and muscle that could enable them to talk to each other. The physiological relevance of this cross talk clearly warrants further study. PMID- 15294060 TI - IL-8 inhibits isoproterenol-stimulated ciliary beat frequency in bovine bronchial epithelial cells. AB - Mucociliary clearance is a critical host defense that protects the lung. The mechanisms by which mucociliary function is altered by inflammation are poorly defined. Chronic exposure to cigarette smoke decreases ciliary beating and interferes with proper airway clearance. Bronchoalveolar lavage (BAL) fluid from smokers and ex-smokers has increased amounts of IL-8, which has played a critical role in airway inflammation. We hypothesized that IL-8 might interfere with stimulated ciliary beating in airway epithelium. To test this hypothesis, we stimulated bovine ciliated bronchial epithelial cells (BBECs) with a known activator of ciliary beat frequency (CBF), isoproterenol (ISO; 100 microM), in the presence or absence of IL-8 (100 pg/mL). We measured CBF digitally using the Sisson-Ammons Video Analysis (SAVA) system. CBF increased in untreated cells exposed to ISO (approximately 3 Hz) over baseline. In contrast, cells pre incubated with IL-8 failed to respond to ISO. Pretreatment with IL-8 also blocked ISO-stimulated cAMP-dependent kinase (PKA) activation, which is known to control ISO-stimulated CBF. In addition, IL-8 pretreated cells revealed a marked decrease in PKA activity when cells were stimulated with forskolin (FSK; 10 microM). Cells were assayed specifically for cAMP-phosphodiesterase (PDE) activity. ISO stimulated cells demonstrated an increase in PDE activity as compared to control. Pretreatment with IL-8 had no effect on ISO-stimulated PDE activity. Collectively, these data suggest that IL-8 appears to mediate its effect at the level of adenylyl cyclase. It is also possible that IL-8 may not only act as a chemotactic agent, but also as a potential autocrine/paracrine inhibitor of PKA mediated stimulation of ciliary motility. In conclusion, IL-8 inhibits beta agonist dependent ciliostimulation and such inhibition of stimulated ciliary activity may contribute to the impaired mucociliary clearance seen in airway diseases. Furthermore, since IL-8 levels are increased in the airway of cigarette smokers, it is likely they may be more resistant to the cilio and muco ciliostimulating effects of beta-agonists. PMID- 15294061 TI - Does wearing a noseclip during inhalation improve lung deposition? AB - Using nebulization, only a small proportion of the dose reaches the lungs, while the remainder is swallowed, exhaled into the atmosphere, or remains in the nebulizer. It was the purpose of this study to investigate whether wearing a noseclip during inhalation can improve lung deposition. Relative lung deposition was compared by inhalation of the marker substance, sodium cromoglycate (SCG), and measurement of urinary excretion of SCG. The SCG absorption half-life allows one to differentiate indirectly between a more or less peripheral deposition. Ten CF patients (9-18 years old) inhaled, under routine conditions, a solution containing 20 mg of SCG in a randomized crossover design through a mouthpiece, without and with a noseclip being worn. Following inhalation without and with a noseclip, no statistically significant difference was seen in the amount of SCG excreted in urine (0.9 +/- 0.4 mg vs. 1.0 +/- 0.5 mg; p = 0.402) and absorption half-life (93 +/- 25 min vs. 113 +/- 36 min; p = 0.083). In conclusion, wearing a noseclip during inhalation under conditions relevant to practice does not increase the amount deposited into the lungs of CF patients and, also, there has been no indication of a more peripheral lung deposition. PMID- 15294062 TI - The development of a novel high-dose pressurized aerosol dry-powder device (PADD) for the delivery of pumactant for inhalation therapy. AB - The performance of a novel dry powder inhaler designed to deliver exceptionally high doses was investigated using pumactant as a model powder. Pumactant (a synthetic lung surfactant consisting of a phospholipid mixture), with a 90th percentile particle size of 2.92 microm is highly cohesive, has a high moisture affinity (6.2% w/w at 45% RH), and is predominantly amorphous. The device (pressurized aerosol dry-powder delivery [PADD]) utilizes pressurized gas to aerosolize a powder bed from a reservoir and delivers it through a conventional mouthpiece. The influence of loaded dose on dry powder delivery and can pressure on aerosolization efficiency was investigated. Analysis of the delivered dose studies suggested a linear relationship between loaded dose and delivered dose (R(2) = 0.96, for loaded doses of 0-250 mg), with a delivery efficiency of 70%. Analysis of the aerosolization efficiency using a Marple Miller type impactor suggested fine particle fractions (particles with an aerodynamic diameter of <5 microm) of approximately 30% using canister pressures of 8-14 bars. These results indicate that the PADD device may be a useful tool in delivering high-dose medicaments, as a carrier-free formulation, to the deep lung. PMID- 15294063 TI - Comparison of patient preference and ease of teaching inhaler technique for Pulmicort Turbuhaler versus pressurized metered-dose inhalers. AB - A multicenter, randomized, open-label, crossover study with two 4-week evaluation periods compared patient preference and ease of teaching correct inhaler technique for Pulmicort Turbuhaler versus pressurized metered-dose inhalers (pMDIs). Patients 18 to 65 years of age with stable, mild to moderate asthma, who required or were eligible for inhaled corticosteroid therapy, were randomized to treatment sequences consisting of 4-week evaluation periods with Pulmicort Turbuhaler (budesonide inhalation powder) two puffs (400 microg) bid and one of three inhaled corticosteroids via pMDI: Aerobid-M (flunisolide) four puffs (1 mg) bid, Flovent (fluticasone propionate) two puffs (440 microg) bid, or Vanceril Double Strength (beclomethasone dipropionate) five puffs (420 microg) bid. Patients indicated device preference at study end and completed the Patient Device Experience Assessment (PDEA) questionnaire after each evaluation period. Ease of teaching, time required to master use of the device, percentage of patients demonstrating mastery on the first attempt, and the number of attempts required to demonstrate mastery were assessed. Despite previous use of pMDIs by most patients, Pulmicort Turbuhaler was significantly preferred (p < 0.001) and required significantly less time to master than pMDIs (p < 0.001). Median times to device mastery were 3.67 min for Pulmicort Turbuhaler versus 5.33 min for pMDIs. Patients rated Pulmicort Turbuhaler significantly better than pMDIs on PDEA ease of use (p = 0.0005) and overall satisfaction (p < 0.0001) single-item scales and all four multi-item scales (pharyngeal symptoms, oral sensation, operational use, and inhaler attributes; p < 0.05). Overall, patients preferred Pulmicort Turbuhaler over pMDIs and required less time to be taught how to correctly use Turbuhaler trade mark. PMID- 15294064 TI - Dosimetry and toxicology of ultrafine particles. AB - While epidemiological studies indicate an association between adverse health effects and ambient ultrafine particle concentrations in susceptible individuals, toxicological studies aim to identify mechanisms which are causal for the gradual transition from the physiological status towards patho-physiological disease. Impressive progress has been made in recent years when objectives changed from classical tests like lung function, etc. to endpoints comprising of particle induced oxidative stress, cell signaling and activation, release of mediators initiating inflammatory processes not only in the respiratory tract but also in the cardio-vascular system. Particularly, the large surface area of ultrafine particles provides a unique interface for catalytic reactions of surface-located agents with biological targets like proteins, cells, etc. However, toxicological studies are hampered by a number of imminent complications when simulating long term exposure of humans in urban environments with inherited and/or acquired susceptibility (e.g., acute exposure studies at high concentrations either in human subjects or animal models). Yet, based on a conservative estimate results available begin to show an adverse health risk for susceptible individuals and support the epidemiological evidence. PMID- 15294065 TI - Deposition of aerosols in infants and children. AB - There is still a lack of knowledge in the field of aerosol therapy in children, particularly in young children. The amount of drug delivered from a commercially available inhalation device that reaches the lungs of children is generally low. The choice of an optimal combination of delivery device and drug formulation based on individual patient related factors is crucial. Aerosols with a small MMAD and a narrow GSD are required for a sufficient inhalation therapy in early childhood. The development of combinations of delivery devices and drug formulations fulfilling the requirements for an efficient inhalation therapy in young children is likely to increase the therapeutical options in this age group. PMID- 15294066 TI - Biological effects of Utah Valley ambient air particles in humans: a review. AB - The Utah Valley provided a unique opportunity to evaluate the health effects of particulate matter (PM) in humans. The area has had intermittently high particle levels with the principal point source being a steel mill. Due to a labor dispute, the mill was shut down. The closure and reopening of the steel mill allowed for an examination of potential correlates between epidemiological observations and measures of the biological effect of PM with experimental cell and human exposure. Epidemiologic investigation demonstrated an association of both the closure of the steel mill and the reduction in exposure to air pollution particles with changes in morbidity and mortality. Changes in these parameters were not fully accounted for by variation in the mass of PM. Metal content, in vitro oxidative stress, and release of pro-inflammatory mediators by cultured respiratory epithelial cells were all elevated in those aqueous extracts collected from the Utah Valley while the steel mill was open. Similarly, inflammatory injury in the lower respiratory tract of humans after instillation of aqueous extracts of filter PM was increased in those volunteers exposed to particles collected while the mill was open. These results indicate that equal masses of PM can induce disparate lung injuries suggesting that particle components may be relevant in assessing health effects after their exposure. Specifically, metals can participate in the biological effects of PM collected from the Utah Valley. In addition, correlates between findings of epidemiological studies and the biological effects of PM in cell and human investigation were demonstrated. PMID- 15294067 TI - House dust mite and cockroach exposure: risk factors for asthma. PMID- 15294068 TI - Virus-induced asthma attacks. AB - Viral infections are responsible for the majority of asthma attacks in both children and adults. Vagally mediated reflex bronchoconstriction is potentiated due to loss of function of inhibitory M2 muscarinic receptors on the airway parasympathetic nerves. Multiple mechanisms are involved. Production of interferons may down regulate the expression of the M2 receptor gene. This effect is reversed by steroids. In allergic animals (and perhaps in atopic humans) eosinophils are recruited to the airway nerves, where they are activated, releasing major basic protein, which binds to the M2 receptors, blocking their function. Despite these negative physiological consequences of eosinophil activation, eosinophils are capable of exerting a potent antiviral effect. Thus, the inflammatory response to viral infections may have positive, as well as negative, consequences. PMID- 15294069 TI - PEG-PLA nanoparticles as carriers for nasal vaccine delivery. AB - This report presents an overview of the potential of nanoparticles as nasal carriers for drug/vaccine administration. In addition, this report shows, for the first time, the efficacy of polylactic acid nanoparticles coated with a hydrophilic polyethyleneglycol coating (PEG-PLA nanoparticles) as carriers for the nasal transport of bioactive compounds. For this purpose, tetanus toxoid (TT), a high molecular weight protein (Mw 150,000 Da), was chosen as a model antigen and encapsulated in the PEG-PLA nano- and microparticles (200 nm and 1.5 microm respectively). These nanosystems were first characterized for their stability in the presence of lysozyme and also for their size, electrical charge, loading efficiency, in vitro release of antigenically active toxoid and afterwards, these formulations were administered intranasally to mice and the systemic and mucosal anti-tetanus responses were evaluated for up to 24 weeks. Additionally, PEG-PLA particles labeled with rhodamine 6G were administered intranasally to rats in order to visualize their interaction with the nasal mucosae by fluorescence microscopy. Their behavior was compared with that of the well known PLA nanoparticles (200 nm). The results showed that PLA nanoparticles suffered an immediate aggregation upon incubation with lysozyme, whereas the PEG coated nanoparticles remained totally stable. The antibody levels elicited following i.n. administration of PEG-coated nanoparticles were significantly higher than those corresponding to PLA nanoparticles. Furthermore, PEG-PLA nanoparticles generated an increasing and a long lasting response. The qualitative fluorescence microscopy studies revealed that PEG-PLA particles are able to cross the rat nasal epithelium. These studies indicate that the PEG coating around the particles has a role in stabilizing PLA particles in mucosal fluids and that it facilitates the transport of the nanoencapsulated antigen, hence eliciting a high and long lasting immune response. PMID- 15294070 TI - Inhaler devices in infants and children: challenges and solutions. PMID- 15294071 TI - Setting standards in gene repair. PMID- 15294072 TI - Inhibition of ampicillin-resistant bacteria by novel mono-DNAzymes and di-DNAzyme targeted to beta-lactamase mRNA. AB - In view of the weakness of antibiotics and the properties of antisense drugs, we applied DNAzymes to the field of drug resistance in bacteria. Two 10-23 mono DNAzymes (Dz1, Dz2) and a di-DNAzyme (Dz1-2) targeted to beta-lactamase mRNA were designed to determine to what degree the growth of ampicillin-resistant bacteria (TEM-1, TEM-3) was inhibited. All three DNAzymes can play a role both in vitro and in vivo. In vitro, they exhibited high catalytic efficiency (kcat/KM) of 63.5, 91.1, and 30.8 pM(-1) min(-1), respectively, under multiple-turnover conditions. In vivo, after 9 hours' incubation, the degree of inhibition of Dz1, Dz2, and Dz1-2 for TEM-1 bacteria was 27.2%, 39.6%, and 57.7%, respectively, and that for TEM-3 bacteria was 39.1%, 44%, and 62.6%, respectively. Dz1-2 showed the greatest inhibiting effect, demonstrating in vivo activity may be increased by constructing multiple-target DNAzymes. The results indicated a potential possibility for DNAzymes to act as a new type of antibacterial or a tool of gene functional analysis for prokaryocytes. PMID- 15294073 TI - Poly-2'-DNP-RNAs with enhanced efficacy for inhibiting cancer cell growth. AB - It is often believed that small interfering RNA (siRNA) is at least 10-fold more effective than the single-stranded antisense oligonucleotide for silencing the same target gene in the same cells. In view of the recent discovery that the RNA induced silencing complex (RISC) contains only a single-stranded RNA (ssRNA) molecule and can be reconstituted using single-stranded antisense RNA, such a large difference in efficacy seems puzzling. One possible reason is that hybridization protects siRNA from hydrolysis by endogenous RNase activity until it is incorporated in the RISC, whereas ssRNA is rapidly hydrolyzed. Because the single-stranded poly-2'-O-(2,4-dinitrophenyl)-RNA (DNP-ssRNA) is both RNase resistant and membrane permeable, we synthesized homologous native siRNAs, DNP siRNAs, native ssRNAs, and DNP-ssRNAs and made a comparative study of their efficacies for inhibiting the growth of two cancer cell lines with different overexpressed target genes under equivalent experimental conditions. It was found that the efficacy of antisense DNP-ssRNA is higher than that of the corresponding siRNA and that the efficacy of native siRNA for inhibiting cell growth can also be enhanced from 2-fold to 6-fold by replacing the native strands of RNA in siRNA with homologous DNP-RNA. Thermal denaturation data show that the hybridization affinity of the DNP-RNA/RNA duplex is higher than that of the native RNA/RNA duplex. Western blotting analysis of A549 cells treated with antisense DNP-ssRNAs containing single mismatching bases shows that the gene silencing by antisense DNP-ssRNA is as sequence specific as that by siRNA. The observed large enhancement of inhibition efficacy of native RNAs by DNP derivatization should be advantageous for both gene silencing studies and therapeutic applications. PMID- 15294074 TI - Use of siRNAs and antisense oligonucleotides against survivin RNA to inhibit steps leading to tumor angiogenesis. AB - The antiapoptotic protein survivin is an attractive target in cancer therapy because it is expressed differently in tumors and normal tissues and it is potentially required for cancer cells to remain viable. Given that survivin is also overexpressed in endothelial cells (ECs) of newly formed blood vessels found in tumors, its RNA targeting might compromise EC viability and interfere with tumor angiogenesis. We used two antisense strategies against survivin expression, antisense oligonucleotides (aODN) and small interfering RNA (siRNA), to study in ECs the contribution of survivin in various steps leading to tumor angiogenesis. A 21-mer phosphorothioate aODN and two siRNA oligonucleotides against survivin mRNA were designed to downregulate survivin expression. Survivin targeting caused (1) a strong growth-inhibitory effect, (2) a 4-fold increase in apoptosis, (3) an accumulation of cells in the S phase and a decrease in G2/M phase, (4) a dose dependent inhibition of EC migration on Vitronectin, and (5) a decrease in capillary formation. Control oligonucleotides, an unrelated oligonucleotide, and one with four mismatches, had no significant effect. All these results show that survivin is a suitable target in cancer therapy because its inhibition in EC causes both a proapoptotic effect and an interruption of tumor angiogenesis. The two strategies used, classic aODN and siRNA technology, were very effective. Moreover, the latter can be used in the low nanomolar range, thus increasing the sensitivity of the treatment. PMID- 15294075 TI - In vitro selection of RNA aptamers against the HCV NS3 helicase domain. AB - Nonstructural protein 3 (NS3) of hepatitis C virus (HCV) has two distinct domains, protease and helicase, that are essential for HCV proliferation. Therefore, NS3 is considered a target for anti-HCV treatment. To study RNA aptamers of the NS3 helicase domain, we carried out in vitro selection against the HCV NS3 helicase domain. RNA aptamers obtained after eight generations possessed 5' extended single-stranded regions and the conserved sequence (5' GGA(U/C)GGAGCC-3') at stem-loop regions. Aptamer 5 showed strong inhibition of helicase activity in vitro. Deletion and mutagenesis analysis clarified that the conserved stem-loop is important and that the whole structure is needed for helicase inhibition. We compared the inhibition of helicase activity between aptamer 5 and 3'+-UTR of HCV. PMID- 15294076 TI - Locked nucleic acid: a potent nucleic acid analog in therapeutics and biotechnology. AB - Locked nucleic acid (LNA) is a class of nucleic acid analogs possessing very high affinity and excellent specificity toward complementary DNA and RNA, and LNA oligonucleotides have been applied as antisense molecules both in vitro and in vivo. In this review, we briefly describe the basic physiochemical properties of LNA and some of the difficulties that may be encountered when applying LNA technology. The central part of the review focuses on the use of LNA molecules in regulation of gene expression, including delivery to cells, stability, unspecific effects, toxicity, pharmacokinetics, and design of LNA oligonucleotides. The last part evaluates LNA as a diagnostic tool in genotyping. PMID- 15294077 TI - Downregulation of p21(WAF1/CIP1) and estrogen receptor alpha in MCF-7 cells by antisense oligonucleotides containing locked nucleic acid (LNA). AB - Locked nucleic acid (LNA) is a nucleic acid analog with very high affinity to complementary RNA and a promising compound in the field of antisense research. The intracellular localization and quantitative uptake of oligonucleotides containing LNA were found to be equivalent to those of phosphorothioate oligonucleotides (PS AONs). The antisense efficiency of LNA-containing oligonucleotides was systematically compared with standard PS AONs targeting expression of two endogenous proteins in the human breast cancer cell line MCF-7, namely, the cyclin-dependent kinase inhibitor p21(WAF1/CIP1) and the estrogen receptor alpha (ERalpha). For downregulation of both target proteins, the most efficient design was achieved with oligonucleotides containing LNA monomers in the extremities and a central gap of PS-linked DNA monomers, so called LNA gapmers. Such LNA gapmers caused more potent downregulation of the targeted proteins than PS AONs, whereas fully modified LNA AONs or LNA mixmers (LNA nucleotides interspersed) were inactive. PMID- 15294078 TI - Oligonucleotide-based gene targeting approaches. PMID- 15294081 TI - The experts speak. Public policy issues in HIV/AIDS. Interview by Vicki Glaser. PMID- 15294082 TI - Tolerance of didanosine as enteric-coated capsules versus buffered tablets. PMID- 15294083 TI - HIV infection and antiphospholipid antibody: literature review and link to the antiphospholipid syndrome. AB - There is a high incidence of antiphospholipid antibodies, detected by assays for anticardiolipin or lupus-like anticoagulant, in HIV disease. However, a link to the antiphospholipid syndrome, with clinical thrombosis, is tenuous. We report a case of a 25-year-old man with undetermined risk factors for HIV presenting with possible antiphospholipid syndrome manifesting as necrotic skin lesions as the initial clinical presentation for HIV. We also review the literature exploring the association between HIV and antiphospholipid syndrome. PMID- 15294084 TI - Development of opportunistic infections after diagnosis of active tuberculosis in HIV-infected patients. AB - Despite advances in highly active antiretroviral therapy (HAART), its use during tuberculosis (TB) treatment is fraught with challenges, often leading to delayed therapy. This report describes the course of HIV infection and outcomes of 26 consecutive TB/HIV coinfected patients who received TB treatment in Rhode Island. HIV infection was diagnosed in 26 (4%) of 598 TB cases in during a 10-year period. Of these patients, TB was the first indication of HIV infection in 15 patients (58%). Of the 21 patients who were not on antiretrovirals at the time of TB diagnosis, 17 (81%) met criteria for immediate initiation of HAART. The median CD4 cell counts and HIV-1 plasma viral load were 80 cells per microliter (range, 2-800 cells per microliter) and 255,631 copies per milliliter (range, 50,000 to > 500,000 copies per milliliter), respectively, for the patients whose baseline measurements were available. CD4 lymphocyte count was less than 200 cells per microliter in 13 (76%) of the 17 patients whose measurements were available. Three (30%) of the 10 patients whose CD4 cell count was less than 100 cells per microliter developed subsequent AIDS-defining illness prior to the initiation of HAART and a fourth patient, within 30 days of starting HAART. None of the patients who had CD4 cell counts 100 cells per microliter or greater developed subsequent AIDS-defining illness during TB treatment. The median time to starting HAART after starting TB treatment was 12 weeks (range, 3-36 weeks). From our limited experience based on this case series, we recommend early initiation of HAART in coinfected patients with CD4 cell counts less than 100 cells per microliter. PMID- 15294085 TI - Screening for potentially transmitting sexual risk behaviors, urethral sexually transmitted infection, and sildenafil use among males entering care for HIV infection. AB - The study aims were to evaluate the prevalence and predictors of sexual risk behaviors and urethral sexually transmitted disease (STD) among males entering care for HIV infection and to examine if sildenafil prescriptions are associated with potentially transmitting sexual risk behavior (PTSRB). The research design included (1) self-administered questionnaire of symptoms of sexually transmitted infection (STI), number of recent sex partners, unprotected sexual risk behaviors, use of drugs/alcohol during sex, and HIV disclosure; (2) urine gonorrhea/chlamydia polymerase chain reaction (PCR); and (3) record review for sildenafil prescriptions. A PTSRB was defined as insertive anal, vaginal, or oral sex without a condom. Between March 2001 and March 2002, 413 entrants were surveyed. The prevalence of positive urine PCR among those with and without urethral symptoms was 16.7% and 2.4%, respectively. Fifty-one percent met criteria for PTSRB during the preceding month. Those reporting PTSRB were more likely to report multiple partners. In a multiple logistic regression model, the following were significant (p < 0.05) predictors of PTSRB: drug or alcohol use during sex; white race; only male partners, and sildenafil use. Drug use during sex was associated both with more sex partners and more sexual risk behaviors. Always disclosing HIV status was associated with fewer partners. There was a high prevalence of PTSRB among HIV-infected males entering care. Men who have sex with men (MSM), white race, drug/alcohol use during sex, and sildenafil use were independent risk factors. PTSRB was associated with having multiple partners. Physicians should discuss risk behaviors before prescribing sildenafil. PMID- 15294087 TI - Guidelines updated. PMID- 15294086 TI - Adherence to antiretroviral therapy in HIV-infected pediatric patients improves with home-based intensive nursing intervention. AB - Adherence to combination antiretroviral therapy (ART) has been shown to be a determining factor in controlling viral replication, maintaining immunologic function and long-term survival in HIV-positive individuals. Little information is available on strategies to improve adherence in pediatric HIV-infected patients. We conducted a randomized, nonblinded, pilot study to determine if a home-based nursing intervention would improve medication adherence. The study was offered to all eligible HIV-positive patients receiving care at Connecticut Children's Medical Center's (CCMC) Pediatric and Youth HIV Program. Sixty-seven percent (37/55) of the patients and their caretakers participated. We randomized participants to either standard of care or the intervention trial. The intervention was designed to improve knowledge and understanding of HIV infection and HIV medications and to resolve or modify barriers to adherence. Both groups completed pre- and post-intervention questionnaires, assessing their knowledge and understanding of HIV, ART, and adherence. Adherence was estimated objectively from medication refill history and subjectively from a self-report score. We also inferred adherence from pre- to post-test plasma viral load and CD4+ T-cell percentages. The knowledge score (p = 0.02) and medication refill history (p = 0.002) improved significantly in the intervention group. The adherence self report score improved, although not significantly (p = 0.07). We did not observe statistical differences in CD4+ T-cell counts or viral load between groups. We conclude that our home-based nursing intervention helped HIV-positive children and their families in better adhering to prescribed medication regimens. PMID- 15294088 TI - Asian language materials available. PMID- 15294089 TI - Endoplasmic reticulum resident, immunoglobulin joining chain, can be secreted by perturbation of the calcium concentration in the endoplasmic reticulum. AB - We established a transient human joining (J)-chain gene expression system in the baby hamster kidney (BHK) cell. The J-chain was detected as a 29-kDa single band on Western blotting. Immunofluorescent staining of the transfectant revealed an exclusive localization of the J-chain in the endoplasmic reticulum (ER). Intracellular transport experiment revealed that incubating conditions favorable for vesicular stomatitis virus glycoprotein (VSV-G) transport did not allow the J chain to exit from the ER. Analysis of glycosylation status of the J-chain in the transfectant was examined by tunicamycin treatment, endoglycosidase H digestion, and also by treatment with brefeldin A. It was found that an N-glycosylation consensus site of the J-chain was functional, and intracellular J-chain was endoglycosidase H sensitive. These results indicate that, in the absence of any immunoglobulin molecules, J-chain localizes exclusively in the ER. We also tested whether the J-chain could be exported from the ER by perturbing the Ca2+ concentration in the ER. Cultivation of the J-chain transfectant in the presence of ionomycin resulted in the time-dependent secretion of the J-chain. The secreted J-chain was modified by the Golgi resident glycosylation enzymes, indicating that the secreted J-chain passed through the normal exocytic pathway. PMID- 15294090 TI - IL-5-Induced Eosinophils Suppress the Growth of Leishmania amazonensis In Vivo and Kill Promastigotes In Vitro in Response to Either IL-4 or IFN-gamma. AB - In IL-5 transgenic mice (C3H/HeN-TgN(IL-5)-Imeg), in which 50% of peripheral blood leukocytes are eosinophils, the development of infection by Leishmania amazonensis was clearly suppressed. To determine mechanistically how this protozoan parasite is killed, we performed in vitro killing experiments. Either IL-4 or IFN-gamma effectively stimulated eosinophils to kill Leishmania amazonensis promastigotes, and most of the killing was inhibited by catalase but not by the NO inhibitor L-N5-(1-iminoethyl)-ornithine, suggesting that hydrogen peroxide is responsible for the killing of L. amazonensis by eosinophils. There was no significant degranulation of eosinophils in the culture, because eosinophil peroxidase was not detected in culture supernatants when L. amazonensis promastigotes were killed by activated eosinophils. Such resistance was also observed in BALB/c mice, which are highly susceptible to L. amazonensis. Expression plasmids for IL-4, IL-5, and IFN-gamma were transferred into muscle by electroporation in vivo starting 1 week before infection. Expression plasmid for IL-5 was most effective in slowing the development of infection among three expression plasmids. Expression plasmid for IL-4 was slightly effective and that for IFN-gamma had no effect on the progress of disease. These results suggest that IL-5 gene transfer into muscle by electroporation is useful as a supplementary protection method against L. amazonensis infection. PMID- 15294091 TI - A mutation found in the promoter region of the human survivin gene is correlated to overexpression of survivin in cancer cells. AB - Survivin, a unique antiapoptotic factor, plays an important role in cell cycle regulation. Numerous clinical studies have shown that survivin is markedly overexpressed in most common types of cancer, suggesting that transcriptional deregulation is a major mechanism involved in aberrant expression of survivin in cancers. In this study, we have identified several polymorphisms in the survivin gene promoter. One of these polymorphisms is located at CDE/CHR repressor elements and appears to be a common mutation with high frequency among cancer cell lines compared to normal cell line controls. The presence of the mutation was correlated in these cell lines with increased survivin expression at the both mRNA and protein levels. Furthermore, gel mobility shift analysis and transcriptional analysis showed the mutation changed cell cycle-dependent transcription by modifying the binding motif of the CDE/CHR repressor. These results indicate that the high level of survivin in some cancers is, at least in part, due to a genetic defect in the promoter region of the human survivin gene, which causes derepression of survivin transcription apparently due to the mutated CDE/CHR repressor binding motifs. PMID- 15294092 TI - Effects of transforming growth factor-alpha (TGF-alpha) in vitro and in vivo on reovirus replication. AB - We have utilized growth factors in in vitro and in vivo systems to examine the role of cellular proliferation in reovirus replication. In vitro, proliferating RIE-1 cells can be infected with whole reovirus virions, but are relatively resistant to infection once confluent (Go arrest). It has been shown that TGF alpha, which signals through the EGF-receptor (EGF-R), is capable of dramatically increasing the number of RIE-1 cells entering the S-phase in the presence of additional serum factors. Stimulation of the EGF-R without serum results in minimal increases in cells entering the S-phase with a restriction in reovirus replication. Therefore, other factors in serum are essential for fully permissive infection. In vivo, we used metallothionein (MT) promoter/enhancer-TGF-alpha transgenic mice to study the effect of cytokine activation on reovirus type 1 infection. Virus replication decreased following oral infection in these transgenic mice at 1 month of age, concordant with increased mucin production. Titers of reovirus obtained from the livers of 1 year old transgenic mice were approximately 10-fold higher than titers obtained in control mice. Taken together, these data indicate that while growth factor activation ultimately leads to an increase in virus infectivity, other factors may be necessary for reovirus replication. PMID- 15294093 TI - Dominant negative effect of novel mutations in pyruvate kinase-M2. AB - The natural mutations observed in pyruvate kinase (PK)-M2, a homotetramer isozyme, in this study correlated with the differential activity of the enzyme in a dominant negative manner in B-lymphoblastoid cells, established from two Bloom syndrome (BS) patients, BS1 and BS3 by 50 and 90%, respectively; and by 75% in the freshly obtained PHA stimulated lymphocytes of a BS patient diagnosed for the first time in India. A gene screen involving the critical domains of the PK-M gene in BS cells resulted in the observation of a missense mutation in BS1 and the BS patient and a frame shift mutation in BS3, in exon-10, coding for the intersubunit contact domain (ISCD) of the PK-M2 protein. Apart from these mutations, other variations in this region of the gene, both in normal and BS cells, did not affect the enzyme activity, since these were silent. Computer based modeling studies of the PK-M2 protein with each mutation was suggestive of a changed interaction between two domains within a subunit in BS1, a gross structural change in BS3, and a changed interaction between two subunits of the tetramer in the BS patient. An absence of such mutations in other regions of the PK-M2 gene in normal subjects and in the randomly chosen unrelated genes in the DNA from BS cell lines and the cells from the BS patient, authenticated the presence of the observed mutations in Bloom syndrome cells. A correlation observed between the differential enzyme activity and the nature of mutation in the intersubunit contact domain (ISCD) region of the PK-M2 gene was interesting, and indicted how the site and the nature of mutation in a heterozygous state could influence the enzyme activity differentially and in a dominant negative manner. The importance of these mutations in Bloom syndrome cells, however, remains to be elucidated, and can only be conjectured. PMID- 15294094 TI - Combined DNA vaccines formulated in DDA enhance protective immunity against tuberculosis. AB - This study evaluated the adjuvant Dimethyldioctyldecyl Ammonium Bromide (DDA) effect on the protective immunity induced by a combination of plasmids containing genes encoding antigens Ag85B, MPT-83, and ESAT-6 from Mycobacterium tuberculosis. The combined DNA vaccines in DDA resulted in significant increases in both specific IgG and splenic T-cell-derived Th1-type cytokine gamma interferon (IFN-gamma) production in response to the three purified antigens when compared to that of combined DNA vaccines in saline. Vaccines in DDA increased the protective efficacy of mice challenged with M. tuberculosis H37Rv as measured by reduced relative CFU counts in their lungs. Mice immunized with the combined DNA vaccines were shown to limit the growth of tubercle bacilli both in lungs and in spleens. Histopathological analyses showed that vaccinated mice had substantially improved postinfection lung pathology relative to the controls. We suggest that our combination of antigens together with DDA formulation may provide a new insight into tuberculosis prevention. PMID- 15294095 TI - Selection of internalizing ligand-display phage using rolling circle amplification for phage recovery. AB - Selection of phage libraries against complex living targets such as whole cells or organs can yield valuable targeting ligands without prior knowledge of the targeted receptor. Our previous studies have shown that noninfective multivalent ligand display phagemids internalize into mammalian cells more efficiently than their monovalent counterparts suggesting that cell-based selection of internalizing ligands might be improved using multivalently displayed peptides, antibodies or cDNAs. However, alternative methods of phage recovery are needed to select phage from noninfective libraries. To this end, we reasoned that rolling circle amplification (RCA) of phage DNA could be used to recover noninfective phage. In feasibility studies, we obtained up to 1.5 million-fold enrichment of internalizing EGF-targeted phage using RCA. When RCA was applied to a large random peptide library, eight distinct human prostate carcinoma cell internalizing peptides were isolated within three selection rounds. These data establish RCA as an alternative to infection for phage recovery that can be used to identify peptides from noninfective phage display libraries or infective libraries under conditions where there is the potential for loss of phage infectivity. PMID- 15294096 TI - Effects of lipopolysaccharide on actin reorganization and actin pools in endothelial cells. AB - OBJECTIVE: To investigate the dose and time-dependent effects of lipopolysaccharide (LPS) on cytoskeletal F-acitn and G-actin reorganizations by visualizing their distribution and measuring their contents in human umbilical vein endothelial cell line ECV-304. METHODS: F-actin was labeled with rhodamine phalloidin and G-actin with deoxyribonuclease I (DNase I)conjugated with fluorescein isothiocyanate (FITC). Contents of cytoskeletal proteins were obtained by flow cytometry. RESULTS: F-actin was mainly distributed peripherally in endothelial cells under normal conditions. LPS stimulation caused the formation of stress fibers and filopodia. G-actin was normally seen in perinuclear and nuclear areas in control ECV-304 cells. Under LPS stimulation, G actin dots appeared in the cytoplasmic region. The actin disorganization was accompanied by the time- and dose- dependent decrease in F-actin pool and increase in G-actin pool. CONCLUSIONS: LPS can induce characteristic morphological alterations of actin cytoskeleton and formation of intercellular gap in endothelial cells, accompanied by changes in F-actin and G-actin pools. PMID- 15294097 TI - Effect of cefazolin loaded bone matrix gelatin on repairing large segmental bone defects and preventing infection. AB - OBJECTIVE: To explore the possibility of repairing long segmental bone defects and preventing infection with cefazolin loaded bone matrix gelatin (C-BMG). METHODS: C-BMG was made from putting cefazolin into BMG by vacuum absorption and lyophilization techniques. The sustaining period of effective drug concentration in vitro and in vivo was detected. The time of inhibiting bacteria, and the drug concentration in local tissues (bone and muscle) and plasma after implantation of C-BMG were examined by high performance liquid chromatography. RESULTS: The effective inhibition time to staphylococcus aureus of C-BMG was 22 days in vitro; while 14 days in vivo. The cefazolin concentration in local tissues was higher in early stage, and later it kept a stable and low drug release. C-BMG showed an excellent ability to repair segmental long bone defects. CONCLUSIONS: C-BMG can gradually release cefazolin with effective drug concentration and has excellent ability to repair segmental bone defects. It can be used to repair segmental long bone defects and prevent infection after operation. PMID- 15294098 TI - Morphological changes of roof of subacromial bursa in patients with rotator cuff tear. AB - OBJECTIVE: To investigate the morphological changes of the roof of the subacromial bursa (SAB) and its involvement extent after rotator cuff tear. METHODS: In the experimental group, the roof of SAB was obtained from 30 cases of rotator cuff tear both at the tear site and a site 2.5-3.0 cm distal to the tear site during rotator cuff repair. In the control group, the roof of SAB was obtained from the exposed site of recurrently dislocated shoulder or fractured humeral shaft of 8 cases. The specimens were stained with hematoxylin and eosin and observed under a transmission electron microscope. The cell number was quantitated through counting the blue-stained nucleus in SAB with a computer image analysis system. RESULTS: The number of cells increased significantly in the roof of SAB in the experimental group compared with that of the control group. However, no difference of the bursal reaction was found among the type of rotator cuff tear, the bursa thickness and the presence of fluid in the bursa. The great majority of cells were type B cells observed under the transmission electron microscope. CONCLUSIONS: The increase in cell number in the roof of SAB in the experimental group is a reactive increase rather than an inflammatory process and the involvement of SAB is not limited in extent. The change of the roof of SAB is a secondary reaction to the rotator cuff tear. PMID- 15294099 TI - Posterolateral dislocation of the knee joints: analysis of 9 cases. AB - OBJECTIVE: To analyze the traumatic pathological characteristics of posterolateral dislocation of knee joints and its treatment. METHODS: Nine cases of posterolateral dislocation of knee joint, 5 cases of fresh injuries (the fresh injury group) and 4 cases of old injuries ( the old injury group) were reviewed and analyzed. In the fresh injury group 4 cases failed in close reduction due to "buttonholing" through the medial joint, among them 3 cases underwent repair of the damaged ligaments. In the old injury group 2 cases underwent ACL and MCL repair only in acute stage, but re-dislocated. Of the rest 2 cases 1 was associated with peroneal nerve injury and the other was not treated in acute stage. One case was associated with comminuted fracture of the tibial condyle and popliteal artery injury. Open reduction was performed in 3 cases. One case was fixed with 2-crossed pin and another was fixed with one pin through the tibial and femoral condyle and second pin with olecranization fixation. Plaster immobilization for 6-8 weeks respectively was required. In the old injury group in 1 case ACL and PCL repair (Augustine method) and posterolateral structure were performed and olecranization fixation and plaster immobilization for 6 weeks was needed. Arthrodesis of the knee was done for the patient with comminuted fracture of the tibial condyle and popliteal artery injury. RESULTS: All the cases were followed up for 1-23 years (average 6 years). Knee stability in 4 cases with repair of the ligaments was improved, although PDT showed (+) with different degrees. The results of the patients treated with ligamentous reconstruction were much better than those of the patients without any repair. CONCLUSIONS: Well understanding of the traumatic pathological characteristics, repair of the damaged ligaments, augmentation of olecranization fixation and postoperative immobilization for 6 weeks are the key points of successful treatment. PMID- 15294100 TI - Coculture of elongated neuron axon with poly (D,L-lactide-co-glycolide) biomembrane in vitro. AB - OBJECTIVE: To elongate human nerve axon in culture and search for suitable support matrices for peripheral nervous system transplantation. METHODS: Human embryo cortical neuronal cells, seeded on poly (D,L-lactide-co-glycolide) (PLGA) membrane scaffolds, were elongated with a self-made neuro-axon extending device. The growth and morphological changes of neuron axons were observed to measure axolemmal permeability after elongation. Neurofilament protein was stained by immunohistochemical technique. RESULTS: Human embryo neuron axon could be elongated and cultured on the PLGA membrane and retain their normal form and function. CONCLUSIONS: Three dimensional scaffolds with elongated neuron axon have the basic characteristics of artificial nerves, indicating a fundemental theory of nerve repair with elongated neuron axon. PMID- 15294101 TI - Schwann cell apoptosis in Wallerian-degenerated sciatic nerve of the rat. AB - OBJECTIVE: To investigate systematically Schwann cell apoptosis in Wallerian degenerated sciatic nerve of the rat, and evaluate its time-related feature. METHODS: Ninety-five SD rats were divided randomly into one normal group (8 rats) and 11 experimental groups (66 rats, 6 in each). Both hind legs of each rat in experimental groups were randomly divided into test leg (sciatic nerve transected) and control one (nerve uninjured). All test legs constituted a test group and all control legs constituted a control one. After operation, all rats were respectively sacrificed at 1 h, 6 h, 12 h, 24 h, 2 d, 3 d, 4 d, 8 d, 14 d, 21 d, and 30 d. We analyzed the specimens of mid-distal sciatic nerve, especially the morphological changes of the nerve, the different expression levels of S-100 protein and apoptosis-related proteins such as Bcl-2, Bax, and Fas in Schwann cells. The TUNEL method was used to detect the apoptotic rate of Schwann cells. RESULTS: (1) The test group showed Wallerian degeneration. The number of Schwann cells began to decrease at 24 h, obviously decreased on day 3 and 4, then began to increase from day 8 and formed Bungner belt after 14 days. (2) Schwann cells generally expressed S-100 at a low level in all groups. The control group was not significantly different from the normal group. The test group had statistical significance at 1 h and day 21. (3) As an inhibitory gene protein of Schwann cell apoptosis, Bcl-2 positive rates in the control and test groups apparently elevated and were statistically different from the normal group. (4) As a promotive gene protein of Schwann cell apoptosis, the control and test groups expressed Bax at a high level and were statistically different from the normal group. (5) As a promotive gene protein of Schwann cell apoptosis, Fas positive rate in control group was slightly elevated, but had no statistical significance compared with the normal group. Fas positive rate in test group continuously elevated in a fluctuant way, with highly statistical significance compared with the normal group. (6) TUNEL detection further proved that Schwann cell apoptosis rarely existed in the normal group, and the left sciatic nerve had no statistical significance compared with the right sciatic nerve. While the test group showed lots of apoptotic nuclei at 6 h, 2 d, 4 d, and 21 d. It had highly statistical significance compared with the normal group. CONCLUSIONS: Schwann cell apoptosis does exist in Wallerian-degenerated sciatic nerve of the rat after transection. Schwann cell apoptosis and its apoptotic genes expression have a time-related feature. PMID- 15294102 TI - Changes of free iron contents and its correlation with lipid peroxidation after experimental spinal cord injury. AB - OBJECTIVE: To observe the dynamic changes of free iron contents and its relationship to the changes of lipid peroxidation after experimental spinal cord injury (SCI). METHODS: Sprague Dawley rats were randomly divided into three groups: Group A (n=6) received no operation; Group B (n=48) received only laminectomy (sham); and Group C (n=48) received both laminectomy and traumatic injury (SCI model). The SCI animal models were made by using an modified Allen's weight-drop device (50 g.cm) on T(12). Rats were sacrificed at 0.5, 1, 3, 6, 12, 24 hours after injury. The levels of free iron involved in spinal cord segments at different time points were measured by bleomycin assay. The malondialdehyde (MDA) was also measured by the thiobarbituric acid (TBA). RESULTS: After SCI in Group C, the level of free iron showed a significant increase at 0.5 hour compared to Groups B and A, restored to the control level at 6 h; the level of MDA was increased at 0.5 hour, peaked at 3 hours, returned to the control level at 12 hours; the concentrations of free iron and lipid peroxidation in injured rats were significantly and positively correlated at 0.5-3 hours. CONCLUSIONS: After SCI the levels of free iron are increased quickly and might be a major contributor to lipid peroxidation in injured spinal cord. PMID- 15294103 TI - Changes in metabotropic glutamate receptor 4 expression and the effects of L-2 amino-4-phosphonobutyrate in a rodent model of diffuse brain injury. AB - OBJECTIVE: To examine the changes in the expression of mGluR4 after diffuse brain injury (DBI) and to determine the role of its specific agonist L-2-amino-4 phosphonobutyrate (L-AP4) in vivo. METHODS: A total of 161 male SD rats were randomized into the following groups. Group A included normal control, sham operated control and DBI group. DBI was produced according to Marmarou's diffuse head injury model. mRNA expression of mGluR4 was detected by hybridization in situ. Group B included DBI alone, DBI treated with normal saline and DBI treated with L-AP4. All DBI rats were trained in a series of performance tests, following which they were subjected to DBI. At 1 and 12 hours, animals were injected intraventricularly with L-AP4 (100 mmol/L, 10 microl) or normal saline. Motor and cognitive performances were tested at 1, 3, 7, 14 days after injury and the damaged neurons were also detected. RESULTS: There was no significant difference between normal control group and sham-operated group in the expression of mGluR4 (P>0.05). The animals exposed to DBI showed significantly increased expression of mRNA of mGluR4 compared with the sham-operated animals 1 h after injury (P<0.05). At 6 hours, the evolution of neuronal expression of mGluR4 in the trauma alone group was relatively static. Compared with saline-treated control animals, rats treated with L-AP4 showed an effective result of decreased number of damaged neurons and better motor and cognitive performances. CONCLUSIONS: Increased expression of mGluR4 is important in the pathophysiological process of DBI and its specific agonist L-AP4 can provide remarkable neuroprotection against DBI not only at the histopathological level but also in the motor and cognitive performance. PMID- 15294104 TI - Effect of polydatin on phospholipase A2 in lung tissues in rats with endotoxic shock. AB - OBJECTIVE: To study the effect of polydatin on phospholipase A(2) in lung tissues in rats with endotoxic shock. METHODS: Thirty-two healthy male Wistar rats were employed in this study. A total of 8 rats received normal saline intravenously (control group), 8 rats received 10 mg/kg of endotoxin (endotoxic shock group), 8 rats received 1 mg/kg of polydatin after endotoxin injection (polydatin treatment group), and 8 rats received 1 mg/kg of polydatin (polydatin prevention group) 30 minutes before endotoxin injection. Mean arterial pressure was measured once half an hour. Lung tissues were collected 6 hours later. Phospholipase A(2) activity was measured with acid titration. The gene expression of secretory phospholipase A(2) type IIA was detected with reverse transcription polymerase chain reaction. Meanwhile, the histological changes of the lungs among four groups were compared through microscopic examination. RESULTS: Phospholipase A(2) activity and the gene expression of secretory phospholipase A(2) type IIA increased after endotoxin injection, but polydatin could inhibit these effects of endotoxin. Obvious morphological evidence could be found in the lung pathological sections and the protective effect of polydatin was most significant in the polydatin prevention group. CONCLUSIONS: Polydatin has prophylactic and therapeutic effects (the former is more distinct than the latter) on acutely injured lungs in rats with endotoxic shock and which suggests that polydatin may be a phospholipase A(2) inhibitor. PMID- 15294105 TI - Management of femoral artery pseudoaneurysm due to addictive drug injection. AB - OBJECTIVE: To study surgical management for patients with femoral pseudoaneurysm resulting from addictive drug injection. METHODS: Clinical data of 34 patients with femoral pseudoaneurysm resulting from addictive drug injection were retrospectively reviewed. RESULTS: Thirteen patients underwent bypass graft (end to side) of external iliac artery and superficial femoral artery using expanded polytetrafluoroethylene (ePTFE). Three patients who had an autogenous saphenous vein graft in situs, one of whom was then performed an ePTFE graft when rupture and bleeding occurred at the anastomotic site. Color Doppler image showed patent grafted blood vessels in all the patients after operation. Eighteen patients had their femoral arteries ligated. Limbs of all the 34 patients were saved. CONCLUSIONS: Ligating femoral artery is an effective way to treat femoral artery pseudoaneurysm if autogenous saphenous vein graft or artificial vessel graft is not applicable. PMID- 15294106 TI - Post-traumatic osteolysis of the distal clavicle, pubis and ischium in 7 patients. AB - Post-traumatic osteolysis (PTOL) is a very rare disease occurring after acute trauma or repetitive micro-trauma, which is characterized by persistent pain in the injured site. In this study, we reported 7 patients, in whom osteolysis developed in the distal clavicle, pubis and ischium. PMID- 15294107 TI - Dural arteriovenous fistulae after forehead knife-cut injury. AB - Dural arteriovenous fistulae (DAVF) is a rare intracranial vascular disease. It is pathologically characterized by direct shunting of the intracranial artery and vein, which results in cerebral ischemia, intracranial hemorrhage, neural deficit and intracranial murmur. The etiological mechanism of DAVF is not well known, but most researchers think it is associated with congenital abnormal development, especially abnormal development of dural blood vessels at the stage of embryogenesis. Recently, some researchers have found that DAVF is also associated with some acquired factors. This article reports a case who developed DAVF within 2 years after debridement of frontal bone fragmentation, depressed fracture, left frontal lobe contusion and superior sagittal sinus injury due to forehead knife cut injury. The pathogenic mechanism was explored through a review of the related literatures. PMID- 15294108 TI - Change and role of heme oxygenase-1 in injured lungs following limb ischemia/reperfusion in rats. AB - OBJECTIVE: To study the change and role of heme oxygenase-1 (HO-1) in injured lungs following limb ischemia/reperfusion in rats. METHODS: A total of 96 healthy male Sprague-Dawley rats, weighing 250-300 g, were used in this study. Hind limb ischemia was made on 40 rats through clamping the infrarenal aorta for 2 hours with a microvascular clip, then limb reperfusion for 0, 4, 8, 16 and 24 hours (n=8 in each time point) was performed, respectively. Other 8 rats undergoing full surgical operation including isolation of the infrarenal aorta without occlusion were taken as the sham operation group. Lung tissues were obtained from the 48 animals and Northern blotting and Western blotting were employed to measure the changes of HO-1 mRNA and protein expression, respectively. Immunohistochemistry technique was used to determine the cell types responsible for HO-1 expression after limb ischemia/reperfusion. Then hind limb ischemia was made on other 12 rats through clamping the infrarenal aorta for 2 hours with a microvascular clip, among whom, 6 rats were given zinc protoporphyrin (ZnPP), an inhibitor of HO. Then limb reperfusion for 16 hours was performed on all the 12 rats. And other 12 rats underwent full surgical operation including isolation of the infrarenal aorta without occlusion, among whom, 6 rats were then given ZnPP. Then lung tissues were obtained from the 24 animals and lung injury markers, lung histology, polymorphonuclear leukocyte (PMN) count and malondialdehyde (MDA) content were detected, respectively. HO activity was determined through measuring the carboxyhemoglobin (COHb) level in artery blood with a CO-oximeter after limb ischemia/reperfusion. And the animal mortality was observed on the other 24 rats. RESULTS: Northern blotting analysis showed that HO-1 mRNA increased significantly at 4 hours after reperfusion, peaked at 16 hours, and began to decrease at 24 hours. In contrast, no positive signal was observed in the sham and simple ischemia animals. Increased HO-1 mRNA levels were accompanied by similar increases in HO-1 protein. Lung PMNs and MDA content increased significantly at 4, 8, 16 and 24 hours after reperfusion, compared with the sham controls (P<0.001), while they decreased in rats with reperfusion for 16 hours when compared with rats with reperfusion for 4 hours (P<0.001). Immunohistochemical studies showed that HO-1 was expressed in a variety of cell types, including the airway epithelia, alveolar macrophages and vascular smooth muscular cells. The blood COHb level and animal mortality increased significantly after limb ischemia/reperfusion compared with the sham controls (P<0.001). ZnPP administrated to the ischemia/reperfusion animals led to a decrease in the COHb level and an increase in lung PMN number, MDA content and animal mortality (P<0.001 compared with ischemia/reperfusion group), and the lung injury was aggravated. CONCLUSIONS: Limb ischemia/reperfusion up-regulates pulmonary HO-1 expression, which serves as a compensatory protective response to the ischemia/reperfusion-induced lung injury in rats. PMID- 15294109 TI - Analysis of prosthetic replacement in treatment of femoral neck fracture on the hemiplegia side in the elderly. AB - OBJECTIVE: To study the effect of prosthetic replacement in treatment of femoral neck fractures on the hemiplegia side in the elderly. METHODS: From May 1990 to May 2000, 189 elderly patients with femoral neck fractures were treated with prosthetic replacement in my hospital. Twenty-nine hemiplegia patients, who suffered from stroke previously, had Garden type III and type IV femoral neck fractures on the hemiplegia side. Thirty non-hemiplegia patients were chosen randomly. The two groups were followed-up for 27-98 months (average: 59 months). The age, hospitalization days, operating time, blood loss, blood transfusion, complications during perioperative period and long-term complications were compared between the two groups and the results of femoral head replacement and total hip replacement in the hemiplegia group were also compared. RESULTS: All the patients of the two groups survived the perioperative period. No significant difference was found in the age, hospitalization days, operation time, blood loss and blood transfusion and long-term complications between the two groups (P>0.05). However there was significant difference in complications during perioperative period between the two groups (P<0.05). Five patients died in the hemiplegia group with the mortality of 17.2% and two died in the non-hemiplegia group with the mortality of 6.7% 11 months to 5 years after operation. There was significant difference in long-term complications between the femoral head replacement and the total hip replacement in the hemiplegia group (P<0.05). The result of the total hip replacement was better than that of the femoral head replacement. CONCLUSIONS: Prosthetic replacement is a reliable method in treatment of Garden type III and type IV femoral neck fractures on the hemiplegia side in the elderly, and patients are safe during perioperative period. More complications during perioperative period occur in the hemiplegia group, and long term complications are insignificantly different between the two groups. The mortality rate is higher in the hemiplegia group than in the non-hemiplegia group within 5 years after operation. Since the result of the total hip replacement is better than that of the femoral head replacement, total hip replacement should be chosen firstly to treat Garden type III and type IV femoral neck fractures on the hemiplegia side in the elderly if the muscular strength of the hip is beyond IV degree. PMID- 15294110 TI - Effect of puerarin on neural function and histopathological damages after transient spinal cord ischemia in rabbits. AB - OBJECTIVE: To investigate the effect of puerarin on the neural function and the histopathological changes after ischemic spinal cord injury in rabbits. METHODS: Thirty male New Zealand white rabbits were randomly divided into three groups as follows: puerarin group (n=10) receiving intravenous infusion of 30 mg/kg puerarin for 10 minutes, control group (n=10) receiving intravenous infusion of the same volume of normal saline as puerarin for 10 minutes, and sham operation group (n=10) undergoing only the surgical exposure of the abdominal aorta. Temporary spinal cord ischemia was induced by infrarenal aortic occlusion for 20 minutes and followed by reperfusion. The neural status was scored with the Tarlov criteria at 8, 12, 24 and 48 hours after reperfusion. All the animals were killed at 48 hours after reperfusion and the spinal cords (L5) were removed immediately for histopathological study. RESULTS: The neural function scores at 8, 12, 24 and 48 hours after reperfusion were higher in the puerarin group and sham operation group than those in the control group (P<0.05). More normal motor neurons in the anterior horn of spinal cord were present in the puerarin group and sham operation group than those in the control group (P<0.01). There was a strong correlation between the final neural function scores and the number of normal motor neurons in the anterior horn of spinal cord (r=0.839, P<0.01). CONCLUSIONS: Puerarin can significantly ameliorate the neural function and the histopathological damages after transient spinal cord ischemia in rabbits. PMID- 15294111 TI - Effects of magnesium sulfate on traumatic brain edema in rats. AB - OBJECTIVE: To investigate the effects of magnesium sulfate on traumatic brain edema and explore its possible mechanism. METHODS: Forty-eight Sprague-Dawley (SD) rats were randomly divided into three groups: Control, Trauma and Treatment groups. In Treatment group, magnesium sulfate was intraperitoneally administered immediately after the induction of brain trauma. At 24 h after trauma, total tissue water content and Na(+), K(+), Ca(2+), Mg(2+) contents were measured. Permeability of blood-brain barrier (BBB) was assessed quantitatively by Evans Blue (EB) dye technique. The pathological changes were also studied. RESULTS: Water, Na(+), Ca(2+) and EB contents in Treatment group were significantly lower than those in Trauma group (P<0.05). Results of light microscopy and electron microscopy confirmed that magnesium sulfate can attenuate traumatic brain injury and relieve BBB injury. CONCLUSIONS: Treatment with MgSO4 in the early stage can attenuate traumatic brain edema and prevent BBB injury. PMID- 15294112 TI - Clinical application of axonal repair technique for treatment of peripheral nerve injury. AB - OBJECTIVE: To evaluate the efficacy of axonal repair technique for treatment of peripheral nerve injury clinically. METHODS: In 1998, the authors applied axonal repair technique to treat peripheral nerve injuries in 12 patients with 13 nerves. It consists of four steps, ie, stumps of the nerve being soaked in a modified Collins fluid, freezed, trimmed, and coapted with glue, making the injured nerve repaired at the axonal level. RESULTS: The patients were followed up for an average of 13 months. Results showed that in 4 cases of first-stage contralateral C7 transfer, regenerating axons reached to the sternoclavicular joint or axilla at 4 to 7 months, offering the timing for performing the second stage contralateral C7 transfer. In 5 cases of accessory nerve transferred to the suprascapular nerve, the abduction of the shoulder was 40 degree on average. In the other 3 patients with four different nerves repaired, results were also satisfactory. CONCLUSIONS: This technique is promising in the treatment of peripheral nerve injury. PMID- 15294113 TI - S-100B and neuron specific enolase in outcome prediction of severe head injury. AB - OBJECTIVE: To elucidate the role of S-100B and neuron specific enolase (NSE) in predicting the outcomes of patients with severe head injury. METHODS: Forty patients with severe head injury were included in this study. The serum concentrations of S-100B and NSE were measured within 12 hours after head injury to investigate the correlation between serum levels of S-100B and NSE and outcome. Validity of both S-100B and NSE in outcome prediction was assessed with Receiver Operator Characteristic (ROC) curve. RESULTS: The serum concentrations of S-100B and NSE of both groups, with favorable or unfavorable outcomes, were significantly higher than those of the normal group. The serum concentrations within 12 hours after head injury were closely correlated with the prognosis. Furthermore, according to the ROC curves of S-100B and NSE, S-100B was found better in predicting outcomes than NSE. CONCLUSIONS: S-100B and NSE may play important roles in outcome prediction after severe head injury. Moreover, S-100B is clearly superior to NSE in terms of predictive value and appears to be a more promising serum marker in outcome prediction after severe head injury. PMID- 15294114 TI - Human neuronal apoptosis secondary to traumatic brain injury and the regulative role of apoptosis-related genes. AB - OBJECTIVE: To observe human neuronal apoptosis secondary to traumatic brain injury, and to elucidate its regulative mechanism and the change of expression of apoptosis-related genes. METHODS: Specimens of brain were collected from cases of traumatic brain injury in humans. The histological and cellular morphology was examined by light and electron microscopy. The extent of DNA injury to cortical neurons was detected by using TUNEL. By in situ hybridisation and immunohistochemistry the mRNA changes and protein expression of Bcl-2, Bax, p53, and caspase 3 p20 subunit were observed. RESULTS: Apoptotic neurons appeared following traumatic brain injury, peaked at 24 hours and lasted for 7 days. In normal brain tissue activated caspase 3 was rare, but a short time after trauma it became activated. The activity peaked at 20-28 hours and remained higher than normal for 5-7 days. There was no expression of Bcl-2 mRNA and Bcl-2 protein in normal brain tissue but 8 hours after injury their expression became evident and then increased, peaked at 2-3 days and remained higher than normal for 5-7 days. The primary expression of Bax-mRNA and Bax protein was high in normal brain tissue. At 20-28 hours they increased and remained high for 2-3 days; on the 7th days they returned to a normal level. In normal brain tissue, p53mRNA and P53 were minimally expressed. Increased expression was detected at the 8th hour, and decreased at 20-28 hours but still remained higher than normal on the 5th day. CONCLUSIONS: Following traumatic injury to the human brain, apoptotic neurons appear around the focus of trauma. The mRNA and protein expression of Bcl-2, Bax and p53 and the activity of caspase 3 enzyme are increased. PMID- 15294115 TI - An anatomical study of corona mortis and its clinical significance. AB - OBJECTIVE: To provide detailed information of corona mortis for ilioinguinal approach as an anterior approach to the acetabulum and pelvis. METHODS: The course, branches and distribution of the vascular connection between the obturator system and the external iliac or inferior epigastric systems located over the superior pubic ramus were observed on 50 hemipelvises with intact soft tissues. RESULTS: During the dissections, 72% of the cadaveric sides had at least one communicating vessel between the obturator system and the external iliac or inferior epigastric systems on the superior pubic ramus. The average diameter of the connecting vessel was 2.6 mm (range, 2.0-4.2 mm). It coursed over the superior pubic ramus or iliopubic eminence vertically to enter the obturator foramen and exit the pelvis. The average distance from pubic symphysis to the vascular connections between the obturator and external iliac systems was 52 mm (range, 38-68 mm). CONCLUSIONS: Vascular connections between the obturator system and the external iliac or inferior epigastric systems were found over the superior pubic ramus with a high incidence. They are prone to damage during the ilioinguinal approach as an anterior approach to the acetabulum and pelvis. Thus, corona mortis located over the superior pubic ramus deserves great attention during the ilioinguinal approach. PMID- 15294116 TI - Subciliary incision and lateral cantholysis in rigid internal fixation of zygomatic complex fractures. AB - OBJECTIVE: To introduce the technique of subciliary incision and lateral cantholysis with tri-dimension reduction and rigid internal fixation to treat zygomatic complex fractures. METHODS: The subciliary incision and lateral cantholysis combined with tri-dimension reduction and rigid internal fixation of zygomatic complex fractures with titanium microplates were applied in 56 patients with zygomatic complex fractures. Another lateral eyebrow incision or sublabial incision was used to simplify the operation. RESULTS: The postoperative follow-up period ranged from 6 months to 5 years. During the follow-up period, all the patients had satisfying postoperative results. All clinical symptoms disappeared except the numbness in the infraorbital region in 2 patients. In 94.6% patients no complications such as obvious scar, ectropion, entropion or blepharoedema were found, only 5.4% of the patients had slight ectropion 6 months after operation. CONCLUSIONS: The subciliary incision and lateral cantholysis have many advantages such as invisible scar, sufficient exposure, minimal injury, and few complications and combined with rigid internal fixation with titanium microplates this technique could be used as one of the routine operation methods to treat zygomatic complex fractures. PMID- 15294117 TI - Management of posttraumatic brain swelling based on clinical typing. AB - OBJECTIVE: To investigate the clinical typing and prophylactico-therapeutic measures for acute posttraumatic brain swelling (BS). METHODS: A retrospective study was performed in 66 cases of acute posttraumatic BS. There were 3 groups based on computered tomography (CT) scanning: 23 cases of hemisphere brain swelling (HBS) with middle line shift for less than 5 mm within 24 hours (Group A), 20 with middle line shift for more than 5 mm (Group B), and 23 with bilateral diffuse brain swelling (Group C). RESULTS: (1) The mortality rates of the operative and nonoperative management in Group A, Group B, and Group C were 20.0%, 31.6%, and 75.0% versus 44.4%, 0, and 85.7%, respectively (P>0.05); while the rates in subgroups with different middle line shift (more than 5 mm and less or equal 5 mm) were 29.2% and 75.0% versus 75.0% and 44.4%, respectively (0.05>P>0.01). (2) The good recovery rate and mortality in Group A were 47.8% and 39.1%, respectively and in Group C, 8.7% and 78.3%, respectively. There was a very significant difference between Group A and Group C (P<0.01). (3) The total survival rate of the selective comprehensive therapy was 53.1%. CONCLUSIONS: (1) Acute posttraumatic BS needs to be diagnosed correctly and promptly with CT scanning within 4 hours. (2) For patients with midline shift for more than 5 mm, especially with thin-layered subdural hematoma, surgical intervention is essential to reduce the fatality of acute posttraumatic BS. PMID- 15294118 TI - Influence of cryopreserved olfactory ensheathing cells transplantation on axonal regeneration in spinal cord of adult rats. AB - OBJECTIVE: To observe the effects of cryopreserved olfactory ensheathing cells (OECs) transplantation on axonal regeneration and functional recovery following spinal cord injury in adult rats. METHODS: Twenty-four rats were divided into experimental and control groups, each group having 12 rats. The spinal cord injury was established by transecting the spinal cord at T10 level with microsurgery scissors. OECs were purified from SD rat olfactory bulb and cultured in DMEM (Dulbecco's minimum essential medium) and cryopreserved (-120 degree) for two weeks. OECs suspension [(1-1.4)x10(5)/ul] was transplanted into transected spinal cord, while the DMEM solution was injected instead in the control group. At 6 and 12 weeks after transplantation, the rats were evaluated with climbing test and MEP (moter evoked potentials) monitoring. The samples of spinal cord were procured and studied with histological and immunohistochemical stainings. RESULTS: At 6 weeks after transplantation, all of the rats in both transplanted and control groups were paraplegic, and MEPs could not be recorded. Morphology of transplanted OECs was normal, and OECs were interfused with host well. Axons could regrow into gap tissue between the spinal cords. Both OECs and regrown axons were immunoreactive for MBP. No regrown axons were found in the control group. At 12 weeks after transplantation, 2 rats (2/7) had lower extremities muscle contraction, 2 rats (2/7) had hip and/or knee active movement, and MEP of 5 rats (5/7) could be recorded in the calf in the transplantation group. None of the rats (7/7) in the control group had functional improvement, and none had MEPs recorded. In the transplanted group, histological and immunohistochemical methods showed the number of transplanted OECs reduced and some regrown axons had reached the end of transected spinal cord. However, no regrown axons could be seen except scar formation in the control group. CONCLUSIONS: Cryopreserved OECs could integrated with the host and promote regrowing axons across the transected spinal cord ends. PMID- 15294119 TI - Clinical analysis of craniocerebral trauma complicated with thoracoabdominal injuries in 2165 cases. AB - OBJECTIVE: To explore the optimal treatment for craniocerebral trauma complicated with thoraco-abdominal injuries. METHODS: A total of 2165 cases of craniocerebral trauma complicated with thoraco-abdominal injuries admitted to our hospital between July 1993 and June 2003 were retrospectively studied. Among them, 382 cases sustained severe craniocerebral trauma (in which 167 were complicated with shock), 733 thoracic injuries, 645 abdominal injuries and 787 thoraco-abdominal injuries. On admittance, 294 cases had developed shock. With the prime goal of saving life, respiratory and circulatory systems and encephalothilipsis were especially treated and monitored. Priority in management was directed to severe or open injures rather than to moderate or closed injures. For cases with cerebral hernia due to intracranial hematoma and severe shock due to blood loss, cerebral hernia and shock were treated concurrently. RESULTS: After treatment, 2024 (93.49%) cases survived and the other 141 (6.51%) died. Among patients who had severe craniocerebral injury with shock and those without, 78 (46.71%) and 53 (24.56%) died, respectively. For patients who had underwent craniocerebral and thoraco-abdominal operations concurrently and those who had not, the death rates were 58.49%-65.96% and 28.57% respectively, indicating a significant difference (P<0.05). CONCLUSIONS: Treatment for hematoma hernia, shock and disturbed respiration is the key in the management of multiple trauma of craniocerebral, thoracic or abdominal injuries, especially when two or three conditions occurred simultaneously. Unless it is necessary, operations at two different parts at the same time is not recommended. It is preferred to start two concurrent operations at different time. PMID- 15294120 TI - Traumatic subdural hydroma developing into chronic subdural hematoma. AB - OBJECTIVE: To probe the incidence, pathogenesis and clinical characteristics of traumatic subdural hydroma (TSH) developing into chronic subdural hematoma (CSDH). METHODS: We retrospectively analyzed the clinical data of 32 patients with TSH developing into CSDH and reviewed related literature. RESULTS: 16.7% of TSH developed into CSDH in this study. The time of evolution was from 22 to 100 days after head injury. All the patients were cured with hematoma drainage. CONCLUSIONS: TSH is one of the origins of CSDH. The clinical characteristics of TSH developing into CSDH follow that the ages of the patients are polarized, that the evolution often happens in the patients with small chronic hydromas and being treated conservatively, that the patients are usually injured deceleratedly and that the accompanying cerebral damage is often very mild. PMID- 15294121 TI - White blood cells and hemoglobin assay for the evaluation of multiple traumas. PMID- 15294122 TI - Road safety is no accident--for celebrating World Health Day 2004. PMID- 15294123 TI - Effects of phospholipase D on cardiopulmonary bypass-induced neutrophil priming. AB - OBJECTIVE: To investigate the relationship between phospholipase D (PLD) activation and neutrophil priming induced by cardiopulmonary bypass (CPB), and try to clarify whether CPB-induced systemic inflammatory response can be attenuated by inhibiting neutrophilic PLD activation. METHODS: Neutrophils were isolated from arterial blood of 8 patients undergoing valve replacement before operation and 30 min after initiation of CPB respectively. Both the preoperative and CPB-stirred neutrophils were subdivided into 5 groups by receiving different experimental interventions: (1) bacterial lipopolysaccharide (LPS, 10 ng x ml( 1)), (2) N-formylmethionylphenylalanine (fMLP, 1 micromol x L(-1)), (3) LPS+fMLP, (4) 1-butanol (0.5%)+LPS+fMLP, (5) vehicle. Elastase and myeloperoxidase (MPO) release was measured for the parameters of neutrophil activation, neutrophil PLD activity was determined by quantitation of choline produced from the stable product of phosphatidylcholine catalyzed by PLD. RESULTS: (1) Preoperative neutrophils treated with LPS+fMLP presented significantly higher PLD activity (13.48+/-2.61 nmol choline x h(-1) x mg(-1)) and released more elastase and MPO than cells treated with vehicle (PLD activity 3.70+/-0.49 nmol choline x h(-1) x mg(-1)), P<0.01), LPS (P<0.01) and fMLP respectively. In 1-butanol+LPS+fMLP group, PLD activity of preoperative neutrophils was lower than that in LPS+fMLP group (P<0.01), besides the release of elastase and MPO decreased sharply below both LPS+fMLP and fMLP groups (P<0.01). In LPS group, PLD activity was higher (P<0.01), while elastase and MPO release did not differ from control. fMLP group presented PLD activity, elastase and MPO release higher than control (P<0.01); nevertheless, lower than LPS+fMLP group (P<0.01). (2) CPB-stirred neutrophils presented prominent PLD activity increment, and even the control level was 3.59 fold of the pre-operative control (P<0.01). PLD activity in LPS+fMLP group was higher than that in other groups. Notably, PLD activity was even nonstatistically lower in 1-butanol+LPS+fMLP group than that in LPS or fMLP group. CPB-stirred neutrophils in LPS+fMLP group released more elastase and MPO than control, LPS, and 1-butanol+LPS+fMLP groups did (P<0.01); however, neither of the release was statistically different from that of fMLP group. CONCLUSIONS: Cardiopulmonary bypass enables neutrophil priming accompanied with significant increase in PLD activity. Inhibition of neutrophil PLD activation attenuates its priming and may alleviate CPB-induced systemic inflammatory reaction. PMID- 15294124 TI - Gene expression of collagen types IX and X in the lumbar disc. AB - OBJECTIVE: To study gene expression of collagen types IX and X in human lumbar intervertebral discs during aging and degeneration and to explore the role of collagen types IX and X in disc degeneration. METHODS: Fetal, adult and pathologic specimens were subjected to in situ hybridization with cDNA probes to investigate mRNA-expressions of types IX and X collagen gene. RESULTS: In fetal intervertebral discs, positive mRNA hybridization signals of type IX collagen were concentrated in the nucleus pulposus and the inner layer of anulus fibrosus. Interstitial matrix of the nucleus pulposus also showed positive type X collagen staining. Positive mRNA hybridization signals of types IX and X were not detected in the middle and outer layers of anulus fibrosus. In adult specimens, expression of type IX collagen mRNA was markedly decreased. No hybridization signals of type X collagen was observed. As for pathological specimens, there was no gene expression of type IX collagen. In severe degenerated discs from adults, there were focal positive expressions of type X collagen. CONCLUSIONS: Obvious changes of collagen gene expression occur with aging. Expression of type IX collagen decreases in adult and pathological discs. Results of type X collagen expression suggest that type X collagen is expressed only in older adult and senile discs (i.e., when disc degeneration has already reached a terminal stage), indicating the terminal stage of degeneration. PMID- 15294125 TI - Effect of nerve growth factor and Schwann cells on axon regeneration of distracted inferior alveolar nerve following mandibular lengthening. AB - OBJECTIVE: To study the effect of nerve growth factor (NGF) and Schwann cells on axon regeneration of the inferior alveolar nerve following mandibular lengthening with distraction osteogenesis. METHODS: Unilateral mandibular osteodistraction was performed in 9 healthy adult male goats with a distraction rate of 1 mm/d. Every 3 goats were killed on days 7, 14 and 28 after mandibular lengthening, respectively. The inferior alveolar nerves in the distraction callus were harvested and processed for ultrastructural and NGF immunohistochemical study. The inferior alveolar nerves from the contralateral side were used as controls. RESULTS: On day 7 after distraction, axon degeneration and Schwann cell proliferation were observed, and very strong staining of NGF in the distracted nerve was detected. On day 14 after distraction, axon regeneration and remyelination were easily observed, and NGF expression started to decline. On day 28 after distraction, the gray scale of NGF immunoreactivity recovered to the normal value and the Schwann cells almost recovered to their normal state. CONCLUSIONS: Gradual mandibular osteodistraction can result in mild or moderate axon degeneration of the inferior alveolar nerve. Nerve trauma may stimulate the proliferation of Schwann cells and promote the synthesis and secretion of NGF in the Schwann cells. Schwann cells and NGF might play important roles in axon regeneration of the injured inferior alveolar nerve following mandibular lengthening. PMID- 15294126 TI - Amino acids protects against renal ischemia-reperfusion injury and attenuates renal endothelin-1 disorder in rats. AB - OBJECTIVE: To investigate nephroprotective effects of a mixture of 8 L-amino acids on renal ischemia-reperfusion injury and its effects on renal endothelin-1 (ET-1). METHODS: The mixture of 8 L-amino acids includes glycine, alanine, threonine, serine, valine, leucine, isoleucine and proline. Acute ischemic renal injury was induced by clamping renal pedicle for 45 minutes in rats. Sixty male Sprague-Dawley rats were randomly divided into 3 groups: a sham-operated group (Group A, n=8), a control group (Group B, n=26) and an amino acid-treated group (Group C, n=26). Amino acids were infused at a rate of 1 ml x 100g(-1) x h(-1) I hour before ischemia and during 3 hours of the whole reperfusion. The serum creatinine values, BUN levels, creatinine clearance, urine sodium and potassium excretion, urine lactate dehydrogenase (LDH), the rate of urine flow and histological examination were measured. Renal ET-1 levels were assayed with radioimmunological assay (RIA) RESULTS: The creatinine clearance was 471.0 microl/min+/-121.5 microl/min in Group C and 227.0 microl/min+/-27.0 microl/min in Group B 3 hours after reperfusion, P<0.01). The urine flow rate was 63.6 microl/min+/-15.2 microl/min in Group C and 24.3 microl/min+/-7.7 microl/minin Group B, P<0.01) 1.5 hours after reperfusion. The serum creatinine was 85.0 microl/min+/-7.7 micromol/L and BUN concentration 11.4 mmol/L+/-3.9 mmol/L in Group C and 112.7 micromol/L+/-19.5 micromol/L and 20.7 mmol/L+/-6.6 mmol/L respectively in Group B after 24 hours of reperfusion (P<0.05). The mean histological score by standards of Paller in kidneys was 108.7+/-15.7 in Group C, and 168.8+/-14.8in Group B (P<0.01). The renal ET-1 levels 15 minute and 3 hours after reperfusion were 7.2 pg/mg+/-0.8 pg/mg and 9.6 pg/ml+/-1.0 pg/ml in Group C, and 10.1 pg/ml+/-2.8 pg/ml and 13.0 pg/ml+/-2.7pg/ml in Group B (P<0.01). CONCLUSIONS: The mixture of 8 L-amino acids can provide remarkable protection against renal ischemia-reperfusion injury in rats. This may associate with attenuation of renal ET-1 disorder. PMID- 15294127 TI - Culture of skin-derived precursors and their differentiation into neurons. AB - OBJECTIVE: To investigate the culture method of skin-derived precursors (SKPs) and to explore a new cell source for cell transplantation of central nervous system. METHODS: Cells from skins of juvenile and adult mice were isolated and cultured in serum-free medium. A mechanical method was chosen to passage these cells and they were identified by the immunocytochemistry assay. RESULTS: SKPs could be isolated from adult and neonatal skins. They could be maintained in vitro for long periods with stable proliferation, and expanded as undifferentiated cells in culture for more than 12 passages. About 50% of SKPs expressed nestin and majority of these cells expressed fibronectin when they were plated on polyornithine and laminin coated plates. About 5% cells showed neuronal differentiation and expressed neurofilament-M (NF-M) and NSE when SKPs were plated in serum-containing medium, and these cells could also differentiate into adipocytes and fibroblast-like cells. CONCLUSIONS: The data support the hypothesis that adult skin contains stem cells capable of differentiating into neurons, adipocytes, and fibroblast-like cells. They may represent an alternative autologous stem cell source for CNS cell transplantation. PMID- 15294128 TI - Rapid natural resolution of intracranial hematoma. AB - OBJECTIVE: To investigate the clinical characteristics of intracranial hematoma and the mechanism involved in its rapid natural resolution. METHODS: Seventeen cases of intracranial hematoma with typical clinical and CT manifestations were retrospectively studied. RESULTS: Intracranial hematoma was found obviously decreased in size within 72 h after its occurrence in 8 cases. The rest 9 cases presented complete resolution. CONCLUSIONS: Rapid natural resolution of acute epidural hematoma is mostly found in teenagers and the resolution is correlated with cranial fracture at the hematoma site. As for acute subdural hematoma, its rapid resolution is associated with the transfer of cerebrospinal fluid toward subdural space, the lavage effect, and the compression caused by the increased intracranial pressure or the space left resulting from redistribution of the hematoma in brain atrophy. PMID- 15294129 TI - Comparative study on treating complete dislocation of acromioclavicular joint with three different methods. AB - OBJECTIVE: To comparatively study complete dislocation of acromioclavicular joint treated with three different methods. METHODS: A total of 96 patients (81 males and 15 females, aged 16-59 years, mean=45 years) with complete dislocation of acromioclavicular joint were treated with Dewar's operation (Group A, n=32), internal fixation with Kirschner tension band wires (Group B, n=44), or internal fixation with Wolter plates (Group C, n=20), respectively, in this study. Eighty five patients suffered from acute dislocations and eleven from chronic dislocations. RESULTS: The patients were followed up for 50 months on an average. According to Karlsson's standard, in Group A, 26 patients were assessed as good, 5 as fair and 1 as poor. In Group B, 20 patients were assessed as good, 13 as fair and 11 as poor. In Group C, 15 patients were assessed as good, 4 as fair and 1 as poor. The good and fair rates were significantly different between Group A and Group B, and between Group C and Group B, but no statistical difference was found between Group A and Group C. The operating time was (52.36+/-7.24) minutes, (67.43+/-8.11) minutes and (69.73+/-8.04) minutes in Groups A, B and C, respectively. And the hospitalizing fees were (2400+/-270) yuan, (2100+/-190) yuan and (8450+/-360) yuan in Groups A, B and C, respectively. CONCLUSIONS: Dewar's operation is a good and safe method with shorter operating time and lower hospitalizing fee for treating complete dislocation of acromioclavicular joint. The method is simple without the need of a second operation to remove the implants and with few complications. PMID- 15294130 TI - Spiral CT arthrography of multiplanar reconstruction and virtual arthroscopy technique in diagnosis of knee with internal derangements. AB - OBJECTIVE: To evaluate the values of spiral CT arthrography with multiplanar reconstruction and virtual arthroscopy technique in diagnosis of internal derangements of the knee. METHODS: Ten bovine knees were used for experiment. The menisci, anterior and posterior cruciate ligaments and cartilage of these 10 bovine knees were injured with a hook. Each of the joints was injected with 100 ml air, then soon scanned with a PQ6000 spiral computed tomography scanner. The data obtained was input into the work station, and multiplanar reconstruction technique was used to illustrate lesions in the knees. The results of CT diagnosis were compared to those found by gross inspection of the specimens. Clinically, 10 knees of 9 patients diagnosed as internal derangement were evaluated with the same method after 50-70 ml air was injected into each of the joints. Nine months later, the data of 2 patients were used for CT endoscopy reconstruction. The results were compared with intraoperative findings. RESULTS: Experimentally, the sensitivity and specificity were 88.9% and 93.9% by detection of meniscal abnormalities, 85.7% and 100% by detection of cruciate ligament lesions, and 72.7% and 100% by detection of cartilage damage, respectively. Clinically, the sensitivity and specificity were 90.0% and 95.0% by detection of meniscal lesion. As to ligament, the figures were 85.7% and 100% respectively. Images of virtual arthroscopy simulated the images of real arthroscopy. CONCLUSIONS: Spiral CT arthrography of multiplanar reconstruction technique offers fine images of internal structures of the knee, with clear border and internal structure. It is an accurate method for detecting meniscal, cruciate and collateral ligament and cartilaginous lesions that cause internal derangement of the knee. Virtual arthroscopy technique is a hopeful method for detecting reasons of derangement of the knee. PMID- 15294131 TI - Direct evidence of the existence of specific LPS binding sites on vascular endothelial cells. AB - OBJECTIVE: To investigate the binding characteristics of endothelial cell (EC) with LPS free from the participation of serum factors. METHODS: Laser confocal microscope was employed in the observation of the binding of EC with FITC-LPS. The KD and the binding sites of each EC were calculated by radioligand binding assay of receptors (RBA) using [(3)H]-LPS. RESULTS: The binding of EC with LPS was saturable, time and concentration dependent and it could be competed with overdosed LPS of the same type. The fluorescence mainly distributed in cytoplasm, especially near the nucleus, which could also be stained. CONCLUSIONS: There might be some specific LPS binding sites existing on ECs and LPS could function intracellularily. PMID- 15294132 TI - Application of CT 3D reconstruction in diagnosing atlantoaxial subluxation. AB - OBJECTIVE: To evaluate and compare the diagnostic value in atlantoaxial subluxation by CT three-dimensional (3D) reconstruction. METHODS: 3D reconstruction findings of 41 patients with atlantoaxial subluxation were retrospectively analyzed, and comparisons were made among images of transverse section, multiplanar reformorting (MPR), surface shade display (SSD), maximum intensity project (MIP), and volume rendering (VR). RESULTS: Of 41 patients with atlantoaxial subluxation, 31 belonged to rotary dislocation, 5 antedislocation, and 5 hind dislocation. All the cases showed the dislocated joint panel of atlantoaxial articulation. Fifteen cases showed deviation of the odontoid process and 8 cases widened distance between the dens and anterior arch of the atlas. The dislocated joint panel of atlantoaxial articulation was more clearly seen with SSD-3D imaging than any other methods. CONCLUSIONS: Atlantoaxial subluxation can well be diagnosed by CT 3D reconstruction, in which SSD-3D imaging is optimal. PMID- 15294133 TI - Changes in liquid clearance of alveolar epithelium after oleic acid-induced acute lung injury in rats. AB - OBJECTIVE: Impaired active fluid transport of alveolar epithelium may involve in the pathogenesis and resolution of alveolar edema. The objective of this study was to explore the changes in alveolar epithelial liquid clearance during lung edema following acute lung injury induced by oleic acid. METHODS: Forty-eight Wistar rats were randomly divided into six groups, i.e., injured, amiloride, ouabain, amiloride plus ouabain and terbutaline groups. Twenty-four hours after the induction of acute lung injury by intravenous oleic acid (0.25 ml/kg), 5% albumin solution with 1.5 microCi (125)I-labeled albumin (5 ml/kg) was delivered into both lungs via trachea. Alveolar liquid clearance (ALC), extravascular lung water (EVLW) content and arterial blood gases were measured one hour thereafter. RESULTS: At 24 h after the infusion of oleic acid, the rats developed pulmonary edema and severe hypoxemia, with EVLW increased by 47.9% and ALC decreased by 49.2%. Addition of either 2x10(-3) M amiloride or 5x10(-4) M ouabain to the instillation further reduced ALC and increased EVLW. ALC increased by approximately 63.7% and EVLW decreased by 46.9% with improved hypoxemia in the Terbutaline (10(-4) M) group, compared those in injured rats. A significant negative correlation was found between the increment of EVLW and the reduction of ALC. CONCLUSIONS: Active fluid transport of alveolar epithelium might play a role in the pathogenesis of lung edema in acute lung injury. PMID- 15294134 TI - Chondromalacia of sesamoids in first metatarsophalangeal joint. PMID- 15294135 TI - Clearing tau pathology with Abeta immunotherapy--reversible and irreversible stages revealed. AB - The report by Oddo and colleagues in this issue of Neuron demonstrates for the first time that clearance of amyloid also results in the removal of early-stage tau pathology in mice that develop both amyloid plaques and neurofibrillary tangles (NFT), the two hallmark lesions of Alzheimer's disease (AD). This result supports a primary role for Abeta in AD etiology. PMID- 15294136 TI - Generation of the cortical area map; emx2 strikes back. AB - The recent identification of molecular cues involved in the generation of the cortical area map, such as the patterning molecule FGF8 and transcription factors such as Emx2, represents an important breakthrough. In this issue of Neuron, Hamasaki et al. use a genetic approach to explore how these signals interact and propose that Emx2 plays a direct, largely FGF8-independent role in the control of the relative size and position that each area occupies within the cortex. PMID- 15294137 TI - Don't go there. AB - Response inhibition, or impulse control, is critical for normal cognitive function. In this issue of Neuron, Hasegawa and colleagues use a spatial nonmatch to-sample task to reveal neurons in and around the frontal eye fields that encode where an animal should not look. PMID- 15294138 TI - Mitochondria and dopamine: new insights into recessive parkinsonism. AB - Recessively inherited mutations in parkin, DJ-1, and PINK1 have recently been linked to familial forms of parkinsonism. These syndromes are often clinically indistinguishable from Parkinson's disease, as similar neuronal groups, notably dopaminergic neurons, are selectively affected. Studies of the functions of these gene products may provide insights into the pathogenic mechanisms underlying the selective degeneration of dopaminergic neurons. Emerging evidence that one or several of these genes play important roles in mitochondrial function and the dopaminergic system suggests that these events may be early steps of the pathophysiological changes of the disease. This review will summarize recent advances in our understanding of these gene products, with emphasis on the surprising convergence of their functions. PMID- 15294139 TI - Tethering naturally occurring peptide toxins for cell-autonomous modulation of ion channels and receptors in vivo. AB - The physiologies of cells depend on electrochemical signals carried by ion channels and receptors. Venomous animals produce an enormous variety of peptide toxins with high affinity for specific ion channels and receptors. The mammalian prototoxin lynx1 shares with alpha-bungarotoxin the ability to bind and modulate nicotinic receptors (nAChRs); however, lynx1 is tethered to the membrane via a GPI anchor. We show here that several classes of neurotoxins, including bungarotoxins and cobratoxins, retain their selective antagonistic properties when tethered to the membrane. Targeted elimination of nAChR function in zebrafish can be achieved with tethered alpha-bungarotoxin, silencing synaptic transmission without perturbing synapse formation. These studies harness the pharmacological properties of peptide toxins for use in genetic experiments. When combined with specific methods of cell and temporal expression, the extension of this approach to hundreds of naturally occurring peptide toxins opens a new landscape for cell-autonomous regulation of cellular physiology in vivo. PMID- 15294140 TI - Visualizing sexual dimorphism in the brain. AB - Sexually dimorphic behaviors are likely to involve neural pathways that express the androgen receptor (AR). We have genetically modified the AR locus to visualize dimorphisms in neuronal populations that express AR. Analysis of AR positive neurons reveals both known dimorphisms in the preoptic area of the hypothalamus and the bed nucleus of the stria terminalis as well as novel dimorphic islands in the basal forebrain with a clarity unencumbered by the vast population of AR-negative neurons. This genetic approach allows the visualization of dimorphic subpopulations of AR-positive neurons along with their projections and may ultimately permit an association between neural circuits and specific dimorphic behaviors. PMID- 15294141 TI - Abeta immunotherapy leads to clearance of early, but not late, hyperphosphorylated tau aggregates via the proteasome. AB - Amyloid-beta (Abeta) plaques and neurofibrillary tangles are the hallmark neuropathological lesions of Alzheimer's disease (AD). Using a triple transgenic model (3xTg-AD) that develops both lesions in AD-relevant brain regions, we determined the consequence of Abeta clearance on the development of tau pathology. Here we show that Abeta immunotherapy reduces not only extracellular Abeta plaques but also intracellular Abeta accumulation and most notably leads to the clearance of early tau pathology. We find that Abeta deposits are cleared first and subsequently reemerge prior to the tau pathology, indicative of a hierarchical and direct relationship between Abeta and tau. The clearance of the tau pathology is mediated by the proteasome and is dependent on the phosphorylation state of tau, as hyperphosphorylated tau aggregates are unaffected by the Abeta antibody treatment. These findings indicate that Abeta immunization may be useful for clearing both hallmark lesions of AD, provided that intervention occurs early in the disease course. PMID- 15294142 TI - LRP/amyloid beta-peptide interaction mediates differential brain efflux of Abeta isoforms. AB - LRP (low-density lipoprotein receptor-related protein) is linked to Alzheimer's disease (AD). Here, we report amyloid beta-peptide Abeta40 binds to immobilized LRP clusters II and IV with high affinity (Kd = 0.6-1.2 nM) compared to Abeta42 and mutant Abeta, and LRP-mediated Abeta brain capillary binding, endocytosis, and transcytosis across the mouse blood-brain barrier are substantially reduced by the high beta sheet content in Abeta and deletion of the receptor-associated protein gene. Despite low Abeta production in the brain, transgenic mice expressing low LRP-clearance mutant Abeta develop robust Abeta cerebral accumulations much earlier than Tg-2576 Abeta-overproducing mice. While Abeta does not affect LRP internalization and synthesis, it promotes proteasome dependent LRP degradation in endothelium at concentrations > 1 microM, consistent with reduced brain capillary LRP levels in Abeta-accumulating transgenic mice, AD, and patients with cerebrovascular beta-amyloidosis. Thus, low-affinity LRP/Abeta interaction and/or Abeta-induced LRP loss at the BBB mediate brain accumulation of neurotoxic Abeta. PMID- 15294143 TI - Cell behaviors and genetic lineages of the mesencephalon and rhombomere 1. AB - Brain structures derived from the mesencephalon (mes) and rhombomere 1 (r1) modulate distinct motor and sensory modalities. The precise origin and cellular behaviors underpinning the cytoarchitectural organization of the mes and r1, however, are unknown. Using a novel inducible genetic fate mapping approach in mouse, we determined the fate and lineage relationships of mes/r1 cells with fine temporal and spatial resolution. We demonstrate that the mes and r1 are neuromeres that along with the isthmic organizer are partitioned along the anterior-posterior axis by lineage restriction boundaries established sequentially between E8.5 and E9.5. Furthermore, a small group of cells originating from the most posterior mes exhibit anterior intracompartmental expansion and contribute throughout the inferior colliculus. Finally, we also uncovered transient and differential genetic lineages of ventral midbrain dopaminergic and ventral hindbrain serotonergic neuronal precursors with respect to Wnt1 and Gli1 expression. PMID- 15294144 TI - EMX2 regulates sizes and positioning of the primary sensory and motor areas in neocortex by direct specification of cortical progenitors. AB - Genetic studies of neocortical area patterning are limited, because mice deficient for candidate regulatory genes die before areas emerge and have other complicating issues. To define roles for the homeodomain transcription factor EMX2, we engineered nestin-Emx2 transgenic mice that overexpress Emx2 in cortical progenitors coincident with expression of endogenous Emx2 and survive postnatally. Cortical size, lamination, thalamus, and thalamocortical pathfinding are normal in homozygous nestin-Emx2 mice. However, primary sensory and motor areas are disproportionately altered in size and shift rostrolaterally. Heterozygous transgenics have similar but smaller changes. Opposite changes are found in heterozygous Emx2 knockout mice. Fgf8 expression in the commissural plate of nestin-Emx2 mice is indistinguishable from wild-type, but Pax6 expression is downregulated in rostral cortical progenitors, suggesting that EMX2 repression of PAX6 specification of rostral identities contributes to reduced rostral areas. We conclude that EMX2 levels in cortical progenitors disproportionately specify sizes and positions of primary cortical areas. PMID- 15294145 TI - Serotonin regulates the secretion and autocrine action of a neuropeptide to activate MAPK required for long-term facilitation in Aplysia. AB - In Aplysia, long-term facilitation (LTF) of sensory neuron synapses requires activation of both protein kinase A (PKA) and mitogen-activated protein kinase (MAPK). We find that 5-HT through activation of PKA regulates secretion of the sensory neuron-specific neuropeptide sensorin, which binds autoreceptors to activate MAPK. Anti-sensorin antibody blocked LTF and MAPK activation produced by 5-HT and LTF produced by medium containing sensorin that was secreted from sensory neurons after 5-HT treatment. A single application of 5-HT followed by a 2 hr incubation with sensorin produced protein synthesis-dependent LTF, growth of new presynaptic varicosities, and activation of MAPK and its translocation into sensory neuron nuclei. Inhibiting PKA during 5-HT applications and inhibiting receptor tyrosine kinase or MAPK during sensorin application blocked both LTF and MAPK activation and translocation. Thus, long-term synaptic plasticity is produced when stimuli activate kinases in a specific sequence by regulating the secretion and autocrine action of a neuropeptide. PMID- 15294146 TI - Presynaptic Ca2+ channels compete for channel type-preferring slots in altered neurotransmission arising from Ca2+ channelopathy. AB - Several human channelopathies result from mutations in alpha1A, the pore-forming subunit of P/Q-type Ca2+ channels, conduits of presynaptic Ca2+ entry for evoked neurotransmission. We found that wild-type human alpha1A subunits supported transmission between cultured mouse hippocampal neurons equally well as endogenous mouse alpha1A, whereas introduction of impermeant human alpha1A hampered the effect of endogenous subunits. Thus, presynaptic P/Q-type channels may compete for channel type-preferring "slots" that limit their synaptic effectiveness. The existence of slots generates predictions for how neurotransmission might be affected by changes in Ca2+ channel properties, which we tested by studying alpha1A mutations that are associated with familial hemiplegic migraine type 1 (FHM1). Mutant human P/Q-type channels were impaired in contributing to neurotransmission in precise accord with their deficiency in supporting whole-cell Ca2+ channel activity. Expression of mutant channels in wild-type neurons reduced the synaptic contribution of P/Q-type channels, suggesting that competition for type-preferring slots might support the dominant inheritance of FHM1. PMID- 15294147 TI - Homer proteins regulate sensitivity to cocaine. AB - Drug addiction involves complex interactions between pharmacology and learning in genetically susceptible individuals. Members of the Homer gene family are regulated by acute and chronic cocaine administration. Here, we report that deletion of Homer1 or Homer2 in mice caused the same increase in sensitivity to cocaine-induced locomotion, conditioned reward, and augmented extracellular glutamate in nucleus accumbens as that elicited by withdrawal from repeated cocaine administration. Moreover, adeno-associated virus-mediated restoration of Homer2 in the accumbens of Homer2 KO mice reversed the cocaine-sensitized phenotype. Further analysis of Homer2 KO mice revealed extensive additional behavioral and neurochemical similarities to cocaine-sensitized animals, including accelerated acquisition of cocaine self-administration and altered regulation of glutamate by metabotropic glutamate receptors and cystine/glutamate exchange. These data show that Homer deletion mimics the behavioral and neurochemical phenotype produced by repeated cocaine administration and implicate Homer in regulating addiction to cocaine. PMID- 15294148 TI - Prefrontal neurons coding suppression of specific saccades. AB - The prefrontal cortex has been implicated in the suppression of unwanted behavior, based upon observations of humans and monkeys with prefrontal lesions. Despite this, there has been little direct neurophysiological evidence for a mechanism that suppresses specific behavior. In this study, we used an oculomotor delayed match/nonmatch-to-sample task in which monkeys had to remember a stimulus location either as a marker of where to look or as a marker of where not to look. We found a group of neurons in both the frontal eye field and the caudal prefrontal cortex that carried signals selective for the forbidden stimulus. The activity of these "don't look" neurons correlated with the monkeys' success or failure on the task. These results demonstrate a frontal signal that is related to the active suppression of one action while the subject performs another. PMID- 15294149 TI - Interaction between the human hippocampus and the caudate nucleus during route recognition. AB - Navigation through familiar environments can rely upon distinct neural representations that are related to different memory systems with either the hippocampus or the caudate nucleus at their core. However, it is a fundamental question whether and how these systems interact during route recognition. To address this issue, we combined a functional neuroimaging approach with a naturally occurring, well-controlled human model of caudate nucleus dysfunction (i.e., preclinical and early-stage Huntington's disease). Our results reveal a noncompetitive interaction so that the hippocampus compensates for gradual caudate nucleus dysfunction with a gradual activity increase, maintaining normal behavior. Furthermore, we revealed an interaction between medial temporal and caudate activity in healthy subjects, which was adaptively modified in Huntington patients to allow compensatory hippocampal processing. Thus, the two memory systems contribute in a noncompetitive, cooperative manner to route recognition, which enables the hippocampus to compensate seamlessly for the functional degradation of the caudate nucleus. PMID- 15294150 TI - Positioning the Golgi apparatus. AB - Rios et al. (2004) report in this issue that the Golgi protein GMAP-210 is sufficient to confer pericentrosomal positioning and recruits gamma-tubulin and associated microtubule-nucleating ring complex proteins to Golgi membranes. The results raise the possibility that short microtubules emanate from the Golgi to mediate its organization and positioning. PMID- 15294151 TI - Myosin assembly: the power of multiubiquitylation. AB - Ubiquitylation provides a means of targeting substrate proteins for degradation by the proteasome. Novel findings in C. elegans (Hoppe et al., 2004, this issue of Cell) establish that two ubiquitin-ligases team up to multiubiquitylate the myosin chaperone UNC-45, suggesting a novel link between regulated protein degradation and myosin assembly. PMID- 15294152 TI - Gene targeting: attention to detail. AB - When it comes to silencing genes in mice, not all approaches are equal. An example published in this issue of Cell (Patrucco et al., 2004) suggests that caution should be used when validating potential drug targets by genetic disruption. PMID- 15294153 TI - Epithelial-mesenchymal transitions: twist in development and metastasis. AB - Epithelial-mesenchymal transitions (EMT) are vital for morphogenesis during embryonic development and are also implicated in the conversion of early stage tumors into invasive malignancies. Several key inducers of EMT are transcription factors that repress E-cadherin expression. A recent report in Cell (Yang et al., 2004) adds Twist to this list and links EMT to the ability of breast cancer cells to enter the circulation and seed metastases. PMID- 15294154 TI - Regulation of RNA polymerase through the secondary channel. AB - High-resolution crystal structures have highlighted functionally important regions in multisubunit RNA polymerases, including the secondary channel, or pore, which is postulated to allow the diffusion of small molecules both into and out of the active center of the enzyme. Recent work from several groups has illustrated how regulatory factors and small molecules can exploit the secondary channel to gain access to the active site and modify the transcription properties of RNA polymerase. PMID- 15294155 TI - IKKbeta links inflammation and tumorigenesis in a mouse model of colitis associated cancer. AB - A link between inflammation and cancer has long been suspected, but its molecular nature remained ill defined. A key player in inflammation is transcription factor NF-kappaB whose activity is triggered in response to infectious agents and proinflammatory cytokines via the IkappaB kinase (IKK) complex. Using a colitis associated cancer model, we show that although deletion of IKKbeta in intestinal epithelial cells does not decrease inflammation, it leads to a dramatic decrease in tumor incidence without affecting tumor size. This is linked to increased epithelial apoptosis during tumor promotion. Deleting IKKbeta in myeloid cells, however, results in a significant decrease in tumor size. This deletion diminishes expression of proinflammatory cytokines that may serve as tumor growth factors, without affecting apoptosis. Thus, specific inactivation of the IKK/NF kappaB pathway in two different cell types can attenuate formation of inflammation-associated tumors. In addition to suppressing apoptosis in advanced tumors, IKKbeta may link inflammation to cancer. PMID- 15294156 TI - Regulation through the secondary channel--structural framework for ppGpp-DksA synergism during transcription. AB - Bacterial transcription is regulated by the alarmone ppGpp, which binds near the catalytic site of RNA polymerase (RNAP) and modulates its activity. We show that the DksA protein is a crucial component of ppGpp-dependent regulation. The 2.0 A resolution structure of Escherichia coli DksA reveals a globular domain and a coiled coil with two highly conserved Asp residues at its tip that is reminiscent of the transcript cleavage factor GreA. This structural similarity suggests that DksA coiled coil protrudes into the RNAP secondary channel to coordinate a ppGpp bound Mg2+ ion with the Asp residues, thereby stabilizing the ppGpp-RNAP complex. Biochemical analysis demonstrates that DksA affects transcript elongation, albeit differently from GreA; augments ppGpp effects on initiation; and binds directly to RNAP, positioning the Asp residues near the active site. Substitution of these residues eliminates the synergy between DksA and ppGpp. Thus, the secondary channel emerges as a common regulatory entrance for transcription factors. PMID- 15294157 TI - DksA: a critical component of the transcription initiation machinery that potentiates the regulation of rRNA promoters by ppGpp and the initiating NTP. AB - Ribosomal RNA (rRNA) transcription is regulated primarily at the level of initiation from rRNA promoters. The unusual kinetic properties of these promoters result in their specific regulation by two small molecule signals, ppGpp and the initiating NTP, that bind to RNA polymerase (RNAP) at all promoters. We show here that DksA, a protein previously unsuspected as a transcription factor, is absolutely required for rRNA regulation. In deltadksA mutants, rRNA promoters are unresponsive to changes in amino acid availability, growth rate, or growth phase. In vitro, DksA binds to RNAP, reduces open complex lifetime, inhibits rRNA promoter activity, and amplifies effects of ppGpp and the initiating NTP on rRNA transcription, explaining the dksA requirement in vivo. These results expand our molecular understanding of rRNA transcription regulation, may explain previously described pleiotropic effects of dksA, and illustrate how transcription factors that do not bind DNA can nevertheless potentiate RNAP for regulation. PMID- 15294158 TI - GMAP-210 recruits gamma-tubulin complexes to cis-Golgi membranes and is required for Golgi ribbon formation. AB - Mammalian cells concentrate Golgi membranes around the centrosome in a microtubule-dependent manner. The mechanisms involved in generating a single Golgi ribbon in the periphery of the centrosome remain unknown. Here we show that GMAP-210, a cis-Golgi microtubule binding protein, recruits gamma-tubulin containing complexes to Golgi membranes even in conditions where microtubule polymerization is prevented and independently of Golgi apparatus localization within the cell. Under overexpression conditions, very short microtubules, or tubulin oligomers, are stabilized on Golgi membranes. GMAP-210 depletion by RNA interference results in extensive fragmentation of the Golgi apparatus, supporting a role for GMAP-210 in Golgi ribbon formation. Targeting of GMAP-210 or its C terminus to mitochondria induces the recruitment of gamma-tubulin to their surface and redistribution of mitochondria to a pericentrosomal location. All our experiments suggest that GMAP-210 displays microtubule anchoring and membrane fusion activities, thus contributing to the assembly and maintenance of the Golgi ribbon around the centrosome. PMID- 15294159 TI - Regulation of the myosin-directed chaperone UNC-45 by a novel E3/E4 multiubiquitylation complex in C. elegans. AB - The organization of the motor protein myosin into motile cellular structures requires precise temporal and spatial control. Caenorhabditis elegans UNC-45 facilitates this by functioning both as a chaperone and as a Hsp90 cochaperone for myosin during thick filament assembly. Consequently, mutations in C. elegans unc-45 result in paralyzed animals with severe myofibril disorganization in striated body wall muscles. Here, we report a new E3/E4 complex, formed by CHN-1, the C. elegans ortholog of CHIP (carboxyl terminus of Hsc70-interacting protein), and UFD-2, an enzyme known to have ubiquitin conjugating E4 activity in yeast, as necessary and sufficient to multiubiquitylate UNC-45 in vitro. The phenotype of unc-45 temperature-sensitive animals is partially suppressed by chn-1 loss of function, while UNC-45 overexpression in worms deficient for chn-1 results in severely disorganized muscle cells. These results identify CHN-1 and UFD-2 as a functional E3/E4 complex and UNC-45 as its physiologically relevant substrate. PMID- 15294160 TI - MAP kinase-mediated stress relief that precedes and regulates the timing of transcriptional induction. AB - In yeast, hyperosmotic stress causes an immediate dissociation of most proteins from chromatin, presumably because cells are unprepared for, and initially unresponsive to, increased ion concentrations in the nucleus. Osmotic stress activates Hog1 MAP kinase, which phosphorylates at least two proteins located at the plasma membrane, the Nha1 Na+/H+ antiporter and the Tok1 potassium channel. Hog1 phosphorylation stimulates Nha1 activity, and this is crucial for the rapid reassociation of proteins with their target sites in chromatin. This initial response to hyperosmolarity precedes and temporally regulates the activation of stress-response genes that depends on Hog1 phosphorylation of transcription factors in the nucleus. Thus, a single MAP kinase coordinates temporally, spatially, and mechanistically distinct responses to stress, thereby providing very rapid stress relief that facilitates subsequent changes in gene expression that permit long-term adaptation to harsh environmental conditions. PMID- 15294161 TI - Lamellipodial versus filopodial mode of the actin nanomachinery: pivotal role of the filament barbed end. AB - Understanding how a particular cell type expresses the lamellipodial or filopodial form of the actin machinery is essential to understanding a cell's functional interactions. To determine how a cell "chooses" among these alternative modes of "molecular hardware," we tested the role of key proteins that affect actin filament barbed ends. Depletion of capping protein (CP) by short hairpin RNA (shRNA) caused loss of lamellipodia and explosive formation of filopodia. The knockdown phenotype was rescued by a CP mutant refractory to shRNA, but not by another barbed-end capper, gelsolin, demonstrating that the phenotype was specific for CP. In Ena/VASP deficient cells, CP depletion resulted in ruffling instead of filopodia. We propose a model for selection of lamellipodial versus filopodial organization in which CP is a negative regulator of filopodia formation and Ena/VASP has recruiting/activating functions downstream of actin filament elongation in addition to its previously suggested anticapping and antibranching activities. PMID- 15294162 TI - PI3Kgamma modulates the cardiac response to chronic pressure overload by distinct kinase-dependent and -independent effects. AB - The G protein-coupled, receptor-activated phosphoinositide 3-kinase gamma (PI3Kgamma) mediates inflammatory responses and negatively controls cardiac contractility by reducing cAMP concentration. Here, we report that mice carrying a targeted mutation in the PI3Kgamma gene causing loss of kinase activity (PI3KgammaKD/KD) display reduced inflammatory reactions but no alterations in cardiac contractility. We show that, in PI3KgammaKD/KD hearts, cAMP levels are normal and that PI3Kgamma-deficient mice but not PI3KgammaKD/KD mice develop dramatic myocardial damage after chronic pressure overload induced by transverse aortic constriction (TAC). Finally, our data indicate that PI3Kgamma is an essential component of a complex controlling PDE3B phosphodiesterase-mediated cAMP destruction. Thus, cardiac PI3Kgamma participates in two distinct signaling pathways: a kinase-dependent activity that controls PKB/Akt as well as MAPK phosphorylation and contributes to TAC-induced cardiac remodeling, and a kinase independent activity that relies on protein interactions to regulate PDE3B activity and negatively modulates cardiac contractility. PMID- 15294163 TI - Calmodulin and Munc13 form a Ca2+ sensor/effector complex that controls short term synaptic plasticity. AB - The efficacy of synaptic transmission between neurons can be altered transiently during neuronal network activity. This phenomenon of short-term plasticity is a key determinant of network properties; is involved in many physiological processes such as motor control, sound localization, or sensory adaptation; and is critically dependent on cytosolic [Ca2+]. However, the underlying molecular mechanisms and the identity of the Ca2+ sensor/effector complexes involved are unclear. We now identify a conserved calmodulin binding site in UNC-13/Munc13s, which are essential regulators of synaptic vesicle priming and synaptic efficacy. Ca2+ sensor/effector complexes consisting of calmodulin and Munc13s regulate synaptic vesicle priming and synaptic efficacy in response to a residual [Ca2+] signal and thus shape short-term plasticity characteristics during periods of sustained synaptic activity. PMID- 15294164 TI - Musings for young scientists. PMID- 15294165 TI - Clinical gene therapy: lessons from the ether dome. PMID- 15294166 TI - Opening the floodgates: clinically applicable hydrodynamic delivery of plasmid DNA to skeletal muscle. PMID- 15294167 TI - Cytokines: value-added products in hematopoietic stem cell gene therapy. PMID- 15294168 TI - Trends in enzyme therapy for phenylketonuria. AB - Phenylketonuria (PKU) is an inborn error of amino acid metabolism caused by phenylalanine hydroxylase (PAH) deficiency. Dietary treatment has been the cornerstone for controlling systemic phenylalanine (Phe) levels in PKU for the past 4 decades. Over the years, it has become clear that blood Phe concentration needs to be controlled for the life of the patient, a difficult task taking into consideration that the diet becomes very difficult to maintain. Therefore alternative models of therapy are being pursued. This review describes the progress made in enzyme replacement therapy for PKU. Two modalities are discussed, PAH and phenylalanine ammonia-lyase PAH. Developing stable and functional forms of both enzymes has proven difficult, but recent success in producing polyethylene glycol-modified forms of active and stable PAH shows promise. PMID- 15294169 TI - Uncertain benefit: investigators' views and communications in early phase gene transfer trials. AB - We report on a study of potential sources of therapeutic misconception in early phase gene transfer research, examining how investigators and their consent forms represent the prospect for direct benefit. Our analysis demonstrates that even though half of PIs said they expected direct medical benefit for their subjects, they did not necessarily convey this to their subjects. What they reported telling subjects resembled what was written in their consent form, which suggests that, far from being irrelevant, the consent form is an influential component of the consent process. We also demonstrate that the language used to describe direct benefit in consent forms and PIs' discussions was mostly vague, ambiguous, and indeterminate about benefit, rather than clearly negative. This was especially true for cancer and vascular disease trials. Our respondents found the problem of balancing hopes and expectations, for themselves and for their subjects, extraordinarily challenging. In the current era, investigators face such challenges without consistent normative guidance or agreed-upon standards for how to talk about scientific promise and uncertainty in early phase trials. This dilemma cannot be effectively addressed by individual investigators alone, but must be acknowledged and openly discussed by the scientific community at large. PMID- 15294170 TI - Targeted exon skipping in transgenic hDMD mice: A model for direct preclinical screening of human-specific antisense oligonucleotides. AB - The therapeutic potential of frame-restoring exon skipping by antisense oligonucleotides (AONs) has recently been demonstrated in cultured muscle cells from a series of Duchenne muscular dystrophy (DMD) patients. To facilitate clinical application, in vivo studies in animal models are required to develop safe and efficient AON-delivery methods. However, since exon skipping is a sequence-specific therapy, it is desirable to target the human DMD gene directly. We therefore set up human sequence-specific exon skipping in transgenic mice carrying the full-size human gene (hDMD). We initially compared the efficiency and toxicity of intramuscular AON injections using different delivery reagents in wild-type mice. At a dose of 3.6 nmol AON and using polyethylenimine, the skipping levels accumulated up to 3% in the second week postinjection and lasted for 4 weeks. We observed a correlation of this long-term effect with the intramuscular persistence of the AON. In regenerating myofibers higher efficiencies (up to 9%) could be obtained. Finally, using the optimized protocols in hDMD mice, we were able to induce the specific skipping of human DMD exons without affecting the endogenous mouse gene. These data highlight the high sequence specificity of this therapy and present the hDMD mouse as a unique model to optimize human-specific exon skipping in vivo. PMID- 15294171 TI - Nitric oxide is a regulator of hematopoietic stem cell activity. AB - Hematopoietic stem cells give rise to various multipotent progenitor populations, which expand in response to cytokines and which ultimately generate all of the elements of the blood. Here we show that it is possible to increase the number of stem and progenitor cells in the bone marrow (BM) by suppressing the activity of NO synthases (NOS). Exposure of mice to NOS inhibitors, either directly or after irradiation and BM transplantation, increases the number of stem cells in the BM. In the transplantation model, this increase is followed by a transient increase in the number of neutrophils in the peripheral blood. Thus, our results indicate that NO is important for the control of hematopoietic stem cells in the BM. They further suggest that suppression of NO synthase activity may allow expansion of the number of hematopoietic stem and progenitor cells or neutrophils for therapeutic purposes. PMID- 15294172 TI - Self-inactivating lentiviral vectors resist proviral methylation but do not confer position-independent expression in hematopoietic stem cells. AB - Oncoretroviral expression is transcriptionally silenced in embryonic and hematopoietic stem cells (HSCs). This is associated with methylation of viral and internal promoters. We determined whether self-inactivating (SIN) lentiviral vectors (LV) would circumvent proviral silencing in HSCs. We studied long-term expression, methylation, and position effects (PE) from two GFP-encoding SIN-LV containing erythroid enhancers and the human ankyrin-1 promoter (h-Ank-P) using the murine secondary bone marrow (BM) transplant assay. Proviral expression was detected in RBC 6-11 months following transplant only in 28 of 49 secondary mice, with 0.9 +/- 0.2 copy/cell and oligoclonally integrated provirus in BM, spleen, and thymus. Twenty-one of 49 secondary mice lacked integrated provirus. Secondary mice containing provirus also had GFP-expressing RBCs, although proviral copy number did not always correlate with expression, suggesting either proviral methylation or chromatin PE. The endogenous h-Ank-P was partially methylated in nonerythroid cell lines and unmethylated in erythroid cell lines. However, h-Ank P in the provirus was unmethylated in erythroid and nonerythroid cells within secondary murine BM. Despite lack of methylation, GFP expression was variable in secondary BFU-e and in single-copy mouse erythroleukemia cell clones. Taken together, these data show that erythroid-specific SIN-LV express long term and resist methylation-associated proviral silencing, but may require additional elements to confer position-independent expression. PMID- 15294173 TI - Integration and long-term expression in xenografted human glioblastoma cells using a plasmid-based transposon system. AB - Gene therapy has the potential to become an effective component of cancer treatment by transferring genes that cause immunomodulation or tumor cell death or that inhibit angiogenesis into tumor cells or tumor-associated stroma. Viral vectors have been the primary gene transfer vehicles used for intratumoral gene transfer to date. Plasmid-based vectors may be safer and more scalable than viral vectors. However, attempts at plasmid-based intratumoral gene transfer have been met with transient expression and poor gene transfer efficiency. Here we report integration and long-term expression of reporter genes in human glial tumors, growing in nude mice, using the Sleeping Beauty (SB) transposon system. A two plasmid system was used, in which linear polyethylenimine was complexed with a GFP, NEO, or luciferase transposon plasmid and a SB transposase-expressing plasmid. SB-mediated transposition led to chromosomal integration of the NEO transgene in roughly 8% of tumor cells. SB-mediated insertions were cloned from the genomes of glial tumor cells to provide molecular proof of transposase mediated integration. Luciferase studies showed that SB facilitated long-term expression of the transgene in glial tumors. SB-mediated intratumoral gene transfer is a novel, nonviral technique that could be used to augment conventional therapy for glioblastoma or other cancers. PMID- 15294174 TI - Long-term transgene expression from plasmid DNA gene therapy vectors is negatively affected by CpG dinucleotides. AB - CpG-reduced, CMV-based plasmid DNA constructs encoding human alpha-galactosidase A and factor IX were injected into C57Bl/6, BALB/c, and CD1 mice using hydrodynamics-based delivery of plasmid DNA (pDNA), and gene expression was monitored for 6 months. Linearized and supercoiled pDNAs were compared for their abilities to support long-term expression and to generate immune responses to the transgene product. In all mouse strains supercoiled CpG-reduced pDNA encoding alpha-galactosidase A and factor IX generated higher and more sustained levels of circulating gene product than their supercoiled CpG-replete analogs. Linearizing supercoiled CpG-reduced pDNA did not significantly increase levels of circulating gene product beyond levels supercoiled CpG-reduced pDNA could achieve. Linearizing supercoiled CpG-replete pDNA vectors significantly increased expression compared to their supercoiled CpG-replete analogs, but the increase was short-lived or subtherapeutic. Regardless of vector, liver depot expression did not elicit significant antibody responses to human alpha-galactosidase A or factor IX. Taken together, these data suggest that a clinically acceptable hydrodynamics-based approach targeting the liver combined with CpG-reduced pDNA vectors may represent a viable option for individuals with hemophilia, a lysosomal storage disease, or other disease in which prolonged depot expression of a therapeutic protein from the liver is desirable. PMID- 15294175 TI - HLA-A*0201-restricted cytolytic responses to the rtTA transactivator dominant and cryptic epitopes compromise transgene expression induced by the tetracycline on system. AB - The tetracycline-controlled transcription system (Tet-on) is widely used to regulate gene expression in mammalian cells. In gene therapy applications, immune responses to Tet-on proteins such as the rtTA transcription factor have been reported, raising concerns about their occurrence in humans. To monitor the HLA class I cytolytic responses against Tet-on regulators, we characterized the immunogenic CD8+ epitopes within rtTA and tTS regulators using HLA-A*0201 class I transgenic mice. Epitope prediction programs, HLA-A*0201 binding assays, and peptide immunization were used to select a set of immunogenic peptides within rtTA and tTS sequences. To identify further the rejection epitopes, we expressed Tet-on protein components in vivo and found a single dominant rtTA186 CTL epitope in the rtTA tetracycline repressor domain. Target cells expressing rtTA were susceptible to CTL lysis, and rtTA expression compromised muscle transgene engraftment. To reduce the occurrence of immune responses to rtTA protein, we mutated the dominant rtTA186 epitope and found that this leads to the appearance of subdominant epitopes. As a result, we think that an epitope modification strategy is not applicable to blunt the immune response in this model. Moreover, the identification of HLA-A*0201 rtTA epitopes allowed us to demonstrate here that the delivery of the Tet-on system with weakly immunogenic rAAV vectors does not trigger primary CTL responses in mice, in contrast to DNA transfer. Altogether, the existence of HLA-A*0201 rtTA epitopes may lead to the occurrence of immune responses depending on vectors and local inflammation in gene therapy applications involving rtTA-based regulatory systems. PMID- 15294176 TI - Therapeutic HER2/Neu DNA vaccine inhibits mouse tumor naturally overexpressing endogenous neu. AB - The therapeutic efficacy of HER2/c-erbB-2/neu DNA immunization on mouse tumor cells expressing exogenous human or rat p185neu but not on mouse tumor cells naturally expressing mouse p185neu has been demonstrated. We investigated the feasibility of using N-terminal rat neu DNA immunization on mouse tumor overexpressing endogenous p185neu and enhancing the therapeutic efficacy of this vaccine by fusion to various cytokine genes, including interleukin-2 (IL-2), interleukin-4 (IL-4), or granulocyte-macrophage colony-stimulating factor. In a therapeutic model, N'-neu-IL-2 DNA vaccine was significantly better than N'-neu DNA vaccine, and N'-neu DNA vaccine was significantly better than control DNA or N'-neu-IL-4 DNA vaccine. The therapeutic efficacy of DNA vaccines was correlated with tumor infiltration of CD8+ T cells. Depletion of CD8+ T cells completely abolished the therapeutic effects of N'-neu-IL-2 DNA vaccine and N'-neu DNA vaccine. Depletion of CD4+ T cells after tumor implantation had no influence on N'-neu-IL-2 DNA vaccine, but enhanced the therapeutic efficacy of N'-neu DNA vaccine. Our results demonstrate that rat N'-neu DNA vaccine has a therapeutic effect on established tumor through the CD8+ T-cell-dependent pathway. Depletion of CD4+ T cells or fusion to the IL-2 gene can thus further enhance the therapeutic effects of N'-neu DNA immunization on mouse tumor expressing endogenous p185neu. PMID- 15294177 TI - Recombinant AAV viral vectors pseudotyped with viral capsids from serotypes 1, 2, and 5 display differential efficiency and cell tropism after delivery to different regions of the central nervous system. AB - Recombinant adeno-associated virus 2 (rAAV2) has been shown to deliver genes to neurons effectively in the brain, retina, and spinal cord. The characterization of new AAV serotypes has revealed that they have different patterns of transduction in diverse tissues. We have investigated the tropism and transduction frequency in the central nervous system (CNS) of three different rAAV vector serotypes. The vectors contained AAV2 terminal repeats flanking a green fluorescent protein expression cassette under the control of the synthetic CBA promoter, in AAV1, AAV2, or AAV5 capsids, producing the pseudotypes rAAV2/1, rAAV2/2, and rAAV2/5. Rats were injected with rAAV2/1, rAAV2/2, or rAAV2/5 into selected regions of the CNS, including the hippocampus (HPC), substantia nigra (SN), striatum, globus pallidus, and spinal cord. In all regions injected, the three vectors transduced neurons almost exclusively. All three vectors transduced the SN pars compacta with high efficiency, but rAAV2/1 and rAAV2/5 also transduced the pars reticulata. Moreover, rAAV2/1 showed widespread distribution throughout the entire midbrain. In the HPC, rAAV2/1 and rAAV2/5 targeted the pyramidal cell layers in the CA1-CA3 regions, whereas AAV2/2 primarily transduced the hilar region of the dentate gyrus. In general, rAAV2/1 and rAAV2/5 exhibited higher transduction frequencies than rAAV2/2 in all regions injected, although the differences were marginal in some regions. Retrograde transport of rAAV1 and rAAV5 was also observed in particular CNS areas. These results suggest that vectors based on distinct AAV serotypes can be chosen for specific applications in the nervous system. PMID- 15294178 TI - Intradermal injection of lentiviral vectors corrects regenerated human dystrophic epidermolysis bullosa skin tissue in vivo. AB - Dystrophic epidermolysis bullosa (DEB) is a family of inherited mechanobullous disorders caused by mutations in the gene, COL7A1, that codes for type VII, (anchoring fibril), collagen, which is critical for epidermal-dermal adherence. Most gene therapy approaches have been ex vivo, involving cell culture and culture graft transplantation, which is logistically difficult. To develop a more simplified approach, we engineered a self-inactivating lentiviral vector expressing human type VII collagen and injected this vector intradermally into hairless, immunodeficient mice and into a human DEB composite skin equivalent grafted onto immunodeficient mice. In both situations, the vector transduced dermal cells, which in turn synthesized and exported type VII collagen into the extracellular space. Remarkably, the type VII collagen selectively adhered to and incorporated into the basement membrane zone (BMZ) between the dermis and the epidermis, where it formed anchoring fibril structures. In the case of the DEB skin equivalent, the newly expressed type VII collagen reversed the DEB phenotype characterized by poor epidermal-dermal adherence and anchoring fibril defects. A single lentiviral vector injection provided stable type VII collagen at the BMZ for at least 3 months. These data demonstrate efficient and long-term type VII collagen gene transfer in vivo using direct intradermal injection of an engineered lentiviral vector. PMID- 15294179 TI - Enhanced repair of the anterior cruciate ligament by in situ gene transfer: evaluation in an in vitro model. AB - The inability of the ruptured anterior cruciate ligament (ACL) of the knee joint to heal spontaneously presents numerous clinical problems. Here we describe a novel, gene-based approach to augment ACL healing. It is based upon the migration of cells from the ruptured ends of the ligament into a collagen hydrogel laden with recombinant adenovirus. Cells entering the gel become transduced by the vector, which provides a basis for the local synthesis of gene products that aid repair. Monolayers of bovine ACL cells were readily transduced by first generation, recombinant adenovirus, and transgene expression remained high after the cells were incorporated into collagen hydrogels. Using an in vitro model of ligament repair, cells migrated from the cut ends of the ACL into the hydrogel and were readily transduced by recombinant adenovirus contained within it. The results of experiments in which GFP was used as the transgene suggest highly efficient transduction of ACL cells in this manner. Moreover, during a 21-day period GFP+ cells were observed more than 6 mm from the severed ligament. This distance is ample for the projected clinical application of this technology. In response to TGF-beta1 as the transgene, greater numbers of ACL cells accumulated in the hydrogels, where they deposited larger amounts of type III collagen. These data confirm that it is possible to transduce ACL cells efficiently in situ as they migrate from the ruptured ACL, that transduction does not interfere with the cells' ability to migrate distances necessary for successful repair, and that ACL cells will respond in a suitable manner to the products of the transgenes they express. This permits optimism over a possible clinical use for this technology. PMID- 15294180 TI - Adenoviral-mediated expression of porphobilinogen deaminase in liver restores the metabolic defect in a mouse model of acute intermittent porphyria. AB - The aim of this study was to investigate the potential of gene therapy in the treatment of acute intermittent porphyria (AIP), a disorder caused by a partial deficiency of porphobilinogen deaminase (PBGD), the third enzyme in heme synthesis. The condition is biochemically characterized by accumulation of the porphyrin precursors 5-aminolevulinic acid (ALA) and porphobilinogen (PBG). Here we present the first experiments in vivo using adenoviral vectors to replace the deficient enzyme in the liver of an AIP mouse model. The use of adenoviral vector carrying the cDNA of luciferase in wild-type mice confirmed that transgene expression after intravenous administration was found mainly in liver. When PBGD deficient mice were administered with adenoviral vector carrying the cDNA of mouse PBGD, the hepatic PBGD activity increased in a dose- and time-dependent manner. The highest activity was found 7 days after injection and remained high after 29 days. The expressed enzyme was shown to correct the metabolic defect in the PBGD-deficient mice as no accumulation of ALA or PBG occurred in plasma, liver, or kidney after induction of heme synthesis by phenobarbital. The study demonstrates that hepatic PBGD expression prevents the accumulation of porphyrin precursors, suggesting a future potential for gene therapy in AIP. PMID- 15294181 TI - Effect of adenovirus serotype 5 fiber and penton modifications on in vivo tropism in rats. AB - Sequestration of adenovirus serotype 5 (Ad5) in liver restricts its use for gene delivery to other target sites in vivo. To date, no studies have systematically assessed the impact of genetic capsid modifications on in vivo tropism in rats, an important preclinical model for many disease types. We evaluated a panel of Ad5 vectors with capsid mutations or pseudotyped with the short fiber from serotype 41 (Ad41s) for infectivity in Wistar Kyoto rats in vitro and systemically in vivo. In vitro studies demonstrated that both coxsackie and adenovirus receptor (CAR) and heparan sulfate proteoglycan (HSPG) binding were predominant predictors of Ad5 tropism. In vivo, neither CAR nor integrin mutations alone affected liver transduction. The HSPG-binding mutation alone moderately reduced rat liver transgene levels by 2-fold (P < 0.05). This was further substantially decreased by additional mutation of CAR binding (95-fold). Combining CAR and integrin mutations reduced transgene levels by >99% (509-fold, P < 0.01), an effect not observed in parallel experiments in mice and highly variable when studied further in an additional two strains of rat. Ad41s mediated very low liver transduction (58-fold lower than AdCTL). Moreover, CAR-binding mutants (KO1-containing) or pseudotyping 41s eliminated hemagglutination of rat and human red blood cells in vitro. This highlights some important potential species and strain differences dictating Ad5 tropism in vivo and identifies vectors that are substantially detargeted from rat liver in vivo. PMID- 15294182 TI - A novel TARP-promoter-based adenovirus against hormone-dependent and hormone refractory prostate cancer. AB - TARP (T cell receptor gamma-chain alternate reading frame protein) is a protein that in males is uniquely expressed in prostate epithelial cells and prostate cancer cells. We have previously shown that the transcriptional activity of a chimeric sequence comprising the TARP promoter (TARPp) and the PSA enhancer (PSAe) is strictly controlled by testosterone and highly restricted to cells of prostate origin. Here we report that a chimeric sequence comprising TARPp and the PSMA enhancer (PSMAe) is highly active in testosterone-deprived prostate cancer cells, while a regulatory sequence comprising PSAe, PSMAe, and TARPp (PPT) has high prostate-specific activity both in the presence and in the absence of testosterone. Therefore, the PPT sequence may, in a gene therapy setting, be beneficial to prostate cancer patients that have been treated with androgen withdrawal. A recombinant adenovirus vector with the PPT sequence, shielded from interfering adenoviral sequences by the mouse H19 insulator, yields high and prostate-specific transgene expression both in cell cultures and when prostate cancer, PC-346C, tumors were grown orthotopically in nude mice. Intravenous virus administration reveals both higher activity and higher selectivity for the insulator-shielded PPT sequence than for the immediate-early CMV promoter. Therefore, we believe that an adenovirus with therapeutic gene expression controlled by an insulator-shielded PPT sequence is a promising candidate for gene therapy of prostate cancer. PMID- 15294183 TI - Ribozyme-mediated induction of apoptosis in human cancer cells by targeted repair of mutant p53 RNA. AB - A variety of mutations in the p53 tumor suppressor gene have been found in over half of human tumors. Thus, restoration of wild-type p53 activity by repair of mutant RNA has been previously suggested as an approach to cancer treatment. To explore the potential utility of RNA repair for cancer therapy, we developed a group I intron-based ribozyme that can replace mutant p53 RNA with a wild-type RNA sequence attached to the 3' end of the ribozyme by trans-splicing reaction. First, RNA mapping analysis demonstrated that the leader sequences upstream of the AUG start codon in the mutant p53 RNA appeared to be particularly accessible to the ribozymes. Then, the trans-splicing ribozyme specifically recognizing the most accessible sequence induced functional p53 activity, resulting in an 8- and a 2.6-fold induction of transactivation of p53-responsive promoters in two mutant p53-related ovarian cancer cell lines, SKOV3 cells and 2774 cells, respectively, by repairing defective p53 RNA. The repair efficiency of the mutant p53 RNA was almost 10% in 2774 cells. Moreover, the ribozyme activated the expression level of endogenous p21 and Bax genes in the cells. Furthermore, apoptosis was efficiently triggered in the human cancer cells transfected with the specific ribozyme. These results suggest that a trans-splicing ribozyme could be a potent anti-cancer agent that can revert the defective p53-related neoplastic phenotype. PMID- 15294184 TI - Macropinocytosis of polyplexes and recycling of plasmid via the clathrin dependent pathway impair the transfection efficiency of human hepatocarcinoma cells. AB - Knowledge of the entry mechanism and intracellular routing of polyplexes is of major importance for designing efficient gene delivery systems. We therefore investigated the internalization and trafficking of polyplexes in HepG2 cells. pDNA encoding the luciferase was complexed with histidylated polylysine (His pLK), a polymer that requires acidic pH for pDNA endosomal release. Fluoresceinylated polyplexes (F-His-pLK or F-pDNA) were internalized by clathrin dependent and -independent pathways. The latter most likely occurred by macropinocytosis since it was stimulated by phorbol myristate and blocked by dimethylamiloride. Intracellular routing of the plasmid was analyzed by confocal microscopy and flow cytometry. These data revealed that: (i) one part of the plasmid was present in vesicles that were not labeled with any known organelle specific marker, (ii) the other part was in transferrin receptor-positive vesicles, and (iii) the plasmid was not transferred to late endosomes/lysosomes. Using luciferase activity as a readout for gene expression, we found that it was strongly reduced when macropinocytosis was stimulated, whereas macropinocytosis inhibitors had no effect. However, blocking clathrin-dependent internalization by chlorpromazine completely prevented gene expression. These findings demonstrate that: (i) macropinocytosis of polyplexes and (ii) plasmid recycling impair the transfection efficiency and (iii) clathrin-dependent endocytosis is the most productive route for transfection of HepG2 cells. PMID- 15294185 TI - A facile nonviral method for delivering genes and siRNAs to skeletal muscle of mammalian limbs. AB - Delivery is increasingly being recognized as the critical hurdle holding back the tremendous promise of nucleic acid-based therapies that include gene therapy and more recently siRNA-based therapeutics. While numerous candidate genes (and siRNA sequences) with therapeutic potential have been identified, their utility has not yet been realized because of inefficient and/or unsafe delivery technologies. We now describe an intravascular, nonviral methodology that enables efficient and repeatable delivery of nucleic acids to muscle cells (myofibers) throughout the limb muscles of mammals. The procedure involves the injection of naked plasmid DNA or siRNA into a distal vein of a limb that is transiently isolated by a tourniquet or blood pressure cuff. Nucleic acid delivery to myofibers is facilitated by its rapid injection in sufficient volume to enable extravasation of the nucleic acid solution into muscle tissue. High levels of transgene expression in skeletal muscle were achieved in both small and large animals with minimal toxicity. Evidence of siRNA delivery to limb muscle was also obtained. The simplicity, effectiveness, and safety of the procedure make this methodology well suited to limb muscle gene therapy applications. PMID- 15294186 TI - Parvalbumin gene delivery improves diastolic function in the aged myocardium in vivo. AB - Abnormal relaxation of the heart, termed diastolic dysfunction, is a significant and growing problem that is a major cause of heart failure in the aged population. The potential of gene transfer of parvalbumin (Parv), a cytoplasmic calcium-binding protein, to improve diastolic function in the aged myocardium in vivo was evaluated. Despite evidence for an early developmental influence on the efficiency of Ad5 striated muscle transduction, results show that Ad5 gene transfer efficiency to adult cardiac myocytes in vitro is identical in young and old rats, suggesting that the basic processes of adenovirus binding and internalization are unaffected by aging. In contrast, Ad5-mediated Parv gene transfer to the myocardium in vivo is reduced in old rats compared to young rats. Nonetheless, Parv gene transfer and expression in vivo were sufficient to improve tau, a load-independent indicator of diastolic function, assessed using catheter based micromanometry in the aged myocardium. These results suggest that expression of the calcium buffer Parv may represent an effective approach to functional correction of the failing heart in the aging. PMID- 15294188 TI - Therapeutic apheresis in low weight patients: technical feasibility, tolerance, compliance, and risks. AB - The use of therapeutic apheresis in very low weight patients is generally thought to have limitations, because of possible severe adverse reactions, potential risk related to the extracorporeal procedure, due to the low weight of the young patients. A careful therapeutic approach using appropriate precautions, and also introducing modifications to the standard procedure, can minimise the risk without compromising the efficacy of the plasmapheresis. The aim of the study was to evaluate apheresis tolerance and acceptability in children [Artif. Organs. 21 (1997) 1126] and infants [J. Clin. Apheresis 5 (1989) 21] with inherited lipid metabolism disorder, familial hypercholesterolemia (FH), primary hyperlipoproteinemia (lipoprotein phenotype I), and acute leukemia, weighing on average 20.55 kg. One thousand one hundred twenty three aphereses were completed. Three types of apheresis were performed: leukapheresis, plasma exchange, dextran sulphate cellulose (DSC) low density lipoprotein (LDL)-apheresis. Three different types of continuous flow systems were used. Technical adaptation depending on patients blood volume, body mass index, hematocrit, type of system used, permitted us to perform complete aphereses, obtaining a high degree of tolerance and acceptability of the treatment. The use of plasmapheresis is regarded to be an extreme therapeutic measure in children. However, when the need for such treatment is undebatable, plasmapheresis must be done. A well-trained and experienced team can overcome the technical difficulties in order to complete the procedures without complications. The most frequently observed adverse effects are vascular relative access insufficiency (2.0%), and mild hypotension (2.0%). PMID- 15294189 TI - Transferrin receptor in serum. A new tool in the diagnosis and prevention of iron deficiency in blood donors. AB - BACKGROUND: Transferrin receptor mediates cellular uptake of iron, and the expression on cells reflects iron needs and erythropoietic activity. The results of measuring transferrin receptor in serum (sTfR) in blood donors are presented. STUDY DESIGN AND METHODS: Haemoglobin, serum-ferritin and sTfR were measured in 172 female and 174 male donors that had donated whole blood six or more times during the previous 3 years and in 96 female and 56 male new donors. RESULTS: Haemoglobin and sTfR were not significant different in new and repeat donors. New donors had significantly higher s-ferritin than repeat donors. Twenty donors had a Hb above the low limit for normal, but below the determined cut-off for donation. Only three of these had high sTfR and/or low serum-ferritin. Hence, of the total 492 donors 3.5% were below the Hb cut-off, but having Hb, s-ferritin and sTfR within normal ranges. 11.6% of new female donors belonged in this category. CONCLUSION: STfR is better than s-ferritin as a screening for iron deficiency. Most donors with low tissue iron neither have high sTfR, nor anaemia. There is probably no need to have a separate, higher than the lower normal range, requirement for Hb in donors. STfR measurements are probably most valuable in a setting where most donors are repeat donors. PMID- 15294190 TI - Systemic lupus erythematosus and thrombotic thrombocytopenic purpura: a case report. AB - BACKGROUND: Thrombotic thrombocytopenic purpura (TTP) is a multisystem disorder characterised by platelet aggregation causing microvascular occlusion. Early diagnosis and utilization of plasmapheresis can provide an improvement in prognosis. CASE REPORT: A 17 year old male with classical findings of TTP was later diagnosed as systemic lupus erythematosus (SLE). Plasmapheresis resulted in the significant amelioration of the course. CONCLUSION: The coexistence of TTP and SLE may facilitate a better understanding of in the pathophysiology of TTP. These association may provide the role of autoimmunity in TTP. SLE should be considered in the differential diagnosis of patients with TTP because of therapeutic implications. PMID- 15294191 TI - A novel method of preoperative autologous blood donation with a large volume of plasma for surgery in gynecologic malignancies. AB - The objective of this study was to establish a novel method of preoperative autologous blood donation (PAD) for surgery of gynecologic malignancies, which requires considerable amounts of plasma relative to the red blood cell component. To collect a double volume of plasma over the amount obtained from whole blood without using an aphaeresis system, we first collected 500 ml of whole blood (2.5 units), and centrifuged it. We gave back the resultant red cell component alone, and retained the plasma component. We further collected an additional 500 ml of whole blood, and centrifuged it. The red cell component (2.5 units) was stored in the refrigerator (as a concentrated red cell, CRC). The resultant plasma together with the plasma collected first (5 units) was frozen and stored in the freezer (fresh frozen plasma, FFP), We repeated this procedure at most three times at intervals of 1 week. Erythropoietin was injected once a week and iron tablets were prescribed. Ninety-nine patients undergoing surgery for a gynecological malignancy were subjected to this method and 86 patients without PAD served as a control. We conducted the procedure for PAD without any noticeable side effects. The amount of actual use of allogeneic CRC and FFP were significantly reduced in the PAD group compared with the control group. In particular, 93.6% of the PAD cases who gave 10 or less units of FFP could go without allogeneic FFP. Postoperative serum albumin levels were higher in the PAD group compared with the control. We have established a novel PAD method which can yield a greater volume of FFP relative to CRC, thus meeting requirements for surgery for gynecological malignancies. PMID- 15294192 TI - Application of noninvasive phagocytic cellular assays using autologous monocytes to assess red cell alloantibodies in sickle cell patients. AB - We investigated red cell (RBC) alloantibodies in 125 sickle cell anemia (SCA) patients using tube indirect antiglobulin test (PEG, LISS or enzyme) and gel centrifugation test (LISS or enzyme). Prediction of clinical significance of alloantibodies was evaluated by the monocyte monolayer assay (MMA) and the chemiluminescence test (CLT) using autologous monocytes. The alloimmunization rate was 20.8% and the gel test detected a higher number of alloantibodies than the tube test (26 v 21, p = 0.02). We observed 58.3% and 69.2% positive MMA and CLT results, respectively. Eighteen (69.2%) antibodies exhibited clinical relevance, 14 (58.3%) antibodies reacted by both MMA and CLT, while 4 (15.4%) antibodies reacted only by CLT. In conclusion, the application of phagocytic cellular assays using autologous monocytes defined clinical significance of about 70% of RBC alloantibodies detected in SCA patients. The data also suggest that the CLT may be more valuable than the MMA as a noninvasive test for predicting hemolysis after transfusion of incompatible blood in SCA patients. PMID- 15294193 TI - Specific aspects of transfusion therapy in cardiovascular surgery. PMID- 15294194 TI - A novel role for an established player: anemia drug erythropoietin for the treatment of cerebral hypoxia/ischemia. AB - Erythropoietin, a hematopoietic growth-factor possessing manifold, potent neuroprotective properties, after multiple testing in cell culture and animal studies now gradually finds its way into clinical neuroscience. The first time this took place was in 1998 with a pilot study in stroke patients, the "Gottingen EPO-Stroke-Trial". This study was able to demonstrate that EPO is perfectly well tolerated and safe with this indication. Furthermore, the EPO-treated patients showed a significantly better outcome regarding their clinical progress as well as regarding the infarct size as observed by MRI, when compared to the placebo treated patients. At the moment a multicenter study is being carried out in Germany. PMID- 15294195 TI - Tissue oxygenation and capacity to deliver O2 do the two go together? AB - Oxidative phosphorylation is the most important source of energy in mammals. Oxygen capture, convective and diffusive oxygen transport as well as the final intracellular oxygen utilization within the mitochondria represent highly refined mechanisms, supervised by a variety of physiological control systems. Any disease process interfering with the delivery of oxygen to tissue will ultimately lead to an impairment of cellular energy production. Generally, cellular hypoxia may result from either reduced oxygen uptake (hypoxic hypoxia), reduced convective and diffusive oxygen transport (circulatory and anemic hypoxia), impaired oxygen consumption (histotoxic hypoxia), or a combination of these states. To effectively treat any of these conditions, it is mandatory to recognize the underlying specific alterations of oxidative metabolism. Identification of the various types of hypoxia as well as contemporary treatment surveillance strategies depend primarily on measuring oxygen partial pressure in inspiratory gas, blood (arterial, mixed-venous) and tissue (extracellular fluid), next to monitoring of various circulatory parameters. This review focuses (a) on the diagnostic value of different techniques used to monitor blood and tissue oxygenation and (b) on the effects of impaired capacity to deliver O2 on tissue oxygen delivery and consumption. The potential value of multiparametric monitoring in guiding specific treatment measures to improve oxygen delivery to tissue is highlighted. PMID- 15294196 TI - Current status of transfusion triggers for red blood cell concentrates. AB - Clinical practice guidelines on red blood cell transfusion (RBC) are based on expert opinion, animal studies and the few human trials available. Twelve randomized controlled trials on the benefits of RBC transfusions in humans have been published. In the absence of definitive outcome studies, numerous theoretical arguments have been put forward in favor or against the classic transfusion threshold of 100 g/l. However, data from randomized controlled trials suggest that overall morbidity (including cardiac) and mortality, hemodynamic, pulmonary and oxygen transport variables are not different between restrictive (transfusion threshold between 70 and 80 g/l) and liberal transfusion strategies and that a restrictive transfusion strategy is not associated with increased adverse outcomes. In fact, a restrictive strategy may be associated with decreased adverse outcomes in younger and less sick critical care patients. The majority of existing guidelines conclude that transfusion is rarely indicated when the hemoglobin concentration is greater than 100 g/l and is almost always indicated when it falls below a threshold of 60 g/l in healthy, stable patients or more in older, sicker patients. In anesthetized patients, this threshold should be modulated by factors related to the dynamic nature of surgery such as uncontrolled hemorrhage, microvascular bleeding, etc. Another important role of RBC relates to primary hemostasis and higher triggers may be appropriate in coagulopathic patients. RBC concentrates are administered to correct inadequate oxygen delivery and/or to sustain primary hemostasis. Reliable monitors of tissue oxygenation and hemostasis will be required to study the benefits (or lack thereof) of RBC transfusions. The quest for a universal transfusion trigger, i.e., one that would be applicable to patients of all ages under all circumstances, must be abandoned. All RBC transfusions must be tailored to the patient's needs, at the moment the need arises. In conclusion published recommendations are commensurate with existing knowledge and, unfortunately, their conclusions are limited. Future research and development should focus on the determination of optimal transfusion strategies in various patient populations and on reliable monitors to guide transfusion therapy. PMID- 15294197 TI - What is happening? Are the current acceptance criteria for therapeutic plasma adequate? AB - The main objective of this overview is to highlight the impact of new developments in the processing of plasma, on the quality, safety and efficacy fresh frozen plasma (FFP). Based on the currently available information, it is suggested that current guidelines need an urgent review to include more relevant criteria of acceptability for therapeutic plasma. Moreover additional streamlining of FFP production, using selective donor panel, will improve the uniformity of therapeutic plasma and lower the potential risks of transfusion reactions. PMID- 15294198 TI - Recombinant coagulation factor VIIa in the management of bleeding in patients with von Willebrand disease type 2A. PMID- 15294199 TI - The clinical neurophysiology of the restless legs syndrome and periodic limb movements. Part I: diagnosis, assessment, and characterization. AB - OBJECTIVE: The restless legs syndrome is a common sensorimotor disorder impacting on sleep which has been known for centuries, but only recently become recognized as a significant clinical and pathophysiological problem. The definition of RLS has evolved until certain key clinical features have been defined as diagnostic, while others are strongly associated: the urge to move is seen as primary. Epidemiology suggests ethnic variation with highest frequency in populations of European origin; family and genetic studies support a genetic basis to many idiopathic cases while links to secondary disorders usually involving low iron stores are also known. Abnormalities of brain iron transport and consequent dysfunction of the dopamine system are suspected sources of the disorder. METHODS: The literature was searched for all references relating clinical neurophysiologic investigations to the diagnosis, assessment, and characterization of RLS. RESULTS: RLS is defined clinically and diagnosed by medical history while its frequent concomitant, periodic limb movements (PLM), must be diagnosed by polysomnography or movement recording. Severity of RLS is generally assessed by subjective measures, but sleep recording and measurement of PLM frequency and association with sleep disruption are also used to measure severity. A provocative test, the suggested immobilization test, can also be used with both subjective and movement recording. RLS and PLM in RLS are both associated with the circadian cycle and are maximal early in the sleep period. PLM appear to be associated both with unstable EEG phases involving the cyclic alternating pattern and cyclical autonomic changes whose initiation may precede the muscle activity. CONCLUSIONS: While RLS remains a subjective disorder, neurophysiologic measures have been important, especially for assessment. Ambulatory methodologies may offer the most accurate and economical means of assessing motor activity as a key marker of RLS and of accurately measuring PLM from night to night. As the pathophysiology of RLS is better understood, more focused techniques may be developed to measure its presence and severity in individual patients. PMID- 15294200 TI - Why do restless legs occur at rest?--pathophysiology of neuronal structures in RLS. Neurophysiology of RLS (part 2). AB - Restless legs syndrome (RLS) is a heterogeneous disorder encompassing genetically caused types with early onset and acquired varieties occurring later in life. Genetic studies in the near future will most likely discover more than one causative gene. The acquired cases too have different etiologies ranging from idiopathic types to secondary forms with uremia, iron depletion, polyneuropathy and others. Here we aim to correlate typical RLS symptoms, such as the sensory symptoms at rest, the reduction of the complaint in response to movement or other physical stimuli, the dominant involvement of the legs, pain, circadian rhythm, and the responsiveness to dopaminergic drugs with neurophysiological features of the central nervous system. We outline the complexity of the neural structures involved and their connections. A diversity of hypothetical affections of different neuronal levels might lead to various combinations of RLS symptomatology. No single pathophysiological explanation has yet been developed that covers all clinical features. PMID- 15294201 TI - Generalization of the generation of an MMN by illusory McGurk percepts: voiceless consonants. AB - OBJECTIVE: The existence of a Mismatch Negativity (MMN) evoked by McGurk percepts elicited by audiovisual syllables with constant auditory components has been previously demonstrated with voiced consonants [Clin. Neurophysiol. 113 (2002) 495]. The present study aimed at generalizing such results with voiceless consonants. In a first experiment, the MMN was computed using the classical subtraction method (standard minus deviant). Since results showed a possible contamination by exogenous visual components, a technique preventing from including those components in the differential waveform was used in a second experiment (deviant in sequence minus deviant presented alone). METHODS: Cortical evoked potentials were recorded using the oddball paradigm on eight adults in three experimental conditions (auditory alone, visual alone and audiovisual) for experiment one and in two conditions (visual alone and audiovisual) for experiment two. Obtaining illusory percepts was confirmed in additional psychophysical experiments. RESULTS: Significant MMNs were recorded in the three conditions of experiment one, whereas only the audiovisual condition of experiment two gave rise to a significant MMN. CONCLUSIONS: Provided that the MMN is computed with deviant stimuli only, the present results confirm the elicitation of genuine audiovisual MMN. Possible refractoriness effects and N2b confound have, however, to be controlled for in further studies. PMID- 15294202 TI - Electrophysiological correlates of attention, inhibition, sensitivity and bias in a continuous performance task. AB - OBJECTIVE: The aim was to verify the occurrence of proposed electrophysiological correlates of attention, inhibition, sensitivity and bias in a continuous performance task and to support their functional interpretation by using a manipulation intended to enhance subjects' response bias. METHODS: Electroencephalographic activity was recorded during administration of a transformed version of the AX continuous performance task in which cues signaled response alternatives. RESULTS: The previously reported parietal P3, NoGo-N2, NoGo-P3 and contingent negative variation were replicated. Besides, the frontal selection positivity and the lateralized readiness potential were demonstrated. With increasing Go-probability, the parietal P3 to the cue increased without changes in other cue-related correlates. In addition, reaction times decreased, non-parametric measures of sensitivity and bias decreased, the NoGo-N2 increased, and the parietal Go-P3 decreased. CONCLUSIONS: The proposed electrophysiological correlates were identified. Sub-threshold LRPs suggested a central inhibition mechanism. Cue-related correlates revealed that anticipation of a high probability Go-stimulus involves attention rather than bias. This implies that the increased NoGo-N2 reflected an increase in conflict rather than an increase in inhibition. SIGNIFICANCE: Electrophysiological measures can greatly enhance our understanding of normal and abnormal information processing during continuous performance and related tasks. PMID- 15294203 TI - Response monitoring without sensory feedback. AB - OBJECTIVE: The elicitation of an evoked potential, the 'error negativity' (Ne) when subjects commit errors in speeded tasks, is often taken as an index of response monitoring processes. The presence of a Ne-like wave on purely correct trials challenges the current conceptions about the nature of such a monitoring system. Here, we evaluate the possibility that the Ne-like wave on correct trials is merely due to reafferences, and at the same time, we test directly the general opinion according to which the Ne is generated by an internal signal. METHODS: We studied the presence of a Ne-like wave in a completely deafferented patient. The patient performed two reaction time (RT) tasks: a two-responses RT task and a go/no-go task. RESULTS: In this patient, a Ne occurs on errors, on incorrect EMG activations, and on purely correct responses. On errors, the Ne was clearly followed by an error positivity (Pe). CONCLUSIONS: The Ne and the Ne-like wave are not generated by reafferences. This similarity is a further argument to consider that these two waves are of same nature. SIGNIFICANCE: The present data demonstrate that sensory information is not mandatory for the brain to monitor and correct ongoing responses. PMID- 15294204 TI - Brainstem responses to speech syllables. AB - OBJECTIVE: To establish reliable procedures and normative values to quantify brainstem encoding of speech sounds. METHODS: Auditory brainstem responses to speech syllables presented in quiet and in background noise were obtained from 38 normal children. Brainstem responses consist of transient and sustained, periodic components-much like the speech signal itself. Transient peak responses were analyzed with measures of latency, amplitude, area, and slope. Magnitude of sustained, periodic frequency-following responses was assessed with root mean square, fundamental frequency, and first formant amplitudes; timing was assessed by stimulus-to-response and quiet-to-noise inter-response correlations. RESULTS: Measures of transient and sustained components of the brainstem response to speech syllables were reliably obtained with high test-retest stability and low variability across subjects. All components of the brainstem response were robust in quiet. Background noise disrupted the transient responses whereas the sustained response was more resistant to the deleterious effects of noise. CONCLUSIONS: The speech-evoked brainstem response faithfully reflects many acoustic properties of the speech signal. Procedures to quantitatively describe it have been developed. SIGNIFICANCE: Accurate and precise manifestation of stimulus timing at the auditory brainstem is a hallmark of the normal perceptual system. The brainstem response to speech sounds provides a mechanism for understanding the neural bases of normal and deficient attention-independent auditory function. PMID- 15294205 TI - Interdependency between heart rate variability and sleep EEG: linear/non-linear? AB - OBJECTIVE: To investigate whether the interdependency between heart rate variability (HRV) and sleep electroencephalogram (EEG) power spectra is linear or non-linear. METHODS: Heart rate and sleep EEG signals were recorded in 8 healthy young men. Spectral analysis was applied to electrocardiogram and EEG sleep recordings. Synchronization likelihood was computed over the first 3 non-rapid eye movement-rapid eye movement sleep cycles between normalized high frequency of RR intervals (RRI) and all electroencephalographic frequency bands. Comparison to surrogate data of different types was used to attest statistical significance of the coupling between RRI and EEG power bands and its linear or non-linear character. RESULTS: Synchronization likelihood values were statistically greater than univariate surrogate synchronization for all sleep bands both at the individual and the group levels. With reference to multivariate surrogates, synchronization values were statistically greater at the group level and, in a majority of cases, for individual comparison except for sigma and beta bands. CONCLUSIONS: While all electroencephalographic power bands are linked to normalized high frequency RRI band, this interdependency is non-linear for delta, theta and alpha bands. SIGNIFICANCE: Non-linear description is required to capture the full interdependent dynamics of HRV and sleep EEG power bands. PMID- 15294206 TI - Magnetoencephalographic spikes not detected by conventional electroencephalography. AB - OBJECTIVE: To investigate some of the reasons why magnetoencephalographic (MEG) spikes are at times not apparent in conventional electroencephalograms (EEG) when the data are co-registered, and to explore to what extent modern EEG analysis methods can improve the yield. METHODS: Seventy seconds of MEG-EEG co registration on a 122 channel Neuromag system were studied in a 10-year-old boy with Landau-Kleffner syndrome. Twenty-six EEG channels were originally recorded with a left ear reference. The EEG data were subsequently reformatted (BESA) to a variety of montages for the 10-20 and 10-10 electrode array. A 10 s data epoch was compared in detail for concordance between MEG and EEG spikes. To detect the characteristics of hidden low voltage EEG spikes, MEG spikes were averaged and compared with the concomitant averaged EEG spike. RESULTS: While there was an abundance of EEG as well as MEG spikes on the left; definite right-sided spikes were not visible in the EEG. Right hemispheric MEG spikes were, however, plentiful with an average strength of 757 fT. When the individual MEG spikes from the right hemisphere were compared with the corresponding EEG events their amplitude ranged between 24 and 31 microV and were, therefore, indistinguishable from background activity. The majority of them became visible, however, with further sophisticated data analysis. CONCLUSIONS: When the relative merits of MEG versus EEG recordings for the detection of epileptogenic spike are investigated the 10-20 system of electrode placement and conventional methods of EEG analysis do not provide optimal data assessment. The use of the 10-10 electrode array combined with modern methods of digital data analysis can provide better concordance with MEG data. PMID- 15294207 TI - Abnormal EEG synchronisation in heavily drinking students. AB - OBJECTIVE: In alcoholics, grey and white brain matter is damaged. In addition, functional brain connectivity as measured by EEG coherence is abnormal. We investigated whether heavily drinking students, although drinking for a shorter period than alcoholics, already show differences in functional connectivity compared to light-drinking controls. METHODS: EEG was recorded in 11 light and 11 heavy male student drinkers during eyes closed, and eyes closed plus mental rehearsal of pictures. Functional connectivity was assessed with the Synchronisation Likelihood method. RESULTS: Heavily drinking students had more synchronisation in the theta (4-8 Hz) and gamma (30-45 Hz) band than lightly drinking students during eyes closed, both with and without a mental-rehearsal task. CONCLUSIONS: Heavy student drinkers have increases in EEG synchronisation that are indicative of changes in hippocampal-neocortical connectivity. SIGNIFICANCE: Heavy student drinkers show differences in functional connectivity as compared to their lightly drinking counterparts, even though they have a relatively short drinking history. PMID- 15294208 TI - Subregions of human MT complex revealed by comparative MEG and direct electrocorticographic recordings. AB - OBJECTIVE: To locate the visual motion complex (MT+) and study its response properties in an epilepsy surgery patient. METHODS: A 17-year-old epilepsy patient underwent invasive monitoring with subdural electrodes in the right temporo-parieto-occipital area. MT+ was investigated by cortical electric stimulation and by epicortical visual evoked potentials time-locked to motion onset of sinusoidal gratings (motion VEP). Motion-related visual evoked magnetic field (motion VEF) was also recorded before the electrode implantation to complement the invasive recording. RESULTS: Motion VEPs revealed two subregions within MT+, generating early and late potentials respectively. The early activity with a peak around 130 ms was localized at a single electrode situated immediately caudal to the initial portion of the ascending limb of the superior temporal sulcus (AL-STS). The late activity, peaking at 242-274 ms, was located ventro-rostrally over three electrodes. Among the four electrodes with motion VEPs, cortical stimulation at the most caudal pair elicited motion-in-depth perception involving the whole visual field. In addition to two subregions revealed on the gyral crown, magnetoencephalography (MEG) demonstrated another subregion with a late motion VEF in AL-STS immediately rostral to the electrode with the early motion VEP. CONCLUSIONS: In combination with MEG recording, the present invasive exploration demonstrated human MT+ in a focal area of the temporo-parieto-occipital junction and delineated possible three subregions as indicated by the different latencies and distributions of the motion VEP/VEFs. SIGNIFICANCE: Comparative MEG and direct electrocorticographic recordings delineated possible subregions within the human MT complex. PMID- 15294209 TI - Magnetoencephalography source localization and surgical outcome in temporal lobe epilepsy. AB - OBJECTIVE: We prospectively investigated the role of magnetoencephalography (MEG) in localizing the seizure focus and in predicting outcome to surgical resections for intractable temporal lobe epilepsy (TLE). METHODS: We performed simultaneous interictal EEG and MEG recording (two 37-channel system) in 26 TLE patients followed by MEG source localization. We correlated early modeling dipoles with intracranial EEG, temporal surgical resection and surgical outcome. RESULTS: There were 12 patients who had anterior temporal horizontal or tangential dipoles to the anterior infero-lateral temporal tip cortex. Two patients underwent selective amygdalo-hippocampectomy (SAH) and nine patients had antero-medial temporal lobectomy (AMTL). All patients had successful outcome except for one patient who initially failed SAH, but became seizure-free after AMTL. There were 11 patients who demonstrated anterior temporal vertical or tangential oblique dipoles. Five patients had AMTL and three had SAH; all became seizure free. Five of above 23 patients had invasive EEG and demonstrated mesial seizure onset. Three TLE patients had lateral vertical dipoles that were concordant with intracranial EEG and these became seizure free after temporal neocortical resections. CONCLUSIONS: MEG source analysis produces distinct source patterns that provide useful localizing information, predict surgical outcome, and may aid in planning limited surgical resection in TLE. PMID- 15294210 TI - Origin, structure, and role of background EEG activity. Part 1. Analytic amplitude. AB - OBJECTIVE: To explain the neural mechanisms of spontaneous EEG by measuring the spatiotemporal patterns of synchrony among beta-gamma oscillations during perception. METHODS: EEGs were measured from 8 x 8 (5.6 x 5.6 mm2) arrays fixed on the surfaces of primary sensory areas in rabbits that were trained to discriminate visual, auditory or tactile conditioned stimuli (CSs) eliciting conditioned responses (CRs). EEG preprocessing was by (i) bandpass filtering to extract the beta-gamma range (deleting theta-alpha); (ii) low-pass spatial filtering (not high-pass Laplacians used for localization), (iii) spatial averaging (not time averaging used for evoked potentials), and (iv) close spacing of 64 electrodes for simultaneous recording in each area (not sampling single signals from several areas); (v) novel algorithms were devised to measure synchrony and spatial pattern stability by calculating variances among patterns in 64-space derived from the 8 x 8 arrays (not by fitting equivalent dipoles). These methodological differences are crucial for the proposed new perspective on EEG. RESULTS: Spatial patterns of beta-gamma EEG emerged following sudden jumps in cortical activity called 'state transitions'. Each transition began with an abrupt phase re-setting to a new value on every channel, followed sequentially by re-synchronization, spatial pattern stabilization, and a dramatic increase in pattern amplitude. State transitions recurred at varying intervals in the theta range. A novel parameter was devised to estimate the perceptual information in the beta-gamma EEG, which disclosed 2-4 patterns with high information content in the CS-CR interval on each trial; each began with a state transition and lasted approximately 0.1 s. CONCLUSIONS: The function of each primary sensory neocortex was discontinuous; discrete spatial patterns occurred in frames like those in cinema. The frames before and after the CS-CR interval had low content. SIGNIFICANCE: Derivation and interpretation of unit data in studies of perception might benefit from using multichannel EEG recordings to define distinctive epochs that are demarcated by state transitions of neocortical dynamics in the CS-CR intervals, particularly in consideration of the possibility that EEG may reveal recurring episodes of exchange and sharing of perceptual information among multiple sensory cortices. Simultaneously recorded, multichannel beta-gamma EEG might assist in the interpretation of images derived by fMRI, since high beta gamma EEG amplitudes imply high rates of energy utilization. The spatial pattern intermittency provides a tag to distinguish gamma bursts from contaminating EMG activity in scalp recording in order to establish beta-gamma recording as a standard clinical tool. Finally, EEG cannot fail to have a major impact on brain theory. PMID- 15294211 TI - Origin, structure, and role of background EEG activity. Part 2. Analytic phase. AB - OBJECTIVE: To explain spontaneous EEG through measurements of spatiotemporal patterns of phase among beta-gamma oscillations. METHODS: High-density 8 x 8 intracranial arrays were fixed over sensory cortices of rabbits. EEGs were spatially low pass filtered, temporally bandpass filtered and segmented in overlapping windows stepped at 2 ms. Phase was measured with the cosine as the temporal basis function, using both Fourier and Hilbert transforms to compensate for their respective limitations. Spatial patterns in 2D phase surfaces were measured with the geometric form of the cone as the spatial basis function. RESULTS: Two fundamental state variables were measured at each digitizing step in the 64 EEGs: the rate of change in phase with time (frequency) and the rate of change in phase with distance (gradient). The parameters of location, diameter, duration, and phase velocity of the cone of phase were derived from these two state variables. Parameter distributions including recurrence intervals extending into the low theta range were fractal; the mean values varied with window duration and interelectrode distance. CONCLUSIONS: The formation of spatial amplitude patterns began with state transitions that were documented by phase discontinuities and phase cones. The multiplicity of overlapping cones indicated that sensory neocortices maintained a scale-free state of self-organized criticality (SOC) in each hemisphere as the basis for its rapid integration of sensory input with prior learning stored in cortical synaptic webs. Further evidence came from the fractal properties of the phase parameters and the self similarity of phase patterns in the ms/mm to m/s ranges. SIGNIFICANCE: These EEG data suggest that neocortical dynamics is analogous to the dynamics of self stabilizing systems, such as a sand pile that maintains its critical angle by avalanches, and a pan of boiling water that maintains its critical temperature by bubbles that release heat. Beta-gamma oscillations stem from the ability of neocortex to maintain its stability under continuous sensory bombardment. Modeling implies that the critical parameter of neocortex (analogous to angle of repose or temperature) is the mean firing rates of neurons that are homeostatically regulated by refractory periods everywhere at all times in cortex. The advantage of SOC in perception may be the ability it gives neocortex to generate instantaneous global state transitions (avalanches, bubbles) large enough to include the multiple sensory areas that are necessary to form gestalts (multisensory percepts). PMID- 15294212 TI - Dipole localization accuracy using grand-average EEG data sets. AB - OBJECTIVE: Dipole localization of grand-average event related potentials only give a tentative description of the estimated underlying neural sources. This study evaluates the differences in dipole solutions between individual and group average data sets using a standard realistic head model. METHODS: Auditory evoked potentials were recorded from 14 right-handed healthy subjects using a 64 electrode montage. Inverse dipole solutions were obtained for each individual data set, as well as for all individual responses averaged together (grand average). Differences in dipole solutions between individual and grand-average responses are reported. Simulations using a two dipole model with 15 different electrode sets are then used to investigate the effects of electrode misplacement and random noise on dipole localization. These effects are compared to those due to grand-averaging. RESULTS: The average differences in dipole locations between the individual and grand-averaged data sets were approximately 1.1 cm (SD=0.7 cm). This difference is larger than typical localization errors due to electrode misplacement and typical noise. CONCLUSIONS: Using a standard realistic head model, it is concluded that dipole solutions based on group-averaged EEG datasets are significantly different than those obtained using subject-specific data. PMID- 15294213 TI - Visualization of incomplete conduction block by neuromagnetic recording. AB - OBJECTIVE: We previously reported on evoked compound action magnetic fields (CAFs) in isolated sciatic nerves with complete conduction block. In this study, we examined evoked CAFs of the nerve with incomplete conduction block, which is clinically common. METHODS: Rabbits' isolated nerves were electrically stimulated in a chamber containing Ringer's solution. Compound action potentials (CAPs) and CAFs were recorded before and after the incomplete conduction block induced by a vascular clip. The positions of the lesion were estimated by dipole localization. RESULTS: Before the nerve clipping, magnetic contour maps showed CAFs with a characteristic quadrupolar pattern. After the clipping, CAFs attenuated in the amplitude and decelerated through the lesion. Estimated position of the lesion was 0.12+/-3.23 mm (mean+/-SD, n=10) assuming that the real position of the clip was 0 mm. CONCLUSIONS: The time-course of changes of CAFs in the incomplete conduction block was visualized by magnetic contour maps, and the lesions were closely localized focusing on the velocity change of the leading dipole. SIGNIFICANCE: The neural conduction with incomplete conduction block was visualized and the lesion was closely localized by neuromagnetic recordings. PMID- 15294214 TI - Interside comparison of cutaneous silent periods in thenar muscles of healthy male and female subjects. AB - OBJECTIVE: The cutaneous silent period (CSP) is a spinal inhibitory reflex mediated by A-delta fibers. To date, no data are available about normal interside differences. METHODS: Twenty healthy subjects underwent comparison of CSPs in the dominant and non-dominant hand. Surface electromyographic (EMG) recordings were obtained from thenar muscles on either side following ipsilateral recurrent nociceptive digit II stimulation. RESULTS: Group average CSP onset and end latency, CSP duration, and the magnitude of EMG suppression were not significantly different between both sides. Regression analysis revealed powerful correlations between individual CSP parameters of dominant and non-dominant hands. Calculated upper normal limits of maximum interside differences were 17% for CSP onset latency, 14% for CSP end latency, 22% for CSP duration, and 45% for the index of suppression. CSP parameters in right-handed subjects did not differ significantly from those in left-handed subjects. Female subjects tended to have shorter CSP onset latencies, longer CSP duration, and a smaller index of suppression, resulting in a larger overall suppression. CONCLUSIONS: CSPs prove to be robust nociceptive cutaneomuscular reflexes with little side-to-side difference. SIGNIFICANCE: The presented normative values of interside differences enable a more thorough comparison with patient data in various conditions. PMID- 15294215 TI - Post-exercise facilitation and depression of motor evoked potentials to transcranial magnetic stimulation: a study in multiple sclerosis. AB - OBJECTIVE: To evaluate motor cortex excitability changes by transcranial magnetic stimulation (TMS) following repetitive muscle contractions in patients with multiple sclerosis (MS); to state whether a typical pattern of post-exercise motor evoked potentials (MEPs) is related to clinical fatigue in MS. METHODS: In 41 patients with definite MS (32 with fatigue and 9 without fatigue according to Fatigue Severity Scale) and 13 controls, MEPs were recorded at rest: at baseline condition, following repetitive contractions until fatigue, and after fatigue, to evaluate post-exercise MEP facilitation (PEF) and depression (PED). RESULTS: After exercise, MEP amplitude significantly increased both in patients and controls (PEF). When fatigue set in, MEP amplitude was significantly reduced in normal subjects (PED), but not in patients. Post-exercise MEP findings were similar both in patients with and without fatigue. CONCLUSIONS: Our findings suggest an intracortical motor dysfunction following a voluntary contraction in MS patients, possibly due to failure of depression of facilitatory cortical circuits, or alternatively of inhibitory mechanisms. PMID- 15294216 TI - The influence of age on learning a locomotor task. AB - OBJECTIVE: Knowledge about locomotor task performance and learning in the elderly is important in optimizing rehabilitation strategies. The aim of this study was to evaluate differences between young and elderly subjects in the acquisition and performance of a precision locomotor task, with full and restricted vision. METHODS: The subjects walked on a treadmill and had to step as low as possible over an obstacle, without touching it. They received acoustic warning and feedback signals, indicating obstacle appearance and foot clearance, respectively. Full vision was provided during the first two runs and became restricted during the third run. The number of obstacle hits and adaptations in foot clearance, leg muscle activity, range of motion of leg joints and swing phase duration were assessed. RESULTS: With vision, the performance improved in both groups. Restricted vision reduced the task accuracy in both the young and the elderly. However, only the young subjects regained optimal foot clearance with practice. CONCLUSIONS: Elderly subjects rely more on visual control when acquiring and performing a precision locomotor task. We suggest that this is due to an impaired function of proprioceptive feedback mechanisms, which can replace visual information in young subjects. SIGNIFICANCE: In the elderly, therapeutical attention should be directed towards optimizing the use of the remaining proprioceptive inputs. PMID- 15294217 TI - Motor outcome after subcortical stroke correlates with the degree of cortical reorganization. AB - OBJECTIVE: The contribution of cortical reorganization to motor recovery after a subcortical stroke is uncertain. The purpose of the study was to investigate the relationship between changes in motor cortex organization, and the degree of motor function after a subcortical stroke. METHODS: Transcranial magnetic stimulation mapping of the corticomotor projection to the hand was performed in 27 patients who had suffered a subcortical ischemic stroke resulting in an upper limb motor deficit up to 23 years previously. Corticospinal conduction was assessed by measurements of motor evoked potential latency, amplitude and threshold. Motor function in the upper limb was assessed using the Motor Assessment Scale for Stroke and measurements of grip strength. RESULTS: Motor maps for the hand were displaced on the affected side relative to the unaffected side in 17 patients. In 10 of these patients in whom corticospinal conduction had normalized, there was a strong positive correlation between the magnitude of the map shift and grip strength in the affected hand (r=0.79; P=0.006). In the other seven patients with a map shift, in whom corticospinal conduction was still impaired, there was a tendency for a larger map area to be associated with better motor function, and in the group as a whole there was a correlation between map area and grip strength (r=0.52; P=0.005). CONCLUSIONS: The present findings provide evidence that the cortical plasticity and reorganization that occurs after a subcortical stroke is functionally significant and contributes to motor outcome. PMID- 15294218 TI - Tremor analysis in two normal cohorts. AB - OBJECTIVE: Quantitative tremor analyses using almost identical methods were compared between two independent large normal cohorts, to separate robust measures that may readily be used diagnostically from more critical ones needing lab-specific normalization. METHODS: Hand accelerometry and surface EMG from forearm flexors and extensors were recorded with (500 and 1000 g) and without weight loading under postural conditions in 117 and 67 normal volunteers in two different specialty centers for movement disorders in Germany. RESULTS: Tremor amplitude (total power) and frequency fell within a similar range but differed significantly. A significant reduction of tremor frequency under 1000 g weight load (>1 Hz), and a lack of rhythmic EMG activity at the tremor frequency in around 85-90% of the recordings were robust findings in both centers. CONCLUSIONS: The differences in frequency and total power indicate that these measures critically depend on the details of the recording conditions being slightly different between the two centers. Thus each lab needs to establish its own normative data. We estimate that at least 25 normal subjects have to be recorded to obtain normal values. The reduction of tremor frequency under load and lacking tremor-related EMG activity were well reproducible allowing a differentiation of physiological from low amplitude pathological tremor. SIGNIFICANCE: This study provides a framework for more standardized tremor analyses in clinical neurophysiology. PMID- 15294219 TI - Fluoxetine facilitates use-dependent excitability of human primary motor cortex. AB - OBJECTIVES: In poststroke patients, fluoxetine, a selective serotonin-reuptake inhibitor, as an adjunct to physical therapy provided a better functional recovery from motor deficits. The aim of this study was to investigate the effect of a single dose of 20 mg fluoxetine on motor learning and associated cortical changes in healthy right-handed subjects in order to get deeper insight into its facilitating influence on human motor cortex. METHODS: Subjects performed a motor task consisting of a simultaneous co-contraction of the abductor pollicis brevis (APB) and the deltoid muscle with and without fluoxetine in a placebo-controlled double-blinded crossover study design. Immediately before and after motor learning motor output maps of the APB muscle were assessed in order to get insight into plastic changes of the muscle representation. RESULTS: We found a significantly improved motor performance under both conditions without having substantial differences between placebo and fluoxetine. After the completion of the motor task there was a medial shift of the APB muscle motor output map. Only after the administration of fluoxetine the sum of MEP amplitudes (SOA) increased and the motor output map enlarged. CONCLUSIONS: These findings provide evidence for a use-dependent facilitating effect of fluoxetine on cortical excitability but not on motor performance. SIGNIFICANCE: Our findings are not in line with previous experiments in poststroke patients. However, long-term treatment with fluoxetine may additionally improve motor function by upregulating serotonergic receptors. Further studies investigating the influence of long-term treatment on cortical excitability and psychophysics may therefore provide deeper insight into a possible therapeutical efficiency of fluoxetine in poststroke patients. PMID- 15294220 TI - Response selection and motor areas: a behavioural and electrophysiological study. AB - OBJECTIVE: The involvement of the supplementary motor areas (SMAs) and primary motor areas (M1s) in motor processes was studied. METHODS: A between-hand choice and a simple reaction time (RT) task were mixed in a precueing paradigm. Laplacians were estimated by the source derivation method from the electroencephalogram recorded over the SMAs and M1s. RESULTS: RT was shorter in the simple than in the choice RT task. Response-locked averages showed a negative potential over M1 contralateral to the response and a positive wave over M1 ipsilateral. This ipsilateral positivity was much smaller in the simple than in the choice RT task, whereas the contralateral negativity was not different. A negativity preceding the activations of the M1s developed over the SMAs. This negativity was larger in the choice than in the simple RT task. CONCLUSIONS: In light of previous results, the present data confirm that, in between-hand choice tasks, response execution is implemented by an activation of the contralateral M1 and by an inhibition of the ipsilateral M1. SMAs and contralateral M1 appear hierarchically organized, the SMAs being more involved in response preparation and M1s in response execution. The task-dependent inhibition of ipsilateral M1 could reflect an active suppression of the erroneous response in the choice task. SIGNIFICANCE: The task context in which one movement is executed can affect the pattern of activities recorded over cortical motor structures. Cognitive context is of importance for understanding the nature of the motor command. PMID- 15294221 TI - MRI compatible EEG electrode system for routine use in the epilepsy monitoring unit and intensive care unit. AB - OBJECTIVE: We report on the development of an electroencephalographic (EEG) recording system that is Magnetic Resonance Imaging (MRI) compatible and can safely be left on the scalp during anatomical imaging or used to obtain simultaneous EEG and metabolic or hemodynamic data using functional imaging techniques such as functional MRI or MR spectroscopy. METHODS: We assembled a versatile EEG recording set-up with medically acceptable materials that contained no ferromagnetic components. It was tested for absence of excess heating and distortion of the image quality in a spherical phantom similar in size to average adult human head in a clinical 1.5 T GE scanner. After testing its safety in four volunteers, 100 consecutive patients from our epilepsy long-term monitoring unit were studied. RESULTS: There was no change in the temperature of the EEG electrode discs during the various anatomical MRI sequences used in our routine clinical studies (maximum temperature change was -0.45 degrees C with average head SAR<==1.6 W/Kg in the selected subjects) nor were there any reported complications in the others. The brain images were not distorted by the susceptibility artifact of the EEG electrodes. CONCLUSIONS: Our MRI compatible EEG set-up allows safe and artifact free brain imaging in 1.5T MR scanner with average SAR<==1.6 W/Kg. This EEG system can be used for EEG recording during anatomical MRI studies as well as functional imaging studies in patients requiring continuous EEG recordings. PMID- 15294222 TI - Removal of time-varying gradient artifacts from EEG data acquired during continuous fMRI. AB - OBJECTIVE: Recording low amplitude electroencephalography (EEG) signals in the face of large gradient artifacts generated by changing functional magnetic resonance imaging (fMRI) magnetic fields continues to be a challenge. We present a new method of removing gradient artifacts with time-varying waveforms, and evaluate it in continuous (non-interleaved) simultaneous EEG-fMRI experiments. METHODS: The current method consists of an analog filter, an EEG-fMRI timing error correction algorithm, and a temporal principal component analysis based gradient noise removal algorithm. We conducted a phantom experiment and a visual oddball experiment to evaluate the method. RESULTS: The results from the phantom experiment showed that the current method reduced the number of averaged samples required to obtain high correlation between injected and recovered signals, compared to a conventional average waveform subtraction method with adaptive noise cancelling. For the oddball experiment, the results obtained from the two methods were very similar, except that the current method resulted in a higher P300 amplitude when the number of averaged trials was small. CONCLUSIONS: The current method enabled us to obtain high quality EEGs in continuous simultaneous EEG-fMRI experiments. SIGNIFICANCE: Continuous simultaneous EEG-fMRI acquisition enables efficient use of data acquisition time and better monitoring of rare EEG events. PMID- 15294223 TI - BP and CNV. PMID- 15294225 TI - Photophysical properties of safranine O in protic solvents. AB - Spectroscopic and photophysical properties of safranine O (Sf) were investigated in binary water/solvent mixtures. It was found that these properties are strongly solvent-dependent. A blue shift is observed for both the ground-state absorption and the triplet-triplet main absorption band when the solvent polarity augments. At the same time a red shift of the fluorescence emission band takes place. These facts are interpreted in terms of higher dipole moment of the dye molecule in the S(1) state as compared with the S(0) state, while a decrease in the dipole moment of the triplet state T(n) with respect to the triplet state T(1) occurs. The Stokes' shift and the fluorescence lifetime shows a linear correlation with the E(T)(30) parameter, while a non-linear behavior is observed when a correlation with models of a continuous dielectric solvent is attempted. These results suggest the operation of strong specific interactions of Sf with solvent molecules, most likely hydrogen bonding. From fluorescence lifetime and quantum yield determinations, as well as intersystem-crossing quantum yields, the solvent dependence of the photophysical kinetic parameters were obtained. The radiative fluorescence rate constant can be adequately reproduced by calculations based on the UV-Vis absorption and emission spectra, as given by the Strickler-Berg equation. PMID- 15294226 TI - EPR, optical, infrared and Raman spectral studies of Actinolite mineral. AB - Electron paramagnetic resonance (EPR), optical, infrared and Raman spectral studies have been performed on a natural Actinolite mineral. The room temperature EPR spectrum reveals the presence of Mn(2+) and Fe(3+) ions giving rise to two resonance signals at g = 2.0 and 4.3, respectively. The resonance signal at g = 2.0 exhibits a six line hyperfine structure characteristic of Mn(2+) ions. EPR spectra have been studied at different temperatures from 123 to 433 K. The number of spins (N) participating in the resonance at g = 2.0 has been calculated at different temperatures. A linear relationship is observed between log N and 1/T in accordance with Boltzmann law and the activation energy was calculated. The paramagnetic susceptibility (chi) has been calculated at different temperatures and is found to be increasing with decreasing temperature as expected from Curie's law. From the graph of 1/chi versus T, the Curie constant and Curie temperature have been evaluated. The optical absorption spectrum exhibits bands characteristic of Fe(2+) and Fe(3+) ions. The crystal field parameter Dq and the Racah parameters B and C have been evaluated from the optical absorption spectrum. The infrared spectral studies reveal the formation of Fe(3+)--OH complexes due to the presence of higher amount of iron in this mineral. The Raman spectrum exhibits bands characteristic of Si--O--Si stretching and Mg?OH translation modes. PMID- 15294227 TI - Spectroscopic investigations of Nd(3+)-doped alkali chloroborophosphate glasses. AB - Optical absorption spectra were studied in wavelength region 400-900 nm for the Nd(3+)-doped alkali (R = Li, Na and K) chloroborophosphate glasses at room temperature. The energy level scheme of the 4f(3) electron configuration was deduced from the observed energy level data using a parametrized Hamiltonian (H(F1)) model which includes 20 free-ion interaction parameters. Reasonable correlation was obtained between the experimental and calculated energy levels. The Judd-Ofelt model for the intensity analysis of induced electric dipole transitions has been applied to the measured oscillator strengths of the absorption bands to determine the three phenomenological intensity parameters Omega(2), Omega(4) and Omega(6) for each glass. Using these parameters, the total radiative transition rates (A(T)), non-radiative relaxation rates (W(NR)), branching ratios (beta(R)), integrated cross-sections for the stimulated emission (Sigma), excited state emission intensities (f(ESE)) and excited state absorption intensities (f(ESA)) have been theoretically calculated for certain excited Nd(3+) fluorescent levels. From the results obtained, the conclusion is made about the possibility of using these glasses as laser media. PMID- 15294228 TI - Studies on 1H NMR, infrared and fluorescence spectra of rare earth complexes with a new binaphthylamide. AB - Solid complexes of lanthanide picrates with N,N-dibenzyl-2-[2' [(dibenzylcarbamoyl)-methoxy]-[1,1']binaphthalenyl-2-yloxy]-acetamide (L), [Ln(pic)(3)L] (Ln = La, Nd, Eu, Gd, Tb, Dy, Y), have been prepared and characterized by elemental analysis, IR, 1H NMR, UV-Vis spectra and conductivity measurements. The fluorescence property of the europium complex in solid state and in CHCl(3), acetone, AcOEt and DMF was studied. PMID- 15294229 TI - Selective fluorescence determination of chromium(VI) with poly-4-vinylaninline nanoparticles. AB - The strong fluorescence poly-4-vinylaniline nanoparticles (PVN) has been prepared under ultrasonic radiation. Based on the fluorescence quenching of PVN by Cr(VI), a method for the selective determination of Cr(VI), without separation of Cr(III) in water, was developed. The reaction conditions between Cr(VI) and PVN were investigated in detail. The assay is characterized by short reaction time (<1 min even at 0 degrees C temperature), very few interference stable fluorescence signals (at least 2.5 h), simple instrument (common spectrofluorometer) and simplicity (a one-step assay). Under optimal experimental conditions, a limit of detection of 0.02 microg ml(-1) was achieved. The calibration curve was linear over the concentration range 0.1-13.0 microg ml(-1) with a correlation coefficient of 0.9984. The proposed method has been applied to the selective quantification of Cr(VI) in synthetic samples and waste-water samples with the satisfactory results. PMID- 15294230 TI - Determination of proteins at nanogram levels by synchronous fluorescence scan technique with a novel composite nanoparticle as a fluorescence probe. AB - A novel composite nanoparticle has been prepared by an in situ polymerization method and applied as a protein fluorescence probe. The nano-CdS has been prepared, then the polymerization of acrylic acid (AA) was carried out by initiator potassium persulfate (KPS) under ultrasonic irradiation. The surface of the composite nanoparticles was covered with abundant carboxylic groups (--COOH). The nanoparticles are water-soluble, stable, and biocompatible. The synchronous fluorescence intensity of the composite nanoparticles is significantly increased in the presence of trace protein at pH 6.90. Based on this, a new synchronous fluorescence scan (SFS) analysis was developed for the determination of proteins including BSA, HSA, and human gamma-IgG. When Delta lambda = 280 nm, maximum synchronous fluorescence is produced at 290 nm. Under the optimum conditions, the response is linearly proportional to the concentration of proteins. The linear range is 0.1-10 microg ml(-1) for HSA, 0.09-8.0 microg ml(-1) for BSA, and 0.08 15 microg ml(-1) for human gamma-IgG, respectively. The method has been applied to the determination of the total protein in human serum samples collected from the hospital and the results are satisfactory. PMID- 15294231 TI - Spectroscopy behavior of diiodofluorescein and tetrabromofluorescein. AB - The luminescence behavior of diiodofluorescein (DIF) and tetrabromofluorescein (TBF) have been investigated including the solid surface room temperature phosphorescence (SS-RTP) and the room temperature fluorescence (RTF). The luminescence intensities of the two compounds are strongest in alkaline solution. RTP lifetime of the two compounds are in the range of 130-140 ms. The RTP and RTF polarization was in the range of 0.01-0.05. The two analytical methods--SS-RTP and RTF, of the two compounds have been established. PMID- 15294232 TI - Preparation and characterization of Eu(3+) doped powder spinel phosphors (MgAl(1.8)Y(0.2-x)O(4)). AB - The photoluminescence (PL) studies of powder phosphors are under rigorous study in view of the applications they have in the field of technology. Different methods are available for the preparation of rare earth ions doped in different host environment of powder phosphors. In the present work, a novel route known as sol-gel technique is employed to prepare spinel phosphor MgAl(1.8)Y(0.2 x)O(4):Eu(x) (x = 2-6 mol%). Then the studies have been carried out to optimize the dopant concentration in the host lattice with the help of photoluminescence spectra. These phosphors have displayed bright red color under UV source. The emission intensities were determined and the relative fluorescence intensities have been estimated. The richness of the red color has been verified by determining their chromaticity coordinates (X, Y) from the CIE standard charts. With the help of XRD, electron spin resonance (ESR), and photo-acoustic (PA) spectra of the samples prepared are also used for the confirmity of the host and analyzing of the data. PMID- 15294233 TI - Reactive oxygen species scavenging ability of a new compound derived from weathered coal. AB - The scavenging activity of three fulvic acids (named XWCS-1, XWCS-4, and XWCS-8 according to time taken for ozonolysis) obtained by ozonolysis of humic acid extracted from Xinjiang (China) weathered coal and a fulvic acid (named XWCFA) extracted from the same coal towards reactive oxygen species such as superoxide radical (O(2)(.)(-)) and hydroxyl radical ((.)OH) was investigated with an electron spin resonance (ESR)-spin trapping method using 5,5-dimethyl-1-pyrroline N-oxide (DMPO) as a spin trap. O(2)(.)(-) was generated with a hypoxanthine xanthine oxidase system. (.)OH was generated by three different methods; (i) FeSO(4)-hydrogen peroxide (H(2)O(2)) system, (ii) Cu(en)(2)-H(2)O(2) system, and (iii) UVB photolysis of H(2)O(2). At physiological pH, XWCS-1 had the greatest O(2)(.)(-) scavenging activity, followed by XWCS-4, XWCS-8 and XWCFA. XWCFA had the greatest ?OH scavenging activity among the four fulvic acids, whereas XWCS-1 and XWCS-4 enhanced the production of (.)OH from a metal-catalyzed hydroxyl radical generating system, suggesting that these molecules act as prooxidants in the presence of metal ion. PMID- 15294234 TI - Vibrational frequencies and structural determination of digermylcarbodiimide. AB - The vibrational frequencies and corresponding normal mode assignments of digermylcarbodiimide are examined theoretically using the Gaussian98 set of quantum chemistry codes. All normal modes were successfully assigned to one of eight types of motion (N=C=N asymmetric stretch, N=C=N symmetric stretch, Ge-H stretch, Ge-N stretch, H-Ge-H bend, GeH(3) wag, GeH(3) twist, and Ge-N. . .N-Ge torsion) utilizing the C(2) symmetry of the molecule. Uniform scaling factors were derived for each type of motion. Predicted infrared and Raman intensities are reported. PMID- 15294235 TI - A study of estimation of excited state dipole moments of di(4 bromophenyl)carbazone and its Cu(II), Zn(II), Cd(II) complexes from the solvent effect on their electronic spectra. AB - The solvent effect on the electronic spectra of di(4-bromophenyl)carbazone and its Cu(II), Zn(II), Cd(II) complexes have been studied by synthesizing and characterizing them by magnetic moment, IR, EPR and 1H NMR spectral measurements. The electric dipole moments of these compounds in the first electronically excited state have been determined. The results indicate that the observed band systems in these compounds may be attributed to pi(*) <-- pi transition. PMID- 15294236 TI - Cetyltrimethylammonium bromide sensitized resonance light-scattering of nucleic acid--Pyronine B and its analytical application. AB - This is the first report on the determination of nucleic acids with Pyronine B (PB) sensitized by cetyltrimethylammonium bromide (CTMAB) with resonance light scattering (RLS) technique. Under the experimental conditions (1 x 10(-5) mol l( 1) PB, 1 x 10(-5) mol l(-1) CTMAB, pH 7.4, at room temperature, ionic strength 0.02 mol l(-1) NaCl), the interaction of PB with DNA sensitized by CTMAB results in enhanced RLS signals at 328 and 377 nm in the enhanced regions. It was found that the enhanced RLS intensity at 328 nm was proportional to the concentration of DNA in the suitable ranges. The linear range of this assay is 0.0-1.2 microg ml(-1) for calf thymus, 0.0-0.8 microg ml(-1) for fish sperm DNA (fsDNA), and 0.04-1.4 microg ml(-1) for yeast RNA, respectively. The detection limits (3 sigma) are 6.1 ng ml(-1) for calf thymus DNA (ctDNA), 11.2 ng ml(-1) for fish sperm DNA, and 8.6 ng ml(-1) for yeast RNA, respectively. Six synthetic samples were determined satisfactorily. This method is simple, rapid and the dye is inexpensive and stable. PMID- 15294237 TI - Vibrational spectra and ab initio analysis of tert-butyl, trimethylsilyl, and trimethylgermyl derivatives of 3,3-dimethylcyclopropene. VI: Application of observed trends to stannyl derivatives. AB - The effects of substitution of X = C by Si or Ge in X(CH(3))(3) moieties attached to the formal double bond of 3,3-dimethylcyclopropene are examined. Regularities in observed trends of vibrational frequencies implicating the moieties containing the X atom, as the X atomic mass is increased, are extrapolated to X = Sn. The results of this extrapolation made it possible to assign the known experimental vibrational frequencies of 3,3-dimethyl-1-(trimethylstannyl)cyclopropene and 3,3 dimethyl-1,2-bis(trimethylstannyl)cyclopropene. PMID- 15294238 TI - Detection of Bacillus endospores using total luminescence spectroscopy. AB - Detection and analysis of bacteria from environmental samples (e.g. water, air, and food) are usually accomplished by standard culture techniques or by analyses that target specific DNA sequences, antigens or chemicals. For large cell numbers in aqueous suspensions, an alternative technique that has proven useful is total luminescence spectroscopy (TLS). TLS is the acquisition of fluorescence data that records the unique excitation-emission matrix (EEM) of compound fluorophores. Past work has shown that one type of bacterial endospore, Bacillus megaterium, possessed a distinct EEM pattern useful for differentiating it in complex biological fluids and suspensions. The work described here extends those observations to establish some limits on the sensitivity and specificity of TLS for the detection and analysis of bacterial endospores versus (bacterial) vegetative cells in aqueous culture. Our findings show Bacillus endospores exhibit a dramatic blue shift of 130 nm in excitation and a smaller shift of 50 nm in emission when compared to ancillary endospore and non-endospore forming bacterial cells. PMID- 15294239 TI - Conformational changes monitored by fluorescence study on reconstituted hemoglobins. AB - Intrinsic steady state fluorescence measurements were performed on a series of reconstituted metal ion and hybrid hemoglobins (Hbs). At 296 nm excitation, the spectrum exhibits a broad and asymmetric feature in the case of copper and nickel reconstituted hemoglobins. Deconvolution of the fluorescence bands clearly reveals the existence of two definite peaks. A similar trend was also observed for hybrid hemoglobins (CuNi, NiCu, CuFe-CO, and NiFe-CO). A guassian fit of the fluorescence bands in these proteins again yields two prominent peaks, which are assigned as due to two different tryptophan (Trp) environments. A relative ratio of the amplitudes of these peaks indicates the percentage of T-character in these molecules. This is in support to our previous findings by other spectroscopic studies on the same molecules. These studies therefore, suggest the presence of two different environments of a tryptophan thereby revealing structural heterogeneity among the subunits. PMID- 15294241 TI - Vibrational frequencies and structural determination of phosphorous tricyanide. AB - The normal mode frequencies and corresponding vibrational assignments of phosphorous tricyanide (P(CN)(3)) are examined theoretically using the Gaussian98 set of quantum chemistry codes. Each of the vibrational modes was assigned to one of four types of motion predicted by a group theoretical analysis P-C stretch, CN stretch, P-C[triple bond]C bend, and C-P-C bend) utilizing the C(3v) symmetry of the molecule. A uniform scaling factor was derived for each type of motion. Predicted infrared and Raman intensities are reported. PMID- 15294240 TI - Theoretical studies of the local structure for the trigonal Ti(3+) center in LiF crystal. AB - The local structure of the trigonal Ti(3+) center in LiF crystal is theoretically investigated by using the perturbation formulas of the anisotropic g factors and g(//) and g(/_) for a 3d(1) ion in trigonally distorted octahedra based on the cluster approach. From the studies on the basis of various possible structure models, the local structure of the trigonal Ti(3+) center may be characterized as [TiF(3)O(3)](6-) cluster (or model I). In this model, the impurity Ti(3+) is expected to substitute for the host Li(+) ion and shift away from its regular lattice site along the [111] (or C(3)) axis by about 0.19 A due to the strong electrostatic attraction of the O(2-) triangle replacing the original F(-) triangle. The magnitude of the above displacement obtained in this work is comparable with that ( approximately 0.2-0.3A) given by ENDOR experiment. Moreover, the cubic field parameter Dq (approximately 1497 cm(-1)) based on the above structure model is also in agreement with that (approximately 1500 cm(-1)) obtained from the experimental optical spectra of the studied system. The theoretical investigations of the local structure in this work may be useful to understand optical properties of Ti-doped LiF. PMID- 15294242 TI - Vibrational frequencies and structural determination of tetrafluoroformaldazine. AB - The normal mode frequencies and corresponding vibrational assignments of tetrafluoroformaldazine (F(2)CNNCF(2)) are examined theoretically using the Gaussian98 set of quantum chemistry codes. Each of the vibrational modes was assigned to one of nine types of motion predicted by a group theoretical analysis (C-F stretch, C[triple bond]N stretch, N-N stretch, C=C-N bend, CF(2) wag, CF(2) rock CF(2) scissors, CF(2) twist, and C=N-N=C torsion) utilizing the C(2h) symmetry of the molecule. Uniform scaling factors was derived for each type of motion. Predicted infrared and Raman intensities are reported. PMID- 15294243 TI - Vibrational frequencies and structural determination of Al(8)S(12). AB - The normal mode frequencies and corresponding vibrational assignments of Al(8)S(12) in T(h) symmetry are examined theoretically using the Gaussian98 set of quantum chemistry codes. All normal modes were successfully assigned to one of four types of motion (Al-S stretch, Al-S-Al bend, S-Al-S bend, and Al-S-Al wag) predicted by a group theoretical analysis. Normal mode frequencies are predicted and calculated infrared intensities and Raman activities are presented. The thermodynamics of the reaction 2Al(4)S(6)-->Al(8)S(12) are examined. PMID- 15294244 TI - Theoretical and experimental vibrational study of phenylurea: structure, solvent effect and inclusion process with the beta-cyclodextrin in the solid state. AB - FTIR and Raman vibrational spectroscopic techniques as well as DFT quantum chemical calculation were used for characterizing conformational changes of phenylurea (a herbicide parent molecule) occurring from solid state to aqueous medium. Experimental infrared frequencies were assigned on the base of urea and benzenic derivatives spectroscopic data available in the literature and vibrational normal modes analytical calculation at the fully optimized geometry. Investigation of isotopic and solvent effects has revealed that the structure of phenylurea is particularly sensitive to the electrostatic environment via hydrogen non covalent bonds. The ability of beta-cyclodextrin (beta-CD) to form host-guest inclusion complex with phenylurea in the solid state was also evidenced by significant frequency shifts observed in the 1400-1800 cm(-1) spectral range. PMID- 15294245 TI - Vibrational frequencies and structural determination of triethynylmethylstannane. AB - The normal mode frequencies and corresponding vibrational assignments of Triethynylmethylstannane (SnCH(3)(CCH)(3)) are examined theoretically using the Gaussian 98 set of quantum chemistry codes. Each of the vibrational modes was assigned to one of nine types of motion predicted by a group theoretical analysis (Sn-C stretch, C[triple bond]C stretch, C-H stretch, C[triple bond]C-H bend, Sn C[triple bond]C bend, C-Sn-C bend, H-C-H bend, CH(3) wag, and CH(3) twist) utilizing the C(3v) symmetry of the molecule. A set of uniform scaling factors was derived for each type of motion. Predicted infrared and Raman intensities are reported. PMID- 15294246 TI - The infrared spectra and theoretical calculations of frequencies of fac tricarbonyl octahedral complexes of manganese(I). AB - The carbonyl stretching frequencies in the infrared spectra of 38 fac-tricarbonyl octahedral complexes of manganese(I) prepared in this laboratory were determined. These complexes may be grouped into three types: (a) neutral complexes of the structure (CO)(3)Mn(P-P)Z where P-P represents depe, dppe, or dppp, and Z represents various anionic functional groups bonded to the manganese; (b) ionic complexes of the structure [(CO)(3)Mn(P-P)Z](+)BF(4)(-) where Z represents various neutral molecules possessing one phosphorous, nitrogen, or oxygen atom coordinated to the manganese; (c) complexes of the structure (CO)(3)Mn(pn)Z where the chelating pn represents 1,1-diphenylphosphino-2,2-dimethylaminoethane, Ph(2)PCH(2)CH(2)NMe(2). All of these complexes show three carbonyl stretching modes (2A' + A"). The effects on the frequencies of these modes induced by both the various Z groups and the various ligands are discussed. Theoretical calculations (B3LYP/6-31G) with optimization of the full molecule make it possible to distinguish between the three stretching modes and to make unambiguous assignments of appropriate symmetry species to each. PMID- 15294247 TI - C-S barrier and vibrational analyses of (halocarbonyl)sulfenyl halides XCO-SX (X = F, Cl, and Br). AB - The structural stability of (halocarbonyl)sulfenyl halides XCO-SX (X is F, Cl, and Br) was investigated by DFT-B3LYP and ab initio MP2 calculations using 6-311 + G(**) basis set. From the calculations the molecules were found to exist predominantly in the trans conformation (two halogen atoms are trans to each other). Full energy optimizations were carried out for the minima and the transition states (TS) at the two levels, from which the rotational barriers about C-S bond in the three molecules were calculated to be about 12-13 kcal mol( 1). The vibrational frequencies of (fluorocarbonyl)sulfenyl fluoride (FCO-SF), (chlorocarbonyl)-sulfenyl chloride (ClCO-SCl), and (bromocarbonyl)-sulfenyl bromide (BrCO-SBr) were computed at the DFT-B3LYP level and the vibrational assignments for the normal modes of the stable forms of the compounds were made on the basis of normal coordinate calculations and experimental data of the chloride. PMID- 15294248 TI - Vibrational frequencies and structural determination of digermyl ether. AB - The normal mode frequencies and corresponding vibrational assignments of digermyl ether in C(2v) symmetry are examined theoretically using the Gaussian 98 set of quantum chemistry codes. All normal modes were successfully assigned to one of six types of motion (Ge-H stretch, Ge-O stretch, Ge-O-Ge bend, H-Ge-H bend, GeH(3) wag, and GeH(3) twist) predicted by a group theoretical analysis. By comparing the vibrational frequencies with IR and Raman spectra available in the literature, a set of scaling factors is derived. Predicted infrared and Raman intensities are reported. PMID- 15294249 TI - Structural analysis and Fourier transform infrared and Raman spectra of dibutoxyphosphoryl benzylisothiourea. AB - A structural analysis for dibutoxyphosphoryl benzylisothiourea (DBBT) was carried out by mass spectrometry, 1H NMR, 13C NMR, infrared and Raman spectroscopy. The Fourier transform infrared and Fourier transform Raman spectra of liquid of DBBT were carried out with the purpose of studying the tautomerism (structures I and II) and the behavior of the more polar absorption's bands in different solvents, i.e., absorption's of the P=O and C=N bands. The results suggest the existence of tautomerism in the pure (liquid) compound and in solution of CHCl(3), CH(2)Cl(2), CHBr(3), and THF, C(2)H(4)Cl(2) and C(2)H(4)Br(2). The solvent interaction with the P=O band was characterized by the presence of a new band in the region of the O-H absorption. A vibrational assignment of the IR bands and Raman shifts was done and is proposed in this paper. PMID- 15294250 TI - Spectrochemical, ab initio and density functional studies on the conversion of 2 hydroxybenzonitrile (o-cyanophenol) into the oxyanion. AB - The spectral and structural changes caused by the conversion of 2 hydroxybenzonitrile (o-cyanophenol) into the corresponding oxyanion have been followed by IR spectra, ab initio and density functional force field calculations. In agreement between theory and experiment, the conversion is accompanied by a 29 cm(-1) frequency decrease of the cyano stretching band, 2.7 fold increase in its integrated intensity, 5.8-fold (total value) intensification of the aromatic skeletal bands of Wilson's 8 and 19 types, and other essential spectral changes. According to the calculations, the strongest structural changes are the shortening of the Ph-O bond with 0.10 A, lengthenings of the adjacent CC bonds in the phenylene ring with 0.06 A and bond angle variations near the oxyanionic center. All these changes are connected with the formation of a quasi ortho-quinonoidal structure of the o-phenylene ring in the oxyanion. According to the electronic density analysis, 0.41 e(-) (Mulliken) or 0.56 e(-) (natural bond orbital, NBO) of the anionic charge remain localized at the oxyanionic center. Conformations and hydrogen bonds have also been discussed on the basis of experimental and theoretical data. PMID- 15294251 TI - Infrared spectra of H(2)16O, H(2)18O and D(2)O in the liquid phase by single-pass attenuated total internal reflection spectroscopy. AB - Mid-infrared attenuated total internal reflection (ATR) spectra of H(2)16O, H(2)18O and D(2)16O in the liquid state were obtained and normal coordinate analysis was performed based on the potential energy surface obtained from density functional theory (DFT) calculations. Fits of the spectra to multiple Gaussians showed a consistent fit of three bands for the bending region and five bands for the stretching region for three isotopomers, H(2)16O, H(2)18O and D(2)16O. The results are consistent with previous work and build on earlier studies by the inclusion of three isotopomers and mixtures using the advantage of single-pass ATR to obtain high quality spectra of the water stretching bands. DFT calculation of the vibrational spectrum of liquid water was conducted on seven model systems, two systems with periodic boundary conditions (PBC) consisting of four and nine H(2)16O molecules, and five water clusters consisting of 4, 9, 19, 27 and 32 H(2)16O molecules. The PBC and cluster models were used to obtain a representation of bulk water for comparison with experiment. The nine-water PBC model was found to give a good fit to the experimental line shapes. A difference is observed in the broadening of the water bending and stretching vibrations indicative of a difference in the rate of pure dephasing. The nine-water PBC calculation was also used to calculate the wavenumber shifts observed in the water isotopomers. PMID- 15294252 TI - Crystal structure of 1,10-dibromodecane and its infrared intensity in a urea clathrate and in the crystal. AB - The crystal structure of 1,10-dibromodecane belongs to the monoclinic system and the space group is P2(1)/c with lattice dimensions of a = 5.4574(3) A, b = 5.2814(4) A, c = 21.088(1) A and beta = 92.897(2) degrees and zeta = 2. Infrared spectra of 1,10-dibromodecane in a urea clathrate and in the crystal were observed to investigate the effect of molecular interaction on infrared intensity. The infrared intensity of the CH(2) waggings in the crystalline state is 1.5-1.9 times stronger on the relative basis than that in a urea clathrate, whereas those of CH(2) stretching, CH(2) rocking and CH(2) bending are almost the same in both states. The former enhancement is explained in terms of increase in the bond moment of the C(alpha)H(2) group on the basis of crystal structure and the electrostatic model. The relative intensity of two CH(2) asymmetric stretching changes between the two states. This is also analyzed by the use of the electrostatic model. PMID- 15294253 TI - Vibrational infrared intensities and polar tensors of 1,2-difluoroethylenes. AB - The polar tensors of cis and trans-1,2-difluoroethylenes have been determined with new normal modes based in a reassignment of nu(7) and nu(12) bands of the trans isomers and frequency values corrected for Fermi resonances and phase shifts. The signs of the dipole moment derivatives (and its directions, for B(u) symmetry species) were considered to be those of MP2/6-31G** estimates. Root mean square errors calculated for the new tensor element values from each pair of isotopomers (trans-1,2-C(2)H(2)F(2)/trans-1,2-C(2)D(2)F(2) and cis-1,2 C(2)H(2)F(2)/cis-1,2-C(2)D(2)F(2)) show that the new polar tensor sets fit the isotopic invariance criterion better than previously reported sets. The accuracy of polar tensor transference procedures was tested by calculating the infrared intensities of trans-1,2-C(2)H(2)F(2) through the new polar tensors of the cis isomer. The resulting estimates are very accurate and also support the new band assignment, though the A(6) intensity remains still somewhat underestimated. PMID- 15294254 TI - [Ruthenium(II)(bpy)(2)L](2+), where L are imidazo[f]-1,10-phenanthrolines: synthesis, photophysics and binding with DNA. AB - Three Ru(II) complexes of type as [Ru(II)(bpy)(2)L](2+) were synthesized, where L are l,10-phenanthroline derivatives of imidazole (1), having at position 2 alpha naphthyl (2), 3-methoxy-4-hydroxy-phenyl (3). All complexes show intense MLCT transition both in acetonitrile and in water and also exhibit strong emission at room temperature, which is efficiently quenched by oxygen as well as, to some extent, by water. The binding of complexes 1-3 to calf thymus DNA was investigated by using electronic absorption, steady-state luminescence, luminescence quenching, excited-state lifetime and circular dichroism spectra. Hypochromic effect, luminescence enhancement, and quenching studies demonstrate the existence of intercalation mode. Circular dichroism spectra indicate the stereoselectivity of the binding. The binding of 1-3 with DNA is sensitive to the nature of ligands, such as planarity, pi-electron extension and hydrophobicity. Complex 3 exhibits the strongest binding with DNA, which can be attributed to hydrogen bonding. PMID- 15294255 TI - Vibrational spectral studies of L-methionine L-methioninium perchlorate monohydrate. AB - The infrared and laser Raman spectra of L-methionine L-methioninium perchlorate monohydrate were recorded at room temperature and the vibrational assignments of the observed wave numbers were made. The presence of both the carbonyl and the ionized carboxylic groups has been identified in the title complex. The L methionine and L-methioninium the cation have different conformations. This together with the different environment has seen by the two -CH(2)- groups in each skeleton cause several of the group wave numbers to occur as doublets or as broad bands. The perchlorate anion was found to be in the T(d) symmetry in the methionine environment. The coordination to other ligands in the crystal through hydrogen bonding does not affect this symmetry. The extensive intermolecular hydrogen bonding in the crystal was identified by the shifting of bands due to the stretching and bending modes of the various functional groups. Fermi resonance has also been observed. PMID- 15294256 TI - Single crystal EPR study of Cu(2+) in cobalt ammonium phosphate hexahydrate: a case of low hyperfine coupling constant and measurement of spin-lattice relaxation times. AB - Single crystal electron paramagnetic resonance (EPR) studies of Cu(II) doped cobalt ammonium phosphate hexahydrate have been carried out from 300 to 77 K, with single crystal rotation in all the three planes at 153 K, since the spectra are well resolved at this temperature. The angular variation studies indicate only one site in substitutional position with spin-Hamiltonian parameters as g: 2.404, 2.155, 2.063 and A: 11.58, 3.49, 2.07 mT. The reduction in one of the principal A value has been explained by considering considerable admixture of d(x(2)-y(2)) ground state with d(zeta(2)) excited state. The admixture coefficients of ground state wave function are: a = 0.2500; b = 0.9663; c = 0.0520; d = 0.0210; e = -0.0210, where a and b correspond to admixture coefficients for d(zeta(2)) and d(x(2)-y(2)) , respectively. Parameters kappa = 0.5140; P = 113 X 10(-4) cm(-1); alpha(2) = 0.7897; alpha = 0.8887; and alpha' = 0.5262 have also been calculated, indicating considerable covalency. The powder spectrum at room temperature is unresolved, whereas it is better resolved at 77 K, with spin-Hamiltonian parameters matching well with the single crystal values of 153 K. Powder spectrum at 77 K has been simulated, which agrees with the experimental one. The spin-lattice relation times are measured from the line width of the resonance lines recorded at different temperature. PMID- 15294258 TI - The intramolecular vibrational energy redistribution threshold in S(1) deuterated p-difluorobenzene. AB - The intramolecular vibrational energy redistribution (IVR) in S(1) deuterated p difluorobenzene (pDFB-d(4) or -d(4)) has been studied to determine the IVR threshold. For this, the S(1) <-- S(0) fluorescence excitation (FE) spectrum of jet-cooled d(4) was investigated in the 2000-3250 cm(-1) vibronic energy range of the S(1) electronic state, and single vibronic level fluorescence (SVLF) spectra have been acquired by exciting selected levels lying between 750 and 2850 cm(-1) in vibrational energy in the S(1) excited state. Congestion of the dispersed fluorescence in this molecule first appears as the vibrational level energy climbs above 2000 cm(-1). By comparing the SVLF spectra of pDFB-d(4) with those of p-difluorobenzene (pDFB or -h(4)), it is obvious that IVR threshold in -d(4) is localized with a few hundreds cm(-1) lower than that in pDFB. This decrease is entirely due to the increase in vibrational state density due to deuteration. PMID- 15294257 TI - Single crystal EPR studies of paramagnetic ions doped zinc potassium phosphate hexahydrate; Part III: Mn(II)-a case of rhombic distortion. AB - Single crystal electron paramagnetic resonance (EPR) studies of Mn(II) doped zinc potassium phosphate hexahydrate have been carried out at room temperature. Single crystal rotations along the three orthogonal axes indicate orthorhombic symmetry with spin-Hamiltonian parameters as: g(xx) = 1.9997; g(yy) = 1.9538; g(zz) = 1.9524, D(xx) = 15.49 mT; D(yy) = 0.22 mT; D(zz) = -15.71 mT, A(xx) = 11.70 mT; A(yy) = 10.53 mT; A(zz) = 10.42 mT and a = 0.8 x 10(-4) cm(-1). A large E term indicates considerable distortion from axial symmetry. The impurity is found to enter the lattice substitutionally. The distortion axis for the impurity has been identified along one of the Zn-O bond directions in the crystal. PMID- 15294259 TI - Principal and independent components analysis of overlapping spectra in the context of multichannel time-resolved absorption spectroscopy. AB - The separation of overlapping absorption spectra in the context of multichannel time-resolved absorption spectroscopy and chemical kinetics is a particular case in the general problem of splitting the observed data into several linear components. Here, principal and independent components analysis are applied to kinetic data of iodine--ozone chemistry, which contains overlapping spectra of different absorbers. The objective of this work is to demonstrate a method which in spite of this overlap is able to extract separated time traces of such absorbers. These time traces are clearly a pre-requisite for any further accurate quantitative analysis. The statistical properties of data recordings obtained from flash photolysis of I(2) and O(3) have been studied to check if the requirements of the model are fulfilled. Results of separation in appropriate spectral windows displaying overlapped vibrational features are presented. Validation is made using prior information and conventional techniques. PMID- 15294260 TI - Atrial pacing for prevention of atrial fibrillation: assessment of simultaneously implemented algorithms. AB - AIMS: Several preliminary studies indicated that right atrial pacing could prevent atrial tachyarrhythmias (ATA). We sought to compare the safety and the efficacy of atrial-based pacing supplemented by dedicated combined algorithms with conventional atrial pacing in the prevention of ATA. METHODS: Fifty-five patients with a history of recurrent paroxysmal ATA implanted with a dual-chamber pacemaker were studied during two randomized cross-over pacing periods (conventional DDD and DDD with ATA prevention algorithms) of 6 months duration. The primary endpoint was the burden of ATA episodes recorded by the device mode switch algorithm. RESULTS: The cross-over analysis did not demonstrate any significant difference between the two pacing modes: 254+/-533 h of ATA during the control period versus 238+/-518 h during the ATA prevention period. Analysis of a subgroup of patients found that those with the lower percentage of ventricular pacing benefited from ATA prevention algorithms (120+/-182 h versus 225+/-350 h during the control period; P < 0.04). CONCLUSION: When compared with DDD pacing at 70 bpm, ATA prevention algorithms have not demonstrated significant efficacy. However, a subgroup of patients with preserved native AV conduction (low percentage of ventricular pacing) responded to ATA prevention algorithms. PMID- 15294261 TI - Is there a role for pacing in the prevention of atrial tachyarrhythmias? PMID- 15294262 TI - Evaluation of KCB-328, a new IKr blocking antiarrhythmic agent in pacing induced canine atrial fibrillation. AB - HYPOTHESIS: KCB-328 is a new potassium channel blocker, which prolongs action potential duration with exhibition of minimal reverse use dependence. We tested the efficacy and proarrhythmic potential of KCB-328, dofetilide and propafenone in the pacing induced canine model of atrial fibrillation (AF). METHODS: Mongrel dogs in complete heart block were paced for 1-6 weeks to produce AF, and given KCB-328 or dofetilide. A subset then received propafenone 14+/-3 days after testing the first drug. RESULTS: KCB-328 prolonged right and left atrial (RA and LA) activation times and AF cycle length (CL), terminating AF in 3 of 6 dogs. RA effective refractory period (ERP) and ventricular ERP and QT interval were prolonged. Dofetilide terminated AF in 1/6 dogs, and increased AF CL and ventricular ERP and QT interval. Dofetilide's reverse use dependency on the QT interval was greater than KCB-328. Propafenone prolonged RA and LA activation times and AF CL and terminated AF in 8 of 9 dogs. One death occurred with dofetilide, none with KCB-328 or propafenone. CONCLUSION: The spectrum of effect of the three drugs differed significantly: propafenone showed the greatest success in AF termination, and both propafenone and KCB-328 appeared less proarrhythmic than dofetilide in this model. PMID- 15294263 TI - Immediate Risk-Stratification Improves Survival (IRIS): study protocol. AB - BACKGROUND: To date, the implantable cardioverter-defibrillator (ICD) has been shown to be effective for primary prevention of sudden cardiac death only in selected groups of patients in the chronic phase after myocardial infarction. METHODS AND RESULTS: The Immediate Risk-Stratification Improves Survival (IRIS) Study compares ICD therapy with no ICD therapy in selected high risk patients early after myocardial infarction. Special emphasis is placed on optimal acute and long term medical therapy in all patients including metoprolol CR/ZOK. The hypothesis is tested that use of the ICD reduces overall mortality. For that purpose, consecutive acute ST elevation or non-ST elevation myocardial infarction patients are collected in a registry. From this denominator, patients are screened, and enrolled early after myocardial infarction (day 5 to day 31) if they exhibit both a reduced left ventricular ejection fraction < or =40% and a heart rate > or =100 bpm on the first available electrocardiogram (criterion I), or non-sustained ventricular tachycardia at a rate > or =150 bpm during Holter (criterion II). CONCLUSIONS: IRIS is a large scale prospective, randomized trial to evaluate the benefit of ICD therapy for reduction of total mortality in patients considered at high risk of sudden death early after acute myocardial infarction. PMID- 15294264 TI - The role of implantable cardioverter defibrillator for primary vs secondary prevention of sudden death in patients with idiopathic dilated cardiomyopathy. AB - AIM: To analyse the characteristics and outcome of patients with idiopathic dilated cardiomyopathy (DC) considered at high risk of sudden death (SD) and treated with implantable cardioverter defibrillators (ICD) for primary prevention (Group A) in comparison with patients treated with ICDs because of previous sustained ventricular tachyarrhythmias or syncope (Group B). METHODS: Group A consisted of 27 patients with at least two of the following criteria: left ventricular end-diastolic diameter (LVEDD) > or =70 mm (74%), LV ejection fraction (LVEF) < or =30% (78%), non-sustained ventricular tachycardia (VT) (56%), long history of disease (> or =48 months since diagnosis, 85%) and family history of SD (11%). Group B consisted of 27 patients treated with ICDs because of sustained VT/fibrillation (n=18) or syncope (n=9). RESULTS: NYHA class, LVEF, LVEDD and amiodarone treatment were similar in the two groups. Patients in group A were younger (46+/-15 vs 59+/-17 years, P=0.0008), were more often treated with beta-blockers (89% vs 62%; P=0.02) and had a longer interval since diagnosis (86+/-60 vs 40+/-50 months; P=0.004). Twelve month rates of appropriate intervention (AI) were 41% in Group A and 57% in group B (P NS). In group A, after a mean follow-up of 21+/-14 months, patients showing the combination of LVEF < or =30% and LVEDD > or =70 mm had the highest frequency of AI (76% vs 10%, P=0.005). In group B, after a mean follow-up of 33+/-23 months, 78% of patients with syncope had AI. Total and sudden deaths were 11% and 4% in group A and 19% and 4% in group B (P NS). CONCLUSIONS: Patients with idiopathic DC treated with ICD for primary prevention because they were considered at high risk of SD according to clinical criteria showed a high rate of AI, similar to that of patients treated for secondary prevention. The highest rate of AI was seen in patients with both severe dysfunction and dilatation and in those with previous syncope. PMID- 15294265 TI - Typical atrial flutter ablation outcome: correlation with isthmus anatomy using intracardiac echo 3D reconstruction. AB - AIMS: To verify if sites of conduction gaps on the isthmus correlate with anatomical peculiarities using the intracardiac echo (ICE) and a new 3D device to reconstruct the isthmus in patients undergoing cavotricuspid isthmus ablation. METHODS AND RESULTS: Twenty patients underwent isthmus ablation using an 8 mm tip ablation catheter. Two-dimensional and 3D ICE reconstruction of the isthmus was made before, during and after ablation. At the end of the lesion line isthmus block was validated by electrophysiological criteria. In case of its absence we closed the remaining conduction gaps verifying the position of the sites with ICE. Fourteen patients required a median of 8 RF pulses to obtain complete isthmus block (Group A). In the remaining 6 patients isthmus block was obtained with a median of 25 RF pulses due to conduction gaps 'resistant' to ablation (Group B). Conduction gap positions assessed by ICE were located in the central portion of the isthmus below the coronary sinus os in 71% of cases in Group A and along a prominent Eustachian ridge in Group B patients, respectively. 3D reconstruction showed a smooth isthmus in Group A with a 'peak and valleys' isthmus in Group B. In these latter patients isthmus block was obtained only after the complete ablation of the prominent Eustachian ridge. CONCLUSION: The isthmus presents anatomical variants particularly due to Eustachian ridge peculiarities which may represent a site of conduction gaps "resistant" to ablation. PMID- 15294266 TI - Electrical stunning and hibernation: suggestion of new terms for short- and long term cardiac memory. AB - Persistent T wave changes following resumption of sinus rhythm induced by pacing or arrhythmias that cause altering of ventricular activation sequence are named "cardiac memory". After this initial definition there has been a discussion whether such T wave changes are primary, secondary or pseudoprimary. In addition according to the results of pathophysiological studies investigating the mechanism and nature of this repolarization abnormality some authors have preferred to use the term "electrical remodelling" instead of cardiac memory. But these two terms are still not well defined. In this article, the previous terms are discussed and a new term instead of cardiac memory is introduced. PMID- 15294267 TI - Recurrence of left atrium-pulmonary vein conduction following successful disconnection in asymptomatic patients. AB - BACKGROUND: The clinical significance of conduction recurrences in isolated pulmonary veins of patients with atrial fibrillation is not established. METHODS: Twenty-two patients with paoxysmal atrial fibrillation underwent successful pulmonary vein isolation. Six months after the procedure, 14 patients were free of atrial fibrillation. Two of these patients were subjected to repeat mapping of the left superior pulmonary vein. RESULTS: There was recurrence of pulmonary vein to left atrium conduction despite complete lack of symptoms or evidence of recurrent atrial fibrillation. CONCLUSION: Successful pulmonary vein isolation with abolition of paroxysmal atrial fibrillation does not confer permanent disconnection of the pulmonary vein musculature from the left atrium. PMID- 15294268 TI - Depletion of atrial natriuretic peptide during longstanding atrial fibrillation. AB - This review focuses on the relation between atrial fibrillation (AF) and atrial natriuretic peptide (ANP). ANP is produced by the atria secondary to atrial stretch. By causing atrial stretch, acute AF leads to an increase in plasma ANP concentration, which serves to normalize haemodynamics through natriuresis and vasodilation. However, data have been reported suggesting that prolonged AF, by inflicting structural atrial damage, is associated with a reduced capacity by the atria to produce ANP. An inverse relation was thus demonstrated between the duration of AF and plasma ANP concentration. In addition, a reduced ANP response to exercise has been shown to be predictive of unsuccessful cardioversion of AF to sinus rhythm. Finally, ANP has also been shown to predict outcome after a maze operation. Outcome was poor when preoperative plasma ANP concentration was low. Moreover, a high atrial collagen content, as a measure of atrial degeneration, correlated with low ANP. These data indicate that ANP may serve as a marker of atrial integrity, which may help in selecting AF patients for therapeutic interventions. PMID- 15294269 TI - Assessment of upgrading to biventricular pacing in patients with right ventricular pacing and congestive heart failure after atrioventricular junctional ablation for chronic atrial fibrillation. AB - AIMS: Effects of cardiac resynchronization therapy (CRT) in patients with right ventricular pacing and congestive heart failure (CHF) have only been reported in limited series. CRT in patients with atrial fibrillation remains controversial. Patients with AV junctional ablation offer a unique opportunity to study the effects of CRT in patients with right ventricular pacing combined with atrial fibrillation. The aims of the present study were to evaluate the effects of upgrading to biventricular pacing patients with CHF, permanent atrial fibrillation, and prior ablation of the atrioventricular (AV) junction followed by conventional right ventricular pacing. METHODS AND RESULTS: We studied 16 consecutive patients with permanent atrial fibrillation treated by AV junctional ablation. After a mean follow-up of 20+/-19 months (6 weeks to 5 years) they were successfully upgraded to biventricular pacing for severe CHF. Parameters were prospectively evaluated at baseline and at 6 months. The 14 surviving patients at 6 months demonstrated significant improvement (P<0.02) in New York Heart Association class but the exercise test parameters remained unchanged. Cardiothoracic ratio decreased by 5% (P=0.04), end-systolic diameter by 8% (P=0.001), end-diastolic diameter by 4% (P=0.08), systolic pulmonary artery pressure by 17% (P<0.0001) and mitral regurgitation area by 40% (P<0.05). Ejection fraction increased by 17% (P=0.11) and fractional shortening by 24% (P=0.01). CONCLUSION: CRT improves left ventricular performance and functional status in patients with permanent atrial fibrillation and prior remote right ventricular pacing. PMID- 15294270 TI - Long-term clinical performance of AAI pacing in patients with sick sinus syndrome: a comparison with dual-chamber pacing. AB - AIMS: In this clinical study, we compared two groups of age-matched patients, AAI and DDD, to evaluate the clinical benefits of AAI pacing in patients with sick sinus syndrome (SSS) and normal atrioventricular (AV) conduction. METHODS AND RESULTS: Ninety-five patients with SSS implanted with AAI pacemakers were compared with 101 SSS patients implanted with DDD pacemakers. Mortality, chronic atrial fibrillation, lead survival rates, and reoperation rates were compared by Kaplan-Meier analysis. Eight AAI devices were switched to DDD due to high-degree (grade 2-3) AV block. The incidence of high-degree AV block was 1.104%/year, with a freedom rate of 88.6% at 10 years. There were no significant differences between the two groups in survival rates (87.8% in AAI vs. 93.4% in DDD at 10 years), freedom from atrial fibrillation (93.6% vs. 90.6%), or freedom from reoperation (71.3% vs. 76.3%). On the other hand, lead failure was twice as frequent in the DDD group than in the AAI group (relative risk=2.045, P=0.0382). CONCLUSION: AAI pacing, a simple system using a single lead and single-chamber pacemaker, can achieve a clinical outcome similar to that of the DDD mode in patients with SSS and normal AV conduction. PMID- 15294271 TI - Anomaly of the middle cardiac vein? PMID- 15294272 TI - Multiple-vein thrombosis and pulmonary embolism after pacemaker implantation treated by thrombolysis. AB - This report describes a patient who suffered multiple-vein thrombosis following permanent pacemaker implantation and developed a pulmonary embolism while on anticoagulation treatment, which was successfully treated by thrombolytic therapy. PMID- 15294273 TI - Impact of age and blood pressure on the lower arterial pressure limit for maintenance of consciousness during passive upright posture in healthy vasovagal fainters: preliminary observations. AB - Maintenance of consciousness importantly depends on systemic arterial blood pressure (BP) remaining above the lower pressure limit for cerebrovascular autoregulation. This study evaluated the impact of age and baseline arterial blood pressure (BP) on the BP recorded at onset of syncope in otherwise healthy individuals undergoing passive head-up tilt (HUT) testing for suspected vasovagal syncope. Since hypertension is thought to shift the lower autoregulation point to higher values, and since older healthy patients tend to have higher BP than younger individuals, we hypothesized that even among healthy individuals HUT induced syncope would occur at higher BP in older compared with younger subjects. Three groups of otherwise healthy individuals who had positive HUT were identified: Group 1: <25 years, n=17; Group 2: 25-59 years, n=18; and Group 3: > or =60 years, n=7. As expected, baseline arterial systolic blood pressure of patients > or =60 years (162+/-37 mmHg) was significantly higher than in the other two groups (Group 1: <25 years, 116+/-15 mmHg; Group 2: 25-59 years, 128+/ 12 mmHg). Further, the > or =60 age group tolerated upright posture for a longer period before syncope than did younger patients. However, despite a trend for BP at syncope to increase with age, differences were small (Group 3: > or =60 years, 61+/-15 mmHg, Group 2: 25-59 years, 58+/-6 mmHg, and Group 1: 54+/-16 mmHg) and were not statistically significant. Thus, in generally healthy individuals, age and baseline BP has only a minor effect on the lower limit of BP necessary for maintenance of consciousness. On the other hand, higher baseline BP provides older individuals a greater blood pressure 'reserve' for maintenance of consciousness compared with younger subjects. PMID- 15294274 TI - Engineering, cloning, and expression of genes encoding the multimeric luteinizing hormone-releasing hormone linked to T cell determinants in Escherichia coli. AB - Two synthetic genes were designed and engineered to encode for multimeric luteinizing-hormone-releasing hormone (LHRH) peptides linked to T cell determinants. These genes were cloned into the prokaryotic expression vectors under control of strong inducible promoters, to overexpress the multimeric LHRH peptides as recombinant proteins. Multimeric LHRH-T cell peptides were expressed as insoluble inclusion bodies in Escherichia coli cultures. Cell extracts containing the recombinant proteins showed immunoreactivity on Western blots with monoclonal antibody recognizing the native hormonal peptide. These gene constructs have potential applications in therapy of sex-steroid-hormone dependent cancers. PMID- 15294275 TI - Purification, refolding, and characterization of recombinant LHRH-T multimer. AB - To make the native LHRH immunogenic, a multimer of LHRH interspersed with T non-B peptides (r-LHRH-d2) was expressed as recombinant protein in Escherichia coli. The expression level of the recombinant protein was around 15% of the total cellular protein and it aggregated as inclusion bodies. Inclusion bodies from the bacterial cells were isolated and purified to homogeneity. Instead of high concentrations of chaotropic agents, r-LHRH- d2 was solubilized in 50 mM citrate buffer at pH 3 containing 2 M urea. The protein was refolded by 5-fold dilution (pulsatile) with cold 10 mM citrate buffer at pH 6 in presence of 0.3 M L arginine. Purification of r-LHRH-d2 was carried out by successive passages on CM Sepharose column at pH 6.0 which retained extraneous proteins and pH 4.8 at which r-LHRH-d2 bound to the resin. The elution was carried out by using linear salt gradient (0.1-1 M NaCl). The overall yield of the purified r-LHRH-d2 was 40% of the initial inclusion body proteins. The purity and homogeneity were confirmed by a single homogeneous peak on analytical HPLC eluting out at 29.51 min and by single band on SDS-PAGE reactive with polyvalent anti-LHRH antibodies. Mass spectroscopic analysis indicated the protein to be of 16.6 kDa which equals the theoretically expected mass. The N-terminal amino acid analysis of r-LHRH-d2 showed the sequence which corresponded to the designed protein. The CD spectrum of the refolded r-LHRH-d2 showed that the multimer has considerable beta sheet structure like the monomeric LHRH protein. PMID- 15294276 TI - An optimized fermentation process for high-level production of a single-chain Fv antibody fragment in Pichia pastoris. AB - The expression of a humanized single-chain variable domain fragment antibody (A33scFv) was optimized for Pichia pastoris with yields exceeding 4 g L(-1). A33scFv recognizes a cell surface glycoprotein (designated A33) expressed in colon cancer that serves as a target antigen for immunotherapy of colon cancer. P. pastoris with a MutS phenotype was selected to express A33scFv, which was cloned under regulation of the methanol-inducible AOX1 promoter. We report the optimization of A33scFv production by examining methanol concentrations using fermentation technology with an on-line methanol control in fed-batch fermentation of P. pastoris. In addition, we examined the effect of pH on A33scFv production and biomass accumulation during the methanol induction phase. A33scFv production was found to increase with higher methanol concentrations, reaching 4.3 g L(-1) after 72 h induction with 0.5% (v/v) methanol. Protein production was also greatly affected by pH, resulting in higher yields (e.g., 4.88 g L(-1)) at lower pH values. Biomass accumulation did not seem to vary when cells were induced at different pH values, but was greatly affected by lower concentration of methanol. Purification of A33scFv from clarified medium was done using a two step chromatographic procedure using anion-exchange and hydrophobic interaction chromatography, resulting in 25% recovery and >90% purity. Pure A33scFv was tested for functionality using surface plasmon resonance and showed activity against immobilized A33 antigen. Our results demonstrate that functional A33scFv can be produced in sufficient quantities using P. pastoris for use in further functionality studies and diagnostic applications. PMID- 15294277 TI - Two-step purification of Bacillus circulans chitinase A1 expressed in Escherichia coli periplasm. AB - A protein purification procedure was developed to efficiently and effectively purify the target enzyme, chitinase A1 of Bacillus circulans WL-12, from Escherichia coli DH5alpha carrying the chiA gene with its natural promoter in the plasmid pNTU110. Chitinase A1 was purified to apparent homogeneity from E. coli periplasm with a final recovery of 90.6%. Two main steps were included in this protein purification procedure, ammonium sulfate precipitation (40% saturation) and anion-exchange chromatography at pH 6.0 using Q Ceramic HyperD column. The yield of chitinase A1 was estimated at 95 microg/L. A polyclonal antibody against chitinase A1 was raised by immunizing BALB/c mice with chitinase A1 purified from E. coli DH5alpha(pNTU110). As indicated by Western blot analysis, a 3000-fold diluted antibody detected purified chitinase A1 from E. coli DH5alpha(pNTU110) in an amount of at least 1 ng and specifically detected chitinase A1 produced by B. circulans WL-12. PMID- 15294278 TI - Purification of ArcR, an oxidation-sensitive regulatory protein from Bacillus licheniformis. AB - In Bacillus licheniformis, ArcR, a transcriptional activator of the Crp/Fnr family, is required for expression of the anaerobic pathway of arginine catabolism, the arginine deiminase pathway. The method described here allows the purification of milligram quantities of functional ArcR from a recombinant Escherichia coli strain. The solubility properties of ArcR were much exploited during the purification process. The protein appeared highly sensitive to oxidation. Oxidation-induced precipitation of the protein was attributed to the formation of intermolecular disulfide bridges. Alkylation of mutant proteins with single substitutions showed that both cysteine residues of the protein, C178 and C205, are involved in formation of the disulfide bridges. Substitution of both cysteines yielded a functional protein insensitive to oxidation and able to form a complex with its cognate target on the DNA. PMID- 15294279 TI - High-level expression, purification, and characterization of recombinant wheat xylanase inhibitor TAXI-I secreted by the yeast Pichia pastoris. AB - Triticum aestivum xylanase inhibitor I (TAXI-I) is a wheat protein that inhibits microbial xylanases belonging to glycoside hydrolase family 11. In the present study, recombinant TAXI-I (rTAXI-I) was successfully produced by the methylotrophic yeast Pichia pastoris at high expression levels (approximately 75 mg/L). The rTAXI-I protein was purified from the P. pastoris culture medium using cation exchange and gel filtration chromatographic steps. rTAXI-I has an iso electric point of at least 9.3 and a mass spectrometry molecular mass of 42,013 Da indicative of one N-linked glycosylation. The recombinant protein fold was confirmed by circular dichroism spectroscopy. Xylanase inhibition by rTAXI-I was optimal at 20-30 degrees C and at pH 5.0. rTAXI-I still showed xylanase inhibition activity at 30 degrees C after a 40 min pre-incubation step at temperatures between 4 and 70 degrees C and after 2 h pre-incubation at room temperature at a pH ranging from 3.0 to 12.0, respectively. All tested glycoside hydrolase family 11 xylanases were inhibited by rTAXI-I whereas those belonging to family 10 were not. Specific inhibition activities against family 11 Aspergillus niger and Bacillus subtilis xylanases were 3570 and 2940IU/mg protein, respectively. The obtained biochemical characteristics of rTAXI-I produced by P. pastoris (no proteolytical cleft) were similar to those of natural TAXI-I (mixture of proteolytically processed and non-processed forms) and non glycosylated rTAXI-I expressed in Escherichia coli. The present results show that xylanase inhibition activity of TAXI-I is only affected to a limited degree by its glycosylation or proteolytic processing. PMID- 15294280 TI - Prokaryotic expression, polyclonal antibody preparation, and sub-cellular localization analysis of Na+, K+-ATPase beta2 subunit. AB - Na+, K+-ATPase beta2 subunit (NKA1b2) is not only a regulator of Na+, K+-ATPase, but also functions in the interaction between neuron and glia cells as a Ca2+ dependent adhesion molecule. To further study the function of NKA1b2, the anti NKA1b2 polyclonal antibody was prepared to recognize the outer-membrane carboxyl portion segment of NKA1b2. The coding region for amino acids 190-290 at the carboxyl portion of NKA1b2 (NKA1b2-CP) was sub-cloned into the vector pGEX-4T-2 and introduced into the Escherichia coli BL21(DE3) cell for efficient soluble expression. The amino acid sequence of expressed protein was determined using mass spectrometry following Mascot analysis. After purification, GST-NKA-beta2-CP was used to immunize the adult rabbits following standard protocols. The produced antiserum could detect the NKA1b2 protein expressed not only in the prokaryotic cells (E. coli) but also in the eukaryotic cells (COS7) transfected with NKA1b2 expression vector (pEGFP-NKA1b2). Furthermore, the antiserum was used for determining the localization of NKA1b2 in primary culture of neonatal rat neurons using immunohistochemical technique. Results demonstrated that NKA1b2 was localized both in the cytoplasm and cellular membrane. The preparation of anti NKA-beta2-CP polyclonal antibody will facilitate further functional study on NKA1b2. PMID- 15294281 TI - Purification and characterization of recombinant human cathepsin E expressed in human kidney cell line 293. AB - A cDNA encoding human prepro-cathepsin E was introduced into the adenovirus transformed HEK-293 (human embryonic kidney) cell line. The construct contained both a V5 peptide epitope and histidine tags at the carboxy terminus. Transfected cells efficiently secreted recombinant pro-cathepsin E into the culture medium. The secreted pro-cathepsin E was purified in a single step using Ni affinity chromatography yielding a protein of about 92 kDa under non-reducing conditions. The amino-terminal sequence of the purified protein began at Ser20, suggesting human cathepsin E accumulated in the culture supernatant as the pro-enzyme. The purified protein was rapidly and completely converted to the active form by treatment at pH 4.0 or below. Steady state kinetic parameters for hydrolysis of the fluorogenic peptide substrate MOCAc-Gly-Lys-Pro-Ile-Leu-Phe-Phe-Arg-Leu Lys(Dnp)-d-Arg-NH2 (cleavage at the Phe-Phe bond) were consistent with previously reported values for purified human enzyme (kc/Ki= 53 x 10(6) M(-1) s(-1), Km= 6.3 microM, and kcat= 3 x 10(2) s(-1)). The activated protein was potently inhibited by pepstatin with Ki= 0.2 nM, as well as a reported beta secretase inhibitor. This work demonstrates the potential for producing large quantities of highly purified human cathepsin E from HEK-293 cells in quantities to support both biochemical and structural characterization of the enzyme. PMID- 15294282 TI - Purification and biochemical characterization of simplified eukaryotic nitrate reductase expressed in Pichia pastoris. AB - NAD(P)H:nitrate reductase (NaR, EC 1.7.1.1-3) is a useful enzyme in biotechnological applications, but it is very complex in structure and contains three cofactors-flavin adenine dinucleotide, heme-Fe, and molybdenum molybdopterin (Mo-MPT). A simplified nitrate reductase (S-NaR1) consisting of Mo MPT-binding site and nitrate-reducing active site was engineered from yeast Pichia angusta NaR cDNA (YNaR1). S-NaR1 was cytosolically expressed in high density fermenter culture of methylotrophic yeast Pichia pastoris. Total amount of S-NaR1 protein produced was approximately 0.5 g per 10 L fermenter run, and methanol phase productivity was 5 microg protein/g wet cell weight/h. Gene copy number in genomic DNA of different clones showed direct correlation with the expression level. S-NaR1 was purified to homogeneity in one step by immobilized metal affinity chromatography (IMAC) and total amount of purified protein per run of fermentation was approximately 180 mg. Polypeptide size was approximately 55 kDa from electrophoretic analysis, and S-NaR1 was mainly homo-tetrameric in its active form, as shown by gel filtration. S-NaR1 accepted electrons efficiently from reduced bromphenol blue (kcat = 2081 s(-1)) and less so from reduced methyl viologen (kcat = 159 s(-1)). The nitrate KM for S-NaR1 was 30 +/- 3 microM, which is very similar to YNaR1. S-NaR1 is capable of specific nitrate reduction, and direct electric current, as shown by catalytic nitrate reduction using protein film cyclic voltammetry, can drive this reaction. Thus, S-NaR1 is an ideal form of this enzyme for commercial applications, such as an enzymatic nitrate biosensor formulated with S-NaR1 interfaced to an electrode system. PMID- 15294283 TI - High-level expression and characterization of a highly functional Comamonas acidovorans xanthine dehydrogenase in Pseudomonas aeruginosa. AB - An improved procedure is described for the high-level expression of Comamonas acidovorans XDH in Pseudomonas aeruginosa PAO1-LAC. The level of functional expression (56 mg protein/L culture) is found to be 7-fold higher than that observed in Escherichia coli and 30-fold higher than that induced in C. acidovorans. Co-expression of the xdhC gene is required for maximal level of functional expression. Comparison of purified preparations of XDH expressed in the absence of xdhC (XDH(AB)) with that expressed in its presence (XDH(ABC)) shows the increased level of activity due to the level of Mo incorporation. The Fe and FAD contents of expressed enzymes are independent of xdhC co-expression. Electron paramagnetic resonance spectroscopy, circular dichroism spectroscopy, metal analysis, and kinetic properties of recombinant purified XDH(ABC) are identical with those exhibited by the native enzyme. This expression system should serve as a valuable tool for further biophysical and mechanistic investigations of xanthine dehydrogenase by site-directed mutagenesis. A method is also described to evaluate the suitability of P. aeruginosa and other organisms as potential expression hosts for five different sources of xdh genes. PMID- 15294284 TI - Codon optimization, expression, and characterization of recombinant lumbrokinase in goat milk. AB - Lumbrokinase is an important fibrinolytic enzyme derived from earthworm. Although its cDNA has been isolated and sequenced, there is still no report on expression of the lumbrokinase due to unknown reasons. To determine the elements affecting the expression of lumbrokinase, two copies of a lumbrokinase cDNA(w) obtained by RT-PCR and a synthesized lumbrokinase cDNA(m) with optimized codons were cloned into a mammary-gland-specific expression vector pIbCP. The pIbCP-LK-LK vector preparations were directly injected in the lactating goat mammary glands. Results showed that both LK-w and LK-m were successfully expressed in goat milk. The fibrinolytic activity of the LK-w in milk was 225,000 +/- 13,200 tPA units/L, while that of the LK-m was 550,000 +/- 21,600 tPA units/L, indicating that the codon optimization plays an important role in improving the lumbrokinase expression. The molecular weight of the recombinant lumbrokinase is 31.8 kDa. The main physiochemical features of the recombinant lumbrokinase, including temperature stability, pH resistance, and sensitivity to pepsin, were also clarified. This is the first report on expression and characterization of a genetically engineered lumbrokinase. PMID- 15294285 TI - Expression of functional human coagulation factor XIII A-domain in plant cell suspensions and whole plants. AB - Coagulation factor XIII, a zymogen present in blood as a tetramer (A2B2) of A- and B-domains, is one of the components of many "wound sealants" which are proposed for use or currently in use as effective hemostatic agents, sealants, and tissue adhesives in surgery. After activation by alpha-thrombin cleavage, coagulation factor XIII A-domain, a transglutaminase, is formed and catalyzes the covalent cross-linking of the alpha- and gamma-chains of linear fibrin to form homopolymers, which can quickly stop bleeding. We have successfully expressed the A-domain of factor XIII in both plant cell cultures and whole plants. Transgenic plant cell culture allows a rapid method for testing production feasibility while expression in whole plants demonstrates an economic production system for recombinant human plasma-based proteins. The expressed factor XIII A-domain had a similar size as that of human plasma-derived factor XIII. Crude plant extract containing recombinant factor XIII A-domain showed transglutaminase activity with monodansylcadaverine and casein as substrates and cross-linking activity in the presence of linear fibrin. The expression of factor XIII A-domain was not affected by plant leaf position. PMID- 15294286 TI - Functional expression of hexahistidine-tagged beta-subunit of yeast F1-ATPase and isolation of the enzyme by immobilized metal affinity chromatography. AB - Mitochondrial ATP synthase (F1Fo-ATPase) catalyzes the terminal step of oxidative phosphorylation. In this paper, we demonstrate the functional expression of the hexahistidine-tagged beta-subunit of yeast ATP synthase and the purification of the F1-ATPase from yeast cells. A gene encoding the beta-subunit from Saccharomyces cerevisiae was modified to encode a protein of which the original N terminus import signal sequence was replaced by a sequence containing the import signal sequence of a mitochondrial ATPase inhibitor, its processing site, and six consecutive histidines. Expression of the modified gene generated a functional F1Fo complex in host yeast cells lacking a functional copy of the endogenous ATP2 gene, as judged by growth of rescued cells on lactate medium. F1 was extracted from the yeast mitochondria by chloroform treatment and purified by immobilized metal affinity chromatography and gel filtration chromatography. The specific activity of the purified F1 was comparable to that of the wild-type enzyme, and the F1 contained all of the 5 known subunits (alpha, beta, gamma, delta, and epsilon). Moreover, the activity of the F1 was completely inhibited by the specific ATPase inhibitor protein, IF1. These results indicate that F1 containing the tagged beta-subunit is fully assembled and active. The application of this novel procedure simplifies the number of steps required for the isolation of F1 used for studying the molecular mechanism of catalysis and regulation of the enzyme. PMID- 15294287 TI - Cloning and expression of an acidic platelet aggregation inhibitor phospholipase A2 cDNA from Bothrops jararacussu venom gland. AB - The phospholipase A2 (PLA2, E.C. 3.1.1.4) superfamily is defined by enzymes that catalyze the hydrolysis of the sn-2 bond of phosphoglycerides. Most PLA2s from the venom of Bothrops species are basic proteins, which have been well characterized both structurally and functionally, however, little is known about acidic PLA2s from this venom. Nevertheless, it has been demonstrated that they are non-toxic, with high catalytic and hypotensive activities and show the ability to inhibit platelet aggregation. To further understand the function of these proteins, we have isolated a cDNA that encodes an acidic PLA2 from a cDNA library prepared from the poly(A)+ RNA of venom gland of Bothrops jararacussu. The full-length nucleotide sequence of 366 base pairs encodes a predicted gene product with 122 amino acid with theoretical isoelectric point and size of 5.28 and 13,685 kDa, respectively. This acidic PLA2 sequence was cloned into expression vector pET11a (+) and expressed as inclusion bodies in Escherichia coli BL21(DE3)pLysS. The N-terminal amino acid sequence of the 14 kDa recombinant protein was determined. The recombinant acidic PLA2 protein was submitted to refolding and to be purified by RP-HPLC chromatography. The structure and function of the recombinant protein was compared to that of the native protein by circular dichroism (CD), enzymatic activity, edema-inducing, and platelet aggregation inhibition activities. PMID- 15294288 TI - Engineering of Escherichia coli to improve the purification of periplasmic Fab' fragments: changing the pI of the chromosomally encoded PhoS/PstS protein. AB - Escherichia coli is a widely used host for the heterologous expression of proteins of therapeutic and commercial interest. The scale and speed at which it can be cultured can result in the rapid generation of large quantities of product. However, to achieve low costs of production a simple and robust purification process is also required. The general factors that impact on the cost of a purification process are the scale at which a process can be performed, the cost of the purification matrix, and the number and complexity of the chromatographic steps employed. Purification of Fab' fragments of antibodies from the periplasm of E. coli using ion exchange chromatography can result in the co purification of E. coli host proteins having similar functional pI: such as the periplasmic phosphate binding protein, PhoS/PstS. In such circumstances, an additional chromatographic step is required to separate Fab' from PhoS. Here, we change the functional pI of the chromosomally encoded PhoS/PstS to effect its non purification with Fab' fragments, enabling the removal of an entire chromatographic step. This exemplifies the strategy of the modification of host proteins with the aim of simplifying the production of heterologous proteins. PMID- 15294289 TI - Characterisation of bacterially expressed structural protein E2 of hepatitis C virus. AB - The E2 glycoprotein is a structural component of the hepatitis C virus (HCV) virion. It interacts with putative cellular receptors, elicits production of neutralising antibodies against the virus, and is involved in viral morphogenesis. The protein is considered as a major candidate for anti-HCV vaccine. Despite this, relatively little is known about this protein. Previous studies have focused on the antigenic and functional analysis of the glycosylated forms. This report describes expression of the ectodomain of E2 (recE2) in Escherichia coli cells, its purification, and initial characterisation of its structural and functional properties. It is demonstrated that the purified protein forms small soluble aggregates, which retain functional characteristics of its native counterpart, i.e., it interacts with a putative cellular receptor, CD81, and is recognised by both conformation-dependent and -independent anti-E2 monoclonal antibodies. PMID- 15294290 TI - Homologous expression of the feruloyl esterase B gene from Aspergillus niger and characterization of the recombinant enzyme. AB - The faeB gene encoding the feruloyl esterase B (FAEB) was isolated from Aspergillus niger BRFM131 genomic DNA. The faeB gene, with additional sequence coding for a C-terminal histidine tag, was inserted into an expression vector under the control of the gpd promoter and trpC terminator and expressed in a protease deficient A. niger strain. Homologous overproduction allows to reach an esterase activity of 18 nkat mL(-1) against MCA as substrate. The improvement factor was 16-fold higher as compared to the production level obtained with non transformed A. niger strain induced by sugar beet pulp. The corresponding secretion yield was estimated to be around 100 mg L(-1). Recombinant FAEB was purified 14.6-fold to homogeneity from an 8-day-old culture by a single affinity chromatographic step with a recovery of 64%. SDS-PAGE revealed a single band with a molecular mass of 75 kDa, while under non-denatured conditions, native enzyme has a molecular mass of around 150 kDa confirming that the recombinant FAEB is a homodimer. The recombinant and native FAEB have the same characteristics concerning temperature and pH optima, i.e., 50 degrees C and 6, respectively. In addition, the recombinant FAEB was determined to be quite stable up to 50 degrees C for 120 min. Kinetic constants for MCA, MpCA, and chlorogenic acid (5-O caffeoyl quinic acid) were as follows: Km: 0.13, 0.029, and 0.16 mM and Vmax: 1101, 527.6, and 28.3 nkat mg(-1), respectively. This is the first report on the homologous overproduction of feruloyl esterase B in A. niger. PMID- 15294291 TI - Decreases in yeast expression yields of the human adenosine A2a receptor are a result of translational or post-translational events. AB - The human adenosine receptor (A2a), a G-protein-coupled receptor (GPCR), was C terminally tagged with the green fluorescent protein (GFP) and expressed in the yeast Saccharomyces cerevisiae to gain an understanding of the expression limitations of this medically relevant class of membrane proteins. The A2a-GFP protein was able to bind adenosine analogs indicating that the GFP tag did not alter the ligand binding activity of the receptor. A screen based on whole cell fluorescence was developed and a library of clones with various gene copy numbers was screened via flow cytometry to isolate clones with the highest protein expression levels. All clones studied exhibited a decrease in the net A2a-GFP protein production rate over time as determined by whole cell fluorescence, Western blotting, confocal microscopy, and ligand binding. Quantitative PCR showed that A2a-GFP mRNA levels remained relatively high even as the protein production rate decreased. A cycloheximide chase experiment showed that the mature protein was stable over time and was not significantly degraded. Taken together, these results suggest that heterologous expression of GPCRs is limited by a translational or post-translational bottleneck that is unique from expression limitations seen for soluble proteins. PMID- 15294292 TI - Purification and characterization of hepatitis C virus non-structural protein 5A expressed in Escherichia coli. AB - We have employed a pET-ubiquitin expression system to produce two his-tagged forms of hepatitis C virus (HCV) non-structural protein 5A (NS5A) in Escherichia coli. One derivative contains the full-length protein extended to include a carboxy-terminal hexahistidine tag; the other derivative contains an amino terminal hexahistidine tag in place of the 32 amino acid amphipathic helix that mediates membrane association. At least 1 mg of each derivative at a purity of 90% could be produced from a 1-L culture. The purified derivatives produced high titer antibody that recognized both p56 and p58 forms of NS5A in Huh-7.5 cells expressing an HCV subgenomic replicon. The NS5A derivatives were efficiently phosphorylated by casein kinase II, leading to at least 5 mol of phosphate incorporated per mole of protein. Interestingly, this level of phosphorylation did not alter the migration of the protein in an SDS-polyacrylamide gel, suggesting that hyperphosphorylation alone is not sufficient to generate the p58 form of NS5A observed in Huh-7 cells. Neither NS5A derivative was capable of inhibiting the eIF2alpha-phosphorylation activity of the activated form of the double-stranded RNA-activated protein kinase, PKR, suggesting that NS5A phosphorylation may be required for this function of NS5A. However, both unphosphorylated derivatives were shown to interact with NS5B, the HCV RNA dependent RNA polymerase, in solution by using a novel kinase-protection assay. The availability of purified HCV NS5A will permit rigorous biochemical and biophysical characterization of this protein, ultimately providing insight into the function of this protein during HCV genome replication. PMID- 15294293 TI - Improved expression, purification, and crystallization of p38alpha MAP kinase. AB - p38alpha mitogen-activated protein (MAP) kinase is widely expressed in many mammalian tissues and is activated as a part of signal transduction cascades that respond to inflammatory stimuli. The activation of p38 is known to trigger various biological effects, including cell death, differentiation, and proliferation. The central role played by p38alpha in cellular signaling events, including those that control a wide range of inflammatory and autoimmune diseases, makes it an attractive drug target. To develop optimized small molecule therapeutics targeting p38alpha, different techniques must be employed for the detailed biochemical, biophysical, and structural characterization of the interactions of p38alpha with lead compounds. These methods typically require large quantities of highly purified p38alpha protein. We describe here an improved expression and purification method for recombinant p38alpha production that reproducibly yields over 70 mg of highly purified protein per liter of shake flask bacterial culture. This yield is significantly higher than that previously reported for p38alpha production in Escherichia coli. We achieved a significant increase in soluble p38alpha protein expression by using the genetically modified E. coli strain BL21 DE3 Rosetta, which is optimized for expression of eukaryotic proteins with codons rarely used in E. coli. The p38alpha protein was purified to near homogeneity using a simple two-step procedure including nickel-chelating Sepharose chromatography followed by anion-exchange chromatography using MonoQ resin. Purified p38alpha was characterized using the standard commercially available small molecule inhibitor SB-203580. The binding association and dissociation rate constants determined by Biacore are in excellent agreement with previously reported values. The purified p38alpha protein was efficiently activated by MKK6 kinase to yield phosphorylated p38alpha. Purified p38alpha protein was also successfully crystallized, producing crystals diffracting to 1.9 angstroms, exceeding the highest resolution for p38alpha reported in the Protein DataBank. The simplicity and efficiency of this approach should prove useful for many laboratories that are interested in production of p38alpha for biochemical and biophysical studies and structure-based drug design. PMID- 15294294 TI - High level production and one-step purification of biologically active ectodysplasin A1 and A2 immunoadhesins using the baculovirus/insect cell expression system. AB - Ectodysplasin A (EDA) is a ligand of the tumor necrosis factor (TNF) family that has been shown to play a crucial role in ectodermal differentiation. Mutations of the syntenic ectodysplasin A gene (Eda) are responsible for Tabby (Ta) phenotype in mice and human X-linked hypohidrotic ectodermal dysplasia (XLHED). EDA-A1 and EDA-A2 are the two main splice variants of Eda, which differ from each other in only two amino acid residues and engage the tumor necrosis factor (TNF) family receptors EDAR and XEDAR, respectively. We have used the baculovirus/insect cell system to express the recombinant EDA proteins fused to the Fc portion of a truncated human IgG1 immunoglobulin heavy chain. Immunoadhesins (4.5-4.7 mg/L) from crude supernatant could be purified to near homogeneity by using rProtein A affinity chromatography. The purified EDA immunoadhesins were endowed with ligand binding activity as they could bind EDAR or XEDAR on the surface of 293T cells that had been transiently transfected with the corresponding plasmids. Functional activities of EDA immunoadhesins were demonstrated by their ability to activate the NF-kappaB pathway in cells expressing their cognate receptors. These results open up the possibility of obtaining large amounts of purified EDA proteins to investigate EDAR/XEDAR related signaling pathways and for the treatment of patients with X-linked hypohidrotic ectodermal dysplasia. PMID- 15294295 TI - Overexpression and characterization of two unknown proteins, YicI and YihQ, originated from Escherichia coli. AB - The proteins encoded in the yicI and yihQ gene of Escherichia coli have similarities in the amino acid sequences to glycoside hydrolase family 31 enzymes, but they have not been detected as the active enzymes. The functions of the two proteins have been first clarified in this study. Recombinant YicI and YihQ produced in E. coli were purified and characterized. YicI has the activity of alpha-xylosidase. YicI existing as a hexamer shows optimal pH at 7.0 and is stable in the pH range of 4.7-10.1 with incubation for 24h at 4 degrees C and also is stable up to 47 degrees C with incubation for 15 min. The enzyme shows higher activity against alpha-xylosyl fluoride, isoprimeverose (6-O-alpha xylopyranosyl-glucopyranose), and alpha-xyloside in xyloglucan oligosaccharides. The alpha-xylosidase catalyzes the transfer of alpha-xylosyl residue from alpha xyloside to xylose, glucose, mannose, fructose, maltose, isomaltose, nigerose, kojibiose, sucrose, and trehalose. YihQ exhibits the hydrolysis activity against alpha-glucosyl fluoride, and so is an alpha-glucosidase, although the natural substrates, such as alpha-glucobioses, are scarcely hydrolyzed. alpha-Glucosidase has been found for the first time in E. coli. PMID- 15294296 TI - On-column refolding of recombinant human interleukin-4 from inclusion bodies. AB - Interleukin-4 (IL4) is a multifunctional cytokine which plays a key role in the immune system. Several antagonists/agonists of IL4 are reported through mutagenesis studies, but their solution structural studies using nuclear magnetic resonance (NMR) spectroscopy are hindered as milligram quantities of isotopically labeled protein are required for structural refinements. In this work, a His tagged recombinant form of human IL4 was overexpressed in Escherichia coli under the control of a T7 promoter. The resulting inclusion bodies were separated from cellular debris by centrifugation and solubilized by 6M guanidine-HCl in the presence of reducing agents. The denatured IL4 was immobilized on Ni2+-fractogel beads and refolded in a single chromatographic step by gradual removal of denaturant. This protocol yielded 15-20 mg of isotope-enriched protein from 1L of culture grown in minimal medium. The refolded protein was highly pure and was correctly folded as judged by its two-dimensional NMR spectrum. To show the successful application of this refolding protocol to IL4 variants, 15N-labeled Y124D-IL4 was also prepared and its first two-dimensional NMR spectrum was presented. PMID- 15294297 TI - The C-terminus of botulinum neurotoxin type A light chain contributes to solubility, catalysis, and stability. AB - Botulinum neurotoxin type A (BoNT/A) is the etiological agent responsible for botulism, a disease characterized by peripheral neuromuscular blockade. BoNT/A is produced by Clostridium botulinum as a single chain protein that is activated by proteolytic cleavage to form a 50 kDa light chain (LC, 448 amino acids) and a disulfide bond-linked 100 kDa heavy chain (HC, 847 amino acids). Whilst HC comprises the receptor binding and translocation domains, LC is a Zn2+ endopeptidase that cleaves at a single glutaminyl-arginine bond corresponding to residues 197 and 198 at the C-terminus of SNAP25. Cleavage of SNAP25 uncouples the neural exocytosis docking/fusion machinery. LC/A (LC 1-448) and several C terminal deletion proteins of LC/A were engineered and expressed as His-tagged fusion proteins in Escherichia coli. LC 1-448 was purified, but precipitated upon storage. Approximately 40% of LC 1-448 was a covalent dimer due to the formation of inter-chain disulfide bond formation at Cys430. Conversion of Cys430 to Ser abolished dimer formation of LC 1-448, but did not improve solubility. Three C terminal deletion peptides were engineered; LC 1-425 and LC 1-418 were expressed and could be purified as soluble and stable proteins, whilst LC 1-398 was soluble, but not stable to storage. Kinetic studies showed that LC 1-448 and LC 1 425 efficiently cleaved GST-SNAP25 and the fluorescent substrate SNAPtide, while LC 1-418 catalyzed the cleavage of both the SNAP25 and the fluorescent substrate SNAPtide with a similar Km, but at a 10-fold slower kcat. Thus, regions within the C-terminus of LC/A contribute to solubility, stability, and catalysis. PMID- 15294299 TI - The toxicity of recombinant proteins in Escherichia coli: a comparison of overexpression in BL21(DE3), C41(DE3), and C43(DE3). AB - Two mutant strains of Escherichia coli BL21(DE3), called C41(DE3) and C43(DE3) and originally described by Miroux and Walker, are frequently used to overcome the toxicity associated with overexpressing recombinant proteins using the bacteriophage T7 RNA polymerase expression system. Even when the toxicity of the plasmids is so high that it prevents transformation in the strain BL21(DE3), the toxic proteins can often be expressed successfully in C41(DE3) and/or C43(DE3). In this work, using a range of plasmids coding for several types of proteins, we investigated in BL21(DE3), C41(DE3), and C43(DE3) their ability to undergo transformation and to express. While transformation was always possible in C41(DE3) and C43(DE3), we could not obtain transformants in BL21(DE3) for 62% of the expression vectors tested. Moreover, after induction, the expression of heterologous proteins in both mutant strains is generally better than in BL21(DE3). In this study, we also enhanced the stability of plasmids in culture during the expression of proteins by adding the par locus from the plasmid pSC101 to the vector backbone. The stability of a subset of the plasmids (measured 3 h after induction) was determined in C41(DE3) and C43(DE3) and varies from 62 to 92% for C43(DE3) and from 10 to 90% for C41(DE3). This study demonstrates the usefulness of these strains C41(DE3) and C43(DE3) in solving the problem of plasmid instability during the expression of toxic recombinant proteins. PMID- 15294298 TI - HIV-1 gp120 V3 cholera toxin B subunit fusion gene expression in transgenic potato. AB - A cDNA fragment encoding the V3 loop of human immunodeficiency virus type-1 (HIV 1) envelope glycoprotein gp120 was fused to the cholera toxin B subunit gene (CTB gp120) and transferred into Solanum tuberosum cells by Agrobacterium tumefaciens mediated transformation. The CTB-gp120 fusion gene was detected in genomic DNA from transformed potato leaves by PCR DNA amplification. Synthesis and assembly of the CTB-gp120 fusion protein into oligomeric structures of pentamer size was detected in transformed tuber extracts by immunoblot analysis. The binding of CTB gp120 fusion protein pentamers to intestinal epithelial cell membrane glycolipid receptors was quantified by GM1-ganglioside enzyme-linked immunosorbent assay (GM1-ELISA). The ELISA results indicated that CTB-gp120 fusion protein made up 0.002-0.004% of the total soluble tuber protein. Synthesis of CTB-gp120 monomers and their assembly into biologically active oligomers in transformed potato tuber tissues demonstrates for the first time the expression of HIV-1 gp120 in plants and emphasizes the feasibility of using edible plant-based vaccination for protection against HIV-1 infection. PMID- 15294300 TI - High-level expression and purification of a recombinant hBD-1 fused to LMM protein in Escherichia coli. AB - In this work, we present the production of an active 43 aa recombinant human beta defensin-1 (rhBD-1(43)) in Escherichia coli AD202 cells using specific pLMM1-rhBD 1 expression system. Unique solubility properties of the C-terminal fragment of light meromyosin (LMM) allowed us to overcome foreseeable problems with isolation procedures and toxicity caused by rhBD-1 to the host organism. As a result, the majority of fusion protein (LMM-rhBD-1(43)) was obtained in the soluble state, isolated by a low salt-high salt treatment of total cell protein. The rhBD-1(43) was cleaved from the fusion with Protease 4 and purified on CM Sepharose Fast Flow column with the yield of approximately 1 mg rhBD-1(43) from 6 g of wet weight cells. Purified rhBD-1(43) showed antimicrobial activity against E. coli ML-35p at a concentration of 129 microM. The procedure of rhBD-1 expression and purification we present can provide a reliable and simple method for production of different cationic peptides for biological studies. PMID- 15294301 TI - Expression of the melittin gene of Apis cerana cerana in Escherichia coli. AB - A cDNA encoding melittin in Apis cerana cerana was obtained by PCR from the recombinant plasmid and cloned into the GST fusion expression vector pGEX-4T-2 for expression of the protein. The expressed protein of about 29 kDa was detected by Western blot and triple antibody sandwich ELISA, indicating that the recombinant protein is the fusion protein of GST-AccM. The expression conditions of GST-AccM fusion protein for Escherichia coli BL21 transformant were optimized. Thin layer scanning on the SDS-PAGE profiles of GST-AccM showed that the expressed protein accumulated up to about 15.2% of total protein of bacterial cells under the optimized expression condition. Purified and recovered recombinant melittin of A. cerana cerana showed bioactivity in activating rabbit platelets to aggregate. PMID- 15294302 TI - Molecular cloning, expression, purification, and characterization of fructose 1,6 bisphosphate aldolase from Mycobacterium tuberculosis--a novel Class II A tetramer. AB - The Class II fructose 1,6-bisphosphate aldolase (fda, Rv0363c) from the pathogen Mycobacterium tuberculosis H37RV was subcloned in the Escherichia coli vector pT7 7 and purified to near homogeneity. The specific activity (35 U/mg) is approximately 9 times higher than previously reported for the enzyme partially purified from the pathogen. Attempts to express the enzyme with an N-terminal fusion tag yielded inactive, mostly insoluble protein. The native recombinant enzyme is zinc-dependent and has a catalytic efficiency for fructose 1,6 bisphosphate cleavage higher than most Class II aldolases characterized to date. The aldolase has a Km of 20 microM, a kcat of 21 s(-1), and a pH optimum of 7.8. The molecular mass of the enzyme subunits as determined by mass spectrometry is in agreement with the mass calculated on the basis of its gene sequence minus the terminal methionine, 36,413 Da. The enzyme is a homotetramer and retains only two zinc ions per tetramer when transferred to a metal-free buffer, as determined by ICP-MS and by a colorimetric assay using 4-(2-pyridylazo)-resorcinol (PAR) as a chelator. The E. coli expression system reported in this study will facilitate the further characterization of this enzyme and the screening for potential inhibitors. PMID- 15294303 TI - Expression of bioactive recombinant GSLL-39, a variant of human antimicrobial peptide LL-37, in Escherichia coli. AB - The human cationic antimicrobial peptide hCAP-18/LL-37 is the unique cathelicidin identified in human to date. It has broad spectrum of antimicrobial activities and LPS-neutralizing activity and is involved in angiogenesis. Both purified and synthetic LL-37 or its derivatives were used in the study on LL-37. However, production of LL-37 in Escherichia coli has not been established. In this study, its precursor instead of the mature peptide was adopted for expression to avoid the lethal effect of recombinant LL-37 on host cells. A thrombin recognition site was introduced between the cathelin-like domain and LL-37 domain by overlap PCR to construct fragment encoding modified precursor (mhCAP-18) to facilitate the final release of the recombinant peptide. Then mhCAP-18 was fused in-frame to thioredoxin gene under the control of inducible T7 promoter to construct expression vector pET-mhCAP-18. The soluble form fusion protein was expressed in E. coli and purified by Chelating Sepharose column chromatography. Thrombin digestion of the fusion protein yielded recombinant GSLL-39, which was then purified by cation-exchange chromatography. Recombinant GSLL-39, which has two extra residues on its N-terminus when compared with its native counterpart, showed similar antimicrobial activities against both Gram-negative and Gram positive bacteria. PMID- 15294304 TI - A novel strategy for the expression and purification of the DNA methyltransferase, M.AhdI. AB - Biochemical and structural studies of the methylase from the type 1 1/2 R-M system AhdI require the ability to purify this multi-subunit enzyme in significant quantities in a soluble and active form. Several Escherichia coli expression systems were tested for their ability to produce the intact methylase but this could not be achieved in a simple co-expression system. Expression experiments were optimised to produce high yields of soluble M and S subunits as individual proteins. Temperature and conditions of induction proved to be the most useful factors and although purification of the S subunit was successful, an efficient strategy for the M subunit remained elusive. A novel strategy was developed in which individual subunits are expressed separately and the bacterial cells mixed before lysis. This method produced a high yield of the multi-subunit methylase when purified to homogeneity by means of heparin and size-exclusion chromatography. It was found to be essential, however, to remove tightly bound DNA by ammonium sulphate precipitation in 1 M NaCl. The intact methylase can now be consistently produced, avoiding the use of fusion proteins. The purified enzyme is stable over long time periods, unlike the individual subunits. This method may be of general application where the expression of multi-subunit proteins, or indeed their individual components, is problematic. PMID- 15294305 TI - In vivo biotinylated proteins as targets for phage-display selection experiments. AB - Screening phage-displayed combinatorial libraries represents an attractive method for identifying affinity reagents to target proteins. Two critical components of a successful selection experiment are having a pure target protein and its immobilization in a native conformation. To achieve both of these requirements in a single step, we have devised cytoplasmic expression vectors for expression of proteins that are tagged at the amino- or carboxy-terminus (pMCSG16 and 15) via the AviTag, which is biotinylated in vivo with concurrent expression of the BirA biotin ligase. To facilitate implementation in high-throughput applications, the engineered vectors, pMCSG15 and pMCSG16, also contain a ligase-independent cloning site (LIC), which permits up to 100% cloning efficiency. The expressed protein can be purified from bacterial cell lysates with immobilized metal affinity chromatography or streptavidin-coated magnetic beads, and the beads used directly to select phage from combinatorial libraries. From selections using the N-terminally biotinylated version of one target protein, a peptide ligand (Kd= 9 microM) was recovered that bound in a format-dependent manner. To demonstrate the utility of pMCSG16, a set of 192 open reading frames were cloned, and protein was expressed and immobilized for use in high-throughput selections of phage-display libraries. PMID- 15294306 TI - A self-cleavable sortase fusion for one-step purification of free recombinant proteins. AB - A new protein fusion system has been developed to generate free recombinant protein in a single affinity chromatographic step. The key component in the fusion is the catalytic core of sortase A from Staphylococcus aureus (SrtAc), which recognizes and cleaves the Thr-Gly bond at an LPXTG sequence with moderate activity. The fusion here consists of an N-terminal His6 tag, SrtAc, and an LPETG linker followed by protein of interest at the C-terminus. The fusion protein is expressed in Escherichia coli and purified by immobilized metal-ion affinity chromatography (IMAC). The immobilized fusion then undergoes on-column SrtAc mediated cleavage at the LPETG site in the presence of Ca2+ and/or triglycine. The target protein with an extra N-terminal glycine is released from the fusion while the N-terminal portion remains bound to the column. Because the cleavage enzyme SrtAc is co-expressed as a fusion with the target protein, the purification system eliminates exogenous proteolysis. This purification approach is simple, robust, inexpensive, time saving, and allows purification of free recombinant protein via one-step chromatography. PMID- 15294307 TI - QSPR treatment of rat blood:air, saline:air and olive oil:air partition coefficients using theoretical molecular descriptors. AB - A QSPR treatment has been applied to a data set that consists of 100 diverse organic compounds to relate the logarithmic function of rat blood:air, saline:air and olive oil:air partition coefficients (denoted by log K(b:a), log K(s:a), and log K(o:a), respectively), with theoretical molecular and fragment descriptors. Three QSPR models with squared correlation coefficients of 0.881, 0.926, and 0.922, respectively, were obtained. The verification of the predictive power of these models on a test set of 33 organic chemicals that were not included in the training set gave satisfactory squared correlation coefficients: 0.791 for rat blood:air, 0.794 for saline:air and 0.846 for olive oil:air. PMID- 15294308 TI - A reference data set for the evaluation of medical image retrieval systems. AB - Content-based image retrieval is starting to become an increasingly important factor in medical imaging research and image management systems. Several retrieval systems and methodologies exist and are used in a large variety of applications from automatic labelling of images to diagnostic aid and image classification. Still, it is very hard to compare the performance of these systems as the used databases often contain copyrighted or private images and are thus not interchangeable between research groups, also for patient privacy. Most of the currently used databases for evaluating systems are also fairly small which is partly due to the high cost in obtaining a gold standard or ground truth that is necessary for evaluation. Several large image databases, though without a gold standard, start to be available publicly, for example by the NIH (National Institutes for Health). This article describes the creation of a large medical image database that is used in a teaching file containing more than 8,700 varied medical images. The images are anonymised and can be exchanged free of charge and copyright. Ground truth (a gold standard) has been obtained for a set of 26 images being selected as query topics for content-based query by image example. To reduce the time for the generation of ground truth, pooling methods well known from the text or information retrieval field have been used. Such a database is a good starting point for comparing the current image retrieval systems and to measure the retrieval quality, especially within the context of teaching files, image case databases and the support of teaching. For a comparison of retrieval systems for diagnostic aid, specialised image databases, including the diagnosis and a case description will need to be made available, as well, including gold standards for a proper system evaluation. A first evaluation event for image retrieval is foreseen at the 2004 CLEF conference (Cross Language Evaluation Forum) to compare text-and content-based access mechanism to images. PMID- 15294309 TI - Automatic spinal disease diagnoses assisted by 3D unaligned transverse CT slices. AB - This paper describes a three-dimensional (3D) reconstruction method and an automatic diagnosis method for spinal diseases using unaligned transverse slices that have arbitrary angles and intervals but do not intersect to each other at regions of interest. The 3D reconstruction method extends the marching cube algorithm to generate triangulated isosurfaces for these unaligned slices. The automatic spinal disease diagnostic method analyses boundaries of discs and vertebral bodies on unaligned transverse slices to estimate the presence and extent of disc herniation and canal compression, and deformities in the spinal curve. The prototype system can be used as a qualitative and quantitative tool for the diagnosis of various spinal diseases using unaligned transverse slices. PMID- 15294310 TI - Processing X-ray images to eliminate irrelevant structures that mask important features. AB - In a plane radiographic image, there generally is an important area of interest (AOI). Too often, the AOI is partially masked by images of other overlapping and underlying structures that may be in front of or behind the AOI. An important adjunct to radiological diagnosis would be the capability of eliminating images of such masking structures to isolate the AOI for more detailed examination. We described a computerized method that utilizes a stereo pair of plane X-ray images to enable radiologists to interact with these images for first identifying for the computer the AOI and then directing the computer to eliminate all structures in front of and behind the AOI. The result is a plane X-ray image or a stereo X ray image pair that includes only the AOI, but not any overlapping or underlying structures. The method uses a stereo pair of X-rays and the 3D perception of radiologists. 3D perception involves eye convergence and lens focus as well as cues, such as parallax and relative sizes. Convergence of the eyes is by far the strongest factor in 3D visualization. The horizontal separation or disparity between points in the left and right eye images on a screen or X-ray film produces convergence which determines an object's perceived depth in visual 3D space. All points in a given perceived depth plane have the same disparity on the screen. In theory, a given depth plane can be eliminated from the 3D image by shifting one image and then the other image of a stereo pair horizontally by the distance of the disparity of the depth plane, and subtracting. A new stereo image pair is thereby produced in which points only of the depth plane do not appear. However, in practical situations, certain artifacts arise that must be considered. The method has the potential for important applications in many areas of medical imaging processing. PMID- 15294311 TI - A relational-tubular (ReTu) deformable model for vasculature quantification of zebrafish embryo from microangiography image series. AB - Embryonic cardiovascular system plays a vital role in embryonic development of human and animal. In this work, we introduce a novel deformable model, which we called Relational-tubular (ReTu) deformable model for segmenting and quantifying the embryonic vasculature of zebrafish embryo from microangiography image series. Particularly, to incorporate additional constraints on the spatial relationships among vessel branches, we introduce a new energy term called relation energy into the model energy function. This energy item acts as a repulsion force between neighboring vessels during the deformation to encourage them to move towards their respective volume data. Using the ReTu deformable model, the deformation process is an iterative two-stage procedure: vascular axis deformation and vascular surface deformation. The efficiency and robustness of this approach are demonstrated by experiments which show that satisfactory quantifications of the vasculature can be obtained after 3-4 iterations. PMID- 15294312 TI - Cross-sectional area reduction of the aortic ostium by suprarenal stent wires: in vitro phantom study by CT virtual angioscopy. AB - The study aims to investigate the reduction of cross-sectional area of the aortic ostium by the presence of aortic stent wires observed using CT virtual angioscopy in an aorta phantom. A human aorta phantom was built with a commercial stent graft placed in situ to simulate a repaired aortic aneurysm. Virtual angioscopic images of the aortic ostium and stent wires were generated in the locations of renal arteries, superior mesenteric artery and corresponding cross-sectional area reduction caused by stent wires was measured by virtual angioscopy in various scanning parameters. Our study showed that cross-sectional area reduction of the aortic ostium was determined by the diameter of renal ostium and stent wires, as well as the number of stent wires crossing the aortic ostium. PMID- 15294313 TI - Computer-assisted evaluation of aortic stiffness using data acquired via magnetic resonance. AB - Aortic stiffness is frequently assessed through pulse wave velocity (PWV) measurements. Based on data acquired by magnetic resonance (MR) using a one dimensional time-of-flight technique, a new computational tool has been developed to rapidly construct flow velocity images and automatically calculate PWV. Comparison between PWV results obtained from this and a manual analysis demonstrates good agreement (correlation coefficient of 0.9951), while the new method improves the time efficiency by more than 20 times. The new method can also significantly improve flow signal quality and yield more credible results when strong interfering background signals are present. PMID- 15294314 TI - Three-dimensional fast field echo MR myelography using water excitation. AB - The purpose of this study is to determine if 3D FFE MRM using WE provides better image quality in comparison with that using SPIR. Twenty subjects were referred for the MRM images using SPIR, and WE with different TR. SNR and CNR were measured in the ganglions and adjacent tissues, in addition, the uniformity of the fat tissues was scored on a three-point scale by two radiologists with consensus. The WE sequence showed the higher SNR and CNR values than that using SPIR, in addition, the WE technique was helpful to achieve the shorter acquisition time of MRM. PMID- 15294315 TI - Molecular detection and characterization of Dichelobacter nodosus in ovine footrot in India. AB - Dichelobacter nodosus was detected in three clinical cases of ovine footrot in Kashmir, India. The detection was done by PCR in three clinical specimens directly, without isolating the organism, using species-specific 16S rDNA primers. Positive results were indicated by amplification of a 783 bp product. All the three samples were subjected to serogrouping by multiplex PCR using group (A-I) specific primers. All the three samples revealed the presence of serogroup B of D. nodosus by yielding a single band of 283 bps. PMID- 15294316 TI - Development of a multiplex PCR for detection of avian adenovirus, avian reovirus, infectious bursal disease virus, and chicken anemia virus. AB - A multiplex polymerase chain reaction (mPCR) was developed and optimized for the simultaneous detection and differentiation of avian reovirus (ARV), avian adenovirus group I (AAV-I), infectious bursal disease virus (IBDV), and chicken anemia virus (CAV). Four sets of specific oligonucleotide primers were used in this test for ARV, AAV-I, IBDV, and CAV. The mPCR DNA products were visualized by gel electrophoresis and consisted of fragments of 365 bp for IBDV, 421 bp for AAV I, 532 bp for ARV, and 676 bp for CAV. The mPCR assay developed in this study was found to be sensitive and specific. Detection of PCR-amplified DNA products was 100 pg for both CAV and IBDV, and 10pg for both ARV and AAV-I and this mPCR did not amplify nucleic acids from the other avian pathogens tested. The mPCR demonstrated similar sensitivity in tests using experimental fecal cloacal swab specimens that were spiked with ARV, AAV-1, IBDV, and CAV, and taken from specific pathogen free (SPF) chickens. This mPCR detected and differentiated various combinations of RNA/DNA templates from ARV, AAV-I, CAV, and IBDV without reduction of amplification from feces. PMID- 15294317 TI - Rapid detection of ethambutol-resistant Mycobacterium tuberculosis strains by PCR RFLP targeting embB codons 306 and 497 and iniA codon 501 mutations. AB - Mutations at embB gene codons 306 and 497 and iniA gene codon 501 occur frequently in ethambutol (EMB)-resistant Mycobacterium tuberculosis strains worldwide. The identification of these mutations in resistant strains has been achieved by labor-intensive DNA sequencing or by tedious amplification protocols followed by restriction endonuclease digestion. In this report, we describe PCR restriction fragment length polymorphism (RFLP)-based methods for determining substitutions at embB codons 306 and 497 and iniA codon 501 directly in BACTEC cultures of M. tuberculosis isolates. The wild-type and mutant alleles are revealed by easily interpretable and different RFLP patterns. The methods optimized initially on reference strains were tested directly on BACTEC cultures of 25 randomly selected clinical M. tuberculosis isolates, seven of which were determined to contain EMB-resistant strains by phenotypic drug susceptibility testing. The PCR-RFLP methods identified mutations in four of seven EMB-resistant strains with three isolates containing mutated embB codon 306 and one isolate containing mutated embB codon 497. The results of PCR-RFLP were confirmed by DNA sequencing. The worldwide prevalence figures for mutations at embB codons 306 and 497 and iniA codon 501 suggest that nearly half of EMB-resistant M. tuberculosis strains could be identified within one working day even in developing countries equipped with simple PCR technology instead of weeks required for phenotypic drug susceptibility testing. Further, since EMB resistance is also associated with multiple-drug resistance from some geographical locations, detection of EMB resistance may also lead to rapid identification of multidrug-resistant strains of M. tuberculosis. PMID- 15294318 TI - Rapid detection and differentiation of Bartonella spp. by a single-run real-time PCR. AB - A Real-time PCR for the identification and differentiation of Bartonella spp. based on the detection of mutations in an internal region of the gltA gene by thermal analysis was developed. The assay included a simultaneous detection of the amplicons by direct hybridization of LCRed and fluorescein-labelled probes coupled with melting curve analysis by the use of fluorescence resonance energy transfer technology. The protocol allowed establishing genospecie identity in less than 1 h without a need for restriction enzyme digestion or gel electrophoresis. The method is simple, reproducible, rapid, specific and potentially transferable to clinical samples. PMID- 15294319 TI - Extraction, detection and persistence of extracellular DNA in forest litter microcosms. AB - A DNA extraction method was developed that preferentially extracted extracellular DNA rather than intracellular DNA from forest litter. The method purposely avoided the use of harsh chemicals and physical disruption steps used in total DNA extraction to release DNA from cells. The detection limit of PCR, determined by spiking forest litter samples with a dilution series of Choristoneura fumiferana MNPVegt(-)/lacZ(+) genomic DNA, was about 1 ng DNA or 6.85 x 10(6) target copies 0.5 g(-1) moist forest litter or 0.14 g(-1) dry forest litter. In this study, outdoor terrestrial microcosms, each spiked with 49.2 microg of genomic DNA (from the baculovirus CfMNPVegt(-)/lacZ(+)), were exposed to summer conditions. A 530 bp DNA fragment from the genome of the baculovirus CfMNPVegt( )/lacZ(+) was detected in these microcosms for about 3 months. The DNA may have persisted for a longer period but was below the detection limit of the PCR analysis. PMID- 15294320 TI - A LightCycler real-time PCR hybridization probe assay for detecting food-borne thermophilic Campylobacter. AB - In a previous study, we reported the performance of a PCR assay amplifying 287-bp of the 16S rRNA gene of thermo-tolerant Campylobacter (C. jejuni, C. lari, C. coli) through an international ring-trial involving 12 participating laboratories. Based on the validated set of primers, a LightCycler real-time PCR assay (LC-PCR), which used fluorescent hybridization probes was developed. The test incorporated an internal amplification control co-amplified with the 16S rRNA gene of Campylobacter to monitor potential PCR inhibitors and ensure successful amplifications. The specificity study involving 39 Campylobacter and nine strains of other species indicated that the LC-PCR test was highly specific, giving cross-reactivity with only one strain of C. upsaliensis (CCUG19559). The sensitivity of the LC-PCR assay, evaluated in 32 spiked poultry-rinse or pork carcass-swab samples, was determined at 10CFU/ml carcass-rinse. The prevalence of samples positive for thermo-tolerant Campylobacter was 58.8% in 68 naturally contaminated poultry rinse samples tested by LC-PCR and the data were in good concordance with those of bacteriological method. The Ct values of the three replicates obtained for each sample tested in three different runs demonstrate that the LC-PCR was highly reproducible and afford a powerful tool for rapid detection of the thermo-tolerant Campylobacter strains. PMID- 15294321 TI - An improved 'cold SSCP' method for the genotypic and subgenotypic characterization of Cryptosporidium. AB - A simple, non-isotopic PCR-based single-strand conformation polymorphism ('cold SSCP') method is described which allows the efficient detection of genetic variation among and within genotypes of Cryptosporidium parvum. This low cost approach has important advantages over other 'genotyping' methods and is applicable to a wide range of genetic loci and organisms. PMID- 15294322 TI - Rapid detection of Salmonella by polymerase chain reaction. AB - Salmonella enterotoxin gene (stn) was sequenced from Salmonella enterica serotypes: Typhimurium, Typhi, Paratyphi A and B. The sequences from all the four serotypes showed complete homology with the already reported stn gene sequence of the serotype Typhimurium. As a tool for detection of this organism, four pairs of oligonucleotide primers were designed to amplify different fragments of this important pathological marker. The protocols were standardized with serotype Typhimurium in such a way so as to complete the PCR reaction in 75-90 min. These primers were found to generate specific amplicons with all the serotypes of Salmonella tested. The PCR protocols were found to be highly specific as no amplifications, specific or non-specific, were found when reactions were run using non-Salmonella DNA as template. The employment of a nested PCR markedly increased the sensitivity of the assay system in natural water samples. The protocol described herein is highly sensitive as it detects less than 10 cells of Salmonella in 250 microl of blood and approx. 1 cell in 1 ml of water without any enrichment. For the validation of this protocol, 72 coded samples of 11% skimmed milk spiked with different pathogens were received from NICED, Kolkata and analyzed for the presence of Salmonella. Our procedures detected correctly the presence of Salmonella in nine samples. 50 samples of raw milk were subjected to this PCR after enrichment for 8 h and 6 samples were found positive for Salmonella. The study indicates that Salmonella enterotoxin (stn) gene is highly conserved and the protocol devised in this study can be used as rapid and reliable method for detection of Salmonella spp. in water, milk and blood samples. PMID- 15294323 TI - Real-time allele-specific amplification for sensitive detection of the BRAF mutation V600E. AB - BRAF is a cytoplasmic serine/threonine kinase in the MAPK pathway that transduces signals from RAS family members to MEK1/2. Mutations in the BRAF gene have been described in the majority of cutaneous melanomas, papillary thyroid carcinoma and to a lesser extent in other cancers. The predominant mutation reported is a single transversion in exon 15 (T1799A). We designed a real-time allele-specific PCR to detect this mutation. This assay allowed us to detect this alteration in samples containing 2% of cells harboring this mutation, which is equivalent to 1% mutated DNA, assuming heterozygosity for the allele. Using this assay, we then tested 44 human primary colorectal tumors. We found the V600E mutation in four samples (9.1%). Analysis of DNA extracted from paraffin-embedded sections gave similar results, indicating that archived tissues can also be analyzed using this assay. The advantages of this method include: (1) rapidity; (2) sensitivity; (3) ease; (4) large throughput; (5) low cost and (6) the small quantities of DNA needed. PMID- 15294324 TI - Haplotype frequencies and linkage disequilibrium analysis of four frequent polymorphisms at the PTH/PTH-related peptide receptor gene locus. AB - The parathyroid hormone (PTH)/PTH-related peptide receptor is a critical component in the control of mineral ion metabolism and in bone development. This receptor is encoded by a single gene (PTHR1) on chromosome 3p21.1-p24.2, and mutations in this gene have been found in several clinical disorders of bone and mineral metabolism. To facilitate future genetic studies of this important gene, we determined haplotype frequencies and performed linkage disequilibrium (LD) analysis of four different polymorphisms at the PTHR1 locus. Combined analysis of Caucasian, African-American and Asian individuals indicated that LD exists between all but one pair of the four polymorphisms. However, the pattern of LD differed substantially among the three subpopulations; for example, LD between two closely spaced (154-bp apart) single nucleotide polymorphisms appeared to be present only in Asians. Depending on the population under study, genetic association studies may need to test even more closely spaced polymorphic markers when screening the PTHR1 locus. These findings may thus affect the design and interpretation of future genetic studies involving PTHR1. PMID- 15294325 TI - MRI measurements of water diffusion: impact of region of interest selection on ischemic quantification. AB - OBJECTIVE: To investigate the effect of ADC heterogeneity on region of interest (ROI) measurement of isotropic and anisotropic water diffusion in acute (< 12 h) cerebral infarctions. METHODS AND MATERIALS: Full diffusion tensor images were retrospectively analyzed in 32 patients with acute cerebral infarction. Fractional anisotropy (FA) and apparent diffusion coefficient (ADC) values were measured in ischemic lesions and in the corresponding contralateral, normal appearing brain by using four ROIs for each patient. The 2 x 2 pixel square ROIs were placed in the center, the lateral rim and the medial rim of the infarction. In addition, the whole volume of the infarction was measured using a free hand method. Each ROI value obtained from the ischemic lesion was normalized using contralateral normal ROI values. RESULTS: The localization of the ROIs in relation to the ischemic lesion significantly affected ADC measurement (P < 0.01, using Friedman test), but not FA measurement (P = 0.25). Significant differences were found between ADC values of the center of the infarction versus whole volume (P < 0.01), and medial rim versus whole volume of infarction (P < 0.001) with variation of relative ADC values up to 11%. The differences of absolute ADC for these groups were 22 and 23%, respectively. The lowest ADC was found in the center, followed by medial rim, lateral rim and whole volume of infarction. CONCLUSION: ADC quantification may provide variable results depending on ROI method. The ADC and FA values, obtained from the center of infarction tend to be lower compared to the periphery. The researchers who try to compare studies or work on ischemic quantification should be aware of these differences and effects. PMID- 15294326 TI - Observer variability based on the strength of MR scanners in the assessment of lumbar degenerative disc disease. AB - OBJECT: aim of this study was to analyse the observer variability in the diagnosis and definition of disc pathologies with low and high-field strength MR scanners. MATERIAL AND METHODS: 95 patients with low back pain or radicular pain who were referred from two different centers were included in the study. Fifty seven patients were scanned with 0.3 T MR (group 1) and 38 patients with 1.5 T (group 2). The intraobserver and interobserver reliability were assessed with the cappa coefficient which was characterised as follows: values less than 0.0 = 'poor' agreement, values 0.01-0.2 = 'slight' agreement beyond chance, 0.21-0.4 = 'fair' agreement, 0.41-0.60 = 'moderate' agreement, 0.61-0.80 = 'substantial' agreement and 0.81-1.00 = 'almost perfect' agreement. RESULTS: intraobserver agreement in group 1 and group 2 for both readers was 'almost perfect' in differentiating normal and pathological discs; 'substantial-almost perfect' in defining the disc pathologies, 'moderate-substantial' in root compression, and 'moderate-substantial' in spinal stenosis. Interobserver agreement was 'almost perfect' in differentiating normal and pathological discs, 'substantial' in defining disc pathologies, 'moderate' in root compression and 'moderate' in spinal stenosis in the group 1, whereas in group 2, it was 'almost perfect' in differentiating normal and pathological discs, 'almost perfect' in defining disc pathologies, 'slight-substantial' in root compression and 'moderate' in spinal stenosis. CONCLUSION: in the diagnosis of root compression and spinal stenosis, the intra and interobserver agreements were relatively poor with both high and low-strength field MRIs, indicating a need for more objective criteria. In differentiating normal and pathologic appearance of disc, the interobserver agreement was considerably better with high-field compared to low-field strength MRI. In cases where this definition is important, high-field strength scanners should be preferred. PMID- 15294327 TI - Dynamic MR imaging in Tolosa-Hunt syndrome. AB - OBJECTIVE: To evaluate the cavernous sinuses with dynamic magnetic resonance (MR) imaging in patients with Tolosa-Hunt syndrome (THS). METHODS: The sellar and parasellar regions of five patients with THS and 12 control subjects were examined with dynamic MR (1.5 T) imaging in the coronal plane. Dynamic images were obtained with spin-echo (SE) sequences in three patients, and with fast spin echo (FSE) sequences in two patients and control subjects. Conventional MR images of the cranium including sellar and parasellar regions were also obtained on T1 weighted pre- and post-contrast SE, and T2-weighted FSE sequences in the coronal plane. RESULTS: MR images revealed affected cavernous sinus with bulged convex lateral wall in three patients and concave lateral wall in two patients. In all control subjects, cavernous sinuses were observed with concave lateral wall. The signal intensity on T1- and T2-weighted images and contrast enhancement on post contrast images of the affected cavernous sinuses in patients were similar to those of the unaffected cavernous sinuses in patients and control subjects. The dynamic images in all patients disclosed small areas adjacent to the cranial nerve filling-defects within the enhanced venous spaces of the affected cavernous sinus, which showed slow and gradual enhancement from the early to the late dynamic images. No such gradually enhancing area was observed in control subjects except one. The follow-up dynamic MR images after corticosteroid therapy revealed complete resolution of the gradually enhancing areas in the previously affected cavernous sinus. CONCLUSION: Dynamic MR imaging may facilitate the diagnosis of THS. PMID- 15294328 TI - The cisternal segment of the abducens nerve in man: three-dimensional MR imaging. AB - PURPOSE: The goal of this study was to identify the abducens nerve in its cisternal segment by using three-dimensional turbo spin echo T2-weighted image (3DT2-TSE). The abducens nerve may arise from the medullopontine sulcus by one singular or two separated rootlets. MATERIAL AND METHODS: We studied 285 patients (150 males, 135 females, age range: 9-72 years, mean age: 33.3 +/- 14.4) referred to MR imaging of the inner ear, internal auditory canal and brainstem. All 3D T2 TSE studies were performed with a 1.5 T MR system. Imaging parameters used for 3DT2-TSE sequence were TR:4000, TE:150, and 0.70 mm slice thickness. A field of view of 160 mm and 256 x 256 matrix were used. The double rootlets of the abducens nerve and contralateral abducens nerves and their relationships with anatomical structures were searched in the subarachnoid space. RESULTS: We identified 540 of 570 abducens nerves (94.7%) in its complete cisternal course with certainty. Seventy-two cases (25.2%) in the present study had double rootlets of the abducens nerve. In 59 of these cases (34 on the right side and 25 on the left) presented with unilateral double rootlets of the abducens. Thirteen cases presented with bilateral double rootlets of the abducens (4.5%). CONCLUSION: An abducens nerve arising by two separate rootlets is not a rare variation. The detection of this anatomical variation by preoperative MR imaging is important to avoid partial damage of the nerve during surgical procedures. The 3DT2-TSE as a noninvasive technique makes it possible to obtain extremely high quality images of microstructures as cranial nerves and surrounding vessels in the cerebellopontine cistern. Therefore, preoperative MR imaging should be performed to detect anatomical variations of abducens nerve and to reduce the chance of operative injuries. PMID- 15294329 TI - Different metabolic patterns analysis of Parkinsonism on the 18F-FDG PET. AB - Idiopathic Parkinson's disease (IPD), progressive supranuclear palsy (PSP) and multiple system atrophy (MSA) are the most common movement disorders associated with neurodegenerative disease. A clinical differential diagnosis of IPD and atypical Parkinsonian disorders, such as MSA and PSP, is often complicated by the presence of symptoms common to both groups. Since Parkinsonism has a different pathophysiology in the cortical and subcortical brain structures, assessing the regional cerebral glucose metabolism may assist in making a differential diagnosis of Parkinsonism. The 18F-FDG PET images of IPD, MSA and PSP were assessed using statistical parametric mapping (SPM) in order to determine the useful metabolic patterns. Twenty-four patients with Parkinsonism: eight patients (mean age 67.9 +/- 10.7 years; M/F: 3/5) with IPD, nine patients (57.9 +/- 9.2 years; M/F: 4/5) with MSA and seven patients (67.6 +/- 4.8 years; M/F: 3/4) with PSP were enrolled in this study. All patients with Parkinsonism and 22 age matched normal controls underwent 18F-FDG PET, (after 370 MBq 18F-FDG). The three groups and the individual IPD, MSA and PSP patients were compared with a normal control group using a two-sided t-test of SPM (uncorrected P < 0.01, extent threshold > 100 voxel). The IPD, MSA and PSP groups showed significant hypometabolism in the cerebral neocortex compared to the normal control group. The MSA group showed significant hypometabolism in the putamen, pons and cerebellum compared to the normal controls and IPD groups. In addition, PSP showed significant hypometabolism in the caudate nucleus, the thalamus, midbrain and the cingulate gyrus compared to the normal controls, the IPD and the MSA groups. In conclusion, an assessment of the 18F-FDG PET images using SPM may be a useful adjunct to a clinical examination when making a differential diagnosis of Parkinsonism. PMID- 15294330 TI - Driven equilibrium (drive) MR imaging of the cranial nerves V-VIII: comparison with the T2-weighted 3D TSE sequence. AB - PURPOSE: The aim of this study is to evaluate the efficacy of the driven equilibrium radio frequency reset pulse (DRIVE) on image quality and nerve detection when used in adjunction with T2-weighted 3D turbo spin-echo (TSE) sequence. MATERIALS AND METHODS: Forty-five patients with cranial nerve symptoms referable to the cerebellopontine angle (CPA) were examined using a T2-weighted 3D TSE pulse sequence with and without DRIVE. MR imaging was performed on a 1.5-T MRI scanner. In addition to the axial resource images, reformatted oblique sagittal, oblique coronal and maximum intensity projection (MIP) images of the inner ear were evaluated. The nerve identification and image quality were graded for the cranial nerves V-VIII as well as inner ear structures. These structures were chosen because fluid-solid interfaces existed due to the CSF around (the cranial nerves V-VIII) or the endolymph within (the inner ear structures). Statistical analysis was performed using the Wilcoxon test. P < 0.05 was considered significant. RESULTS: The addition of the DRIVE pulse shortens the scan time by 25%. T2-weighted 3D TSE sequence with DRIVE performed slightly better than the T2-weighted 3D TSE sequence without DRIVE in identifying the individual nerves. The image quality was also slightly better with DRIVE. CONCLUSION: The addition of the DRIVE pulse to the T2-weighted 3D TSE sequence is preferable when imaging the cranial nerves surrounded by the CSF, or fluid-filled structures because of shorter scan time and better image quality due to reduced flow artifacts. PMID- 15294331 TI - Classification of the inferior turbinate bones: a computed tomography study. AB - BACKGROUND: There are only few reports describing the texture of the inferior turbinate bone in normal and pathologic conditions. In this study, different types of human inferior turbinate bones were classified and radiological features of each type were defined. METHODS: The shape, structure and density of the inferior turbinate bones were evaluated using paranasal sinus computed tomography images of 283 patients. The cross-sectional areas of the bony part of the inferior turbinate were measured in bone windows. RESULTS: Human inferior turbinate bones were classified into four groups on the basis of different shape and structure as: Type I, lamellar; Type II, compact; Type III, combined type (compact with spongious component); Type IV, bullous. The distribution was as follows: 352 (62.19%) lamellar, 50 (8.83%) compact, 162 (28.63%) combined, and 2(0.35%) bullous type. CONCLUSION: Inferior turbinate bone is not in a uniform shape and structure. These diversities should be taken into consideration in radiological and clinical evaluation. PMID- 15294332 TI - The diagnostic relevance of colour Doppler artefacts in carotid artery examinations. AB - PURPOSE: Physical and technical artefacts of the colour Doppler method are examined with regard to their diagnostic relevance for the carotid artery. MATERIALS AND METHODS: After recording all diagnostic problems arising from physical and technical artefacts in 30,000 consecutively carotid arteries, the quantitative significance of relevant artefacts was determined in minor subgroups of the collective. RESULTS: Acoustic shadowing causes diagnostic problems in morphological and haemodynamic evaluation of stenoses. Mirror image artefacts simulate flow in cases of actual vessel occlusion or mimic vessel wall ulceration in carotid plaques and stenoses. Insonation angle artefacts inhibit detection of flow or mimic flow reversal. Problems of spatial resolution lead to incorrect demonstration of the vessel lumen in stenotic findings. Aliasing, perivascular colour artefacts, and ghosting do not cause any diagnostic problems. Relevant shadowing artefacts occurred in 14.7%, and relevant mirror image artefacts in 2.5% of the pathological cases. Insonation angle artefacts occurred in 17.3% of the cases (when using standard apparatus setting). However, with an exact knowledge of the artefact phenomena, insonation angle artefacts could be eliminated and mirror image artefacts were recognized in all cases. Resolution artefacts resulted in underestimation of carotid stenoses by on average 13.3% of the degree of stenosis. CONCLUSION: In principle, four artefact phenomena give rise to diagnostic problems; however, with a good knowledge of the phenomena, only artefacts due to acoustic shadowing and limited spatial resolution are of diagnostic significance. PMID- 15294333 TI - Assessment of MRI and dynamic contrast-enhanced MRI in the differential diagnosis of adenomatoid odontogenic tumor. AB - The radiographical differentiation of adenomatoid odontogenic tumor (AOT) from dentigerous cysts, calcifying odontogenic cysts, calcifying epithelial odontogenic tumors, odontogenic keratocysts and amelobastomas is sometimes difficult. We attempted to differentiate AOT from other lesions similar to AOT in radiographic findings using MRI. The MRI features of AOT in our three cases included homogeneous low SI in the cystic portion and homogeneous intermediate SI in the solid portion on T1WI, homogeneous high SI in the cystic portion and intermediate to slightly high SI in the solid portion on T2WI and enhancement of only the solid portion on CE-T1WI although none of the sequences included SI of calcifications. The contrast index curves in the three cases of AOT showed a gradual increase to 300 s, which signified a benign tumor. These MRI features were characteristic features of AOT and might be a basis for differentiating AOT from the above possible lesions in radiographic examinations. PMID- 15294334 TI - Dental CT and orthodontic implants: imaging technique and assessment of available bone volume in the hard palate. AB - PURPOSE: Palatal implants (PI) have been introduced for orthodontic treatment of dental and skeletal dysgnathia. Due to the restricted amount of bone in this region, precise preoperative anatomic information is necessary. The aim of this study was to determine whether dental CT could serve as a tool to locate the optimal size and position for orthodontic implant placement. MATERIALS AND METHODS: In 32 patients, where palatal implant placement was planned, axial CT scans of the maxillary bone were acquired. Using a standard dental software package (Easy Vision dental software package 2.1, Philips; Best, The Netherlands), paracoronal views were reconstructed and measurements of palatal bone height in 3 mm increments, dorsally from the incisive canal, were performed in the median and both paramedian regions. RESULTS: The overall mean bone height was 5.01 mm (S.D. 2.60), ranging from 0 to 16.9 mm. The maximum palatal bone height was 6.17 mm (S.D. 2.81) at 6 mm dorsally from the incisive canal. Due to the lack of adequate bone (less than 4 mm), implant placement was not performed in 3 cases (7%). In the remaining 39 cases (93.0%), primary implant stability was achieved and complications, such as perforation of the palate, could be avoided. CONCLUSION: The results demonstrate that dental CT promises to be a valuable tool in evaluating the potential and optimal size and site for orthodontic implant placement. PMID- 15294335 TI - Rotator cuff tears in asymptomatic individuals: a clinical and ultrasonographic screening study. AB - OBJECTIVE: To determine the prevalence and clinical impact of rotator cuff tears in asymptomatic volunteers. MATERIALS AND METHODS: Sonographic examinations of the shoulder of 212 asymptomatic individuals between 18 and 85 years old were performed by a single experienced operator. The prevalence and location of complete rotator cuff tears were evaluated. The clinical assessment was based on the Constant Score. Magnetic resonance imaging (MRI) of the shoulder was obtained in those patients where US showed rotator cuff pathology. RESULTS: Ultrasound showed a complete rupture of the supraspinatus tendon in 6% of 212 patients from 56 to 83 years of age (mean: 67 years). MRI confirmed a complete rupture of the supraspinatus tendon in 90%. All patients reported no functional deficits, although strength was significantly lower in the patient group with complete supraspinatus tendon tear (P < 0.01). CONCLUSION: There is a higher prevalence in older individuals of rotator cuff tendon tears that cause no pain or decrease in activities of daily living. PMID- 15294336 TI - Relationship of condylar position to disc position and morphology. AB - INTRODUCTION/OBJECTIVE: The purpose of this study was to assess whether condylar position, as depicted by magnetic resonance imaging, was an indicator of disc morphology and position. METHODS AND MATERIAL: One hundred and twenty two TMJs of 61 patients with temporomandibular joint disorder were examined. Condylar position, disc deformity and degree of anterior disc displacement were evaluated by using magnetic resonance imaging. RESULTS AND DISCUSSION: Posterior condyle position was found to be the main feature of temporomandibular joints with slight and moderate anterior disc displacement. No statistical significance was found between the condylar position, and reducing and nonreducing disc positions. On the other hand, superior disc position was found to be statistically significant for centric condylar position. CONCLUSION: It was concluded that posterior condyle position could indicate anterior disc displacement whereas there was no relation between the position of condyle and the disc deformity. PMID- 15294337 TI - Sensitivity and specificity of thallium-201 scintigraphy for the diagnosis of malignant vertebral fractures. AB - OBJECTIVE: To evaluate the accuracy of thallium-201 (201TI) scintigraphy in distinguishing a benign from a malignant recent non-traumatic vertebral fracture. METHODS: STUDY DESIGN--Single center, prospective study. PARTICIPANTS--Patients hospitalized for a recent non-traumatic vertebral fracture. EVALUATION--Usual clinical, laboratory and radiological assessment; 201TI vertebral scintigraphy: patients were injected with iv 3 mCi 201TI. Early and delayed images of the fractured vertebra were obtained. DATA ANALYSIS--(1) Two examinators, unaware of the other findings, rated the images as hyperfixation or not of the fractured vertebra; (2) the ratio (average count per pixel of the fractured vertebra/normal adjacent vertebrae) were calculated. The FINAL DIAGNOSIS was established on the result of vertebral biopsy or on follow-up. RESULTS: Twenty-one patients were included. The final diagnosis was a benign vertebral fracture in 14 patients and a malignant vertebral fracture in 7. The sensitivity, specificity, positive and negative predictive values for a malignant fracture on early 201TI vertebral scintigraphy images were 28.6, 92.9, 66.6, and 72.2%, respectively, and on delayed images were 28.6, 100, 100, and 73.7%, respectively. The ratio of lesioned over normal tissue was not increased in malignant, compared with benign fractures. CONCLUSION: The weak sensitivity does not support the wide use of 201TI bone scintigraphy to distinguish a benign from a malignant recent non traumatic vertebral fracture. However, the high specificity suggests that such evaluation might be proposed prior to vertebral biopsy in some difficult cases. PMID- 15294338 TI - Foci of decreased signal on T2-weighted MR images in leiomyosarcomas of soft tissue: correlation between MR and histological findings. AB - OBJECTIVE: Leiomyosarcomas are rare soft tissue sarcomas with varying MR signal characteristics and histologic pictures. The purpose of this study was to investigate the histological features of foci, which showed decreased signal on T2-weighted images in leiomyosarcomas of soft tissue. MATERIAL AND METHODS: We reviewed the MR images of six histologically proved cases of leiomyosarcomas of soft tissue and correlated the foci, which showed decreased signal on T2-weighted images with the histologic findings. RESULT: Microscopic examination revealed that these foci were composed of hyalinization of neoplastic tissue, internal septations, deposition of hemosiderin, or corresponded to metaplastic bone. CONCLUSION: The authors explain that the foci of decreased signal on T2-weighted MR images correspond to tissue components of the lesion, particularly fibrous tissue, hemosiderin and metaplastic bone. So, the suggestion is that leiomyosarcoma should be considered in the differential diagnosis of soft-tissue tumors that show foci of decreased signal on T2-weighted MR images. PMID- 15294339 TI - Patient data security in the DICOM standard. AB - The DICOM committee added the section "Security Profiles" to the DICOM standard, in order to provide the opportunity of safe communication between health care system partners. Data complying with the DICOM standard--e.g. pictures, signals or reports of examinations can be provided with one or more digital signatures. Attention should be paid to the fact that these possibilities of the DICOM standard are available or can be supplied subsequently by new acquisitions of radiological modalities. The required information to check these prerequisites are given. PMID- 15294340 TI - Hemispheric dominance for the cortical control of swallowing in humans: a contribution to better understand cortical organization? PMID- 15294341 TI - Altered expression of genes related to zinc homeostasis in early mouse embryos exposed to di-2-ethylhexyl phthalate. AB - Numerous studies have shown that di-2-ethylhexyl phthalate (DEHP) is teratogenic in animals but the mechanism of developmental toxicity is not well understood. One hypothesis is altered zinc homeostasis. The present study has investigated the effect of DEHP exposure on several key genes in zinc metabolism (MT-I, MT-II, ZnT-1) for early mouse embryos exposed in utero. Time- and dose-dependent effects were examined using expression polymerase chain reaction (PCR) (relative to ACTB) and Western blot analysis of the maternal liver, embryonic brain, and visceral yolk sac at 9 days post-coitus (d.p.c.). Maternal exposure to 800 mg/kg DEHP increased the abundance of MT-I and MT-II transcripts in maternal liver at 3.0, 4.5 and 6.0 h after administration. MT-I and MT-II protein induction was confirmed by Western blot analysis. On the other hand, this exposure down regulated both transcripts (MT-I, MT-II), as well as transcripts for a zinc transporter (ZnT-1), in the embryonic brain, but not the visceral yolk sac. To examine dose-response relationships, the experiment was repeated for DEHP exposures of 50, 200 and 800 mg/kg. The effect to MT-I and MT-II expression in the maternal liver became significant at the 200 mg/kg dose level. The contrasting effect to MT-I, MT-II and ZnT-1 expression in the embryo was also dose-dependent, and a benchmark computation for the dose resulting in a 5% change in the mean (BMD5) was estimated as 11.6 mg/kg for MT-I, 8.9 mg/kg for MT-II, and 6.6 mg/kg for ZnT-1. We conclude that DEHP exposure to pregnant dams at reasonably low levels during organogenesis stages can alter the expression of several key genes in embryonic zinc homeostasis. PMID- 15294342 TI - CpG methylation of the mouse CYP1A2 promoter. AB - Cytochrome P450 1A2 (CYP1A2) is a xenobiotic metabolizing enzyme that is tissue specifically regulated in the mammalian liver by arylhydrocarbon receptor (AhR) dependent and -independent pathways. In this study, CpG methylation of the CYP1A2 promoter was analyzed in mouse tissues and liver-derived cells. Compared to lung and kidney, the CYP1A2 promoter is undermethylated in the liver in a promoter domain-specific manner. The CYP1A2 promoter showed a similar methylation pattern in wild-type and AhR-null liver. At birth, the promoter was hypermethylated and CYP1A2 was negligibly expressed in the liver. However, CYP1A2 expression increased following birth, coincident with the demethylation of the promoter. In hepatoma Hepa1c1c7 cells not expressing CYP1A2, the promoter was hypermethylated at specific CpG sites. In isolated hepatocytes, CYP1A2 expression declined over time and the degree of CYP1A2 methylation increased, albeit only after a delay. Exposure to 5-aza-2'-deoxycytidine did not induce CYP1A2 in Hepa1c1c7 cells and hepatocytes. Taken together, our findings suggest that CpG methylation is involved in the tissue-specific and developmental regulation of CYP1A2, but the de novo methylation of the CYP1A2 promoter is induced by the silent state of the gene rather than causing it. PMID- 15294343 TI - Dietary iron regulates intestinal cadmium absorption through iron transporters in rats. AB - The intestinal absorption of cadmium (Cd) is influenced by body iron (Fe) status in laboratory animals and humans. In this study we investigated the role of the apical Fe transporter divalent metal transporter 1 (DMT1) and the basolateral Fe exporter metal transporter protein 1 (MTP1) in Cd absorption. Rats were divided into the following groups; an Fe-sufficient (FeS) control group that was fed an FeS diet for 4 weeks (FeS, 4 weeks); an Fe-deficient (FeD) group that was fed an FeD diet for 4 weeks (FeD, 4 weeks); an FeS control group that was fed an FeS diet for 8 weeks (FeS, 8 weeks); an FeD/FeS group that was fed an FeD diet for 4 weeks and then an FeS diet for the following 4 weeks (FeD/FeS, 4 weeks/4 weeks); and an FeD group that was fed an FeD diet for 8 weeks (FeD, 8 weeks). After the 4 and 8-week feeding periods, rats were given a single oral gavage of Cd and were sacrificed 24 h later. The FeD (4 weeks) group developed Fe deficient anemia, but the parameters returned to control levels in the FeD/FeS group (4 weeks/4 weeks). The Cd body burden was greater in FeD (4 weeks) rats compared to FeS control (4 weeks), but returned to control Cd levels in FeD/FeS (4 weeks/4 weeks) rats. In addition, the expression of DMT1 and MTP1 was induced by Fe deficiency in the duodenum of FeD (4 weeks) rats, but was down-regulated to control values in FeD/FeS (4 weeks/4 weeks) rats. The correlation between duodenal DMT1 and MTP1 expression and Cd body burden in rats suggests an important role of DMT1 and MTP1 in Cd absorption. PMID- 15294344 TI - Similarity and difference in the acute lung injury induced by a radiographic contrast medium and an anticancer agent paclitaxel in rats. AB - Paclitaxel is one of the most frequently used anticancer agents but its use is sometimes limited because of the incidence of severe hypersensitivity reactions. The clinical symptoms of the reactions, including dyspnea and pulmonary edema, are similar to those induced by iodinated contrast medium during radiographic examination. Therefore, the premedication for the prophylaxis of hypersensitivity reactions to paclitaxel is carried out in accordance with that for radiographic contrast medium. In the present study, we compared the effects of paclitaxel and an iodinated radiocontrast medium ioxaglate on vascular permeability and pulmonary function in rats. Both paclitaxel (15 mg/kg) and ioxaglate (4 g iodine/kg) caused perivascular edema, plasma extravasation and decrease in arterial PaO2. Dexamethasone inhibited plasma extravasation induced by the two compounds. In contrast, histamine H1 and H2 antagonists attenuated the effects of ioxaglate without inhibiting those of paclitaxel. On the other hand, a neurokinin NK1 antagonist (LY303870: 0.5 mg/kg) significantly inhibited the pulmonary responses induced by paclitaxel but not by ioxaglate. Therefore, it is suggested that paclitaxel and ioxaglate cause similar acute lung injury but the mechanisms are different between the two compounds, in which histamine and substance P are involved in the pulmonary dysfunction induced by ioxaglate and paclitaxel, respectively. These findings also raise a possibility that more effective premedication is required for the prophylaxis of paclitaxel hypersensitivity. PMID- 15294345 TI - Neurotoxic effects of trimethyltin and triethyltin on human fetal neuron and astrocyte cultures: a comparative study with rat neuronal cultures and human cell lines. AB - Trimethyltin (TMT) and triethyltin (TET) caused cell death in cultures of primary human neurons and astrocytes, rat neurons and human neuroblastoma cell lines. Human neurons and astrocytes showed a delayed response to TMT cytotoxicity. After 24h of TMT exposure, LC50 values were 148.1, 335.5 and 609.7 microM for SK-N-MC neuroblastoma cell line, neurons and astrocytes, respectively. Over 5 days of exposure, the cytotoxic potency of TMT increased about 70-fold in human cortical neurons. Rat hippocampal neurons were the most vulnerable cells to TMT cytotoxicity, exhibiting an LC50 value 30-fold lower (1.4 microM) than that of rat cerebellar granule cells (44.28 microM). With the exception of rat hippocampal neurons, TET was more potent than TMT in inducing cell death (LC50 values of 3.5-16.9 microM). Moreover, TET was more effective than TMT in increasing intracellular free Ca2+ concentration in human and rat neurons. This work shows that human fetal neuron and astrocyte cultures are a useful model for studying the neurotoxic effects of these environmental contaminants and, thus, predicting their impact on human health. PMID- 15294346 TI - Production of macrophage IL-1beta was inhibited both at the levels of transcription and maturation by caspase-1 following inhalation exposure to isobutyl nitrite. AB - Epidemiological studies have identified abuse of nitrite inhalants as an independent co-factor in HIV infection and in Kaposi's sarcoma (KS) in AIDS patients. In the present study we investigated the ability of macrophages from mice exposed to isobutyl nitrite to produce the inflammatory cytokine IL-1beta, upon stimulation with IFN-gamma and LPS. The production of IL-1beta was inhibited up to 55%. IL-1beta mRNA transcription was reduced by 35% following nitrite inhalant exposure, consistent with inhibition of activation-induced phosphorylation of macrophage mitogen-activated protein kinase p38. However, synthesis of the 31 kDa IL-1beta precursor protein was only marginally inhibited. Caspase-1, which cleaves the precursor IL-1beta into mature 17 kDa IL-1beta, was examined. Nitrite inhalant exposure blocked activation-induced increases in caspase-1 activity, consistent with a 50% reduction in 17 kDa IL-1beta shown in Western blots. Thus, exposure to nitrite inhalants reduced macrophage production of IL-1beta by reducing transcription, as well as post-translational processing mediated by caspase-1. PMID- 15294347 TI - The relation of individual cadmium concentration in urine with total cadmium intake in Kakehashi River basin, Japan. AB - The aim of the present study was to clarify the relationship between urinary cadmium concentration (U-Cd) and total cadmium intake (T-Cd) based on individual data of Cd in urine. Morning urine specimens were collected from 198 persons (100 men with average age of 45.8 years and 98 women with average age of 50.7 years) living in the Kakehashi River basin in Ishikawa Prefecture, Japan. The method used to calculate T-Cd basically mirrored that used in our previous study. Correlation coefficients between U-Cd and T-Cd of 0.879 and 0.835 were obtained in men and women, respectively, with both values statistically significant. Therefore, T-Cd is confirmed to be a useful indicator for external Cd exposure even on the basis of individual data of U-Cd. PMID- 15294348 TI - Locomotor hyperactivity following prenatal exposure to 5-bromo-2'-deoxyuridine: neurochemical and behavioral evidence of dopaminergic and serotonergic alterations. AB - Prenatal exposure to 5-bromo-2'-deoxyuridine (BrdU) has been reported to induce abnormal behaviors in offspring, including marked hyperactivity. In this study, the contribution of the serotonin (5-HT) and dopamine (DA) systems to BrdU induced developmental neurotoxicity was investigated. Sprague-Dawley rats were treated with BrdU on gestational days 9 through 15 (50mg/kg, i.p.) and male offspring (BrdU-rats) were examined. The BrdU-rats exhibited a 3.5-fold increase in locomotor activity. The dopamine D2 receptor antagonist sulpiride increased locomotor activity in the BrdU-rats, but decreased it in control rats. The BrdU rats responded to the 5-HT1A receptor antagonist NAN190 much more than the controls. The measurement of monoamines revealed significant decreases in DA, dihydroxyphenylacetic acid, and homovanilic acid, and significant increases in 5 HT and 5-hydroxy-3-indolacetic acid, with a decrease in the 5-HT turnover ratio in the striatum of BrdU-rats. Thus, prenatal exposure to BrdU induced alterations in both the DA and 5-HT systems. PMID- 15294349 TI - Phenol and catechol induce prehemolytic and hemolytic changes in human erythrocytes. AB - The toxic potency of two industrially used compounds (phenol and catechol) was studied in human blood cells in vitro. Catechol was found to be a more harmful toxin than phenol, since it provokes statistically significant changes in the function of erythrocytes even at low doses. Most of the changes was statistically significant for the doses of 50 ppm of catechol and 250 ppm of phenol. Both compounds induced methaemoglobin formation, glutathione depletion and conversion of oxyhaemoglobin to methaemoglobin, which is associated with superoxide anion production and lead to formation of ferryl hemoglobin, hydrogen peroxide or hydroxyl radicals. It is known that oxidation of catechol leads to formation of semiquinone radicals. Semiquinones are able to bind to nucleophilic residues like -SH or -NH2 of proteins and these macromolecules may undergo inactivation. We observed among especially susceptible to action of catechol are catalase (CAT) (100 ppm) and superoxide dismutase (SOD) (250 ppm). Decrease of the activity of catalase and SOD by catechol induced radical species formation. This lead to inhibition of another protective enzymes such as glutathione-S-transferase (500 ppm), glutathione reductase (1000 ppm), glucose-6-phosphate dehydrogenase activity (1000 ppm). Cytotoxicity of phenol or catechol was noted as hemolysis. Haemoglobin liberated from erythrocytes in this process may further generate oxygen free radicals and subsequently initiate enzymes damage. It seems to be essential that in phenol and catechol toxicity special role play damages of heme proteins and other proteins molecule, and damages of lipids are not so important. PMID- 15294350 TI - Protective activity of cedron (Aloysia triphylla) infusion over genetic damage induced by cisplatin evaluated by the comet assay technique. AB - Using the comet assay technique, this paper examines the protection from the cisplatin-induced genetic damage in mouse bone marrow cells provided by cedron leaf infusion. Animals were separated into six groups: (I) untreated, (II) negative control, (III) treated with cedron-leaf infusion (5%), (IV) treated with cisplatin (6 mg/kg b.w.), (V) pretreated with infusion and treated with cisplatin and (VI) positive control (cyclophosphamide, 20 mg/kg b.w.). Based on the tail moment values found, four types of comets were distinguished. No statistical differences (P<0.01) were found between untreated animals, negative control and infusion treated mice. As expected, treatment of mice with a single dose of cis DDP-induced genetic damage and the pretreatment with infusion prior to cis-DDP injection inhibited the capacity of cisplatin to induce genetic damage. Cell viability was up to 90% in all cases. The results suggest that infusion could exert its in vivo antigenotoxic action by enhancing the antioxidant status of bone marrow cells. The found could be attributed to its scavenging potency towards free radicals. PMID- 15294351 TI - Suppression of aromatase activity in vitro by PCBs 28 and 105 and Aroclor 1221. AB - The effects of polychlorinated biphenyls (PCBs) on human cytochrome P450 aromatase activity in vitro were investigated using a commercially available microsomal fraction obtained from baculovirus infected insects that had been transfected with the human CYP19 gene and cytochrome P450 reductase. The assay measured the conversion of tritiated testosterone to estradiol in Tris buffer at pH 7.4. When aroclors, commercial preparations of PCBs, were added to aromatase assays at a 10 microM concentration, Aroclor 1221 caused a reduction in the aromatase activity, whereas other aroclors (1016, 1232, 1242, 1248, 1254, 1260, 5432, 5442 and 5460) were without effect. Further investigation of the effect of Aroclor 1221 on aromatase activity showed that the inhibition was dose dependent. When a reconstituted mixture (RM) of PCBs that represented the congeneric content of human milk was investigated, no inhibition of aromatase activity at the maximum treatment of 15.0 microM was observed. None of the congeners present in the reconstituted mixture, except PCB 28 and 105, affected P450 arom activity. PCB 28 showed a statistically significant inhibition of aromatase activity (P<0.05) at 1.5 and 15 microM and a significant inhibition of aromatase activity by PCB 105 was also observed, but only at 15 microM. In three separate kinetic analyses the Km(app) for aromatase was 64, 89 and 69 nM (mean 74 nM). In addition, PCB 28 resulted in an increase in the Km(app) without a significant effect on Vmax(app), suggesting competitive inhibition by this congener. This conclusion was supported by slope (Km(app)/Vmax(app) versus [inhibitor]) and intercept (1/Vmax(app) versus [inhibitor]) replots. The slope replots gave Ki(app) values for PCB 28 of 0.9, 1.3 and 2.0 microM (mean 1.4 microM), whereas intercept replots were almost horizontal. Thus, PCB 28 is a competitive inhibitor of aromatase with a Ki(app) value approximately 20-fold the Km(app) value. Based on these studies, we conclude that most PCBs are not inhibitors of aromatase activity in vitro. However, as being inhibitors of aromatase activity, Aroclor 1221, PCB 28 and PCB 105 would remain a priority for further study as possible endocrine disrupters. PMID- 15294352 TI - Removal of copper from aqueous solution by Ascophyllum nodosum immobilised in hydrophilic polyurethane foam. AB - The seaweed Ascophyllum nodosum was pre-treated by successive washes in distilled water and dilute acid, dried, and pulverised to produce particles of <150 microm. These were immobilised during the manufacturing process of Hypol 2002 polymer to form a biomass/polymer matrix that was stable and easy to handle. In making the composite a mixing speed of 360 rpm for 20-30 s with 2% (w/w) addition of surfactant to pre-polymer was found to be ideal. The average pore sizes for different water polymer mixes (expressed as volume ratios) were 1.66 mm +/- 0.98 (ratio 0.75:1), 1.58 mm +/- 0.76 (ratio 1:1), 1.64 mm +/- 0.6 (ratio 1.5:1) and 1.11 mm +/- 0.615 (ratio 2:1). The biomass/polymer was used alongside free native biomass in an initial adsorption experiment using a 0.0315 mmol dm(-3) Cu (II) solution and gave a copper uptake capacity (q)(max) of 0.037 mmol Cu g(-1) dry weight seaweed in both cases which represented approximately 85% of total initially available copper. In later adsorption isotherm experiments using Cu concentrations between 0.0315 and 0.944 mmol dm(-3) at pH 5.0 and immobilized biomass over five consecutive adsorption/desorption cycles the biomass/polymer showed an initial lowering of adsorption capacity but stabilised at 0.23 mmol g( 1) dry weight by the third re-use. The q(max) of the immobilised biomass decreased from 0.55 and 0.416 mmol of Cu g(-1) dry weight when pH was lowered from 4.0 to 3.0, and increased from 0.576 mmol g(-1) dry weight (biomass) at 283 K to 0.636 mmol g(-1)(biomass) at 303 K. PMID- 15294353 TI - The environmental history of Chatthin Wildlife Sanctuary, a protected area in Myanmar (Burma). AB - We reconstructed the history of Chatthin Wildlife Sanctuary (CWS) to understand how social and economic events, and policy changes affected the sanctuary's condition. We surveyed 25 villages surrounding CWS to evaluate past and present ecological conditions, compare the results with historical accounts and identify causal relationships. During the first half of the 20th century, the primary threat was the government's reduction of old growth forest to supply fuel wood for the British-built railway. The railroad opened the area to colonization, but the villagers' impact on timber and wildlife was low. From 1945 to 1988, villagers became the primary force of landscape degradation. The post-war windfall of firearms increased hunting pressure, and populations of large mammal started to decline. With the economic decline of the 1970s and 1980s, the community's demand for game and forest products intensified, and the large mammal fauna was reduced from eleven to four species. From 1988-2003, the forests surrounding the sanctuary were fragmented and degraded. The absence of large predators rendered the park safe for livestock, and the combined effects of grazing and removal of forest products seriously degraded habitat within CWS. Major threats to CWS during the past two decades have resulted from land use decisions in which government-planned economic enterprises caused encroachment by villagers. Stabilization and recovery of this sanctuary will require management compatible with human needs, including expanded buffer zones, better core area protection, community forestry projects, and probably relocation of villages within the park. PMID- 15294354 TI - Spatial assessment of the economic feasibility of short rotation coppice on radioactively contaminated land in Belarus, Ukraine, and Russia. I. Model description and scenario analysis. AB - The economic feasibility of short rotation coppice (SRC) production and energy conversion in areas contaminated by Chernobyl-derived (137)Cs was evaluated taking the spatial variability of environmental conditions into account. Two sequential GIS-embedded submodels were developed for a spatial assessment, which allow for spatial variation in soil contamination, soil type, and land use. These models were applied for four SRC production and four energy conversion scenarios for the entire contaminated area of Ukraine, Belarus, and Russia and for a part of the Bragin district, Belarus. It was concluded that in general medium-scale SRC production using local machines is most profitable. The areas near Chernobyl are not suitable for SRC production since the contamination levels in SRC wood exceed the intervention limit. Large scale SRC production is not profitable in areas where dry and sandy soils predominate. If the soil contamination does not exceed the intervention limit and sufficient SRC wood is available, all energy conversion scenarios are profitable. PMID- 15294355 TI - Spatial assessment of the economic feasibility of short rotation coppice on radioactively contaminated land in Belarus, Ukraine, and Russia. II. Monte Carlo analysis. AB - A Monte Carlo analysis of two sequential GIS-embedded submodels, which evaluate the economic feasibility of short rotation coppice (SRC) production and energy conversion in areas contaminated by Chernobyl-derived (137)Cs, was performed to allow for variability of environmental conditions that was not contained in the spatial model inputs. The results from this analysis were compared to the results from the deterministic model presented in part I of this paper. It was concluded that, although the variability in the model results due to within-gridcell variability of the model inputs was considerable, the prediction of the areas where SRC and energy conversion is potentially profitable was robust. If the additional variability in the model input that is not contained in the input maps is also taken into account, the SRC production and energy conversion appears to be potentially profitable at more locations for both the small scale and large scale production scenarios than the model predicted using the deterministic model. PMID- 15294356 TI - Influence of organic loading on an anaerobic sequencing biofilm batch reactor (ASBBR) as a function of cycle period and wastewater concentration. AB - The effect of organic loading on the performance of a mechanically stirred anaerobic sequencing biofilm batch reactor (ASBBR) has been investigated, by varying influent concentration and cycle period. For microbial immobilization 1 cm polyurethane foam cubes were used. An agitation rate of 500 rpm and temperature of 30+/-2 degrees C were employed. Organic loading rates (OLR) of 1.5 6.0gCODl(-1)d(-1) were applied to the 6.3-l reactor treating 2.0 l synthetic wastewater in 8 and 12-h batches and at concentrations of 500-2000mgCODl(-1), making it possible to analyze the effect of these two operation variables for the same organic loading range. Microbial immobilization on inert support maintained approximately 60 gTVS in the reactor. Filtered sample organic COD removal efficiencies ranged from 73 to 88% for organic loading up to 5.4gCODl(-1)d(-1). For higher organic loading (influent concentration of 2000mgCODl(-1) and 8-h cycle) the system presented total volatile acids accumulation, which reduced organics removal efficiency down to 55%. In this way, ASBBR with immobilized biomass was shown to be efficient for organic removal at organic loading rates of up to 5.4gCODl(-1)d(-1) and to be more stable to organic loading variations for 12-h cycles. This reactor might be an alternative to intermittent systems as it possesses greater operational flexibility. It might also be an alternative to batch systems suspended with microorganisms since it eliminates both the uncertainties regarding granulation and the time necessary for biomass sedimentation, hence reducing the total cycle period. PMID- 15294357 TI - The effect of Oja-titun market effluent on the chemical quality of receiving OPA reservoir in Ile-Ife, Nigeria. AB - The effect of market effluent from the Oja-titun market in Ile-Ife, Nigeria on the chemical quality of the Opa Reservoir located 3.5 km downstream was investigated between February and November 2000. Water samples were collected in February, May, August and November from 16 sites, four along each of the market drainage channels (MDC), market stream, tributary stream and the Opa River and Reservoir. The peak level of each variable-biochemical oxygen demand, temperature, total alkalinity, Na(+), K(+), Ca(2+), Mg(2+), PO(4)(3+), SO(4)(2+), Cl(-), NO(3)(-), Pb and Zn-occurred at the MDC, and decreased significantly downstream, except pH, conductivity and dissolved oxygen, which increased. Seasonal fluctuation in most variables was pronounced. Generally, there were high values in the early dry and dry seasons and low values in the rainy and early rainy seasons. Comparison of the reservoir water with international limitation standards for drinking water supply showed that the quality of the reservoir water was very low and that treatment required to achieve minimum limitation standards for drinking water would be both intensive and expensive. The study concluded that the stream borne effluent from the market impacts significantly on the chemical quality of the reservoir water although other tributaries within the Reservoir's catchment are other possible sources of pollutants in the reservoir. PMID- 15294358 TI - Respective influence of habitat conditions and management regimes on prealpine calcareous grasslands. AB - The calcareous grasslands of the south-western French Alps have been poorly studied, although they provide suitable habitat for rare plant species and communities. The separate and combined effects on calcareous grassland communities of habitat conditions (lithology, soil moisture) and management regimes (grazing intensity, cutting regime) were studied using constrained ordination techniques (canonical correspondence analysis with variance partitioning). Among the explanatory variables considered, the most important factor determining floristic composition was lithology, which explained 11.9% of floristic variability, followed by grazing intensity (6.0%). Additive effects of management and lithology explained 23.9% of floristic variability. Species niche amplitude was measured by conditional variances of samples along main ordination axes, in order to define adequate conservation management for the rarest short lived species with narrow niche breadth on both habitat and management gradients. PMID- 15294360 TI - Risk factors for low birth weight: a review. AB - Low birth weight (LBW) is one of the main predictors of infant mortality. The global incidence of LBW is around 17%, although estimates vary from 19% in the developing countries (countries where it is an important public health problem) to 5-7% in the developed countries. The incidence in Spain in the decade 1980 1989 was about 5.7%. LBW is generally associated with situations in which uterine malnutrition is produced due to alterations in placental circulation. There are many known risk factors, the most important of which are socio-economic factors, medical risks before or during gestation and maternal lifestyles. However, although interventions exist to prevent many of these factors before and during pregnancy, the incidence of LBW has not decreased. PMID- 15294361 TI - Medical management for termination of second and third trimester pregnancies: a comparison of strategies. AB - OBJECTIVE: Misoprostol and sulprostone are prostaglandins that can be used for the termination of second and third trimester pregnancy. The aim of the present study was to compare the effectiveness of both agents for the termination of second and third trimester pregnancy in cases of congenital or genetic abnormalities, and for the induction of labour in cases of intra-uterine foetal death. STUDY DESIGN: We collected data from all women who had been treated with misoprostol in the second or third trimester of pregnancy between January 2001 and July 2002 in cases of congenital or genetic abnormalities, and for the induction of labour in cases of intra-uterine foetal death. In cases where the foetus was alive, misoprostol was usually (77%) combined with mifepristone. Women were matched to women who had been treated with sulprostone for termination of second and third trimester pregnancy before 2001. We matched for hospital, previous vaginal delivery, intra-uterine death and duration of pregnancy. The primary outcome measure was time to delivery. RESULTS: Since the treatment effect was different in patients in whom labour was induced for intra-uterine death and patients in whom labour was induced while the foetus was alive, the analysis was stratified for this parameter. In 94 patients with intra-uterine death, there was no significant difference in time to delivery, blood loss, operative removal of the placenta and need for pain relief between misoprostol and sulprostone. In vital pregnancy (n = 96), time to delivery was significantly shorter in the misoprostol group. The relative risk for haemorrhage exceeding 1000 ml in this group was 0.40 (95% confidence interval, CI, 0.13-1.2). We observed no significant differences with respect to operative removal of the placenta or need for pain relief. CONCLUSION: In cases of intra-uterine death, the effectiveness of misoprostol for termination of pregnancy is comparable to that of sulprostone. In vital pregnancy, combination of mifepristone and misoprostol is more effective than sulprostone alone. PMID- 15294362 TI - The quality of intrapartum fetal heart rate monitoring. AB - OBJECTIVE: To determine the quality of fetal heart rate (FHR) recordings during the first and second stage of labor by quantifying the amount of fetal signal loss in relation to the method of monitoring: external ultrasound or directly via a scalp electrode. STUDY DESIGN: Analysis of 239 intrapartum recordings stored between 1 January 2001 and 1 July 2001 from consecutive deliveries at the Vrije Universiteit Medical Center in Amsterdam. Singletons delivered via the vaginal route were included in the study. FHR recordings had duration of at least 1h prior to birth of the infant. Subdivision in three groups took place on the basis of the recording technique which had been used; i.e. ultrasound, scalp electrode or a combination of both methods. FHR data was obtained using HP-M1350 cardiotocographs. The status (pen on, pen off, maternal signal) and the mode of the signals were acquired. The duration of pen lifts and maternal signals was divided by the total duration of the recording. Statistical analyses were performed with the Mann-Whitney U-test and the Wilcoxon signed ranks test. RESULTS: Recordings obtained via ultrasound demonstrated significantly more fetal signal loss than those obtained via the direct mode, particularly in the second stage. The FIGO criteria for fetal signal loss with external ultrasound were not fulfilled during this stage for about half the cases. CONCLUSION: Intrapartum FHR monitoring via a scalp electrode provides far better quality FHR signals than external ultrasound and deserves a more prominent position in fetal surveillance than it currently has. PMID- 15294363 TI - Evaluating the decision--to--delivery interval in emergency caesarean sections. AB - OBJECTIVE: To assess the interval between the decision to carry out an emergency caesarean section and delivery, and to determine whether this interval can be shortened. STUDY DESIGN: A retrospective study was performed in a French maternity hospital over a 6-month period. All caesarean sections performed during labour were included. These caesarean sections were divided into two groups according to Lucas's classification: (1) emergency and urgent caesarean sections and (2) scheduled caesarean sections. RESULTS: The mean decision--to--delivery interval was 39.5 min in the first group and 55.9 min in the second group. It was mainly influenced by the time taken to get the patient into theatre. The mean decision-to-operating theatre interval accounted for 45.6 and 53.8% of the mean decision-to delivery-interval, respectively. CONCLUSION: The recommended interval of 30 min is not routinely achieved. Improving communication within the perinatal team could decrease the decision--to--operating theatre interval and should be promoted. PMID- 15294364 TI - Influence of misoprostol or prostaglandin E(2) for induction of labor on the incidence of pathological CTG tracing: a randomized trial. AB - OBJECTIVE: To compare the efficacy and safety of misoprostol (prostaglandin E(1) (PGE(1))) with dinoprostone (prostaglandin E(2) (PGE(2))) for third trimester cervical ripening and labor induction. STUDY DESIGN: Patients requiring induction of labor were randomly assigned to receive either 50 microg of intravaginal misoprostol every 4 h or 0.5 mg of intracervical dinoprostone gel every 6 h. Eligibility criteria included gestation = 36 weeks. Primary outcome was the time interval from induction to delivery; secondary outcomes were mode of delivery, perinatal outcome, and interpretation of cardiotocogram (CTG) records. RESULTS: Two hundred women were randomly enrolled to receive either misoprostol (n = 100) or dinoprostone (n = 100). Time induction-to-delivery at 12, 24 and 48 h and the need for oxytocin were reduced with misoprostol (P < 0.05). Pathological CTG tracing according to FIGO and Melchior scores were more frequent in the misoprostol-treated group (P < 0.001). CONCLUSION: Misoprostol shortened the induction-to-delivery interval, but is associated with a higher incidence of abnormal CTG than prostaglandin E(2). PMID- 15294365 TI - Prediction of successful vaginal delivery in women undergoing external cephalic version at term for breech presentation. AB - OBJECTIVE: The aim of this study is to evaluate the clinical and sonographic predictors of ultimate successful vaginal delivery in women undergoing external cephalic version. STUDY DESIGN: The study population consisted of women with external cephalic version performed at or after 36 weeks of gestation. They were randomized into group A or B, each consisted of half of the total study population. Logistic regression was performed on group A to identify the significant independent variables in predicting successful cephalic vaginal delivery, which were used to construct a prediction model. The derived regression model was then tested in group B to assess its accuracy. RESULTS: The study included 407 pregnancies. Maternal weight (kg) at the time of version, maternal height (cm), multiparity, engagement of fetal presenting part, and fetal head palpable were significant independent variables of successful version and vaginal delivery (regression coefficients are: -0.084, 0.085, 1.752, -1.271, and 0.725, respectively). A prediction model was constructed based on these independent variables. The weighted average of the overall accuracy in predicting success or failure of version and vaginal delivery was 70.9%. The regression model was then applied to study group B. Using 0.50 as the cutoff value, the sensitivity, specificity, positive, and negative predictive values were 75.4, 58.8, 73.7, and 61.0%, respectively. CONCLUSION: A regression model constructed based on clinical variables failed to provide an accurate predictive tool of successful external cephalic version and vaginal delivery. However, in women who are equivocal about external cephalic version, a high prediction of success would be encouraging. PMID- 15294366 TI - Precipitate labor: higher rates of maternal complications. AB - OBJECTIVE: The study was aimed to identify risk factors and to elucidate pregnancy outcome following precipitate labor, i.e. expulsion of the fetus within less than 3 h of commencement of contractions. METHODS: A comparison of patients with and without precipitate labor, delivered during the years 1988-2002, was conducted. Patients who underwent cesarean deliveries were excluded from the analysis. A multiple logistic regression model, with backward elimination, was performed to investigate independent risk factors for precipitate labor. RESULTS: The number of vaginal deliveries that occurred during the study period was 137,171. Of these, 99 were precipitate. Independent risk factors for precipitate labor, using a backward, stepwise multivariate analysis were: placental abruption (odds ratio (OR) = 30.9, 95% confidence interval (CI) 15.9-60.4, P < 0.001); fertility treatments (OR = 3.9, 95% CI 1.7-9.0, P = 0.002); chronic hypertension (OR = 3.1, 95% CI 1.2-7.8, P = 0.015); intrauterine growth restriction (IUGR) (OR = 2.9, 95% CI 1.2-6.8, P = 0.014); prostaglandin E2 induction (OR = 1.9, 95% CI 1.1-3.5, P = 0.045); birth weight < 2,500 g (OR = 1.8, 95% CI 1.1-3.1, P = 0.020); and nulliparity (OR = 1.7, 95% CI 1.1-2.6, P = 0.014). No significant differences were noted between the groups regarding perinatal complications such as meconium stained amniotic fluid, perinatal mortality and low Apgar scores. However, there were higher rates of maternal complications in the precipitate labor group such as cervical tears and grade 3 perineal tears (18.2% versus 0.3%, P < 0.001; and 2.0% versus 0.1%, P < 0.001, respectively), post-partum hemorrhage (13.1% versus 0.4%, P < 0.001); retained placenta (2.0% versus 0.5%, P = 0.02); the need for revision of uterine cavity and packed-cells transfusions (34.3% versus 4.9%, P < 0.001; and 11.1% versus 1.1%, P < 0.001, respectively) and prolonged hospitalization (27.6% versus 19.2%, P = 0.035) as compared to the controls. CONCLUSION: Precipitate labor is associated with higher rates of maternal complications. PMID- 15294367 TI - Associations of polymorphisms of the angiotensinogen M235 polymorphism and angiotensin-converting-enzyme intron 16 insertion/deletion polymorphism with preeclampsia in Korean women. AB - OBJECTIVE: The contribution of genetic factors to preeclampsia has been well documented. However, there has not been any study done on the association between preeclampsia and the angiotensinogen (AGT) M235T polymorphism and angiotensin converting-enzyme (ACE) intron 16 insertion/deletion (I/D) polymorphism among Korean preeclampsia women. We performed a hospital-based case-control study on Korean women to investigate the association between preeclampsia and the angiotensinogen M235T polymorphism and also to determine the association between preeclampsia and the angiotensin-converting-enzyme intron 16 polymorphism. METHODS: DNA was extracted from whole blood of 104 preeclampsia patients and 114 healthy pregnant women. All samples were genotyped for all the polymorphisms using amplification after PCR of known allelic variants. Results were analyzed with the chi-square test, Student's t-test, and logistic regression. RESULTS: 18 of 50 women with preeclampsia (36.0%) in nulliparous women and 15 of 37 women with preeclampsia (40.5%) in parous women were homozygous for methionine (M235) to threonine (T235) substitution at residue 235 of AGT gene, versus 12 of 38 women in nulliparous control women and 18 of 50 women in parous control women. There was no association between the AGT M235T polymorphism and preeclampsia according to age. Fourteen of 55 women with preeclampsia (25.5%) in nulliparous women and 11 of 39 women with preeclampsia (28.2%) in parous women were homozygous for the D allele of the ACE intron 16, versus 9 of 52 women in nulliparous control women and 16 of 53 women in parous control women. No association was demonstrated between D allele of ACE intron 16 and preeclampsia according to age. There were significant differences in birth weight and delivery weeks between controls and preeclampsia patients (P < 0.001). There were no significant differences in age and nulliparity between controls and preeclampsia patients. CONCLUSION: The result indicates that the AGT M235T polymorphism and the ACE intron 16 polymorphism play no significant role in preeclampsia observed in Korean women. PMID- 15294368 TI - A trial of a new regimen with clomiphene citrate administration to reduce the antiestrogenic effects on reproductive end organs. AB - OBJECTIVE: To determine the endocrinological and clinical outcomes of a 3-day clomiphene citrate (CC) regimen and compare it with the classical 5-day clomiphene citrate regimen. STUDY DESIGN: 59 patients, diagnosed with Class II ovulatory deficiency according to the criteria defined by WHO, were randomized into two groups. Patients in Group I received 50mg per day of CC for 3 days starting on the first day of the cycle during 72 cycles. Group II received 50mg per day of CC for 5 days starting on the fifth day during 64 cycles. RESULT(S): The ovulation rate was significantly higher in Group II (78.11%) compared to Group I (63.88%) (P < 0.05). However, the implantation rate was higher in Group I than Group II. CONCLUSION: We observed that starting CC on the first day of the cycle for 3 days would lead to higher implantation rates compared to the classical 5-day CC therapy. PMID- 15294369 TI - Parental decisions following the prenatal diagnosis of sex chromosome abnormalities. AB - OBJECTIVE: To report parental decisions regarding pregnancy termination following the prenatal diagnosis of a sex chromosome abnormality (SCA) in the fetus. METHODS: Retrospective collection of data from records of 61 families receiving genetic counseling after prenatal diagnosis of a sex chromosome abnormality in the fetus in the Division of Medical Genetics, University Hospital of Geneva during the time period 1980-2001. RESULTS: Among 61 couples with a prenatal diagnosis of a sex chromosome abnormality (SCA), 44 couples (72.1%) decided to terminate pregnancy. Pregnancy termination rates were 100, 73.9, 70, 50 and 42.9% for Turner syndrome, Klinefelter syndrome, 47,XXX females, 47,XYY males, and mosaic cases, respectively. In all 11 cases with a fetal abnormality seen on ultrasound, pregnancy was terminated. Termination rates were higher among couples with a higher mean number of previous children. Maternal age and year of test did not influence parental decisions. CONCLUSIONS: Parental decision to terminate a pregnancy for a fetus with a SCA varied by type of sex chromosome abnormality, by presence of fetal ultrasound anomalies, and by the mean number of previous children. PMID- 15294370 TI - Effect of cryopreservation on quality and fertilization capacity of human sperm. AB - OBJECTIVE: To analyze retrospectively the effect of cryopreservation on donor's sperm. STUDY DESIGN: Data were collected on 178 cryopreserved-thawed sperm specimens from 44 donors and 624 oocytes from 62 women, which underwent in vitro fertilization-embryo transfer (IVF-ET) treatments with donor's sperm. Data on fresh sperm, 175 sperm specimens from 76 couples which underwent IVF-ET treatments, were used as a control group. Semen analysis was done by cell concentration, percent of motility, and quality of motility according to the World Health Organization (WHO) recommendation. Sperm quality parameters which had the strongest impact on fertilization capacity were determined using the statistical response surface model and conjoint analysis. RESULTS: Passing sperm through Percoll column decreased sperm concentration, with no improvement in sperm motility but with a slight increase in quality of motility. Quality of motility of donor's sperm had the strongest impact on fertilization capacity. CONCLUSION: Current freezing-thawing protocols of sperm cause a decrease in sperm parameters without affecting fertilization capacity. Furthermore, quality of motility of frozen-thawed sperm seems to be a significant measure of sperm fertilization capacity. PMID- 15294371 TI - Tension-free vaginal tape (TVT) in women with low urethral closure pressure. AB - OBJECTIVE(S): Aim of the study was to assess the effectiveness and the complications associated with the use of tension-free vaginal tape (TVT) in women with stress urinary incontinence and low urethral closure pressure (LUCP). STUDY DESIGN: Thirty-seven patients with stress urinary incontinence and LUCP who were treated with the TVT procedure have been included in the study. Physical examination and urodynamic investigations were carried out to all women preoperatively and at 6, 12 and 26 months (average, range: 22-30 months), postoperatively. The mean age of the patients was 69 years (+/-13), while mean parity was 2.2 (range 0-3). RESULTS: TVT procedure was carried out in all patients with epidural anesthesia. Postoperative evaluation showed 27 patients (73%) to have been completely cured, four (9.25%) to have a considerable improvement, whereas six patients (16.2%) were classified as failures. Only a few complications occurred. CONCLUSION(S): Our study indicates that the TVT procedure is an effective and well-accepted minimal invasive surgery for treatment of urinary stress incontinence in women with LUCP. The cure rate of 73% could be considered satisfactory. Women with LUCP and 'fixed' urethra, are at significantly increased risk of failure of the procedure. PMID- 15294372 TI - Sacro-spinous ligament fixation peri-operative complications in 195 cases: visual approach versus digital approach of the sacro-spinous ligament. AB - OBJECTIVE: To evaluate sacro-spinous ligament fixation (SLF) peri-operative complications. STUDY DESIGN: Monocentric, retrospective study. Department of Gynecology, SIHCUS-CMCO, University Hospital, Strasbourg, France. Between January 1990 and December 2000, 195 women, mean age 63.2 years old (40-90), underwent a vaginal SLF. Ninety point eight percent of women were post-menopaused and 27.9% of these had a hormonal substitution. About 24% of patients had prior hysterectomy, 20% vaginal prolapse repair and 22% urinary stress incontinence repair. SLF was performed in 1.5% of cases without any other procedures and it was combined with the following: rectocele and elytrocele repair in 89.2%, hysterectomy in 72.3%, cystocele repair in 52.8% and stress incontinence repair in 15.3% of cases. In 107 cases, the SLF attachment was placed under digital control and in 88 cases under visual control. RESULTS: The mean hospitalisation stay was of 8.5 +/- 2.6 days (4-26). About 41% of women presented a complication. Major complications were represented by 3.6% of bladder injury, 0.5% of uretero vaginal fistula, 0.5% of vascular injuries, 0.5% of thromboembolic events. In 38% of cases patients had minor complications: urinary tract infections (29%), temporary urinary retention (5.6%), local complications (4.5%), and other complications (3%). The only specific SLF complication in this data was a vascular injury and in this case the SLF was performed under digital control. CONCLUSIONS: The global peri-operative complication frequency of SLF is high. It is mainly represented by non-specific complications, secondary to the combined procedures and not to the SLF itself. The specific complications due to SLF, all of which are major ones, can be avoided or diagnosed earlier, by using the visual approach technique. PMID- 15294373 TI - Laparoscopic burch colposuspension: comparison of effectiveness of extraperitoneal and transperitoneal techniques. AB - OBJECTIVE: To compare the effectiveness of two different laparoscopic colposuspension procedures: extraperitoneal approach using mesh fixed with tacks, and transperitoneal approach using sutures. STUDY DESIGN: We conducted a retrospective study of all patients (n = 64) who had undergone two different techniques of laparoscopic Burch colposuspension without additional surgeries over a 6-year period. Thirty-six women underwent laparoscopic transperitoneal colposuspension with using nonabsorbable sutures (group A), whereas 28 women underwent laparoscopic extraperitoneal colposuspension with using mesh and preperitoneal balloon dissection technique (group B). Cure rate was assessed by simple cystometry with a cough stress test in the standing position. Both groups were compared with regard to cure rates, operative time, length of hospital stay, complications, estimated blood loss, and total hospital charges. RESULTS: The mean times to follow-up were 25.7 months in the group A and 27.3 months in the group B (P = 0.082). At last follow-up, 33 of 36 (91.7%) patients in the group A and 23 of 28 (82.1%) patients in the group B were continent (P = 0.22). The other results were as follows for group A and B, respectively: average duration of surgery, 58.1 compared with 46.8 min (P = 0.001); average hospital stay, 2.05 compared with 1.57 days (P = 0.02); the intraoperative complication rate, 8.3% compared with 7.1% (P = 0.62). The total hospital charges for the group B were found significantly higher (US dollars 2,234 versus US dollars 1,348, P = 0.001). CONCLUSION: Although we found higher cure rates in laparoscopic colposuspension with the transperitoneal approach using sutures than the extraperitoneal approach using mesh fixed with tacks, there was no statistically significant difference between the two procedures. In comparison with extraperitoneal mesh technique, lower cost is the superiority of the transperitoneal suture technique. PMID- 15294374 TI - Use of CA125 fluctuation during the menstrual cycle as a tool in the clinical diagnosis of endometriosis; a preliminary report. AB - OBJECTIVE: To elucidate whether endometriosis can be diagnosed clinically by assessing the differences between serum CA125 levels during menstruation and during the rest of the menstrual cycle. METHODS: The study was performed in 28 patients who underwent laparoscopy to check for pelvic causes of infertility. Patients with endometriosis were selected as the study group, and patients with normal laparoscopic findings functioned as the control group. Blood specimens were taken for CA125 determination during menstruation and during the rest of the menstrual cycle. Mean serum CA125 concentrations were compared by the two-sample t-test for between-group comparisons and the paired t-test for within-group comparisons. The receiver operating characteristic curve was applied to assess the usefulness of CA125 level changes during the menstrual cycle in the clinical diagnosis of endometriosis. RESULTS: The mean CA125 concentrations of healthy women during menstruation and during the rest of the menstrual cycle were 12.2 and 10 U ml(-1), respectively. In this group, the mean CA125 concentration was an average of 22% higher during menstruation than during the rest of the menstrual cycle (P < 0.001). The patients with endometriosis showed a similar pattern to that of normal women, but the levels differed by 198.3% in these patients (P < 0.001). Mean CA125 concentrations of these patients during menstruation and in the rest of the cycle were 35.8 and 12 U ml(-1), respectively. The mean CA125 concentration during menstruation was significantly higher in patients with endometriosis than in normal women (P < 0.001), but CA125 concentrations at other points in the menstrual cycle were found to be similar in both groups (P > 0.05). ROC curve analyses set a cutoff of 83% (percentage increment of CA125 level during menstruation compared with that on days without menstrual bleeding), which gives a sensitivity of 93% and specificity of 92%, with a corresponding likelihood ratio of 11.3. CONCLUSIONS: It may be possible to diagnose endometriosis clinically by assessment of the differences in CA125 level during menstruation as against the remainder of the menstrual cycle. PMID- 15294375 TI - Genomic alterations in the endometrium may be a proximate cause for endometriosis. AB - OBJECTIVE: To test the hypothesis that endometriosis may originate from genomic alterations in the endometrium by genomic analysis of endometrial tissues in patients with endometriosis and compare them with those from normal controls. METHODS: Endometrial tissue samples were taken from five women with endometriosis. For controls, we used endometrial tissue samples from four women who underwent elective abortions and one sample from placenta. Using array-based comparative genomic hybridization (CGH), we determined the normal range of variation in CGH signals using normal controls. CGH results were further confirmed by real-time quantitative PCR and loss of heterozygosity analysis. RESULTS: We identified several regions of genomic alterations in all five patients. Some of these regions were the same regions identified previously in endometriotic lesions. For select markers, the genomic alterations were confirmed by real-time PCR and LOH analyses. CONCLUSIONS: There is evidence that the endometrium in women with endometriosis has genomic alterations. This is consistent with numerous reports that the endometrium of women with endometriosis differ from those of women without. Our finding suggests that genomic alterations in the endometrium may be a proximate cause for endometriosis. PMID- 15294376 TI - The treatment with a COX-2 specific inhibitor is effective in the management of pain related to endometriosis. AB - OBJECTIVE: To evaluate the efficacy and safety of a cyclooxygenase (COX)-2 specific inhibitors versus placebo in the treatment of endometriosis-associated pelvic pain. STUDY DESIGN: A group of women (n = 28) with pelvic pain after conservative surgery for symptomatic endometriosis (Stage I and II) were enrolled at the Department of Pediatric, Obstetrics and Reproductive Medicine of University of Siena. A treatment with a COX-2 specific inhibitors (rofecoxib, 25mg per day) (n = 16) or placebo (n = 12) was given for 6 months. Pelvic pain quantification with a clinical evaluation, including Visual Analogue Scale (VAS) for pain, was performed before and up to 6 months after treatment. RESULTS: A significant improvement of both pelvic pain and dyspareunia was observed after a 6 months persisting since the end of the treatment (P < 0.0001). The efficacy of rofecoxib was higher than placebo and no recurrence occurred, while in the placebo-treatment a 16% (2/12) occurred. No significant side effects have been found with the use of rofecoxib. CONCLUSIONS: The use of COX-2 specific inhibitors was effective, safe and low cost therapy in the management of pelvic pain associated to endometriosis and might be also proposed in early stage of endometriosis. PMID- 15294377 TI - The usefulness of Tc-99m-HMPAO-labeled leukocyte scintigraphy in the diagnosis of multiple intra-abdominal abscesses following in vitro fertilization (IVF) procedure. AB - Pelvic infection is a rare but well-known complication following IVF procedures with a reported incidence <1%. A case of multiple abdominal abscesses following IVF procedure was established by Tc-99m-HMPAO-labeled leukocyte scintigraphy and confirmed by laparotomy. This imaging technique should be recommended for early and precise diagnosis of pelvic infection following IVF. PMID- 15294378 TI - Spontaneous reversal of mirror syndrome in a twin pregnancy after a single fetal death. AB - The case report illustrates that pre-eclampsia like symptoms can arise as a consequence of pathological changes in a single feto-placental unit of a twin pregnancy and may resolve spontaneously when the cause is removed. PMID- 15294379 TI - Myasthenia and HELLP syndrome. PMID- 15294380 TI - Exposure to sibutramine during pregnancy: a case series. PMID- 15294381 TI - Sertoli-Leydig cell androgens-estrogens secreting tumor of the ovary: ultra conservative surgery. PMID- 15294382 TI - Pyoderma gangrenosum at the incision site following gynecologic surgery. PMID- 15294384 TI - The influence of perceived control and locus of control on the cortisol and subjective responses to stress. AB - Stress has been implicated in the etiology of numerous mental and physical illnesses. Thus, it is important to identify factors that buffer individuals against stress. The purpose of this study was to examine the influence of perceived control (PC) on the biological and subjective stress responses, and to investigate the potential moderating effect of locus of control (LOC) on this relationship. Stress was induced with a noise-cognitive paradigm, and PC was manipulated by offering the option of manual control over noise intensity. Saliva cortisol and subjective stress were measured. There was no main effect of control on cortisol. However, LOC moderated the relation between control and cortisol; participants with more internal LOC, who also perceived themselves to have control over the stressor, showed a reduced cortisol response in the PC condition. The results are discussed in light of their implications for elucidating the determinants of the effects of perceived control on stress. PMID- 15294385 TI - Higher Beck depression scores predict delayed epinephrine recovery after acute psychological stress independent of baseline levels of stress and mood. AB - Depressive symptoms in the non-clinical range have been linked to increased health risks. Recent theorizing raises the possibility that heightened physiologic responses to acute stress and/or slowed stress recovery in individuals with depressive symptoms may contribute to increased risk. We investigated stress-induced catecholamine responses and recovery patterns using a modified version of the Trier Social Stress Test (15 min) with a sample of 52 healthy women and compared subgroups with high normal versus low scores on the Beck Depression Inventory (BDI, median split) to 29 women randomly assigned to a non-stressed control group. The BDI-high normal and BDI-low groups showed similar acute increases in epinephrine immediately post stressor, but only the BDI-high normal group remained significantly elevated above control group levels during the recovery period. No differences were found in norepinephrine responses. Elevations in BDI scores within the normal range may selectively predict slower physiological recovery following acute stress. PMID- 15294386 TI - Impact of stress reduction instructions on stress and cortisol levels during pregnancy. AB - This pilot study examined whether giving stress reduction (SR) instructions to pregnant women would be effective in regulating stress, mood, and cortisol levels during pregnancy. Forty-one predominantly low-income Latina women, receiving prenatal services at a public county hospital, completed measures of stress and mood (depressive symptoms, positive and negative affect) and provided morning and evening saliva samples to measure cortisol prior to and after receiving SR instructions. We hypothesized that adherence to these SR instructions would result in lower levels of stress, negative mood states, and cortisol levels when compared to baseline values. Repeated measures ANOVA analyses demonstrated significantly lower levels of stress (P < 0.001), lower symptoms of depression and negative affect (P < 0.001), and lower levels of morning cortisol (P = 0.01) under the SR condition, compared to baseline. Health behaviors that were engaged in during the SR condition and implications for prenatal health interventions are discussed. PMID- 15294387 TI - An evaluation of an adaptive automation system using a cognitive vigilance task. AB - The performance of an adaptive automation system was evaluated using a cognitive vigilance task. Participants responded to the presence of a green "K" in an array of two, five, or nine distractor stimuli during a 40-min vigil. The array with the target stimulus was presented once each minute. Participants EEG was recorded and an engagement index (EI = 20 x beta/(alpha + theta)) was derived. In the negative feedback condition, increases in the EI caused the number of stimuli in the array to decrease while decreases in the EI caused the number of stimuli to increase. For the positive feedback condition, increases in the index caused an increase in the array size (AS) while decreases caused a decrease in the array size. Each experimental participant had a yoked control partner who received the same pattern of changes in array irrespective of their engagement index. A vigilance decrement was seen only for the positive feedback, experimental group. PMID- 15294388 TI - Atypical brainstem representation of onset and formant structure of speech sounds in children with language-based learning problems. AB - This study investigated how the human auditory brainstem represents constituent elements of speech sounds differently in children with language-based learning problems (LP, n = 9) compared to normal children (NL, n = 11), especially under stress of rapid stimulation. Children were chosen for this study based on performance on measures of reading and spelling and measures of syllable discrimination. In response to the onset of the speech sound /da/, wave V-V(n) of the auditory brainstem response (ABR) had a significantly shallower slope in LP children, suggesting longer duration and/or smaller amplitude. The amplitude of the frequency following response (FFR) was diminished in LP subjects over the 229 686 Hz range, which corresponds to the first formant of the/da/ stimulus, while activity at 114 Hz, representing the fundamental frequency of /da/, was no different between groups. Normal indicators of auditory peripheral integrity suggest a central, neural origin of these differences. These data suggest that poor representation of crucial components of speech sounds could contribute to difficulties with higher-level language processes. PMID- 15294389 TI - Roughness perception in sounds: behavioral and ERP evidence. AB - The mismatch negativity (MMN) correlates of the perception of roughness, the unpleasant character of sounds caused by the perception of amplitude fluctuation in the range of 20-200 Hz, were studied on the basis of a variation in the degree of modulation (=modulation index m), which is a main parameter influencing roughness. A psychophysical study showed that perceived roughness of tones increased with modulation index for m-values from 0 up to 1.2. For larger values of m, roughness perception remained stable. In a subsequent ERP-study, infrequent amplitude modulated (AM) tones with varying modulation index were presented in the context of a series of pure tones in an ignore condition. The amplitude of the mismatch negativity correlated highly with the roughness ratings (r = -0.93) and did not increase monotonously with increasing modulation index. We conclude that perceived roughness rather than its physical correlate in sounds is reflected by the MMN and that roughness is thus preattentively encoded. PMID- 15294390 TI - Impact of low frequency transcranial magnetic stimulation on event-related brain potentials. AB - Contradictory findings exist concerning the inhibitory function of low frequency repetitive transcranial magnetic stimulation (rTMS). Therefore, the study examines the impact of different duration of low frequency rTMS on ERPs. In 17 subjects, auditory ERPs were measured before and after 1 Hz rTMS delivered over the left prefrontal cortex during 10 min (600 pulses) and 15 min (900 pulses). Results showed that 15 min of 1 Hz rTMS induced a significant increase of P300 latency. There was no effect for early ERP components (N100, P200 and N200). This study confirms and extends that 1 Hz rTMS produces a real inhibitory effect only when the duration of the stimulation is about 15 min. The data suggest that rTMS modifies the speed of cognitive processing rather than the energetical aspect of information processing, and that cortical inhibition induced by the magnetic stimulation affects principally the controlled cognitive processes and not the automatic ones. PMID- 15294391 TI - Differences in startle modulation during instructed threat and selective attention. AB - This study investigated whether attentional processes contribute to fear potentiated startle. Ten subjects participated in a threat of shock experiment and an attentional control condition. In the threat of shock experiment, visual cues indicated whether or not an aversive shock might occur. In the attentional control, the shocks were replaced by faint vibrotactile stimuli that had to be counted. The P300 amplitudes of the ERP evoked by the visual cues did not differ under threat and counting, which suggested that both conditions engaged attention to the same extent. In contrast, startle potentiation in the threat condition was an order of magnitude larger than the marginally significant attentional startle facilitation in the counting condition. These results indicate that an attentional contribution to fear-potentiated startle under the present experimental conditions is small. In addition, contextual effects of threat of shock became manifest as baseline startle was facilitated relative to the attention condition. This may reflect a more sustained state of anxiety on which cue-specific fear responses are superimposed. PMID- 15294392 TI - Respiratory responses during affective picture viewing. AB - Previous research has demonstrated covariation of physiological responding with judgments of valence and arousal. However, until now links between these affective dimensions and respiratory measures have not been extensively investigated. In this study, eight picture series of different affective valence and arousal level were shown to 30 subjects, while respiration, skin conductance level (SCL), heart rate (HR) and affective judgments were measured. With increasing pleasantness, inspiratory time lengthened, mean inspiratory flow decreased and thoracic breathing increased. With increasing arousal, inspiratory time and total breath duration shortened and mean inspiratory flow, minute ventilation, thoracic breathing and electrodermal activity increased. These findings confirm the importance of arousal in respiratory responding, but also indicate a modulatory role of affective valence. We propose that the arousal effects reflect energy mobilization in preparation to act, and that the valence effects might be a manifestation of an attention bias toward negative stimuli. PMID- 15294393 TI - Second to fourth digit ratio and hand skill in Austrian children. AB - Prenatal exposure to testosterone is thought to promote the development of the right-hemisphere and increase the incidence of sinistrality. A direct test of this hypothesis has previously been problematic because of the difficulty of indirectly assessing prenatal sex steroid exposure. Evidence now suggests that the ratio between the length of the second and fourth digits (2D:4D) is related to prenatal testosterone exposure. We tested whether digit ratio is related to the degree of hand skill such that low 2D:4D (indicating high levels of testosterone in utero) may be correlated with enhanced left-hand performance. In right-handed children, high 2D:4D correlated with improved right-hand skill and low 2D:4D correlated with enhanced left-hand skill. Correlations were found to be similar for girls and for boys. Since low 2D:4D has been previously reported to be associated with faster left-hand speed compared to right in Afro-Caribbean children with very low mean 2D:4D, the present finding in a Caucasian population with high mean 2D:4D suggests that a tendency of improved left-hand performance due to prenatal testosterone may be found across ethnic groups. PMID- 15294394 TI - Baroreceptor sensitivity and effectiveness varies differentially as a function of cognitive-attentional demands. AB - From the spontaneous sequence method, a new index of baroreceptor function has recently been proposed, the baroreflex effectiveness index (BEI). BEI quantifies the number of times the baroreflex is effective in driving the sinus node. In this study we examined how different cognitive-attentional demands modulates BEI and baroreceptor reflex sensitivity (BRS). Eighty three students performed three tasks: mental arithmetic, memory, and visual attention. Results indicate that BRS reliably decreases during mental arithmetic and increases slightly during visual attention. BEI increases during the visual attention task. The overall pressure change of the systolic blood pressure ramps decreases during tasks with respect to baseline periods and cannot explain the effect found in BEI (in effect, BEI works against this underlining influence). The modulation found in BRS and BEI as a function of cognitive demand is in accordance with the Laceys' intake/rejection theory. Specifically, it is suggested that BRS is more sensitive to internal cognitive elaboration conditions, while BEI is more sensitive to external attention conditions. PMID- 15294395 TI - Abstracts of the 10th annual meeting of the International Society for the Advancement of Respiratory Psychophysiology (ISARP). PMID- 15294397 TI - Resuscitation greats. Ambroise Pare and the arrest of haemorrhage. PMID- 15294398 TI - Association between clinically abnormal observations and subsequent in-hospital mortality: a prospective study. AB - BACKGROUND: Patients with unexpected in-hospital cardiac arrest often have an abnormal clinical observation prior to the arrest. Previous studies have suggested that a medical emergency team responding to such patients may decrease in-hospital mortality from cardiac arrest, but the association between any abnormal clinical observation and subsequent increased mortality has not been studied prospectively. The aim of this study was to determine the predictive value of selected abnormal clinical observations in a ward population for subsequent in-hospital mortality. DESIGN AND SETTING: Prospective data collection in five general hospital ward areas at Dandenong Hospital, Victoria, Australia. INTERVENTIONS: None. RESULTS: During the study period, 6303 patients were admitted to the study areas. Of those, 564 (8.9%) experienced 1598 pre-determined clinically abnormal events and 146 of these patients (26%) died. The two commonest abnormal clinical events were arterial oxygen desaturation (51% of all events), and hypotension (17.3% of all events). Using a multiple linear logistic regression model, there were six clinical observations which were significant predictors of mortality. These were: a decrease in Glasgow Coma Score by two points, onset of coma, hypotension (<90 mmHg), respiratory rate <6 min(-1), oxygen saturation <90%, and bradycardia >30 min(-1). The presence of any one of the six events was associated with a 6.8-fold (95% CI: 2.7-17.1) increase in the risk of mortality. CONCLUSIONS: Six abnormal clinical observations are associated with a high risk of mortality for in-hospital patients. These observations should be included as criteria for the early identification of patients at higher risk of unexpected in-hospital cardiac arrest. PMID- 15294399 TI - A prospective, multicenter pilot study to evaluate the feasibility and safety of using the CoolGard System and Icy catheter following cardiac arrest. AB - BACKGROUND: Cardiac arrest causes devastating neurological morbidity and mortality. Mild/moderate hypothermia is neuroprotective after global cerebral ischemia. More rapid controlled attainment of the target temperature may increase efficacy. METHODS: We assessed the safety and feasibility of endovascular cooling in a single arm study of comatose patients who had been successfully resuscitated after cardiac arrest. Core temperature was reduced to a target of 33 degrees C for 24 h using a closed loop endovascular system placed in the inferior vena cava, followed by controlled rewarming. Primary outcomes were speed and accuracy of cooling, survival and GOS after 30 days. RESULTS: Thirteen patients were enrolled, six male, age 60 +/- 19 years. Time from cardiac arrest to return of spontaneous circulation was 14.3 min (range 5-32.5). It took 3h and 39 min (median 210 min, IQ 80-315) to reach 33 degrees C; cooling averaged 0.8 +/- 0.3 degrees C/h (range 0.22-1.12 degrees C/h). Temperature was tightly maintained for all patients averaging 32.7 +/- 0.5 degrees C. Rewarming lasted 18.3 +/- 5.9 h. Five patients (38%) had 30-day Glasgow Outcome Scores of 1-2. Four patients died, none related to the hypothermia procedure. No unanticipated or procedure-related adverse events occurred. CONCLUSION: In comatose survivors of cardiac arrest, hypothermia via endovascular methods is safe and feasible, and target temperatures can be achieved and controlled rapidly and precisely. More studies are needed to assess the efficacy of rapid endovascular hypothermia after cardiac arrest. PMID- 15294400 TI - Laryngeal mask airway: is the management of neonates requiring positive pressure ventilation at birth changing? AB - OBJECTIVE: To evaluate the impact of the laryngeal mask airway (LMA) on neonatal resuscitation policy. DESIGN: We analyzed retrospectively the records of neonates requiring positive pressure ventilation (PPV) at birth before (1996) and after (2000) the introduction of the LMA into our delivery suites. In addition, the outcome of neonates treated with the LMA was compared with that of neonates matched for gestational age and mode of delivery who were resuscitated using a face mask. RESULTS: During the year 2000, 95 out of 380 (25%) resuscitated neonates were treated with the LMA. The LMA was effective in 94 out of 95 (99%) of these infants. Over the same period, the percentage of neonates receiving tracheal intubation (TI) at birth (34%) was significantly reduced compared with the figure for 1996 (67%). There were no reported complications associated with the use of the LMA. Seventy-four out of the 95 neonates treated with the LMA were considered suitable for matching for gestational age and mode of delivery with 74 neonates treated with a face mask. No differences were found between the two groups for birth weight, Apgar scores, need for tracheal intubation, need for admission to the Neonatal Intensive Care Unit (NICU), primary diagnosis at discharge and primary outcomes. The LMA provided effective ventilation in four neonates in whom the face mask failed. CONCLUSIONS: The LMA is changing neonatal resuscitation practice in our Institution. Our data suggest that it is a safe and useful alternative method for respiratory support in neonates requiring PPV at birth, which merits further study. PMID- 15294401 TI - Learning effect of a novel interactive basic life support CD: the JUST system. AB - INTRODUCTION: Electronic interactive learning environments can enhance the learning experience and may prove beneficial in basic life support (BLS) training. As part of the European Union funded project "JUST-in-time health emergency interventions-training of non-professionals by virtual reality and advanced IT tools", an innovative interactive CD-ROM on BLS and other emergency medicine topics was developed. We hypothesised that individuals without previous BLS training could learn CPR techniques from this CD. METHODS: Sixty-two students were randomised into a group studying the JUST CD in a computer class room for up to 60 min, and a control group who did not receive any training (serving as a reference). CD users also completed a satisfaction questionnaire immediately after studying the CD. The BLS skills of both groups were assessed in a mock BLS scenario on a training manikin. BLS performance was video recorded and analysed. RESULTS: After studying the CD for a mean period of 42 min, users of the CD had better assessment skills and were more likely to show a positive helping attitude, but chest compression and breathing techniques were ineffective. Most users rated the CD as very good and a positive learning tool. CONCLUSION: Individuals without prior BLS training showed improved behaviour and assessment skills after exposure to the CD, but motor skill acquisition requires alternative learning strategies. PMID- 15294402 TI - Differing operational outcomes with six commercially available automated external defibrillators. AB - INTRODUCTION: In general automated external defibrillators (AED) are handled easily, but some untrained lay rescuers may have major problems with the use of such products. This may result in delayed shock delivery and delay in basic life support (BLS) after use of the AED. To study the effect of voice prompts and design solutions we tested the time from the first shock to the initiation of BLS for six defibrillators available in Austria. METHODS: Volunteers, who had no AED training, were evaluated to see when they delivered the first shock and how often BLS was started after the voice prompts were given by the defibrillators. RESULTS: Time to first shock delivered ranged from 78 (95% CI: 68-89) to 128 (95% CI: 110-146)s. The defibrillator-type had a significant influence on the time to first shock delivered (P < 0.0001). The proportion of volunteers who started BLS after defibrillation ranged from 93 to 33% and differed significantly between the AEDs used (P < 0.03). CONCLUSIONS: We demonstrated that there are significant differences between AEDs, concerning important operational outcomes like time to first shock and the start of BLS. Further research and development is urgently required to optimise user-friendliness and operational outcomes. PMID- 15294403 TI - Arrhrythmias and haemodynamic effects associated with early versus late prehospital thrombolysis for acute myocardial infarction. AB - The occurrence of arrhythmias and haemodynamic changes was studied prospectively in 226 consecutive patients who received prehospital thrombolysis for acute ST elevation myocardial infarction (STEMI) in two Helicopter Emergency Medical Service (HEMS) systems in Southern Finland. Of the 226 patients, 129 were classified as receiving early (pain to treatment-time <90 min) and 97 as late (pain to treatment-time >90 min) treatment. Data on all arrhythmias and haemodynamic disturbances during the prehospital phase were collected. Arrhythmias occurred in 39% of all patients (40% in the early and 38% in the late group). A third of the patients received treatment for their arrhythmia (38% in the early group and 24% in the late group, P = NS). The most common arrhythmia was ventricular extrasystoles, which did not require any treatment in the majority of patients. On arrival of the EMS crew, 14% of all patients were hypotensive (14% in the early and 13% in the late group). After thrombolytic treatment, 7% of all patients became hypotensive (7% of the patients in both groups). The most common treatment for hypotension was fluid administration. Of the 15 patients who received thrombolysis after cardiopulmonary resuscitation (CPR), four patients suffered from arrhythmias and six patients developed hypotension after initiation of thrombolytic treatment. Although arrhythmias and haemodynamic changes were frequent in the prehospital setting after initiation of thrombolytic therapy, severe adverse events were rare. Those requiring therapeutic measures responded well to treatment. The occurrence of events was not related to the timing of thrombolysis in relation to the duration of pain. PMID- 15294404 TI - General medicine practitioners' attitudes towards "do not attempt resuscitation" orders. AB - OBJECTIVE: To examine whether "all-or-none" guidelines for cardiopulmonary resuscitation (CPR) are being applied by practitioners on general medical wards (GMWs). HYPOTHESIS: Do not attempt resuscitation (DNAR) orders are rarely related to patient preferences. Limited resuscitation efforts are being practiced to circumvent the need for DNAR orders. DESIGN: A surprise opinion survey (presented below), based on case vignette and practice description, and performed by remote control votes. SETTING: The multi-centre forum for practitioners on GMWs within the greater Jerusalem district. PARTICIPANTS: 79/85 clinicians practicing/training on GMWs in six teaching hospitals, who attended the forum and responded within 3 min to the survey. RESULTS: Fifty-eight practitioners (73%) assigned a DNAR order for a patient unable to express a preference and only 43 (55%) complied with the request of a competent patient for a DNAR order (P < 0.05]; 95% CI: 2-34). During the past year, only five practitioners (9% of respondents) had performed CPR solely when pathophysiological benefit was expected, 31 (59%) had performed limited CPR efforts and only 13 (28%) had discussed the subject of DNAR with patients and their next of kin >5 times. CONCLUSIONS: (1) DNAR orders are rarely discussed with patients and their next of kin in GMWs within the region examined; (2) even when DNAR is discussed, physicians tend to confer DNAR orders based on their personal value judgements rather than on patient preferences; (3) practitioners on GMWs perform CPR when no pathophysiological benefit is expected; (4) limited resuscitation efforts are performed frequently in GMWs. PMID- 15294405 TI - Susceptibility of automated external defibrillators to train overhead lines and metro third rails. AB - INTRODUCTION: Immediate accessibility to automated external defibrillators (AED) is recommended for highly frequented public areas. In train terminals and metro stations electromagnetic interference (EMI) is present. In preparation for a public access defibrillation (PAD) programme in this environment possible effects on AED safety and accuracy were studied. METHODS: In typical public transportation settings 11 different AED models were bench tested for their sensitivity and specificity of ECG analysis with shockable and nonshockable rhythms provided by an ECG simulator. The devices were exposed to the electromagnetic interference of a rail system operating with 15 kV alternating current (ac) with a frequency of 16 2/3 Hz and a subway system powered with 750 V direct current (dc). AED cables were setup parallel and perpendicular to the tracks, the tests were carried out at 3 m distance from the rails in an empty station and with incoming trains. RESULTS: A total of 5280 tests were recorded, each device was tested a total of 480 times. Fifteen kilovolts 16 2/3 Hz ac interfered more than 750 V dc with the tachyarrhythmia detection systems (P < 0.0001). An AED setup with electrode cables perpendicular to track and power line reduced interference (P < 0.0001), while incoming trains had no significant effect on ECG analysis (P = 0.19). Depending on the AED model, sensitivity ranged from 60 to 100% and specificity from 54 to 100%, representing a positive likelihood-ratio from 1.3 to 241 and a negative likelihood-ratio from 0.7 to 0.0. In the public transportation setting tested, four AED models were unsuitable for automated defibrillation as these devices demonstrated an unacceptable performance in respect of accuracy and safety. In the train setting two devices performed with an accuracy of 57 and 65%. One AED recommended shocks for sinus rhythm at normal frequency. In the metro setting one AED did not advise shocks for ventricular tachycardia. CONCLUSION: Shock advisory systems of some AED models are susceptible to electromagnetic interference, especially in terminals with 15 kV 16 2/3 Hz ac power supplies. Interference is minimized, if patient position is parallel and electrode cables are perpendicular to overhead line. The choice of AED model for train or metro stations depends on its lack of susceptibility to typical electromagnetic interference. PMID- 15294406 TI - Physico-chemical stability and sterility of previously prepared saline infusion solutions for use in out-of-hospital emergencies. AB - OBJECTIVE: The topic of this research was to determine whether out-of-hospital emergency teams could make use of previously prepared saline solutions (SS). The objective was to discover the physical, chemical and sterility characteristics of previously prepared saline infusions stored in ambulances and ascertain how long they remained in optimum condition. METHOD: Randomised clinical trial, triple blind, where study units consisted of saline solutions prepared with an infusion system and a three-way valve. The duration of the study was 12 months. Six intervention groups were designed on the basis of time of exposure and location. Samples consisted of 672 units. Twelve microbiological cultures were made and the pH, density, viscosity and CINa concentration were determined. We compared hypotheses with four models of linear regression for the variables and a model of logistic regression for the variables. A value of P < 0.05 was considered significant. RESULTS: We obtained results from 669 saline solutions (98.82%). Neither multivariant analysis nor ANOVA tests showed any significant association for a power greater than 99% with regard to the physical-chemical characteristics. The model of logistic regression also did not find any significant association for sterility. Colonisation was present in 1.7% of the 8,028 cultures made and more than 5 CFU per millilitre was found in only two cases. CONCLUSION: There is no evidence to suggest that recently prepared saline infusion solutions are any different from a physical-chemical and sterility point of view than those exposed for 24, 48, or 72 h. It was concluded that use can be made of previously prepared saline solutions with a guarantee their stability and sterility. PMID- 15294407 TI - Optimum cardiopulmonary resuscitation for basic and advanced life support: a simulation study. AB - Optimum cardiopulmonary resuscitation (CPR) for both basic and advanced cardiac life support depends on a compromise between the number of chest compressions delivered and the amount of ventilation provided. This study used theoretical models of blood flow and both arterial and venous blood gas values to investigate the influence of different compression to ventilation ratios on CPR efficiency, as well as the effects of different inspired oxygen concentrations. With mouth-to mouth ventilation, greater numbers of compressions between each ventilation provided progressively greater blood flow. However, a greater the number of compressions, reduced the arterial oxygen levels and carbon dioxide clearance. There was an optimum ratio, in terms of both oxygen delivery and carbon dioxide clearance, of around 20:1 compressions to ventilation. Optimum oxygen delivery was 0.19 L/min at 20:1, which was better than the oxygen delivery for standard CPR based on a ratio of 15:2 (0.13 L/min). When patients were ventilated with supplemental oxygen (either 50 or 85%) the lungs rapidly became saturated with oxygen, and oxygen delivery depended more on blood flow. Higher numbers of compressions provided greater oxygen delivery, but at the cost of increasing hypercarbia, which is thought to affect resuscitation success rates adversely. The simulation results suggested ratios around 20:1 would be the best compromise between blood flow, oxygen delivery (0.25 L/min) and avoidance of hypercarbia. The best results were provided by continuous chest compressions and simultaneous, asynchronous ventilation in an intubated patient. Arterial and venous oxygen and carbon dioxide levels were well maintained, with very good oxygen delivery (0.32 L/min). Intubation with continuous chest compressions and asynchronous ventilation can therefore significantly improve the quality of CPR as a whole, and not just ventilation. PMID- 15294408 TI - Continuous intratracheal insufflation of oxygen improves the efficacy of mechanical chest compression-active decompression CPR. AB - The aim of the present study was to compare the efficacy of intratracheal continuous insufflation of oxygen (CIO) with intermittent positive pressure ventilation (IPPV) regarding gas exchange and haemodynamics during mechanical chest compression-active decompression cardiopulmonary resuscitation (mCPR) provided by the LUCAS device. Ventricular fibrillation (VF) was induced electrically and ventilation was discontinued in 16 pigs, mean body weight 23 kg (range 22-27 kg). They were randomized into two groups (CIO versus IPPV). After 8 min of VF, mCPR was started and run for 30 min in normothermia, after which defibrillation was attempted during on-going mCPR. Return of spontaneous circulation was obtained in eight of eight CIO pigs and in four of eight IPPV pigs. Arterial oxygen tension (P < 0.05) and coronary perfusion pressure (P < 0.01) were significantly higher in the CIO pigs. Arterial CO(2)-tension was subnormal in both groups and significantly (P < 0.05) lower in the IPPV-pigs (around 4.5 versus 3.0 kPa). The intratracheal pressure differed significantly (P < 0.001) between the two groups. It was negative in each decompression phase in the IPPV pigs in spite of 6 mmHg of PEEP. The CIO pigs had a positive intratracheal pressure during the whole cycle of mCPR, with a minimum pressure of 8 mmHg during each decompression phase. To conclude, mCPR combined with CIO gave adequate ventilation and significantly better oxygenation and coronary perfusion pressure than mCPR combined with IPPV. PMID- 15294409 TI - Effects of vasopressin on left anterior descending coronary artery blood flow during extremely low cardiac output. AB - Because of the possibility of vasopressin-mediated coronary vasospasm, this study was designed to assess effects of vasopressin compared to saline placebo on left anterior descending (LAD) coronary artery blood flow. Twelve anaesthetized domestic swine were prepared for LAD coronary artery blood flow measurement with ultrasonic flow probes, using cardiopulmonary by-pass adjusted to 10% of the prearrest cardiac output. This 10% value approximates that reported for cardiac output during conventional closed-chest CPR. After 4 min of untreated ventricular fibrillation, and 3 min of cardiopulmonary by-pass blood flow, 12 pigs were randomly assigned to receive intravenously, every 5 min, either vasopressin (0.4, 0.4, and 0.8 U/kg; n = 6) or saline placebo (n = 6). The mean +/- S.D. LAD coronary artery blood flow in the vasopressin and placebo pigs was comparable before cardiac arrest, and during cardiopulmonary by-pass low flow; but increased significantly (P < 0.05) 90 s after each of three vasopressin injections compared to placebo (78 +/- 1 versus 42 +/- 2 ml/min; 62 +/- 2 versus 36 +/- 1 ml/min; and 54 +/- 1 versus 27 +/- 1 ml/min), respectively. Coronary vascular resistance decreased significantly (P < 0.05 ) 90 s after each of three vasopressin and placebo injections. In this model, repeated bolus administration of vasopressin, given during simulated extremely low cardiac output improved LAD coronary artery blood flow to prearrest levels without affecting coronary vascular resistance. CONCLUSIONS: during extremely low blood flow using cardiopulmonary by-pass, vasopressin improves LAD coronary artery blood flow without affecting coronary vascular resistance. PMID- 15294410 TI - Ketamine improves survival and suppresses IL-6 and TNFalpha production in a model of Gram-negative bacterial sepsis in rats. AB - OBJECTIVE: In a previous study, ketamine suppressed Escherichia coli-induced production of the cytokines interleukin (IL)-6 and tumor necrosis factor alpha (TNF). In other previous studies ketamine improved survival after E. coli inoculation. However, the relationship between cytokines and survival following ketamine treatment is uncertain because no study has examined both cytokines and survival after E. coli inoculation. METHODS: Rats were given E. coli (0.4 x 10(9) colony forming unit (CFU)) at time 0, followed by ketamine (50 mg/kg, n=30) or saline (n=30) at 5 min or 2 h. IL-6 and TNF were measured in serum at 6 h, and mortality was recorded for 7 days. RESULTS: Survival rate with ketamine was 57% (17/30) and was significantly increased compared to saline (27%, 8/30, P=0.01). IL-6 and TNF were lower with ketamine than saline (15,197 +/- 3444 versus 30,725 +/- 4623 pg/ml [mean +/- S.E.M.], P=0.013 and 38.5 +/- 9.5 versus 122.5 +/- 14.0 pg/ml, P=0.001, respectively). With ketamine, IL-6 (but not TNF) concentrations were lower in the survivors (10,900 +/- 776 pg/ml) as compared to the non survivors (P=0.01). IL-6 in ketamine-treated survivors was not different from that in saline-treated survivors. CONCLUSION: We conclude that ketamine given 5 min or 2 h after induction of E. coli sepsis significantly improves survival, possibly by interfering with the inflammatory cascade (as evidenced by attenuation of cytokine production). PMID- 15294411 TI - Emergency drug availability on general paediatric units. AB - Following an incident where intravenous lorazepam was not available on a general paediatric ward we undertook a national survey of emergency drug availability on general paediatric units in the United Kingdom. Drugs chosen were those recommended in the Advanced Paediatric Life Support manual and the British National Formulary for the management of the most common paediatric emergencies. Twelve drugs were chosen covering emergencies in the following systems: cardiovascular (adrenaline (epinephrine), atropine and adenosine); neurological (flumazenil, lorazepam, paraldehyde, phenytoin and mannitol); metabolic (Hypostop Gel and glucagon); analgesia related (naloxone); and respiratory (aminophylline). A thirteenth drug, intravenous salbutamol was included in a reminder letter sent to non-responding units. Questionnaires were sent to 274 units. Replies were received from 242 (88.3%), of whom 20 did not have a general paediatric ward, leaving 222 units (81.0%). Drug availability varied for the different drugs: adrenaline (available on 100% of units), atropine (98.2%), naloxone (96.4%), phenytoin (95.9%), aminophylline (93.2%), paraldehyde (92.3%), mannitol (87.8%), lorazepam (86.9%), glucagon (86.5%), Hypostop Gel (80.6%), adenosine (72.1%) and flumazenil (66.7%). Six of the drugs were classified as first line agents (adrenaline, atropine, adenosine, lorazepam, paraldehyde and aminophylline). Over one in 10 units did not stock two or more of these first line drugs. Consideration needs to be given to the compiling of a consensus based list of drugs that ought to be stocked on all general paediatric units. PMID- 15294412 TI - Early and exclusive use of norepinephrine in septic shock. AB - BACKGROUND: The timing and use of norepinephrine (noradrenaline) (NE) in septic shock remain a matter of controversy. AIM: To study the outcome of septic patients treated with early and exclusive NE. SETTING: Tertiary Intensive Care Unit. PATIENTS: 142 patients with septic shock. INTERVENTION: Exclusive NE infusion within 24 hours of admission to ICU. METHODS AND MAIN RESULTS: Retrospective analysis of data from a unit database identified 142 patients. Their median admission simplified acute physiology score (SAPS II) score was 46 [38, 56] with 98 (69%) receiving mechanical ventilation. Mean arterial pressure (MAP) at the start of NE infusion was 60 [58, 68]mmHg. NE infusion was started at a median of 1.3 [0.3, 5.0]h after ICU admission. Restoration and maintenance of target MAP was achieved initially in all patients and, in 61.3%, within 30 min. The median peak dose of NE was 0.28 [0.14, 0.61]microg/(kg min) and the duration of infusion was 88 [42, 175]h. SAPS II predicted mortality was 40.8%, however, only 34.5% (P = 0.27) died. Among the most severely ill patients (SAPS II score >56) actual mortality was 50.0% versus 74.7% predicted (P = 0.07). CONCLUSIONS: Early and exclusive use of NE in hyperdynamic septic shock achieved a stable MAP >75 mmHg in all patients. Survival compared favorably with that predicted by illness severity scores. PMID- 15294413 TI - Investigation of the Bernoulli model for RNA secondary structures. AB - Within this paper we investigate the Bernoulli model for random secondary structures of ribonucleic acid (RNA) molecules. Assuming that two random bases can form a hydrogen bond with probability p we prove asymptotic equivalents for the averaged number of hairpins and bulges, the averaged loop length, the expected order, the expected number of secondary structures of size n and order k and further parameters all depending on p. In this way we get an insight into the change of shape of a random structure during the process 1 p -->0. Afterwards we compare the computed parameters for random structures in the Bernoulli model to the corresponding quantities for real existing secondary structures of large subunit rRNA molecules found in the database of Wuyts et al. That is how it becomes possible to identify those parameters which behave (almost) randomly and those which do not and thus should be considered as interesting, e.g., with respect to the biological functions or the algorithmic prediction of RNA secondary structures. PMID- 15294414 TI - Genetic code, attributive mappings and stochastic matrices. AB - In this paper we construct three primitive mappings based on three kinds of genetic attribute equivalences. We then apply the mappings and basic addition operation to the universal genetic code to generate three square matrices. We show that these square matrices are stochastic in nature. They demonstrate some fractal similarity properties and resemble the similar properties to the original stochastic matrices. PMID- 15294415 TI - Climate and competition: the effect of moving range boundaries on habitat invasibility. AB - Predictions for climate change include movement of temperature isoclines up to 1000 m/year, and this is supported by recent empirical studies. This paper considers effects of a rapidly changing environment on competitive outcomes between species. The model is formulated as a system of nonlinear partial differential equations in a moving domain. Terms in the equations describe competition interactions and random movement by individuals. Here the critical patch size and travelling wave speed for each species, calculated in the absence of competition and in a stationary habitat, play a role in determining the outcome of the process with competition and in a moving habitat. We demonstrate how habitat movement, coupled with edge effects, can open up a new niche for invaders that would be otherwise excluded. PMID- 15294416 TI - Sensitivity analysis of a nonlinear lumped parameter model of HIV infection dynamics. AB - A formal sensitivity analysis is performed on a delay differential equation model for the viral dynamics of an in vivo HIV infection during protease inhibitor therapy. We present results of both a differential analysis as well as a principle component based analysis and provide evidence that suggests the exact times at which specific parameters have the most influence over the solution. We offer insight into the pairwise mathematical relationships between the productively infected T-cell death rate delta, the viral plasma clearance rate c, and the time delay tau between infection and viral production as they relate to the viral dynamics. The results support the claim that the presence of a nonzero delay has a major impact on the model dynamics. Lastly, we comment upon the inadequacies of an alternative principle component based analysis. PMID- 15294417 TI - On 'Analytical models for the patchy spread of plant disease'. AB - Epidemiologists are interested in using models that incorporate the effects of clustering in the spatial pattern of disease on epidemic dynamics. Bolker (1999, Bull. Math. Biol. 61, 849-874) has developed an approach to study such models based on a moment closure assumption. We show that the assumption works above a threshold initial level of disease that depends on the spatial dispersal of the pathogen. We test an alternative assumption and show that it does not have this limitation. We examine the relation between lattice and continuous-medium implementations of the approach. PMID- 15294418 TI - A history of the study of solid tumour growth: the contribution of mathematical modelling. AB - A miscellany of new strategies, experimental techniques and theoretical approaches are emerging in the ongoing battle against cancer. Nevertheless, as new, ground-breaking discoveries relating to many and diverse areas of cancer research are made, scientists often have recourse to mathematical modelling in order to elucidate and interpret these experimental findings. Indeed, experimentalists and clinicians alike are becoming increasingly aware of the possibilities afforded by mathematical modelling, recognising that current medical techniques and experimental approaches are often unable to distinguish between various possible mechanisms underlying important aspects of tumour development. This short treatise presents a concise history of the study of solid tumour growth, illustrating the development of mathematical approaches from the early decades of the twentieth century to the present time. Most importantly these mathematical investigations are interwoven with the associated experimental work, showing the crucial relationship between experimental and theoretical approaches, which together have moulded our understanding of tumour growth and contributed to current anti-cancer treatments. Thus, a selection of mathematical publications, including the influential theoretical studies by Burton, Greenspan, Liotta et al., McElwain and co-workers, Adam and Maggelakis, and Byrne and co workers are juxtaposed with the seminal experimental findings of Gray et al. on oxygenation and radio-sensitivity, Folkman on angiogenesis, Dorie et al. on cell migration and a wide variety of other crucial discoveries. In this way the development of this field of research through the interactions of these different approaches is illuminated, demonstrating the origins of our current understanding of the disease. PMID- 15294419 TI - Predicting differential responses to structured treatment interruptions during HAART. AB - Highly active antiretroviral therapy (HAART) has been used clinically in various administration schemes for several years. However, due to the development of drug resistance, evolution of viral strains, serious side effects, and poor patient compliance, the combination of drugs used in HAART fails to effectively contain virus long term in a high proportion of patients. Our group and others have suggested a change to the usual regimen of continuous HAART through structured treatment interruptions (STIs). STIs may provide similar clinical benefits as continuous treatment such as reduced viral loads and reestablishment of CD4(+)T cells while allowing patients drug holidays. We explore the use of STIs using a previously published model that accurately represents CD4(+)T-cell counts and viral loads during both untreated HIV-1 infection and HAART therapy. We simulate the effects of different STI regimens including weekly and monthly interruptions together with variations in treatment initiation time. We predict that differential responses to STIs as observed in conflicting clinical trial data are impacted by the duration of the interruption, stage of infection at initiation of treatment, strength of the immune system in suppressing virus, or pre-therapy CD4(+)T-cell count or virus load. Our results indicate that dynamics occurring below the limit of detection (LOD) are influenced by these factors, and contribute to reemergence or suppression of virus during interruptions. Simulations predict that short-term viral suppression with varying interruption strategies does not guarantee long-term clinical benefit. PMID- 15294420 TI - A mathematical model for the dynamics of large membrane deformations of isolated fibroblasts. AB - In this paper we develop and extend a previous model of cell deformations, initially proposed to describe the dynamical behaviour of round-shaped cells such as keratinocytes or leukocytes, in order to take into account cell pseudopodial dynamics with large amplitude membrane deformations such as those observed in fibroblasts. Beyond the simulation (from a quantitative, parametrized model) of the experimentally observed oscillatory cell deformations, a final goal of this work is to underline that a set of common assumptions regarding intracellular actin dynamics and associated cell membrane local motion allows us to describe a wide variety of cell morphologies and protrusive activity. The model proposed describes cell membrane deformations as a consequence of the endogenous cortical actin dynamics where the driving force for large-amplitude cell protrusion is provided by the coupling between F-actin polymerization and contractility of the cortical actomyosin network. Cell membrane movements then result of two competing forces acting on the membrane, namely an intracellular hydrostatic protrusive force counterbalanced by a retraction force exerted by the actin filaments of the cell cortex. Protrusion and retraction forces are moreover modulated by an additional membrane curvature stress. As a first approximation, we start by considering a heterogeneous but stationary distribution of actin along the cell periphery in order to evaluate the possible morphologies that an individual cell might adopt. Then non-stationary actin distributions are considered. The simulated dynamic behaviour of this cytomechanical model not only reproduces the small amplitude rotating waves of deformations of round-shaped cells such as keratinocytes [as proposed in the original model of Alt and Tranquillo (1995, J. Biol. Syst. 3, 905-916)] but is furthermore in very good agreement with the protrusive activity of cells such as fibroblasts, where large amplitude contracting/retracting pseudopods are more or less periodically extended in opposite directions. In addition, the biophysical and biochemical processes taken into account by the cytomechanical model are characterized by well-defined parameters which (for the majority) can be discussed with regard to experimental data obtained in various experimental situations. PMID- 15294421 TI - A computer model of fractal myocardial perfusion heterogeneity to elucidate mechanisms of changes in critical coronary stenosis and hypotension. AB - Critical coronary stenosis (critical CS) alone does not lead to an alteration of fractal dimension (D) under resting conditions in a pig model, indicating undisturbed local myocardial perfusion. If critical CS is combined with hypovolemic anemia the resulting hypotension leads to a significant decline of D. The mechanisms involved in this phenomenon have not yet been elucidated. A computer program was developed enabling calculation of D for normal vascular trees, for single vessel coronary stenosis (CS), and for CS in combination with reduced coronary perfusion pressure (CPP). The values of D obtained by the computer program were compared to those available from an existing animal study to confirm that changes of D can largely be explained by changes of arterial branching pattern simulated by the computer program. Using our computer model, D was 1.15+/-0.06 in normal vascular trees. Third branch critical CS did not alter (1.14+/-0.06; n.s.), whereas critical CS combined with a reduction of CPP to 40 mmHg reduced D (1.07+/-0.03; P < 0.05). These data are comparable to those obtained in the animal study, and therefore show that alterations of vessel diameter and regional blood flow can largely explain changes of fractal dimension during critical CS and hypotension while changes of functional myocardial parameters might play a minor role. PMID- 15294422 TI - MCMC for hidden Markov models incorporating aggregation of states and filtering. AB - This paper is concerned with the statistical analysis of single ion channel records. Single channels are modelled by using hidden Markov models and a combination of Bayesian statistics and Markov chain Monte Carlo methods. The techniques presented here provide a straightforward generalization to those in Rosales et al. (2001, Biophys. J., 80, 1088-1103), allowing to consider constraints imposed by a gating mechanism such as the aggregation of states into classes. This paper also presents an extension that allows to consider correlated background noise and filtered data, extending the scope of the analysis toward real experimental conditions. The methods described here are based on a solid probabilistic basis and are less computationally intensive than alternative Bayesian treatments or frequentist approaches that consider correlated data. PMID- 15294423 TI - Modeling epithelial cell homeostasis: assessing recovery and control mechanisms. AB - Critical to epithelial cell viability is prompt and direct recovery, following a perturbation of cellular conditions. Although a number of transporters are known to be activated by changes in cell volume, cell pH, or cell membrane potential, their importance to cellular homeostasis has been difficult to establish. Moreover, the coordination among such regulated transporters to enhance recovery has received no attention in mathematical models of cellular function. In this paper, a previously developed model of proximal tubule (Weinstein, 1992, Am. J. Physiol. 263, F784-F798), has been approximated by its linearization about a reference condition. This yields a system of differential equations and auxiliary linear equations, which estimate cell volume and composition and transcellular fluxes in response to changes in bath conditions or membrane transport coefficients. Using the singular value decomposition, this system is reduced to a linear dynamical system, which is stable and reproduces the full model behavior in a useful neighborhood of the reference. Cost functions on trajectories formulated in the model variables (e.g., time for cell volume recovery) are translated into cost functions for the dynamical system. When the model is extended by the inclusion of linear dependence of membrane transport coefficients on model variables, the impact of each such controller on the recovery cost can be estimated with the solution of a Lyapunov matrix equation. Alternatively, solution of an algebraic Riccati equation provides the ensemble of controllers that constitute optimal state feedback for the dynamical system. When translated back into the physiological variables, the optimal controller contains some expected components, as well as unanticipated controllers of uncertain significance. This approach provides a means of relating cellular homeostasis to optimization of a dynamical system. PMID- 15294424 TI - On the 28-gon symmetry inherent in the genetic code intertwined with aminoacyl tRNA synthetases--the Lucas series. AB - Despite considerable efforts it has remained unclear what principle governs the selection of the 20 canonical amino acids in the genetic code. Based on a previous study of the 28-gonal and rotational symmetric arrangement of the 20 amino acids in the genetic code, new analyses of the organization of the genetic code system together with their intrinsic relation to the two classes of aminoacyl-tRNA synthetases are reported in this work. A close inspection revealed how the enzymes and the 20 gene-encoded amino acids are intertwined on the polyhedron model. Complementary and cooperative symmetries between class I and class II aminoacyl-tRNA synthetases displayed by a 28-gon organization are discussed, and we found that the two previously suggested evolutionary axes within the genetic code overlap the symmetry axes within the two classes of aminoacyl-tRNA synthetases. Moreover, it has been shown that the side-chain carbon-atom numbers (2, 1, 3, 4 and 7) in the overwhelming majority of the amino acids recognized by each of the two classes of aminoacyl-tRNA synthetases are determined by a mathematical relationship, the Lucas series. A stepwise co evolutionary selection logic of the amino acids is manifested by the amino acid side-chain carbon-atom number balance at '17', when grouping the genetic code doublets in the 28-gon organization. The number '17' equals the sum of the initial five numbers in the Lucas series, which are 2, 1, 3, 4 and 7. PMID- 15294425 TI - Distinct effects of protease and reverse transcriptase inhibition in an immunological model of HIV-1 infection with impulsive drug effects. AB - We present an immunological model that considers the dynamics of CD4+ T cells interacting with free virions, reverse transcriptase inhibiting drugs and protease inhibiting drugs. We divide the T cells into multiple classes and use impulsive differential equations to describe the drug activity. As expected, we find that insufficient dosing of either drug corresponds to high viral load and a large population of infectious T cells. The model further predicts that, in the absence of physiological limits on tolerable drug concentrations, sufficiently frequent dosing with the reverse transcriptase inhibitor alone could theoretically maintain the CD4+ T cell count arbitrarily close to the T cell count in the uninfected immune system. However, for frequent dosing of the protease inhibitor alone, the limiting T cell populations may not be enough to maintain the immune system. Furthermore, frequent dosing of both drugs has the same net effect on the T cell population as frequent dosing of the reverse transcriptase inhibitor only. Thus, the two drug classes can have fundamentally different effects on the long-term dynamics and the reverse transcriptase inhibitor, in particular, plays a crucial role in maintaining the immune system. We also provide estimates for the dosing intervals of each drug that could theoretically sustain the T cell population at a desired level. PMID- 15294426 TI - Simple stochastic fingerprints towards mathematical modelling in biology and medicine. 1. The treatment of coccidiosis. AB - We have developed a classification function that is capable of discriminating between anticoccidial and nonanticoccidial compounds with different structural patterns. For this purpose, we calculated the Markovian electron delocalization negentropies of several compounds. These molecular descriptors, which act as molecular fingerprints, are derived from an electronegativity-weighted stochastic matrix (1Pi). The method attempts to describe the delocalization of electrons with time during the process of molecule formation by considering the 3D environment of the atoms. Accordingly, the entropies of this random process are used as molecular descriptors. The present study involves a stochastic generalization of the original idea described by Kier, which concerned the use of molecular negentropies in QSAR. Linear discriminant analysis allowed us to fit the discriminant function. This function has given rise to a good classification of 82.35% (28 anticoccidials out of 34) and 91.8% of inactive compounds (56/61) in training series. An overall classification of 88.42% (84/95) was achieved. Validation of the model was carried out by means of an external predicting series and this gave a global predictability of 93.1%. Finally, we report the experimental assay (more than 95% of lesion control) of two compounds selected from a large data set through virtual screening. We conclude that the approach described here seems to be a promising 3D-QSAR tool based on the mathematical theory of stochastic processes. PMID- 15294427 TI - A calcium-based phantom bursting model for pancreatic islets. AB - Insulin-secreting beta-cells, located within the pancreatic islets of Langerhans, are excitable cells that produce regular bursts of action potentials when stimulated by glucose. This system has been the focus of mathematical investigation for two decades, spawning an array of mathematical models. Recently, a new class of models has been introduced called 'phantom bursters' [Bertram et al. (2000) Biophys. J. 79, 2880-2892], which accounts for the wide range of burst frequencies exhibited by islets via the interaction of more than one slow process. Here, we describe one implementation of the phantom bursting mechanism in which intracellular Ca2+ controls the oscillations through both direct and indirect negative feedback pathways. We show how the model dynamics can be understood through an extension of the fast/slow analysis that is typically employed for bursting oscillations. From this perspective, the model makes use of multiple degrees of freedom to generate the full range of bursting oscillations exhibited by beta-cells. The model also accounts for a wide range of experimental phenomena, including the ubiquitous triphasic response to the step elevation of glucose and responses to perturbations of internal Ca2+ stores. Although it is not presently a complete model of all beta-cell properties, it demonstrates the design principles that we anticipate will underlie future progress in beta-cell modeling. PMID- 15294428 TI - Expansion and contraction of the cytotoxic T lymphocyte response--an optimal control approach. AB - The kinetics of the cytotoxic T lymphocyte (CTL) response against intracellular pathogens has been found to have many stereotypical features that appear to be programmed early in the infection. We explain these findings here in terms of CTL response kinetics that minimize the probability that a pathological symptom will occur in association with the infection and its eradication. We assume that both the infection and the CTLs contribute to this pathology. We find that contraction kinetics is influenced by the relative pathogenicities of infection and CTLs, as well as on the virulence of the infection and the efficiency of the CTLs, but not by the magnitude of expansion or the dose and duration of infection. Our analysis explains the finding that the duration of the CTL expansion is highly stereotypical, with the maximum expansion of the CTL response dependent on the dose of the infection. Finally, we show that the stereotypical nature of CTL kinetics relies upon stringent regulation of the rates at which CTLs proliferate and die. PMID- 15294429 TI - Thermodynamical interpretation of evolutionary dynamics on a fitness landscape in a evolution reactor, I. AB - A theory for describing evolution as adaptive walks by a finite population with M walkers (M > or = 1) on an anisotropic Mt. Fuji-type fitness landscape is presented, from a thermodynamical point of view. Introducing the 'free fitness' as the sum of a fitness term and an entropy term and 'evolutionary force' as the gradient of free fitness on a fitness coordinate, we demonstrate that the behavior of these theoretical walkers is almost consistent with the thermodynamical schemes. The major conclusions are as follows: (1) an adaptive walk (=evolution) is driven by an evolutionary force in the direction in which free fitness increases; (2) the expectation of the climbing rate obeys an equation analogous to the Einstein relation in Brownian motion; (3) the standard deviation of the climbing rate is a quantity analogous to the mean thermal energy of a particle, kT (x constant). In addition, on the interpretation that the walkers climb the landscape by absorbing 'fitness information' from the surroundings, we succeeded in quantifying the fitness information and formulating a macroscopic scheme from an informational point of view. PMID- 15294430 TI - Genetic code, hamming distance and stochastic matrices. AB - In this paper we use the Gray code representation of the genetic code C=00, U=10, G=11 and A=01 (C pairs with G, A pairs with U) to generate a sequence of genetic code-based matrices. In connection with these code-based matrices, we use the Hamming distance to generate a sequence of numerical matrices. We then further investigate the properties of the numerical matrices and show that they are doubly stochastic and symmetric. We determine the frequency distributions of the Hamming distances, building blocks of the matrices, decomposition and iterations of matrices. We present an explicit decomposition formula for the genetic code based matrix in terms of permutation matrices, which provides a hypercube representation of the genetic code. It is also observed that there is a Hamiltonian cycle in a genetic code-based hypercube. PMID- 15294431 TI - Pairwise alignment of the DNA sequence using hypercomplex number representation. AB - A new set of DNA base-nucleic acid codes and their hypercomplex number representation have been introduced for taking the probability of each nucleotide into full account. A new scoring system has been proposed to suit the hypercomplex number representation of the DNA base-nucleic acid codes and incorporated with the method of dot matrix analysis and various algorithms of sequence alignment. The problem of DNA sequence alignment can be processed in a rather similar way to pairwise alignment of the protein sequence. PMID- 15294432 TI - Concentration estimation via curve fitting: quantification of negative inotropic agents by using a simple mathematical method in guinea pig atria. AB - We created a simple method based on curve fitting in order to assess the concentration of pharmacological agonists or antagonists in the microenvironment of the receptors. We tested our method in electrically driven guinea pig left atria by estimating the concentration of N6-cyclopentyladenosine (CPA; A1 adenosine receptor agonist), acetyl-beta-methylcholine (muscarinic receptor agonist) and verapamil (L-type Ca2+ channel inhibitor) added previously to the atria in known amounts. Our results validated the fitness of the model under specified conditions. In addition, our data suggest a relatively slow elimination of CPA in isolated, practically bloodless guinea pig atrial myocardium. PMID- 15294433 TI - Translocation of metals and its effects in the tomato plants grown on various amendments of tannery waste: evidence for involvement of antioxidants. AB - A field experiment was conducted to study the effect of different amendments of tannery sludge on physiological and biochemical parameters of tomato plant (Lycopersicon esculentum L. Mill). The accumulation of metals (Cr, Fe) in different parts of tomato plants grown on tannery sludge amended soil increased in a concentration and duration-dependent manner. The accumulation of both the metals was found lowest in the fruits of the plant. The statistical analysis of the results showed an increase in chlorophyll and protein contents in lower sludge amendment ratio at all exposures followed by a decrease at highest (100%) sludge amendment ratio. Lipid peroxidation enhanced in both root and leaves of sludge grown plants of tomato at all the sludge amendments and exposure periods, which is evidenced by increased malondialdehyde content, however the maximum increase was found in the roots (43.63%) and leaves (56.66%) of the plant grown on 100% tannery sludge at 60 d, over respective controls. The level of antioxidants, cysteine, non-protein thiol and ascorbic acid increased in the sludge grown plants of tomato to cope up with stress induced by the excess amount of the heavy metals present in the tannery sludge. The maximum increase was found in cysteine content (75.53% in the leaves), non-protein thiol content (92.68% in the roots) and ascorbic acid content (29.66% in the roots) of the plant at 75% tannery sludge after 30 d. The tomato plants were found well adopted for minimizing damage induced by reactive oxygen species, when grown on tannery sludge amendments in the present study. PMID- 15294434 TI - Root growth inhibition and induction of DNA damage in soybean (Glycine max) by chlorobenzenes in contaminated soil. AB - The cytotoxic and genotoxic effects of 1,2,4-trichlorobenzene (TCB), chlorobenzene (CB), and hexachlorobenzene (HCB) on root growth and DNA strand breakage damage of soybean nuclei in the test soil were studied using the comet assay. Results indicated that the root growth was significantly inhibited, and DNA strand breaks and the comet tail in the root tip nuclei were both induced after 48 h exposure with TCB concentrations of 50, 100, 200, 300 microg g(-1) in the soil. DNA strand breakage was more sensitive to the TCB than the root growth. There was a significant dose-response relationship between the TCB exposure and DNA strand breakage in the soybean nuclei. Thus it is possible for DNA strand breakage to be used as a biomarker of soybean exposed to TCB contamination. Significant cytotoxic threshold concentration of the TCB exposure on the root growth inhibition was determined as 61 microg g(-1) in the soil. The toxicity of 100-1,000 microg g(-1) CB and HCB to the soybean seedlings in the soil were not observed after 48 h or longer exposure. PMID- 15294435 TI - Pharmaceuticals in the river Elbe and its tributaries. AB - Medicinal drugs were found to be ubiquitous in the river Elbe, its tributary the river Saale and in other tributaries at their points of entry into the Elbe. The distribution of concentration peaks along the investigated river stretches provides an indication that they are mainly due to the emission of treated waste water from municipal sewage treatment works. This leads to the conclusion that medicinal substances can be regarded as faecal indicators for water pollution caused by human activity. The main substances found in the Elbe in 1998 were diclofenac, ibuprofen and carbamazepine as well as various antibiotics and lipid regulators in the concentration range of <20-140 ng/l. The more thorough investigations carried out in 1999 and 2000 show that in addition to the drugs (phenazone, isopropyl-phenazone and paracetamol) metabolite concentrations contributed significantly to the total concentration of pharmaceuticals in the Elbe. The metamizole metabolites N-acetyl-4-aminoantipyrine (AAA) and N-formyl-4 aminoantipyrine (FAA) were found in concentrations from <20 to 939 ng/l. A multivariate statistical analysis revealed a high correlation in respect of the distribution of persistent substances. The metoprolol distribution throughout the Saale demonstrated that the tributaries cause either an increase (Weisse Elster, Unstrut, Ilm) or a reduction (Wipper, Bode) in the concentration, depending on the respective load of waste water. Wide scale sampling in Saxony during 2002 showed the ubiquitous occurrence of carbamazepine in surface waters. The ecotoxicological effects of this contamination cannot be assessed at present. This is due to the fact that no legal framework in respect of these medicinal drugs for human consumption has been established and therefore little research and no risk assessment has been carried out. Therefore it is urgently necessary to include at least the quantitatively most significant substances in the new assessment concept of the EC White Paper. PMID- 15294436 TI - Effects of four chlorobenzenes on serum sex steroids and hepatic microsome enzyme activities in crucian carp, Carassius auratus. AB - Four chlorobenzenes (chlorobenzene, 1,3-dichlorobenzene, 1,4-dichlorobenzene, p chloro-methylbenzene) were administrated to the crucian carps (Carassius auratus) by peritoneal injections in the laboratory for 30 days. Serum testosterone and 17 beta-estradiol concentrations were detected using radioimmunology assay (RIA), and the activities of two hepatic microsome enzymes, glutathione s-transferase (GST) and UDP-glucuronosyltransferase (UDPGT), were measured using the modified methods as described by Habig and Owens. Results showed that the four chlorobenzenes caused significant increases in serum testosterone concentration in the crucian carps (P < 0.05) compared to the controls, but they caused no significant effect on 17 beta-estradiol level. All test chemicals caused a change in hepatic GST activity in crucian carps, with significant increases in enzyme activity (P < 0.05). Chlorobenzene, 1,3-dichlorobenzene and p-chloro methylbenzene resulted in a marked inhibition to UDPGT activity in crucian carp (P < 0.05) except 1,4-dichlorobenzene. The changes in hepatic microsome enzyme activities may have resulted in the alterations of serum sex steroids levels in the crucian carps. The results indicated that these four chlorobenzenes may result in the changes of endocrine functions and may affect the reproductive success of this and other species. PMID- 15294437 TI - Monitoring of toxicity during degradation of selected pesticides using ionizing radiation. AB - The optimization of experimental conditions for radiolytic removal of organic pollutants from water and waste with the use of ionizing radiation via controlling the concentration of target compound(s) requires also monitoring the toxicity changes during the process. Commonly used herbicides 2,4-D and dicamba were shown to increase toxicity measured with the Microtox test at low irradiation doses resulting from formation of more toxic transient products, which can be decomposed at larger doses. The changes of toxicity were examined with respect to dose magnitude and the presence of commonly occurring scavengers of radiation. PMID- 15294438 TI - Juvenile sea bass biotransformation, genotoxic and endocrine responses to beta naphthoflavone, 4-nonylphenol and 17 beta-estradiol individual and combined exposures. AB - Juvenile sea bass, Dicentrarchus labrax L., were exposed during 2, 4, 8, and 24 h to 0.9 microM beta-naphthoflavone (BNF), 131 nM 17 beta-estradiol (E(2)), 4.05 microM 4-nonylphenol (NP), as well as to BNF combined either to E(2) or NP (maintaining the previous concentrations). Liver cytochrome P450 content (P450), ethoxyresorufin-O-deethylase (EROD), and glutathione S-transferase (GST) activities were measured in order to evaluate biotransformation responses. Genotoxicity was assessed as erythrocytic nuclear abnormalities (ENA) frequency. The effects on endocrine function were evaluated as plasma cortisol and glucose. Cortisol was not affected by xeno/estrogens tested, either in single exposure or mixed with BNF. Nevertheless, the intermediary metabolism was affected since glucose concentration increased after 4 h exposure to E(2), and after all BNF+NP exposure lengths. Moreover, a synergism between BNF and NP was thoroughly demonstrated, whereas a sporadic antagonistic interaction was found at 4 h BNF + E(2) exposure. Liver EROD and GST activities were not significantly altered by single E(2) or NP exposure. However, both compounds were able to induce EROD activity in the presence of BNF. NP single exposure was able to significantly increase liver P450 content, while its mixture with BNF displayed an antagonistic interference. Considering the xeno/estrogens single exposures, only NP induced an ENA increase; however, both mixtures (BNF + E(2) and BNF + NP) displayed genotoxic effects. Fish responses to mixtures of xenobiotics are complex and the type of interaction (synergism/potentiation or antagonism) in a particular mixture can vary with the evaluated biological response. PMID- 15294439 TI - Bioaccumulation of PAHs from creosote-contaminated sediment in a laboratory exposed freshwater oligochaete, Lumbriculus variegatus. AB - The oligochaete, Lumbriculus variegatus, was used for a bioaccumulation assay in the creosote-contaminated sediment of Lake Jamsanvesi in a 28-day experiment. The PAH concentrations of the whole body tissue of worms, sediments and water samples were determinated by GC-MS. Chemical analyses showed that benzo(k)fluoranthene, anthracene and fluorene were the main PAH compounds present in the tissue of oligochaetes, just as in the sediment. The biota-sediment accumulation factors (BSAFs) of the individual PAHs varied from 1.2 to 5.7. It is concluded that oligochaetes have a marked ability to accumulate and retain PAHs from creosote contaminated sediment. PMID- 15294440 TI - Prophylactic role of phycocyanin: a study of oxalate mediated renal cell injury. AB - Oxalate induced renal calculi formation and the associated renal injury is thought to be caused by free radical mediated mechanisms. An in vivo model was used to investigate the effect of phycocyanin (from Spirulina platensis), a known antioxidant, against calcium oxalate urolithiasis. Male Wistar rats were divided into four groups. Hyperoxaluria was induced in two of these groups by intraperitoneal infusion of sodium oxalate (70 mg/kg) and a pretreatment of phycocyanin (100 mg/kg) as a single oral dosage was given, 1h prior to sodium oxalate infusion. An untreated control and drug control (phycocyanin alone) were also included in the study. We observed that phycocyanin significantly controlled the early biochemical changes in calcium oxalate stone formation. The antiurolithic nature of the drug was evaluated by the assessment of urinary risk factors and light microscopic observation of urinary crystals. Renal tubular damage as divulged by urinary marker enzymes (alkaline phosphatase, acid phosphatase and gamma-glutamyl transferase) and histopathological observations such as decreased tubulointerstitial, tubular dilatation and mononuclear inflammatory cells, indicated that renal damage was minimised in drug-pretreated group. Oxalate levels (P < 0.001) and lipid peroxidation (P < 0.001) in kidney tissue were significantly controlled by drug pretreatment, suggesting the ability of phycocyanin to quench the free radicals, thereby preventing the lipid peroxidation mediated tissue damage and oxalate entry. This accounts for the prevention of CaOx stones. Thus, the present analysis revealed the antioxidant and antiurolithic potential of phycocyanin thereby projecting it as a promising therapeutic agent against renal cell injury associated kidney stone formation. PMID- 15294441 TI - Induction of renal oxidative stress and cell proliferation response by ferric nitrilotriacetate (Fe-NTA): diminution by soy isoflavones. AB - Ferric nitrilotriacetate (Fe-NTA) is a known potent nephrotoxic agent. In this communication, we report the chemopreventive effect of soy isoflavones on renal oxidative stress, toxicity and cell proliferation response in Wistar rats. Fe-NTA (9 mg Fe/kg body weight, intraperitoneally) enhances gamma-glutamyl transpeptidase, renal lipid peroxidation, xanthine oxidase and hydrogen peroxide (H2O2) generation with reduction in renal glutathione content, antioxidant enzymes, viz., glutathione peroxidase, glutathione reductase, catalase, glucose-6 phosphate dehydrogenase and phase-II metabolising enzymes such as glutathione-S transferase and quinone reductase. Fe-NTA treatment also induced tumor promotion markers, viz., ornithine decarboxylase (ODC) activity and thymidine [3H] incorporation into renal DNA. A sharp elevation in the levels of blood urea nitrogen and serum creatinine has also been observed. Treatment of rats orally with soy isoflavones (5 mg/kg body weight and 10 mg/kg body weight) resulted in significant decreases in gamma-glutamyl transpeptidase, lipid peroxidation, xanthine oxidase, H2O2 generation, blood urea nitrogen, serum creatinine, renal ODC activity and DNA synthesis (P < 0.001). Renal glutathione content (P < 0.01), glutathione metabolizing enzymes (P < 0.001) and antioxidant enzymes were also returned to normal levels (P < 0.001). Thus, our data suggest that soy isoflavones may be used as an effective chemopreventive agent against Fe-NTA mediated renal oxidative stress, toxicity and cell proliferation response in Wistar rats. PMID- 15294442 TI - G protein activation by G protein coupled receptors: ternary complex formation or catalyzed reaction? AB - G protein coupled receptors catalyze the GDP/GTP exchange on G proteins, thereby activating them. The ternary complex model, designed to describe agonist binding in the absence of GTP, is often extended to G protein activation. This is logically unsatisfactory as the ternary complex does not accumulate when G proteins are activated by GTP. Extended models taking into account nucleotide binding exist, but fail to explain catalytic G protein activation. This review puts forward an enzymatic model of G protein activation and compares its predictions with the ternary complex model and with observed receptor phenomenon. This alternative model does not merely provide a new set of formulae but leads to a new philosophical outlook and more readily accommodates experimental observations. The ternary complex model implies that, HRG being responsible for efficient G protein activation, it should be as stable as possible. In contrast, the enzyme model suggests that although a limited stabilization of HRG facilitates GDP release, HRG should not be "too stable" as this might trap the G protein in an inactive state and actually hinder G protein activation. The two models also differ completely in the definition of the receptor "active state": the ternary complex model implies that the active state corresponds to a single active receptor conformation (HRG); in contrast, the catalytic model predicts that the active receptor state is mobile, switching smoothly through various conformations with high and low affinities for agonists (HR, HRG, HRGGDP, HRGGTP, etc.). PMID- 15294443 TI - Potassium-specific effects of levosimendan on heart mitochondria. AB - In this study, we evaluated levosimendan, a new drug developed for the treatment of acute and decompensated heart failure, as a potential activator of ATP sensitive potassium flux to the matrix of cardiac mitochondria. We estimated the KATP channel openers-induced increase in mitochondrial inner membrane permeability for potassium by registering changes in membrane potential of heart mitochondria, oxidizing endogenous substrates. We compared the effect of levosimendan with the effects of the known KATP channel openers diazoxide and pinacidil. Levosimendan (1 microM) accelerated potassium-specific DeltaPsi decrease by 0.15%/s, whereas 50 microM diazoxide by 0.10%/s, and 50 microM pinacidil by 0.08%/s, respectively. These results were confirmed by swelling experiments of non-respiring mitochondria in potassium nitrate medium. We found that levosimendan with an EC50 of 0.83 +/- 0.24 microM activates potassium flux to the mitochondrial matrix. This effect is discussed as a possible explanation of the anti-ischemic action of levosimendan. PMID- 15294444 TI - Inhibition of plasma lipid peroxidation by anti-atherogenic antioxidant BO-653, 2,3-dihydro-5-hydroxy-4,6-di-tert-butyl-2,2-dipentylbenzofuran. AB - BO-653, 2,3-dihydro-5-hydroxy-4,6-di-tert-butyl-2,2-dipentylbenzofuran, is a synthetic antioxidant which is now being developed as an anti-atherogenic drug. The antioxidant action of BO-653 against lipid peroxidation in rat plasma was studied and compared with its analogue BO-653M, 2,3-dihydro-5-hydroxy-4,6-di methyl-2,2-dipentylbenzofuran, and vitamin E. BO-653 was readily incorporated into plasma by oral administration and it inhibited plasma lipid peroxidation more efficiently than vitamin E independent of the presence or absence of vitamin C. On the other hand, its analogue BO-653M having two methyl substituents in place of tert-butyl groups of BO-653 did not inhibit the lipid peroxidation in plasma as efficiently as BO-653, demonstrating clearly that the tert-butyl groups at the ortho-position play a key role in determining the antioxidant efficacy. PMID- 15294445 TI - Multiple structural features of steroids mediate subtype-selective effects on human alpha4beta3delta GABAA receptors. AB - Neurosteroids have been shown to mediate some of their physiological effects via a modulatory site on type A inhibitory gamma-aminobutyric acid (GABAA) receptors. In particular, recent evidence has implicated selective potentiation of the delta subunit of GABAA receptors as an important mediator of in vitro and in vivo neurosteroid activity. However, this has been demonstrated for only a very small number of steroids, so both the generality of this finding, and the structural features of steroids which mediate functional delta-selectivity, are unclear. We have used a potentiometric assay based on fluorescence resonance energy transfer to measure GABA-activated responses in L(tk-) cells stably transfected with human GABAA receptor alpha4beta3delta and alpha4beta3gamma2 receptor subtypes. A set of 28 steroids were evaluated on these subtypes to characterise their functional potency and efficacy in modulating GABA responses. For most compounds there was a clear separation of their efficacy profiles between the receptor subtypes, with a substantially larger maximal response at the alpha4beta3delta receptor. 5beta Pregnan-3beta-ol-20-one, 5beta-pregnane-3alpha,20beta-diol and 5beta-pregnane 3alpha,17alpha-diol-11,20-dione showed particularly high efficacy for alpha4beta3delta. No compounds were identified that simply inhibited responses at delta-containing receptors. However, 5beta-pregnane-3alpha,17alpha,20beta-triol, prednisolone 21-acetate, 4-pregnene-17alpha,20alpha-diol-3-one-20-acetate, 4 pregnen-20alpha-ol-3-one, and 5beta-pregnane-3alpha,17alpha,21-triol-20-one inhibited, though did not abolish, GABA responses at the alpha4beta3gamma2 subtype, while evoking modest-amplitude potentiation of alpha4beta3delta responses. Molecular modelling on this compound series using principal components analysis indicates that several structural features of steroids underlie their relative functional selectivity for potentiation of delta-containing GABAA receptors. PMID- 15294446 TI - Blockade of calcium entry in smooth muscle cells by the antidepressant imipramine. AB - The present study was designed to evaluate the effects of antidepressants on smooth muscle contractile activity. In rat aortic rings, the antidepressants imipramine, mianserin and sertraline provoked concentration-dependent inhibitions of the mechanical responses evoked by K+ (30 mM) depolarization. These myorelaxant effects were not modified by the presence of glibenclamide or 80 mM K+ in the bathing medium. Moreover, the vasodilator properties of imipramine were not affected by atropine, phentolamine and pyrilamine. Radioisotopic experiments indicated that imipramine failed to enhance 86Rb outflow from prelabelled and perifused aortic rings whilst counteracting the increase in 45Ca outflow provoked by a rise in the extracellular K+ concentration. Simultaneous measurements of contractile activity and fura-2 fluorescence revealed that, in aortic rings, imipramine reduced the mechanical and fluorimetric response to K+ challenge. In A7r5 smooth muscle cells, whole cell recordings further demonstrated that imipramine inhibited the inward Ca2+ current. Under different experimental conditions, the ionic and relaxation responses to the antidepressants were reminiscent of those mediated by the Ca2+ entry blocker verapamil. Lastly, it should be pointed out that imipramine exhibited a myorelaxant effect of similar amplitude on rat aorta and on rat distal colon. All together, these findings suggest that the myorelaxant properties of imipramine, and probably also setraline and mianserin, could result from their capacity to inhibit the voltage sensitive Ca2+ channels. PMID- 15294447 TI - Reversal of P-glycoprotein-mediated multidrug resistance by Alisol B 23-acetate. AB - Herbal drugs were screened for their activity in reversing multidrug resistance (MDR) in P-glycoprotein (P-gp) over-expressing cancer cells. Through bio-assay guided fractionation an active compound was isolated from Rhizoma Alismatis, the underground part of Alisma orientale and the chemical structure of the isolate compound was confirmed by HPLC, LC-MS and NMR as Alisol B 23-acetate (ABA). ABA restored the sensitivity of MDR cell lines HepG2-DR and K562-DR to anti-tumor agents that have different modes of action but are all P-gp substrates. It restored the activity of vinblastine, a P-gp substrate, in causing G2/M arrest in MDR cells. In a dose-dependent manner, ABA increased doxorubicin accumulation and slowed down the efflux of rhodamin-123 from MDR cells. ABA inhibited the photoaffinity labeling of P-gp by [125I]iodoarylazidoprazosin and stimulated the ATPase activity of P-gp in a concentration-dependent manner, suggesting that it could be a transporter substrate for P-gp. In addition, ABA was also a partial non-competitive inhibitor of P-gp when verapamil was used as a substrate. Our results suggest that ABA may be a potential MDR reversal agent and could serve as a lead compound in the development of novel drugs. PMID- 15294448 TI - Cytotoxicity, DNA strand breakage and DNA-protein crosslinking by a novel transplatinum compound in human A2780 ovarian and MCF-7 breast carcinoma cells. AB - Cisplatin, cis-[PtCl2(NH3)2], is commonly utilized in various combination chemotherapy protocols for the treatment of both ovarian and breast cancer while the corresponding trans isomer is therapeutically inactive. This work describes efforts to elucidate the cellular mechanism of action of a novel trans-platinum compound, trans-(dichloroamminethiazole)platinum(II) (ATZ), which demonstrates antiproliferative and cytotoxic effects against both MCF-7 human breast and A2780 human ovarian carcinoma cells in culture. A2780 cells were approximately twofold more sensitive to ATZ than MCF-7 cells in both cell growth and clonogenic survival assays. Dye exclusion studies revealed a 50-70% loss in cell viability within the first 12 h of drug treatment in both cell lines. This initial wave of cell death was succeeded by a prolonged interval of growth arrest during which a small fraction of apoptotic cells was detected. Binding of ATZ to DNA, as estimated by atomic absorption spectroscopy, was similar for the two cell lines and was almost completely reversed 24 h after drug removal. ATZ also induced DNA strand breakage as well as DNA-protein crosslinking during the initial 12 h period when the bulk of cell death was evident. However, neither the extent of DNA strand breakage nor that of DNA protein crosslinking was sufficient to explain the different drug sensitivity in the two cell lines. At 24 and 48 h after exposure of MCF-7 cells to high concentrations of ATZ, the formation of DNA topoisomerase I complexes is detected, coincident with a high degree of apoptosis. These studies suggest that ATZ has the capacity to interfere with topoisomerase I in the tumor cell, a function not evident in cis-platinum-based drugs. PMID- 15294449 TI - Differential inhibitor sensitivity between human recombinant and native photoreceptor cGMP-phosphodiesterases (PDE6s). AB - Human photoreceptor cGMP-phosphodiesterases (PDE6s) are important reagents in PDE inhibitor discovery. However, recombinant human PDE6s have not been expressed, and isolation of native human PDE6s is highly difficult. In this study, the catalytic subunit(s) of human rod and cone PDE6s (PDE6alphabeta and PDE6alpha', respectively) were co-expressed or expressed separately as catalytically active enzymes. Sildenafil inhibited both the recombinant PDE6s in a dose-dependent manner with Ki values of 94 and 98 nM, respectively. These Ki values were four fold higher than that (25 nM) of a human native PDE6 preparation. Similarly, 3 isobutyl-1-methylxanthine (IBMX)'s Ki values for the recombinant PDE6s were five- to eight-fold higher than that of the native enzyme. However, E4021 and zaprinast exhibited much (30-80-fold) lower potencies for the recombinant PDE6s than for the native enzyme. Additional PDE5 inhibitors representing other structural classes and possessing different selectivity against native PDE6 also showed different potencies against the recombinant and native PDE6s. In particular, one class of xanthine analogues exhibited significantly (5-15-fold) higher potencies for the recombinant PDE6s than for the native enzyme. Our data demonstrates that the recombinant and native PDE6s exhibit differential sensitivity to inhibitors, and cautions the use of recombinant catalytic subunits of PDE6 in drug discovery or in structural/functional studies. PMID- 15294450 TI - Calcium ionophoretic and apoptotic effects of ferutinin in the human Jurkat T cell line. AB - We have investigated the ionophoretic and apoptotic properties of the daucane sesquiterpene ferutinin and three related compounds, ferutidin, 2-alpha hydroxyferutidin and teferin, all isolated from various species of plants from the genus Ferula. Ferutinin induced a biphasic elevation of intracellular Ca2+ in the leukemia T-cell line, Jurkat. First, a rapid calcium peak was observed and inhibited by BAPTA-AM. This initial calcium mobilization was followed by a sustained elevation, mediated by the entry of extracellular calcium through L type calcium channels and sensitive to inhibition by EGTA. Moreover, ferutinin induced apoptosis in Jurkat cells, and this event was preceded, in a cyclosporine A sensitive manner, by a loss of mitochondrial transmembrane potential (DeltaPsim) and by an increase in intracellular reactive oxygen species. Ferutinin-induced DNA fragmentation was mediated by a caspase-3-dependent pathway, and was initiated independently of any specific phase of the cell cycle. The evaluation of ferutinin analogs in calcium mobilization and apoptosis assays showed strict structure-activity relationships, with p-hydroxylation of the benzoyl moiety being requested for activity. PMID- 15294451 TI - Endogenous cystinyl aminopeptidase in Chinese hamster ovary cells: characterization by [125I]Ang IV binding and catalytic activity. AB - The angiotensin II C-terminal hexapeptide fragment angiotensin IV (Ang IV) exerts central and cardiovascular effects. Cystinyl aminopeptidase (EC 3.4.11.3), a membrane-associated zinc-dependent metallopeptidase of the M1 family, has recently been found to display high affinity for Ang IV and it was proposed to represent the AT4 receptor. We present evidence for the presence of endogenous cystinyl aminopeptidase in membranes from Chinese hamster ovary (CHO-K1) cells by binding studies with [125I]Ang IV and by measuring the cleavage of L-leucine-p nitroanilide. The equilibrium dissociation constant of [125I]Ang IV in saturation binding studies (KD= 0.90 nM) was similar to the value (KD= 0.70 nM) calculated from the association and dissociation rates. Binding was displaced with high potency by the "AT4 receptor" ligands (Ang IV > divalinal1-Ang IV approximately LVV-hemorphin-7 approximately LVV-hemorphin-6 > Ang (3-7) > Ang III > Ang (4-8)) but not by AT1/AT2 receptor antagonists. Enzymatic activity in CHO-K1 cell membranes was competitively inhibited upto 94% by Ang IV and other "AT4 receptor" ligands (Ang IV > Ang III approximately divalinal1-Ang IV approximately Ang (3-7) approximately LVV-hemorphin-7 > Ang (4-8) approximately LVV-hemorphin-6). High affinity binding of [125I]Ang IV required the presence of metal chelators and the ligands such as Ang IV and LVV-hemorphin-7 displayed higher potency in the binding studies as in the enzyme assay. This difference in potency varied from one peptide to another. These pharmacological properties match those previously reported for the recombinantly-expressed human cystinyl aminopeptidase in embryonal kidney cells. PMID- 15294452 TI - Synergistic modulation of cystinyl aminopeptidase by divalent cation chelators. AB - Membranes of Chinese hamster ovary (CHO-K1) cells were used to study the opposite modulation of enzyme activity and [125I]Ang IV binding to cystinyl aminopeptidase (EC 3.4.11.3) by divalent cation chelators. Whereas ethylene diamine tetraacetic acid (EDTA) or ethylene glycol-bis(2-aminoethylether)-N,N,N',N'-tetraacetic acid (EGTA) alone only slightly affected the enzyme activity, 1,10-phenanthrolin (1,10 PHE) produced a complete and concentration-dependent inhibition. Interestingly EDTA (> or =0.05 mM) or EGTA (> or =0.15 mM) enhanced the inhibitory effect of 1,10-PHE. Two-site analysis of the corresponding inhibition curves revealed that EDTA and EGTA converted enzymes with low sensitivity towards 1,10-PHE into enzymes with high sensitivity. The combined inhibition by EDTA (0.1 mM) and 1,10 PHE (0.1 mM) could be prevented and reversed by addition of Zn2+ (at about 0.04 0.1 mM). In contrast, specific binding of [125I]Ang IV was enhanced in the presence of 1,10-PHE. Binding was only slightly affected by EDTA or EGTA alone. Furthermore, the stimulatory effect of 1,10-PHE was potentiated by EDTA (> or =0.05 mM) as well as EGTA (> or =0.15 mM). In the presence of EDTA (0.1 mM) and 1,10-PHE (0.1 mM), specific [125I]Ang IV binding was completely inhibited by Zn2+ (IC50= 39.7 +/- 6.2 microM). The present data show that divalent cations such as Zn2+ are essential for the enzyme activity of cystinyl aminopeptidase and inhibitory for [125I]Ang IV binding. Modulation of the effects of 1,10-PHE by other chelators such as EDTA or EGTA, suggests that, in addition to the binding site for zinc in the catalytic site, cystinyl aminopeptidase also bears a regulatory divalent cation binding site. PMID- 15294453 TI - Taurine block of cloned ATP-sensitive K+ channels with different sulfonylurea receptor subunits expressed in Xenopus laevis oocytes. AB - Taurine has been found to inhibit ATP-sensitive K+ (KATP) channels in rat pancreatic beta-cells [Park et al., Biochem Pharmacol 2004;67:1089-1096] which could be due to its interaction with a benzamido-binding site on SUR1. In present study, we further evaluated the mechanism of taurine action on the KATP-channel inhibition, using cloned KATP-channels with different types of SUR subunit expressed in Xenopus laevis oocytes. The oocytes were coinjected with Kir6.2 mRNA, and mRNA encoding SUR1, SUR2A or SUR2B. Macroscopic currents were recorded from giant excised inside-out patches. The binding of glibenclamide to SUR1 was assessed by using a glibenclamide-fluorescent probe. Intracellular taurine inhibited all three types of KATP-channels to a similar extent. They were fit to the Hill equation, showing IC50 of 11.0 mM for Kir6.2/SUR1, 10.9 mM for Kir6.2/SUR2A, and 9.0 mM for Kir6.2/SUR2B currents. Taurine at the concentration of 10 mM enhanced the high-affinity bindings of glibenclamide and repaglinide on all types of SUR, whereas the low-affinity binding on Kir6.2 was not affected. The intensity of glibenclamide fluorescence was higher in the plasma membrane of taurine-pretreated oocytes. The high-affinity binding of tolbutamide or gliclazide on SUR was not modified by taurine. These results suggest that the taurine inhibition of KATP-channels is mediated by an interaction with the site on SUR where the benzamido group is bound. Therefore, intracellular concentrations of taurine in different tissues may be more important in determining taurine modulation of the KATP-channel rather than distinct types of SUR subunit. PMID- 15294454 TI - Resistance to purine and pyrimidine nucleoside and nucleobase analogs by the human MDR1 transfected murine leukemia cell line L1210/VMDRC.06. AB - Overexpression of human MDR1 P-glycoprotein [Pgp] is associated with cellular resistance to bulky amphipathic drugs, such as taxol, anthracyclines, vinca alkaloids, and epipodophyllotoxins by actively effluxing drugs from cells. We have found that human MDR1 transfected murine L1210/VMDRC.06 leukemia cells exhibit relatively large amounts of Pgp and high levels of resistance to 6 mercaptopurine [6-MP] and other purine and pyrimidine nucleobase and nucleoside analogs. L1210/VMDRC.06 cells accumulated 6-MP as the nucleotide in vitro at only about one-third of that formed by parental L1210 cells in normal medium; however, under conditions of ATP-depletion, the amount of 6-MP nucleotide formed was essentially the same in both cell lines. The findings support active efflux of 6 MP in L1210 cells, suggesting involvement of Pgp in 6-MP resistance even though it is generally believed that Pgp does not transport such agents. The resistance pattern observed in L1210/VMDRC.06 cells was not duplicated in P388/VMDRC.04 leukemia cells transfected with the same MDR1 cDNA, even though a similar amount of Pgp was present in both cell lines. Immunofluorescent staining of surface membrane Pgp showed that L1210/VMDRC.06 cells contained at least three-fold more surface Pgp than P388/VMDRC.04, implying that P388/VMDRC.04 cells are unable to actively efflux 6-MP and other antimetabolites as effectively as L1210/VMDRC.06, because of significantly lower membrane Pgp. The findings suggest that the exceedingly large concentration of overexpressed Pgp in the surface membrane of L1210/MDRC.06 cells is responsible for resistance to 6-MP and other purine and pyrimidine analogs, even though these agents usually are not considered to be substrates for Pgp. PMID- 15294455 TI - Role of c-myc protein in hormone refractory prostate carcinoma: cellular response to paclitaxel. AB - Amplification of the c-MYC proto-oncogene is a frequent alteration in hormone refractory prostate carcinomas (HRPC). In an attempt to investigate the role of c myc in the cellular response to paclitaxel (PTX), we used two HRPC cell lines, DU145 and PC3, characterised by different levels of the protein and by different behaviour in response to taxane. In both cell lines, PTX-induced cell death was a caspase-mediated apoptosis. In DU145 cells, PTX induced an early apoptotic response associated with upregulation of c-myc restricted to the G2/M cell population. This event appeared delayed in the presence of c-myc antisense (AS-c myc), suggesting an upstream regulation of the protein expression. In addition, the antisense approach provided evidence of an involvement of c-myc in the apoptotic response to the taxane. In contrast, in PC3 cells, the overexpressed c myc was not modulated by drug-treatment and the addition of AS-c-myc did not affect the cell growth inhibition of PTX. In both cell lines, PTX-induced c-myc phosphorylation was concomitant with the mitotic arrest and not related to the modulation of the activation state of AKT and MAPK kinases. Our data indicate that the cellular response to PTX of HRPC cells can involve c-myc and suggest that its pro-apoptotic role is affected by the genetic background, thus supporting a complex and differentiated HRPC cell response to taxanes. PMID- 15294456 TI - Pharmacological and behavioral properties of A-349821, a selective and potent human histamine H3 receptor antagonist. AB - Histamine H3 receptors regulate the release of a variety of central neurotransmitters involved in cognitive processes. A-349821 ((4'-(3-((R,R)2,5 dimethyl-pyrrolidin-1-yl)-propoxy)-biphenyl-4-yl)-morpholin-4-yl-methanone) is a novel, non-imidazole H3 receptor ligand, displaying high affinity for recombinant rat and human H3 receptors, with pKi values of 9.4 and 8.8, respectively, and high selectivity for the H3 receptor versus H1, H2, and H4 histamine receptors. A 349821 is a potent H3 receptor antagonist in a variety of models using recombinant human and rat receptors, reversing agonist induced changes in cyclic AMP formation (pKb= 8.2 and pKb= 8.1, respectively), [35S]-GTPgammaS binding (pKb= 9.3 and pKb= 8.6, respectively) and calcium levels (human pKb= 8.3). In native systems, A-349821 competitively reversed agonist induced inhibition of electric field stimulated guinea-pig ileum (pA2= 9.5) and histamine-mediated inhibition of [3H]-histamine release from rat brain cortical synaptosomes (pKb= 9.2). Additionally, A-349821 inhibited constitutive GTPgammaS binding at both rat and human H3 receptors with respective pEC50 values of 9.1 and 8.6, demonstrating potent inverse agonist properties. In behavioral studies, A-349821 (0.4 mg/kg-4 mg/kg) potently blocked (R)-alpha-methylhistamine-induced dipsogenia in mice. The compound also enhanced cognitive activity in a five-trial inhibitory avoidance model in spontaneously hypertensive rat (SHR) pups at doses of 1-10mg/kg, with the 1mg/kg dose showing comparable efficacy to a fully efficacious dose of ciproxifan (3mg/kg). These doses of A-349821 were without effect on spontaneous locomotor activity. Thus, A-349821 is a novel, selective non-imidazole H3 antagonist/inverse agonist with balanced high potency across species and favorable cognition enhancing effects in rats. PMID- 15294457 TI - Fenproporex N-dealkylation to amphetamine--enantioselective in vitro studies in human liver microsomes as well as enantioselective in vivo studies in Wistar and Dark Agouti rats. AB - Fenproporex (FP) is known to be N-dealkylated to R(-)-amphetamine (AM) and S(+) amphetamine. Involvement of the polymorphic cytochrome P450 (CYP) isoform CYP2D6 in metabolism of such amphetamine precursors is discussed controversially in literature. In this study, the human hepatic CYPs involved in FP dealkylation were identified using recombinant CYPs and human liver microsomes (HLM). These studies revealed that not only CYP2D6 but also CYP1A2, CYP2B6 and CYP3A4 catalyzed this metabolic reaction for both enantiomers with slight preference for the S(+)-enantiomer. Formation of amphetamine was not significantly changed by quinidine and was not different in poor metabolizer HLM compared to pooled HLM. As in vivo experiments, blood levels of R(-)-amphetamine and S(+)-amphetamine formed after administration of FP were determined in female Dark Agouti rats (fDA), a model of the human CYP2D6 poor metabolizer phenotype (PM), male Dark Agouti rats (mDA), an intermediate model, and in male Wistar rats (WI), a model of the human CYP2D6 extensive metabolizer phenotype. Analysis of the plasma samples showed that fDA exhibited significantly higher plasma levels of both amphetamine enantiomers compared to those of WI. Corresponding plasma levels in mDA were between those in fDA and WI. Furthermore, pretreatment of WI with the CYP2D inhibitor quinine resulted in significantly higher amphetamine plasma levels, which did not significantly differ from those in fDA. The in vivo studies suggested that CYP2D6 is not crucial to the N-dealkylation but to another metabolic step, most probably to the ring hydroxylation. Further studies are necessary for elucidating the role of CYP2D6 in FP hydroxylation. PMID- 15294458 TI - S-oxygenation of the thioether organophosphate insecticides phorate and disulfoton by human lung flavin-containing monooxygenase 2. AB - Phorate and disulfoton are organophosphate insecticides containing three oxidizable sulfurs, including a thioether. Previous studies have shown that only the thioether is oxygenated by flavin-containing monooxygenase (FMO) and the sole product is the sulfoxide with no oxygenation to the sulfone. The major FMO in lung of most mammals, including non-human primates, is FMO2. The FMO2*2 allele, found in all Caucasians and Asians genotyped to date, codes for a truncated, non functional, protein (FMO2.2A). Twenty-six percent of individuals of African descent and 5% of Hispanics have the FMO2*1 allele, coding for full-length, functional protein (FMO2.1). We have here demonstrated that the thioether containing organophosphate insecticides, phorate and disulfoton, are substrates for expressed human FMO2.1 with Km of 57 and 32 microM, respectively. LC/MS confirmed the addition of oxygen and formation of a single polar metabolite for each chemical. MS/MS analysis confirmed the metabolites to be the respective sulfoxides. Co-incubations with glutathione did not reduce yield, suggesting they are not highly electrophilic. As the sulfoxide of phorate is a markedly less effective acetylcholinesterase inhibitor than the cytochrome P450 metabolites (oxon, oxon sulfoxide or oxon sulfone), humans possessing the FMO2*1 allele may be more resistant to organophosphate-mediated toxicity when pulmonary metabolism is an important route of exposure or disposition. PMID- 15294459 TI - Retention of basic science knowledge: a comparison between body system-based and clinical presentation curricula. AB - BACKGROUND: When the University of Calgary implemented the clinical presentation (CP) curriculum in 1994, it was prospectively decided to administer the National Board of Medical Examiner's Comprehensive Basic Science Exam (CBSE) as a measure of students' basic science knowledge retention. PURPOSE: The exam performance from 2 classes (1995, 1996) of the previous system-based (SB) curriculum was compared to exam performance of 2 classes (2000, 2002) of the CP curriculum. METHODS: Data analyses employed 2 statistical models (covariate multiple linear regression and hierarchical mixed effects), and effect sizes were computed. RESULTS: Differences between CBSE mean scores produced by students from the SB and CP curricula showed a curricular effect on students' retention of basic science knowledge. However, preexisting differences between groups were found to be in the small-to-medium range. CONCLUSION: Evidence supporting the potential of schemes within a CP curriculum and their relation to basic science knowledge retention was observed. Effect size for the CP curriculum on students' retention of basic science knowledge was substantial; however, a notable part of that difference can be accounted for by extraneous and confounding factors. Further research utilizing more rigorous designs to investigate the relation between schemes and basic science retention is warranted. PMID- 15294460 TI - Relationship between specialty choice and medical student temperament and character assessed with Cloninger Inventory. AB - BACKGROUND: Multiple variables affect medical specialty choice, including temperament, sociodemographic factors, and personal experiences. Many studies address specific variables for specific specialties, but few assess the relative impact of each factor. PURPOSE: To identify the relative influence of temperament in choosing a specialty. METHODS: A sociodemographic and personal experiences questionnaire and a 240-question temperament and character inventory was distributed to 682 medical students. Their scores for 6 medical specialties were examined using analyses of variance, multivariate analyses of variance, and discriminant analysis. RESULTS: Students choosing surgery, emergency medicine, and obstetrics and gynecology were higher on novelty seeking than other students. Future surgeons were lower in harm avoidance and reward dependence (RD) than the others. Students choosing primary care specialties, emergency medicine, and obstetrics and gynecology were all high on RD; with pediatrics being highest. Students differed from college students, the women differed from the men, and the Asian Americans differed from the other groups. CONCLUSION: The implications of these findings are discussed for career counseling and future research. PMID- 15294461 TI - What is the impact of commercial test preparation courses on medical examination performance? AB - BACKGROUND: Commercial test preparation courses are part of the fabric of U.S. medical education. They are also big business with 2,000 sales for 1 firm listed at nearly $250 million. This article systematically reviews and evaluates research published in peer-reviewed journals and in the "grey literature" that addresses the impact of commercial test preparation courses on standardized, undergraduate medical examinations. SUMMARY: Thirteen computerized English language databases were searched using 29 search terms and search concepts from their onset to October 1, 2002. Also manually searched was medical education conference proceedings and publications after the end date; and medical education journal editors were contacted about articles accepted for publication, but not yet in print, that were deemed pertinent to this review. Studies that met three criteria were selected: (a) a commercial test preparation course or service was an educational intervention, (b) the outcome variable was one of several standardized medical examinations, and (c) results are published in a peer reviewed journal or another outlet that insures scholarly scrutiny. The criteria were applied and data extracted by consensus of 2 reviewers. The search identified 11 empirical studies, of which 10 (8 journal articles, 2 unpublished reports) are included in this review. Qualitative data synthesis and tabular presentation of research methods and outcomes are used. CONCLUSION: The articles and unpublished reports reveal that current research lacks control and rigor; the incremental validity of the commercial courses on medical examination performance, if any, is extremely small; and evidence in support of the courses is weak or nonexistent; almost no details are given about the form and conduct of the commercial test preparation courses; studies are confined to courses in preparation for the Medical College Admission Test, the former National Board of Medical Examiners Part 1, and the United States Medical Licensing Examination Step 1, not tests of clinical science; and that cost-benefit analyses of the test preparation courses have not been done. It is concluded that the utility and value of commercial test preparation courses in medicine have not been demonstrated, and that evaluation apprehension in the medical profession and aggressive marketing practices are most likely responsible for commercial course prosperity. PMID- 15294462 TI - Teaching, digression, and implicit curriculum. AB - BACKGROUND: Although medicine resides within contexts that have historical, cultural, and societal determinants, these are rarely addressed explicitly in current medical teaching. SUMMARY: The article describes a method of teaching in which mainstream biomedical learning is linked to digressions, which serve as the medium for considering the contexts of medicine, unmasking hidden messages, and broadening the scope of medical instruction. CONCLUSION: Teaching by digression encourages students to learn core clinical science while considering such otherwise neglected areas such as medical values, contexts, habits, and history. Integrating this consideration of the hidden assumptions of medical practice into mainstream medical learning allows students to understand modern biomedicine as a system that is historically, culturally, and socially conditioned. PMID- 15294754 TI - Variation in transcript abundance during somatic embryogenesis in gymnosperms. AB - Somatic embryogenesis of Norway spruce (Picea abies L.) is a versatile model system to study molecular mechanisms regulating embryo development because it proceeds through defined developmental stages corresponding to specific culture treatments. Normal embryonic development involves early differentiation of proembryogenic masses (PEMs) into somatic embryos, followed by early and late embryogeny leading to the formation of mature cotyledonary embryos. In some cell lines there is a developmental arrest at the PEM-somatic embryo transition. To learn more about the molecular mechanisms regulating embryogenesis, we compared the transcript profiles of two normal lines and one developmentally arrested line. Ribonucleic acid, extracted from these cell lines at successive developmental stages, was analyzed on DNA microarrays containing 2178 expressed sequence tags (ESTs) (corresponding to 2110 unique cDNAs) from loblolly pine (Pinus taeda L.). Hybridization between spruce and pine species on microarrays has been shown to be effective (van Zyl et al. 2002, Stasolla et al. 2003). In contrast to the developmentally arrested line, the early phases of normal embryo development are characterized by a precise pattern of gene expression, i.e., repression followed by induction. Comparison of transcript levels between successive stages of embryogenesis allowed us to identify several genes that showed unique expression responses during normal development. Several of these genes encode proteins involved in detoxification processes, methionine synthesis and utilization, and carbohydrate metabolism. The potential role of these genes in embryo development is discussed. PMID- 15294755 TI - Photosynthesis, carbohydrate storage and survival of a native and an introduced tree species in relation to light and defoliation. AB - Fraxinus uhdei (Wenz.) Lingelsh (tropical ash), a species introduced to Hawaii from Mexico, invades forests of the endemic tree Acacia koa A.Gray (koa). We examined physiological and morphological characteristics of koa and tropical ash to explore possible mechanisms that may facilitate invasion of koa forests by tropical ash. Seedlings of both species were grown in a greenhouse in three light treatments: 100% photosynthetic photon flux (PPF); 18% PPF; and 2% PPF inside the greenhouse. Light compensation point, maximum CO2 assimilation rate and dark respiration rate of seedlings differed significantly among light treatments, but were similar between species. A defoliation experiment indicated that tropical ash was better able to survive defoliation than koa, especially under high-light conditions. Tropical ash seedlings allocated more carbon (C) and nitrogen (N) to storage per unit PPF than koa seedlings. Total nonstructural carbohydrates were positively correlated with plant survival in both species. The patterns of C and N allocation associated with tropical ash seedlings favor their survival in high light, under intense herbivory and on sites where N availability is seasonal or highly variable. Variation in carbohydrate storage between koa and tropical ash greatly exceeded variation in photosynthetic performance at the leaf level. PMID- 15294756 TI - Growth response and sapwood hydraulic properties of young lodgepole pine following repeated fertilization. AB - We examined how tree growth and hydraulic properties of branches and boles are influenced by periodic (about 6 years) and annual fertilization in two juvenile lodgepole pine (Pinus contorta Dougl. var. latifolia Engelm.) stands in the interior of British Columbia, Canada. Mean basal area (BA), diameter at breast height (DBH) and height increments and percent earlywood and sapwood hydraulic parameters of branches and boles were measured 7 or 8 years after the initial treatments at Sheridan Creek and Kenneth Creek. At Sheridan Creek, fertilization significantly increased BA and DBH increments, but had no effect on height increment. At Kenneth Creek, fertilization increased BA, but fertilized trees had significantly lower height increments than control trees. Sapwood permeability was greater in lower branches of repeatedly fertilized trees than in those of control trees. Sapwood permeabilities of the lower branches of trees in the control, periodic and annual treatments were 0.24 x 10(-12), 0.35 x 10(-12) and 0.45 x 10(-12) m2 at Kenneth Creek; and 0.41 x 10(-12), 0.54 x 10(-12) and 0.65 x 10(-12) m2 at Sheridan Creek, respectively. Annual fertilization tended to increase leaf specific conductivities and Huber values of the lower branches of trees at both study sites. We conclude that, in trees fertilized annually, the higher flow capacity of lower branches may reduce the availability of water to support annual growth of the leader and upper branches. PMID- 15294757 TI - Interstock-induced mechanism of increased growth and salt resistance of orange (Citrus sinensis) trees. AB - Interstocks improve the growth and salt resistance of lemon (Citrus limon (L.) Burm. f.) trees, but their effects on orange (Citrus sinensis (L.) Osbeck) trees are unknown. We grew 'Cleopatra' mandarin (CM) seedlings, budded trees of 'Salustiano' orange (SAO) on CM, 'Valencia Late' orange (VLO) on CM (VLO/CM), and interstock trees VLO/SAO/CM in pots of sand watered with nutrient solution containing 5 (control) or 50 mM NaCl for 12 weeks. Plants were harvested on six successive occasions and the time trends in relative growth rate (RGR) and its components were estimated by fitting a Richards function regression to the harvest data. At low and high salinities, the VLO/SAO/CM combination had higher mean RGR than VLO/CM. Under control conditions, the increase in RGR caused by the interstock was the result of an increase in leaf mass fraction (LMF; leaf dry mass/plant dry mass ratio). Increases in net assimilation rate on a leaf mass basis (NARm) and LMF contributed equally to the increase in RGR in saline conditions, their growth response coefficients being 0.52 and 0.48, respectively. The structural modifications, specific leaf area (SLA) and leaf area ratio (LAR; leaf area:plant dry mass ratio), had a slight influence on the reduction in RGR by salinity. However, NARm had a large influence on RGR, except in CM. The interstock-induced mechanism increased biomass allocation to the assimilatory organs and, under saline conditions, increased Cl- and Na+ allocations to roots. Thus, the flux of ions to the leaves was either delayed or reduced or both. The dilution of imported ions by foliar growth reduced ion concentrations in leaves, resulting in higher NARm, which together with higher LMF, increased RGR. PMID- 15294758 TI - Processes preventing nocturnal equilibration between leaf and soil water potential in tropical savanna woody species. AB - The impact of nocturnal water loss and recharge of stem water storage on predawn disequilibrium between leaf (psiL) and soil (psiS) water potentials was studied in three dominant tropical savanna woody species in central Brazil (Cerrado). Sap flow continued throughout the night during the dry season and contributed from 13 to 28% of total daily transpiration. During the dry season, psiL was substantially less negative in covered transpiring leaves, throughout the day and night, than in exposed leaves. Before dawn, differences in psiL between covered and exposed leaves were about 0.4 MPa. When relationships between sap flow and psiL of exposed leaves were extrapolated to zero flow, the resulting values of psiL (a proxy of weighted mean soil water potential) in two of the species were similar to predawn values of covered leaves. Consistent with substantial nocturnal sap flow, stomatal conductance (gs) never dropped below 40 mmol m(-2) s(-1) at night, and in some cases, rose to as much as 100 mmol m(-2) s(-1) before the end of the dark period. Nocturnal gs decreased linearly with increasing air saturation deficit (D), but there were species-specific differences in the slopes of the relationships between nocturnal gs and D. Withdrawal and recharge of water from stem storage compartments were assessed by monitoring diel fluctuations of stem diameter with electronic dendrometers. Stem water storage compartments tended to recharge faster when nocturnal transpiration was reduced by covering the entire plant. Water potential of covered leaves did not stabilize in any of the plants before the end of the dark period, suggesting that, even in covered plants, water storage tissues were not fully rehydrated by dawn. Patterns of sap flow and expansion and contraction of stems reflected the dynamics of water movement during utilization and recharge of stem water storage tissues. This study showed that nighttime transpiration and recharge of internal water storage contribute to predawn disequilibrium in water potential between leaves and soil in neotropical savanna woody plants. PMID- 15294759 TI - A method for routine measurements of total sugar and starch content in woody plant tissues. AB - Several extraction and measurement methods currently employed in the determination of total sugar and starch contents in plant tissues were investigated with the view to streamline the process of total sugar and starch determination. Depending on the type and source of tissue, total sugar and starch contents estimated from samples extracted with 80% hot ethanol were significantly greater than from samples extracted with a methanol:chloroform:water solution. The residual ethanol did not interfere with the sugar and starch determination, rendering the removal of ethanol from samples unnecessary. The use of phenol sulfuric acid with a phenol concentration of 2% provided a relatively simple and reliable colorimetric method to quantify the total soluble-sugar concentration. Performing parallel sugar assays with and without phenol was more useful for accounting for the interfering effects of other substances present in plant tissue than using chloroform. For starch determination, an enzyme mixture of 1000 U alpha-amylase and 5 U amyloglucosidase digested starch in plant tissue samples more rapidly and completely than previously recommended enzyme doses. Dilute sulfuric acid (0.005 N) was less suitable for starch digestion than enzymatic hydrolysis because the acid also broke down structural carbohydrates, resulting in overestimates of starch content. After the enzymatic digestion of starch, the glucose hydrolyzate obtained was measured with a peroxidase-glucose oxidase/o dianisidine reagent; absorbance being read at 525 nm after the addition of sulfuric acid. With the help of this series of studies, we developed a refined and shortened method suitable for the rapid measurement of total sugar and starch contents in woody plant tissues. PMID- 15294760 TI - Growth CO2 concentration modifies the transpiration response of Populus deltoides to drought and vapor pressure deficit. AB - Cottonwood (Populus deltoides Bartr. ex Marsh.) trees grown for 9 months in elevated carbon dioxide concentration ([CO2]) showed significant increases in height, leaf area and basal diameter relative to trees in a near-ambient [CO2] control treatment. Sample trees in the CO2 treatments were subjected to high and low atmospheric vapor pressure deficits (VPD) over a 5-week period at both high and low soil water contents (SWC). During these periods, transpiration rates at both the leaf and canopy levels were calculated based on sap flow measurements and leaf-to-sapwood area ratios. Leaf-level transpiration rates were approximately equivalent across [CO2] treatments when soil water was not limiting. In contrast, during drought stress, canopy-level transpiration rates were approximately equivalent across [CO2] treatments, indicating that leaf-level fluxes during drought stress were reduced in elevated [CO2] by a factor equal to the leaf area ratio of the two canopies. The shift from equivalent leaf-level transpiration to equivalent canopy-level transpiration with increasing drought stress suggests maximum water use rates were controlled primarily by atmospheric demand at high SWC and by soil water availability at low SWC. Changes in VPD had less effect on transpiration than changes in SWC for trees in both CO2 treatments. Transpiration rates of trees in both CO2 treatments reached maximum values at a VPD of about 2.0 kPa at high SWC, but leveled off and decreased slightly in both canopies as VPD increased above this value. At low SWC, increasing VPD from approximately 1.4 to 2.5 kPa caused transpiration rates to decline slightly in the canopies of trees in both treatments, with significant (P = 0.004) decreases occurring in trees in the near-ambient [CO2] treatment. The transpiration responses at high VPD in the presence of high SWC and throughout the low SWC treatment suggest some hydraulic limitations to water use occurred. Comparisons of midday leaf water potentials of trees in both CO2 treatments support this conclusion. PMID- 15294761 TI - Drought tolerance and transplanting performance of holm oak (Quercus ilex) seedlings after drought hardening in the nursery. AB - Drought stress is the main cause of mortality of holm oak (Quercus ilex L.) seedlings in forest plantations. We therefore assessed if drought hardening, applied in the nursery at the end of the growing season, enhanced the drought tolerance and transplanting performance of holm oak seedlings. Seedlings were subjected to three drought hardening intensities (low, moderate and severe) for 2.5 and 3.5 months, and compared with control seedlings. At the end of the hardening period, water relations, gas exchange and morphological attributes were determined, and survival and growth under mesic and xeric transplanting conditions were assessed. Drought hardening increased drought tolerance primarily by affecting physiological traits, with no effect on shoot/root ratio or specific leaf mass. Drought hardening reduced osmotic potential at saturation and at the turgor loss point, stomatal conductance, residual transpiration (RT) and new root growth capacity (RGC), but enhanced cell membrane stability. Among treated seedlings, the largest response occurred in seedlings subjected to moderate hardening. Severe hardening reduced shoot soluble sugar concentration and increased shoot starch concentration. Increasing the duration of hardening had no effect on water relations but reduced shoot mineral and starch concentrations. Variation in cell membrane stability, RT and RGC were negatively related to osmotic adjustment. Despite differences in drought tolerance, no differences in mortality and relative growth rate were observed between hardening treatments when the seedlings were transplanted under either mesic or xeric conditions. PMID- 15294762 TI - Relationships between light, leaf nitrogen and nitrogen remobilization in the crowns of mature evergreen Quercus glauca trees. AB - We estimated the amount of nitrogen (N) remobilized from 1-year-old leaves at various positions in the crowns of mature Quercus glauca Thunb. ex Murray trees and related this to the production of new shoots. Leaf N concentration on an area basis (Na) and total N (Nt= Na x lamina area of all leaves on a shoot) were related to photosynthetic photon flux (PPF) on the leaves of current-year and 1 year-old shoots. When new shoots (S02 shoots; flushed in 2002) flushed, only a portion of the leaves on the previous year's shoots (S01 shoots; flushed in 2001) were shed. After the S02 shoots flushed, S01 shoots were defined as 1-year-old shoots (S01* shoots). Both Na and Nt were positively correlated with PPF for S01 shoots, but not for S01* shoots. The fraction of remobilized N (% of the maximum Na in S01 leaves) from remaining leaves was 5-35%, with the fraction size being positively correlated with the number of S02 shoots on an S01* shoot (new shoot number). However, the mean fraction of remobilized N from fallen leaves was 45% and was unrelated to new shoot number. The total amount of N remobilized from both fallen and remaining leaves was 1-20 mg per S01* shoot. Total remobilized N was positively correlated with new shoot number. There was a statistically significant positive relationship between the light-saturated net photosynthetic rate on a leaf area basis (Amax) and Na for both S01* and S02 leaves. However, when we compared leaves with similar Na, Amax of S01* leaves was only half that of S02 leaves, indicating that 1-year-old leaves had lower instantaneous N-use efficiency (Amax per unit Na) than current-year leaves. Ratios of chlorophyll a:b and Rubisco:chlorophyll were lower in S01* leaves than in S02 leaves, indicating that 1-year-old leaves were acclimatized to lower light environments. Thus, in Q. glauca, the N allocation theory (i.e., that N is distributed according to local PPF) applied only to the current-year shoots. Although the amount of foliar N in 1-year-old shoots was not strongly affected by the PPF on 1-year-old leaves, it was affected by interactions with current-year shoots. PMID- 15294763 TI - Responses to water stress in two Eucalyptus globulus clones differing in drought tolerance. AB - We evaluated drought resistance mechanisms in a drought-tolerant clone (CN5) and a drought-sensitive clone (ST51) of Eucalyptus globulus Labill. based on the responses to drought of some physiological, biophysical and morphological characteristics of container-grown plants, with particular emphasis on root growth and hydraulic properties. Water loss in excess of that supplied to the containers led to a general decrease in growth and significant reductions in leaf area ratio, specific leaf area and leaf-to-root area ratio. Root hydraulic conductance and leaf-specific hydraulic conductance decreased as water stress became more severe. During the experiment, the drought-resistant CN5 clone maintained higher leaf water status (higher predawn and midday leaf water potentials), sustained a higher growth rate (new leaf area expansion and root growth) and displayed greater carbon allocation to the root system and lower leaf to-root area ratio than the drought-sensitive ST51 clone. Clone CN5 possessed higher stomatal conductances at moderate stress as well as higher hydraulic conductances than Clone ST51. Differences in the response to drought in root biomass, coupled with changes in hydraulic properties, accounted for the clonal differences in drought tolerance, allowing Clone CN5 to balance transpiration and water absorption during drought treatment and thereby prolong the period of active carbon assimilation. PMID- 15294764 TI - Effects of root medium pH on water transport in paper birch (Betula papyrifera) seedlings in relation to root temperature and abscisic acid treatments. AB - We investigated the effects of root medium pH on water transport in whole-plant and detached roots of paper birch (Betula papyrifera Marsh.). Exposure of seedling roots to pH 4 and 8 significantly decreased root hydraulic conductivity (Lp) and stomatal conductance (gs), compared with pH 6. When roots of solution culture-grown (pH 6) seedlings were transferred to pH 4 or 8, their steady-state water flow (Qv) declined within minutes, followed by a decline in gs. The root oxygen uptake rates were not significantly affected by the pH treatments. Treatment of roots with mercuric chloride resulted in a large decrease in Qv at pH 6; the extent of this decrease was similar to that brought about by pH 4 and 8. Lowering root temperature from 21 to 4 degrees C decreased Qv irrespective of medium pH. Low root temperatures did not offset the effects of medium pH 4 on Qv and the roots in this treatment had a high activation energy for water flow. Conversely, roots exposed to pH 8 had a low activation energy, similar to that at pH 6. When 2 micro M abscisic acid, (+/-)-cis-trans-ABA, was added to the root medium, Qv increased in roots that were incubated at pH 6. It also increased slightly in roots incubated at pH 4, but not at pH 8. The increase at pH 4 and 6 was temperature-dependent, occurring at 21 degrees C, but not 4 degrees C. We suggest that the pH treatments are responsible for altering root water flow properties through their effects on the activity of water channels. These results support the concept that ABA effects on water channels are modulated by other, possibly metabolic- and pH-dependent factors. PMID- 15294765 TI - High stability of nuclear microsatellite loci during the early stages of somatic embryogenesis in Norway spruce. AB - Somatic embryos of Norway spruce (Picea abies (L.) Karst.) differentiate from proembryogenic masses (PEMs), which are subject to autodestruction through programmed cell death. In PEMs, somatic embryo formation and activation of programmed cell death are interrelated processes. We sought to determine if activation of programmed cell death in PEMs is caused by genetic aberrations during somatic embryogenesis. Based on the finding that withdrawal of auxin and cytokinin induces programmed cell death in PEMs, 1-week-old cell suspensions were cultured in medium either with or without auxin and cytokinin and then transferred to maturation medium containing abscisic acid. We analyzed the stability of three nuclear simple sequence repeat (SSR) microsatellite markers at successive stages of somatic embryogenesis in two cell lines. There were no mutations at the SSR loci at any of the successive developmental stages from PEMs to cotyledonary embryos, irrespective of whether or not the proliferation medium in which cell suspensions had been cultured contained auxin or cytokinin. The morphologies of plants regenerated from the cultures were similar, although withdrawal of auxin and cytokinin significantly stimulated the yield of both embryos and plants. We conclude, therefore, that the high genetic stability of somatic embryos in Norway spruce is unaffected by the induction of programmed cell death caused by withdrawal of auxin and cytokinin. PMID- 15294766 TI - Interspecific variation of photosynthesis and leaf characteristics in canopy trees of five species of Dipterocarpaceae in a tropical rain forest. AB - Photosynthetic rate, nitrogen concentration and morphological properties of canopy leaves were studied in 18 trees, comprising five dipterocarp species, in a tropical rain forest in Sarawak, Malaysia. Photosynthetic rate at light saturation (Pmax) differed significantly across species, varying from 7 to 18 micro mol m(-2) s(-1). Leaf nitrogen concentration and morphological properties, such as leaf blade and palisade layer thickness, leaf mass per area (LMA) and surface area of mesophyll cells per unit leaf area (Ames/A), also varied significantly across species. Among the relationships with leaf characteristics, Pmax had the strongest correlation with leaf mesophyll parameters, such as palisade cell layer thickness (r2 = 0.76, P < 0.001) and Ames/A (r2 = 0.73, P < 0.001). Leaf nitrogen concentration and Pmax per unit area also had a significant but weaker correlation (r2 = 0.46, P < 0.01), whereas Pmax had no correlation, or only weakly significant correlations, with leaf blade thickness and LMA. Shorea beccariana Burck, which had the highest P(max) of the species studied, also had the thickest palisade layer, with up to five or more layers. We conclude that interspecific variation in photosynthetic capacity in tropical rain forest canopies is influenced more by leaf mesophyll structure than by leaf thickness, LMA or leaf nitrogen concentration. PMID- 15294767 TI - Urease-positive thermophilic Campylobacter species. PMID- 15294768 TI - Mating type sequences in asexually reproducing Fusarium species. AB - To assess the potential for mating in several Fusarium species with no known sexual stage, we developed degenerate and semidegenerate oligonucleotide primers to identify conserved mating type (MAT) sequences in these fungi. The putative alpha and high-mobility-group (HMG) box sequences from Fusarium avenaceum, F. culmorum, F. poae, and F. semitectum were compared to similar sequences that were described previously for other members of the genus. The DNA sequences of the regions flanking the amplified MAT regions were obtained by inverse PCR. These data were used to develop diagnostic primers suitable for the clear amplification of conserved mating type sequences from any member of the genus Fusarium. By using these diagnostic primers, we identified mating types of 122 strains belonging to 22 species of Fusarium. The alpha box and the HMG box from the mating type genes are transcribed in F. avenaceum, F. culmorum, F. poae, and F. semitectum. The novelty of the PCR-based mating type identification system that we developed is that this method can be used on a wide range of Fusarium species, which have proven or expected teleomorphs in different ascomycetous genera, including Calonectria, Gibberella, and Nectria. PMID- 15294769 TI - Salt-inducible multidrug efflux pump protein in the moderately halophilic bacterium Chromohalobacter sp. AB - It has been known that halophilic bacteria often show natural resistance to antibiotics, dyes, and toxic metal ions, but the mechanism and regulation of this resistance have remained unexplained. We have addressed this question by identifying the gene responsible for multidrug resistance. A spontaneous ofloxacin-resistant mutant derived from the moderately halophilic bacterium Chromohalobacter sp. strain 160 showed a two- to fourfold increased resistance to structurally diverse compounds, such as tetracycline, cefsulodin, chloramphenicol, and ethidium bromide (EtBr), and tolerance to organic solvents, e.g., hexane and heptane. The mutant produced an elevated level of the 58-kDa outer membrane protein. This mutant (160R) accumulated about one-third the level of EtBr that the parent cells did. An uncoupler, carbonyl cyanide m chlorophenylhydrazone, caused a severalfold increase in the intracellular accumulation of EtBr, with the wild-type and mutant cells accumulating nearly equal amounts. The hrdC gene encoding the 58-kDa outer membrane protein has been cloned. Disruption of this gene rendered the cells hypersusceptible to antibiotics and EtBr and led to a high level of accumulation of intracellular EtBr. The primary structure of HrdC has a weak similarity to that of Escherichia coli TolC. Interestingly, both drug resistance and the expression of HrdC were markedly increased in the presence of a high salt concentration in the growth medium, but this was not observed in hrdC-disrupted cells. These results indicate that HrdC is the outer membrane component of the putative efflux pump assembly and that it plays a major role in the observed induction of drug resistance by salt in this bacterium. PMID- 15294770 TI - Culture-independent analysis of fecal enterobacteria in environmental samples by single-cell mRNA profiling. AB - A culture-independent method called mRNA profiling has been developed for the analysis of fecal enterobacteria and their physiological status in environmental samples. This taxon-specific approach determines the single-cell content of selected gene transcripts whose abundance is either directly or inversely proportional to growth state. Fluorescence in situ hybridization using fluorochrome-labeled oligonucleotide probes was used to measure the cellular concentration of fis and dps mRNA. Relative levels of these transcripts provided a measure of cell growth state and the ability to enumerate fecal enterobacterial cell number. Orthologs were cloned by inverse PCR from several major enterobacterial genera, and probes specific for fecal enterobacteria were designed using multiple DNA sequence alignments. Probe specificity was determined experimentally using pure and mixed cultures of the major enterobacterial genera as well as secondary treated wastewater samples seeded with pure culture inocula. Analysis of the fecal enterobacterial community resident in unseeded secondary treated wastewater detected fluctuations in transcript abundance that were commensurate with incubation time and nutrient availability and demonstrated the utility of the method using environmental samples. mRNA profiling provides a new strategy to improve wastewater disinfection efficiency by accelerating water quality analysis. PMID- 15294771 TI - Polyhydroxyalkanoate (PHA) accumulation in sulfate-reducing bacteria and identification of a class III PHA synthase (PhaEC) in Desulfococcus multivorans. AB - Seven strains of sulfate-reducing bacteria (SRB) were tested for the accumulation of polyhydroxyalkanoates (PHAs). During growth with benzoate Desulfonema magnum accumulated large amounts of poly(3-hydroxybutyrate) [poly(3HB)]. Desulfosarcina variabilis (during growth with benzoate), Desulfobotulus sapovorans (during growth with caproate), and Desulfobacterium autotrophicum (during growth with caproate) accumulated poly(3HB) that accounted for 20 to 43% of cell dry matter. Desulfobotulus sapovorans and Desulfobacterium autotrophicum also synthesized copolyesters consisting of 3-hydroxybutyrate and 3-hydroxyvalerate when valerate was used as the growth substrate. Desulfovibrio vulgaris and Desulfotalea psychrophila were the only SRB tested in which PHAs were not detected. When total DNA isolated from Desulfococcus multivorans and specific primers deduced from highly conserved regions of known PHA synthases (PhaC) were used, a PCR product homologous to the central region of class III PHA synthases was obtained. The complete pha locus of Desulfococcus multivorans was subsequently obtained by inverse PCR, and it contained adjacent phaE(Dm) and phaC(Dm) genes. PhaC(Dm) and PhaE(Dm) were composed of 371 and 306 amino acid residues and showed up to 49 or 23% amino acid identity to the corresponding subunits of other class III PHA synthases. Constructs of phaC(Dm) alone (pBBRMCS-2::phaC(Dm)) and of phaE(Dm)C(Dm) (pBBRMCS-2::phaE(Dm)C(Dm)) in various vectors were obtained and transferred to several strains of Escherichia coli, as well as to the PHA negative mutants PHB(-)4 and GPp104 of Ralstonia eutropha and Pseudomonas putida, respectively. In cells of the recombinant strains harboring phaE(Dm)C(Dm) small but significant amounts (up to 1.7% of cell dry matter) of poly(3HB) and of PHA synthase activity (up to 1.5 U/mg protein) were detected. This indicated that the cloned genes encode functionally active proteins. Hybrid synthases consisting of PhaC(Dm) and PhaE of Thiococcus pfennigii or Synechocystis sp. strain PCC 6308 were also constructed and were shown to be functionally active. PMID- 15294772 TI - Benzoic acid, a weak organic acid food preservative, exerts specific effects on intracellular membrane trafficking pathways in Saccharomyces cerevisiae. AB - Microbial spoilage of food causes losses of up to 40% of all food grown for human consumption worldwide. Yeast growth is a major factor in the spoilage of foods and beverages that are characterized by a high sugar content, low pH, and low water activity, and it is a significant economic problem. While growth of spoilage yeasts such as Zygosaccharomyces bailii and Saccharomyces cerevisiae can usually be retarded by weak organic acid preservatives, the inhibition often requires levels of preservative that are near or greater than the legal limits. We identified a novel synergistic effect of the chemical preservative benzoic acid and nitrogen starvation: while exposure of S. cerevisiae to either benzoic acid or nitrogen starvation is cytostatic under our conditions, the combination of the two treatments is cytocidal and can therefore be used beneficially in food preservation. In yeast, as in all eukaryotic organisms, survival under nitrogen starvation conditions requires a cellular response called macroautophagy. During macroautophagy, cytosolic material is sequestered by intracellular membranes. This material is then targeted for lysosomal degradation and recycled into molecular building blocks, such as amino acids and nucleotides. Macroautophagy is thought to allow cellular physiology to continue in the absence of external resources. Our analyses of the effects of benzoic acid on intracellular membrane trafficking revealed that there was specific inhibition of macroautophagy. The data suggest that the synergism between nitrogen starvation and benzoic acid is the result of inhibition of macroautophagy by benzoic acid and that a mechanistic understanding of this inhibition should be beneficial in the development of novel food preservation technologies. PMID- 15294773 TI - Ecology and transmission of Listeria monocytogenes infecting ruminants and in the farm environment. AB - A case-control study involving 24 case farms with at least one recent case of listeriosis and 28 matched control farms with no listeriosis cases was conducted to probe the transmission and ecology of Listeria monocytogenes on farms. A total of 528 fecal, 516 feed, and 1,012 environmental soil and water samples were cultured for L. monocytogenes. While the overall prevalence of L. monocytogenes in cattle case farms (24.4%) was similar to that in control farms (20.2%), small ruminant (goat and sheep) farms showed a significantly (P < 0.0001) higher prevalence in case farms (32.9%) than in control farms (5.9%). EcoRI ribotyping of clinical (n = 17) and farm (n = 414) isolates differentiated 51 ribotypes. L. monocytogenes ribotypes isolated from clinical cases and fecal samples were more frequent in environmental than in feed samples, indicating that infected animals may contribute to L. monocytogenes dispersal into the farm environment. Ribotype DUP-1038B was significantly (P < 0.05) associated with fecal samples compared with farm environment and animal feedstuff samples. Ribotype DUP-1045A was significantly (P < 0.05) associated with soil compared to feces and with control farms compared to case farms. Our data indicate that (i) the epidemiology and transmission of L. monocytogenes differ between small-ruminant and cattle farms; (ii) cattle contribute to amplification and dispersal of L. monocytogenes into the farm environment, (iii) the bovine farm ecosystem maintains a high prevalence of L. monocytogenes, including subtypes linked to human listeriosis cases and outbreaks, and (iv) L. monocytogenes subtypes may differ in their abilities to infect animals and to survive in farm environments. PMID- 15294774 TI - Identification of Carnobacterium species by restriction fragment length polymorphism of the 16S-23S rRNA gene intergenic spacer region and species specific PCR. AB - The genus Carnobacterium is currently divided into the following eight species: Carnobacterium piscicola, C. divergens, C. gallinarum, C. mobile, C. funditum, C. alterfunditum, C. inhibens, and C. viridans. An identification tool for the rapid differentiation of these eight Carnobacterium species was developed, based on the 16S-23S ribosomal DNA (rDNA) intergenic spacer region (ISR). PCR-restriction fragment length polymorphism (PCR-RFLP) analysis of this 16S-23S rDNA ISR was performed in order to obtain restriction profiles for all of the species. Three PCR amplicons, which were designated small ISR (S-ISR), medium ISR (M-ISR), and large ISR (L-ISR), were obtained for all Carnobacterium species. The L-ISR sequence revealed the presence of two tRNA genes, tRNA(Ala) and tRNA(Ile), which were separated by a spacer region that varied from 24 to 38 bp long. This region was variable among the species, allowing the design of species-specific primers. These primers were tested and proved to be species specific. The identification method based on the 16S-23S rDNA ISR, using PCR-RFLP and specific primers, is very suitable for the rapid low-cost identification and discrimination of all of the Carnobacterium species from other phylogenetically related lactic acid bacteria. PMID- 15294775 TI - Sample size, library composition, and genotypic diversity among natural populations of Escherichia coli from different animals influence accuracy of determining sources of fecal pollution. AB - A horizontal, fluorophore-enhanced, repetitive extragenic palindromic-PCR (rep PCR) DNA fingerprinting technique (HFERP) was developed and evaluated as a means to differentiate human from animal sources of Escherichia coli. Box A1R primers and PCR were used to generate 2,466 rep-PCR and 1,531 HFERP DNA fingerprints from E. coli strains isolated from fecal material from known human and 12 animal sources: dogs, cats, horses, deer, geese, ducks, chickens, turkeys, cows, pigs, goats, and sheep. HFERP DNA fingerprinting reduced within-gel grouping of DNA fingerprints and improved alignment of DNA fingerprints between gels, relative to that achieved using rep-PCR DNA fingerprinting. Jackknife analysis of the complete rep-PCR DNA fingerprint library, done using Pearson's product-moment correlation coefficient, indicated that animal and human isolates were assigned to the correct source groups with an 82.2% average rate of correct classification. However, when only unique isolates were examined, isolates from a single animal having a unique DNA fingerprint, Jackknife analysis showed that isolates were assigned to the correct source groups with a 60.5% average rate of correct classification. The percentages of correctly classified isolates were about 15 and 17% greater for rep-PCR and HFERP, respectively, when analyses were done using the curve-based Pearson's product-moment correlation coefficient, rather than the band-based Jaccard algorithm. Rarefaction analysis indicated that, despite the relatively large size of the known-source database, genetic diversity in E. coli was very great and is most likely accounting for our inability to correctly classify many environmental E. coli isolates. Our data indicate that removal of duplicate genotypes within DNA fingerprint libraries, increased database size, proper methods of statistical analysis, and correct alignment of band data within and between gels improve the accuracy of microbial source tracking methods. PMID- 15294776 TI - Bactericidal activity of glycinecin A, a bacteriocin derived from Xanthomonas campestris pv. glycines, on phytopathogenic Xanthomonas campestris pv. vesicatoria cells. AB - The ability of glycinecin A, a bacteriocin derived from Xanthomonas campestris pv. glycines 8ra, to kill closely related bacteria has been demonstrated previously by our group. In the present study, we aimed at determining the glycinecin A-induced cause of death. Treatment with glycinecin A caused slow dissipation of membrane potential and rapid depletion of the pH gradient. Glycinecin A treatment also induced leakage of potassium ions from X. campestris pv. vesicatoria YK93-4 cells and killed sensitive bacterial cells in a dose dependent manner. Sensitive cells were killed within 2 h of incubation, most likely due to the potassium ion efflux caused by glycinecin A. These results suggest that the bactericidal mechanism of action of glycinecin A is correlated with the permeability of membranes to hydroxyl and potassium ions, leading to the lethal activity of the bacteriocin on the target bacteria. PMID- 15294777 TI - Differential protein expression during growth of Acidithiobacillus ferrooxidans on ferrous iron, sulfur compounds, or metal sulfides. AB - A set of proteins that changed their levels of synthesis during growth of Acidithiobacillus ferrooxidans ATCC 19859 on metal sulfides, thiosulfate, elemental sulfur, and ferrous iron was characterized by using two-dimensional polyacrylamide gel electrophoresis. N-terminal amino acid sequencing and mass spectrometry analysis of these proteins allowed their identification and the localization of the corresponding genes in the available genomic sequence of A. ferrooxidans ATCC 23270. The genomic context around several of these genes suggests their involvement in the energetic metabolism of A. ferrooxidans. Two groups of proteins could be distinguished. The first consisted of proteins highly upregulated by growth on sulfur compounds (and downregulated by growth on ferrous iron): a 44-kDa outer membrane protein, an exported 21-kDa putative thiosulfate sulfur transferase protein, a 33-kDa putative thiosulfate/sulfate binding protein, a 45-kDa putative capsule polysaccharide export protein, and a putative 16-kDa protein of unknown function. The second group of proteins comprised those downregulated by growth on sulfur (and upregulated by growth on ferrous iron): rusticyanin, a cytochrome c(552), a putative phosphate binding protein (PstS), the small and large subunits of ribulose biphosphate carboxylase, and a 30-kDa putative CbbQ protein, among others. The results suggest in general a separation of the iron and sulfur utilization pathways. Rusticyanin, in addition to being highly expressed on ferrous iron, was also newly synthesized, as determined by metabolic labeling, although at lower levels, during growth on sulfur compounds and iron-free metal sulfides. During growth on metal sulfides containing iron, such as pyrite and chalcopyrite, both proteins upregulated on ferrous iron and those upregulated on sulfur compounds were synthesized, indicating that the two energy-generating pathways are induced simultaneously depending on the kind and concentration of oxidizable substrates available. PMID- 15294778 TI - Expression of a temperature-sensitive esterase in a novel chaperone-based Escherichia coli strain. AB - A new principle for expression of heat-sensitive recombinant proteins in Escherichia coli at temperatures close to 4 degrees C was experimentally evaluated. This principle was based on simultaneous expression of the target protein with chaperones (Cpn60 and Cpn10) from a psychrophilic bacterium, Oleispira antarctica RB8(T), that allow E. coli to grow at high rates at 4 degrees C (maximum growth rate, 0.28 h(-1)). The expression of a temperature sensitive esterase in this host at 4 to 10 degrees C yielded enzyme specific activity that was 180-fold higher than the activity purified from the non chaperonin-producing E. coli strain grown at 37 degrees C (32,380 versus 190 micromol min(-1) g(-1)). We present evidence that the increased specific activity was not due to the low growth temperature per se but was due to the fact that low temperature was beneficial to folding, with or without chaperones. This is the first report of successful use of a chaperone-based E. coli strain to express heat-labile recombinant proteins at temperatures below the theoretical minimum growth temperature of a common E. coli strain (7.5 degrees C). PMID- 15294779 TI - Polydextrose, lactitol, and fructo-oligosaccharide fermentation by colonic bacteria in a three-stage continuous culture system. AB - In vitro fermentations were carried out by using a model of the human colon to simulate microbial activities of lower gut bacteria. Bacterial populations (and their metabolic products) were evaluated under the effects of various fermentable substrates. Carbohydrates tested were polydextrose, lactitol, and fructo oligosaccharide (FOS). Bacterial groups of interest were evaluated by fluorescence in situ hybridization as well as by species-specific PCR to determine bifidobacterial species and percent-G+C profiling of the bacterial communities present. Short-chain fatty acids (SCFA) produced during the fermentations were also evaluated. Polydextrose had a stimulatory effect upon colonic bifidobacteria at concentrations of 1 and 2% (using a single and pooled human fecal inoculum, respectively). The bifidogenic effect was sustained throughout all three vessels of the in vitro system (P = 0.01 seen in vessel 3), as corroborated by the bacterial community profile revealed by %G+C analysis. This substrate supported a wide variety of bifidobacteria and was the only substrate where Bifidobacterium infantis was detected. The fermentation of lactitol had a deleterious effect on both bifidobacterial and bacteroides populations (P = 0.01) and decreased total cell numbers. SCFA production was stimulated, however, particularly butyrate (beneficial for host colonocytes). FOS also had a stimulatory effect upon bifidobacterial and lactobacilli populations that used a single inoculum (P = 0.01 for all vessels) as well as a bifidogenic effect in vessels 2 and 3 (P = 0.01) when a pooled inoculum was used. A decrease in bifidobacteria throughout the model was reflected in the percent-G+C profiles. PMID- 15294780 TI - Analysis of structural and physiological profiles to assess the effects of Cu on biofilm microbial communities. AB - We investigated the effects of copper on the structure and physiology of freshwater biofilm microbial communities. For this purpose, biofilms that were grown during 4 weeks in a shallow, slightly polluted ditch were exposed, in aquaria in our laboratory, to a range of copper concentrations (0, 1, 3, and 10 microM). Denaturing gradient gel electrophoresis (DGGE) revealed changes in the bacterial community in all aquaria. The extent of change was related to the concentration of copper applied, indicating that copper directly or indirectly caused the effects. Concomitantly with these changes in structure, changes in the metabolic potential of the heterotrophic bacterial community were apparent from changes in substrate use profiles as assessed on Biolog plates. The structure of the phototrophic community also changed during the experiment, as observed by microscopic analysis in combination with DGGE analysis of eukaryotic microorganisms and cyanobacteria. However, the extent of community change, as observed by DGGE, was not significantly greater in the copper treatments than in the control. Yet microscopic analysis showed a development toward a greater proportion of cyanobacteria in the treatments with the highest copper concentrations. Furthermore, copper did affect the physiology of the phototrophic community, as evidenced by the fact that a decrease in photosynthetic capacity was detected in the treatment with the highest copper concentration. Therefore, we conclude that copper affected the physiology of the biofilm and had an effect on the structure of the communities composing this biofilm. PMID- 15294781 TI - Purification and characterization of chitosanase from Bacillus sp. strain KCTC 0377BP and its application for the production of chitosan oligosaccharides. AB - For the enzymatic production of chitosan oligosaccharides from chitosan, a chitosanase-producing bacterium, Bacillus sp. strain KCTC 0377BP, was isolated from soil. The bacterium constitutively produced chitosanase in a culture medium without chitosan as an inducer. The production of chitosanase was increased from 1.2 U/ml in a minimal chitosan medium to 100 U/ml by optimizing the culture conditions. The chitosanase was purified from a culture supernatant by using CM Toyopearl column chromatography and a Superose 12HR column for fast-performance liquid chromatography and was characterized according to its enzyme properties. The molecular mass of the enzyme was estimated to be 45 kDa by means of sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The enzyme demonstrated bifunctional chitosanase-glucanase activities, although it showed very low glucanase activity, with less than 3% of the chitosanase activity. Activity of the enzyme increased with an increase of the degrees of deacetylation (DDA) of the chitosan substrate. However, the enzyme still retained 72% of its relative activity toward the 39% DDA of chitosan, compared with the activity of the 94% DDA of chitosan. The enzyme produced chitosan oligosaccharides from chitosan, ranging mainly from chitotriose to chitooctaose. By controlling the reaction time and by monitoring the reaction products with gel filtration high-performance liquid chromatography, chitosan oligosaccharides with a desired oligosaccharide content and composition were obtained. In addition, the enzyme was efficiently used for the production of low-molecular-weight chitosan and highly acetylated chitosan oligosaccharides. A gene (csn45) encoding chitosanase was cloned, sequenced, and compared with other functionally related genes. The deduced amino acid sequence of csn45 was dissimilar to those of the classical chitosanase belonging to glycoside hydrolase family 46 but was similar to glucanases classified with glycoside hydrolase family 8. PMID- 15294782 TI - Purification, cloning, and sequencing of a 3,5-dichlorophenol reductive dehalogenase from Desulfitobacterium frappieri PCP-1. AB - A membrane-associated 3,5-dichlorophenol reductive dehalogenase was isolated from Desulfitobacterium frappieri PCP-1. The highest dehalogenase activity was observed with the biomass cultured at 22 degrees C, compared to 30 and 37 degrees C, where the cell suspensions were 2.2 and 9.6 times less active, respectively. The reductive dehalogenase was purified 12.7-fold to apparent homogeneity. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a single band with an apparent molecular mass of 57 kDa. Its dechlorinating activity was not inhibited by sulfate and nitrate but was completely inhibited by 2.5 mM sulfite and 10 mM KCN. A mixture of iodopropane and titanium citrate caused a light-reversible inhibition of the dechlorinating activities, suggesting the involvement of a corrinoid cofactor. Several polychlorophenols were dechlorinated at the meta and para positions. The apparent K(m) for 3,5-dicholorophenol was 49.3 +/- 3.1 microM at a methyl viologen concentration of 2 mM. Six internal tryptic peptides were sequenced by mass spectrometry. One open reading frame (ORF) was found in the Desulfitobacterium hafniense genome containing these peptide sequences. This ORF corresponds to a gene coding for a CprA-type reductive dehalogenase. The corresponding ORF (named cprA5) in D. frappieri PCP-1 was cloned and sequenced. The cprA5 gene codes for a 548-amino-acid protein that contains a twin-arginine type signal for secretion. The gene product has a cobalamin binding site motif and two iron-sulfur binding motifs and shows 66% identity (76 to 77% similarity) with some tetrachloroethene reductive dehalogenases. This is the first CprA-type reductive dehalogenase that can dechlorinate chlorophenols at the meta and para positions. PMID- 15294783 TI - Inactivation of caliciviruses. AB - The viruses most commonly associated with food- and waterborne outbreaks of gastroenteritis are the noroviruses. The lack of a culture method for noroviruses warrants the use of cultivable model viruses to gain more insight on their transmission routes and inactivation methods. We studied the inactivation of the reported enteric canine calicivirus no. 48 (CaCV) and the respiratory feline calicivirus F9 (FeCV) and correlated inactivation to reduction in PCR units of FeCV, CaCV, and a norovirus. Inactivation of suspended viruses was temperature and time dependent in the range from 0 to 100 degrees C. UV-B radiation from 0 to 150 mJ/cm(2) caused dose-dependent inactivation, with a 3 D (D = 1 log(10)) reduction in infectivity at 34 mJ/cm(2) for both viruses. Inactivation by 70% ethanol was inefficient, with only 3 D reduction after 30 min. Sodium hypochlorite solutions were only effective at >300 ppm. FeCV showed a higher stability at pH <3 and pH >7 than CaCV. For all treatments, detection of viral RNA underestimated the reduction in viral infectivity. Norovirus was never more sensitive than the animal caliciviruses and profoundly more resistant to low and high pH. Overall, both animal viruses showed similar inactivation profiles when exposed to heat or UV-B radiation or when incubated in ethanol or hypochlorite. The low stability of CaCV at low pH suggests that this is not a typical enteric (calici-) virus. The incomplete inactivation by ethanol and the high hypochlorite concentration needed for sufficient virus inactivation point to a concern for decontamination of fomites and surfaces contaminated with noroviruses and virus safe water. PMID- 15294784 TI - Oxidation of methyl tert-butyl ether by alkane hydroxylase in dicyclopropylketone induced and n-octane-grown Pseudomonas putida GPo1. AB - The alkane hydroxylase enzyme system in Pseudomonas putida GPo1 has previously been reported to be unreactive toward the gasoline oxygenate methyl tert-butyl ether (MTBE). We have reexamined this finding by using cells of strain GPo1 grown in rich medium containing dicyclopropylketone (DCPK), a potent gratuitous inducer of alkane hydroxylase activity. Cells grown with DCPK oxidized MTBE and generated stoichiometric quantities of tert-butyl alcohol (TBA). Cells grown in the presence of DCPK also oxidized tert-amyl methyl ether but did not appear to oxidize either TBA, ethyl tert-butyl ether, or tert-amyl alcohol. Evidence linking MTBE oxidation to alkane hydroxylase activity was obtained through several approaches. First, no TBA production from MTBE was observed with cells of strain GPo1 grown on rich medium without DCPK. Second, no TBA production from MTBE was observed in DCPK-treated cells of P. putida GPo12, a strain that lacks the alkane-hydroxylase-encoding OCT plasmid. Third, all n-alkanes that support the growth of strain GPo1 inhibited MTBE oxidation by DCPK-treated cells. Fourth, two non-growth-supporting n-alkanes (propane and n-butane) inhibited MTBE oxidation in a saturable, concentration-dependent process. Fifth, 1,7-octadiyne, a putative mechanism-based inactivator of alkane hydroxylase, fully inhibited TBA production from MTBE. Sixth, MTBE-oxidizing activity was also observed in n octane-grown cells. Kinetic studies with strain GPo1 grown on n-octane or rich medium with DCPK suggest that MTBE-oxidizing activity may have previously gone undetected in n-octane-grown cells because of the unusually high K(s) value (20 to 40 mM) for MTBE. PMID- 15294785 TI - Effects of phosphate and light on growth of and bioactive peptide production by the Cyanobacterium anabaena strain 90 and its anabaenopeptilide mutant. AB - Cyanobacteria synthesize several types of bioactive secondary metabolites. Anabaena strain 90 produces three types of bioactive peptides, microcystins (inhibitors of protein phosphatases 1 and 2A), anabaenopeptilides, and anabaenopeptins (serine protease inhibitors). To investigate the role of the anabaenopeptilides in Anabaena, wild-type strain 90 (WT) and its anabaenopeptilide deficient mutant (MU) were cultured with various light and phosphate levels to evaluate the effects and coeffects of these growth factors on the concentrations of the three classes of peptides and the growth characteristics. WT and MU grew in comparable ways under the different growth conditions. The total peptide concentration in WT was significantly higher than that in MU (2.5 and 1.4 microg/mg [dry weight], respectively). Interestingly, the average concentration of anabaenopeptins was significantly higher in MU than in WT (0.59 and 0.24 microg/mg [dry weight], respectively). The concentration of microcystins was slightly but not statistically significantly higher in MU than in WT (1.0 and 0.86 microg/mg [dry weight], respectively). In WT, the highest peptide concentrations were usually found after 13 days in cultures grown at medium light intensities (23 micromol m(-2) s(-1)) and with the highest phosphate concentrations (2,600 microg liter(-1)). In MU, the highest peptide concentrations were found in 13-day-old cultures grown at medium light intensities (23 micromol m(-2) s(-1)) and with phosphate concentrations greater than 100 microg liter(-1). The higher concentrations of anabaenopeptins in MU may compensate for the absence of anabaenopeptilides. These findings clearly indicate that these compounds may have some linked function in the producer organism, the nature of which remains to be discovered. PMID- 15294786 TI - Rapid real-time PCR assay for detection and quantitation of Mycobacterium avium subsp. paratuberculosis DNA in artificially contaminated milk. AB - Using fluorescence resonance energy transfer technology and Lightcycler analysis, we developed a real-time PCR assay with primers and probes designed by using IS900 which allowed rapid detection of Mycobacterium avium subsp. paratuberculosis DNA in artificially contaminated milk. Initially, the PCR parameters (including primer and probe levels, assay volume, Mg(2+) concentration, and annealing temperature) were optimized. Subsequently, the quantitative ability of the assay was tested and was found to be accurate over a broad linear range (3 x 10(6) to 3 x 10(1) copies). The assay sensitivity when purified DNA was used was determined to be as low as five copies, with excellent reproducibility. A range of DNA isolation strategies was developed for isolating M. avium subsp. paratuberculosis DNA from spiked milk, the most effective of which involved the use of 50 mM Tris HCl, 10 mM EDTA, 2% Triton X-100, 4 M guanidinium isothiocyante, and 0.3 M sodium acetate combined with boiling, physical grinding, and nucleic acid spin columns. When this technique was used in conjunction with the real-time PCR assay, it was possible to consistently detect <100 organisms per ml of milk (equivalent to 2,000 organisms per 25 ml). Furthermore, the entire procedure (extraction and PCR) was performed in less than 3 h and was successfully adapted to quantify M. avium subsp. paratuberculosis in spiked milk from heavily and mildly contaminated samples. PMID- 15294787 TI - Expression and immunogenicity of a recombinant diphtheria toxin fragment A in Streptococcus gordonii. AB - A nontoxic mutant diphtheria toxin fragment A (DTA) was genetically fused in single, double, or triple copy to the major surface protein antigen P1 (SpaP) and surface expressed in Streptococcus gordonii DL-1. The expression was verified by Western immunoblotting. Mouse antisera raised against the recombinant S. gordonii recognized the native diphtheria toxinm suggesting the recombinant DTA was immunogenic. When given intranasally to mice with cholera toxin subunit B as the adjuvant, the recombinant S. gordonii expressing double copies of DTA (SpaP DTA(2)) induced a mucosal immunoglobulin A response and a weak systemic immunoglobulin G response. S. gordonii SpaP-DTA(2) was able to orally colonize BALB/c mice for a 15-week period and elicited a mucosal response, but a serum immunoglobulin G response was not apparent. The antisera failed to neutralize diphtheria toxin cytotoxicity in a Vero cell assay. PMID- 15294788 TI - Novel haloperoxidase from the agaric basidiomycete Agrocybe aegerita oxidizes aryl alcohols and aldehydes. AB - Agrocybe aegerita, a bark mulch- and wood-colonizing basidiomycete, was found to produce a peroxidase (AaP) that oxidizes aryl alcohols, such as veratryl and benzyl alcohols, into the corresponding aldehydes and then into benzoic acids. The enzyme also catalyzed the oxidation of typical peroxidase substrates, such as 2,6-dimethoxyphenol (DMP) or 2,2'-azinobis-(3-ethylbenzothiazoline-6-sulfonate) (ABTS). A. aegerita peroxidase production depended on the concentration of organic nitrogen in the medium, and highest enzyme levels were detected in the presence of soybean meal. Two fractions of the enzyme, AaP I and AaP II, which had identical molecular masses (46 kDa) and isoelectric points of 4.6 to 5.4 and 4.9 to 5.6, respectively (corresponding to six different isoforms), were identified after several steps of purification, including anion- and cation exchange chromatography. The optimum pH for the oxidation of aryl alcohols was found to be around 7, and the enzyme required relatively high concentrations of H(2)O(2) (2 mM) for optimum activity. The apparent K(m) values for ABTS, DMP, benzyl alcohol, veratryl alcohol, and H(2)O(2) were 37, 298, 1,001, 2,367 and 1,313 microM, respectively. The N-terminal amino acid sequences of the main AaP II spots blotted after two-dimensional gel electrophoresis were almost identical and exhibited almost no homology to the sequences of other peroxidases from basidiomycetes, but they shared the first three amino acids, as well as two additional amino acids, with the heme chloroperoxidase (CPO) from the ascomycete Caldariomyces fumago. This finding is consistent with the fact that AaP halogenates monochlorodimedone, the specific substrate of CPO. The existence of haloperoxidases in basidiomycetous fungi may be of general significance for the natural formation of chlorinated organic compounds in forest soils. PMID- 15294789 TI - Enhanced arsenic accumulation in engineered bacterial cells expressing ArsR. AB - The metalloregulatory protein ArsR, which offers high affinity and selectivity toward arsenite, was overexpressed in Escherichia coli in an attempt to increase the bioaccumulation of arsenic. Overproduction of ArsR resulted in elevated levels of arsenite bioaccumulation but also a severe reduction in cell growth. Incorporation of an elastin-like polypeptide as the fusion partner to ArsR (ELP153AR) improved cell growth by twofold without compromising the ability to accumulate arsenite. Resting cells overexpressing ELP153AR accumulated 5- and 60 fold-higher levels of arsenate and arsenite than control cells without ArsR overexpression. Conversely, no significant improvement in Cd(2+) or Zn(2+) accumulation was observed, validating the specificity of ArsR. The high affinity of ArsR allowed 100% removal of 50 ppb of arsenite from contaminated water with these engineered cells, providing a technology useful to comply with the newly approved U.S. Environmental Protection Agency limit of 10 ppb. These results open up the possibility of using cells overexpressing ArsR as an inexpensive, high affinity ligand for arsenic removal from contaminated drinking and ground water. PMID- 15294790 TI - Rectal administration of Escherichia coli O157:H7: novel model for colonization of ruminants. AB - Escherichia coli O157:H7 causes hemorrhagic colitis and life-threatening complications. Because healthy cattle are reservoirs for the bacterium, ruminant infection models have applications in analyzing the relationship between cattle and this human pathogen and in testing interventions to reduce or prevent bovine colonization with this bacterium. Current approaches often do not reliably mimic natural, long-term bovine colonization with E. coli O157:H7 in older calves and adult animals (ages that enter our food chain). Based on the recent identification of the bovine rectoanal junction mucosa as a site of E. coli O157:H7 colonization, we developed a novel rectal swab administration colonization model. We compared this method with oral dosing and direct contact transmission (Trojan) methods. E. coli O157:H7 carriage status was determined by fecal or rectoanal mucosa swab culture. High ( approximately 10(10) CFU) and low ( approximately 10(7) CFU) oral doses of E. coli O157:H7 in sheep and cattle resulted in variable infection with the bacterium. Some animals became colonized with the bacteria and remained culture positive for several weeks, and some animals did not become colonized and rapidly cleared the bacteria in a few days. Pen mates of E. coli O157:H7 culture-positive Trojan cattle had a low infection rate and variable colonization status. However, rectal swab administration of E. coli O157:H7 to cattle resulted in consistent long-term colonization in all animals. The surprising ease with which long-term infections resulted from a single application of bacteria to the rectoanal mucosa also strongly supported this location as a site of E. coli O157:H7 colonization in cattle. PMID- 15294791 TI - Characterization of a galactokinase-positive recombinant strain of Streptococcus thermophilus. AB - The lactic acid bacterium Streptococcus thermophilus is widely used by the dairy industry for its ability to transform lactose, the primary sugar found in milk, into lactic acid. Unlike the phylogenetically related species Streptococcus salivarius, S. thermophilus is unable to metabolize and grow on galactose and thus releases substantial amounts of this hexose into the external medium during growth on lactose. This metabolic property may result from the inability of S. thermophilus to synthesize galactokinase, an enzyme of the Leloir pathway that phosphorylates intracellular galactose to generate galactose-1-phosphate. In this work, we report the complementation of Gal(-) strain S. thermophilus SMQ-301 with S. salivarius galK, the gene that codes for galactokinase, and the characterization of recombinant strain SMQ-301K01. The recombinant strain, which was obtained by transformation of strain SMQ-301 with pTRKL2TK, a plasmid bearing S. salivarius galK, grew on galactose with a generation time of 55 min, which was almost double the generation time on lactose. Data confirmed that (i) the ability of SMQ-301K01 to grow on galactose resulted from the expression of S. salivarius galK and (ii) transcription of the plasmid-borne galK gene did not require GalR, a transcriptional regulator of the gal and lac operons, and did not interfere with the transcription of these operons. Unexpectedly, recombinant strain SMQ 301K01 still expelled galactose during growth on lactose, but only when the amount of the disaccharide in the medium exceeded 0.05%. Thus, unlike S. salivarius, the ability to metabolize galactose was not sufficient for S. thermophilus to simultaneously metabolize the glucose and galactose moieties of lactose. Nevertheless, during growth in milk and under time-temperature conditions that simulated those used to produce mozzarella cheese, the recombinant Gal(+) strain grew and produced acid more rapidly than the Gal(-) wild-type strain. PMID- 15294792 TI - Characterization of a novel plasma membrane protein, expressed in the midgut epithelia of Bombyx mori, that binds to Cry1A toxins. AB - We describe the properties of a novel 252-kDa protein (P252) isolated from brush border membranes of Bombyx mori. P252 was found in a Triton X-100-soluble brush border membrane vesicle fraction, suggesting that it may be a component of the midgut epithelial cell membrane. P252 was purified to homogeneity, and the amino acid sequence of two internal peptides was determined, but neither of the peptides matched protein sequences in the available databases. The apparent molecular mass of the purified protein was estimated by denaturing gel electrophoresis to be 252 kDa, and it migrated as a single band on native gels. However, gel filtration chromatography indicated an apparent mass of 985 kDa, suggesting that P252 may exist as a homo-oligomer. The associations of P252 with Cry1Aa, Cry1Ab, and Cry1Ac were specific, and K(d) constants were determined to be 28.9, 178.5, and 20.0 nM, respectively. A heterologous competition assay was also done. P252 did not exhibit Leu-pNA hydrolysis activity, and binding to the Cry1A toxins was not inhibited by GalNAc. Binding assays of P252 with various lectins indicated the presence of three antennal N-linked high-mannose-type as well as O-linked mucin-type sugar side chains. While the function of P252 is not yet clear, we propose that it may function with Cry1A toxins during the insecticidal response and/or Cry toxin resistance mechanism. PMID- 15294794 TI - Development and evaluation of an online CO(2) evolution test and a multicomponent biodegradation test system. AB - Well-established biodegradation tests use biogenously evolved carbon dioxide (CO(2)) as an analytical parameter to determine the ultimate biodegradability of substances. A newly developed analytical technique based on the continuous online measurement of conductivity showed its suitability over other techniques. It could be demonstrated that the method met all criteria of established biodegradation tests, gave continuous biodegradation curves, and was more reliable than other tests. In parallel experiments, only small variations in the biodegradation pattern occurred. When comparing the new online CO(2) method with existing CO(2) evolution tests, growth rates and lag periods were similar and only the final degree of biodegradation of aniline was slightly lower. A further test development was the unification and parallel measurement of all three important summary parameters for biodegradation--i.e., CO(2) evolution, determination of the biochemical oxygen demand (BOD), and removal of dissolved organic carbon (DOC)--in a multicomponent biodegradation test system (MCBTS). The practicability of this test method was demonstrated with aniline. This test system had advantages for poorly water-soluble and highly volatile compounds and allowed the determination of the carbon fraction integrated into biomass (heterotrophic yield). The integrated online measurements of CO(2) and BOD systems produced continuous degradation curves, which better met the stringent criteria of ready biodegradability (60% biodegradation in a 10-day window). Furthermore the data could be used to calculate maximal growth rates for the modeling of biodegradation processes. PMID- 15294793 TI - Treatment of Salmonella enterica serovar Enteritidis with a sublethal concentration of trisodium phosphate or alkaline pH induces thermotolerance. AB - The responses of Salmonella enterica serovar Enteritidis to a sublethal dose of trisodium phosphate (TSP) and its equivalent alkaline pH made with NaOH were examined. Pretreatment of S. enterica serovar Enteritidis cells with 1.5% TSP or pH 10.0 solutions resulted in a significant increase in thermotolerance, resistance to 2.5% TSP, resistance to high pH, and sensitivity to acid and H(2)O(2). Protein inhibition studies with chloramphenicol revealed that thermotolerance, unlike resistance to high pH, was dependent on de novo protein synthesis. Two-dimensional polyacrylamide gel electrophoresis (PAGE) of total cellular proteins from untreated control cells resolved as many as 232 proteins, of which 22 and 15% were absent in TSP- or alkaline pH-pretreated cells, respectively. More than 50% of the proteins that were either up- or down regulated by TSP pretreatment were also up- or down-regulated by alkaline pH pretreatment. Sodium dodecyl sulfate-PAGE analysis of detergent-insoluble outer membrane proteins revealed the up-regulation of at least four proteins. Mass spectrometric analysis showed the up-regulated proteins to include those involved in the transport of small hydrophilic molecules across the cytoplasmic membrane and those that act as chaperones and aid in the export of newly synthesized proteins by keeping them in open conformation. Other up-regulated proteins included common housekeeping proteins like those involved in amino acid biosynthesis, nucleotide metabolism, and aminoacyl-tRNA biosynthesis. In addition to the differential expression of proteins following TSP or alkaline pH treatment, changes in membrane fatty acid composition were also observed. Alkaline pH- or TSP-pretreated cells showed a higher saturated and cyclic to unsaturated fatty acid ratio than did the untreated control cells. These results suggest that the cytoplasmic membrane could play a significant role in the induction of thermotolerance and resistance to other stresses following TSP or alkaline pH treatment. PMID- 15294795 TI - Biodegradation of chloromethane by Pseudomonas aeruginosa strain NB1 under nitrate-reducing and aerobic conditions. AB - Pseudomonas aeruginosa strain NB1 uses chloromethane (CM) as its sole source of carbon and energy under nitrate-reducing and aerobic conditions. The observed yield of NB1 was 0.20 (+/-0.06) (mean +/- standard deviation) and 0.28 (+/-0.01) mg of total suspended solids (TSS) mg of CM(-1) under anoxic and aerobic conditions, respectively. The stoichiometry of nitrate consumption was 0.75 (+/ 0.10) electron equivalents (eeq) of NO(3)(-) per eeq of CM, which is consistent with the yield when it is expressed on an eeq basis. Nitrate was stoichiometrically converted to dinitrogen (0.51 +/- 0.05 mol of N(2) per mol of NO(3)(-)). The stoichiometry of oxygen use with CM (0.85 +/- 0.21 eeq of O(2) per eeq of CM) was also consistent with the aerobic yield. Stoichiometric release of chloride and minimal accumulation of soluble metabolic products (measured as chemical oxygen demand) following CM consumption, under anoxic and aerobic conditions, indicated complete biodegradation of CM. Acetylene did not inhibit CM use under aerobic conditions, implying that a monooxygenase was not involved in initiating aerobic CM metabolism. Under anoxic conditions, the maximum specific CM utilization rate (k) for NB1 was 5.01 (+/-0.06) micromol of CM mg of TSS(-1) day(-1), the maximum specific growth rate (micro(max)) was 0.0506 day(-1), and the Monod half-saturation coefficient (K(s)) was 0.067 (+/-0.004) microM. Under aerobic conditions, the values for k, micro(max), and K(s) were 10.7 (+/-0.11) micromol of CM mg of TSS(-1) day(-1), 0.145 day(-1), and 0.93 (+/-0.042) microM, respectively, indicating that NB1 used CM faster under aerobic conditions. Strain NB1 also grew on methanol, ethanol, and acetate under denitrifying and aerobic conditions, but not on methane, formate, or dichloromethane. PMID- 15294796 TI - Mining the microbiota of the neonatal gastrointestinal tract for conjugated linoleic acid-producing bifidobacteria. AB - This study was designed to isolate different strains of the genus Bifidobacterium from the fecal material of neonates and to assess their ability to produce the cis-9, trans-11 conjugated linoleic acid (CLA) isomer from free linoleic acid. Fecal material was collected from 24 neonates aged between 3 days and 2 months in a neonatal unit (Erinville Hospital, Cork, Ireland). A total of 46 isolates from six neonates were confirmed to be Bifidobacterium species based on a combination of the fructose-6-phosphate phosphoketolase assay, RAPD [random(ly) amplified polymorphic DNA] PCR, pulsed-field gel electrophoresis (PFGE), and partial 16S ribosomal DNA sequencing. Interestingly, only 1 of the 11 neonates that had received antibiotic treatment produced bifidobacteria. PFGE after genomic digestion with the restriction enzyme XbaI demonstrated that the bifidobacteria population displayed considerable genomic diversity among the neonates, with each containing between one and five dominant strains, whereas 11 different macro restriction patterns were obtained. In only one case did a single strain appear in two neonates. All genetically distinct strains were then screened for CLA production after 72 h of incubation with 0.5 mg of free linoleic acid ml(-1) by using gas-liquid chromatography. The most efficient producers belonged to the species Bifidobacterium breve, of which two different strains converted 29 and 27% of the free linoleic acid to the cis-9, trans-11 isomer per microgram of dry cells, respectively. In addition, a strain of Bifidobacterium bifidum showed a conversion rate of 18%/microg dry cells. The ability of some Bifidobacterium strains to produce CLA could be another human health-promoting property linked to members of the genus, given that this metabolite has demonstrated anticarcinogenic activity in vitro and in vivo. PMID- 15294797 TI - Construction of a chimeric thermostable pyrophosphatase to facilitate its purification and immobilization by using the choline-binding tag. AB - The thermophilic inorganic pyrophosphatase (Pyr) from Thermus thermophilus has been produced in Escherichia coli fused to the C terminus of the choline-binding tag (ChB tag) derived from the choline-binding domain (ChBD) of pneumococcal LytA autolysin. The chimeric ChBD-Pyr protein retains its thermostable activity and can be purified in a single step by DEAE-cellulose affinity chromatography. Pyr can be further released from the ChBD by thrombin, using the specific protease recognition site incorporated in the C terminus of this tag. Remarkably, the ChB tag provides a selective and very strong thermostable noncovalent immobilization of ChBD-Pyr in the DEAE-cellulose matrix. The binding of choline or choline analogues, such as DEAE, confers a high thermal stability to this tag; therefore, the immobilized chimeric enzyme can be assayed at high temperature without protein leakage, demonstrating the usefulness of the ChB tag for noncovalent immobilization of thermophilic proteins. Moreover, ChBD-Pyr can be purified and immobilized in a single step on commercial DEAE-cellulose paper. The affinity of the ChB tag for this versatile solid support can be very helpful in developing many biotechnological applications. PMID- 15294798 TI - Use of microautoradiography combined with fluorescence in situ hybridization to determine dimethylsulfoniopropionate incorporation by marine bacterioplankton taxa. AB - The fraction of planktonic heterotrophic bacteria capable of incorporating dissolved dimethylsulfoniopropionate (DMSP) and leucine was determined at two coastal sites by microautoradioagraphy (AU). In Gulf of Mexico seawater microcosm experiments, the proportion of prokaryotes that incorporated sulfur from [(35)S]DMSP ranged between 27 and 51% of 4',6-diamidino-2-phenylindole (DAPI) positive cells, similar to or slightly lower than the proportion incorporating [(3)H]leucine. In the northwest Mediterranean coast, the proportion of cells incorporating sulfur from [(35)S]DMSP increased from 5 to 42% from January to March, coinciding with the development of a phytoplankton bloom. At the same time, the proportion of cells incorporating [(3)H]leucine increased from 21 to 40%. The combination of AU and fluorescence in situ hybridization (FISH) revealed that the Roseobacter clade (alpha-proteobacteria) accounted for 13 to 43% of the microorganisms incorporating [(35)S]DMSP at both sampling sites. Significant uptake of sulfur from DMSP was also found among members of the gamma proteobacteria and Cytophaga-Flavobacterium groups. Roseobacter and gamma proteobacteria exhibited the highest percentage of DAPI-positive cells incorporating (35)S from DMSP (around 50%). Altogether, the application of AU with [(35)S]DMSP combined with FISH indicated that utilization of S from DMSP is a widespread feature among active marine bacteria, comparable to leucine utilization. These results point toward DMSP as an important substrate for a broad and diverse fraction of marine bacterioplankton. PMID- 15294800 TI - Effect of nematodes on rhizosphere colonization by seed-applied bacteria. AB - There is much interest in the use of seed-applied bacteria for biocontrol and biofertilization, and several commercial products are available. However, many attempts to use this strategy fail because the seed-applied bacteria do not colonize the rhizosphere. Mechanisms of rhizosphere colonization may involve active bacterial movement or passive transport by percolating water or plant roots. Transport by other soil biota is likely to occur, but this area has not been well studied. We hypothesized that interactions with soil nematodes may enhance colonization. To test this hypothesis, a series of microcosm experiments was carried out using two contrasting soils maintained under well-defined physical conditions where transport by mass water flow could not occur. Seed applied Pseudomonas fluorescens SBW25 was capable of rhizosphere colonization at matric potentials of -10 and -40 kPa in soil without nematodes, but colonization levels were substantially increased by the presence of nematodes. Our results suggest that nematodes can have an important role in rhizosphere colonization by bacteria in soil. PMID- 15294799 TI - Genetic diversity of Escherichia coli isolated from urban rivers and beach water. AB - Repetitive element anchored PCR was used to evaluate the genetic profiles of Escherichia coli isolated from surface water contaminated with urban stormwater, sanitary sewage, and gull feces to determine if strains found in environmental samples reflect the strain composition of E. coli obtained from host sources. Overall, there was less diversity in isolates collected from river and beach sites than with isolates obtained from human and nonhuman sources. Unique strain types comprised 28.8, 29.2, and 15.0% of the isolate data sets recovered from stormwater, river water, and beach water, respectively. In contrast, 50.4% of gull isolates and 41.2% of sewage isolates were unique strain types. River water, which is expected to contain E. coli strains from many diffuse sources of nonpoint source pollution, contained strains most closely associated with other river water isolates that were collected at different sites or on different days. However, river sites impacted by sewage discharge had approximately 20% more strains similar to sewage isolates than did sites impacted by stormwater alone. Beach sites with known gull fecal contamination contained E. coli most similar to other beach isolates rather than gull isolates collected at these same sites, indicating underrepresentation of possible gull strains. These results suggest large numbers of strains are needed to represent contributing host sources within a geographical location. Additionally, environmental survival may influence the composition of strains that can be recovered from contaminated waters. Understanding the ecology of indicator bacteria is important when interpreting fecal pollution assessments and developing source detection methodology. PMID- 15294801 TI - Mutational analysis of mesentericin y105, an anti-Listeria bacteriocin, for determination of impact on bactericidal activity, in vitro secondary structure, and membrane interaction. AB - Mesentericin Y105 is a 37-residue bacteriocin produced by Leuconostoc mesenteroides Y105 that displays antagonistic activity against gram-positive bacteria such as Enterococcus faecalis and Listeria monocytogenes. It is closely related to leucocin A, an antimicrobial peptide containing beta-sheet and alpha helical structures. To analyze structure-function relationships and the mode of action of this bacteriocin, we generated a collection of mesentericin derivatives. Mutations were obtained mostly by PCR random mutagenesis, and the peptides were produced by an original system of heterologous expression recently described. Ten derivatives were obtained displaying modifications at eight different positions in the mesentericin Y105 sequence. Purified peptides were incorporated into lysophosphatidylcholine micelles and analyzed by circular dichroism. The alpha-helical contents of these peptides were compared and related to their respective bactericidal activities. Moreover, studies of the intrinsic fluorescence of tryptophan residues naturally occurring at positions 18 and 37 revealed information about insertion of the peptides in micelles. A model for the mode of action of mesentericin Y105 and related bacteriocins is proposed. PMID- 15294802 TI - Altering the substrate specificity of organophosphorus hydrolase for enhanced hydrolysis of chlorpyrifos. AB - Chlorpyrifos is one of the most popular pesticides used for agriculture crop protection, and widespread contamination is a potential concern. However, chlorpyrifos is hydrolyzed almost 1,000-fold slower than the preferred substrate, paraoxon, by organophosphorus hydrolase (OPH), an enzyme that can degrade a broad range of organophosphate pesticides. We have recently demonstrated that directed evolution can be used to generate OPH variants with up to 25-fold improvement in hydrolysis of methyl parathion. The obvious question and challenge are whether similar success could be achieved with this poorly hydrolyzed substrate, chlorpyrifos. For this study, five improved variants were selected from two rounds of directed evolution based on the formation of clear haloes on Luria Bertani plates overlaid with chlorpyrifos. One variant, B3561, exhibited a 725 fold increase in the k(cat)/K(m) value for chlorpyrifos hydrolysis as well as enhanced hydrolysis rates for several other OP compounds tested. Considering that wild-type OPH hydrolyzes paraoxon at a rate close to the diffusion control limit, the 39-fold improvement in hydrolysis of paraoxon by B3561 suggests that this variant is one of the most efficient enzymes available to attack a wide spectrum of organophosphate nerve agents. PMID- 15294803 TI - Genetic instability of heterozygous, hybrid, natural wine yeasts. AB - We describe a genetic instability found in natural wine yeasts but not in the common laboratory strains of Saccharomyces cerevisiae. Spontaneous cyh2(R)/cyh2(R) mutants resistant to high levels of cycloheximide can be directly isolated from cyh2(S)/cyh2(S) wine yeasts. Heterozygous cyh2(R)/cyh2(S) hybrid clones vary in genetic instability as measured by loss of heterozygosity at cyh2. There were two main classes of hybrids. The lawn hybrids have high genetic instability and generally become cyh2(R)/cyh2(R) homozygotes and lose the killer phenotype under nonselective conditions. The papilla hybrids have a much lower rate of loss of heterozygosity and maintain the killer phenotype. The genetic instability in lawn hybrids is 3 to 5 orders of magnitude greater than the highest loss-of-heterozygosity rates previously reported. Molecular mechanisms such as DNA repair by break-induced replication might account for the asymmetrical loss of heterozygosity. This loss-of-heterozygosity phenomenon could be economically important if it causes sudden phenotype changes in industrial or pathogenic yeasts and of more basic importance to the degree that it influences the evolution of naturally occurring yeast populations. PMID- 15294804 TI - Chemotaxis of Silicibacter sp. strain TM1040 toward dinoflagellate products. AB - The alpha-proteobacteria phylogenetically related to the Roseobacter clade are predominantly responsible for the degradation of organosulfur compounds, including the algal osmolyte dimethylsulfoniopropionate (DMSP). Silicibacter sp. strain TM1040, isolated from a DMSP-producing Pfiesteria piscicida dinoflagellate culture, degrades DMSP, producing 3-methylmercaptopropionate. TM1040 possesses three lophotrichous flagella and is highly motile, leading to a hypothesis that TM1040 interacts with P. piscicida through a chemotactic response to compounds produced by its dinoflagellate host. A combination of a rapid chemotaxis screening assay and a quantitative capillary assay were used to measure chemotaxis of TM1040. These bacteria are highly attracted to dinoflagellate homogenates; however, the response decreases when homogenates are preheated to 80 degrees C. To help identify the essential attractant molecules within the homogenates, a series of pure compounds were tested for their ability to serve as attractants. The results show that TM1040 is strongly attracted to amino acids and DMSP metabolites, while being only mildly responsive to sugars and the tricarboxylic acid cycle intermediates. Adding pure DMSP, methionine, or valine to the chemotaxis buffer resulted in a decreased response to the homogenates, indicating that exogenous addition of these chemicals blocks chemotaxis and suggesting that DMSP and amino acids are essential attractant molecules in the dinoflagellate homogenates. The implication of Silicibacter sp. strain TM1040 chemotaxis in establishing and maintaining its interaction with P. piscicida is discussed. PMID- 15294805 TI - Expression of a heterologous manganese superoxide dismutase gene in intestinal lactobacilli provides protection against hydrogen peroxide toxicity. AB - In living organisms, exposure to oxygen provokes oxidative stress. A widespread mechanism for protection against oxidative stress is provided by the antioxidant enzymes: superoxide dismutases (SODs) and hydroperoxidases. Generally, these enzymes are not present in Lactobacillus spp. In this study, we examined the potential advantages of providing a heterologous SOD to some of the intestinal lactobacilli. Thus, the gene encoding the manganese-containing SOD (sodA) was cloned from Streptococcus thermophilus AO54 and expressed in four intestinal lactobacilli. A 1.2-kb PCR product containing the sodA gene was cloned into the shuttle vector pTRK563, to yield pSodA, which was functionally expressed and complemented an Escherichia coli strain deficient in Mn and FeSODs. The plasmid, pSodA, was subsequently introduced and expressed in Lactobacillus gasseri NCK334, Lactobacillus johnsonii NCK89, Lactobacillus acidophilus NCK56, and Lactobacillus reuteri NCK932. Molecular and biochemical analyses confirmed the presence of the gene (sodA) and the expression of an active gene product (MnSOD) in these strains of lactobacilli. The specific activities of MnSOD were 6.7, 3.8, 5.8, and 60.7 U/mg of protein for L. gasseri, L. johnsonii, L. acidophilus, and L. reuteri, respectively. The expression of S. thermophilus MnSOD in L. gasseri and L. acidophilus provided protection against hydrogen peroxide stress. The data show that MnSOD protects cells against hydrogen peroxide by removing O(2)(.-) and preventing the redox cycling of iron. To our best knowledge, this is the first report of a sodA from S. thermophilus being expressed in other lactic acid bacteria. PMID- 15294806 TI - Identification of an Na(+)-dependent transporter associated with saxitoxin producing strains of the cyanobacterium Anabaena circinalis. AB - Blooms of the freshwater cyanobacterium Anabaena circinalis are recognized as an important health risk worldwide due to the production of a range of toxins such as saxitoxin (STX) and its derivatives. In this study we used HIP1 octameric palindrome repeated-sequence PCR to compare the genomic structure of phylogenetically similar Australian isolates of A. circinalis. STX-producing and nontoxic cyanobacterial strains showed different HIP1 (highly iterated octameric palindrome 1) DNA patterns, and characteristic interrepeat amplicons for each group were identified. Suppression subtractive hybridization (SSH) was performed using HIP1 PCR-generated libraries to further identify toxic-strain-specific genes. An STX-producing strain and a nontoxic strain of A. circinalis were chosen as testers in two distinct experiments. The two categories of SSH putative tester specific sequences were characterized by different families of encoded proteins that may be representative of the differences in metabolism between STX-producing and nontoxic A. circinalis strains. DNA-microarray hybridization and genomic screening revealed a toxic-strain-specific HIP1 fragment coding for a putative Na(+)-dependent transporter. Analysis of this gene demonstrated analogy to the mrpF gene of Bacillus subtilis, whose encoded protein is involved in Na(+) specific pH homeostasis. The application of this gene as a molecular probe in laboratory and environmental screening for STX-producing A. circinalis strains was demonstrated. The possible role of this putative Na(+)-dependent transporter in the toxic cyanobacterial phenotype is also discussed, in light of recent physiological studies of STX-producing cyanobacteria. PMID- 15294807 TI - Enhanced anaerobic biodegradation of benzene-toluene-ethylbenzene-xylene-ethanol mixtures in bioaugmented aquifer columns. AB - Methanogenic flowthrough aquifer columns were used to investigate the potential of bioaugmentation to enhance anaerobic benzene-toluene-ethylbenzene-xylene (BTEX) degradation in groundwater contaminated with ethanol-blended gasoline. Two different methanogenic consortia (enriched with benzene or toluene and o-xylene) were used as inocula. Toluene was the only hydrocarbon degraded within 3 years in columns that were not bioaugmented, although anaerobic toluene degradation was observed after only 2 years of acclimation. Significant benzene biodegradation (up to 88%) was observed only in a column bioaugmented with the benzene-enriched methanogenic consortium, and this removal efficiency was sustained for 1 year with no significant decrease in permeability due to bioaugmentation. Benzene removal was hindered by the presence of toluene, which is a more labile substrate under anaerobic conditions. Real-time quantitative PCR analysis showed that the highest numbers of bssA gene copies (coding for benzylsuccinate synthase) occurred in aquifer samples exhibiting the highest rate of toluene degradation, which suggests that this gene could be a useful biomarker for environmental forensic analysis of anaerobic toluene bioremediation potential. bssA continued to be detected in the columns 1 year after column feeding ceased, indicating the robustness of the added catabolic potential. Overall, these results suggest that anaerobic bioaugmentation might enhance the natural attenuation of BTEX in groundwater contaminated with ethanol-blended gasoline, although field trials would be needed to demonstrate its feasibility. This approach may be especially attractive for removing benzene, which is the most toxic and commonly the most persistent BTEX compound under anaerobic conditions. PMID- 15294808 TI - Detection and quantification of the red tide dinoflagellate Karenia brevis by real-time nucleic acid sequence-based amplification. AB - Nucleic acid sequence-based amplification (NASBA) is an isothermal method of RNA amplification that has been previously used in clinical diagnostic testing. A real-time NASBA assay has been developed for the detection of rbcL mRNA from the red tide dinoflagellate Karenia brevis. This assay is sensitive to one K. brevis cell and 1.0 fg of in vitro transcript, with occasional detection of lower concentrations of transcript. The assay did not detect rbcL mRNA from a wide range of nontarget organisms and environmental clones, while 10 strains (all tested) of K. brevis were detected. By the use of standard curves based on time to positivity, concentrations of K. brevis in environmental samples were predicted by NASBA and classified into different levels of blooms per the Florida Fish and Wildlife Conservation Commission (FWC) system. NASBA classification matched FWC classification (based on cell counts) 72% of the time. Those samples that did not match were off by only one class. NASBA is sensitive, rapid, and effective and may be used as an additional or alternative method to detect and quantify K. brevis in the marine environment. PMID- 15294809 TI - veA is required for toxin and sclerotial production in Aspergillus parasiticus. AB - It was long been noted that secondary metabolism is associated with fungal development. In Aspergillus nidulans, conidiation and mycotoxin production are linked by a G protein signaling pathway. Also in A. nidulans, cleistothecial development and mycotoxin production are controlled by a gene called veA. Here we report the characterization of a veA ortholog in the aflatoxin-producing fungus A. parasiticus. Cleistothecia are not produced by Aspergillus parasiticus; instead, this fungus produces spherical structures called sclerotia that allow for survival under adverse conditions. Deletion of veA from A. parasiticus resulted in the blockage of sclerotial formation as well as a blockage in the production of aflatoxin intermediates. Our results indicate that A. parasiticus veA is required for the expression of aflR and aflJ, which regulate the activation of the aflatoxin gene cluster. In addition to these findings, we observed that deletion of veA reduced conidiation both on the culture medium and on peanut seed. The fact that veA is necessary for conidiation, production of resistant structures, and aflatoxin biosynthesis makes veA a good candidate gene to control aflatoxin biosynthesis or fungal development and in this way to greatly decrease its devastating impact on health and the economy. PMID- 15294810 TI - Determination of the efficacy of two building decontamination strategies by surface sampling with culture and quantitative PCR analysis. AB - The efficacy of currently available decontamination strategies for the treatment of indoor furnishings contaminated with bioterrorism agents is poorly understood. Efficacy testing of decontamination products in a controlled environment is needed to ensure that effective methods are used to decontaminate domestic and workplace settings. An experimental room supplied with materials used in office furnishings (i.e., wood laminate, painted metal, and vinyl tile) was used with controlled dry aerosol releases of endospores of Bacillus atrophaeus ("Bacillus subtilis subsp. niger," also referred to as BG), a Bacillus anthracis surrogate. Studies were performed using two test products, a foam decontaminant and chlorine dioxide gas. Surface samples were collected pre- and posttreatment with three sampling methods and analyzed by culture and quantitative PCR (QPCR). Additional aerosol releases with environmental background present on the surface materials were also conducted to determine if there was any interference with decontamination or sample analysis. Culture results indicated that 10(5) to 10(6) CFU per sample were present on surfaces before decontamination. After decontamination with the foam, no culturable B. atrophaeus spores were detected. After decontamination with chlorine dioxide gas, no culturable B. atrophaeus was detected in 24 of 27 samples (89%). However, QPCR analysis showed that B. atrophaeus DNA was still present after decontamination with both methods. Environmental background material had no apparent effect on decontamination, but inhibition of the QPCR assay was observed. These results demonstrate the effectiveness of two decontamination methods and illustrate the utility of surface sampling and QPCR analysis for the evaluation of decontamination strategies. PMID- 15294811 TI - New strategies for cultivation and detection of previously uncultured microbes. AB - An integrative approach was used to obtain pure cultures of previously uncultivated members of the divisions Acidobacteria and Verrucomicrobia from agricultural soil and from the guts of wood-feeding termites. Some elements of the cultivation procedure included the following: the use of agar media with little or no added nutrients; relatively long periods of incubation (more than 30 days); protection of cells from exogenous peroxides; and inclusion of humic acids or a humic acid analogue (anthraquinone disulfonate) and quorum-signaling compounds (acyl homoserine lactones) in growth media. The bacteria were incubated in the presence of air and in hypoxic (1 to 2% O(2) [vol/vol]) and anoxic atmospheres. Some bacteria were incubated with elevated concentrations of CO(2) (5% [vol/vol]). Significantly more Acidobacteria were found on isolation plates that had been incubated with 5% CO(2). A simple, high-throughput, PCR-based surveillance method (plate wash PCR) was developed. This method greatly facilitated detection and ultimate isolation of target bacteria from as many as 1,000 colonies of nontarget microbes growing on the same agar plates. Results illustrate the power of integrating culture methods with molecular techniques to isolate bacteria from phylogenetic groups underrepresented in culture. PMID- 15294812 TI - Determining rates of change and evaluating group-level resiliency differences in hyporheic microbial communities in response to fluvial heavy-metal deposition. AB - Prior field studies by our group have demonstrated a relationship between fluvial deposition of heavy metals and hyporheic-zone microbial community structure. Here, we determined the rates of change in hyporheic microbial communities in response to heavy-metal contamination and assessed group-level differences in resiliency in response to heavy metals. A controlled laboratory study was performed using 20 flowthrough river mesocosms and a repeated-measurement factorial design. A single hyporheic microbial community was exposed to five different levels of an environmentally relevant metal treatment (0, 4, 8, 16, and 30% sterilized contaminated sediments). Community-level responses were monitored at 1, 2, 4, 8, and 12 weeks via denaturing gradient gel electrophoresis and quantitative PCR using group-specific primer sets for indigenous populations most closely related to the alpha-, beta-, and gamma-proteobacteria. There was a consistent, strong curvilinear relationship between community composition and heavy-metal contamination (R(2) = 0.83; P < 0.001), which was evident after only 7 days of metal exposure (i.e., short-term response). The abundance of each phylogenetic group was negatively affected by the heavy-metal treatments; however, each group recovered from the metal treatments to a different extent and at a unique rate during the course of the experiment. The structure of hyporheic microbial communities responded rapidly and at contamination levels an order of magnitude lower than those shown to elicit a response in aquatic macroinvertebrate assemblages. These studies indicate that hyporheic microbial communities are a sensitive and useful indicator of heavy-metal contamination in streams. PMID- 15294813 TI - Enhanced mineralization of [U-(14)C]2,4-dichlorophenoxyacetic acid in soil from the rhizosphere of Trifolium pratense. AB - Enhanced biodegradation in the rhizosphere has been reported for many organic xenobiotic compounds, although the mechanisms are not fully understood. The purpose of this study was to discover whether rhizosphere-enhanced biodegradation is due to selective enrichment of degraders through growth on compounds produced by rhizodeposition. We monitored the mineralization of [U-(14)C]2,4 dichlorophenoxyacetic acid (2,4-D) in rhizosphere soil with no history of herbicide application collected over a period of 0 to 116 days after sowing of Lolium perenne and Trifolium pratense. The relationships between the mineralization kinetics, the number of 2,4-D degraders, and the diversity of genes encoding 2,4-D/alpha-ketoglutarate dioxygenase (tfdA) were investigated. The rhizosphere effect on [(14)C]2,4-D mineralization (50 microg g(-1)) was shown to be plant species and plant age specific. In comparison with nonplanted soil, there were significant (P < 0.05) reductions in the lag phase and enhancements of the maximum mineralization rate for 25- and 60-day T. pratense soil but not for 116-day T. pratense rhizosphere soil or for L. perenne rhizosphere soil of any age. Numbers of 2,4-D degraders in planted and nonplanted soil were low (most probable number, <100 g(-1)) and were not related to plant species or age. Single strand conformational polymorphism analysis showed that plant species had no impact on the diversity of alpha-Proteobacteria tfdA-like genes, although an impact of 2,4-D application was recorded. Our results indicate that enhanced mineralization in T. pratense rhizosphere soil is not due to enrichment of 2,4-D degrading microorganisms by rhizodeposits. We suggest an alternative mechanism in which one or more components of the rhizodeposits induce the 2,4-D pathway. PMID- 15294814 TI - Uptake of the beta-lactam precursor alpha-aminoadipic acid in Penicillium chrysogenum is mediated by the acidic and the general amino acid permease. AB - External addition of the beta-lactam precursor alpha-aminoadipic acid to the filamentous fungus Penicillium chrysogenum leads to an increased intracellular alpha-aminoadipic acid concentration and an increase in penicillin production. The exact route for alpha-aminoadipic acid uptake is not known, although the general amino acid and acidic amino acid permeases have been implicated in this process. Their corresponding genes, PcGAP1 and PcDIP5, of P. chrysogenum were cloned and functionally expressed in a mutant of Saccharomyces cerevisiae (M4276) in which the acidic amino acid and general amino acid permease genes (DIP5 and GAP1, respectively) are disrupted. Transport assays show that both PcGap1 and PcDip5 mediated the uptake of alpha-aminoadipic acid, although PcGap1 showed a higher affinity for alpha-aminoadipic acid than PcDip5 (K(m) values, 230 and 800 microM, respectively). Leucine strongly inhibits alpha-aminoadipic acid transport via PcGap1 but not via PcDip5. This difference was exploited to estimate the relative contribution of each transport system to the alpha-aminoadipic acid flux in beta-lactam-producing P. chrysogenum. The transport measurements demonstrate that both PcGap1 and PcDip5 contribute to the alpha-aminoadipic acid flux. PMID- 15294815 TI - Characterization of two Bacillus thuringiensis genes identified by in vivo screening of virulence factors. AB - Bacillus thuringiensis vegetative cells are known to be highly pathogenic when injected into the hemocoel of susceptible insect larvae. This pathogenicity is due to the capacity of B. thuringiensis to cause septicemia in the host. We screened a B. thuringiensis mini-Tn10 insertion library for loss of virulence against Bombyx mori larvae on injection into the hemocoel. Three clones with attenuated virulence were isolated, corresponding to two different mini-Tn10 insertions mapping to the yqgB/yqfZ locus. Single disruptions of the yqgB and yqfZ genes did not affect virulence against B. mori. In contrast, the inactivation of both genes simultaneously reproduced the effect of the mini-Tn10 insertion and resulted in a significant delay to infection. The double DeltayqgB DeltayqfZ mutant was also nonmotile, and its growth was affected at 25 degrees C. We analyzed lacZ transcriptional fusions and detected promoter activity upstream from yqgB at 25 and 37 degrees C. Overall, our findings suggest that the yqgB and yqfZ genes encode adaptive factors that may act in synergy, enabling the bacteria to cope with the physical environment in vivo, facilitating colonization of the host. PMID- 15294816 TI - Acid resistance systems required for survival of Escherichia coli O157:H7 in the bovine gastrointestinal tract and in apple cider are different. AB - Escherichia coli O157:H7 is a highly acid-resistant food-borne pathogen that survives in the bovine and human gastrointestinal tracts and in acidic foods such as apple cider. This property is thought to contribute to the low infectious dose of the organism. Three acid resistance (AR) systems are expressed in stationary phase cells. AR system 1 is sigma(S) dependent, while AR systems 2 and 3 are glutamate and arginine dependent, respectively. In this study, we sought to determine which AR systems are important for survival in acidic foods and which are required for survival in the bovine intestinal tract. Wild-type and mutant E. coli O157:H7 strains deficient in AR system 1, 2, or 3 were challenged with apple cider and inoculated into calves. Wild-type cells, adapted at pH 5.5 in the absence of glucose (AR system 1 induced), survived well in apple cider. Conversely, the mutant deficient in AR system 1, shown previously to survive poorly in calves, was susceptible to apple cider (pH 3.5), and this sensitivity was shown to be caused by low pH. Interestingly, the AR system 2-deficient mutant survived in apple cider at high levels, but its shedding from calves was significantly decreased compared to that of wild-type cells. AR system 3 deficient cells survived well in both apple cider and calves. Taken together, these results indicate that E. coli O157:H7 utilizes different acid resistance systems based on the type of acidic environment encountered. PMID- 15294817 TI - Comparisons of different hypervariable regions of rrs genes for use in fingerprinting of microbial communities by PCR-denaturing gradient gel electrophoresis. AB - Denaturing gradient gel electrophoresis (DGGE) has become a widely used tool to examine microbial diversity and community structure, but no systematic comparison has been made of the DGGE profiles obtained when different hypervariable (V) regions are amplified from the same community DNA samples. We report here a study to make such comparisons and establish a preferred choice of V region(s) to examine by DGGE, when community DNA extracted from samples of digesta is used. When the members of the phylogenetically representative set of 218 rrs genes archived in the RDP II database were compared, the V1 region was found to be the most variable, followed by the V9 and V3 regions. The temperature of the lowest melting-temperature (T(m(L))) domain for each V region was also calculated for these rrs genes, and the V1 to V4 region was found to be most heterogeneous with respect to T(m(L)). The average T(m(L)) values and their standard deviations for each V region were then used to devise the denaturing gradients suitable for separating 95% of all the sequences, and the PCR-DGGE profiles produced from the same community DNA samples with these conditions were compared. The resulting DGGE profiles were substantially different in terms of the number, resolution, and relative intensity of the amplification products. The DGGE profiles of the V3 region were best, and the V3 to V5 and V6 to V8 regions produced better DGGE profiles than did other multiple V-region amplicons. Introduction of degenerate bases in the primers used to amplify the V1 or V3 region alone did not improve DGGE banding profiles. Our results show that DGGE analysis of gastrointestinal microbiomes is best accomplished by the amplification of either the V3 or V1 region of rrs genes, but if a longer amplification product is desired, then the V3 to V5 or V6 to V8 region should be targeted. PMID- 15294818 TI - Effects of different spices used in production of fermented sausages on growth of and curvacin A production by Lactobacillus curvatus LTH 1174. AB - Lactobacillus curvatus LTH 1174, a fermented sausage isolate, produces the listericidal bacteriocin curvacin A. The effect of different spices relevant for the production of fermented sausages was investigated in vitro through laboratory fermentations with a meat simulation medium and an imposed pH profile relevant for Belgian-type fermented sausages. The influence on the growth characteristics and especially on the kinetics of curvacin A production with L. curvatus LTH 1174 was evaluated. Pepper, nutmeg, rosemary, mace, and garlic all decreased the maximum specific growth rate, while paprika was the only spice that increased it. The effect on the lag phase was minor except for nutmeg and especially for garlic, which increased it, yet garlic was stimulatory for biomass production. The maximum attainable biomass concentration (X(max)) was severely decreased by the addition of 0.40% (wt/vol) nutmeg, while 0.35% (wt/vol) garlic or 0.80% (wt/vol) white pepper increased X(max). Nutmeg decreased both growth and bacteriocin production considerably. Garlic was the only spice enhancing specific bacteriocin production, resulting in higher bacteriocin activity in the cell-free culture supernatant. Finally, lactic acid production was stimulated by the addition of pepper, and this was not due to the manganese present because an amount of manganese that was not growth limiting was added to the growth medium. Addition of spices to the sausage mixture is clearly a factor that will influence the effectiveness of bacteriocinogenic starter cultures in fermented-sausage manufacturing. PMID- 15294819 TI - Overexpression of Lactobacillus casei D-hydroxyisocaproic acid dehydrogenase in cheddar cheese. AB - Metabolism of aromatic amino acids by lactic acid bacteria is an important source of off-flavor compounds in Cheddar cheese. Previous work has shown that alpha keto acids produced from Trp, Tyr, and Phe by aminotransferase enzymes are chemically labile and may degrade spontaneously into a variety of off-flavor compounds. However, dairy lactobacilli can convert unstable alpha-keto acids to more-stable alpha-hydroxy acids via the action of alpha-keto acid dehydrogenases such as d-hydroxyisocaproic acid dehydrogenase. To further characterize the role of this enzyme in cheese flavor, the Lactobacillus casei d-hydroxyisocaproic acid dehydrogenase gene was cloned into the high-copy-number vector pTRKH2 and transformed into L. casei ATCC 334. Enzyme assays confirmed that alpha-keto acid dehydrogenase activity was significantly higher in pTRKH2:dhic transformants than in wild-type cells. Reduced-fat Cheddar cheeses were made with Lactococcus lactis starter only, starter plus L. casei ATCC 334, and starter plus L. casei ATCC 334 transformed with pTRKH2:dhic. After 3 months of aging, the cheese chemistry and flavor attributes were evaluated instrumentally by gas chromatography-mass spectrometry and by descriptive sensory analysis. The culture system used significantly affected the concentrations of various ketones, aldehydes, alcohols, and esters and one sulfur compound in cheese. Results further indicated that enhanced expression of d-hydroxyisocaproic acid dehydrogenase suppressed spontaneous degradation of alpha-keto acids, but sensory work indicated that this effect retarded cheese flavor development. PMID- 15294820 TI - Biogeography, evolution, and diversity of epibionts in phototrophic consortia. AB - Motile phototrophic consortia are highly regular associations in which numerous cells of green sulfur bacteria surround a flagellated colorless beta proteobacterium in the center. To date, seven different morphological types of such consortia have been described. In addition, two immotile associations involving green sulfur bacteria are known. By employing a culture-independent approach, different types of phototrophic consortia were mechanically isolated by micromanipulation from 14 freshwater environments, and partial 16S rRNA gene sequences of the green sulfur bacterial epibionts were determined. In the majority of the lakes investigated, different types of phototrophic consortia were found to co-occur. In all cases, phototrophic consortia with the same morphology from the same habitat contained only a single epibiont phylotype. However, morphologically indistinguishable phototrophic consortia collected from different lakes contained different epibionts. Overall, 19 different types of epibionts were detected in the present study. Whereas the epibionts within one geographic region were very similar (Dice coefficient, 0.582), only two types of epibionts were found to occur on both the European and North American continents (Dice coefficient, 0.190). None of the epibiont 16S rRNA gene sequences have been detected so far in free-living green sulfur bacteria, suggesting that the interaction between epibionts and chemotrophic bacteria in the phototrophic consortia is an obligate interaction. Based on our phylogenetic analysis, the epibiont sequences are not monophyletic. Thus, the ability to form symbiotic associations either arose independently from different ancestors or was present in a common ancestor prior to the radiation of green sulfur bacteria and the transition to the free-living state in independent lineages. The present study thus demonstrates that there is great diversity and nonrandom geographical distribution of phototrophic consortia in the natural environment. PMID- 15294821 TI - Ecological significance of microdiversity: identical 16S rRNA gene sequences can be found in bacteria with highly divergent genomes and ecophysiologies. AB - A combination of cultivation-based methods with a molecular biological approach was used to investigate whether planktonic bacteria with identical 16S rRNA gene sequences can represent distinct eco- and genotypes. A set of 11 strains of Brevundimonas alba were isolated from a bacterial freshwater community by conventional plating or by using a liquid most-probable-number (MPN) dilution series. These strains had identical 16S rRNA gene sequences and represented the dominant phylotype in the plateable fraction, as well as in the highest positive dilutions of the MPN series. However, internally transcribed spacer and enterobacterial repetitive intergenic consensus PCR fingerprinting analyses, as well as DNA-DNA hybridization analyses, revealed great genetic diversity among the 11 strains. Each strain utilized a specific combination of 59 carbon substrates, and the niche overlap indices were low, suggesting that each strain occupied a different ecological niche. In dialysis cultures incubated in situ, each strain had a different growth rate and cell yield. We thus demonstrated that the B. alba strains represent distinct populations with genetically determined adaptations and probably occupy different ecological niches. Our results have implications for assessment of the diversity and biogeography of bacteria and increase the perception of natural diversity beyond the level of 16S rRNA gene sequences. PMID- 15294822 TI - Protein folding failure sets high-temperature limit on growth of phage P22 in Salmonella enterica serovar Typhimurium. AB - The high-temperature limit for growth of microorganisms differs greatly depending on their species and habitat. The importance of an organism's ability to manage thermal stress is reflected in the ubiquitous distribution of the heat shock chaperones. Although many chaperones function to reduce protein folding defects, it has been difficult to identify the specific protein folding pathways that set the high-temperature limit of growth for a given microorganism. We have investigated this for a simple system, phage P22 infection of Salmonella enterica serovar Typhimurium. Production of infectious particles exhibited a broad maximum of 150 phage per cell when host cells were grown at between 30 and 39 degrees C in minimal medium. Production of infectious phage declined sharply in the range of 40 to 41 degrees C, and at 42 degrees C, production had fallen to less than 1% of the maximum rate. The host cells maintained optimal division rates at these temperatures. The decrease in phage infectivity was steeper than the loss of physical particles, suggesting that noninfectious particles were formed at higher temperatures. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed a decrease in the tailspike adhesins assembled on phage particles purified from cultures incubated at higher temperatures. The infectivity of these particles was restored by in vitro incubation with soluble tailspike trimers. Examination of tailspike folding and assembly in lysates of phage-infected cells confirmed that the fraction of polypeptide chains able to reach the native state in vivo decreased with increasing temperature, indicating a thermal folding defect rather than a particle assembly defect. Thus, we believe that the folding pathway of the tailspike adhesin sets the high-temperature limit for P22 formation in Salmonella serovar Typhimurium. PMID- 15294823 TI - Influence of humic substances on bacterial and viral dynamics in freshwaters. AB - Bacterial and viral abundances were measured in 24 lakes with dissolved organic carbon (DOC) concentrations ranging from 3 to 19 mg of C liter(-1). In addition, a laboratory experiment was performed to test the effects of different sources of carbon (i.e., glucose and fulvic acids) and nutrients on the dynamics of viruses and bacteria. In the lake survey, no correlation was found between virus abundance and DOC concentration, yet there was a significant positive correlation between bacterial abundance and DOC concentration. A negative correlation was found between the virus-to-bacteria ratio and DOC level. These results are in agreement with our findings in the laboratory, where virus counts were significantly lower in treatments with fulvic acid additions than in a control (mean, 67.4% +/- 6.5% of the control). Virus counts did not differ significantly among the control and treatments with glucose, indicating that it was the type of organic carbon and not quantity which had an impact on viruses. Results from this study suggest that the way viruses control bacterial assemblages in humic lakes is different from the mechanism in clear water systems. PMID- 15294824 TI - Biodegradation of chlorpyrifos by enterobacter strain B-14 and its use in bioremediation of contaminated soils. AB - Six chlorpyrifos-degrading bacteria were isolated from an Australian soil and compared by biochemical and molecular methods. The isolates were indistinguishable, and one (strain B-14) was selected for further analysis. This strain showed greatest similarity to members of the order Enterobacteriales and was closest to members of the Enterobacter asburiae group. The ability of the strain to mineralize chlorpyrifos was investigated under different culture conditions, and the strain utilized chlorpyrifos as the sole source of carbon and phosphorus. Studies with ring or uniformly labeled [(14)C]chlorpyrifos in liquid culture demonstrated that the isolate hydrolyzed chlorpyrifos to diethylthiophospshate (DETP) and 3, 5, 6-trichloro-2-pyridinol, and utilized DETP for growth and energy. The isolate was found to possess mono- and diphosphatase activities along with a phosphotriesterase activity. Addition of other sources of carbon (glucose and succinate) resulted in slowing down of the initial rate of degradation of chlorpyrifos. The isolate degraded the DETP-containing organophosphates parathion, diazinon, coumaphos, and isazofos when provided as the sole source of carbon and phosphorus, but not fenamiphos, fonofos, ethoprop, and cadusafos, which have different side chains. Studies of the molecular basis of degradation suggested that the degrading ability could be polygenic and chromosome based. Further studies revealed that the strain possessed a novel phosphotriesterase enzyme system, as the gene coding for this enzyme had a different sequence from the widely studied organophosphate-degrading gene (opd). The addition of strain B-14 (10(6) cells g(-1)) to soil with a low indigenous population of chlorpyrifos-degrading bacteria treated with 35 mg of chlorpyrifos kg(-1) resulted in a higher degradation rate than was observed in noninoculated soils. These results highlight the potential of this bacterium to be used in the cleanup of contaminated pesticide waste in the environment. PMID- 15294825 TI - Characterization of coliphage PR772 and evaluation of its use for virus filter performance testing. AB - Virus filtration is a key clearance unit operation in the manufacture of recombinant protein, monoclonal antibody, and plasma-derived biopharmaceuticals. Recently, a consensus has developed among filter manufacturers and end users about the desirability of a common nomenclature and a standardized test for classifying and identifying virus-retentive filters. The Parenteral Drug Association virus filter task force has chosen PR772 as the model bacteriophage to standardize nomenclature for large-pore-size virus-retentive filters (filters designed to retain viruses larger than 50 to 60 nm in size). Previously, the coliphage PR772 (Tectiviridae family) has been used in some filtration studies as a surrogate for mammalian viruses of around 50 to 60 nm. In this report, we describe specific properties of PR772 critical to the support of its use for the standardization of virus filters. The complete genomic sequence of virulent phage PR772 was determined. Its genome contains 14,946 bp with an overall G+C content of 48.3 mol%, and 32 open reading frames of at least 40 codons. Comparison of the PR772 nucleotide sequence with the genome of Tectiviridae family prototype phage PRD1 revealed 97.2% identity at the DNA level. By dynamic light-scattering analysis, its hydrodynamic diameter was measured as 82 +/- 6 nm, consistent with use in testing large-virus-retentive filters. Finally, dynamic light-scattering analysis of PR772 preparations purified on CsCl gradients showed that the phage preparations are largely monodispersed. In summary, PR772 appears to be an appropriate model bacteriophage for standardization of nomenclature for larger pore-size virus-retentive filters. PMID- 15294826 TI - Cloning and characterization of three fatty alcohol oxidase genes from Candida tropicalis strain ATCC 20336. AB - Candida tropicalis (ATCC 20336) converts fatty acids to long-chain dicarboxylic acids via a pathway that includes among other reactions the oxidation of omega hydroxy fatty acids to omega-aldehydes by a fatty alcohol oxidase (FAO). Three FAO genes (one gene designated FAO1 and two putative allelic genes designated FAO2a and FAO2b), have been cloned and sequenced from this strain. A comparison of the DNA sequence homology and derived amino acid sequence homology between these three genes and previously published Candida FAO genes indicates that FAO1 and FAO2 are distinct genes. Both genes were individually cloned and expressed in Escherichia coli. The substrate specificity and K(m) values for the recombinant FAO1 and FAO2 were significantly different. Particularly striking is the fact that FAO1 oxidizes omega-hydroxy fatty acids but not 2-alkanols, whereas FAO2 oxidizes 2-alkanols but not omega-hydroxy fatty acids. Analysis of extracts of strain H5343 during growth on fatty acids indicated that only FAO1 was highly induced under these conditions. FAO2 contains one CTG codon, which codes for serine (amino acid 177) in C. tropicalis but codes for leucine in E. coli. An FAO2a construct, with a TCG codon (codes for serine in E. coli) substituted for the CTG codon, was prepared and expressed in E. coli. Neither the substrate specificity nor the K(m) values for the FAO2a variant with a serine at position 177 were radically different from those of the variant with a leucine at that position. PMID- 15294827 TI - Molecular identification of the catabolic vinyl chloride reductase from Dehalococcoides sp. strain VS and its environmental distribution. AB - Reductive dehalogenation of vinyl chloride (VC) to ethene is the key step in complete anaerobic degradation of chlorinated ethenes. VC-reductive dehalogenase was partially purified from a highly enriched culture of the VC-respiring Dehalococcoides sp. strain VS. The enzyme reduced VC and all dichloroethene (DCE) isomers, but not tetrachloroethene (PCE) or trichloroethene (TCE), at high rates. By using reversed genetics, the corresponding gene (vcrA) was isolated and characterized. Based on the predicted amino acid sequence, VC reductase is a novel member of the family of corrinoid/iron-sulfur cluster containing reductive dehalogenases. The vcrA gene was found to be cotranscribed with vcrB, encoding a small hydrophobic protein presumably acting as membrane anchor for VC reductase, and vcrC, encoding a protein with similarity to transcriptional regulators of the NosR/NirI family. The vcrAB genes were subsequently found to be present and expressed in other cultures containing VC-respiring Dehalococcoides organisms and could be detected in water samples from a field site contaminated with chlorinated ethenes. Therefore, the vcrA gene identified here may be a useful molecular target for evaluating, predicting, and monitoring in situ reductive VC dehalogenation. PMID- 15294828 TI - Binary toxins from Bacillus thuringiensis active against the western corn rootworm, Diabrotica virgifera virgifera LeConte. AB - The western corn rootworm, Diabrotica virgifera virgifera LeConte, is a significant pest of corn in the United States. The development of transgenic corn hybrids resistant to rootworm feeding damage depends on the identification of genes encoding insecticidal proteins toxic to rootworm larvae. In this study, a bioassay screen was used to identify several isolates of the bacterium Bacillus thuringiensis active against rootworm. These bacterial isolates each produce distinct crystal proteins with approximate molecular masses of 13 to 15 kDa and 44 kDa. Insect bioassays demonstrated that both protein classes are required for insecticidal activity against this rootworm species. The genes encoding these proteins are organized in apparent operons and are associated with other genes encoding crystal proteins of unknown function. The antirootworm proteins produced by B. thuringiensis strains EG5899 and EG9444 closely resemble previously described crystal proteins of the Cry34A and Cry35A classes. The antirootworm proteins produced by strain EG4851, designated Cry34Ba1 and Cry35Ba1, represent a new binary toxin. Genes encoding these proteins could become an important component of a sustainable resistance management strategy against this insect pest. PMID- 15294829 TI - Persistence of Mycobacterium paratuberculosis during manufacture and ripening of cheddar cheese. AB - Model Cheddar cheeses were prepared from pasteurized milk artificially contaminated with high 10(4) to 10(5) CFU/ml) and low (10(1) to 10(2) CFU/ml) inocula of three different Mycobacterium paratuberculosis strains. A reference strain, NCTC 8578, and two strains (806PSS and 796PSS) previously isolated from pasteurized milk for retail sale were investigated in this study. The manufactured Cheddar cheeses were similar in pH, salt, moisture, and fat composition to commercial Cheddar. The survival of M. paratuberculosis cells was monitored over a 27-week ripening period by plating homogenized cheese samples onto HEYM agar medium supplemented with the antibiotics vancomycin, amphotericin B, and nalidixic acid without a decontamination step. A concentration effect was observed in M. paratuberculosis numbers between the inoculated milk and the 1-day old cheeses for each strain. For all manufactured cheeses, a slow gradual decrease in M. paratuberculosis CFU in cheese was observed over the ripening period. In all cases where high levels (>3.6 log(10)) of M. paratuberculosis were present in 1-day cheeses, the organism was culturable after the 27-week ripening period. The D values calculated for strains 806PSS, 796PSS, and NCTC 8578 were 107, 96, and 90 days, respectively. At low levels of contamination, M. paratuberculosis was only culturable from 27-week-old cheese spiked with strain 806PSS. M. paratuberculosis was recovered from the whey fraction in 10 of the 12 manufactured cheeses. Up to 4% of the initial M. paratuberculosis load was recovered in the culture-positive whey fractions at either the high or low initial inoculum. PMID- 15294830 TI - Investigation of spa pools associated with lung disorders caused by Mycobacterium avium complex in immunocompetent adults. AB - Three cases of Mycobacterium avium complex-related lung disorders were associated with two poorly maintained spa pools by genotypic investigations. Inadequate disinfection of the two spas had reduced the load of environmental bacteria to less than 1 CFU/ml but allowed levels of M. avium complex of 4.3 x 10(4) and 4.5 x 10(3) CFU/ml. Persistence of the disease-associated genotype was demonstrated in one spa pool for over 5 months until repeated treatments with greater than 10 mg of chlorine per liter for 1-h intervals eliminated M. avium complex from the spa pool. A fourth case of Mycobacterium avium complex-related lung disease was associated epidemiologically but not genotypically with another spa pool that had had no maintenance undertaken. This spa pool contained low numbers of mycobacteria by smear and was culture positive for M. avium complex, and the nonmycobacterial organism count was 5.2 x 10(6) CFU/ml. Public awareness about the proper maintenance of private (residential) spa pools must be promoted by health departments in partnership with spa pool retailers. PMID- 15294831 TI - Change in bacterial community structure during in situ biostimulation of subsurface sediment cocontaminated with uranium and nitrate. AB - Previous studies have demonstrated that metal-reducing microorganisms can effectively promote the precipitation and removal of uranium from contaminated groundwater. Microbial communities were stimulated in the acidic subsurface by pH neutralization and addition of an electron donor to wells. In single-well push pull tests at a number of treated sites, nitrate, Fe(III), and uranium were extensively reduced and electron donors (glucose, ethanol) were consumed. Examination of sediment chemistry in cores sampled immediately adjacent to treated wells 3.5 months after treatment revealed that sediment pH increased substantially (by 1 to 2 pH units) while nitrate was largely depleted. A large diversity of 16S rRNA gene sequences were retrieved from subsurface sediments, including species from the alpha, beta, delta, and gamma subdivisions of the class Proteobacteria, as well as low- and high-G+C gram-positive species. Following in situ biostimulation of microbial communities within contaminated sediments, sequences related to previously cultured metal-reducing delta Proteobacteria increased from 5% to nearly 40% of the clone libraries. Quantitative PCR revealed that Geobacter-type 16S rRNA gene sequences increased in biostimulated sediments by 1 to 2 orders of magnitude at two of the four sites tested. Evidence from the quantitative PCR analysis corroborated information obtained from 16S rRNA gene clone libraries, indicating that members of the delta Proteobacteria subdivision, including Anaeromyxobacter dehalogenans-related and Geobacter-related sequences, are important metal-reducing organisms in acidic subsurface sediments. This study provides the first cultivation-independent analysis of the change in metal-reducing microbial communities in subsurface sediments during an in situ bioremediation experiment. PMID- 15294832 TI - "Candidatus Endobugula glebosa," a specific bacterial symbiont of the marine bryozoan Bugula simplex. AB - The bryozoans Bugula neritina and Bugula simplex harbor bacteria in the pallial sinuses of their larvae as seen by electron microscopy. In B. neritina, the bacterial symbiont has been characterized as a gamma-proteobacterium, "Candidatus Endobugula sertula." "Candidatus E. sertula" has been implicated as the source of the bryostatins, polyketides that provide chemical defense to the host and are also being tested for use in human cancer treatments. In this study, the bacterial symbiont in B. simplex larvae was identified by 16S rRNA-targeted PCR and sequencing as a gamma-proteobacterium closely related to and forming a monophyletic group with "Candidatus E. sertula." In a fluorescence in situ hybridization, a 16S ribosomal DNA probe specific to the B. simplex symbiont hybridized to long rod-shaped bacteria in the pallial sinus of a B. simplex larva. The taxonomic status "Candidatus Endobugula glebosa" is proposed for the B. simplex larval symbiont. Degenerate polyketide synthase (PKS) primers amplified a gene fragment from B. simplex that closely matched a PKS gene fragment from the bryostatin PKS cluster. PCR surveys show that the symbiont and this PKS gene fragment are consistently and uniquely associated with B. simplex. Bryostatin activity assays and chemical analyses of B. simplex extracts reveal the presence of compounds similar to bryostatins. Taken together, these findings demonstrate a symbiosis in B. simplex that is similar and evolutionarily related to that in B. neritina. PMID- 15294833 TI - Dominant microbial composition and its vertical distribution in saline meromictic Lake Kaiike (Japan) as revealed by quantitative oligonucleotide probe membrane hybridization. AB - Vertical distributions of dominant bacterial populations in saline meromictic Lake Kaiike were investigated throughout the water column and sediment by quantitative oligonucleotide probe membrane hybridization. Three oligonucleotide probes specific for the small-subunit (SSU) rRNA of three groups of Chlorobiaceae were newly designed. In addition, three general domain (Bacteria, Archaea, and Eukarya)-specific probes, two delta-Proteobacteria-specific probes, a Chlorobiaceae-specific probe, and a Chloroflexi-specific probe were used after optimization of their washing conditions. The abundance of the sum of SSU rRNAs hybridizing with probes specific for three groups of Chlorobiaceae relative to total SSU rRNA peaked in the chemocline, accounting for up to 68%. The abundance of the delta-proteobacterial SSU rRNA relative to total SSU rRNA rapidly increased just below the chemocline up to 29% in anoxic water and peaked at the 2 to 3-cm sediment depth at ca. 34%. The abundance of SSU rRNAs hybridizing with the probe specific for the phylum Chloroflexi relative to total SSU rRNA was highest (31 to 54%) in the top of the sediment but then steeply declined with depth and became stable at 11 to 19%, indicating the robust coexistence of sulfate-reducing bacteria and Chloroflexi in the top of the sediment. Any SSU rRNA of Chloroflexi in the water column was under the detection limit. The summation of the signals of group-specific probes used in this study accounted for up to 89% of total SSU rRNA, suggesting that the DGGE-oligonucleotide probe hybridization approach, in contrast to conventional culture-dependent approaches, was very effective in covering dominant populations. PMID- 15294834 TI - Differential gene expression to investigate the effect of (5Z)-4-bromo- 5 (bromomethylene)-3-butyl-2(5H)-furanone on Bacillus subtilis. AB - (5Z)-4-Bromo-5-(bromomethylene)-3-butyl-2(5H)-furanone (furanone) from the red marine alga Delisea pulchra was found previously to inhibit the growth, swarming, and biofilm formation of gram-positive bacteria. Using the gram-positive bacterium Bacillus subtilis as a test organism, we observed cell killing by 20 microg of furanone per ml, while 5 microg of furanone per ml inhibited growth approximately twofold without killing the cells. To discover the mechanism of this inhibition on a genetic level and to investigate furanone as a novel antibiotic, full-genome DNA microarrays were used to analyze the gene expression profiles of B. subtilis grown with and without 5 microg of furanone per ml. This agent induced 92 genes more than fivefold (P < 0.05) and repressed 15 genes more than fivefold (P < 0.05). The induced genes include genes involved in stress responses (such as the class III heat shock genes clpC, clpE, and ctsR and the class I heat shock genes groES, but no class II or IV heat shock genes), fatty acid biosynthesis, lichenan degradation, transport, and metabolism, as well as 59 genes with unknown functions. The microarray results for four genes were confirmed by RNA dot blotting. Mutation of a stress response gene, clpC, caused B. subtilis to be much more sensitive to 5 microg of furanone per ml (there was no growth in 8 h, while the wild-type strain grew to the stationary phase in 8 h) and confirmed the importance of the induction of this gene as identified by the microarray analysis. PMID- 15294835 TI - Correspondence between community structure and function during succession in phenol- and phenol-plus-trichloroethene-fed sequencing batch reactors. AB - The effects of more than 2 years of trichloroethene (TCE) application on community succession and function were studied in two aerobic sequencing batch reactors. One reactor was fed phenol, and the second reactor was fed both phenol and TCE in sequence twice per day. After initiation of TCE loading in the second reactor, the TCE transformation rates initially decreased, but they stabilized with an average second-order rate coefficient of 0.044 liter mg(-1) day(-1) for 2 years. In contrast, the phenol-fed reactor showed higher and unstable TCE transformation rates, with an average rate coefficient of 0.093 liter mg(-1) day( 1). Community analysis by terminal restriction fragment length polymorphism (T RFLP) analysis of the 16S rRNA genes showed that the phenol-plus-TCE-fed reactor had marked changes in community structure during the first 100 days and remained relatively stable afterwards, corresponding to the period of stable function. In contrast, the community structure of the phenol-fed reactor changed periodically, and the changes coincided with the periodicity observed in the TCE transformation rates. Correspondence analysis of each reactor community showed that different community structures corresponded with function (TCE degradation rate). Furthermore, the phenol hydroxylase genotypes, as determined by restriction fragment length polymorphism analysis, corresponded to community structure patterns identified by T-RFLP analysis and to periods when the TCE transformation rates were high. Long-term TCE stress appeared to select for a different and stable community structure, with lower but stable TCE degradation rates. In contrast, the community under no stress exhibited a dynamic structure and dynamic function. PMID- 15294836 TI - Biphenyl and benzoate metabolism in a genomic context: outlining genome-wide metabolic networks in Burkholderia xenovorans LB400. AB - We designed and successfully implemented the use of in situ-synthesized 45-mer oligonucleotide DNA microarrays (XeoChips) for genome-wide expression profiling of Burkholderia xenovorans LB400, which is among the best aerobic polychlorinated biphenyl degraders known so far. We conducted differential gene expression profiling during exponential growth on succinate, benzoate, and biphenyl as sole carbon sources and investigated the transcriptome of early-stationary-phase cells grown on biphenyl. Based on these experiments, we outlined metabolic pathways and summarized other cellular functions in the organism relevant for biphenyl and benzoate degradation. All genes previously identified as being directly involved in biphenyl degradation were up-regulated when cells were grown on biphenyl compared to expression in succinate-grown cells. For benzoate degradation, however, genes for an aerobic coenzyme A activation pathway were up-regulated in biphenyl-grown cells, while the pathway for benzoate degradation via hydroxylation was up-regulated in benzoate-grown cells. The early-stationary phase biphenyl-grown cells showed similar expression of biphenyl pathway genes, but a surprising up-regulation of C(1) metabolic pathway genes was observed. The microarray results were validated by quantitative reverse transcription PCR with a subset of genes of interest. The XeoChips showed a chip-to-chip variation of 13.9%, compared to the 21.6% variation for spotted oligonucleotide microarrays, which is less variation than that typically reported for PCR product microarrays. PMID- 15294837 TI - Development and application of a real-time PCR approach for quantification of uncultured bacteria in the central Baltic Sea. AB - We have developed a highly sensitive approach to assess the abundance of uncultured bacteria in water samples from the central Baltic Sea by using a noncultured member of the "Epsilonproteobacteria" related to Thiomicrospira denitrificans as an example. Environmental seawater samples and samples enriched for the target taxon provided a unique opportunity to test the approach over a broad range of abundances. The approach is based on a combination of taxon- and domain-specific real-time PCR measurements determining the relative T. denitrificans-like 16S rRNA gene and 16S rRNA abundances, as well as the determination of total cell counts and environmental RNA content. It allowed quantification of T. denitrificans-like 16S rRNA molecules or 16S rRNA genes as well as calculation of the number of ribosomes per T. denitrificans-like cell. Every real-time measurement and its specific primer system were calibrated using environmental nucleic acids obtained from the original habitat for external standardization. These standards, as well as the respective samples to be measured, were prepared from the same DNA or RNA extract. Enrichment samples could be analyzed directly, whereas environmental templates had to be preamplified with general bacterial primers before quantification. Preamplification increased the sensitivity of the assay by more than 4 orders of magnitude. Quantification of enrichments with or without a preamplification step yielded comparable results. T. denitrificans-like 16S rRNA molecules ranged from 7.1 x 10(3) to 4.4 x 10(9) copies ml(-1) or 0.002 to 49.7% relative abundance. T. denitrificans-like 16S rRNA genes ranged from 9.0 x 10(1) to 2.2 x10(6) copies ml(-1) or 0.01 to 49.7% relative abundance. Detection limits of this real-time PCR approach were 20 16S rRNA molecules or 0.2 16S rRNA gene ml(-1). The number of ribosomes per T. denitrificans-like cell was estimated to range from 20 to 200 in seawater and reached up to 2,000 in the enrichments. The results indicate that our real-time PCR approach can be used to determine cellular and relative abundances of uncultured marine bacterial taxa and to provide information about their levels of activity in their natural environment. PMID- 15294838 TI - Model system for growing and quantifying Streptococcus pneumoniae biofilms in situ and in real time. AB - Streptococcus pneumoniae forms biofilms, but little is known about its extracellular polymeric substances (EPS) or the kinetics of biofilm formation. A system was developed to enable the simultaneous measurement of cells and the EPS of biofilm-associated S. pneumoniae in situ over time. A biofilm reactor containing germanium coupons was interfaced to an attenuated total reflectance (ATR) germanium cell of a Fourier transform infrared (FTIR) laser spectrometer. Biofilm-associated cells were recovered from the coupons and quantified by total and viable cell count methods. ATR-FTIR spectroscopy of biofilms formed on the germanium internal reflection element (IRE) of the ATR cell provided a continuous spectrum of biofilm protein and polysaccharide (a measure of the EPS). Staining of the biofilms on the IRE surface with specific fluorescent probes provided confirmatory evidence for the biofilm structure and the presence of biofilm polysaccharides. Biofilm protein and polysaccharides were detected within hours after inoculation and continued to increase for the next 141 h. The polysaccharide band increased at a substantially higher rate than did the protein band, demonstrating increasing coverage of the IRE surface with biofilm polysaccharides. The biofilm total cell counts on germanium coupons stabilized after 21 h, at approximately 10(5) cells per cm(2), while viable counts decreased as the biofilm aged. This system is unique in its ability to detect and quantify biofilm-associated cells and EPS of S. pneumoniae over time by using multiple, corroborative techniques. This approach could prove useful for the study of biofilm processes of this or other microorganisms of clinical or industrial relevance. PMID- 15294839 TI - VdNEP, an elicitor from Verticillium dahliae, induces cotton plant wilting. AB - Verticillium wilt is a vascular disease of cotton. The causal fungus, Verticillium dahliae, secretes elicitors in culture. We have generated approximately 1,000 5'-terminal expressed sequence tags (ESTs) from a cultured mycelium of V. dahliae. A number of ESTs were found to encode proteins harboring putative signal peptides for secretion, and their cDNAs were isolated. Heterologous expression led to the identification of a protein with elicitor activities. This protein, named V. dahliae necrosis- and ethylene-inducing protein (VdNEP), is composed of 233 amino acids and has high sequence identities with fungal necrosis- and ethylene-inducing proteins. Infiltration of the bacterially expressed His-VdNEP into Nicotiana benthamiana leaves resulted in necrotic lesion formation. In Arabidopsis thaliana, the fusion protein also triggered production of reactive oxygen species and induced the expression of PR genes. When added into suspension cultured cells of cotton (Gossypium arboreum), the fusion protein elicited the biosynthesis of gossypol and related sesquiterpene phytoalexins at low concentrations, and it induced cell death at higher concentrations. On cotton cotyledons and leaves, His-VdNEP induced dehydration and wilting, similar to symptoms caused by a crude preparation of V. dahliae elicitors. Northern blotting showed a low level of VdNEP expression in the mycelium during culture. These data suggest that VdNEP is a wilt-inducing factor and that it participates in cotton-V. dahliae interactions. PMID- 15294840 TI - Biological effects of Trichoderma harzianum peptaibols on mammalian cells. AB - Trichoderma species isolated from water-damaged buildings were screened for toxicity by using boar sperm cells as indicator cells. The crude methanolic cell extract from Trichoderma harzianum strain ES39 inhibited the boar sperm cell motility at a low exposure concentration (50% effective concentration, 1 to 5 microg [dry weight] ml of extended boar semen(-1)). The same exposure concentration depleted the boar sperm cells of NADH(2). Inspection of the exposed boar sperm cells by transmission electron microscopy revealed damage to the plasma membrane. By using the black lipid membrane technique, it was shown that the semipurified metabolites (eluted from a SepPak C(18) cartridge) of T. harzianum strain ES39 induced voltage-dependent conductivity. The high performance liquid chromatography-purified metabolites of T. harzianum strain ES39 dissipated the mitochondrial membrane potential (Deltapsi(m)) of human lung epithelial carcinoma cells (cell line A549). The semipurified metabolites (eluted from a SepPak C(18) cartridge) of T. harzianum strain ES39 were analyzed by mass spectrometry (MS). Matrix-assisted laser desorption ionization and nanoflow electrospray ionization MS revealed five major peptaibols, each of which contained 18 residues and had a mass ranging from 1,719 to 1,775 Da. Their partial amino acid sequences were determined by collision-induced dissociation tandem MS. PMID- 15294841 TI - Host physiology and pathogenic variation of Cochliobolus heterostrophus strains with mutations in the G protein alpha subunit, CGA1. AB - Conserved eukaryotic signaling proteins participate in development and disease in plant-pathogenic fungi. Strains with mutations in CGA1, a heterotrimeric G protein G alpha subunit gene of the maize pathogen Cochliobolus heterostrophus, are defective in several developmental pathways. Conidia from CGA1 mutants germinate as abnormal, straight-growing germ tubes that form few appressoria, and the mutants are female sterile. Nevertheless, these mutants can cause normal lesions on plants, unlike other filamentous fungal plant pathogens in which functional homologues of CGA1 are required for full virulence. Deltacga1 mutants of C. heterostrophus were less infective of several maize varieties under most conditions, but not all, as virulence was nearly normal on detached leaves. This difference could be related to the rapid senescence of detached leaves, since delaying senescence with cytokinin also had differential effects on the virulence of the wild type and the Deltacga1 mutant. In particular, detached leaves may provide a more readily available nutrient source than attached leaves. Decreased fitness of Deltacga1 as a pathogen may reflect conditions under which full virulence requires signal transduction through CGA1-mediated pathways. The virulence of these signal transduction mutants is thus affected differentially by the physiological state of the host. PMID- 15294842 TI - Explorative multivariate analyses of 16S rRNA gene data from microbial communities in modified-atmosphere-packed salmon and coalfish. AB - Modified-atmosphere packaging (MAP) of foods in combination with low-temperature storage extends product shelf life by limiting microbial growth. We investigated the microbial biodiversity of MAP salmon and coalfish by using an explorative approach and analyzing both the total amounts of bacteria and the microbial group composition (both aerobic and anaerobic bacteria). Real-time PCR analyses revealed a surprisingly large difference in the microbial loads for the different fish samples. The microbial composition was determined by examining partial 16S rRNA gene sequences from 180 bacterial isolates, as well as by performing terminal restriction fragment length polymorphism analysis and cloning 92 sequences from PCR products of DNA directly retrieved from the fish matrix. Twenty different bacterial groups were identified. Partial least-squares (PLS) regression was used to relate the major groups of bacteria identified to the fish matrix and storage time. A strong association of coalfish with Photobacterium phosphoreum was observed. Brochothrix spp. and Carnobacterium spp., on the other hand, were associated with salmon. These bacteria dominated the fish matrixes after a storage period. Twelve Carnobacterium isolates were identified as either Carnobacterium piscicola (five isolates) or Carnobacterium divergens (seven isolates), while the eight Brochothrix isolates were identified as Brochothrix thermosphacta by full-length 16S rRNA gene sequencing. Principal-component analyses and PLS analysis of the growth characteristics (with 49 different substrates) showed that C. piscicola had distinct substrate requirements, while the requirements of B. thermosphacta and C. piscicola were quite divergent. In conclusion, our explorative multivariate approach gave a picture of the total microbial biodiversity in MAP fish that was more comprehensive than the picture that could be obtained previously. Such information is crucial in controlled food production when, for example, the hazard analysis of critical control points principle is used. PMID- 15294843 TI - Strategy for cloning large gene assemblages as illustrated using the phenylacetate and polyhydroxyalkanoate gene clusters. AB - We report an easy procedure for isolating chromosome-clustered genes. By following this methodology, the entire set of genes belonging to the phenylacetic acid (PhAc; 18-kb) pathway as well as those required for the synthesis and mobilization of different polyhydroxyalkanoates (PHAs; 6.4 kb) in Pseudomonas putida U were recovered directly from the bacterial chromosome and cloned into a plasmid for the first time. The transformation of different bacteria with these genetic constructions conferred on them the ability to either degrade PhAc or synthesize bioplastics (PHAs). PMID- 15294844 TI - Two morphological types of cell appendages on a strongly adhesive bacterium, Acinetobacter sp. strain Tol 5. AB - Two morphological types of appendages, an anchor-like appendage and a peritrichate fibril-type appendage, have been observed on cells of an adhesive bacterium, Acinetobacter sp. strain Tol 5, by use of recently developed electron microscopic techniques. The anchor extends straight to the substratum without branching and tethers the cell body at its end at distances of several hundred nanometers, whereas the peritrichate fibril attaches to the substratum in multiple places, fixing the cell at much shorter distances. PMID- 15294845 TI - Nisin-controlled production of pediocin PA-1 and colicin V in nisin- and non nisin-producing Lactococcus lactis strains. AB - The introduction of chimeric genes encoding the fusion leader of lactococcin A propediocin PA-1 or procolicin V under the control of the inducible nisA promoter and the lactococcin A-dedicated secretion genes (lcnCD) into Lactococcus lactis strains, including a nisin producer, expressing the two component regulator NisRK led to the production or pediocin PA-1 or colicin V, respectively. PMID- 15294846 TI - Translational features of human alpha 2b interferon production in Escherichia coli. AB - The yield of human alpha 2b interferon in Escherichia coli was optimized by replacement of low-usage arginine codons located in the mRNA 5' end. The differences observed among the various gene variants suggest that codon usage, Shine-Dalgarno-like sequences, and mRNA secondary structure contribute to the performance of E. coli translation machinery. PMID- 15294847 TI - Direct production of ethanol from raw corn starch via fermentation by use of a novel surface-engineered yeast strain codisplaying glucoamylase and alpha amylase. AB - Direct and efficient production of ethanol by fermentation from raw corn starch was achieved by using the yeast Saccharomyces cerevisiae codisplaying Rhizopus oryzae glucoamylase and Streptococcus bovis alpha-amylase by using the C-terminal half region of alpha-agglutinin and the flocculation functional domain of Flo1p as the respective anchor proteins. In 72-h fermentation, this strain produced 61.8 g of ethanol/liter, with 86.5% of theoretical yield from raw corn starch. PMID- 15294848 TI - Temporal transcription map of the virulent Streptococcus thermophilus bacteriophage Sfi19. AB - A transcription map was developed for the virulent Streptococcus thermophilus phage Sfi19 on the basis of systematic Northern blot hybridizations. All deduced 5' ends were confirmed by primer extension experiments. Three classes of transcripts were detected based on the different times of appearance. Early transcripts were identified in three genome regions; middle transcripts covered cro-like, DNA replication, and transcriptional regulation genes; and late genes consisted of structural and lysis genes. Chloramphenicol treatment suppressed the translation of a putative transcriptional factor necessary for the production of late transcripts and shifted middle transcripts to early transcription times. PMID- 15294849 TI - Cyanobacterial protease inhibitor microviridin J causes a lethal molting disruption in Daphnia pulicaria. AB - Laboratory experiments identified microviridin J as the source of a fatal molting disruption in Daphnia species organisms feeding on Microcystis cells. The molting disruption was presumably linked to the inhibitory effect of microviridin J on daphnid proteases, suggesting that hundreds of further cyanobacterial protease inhibitors must be considered potentially toxic to zooplankton. PMID- 15294850 TI - Nisin-producing Lactococcus lactis strains isolated from human milk. AB - Characterization by partial 16S rRNA gene sequencing, ribotyping, and green fluorescent protein-based nisin bioassay revealed that 6 of 20 human milk samples contained nisin-producing Lactococcus lactis bacteria. This suggests that the history of humans consuming nisin is older than the tradition of consuming fermented milk products. PMID- 15294851 TI - Modelling ontogenetic changes of nitrogen and water content in lettuce. AB - BACKGROUND AND AIMS: It is well established that the nitrogen content of plants, including lettuce, decreases with time. It has also been observed that water content of lettuce increases between planting and harvest. This paper is an attempt at modelling these observations. METHODS: An existing dynamic model (Nicolet), designed to predict growth and nitrate content of glasshouse lettuce, is modified to accommodate the ontogenetic changes of reduced-nitrogen and water contents (on a dry matter basis). The decreasing reduced-N content and the increasing water content are mimicked by dividing the originally uniform plant into 'metabolically active' tissue and 'support' tissue. The 'metabolic' tissue is assumed to contain a higher nitrogen content and a lower water content than the 'support' tissue. As the plants grow, the ratio of 'support' to 'metabolic' tissue increases, resulting in an increased mean water content and a decreased reduced-N content. Simulations with the new model are compared with experimental glasshouse data over four seasons. KEY RESULTS: The empirical linear relationship between water and reduced-N contents, matches, to a good approximation, the corresponding relationship based on the model. The agreement between the two makes it possible to effectively uncouple the estimation of the 'ontogenetic' parameters from the estimation of the other parameters. The growth and nitrate simulation results match the data rather well and are hardly affected by the new refinement. The reduced-N and water contents are predicted much better with the new model. CONCLUSION: Prediction of nitrogen uptake for the substantial nitrate pool of lettuce depends on the water content. Hence, the modified model may assist in making better fertilization decisions and better estimates of nitrogen leaching. PMID- 15294852 TI - PKC-epsilon is upstream and PKC-alpha is downstream of mitoKATP channels in the signal transduction pathway of ischemic preconditioning of human myocardium. AB - Protein kinase C (PKC) is involved in the process of ischemic preconditioning (IPC), although the precise mechanism is still a subject of debate. Using specific PKC inhibitors, we investigated which PKC isoforms were involved in IPC of the human atrial myocardium sections and to determine their temporal relationship to the opening of mitochondrial potassium-sensitive ATP (mitoKATP) channels. Right atrial muscles obtained from patients undergoing elective cardiac surgery were equilibrated and then randomized to receive any of the following protocols: aerobic control, 90-min simulated ischemia/120-min reoxygenation, IPC using 5-min simulated ischemia/5-min reoxygenation followed by 90-min simulated ischemia/120-min reoxygenation and finally, PKC inhibitors were added 10 min before and 10 min during IPC followed by 90-min simulated ischemia/120-min reoxygenation. The PKC isoforms inhibitors investigated were V1-2 peptide, GO 6976, rottlerin, and LY-333531 for PKC-epsilon, -alpha, -delta and -beta, respectively. To investigate the relation of PKC isoforms to mitoKATP channels, PKC inhibitors found to be involved in IPC were added 10 min before and 10 min during preconditioning by diazoxide followed by 90-min simulated ischemia/120-min reoxygenation in a second experiment. Creatine kinase leakage and methylthiazoletetrazolium cell viability were measured. Phosphorylation of PKC isoforms after activation of the sample by either diazoxide or IPC was detected by using Western blot analysis and then analyzed by using Scion image software. PKC-alpha and -epsilon inhibitors blocked IPC, whereas PKC-delta and -beta inhibitors did not. The protection elicited by diazoxide, believed to be via mitoKATP channels opening, was blocked by the inhibition of PKC-alpha but not epsilon isoforms. In addition, diazoxide caused increased phosphorylation of PKC alpha to the same extent as IPC but did not affect the phosphorylation of PKC epsilon, a process believed to be critical in PKC activation. The results demonstrate that PKC-alpha and -epsilon are involved in IPC of the human myocardium with PKC-epsilon being upstream and PKC-alpha being downstream of mitoKATP channels. PMID- 15294853 TI - Lysophospholipids increase ICAM-1 expression in HUVEC through a Gi- and NF-kappaB dependent mechanism. AB - Lysophosphatidic acid (LPA) and sphingosine 1-phosphate (S-1-P) are both low molecular weight lysophospholipid (LPL) ligands that are recognized by the Edg family of G protein-coupled receptors. In endothelial cells, these two ligands activate Edg receptors, resulting in cell proliferation and cell migration. The intercellular adhesion molecule-1 (ICAM-1, CD54) is one of many cell adhesion molecules belonging to the immunoglobulin superfamily. This study showed that LPA and S-1-P enhance ICAM-1 expression at both the mRNA and protein levels in human umbilical cord vein endothelial cells (HUVECs). This enhanced ICAM-1 expression in HUVECs was first observed at 2 h postligand treatment. Maximal expression appeared at 8 h postligand treatment, as detected by flow cytometry and Western blotting. Furthermore, the effects of S-1-P on ICAM-1 expression were shown to be concentration dependent. Prior treatment of HUVECs with pertussis toxin, a specific inhibitor of G(i), ammonium pyrrolidinedithiocarbamate and BAY 11-7082, inhibitors of the nuclear factor (NF)-kappaB pathway, or Clostridium difficile toxin B, an inhibitor of Rac, prevented the enhanced effect of LPL-induced ICAM-1 expression. However, pretreatment of HUVECs with exoC3, an inhibitor of Rho, had no effect on S-1-P-enhanced ICAM-1 expression. In a static cell-cell adhesion assay system, pretreatment of LPL enhanced the adhesion between HUVECs and U-937 cells, a human mononucleated cell line. The enhanced adhesion effect could be prevented by preincubation with a functional blocking antibody against human ICAM 1. These results suggest that LPLs released by activated platelets might enhance interactions of leukocytes with the endothelium through a G(i)-, NF-kappaB-, and possibly Rac-dependent mechanism, thus facilitating wound healing and inflammation processes. PMID- 15294854 TI - Trafficking of cholera toxin-ganglioside GM1 complex into Golgi and induction of toxicity depend on actin cytoskeleton. AB - Intestinal epithelial lipid rafts contain ganglioside GM1 that is the receptor for cholera toxin (CT). The ganglioside binds CT at the plasma membrane (PM) and carries the toxin through the trans-Golgi network (TGN) to the endoplasmic reticulum (ER). In the ER, a portion of the toxin unfolds and translocates to the cytosol to activate adenylyl cyclase. Activation of the cyclase leads to an increase in intracellular cAMP, which results in apical chloride secretion. Here, we find that an intact actin cytoskeleton is necessary for the efficient transport of CT to the Golgi and for subsequent activation of adenylyl cyclase. CT bound to GM1 on the cell membrane fractionates with a heterogeneous population of lipid rafts, a portion of which is enriched in actin and other cytoskeletal proteins. In this actin-rich fraction of lipid rafts, CT and actin colocalize on the same membrane microdomains, suggesting a possible functional association. Depolymerization or stabilization of actin filaments interferes with transport of CT from the PM to the Golgi and reduces the levels of cAMP generated in the cytosol. Depletion of membrane cholesterol, which also inhibits CT trafficking to the TGN, causes displacement of actin from the lipid rafts while CT remains stably raft associated. On the basis of these observations, we propose that the CT-GM1 complex is associated with the actin cytoskeleton via the lipid rafts and that the actin cytoskeleton plays a role in trafficking of CT from the PM to the Golgi/ER and the subsequent activation of adenylyl cyclase. PMID- 15294855 TI - Regulation of dihydropyridine receptor gene expression in mouse skeletal muscles by stretch and disuse. AB - This study examined dihydropyridine receptor (DHPR) gene expression in mouse skeletal muscles during physiological adaptations to disuse. Disuse was produced by three in vivo models-denervation, tenotomy, and immobilization-and DHPR alpha1s mRNA was measured by quantitative Northern blot. After 14-day simultaneous denervation of the soleus (Sol), tibialis anterior (TA), extensor digitorum longus (EDL), and gastrocnemius (Gastr) muscles by sciatic nerve section, DHPR mRNA increased preferentially in the Sol and TA (+1.6-fold), whereas it increased in the EDL (+1.6-fold) and TA (+1.8-fold) after selective denervation of these muscles by peroneal nerve section. It declined in all muscles (-1.3- to -2.6-fold) after 14-day tenotomy, which preserves nerve input but removes mechanical tension. Atrophy was comparable in denervated and tenotomized muscles. These results suggest that factor(s) in addition to inactivity per se, muscle phenotype, or associated atrophy can regulate DHPR gene expression. To test the contribution of passive tension to this regulation, we subjected the same muscles to disuse by limb immobilization in a maximally dorsiflexed position. DHPR alpha1s mRNA increased in the stretched muscles (Sol, +2.3-fold; Gastr, +1.5-fold) and decreased in the shortened muscles (TA, -1.4 fold; EDL, -1.3-fold). The effect of stretch was confirmed in vitro. DHPR protein did not change significantly after 4-day immobilization, suggesting that additional levels of regulation may exist. These results demonstrate that DHPR alpha1s gene expression is regulated as an integral part of the adaptive response of skeletal muscles to disuse in both slow- and fast-twitch muscles and identify passive tension as an important signal for its regulation in vivo. PMID- 15294856 TI - Prospective randomised controlled trial of an infection screening programme to reduce the rate of preterm delivery. AB - OBJECTIVE: To evaluate whether a screening strategy in pregnancy lowers the rate of preterm delivery in a general population of pregnant women. DESIGN: Multicentre, prospective, randomised controlled trial. SETTING: Non-hospital based antenatal clinics. PARTICIPANTS: 4429 pregnant women presenting for their routine prenatal visits early in the second trimester were screened by Gram stain for asymptomatic vaginal infection. In the intervention group, the women's obstetricians received the test results and women received standard treatment and follow up for any detected infection. In the control group, the results of the vaginal smears were not revealed to the caregivers. MAIN OUTCOME MEASURES: The primary outcome variable was preterm delivery at less than 37 weeks. Secondary outcome variables were preterm delivery at less than 37 weeks combined with different birth weight categories equal to or below 2500 g and the rate of late miscarriage. RESULTS: Outcome data were available for 2058 women in the intervention group and 2097 women in the control group. In the intervention group, the number of preterm births was significantly lower than in the control group (3.0% v 5.3%, 95% confidence interval 1.2 to 3.6; P = 0.0001). Preterm births were also significantly reduced in lower weight categories at less than 37 weeks and 6-linked ManNAc 1-phosphate that is distinct from the capsule structures of the other meningococcal disease-causing serogroups, B, C, Y, and W-135. The serogroup A capsule biosynthetic genetic cassette consists of four open reading frames, mynA-D (sacA-D), that are specific to serogroup A, but the functions of these genes have not been well characterized. mynC was found to encode an inner membrane-associated acetyltransferase that is responsible for the O-acetylation of the CPS of serogroup A. The wild-type CPS as revealed by 1H NMR had 60-70% O-acetylated ManNAc residues that contained acetyl groups at O-3, with some species acetylated at O-4 and at both O-3 and O-4. A non-polar mynC mutant generated by introducing an aphA-3 kanamycin resistance cassette produced CPS with no O-acetylation. A serogroup A capsule-specific monoclonal antibody was shown to recognize the wild type O-acetylated CPS, but not the CPS of the mynC mutant, which lacked O acetylation. MynC was C-terminally His-tagged and overexpressed in Escherichia coli to obtain the predicted approximately 26-kDa protein. The acetyltransferase activity of purified MynC was demonstrated in vitro using [14C]acetyl-CoA. MynC O acetylated the O-acetylated CPS of the mynC mutant and further acetylated the wild-type CPS of serogroup A meningococci, but not the CPS of serogroup B or C meningococci. Genetic complementation of the mynC mutant confirmed the function of MynC as the serogroup A CPS O-3 and O-4 acetyltransferase. MynC represents a new subclass of O-acetyltransferases that utilize acetyl-CoA to decorate the D mannosamine capsule of N. meningitidis serogroup A. PMID- 15294917 TI - Social control of health behaviors: a comparison of young, middle-aged, and older adults. AB - Social control can positively influence health behaviors, but changes in social networks over time may cause older adults to experience less health-related social control. The size and composition of social control networks, and receipt of health-related social control, were examined in a probability sample of 509 household residents (aged 25-80 years) in Los Angeles County who completed a telephone survey. Compared with younger and middle-aged adults, older adults identified fewer people who attempted to influence their health behaviors and fewer health behaviors that others urged them to change. Older adults also reported less frequent social control attempts aimed at modifying their health behaviors, even after health status, health habits, and social network characteristics were controlled for. Possible explanations for these age-related differences are discussed. PMID- 15294918 TI - The role of health congruence in functional status and depression. AB - Few studies have attempted to examine the meaning of health congruence, particularly in the oldest old. Participants were drawn from a longitudinal study of the oldest old (N = 151; M = 90 years). Dichotomized objective health was cross classified with dichotomized subjective health, producing four health congruence groups: good health realists, poor health realists, optimists, and pessimists. Both good health realists and pessimists had good objective health, yet pessimists had the highest depression, lowest functional status, and frequent reports of hospitalization. By contrast, the poor health realists and optimist groups had poor objective health, but the optimists had better outcomes on depression. This suggests that discrepancies between objective and subjective health may have significant implications for health outcomes. PMID- 15294919 TI - Emotional well-being in recently bereaved widows: a dynamical systems approach. AB - A dynamical systems approach was used to model the intraindividual variability in emotional well-being following conjugal loss. Well-being in a sample of 19 recently bereaved older adult widows was measured every day for 3 months. The pattern of variability of well-being was hypothesized to be an oscillating process that damps across time (i.e., large swings followed by a gradual damping). Results indicated that there was significant patterned variability in the emotional well-being adjustment that can be modeled by a linear oscillator model (R2=.77), in addition to an overall positive trend. Applying dynamical systems analyses to capture variability and subsequent well-being trajectories following spousal loss is an important step in delineating the complex adjustment to widowhood. PMID- 15294920 TI - The role of daily positive emotions during conjugal bereavement. AB - The role of daily positive emotions in the stress process was examined in a sample of 34 recently bereaved older adult widows. Humor coping and perceived stress were measured in questionnaires, and positive emotions, depression, anxiety, and stress were assessed daily for 98 days. Results highlight the critical role of daily positive emotions in the months immediately following conjugal loss. Intraindividual analyses revealed significant reductions in the magnitude of the stress-depression correlation on days in which greater positive emotions were present. Results also suggest that different vulnerability and resilience factors are implicated in the emotion differentiation process. For widows with greater humor coping skills, there was less overlap in daily ratings of positive emotions and depressive symptoms. In contrast, higher levels of chronic stress resulted in less differentiation of emotional responses. PMID- 15294921 TI - Longitudinal changes in the well-being of Japanese caregivers: variations across kin relationships. AB - This study examined how the psychological well-being of Japanese caregivers changed over time; it also examined the variation across kin relationships with care recipients. Three interviews over the course of 30 months were conducted with a representative sample of community-dwelling caregivers of frail elderly persons living in a Tokyo suburb. Latent growth modeling demonstrated that mean levels of both depression and emotional exhaustion worsened over time. Change in emotional exhaustion over time showed significant individual variability, whereas change in depression showed little individual variability. Although wife caregivers tended to experience the worst trajectory of emotional exhaustion, daughters-in-law also showed a similar negative trend. The difference in individuals' well-being trajectories by kinship may be explained partly by differences in care recipients' disabilities. PMID- 15294922 TI - Positive aspects of Alzheimer's caregiving: the role of race. AB - We examined differences in positive aspects of caregiving (PAC) among 275 African American and 343 Caucasian caregivers of individuals with Alzheimer's disease from the National Institutes of Health Resources for Enhancing Alzheimer's Care Health (REACH) study sites in Birmingham, Memphis, and Philadelphia. African Americans reported higher scores on PAC than did Caucasians. African Americans' higher religiosity partially mediated the relationship between race and PAC. Additional variables that contributed to their higher PAC scores were African Americans' lower anxiety, lower feelings of bother by the care recipient's behavior, and lower socioeconomic status. PMID- 15294923 TI - Age differences in feature selection in triple conjunction search. AB - Younger and older participants were trained in a triple conjunction visual search task to examine age differences in the development of proficient performance. For the first 8 days, participants searched for a target defined by its contrast polarity, shape, and orientation. On Days 9 through 16, the target identity was switched to one defined by opposing feature values. On Day 17, the target was returned to the original feature values. Results indicated that, after training, younger adults reduced their display size effects more than elderly adults. Disruption occurred after the first but not after the second transfer. However, each time the target was switched, there were no age differences in disruption. Eye movement data suggest that older adults use a similar feature selection strategy as younger adults but may be more susceptible to distraction. The results are discussed in terms of current models of attention and search. PMID- 15294924 TI - Managing decline in assisted living: the key to aging in place. AB - OBJECTIVE: This study investigates the process of aging in place in assisted living facilities (ALFs) and seeks to gain an in-depth understanding of the factors influencing this phenomenon in a variety of ALF contexts. METHODS: Qualitative methods were used to study five ALFs for 1 year. Data collection methods included participant observation; informal and in-depth interviewing of providers, residents, and residents' families; and review of resident and facility records. Analysis was conducted using the grounded theory approach. RESULTS: The ability of residents to remain in assisted living was principally a function of the "fit" between the capacity of both residents and facilities to manage decline. Multiple community, facility, and resident factors influenced the capacity to manage decline, and resident-facility fit was both an outcome and an influence on the decline management process. Resident and facility risk also was an intervening factor and a consequence of decline management. DISCUSSION: Findings point out the complexity of aging in place in ALFs and the need for a coordinated effort by facilities, residents, and families in the management of resident decline. Findings further highlight the necessity of residents being well informed about both their own needs and the capacity of a facility to meet them. PMID- 15294925 TI - Grandparent identity, intergenerational family identity, and well-being. AB - OBJECTIVE: A new grandparent identity measure is constructed that allows us to compare grandparent identity meanings with the meanings of other adult identities and to investigate the relationships between identities and well-being. METHODS: Data were collected in 1997 from 203 older grandmothers and grandfathers living in a metropolitan area. Grandparent and parent identity meanings are measured with an introductory identifier focusing attention on being a grandparent or a parent, followed by a set of 10 adjective pairs to capture identity meanings. Intergenerational family identity combined grandparent and parent identity meanings. Self-esteem and depressive symptoms serve as two indicators of well being. RESULTS: We find that there are no significant differences between grandparent and parent identity meanings and that men and women are more positive about their grandparent identities than they are about other adult identity meanings. Further, grandparent identity is significantly related to well-being when it is the only identity in the model but not when parent identity is included in another model. Finally, intergenerational family identity is positively related to well-being. DISCUSSION: The findings confirm the expectation that grandparent identity meanings may encourage well-being. Further, the intergenerational identity reflects the overlapping meanings and experiences of being a parent and a grandparent. PMID- 15294926 TI - Is there equity in the home health care market? Understanding racial patterns in the use of formal home health care. AB - OBJECTIVE: This article explores whether the formal home health care (HHC) market is equitable or manifests unexplained racial disparities in use. METHODS: The database is the 1994 National Long Term Care Survey. We estimate logit regression models with a race dummy variable, race interaction terms, and stratification by race. We apply the Oaxaca decomposition technique to quantify whether the observed racial gap in formal HHC use is explained by racial differences in predisposing, enabling, need, and environmental characteristics. RESULT: We find numerous unique racial patterns in HHC use. Blacks with diabetes and low income have higher probabilities of HHC use than their White counterparts. Black older persons have a 25% higher chance of using HHC than Whites. Our Oaxaca analysis indicates that racial differences in predisposing, enabling, need, and environmental characteristics account for the racial gap in use of HHC. DISCUSSION: We find that the HHC market is equitable, enhancing availability, acceptability, and accessibility of care for older Black persons. Thus, the racial differences that we find are not racial disparities. PMID- 15294927 TI - Stability and change in social negativity in later life: reducing received while maintaining initiated negativity. AB - OBJECTIVE: This article examines stability and change in social negativity and the links between social negativity and instrumental support over time among disabled older adults. The analyses focused on family relationships because social negativity tends to be more prevalent in family compared with nonkin relationships. Social negativity received and initiated are each addressed separately to determine whether or not they show similar patterns and links to instrumental support over time. METHODS: Latent growth curve methodology was used to examine change over time in social negativity and instrumental family support in relation to age, gender, and family network size at baseline. Participants, 570 older adults with chronic visual impairment, were interviewed three times over an 18-month period. RESULTS: Social negativity received showed a decrease over time, whereas levels of social negativity initiated remained more stable. Links with instrumental support were positive but stronger for received compared with initiated social negativity. DISCUSSION: The differential pattern of stability and change over time in received versus initiated social negativity and their links to instrumental support suggest different origins for the initiation versus receipt of social negativity. PMID- 15294928 TI - Trajectories of impairment, social support, and depressive symptoms in later life. AB - OBJECTIVE: Research has increasingly focused on the dynamic nature of disability and depressive symptoms in later life. Little research, however, has modeled disability and depressive symptoms as dynamic, related processes. Furthermore, virtually no research has considered social support as dynamic across age. Here, we investigate the relationship between long-term patterns of disability, perceived and received social support, and depressive symptoms in later life. METHODS: We use random coefficient (growth) models of four waves of Duke Established Populations for Epidemiologic Studies of the Elderly data to determine whether disability, support, and depressive symptoms follow linear trajectories across age and whether support mediates the relationship between disability and depressive symptoms. RESULT: The results show that (a) trajectories of disability are strongly related to trajectories of depressive symptoms and (b) trajectories of perceived support mediate the relationship between trajectories of disability and depressive symptoms, whereas trajectories of received support do not. DISCUSSION: Disability, social support, and depressive symptoms are strongly interrelated processes in later life. Our results are consistent with previous research in showing that perceived, rather than received, support mediates the relationship between disability and depressive symptoms, but our results extend previous research in showing that this mediation occurs across time. PMID- 15294929 TI - Regulatory cells and infectious agents: detentes cordiale and contraire. AB - This brief review describes the types of interactions that occur between CD4(+)CD25(+) regulatory T cells (Treg) and microbial pathogens. These interactions range from one of mutual benefit (detente cordiale) such as occurs in Leishmania major infection of resistant mouse strains, to instances where the Treg response appears to mainly favor the pathogen and be detrimental to the host (detente contraire). Examples of the latter include chronic persistent infections with retroviruses, perhaps including HIV, and hepatitis C virus. The Treg response also hampers the effectiveness of immunity against some acute virus infections such as HSV. Evidence is also discussed showing that Treg can play a benevolent role to limit the severity of bystander tissue damage in circumstances where the immune response to pathogens is immunopathological. Finally, emerging approaches are discussed that either blunt or activate Treg and that could be used practically to manage host-pathogen interaction. PMID- 15294930 TI - Cutting edge: distinct roles for T help and CD40/CD40 ligand in regulating differentiation of proliferation-competent memory CD8+ T cells. AB - Murine primary antiviral cytotoxic T cell (CTL) responses are often induced in the absence of Th cells. In this study, we show that virus-like particles, if combined with DNA rich in CpG motifs, efficiently trigger primary CTL responses and comparable frequencies of memory CTLs in the presence or absence of T help. However, memory CTLs primed in the absence of T help failed to proliferate upon viral challenge. Nevertheless, they were efficiently recruited to sites of inflammation, indicating that T help may regulate the balance between proliferation-competent and migration-competent memory CTLs. Surprisingly, generation of proliferation-competent memory CTLs was completely independent of CD40 or CD40L, molecules commonly assumed to be central for mediating the beneficial effects of Th cells on CTL development. Thus, Th cells but not CD40/CD40L are key for the differentiation of proliferation-competent central memory CD8(+) T cells. PMID- 15294931 TI - Cutting edge: LFA-1 integrin-dependent T cell adhesion is regulated by both ag specificity and sensitivity. AB - Ab stimulation of the TCR rapidly enhances the functional activity of the LFA-1 integrin. Although TCR-mediated changes in LFA-1 activity are thought to promote T cell-APC interactions, the Ag specificity and sensitivity of TCR-mediated triggering of LFA-1 is not clear. We demonstrate that peptide/MHC (pMHC) tetramers rapidly enhance LFA-1-dependent adhesion of OT-I TCR transgenic CD8(+) T cells to purified ICAM-1. Inhibition of src family tyrosine kinase or PI3K activity blocked pMHC tetramer- and anti-CD3-stimulated adhesion. These effects are highly specific because partial agonist and antagonist pMHC tetramers are unable to stimulate OT-I T cell adhesion to ICAM-1. The Ag thresholds required for T cell adhesion to ICAM-1 resemble those of early T cell activation events, because optimal LFA-1 activation occurs at tetramer concentrations that fail to induce maximal T cell proliferation. Thus, TCR signaling to LFA-1 is highly Ag specific and sensitive to low concentrations of Ag. PMID- 15294932 TI - Cutting edge: estrogen drives expansion of the CD4+CD25+ regulatory T cell compartment. AB - CD4(+)CD25(+) regulatory T cells are crucial to the maintenance of tolerance in normal individuals. However, the factors regulating this cell population and its function are largely unknown. Estrogen has been shown to protect against the development of autoimmune disease, yet the mechanism is not known. We demonstrate that estrogen (17-beta-estradiol, E2) is capable of augmenting FoxP3 expression in vitro and in vivo. Treatment of naive mice with E2 increased both CD25(+) cell number and FoxP3 expression level. Further, the ability of E2 to protect against autoimmune disease (experimental autoimmune encephalomyelitis) correlated with its ability to up-regulate FoxP3, as both were reduced in estrogen receptor alpha deficient animals. Finally, E2 treatment and pregnancy induced FoxP3 protein expression to a similar degree, suggesting that high estrogen levels during pregnancy may help to maintain fetal tolerance. In summary, our data suggest E2 promotes tolerance by expanding the regulatory T cell compartment. PMID- 15294933 TI - Cutting edge: expression of chemokine receptor CXCR1 on human effector CD8+ T cells. AB - IL-8 is a potent inflammatory cytokine that induces chemotaxis of neutrophils expressing CXCR1 and CXCR2, thus indicating its involvement in the migration of these cells to inflammatory sites where bacteria proliferate. Presently, we showed that CXCR1(+) cells were predominantly found among CD8(+) T cells having effector phenotype, and that the expression of CXCR1 was positively correlated with that of perforin, suggesting that CXCR1 is expressed on effector CD8(+) T cells. Indeed, human CMV-specific CD8(+) T cells from healthy individuals, which mostly express the effector phenotype and have cytolytic function, expressed CXCR1, whereas EBV-specific CD8(+) T cells, which mostly express the memory phenotype and have no cytolytic function, did not express this receptor. The results of a chemotaxis assay showed that the migration of CXCR1(+)CD8(+) T cells was induced by IL-8. These results suggest that the IL-8-CXCR1 pathway plays an important role in the homing of effector CD8(+) T cells. PMID- 15294934 TI - Cutting edge: differential roles for phosphoinositide 3-kinases, p110gamma and p110delta, in lymphocyte chemotaxis and homing. AB - Despite the established role for PI3Ks in cell migration, the PI3Ks involved in lymphocyte chemotaxis are poorly defined. In this study, we report that p110gamma deficient T cells, but not B cells, show reduced chemotactic responses to the lymphoid chemokines, CCL19, CCL21, and CXCL12. As B cell and T cell chemotactic responses were both sensitive to the general PI3K inhibitors, wortmannin (WMN) and LY294002, we explored whether B cell responses were affected in mice lacking p110delta, a major PI3K isoform in lymphocytes. B cells deficient in p110delta showed diminished chemotactic responses, especially to CXCL13. Adoptive transfer experiments with WMN-treated wild-type B cells and with p110delta-deficient B cells revealed diminished homing to Peyer's patches and splenic white pulp cords. WMN selectively inhibited CXCR5-dependent B cell homing to Peyer's patches. These observations establish that p110gamma and p110delta function in lymphocyte chemotaxis, and show differential roles for PI3K family members in B and T cell migration. PMID- 15294935 TI - Cutting edge: prolonged antigen presentation after herpes simplex virus-1 skin infection. AB - It has been reported that MHC class I-restricted Ag presentation persists for only a short period following infection with certain pathogens, declining in parallel with the emergence of specific CTL activity. We have examined this issue in the case of murine infection with HSV-1. We found that the period of Ag presentation capable of priming naive CD8(+) T cells is comparatively prolonged, persisting for at least 7 days after infection, and continuing despite the appearance of localized CTL activity. Ag presentation was abbreviated to 3 or 4 days postinfection by surgical excision of the inoculation site early after infection. This intervention attenuated the size of the primary CTL response, implying that prolonged presentation is necessary to drive maximal CTL expansion. Combined, these data show that, in some types of infection, CTL priming can extend well beyond the first 24-48 h after primary inoculation. PMID- 15294936 TI - TNF family member B cell-activating factor (BAFF) receptor-dependent and independent roles for BAFF in B cell physiology. AB - The cytokine TNF family member B cell-activating factor (BAFF; also termed BLyS) is essential for B cell generation and maintenance. Three receptors have been identified that bind to BAFF: transmembrane activator, calcium modulator, and cyclophilin ligand interactor (TACI); B cell maturation Ag (BCMA); and BAFF-R. Recently, it was shown that A/WySnJ mice, which contain a dramatically reduced peripheral B cell compartment due to decreased B cell life span, express a mutant BAFF-R. This finding, together with normal or enhanced B cell generation in mice deficient for BCMA or TACI, respectively, suggested that the interaction of BAFF with BAFF-R triggers signals essential for the generation and maintenance of mature B cells. However, B cells in mice deficient for BAFF differ phenotypically and functionally from A/WySnJ B cells. Residual signaling through the mutant BAFF R could account for these differences. Alternatively, dominant-negative interference by the mutant receptor could lead to an overestimation of the importance of BAFF-R. To resolve this issue, we generated BAFF-R-null mice. Baff r(-/-) mice display strongly reduced late transitional and follicular B cell numbers and are essentially devoid of marginal zone B cells. Overexpression of Bcl-2 rescues mature B cell development in Baff-r(-/-) mice, suggesting that BAFF R mediates a survival signal. CD21 and CD23 surface expression are reduced on mature Baff-r(-/-) B cells, but not to the same extent as on mature B cells in BAFF-deficient mice. In addition, we found that Baff-r(-/-) mice mount significant, but reduced, Ag-specific Ab responses and are able to form spontaneous germinal centers in mesenteric lymph nodes. The reduction in Ab titers correlates with the reduced B cell numbers in the mutant mice. PMID- 15294937 TI - B7-CD28 interaction promotes proliferation and survival but suppresses differentiation of CD4-CD8- T cells in the thymus. AB - Costimulatory molecules play critical roles in the induction and effector function of T cells. More recent studies reveal that costimulatory molecules enhance clonal deletion of autoreactive T cells as well as generation and homeostasis of the CD25(+)CD4(+) regulatory T cells. However, it is unclear whether the costimulatory molecules play any role in the proliferation and differentiation of T cells before they acquire MHC-restricted TCR. In this study, we report that targeted mutations of B7-1 and B7-2 substantially reduce the proliferation and survival of CD4(-)CD8(-) (double-negative (DN)) T cells in the thymus. Perhaps as a result of reduced proliferation, the accumulation of RAG-2 protein in the DN thymocytes is increased in B7-deficient mice, which may explain the increased expression of TCR gene and accelerated transition of CD25(+)CD44(-) (DN3) to CD25(-)CD44(-) (DN4) stage. Qualitatively similar, but quantitatively less striking effects were observed in mice with a targeted mutation of CD28, but not CTLA4. Taken together, our results demonstrate that the development of DN in the thymus is subject to modulation by the B7-CD28 costimulatory pathway. PMID- 15294938 TI - Relevance of CD6-mediated interactions in T cell activation and proliferation. AB - CD6 is a cell surface receptor expressed on immature thymocytes and mature T and B1a lymphocytes. The ultimate function of CD6 has not been deciphered yet, but much evidence supports a role for CD6 in T cell activation and differentiation. In this study, we show that a fraction of CD6 molecules physically associates with the TCR/CD3 complex by coimmunoprecipitation, cocapping, and fluorescence resonance energy transfer experiments. Image analysis of Ag-specific T-APC conjugates demonstrated that CD6 and its ligand, activated leukocyte cell adhesion molecule (CD166), colocalize with TCR/CD3 at the center of the immunological synapse, the so-called central supramolecular activation cluster. The addition of a soluble rCD6 form significantly reduced the number of mature Ag specific T-APC conjugates, indicating that CD6 mediates early cell-cell interactions needed for immunological synapse maturation to proceed. This was in agreement with the dose-dependent inhibition of CD3-mediated T cell proliferation induced by soluble rCD6. Taken together, our data illustrate the important role played by the intra- and intercellular molecular interactions mediated by CD6 during T cell activation and proliferation processes. PMID- 15294939 TI - A stroma-derived defect in NF-kappaB2-/- mice causes impaired lymph node development and lymphocyte recruitment. AB - The NF-kappaB family of transcription factors is vital to all aspects of immune function and regulation in both the hemopoietic and stromal compartments of immune environments. Recent studies of mouse models deficient for specific members of the NF-kappaB family have revealed critical roles for these proteins in the process of secondary lymphoid tissue organogenesis. In this study, we investigate the role of NF-kappaB family member NF-kappaB2 in lymph node development and lymphocyte recruitment. Inguinal lymph nodes in nfkappab2(-/-) mice are reduced in size and cellularity, most notably in the B cell compartment. Using in vitro and in vivo lymph node grafting assays, we show that the defect resides in the stromal compartment. Further examination of the nfkappab2(-/-) inguinal lymph nodes revealed that expression of peripheral node addressin components CD34 and glycosylation-dependent cell adhesion molecule-1 along with the high endothelial venule-restricted sulfotransferase HEC-GlcNAc6ST was markedly reduced. Furthermore, expression of the lymphocyte homing chemokines CCL19, CCL21, and CXCL13 was down-regulated. These data highlight the role of NF kappaB2 in inguinal lymph node organogenesis and recruitment of lymphocytes to these organs due to its role in up-regulation of essential cell adhesion molecules and chemokines, while suggesting a potential role for NF-kappaB2 in organization of lymph node endothelium. PMID- 15294940 TI - A vitamin D analog down-regulates proinflammatory chemokine production by pancreatic islets inhibiting T cell recruitment and type 1 diabetes development. AB - Type 1 diabetes (T1D) is an autoimmune disease characterized by leukocyte infiltration into the pancreatic islets, and we have previously shown that treatment of adult NOD mice with a vitamin D analog arrests the progression of insulitis, blocks Th1 cell infiltration into the pancreas, and markedly reduces T1D development, suggesting inhibition of chemokine production by islet cells. In this study, we show that all TLRs are expressed by mouse and human islet cells, and their engagement by pathogen-derived ligands markedly enhances proinflammatory chemokine production. The vitamin D analog significantly down regulates in vitro and in vivo proinflammatory chemokine production by islet cells, inhibiting T cell recruitment into the pancreatic islets and T1D development. The inhibition of islet chemokine production in vivo persists after restimulation with TLR ligands and is associated with up-regulation of IkappaBalpha transcription, an inhibitor of NF-kappaB and with arrest of NF kappaBp65 nuclear translocation, highlighting a novel mechanism of action exerted by vitamin D receptor ligands potentially relevant for the treatment of T1D and other autoimmune diseases. PMID- 15294941 TI - Immunoprevention of mammary carcinoma in HER-2/neu transgenic mice is IFN-gamma and B cell dependent. AB - A vaccine combining IL-12 and allogeneic mammary carcinoma cells expressing p185(neu) completely prevents tumor onset in HER-2/neu transgenic BALB/c mice (NeuT mice). The immune protection elicited was independent from CTL activity. We now formally prove that tumor prevention is mainly based on the production of anti-p185(neu) Abs. In the present studies, NeuT mice were crossed with knockout mice lacking IFN-gamma production (IFN-gamma(-/-)) or with B cell-deficient mice (microMT). Vaccination did not protect NeuT-IFN-gamma(-/-) mice, thus confirming a central role of IFN-gamma. The block of Ab production in NeuT-microMT mice was incomplete. About one third of NeuT-microMT mice failed to produce Abs and displayed a rapid tumor onset. By contrast, those NeuT-microMT mice that responded to the vaccine with a robust production of anti-p185(neu) Ab displayed a markedly delayed tumor onset. In these NeuT-microMT mice, the vaccine induced a lower level of IgG2a and IgG3 and a higher level of IgG2b than in NeuT mice. Moreover, NeuT-microMT mice failed to produce anti-MHC class I Abs in response to allogeneic H-2(q) molecules present in the cell vaccine. These findings show that inhibition of HER-2/neu carcinogenesis depends on cytokines and specific Abs, and that a highly effective vaccine can rescue Ab production even in B cell-deficient mice. PMID- 15294942 TI - Analyses of the in vivo trafficking of stoichiometric doses of an anti-complement receptor 1/2 monoclonal antibody infused intravenously in mice. AB - Complement plays a critical role in the immune response by opsonizing immune complexes (IC) and thymus-independent type 2 Ags with C3 breakdown product C3dg, a CR2-specific ligand. We used a C3dg-opsonized IC model, anti-CR1/2 mAb 7G6, to investigate how such substrates are processed. We used RIA, whole body imaging, flow cytometry, and fluorescence immunohistochemistry to examine the disposition of 0.1- to 2-microg quantities of mAb 7G6 infused i.v. into BALB/c mice. The mAb is rapidly taken up by the spleen and binds preferentially to marginal zone (MZ) B cells; within 24 h, the MZ B cells relocate and transfer mAb 7G6 to follicular dendritic cells (FDC). Transfer occurs coincident with loss of the extracellular portion of MZ B cell CR2, suggesting that the process may be mediated by proteolysis of CR2. Intravenous infusion of an FDC-specific mAb does not induce comparable splenic localization or cellular reorganization, emphasizing the importance of MZ B cells in intrasplenic trafficking of bound substrates. We propose the following mechanism: binding of C3dg-opsonized IC to noncognate MZ B cells promotes migration of these cells to the white pulp, followed by CR2 proteolysis, which allows transfer of the opsonized IC to FDC, thus facilitating presentation of intact Ags to cognate B cells. PMID- 15294943 TI - STAT5 is required for thymopoiesis in a development stage-specific manner. AB - Diverse cytokines necessary for normal lymphopoiesis and lymphocyte homeostasis activate STAT5 in responder cells. Although STAT5 has been suggested to be a central molecular effecter of IL-7 function, its essential role during IL-7 dependent T cell development in vivo remained unclear. Using Stat5(-/-) mice we now show that STAT5 is essential for various functions ascribed to IL-7 in vivo. STAT5 is required for embryonic thymocyte production, TCRgamma gene transcription, and Peyer's patch development. In sharp contrast, normal STAT5 is dispensable for adult thymopoiesis. In peripheral lymphocytes, STAT5 is primarily required for the generation and/or maintenance of gammadelta T cells and TCRgammadelta(+) intraepithelial lymphocytes. Collectively, these results demonstrate that STAT5 is critical for many, but not all, aspects of steady state lymphoid lineage development and maintenance and suggest the existence of previously undocumented cytokine signaling traits and/or cytokine milieu during adult thymopoiesis. PMID- 15294944 TI - Genetic control of autoimmunity: protection from diabetes, but spontaneous autoimmune biliary disease in a nonobese diabetic congenic strain. AB - At least 20 insulin-dependent diabetes (Idd) loci modify the progression of autoimmune diabetes in the NOD mouse, an animal model of human type 1 diabetes. The NOD.c3c4 congenic mouse, which has multiple B6- and B10-derived Idd-resistant alleles on chromosomes 3 and 4, respectively, is completely protected from autoimmune diabetes. We demonstrate in this study, however, that NOD.c3c4 mice develop a novel spontaneous and fatal autoimmune polycystic biliary tract disease, with lymphocytic peribiliary infiltrates and autoantibodies. Strains having a subset of the Idd-resistant alleles present in the NOD.c3c4 strain show component phenotypes of the liver disease: NOD mice with B6 resistance alleles only on chromosome 3 have lymphocytic liver infiltration without autoantibody formation, while NOD mice with B10 resistance alleles only on chromosome 4 show autoantibody formation without liver infiltration. The liver disease is transferable to naive NOD.c3c4 recipients using splenocytes from affected NOD.c3c4 mice, demonstrating an autoimmune etiology. Thus, substitution of non NOD genetic intervals into the NOD strain can prevent diabetes, but in turn cause an entirely different autoimmune syndrome, a finding consistent with a generalized failure of self-tolerance in the NOD genetic background. The complex clinical phenotypes in human autoimmune conditions may be similarly resolved into largely overlapping biochemical pathways that are then modified, potentially by alleles at a few key chromosomal regions, to produce specific autoimmune syndromes. PMID- 15294945 TI - Regulated expression of the receptor-like tyrosine phosphatase CD148 on hemopoietic cells. AB - CD148 is a receptor-like protein tyrosine phosphatase expressed on a wide variety of cell types. Through the use flow cytometry and immunofluorescence microscopy on tissue sections, we examined the expression of CD148 on multiple murine hemopoietic cell lineages. We found that CD148 is moderately expressed during all stages of B cell development in the bone marrow, as well as peripheral mature B cells. In contrast, CD148 expression on thymocytes and mature T cells is substantially lower. However, stimulation of peripheral T cells through the TCR leads to an increase of CD148 expression. This up-regulation on T cells can be partially inhibited by reagents that block the activity of src family kinases, calcineurin, MEK, or PI3K. Interestingly, CD148 levels are elevated on freshly isolated T cells from MRL lpr/lpr and CTLA-4-deficient mice, two murine models of autoimmunity. Together, these expression data along with previous biochemical data suggest that CD148 may play an important regulatory role to control an immune response. PMID- 15294946 TI - B cell-activating factor belonging to the TNF family acts through separate receptors to support B cell survival and T cell-independent antibody formation. AB - The TNF-related ligand, B cell-activating factor belonging to the TNF family (BAFF), is necessary for normal B cell development and survival, and specifically binds the receptors transmembrane activator and calcium-modulator and cyclophilin ligand interactor (TACI), B cell maturation Ag (BCMA), and BAFF-R. Similarities between mice completely lacking BAFF and A/WySnJ strain mice that express a naturally occurring mutant form of BAFF-R suggest that BAFF acts primarily through BAFF-R. However, the nearly full-length BAFF-R protein expressed by A/WySnJ mice makes unambiguous interpretation of receptor function in these animals impossible. Using homologous recombination we created mice completely lacking BAFF-R and compared them directly to A/WySnJ mice and to mice lacking BAFF. BAFF-R-null mice exhibit loss of mature B cells similar to that observed in BAFF(-/-) and A/WySnJ mice. Also, mice lacking both TACI and BCMA simultaneously exhibit no B cell loss, thus confirming that BAFF-R is the primary receptor for transmitting the BAFF-dependent B cell survival signal. However, while BAFF-R null mice cannot carry out T cell-dependent Ab formation, they differ from BAFF deficient mice in generating normal levels of Ab to at least some T cell independent Ags. These studies clearly demonstrate that BAFF regulates Ab responses in vivo through receptors in addition to BAFF-R. PMID- 15294947 TI - TIRC7 deficiency causes in vitro and in vivo augmentation of T and B cell activation and cytokine response. AB - The membrane protein T cell immune response cDNA 7 (TIRC7) was recently identified and was shown to play an important role in T cell activation. To characterize the function of TIRC7 in more detail, we generated TIRC7-deficient mice by gene targeting. We observed disturbed T and B cell function both in vitro and in vivo in TIRC7(-/-) mice. Histologically, primary and secondary lymphoid organs showed a mixture of hypo-, hyper-, and dysplastic changes of multiple lymphohemopoietic compartments. T cells from TIRC7(-/-) mice exhibited significantly increased proliferation and expression of IL-2, IFN-gamma, and IL-4 in response to different stimuli. Resting T cells from TIRC7(-/-) mice exhibited decreased CD62L, but increased CD11a and CD44 expression, suggesting an in vivo expansion of memory/effector T cells. Remarkably, activated T cells from TIRC7(-/ ) mice expressed lower levels of CTLA-4 in comparison with wild-type cells. B cells from TIRC7-deficient mice exhibited significantly higher in vitro proliferation following stimulation with anti-CD40 Ab or LPS plus IL-4. B cell hyperreactivity was reflected in vivo by elevated serum levels of various Ig classes and higher CD86 expression on B cells. Furthermore, TIRC7 deficiency resulted in an augmented delayed-type hypersensitivity response that was also reflected in increased mononuclear infiltration in the skin obtained from TIRC7 deficient mice food pads. In summary, the data strongly support an important role for TIRC7 in regulating both T and B cell responses. PMID- 15294948 TI - Initiation of immune responses in brain is promoted by local dendritic cells. AB - The contribution of dendritic cells (DCs) to initiating T cell-mediated immune response in and T cell homing into the CNS has not yet been clarified. In this study we show by confocal microscopy and flow cytometry that cells expressing CD11c, CD205, and MHC class II molecules and containing fluorescently labeled, processed Ag accumulate at the site of intracerebral Ag injection. These cells follow a specific pattern upon migrating out of the brain. To track their pathway out of the CNS, we differentiated DCs from bone marrow of GFP-transgenic mice and injected them directly into brains of naive C57BL/6 mice. We demonstrate that DCs migrate from brain to cervical lymph nodes, a process that can be blocked by fixation or pertussis toxin treatment of the DCs. Injection of OVA-loaded DCs into brain initiates a SIINFEKL (a dominant OVA epitope)-specific T cell response in lymph nodes and spleen, as measured by specific tetramer and LFA-1 activation marker staining. Additionally, a fraction of activated SIINFEKL-specific T cells home to the CNS. Specific T cell homing to the CNS, however, cannot be induced by i.v. injection of OVA-loaded DCs alone. These data suggest that brain-emigrant DCs are sufficient to support activated T cells to home to the tissue of DC origination. Thus, initiation of immune reactivity against CNS Ags involves the migration of APCs from nervous tissue to peripheral lymphoid tissues, similarly to that in other organs. PMID- 15294949 TI - IL-10-producing B220+CD11c- APC in mouse spleen. AB - APC acting at the early stages of an immune response can shape the nature of that response. Such APC will include dendritic cells (DCs) but may also include populations of B cells such as marginal zone B cells in the spleen. In this study, we analyze APC populations in mouse spleen and compare the phenotype and function of B220(+)CD11c(-) populations with those of CD11c(+) spleen DC subsets. Low-density B220(+) cells had morphology similar to DCs and, like DCs, they could stimulate naive T cells, and expressed high levels of MHC and costimulatory molecules. However, the majority of the B220(+) cells appeared to be of B cell lineage as demonstrated by coexpression of CD19 and surface Ig, and by their absence from RAG-2(-/-) mice. The phenotype of these DC-like B cells was consistent with that of B cells in the marginal zone of the spleen. On bacterial stimulation, they preferentially produced IL-10 in contrast to the DCs, which produced IL-12. Conventional B cells did not produce IL-10. The DC-like B cells could be induced to express low levels of the DC marker CD11c with maturational stimuli. A minority of the B220(+)CD11c(-) low-density cells did not express CD19 and surface Ig and may be a DC subset; this population also produced IL-10 on bacterial stimulation. B220(+) APC in mouse spleen that stimulate naive T cells and preferentially produce IL-10 may be involved in activating regulatory immune responses. PMID- 15294950 TI - Activation through cannabinoid receptors 1 and 2 on dendritic cells triggers NF kappaB-dependent apoptosis: novel role for endogenous and exogenous cannabinoids in immunoregulation. AB - The precise role of cannabinoid receptors (CB)1 and CB2, as well as endogenous ligands for these receptors, on immune cells remains unclear. In the current study, we examined the effect of endogenous and exogenous cannabinoids on murine bone marrow-derived dendritic cells (DCs). Addition of Delta(9) tetrahydrocannabinol (THC), a major psychoactive component found in marijuana or anandamide, an endogenous cannabinoid, to DC cultures induced apoptosis in DCs. DCs expressed CB1 and CB2 receptors and the engagement of both receptors was necessary to trigger apoptosis. Treatment with THC induced caspase-2, -8, and -9 activation, cleavage of Bid, decreased mitochondrial membrane potential, and cytochrome c release, suggesting involvement of death-receptor and mitochondrial pathways. DCs from Bid-knockout mice were sensitive to THC-induced apoptosis thereby suggesting that Bid was dispensable. There was no induction of p44/p42 MAPK, p38 MAPK, or stress-activated protein/JNK pathway in THC-treated DCs. However, THC treatment induced phosphorylation of IkappaB-alpha, and enhanced the transcription of several apoptotic genes regulated by NF-kappaB. Moreover, inhibition of NF-kappaB was able to block THC-induced apoptosis in DCs. Lastly, in vivo treatment of mice with THC caused depletion of splenic DCs. Together, our study demonstrates for the first time that endogenous and exogenous cannabinoids may suppress the immune response through their ability to induce apoptosis in DCs. PMID- 15294951 TI - Identification of multiple cell cycle regulatory functions of p57Kip2 in human T lymphocytes. AB - The specific functions of p57(Kip2) in lymphocytes have not yet been fully elucidated. In this study, it is shown that p57(Kip2), which is a member of the Cip/Kip family of cyclin-dependent kinase inhibitors, is present in the nuclei of normal resting (G(0)) T cells from peripheral blood and in the nuclei of the T cell-derived Jurkat cell line. Activation through the TCR results in rapid transport of cytoplasmic cyclin-dependent kinase 6 (cdk6) to nuclei, where it associates with cyclin D and p57(Kip2) in active enzyme complexes. Using purified recombinant proteins, it was shown in vitro that addition of p57(Kip2) protein to a mixture of cyclin D2 and cdk6 enhanced the association of the latter two proteins and resulted in phosphorylation of p57(Kip2). To probe further the function of p57(Kip2), Jurkat cells stably transfected with a plasmid encoding p57(Kip2) under control of an inducible (tetracycline) promoter were made. Induction of p57(Kip2) resulted in increased association of cdk6 with cyclin D3, without receptor-mediated T cell stimulation. The overall amounts of cdk6 and cyclin D3, and also of cdk4 and cyclin E, remained unchanged. Most notably, increased p57(Kip2) levels resulted in marked inhibition of both cyclin E- and cyclin A-associated cdk2 kinase activities and a decrease in cyclin A amounts. Therefore, although facilitating activation of cdk6, the ultimate outcome of p57(Kip2) induction was a decrease in DNA synthesis and cell proliferation. The results indicate that p57(Kip2) is involved in the regulation of several aspects of the T cell cycle. PMID- 15294952 TI - Membrane-associated heparan sulfate proteoglycans are involved in the recognition of cellular targets by NKp30 and NKp46. AB - Lysis of virus-infected and tumor cells by NK cells is mediated via natural cytotoxicity receptors (NCRs). We have recently shown that the NKp44 and NKp46 NCRs, but not the NKp30, recognize viral hemagglutinins. In this study we explored the nature of the cellular ligands recognized by the NKp30 and NKp46 NCRs. We demonstrate that target cell surface heparan sulfate proteoglycans (HSPGs) are recognized by NKp30 and NKp46 and that 6-O-sulfation and N acetylation state of the glucose building unit affect this recognition and lysis by NK cells. Tumor cells expressing cell surface heparanase, CHO cells lacking membranal heparan sulfate and glypican-1-suppressed pancreatic cancer cells manifest reduced recognition by NKp30 and NKp46 and are lysed to a lesser extent by NK cells. Our results are the first clue for the identity of the ligands for NKp30 and NKp46. Whether the ligands are particular HSPGs, unusual heparan sulfate epitopes, or a complex of HSPGs and either other protein or lipid moieties remains to be further explored. PMID- 15294953 TI - TNF-alpha controls intrahepatic T cell apoptosis and peripheral T cell numbers. AB - At the end of an immune response, activated lymphocyte populations contract, leaving only a small memory population. The deletion of CD8(+) T cells from the periphery is associated with an accumulation of CD8(+) T cells in the liver, resulting in both CD8(+) T cell apoptosis and liver damage. After adoptive transfer and in vivo activation of TCR transgenic CD8(+) T cells, an increased number of activated CD8(+) T cells was observed in the lymph nodes, spleen, and liver of mice treated with anti-TNF-alpha. However, caspase activity was decreased only in CD8(+) T cells in the liver, not in those in the lymphoid organs. These results indicate that TNF-alpha is responsible for inducing apoptosis in the liver and suggest that CD8(+) T cells escaping this mechanism of deletion can recirculate into the periphery. PMID- 15294954 TI - CD8+ T cells specific for EBV, cytomegalovirus, and influenza virus are activated during primary HIV infection. AB - Primary viral infections, including primary HIV infection, trigger intense activation of the immune system, with marked expansion of CD38(+)CD8(+) T cells. Whether this expansion involves only viral-specific cells or includes a degree of bystander activation remains a matter of debate. We therefore examined the activation status of EBV-, CMV-, and influenza virus (FLU)-specific CD8(+) T cells during primary HIV infection, in comparison to HIV-specific CD8(+) T cells. The activation markers CD38 and HLA-DR were strongly expressed on HIV-specific CD8(+) T cells. Surprisingly, CD38 expression was also up-regulated on CD8(+) T cells specific for other viruses, albeit to a lesser extent. Activation marker expression returned to normal or near-normal values after 1 year of highly active antiretroviral therapy. HIV viral load correlated with CD38 expression on HIV specific CD8(+) T cells but also on EBV-, CMV-, and FLU-specific CD8(+) T cells. In primary HIV infection, EBV-specific CD8(+) T cells also showed increased Ki67 expression and decreased Bcl-2 expression, compared with values observed in HIV seronegative control subjects. These results show that bystander activation occurs during primary HIV infection, even though HIV-specific CD8(+) T cells express the highest level of activation. The role of this bystander activation in lymphocyte homeostasis and HIV pathogenesis remains to be determined. PMID- 15294955 TI - Parasite-induced Th2 polarization is associated with down-regulated dendritic cell responsiveness to Th1 stimuli and a transient delay in T lymphocyte cycling. AB - The nature of the signals that bias Th effector choice is still not completely understood. Using parasite extracts from pathogens known to induce polarized Th1 or Th2 responses and an in vitro experimental model for priming murine CD4(+) cells, we demonstrated that splenic dendritic cells (DC), but not B cells, promote Th1/Th2 differentiation of naive CD4(+) lymphocytes. Th polarization in this system was found not to depend on DC secretion of the polarizing cytokines IL-12/IL-4, but instead correlated with distinct states of DC activation induced by the different parasite preparations. As expected, conditioning of DC for Th1 development was associated with up-regulation of costimulatory molecules and enhanced chemokine production and required intact MyD88 signaling. In contrast, conditioning of DC for Th2 differentiation correlated with down-regulation of many of the same functions and was MyD88 independent. This dampened DC activation was accompanied in the cocultures by a reduction in the frequency of CD4(+) lymphocytes exiting the first division of the cell cycle. When the latter was mimicked by drug-induced arrest of peptide-primed CD4(+) cells after the S phase of the first cycle, a marked Th2 polarization was also observed. Together, these findings suggest that the emergence of IL-4-producing CD4(+) lymphocytes results from a suppression in DC function leading to a temporary delay in initial T cell cycling. PMID- 15294956 TI - The 4-1BB costimulation augments the proliferation of CD4+CD25+ regulatory T cells. AB - The thymus-derived CD4(+)CD25(+) T cells belong to a subset of regulatory T cells potentially capable of suppressing the proliferation of pathogenic effector T cells. Intriguingly, these suppressor cells are themselves anergic, proliferating poorly to mitogenic stimulation in culture. In this study, we find that the 4-1BB costimulator receptor, best known for promoting the proliferation and survival of CD8(+) T cells, also induces the proliferation of the CD4(+)CD25(+) regulatory T cells both in culture and in vivo. The proliferating CD4(+)CD25(+) T cells produce no detectable IL-2, suggesting that 4-1BB costimulation of these cells does not involve IL-2 production. The 4-1BB-expanded CD4(+)CD25(+) T cells are functional, as they remain suppressive to other T cells in coculture. These results support the notion that the peripheral expansion of the CD4(+)CD25(+) T cells is controlled in part by costimulation. PMID- 15294957 TI - Estrogen receptor alpha signaling in inflammatory leukocytes is dispensable for 17beta-estradiol-mediated inhibition of experimental autoimmune encephalomyelitis. AB - Estrogen treatment has been shown to exert a protective effect on experimental autoimmune encephalomyelitis (EAE), and is under clinical trial for multiple sclerosis. Although it is commonly assumed that estrogens exert their effect by modulating immune functions, we show in this study that 17beta-estradiol (E2) treatment can inhibit mouse EAE without affecting autoantigen-specific T cell responsiveness and type 1 cytokine production. Using mutant mice in which estrogen receptor alpha (ERalpha) has been unambiguously inactivated, we found that ERalpha was responsible for the E2-mediated inhibition of EAE. We next generated irradiation bone marrow chimeras in which ERalpha expression was selectively impaired in inflammatory T lymphocytes or was limited to the radiosensitive hemopoietic compartment. Our data show that the protective effect of E2 on clinical EAE and CNS inflammation was not dependent on ERalpha signaling in inflammatory T cells. Likewise, EAE development was not prevented by E2 treatment in chimeric mice that selectively expressed ERalpha in the systemic immune compartment. In conclusion, our data demonstrate that the beneficial effect of E2 on this autoimmune disease does not involve ERalpha signaling in blood-derived inflammatory cells, and indicate that ERalpha expressed in other tissues, such as CNS-resident microglia or endothelial cells, mediates this effect. PMID- 15294958 TI - CD40 engagement enhances antigen-presenting langerhans cell priming of IFN-gamma producing CD4+ and CD8+ T cells independently of IL-12. AB - The delivery of CD40 signaling to APCs during T cell priming enhances many T cell mediated immune responses. Although CD40 signaling up-regulates APC production of IL-12, the impact of this increased production on T cell priming is unclear. In this study an IL-12-independent T cell-mediated immune response, contact hypersensitivity (CHS), was used to further investigate the effect of CD40 ligation on the phenotypic development of Ag-specific CD4(+) and CD8(+) T cells. Normally, sensitization for CHS responses induces hapten-specific CD4(+) T cells producing type 2 cytokines and CD8(+) T cells producing IFN-gamma. Treatment of mice with agonist anti-CD40 mAb during sensitization with the hapten 2,4 dinitrofluorobenzene resulted in CHS responses of increased magnitude and duration. These augmented responses in anti-CD40 Ab-treated mice correlated with increased numbers of hapten-specific CD4(+) and CD8(+) T cells producing IFN gamma in the skin draining lymph nodes. Identical results were observed using IL 12(-/-) mice, indicating that CD40 ligation promotes CHS responses and development of IFN-gamma-producing CD4(+) and CD8(+) T cells in the absence of IL 12. Engagement of CD40 on hapten-presenting Langerhans cells (hpLC) up-regulated the expression of both class I and class II MHC and promoted hpLC migration into the T cell priming site. These results indicate that hpLC stimulated by CD40 ligation use a mechanism distinct from increased IL-12 production to promote Ag specific T cell development to IFN-gamma-producing cells. PMID- 15294959 TI - CD40 ligand rescues inhibitor of differentiation 3-mediated G1 arrest induced by anti-IgM in WEHI-231 B lymphoma cells. AB - The engagement of membrane-bound Igs (mIgs) results in growth arrest, accompanied by apoptosis, in the WEHI-231 murine B lymphoma cells, a cell line model representative of primary immature B cells. Inhibitor of differentiation (Id) proteins, members of the helix-loop-helix protein family, functions in proliferation, differentiation, and apoptosis in a variety of cell types. In this study, we analyzed the involvement of Id protein in mIg-induced growth arrest and apoptosis in WEHI-231 cells. Following stimulation with anti-IgM, expression of Id3 was up-regulated at both the mRNA and protein levels; this up-regulation could be reversed by CD40L treatment. Retrovirus-mediated transduction of the Id3 gene into WEHI-231 cells resulted in an accumulation of the cells in G(1) phase, but did not induce apoptosis. E box-binding activity decreased in response to anti-IgM administration, but increased after stimulation with either CD40L alone or anti-IgM plus CD40L, suggesting that E box-binding activity correlates with cell cycle progression. WEHI-231 cells overexpressing Id3 accumulated in G(1) phase, which was accompanied by reduced levels of cyclin D2, cyclin E, and cyclin A, and a reciprocal up-regulation of p27(Kip1). Both the helix-loop-helix and the C-terminal regions of Id3 were required for growth-suppressive activity. These data suggest that Id3 mimics mIg-mediated G(1) arrest in WEHI-231 cells. PMID- 15294960 TI - Ionizing radiation affects human MART-1 melanoma antigen processing and presentation by dendritic cells. AB - Radiation is generally considered to be an immunosuppressive agent that acts by killing radiosensitive lymphocytes. In this study, we demonstrate the noncytotoxic effects of ionizing radiation on MHC class I Ag presentation by bone marrow-derived dendritic cells (DCs) that have divergent consequences depending upon whether peptides are endogenously processed and loaded onto MHC class I molecules or are added exogenously. The endogenous pathway was examined using C57BL/6 murine DCs transduced with adenovirus to express the human melanoma/melanocyte Ag recognized by T cells (AdVMART1). Prior irradiation abrogated the ability of AdVMART1-transduced DCs to induce MART-1-specific T cell responses following their injection into mice. The ability of these same DCs to generate protective immunity against B16 melanoma, which expresses murine MART-1, was also abrogated by radiation. Failure of AdVMART1-transduced DCs to generate antitumor immunity following irradiation was not due to cytotoxicity or to radiation-induced block in DC maturation or loss in expression of MHC class I or costimulatory molecules. Expression of some of these molecules was affected, but because irradiation actually enhanced the ability of DCs to generate lymphocyte responses to the peptide MART-1(27-35) that is immunodominant in the context of HLA-A2.1, they were unlikely to be critical. The increase in lymphocyte reactivity generated by irradiated DCs pulsed with MART-1(27-35) also protected mice against growth of B16-A2/K(b) tumors in HLA-A2.1/K(b) transgenic mice. Taken together, these results suggest that radiation modulates MHC class I-mediated antitumor immunity by functionally affecting DC Ag presentation pathways. PMID- 15294961 TI - A Structural basis for the association of DAP12 with mouse, but not human, NKG2D. AB - Prior studies have revealed that alternative mRNA splicing of the mouse NKG2D gene generates receptors that associate with either the DAP10 or DAP12 transmembrane adapter signaling proteins. We report that NKG2D function is normal in human patients lacking functional DAP12, indicating that DAP10 is sufficient for human NKG2D signal transduction. Further, we show that human NKG2D is incapable of associating with DAP12 and provide evidence that structural differences in the transmembrane of mouse and human NKG2D account for the species specific difference for this immune receptor. PMID- 15294962 TI - Induction of IgG2a class switching in B cells by IL-27. AB - IL-27 is a novel IL-12 family member that plays a role in the early regulation of Th1 initiation. However, its role in B cells remains unexplored. We here show a role for IL-27 in the induction of T-bet expression and regulation of Ig class switching in B cells. Expression of WSX-1, one subunit of IL-27R, was detected at the mRNA level in primary mouse spleen B cells, and stimulation of these B cells by IL-27 rapidly activated STAT1. IL-27 then induced T-bet expression and IgG2a, but not IgG1, class switching in B cells activated with anti-CD40 or LPS. In contrast, IL-27 inhibited IgG1 class switching induced by IL-4 in activated B cells. Similar induction of STAT1 activation, T-bet expression and IgG2a class switching was observed in IFN-gamma-deficient B cells, but not in STAT1-deficient ones. The induction of IgG2a class switching was abolished in T-bet-deficient B cells activated with LPS. These results suggest that primary spleen B cells express functional IL-27R and that the stimulation of these B cells by IL-27 induces T-bet expression and IgG2a, but not IgG1, class switching in a STAT1 dependent but IFN-gamma-independent manner. The IL-27-induced IgG2a class switching is highly dependent on T-bet in response to T-independent stimuli such as LPS. Thus, IL-27 may be a novel attractive candidate as a therapeutic agent against diseases such as allergic disorders by not only regulating Th1 differentiation but also directly acting on B cells and inducing IgG2a class switching. PMID- 15294963 TI - The positive regulatory effect of TGF-beta2 on primitive murine hemopoietic stem and progenitor cells is dependent on age, genetic background, and serum factors. AB - TGF-beta is considered a negative regulator of hemopoietic stem and progenitor cells. We have previously shown that one TGF-beta isoform, TGF-beta2, is, in fact, a positive regulator of murine hemopoietic stem cell function in vivo. In vitro, TGF-beta2, but not TGF-beta1 and TGF-beta3, had a biphasic dose response on the proliferation of purified lin-Sca1(++)kit(+) (LSK) cells, with a stimulatory effect at low concentrations, which was subject to mouse strain dependent variation. In this study we report that the stimulatory effect of TGF beta2 on the proliferation of LSK cells increases with age and after replicative stress in C57BL/6, but not in DBA/2, mice. The age-related changes in the TGF beta2 effect correlated with life span in BXD recombinant strains. The stimulatory effect of TGF-beta2 on the proliferation of LSK cells requires one or more nonprotein, low m.w. factors present in fetal calf and mouse sera. The activity of this factor(s) in mouse serum increases with age. Taken together, our data suggest a role for TGF-beta2 and as yet unknown serum factors in the aging of the hemopoietic stem cell compartment and possibly in organismal aging. PMID- 15294964 TI - Functional characterization of MHC class II-restricted CD8+CD4- and CD8-CD4- T cell responses to infection in CD4-/- mice. AB - Classical CD4(+) and CD8(+) T cells recognize Ag presented by MHC class II (MHCII) and MHC class I (MHCI), respectively. However, our results show that CD4( /-) mice mount a strong, readily detectable CD8(+) T cell response to MHCII restricted epitopes after a primary bacterial or viral infection. These MHCII restricted CD8(+)CD4(-) T cells are more similar to classical CD8(+) T cells than to CD4(+) T cells in their expression of effector functions during a primary infection, yet they also differ from MHCI-restricted CD8(+) T cells by their inability to produce high levels of the cytolytic molecule granzyme B. After resolution of a primary infection, epitope-specific MHCII-restricted T cells in CD4(-/-) mice persist for a long period of time as memory T cells. Surprisingly, upon reinfection the secondary MHCII-restricted response in CD4(-/-) mice consists mainly of CD8(-)CD4(-) T cells. In contrast to CD8(+) T cells, MHCII restricted CD8(-)CD4(-) T cells are capable of producing IL-2 in addition to IFN gamma and thus appear to have attributes characteristic of CD4(+) T cells rather than CD8(+) T cells. Therefore, MHCII-restricted T cells in CD4(-/-) mice do not share all phenotypic and functional characteristics with MHCI-restricted CD8(+) T cells or with MHCII-restricted CD4(+) T cells, but, rather, adopt attributes from each of these subsets. These results have implications for understanding thymic T cell selection and for elucidating the mechanisms regulating the peripheral immune response and memory differentiation. PMID- 15294965 TI - Murine B7-H3 is a negative regulator of T cells. AB - T cell activation is regulated by the innate immune system through positive and negative costimulatory molecules. B7-H3 is a novel B7-like molecule with a putative receptor on activated T cells. Human B7-H3 was first described as a positive costimulator, most potently inducing IFN-gamma production and cellular immunity. In this study we examined the expression and function of mouse B7-H3. B7-H3 is mostly expressed on professional APCs; its expression on dendritic cells appears to be up-regulated by LPS. In contrast to human B7-H3, we found that mouse B7-H3 protein inhibited T cell activation and effector cytokine production. An antagonistic mAb to B7-H3 enhanced T cell proliferation in vitro and led to exacerbated experimental autoimmune encephalomyelitis in vivo. Therefore, mouse B7-H3 serves as a negative regulator of T cell activation and function. PMID- 15294966 TI - Up-regulation of vascular endothelial growth factor (VEGF) in small-for-size liver grafts enhances macrophage activities through VEGF receptor 2-dependent pathway. AB - This study aims to investigate the potential role of vascular endothelial growth factor (VEGF) and VEGF-R2 (fetal liver kinase (Flk)-1) in mediating macrophage activities in small-for-size liver transplantation. A rat orthotopic liver transplantation model was performed using either whole, 50, or 30% liver grafts (both 50 and 30% were regarded as small-for-size) in syngeneic or allogeneic combinations, respectively. Firstly, the mRNA and protein levels of VEGF and Flk 1 in liver grafts were detected by RT-PCR and Western blot, and the number of Flk 1(+) macrophages (labeled by ED1) was determined by flow cytometry. It was found that the small-for-size isografts and allografts presented higher levels of VEGF and Flk-1 expression than the whole isograft and allograft. In addition, a higher number of Flk-1(+)ED1(+) cells were detected in the small-for-size isografts and allografts than the whole isograft and allograft. Secondly, our study revealed that macrophage cell lines did not initially express detectable Flk-1, but could be induced by VEGF, and the inducible expression of Flk-1 in macrophages was related to their migration and proliferation activities. Finally, our study demonstrated that the induction of Flk-1 expression on macrophages by VEGF was associated with the expression of NF-kappaB and heat shock protein 90. In conclusion, the present study showed that the up-regulated expression of VEGF and its interaction with Flk-1 in small-for-size liver grafts might facilitate the activities of macrophages. PMID- 15294967 TI - Early expression of a functional TCRbeta chain inhibits TCRgamma gene rearrangements without altering the frequency of TCRgammadelta lineage cells. AB - To investigate the consequences of the simultaneous expression in progenitor cells of a TCRgammadelta and a pre-TCR on alphabeta/gammadelta lineage commitment, we have forced expression of functionally rearranged TCRbeta, TCRgamma, and TCRdelta chains by means of transgenes. Mice transgenic for the three TCR chains contain numbers of gammadelta thymocytes comparable to those of mice transgenic for both TCRgamma and TCRdelta chains, and numbers of alphabeta thymocytes similar to those found in mice solely transgenic for a rearranged TCRbeta chain gene. gammadelta T cells from the triple transgenic mice express the transgenic TCRbeta chain, but do not express a TCRalpha chain, and, by a number of phenotypic and molecular parameters, appear to be bona fide gammadelta thymocytes. Our results reveal a remarkable degree of independence in the generation of alphabeta and gammadelta lineage cells from progenitor cells that, in theory, could simultaneously express a TCRgammadelta and a pre-TCR. PMID- 15294968 TI - Complement C5a receptor is essential for the optimal generation of antiviral CD8+ T cell responses. AB - The complement system has been long regarded as an important effector of the innate immune response. Furthermore, complement contributes to various aspects of B and T cell immunity. Nevertheless, the role of complement in CD8(+) T cell antiviral responses has yet to be fully delineated. We examined the CD8(+) T cell response in influenza type A virus-infected mice treated with a peptide antagonist to C5aR to test the potential role of complement components in CD8(+) T cell responses. We show that both the frequency and absolute numbers of flu specific CD8(+) T cells are greatly reduced in C5aR antagonist-treated mice compared with untreated mice. This reduction in flu-specific CD8(+) T cells is accompanied by attenuated antiviral cytolytic activity in the lungs. These results demonstrate that the binding of the C5a component of complement to the C5a receptor plays an important role in CD8(+) T cell responses. PMID- 15294969 TI - Requirement for both H-2Db and H-2Kd for the induction of diabetes by the promiscuous CD8+ T cell clonotype AI4. AB - The NOD mouse is a model for autoimmune type 1 diabetes in humans. CD8(+) T cells are essential for the destruction of the insulin-producing pancreatic beta cells characterizing this disease. AI4 is a pathogenic CD8(+) T cell clone, isolated from the islets of a 5-wk-old female NOD mouse, which is capable of mediating overt diabetes in the absence of CD4(+) T cell help. Recent studies using MHC congenic NOD mice revealed marked promiscuity of the AI4 TCR, as the selection of this clonotype can be influenced by multiple MHC molecules, including some class II variants. The present work was designed, in part, to determine whether similar promiscuity also characterizes the effector function of mature AI4 CTL. Using splenocyte and bone marrow disease transfer models and in vitro islet-killing assays, we report that efficient recognition and destruction of beta cells by AI4 requires the beta cells to simultaneously express both H-2D(b) and H-2K(d) class I MHC molecules. The ability of the AI4 TCR to interact with both H-2D(b) and H 2K(d) was confirmed using recombinant peptide libraries. This approach also allowed us to define a mimotope peptide recognized by AI4 in an H-2D(b) restricted manner. Using ELISPOT and mimotope/H-2D(b) tetramer analyses, we demonstrate for the first time that AI4 represents a readily detectable T cell population in the islet infiltrates of prediabetic NOD mice. Our identification of a ligand for AI4-like T cells will facilitate further characterization and manipulation of this pathogenic and promiscuous T cell population. PMID- 15294970 TI - CD45-deficient mice accumulate Pro-B cells both in vivo and in vitro. AB - Efficient generation of mature B lineage cells requires the participation of the BCR, the pre-BCR, accessory coreceptors, and growth factor receptors. Together these receptors integrate cell intrinsic signals with regulatory pathways initiated by surrounding cells and structures. CD45 is a receptor tyrosine phosphatase expressed at high levels on all hemopoietic cells, and has been shown to modulate many signaling cascades in both positive and negative manners. In the absence of B220, the B lineage isoform of CD45, differentiation to the mature B cell stage is incomplete. We demonstrate that CD45-deficient mice also accumulate pro-B cells in the bone marrow. In vitro differentiation is altered in that B lineage populations exhibit prolonged survival in the presence of high concentrations of IL-7. Cell lines derived from CD45-deficient animals experience prolonged JAK/STAT activation in response to IL-7 stimulation, and constitutively elevated levels of phosphorylated src kinases. Aberrant IL-7Ralpha expression is observed in vivo, and may be responsible for the skewed development present in CD45(-/-) animals. Demonstrating that CD45-deficient pro-B cells are affected by the absence of B220 highlights a previously unrecognized parallel in B and T lineage precursors, and emphasizes that the presence of normal numbers of peripheral B cells does not assure that the bone marrow compartment is intact. PMID- 15294971 TI - Transcriptional regulation of the human TLR9 gene. AB - To clarify the molecular basis of human TLR9 (hTLR9) gene expression, the activity of the hTLR9 gene promoter was characterized using the human myeloma cell line RPMI 8226. Reporter gene analysis and EMSA demonstrated that hTLR9 gene transcription was regulated via four cis-acting elements, cAMP response element, 5'-PU box, 3'-PU box, and a C/EBP site, that interacted with the CREB1, Ets2, Elf1, Elk1, and C/EBPalpha transcription factors. Other members of the C/EBP family, such as C/EBPbeta, C/EBPdelta, and C/EBPepsilon, were also important for TLR9 gene transcription. CpG DNA-mediated suppression of TLR9 gene transcription led to decreased binding of the trans-acting factors to their corresponding cis acting elements. It appeared that suppression was mediated via c-Jun and NF kappaB p65 and that cooperation among CREB1, Ets2, Elf1, Elk1, and C/EBPalpha culminated in maximal transcription of the TLR9 gene. These findings will help to elucidate the mechanism of TLR9 gene regulation and to provide insight into the process by which TLR9 evolved in the mammalian immune system. PMID- 15294972 TI - Human lymphocytes interact directly with CD47 through a novel member of the signal regulatory protein (SIRP) family. AB - Two closely related proteins, signal regulatory protein alpha (SIRPalpha; SHPS 1/CD172) and SIRPbeta, have been described in humans. The existence of a third SIRP protein has been suggested by cDNA sequence only. We show that this third SIRP is a separate gene that is expressed as a protein with unique characteristics from both alpha and beta genes and suggest that this gene should be termed SIRPgamma. We have expressed the extracellular region of SIRPgamma as a soluble protein and have shown that, like SIRPalpha, it binds CD47, but with a lower affinity (K(d), approximately 23 microM) compared with SIRPalpha (K(d), approximately 2 microM). mAbs specific to SIRPgamma show that it was not expressed on myeloid cells, in contrast to SIRPalpha and -beta, being expressed instead on the majority of T cells and a proportion of B cells. The short cytoplasmic tail of SIRPgamma does not contain any known signaling motifs, nor does it contain a characteristic lysine, as with SIRPbeta, that is required for DAP12 interaction. DAP12 coexpression is a requirement for SIRPbeta surface expression, whereas SIRPgamma is expressed in its absence. The SIRPgamma-CD47 interaction may therefore not be capable of bidirectional signaling as with the SIRPalpha-CD47, but, instead, use unidirectional signaling via CD47 only. PMID- 15294973 TI - c-Fos as a regulator of degranulation and cytokine production in FcepsilonRI activated mast cells. AB - The AP-1 complex is composed of c-Jun and c-Fos and is a key component in the regulation of proinflammatory genes. Mast cells play a significant role in the initiation of many inflammatory responses, such as allergy and allergy-associated diseases. In the present work, we characterized the role of c-Fos in mast cell function by investigating IL-3-dependent cell proliferation, degranulation capability, and cytokine expression in c-Fos-deficient mice. In c-Fos-deficient mast cells, we found that FcepsilonRI-mediated degranulation was significantly inhibited, which correlates with the reduced expression of SWAP-70, VAMP-7, and Synaptotagmin I genes, which are involved directly in the degranulation process. These findings show that c-Fos plays an important role in FcepsilonRI-mediated regulation of mast cell function. PMID- 15294974 TI - pH-triggered microparticles for peptide vaccination. AB - Improving vaccine delivery to human APCs is a way to increase the CTL response to vaccines. We report the use of a novel pH-triggered microparticle that exploits the ability of APCs to cross-present MHC I-restricted Ags that have been engulfed in the low pH environment of the phagosome. A model MHC class I-restricted peptide Ag from the influenza A matrix protein was encapsulated in spray-dried microparticles composed of dipalmitoylphosphatidylcholine and the pH-sensitive polymethacrylate Eudragit E100. Release of the peptide from the particle was triggered by a drop in pH to the acidity normally found in the phagosome. The particles were efficiently phagocytosed by human monocytes and dendritic cells with minimal cellular toxicity and no functional impairment. Encapsulation of the peptide in the microparticles resulted in efficient presentation of the peptide to CD8(+) T cells by human dendritic cells in vitro, and was superior to unencapsulated peptide or peptide encapsulated in an analogous pH-insensitive particle. Vaccination of human HLA-A*0201 transgenic mice with peptide encapsulated in pH-triggering microparticles resulted in priming of CTL responses. These microparticles can be modified to coencapsulate a range of adjuvants along with the Ag of interest. Encapsulation of MHC I epitopes in pH triggered microparticles increases Ag presentation and may improve CD8(+) T cell priming to peptide vaccines against viruses and cancer. PMID- 15294975 TI - Endoplasmic reticulum-associated degradation-induced dissociation of class II invariant chain complexes containing a glycosylation-deficient form of p41. AB - The quality control system in the secretory pathway can identify and eliminate misfolded proteins through endoplasmic reticulum-associated degradation (ERAD). ERAD is thought to occur by retrotranslocation through the Sec61 complex into the cytosol and degradation by the proteasome. However, the extent of disassembly of oligomeric proteins and unfolding of polypeptide chains that is required for retrotranslocation is not fully understood. In this report we used a glycosylation mutant of the p41 isoform of invariant chain (Ii) to evaluate the ability of ERAD to discriminate between correctly folded and misfolded subunits in an oligomeric complex. We show that loss of glycosylation at position 239 of p41 does not detectably affect Ii trimerization or association with class II but does result in a defect in endoplasmic reticulum export of Ii that ultimately leads to its degradation via the ERAD pathway. Although class II associated with the mutated form of p41 is initially retained in the endoplasmic reticulum, it is subsequently released and traffics through the Golgi to the plasma membrane. ERAD mediated degradation of the mutant p41 is dependent on mannose trimming and inhibition of mannosidase I stabilizes Ii. Interestingly, inhibition of mannosidase I also results in prolonged association between the mutant Ii and class II, indicating that complex disassembly and release of class II is linked to mannosidase-dependent ERAD targeting of the misfolded Ii. These results suggest that the ERAD machinery can induce subunit disassembly, specifically targeting misfolded subunits to degradation and sparing properly folded subunits for reassembly and/or export. PMID- 15294976 TI - Mechanisms regulating the positioning of mouse p47 resistance GTPases LRG-47 and IIGP1 on cellular membranes: retargeting to plasma membrane induced by phagocytosis. AB - The recently identified p47 GTPases are one of the most effective cell-autonomous resistance systems known against intracellular pathogens in the mouse. One member of the family, LRG-47, has been shown to be essential for immune control in vivo of Listeria monocytogenes, Toxoplasma gondii, Mycobacterium tuberculosis, and Mycobacterium avium, possibly by promoting acidification of the phagosome. However, the intracellular localization of LRG-47, and the nature of its association with the phagosomal or any other membrane system is unknown. In this study, we show that LRG-47 is a Golgi-associated protein in the IFN-stimulated cell, which is rapidly recruited to active plasma membrane upon phagocytosis and remains associated with phagosomes as they mature. We show that the Golgi localization of LRG-47 is dependent on the integrity of an amphipathic helix near the C terminus, whereas the plasma membrane localization depends on an unidentified signal associated with the G domain. Unlike LRG-47, but like the published p47 resistance GTPase, IGTP, a further p47 GTPase, IIGP1, is associated with the endoplasmic reticulum. However, unlike IGTP, IIGP1 is associated with the endoplasmic reticulum by an N-terminal myristoylation modification. Thus, the p47 GTPases are a diverse battery of intracellular defense factors dynamically associated with different membrane systems. PMID- 15294977 TI - Characterization of bovine homologues of granulysin and NK-lysin. AB - Granulysin and NK-lysin are antimicrobial proteins found in the granules of human and swine cytotoxic lymphocytes. A murine counterpart to granulysin has not been identified to date, indicating the importance of additional models to fully characterize the role of granulysin-like molecules in the immune response to infectious disease. Two partial nucleotide sequences corresponding to the complete functional domain of granulysin and NK-lysin were amplified from bovine PBMC mRNA. Following stimulation with phorbol ester and calcium ionophore, expression of the bovine gene was detected in CD3(+) T cells, CD4(+) T cells, CD8(+) T cells, WC1(+) gammadelta T cells, and PBMC depleted of CD3(+) T cells, but was absent in CD21(+) cells and CD14(+) cells. Intracellular flow cytometry and immunoblotting confirmed the presence of protein corresponding to the bovine granulysin homologue in activated T lymphocytes and PBMC. Synthetic human, bovine, and swine peptides corresponding to the C terminus of helix 2 through helix 3 region of granulysin displayed potent antimicrobial activity against Escherichia coli, Salmonella enteritidis, Staphylococcus aureus, and Mycobacterium bovis bacillus Calmette-Guerin. Human and bovine peptides corresponding to helix 2 displayed antimycobacterial activity against M. bovis bacillus Calmette-Guerin. Expression of the bovine gene was detected in laser microscopy-dissected lymph node lesions from an M. bovis-infected animal. The identification of a biologically active bovine homologue to granulysin demonstrates the potential of the bovine model in characterizing the role of granulysin in the immune response to a variety of infectious agents. PMID- 15294978 TI - Phospholipases D1 and D2 coordinately regulate macrophage phagocytosis. AB - Phagocytosis is a fundamental feature of the innate immune system, required for antimicrobial defense, resolution of inflammation, and tissue remodeling. Furthermore, phagocytosis is coupled to a diverse range of cytotoxic effector mechanisms, including the respiratory burst, secretion of inflammatory mediators and Ag presentation. Phospholipase D (PLD) has been linked to the regulation of phagocytosis and subsequent effector responses, but the identity of the PLD isoform(s) involved and the molecular mechanisms of activation are unknown. We used primary human macrophages and human THP-1 promonocytes to characterize the role of PLD in phagocytosis. Macrophages, THP-1 cells, and other human myelomonocytic cells expressed both PLD1 and PLD2 proteins. Phagocytosis of complement-opsonized zymosan was associated with stimulation of the activity of both PLD1 and PLD2, as demonstrated by a novel immunoprecipitation-in vitro PLD assay. Transfection of dominant-negative PLD1 or PLD2 each inhibited the extent of phagocytosis (by 55-65%), and their combined effects were additive (reduction of 91%). PLD1 and PLD2 exhibited distinct localizations in resting macrophages and those undergoing phagocytosis, and only PLD1 localized to the phagosome membrane. The COS-7 monkey fibroblast cell line, which has been used as a heterologous system for the analysis of receptor-mediated phagocytosis, expressed PLD2 but not PLD1. These data support a model in which macrophage phagocytosis is coordinately regulated by both PLD1 and PLD2, with isoform-specific localization. Human myelomonocytic cell lines accurately model PLD-dependent signal transduction events required for phagocytosis, but the heterologous COS cell system does not. PMID- 15294979 TI - Human papillomavirus type-16 virus-like particles activate complementary defense responses in key dendritic cell subpopulations. AB - Human papillomavirus type-16 (HPV16) L1 virus-like particles (VLPs) activate dendritic cells (DCs) and induce protective immunity. In this study, we demonstrate, using global gene expression analysis, that HPV16 VLPs produce quite distinct innate responses in murine splenic DC subpopulations. While HPV16 VLPs increase transcription of IFN-gamma and numerous Th1-related cytokines and chemokines in CD8alpha(+)CD11c(+) DCs, CD4(+)CD11c(+) DCs up-regulate only type I IFN and a different set of Th2-associated cytokines and chemokines. Type I IFN, but not IFN-gamma, potentiates humoral immunity, notably production of VLP specific IgG2a. However, HPV16 VLP-stimulated IL-12 production by CD8alpha(+)CD11c(+) DCs is augmented by autocrine IFN-gamma signaling. Thus, before adaptive immunity, HPV16 VLPs signal complementary defense responses in key DC subpopulations, indicating specialized DC lineages with predetermined polarization. PMID- 15294980 TI - Functional inactivation of immature dendritic cells by the intracellular parasite Toxoplasma gondii. AB - Despite its noted ability to induce strong cellular immunity, and its known susceptibility to IFN-gamma-dependent immune effector mechanisms, the protozoan Toxoplasma gondii is a highly successful parasite, able to replicate, disseminate, and either kill the host or, more commonly, establish resistant encysted life forms before the emergence of protective immune responses. We sought to understand how the parasite gains the advantage. Using transgenic clonal parasite lines engineered to express fluorescent markers in combination with dendritic cells (DC) grown from the bone marrow of wild-type mice or transgenic mice expressing fluorescent protein-tagged MHC class II molecules, we used flow cytometry and fluorescence microscopy to analyze the responses of infected DC to both invasion by the parasite and subsequent DC maturation signals. We found that T. gondii preferentially invades immature dendritic cells but fails to activate them in the process, and renders them resistant to subsequent activation by TLR ligands or the immune-system-intrinsic maturation signal CD40L. The functional consequences of T. gondii-mediated suppression of DC activation are manifested in a relative inability of infected immature DC to activate naive CD4(+) Th lymphocytes, or to secrete cytokines, such IL-12 and TNF alpha, that play important roles in innate and/or adaptive immunity. The findings reveal that T. gondii suppresses the ability of immature DC to participate in innate immunity and to induce adaptive immune responses. The ability of T. gondii to temporarily evade recognition could provide a selective advantage that permits dissemination and establishment before adaptive immune response initiation. PMID- 15294981 TI - Cytoplasmic entry of Listeria monocytogenes enhances dendritic cell maturation and T cell differentiation and function. AB - Protective immunity to the intracellular bacterial pathogen, Listeria monocytogenes, is mediated by a vigorous T cell response. In particular, CD8(+) cytolytic T cells provide essential effector function in the clearance of bacterial infection. The cytoplasmic entry of Listeria facilitated by listeriolysin O is an essential feature not only of the bacteria's virulence, but of the ability of the bacteria to elicit protective immunity in the host. To determine how cytoplasmic entry of Listeria regulates the development of protective immunity, we examined the effects of this process on the maturation of murine dendritic cells (DC) and on their ability to prime naive CD8(+) T cell responses. Costimulatory molecules (CD40, CD80, and CD86) were induced by listerial infection only when the bacteria invaded the cytoplasm. In addition, the production of IL-12, IL-10, IL-6, and TNF-alpha was most efficiently triggered by cytosolic Listeria. Naive T cells primed by peptide-loaded DC infected with either wild-type or nonhemolytic mutant Listeria proliferated equivalently, but a much larger proportion of those primed by wild-type Listeria monocytogenes produced IFN-gamma. Costimulatory molecules induced by cytosolic entry regulated T cell proliferation and, as a result, the number of functional T cells generated. DC-produced cytokines (specifically IL-12 and IL-10) were the major factors determining the proportion of T cells producing IFN-gamma. These data highlight the requirement for listerial cytoplasmic invasion for the optimal priming of T cell cytokine production and attest to the importance of this event to the development of protective CTL responses to this pathogen. PMID- 15294982 TI - Temporin A and related frog antimicrobial peptides use formyl peptide receptor like 1 as a receptor to chemoattract phagocytes. AB - Many mammalian antimicrobial peptides (AMPs) have multiple effects on antimicrobial immunity. We found that temporin A (TA), a representative frog derived AMP, induced the migration of human monocytes, neutrophils, and macrophages with a bell-shaped response curve in a pertussis toxin-sensitive manner, activated p44/42 MAPK, and stimulated Ca(2+) flux in monocytes, suggesting that TA is capable of chemoattracting phagocytic leukocytes by the use of a G(ialpha) protein-coupled receptor. TA-induced Ca(2+) flux in monocytes was cross-desensitized by an agonistic ligand MMK-1 specific for formyl peptide receptor-like 1 (FPRL1) and vice versa, suggesting that TA uses FPRL1 as a receptor. This conclusion was confirmed by data showing that TA selectively stimulated chemotaxis of HEK 293 cells transfected with human FPRL1 or its mouse ortholog, murine formyl peptide receptor 2. In addition, TA elicited the infiltration of neutrophils and monocytes into the injection site of mice, indicating that TA is also functionally chemotactic in vivo. Examination of two additional temporins revealed that Rana-6 was also able to attract human phagocytes using FPRL1, but temporin 1P selectively induced the migration of neutrophils using a distinct receptor. Comparison of the chemotactic and antimicrobial activities of several synthetic analogues suggested that these activities are likely to rely on different structural characteristics. Overall, the results demonstrate that certain frog-derived temporins have the capacity to chemoattract phagocytes by the use of human FPRL1 (or its orthologs in other species), providing the first evidence suggesting the potential participation of certain amphibian antimicrobial peptides in host antimicrobial immunity. PMID- 15294983 TI - Mycobacterium tuberculosis LprG (Rv1411c): a novel TLR-2 ligand that inhibits human macrophage class II MHC antigen processing. AB - MHC class II (MHC-II)-restricted CD4(+) T cells are essential for control of Mycobacterium tuberculosis infection. This report describes the identification and purification of LprG (Rv1411c) as an inhibitor of primary human macrophage MHC-II Ag processing. LprG is a 24-kDa lipoprotein found in the M. tuberculosis cell wall. Prolonged exposure (>16 h) of human macrophages to LprG resulted in marked inhibition of MHC-II Ag processing. Inhibition of MHC-II Ag processing was dependent on TLR-2. Short-term exposure (<6 h) to LprG stimulated TLR-2-dependent TNF-alpha production. Thus, LprG can exploit TLR-2 signaling to inhibit MHC-II Ag processing in human macrophages. Inhibition of MHC-II Ag processing by mycobacterial lipoproteins may allow M. tuberculosis, within infected macrophages, to avoid recognition by CD4(+) T cells. PMID- 15294984 TI - Partial activation of neonatal CD11c+ dendritic cells and induction of adult-like CD8+ cytotoxic T cell responses by synthetic microspheres. AB - Neonatal cytotoxic T cell responses have only been elicited to date with immunogens or delivery systems inducing potent direct APC activation. To define the minimal activation requirements for the induction of neonatal CD8(+) cytotoxic responses, we used synthetic microspheres (MS) coated with a single CD8(+) T cell peptide from lymphocytic choriomeningitis virus (LCMV) or HIV-1. Unexpectedly, a single injection of peptide-conjugated MS without added adjuvant induced CD4-dependent Ag-specific neonatal murine cytotoxic responses with adult like CTL precursor frequency, avidity for Ag, and frequency of IFN-gamma secreting CD8(+) splenocytes. Neonatal CD8(+) T cell responses to MS-LCMV were elicited within 2 wk of a single immunization and, upon challenge, provided similar protection from viral replication as adult CTLs, demonstrating their in vivo competence. As previously reported, peptide-coated MS elicited no detectable activation of adult CD11c(+) dendritic cells (DC). In contrast, CTL responses were associated with a partial activation of neonatal CD11c(+) DC, reflected by the up-regulation of CD80 and CD86 expression but no concurrent changes in MHC class II or CD40 expression. However, this partial activation of neonatal DC was not sufficient to circumvent the requirement for CD4(+) T cell help. The effective induction of neonatal CD8(+) T cell responses by this minimal Ag delivery system demonstrates that neonatal CD11c(+) DC may mature sufficiently to stimulate naive CD8(+) neonatal T cells, even in the absence of strong maturation signals. PMID- 15294985 TI - Cloning and characterization of chicken IL-10 and its role in the immune response to Eimeria maxima. AB - We isolated the full-length chicken IL-10 (chIL-10) cDNA from an expressed sequence tag library derived from RNA from cecal tonsils of Eimeria tenella infected chickens. It encodes a 178-aa polypeptide, with a predicted 162-aa mature peptide. Chicken IL-10 has 45 and 42% aa identity with human and murine IL 10, respectively. The structures of the chIL-10 gene and its promoter were determined by direct sequencing of a bacterial artificial chromosome containing chIL-10. The chIL-10 gene structure is similar to (five exons, four introns), but more compact than, that of its mammalian orthologues. The promoter is more similar to that of Fugu IL-10 than human IL-10. Chicken IL-10 mRNA expression was identified mainly in the bursa of Fabricius and cecal tonsils, with low levels of expression also seen in thymus, liver, and lung. Expression was also detected in PHA-activated thymocytes and LPS-stimulated monocyte-derived macrophages, with high expression in an LPS-stimulated macrophage cell line. Recombinant chIL-10 was produced and bioactivity demonstrated through IL-10-induced inhibition of IFN gamma synthesis by mitogen-activated lymphocytes. We measured the expression of mRNA for chIL-10 and other signature cytokines in gut and spleen of resistant (line C.B12) and susceptible (line 15I) chickens during the course of an E. maxima infection. Susceptible chickens showed higher levels of chIL-10 mRNA expression in the spleen, both constitutively and after infection, and in the small intestine after infection than did resistant chickens. These data indicate a potential role for chIL-10 in changing the Th bias during infection with an intracellular protozoan, thereby contributing to susceptibility of line 15I chickens. PMID- 15294986 TI - Lipopolysaccharide binding protein binds to triacylated and diacylated lipopeptides and mediates innate immune responses. AB - LPS binding protein (LBP) is an acute-phase protein synthesized predominantly in the liver of the mammalian host. It was first described to bind LPS of Gram negative bacteria and transfer it via a CD14-enhanced mechanism to a receptor complex including TLR-4 and MD-2, initiating a signal transduction cascade leading to the release of proinflammatory cytokines. In recent studies, we found that LBP also mediates cytokine induction caused by compounds derived from Gram positive bacteria, including lipoteichoic acid and peptidoglycan fragments. Lipoproteins and lipopeptides have repeatedly been shown to act as potent cytokine inducers, interacting with TLR-2, in synergy with TLR-1 or -6. In this study, we show that these compounds also interact with LBP and CD14. We used triacylated lipopeptides, corresponding to lipoproteins of Borrelia burgdorferi, mycobacteria, and Escherichia coli, as well as diacylated lipopeptides, corresponding to, e.g., 2-kDa macrophage activating lipopeptide of Mycoplasma spp. Activation of Chinese hamster ovary cells transfected with TLR-2 by both lipopeptides was enhanced by cotransfection of CD14. Responsiveness of human mononuclear cells to these compounds was greatly enhanced in the presence of human LBP. Binding of lipopeptides to LBP as well as competitive inhibition of this interaction by LPS was demonstrated in a microplate assay. Furthermore, we were able to show that LBP transfers lipopeptides to CD14 on human monocytes using FACS analysis. These results support that LBP is a pattern recognition receptor transferring a variety of bacterial ligands including the two major types of lipopeptides to CD14 present in different receptor complexes. PMID- 15294987 TI - Oral administration with papillomavirus pseudovirus encoding IL-2 fully restores mucosal and systemic immune responses to vaccinations in aged mice. AB - Infectious diseases are one of the major threats for the elderly because their immune system is often compromised, and vaccinations to prevent these infections are not effective. A major defect in their immune system seems to be the inability of T cells to produce IL-2. We used papillomavirus (PV) pseudoviruses (PSVs) as a model vaccine and a gene delivery vector to address how to enhance immune responses to vaccinations. We found that oral immunization with PV PSV induced minimal mucosal and systemic Abs and CTLs specific for the PSVs in aged mice compared with young adult mice. In addition, fewer specific Th cells were generated in the aged mice. When aged mice were immunized with PV PSVs encoding human IL-2, specific Th cells were generated, producing murine IL-2, IL-4, and IFN-gamma. Further, specific Abs and CTLs were induced, resulting in protection against mucosal viral challenge. Thus, this study provided a basis for clinical trials using PV PSVs encoding IL-2 for vaccination of the elderly. PMID- 15294988 TI - A secreted protein from the human hookworm necator americanus binds selectively to NK cells and induces IFN-gamma production. AB - Parasitic helminths induce chronic infections in their hosts although, with most human helminthiases, protective immunity gradually develops with age or exposure of the host. One exception is infection with the human hookworm, Necator americanus, where virtually no protection ensues over time. Such observations suggest these parasites have developed unique mechanisms to evade host immunity, leading us to investigate the role of the excretory/secretory (ES) products of adult N. americanus in manipulating host immune responses. Specifically, we found that a protein(s) from ES products of adult N. americanus bound selectively to mouse and human NK cells. Moreover, incubation of purified NK cells with N. americanus ES products stimulated the production of augmented (4- to 30-fold) levels of IFN-gamma. This augmentation was dependent on the presence of both IL-2 and IL-12 and was endotoxin-independent. This is the first report of a pathogen protein that binds exclusively to NK cells and the first report of a nematode derived product that induces abundant levels of cytokines from NK cells. Such an interaction could provide a means of cross-regulating deleterious Th2 immune responses in the host, thereby contributing to the long-term survival of N. americanus. PMID- 15294989 TI - IL-15-independent proliferative renewal of memory CD8+ T cells in latent gammaherpesvirus infection. AB - IL-15 is known to be critical in the homeostasis of Ag-specific memory CD8(+) T cells following acute viral infection. However, little is known about the homeostatic requirements of memory CD8(+) T cells during a latent viral infection. We have used the murine gammaherpesvirus-68 (MHV-68) model system to investigate whether IL-15 is necessary for the maintenance of memory CD8(+) T cells during a latent viral infection. IL-15 is not essential either for the initial control of MHV-68 infection or for the maintenance of MHV-68-specific memory CD8(+) T cells. Even at 140 days postinfection, the proportion of CD8(+) T cells recognizing the MHV-68 epitopes were the same as in control mice. The maintenance of these memory CD8(+) T cells was attributable to their ability to turn over in vivo, probably in response to the presence of low levels of Ag. IL 15(-/-) mice had a significantly higher turnover rate within the virus-specific memory CD8(+) T cell population, which was the result of increased levels of viral gene expression rather than an increase in viral load. These cells did not accumulate in the spleens of the IL-15(-/-) mice due to an increased sensitivity to apoptosis as a result of decreased Bcl-2 levels. Intriguingly, memory CD8(+) T cells from latently infected mice failed to undergo homeostatic proliferation in a naive secondary host. These data highlight fundamental differences between memory CD8(+) T cells engaged in active immune surveillance of latent viral infections vs memory CD8(+) T cells found after acute viral infections. PMID- 15294990 TI - Suppression of innate immunity by acute ethanol administration: a global perspective and a new mechanism beginning with inhibition of signaling through TLR3. AB - Excessive consumption of ethanol (EtOH) suppresses innate immunity, but the mechanisms have not been fully delineated. The present study was conducted to determine whether EtOH suppresses TLR signaling in vivo in mice and to characterize the downstream effects of such suppression. Degradation of IL-1R associated kinase 1 induced by a TLR3 ligand in peritoneal cells ( approximately 90% macrophages) was suppressed by EtOH. Phosphorylation of p38 kinase in peritoneal macrophages (F4/80(+)) was suppressed, as was nuclear translocation of p-c-Jun and p65 in peritoneal cells. EtOH decreased IL-6 and IL-12 (p40), but did not significantly affect IL-10 in peritoneal lavage fluid or in lysates of peritoneal cells. Changes in cytokine mRNAs (by RNase protection assay) in macrophages isolated by cell sorting or using Ficoll were generally consistent with changes in protein levels in cell lysates and peritoneal lavage fluid. Thus, suppression of TLR signaling and cytokine mRNA occurred in the same cells, and this suppression generally corresponded to changes in i.p. and intracellular cytokine concentrations. DNA microarray analysis revealed the suppression of an IFN-related amplification loop in peritoneal macrophages, associated with decreased expression of numerous innate immune effector genes (including cytokines and a chemokine also suppressed at the protein level). These results indicate that EtOH suppresses innate immunity at least in part by suppressing TLR3 signaling, suppressing an IFN-related amplification loop, and suppressing the induction of a wide range of innate effector molecules in addition to proinflammatory cytokines and chemokines. PMID- 15294991 TI - The induction of acute ileitis by a single microbial antigen of Toxoplasma gondii. AB - The role of specific microbial Ags in the induction of experimental inflammatory bowel disease is poorly understood. Oral infection of susceptible C57BL/6 mice with Toxoplasma gondii results in a lethal ileitis within 7-9 days postinfection. An immunodominant Ag of T. gondii (surface Ag 1 (SAG1)) that induces a robust B and T cell-specific response has been identified and a SAG1-deficient parasite (Deltasag1) engineered. We investigated the ability of Deltasag1 parasite to induce a lethal intestinal inflammatory response in susceptible mice. C57BL/6 mice orally infected with Deltasag1 parasites failed to develop ileitis. In vitro, the mutant parasites replicate in both enterocytes and dendritic cells. In vivo, infection with the mutant parasites was associated with a decrease in the chemokine and cytokine production within several compartments of the gut associated cell population. RAG-deficient (RAG1(-/-)) mice are resistant to the development of the ileitis after T. gondii infection. Adoptive transfer of Ag specific CD4(+) effector T lymphocytes isolated from C57BL/6-infected mice into RAG(-/-) mice conferred susceptibility to the development of the intestinal disease. In contrast, CD4(+) effector T lymphocytes from mice infected with the mutant Deltasag1 strain failed to transfer the pathology. In addition, resistant mice (BALB/c) that fail to develop ileitis following oral infection with T. gondii were rendered susceptible following intranasal presensitization with the SAG1 protein. This process was associated with a shift toward a Th1 response. These findings demonstrate that a single Ag (SAG1) of T. gondii can elicit a lethal inflammatory process in this experimental model of pathogen-driven ileitis. PMID- 15294992 TI - Tolerance induced by the lipopeptide Pam3Cys is due to ablation of IL-1R associated kinase-1. AB - Stimulation of the human monocytic cell line Mono Mac 6 with the synthetic lipopeptide (S)-(2,3-bis(palmitoyloxy)-(2RS)-propyl)-N-palmitoyl-(R)-Cys-(S) Ser(S)-Lys(4)-OH, trihydrochloride (Pam(3)Cys) at 10 microg/ml induces a rapid expression of the TNF gene in a TLR2-dependent fashion. Preculture of the cells with Pam(3)Cys at 1 microg/ml leads to a reduced response after subsequent stimulation with Pam(3)Cys at 10 microg/ml, indicating that the cells have become tolerant to Pam(3)Cys. The CD14 and TLR2 expression is not decreased on the surface of the tolerant cells, but rather up-regulated. Analysis of the NF-kappaB binding in Pam(3)Cys-tolerant cells shows a failure to mobilize NF-kappaB-p50p65 heterodimers, while NF-kappaB-p50p50 homodimers remain unchanged. Pam(3)Cys tolerant cells showed neither IkappaBalpha-Ser(32) phosphorylation nor IkappaBalpha degradation but MyD88 protein was unaltered. However, IRAK-1 protein was absent in Pam(3)Cys-induced tolerance, while IRAK-1 mRNA was still detectable at 30% compared with untreated cells. In contrast, in LPS-tolerized cells, p50p50 homodimers were induced, IRAK-1 protein level was only partially decreased, and p50p65 mobilization remained intact. It is concluded that in Mono Mac 6 monocytic cells, inhibition of IRAK-1 expression at the mRNA and protein levels is the main TLR-2-dependent mechanism responsible for Pam(3)Cys-induced tolerance, but not for TLR-4-dependent LPS-induced tolerance. PMID- 15294993 TI - Impact of fibronectin fragments on the transendothelial migration of HIV-infected leukocytes and the development of subendothelial foci of infectious leukocytes. AB - Leukocyte infiltrates that can serve as viral reservoirs, and sites for viral replication are found in many organs of HIV-1-infected patients. Patients whose blood leukocytes migrate across confluent endothelial monolayers ex vivo and transmit infectious virus to mononuclear leukocytes (MNLs) lodged beneath this endothelial barrier have a worse prognosis. We evaluated the ability of 110- to 120-kDa fibronectin fragments (FNf), which are found in the blood of >60% of HIV 1-infected patients, to stimulate transendothelial migration and drive productively infected MNLs into a potential perivascular space. FNf induced MNLs to release TNF-alpha in a dose-dependent fashion; the resulting increase in lymphocyte and monocyte transendothelial migration could be blocked with soluble TNF receptor I. Rather than penetrate deeply into the subendothelial matrix, as is seen with untreated controls, FNf-treated MNLs clustered just below the endothelial monolayer. Treatment with FNf during migration increased subsequent recovery of HIV-infected cells from the subendothelial compartment. FNf treatment also significantly increased the numbers of HLA-DR(bright), dendritic-type cells that reverse-migrated from the subendothelial depot to the apical endothelial surface 48 h after migration. Fibronectin fragments can be produced by viral and host proteases in the course of inflammatory conditions. The ability of FNf to stimulate transendothelial migration of HIV-1-infected MNLs may help to explain the dissemination of this infection into cardiac, renal, and CNS tissues. PMID- 15294994 TI - Toll IL-1 receptors differ in their ability to promote the stabilization of adenosine and uridine-rich elements containing mRNA. AB - Several ligands for Toll IL-1R (TIR) family are known to promote stabilization of a subset of short-lived mRNAs containing AU-rich elements (AREs) in their 3' untranslated regions. It is now evident however, that members of the TIR family may use distinct intracellular signaling pathways to achieve a spectrum of biological end points. Using human embryonic kidney 293 cells transfected to express different TIRs we now report that signals initiated through IL-1R1 or TLR4 but not TLR3 can promote the stabilization of unstable chemokine mRNAs. Similar results were obtained when signaling from endogenous receptors was examined using a mouse endothelial cell line (H5V). The ability of TIR family members to stabilize ARE-containing mRNAs results from their differential use of signaling adaptors MyD88, MyD88 adaptor-like protein, Toll receptor IFN-inducing factor (Trif), and Trif-related adaptor molecule. Overexpression of MyD88 or MyD88 adaptor-like protein was able to promote enhanced stability of ARE containing mRNA, whereas Trif and Trif-related adaptor molecule exhibited markedly reduced capacity. Hence the ability of TIRs to signal stabilization of mRNA appears to be linked to the MyD88-dependent signaling pathway. PMID- 15294995 TI - Specific modulation of astrocyte inflammation by inhibition of mixed lineage kinases with CEP-1347. AB - Inflammatory conversion of murine astrocytes correlates with the activation of various MAPK, and inhibition of terminal MAPKs like JNK or p38 dampens the inflammatory reaction. Mixed lineage kinases (MLKs), a family of MAPK kinase kinases, may therefore be involved in astrocyte inflammation. In this study, we explored the effect of the MLK inhibitors CEP-1347 and CEP-11004 on the activation of murine astrocytes by either TNF plus IL-1 or by a complete cytokine mix containing additional IFN-gamma. The compounds blocked NO-, PG-, and IL-6 release with a median inhibitory concentration of approximately 100 nM. This activity correlated with a block of the JNK and the p38 pathways activated in complete cytokine mix-treated astrocytes. Although CEP-1347 did not affect the activation of NF-kappaB, it blocked the expression of cyclooxygenase-2 and inducible NO synthase at the transcriptional level. Quantitative transcript profiling of 17 inflammation-linked genes revealed a specific modulation pattern of astrocyte activation by MLK inhibition, for instance, characterized by up regulation of the anti-stress factors inhibitor of apoptosis protein-2 and activated transcription factor 4, no effect on manganese superoxide dismutase and caspase-11, and down-regulation of major inflammatory players like TNF, GM-CSF, urokinase-type plasminogen activator, and IL-6. In conclusion, MLK inhibitors like CEP-1347 are highly potent astrocyte immune modulators with a novel spectrum of activity. PMID- 15294996 TI - IL-9-mediated induction of eotaxin1/CCL11 in human airway smooth muscle cells. AB - Recent work has shown the potential importance of IL-9 in allergic diseases. The development of transgenic mice overexpressing IL-9 has suggested a key role for this cytokine in the development of the asthmatic phenotype including airway eosinophilia. In this study, we evaluated the expression of the IL-9R and the effects of IL-9 on human ASM cells by examining the release of Th2-associated chemokines (eotaxin1/CCL11 and thymus- and activation-regulated chemokine (TARC)/CCL17). IL-9R alpha-chain mRNA and surface expression were detected in cultured human airway smooth muscle (ASM) cells. In addition, primary cultured ASM cells, as well as bronchial smooth muscle cells within biopsies of asthmatics and not control subjects, revealed IL-9R protein expression. IL-9 stimulation of human ASM cells resulted in release of eotaxin1/CCL11, but had no effect on the release of TARC/CCL17, in time- and dose-dependent manner. Moreover, in vitro chemotaxis assay demonstrated that conditioned medium from IL-9-stimulated ASM cells attracted human eosinophils. Neutralizing Abs to IL-9, but not to IL-4 or IL-13, reduced significantly IL-9-induced production of eotaxin1/CCL11 from ASM cells. Interestingly, real-time RT-PCR showed that IL-9 up-regulated eotaxin1/CCL11 mRNA expression, but had no effect on TARC/CCL17. Treatment with Act D abrogates IL-9-induced eotaxin1/CCL11 mRNA and protein release by ASM cells. Finally, transfection study using eotaxin1/CCL11 promoter luciferase construct confirmed that IL-9 induced eotaxin1/CCL11 at the transcriptional level. Taken together, these data provide new evidence demonstrating that IL-9 dependent activation of ASM cells contributes to eosinophilic inflammation observed in asthma. PMID- 15294997 TI - Mature human Langerhans cells derived from CD34+ hematopoietic progenitors stimulate greater cytolytic T lymphocyte activity in the absence of bioactive IL 12p70, by either single peptide presentation or cross-priming, than do dermal interstitial or monocyte-derived dendritic cells. AB - The emerging heterogeneity of dendritic cells (DCs) mirrors their increasingly recognized division of labor at myriad control points in innate and acquired cellular immunity. We separately generated blood monocyte-derived DCs (moDCs), as well as Langerhans cells (LCs) and dermal-interstitial DCs (DDC-IDCs) from CD34(+) hematopoietic progenitor cells. Differential expression of CD11b, CD52, CD91, and the CD1 isoforms proved useful in distinguishing these three DC types. All mature DCs uniformly expressed comparable levels of HLA-DR, CD83, CD80, and CD86, and were potent stimulators of allogeneic T cells after exposure either to recombinant human CD40L trimer or a combination of inflammatory cytokines with PGE(2). moDCs, however, required 0.5-1 log greater numbers than LCs or DDC-IDCs to stimulate comparable T cell proliferation. Only moDCs secreted the bioactive heterodimer IL-12p70, and moDCs phagocytosed significantly more dying tumor cells than did either LCs or DDC-IDCs. LCs nevertheless proved superior to moDCs and DDC-IDCs in stimulating CTL against a recall viral Ag by presenting passively loaded peptide or against tumor Ag by cross-priming autologous CD8(+) T cells. LCs also secreted significantly more IL-15 than did either moDCs or DDC-IDCs, which is especially important to the generation of CTL. These findings merit further comparisons in clinical trials designed to determine the physiologic relevance of these distinctions in activity between LCs and other DCs. PMID- 15294998 TI - MHC class II isotype- and allele-specific attenuation of experimental autoimmune encephalomyelitis. AB - Most autoimmune diseases are associated with certain MHC class II haplotypes. Autoantigen-based specific immune therapy can lead either to beneficial or, in the context of inflammatory conditions, detrimental outcomes. Therefore, we designed a platform of peptides by combinatorial chemistry selected in a nonbiased Ag-independent approach for strong binding to the rat MHC class II isotype RT1.D(n) allelic product of the RT1(n) haplotype that is presenting autoantigen in myelin oligodendrocyte glycoprotein-induced experimental autoimmune encephalomyelitis in LEW.1N rats. Peptide p17 (Ac-FWFLDNAPL-NH(2)) was capable of suppressing the induction of and also ameliorated established experimental autoimmune encephalomyelitis. MHC class II isotype and allele specificity of the therapeutic principle were demonstrated in myelin basic protein-induced experimental autoimmune encephalomyelitis in LEW rats bearing the RT1(l) haplotype. A general immunosuppressive effect of the treatment was excluded by allogeneic heart transplantation studies. In vitro studies demonstrated the blocking effect of p17 on autoantigenic T cell responses. We thus demonstrate a rational design of strong MHC class II-binding peptides with absolute isotype and allele specificity able to compete for autoantigenic sequences presented on disease-associated MHC class II molecules. PMID- 15294999 TI - Complete complement components C4A and C4B deficiencies in human kidney diseases and systemic lupus erythematosus. AB - Although a heterozygous deficiency of either complement component C4A or C4B is common, and each has a frequency of approximately 20% in a Caucasian population, complete deficiencies of both C4A and C4B proteins are extremely rare. In this paper the clinical courses for seven complete C4 deficiency patients are described in detail, and the molecular defects for complete C4 deficiencies are elucidated. Three patients with homozygous HLA A24 Cw7 B38 DR13 had systemic lupus erythematosus, mesangial glomerulonephritis, and severe skin lesions or membranous nephropathy. Immunofixation, genomic restriction fragment length polymorphisms, and pulsed field gel electrophoresis experiments revealed the presence of monomodular RP-C4-CYP21-TNX (RCCX) modules, each containing a solitary, long C4A mutant gene. Sequencing of the mutant C4A genes revealed a 2 bp, GT deletion in exon 13 that leads to protein truncation. The other four patients with homozygous HLA A30 B18 DR7 had SLE, severe kidney disorders including mesangial or membranoproliferative glomerulonephritis, and/or Henoch Schoenlein purpura. Molecular genetic analyses revealed an unusual RCCX structure with two short C4B mutant genes, each followed by an intact gene for steroid 21 hydroxylase. Nine identical, intronic mutations were found in each mutant C4B. In particular, the 8127 g-->a mutation present at the donor site of intron 28 may cause an RNA splice defect. Analyses of 12 complete C4 deficiency patients revealed two hot spots of deleterious mutations: one is located at exon 13, the others within a 2.6-kb genomic region spanning exons 20-29. Screening of these mutations may facilitate epidemiologic studies of C4 in infectious, autoimmune, and kidney diseases. PMID- 15295000 TI - Characterization and recruitment of plasmacytoid dendritic cells in synovial fluid and tissue of patients with chronic inflammatory arthritis. AB - Dendritic cells (DCs) are thought to play a key role in driving the immunopathogenic response underlying chronic inflammatory arthritis. In this study, we have examined the presence and phenotype of plasmacytoid DCs (pDCs) in the synovial fluids (SF) of patients with rheumatoid arthritis (RA), psoriatic arthritis (PA), and osteoarthritis (OA) and determined the chemotactic properties of SF from these patients toward pDCs. Flow cytometry analysis showed that the percentage of pDCs, identified as a population of Lin(-)CD123(++) cells, is 4- to 5-fold higher in RA SF and PA SF than in OA SF. The morphological and immunophenotypic characterization of pDCs isolated from PA and RA SF indicates that they are in an immature state, most likely due to inhibitory factors present in RA SF, but are still able to undergo maturation when exposed ex vivo to viral agent or unmethylated DNA. CD123(+) and BDCA2(+) pDCs were detected by immunohistochemistry in RA synovial tissue in which expression of the IFN-alpha inducible protein MxA was also found, suggesting production of type I IFN by maturing pDCs. We also show that CXCR3 and CXCR4 are expressed by both blood derived pDCs and pDCs isolated from RA and PA SF and that CXCL-10, CXCL-11, and CXCL-12 present in RA and PA SF stimulate chemotaxis of blood-derived pDCs. Altogether, these findings suggest that chemokine-driven recruitment of pDCs from the blood to the inflamed synovium could be important in the regulation of the immune response in chronic inflammatory arthritis. PMID- 15295001 TI - Ex vivo homeostatic proliferation of CD4+ T cells in rheumatoid arthritis is dysregulated and driven by membrane-anchored TNFalpha. AB - The systemic CD4(+) T cell compartment in patients with rheumatoid arthritis (RA) is characterized by TCR repertoire contraction, shortened telomere lengths, and decreased numbers of recent thymic emigrants, suggesting a disturbed CD4(+) T cell homeostasis. In mice, homeostatic proliferation of peripheral CD4(+) T cells is regulated by TCR interaction with self peptide-MHC complexes (pMHC) and can be reproduced in vitro. We have established an ex vivo model of homeostatic proliferation, in which self-replication of human CD4(+) T cells is induced by cell-cell contact with autologous monocytes. In healthy individuals, blockade of TCR-pMHC class II contact resulted in decreased CD4(+) T cell division. In contrast, homeostatic proliferation in RA patients was not inhibited by pMHC blockade, but increased during the initial culture period. The anti-TNF-alpha Ab cA2 inhibited homeostasis-driven ex vivo proliferation in healthy controls and in RA patients. In addition, treatment of RA patients with infliximab decreased the ex vivo rate of homeostatic proliferation of CD4(+) T cells. Our results suggest a disturbed regulation of CD4(+) T cell homeostasis leading to the repertoire aberrations reported in RA. Membrane-anchored TNF-alpha appears to be a cell-cell contact-dependent stimulus of homeostatic proliferation of CD4(+) T cells, possibly favoring self-replication of autoreactive CD4(+) T cells in patients with RA. PMID- 15295002 TI - Molecular recognition patterns of serum anti-DNA topoisomerase I antibody in systemic sclerosis. AB - Autoreactive anti-DNA topoisomerase I (anti-Topo I) Abs are commonly detected in sera of systemic sclerosis (SSc) patients. Our studies have established a positive correlation between the levels of serum anti-Topo I Abs and both disease severity and activity of SSc. The molecular targets of anti-Topo I Ab on Topo I domains remain to be further defined. In this report, we studied the molecular recognition pattern of serum anti-Topo I Ab in 52 SSc patients. The highest reactivity of serum anti-Topo I Abs was against the core subdomains I and II (aa 207-441) and, to a lesser extent, against the core subdomain III (aa 433-636) of Topo I. The linker domain (aa 636-712) and the C-terminal domain (aa 713-765) had much less reactivity than the core domain (aa 207-636). Strikingly, very little reactivity was directed against the N-terminal domain (aa 1-213) by serum anti Topo I Ab. This molecular recognition pattern was consistent among all SSc serum samples studied. Results from patients with serial serum samples indicated that this pattern remained unchanged over time. Interestingly, some naive B cells from healthy controls, upon transformation by EBV, produced IgM Abs against Topo I. These Abs had low affinity for Topo I and reacted equally to all domains of Topo I. The molecular recognition pattern of serum anti-Topo I Ab in SSc suggests the presence of a unique antigenic stimulation in vivo in this disease. PMID- 15295003 TI - Epitope-dependent inhibition of T cell activation by the Ea transgene: an explanation for transgene-mediated protection from murine lupus. AB - A high level expression of the Ea(d) transgene encoding the I-E alpha-chain is highly effective in the suppression of lupus autoantibody production in mice. To explore the possible modulation of the Ag-presenting capacity of B cells as a result of the transgene expression, we assessed the ability of the transgenic B cells to activate Ag-specific T cells in vitro. By using four different model Ag MHC class II combinations, this analysis revealed that a high transgene expression in B cells markedly inhibits the activation of T cells in an epitope dependent manner, without modulation of the I-E expression. The transgene mediated suppression of T cell responses is likely to be related to the relative affinity of peptides derived from transgenic I-E alpha-chains (Ealpha peptides) vs antigenic peptides to individual class II molecules. Our results support a model of autoimmunity prevention based on competition for Ag presentation, in which the generation of large amounts of Ealpha peptides with high affinity to I A molecules decreases the use of I-A for presentation of pathogenic self-peptides by B cells, thereby preventing excessive activation of autoreactive T and B cells. PMID- 15295004 TI - Characterization of rat CD8+ uveitogenic T cells specific for interphotoreceptor retinal-binding protein 1177-1191. AB - The uveitogenic T cells that mediate experimental autoimmune uveitis are commonly assumed to be exclusively CD4(+). In the present study, we showed that, although a panel of long-term cultured rat uveitogenic T cell lines specific for the interphotoreceptor retinal-binding protein peptide, R16, all expressed CD4, approximately 40% of the R16-specific uveitogenic T cells freshly prepared from Ag-immunized rats were CD8(+)alphabetaTCR(+), as demonstrated by CFSE staining. We showed that the expansion of these CD8(+)alphabetaTCR(+) T cells was Ag specific and that highly purified CD8(+) R16-specific T cells were able to induce uveitis on transfusion into naive rats. Moreover, CD8(+) uveitogenic T cells more readily switched phenotype from, and to, TCR(-)CD8(-)CD4(-) during in vivo or in vitro activation compared with their CD4(+) counterparts. In a previous study, we showed that highly purified CD8(+) myelin oligodendrocyte glycoprotein-specific T cells induced more severe autoimmune encephalomyelitis than the corresponding CD4(+) T cells. In this study, we show that an interphotoreceptor retinal-binding protein peptide consistently activated a high proportion of CD8(+)alphabetaTCR(+) T cells, which were uveitogenic in Lewis rats. PMID- 15295005 TI - Leukemia-derived immature dendritic cells differentiate into functionally competent mature dendritic cells that efficiently stimulate T cell responses. AB - Primary acute myeloid leukemia cells can be induced to differentiate into dendritic cells (DC). In the presence of GM-CSF, TNF-alpha, and/or IL-4, leukemia derived DC are obtained that display features of immature DC (i-DC). The aim of this study was to determine whether i-DC of leukemic origin could be further differentiated into mature DC (m-DC) and to evaluate the possibility that leukemic m-DC could be effective in vivo as a tumor vaccine. Using CD40L as maturating agent, we show that leukemic i-DC can differentiate into cells that fulfill the phenotypic criteria of m-DC and, compared with normal counterparts, are functionally competent in vitro in terms of: 1) production of cytokines that support T cell activation and proliferation and drive Th1 polarization; 2) generation of autologous CD8(+) CTLs and CD4(+) T cells that are MHC-restricted and leukemia-specific; 3) migration from tissues to lymph nodes; 4) amplification of Ag presentation by monocyte attraction; 5) attraction of naive/resting and activated T cells. Irradiation of leukemic i-DC after CD40L stimulation did not affect their differentiating and functional capacity. Our data indicate that acute myeloid leukemia cells can fully differentiate into functionally competent m-DC and lay the ground for testing their efficacy as a tumor vaccine. PMID- 15295006 TI - Targeted CTLA-4 engagement induces CD4+CD25+CTLA-4high T regulatory cells with target (allo)antigen specificity. AB - CTLA-4 (CD152) is actively involved in down-regulating T cell activation and maintaining lymphocyte homeostasis. Our earlier studies showed that targeted engagement of CTLA-4 can down-modulate T cell response and suppress allo- and autoimmune responses. In this study, we report that targeted CTLA-4 engagement can induce immune tolerance to a specific target through selective induction of an Ag-specific CD4(+)CD25(+)CTLA-4(high) regulatory T cell (Treg cell) population. Allogeneic cells coated with anti-CTLA-4 Ab induced immune hyporesponsiveness through suppression of proinflammatory cytokines IFN-gamma and IL-2, and up-regulation of the regulatory cytokines IL-10, TGF-beta1, and IL-4, presumably through the engagement of CTLA-4 on activated T cells. Although rechallenge with alloantigen failed to break the unresponsiveness, a transient recovery from tolerance was observed in the presence of high concentrations of exogenous IL-2, saturating concentrations of neutralizing anti-TGF-beta1 and anti IL-10 Abs, and blocking anti-CTLA-4 Ab, and upon depletion of CD4(+)CD25(+) Treg cells. The CD4(+)CD25(+)CTLA-4(high) Treg cells from tolerant mice suppressed the effector function of CD25(-) T cells from Ag-primed mice. Adoptive transfer of these Treg cells into Ag-primed mice resulted in a significantly reduced alloantigen-specific response. Further characterization demonstrated that the Treg cells with memory phenotype (CD62L(-)) were more potent in suppressing the alloantigen-specific T cell response. These results strongly support that the targeted engagement of CTLA-4 has therapeutic potential for the prevention of transplant rejection. PMID- 15295007 TI - Opioid-like actions of neuropeptide Y in rat substantia gelatinosa: Y1 suppression of inhibition and Y2 suppression of excitation. AB - Neuropathic pain that results from injury to the peripheral or CNS responds poorly to opioid analgesics. Y1 and Y2 receptors for neuropeptide Y (NPY) may, however, serve as targets for analgesics that retain their effectiveness in neuropathic pain states. In substantia gelatinosa neurons in spinal cord slices from adult rats, we find that NPY acts via presynaptic Y2 receptors to attenuate excitatory postsynaptic currents (EPSCs) and predominantly on presynaptic Y1 receptors to attenuate glycinergic and GABAergic inhibitory postsynaptic currents (IPSCs). Because NPY attenuates the frequency of TTX-resistant miniature EPSCs and IPSCs, perturbation of the neurotransmitter release process contributes to its actions at both excitatory and inhibitory synapses. These effects, which are reminiscent of those produced by analgesic opioids, provide a cellular basis for previously documented spinal analgesic actions mediated via Y1 and Y2 receptors in neuropathic pain paradigms. They also underline the importance of suppression of inhibition in spinal analgesic mechanisms. PMID- 15295008 TI - Neuronal activity in macaque SEF and ACC during performance of tasks involving conflict. AB - It has been suggested on the basis of previous studies involving functional MRI (fMRI) and single-neuron recording that neurons of the supplementary eye field (SEF) and anterior cingulate cortex (ACC) monitor conflict. To test this idea, we carried out microelectrode recording in monkeys performing a color-conditional eye movement task in which red and green cues instructed leftward and rightward saccades, respectively. In a variant inducing conflict by spatial incompatibility, the cue was presented either at the location of the target (no conflict) or opposite the location of the target (conflict). In a variant inducing conflict by reversal, the foveal cue either remained one color (no conflict) or reversed color after 100 ms (conflict), with the monkey required to follow the instruction conveyed by the second color. In both tasks, conflict was evident in behavioral measures (reduced percent correct and slowed reaction time) and in physiological measures (reduced strength of directional activity among direction-selective neurons). In the SEF, there was a tendency for neurons to fire more strongly on trials involving conflict, but this effect took the form of modulation of task-related activity among direction-selective neurons, not of a pure conflict-monitoring signal. In the ACC, there was no conflict-related enhancement. These results are incompatible with the idea that the SEF and ACC contain populations of neurons specialized for monitoring conflict. PMID- 15295009 TI - Very fast oscillations evoked by median nerve stimulation in the human thalamus and subthalamic nucleus. AB - Very fast oscillations (VFOs; 500-1,500 Hz) are associated with sensory-evoked potentials (SEPs), but their origin is unknown. To characterize the origins of VFOs, we studied 35 patients with deep brain stimulation (DBS) electrodes [15 with thalamic and 20 with the subthalamic nucleus (STN) electrodes]. We recorded median nerve stimulation-evoked SEPs from the thalamus and STN with microelectrodes during stereotactic surgery and from the contacts of the DBS electrodes postoperatively. We also examined the firing of individual neurons in thalamus in relation to the VFOs. In the thalamus, VFOs with frequencies around 1,000 Hz were superimposed on slow potentials. Both slow and fast SEP components showed phase reversals in the somatosensory thalamus [ventralis caudalis (Vc)]. Median nerve poststimulus time histograms showed that single thalamic neurons fired at preferred times at intervals between 0.8 to 1.2 ms that were synchronous with the VFOs, although the neurons fired only once or a few times per trial. In the STN, low-amplitude SEPs with VFOs were observed at a latency similar to the thalamic SEPs. The VFOs from STN probably represent volume conduction, possibly from the medial lemniscus. We conclude that the thalamic VFOs are generated within Vc and that they induce time-locked firing in a network of neurons. PMID- 15295010 TI - Control of dynamic stability during gait termination on a slippery surface. AB - There are three common ways by which to successfully terminate gait: decreased acceleration of whole-body center of mass (COM) through a flexor synergy in the trail leg, increased deceleration of whole-body COM through an extensor synergy in the front limb, and an energy/momentum transfer to dissipate any remaining momentum if the first two strategies are unsuccessful. Healthy individuals were asked to stop on a slippery surface while we examined their unexpected response to the slippery surface. Kinetic data from the forceplates revealed lower braking forces in the slip trials compared with normal gait-termination trials. Subjects were unable to control their center of pressure (COP) to manipulate the COM as revealed by increased deviations and maximum absolute ranges of COP movement. Subject COM deviated farther in both horizontal planes and lowered further during the slip compared with normal gait-termination trials. Arm movements were effective in dissipating forward COM movement. In addition, there likely was a transfer of forward to lateral momentum to stop forward progression. All recorded muscle activity in the lower limbs and back increased during the slip to provide support to the lower limbs and correct upright balance. The trailing limb shortened its final step to provide support to the lowering COM. The balance correction response seen here resembles previous reactions to perturbations during locomotion suggesting there is a generalized strategy employed by the nervous system to correct for disturbances and maintain balance. PMID- 15295011 TI - Developmental changes in release properties of the CA3-CA1 glutamate synapse in rat hippocampus. AB - Developmental changes in release probability (Pr) and paired-pulse plasticity at CA3-CA1 glutamate synapses in hippocampal slices of neonatal rats were examined using field excitatory postsynaptic potential (EPSP) recordings. Paired-pulse facilitation (PPF) at these synapses was, on average, absent in the first postnatal week but emerged and became successively larger during the second postnatal week. This developmental increase in PPF was associated with a reduction in Pr, as indicated by the slower progressive block of the N-methyl-D aspartate (NMDA) EPSP by the noncompetitive NMDA receptor antagonist MK-801. This developmental reduction in Pr was not homogenous among the synapses. As shown by the MK-801 analysis, the Pr heterogeneity observed among adult CA3-CA1 synapses is present already during the first postnatal week, and the developmental Pr reduction was found to be largely selective for synapses with higher Pr values, leaving Pr of the vast majority of the synapses essentially unaffected. A reduction in Pves, the release probability of the individual vesicle, possibly caused by reduction in Ca2+ influx, seems to explain the reduction in Pr. In vivo injection of tetanus toxin at the end of the first postnatal week did not prevent the increase in PPF, indicating that this developmental change in release is not critically dependent on normal neural activity during the second postnatal week. PMID- 15295012 TI - Selectivity for the human body in the fusiform gyrus. AB - Functional neuroimaging studies have revealed human brain regions, notably in the fusiform gyrus, that respond selectively to images of faces as opposed to other kinds of objects. Here we use fMRI to show that the mid-fusiform gyrus responds with nearly the same level of selectivity to images of human bodies without faces, relative to tools and scenes. In a group-average analysis (n = 22), the fusiform activations identified by contrasting faces versus tools and bodies versus tools are very similar. Analyses of within-subjects regions of interest, however, show that the peaks of the two activations occupy close but distinct locations. In a second experiment, we find that the body-selective fusiform region, but not the face-selective region, responds more to stick figure depictions of bodies than to scrambled controls. This result further distinguishes the two foci and confirms that the body-selective response generalizes to abstract image formats. These results challenge accounts of the mid-fusiform gyrus that focus solely on faces and suggest that this region contains multiple distinct category-selective neural representations. PMID- 15295013 TI - Deficits in movements of the wrist ipsilateral to a stroke in hemiparetic subjects. AB - We examined step-tracking movements of the wrist and associated EMG activity in seven patients (age range, 27-73 yr) and in seven normal subjects that were matched to patients in age, sex, and handedness. All patients exhibited a hemiparesis that resulted from a unilateral cerebrovascular accident (CVA) that included motor areas in the frontal lobe or their efferents. The lesion in three patients was in their dominant hemisphere. The patients were tested 1-48 mo following their CVA. They had great difficulty in performing or were unable to perform step-tracking movements with the contralesional wrist. In addition, the patients displayed striking deficits in wrist movements and muscle activity of the ipsilesional wrist. These movements were >50% slower than those of controls. The initial movement step routinely undershot the target and was only 63% as large as that of controls. The patients made wrist movements with marked directional errors requiring corrective responses. These errors were due largely to inappropriate temporal sequencing of muscle activity. The deficits in movement and muscle activity in the wrist ipsilesional to a CVA were marked, regardless of whether the lesion was in the dominant or nondominant hemisphere. These observations indicate that unilateral lesions can have significant bilateral effects on the generation and control of distal limb movements. PMID- 15295014 TI - Modulation of an integrated central pattern generator-effector system: dopaminergic regulation of cardiac activity in the blue crab Callinectes sapidus. AB - Theoretical studies have suggested that the output of a central pattern generator (CPG) must be matched to the properties of its peripheral effector system to ensure production of functional behavior. One way that such matching could be achieved is through coordinated central and peripheral modulation. In this study, morphological and physiological methods were used to examine the sources and actions of dopaminergic modulation in the cardiac system of the blue crab, Callinectes sapidus. Immunohistochemical localization of tyrosine hydroxylase (TH) revealed a prominent neuron in the commissural ganglion, the L-cell, that projected a large-diameter axon to the pericardial organ (PO) by an indirect and circuitous route. Within the PO, the L-cell axon gave rise to fine varicose fibers, suggesting that it releases dopamine in a neurohormonal fashion onto the heart musculature. In addition, one branch of the axon continued beyond the PO to the heart, where it innervated the anterior motor neurons and the posterior pacemaker region of the cardiac ganglion (CG). In physiological experiments, exogenous dopamine produced multiple effects on contraction and motor neuron burst parameters that corresponded to the dual central-peripheral modulation suggested by the L-cell morphology. Interestingly, parameters of the ganglionic motor output were modulated differently in the isolated CG and in a novel semi intact system where the CG remained embedded within the heart musculature. These observations suggest a critical role of feedback from the periphery to the CG and underscore the requirement for integration of peripheral (neurohormonal) actions and direct ganglionic modulation in the regulation of this exceptionally simple system. PMID- 15295015 TI - Neural correlates of the precedence effect in the inferior colliculus of behaving cats. AB - Several auditory spatial illusions, collectively called the precedence effect (PE), occur when transient sounds are presented from two different spatial locations but separated in time by an interstimulus delay (ISD). For ISDs in the range of localization dominance (<10 ms), a single fused sound is typically located near the leading source location only, as if the location of the lagging source were suppressed. For longer ISDs, both the leading and lagging sources can be heard and localized, and the shortest ISD where this occurs is called the echo threshold. Previous physiological studies of the extracellular responses of single neurons in the inferior colliculus (IC) of anesthetized cats and unanesthetized rabbits with sounds known to elicit the PE have shown correlates of these phenomena though there were differences in the physiologically measured echo thresholds. Here we recorded in the IC of awake, behaving cats using stimuli that we have shown to evoke behavioral responses that are consistent with the precedence effect. For small ISDs, responses to the lag were reduced or eliminated consistent with psychophysical data showing that sound localization is based on the leading source. At longer ISDs, the responses to the lagging source recovered at ISDs comparable to psychophysically measured echo thresholds. Thus it appears that anesthesia, and not species differences, accounts for the discrepancies in the earlier studies. PMID- 15295016 TI - Influence of postural anxiety on postural reactions to multi-directional surface rotations. AB - Previous studies have shown significant effects of increased postural anxiety in healthy young individuals when standing quietly or performing voluntary postural tasks. However, little is known about the influence of anxiety on reactive postural control. The present study examined how increased postural anxiety influenced postural reactions to unexpected surface rotations in multiple directions. Ten healthy young adults (mean age: 25.5 yr, range: 22-27 yr) were required to recover from unexpected rotations of the support surface (7.5 degrees amplitude, 50 degrees/s velocity) delivered in six different directions while standing in a low postural threat (surface height: 60 cm above ground) or high postural threat (surface height: 160 cm above ground) condition. Electromyographic data from 12 different postural leg, hip, and trunk muscles was collected simultaneously. Full body kinematic data were also used to determine total body center of mass (COM) and segment displacements. Four distinct changes were observed with increased postural anxiety: increased amplitude in balance correcting responses (120-220 ms) in all leg, trunk, and arm muscles; decreased onset latency of deltoid responses; reduced magnitude of COM displacement; and reduced angular displacement of leg, pelvis, and trunk. These observations suggest that changes in dynamic postural responses with increased anxiety are mediated by alterations in neuro-muscular control mechanisms and thus may contribute significantly to the pathophysiology of balance deficits associated with aging or neurological disease. PMID- 15295017 TI - Localization of organelle proteins by isotope tagging (LOPIT). AB - We describe a proteomics method for determining the subcellular localization of membrane proteins. Organelles are partially separated using centrifugation through self-generating density gradients. Proteins from each organelle co fractionate and therefore exhibit similar distributions in the gradient. Protein distributions can be determined through a series of pair-wise comparisons of gradient fractions, using cleavable ICAT to enable relative quantitation of protein levels by MS. The localization of novel proteins is determined using multivariate data analysis techniques to match their distributions to those of proteins that are known to reside in specific organelles. Using this approach, we have simultaneously demonstrated the localization of membrane proteins in both the endoplasmic reticulum and the Golgi apparatus in Arabidopsis. Localization of organelle proteins by isotope tagging is a new tool for high-throughput protein localization, which is applicable to a wide range of research areas such as the study of organelle function and protein trafficking. PMID- 15295018 TI - Changes in phospholipid content and myocardial calcium-independent phospholipase A2 activity during chronic anthracycline administration. AB - Despite numerous investigations, the causes underlying anthracycline cardiomyopathy are yet to be established. We have recently reported that acute treatment with anthracyclines inhibits membrane-associated calcium-independent phospholipase A(2) (iPLA(2)) activity both in vitro and in vivo. This study presents data that iPLA(2) activity is also suppressed during chronic drug administration. Adult Sprague-Dawley rats were given weekly 1 mg/kg i.v. injections of doxorubicin for a total of 8 weeks. One week after the last injection, the animals were sacrificed, and heart tissue was assessed for phospholipid content and iPLA(2) activity. Membrane-associated iPLA(2) activity in the myocardium of doxorubicin-treated animals was 40% lower than that in control hearts. In addition, doxorubicin treatment resulted in significant alterations in the distribution of fatty acyl moieties esterified to the sn-2 position of choline glycerophospholipids. The ethanolamine species remained unaffected. Elevation in the amount of arachidonate and linoleate esterified to the sn-2 position of choline plasmalogens was consistent with the hypothesis that iPLA(2) displays selectivity for plasmalogen phospholipids; therefore, enzyme inhibition may affect hydrolysis of these phospholipid subclasses. Notably, the changes in phospholipid content occurred at a low cumulative dose of 8 mg/kg at which appearance of structural lesions was minimal. Therefore, these alterations seem to be both specific and early signs of cardiomyocyte pathology. The results support our hypothesis that myocardial iPLA(2) inhibition may be one of the steps that leads to the functional and structural changes associated with chronic anthracycline treatment. PMID- 15295019 TI - Multiple sites of adaptive plasticity in the owl's auditory localization pathway. AB - In the midbrain auditory localization pathway of the barn owl, a map of auditory space is relayed from the external nucleus of the inferior colliculus (ICX) to the deep and intermediate layers of the optic tectum (OT) and from these layers to the superficial layers. Within the OT, the auditory space map aligns with a visual map of space. Raising young barn owls with a prismatic displacement of the visual field leads to progressive changes in auditory tuning in the OT that tend to realign the auditory space map with the prismatically displaced visual space map. The only known site of this adaptive plasticity is in the ICX, in which the auditory system first creates a map of space. In this study, we identified an additional site of plasticity in the OT. In owls that experienced prisms beginning late in the juvenile period, adaptive shifts in auditory tuning in the superficial layers of the OT exceeded the adaptive shifts that occurred in the deep layers of the OT or in the ICX. Anatomical results from these owls demonstrated that the topography of intrinsic OT connections was systematically altered in the adaptive direction. In juvenile owls, plasticity in the OT increased as plasticity in the ICX decreased. Because plasticity at both sites has been shown to decline substantially in adults, these results suggest that an age-dependent decrease in auditory map plasticity occurs first in the ICX and later at the higher level, in the OT. PMID- 15295020 TI - The sleep slow oscillation as a traveling wave. AB - During much of sleep, virtually all cortical neurons undergo a slow oscillation (<1 Hz) in membrane potential, cycling from a hyperpolarized state of silence to a depolarized state of intense firing. This slow oscillation is the fundamental cellular phenomenon that organizes other sleep rhythms such as spindles and slow waves. Using high-density electroencephalogram recordings in humans, we show here that each cycle of the slow oscillation is a traveling wave. Each wave originates at a definite site and travels over the scalp at an estimated speed of 1.2-7.0 m/sec. Waves originate more frequently in prefrontal-orbitofrontal regions and propagate in an anteroposterior direction. Their rate of occurrence increases progressively reaching almost once per second as sleep deepens. The pattern of origin and propagation of sleep slow oscillations is reproducible across nights and subjects and provides a blueprint of cortical excitability and connectivity. The orderly propagation of correlated activity along connected pathways may play a role in spike timing-dependent synaptic plasticity during sleep. PMID- 15295021 TI - Coordinating structural and functional synapse development: postsynaptic p21 activated kinase independently specifies glutamate receptor abundance and postsynaptic morphology. AB - Here, we show that postsynaptic p21-activated kinase (Pak) signaling diverges into two genetically separable pathways at the Drosophila neuromuscular junction. One pathway controls glutamate receptor abundance. Pak signaling within this pathway is specified by a required interaction with the adaptor protein Dreadlocks (Dock). We demonstrate that Dock is localized to the synapse via an Src homology 2-mediated protein interaction. Dock is not necessary for Pak localization but is necessary to restrict Pak signaling to control glutamate receptor abundance. A second genetically separable function of Pak kinase signaling controls muscle membrane specialization through the regulation of synaptic Discs-large. In this pathway, Dock is dispensable. We present a model in which divergent Pak signaling is able to coordinate two different features of postsynaptic maturation, receptor abundance, and muscle membrane specialization. PMID- 15295022 TI - Protein kinase C delta mediates cerebral reperfusion injury in vivo. AB - Protein kinase C (PKC) has been implicated in mediating ischemic and reperfusion damage in multiple organs. However, conflicting reports exist on the role of individual PKC isozymes in cerebral ischemic injury. Using a peptide inhibitor selective for deltaPKC, deltaV1-1, we found that deltaPKC inhibition reduced cellular injury in a rat hippocampal slice model of cerebral ischemia [oxygen glucose deprivation (OGD)] when present both during OGD and for the first 3 hr of reperfusion. We next demonstrated peptide delivery to the brain parenchyma after in vivo delivery by detecting biotin-conjugateddeltaV1-1 and by measuring inhibition of intracellular deltaPKC translocation, an indicator of deltaPKC activity. Delivery of deltaV1-1 decreased infarct size in an in vivo rat stroke model of transient middle cerebral artery occlusion. Importantly, deltaV1-1 had no effect when delivered immediately before ischemia. However, delivery at the onset, at 1 hr, or at 6 hr of reperfusion reduced injury by 68, 47, and 58%, respectively. Previous work has implicated deltaPKC in mediating apoptotic processes. We therefore determined whether deltaPKC inhibition altered apoptotic cell death or cell survival pathways in our models. We found that deltaV1-1 reduced numbers of terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick end labeling-positive cells, indicating decreased apoptosis, increased levels of phospho-Akt, a kinase involved in cell survival pathways, and inhibited BAD (Bcl-2-associated death protein) protein translocation from the cell cytosol to the membrane, indicating inhibition of proapoptotic signaling. These data support a deleterious role for deltaPKC during reperfusion and suggest that deltaV1-1 delivery, even hours after commencement of reperfusion, may provide a therapeutic advantage after cerebral ischemia. PMID- 15295023 TI - Altered prelimbic cortex output during cue-elicited drug seeking. AB - Cocaine treatment paired with environmental cues establishes a conditioned place preference (CPP) for that environment. After expression of this preference, rats show elevated levels of immediate early genes (IEGs; e.g. c-fos) in the prelimbic cortex (PrL), basolateral amygdala complex (BLC), and nucleus accumbens core (NAcc) compared with drug-unpaired controls. These findings, together with the known connections between these regions, suggest that they function as a circuit contributing to cue-elicited craving. To investigate the function of this circuit during drug-seeking, we characterized Fos immunoreactivity of particular neuron classes in each region. To distinguish between IEG activation of GABAergic and non-GABAergic (principally, excitatory projection) neurons, we combined Fos immunohistochemistry with immunohistochemistry for glutamic acid decarboxylase 67 (GAD67) or calcium/calmodulin-dependent protein kinase II (CAMKII) proteins. Within the BLC and NAcc of drug-paired and drug-unpaired animals tested for CPP, we observed no significant differences in the percentage of Fos-immunoreactive (IR) cells that were also GAD67-IR. We also observed no group difference in the degree of Fos/CAMKII overlap in the BLC. However, in PrL, the degree of Fos/GAD67 overlap in the drug-paired group was significantly higher than in the drug unpaired group. Also, the Fos/CAMKII overlap in the entire PrL as well as just its layer V was significantly lower in the drug-paired animals compared with controls. These findings suggest that, during CPP expression in cocaine-paired animals, the PrL GABAergic interneurons are preferentially activated while PrL output is attenuated, perhaps through greater inhibition of layer V pyramidal neurons. These results suggest a shifting prefrontal cortex cell population response during cocaine-seeking. PMID- 15295024 TI - Setting boundaries: brain dynamics of modal and amodal illusory shape completion in humans. AB - Normal visual perception requires differentiating foreground from background objects. Differences in physical attributes sometimes determine this relationship. Often such differences must instead be inferred, as when two objects or their parts have the same luminance. Modal completion refers to such perceptual "filling-in" of object borders that are accompanied by concurrent brightness enhancement, in turn termed illusory contours (ICs). Amodal completion is filling-in without concurrent brightness enhancement. Presently there are controversies regarding whether both completion processes use a common neural mechanism and whether perceptual filling-in is a bottom-up, feedforward process initiating at the lowest levels of the cortical visual pathway or commences at higher-tier regions. We previously examined modal completion (Murray et al., 2002) and provided evidence that the earliest modal IC sensitivity occurs within higher-tier object recognition areas of the lateral occipital complex (LOC). We further proposed that previous observations of IC sensitivity in lower-tier regions likely reflect feedback modulation from the LOC. The present study tested these proposals, examining the commonality between modal and amodal completion mechanisms with high-density electrical mapping, spatiotemporal topographic analyses, and the local autoregressive average distributed linear inverse source estimation. A common initial mechanism for both types of completion processes (140 msec) that manifested as a modulation in response strength within higher tier visual areas, including the LOC and parietal structures, is demonstrated, whereas differential mechanisms were evident only at a subsequent time period (240 msec), with amodal completion relying on continued strong responses in these structures. PMID- 15295025 TI - Synaptic strengthening mediated by bone morphogenetic protein-dependent retrograde signaling in the Drosophila CNS. AB - Retrograde signaling is an essential component of synaptic development and physiology. Previous studies show that bone morphogenetic protein (BMP)-dependent retrograde signaling is required for the proper development of the neuromuscular junction (NMJ) in Drosophila. These studies, moreover, raised the significant possibility that the development of central motor circuitry might similarly be reliant on such signaling. To test this hypothesis, retrograde signaling between postsynaptic motoneurons and their presynaptic interneurons is examined. Postsynaptic expression of an adenylate cyclase encoded by rutabaga (rut), is sufficient to strengthen synaptic transmission at these identified central synapses. Results are presented to show that the underlying mechanism is dependent on BMP retrograde signaling. Thus, presynaptic expression of an activated TGF-beta receptor, thickvien (tkv), or postsynaptic expression of a TGF beta ligand, glass-bottom boat (gbb), is sufficient to phenocopy strengthening of synaptic transmission. In the absence of gbb, endogenous synaptic transmission is significantly weakened and, moreover, postsynaptic overexpression of rut is unable to potentiate synaptic function. Potentiation of presynaptic neurotransmitter release, mediated by increased postsynaptic expression of gbb, is dependent on normal cholinergic activity, indicative that either the secretion of this retrograde signal, or its transduction, is activity dependent. Thus, in addition to the development of the NMJ and expression of myoactive FMRFamide-like peptides in specific central neurons, the results of the present study indicate that this retrograde signaling cascade also integrates the development and function of central motor circuitry that controls movement in Drosophila larvae. PMID- 15295026 TI - Opioid receptors in the midbrain periaqueductal gray regulate extinction of pavlovian fear conditioning. AB - Four experiments studied the role of opioid receptors in the midbrain periaqueductal gray matter (PAG), an important structure eliciting conditioned fear responses, in the extinction of Pavlovian fear. Rats received pairings of an auditory conditioned stimulus (CS) with a foot shock unconditioned stimulus (US). The freezing conditioned response (CR) elicited by the CS was then extinguished via nonreinforced presentations of the CS. Microinjection of the opioid receptor antagonist naloxone into the ventrolateral PAG (vlPAG) before nonrein-forced CS presentations impaired development of extinction, but such microinjections at the end of extinction did not reinstate an already extinguished freezing CR. This role for opioid receptors in fear extinction was specific to the vlPAG because infusions of naloxone into the dorsal PAG did not impair fear extinction. Finally, the impairment of fear extinction produced by vlPAG infusions of naloxone was dose-dependent. These results show for the first time that the midbrain PAG contributes to fear extinction and specifically identify a role for vlPAG opioid receptors in the acquisition but not the expression of such extinction. Taken together with our previous findings, we suggest that, during fear conditioning, activation of vlPAG opioid receptors contributes to detection of the discrepancy between the actual and expected outcome of the conditioning trial. vlPAG opioid receptors regulate the learning that accrues to the CS and other stimuli present on a trial because they instantiate an associative error correction process influencing US information reaching the site of CS-US convergence in the amygdala. During nonreinforcement, this vlPAG opioid receptor contribution signals extinction. PMID- 15295027 TI - Glutamate released from glial cells synchronizes neuronal activity in the hippocampus. AB - Glial cells of the nervous system directly influence neuronal and synaptic activities by releasing transmitters. However, the physiological consequences of this glial transmitter release on brain information processing remain poorly understood. We demonstrate here in hippocampal slices of 2- to 5-week-old rats that glutamate released from glial cells generates slow transient currents (STCs) mediated by the activation of NMDA receptors in pyramidal cells. STCs persist in the absence of neuronal and synaptic activity, indicating a nonsynaptic origin of the source of glutamate. Indeed, STCs occur spontaneously but can also be induced by pharmacological tools known to activate astrocytes and by the selective mechanical stimulation of single nearby glial cells. Bath application of the inhibitor of the glutamate uptake dl-threo-beta-benzyloxyaspartate increases both the frequency of STCs and the amplitude of a tonic conductance mediated by NMDA receptors and probably also originated from glial glutamate release. By using dual recordings, we observed synchronized STCs in pyramidal cells having their soma distant by <100 microm. The degree of precision (<100 msec) of this synchronization rules out the involvement of calcium waves spreading through the glial network. It also indicates that single glial cells release glutamate onto adjacent neuronal processes, thereby controlling simultaneously the excitability of several neighboring pyramidal cells. In conclusion, our results show that the glial glutamate release occurs spontaneously and synchronizes the neuronal activity in the hippocampus. PMID- 15295028 TI - A morphological correlate of synaptic scaling in visual cortex. AB - We studied the response of dendritic spines in the thalamic-recipient zone of rat visual cortex to simple manipulations of the visual environment. We measured the morphologies of a total of 3824 spines located on the basal dendrites of 60 layer 3 pyramidal cells. As expected from previous studies, we found a significantly lower spine density in dark-reared animals at postnatal day 30 (P30) compared with light-reared controls. Additional analysis revealed that the spines in dark reared animals were significantly shorter and more bulbous than in light-reared animals. When these two results were combined, we found that the total synaptic area per unit length of dendrite was conserved, compatible with the phenomenon of "synaptic scaling." We also found that the increase in average spine head diameter is reversed by 10 d of light exposure (starting at P20), but surprisingly, the decrease in spine density is not. Thus, not all effects of dark rearing can be reversed by subsequent visual experience, even when the experience occurs during the third postnatal week. PMID- 15295029 TI - A single in vivo exposure to cocaine abolishes endocannabinoid-mediated long-term depression in the nucleus accumbens. AB - In the nucleus accumbens (NAc), a key structure to the effects of all addictive drugs, presynaptic cannabinoid CB1 receptors (CB1Rs) and postsynaptic metabotropic glutamate 5 receptors (mGluR5s) are the principal effectors of endocannabinoid (eCB)-mediated retrograde long-term depression (LTD) (eCB-LTD) at the prefrontal cortex-NAc synapses. Both CB1R and mGluR5 are involved in cocaine related behaviors; however, the impact of in vivo cocaine exposure on eCB mediated retrograde synaptic plasticity remains unknown. Electrophysiological and biochemical approaches were used, and we report that a single in vivo cocaine administration abolishes eCB-LTD. This effect of cocaine was not present in D1 dopamine receptor (D1R) -/- mice and was prevented when cocaine was coadministered with the selective D1R antagonist 8-chloro-2,3,4,5-tetrahydro-3-5 1h-3-benzazepin-7-ol (0.5 mg/kg) or with the NMDA receptor (NMDAR) blocker (+)-5 methyl-10,11-dihydro-5H-dibenzo [a,d] cyclohepten-5,10-imine maleate (1 mg/kg), suggesting the involvement of D1R and NMDAR. We found that the cocaine-induced blockade of retrograde signaling was correlated with enhanced expression levels of Homer scaffolding proteins containing the coiled-coil domain and accompanied by a strong reduction of mGluR5 surface expression. The results suggest that cocaine-induced loss of eCB retrograde signaling is caused by a reduction in the ability of mGluR5 to translate anterograde glutamate transmission into retrograde eCB signaling. PMID- 15295030 TI - Action potential-evoked and ryanodine-sensitive spontaneous Ca2+ transients at the presynaptic terminal of a developing CNS inhibitory synapse. AB - The existence of spontaneous calcium transients (SCaTs) dependent on intracellular store activation has been reported in putative axonal terminals of cerebellar basket interneurons. We used the two-photon imaging technique to optically identify basket terminals in acute cerebellar slices of young rats (11 16 d old) and study the properties of SCaTs unambiguously localized in these regions. The whole-cell recording configuration and preloading technique were alternatively used to load the calcium-dependent dye in the interneuron and compare SCaTs with action potential evoked calcium transients. SCaTs were observed in the basket terminals at frequencies that were significantly increased after bath application of 10 microm ryanodine and did not depend on P/Q- or N type voltage-dependent calcium channel activation. They originated at specific sites where bursts of events with temporal separation as small as 200 msec could be generated. Their sites of origin were spaced on average 6 microm apart and were preferentially located near axonal endings. SCaTs had amplitudes comparable with those of Ca2+ rises evoked by single action potentials that lead to release of neurotransmitter, as confirmed by parallel recordings of preloaded terminals and evoked IPSCs in the postsynaptic Purkinje cells. These results support the hypothesis that SCaTs at basket terminals underlie the large miniature IPSCs characteristic of Purkinje cells. PMID- 15295031 TI - Striatal neuron differentiation from neurosphere-expanded progenitors depends on Gsh2 expression. AB - Neural stem and progenitor cells from the embryonic forebrain can be expanded under growth factor stimulation in vitro, either as free-floating aggregates called neurospheres or as attached monolayer cultures. We have previously shown that despite the maintenance of important regulatory genes such as Gsh2, in vitro expansion of cells from the lateral ganglion eminence (LGE) restricts their differentiation potential. Specifically, their ability to differentiate into striatal projection neurons is compromised. It is not clear whether this restriction is caused by loss of progenitors with the ability to generate striatal projection neurons or whether the restricted differentiation potential is caused by factors lacking during in vitro differentiation. To address this, we have set up an in vitro system, in which expanded LGE-derived cells are differentiated in coculture with primary cells isolated from different regions of the embryonic brain. We provide evidence that the primary cells supply the expanded cells with contact-mediated region-specific developmental cues. Neurosphere-expanded LGE progenitors can, when presented with these cues, differentiate into neurons with characteristics of striatal projection neurons. Furthermore, we show that the ability of the expanded LGE cells to respond to the developmental cues presented by the primary cells depends on the maintained expression of Gsh2 in the expanded cells. PMID- 15295032 TI - Identification of ectodomain regions contributing to gating, deactivation, and resensitization of purinergic P2X receptors. AB - The P2X receptors (P2XRs) are a family of ligand-gated channels activated by extracellular ATP through a sequence of conformational transitions between closed, open, and desensitized states. In this study, we examined the dependence of the activity of P2XRs on ectodomain structure and agonist potency. Experiments were done in human embryonic kidney 293 cells expressing rat P2X2aR, P2X2bR, and P2X3R, and chimeras having the V60-R180 or V60-F301 ectodomain sequences of P2X3R instead of the I66-H192 or I66-Y310 sequences of P2X2aR and P2X2bR. Chimeric P2X2a/V60-F301X3R and P2X2b/V60-F301X3R inherited the P2X3R ligand-selective profile, whereas the potency of agonists for P2X2a/V60-R180X3R was in between those observed at parental receptors. Furthermore, P2X2a/V60-F301X3R and P2X2a/V60-R180X3R desensitized in a P2X2aR-specific manner, and P2X2b/V60-F301X3R desensitized with rates comparable with those of P2X2bR. In striking contrast to parental receptors, the rates of decay in P2X2a/V60-F301X3R and P2X2b/V60-F301X3R currents after agonist withdrawal were 15- to 200-fold slower. For these chimeras, the decays in currents were not dependent on duration of stimuli and reflected both continuous desensitization and deactivation of receptors. Also, participation of deactivation in closure of channels inversely correlated with potency of agonists to activate receptors. The delay in deactivation was practically abolished in P2X2a/V60-R180X3R-expressing cells. However, the recovery from desensitization of P2X2a/V60-F301X3R and P2X2a/V60-R180X3R was similar and substantially delayed compared with that of parental receptors. These results indicate that both ectodomain halves participate in gating, but that the C and N halves influence the stability of open and desensitized conformation states, respectively, which in turn reflects on rates of receptor deactivation and resensitization. PMID- 15295033 TI - Retrieving memories via internal context requires the hippocampus. AB - Episodic memory encodes the unique contexts of events so that people can remember the details of an experience when cued by only a subset of event features (Tulving, 1972). In humans, the hippocampus is crucial for this kind of memory (Scoville and Milner, 1957; Vargha-Khadem et al., 1997). The present study tested whether the hippocampus was required for nonspatial, context-dependent memory retrieval in rats that were trained in a constant external environment to approach different nonspatial goal objects depending on their current internal motivational state (hunger or thirst). The rats learned to reliably approach the correct goal and thus used internal context to guide associative memory retrieval. Both fornix transection and selective neurotoxic hippocampal lesions severely impaired memory performance, but cue and motivational discrimination, as well as stimulus-reward associations, were preserved. The findings suggest that the hippocampus is required for using internal contextual information for flexible associative memory retrieval. PMID- 15295034 TI - LIM genes parcellate the embryonic amygdala and regulate its development. AB - The mechanisms that regulate the development of the amygdaloid complex are as yet poorly understood. Here, we show that in the absence of the LIM-homeodomain (LIM HD) gene Lhx2, a particular amygdaloid nucleus, the nucleus of the lateral olfactory tract (nLOT), is selectively disrupted. LIM family members are well suited for multiple roles in the development of complex structures because they participate in regulatory interactions that permit a diversity of function. To investigate the possible role for other LIM-HD genes as well as LIM-only (Lmo) genes in the developing amygdala, we examined their expression in the embryo. We show that amygdaloid nuclei upregulate distinct patterns of LIM gene expression from embryonic stages. This supports the hypothesis that LIM genes may participate in the mechanisms that control the development of the amygdala. The disruption of the nLOT in the Lhx2 mutant is the first evidence of a role for LIM HD genes in the development of the amygdaloid complex. The combinatorial expression patterns of LIM genes suggest a comprehensive mechanism for patterning this structure. PMID- 15295035 TI - Natural stimulus statistics alter the receptive field structure of v1 neurons. AB - Studies of the primary visual cortex (V1) have produced models that account for neuronal responses to synthetic stimuli such as sinusoidal gratings. Little is known about how these models generalize to activity during natural vision. We recorded neural responses in area V1 of awake macaques to a stimulus with natural spatiotemporal statistics and to a dynamic grating sequence stimulus. We fit nonlinear receptive field models using each of these data sets and compared how well they predicted time-varying responses to a novel natural visual stimulus. On average, the model fit using the natural stimulus predicted natural visual responses more than twice as accurately as the model fit to the synthetic stimulus. The natural vision model produced better predictions in >75% of the neurons studied. This large difference in predictive power suggests that natural spatiotemporal stimulus statistics activate nonlinear response properties in a different manner than the grating stimulus. To characterize this modulation, we compared the temporal and spatial response properties of the model fits. During natural stimulation, temporal responses often showed a stronger late inhibitory component, indicating an effect of nonlinear temporal summation during natural vision. In addition, spatial tuning underwent complex shifts, primarily in the inhibitory, rather than excitatory, elements of the response profile. These differences in late and spatially tuned inhibition accounted fully for the difference in predictive power between the two models. Both the spatial and temporal statistics of the natural stimulus contributed to the modulatory effects. PMID- 15295036 TI - Persistent increase in olfactory type G-protein alpha subunit levels may underlie D1 receptor functional hypersensitivity in Parkinson disease. AB - Although L-dopa remains the most effective treatment of Parkinson disease, its long-term administration is hampered by the appearance of dyskinesia. Hypersensitivity of dopamine D1 receptors in the striatum has been suggested to contribute to the genesis of these delayed adverse effects. However, D1 receptor amounts are unchanged in Parkinson disease, suggesting alterations of downstream effectors. In rodents, striatal D1 receptors activate adenylyl cyclase through olfactory type G-protein alpha subunit (Galphaolf) and G-protein gamma 7 subunit (Ggamma7). We found that Galphaolf was enriched in human basal ganglia and was markedly diminished in the putamen of patients with Huntington disease, in relation with the degeneration of medium spiny neurons. In contrast, in the putamen of patients with Parkinson disease, Galphaolf and Ggamma7 levels were both significantly increased. In the rat, the degeneration of dopamine neurons augmented Galphaolf levels in the striatal neurons, specifically at the plasma membrane, an effect accounting for the increase of D1 response on cAMP production in dopamine-depleted striatum. In lesioned rats, Galphaolf levels were normalized by a 3 week treatment with l-dopa or a D1 agonist but not with aD2-D3 agonist, supporting a Galphaolf regulation by D1 receptor usage. In contrast, the increases of Galphaolf levels in patients were not affected by the duration of l dopa treatment but correlated with duration of disease. In conclusion, our results revealed in the parkinsonian putamen a prolonged elevation of Galphaolf levels that may lead to a persistent D1 receptor hypersensitivity and contribute to the genesis of long-term complications of L-dopa. PMID- 15295037 TI - Putting fear in its place: remapping of hippocampal place cells during fear conditioning. AB - We recorded hippocampal place cells in two spatial environments: a training environment in which rats underwent fear conditioning and a neutral control environment. Fear conditioning caused many place cells to alter (or remap) their preferred firing locations in the training environment, whereas most cells remained stable in the control environment. This finding indicates that aversive reinforcement can induce place cell remapping even when the environment itself remains unchanged. Furthermore, contextual fear conditioning caused significantly more remapping of place cells than auditory fear conditioning, suggesting that place cell remapping was related to the rat's learned fear of the environment. These results suggest that one possible function of place cell remapping may be to generate new spatial representations of a single environment, which could help the animal to discriminate among different motivational contexts within that environment. PMID- 15295038 TI - Surface targeting of the dopamine transporter involves discrete epitopes in the distal C terminus but does not require canonical PDZ domain interactions. AB - The human dopamine transporter (hDAT) contains a C-terminal type 2 PDZ (postsynaptic density 95/Discs large/zona occludens 1) domain-binding motif (LKV) known to interact with PDZ domain proteins such as PICK1 (protein interacting with C-kinase 1). As reported previously, we found that, after deletion of this motif, hDAT was retained in the endoplasmic reticulum (ER) of human embryonic kidney (HEK) 293 and Neuro2A cells, suggesting that PDZ domain interactions might be critical for hDAT targeting. Nonetheless, substitution of LKV with SLL, the type 1 PDZ-binding sequence from the beta2-adrenergic receptor, did not disrupt plasma membrane targeting. Moreover, the addition of an alanine to the hDAT C terminus (+Ala), resulting in an LKVA termination sequence, or substitution of LKV with alanines (3xAla_618-620) prevented neither plasma membrane targeting nor targeting into sprouting neurites of differentiated N2A cells. The inability of +Ala and 3xAla_618-620 to bind PDZ domains was confirmed by lack of colocalization with PICK1 in cotransfected HEK293 cells and by the inability of corresponding C-terminal fusion proteins to pull down purified PICK1. Thus, although residues in the hDAT C terminus are indispensable for proper targeting, PDZ domain interactions are not required. By progressive substitutions with beta2 adrenergic receptor sequence, and by triple-alanine substitutions in the hDAT C terminus, we examined the importance of epitopes preceding the LKV motif. Substitution of RHW(615-617) with alanines caused retention of the transporter in the ER despite preserved ability of this mutant to bind PICK1. We propose dual roles of the hDAT C terminus: a role independent of PDZ interactions for ER export and surface targeting, and a not fully clarified role involving PDZ interactions with proteins such as PICK1. PMID- 15295039 TI - A minimum structure of aminoglycosides that causes an initiation shift of trans translation. AB - Trans-translation is an unusual translation in which transfer-messenger RNA plays a dual function--as a tRNA and an mRNA--to relieve the stalled translation on the ribosome. It has been shown that paromomycin, a typical member of a 4,5 disubstituted class of aminoglycosides, causes a shift of the translation resuming point on the tmRNA by -1 during trans-translation. To address the molecular basis of this novel effect, we examined the effects of various aminoglycosides that can bind around the A site of the small subunit of the ribosome on trans-translation in vitro. Tobramycin and gentamicin, belonging to the 4,6-disubstituted class of aminoglycosides having rings I and II similar to those in the 4,5-disubstituted class, possess similar effects. Neamine, which has only rings I and II, a common structure shared by 4,5- and 4,6-disubstituted classes of aminoglycosides, was sufficient to cause an initiation shift of trans translation. In contrast, streptomycin or hygromycin B, lacking ring I, did not cause an initiation shift. The effect of each aminoglycoside on trans-translation coincides with that on conformational change in the A site of the small subunit of the ribosome revealed by recent structural studies: paromomycin, tobramycin and geneticin which is categorized into the gentamicin subclass, but not streptomycin and hygromycin B, flip out two conserved adenine bases at 1492 and 1493 from the A site helix. The pattern of initiation shifts by paromomycin fluctuates with variation of mutations introduced into a region upstream of the initiation point. PMID- 15295040 TI - Complex cis-elements determine an RNA editing site in pea mitochondria. AB - The cis-requirements for the first editing site in the atp9 mRNA from pea mitochondria were investigated in an in vitro RNA editing system. Template RNAs deleted 5' of -20 are edited correctly, but with decreased efficiency. Deletions between -20 and the edited nucleotide abolish editing activity. Substitution of the sequences 3' of the editing site has little effect, which suggests that the major determinants reside upstream. Stepwise mutated RNA sequences were used as templates or competitors that divide the cis-elements into several distinct regions. In the template RNAs, mutation of the sequence between -40 and -35 reduces the editing activity, while the region from -15 to -5 is essential for the editing reaction. In competition experiments the upstream region can be titrated, while the essential sequence near the editing site is largely resistant to excess competitor. This observation suggests that either one trans-factor attaches to these separate cis-regions with different affinities or two distinct trans-factors bind to these sequences, and one of which is present in limited amounts, whereas the other one is more abundant in the lysate. PMID- 15295041 TI - The antibiotic pipeline--challenges, costs, and values. PMID- 15295042 TI - Producing penicillin. PMID- 15295043 TI - Treatment of heart failure with spironolactone--trial and tribulations. PMID- 15295044 TI - TGF-beta signaling, tumor suppression, and acute lymphoblastic leukemia. PMID- 15295045 TI - Airway smooth muscle in asthma--not just more of the same. PMID- 15295046 TI - Gene-expression patterns in drug-resistant acute lymphoblastic leukemia cells and response to treatment. AB - BACKGROUND: Childhood acute lymphoblastic leukemia (ALL) is curable with chemotherapy in approximately 80 percent of patients. However, the cause of treatment failure in the remaining 20 percent of patients is largely unknown. METHODS: We tested leukemia cells from 173 children for sensitivity in vitro to prednisolone, vincristine, asparaginase, and daunorubicin. The cells were then subjected to an assessment of gene expression with the use of 14,500 probe sets to identify differentially expressed genes in drug-sensitive and drug-resistant ALL. Gene-expression patterns that differed according to sensitivity or resistance to the four drugs were compared with treatment outcome in the original 173 patients and an independent cohort of 98 children treated with the same drugs at another institution. RESULTS: We identified sets of differentially expressed genes in B-lineage ALL that were sensitive or resistant to prednisolone (33 genes), vincristine (40 genes), asparaginase (35 genes), or daunorubicin (20 genes). A combined gene-expression score of resistance to the four drugs, as compared with sensitivity to the four, was significantly and independently related to treatment outcome in a multivariate analysis (hazard ratio for relapse, 3.0; P=0.027). Results were confirmed in an independent population of patients treated with the same medications (hazard ratio for relapse, 11.85; P=0.019). Of the 124 genes identified, 121 have not previously been associated with resistance to the four drugs we tested. CONCLUSIONS: Differential expression of a relatively small number of genes is associated with drug resistance and treatment outcome in childhood ALL. PMID- 15295047 TI - Rates of hyperkalemia after publication of the Randomized Aldactone Evaluation Study. AB - BACKGROUND: The Randomized Aldactone Evaluation Study (RALES) demonstrated that spironolactone significantly improves outcomes in patients with severe heart failure. Use of angiotensin-converting-enzyme (ACE) inhibitors is also indicated in these patients. However, life-threatening hyperkalemia can occur when these drugs are used together. METHODS: We conducted a population-based time-series analysis to examine trends in the rate of spironolactone prescriptions and the rate of hospitalization for hyperkalemia in ambulatory patients before and after the publication of RALES. We linked prescription-claims data and hospital admission records for more than 1.3 million adults 66 years of age or older in Ontario, Canada, for the period from 1994 through 2001. RESULTS: Among patients treated with ACE inhibitors who had recently been hospitalized for heart failure, the spironolactone-prescription rate was 34 per 1000 patients in 1994, and it increased immediately after the publication of RALES, to 149 per 1000 patients by late 2001 (P<0.001). The rate of hospitalization for hyperkalemia rose from 2.4 per 1000 patients in 1994 to 11.0 per 1000 patients in 2001 (P<0.001), and the associated mortality rose from 0.3 per 1000 to 2.0 per 1000 patients (P<0.001). As compared with expected numbers of events, there were 560 (95 percent confidence interval, 285 to 754) additional hyperkalemia-related hospitalizations and 73 (95 percent confidence interval, 27 to 120) additional hospital deaths during 2001 among older patients with heart failure who were treated with ACE inhibitors in Ontario. Publication of RALES was not associated with significant decreases in the rates of readmission for heart failure or death from all causes. CONCLUSIONS: The publication of RALES was associated with abrupt increases in the rate of prescriptions for spironolactone and in hyperkalemia-associated morbidity and mortality. Closer laboratory monitoring and more judicious use of spironolactone may reduce the occurrence of this complication. PMID- 15295048 TI - Loss of Smad3 in acute T-cell lymphoblastic leukemia. AB - BACKGROUND: The receptors for transforming growth factor beta (TGF-beta) and their signaling intermediates make up an important tumor-suppressor pathway. The role of one of these intermediates--Smad3--in the pathogenesis of lymphoid neoplasia is unknown. METHODS: We measured Smad3 messenger RNA (mRNA) and protein in leukemia cells obtained at diagnosis from 19 children with acute leukemia, including 10 with T-cell acute lymphoblastic leukemia (ALL), 7 with pre-B-cell ALL, and 2 with acute nonlymphoblastic leukemia (ANLL). All nine exons of the SMAD3 gene (MADH3) were sequenced. Mice in which one or both alleles of Smad3 were inactivated were used to evaluate the role of Smad3 in the response of normal T cells to TGF-beta and in the susceptibility to spontaneous leukemogenesis in mice in which both alleles of the tumor suppressor p27Kip1 were deleted. RESULTS: Smad3 protein was absent in T-cell ALL but present in pre-B cell ALL and ANLL. No mutations were found in the MADH3 gene in T-cell ALL, and Smad3 mRNA was present in T-cell ALL and normal T cells at similar levels. In mice, the loss of one allele for Smad3 impairs the inhibitory effect of TGF-beta on the proliferation of normal T cells and works in tandem with the homozygous inactivation of p27Kip1 to promote T-cell leukemogenesis. CONCLUSIONS: Loss of Smad3 protein is a specific feature of pediatric T-cell ALL. A reduction in Smad3 expression and the loss of p27Kip1 work synergistically to promote T-cell leukemogenesis in mice. PMID- 15295049 TI - Dysfunctional interaction of C/EBPalpha and the glucocorticoid receptor in asthmatic bronchial smooth-muscle cells. AB - BACKGROUND: Increased proliferation of bronchial smooth-muscle cells may lead to increased muscle mass in the airways of patients with asthma. The antiproliferative effect of glucocorticoids in bronchial smooth-muscle cells in subjects without asthma is mediated by a complex of the glucocorticoid receptor and the CCAAT/enhancer binding protein alpha (C/EBPalpha). We examined the signaling pathway controlling the inhibitory effect of glucocorticoids on cell proliferation and interleukin-6 synthesis in bronchial smooth-muscle cells of subjects with asthma and those without asthma. METHODS: Lines of bronchial smooth muscle cells were established from cells from 20 subjects with asthma, 8 subjects with emphysema, and 26 control subjects. Cell proliferation was determined by means of cell counts and [3H]thymidine incorporation. Signal transduction was studied by means of an electrophoretic DNA mobility-shift assay, a supershift electrophoretic-mobility assay, immunoblotting, use of C/EBPalpha antisense oligonucleotides, and use of a human C/EBPalpha expression vector. Interleukin-6 release was determined by means of an enzyme-linked immunosorbent assay. RESULTS: Glucocorticoids activated the glucocorticoid receptor and inhibited serum-induced secretion of interleukin-6 in bronchial smooth-muscle cells from both subjects with asthma and those without asthma; however, glucocorticoids inhibited proliferation only in bronchial smooth-muscle cells from subjects without asthma. C/EBPalpha protein was detected by immunoblotting in all bronchial smooth-muscle cells from subjects without asthma but not in those with asthma, whereas the protein was expressed in lymphocytes from both groups of subjects. C/EBPalpha antisense oligonucleotides or the glucocorticoid-receptor inhibitor mifepristone reversed the antiproliferative effect of glucocorticoids in bronchial smooth muscle cells from subjects without asthma. When bronchial smooth-muscle cells from subjects with asthma were transiently transfected with an expression vector for human C/EBPalpha, two forms of the protein were expressed, and subsequent administration of glucocorticoids inhibited cell proliferation. CONCLUSIONS: We hypothesize that a cell-type-specific absence of C/EBPalpha is responsible for the enhanced proliferation of bronchial smooth-muscle cells derived from subjects with asthma and that it explains the failure of glucocorticoids to inhibit proliferation in vitro. PMID- 15295050 TI - Primary care physicians who treat blacks and whites. AB - BACKGROUND: In the United States, black patients generally receive lower-quality health care than white patients. Black patients may receive their care from a subgroup of physicians whose qualifications or resources are inferior to those of the physicians who treat white patients. METHODS: We performed a cross-sectional analysis of 150,391 visits by black Medicare beneficiaries and white Medicare beneficiaries 65 years of age or older for medical "evaluation and management" who were seen by 4355 primary care physicians who participated in a biannual telephone survey, the 2000-2001 Community Tracking Study Physician Survey. RESULTS: Most visits by black patients were with a small group of physicians (80 percent of visits were accounted for by 22 percent of physicians) who provided only a small percentage of care to white patients. In a comparison of visits by white patients and black patients, we found that the physicians whom the black patients visited were less likely to be board certified (77.4 percent) than were the physicians visited by the white patients (86.1 percent, P=0.02) and also more likely to report that they were unable to provide high-quality care to all their patients (27.8 percent vs. 19.3 percent, P=0.005). The physicians treating black patients also reported facing greater difficulties in obtaining access for their patients to high-quality subspecialists, high-quality diagnostic imaging, and nonemergency admission to the hospital. CONCLUSIONS: Black patients and white patients are to a large extent treated by different physicians. The physicians treating black patients may be less well trained clinically and may have less access to important clinical resources than physicians treating white patients. Further research should be conducted to address the extent to which these differences may be responsible for disparities in health care. PMID- 15295051 TI - Managing hyperkalemia caused by inhibitors of the renin-angiotensin-aldosterone system. PMID- 15295052 TI - Images in clinical medicine. "Pseudoinfarction" pattern due to hyperkalemia. PMID- 15295053 TI - Clinical problem-solving. A bitter tale. PMID- 15295054 TI - Childhood leukemia--new advances and challenges. PMID- 15295055 TI - Health care in America--still too separate, not yet equal. PMID- 15295056 TI - Can curcumin cure cystic fibrosis? PMID- 15295057 TI - Evidence of airborne transmission of SARS. PMID- 15295058 TI - Dying and decision making. PMID- 15295059 TI - Registries and informed consent. PMID- 15295060 TI - Treatment of photoaging. PMID- 15295061 TI - Long-term leptin-replacement therapy for lipoatrophic diabetes. PMID- 15295062 TI - Images in clinical medicine. Gastrothorax simulating acute tension pneumothorax. PMID- 15295064 TI - Actin control over microtubules suggested by DISTORTED2 encoding the Arabidopsis ARPC2 subunit homolog. AB - In Arabidopsis, based on the randomly misshapen phenotype of leaf epidermal trichomes, eight genes have been grouped into a 'DISTORTED' class. Three of the DIS genes, WURM, DISTORTED1 and CROOKED have been cloned recently and encode the ARP2, ARP3 and ARPC5 subunits respectively, of a conserved actin modulating ARP2/3 complex. Here we identify a fourth gene, DISTORTED2 as the Arabidopsis homolog of the ARPC2 subunit of the ARP2/3 complex. Like other mutants in the complex dis2 trichomes also display supernumerary, randomly localized cortical actin patches. In addition dis2 trichomes possess abnormally clustered endoplasmic microtubules near sites of actin aggregation. Since microtubules are strongly implicated in the establishment and maintenance of growth directionality in higher plants our observations of aberrant microtubule clustering in dis2 trichomes suggests a convincing explanation for the randomly distorted trichome phenotype in dis mutants. In addition, the close proximity of microtubule clusters to the arbitrarily dispersed cortical actin patches in the dis mutants provides fresh insights into cytoskeletal interactions leading us to suggest that in higher plants microtubule arrangements directed towards the establishment and maintenance of polar growth-directionality are guided by cortical actin behavior and organization. PMID- 15295065 TI - Water channel activity of radish plasma membrane aquaporins heterologously expressed in yeast and their modification by site-directed mutagenesis. AB - Plants contain a number of aquaporin isoforms. We developed a method for determining the water channel activity of individual isoforms of aquaporin. Six plasma membrane aquaporins (RsPIPs) and two vacuolar membrane aquaporins (RsTIPs) of radish (Raphanus sativus) were expressed heterologously in Saccharomyces cerevisiae BJ5458, which is deficient in endogenous functional aquaporin. Aquaporins were detected by immunoblot analysis with corresponding antibodies. Water permeability of membranes from yeast transformants was assayed by stopped flow spectrophotometry. The water channel activity of members of the RsPIP2 and RsTIP subfamilies was about 10 times and 5 times greater, respectively, than that of the control; however, RsPIP1s had little (RsPIP1-2 and RsPIP1-3) or no activity (RsPIP1-1). Site-directed mutation of several residues conserved in RsPIP1s or RsPIP2s markedly altered the water transport activity. Exchange of Ile244 of RsPIP1-3 with valine increased the activity to 250% of the wild type RsPIP1-3. On the other hand, exchange of Val235 of RsPIP2-2, which corresponds to RsPIP1-3 Ile244, with isoleucine caused a marked inactivation to 45% of the original RsPIP2-2. Mutation at possible phosphorylation sites at the N- and C terminal tails also altered the activity. These results suggest that these residues in the half-helix loop E and the tails are involved in the water transport and the functional regulation of RsPIP1 and RsPIP2. PMID- 15295066 TI - Four DEF-like MADS box genes displayed distinct floral morphogenetic roles in Phalaenopsis orchid. AB - The complex flower organization of orchids offers an opportunity to discover new variant genes and different levels of complexity in the morphogenesis of flowers. In this study, four B-class Phalaenopsis DEF-like MADS-box genes were identified and characterized, including PeMADS2, PeMADS3, PeMADS4 and PeMADS5. Differential expression profiles of these genes were detected in the floral organs of P. equestris, suggesting distinctive roles in the floral morphogenesis of orchids. Furthermore, expressions of these genes were varied to different extents in the peloric mutants with lip-like petals. Expression of PeMADS4 was in lips and columns of wild type, and it extended to the lip-like petals in the peloric mutant. Expression of PeMADS5 was mainly in petals and to a lesser extent in columns in the wild type, whereas it was completely eliminated in the peloric mutant. Disruption of the PeMADS5 promoter region of the peloric mutant was detected at nucleotide +312 relative to the upstream of translational start codon, suggesting that a DNA rearrangement has occurred in the peloric mutant. Genomic structure analysis of the PeMADS5 showed that the exon length was conserved in exons 1-6, similar to DEF-like genes of other plants. Collectively, this is the first report that four DEF-like MADS genes were identified in a single monocotyledonous species and that they may play distinctive morphogenetic roles in the floral development of an orchid. PMID- 15295067 TI - VOZ; isolation and characterization of novel vascular plant transcription factors with a one-zinc finger from Arabidopsis thaliana. AB - A 38-bp pollen-specific cis-acting region of the AVP1 gene is involved in the expression of the Arabidopsis thaliana V-PPase during pollen development. Here, we report the isolation and structural characterization of AtVOZ1 and AtVOZ2, novel transcription factors that bind to the 38-bp cis-acting region of A. thaliana V-PPase gene, AVP1. AtVOZ1 and AtVOZ2 show 53% amino acid sequence similarity. Homologs of AtVOZ1 and AtVOZ2 are found in various vascular plants as well as a moss, Physcomitrella patens. Promoter-beta-glucuronidase reporter analysis shows that AtVOZ1 is specifically expressed in the phloem tissue and AtVOZ2 is strongly expressed in the root. In vivo transient effector-reporter analysis in A. thaliana suspension-cultured cells demonstrates that AtVOZ1 and AtVOZ2 function as transcriptional activators in the Arabidopsis cell. Two conserved regions termed Domain-A and Domain-B were identified from an alignment of AtVOZ proteins and their homologs of O. sativa and P. patens. AtVOZ2 binds as a dimer to the specific palindromic sequence, GCGTNx7ACGC, with Domain-B, which is comprised of a functional novel zinc coordinating motif and a conserved basic region. Domain-B is shown to function as both the DNA-binding and the dimerization domains of AtVOZ2. From highly the conservative nature among all identified VOZ proteins, we conclude that Domain-B is responsible for the DNA binding and dimerization of all VOZ-family proteins and designate it as the VOZ domain. PMID- 15295068 TI - Capturing in vivo dynamics of the actin cytoskeleton stimulated by auxin or light. AB - We present here a transient expression system that allows the response of actin microfilaments to physiological stimuli (changes in auxin content, light) to be observed in single cells in vivo. Etiolated, intact rice seedlings are attached to glass slides, transfected biolistically with talin fused to yellow-fluorescent protein to visualize actin microfilaments, and either treated with auxin or irradiated. The talin marker labels distinct populations of actin that are differentially expressed depending on the physiological state of the coleoptile (active elongation versus ceased elongation). Whereas longitudinal transvacuolar bundles prevail in cells that have ceased to elongate, fine cortical strands are characteristic for elongating cells. The visualized actin structures remain dynamic and responsive to signals. Exogenous auxin triggers a loosening of the bundles and an extension of the cortical strands, whereas irradiation reorientates cortical strands into longitudinal arrays. These responses correspond in quality and timing to the signal responses inferred previously from fixed specimens and biochemical studies. In big advantage over those methods it is now possible to observe them directly at the single cell level. Thus, the rice coleoptile system can be used as a convenient model to study actin dynamics in vivo, in response to physiologically relevant stimuli. PMID- 15295069 TI - The accumulation of alpha-zein in transgenic tobacco endosperm is stabilized by co-expression of beta-zein. AB - The cysteine-poor alpha-zein is the major prolamin storage protein fraction in maize endosperm and is localized in the interior of protein bodies with delta zein, whereas the hydrophobic cysteine-rich beta- and gamma-zein are found on the exterior of the PB. In transgenic tobacco endosperm expressing zein genes, alpha zein was unstable unless co-expressed with gamma-zein. Here we showed that alpha zein was also stabilized by beta-zein. Small accretions of alpha- and beta-zeins, similar in appearance to maize protein bodies, were localized to the endoplasmic reticulum within tobacco endosperm cells. The zein proteins were also localized to protein storage vacuoles in a more dispersed pattern, suggesting that they were transported there after they were post-translationally sequestered into the ER. PMID- 15295070 TI - Deactivation of photosynthetic activities is triggered by loss of a small amount of water in a desiccation-tolerant cyanobacterium, Nostoc commune. AB - Changes in photosynthetic activities under hypertonic conditions were studied in a terrestrial, highly desiccation-tolerant cyanobacterium, Nostoc commune, and in some desiccation-sensitive cyanobacteria. The amounts of water sustained in the colony matrix outside the N. commune cells and the cellular solute concentration were estimated by measuring the water potential, and the solute concentration was supposed to correspond to around 0.22 M sorbitol. Incubation of the colonies in 0.8 M sorbitol solution inhibited the energy transfer from the phycobilisome (PBS) anchor to PSII core complexes. At higher sorbitol concentrations, light energy absorbed by PSI, PSII, and PBS was dissipated to heat. PSI and cyclic electron flow around PSI was also deactivated by hypertonic treatment. Fv/Fm and (Fm'-F)/Fm' values started to decrease at 0.6 and 0.3 M sorbitol and reached zero at 1.0 and 0.8 M, respectively. Decreases in these two fluorescence parameters corresponded to the decreases in PSII fluorescence (F695) and photosynthetic CO2 fixation, respectively. The intensity of delayed light emission started to decrease at 1.0 M sorbitol and became negligible at 4.0 M. Comparing these changes in N. commune with those in desiccation-sensitive species, we found that N. commune cells actively deactivates photosynthetic systems on sensing water loss. PMID- 15295071 TI - Biochemical characterization of cytosolic fructose-1,6-bisphosphatase from apple (Malus domestica) leaves. AB - Cytosolic fructose-1,6-bisphosphatase was purified to apparent homogeneity from the leaves of apple, a sorbitol synthesizing species. The enzyme was a homotetramer with a subunit mass of 37 kDa, and was highly specific for fructose 1,6-bisphosphate (F1,6BP) with a Km of 3.1 micro M and a Vmax of 48 units (mg protein)(-1). Either Mg2+ or Mn2+ was required for its activity with a Km of 0.59 mM and 62 micro M, respectively. Li+, Ca2+, Zn2+, Cu2+ and Hg2+ inhibited whereas Mn2+ enhanced the Mg2+ activated enzyme activity. Fructose 6-phosphate (F6P) was found to be a mixed type inhibitor with a Ki of 0.47 mM. Fructose 2,6 bisphosphate (F2,6BP) competitively inhibited the enzyme activity and changed the substrate saturation curve from hyperbolic to sigmoidal. AMP was a non competitive inhibitor for the enzyme. F6P interacted with F2,6BP and AMP in a synergistic way to inhibit the enzyme activity. Dihydroxyacetone phosphate slightly inhibited the enzyme activity in the presence or absence of F2,6BP. Sorbitol increased the susceptibility of the enzyme to the inhibition by high concentrations of F1,6BP. High concentrations of sorbitol in the reaction mixture led to a reduction in the enzyme activity. PMID- 15295072 TI - Thermal and chlorophyll-fluorescence imaging distinguish plant-pathogen interactions at an early stage. AB - Different biotic stresses yield specific symptoms, owing to their distinct influence on a plant's physiological status. To monitor early changes in a plant's physiological status upon pathogen attack, chlorophyll fluorescence imaging (Chl-FI) and thermography, which respectively visualize photosynthetic efficiency and transpiration, were carried out in parallel for two fundamentally different plant-pathogen interactions. These non-destructive imaging techniques were able to visualize infections at an early stage, before damage appeared. Under growth-room conditions, a robotized set-up captured time series of visual, thermal and chlorophyll fluorescence images from infected regions on attached leaves. As a first symptom of the plant-virus interaction between resistant tobacco and tobacco mosaic virus (TMV), thermal imaging detected a local rise in temperature while Chl-FI monitored a co-localized increase in fluorescence intensity. Chl-FI also revealed pre-symptomatic high-intensity spots for the plant-fungus system sugar beet-Cercospora beticola. Concomitantly, spots of lower temperature were monitored with thermography, in marked contrast with our observations on TMV-infection in tobacco. Knowledge of disease signatures for different plant-pathogen interactions could allow early identification of emerging biotic stresses in crops, facilitating the containment of disease outbreaks. Presymptomatic monitoring clearly opens perspectives for quantitative screening for disease resistance, either on excised leaf pieces or attached leaves. PMID- 15295073 TI - Whole genome analysis of the OsGRF gene family encoding plant-specific putative transcription activators in rice (Oryza sativa L.). AB - OsGRF1 (Oryza sativa GROWTH-REGULATING FACTOR1) is a rice gene encoding a putative novel transcriptional regulator. We identified and characterized eleven homologs of OsGRF1 in the rice genome. All twelve OsGRF proteins have two highly conserved regions, the QLQ (Gln, Leu, Gln) and WRC (Trp, Arg, Cys) domains, and sequences reminiscent of transcription factors. OsGRF genes were preferentially expressed in young and growing tissues, and applied gibberellic acid (GA3) enhanced the expression of seven OsGRF genes. In situ hybridization showed high levels of OsGRF1 transcripts in the shoot apical meristem and in cells surrounding the vasculature of the intercalary meristem. In a GAL4-based yeast assay, the C-terminal region of OsGRF1 was found to have transactivation activity. These results indicate that OsGRF1 acts as a transcriptional activator. Based on the in situ expression pattern of OsGRF1, we postulate that it may be involved in regulating vegetative growth in rice. PMID- 15295074 TI - The ABNORMAL GAMETOPHYTES (AGM) gene product of Arabidopsis demonstrates a role in mitosis during gamete development. AB - Screening a T-DNA mutagenized population of Arabidopsis thaliana for reduced seed set and segregation distortion led to the isolation of the ABNORMAL GAMETOPHYTES (AGM) mutant. Homozygous plants were never recovered, but heterozygous plants showed mitotic defects during gametogenesis resulting in approximately 50% abortion of both the male and female gametes. Isolation of the genomic sequence flanking the co-segregating T-DNA element led to the identification of a gene located on chromosome 5, predicted to encode a transmembrane protein. BLAST homology searches identified two homologous proteins that are not redundant, as is clear from the existence of the agm mutant. Unexpectedly, expression studies using the beta-glucuronidase reporter gene suggest that AGM and its closest Arabidopsis homolog are mostly expressed in cells undergoing mitosis. Thus, AGM is not a gametophytic gene as originally speculated on the basis of segregation distortion, but rather classified as an essential gene crucial to the process of mitosis in plants. PMID- 15295075 TI - Control of nodule number by the phytohormone abscisic Acid in the roots of two leguminous species. AB - The effects of the phytohormone abscisic acid (ABA) on plant growth and root nodule formation were analyzed in Trifolium repense (white clover) and Lotus japonicus, which form indeterminate and determinate nodules, respectively. In T. repense, although the number of nodules formed after inoculation with Rhizobium leguminosarum bv. trifolii strain 4S (wild type) was slightly affected by exogenous ABA, those formed by strain H1(pC4S8), which forms ineffective nodules, were dramatically reduced 28 days after inoculation (DAI). At 14 and 21 DAI, the number of nodules formed with the wild-type strain was decreased by exogenous ABA. In L. japonicus, the number of nodules was also reduced by ABA treatment. Thus, exogenous ABA inhibits root nodule formation after inoculation with rhizobia. Observation of root hair deformation revealed that ABA blocked the step between root hair swelling and curling. When the ABA concentration in plants was decreased by using abamine, a specific inhibitor of 9-cis-epoxycarotenoid dioxygenase, the number of nodules on lateral roots of abamine-treated L. japonicus increased dramatically, indicating that lower-than-normal concentrations of endogenous ABA enhance nodule formation. We hypothesize that the ABA concentration controls the number of root nodules. PMID- 15295076 TI - The delayed leaf senescence mutants of Arabidopsis, ore1, ore3, and ore9 are tolerant to oxidative stress. AB - Reactive oxygen species play a critical role in mediating the oxidative damage that causes senescence in a variety of aerobic organisms, from yeast to mammals. Genetic studies of these organisms have revealed that extended longevity is frequently associated with an increased resistance to stress. However, the relationship between life span and oxidative stress tolerance in plants is poorly understood. We have investigated the responses to oxidative stress in the delayed leaf senescence mutants of Arabidopsis thaliana, ore1, ore3, and ore9. The detached leaves of these mutants exhibit increased tolerance to various types of oxidative stress. The ore1, ore3, and ore9 mutants were also more tolerant to oxidative stress at the level of the whole plant, as determined by measuring physiological and molecular changes associated with oxidative stress. However, the activities of antioxidant enzymes were similar or lower in the mutants, as compared to wild type. These results suggest that the increased resistance to oxidative stress in the ore1, ore3, and ore9 mutants is not due to enhanced activities of these antioxidant enzymes. Taken together, our findings provide genetic evidence that oxidative stress tolerance is linked to control of leaf longevity in plants. PMID- 15295077 TI - Characterization of Tpn1 family in the Japanese morning glory: En/Spm-related transposable elements capturing host genes. AB - Some mutant phenotypes are known to be unstable somatically and germinally due to the insertion of transposable elements in the Japanese morning glory (Ipomoea nil). Several transposable elements that cause mutable phenotypes have recently been isolated. All of these elements show characteristic features of the En/Spm (Enhancer/Suppressor-mutator) or CACTA family. They carry common 28 bp terminal inverted repeats and subterminal repetitive regions and are known as the Tpn1 family. All of these elements are thought to be non-autonomous and mobilized by unidentified autonomous element(s). Using a probe corresponding to the subterminal region, we isolated many genomic Tpn clones, 120 of which were classified into 28 types based on their restriction maps. The copy number of the Tpn1 family was estimated to be between 500 and 1,000 copies per haploid genome. We then determined the complete sequences of 28 representative clones from each Tpn type. Most Tpn elements showed a high degree of similarity to plant genes in their internal sequences, suggesting that the Tpn1 family captured host gene sequences during the process of evolution. Detailed analyses of Tpn104 in comparison with an orthologous host gene InAP2B confirmed this assumption. PMID- 15295078 TI - GMPOZ, a BTB/POZ domain nuclear protein, is a regulator of hormone responsive gene expression in barley aleurone. AB - GAMYB is a GA-responsive activator of hydrolase gene expression in the aleurone layer of germinated cereal grains. We have isolated a putative GAMYB-binding protein, GMPOZ, which contains a BTB/POZ domain found in certain animal transcriptional regulators. Although BTB/POZ domain proteins are numerous in plants, very few are yet characterized. We found that GMPOZ is nuclear localized and that GMPOZ mRNA is expressed highly in anthers as well as aleurone. Transient silencing of the GMPOZ gene suggests that GMPOZ is involved in hormone responsive gene expression in aleurone. PMID- 15295079 TI - Contribution of the plasma membrane and central vacuole in the formation of autolysosomes in cultured tobacco cells. AB - Autolysosomes accumulate in tobacco cells cultured under sucrose starvation conditions in the presence of a cysteine protease inhibitor. We characterized these plant autolysosomes using fluorescent dyes and green fluorescent protein (GFP). Observation using the endocytosis markers, FM4-64 and Lucifer Yellow CH, suggested that there is a membrane flow from the plasma membrane to autolysosomes. Using these dyes as well as GFP-AtVam3p, sporamin-GFP and gamma VM23-GFP fusion proteins as markers of the central vacuole, we found transport of components of the central vacuole to autolysosomes. Thus endocytosis and the supply from the central vacuole may contribute to the formation of autolysosomes. PMID- 15295080 TI - Proton spectroscopy detected myoinositol in children with traumatic brain injury. AB - Previous studies have shown that proton magnetic resonance spectroscopy (MRS) is useful in predicting neurologic prognosis in children with traumatic brain injury (TBI). Reductions in N-acetyl derived metabolites and presence of lactate have been predictive of poor outcomes. We examined another spectroscopy metabolite, myoinositol (mI), to determine whether it is altered after TBI. Found primarily in astrocytes, mI functions as an osmolyte and is involved in hormone response pathways and protein-kinase C activation. Myoinositol is elevated in the newborn brain and is increased in a variety of diseases. We studied 38 children (mean age 11 y; range 1.6-17 y) with TBI using quantitative short echo time occipital gray and parietal white matter proton MRS at a mean of 7 d (range 1-17 d) after injury. We found that occipital gray matter mI levels were increased in children with TBI (4.30 +/- 0.73) compared with controls (3.53 +/- 0.48; p = 0.003). We also found that patients with poor outcomes 6-12 mo after injury had higher mI levels (4.78 +/- 0.68) than patients with good outcomes (4.15 +/- 0.69; p < 0.05). Myoinositol is elevated after pediatric TBI and is associated with a poor neurologic outcome. The reasons for its elevation remain unclear but may be due to astrogliosis or to a disturbance in osmotic function. PMID- 15295081 TI - Aerosolized PGE1: a selective pulmonary vasodilator in neonatal hypoxemic respiratory failure results of a Phase I/II open label clinical trial. AB - Twenty term/near term neonates with hypoxemic respiratory failure and oxygenation index >/=20 were enrolled in a Phase I/II feasibility, safety and dose escalation study of inhaled PGE(1) (IPGE(1)). Incremental doses of IPGE(1), delivered by a jet nebulizer over a 2-h period, followed by weaning over 1 h, were given to 13 patients before receiving inhaled nitric oxide (INO) (Group I), and to seven patients, who failed to respond to INO (Group II). Response was defined as an increase in P(a)O(2) of either >/= 25 (full) or 10-25 (partial) torr. Exit criteria included an acute deterioration in oxygenation status, a persistent oxygenation index above 35 in Group I, or the availability of extracorporeal membrane oxygenation (ECMO) in Group II. The mean (SD) increase in P(a)O(2) at the end of IPGE(1) administration was 63 (62.3) in Group I (p = 0.024), and 40 (62.1) in Group II (p > 0.05). In Group I, 8 of 13 neonates had a full response, but 4 deteriorated following discontinuation of IPGE(1). Of these four, two responded to INO and two were placed on ECMO. Five patients deteriorated before or during IPGE(1,) and none of them responded to INO. In Group II, three of seven patients had a full response to IPGE(1). One patient with a partial response and all patients exiting before or during IPGE(1) administration were placed on ECMO. The results of our study indicate that IPGE(1) may be a safe, selective pulmonary vasodilator in neonatal hypoxemic respiratory failure. PMID- 15295082 TI - Effects of citrin deficiency in the perinatal period: feasibility of newborn mass screening for citrin deficiency. AB - Deficiency of citrin due to mutations of the SLC25A13 gene causes adult-onset type II citrullinemia (CTLN2) and one type of neonatal intrahepatic cholestasis (NICCD). About half of the NICCD patients are detected based on high galactose, phenylalanine, and/or methionine concentrations on newborn mass screening (NMS). To clarify the perinatal and neonatal effects and the inconsistent results on NMS, we examined aminograms, the levels of bile acids and galactose in dried blood spots for NMS from 20 patients with NICCD. Birth weight was low for gestational age (-1.4 +/- 0.7 SD). Affected fetuses may have suffered intrauterine citrin deficiency. The first abnormality detected after birth was citrullinemia, and 19 of 20 patients had citrulline levels higher than +2 SD of controls. Tyrosine, phenylalanine, methionine, galactose, and bile acids were less affected than citrulline on d 5 after birth. Galactose and bile acids levels were increased at 1 mo in comparison with d 5 after birth due to impairment of the cytosolic NADH reducing-equivalent supply into mitochondria of hepatocytes. Patients with negative findings on NMS had low levels of total 20 amino acids. Citrulline/serine, citrulline /leucine plus isoleucine, and citrulline/total amino acids ratios, controlled for the confounding effect of low amount of total amino acids, were higher in all patients than +2 SD, +2 SD, and +3 SD of controls, respectively. NMS for citrin deficiency (frequency of homozygote with SLC25A13 mutation: 1/10,000-1/38,000 in East Asia) will be useful for clarification of the clinical course, treatment, and prevention of this disease. PMID- 15295083 TI - Postnatal maturation of cytochrome oxidase and lactate dehydrogenase activity and age-dependent consequences of lithium-pilocarpine status epilepticus in the rat: a regional histoenzymology study. AB - The lithium-pilocarpine (Li-Pilo) model of epilepsy reproduces some pathophysiological, temporal, and developmental features of human temporal lobe epilepsy. In this model, rates of cerebral glucose utilization measured by the [(14)C]2-deoxyglucose technique increased during the initial status epilepticus (SE) and decreased during the latent or chronic periods. To correlate these metabolic changes with the activities of the enzymes of the glycolytic and tricarboxylic acid cycle pathways, we measured by histoenzymology the regional activity of two key enzymes of glucose metabolism, lactate dehydrogenase (LDH) for the anaerobic pathway and cytochrome oxidase (CO) for the aerobic pathway coupled to oxidative phosphorylation, at various times after SE induced by Li Pilo in 10- (P10), 21-d-old (P21) and adult rats for CO and in adult rats only for LDH. CO activity was slightly affected in P10 and P21 rats only at 4 and 24 h and normalized by 14 d after SE. In adult rats, CO activity decreased at 4 and 24 h in damaged areas, like entorhinal cortex, hippocampal CA3 area, amygdala, and thalamus. At 14 d after SE, CO activity was decreased only in entorhinal cortex and increased in brainstem regions involved in the remote control of seizures. In adult rats, LDH activity decreased at 24 h and 14 d after SE in sensorimotor and entorhinal cortex. These data show that the enzymatic equipment underlying the metabolism of glucose is not severely affected by Li-Pilo SE and confirm our previous observations concerning the relative metabolic hyperactivity of brain regions involved in the seizure circuit despite marked neuronal loss. PMID- 15295084 TI - Cytomegalovirus (CMV) inactivation in breast milk: reassessment of pasteurization and freeze-thawing. AB - Breast-feeding mothers frequently transmit cytomegalovirus (CMV) to preterm infants of very low birth weight. Current recommendations for prevention of virus transmission are based on data published 20 y ago in the context of human milk banking. Two recent clinical trials examined storage of breast milk at -20 degrees Celsius to reduce virus transmission. However, in both studies, CMV transmission occurred. Using sensitive tools like quantitative PCR, CMV pp67 late mRNA assay, and a high-speed, centrifugation-based microculture assay for quantification of CMV infectivity, we reassessed the virological and biochemical characteristics of freeze-storing breast milk at -20 degrees Celsius, compared it with traditional Holder pasteurization (30 min at 62.5 degrees Celsius), and a new short-term pasteurization (5 s at 72 degrees Celsius) based on the generation of a milk film. Both heat treatment procedures were able to destroy viral infectivity and pp67 RNA completely. Preliminary results showed short-term heat inactivation below 72 degrees Celsius was less harmful in reducing the activity of marker enzymes than Holder pasteurization. Freezing breast milk preserved the biochemical and immunologic quality of the milk; however, late viral RNA and viral infectivity was also preserved. Compared with viral DNA, CMV-RNA more directly reflects infectious CMV in human milk samples. Further studies are necessary to evaluate short-term heat treatment below 72 degrees Celsius as an effective tool for prevention of CMV transmission. PMID- 15295085 TI - Cholesterol synthesis and de novo lipogenesis in premature infants determined by mass isotopomer distribution analysis. AB - Premature infants change from placental supply of mainly carbohydrates to an enteral supply of mainly lipids earlier in their development than term infants. The metabolic consequences hereof are not known but might have long-lasting health effects. In fact, knowledge of lipid metabolism in premature infants is very limited. We have quantified de novo lipogenesis and cholesterogenesis on d 3 of life in seven premature infants (birth weight, 1319 +/- 417 g; gestational age, 30 +/- 2 wk). For comparison, five healthy adult subjects were also studied. All subjects received a 12-h [1-(13)C] acetate infusion, followed by mass isotopomer distribution analysis (MIDA) on lipoprotein-palmitate and plasma unesterified cholesterol. The fraction of lipoprotein-palmitate synthesized at the end of the infusion period was 5.4 +/- 3.9% in infants, which was in the same range as found in adult subjects on a normal diet, suggesting that hepatic de novo lipogenesis is not a major contributor to fat accumulation in these premature neonates. The fractional contribution of newly synthesized cholesterol to plasma unesterified cholesterol was 7.4 +/- 1.3% after a 12-h infusion. The calculated rate of endogenous cholesterol synthesis was 31 +/- 7 mg/kg/d, a value approximately three times higher than that found in adult subjects (10 +/- 6 mg/kg/d). These results indicate that the cholesterol-synthesizing machinery is well developed in premature infants. PMID- 15295086 TI - Primary pulmonary hypertension in children may have a different genetic background than in adults. AB - Mutations of the bone morphogenetic protein receptor II (BMPR2) gene on chromosome 2q33 can cause familial primary pulmonary hypertension (PPH) and may occur in 26% adult patients with sporadic disease. Other disease-related genes have been localized to chromosomes 2q31 (PPH2) and 12q13 (ALK1). The genetic background in affected children remains unclear. Thirteen children (age at diagnosis, 6 mo to 13 y; mean, 5.6 +/- 3.9 y) with invasively confirmed PPH were screened for BMPR2 mutations using denaturing HPLC and sequence analysis. In addition, all children were scanned for BMPR2 deletions by Southern blot analysis. Pulmonary artery pressure was assessed using echocardiography at rest and during exercise in 57 family members of six infants. The six families were subjected to linkage analysis. None of the 13 children had a BMPR2 mutation or deletion. Linkage to chromosome 2 or 12 could not be confirmed in any of the families investigated. In all assessed families, both parents of the index patient and/or members of both branches revealed an abnormal pulmonary artery systolic pressure (PASP)-response to exercise. PPH in children may have a different genetic background than in adults. We postulate a recessive mode of inheritance in a proportion of infantile cases. PMID- 15295087 TI - Zinc homeostasis in premature infants does not differ between those fed preterm formula or fortified human milk. AB - The objectives of this study were to compare zinc homeostasis in premature infants enterally fed with either preterm infant formula or fortified human milk; to examine interrelationships of variables of zinc homeostasis; and to examine the findings in relation to estimated zinc requirements of preterm infants. Zinc homeostasis was studied in 14 infants (8 male), with mean gestational age of 31 wk and birth weight appropriate for gestational age, who were exclusively fed either preterm formula (n = 9) or own mother's milk with human milk fortifier (n = 5). Zinc stable isotopes were administered intravenously ((70)Zn) and orally as an extrinsic label ((67)Zn) over multiple feeds for determination of fractional absorption by dual isotope tracer ratio in urine; endogenous fecal zinc was determined by isotope dilution; and exchangeable zinc pool (EZP) size was estimated from linear regression of log-transformed urine (70)Zn enrichment data. Results indicated no significant differences in the variables of zinc homeostasis between the feeding groups; data for all subjects were thus combined. Mean (+/- SD) fractional absorption was 0.26 +/- 0.07; net absorbed zinc 0.43 +/- 0.25 mg/d (0.31 +/- 0.19 mg/kg/d). Mean EZP was 20 +/- 10 mg/kg, and was positively correlated with total absorbed zinc and with net absorbed zinc. Feeding type and total absorbed zinc were significantly related to daily weight gain (p = 0.003). Current zinc intakes from fortified human milk or formula are associated with acceptable weight gain, but whether the observed net zinc absorption was optimal in the human milk group cannot be definitively determined from these data. PMID- 15295088 TI - Respiratory adaptations to lung morphological defects in adult mice lacking Hoxa5 gene function. AB - The Hoxa5 mutation is associated with a high perinatal mortality rate caused by a severe obstruction of the laryngotracheal airways, pulmonary dysmorphogenesis, and a decreased production of surfactant proteins. Surviving Hoxa5(-/-) mutant mice also display lung anomalies with deficient alveolar septation and areas of collapsed tissue, thus demonstrating the importance of Hoxa5 throughout lung development and maturation. Here, we address the functional consequences of the Hoxa5 mutation on respiration and chemoreflexes by comparing the breathing pattern of Hoxa5(-/-) mice to that of wild-type animals under resting conditions and during exposure to moderate ventilatory stimuli such as hypoxia and hypercapnia. Resting Hoxa5(-/-) mice present a higher breathing frequency and overall minute ventilation that likely compensate for their reduced lung alveolar surface available for gas exchange and their increased upper airway resistance. When exposed to ventilatory stimuli, Hoxa5(-/-) mice maintain the higher minute ventilation by adapting the tidal volume and/or the breathing frequency. The minute ventilation increase seen during hypoxia was similar for both groups of mice; however, the dynamics of the frequency response was genotype-dependent. The hypercapnic ventilatory response did not differ between genotypes. These findings reveal the strategies allowing survival of Hoxa5(-/-) mice facing morphologic anomalies leading to a significant deficit in gas exchange capacity. PMID- 15295089 TI - Possible synergic effect of angiotensin-I converting enzyme gene insertion/deletion polymorphism and angiotensin-II type-1 receptor 1166A/C gene polymorphism on ischemic heart disease in patients with Kawasaki disease. AB - ACE I/D and AT1R 1166A/C polymorphisms are considered to comprise individual risk factors for the development of coronary disease. We sought to demonstrate that the ACE I/D and AT1R 1166A/C polymorphisms affect coronary artery stenosis in patients with Kawasaki disease (KD). We examined 147 healthy controls and 281 Japanese children with KD. The patients were further divided into group N (n = 246, no ischemia) and group I (n = 35, severe coronary artery stenosis with myocardial ischemia), and we studied the genotype of ACE I/D and AT1R 1166A/C polymorphisms. We also examined ACE activity in patients with acute KD. We did not detect any prevalent genotypes of the ACE and AT1R polymorphisms between controls and KD patients. However, the prevalence of the D allele in the ACE polymorphism and of the C allele in the AT1R polymorphism tended to be higher in group I than in group N (odds ratios, 2.00 and 2.32, respectively). In addition, the presence of the D and/or C alleles significantly increased the relative risk of developing myocardial ischemia (odds ratio, 2.71; p = 0.038). During the convalescent phase of KD, ACE activity was increased despite significant attenuation during the acute phase. These results suggested that the renin angiotensin system is associated with the formation of severe coronary artery stenosis and myocardial ischemia. PMID- 15295090 TI - Intrauterine growth restriction ameliorates the effects of gradual hemorrhagic hypotension on regional cerebral blood flow and brain oxygen uptake in newborn piglets. AB - Data are scant regarding the development of cerebrovascular autoregulation in intrauterine growth-restricted (IUGR) newborns. We tested the hypothesis that IUGR improves the ability of neonates to withstand critical periods of gradual hemorrhagic hypotension by optimizing cerebrovascular autoregulation. Studies were conducted on 1-d-old anesthetized piglets divided into groups of normal weight (NW, n = 14, body weight = 1518 +/- 122 g) and IUGR (n = 14, body weight = 829 +/- 50 g) animals. Physiologic parameters, including regional cerebral blood flow (CBF) and cerebral metabolic rate of oxygen (CMRO(2)), were similar in NW and IUGR piglets under baseline conditions. Controlled arterial blood loss [hemorrhagic hypotension (HH)] induced a stepwise reduction of the mean arterial blood pressure of 49 +/- 3 mm Hg (mild HH), 39 +/- 3 mm Hg (moderate HH), and 30 +/- 3 mm Hg (severe HH) in seven NW and seven IUGR piglets (p < 0.05). In NW piglets, cortical CBF and CMRO(2) was reduced already at moderate HH (p < 0.05). A similar CMRO(2) reduction occurred during severe HH in NW and IUGR piglets (p < 0.05). In addition, during mild and moderate HH, primarily in IUGR piglets, an increase in regional CBF of brainstem, cerebellum, and thalamus was shown compared with baseline values (p < 0.05). Furthermore, under these conditions, cerebral cortex blood flow was maintained in newborn IUGR animals. In contrast, NW piglets exhibited a significant reduction in CBF (p < 0.05) during moderate HH. Thus, IUGR resulted in an improved ability to withstand critical periods of gradual oxygen deficit as shown by improved cerebrovascular autoregulation during hemorrhagic hypotension. PMID- 15295091 TI - Manipulation of antioxidant pathways in neonatal murine brain. AB - To assess the role of brain antioxidant capacity in the pathogenesis of neonatal hypoxic-ischemic brain injury, we measured the activity of glutathione peroxidase (GPX) in both human-superoxide dismutase-1 (hSOD1) and human-GPX1 overexpressing transgenic (Tg) mice after neonatal hypoxia-ischemia (HI). We have previously shown that mice that overexpress the hSOD1 gene are more injured than their wild type (WT) littermates after HI, and that H(2)O(2) accumulates in HI hSOD1-Tg hippocampus. We hypothesized that lower GPX activity is responsible for the accumulation of H(2)O(2). Therefore, increasing the activity of this enzyme through gene manipulation should be protective. We show that brains of hGPX1-Tg mice, in contrast to those of hSOD-Tg, have less injury after HI than WT littermates: hGPX1-Tg, median injury score = 8 (range, 0-24) versus WT, median injury score = 17 (range, 2-24), p < 0.01. GPX activity in hSOD1-Tg mice, 2 h and 24 h after HI, showed a delayed and bilateral decline in the cortex 24 h after HI (36.0 +/- 1.2 U/mg in naive hSOD1-Tg versus 29.1 +/- 1.7 U/mg in HI cortex and 29.2 +/- 2.0 for hypoxic cortex, p < 0.006). On the other hand, GPX activity in hGPX1-Tg after HI showed a significant increase by 24 h in the cortex ipsilateral to the injury (48.5 +/- 5.2 U/mg, compared with 37.2 +/- 1.5 U/mg in naive hGPX1 Tg cortex, p < 0.008). These findings support the hypothesis that the immature brain has limited GPX activity and is more susceptible to oxidative damage and may explain the paradoxical effect seen in ischemic neonatal brain when SOD1 is overexpressed. PMID- 15295092 TI - Postnatal maturation of phosphodiesterase 5 (PDE5) in piglet pulmonary arteries: activity, expression, effects of PDE5 inhibitors, and role of the nitric oxide/cyclic GMP pathway. AB - After birth and during the first days of extrauterine life, pulmonary arterial pressure is progressively reduced to reach the adult values. We hypothesized that changes in PDE5 activity might be involved in the pulmonary postnatal maturation of the nitric oxide (NO)/cGMP pathway. The PDE5 inhibitor sildenafil produced vasorelaxant responses in isolated pulmonary arteries. These effects were similar in newborn (3-18 h) and 2-wk-old piglets, unchanged by endothelium removal, and markedly inhibited by the soluble guanylyl cyclase inhibitor ODQ. The peak of the transient vasorelaxant response to NO gas increased with postnatal age but was unaffected by PDE inhibition. However, the duration of the response to NO was significantly increased. The vasorelaxant response to sodium nitroprusside was potentiated by sildenafil in both age groups. The PDE5 inhibitors dipyridamole and zaprinast, produced qualitatively similar effects but with lower potency. Both total and PDE5-dependent cGMP hydrolytic activity and PDE5 protein expression increased with postnatal age. All these results suggest that PDE5 is a key regulator of NO-induced vasodilation in the postnatal pulmonary arteries. PDE5 inhibition is able to produce pulmonary vasodilation even in the absence of a functional endothelium and potentiates the vasorelaxant response to exogenous NO and nitroprusside. However, PDE5 is not responsible for the maturational increase of NO bioactivity during the first days of extrauterine life. PMID- 15295093 TI - Human milk--derived oligosaccharides and plant-derived oligosaccharides stimulate cytokine production of cord blood T-cells in vitro. AB - Human milk contains large amounts of free oligosaccharides (HMOs). HMOs have been shown to exert antiinflammatory properties, and evidence for their immunomodulatory effects is increasing. The purpose of this study was to evaluate influences of two human breast milk-derived oligosaccharide samples (neutral and acidic oligosaccharides), and of a low-molecular-weight fucoidan on cytokine production and activation of cord blood mononuclear cells. Cord blood mononuclear cells from randomly chosen healthy newborns were co-cultured with the oligosaccharide samples. By means of flow cytometry, intracellular cytokine production (d 20) and surface marker expression of T cells (d 5) were measured. In vitro-induced Ig levels were quantified nephelometrically (total IgG1) and by ELISA (total IgE) in the supernatant of cell cultures. The acidic oligosaccharide fraction increased the percentage of interferon-gamma producing CD3+CD4+ and CD3+CD8+ cells (p < 0.05) and the IL-13 production in CD3+CD8+ cells (p < 0.05). In acidic oligosaccharide cultures, CD25+ expression on CD3+CD4+ cells was significantly elevated (p < 0.05). Low-molecular-weight fucoidan induced IL-4 production in CD3+CD4+ T cells (p < 0.05) and IL-13 production in CD3+CD8+ T cells (p < 0.05), whereas interferon-gamma production remained unaffected in both T-cell populations. Ig production (total IgE and total IgG1) remained unaffected. Human milk-derived oligosaccharides and plant-derived oligosaccharides affect the cytokine production and activation of cord blood derived T cells in vitro. Therefore, oligosaccharides and, in particular, acidic oligosaccharides may influence lymphocyte maturation in breast-fed newborns. PMID- 15295094 TI - An EP4 receptor agonist prevents indomethacin-induced closure of rat ductus arteriosus in vivo. AB - Indomethacin exerts a strong tocolytic effect by suppressing uterine contractions mediated by prostaglandins. However, indomethacin also induces in utero closure of fetal ductus arteriosus (DA), leading to serious neonatal consequences. Using rats, we tested the effect of an agonist for a subtype of prostaglandin E2 receptor (EP4), ONO-AE1-437 and its prodrug ONO-4819, as a DA dilator during indomethacin treatment. In vitro, ONO-AE1-437 exhibited a potent dilatory effect on DA against O(2)- and indomethacin-induced contractions in a concentration dependent manner. In vivo, rat dams were given indomethacin (10 mg/kg, p.o.) alone or with ONO-4819 (0.3 micrograms/kg/h, s.c.) on d 21 of gestation and pups were delivered 4 h later through cesarean section to evaluate the ratio of diameter of DA to that of pulmonary artery. Pups from dams with no drug had DA/PA ratio of 0.9 +/- 0.05, whereas those from dams with indomethacin alone had a decreased ratio of 0.2 +/- 0.03. When ONO-4819 was co-administered to the dams, the ratio recovered significantly to 0.7 +/- 0.06. The administration of ONO-4819 to the dams did not induce any increase in the uterine activity. These results suggest that administration of an EP4 agonist in addition to indomethacin might prevent adverse reactions of indomethacin on fetal DA without restricting its tocolytic effects. PMID- 15295095 TI - Endotracheal colonization at birth is associated with a pathogen-dependent pro- and antiinflammatory cytokine response in ventilated preterm infants: a prospective cohort study. AB - The possible association between mediators of inflammation such as cytokines and perinatal colonization of the respiratory tract remains unclear. This prospective cohort study evaluated endotracheal colonization in 141 ventilated preterm infants at birth. The relation with cytokine response in the airways and C reactive protein (CRP) in umbilical blood was investigated. Of the 141 preterm infants enrolled in this study, 37 (26%) were colonized. In addition to traditional pathogens (61%), commensal species (26%) and Mycoplasmataceae (13%) were isolated. Both the pro-inflammatory cytokines IL-1 beta, IL-6, IL-8, and tumor necrosis factor (TNF)-alpha as well as the antiinflammatory IL-10 are increased in colonized patients in a dose-dependent manner, with the strongest response in neonates colonized with Gram-negative organisms. There was no antimicrobial IL-12p70 response in colonized infants. Commensal flora is associated with the same inflammatory response as traditional pathogens. Although the umbilical cord blood CRP level was significantly higher in neonates with endotracheal colonization, it was highest in those colonized with Gram-negative organisms but still close to normal limits. Microorganisms in the endotracheal fluid of ventilated preterm infants are associated with a pathogen-specific and dose-dependent cytokine response in the airways and systemic CRP response. PMID- 15295096 TI - Determining the fetal inflammatory response in an experimental model of intrauterine inflammation in rats. AB - Intrauterine infection is a risk factor for developmental brain injuries in childhood. A variety of cytokines known to be toxic to developing brain cells have been isolated from mothers or children at risk for developmental disabilities, and these cytokines have been proposed as mediators of these injuries. We have developed a model of intrauterine inflammation that damages the developing white matter and we now hypothesize that selected cytokines are increased after our experimental inflammatory stimulus. Timed-pregnant Fischer 344 and Lewis rats were injected with 0.1 mg/kg of lipopolysaccharide (LPS) into the cervix at E15. Tumor necrosis factor-alpha (TNF-alpha), interferon-gamma (IFN gamma), IL-6, and IL-10 were measured in homogenates of fetal brain and placenta at serial time periods within the first 24 h after the inflammatory stimulus. TNF alpha was increased 20-fold in the placenta and more than 5-fold in the fetal brain after the stimulus. IFN-gamma was only increased within the fetal brain (20 fold) and IL-6 was only increased in the placenta (10-fold). IL-10 was mildly increased in the placenta and was decreased slightly in the fetal brain. Our observations show that an intrauterine inflammatory stimulus can cause large increases in Th1 cytokines within the fetal brain. The placenta can produce selected cytokines but fails to produce IFN-gamma, suggesting that the fetal immune system produces this cytokine in response to our stimulus. By studying placental and brain cytokine responses in models such as ours, the mechanisms responsible for the damage to developing white matter can be determined. PMID- 15295097 TI - Modulating effects of mannose binding lectin genotype on arterial stiffness in children after Kawasaki disease. AB - Systemic arterial stiffness is increased in patients after Kawasaki disease (KD). Recently, associations between mannose-binding lectin (MBL) gene mutation and coronary complications in infants with KD and atherosclerosis in adults have been reported. We tested the hypothesis that MBL genotype modulates arterial stiffness in children after KD. Seventy-one KD patients (42 with and 29 without coronary aneurysms), aged 9.5 +/- 3.7 y, and 41 age-matched controls were studied. We determined and compared their blood pressure, brachioradial arterial stiffness as determined by pulse wave velocity (PWV), fasting total cholesterol, serum MBL level, and MBL genotype. Additionally, the modulating effects of different MBL expression genotypes [high level (HL) versus intermediate or low level (IL/LL)] on arterial stiffness in different groups were assessed. The MBL genotype distributions did not differ between patients and controls (p = 0.41) or between patients with and without coronary aneurysms (p = 0.42). Patients with IL/LL expression genotypes had significantly faster PWV than those with HL expression genotypes (7.93 +/- 1.38 m/s versus 6.67 +/- 2.28 m/s, p = 0.027). This genotype modulating effect is more pronounced in patients without (HL 8.86 +/- 0.77 m/s versus IL/LL 6.48 +/- 2.32 m/s, p = 0.02) than those with (HL 7.50 +/- 1.41 m/s versus IL/LL 6.80 +/- 2.28 m/s, p = 0.32) coronary aneurysms. Multiple linear regression analysis identified age (beta = 0.26, p = 0.012), being a Kawasaki patient (beta = 0.22, p = 0.015), and MBL IL/LL genotype subgroup (beta = 0.20, p = 0.03) as significant determinants of arterial stiffness in the entire cohort. In conclusion, MBL genotype modulates arterial stiffness, an important cardiovascular risk factor, in children after KD. PMID- 15295098 TI - Auxin-induced SCFTIR1-Aux/IAA interaction involves stable modification of the SCFTIR1 complex. AB - The plant hormone auxin can regulate gene expression by destabilizing members of the Aux/IAA family of transcriptional repressors. Auxin-induced Aux/IAA degradation requires the protein-ubiquitin ligase SCF(TIR1), with auxin acting to enhance the interaction between the Aux/IAAs and SCF(TIR1). SKP1, Cullin, and an F-box-containing protein (SCF)-mediated degradation is an important component of many eukaryotic signaling pathways. In all known cases to date, the interaction between the targets and their cognate SCFs is regulated by signal-induced modification of the target. The mechanism by which auxin promotes the interaction between SCF(TIR1) and Aux/IAAs is not understood, but current hypotheses propose auxin-induced phosphorylation, hydroxylation, or proline isomerization of the Aux/IAAs. We found no evidence to support these hypotheses or indeed that auxin induces any stable modification of Aux/IAAs to increase their affinity for SCF(TIR1). Instead, we present data suggesting that auxin promotes the SCF(TIR1) Aux/IAA interaction by affecting the SCF component, TIR1, or proteins tightly associated with it. PMID- 15295100 TI - A mechanism for slow release of biomagnified cyanobacterial neurotoxins and neurodegenerative disease in Guam. AB - As root symbionts of cycad trees, cyanobacteria of the genus Nostoc produce beta methylamino-l-alanine (BMAA), a neurotoxic nonprotein amino acid. The biomagnification of BMAA through the Guam ecosystem fits a classic triangle of increasing concentrations of toxic compounds up the food chain. However, because BMAA is polar and nonlipophilic, a mechanism for its biomagnification through increasing trophic levels has been unclear. We report that BMAA occurs not only as a free amino acid in the Guam ecosystem but also can be released from a bound form by acid hydrolysis. After first removing free amino acids from tissue samples of various trophic levels (cyanobacteria, root symbioses, cycad seeds, cycad flour, flying foxes eaten by the Chamorro people, and brain tissues of Chamorros who died from amyotrophic lateral sclerosis/Parkinsonism dementia complex), we then hydrolyzed the remaining fraction and found BMAA concentrations increased 10- to 240-fold. This bound form of BMAA may function as an endogenous neurotoxic reservoir, accumulating and being transported between trophic levels and subsequently being released during digestion and protein metabolism. Within brain tissues, the endogenous neurotoxic reservoir can slowly release free BMAA, thereby causing incipient and recurrent neurological damage over years or even decades, which may explain the observed long latency period for neurological disease onset among the Chamorro people. The presence of BMAA in brain tissues from Canadian patients who died of Alzheimer's disease suggests that exposure to cyanobacterial neurotoxins occurs outside of Guam. PMID- 15295099 TI - Chemical chaperones protect from effects of apoptosis-inducing mutation in carbonic anhydrase IV identified in retinitis pigmentosa 17. AB - Carbonic anhydrase (CA) IV is a glycosylphosphotidylinositol-anchored enzyme highly expressed on the plasma face of microcapillaries and especially strongly expressed in the choriocapillaris of the human eye. In collaboration with scientists at the University of Cape Town (Rondebosch, South Africa), we recently showed that the R14W mutation in the signal sequence of CA IV, which they identified in patients with the retinitis pigmentosa (RP) 17 form of autosomal dominant RP, results in accumulation of unfolded protein in the endoplasmic reticulum (ER), leading to ER stress, the unfolded protein response, and apoptosis in a large fraction of transfected COS-7 cells expressing mutant, but not wild-type, CA IV. Here we present experiments showing that several well characterized CA inhibitors largely prevent the adverse effects of expressing R14W CA IV in transfected COS-7 cells. Specifically, CA inhibitors prevent the accelerated turnover of the mutant protein, the up-regulation of Ig-binding protein, double-stranded RNA-regulated protein kinase-like ER kinase, and CCAAT/enhancer-binding protein homologous protein (markers of the unfolded protein response and ER stress), the inhibition of production of other secretory proteins expressed from COS-7-transfecting plasmids, and the induction of apoptosis, all characteristics of transfected cells expressing R14W CA IV. Furthermore, treatment with 4-phenylbutyric acid, a nonspecific chemical chaperone used in other protein-folding disorders, also dramatically reduces the apoptosis-inducing effect of expressing R14W CA IV cDNA in transfected COS-7 cells. These experiments suggest a promising approach to treatment of RP17 that might delay the onset or possibly prevent this autosomal dominant form of RP. PMID- 15295101 TI - Accelerated immunosenescence in preindustrial twin mothers. AB - Life-history theory predicts a tradeoff between reproductive effort and lifespan. It has been suggested that this tradeoff is a result of reproductive costs accelerating senescence of the immune system, leading to earlier death. Longevity costs of reproduction are suggested for some human populations, but whether high reproductive effort leads to impaired immune function is unknown. We examined how reproductive effort affected postreproductive survival and the probability of dying of an infectious disease in women born in preindustrial Finland between 1702 and 1859. We found that mothers delivering twins had reduced postreproductive survival after age 65. This effect arose because mothers of twins had a higher probability of succumbing to an infectious disease (mainly tuberculosis) than mothers delivering singletons. The risk among mothers of twins of dying of an infectious disease was further elevated if mothers had started reproducing early. In contrast, neither female postreproductive survival nor the risk of succumbing to an infectious disease was influenced by the total number of offspring produced. Our results provide evidence of a long-term survival cost of twinning in humans and indicate that the mechanism mediating this cost might have been accelerated immunosenescence. PMID- 15295102 TI - YY1 inhibits the activation of the p53 tumor suppressor in response to genotoxic stress. AB - The tumor suppressor p53 regulates cell-cycle progression and apoptosis in response to genotoxic stress, and inactivation of p53 is a common feature of cancer cells. The levels and activity of p53 are tightly regulated by posttranslational modifications, including phosphorylation, ubiquitination, and acetylation. Here, we demonstrate that the transcription factor Yin Yang 1 (YY1) interacts with p53 and inhibits its transcriptional activity. We show that YY1 disrupts the interaction between p53 and the coactivator p300 and that expression of YY1 blocks p300-dependent acetylation and stabilization of p53. Furthermore, expression of YY1 inhibits the accumulation of p53 and the induction of p53 target genes in response to genotoxic stress. YY1 also interacts with Mdm2 and the expression of YY1 promotes the assembly of the p53-Mdm2 complex. Consequently, YY1 enhances Mdm2-mediated ubiquitination of p53. Inactivation of endogenous YY1 enhances the accumulation of p53 as well as the expression of p53 target genes in response to DNA damage, and it sensitizes cells to DNA damage induced apoptosis. Hence, our results demonstrate that YY1 regulates the transcriptional activity, acetylation, ubiquitination, and stability of p53 by inhibiting its interaction with the coactivator p300 and by enhancing its interaction with the negative regulator Mdm2. YY1 may, therefore, be an important negative regulator of the p53 tumor suppressor in response to genotoxic stress. PMID- 15295103 TI - The xipotl mutant of Arabidopsis reveals a critical role for phospholipid metabolism in root system development and epidermal cell integrity. AB - Phosphocholine (PCho) is an essential metabolite for plant development because it is the precursor for the biosynthesis of phosphatidylcholine, which is the major lipid component in plant cell membranes. The main step in PCho biosynthesis in Arabidopsis thaliana is the triple, sequential N-methylation of phosphoethanolamine, catalyzed by S-adenosyl-l-methionine:phosphoethanolamine N methyltransferase (PEAMT). In screenings performed to isolate Arabidopsis mutants with altered root system architecture, a T-DNA mutagenized line showing remarkable alterations in root development was isolated. At the seedling stage, the mutant phenotype is characterized by a short primary root, a high number of lateral roots, and short epidermal cells with aberrant morphology. Genetic and biochemical characterization of this mutant showed that the T-DNA was inserted at the At3g18000 locus (XIPOTL1), which encodes PEAMT (XIPOTL1). Further analyses revealed that inhibition of PCho biosynthesis in xpl1 mutants not only alters several root developmental traits but also induces cell death in root epidermal cells. Epidermal cell death could be reversed by phosphatidic acid treatment. Taken together, our results suggest that molecules produced downstream of the PCho biosynthesis pathway play key roles in root development and act as signals for cell integrity. PMID- 15295105 TI - Suppressed RAGE. PMID- 15295106 TI - The thing that wouldn't leave. PMID- 15295104 TI - Tissue-specific expression of head-to-tail cyclized miniproteins in Violaceae and structure determination of the root cyclotide Viola hederacea root cyclotide1. AB - The plant cyclotides are a family of 28 to 37 amino acid miniproteins characterized by their head-to-tail cyclized peptide backbone and six absolutely conserved Cys residues arranged in a cystine knot motif: two disulfide bonds and the connecting backbone segments form a loop that is penetrated by the third disulfide bond. This knotted disulfide arrangement, together with the cyclic peptide backbone, renders the cyclotides extremely stable against enzymatic digest as well as thermal degradation, making them interesting targets for both pharmaceutical and agrochemical applications. We have examined the expression patterns of these fascinating peptides in various Viola species (Violaceae). All tissue types examined contained complex mixtures of cyclotides, with individual profiles differing significantly. We provide evidence for at least 57 novel cyclotides present in a single Viola species (Viola hederacea). Furthermore, we have isolated one cyclotide expressed only in underground parts of V. hederacea and characterized its primary and three-dimensional structure. We propose that cyclotides constitute a new family of plant defense peptides, which might constitute an even larger and, in their biological function, more diverse family than the well-known plant defensins. PMID- 15295107 TI - Murine models of life span extension. AB - Mice are excellent experimental models for genetic research and are being used to investigate the genetic component of organismal aging. Several mutant mice are known to possess defects in the growth hormone/insulin-like growth factor 1 (GH/IGF-1) neurohormonal pathway and exhibit dwarfism together with extended life span. Their phenotypes resemble those of mice subjected to caloric restriction. Targeted mutations that affect components of this pathway, including the GH receptor, p66Shc, and the IGF-1 receptor (IGF-1R), also extend life span; mutations that affect IGF-1R or downstream components of the pathway decouple longevity effects from dwarfism. These effects on life span may result from an increased capacity to resist oxidative damage. PMID- 15295108 TI - Both chaperone and isomerase functions of protein disulfide isomerase are essential for acceleration of the oxidative refolding and reactivation of dimeric alkaline protease inhibitor. AB - Oxidative refolding of the dimeric alkaline protease inhibitor (API) from Streptomyces sp. NCIM 5127 has been investigated. We demonstrate here that both isomerase and chaperone functions of the protein folding catalyst, protein disulfide isomerase (PDI), are essential for efficient refolding of denatured reduced API (dr-API). Although the role of PDI as an isomerase and a chaperone has been reported for a few monomeric proteins, its role as a foldase in refolding of oligomeric proteins has not been demonstrated hitherto. Spontaneous refolding and reactivation of dr-API in redox buffer resulted in 45% to 50% reactivation. At concentrations <0.25 microM, reactivation rates and yields of dr API are accelerated by catalytic amounts of PDI through its isomerase activity, which promotes disulfide bond formation and rearrangement. dr-API is susceptible to aggregation at concentrations >25 microM, and a large molar excess of PDI is required to enhance reactivation yields. PDI functions as a chaperone by suppressing aggregation and maintains the partially unfolded monomers in a folding-competent state, thereby assisting dimerization. Simultaneously, isomerase function of PDI brings about regeneration of native disulfides. 5 Iodoacetamidofluorescein-labeled PDI devoid of isomerase activity failed to enhance the reactivation of dr-API despite its intact chaperone activity. Our results on the requirement of a stoichiometric excess of PDI and of presence of PDI in redox buffer right from the initiation of refolding corroborate that both the functions of PDI are essential for efficient reassociation, refolding, and reactivation of dr-API. PMID- 15295109 TI - Hexa-histidin tag position influences disulfide structure but not binding behavior of in vitro folded N-terminal domain of rat corticotropin-releasing factor receptor type 2a. AB - The oxidative folding, particularly the arrangement of disulfide bonds of recombinant extracellular N-terminal domains of the corticotropin-releasing factor receptor type 2a bearing five cysteines (C2 to C6), was investigated. Depending on the position of a His-tag, two types of disulfide patterns were found. In the case of an N-terminal His-tag, the disulfide bonds C2-C3 and C4-C6 were found, leaving C5 free, whereas the C-terminal position of the His-tag led to the disulfide pattern C2-C5 and C4-C6, and leaving C3 free. The latter pattern is consistent with the disulfide arrangement of the extracellular N-terminal domain of the corticotropin-releasing factor (CRF) receptor type 1, which has six cysteines (C1 to C6) and in which C1 is paired with C3. However, binding data of the two differently disulfide-bridged domains show no significant differences in binding affinities to selected ligands, indicating the importance of the C terminal portion of the N-terminal receptor domains, particularly the disulfide bond C4-C6 for ligand binding. PMID- 15295110 TI - Crystal structure of a dimeric form of streptococcal pyrogenic exotoxin A (SpeA1). AB - Streptococcal pyrogenic exotoxin A (SpeA1) is a bacterial superantigen associated with scarlet fever and streptococcal toxic shock syndrome (STSS). SpeA1 is found in both monomeric and dimeric forms, and previous work suggested that the dimer results from an intermolecular disulfide bond between the cysteines at positions 90 of each monomer. Here, we present the crystal structure of the dimeric form of SpeA1. The toxin crystallizes in the orthorhombic space group P212121, with two dimers in the crystallographic asymmetric unit. The final structure has a crystallographic R-factor of 21.52% for 7248 protein atoms, 136 water molecules, and 4 zinc atoms (one zinc atom per molecule). The implications of SpeA1 dimer on MHC class II and T-cell receptor recognition are discussed. PMID- 15295111 TI - Insulin amyloid fibrillation at above 100 degrees C: new insights into protein folding under extreme temperatures. AB - To investigate the folding behavior of amyloidogenic proteins under extreme temperatures, the kinetics of fibrillation and accompanying secondary structure transitions of bovine insulin were studied for temperatures ranging up to 140 degrees C. The presence of extreme heat stress had traditionally been associated with irreversible denaturation of protein while the initial steps of such a denaturation process may be common with a fibril formation pathway of amyloidogenic proteins. The present work demonstrates the ability of insulin to form amyloid fibrils at above 100 degrees C. Amyloid formation was gradually replaced by random coil generation after approximately 80 degrees C until no amyloid was detected at 140 degrees C. The morphology of insulin amyloid fibrils underwent sharp changes with increasing the temperature. The dependence of amyloid formation rate on incubation temperature followed non-Arrhenius kinetics, which is explained by temperature-dependent enthalpy change for amyloid formation. The intermediate stage of amyloid formation and random coil generation consisted of a partially folded intermediate common to both pathways. The fully unfolded monomers in random coil conformation showed partial reversibility through this intermediate by reverting back to the amyloid pathway when formed at 140 degrees C and incubated at 100 degrees C. This study highlights the non Arrhenius kinetics of amyloid fibrillation under extreme temperatures, and elucidates its intermediate stage common with random coil formation. PMID- 15295112 TI - Stabilizing roles of residual structure in the empty heme binding pockets and unfolded states of microsomal and mitochondrial apocytochrome b5. AB - The microsomal (Mc) and mitochondrial (OM) isoforms of mammalian cytochrome b5 are the products of different genes, which likely arose via duplication of a primordial gene and subsequent functional divergence. Despite sharing essentially identical folds, heme-polypeptide interactions are stronger in OM b5s than in Mc b5s due to the presence of two conserved patches of hydrophobic amino acid side chains in the OM heme binding pockets. This is of fundamental interest in terms of understanding heme protein structure-function relationships, because stronger heme-polypeptide interactions in OM b5s in comparison to Mc b5s may represent a key source of their more negative reduction potentials. Herein we provide evidence that interactions amongst the amino acid side chains contributing to the hydrophobic patches in rat OM (rOM) b5 persist when heme is removed, rendering the empty heme binding pocket of rOM apo-b5 more compact and less conformationally dynamic than that in bovine Mc (bMc) apo-b5. This may contribute to the stronger heme binding by OM apo-b5 by reducing the entropic penalty associated with polypeptide folding. We also show that when bMc apo-b5 unfolds it adopts a structure that is more compact and contains greater nonrandom secondary structure content than unfolded rOM apo-b5. We propose that a more robust beta sheet in Mc apo-b5s compensates for the absence of the hydrophobic packing interactions that stabilize the heme binding pocket in OM apo-b5s. PMID- 15295113 TI - Delineation and analysis of the conceptual data model implied by the "IUPAC Recommendations for Biochemical Nomenclature". AB - Computational analysis of the bonding, geometric, and topological relationships within proteins typically takes on the order of hours, mainly devoted to the writing of scripts and code to correctly parse the data. The Structured Query Language (SQL) built into modern database management systems eliminates the need for data parsing, effectively reducing the analysis time to seconds. To this end, we have formulated a conceptual data model (CDM) for proteins based on the IUPAC recommendations for biochemical nomenclature. This conceptual data model makes explicit the inherent bonding relationships between the atoms of a protein, as well as the geometric (bond angle and torsion angle) and topological (chirality) relationships between the bonds. The validity of the CDM has been tested with a reduced implementation using commercial database software. The ease in both populating the database with data from the Protein Data Bank and formulating/executing queries supports the correctness of the model. The ability to conduct truly interactive analyses of protein structure is essential to fully capitalize on the explosion in postgenomic protein structure data. PMID- 15295114 TI - Lipid binding in rice nonspecific lipid transfer protein-1 complexes from Oryza sativa. AB - Nonspecific lipid transfer proteins (nsLTPs) facilitate the transfer of phospholipids, glycolipids, fatty acids and steroids between membranes, with wide ranging binding affinities. Three crystal structures of rice nsLTP1 from Oryza sativa, complexed with myristic (MYR), palmitic (PAL) or stearic acid (STE) were determined. The overall structures of the rice nsLTP1 complexes belong to the four-helix bundle folding with a long C-terminal loop. The nsLTP1-MYR and the nsLTP1-STE complexes bind a single fatty acid while the nsLTP1-PAL complex binds two molecules of fatty acids. The C-terminal loop region is elastic in order to accommodate a diverse range of lipid molecules. The lipid molecules interact with the nsLTP1-binding cavity mainly with hydrophobic interactions. Significant conformational changes were observed in the binding cavity and the C-terminal loop of the rice nsLTP1 upon lipid binding. PMID- 15295115 TI - Arginine 60 in the ArsC arsenate reductase of E. coli plasmid R773 determines the chemical nature of the bound As(III) product. AB - Arsenic is a ubiquitous environmental toxic metal. Consequently, organisms detoxify arsenate by reduction to arsenite, which is then excreted or sequestered. The ArsC arsenate reductase from Escherichia coli plasmid R773, the best characterized arsenic-modifying enzyme, has a catalytic cysteine, Cys 12, in the active site, surrounded by an arginine triad composed of Arg 60, Arg 94, and Arg 107. During the reaction cycle, the native enzyme forms a unique monohydroxyl Cys 12-thiol-arsenite adduct that contains a positive charge on the arsenic. We hypothesized previously that this unstable intermediate allows for rapid dissociation of the product arsenite. In this study, the role of Arg 60 in product formation was evaluated by mutagenesis. A total of eight new structures of ArsC were determined at resolutions between 1.3 A and 1.8 A, with R(free) values between 0.18 and 0.25. The crystal structures of R60K and R60A ArsC equilibrated with the product arsenite revealed a covalently bound Cys 12-thiol dihydroxyarsenite without a charge on the arsenic atom. We propose that this intermediate is more stable than the monohydroxyarsenite intermediate of the native enzyme, resulting in slow release of product and, consequently, loss of activity. PMID- 15295116 TI - The effect of an ionic detergent on the natively unfolded beta-dystroglycan ectodomain and on its interaction with alpha-dystroglycan. AB - Dystroglycan (DG) is an adhesion complex, expressed in a wide variety of tissues, formed by an extracellular and a transmembrane subunit, alpha-DG and beta-DG, respectively, interacting noncovalently. Recently, we have shown that the recombinant ectodomain of beta-DG, beta-DG(654-750), behaves as a natively unfolded protein, as it is able to bind the C-terminal domain of alpha-DG, while not displaying a defined structural organization. We monitored the effect of a commonly used denaturing agent, the anionic detergent sodium dodecylsulphate (SDS), on beta-DG(654-750) using a number of biophysical techniques. Very low concentrations of SDS (< or =2 mM) affect both tryptophan fluorescence and circular dichroism of beta-DG, and significantly perturb the interaction with the alpha-DG subunit as shown by solid-phase binding assays and fluorescence titrations in solution. This result confirms, as recently proposed for natively unfolded proteins, that beta-DG(654-750) exists in a native state, which is crucial to fulfill its biological function. Two-dimensional NMR analysis shows that SDS does not induce any evident conformational rearrangement within the ectodomain of beta-DG. Its first 70 amino acids, which show a lower degree of mobility, interact with the detergent, but this does not change the amount of secondary structure, whereas the highly flexible and mobile C-terminal region of beta-DG(654-750) remains largely unaffected, even at a very high SDS concentration (up to 50 mM). Our data indicate that SDS can be used as a useful tool for investigating natively unfolded proteins, and confirm that the beta-DG ectodomain is an interesting model system. PMID- 15295117 TI - Characterization of SLAC: a small laccase from Streptomyces coelicolor with unprecedented activity. AB - Laccases and other four-copper oxidases are usually constructed of three domains: Domains one and three house the copper sites, and the second domain often helps form a substrate-binding cleft. In contrast to this arrangement, the genome of Streptomyces coelicolor was found to encode a small, four-copper oxidase that lacks the second domain. This protein is representative of a new family of enzymes--the two-domain laccases. Disruption of the corresponding gene abrogates laccase activity in the growth media. We have recombinantly expressed this enzyme, called SLAC, in Escherichia coli and characterized it. The enzyme binds four copper ions/monomer, and UV-visible absorption and EPR measurements confirm that the conserved type 1 copper site and trinuclear cluster are intact. We also report the first known paramagnetic NMR spectrum for the trinuclear copper cluster of a protein from the laccase family. The enzyme is highly stable, retaining activity as a dimer in denaturing gels after boiling and SDS treatment. The activity of the enzyme against 2,6-dimethoxyphenol (DMP) peaks at an unprecedentedly high pH (9.4), whereas the activity against ferrocyanide decreases with pH. SLAC binds negatively charged substrates more tightly than positively charged or uncharged molecules. PMID- 15295119 TI - What is endometritis and does it require treatment? PMID- 15295118 TI - What can one learn from experiments about the elusive transition state? AB - We present the results of an exact analysis of a model energy landscape of a protein to clarify the idea of the transition state and the physical meaning of the phi values determined in protein engineering experiments. We benchmark our findings to various theoretical approaches proposed in the literature for the identification and characterization of the transition state. PMID- 15295120 TI - Notice of redundant publication. PMID- 15295121 TI - Recent trends in the epidemiology of sexually transmitted infections in the European Union. AB - Sexually transmitted infections (STIs) are a major public health problem in Europe. We review recent trends in the epidemiology of the major acute STIs in the European Union and Norway, their key determinants, and opportunities for enhancing STI prevention interventions in the region. PMID- 15295122 TI - Surveillance systems for STIs in the European Union: facing a changing epidemiology. AB - OBJECTIVES: To characterise the nature, content, and performance characteristics of existing national STI surveillance systems in the European Union (EU) and Norway, to facilitate collection of comparable surveillance data. METHODS: Cross sectional survey using a structured questionnaire. RESULTS: Case reporting from clinicians and/or laboratories is the mainstay of EU surveillance systems for bacterial STIs. Coverage of case reporting varies from less than 10% to over 75%, and lack of and/or heterogeneity in case definitions affect the relative specificity and sensitivity of reporting systems. Considerable variations also exist in STI care sites; the populations who use these services; and in partner notification practices, STI screening practices, and STI laboratory diagnostic tests employed, affecting the representativeness of reported data and the sensitivity of surveillance systems for detecting the true number of STI cases. CONCLUSIONS: The heterogeneity of current surveillance systems complicates direct comparison of STI incidence rates across Europe. Introduction of standardised case definitions for reporting, and increased coverage of mandatory reporting systems where necessary, are needed. Definition of standardised minimum datasets and use of sentinel and enhanced surveillance systems to supplement universal case/laboratory notification data, could improve our understanding of the distribution and determinants of STIs across Europe, and aid in the design of effective public health responses. In the context of the changing epidemiology of STIs, systems for detection and monitoring of localised outbreaks of acute bacterial STIs (syphilis and antimicrobial resistant gonorrhoea), as well as prevalence monitoring systems for frequently asymptomatic STIs (chlamydial infection and viral STIs), are also necessary. PMID- 15295123 TI - Oral shedding of herpes simplex virus type 2. AB - OBJECTIVES: Herpes simplex virus (HSV) 1 and HSV-2 reactivate preferentially in the oral and genital area, respectively. We aimed to define frequency and characteristics associated with oral shedding of HSV-2. METHODS: Demographic, clinical and laboratory data of patients with documented HSV-2 infection and at least one oral viral culture obtained were selected from the University of Washington Virology Research Clinic database. RESULTS: Of 1388 people meeting the entry criteria, 44 (3.2%) had HSV-2 isolated at least once from their mouths. In comparison with the 1344 people who did not have HSV-2 isolated from their mouth, participants with oral HSV-2 were more likely to be male (OR = 1.9, 95% CI 1.0 to 3.7), HIV positive (OR = 2.9, 95% CI 1.4 to 6.0), and homosexual (OR = 2.2, 95% CI 1.1 to 4.2), and to have collected a larger number of oral specimens (median 32 v 4, p<0.001). Of the 58 days with oral HSV-2 isolation, 15 (25%) occurred during newly acquired HSV-2 infection, 12 (21%) during a recurrence with genital lesions, three (5%) during a recurrence with oral lesions, and three (5%) during a recurrence with oral and genital lesions; 25 (43%) occurred during asymptomatic shedding. Oral HSV-2 was found less frequently than oral HSV-1 (0.06% v 1%, p<0.001) in people with HSV-1 and HSV-2 antibody, and less frequently than genital HSV-2 (0.09% v 7%, p<0.001). CONCLUSIONS: Oral reactivation of HSV-2 as defined by viral isolation is uncommon and usually occurs in the setting of first episode of genital HSV-2 or during genital recurrence of HSV-2. PMID- 15295125 TI - Changing epidemiology of genital herpes simplex virus infection in Melbourne, Australia, between 1980 and 2003. AB - OBJECTIVE: To investigate changes in the proportions of patients infected with genital herpes simplex virus (HSV) types 1 and 2 from 1980 to 2003 in Melbourne, Australia. METHODS: A total of 25 372 patients were studied retrospectively. The proportions of HSV-1 and HSV-2 detected in these individuals were analysed by age, sex, and genital site. RESULTS: In 1980 only 15.8% of HSV positive genital specimens were HSV-1 compared to 34.9% in 2003. In 2003 HSV-1 was detected in 77% of patients aged less than 20 years. Females were more likely to be infected with HSV-1, although the rate of increased detection was more pronounced in males. Except for females over the age of 40, the trend for the increase in HSV-1 was detected in all age groups. No specific genital site in either sex was associated with the increase. CONCLUSIONS: The proportion of genital HSV-1 has increased in Australian patients, although HSV-2 is still the most common cause of genital infection. Confirmation of HSV type is necessary for optimal patient management. PMID- 15295126 TI - Sexual network analysis of a gonorrhoea outbreak. AB - OBJECTIVES: Sexual partnerships can be viewed as networks in order to study disease transmission. We examined the transmission of Neisseria gonorrhoeae in a localised outbreak in Alberta, Canada, using measures of network centrality to determine the association between risk of infection of network members and their position within the sexual network. We also compared risk in smaller disconnected components with a large network centred on a social venue. METHODS: During the investigation of the outbreak, epidemiological data were collected on gonorrhoea cases and their sexual contacts from STI surveillance records. In addition to traditional contact tracing information, subjects were interviewed about social venues they attended in the past year where casual sexual partnering may have occurred. Sexual networks were constructed by linking together named partners. Univariate comparisons of individual network member characteristics and algebraic measures of network centrality were completed. RESULTS: The sexual networks consisted of 182 individuals, of whom 107 were index cases with laboratory confirmed gonorrhoea and 75 partners of index cases. People who had significantly higher information centrality within each of their local networks were found to have patronised a popular motel bar in the main town in the region (p = 0.05). When the social interaction through the bar was considered, a large network of 89 individuals was constructed that joined all eight of the largest local networks. Moreover, several networks from different communities were linked by individuals who served as bridge populations as a result of their sexual partnering. CONCLUSION: Asking clients about particular social venues emphasised the importance of location in disease transmission. Network measures of centrality, particularly information centrality, allowed the identification of key individuals through whom infection could be channelled into local networks. Such individuals would be ideal targets for increased interventions. PMID- 15295127 TI - The diversity of the opa gene in gonococcal isolates from men who have sex with men. AB - OBJECTIVES: To use a molecular typing method (opa-typing) to characterise gonococcal isolates obtained from men who have sex with men (MSM) attending a genitourinary medicine (GUM) clinic in Edinburgh during 2001. To compare the results of opa-typing with A/S (auxotype/serovar) phenotyping, and with epidemiological information obtained by contact tracing. METHODS: Isolates were opa-typed by a PCR-RFLP method where the restriction fragments resulting from digestion with three separate restriction enzymes were resolved by polyacrylamide gel electrophoresis. Isolates were considered to have the same opa-type if the band patterns were identical or if they differed by one band between the three digestions. RESULTS: 40 opa-types were detected from a collection of 73 gonococcal isolates from 61 patients. 26 opa-types were unique, being found in one individual each, 14 different opa-types were found in more than one patient. Opa-typing was found to have a discrimination index (DI) of 0.96, compared to a DI of 0.87 for A/S phenotyping, indicating that opa-typing is better at discriminating between unrelated isolates. Opa-typing confirmed three epidemiological links established by contact tracing, and uncovered a further 13 clusters of isolates. CONCLUSIONS: Opa-typing is a more discriminative method than A/S phenotyping when determining relatedness in gonococcal isolates. The ability of opa-typing to identify sexual networks not disclosed by contact tracing and conventional phenotyping make it a useful method for studying the spread of gonorrhoea with the potential to contribute to the control of this infection. PMID- 15295128 TI - Symptomatic urethritis is more prevalent in men infected with Mycoplasma genitalium than with Chlamydia trachomatis. AB - OBJECTIVES: To study the prevalence, symptoms, and signs of Mycoplasma genitalium and Chlamydia trachomatis infections in men attending a Swedish STD clinic and to study the criteria for urethritis. METHODS: A cross sectional study among STD clinic attendees in Orebro, Sweden. Attendees were examined for microscopic urethritis and first void urine (FVU) was tested for M genitalium and C trachomatis. RESULTS: The prevalence of M genitalium and C trachomatis was 7% (34/512) and 12% (61/512), respectively. Dual infection was diagnosed in four men. In both infections 90% of the patients had signs of microscopic urethritis. M genitalium positive men had symptomatic urethritis significantly more often than those infected with C trachomatis (73% v 40%, RR 1.8; 95% CI 1.2 to 2.7). 63% of female partners of men infected with M genitalium were infected with M genitalium compared with chlamydial infection in 67% of female partners of men infected with C trachomatis. Non-chlamydial non-gonococcal urethritis without evidence of M genitalium infection was diagnosed in 180 men (35%). Symptoms and/or visible discharge were reported in 49% in this group. CONCLUSIONS: M genitalium is a common infection associated with symptomatic urethritis and with a high prevalence of infected sexual partners supporting its role as a sexually transmitted infection. PMID- 15295129 TI - Spatial analysis and mapping of sexually transmitted diseases to optimise intervention and prevention strategies. AB - OBJECTIVE: We analysed and mapped the distribution of four reportable sexually transmitted diseases, chlamydial infection/non-gonococcal urethritis (chlamydial infection), gonorrhoea, primary and secondary syphilis (syphilis), and HIV infection, for Wake County, North Carolina, to optimise an intervention. METHODS: We used STD surveillance data reported to Wake County, for the year 2000 to analyse and map STD rates. STD rates were mathematically represented as a spatial random field. We analysed spatial variability by calculating and modelling covariance functions of random field theory. Covariances are useful in assessing spatial patterns of disease locally and at a distance. We combined observed STD rates and appropriate covariance models using a geostatistical method called kriging, to predict STD rates and associated prediction errors for a grid covering Wake County. Final disease estimates were interpolated using a spline with tension and mapped to generate a continuous surface of infection. RESULTS: Lower incidence STDs exhibited larger spatial variability and smaller neighbourhoods of influence than higher incidence STDs. Each reported STD had a clustered spatial distribution with one primary core area of infection. Core areas overlapped for all four STDs. CONCLUSIONS: Spatial heterogeneity within STD suggests that STD specific prevention strategies should not be targeted uniformly across Wake County, but rather to core areas. Overlap of core areas among STDs suggests that intervention and prevention strategies can be combined to target multiple STDs effectively. Geostatistical techniques are objective, population level approaches to spatial analysis and mapping that can be used to visualise disease patterns and identify emerging outbreaks. PMID- 15295130 TI - Investigating ethnic differences in sexual health: focus groups with young people. AB - OBJECTIVES: To compare normative beliefs about sexual health in young men and women from black Caribbean, black African, and white ethnic groups in order to better understand ethnic inequalities in sexual health. METHODS: Focus group discussions with young people living in an area with known high prevalence of gonorrhoea and chlamydia. Groups were stratified by sex and self defined ethnicity. RESULTS: 22 male and 20 female 16-25 year olds of black Caribbean, black African, and white ethnicity took part in eight discussions. Participants from black ethnic groups were more aware of gonorrhoea than white participants but all ethnic groups regarded these as being less important than unplanned pregnancy or HIV/AIDS. Most participants believed that they would have obvious symptoms if they had a sexually transmitted infection and could determine the cleanliness of sexual partners by visual or behavioural cues. Black Caribbean women were alone in acknowledging the likelihood of their partners having concurrent sexual relationships. Some black Caribbean women described negative attitudes of staff in genitourinary medicine clinics who were from the same ethnic background. CONCLUSION: In this focus group study we identified ethnic differences in terminology, awareness of sexually transmitted infections, non exclusive sexual relationships, and experience of sexual health services but gender had a greater influence on normative beliefs. The similarities in norms for all ethnic groups might reflect common social and cultural exposures. The low priority given to sexually transmitted infections by young people from all ethnic groups needs to be addressed if they are to be tackled successfully. PMID- 15295131 TI - If the condom fits, wear it: a qualitative study of young African-American men. AB - OBJECTIVE: To extend the current knowledge base pertaining to condom failure among young African-American men by assessing their experiences with male condom use. METHODS: Qualitative assessments were conducted with 19 African-American men (aged 18-29 years) who had just been diagnosed with an STI and reported using condoms in the previous 3 months. RESULTS: Five categories were identified from the data. These categories pertained to: (1) the "fit and feel" of condoms; (2) condom brand and size; (3) application problems; (4) availability of condoms and lubricants; and (5) commitment to condom use. Common themes included reasons why men believed condoms would break or slip off during sex. Comfort problems, including tightly fitting condoms and condoms drying out during intercourse, were mentioned frequently. Condom associated erection problems were often described. Many men also noted that condom use reduced the level of sexual satisfaction for their female partners. Men noted that finding the right kind of condom was not always easy and it became apparent during the interviews that men typically did not acquire lubrication to add to condoms. Despite their expressed problems with using condoms, men were, none the less, typically emphatic that condom use is an important part of their protective behaviour against STIs. CONCLUSION: Men were highly motivated to use condoms; however, they experienced a broad range of problems with condom use. With the exception of losing the sensation of skin to skin contact, the vast majority of these problems may be amenable to behavioural interventions. PMID- 15295132 TI - Determinants of inconsistent condom use with female sex workers among men attending the STD clinic in Singapore. AB - BACKGROUND/OBJECTIVES: Female sex workers and their male clients have been identified as risk groups for the transmission of STDs and HIV. Behavioural interventions targeting clients need to address inconsistent condom use among them. The aim of the study is to assess the sociodemographic, behavioural, and psychological factors associated with inconsistent condom use among clients of sex workers. METHODS: 229 male patients attending the STD clinic in Singapore who reported paying for sex in the previous 6 months were interviewed. Response rate was 91%. RESULTS: Overall, 45% used condoms inconsistently; these clients were more likely to have poor STD knowledge, visit sex workers five or more times in the past 6 months, have lower self efficacy, less favourable social norms for condom use, and more likely to forget condom use when intoxicated (alcohol impaired decision making). CONCLUSIONS: Behavioural interventions for clients need to improve STD/HIV transmission knowledge and focus on improving client's self efficacy in using condoms. PMID- 15295133 TI - Unusual muscle disease in HIV infected patients. AB - Two patients presented with proximal muscle weakness, a normal or minor elevation of creatine phosphokinase (CPK) and normal findings on electromyography. Muscle biopsy in one patient revealed CD8+ polymyositis, and in the other showed ddI induced myopathy. These cases illustrate the importance of muscle biopsy in identifying the underlying pathology in HIV infected patients with muscle weakness and little or no abnormality in laboratory investigations. PMID- 15295134 TI - Impact of measurement error in the study of sexually transmitted infections. AB - Measurement is a fundamental part of all scientific research, and the introduction of errors of different sorts is an inevitable part of the measurement process in epidemiological and clinical research. Despite the ubiquity of measurement error in research, the substantial impacts which measurement error can have on data and subsequent study inferences are frequently overlooked. This review introduces the basic concepts of measurement error that are most relevant to the study of sexually transmitted infections, and demonstrates the impacts of several of the most common forms of measurement error on study results. A self assessment test and MCQs follow this paper. PMID- 15295136 TI - HIV transmission among men who have sex with men through oral sex. PMID- 15295137 TI - The correct approach to modelling and evaluating chlamydia screening. PMID- 15295138 TI - Haryana state in India, still a low HIV prevalence state. PMID- 15295139 TI - Increased numbers of acute hepatitis C infections in HIV positive homosexual men; is sexual transmission feeding the increase? PMID- 15295140 TI - Transmission of Neisseria gonorrhoeae from a toilet seat. PMID- 15295141 TI - Detection of Chlamydia trachomatis by polymerase chain reaction in male patients with non-gonococcal urethritis attending an STD clinic. PMID- 15295143 TI - Cybertherapy in practice: the VEPSY updated project. AB - Rapid and far-reaching technological advances are changing the ways in which people relate, communicate, and live. Technologies that were hardly used ten years ago, such as the Internet, e-mail, and video teleconferencing are becoming familiar methods for diagnosis, therapy, education and training. This is producing an emerging field (cybertherapy) whose focus is the use of communication and information technologies to improve the health care processes. To exploit and understand this potential was the aim of the Telemedicine and Portable Virtual Environment in Clinical Psychology, VEPSY UPDATED, an European Community funded research project (IST-2000-25323, http://www.cybertherapy.info). The chapter describes the clinical and technical rationale behind the cybertherapy applications developed by the project. Further, the actual role of virtual reality in the cybertherapy field is discussed, focusing on the advantages provided by its three different faces: technological, experiential and communicative. PMID- 15295144 TI - New tools in cybertherapy: the VEPSY web site. AB - In the last years the rapid development of the Internet and new communication technologies has had a great impact on psychology and psychotherapy. Psychotherapists seem to rely with more and more interest on the new technological tools such as videophone, audio and video chat, e-mail, SMS and the new Instant Messaging Tools (IMs). All these technologies outline a stimulating as well as complex scenario: in order to effectively exploit their potential, it is important to study which is the possible role played by the Internet-based tools inside a psychotherapeutic iter. Could the technology substitute the health care practitioners or are these tools only a resource in addition to the traditional ones in the therapist's hand? The major aim of this chapter is to provide a framework for the integration of old and new tools in mental health care. Different theoretical positions about the possible role played by e-therapy are reported showing the possible changes that psychotherapy will necessarily face in a cyber setting. The VEPSY website, an integration of different Internet based tools developed within the VEPSY UPDATED Project, is described as an example of clinical application matching between old (and functional) practices with new (and promising) media for the treatment of different mental disorders. A rationale about the possible scenarios for the use of the VEPSY website in the clinical process is provided. PMID- 15295145 TI - Virtual reality and psychotherapy. AB - Virtual Reality (VR) is a new technology consisting on a graphic environment in which the user, not only has the feeling of being physically present in a virtual world, but he/she can interact with it. The first VR workstations were designed for big companies in order to create environments that simulate certain situations to train professionals. However, at this moment a great expansion of this technology is taking place in several fields, including the area of health. Especially interesting for us is the use of VR as a therapeutic tool in the treatment of psychological disorders. Compared to the traditional treatments, VR has many advantages (e.g., it is a protected environment for the patient, he/she can re-experience many times the feared situation, etc.). There are already data on the effectiveness of this technology in the treatment of different psychological disorders; here anxiety disorders, eating disorders and sexual disorders are reviewed. Finally, this chapter ends with some words about the limitations of VR and future perspectives. PMID- 15295146 TI - Virtual interaction in cognitive neuropsychology. AB - Several recent studies have investigated whether knowledge representation turns possible within virtual reality simulated environments. According to these affirmative results different clinical applications were developed in psychology. Among these applications virtual reality seems to have a specific role in assessment and treatment of neuropsychological diseases. This chapter will firstly investigate possibilities and challenges carried from virtual-reality based neuropsychological applications focusing both on patient's and therapist's point of view. Afterward it will provide a survey of research and intervention application examples. More in detail a clear explanation of contribution goals will be discussed, in order to place research and applied works within a cognitive neuroscience frame of reference, according with their usefulness and effectiveness in clinical treatment. Fulfilling these objectives neuropsychological virtual reality approaches in memory, motor abilities, executive functions and spatial representation will be shown. PMID- 15295147 TI - The use of VR in the treatment of panic disorders and agoraphobia. AB - Panic disorder with agoraphobia (PDA) is considered an important public health problem. The efficacy of cognitive-behavioral therapy (CBT) for PDA has been widely demonstrated. The American National Institute of Health recommended Cognitive-Behavioral programs as the treatment of choice for this disorder. This institution also recommended that researchers develop treatments whose mode of delivery increases the availability of these programs. Virtual Reality based treatments can help to achieve this goal. VR has several advantages compared with conventional techniques. One of the essential components to treat these disorders is exposure. In VR the therapist can control the feared situations at will and with a high degree of safety for the patient, as it is easier to grade the feared situations. Another advantage is that VR is more confidential because treatment takes place in the therapist's office. It is also less time consuming as it takes place in the therapist's office. Considering the wide number of situations and activities that agoraphobic patients use to avoid, VR can save time and money significantly. Another advantage in treating PDA using VR is the possibility of doing VR interoceptive. VR could be a more natural setting for interoceptive exposure than the consultation room because we can elicit bodily sensations while the patient is immerse in VR agoraphobic situations. Finally, we think that VR exposure can be a useful intermediate step for those patients who refuse in vivo exposure because the idea of facing the real agoraphobic situations is too aversive for them. In this chapter we offer the work done by our research team at the VEPSY-UPDATED project. We describe the VR program we have developed for the treatment of PDA and we summarize the efficacy and effectiveness data of a study where we compare a cognitive-behavioral program including VR for the exposure component with a standard cognitive-behavioral program including in vivo exposure and with a waiting list control condition. Our findings support the efficacy and effectiveness of VR for the treatment of PDA. PMID- 15295148 TI - Virtual reality exposure in the treatment of social phobia. AB - Social phobia is one of the most frequent psychiatric disorders and is accessible to two forms of scientifically validated treatments: anti-depressant drugs and cognitive-behavioral therapies. Graded exposure to feared social situations (either in vivo or by imagining the situations) is fundamental to obtain an improvement of the anxious symptoms. Virtual reality (VR) may be an alternative to these standard exposure techniques and seems to bring significant advantages by allowing exposures to numerous and varied situations. Moreover studies have shown that human subjects are appropriately sensitive to virtual environments. This chapter reports the definition of a VR-based clinical protocol and a study to treat social phobia using virtual reality techniques. The virtual environments used in the treatment reproduce four situations that social phobics feel the most threatening: performance, intimacy, scrutiny and assertiveness. With the help of the therapist, the patient learns adapted cognitions and behaviors when coping with social situations, with the aim of reducing her or his anxiety in the corresponding real life situations. Some studies have been carried out using virtual reality in the treatment of fear of public speaking, which is only a small part of the symptomatology of most of social phobic patients. The novelty of our work is to address a larger group of situations that the phobic patients experience with high anxiety. In our protocol, the efficacy of the virtual reality treatment is compared to well established and well validated group cognitive-behavioral treatment. PMID- 15295149 TI - The use of VR in the treatment of eating disorders. AB - In the treatment of eating disorders, the cognitive behavioral therapy (CBT) is still considered the best approach but could present different limitations related to costs of behavioural procedures (such as exposure and desensitization) or difficulty of cognitive techniques (such as imagination of daily scenarios). The major aim of this contribution is the description of a new Virtual Reality enhanced treatment named Experiential Cognitive Therapy (ECT). Rationale and protocols about this new approach are explained. Moreover data about clinical trials, carried on with the VEPSY Project, are shown comparing different groups: experimental group (ECT), cognitive-behavioural therapy group (CBT), nutritional group and control group. PMID- 15295150 TI - Male sexual dysfunctions: immersive virtual reality and multimedia therapy. AB - The study describes a therapeutic approach using psycho-dynamic psychotherapy integrating virtual environment (VE) for resolving impotence or better erectile dysfunction (ED) of presumably psychological or mixed origin and premature ejaculation (PE). The plan for therapy consists of 12 sessions (15 if a sexual partner was involved) over a 25-week period on the ontogenetic development of male sexual identity, and the methods involved the use of a laptop PC, joystick, Virtual Reality (VR) helmet with miniature television screen showing a new specially-designed CD-ROM programs using Virtools with Windows 2000 and an audio CD. This study was composed of 30 patients, 15 (10 suffering from ED and 5 PE) plus 15 control patients (10 ED and 5 PE), that underwent the same therapeutic protocol but used an old VR helmet to interact with the old VE using a PC Pentium 133 16 Mb RAM. We also compared this study with another study we carried out on 160 men affected by sexual disorders, underwent the same therapeutic protocol, but treated using a VE created (in Superscape VRT 5.6) using always Windows 2000 with portable tools. Comparing the groups of patients affected by ED and PE, there emerged a significant positive results value without any important differences among the different VE used. However, we had a % increase of undesirable physical reactions during the more realistic 15-minute VR experience using Virtools development kit. Psychotherapy alone normally requires long periods of treatment in order to resolve sexual dysfunctions. Considering the particular way in which full-immersion VR involves the subject who experiences it (he is totally unobserved and in complete privacy), we hypothesise that this methodological approach might speed up the therapeutic psycho-dynamic process, which eludes cognitive defences and directly stimulates the subconscious, and that better results could be obtained in the treatment of these sexual disorders. This method can be used by any psychotherapist and it can be used alone or associated with pharmacotherapy prescribed by the urologist/andrologist as part of a therapeutic alliance. PMID- 15295151 TI - New technologies for providing remote psychological treatments. AB - New technologies lead us to a series of new applications that we could not imagine just a few years before. Many services have appeared for Internet, the global computer network: FTP, e-mail, World Wide Web. Psychological treatments are one of the multiple applications that can be developed using these tools. Dynamic web pages that include information prepared by the therapist for different patients and that receive information from them can be generated. Other tools such as e-mail or chats can be used to provide a direct communication. Databases can be integrated in web applications for storing data about different patients. Several formats can be used for storing the information, and some of them such as XML provide a promising method of psychological data standardization. Using different development tools, virtual environments can also be generated and integrated in web pages, so new psychological treatments such as virtual environment exposure are also possible from web applications. This entire basis provides the structure that allows that new applications can be imagined and developed. In a few years, new trends will appear, probably one of them will be the use of wireless devices to provide psychological treatment and help at any place and any time. PMID- 15295152 TI - Technological background of VR. AB - User interface (UI) design is a critical component of any virtual environment (VE) application, and especially for VE applied to medicine. User interfaces for VE are becoming more diverse. Mice, keyboards, windows, menus, and icons--the standard parts of traditional WIMP interfaces--are still prevalent, but nontraditional devices and interface components are proliferating rapidly. These include spatial input devices such as trackers, 3-D pointing devices, and whole hand devices allowing gestural input. Three-dimensional, multisensory output technologies--such as stereoscopic projection displays, head-mounted displays (HMDs), spatial audio systems, and haptic devices--are also becoming more common. In this chapter we present a brief overview of 3-D interaction and user interfaces technologies for VE. PMID- 15295153 TI - Ergonomics of virtual environments for clinical use. AB - Usability is defined by the International Standards Organization as "the effectiveness, efficiency and satisfaction with which a certain user may achieve a specific objective in a particular environment" (ISO DIS 9241-11). This definition highlights the need for considering the specific destination of a certain technology and reflects the current trends in Human-Computer Interaction (HCI). In compliance with these remarks, the evaluation will be described here of a Virtual Environment (VE) for the treatment of male sexual dysfunctions; the common assumption will be avoided according to which a VE is a space separated from its "real" surroundings [6, 7] and the full environment where the Virtual session takes place will be addressed instead. After a description of the conceptual framework adopted, the paper will dwell on one method among those deployed for the evaluation, namely the analysis of "situated actions". Four aspects will be dealt with: (a) the interplay of various concurrent settings during the virtual session; (b) the users' comprehension of the symbols used in the VE; (c) the structure of the relationship between users and guide; (d) the breakdowns during the human-VE interaction. The goals and the intended meanings of the simulation as set by the designers became the main parameters for the evaluation. PMID- 15295154 TI - An integrated approach to the ergonomic analysis of VR in psychotherapy: panic disorders, agoraphobia and eating disorders. AB - To face the aspects connected with VR environments usability for psychotherapeutic applications means to dare a double challenge from a methodological point of view: from one side, the need to adapt and to integrate on a heuristic basis classic usability evaluation methods to specific artefacts such as 3D Virtual Environments for clinical applications; from the other hand, the problems arisen by integration of expert evaluation of VR environments user based tests carried out in real context of use. The theoretical background of our analytical stance is based upon an ethnomethodological approach, a perspective that gives evidence of how people, in specific social situations, are able to solve complex tasks producing shared meanings and achieving their goals during interaction. According to this perspective, the methodological objective consisted also in the identification of the usability requirements of the specific community of practice. The virtual environments considered were two of the four VR modules in the framework of the VEPSY project: Panic Disorders- Agoraphobia and Eating Disorders. PMID- 15295155 TI - Immersive Virtual Telepresence: virtual reality meets eHealth. AB - Immersive Virtual Telepresence (IVT) tools are virtual reality environments combined with wireless multimedia facilities--real-time video and audio--and advanced input devices--tracking sensors, biosensors, brain-computer interfaces. For its features IVT can be considered an innovative communication interface based on interactive 3D visualization, able to collect and integrate different inputs and data sets in a single real-like experience. In this paper we try to outline the current state of research and technology that is relevant to the development of IVT in medicine. Moreover, we discuss the clinical principles and possible advantages associated with the use of IVT in this field. PMID- 15295156 TI - The future of cybertherapy: improved options with advanced technologies. AB - Cybertherapy is a field that is growing rapidly due to today's technology and information boom. Virtual reality and advanced technologies have been used successfully to in a variety of healthcare issues, including treatment of anxiety disorders and phobias, treatment of eating and body dysmorphic disorders, neuropsychological assessment and rehabilitation and distraction during painful or unpleasant medical procedures. The novel applications of these technologies yield many advantages over traditional treatment modalities, and the disadvantages that accompanied the first trials of virtual reality are quickly being addressed and eliminated. Virtual reality peripherals such as data gloves, physiological monitoring and Internet worlds are swiftly demonstrating their usefulness in cybertherapy applications. Future directions for research include improvements of objective measures of efficacy such as fMRI and physiological monitoring devices, and investigations are being carried out to determine if virtual reality and advanced technologies can be used to treat a broader scope of disorders, including depression, schizophrenia, drug addiction, and autism. PMID- 15295159 TI - Cerium iron sulfide, Ce3Fe1.94S7. PMID- 15295160 TI - Lithium and beryllium hypophosphites. PMID- 15295161 TI - [Beta-LiMoO2(AsO4)]. PMID- 15295162 TI - catena-Poly[[diaquabis(2-chloronicotinato-kappa2O,O')cadmium(II)]-mu-2 chloronicotinato-kappa3O,O':N]. PMID- 15295163 TI - fac-Tricarbonyl(4,4'-dimethyl-2,2'-bipyridine)triphenyltin(II)rhenium(I). PMID- 15295164 TI - Catena-poly[[[cis-bis(1-phenyltetrazole-kappaN4)copper(II)]-di-mu-chloro] hemi(1 phenyltetrazole)]. PMID- 15295165 TI - Catena-poly[[[aquacopper(I)]-mu-4,4'-bipyridyl-kappa2N:N'] hemi(isophthalate) monohydrate]. PMID- 15295166 TI - A novel two-dimensional Mn(II) coordination polymer with 1,2-bis(imidazol-1 yl)ethane. PMID- 15295167 TI - Three zinc(II) complexes presenting a ZnN6 chromophore and with peroxodisulfate as the counter-ion. PMID- 15295168 TI - Bis(acetato-kappa2O,O')(2,9-dimethyl-1,10-phenanthroline-kappa2N,N')mercury(II) in two differently hydrated crystal forms. PMID- 15295169 TI - Triferrocenylphosphine oxide monohydrate. PMID- 15295170 TI - Catena-poly[[diaqua(nicotinato-kappa2O,O')cadmium(II)]-mu-nicotinato kappa3N:O,O']. PMID- 15295171 TI - A capped trigonal prismatic cadmium complex with tetra- and tridentate ligands: bis(triethanolamine)-kappa3N,O,O';kappa4N,O,O',O"-cadmium(II) squarate monohydrate. PMID- 15295172 TI - Intramolecular hydrogen-bond-directed coordination: trans-bis(N-benzoyl-N' propylthiourea-kappaS)diiodoplatinum(II) and trans-bis(N-benzoyl-N' propylthiourea-kappaS)dibromoplatinum(II). PMID- 15295173 TI - Hexa-mu2-chloro-mu4-oxo-tetrakis[(2-ethyltetrazole-kappaN4)copper(II)]. PMID- 15295174 TI - Trans-bis[N-(2-hydroxyethyl)ethylenediamine-kappa2N,N']bis(isothiocyanato kappaN)nickel(II). PMID- 15295175 TI - Chloro(phenyl)(1H-pyrazole-kappaN2)(triphenylphosphine-kappaP)palladium(II). PMID- 15295176 TI - The first suberate lanthanum(III) complex without uncoordinated water. PMID- 15295177 TI - A compound of a novel tetraaza-macrocycle with trinuclear tetracyanonickelate bridged cations. PMID- 15295178 TI - catena-Poly[bromo(omega-thiocaprolactam-kappaS)gold(I)](Au-Au). PMID- 15295179 TI - Hydrate isomerism in [Cu(en)2(H2O)1.935]2[Fe(CN)6].4H2O. PMID- 15295180 TI - 4-[N,N-Bis(2-hydroxyimino-2-phenylethyl)amino]-1,5-dimethyl-2-phenyl-2,3-dihydro 1H-pyrazol-3-one monohydrate. PMID- 15295181 TI - 4-Phenyl-1H-imidazole (a low-temperature redetermination), 1-benzyl-1H-imidazole and 1-mesityl-1H-imidazole. PMID- 15295182 TI - Tetramethylammonium pentaborate 0.25-hydrate. PMID- 15295183 TI - 1-(2,5-Dichlorophenyl)-3-methyl-5-phenyl-1H-pyrazole. PMID- 15295184 TI - (E)-N-[(Benzo[b]thiophen-3-yl)methylene]-N'-(2,5-dichlorophenyl)hydrazine. PMID- 15295185 TI - p-Chloro-, p-bromo- and two polymorphs of p-iodoacetophenone. PMID- 15295186 TI - Redetermination of triethylammonium chloride in the space group P31c. PMID- 15295187 TI - 5-Acetyl-2-amino-6-methyl-4-(1-naphthyl)-4H-pyran-3-carbonitrile, methyl 6-amino 5-cyano-2-methyl-4-(1-naphthyl)-4H-pyran-3-carboxylate and tert-butyl 6-amino-5 cyano-2-methyl-4-(1-naphthyl)-4H-pyran-3-carboxylate. PMID- 15295188 TI - A non-planar peptide bond in L-seryl-L-valine. PMID- 15295189 TI - 1,N6-Etheno derivative of 7-deaza-2,8-diazaadenosine. PMID- 15295190 TI - Non-mutagenic organic pigment intermediates. II. Isomorphous 2,2'-dichloro-5,5' dipropoxybenzidine and 2,2'-dimethyl-5,5'-dipropoxybenzidine. PMID- 15295191 TI - S-[4-(Trimethylammonio)phenyl]thiosulfate, an aromatic organic thiosulfate. PMID- 15295192 TI - Hydrogen bonding in proton-transfer compounds of 5-sulfosalicylic acid with bicyclic heteroaromatic Lewis bases. PMID- 15295193 TI - 2-(6-Bromopyridin-2-yl)-6-methyl-[1,3,6,2]dioxazaborocane, a new stable (pyridin 2-yl)boronic acid derivative. PMID- 15295194 TI - Picolinamidium squarate and di-p-toluidinium squarate dihydrate. PMID- 15295195 TI - A supramolecular ladder motif in 2-(2,2,6,6-tetramethylpiperidin-1-yloxy)propane 1,3-diol. PMID- 15295196 TI - Ethyl 2-amino-4-tert-butyl-1,3-thiazole-5-carboxylate and 6-methylimidazo[2,1 b]thiazole-2-amino-1,3-thiazole (1/1). PMID- 15295197 TI - Dimethyl 3,4,5,5-tetraphenyl-1,3-thiazolidine-2,2-dicarboxylate and 3,3-dichloro 2,2,4,4,3'-pentamethyl-r-2',t-4'-diphenylcyclobutane-1-spiro-5'-1,3-thiazolidine. PMID- 15295198 TI - Hydrogen bonding in proton-transfer compounds of 8-quinolinol (oxine) with aromatic sulfonic acids. PMID- 15295199 TI - 10-(4-Chlorophenyl)-7-methyl-5,6-dihydrobenzo[h]pyrazolo[5,1-b]quinazoline and 2 (4-chlorophenyl)-5-methyl-6,7-dihydrobenzo[h]pyrazolo[1,5-a]quinazoline: isomeric molecules linked into hydrogen-bonded dimers or pi-stacked chains. PMID- 15295200 TI - Curcumin and derivatives. PMID- 15295201 TI - Hydrogen-bonding and C-H...pi interactions in 1,7-bis(4-hydroxy-3 methoxyphenyl)heptane-3,5-dione (tetrahydrocurcumin). PMID- 15295202 TI - 4-(3,4-Dichlorophenyl)-9-(4-methylphenylsulfonyl)-4,9 diazatetracyclo[5.3.1.0(2,6).0(8,10)]undecane-3,5-dione and 4-(4-chlorophenyl)-9 (4-methylphenylsulfonyl)-4,9-diazatetracyclo[5.3.1.0(2,6).0(8,10)]undecane-3,5 dione. PMID- 15295203 TI - Supramolecular structures of three configurational isomers of 1 phenylethanaminium malate(1-). PMID- 15295204 TI - Serotonin1A receptor agonist acquires an antimalarial connection. PMID- 15295205 TI - A gene involved in nematode feeding behaviour. PMID- 15295206 TI - What impact, if any, has feminism had on science? PMID- 15295207 TI - How to design a highly effective siRNA. PMID- 15295208 TI - The enigma of carcinogenesis - stroma or epithelial cells? PMID- 15295209 TI - Bhattacharyya's distance measure as a precursor of genetic distance measures. PMID- 15295210 TI - John Maynard Smith: 6 January 1920-19 April 2004. PMID- 15295211 TI - Exploring germ-soma differentiation in Volvox. PMID- 15295217 TI - Comprehensive pediatric human immunodeficiency virus care and treatment in Constanta, Romania: implementation of a program of highly active antiretroviral therapy in a resource-poor setting. AB - BACKGROUND: Relatively few human immunodeficiency virus (HIV)-infected children worldwide have access to care and treatment. The Romanian-American Children's Center, a collaborative project of a U.S. health care institution and the Romanian government, has established a comprehensive program of highly active antiretroviral therapy for children in Constanta, Romania. OBJECTIVES: To describe the design and outcomes of a program of pediatric HIV/acquired immunodeficiency syndrome (AIDS) care and treatment in a resource-poor setting. SETTING: Outpatient center providing comprehensive primary and HIV/AIDS specialty care and treatment to all known HIV-infected children living in Constanta County, Romania. OUTCOMES: As of August 2003, a total of 452 children were receiving highly active antiretroviral therapy. Therapy has been well-tolerated, with approximately 90% of children continuing to receive treatment after a median duration of follow-up of 67 weeks. Normal weight and height growth velocities have been observed among treated children. Marked decreases have been observed in rates of hospitalization and mortality. The mean change in CD4+ lymphocyte count for 173 children who have both a baseline count and at least 1 follow-up count is +284 cells/microL (P < 0.0001). CONCLUSIONS: Highly active antiretroviral therapy can be administered safely and effectively to children in a resource-poor setting, with outcomes comparable with those observed in U.S. pediatric antiretroviral clinical trials. PMID- 15295218 TI - Community-acquired, methicillin-resistant and methicillin-susceptible Staphylococcus aureus musculoskeletal infections in children. AB - BACKGROUND: The clinical characteristics and virulence factors related to musculoskeletal infections caused by community-acquired, methicillin-resistant Staphylococcus aureus (MRSA) in children are not well-defined. METHODS: In this retrospective study, the demographics, hospital course and outcome of children with musculoskeletal infections were reviewed from medical records and by contacting patients or their physicians. Antimicrobial susceptibilities were determined by disk diffusion. Polymerase chain reaction was performed to detect genes encoding for virulence factors. Mann-Whitney, chi2 and Kaplan-Meier tests were used for statistical analysis. RESULTS: Community-acquired MRSA and community-acquired methicillin-susceptible S. aureus (MSSA) caused musculoskeletal infections in 31 and 28 children, respectively. The median numbers of febrile days after start of therapy were 4 and 1 for MRSA and MSSA patients, respectively (P = 0.001). The median numbers of hospital days were 13 and 8 for the MRSA and MSSA groups, respectively (P = 0.014). At follow-up, 2 patients in the MRSA and 1 in the MSSA group had developed chronic osteomyelitis. pvl and fnbB genes were found in 87 and 90% versus 24 and 64% in the MRSA versus MSSA groups, respectively. (P = 0.00001 and 0.017). Ten patients with pvl positive strains had complications versus no patients with pvl-negative isolates (P = 0.002). CONCLUSIONS: Febrile days and hospital days were greater in children with musculoskeletal infection caused by MRSA than in those affected by MSSA, but no significant differences were found in the final outcome. pvl and fnbB genes were more frequent in the MRSA than in the MSSA strains. The presence of the pvl gene may be related to an increased likelihood of complications in children with S. aureus musculoskeletal infections. PMID- 15295219 TI - Safety and pharmacokinetics of palivizumab therapy in children hospitalized with respiratory syncytial virus infection. AB - BACKGROUND: Respiratory syncytial virus (RSV) infection represents a major cause of pediatric respiratory hospitalizations. Limited treatment options exist. Palivizumab is a humanized monoclonal IgG1 antibody to the fusion protein of RSV that is highly active against RSV A and B strains. METHODS: A phase I/II, multicenter, randomized, double-blind, placebo-controlled, escalating dose clinical trial to describe the safety, tolerance, pharmacokinetics and clinical outcome of a single intravenous dose of palivizumab in previously healthy children hospitalized with acute RSV infection. RESULTS: Fifty-nine children < or =2 years of age received study drug. Sixteen children received 5 mg/kg of palivizumab (n = 8) or placebo (n = 8); 43 received 15 mg/kg of palivizumab (n = 22) or placebo (n = 21). Adverse events judged to be related to study drug were seen in one 5-mg/kg palivizumab patient and one 15-mg/kg palivizumab patient. These events were transient or consistent with progression of RSV disease. No discontinuations of study drug infusion because of adverse events occurred. Mean serum concentrations of palivizumab in the 5- and 15-mg/kg groups, respectively, were 61.2 and 303.4 microg/mL at 60 min and 11.2 and 38.4 microg/mL after 30 days. There were no significant differences in clinical outcomes between placebo and palivizumab groups for either dose. CONCLUSIONS: Intravenous palivizumab was safe and well-tolerated in children hospitalized with RSV disease. A single 15 mg/kg dose achieved serum palivizumab concentrations above the 25- to 30 microg/mL concentration associated with 2-log reduction of pulmonary RSV titer in the cotton rat model. PMID- 15295220 TI - Long term tolerability and safety of enfuvirtide for human immunodeficiency virus 1-infected children. AB - BACKGROUND: Enfuvirtide, a peptide inhibitor of human immunodeficiency virus (HIV)-1-host cell membrane fusion, is the first of a new class of antiretroviral agents, the entry inhibitors. The safety and antiretroviral activity of enfuvirtide treatment of 24 weeks in HIV-1-infected children has been previously documented. Here we present the long term tolerability and safety of enfuvirtide. METHODS: Fourteen children, 4 to 12 years of age, with incompletely suppressed HIV-1 infection were evaluated. Enfuvirtide was administered twice daily by subcutaneous injection. After the first 24 weeks of enfuvirtide dosing, subjects were evaluated every 8 weeks up to 96 weeks of therapy. At each visit, each subject had a physical examination and an assessment for adverse events with particular attention to evaluation of injection site reactions. Laboratory studies obtained at each visit included hematology and blood chemistry values, plasma HIV-1 RNA concentrations and CD4+ T cell counts. RESULTS: Of 14 subjects, 6 completed at least 96 weeks of treatment. One child discontinued enfuvirtide after 22 days of treatment because of an aversion to injections, and 1 child electively discontinued after week 24 because of surgical complications unrelated to study drug. Four subjects discontinued study because of virologic failure, defined as an increase or persistence of plasma HIV-1 RNA 1.0 copies/mL above baseline, which occurred between >or =log10 weeks 40 and 63. Two children experienced grade 3 adverse events resulting in discontinuation of the study drug; 1 subject developed grade 3 thrombocytopenia and 1 developed grade 3 edema at weeks 65 and 77, respectively. Eleven of 14 children had local injection site reactions during the first 24 weeks of treatment, 4 of the 12 subjects who continued treatment beyond week 24 reported local reactions. Generally, these local reactions were 1- to 3-cm tender nodules that developed after the injections and lasted for 1-2 days. Twelve children developed new diagnoses during treatment with enfuvirtide. None was judged to be definitively related to the study drug. Thirty-six percent of children starting enfuvirtide had HIV-1 RNA levels > 1 log10 copies/mL below baseline levels at week 96. Children remaining on enfuvirtide for the entire 96 weeks had a median of 65 cells/mm and 9% increase in CD4+ T cells. CONCLUSIONS: Enfuvirtide was generally safe and, except for a high rate of injection site reactions, well-tolerated in HIV-1-infected children for as long as 96 weeks. PMID- 15295221 TI - Topically applied sunflower seed oil prevents invasive bacterial infections in preterm infants in Egypt: a randomized, controlled clinical trial. AB - BACKGROUND: Because the therapeutic options for managing infections in neonates in developing countries are often limited, innovative approaches to preventing infections are needed. Topical therapy with skin barrier-enhancing products may be an effective strategy for improving neonatal outcomes, particularly among preterm, low birth weight infants whose skin barrier is temporarily but critically compromised as a result of immaturity. METHODS: We tested the impact of topical application of sunflower seed oil 3 times daily to preterm infants <34 weeks gestational age at the Kasr El-Aini neonatal intensive care unit at Cairo University on skin condition, rates of nosocomial infections and mortality. RESULTS: Treatment with sunflower seed oil (n = 51) resulted in a significant improvement in skin condition (P = 0.037) and a highly significant reduction in the incidence of nosocomial infections (adjusted incidence ratio, 0.46; 95% confidence interval, 0.26-0.81; P = 0.007) compared with infants not receiving topical prophylaxis (n = 52). There were no reported adverse events as a result of topical therapy. CONCLUSIONS: Given the low cost (approximately .20 dollars for a course of therapy) and technologic simplicity of the intervention and the effect size observed in this study, a clinical trial with increased numbers of subjects is indicated to evaluate the potential of topical therapy to reduce infections and save newborn lives in developing countries. PMID- 15295222 TI - Elimination of racial differences in invasive pneumococcal disease in young children after introduction of the conjugate pneumococcal vaccine. AB - BACKGROUND: Racial differences in the epidemiology of invasive pneumococcal disease (IPD) have been widely recognized, but the impact of conjugate pneumococcal vaccine (PCV) introduction in 2000 on these differences has not been extensively studied. METHODS: IPD episodes in 5 Tennessee counties from January 1995 through December 2002 were collected prospectively using the Centers for Disease Control and Prevention's Active Bacterial Core Surveillance system (ABCs). Trained nurses collected clinical data, and antibiotic susceptibility testing was performed on available isolates. RESULTS: Before vaccine licensure, IPD rates were highest in children younger than 2 years and in blacks. The disparity in IPD rates between blacks and whites younger than 2 years of age substantially diminished after PCV introduction. In 1999, the IPD rate in black children younger than 2 years was 340.2 per 100,000, representing 176.5 more events per 100,000 than in white children (P < 0.001). In 2002, this rate had decreased 83% to 57.4 per 100,000, similar to the rate in white children (39.6 per 100,000; P = 0.31). Before vaccine licensure, a higher percentage of isolates from whites were antibiotic-nonsusceptible. In 2002, the proportion of antibiotic nonsusceptible pneumococcal isolates was similar in whites and blacks of all ages for the first time during the study period (P > 0.05 for separate comparisons of penicillin, cephalosporin and erythromycin nonsusceptibility). These changes occurred despite a lower PCV vaccination coverage in Tennessee in blacks than in whites (31.2% versus 47.6%). CONCLUSIONS: With conjugate pneumococcal vaccine introduction in Tennessee, racial differences in the incidence rates of IPD have largely been eliminated, particularly in young children. PMID- 15295223 TI - The seven-valent pneumococcal conjugate vaccine reduces tympanostomy tube placement in children. AB - BACKGROUND: The novel pneumococcal conjugate vaccine, PncCRM, has been shown to prevent acute otitis media caused by vaccine serotypes and to reduce otitis surgery. Our aim was to assess long term efficacy of the vaccine on tympanostomy tube placements. METHODS: Children with complete follow-up in the Finnish Otitis Media Vaccine Trial up to 24 months of age and still living in the study area (1490 of 1662 randomized at 2 months of age) were invited to a single visit at 4 5 years of age. The children had been vaccinated at 2, 4, 6 and 12 months of age with PncCRM or hepatitis B vaccine (control). Tympanostomy tube placements reported by parents at the visit were verified from hospital and private medical center records. Additionally, tympanostomy tube placements of all children were verified from the hospital discharge registry. Vaccine efficacy (VE) was estimated by comparing all events of tympanostomy tube placement between vaccine groups. RESULTS: During the vaccine trial (2-24 months of age), VE (95% confidence interval) in preventing tympanostomy tube placement was only 4% (-19 23%). Altogether 756 children were enrolled for the follow-up study. After 24 months of age, the rate of surgery was 3.5 per 100 person-years in the PncCRM and 5.7 per 100 person-years in the control children, giving VE of 39% (4-61%). In the hospital-based data of all children (N = 1490), VE of 44% was obtained (19 62%). CONCLUSIONS: Receipt of PncCRM vaccine at infancy was associated with a reduction in tympanostomy tube placement from 2 to 4-5 years of age. PMID- 15295224 TI - Improvements in nutritional management as a determinant of reduced mortality from community-acquired lower respiratory tract infection in hospitalized children from rural central Africa. AB - BACKGROUND: In-hospital mortality from lower respiratory tract infections (LTRI) is unacceptably high in developing countries where LTRI are still a leading cause of death. OBJECTIVE: To identify new approaches to reduce in-hospital mortality of LRTI through the improvement of its management. METHODS: The prospectively collected database of children admitted during an 11-year period with LRTI in a pediatric rural hospital in Central Africa was reviewed to determine the predictors of death and to evaluate the impact on mortality of 4 different protocols for the management of malnutrition. RESULTS: During the study period, 859 children were admitted with a nonmeasles severe LRTI. In the 3-year period during which blood cultures were obtained, 29.0% of the children with LRTI were bacteremic, and multiresistant Enterobacteriaceae were recovered in 81.4% of positive blood cultures. Independent predictors of death in children without edema were age <24 months, dehydration and hepatomegaly with adjusted odds ratios (numbers in parentheses, 95% confidence interval) of 3.47 (1.70-7.08), 4.24 (2.11 8.50) and 2.90 (1.43-5.85), respectively. In those with edema, a significantly increased risk of death was noted for girls [4.31 (1.71-10.90)], in children with z-score of weight to height < or = -3 [5.45 (1.67-17.79)] and when the serum albumin was <16 g/l [2.58 (1.01-6.58)]. The improvement in the management of malnutrition was followed by a reduction of LRTI-related mortality in children with edema from 32.4 to 8.9% (P < 0.001). In children without edema, the LRTI related mortality decreased from approximately 12% to 3.5% when their diet was supplemented with micronutrients. CONCLUSION: This study indicates that the improvement of the management of underlying nutritional deficiencies is crucial for the reduction of the high in-hospital case fatality rate associated with severe nonmeasles LRTI. The empiric antibiotic regimen should be modified to cover for multiresistant Enterobacteriaceae. PMID- 15295225 TI - Serum group a anticapsular antibodies in a Sudanese population immunized with meningococcal polysaccharide vaccine during a group A epidemic. AB - BACKGROUND: Vaccination during group A meningococcal epidemics is reported to decrease the number of new cases of disease. However, implementing mass vaccination is often delayed, and little information is available on whether immunity increases in a population vaccinated during an epidemic when exposure to the epidemic strain is common. METHODS: A convenience sample of 134 previously unimmunized persons, ages 3-49 years, were immunized with a meningococcal polysaccharide vaccine. Their serum group A antibody responses were compared with those of 26 adults immunized in California with no known group A exposure. RESULTS: Before immunization, serum anticapsular antibody concentrations were 10 fold higher in Sudanese adults and 4-fold higher in Sudanese, ages 3-17 years, than in North American adults (geometric means, 29 and 13 microg/ml, respectively, versus 3 microg/ml, P < 0.001). Seventy-five percent of the Sudanese had serum bactericidal titers that correlate with protection (> or 1/128). Nearly all Sudanese with low bactericidal titers before vaccination developed protective bactericidal antibody responses after vaccination, and the magnitude of the anticapsular antibody responses of the Sudanese was similar to that of the immunized North Americans. INTERPRETATION: The high titers of naturally acquired antibody in the Sudanese may reflect widespread exposure to the epidemic strain and underscore difficulties of instituting immunization before exposure occurs. Also epidemics in sub-Saharan Africans may not abate even if 75% of the population is immune to disease as long as the organism is transmitted widely among both immune and susceptible persons. PMID- 15295226 TI - Acute disseminated encephalomyelitis in childhood: epidemiologic, clinical and laboratory features. AB - BACKGROUND: Acute disseminated encephalomyelitis (ADEM) is a central nervous system demyelinating disease that usually follows an apparently benign infection in otherwise healthy young persons. The epidemiology, infectious antecedents and pathogenesis of ADEM are poorly characterized, and some ADEM patients are subsequently diagnosed with multiple sclerosis (MS). METHODS: We retrospectively (1991-1998) and prospectively (1998-2000) studied all persons aged < 20 years diagnosed with ADEM from the 3 principal pediatric hospitals in San Diego County, CA, during 1991-2000. Acute neurologic abnormalities and imaging evidence of demyelination were required for study inclusion. Epidemiologic variables, risk factors, clinical course, laboratory and radiographic findings, neuropathology and treatment data were analyzed. Interleukin (IL)-12, interferon-gamma (IFN gamma) and IL-10 were assayed in blinded manner on cerebrospinal fluid (CSF) obtained prospectively from a subset of ADEM cases and compared with CSF from patients with enteroviral (EV) meningoencephalitis confirmed by polymerase chain reaction (PCR) and controls without pleocytosis. RESULTS: Data were analyzed on 42 children and adolescents diagnosed with ADEM during 1991-2000, and CSF IL-12, IFN-gamma and IL-10 levels were compared among ADEM (n = 14), EV meningoencephalitis (n = 14) and controls without pleocytosis (n = 28). Overall incidence of ADEM was 0.4/100,000/year; incidence quadrupled during 1998-2000 compared with earlier years. No gender, age stratum, ethnic group or geographic area was disproportionately affected. A total of 4 (9.5%) patients initially diagnosed with ADEM were subsequently diagnosed with MS after multiple episodes of demyelination. Although most children eventually recovered, 2 died, including 1 of the 3 ultimately diagnosed with MS. Magnetic resonance imaging was required for diagnosis among 74% of patients; computerized tomography findings were usually normal. Patients with EV had significantly higher mean CSF IFN-gamma (P = 0.005) and IL-10 (P = 0.05) than patients with ADEM and controls without CSF pleocytosis. CSF from ADEM patients had CSF cytokine values statistically similar to those of 3 patients subsequently diagnosed with MS. CONCLUSIONS: ADEM is a potentially severe demyelinating disorder likely to be increasingly diagnosed as more magnetic resonance imaging studies are performed on patients with acute encephalopathy. Further characterization of the central nervous system inflammatory response will be needed to understand ADEM pathogenesis, to improve diagnostic and treatment strategies and to distinguish ADEM from MS. PMID- 15295227 TI - Progress in group A streptococcal vaccine development. PMID- 15295228 TI - Cochlear implants and meningitis in children. PMID- 15295229 TI - Interferon-gamma and colony-stimulating factors as adjuvant therapy for refractory fungal infections in children. AB - A human immunodeficiency virus-infected boy with Scedosporium apiospermum otomastoiditis and a girl with diabetes mellitus and Mucor sinusitis and orbital cellulitis had life-threatening disease progression despite antifungal treatment. Interferon-gamma and granulocyte-macrophage or granulocyte colony-stimulating factor were added, with good functional outcome in both children. Adjunctive therapy with interferon-gamma, granulocyte-macrophage colony-stimulating factor and granulocyte colony-stimulating factor can be considered for refractory invasive fungal infections. PMID- 15295230 TI - Fungal biofilm formation on cochlear implant hardware after antibiotic-induced fungal overgrowth within the middle ear. AB - Cochlear implantation in patients with chronic suppurative otitis media is managed with perioperative antibiotics; however, fungal overgrowth can occur. We present a child who received oral cefdinir and topical ofloxacin (Floxin). After 6 weeks, a fungal (Candida) biofilm was demonstrated on the implant surface. In this clinical setting, an antimicrobial strategy using an oral antifungal to prevent fungal overgrowth is a possibility. PMID- 15295231 TI - Epidemiology of invasive Streptococcus pneumoniae infections and vaccine implications among children in a West Virginia community, 1978-2003. AB - From 1978 to 2003, in Huntington, WV, we investigated Streptococcus pneumoniae invasive disease and the effect of conjugated pneumococcal vaccine among 161 children 14 years of age and younger admitted to the hospital. During 2002 and 2003, the number and proportion of invasive disease caused by vaccine strains declined; and in 2003, no invasive disease occurred in young children, suggesting a vaccine effect. PMID- 15295232 TI - Disseminated histoplasmosis in a nonendemic area. AB - Health care providers in the areas where histoplasmosis is not endemic can benefit greatly from understanding the clinical presentation, diagnosis and management of disseminated histoplasmosis as patients from the endemic areas may travel to and require medical attention in areas of low disease prevalence. Use of effective diagnostic tools such as Histoplasma antigen detection can aid in providing timely and appropriate therapy. PMID- 15295233 TI - Pulmonary symptoms in Kawasaki disease. AB - The diagnosis of Kawasaki disease is based on 6 clinical criteria, 5 of which must be fulfilled. The presence of uncommon symptoms in addition to the classic criteria can be as misleading as the lack of common ones. Here we report 2 infants with marked pulmonary symptoms in the course of Kawasaki disease who were initially diagnosed with pneumonia. PMID- 15295234 TI - Calmette-Guerin bacillus sternal osteomyelitis diagnosed by DNA sequencing analysis of PNC A. AB - Sternal osteomyelitis is an uncommon complication of Calmette-Guerin bacillus vaccination. Herein we describe a 4-month-old Taiwanese infant with a growing parasternal mass resulting from sternal osteomyelitis. By using DNA sequencing analysis, we identified the etiology as Calmette-Guerin bacillus vaccination. PMID- 15295235 TI - Cerebrospinal latex agglutination fails to contribute to the microbiologic diagnosis of pretreated children with meningitis. AB - We conducted a 10-year retrospective study of all children who had cerebrospinal fluid latex agglutination testing for bacterial antigens performed at 1 tertiary care urban children's hospital. Of the 176 patients with culture-negative meningitis who were pretreated with antibiotics before lumbar puncture, none had a positive latex agglutination study (0 of 176; 95% confidence interval, 0-2%). Latex agglutination studies identified no additional cases of bacterial meningitis beyond those identified by culture in pretreated patients. Clinical decision-making algorithms for the management of pretreated patients at risk for bacterial meningitis should not include latex agglutination testing. PMID- 15295236 TI - Poliomyelitis-like syndrome in a child with West Nile virus infection. AB - West Nile virus (WNV) infections of the central nervous system are very uncommon in U.S. children. We report a child with a poliomyelitis-like presentation in which WNV was the only detected pathogen. WNV has not previously been associated with a poliomyelitis-like presentation in children. PMID- 15295237 TI - Failure to diagnose Kawasaki disease at the extremes of the pediatric age range. AB - To learn about physician practices in diagnosing Kawasaki disease, we surveyed general pediatricians and pediatric infectious disease physicians by questionnaire. A high proportion of general pediatricians (>50%) and infectious disease subspecialists (25%) did not consider the diagnosis of Kawasaki disease in children younger than 6 months and older than 8 years. Failure to consider the diagnosis at the extremes of the pediatric age range puts children at risk because coronary artery abnormalities occur more often in young infants and adolescents with Kawasaki disease. PMID- 15295238 TI - Wound drainage after trauma in a fourteen-year-old girl. PMID- 15295239 TI - Avian influenza virus infection of children in Vietnam and Thailand. AB - Influenza viruses from chickens (H5N1) have caused outbreaks in children from both Vietnam and Thailand in 2004. All infected patients presented with fever and cough. Striking laboratory findings included leukopenia and thrombocytopenia. All children who developed progressive pneumonia with acute respiratory distress syndrome died. However, very few children received antiviral therapy. PMID- 15295240 TI - Treatment of community-associated methicillin-resistant Staphylococcus aureus skin and soft tissue infections with drainage but no antibiotic therapy. PMID- 15295243 TI - Activated charcoal administration in a pediatric emergency department. AB - OBJECTIVES: Activated charcoal is the commonest form of gastrointestinal decontamination offered to potentially poisoned children within United States emergency departments. Our aim was to describe this practice with regard to timing, route of administration, use of flavoring agents, and occurrence of adverse events other than vomiting. METHODS: Descriptive data were prospectively collected from consecutive administrations of single-dose activated charcoal, within an urban, academic pediatric emergency department, over a period of 2.5 years. RESULTS: Two hundred seventy-five subjects were enrolled. The median time elapsed between ingestion and emergency department arrival was 1.2 hours. Although 55% of children were administered charcoal within 1 hour of emergency department presentation, only 7.8% received charcoal within 1 hour of poisoning exposure. Forty-four percent of children younger than 6 years, 50% of 6-year to 12-year olds, and 89% of 12-year to 18-year olds drank the charcoal voluntarily (P < 0.01). Medical staff chose not to offer charcoal orally to 42 asymptomatic children among the 176 subjects under the age of 6 years. Of the 114 young children offered oral charcoal, 36 (32%) refused or were intolerant. Nurses added flavoring agents to the charcoal in 59% of oral administrations, but this act did not enhance observed palatability. Among children younger than 6 years, the median time from first sip to complete ingestion of charcoal slurry was 15 minutes. One pulmonary aspiration event and a case of constipation were noted. CONCLUSIONS: Despite published guidelines, children treated in an emergency department rarely received charcoal within 1 hour of ingestion. Gastric tube administration of charcoal varies by age and is partly subjective in its application. We found no evidence that excipient flavoring of charcoal improved success of administration. Pulmonary aspiration of charcoal, although uncommon, should be considered when assessing the risk of therapy. We offer a report of symptomatic constipation from single-dose charcoal. PMID- 15295244 TI - Etomidate versus pentobarbital for sedation of children for head and neck CT imaging. AB - OBJECTIVES: We compare etomidate to pentobarbital for sedation of children for head and neck computed tomography imaging. METHODS: We performed a prospective, randomized, double-blinded trial of patients aged 6 months to 6 years enrolled from the emergency department or radiology department at a large urban children's hospital. The primary outcome measure was sedation success rate. RESULTS: A total of 61 patients were enrolled in the study (27 etomidate group, 34 pentobarbital group) at 2 different dosing regimens for etomidate. The final analysis group included 17 etomidate patients and 33 pentobarbital patients. The success rate for the etomidate group was 57% at total doses of up to 0.3 mg/kg (n = 7) and 76% at total doses of up to 0.4 mg/kg (n = 17), in contrast to a success rate of 97% for pentobarbital at a total dose of up to 5 mg/kg (n = 33). The success rate for pentobarbital was significantly greater than the final etomidate group (P = 0.04; difference in proportions 20.5%, 95% CI 1.9% to 44.4%). Patients receiving etomidate had significantly shorter induction times (P = 0.02; difference of means 2.1 minutes, 95% CI 0.35 to 3.86), sedation times (P < 0.001; difference of means 31.3 minutes, 95% CI 24.0 to 38.5), and total examination times (P < 0.001; difference of means 53.1 minutes, 95% CI 40.8 to 65.3). Significantly more parents in the etomidate group perceived their child to be back to baseline by discharge from the hospital (P < 0.001; difference of proportions 60.7, 95% CI 29.1 to 92.4) and expressed fewer concerns about their child's behavior after discharge (P = 0.024; difference of proportions 28.6, 95% CI 6.5 to 50.7). CONCLUSIONS: At the dosing used in this study, pentobarbital is superior to etomidate when comparing success rates for sedation. However, among the successful sedations, the duration of sedation was shorter in the etomidate group than in the pentobarbital group. Pentobarbital is associated with more frequent side effects and parental concerns compared to etomidate. PMID- 15295245 TI - Children referred to an emergency department by an after-hours call center: complaint-specific analysis. AB - BACKGROUND AND OBJECTIVES: Approximately 20% of phone calls to after-hours call centers result in referrals to the emergency department (ED), but data regarding ED management and disposition are lacking. We sought to determine the acuity of illness of referred children as reflected by triage classifications and need for therapeutic interventions, diagnostic testing, and hospitalization, and to stratify the analysis of ED management and dispositions by chief complaints. DESIGN AND METHODS: Patients referred to the ED by the after-hours call centers, without physician consultation, were identified. The 4 most common groups of chief complaints resulting in ED referral were determined, and the records of these children were analyzed. RESULTS: The 525 patients with chief complaints related to the following organ systems were studied: lower respiratory tract, 263 (50%); gastrointestinal, 104 (20%); head, ears, eyes, nose, and throat, 84 (16%); and upper respiratory tract, 74 (14%). The proportion of children referred for lower respiratory tract complaints who received the after-hours call centers call dispositions (99%) or ED triage classifications (38%) of highest priorities, or who required therapeutic interventions (73%), diagnostic testing (40%), or hospitalization (22%) was significantly higher than for all other categories. Thirteen percent with gastrointestinal complaints, referred primarily for dehydration, required intravenous fluids, and 2% of head, ears, eyes, nose, and throat patients required hospitalization. CONCLUSIONS: Children referred to the ED for illnesses related to the lower respiratory tract, principally wheezing, had illnesses of high acuity. On the other hand, current criteria for ED referral for children in the gastrointestinal, head, ears, eyes, nose, and throat, and upper respiratory tract categories result in the referral of many children with nonurgent problems. These data support a reassessment of current referral practices for children with these complaints. PMID- 15295246 TI - Agreement among pediatric health care professionals with the pediatric Canadian triage and acuity scale guidelines. AB - OBJECTIVES: To compare triage level assignment, using case scenarios, in a pediatric emergency department between registered nurses (RNs) and pediatric emergency physicians (PEPs) based on the Pediatric Canadian Triage and Acuity Scale (P-CTAS) guidelines. To compare triage level assignment of the RNs and PEPs to that done by a panel of experts using the same P-CTAS guidelines. METHODS: A cross-sectional questionnaire survey (55 case scenarios) was sent to all RNs and PEPs working in the emergency department after the P-CTAS was implemented. Participants were instructed to assign a triage level for each case. A priori, all cases were assigned a triage level by a panel of experts using the P-CTAS guidelines. Kappa statistics and the mean number (+/-1SD) of correct responses were calculated. RESULTS: A response rate of 85% was achieved (29 RNs, 15 PEPs). The kappa level of agreement (95% CI) among RNs was 0.51 (0.50-0.52) and was 0.39 (0.38-0.41) among PEPs (P < 0.001). The mean number of correct responses (+/-1SD) for RNs was 64% +/- 27% and for PEPs 60% +/- 22% (P = 0.31). Levels of agreement did not vary according to experience or type of shift work done or work status of RNs and PEPs. CONCLUSIONS: With the introduction of the P-CTAS, the level of agreement and accuracy of triage categorization remained moderate for both RNs and PEPs. The reliability of the P-CTAS needs to be further assessed and the requirements for revisions considered prior to its widespread use. PMID- 15295247 TI - Randomized controlled comparison of cosmetic outcomes of simple facial lacerations closed with Steri Strip Skin Closures or Dermabond tissue adhesive. AB - OBJECTIVE: To compare the short-term complications and long-term cosmetic outcomes of simple facial lacerations closed with 3M Steri Strip Skin Closures or Dermabond. METHODS: Prospective, randomized controlled trial of children ages 1 to 18 presenting to a pediatric emergency department with simple low-tension lacerations of the face. After standard wound care, patients received wound closure with either Steri Strip Skin Closure or Dermabond. Pain associated with closure was evaluated on a 100-mm visual analogue scale (0 = no pain, 100 = worst pain). A follow-up telephone call was made a week after enrollment to determine short-term complications. Patients returned 2 months after would closure for wound photography. Cosmetic outcomes were evaluated by 2 plastic surgeons blinded to the method of wound closure on a 100-mm visual analogue scale (0 = best scar, 100 = worst scar). RESULTS: One hundred children aged 1 to 18 were enrolled. Ninety-seven patients had results analyzed. Forty-eight received Steri Strip Skin Closures and 49 received Dermabond. Patient demographics and wound characteristics were similar between groups. Pain scores on a 100-mm visual analogue scale were 9.0 mm for the Steri Strip group and 6.2 mm for the Dermabond group (P = ns). At short-term follow-up, there was one wound complication in the Steri Strip group and 7 complications in the Dermabond group (P = 0.06). Eighty nine patients received 2-month evaluation (41 Steri Strip, 45 Dermabond). There was no difference in the mean visual analogue scale cosmesis scores: 37.2 mm (95% CI = 30.8-43.7) versus 43.8 mm (95% CI = 38.4-49.2) (P = 0.12). CONCLUSIONS: Steri Strip Skin Closures and Dermabond provide similar cosmetic outcomes for closure of simple facial lacerations. Steri Strip Skin Closure may represent a low-cost alternative for closure of simple facial lacerations. PMID- 15295248 TI - Cecal retention of a swallowed penny mimicking appendicitis in a healthy 2 year old. AB - Toddlers commonly ingest coins. Studies of the evaluation and management of such ingestions have focused on the risk of complications from impaction in the esophagus. It is commonly assumed that coins that have passed through the esophagus present little or no risk for distal complications. We present the first report of cecal retention of a penny in a previously healthy 2 year old, ultimately resulting in surgical intervention. PMID- 15295249 TI - Pneumococcal-associated purpura fulminans in a healthy infant. PMID- 15295250 TI - Abdominal pain-an uncommon cause. PMID- 15295251 TI - Accidental ecstasy poisoning in a toddler. AB - A child of 14 months accidentally swallowed a portion of an Ecstasy pill. Forty minutes after ingestion, he started a generalized convulsion. He also presented hyperthermia (38 degrees C), hypertension, tachycardia (130 bpm), ventricular extrasystoles, tachypnea (50 rpm), and mydriasis. At the hospital 5 hours later, the urine levels of amphetamine/metamphetamine were >16 mg/L. He was treated with general support measures and benzodiazepines intravenously and admitted to the pediatric intensive care unit. During the first 12 hours, he continued with hypertension, tachycardia, and long periods of trigeminy, without hemodynamic repercussion. He was discharged fully recovered. Gas chromatography/mass spectrometry analysis (8 hours after ingestion) showed a serum level of 3,4 methylenedioximethylamphetamine (MDMA) of 0.591 mg/L. The 3 cases described in the literature have shown a good evolution of Ecstasy poisoning in toddlers and infants, despite an initial critical situation. Regarding adults, the toddler intoxication seems to present symptoms sooner (20 to 30 minutes), having as an initial manifestation convulsions. However, great care must be taken on accidental ingestion of these attractive design pills. PMID- 15295252 TI - Clearing the fog (a bit): the 2003 EMTALA regulations and their implications for pediatric emergency departments. PMID- 15295253 TI - "Doctor, my child needs some medicine!". PMID- 15295254 TI - Painful, red umbilicus. PMID- 15295255 TI - Motor vehicle accident and alarming chest X-ray. PMID- 15295256 TI - ECGs in the ED. PMID- 15295257 TI - Pulmonary embolism in the pediatric patient. PMID- 15295259 TI - RN-BSN programs: associate degree and diploma nurses' perceptions of the benefits and barriers to returning to school. AB - The nursing profession is facing a serious shortage in all areas including BSN completion programs. The purpose of this study was to explore associate degree and diploma nurses' perceptions of the benefits and barriers to RN-BSN programs. In addition, factors that would facilitate degree completion in academia and work environments were examined. PMID- 15295261 TI - Teaching old dogs new tricks: RN "Returnship" Program. AB - RN shortages, lack of experienced RNs, technological advances, closed doors to RNs without recent acute care experience-these are the common problems making it difficult for experienced RNs to return to the hospital setting after an extended absence. Solution? A specialized orientation-the "RN Returnship Program." This creative and adaptable medical-surgical nursing program met these unique needs. The Returnship Program was designed to support "old dog" RNs and teach them the "new tricks" of acute care nursing. PMID- 15295262 TI - Practice makes perfect: preparing graduate nurses for professional licensure. AB - In the fall of 2000, the licensing examination for nursing taken by candidates in the Province of Quebec was altered to include an oral practical component. This article describes how a group of nurse clinician educators, despite severe time and human resources constraints, developed a unique and interactive program to assist graduate unlicensed nurses to successfully prepare for this new examination process. PMID- 15295263 TI - Post-anesthesia care unit nurses' knowledge of pulse oximetry. AB - The purpose of this study was to assess post-anesthesia care unit (PACU) nurses' knowledge of pulse oximetry. A convenience sample of 19 nurses completed a 32 item questionnaire that included a 20-item true-false test on pulse oximetry. Overall, nurses demonstrated a knowledge deficit in pulse oximetry. Competency in the use of pulse oximetry is vital to ensure a positive clinical outcome. Nurse educators are responsible for identifying knowledge deficits among staff and implementing strategies to correct these deficits. It is incumbent on nurse educators to provide research-based education on pulse oximetry and opportunities to participate in continuing education. PMID- 15295264 TI - A model of nurse assistant care to promote functional status in hospitalized elders. AB - After completing 20 hours of classes on promoting the functional status of hospitalized elders, the certified nursing assistants on this medical unit participated in developing a new model of care delivery. Discharge destination (home or nursing home) and length of stay were compared for patients pre- and post-implementation. Length of stay decreased by 2.4 days (p =.0007), and there was a significant increase in the number of elders who were able to return home (p =.024). PMID- 15295265 TI - Is obtaining magnet status valuable? PMID- 15295266 TI - Writing continuing education courses. PMID- 15295267 TI - Instant teaching tools. PMID- 15295270 TI - Techniques to assess in-vivo tissue metabolism directly in humans without biopsy samples. PMID- 15295271 TI - Microdialysis methods for measuring human metabolism. AB - PURPOSE OF REVIEW: To discuss the advantages and limitations of the microdialysis technique as a diagnostic and research tool using recent findings on human metabolism. RECENT FINDINGS: Results from several studies have supported the potential of microdialysis as a diagnostic tool for metabolic monitoring in difficult accessible tissues (brain, liver, intestine). However, despite promising results, no clear diagnostic measures have yet emerged. Several studies combining microdialysis with stable isotope tracers have shown that this approach has great potential for studying human metabolism non-invasively in specific tissue beds in a more dynamic way. SUMMARY: Bearing in mind the limitations and assumptions, the microdialysis technique is a useful tool in investigations of human metabolism. Its main advantages are that it can be used safely with low grade invasiveness in humans, and thereby allows continuous sampling over prolonged periods of time from specific tissues without taking any biopsies. At present, microdialysis would seem to be useful as a diagnostic tool when integrated in the total clinical, physiological and pharmacological evaluation. Within human metabolic research, microdialysis has been and will be a very useful technique. PMID- 15295272 TI - Methods of measuring metabolism during surgery in humans: focus on the liver brain relationship. AB - PURPOSE OF REVIEW: The purpose of this work is to review recent advances in setting methods and models for measuring metabolism during surgery in humans. Surgery, especially solid organ transplantation, may offer unique experimental models in which it is ethically acceptable to gain information on difficult problems of amino acid and protein metabolism. RECENT FINDINGS: Two areas are reviewed: the metabolic study of the anhepatic phase during liver transplantation and brain microdialysis during cerebral surgery. The first model offers an innovative approach to understand the relative role of liver and extrahepatic organs in gluconeogenesis, and to evaluate whether other organs can perform functions believed to be exclusively or almost exclusively performed by the liver. The second model offers an insight to intracerebral metabolism that is closely bound to that of the liver. SUMMARY: The recent advances in metabolic research during surgery provide knowledge immediately useful for perioperative patient management and for a better control of surgical stress. The studies during the anhepatic phase of liver transplantation have showed that gluconeogenesis and glutamine metabolism are very active processes outside the liver. One of the critical organs for extrahepatic glutamine metabolism is the brain. Microdialysis studies helped to prove that in humans there is an intense trafficking of glutamine, glutamate and alanine among neurons and astrocytes. This delicate network is influenced by systemic amino acid metabolism. The metabolic dialogue between the liver and the brain is beginning to be understood in this light in order to explain the metabolic events of brain damage during liver failure. PMID- 15295273 TI - An integrative approach to in-vivo protein synthesis measurement: from whole tissue to specific proteins. AB - PURPOSE OF REVIEW: In-vivo estimation of protein turnover by stable isotopes in animals and humans has provided much relevant information on metabolic regulation and alterations for decades. While it was first appreciated at the whole body level in the 1970s and 1980s, new approaches have allowed inter-organ or tissue protein turnover rates to be measured, notably the incorporation rate of a labelled amino acid in muscle. These technical improvements have recently been completed by new isolation methods for the study of protein synthesis rates in various muscle and hepatic protein fractions in different blood cells or tissues such as bone and skin. RECENT FINDINGS: This new insight into tissue protein synthesis opens the door for exploration of single proteins, which may be fully achievable in the near future through the combination of proteomics analysis and technical progress in mass spectrometry. This is, therefore, a new area in which not only quantitative but also qualitative changes in specific proteins will be considered for a fully integrative approach to assessing protein metabolism in physiology and disease. SUMMARY: To understand the mechanisms by which protein metabolism is altered during physiopathological situations, it is of importance to measure the effect on specific proteins rather than on the body as a whole. Procedures are currently under development with the aim of isolating individuals proteins and to measure their synthesis rates by isotopic methods. Such technical progress is needed to gain a better understanding of the regulation of protein metabolism in situations in which loss of body protein mass occurs. PMID- 15295274 TI - Quantifying folate bioavailability: a critical appraisal of methods. AB - PURPOSE OF REVIEW: Dietary reference intakes for folate rely on a good estimate of folate bioavailability from the general diet. In this review, current methods for quantifying the bioavailability of dietary folate and specific folate vitamers in humans are reviewed. Emphasis is on isotopic labeling techniques that have been developed during the past 15 years. RECENT FINDINGS: Most reported studies applied single-dose designs, in which blood or urine concentrations of folate are measured for several hours after oral folate administration. To obtain a measurable biochemical response, however, relatively high doses of folic acid are administered and individuals are often saturated with large doses of folic acid prior to study. The effect of this on folate absorption and metabolism is poorly understood. Therefore, study designs in which multiple oral doses are administered are preferred. Several such studies, both with unlabeled and isotopically labeled folic acid, are discussed. Although many studies have been performed on the bioavailability of specific folate compounds and of folate from single foods, reliable data in which the bioavailability of folate from total diets have been measured are currently lacking. SUMMARY: A multiple oral dose design is the best approach for measuring folate bioavailability because there are several serious drawbacks to designs based on the use of a single oral dose. Studies on folate bioavailability from total diets are urgently required in order to evaluate current recommendations for folate intake. PMID- 15295275 TI - Measuring very low density lipoprotein-triglyceride kinetics in man in vivo: how different the various methods really are. AB - PURPOSE OF REVIEW: The purpose of this article is to briefly outline the methods that are currently available for the determination of very low density lipoprotein-triglyceride (VLDL-TG) kinetics in man in vivo. RECENT FINDINGS: A number of novel methodologies have been developed over the years for quantifying VLDL-TG production, clearance, and turnover rates. Besides the splanchnic arteriovenous balance technique, tracer methods with radioactive and, more recently, stable isotopes have been widely used. Most of the latter approaches utilize an isotopically labelled substrate, such as glycerol, fatty acid or acetate, which is eventually incorporated into a VLDL-TG moiety, and monitor the time course of change in specific activity or enrichment. A procedure of in vivo labelling of VLDL-TG with stable isotopes and use of the labelled VLDL-TG as a tracer has also been described in man. There is, however, considerable variability in estimates of VLDL-TG kinetics obtained by the various techniques, which cannot be readily attributed to normal physiological variation. Still, a large part of this discrepancy may be related to differences in VLDL-TG pool size within the normal range, which seem to account for approximately 40-50% of the total variance in VLDL-TG kinetics in both men and women. SUMMARY: Several methods are available for quantifying VLDL-TG kinetics in man in vivo, varying in the selection of tracer, mode of administration and sampling, and data analysis. These inherent features, along with different pool sizes, result in multifold variable estimations of VLDL-TG kinetic parameters. PMID- 15295276 TI - The liquid chromatography mass spectrometry approach to measure amino acid isotope ratios. AB - PURPOSE OF REVIEW: Although the application of liquid chromatography mass spectrometry has seen a rapid development in the world of clinical science since the introduction of atmospheric pressure ionization, its potential is still hardly recognized in the field of stable isotope biomedical research, as indicated by the small number of publications on this topic. Nevertheless, considering the polar nature of many biological substrates of interest as well as the non-destructive nature of electrospray ionization, a liquid chromatography mass spectrometry approach simplifies mass spectrometric spectra, allows the application of multiple labelled tracers, and provides an easy on-line sample processing, thus minimizing labour while at the same time providing concentration data and good reproducibility in low quantities of sample. RECENT FINDINGS: These features make it an excellent tool to perform both clinical and animal experiments that usually generate large numbers of samples, which when processed through the classic approach employing gas chromatography combustion isotope ratio mass spectrometry systems would require much more work and time and thus analytical costs. SUMMARY: The present paper is intended to introduce the functionality, limitations and options of representatives from the present family of modern liquid chromatography mass spectrometry systems into the field of stable isotope biomedical research, with a focus on amino acid metabolism. PMID- 15295278 TI - Nutrient deficiencies secondary to bariatric surgery. AB - PURPOSE OF REVIEW: The number of adolescent and adult patients submitting to bariatric surgery is increasing rapidly around the world. This review describes the literature published in the last few years concerning nutritional deficiencies after bariatric surgery as well as their etiology, incidence, treatment and prevention. RECENT FINDINGS: Although bariatric surgery was first introduced in the 1950s, safe and successful surgical management has progressed over the last two decades and longer post-surgical follow-up data are now available. Most of the patients undergoing malabsorptive procedures will develop some nutritional deficiency, justifying mineral and multivitamin supplementation to all postoperatively. Nutrient deficiency is proportional to the length of absorptive area and to the percentage of weight loss. Low levels of iron, vitamin B12, vitamin D and calcium are predominant after Roux-en-Y gastric bypass. Protein and fat-soluble vitamin deficiencies are mainly detected after biliopancreatic diversion. Thiamine deficiency is common in patients with frequent vomiting. As the incidence of these deficiencies progresses with time, the patients should be monitored frequently and regularly to prevent malnutrition. SUMMARY: Nutritional deficiencies can be prevented if a multidisciplinary team regularly assists the patient. Malnutrition is generally reverted with nutrient supplementation, once it is promptly diagnosed. Especial attention should be given to adolescents, mainly girls at reproductive age who have a substantial risk of developing iron deficiency. Future studies are necessary to detect nutrient abnormalities after new procedures and to evaluate the safety of bariatric surgery in younger obese patients. PMID- 15295277 TI - The butyrate story: old wine in new bottles? AB - PURPOSE OF REVIEW: Short-chain fatty acids are important end products of bacterial carbohydrate fermentation in the colon. In particular, n-butyrate is thought to play a regulatory role in the maintenance of a physiological environment. Disturbances in the interplay between the microflora and the lining epithelium may lead to mucosal inflammation and promote carcinogenesis. The purpose of this article is to review the literature between March 2003 and February 2004 and to determine if recent studies have improved the understanding of butyrate effects in health and disease. RECENT FINDINGS: Preclinical studies (cell culture experiments, animal studies) using modern molecular biological tools (including cDNA arrays) have provided new insights into the action of butyrate on colonic epithelial, vascular endothelial and extracolonic cell types. The new information adds pieces of evidence to the assumption that butyrate may ameliorate colonic inflammation and may be chemopreventive in carcinogenesis. In contrast, new data from clinical studies have been limited in the review period. SUMMARY: In the era of molecular biology our understanding of subcellular processes that ultimately lead to inflammatory bowel disease or colorectal cancer has widened considerably. The new powerful technology of genomics and proteomics, however, raises new questions without easy answers. With this new information in mind, we will have to go back to human intervention trials to test the hypotheses generated in vitro. The preclinical data from the review period justify the need for carefully designed clinical trials to test the benefits derived from butyrate production. PMID- 15295279 TI - The impact of early nutrition on metabolic response and postoperative ileus. AB - PURPOSE OF REVIEW: Early nutrition has been evaluated and used as a possible strategy to decrease the negative impact of the metabolic response to injury and postoperative ileus. The metabolic response to injury, be it surgical or traumatic, is a physiological mechanism that, according to the magnitude and duration of the event, can impact on the patient's morbidity and survival. The adequate initial approach is a determinant factor that might influence its outcome. Simultaneously, gastrointestinal tract motility is transiently impaired, leading to the so-called postoperative ileus. The latter not only causes patient discomfort, but is also related to abdominal complications and worsening of the nutritional status, as well as increased length of hospital stay and costs. RECENT FINDINGS: Multimodal surgical strategies such as preoperative intake of a carbohydrate drink, together with patient education of the postoperative care plan, efficacious analgesia and early nutrition have been described to significantly decrease the stress response and improve the ileus. Therefore, these strategies accelerate rehabilitation and, as a consequence, decrease complications and length of hospital stay and its related costs. SUMMARY: Understanding perioperative pathophysiology and implementing care regimes through a multimodal approach in order to reduce the stress of the operation and the related postoperative ileus are major challenges. These factors will certainly impact on patient outcomes. PMID- 15295280 TI - Bibliography. Current world literature. Assessment of nutritional status and analytical methods. PMID- 15295297 TI - [Recommendations of SPC/ALFEDIAM on the care of diabetic patient as seen by the cardiologist]. PMID- 15295299 TI - Peripheral arterial disease. When leg pain is more than a simple ache. You're climbing a flight of stairs and you feel it--a familiar achy feeling or painful cramping in your calves or thighs. PMID- 15295300 TI - Postmenopausal estrogen therapy not helpful for conditions of aging. PMID- 15295301 TI - Component in green tea helps kill leukemia cells. PMID- 15295302 TI - Your adrenal glands. Vital for your good health. AB - You probably don't think much about your adrenal glands, possibly not even what they are or what they do. Although tiny, these glands produce hormones such as cortisol and adrenaline, which help regulate crucial body functions, including your reactions to stress. PMID- 15295303 TI - Second opinions. When to seek additional medical advice. PMID- 15295305 TI - Exercising when it hurts. PMID- 15295306 TI - The beta blocker I take has slowed my heart rate, making it difficult to achieve my target heart rate when I exercise. Does this mean the exercise isn't doing me any good? PMID- 15295307 TI - What can I do to prevent embarrassing gas? PMID- 15295308 TI - A friend of mine was recently diagnosed with mammary duct ectasia. Can you tell me more about this condition? PMID- 15295309 TI - Follow-up care after breast cancer treatment. PMID- 15295310 TI - Nonfatal motor-vehicle animal crash-related injuries--United States, 2001-2002. AB - In 2000, an estimated 6.1 million light-vehicle (e.g., passenger cars, sport utility vehicles, vans, and pickup trucks) crashes on U.S. roadways were reported to police. Of these reported crashes, 247,000 (4.0%) involved incidents in which the motor vehicle (MV) directly hit an animal on the roadway. Each year, an estimated 200 human deaths result from crashes involving animals (i.e., deaths from a direct MV animal collision or from a crash in which a driver tried to avoid an animal and ran off the roadway). To characterize nonfatal injuries from these incidents, CDC analyzed data from the National Electronic Injury Surveillance System-All Injury Program (NEISS-AIP). This report summarizes the results of that analysis, which indicated that, during 2001-2002, an estimated 26,647 MV occupants per year were involved in crashes from encounters with animals (predominantly deer) in a roadway and treated for nonfatal injuries in U.S. hospital emergency departments (EDs). Cost-effective measures targeting both drivers (e.g., speed reduction and early warnings) and animals (e.g., fencing and underpasses) are needed to reduce injuries associated with MV collisions involving animals. PMID- 15295311 TI - Transmission of hepatitis B virus in correctional facilities--Georgia, January 1999-June 2002. AB - Incarcerated persons have a disproportionate burden of infectious diseases, including hepatitis B virus (HBV) infection. Among U.S. adult prison inmates, the overall prevalence of current or previous HBV infection ranges from 13% to 47%. The prevalence of chronic HBV infection among inmates is approximately 1.0%-3.7%, two to six times the prevalence among adults in the general U.S. population. Incarcerated persons can acquire HBV infection in the community or in correctional settings. This report summarizes the results of 1) an analysis of hepatitis B cases among Georgia inmates reported to the Georgia Department of Human Resources, Division of Public Health (DPH) during January 1999-June 2002, including a retrospective investigation of cases reported during January 2001 June 2002; and 2) a prevalence survey conducted in prison intake centers during February-March 2003. These efforts identified cases of acute hepatitis B in multiple Georgia prisons and documented evidence of ongoing transmission of HBV in the state correctional system. The findings underscore the need for hepatitis B vaccination programs in correctional facilities. PMID- 15295312 TI - Hepatitis B vaccination of inmates in correctional facilities--Texas, 2000-2002. AB - In December 2002, approximately 2.2 million persons were incarcerated in the United States; an estimated 8 million were released to the community that year. In 2001, approximately 22,000 acute hepatitis B cases and 78,000 new hepatitis B virus (HBV) infections occurred in the United States (3); an estimated 29% of these cases were in persons who had been incarcerated previously. The majority of HBV infections among incarcerated persons are acquired in the community; however, infection also is transmitted within correctional settings. Hepatitis B vaccination of incarcerated persons is recommended to prevent transmission in correctional facilities and in previously incarcerated persons on their return to the community. In May 2000, the Texas Department of Criminal Justice (TDCJ), which oversees custody of state jail and prison inmates, implemented a hepatitis B vaccination program. To determine hepatitis B vaccination rates of inmates during 2000-2002, TDCJ reviewed charts of inmates released during a 3-day period for documentation of vaccination. This report summarizes the results of that study, which indicated that rates of vaccine acceptance and vaccine series completion among inmates were high. Establishing hepatitis B vaccination programs in prisons and jails can prevent a substantial proportion of HBV infections among adults in the outside community. PMID- 15295313 TI - Tuberculosis associated with blocking agents against tumor necrosis factor-alpha- California, 2002-2003. AB - The Food and Drug Administration (FDA) has determined that tuberculosis (TB) disease is a potential adverse reaction from treatment with the tumor necrosis factor-alpha (TNF-alpha) antagonists infliximab (Remicade), etanercept (Enbrel), and adalimumab (Humira); the three products are labeled accordingly. These products work by blocking TNF-alpha, an inflammatory cytokine, and are approved for treating rheumatoid arthritis and other selected autoimmune diseases. TNF alpha is associated with the immunology and pathophysiology of certain infectious diseases, notably TB; blocking TNF-alphacan allow TB disease to emerge from latent Mycobacterium tuberculosis infection. In 2002, a California county health department reported three cases of TB disease occurring in association with infliximab therapy. This report summarizes those cases and nine subsequently reported cases and provides interim recommendations for TB prevention and management in recipients of these blocking agents. Health-care providers should take steps to prevent TB in immunocompromised patients and remain vigilant for TB as a cause of unexplained febrile illness. PMID- 15295314 TI - West Nile virus activity--United States, July 28-August 3, 2004. AB - During July 28-August 3, a total of 141 cases of human West Nile virus (WNV) illness were reported from 11 states (Alabama, Arizona, California, Colorado, Florida, Illinois, Nevada, New York, North Dakota, South Dakota, and Texas). During 2004, a total of 20 states have reported a total of 406 cases of human WNV illness to CDC through ArboNET. Of these, 247 (61%) were reported from Arizona. A total of 226 (57%) of the 406 cases occurred in males; the median age of patients was 51 years (range: 1 month-99 years). Illness onset ranged from April 20 to July 29; seven cases were fatal. PMID- 15295315 TI - Masticatory performance and efficiency in denture wearers. 1964. PMID- 15295316 TI - An interdisciplinary approach for restoring function and esthetics in a patient with amelogenesis imperfecta and malocclusion: a clinical report. AB - This clinical report describes an interdisciplinary approach for the coordinated treatment of a patient diagnosed with amelogenesis imperfecta and malocclusion. The patient's functional and esthetic expectations were successfully met with interdisciplinary treatments, including orthodontics, porcelain laminate veneers, metal-ceramic fixed partial dentures, and direct composite restorations. PMID- 15295318 TI - Altered prosthodontic treatment approach for bilateral complete maxillectomy: a clinical report. AB - Bilateral maxillary resection offers a significant challenge for prosthodontic rehabilitation with a removable tissue-borne prosthesis, owing in large part to a lack of adequate retention, support, and stability. The clinical management of a patient with a bilateral maxillectomy using an altered prosthodontic treatment sequence is described. PMID- 15295317 TI - Esthetic enhancement of ceramic crowns with zirconia dowels and cores: a clinical report. AB - There has been an increase in the use of esthetic metal-free ceramic crowns in restoring endodontically treated teeth or teeth with severe coronal destruction. Tooth-colored dowels and cores are used to enhance the esthetic result. This report describes the treatment of a patient with bilateral maxillary supernumerary lateral incisors, a severe malocclusion, and maxillary anterior tooth discoloration. Treatment included heat-pressed, metal-free ceramic crowns supported by zirconia ceramic dowel-and-core foundations. PMID- 15295319 TI - A simple procedure for minimizing adjustment of immediate complete denture: a clinical report. AB - This article describes fabrication of a clear surgical template that minimizes pressure caused by immediate complete dentures on a surgical area. The trimmed areas on the maxillary definitive stone cast were further trimmed on the duplicated stone cast for making the clear surgical template. The procedure provided proper seating of the immediate complete denture, comfort for the patient, and reduced postoperative soreness and denture adjustments. PMID- 15295320 TI - Flexible denture flanges for patients exhibiting undercut tuberosities and reduced width of the buccal vestibule: a clinical report. AB - Buccal undercut of the maxillary tuberosity together with reduced width of the buccal vestibule can complicate denture fabrication. This clinical report describes the treatment options for this situation, the rationale for the design and use of flexible denture flanges in the maxillary posterior buccal vestibule, and the laboratory procedure for incorporation of flexible denture flanges in the undercut area. PMID- 15295321 TI - In vitro comparative analysis of the fit of gold alloy or commercially pure titanium implant-supported prostheses before and after electroerosion. AB - STATEMENT OF PROBLEM: For implant-supported prostheses, passive fit is critical for the success of rehabilitation, especially when alternative materials are used. PURPOSE: The purpose of this study was to compare interfacial fit of implant-supported prostheses cast in titanium to those cast in gold alloy. MATERIAL AND METHODS: Five 3-unit fixed partial dentures were fabricated in gold alloy (Degudent U) as 1-piece castings, and 5 others were similarly cast in commercially pure titanium (Grade 1). The interfacial gaps between the prostheses and the abutments were evaluated with an optical microscope, before and after electroerosion. Readings were made with both screws tightened (10 N.cm torque), and with only 1 side tightened, so as to also evaluate the passive fit of the prostheses. Data were compared statistically by 2-way analysis of variance and the post hoc Tukey multiple range test (alpha=.05). RESULTS: Before electroerosion, the interfacial gaps for the 1-piece prostheses were significantly smaller (P<.001) in the gold alloy group when the screws were tightened (Au=12.6 +/- 3.0 microm, compared to Ti=30.1 +/- 6.4 microm). When the side opposite the tightened side was analyzed, there was no significant difference between the gold alloy and titanium groups (Au=69.2 +/- 24.9 microm and Ti=94.2 +/- 39.6 microm). The electroerosion procedure significantly (P<.001) reduced the gaps at the interfaces for both groups under all conditions. Comparison between groups after electroerosion did not present significant differences when the side opposite the tightened side was analyzed, but the gold alloy group showed better fit when the tightened side was analyzed (12.8 +/- 1.4 microm for gold alloy; 29.6 +/- 4.4 microm for titanium) and when both screws were tightened (5.4 +/- 2.3 microm for gold alloy; 16.1 +/- 5.5 microm for titanium). CONCLUSIONS: Cast titanium prostheses, despite showing larger interfacial gaps between the prosthesis and abutment than those obtained with gold alloy, had improved fit after being subjected to electroerosion. PMID- 15295322 TI - The use of short, wide implants in posterior areas with reduced bone height: a retrospective investigation. AB - STATEMENT OF PROBLEM: Reduced bone height frequently presents a challenge for implant-assisted tooth replacement in partially edentulous patients. PURPOSE: This retrospective study evaluated the success rate of short, wide hydroxyapatite (HA)-coated implants placed in mandibular and maxillary molar areas with reduced bone height. MATERIAL AND METHODS: A total of 168 HA-coated implants (6-mm diameter x 8-mm length) were placed in 167 patients in a private-practice setting. A minimal 6-mm workable ridge height and 8-mm ridge width was available in all situations. Patients were referred back to 1 of 7 referring restorative dentists for restoration of the implants. No attempt was made to standardize the restoration of the implants except to avoid working and nonworking contacts in lateral excursions. Implant success was evaluated according to the following criteria: (1) absence of complaints, (2) absence of recurring peri-implant infection or suppuration, (3) absence of perceptible implant mobility, and (4) absence of radiolucencies at implant-bone junction. The data were analyzed with descriptive statistics. RESULTS: Fifty-four (32.1%), 35 (20.8%), 36 (21.4%), and 42 (25.0%) implants replaced maxillary first and second and mandibular first and second molars, respectively. There were 128 implant-supported single crowns. Thirty-eight implants served as abutments for fixed partial dentures connected to other implants of various sizes. Two implants were involved in cantilevered fixed partial dentures. Patients were followed for up to 68 months (mean=34.9 months) after loading of implants. The overall cumulative success rate was found to be 100%. CONCLUSIONS: For residual ridges with minimal height but adequate width, the use of short, wide HA-coated implants may offer a simple and predictable treatment alternative in posterior areas. PMID- 15295323 TI - A descriptive 18-year retrospective review of subperiosteal implants for patients with severely atrophied edentulous mandibles. AB - STATEMENT OF PROBLEM: Fabricating dentures for the patient with severe mandibular atrophy can be a challenge for both the dentist and patient. Subperiosteal implants with a mandibular overdenture may be a solution for the atrophic mandible. PURPOSE: The purpose of this retrospective study was to review the survival of mandibular subperiosteal implants placed at the University of Missouri Kansas City (UMKC) School of Dentistry Graduate Prosthodontics program between 1982 and 2000. MATERIAL AND METHODS: Forty subperiosteal implants were placed in atrophic mandibles of 40 patients (33 women, 7 men) between 1982 and 2000. The age range of the patients was 47 to 80 years of age at time of placement (mean=62 years). Each patient was reviewed clinically by an author (DJM). Manual depression and lifting of the framework were used to evaluate the stability of the implant. Additionally, the implants were observed for any movement. Each patient was questioned for pain or discomfort. Each patient was examined for observable inflammation and intraoral exposure of the framework and questioned as to whether the implant had satisfied the patient and met the patient's expectations. RESULTS: Thirty-nine of the 40 original patients were recalled in 2000. One patient had died. Fourteen patients had implants for over 10 years, 12 patients had implants between 5 and 10 years, and 12 patients had implants for less than 5 years (mean time of implant service=8 years). Thirty eight patients had the implant in place with no sign of inflammation or mobility, 1 patient with diabetes had inflammation around one of the struts. All patients were wearing their prostheses, and there was no sign of exposed implant framework for any patient. All patients reported a high level of satisfaction with the implant. CONCLUSIONS: Within the limitations of this study, the mandibular implants placed at UMKC were still functioning, and all patients denied any discomfort or pain from the prostheses. Patients reported they were comfortable and able to function with the implant-supported prosthesis. PMID- 15295324 TI - Mechanical properties of 3 hydrophilic addition silicone and polyether elastomeric impression materials. AB - STATEMENT OF PROBLEM: New "hydrophilic" elastomeric impression materials have been introduced with the goals of reducing marginal voids and distortion in the impressions and improving the quality of gypsum dies, but there are insufficient data on the mechanical properties of these materials. PURPOSE: Mechanical properties, including elastic recovery, strain in compression, tear energy, and tensile strength of 3 hydrophilic impression materials with low and high consistencies were compared. MATERIAL AND METHODS: Two addition silicone impression brands (Imprint II, 3M ESPE; Flexitime, Heraeus Kulzer) and a polyether brand (Impregum, 3M ESPE) were studied. Two consistencies of each material (light-body and heavy-body) were investigated. Elastic recovery (%) and strain in compression (%) were tested according to ISO 4823; tear energy (J/m2) and tensile strength (MPa) were tested following Webber and Ryge's method and ASTM D412 (Test Method A), respectively. Five specimens were made for each group for a total of 24 groups and 120 specimens. Results were analyzed by 2-way analysis of variance, and Fisher's protected least significance difference intervals were calculated (alpha=.05). Correlation analysis was used to evaluate the relationships among properties. RESULTS: P values were smaller than .0001 for material, consistency, and interaction for strain in compression, tear energy, and tensile strength. For elastic recovery, P values were smaller than .0001 for material and the interaction between material and consistency, but equal to .4150 for consistency. Strain in compression correlated with other mechanical properties (P<.05), but tensile strength and tear resistance were not correlated. CONCLUSIONS: In general, new "soft" polyether impression materials had higher strain in compression and lower tensile strength compared to new "hydrophilic" addition silicone materials. Heavy-body materials had higher tear properties and tensile strength than light-body materials. Strain in compression was correlated with elastic recovery, tear energy, and tensile strength. Tear resistance and tensile strength were not correlated. PMID- 15295325 TI - An in vitro study evaluating the effect of ferrule length on fracture resistance of endodontically treated teeth restored with fiber-reinforced and zirconia dowel systems. AB - STATEMENT OF PROBLEM: There are few published studies analyzing the effects of different ferrule lengths of endodontically treated teeth in relationship to newly developed fiber-reinforced and zirconia dowel systems. PURPOSE: This in vitro study compared the effect of 3 different ferrule lengths on the fracture resistance and fracture patterns of crowned endodontically treated teeth restored with 4 different esthetic dowel systems. MATERIAL AND METHODS: The crowns of 123 human maxillary canines were removed at the cementoenamel junction and the roots were endodontically treated. Three master tooth models were prepared to ferrule lengths of 1.0 mm, 1.5 mm, and 2.0 mm to produce 3 master analogs. Each root was embedded in autopolymerizing resin with a 0.2-mm layer of silicone impression material to simulate the periodontal ligament. Forty analogs of each master tooth, with ferrule lengths of 1.0 mm, 1.5 mm, and 2.0 mm were produced with copy milling (Celay system). Each group was further subdivided into 4 groups of 10 specimens each and restored with 4 different esthetic dowel systems (quartz fiber, glass fiber, glass fiber plus zirconia, and zirconia). All dowels were luted with adhesive resin cement (RelyX ARC), restored with composite cores (Valux Plus), and Ni-Cr alloy (Wiron 99) complete crowns. All specimens were loaded at 130 degrees to the long axes in a universal testing machine at a crosshead speed of 1 mm/min until fracture. Fracture patterns were classified as failures above or below the incisal third of the roots. The data were analyzed with 2-way ANOVA and Tukey HSD tests (alpha=.05). A Fisher exact test was conducted for evaluation of the mode of failure (alpha=.05). RESULTS: Mean failure loads (kg) for quartz fiber, glass fiber, glass fiber plus zirconia, and zirconia groups, respectively, with the 3 ferrule lengths were: 1.0-mm ferrule specimens: 98.09 +/- 2.90, 85.36 +/- 2.82, 80.24 +/- 1.88, 70.11 +/- 2.48; 1.5-mm ferrule specimens: 101.0 +/- 2.88, 87.58 +/- 2.83, 89.8 +/- 2.09, 82.71 +/- 2.14; 2.0-mm ferrule specimens: 119.5+/-1.78, 99.84+/-1.23, 98.6 +/- 1.64, 95.42 +/- 1.02. Teeth prepared with 2.0-mm ferrules demonstrated significantly higher fracture thresholds (P<.001). There were no significant differences in fracture patterns. CONCLUSION: Increasing the ferrule length of the endodontically treated teeth from 1 mm to 1.5 mm in specimens restored with quartz-fiber and glass-fiber dowels did not produce significant increases in the failure loads (P=.084, P=.119, respectively). No significant difference was detected between glass-fiber and glass-fiber plus zirconia dowels with 1.5-mm and 2.0-mm ferrules (P=.218, P=.244, respectively). However, fracture thresholds were higher for all 4 dowel systems when the specimens were prepared with a 2.0-mm ferrule length (P<.001). PMID- 15295327 TI - Effect of different high-palladium metal-ceramic alloys on the color of opaque and dentin porcelain. AB - STATEMENT OF PROBLEM: The color of dental porcelain depends on the type of metal substrate. Little research has been done to document the effects of different types of high-palladium alloys on the color of dental porcelain. PURPOSE: The purpose of this in vitro study was to evaluate the effects of different high palladium alloys on the resulting color of dentin porcelain, as well as on that of opaque porcelain after simulated dentin and glazing firing cycles. MATERIAL AND METHODS: Three Pd-Cu-Ga alloys, Spartan Plus (S), Liberty (B), and Freedom Plus (F), and 5 Pd-Ga alloys, Legacy (L), IS 85 (I), Protocol (P), Legacy XT (X), and Jelenko No.1 (N), were examined. A Pd-Ag alloy, Super Star (T), was included for comparison to the high-palladium alloys, and the Au-Pd alloy, Olympia (O), served as the control. Six cast discs (16 x 1 mm) were prepared from each of the alloys. Shade B1 opaque porcelain (Vita-Omega) was applied at a final thickness of 0.1 mm. After 2 opaque porcelain firing cycles, the surfaces were airborne particle abraded, and the specimens were divided into 2 groups. In the first group, 0.9 mm of B1 dentin porcelain was applied. The other group of specimens with only opaque porcelain underwent the same dentin porcelain and glazing firing cycles. Color differences (DeltaE) were determined with a colorimeter between the control and each experimental group, after the second opaque porcelain, second dentin porcelain, and glazing firing cycles. One-way analysis of variance and Dunnett's multiple range test were performed on the DeltaE data (alpha=.05). RESULTS: After the application of dentin porcelain, the 3 Pd-Cu-Ga alloys showed significantly different (P<.05) DeltaE values (S=2.3 +/- 0.5, B=1.4 +/- 0.3, and F=1.3 +/- 0.7) than the control group. After the glazing cycle of this group, the 3 Pd-Cu-Ga alloys and the Pd-Ag alloy exhibited significantly different (P<.05) DeltaE values (S=2.8 +/- 0.8, B=2.2 +/- 0.3, F=1.9 +/- 1.0, and T=1.4 +/- 0.5) than the control group. After the simulated dentin porcelain firing cycles, the specimens with only opaque porcelain exhibited significantly different (P<.05) DeltaE values (S=5.2 +/- 1.4, B=5.4 +/- 0.6, and F=3.9 +/- 0.2) than the control group. The color difference between the 3 Pd-Cu-Ga alloys with only opaque porcelain and the control group increased more after the simulated glazing cycle (S=6.6 +/- 1.5, B=6.3 +/- 0.5, and F=4.6 +/- 0.1). The observed color differences between the Pd-Ga alloys and the control group were not statistically significant at any point. CONCLUSIONS: The Pd-Cu-Ga alloys with only opaque porcelain, after the simulated dentin porcelain and glazing firing cycles, exhibited clinically unacceptable color differences. The application of dentin porcelain to the Pd-Cu Ga alloys resulted in clinically acceptable color differences. The application of dentin porcelain to the Pd-Ag alloy, after the glazing firing cycle, resulted in clinically acceptable color differences (approximately 2.8 to 3.7 DeltaE CIELAB units). The Pd-Ag alloy specimens with only opaque porcelain did not exhibit significant color differences from the control group, whereas significant color differences from the control group after the dentin porcelain and glazing firing cycles were still clinically acceptable. PMID- 15295326 TI - Microleakage of endodontically treated teeth with different dowel systems. AB - STATEMENT OF PROBLEM: Several new esthetic dowel systems are available for the restoration of endodontically treated teeth, but little is known about how effectively these dowels seal the restored teeth. PURPOSE: The purpose of this in vitro study was to compare microleakage of 3 esthetic, adhesively luted dowel systems with a conventional dowel system. MATERIAL AND METHODS: The root canals of 41 human intact single-rooted extracted teeth were prepared using a step-back technique. The teeth were randomly divided into 4 experimental groups (n=10), and 1 tooth served as a positive control. The decoronated roots were obturated with gutta-percha using lateral condensation. Roots were restored with 1 of the following dowel systems according to the manufacturer's instructions: (1) stainless steel dowels (ParaPost), (2) glass fiber dowels (Snowpost), (3) resin supported polyethylene fiber (Ribbond) dowels, or (4) zirconia dowels (Cosmopost). Using a fluid filtration method, coronal leakage of the specimens along the dowel space and root canal restorative material was measured. Fluid movement measurements were made at 2-minute intervals for 8 minutes to measure the presence of voids existing in the obturated canals, at 1 week, 3 months, and 6 months following dowel insertion. A repeated-measures analysis of variance (ANOVA) was used to analyze logarithmic transformations of data (time and dowel material) for significant differences. The Tukey HSD test and paired 2-tailed tests were used to perform multiple comparisons (alpha=.05). RESULTS: The data indicated that the leakage values varied according to the dowel system used (P<.01). There was significant interaction between dowel systems and time of testing (P<.01). The sealing ability of zirconia dowels decreased over time (P<.01), but sealing abilities of stainless steel and resin-supported polyethylene fiber dowels remained constant (P>.05). The sealing ability of glass fiber dowels increased at 3 months (P=.032) and remained constant over the next 3 months (P=.758). Statistically, resin-supported polyethylene fiber and glass fiber dowels showed the lowest coronal leakage when compared with stainless steel and zirconia dowels at all time periods (P<.01). There were no significant differences between resin-supported polyethylene fiber and glass fiber dowels at any time period. The initial leakage measurement in zirconia dowel and stainless steel dowels were similar (P=.914), but became significantly different at 3 and 6 months (P<.01). CONCLUSION: Resin-supported polyethylene fiber dowels and glass fiber dowels tested exhibited less microleakage compared to zirconia dowel systems. The latter system should be further evaluated because of its unacceptable level of leakage. PMID- 15295328 TI - Effect of fluoride and 10% carbamide peroxide on the surface roughness of low fusing and ultra low-fusing porcelain. AB - STATEMENT OF PROBLEM: The effect of repeated applications of fluoride solutions and 10% carbamide peroxide on the surface roughness of newer dental porcelains is not completely known. PURPOSE: The purpose of this study was to compare the surface roughness of 3 different porcelains when exposed to 2 fluoride solutions, a 10% solution of carbamide peroxide, and distilled water. MATERIAL AND METHODS: Forty discs (10-mm diameter, 2 mm thick) were made of each of the following porcelains: feldspathic porcelain (Ceramco II), low-fusing porcelain (Finesse), and an aluminous porcelain (All-Ceram). Each disc was abraded with a medium-grit diamond bur and auto-glazed. One side of each disc was abraded with a diamond bur and polished using a porcelain polishing kit to simulate a chairside adjustment and polishing. The discs (10 specimens/group) were immersed in 1.23% APF, 0.4% stannous fluoride, 10% carbamide peroxide, and distilled water for 50 seconds (control). The discs in the 10% carbamide peroxide solution were immersed for 48 hours. The surface of each disc was evaluated with surface profilometry (0.1 mm/s speed, 600-microm range). The data were analyzed by factorial analysis of variance and a Tukey multiple comparison test, (alpha=.05). RESULTS: The data showed that the acidulated phosphate fluoride etched the auto-glazed surface of all 3 porcelains. For Finesse specimens, the mean Ra values for the auto-glazed surface were significantly higher than that of the control after immersion in 1.23% APF (mean Ra 0.3 +/- 0.06 microm, P<.031). All-Ceram auto-glazed surface specimens had a significantly higher mean Ra value when immersed in the 3 solutions than the control (1.23% APF, 0.4% stannous fluoride, and 10% carbamide peroxide, 0.245 +/- 0.115 microm, 0.22 +/- 0.104 microm, 0.22 +/- 0.04 microm, respectively; P<.002). Ceramco II specimens were affected by all 3 solutions, with the auto-glazed surface having higher Ra values (1.23% APF, 0.4% stannous fluoride, and 10% carbamide peroxide, with mean Ra values of 0.35 +/- 0.1 microm, 0.26 +/- 0.08 microm, and 0.24 +/-.0.05 microm, respectively, P=.001). Immersion in the 3 solutions had no effect on the polished surfaces of all-ceramic specimens tested. CONCLUSION: Prior to the use of fluoride and 10% carbamide peroxide, dentists should ascertain the type of porcelain restoration present to prevent a roughened surface from occurring. PMID- 15295329 TI - Computerized tomographic evaluation of effects of mandibular anterior repositioning on the upper airway: a pilot study. AB - STATEMENT OF PROBLEM: Snoring is related to narrowing of the upper airway. Treatment options for snoring may have potential side effects or discomfort which may not be tolerated by some patients. Mandibular advancement devices may cause fewer negative side effects than other treatment modalities and have been advocated as a treatment for snoring. PURPOSE: The aim of the pilot study was to evaluate effects of the anterior mandibular positioner (AMP) on upper airway structures in patients with a snoring problem. MATERIAL AND METHODS: Fifteen subjects with snoring problems as identified by spouses or close relatives were selected to complete a questionnaire related to snoring, respiratory symptoms, body mass index (BMI), and smoking habits. Computerized tomography (CT) scans were obtained with and without AMP device fabricated as a monoblock of acrylic resin. Measurements of the oropharynx including length (mm), cross-sectional area (mm2), and plane angle (degrees) were made using anatomic landmarks on standardized CT images and lateral scanograms. The data were statistically evaluated using Wilcoxon signed rank test (alpha=.05). RESULTS: When the AMP was inserted, the cross-sectional area of the oropharynx increased by 60 mm2 on average (P<.05). A mean increase of 2.4 mm (range 1-7 mm) was observed in the posterior airway space on lateral scanograms (P<.005). The range for mandibular advancement was between 4 and 8 mm (mean 5.73 mm). CONCLUSION: The primary effect of the AMP was enlargement of the oropharyngeal airway. Within the limitations of this pilot study, the results suggest that CT evaluation of the upper airway structures may be helpful in determining treatment modality and monitoring the effectiveness of the positioner. PMID- 15295330 TI - Quantification of the relative risk of multiple occlusal variables for muscle disorders of the stomatognathic system. AB - STATEMENT OF PROBLEM: There is no consensus on the association between occlusion and temporomandibular disorders (TMD). PURPOSE: The purpose of this study was to quantify the relative risk of multiple occlusal variables for muscle disorders of the stomatognathic system. MATERIAL AND METHODS: Eight occlusal features: retruded contact position (RCP) to maximum intercuspation (MI) slide length, vertical overlap, horizontal overlap, unilateral posterior reverse articulation, anterior open occlusal relationship, incisor dental midline discrepancy, mediotrusive interferences, and laterotrusive interferences, were clinically assessed by the same trained operator. The sample consisted of 81 women with a Research Diagnostic Criteria for Temporomandibular Disorders (RDC/TMD) axis I diagnosis of muscle disorder, for example myofascial pain, with or without limited opening, and 48 healthy women (control group). A stepwise multiple logistic regression was used to identify the significant associations between occlusal features and disease. RESULTS: A slide from the retruded contact position to maximum intercuspation > or =2 mm and mediotrusive interferences were the only 2 occlusal features significantly associated with the presence of myofascial pain according to the RDC/TMD criterion symptoms. The odds ratio for myofascial pain was 2.57 for a slide from RCP to MI > or =2 mm and 2.45 for mediotrusive interferences. The percentage of the total log likelihood for myofascial pain explained by the significant occlusal factors amounted to 10.8% (Nagelkerke's R2=0.108). The multifactorial model, including the 2 significant occlusal factors, showed an acuracy to predict disease of 66.7% (sensitivity 71.6%; specificity 58.3%). CONCLUSION: Occlusal features showed a low predictive value to detect muscle disorders of the stomatognathic system. Multifactorial complex pathologies, such as TMD, should be studied using multivariate statistical analyses, as univariate models may overestimate some resulting associations. PMID- 15295332 TI - Technique for preparation of parallel guiding planes for removable partial dentures. PMID- 15295331 TI - Fabrication and use of a simple implant placement guide. AB - Proper placement and orientation of dental implants is a requirement for optimum function and esthetics to be obtained with the definitive restoration. This article describes an acrylic resin implant placement guide which is simple to fabricate and easy to use. This device guides the surgeon in the precise position and angulation planned for the implant, yet allows for some flexibility in the event slight adjustments are necessary during surgery. PMID- 15295333 TI - Simplifying the correction of the digital image in shade communication. PMID- 15295334 TI - Improving access to the lingual dental surface of a mandibular cast. PMID- 15295335 TI - Management of clinical chorioamnionitis at term. PMID- 15295336 TI - Changing times. PMID- 15295337 TI - Pelvic floor morbidity up to one year after difficult instrumental delivery and cesarean section in the second stage of labor: a cohort study. AB - OBJECTIVE: This study was undertaken to assess symptoms of pelvic floor morbidity at 6 weeks and at 1 year after difficult instrumental vaginal delivery or cesarean section during the second stage of labor. STUDY DESIGN: Prospective cohort study of 393 women with term, singleton, cephalic pregnancies who required operative delivery in surgery at full dilatation between February 1999 and February 2000. Postal questionnaires were used for follow-up at 6 weeks and at 1 year. RESULTS: Instrumental delivery was associated with a greater risk of urinary incontinence at 6 weeks and at 1-year postdelivery, adjusted odds ratio [OR] 7.8 (95% CI, 2.6-23.6) and OR 3.1 (95% CI, 1.3-7.6), respectively. Although instrumental delivery was associated with an increased risk of moderate-to-severe dyspareunia at 6 weeks, adjusted OR 3.35 (95% CI, 1.36-8.25), this difference was not significant at 1 year. Cesarean section after attempted instrumental delivery was associated with an increased risk of moderate-to-severe pain during intercourse at 1 year compared with immediate cesarean section, (18% vs 9%) P=.01. CONCLUSION: Although cesarean section at full dilatation does not completely protect women from pelvic floor morbidity, those that followed instrumental delivery had a significantly greater prevalence of urinary symptoms and dyspareunia. Urinary symptoms persist up to 1 year after delivery. PMID- 15295338 TI - Marginal maternal vitamin B12 status increases the risk of offspring with spina bifida. AB - OBJECTIVE: The purpose of this study was to investigate B vitamins and homocysteine as risk factor for offspring with spina bifida. STUDY DESIGN: Blood samples from 45 mothers and their children with spina bifida and from 83 control mothers and their children were obtained to determine the levels of serum and red blood cell folate, serum vitamin B(12), whole blood vitamin B(6), and total plasma homocysteine. RESULTS: In the case mothers, the vitamin B(12) concentration was 21% lower (95% CI, 8%-33%) compared with control mothers. Unlike folate, vitamin B(6,) and homocysteine, a vitamin B(12) concentration of .001) and diastolic (59.3 +/- 1.6 vs 64.2 +/- 1.5 mm Hg) (P=.038). Logistic regression revealed that dysmenorrhea and lower systolic blood pressure are associated with primary VVS. CONCLUSION: Women with primary and secondary VVS differ in their systemic pain perception and psychophysical characteristics. The impact of these findings on treatment modalities should be further evaluated. PMID- 15295355 TI - Translocation of lysophosphatidic acid phosphatase in response to gonadotropin releasing hormone to the plasma membrane in ovarian cancer cell. AB - OBJECTIVE: Lysophosphatidic acid mediates proliferative and/or morphologic effects on multiple cell lineages, which include ovarian cancer cells. Lysophosphatidic acid hydrolysis limits the duration of lysophosphatidic acid's action. We examined hormonal translocation of lipid phosphate phosphatase type 3 to the plasma membrane in gonadotropin-releasing hormone-responsive ovarian cancers. STUDY DESIGN: Ovarian cancers that were removed surgically and the ovarian cancer cell lines Caov-3 and SK-OV-3 were examined. Lipid phosphate phosphatase type 3 protein and activity in plasma membranes were assessed by immunohistochemical staining with lipid phosphate phosphatase type 3-specific antibodies and by the measurement of the conversion of exogenous [(3)H oleoyl]lysophosphatidic acid to mono[(3)H-oleoyl]glycerol, respectively. RESULTS: In ovarian cancers that were removed surgically, the cell surface staining and activity measurements indicated that a portion of the enzyme was localized to the plasma membrane. In Caov-3 cells and SK-OV-3 cells, lipid phosphate phosphatase type 3 protein was present both in the cytoplasm and at the plasma membrane. Treatment of the cells with a gonadotropin-releasing hormone agonist buserelin produced a rapid and progressive translocation of lipid phosphate phosphatase type 3 protein to the plasma membrane, with a concomitant loss of cytoplasmic staining. The enzyme activity in plasma membrane was also increased when the cell lines were exposed to the gonadotropin-releasing hormone agonist in intact cells before the assay of the cell membranes. CONCLUSION: These findings support the presence of lipid phosphate phosphatase type 3 in plasma membrane of ovarian cancers and provide for the ability of agonists (such as gonadotropin-releasing hormone) to induce the translocation of lipid phosphate phosphatase type 3 to plasma membrane in ovarian cancer cells. PMID- 15295356 TI - Knowledge of emergency contraception among women aged 18 to 44 in California. AB - OBJECTIVE: The State of California has taken several steps to make emergency contraceptives (ECs) available to women in the state. By using data from the 1999 2001 California Women's Health Survey, we estimated the knowledge of emergency contraception among adult women of reproductive age at risk of pregnancy (n=6209). STUDY DESIGN: This study is based on 3 years of data (1999-2001) from the California Women's Health Survey (CWHS), an annual population-based survey of more than 4000 randomly selected adult women (aged 18 years and older) in California. A total of 6198 women aged 18 to 44 responded to the 2 emergency contraception questions: "To the best of your knowledge, if a woman has unprotected sex is there anything she can do in the 3 days after intercourse that will prevent pregnancy?" and "What can she do?" RESULTS: We find that 38% of California women were able to correctly identify emergency contraception. Most importantly, the women who are most likely to need emergency contraception-those who are at risk of an unintended pregnancy but not using any method of contraception-have among the lowest levels of knowledge (only 29% identified a method of ECs). CONCLUSION: Results show that family planning providers may be reaching their clients, but broader outreach to the public has not yet achieved sufficiently high information levels among women in greatest need of the method. PMID- 15295357 TI - A new device for "no touch" biopsy at "no touch" hysteroscopy: the H Pipelle. AB - A new device for endometrial biopsy at "no touch" hysteroscopy has been developed based on the Pipelle. The modification (H Pipelle) facilitates endometrial sampling after hysteroscopy without the need to insert additional instruments into the vagina. PMID- 15295358 TI - Patient characteristics that are associated with continued pessary use versus surgery after 1 year. AB - OBJECTIVE: The purpose of this study was to identify patient characteristics in women with symptomatic pelvic organ prolapse that is associated with continued pessary use versus surgery after 1 year. STUDY DESIGN: Fifty-nine women with symptomatic pelvic organ prolapse who were satisfied with their pessary at 2 months were evaluated prospectively at 1 year. Characteristics of women who continued to use a pessary were compared with women who underwent pelvic reconstructive surgery to identify predictors for continued pessary use versus surgery. RESULTS: Forty-three women (73%) continued pessary use, and 16 women (27%) underwent surgery. Characteristics that were associated with continued pessary use were older age (76 vs 61 years; p <.001) and poor surgical risk (26% vs 0%; P =.03). Characteristics that were associated with surgery were sexual activity (81% vs 26%; P <.001), stress incontinence (44% vs 16%; P =.03), stage III-IV posterior vaginal wall prolapse (44% vs 16%; P =.03), and desire for surgery at the first visit (63% vs 12%; P <.001). Age >or=65 years was the best cut-off value for continued pessary use, with sensitivity of 95% (95% CI, 84%, 99%) and a positive predictive value of 87% (95% CI, 74%, 94%). Logistic regression demonstrated that age >or=65 years ( P <.001), stage III-IV posterior vaginal wall prolapse ( P =.007), and desire for surgery ( P =.04) were independent predictors. CONCLUSION: Age >or=65 years was associated highly with continued pessary use. Desire for surgery and stage III-IV posterior vaginal wall prolapse were associated with discontinued pessary use and pelvic reconstructive surgery. PMID- 15295359 TI - The use of Visual Analog Scale in urogynecologic research: a psychometric evaluation. AB - OBJECTIVE: The purpose of this study was to assess the validity of Visual Analog Scales in urogynecologic research. STUDY DESIGN: In phase I, 35 women completed short forms of the urogenital distress inventory, incontinence impact questionnaire, and Beck depression inventory fast screen using the Likert-type scale and Visual Analog Scale formats. Reliability was estimated with Spearman's correlations and Cronbach's alpha; construct validity was assessed with the use of factor analyses. In phase II, 101 women were recruited for the test-retest reliability assessment of the Visual Analog Scale formats of the urogenital distress inventory and incontinence impact questionnaire short forms. Reproducibility was analyzed with intraclass correlations. RESULTS: In phase I, correlations between the Likert-type scale and the Visual Analog Scale were good: urogenital distress inventory (0.748), incontinence impact questionnaire (0.787), and Beck depression inventory fast screen (0.852; P <.05). In phase II, intraclass correlations were 0.898 and 0.938 for the urogenital distress inventory and incontinence impact questionnaire scores, respectively ( P <.001). CONCLUSION: The Visual Analog Scale is a simple, reliable, and reproducible method for the assessment of quality of life in urogynecologic research. PMID- 15295360 TI - Can women without visible pubococcygeal muscle in MR images still increase urethral closure pressures? AB - OBJECTIVE: The purpose of this study was to determine if the ability to increase maximum urethral closure pressure (MUCP) with a pelvic muscle contraction is impaired in women without pubococcygeal muscle (PCM). STUDY DESIGN: This was a cross-sectional study of continent women comparing those with (n=28) and those without (n=17) PCM as identified by MR scans. A pelvic muscle contraction was performed simultaneously with recordings of urethral and bladder pressures. RESULTS: Eighty-six percent of the women with PCM compared with 41% of the women without could volitionally increase (>5 cm H(2)O) their MUCP. Those with PCM generated a mean intraurethral pressure rise of 14.0 (10.8) cm H(2)O, compared with 6.2 (8.7) cm H(2)O in those without (P=.015). Among women who could produce a visible pressure rise, there was not a statistically significant difference between groups (with PCM=17.2 [7.8] cm H(2)O; without PCM=14.7 [7.5] cm H(2)O; P=.457). CONCLUSION: Selective women without visible PCM can increase MUCP. PMID- 15295361 TI - Determinants of patient satisfaction after the tension-free vaginal tape procedure. AB - OBJECTIVE: This study was undertaken to compare patient satisfaction with subjective and objective measures of success. STUDY DESIGN: Satisfaction at 1 year was measured by using a self-administered written questionnaire. Success was assessed both subjectively and objectively by using the short form Urogenital Distress Inventory, stress testing, and multichannel urodynamics. Outcomes were compared to determine the relationship between satisfaction and success. Satisfied and dissatisfied patients were compared with respect to pre-, intra-, peri-, and postoperative characteristics. Student t and chi(2) tests were used for continuous and ordinal data, respectively. RESULTS: Of 66 (80%) patients, 53 were satisfied. These patients were more likely to achieve subjective cure (91% vs 62%, P=.009) but not statistically more likely to be objectively cured (92% vs 75%, P=.08). Dissatisfied patients tended to have overactive bladder symptoms, voiding difficulty, and required sling release. Other patient characteristics did not influence satisfaction rates. CONCLUSION: Satisfaction after tension-free vaginal tape procedure is highly dependent on normal bladder function. Dissatisfied patients deserve comprehensive evaluation, although objective measures may not reflect patient satisfaction. PMID- 15295362 TI - A randomized, double-blind, placebo-controlled comparison of the effect of nitrofurantoin monohydrate macrocrystals on the development of urinary tract infections after surgery for pelvic organ prolapse and/or stress urinary incontinence with suprapubic catheterization. AB - OBJECTIVE: The purpose of this study was to determine if antibiotic prophylaxis with nitrofurantoin monohydrate macrocrystals (study drug) after pelvic organ prolapse and/or urinary incontinence surgery with suprapubic catheterization (SPC) decreases urinary tract infection (uti) compared with placebo in a randomized, double-blind, multicenter trial. STUDY DESIGN: Six centers participated in this study. After a negative preoperative urine culture, history, surgical and postoperative course, urine culture and symptoms at SPC removal, and at 6 to 8 weeks postoperative, any other UTI and adherence were recorded. To demonstrate a 50% decrease in the bacteruria rate from 20%, with 80% power and alpha of 0.05, 438 patients were required. Data were evaluated with Student t test and Fisher exact test. RESULTS: Of 449 patients enrolled, 211 randomized to study drug, and 224 randomized to placebo. No pre- or perioperative differences existed between groups (all P>.05). Antibiotic prophylaxis decreased positive urine cultures compared with placebo (46% vs 61%, P=.002), symptomatic UTI at SPC removal (7.2% vs 19.8%, P=.001), and any other symptomatic UTI 6 to 8 weeks postoperatively (18.9% vs 32.6%, P=.002). Antibiotic prophylaxis did not decrease symptomatic UTI at the 6- to 8-week postoperative visit (1.8% vs 5.4%, P=.10). CONCLUSION: Antibiotic prophylaxis with nitrofurantoin monohydrate macrocrystals decreases UTI compared with placebo after pelvic organ prolapse and/or urinary incontinence surgery with suprapubic catheterization. PMID- 15295363 TI - Does socioeconomic status explain racial differences in urinary incontinence knowledge? AB - OBJECTIVE: This study was undertaken to discover whether urinary incontinence knowledge differs between racial/ethnic groups. If incontinence knowledge differs between racial/ethnic groups, is the difference explained by socioeconomic status (SES). STUDY DESIGN: In this cross-sectional study, 212 women from 3 counties in California were interviewed by telephone. Urinary incontinence knowledge was measured by an Incontinence Quiz. SES was calculated. Race was categorized as white or minority (non-Hispanic black, Hispanic, and other). RESULTS: White women scored better than minority women on the Incontinence Quiz (6.16 +/- 2.86 vs 5.46 +/- 2.66, P=.071). Higher SES was significantly associated with higher Incontinence Quiz total score (r=0.177, P=.010). Racial differences in Incontinence Quiz total score no longer trended toward significance after adjusting for SES in multivariable analysis. Race/ethnicity and SES were analyzed for association with each question of the Incontinence Quiz CONCLUSION: Socioeconomic status explains racial differences in total urinary incontinence knowledge. Consideration of socioeconomic status may improve the effectiveness of urinary incontinence educational programs. PMID- 15295364 TI - The effect of previous treatment experience and incontinence severity on the placebo response of stress urinary incontinence. AB - OBJECTIVE: The placebo response associated with stress urinary incontinence (SUI) is sizeable but poorly understood. The aim of this study was to examine the relationship between previous treatment experience and baseline urinary incontinence severity with placebo response. STUDY DESIGN: Nine hundred twenty one women ages 24 to 83 years received placebo in 4 12-week randomized trials evaluating duloxetine for SUI in 16 countries in Africa, Australia, Europe, and North and South America. Incontinence episode frequency (IEF) was calculated before and after randomization with entries from subject-completed real-time diaries. At baseline, subjects reported on their experience with previous continence surgery and with current pelvic floor muscle training (PFMT) with standardized questions. Analyses included Pearson correlations and the Wilcoxon two-sample test, and were based on intent-to-treat principles. RESULTS: The placebo group averaged 17 IEF per week at baseline. Fifty-five percent of placebo treated subjects averaged >or=14 IEF/wk, 11.8% had previous continence surgery, and 16.5% currently performed PFMT. The overall median decrease in IEF with placebo was 33%, but ranged from 11% to 57% for individual countries. The placebo response was lower in women with more severe SUI (29.6% vs 36.4%, P=.07), in those who had previous continence surgery (25.0% vs 33.3%, P=.26), and for those using PFMT (23.6% vs 33.3%, P=.02). There was a significant positive correlation (rho=.44; P <.0001) between the placebo response within a country and that country's use rate for PFMT. CONCLUSION: Treatment naivete and less severe incontinence are associated with an increased placebo response in clinical trials for stress urinary incontinence, although this difference was statistically significant only for PFMT. PMID- 15295365 TI - Sexual activity predicts continued pessary use. AB - OBJECTIVE: The purpose of this study was to determine which clinical variables predict continued pessary use. STUDY DESIGN: After IRB approval, charts of consecutive women at Loyola Women's Pelvic Medicine Center who bought a pessary from August 2000 to December 2002 were reviewed. Demographic information, length of pessary use, and reason for pessary discontinuation were recorded. Current pessary "users" were compared with "nonusers" (women who stopped wearing the pessary during the study period). RESULTS: Of the 136 study women, 82 (60%) were "users," and 54 (40%) were "nonusers." Women who were sexually active were more likely to continue pessary use (beta=2.204, P=.021). This was true regardless of indication for pessary placement. Women with prolapse were more likely than women with incontinence to continue with long-term pessary use (beta=2.031, P=.049). No other demographic characteristics predicted continued pessary use. CONCLUSION: Long-term pessary use is acceptable to sexually active women. Women being treated for pelvic organ prolapse are more likely to continue pessary use than women being treated for urinary incontinence. Additionally, the majority of women (60%) who accept a pessary for prolapse or urinary incontinence continue this treatment. PMID- 15295366 TI - Patient-centered goals for pelvic floor dysfunction surgery: long-term follow-up. AB - OBJECTIVE: This study was undertaken to describe long-term postoperative perceived achievement of subjective preoperative goals for pelvic floor dysfunction (PFD) surgery. STUDY DESIGN: From March 2000 through December 2001, 123 PFD surgical patients completed a preoperative open-ended questionnaire on which they described up to 5 personal outcome goals for PFD surgery. Patients were asked to review their original goals list and assess the degree to which they had met their goals on a 5-point scale (-2=strongly disagree the goal had been met to +2=strongly agree that the goal had been met) 3 months after surgery and again between 1 and 3 years after surgery. At the second follow-up, patients also completed the Incontinence Impact Questionnaire (IIQ-7) and Urogenital Distress Inventory (UDI-6) instruments to assess life impact and symptom distress, respectively. RESULTS: Of 50 women to date with long-term follow-up, 98% were white, 96% had delivered at least 1 child, 38% had previous surgery for PFD, mean weight was 74.2 +/- 14.1 kilos, and mean age was 65.4 +/- 11.5 years. Mean follow-up duration was 1.8 years, and ranged from 0.98 to 3.01 years. Of 194 goals listed by participants, 40.2% had to do with resuming previous activities or lifestyle, 38.1% with symptom relief, 9.3% with improving self-image and social relationships, 7.7% with improving general health, and 4.6% with improving physical appearance. At the individual goal level, 72% of goals were attained at short-term, and 68% attained at long-term follow-up. Long-term goal achievement did not vary significantly by category of goal. Goal achievement was lower only for symptom relief at long-term follow-up (68.9%) than at short-term follow-up (87.4%, P <.001). At the person level, 45.8% of women reported achieving all listed goals in the short term, and 42.0% in the long term. Long-term goal achievement was associated with PFD-specific quality of life (UDI-6 and IIQ-7 scores) and inversely associated with surgical complications, but was not associated with other clinical or demographic variables, including weight, parity, PFD diagnosis, psychiatric comorbidity, surgical route, or previous surgical history. CONCLUSION: Self-reported achievement of preoperatively recorded goals for PFD surgery persisted 1 to 3 years after surgery. The association of goal achievement to IIQ-7 and UDI-6 scores suggests that goal achievement is related to, but not identical to, overall measures of PFD life impact and symptom distress. Future work should examine the association of goal achievement to clinical measures of PFD severity, and compare surgically and medically managed patients. Preoperative assessment of goals may be a useful addition to clinical and subjective data in the long-term management of women with pelvic floor disorders. PMID- 15295367 TI - Sexual function after surgery for stress urinary incontinence and/or pelvic organ prolapse: a multicenter prospective study. AB - OBJECTIVE: The purpose of this study was to assess sexual function in women after surgery for stress urinary incontinence and/or pelvic organ prolapse (UI/POP) at 3 and 6 months with the Pelvic Organ Prolapse Urinary Incontinence Sexual Questionnaire (PISQ). STUDY DESIGN: Of 269 eligible women participating in a trial of prophylactic antibiotic use with suprapubic catheters, 102 (37.9%) agreed to participate in a sexual function study. Women underwent a variety of anti-incontinence and reconstructive surgeries. Sexual function and urinary incontinence were assessed preoperatively and at 3 and 6 months postoperatively with the PISQ and Incontinence Impact Questionnaires (IIQ-7). Paired t tests compared changes over time. Logistic regression compared worsening PISQ vs other variables. Generalized McNemar's test compared individual questions pre- and postoperatively. Significance was set at P <.05. RESULTS: Mean age was 47.1 (23 to 85) years, and 64% of women were premenopausal. Seventy-five (74%) women completed questionnaires at 3 or 6 months. Sexual function scores declined after surgery despite improvement in IIQ-7 scores (PISQ=86 vs 78, P <.001; IIQ-7=52 vs 13, P <.001). Behavioral Emotive domain scores worsened at 3 to 6 months compared with preoperative scores, while the Physical domain improved (all P <.001). Worsening PISQ scores were independent of age, type of surgery, hysterectomy, complications, or hormonal status (logistic regression, all P <.05). CONCLUSION: Sexual function scores in women after surgery for UI/POP do not improve despite improvement of incontinence at 3 to 6 months after surgery. PMID- 15295369 TI - Two-year infant neurodevelopmental outcome after single or multiple antenatal courses of corticosteroids to prevent complications of prematurity. AB - OBJECTIVE: This study was undertaken to evaluate the effect of exposure to multiple antenatal steroid courses on short-term neonatal morbidity and 2-year infant neurodevelopmental outcome. STUDY DESIGN: This was a prospective observational study of 201 preterm singleton infants who received 1 or more courses of corticosteroids to prevent complications of prematurity and were delivered between 24 and 34 weeks' gestation at a single institution. Neurodevelopmental outcome of the infants was evaluated at 2 years corrected age. Logistic regression analysis was used to perform multivariate analyses of associations and trends. RESULTS: One hundred thirty-eight subjects (68.7%) received at least 1 complete course of betamethasone, whereas 63 (31.3%) patients were treated with dexamethasone. The prevalence of multiple steroid doses exposure was 26.8% (37/138) in betamethasone and 52.4% (33/63) in dexamethasone group. The prevalence of infant leukomalacia, including both prolonged echogenicity and cystic leukomalacia, was 25.9% (34/131) after a complete corticosteroid course, 40% (6/15) after 1, 42.3% (12/28) after 2, and 44.4% (12/27) after more than 2 additional courses, respectively (adjusted P for trend=.011). In the same categories of steroid exposure, the corresponding prevalences of 2-year infant neurodevelopmental abnormalities were 18% (20/111), 21.4% (3/14), 29.2% (7/24), and 34.8% (8/23), respectively (adjusted P for trend=.038). Multivariate study of first grade interaction suggested that the risk of leukomalacia and 2-year infant neurodevelopmental abnormalities associated with multiple doses exposure was confined to dexamethasone. In fact, compared with betamethasone, exposure to multiple doses of dexamethasone was associated with an increased risk of leukomalacia (19/33 compared with 11/37; odds ratio [OR]=3.21, 95% CI=1.07-9.77) and overall 2-year infant neurodevelopmental abnormalities (12/28 compared with 6/35; OR=3.63, 95% CI=1.03 13.58). CONCLUSION: In this study, multiple antenatal courses of dexamethasone but not betamethasone were associated with an increased risk of leukomalacia and 2-year infant neurodevelopmental abnormalities. PMID- 15295368 TI - The Maternal-Fetal Medicine Units cesarean registry: chorioamnionitis at term and its duration-relationship to outcomes. AB - OBJECTIVE: The purpose of this study was to evaluate the relationship between chorioamnionitis and its duration to adverse maternal, fetal, and neonatal outcomes. STUDY DESIGN: This was a 13-university center, prospective observational study. All women at term carrying a singleton gestation who underwent primary cesarean from January 1, 1999 to December 31, 2000 were eligible. Data abstraction was systematic and performed by trained research nurses. Selected adverse outcomes were compared between pregnancies with, and without, clinically diagnosed chorioamnionitis using relative risks (RRs) and 95% CIs. The duration of chorioamnionitis was stratified into 5 intervals (3 6 h,>6-9 h,>9-12 h, and>12 h), and respective outcomes compared by Mantel Haenszel test for trend. Additionally, regression analysis was used to compute odds ratios (ORs) and 95% CIs for chorioamnionitis duration length as a continuous explanatory variable. RESULTS: 16,650 pregnancies were analyzed, 1965 (12%) with chorioamnionitis, which was associated with significantly increased risks of maternal blood transfusion, uterine atony, septic pelvic thrombophlebitis, and pelvic abscess (RR 2.3-3.7), as well as 5-minute Apgar 12 weeks of gestation, compared with a preoperative fetal survival rate of 12%. There were no significant intraoperative, antenatal, intrapartum or neonatal complications. CONCLUSION: Within this case series, preconception transabdominal cervicoisthmic cerclage was a safe alternative to transabdominal cervicoisthmic cerclage that was performed in pregnancy with no risk to a fetus. It should be considered in appropriate cases in women seen for prepregnancy counseling. PMID- 15295372 TI - Does fetal fibronectin use in the diagnosis of preterm labor affect physician behavior and health care costs? A randomized trial. AB - OBJECTIVE: The purpose of this study was to determine whether a knowledge of fetal fibronectin results affects patient treatment and health care costs. STUDY DESIGN: Women between 24 and 34 weeks of gestation with a singleton pregnancy and preterm uterine contractions were eligible for enrollment. Once informed consent was given, a fetal fibronectin specimen was obtained, and women were assigned randomly into 2 groups. In 1 group, results of the fetal fibronectin test were available; in the other group, results were not available. The use of inpatient and outpatient health care resources subsequent to enrollment was ascertained through the use of medical records, hospital billing data, and patient interviews. This study was powered to allow the detection in the fetal fibronectin group of a 20% reduction in total health care-related costs. RESULTS: The 2 groups were similar with respect to maternal age, parity, race, cervical examination at admission, and estimated gestational age at enrollment and at delivery. Women who did not have fetal fibronectin results available were no different than those women who did with respect to initial length of labor and delivery observation (median, 4 hours vs 3 hours), hospital admission (28% vs 26%), tocolysis (18% vs 16%), cessation of work (27% vs 26%), or total health care-related costs (log mean +/- SD, 7.6 +/- 1.2 vs 7.5 +/- 1.1). CONCLUSION: In this study population, the use of fetal fibronectin did not affect physician behavior or health care costs related to preterm contractions. PMID- 15295373 TI - Number and gestational age of prior preterm births does not modify the predictive value of a short cervix. AB - OBJECTIVE: This study was undertaken to determine whether the number and gestational age of prior preterm deliveries modifies the significance of endovaginal sonographic cervical length less than 25 mm for the prediction of recurrent preterm birth less than 35 weeks' gestation. STUDY DESIGN: Secondary analysis of a multicenter, blinded, observational study. Endovaginal ultrasonographic examinations were scheduled at 2-week intervals between 16 and 23 weeks' gestation in singleton pregnancies of 181 gravid women with at least 1 prior spontaneous preterm birth between 16 and 32 weeks' gestation. RESULTS: The earliest prior preterm birth occurred before 23 weeks in 61 women and at 23.0 to 31 weeks in 115; 5 had missing gestational age data. Cervical length was not different between these 2 groups both at the initial scan (median 38 vs 37 mm, P=.54) and considering the shortest ever observed cervical length over the entire study period (median 30 vs 30 mm, P=.97). Cervical length less than 25 mm was associated with spontaneous preterm birth less than 35 weeks for both groups (positive predictive value 80% vs 71%, P>.99). There were 134 women with 1 prior preterm delivery (74%) and 47 with 2 or more. Cervical lengths were not different between these 2 groups at the initial scan (median 36.5 vs 37 mm, P=.52) or over the entire study period (median 30 vs 32 mm, P=.31). The positive predictive value of cervical length less than 25 mm for subsequent spontaneous premature birth was not significantly higher in gravid women with multiple prior preterm births (100% vs 73%, P>.99). CONCLUSION: Neither the number nor the gestational age of prior preterm births modify the predictive value of a cervical length less than 25 mm at 16 to 19 weeks for recurrent spontaneous preterm birth. PMID- 15295374 TI - A randomized trial that compared intravaginal misoprostol and dinoprostone vaginal insert in pregnancies at high risk of fetal distress. AB - OBJECTIVE: The purpose of this study was to compare the safety and efficacy of misoprostol and dinoprostone in pregnancies at high risk of fetal distress. STUDY DESIGN: Medical indications for the induction of labor with postdate pregnancy or intrauterine growth restriction were randomized. A sequential design that was based on the triangular test was used. RESULTS: At the fourth interim analysis, which included 140 patients, the trial was stopped because no significant difference was found in neonatal safety between misoprostol and dinoprostone, which was assessed on arterial cord pH <7.20 (14.3% vs 10.0%, respectively; P=.60). Neonatal tolerance was similar in the 2 groups, with no difference in the cesarean delivery rate for fetal distress or in the incidence of meconium-stained amniotic fluid. Time to vaginal delivery was shortened by misoprostol (P=.03). CONCLUSION: Misoprostol and dinoprostone are equally safe for the induction of labor in pregnancies that are at high risk of fetal distress; however, misoprostol allowed the earlier induction of labor than did dinoprostone. PMID- 15295375 TI - Maternal morbidity and obstetric complications in triplet pregnancies and quadruplet and higher-order multiple pregnancies. AB - OBJECTIVE: The purpose of this study was to assess the risk of maternal morbidity and obstetric complications in women with triplet pregnancies and quadruplet and higher-order multiple pregnancies. STUDY DESIGN: We compared the outcomes in women with triplet pregnancies (n=5491) and quadruplet and higher-order multiple pregnancies (n=423) with women with twin pregnancies (n=152,238), with the use of the 1995 to 1997 Multiple Birth File of the United States. RESULTS: After an adjustment was made for important confounding factors, the risks of pregnancy associated hypertension and eclampsia, anemia, diabetes mellitus, abruptio placenta, premature rupture of membrane, and cesarean delivery were increased in women with triplet pregnancies and quadruplet and higher-order multiple gestations than in women with twin pregnancies. A dose-response relationship was observed for pregnancy-associated hypertension, diabetes mellitus, and placental abruption, with higher odds ratios in women with quadruplet and higher-order multiple gestations than in women with triplet pregnancies. CONCLUSION: The risks of maternal morbidity and obstetric complications are increased in triplet pregnancies and quadruplet and higher-order multiple pregnancies than in twin pregnancies; for certain outcomes, there is a dose-response relationship. PMID- 15295376 TI - The effect of early epidural versus early intravenous analgesia use on labor progression: a natural experiment. AB - OBJECTIVE: The purpose of this study was to compare the effect of early epidural with the use of early intravenous analgesia on labor progression. STUDY DESIGN: We systematically selected singleton, nulliparous term pregnancies with a spontaneous labor and analgesia placement or =100 Agatston units) were prospectively evaluated by technetium 99m sestamibi single photon emission computed tomography (SPECT). Patients were followed up (mean follow-up, 14 months) and data regarding their subsequent clinical management recorded. Of the 220 patients, 119 had moderate atherosclerosis (CAC score of 100-400 Agatston units) and 101 had severe atherosclerosis (CAC score > or =400 Agatston units). Abnormal SPECT findings were seen in 18% of patients with moderate atherosclerosis (n = 21) and 45% of patients with severe atherosclerosis (n = 45). Increasing severity of atherosclerosis was related to increasing ischemic burden (summed difference score = 1 +/- 0.2 for CAC score of 100-400 Agatston units and 3.2 +/- 0.5 for CAC score > or =400 Agatston units). In a multivariate linear regression model incorporating risk factors, CAC was the only predictor of silent ischemia. CONCLUSION: In comparison to previously published data, we detected a higher prevalence of silent ischemia even in patients with moderate coronary atherosclerosis (18%). This may reflect the differing risk factor profile of our patient population. When coronary calcium screening is used to preselect asymptomatic patients with cardiovascular risk factors for myocardial perfusion imaging, the optimum coronary calcium score threshold will depend on the population prevalence of risk factors and asymptomatic obstructive CAD. PMID- 15295415 TI - Adenosine sestamibi SPECT post-infarction evaluation (INSPIRE) trial: A randomized, prospective multicenter trial evaluating the role of adenosine Tc-99m sestamibi SPECT for assessing risk and therapeutic outcomes in survivors of acute myocardial infarction. AB - BACKGROUND: Preliminary studies indicate that adenosine myocardial perfusion single photon tomography (SPECT) can safely and accurately stratify patients into low and high risk groups early after acute myocardial infarction (AMI). METHODS AND RESULTS: INSPIRE is a prospective, randomized multicenter trial which enrolled 728 clinically stable survivors of AMI. Following baseline adenosine sestamibi gated SPECT, patients were classified as low, intermediate or high risk based on the quantified total and ischemic left ventricular (LV) perfusion defect size (PDS). A subset of high risk patients with a LV ejection fraction > or =35% were randomized to a strategy of either intensive medical therapy or coronary revascularization. Adenosine SPECT was repeated at 6-8 weeks to determine the relative effects of anti-ischemic therapies on total and ischemic PDS (primary endpoint). All patients were followed for one year. The baseline demographic, clinical and scintigraphic characteristics of the study population are presented. Adenosine SPECT was performed within 1 day of admission in 12% of patients and in 64% by Day 4. CONCLUSION: The unique study design features of INSPIRE will further clarify the role of adenosine sestamibi SPECT in defining initial patient risk after AMI and in monitoring the benefits of intensive anti-ischemic therapies. PMID- 15295416 TI - Acute rest myocardial perfusion imaging for chest pain. PMID- 15295417 TI - Positron emission tomography for quantitation of myocardial perfusion. PMID- 15295418 TI - Molecular imaging of cardiovascular gene products. PMID- 15295419 TI - The impact of caffeine on vasodilator stress perfusion studies. PMID- 15295421 TI - Myocardial perfusion imaging in a patient with chest pain. PMID- 15295420 TI - Duodenogastroesophageal reflux in a patient with postoperative esophageal cancer shown on Tc-99m tetrofosmin raw data images of dual-isotope gated cardiac SPECT. PMID- 15295553 TI - On the campaign trail. PMID- 15295554 TI - Weak at the centre. PMID- 15295555 TI - Nobel laureates spearhead effort to put Kerry in the White House. PMID- 15295556 TI - Lab closure sparks fears that US prion research is waning. PMID- 15295557 TI - Reform of land use urged as floodwaters rise across Asia. PMID- 15295558 TI - From DNA to consciousness--Crick's legacy. PMID- 15295559 TI - 'Militant' animal activists trigger British law change. PMID- 15295560 TI - Biotech funding deal judged to be 'a mistake' for Berkeley. PMID- 15295561 TI - Review of tenure refusal uncovers conflicts of interest. PMID- 15295562 TI - Court ruling upsets hopes for career reforms. PMID- 15295563 TI - Size matters when it comes to safety, report warns. PMID- 15295566 TI - A breed apart. PMID- 15295567 TI - The medals and the damage done. PMID- 15295568 TI - Heart-stopping action. PMID- 15295569 TI - Gasping for victory. PMID- 15295570 TI - Traumatic events take their toll on mental health. PMID- 15295571 TI - OvaCheck: let's not dismiss the concept. PMID- 15295572 TI - OvaCheck: doubts voiced soon after publication. PMID- 15295578 TI - The pleasure of learning. PMID- 15295579 TI - Palaeontology: inside the oldest bird brain. PMID- 15295580 TI - Semiconductor physics: the value of seeing nothing. PMID- 15295581 TI - Fisheries science: why mothers matter. PMID- 15295583 TI - Ultrafast physics: quantum control with a twist. PMID- 15295582 TI - Astronomy: a faint population of bursts? PMID- 15295584 TI - Genomes: worming into genetic instability. PMID- 15295587 TI - Biotechnology: surrogate broodstock produces salmonids. AB - A worldwide decline in the number of wild salmonids calls for strategies to restore endangered populations. Here we show that germ cells can be transplanted between two different salmonid species, with the subsequent production of xenogenic, donor-derived offspring. This pioneering xenotransplantation technology may eventually find applications in facilitating the production of commercially valuable fish, as well as in species conservation. PMID- 15295588 TI - Thin films: unexpected magnetism in a dielectric oxide. AB - It is generally accepted that magnetic order in an insulator requires the cation to have partially filled shells of d or f electrons. Here we show that thin films of hafnium dioxide (HfO2), an insulating oxide better known as a dielectric layer for nanoscale electronic devices, can be ferromagnetic even without doping. This discovery challenges our understanding of magnetism in insulators, because neither Hf4+ nor O2- are magnetic ions and the d and f shells of the Hf4+ ion are either empty or full. PMID- 15295590 TI - Mechanism of transfer RNA maturation by CCA-adding enzyme without using an oligonucleotide template. AB - Transfer RNA nucleotidyltransferases (CCA-adding enzymes) are responsible for the maturation or repair of the functional 3' end of tRNAs by means of the addition of the essential nucleotides CCA. However, it is unclear how tRNA nucleotidyltransferases polymerize CCA onto the 3' terminus of immature tRNAs without using a nucleic acid template. Here we describe the crystal structure of the Archaeoglobus fulgidus tRNA nucleotidyltransferase in complex with tRNA. We also present ternary complexes of this enzyme with both RNA duplex mimics of the tRNA acceptor stem that terminate with the nucleotides C74 or C75, as well as the appropriate incoming nucleoside 5'-triphosphates. A single nucleotide-binding pocket exists whose specificity for both CTP and ATP is determined by the protein side chain of Arg 224 and backbone phosphates of the tRNA, which are non complementary to and thus exclude UTP and GTP. Discrimination between CTP or ATP at a given addition step and at termination arises from changes in the size and shape of the nucleotide binding site that is progressively altered by the elongating 3' end of the tRNA. PMID- 15295591 TI - An apparently normal gamma-ray burst with an unusually low luminosity. AB - Much of the progress in understanding gamma-ray bursts (GRBs) has come from studies of distant events (redshift z approximately 1). In the brightest GRBs, the gamma-rays are so highly collimated that the events can be seen across the Universe. It has long been suspected that the nearest and most common events have been missed because they are not as collimated or they are under-energetic (or both). Here we report soft gamma-ray observations of GRB 031203, the nearest event to date (z = 0.106; ref. 2). It had a duration of 40 s and peak energy of >190 keV, and therefore appears to be a typical long-duration GRB. The isotropic gamma-ray energy of < or =10(50) erg, however, is about three orders of magnitude smaller than that of the cosmological population. This event--as well as the other nearby but somewhat controversial GRB 980425--is a clear outlier from the isotropic-energy/peak-energy relation and luminosity/spectral-lag relations that describe the majority of GRBs. Radio calorimetry shows that both of these events are under-energetic explosions. We conclude that there does indeed exist a large population of under-energetic events. PMID- 15295589 TI - Pathways towards and away from Alzheimer's disease. AB - Slowly but surely, Alzheimer's disease (AD) patients lose their memory and their cognitive abilities, and even their personalities may change dramatically. These changes are due to the progressive dysfunction and death of nerve cells that are responsible for the storage and processing of information. Although drugs can temporarily improve memory, at present there are no treatments that can stop or reverse the inexorable neurodegenerative process. But rapid progress towards understanding the cellular and molecular alterations that are responsible for the neuron's demise may soon help in developing effective preventative and therapeutic strategies. PMID- 15295592 TI - The sub-energetic gamma-ray burst GRB 031203 as a cosmic analogue to the nearby GRB 980425. AB - Over the six years since the discovery of the gamma-ray burst GRB 980425, which was associated with the nearby (distance approximately 40 Mpc) supernova 1998bw, astronomers have debated fiercely the nature of this event. Relative to bursts located at cosmological distance (redshift z approximately 1), GRB 980425 was under-luminous in gamma-rays by three orders of magnitude. Radio calorimetry showed that the explosion was sub-energetic by a factor of 10. Here we report observations of the radio and X-ray afterglow of the recent GRB 031203 (refs 5 7), which has a redshift of z = 0.105. We demonstrate that it too is sub energetic which, when taken together with the low gamma-ray luminosity, suggests that GRB 031203 is the first cosmic analogue to GRB 980425. We find no evidence that this event was a highly collimated explosion viewed off-axis. Like GRB 980425, GRB 031203 appears to be an intrinsically sub-energetic gamma-ray burst. Such sub-energetic events have faint afterglows. We expect intensive follow-up of faint bursts with smooth gamma-ray light curves (common to both GRB 031203 and 980425) to reveal a large population of such events. PMID- 15295593 TI - Two-dimensional geometry of spin excitations in the high-transition-temperature superconductor YBa2Cu3O6+x. AB - The fundamental building block of the copper oxide superconductors is a Cu4O4 square plaquette. The plaquettes in most of these materials are slightly distorted to form a rectangular lattice, for which an influential theory predicts that high-transition-temperature (high-T(c)) superconductivity is nucleated in 'stripes' aligned along one of the axes. This theory received strong support from experiments that indicated a one-dimensional character for the magnetic excitations in the high-T(c) material YBa2Cu3O6.6 (ref. 4). Here we report neutron scattering data on 'untwinned' YBa2Cu3O6+x crystals, in which the orientation of the rectangular lattice is maintained throughout the entire volume. Contrary to the earlier claim, we demonstrate that the geometry of the magnetic fluctuations is two-dimensional. Rigid stripe arrays therefore appear to be ruled out over a wide range of doping levels in YBa2Cu3O6+x, but the data may be consistent with liquid-crystalline stripe order. The debate about stripes has therefore been reopened. PMID- 15295594 TI - Controlling the dynamics of spontaneous emission from quantum dots by photonic crystals. AB - Control of spontaneously emitted light lies at the heart of quantum optics. It is essential for diverse applications ranging from miniature lasers and light emitting diodes, to single-photon sources for quantum information, and to solar energy harvesting. To explore such new quantum optics applications, a suitably tailored dielectric environment is required in which the vacuum fluctuations that control spontaneous emission can be manipulated. Photonic crystals provide such an environment: they strongly modify the vacuum fluctuations, causing the decay of emitted light to be accelerated or slowed down, to reveal unusual statistics, or to be completely inhibited in the ideal case of a photonic bandgap. Here we study spontaneous emission from semiconductor quantum dots embedded in inverse opal photonic crystals. We show that the spectral distribution and time-dependent decay of light emitted from excitons confined in the quantum dots are controlled by the host photonic crystal. Modified emission is observed over large frequency bandwidths of 10%, orders of magnitude larger than reported for resonant optical microcavities. Both inhibited and enhanced decay rates are observed depending on the optical emission frequency, and they are controlled by the crystals' lattice parameter. Our experimental results provide a basis for all-solid-state dynamic control of optical quantum systems. PMID- 15295595 TI - Atomic-scale imaging of nanoengineered oxygen vacancy profiles in SrTiO3. AB - At the heart of modern oxide chemistry lies the recognition that beneficial (as well as deleterious) materials properties can be obtained by deliberate deviations of oxygen atom occupancy from the ideal stoichiometry. Conversely, the capability to control and confine oxygen vacancies will be important to realize the full potential of perovskite ferroelectric materials, varistors and field effect devices. In transition metal oxides, oxygen vacancies are generally electron donors, and in strontium titanate (SrTiO3) thin films, oxygen vacancies (unlike impurity dopants) are particularly important because they tend to retain high carrier mobilities, even at high carrier densities. Here we report the successful fabrication, using a pulsed laser deposition technique, of SrTiO3 superlattice films with oxygen doping profiles that exhibit subnanometre abruptness. We profile the vacancy concentrations on an atomic scale using annular-dark-field electron microscopy and core-level spectroscopy, and demonstrate absolute detection sensitivities of one to four oxygen vacancies. Our findings open a pathway to the microscopic study of individual vacancies and their clustering, not only in oxides, but in crystalline materials more generally. PMID- 15295596 TI - Vigorous exchange between the Indian and Atlantic oceans at the end of the past five glacial periods. AB - The magnitude of heat and salt transfer between the Indian and Atlantic oceans through 'Agulhas leakage' is considered important for balancing the global thermohaline circulation. Increases or reductions of this leakage lead to strengthening or weakening of the Atlantic meridional overturning and associated variation of North Atlantic Deep Water formation. Here we show that modern Agulhas waters, which migrate into the south Atlantic Ocean in the form of an Agulhas ring, contain a characteristic assemblage of planktic foraminifera. We use this assemblage as a modern analogue to investigate the Agulhas leakage history over the past 550,000 years from a sediment record in the Cape basin. Our reconstruction indicates that Indian-Atlantic water exchange was highly variable: enhanced during present and past interglacials and largely reduced during glacial intervals. Coherent variability of Agulhas leakage with northern summer insolation suggests a teleconnection to the monsoon system. The onset of increased Agulhas leakage during late glacial conditions took place when glacial ice volume was maximal, suggesting a crucial role for Agulhas leakage in glacial terminations, timing of interhemispheric climate change and the resulting resumption of the Atlantic meridional overturning circulation. PMID- 15295597 TI - The avian nature of the brain and inner ear of Archaeopteryx. AB - Archaeopteryx, the earliest known flying bird (avialan) from the Late Jurassic period, exhibits many shared primitive characters with more basal coelurosaurian dinosaurs (the clade including all theropods more bird-like than Allosaurus), such as teeth, a long bony tail and pinnate feathers. However, Archaeopteryx possessed asymmetrical flight feathers on its wings and tail, together with a wing feather arrangement shared with modern birds. This suggests some degree of powered flight capability but, until now, little was understood about the extent to which its brain and special senses were adapted for flight. We investigated this problem by computed tomography scanning and three-dimensional reconstruction of the braincase of the London specimen of Archaeopteryx. Here we show the reconstruction of the braincase from which we derived endocasts of the brain and inner ear. These suggest that Archaeopteryx closely resembled modern birds in the dominance of the sense of vision and in the possession of expanded auditory and spatial sensory perception in the ear. We conclude that Archaeopteryx had acquired the derived neurological and structural adaptations necessary for flight. An enlarged forebrain suggests that it had also developed enhanced somatosensory integration with these special senses demanded by a lifestyle involving flying ability. PMID- 15295598 TI - Processing of wild cereal grains in the Upper Palaeolithic revealed by starch grain analysis. AB - Barley (Hordeum vulgare L.) and wheat (Triticum monococcum L. and Triticum turgidum L.) were among the principal 'founder crops' of southwest Asian agriculture. Two issues that were central to the cultural transition from foraging to food production are poorly understood. They are the dates at which human groups began to routinely exploit wild varieties of wheat and barley, and when foragers first utilized technologies to pound and grind the hard, fibrous seeds of these and other plants to turn them into easily digestible foodstuffs. Here we report the earliest direct evidence for human processing of grass seeds, including barley and possibly wheat, in the form of starch grains recovered from a ground stone artefact from the Upper Palaeolithic site of Ohalo II in Israel. Associated evidence for an oven-like hearth was also found at this site, suggesting that dough made from grain flour was baked. Our data indicate that routine processing of a selected group of wild cereals, combined with effective methods of cooking ground seeds, were practiced at least 12,000 years before their domestication in southwest Asia. PMID- 15295599 TI - Effect of trace metal availability on coccolithophorid calcification. AB - The deposition of atmospheric dust into the ocean has varied considerably over geological time. Because some of the trace metals contained in dust are essential plant nutrients which can limit phytoplankton growth in parts of the ocean, it has been suggested that variations in dust supply to the surface ocean might influence primary production. Whereas the role of trace metal availability in photosynthetic carbon fixation has received considerable attention, its effect on biogenic calcification is virtually unknown. The production of both particulate organic carbon and calcium carbonate (CaCO3) drives the ocean's biological carbon pump. The ratio of particulate organic carbon to CaCO3 export, the so-called rain ratio, is one of the factors determining CO2 sequestration in the deep ocean. Here we investigate the influence of the essential trace metals iron and zinc on the prominent CaCO3-producing microalga Emiliania huxleyi. We show that whereas at low iron concentrations growth and calcification are equally reduced, low zinc concentrations result in a de-coupling of the two processes. Despite the reduced growth rate of zinc-limited cells, CaCO3 production rates per cell remain unaffected, thus leading to highly calcified cells. These results suggest that changes in dust deposition can affect biogenic calcification in oceanic regions characterized by trace metal limitation, with possible consequences for CO2 partitioning between the atmosphere and the ocean. PMID- 15295600 TI - Aggression by polyembryonic wasp soldiers correlates with kinship but not resource competition. AB - Kin selection theory predicts that individuals will show less aggression and more altruism towards relatives. However, recent theoretical developments suggest that with limited dispersal, competition between relatives can override the effects of relatedness. The predicted and opposing influences of relatedness and competition are difficult to approach experimentally because conditions that increase average relatedness among individuals also tend to increase competition. Polyembryonic wasps in the family Encyrtidae are parasites whose eggs undergo clonal division to produce large broods. These insects have also evolved a caste system: some embryos in a clone develop into reproductive larvae that mature into adults, whereas others develop into sterile soldier larvae that defend siblings from competitors. In a brood from a single egg, reproductive altruism by soldiers reflects clone-level allocation to defence at the cost of reproduction, with no conflict between individuals. When multiple eggs are laid into a host, inter clone conflicts of interest arise. Here we report that soldier aggression in Copidosoma floridanum is inversely related to the genetic relatedness of competitors but shows no correlation with the level of resource competition. PMID- 15295601 TI - High mutation rate and predominance of insertions in the Caenorhabditis elegans nuclear genome. AB - Mutations have pivotal functions in the onset of genetic diseases and are the fundamental substrate for evolution. However, present estimates of the spontaneous mutation rate and spectrum are derived from indirect and biased measurements. For instance, mutation rate estimates for Caenorhabditis elegans are extrapolated from observations on a few genetic loci with visible phenotypes and vary over an order of magnitude. Alternative approaches in mammals, relying on phylogenetic comparisons of pseudogene loci and fourfold degenerate codon positions, suffer from uncertainties in the actual number of generations separating the compared species and the inability to exclude biases associated with natural selection. Here we provide a direct and unbiased estimate of the nuclear mutation rate and its molecular spectrum with a set of C. elegans mutation-accumulation lines that reveal a mutation rate about tenfold higher than previous indirect estimates and an excess of insertions over deletions. Because deletions dominate patterns of C. elegans pseudogene variation, our observations indicate that natural selection might be significant in promoting small genome size, and challenge the prevalent assumption that pseudogene divergence accurately reflects the spontaneous mutation spectrum. PMID- 15295602 TI - Optimal neural population coding of an auditory spatial cue. AB - A sound, depending on the position of its source, can take more time to reach one ear than the other. This interaural (between the ears) time difference (ITD) provides a major cue for determining the source location. Many auditory neurons are sensitive to ITDs, but the means by which such neurons represent ITD is a contentious issue. Recent studies question whether the classical general model (the Jeffress model) applies across species. Here we show that ITD coding strategies of different species can be explained by a unifying principle: that the ITDs an animal naturally encounters should be coded with maximal accuracy. Using statistical techniques and a stochastic neural model, we demonstrate that the optimal coding strategy for ITD depends critically on head size and sound frequency. For small head sizes and/or low-frequency sounds, the optimal coding strategy tends towards two distinct sub-populations tuned to ITDs outside the range created by the head. This is consistent with recent observations in small mammals. For large head sizes and/or high frequencies, the optimal strategy is a homogeneous distribution of ITD tunings within the range created by the head. This is consistent with observations in the barn owl. For humans, the optimal strategy to code ITDs from an acoustically measured distribution depends on frequency; above 400 Hz a homogeneous distribution is optimal, and below 400 Hz distinct sub-populations are optimal. PMID- 15295603 TI - Structural basis for template-independent RNA polymerization. AB - The 3'-terminal CCA nucleotide sequence (positions 74-76) of transfer RNA is essential for amino acid attachment and interaction with the ribosome during protein synthesis. The CCA sequence is synthesized de novo and/or repaired by a template-independent RNA polymerase, 'CCA-adding enzyme', using CTP and ATP as substrates. Despite structural and biochemical studies, the mechanism by which the CCA-adding enzyme synthesizes the defined sequence without a nucleic acid template remains elusive. Here we present the crystal structure of Aquifex aeolicus CCA-adding enzyme, bound to a primer tRNA lacking the terminal adenosine and an incoming ATP analogue, at 2.8 A resolution. The enzyme enfolds the acceptor T helix of the tRNA molecule. In the catalytic pocket, C75 is adjacent to ATP, and their base moieties are stacked. The complementary pocket for recognizing C74-C75 of tRNA forms a 'protein template' for the penultimate two nucleotides, mimicking the nucleotide template used by template-dependent polymerases. These results are supported by systematic analyses of mutants. Our structure represents the 'pre-insertion' stage of selecting the incoming nucleotide and provides the structural basis for the mechanism underlying template-independent RNA polymerization. PMID- 15295606 TI - Back to basics: from industry to academia. PMID- 15295607 TI - A long-term follow-up of 33 patients with non-Hodgkin's lymphoma who received the BEAM high-dose intensification regimen with cytokine support only and no transplant. AB - High-dose intensification and autologous stem-cell transplantation (ASCT) is widely used to consolidate patients with non-Hodgkin's lymphoma (NHL), who have reached a stage of minimal residual disease. However, patients with persisting marrow and/or blood involvement and those who fail peripheral blood hemopoietic progenitor mobilization are excluded from ASCT. For such patients with no available graft to infuse, we developed 15 years ago, before the anti-CD20 monoclonal antibody therapeutic era, the use of the BEAM pretransplant regimen followed only by the administration of three cytokines (erythropoietin, granulocyte colony-stimulating factor, granulocyte-macrophage colony-stimulating factor). We report here on the long-term follow-up of 33 patients treated with this approach. In all, 33 NHL patients underwent the BEAM (carmustine, VP-16, cytosine-arabinoside, melphalan) followed by the administration of the three cytokines from January 1994-2000. A backup marrow, albeit infiltrated by tumor cells, had been collected earlier and stored in all. A total of 30 patients (91%) recovered normal hematopoiesis. In total, 32 patients (97%) recovered neutrophils (>500/microl) at a median of 19 days and 30 patients (91%) recovered platelets (>20,000/microl) at a median of 26 days. Age, richness of backup graft and blood hemoglobin level at intensification had an impact on the time for hematopoietic recovery (P=0.014, P=0.014, P=0.048). The median follow-up was 62 months. Five patients died from toxicity related to the procedure. Eight patients relapsed and died. A total of 20 patients (61%) are alive, 16 (49%) in complete remission. A 5 year disease-free survival was 52+/-9%, relapse incidence 35+/-16%, mortality due to the procedure 12+/-12% and overall survival 61+/-10%. The BEAM regimen is not myeloablative. The BEAM+3CK procedure is a feasible therapeutic option that has shown efficacy in poor risk NHL patients who were not eligible for autografting because of persisting marrow/blood tumor contamination, or poor hemopoietic progenitor harvesting. It is unclear today whether some of these patients would have cleared their marrow/peripheral blood with the additional use of anti-CD20 treatment, thereby making the classical approach (BEAM followed by the infusion of a clean autograft) feasible. PMID- 15295608 TI - Analysis of minimal residual disease in childhood acute lymphoblastic leukemia: comparison between RQ-PCR analysis of Ig/TcR gene rearrangements and multicolor flow cytometric immunophenotyping. AB - Detection of minimal residual disease (MRD) in follow-up samples from patients with ALL is essential for evaluation of treatment response. We applied multicolor flow cytometry and real-time quantitative PCR (RQ-PCR) to compare MRD results in 71 follow-up samples from 22 children treated for ALL. When results obtained by flow cytometry and RQ-PCR were grouped into positive-negative categories, a significant level of agreement was found in 72% of samples (P<0.001). However, if a cutoff level of 0.01% was applied, the concordance was 89%. MRD could be quantified in 19 samples by both methods, showing a strong correlation (P<0.01). Nevertheless, MRD levels differed more than five-fold between both methods in 4/19 samples. In 20 (28%) samples, the two techniques showed discordant results. Most discordant results (17/20) were due to the limited sensitivity of flow cytometry analysis within the range 0.01-0.001%; remaining discordant results were due to the instable or subclonal IG/TCR gene rearrangements or a limited quantitative range of the applied RQ-PCR targets. Although concordant results could be obtained by flow cytometry and RQ-PCR analysis, MRD levels may differ. Therefore, MRD data obtained by these two techniques are not yet easily exchangeable. PMID- 15295609 TI - Comparison of skin sites for estimating serum total bilirubin in in-patients and out-patients: chest is superior to brow. AB - OBJECTIVE: [corrected] To compare transcutaneous bilirubin readings from the chest and forehead of inpatient and outpatient infants to investigate whether one site is more accurate for estimating serum bilirubin concentration. METHODS: In all, 31 infants were followed with serum and transcutaneous bilirubins using BiliChek trade mark at two skin sites. RESULTS: For inpatients average chest bilirubin was 0.4 mg/dl (7 micromol/l) higher than serum while brow was 0.3 mg/dl (5 micromol/l) lower. For outpatients, skin readings from both sites underestimated serum values. Chest estimates were 0.6 mg/dl (10 micromol/l) lower; brow was 2.1 mg/dl (36 micromol/l) lower (p<0.0001). Correlation coefficients and mean differences between skin and serum values for Hispanic and non-Hispanic infants were similar. CONCLUSIONS: In our inpatients, chest and brow readings approximated serum values. After discharge, brow readings were lower than serum values by almost 20%, while chest readings were underestimated by 5%. We recommend using the chest for transcutaneous bilirubin estimates. PMID- 15295610 TI - Attitudes of health-care providers towards research with newborn babies. AB - CONTEXT: By providing information and possibly shaping parents' preferences, health-care providers are thought to play a critical role in parental decisions to enroll their infants in research. Yet, little is known about health-care providers' beliefs about research with newborns. Previous studies suggest that parents and health-care providers are often at odds regarding attitudes towards research. OBJECTIVE: To examine the attitudes of health-care providers concerning the acceptability of research with newborn babies and the degree of research related risk to which they would be willing to expose their own infant. These findings were compared with a previous study of parental attitudes. DESIGN, SETTING AND PARTICIPANTS: A survey (pretested questionnaire with 20 scaled items and five case scenarios) of 50 doctors and 64 nurses conducted in a large tertiary care center in western Canada. RESULTS: Study limitations were a response rate of 64.5% among nurses but only 22% among physicians. Both doctors and nurses were strongly supportive of research with newborns, but nurses were more averse to exposing infants to risk. Only 76.0% of nurses, compared to 92.2% of physicians, agreed that informed consent should be sought for all forms of research. When results were compared with parental perceptions, health-care providers were more likely to believe that research should be conducted for the good of all babies. Parents were generally less aware of the existence of an approval process for research in general. In responding to hypothetical scenarios with risk and direct benefit, parents were less willing to enroll their infants than were health-care providers. Approximately 30% of both groups would be willing to enroll their infants in a study involving moderate risk and no direct benefit. CONCLUSIONS: Views of nurses, physicians, and parents regarding research with newborns are different. Overall, there is support for research; however, nurses are more likely to never enroll their own baby and enroll babies into minor studies without consent. PMID- 15295612 TI - Association of ACE gene haplotype with essential hypertension. PMID- 15295613 TI - Beneficial action of candesartan cilexetil plus amlodipine or ACE inhibitors in chronic nondiabetic renal disease. AB - Although multiple antihypertensive agents are required to control blood pressure (BP) in chronic renal disease, it remains undetermined whether the combination therapy with angiotensin receptor blockers (ARB) plus calcium antagonists or angiotensin-converting enzyme inhibitors (ACEI) confers more preferable action on renal disease than the ARB monotherapy. In the present study, we compared the effect of the combination therapy with ARB plus calcium antagonists/ACEI on proteinuria with that of the ARB monotherapy in chronic nondiabetic renal disease. At 1 month of the drug treatment, the candesartan monotherapy (n=19) reduced BP from 154+/-3/93+/-2 to 146+/-3/88+/-2 mmHg (P<0.05), and a similar magnitude of BP reductions was observed with the combination therapy with candesartan plus ACEI/amlodipine (from 153+/-2/95+/-2 to 144+/-2/88+/-2 mmHg, P<0.05, n=39). The depressor action of these therapies was sustained throughout the 12-month treatment. In contrast, the reduction in proteinuria was greater with the combination therapy (-52+/-3% at 12 months, n=39) than with the candesartan monotherapy (-25+/-3%, n=19), although the baseline values of proteinuria were nearly the same in the candesartan monotherapy group (1.74+/ 0.22 g/day) and the combination therapy group (2.10+/-0.19 g/day, P>0.2). Of note, the proteinuria-sparing effect did not differ between the candesartan+ACEI group and the candesartan+amlodipine group. In conclusion, the present study suggests more beneficial action of the combination therapy with ARB plus ACEI/amlodipine than the ARB monotherapy in nondiabetic renal disease. Since the reduction in BP was achieved to the same level, the distinct proteinuria-sparing action of these therapies is attributed to BP-independent mechanisms, which should vary depending on the agents used. PMID- 15295614 TI - Effect of Pseudomonas-induced chronic lung inflammation on specific cytotoxic T cell responses to adenoviral vectors in mice. AB - A mouse model of chronic Pseudomonas-induced bronchopulmonary inflammation that mimics chronic cystic fibrosis (CF) lung disease was employed to determine whether this inflammatory milieu influences immune responses to adenoviral vectors. Pseudomonas-infected and control mice were inoculated intranasally with a second-generation type 2 adenovirus (Ad2) vector (Ad2/betagal-2). After 3 weeks, serum and airway Ad2-specific antibodies and Ad2 vector-directed, cytotoxic T-lymphocyte (CTL) activity in splenocytes were measured. No differences in humoral immunity were observed between Pseudomonas-infected mice and controls. However, there was a two- to three-fold increase in Ad-specific CTL activity in the Pseudomonas-infected mice compared to control mice. MHC class I dependent antigen presentation by antigen-presenting cells (APC) from lungs of Pseudomonas-infected mice was also significantly increased compared to APC from control mice, suggesting a mechanism that may contribute to increased Ad-specific CD8+ CTL responses. It was concluded that Ad-specific CTL activity is enhanced in the setting of pre-existing chronic Pseudomonas-induced lung inflammation similar to CF lung disease, and that increased antigen presentation via MHC class I in this setting may be one underlying mechanism. These findings underscore the importance of considering the influence of the disease milieu when evaluating modes of gene therapy for such diseases in animal models. PMID- 15295615 TI - Protecting from R5-tropic HIV: individual and combined effectiveness of a hammerhead ribozyme and a single-chain Fv antibody that targets CCR5. AB - The CCR5 chemokine receptor is important for most clinical strains of HIV to establish infection. Individuals with naturally occurring polymorphisms in the CCR5 gene who have reduced or absent CCR5 are apparently otherwise healthy, but are resistant to HIV infection. With the goal of reducing CCR5 and protecting CCR5+ cells from R5-tropic HIV, we used Tag-deleted SV40-derived vectors to deliver several anti-CCR5 transgenes: 2C7, a single-chain Fv (SFv) antibody; VCKA1, a hammerhead ribozyme; and two natural CCR5 ligands, MIP-1alpha and MIP 1beta, modified to direct these chemokines, and hence their receptor to the endoplasmic reticulum. These transgenes were delivered using recombinant, Tag deleted SV40-derived vectors to human CCR5+ cell lines and primary cells: monocyte-derived macrophages and brain microglia. All transgenes except MIP 1alpha decreased CCR5, as assayed by immunostaining, Northern blotting, and cytofluorimetry (FACS). Individually, all transgenes except MIP-1alpha protected from low challenge doses of HIV. At higher dose HIV challenges, protection provided by all transgenes diminished, the SFv and the ribozyme being most potent. Vectors carrying these two transgenes were used sequentially to deliver combination anti-CCR5 genetic therapy. This approach gave approximately additive reduction in CCR5, as measured by FACS and protected from higher dose HIV challenges. Reducing cell membrane CCR5 using anti-CCR5 transgenes, alone or in combinations, may therefore provide a degree of protection from R5-tropic strains of HIV. PMID- 15295616 TI - Bioluminescence imaging reveals a significant role for complement in liver transduction following intravenous delivery of adenovirus. AB - The effect of complement on transgene expression was evaluated in vivo and in vitro using mice lacking complement components. Complement component 3 (C3) deficient mice (C3-/-) and appropriate wild-type controls were intravenously injected with a replication incompetent, luciferase-expressing normal Ad5 (Ad5Luc1), or fibritin-fiber Ad5 (Ad5FFLuc1). Repeated, noninvasive bioluminescence imaging was conducted over 35 days. Our data show for the first time that C3 facilitates both short- and long-term hepatic expression of luciferase following systemic delivery. C3-/- mice showed significantly less (P < 0.05) luciferase expression in their liver than treatment-matched wild-type mice when 2.3 x 10(9) (Ad5Luc1) and 4.0 x 10(9) (Ad5Luc1 or Ad5FFLuc1) viral particles (v.p.) were infused. The maximal difference in luciferase activity between C3-/- and wild-type mice was 99-fold difference at 3 days for the 2.3 x 10(9) v.p. dose (Ad5Luc1), 35-fold at 13 days for the 4.0 x 10(9) v.p. dose (Ad5Luc1), and 22 fold at 13 days for the 4.0 x 10(9) v.p. dose (Ad5FFLuc1). Preincubation of Ad5Luc1 with wild-type, C1q-/-, or factor B (FB) deficient mouse sera for 5 min significantly (P < 0.05) increased transduction of mouse liver cells, as compared to preincubation with C3-/- sera or PBS. These results suggest the classical or alternate complement pathway enhances Ad5-mediated liver transduction. PMID- 15295617 TI - Vaccination with helper-dependent adenovirus enhances the generation of transgene specific CTL. AB - Recombinant adenoviral vectors (AdV) have been used experimentally as vaccines to present antigenic transgenes in vivo. However, administration of first-generation vectors (FG-AdV) is often limited by their induction of antiviral immunity. To address this limitation, helper-dependent vectors (HD-AdV) were developed that lack viral coding regions. While the administration of HD-AdV results in long term gene expression in vivo, their utility as immunogens has never been examined. Direct vaccination with 10(8) blue-forming units (BFU) of HD-AdV injected into C57BL/6 mice lead to superior transgene-specific CTL and antibody responses when compared to the same amount of a FG-AdV. The antibody responses to viral antigens were high in response to both the vectors. As a mechanism to reduce viral exposure, dendritic cells (DC) were transduced with HD-AdV in vitro and then used as a cell-based vaccine. DC transduced with HD-AdV expressed higher levels of transgene-specific mRNA and up to 1200-fold higher levels of transgene protein than did DC transduced with a FG-AdV. In addition, HD-AdV-transduced DC stimulated superior transgene-specific CTL responses when administered in vivo, an effect that was further enhanced by maturing the DC with LPS prior to administration. In contrast to direct immunization with HD-AdV, vaccination with HD-AdV-transduced DC was associated with limited antibody responses against the AdV. We conclude that HD-AdV stimulates superior transgene-specific immune responses when compared to a FG-AdV, and that immunization with a DC-based vaccine maintains this efficacy while limiting antiviral reactivity. PMID- 15295618 TI - Gene therapy progress and prospects: electroporation and other physical methods. AB - Over the last 5 years, physical methods of plasmid delivery have revolutionized the efficiency of nonviral gene transfer, in some cases reaching the efficiencies of viral vectors. In vivo electroporation dramatically increases transfection efficiency for a variety of tissues. Other methods with clinical precedent, pressure-perfusion and ultrasound, also improve plasmid gene transfer. Alternatives such as focused laser, magnetic fields and ballistic (gene gun) approaches can also enhance delivery. As plasmid DNA appears to be a safe gene vector system, it seems likely that plasmid with physically enhanced delivery will be used increasingly in clinical trials. PMID- 15295619 TI - Brain transplantation of genetically modified bone marrow stromal cells corrects CNS pathology and cognitive function in MPS VII mice. AB - Current therapies for lysosomal storage diseases (LSDs), enzyme replacement therapy and bone marrow transplantation are effective for visceral organ pathology of LSD, but their effectiveness for brain involvement in LSDs is still a subject of controversy. As an alternative approach, we transplanted genetically modified bone marrow stromal (BMS) cells to lateral ventricle of newborn mucopolysaccharidosis VII (MPS VII) mice. MPS VII is one of LSDs and caused by deficiency of beta-glucuronidase (GUSB), resulting in accumulation of glycosaminoglycans (GAGs) in brain. At 2 weeks after transplantation, the GUSB enzyme-positive cells were identified in olfactory bulb, striatum and cerebral cortex, and the enzymatic activities in various brain areas increased. The GAGs contents in brain were reduced to near normal level at 4 weeks after transplantation. Although GUSB activity declined to homozygous level after 8 weeks, the reduction of GAGs persisted for 16 weeks. Microscopic examination indicated that the lysosomal distention was not found in treated animal brain. Cognitive function in MPS VII animals as evaluated by Morris Water Maze test in treated mice showed a marked improvement over nontreated animals. Brain transplantation of genetically modified BMS cells appears to be a promising approach to treat diffuse CNS involvement of LSDs. PMID- 15295620 TI - Messenger RNA electroporation is highly efficient in mouse embryonic stem cells: successful FLPe- and Cre-mediated recombination. AB - Development of efficient short-term gene transfer technologies for embryonic stem (ES) cells is urgently needed for various existing and new ES cell-based research strategies. In this study, we present a highly efficient, nonviral non-DNA technology for genetic loading of mouse ES cells based on electroporation of defined mRNA. Here, we show that mouse ES cells can be efficiently loaded with mRNA encoding a green fluorescent reporter protein, resulting in a level of at least 90% of transgene expression without loss of cell viability and phenotype. To show that transgenes, introduced by mRNA electroporation, exert a specific cellular function in transfected cells, we electroporated stably transfected ES cell lines with mRNA encoding FLPe or Cre recombinase proteins in order to excise an FRT- or LoxP-flanked reporter gene. The results, as determined by the disappearance and/or appearance of a fluorescent reporter gene expression, show that FLPe and Cre recombinase proteins, introduced by mRNA electroporation, efficiently exert their function without influence on further culture of undifferentiated ES cell populations and their ability to differentiate towards a specific lineage. PMID- 15295621 TI - Association of interleukin-10 promoter polymorphisms with systemic lupus erythematosus. AB - Several lines of evidence suggest interleukin-10 gene (IL-10) is a candidate gene in susceptibility to systemic lupus erythematosus (SLE). We investigated the association of IL-10 promoter single-nucleotide polymorphisms (SNPs) (-3575T/A, 2849G/A, -2763C/A, -1082A/G, -819T/C and -592A/C) and microsatellites (IL10.R, IL10.G) with SLE in 554 Hong Kong Chinese patients and 708 ethnically matched controls. Six haplotypes (hts) were identified from the SNPs. The genotype distribution of the ht1 (T-C-A-T-A), which is associated with low IL-10 production, was different in patients and controls (P=0.009). The homozygous genotype of non-ht1 was significantly increased in patients (P=0.009, odds ratio (OR)=1.80, 95% CI: 1.15-2.82). The frequency of IL10.G4 of IL10.G was also significantly increased in patients (P=0.017, OR=2.53, 95% CI: 1.18-5.40). We found that the homozygous non-ht1 combined with short allele (CA repeat number < or =21) of IL10.G has a dose-dependent effect on SLE susceptibility: non-ht1/non ht1 with homozygous short allele showed a higher OR (OR=4.11, 95% CI: 1.27-13.2, P=0.018) of association with SLE than the genotype of non-ht1/non-ht1 with heterozygous short/long allele (OR=2.98, 95% CI: 1.26-7.07, P=0.013) and homozygous long allele (OR=1.05, 95% CI: 0.62-1.78, P=0.848). The frequency of non-ht1 was significantly increased in patients with serositis (P<0.0001, OR=2.42, 95% CI: 1.55-3.80). In conclusion, the high expression promoter genotype is associated with SLE in Chinese. PMID- 15295622 TI - How should cataracts be measured? PMID- 15295623 TI - Plasma catalase activity and malondialdehyde level in patients with cataract. AB - PURPOSE: Oxidative mechanisms play a major role in the aetiology and pathogenesis of cataract, especially in age-related cataract. Our study aims to investigate systemic oxidant and antioxidant markers in cataract patients. METHODS: The activity of erythrocyte catalase and the level of malondialdehyde in plasma were measured in 40 patients with cataract and 60 healthy control subjects. The malondialdehyde level, as an index of lipid peroxidation, was determined by thiobarbituric acid reaction according to Yagi. The determination of catalase activity was measured by a method that was defined by Beutler. Catalase enzyme activity and malondialdehyde level were evaluated to find out whether there was a significant difference in these variables. Analysis of variance was used by forming a general linear model that takes age and gender as the covariate. RESULTS: CAT activity was found to be 13 920.2 +/- 847.9 U/l in cataract patients and 16 061.3 +/- 1126.6 U/l in control subjects. CAT activity in cataract patients was significantly lower than the control subjects (P = 0.008). Plasma MDA level is significantly higher in patients with cataract 4.47 +/- 0.35 nmol/ml compared to the control subjects 2.94 +/- 0.26 nmol/ml (P = 0.0001). There was no significant difference between different cataract subgroups when erythrocyte CAT activities and plasma MDA levels were compared (P = 0.322, 0.062). CONCLUSION: This study shows that oxidant/antioxidant balances alter in the presence of cataract. PMID- 15295624 TI - Tumour-lysis-related elevation of intraocular pressure following high-dose-rate brachytherapy for choroidal melanoma. PMID- 15295625 TI - Divergent mechanisms for the tuning of shortwave sensitive visual pigments in vertebrates. AB - Of the four classes of vertebrate cone visual pigments, the shortwave-sensitive SWS1 class shows the shortest lambda(max) values with peaks in different species in either the violet (390-435 nm) or ultraviolet (around 365 nm) regions of the spectrum. Phylogenetic evidence indicates that the ancestral pigment was probably UV-sensitive (UVS) and that the shifts between violet and UV have occurred many times during evolution. This is supported by the different mechanisms for these shifts in different species. All visual pigments possess a chromophore linked via a Schiff base to a Lys residue in opsin protein. In violet-sensitive (VS) pigments, the Schiff base is protonated whereas in UVS pigments, it is almost certainly unprotonated. The generation of VS from ancestral UVS pigments most likely involved amino acid substitutions in the opsin protein that serve to stabilise protonation. The key residues in the opsin protein for this are at sites 86 and 90 that are adjacent to the Schiff base and the counterion at Glu113. In this review, the different molecular mechanisms for the UV or violet shifts are presented and discussed in the context of the structural model of bovine rhodopsin. PMID- 15295626 TI - Regulatory pathways in photodynamic therapy induced apoptosis. AB - Photodynamic therapy is an approved treatment for several types of tumors and certain benign diseases, based on the use of a light-absorbing compound (photosensitizer) and light irradiation. In the presence of molecular oxygen, light-activation of the photosensitizer, which accumulates in cancer tissues, leads to the local production of reactive oxygen species that kill the tumor cells. Mitochondria are central coordinators of the mechanisms by which PDT induces apoptosis in the target cells. Recent studies indicate that concomitant to the permeabilization of the outer mitochondrial membrane (which leads to the release of several apoptogenic factors in the cytosol and to the activation of effector caspases), regulatory signaling pathways are activated in a photosensitizer, PDT dose and cell-dependent fashion. Signaling pathways regulated by members of mitogen activated protein kinases and their downstream targets, such as cyclooxygenase-2, appear to critically modulate cancer cell sensitivity to PDT. Understanding the molecular events that contribute to PDT induced apoptosis, and how cancer cells can evade apoptotic death, should enable a more rationale approach to drug design and therapy. PMID- 15295627 TI - Protein oxidation in plant mitochondria as a stress indicator. AB - Plant mitochondria produce reactive oxygen species (ROS) as an unavoidable side product of aerobic metabolism, but they have mechanisms for regulating this production such as the alternative oxidase. Once produced, ROS can be removed by several different enzyme systems. Finally, should the first two strategies fail, the ROS produced can act as a signal to the rest of the cell and/or cause damage to DNA, lipids and proteins. Proteins are modified in a variety of ways by ROS, some direct, others indirect e.g. by conjugation with breakdown products of fatty acid peroxidation. Reversible oxidation of cysteine and methionine side chains is an important mechanism for regulating enzyme activity. Mitochondria from both mammalian and plant tissues contain a number of oxidised proteins, but the relative abundance of these post-translationally modified forms is as yet unknown, as are the consequences of the modification for the properties and turnover time of the proteins. Specific proteins appear to be particularly vulnerable to oxidative carbonylation in the matrix of plant mitochondria; these include several enzymes of the Krebs cycle, glycine decarboxylase, superoxide dismutase and heat shock proteins. Plant mitochondria contain a number of different proteases, but their role in removing oxidatively damaged proteins is, as yet, unclear. PMID- 15295628 TI - The suppression of immunity by ultraviolet radiation: UVA, nitric oxide and DNA damage. AB - We have examined the mechanism by which solar-simulated ultraviolet radiation (ssUV) suppresses memory immunity to nickel in allergic humans. In initial studies, we used inbred mice to determine the contribution of different wavebands to sunlight-induced immunosuppression. We found that low dose UVA can enhance memory, medium dose UVA (half the amount in one minimum erythemal dose of ssUV) is immunosuppressive, but higher doses protect from UVB. This is genetically dependent, as it is not observed in all mouse strains. UVA caused a similar dose related change in recall immunity in humans. ssUV dose responses determined the limits of protection provided by sunscreens from immunosuppression in humans. Immune protection factors calculated from these data correlated with UVA protection, but not with sun protection factor, showing that in commercial sunscreens that provide good UVB protection, UVA protection limits prevention of immunosuppression. N(G)-monomethyl-l-arginine acetate (l-NMMA) was used to inhibit nitric oxide (NO) production and T4N5 liposomes containing T4 endonuclease V to enhance DNA repair. Sub-erythemal ssUV caused a dose-related local suppression of recall immunity to nickel in humans. l-NMMA and the liposomes protected the nickel reaction, suggesting that NO and DNA damage are mediators of UV-induced immunosuppression in humans. PMID- 15295629 TI - Photochemical and electrophysical production of radicals on millisecond timescales to probe the structure, dynamics and interactions of proteins. AB - The reaction of hydroxyl and other oxygen-based radicals with the side chains of proteins on millisecond timescales has been used to probe the structure of proteins, their dynamics in solution and interactions with other macromolecules. Radicals are generated in high flux within microseconds from synchrotron radiation and discharge sources and react with proteins on timescales that are less than those often attributed to structural reorganisation and folding. The oxygen-based radicals generated in aqueous solution react with proteins to effect limited oxidation at specific amino acids throughout the sequence of the protein. The extent of oxidation at these residue markers is highly influenced by the accessibility of the reaction site to the bulk solvent. The extent of oxidation allows protection levels to be measured based on the degree to which a reaction occurs. A map of a protein's three-dimensional structure is subsequently assembled as in a footprinting experiment. Protein solutions that contain various concentrations of substrates that either promote or disrupt structural transitions can be investigated to facilitate site-specific equilibrium and time resolved studies of protein folding. The radical-based strategies can also be employed in the study of protein-protein interactions to provide a new avenue for investigating protein complexes and assemblies with high structural resolution. The urea-induced unfolding of apomyoglobin, and the binding domains within the ribonuclease S and calmodulin-melittin protein-peptide complexes are presented to illustrate the approach. PMID- 15295630 TI - Carotenoids and UV protection. AB - Photooxidative processes play a role in the pathobiochemistry of various disorders of light-exposed tissue. After irradiation of skin with UV light, erythema (sunburn) is an initial effect suitable for monitoring the direct biological response. Carotenoids are efficient in photoprotection, scavenging singlet oxygen and peroxyl radicals. Intervention studies with supplements or a carotenoid-rich diet documented efficiency in systemic photoprotection, measuring a decreased sensitivity against UV-induced erythema. For successful intervention, treatment with carotenoids is needed for a period of at least ten weeks. An increased consumption of carotenoids may contribute to life-long protection against UV-induced damage. PMID- 15295631 TI - Pro-carcinogenic activity of beta-carotene, a putative systemic photoprotectant. AB - Beta-carotene is a strong singlet oxygen quencher and antioxidant. Epidemiologic studies have implied that an above average intake of the carotenoid might reduce cancer risks. Earlier studies found that the carotenoid, when added to commercial closed-formula rodent diets, provided significant photoprotection against UV carcinogenesis in mice. Clinical intervention trials found that beta-carotene supplementation evoked no change in incidence of nonmelanoma skin cancer. However, when smokers were supplemented with the carotenoid a significant increase in lung cancer resulted. Recently, employing a beta-carotene supplemented semi-defined diet, not only was no photoprotective effect found, but significant exacerbation of UV-carcinogenesis occurred. Earlier, a mechanism, based upon redox potential of interacting antioxidants, was proposed in which beta-carotene participated with vitamins E and C to efficiently repair oxy radicals and, thus, thought to provide photoprotection. In this schema, alpha tocopherol would first intercept an oxy radical. In terminating the radical propagating reaction, the tocopherol radical cation is formed which, in turn, is repaired by beta-carotene to form the carotenoid radical cation. This radical is repaired by ascorbic acid (vitamin C). As the carotenoid radical cation is a strongly oxidizing radical, unrepaired it could contribute to the exacerbating effect on UV-carcinogenesis. Thus, vitamin C levels could influence the levels of the pro-oxidant carotenoid radical cation. However, when hairless mice were fed beta-carotene supplemented semi-defined diet with varying levels of vitamin C (0 5590 mg kg(-1) diet) no effect on UV-carcinogenesis was observed. Lowering alpha tocopherol levels did result in further increase of beta-carotene exacerbation, suggesting beta-carotene and alpha-tocopherol interaction. It was concluded that the non-injurious or protective effect of beta-carotene found in the closed formula rations might depend on interaction with other dietary factors that are absent in the semi-defined diet. At present, beta-carotene use as a dietary supplement for photoprotection should be approached cautiously. PMID- 15295632 TI - The photoreactivity of ocular lipofuscin. AB - Lipofuscin or "age pigment" is a lipid-protein complex which accumulates in a variety of postmitotic, metabolically active cells throughout the body. These complexes, which are thought to result from the incomplete degradation of oxidised substrate, have the potential for photoreactivity. This is particularly so in the retina in which the lipofuscin not only contains retinoid metabolites but is also exposed to high oxygen and fluxes of visible light all of which provide an ideal environment for the generation of reactive oxygen species (ROS). Lipofuscin is a potent photoinducible generator of ROS with the potential to damage proteins, lipids and DNA. Retinal cell dysfunction may be strongly associated with photoreactivity of lipofuscin and may contribute to age-related disease and vision loss. PMID- 15295633 TI - Targeting of the vascular system of solid tumours by photodynamic therapy (PDT). AB - Owing to morphological and rheological differences of the tumour vascular system as compared to the vascular system of the surrounding tissue, the efficacy of several experimental and clinical therapeutic approaches is limited. This fact has put the vascular system of solid tumours into focus and two new therapeutic strategies, anti-angiogenesis and vascular targeting, have emerged. Under the term vascular targeting various therapeutic approaches are summarized, e.g. chemoembolization, chemotherapy, hyperthermia, vascular targeting agents (VTA) and photodynamic therapy (PDT). As shown using the clinically approved photosensitiser Photofrin the irreversible destruction of the tumour vascular system is primarily responsible for an effective PDT of solid tumours. However, the clinical disadvantages of Photofrin are well known. Thus, several new photosensitisers, e.g. aminolaevulinic acid (ALA), porphycenes and indocyanine green (ICG), have been evaluated in vitro and in vivo regarding their suitability for vascular targeting of solid tumours. The promising experimental findings with the photosensitiser ICG led to first clinical results in treating Kaposi's sarcomas. In summary, systemic PDT is only effective when leading to complete ischaemia of solid tumours with subsequent necrosis. An essential prerequisite is the use of a chemically and photophysically defined photosensitiser localizing in the intravascular space due to e.g. a high molecular weight. The specific properties of such a photosensitiser are outlined. PMID- 15295634 TI - Hypericin as a potential phototherapeutic agent in superficial transitional cell carcinoma of the bladder. AB - Photodynamic therapy (PDT) involves the local or systemic administration of a photosensitizing drug that, upon light irradiation and presence of oxygen, results in tissue damage such as tumor destruction. Hypericin, a hydroxylated phenanthroperylenequinone, is obtained from Hypericum perforatum plants. Hypericin exhibits a high fluorescence quantum yield, and its presence in the tissue can easily be visualized. Interestingly, when instilled into the human bladders, hypericin selectively accumulates in the bladder carcinoma lesions, with the specificity and sensitivity of detecting CIS reaching up to 98.5 and 93%, respectively. Due to this selective accumulation of hypericin in bladder carcinoma lesions, the compound is now used as a fluorescent diagnostic tool for superficial bladder cancer. However, hypericin is also a photosensitizer with a potent photocytotoxic activity. Taken together, these data indicate that hypericin could be used for whole bladder wall PDT of superficial bladder tumors. This review focuses on the more recent in vitro and in vivo evaluation of hypericin as a photodynamic agent in the treatment of superficial transitional cell carcinoma (TCC) bladder tumors. PMID- 15295635 TI - Biological UV dosimetry using the DLR-biofilm. AB - Changes of environmental UV radiation as part of global atmospheric changes will influence the biosphere substantially. The determination of the biological effects of these changes requires accurate and reliable UV monitoring systems that weight the spectral irradiance according to the biological responses under consideration. Biological UV dosimeters, which directly weight the incident UV components of sunlight in relation to the effectiveness of the different wavelengths and the potential interactions between them, can complement weighted physical UV measurements. Up to now several UV-dependent endpoints in biomolecules (e.g. uracil, DNA, provitamin D3), bacteriophages (e.g. T7), bacteria (e.g.E. coli, B. subtilis) and cultured eukaryotic cells have been suggested as sensing elements in biological UV dosimeters. One example is the DLR biofilm consisting of immobilised spores of the bacterium B. subtilis as a UV sensor. It weights per se the incident UV radiation according to its DNA-damaging effectiveness. In several examples the applicability of the DLR-biofilm technique for personal UV dosimetry as well as for the measurement of the biologically weighted irradiance of the sun and of artificial UV sources is demonstrated. PMID- 15295636 TI - The digital photobiology compendium: perspectives on a web-based teaching tool from learners and developers. AB - The DPC offers many benefits for learners, teachers and developers involved in creation of teaching materials in photobiology. Modifications and additions can be made relatively easily. Anonymous peer review of modules, allowing them to be cited as peer reviewed publications, is likely to encourage new submissions. PMID- 15295637 TI - Time-domain fluorescence lifetime imaging applied to biological tissue. AB - Fluorescence lifetime imaging (FLIM) is a functional imaging methodology that can provide information, not only concerning the localisation of specific fluorophores, but also about the local fluorophore environment. It may be implemented in scanning confocal or multi-photon microscopes, or in wide-field microscopes and endoscopes. When applied to tissue autofluorescence, it reveals intrinsic excellent contrast between different types and states of tissue. This article aims to review our recent progress in developing time-domain FLIM technology for microscopy and endoscopy and applying it to biological tissue. PMID- 15295638 TI - Are dietary carotenoids beneficial? Reactions of carotenoids with oxy-radicals and singlet oxygen. AB - Carotenoids play diverse roles in biology and medicine. Both the quenching of singlet oxygen (energy transfer) and interaction with oxy-radicals (electron transfer, H-atom transfer and addition reactions) are key processes in understanding many of these roles. Much previous work in 'simple' solvents is reviewed and new results in cell membrane models are presented. The possible consequences of using carotenoids as dietary supplements are discussed. PMID- 15295639 TI - Adjuvant treatment for complement activation increases the effectiveness of photodynamic therapy of solid tumors. AB - Phototoxic lesions generated in tumor tissue by photodynamic therapy (PDT) are recognized by the host as a threat to the integrity and homeostasis at the affected site. Among the canonical pathways invoked by the host for dealing with this type of challenge is the activation of the complement system, integrating proteins that serve as molecular sensors of danger signals produced by PDT and those initiating signalling cascades coupled into the network of inflammatory and immune responses. Since the activated complement system is a salient participant of the antitumor response produced by PDT, it is worth exploring whether its manipulation can be exploited for the therapeutic benefit. Using mouse tumor models, the present study examined the potential of representative complement activating agents to act as effective adjuvants to PDT. Tumor-localized treatment with zymosan, an alternative complement pathway activator, reduced the recurrence rate of PDT-treated tumors, markedly increasing the percentage of permanent cures. In contrast, a similar treatment with heat aggregated gamma globulin (complement activator via the classical pathway) was of no significant benefit as a PDT adjuvant. Systemic complement activation with streptokinase treatment had no detectable effect on complement deposition at the tumor site without PDT, but it augmented the extent of complement activity in PDT-treated tumors. This finding based on immunohistochemistry analysis explains the results of tumor therapy experiments, which showed that systemic treatment with streptokinase or a similar agent, urokinase, enhances the PDT-mediated tumor response. Zymosan and streptokinase administrations produced no beneficial results with PDT of tumors growing in complement-deficient mice. This study, therefore, establishes the potential of complement-activating agents to serve as effective adjuvants to PDT for cancer treatment. PMID- 15295640 TI - Laser-assisted fluorescence microscopy for measuring cell membrane dynamics. AB - Membranes of living cells are characterized by laser-assisted fluorescence microscopy, in particular a combination of microspectrofluorometry, total internal reflection fluorescence microscopy (TIRFM), fluorescence lifetime imaging (FLIM) and Forster resonance energy transfer (FRET) spectroscopy. The generalized polarization (GP, characterizing a spectral shift which depends on the phase of membrane lipids) as well as the effective fluorescence lifetime (tau(eff)) of the membrane marker laurdan were revealed to be appropriate parameters for membrane stiffness and fluidity. GP decreased with temperature, but increased during cell growth and was always higher for the plasma membrane than for intracellular membranes. Microdomains of different fluorescence lifetimes tau(eff) were observed at temperatures above 30 degree C and disappeared during cell aging. Non-radiative energy transfer was used to detect laurdan selectively in close proximity to a molecular acceptor (DiI) and may present a possibility for measuring membrane dynamics in specific microenvironments. PMID- 15295641 TI - Early molecular events in the photoactive yellow protein: role of the chromophore photophysics. AB - We report a comparative study of the isomerization reaction in native and denatured photoactive yellow protein (PYP) and in various chromophore analogues in their trans deprotonated form. The excited-state relaxation dynamics was followed by subpicosecond transient absorption and gain spectroscopy. The free p hydroxycinnamate (pCA(2-)) and its amide analogue (pCM(-)) are found to display a quite different transient spectroscopy from that of PYP. The excited-state deactivation leads to the formation of the ground-state cis isomer without any detectable intermediate with a mechanism comparable to trans-stilbene photoisomerization. On the contrary, the early stage of the excited-state deactivation of the free thiophenyl-p-hydroxycinnamate (pCT(-)) and of the denatured PYP is similar to that of the native protein. It involves the formation of an intermediate absorbing in the spectral region located between the bleaching and gain bands in less than 2 ps. However, in these two cases, the formation of the cis isomer has not been proved yet. This difference with pCA(-) and pCM(-) might result from the fact that, in the thioester substituted chromophore, simultaneous population of two quasi-degenerate excited states occurs upon excitation. This comparative study highlights the determining role of the chromophore structure and of its intrinsic properties in the primary molecular events in native PYP. PMID- 15295643 TI - Identification and characterization of hmr19 gene encoding a multidrug resistance efflux protein from Streptomyces hygroscopicus subsp. yingchengensis strain 10 22. AB - The hmr19 gene was cloned from Streptomyces hygroscopicus subsp. yingchengensis strain 10-22, a bacterium strain producing agricultural antibiotics. Sequence similarity comparison indicates that hmr19 gene may encode a predicted protein with 14 putative transmembrane alpha-helical spanners, belonging to the drug:H(+) antiporter-2 family of the major facilitator superfamily. The expression of hmr19 in the mycelium of strain 10-22 was detected by Western blotting analysis. Gene replacement technology was employed to construct an hmr19 disruption mutant. The growth inhibition test against different antibiotics indicated that the mutant strain was 5-20 fold more susceptible to tetracycline, vancomycin and mitomycin C than the parental wild type strain. The mutant took up tetracycline much faster and accumulated more antibiotics than the wild type strain 10-22. While with the addition of an energy uncoupler, carbonyl cyanide m-chlorophenylhydrazone, the characteristics of the accumulation of [(3)H]tetracycline in these two strains were almost the same. It was thus concluded that hmr19 encoded a multidrug resistance efflux protein. PMID- 15295644 TI - Influence of trace amount of calponin on smooth muscle myosin in different states. AB - Calponin (CaP), a thin filament-associated protein, is thought to be involved in modulating smooth muscle contractile activity, but the role and mechanism keep unknown. In this study, trace amount of calponin (TAC) was found to obviously influence myosin in different states in Ca(2+)-independent manner, suggesting a high efficient interaction between TAC and myosin. In this assay, the lowest ratio of CaP vs. myosin was 1:10,000, with the concentration of CaP 10,000-fold lower than that used previously. Myosin phosphorylation, myosin Mg(2+)-ATPase activity and protein binding activity were detected to determine the effects of TAC on the myosin in different states. The amount of precipitated myosin that bound to TAC was used as the index to determine the interaction between myosin and TAC in binding assay. Our data showed that in the absence of actin, TAC significantly increased the precipitation of unphosphorylated myosin, Ca(2+) dependently or independently phosphorylated myosin by MLCK, and stimulated the Mg(2+)-ATPase activities of these myosins slightly but significantly. However, no obvious change of precipitation of myosin phosphorylated by PKA was observed, indicating the relatively selective effect of TAC. In the presence of actin, the increase of myosin precipitations was abolished, and no obvious change of actin precipitations and actin-activated myosin Mg(2+)-ATPase activities were observed implicating the high efficiency of TAC on myosin being present in the absence of actin. Although we can not give conclusive comments to our results, we propose that the high efficiency of TAC-myosin interaction is present when actin is dissociated from myosin, even if CaP/myosin ratio is very low; this high efficient interaction can be abolished by actin. However, why and how TAC can possess such a high efficiency to influence myosin and how the physiological significance of the high efficiency of TAC is in regulating the interaction between myosin and actin remain to be investigated. PMID- 15295645 TI - Antisense Tiam1 down-regulates the invasiveness of 95D cells in vitro. AB - As a specific guanine nucleotide exchange factor of Rac1, Tiam1 (T-lymphoma invasion and metastasis inducing protein 1) is involved in a number of cellular events, such as cytoskeleton reorganization, cell adhesion, and cell migration. Since Tiam1 was implicated in the invasion and metastasis of T-lymphoma cells and breast tumor cells, we compared the expression level of Tiam1 in two human giant cell lung carcinoma cell strains with high or low metastasis potential, and found that Tiam1 expression level in high-metastatic 95D cells was higher than that in low-metastatic 95C cells. To further confirm the role of Tiam1 in invasion and metastasis, we constructed the antisense Tiam1 expression plasmid (pcDNA3-anti Tiam1), which was transfected into 95D cells. A stable transfected clone with decreased Tiam1 expression was screened and selected for further research. Transwell assay showed that down-regulation of endogenous Tiam1 by anti-Tiam1 can reduce the in vitro invasiveness of 95D cells. Our results suggested that Tiam1 signaling contributed to the invasion and metastasis of the human giant-cell lung carcinoma cells. PMID- 15295646 TI - Recombinant scFv antibodies against E protein and N protein of severe acute respiratory syndrome virus. AB - Three single chain antibodies (scFv) against the proteins of severe acute respiratory syndrome coronavirus (SARS-CoV) were isolated by phage display from an scFv antibody library. Bio-panning was carried out against immobilized purified envelope (E) and nucleocapsid (N) proteins of SARS-CoV. Their binding activity and specificity to E or N protein of SARS-CoV were characterized by phage-ELISA. Two of them, B10 and C20, could recognize non-overlapping epitopes of the E protein according to the two-site binding test result. Clone A17 could recognize N protein. The sequence of the epitope or overlapping epitope of scFv antibody A17 was PTDSTDNNQNGGRNGARPKQRRPQ. The affinity (equilibrium dissociation constant, K(d)) of SARS-CoV E protein was 5.7 x 10(-8) M for B10 and 8.9 x 10(-8) M for C20. The affinity of A17 for N protein was 2.1 x 10(-6) M. All three scFv antibodies were purified with affinity chromatography and determined by Western blot. PMID- 15295647 TI - Highly efficient and economical baculovirus expression system for preparing human papillomavirus type16 virus-like particle. AB - To improve the existing human papillomavirus type16 (HPV16) virus-like particle (VLP) preparation, a highly efficient, economical and timesaving system was established. Sf-9 cells were infected with recombinant baculovirus containing the target gene encoding HPV16L1 protein with 6xHis tag, and harvested 72 h postinfection (p.i.) at 27 degrees. The ProBond(TM) purification system was used for protein purification. The molecular weight of expressed HPV16L1 protein was 58 kD as revealed by SDS-PAGE, and confirmed by Western blot. The purity of denatured and native HPVL1 proteins that were prepared were 91.9% and 71.5%, respectively, which corresponded to a yield of 2.26 mg denatured protein and 1.84 mg native protein per 2x10(7) cells. The proteins were further analyzed by mouse erythrocyte hemagglutination assay and hemagglutination inhibition assay, and there effects on VLP formation were also visualized by transmission electron microscopy. Results showed that the native protein purified was biologically active as natural HPVL1 protein, inducing the murine erythrocyte agglutination and VLP formation. In addition, the purified recombinant HPV16L1 native protein with 6xHis tag could self-assemble into virions in vitro. Hopefully, the present expression and purification system is promising to be convenient, timesaving and economical for preparation of HPV16 VLP vaccine. PMID- 15295648 TI - Signal peptide of potato PinII enhances the expression of Cry1Ac in transgenic tobacco. AB - The modified Cry1Ac was expressed in transgenic tobacco plants. To allow secretion of the Cry1Ac protein into the intercellular space, the signal peptide sequence of potato proteinase inhibitor II (pinII) was N-terminally fused to the Cry1Ac encoding region. Expression of Cry1Ac in transgenic tobacco plants was assayed with ELISA. The results showed that pinII signal peptide sequence enhanced the expression of Cry1Ac protein and led to the secretion of the Cry1Ac protein in transgenic tobacco plants. GFP gene was also fused to the signal peptide sequence and transformed to tobacco. The results of fluorescent detection showed that GFP had localized in the apoplast of transgenic plants. PMID- 15295649 TI - Inhibiting apoptosis of CTLL-2 cells to enhance their GVL effects via anti-Fas ribozyme. AB - To investigate the inhibition role of anti-Fas hammerhead ribozyme on fas expression and Fas-mediated apoptosis of CTL cell line CTLL-2 cells, the cDNA of an anti-Fas hammerhead ribozyme was synthesized, its expression plasmid was constructed and transfected into CTLL-2 cells by electroporation. fas expression of CTLL-2 cells was detected by RT-PCR and Western blot. CTLL-2 cell viability was measured using MTT assay when co-cultured with mouse T cell leukemia cell line EL4 cells that highly expressed Fas ligand (FasL). Meanwhile, caspase-3 proteolytic activity was detected, and cell apoptosis was measured by flow cytometry and Hochest-PI double staining. Killing activity of CTLL-2 cells was detected by lactate dehydrogenase (LDH) releasing assay in vitro. Results showed that the expression of both Fas mRNA and protein in CTLL-2 cells were decreased after transfection of anti-Fas ribozyme. Compared with mock-transfected group and mutant ribozyme-transfected group, viability of CTLL-2 cells co-cultured with EL4 cells was increased significantly and cells killing activity was enhanced after transfected with anti-Fas ribozyme, while the caspase-3 activity and apoptosis rate was significantly decreased. The results demonstrated anti-Fas ribozyme could efficiently cleave Fas and inhibit Fas-mediated apoptosis of CTLL-2 cells to improve their viability. Our study made a basis for enhancing CTLL-2 cells anti-leukemia effect in DLI. PMID- 15295650 TI - Cloning, expression, purification and crystallization of NHR3 domain from acute myelogenous leukemia-related protein AML1-ETO. AB - The t(8;21) translocation is one of the most frequent chromosome abnormalities in acute myeloid leukemia. This translocation creates a fusion between the acute myelogenous leukemia 1 (AML1, a transcription factor) gene on chromosome 21 and the eight-twenty-one (ETO, a zinc finger nuclear protein) gene on chromosome 8, leading to the repression of certain AML1 target genes. We cloned NHR3 domain coding fragment into vector pGEX-6p-1 using PCR and obtained recombinant plasmid pGEX-6p-1-NHR3, which can be induced to stably overexpress fusion protein in E. coli. Through the purification on GST affinity chromatography column and PreScission protease cleavage, a large amount of NHR3 protein with high purity was obtained. In order to avoid possible interference of some strong negative charged molecules, NHR3 protein was further purified by Mono Q anion exchange chromatography. The NHR3 crystals were grown with hanging drop/vapor diffusion method and the first crystals appeared after four weeks at 18 degrees in 0.2 M Tris-sodium citrate dihydrate, 0.1 M sodium cacodylate, pH 6.5, and 30% iso propanol (V/V). ESI mass spectrum showed that the molecular weight of this domain was in agreement with its primary structure sequence prediction, and circular dichroism spectral data (190-250 nm) showed that NHR3 had a well-defined secondary structure of 25.9% alpha-helix, 23.2% random coil and 50.9% turn, which was consistent with GOV4 software prediction. PMID- 15295651 TI - An effective method for raising antisera against beta-defensins: double-copy protein expression of mBin1b in E. coli. AB - Bin1b is a rat epididymis specific beta-defensin which may have fertility related functions in addition to its antimicrobial activity. beta-defensins are cysteine rich cationic antimicrobial peptides that have their important implications in innate and adaptive immunity. Though considerable numbers of new beta-defensins have been discovered, few corresponding antibodies have been reported. The small peptide with special structure and antimicrobial nature of beta-defensins make them very difficult to express in prokaryotic system. Here we adopted a double copy protein expression scheme based on which not only the mBin1b protein was successfully expressed but also the immunity of the antigen was enhanced. The validity of the antisera was verified by using Western blotting and immunohistochemical analyses. It will be a useful tool for deeply investigating the roles of Bin1b and also provide a simple but effective method in raising antisera against other members of the beta-defensin gene family. PMID- 15295652 TI - Development of cell lines stably expressing staphylococcal nuclease fused to dengue 2 virus capsid protein for CTVI. AB - To explore the potential application of capsid-targeted viral inactivation (CTVI) strategy in prophylactic model against dengue virus (DV) infection, here we fused a Ca(2+)-dependent nuclease, staphylococcal nuclease (SN), to the capsid protein of dengue 2 virus (D2C) at the carboxyl terminal, and constructed the desired expression plasmid pc/D2C-SN and control plasmids pc/D2C-SN* and pc/D2C. A mammalian cell line BHK-21 was transfected by electroporation with those plasmids and thereafter selected by 5mgr;/ml blasticidin. The resistant cell clones were then expanding cultured and screened by RT-PCR and Western Blot assays. The nuclease activity of the expressed fusion protein D2C-SN was analyzed by in vitro DNA digestion assay. It was confirmed cell lines stably expressing D2C-SN and control constructs were obtained. The intracellular expressed fusion protein D2C SN had ideal nuclease activity and no cytotoxicity on mammalian cells. Those engineered cell lines provided the experimental system for CTVI application in prophylactic model and paved the new road for combating DV infection with CTVI. PMID- 15295653 TI - Abstracts of the Second European Conference on Head and Neck Cancer, Lille, France, October 16 - 18, 2003 and the foundation of the European Head and Neck Society (EHNS). PMID- 15295654 TI - Mutations in the membrane component, chromosome 1, surface marker 1 (M1S1) gene in gelatinous drop-like corneal dystrophy. AB - PURPOSE: To report mutations in the membrane component, chromosome 1, surface marker 1 ( M1S1) gene in two members of the same family who showed symptoms of gelatinous drop-like corneal dystrophy (GDLD). METHODS: DNA was extracted from leukocytes of peripheral blood of the two affected members of the family and from controls, and the coding region of M1S1 was amplified by polymerase chain reaction (PCR). The PCR products were analyzed by direct sequencing. Normal and mutant M1S1 expression vectors were constructed and transfected into CHO cells to identify the cellular location of the gene products. RESULTS: The affected members had compound heterozygous mutations consisting of a nonsense change at codon 84 (K84X) and a missense mutation resulting in a substitution of arginine for cysteine at codon 108 (C108R). Neither of these mutations was found in the 50 controls. Protein expression analysis showed that the C108R product was distributed diffusely in the cytoplasm, whereas the normal gene product accumulated at cell-to-cell adhesion borders. CONCLUSION: These data indicate that the K84X and C108R mutations in M1S1 cause GDLD. PMID- 15295655 TI - Trehalose versus hyaluronan or cellulose in eyedrops for the treatment of dry eye. AB - PURPOSE: Trehalose eyedrops were found by a previous study to be safe and effective compared with saline in the treatment of moderate-to-severe dry eye syndrome. The present study was designed to compare the efficacy of trehalose eyedrops with that of the commercially available eyedrops containing hyaluronan or cellulose now used in the treatment of moderate-to-severe dry eye syndrome. METHODS: In a randomized, double-masked, 4-week crossover, controlled clinical trial, 36 patients with moderate-to-severe dry eye syndrome were divided into two groups: the hyaluronan (Hyalein)-comparison group (18 patients) and the hydroxyethylcellulose (Mytear)-comparison group (18 patients). Each group used either trehalose or one of the commercially available medications contained in a masked eyedrop container for the first 4 weeks, and then for the second 4 weeks, switched to either trehalose or the commercial eyedrop not used for the first 4 weeks. Symptoms and signs in both eyes were recorded at the baseline, at 4 weeks, and at 8 weeks. RESULTS: At 4 weeks after the treatment, fluorescein and rose bengal staining scores of the ocular surface as well as the tear film breakup time had improved significantly with trehalose eyedrops compared with the commercially available eyedrops containing either hyaluronan or hydroxyethylcellulose (P < 0.001, Wilcoxon signed ranks test). In addition, all the objective signs were significantly better in patients who finished with trehalose at the end of the 8-week trial compared with those who finished with either of the two commercially available drugs. A larger number of patients evaluated trehalose as a better treatment than the commercially available eyedrops. CONCLUSIONS: Trehalose solution was a better treatment for moderate-to severe dry eye syndrome in comparison with two commercially available eyedrops containing hyaluronan or hydroxyethylcellulose. PMID- 15295656 TI - Study of a polymerase chain reaction-based method for detection of herpes simplex virus type 1 DNA among Iranian patients with ocular herpetic keratitis infection. AB - PURPOSE: To study the presence of the herpes simplex virus type 1 (HSV-1) glycoprotein D gene in tear films of Iranian patients with herpetic keratitis. METHODS: Twenty-five tear film and eye swab specimens from 25 herpetic keratitis patients and 10 specimens from 10 healthy volunteers were collected in the Farabi Eye Hospital, Tehran, Iran. HSV-1 DNA was detected by using the nested polymerase chain reaction (nPCR) method. Viral isolation was done using conventional viral techniques. A monoclonal antibody-based enzyme-linked immunosorbent assay (ELISA) was used for confirmation of positive cytopathic effect cell culture. The results of a diagnosis by an ophthalmologist team were compared with those of nPCR. RESULTS: HSV-1 DNA was identified in tear films of 88% (23/25) of suspected herpetic keratitis patients. All healthy controls (100%) had negative PCR results. HSV-1 was isolated in cell culture and confirmed by ELISA in 12% (3/25) of herpetic keratitis patients who had epithelial keratitis. The kappa value showed a high level of agreement between ophthalmologist team diagnosis and the PCR results (kappa = 0.86, P < 0.0001). CONCLUSIONS: nPCR is a sensitive, rapid, and powerful tool for detection of HSV-1 DNA in tear films of ocular herpetic keratitis patients and can serve as a supplemental method for diagnosis of herpetic keratitis infection. PMID- 15295657 TI - Phenotypic changes and inflammatory cell distribution in the cornea during development of experimental immune-mediated blepharoconjunctivitis. AB - PURPOSE: To investigate the morphological changes in the cornea during the development of experimental immune-mediated blepharoconjunctivitis (EC). METHODS: EC was induced in Brown Norway (BN) rats by active immunization with ovalbumin (OVA) emulsified in complete Freund's adjuvant and a subsequent challenge by OVA eyedrops. The corneas were analyzed immunohistochemically. RESULTS: Before the induction of EC, cells stained with OX6 (rat MHC class 2, RT1B), ED1 (tissue macrophages), ED2 (resident macrophages), CD4, or major basic protein were present in the peripheral corneal stroma. ED1- and OX6-stained cells were also observed in the central corneal stroma, and their number increased after the antigen challenge. Infiltration of cells stained with ED1, ED2, OX62 (dendritic cells), CD4, or CD3 (T cells) from the limbus to the peripheral corneal stroma started 6 h after the antigen challenge. Expression of MHC class 2 molecules was induced on the corneal epithelium by the antigen challenge. CONCLUSIONS: The present study demonstrates for the first time the phenotypic changes and distribution of inflammatory cells in the cornea during the development of EC. PMID- 15295658 TI - Visual field damage in normal-tension glaucoma patients with or without ischemic changes in cerebral magnetic resonance imaging. AB - PURPOSE: To compare the pattern of visual field damage between normal-tension glaucoma (NTG) patients with signs indicative of ischemic changes and those NTG patients without signs of ischemic changes, using brain magnetic resonance imaging (MRI), in a single center, cross-sectional study. METHODS: In 94 consecutive NTG patients who were younger than 61 years old, brain MRI images were obtained using fluid-attenuated inversion recovery pulse sequences. The presence of signs indicative of ischemic changes in brain MRI images was decided separately by two neuroradiologists masked to the diagnosis and stage of glaucoma. Visual field testing was performed using the 30-2 program of the Humphrey Visual Field Analyzer. Between the patients with signs indicative of ischemic changes in brain MRI (ischemic group) and those without MRI signs (nonischemic group), total deviation (TD) at each test point less the average of TDs of the 30-2 program ([TD - TD(mean)])--was compared at each test point. RESULTS: Signs indicative of ischemic changes in brain MRI were found in 32 of the 94 patients (34.0%). Age, blood pressure, refraction, intraocular pressure, the average of TDs, mean deviation, and corrected pattern standard deviation were not significantly different between the ischemic (N = 32) and nonischemic (N = 62) groups (P > 0.2). [TD - TD(mean)] in the ischemic group was significantly smaller than that in the nonischemic group at 6 nonedge contiguous test points in the inferior pericentral to nasal field (P = 0.005-0.047). CONCLUSION: NTG patients with signs indicative of ischemic changes in brain MRI had a relatively deeper depression in the inferior pericentral visual field. PMID- 15295659 TI - Comparison of electrical stimulation thresholds in normal and retinal degenerated mouse retina. AB - PURPOSE: To compare the threshold for electrically elicited action potentials of retinal ganglion cells in normal mouse retina and photoreceptor degenerated (rd) mouse retina. METHODS: Microelectrode recordings were made from retinal ganglion cells of normal and rd mice. Mice with a genetically based retinal degeneration (rd mice) were grown to the age of 16 weeks, when light-evoked responses could no longer be recorded. A bare wire was placed in the vitreous to stimulate the retina with charge-balanced current pulses. The following pulse shapes were investigated: single, square biphasic pulse, single sine wave, and biphasic pulse trains. RESULTS: Normal mice had significantly lower stimulus thresholds than rd mice for all pulse shapes. In normal and rd mice, short pulses were more efficient with respect to total charge used, but required a higher current. In normal mice, sine wave stimulation was significantly more efficient than a biphasic pulse of the same duration. No difference was noted between sine wave and square wave stimulation in rd mice. Pulse trains offered little benefit over single pulses. CONCLUSION: The amount of electrical charge required to elicit an action potential is dependent on the condition of the retina and the shape of the stimulus pulse used to deliver the charge. PMID- 15295660 TI - Gene analysis and evaluation of the single founder effect in Japanese patients with Oguchi disease. AB - PURPOSE: To analyze mutations of the arrestin/S-antigen (SAG) gene in nine newly identified Oguchi disease patients, and to examine whether the 926delA (formerly called 1147delA) mutation in the SAG gene is inherited from a single founder. METHODS: DNA samples were assayed for mutations around nucleotide 926 of the SAG gene by direct sequencing, and analyzed for polymorphisms at codon 403 and IVS6 18 of the SAG gene by restriction analysis of polymerase chain reaction products. RESULTS: All nine newly identified patients were homozygous for the 926delA mutation and had the same haplotype at codon 403 and IVS6-18. These findings are identical to those of previous reports of four Japanese Oguchi disease patients. CONCLUSIONS: Mutation 926delA of the SAG gene is the main cause of Oguchi disease in Japanese. This mutation appears to have been inherited from a single founder. PMID- 15295661 TI - N-acetylaspartate concentration in the chiasm measured by in vivo proton magnetic resonance spectroscopy. AB - PURPOSE: To measure the concentration of N-acetylaspartate (NAA) in the chiasm of both normal controls and patients with chiasmal optic neuritis by in vivo proton magnetic resonance spectroscopy (1H-MRS). METHODS: Ten normal subjects (age range, 22 to 40 years; mean, 31 years; seven men and three women) and two patients with chiasmal optic neuritis were examined at the Sapporo Medical University Hospital. Localized 1H-MRS spectra of the chiasm were obtained using a whole-body 1.5-T magnetic resonance system. RESULTS: 1H-MRS spectra of the chiasm were obtained in all subjects. The level of NAA concentration in the optic chiasm was 15.73 +/- 1.43 mM (mean +/- SD) in normal subjects. In contrast, the NAA levels in the two patients with chiasmal optic neuritis were significantly lower (P < 0.001). The level of NAA in the two patients was significantly increased after improvement of visual acuity and visual fields following corticosteroid pulse therapy. CONCLUSIONS: These results suggest that 1H-MRS enables us to estimate the concentration of NAA in the chiasm. The NAA concentration in the optic nerve measured by 1H-MRS may be a new clinical parameter to monitor the axonal damage following optic neuritis. PMID- 15295662 TI - Retinal blood flow in the macular area before and after scleral buckling procedures for rhegmatogenous retinal detachment without macular involvement. AB - PURPOSE: To investigate retinal microcirculation changes in patients with rhegmatogenous retinal detachment (RRD). METHODS: The tissue blood flow in the macular area was measured in 28 patients with RRD without macular involvement by scanning laser Doppler flowmetry before and after scleral buckling procedures. The mean blood flow (MBF) was calculated by the automatic full-field analysis program. The MBF ratios of the affected eye to the fellow eye (a/f ratio) in patients were compared with those of the right eye to the left eye (R/L ratio) in the control subjects. RESULTS: The mean preoperative a/f ratio in the patients (0.81 +/- 0.11) was lower than the mean R/L ratio in the control subjects (1.02 +/- 0.11, P < 0.0001) and correlated with the extent of RRD (P < 0.05). The mean a/f ratio tended to decrease 2 weeks after surgery (0.72 +/- 0.09) and recovered to an almost normal level after 1 month (0.96 +/- 0.09). The blood-flow change was not influenced by the type of buckling. CONCLUSIONS: The retinal microcirculation in the macular area was disturbed in RRD patients without macular involvement. It correlated with the extent of the RRD, and subsided 1 month after successful scleral buckling procedures. PMID- 15295663 TI - Magnetic resonance imaging of pre- and postoperative lower eyelid states in involutional entropion. AB - PURPOSE: To disclose pre- and postoperative lower eyelid gradients in involutional entropion using sagittal magnetic resonance imaging (MRI). METHODS: Three female patients, average age 82 (two right eyes and one left), were operated on for involutional entropion by the Jones procedure. Before and after the surgery, the lower eyelid gradient was evaluated by MRI and photography. RESULTS: Preoperatively, each lower eyelid presented anterior protrusion, and the retractor was apart from the globe. Postoperatively, no anterior protrusion was observed, and the retractor was pulled posteroinferiorly and located parallel to the globe. CONCLUSIONS: The MRI visualization of pre- and postoperative changes of gradient in the lower involutional entropion supports surgical reconstruction. PMID- 15295664 TI - Results of therapeutic penetrating keratoplasty. AB - PURPOSE: By a retrospective study of patients in the Ankara Hospital Eye Clinic, to determine the anatomical and visual results of therapeutic penetrating keratoplasty (PK) and its role in the management of corneal disease. METHODS: Therapeutic PK was performed in 36 patients (37 eyes) who had corneal perforation due to corneal disease (23 eyes) or eyes in which perforation was imminent (14 eyes). Initial indications for grafting were nonperforated descemetocele without inflammation (six eyes, 16.2%); nonperforated bacterial corneal ulcer (five eyes, 13.5%); nonperforated herpetic keratitis with active stromal inflammation (two eyes, 5.4%); acanthamoeba keratitis (one eye, 2.7%); perforation due to herpetic keratitis (13 eyes, 35.2%); perforation due to persistent epithelial defect (8 eyes, 21.6%); or perforation due to bacterial corneal ulcer (two eyes; 5.4%). The results were evaluated for each of the following criteria: anatomical integrity of the eye, cure of the disease, complications, graft clarity, and visual acuity. RESULTS: Anatomical integrity was achieved in 21 of the 23 eyes (91.3%) perforated from corneal disease. Therapeutic PK cured the disease in all bacterial keratitis cases. The proportion of clear grafts was 60.9% in the 23 eyes perforated from corneal disease, and 57.1% in the 14 eyes in which perforation was imminent. Fifteen eyes (40.5%) obtained a final visual acuity of 20/100 or better; five of these eyes were not yet perforated before the PK. CONCLUSIONS: Therapeutic PK is effective in the management of the eye with active uncontrolled infection or perforation from corneal disease. Approximately half of our patients maintained a clear graft at the last visit. Without therapeutic surgery, these eyes would have been lost. PMID- 15295665 TI - Increased tear evaporation in a patient with ectrodactyly-ectodermal dysplasia clefting syndrome. AB - PURPOSE: To describe the tear function and ocular surface disorders in a patient with ectrodactyly-ectodermal dysplasia-clefting (EEC) syndrome. METHODS: Routine ophthalmic examinations were performed, including slit-lamp biomicroscopy, anterior segment photography including transillumination photos of the lids, Schirmer tests I and II, tear film break-up time (BUT) assessment, corneal fluorescein staining, DR-1 tear film lipid layer interferometry, and tear evaporation rate measurements. RESULTS: Slit-lamp examination revealed conjunctival hyperemia, superficial punctate keratopathy, and corneal leucoma with neovascularization. Although the Schirmer test values were within normal limits, the BUT value was 0 s in both eyes. Transillumination of the lids showed the absence of meibomian glandular structures. DR-1 tear film lipid layer interferometry results were dry eye grade 5 with an irregular tear film, areas of corneal surface exposure, and several dry spots. The tear evaporation rate was elevated and was measured as 6.98 x 10(-7) g/cm2 per second (normal, 4.1 +/- 1.4 x 10(-7) g/cm2 per second). CONCLUSION: The ocular surface disorder and shortened BUT in EEC syndrome were attributed to the absence of meibomian glands, leading to lipid layer deficiency in the tear film with a concomitant increase in tear evaporation. PMID- 15295666 TI - Ocular discomfort at the initial wearing of rigid gas permeable contact lenses. AB - PURPOSE: To evaluate the relationship between ocular discomfort in the initial wearing of rigid gas permeable contact lenses (RGPCL) and the time required to adapt to RGPCL as measured by using the visual analogue scale (VAS). METHODS: RGPCL were prescribed for 89 patients (178 eyes) with myopia or myopic astigmatism and no history of wearing contact lenses. The patients who stopped wearing RGPCL after 1 week because of ocular discomfort were defined as dropouts, and those who adapted to regular full-time RGPCL wear were defined as successful wearers. VAS scores were measured at several intervals for 1 month after the initial wearing of the lenses. The number of days required to adapt to wearing RGPCL was defined as the adaptation time. RESULTS: Four patients were dropouts due to ocular discomfort (eight eyes; 4.5%). VAS scores were significantly lower in successful wearers than in dropouts at 1 week, although their VAS scores were not different at 1 day. VAS scores of the successful wearers at 1 week were significantly lower than those at 1 day. The average time required to adapt to RGPCL was 23.0 +/- 22.1 days for the 85 successful wearers. Lower VAS scores at 1 day or 1 week were significantly correlated with a shorter adaptation time in the successful RGPCL wearers. CONCLUSIONS: Our results indicate that the VAS score might be useful for predicting successful RGPCL use and the adaptation time required by successful wearers. PMID- 15295667 TI - Association between nocturnal blood pressure reduction and progression of visual field defect in patients with primary open-angle glaucoma or normal-tension glaucoma. AB - PURPOSE: To assess prospectively the relationship between nocturnal blood pressure reduction (dip) and progression of the visual field defect in patients with normal-tension glaucoma (NTG) or primary open-angle glaucoma (POAG). PATIENTS AND METHODS: The subjects studied were 38 patients with glaucoma (23 patients with NTG, 15 patients with POAG), in whom 48-h ambulatory blood pressure monitoring was conducted, who were followed for at least 4 years, and in whom reliable visual field tests were performed at least 5 times. The progression was determined by the mean deviation (MD) slope analysis system installed in the Humphrey field analyzer Statpac program. Glaucoma patients with a dip of <10% were assigned to the nondipper group, those with a dip of 10%-20% to the tipper group, and those with a dip of >20% to the extreme dipper group. The dipper group was defined as physiologic dippers, while the nondipper and the extreme dipper groups were defined as nonphysiologic dippers, to study the relationship between the disturbance of the dip and progression of the visual field defect. RESULTS: Thirteen patients showed significant progression, while 25 patients did not. There were no significant differences in the mean intraocular pressure and follow up period between the patients with progression and those without progression. Half (7/14) of the nondippers, 20% (4/20) of the dippers, and 50% (2/4) of the extreme dippers showed progression, indicating a tendency of progression in the nondipper and the extreme dipper groups. The nonphysiologic dippers had a higher incidence of progression compared with the physiologic dippers (P = 0.05). Among the glaucoma patients in the nondipper and dipper categories only, those with progression had significantly smaller dips (P = 0.02). CONCLUSION: These results suggest that disturbance in the physiologic dip may be involved in the progression of glaucoma. PMID- 15295668 TI - Clinical features and prognosis in ocular toxoplasmosis. AB - PURPOSE: To evaluate retrospectively the clinical characteristics, complications, and prognosis in patients with ocular toxoplasmosis. PATIENTS AND METHODS: We reviewed the records of 189 patients (243 eyes) with ocular toxoplasmosis who were examined between 1972 and 1999. Color fundus photography and, in some patients, fluorescein angiography and indocyanine green angiography were performed. There were 98 male (52%) and 91 female (48%) patients with a mean age of 22.8 +/- 8.9 years. RESULTS: Of the patients, 140 (74%) had congenital and 49 (26%) had acquired toxoplasmosis. At the initial examination, there were active lesions in 65 eyes and inactive lesions in 178 eyes. Active lesions included retinochoroiditis in 59 (91%), papillitis in 2 (3%), and neuroretinitis in 4 (6%) eyes. There was also an inactive scar in 17 eyes with active retinochoroiditis. Localisation of the active retinochoroiditis was the macula in 44 (74%), the macula and peripheral retina in 3 (5%), the peripheral retina in 9 (15%) and the peripapillary retina in 3 (5%) eyes. Optic atrophy, pigment epithelial detachment, choroidal neovascularization, lamellar macular hole, and retinal neovascularization were seen during the follow-up period. CONCLUSIONS: Ocular toxoplasmosis commonly affects the macula and seriously impairs visual acuity. The prevention of acquired and congenital infections is very important in controlling ocular toxoplasmosis. Patients should be followed to avoid late complications. PMID- 15295669 TI - Choroidal circulatory disturbance in ocular sarcoidosis without the appearance of retinal lesions or loss of visual function. AB - BACKGROUND: Birdshot chorioretinopathy, acute posterior multifocal placoid pigment epitheliopathy, and retinal pigment epithelial detachment have been reported as rare manifestations associated with sarcoidosis, suggesting that ocular sarcoidosis may affect the choroidal circulation. We report a case of ocular sarcoidosis representing a choroidal circulatory disturbance without the appearance of retinal lesions or loss of retinal function. CASE: A 20-year-old woman was referred with blurred vision in the left eye. Inflammatory change in the anterior segment of the eye was noted with multiple nodules on the iris. OBSERVATIONS: Hematological examination revealed elevated lysozyme levels. Bilateral hilar lymphadenopathy was noted on chest X-ray. Specimens obtained by transbronchial lung biopsy revealed granuloma with Langhans giant cells, which led to the diagnosis of sarcoidosis. The eye was treated with topical steroid. The symptoms and the inflammatory change in the anterior segment disappeared within 10 days. However, despite the normal appearance of the ocular fundus, fluorescein angiography revealed multiple puncta of hyperfluorescence. In indocyanine green angiography, a filling delay was noted in the area corresponding to the punctate lesions. Static visual field testing and multifocal electroretinography showed no significant changes. At the last visit, 15 months after the left eye became asymptomatic, the choroidal lesions had disappeared with no residual alteration of the funduscopic appearance or visual function. CONCLUSIONS: This case indicates that choroidal circulatory disturbance can underlie ocular sarcoidosis even in the absence of funduscopically detectable lesions and loss of visual function. PMID- 15295670 TI - Isolated caruncular approach for orbital decompression. AB - PURPOSE: To evaluate the effectiveness of the isolated caruncular approach to orbital decompression of thyroid ophthalmopathy. METHODS: In a retrospective, noncomparative, interventional case series, we reviewed the records of 29 patients (48 orbits) who had thyroid ophthalmopathy and had undergone orbital decompression using the caruncular approach. The medial wall was decompressed in two patients (three orbits), and the medial and inferior walls were decompressed in 27 patients (45 orbits). RESULTS: The mean retrodisplacement achieved was 2.7 mm of decompression of the medial wall, and 4.2 mm of decompression of the medial and inferior walls. Diplopia arose in the primary position in 4 of 17 previously asymptomatic patients. Persistent postdecompression strabismus was managed successfully with adjustable strabismus surgery. Other complications were minimal, including a hypertrophic scar in one eye and a pyogenic granuloma in another. CONCLUSIONS: Orbital decompression using the isolated caruncular approach offers rapid access to the medial and inferior orbital walls and makes graded decompression possible in each case. It is a useful approach for patients wishing surgery for cosmetic purposes and for those with compressive optic neuropathy as well. PMID- 15295671 TI - A comparison of viscogoniotomy with classical goniotomy in Turkish patients. AB - PURPOSE: To compare the success and complication rates of viscogoniotomy and goniotomy in Turkish patients. METHODS: In a retrospective review, the medical records of patients with primary congenital glaucoma were divided into two groups. Group 1 consisted of 21 patients (38 eyes) who had undergone classical goniotomy, and group 2 consisted of 25 patients (44 eyes) who had undergone viscogoniotomy. The success rates and intra- and postoperative complications were compared between the two groups. RESULTS: Mean preoperative introcular pressure (IOP) was 28.9 +/- 3.6 mmHg in group 1 and 29.3 +/- 2.8 mmHg in group 2. At the last postoperative visit, it was 17.3 +/- 3.1 mmHg and 16.2 +/- 2.1 mmHg, respectively (P < 0.001). The success rates at the last visit of group 1 and group 2 were 68.4% and 88.6%, respectively (P < 0.05). The most common early postoperative complication was hyphema in group 1 and transient IOP elevation in group 2. CONCLUSION: Viscogoniotomy may increase the success rate and decrease the complications rate by preventing hyphema and flat anterior chamber. PMID- 15295672 TI - Phacotrabeculectomy in treatment of primary angle-closure glaucoma and primary open-angle glaucoma. AB - PURPOSE: To evaluate the surgical outcome of combined phacoemulsification, posterior chamber intraocular lens implantation, and trabeculectomy (phacotrabeculectomy) in patients with primary angle-closure glaucoma (PACG) or primary open-angle glaucoma (POAG). METHODS: The records of 57 consecutive patients (65 eyes) with PACG or POAG that were treated with phacotrabeculectomy were reviewed retrospectively. There were 31 eyes with PACG and 34 with POAG. The mean follow-up period was 21.0 +/- 8.3 months. The visual acuity, intraocular pressure (IOP), number of medications, and complications were evaluated. RESULTS: The mean IOP and the number of glaucoma medications decreased significantly after phacotrabeculectomy in both groups. The mean IOP reduction was significantly greater in eyes with PACG (P < 0.05). The absolute success rates were 87.1% and 70.6% in PACG and POAG, respectively. The difference in the success rates was not significant (P = 0.297). The early postoperative complication rates were similar in both groups. CONCLUSIONS: Phacotrabeculectomy results in greater IOP reduction in eyes with PACG than in those with POAG, but the overall success rates were not significantly different. PMID- 15295673 TI - Ultrasound biomicroscopic findings of ciliary body malignant melanoma. AB - BACKGROUND: Malignant melanoma of the uveal tract occurs rarely in Asian populations, and melanoma of the ciliary body is extremely rare. We treated an Asian man with uveal melanoma, relying on ultrasound biomicroscopy (UBM) findings for diagnosis and evaluation of the tumor. CASE. A 65-year-old man with uveal melanoma was examined by slit-lamp microscopy, UBM, and magnetic resonance imaging (MRI), and histopathological examinations were performed as well. OBSERVATIONS: Slit-lamp examination and MRI gave rough images of the tumor, but the exact origin and margin of the mass could not be determined. UBM clearly characterized the tumor as a medium echoic solid mass with acoustic hollowing attached to the ciliary body by a thin stem. Ciliary body detachment at the pars plicata was also found by UBM. Histopathological examination confirmed that the tumor was an epithelioid cell malignant melanoma of the ciliary body. CONCLUSIONS: UBM is useful in detecting, diagnosing, and evaluating a malignant melanoma of the ciliary body. PMID- 15295674 TI - Concomitant cavernous hemangioma and venous angioma of the orbit. AB - BACKGROUND: An unusual case of cavernous hemangioma coexisting with venous angioma in the ipsilateral orbit is described. CASE: A 67-year-old woman had a mass in the lower eyelid of her right eye and proptosis. Imaging examinations showed two masses connected to each other in the extraconal space of the right orbit. OBSERVATIONS: Anterior orbitotomy was performed to remove the tumors. Histopathological evaluation of the anteriorly located tumor revealed cavernous hemangioma. The posteriorly located tumor was found to be a venous angioma. Endothelial cells in both tumors showed positive immunostaining for factor VIII related antigen and smooth muscle actin. Immunoreactivity of smooth muscle actin was more prominent in the interstitium of the posteriorly located tumor. CONCLUSION: Coexistence of cavernous hemangioma and venous angioma in the same area suggests that they are a continuum and of the same origin. PMID- 15295675 TI - Glomus tumour of the eyelid. PMID- 15295676 TI - A case of dacryocystocele in an adult. PMID- 15295677 TI - Adrenomedullin levels in early and late rhegmatogenous retinal detachments. PMID- 15295678 TI - Photodynamic therapy in patients with idiopathic choroidal neovascularization. PMID- 15295679 TI - Nonarteritic anterior ischemic optic neuropathy after photodynamic therapy for choroidal neovascularization. PMID- 15295682 TI - [Prognosis of hepatocellular carcinoma according to new staging classifications]. AB - BACKGROUND AND OBJECTIVE: We investigated prognostic factors and survival of patients with hepatocellular carcinoma (HCC) in North-Germany. Established staging systems (Child-Pugh, Okuda and UICC classification) were compared with new prognostic scores from Italy (CLIP) and Spain (BCLC). PATIENTS AND METHODS: The clinical course of 62 consecutive patients (34-82 years, 48 males, 14 females) with HCC observed in the Medical School of Hannover from October 1996 to September 1998 were retrospectively analyzed. The patients were classified according to the staging systems of Child Pugh, Okuda, UICC, CLIP and BCLC. Follow-up ended on December 31 (st) 2001. RESULTS: Overall median survival was 11,3 (1 - 59,5) months. At univariate analysis (log-rank test) Okuda, UICC, CLIP and BCLC Score were each associated with a shorter survival. In contrast Child Pugh score provided no significant prognostication. By multiple regression analysis (Cox regression analysis), only the CLIP and UICC score and chronic hepatitis B infection were shown to be independent risk factors. CONCLUSION: Our investigations indicate, that the CLIP and UICC classifications identified those patients with the best prognosis and they, as well as chronic hepatitis C, were shown to be independent risk factors. PMID- 15295683 TI - [Encoding of diagnosis by medical documentation assistant or ward physician. Influence on the mapping of Diagnostic Related Group (DRG) performance]. AB - BACKGROUND AND OBJECTIVE: Starting in 2004 the patient budget in Germany will be calculated according to the Diagnosis Related Group (DRG) system, by which system the monetary reward of a unit will be directly related to the quality of documentation e. g. diagnosis and procedures. The aim of this study was to compare the quality of documentation by a medical documentation assistant (MDA) with the usual practice of documentation by the ward physician (WP). Additionally, the effect of introducing a completely changed organizational process was tested. METHODS: In a prospective study on the ward of a gastroenterology unit two different approaches of medical documentation were compared. In a first six-month period diagnosis and procedures were encoded by WP. In the following six months an MDA was introduced and involved in the encoding process. RESULTS: In the first six months 221 patients (mean age 55 +/- 16,2 years, 55,7 % males) were evaluated, whereas in the following six months 305 patients (mean age 53 +/- 15,4 years, 59,9 % males) were included. The introduction of an MDA improved medical documentation and economical reference numbers: with an increase of diagnosis per case to 7,43 (in first six months 5,53), patient complexity and comorbidity level (PCCL) to 2,5 (in first six months 2,13), case-mix index to 1,04 (in first six months 0,98). Additionally the medial hospitalization time decreased from 11,2 to 8,1 days. The average daily reimbursement increased in the MDA group from 423 Euro to 603 Euro. This was calculated on the basis of a basic case factor of 2900 Euro. CONCLUSION: Introduction of an MDA in a gastroenterology ward increases the quality of documentation and results in an improved presentation of DRG-relevant efforts with a better reimbursement of medical costs. PMID- 15295684 TI - [Long-term effects of sildenafil in a patient with scleroderma-associated pulmonary hypertension and Raynaud's syndrome]. AB - HISTORY: A 65-year-old woman was admitted because of dyspnea at rest and peripheral edema due to scleroderma-associated pulmonary fibrosis and hypertension, as well as Raynaud's phenomenon. DIAGNOSTIC FEATURES, TREATMENT AND COURSE: She had a marked restrictive ventilatory disorder with severe impairment of diffusion capacity. Right heart catheterization demonstrated a mean pulmonary artery pressure of 50 mmHg. She was able to walk only 220 m. All usual methods of treatment failed to give satisfactory results so that sildenafil (phospherodiesterase type-5 |PDE-5| inhibitor; Viagra ((R)) was given, even though it is not licensed for this indications ("off-label", as a therapeutic attempt. This achieved definite reduction in pulmonary arterial pressure and significantly improved the clinical symptoms. In particular, it drastically reduced the level of atrial natriuretic peptide, an important prognostic marker in right heart failure. Sildenafil also significantly raised peripheral perfusion and the signs of Raynaud's syndrome. CONCLUSION: PDE-5 inhibitors are efficacious in scleroderma-associated pulmonary hypertension and may also provide a new option in the treatment of Raynaud's disease. PMID- 15295685 TI - [Paraneoplastic neurological autoimmune diseases]. PMID- 15295686 TI - [Intestinal tuberculosis: a clinical and diagnostic challenge]. PMID- 15295687 TI - [Immediate hemostasis of the femoral artery after heart catheterization: the present situation of closure systems]. AB - The femoral approach is the most commonly used route for diagnostic cardiac catheterization and coronary interventions today. Manual compression and pressure bandages usually lead to immobilisation of the patient for several hours and may result in significant discomfort. Since the introduction of the first femoral closure device in 1991, many devices have proven their efficacy in significantly reducing time to hemostasis while simultaneously improving patient comfort. Twenty four closure device systems with different concepts are on the market, e. g. pure collagen, collagen + thrombin, collagen + anchor, vascular suture, hemostatic patches and pads, staples and more. The four predominantly used are Angio-Seal (46 %), Perclose (32 %), VasoSeal (14 %) and Duett (3 %). The effectiveness of all four systems has been proven in a prospective, randomized, controlled multicenter trial each. Efficacy and safety were analyzed using data from ten comparative studies in 8832 predominantly or exclusively interventional patients, however none of the closure systems proved to be superior. Fortunately, recent years have shown a trend toward a reduction in local complications by vascular closure devices compared to manual compression. Closure devices are thus becoming increasingly cost effective. Vascular closure systems should be preferred when the prolonged supine position is not tolerated, a protein IIb/IIIa inhibitor was used during the procedure, or early discharge of patient is anticipated. In the presence of peripheral vascular disease, small diameter of the femoral vessels or stenotic lesions in the femoral artery, closure devices should be used with caution. Closure systems for immediate femoral puncture site hemostasis are now an important tool of invasive cardiology today. PMID- 15295688 TI - What is the purpose of the embryonic heart beat? Or how facts can ultimately prevail over physiological dogma. AB - Embryonic physiology is often viewed as merely those processes understood for the adult but conducted on a smaller physical scale. Yet striking examples of the inaccuracy of this perspective can be identified in the embryonic cardiovascular system. For example, dogma holds that the embryonic heart begins to beat to pump blood for convective transport, just like that of the adult. This is the major assumption inherent in the hypothesis we have called "convective synchronotropy"; that is, the embryonic heart starts to beat synchronously with the need for convective blood flow. However, there is compelling evidence on many fronts that the convective flow of blood generated by the early embryonic vertebrate heart is simply not required for transport of oxygen, nutrients, metabolic wastes, or hormones, all of which can be achieved entirely by diffusion. In fact, fish, amphibian, and bird embryos lacking a functional heart (either through surgical intervention or mutation) or whose oxygen-hemoglobin transport has been chemically eliminated nonetheless continue to function and grow in size for extended periods up to the point at which diffusion alone can no longer serve oxygen transport needs. We advocate the alternative hypothesis of "prosynchronotropy" (i.e., the heart starts to beat well before convective blood flow is needed for bulk transport). So, what is the purpose of the early embryonic heart beat? Evidence is presented herein in support of a morphogenic rationale for prosynchronotropy. Specifically, it appears that the initial rationale for the beat of the vertebrate embryonic heart may be two-fold: to aid in subtle but significant aspects of cardiac growth, shaping, and maturation, and to facilitate cardiac maturation angiogenesis--the formation of new vessels by sprouting from vessel tips. Ultimately, the embryonic cardiovascular system provides a graphic demonstration of how adult physiological functions should not, without verification, be interpolated back to the embryo of that species. PMID- 15295689 TI - Accumulation of lactate by frozen painted turtles (Chrysemys picta) and its relationship to freeze tolerance. AB - Hatchling painted turtles (Chrysemys picta) survived freezing at -2 degrees C for 4 d, few recovered from freezing lasting 6 d, and none survived being frozen for 8 d. Whole-body glucose and lactate were low in animals that had not been subjected to cold and ice but increased precipitously in animals that were frozen for 2 d. Both metabolites continued to increase, but at a somewhat lower rate, in animals frozen for 4, 6, or 8 d. The increase in whole-body lactate reflects a reliance by frozen hatchlings on anaerobiosis, whereas the increase in glucose presumably results from mobilization of glycogen reserves to support anaerobic metabolism. Mortality of frozen hatchlings is correlated with the increase in whole-body lactate. Factors that may contribute to the observed correlation include a compromised capacity for individual organs to cope with the lactic acidosis that accompanies anaerobic metabolism and organ-specific depletion of energy reserves. Individual organs must rely on buffering and glucose reserves available in situ because blood of frozen hatchlings does not circulate. Thus, buffer from the shell cannot be transported to other organs, lactate cannot be sequestered in the shell, and glucose mobilized from liver glycogen is not available to supplement glucose reserves of other tissues. This integrated suite of physiological disruptions may limit tolerance of freezing to conditions with little or no ecological relevance. PMID- 15295690 TI - A randomized, double-blind, placebo-controlled trial of the use of prednisolone as an adjunct to treatment in HIV-1-associated pleural tuberculosis. AB - BACKGROUND: Active tuberculosis may accelerate progression of human immunodeficiency virus (HIV) infection by promoting viral replication in activated lymphocytes. Glucocorticoids are used in pleural tuberculosis to reduce inflammation-induced pathology, and their use also might reduce progression of HIV by suppressing immune activation. We examined the effect that prednisolone has on survival in HIV-1-associated pleural tuberculosis. METHODS: We conducted a randomized, double-blind, placebo-controlled trial of prednisolone as an adjunct to tuberculosis treatment, in adults with HIV-1-associated pleural tuberculosis. The primary outcome was death. Analysis was by intention to treat. RESULTS: Of 197 participants, 99 were assigned to the prednisolone group and 98 to the placebo group. The mortality rate was 21 deaths/100 person-years (pyr) in the prednisolone group and 25 deaths/100 pyr in the placebo group (age-, sex-, and initial CD4+ T cell count-adjusted mortality rate ratio, 0.99 [95% confidence interval, 0.62-1.56] [P =.95]). Resolution of tuberculosis was faster in the prednisolone group, but recurrence rates were slightly (though not significantly) higher, and use of prednisolone was associated with a significantly higher incidence of Kaposi sarcoma (4.2 cases/100 pyr, compared with 0 cases/100 pyr [P =.02]). CONCLUSIONS: In view of the lack of survival benefit and the increased risk of Kaposi sarcoma, the use of prednisolone in HIV-associated tuberculous pleurisy is not recommended. PMID- 15295691 TI - Decreased HIV transmission after a policy of providing free access to highly active antiretroviral therapy in Taiwan. AB - BACKGROUND: Taiwan established a nationwide surveillance system for human immunodeficiency virus (HIV) infection in 1989 and adopted a policy to provide all HIV-infected citizens with free access to highly active antiretroviral therapy (HAART) beginning in April 1997. This provided an opportunity to determine the effect of the widespread use of HAART on the evolution of the HIV epidemic. METHODS: We analyzed national HIV surveillance data. The HIV transmission rate was estimated by use of an exponential model of HIV epidemic evolution, with statistical projection over the interval between infection and detection to fit the surveillance data. RESULTS: By the end of 2002, the cumulative number of HIV-infected citizens in Taiwan had reached 4390 (0.019% of the total population). After free access to HAART was established, the estimated HIV transmission rate decreased by 53% (0.391 vs. 0.184 new cases/prevalent case year [95% confidence interval, 31%-65%]). There was no statistically significant change in the incidence of syphilis, in the general population or among HIV positive patients, during the same period. CONCLUSION: Providing free HAART to all HIV-infected citizens was associated with a 53% decrease in the HIV transmission rate and contributed to the control of the HIV epidemic in Taiwan. PMID- 15295692 TI - Weighted phenotypic susceptibility scores are predictive of the HIV-1 RNA response in protease inhibitor-experienced HIV-1-infected subjects. AB - We analyzed retrospectively, by use of logistic regression, the role of the phenotypic susceptibility score (PSS) as a determinant of virologic outcome in a study of treatment-experienced human immunodeficiency virus (HIV) type 1-infected subjects. A model for PSS in which each drug was treated equally and scored as active or inactive was not predictive of HIV-1 RNA load (VL) suppression. However, weighting the potential contribution of each drug on the basis of inferred potency and by use of a continuous scale to quantify drug susceptibility revealed significant associations between PSS and outcome. Entry VL was a strong independent predictor of therapy outcome, and PSS was a strong predictor of the magnitude of the initial decrease in VL. This suggests that the prospective evaluation of PSS to identify regimens that maximize decreases in VL to reduce the probability of virologic rebound could improve antiretroviral treatment. PMID- 15295693 TI - Susceptibility of HIV type 1 to the fusion inhibitor T-20 is reduced on insertion of host intercellular adhesion molecule 1 in the virus membrane. AB - BACKGROUND: T-20 is a synthetic peptide that targets and blocks the entry of human immunodeficiency virus (HIV)-1 inside target cells. Data from clinical studies have shown that primary HIV-1 variants display a broad range of susceptibilities to T-20 that are mainly caused by mutations in the N-terminal heptad repeat of gp41 and by other determinants, such as envelope and coreceptor affinity, coreceptor specificity, and receptor density. We investigated whether sensitivity to this drug is affected by the incorporation of intercellular adhesion molecule (ICAM)-1 in HIV-1. METHODS: We used laboratory isolates of HIV 1 (X4- and R5-tropic) either lacking or bearing ICAM-1 and clinical variants. RESULTS: We demonstrate that ICAM-1-bearing virions are more resistant to T-20 than are isogenic HIV-1 particles without it. This effect is lost when ICAM 1/leukocyte function antigen-1 interactions are blocked by antibody. The mechanism through which virus-anchored host ICAM-1 is acting seems to be a reduction of the kinetic window during which the viral envelope is sensitive to T 20. CONCLUSIONS: Altogether, these results demonstrate that the insertion of host derived ICAM-1 into HIV-1 represents another factor that affects virus sensitivity to the entry inhibitor. These findings have potential implications for the therapeutic use of T-20. PMID- 15295694 TI - Breakthrough infections during phase 1 and 2 prime-boost HIV-1 vaccine trials with canarypox vectors (ALVAC) and booster dose of recombinant gp120 or gp160. AB - Candidate human immunodeficiency virus (HIV)-1 vaccines that elicit cytotoxic T lymphocytes may modulate HIV infection, requiring a prototype evaluation to assess participants who become infected with HIV. Of 1497 participants in canarypox HIV-1 vaccine prime-boost trials, 28 (1.9%) acquired HIV-1 infection after vaccination. Median plasma HIV-1 RNA levels (vaccinees, 4.78 log10 copies/mL; placebo recipients, 4.27 log10 copies/mL) and CD4 cell counts (vaccinees, 552 cells/mm3; placebo recipients, 657 cells/mm3) before administration of antiretroviral therapy (ART) and time to a composite end point (plasma HIV-1 RNA level >55,000 copies/mL, CD4 cell count <350 cells/mm3, or initiation of ART) did not differ significantly between vaccinees and placebo recipients (P =.4, P =.1, and P =.7, respectively). Persons who acquire HIV-1 infection while enrolled in HIV-1 vaccine trials can be successfully followed after infection, to determine whether vaccines alter the course of HIV-1 infection. PMID- 15295695 TI - Endocrine pancreatic dysfunction in HIV-infected children: association with growth alterations. AB - BACKGROUND: The pancreatic endocrine system normally guarantees a quick and efficient response to daily metabolic perturbations, but associated data for human immunodeficiency virus (HIV)-infected patients are lacking. A prospective study was performed to evaluate pancreatic endocrine secretion and its possible association with failure to thrive among HIV-infected children. METHODS: Fourteen well-nourished, prepubertal, HIV-infected children (6 boys and 8 girls; age range, 5-11 years), none of whom were receiving protease inhibitors, and 16 clinically healthy sex- and age-matched children formed the patient group and the control group, respectively. At yearly follow-up examinations, insulin, glucagon, C-peptide, and glucose levels were measured; the ratio of insulin to glucose, the ratio of insulin to glucagon, and the homeostasis model assessment (HOMA) index were calculated; the glucagon test was administered; and growth hormone, thyroid stimulating hormone, adrenocorticotropic hormone, cortisol, and lipid patterns were evaluated. RESULTS: Insulin, glucagon, C-peptide, glucose, and HOMA measurements were significantly higher among patients, compared with control subjects, at all 3 follow-ups performed to date. The glucagon test revealed a normal glycemic response in all the healthy control subjects and a significantly impaired response in 11 patients. A significant correlation emerged between the ratio of insulin to glucagon and the growth velocity of HIV-infected children. CONCLUSION: To our knowledge, the present study provides the first evidence of altered pancreatic endocrine secretion and its association with growth failure among HIV-infected children. PMID- 15295696 TI - Associations between IL12B polymorphisms and tuberculosis in the Hong Kong Chinese population. AB - BACKGROUND: Interleukin (IL)-12 plays a vital role in regulating cell-mediated immunity against tuberculosis (TB). METHODS: To test whether IL12B genetic polymorphisms might contribute to human TB susceptibility, we examined the genotype frequencies of 5 IL12B polymorphisms (at promoter, intron 2, intron 4, exon 5, and 3' untranslated region [UTR]) in 516 patients with TB and 514 healthy control subjects from the Hong Kong Chinese population. RESULTS: Individuals homozygous for the IL12B intron 2-repeat marker (ATT)8 had a 2.1-fold increased risk of developing TB (P < .001) (odds ratio, 2.14 [95% confidence interval, 1.45 3.19]). Estimation of the frequencies of multiple-locus haplotypes composed of IL12B promoter, intron 2, intron 4, and 3' UTR alleles revealed potential risk haplotypes (designated "A" and "K") and protective haplotypes (designated "B") for TB. Furthermore, combining the genotype data of the 4 informative IL12B loci revealed a strong association between a specific genotype pattern, termed "diplotype I" (heterozygous A and K haplotypes), and TB. In contrast, diplotype II (homozygous BB haplotypes) appeared protective against TB. CONCLUSIONS: These findings support the association between IL12B intron 2 polymorphism and TB and between specific IL12B haplotypes and TB. PMID- 15295697 TI - Association between vitamin D receptor gene polymorphisms and response to treatment of pulmonary tuberculosis. AB - BACKGROUND: Polymorphisms in the gene that encodes the vitamin D receptor (VDR) may influence the host response to Mycobacterium tuberculosis infection. METHODS: In a Peruvian community with a high incidence of tuberculosis (TB), VDR TaqI and FokI polymorphisms were compared among 103 patients with pulmonary TB and 206 matched healthy control subjects. Associations of VDR polymorphisms with treatment outcome were analyzed among 78 patients undergoing treatment of pulmonary TB. RESULTS: Sputum mycobacterial culture and auramine stain conversions were significantly faster among participants with the FokI FF genotype, compared with participants with the non-FF genotypes. Sputum culture conversion was faster among participants with the TaqI Tt genotype, compared with those with the TT genotype. Increased probability of culture conversion during TB treatment was independently associated with the TaqI Tt genotype (age- and sex adjusted relative risk, 4.28; 95% confidence interval, 1.88-9.75; P = .001). VDR polymorphisms were not significantly associated with susceptibility to TB in the case-control study. CONCLUSIONS: VDR gene polymorphisms are associated with the time to sputum culture and auramine stain conversion during TB treatment. To our knowledge, the present study is the first report of a specific host gene influence on the outcome of TB treatment. These findings demonstrate the potential clinical relevance of immunomodulatory functions of vitamin D metabolites acting via the VDR in the host response against pulmonary TB. PMID- 15295698 TI - Level of maternal IgG anti-group B streptococcus type III antibody correlated with protection of neonates against early-onset disease caused by this pathogen. AB - The present study estimates the level of maternal immunoglobulin (Ig) G anti group B streptococcus (GBS) type III required to protect neonates against early onset disease (EOD) caused by this pathogen. Levels of maternal serum IgG anti GBS type III, measured by enzyme-linked immunosorbent assay, in 26 case patients (neonates with EOD caused by GBS type III) and 143 matched control subjects (neonates colonized by GBS type III who did not develop EOD) of > or = 34 weeks gestation were compared. The probability of EOD decreased with increasing levels of maternal IgG anti-GBS type III (P = .01). Neonates whose mothers had > or = 10 microg/mL IgG anti-GBS type III had a 91% lower risk for EOD, compared with those whose mothers had levels of < 2 microg/mL. A vaccine that induces IgG anti-GBS type III levels of > or = 10 microg/mL in mothers can be predicted to offer a significant degree of protection against EOD caused by this pathogen. PMID- 15295699 TI - Cardiac valves in patients with Whipple endocarditis: microbiological, molecular, quantitative histologic, and immunohistochemical studies of 5 patients. AB - The pathological features of Whipple endocarditis, which is caused by Tropheryma whipplei, were histologically evaluated in cardiac valves from 5 patients. We used quantitative image analysis to compare the valvular fibrosis, calcifications, vegetations, inflammation, and vascularization due to Whipple endocarditis with those due to non-Whipple endocarditis and degenerative valves. We also studied the presence of T. whipplei in valves by immunohistochemical analysis, culture, and polymerase chain reaction (PCR). In histologic analysis, Whipple endocarditis was characterized by significant fibrosis, a lack of calcifications, slight inflammation and vascularization, and vegetations of intermediate size. Inflammatory infiltrates consisted mainly of foamy macrophages and lymphocytes. We found that the detection of T. whipplei in cardiac valves, by immunohistochemical analysis, was correlated with the detection of the bacterium by culture and PCR. We report, for the first time, the immunodetection of T. whipplei in a surgically removed arterial embolus. Pathological and immunohistologic analyses may contribute to the diagnosis of Whipple endocarditis. PMID- 15295700 TI - Transcription profile of Helicobacter pylori in the human stomach reflects its physiology in vivo. AB - BACKGROUND: Little is known about levels of expression of Helicobacter pylori genes in the human host. We therefore developed a quantitative real-time reverse transcriptase polymerase chain reaction (qRT-PCR) assay to measure transcript profiles of H. pylori in the human stomach. METHODS: In vivo expression of 16 genes on the cag pathogenicity island and of 18 putative virulence genes was quantitated by isolation of total RNA directly from infected human gastric mucosa. The results were compared with in vitro expression determined from H. pylori cells grown in culture. RESULTS: The highest levels of expression were found for cag1 and cag25 and for genes, such as urease and catalase, that may be important for bacterial homeostasis in the relatively hostile environment of the gastric mucosa. Transcript abundance, relative to 16S rRNA, was lower in vivo than in vitro, which suggests that H. pylori cells are in stationary phase in the gastric environment. This was particularly apparent for cagA. Since CagA is arguably of unique importance, in terms of interaction with the host, tight control of its in vivo expression might be particularly important. CONCLUSIONS: qRT-PCR is a powerful tool to measure gene expression in human or animal tissue that contains minute amounts of microbial mRNA, and the results reflect on the physiology of the pathogen in its natural host. PMID- 15295701 TI - Production of exopolysaccharide by Burkholderia cenocepacia results in altered cell-surface interactions and altered bacterial clearance in mice. AB - Despite the characterization of some Burkholderia cepacia complex exopolysaccharides (EPSs), little is known about the role of EPSs in the pathogenicity of B. cepacia complex organisms. We describe 2 Burkholderia cenocepacia (genomovar III) isolates obtained from a patient with cystic fibrosis (CF): the nonmucoid isolate C8963 and the mucoid isolate C9343. Both isolates had identical random amplified polymorphic DNA patterns. C9343 produced a capsule composed of the EPSs PS-I and PS-II, as well as alpha -1,6-glucan. These isolates exhibited several phenotypic differences: C8963 synthesized octanoyl-homoserine lactone and produced biofilms, but C9343 did not; in a mouse model of pulmonary infection, C8963 was cleared more rapidly than was C9343; and C9343 interacted poorly with macrophages and neutrophils, compared with C8963, suggesting that the C9343 capsule interfered with cell-surface interactions. Overproduction of EPS by C9343 resulted in a mucoid appearance and interfered with cell-surface interactions and clearance in an animal model. This mucoid colonial appearance could enhance the persistence and virulence of this important CF-related pathogen. PMID- 15295702 TI - Effects of glucose on fsr-mediated biofilm formation in Enterococcus faecalis. AB - Biofilm production is frequently dependent on such environmental factors as cell density and glucose concentration. The Enterococcus faecalis quorum-sensing locus (fsr) increases enterococcal virulence in multiple animal models. To date, fsr has been shown to regulate the transcription of 2 downstream protease genes. We demonstrate that the effect of fsr mutations on biofilm formation, as well as the fsr-mediated catabolite control of biofilm, is mediated via these proteases. The present study provides additional insight into the mechanisms used by E. faecalis to establish nosocomial infection. PMID- 15295703 TI - Correlation between Torque tenovirus infection and serum levels of eosinophil cationic protein in children hospitalized for acute respiratory diseases. AB - Children with bronchopneumonia have considerably higher Torque tenovirus (TTV) loads than do children with milder acute respiratory diseases (ARDs). Moreover, in children with ARDs, high TTV loads correlate with low percentages of circulating CD3+ and CD4+ T cells and with elevated percentages of B cells, suggesting that TTV might be immunomodulatory. Here, we show that, in children with ARDs, the presence of TTV and TTV load correlate with concentrations of serum eosinophil cationic protein. The possible mechanisms whereby TTV infection might lead to augmented activity of eosinophils and the implications for pathogenesis are discussed. PMID- 15295704 TI - Surveillance of clinical isolates of respiratory syncytial virus for palivizumab (Synagis)-resistant mutants. AB - Premature infants and those with chronic lung disease or congenital heart disease are at high risk of severe respiratory syncytial virus (RSV) disease. Palivizumab (Synagis), a humanized anti-RSV monoclonal antibody, has been used extensively since 1998 to prevent severe RSV disease in high-risk infants. To monitor for possible palivizumab-resistant mutants, an immunofluorescence binding assay that predicts palivizumab neutralization of RSV was developed. RSV isolates were collected at 8 US sites from 458 infants hospitalized for RSV disease (1998 2002). Palivizumab bound to all 371 RSV isolates able to be evaluated, including 25 from active-palivizumab recipients. The palivizumab epitope appears to be highly conserved, even in infants receiving prophylaxis with palivizumab. PMID- 15295705 TI - Peripheral blood lymphocytes express recombination-activating genes 1 and 2 during Epstein-Barr virus-induced infectious mononucleosis. AB - Implicit in the persistence of Epstein-Barr virus (EBV) in B lymphocytes is the successful circumvention of ongoing cell selection for competence of B cell receptors (BCRs). Because the EBV infection of B cells in vitro induces enzymatic machinery that is responsible for secondary immunoglobulin gene rearrangement, we examined the expression of the recombination-activating genes (RAGs) in peripheral blood mononuclear cells (PBMCs) from 26 patients with infectious mononucleosis (IM). RAG1 and/or RAG2 RNA was detected in PBMCs from 42% of patients with IM but not from healthy control subjects. EBV may usurp the cellular mechanism that diversifies the BCR, to guarantee a level of survival signaling sufficient for its own persistence. PMID- 15295706 TI - Quantitative analysis of Epstein-Barr virus (EBV)-specific CD8+ T cells in patients with chronic active EBV infection. AB - To clarify the pathogenesis of chronic active Epstein-Barr virus (EBV) infection, EBV-specific CD8+ T cells were enumerated, by use of human leukocyte antigen (HLA)-A*2402-restricted tetramers, in 8 patients with chronic active EBV infection, 10 patients with infectious mononucleosis, and 16 EBV-seropositive healthy control subjects. In most of the patients with chronic active EBV infection, EBV-specific CD8+ T cells were not detected. Of note, latent membrane protein 2-specific CD8+ T cells were not detectable in any patients with chronic active EBV infection. In contrast, EBV-specific CD8+ T cells were detected in patients with infectious mononucleosis and in healthy control subjects. Low frequencies of EBV-specific CD8+ T cells may be one of the immunological features of chronic active EBV infection. PMID- 15295707 TI - Expression of chemokine receptors on intrahepatic and peripheral lymphocytes in chronic hepatitis C infection: its relationship to liver inflammation. AB - BACKGROUND: Intrahepatic lymphocytes are believed to be directly involved in the immunopathogenesis of chronic liver diseases. Little is known about the trafficking of lymphocytes into the liver and their role in chronic hepatitis C infection. METHODS: The expression of 4 chemokine receptors and an activation marker on multiple lymphocyte subsets in paired liver biopsy and peripheral blood specimens from 23 patients with chronic hepatitis C infection were analyzed by a 6-color flow-cytometric assay. RESULTS: CCR5, CXCR3, and CXCR6 were expressed on intrahepatic CD4+ and CD8+ T cells, natural killer (NK) T cells, NK cells, and B cells at significantly higher frequencies than on peripheral lymphocyte subsets. The expression of these receptors and the activation marker CD38 tended to increase with the severity of liver inflammation. This increase was significant for several intrahepatic lymphocytes subsets. Correlations in expression differed among pairs of these extralymphoid homing receptors on the intrahepatic T cells. CONCLUSIONS: The homing program for intrahepatic lymphocytes involves multiple extralymphoid chemokine receptors that are regulated by >1 pathway. The expression of homing receptors on intrahepatic lymphocytes is associated with the immunopathogenesis of chronic hepatitis C disease. These preliminary results indicate that confirmational studies with larger sample sizes are warranted. PMID- 15295708 TI - Limited contribution of humoral immunity to the clearance of measles viremia in rhesus monkeys. AB - The development of an improved vaccine for controlling measles virus (MV) infections in the developing world will require an understanding of the immune mechanisms responsible for the clearance of this virus. To evaluate the role of humoral immunity in the containment of MV, rhesus monkeys were treated at the time of MV challenge with either anti-CD20 monoclonal antibody (MAb) infusion, to deplete B lymphocytes, or both anti-CD20 and anti-CD8 MAb, to deplete both B lymphocytes and CD8+ effector T lymphocytes. Although the MV-specific antibody response in CD20+ lymphocyte-depleted monkeys was delayed by >1 week, the kinetics of MV clearance did not differ from those for monkeys that received control MAb. Furthermore, unusual clinical sequelae of MV infection were not observed in these monkeys. In contrast, MV-infected rhesus monkeys depleted of both CD20+ and CD8+ lymphocytes had a prolonged duration of viremia and developed a desquamating skin rash. These findings indicate that humoral immunity plays a limited role in the control of MV replication in an MV-naive individual and suggest that new measles vaccination strategies should focus on the elicitation of cell-mediated immune responses, in addition to neutralizing antibodies, to facilitate rapid elimination of locally replicating virus. PMID- 15295709 TI - Hemoglobin C and resistance to severe malaria in Ghanaian children. AB - Hemoglobin (Hb) C has been reported to protect against severe malaria. It is unclear whether relative resistance affects infection, disease, or both. Its extent may vary between regions and with disease pattern. We conducted a case control study of children with severe malaria, asymptomatic parasitemic children, and healthy children in Ghana. HbAC did not prevent infection but reduced the odds of developing severe malaria and severe anemia. Protection was stronger with HbAS. The frequencies of HbCC and HbSC decreased, from healthy children to asymptomatic parasitemic children to children with severe malaria. These data support the notion that natural selection of HbC occurs because of the relative resistance it confers against severe malaria but argue against the notion that HbC offers resistance to infection. PMID- 15295710 TI - Identification of a conserved region of Plasmodium falciparum MSP3 targeted by biologically active antibodies to improve vaccine design. AB - Merozoite surface protein 3 (MSP3) is a target of antibody-dependent cellular inhibition (ADCI), a protective mechanism against Plasmodium falciparum malaria. From the C-terminal half of the molecule, 6 overlapping peptides were chosen to characterize human immune responses. Each peptide defined at least 1 non-cross reactive B cell epitope. Distinct patterns of antibody responses, by level and IgG subclass distribution, were observed in inhabitants of a malaria-endemic area. Antibodies affinity purified toward each peptide differed in their functional capacity to mediate parasite killing in ADCI assays: 3 of 6 overlapping peptides had a major inhibitory effect on parasite growth. This result was confirmed by the passive transfer of anti-MSP3 antibodies in vivo in a P. falciparum mouse model. T helper cell epitopes were identified in each peptide. Antigenicity and functional assays identified a 70-amino acid conserved domain of MSP3 as a target of biologically active antibodies to be included in future vaccine constructs based on MSP3. PMID- 15295711 TI - Protection against the early acute phase of Cryptosporidium parvum infection conferred by interleukin-4-induced expression of T helper 1 cytokines. AB - Immunity to Cryptosporidium parvum infection involves a T helper (Th) 1 response with interferon (IFN)- gamma and interleukin (IL)-12 activity, but the role of Th2 cytokines, such as IL-4, is unclear. Around the peak of infection, production of oocysts in IL-4-deficient and IL-4 receptor alpha -deficient neonatal BALB/c mice was greater than that in wild-type (wt) mice. Susceptibility to infection was increased or decreased, respectively, in wt mice treated with anti-IL-4 neutralizing antibodies or recombinant IL-4. Excretion of oocysts by IFN- gamma deficient mice was unaffected by treatment with anti-IL-4, indicating that IL-4 stimulated IFN- gamma activity. Early during infection, wt mice had increased intestinal expression of IFN- gamma and IL-12 mRNA, compared with IL-4-deficient mice. Intestinal IL-4 was detected by Western blotting in wt mice 24 h after infection but not in uninfected control mice. These findings suggest that, early during C. parvum infection of BALB/c mice, there is production of IL-4 that promotes Th1-mediated immunity. PMID- 15295712 TI - Modification of prostanoid secretion in endothelial cells by amphotericin B acting synergistically with interleukin-1beta: possible explanation of proinflammatory effects. AB - Human umbilical vein endothelial cells were exposed to free-form and lipid complexed versions of amphotericin B (alone or in combination with human recombinant interleukin [IL]-1 beta) and to culture medium from the human macrophage cell line THP-1 that had been exposed to amphotericin B. Endothelial cells were then incubated with exogenous-labeled arachidonic acid or were stimulated with histamine. Measurement of the resulting prostanoids indicated that amphotericin B and IL-1 beta acted synergistically to increase the ability of endothelial cells to synthesize prostanoids from endogenous and exogenous substrate and to increase expression of cyclooxygenase-2. This resulted in an increase of the ratio of untransformed prostaglandin (PG) H2 to PGI2 released by endothelial cells. Culture medium from amphotericin B-activated macrophages caused similar effects in endothelial cells. The synergistic effect with IL-1 beta was observed with free-form amphotericin B and, to a lesser extent, with lipid-complexed amphotericin B (Abelcet). Differences between Abelcet and the lipisome carrier (AmBisome) were not significantly different with respect to any of the parameters analyzed. PMID- 15295713 TI - Commentary: McIntosh K, Chao RK, Krause HE, Wasil R, Mocega HE, Mufson MA. Coronavirus infection in acute lower respiratory tract disease of infants. J Infect Dis 1974; 130:502-7. PMID- 15295714 TI - Oligomerisation of angiotensin receptors: novel aspects in disease and drug therapy. AB - The concept that 7 transmembrane receptors (7TMRs) exist and function as independent monomers has facilitated a therapeutic approach of selective targeting of receptors. However, oligomerisation of 7TMRs is now being recognised as an important biological phenomenon that adds a level of complexity to their signalling. In vitro, many 7TMR heterodimers display altered binding, signalling, and trafficking properties compared to their constituent monomeric units. This review discusses an emerging concept regarding the role of 7TMR heterodimerisation in vivo, and its significance to drug therapy. Studies of angiotensin receptor signalling indicate a pivotal role for heterodimerisation in the pathogenesis of human disorders. Furthermore, the occurrence of angiotensin receptor heterodimers has a profound effect on the responsiveness to treatment with 7TMR blockers. Global assessment of the prevalence of different heterodimers during disease and their responsiveness to drug therapy is likely to optimise patient treatment and reduce side-effects associated with 7TMR blockers. PMID- 15295715 TI - The utility of BNP in clinical practice. AB - Brain natriuretic peptide (BNP) is a cardiac neurohormone of increasing interest over recent years, with research applications expanding at a rapid rate and new data published on a monthly basis. Initially developed as a diagnostic aid for those with acute shortness of breath, clinical applications are now increasing, and this article reviews these clinical applications of BNP and the evidence for effectiveness of the synthetic BNP analogue nesiritide. PMID- 15295716 TI - A putative placebo comparison of the SCOPE and LIFE trials. AB - The Losartan Intervention For Endpoint reduction in hypertension (LIFE) trial and the Study on Cognition and Prognosis in the Elderly (SCOPE) superficially produced comparable outcomes, with effects on stroke greater than those anticipated from blood pressure (BP) lowering alone. This, however, ignores important features of both studies. It ignores firstly the disparate comparator agents - atenolol in LIFE and predominantly hydrochlorthiazide in SCOPE, secondly the small, but potentially important BP differential between the treatment arms in SCOPE and finally the small, statistically non-significant increase in coronary heart disease (CHD) in both trials. This analysis compares the major cardiovascular outcomes in these trials with reference to placebo. Two alternative reference populations were employed to calculate the imputed placebo, firstly the MRC Trial in Elderly Hypertensives and secondly a meta-analysis of trials in the elderly, which included comparisons between diuretic- and b-blocker based regimens. Overall, the choice of 'comparator placebo' did not substantially influence the derived results. Accounting for BP differences and based on the meta-analysis, both trials demonstrated statistically significant reductions in fatal/non-fatal stroke compared with placebo - relative risks (95% confidence intervals [CI]) of 0.53 (0.39, 0.73) and 0.56 (0.41, 0.76) for SCOPE and LIFE, respectively. For fatal/non-fatal MI, there were greater discrepancies between the studies, but with neither achieving statistical significance compared with placebo - relative risks of 0.85 (0.59, 1.24) and 1.08 (0.80, 1.46) for SCOPE and LIFE, respectively. This analysis clearly demonstrates that both candesartan in SCOPE and losartan in LIFE are associated with reductions in stroke events compared with placebo, greater than that observed in the well-established meta analysis of placebo-controlled hypertensive trials. However, the CIs are such that it is not possible to suggest definitively that this is a benefit beyond BP reduction alone. Neither trial is sufficiently 'powered' to demonstrate a benefit in CHD outcomes, but with SCOPE there was a trend towards benefit with a point estimate compatible with the major meta-analysis. PMID- 15295717 TI - Effect of candesartan and lisinopril alone and in combination on blood pressure and microalbuminuria. AB - BACKGROUND: Angiotensin-converting enzyme (ACE) inhibitors and angiotensin receptor blocking drugs (ARB) block the effect of angiotensin II by different mechanisms. It has been suggested that combined therapy may be more effective at reducing blood pressure (BP) than higher doses of either drug. METHODS: Twenty three elderly patients with systolic hypertension completed a double-blind crossover study comparing placebo, candesartan (C) 16 mg, C32 mg, lisinopril (L) 20 mg, L40 mg and C16 mg + L20 mg. Treatment periods were one month and ambulatory BP measurements were performed at the end of each period. The effects on albumin excretion in eight patients with microalbuminuria were determined. RESULTS: All treatments lowered BP. The falls in systolic and diastolic BP with C16, C32, L20 and L40 were similar. Plasma renin rose to a similar extent. A plateau effect was reached with C16 and L20. Systolic BP on the combination of C16 + L20 was lower than on each monotherapy (C16, 3.8 mmHg [p=0.002]; C32, 6.4 [0.0003]; L20, 2.9 [0.05]; L40, 3.3 [0.003]). The additional fall in BP with the combination appeared to be due to recruitment of non-responders, rather than to an additive effect in most patients. All treatments reduced microalbuminuria to a similar extent. The combination was well tolerated and there was no deterioration in renal function. CONCLUSION: When patients are on a plateau dose of an ACE inhibitor or an ARB, addition of the other drug class has a small but significant incremental effect on BP in the overall group. However, some patients respond better to one drug class than to the other and this may explain the results. This study lends no support to the use of these two drugs in combination to treat hypertension. PMID- 15295718 TI - Macroarray analysis in the hypertrophic left ventricle of renin-dependent hypertensive rats: identification of target genes for renin. AB - INTRODUCTION: The aim of this work was to identify new renin target genes in left ventricular hypertrophy during hypertension. MATERIALS AND METHODS: We compared left ventricle gene expression from four transgenic TGR(mRen2)27 (TG+/-) rats and four non-transgenic littermates (TG-/-) using cDNA macroarray. Hybridisation signals were quantified with a phosphorimager, and normalised to an external scale. Data analysis was performed with Statistical Analysis for Microarrays (SAM 1.21) software. The mRNA levels of candidate genes were determined by semi quantitative RT-PCR in three different hypertensive rats: TG+/-, spontaneously hypertensive (SHR) and genetically Lyon hypertensive (LH) rats, compared to their respective controls (TG-/-, Wistar-Kyoto, Lyon low blood pressure rats). RESULTS: Out of 1,200 genes present on the macroarray, 233 were reliably measured and only three were overexpressed (Biglycan, beta1-adenosine monophosphate-activated protein kinase [AMPK] and amyloid precursor like protein 2 [APLP2]) and 19 were underexpressed in the left ventricle of TG+/- compared with TG-/-. APLP2 is a member of the amyloid precursor protein (APP) family. APLP2 and APP mRNA levels were increased in TGR(mRen2)27 but significantly decreased in LH rats, while only APP was increased in SHR rats. CONCLUSIONS: We report new genes associated with renin-dependent left ventricular hypertrophy. Moreover, this work shows for the first time that the APP family gene expression could be altered in response to high renin activity and this effect is independent of cardiac remodelling and hypertension. PMID- 15295719 TI - Comparative study of the inhibitory effects of adrenomedullin on angiotensin II contraction in rat conductance and resistance arteries. AB - Adrenomedullin (ADM), a ubiquitous vasoactive peptide, has been the target of a multitude of studies concerning its effect on the vascular tone. The present work aims at clarifying a series of its interactions with the renin-angiotensin system. The study uses the rat aorta ring as a model of conductance vessels, with or without vascular endothelium, and the second order branch of rat mesenteric arteries as a model of resistance arteries. Interactions between various concentrations of ADM and angiotensin II (Ang II) were studied, in the presence of L-NAME (a nitric oxide [NO] synthase inhibitor) and methylene blue (MB; a soluble guanylate cyclase inhibitor). Results point out differences in the mechanism of the inhibitory action of ADM upon Ang II effects in the two vessel types studied. Inhibition of Ang II contraction by ADM involves guanylate cyclase in both cases. However, NO is involved in ADM-induced inhibition of angiotensinergic vasoconstriction only in the conductance arteries, not in the resistance ones. PMID- 15295720 TI - Tonic levels of angiotensin II reduce tonic levels of vascular nitric oxide even in salt-replete man. AB - INTRODUCTION: Losartan improves stimulated endothelial function in patients with cardiovascular disease, but there are no data to establish whether losartan has this effect in normal man. Furthermore, whether losartan improves basal nitric oxide (NO) activity is controversial. We therefore examined whether treatment with losartan improved basal NO activity in normal, salt-replete man. If so, this would imply that tonic levels of angiotensin II (Ang II) reduce tonic basal levels of NO, even in salt-replete normal man. METHODS: We performed a randomised, placebo-controlled, double-blind crossover study in 24 healthy volunteers, comparing losartan 50 mg daily for one month versus placebo. Brachial artery endothelial function was assessed by bilateral venous occlusion plethysmography, measuring the response to intra-arterial infusions of the endothelial-dependent and endothelial-independent vasodilators, acetylcholine and sodium nitroprusside respectively and the endothelial-dependent vasoconstrictor NG-monomethyl-L-arginine. Results were analysed by multiple analysis of variance and statistical significance was taken as a p value of 10(-9) M) inhibit. Ang II is secreted into the lumen in the proximal tubule and the concentration of Ang II in the proximal lumen has been reported to be in the nanomolar range, 100-1,000 times higher than in peritubular blood. We investigated the regulation of renal proximal fluid transport by luminal (predominantly locally produced) and peritubular capillary (circulatory) Ang II in anaesthetised rats, using a selective AT(1)-receptor antagonist, candesartan. METHODS: Experiments were performed in inactin anaesthetised male Wistar rats. Proximal fluid uptake rate was measured using computerised capture and analysis of shrinking-split droplet microperfusion in response to either luminal addition or luminal addition with simultaneously peritubular capillary perfusion of 10(-8) M candesartan. RESULTS: Luminal addition of candesartan (10(-8) M) decreased fluid absorption by 19-25%. Perfusion of the peritubular capillaries with an electrolyte solution (containing no Ang II) reduced fluid uptake by 27%, and blockade of the peritubular actions of Ang II by addition of candesartan (10(-8) M) resulted in 33% decrease in fluid uptake. However, when candesartan (10(-8) M) was added to both luminal and capillary perfusates, there was a 43% reduction in fluid transport when compared with initial values. CONCLUSION: These results suggest that the presence of endogenous Ang II in both peritubular blood and luminal fluid is important for maximal expression of the stimulatory influence of this peptide on proximal tubule fluid uptake. PMID- 15295722 TI - Losartan may prevent the elevation of plasma glucose, corticosterone and catecholamine levels induced by chronic stress. AB - INTRODUCTION: Stress is a stimulus that activates the hypothalamic pituitary adrenal (HPA) axis and sympathetic nervous system (SNS). Increased activity of the SNS causes to increment or impairment in blood pressure, heart rate, body temperature and plasma glucose and adreno- corticotrophic hormone (ACTH) levels. Angiotensin II (Ang II), which is a product of the renin-angiotensin system (RAS), is an important factor affecting the activity of the SNS and responses to stress. We suggest that the blockade of Ang II may be worthwhile in the prevention and treatment of diabetes mellitus and cardiovascular diseases affected by stress. Therefore, we investigated the effects of immobilisation stress on blood glucose, norepinephrine (NE), epinephrine (E) and corticosterone levels and the effects of an Ang II receptor antagonist, losartan, on these parameters. MATERIALS AND METHODS: The rats were kept in small cylindrical cages for 60 min/day for 10 consecutive days to perform chronic immobilisation stress. Losartan (10 mg/kg) was given daily by gavage to Losartan (L) and Losartan + Chronic Stress (L+CS) groups. Control (C) and Chronic Stress (CS) groups received an equal volume of saline daily by gavage for 10 days. After the last stress regimen, blood samples were collected for plasma glucose, NE, E and corticosteroid measurements. RESULTS: Plasma glucose, NE, E and corticosterone levels in the CS Group increased significantly compared with the C group. In Group L+CS, the plasma glucose, NE, E and corticosterone levels decreased significantly vs. Group CS. In Group L there was no significant difference vs. Group C. CONCLUSION: It can be speculated that chronic blockade of RAS may decrease the excess sympathetic responses to stress in cardiovascular diseases and prevent the likely development of Type II diabetes mellitus. PMID- 15295723 TI - Linkages of acute care and emergency medical services to state and local public health programs: the role of interactive information systems for responding to events resulting in mass injury. PMID- 15295724 TI - An evaluation of the use of guidelines in prehospital management of brain injury. AB - OBJECTIVES: The Brain Trauma Foundation (BTF) Guidelines for Prehospital Management of Traumatic Brain Injury (TBI) are intended to standardize treatment and improve outcomes in severe TBI patients. The key guideline components focus on airway management, blood pressure support, Glasgow Coma Score assessment, and transport. The purposes of this study were to determine if providers could learn and retain the guidelines (education), assess if providers would use the guidelines in practice (implementation), and evaluate the effect of guideline implementation on patients (outcomes). METHODS: Data were collected prospectively on all trauma patients for five months. Providers were then educated on the TBI guidelines over two months, and five additional months of data were collected. A knowledge test was given before and after the course and three months later to assess education. To assess implementation, data were analyzed to determine whether providers were using the key interventions more consistently after education. The clinical courses of TBI patients before and after guideline implementation were measured to assess outcomes. RESULTS: Knowledge of TBI care improved significantly after education and remained elevated at three months (62% vs. 82% vs. 79%, p < 0.001). For the 1,044 patients seen, providers demonstrated higher rates of appropriate care, resulting in lower rates of hypoxia (2.8% vs. 1.1%, p=0.010) and hypotension (4.8% vs. 2.0%, p=0.018). Mortality was significantly decreased (34.6% vs. 17.0%, p=0.039), and rates of patients with maximum functional scores at 14 days significantly increased (Glasgow Outcome Score 44.2% vs. 66.0%, p=0.025; Rancho Los Amigos Scale 55.9% vs. 77.3%, p=0.045). CONCLUSION: Providers were able to learn and implement the BTF guidelines, and outcomes in TBI patients were significantly improved. All emergency medical services providers should be trained in these potentially lifesaving guidelines. PMID- 15295725 TI - Efficacy of lower-energy biphasic shocks for transthoracic defibrillation: a follow-up clinical study. AB - OBJECTIVE: This clinical study prospectively evaluated the first-shock defibrillation efficacy of 150-joule impedance-compensated, 200-microF biphasic truncated exponential (BTE) shocks in patients with electrically-induced ventricular fibrillation (VF), and compared it with a historical control group treated with 200-J monophasic damped sine (MDS) shocks. METHODS: Ventricular tachyarrhythmias were induced in patients undergoing electrophysiologic (EP) testing for ventricular arrhythmias or testing of an implantable cardioverter defibrillator (ICD). A 150-J shock was delivered as the primary therapy to terminate induced arrhythmias in the EP group, and as a "rescue" shock when a single ICD shock failed to terminate the arrhythmias in the ICD group. RESULTS: Ninety-six patients received study shocks. The preshock rhythm was classified as VF in 77 patients and as ventricular tachycardia (VT) in 19 patients. First-shock success rates for VF and VT were 75 out of 77 (97.4%) and 19 out of 19 (100%) for the 150-J BTE compared with the historical control rates of 61 out of 68 (89.7%) and 29 out of 31 (94%) for 200-J MDS. The first-shock success rate for VF treated with 150-J BTE was technically equivalent to that of 200-J MDS (p=0.001). The transthoracic impedance did not vary between groups, yet the peak current delivered by the 150-J BTE shock was about 50% lower. CONCLUSIONS: This study demonstrated that 150-J shocks of this impedance-compensated, 200-microF BTE waveform provided very high efficacy for defibrillation of short duration, electrically-induced VF. These lower-energy biphasic shocks had a success rate equivalent to that of 200-J MDS shocks, and they provided this efficacy while exposing patients to much less current than the monophasic shocks. PMID- 15295726 TI - Trauma center versus non-trauma center admissions in adult trauma victims by age and gender. AB - OBJECTIVE: The admission types and appropriateness of admission of adults with differing levels of injury severity were compared, based on patient age and gender. METHODS: This retrospective study used a statewide hospital discharge dataset. The patients included adults who had sustained trauma related to motor vehicles and were admitted to trauma center (TCs) and non-trauma center (NTCs) hospitals. Using injury severity scores (ISSs) >or= 16 to denote major trauma, the proportion of patients with major traumatic injuries who were admitted to TCs and NTCs was determined. Types of admission (TC versus NTC) were compared by age and gender for four subgroups of men and women, aged 25 to 64 years, and aged 65 years and older. RESULTS: The sample included 5,712 patients. Of those patients with ISS >or= 16, younger men were most likely to be admitted to a TC (82%), and older women were least likely to be admitted to a TC (60%). The proportions of older men and women with ISS >or= 16 who were admitted to a TC were comparable. Among patients with ISS < 16 admitted to NTCs, older women were the highest proportion (65%), and younger men were the lowest proportion (43%). Overall, more older men and women with ISS >or= 16 were admitted to NTCs than would have been expected. Conversely, a statistically significant proportion of younger men and women with ISS<16 were admitted to TCs. CONCLUSION: Findings suggest that older trauma victims whose injuries appear to be serious are admitted to NTCs more often than are younger trauma victims with similarly serious injuries. Additional studies to examine this phenomenon are warranted. PMID- 15295727 TI - Comparison of etomidate and midazolam for prehospital rapid-sequence intubation. AB - OBJECTIVE: This study compares etomidate with midazolam for prehospital rapid sequence intubation (RSI). METHODS: The authors conducted a retrospective review of consecutive intubations at a university-based air medical program from January 1995 to December 2000. Exclusion criteria were patients not undergoing RSI, age <15 years, and incomplete chart data. Outcome measures included intubation success, incidence of hypotension, and percentage of change in heart rate (HR) and systolic blood pressure (SBP). RESULTS: The intubation success rate was 110 out of 112 (98%) with etomidate, and 96 out of 97 (99%) with midazolam. Mean ages, patient gender distributions, and initial SBPs and HRs did not differ between the two groups. The mean dose of etomidate was 24 mg, the mean percentage of change in HR was -1% (95% confidence interval [CI], -6 to 4), and the mean percentage of change in SBP was 2% (95% CI, -3 to 7). The mean dose of midazolam was 3.5 mg, the mean percentage of change in HR was 1% (95% CI, -5 to 7), and the mean percentage change in SBP was 3% (95% CI, -3 to 9). The number of hypotensive episodes with etomidate (7 out of 74) compared with midazolam (3 out of 56) did not differ significantly (Fisher's exact test, p=0.51). CONCLUSION: Intubation success rate was very high with both etomidate (98%) and midazolam (99%). There was no statistically significant mean percentage of change in SBP or HR with either agent. The authors found a low incidence of hypotension with both agents, although the mean dose of midazolam used was considerably less than typically recommended for induction. PMID- 15295728 TI - Prevalence of automated external defibrillators at cardiac arrest high-risk sites. AB - OBJECTIVE: Automated external defibrillators (AEDs) distributed throughout communities may improve survival from cardiac arrest. The purpose of this study was to determine if AEDs were present at high-risk locations for cardiac arrest in King County, Washington. METHODS: The authors compiled a list of sites based on a five-year study that identified public sites with the highest incidence of cardiac arrests in King County. They conducted a structured telephone survey with the manager, director, or owner of those high-risk sites. RESULTS: Of the 263 identified high-risk cardiac arrest sites, we obtained information for 228 (87%) sites. Overall, 87 of 228 (38%) high-risk sites had one or more AEDs. The AED dissemination varied greatly by type of site. The airport, the two county jails, the five public sports venues, and the nine ferries/train terminals each reported at least one AED on site. In contrast, none of the 13 shelters and 19% of health clubs/gyms reported an AED on site. Nearly half (44%) of sites without AEDs cited cost as a factor preventing them from purchasing AEDs in the future. CONCLUSION: Although AEDs have diffused into high-risk sites in this community, the diffusion appears to vary by the type of site. PMID- 15295729 TI - Automated external defibrillator use by untrained bystanders: can the public-use model work? AB - OBJECTIVE: For automated external defibrillators (AEDs) to be practical for broad public use, responders must be able to use them safely and effectively. This study's objective was to determine whether untrained laypersons could accurately follow the visual and voice prompt instructions of an AED. METHODS: Each of four different AED models (AED1, AED2, AED3, and AED4) was randomly assigned to a different group of 16 untrained volunteers in a simulated cardiac arrest. Four usability indicators were observed: 1) number of volunteers able to apply the pads to the manikin skin, 2) appropriate pad positioning, 3) time from room entry to shock delivery, and 4) safety in terms of touching the patient during shock delivery. RESULTS: Some of the 64 volunteers who participated in the study failed to open the pad packaging or remove the lining, or placed the pads on top of clothing. Fifty-percent of AED2 pads and 44% of AED3 pads were not placed directly on the manikin skin compared with 100% of AED1 and AED4 pads. Adjacent pad displacements that potentially could affect defibrillation efficacy were observed in 6% of AED1, 11% of AED2, 0% of AED3, and 56% of AED4 usages. Time to deliver a shock was within 3.5 minutes for all AEDs, although the median times for AED1 and AED4 were the shortest at 1.6 and 1.7 minutes, respectively. No significant volunteer contact with the manikin occurred during shock delivery. CONCLUSIONS: This study demonstrated that the AED user interface significantly influences the ability of untrained caregivers to appropriately place pads and quickly deliver a shock. Avoiding grossly inappropriate pad placement and failure to place AED pads directly on skin may be correctable with improvements in the AED instruction user interface. PMID- 15295730 TI - Mathematical determination of external defibrillators needed at mass gatherings. AB - OBJECTIVE: To develop a mathematical formula that assists in determining the number of automated external defibrillators (AEDs) needed at sites of mass gatherings. METHODS: Twenty (10 male, 10 female) healthy volunteers (equally divided between age groups 21-30 and 31-40 years) responded to mock cardiac arrests in a sports stadium. Seven different first-responder scenarios were simulated (ascending and descending three separate stairway slopes (22 degrees, 39 degrees, and 69 degrees ), as well as a response across a horizontal (0 degrees ) surface. To assess the impact of spectator congestion, the same volunteers conducted each scenario in an empty and full stadium. The quantitative relationship between time and distance was then plotted for each situation. Using the quantitative relationship, the area a first responder can cover in a specified time was calculated. RESULTS: The formula for the total number of AEDs needed in a stadium (or other mass gathering site) can be expressed as follows: Total AEDs=[A(1)/(Ds(1)xDh(1))]+[A(2)/(Ds(2)xDh(2))]+[A(3)/(Ds(3)xDh(3))] where A(1), A(2), and A(3) represent the total areas of a stadium with a slight, moderate, or steep stairway slope, respectively; Ds(1), Ds(2), and Ds(3) represent the stairway distance a first responder must ascend or descend for each slope; and Dh(1), Dh(2), and Dh(3) are the horizontal distances a responder can run in the time remaining. CONCLUSION: Given a medical director's targeted response times and goals, the optimal number of AEDs required at a mass gathering can be calculated using time versus distance relationships. Future studies should evaluate the impact of the mathematically derived optimal number of AEDs at mass gatherings. PMID- 15295732 TI - An evaluation of paramedics' ability to screen older adults during emergency responses. AB - OBJECTIVES: This study evaluated the feasibility of using the emergency medical services (EMS) system as a public health provider by having paramedics screen older adults (age >or= 65 years) for influenza immunization status during emergency responses. It also determined the proportion of older-adult EMS patients who lacked an influenza vaccination. METHODS: A retrospective descriptive study was performed, with medical-record review for patients treated between January 2003 and April 2003. Patients were included if they were age 65 years and older, requested assistance via a 9-1-1 call, and were treated by one of 13 paramedics using a directed medical record. The authors calculated the proportion of patients successfully screened and the proportion who reported being nonimmunized. They also compared the patients screened and not screened by the EMS providers and patients who reported being immunized and reported being nonimmunized. RESULTS: Two hundred eighty-eight patients were eligible; the median age was 80 years, 53% were women, 73% were white, and 59% required advanced life support care. Paramedics successfully screened 177 patients (61%; 95% CI, 56-67%). Sixty-five patients (37%; 95% CI, 30-44%) reported being nonimmunized. Failure to screen was associated with a Glasgow Coma Scale score of 13 or less. Lack of immunization was associated with younger age and female gender. CONCLUSION: Paramedics can screen a majority of older adults for influenza immunization status during emergency responses. Older adult users of EMS reported lacking influenza vaccination at levels similar to national estimates. An EMS-based, paramedic-implemented screening program has the potential to identify older adults at risk for preventable illnesses and to augment traditional screening programs, but additional measures are needed to enhance screening rates. PMID- 15295733 TI - New paradigm for protection: the emergency ambulance services in the time of severe acute respiratory syndrome. AB - Severe acute respiratory syndrome (SARS) is a newly emerging and highly infectious form of atypical pneumonia with a high rate of transmission, especially among health care workers. With SARS, certain policies had to be implemented rapidly by the emergency ambulance services and the Ministry of Health to support and protect all personnel adequately. The authors discuss the changes in policies and personnel behavior, the training and education that had to be disseminated widely, and certain alternatives in policies such as transportation. The authors hope to share their experience in the implementation of these strategies by the Singapore Civil Defence Force and stress the importance of the psychological preparedness of the paramedics and prehospital care providers worldwide in this era of SARS. PMID- 15295734 TI - Development of emergency medical services in Guatemala. AB - Guatemala has recently undergone many advances in emergency medical services (EMS) training and disaster management. Industrialization and demographic changes have led to a continuing decline in the prevalence of infectious disease, while trauma and cardiovascular-related deaths have become increasingly important. Trauma now accounts for the nation's single greatest cause of productive years of life lost, a major indicator of a disease's impact on society. This "demographic transition" has dramatically increased the number of incidents where early prehospital intervention can have a positive impact on morbidity and mortality. However, until recently, prehospital medical care was provided by firefighters, who lacked formal medical training. Responding to a perceived need, increased collaborative efforts between prehospital care providers and governmental and nongovernmental agencies have rapidly improved provider training, initiated care standardization, and improved disaster preparedness. These efforts may serve as a model to other developing nations seeking to improve their EMS systems. PMID- 15295735 TI - High-dose steroids for acute spinal cord injury in emergency medical services. PMID- 15295736 TI - National Association of EMS Physicians Board Position Paper Review Actions 2002 2003. PMID- 15295737 TI - Pre-EMS education and instructor development. PMID- 15295738 TI - Helicopter emergency medical services transport outcomes literature: annotated review of articles published 2000-2003. AB - Helicopter emergency medical services (HEMS) and its possible association with outcomes improvement continues to be a subject of debate. As is the case with other scientific endeavors, debate over HEMS usefulness should be framed around an evidence-based assessment of the relevant literature. In an effort to facilitate the academic pursuit of assessment of HEMS utility, in late 2000 the National Association of EMS Physicians' Air Medical Committee prepared annotated bibliographies of the HEMS-related outcomes literature. As a result of that work, two review articles-one covering HEMS use in nontrauma and the other in trauma published in 2002 in Prehospital Emergency Care surveyed HEMS outcomes-related literature published between 1980 and mid-2000. Given the broad interest in the earlier reviews, and the increasing rate of publication of HEMS studies, the current project was executed with the intent of updating the annotated HEMS outcomes-related bibliography, covering a three-year time interval (through 2003) since the prior reviews. PMID- 15295743 TI - Resistance training effectiveness in patients with Charcot-Marie-Tooth disease: recommendations for exercise prescription. AB - OBJECTIVES: To determine the effects of a 12-week, home-based resistance exercise program on strength, body composition, and activities of daily living (ADLs) in men and women with Charcot-Marie-Tooth (CMT) disease and to design an ADL-based resistance exercise prescription template. DESIGN: Double-blind, placebo controlled study. SETTING: Testing in a university setting; exercise in patients' homes. PARTICIPANTS: Twenty CMT patients who volunteered. INTERVENTION: Subjects progressively strength trained at home 3 d/wk for 12 weeks. MAIN OUTCOME MEASURES: Timed ADLs, isometric strength, and body composition. RESULTS: Absolute strength was greater in men with CMT in only 4 of 10 baseline measures (P<.05), but not when strength was normalized by lean mass. Training compliance was 87% with no gender differences. At baseline, women had 80% of normal strength in 4 of 10 measures, whereas men did not achieve 80% of normal strength in any measure. After training, women had 80% of normal strength in 8 of 10 measures, whereas men only had 80% of normal strength in 1. Training volumes and strength change scores showed no gender differences. ADLs improved after training with no gender differences (P<.05). An exercise prescription template was developed by using chair-rise time to estimate starting weights for lower body and supine rise for upper body. CONCLUSIONS: Resistance training improved strength and ADLs equally in men and women. We designed an exercise prescription recommendation, based on ADL performance. PMID- 15295744 TI - Plantar- and dorsiflexor strength in prepubertal girls with juvenile idiopathic arthritis. AB - OBJECTIVE: To compare lower-leg strength of young girls with polyarticular juvenile idiopathic arthritis (JIA) with that of healthy, age-matched controls. DESIGN: Isometric and isokinetic strength tests of the plantar- and dorsiflexors. All strength measures were made at an ankle angle of 90 degrees. Isokinetic plantar- and dorsiflexor measures were made at 15 degrees/s during shortening (concentric) and lengthening (eccentric) actions. SETTING: Strength testing laboratory. PARTICIPANTS: Ten prepubertal girls diagnosed with JIA and 10 healthy girls. INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURES: Isometric and isokinetic plantar- and dorsiflexor strength. RESULTS: Isometric plantar- and dorsiflexion torques were significantly lower (48% and 38% respectively; P<.05) for the children with JIA than for the controls. The JIA group also produced lower shortening plantarflexion torques (52%, P<.05). Lengthening plantarflexor torques did not differ significantly between the 2 groups (P<.05). Controls were stronger than the JIA group for both shortening and lengthening maximal dorsiflexor actions (P<.05). All children were 4 to 5 times stronger in plantarflexion than in dorsiflexion. CONCLUSIONS: Girls with JIA had significantly less plantar- and dorsiflexor strength than age-matched, healthy peers. The reduced strength of children with JIA is likely to affect function in daily activities and probably contributes to reduced levels of physical activity. PMID- 15295745 TI - Conservative treatment for shoulder pain: prognostic indicators of outcome. AB - OBJECTIVES: To investigate the long-term clinical outcome and to identify factors that predict that outcome, after conservative treatment of patients who have shoulder pain with or without accompanying stiffness. DESIGN: Cohort study. SETTING: Outpatient clinic. PARTICIPANTS: Eighty-two subjects who had participated in a randomized controlled trial that compared the short-term effectiveness of conservative treatment for chronic, unilateral shoulder pain of mechanical origin with and without accompanying stiffness, and who were available for longer term follow-up 6 months after the cessation of formal treatment. INTERVENTIONS: Conservative treatment consisting of various combinations of exercise therapy, passive joint mobilization, electrophysical modalities, and corticosteroid injections. MAIN OUTCOME MEASURES: Pain intensity, functional limitation, perceived change in symptoms, active range of motion, muscle force, and clinical and demographic variables. RESULTS: Patients showed significant improvement in all outcome measurements in the long-term whether or not their shoulder pain was accompanied by stiffness. Long-term outcome was not predicted by hand dominance, clinical history of the shoulder condition, severity of the shoulder problem, or shoulder mechanics. CONCLUSIONS: Patients with chronic shoulder pain, with or without accompanying stiffness, can expect significant decreases in shoulder pain and improvements in shoulder function in the long term after conservative treatment. PMID- 15295746 TI - Manipulation and arthroscopy under general anesthesia and early rehabilitative treatment for frozen shoulders. AB - OBJECTIVE: To evaluate the efficacy of manipulation followed by arthroscopic release of the glenohumeral joint in conjunction with an immediate and intensive rehabilitation program. DESIGN: Retrospective, descriptive study. SETTING: A free standing, university-affiliated orthopedics and rehabilitation hospital. PARTICIPANTS: Forty patients with a clinical diagnosis of adhesive capsulitis resistant to pharmacologic and physical therapy (PT). INTERVENTIONS: Patients underwent manipulation and arthroscopic release of the capsular joint and were given an intensive PT program on the first postoperative day. MAIN OUTCOME MEASURES: All patients were evaluated pre- and postoperatively at follow-up at an average of 42 months by using the Simple Shoulder Test (SST), the Constant-Murley system score, and passive (PROM) and active (AROM) range of motion. RESULTS: The SST, which showed a mean preoperative score of 2.2+/-0.7, was 10.8+/-0.7 (P<.001) after surgery. Preoperatively, the mean Constant-Murley score was 33.2%+/-1.9%; postoperatively, the mean score was 91.7%+/-2.9% (P<.001). PROM increased from 90 degrees to 165 degrees for anterior elevation, from 85 degrees to 160 degrees for abduction, from 20 degrees to 60 degrees for external rotation, and from 10 degrees to 40 degrees for internal rotation. AROM improved for anterior elevation from 82 degrees to 155 degrees; for abduction from 77 degrees to 143 degrees, and for external rotation, with the arm along the patient's side, from 5 degrees to 50 degrees. CONCLUSIONS: Results support the efficacy of manipulation follow by arthroscopic release and rehabilitative treatment for patients with resistant adhesive capsulitis of the shoulder. PMID- 15295747 TI - Cryotherapy does not impair shoulder joint position sense. AB - OBJECTIVE: To determine the effects of a cryotherapy treatment on shoulder proprioception. DESIGN: Crossover design with repeated measures. SETTING: University athletic training and sports medicine research laboratory. PARTICIPANTS: Thirty healthy subjects (15 women, 15 men). INTERVENTION: A 30 minute cryotherapy treatment. MAIN OUTCOME MEASURES: Joint position sense was measured in the dominant shoulder by using an inclinometer before and after receiving 30 minutes of either no ice or a 1-kg ice bag application. Skin temperature was measured below the tip of the acromion process and recorded every 5 minutes for the entire 30 minutes and immediately after testing. Three different types of error scores were calculated for data analyses and used to determine proprioception. RESULTS: Separate analyses of absolute, constant, and variable error failed to identify changes in shoulder joint proprioception as a function of the cryotherapy application. CONCLUSIONS: Application of an ice bag to the shoulder does not impair joint position sense. The control of proprioception at the shoulder may be more complex than at other joints in the body. Clinical implications may involve modifying rehabilitation considerations when managing shoulder injuries. PMID- 15295748 TI - Virtual reality and haptics as a training device for movement rehabilitation after stroke: a single-case study. AB - OBJECTIVE: To investigate whether training in a virtual environment with a haptic device will improve motor function in the left hemiparetic arm of a stroke subject. DESIGN: Single case, A-B-A design. SETTING: University hospital research laboratory. PARTICIPANT: A man in his late fifties (right handed), with a right hemisphere lesion that caused a deficit in the left upper extremity. INTERVENTION: The subject trained with a 3-dimensional computer game during a 4 week period that consisted of twelve 90-minute sessions. MAIN OUTCOME MEASURES: Three tests (Purdue pegboard test, dynamometer hand-grip strength, upper extremity test) and a subjective interview were used to evaluate motor performance. RESULTS: Improvements were found in fine manual dexterity, grip force, and motor control of the affected upper extremity. The subject reported that there was a change in his day-to-day use of the upper extremity and that he was able to use it in activities that were previously impossible for him. CONCLUSIONS: Training with virtual reality and haptics can promote motor rehabilitation. PMID- 15295749 TI - Quantitative evaluation of long sitting in paraplegic patients with spinal cord injury. AB - OBJECTIVES: To evaluate the characteristics of long sitting (ie, sitting with legs extended) in patients with spinal cord injury (SCI) and to compare these results with able-bodied control subjects. DESIGN: A kinematic study using a video camera and forceplate with a strain-gauge type load cell. SETTING: A referral center for patients with SCI in Japan. PARTICIPANTS: Twenty-four subjects, including 11 able-bodied, matched control subjects and 13 SCI patients with complete paraplegia. INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURES: Sitting posture in the sagittal plane as well as the movement pattern and distance of the center of pressure (COP). RESULTS: Patients with SCI kept their pelvis tilted posteriorly and the lumbar spine was less lordotic during long sitting. The changing COP pattern during long sitting differed in able-bodied subjects as compared patients with SCI. During long sitting with arms outstretched over the thighs, COP movement in the subjects with SCI was significantly greater than that in the able-bodied subjects. When the arms were outstretched over the thighs, the COP shifted anteriorly in the able-bodied subjects and posteriorly in the patients with SCI. CONCLUSIONS: Long sitting in the paraplegic patients with SCI was unstable compared with the able-bodied subjects. The COP distribution pattern differed significantly between the 2 groups. The support and function of the upper extremities may influence balance during long sitting in the patients with SCI. The method of seating evaluation using a video camera and gravicorder was easy to use and appeared to provide an objective measurement of dynamic seating function in the patients with SCI. PMID- 15295750 TI - Patterns of recurrent pressure ulcers after spinal cord injury: identification of risk and protective factors 5 or more years after onset. AB - OBJECTIVES: To identify risk and protective factors associated with a history of recurrent pressure ulcers among participants with spinal cord injury (SCI). DESIGN: A mail survey was used to identify factors associated with the presence or absence of recurrent pressure ulcers. SETTING: A large specialty hospital in the southeastern United States. PARTICIPANTS: All participants had traumatic SCI, were nonambulatory, 18 years or older, and had been injured at least 5 years. A total of 826 subjects participated, 633 of whom reported a pressure ulcer history that could be classified as to whether they did or did not have a history of recurrent pressure ulcers. INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURES: History of pressure ulcers was measured by a single item that required participants to classify their history into 1 of 5 options, ranging from never having any pressure ulcers to having almost continuous pressure ulcers, often requiring hospitalization. Those who either never had a pressure ulcer or had them mostly for a short period after SCI onset were classified as nonrecurrent, whereas those who reported at least 1 per year were classified as recurrent. RESULTS: Seventy percent of the participants failed to report recurrent pressure ulcers (never had any or had them only immediately after SCI onset), whereas 13% reported a clear pattern of recurring pressure ulcers of 1 or more per year. Logistic regression analyses suggested several general behaviors were protective for recurrent pressure ulcers, including lifestyle, exercise, and diet. Yet none of the behaviors generally recommended during inpatient rehabilitation specifically to prevent pressure ulcers (eg, skin checks weight shifts) were associated with pressure ulcer history. Only 2 risk behaviors were identified (number of cigarettes smoked, use of medication for sleep), although several proxy variables were related to pressure ulcer history. CONCLUSIONS: Pressure ulcer history is a more viable measure of pressure ulcer outcomes than measures taken at a single point in time (current), over a brief period (eg, 1y), or those relying on critical events occurring at any time since SCI onset (ie, surgeries to repair pressure ulcers). A healthy lifestyle appears to be strongly associated with avoiding pressure ulcers, whereas the efficacy of specific prevention behaviors was not demonstrated. Problem solving and coping strategies should be targets for further research. PMID- 15295751 TI - Can functional electric stimulation-assisted rowing reproduce a race-winning rowing stroke? AB - OBJECTIVE: To compare the ergometer rowing technique of a person with spinal cord injury (SCI), using functional electric stimulation (FES) of his leg muscles, with that of a well-defined group of able-bodied rowers. DESIGN: Whole-body kinematics and kinetics and electric activity of selected muscles were measured during ergometer rowing. SETTING: A hospital-based motion analysis laboratory. PARTICIPANTS: Five male university varsity-level rowers and 1 male rower with SCI. INTERVENTIONS: Eight rowing trials were collected on the university-level rowers, 2 trials each at 20, 24, 28, and 32 strokes/min. The rower with SCI had surface electrodes applied to his medial hamstrings and medial quadriceps muscle bellies. The electrodes were attached to a stimulator that was activated using a button in the ergometer handle. The subject with SCI rowed at a self-selected stroke rate. MAIN OUTCOME MEASURES: Forces at the ergometer handle and foot cradle, 3-dimensional whole-body kinematics, net joint moments, and phasic activity of muscles. RESULTS: Motion of the arms, ankles, and knees of the rower with SCI was similar to those of the university-level rowers; other joint motions and forces applied to the ergometer differed. CONCLUSIONS: FES-assisted rowing in its current implementation cannot reproduce a race-winning rowing stroke. Further development work is required. PMID- 15295752 TI - Reproducibility of maximal quadriceps strength and its relationship to maximal voluntary activation in postpoliomyelitis syndrome. AB - OBJECTIVES: To determine what changes in maximal isometric strength can be detected in a symptomatic quadriceps muscle in patients with postpoliomyelitis syndrome (PPS) and to investigate the association between the variability in maximal strength and maximal voluntary activation (MVA). DESIGN: Repeated measures over a 3-week interval. SETTING: University hospital. PATIENTS: Convenience sample of 65 patients with PPS. INTERVENTION: Dynamometer testing. MAIN OUTCOME MEASURES: Maximal voluntary contraction (MVC) torque of the quadriceps was measured with a Kin-Com dynamometer and MVA was determined by twitch interpolation. RESULTS: The mean difference between the 2 consecutive measurements was -0.7+/-12.8 Nm (95% confidence interval [CI], -3.9 to 2.5). The test-retest reliability was excellent for MVC torque (intraclass correlation coefficient [ICC]=.96; 95% CI,.93-.98) and moderate for MVA (ICC=.73; 95% CI,.56 .85). The smallest detectable change in MVC torque was 25% for an individual. The variability in MVA explained 18% of the variability in maximal strength. CONCLUSIONS: Variability in maximal quadriceps strength, measured with a fixed dynamometer, was large and partly related to variability in MVA. This implies that even with optimally standardized strength testing, a follow-up of many years is required to objectify progression of quadriceps weakness in an individual patient with PPS. To demonstrate changes in strength in groups of patients in follow-up or intervention studies, feasible sample sizes are required. PMID- 15295753 TI - Lower-limb force production in individuals after unilateral total knee arthroplasty. AB - OBJECTIVE: To determine and compare lower-limb force production during a single leg horizontal press in involved and uninvolved limbs in people with unilateral total knee arthroplasty (TKA) and the limb of age- and gender-matched subjects. DESIGN: Quasi-experimental, posttest only control group design. SETTING: Subjects living in the community and were tested at a university facility. PARTICIPANTS: Nine people with unilateral TKA and 9 subjects without TKA volunteered. The mean postsurgery time for the TKA group was 15.89+/-6.62 months. INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURE: Force production in kilograms during a single leg press. RESULTS: Lower-limb force production differed significantly between involved and uninvolved limbs (P=.007) for the unilateral TKA group. Lower-limb force production of the involved limb in persons with unilateral TKA was significantly lower than the limb of the age- and gender-matched controls (P=.0001). CONCLUSIONS: Lower-limb force production on the involved side was significantly lower than the uninvolved side. When compared with an age-matched control group, subjects with unilateral TKA produced less force during a single leg horizontal press. PMID- 15295754 TI - Heart rate variability at rest and during exercise in persons with Down syndrome. AB - OBJECTIVES: To determine whether autonomic dysfunction explains chronotropic incompetence observed in persons with Down syndrome (DS) and to measure heart rate variability (HRV) at rest and during exercise in persons with mental retardation with and without DS. DESIGN: Comparative study. SETTING: University exercise science laboratory. PARTICIPANTS: Thirty-one subjects with mental retardation (age, 20.2 y) with DS (n=16; 10 men, 6 women) and without DS (n=15; 8 men, 7 women). INTERVENTIONS: Not applicable. Main outcome measures HRV was determined at rest and at 2 steady-state exercise intensities on the treadmill in both time (standard deviation of the R-R interval, percentage of R-R intervals deviating by more than 50 ms from the previous R-R interval [deviation >50], square root of the mean squared differences of successive differences) and frequency (low-frequency power [LF]), high-frequency power [HF], the LF/HF ratio) domains. RESULTS: The DS group demonstrated a statistically lower peak heart rate (161 beats/min vs 178 beats/min, P<.05), and peak oxygen consumption (27.4 mL.kg( 1).min(-1) vs 34.3 mL. kg(-1).min(-1), P<.05) than did the group with mental retardation without DS. At rest, all time domain measures of HRV and absolute HF power were significantly higher in the DS group (P<.05). Yet, LF power and LF/HF values did not differ between groups. All HRV variables decreased significantly at both exercise intensities, with no differences between groups during exercise (P<.05). CONCLUSIONS: People with DS have greater parasympathetic activity at rest, but group differences disappear with the onset of exercise, which suggests that other variables are responsible for chronotropic incompetence in persons with DS. PMID- 15295755 TI - Relationship between strength, balance, and swallowing deficits and outcome after traumatic brain injury: a multicenter analysis. AB - OBJECTIVE: To examine the relation among strength, balance, and swallowing deficits, as measured on rehabilitation admission, and functional outcome at discharge and 1 year after traumatic brain injury (TBI). DESIGN: Multicenter analysis of consecutive admissions to designated Traumatic Brain Injury Model Systems (TBIMS) facilities. SETTING: Seventeen TBIMS centers. PARTICIPANTS: Adults and children older than 16 years of age with TBI (N=2363) enrolled in the national database from January 1989 to November 2000. INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURES: Transfers, locomotion, stairs, lower-body dressing, grooming, bathing, upper-body dressing, toileting, and eating as measured by the FIM instrument at acute rehabilitation discharge and at 1 year after TBI. RESULTS: Lower-extremity strength less than 3/5 on admission to acute rehabilitation was associated with increased need for assistance in locomotion, transfers, and lower-body dressing and less than 3/5 upper-extremity strength was associated with the need for assistance in self-care at rehabilitation discharge and at 1 year postinjury. Similar relations were found between impaired swallowing and assistance with eating, grossly impaired dynamic sitting, or standing balance and assistance with locomotion, transfers, eating, and self-care at rehabilitation discharge and at 1 year after TBI. CONCLUSIONS: Assessments of physical strength, swallowing ability, and dynamic balance on acute rehabilitation admission are helpful as screening tests in predicting the need for assistance of another person for mobility and self-care at rehabilitation discharge. This association remains strong at 1 year after TBI. By using this information, clinicians should initiate therapeutic interventions that optimize rehabilitation of the identified impairments and should make necessary arrangement for the patient's anticipated postdischarge needs. Further studies are necessary to delineate the amount of unique variance that these early physical examination findings contribute to outcome prediction. PMID- 15295756 TI - Cognitive behavioral therapy for insomnia associated with traumatic brain injury: a single-case study. AB - OBJECTIVE: To test the efficacy of a cognitive behavioral therapy (CBT) for insomnia with a patient with traumatic brain injury (TBI). DESIGN: Single-case study. SETTING: Outpatient rehabilitation center. PARTICIPANT: A man in his late thirties who sustained a moderate TBI in a motor vehicle crash and who developed insomnia. He complained of difficulties falling asleep and staying asleep, despite pharmacotherapy with zopiclone. INTERVENTIONS: Eight weekly individual CBT sessions. Treatment included stimulus control, sleep restriction, cognitive therapy, and sleep hygiene education. MAIN OUTCOME MEASURES: Sleep diary and polysomnography data. RESULTS: Sleep onset decreased from 47 to 18 minutes, and nocturnal awakenings dropped from 85 to 28 minutes on average at posttreatment. Sleep efficiency also increased substantially (58% to 83%). Polysomnography evaluations corroborated the diary data by showing a decrease in total time awake (63.2 to 26.3 min) and in the number of awakenings (21 to 7.5). The majority of gains were well maintained at 1- and 3-month follow-up assessments. CONCLUSIONS: These preliminary results suggest that sleep disturbances after TBI can be alleviated with a nonpharmacologic intervention. CBT for post-TBI insomnia is a promising therapeutic avenue deserving more scientific and clinical attention. PMID- 15295757 TI - Decreased isometric neck strength in women with chronic neck pain and the repeatability of neck strength measurements. AB - OBJECTIVES: To evaluate neck flexion, extension, and, especially, rotation strength in women with chronic neck pain compared with healthy controls and to evaluate the repeatability of peak isometric neck strength measurements in patients with neck pain. DESIGN: Cross-sectional. SETTINGS: Rehabilitation center and physical and rehabilitation medicine department at a Finnish hospital. PARTICIPANTS: Twenty-one women with chronic neck pain and healthy controls matched for sex, age, anthropometric measures, and occupation. INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURES: Peak isometric strength of the cervical muscles was tested in rotation, flexion, and extension. RESULTS: Significantly lower flexion (29%), extension (29%), and rotation forces (23%) were produced by the chronic neck pain group compared with controls. When the repeated test results were compared pairwise against their mean, considerable variation was observed in the measures on the individual level. Intratester repeatability of the neck muscle strength measurements was good in all the 4 directions tested in the chronic neck pain group (intraclass correlation coefficient range,.74-.94). The coefficient of repeatability was 15N, both in flexion and extension, and 1.8 Nm in rotation. On the group level, improvement up to 10% due to repeated testing was observed. CONCLUSIONS: The group with neck pain had lower neck muscle strength in all the directions tested than the control group. This factor should be considered when planning rehabilitation programs. Strength tests may be useful in monitoring training progress in clinical settings, but training programs should be planned so that the improvement in results is well above biologic variation, measurement error, and learning effect because of repeated testing. PMID- 15295758 TI - Intratester and intertester reliability of neck isometric dynamometry. AB - OBJECTIVE: To evaluate the reproducibility of measurement for maximum voluntary isometric contractions of the cervical musculature in different movements. DESIGN: Repeated test-retest measurements. SETTING: A department of physiotherapy. PARTICIPANTS: Thirty-three healthy subjects (17 men, 16 women; age range, 19-63 y) for the intraexaminer study and 10 healthy subjects (4 men, 6 women; age range, 20-37 y) for the interexaminer study. INTERVENTIONS: Maximum isometric strength in sitting and standing for flexion, extension, lateral flexion, and rotation using a custom isomyometer device. Three tests, performed 5 to 8 days apart, to assess intraexaminer reliability. Two examiners, each performing 1 trial, measuring on the same day to assess interexaminer reliability. MAIN OUTCOME MEASURES: Intraexaminer and interexaminer reliability of neck muscle strength. RESULTS: The standing position showed better reproducibility than the sitting position. The intraclass correlation coefficient (ICC1,3) was above .84 for all tests in any movement and position and above .93 when the first test was excluded. The standard error (SE) of measurement (<16.5 N; <.13 N-m for rotation) and smallest detectable difference (SDD) (<20.1%) were also small. For interexaminer reliability, the ICC(2,1) ranged from.88 to.94 and the SE from 10.7 to 20.8 N (<1.15 N-m for rotation); the SDD was less than 29.8% (except right rotation, which was 38.8%). CONCLUSIONS: A reliable protocol for measuring neck strength has been developed. Standing position and a full practice session produces more reliable measurements. PMID- 15295759 TI - Measuring quality of life in multiple sclerosis patients with urinary disorders using the Qualiveen questionnaire. AB - OBJECTIVES: To assess the impact of urinary disorders on multiple sclerosis (MS) patients' health-related quality of life and to examine the cross-sectional construct validity of Qualiveen, a questionnaire originally developed for spinal cord injury patients with urinary disorders, in patients with MS. DESIGN: Cohort study. SETTING: Neurourodynamic units in 3 French university hospitals. PARTICIPANTS: Patients with MS (N=197). INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURES: We tested predictions about the relationships among clinical features, the French version of the Multiple Sclerosis Quality of Life questionnaire (SEP-59), the Expanded Disability Status Scale (EDSS), and the 4 domains of the 30-item Qualiveen. RESULTS: Cross-sectional correlations among the 4 Qualiveen domains and type (range, .36-.54), number of symptoms (range, .23 .50), and severity of incontinence (.39-.68) were generally moderate to strong. The SEP-59 bowel and bladder function domain showed moderate to strong relationships with the Qualiveen (range, .39-.59). Relationships with other SEP 59 domains were generally weak (range, .22-.35), and with the EDSS they were very weak. Predictions proved generally accurate (weighted kappa=.61). CONCLUSIONS: Our data supported the Qualiveen's validity as a discriminative instrument for use with patients with MS. Further studies should explore the Qualiveen's longitudinal validity and responsiveness. PMID- 15295760 TI - Quantifying environmental factors: a measure of physical, attitudinal, service, productivity, and policy barriers. AB - OBJECTIVE: To develop and test a new instrument to assess environmental barriers encountered by people with and without disabilities by using a questionnaire format. DESIGN: New instrument development. SETTING: A rehabilitation hospital and community. PARTICIPANTS: Two convenience samples: (1) 97 subjects, 50 with disabilities and 47 without disability, and (2) 409 subjects with disabilities from spinal cord injury, traumatic brain injury, multiple sclerosis, amputation, or auditory or visual impairments. In addition, a population-based sample in Colorado of 2269 people (mean age, 44 y; 57% men) with and without disabilities. INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURES: Item development; factor structure; test-retest, subject-proxy and internal consistency reliability; content, construct, and discriminant validity; and subscale and abbreviated version development. RESULTS: Panels of experts on disability developed items for the Craig Hospital Inventory of Environmental Factors (CHIEF). The instrument measured the frequency and magnitude of environmental barriers reported by individuals. Five subscales were derived from factor analysis measuring (1) attitudes and support, (2) services and assistance, (3) physical and structural, (4) policy, and (5) work and school environmental barriers. The CHIEF total score had high test-retest reliability (intraclass correlation coefficient [ICC]=.93) and high internal consistency (Cronbach alpha=.93), but lower participant-proxy agreement (ICC=.62). Significant differences were found in CHIEF scores among groups of people with known differences in disability levels and disability categories. CONCLUSIONS: The CHIEF has good test-retest and internal consistency reliability with evidence of content, construct, and discriminant validity resulting from its development strategy and psychometric assessments in samples of the general population and among people with a variety of disabilities. PMID- 15295761 TI - Positive straight-leg raising in lumbar radiculopathy: is documentation affected by insurance coverage? AB - OBJECTIVE: To evaluate whether differences exist in documentation of straight-leg raising (SLR), based on insurance coverage. DESIGN: Retrospective study. SETTING: Managed care organization (MCO). PARTICIPANTS: Two hundred people with a diagnosis of lumbar radiculopathy or herniated disk were referred to an MCO for authorization of further treatment. Half were self-directed under a personal injury program (PIP) after automobile collisions, and half were covered under a managed care workmen's compensation (WC) program. INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURES: Documentation of an SLR test, strength, sensation, and/or reflexes were eligible for the study. The results of SLR were coded as 0, 1, or 2, for absent, positive unilateral, and positive bilateral, respectively. Additional information included subject age, sex, date of injury, provider type, and presence of attorney representation RESULTS: A positive (unilateral, bilateral) SLR in women was 7.4 times more likely if they were covered by PIP than by WC (95% confidence interval [CI], 1.4-38.7; P=.018). For men, a positive SLR was 23.5 times more likely if they were covered by a PIP (95% CI, 2.9-189.9; P=.003). The odds of bilateral SLR (radicular pain on both sides) were even more strongly associated with type of reimbursement. For women, bilateral SLR was 105.1 times more likely if they were covered by a PIP than by WC (95% CI, 11.1 992.6; P<.001). For men, bilateral SLR was 38.9 times more likely if covered by a PIP (95% CI, 11.3-133.6; P<.001). CONCLUSIONS: Reasons for reporting higher rates of positive SLR in the PIP group include an added incentive to treat, poor knowledge of proper interpretation of the SLR test, and/or an increased exaggeration of symptoms. PMID- 15295762 TI - Houghton Scale of prosthetic use in people with lower-extremity amputations: Reliability, validity, and responsiveness to change. AB - OBJECTIVE: To evaluate the responsiveness to change and the floor and ceiling effects of the Houghton Scale. DESIGN: One-week and 3-month test-retest to evaluate reliability, validity, and responsiveness to change. SETTING: Amputee rehabilitation program. PARTICIPANTS: Persons (N=125) with unilateral or bilateral lower-extremity amputation who were wearing a prostheses: 1 group (n=49) for the reliability component and another group (n=76) for the responsiveness and validity component. INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURES: Responsiveness to change, ceiling and floor effects, and reliability and convergent validity. RESULTS: Evaluation of responsiveness to change (n=76) showed that the total score increased from a mean +/- standard deviation of 6.14+/-2.40 at discharge to 7.70+/-2.62 (P<.001) at follow-up 3 months later. Floor and ceiling effects were not detected for the overall score but were noted for the individual subscales. The internal consistency was moderate at discharge (Cronbach alpha=.71) and follow-up (Cronbach alpha=.70). The Houghton Scale correlated significantly, although moderately, with the physical composite score of the Medical Outcomes Study 36-Item Short-Form Health Survey (r=.393, P<.01) and the 2-minute walk test at admission (r=.620, P<.01) and discharge (r=.653, P<.01). The reliability (intraclass correlation coefficient=.96) of the Houghton Scale was high (n=49). CONCLUSIONS: The Houghton Scale is appropriately responsive to change in prosthetic use in individuals with lower-limb amputation after rehabilitation. PMID- 15295763 TI - The supine hip extensor manual muscle test: a reliability and validity study. AB - OBJECTIVES: To define the relative hip extensor muscle strengths values identified by the 4 grades obtained with a supine manual muscle test (MMT) and to compare these values with those indicated by the traditional prone test. DESIGN: Comparison of 4 manual supine strength grades with isometric hip extension joint torque; kappa statistic-determined interrater reliability, and analyses of variance identified between grade differences in torque. SETTING: Pathokinesiology laboratory. PARTICIPANTS: Adult volunteers recruited from local community and outpatient clinics. Reliability testing: 16 adults with postpolio (31 limbs). Validity testing (2 groups): 18 subjects without pathology (18 limbs), and 26 people with clinical signs of hip extensor weakness (51 limbs). INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURES: Supine hip extensor manual muscle grade and isometric hip extension torque. RESULTS: Reliability testing showed excellent agreement (82%). Subjects with pathology had significant differences in mean torque (P<.01) for the assigned grade 5 (176 Nm), grade 4 (103 Nm), grade 3 (67 Nm), and grade 2 (19 Nm). Healthy adults showed significant differences between grade 5 (212 Nm) and grade 4 (120 Nm) in mean torque (P<.05). CONCLUSIONS: The supine MMT is a reliable and valid method with which to assess hip extension strength. PMID- 15295764 TI - Ipsilateral motor pathway confirmed by combined brain mapping of a patient with hemiparetic stroke: a case report. AB - This study investigated the motor control pathway using both functional magnetic resonance imaging (fMRI) and transcranial magnetic stimulation (TMS) in a patient with left hemiparesis with an infarction on the posterior limb of the right internal capsule. fMRI was performed using the blood oxygen level-dependent technique at 1.5 T with a standard head coil. The motor activation task consisted of hand grasp-release movements in 1-Hz cycles. TMS was performed using a butterfly coil; the intersection of the wings (center of the coil) was applied tangentially to the scalp 1.0 cm apart. Stimulation was performed at 100% of maximal output. Motor evoked potentials (MEPs) from both abductor pollicis brevis (APB) muscles were obtained simultaneously. fMRI showed that the unaffected (left) primary sensorimotor cortex (SM1) was activated by movements of the unaffected (right) hand. Conversely, the bilateral SM1 were activated by movements of the affected (left) hand. Brain mapping using TMS showed that ipsilateral MEPs were obtained at the affected (left) APB muscle when the unaffected (left) motor cortex was stimulated. We concluded that the ipsilateral motor pathway from the unaffected motor cortex to the affected hand was present in this patient. PMID- 15295765 TI - Rehabilitation of a child with meningococcal septicemia and quadrilateral limb loss: a case report. AB - Acquired quadrilateral limb loss is a rare occurrence in children. One cause of this condition is severe meningococcal septicemia. We present the case of a boy who, at 14 months of age, required extensive amputation after an episode of meningococcal septicemia. We review his medical recovery and rehabilitation, including upper- and lower-limb prosthetic prescription and training, and adaptation to his altered body. A multidisciplinary approach led to effective management of his complex clinical and psychologic needs. This case illustrates the need to address a range of medical, prosthetic, and family issues central to successful clinical outcome. PMID- 15295766 TI - Postpartum sacral fracture presenting as lumbar radiculopathy: a case report. AB - Although rare, sacral stress fractures may occur in pregnant women, and osteoporosis of pregnancy is a poorly understood entity. We present the case of a young, postpartum, recreational runner who developed low back pain (LBP) and radicular symptoms suggestive of L5 radiculopathy found to be secondary to sacral stress fracture. The patient had a good clinical outcome after several months and was able to resume her normal activities. This case illustrates that clinicians should have a high index of suspicion for sacral stress fracture in athletic pregnant or postpartum women presenting with LBP and/or lumbar radiculopathy. Also included are a brief review of osteoporosis in pregnancy and guidelines on the diagnosis and management of sacral stress fractures. PMID- 15295767 TI - Factors affecting recovery from work-related, low back disorders in autoworkers. AB - OBJECTIVE: To simultaneously evaluate personal, medical, and job factors that could affect recovery from work-related, low back disorders, specifically focusing on an active working sample. DESIGN: Observational, longitudinal study. SETTING: Two US automotive plants. PARTICIPANTS: Employees (N=352; 289 men, 63 women; mean age +/- standard deviation, 45.1+/-7.5 y) who were active hourly autoworkers, diagnosed with work-related, low back disorder by the plant's medical department. INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURE: Oswestry Disability Questionnaire for back pain was used to evaluate recovery. RESULTS: Factors associated with better recovery were lower stress levels (P<.001) and exercise or physical activity outside work (P<.001); factors associated with higher disability levels over time were current cigarette smoking (P<.01) and bedrest (P<.001). CONCLUSIONS: Personal modifiable factors are major influences in the recovery from work-related, low back disorders, even in active working populations. Interventions aimed at increasing exercise and decreasing stress should also be considered as a part of rehabilitation in employed persons with low levels of disability. PMID- 15295768 TI - Impact of a viral respiratory epidemic on the practice of medicine and rehabilitation: severe acute respiratory syndrome. AB - Severe acute respiratory syndrome (SARS) is a new respiratory viral epidemic that originated in China but has affected many parts of the world, with devastating impact on economies and the practice of medicine and rehabilitation. A novel coronavirus has been implicated, with transmission through respiratory droplets. Rehabilitation was significantly affected by SARS, because strict infection control measures run counter to principles such as multidisciplinary interactions, patients encouraging and learning from each other, and close physical contact during therapy. Immunocompromised patients who may silently carry SARS are common in rehabilitation and include those with renal failure, diabetes, and cancer. Routine procedures such as management of feces and respiratory secretions (eg, airway suctioning, tracheotomy care) have been classified as high risk. Personal protection equipment presented not only a physical but also a psychologic barrier to therapeutic human contact. Visitor restriction to decrease chances of disease transmission are particularly difficult for long-staying rehabilitation patients. At the height of the epidemic, curtailment of patient movement stopped all transfers for rehabilitation, and physiatrists had to function as general internists. Our experiences strongly suggest that rehabilitation institutions should have emergency preparedness plans because such epidemics may recur, whether as a result of nature or of bioterrorism. PMID- 15295769 TI - The kappa statistic in rehabilitation research: an examination. AB - The number and sophistication of statistical procedures reported in medical rehabilitation research is increasing. Application of the principles and methods associated with evidence-based practice has contributed to the need for rehabilitation practitioners to understand quantitative methods in published articles. Outcomes measurement and determination of reliability are areas that have experienced rapid change during the past decade. In this study, distinctions between reliability and agreement are examined. Information is presented on analytical approaches for addressing reliability and agreement with the focus on the application of the kappa statistic. The following assumptions are discussed: (1) kappa should be used with data measured on a categorical scale, (2) the patients or objects categorized should be independent, and (3) the observers or raters must make their measurement decisions and judgments independently. Several issues related to using kappa in measurement studies are described, including use of weighted kappa, methods of reporting kappa, the effect of bias and prevalence on kappa, and sample size and power requirements for kappa. The kappa statistic is useful for assessing agreement among raters, and it is being used more frequently in rehabilitation research. Correct interpretation of the kappa statistic depends on meeting the required assumptions and accurate reporting. PMID- 15295770 TI - Reconsidering the motor recovery plateau in stroke rehabilitation. AB - Termination of motor rehabilitation is often recommended as patients with cerebrovascular accident (CVA) become more chronic and/or when they fail to respond positively to motor rehabilitation (commonly termed a "plateau"). Managed care programs frequently reinforce this practice by restricting care to patients responding to therapy and/or to the most acute patients. When neuromuscular adaptation occurs in exercise, rather than terminating the current regimen, a variety of techniques (eg, modifying intensity, attempting different modalities) are used to facilitate neuromuscular adaptations. After presenting the concepts of the motor recovery plateau and adaptation, we similarly posit that patients with CVA adapt to therapeutic exercise but that this is not indicative of a diminished capacity for motor improvement. Instead, like traditional exercise circumstances, adaptive states can be overcome by modifying regimen aspects (eg, intensity, introducing new exercises). Findings suggesting that patients with chronic CVA can benefit from motor rehabilitation programs that apply novel or different parameters and modalities. The objectives of this commentary are to (1) to encourage practitioners to reconsider the notion of the motor recovery plateau, (2) to reconsider chronic CVA patients' ability to recover motor function, and (3) to use different modalities when accommodation is exhibited. PMID- 15295771 TI - Pediatric rheumatology: what does the future hold? AB - Effectiveness of the traditional rehabilitation approaches used in pediatric rheumatology has been difficult to prove and, in times of cost containment, this lack of evidence may lead to undertreatment with physical and occupational therapies. Quantitative methods such as those described in this issue by Brostrom and colleagues can be used to validate those approaches and to reinforce the need for careful attention to the effects of even minor loss of range and strength in children with juvenile arthritis. Historically, up to half of the children affected by polyarticular juvenile arthritis became disabled. Some factors that have led to improved outcomes for childhood rheumatic diseases are discussed, including medications (use of weekly low-dose methotrexate, intra-articular steroid injections, new biologic agents that specifically block mediators of inflammation, for example, tumor necrosis factor and interleukin-1), surgery (joint replacements), and psychosocial interventions (with schools and families). The importance of maintaining range of movement, strength, weight bearing, and ambulation, in an effort to prevent sequelae such as osteoporosis and wheelchair dependence, is emphasized. Early identification of children with rheumatic diseases and aggressive intervention, with a combined medical, rehabilitation, psychosocial, and, rarely, surgical approach, should now allow most affected children to reach adulthood with little or no disability. PMID- 15295772 TI - Quality of life and psychological adaptation in siblings of paediatric cancer patients, 2 years after diagnosis. AB - Several studies have been conducted on sibling psychosocial adaptation to cancer in a brother or sister, but little is known on how the long-term adaptation of siblings to the illness develops. The concept quality of life has primarily been applied in research on the effects of chronic illness on the affected patient, but has not yet been studied in siblings. AIMS: To investigate the prevalence of self-reported psychosocial problems in siblings of pediatric cancer patients, 2 years after the onset of the illness. MEASUREMENTS: Two Dutch quality of life questionnaires, the TACQOL and the DUCATQOL are used, as well as generic non health-related measures of emotional and behaviour problems (CBCL-YSR) and anxiety (STAI-C). PARTICIPANTS: Participants were 103 siblings aged 7-18 years old. Fifty seven Siblings participated in a prospective and 46 in a retrospective study group. RESULTS: Siblings aged 7-11 report lower overall quality of life than children in the norm group. No differences in mean scores were found on any of the other domains that were investigated. When the prevalence of problems was investigated, however, relatively more siblings compared to normative data had scores in the impaired group based on the 20th percentile norm. A relatively high number of siblings aged 7-11 reported impaired emotional (42%), social (34%) and total quality of life (47%) (DUCATQOL) and physical problems (26%) (TACQOL). Relatively many adolescent siblings (26%) reported significant internalising problems on the CBCL-YSR. CONCLUSIONS: Although acute emotional distress reactions seem to have normalised in most siblings as has been suggested in the literature, emotional distress of having a brother or sister with cancer may continue beyond diagnosis for a subgroup of children. Young siblings seem to be affected in their quality of life, whereas a subgroup of adolescent siblings experience clinically relevant internalising problems. The results support the use of quality of life measures for siblings. Predictors of long-term adaptation in siblings need to be investigated. PMID- 15295773 TI - Interest in services among prostate cancer patients receiving androgen deprivation therapy. AB - Treatment side effects and decreased quality of life associated with androgen deprivation therapy (ADT) suggest the need for supportive services for prostate cancer (PC) patients receiving ADT. Nonetheless, uptake of services is low, suggesting that PC patients' preferences are not being addressed. We examined interest in supportive services and predictors of interest among 118 PC patients receiving ADT. Overall interest in services was associated with lower quality of life (p = 0.01). The majority of participants expressed interest in informational services (70%), with a minority (22%) expressing interest in psychosocial services. Interest in psychosocial services was associated with younger age (p = 0.02), and shorter duration of ADT (p < 0.04), but was unrelated to psychological distress or social support. Although most men (68%) reported that they would prefer not to take medication for depression, 75% would do so if advised by their physician. Overall, results suggest that PC patients on ADT prefer individualized informational support. Substantial interest (61%) in Oncolink, an internet-based informational resource, suggests the Internet may provide an acceptable mode of service delivery. Health care providers should consider integrating increased informational support into routine care and, more generally, consider patient preferences in prioritizing and designing support services. PMID- 15295774 TI - Longitudinal course of depression, fatigue, and quality of life in patients with high risk melanoma receiving adjuvant interferon. AB - PURPOSE: Treatment of malignant melanoma with interferon-alpha has been associated with a variety of side effects ranging from fatigue to depression, and a concomitant impact on quality of life (QOL), in a variety of case reports and cross-sectional clinical trials. Few, if any, studies have been conducted with the express purpose of assessing the longitudinal course of depression, fatigue, and QOL before and during interferon therapy. DESCRIPTION OF STUDY: The current study reports on 16 patients who were assessed at 6 points in time: baseline, post high dose, and 1, 2, 3, and 6 months post high dose treatment with interferon-alpha with the Brief Symptom Inventory, Beck Depression Inventory, Revised Piper Fatigue Scale, and Functional Assessment of Cancer Therapy Biological Response Modifiers. RESULTS: Results revealed consistent changes from baseline through 6 month assessment. Specifically, increased somatic complaints, depression, and fatigue were observed on the BSI, BDI, and RPFS, respectively. Additional reductions in QOL on the FACT-BRM were also identified. CLINICAL IMPLICATIONS: The findings suggest that IFN has a significant effect on QOL, but that it may be the somatic symptoms of fatigue that contribute to changes on measures of mood. Limiting the amount of fatigue and depression would appear to be significant if individuals are to successfully complete IFN therapy. PMID- 15295775 TI - Parent-reported environmental tobacco smoke exposure among preadolescents and adolescents treated for cancer. AB - Exposure to environmental tobacco smoke (ETS) poses serious health risks for children with cancer. Parental smoke is a primary source of exposure for these children. Parent smoking behaviors and parent-reported ETS exposure among children treated for cancer were examined in this study. In addition, reports of ETS exposure among children with cancer who currently smoked or who had smoked in the past were compared to those of children with cancer who never smoked. Written questionnaires about smoking behaviors and ETS exposure were administered to 47 smoking parents of youngsters diagnosed with cancer, 10-18 years of age (57.4% male, 78.7% Caucasian). Child reports of smoking status were also obtained. Results indicated that children with cancer are exposed to ETS from a number of sources and settings, as reported by their parents. Current or previous child smokers had greater ETS exposure than non-smoking children. Older children and Caucasian children also had greater ETS exposure. Level of ETS exposure did not differ based on the child's treatment status. Interventions that teach parents to protect their youngster from ETS exposure have potential for reducing adverse health outcomes in this vulnerable population. PMID- 15295776 TI - Repressive coping before and after diagnosis of breast cancer. AB - OBJECTIVE: The aim was to investigate to which extent emotional repression is a premorbid coping tendency of cancer patients and/or a coping response to the threat posed by a cancer diagnosis. The results of one previous study of breast cancer patients suggest the latter possibility, and our aim was to replicate and extend these findings. METHODS: Of 646 women referred to mammographic examination for breast cancer, 71 women were diagnosed with primary breast cancer. Repressive coping, defined as having high scores on defensiveness (Marlowe-Crowne Social Desirability Scale) and low scores on anxiety (Taylor Manifest Anxiety Scale), was measured (1) before, (2) 4 weeks after, and (3) 12 weeks after diagnosis. The women were not aware of their disease status before the examination, and there were no significant differences between groups in their perceived risk of having breast cancer. RESULTS: Four weeks after diagnosis, increased repression (p < 0.01) was found in the group of women diagnosed with breast cancer but not in women without cancer, with women with a breast cancer diagnosis being 1.5 times more likely to be repressive than women with cancer. There were no group differences in defensiveness, anxiety, or repression before diagnosis and 12 weeks after diagnosis. When controlling for repressive coping prior to diagnosis, age, and other demographic factors with a multiple, logistic regression, only cancer diagnosis (odds ratio: 2.39; p< 0.05) and having biological children (odds ratio: 2.83; p< 0.02) emerged as significant predictors of repressive coping 4 weeks after diagnosis. Before diagnosis, only higher age predicted later diagnosis of breast cancer. CONCLUSION: Previously found higher repression in cancer patients vs. controls could be a response to the threat associated with cancer diagnosis and may not necessarily reflect premorbid differences. PMID- 15295777 TI - The psychological impact of a cancer diagnosis on families: the influence of family functioning and patients' illness characteristics on depression and anxiety. AB - A diagnosis of cancer is a very stressful event for the patients and their families. Patients, partners and other family members can suffer from clinical levels of depression and severe levels of anxiety and stress reactions. The similarity in levels of distress between patients and partners and patients and offspring suggests that there are common factors that impact on families' distress levels. The current study examined levels of depression and anxiety in newly diagnosed adult patients (n = 48) and their adult relatives (n = 99). Family functioning and patients' illness characteristics were identified as factors that might impact on families' depression and anxiety. Results from multilevel models indicated that family functioning was important. Families that were able to act openly, express feelings directly, and solve problems effectively had lower levels of depression. Direct communication of information within the family was associated with lower levels of anxiety. Aside from differences anxiety due to cancer type, patients' illness characteristics appear to be risk factors in patients' but not relatives' depression and anxiety. The results from the current study suggest that researchers and clinicians need to be family-focused as cancer affects the whole family, not just the patient. PMID- 15295778 TI - Validation of a questionnaire for self-assessment of sexual function and vaginal changes after gynaecological cancer. AB - The Sexual function-Vaginal changes Questionnaire (SVQ), was developed to investigate sexual and vaginal problems in gynaecological cancer patients. The instrument consists of 20 core items, measuring sexual interest, lubrication, orgasm, dyspareunia, vaginal dimensions, intimacy, sexual problems of partner, sexual activity, sexual satisfaction, and body image. Seven additional items assessing current levels of sexual and vaginal problems compared to pre-diagnosis are intended to be used only once in longitudinal studies. The SVQ was validated in two ways: first, the comprehensibility of each item was investigated through combined quantitative and qualitative assessment of patient-observer agreement in 75 gynaecological cancer patients, second, multitrait analyses and principal component analyses were applied to responses from 257 patients with cervical cancer to investigate the scale properties. The level of agreement between the patients' and the observer's ratings was high (median overall agreement 0.84, range 0.46-1.00; median kappa: 0.80, range 0.52-1.00). From the 10 items applicable to all patients, three scales were hypothesized: intimacy, sexual interest and global sexual satisfaction. For sexually active respondents an additional two scales were hypothesized: vaginal changes and sexual functioning. The psychometric analyses confirmed these scales. The internal consistency of the scales ranged 0.76-0.83 (Cronbach's alpha). The study supports the validity and reliability of the SVQ. PMID- 15295779 TI - Effect of lipid bilayer alteration on transdermal delivery of a high-molecular weight and lipophilic drug: studies with paclitaxel. AB - Skin forms an excellent barrier against drug permeation, due to the rigid lamellar structure of the stratum corneum (SC) lipids. Poor permeability of drugs can be enhanced through alteration in partition and diffusion coefficients, or concentration gradient of drug with an appropriate choice of solvent system, along with penetration enhancers. The aim of the current investigation was to assess applicability of lipid bilayer alteration by fatty acids and terpenes toward the permeation enhancement of a high-molecular-weight, lipophilic drug, paclitaxel (PCL) through rat skin. From among the fatty acids studied using ethanol/isopropyl myristate (1:1) vehicle, no significant enhancement in flux of PCL was observed (p > 0.05). In the case of cis mono and polyunsaturated fatty acids lag time was found to be similar to control (p > 0.05). This suggests that the permeation of a high-molecular-weight, lipophilic drug may not be enhanced by the alteration of the lipid bilayer, or the main barrier to permeation could lie in lower hydrophilic layers of skin. A significant increase in lag time was observed with trans unsaturated fatty acids unlike the cis isomers, and this was explained on the basis of conformation and preferential partitioning of fatty acids into skin. From among the terpenes, flux of PCL with cineole was significantly different from other studied terpenes and controls, and after treatment with menthol and menthone permeability was found to be reduced. Menthol and menthone cause loosening of the SC lipid bilayer due to breaking of hydrogen bonding between ceramides, resulting in penetration of water into the lipids of the SC lipid bilayer that leads to creation of new aqueous channels and is responsible for increased hydrophilicity of SC. This increased hydrophilicity of the SC bilayer might have resulted in unfavorable conditions for ethanol/isopropyl myristate (1:1) along with PCL to penetrate into skin, therefore permeability was reduced. The findings of this study suggest that the permeation of a high-molecular-weight and lipophilic drug cannot be enhanced through bilayer alteration by penetration enhancers, and alteration in partitioning of drug into skin could be a feasible mode to enhance the permeation of drug. PMID- 15295780 TI - Pharmacokinetic aspects of biotechnology products. AB - In recent years, biotechnologically derived peptide and protein-based drugs have developed into mainstream therapeutic agents. Peptide and protein drugs now constitute a substantial portion of the compounds under preclinical and clinical development in the global pharmaceutical industry. Pharmacokinetic and exposure/response evaluations for peptide and protein therapeutics are frequently complicated by their similarity to endogenous peptides and proteins as well as protein nutrients. The first challenge frequently comes from a lack of sophistication in various analytical techniques for the quantification of peptide and protein drugs in biological matrices. However, advancements in bioassays and immunoassays--along with a newer generation of mass spectrometry-based techniques -can often provide capabilities for both efficient and reliable detection. Selection of the most appropriate route of administration for biotech drugs requires comprehensive knowledge of their absorption characteristics beyond physicochemical properties, including chemical and metabolic stability at the absorption site, immunoreactivity, passage through biomembranes, and active uptake and exsorption processes. Various distribution properties dictate whether peptide and protein therapeutics can reach optimum target site exposure to exert the intended pharmacological response. This poses a potential problem, especially for large protein drugs, with their typically limited distribution space. Binding phenomena and receptor-mediated cellular uptake may further complicate this issue. Elimination processes--a critical determinant for the drug's systemic exposure--may follow a combination of numerous pathways, including renal and hepatic metabolism routes as well as generalized proteolysis and receptor mediated endocytosis. Pharmacokinetic/pharmacodynamic (PK/PD) correlations for peptide and protein-based drugs are frequently convoluted by their close interaction with endogenous substances and physiologic regulatory feedback mechanisms. Extensive use of pharmacokinetic and exposure/response concepts in all phases of drug development has in the past been identified as a crucial factor for the success of a scientifically driven, evidence-based, and thus accelerated drug development process. Thus, PK/PD concepts are likely to continue and expand their role as a fundamental factor in the successful development of biotechnologically derived drug products in the future. PMID- 15295781 TI - Correlation of tetradecylmaltoside induced increases in nasal peptide drug delivery with morphological changes in nasal epithelial cells. AB - The effect of tetradecylmaltoside (TDM) on nasal peptide drug absorption was assessed with four peptides of distinct molecular size: insulin (5.7 kDa), leptin (16 kDa), somatropin (22.1 kDa), and epoetin alfa (30.4 kDa). The nasal uptake of the smallest peptides, insulin and leptin, was significantly increased at a TDM concentration of only 0.06%. The uptake of somatropin was significantly increased when concentrations of 0.125% or more were used. The uptake of the largest peptide, epoetin alfa, was not significantly increased, in the presence of 0.125 0.5% TDM. Light microscopy revealed that formulations containing 0.125% TDM caused moderate alterations in nasal epithelial cell morphology, while higher concentrations of TDM (0.5%), caused more extensive morphological changes. Following treatment with 0.125% TDM, the distribution of cilia was altered and the number of pinocytotic vesicles was increased, at a time that correlated with increased nasal absorption of insulin. Consistent with these findings, FITC insulin applied nasally in the absence of TDM did not enter nasal epithelial cells, whereas FITC-insulin co-administered with 0.125% TDM was internalized into the cells, with a uniform distribution, consistent with transcellular movement of the peptide through the cells. PMID- 15295783 TI - Hydration, stability, and phase transformations of a new antitumor drug. AB - We studied hydration equilibria and phase transformations in a cytotoxic drug (BBR3576). The original sample is a hydrated compound that undergoes a structural phase transition when it looses about half of its structural water. Such a structural transition is completely reversible: the partially dehydrated form is stable up to 130 degrees C (or up to 140 degrees C for several minutes) and reverts to the original form upon rehydration. Completely different phase relationships are observed if an original sample is fully dehydrated: when all water has been released, a metastable anhydrous phase forms, which undergoes an irreversible exothermic conversion to a stable phase. Upon rehydration at room temperature of such an anhydrous phase, a new hydrated form is obtained, which is definitely different from the original one. PMID- 15295782 TI - Pre-clinical and clinical evaluation of solution and soft gelatin capsule formulations for a BCS class 3 compound with atypical physicochemical properties. AB - R1481 is a sub-type selective muscarinic receptor antagonist with the potential treatment of overactive bladder. R1481 presents two challenges for drug development. The first is the viscous semi-solid nature of the active pharmaceutical ingredient (API). The second challenge is the poor oral bioavailability of this water soluble, metabolically stable compound due to low intestinal permeability, and the P-glycoprotein (P-gp) efflux mechanism. Vitamin E TPGS is reported by others to enhance bioavailability by increasing the solubility of active compounds and by inhibiting P-gp in the intestine. In this report, compatibility of R1481 in Capmul MCM-based formulations with and without vitamin E TPGS is summarized. Review of accelerated stability studies of oral formulations led to the identification of a soft gelatin capsule formulation using neat Capmul MCM as an acceptable formulation for Phase 1 clinical studies. Soft gelatin capsules (5 mg strength) were manufactured with and without the addition of vitamin E TPGS. Clinical data show that vitamin E TPGS does not improve systemic exposure of R1481 in humans. PMID- 15295784 TI - Comparison of torque measurements and near-infrared spectroscopy in characterization of a wet granulation process. AB - The purpose of this study was to compare impeller torque measurements and near infrared (NIR) spectroscopy in the characterization of the water addition phase of a wet granulation process. Additionally, the effect of hydrate formation during granulation on the impeller torque was investigated. Anhydrous theophylline, alpha-lactose monohydrate, and microcrystalline cellulose (MCC) were used as materials for the study. The materials and mixtures of them were granulated using purified water in a small-scale high-shear mixer. The impeller torque was registered and NIR spectra of wet samples were recorded at-line. The torque and the NIR baseline-corrected water absorbances increased with increasing water content. A plateau in the NIR baseline-corrected water absorbances was observed for wet masses containing MCC. This was at the region of optimal water amount for granulation according to the torque results. In the case of anhydrous theophylline, the slope of baseline-corrected water absorbance values increased at the same water amount as the impeller torque started to increase. The hydrate formation of theophylline during granulation was observed as a slight decrease in the impeller torque. In addition, the hydrate formation during granulation affected the granulation liquid requirement. The liquid requirement was different for monohydrate formed during granulation compared to one formed in high relative humidity before the granulation. The results suggest that NIR spectroscopy may be applicable to process monitoring of wet granulation, also in cases where monitoring of impeller torque is difficult to apply. PMID- 15295785 TI - Lyophilization of polyethylene glycol mixtures. AB - Lyophilization of cosolvent systems may be a beneficial way of enhancing both physical and chemical stability of a drug product. The objective of this research is to establish whether cosolvent systems commonly used in the formulation of poorly water-soluble drugs can be successfully lyophilized. Polyethylene glycol (PEG) 400 was selected because it is widely used and can be easily frozen. The addition of PEG 400 to commonly used bulking agents, such as mannitol, sucrose, or polyvinylpyrrolidone, caused a significant change in the thermal properties of the bulking agents as observed by modulated differential scanning calorimetry. In addition, PEG 8000 was evaluated as a bulking agent because it also can function as a cosolvent in solution and forms an acceptable cake after lyophilization. Addition of PEG 400 to PEG 8000 caused negligible changes in the thermogram of this bulking agent. Surprisingly, the combination of PEG 8000 and PEG 400 forms a solid lyophilized cake. The current system can be best described as the lyophilization of a miscible solution of PEG 8000 and PEG 400 resulting in a lyophile that has a crystalline structure of PEG 8000 which is able to support PEG 400. PMID- 15295786 TI - Characterization of pharmaceutical solids by scanning probe microscopy. AB - The force-displacement profiles of four well-characterized materials that represent both soft/hard and plastic/brittle materials have been obtained using a novel nanoindentation technique. Flat surfaces of acetaminophen, potassium chloride, sucrose, and sodium stearate were prepared by melting or recrystallization, and the melting points were measured. Topographic and the corresponding first derivative images were obtained both before and after indentation. The materials were indented using a 10 s loading time, followed by a 2 s hold, and ending with a 10 s unloading time thereby providing a unique force displacement profile for each material. The loading profile of acetaminophen was discontinuous, whereas the profiles for the other three materials were smooth. The profiles were analyzed and the rank order of hardness was sucrose > acetaminophen > KCl > sodium stearate, which is consistent with the literature. The rank order of the stiffness, which is related to the modulus of elasticity, was sucrose > KCl > acetaminophen > sodium stearate. Given the flexibility and power of this approach, nanoindentation coupled with atomic force microscopy may be a useful means to characterize materials for evaluating tablet-processing conditions, perhaps at a preformulation stage. PMID- 15295787 TI - The stability of lyophilized lipid/DNA complexes during prolonged storage. AB - It is well known that excipients are required to protect nonviral vectors during the lyophilization process. The goal of this study is to describe the stability of lyophilized nonviral vector preparations on pharmaceutically relevant timescales and provide insight into the factors that govern long-term stability of vectors in the dried state. Lipid/DNA complexes were lyophilized in glucose, sucrose, or trehalose and stored for a period of up to 2 years at five different temperatures (-20, 4, 22, 40, 60 degrees C). We evaluated simultaneously the physico-chemical characteristics (size, zeta potential, ethidium bromide (EtBr) accessibility, supercoiled DNA content) and the ability of vector formulations to transfect COS-7 cells at different time intervals. In addition, a fluorescence assay was utilized to assess levels of ROS in the dried cake after storage. The physical state of each formulation was evaluated by determination of the glass transition temperature and residual moisture content, before and after storage. Results from our stability study show that a progressive degradation of lipid/DNA complexes occurs in terms of transfection rates, particle size, dye accessibility, and supercoil content, even when samples are stored at low temperatures (e.g., -20 degrees C). Furthermore, our preliminary results on the quantification of free radicals in rehydrated formulations emphasize the importance of developing strategies to prevent the formation of reactive oxygen species (ROS) during prolonged storage in the dried state. PMID- 15295788 TI - GARField magnetic resonance profiling of the ingress of model skin-care product ingredients into human skin in vitro. AB - A preliminary study of the ingress of mineral oil, decanol, and glycerine into samples of human abdominal skin tissue in vitro made using magnetic resonance profiling with a GARField magnet is reported. Two layers, each circa 50 microm thick and attributed to stratum corneum and viable epidermis, are spatially resolved. Clear differences are observed in the magnetic resonance response of these layers arising from the application of the model skin-care product ingredients. In the case of decanol and glycerine, it is suggested that the profiles show evidence for the effects of moisturization, as distinct from hydration. In the case of glycerine, the effective ingress diffusion coefficient is calculated to be 1.3 +/- 0.5 x 10(-9) cm2s(-1). PMID- 15295789 TI - Microbore HPLC method with online microdialysis for measurement of topotecan lactone and carboxylate in murine CSF. AB - We developed a chromatography method to measure lactone and carboxylate forms of topotecan (TPT) in mouse cerebrospinal fluid (CSF) using microdialysis sampling. The chromatography method utilized a microbore (0.8 mm) column. Analytes, which eluted in less than 5 min, were detected with a fluorescence detector. The calibration range was 0.25-100.0 ng/mL for both forms. The within-day and between day precision was < or =16% for 0.8 ng/mL and < or =8.0% for 3, 12, and 80 ng/mL. Accuracy was +/-15% (0.8 and 3 ng/mL) and +/-10% (12 and 80 ng/mL). TPT lactone hydrolyzes to the carboxylate during sampling, so we developed an equation and parameters to describe the TPT lactone hydrolysis in artificial CSF (aCSF). After TPT administration, CSF dialysate samples (2 microL) were analyzed for lactone and carboxylate using online injection. The hydrolysis of each dialysate sample was then estimated and a correction applied. We conclude that this HPLC method coupled with online microdialysis sampling allows for the rapid measurement of both TPT forms in small volumes of murine CSF dialysate. The system allows for the determination of TPT pharmacokinetics in murine CSF and provides a tool to extend pharmacological studies in this brain compartment. PMID- 15295790 TI - Pharmaceutical impurity identification: a case study using a multidisciplinary approach. AB - A multidisciplinary team approach to identify pharmaceutical impurities is presented in this article. It includes a representative example of the methodology. The first step is to analyze the sample by LC-MS. If the structure of the unknown impurity cannot be conclusively determined by LC-MS, LC-NMR is employed. If the sample is unsuitable for LC-NMR, the impurity needs to be isolated for conventional NMR characterization. Although the technique of choice for isolation is preparative HPLC, enrichment is often necessary to improve preparative efficiency. One such technique is solid-phase extraction. For complete verification, synthesis may be necessary to compare spectroscopic characteristics to those observed in the original sample. Although not widely practiced, an effective means of getting valuable structural information is to conduct a degradation study on the purified impurity itself. This systematic strategy was successfully applied to the identification of an impurity in the active pharmaceutical ingredient 1-(1,2,3,5,6,7-hexahydro-s-indacen-4-yl)-3-[4-(1 hydroxy-1-methyl-ethyl)-furan-2-sulphonylurea. Identification required the use of all of the previously mentioned techniques. The instability of the impurity under acidic chromatographic conditions presented an additional challenge to purification and identification. However, we turned this acidic instability to an advantage, conducting a degradation study of the impurity, which provided extensive and useful information about its structure. The following discussion describes how the information gained from each analytical technique was brought together in a complementary fashion to elucidate a final structure. PMID- 15295791 TI - Intrinsic adhesion force of lubricants to steel surface. AB - The intrinsic adhesion forces of lubricants and other pharmaceutical materials to a steel surface were quantitatively compared using Atomic Force Microscopy (AFM). A steel sphere was attached to the tip of an AFM cantilever, and its adhesion forces to the substrate surfaces of magnesium stearate, sodium stearyl fumarate, lactose, 4-acetamidophenol, and naproxen were measured. Surface roughness varied by an order of magnitude among the materials. However, the results clearly showed that the two lubricants had about half the intrinsic adhesion force as lactose, 4 acetamidophenol, and naproxen. Differences in the intrinsic adhesion forces of the two lubricants were insignificant. The lubricant molecules were unable to cover the steel surface during AFM measurements. Intrinsic adhesion force can slightly be modified by surface treatment and compaction, and its tip-to-tip variation was not greater than its difference between lubricants and other pharmaceutical particles. This study provides a quantitative fundamental basis for understanding adhesion related issues. PMID- 15295792 TI - Influence of methacrylic and acrylic acid polymers on the release performance of weakly basic drugs from sustained release hydrophilic matrices. AB - Weakly basic drugs and their salts exhibit a drop in aqueous solubility at high pH conditions, which can result in low and incomplete release of these drugs from sustained release formulations. The objective of this study is to modulate matrix microenvironmental pH by incorporation of acidic polymers and thus enhance the local solubility and release of basic drugs in high pH environment. Two weakly basic drugs, papaverine hydrochloride and verapamil hydrochloride with widely different pKa and aqueous solubilities at the pH of interest (6.8), were investigated for their release from hydrophilic matrices and the effect of a methacrylic (Eudragit L100-55) and an acrylic acid polymer (Carbopol 71G), were studied. For papaverine HCl, release increased with an increase in the levels of the acidic polymer used. Direct measurement of matrix pH using microelectrodes illustrated that the mechanism of release enhancement was based on modulation of microenvironmental pH. For verapamil HCl, incorporation of L100-55 resulted in release retardation due to an interaction between the anionic polymer and the cationic drug and the extent of retardation increased with an increase in the polymer level. The interaction product was characterized by NIR, FT-IR, and MTDSC techniques. Verapamil HCl release from Carbopol 71G based matrix tablets was higher than that from conventional hydroxypropyl methylcellulose (HPMC) based matrices, without any incorporated acidic additives. PMID- 15295793 TI - Protein-solute interactions affect the outcome of ultrafiltration/diafiltration operations. AB - Protein production operations often involve a final diafiltration of the protein into formulation buffer. For several Amgen product proteins, post-diafiltration assays revealed a significant difference in molar excipient concentrations on the retentate and the permeate side of the membrane. For example, post-diafiltration assays of formulated 200 mg/mL human interleukin-1 receptor antagonist showed molar chloride concentrations up to 30% lower than those of the diafiltration buffer. Deviations from expected results were also observed in cases where a fusion conjugate protein (AMG-719) was formulated by dialysis in 10 mM acetate and where PEGylated soluble tumor necrosis factor receptor (PEG-sTNF-RI) was formulated in 270 mM glycine and 10 mM histidine. Classical thermodynamic theory describing intermolecular interactions predicts that the partitioning of small solutes during dialysis will be dependent on the protein concentration, charge, and surface area. This study illustrates methods to approximate these effects using readily available protein data (theoretical titration curves based on protein sequence, density information, etc.). Additionally, guidelines are provided to determine when intermolecular interactions are likely to significantly impact the outcome of dialysis/diafiltration operations. PMID- 15295794 TI - Physicochemical, tissue distribution, and vasodilation characteristics of nitrosated serum albumin: delivery of nitric oxide in vivo. AB - Conjugates of nitric oxide (NO) to serum albumins are candidates for controlled delivery of NO in vivo, but their physicochemical and tissue distribution characteristics have hardly been examined yet. In this study, to achieve its in vivo delivery, bovine serum albumin (BSA) was reacted with sodium nitrite to obtain NO-BSA, which had 0.25-0.28 molecules of S-nitrosothiol/BSA. In addition to cystein, other amino acid residues were modified by the reaction. The conjugation had no significant effect on the molecular weight, but reduced the electric charge and induced reversible changes in the secondary structure of BSA. After intravenous injection in mice at a dose of 1 mg/kg, 111In-NO-BSA slowly disappeared from plasma in a similar manner to 111In-BSA, but showed greater accumulation in the liver and kidney. NO-BSA induced a transient decrease in arterial pressure after intravenous injection in rats at a dose of 100 mg/kg, and significantly increased the distribution of 111In-BSA to the lung in mice. These results indicate that NO is released from NO-BSA shortly after injection, and this NO decreases blood pressure and increases the distribution of macromolecules to the lung. These findings provide useful basic information for designing macromolecular NO donors able to achieve controlled delivery of NO. PMID- 15295795 TI - Effects of acute renal failure on the pharmacokinetics of oltipraz in rats. AB - Pharmacokinetic parameters of oltipraz were compared after intravenous and oral administration at a dose of 30 mg/kg to control rats and rats with U-ARF. After intravenous administration to rats with U-ARF, the AUC was significantly greater (1100 versus 1730 microg x min/mL) than that in control rats, and this could be due to significantly slower CL (27.2 versus 17.3 mL/min/kg). The slower CL could be mainly due to significantly slower CL(NR) (27.2 versus 17.3 mL/min/kg), and this could be supported by significantly slower in vitro CL(int) (32.1 versus 13.2 mL/min/whole liver) in the rats. The Vss was significantly larger in rats with U-ARF (4050 versus 5680 mL/kg), and this was not due to a significant increase in free fractions (unbound in plasma proteins) of oltipraz in the rats because the free fractions were 17.0 and 15.7% for control rats and rats with U ARF, respectively. Unexpectedly, after oral administration to rats with U-ARF, the AUC of oltipraz was significantly smaller than that in control rats (329 versus 149 microg x min/mL), and this could be mainly due to a decrease in the absorption of oltipraz from the gastrointestinal tract in the rats (95 and 72% of the oral dose were absorbed in control rats and rats with U-ARF, respectively). PMID- 15295796 TI - Effects of bacterial lipopolysaccharide on the pharmacokinetics of DA-7867, a new oxazolidinone, in rats. AB - Pharmacokinetic parameters of DA-7867 were compared after intravenous and oral administration at a dose of 10 mg/kg to control rats and rats pretreated with Klebsiella pneumoniae lipopolysaccharide (KPLPS). After intravenous administration of DA-7867 at a dose of 10 mg/kg to 10 rats, metabolism of DA-7867 was minimal; however, the urinary and gastrointestinal excretion of DA-7867 were approximately 85% of intravenous dose when collected for up to 14 days. After intravenous administration to rats pretreated with KPLPS, the AUC was significantly greater (14,100 versus 9810 microg x min/mL), and this could be due to significantly slower total body clearance (CL, 0.709 versus 1.02 mL/min/kg). The slower CL in the rats could be due to significantly smaller fecal excretion of DA-7867 for up to 14 days (41.1 versus 58.8% of intravenous dose of DA-7867) because urinary excretion of DA-7867 was not significantly different between two groups of rats. After oral administration, the AUC values were not significantly different between two groups of rats and this was mainly due to decrease in absorption from the gastrointestinal tract in rats pretreated with the KPLPS (approximately 82 and 95% of oral dose were absorbed for rats with KPLPS and control rats, respectively). PMID- 15295797 TI - Pharmacokinetic changes of DA-8159, a new erectogenic, after intravenous and oral administration to rats with diabetes mellitus induced by streptozotocin. AB - Intravenous administration of DA-8159, 30 mg/kg, to rats with diabetes mellitus induced by streptozotocin (DMIS), AUC of DA-8164 (a metabolite) was significantly smaller in rats with DMIS (57.9 compared with 81.8 microg x min/mL). This may be due to more contribution of significantly faster clearance of DA-8164 than that of significantly greater formation of DA-8164 in the rats. For example, the CL of DA-8164 was significantly faster (9.68 compared with 6.29 mL/min/kg) after intravenous administration of DA-8164, 10 mg/kg, to rats with DMIS and in vitro intrinsic clearance for the formation of DA-8164 was significantly faster (1.92 compared with 1.59 microL/min/mg protein) in hepatic microsomal fraction of rats with DMIS due to significant increase in expression of CYP3A1(23) in the rats. DA 8164 was formed mainly via CYP3A1/2 in rats. After intravenous administration of DA-8159, renal clearance was significantly faster in rats with DMIS (5.79 compared with 2.80 mL/min/kg) due to urine flow-dependent renal clearance of DA 8159 in rats. After oral administration of DA-8159, the AUC values of both DA 8159 and DA-8164 were not significantly different between two groups of rats. Although the exact reason is not known it may be due to changes in first-pass effect of DA-8159 in rats with DMIS. PMID- 15295798 TI - Effects of cysteine on the pharmacokinetics of intravenous torasemide in rats with protein-calorie malnutrition. AB - Effects of cysteine on the pharmacokinetics of torasemide were investigated after intravenous administration at a dose of 2 mg/kg to control rats and rats with PCM and PCMC. Torasemide was reported to be mainly metabolized via hepatic CYP2C9 in humans, and human CYP2C9 and male rat CYP2C11 proteins have 77% homology. It has also been reported that in male rats with PCM, the CYP2C11 level decreased to approximately 20% of the control level, but the decreased CYP2C11 level in rats with PCM partially returned to the control level by oral cysteine supplementation (rats with PCMC). Hence, it could be expected that in rats with PCM, some pharmacokinetic parameters of torasemide could be significantly different compared with those in control rats and rats with PCMC; however, they could be not significantly different between control rats and rats with PCMC. This was proven by the following parameters; the AUC (1880, 4080, and 2290 microg x min/mL for control rats and rats with PCM and PCMC, respectively), terminal half-life (188, 277, and 139 min), MRT (154, 323, and 155 min), CL (1.06, 0.491, and 0.943 mL/min/kg), CL(NR) (0.992, 0.430, and 0.874 mL/min/kg), and in vitro intrinsic torasemide disappearance clearance, CL(int) (0.102, 0.0842, and 0.0997 mL/min/mg protein). PMID- 15295799 TI - Ionic interaction of amiloride and uridine 5'-triphosphate in nebulizer solutions. AB - Combination therapy using nebulized amiloride hydrochloride and uridine-5' triphosphate (UTP) trisodium salt aerosols has been investigated for the treatment of cystic fibrosis (CF). Amiloride in aqueous solution precipitates in the presence of UTP, reducing drug concentrations. Interactions between these drugs and NaCl in solution were studied using phase-solubility techniques monitored by UV spectrophotometry. Elemental analyses were employed for precipitate characterization. Amiloride solubility was reduced by more than 85% in saline. Amiloride solubility decreased with increasing UTP concentration, resulting in formation of a precipitated complex. The theoretical molar ratio of complexes range from 1-3 amiloride:1 UTP. At most concentrations only 3 amiloride:1 UTP complex was observed in precipitate. This is a reflection of low Ksp for the 3:1 complex of 2.92 x 10(-11) M4 compared with 2.09 x 10(-4) M2 for amiloride alone. Equilibration over excess bulk solid resulted in higher solubility estimates and different phase solubility diagrams than solubility studies utilizing precipitation technique. This may be explained by the absence of amiloride in the solid state and its impact on complex equilibria with UTP. The solubility suppressing effects of UTP and saline were largely additive. A number of ionic interactions increase complex solubility profile of amiloride hydrochloride in the presence of UTP and NaCl. PMID- 15295801 TI - Festschrift dedicated to Prof. Manfred Zimmermann on the occasion of his 70th birthday. PMID- 15295800 TI - Dietary polyphenols (-)-epicatechin and chrysin inhibit intestinal glucuronidation metabolism to increase drug absorption. AB - The effect of dietary polyphenols on the intestinal glucuronidation and absorption of a model phenolic drug, alpha-naphthol (alpha-NA), was studied in isolated rat small intestine. (-)-Epicatechin significantly inhibited the glucuronidation of alpha-NA. Chrysin, (-)-epigallocatechin galleate (EGCG), and quercetin decreased the rate of glucuronidation, although not significantly. Baicalin did not affect the glucuronidation. The rate of absorption of alpha-NA in the presence of these polyphenols also varied. The absorption clearance (CLabs) and the metabolic clearance (CLmet) were inversely correlated, and this relationship was well explained in the metabolic inhibition model with kinetic parameters (knowledge-based prediction) which characterizes the relationship between the CLabs and CLmet of alpha-NA (Biochim Biophys Acta 1998, 1425, 398.). These results indicate that the concomitant intake of some polyphenols can increase the absorption of a phenolic drug, and the effect is predictable. (-) Epicatechin and chrysin are effective for the inhibition of glucuronidation and promotion of intestinal drug absorption. PMID- 15295802 TI - Diabetes care. Defining standards part I. PMID- 15295803 TI - Use your beans. The wonders of fiber. PMID- 15295804 TI - Smart snacking. A new look at a favorite pastime. PMID- 15295805 TI - Medicare and you. New prescription drug discounts. PMID- 15295806 TI - Della Reese: "Diabetes will not make me a victim." Della Reese has type 2 diabetes, but you'll never hear her complain about her life. PMID- 15295807 TI - Tired of beef? Try lamb chops. For an easy, memorable dinner, choose lamb chops. PMID- 15295808 TI - Tips for site rotation. When it comes to insulin, where you inject is just as important as how much and when. PMID- 15295809 TI - Beverages count. It's easy to think that we don't have to count anything we drink in our meal plan. Not true. PMID- 15295810 TI - Diabetes: a family affair. Tom and Evelyn Parks want their children to know that there is diabetes in the family. PMID- 15295812 TI - Abstracts from the National Research Forum for Young Health Investigators in Circulatory and Respiratory Health. May 6-8, 2004. Winnipeg, Manitoba, Canada. PMID- 15295811 TI - Research profile. Chromium. Can it help kids with type 2? Sushil K. Jain, PhD. PMID- 15295813 TI - Sepsis, kidney and multiple organ dysfunction. Proceedings of the Third International Course on Critical Care Nephrology. June 1-4, 2004. Vicenza, Italy. PMID- 15295814 TI - ICD's in congestive heart failure-is there bang for the beat? PMID- 15295815 TI - Work stress and hypertension: a call from research into intervention. PMID- 15295816 TI - [The development of psychophysiological measurement,1850-1865]. AB - Toward the end of the 1840s, Hermann von Helmholtz (1821-1894) began to investigate experimentally the propagation of stimuli within nerves. Helmholtz's experiments on animals and human subjects opened a research field that in the following decades was intensively explored by neurophysiologists and experimental psychologists. Studying the concrete experiemental settings and their local contexts shows how deeply the work of Helmholtz, Adolphe hirsch (1830-1901), Franciscus Donders (1818-1889) and others was embedded in the history of culture and technology. In particular, the rapidly growing technologies of electromagnetism, which gave rise to telegraphy and electric clocks, facilitated the time measurements of 19th-century physiologists and psychologists. However, the transition from frogs to human beings as model organisms confronted the time measuring psychophysiologists with a whole range of experimental parameters that were difficult to control (temperature, attention etc.). It is no wonder then that it took some 20 years before this branch of research stabilised. PMID- 15295818 TI - Abstracts of the 10th International Congress of Toxicology. 11-15 July 2004, Tampere, Finland. PMID- 15295817 TI - Portrait of an instrument-maker:Wenceslaus Hollar's engraving of Elias Allen. AB - Among the many engravings of landscapes, buildings, portraits and other illustrations produced by the seventeenth-century artist Wenceslaus Hollar, there are a small number of images of contemporary men of science. Of particular interest is the portrait of the instrument-maker Elias Allen, both because portraits of men of his social status were extremely uncommon at this time, and also because the cluttered mass of instruments shown in the image presents a picture wholly unlike other portraits of the period. The first part of this paper explores the position of portraiture as inherently linked to nobility, and seeks to present an explanation as to why the original oil painting of Allen (made by Hendrik van der Borcht and no longer extant) might have been made. The second part looks at the image itself, and discusses possible reasons for Hollar's production of the engraving some twenty years after the original. PMID- 15295819 TI - Heat, sex, and sugar: pregnancy and childbearing in the slave quarters. AB - This article examines the dynamics of slave fecundity in the antebellum South and analyzes the relationship between the planters' labor requirements and pregnancy on large sugar estates in nineteenth-century Louisiana. In contrast to the cotton states, where the slave population grew, bondspeople in Louisiana's sugar world experienced natural population decrease. This derived in part from imbalanced sex ratios, but as this article explores, it also occurred because of the punishing nature of sugar production that taxed slave women in distinct ways over the entire year. As this article shows, conceptions peaked during the annual harvest season but collapsed at other times because of nutritional stress, overwork, heat, and exhaustion. Addressing the seasonality of slave childbirths, the article posits that workload combined with climatic, ecological, hormonal, nutritional, and lactation factors ultimately shaped the reproductive ecology of American slavery. PMID- 15295820 TI - Abstracts of the Italian Parasitology Society XXIII National Congress. Vietri sul Mare, Italy, 9-12 June 2004. PMID- 15295821 TI - Abstracts of the 2004 Annual Meeting of the Society for Academic Emergency Medicine. May 16-19, 2004, Orlando, Florida, USA. PMID- 15295822 TI - Academic Emergency Medicine and the "Tragedy of the Commons" defined. PMID- 15295824 TI - Membership list of the Belgium Royal Academy of Medicine, 2004. PMID- 15295823 TI - Dynamics of pharmacy regulation in Denmark, 1546-1932: a study of profession state relations. PMID- 15295825 TI - Medicare home health care in rural America. AB - The past decade has brought many changes to the home health care industry, largely as a result of Medicare policy changes. These policy reforms include a new payment system, eligibility restrictions, and stringent fraud and abuse enforcement. In addition, Medicare now pays for home health care based on the location of the beneficiary, not the agency. To examine the impact of these changes on access to care, we evaluated the degree to which beneficiaries are served by agencies outside of their county. We constructed an analytical file by linking the 1997 five percent Medicare Standard Analytical File home health claims file to the Provider of Services file to obtain the characteristics of the beneficiaries' primary agency. This beneficiary-level analytical file included information on 162,241 Medicare home health users - including 43,488 rural residents - of 9,410 home health agencies. We examined the characteristics of rural beneficiaries served by urban agencies as compared with those served by rural agencies. Our findings demonstrate that urban agencies - either directly or through their branch offices - play an important role in providing home health care to rural Medicare beneficiaries. PMID- 15295826 TI - The moral economy of the drug company-medical scientist collaboration in interwar America. AB - This paper explores the exchange relationships underlying collaborations between pharmaceutical companies and preclinical (laboratory-based) researchers, in universities and similar contexts, during the interwar period. It also examines the arguments advanced to justify such collaborations in particular contexts as a way of investigating the perceived costs and benefits, especially among the academic parties in these collaborations, and the way these collaborations were regarded in the US biomedical research community. PMID- 15295827 TI - Financially distressed rural hospitals in four states. AB - The Balanced Budget Act of 1997 dramatically changed the payment environment for institutional providers of non-acute health services by mandating a shift in Medicare reimbursement of outpatient, home health, and skilled nursing services from the traditional cost-based approaches to prospective payment. Although they were designed to slow health care spending, these Medicare payment reforms, particularly the outpatient prospective payment system (OPPS) rules, were projected to have a disproportionately negative impact on many rural hospitals. Subsequent revisions to the Balanced Budget Act (BBA) modified the initial legislation to alleviate or postpone the negative financial impact, including a hold harmless provision for small (100-bed or under) rural hospitals. Due to delays in processing hospital cost reports, sufficient data to assess the impact of the new outpatient payment system on small rural hospitals have only recently become available. We simulated the effect of OPPS on the financial performance of rural hospitals in four states - Iowa, Texas, Washington, and West Virginia. Our findings suggest that the profitability and cash position of small, government owned, and Medicare-dependent hospitals will be adversely impacted by outpatient PPS. The results also suggest that the number of financially distressed rural hospitals will increase significantly. The small rural hospitals currently protected by the hold harmless provision are those that are likely to be hardest hit by OPPS. PMID- 15295828 TI - Olson's embryo problem. PMID- 15295829 TI - Teaching research ethics: illustrating the nature of the researcher-IRB relationship. AB - I collected information on the effects of ethical concerns on research questions asked and methods used by psychological researchers. Faculty researchers described the advantages and disadvantages of ethical concerns on their specialized areas and provided case examples of the effects of institutional review board actions on their research questions and methods. I offer several suggestions for using this information to increase students' ethical awareness and understanding in research methods courses. PMID- 15295831 TI - Manufacturing desire: the commodification of female sexual dysfunction. AB - The process of bringing new drugs to market interweaves commercialism, science, clinical medicine, and governmental regulation. Through their authority and public persona as medical experts, academic clinical trial researchers studying these pharmaceuticals are integral to this process, serving as mediators between producers (the pharmaceutical companies) and consumers (clinicians and patients) of new drugs through a complex set of exchange networks. Using examples from my ethnographic research on the search for pharmaceuticals to treat what has become known as female sexual dysfunction, this paper explores the links academic researchers make with drug manufacturers and consumer markets. Academic researchers have become an integral aspect of drug development, not only by conducting clinical trial research, but also by participating in a number of other activities that assist pharmaceutical companies in identifying and creating new markets. In this paper, i examine how researchers attend professional meetings where they present clinical trial data, lecture at continuing medical education conferences, and offer themselves as ' experts' to raise awareness about disorders and their treatments. Modifying a sociology of technology approach, this paper focuses on the actors in the social network who mediate the junctions between technological producers and consumers. This extends work in this area through theorizing the linkages between exchange networks, commodification techniques, and technoscientific developments. PMID- 15295832 TI - Perspectives of rural hospitals on bioterrorism preparedness planning. AB - Even the smallest, most isolated rural hospitals are now required to have bioterrorism preparedness plans. From the perspective of many rural hospitals, however, there is a disparity between Federal expectations and the realities of small hospitals operating in geographically isolated communities. As part of an effort to better understand how to close this gap, the Walsh Center for Rural Health Analysis convened a panel of representatives of rural hospitals who are responsible for bioterrorism preparedness in their hospitals. Perspectives of rural hospitals on various aspects of preparedness were discussed, in terms of workforce and training, physical capacity and supplies, communication, and coordination with other entities. All of the participants noted the tremendous progress that has been made in the past two years, but also the distance they each need to go. Some of the issues raised by the panelists included the dual benefit of efforts to increase capacity at rural hospitals, the inapplicability of many federal guidelines and directives for small hospitals because of size and less sophisticated infrastructure, the burden of geographic isolation relative to obtaining training and information, and the fragmentation of funding and directives at both the state and federal levels. PMID- 15295833 TI - Regulating clinical research: informed consent, privacy, and IRBs. PMID- 15295834 TI - No pain, no gain, no compensation: exploiting professional athletes through substandard medical care administered by team physicians. PMID- 15295835 TI - A look at the rights and entitlements of posthumously conceived children: no surefire way to tame the reproductive Wild West. PMID- 15295836 TI - Exploring the impact of Medicare's post-acute care transfer payment policy on rural hospitals. PMID- 15295837 TI - Abstracts of the 9th Annual Conference of the Japanese Society for Hemodiafiltration in conjunction with the International Symposium on Hemodiafiltration Therapy. August 30-31, 2003, Kanagawa, Japan. PMID- 15295839 TI - Abstracts from the 9th International Conference on Alzheimer's Disease and Related Disorders. July 17-22, 2004. Philadelphia, Pennsylvania, USA. PMID- 15295840 TI - Abstracts of the 67th Scientific Conference of the Polish Cardiac Society. Zabrze, 13-15 May 2004. PMID- 15295838 TI - Flatlanders. PMID- 15295841 TI - Abstracts of the 13th International Symposium on Bioluminescence and Chemiluminescence. PMID- 15295842 TI - Abstracts of the 20th International Conference on Pharmacoepidemiology and Therapeutic Risk Management. Bordeaux, France, 22-25 August 2004. PMID- 15295843 TI - Abstracts of the 20th Scientific Meeting of the International Society of Hypertension. February 15-19, 2004, Sao Paulo, Brazil. PMID- 15295844 TI - EUROMIT 6. Abstracts of the 6th European Meeting on Mitochondrial Pathology. 1-4 July 2004, Nijmegen, The Netherlands. PMID- 15295846 TI - Abstracts of the 81st Annual Meeting of the Physiological Society of Japan. June 2-4, 2004, Sapporo, Japan. PMID- 15295845 TI - Abstracts of the 18th International Congress on Thrombosis. Ljubljana, Slovenia, June 20-24, 2004. PMID- 15295847 TI - Abstracts of the 40th Annual Meeting of the American Society of Clinical Oncology. New Orleans, Louisiana, USA, June 5-8, 2004. PMID- 15295848 TI - "Geographical morality" revisited: international relations, international law, and the controversy over placebo controlled HIV clinical trials in developing countries. PMID- 15295849 TI - [Radical retropubic prostatectomy]. PMID- 15295850 TI - Fetal personhood and the sorites paradox. PMID- 15295851 TI - Is abortion vicious? PMID- 15295852 TI - Is genetic engineering wrong, per se? PMID- 15295853 TI - Distinctive moralities: the value of life and our duties to the handicapped. PMID- 15295854 TI - Trends in U.S. health insurance coverage, 2001-2003. AB - Against the backdrop of a sluggish economy and rapidly rising health insurance premiums, the proportion of Americans under age 65 covered by employer-sponsored insurance fell dramatically from 67 percent to 63 percent between 2001 and 2003. Although the decline in employer coverage could have spurred a large increase in the uninsured, the proportion of Americans without health insurance did not increase significantly, according to findings from the Center for Studying Health System Change's (HSC) Community Tracking Study Household Survey. Expansion of public health insurance--including Medicaid and the State Children's Health Insurance Program (SCHIP)--forestalled a significant increase in the uninsured, as the proportion of the under-65 population enrolled in public coverage increased from 9 percent to 12 percent. PMID- 15295855 TI - Transsexual prisoners: how much treatment is enough? PMID- 15295856 TI - The ethics of reproductive control. PMID- 15295857 TI - Still stuck in the cuckoo's nest: why do courts continue to rely on antiquated mental illness research? PMID- 15295858 TI - Thyroid cancer has increased in the adult populations of countries moderately affected by Chernobyl fallout. AB - BACKGROUND: The incidence of thyroid carcinoma increased among children affected by Chernobyl fallout. Less evidence exists for a corresponding effect in adolescents and adults. The Cancer Registry of the Czech Republic provides an opportunity to study various determinants of the occurrence of thyroid cancer. MATERIAL/METHODS: Anonymous population-based incidence data on thyroid carcinoma of the Czech Republic from 1976 to 1999 were obtained from the Czech Statistical Office (CSO) and the Institute of Health Information and Statistics (IHIS). This study covers 247 million person-years. Linear logistic regression models allowing for possible changes in slope (change-points) are suggested for the trends of incidence proportions. RESULTS: From 1976 to 1999 a uniform annual increase of 2.0% per year was found in the directly age-standardized thyroid cancer incidence proportion (95%-CI: 1.3-2.7, p<0.0001). From 1990 on, we observed an additional significant increase in the thyroid cancer incidence of 2.6% per year (95%-CI:1.2 4.1, p=0.0003). This effect (change-point) is essentially independent of age but dependent on gender: females 2.9% per year (95%-CI: 1.3-4.7, p=0.0006), males 1.8% per year (95%-CI: -1.0-4.7,p=0.2127). The estimated minimum latency period for the population as a whole is 4 years. CONCLUSIONS: Although the Czech Republic received only a relatively moderate amount of radioactive fallout,an unexpected uniformly accelerated increase of thyroid cancer in all age categories is seen from 1990 onwards. Therefore one should look carefully at collective dose and at the group of persons low in individual organ dose but high in number. PMID- 15295859 TI - Cervical cancer screening 'effective everywhere'. PMID- 15295860 TI - New visa 'will attract scientists to Europe'. PMID- 15295861 TI - Why we need to revive the autopsy. PMID- 15295862 TI - Minimally invasive thoracic surgery taking off. PMID- 15295863 TI - Not-for-profits respond to Scruggs' lawsuits. PMID- 15295864 TI - Hospitals reconsider outsourcing key support and clinical departments. PMID- 15295865 TI - Mesosomes and scientific methodology. AB - In his recent article, Nicolas Rasmussen (2001) is harshly critical of what he terms 'empirical philosophy of science', a philosophy that takes seriously the history of science in advancing philosophical pronouncements about science. He motivates his criticism by reflecting on recent history in microbiology involving the 'discovery' of a new bacterial organelle, the mesosome, during the 1950's and 1960's, and the subsequent retraction of this discovery by experimental microbiologists during the late 1970's and early 1980's. In particular, he argues that there was a lack of constancy in the methods microbiologists used in approaching the issue of the existence of mesosomes, and that in fact a similar sort of 'methodological flux' pervades all experimental work. My goal here is to refute Rasmussen's doctrine of flux, and in turn to re-establish order in our understanding of the methods and strategies of experimenters. My strategy in achieving this goal is to re-visit the same crucial research articles in the history of the mesosome episode that Rasmussen (2001) visits; and what I find upon returning to this literature is not flux, as Rasmussen seems to find, but a constancy of method in experimental reasoning, a constancy codified by what I call 'reliable process reasoning'. PMID- 15295866 TI - Homeosis and atavistic regeneration: the 'biogenetic law' in Entwicklungsmechanik. AB - Homeosis is a developmental abnormality corresponding to the transformation of a part of the body into another one. This term was introduced in 1894 by William Bateson, who aimed to make an inventory of all kinds of biological variation in order to understand how evolution proceeds. But, immediately afterwards experimental embryology, or Entwicklungsmechanik in Germany, adopted and redefined this term to refer to abnormal regenerations in which the newly developed organ was not identical to the initial one but rather resembled another part of the body. At that time, many experimental embryologists, such as Wilhelm Roux, were calling for the elimination of any phylogenetic explanation of development and were attempting to promote more mechanistic, proximate explanations. Despite these recommendations, several biologists continued to account for developmental processes by turning to phylogeny instead of mechanical forces. The case of homeosis is representative. Indeed, abnormal regenerations were often seen as examples of atavisms, or recurrence of ancestral characteristics, and many embryologists appealed to Ernst Haeckel's 'fundamental biogenetic law' to explain these strange phenomena. The break between Haeckelian tradition and Entwicklungsmechanik is thus less radical than often assumed, and the homeosis concept represents one of the factors of this continuity. PMID- 15295867 TI - The dual biological identity of human beings and the naturalization of morality. AB - The last two centuries have been the centuries of the discovery of the cell evolution: in the XIX century of the germinal cells and in the XX century of two groups of somatic cells, namely those of the brain-mind and of the immune systems. Since most cells do not behave in this way, the evolutionary character of the brain-mind and of the immune systems renders human beings formed by t wo different groups of somatic cells, one with a deterministic and another with an indeterministic (say Darwinian) behavior. An inherent consequence is that of the generation, during ontogenesis, of a dual biological identity. The concept of the dual biological identity may be used to explain the Kantian concept of the two metaphysical worlds, namely of the causal necessity and of the free will (Azzone, 2001). Two concepts, namely those of complex adaptive systems (CAS) and of emergence (Holland, 2002), are useful tools for understanding the mechanisms of adaptation and of evolution. The concept of complex adaptive systems indicates that living organisms contain series of stratified components, denoted as building blocks, forming stratified layers of increasing complexity. The concept of emergence implies the use of repeating patterns and of building blocks for the generation of structures of increasing levels of complexity, structures capable of exchanging communications both in the top-down and in the bottom-up direction. Against the concept of emergence it has been argued that nothing can produce something which is really new and endowed of causal efficacy. The defence of the concept of emergence is based on two arguments. The first is the interpretation of the variation-selection mechanism as a process of generation of information and of optimization of free energy dissipation in accord with the second principle of thermodynamics. The second is the objective evidence of the cosmological evolution from the Big Bang to the human mind and its products. Darwin has defended the concept of the continuity of evolution. However evolution should be considered as continuous when there is no increase of information and as discontinuous when there is generation of new information. Examples of such generation of information are the acquisition of the innate structures for language and the transition from absence to presence of morality. There are several discontinuity thresholds during both phylogenesis and ontogenesis. Morality is a relational property dependent on the interactions of human beings with the environment. Piaget and Kohlberg have shown that the generation of morality during childhood occurs through several stages and is accompanied by reorganization of the child mental organization. The children respect the conventions in the first stage and gradually generate their autonomous morality. The transition from absence to presence of morality, a major adaptive process, then, not only has occurred during phylogenesis but it occurs again in every human being during ontogenesis. The religious faith does not provide a logical justification of the moral rules (Ayala, 1987) but rather a psychological and anthropological justification of two fundamental needs of human beings: that of rendering Nature an understandable entity, and that of increasing the cooperation among members of the human societies. The positive effects of the altruistic genes in the animal societies are in accord with the positive effects of morality for the survival and development of the human societies. PMID- 15295868 TI - Biology with information and meaning. AB - It is shown that information and meaning can be defined by operative procedures, and that we need to recognize them as new types of natural entities. They are not quantities (neither fundamental nor derived) because they cannot be measured, and they are not qualities because they are not subjective features. Here it is proposed to call them nominable entities, i.e., entities which can be specified only by naming their components in their natural order. PMID- 15295869 TI - Biology without information. PMID- 15295870 TI - The use of information theory in biology: a historical perspective. PMID- 15295871 TI - Fourier transform infrared (FT-IR) spectrometry. PMID- 15295872 TI - Selected uses of enzymes with critical fluids in analytical chemistry. AB - The use of enzymes coupled with supercritical fluid (SF)-based analytical techniques, such as supercritical fluid extraction (SFE), provides a safer environment platform for the analytical chemist and reduces the use of organic solvents. Incorporation of such techniques not only reduces the use of solvent in analytical laboratories, but it can also lead to overall method simplification and time savings. In this review, some of the fundamental aspects of using enzymes in the presence of SF media are discussed, particularly the influence of extraction (reaction) pressure, temperature, and water content of the extracting fluid and/or the sample matrix. Screening of optimal conditions for conducting reactions in the presence of SF media can be readily accomplished with automated serial or parallel SFE instrumentation, including selection of the proper enzyme. Numerous examples are cited, many based on lipase-initiated conversions of lipid substrates, to form useful analytical derivatives for gas chromatography, liquid chromatography, or SF chromatography analysis. In certain cases, enzymatic-aided processing of samples can permit the coupling of the extraction, sample preparation, and final analysis steps. The derived methods/techniques find application in nutritional food analyses, assays of industrial products, and micro analyses of specific samples. PMID- 15295873 TI - A simple and rapid spectrophotometric method for determination of thiophanate methyl in commercial formulations and its residues in foodstuffs. AB - A new, simple, rapid, and sensitive spectrophotometric method for the determination of thiophanate-methyl, based on its reaction with cobalt(II) in the presence of triethylamine, has been developed. The yellowish green color that develops instantaneously on mixing the fungicide with the reagents in dimethylformamide is stable for at least 2 h and has maximum absorbance at 360 nm. The method has been successfully applied to the determination of thiophanate methyl in its commercial formulations and residues on grains and apples. A photometric titration procedure for formulation analysis of the fungicide has also been developed. PMID- 15295874 TI - Sample preparation and determination of ginkgo terpene trilactones in selected beverage, snack, and dietary supplement products by liquid chromatography with evaporative light-scattering detection. AB - Ginkgo biloba leaf extract has been widely used in dietary supplements and more recently in some foods and beverages. Sample preparation procedures for determination of ginkgo terpene trilactones (including bilobalide and ginkgolides A, B, C, and J) in various sample matrixes were developed in this study. Ginkgo leaves and capsules were extracted with 5% KH2PO4 aqueous solution under sonication. Tea bags were extracted with boiling water, whereas drink samples were taken directly from the bottles. After filtration and the addition of NaCl to approximately 30% (w/v), the terpene trilactones in aqueous solutions were quantitatively extracted with ethyl acetate-tetrahydrofuran (4 + 1, v/v). Puff samples (a cereal-based fried snack item) were first defatted by using hexane or by using supercritical fluid extraction and then extracting under sonication with methanol-acetic acid (99 + 1, v/v). After evaporation of the organic phase, the terpene trilactones were redissolved in methanol and determined on a C18 reversed phase column by liquid chromatography (LC) with evaporative light-scattering detection. The method of standard additions and gas chromatography with flame ionization detection were used for method validation. For most samples, the relative standard deviation was <10%. The identities of target compounds in ginkgo leaves and drink samples were confirmed by LC/electrospray ionization tandem mass spectrometry. PMID- 15295875 TI - Determination of trimetazidine dihydrochloride in the presence of its acid induced degradation products. AB - Three methods are presented for the determination of trimetazidine dihydrochloride in the presence of its acid-induced degradation products. The first method was based on measurement of first-derivative D1 value of trimetazidine dihydrochloride at 282 nm over a concentration range of 8.00-56.00 microg/mL with mean percentage accuracy of 99.80+/-1.17. The second method was based on first derivative of the ratio spectra DD1 at 282 nm over the same concentration range with the percentage accuracy of 99.14+/-0.68. The third method was based on separation of trimetazidine dihydrochloride from its acid induced degradation products followed by densitometric measurement of the spots at 215 nm. The separation was performed on silica gel 60 F254 using methanol ammonia (100+/-1.5, v/v) as mobile phase. This method was applicable for determination of the intact drug in the presence of its degradation products over a concentration range of 2.00-9.00 microg/spot with mean percentage accuracy of 99.86+/-0.92. The proposed methods were successfully applied for the determination of trimetazidine dihydrochloride in bulk powder, laboratory prepared mixtures containing different percentages of degradation products, and pharmaceutical dosage forms. The validity of results was assessed by applying the standard addition technique. The results obtained agreed statistically with those obtained by the reported method. PMID- 15295876 TI - Application of the wavelet method for the simultaneous quantitative determination of benazepril and hydrochlorothiazide in their mixtures. AB - The discrete and continuous wavelet transforms were applied to the overlapping signal analysis of the ratio data signal for simultaneous quantitative determination of the title subject compounds in samples. The ratio spectra data of the binary mixtures containing benazepril (BE) and hydrochlorothiazide (HCT) were transferred as data vectors into the wavelet domain. Signal compression, followed by a 1-dimension continuous wavelet transform (CWT), was used to obtain coincident transformed signals for pure BE and HCT and their mixtures. The coincident transformed amplitudes corresponding to both maximum and minimum points allowed construction of calibration graphs for each compound in the binary mixture. The validity of CWT calibrations was tested by analyzing synthetic mixtures of the investigated compounds, and successful results were obtained. All calculations were performed within EXCEL, C++, and MATLAB6.5 softwares. The obtained results indicated that our approach was flexible and applicable for the binary mixture analysis. PMID- 15295877 TI - New liquid chromatographic method for determination of rabeprazole sodium in coated tablets. AB - Rabeprazole sodium is a proton pump inhibitor that covalently binds and inactivates the gastric parietal cell proton pump (H+/K+ ATPase). Little has been published about the quantitative determination of this drug. The aim of this research was to develop a new liquid chromatographic method for quantitative determination of rabeprazole in coated tablets. The system consisted of a Hypersil Keystone Betabasic C8 column (250 x 4.6 mm, 5 microm particle size), an isocratic acetonitrile-water (35 + 65) mobile phase at a flow rate of 1.0 mL/min, and a diode array detector set at 282 nm. The following validation parameters were evaluated: linearity, precision, accuracy, specificity, detection and quantitation limits, and robustness. The method showed good linearity in the concentration range of 10-70 microg/mL. The quantitation limit was 2.43 microg/mL, and the detection limit was 0.80 microg/mL. The intra- and interday precision data showed that the method has good reproducibility (relative standard deviation = 1.03). Accuracy and robustness were also evaluated, and the results were satisfactory. The mean recovery was 101.61%. The analysis of a placebo mixture demonstrated the method is also specific. PMID- 15295878 TI - Application of derivative spectrophotometry for simultaneous determination of quinapril and hydrochlorothiazide in the combination tablets. AB - A new second-order-derivative spectrophotometric method using zero-crossing technique measures quinapril (QUI) and hydrochlorothiazide (HYD) in 2-component mixtures. The procedure does not require prior separation of components from the sample. QUI was determined at a wavelength of 211.6 nm (zero-crossing wavelength point of HYD). Similarly, HYD was measured at 270.8 nm (zero-crossing wavelength point of QUI ). Calibration graphs were constructed over the concentration range of 4.0 to 24.0 microg/mL for QUI and 2.5 to 15.0 microg/mL for HYD. Detection and quantitation limits were 0.85 and 2.5 microg/mL for QUI and 0.12 and 0.4 microg/mL for HYD, respectively. The accuracy (recovery 100.5-102%), precision (relative standard deviation less than 3.5% for QUI and 1.5% for HYD), selectivity, and sensitivity of the elaborated methods were satisfactory. The proposed method was applied successfully for the determination of both drugs in QUI-HYD tablets. PMID- 15295879 TI - A sensitive extracto-photometric method for determination of residual chlorine in greywater. AB - A new photometric method for chlorine determination based on the oxidative transformation of iodide to iodine and subsequent extraction in ethyl acetate has been developed. The effects of several chemical variables (pH, ionic strength, and iodide concentration) have been studied. Characteristics of the method were linear range 0-0.6 mg C12/L, limit of detection 5 microg Cl2/L, and coefficient of variation 0.6%. The method has been applied to greywater without previous sample treatment. PMID- 15295880 TI - Toward an international standard for PCR-based detection of foodborne Escherichia coli O157: validation of the PCR-based method in a multicenter interlaboratory trial. AB - The performance of a polymerase chain reaction (PCR) method for detection of Escherichia coli O157, previously validated on DNA extracted from pure cultures, was evaluated on spiked cattle swabs through an interlaboratory trial, including 12 participating laboratories from 11 European countries. Twelve cattle swab samples, spiked at 4 levels (0, 1-10, 10-100, and 100-1000 colony-forming units, in triplicate) with E. coli O157 were prepared centrally in the originating laboratory; the receiving laboratories performed pre-PCR treatment followed by PCR. The results were reported as positive when the correct amplicons were present after gel electrophoresis. The statistical analysis, performed on 10 sets of reported results, determined the diagnostic sensitivity to be 92.2%. The diagnostic specificity was 100%. The accordance (repeatability) was 90.0%, calculated from all positive inoculation levels. The concordance (reproducibility) was 85.0%, calculated from all positive inoculation levels. The concordance odds ratio (degree of interlaboratory variation calculated from all positive inoculation levels) was 1.58, indicating the robustness of the PCR method. Thus, the interlaboratory variation due to personnel, reagents, minor temperature or pH fluctuations and, not least, thermal cyclers, did not affect the performance of the method, which is currently being considered as part of an intenational PCR standard. PMID- 15295881 TI - Multicenter validation of PCR-based method for detection of Salmonella in chicken and pig samples. AB - As part of a standardization project, an interlaboratory trial including 15 laboratories from 13 European countries was conducted to evaluate the performance of a noproprietary polymerase chain reaction (PCR)-based method for the detection of Salmonella on artificially contaminated chicken rinse and pig swab samples. The 3 levels were 1-10, 10-100, and 100-1000 colony-forming units (CFU)/100 mL. Sample preparations, including inoculation and pre-enrichment in buffered peptone water (BPW), were performed centrally in a German laboratory; the pre-PCR sample preparation (by a resin-based method) and PCR assay (gel electrophoresis detection) were performed by the receiving laboratories. Aliquots of BPW enrichment cultures were sent to the participants, who analyzed them using a thermal lysis procedure followed by a validated Salmonella-specific PCR assay. The results were reported as negative or positive. Outlier results caused, for example, by gross departures from the experimental protocol, were omitted from the analysis. For both the chicken rinse and the pig swab samples, the diagnostic sensitivity was 100%, with 100% accordance (repeatability) and concordance (reproducibility). The diagnostic specificity was 80.1% (with 85.7% accordance and 67.5% concordance) for chicken rinse, and 91.7% (with 100% accordance and 83.3% concordance) for pig swab. Thus, the interlaboratory variation due to personnel, reagents, thermal cyclers, etc., did not affect the performance of the method, which will be proposed as part of a developing international PCR standard. PMID- 15295882 TI - Evaluation of VIDAS Salmonella (SLM) immunoassay method with Rappaport Vassiliadis (RV) medium for detection of Salmonella in foods: collaborative study. AB - A collaborative study was conducted to compare the VIDAS Salmonella (SLM) with Rappaport-Vassiliadis (RV) method for detection of Salmonella in foods to the current standard method presented in the U.S. Food and Drug Administration's Bacteriological Analytical Manual (BAM) and the culture method presented in AOAC's Official Methods of Analysis. The VIDAS SLM with RV method uses tetrathionate broth in combination with RV medium in place of selenite cystine broth for selective enrichment, thereby eliminating the hazardous waste issue for laboratories. Twenty five laboratories participated in the evaluation, each testing one or more of 8 test products: nonfat dry milk, dried egg, soy flour, lactic casein, milk chocolate, raw ground pork, raw ground turkey, and raw peeled shrimp. Results of the study showed no significant differences in the numbers of confirmed positive samples with the VIDAS SLM with RV procedure and the BAM/AOAC culture procedure. The VIDAS SLM with RV method was effective for rapid detection of Salmonella in foods. It is recommended that AOAC INTERNATIONAL modify the VIDAS Salmonella SLM procedure to include the RV method. PMID- 15295883 TI - Testing green coffee for ochratoxin A, part I: estimation of variance components. AB - The variability associated with testing lots of green coffee beans for ochratoxin A (OTA) was investigated. Twenty-five lots of green coffee were tested for OTA contamination. The total variance associated with testing green coffee was estimated and partitioned into sampling, sample preparation, and analytical variances. All variances increased with an increase in OTA concentration. Using regression analysis, mathematical expressions were developed to model the relationship between OTA concentration and the total, sampling, sample preparation, and analytical variances. The expressions for these relationships were used to estimate the variance for any sample size, subsample size, and number of analyses for a specific OTA concentration. Testing a lot with 5 microg/kg OTA using a 1 kg sample, Romer RAS mill, 25 g subsamples, and liquid chromatography analysis, the total, sampling, sample preparation, and analytical variances were 10.75 (coefficient of variation [CV] = 65.6%), 7.80 (CV = 55.8%), 2.84 (CV = 33.7%), and 0.11 (CV = 6.6%), respectively. The total variance for sampling, sample preparation, and analytical were 73, 26, and 1%, respectively. PMID- 15295884 TI - Preparation and certification of zearalenone mass concentration of two low-level maize reference materials. AB - The contamination of maize by fungi, especially by Fusarium species, is a worldwide problem. One of the most prevalent Fusarium mycotoxins frequently found on European maize is zearalenone (ZON), which has been implicated in a range of human and animal diseases. It shows remarkable estrogenic properties and can cause severe infertility problems in farm animals. Currently, 9 countries have set maximum tolerable levels for ZON in food, ranging from 0 to 1000 microg/kg. This paper describes the preparation of 2 maize reference materials (BCR-716 very low level ZON and BCR-717 low level ZON) and the certification of their individual ZON contents (mass concentration and mass fraction). Uncertainties were calculated in compliance with the Guide to the Expression of Uncertainty in Measurement and include uncertainties that are due to possible inhomogeneity and instability. Finally, BCR-716 was certified at a level of <5 microg/kg and BCR 717 at a level of 83 microg/kg with an expanded uncertainty (k = 2) of 9 microg/kg. PMID- 15295885 TI - Purity assessment of commercially available crystalline deoxynivalenol. AB - Deoxynivalenol (DON) obtained from 2 commercial sources was characterized, and its purity was determined. The structural identity of DON was confirmed by 1H and 13C-nuclear magnetic resonance (NMR) spectroscopy, gas chromatography with mass spectrometric (GC/MS) detection, and infrared/attenuated total reflectance (IR/ATR) spectroscopy. NMR spectra showed shifts that varied from previously published data. However, we established a complete, unambiguous assignment for all signals. Chromatograms obtained by GC/MS were almost identical for both investigated samples and confirmed the structure of DON. Likewise, IR/ATR spectra verified the identity of DON. The degree of purity was determined by liquid chromatography (LC) with a variable wavelength detector, LC/MS/MS, GC with electron-capture detection (GC-ECD), and ultraviolet (UV) spectrophotometry. The purity check using LC showed a single peak in both chromatograms. With LC/MS/MS measurements, we could detect small amounts of impurities in the crystalline DON from both sources. In data obtained by GC-ECD, no differences in purity were observed. The UV measurements showed an absorption maximum at 217 nm. The mean epsilon(m) of the extinction coefficients was calculated as 6727 (L/cm/mol) for DON (Sigma) and 6825 (L/cm/mol) for DON (Biopure). Finally, the purity of DON from the 2 commercial sources was calculated as >96 and >98%, respectively. Although the DON produced by both providers can be considered sufficiently pure for routine analysis of trichothecenes in food and feed, this work again demonstrated that the impurity of the solid mycotoxin constitutes the greatest contribution to the overall uncertainty of a mycotoxin calibrant. PMID- 15295886 TI - Automatic flow procedure for the determination of ethanol in wine exploiting multicommutation and enzymatic reaction with detection by chemiluminescence. AB - An automatic procedure for the determination of ethanol in wines using a flow system based on multicommutation and enzymatic reaction is described. Alcohol oxidase was immobilized on aminopropyl glass beads and packed in an acrylic column. The peroxide due to enzymatic reaction with ethanol reacted with luminol and generated the chemiluminescence radiation that was monitored by using a laboratory-made detector based on photodiodes. The system manifold comprised a set of 3-way solenoid valves controlled by a microcomputer furnished with electronic interfaces, which ran on software written in Quick BASIC 4.5 to provide facilities to perform on-line sample dilution, reagent addition, and data acquisition. After system parameters optimization, ethanol samples were processed without prior pretreatment. The following suitable features were achieved: linear response ranging from 2.5 to 25% (v/v) ethanol, relative standard deviation of 1.8% (n = 10), detection limit of 0.3% (v/v) ethanol, sampling rate of 23 determinations per hour, and low reagent consumption of 0.23 mg luminol and 7 mg hexacyanoferrate (III) per determination. When the results were compared with those obtained using the AOAC Official Method, no significant difference at the 90% confidence level was observed. PMID- 15295887 TI - The modular analytical procedure and validation approach and the units of measurement for genetically modified materials in foods and feeds. AB - Food and feed analysts are confronted with a number of common problems, irrespective of the analytical target. The analytical procedure can be described as a series of successive steps: sampling, sample processing, analyte extraction, and ending, finally, in interpretation of an analytical result produced with, e.g., real-time polymerase chain reaction. The final analytical result is dependent on proper method selection and execution and is only valid if valid methods (modules) are used throughout the analytical procedure. The final step is easy to validate-the measurement uncertainty added from this step is relatively limited and can be estimated with a high degree of precision. In contrast, the front-end sampling and processing steps have not evolved much, and the corresponding methods are rarely or never experimentally validated according to internationally harmonized protocols. In this paper, we outlined a strategy for modular validation of the entire analytical procedure, using an upstream validation approach, illustrated with methods for genetically modified materials that may partially apply also to other areas of food and feed analyses. We have also discussed some implications and consequences of this approach in relation to reference materials, measurement units, and thresholds for labelling and enforcement, and for application of the validated methods (modules) in routine food and feed analysis. PMID- 15295888 TI - Development of a solid-phase extraction method for determination of pheophorbide a and pyropheophorbide a in health foods by liquid chromatography. AB - A simple solid-phase extraction (SPE) method was developed for the liquid chromatography (LC) determination of pheophorbide (Phor) a and pyropheophorbide (Pyro) a in health foods such as chlorella, spirulina, etc. The food sample was extracted with 85% (v/v) acetone. The extract was acidified with hydrochloric acid and loaded on a C18 cartridge. After washing with water, Phor a and Pyro a were eluted with the LC mobile phase. Phor a and Pyro a were separated by isocratic reversed-phase LC and quantitated by fluorescence detection. The recoveries for spiked samples of chlorella and the extract were 87.1-102.0%. Commercial health foods (chlorella, spirulina, aloe, kale, Jews mallow, and green tea leaves) were analyzed using the SPE method. The values found for Phor a and Pyro a ranged from 2 to 788 microg/g and from <1 to 24 microg/g, respectively. There was no significant difference between the SPE method and the official method in Japan (spectrophotometry after liquid-liquid extraction). The advantages of the SPE method are the short extraction times, lack of emulsions, and reduced consumption of organic solvents compared with the official method in Japan. The SPE method is considered to be useful for the screening of Phor a and Pyro a in health foods. PMID- 15295889 TI - Variation of analytical results for peanuts in energy bars and milk chocolate. AB - Peanuts contain proteins that can cause severe allergic reactions in some sensitized individuals. Studies were conducted to determine the percentage of recovery by an enzyme-linked immunosorbent assay (ELISA) method in the analysis for peanuts in energy bars and milk chocolate and to determine the sampling, subsampling, and analytical variances associated with testing energy bars and milk chocolate for peanuts. Food products containing chocolate were selected because their composition makes sample preparation for subsampling difficult. Peanut-contaminated energy bars, noncontaminated energy bars, incurred milk chocolate containing known levels of peanuts, and peanut-free milk chocolate were used. A commercially available ELISA kit was used for analysis. The sampling, sample preparation, and analytical variances associated with each step of the test procedure to measure peanut protein were determined for energy bars. The sample preparation and analytical variances were determined for milk chocolate. Variances were found to be functions of peanut concentration. Sampling and subsampling variability associated with energy bars accounted for 96.6% of the total testing variability. Subsampling variability associated with powdered milk chocolate accounted for >60% of the total testing variability. The variability among peanut test results can be reduced by increasing sample size, subsample size, and number of analyses. For energy bars the effect of increasing sample size from 1 to 4 bars, subsample size from 5 to 20 g, and number of aliquots quantified from 1 to 2 on reducing the sampling, sample preparation, and analytical variance was demonstrated. For powdered milk chocolate, the effects of increasing subsample size from 5 to 20 g and number of aliquots quantified from 1 to 2 on reducing sample preparation and analytical variances were demonstrated. This study serves as a template for application to other foods, and for extrapolation to different sizes of samples and subsamples as well as numbers of analyses. PMID- 15295890 TI - Evaluation of sampling plans to detect Cry9C protein in corn flour and meal. AB - StarLink is a genetically modified corn that produces an insecticidal protein, Cry9C. Studies were conducted to determine the variability and Cry9C distribution among sample test results when Cry9C protein was estimated in a bulk lot of corn flour and meal. Emphasis was placed on measuring sampling and analytical variances associated with each step of the test procedure used to measure Cry9C in corn flour and meal. Two commercially available enzyme-linked immunosorbent assay kits were used: one for the determination of Cry9C protein concentration and the other for % StarLink seed. The sampling and analytical variances associated with each step of the Cry9C test procedures were determined for flour and meal. Variances were found to be functions of Cry9C concentration, and regression equations were developed to describe the relationships. Because of the larger particle size, sampling variability associated with cornmeal was about double that for corn flour. For cornmeal, the sampling variance accounted for 92.6% of the total testing variability. The observed sampling and analytical distributions were compared with the Normal distribution. In almost all comparisons, the null hypothesis that the Cry9C protein values were sampled from a Normal distribution could not be rejected at 95% confidence limits. The Normal distribution and the variance estimates were used to evaluate the performance of several Cry9C protein sampling plans for corn flour and meal. Operating characteristic curves were developed and used to demonstrate the effect of increasing sample size on reducing false positives (seller's risk) and false negatives (buyer's risk). PMID- 15295891 TI - Solid-phase extraction and cleanup procedures for determination of acrylamide in fried potato products by liquid chromatography/mass spectrometry. AB - In response to recent discoveries of acrylamide in heated foods, a solid-phase extraction and cleanup protocol was developed for the determination of acrylamide in fried or baked potato samples by liquid chromatography/mass spectrometry (LC/MS). The analyte was extracted from the matrix by using 2M NaCl, and an aliquot of the initial extract was loaded onto a reversed-phase cartridge. After the analyte was eluted from the cartridge, the eluate was cleaned up on a mixed mode cation-exchange cartridge. The eluate was then evaporated, and the residue was reconstituted in mobile phase before LC/MS analysis. Recoveries, based on the recovery of an added internal standard, ranged from 96 to 101% with relative standard deviations (RSDs) of 5-11%. The response was linear for a concentration range of 100-2000 ng/g with a coefficient of determination (R2) of 0.992 (n = 25). An interday study showed good accuracy and precision of the method over a 3 day period with a recovery of 98% and an RSD of 9.5% (n = 15). The analyses of 6 potato chip samples showed concentrations of incurred acrylamide ranging from 260 to 1500 ng/g. PMID- 15295892 TI - Model proficiency testing scheme for serological diagnosis of Brucellosis: interlaboratory study. AB - A model interlaboratory testing scheme was developed by the Italian National Reference Laboratory for Brucellosis. This scheme was planned for both qualitative (Rose Bengal Plate Test; RBPT) and quantitative (Complement Fixation Test; CFT) serological tests and involved a total of 42 laboratories. In the preparation of this scheme, reference was made to general protocols and guidelines and to methods reported in the literature, which were applicable to analytical chemistry laboratories. Six field sera from naturally infected animals, one positive serum at a titer below the European Union (EU) positivity threshold, and 5 sera positive at titers between 20 and 851 International Units of Complement Fixation Test (IUCFT)/mL plus one negative serum were used to produce a panel of test sera. To evaluate laboratory performances in the quantitative test for each tested sample examined, z-scores based on robust summary statistics (the median and normalized interquartile range) were used. To evaluate overall laboratory performance, 2 types of combined z-scores were used: Rescaled Sum of Scores and Sum of Squared Scores. In the case of the qualitative test (RBPT), results were analyzed by a Bayesian approach. A Beta distribution, based on the result of each laboratory, was calculated and used to estimate the probability of each laboratory giving a correct result and its uncertainty. PMID- 15295893 TI - Determination of multiclass pesticide residues in apple juice by gas chromatography-mass selective detection after extraction by matrix solid-phase dispersion. AB - A multiresidue method is described for the analysis of 106 multiclass pesticides (organochlorine, organophosphate, carbamate, pyrethroid, and triazine classes) in apple juice in a single injection. The determination procedure was based on matrix solid-phase dispersion of juice on diatomaceous earth in a glass column and subsequent extraction with a mixture of hexane-dichloromethane (1 + 1) at a flow rate of 5 mL/min. The analytes were determined by capillary gas chromatography with mass spectrometric detection and confirmed by their retention times and ion ratios. The coefficients of variation for analysis of samples fortified over the range of 0.01-0.2 mg/kg were 1.62 to 18.4%, and the recoveries for all analytes were between 70 and 110%. PMID- 15295894 TI - Determination of thallium in potassium chloride and electrolyte replenishers by electrothermal atomic absorption spectrometry. AB - Thallium in potassium chloride and electrolyte replenishers was determined by electrothermal atomic absorption spectrometry (ETAAS) with direct injection of a resin suspension. Thallium(III) was extracted on fine particles of a cellulose nitrate resin (CNR) from dilute HCl (pH 1.6) in the presence of ammonium pyrrolidine-1-carbodithioate. The CNR particles were collected on a membrane filter by filtration under suction, suspended in 1.0 mL 10mM HNO3, and then delivered directly to ETAAS as the suspension. The effects of chloride ions were thoroughly investigated. The results showed that the addition of 0.5mM NaCl to the suspension (10mM HNO3) was recommended, after CNR and a membrane filter holding the CNR were washed thoroughly with 0.025M HCl, to eliminate interference from chloride ions. No chemical modifier was required. Extraction from the solution containing up to 2M chloride ion was allowable. The proposed method gave a concentration factor of 50 for a 50 mL sample volume. The detection limit (3sigma, n = 5) was 1 ng (20 pg/mL). The relative standard deviation was 4.9% (n = 5) at 30 ng level of thallium. The content of thallium in potassium chloride was 15.7-32.8 ng/g, and in electrolyte replenishers was 0.18-4.16 ng/mL. PMID- 15295895 TI - Evaluation of a method based on liquid chromatography/electrospray tandem mass spectrometry for analyzing eight triazolic and pyrimidine fungicides in extracts of processed fruits and vegetables. AB - The feasibility of using liquid chromatography/electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) for determining 8 fungicides (triadimenol, penconazole, propiconazole, hexaconazole, cyproconazole, myclobutanil, fenarimol, and bitertanol) in extracts of tomato puree and lemon juice concentrate has been evaluated. A miniaturized extraction-partition procedure requiring small amounts of nonchlorinated solvents has been used. The extracts (5 microL) were analyzed by LC/ESI-MS/MS without any previous cleanup step. Chromatographic determination has been performed using a C18 column and isocratic elution. Seventeen MS/MS transitions of precursor ions were monitored simultaneously (2 or 3 for each pesticide). The excellent selectivity and good linearity of the LC/MS/MS method allowed quantitation and identification at low levels (limits of quantitation <0.010 mg/kg), even in difficult matrixes, with a run time of only 1.5 min. PMID- 15295896 TI - Determination of imidacloprid in fruits and vegetables by liquid chromatography with diode array and nitrogen-specific chemiluminescence detection. AB - A national trend is focusing on expanding usage of environmentally friendly nitrogenous pesticides and herbicides. However, some of these chemicals are undetectable by any gas chromatographic method due to their thermal lability or nonvolatility. Using imidacloprid as a model compound, this laboratory has developed a simple, effective, and reliable liquid chromatography (LC) method to address the inherent difficulties of these chemicals. Focusing on nitro, nitrate, and/or nitrite chromophore groups and components contained in the chemical structure of most nitrogenous compounds, LC with diode array detector and a nitrogen-specific detector in tandem enabled determination and confirmation analyses in series. Furthermore, using 2 mixtures of solvents in conjunction with 2-stage solid-phase extraction cleanup on aminopropyl/Florisil cartridges to eliminate and/or minimize interferences resulting from free amino acids, peptides, proteins, pigments, and lipids in the plant tissues made the qualitative and quantitative analysis simple, fast, and cost effective. This method demonstrated the potential for becoming a viable analysis tool for many other types of nonvolatile nitrogenous chemicals. PMID- 15295897 TI - Multiresidue analysis of pesticides in vegetables and fruits by gas chromatography/mass spectrometry after gel permeation chromatography and graphitized carbon column cleanup. AB - A multiresidue method for pesticides that enables quantitative, sequential analysis of a large number of vegetable and fruit samples by gas chromatography/mass spectrometry has been developed. First, 89 important target compounds were selected for monitoring, and then the appropriate internal standards for these pesticides, 14 stable isotopically labeled pesticides (surrogates), were used. The sample was extracted with acetonitrile, and the extract was cleaned up by a salting-out step followed by redissolution in ethyl acetate. Coextractives were removed automatically by gel permeation chromatography with a graphitized carbon column, and then by use of a tandem silica-gel/PSA cartridge column. Recoveries of 82 of the 89 pesticides from fortified spinach, tomato, apple, and strawberry were within a range from 70 to 120%, and the relative standard deviation values of 80 of the 89 pesticides were <5%. The method was applied to 188 commercial vegetable and fruit samples to demonstrate its use in routine analysis. PMID- 15295898 TI - Determination of residues of quaternary ammonium disinfectants in food products by liquid chromatography-tandem mass spectrometry. AB - A liquid chromatography-tandem mass spectrometry method has been developed for the determination of residues of alkylbenzyldimethylammonium, didecyldimethylammonium, didodecyldimethylammonium, and benzyldodecylhydroxyethylammonium compounds in various food matrixes. These quaternary ammonium compounds (QAs) are used in the food industry as disinfectants. According to the Dutch Food Law, the total mass (expressed as cetyltrimethylammonium chloride) of QAs in food products shall not exceed the legislative limit of 0.5 mg/kg. Samples were extracted by a simple salting-out procedure, using acetonitrile and sodium chloride; about 100 samples could be prepared and analyzed daily. Special care had to be taken to thoroughly homogenize samples and to avoid the use of contaminated labware. The method was validated by a procedure in compliance with EU Directive 2002/657. From the matrixes of ice cream and minced meat, recoveries of more than 95% with a relative standard deviation of about 3% were obtained by 3 different analysts (n = 54). Detection limits were in the low microg/kg range. The decision limit (CCalpha) was determined to be 0.55 mg/kg. Dairy and meat products, collected in The Netherlands, were analyzed (761 samples). In 1% of the meat samples, 2% of the ice cream and milkshake samples, and 24% of the whipped cream samples, the Dutch legislative limit was exceeded. Over 2000 injections could be performed on a single column without deterioration of the peak shapes or recoveries. PMID- 15295899 TI - Optimization of solid-phase microextraction for the gas chromatography/mass spectrometry analysis of persistent organic pollutants. AB - In this work, solid-phase microextraction (SPME) has been applied as an alternative for the selective extraction of 3 polybrominated diphenyl ethers (PBDEs), 2,2',4,4'-tetrabromodiphenyl ether (PBDE-47); 2,2',4,4',5 pentabromodiphenyl ether (PBDE-99); and 2,2',4,4',6-pentabromodiphenyl ether (PBDE-100), and 2 alkylphenols, 4-tert-OP and 4-NP, prior to their analysis by gas chromatography/mass spectrometry (GC/MS). The advantages of this technique are mainly its simplicity, cost-effectiveness, and time-saving sample preparation, as well as being a solvent-free technique. With the aim of optimizing the conditions for an efficient extraction of the studied compounds, different fiber coatings and the main parameters affecting the extraction procedure have been evaluated. The results obtained showed a good linearity in the range of concentrations investigated, and adequate relative standard deviation values were found according to the range accepted for SPME. Recovery values were in the range of 78-108%, and good detection and quantitation limits at ppt levels were obtained for both methods, allowing the determination of the selected compounds in samples at trace levels. The results obtained clearly show the potential of SPME for efficient concentration of the target compounds and also demonstrate the reliability of this extraction technique for their GC/MS analysis. PMID- 15295900 TI - Identification of microcystin-RR and [Dha7]microcystin-RR in commercial standards by electrospray ionization mass spectrometry. AB - Three different commercial standards of microcystin-RR were assessed for purity by the liquid chromatography coupled with electrospray ionization mass spectrometry (LC/ESI/MS) technique. Although the liquid chromatograms with photodiode array detector for each standard looked virtually identical, the analysis of corresponding mass spectra revealed that only one of them contained microcystin-RR per purity assay. The second standard was a mixture of microcystin RR, and its demethyl variant identified as [Dha7]microcystin-RR, and the third one contained [Dha7]microcystin-RR only. We strongly recommend applying LC coupled with MS for purity assay of microcystin standards. PMID- 15295901 TI - Silicosis and smoking strongly increase lung cancer risk in silica-exposed workers. AB - It remains controversial whether silica is a human lung carcinogen. In this study, we estimated the relative risks of lung cancer due to silica and silicosis by meta-analysis. We collected papers published from 1966-2001 which epidemiologically reported on the relationship between silica/silicosis and lung cancer. We removed papers which did not exclude the effects of asbestos and radioactive materials including radon. We selected the most recent one if some papers were based on the same cohort. Based on the selected papers, we summarized the lung cancer risks from silica, silicosis and non-silicosis with exposure to silica, by meta-analysis using a random effects model. The pooled relative risks were 1.32 (95% confidence interval (CI), 1.23-1.41) for silica, 2.37 (95% CI, 1.98-2.84) for silicosis and 0.96 (95% CI, 0.81-1.15) for non-silicosis with exposure to silica. Since some papers on silica did not exclude silicosis, the risk due to silica itself may be smaller than 1.32. It was less possible that silica exposure directly increases lung cancer risk. On the other hand, the relative risk, 2.37 for silicosis suggested that silicosis increases lung cancer risk. Meta-analysis also revealed that cigarette smoking strongly increased the lung cancer risk in silicotic patients (relative risk, 4.47; 95% CI, 3.17-6.30). Thus, the present study suggested the great importance of preventing silicosis and smoking cessation in reducing lung cancer incidence in silica-exposed workers. PMID- 15295902 TI - CDNA array analysis of gene expression profiles in brain of mice exposed to manganese. AB - This study is performed to detect changes of gene expression in substantia nigra (SN) and striatum in manganese (Mn)-exposed mice brain. The cDNA array is a recently developed molecular biological method that can detect the differential expression of several hundreds of genes simultaneously and is therefore advantageous in the study of trace metal intoxication effect at the genetic level. Using this technology, we discovered 5 genes in the mouse striatum and 9 genes in SN changed by more than 50% following Mn exposure. Depression were observed in two genes (neural cell adhesion protein BIG2, heavy neurofilament subunit genes) in striatum and three genes (light neurofilament subunit, brain acyl-CoA synthetase II, heavy neurofilament subunit genes) in the SN. However three genes (N-acetylglucosaminyltransferase I, S100beta, and synaptonemal complex protein I genes) in striatum and six genes (noggin, striatin, Ost oncogene, S100beta, calcium/calmodulin-dependent protein kinase kinase beta, and N-acetylglucosaminyltransferase I genes) in SN were elevated following Mn exposure. Immunohistochemical study revealed that protein levels of S100beta also increased following Mn treatment. Activated astrocytes overexpressing S100beta are invariably and intimately associated with decreased expression of heavy and light neurofilament subunits which is a distinguishing feature of neurodegeneration by Mn exposure. All our findings suggested that neuronal degenerations occur in SN as well as striatum of mice exposed to Mn. PMID- 15295903 TI - Cancer risk assessment of toxaphene. AB - The primary purpose is to do cancer risk assessment of toxaphene by using four steps of risk assessment proposed by the United States National Academy of Sciences/National Research Council (NAS/NRC). Four steps of risk assessment including hazard identification, dose-response relationship, exposure assessment, and risk characterization were used to evaluate cancer risk of toxaphene. Toxaphene was the most heavily used insecticide in many parts of the world before it was banned in 1982. It increased incidence of neoplasms of liver and uterus in mice and increased incidence of neoplasms of endocrine organs, thyroid, pituitary, adrenal, mammary glands, and reproductive systems in rats. From mice's and rats' study, slope factor for toxaphene is 0.8557 (mg/ kg/day)(-1). Lifetime average daily dose (LADD) of toxaphene from ambient air, surface water, soil, and fish were 1.08 x 10(-6), 5.71 x 10(-6), 3.43 x 10(-7), and 7.96 x 10(-5) mg/kg/day, respectively. Cancer risk of toxaphene for average exposure is 7.42 x 10(-5). From this study, toxaphene might have carcinogenic risk among humans. PMID- 15295904 TI - Differences in mortality rates due to major specific causes between Japanese male occupational groups over a recent 30-year period. AB - It is assumed that differences in the mortality rates of occupational groups are explained by work-related factors, socioeconomic status, and health practices, etc. The present study focuses on the common factors contributing to differences in the mortality rates from all and major specific causes among Japanese male occupational groups. With respect to mortality rates, the following conditions were adopted as major specific causes of death: cerebrovascular disease (CVD), ischemic heart disease (IHD), stomach cancer (Stomach CA), lung cancer (Lung CA) and suicide. Occupations were classified into eight groups. Age-adjusted mortality rates due to each specific cause of death were calculated, using the age-specific population in 1985 as a standard, for every five years of census from 1965 until 1995. The number of significant correlation coefficients and their magnitude between mortality rates due to major specific causes, among the eight occupational groups, increased with advancing census year. Namely, the order of mortality rates for the major causes in Japanese male occupational groups became more similar over the recent 30 yr period. According to the principal component analysis of mortality rates due to major specific causes, the first main factor contributed 57.9% of the commonality in 1965, 76.5% in 1980, and 86.0% in 1995, respectively. PMID- 15295905 TI - Brain microdialysis study of the effects of hazardous chemicals on the central nervous system 2. Toluene exposure and cerebral acetylcholine. AB - The microdialysis technique was applied to detect the changes in the activity of acetylcholine (ACh) neurons in the rat brain. The effects of intraperitoneal (i.p.) injection of toluene on the amount of ACh release from the nerve terminals of the brain cholinergic neurons were investigated in freely moving rats. In the striatum, injection of toluene decreased the extracellular concentration of ACh in a dose dependent manner in the range 200 to 2,000 mg/kg. Similar effects of toluene on ACh release were observed in the hippocampus after i.p. administration. The increases in ACh content in brain homogenate after i.p. injection of toluene seemed to be caused by the decreased release of ACh from cholinergic nerve endings. Injection of toluene at doses higher than 200 mg/kg decreased ACh release and a similar decrease was suggested to occur in 8-h inhalation exposure to toluene at 1,000 ppm or higher concentrations. PMID- 15295906 TI - Assessment of smoking status among workers using an improved colorimetric method. AB - To monitor smoking status among workers, we improved the colorimetric method to detect cotinine and other nicotine metabolites in urine. In adding ethanol in the reaction mixture, the quantitative measurable time defined as the duration with more than 95% of the peak absorbance, extended to 80 min in contrast to 16 min in the original method. As the analytical condition, a aliquot of urine sample (0.5 ml) was mixed with 0.5 ml of ethanol, with 0.2 ml of 4 M acetate buffer (pH: 4.7), with 0.1 ml of 150 mM KCN, with 0.1 ml of 0.44 M chloramin and then with 0.5 ml of 78 mM barbituric acid. A linear relationship was observed between cotinine concentration up to 80 microM and absorbance at 508 nm (r=0.998, p<0.01). Mean levels of nicotine metabolites among non smokers and smokers were 4.9 and 47.4 microM cotinine equivalent, respectively. Sensitivity and specificity were 96.1% and 96.7%, respectively for nicotine metabolites concentration among workers (n=385) when adopting 6.9 microM cotinine equivalent as a cut off value and the area under the ROC curve was 0.982. This method can be applicable to quantitative detection of smoking status. PMID- 15295907 TI - Occupations and Parkinson's disease: a case-control study in South Korea. AB - We performed a hospital based case-control study in the southeast region of Korea to clarify the role of occupational exposure, especially manganese (Mn), in the etiology of Parkinson's disease (PD) and to discover the association between any occupation and PD. 105 outpatients with PD and 129 neurological disease controls and 101 healthy controls were interviewed. We employed occupational and industrial categories as defined by Section (the most broad category) and Division (sub-category) of the Korea Standard Industry Code and the Korea Standard Classification of Occupations. There was not a significant association between exposure to hazardous materials, especially Mn and PD. There were not any occupations listed under the Section of Industry Classification as a significant risk factor or protective factor for PD. However, the 'clerk' occupation [Section] was positively associated with PD. There is a decreased risk for PD with a subject ever having worked in the 'agriculture, forestry and fishery' occupational group. Ever having worked in 'sales' also was negatively associated with PD. There were not any Divisions of Industry found as a significant risk factor or protective factor for PD. However, ever having worked in an 'agriculture' Division of Occupation was negatively associated with PD. PMID- 15295908 TI - Age-related change in relationship between white blood cell count and some features of the metabolic syndrome. AB - To examine age-related change in the strength of the association of white blood cell count (WBC) with features of the metabolic syndrome (MS), body mass index, blood pressure, total cholesterol, high-density lipoprotein (HDL) cholesterol, triglycerides, fasting plasma glucose, and uric acid were assessed as the components of the MS in 5,218 Japanese male office workers aged 23-59 yr. The subjects were stratified by age into three groups of 23-39, 40-49, and 50-59 yr. WBC count showed a positive crude correlation with the components of the MS, except for HDL cholesterol (negative), in all three age groups. With an increase in age, an association between WBC count and each component of the MS declined, and an interaction with age weakened. After controlling for potential confounders, the largest differences of WBC count for each categorized feature of the MS were found among those aged 23-39 yr. Stratified analyses by smoking status and age showed that in both non-smokers and current smokers the adjusted WBC count increased as the number of features of the MS increased in all three age groups. In each category of the number of clustered features of the MS, differences of WBC count compared with the presence of no features of the MS were the largest in those aged 23-39 yr in both non-smokers and current smokers. The adjusted odds ratios of > or =3 features of the MS also increased with an increase in WBC count in all three age groups in both non-smokers and current smokers. The adjusted odds ratios of > or =3 features of the MS across quartiles of WBC count (lowest to highest) were the largest in the youngest age group. Our results indicate that a variety of features of the MS are associated with WBC count and that these tendencies are more pronounced in younger individuals in both non-smokers and current smokers. PMID- 15295909 TI - Lifting strengths in different exertion heights conditioned on extended legs. AB - This study recruited seven height-matched healthy males to examine their maximum isometric lifting strengths across 13 exertion heights, ranging from 25 cm to 133 cm in increment of 9 cm. The results showed a nonlinear (increasing-decreasing increasing) strength-height relationship for all subjects. The subjects' lifting strength was strongest (mean 1253.2 N) at the exertion height of 61 cm and weakest (mean 454.1 N) at the exertion height of 115 cm. Due to a large variability of strength ratio for the weakest individual strength to the strongest individual strength across the 13 exertion heights, ranging from 59.6% to 83.7%, practitioners should be cautious when assessing workers' lifting capacity based on strength testing. PMID- 15295910 TI - Ergonomic risk factors of work processes in the semiconductor industry in Peninsular Malaysia. AB - A cross-sectional survey of semiconductor factories was conducted to identify the ergonomic risk factors in the work processes, the prevalence of body pain among workers, and the relationship between body pain and work processes. A total of 906 women semiconductor workers took part in the study. In wafer preparation and polishing, a combination of lifting weights and prolonged standing might have led to high pain prevalences in the low back (35.0% wafer preparation, 41.7% wafer polishing) and lower limbs (90.0% wafer preparation, 66.7% wafer polishing). Semiconductor front of line workers, who mostly walked around to operate machines in clean rooms, had the lowest prevalences of body pain. Semiconductor assembly middle of line workers, especially the molding workers, who did frequent lifting, had high pain prevalences in the neck/shoulders (54.8%) and upper back (43.5 %). In the semiconductor assembly end of line work section, chip inspection workers who were exposed to prolonged sitting without back support had high prevalences of neck/shoulder (62.2%) and upper back pain (50.0%), while chip testing workers who had to climb steps to load units had a high prevalence of lower limb pain (68.0%). Workers in the assembly of electronic components, carrying out repetitive tasks with hands and fingers, and standing in awkward postures had high pain prevalences in the neck/shoulders (61.5%), arms (38.5%), and hands/wrists (30.8%). PMID- 15295911 TI - Occupational dermatitis from soldering flux. AB - The aim of this study was to determine the specific agent responsible for eczema on the forearms of 2 electronic assemblers who cleaned out a flux-spraying unit once a week. Soldering flux can be a source of skin irritation as well as allergy. Patch test with dried flux residue (as is) and rosin in dilution series of 20%, 10%, and 1% in olive oil was performed. Readings were taken on day 2 and day 3. The rosin in dilution series was negative; however, the flux residue gave a + reaction on day 2, and by day 3 the reaction had weakened (+?). Similar results were obtained in 2 unexposed controls. Patch tests results in our cases indicate that the flux used in the soldering process caused irritant contact dermatitis. PMID- 15295912 TI - Generalized eruption accompanied by hepatitis in two Thai metal cleaners exposed to trichloroethylene. AB - Two female workers, aged 23 and 24, engaged in cleaning metal straps with trichloroethylene (TCE) in a watch manufacturing plant, experienced generalized eruption, mucosal lesion, fever and hepatitis. The first case suffered fulminant hepatitis and died from liver failure in two weeks after the first symptom appearance. The second case, whose onset of generalized eruption, mucosal lesion and hepatitis without jaundice was nine days after that of the first case, however, recovered in 2 wk. Because the result of working environment measurement suggested heavy exposure to TCE, we deemed that there would be a causal relationship between TCE exposure and the illness. Although there have been considerable number of papers describing the above-mentioned relationship, the fact is not well recognized even among medical personnel in Thailand. Taking the wide use of TCE into account, the prevention of this illness would be very important especially in rapidly industrializing countries. PMID- 15295913 TI - Complementary & alternative mental health treatments. PMID- 15295914 TI - Lighting solutions for contemporary problems of older adults. PMID- 15295915 TI - Mindfulness meditation: a path of transformation & healing. AB - As nurses, we have the unique privilege of witnessing and nurturing the healing process of the whole person--mind, body, and spirit. Teaching mindfulness meditation is a nursing intervention that can foster healing. The consistent practice of mindfulness meditation has been shown to decrease the subjective experience of pain and stress in a variety of research settings. Formal and informal daily practice fosters development of a profound inner calmness and nonreactivity of the mind, allowing individuals to face, and even embrace, all aspects of daily life, regardless of circumstances. By emphasizing being, not doing, mindfulness meditation provides a way through suffering for patients, families, and staff. This practice allows individuals to become compassionate witnesses to their own experiences, to avoid making premature decisions, and to be open to new possibilities, transformation, and healing. PMID- 15295916 TI - Homeopathy: natural mind-body healing. AB - 1. Homeopathy is an accepted form of health care in many countries worldwide. 2. Homeopathy is deeply healing, yet does so without causing side effects, tolerance, or addiction problems. 3. Homeopathic remedies are available over the counter in health food stores and some pharmacies. 4. Homeopathy is distinct from herbal medicine and does not interact with conventional treatments. PMID- 15295917 TI - Spirituality and mental health clients. AB - Spirituality is an important part of human existence but is often overlooked in the conceptualization of the person as a biopsychosocial entity. This article examines spirituality as a concept, relates it to the experience of mental health clients, proposes spiritual assessments and interventions within the role of advanced practice mental health nurses, and discusses the necessity of including spiritual interventions to support healing and wholeness for mental health clients. PMID- 15295918 TI - Sturge-Weber syndrome. AB - Sturge-Weber syndrome is a sporadic neurocutaneous disease characterized by facial port-wine stain, ocular abnormalities (glaucoma and choroidal hemangioma) and leptomeningeal angioma. Although the precise pathogenesis is unknown, available data regarding genetics, embryogenesis, and pathologic features are briefly reviewed. Clinical features vary from mild incomplete forms to full-blown disease with facial stain, seizures, and glaucoma. Frequencies of associated complications are reviewed. To plan treatment and further follow-up, diagnosis of glaucoma and intracranial involvement, even if asymptomatic, is fundamental in children at risk. Early neuroimaging features are important to recognize. Management of patients with Sturge-Weber syndrome is focused on treating associated neurologic and ocular abnormalities. PMID- 15295919 TI - Cutis marmorata telangiectatica congenita. AB - Cutis marmorata telangiectatica congenita is an uncommon vascular malformation composed of capillary and venous sized vessels. It presents with a distinct reticulated pattern that is reminiscent of physiologic cutis marmorata however skin lesions do not resolve with warming of the skin surface. It may have a localized or generalized pattern on the skin. Associated anomalies occur in individuals with cutis marmorata telangiectatica congenita the most commonly reported are limb asymmetry and the coexistence of other vascular birthmarks. Adams Oliver Syndrome and cutis marmorata telangiectatica congenital-macrocephaly syndrome are rare disorders that are associated with cutis marmorata telangiectatica congenita. PMID- 15295920 TI - Potential complications of segmental hemangiomas of infancy. AB - Although the majority of hemangiomas of infancy can be expected to follow a benign course, a significant subset may result in serious complications. Recently, hemangiomas of segmental morphology, or those which are large, plaque like, and patterned in distribution, have been recognized as important markers for potential complications. PHACE syndrome represents the best known example of the variety of problems that can occur in this setting. The PHACE acronym, which stands for posterior fossa brain malformations, segmental cervicofacial hemangiomas, arterial anomalies, cardiac defects and coarctation of the aorta, and eye anomalies, is sometimes referred to as PHACE(S) when ventral developmental defects such as sternal clefting and supraumbilical raphe are present. This article reviews the specific manifestations of PHACE, reflects on pathogenesis, and discusses appropriate work-up and future directions for this complex and fascinating syndrome. We also discuss other complications associated with hemangiomas of segmental morphology, including ulceration, potential visceral involvement, and underlying anomalies related to the lumbosacral location. PMID- 15295921 TI - Incontinentia pigmenti: a window to the role of NF-kappaB function. AB - Incontinentia pigmenti is an uncommon X-linked dominant genodermatosis primarily affecting females. Its hallmark is a unique skin eruption that presents in infancy along the lines of Blaschko and evolves through four stages: inflammatory, verrucous, hyperpigmented, and atrophic. Other persistent findings of the disease include alopecia and dental anomalies. In a minority of cases, serious ophthalmologic and neurological alterations may occur. Mutations in the NF-kappaB essential modulator (NEMO) that lead to an inability to activate the NF kappaB pathway produce IP. Less deleterious mutations in NF-kappaB essential modulator give rise to hypohidrotic ectodermal dysplasia with immune deficiency in affected males, a related but distinct phenotype. These recent discoveries provide insight into the crucial role of NF-kappaB function in regulating the developmental, inflammatory, immune, and anti-apoptotic responses of the skin and other organs. PMID- 15295922 TI - Cutaneous signs of neural tube malformations. PMID- 15295923 TI - Neurocutaneous melanosis. AB - Neurocutaneous melanosis (NCM) is a rare congenital disorder characterized by the presence of large or multiple congenital melanocytic nevi in association with benign or malignant proliferation of melanocytes in the leptomeninges. NCM is believed to occur as a consequence of an error in morphogenesis in neural ectoderm in the developing embryo. Animal models suggest that aberrant expression of the hepatocyte growth factor/scatter factor (HGF/SF) may be involved in the pathogenesis in the NCM. While the majority of patients with NCM have large congenital melanocytic nevi in a posterior axial distribution, a significant proportion of patients present with multiple smaller nevi in the absence of a single larger lesion. Neurologic manifestations generally occur within the first two years of life, and are often related to increased intracranial pressure. Associated structural anomalies of the CNS have been reported in NCM, particularly the Dandy-Walker complex. The long-term clinical significance of characteristic magnetic resonance findings in neurologically asymptomatic patients is unclear. Approximately half of NCM patients develop CNS melanoma. The prognosis of symptomatic patients remains poor. PMID- 15295924 TI - Epidermal nevus syndromes. AB - The term "epidermal nevus syndrome" (ENS) has been used to describe the association of epidermal hamartomas and extra-cutaneous abnormalities. Epidermal nevi follow the lines of Blaschko. The majority of the extra-cutaneous manifestations involve the brain, eye, and skeletal systems. Several subsets with characteristic features have been delineated including the nevus sebaceous syndrome, Proteus syndrome, CHILD syndrome, Becker nevus syndrome, nevus comedonicus syndrome, and phakomatosis pigmentokeratotica. Epidermal nevi have been associated with benign and malignant neoplasms. Advances in molecular biology have revealed that the manifestations of ENS are due to genomic mosaicism. It is likely that the varied clinical manifestations of ENS are due in great part to the functional effects of specific genetic defects. Optimal management of the patient with ENS involves an interdisciplinary approach. Amelioration of the cutaneous features of ENS has been difficult but there have been advances, especially in the use of lasers. PMID- 15295925 TI - [The effectiveness of follow-up care in tumour patients in the early detection of metachronous second primary tumours after curative treatment of carcinoma of the upper aerodigestive tract]. AB - The effectiveness of screening for metachronous second primary tumours by routine follow-up after curative treatment of carcinoma of the upper aerodigestive tract (UADT) was investigated. Data relating to epidemiology, treatment, follow-up (8,014 consultations), and survival from 819 previously untreated patients were analysed to identify risk factors for second primary tumours and to evaluate their impact on survival. During the follow-up period (median 45.2 months), 10.7% of the patients developed a second carcinoma. The annual incidence, which depended on the localisation of the index tumour, was 1.6% for glottic, 3.0% for oropharyngeal, 3.4% for oral cavity, 3.6% for supraglottic, and 4.6% for hypopharyngeal carcinoma. Male sex and the site of the index tumour were identified as relevant prognostic factors for the time to occurrence of a second carcinoma. Radiotherapy as part of initial treatment was associated with a decreased risk for second tumour. Second primary tumour-specific survival was influenced by age, treatment of the index carcinoma, site and stage of the second carcinoma. Patients with curatively treated UADT cancer should have a long-term follow-up with an annual otolaryngologic examination to screen for second primary tumours. More intensive follow-up strategies do not appear to be beneficial. PMID- 15295926 TI - [Acute otitis media and its life-threatening complications]. AB - Acute otitis media is a widespread disease affecting all ages. The introduction of antibiotics has led to a reduction in the incidence of complications from approx. 17% to 1%. Nevertheless, life-threatening complications still occur. The present paper describes the course and treatment of mastoiditis, petrositis and sepsis, as well as intracranial complications, i.e. meningitis, sigmoid sinus thrombosis, extradural abscess, subdural empyema, brain abscess and otitic hydrocephalus. The importance of antibiotic treatment for acute otitis media and the need to assess treatment outcomes are emphasised. PMID- 15295927 TI - [Early diagnosis of hearing impairment in children]. AB - Hearing loss can be attributed to a variety of causes. In infants early detection of hearing loss coupled with an appropriate intervention is critical to their speech, language, cognitive and social development. Despite the fact that several methods exist to uncover hearing impairment beyond any doubt, permanent hearing loss in children is still being detected much too late in Germany. This calls for universal new-born hearing screening and adequate audiometric testing in children that their parents either suspect may suffer from a hearing problem or have other risk factors. PMID- 15295928 TI - [Minimally invasive surgery of the paranasal sinuses]. AB - Surgery of the paranasal sinuses for chronic sinusitis or nasal polyposis has changed profoundly over the last decades. Today, minimally invasive surgery through the nostrils is the standard of care throughout the world. Classic procedures using an extranasal approach have been abandoned almost completely. This development was mainly driven by the introduction of rigid telescopes providing an angled view of the operation field. In addition, new pathophysiological findings have suggested less radical procedures. Functional endoscopic sinus surgery (FESS) shows high success rates, complications are rare, but potentially serious. Computer-assisted surgery systems for intraoperative navigation are at the threshold of being used on a routine basis. However, at the present time their use must not be considered as mandatory for the vast majority of cases. Postsurgical treatment using nasal irrigations with normal saline solution and topical application of steroids is of great importance to provide long-term success following surgical interventions. PMID- 15295929 TI - [Testing the sense of smell using validated procedures]. AB - There is a growing interest into the investigation of smell disorders in both research and clinical practice. For psychophysical ("subjective") investigations related to the sense of smell a variety of test kits are available, namely the various "Sniffin' Sticks" kits, the UPSIT (University of Pennsylvania Smell Identification Test) or the CCSIT (Cross-Cultural Smell Identification Test). Recording of olfactory evoked potentials (OEPs) and respiration olfactometry can be used for diagnosing smell dysfunctions in a more objective way. PMID- 15295930 TI - [Testing the sense of taste using validated procedures]. AB - Psychophysical tests related to the sense of taste are usually less complex than those of the sense of smell. In addition to global tests a regional examination of the sense of taste might be indicated to search for nerve lesions. There are numerous validated psychophysical tests available based on the administration of liquid or solid substrates. The use of electrical currents (electrogustometry) is especially feasible for a rapid regional testing. The registration of gustatory event-related potentials to objectify results from psychophysical examination is possible although limited to a few centers world-wide. PMID- 15295931 TI - [Teaching courses on aspects of medical history taking and communication skills in Germany: a survey among students of 12 medical faculties]. AB - BACKGROUND: Good communication between patients and doctors has positive effects on health and the patients' quality of life. Communication skills can be trained. In many countries communication skills training is an important part of medical education and continuing medical education. In this study German medical students were questioned about current communication training. METHODS: Questionnaires were sent to 28 Medical Schools in Germany and distributed in General Practice courses. Using Likert scales students were asked to rate both existing teaching courses on communication skills and their ability to communicate. RESULTS: 377 students of 12 Medical Schools participated in this study. Two Medical Schools offer teaching courses on communication skills as part of their regular curriculum. On a scale ranging from 1 (no such courses available) to 7 (courses fully available) students assessed the practical teaching of communication skills to be 3 (median). In addition, on a scale ranging from 1 to 7 students rated their general communication skills as 3 (median) and their ability of taking a sexual history and breaking bad news as 4 (median). CONCLUSION: Although these results are not representative, they give a general idea of communication skills teaching in Germany. During their clinical education students should be especially trained for difficult situations in the patient-doctor encounter. The international experience of other Medical Schools should be taken into account when implementing communication skills training as part of medical education. PMID- 15295932 TI - [Evidence-based medicine from the office-based physicians' point of view: a representative survey on its acceptance and demand for continuous medical education]. AB - In 2002 a curriculum on evidence-based medicine (EbM) has been developed and published in Germany on behalf of the national statutory health care administrations. Planning and guiding medical education in EbM requires an empirically-based knowledge of the level of awareness and acceptance of EbM as well as specific educational needs and organisational preferences. In Germany, there is no such systematic knowledge, particularly in regard to general practitioners. A random sample of 900 office-based physicians in the German state of Schleswig-Holstein, stratified by medical specialties, was surveyed by means of a postal questionnaire. After two reminders the response rate was 65.6%. Respondents mainly welcomed EbM, in terms of their own daily work, for the benefit of the patient and the healthcare system as a whole. Integrating EbM into their daily practice and possible restrictions to therapeutic freedom were considered the most serious problems of EbM. 46.8% of the respondents were interested in EbM training. They preferred a rather low-cost 2-day weekend seminar. Office-based physicians were found to be sufficiently motivated to participate in EbM courses, which is in marked contrast to the actual participation rate being rather low in Schleswig-Holstein. The training should primarily focus on integrating EbM into daily routines. In public presentations efforts should be directed towards promoting the way in which EbM enhances professional autonomy and therapeutic freedom. PMID- 15295933 TI - [Patient survey in general practice: development, validation and application of a survey instrument]. AB - Patient surveys are one of the most important sources for quality management in ambulatory care. The quality assurance of the survey instrument itself is of outstanding importance, since only a high-quality instrument is able to supply high-quality data, which can serve as a reliable basis for improvement strategies. The present paper outlines the process of development and optimisation of a survey instrument for patients' rating in all sectors of general practice and primary care on the basis of one of the largest surveys of this kind that have so far been conducted in German-speaking countries. The technique presented here of building quality dimensions of care out of single items has been methodologically validated. On the one hand it allows rapid identification of critical areas at a glance, and on the other hand helps to focus--if necessary--on single aspects in order to define detailed improvement actions. PMID- 15295934 TI - [From the German Society of Quality Assurance: Correction of the nasal septum]. PMID- 15295935 TI - [From the German Society of Quality Assurance: tonsillectomy]. PMID- 15295936 TI - [Geriatric eye diseases]. AB - Ageing of the eye affects all ocular structures. Presbyopia is an early manifestation of physiological ageing resulting in the impossibility to read without additional spectacle correction. Some systemic disorders may interfere with the physiological ageing of the eye and cause blindness. In the western countries blindness is caused by age-related macular degeneration (26%), glaucoma (20.5%), cataract (11.2%), diabetic retinopathy (8.9%) and ischemic optic neuropathy (4 %). PMID- 15295937 TI - [Case report: acute unilateral loss of visual acuity after a visit to the dentist: an unusual complication after the use of an anesthetic combined with adrenaline]. AB - Occurrence of acute and unilateral blindness after local anaesthesia combined with adrenaline, for the treatment of dental caries. The blindness was caused by vasospasm of the central retinal artery. The dentists should be warned about possible visual complaints after use of local anaesthesia, which should urge them to refer the patient to the ophtalmologist. PMID- 15295938 TI - [Results of penetrating keratoplasty in syphilitic interstitial keratitis]. AB - PURPOSE: To compare the results in our patient series after penetrating keratoplasty (PKP) for syphilitic interstitial keratitis (IK) with those described in the literature. METHODS: Retrospective case series in which visual acuity (VA), graft clarity, rejection episodes, intraocular pressure and endothelial cell density (ECD) were examined postoperatively. RESULTS: Postoperative VA improved in all cases. There was no evidence of wound dehiscense or occurrence of retrocorneal membrane formation in any case. Postoperative inflammation was not more severe in our patients with syphilitic IK compared to patients undergoing PKP for other reasons. A normal decline in ECD proved that there was no subclinical inflammation as well. CONCLUSION: PKP for syphilitic IK has a good prognosis in our case series as far as graft survival is concerned. Improvement in VA was present in all cases, though sometimes limited. In our case series, we experienced less postoperative complications than described in the older literature, which is probably due to better microsurgical techniques used nowadays. PMID- 15295939 TI - [The choice of color in fixed prosthetics: what steps should be followed for a reliable outcome?]. AB - The creation of a perfectly-matched esthetic fixed restoration is undeniably one of the most difficult challenges in modern dentistry. The final outcome depends on several essential steps: the use of an appropriate light source, the accurate analysis and correct evaluation of patient's teeth parameters (morphology, colour, surface texture,...), the clear and precise transmission of this data to the laboratory and the sound interpretation of it by a dental technician who masters esthetic prosthetic techniques perfectly. The purpose of this paper was to give a reproducible clinical method to the practitioner in order to achieve a reliable dental colorimetric analysis. PMID- 15295940 TI - [Working with magnification in dental medicine: useful or indispensible?]. AB - In this article, the author describes the different types of magnifying-glasses which can be used in a dental practice. He informs about his own experience in using these glasses in his daily practice. PMID- 15295941 TI - [The microscopy in dental medicine: gadget or necessity?]. AB - In recent years full-blown technical revolutions have changed the face of dentistry. The introduction of the operating microscope in dental surgery was not among the slightest ones. The article wants to show what a brilliant instrument the surgical microscope turned out to be for the dental practice. The microscope offers a homogeneous illumination of the surgical field, without shadows, quite the contrary, with a lot of contrast and a great 3D effect. It provides a clear view throughout treatment (in marked contrast with endoscopes). Thanks to the view optimization by means of the intense light and the magnification, technical precision is very much enhanced during ever expanding therapeutic procedures. PMID- 15295942 TI - Relationship of periodontal disease and tooth loss to prevalence of coronary heart disease. AB - BACKGROUND: Studies relating periodontal disease to coronary heart disease (CHD) have provided equivocal results using tooth loss and/or clinical signs of periodontal disease as measures of periodontal exposure. METHODS: The purpose of this cross-sectional study was to evaluate the relationship of tooth loss and periodontitis to prevalent CHD at the Atherosclerosis Risk in Communities (ARIC) visit 4 using both tooth loss and clinical signs of disease in a population-based sample of 8,363 men and women aged 52 to 75 years from four U.S. communities. Each subject participated in a complete periodontal examination, assessment of missing teeth, assessment of prevalent CHD, and a number of laboratory tests and questionnaires. High attachment loss was defined as > or = 10% of sites with attachment loss > 3 mm and high tooth loss was defined as fewer than 17 remaining teeth. RESULTS: Individuals with both high attachment loss and high tooth loss (odds ratio [OR] 1.5, 95% confidence interval [CI] 1.1 to 2.0) and edentulous individuals (OR 1.8, CI 1.4 to 2.4) had elevated odds of prevalent CHD compared to individuals with low attachment loss and low tooth loss, while controlling for a number of traditional risk factors for CHD. CONCLUSIONS: These results suggest that tooth loss and periodontal disease are associated with prevalent CHD, but only when both are present. The weaker relationships between periodontal disease and CHD that have been found among older adults may be due to older adults having fewer teeth. Future longitudinal studies should be designed to ascertain the cause of tooth loss during follow-up. PMID- 15295943 TI - Relationship between electrocardiographic abnormalities and periodontal disease: the Hisayama Study. AB - BACKGROUND: Recent studies have suggested a relationship between periodontitis and cardiovascular disease (CVD). This study investigated the relationship between periodontitis and electrocardiographic (ECG) abnormalities, which are known predictors of CVD. METHODS: We examined the periodontal status of 1,111 residents of Hisayama Town, Fukuoka, Japan. Nine hundred fifty-seven (957) subjects (374 males, 583 females) with > or = 10 teeth and without a medical history of CVD were included in the analysis. Probing depth (PD) and clinical attachment level (CAL) were measured on two randomly selected quadrants, one maxillary and one mandibular. A 12-lead ECG was recorded using a standard electrocardiograph. ECG abnormalities included left ventricular hypertrophy (Minnesota code 3-1) and ST depression (4-1, 2, 3). The relation of periodontal condition and ECG abnormalities was assessed with logistic regression analysis. RESULTS: Univariate analysis revealed that mean probing depth, mean attachment loss, number of teeth, and plaque index were significantly associated with ECG abnormalities, as well as with known risk factors of CVD. In multivariate analysis, the subjects with deep pockets (mean probing depth > or = 2 mm) had an increased risk for ECG abnormalities (odds ratio [OR] = 1.6; 95% confidence interval [CI] = 1.01 to 2.50) compared to the subjects with mean PD < 2 mm. Subjects with severe attachment loss (mean CAL > or = 2.5 mm) had also significant risk for ECG abnormalities (OR = 1.7; 95% CI = 1.07 to 2.67) compared to those whose mean CAL was < 2.5 mm. CONCLUSION: This study clearly shows the relationship between periodontitis and ECG abnormalities, which are important predictors of CVD. PMID- 15295944 TI - A targeted review of study outcomes with short (< or = 7 mm) endosseous dental implants placed in partially edentulous patients. AB - BACKGROUND: Generally, threaded root-form endosseous dental implants are thought to perform poorly in short lengths (i.e., < 10 mm). However, whether modifications in implant surface geometry will improve performance of short threaded implants is less clear. METHODS: The relationship between dental implant failure rates and their surface geometry, length, and location (maxilla versus mandible) was explored in the published literature. Using a MEDLINE search (1985 through 2001), studies were sought with the following criteria: 1) data suitable to calculate failure rates of implant lengths < or = 7 mm versus > 7 mm; 2) data separable into maxillary versus mandibular results; 3) criteria for "failure" clearly defined; and 4) minimal functional period of 2 years. RESULTS: Twelve papers were identified as follows: eight with machined threaded implants, two with acid-treated threaded implants, and two with sintered porous-surfaced press fit implants. The following results were found: 1) machined surface implants experienced greater failure rates than textured surface implants; 2) with the exception of sintered porous-surfaced implants, 7 mm long dental implants appear to have higher failure rates than those > 7 mm length; and 3) with textured surface implants, higher failure rates were more likely in the maxilla than in the mandible, but with machined surface implants there were no differences in failure rates between maxilla and mandible. CONCLUSIONS: Dental implant surface geometry is a major determinant in how well these implants perform in short lengths, defined here as lengths of < or = 7 mm. While threaded implants show higher failure rates in short versus longer lengths, sintered porous-surfaced implants perform well in the defined "short" lengths. More studies are needed to better assess the performance of short, acid-washed threaded implants. PMID- 15295945 TI - The effects of ovulation induction during infertility treatment on gingival inflammation. AB - BACKGROUND: Ovulation induction is the most common method of infertility treatment in which the ovaries are stimulated to produce multiple follicles. The aim of this study was to assess the effects of three drug protocols of ovulation induction: clomiphene citrate (CC) alone, CC combined with follicle stimulating hormone (FSH), and CC combined with human menopausal gonadotropin (HMG) on the gingival tissues of women who were undergoing infertility treatment. METHODS: Study population was composed of 18 women using CC for three menstrual cycles or less and 16 women using CC for more than three cycles; 21 women using CC-FSH; and 24 women using CC-HMG who had at least four cycles of CC alone the previous year. All subjects were clinically examined for plaque levels (plaque index), gingival inflammation (gingival index), bleeding on probing, and gingival crevicular fluid volume. The results were compared with a control group of 20 women matched for age, educational and professional level, and oral habits and who had never used ovulation drugs. RESULTS: Despite similar plaque levels (P>0.05), women using CC for more than three cycles and combined protocols of CC-FSH and CC-HMG had higher levels of gingival inflammation (P<0.01, P<0.001 and P<0.001, respectively), bleeding (P<0.001), and GCF volume (P<0.001) when compared to the control group and to the users of CC for three cycles or less. CONCLUSIONS: The results of this study have shown that ovulation induction, which is the most common method in the management of infertility, exacerbates gingival inflammation, bleeding, and GCF volume and that the duration of the usage of these drugs is strongly associated with the severity of gingival inflammation. PMID- 15295946 TI - Estrogen and/or calcium plus vitamin D increase mandibular bone mass. AB - BACKGROUND: We have previously reported that estrogen/hormone replacement therapy (E/HRT) has beneficial effects on oral bone density over 3 years and that calcium and vitamin D supplementation has a lesser effect. Here we report on mandibular bone mass for 49 women (of the original cohort of 135) who continued in an additional 2-year, open-label extension. METHODS: Postmenopausal women were randomly assigned to receive calcium and vitamin D plus E/HRT, or calcium and vitamin D only. Regression analysis of mandibular bone mass over time was performed for each woman. RESULTS: Twenty-two of 26 women who took calcium and vitamin D plus E/HRT for 5 years had small mandibular bone mass increases (0.35 +/- 0.38%, P<0.001). Seventeen of 19 women who took only calcium and vitamin D for 3 years had increases in mandibular bone mass (0.74 +/- 0.89%, P<0.002). The largest gains in mandibular bone mass occurred during the first 3 years of the study. CONCLUSIONS: The data of this study indicate that E/HRT and/or calcium and vitamin D may result in increases of mandibular bone mass in postmenopausal women. Because of the long-term risks associated with E/HRT, caution should be exercised in prescribing E/HRT for prevention of chronic menopausal conditions. PMID- 15295947 TI - The impact of ethnicity, gender, and marital status on periodontal and systemic health of older subjects in the Trials to Enhance Elders' Teeth and Oral Health (TEETH). AB - BACKGROUND: Few studies have examined the association between periodontitis risk, gender, and marital status in older adults. The purpose of this study was to assess if the oral health status of older subjects could be explained by differences in: 1) marital status; 2) gender; and 3) ethnicity. METHODS: Clinical and radiographic periodontal oral conditions were studied in 701 older subjects from the TEETH clinical trial. Medical conditions as well as ethnic and marital status and smoking habits were considered. RESULTS: A total of 89 married couples were identified; 40.7% of these were of European descent and 48.1% of Chinese descent. The mean age was 67.7 years (SD +/- 4.7). The men were older than the women (mean difference: 1.5 years, SD +/- 4.6, 95% confidence interval [CI]: 0.5 to 2.5, P<0.01). No significant differences in periodontal conditions were found between spouses or by marital status. Chinese descent was associated with a higher risk for periodontitis, regardless of marital status (odds ratio: 1.5, 95% CI: 1.05 to 2.04, P<0.03). CONCLUSIONS: 1) Married couples have similar social habits, similar oral health perceptions, and similar patterns of periodontal disease. 2) Dental studies including married couples do not bias data for married subjects as such. 3) Marital status has a limited impact on periodontal health but may have a greater impact on several systemic conditions, especially in widowed, divorced, or never married women. 4) Older Chinese subjects perceive themselves as being at lower risk for periodontitis but have more objective signs of periodontitis than older subjects of European descent. PMID- 15295948 TI - Morphometric analysis of the furcation anatomy of mandibular molars. AB - BACKGROUND: Successful treatment of molar furcation defects remains a challenge in clinical practice. Knowledge of anatomic factors facilitates predictable management of furcation involvement lesions. The degree of success in managing furcation involvement is inversely related to the horizontal probing depth. The depth of the horizontal component of attachment loss can vary depending on the external tooth-surface reference points used. However, the anatomical factors affecting horizontal component of attachment loss have not been previously assessed. Therefore, this study determined the bucco-lingual measurements of the cemento-enamel junction and the mesial and distal roots and at the level of root separation. METHODS: One hundred extracted permanent human mandibular first (N = 50) and second (N = 50) molars were studied. Four horizontal bucco-lingual widths were measured with calibrated calipers: 1) furcation entrance/roof (FE); 2) cemento-enamel junction level (CEJ); 3) mesial root width (MRW); and 4) distal root width (DRW). RESULTS: The mean widths at FE, CEJ, MRW, and DRW were, respectively, 5.53 +/- 0.45 mm, 8.71 +/- 0.54 mm, 8.57 +/- 0.54 mm, and 7.97 +/- 0.65 mm in the first molars and 5.61 +/- 0.65 mm, 8.40 +/- 0.65 mm, 7.95 +/- 0.88 mm, and 7.16 +/- 0.84 mm in the second molars. Analysis of variance revealed significant differences between FE and the other variables tested. The results showed that the bucco-lingual width of the furcation roof is considerably shorter than the MRW and DRW. The difference in the mean bucco-lingual dimension between FE and the other measurements occurred in all teeth evaluated and varied between 0.7 and 4.30 mm. CONCLUSIONS: Our findings demonstrate that clinical measurements of horizontal probing depth that use the external surfaces of roots as reference points overestimate the true anatomical component of furcation involvement in mandibular molars. Conversely, positive treatment outcomes in these teeth may be underestimated. This has implications not only for clinical practice but also for clinical research studies evaluating treatment outcomes. PMID- 15295949 TI - Meta-analysis of local metronidazole in the treatment of chronic periodontitis. AB - BACKGROUND: Meta-analysis was used to assess the effectiveness of local delivery of metronidazole alone or as an adjunct to mechanical therapy in patients with chronic periodontitis. METHODS: Studies were identified using MEDLINE and other sources. Meta-analyses were performed on the basis of probing depth (PD) at baseline and experimental and control regimens studied (i.e., metronidazole plus scaling and root planing [SRP] versus SRP and metronidazole versus SRP); the effect of local metronidazole on PD and attachment level (AL) was evaluated for follow-up times of 4, 8, 12, 24, and 36 weeks. The DerSimonian & Laird random effects model was used. RESULTS: Twelve studies that met inclusion criteria were entered into the meta-analysis. A significant mean reduction in PD for the combined metronidazole and SRP was observed in all comparisons with initial PD > or = 4 mm (0.38 mm at 8 weeks to 0.6 mm at 12 weeks); whereas, with initial PD > or = 5 mm a significant mean reduction was observed from 12 weeks (0.29 to 0.48 mm at 24 and 36 weeks, respectively). Meta-analysis could be performed for AL to test the effectiveness of metronidazole as an adjunct to SRP and a significant AL improvement was found in all analyses (0.2 mm at 4 weeks to 0.29 mm at 24 and 36 weeks). Meta-analyses were performed including two to four studies. CONCLUSION: Our results demonstrated the effectiveness of metronidazole as an adjunct to SRP in the treatment of chronic adult periodontitis, but clinical significance and dissemination of antibiotics should be taken into account in the evaluation of metronidazole as an alternative to SRP. PMID- 15295950 TI - Effect of periodontal therapy in smokers and non-smokers with advanced periodontal disease: results after maintenance therapy for a minimum of 5 years. AB - BACKGROUND: Longitudinal clinical studies show smoking is a risk factor for periodontal disease progression. It has also been documented that smoking impairs healing after periodontal therapy. However, the longitudinal effect of smoking on treatment results in patients who undergo long-term maintenance therapy has not been extensively investigated. This study clinically and radiographically compared smoking and non-smoking patients who had been treated for advanced periodontal disease and who received maintenance therapy for a minimum of 5 years. METHODS: Twenty-nine patients were selected over a 6-month period when they presented for a regularly scheduled visit in a private office. Patients were selected on the basis of initially having lost 50% of bone support on 50% of their teeth; had received follow-up therapy for at least 5 years; were compliant at 75% of the appointments; and had plaque scores < 20% in 75% of the visits. All patients had received non-surgical and surgical therapy as required for pocket elimination. Fourteen were active smokers during the entire maintenance period. Clinical measurements of probing depths and presence of plaque and gingivitis and a new set of standardized radiographs were taken. RESULTS: Smokers had higher mean radiographic bone loss values prior to treatment (7.52 +/- 1.39 versus 6.65 +/- 1.39) and at the final examination (7.32 +/- 1.42 versus 6.29 +/- 1.29) mean radiographic bone loss as well as initial, immediate post-therapy, and final percent of pockets > or = 6 mm (1.42% +/- 1.87% versus 0.60% +/- 1.11%). Differences were not statistically significant. Over 5 to 8 years, seven sites in four non-smokers and 11 sites in six smokers exhibited radiographic bone loss > or = 2 mm. One tooth in a non-smoker and three teeth in two smokers were lost. In a logistic regression analysis, smoking increased the odds ratio 10.7 times of having > or = 1 site with bone loss > or = 2 mm. CONCLUSION: The present study on a small group of patients treated for advanced periodontal disease and well maintained over 5 to 8 years showed no statistically significant differences between smokers and non-smokers in clinical probing depth and radiographic bone loss measurements. PMID- 15295951 TI - The association of smoking with vertical periodontal bone loss. AB - BACKGROUND: Knowledge about the relation between smoking and vertical periodontal bone loss is scant. The objective of the present study was to investigate the association between smoking and the prevalence and severity of vertical bone defects. METHODS: Full sets of intraoral radiographs of 249 individuals in 1982 and 229 individuals in 1992 both with an age range 21 to 70 years were assessed with regard to presence or absence of vertical bone defects. A vertical defect was defined as a resorption of the interdental marginal bone > or = 2 mm that had a typical angulation towards either the mesial or distal aspect of the root. RESULTS: The prevalence of vertical defects in 1982 was 47% for current smokers, 49% for former smokers, and 24% for non-smokers. In 1992 the prevalence was 42%, 28%, and 19% for current smokers, former smokers, and non-smokers, respectively. Both in 1982 and 1992 the prevalence was significantly related to smoking status (chi2 = 14.4 and chi2 = 9.9, P<0.01). Furthermore, the severity of vertical defects was significantly associated with smoking after controlling for age both in 1982 and 1992 (P<0.05). The relative risk associated with current smoking was 2.0-fold increased in 1982 and 3.4-fold increased in 1992 (P<0.05). CONCLUSIONS: The observations suggest that smoking is associated with increased levels of prevalence as well as severity of vertical bone loss. Smoking is considered a potential risk factor for vertical periodontal bone loss. PMID- 15295952 TI - The effects of an amine fluoride/stannous fluoride and an antimicrobial host protein mouthrinse on supragingival plaque regrowth. AB - BACKGROUND: Chlorhexidine (CHX)-containing mouthrinses are recommended as adjuncts to mechanical oral hygiene. The problem associated with side effects, however, has stimulated the search for alternative antiplaque agents. The aim of this study was to investigate the plaque inhibitory effects of two mouthrinses containing amine fluoride/stannous fluoride (ASF) and antimicrobial host proteins (lactoperoxidase, lysozyme and lactoferrin; LLL), respectively. METHODS: The study was an observer-masked, randomized 4x4 Latin square cross-over design balanced for carryover effects, involving 12 healthy volunteers in a 4-day plaque regrowth model. A 0.12% CHX mouthrinse and a saline solution served as positive and negative controls, respectively. On day 1, subjects received professional prophylaxis, suspended oral hygiene measures, and commenced rinsing with their allocated rinses. On day 5, subjects were scored for disclosed plaque. RESULTS: The ASF rinse showed a significant inhibition of plaque regrowth in comparison to both saline and LLL solutions, but the lowest plaque indices were obtained with the CHX formulation (P<0.01). There were no significant differences between LLL rinse and saline (P>0.05). Such pattern of efficacy was the same in anterior and posterior teeth and in vestibular and lingual surfaces as well, with the exception of the lingual anterior surfaces. In these sites, differences between the CHX and ASF rinses were not significant (P>0.05). A significantly higher prevalence of side effects was found in subjects using the CHX product (P<0.0042). CONCLUSIONS: Although the effect on plaque regrowth observed with 0.12% CHX rinsing was superior to that with ASF, the ASF rinse was not associated with side effects. These findings, together with those from long-term trials, suggest that the ASF rinse may represent an effective alternative to CHX rinse as an adjunct to oral hygiene. On the contrary, the LLL rinse did not significantly inhibit plaque regrowth. PMID- 15295953 TI - Effect of enamel matrix derivative on periodontal ligament cells in vitro is diminished by Porphyromonas gingivalis. AB - BACKGROUND: Enamel matrix derivative (EMD) has been shown to possess a mitogenic effect to induce effective periodontal regeneration, however, it is unclear whether periodontal pathogens can modulate the effect of EMD. The present study examined the influence of Porphyromonas gingivalis on EMD-stimulated periodontal ligament (PDL) cells. METHODS: P. gingivalis ATCC33277 and its mutants deficient in fimbriae (delta fimA) or gingipains (delta rgpA delta rgpB, delta kgp, and delta rgpA delta rgpB delta kgp) were employed. PDL cells were grown on EMD coated dishes and infected with P. gingivalis wild strain or a mutant. Cell migration and proliferation were then evaluated with an in vitro wound healing assay. The expression of transforming growth factor-beta1 (TGF-beta1) and insulin like growth factor I (IGF-I) mRNA by PDL cells was examined. Further, the degradation and phosphorylation of extracellular signal-regulated kinase 1 and 2 (ERK1/2) as well as paxillin in infected PDL cells were estimated using Western blot analysis. RESULTS: P. gingivalis ATCC33277 inhibited the migration and proliferation of PDL cells on EMD-coated dishes, and the mutants delta fimA, delta rgpA delta rgpB, and delta kgp showed the same effects. Further, each of these organisms diminished the expression of TGF-beta1 and IGF-I mRNA, as well as the phosphorylation of ERK1/2 from EMD-stimulated PDL cells. In addition, total paxillin protein was markedly degraded by both the wild-type strain and each of the mutants except for delta rgpA delta rgpB delta kgp, which showed a negligible effect in all of the assays with EMD-stimulated PDL cells. CONCLUSION: These results suggest that P. gingivalis diminishes the effect of EMD on PDL cells in vitro through a cooperative action of gingipains. PMID- 15295954 TI - Accuracy of supported root ratio estimation from projected length and area using digital radiographs. AB - BACKGROUND: The purpose of this study was to evaluate the accuracy of supported single-root surface ratio estimated from the length and projected area of the tooth, using digital dental radiographs. METHODS: Eight extracted, single-root teeth were three-dimensionally digitized using a contact technique for surface area measurement. The data were then processed using engineering application software and length, projection area, and true surface area of the root at a designated length were obtained. Based on these three measurements, the accuracy of the supported surface area ratio measurement at different lengths of the root was evaluated. RESULTS: The largest mean errors from linear and area estimation were 9.58% and -1.16%, respectively. The 95% confidence intervals were all positive, indicating that linear measurements overestimated supported ratio. T tests showed that linear estimations resulted in significant differences in all eight teeth and area estimations in five teeth. When analyzing the supported ratio of the alveolar bone receding from the cemento-enamel junction (CEJ) toward the apex of the root at each mm, linear estimation showed significant differences down to 8 mm, while area estimation showed significant differences only up to 2 mm. CONCLUSIONS: The results of this study indicate that a reliable estimate of the ratio of root surface area supported by alveolar bone cannot be determined from linear or area data. However, when the marginal bone destruction exceeds 2 mm from the CEJ, area estimation does not show a significant difference in the supported region. As demonstrated in this study, root surface ratio estimation function could be an advantage of digital dental x-ray systems in which projected root area is readily observed. PMID- 15295955 TI - Efficacy of ibuprofen-hydrocodone for the treatment of postoperative pain after periodontal surgery. AB - BACKGROUND: Previous studies have shown that non-steroidal anti-inflammatory drugs (NSAIDs) have significant benefits in the control of postoperative pain after periodontal or oral surgical procedures. The combination of a peripherally acting NSAID with a centrally acting opioid drug is found to be more effective. The purpose of this study was to compare an alternative combination of ibuprofen 400 mg with 5 mg of hydroxycodone to ibuprofen 400 mg used alone in the management of pain following periodontal surgery. METHODS: This study used a double-masked cross-over design with the patients acting as their own controls. Twelve patients underwent two periodontal surgeries in different quadrants of the same dental arch at least 2 weeks apart. A standardized amount of local anesthetic and similar extent and duration of surgery for each side was required. The patients received four doses of medication at predetermined intervals and filled out a visual analog pain scale every 2 hours for the first 12 hours after surgery. RESULTS: The overall pain reported by the patients on visual analog scale was 1.55 (SE +/- 0.16), out of a possible 10. More pain was reported with ibuprofen alone, 1.81 (SE +/- 0.12), compared to the ibuprofen with hydrocodone combination, 1.30 (SE +/- 0.16). The difference was statistically significant (P<0.05). CONCLUSION: The findings suggest that a combination analgesic preparation of ibuprofen (400 mg) with hydrocodone (5 mg) results in better pain control compared to ibuprofen used alone. PMID- 15295956 TI - Polyamines in nickel-titanium archwire-induced gingivitis in adolescents. AB - BACKGROUND: Polyamines spermine, spermidine, and putrescine are involved in a number of inflammatory diseases, but their role in the development of gingivitis and periodontitis has not been fully investigated. The goal of this investigation was to study the levels and the variations of these amines, and the main enzymes related to their metabolism, during archwire orthodontic treatment, a condition which may induce gingivitis. METHODS: Sixty patients (age range: 11 to 27 years) were examined for gingivitis occurring during nickel-titanium (Ni-Ti) archwire orthodontic treatment. Plaque and gingival indexes (PI, GI) as well as salivary polyamine metabolism before the archwire insertion (T0) and at 3 (T1), 6 (T2), and 12 (T3) months of treatment were measured. RESULTS: In patients in the age range of 14 to 17 years, spermine and spermidine, but not putrescine contents, as well as ornithine-decarboxylase (ODC) and S-adenosylmethionine-decarboxylase (SAMDC) activities, significantly rose at 3 months after insertion, without any change in periodontal parameters, and further increased at 6 months reaching the maximum at 12 months. GI increased later, from 6 to 12 months, while PI did not significantly change. Spermidine/spermine-N1-acetyltransferase (SSAT) activity remained unchanged from T0 to T3. On the contrary, in patients whose age was 11 to 13 or over 18 years, no significant variations in polyamine metabolism and periododontal parameters were observed at any examination time. CONCLUSION: These data support the hypothesis that salivary polyamines might be earlier indicators of gingivitis than the gingival index score in adolescents wearing archwire appliances. PMID- 15295957 TI - Relative connective tissue graft size affects root coverage treatment outcome in the envelope procedure. AB - BACKGROUND: This study investigated the correlation between the connective tissue graft size and the percentage of root coverage. METHODS: Fifteen patients with Miller Class I or II recession defects (one tooth/defect per patient) were treated using an envelope connective tissue graft procedure. Clinical parameters including gingival recession depth (RD), clinical attachment level (CAL), keratinized tissue width (KW), and the probing depth (PD) were measured. Visible denuded area (VDA) and graft tissue area (GTA) were indirectly measured in mm2 using computer software. Graft tissue thickness (GTT) and graft tissue width (GTW) were also measured. All clinical parameters and VDA were recorded at baseline and 3 and 6 months. The RD, CAL, KW, PD, and VDA were evaluated by Friedman test. The correlation between the percentage of root coverage (PRC) and factors related to graft size were determined by Spearman rank correlation and nonparametric regression analysis. RESULTS: The percent of root coverage at 3 and 6 months postoperatively was statistically significantly associated with the GTA:VDA ratio (P<0.01); it did not correlate with GTA, GTT, or RD, and was inconsistently correlated to GTW and VDA. In patients who had 100% root coverage, the GTA:VDA ratio ranged between 10.92:1 and 21.95:1; in patients with <100% root coverage, the ratio was between 4.54:1 and 11.06:1. CONCLUSION: The GTA:VDA ratio should be at least 11:1, which is a significant factor for optimal root coverage result in the envelope procedure. PMID- 15295958 TI - Alterations in location of the mucogingival junction 5 years after coronally repositioned flap surgery. AB - BACKGROUND: The aim of this clinical study was to evaluate whether the mucogingival junction (MGJ) reverts back towards its original location following coronally repositioned flap (CRF) procedure over a 5-year follow-up period. METHODS: Thirteen systemically healthy patients with 26 Miller Class I buccal gingival recessions were treated using the CRF technique. At baseline, and 1, 6, 12, and 60 months after surgery, location of gingival margin (LGM), probing depth (PD), clinical attachment level (CAL), width of keratinized gingiua (WKG), and location of mucogingival junction (LMGJ) were recorded. The alterations in the clinical measurements at the different evaluation times were analyzed using the Friedman and the repeated measure analysis of variance (ANOVA) tests, where applicable. Degree of association between continuous variables was calculated by the Pearson's correlation coefficient. RESULTS: The mean percentage of root coverage obtained at the end of 1 month was 68.26% +/- 30.37% (P<0.05) and at 60 months, it was reduced to 44.86% +/- 33.91% (P<0.01). LGM (r=0.592, P<0.001), CAL (r=0.590, P<0.01), WKG (r=0.442, P<0.05), and LMGJ (r=0.653, P<0.001) were found to be significantly correlated with the 60-month postoperative values of LMGJ. At the end of the 60-month follow-up period, the mean apical displacement of LGM was 0.67 +/- 0.72 mm and the same mean apical displacement value for LMGJ was 0.98 +/ 1.19 mm. CONCLUSIONS: Within the limits of this study, the 60-month follow-up findings indicated that the CRF procedure failed to maintain the gingival tissue in a coronal position and that the observed movement of the MGJ back to its original position was partially dependent on the apical movement of gingival margin. PMID- 15295959 TI - Medical grade calcium sulfate hemihydrate in healing of human extraction sockets: clinical and histological observations at 3 months. AB - BACKGROUND: Following tooth extraction, remodeling and resorption of the alveolar bone at the extraction site characterize wound healing. This produces a reduction in ridge volume and difficulties in delayed placement of implants in an ideal position. Medical grade calcium sulfate hemihydrate (MGCSH) has been proposed as a graft material in extraction sockets to minimize the reduction in ridge volume. The aim of the present study was to investigate the influence of MGCSH on the histopathologic pattern of intrasocket regenerated bone and to evaluate histologically the healed MGCSH grafted extraction socket site 3 months postextraction METHODS: MGCSH was grafted in 10 fresh human extraction sockets in 10 patients. Five post-extraction sockets were used as controls. At 3 months a cylindrical tissue specimen, 2.5 mm in diameter, was trephined from the previously grafted site followed by implant placement. Non-decalcified specimens were sectioned at a cross-horizontal plane and stained with fast green, toluidine blue, and Van Kossa stains for histological and histomorphometrical examination. RESULTS: Histologically, MGCSH was not observed in most of the specimens. Newly formed bone with lamellar arrangements was identified in all the horizontal sections with no difference between apical, medium, and coronal areas. The mean trabecular area in the coronal sections was 58.6% +/- 9.2%; in the medium sections, 58.1% +/- 6.2%; and in the apical sections, 58.3% +/- 7.8%. The differences were not statistically significant. CONCLUSION: MGCSH seems to be an ideal graft material in extraction socket bone regeneration because it is almost completely resorbable, and it allows a new trabecular bone arrangement at 3 months. PMID- 15295960 TI - Implant therapy following liver transplantation: clinical and microbiological results after 10 years. AB - BACKGROUND: The number of immunocompromised patients is increasing as a consequence of successful organ transplantation. Placing dental implants in these patients has been questioned because of their increased risk for infections. The 10-year follow-up data of a 71-year-old liver transplant recipient with long-term immunosuppressive therapy is reported. Six months after liver transplantation, two interforaminal implants were inserted in the edentulous mandible, and an overdenture using non-rigid telescopic attachments was fabricated 3 months later. METHODS: Oral clinical parameters included the modified plaque index, sulcus fluid flow rate, modified bleeding index, probing depth, distance implant mucosa, attachment level, width of the keratinized mucosa and mobility values. The distance implant bone (DIB) was determined on digital panoramic radiographs. The levels of eight periodontal marker organisms were established using DNA probe technology. Additionally, swabs of the edentulous oral mucosa were taken for microbiological culture and antimicrobial resistance testing. RESULTS: The peri implant parameters were within normal ranges indicating a stable osseointegration with moderate vertical bone loss. Actinobacillus actinomycetemcomitans, Tannerella forsythensis, Campylobacter rectus, and Treponema denticola were not detected. Low levels of Porphyromonas gingivalis, Fusobacterium nucleatum, and Eikenella corrodens were found. Prevotella intermedia was the only bacterium with a level higher than 10(4). The mucosal swabs indicated the presence of an abundant normal oral flora, including Escherichia coli and Candida albicans. The antibiogram revealed multiple resistance to antibiotics. CONCLUSION: This case report suggests that immunocompromised patients can be successfully rehabilitated with dental implants. PMID- 15295963 TI - The role of high-CF fibers in speech perception: comments on Horwitz et al. (2002). PMID- 15295965 TI - Ultrasonic wave propagation in human cancellous bone: application of Biot theory. AB - Ultrasonic wave propagation in human cancellous bone is considered. Reflection and transmission coefficients are derived for a slab of cancellous bone having an elastic frame using Biot's theory modified by the model of Johnson et al. [J. Fluid Mech. 176, 379-402 (1987)] for viscous exchange between fluid and structure. Numerical simulations of transmitted waves in the time domain are worked out by varying the modified Biot parameters. The variation is applied to the governing parameters and is about 20%. From this study, we can gain an insight into the sensitivity of each physical parameter used in this theory. Some parameters play an important role in slow-wave wave form, such as the viscous characteristic length lambda and pore fluid bulk modulus Kf. However, other parameters play an important role in the fast-wave wave form, such as solid density rhos and shear modulus N. We also note from these simulations that some parameters such as porosity phi, tortuosity alpha(infinty), thickness, solid bulk modulus Ks, and skeletal compressibility frame Kb, play an important role simultaneously in both fast and slow wave forms compared to other parameters which act on the wave form of just one of the two waves. The sensitivity of the modified Biot parameters with respect to the transmitted wave depends strongly on the coupling between the solid and fluid phases of the cancellous bone. Experimental results for slow and fast waves transmitted through human cancellous bone samples are given and compared with theoretical predictions. PMID- 15295969 TI - Spectral estimation for characterization of acoustic aberration. AB - Spectral estimation based on acoustic backscatter from a motionless stochastic medium is described for characterization of aberration in ultrasonic imaging. The underlying assumptions for the estimation are: The correlation length of the medium is short compared to the length of the transmitted acoustic pulse, an isoplanatic region of sufficient size exists around the focal point, and the backscatter can be modeled as an ergodic stochastic process. The motivation for this work is ultrasonic imaging with aberration correction. Measurements were performed using a two-dimensional array system with 80 x 80 transducer elements and an element pitch of 0.6 mm. The f number for the measurements was 1.2 and the center frequency was 3.0 MHz with a 53% bandwidth. Relative phase of aberration was extracted from estimated cross spectra using a robust least-mean-square-error method based on an orthogonal expansion of the phase differences of neighboring wave forms as a function of frequency. Estimates of cross-spectrum phase from measurements of random scattering through a tissue-mimicking aberrator have confidence bands approximately +/- 5 degrees wide. Both phase and magnitude are in good agreement with a reference characterization obtained from a point scatterer. PMID- 15295972 TI - On the feasibility of elastic wave visualization within polymeric solids using magnetic resonance elastography. AB - In this paper, the feasibility of extending previously described magnetic resonance elastography (MRE) dynamic displacement (and associated elasticity) measurement techniques, currently used successfully in tissue, to solid materials which have much higher shear rigidity and much lower nuclear spin densities, is considered. Based on these considerations, the MRE technique is modified in a straightforward manner and used to directly visualize shear wave displacements within two polymeric materials, one of which is relatively stiff. PMID- 15295983 TI - On the acoustic diffraction by the edges of benthic shells. AB - Recent laboratory measurements of acoustic backscattering by individual benthic shells have isolated the edge-diffracted echo from echoes due to the surface of the main body of the shell. The data indicate that the echo near broadside incidence is generally the strongest for all orientations and is due principally to the surface of the main body. At angles well away from broadside, the echo levels are lower and are due primarily to the diffraction from the edge of the shell. The decrease in echo levels from broadside incidence to well off broadside is shown to be reasonably consistent with the decrease in acoustic backscattering from normal incidence to well off normal incidence by a shell-covered seafloor. The results suggest the importance of the edge of the shell in off-normal incidence backscattering by a shell-covered seafloor. Furthermore, when considering bistatic diffraction by edges, there are implications that the edge of the shell (lying on the seafloor) can cause significant scattering in many directions, including at subcritical angles. PMID- 15295984 TI - Tracking sperm whale (Physeter macrocephalus) dive profiles using a towed passive acoustic array. AB - A passive acoustic method is presented for tracking sperm whale dive profiles, using two or three hydrophones deployed as either a vertical or large-aperture towed array. The relative arrival times between the direct and surface-reflected acoustic paths are used to obtain the ranges and depths of animals with respect to the array, provided that the hydrophone depths are independently measured. Besides reducing the number of hydrophones required, exploiting surface reflections simplifies automation of the data processing. Experimental results are shown from 2002 and 2003 cruises in the Gulf of Mexico for two different towed array deployments. The 2002 deployment consisted of two short-aperture towed arrays separated by 170 m, while the 2003 deployment placed an autonomous acoustic recorder in tandem with a short-aperture towed array, and used ship noise to time-align the acoustic data. The resulting dive profiles were independently checked using single-hydrophone localizations, whenever multipath reflections from the ocean bottom could be exploited to effectively create a large-aperture vertical array. This technique may have applications for basic research and for real-time mitigation for seismic airgun surveys. PMID- 15295985 TI - Experimental verification of an interpolation algorithm for improved estimates of animal position. AB - This article presents experimental verification of an interpolation algorithm that was previously proposed in Jaffe [J. Acoust. Soc. Am. 105, 3168-3175 (1999)]. The goal of the algorithm is to improve estimates of both target position and target strength by minimizing a least-squares residual between noise corrupted target measurement data and the output of a model of the sonar's amplitude response to a target at a set of known locations. Although this positional estimator was shown to be a maximum likelihood estimator, in principle, experimental verification was desired because of interest in understanding its true performance. Here, the accuracy of the algorithm is investigated by analyzing the correspondence between a target's true position and the algorithm's estimate. True target position was measured by precise translation of a small test target (bead) or from the analysis of images of fish from a coregistered optical imaging system. Results with the stationary spherical test bead in a high signal-to-noise environment indicate that a large increase in resolution is possible, while results with commercial aquarium fish indicate a smaller increase is obtainable. However, in both experiments the algorithm provides improved estimates of target position over those obtained by simply accepting the angular positions of the sonar beam with maximum output as target position. In addition, increased accuracy in target strength estimation is possible by considering the effects of the sonar beam patterns relative to the interpolated position. A benefit of the algorithm is that it can be applied "ex post facto" to existing data sets from commercial multibeam sonar systems when only the beam intensities have been stored after suitable calibration. PMID- 15295987 TI - On the acoustic vaporization of micrometer-sized droplets. AB - This paper examines the vaporization of individual dodecafluoropentane droplets by the application of single ultrasonic tone bursts. High speed video microscopy was used to monitor droplets in a flow tube, while a focused, single element transducer operating at 3, 4, or 10 MHz was aimed at the intersection of the acoustical and optical beams. A highly dilute droplet emulsion was injected, and individual droplets were positioned in the two foci. Phase transitions of droplets were produced by rarefactional pressures as low as 4 MPa at 3 MHz using single, 3.25 micros tone bursts. During acoustic irradiation, droplets showed dipole-type oscillations along the acoustic axis (average amplitude 1.3 microm, independent of droplet diameter which ranged from 5 to 27 microm). The onset of vaporization was monitored as either spot-like, within the droplet, or homogeneous, throughout the droplet's imaged cross section. Spot-like centers of nucleation were observed solely along the axis lying parallel to the direction of oscillation and centered on the droplet. Smaller droplets required more acoustic intensity for vaporization than larger droplets, which is consistent with other experiments on emulsions. PMID- 15295993 TI - Annoyance with aircraft noise in local recreational areas, contingent on changes in exposure and other context variables. AB - Few socioacoustic studies have examined the effect of noise on outdoor recreationists. The areas studied have been mountain and wilderness areas that people typically travel for a distance to visit. In this article we examine the reactions to aircraft noise in local recreational areas experiencing either decreased (1930 survey respondents), or increased noise exposure (1001 survey respondents). Field studies were conducted before and after the relocation the main airport of Norway in 1998 in one area near each airport. The relationship between individual noise exposure (LAeq for the aircraft events, percentage of time aircraft were audible, and LAsel) for the aircraft events. The analyses included the "situation" in which data were collected (before or after the relocation), and variables describing the recreational context. A strong effect of the "situation" was found in both cases, but the size of the effect was influenced by the choice of exposure variable in one of the study areas. Other context variables were also influencing annoyance. The effect of the situation (before/after a change in exposure) on the dose-response relationship may be influenced by the initial noise levels, the amount of change, and the time elapsed since the change at the time of the second survey. Further research should investigate the significance of these variables. PMID- 15295996 TI - Objective evaluation of chamber-music halls in Europe and Japan. AB - The room acoustical parameters reverberation time, RT; early decay time, EDT; clarity, C80; time gravity, Tg; bass ratio, BR; strength, G; initial time delay gap, ITDG; interaural cross-correlation coefficient, IACC(E), the where binaural quality index BQI equals [1-IACC(E3)]; and stage support, ST1 were measured in 18 major chamber-music halls in Austria, Germany, the Netherlands, Czech Republic, Switzerland, and Japan, employing procedures in accordance with ISO 3382 (1997). In combination with the architectural data, the intrinsic objective parameters for the acoustics of chamber-music halls and their variation range were examined. The results of these studies reveal four pertinent orthogonal parameters: RT, G, ITDG, BQI. General design guidelines for a chamber-music hall are presented. PMID- 15295994 TI - Noise annoyance from stationary sources: relationships with exposure metric day evening-night level (DENL) and their confidence intervals. AB - Relationships between exposure to noise [metric: day-evening-night levels (DENL)] from stationary sources (shunting yards, a seasonal industry, and other industries) and annoyance are presented. Curves are presented for expected annoyance score, the percentage "highly annoyed" (%HA, cutoff at 72 on a scale from 0 to 100), the percentage "annoyed" (%A, cutoff at 50 on a scale from 0 to 100), and the percentage "(at least) a little annoyed" (%LA, cutoff at 28 on a scale from 0 to 100). The estimates of the parameters of the relations are based on the data from a field study (N=1875) at 11 locations (2 shunting yards, 1 seasonal industry, 8 other industries) in the Netherlands. With the same (yearly) DENL, the seasonal industry causes less annoyance than the other industries, while the other industries cause less annoyance than the shunting yards. It appears that annoyance caused by vibrations from shunting yards and annoyance caused by noise from through trains are (partly) responsible for the relatively high annoyance from shunting yards. The relatively low annoyance from the seasonal industry presumably is related to the presence of a relatively quiet period. Results for the two shunting yards and the seasonal industry are based on fewer data than the other industrial sources, and are indicative. The same patterns of influence of age and noise sensitivity that are generally found are also found in this study. For comparison, results regarding transportation sources are also given, including previously unpublished results for expected annoyance. PMID- 15295997 TI - Blind deconvolution of audio-frequency signals using the self-deconvolving data restoration algorithm. AB - A signal processing algorithm has been developed in which a filter function is extracted from degraded data through mathematical operations. The filter function can be used to restore much of the degraded content of the data through use of a deconvolution process. The operation can be performed without prior knowledge of the detection system, a technique known as blind deconvolution. The extraction process, designated self-deconvolving data reconstruction algorithm, is applied here to audio-frequency signals showing significant qualitative improvement. Degradation arising from the process of electronic recording and reproduction is significantly reduced. PMID- 15295999 TI - Accurate analysis of multitone signals using a DFT. AB - Optimum data windows make it possible to determine accurately the amplitude, phase, and frequency of one or more tones (sinusoidal components) in a signal. Procedures presented in this paper can be applied to noisy signals, signals having moderate nonstationarity, and tones close in frequency. They are relevant to many areas of acoustics where sounds are quasistationary. Among these are acoustic probes transmitted through media and natural sounds, such as animal vocalization, speech, and music. The paper includes criteria for multitone FFT block design and an example of application to sound transmission in the atmosphere. PMID- 15296001 TI - Time-frequency analysis of auditory-nerve-fiber and basilar-membrane click responses reveal glide irregularities and non-characteristic-frequency skirts. AB - Although many properties of click responses can be accounted for by a single, frequency-dispersive traveling wave exciting a single, characteristic-frequency (CF) resonance, some properties, such as waxing and waning cannot. Joint time frequency distributions (TFDs) were used to help understand click responses of cat single auditory-nerve (AN) fibers (CFs<4 kHz) and published measurements of chinchilla basilar-membrane (BM) motion. For CFs> 800 Hz, the peak energy of the response decreased in latency and frequency as the level increased, as expected. However, at high levels the trend reversed for AN, but not BM, responses. Normalized TFDs, which show the frequency with the peak energy at each response time, revealed glides, as previously reported. Classical theory predicts smooth, upward glides. Instead, at low CFs there were downward glides, and at other CFs glides had substantial irregularities. Finally, click skirts, defined as the longest-latency part of click responses, sometimes showed deviations from CF for above-threshold sound levels. Most of these phenomena are not explained by a single, frequency-dispersive traveling wave exciting a single CF resonance, but they can be accounted for by the interaction of two (or more) excitation drives with different latencies and frequency contents. PMID- 15296002 TI - Prediction of the characteristics of two types of pressure waves in the cochlea: theoretical considerations. AB - The aim of this study was to predict the characteristics of two types of cochlear pressure waves, so-called fast and slow waves. A two-dimensional finite-element model of the organ of Corti (OC), including fluid-structure interaction with the surrounding lymph fluid, was constructed. The geometry of the OC at the basal turn was determined from morphological measurements of others in the gerbil hemicochlea. As far as mechanical properties of the materials within the OC are concerned, previously determined mechanical properties of portions within the OC were adopted, and unknown mechanical features were determined from the published measurements of static stiffness. Time advance of the fluid-structure scheme was achieved by a staggered approach. Using the model, the magnitude and phase of the fast and slow waves were predicted so as to fit the numerically obtained pressure distribution in the scala tympani with what is known about intracochlear pressure measurement. When the predicted pressure waves were applied to the model, the numerical result of the velocity of the basilar membrane showed good agreement with the experimentally obtained velocity of the basilar membrane documented by others. Thus, the predicted pressure waves appeared to be reliable. Moreover, it was found that the fluid-structure interaction considerably influences the dynamic behavior of the OC at frequencies near the characteristic frequency. PMID- 15296003 TI - A biophysical model of an inner hair cell. AB - Whole-cell patch-clamp recordings on isolated inner hair cells (IHCs) of guinea pig cochleae have revealed the presence of voltage-gated potassium channels. A biophysical model of an IHC is presented that indicates activation of slow voltage-gated potassium channels may lead to receptor potentials whose dc component decreases during the stimulus, and membrane potential hyperpolarizes when the stimulus is turned off. Both the decreasing dc and the hyperpolarization are, respectively, consistent with rapid adaptation and suppression of spontaneous rate in the auditory nerve. Receptor potentials recorded in vivo do not show these features, and when a nonspecific leak is included in the model to simulate microelectrode impalement, the model's receptor potentials become similar to those in vivo. The nonspecific leak creates an electrical shunt that masks slow channel activity and allows the cell to depolarize. Both the decreasing dc and the hyperpolarization are sensitive to the resting potential. Because the reported resting potentials in vivo and in vitro differ greatly, the model is used to investigate homeostatic mechanisms responsible for the resting potential. It is found that the voltage-gated potassium channels have the greatest influence on the resting potential, but that the standing transducer current may be sufficient to eliminate the decreasing dc and after-stimulus hyperpolarization. PMID- 15296004 TI - Periodogram based tests for distortion product otoacoustic emissions. AB - Distortion product otoacoustic emissions (DPOAEs) are an important nonbehavioral measure of cochlear function, which provides a close analogue of the behavioral pure-tone audiogram. DPOAEs are sinusoidal distortion products (DPs) produced by nonlinearities in the healthy cochlea. Detection of DPs is accomplished in the Fourier domain with a periodogram based test. The test compares the power in the DP periodogram bin to a noise estimate derived from a certain number of the surrounding bins. Statistical properties of this test to date have only been examined by constructing receiver operator characteristics curves derived from DPOAE measurements in normal and hearing impaired individuals. In this paper the null distribution of this order-statistic based test is explicitly derived, and via simulations intended to mimic the nonwhite features of real-ear noise measurements, the power of the test is demonstrated. These simulations demonstrate that a local F test is more powerful than this DPOAE test, with critical values that are easier to calculate. Although the power of both tests increase with an increasing number of bins, the improvement is negligible at around four bins. Since the power of both tests decrease at lower DP frequencies, it is not recommended to use a large number of bins. PMID- 15296005 TI - Effects of cochlear-implant pulse rate and inter-channel timing on channel interactions and thresholds. AB - Interactions among the multiple channels of a cochlear prosthesis limit the number of channels of information that can be transmitted to the brain. This study explored the influence on channel interactions of electrical pulse rates and temporal offsets between channels. Anesthetized guinea pigs were implanted with 2-channel scala-tympani electrode arrays, and spike activity was recorded from the auditory cortex. Channel interactions were quantified as the reduction of the threshold for pulse-train stimulation of the apical channel by sub threshold stimulation of the basal channel. Pulse rates were 254 or 4069 pulses per second (pps) per channel. Maximum threshold reductions averaged 9.6 dB when channels were stimulated simultaneously. Among nonsimultaneous conditions, threshold reductions at the 254-pps rate were entirely eliminated by a 1966 micros inter-channel offset. When offsets were only 41 to 123 micros, however, maximum threshold shifts averaged 3.1 dB, which was comparable to the dynamic ranges of cortical neurons in this experimental preparation. Threshold reductions at 4069 pps averaged up to 1.3 dB greater than at 254 pps, which raises some concern in regard to high-pulse-rate speech processors. Thresholds for various paired-pulse stimuli, pulse rates, and pulse-train durations were measured to test possible mechanisms of temporal integration. PMID- 15296006 TI - Age reduces response latency of mouse inferior colliculus neurons to AM sounds. AB - Age and stimulus rise time (RT) effects on response latency were investigated for inferior colliculus (IC) neurons in young-adult and old CBA mice. Single-unit responses were recorded to unmodulated and sinusoidal amplitude modulated (SAM) broadband noise carriers, presented at 35 to 80 dB SPL. Data from 63 young-adult and 76 old phasic units were analyzed to identify the time interval between stimulus onset and driven-response onset (latency). When controlling for stimulus sound level and AM frequency, significant age-related changes in latency were identified. Absolute latency decreased with age at all stimulus AM frequencies, significantly so for equivalent rise times (RT) < or = 12.5 ms. The linear correlation of latency with AM stimulus RT was significant for both young-adult and old units, and increased significantly with age. It is likely that both the decrease in absolute latency and the increase in latency/RT correlation with age are consistent with a reduction of inhibitory drive with age in the IC. These latency changes will result in age-related timing variations in brainstem responses to stimulus onsets, and therefore affect the encoding of complex sounds. PMID- 15296007 TI - External and internal limitations in amplitude-modulation processing. AB - Three experiments are presented to explore the relative role of "external" signal variability and "internal" resolution limitations of the auditory system in the detection and discrimination of amplitude modulations (AM). In the first experiment, AM-depth discrimination performance was determined using sinusoidally modulated broadband-noise and pure-tone carriers. The AM index, m, of the standard ranged from -28 to -3 dB (expressed as 20 log m). AM-depth discrimination thresholds were found to be a fraction of the AM depth of the standard for standards down to -18 dB, in the case of the pure-tone carrier, and down to -8 dB, in the case of the broadband-noise carrier. For smaller standards, AM-depth discrimination required a fixed increase in AM depth, independent of the AM depth of the standard. In the second experiment, AM-detection thresholds were obtained for signal-modulation frequencies of 4, 16, 64, and 256 Hz, applied to either a band-limited random-noise carrier or a deterministic ("frozen") noise carrier, as a function of carrier bandwidth (8 to 2048 Hz). In general, detection thresholds were higher for the random- than for the frozen-noise carriers. For both carrier types, thresholds followed the pattern expected from frequency selective processing of the stimulus envelope. The third experiment investigated AM masking at 4, 16, and 64 Hz in the presence of a narrow-band masker modulation. The variability of the masker was changed from entirely frozen to entirely random, while the long-term average envelope power spectrum was held constant. The experiment examined the validity of a long-term average quantity as the decision variable, and the role of memory in experiments with frozen-noise maskers. The empirical results were compared to predictions obtained with two modulation-filterbank models. The predictions revealed that AM-depth discrimination and AM detection are limited by a combination of the external signal variability and an internal "Weber-fraction" noise process. PMID- 15296008 TI - The effects of real and illusory glides on pure-tone frequency discrimination. AB - Experiment 1 measured pure-tone frequency difference limens (DLs) at 1 and 4 kHz. The stimuli had two steady-state portions, which differed in frequency for the target. These portions were separated by a middle section of varying length, which consisted of a silent gap, a frequency glide, or a noise burst (conditions: gap, glide, and noise, respectively). The noise burst created an illusion of the tone continuing through the gap. In the first condition, the stimuli had an overall duration of 500 ms. In the second condition, stimuli had a fixed 50-ms middle section, and the overall duration was varied. DLs were lower for the glide than for the gap condition, consistent with the idea that the auditory system contains a mechanism specific for the detection of dynamic changes. DLs were generally lower for the noise than for the gap condition, suggesting that this mechanism extracts information from an illusory glide. In a second experiment, pure-tone frequency direction-discrimination thresholds were measured using similar stimuli as for the first experiment. For this task, the type of the middle section hardly affected the thresholds, suggesting that the frequency change detection mechanism does not facilitate the identification of the direction of frequency changes. PMID- 15296009 TI - Limits to the role of a common fundamental frequency in the fusion of two sounds with different spatial cues. AB - Two experiments establish constraints on the ability of a common fundamental frequency (F0) to perceptually fuse low-pass filtered and complementary high-pass filtered speech presented to different ears. In experiment 1 the filter cut-off is set at 1 kHz. When the filters are sharp, giving little overlap in frequency between the two sounds, listeners report hearing two sounds even when the sounds at the two ears are on the same F0. Shallower filters give more fusion. In experiment 2, the filters' cut-off frequency is varied together with their slope. Fusion becomes more frequent when the signals at the two ears share low-frequency components. This constraint mirrors the natural filtering by head-shadow of sound sources presented to one side. The mechanisms underlying perceptual fusion may thus be similar to those underlying auditory localization. PMID- 15296010 TI - Specification of cross-modal source information in isolated kinematic displays of speech. AB - Information about the acoustic properties of a talker's voice is available in optical displays of speech, and vice versa, as evidenced by perceivers' ability to match faces and voices based on vocal identity. The present investigation used point-light displays (PLDs) of visual speech and sinewave replicas of auditory speech in a cross-modal matching task to assess perceivers' ability to match faces and voices under conditions when only isolated kinematic information about vocal tract articulation was available. These stimuli were also used in a word recognition experiment under auditory-alone and audiovisual conditions. The results showed that isolated kinematic displays provide enough information to match the source of an utterance across sensory modalities. Furthermore, isolated kinematic displays can be integrated to yield better word recognition performance under audiovisual conditions than under auditory-alone conditions. The results are discussed in terms of their implications for describing the nature of speech information and current theories of speech perception and spoken word recognition. PMID- 15296011 TI - Blind normalization of speech from different channels. AB - We show how to construct a channel-independent representation of speech that has propagated through a noisy reverberant channel. This is done by blindly rescaling the cepstral time series by a nonlinear function, with the form of this scale function being determined by previously encountered cepstra from that channel. The rescaled form of the time series is an invariant property of it in the following sense: It is unaffected if the time series is transformed by any time independent invertible distortion. Because a linear channel with stationary noise and impulse response transforms cepstra in this way, the new technique can be used to remove the channel dependence of a cepstral time series. In experiments, the method achieved greater channel-independence than cepstral mean normalization, and it was comparable to the combination of cepstral mean normalization and spectral subtraction, despite the fact that no measurements of channel noise or reverberations were required (unlike spectral subtraction). PMID- 15296012 TI - Therelationship between professional operatic soprano voice and high range spectral energy. AB - Operatic sopranos need to be audible over an orchestra yet they are not considered to possess a singer's formant. As in other voice types, some singers are more successful than others at being heard and so this work investigated the frequency range of the singer's formant between 2000 and 4000 Hz to consider the question of extra energy in this range. Such energy would give an advantage over an orchestra, so the aims were to ascertain what levels of excess energy there might be and look at any relationship between extra energy levels and performance level. The voices of six operatic sopranos (national and international standard) were recorded performing vowel and song tasks and subsequently analyzed acoustically. Measures taken from vowel data were compared with song task data to assess the consistency of the approaches. Comparisons were also made with regard to two conditions of intended projection (maximal and comfortable), two song tasks (anthem and aria), two recording environments (studio and anechoic room), and between subjects. Ranking the singers from highest energy result to lowest showed the consistency of the results from both vowel and song methods and correlated reasonably well with the performance level of the subjects. The use of formant tuning is considered and examined. PMID- 15296013 TI - Frequency dependence of acoustic properties of aqueous glucose solutions in the VHF/UHF range. AB - The bioultrasonic spectroscopy system was employed for measurements of velocity and attenuation coefficient of glucose solutions in the VHF/UHF range. The relation between the slope of the square of velocity and the relaxation parameters, and the relation between the frequency exponent on attenuation coefficient and the relaxation parameters are investigated. In order to carry out numerical calculations, a model for a single relaxation process is employed, wherein the attenuation coefficient is expressed as (A/( 1 + (f/falpha)2) + B)f2 where falpha is the attenuation relaxation frequency, and A and B are constants. The numerical calculations show that the slope of the square of the velocity is determined uniquely by the velocity relaxation frequency fv and v(infinity)2 - v(0)2 where v0 is the zero-frequency velocity and v(infinity) is the infinite frequency velocity, and that the frequency exponent on the attenuation coefficient is determined uniquely by falpha and A/B. For experimental considerations, the velocities and the attenuation coefficients of 5, 15, and 25% concentration aqueous solutions of glucose were measured in the frequency range 20 to 700 MHz. The data for the 5 and 15% aqueous solutions can be explained using the single relaxation model. However, the data for the 25% aqueous solution suggest the existence of multirelaxation processes. PMID- 15296014 TI - The corono-apically varying ultrasonic velocity in human hard dental tissues. AB - The speed of ultrasound at 20 MHz is measured for hard dental tissues inside human teeth. This includes the cementum, for which no data are available. The spatial distribution, extrema, and means of the longitudinal ultrasound velocity (LUV) are determined with an emphasis on the apical thirds and an extended spectrum. Tissue areas are investigated by optical means and by acoustical scanning, in order to compare apical regions-of-interest with the complete mineralized wet porous tissue that lies beneath the enamel cap. The maximal LUV in a single dentin specimen varies from 3903 m/s to 4226 m/s. The dentin's frequency distribution of LUV at 20 degrees C exhibits a predominant peak feature comprising several Voigt functions. Introducing standardized relative tooth width portions, the corono-apical decrease in LUV of 21 specimens is approximated by LUV=4224 - (257* ln(y)) along reduced distances in dentin. Abnormal teeth require a higher resolution and an approximate equation of the form LUV= (sigma(ai*yi))/(1 + sigma(bi+1*y(i+1. It can be used each time the corono-apical variation has to be quantified in each of the three tissues. Ten coefficients are numerically exemplified. An error evaluation is performed, which denotes errors of 0.2% +/- 1.3% (enamel), -0.1% +/- 1.6% (cementum), and acceptable residual errors for dentin. PMID- 15296015 TI - Sounds produced by Norwegian killer whales, Orcinus orca, during capture. AB - To date very little is still known about the acoustic behavior of Norwegian killer whales, in particular that of individual whales. In this study a unique opportunity was presented to document the sounds produced by five captured killer whales in the Vestfjord area, northern Norway. Individuals produced 14 discrete and 7 compound calls. Two call types were used both by individuals 16178 and 23365 suggesting that they may belong to the same pod. Comparisons with calls documented in Strager (1993) showed that none of the call types used by the captured individuals were present. The lack of these calls in the available literature suggests that call variability within individuals is likely to be large. This short note adds to our knowledge of the vocal repertoire of this population and demonstrates the need for further studies to provide behavioural context to these sounds. PMID- 15296016 TI - Directionality of dog vocalizations. AB - The directionality patterns of sound emission in domestic dogs were measured in an anechoic environment using a microphone array. Mainly long-distance signals from four dogs were investigated. The radiation pattern of the signals differed clearly from an omnidirectional one with average differences in sound-pressure level between the frontal and rear position of 3-7 dB depending from the individual. Frequency dependence of directionality was shown for the range from 250 to 3200 Hz. The results indicate that when studying acoustic communication in mammals, more attention should be paid to the directionality pattern of sound emission. PMID- 15296017 TI - Non-Gaussian statistics and temporal variations of the ultrasound signal backscattered by blood at frequencies between 10 and 58 MHz. AB - Very little is known about the blood backscattering behavior and signal statistics following flow stoppage at frequencies higher than 10 MHz. Measurements of the radio frequency (rf) signals backscattered by normal human blood (hematocrit = 40%, temperature = 37 degrees C) were performed in a tube flow model at mean frequencies varying between 10 and 58 MHz. The range of increase of the backscattered power during red blood cell (RBC) rouleau formation was close to 15 dB at 10 and 36 MHz, and dropped, for the same blood samples, below 8 dB at 58 MHz. Increasing the frequency from 10 to 58 MHz raised the slope of the power changes at the beginning of the kinetics of aggregation, and could emphasize the non-Gaussian behavior of the rf signals interpreted in terms of the K and Nakagami statistical models. At 36 and 58 MHz, significant increases of the kurtosis coefficient, and significant reductions of the Nakagami parameter were noted during the first 30 s of flow stoppage. In conclusion, increasing the transducer frequency reduced the magnitude of the backscattered power changes attributed to the phenomenon of RBC aggregation, but improved the detection of rapid growth in aggregate sizes and non-Gaussian statistical behavior. PMID- 15296018 TI - Scatterer size estimation in pulse-echo ultrasound using focused sources: theoretical approximations and simulation analysis. AB - The speckle in ultrasound images has long been thought to contain information related to the tissue microstructure. Many different investigators have analyzed the frequency characteristics of the backscattered signals to estimate the scatterer acoustic concentration and size. Previous work has been mostly restricted to unfocused or weakly focused ultrasound sources, thus limiting its implementation with diagnostically relevant fields. Herein, we derive equations capable of estimating the size of a scatterer for any reasonably focused source provided that the velocity potential field in the focal region can be approximated as a three-dimensional Gaussian beam, scatterers are a sufficient distance from the source, and the field is approximately constant across the scatterer. The calculations show that, when estimating the scatterer size, correcting for focusing requires a generalized attenuation-compensation function that includes both attenuation and focusing along the beam axis. The Gaussian approximation is validated by comparing the ideal velocity potential field for three spherically focused sources with f-numbers of 1, 2, and 4 to the Gaussian approximation for frequencies from 2 to 14 MHz. The theoretical derivations are evaluated by simulating the backscatter by using spherically focused sources (f numbers of 1, 2, and 4) adjacent to attenuating media (0.05 to 1 dB/cm/MHz) that contain scatterers with Gaussian impedance distributions. The generalized attenuation-compensation function yielded results accurate to 7.2% while the traditional attenuation-compensation functions that neglected focusing had errors as high as 103%. PMID- 15296019 TI - Scatterer size estimation in pulse-echo ultrasound using focused sources: calibration measurements and phantom experiments. AB - In a companion paper [T. A. Bigelow and W. D. O'Brien Jr., J. Acoust. Soc. Am. 116, 578 (2004)], theory, supported by simulations, showed that accurate scatterer size estimates could be obtained using highly focused sources provided that the derived generalized attenuation-compensation function was used and the velocity potential field near the focus could be approximated as a three dimensional Gaussian. Herein, the theory is further evaluated via experimental studies. A calibration technique is developed to find the necessary equivalent Gaussian dimensions for a focused source using reflections obtained from a rigid plane scanned through the focus. Then, the theoretical analysis of focused sources is validated experimentally using three spherically focused ultrasound transducers to estimate the radius of glass beads imbedded in tissue mimicking phantoms. Both the impact of focusing (f/1, f/2, and f/4) and the effect of scatterer type (comparing glass bead results to simulation results that used scatterers with Gaussian impedance distributions) were tested. The simulated differences agree with the measured differences to within 2.5% provided that the comparison is made between the same scatterer type and sources with the same equivalent Gaussian dimensions. The improvement provided by the generalized attenuation-compensation function is greatly influenced by the type of scatterer whose size is being estimated and decreases as the wavelength dependence of the Gaussian depth of focus is reduced. PMID- 15296020 TI - [Effects of home-visit nutrition education on nutritional status improvement of an urban community-dwelling elderly women in Korea]. AB - PURPOSE: This study was conducted to evaluate the effects on home-visit nutrition education by a dietitian on nutritional status improvement of an urban community dwelling elderly women in Korea. METHODS: In the baseline survey, information on general characteristics, health-related characteristics, anthropometric measurements, biochemical measurements, nutritional knowledge. nutritional attitude, dietary habits, and food and nutrient intakes of 183 elderly people were obtained. The intervention group received weekly home-visit nutrition education over 4 months. RESULTS: After home-visiting nutrition education, nutritional knowledge, nutritional attitude and dietary habit were increased significantly by 1.8, 2.1 and 6.9 in the intervention group (P<0.01), respectively, who also appeared to consume more cereals and their products, legumes and their products, vegetables, seasonings, milk and dairy products than the control group. It was found that the nutrient intake increased significantly regarding energy, protein, calcium, iron, phosphorus, thiamin and riboflavin (P<0.05). The MAR (mean nutrient adequacy ratio) increased by 0.22 during the period of the study in the intervention group, and 0.09 in the control group, the difference being statistically significant (P<0.01). Differences between in mean change of anthropometric and biochemical indices between the intervention and control groups were not significant. CONCLUSIONS: These findings suggest that home-visit nutrition education by a dietitian is effective for improvement of the nutritional status of elderly women in an urban community. In conclusion, home visit nutrition education should be recommended for nutritional status improvement and health promotion in the community elderly. PMID- 15296021 TI - [Efficacy of a smoking relapse prevention program by postdischarge telephone contacts: a randomized trial]. AB - OBJECTIVES: This randomized controlled trial assessed the efficacy of a smoking relapse prevention program featuring 3 postdischarge telephone contacts with subjects who had quit smoking on hospitalization. METHODS: Patients were randomly assigned to public health nurse-mediated behaviorally oriented in-patient counseling focused on relapse prevention (control group, n = 49), or the same inpatient counseling with postdischarge telephone contacts at 7, 21 and 42 days after discharge (intervention group, n = 57). The main outcome measure, smoking cessation rate, was obtained from self-reports at 3, 6 and 12 months after discharge. Smoking cessation at 12 months after discharge was confirmed by urinary nicotine concentration. RESULTS: At 3, 6 and 12 months smoking cessation rates were 83%, 63% and 56% for the intervention group, and 76%, 65% and 51% for control group. After adjustment for sex, age, having any complication, number of family members, smoking status on admission, strength of nicotine dependence and self confidence to quit smoking, the odds ratio of cessation among the intervention group were 1.46 (95% confidence interval (CI): 0.48-4.47), 0.82 (95% CI: 0.31-2.17) and 0.99 (95% CI: 0.40-2.45) at 3, 6 and 12 months after discharge, respectively. CONCLUSION: This program had limited efficacy to maintain postdischarge smoking abstinence. We should re-consider the modality of smoking cessation program for relapse prevention among hospitalized patients. PMID- 15296022 TI - [Health of aged people living with siblings who are professional nurses in China from the view point of lifestyle and health condition]. AB - PURPOSE: Since 1978 in China, rapid economic development has taken place and the nation's quality of life has improved through the introduction of reform-opening policies. Such change has caused new health problems, partially due to aging of the population, with increase in lifestyle-related diseases and environmental pollution, and also expansion of regional variation. In this study, diseases undergoing treatment (relevance) and lifestyles of the elderly living with siblings who are professional nurses were evaluated. METHOD: We conducted a study in 23 provinces to discern characteristics and factors related to lifestyle and situations of patients undergoing treatment. We analyzed 1,548 senior citizens (response rate: 82.1%) over 65 years old living with a sibling who is a professional nurse. The professional nurse provided the replying to the questions. RESULTS: 1. A total of 457 out of 597 males (76.5%) and 725 out of 951 females (76.2%) had diseases under treatment. Males over 75 years old suffered from arteriosclerosis, cerebrovascular diseases, and heart disease. Females over 75 years old suffered from arteriosclerosis, respiratory diseases, and eye diseases. 2. In both males and females over 75 years old (older elderly) there were no significant differences in the Health Practice Index (HPI) from persons under 75 (younger elderly). Older elderly were more likely to snack often. Among males and females, 5 of 8 health-practices, such as a napping and physical exercise, differed. Females were less likely to smoke and drink alcohol. 3. In both males and females, non-diseased participants had a higher HPI than that of diseased participants. This tendency was the same in both younger and older elderly. 4. Cluster analyses of patterns of diseases revealed that the 23 provinces could be classified into 4 areas. The HPI in areas with a low proportion of diseased subjects was significantly higher than that in areas with a high proportion of diseased. One of the areas' HPI appeared to be noticeably lower than that of the other 3 and the number of participants with low physical exercise was higher in this case. CONCLUSION: Our data indicate that individuals having a high HPI appear less likely to develop lifestyle-related diseases. In 4 areas divided by the cluster analysis of patterns of diseases, there were significant differences in HPI, smoking and physical exercise. PMID- 15296023 TI - [Development of an assessment tool for preventive home visits]. AB - PURPOSE: To develop a new version of the Minimum Data Set-Home Care (MDS-HC) for preventive home visits. METHODS: Public health nurses (PHNs) visited community dwelling elderly who were independent in ADL but dependent in IADL, every three months using the MDS-HC. Assessment forms and visiting records documented by the PHNs at the third visit (n = 217) and the assessment forms available from the third to the fifth visit (n = 163) were analyzed. Assessment items appropriate for preventive home visits were selected based on frequency of problem identified, changes observed, and areas requiring follow-up identified by the PHNs. These items were evaluated for their sensitivity and specificity in identifying the areas requiring follow-up. RESULTS: 53 assessment items were selected based on the frequency of the problems identified and the changes observed. An additional 36 assessment items were needed to identify the 24 areas requiring follow-up. In total, 89 of the 247 items in the original were identified as being appropriate for the preventive version of the MDS-HC. The sensitivity of the new version to detect the 24 problem areas were the same or higher than with the original. CONCLUSION: The preventive version of the MDS-HC had about one third of the items in the original and appropriately detected the areas that needed follow up. The use of this tool should lead to a more systematic approach to preventive health visits. PMID- 15296024 TI - [A study of business management of public health office]. PMID- 15296025 TI - [Encryption technique for linkable anonymizing]. AB - Linkage of different records such as health insurance claims or medical records for the purpose of cohort studies or cancer registration usually requires matching with personal names and other personally identifiable data. The present study was conducted to examine the possibility of performing such privacy sensitive procedures in a "linkable anonymizing" manner using encryption. While bidirectional communication entails encryption and deciphering, necessitating both senders and receivers sharing a common secret "key", record linkage entails only encryption and not deciphering because researchers do not need to know the identity of the linked person. This unidirectional nature relieves researchers from the historical problem of "key sharing" and enables data holders such as municipal governments and insurers to encrypt personal names in a relatively easy manner. The author demonstrates an encryption technique using readily available spread-sheet software, Microsoft Excel in a step-by-step fashion. Encoding Chinese characters into the numeric JIS codes and replacing the codes with a randomly assigned case-sensitive alphabet, all names of Japanese nationals will be encrypted into gibberish strings of alphabet, which can not be deciphered without the secret key. Data holders are able to release personal data without sacrificing privacy, even when accidental leakage occurs and researchers are still able to link records of the same name because encrypted texts, although gibberish, are unique to each name. Such a technical assurance of privacy protection is expected to satisfy the Privacy Protection Act or the Ethical Guidelines for Epidemiological Research and enhance public health research. Traditional encryption techniques, however, cannot be applied to cancer or stroke registration, because the registrar receives reports from numerous unspecified senders. The new public key encryption technique will enable disease registry in a linkable anonymizing manner. However various technical problems such as complexity, difficulties in registrar inquiries and risk of code-breaking make the encryption technique unsuitable for disease registry in the foreseeable future. PMID- 15296026 TI - [The vital statistics in Japan before World War 2--the transition of the viewpoint about nation in the listing in vital statistics reports]. PMID- 15296027 TI - Clinical analysis of interstitial pneumonia after surgery for lung cancer. AB - OBJECTIVE: The postoperative development or exacerbation of interstitial pneumonia (IP) in lung cancer patients often affects prognoses. We analyzed the patients who suffered from IP after surgery, to determine treatment and prevention of IP. METHODS: One hundred and one consecutive patients who underwent resection were enrolled in the study. Clinical background and post-, perioperative course were compared between patients who developed IP and those who did not. RESULTS: If IP developed or was exacerbated, steroid pulse (SP) therapy, immunosuppressant (IS) therapy or nitric oxide (NO) inhalation therapy was employed. Of 101 patients, 20 had suffered from IP before surgery. In four of these 20 (20%), postoperative exacerbation was observed. SP therapy was given to all patients. To one patient, IS and NO therapy were added. Three of the four patients died. 81 patients did not have IP; three of them (3.7%) developed IP after the operation and were treated with SP therapy. To one patient, IS and NO therapy were added. Two of the three patients died. IP development or exacerbation after surgery was observed in seven of 101 patients, and five of them died. It was significantly more frequent in patients with poorly differentiated squamous cell carcinoma, restrictive change in pulmonary function tests, and a low percentage diffusion capacity for carbon monoxide. Postoperative development or exacerbation was observed in patients who had undergone lobectomy or pneumonectomy. CONCLUSIONS: Postoperative IP is a serious complication. Further studies are needed to determine definitive therapeutic options. For the patients with the aforementioned risk factors, limited surgery must be considered. PMID- 15296028 TI - Does the lobectomy plus lymph node dissection still remain a standard surgical procedure for patients with cT1N0M0 adenocarcinoma of the lung? AB - OBJECTIVES: Controversies still exists regarding treatment for cT1N0M0 adenocarcinoma of the lung. The following topics need to be answered: 1) Should all patients undergo lobectomy plus lymph node dissection? and 2) Is there poor prognostic subgroup that may need adjuvant therapy? METHODS: Between 1990 and 1999, 141 patients with cT1N0M0 adenocarcinoma of the lung underwent lobectomy plus lymph node dissection. Fifteen clinicopathological characteristics of the entire population were investigated with regard to survival. Forty-seven samples, which were possible to reexamine among 68 patients with small adenocarcinoma 2 cm or less in greatest dimension, were assessed according to Noguchi's classification. RESULTS: Nine of fifteen clinicopathological variables were significant in indicating poor prognostic factors in univariate analysis: gender, differentiation, p-T status, p-N status, pm, lymphatic invasion, vascular invasion, pleural invasion, and serum carcinoembryonic antigen (CEA) level. The p N status and high serum CEA level were independent predictive variables in multivariate analysis. A five-year survival rate for patients with Noguchi's type A and B was 100%. However, six (8.8%) of 68 patients with small adenocarcinoma had lymph node involvement and four patients (5.9%) had pulmonary metastasis. CONCLUSIONS: It is inappropriate and inadequate to omit lobectomy or lymph node dissection only on the basis of tumor size. Therefore, it seems reasonable to conclude that lobectomy plus lymph node dissection still remains as a standard surgical procedure to treat cT1N0M0 adenocarcinoma of the lung. We must continue to search for new deciding factors in order to choose candidates for limited operation among patients with cT1N0M0 adenocarcinoma of the lung. PMID- 15296029 TI - Mitral valve replacement after percutaneous transluminal mitral commissurotomy. AB - OBJECTIVES: We reviewed our experience of mitral valve replacement (MVR) after percutaneous transluminal mitral commissurotomy (PTMC) for mitral stenosis (MS). METHODS: From December 1987 to December 2001, PTMC was conducted in 75 patients with symptomatic rheumatic MS. At mean follow-up of 8.4+/-3.5 years, 11 patients (14.7%) underwent MVR for mitral restenosis (9 cases) and mitral regurgitation (MR) (2 cases). The mean interval between PTMC and MVR was 5.2+/-3.2 years. RESULTS: There were 2 hospital deaths (due to low output syndrome and mediastinitis) and 2 complications (prosthetic valve endocarditis and left ventricular rupture). The mitral valve area (MVA) at pre-PTMC, post-PTMC and pre MVR was 1.02+/-0.48 cm2, 1.55+/-0.59 cm2, 1.04+/-0.23 cm2, respectively. The MVA significantly increased after PTMC (p<0.01), but decreased significantly to the pre-PTMC value at pre-MVR (p<0.05). The left atrial dimension (LAD) significantly increased from 50.4+/-10.8 mm at pre-PTMC to 61.1+/-13.1 mm at pre-MVR (p<0.05). The number of significant tricuspid regurgitation (TR) cases increased from 2 at pre-PTMC to 5 at pre-MVR. The New York Heart Association class got better after PTMC (3 cases in class III at pre-PTMC to 0 at post-PTMC), but at pre-MVR, deteriorated to the same level at pre-PTMC (4 cases in class III). CONCLUSIONS: Our results of MVR after PTMC were reasonable to be considered despite their high risk at MVR resulting in 2 hospital deaths. For the reliable relief of MS and control of TR, not PTMC but MVR combined with tricuspid annuloplasty may be preferable in such two cases suffering from congestive heart failure with significant TR at first intervention. Close follow-ups like periodic ultrasonic cardiography studies should be conducted to gain more information on the mitral restenosis, TR deterioration and dilatation of the cardiac chambers. PMID- 15296030 TI - Subepicardial aneurysm detected during diagnosis of acute cardiac failure. AB - A 55-year-old female developed dyspnea at night and visited our hospital. A diagnosis of acute cardiac failure was made and the patient was immediately admitted. A false ventricular aneurysm with blood flow was found in the lateral wall of the left ventricle. Past history of myocardial infarction was not clear. Repeated cardiac failure may have resulted from decreased cardiac output by increased blood flow in the aneurysm due to its expansion. After patch closure was performed, the symptoms improved. PMID- 15296031 TI - Transventricular annuloplasty for ischemic mitral regurgitation in the Dor procedure. AB - Since the residual mitral regurgitation after the Dor procedure contributes to increasing postoperative mortality, repair of mitral regurgitation has become one of the essential surgical approaches. We describe two cases of transventricular posterior annuloplasty using a trimmed Duran ring for surgical management of ischemic mitral regurgitation performed with the Dor procedure. This procedure is easy to perform and provides secure annuloplasty because the mitral annulus can be easily exposed through the same ventriculotomy incision. PMID- 15296032 TI - Triad of thymoma, myasthenia gravis and pure red cell aplasia combined with Sjogren's syndrome. AB - A 36-year-old woman complained of cough and high fever. Computed tomographic scans demonstrated a mediastinal mass. A couple of months later, she developed dryness in her eyes and mouth. Biopsy of the lip confirmed the diagnosis of Sjogren's syndrome. She underwent thymo-thymomectomy. Pathological findings of the mass revealed thymoma. At two months after surgery, she developed ptosis and dysphagia that were compatible with myasthenia gravis. The clinical symptoms were adequately controlled with prednisolone. At eleven months after surgery, she presented with severe anemia, which led to the diagnosis of pure red cell aplasia. The following treatment with cyclosporin caused hemoglobin concentration to rise. However, she continues to suffer from dryness of her eyes and mouth. The case is the first to be reported with Sjogren's syndrome and the triad of thymoma, myasthenia gravis and pure red cell aplasia, and is compared with previously reported cases of the three conditions. PMID- 15296033 TI - Radiotherapy-induced aortic valve disease associated with porcelain aorta. AB - Mediastinal irradiation has been reported to induce cardiac disease such as pericarditis, valvular dysfunction, conduction abnormalities, accelerated arteriosclerosis of the coronary arteries, and also calcifications of the ascending aorta. We herein describe a case of radiotherapy-induced porcelain aorta and aortic valve disease and their surgical treatment. The patient was diagnosed with myasthenia gravis (MG) in 1965 (Osserman's type II), and mediastinal irradiation was performed in 1970 for treatment of thymic tumor associated with MG. Thirty years after radiation therapy, complete atrioventricular block and aortic valve disease with severe calcification of the ascending aorta and aortic arch (porcelain aorta) were detected on echo cardiogram and cardiac catheterization. A permanent pacemaker was implanted via the left subclavian vein and aortic valve replacement was performed under extracorporeal circulation established by selective cerebral perfusion and balloon occlusion instead of aortic cross-clamping. As no risk factors of arteriosclerosis such as hypercholesterolemia, hyperglycemia and hypertension were apparent, we concluded that the aortic valve disease and porcelain aorta were primarily induced by radiotherapy. PMID- 15296034 TI - Metastasis to the thyroid from lung adenocarcinoma mimicking thyroid carcinoma. AB - We herein report a case of metastasis to the thyroid from lung adenocarcinoma mimicking thyroid carcinoma. The thyroid tumor was palpated in the left lobe of the thyroid and diagnosed as primary thyroid carcinoma by fine-needle aspiration cytology. The patient also had a large pulmonary tumor and tiny pulmonary nodules, which were respectively diagnosed as moderately differentiated adenocarcinoma of the lung and intrapulmonary metastases from the main large lung carcinoma by the pathological examination of the biopsy specimens obtained by video-assisted thoracic surgery. Hemithyroidectomy with radical neck dissection was performed. The thyroid tumor was diagnosed as metastasis to the thyroid from lung adenocarcinoma, because it showed mucin production, positive immunoreactivity for carcinoembryonic antigen and negative immunoreactivities for thyroglobulin and calcitonin. The patient received systemic chemotherapy and died of the disease 1 year and 7 months after the diagnosis was made. PMID- 15296035 TI - Synchronous pulmonary atypical adenomatous hyperplasia and metastatic osteosarcoma in a young female. AB - A 17-year-old female underwent metastasectomy of three synchronous lesions in the bilateral lungs under the diagnosis of metastatic osteosarcoma, however, one of them was found to be atypical adenomatous hyperplasia (AAH). Since AAH is very rare among young people, a careful evaluation of high-resolution computed tomographic image is important in determining the operative indications and procedures in patients with multiple metastatic tumors. PMID- 15296036 TI - Incidence of perioperative myocardial infarction in off-pump coronary artery bypass. PMID- 15296037 TI - Moving the research agenda forward for children and adolescents with cancer. PMID- 15296038 TI - Pediatric cancer research from past successes through collaboration to future transdisciplinary research. AB - Multidisciplinary collaboration in therapeutic research in childhood cancer has been responsible for enormous improvements in outcomes. Many of the improvements have resulted from large clinical trials carried out in multisite settings through the Children's Oncology Group (COG) and its predecessors, the Children's Cancer Group, the Pediatric Oncology Group, the National Wilms' Tumor Study, and Intergroup Rhabdomyosarcoma Study Groups. Childhood acute lymphoblastic leukemia models the past success of 35 years of randomized clinical trials that resulted in survival rates of around 80%. However more can be done to improve both survival rates and the quality of survival. Areas that can benefit from a transdisciplinary model of research are discussed, as well as challenges to this form of collaboration. PMID- 15296039 TI - Advancing biobehavioral research in childhood cancer. AB - Research on individual and family responses to a child's or adolescent's cancer diagnosis and treatment is important because of significant advances in the field of pediatric oncology. To date, the majority of research has been behavioral or psychosocial in nature. It could be argued that a more holistic perspective that includes psychological, sociocultural, and biological dimensions would advance knowledge about individual and family responses to the experience of childhood cancer. Biobehavioral research refers to investigations that link behavioral and biological underpinnings in specific areas of science. The purpose of this article is to review research in two broad areas that could benefit from a biobehavioral perspective-psychosocial functioning/behavioral adjustment, and CNS treatment outcomes. Studies that include biological measures are highlighted. Advantages, challenges, and strategies for advancing biobehavioral research in childhood cancer are proposed. PMID- 15296040 TI - Symptom management research in children with cancer. AB - A review of the clinical research studies published within the past 5 years revealed that efforts to manage symptoms of cancer and its treatments have not kept pace with new advances in the cure for cancer. Children with cancer continue to experience distressing physical symptoms caused by the disease and treatment. The purpose of this article is to provide a concise overview of the most common symptoms experienced by children with cancer. These symptoms include pain, nausea and vomiting, nutritional concerns, mucositis, and fatigue experienced by the child with cancer. Recommendations for future research are addressed. PMID- 15296041 TI - Health-related quality of life in pediatric oncology: current status and future challenges. AB - This article presents information regarding the current status of health-related quality of life (HRQL) assessment in pediatric oncology, including the substantial advances in our knowledge that have occurred during the past 10 years. Additionally, issues that continue to challenge researchers who are attempting to measure health-related quality of life in children from point of diagnosis to cure or to death are identified. It is posited that researchers must conduct investigations of the impact of health-related quality-of-life data on clinical care and outcomes for the field to continue to advance, and for these data to be valued by patients, families, and health care providers. PMID- 15296042 TI - Research priorities for family assessment and intervention in pediatric oncology. AB - Four broad areas of research priority related to family assessment and intervention in pediatric oncology are outlined. The importance of a contextual, systemic approach is highlighted along with the value of systematic empirical assessment of psychosocial risk at diagnosis. Research to advance the development and application of standards of care during treatment is also advocated. The chapter concludes with opportunities for family-oriented research related to survivorship and end-of-life care. PMID- 15296043 TI - The adolescent/young adult experience. AB - Adolescents and young adults with cancer (AYA) have strikingly poorer outcomes when compared to younger or older patients. Contributing reasons include low rates of enrollment in clinical trials and the "invisibility" of the AYA perspectives in research reports. We recommend a shift in research perspectives away from function-based studies that focus on morbidity and mortality outcomes to meaning-based models that will very likely rely upon qualitative methods and findings as the basis for developing psychosocial interventions that are sensitive to the AYA cancer experience. In addition, research on positive health concepts such as hope, spirituality, and positive coping are recommended as a way to learn about the effective strategies used by AYA to adjust to the cancer experience. PMID- 15296044 TI - Good communication in pediatric cancer care: a culturally-informed research agenda. AB - Communication affects people's factual understanding of the situations they are in. This in turn affects consent to be treated and adherence. Although a growing body of literature concerns cancer communication, it generally concerns adult patients; pediatric-specific knowledge is limited. Furthermore, most research focuses shortsightedly on physicians rather than providers who have more patient contact, favors studying the single visit instead of encompassing the cycle of cancer care, does not take the child into account, and ignores the importance of provider-provider communication. Moreover, cultural issues are only narrowly conceived. In exploring culture's role in pediatric cancer care communication, this article demonstrates that culture cannot be defined as just ethnicity or race. Professions also have cultures and resultant differences in communication patterns that can lead to communication failures. It also shows that there is a crucial need for more applied research as well as more qualitative research that can enrich our understanding of the complicated context-related factors facilitating or barring successful pediatric cancer communication. PMID- 15296045 TI - Delivery of culturally competent care to children with cancer and their families- the Latino experience. AB - This article discusses selected cultural factors such as language and interpretation services, beliefs, health care practices, and communication styles of Latino families that can increase and enhance the ability of nurses to work with a child with cancer and his or her family. Suggestions for research and clinical intervention are presented. PMID- 15296046 TI - An overview of progress in childhood cancer survival. AB - Survival for children with cancer has continued to improve over the past 20 years, with 5-year survival rates now approaching 80% and with an increasing number of children surviving into adulthood. In recognition of the need to address important issues for survivors of childhood cancer, the National Cancer Institute (NCI) has supported the Childhood Cancer Survivor Study (CCSS) since 1993. The CCSS has established a cohort of over 14,000 5-year survivors of childhood cancer initially diagnosed between 1970 and 1986 and has obtained comprehensive summaries of treatment received by these survivors through abstraction of medical records for chemotherapy, radiation therapy, and surgical procedures. Survivors in the CCSS cohort completed a baseline questionnaire and follow-up questionnaires to provide information about their current status. The CCSS collected buccal cells for DNA isolation from members of the survivor cohort to allow evaluations of the contribution of genetic factors to long-term sequelae of cancer therapy. The CCSS is now poised to expand its contributions through studies designed to gain greater insight into the biological basis for long-term adverse effects of cancer treatment and is enlarging its efforts to include more recently treated cohorts of patients. The experts present at the work group, Moving the Research Agenda Forward for Children With Cancer, offered several possibilities for utilizing CCSS data in expanding research opportunities concerned with cancer survivorship. PMID- 15296047 TI - Integrating the child into home and community following the completion of cancer treatment. AB - The present article examines the period of time immediately following the completion of treatment for childhood cancer. The unique concerns experienced by families at this stage of the cancer treatment are examined, and the specific challenges that children face as they renegotiate roles and relationships that are necessary for successful reintegration into family, school, and community settings are discussed. Obstacles to successful reintegration that are frequently encountered by patients and families are reviewed, as well as variables that may promote optimal adjustment during this transitional period. The need for continued research in this area is highlighted, and specific research questions are identified. An emphasis is placed on applying a socioecological framework to research and clinical work with pediatric oncology patients at this stage of the cancer experience. PMID- 15296048 TI - A model of care for childhood cancer survivors that facilitates research. AB - The majority of children and adolescents diagnosed with cancer will achieve long term survival after contemporary therapy. Consequent to this success are challenges inherent in coordinating lifelong health care for a group predisposed to a variety of cancer-related complications. With increasing numbers of aging adult survivors of childhood cancer, clinicians now face the additional challenge of studying delayed effects of childhood cancer in the context of organ senescence. Clinicians must also address the transition of survivor health care from the pediatric oncology setting to the adult community. Salient issues influencing health care of long-term childhood cancer survivors are summarized, and a model for monitoring late treatment effects used at a pediatric cancer facility is presented. This model is remarkable for its ability to enhance optimal delivery of long-term survivor care, facilitate the transition of survivor care from the pediatric treatment center to community, providers, and support investigations of late cancer-related morbidity and mortality. PMID- 15296049 TI - End-of-life research as a priority for pediatric oncology. AB - Approximately 2,200 children and adolescents die a cancer-related death each year in the United States; of these, almost 90% will die while experiencing 2 to 8 troubling symptoms. With improved symptom control and end-of-life care, these patients might suffer less before they die and their survivors might experience fewer or less intense adverse physical and mental conditions secondary to their bereavement. The focus of this article is on five key areas related to end of life where research is critically needed; (a) the characteristics of cancer related death and the profiles of survivorship in bereaved family members and health care providers, (b) the trajectory of dying in children and adolescents and a comparison of care delivery preferred by the family and that actually delivered, (c) end-of-life decision making, (d) the financial costs of a child or adolescent dying a cancer-related death and associated policy making, and (e) outcomes of symptom-directed or bereavement interventions. Knowing the characteristics of cancer-related deaths in children and adolescents will help researchers and clinicians develop and test effective interventions related to symptom management, decision making, and availability of care delivery models that match the dying child's needs and preferences. Such interventions could also contribute to the highest quality and cost-effective care being provided to the bereaved survivors. PMID- 15296050 TI - Human suffering: the need for relationship-based research in pediatric end-of life care. AB - Children living with and dying from advanced cancer and their families experience significant suffering. The cure of disease and the relief of suffering are dual moral obligations of our professions. To relieve suffering, health care providers must understand the multiple dimensions of the person who suffers and the complex set of relationships within the natural and the clinical social networks. Pediatric oncology research must include appropriately designed studies with sound methodology and measurement strategies to test and refine theories that account for the link between human relationships and the relief of suffering. Studies should assess as many theoretical models as possible, including the social network, perceptions of support, and provider-recipient interactions; their physical, emotional, behavioral, and spiritual concomitants; and their impact on medical decision making and health outcomes. Future directions in pediatric end-of-life care research must also include evaluating social and spiritual interventions developed on the basis of solid hypotheses regarding the positive and negative influences of interpersonal dynamics on the processes that mediate between suffering and well-being. PMID- 15296051 TI - Moving the research agenda forward for children and adolescents with cancer. PMID- 15296052 TI - Colour vision. PMID- 15296053 TI - Danny Bungaroo. PMID- 15296054 TI - Primary objectives. PMID- 15296055 TI - 'In the midst of difficulty lies opportunity'. PMID- 15296056 TI - Addressing increasing patient acuity and nursing workload. PMID- 15296057 TI - Delegation: a key function of nursing. PMID- 15296058 TI - Shifting boundaries. PMID- 15296059 TI - [The fan-shaped and ellipsoid bodies of the brain central complex are involved in the control of courtship behavior and communicative sound production in Drosophila melanogaster males]. AB - To elucidate the role of the fan-shaped and ellipsoid bodies (FB and EB) of the central complex of Drosophila melanogaster brain in the control of male courtship behavior and singing, we analyzed characteristics of the courtship behavior and parameters of the communicative sound signals accompanying it in wild type flies and in flies from 5 mutant strains with various anatomical defects in FB and EB. The following strains of flies were used for experiments: Canton S (wild type, the control), eboKS263 with defects only in EB and ebo1041, ceb849, ceb892 and cbdKS96 with both the FB and EB damaged in different manner. The data obtained suggest that the FB and EB are involved: 1) in maintenance of a high courtship activity level, 2) in the control of accuracy of male following movements when courting a female, 3) in the control of the form and stability of sound elements in courtship songs, 4) in the control of the rhythmic structure of courtship songs determined by the stability of the pacemakers, and 5) in setting up a correspondence between the current behavior and the external situation. PMID- 15296060 TI - [The second rhythm and brain slow wave potentials reflecting physiological condition in humans]. AB - In 4-hour experiments, dynamics of second brain potentials and the parameters of a conventionally negative wave, were studied. Along with growing fatigue, the amplitude of the second brain potentials was found to increase with a simultaneous increase in correlation relationships among the second oscillations registered in different areas which fact may serve as a diagnostic sign when controlling functional condition in humans. Velocity of a simple sensomotor response turned out to depend on coincidence with a current phase of the second brain potentials. The shortest time of response corresponded to responses against the background of a positive phase of the second oscillations. In fatigue, the dependence is enhanced. PMID- 15296061 TI - [Possible mechanisms of firing patterns reorganization of the neurons of the output nuclei basal ganglia]. AB - Previously proposed unitary modification rules and known experimental data were used for understanding possible mechanisms of reorganization of firing patterns of neurons in the output basal ganglia nuclei. According to the suggested mechanism, a switch from regular-spiking to bursting activity evoked by systemic inactivation of N-methyl-d-aspartate (NMDA) receptors or dopaminic receptors mainly depends on modifications of cortico-striatal synapses whereas the opposite effect of inactivation of the same receptors directly on the output basal ganglia cells is less effective. We hypothesized that some of the output basal ganglia nuclei neurons which can generate bursting discharges due to inactivation of NMDA and dopaminic receptors are glutamatergic or cholinergic cells. PMID- 15296062 TI - [Contribution of D2 dopamine receptors of the rat dorsolateral caudate nucleus to immunomodulation]. AB - The analysis of the immune response changes in Wistar rats under activation or blockade of D2 DA receptors has shown that electrolytic lesion of the dorsolateral caudate nucleus characterized by a high density of D2 DA receptors resulted in a decrease of the immune response to SRBC. At the same time, in rats with similar lesion stimulation of the immune reactions caused by a selective D2 agonist guinpirol (1.0 mg/kg) did not develop completely. Administration of haloperidol (2.0 mg/kg), the immune-inhibitory effect of which is associated with increasing serotoninergic system activity, to rats with impaired dorso-lateral caudate nucleus did not produce more expressed immunosuppression. However, the level of the immune response in sham-operated rats receiving haloperidol was significantly lower than that of animal with the destructed nucleus caudatus. Considering that qunmpirol-induced immunostimulation is related to the selective activation of the DA-ergic brain system, it is concluded that D2 DA receptors of the nucleus caudatus are involved in the mechanisms of immunostimulation, although D2 DA receptors of other brain structures may also impact this process. PMID- 15296063 TI - [Nitric oxide and carbon dioxide in neurotoxicity induced by oxygen under pressure]. AB - Hyperbaric oxygen (HBO2) causes CO2 retention in the brain that leads to the increase in cerebral blood flow (CBF) by poorly understood mechanisms. We have tested the hypothesis that NO is implicated in CBF-responses to hypercapnia under hyperoxic conditions. Alert rats were exposed to HBO2 at 5 ata and blood flow in the striatum measured by H2 clearance every 10 min. Acetazolamide, the inhibitor of carbonic anhydrase, was used to increase brain PCO2. CBF responses to acetazolamide administration (30 mg/kg, i.p.) were assessed in rats breathing air at 1 ata or oxygen at 5 ata with and without NOS inhibition (L-NAME, 30 mg/kg, i.p.). In rats breathing air, acetazolamide increased CBF by 34 +/- 7.4% over 30 min and by 28 +/- 12% over 3 hours while NOS inhibition with L-NAME attenuated acetazolamide-induced cerebral vasodilatation. HBO2 at 5 ata reduced CBF during the first 30 min hyperoxia, after that CBF increased by 55 +/- 19% above pre exposure levels. In acetazolamide-treated animals, no HBO, induced vasoconstricton was observed and striatal blood flow increased by 53 +/- 18% within 10 min of hyperbaric exposure. After NOS inhibition, cerebral vasodilatation in response to acetazolamide during HBO2 exposure was significantly attenuated. The study demonstrates that NO is implicated in acetazolamide (CO2)-induced cerebral hyperemia under hyperbaric oxygen exposure. PMID- 15296064 TI - [Amplitude and frequency characteristics of sympathetic nerve discharge in normotensive rats]. AB - Electrical discharges in the cervical sympathetic trunk and arterial blood pressure were recorded in chloralose-anaesthetized Wistar rats. Some experiments were performed in rats with impaired baroreceptor reflex. The efferent electrical activity was analysed in amplitude and frequency domain. It has been shown that arterial and cardio-pulmonary mechanoreceptors do not affect magnitude of the electrical activity. However, arterial and cardio-pulmonary baroreceptor reflex arc seems to be involved in frequency modulation of the efferent sympathetic activity. PMID- 15296065 TI - [Blood amino acid composition of poikilothermic vertebrate under prolonged starvation]. AB - Free amino acids in the blood plasma of lamprey (Lampetra fluviatilis) and frog (Rana temporaria) have been determined quantitatively for the periods of deprivation of the exogenous feed. The lamprey's total amino acid pool increased by 74% from November to April and reached the lower limit known for the mammals. The amount of free amino acids in frogs decreased by 40% in the spring as compared with the autumn values. The difference is accounted for by certain features of the living cycles of these animals. A more energetic proteolysis in the lamprey tissues as compared with that in the frog tissues has been confirmed by quantitative determining of leucine, isoleucine and valine in the blood of these animals. Apart from the above, alanine, glycine, lysine, threonine and, in certain periods, tyrosine have been found to be quantitatively significant in the plasma of both animal species. The composition and proportion of the amino acids in blood plasma of these animals are due to specific features of their metabolism and connected with the energy state of the liver cells under starvation. PMID- 15296066 TI - [Transcription factors C/EBP inhibition specifically affects synaptic plasticity in the snail's neuron]. AB - In our previous investigations it was found that nociceptive sensitization is followed by a long-term facilitation of synaptic transmission in command neuron of defence behavior. The facilitation is translation- and transcription dependent. Features of plasticity neurochemical mechanisms of different sensory inputs in the neural cells were determined. NMDA glutamate receptors, serotonin receptors, cAMP as well as serotonin-modulated protein SMP-69 are involved in mechanisms of induction of neural response facilitation in LPl 1 neuron evoked by chemical sensory stimulation of snail head. Protein kinase C is involved in synaptic facilitation-induction mechanisms from other sensory neural input- tactile receptors of snail "head". In this work, participation of C/EBP (CCAAT enhancing-binding protein) immediate early gene transcription factor was established in mechanisms of synapse-specific plasticity during sensitization in LPl 1 neuron in snail Helix lucorum. Specific binding with C/EBP oligonucleotides were used as C/EBP inhibitors. It was found that sensitization during intracellular oligonucleotide injection selectively suppressed synaptic facilitation in LPl 1 neuron responses evoked by chemical sensory stimulation of snail "head". At the same time, development of synaptic facilitation of the neuron responses evoked by tactile stimulation of snail head or foot was the same as in control sensitized snails. It seems that C/EBP transcription factor is selectively involved in mechanisms of synapse-specific plasticity of sensory input in LPl 1 neuron from snail head chemoreceptors. PMID- 15296067 TI - [Visual potentials in humans with multiple sclerosis evoked by checkboard pattern of different contrast with additional noise]. AB - A hypothesis is advanced of an effect of internal noise upon signal conduction along visual nerve in an early stage of multiple sclerosis. Visual evoked potentials in response to chess patterns against a homogeneous background and against a noise background were studied in healthy subjects and in patients with multiple sclerosis. Contrast between internal noise in the patients was found to be twice as great as in healthy subjects. PMID- 15296068 TI - [Effect of experimental elevation and decrease of thyroxine level on catalepsy in rats]. AB - Thyroid dysfunction is associated with mental disorders. The present study was aimed to reveal the effects of experimental decrease and increase of thyroxine level on expression of two types of extensive freezing: spontaneous and pinch induced catalepsy, in Wistar rat males. Chronic administration of thyroxine synthesis inhibitor, propylthiouracil (5 mg/kg/day, 28 days), markedly decreased plasma hormone level and at the same time produced a significant increase in percentage of spontaneously cataleptic animals and immobility time, but had no effect on the expression of pinch-induced catalepsy. On the contrary, chronic thyroxin (0.1 mg/kg/day, 28 days) treatment produced no effect on spontaneous catalepsy expression, although it significantly increased percentage of cataleptic animals and immobility time of pinch-induced catalepsy. The results suggest that both the thyroid hormone deficit and excess provoke catalepsy in rats but enhance different forms of freezing reaction. PMID- 15296069 TI - [The temperature and temperature gradients distribution in the rabbit body thermophysical model with evaporation of moisture from its surface]. AB - On created in laboratory heat-physical model of a rabbit body reflecting basic heat-physical parameters of the body such as: weight, size of a relative surface, heat absorption and heat conduction, heat capacity etc., a change of radial distribution of temperature and size was found across a superficial layer of evaporation of water from its surface, that simulates sweating, with various ratio of environmental temperature and capacity of electrical heater simulating heat production in animal. The experiments have shown that with evaporation of moisture from a surface of model in all investigated cases, there is an increase of superficial layer of body of a temperature gradient and simultaneous decrease of temperature of a model inside and on the surface. It seems that, with evaporation of a moisture from a surface of a body, the size of a temperature gradient in a thin superficial layer dependent in our experiments on capacity for heat production and environmental temperature, is increased and can be used in a live organism for definition of change in general heat content of the body with the purpose of maintenance of its thermal balance with environment. PMID- 15296070 TI - [Role of active transport of 5-hydroxyindoleacetic acid in serotonin metabolism in the brain]. PMID- 15296071 TI - [Blood pressure level in normotensive and spontaneously hypertensive rats under urethane anesthesia]. PMID- 15296072 TI - [The cardioamplitudography as a technique for studying inotropic effects of the myocardium]. PMID- 15296074 TI - Beta-carotene is accumulated, metabolized, and possibly converted to retinol in human breast carcinoma cells (MCF-7). AB - The aims of the present study were to investigate the uptake, accumulation, and metabolism of beta-carotene by the human breast carcinoma cell line MCF-7. Beta carotene uptake was time- and dose-dependent, and independent of cell polarity. Beta-carotene accumulation in cells was linear as a function of its concentration in medium (1.3-4.1 micromol/L). It was accompanied by increasing amounts of retinol, which accumulated in cells following a sigmoid pattern, and by other four putative metabolites. Beta-apocarotenals, epoxides, endoperoxides, retinal, retinoic acid, and retinyl esters were not detected in cell extracts. Beta carotene and its metabolites did not induce alterations in cell morphology or subcellular localization of epithelial mucins. Beta-carotene and retinol were released from cells that had previously accumulated beta-carotene, and were further incubated in beta-carotene- and retinol-free medium, but intracellular retinol content remained constant whereas beta-carotene decreased. In conclusion, beta-carotene added to culture medium in physiological concentrations (1-6 micromol/L) is taken up and metabolized in MCF-7 cells, and is possibly converted to retinol. PMID- 15296075 TI - Effect of chromium supplementation on blood glucose and lipid levels in type 2 diabetes mellitus elderly patients. AB - Intervention trials have shown the beneficial effects of chromium supplementation in type 2 diabetes (non-insulin-dependent diabetes mellitus). This study investigated the effects of chromium picolinate on elderly diabetic patients within a rehabilitation program. Thirty-nine diabetic subjects, average age 73 years (18 males and 21 females), undergoing rehabilitation following stroke or hip fracture, were recruited to participate in this study. An additional 39 diabetic patients constituted the control group. Along with standard treatment for diabetes, the study group received 200 microg of chromium twice a day for a three-week period. Blood samples, dietary intake, and anthropometric data were collected prior to and post-intervention. Throughout the study period, participants received a diet of approximately 1500 kcal/day. Significant differences in the fasting blood level of glucose compared to the baseline (190 mg/dL vs 150 mg/dL, p < 0.001) were found at the end of the study. HbA1c also improved from 8.2% to 7.6% (p < 0.01). Total cholesterol was also reduced from 235 mg/dL to 213 mg/dL (p < 0.02). A trend towards lowered triglyceride levels was also observed (152 mg/dL vs 136 mg/dL). We conclude that, in this population of elderly, diabetic patients undergoing rehabilitation, dietary supplementation with chromium is beneficial in moderating glucose intolerance. In addition, chromium intake appears to lower plasma lipid levels. PMID- 15296076 TI - Who and how many people are taking herbal supplements? A survey of 21,923 adults. AB - BACKGROUND: There is little information on the number and characteristics of adults taking herbal supplements and the relationship of this with other health and lifestyle factors. These were examined in the current study. METHODS: Information on herbal supplement use and health and lifestyle characteristics was obtained by postal questionnaire, sent to a sample of the adult population in Northwest England. RESULTS: In summer 2001, 70.5% (15,465/21,923) of questionnaires were returned. The mean age of responders was 49.8 years (SD 17.57) and 45.2% (6,986/15,465) were men. The percentage taking at least one herbal supplement was 12.8% (1,987/15,465). Users of herbal supplements were more likely to be younger, female, white, and to own their home. Herbal supplement use was not strongly associated with any health and lifestyle variables examined. Weak associations were found with physical activity, psychiatric caseness, and use of prescribed medications. The most common herbal supplement was evening primrose oil, taken by 7.7% (1,186/15,465) of respondents (12.7% of women and 1.1% of men). CONCLUSIONS: More than one in ten adults were taking herbal supplements, with evening primrose oil, the most common supplement, used mainly by women. Individual characteristics such as age, sex, ethnicity, and social class influenced the use of herbal supplements, but there was no evidence that this substituted for conventional medical care. The evidence base to support some popular herbal supplements is weak. Large well-designed trials are needed to quantify the value of herbal supplements to health and well-being. PMID- 15296077 TI - Folate status of adults living in the Canary Islands (Spain). AB - BACKGROUND AND AIM: Human studies support the hypothesized contribution of folate deficiency to carcinogenesis and vascular risk. We assess the nutritional folate status and its relationship to folate intake, smoking, alcohol consumption, oral contraceptive use, and multivitamin supplements. METHODOLOGY: A representative sample of 601 individuals from 18 to 75 years of age was selected from the participants in the Canary Islands Nutrition Survey. A food frequency questionnaire was administered. Serum and erythrocyte levels of folate were determined using a method of automated ionic capturing. RESULTS: Mean serum and red cell folate were 8.2 ng/mL and 214.3 ng/mL, respectively. Only one individual had serum folate below 3 ng/mL, and 21.7% showed moderate deficits (3-6 ng/mL); 10.7% of the sample had erythrocyte folate levels falling below 140 ng/mL, 61.3% between 140 and 240 ng/mL and the remaining 27.9% above 240 ng/mL. A positive significant association was observed between these two folate measurements, as well as between folate intake and each of these biomarkers (p < 0.001). Tobacco consumption was negatively correlated with folate status (p < 0.001). Alcohol consumption, oral contraceptive, and vitamin supplement use were not associated with serum and red cell folate levels. CONCLUSIONS: Even though nutritional folate status can be considered minimally acceptable, it may reflect the low level of fruit and vegetable consumption within the Canary Islands population. PMID- 15296078 TI - Concentrations of ascorbic acid in the plasma of pregnant smokers and nonsmokers and their newborns. AB - A cross-sectional study was carried out to determine the differences in vitamin C status of Brazilian pregnant women smokers and nonsmokers and their respective newborn babies, and to assess the prevalence of hypovitaminosis C among these two groups of women. The study involved 127 pregnant women, 40 pregnant smokers and 87 pregnant nonsmokers, admitted to a maternity hospital in Sao Paulo, Brazil. Data concerning the pregnant women's socioeconomic, demographic, obstetric, and nutritional characteristics were collected, as well as data concerning the newborns' anthropometry and Apgar scores. A strongly significant correlation (p < 0.001) was found between the concentrations of ascorbic acid (AA) in both pregnant smokers (r = 0.77) and pregnant nonsmokers (r = 0.61) and their respective umbilical cords. The mean umbilical AA concentration was significantly higher than the meanAA concentration in pregnant women (92.05 +/- 41.13 vs. 33.39 +/- 18.25 micromol/L, p < 0.001). It was observed that the mean AA was significantly lower for the newborns (p = 0.03) and pregnant women (p = 0.02) from the smoking group. Forty percent (40%) of the smokers and 27% of the nonsmokers presented hypovitaminosis C. We suggest an increase in the consumption of fruits and vegetables by pregnant women, especially the smokers. PMID- 15296079 TI - Antioxidant status of red blood cells and liver in hypercholesterolemic rats fed hypolipidemic spices. AB - Animal studies were carried out to examine the beneficial influence of known hypolipidemic spice principles--curcumin, capsaicin, and garlic--on the antioxidant status of red blood cells and liver under induced hypercholesterolemic conditions. Groups of experimental rats rendered hypercholesterolemic were maintained on curcumin (0.2%)/capsaicin (0.015%)/garlic (2.0% dry powder)-containing diets for eight weeks. Erythrocytes isolated at the end of the study were analyzed for intracellular antioxidant molecules and antioxidant enzymes. Intracellular thiols and glutathione content in red blood cells were significantly depleted (by about 35%) in hypercholesterolemic rats. This depletion in intracellular thiols and glutathione was effectively countered by dietary spice principles - curcumin, capsaicin, and garlic. Glutathione reductase activity that was lowered in hypercholesterolemic conditions (by 25%) was completely countered by dietary spice principles and garlic. Activities of glutathione transferase, glutathione peroxidase, and catalase in erythrocytes remained unchanged under hypercholesterolemic conditions. Although hemoglobin levels of erythrocytes were not affected, methemoglobin concentration was significantly increased in hypercholesterolemic rats. This alteration was partially countered by dietary spice principles. Significant fall in hepatic total thiols in the hypercholesterolemic situation was partially corrected by dietary spice treatment. Similarly, the lowered activities of hepatic antioxidant enzymes--glutathione reductase, glutathione-S-transferase, catalase, and superoxide dismutase--in hypercholesterolemic rats were effectively countered by the dietary spices treatment. PMID- 15296081 TI - High intake of magnesium in relation to the ruminal transmural potential difference and magnesium absorption in wethers. AB - High potassium (K) intakes are known to decrease magnesium (Mg) absorption in ruminants by increasing the transmural potential difference (PDt, serosal side = positive). High Mg intakes are known to increase the amount of Mg absorbed, which may be explained by increasing the ruminal Mg concentration, but an effect on the PDt cannot be excluded. The objective of this study was to determine whether or not Mg intake affects the PDt. In a 3 x 3 Latin square design, six ruminally fistulated wethers were fed a low-Mg, low-K ration (3.88 g Mg/kg dry matter (DM); 30.7 g K/kg DM), a high-Mg, low K-ration (16.79 g Mg/kg DM; 30.7 g K/kg DM), and a low-Mg ration high-K (3.88 g Mg/kg DM or 62.1 g K/kg DM). When compared with the low-Mg, low-K ration, the high-Mg, low-K ration raised the absolute apparent Mg absorption (g/day) by 421% and the low-Mg, high-K ration decreased it by 20%. The intake of extra K produced a significant increase in the PDt. The intake of extra Mg did not change the PDt across the rumen wall but produced a significant increase of the ruminal Mg concentrations. On the basis of the individual values for three rations, the mean post feeding ruminal Mg concentrations were found to be unrelated to the PDt (Pearson's r = -0.329, p = 0.183, n = 18). Thus, it is concluded that the observed increase in Mg absorption after a high Mg intake can be explained by an increase in the ruminal Mg concentration rather than by a change in PDt. PMID- 15296080 TI - Jurkat cells respond to biotin deficiency with increased nuclear translocation of NF-kappaB, mediating cell survival. AB - Members of the NF-kappaB family of transcription factors cause transcriptional activation of anti-apoptotic genes. Here we determined whether survival of biotin deficient cells is mediated by nuclear translocation of NF-kappaB. Human T (Jurkat) cells were cultured in biotin-deficient or biotin-supplemented media; nuclear translocation of NF-kappaB was stimulated with phytohemagglutinin and phorbol-12-myristate-13-acetate. Nuclear abundance of two members (p50 and p65) of the NF-kappaB family was greater in biotin-deficient compared to biotin supplemented cells; this effect was mediated by phosphorylation of IkappaBalpha. The nuclear enrichment of p50 and p65 in biotin-deficient cells was associated with transcriptional activation of NF-kappaB-depedent genes such as the tumor suppressor gene p53 and the anti-apoptotic gene Bfl-1/A1. Biotin-deficient cells exhibited smaller activities of the apoptotic enzyme caspase-3 in response to treatment with tumor necrosis factor alpha, and decreased cell death in response to serum starvation compared to biotin-supplemented cells. These findings suggest that NF-kappaB mediates survival of biotin-deficient cells. PMID- 15296082 TI - Risk assessment and risk management of vitamins and minerals. AB - OBJECTIVE OF WORKSHOP: The EANS workshop held in 1998 examined the various approaches to determine requirements and safety of vitamins and minerals. The methodology used and approaches taken on both sides of the Atlantic provided the focus of the event. Over three years later, and with risk assessment much advanced, the progress made was reviewed and inadequacies as well as limitations were defined. In addition, this workshop looked beyond assessment to the broader context in which nutrition science operates. What are the particular problems facing risk managers in the light of risk assessment conclusions? To what extent can nutrition research provide the answers that risk managers require and should the nutrition research agenda be shaped by the needs of the policymaker? PMID- 15296083 TI - The administration of a multivitamin/mineral fortified dairy product improves folate status and reduces plasma homocysteine concentration in women of reproductive age. AB - BACKGROUND AND OBJECTIVES: Folate deficiency during the periconceptional period is related to the occurrence and recurrence of neural tube defects. The aim of the study was to assess whether the administration of folic acid and other vitamins and minerals as a fortified dairy product (400 microg per day of folic acid) improves the folate status in women of reproductive age. DESIGN AND METHODS: Plasma and red blood cell folate, plasma vitamin E, B12, total plasma homocysteine, plasma lipid profile, and serum ferritin and transferrin levels were investigated in 31 healthy nonpregnant women receiving 500 mL/day of the fortified dairy product for eight weeks. RESULTS: The women showed a significant increase in plasma levels of folate and vitamin B12 concentrations after four and eight weeks of supplementation. Moreover, we observed an increase in red blood cell folate concentration during the period of the study. Simultaneously, total plasma homocysteine levels decreased significantly during the intervention period. CONCLUSIONS: The regular consumption of a folic acid and other vitamins (mainly vitamins B6 and B12) and minerals in a fortified dairy product improves folate status and reduces total plasma homocysteine concentration in healthy women of childbearing age. PMID- 15296084 TI - New 3-[4-(2,3-dihydro-14-benzodioxin-5-yl)piperazin-1-yl]-1-(5-substituted benzo[b]thiophen-3-yl)propanol derivatives with dual action at 5-HT(1A) serotonin receptors and serotonin transporter as a new class of antidepressants. AB - Compounds derived from 2,3-dihydro-(1,4-benzodioxin-5-yl)piperazine and benzo[b]thiophene with different substituents in 5 position (H, F, NO2, NH2, CH3 and OH) have been synthesized in order to obtain new dual antidepressant drugs. The final compounds were evaluated for in vitro 5-HT(1A) receptor affinity and serotonin reuptake inhibition by radioligand assays. Compounds 1-(5 nitrobenzo[b]thiophen-3-yl)-3-[4-(2,3-dihydro-1,4-benzodioxin-5-yl)piperazin-1 yl]propan-1-ol (4c) (Ki = 6.8 for 5-HT(1A) receptor and Ki = 14 for 5-HT transporter) and 1-(5-hydroxybenzo[b]thiophen-3-yl)-3-[4-(2,3-dihydro-1,4 benzodioxin-5-yl)piperazin-1-yl] propan-1-ol (4f) (Ki = 6.2 for 5-HT(1A) receptor and Ki = 18.2 for 5-HT transporter) showed the best results for both activities. PMID- 15296085 TI - Antimycobacterial arylidenecyclohexanones and related Mannich bases. AB - Several series of 2-arylidenecyclohexanones and related Mannich bases as well as various 2,6-bis(arylidene)cyclohexanones were evaluated against Mycobacterium tuberculosis H37Rv. Using a concentration of 12.5 microg/ml, nearly half of the unsaturated ketones inhibited the growth of the microorganism by 21-66% while all of the Mannich bases achieved 99% or greater inhibition. The relative hydrophobicities and widths of the molecules may have been contributing factors as to whether bioactivity was present or absent. Two of the Mannich bases demonstrated noteworthy potencies towards Mycobacterium avium. The conclusion was drawn that Mannich bases of 2-arylidenecyclohexanones represent a novel class of antimycobacterials. PMID- 15296086 TI - Absolute configuration of chloro-bisabolene sesquiterpene. AB - The crystal structure of a cholor-bisabolene sesquiterpene has been determined for the absolute configuration. Its structure was elucidated as (-) (1R,2R,3R,4R,6S,8S,10S)-chloro-2,10-diacetoxy-1,8-diangeloyloxy-3-hydroxy-11 methoxy-bisabol-7(14)-ene. The chlorine atom at C4 is axial in the cyclohexane ring. The molecule shows a stable chair conformation, and crystallizes in the monoclinic space group P2(1) with one molecule in the asymmetric unit. PMID- 15296087 TI - New chimera proteins for fluorescence correlation spectroscopy. AB - A new class of chimera proteins has been developed. They are ideally suited for detection by fluorescence correlation spectroscopy (FCS), a new technology to analyze molecular interactions. The molecular structure of these chimera proteins consists of four domains: a N-terminal (His)6-tag for affinity chromatography followed by an eight amino acid epitope for immunodetection, a polypeptide affinity domain (ADF) for target specific interaction and a C-terminal Green Fluorescent Protein (GFPuv) for reporting of interaction with the target by FCS. We designed, prepared and characterized a prototype of ADF-GFP proteins capable of specific interaction with DNA fragments bearing nuclear factor (NF)-kappaB sites. ADF NF-kappaB p50 and a non-DNA-binding deletion mutant (p35) combined with GFPuv were inserted in a procaryotic vector and expressed in E. coli. Following affinity purification the fluoroproteins p50-GFPuv and p35-GFPuv were employed in specific protein-protein and protein-DNA interaction studies. FCS analysis as well as EMSA showed that p50-GFPuv revealed a fully functional ADF. We present a model for the preparation of GFP fusion proteins capable of specific interaction with proteins, lipids or nucleic acids. The rational design allows any polypeptide fragment to be incorporated into the chimeric protein. So a new series of bio-molecules with different binding specificities and assays can be developed. PMID- 15296088 TI - Quantification of allantoin in various Zea mays L. hybrids by RP-HPLC with UV detection. AB - A RP-HPLC method for quantification of allantoin in silk of fifteen maize hybrids (Zea mays L., Poaceae) was described. Following extraction of the plant material with an acetone-water (7:3, VN) mixture, filtration and dilution, the extracts were analyzed without previous chemical derivatization. Separation and quantification were achieved using an Alltech Econosil C18 column under isocratic conditions at 40 degrees C. The mobile phase flow (20% methanol--80% water with 5 mM sodium laurylsulfate added at pH 2.5, adjusted with 85% orthophosphoric acid; pH of water phase was finally adjusted at 6.0 by addition of triethylamine) was maintained at 1.0 mL/min. Column effluent was monitored at 235 nm. This simple procedure afforded efficient separation and quantification of allantoin in plant material, without interference of polyphenols or other plant constituents of medium to high polarity, or similar UV absorption. Our study revealed that the silk of all investigated maize hybrids could be considered relatively rich in allantoin, covering the concentration range between 215 and 289 mg per 100 g of dry plant material. PMID- 15296089 TI - HPTLC densitomeric determination of justicidin B in Linum in vitro cultures. AB - A convenient, sensitive and accurate method for the separation and determination of justicidin B was created. The values were calculated by linear calibration under standard conditions. The method can be applied for the quantification of justicidin B in plants or in vitro cultures. PMID- 15296090 TI - A simple method for determination of surface volume mean diameters of oil in water submicron emulsions. AB - Plots of the droplet size parameter, alpha = (pi(n)D / lambda) versus the total scattering coefficient or specific turbidity, K, are sinusoidal curves. It was noticed that, along one branch of the K-alpha curve (up to 1.1 microm for relative refractive indices (rr(I)) < 1.2), the wavelength dependence on turbidity; i.e. the extent to which light at different wavelengths is scattered showed a progressive decrease with increase in droplet diameter. It was in fact found that when the logarithm of the relative standard deviations (RSD's) in specific turbidities at four different wavelengths (for a particular droplet diameter and relative refractive index) was plotted as a function of that diameter, a straight line resulted. This relationship provides a valuable new method for estimating the surface volume mean droplet size, D(vs), for submicron emulsions by means of simple spectro-turbidimetry. PMID- 15296091 TI - Physical transformation of niclosamide solvates in pharmaceutical suspensions determined by DSC and TG analysis. AB - This study reports the preparation of four niclosamide solvates and the determination of the stability of the crystal forms in different suspension vehicles by DSC and TG analysis. Thermal analysis showed that the niclosamide solvates were extremely unstable in a PVP-vehicle and rapidly changed to monohydrated crystals. A suspension in propylene glycol was more stable and TG analysis showed that crystal transformation was less rapid. In this vehicle, the crystals transformed to the anhydrate, rather than the monohydrate, since the vehicle was non-aqueous. The TEG-hemisolvate was the most stable in suspension and offered the best possibility of commercial exploitation. PMID- 15296092 TI - Statistical method for evaluation of dissolution stability in the formulation development of solid dosage forms: tablets of amonafide. AB - A statistical method for the evaluation of the dissolution stability results and for selecting the most stable formulation within a solid dosage form development is discussed. Three types of tablets of an antineoplastic drug, amonafide, stored at a relative humidities (RH), 45% and 75%, were used. The drug release from tablets was tested before and after storage. The experimental data were statistically fitted to empirical model equations. Furthermore, the best mathematical fit was the statistical comparison of the residuals. From the selected model equation, time-dependent dissolution (Q45 and DE45) and dissolved quantity-dependent parameters (t70, t100 and MDT) were calculated. A useful parameter to present and evaluate the results obtained in comparative stability studies was defined: the Modification Factor (MF). It allowed the selection of the most stable formulation in the easiest and fastest way: the most stable formulation should present the smallest modification of the studied characteristics, in other words, the smallest MF value. In this way, tablets II (manufactured by wet granulation and with Emcompress as main excipient) showed the greater dissolution stability of the three types of tablets studied. Amonafide tablets must be packaged in impermeable containers, since the environmental relative humidity strongly modifies their dissolution characteristics. PMID- 15296093 TI - In vitro cytotoxicity of berberine against HeLa and L1210 cancer cell lines. AB - Previous studies on anti-cancer activity of protoberberine alkaloids against a variety of cancer cell lines were extended to human uterus HeLa nad murine leukemia L1210 cell lines. Cytotoxicity was measured using in vitro techniques and cell morphology changes were examined by light microscopy in both cytostatic and cytocidal concentration ranges. The IC50 was found to be less than 4 microg/ml, a limit put forward by NCI for classification of the compound as a potential anti-cancer drug. The microscopy examination indicated that at cytocidal concentrations the HeLa and L120 cells died apoptotically. The comparative analysis revealed that berberine belongs to the camptothecin family of drugs characterized by the ability to induce DNA topoisomerase poisoning and hence apoptotic cell death. Although the cytotoxic potency of berberine was found to be several orders of magnitude lower compared to camptothecin, its significance may increase in future in view of the lack of unwanted side effects characteristic for camptothecin compounds currently in clinical use for treatment of cancer. PMID- 15296094 TI - Stachyose extract from Rehmannia glutinosa Libosch. to lower plasma glucose in normal and diabetic rats by oral administration. AB - The hypoglycemic effects of water extract and stachyose extract (Part III) from Rehmannia glutinosa Libosch. were investigated in this paper by oral administration to normal, glucose- and adrenaline-induced hyperglycemic and alloxan-induced diabetic rats. The results showed that Part III had the effect of lowering fasted plasma glucose level and partially preventing hyperglycemia induced by glucose (2.5 g x kg(-1), i.p.) and adrenaline (300 microg x kg(-1), i.p.), respectively, but no obvious dose-dependent effect was found when it was administered at the doses of 100, 200 and 400 mg x kg(-1) for 6 days, i.g. In alloxan-induced diabetic rats, Part III (200 mg x kg(-1) for 15 days, i.g.) gave a significant decrease in blood glucose level. The results suggested that Part III, which is mainly composed of stachyose from Rehmannia glutinosa Libosch., had a significant hypoglycemic effect in glucose- and adrenaline-induced hyperglycemic and alloxan-induced diabetic rats. PMID- 15296095 TI - Effect of scoparia dulcis (Sweet Broomweed) plant extract on plasma antioxidants in streptozotocin-induced experimental diabetes in male albino Wistar rats. AB - Clinical research has confirmed the efficacy of several plants in the modulation of oxidative stress associated with diabetes mellitus. Scoparia dulcis plant extract is tried for prevention and treatment of diabetes mellitus induced experimentally by streptozotocin injection. A single dose of streptozotocin (45 mg/kg body weight) produced decrease in insulin, hyperglycemia, increased lipid peroxidation (Thiobarbituric reactive substances and lipid hydroperoxides) and decreased antioxidant levels (vitamin C, vitamin E, reduced glutathione, ceruloplasmin). Oral administration of an aqueous extract of Scoparia dulcis plant (200 mg/kg body weight) for 6 weeks to diabetic rats significantly increased the plasma insulin and plasma antioxidants and significantly decreased lipid peroxidation. The effect of Scoparia dulcis plant extract at 200 mg/kg body weight was better than that of glibenclamide, a reference drug. PMID- 15296096 TI - Possible mechanism(s) for relaxant effects of Foeniculum vulgare on guinea pig tracheal chains. AB - In a previous study the relaxant (bronchodilatory) effect of Foeniculum vulgare on isolated guinea pig tracheal chains was demonstrated. To study mechanisms responsible for this effect the present study evaluated the inhibitory effect of this plant on contracted tracheal chains of guinea pig. The relaxant effects of aqueous and ethanol extracts and an essential oil from Foeniculum vulgare were compared to negative controls (saline for aqueous extract and essential oil and ethanol for ethanol extract) and a positive control (diltiazem) using isolated tracheal chains of the guinea pig precontracted by 10 microM methacholine (group 1) and 60 mM KCl (group 2, n = 7 for each group). In the group 1, experiments diltiazem, ethanol extract, and essential oil from Foeniculum vulgare showed a significant relaxant effect on methacholine induced contraction of tracheal chains compared to those of negative controls (p < 0.05 to p < 0.001). In addition the effect of the ethanol extract was significantly greater than that of diltiazem (p < 0.001). However, the aqueous extract did not show any relaxant effect in group 1. In the group 2 experiments, only diltiazem showed a significant relaxant effect on KCl induced contraction of tracheal chains (p < 0.001). The relaxant effects of ethanol extracts and essential oil obtained in the group 2 experiments were significantly lower than those in group 1 (p < 0.05 to p < 0.001). These results confirm the bronchodilatory effects of ethanol extract and essential oil from Foeniculum vulgare. However with regard to the effect of KCl on calcium channels, the results indicated that the inhibitory effect of ethanol extracts and essential oil from Foeniculum vulgare on calcium channels is not contributing to their relaxant (bronchodilatory) effects on guinea pig tracheal chains. However the results suggest a potassium channel opening effect for this plant, which may contribute on its relaxant effect on guinea pig tracheal chains. PMID- 15296097 TI - Chrysanthemum morifolium attenuated the reduction of contraction of isolated rat heart and cardiomyocytes induced by ischemia/reperfusion. AB - The present study was aimed to investigate the effect of Chrysanthemum morifolium Ramat. (CM) on isolated rat heart and ventricular myocytes during ischemia/anoxia and reperfusion/reoxygenation. The ischemia/reperfusion injury was induced by ligation the left artery descending coronary of isolated rat heart for 30 min followed by 30 min reperfusion with Langendorff equipment. Cell contraction in enzymatically isolated ventricular myocytes was determined by a video tracking system. The results showed CM (0.25 g/L to 1.0 g/L) increased left ventricular developed pressure (LVDP), +/- dp/dt(max), LVDP x HR and coronary flow (CF) and decreased heart rate (HR) in dose dependent manner. CM (0.5 g/L) attenuated the reduction of LVDP, +/- dp/dt(max) and CF caused by ischemia/reperfusion. CM (0.25 g/L to 1.0 g/L) increased peak velocity of cell shortening/relengthening (+/- dL/dt(max)) and contraction amplitude (dL) of isolated ventricular myocytes in a dose-dependent way under control condition, but without significant effect on end diastolic cell length (L0). Under anoxia 5 min followed by 10 min reoxygenation, CM attenuated the reduction in contractile parameters. The results suggest that CM processes cardioprotective effect during ischemia/anoxia and reperfusion/reoxygenation in the isolated rat heart and the ventricular myocytes. PMID- 15296098 TI - A novel antifungal pyrrole derivative from Datura metel leaves. AB - Phytochemical investigation of the leaves of Datura metel Linn. led to the isolation of a new pyrrole derivative 1 which was characterised as 2beta-(3,4 dimethyl-2,5-dihydro-1H-pyrrol-2-yl)-1'-methylethyl pentanoate on the basis of spectral data analyses and chemical reactions. Compound 1 was endowed with antifungal activity and its MIC was found to be 87.5 microg/ml. Two proteins having molecular weights of 42 and 58 kD of Aspergillus fumigatus are potential targets for compound 1. PMID- 15296099 TI - High gene expression of the mutant adenovirus vector, both in vitro and in vivo, with the insertion of integrin-targeting peptide into the fiber. AB - In the present study, a first-generation adenovirus (Ad) vector was modified with the RGD peptide inserted into the fiber. The insertion of an integrin-targeting sequence into the Ad vector notably enhanced the luciferase expression in the Coxsackie virus and Adenovirus Receptor-deficient A2058 and B16BL6 melanoma cells. The results of an in vivo study with tumor-bearing mice also showed that Ad-RGD-Luc had enhanced gene expression in many organs and in the B16BL6 tumor compared to that induced by a conventional Ad vector after intravenous injection. PMID- 15296100 TI - The efficiency of the benzothiazole APB, the echinocandin micafungin, and amphotericin B in fluconazole-resistant Candida albicans and Candida dubliniensis. AB - This study presents the efficiency of the experimental antifungal agents 6-amino 2-n-pentylthiobenzothiazole (APB) and the echinocandin micafungin, and amphotericin B against fluconazole-resistant Candida albicans and Candida dubliniensis (MIC95 for fluconazole > 64 mg/l). The benzothiazole APB was less active against C. albicans and C. dubliniensis (MIC80 = 8 - 32 mg/l, MIC95 = 16 - 64 mg/l) than amphotericin B, which was efficient in a concentration range from 0.125 to 2 mg/l. However, the efficiency of micafungin was very high with MIC80, and MIC100 < or = 0.031 mg/l. PMID- 15296101 TI - Left ventricular hypertrophy and hypertension. AB - BACKGROUND: Hypertension is the commonest non-communicable disease in our environment. Left ventricular hypertrophy is quite prevalent among this category of patients. METHODS: An endeavour was therefore made to review the literature on left ventricular hypertrophy in hypertension using MEDLINE and bibliographic searches for English language studies. RESULTS: The epidemiology, pathogenesis, molecular mechanism, pathophysiology, pathology and diagnosis of left ventricular hypertrophy in hypertension were presented. Effect of left ventricular hypertrophy on morbidity and mortality and the probable consequence of its regression were also discussed. CONCLUSION: Left ventricular hypertrophy in hypertension can be detected through an avalanche of diagnostic modalities and criteria. It carries not just management but also prognostic implications. PMID- 15296102 TI - Postgraduate dental education in Nigeria: professional knowledge self assessed in relationship to skills among resident dental surgeons in Nigerian teaching hospitals. AB - BACKGROUND: Improvement in any professional calling demands periodic assessment. The purpose of this study was to assess the professional knowledge in relation to skills of the resident dental surgeons in Nigeria in an attempt to contribute to their training needs analysis. METHOD: A 33-item, self-evaluation one-page questionnaire focussing on their knowledge and skills in the different aspects of clinical dentistry was distributed to the residents in Nigerian teaching hospitals. Sixty-six residents consisting of 38 (57.6%) males and 28 (42.4%) females with age range of 27-46 years (mean 31.3 +/- 3.4 SD) completed and returned the questionnaires giving a 91.7% response rate. RESULTS: The overall means for knowledge and skills of the residents were 70.1 +/- 13.8 (SD) and 67.3 +/- 12.0 (SD) respectively. Residents whose claims of professional knowledge and skills were found below average were 30 (45.5%) and 29 (43.9%) respectively. No statistically significant sex differences were found in the claimed knowledge and skills of the residents as well as in terms of their professional status (P > 0.05). The lowest grand means for knowledge and skills were found for orthodontics/paedodontics. CONCLUSION: The study has shown that the resident dental surgeons in Nigeria claimed better professional knowledge than skills and needed most attention in orthodontics/paedodontics. PMID- 15296103 TI - A preliminary review of extracapsular cataract extraction with posterior chamber intraocular lens implantation in Kaduna, Nigeria. AB - BACKGROUND: Extracapsular cataract extraction with posterior chamber intraocular lens implantation came into vogue recently in many ophthalmic centres in Nigeria for the management of cataract and its accompanying aphakia. Evaluation of this procedure in the hands of surgeons who converted newly to extracapsular cataract extraction with intraocular lens implant microsurgery was reviewed. This is with the view to assess and improve on their surgical skill considering their delayed take off due to lack of facility in the centre. METHOD: A retrospective study of the first 48 patients (50 eyes) who had extracapsular cataract extraction with posterior chamber intraocular lens implantation between September 1999 and December 2000 was carried out. RESULT: Forty-six patients had extracapsular cataract extraction with posterior chamber intraocular lens implant in one eye only, while 2 had the procedure in both eyes. Fifty-six (56%) percent of the patients were above 55 years old. Male to female ratio was 1.8:1.0. The preoperative visual acuity in 92% of eyes was equal to or less than 3/60. Six (6%) percent of the eyes had good visual outcome (6/6-6/18), 70% had borderline (6/24-6/60) on the 1st postoperative day. Visual outcome improved steadily with the passage of time as the immediate postoperative complications resolved. The main intra-operative complications were large anterior capsular tags (35.7%) and cortical lens remnants (50%); while striae keratopathy and corneal oedema (54.5%) constituted the main postoperative complications. CONCLUSION: Good visual outcome of greater than 80% with available correction is possible in the early postoperative period. Improved surgeons skill through re-training and refresher courses will guarantee a good outcome. Finally, microsurgical facilities must be put in place before a conversion course or training. PMID- 15296104 TI - Maternal mortality in the University of Port Harcourt Teaching Hospital, Port Harcourt in the last year before the new millennium. AB - BACKGROUND: Maternal death in developing countries is still high and the causes are multifactorial. The unbooked primigravidae with severe pre eclampsia/eclampsia constitute a high risk group. METHODS: The data from case notes of all maternal deaths which occurred at the University of Port Harcourt Teaching Hospital (UPTH), Port Harcourt between 1st of January and 31st of December 1999 formed the basis of this study. Case notes of all deaths were stored in the office of the head of department of obstetrics and gynaecology as soon as they occurred. Information was extracted from the case files at the end of the year. The total number of deliveries was obtained from the registers kept in labour and isolation wards respectively. RESULTS: There were 45 maternal deaths; 40 (88.9%) among the unbooked and 5 (11%) among the booked mothers constituting a maternal mortality ratio of 23, 121.4 and 339.7 per 100,000 deliveries respectively. The combined mortality ratio was 2735.6 per 100,000 deliveries. Fifteen (37.5%) unbooked primigravidae died of severe pre eclampsia/eclampsia. A total of 1645 mothers delivered, 1472 (89.5%) were booked while 173 (10.5%) were unbooked with the hospital. CONCLUSION: Severe pre eclampsia/eclampsia, haemorrhage and sepsis were the major causes of death. The high maternal mortality was common among the unbooked primigravidae who usually present late with pre-eclampsia/eclampsia. More research into the causes and management of pre-eclampsia/eclampsia are needed to reduce the high maternal mortality associated with it. Lack of antenatal care is also a high risk factor for maternal mortality. PMID- 15296105 TI - Pattern of substance use among secondary school students in Rivers State. AB - BACKGROUND: The problem of substance use in our secondary schools with cultism has negatively affected learning [corrected]. The aim of this study was to determine the commonly used substances and the factors that influence their use in these adolescents. METHODOLOGY: One thousand and forty-nine students of four secondary schools in Port Harcourt were screened using a self administered 117 items-substance use questionnaires. RESULTS: Males constituted 57.2% and females, 42.8% of the study population. Eighty-seven percent were using at least one substance, at the time of the survey while 3% were past users in the last one year. Ten percent had never used any of the substances. The substances commonly used included alcohol 65%, kolanut 63.1%, cigarettes 61%, paracetamol 41.5%, butazolidine 39.3%, pemoline 28% and cannabis 26%. Others include tetracycline 25.7% and ampicillin 24.3% and valium 24%. Those that were least used included, heroine, cocaine, latex, petrol, pethidine and ativan. The mean age of onset for alcohol was 4 years, kolanut 8 years, cigarettes 11 years, paracetamol, tetracycline, valium, cannabis was, 12 years, lexotan and ativan, was 12.5 years and, latex inhalation, petrol, pethidine, ativan, cocaine and heroine was 13.17 years. Males predominated in all the substances used. CONCLUSION: The use of substances/drugs among our youths is assuming a dangerous dimension, hence calls for immediate and enduring measures to curb this disturbing trend of abuse of substances. PMID- 15296106 TI - Malaria parasite density and splenic status by ultrasonography in stable sickle cell anaemia (HbSS) children. AB - BACKGROUND: To determine the relationship, if any, between malaria parasitaemia, parasite density and presence/size of the spleen, using abdominal ultrasonography in stable sickle cell anaemia subjects aged 6 to 15 years. METHODS: A prospective study of one hundred consecutive sickle cell anaemia (SCA) and 100 matched healthy HbAA controls aged 6 to 15 years was undertaken. The presence of malaria parasite, and parasite density were determined using thick blood film. Splenic status was determined by using abdominal ultrasound. None of the children was symptomatic for malaria. RESULTS: The prevalence of autosplenectomy and splenomegaly were 20% and 27% respectively in SCA subjects compared to 0% and 4% respectively in HbAA controls. Thirty percent and 34% of the SCA and controls respectively had malaria parasitaemia. In SCA subjects, the parasite density ranged from 33 to 4000 per microl with a mean of 1071.10 +/- 895.5 per microl. In HbAA controls, the parasite density ranged from 180 to 5150 per microl with a mean of 1759 +/- 1382.87 per microl. The difference in parasite density between SCA subjects and HbAA was significant p<0.05. The parasite densities were relatively higher among SCA with splenomegaly and normal spleen sizes compared to SCA subjects with autosplenectomy. CONCLUSIONS: The prevalence of malaria parasitaemia is lower in healthy SCA subjects than in HBAA controls. Sickle cell anaemia subjects have lower malaria parasite density. Autosplenectomy may be a positive adaptation in SCA subjects with effective innate immunity to malaria. PMID- 15296107 TI - Perinatal outcome among elderly nulliparae at the University of Port Harcourt Teaching Hospital. AB - BACKGROUND: Maternal and perinatal health is influenced by sociodemographic factors including maternal age, parity, social class and ethnicity. Elderly nulliparity is considered traditionally, to be high risk and has not been evaluated in our centre. The objective of our study was to determine the perinatal outcome in the elderly nullipara at the University of Port Harcourt Teaching Hospital. METHODS: This was a two-year retrospective case controlled study. The study population consisted of all nulliparae aged 35 years and above who delivered in the hospital during the period and the control group consisted of all nulliparae less than 35 years of age. The chi2-test was used for comparison of both groups and statistical significance set at p values of P < or = 0.05. RESULTS: Fifty-nine (2%) of 2967 women who delivered during the period were elderly nulliparae. They accounted for 4.8% of all nulliparae. Compared to other nulliparae, there was no significant difference in the rate of preterm delivery (21.8% vs. 25.8%; p = 0.6), postterm delivery (9.1% vs. 17.3%; p = 0.2), caesarean section rate (41% vs. 32.8%; p = 0.3), low birth weight (5.2% vs. 11.6%; p = 0.2), macrosomia (10.3% vs. 5.7%; p = 0.2), still birth rate (0% vs. 3.2%), birth asphyxia (31% vs. 25.4%; p = 0.4), and sex ratios of babies. There were no congenital malformations in both groups. CONCLUSION: Elderly nulliparae are not at increased risk of adverse perinatal outcome compared to younger nulliparae in our hospital. PMID- 15296108 TI - Awareness and acceptability of prenatal diagnosis of sickle cell anaemia among health professionals and students in North Eastern Nigeria. AB - BACKGROUND: The genetic control of sickle cell anaemia through prenatal diagnosis was recently introduced in Nigeria. Its acceptability will depend on some factors such as religious, political and social attitudes of the populace. The aim of the study was to determine the awareness and acceptability of parental diagnosis of sickle cell anaemia among health professionals and students in North Eastern Nigeria. METHOD: Structured questionnaires were administered to health professionals and students. RESULTS: Of 353 respondents interviewed, 279 (79%) were from Borno State, while 36 (10.2%) and 38 (10.8%) were from Yobe and Bauchi States respectively. Two hundred and four (57.8%) practised Islam while 149 (42.2%) were Christians. Ninety-five (26.9%) of the respondents were doctors, 17 (4.8%) pharmacists 37 (10.5%) technologists while 107 (30.3%) were nurses, 82 (23.2%) medical students and 15 (4.2%) physiotherapists. Two hundred and fifty seven (72.8%) had heard about Prenatal Diagnosis (PND) of Sickle Cell Anaemia (SCA). Fifty three percent (187) of the respondents would not like to terminate pregnancy by abortion if prenatal diagnosis confirmed sickle cell anaemia (SCA) in first trimester with significantly more Christians saying no to abortion. Only 50 (14.2%) of the respondents knew where facilities for prenatal diagnosis are obtainable in Nigeria whereas 85.8% (303) did not. CONCLUSION: Religion may be a major factor militating against acceptability of prenatal diagnosis of SCA in North Eastern part of Nigeria. The awareness of where facilities for prenatal diagnosis are obtainable in Nigeria among health professionals and students is also lacking. There is a need to educate our religious leaders, government and non-government organizations and the populace. PMID- 15296109 TI - Factors militating against delivery among patients booked in Olabisi Onabanjo University Teaching Hospital, Sagamu. AB - BACKGROUND: A preliminary review of antenatal and delivery records of Olabisi Onabanjo University Teaching Hospital (OOUTH), Sagamu revealed that only 47.6% of booked pregnant women subsequently delivered there. This observation stimulated a thought to carry out a study to find out the proportion of pregnant patients who booked in OOUTH and subsequently delivered in OOUTH, patients' preferred place of delivery and factors militating against OOUTH delivery. METHOD: A cross-sectional study was carried out on pregnant patients booked at OOUTH, Sagamu between December, 2001 and May, 2002 to find out factors militating against delivery at OOUTH. Two hundred and sixty six questionnaires were returned properly filled. RESULTS: The age range of the pregnant women was 16 to 42 years with a mean age of 31.2 +/- 3.4 years. The parity ranged from 1-5 with a mean of 2.1 +/- 1.1. The preferred place of delivery was private hospital (58.3%), followed by OOUTH (28%). The higher the educational level, the more they tended to hospital delivery; 57% of the pregnant women who had hospital deliveries had tertiary education. While 52.7% of the pregnant women gave miscellaneous reasons such as relocation, being used for experiment, too many students being present, and frequent strikes by hospital workers for not delivering in OOUTH, as much as 16.5% said the time wasting and bad attitude of the hospital staff were responsible for preferring centres other than OOUTH for delivery. CONCLUSION: OOUTH like most other teaching/government hospitals need to be more user-friendly to encourage better patronage. There is a need to educate the patients that attend teaching hospital on the essence of establishing teaching hospitals and the importance of training medical students and retraining of doctors in order to maintain a continuity and high standard of medical practice. PMID- 15296110 TI - Accidental incision of the fetus at caesarian section. AB - BACKGROUND: Caesarian section is the most common major surgical procedure among women of reproductive age. It is thought to be safer for the baby than the mother. However, the baby is at risk of laceration injury. METHODS: Retrospective review of clinical records of women who were delivered by caesarian section during a seven-year period from January 1, 1991 to December 31, 1997. RESULTS: Fetal laceration injury was documented in 0.55% of live caesarian section births. Accidental incision of the fetus was associated with emergency caesarian section, ruptured membranes and relative inexperience of the operating surgeon. Documentation of injury was poor and there was no evidence that parents were counselled. CONCLUSION: Laceration of the fetus is an occasional complication of caesarian section. Proper documentation and counselling of parents are required especially as there are potential medicolegal implications. Careful attention to the technique of uterine entry at caesarian section should reduce the risk of injury to the fetus. PMID- 15296111 TI - Epidural blood patch for post dural puncture headache: a handy ally [corrected]. AB - BACKGROUND: Epidural blood patch is a well-known item in the management protocol of post dural puncture headache though never been practised or reported in our environment. The aim of this report is to document its efficacy and establish a rationale for its use. METHOD: A review of the case record of patients with post dural puncture headache and the relevant literature. RESULT: A case of post dural puncture headache was encountered at a free medical missionary outreach program following spinal anaesthesia for myomectomy. Having exhausted all the available conservative management options, she was offered epidural blood patch. The recovery profile was collected by assessment of her abilities and disabilities before and after the epidural blood patch, with visual analog scale for pain. The clinical condition improved a lot, with remarkable improvement in neck movement and the visual analog scale. CONCLUSION: We conclude that epidural blood patch is a superior therapy for post dural puncture headache and recommend that post anaesthetic rounds be conducted routinely to identify complications and enable early institution of therapy. PMID- 15296112 TI - Pre-eclampsia presenting as nephrotic syndrome: a case report. AB - BACKGROUND: Nephrotic syndrome occurring for the first time in pregnancy is rare. We report pre-eclampsia presenting with nephrotic syndrome. METHOD: A case report utilizing the case record and review of relevant literature. RESULTS: A 26-year old multi-gravida who presented with nephrotic syndrome for the first time in pregnancy. Proteinuria resolved spontaneously after delivery. CONCLUSION: This report highlights the need for increased awareness of this rare presentation of pre-eclampsia, especially in the tropics where a large majority of women presenting for delivery are unbooked. PMID- 15296113 TI - An uncommon cause of massive penoscrotal haematoma. AB - BACKGROUND: Haematoma is a common complication of surgical operations. Haematoma of traumatic origin is often associated with severe and extensive injuries. In patients with disorders of blood coagulation, haematoma may occur even after trivial injuries. This paper aims to report a case of massive penoscrotal haematoma. METHOD: The case record of a haemophiliac with massive penoscrotal haematoma and relevant literature on the disease were reviewed. RESULT: A 43 year old haemophiliac who developed extensive penoscrotal haematoma following trivial injury to his left groin which had to be evacuated operatively and his haemostatic function stabilised with factor VIII concentrate. He developed secondary haemorrhage post operatively and had antibiotics and further transfusion of whole blood and factor VIII concentrate. His wounds healed and he was discharged after two months of hospitalisation. He was followed up in the out patient clinic for six months without any further bleeding. CONCLUSION: Operative evacuation of the haematoma was necessary to avoid the development of necrotising fascitis of the scrotum (Fournier's gangrene). PMID- 15296114 TI - Clinical photographic quiz. PMID- 15296115 TI - [The decrease in autopsy rates and unconfirmed clinical diagnoses--yes or no?]. PMID- 15296116 TI - N-methyl-3,4-methylenedioxyamphetamine-induced hepatotoxicity in rats: oxidative stress after acute and chronic administration. AB - BACKGROUND: The underlying mechanisms of N-methyl-3,4-methylenedioxyamphetamine- MDMA--induced hepatotoxicity are still unknown. The aim of this study was to evaluate hepatic oxido-reductive status in the rats liver after the single and repeated administration of MDMA. METHODS: MDMA was dissolved in distilled water and administered in the doses of 5 mg, 10 mg, 20 mg, and 40 mg/kg. The animals from the acute experiment were treated per os with the single dose of the appropriate solution, through the orogastric tube. The animals from the chronic experiment were treated per os, with the doses of 5, 10, or 20 mg/kg of MDMA every day during 14 days. The control groups were treated with water only. Eight hours after the last dose, the animals were sacrificed, dissected, their livers were rapidly removed, frozen and stored at -70 degrees C until the moment of analysis. The parameters of oxidative stress in the crude mitochondrial fractions of the livers were analyzed. RESULTS: Superoxide dismutase (SOD) activity increased in the livers of the animals that were treated with single doses of MDMA. Chronically treated animals showed the increased SOD activity only after the highest dose (20 mg/kg). The content of reduced glutathione decreased in both groups, but the depletion was much more expressed after the single administration. Lipid peroxidation index increased in dose-dependent manner in both groups, being much higher after the single administration. CONCLUSION: The increased index of lipid peroxidation and the decreased reduced glutathione levels suggested that MDMA application induced the state of oxidative stress in the liver. These changes were much more expressed after the single administration of MDMA. PMID- 15296117 TI - [Epidural hematomas of the posterior fossa]. AB - BACKGROUND: Posterior fossa epidural hematomas represent 7-14% of all traumatic intracranial epidural hematomas. They are most frequently encountered posttraumatic mass lesions in the posterior fossa. The aim of this study was to identify clinical features that could lead to the early diagnosis of posterior fossa epidural hematoma. METHODS: Between 1980 and 2002, 28 patients with epidural hematoma of the posterior fossa were operated on at the Institute for Neurosurgery, Belgrade. Clinical course, neuroradiological investigations, and the results of surgical treatment of the patients with posterior fossa epidural hematomas were analyzed retrospectively. RESULTS: Almost two thirds of patients were younger than 16 years of age. In 20 cases injury was caused by a fall, in 6 cases by a traffic accident, and in 2 by the assault. Clinical course was subacute or chronic in two thirds of the patients. On the admission Glasgow Coma Scale was 7 or less in 9 injured, 8-14 in 14 injured, and 15 in 5 injured patients. Linear fracture of the occipital bone was radiographically evident in 19 patients, but was intraoperatively encountered in all the patients except for a 4-year old child. In 25 patients the diagnosis was established by computer assisted tomography (CAT) and in 3 by vertebral angiography. All the patients were operated on via suboccipital craniotomy. Four injured patients who were preoperatively comatose were with lethal outcome. Postoperatively, 24 patients were with sufficient neurologic recovery. CONCLUSION: Posterior fossa epidural hematoma should be suspected in cases of occipital injury, consciousness disturbances, and occipital bone fracture. In such cases urgent CAT-scan is recommended. Early recognition, early diagnosis, and prompt treatment are crucial for good neurological recovery after surgery. PMID- 15296118 TI - [Importance of determination of proinflammatory cytokines in the blood of polytraumatized patients with sepsis]. AB - Severe sepsis and trauma complicated with multiple organ dysfunction syndrome (MODS) are among the leading causes of death in intensive therapy units, with mortality rate exceeding 50%. The outcome is not determined only by infection or trauma, but also by the intensity of immuno-inflammatory response, which is essential for host defence, but if uncontrolled leads to MODS. Pro-inflammatory cytokines (tumor necrosis factor-alpha--TNF-alpha, IL-1, IL-8, IL-12, IFN-gamma, etc.) represent a part of this immuno-inflammatory response to an insult. The results of the clinical investigation of correlation between pro-inflammatory cytokines (IL-8, IL-12, TNF-alpha, IFN-gamma), the outcome (survivors, non survivors), and the severity (systemic inflammatory response syndrome--SIRS--less severe, and MODS--more severe) in polytraumatised patients with sepsis are presented in this paper. Mean values of IL-8 were 1.3-fold higher in non survivors (p<0.05), and 60-fold higher in MODS group (p<0.01). Mean values of IL 12 were 1.6-fold higher in survivors (p<0.01), while the values between SIRS and MODS group did not differ significantly; mean values of TNF-alpha were 3-fold higher in survivors (p<0.05), and 46-fold higher in MODS group (p<0.01). Mean values of IFN-gamma did not differ significantly between the two groups regarding the outcome and severity. The obtained results indicated that IL-8 was a reliable predictor of lethal outcome and MODS (p<0.01), IL-12 a reliable predictor of survival (p<0.05), and TNF-alpha a reliable predictor of survival (p<0.05) and MODS (p<0.01). PMID- 15296119 TI - [Selection of the optimal radiotherapy technique for locally advanced maxillary carcinoma using a three-dimensional treatment planning system]. AB - AIM: To compare the isodose distribution of three radiotherapy techniques for locally advanced maxillary sinus carcinoma and analyze the potential of three dimensional (3D) conformal radiotherapy planning in order to determine the optimal technique for target dose delivery, and spare uninvolved healthy tissue structures. METHODS: Computed tomography (CT) scans of fourteen patients with T3 T4, N0, M0 maxillary sinus carcinoma were acquired and transferred to 3D treatment planning system (3D-TPS). The target volume and uninvolved dose limiting structures were contoured on axial CT slices throughout the volume of interest combining three variants of treatment plans (techniques) for each patient: 1. A conventional two-dimensional (2D) treatment plan with classically shaped one anterior + two lateral opposite fields and two types of 3D conformal radiotherapy plans were compared for each patient. 2. Three-dimensional standard (3D-S) plan: one anterior + two lateral opposite coplanar fields, which outlines were shaped with multileaf collimator (MLC) according to geometric information based on 3D reconstruction of target volume and organs at risk as seen in the beam eye's view (BEV) projection. 3. Three-dimensional non-standard (3D-NS) plan: one anterior + two lateral noncoplanar fields, which outlines were shaped in the same manner as in 3D-S plans. The planning parameters for target volumes and the degree of neurooptic structures and parotid glands protection were evaluated for all three techniques. Comparison of plans and treatment techniques was assessed by isodose distribution, dose statistics, and dose-volume histograms. RESULTS: The most enhanced conformity of the dose delivered to the target volume was achieved with 3D-NS technique, and significant differences were found comparing 3D-NS vs. 2D (Dmax: p<0.05; Daver: p<0.01; Dmin: p<0.05; V90: p<0.05, and V95: p<0.01), as well as 3D-NS vs. 3D-S technique (Dmin: p<0.05; V90: p<0.05, and V95: p<0.01), while there were no differences between 2D vs. 3D-S technique. 3D-S conformal plans were significantly superior to 2D plans regarding the protection of parotid glands, and the additional improvement of dose conformity was achieved with 3D-NS technique. 3D-NS technique resulted in the decrease of Dmax for ipsilateral retina compared with 3D-S technique, while the level of Dmax for optic nerve was increased (within an acceptable range) with 3D-NS technique. CONCLUSION: In this study, 3D planning of radiotherapy for locally advanced maxillary sinus carcinoma with noncoplanar fields, which number did not exceed the number of fields for conventional arrangement enabled conformal delivering of the adequate dose to the target volume with the improved sparing of adjacent uninvolved healthy tissue structures. PMID- 15296120 TI - [Predictive significance of residual ischemia detected by the dobutamine stress echocardiography test soon after the first uncomplicated myocardial infarction]. AB - BACKGROUND: To evaluate the long-term prognostic value of dobutamine stress echocardiography (ECG) test for new coronary events (new episodes of angina pectoris, cardiac-related deaths, and reinfarctions) early after the first uncomplicated myocardial infarction. METHODS: Dobutamine stress-echocardiography tests were performed in all of 104 patients 10-20 days after the first myocardial infarction. Patients were followed-up for 36 (29 +/- 7) months. Kaplan-Meier cumulative survival curves were tested by Breslow test (Log Rank). RESULTS: Two cardiac deaths (1.92%), nine nonfatal myocardial infarctions (8.65%), and three cases of recurrent angina pectoris (2.88%) occurred during the prospective follow up. Cumulative survival curves showed that in patients with negative findings of dobutamine stress-echocardiography test, survival time without significant events was 35.31 months, while in the group with positive findings of dobutamine stress echocardiography test it was 30.91 months (log Rank 7.22; p<0.01). Prognostic value of dobutamine stress-echocardiography test was analyzed by Cox regression model and was 2.92, meaning that the risk of significant events was 2.92 times higher in the group of patients with positive findings of dobutamine stress echocardiography test. CONCLUSION: Patients with negative findings of dobutamine stress-echocardiography test were with significantly higher possibility of surviving without significant events in comparison with the patients in whom the findings of dobutamine stress-echocardiography test were positive. In combination with clinical signs and ECG results, the results of dobutamine stress echocardiography test improved prognostic value in the patients with the first uncomplicated myocardial infarction, and in that way influenced the strategy of their further treatment. PMID- 15296121 TI - [Clinical significance of intramammary arterial calcifications in women with diabetes mellitus]. AB - BACKGROUND: It is well known that intramammary arterial calcifications, diagnosed by mammography as a part of generalized diabetic macroangiopathy, may be an indirect sign of diabetes mellitus. Hence, the aim of this study was to determine the incidence of intramammary arterial calcifications, the patient's age when the calcifications occur, as well as to observe the influence of diabetic polyneuropathy, type, and the duration of diabetes on the onset of calcifications, in comparison with nondiabetic women. METHODS: Mammographic findings of 113 diabetic female patients (21 with type 1 diabetes and 92 with type 2), as well as of 208 nondiabetic women (the control group) were analyzed in the prospective study. The data about the type of diabetes, its duration, and polyneuropathy were obtained using the questionnaire. Statistical differences were determined by Mann-Whitney test. RESULTS: Intramammary arterial calcifications were identified in 33.3% of the women with type 1 diabetes, in 40.2% with type 2, and in 8.2% of the women from the control group, respectively. The differences comparing the women with type 1, as well as type 2 diabetes and the controls were statistically significant (p=0.0001). Women with intramammary arterial calcifications and type 1 diabetes were younger comparing to the control group (median age 52 years, comparing to 67 years of age, p=0.001), while there was no statistically significant difference in age between the women with calcifications and type 2 diabetes (61 years of age) in relation to the control group (p=0.176). The incidence of polineuropathy in diabetic women was higher in the group with intramammary arterial calcifications (52.3%), in comparison to the group without calcifications (26.1%), (p=0.005). The association between intramammary arterial calcifications and the duration of diabetes was not found. CONCLUSION: The obtained results supported the theory that intramammary arterial calcifications, detected by mammography, could serve as markers of co-existing diabetes mellitus and therefore should be specified in radiologic report in case of their early development. PMID- 15296122 TI - Evaluation of usefulness of single-strand conformation polymorphism method for rapid detection of rifampicin-resistant Mycobacterium tuberculosis. AB - The aim was to evaluate the Single-Strand Conformation Polymorphism method as a potential tool for rapid detection of rifampicin-resistant strains by the use of 39 rifampicin-resistant Mycobacterium tuberculosis strains isolated in Serbia. SSCP analysis on acrylamide gel detected 56.4% of the rifampicin-resistant M. tuberculosis strains and showed the inability to detect one of the most frequent mutations, TCG-->TTG mutation in codon 531 of the rpoB gene, which was shown by automated sequencing. PMID- 15296123 TI - [Neurological disorders in patients with Kallmann's syndrome]. PMID- 15296124 TI - [Pharmacology of receptors and cellular glutamate transporters]. PMID- 15296125 TI - [Current views on the diagnosis of pulmonary thromboembolism]. PMID- 15296126 TI - [Breast reconstruction by pedicled transverse rectus abdominis myocutaneous flap]. AB - Reconstruction of the amputated breast in female patients after surgical management of breast carcinoma is possible with the use of autologous tissue, synthetic implants, or by combining autologous tissue and synthetic materials. Autologous tissue provides soft and sufficiently elastic tissue, which is usable for breast reconstruction and eventually obtains original characteristics of the surrounding tissue on the chest wall. The use of the TRAM flap for breast reconstruction was introduced in 1982 by Hartrampf, Scheflan, and Black. The amount of the TRAM flap tissue allows breast reconstruction in the shape most adequate to the remaining breast. The possibilities of using the TRAM flap as pedicled myocutaneous flap or as free TRAM flap make this flap a superior choice for breast reconstruction in comparison with other flaps. PMID- 15296127 TI - [Suicide in the military environment]. AB - Suicide of soldiers has its own specifics, because not only it represents the tragedy for the individuals and their family, but also has great psychological effect on social environment and military unit in which it occurs. Suicide can be caused by variety of factors, as reported in the literature. The case reviewed in this article presents multilateral determination of suicide, with particular stress on the character of each individual and social interaction of soldiers. Psychological complex of basic inferiority, low educational level, family problems, and poor integration into military unit could be considered the leading determinants of this suicide. This emphasizes the importance of certain preventive measures such as more rigorous psychological selection for specific military duty, and the education of non-commissioned officers for better recognition and understanding of pre-suicidal syndrome. PMID- 15296128 TI - [Ocular rosacea]. AB - Five cases of ocular rosacea (one male and four females) are reported in this paper. Two of the patients were with keratoconjunctivitis sicca, one with conjunctivitis chronica and blepharitis, one with conjunctivitis chronica and meibomitis, and one with reccurent corneal erosions with meibomitis and chordeloum. In four patients ocular symptoms preceded the occurence of skin lesions. The treatment with oral tetracyclines significantly improved the state of ocular rosacea in four patients, while in one case the changes of the anterior eye segment progressed in more severe state of ulcerative keratitis. It is considered that in almost 20% of the patients with rosacea ocular lesions may precede the skin changes, representing a diagnostic problem. Thus, in those cases multidisciplinary approach is suggested. Such approach is particularly important because of the decrease of morbidity and the prevention of the onset of the eye complications such as drastic worsening of visual acuity, i.e., the blindness. PMID- 15296129 TI - [Rare forms of panniculitis: subcutaneous sarcoidosis and panniculitis associated with dermatomyositis]. AB - Panniculitides represent heterogenous group of disorders involving subcutaneous fat tissue, and are etiologically related to different causes, including systemic diseases. Two female patients having subcutaneous sarcoidosis and panniculitis associated with dermatomyositis are presented in this paper. The first patient, (38 years of age), was with the nodes on forearms and lower legs, which occurred one month after hypophysectomy for adrenocorticoscopic (ACTH)-secreting pituitary adenoma, and showed histopathologically confirmed epithelioid, noncaseating granulomas in the subcutaneous fat tissue. Laboratory and radiographic findings were normal, with the exception of positive rheumatoid factor and incompletely developed cysts in proximal phalanges of the hand. The second patient (56 years of age), was with subcutaneous nodes on the upper arm and the lateral chest wall, which were noticed several months after the diagnosis of dermatomyositis. Histopathological examination showed lobular pannicilitis with hyaline fibrosis and plasmocytic-lymphocytic infiltrate. PMID- 15296130 TI - [Serbian military surgery experience (1876-1918): part 4. Military surgery in Serbia during the Balkan Wars (1912-1913)]. PMID- 15296131 TI - Possibilities for funding of applied biotechnology research in the sixth Framework Programme. PMID- 15296132 TI - Quality without tears--practical applications of quality assurance in biotechnology. AB - Expectations of society and funding bodies are increasing the application of quality management systems to research and development. Implementation of such systems needs to be performed sensibly to ensure that the creativity, which is at the core of the research process, can still thrive whilst increasing the visible reliability of outputs. This paper explores the application of quality principles to research and development and provide practical advice on their application. PMID- 15296133 TI - Xenotransplantation. PMID- 15296134 TI - Fate of fuel oxygenates in the environment. PMID- 15296135 TI - Organomercurial removal from vaccine production wastewaters in a supported liquid membrane bioreactor. AB - Vaccine production effluents are strongly polluted with thiomersal, a highly toxic organomercurial compound, for which there is presently no remediation technology available. This work describes a new remediation process based on the extraction of thiomersal from the wastewater to a biological compartment, where it is degraded by a microbial strain. The selective extraction of thiomersal is achieved by using an ionic liquid immobilized in a porous membrane. In the biological compartment, thiomersal is degraded to metallic mercury, under aerobic conditions, by a Pseudomonas putida strain. The utilization of ionic liquids in supported liquid membranes for thiomersal transport, and the kinetics of thiomersal biodegradation by a Pseudomonas putida strain are presented and discussed. PMID- 15296136 TI - Technology for biodestruction of heptil. PMID- 15296137 TI - Bioluminescence-based assay used for toxicity monitoring. AB - A new ATP bioluminescence-based method is proposed in order to examine the effect of toxic shock on microbial communities in urban wastewater treatment plant (WWTP). As the efficiency of WWTP must be improved, the occurrence and toxic effect of hazardous substances have to be evaluated (Directive 2455/2001/EC). Among bioluminescence assay with firefly luciferase is commonly used for intracellular ATP monitoring. A multienzyme systems catalysed by three enzymes (adenylate kinase, pyruvate kinase and firefly luciferase) for adenine nucleotides (AN = ATP+ ADP+ AMP) estimation is used for monitoring metabolic changes in activated sludge. The AN method provides better sensitivity than ATP quantification alone. The pool of intracellular and extracellular AN is constant during growth and the rate of intracellular AN is correlated with the microbial growth. In the present investigation, the extraction of total adenylate assay is optimized (chemical and mechanical extraction) and applied for monitoring the effect of the toxic shock (eg. Naphthalene) on indigenous biomass activity (intracellular and extracellular AN rates). In the same time, the removal of naphthalene is addressed Laboratory-scale reactors (4 L) are inoculated with urban activated sludge is under continuous-flow aeration. The reactor, containing non acclimated sludge enriched with naphthalene. The other reactor containing sterilised activated sludge is used as abiotic and volatilisation control system. In all this batch cultures, ATP and AN (intracellular and extracellular) rates and naphthalene concentrations are controlled. One of the objectives of this research is to determine a relation between the ATP/AN ratio and the biodegradation of naphthalene. Evidence is given which demonstrates that the ATP/AN parameter is a possible alternative for monitoring very rapid metabolic changes in complex microbial community such as activated sludge. PMID- 15296138 TI - Use of biolog methodology for optimizing the degradation of hydrocarbons by bacterial consortia. AB - Biolog methodology was used for the preliminary screening of different cultural conditions in order to detect the best combination/s of factors influencing the metabolic performance of bacterial consortia active in the degradation of hydrocarbons. Two microbial consortia were tested for their activity on 2 hydrocarbons (nonadecane and eicosane) in presence of the following cultural coadjuvants: vegetal oil, beta-cyclodextrine, sodium acetate, mineral solution. Tests were conducted in Biolog MT plates, where only the redox indicator of microbial growth (tetrazolium violet) and no carbon sources are provided. The microwells were filled with various combinations of hydrocarbons, microbial inoculum and coadjuvants. Blanks were prepared with the same combinations but without hydrocarbons. The results obtained show the suitability of the methodology developed to identify the most active consortium and the conditions for its best degradation performance. The efficacy of Biolog methodology (Biolog Inc., USA) for the characterization of microbial communities on the basis of the metabolic profiles obtained on specific carbon sources in the microwells of Elisa type plates, is widely acknowledged (Garland, 1997; Pietikainen et al., 2000; Dauber and Wolters, 2000). Due to its aptitude to simultaneously evaluate multiple microbial responses and directly organize the results, it can be adapted to meet specific study purposes (Gamo and Shji, 1999). In the present research Biolog methodology was fitted for the preliminary screening of different cultural conditions, in order to detect the best combination/s of factors influencing the metabolic performance of bacterial consortia active in the degradation of aliphatic hydrocarbons, in view of their utilization for the bioremediation of polluted sites. PMID- 15296139 TI - Performance of a submerged aerated filter and a rotating biological contactor under dynamic loading conditions. AB - Two types of small wastewater treatment systems were studied for their performance under normal conditions, including the hydraulic peak flows associated with small systems connected to just one house. Furthermore, the systems were subjected to a 7-day starvation period to simulate the effect of a holiday from home. The systems studied are (1) a combined submerged aerated filter-activated sludge system and (2) a rotating biological contactor system. Both the organic removal and the nitrification process were closely monitored. During normal operation, very good treatment results were achieved. The combined SAF-AS system realized 95% BOD removal, 88% COD removal and 94% NH4-N removal. The RBC system removed 92% of the BOD, 89% of the COD and 99% of the ammonium nitrogen. Both systems do not experience severe problems dealing with the lack of influent for a duration of seven days. The effluent concentrations did not change much, except for a small peak of nitrite which was present in all tests. However, both the ammonium oxidizing and the nitrite oxidizing bacterial populations were still active, as evidenced by the continued removal of ammonium and formation of nitrate. PMID- 15296140 TI - Practical experiences with start-up and operation of a continuously aerated lab scale SHARON reactor. AB - Partial nitrification techniques, such as the continuously aerated SHARON process, have been denoted for quite a while as very promising for improved sustainability of wastewater treatment. Combination of such a SHARON process with the Anammox process, where ammonium is oxidised with nitrite to nitrogen gas under anoxic conditions, leads to cost-efficient and sustainable nitrogen removal from concentrated streams. In this study practical experiences during start-up and operation of a lab-scale SHARON reactor are discussed. Special attention is given to the start-up in view of possible toxic effects of high ammonium and nitrite concentrations (up to 4000 mgN/l) on the nitrifier population and because the reactor was inoculated with sludge from a SBR reactor operated under completely different conditions. Because of these considerations, the reactor was first operated as a SBR to prevent biomass wash out and to allow the selection of a strong nitrifying population. A month after the inoculation the reactor was switched to normal chemostat operation. As a result the nitrite oxidisers were washed out and only the ammonium oxidisers persisted in the reactor. In this contribution also some practical considerations, such as mixing, evaporation and wall growth, concerning the operation of a continuously aerated SHARON reactor are discussed. These considerations are not trivial, since the reactor will be used for kinetic characterisation and modelling studies. Finally the performance of the SHARON reactor under different conditions is discussed in view of its coupling with an Anammox unit. Full nitrification was proven to be feasible for nitrogen loads up to 1.5 g/l d, indicating the possibility of the SHARON process to treat highly loaded nitrogen streams. PMID- 15296141 TI - Nutrient removal in a sequencing batch reactor operated with short anaerobic/aerobic cycles. AB - A single sequencing batch reactor operated with short intermittent aeration cycles was used to simultaneously remove carbon, nitrogen and phosphorus. The complete cycle, comprising feeding, anaerobiosis, aerobiosis, settling and decanting, was only 36 minutes long. The system has shown high and stable nutrient removal at 30 degrees C with acetate as carbon source and it has proved to be rather robust and dynamic, efficiently adapting to most of the changes in operating parameters tested: presence of nitrate in the feeding medium, different substrates (propionate and butyrate), temperature and nutrient shock loads. For the optimum conditions used, a removal efficiency of over 90% was obtained for each nutrient. Description of the population kinetics was obtained for each operating condition, by performing batch tests. Kinetic and stoichiometric parameters were used to infer the relative contribution of each group of microorganisms on SBR performance. Compared to the traditional SBR operated with cycles of 6 hours, the use of short intermittent aeration cycles of 36 minutes corresponds to a 40% reduction on aeration time. PMID- 15296142 TI - Nitrification of ammonia nitrogen high concentration in membrane assisted bioreactor. AB - Use of the membrane-assisted bioreactors (MBR) in wastewater treatment can bring a lot of advantages. Usually COD removal achieves level 90% and nitrification of ammonia nitrogen concentrations typical for municipal wastewater performs without any disturbances, what was confirmed by large number of experiments. However, inhibition of nitrification of the high ammonia nitrogen concentration was noticed. In this study, the performance of ammonia-rich wastewater nitrification in membrane-assisted bioreactor (MBR) was examined. With SRT of 40, 32, 24 and 16 days any nitrification inhibition effect wasn't noticed. Ammonia nitrogen removal was around 98% and full nitrification was observed. At the sludge age equal to 12 d the first stage of nitrification was full, but the second stage of nitrification was incomplete. The variation in number and presence of the higher organisms were noticed at the all examined sludge ages. With SRT of 40, 32, 24 and 16 days the free ciliates were prominent organisms. At the sludge age of 12 days the domination of flagellates was observed The kinetic constants of the high ammonia wastewater nitrification were calculated The average value of Vmax at the sludge age 12, 16 and 24 days was constant (4.7 mg NH4(+)-N/g MLSS h, 4.8 mg NH4(+)-N/g MLSS h and 4.8 mg NH4(+)-N/g MLSS, respectively), but the value of Km rose, when the sludge age increased (6.8 mg NH4(+)-N/L for 12 days, 11.3 mg NH4(+)-N/L for 16 days, 21.6 mg NH4(+)-N/L for 24 days). At the sludge age 32 and 40 days the increase of Vmax was observed (7.2 mg NH4(+)-N/g MLSS h and 12.5 mg NH4(+)-N/g MLSS h, respectively) and, also, changes of Km (6.8 mg NH4(+)-N/L and 44 mg NH4(+)-N/L, respectively. Concerning the second stage of nitrification value of Vmax decreased with the increase of the sludge age and the average Km varied from 20.3 to 31.3 mg NO2(-)-N/L. PMID- 15296143 TI - On-line control of biological nutrient removal processes in practice: a cost benefit analysis. AB - On-line control is able to boost process efficiency and optimise effective capacity of nutrient removal plants. In this paper, costs and benefits of on-line control are presented and economical feasibility is analysed using a simple economical cost-benefit analysis. These concepts are illustrated by a number of full-scale case studies. The on-line control of ferric dosage is illustrated by the case of the Lommel WWTP, and the Harelbeke WWTP and Houthalen WWTP illustrate advanced on-line control of aeration for nitrogen removal. PMID- 15296144 TI - Production of polyhydroxyalkanoates by a mixed culture in a sequencing batch reactor: the use of propionate as carbon source. AB - In this work, sludge adapted to anaerobic/aerobic conditions, AN/AE, showing a high capacity of P accumulation, was submitted to aerobic dynamic substrate feeding (ADF). The fermenter was operated as a Sequencing Batch Reactor, with propionate as carbon substrate. Propionate is an important waste product from several industrial processes that can be valued, using it as a precursor for hydroxyvalerate in PHA production. Under the operational conditions used, apart from 3-hydroxyvalerate as its major component, 3-hydroxy-2-methylvalerate, 2 hydroxyisovalerate and 4-oxovalerate were also produced. A second reactor operated under the same conditions was adapted for the use of acetate as carbon substrate. The global metabolism of the organisms involved on PHA production, utilizing acetate or propionate, was studied using in vivo 13C Nuclear Magnetic Resonance (NMR). PMID- 15296145 TI - Novel trends in anaerobic digestion of municipal solid waste. AB - Anaerobic digestion capacity has been installed on a large scale for the treatment of biowaste coming from municipal solid waste in the 90's. However, in recent years, a new trend has developed in which anaerobic digestion is applied more and more for the treatment of mixed or grey waste. It is expected that the installed capacity for grey/mixed waste will surpass the capacity installed for biowaste digestion. Five years ago, more than 85% of the treatment capacity was for biowaste digestion and only 15% for grey/mired waste digestion, derived from two old plants that were constructed prior to 1990. By the end of 2004, a digestion capacity of 1,285,000 ton per year will be available in Europe for the treatment of grey or mixed MSW, while digestion capacity for biowaste will only amount to 1,270,000 ton per year. Especially dry digestion offers new perspectives. Dry digestion is particularly suited for the treatment of grey/mixed waste due to its insensitivity to the presence of heavy inerts and light materials. Heavy inerts such as sand, glass and stones cause sedimentation and the light materials cause floatation and scum formation in the more conventional wet and semi-dry systems. Due to the high initial dry solids content, the digestate coming from dry digestion can be treated in a variety of ways. Besides the conventional mechanical dewatering, drying with waste heat or aerobic drying by addition of a fresh waste can be utilized for the production of a high-quality compost in case of the treatment of biowaste. Digestate from mixed or grey waste is not immediately suitable for the production of a high-quality compost. However, integration with incineration plants and landfills can be optimized easily with such a dry digestate and offers various interesting alternatives. In case a maximum of recyclables is pursued, the dry digestion can be followed by a wet separation in order to produce marketable endproducts such as sand and fibers. PMID- 15296146 TI - Manure treatment according to the Trevi-concept. AB - In Flanders great amounts of livestock, especially pigs, lead to a surplus of manure that can not be spread on farmland because of the prevailing fertilizing restrictions. Aside that, Flemish government obliges farmers with a surplus of 7500 kg total P2O5 production to treat this excess of manure partially or totally. Trevi N.V. developed a manure treating system that starts with the separation of the raw manure in a solid fraction and a liquid fraction. The solid fraction can be exported after drying and hygienisation in the condensation drier, which is characterised by the absence of off-gases. The liquid fraction is treated in a biological plant and contains only 10 to 15% of the original amount of nutrients. As a result of this, the biological effluent can be spread on farmland at a bigger quantity (100-150 ton/ha) in comparison with raw manure. This biological effluent needs further polishing to obtain an effluent that can be discharged into surface waters. This was realised in coordination with Danis N. V. at the Discover plant in Izegem. In this plant, an evaporator treats the biological effluent and produces a dischargable condensate. PMID- 15296147 TI - Development of a sustainable treatment technology for domestic sewage. AB - We propose an integrated chemical-physical-biological treatment concept for the low-cost treatment of domestic wastewater. Domestic wastewater was subjected to a chemically enhanced primary sedimentation (CEPS), followed by treatment in an upflow anaerobic sludge blanket (UASB) reactor. In addition, a regenerable zeolite was used to remove NH4+, either after CEPS pretreatment or after biological treatment in the UASB reactor. The CEPS pretreatment consisted of the addition of a coagulant (FeCl3) and an anionic organic flocculant and removed on average 73% of the total chemical oxygen demand (CODt), 85% of the total suspended solids, and 80% of PO4(3-) present in the wastewater. The UASB system, which consequently received a low CODt input of approximately 140 mg/L, was operated using a volumetric loading rate of 0.4 g CODt/L.d (hydraulic retention time [HRT] = 10 h) and 0.7 g CODt/L.d (HRT = 5 h). For these conditions, the system removed about 55% of the CODt in its influent, thus producing an effluent with a low CODt of approximately 50 mg/L. The zeolite, when applied in batch mode before the UASB reactor, removed approximately 45% of the NH4+, whereas its application as a post-treatment cartridge resulted in almost 100% NH4+ removal. The simple design and relatively low operating costs, due to low costs of added chemicals and low energy input (estimated at Euro 0.07-0.1 per m3 wastewater treated), combined with excellent treatment performance, suggest that this system can be used as a novel domestic wastewater treatment system for developing countries. Therefore, the system is called a Low-cost, Integrated Sewage Treatment (LIST) system. PMID- 15296148 TI - Growth enhancement of ETBE-degrading bacterial consortium with various carbon sources. AB - In this study, we evaluated Ethyl tert-Butyl Ether (ETBE)-degrading consortia growths in the presence of diverse carbon sources (alcohols, alkanes, ether compounds and carbohydrates). In a second step, we studied the consortium ability to maintain its ETBE degradation activity after growing on these carbon sources in presence or in absence of ETBE. The results indicate that the bacterial growth of ETBE-degrading consortia is enhanced three times more with addition of ethanol than with ETBE alone, while maintaining its ability to degrade ETBE. The bacterial yield growth rate was 0.504 d(-1) when growing on ETBE alone, 1.728 d( 1) on both ETBE and ethanol and 2.856 d(-1) on ethanol alone. Both ETBE and ethanol are completely degraded at 8.33 mg L(-1) h(-1) and 18.55 mg L(-1) h(-1) respectively for an initial OD of 0.4. The frequency of ethanol addition, as growth co-substrate, was studied to preserve the ETBE-degrading capacity of the consortium, and to observe the stability of the genetic character of the ether degradation. PMID- 15296149 TI - Engineering the interaction between micro-organisms and construction materials. AB - The influence of micro-organisms on degradation of mineral materials, cement bound systems, wood and steel is a rather new subject of research slowly becoming recognised by the 'classical' technical disciplines. An increasing amount of literature appears on biodeterioration of construction materials and microbial activity can not be neglected as a determining factor in the deterioration process. Microbial communities interact in many different ways with mineral materials and their external environment. They can be present on the surface or in crevices and fissures within the material and often their actions become organized in a biofilm. The interaction with the material and its environment can give rise to biodeterioration. Yet recent findings show that in some cases the microbial interaction can lead to protection of materials. It is our mission for the future to engineer the microbiological processes with positive impact on construction materials with a view to practical applications. PMID- 15296150 TI - Simple operational tools for secondary settler operation--conceptual validation on an extended full-scale data set. PMID- 15296151 TI - Comparison of yeast (Candida maltosa) and bacterial (Rhodococcus erythropolis) phenol hydroxylase activity and its properties in the phenolic compounds biodegradation. AB - Aromatic contaminants of the environment, to which belongs phenol and its derivatives, are toxic and in most of the cases hard to degrade. Removal of these pollutants by biological, gentle and effective way, depends on specific environmental conditions in the locality and on the biodegradation potential of the used microbial population. Closer characterization of the biodegradation and enzyme mechanisms is therefore an essential assumption of the successful implementation of microbes. This paper is focused on comparison of the biodegradation activity between the soil yeast Candida maltosa and bacteria Rhodococcus erythropolis towards various aromatics connected with determination of the first enzyme of the phenol biodegradation pathway: phenol hydroxylase (PH). The effect of substrate type, substrate concentration, growth phase of the microorganisms and presence of humic acids in the cultivation medium, on phenol biodegradation and PH activity are discussed. PMID- 15296152 TI - Nitrate removal in aquaria systems: use of electrochemically generated hydrogen gas as electron donor for denitrification. AB - Nitrate, the end product of the nitrification process, tends to accumulate in aquaria systems. Removal of nitrate in these systems by means of biological denitrification requires the addition of an external electron donor. In this study, the possibilities for using hydrogen gas in aquaria systems as a harmless alternative to other electron donors such as sulphur or reduced organic carbon was investigated. To circumvent safety issues regarding the storage of large volumes of hydrogen gas, in situ generation of hydrogen gas by means of a separate electrochemical cell was chosen. A plug flow reactor filled with polyester cotton wool as carrier material for denitrifying bacteria received hydrogen gas in the headspace. Removal rates of 15 +/- 6 mg N per day were achieved with a hydraulic residence time of 3.3 hours. During the start-up phase nitrite peaks up to 3 mg N per liter were measured in the effluent of the reactor. PMID- 15296153 TI - Treatment of waters polluted with crude oil and heavy metals by means of a natural wetland. AB - In the Dolni Dubnik oil deposit, Northern Bulgaria, waters polluted with crude oil and heavy metals (iron, manganese, zinc, cadmium, lead, copper) were treated by means of a natural wetland located in the deposit. The waters had a pH in the range of about 4.5-6.5 and contained about 1-5 mg/l oil. The concentrations of heavy metals usually were about 2-4 times higher than the relevant permissible levels for waters intended for use in the agriculture and industry. The watercourse through the wetland covered a distance of about 100 m and the water flow rate varied in the range of about 0.2-0.8 l/s. The wetland was characterized by an abundant water and emergent vegetation and a diverse microflora, including different oil-degrading bacteria and fungi. The treatment of the polluted waters by means of the above-mentioned wetland markedly depended on the temperature but was efficient during the different climatic seasons, even during the cold winter months at temperatures close to 0 degrees C. The oil content in the wetland effluents in most cases was decreased to less than 0.2 mg/l, and the concentrations of heavy metals were decreased below the relevant permissible levels. The removal of oil was connected with its microbial degradation. The removal of heavy metals was due to different processes but the microbial dissimilatory sulphate reduction and the sorption of metals on the organic matter and clay minerals present in the wetland played the main role. PMID- 15296154 TI - Optimized biogas-fermentation by neural network control. AB - In this work several feed-forward back-propagation neural networks (FFBP) were trained in order to model, and subsequently control, methane production in anaerobic digesters. To produce data for the training of the neural nets, four anaerobic continuous stirred tank reactors (CSTR) were operated in steady-state conditions at organic loading rates (Br) of about 2 kg x m(-3) x d(-1) chemical oxygen demand (COD), and disturbed by pulse-like increase of the organic loading rate. For the pulses additional carbon sources were added to the basic feed (surplus- and primary sludge) to simulate cofermentation and to increase the COD. Measured parameters were: gas composition, methane production rate, volatile fatty acid concentration, pH, redox potential, volatile suspended solids and COD of feed and effluent. A hierarchical system of neural nets was developed and embedded in a Decision Support System (DSS). A 3-3-1 FFBP simulated the pH with a regression coefficient of 0.82. A 9-3-3 FFBP simulated the volatile fatty acid concentration in the sludge with a regression coefficient of 0.86. And a 9-3-2 FFBP simulated the gas production and gas composition with a regression coefficient of 0.90 and 0.80 respectively. A lab-scale anaerobic CSTR controlled by this tool was able to maintain a methane concentration of about 60% at a rather high gas production rate of between 5 to 5.6 m3 x m(-3) x d(-1). PMID- 15296155 TI - Anaerobic bioconversion of organic waste into biogas by hot water treatment at near-critical conditions: application in bioregenerative life support. AB - The feasibility of nearly-complete conversion of lignocellulosic waste (70% food crops, 20% faecal matter and 10% green algae) into biogas was investigated in the context of a Life Support Project. The treatment comprised a series of processes, i.e. a mesophilic laboratory scale CSTR (continuously stirred tank reactor), an upflow biofilm reactor and a hydrothermolysis system in near-critical water. By the one-stage CSTR, a biogas yield of 75% with a specific biogas production of 0.37 l biogas g(-1) VSS (volatile suspended solids) added at a HRT (hydraulic retention time) of 20 d was obtained. Biogas yields further increased with 10-15% at HRT > 20 d, indicating the hydrolysis of lignocellulose to be the rate limiting conversion step. The solids present in the CSTR-effluent were subsequently treated by hot water treatment (T approximately 310-350 degrees C, p approximately 240 bar), resulting in effective carbon liquefaction (50-60% without and 83% with carbon dioxide saturation) and complete hygienisation of the residue. Subsequent anaerobic digestion of the hydrolysate allowed further conversion of 48-60% on COD (chemical oxygen demand) basis. Thus, the total process yielded biogas corresponding with a COD conversion up to 90% of the original organic matter. It appears that mesophilic digestion in conjunction with hydrothermolysis at near-critical conditions offers interesting features for (nearly) complete, non-toxic and hygienic carbon and energy recovery from human waste in a bioregenerative life support context. PMID- 15296156 TI - Optimization of the biodegradation of naphthalene by a microorganism isolated from petroleum contaminated soil. AB - Response surface methodology (RSM) was used to optimise the process parameters to obtain maximum biodegradation of naphthalene. The factorial design employed, a face-centred cube, helped in identifying the combined best parameter conditions. Naphthalene-containing medium was inoculated with Pseudomonas fluorescens and incubated at different levels of agitation, pH, and temperature, and analyzed by Gas Chromatography-Mass Spectroscopy. As a result of the optimisation process, the process parameters found for maximum degradation of naphthalene by P. fluorescens were an agitation speed of 186 rpm, a pH of 7.3, and a temperature of 22.8 degrees C. PMID- 15296157 TI - Biotechnology of deactivation of dioxine (on biological technologies of deactivation of some ecotoxicants). PMID- 15296158 TI - The technology of bioremediation of oil polluted objects by biopreparations (project "biodestructor"). PMID- 15296159 TI - Development of microbial granules for PCB dechlorination. AB - The effect of acclimating anaerobic granules from commercial bioreactors with different carbon/electron sources on their ability to reductively dechlorinate a tri- (2,3,4-CB) and heptachlorobiphenyl (2,2',3,3',4,5,6-CB) was studied. The anaerobic granules were first grown in upflow anaerobic sludge blanket (UASB) reactors fed with two different mixtures of carbon/electron sources, i.e., propionate/butyrate/methanol and formate/methanol. Differences in dechlorination patterns for 2,2',3,3',4,5,6-CB were observed in batch experiments inoculated with granules from these two sets of UASB reactors. The reductive dechlorination of 2,2',3,3',4,5,6-CB for propionate/butyrate/methanol acclimated granules proceeded via 2,2',3,3',4,5-CB for all 2,2',3,3',4,5-CB for all 2,2',3',4,5-CB + 2,2',3',4-CB for all 2,4-CB + 2,3-CB + 2,4'-CB and 2,2',3,3',5,6-CB for all 2,2',3,5,6-CB for all 2,2',3,5-CB for all 2',3,5-CB. The dechlorination pathway for formate/methanol acclimated granules followed: 2,2',3,3',4,5-CB + 2,2',3,3',5,6-CB for all 2,2',3,3',4-CB + 2,2',3,3',6-CB for all 2,2',3,3'-CB for all 2,2',3-CB for all 2,2'-CB. This research demonstrates a successful strategy for the development of biocatalysts to serve as the inoculum of partially decontaminated sites in order to provide microorganisms with specificities complementary to those of naturally occuring dechlorinators. PMID- 15296160 TI - Occurrence and origin of phosphine in landfill gas. AB - A landfill (Hooge Maey, Flanders, Belgium) was subjected to an in-depth study in order to explain the origin of phosphine detected in high amounts in landfill gas during a previous study. The spatial and temporal variability of the phosphine concentration in landfill gas was assessed. Twenty four wells were monitored and differences in phosphine concentration up to one log unit were observed (3.2-32.4 microg/m3). The phosphine concentration in each well was constant in time over a period of 4 months. No correlation was found between the phosphine concentration and methane, carbon dioxide, hydrogen sulphide, ethene or ethane concentration. In a series of laboratory tests, it was shown that phosphine was emitted during batch fermentation tests inoculated with landfill leachate when Fe0 or Al0 specimens were added. Conditions favouring corrosion of iron gave rise to higher emissions of phosphine. The phosphine concentration in the headspace of a batch test rose to 1.43 mg/m3 after 27 days of incubation. Weight loss of corroding steel coupons correlated with phosphine emission. Calculations showed that all phosphine emitted from the 0.005 km3 landfill (160 g/year) could be attributed to corrosion of metals. No evidence of de novo synthesis could be established PMID- 15296161 TI - Degradation of isobutyraldehyde and its intermediates in a compost biofilter. AB - This study demonstrates that at low to medium isobutyraldehyde loading rates (191 gm(-3) d(-1)-933 gm(-3) d(-1)), 100% removal efficiencies can be obtained in a compost biofilter. However, increasing the loading rate to 1500-1900 gm(-3) d(-1) caused a drop in degradation efficiency, a pH decrease and production of isobutyl alcohol and isobutyric acid. Additional batch and continuous experiments were performed to study the effect of pH and compost moisture content on the biofiltration of isobutyraldehyde, isobutyl alcohol and isobutyric acid. It was shown that the degradation rate of the three compounds decreased in the order isobutyraldehyde > isobutyl alcohol >> isobutyric acid, with no significant degradation for isobutyric acid. The isobutyl alcohol degradation rate was negatively influenced by the presence of isobutyraldehyde, while isobutyraldehyde degradation was not affected by the presence of either of the two compounds. A pH of 5.2 apparently inhibited the isobutyl alcohol degradation and lowered the isobutyraldehyde degradation rate, although adaptation of the microorganisms to low pH seemed to occur in the biofilters. Moisture content had a smaller effect on the degradation rates, although continuous experiments showed that a very high water content (55% compared to 40%) negatively affected isobutyraldehyde elimination increasingly during the course of the experiment. As a conclusion, it appears that at high loads of isobutyraldehyde, isobutyric acid is accumulated in the biofilter, resulting in a drop of pH. Consequently, isobutyraldehyde removal efficiency decreases and both isobutyl alcohol and isobutyric acid are measured in the effluent. It is suggested that next to moisture control, a pH buffer is necessary to remove high loads of isobutyraldehyde and to avoid persistence of intermediates in the effluent. PMID- 15296162 TI - The enhancement of reproduction and biodegradation activity of eukaryiotic cells by humic acids. AB - Fourteen samples of humic acids (HA) were screened for ability to influence reproduction and biodegradation activity of eukaryotic cells in the presence of chosen toxic pollutants. Microorganisms Candida maltosa and Rhodotorula mucilaginosa (soil isolates) were used for all tests. It was observed during our experiments that some samples of humic acids served as a protection against the high concentration of toxic pollutants (phenol, naphtalene etc). This effect can be widely used in many bioremediation technologies. PMID- 15296163 TI - Modelling of the production of gaseous by-products in anaerobic digestion. AB - Goal of the EU-Project AMONCO (Advanced Prediction, Monitoring and Controlling of Anaerobic Digestion Processes Behaviour towards Biogas Usage in Fuel Cells) is demonstration of the practical use of biogas in fuel cells. The right precondition is a biogas quality which fits into the fuel cells tolerances. Therefore the mission of the workgroup Environmental biotechnology is to control anaerobic digestion in a way that production of potential harmful by-products for fuel cells is reduced. A good understanding of the production of these by products is essential for an applicable decision support tool. This poster presents the modelling of hydrogen sulfide by means of hierarchical neural networks and a classical mathematical method. PMID- 15296164 TI - Comparative biodegradation examination of Pseudomonas aeruginosa (ATCC 27853) and other oil degraders on hydrocarbon contaminated soil. AB - The soil that we investigated in our experiment was extremly high contaminated with petroleum hydrocarbons. The sample was originated from a former truck factory, next to an oil tank. We tried on this sample three different biodegradation treatments in a soil respirometer. The aim of the experiment was investigate if the clinical strain Pseudomonas aeruginosa ATCC 27853 is able to utilize petroleum hydrocarbons as carbon source. In spite of the relevant literature Pseudomonas aeruginosa ATCC 27853 (clinical isolate, from blood) was proven to be a good oil degrader. PMID- 15296165 TI - Examination of the long-term process for purifying gaseous discharges in industrial biofilter. AB - Results of industrial exploitation of the biofilter unit for purifying gaseous discharges of hazardous organic substances are examined. A population of microbial associated composed by selected strains of yeast and bacterial cells in the biofilm on the surface of the filtering sheets and in the liquid phase were monitoring for a long-term (about 2 years) utilization process in industrial biofilter. Overall, the biofilter with productivity 12,000-15,000 m3 purified flow per hour and very low energy consumption (0.3-0.4 wt.h/m3) as an open and autonomic system maintained its microbial association, thereby providing high degree (93-96%) purification of industrial gaseous discharges from organic pollutants. PMID- 15296166 TI - A fuzzy logic approach to control anaerobic digestion. AB - One of the goals of the EU-Project AMONCO (Advanced Prediction, Monitoring and Controlling of Anaerobic Digestion Process Behaviour towards Biogas Usage in Fuel Cells) is to create a control tool for the anaerobic digestion process, which predicts the volumetric organic loading rate (Bv) for the next day, to obtain a high biogas quality and production. The biogas should contain a high methane concentration (over 50%) and a low concentration of components toxic for fuel cells, e.g. hydrogen sulphide, siloxanes, ammonia and mercaptanes. For producing data to test the control tool, four 20 l anaerobic Continuously Stirred Tank Reactors (CSTR) are operated. For controlling two systems were investigated: a pure fuzzy logic system and a hybrid-system which contains a fuzzy based reactor condition calculation and a hierachial neural net in a cascade of optimisation algorithms. PMID- 15296167 TI - Biofiltration in an immobilised system for processing gaseous industrial effluent. PMID- 15296168 TI - Factors influencing biofilm formation of phenol degrader Candida maltosa. PMID- 15296169 TI - From extreme environments to biologically active exopolysaccharides. AB - In the course of the discovery of novel polysaccharides of biotechnological interest, it is now widely accepted that extremophilic microorganisms will provide a valuable resource not only for exploitation in novel biotechnological processes but also as models for investigating how biomolecules are stabilized when subjected to extreme conditions. Microbes isolated from extreme environments offer a great diversity in chemical and physical properties of their EPS as compared to anywhere else in the biosphere. Bacteria from remote areas still remain virtually unexplored and there is not doubt that extreme environments are a rich source of microorganisms of biotechnological importance. A number of interesting and unique bacterial polysaccharides have been isolated from these ecosystems and are expected to find applications in the very near future in different industrial. Further screenings are underway as well as research into understanding the structure-function relationships of these unusual polymers. PMID- 15296170 TI - Metabolic engineering of Escherichia coli: construction of an efficient biocatalyst for D-mannitol formation in a whole-cell biotransformation. AB - A whole-cell biotransformation system for the conversion of D-fructose to D mannitol was developed in Escherichia coli by construction of a recombinant oxidation/reduction cycle. First, the mdh gene encoding for the mannitol dehydrogenase of Leuconostoc pseudomesenteroides ATCC 12291 (MDH) was expressed, effecting a strong catalytic activity of a NADH-dependent reduction of D-fructose to D-mannitol in cell extracts of the recombinant E. coli strain but not enabling whole cells of the strain to produce D-mannitol from D-fructose. To provide a source for reduction equivalents needed for D-fructose reduction, the fdh gene from Mycobacterium vaccae N10 (FDH) encoding formate dehydrogenase was functionally co-expressed. FDH generates NADH used for D-fructose reduction by dehydrogenation of formate to carbon dioxide. These recombinant E. coli cells were able to form D-mannitol from D-fructose in a low but significant quantity (15 mM). The introduction of a further gene, encoding for the glucose facilitator protein of Zymomonas mobilis (GLF) enabled the cells to efficiently take up D fructose into the cells, without simultaneous phosphorylation. Resting cells of this E. coli strain (3 g cell dry weight/l) produced 216 mM D-mannitol in 17 hours. Biotransformations conducted under pH-control by formic acid addition yielded D-mannitol at a concentration of 362 mM within 8 hours. The yield Y(D mannitol D-fructose) was 84 [mol%]. These results show that the recombinant strain of E. coli can be utilized as an efficient biocatalyst for D-mannitol formation. PMID- 15296171 TI - Dynamics and optimal conditions of intracellular ectoine accumulation in Brevibacterium sp. AB - The optimal conditions for the intracellular synthesis of ectoine were determined in a halotolerant Brevibacterium sp. The size of the intracellular ectoine pool in the bacterial cells is shown to depend on the external salt concentrations, type of carbon source and aeration level. In erlenmeyer flasks a maximum concentration of intracellular ectoine of about 0.9 g/l was obtained. Under controlled aeration in a 1.5 l fermentor this level could be increased to 1.2 g/l. Consecutive cell transfers to media with increasingly higher salt concentrations enabled us to reach even higher levels, up to 1.6 g/l on erlenmeyer scale. The ectoine synthesis takes place immediately after the osmotic upshock. Within one generation time, the new corresponding specific intracellular ectoine concentration is reached. PMID- 15296172 TI - A pervaporation-bio-hybridreactor (PBHR) for improved aroma biosynthesis with submerged culture of Ceratocystis fimbriata. AB - In the present study a Pervaporation-bio-hybridreactor was investigated for the improved biotechnological production of volatile flavour compounds. The mixture of esters and alcohols produced by the fungus Ceratocystis fimbriata had a fruity, banana-like odour and can legally be defined as "natural flavour". Increasing consumer demand and high market prices make the production of bioflavours an attractive alternative to chemical synthesises. However, the yield of biotechnological flavour production is often very low due to product inhibition. In situ product-removal processes can yield higher productivities by preventing product inhibition. A bioreactor with an integrated pervaporation membrane was used to determine the effects of in-situ product removal on the metabolic productivity of C. fimbriata. Fed-batch fermentations with and without integrated product removal were conducted and the performance of the processes in terms of productivities was compared. Improved productivities were observed for all products. Figure 1a and 1b show the increased amount of product gained through the application of the PBHR with fed-batch fermentation. PMID- 15296173 TI - Utilization of grape must for gluconic acid production using polyurethane sponge and calcium alginate immobilized cells of Aspergillus niger ORS-4.410. PMID- 15296174 TI - Application of NAD-dependent polyol dehydrogenases for enzymatic mannitol/sorbitol production with coenzyme regeneration. AB - D-Mannitol and D-sorbitol were produced enzymatically from D-fructose using NAD dependent polyol dehydrogenases. For the production of D-mannitol the Leuconostoc mesenteroides mannitol dehydrogenase could be used. Gluconobacter oxydans cell extract contained however both mannitol and sorbitol dehydrogenase. When this cell extract was used, the reduction of D-fructose resulted in a mixture of D sorbitol and D-mannitol. To determine the optimal bioconversion conditions the polyol dehydrogenases were characterized towards pH- and temperature-optimum and stability. As a compromise between enzyme activity and stability, the bioconversion reactions were performed at pH 6.5 and 25 degrees C. Since the polyol dehydrogenases are NADH-dependent, an efficient coenzyme regeneration was needed. Regeneration of NADH was accomplished by formate dehydrogenase-mediated oxidation of formate into CO2. PMID- 15296175 TI - Pycnoporus cinnabarinus laccases: an interesting tool for food or non-food applications. AB - The effects of the addition of ferulic acid and ethanol in P. cinnabarinus ss3 culture medium in fermentor were compared in 15-L fermentor. In the presence of 30 g l(-1) ethanol, laccase activity (270,000 U/L1) was 3-fold higher as compared with ferulic acid-induced cultures, and 150-fold higher as compared with non induced cultures, respectively. High-quality flax pulp was bleached in a totally chlorine free (TCF) sequence using a laccase-mediator system constituted by laccase from Pycnoporus cinnabarinus and 1-hydroxybenzotriazole (HBT) as mediator. Up to 90% delignification and strong brightness increase were attained after the laccase-mediator treatment followed by H2O2 bleaching. This TCF sequence was further improved by applying H2O2 under pressurized O2. In this way, up to 82% ISO brightness was obtained (compared with 37% in the initial pulp and 60% in the peroxide-bleached control) as well as very low kappa number. A positive evaluation of the laccase has been also performed in a food application. The colour of a tea-based beverage was significantly improved by incubating an infusion of green tea with the Pycnoporus laccase. PMID- 15296176 TI - Molecular analysis and functionality of pectin. PMID- 15296177 TI - Characteristics of a newly isolated fungus Geotrichum candidum Dec 1 with broad degradation spectrum of xenobiotic compounds. AB - A newly isolated fungus, Geotrichum candidum Dec 1 (abbreviated as Dec 1), was found to have the ability to degrade many xenobiotic compounds such as synthetic dyes, food coloring agents, molasses, organic halogens, lignin and kraft pulp effluents. The broad spectrum of the degradation of these compounds are associated mainly with peroxidases produced by the fungus. PMID- 15296178 TI - Oligosaccharides and derivatives--integrating biocatalyst selectivity and chemical diversity. AB - Several biocatalytic pathways are available to synthesize rare sugars, oligosaccharides and derivatives. As examples selective hydrolysis, transglycosylation and regioselective oxidation are presented illustrating their potential. From inulin, difructose anhydrides and inulobiose are accessible via specific hydrolases with transglycosylation activity. Glycosyltransferases of the sucrase type utilise sucrose as a versatile substrate for the selective transfer of either glucosyl- or fructosyl units to a broad range of acceptors, sugars as well as derivatives, to yield oligosaccharides of different type. Kinetic analysis and reaction engineering can provide high product yields and concentration, as will be shown for glucose, yielding isomaltooligosaccharides as an example. Several new acceptor saccharides, including sugar alcohols and acids, have been shown to give new oligosaccharides. Regioselective oxidation of disaccharides like sucrose, lactose and others with resting cells of Agrobacterium tumefaciens yield 3-keto-disaccharides. These can further be functionalized in aquous systems without protecting groups via established chemical routes, such as catalytic hydration. As an example allosucrose is thus obtained from 3-keto-sucrose with high yield and stereochemical selectivity, which in turn can easily be hydrolysed and separated from fructose to give allose. Amino acid and peptide conjugates are accessible via reductive amination and acylation, as are building blocks for polymerisation. PMID- 15296179 TI - Reducing environmental pollution using animal feed enzymes. AB - The global livestock population is estimated to be close to 4 billion animals, and to produce around 500 million tons of manure annually (Baidoo, 2003). This is expected to increase in the future with the projected greater demand for meat for human consumption. The problem of manure disposal is exacerbated by the concentration of animal production in increasingly large units, to obtain economies of scale and keep up with the demand for cheap food. The primary environmentalfactors are manure volume, manure nitrogen and phosphorus contents, methane production and odour (Jongbloed and Lenis, 1998). Legislation in many regions now restricts the amount of manure that can be applied per hectare, to prevent environmental pollution (Centner, 2001; Pellini and Morris, 2001). There are a number of strategies the animal production industry can take to reduce environmental impact. These include taking steps to improve the efficiency of conversion of feed into edible products, reduce feed wastage and formulate diets that more closely satisfy animal requirements for specific nutrients. At present 50-80% of the nitrogen and phosphorus fed to animals are not utilized but are excreted via manure and urine to the environment (Baidoo, 2003). Biotechnology could play a very important role in reducing the environmental impact of animal production. Examples include the development of animals more efficient at converting nutrients into edible products, and of higher quality, more digestible feedstuffs. Biotechnology can also be used to produce a range of feed additives that can improve the efficiency of animal production, including for example recombinant somatrophin, amino acids and enzymes. This paper summarizes a series of four experiments looking at the effects of microbial xylanase or phytase supplementation on excretion in swine and poultry. This summary indicates that the inclusion of these enzymes in animal feeds can reduce manure volume by up to 14%, and nitrogen and phosphorus outputs by up to 13% and 70%, respectively. PMID- 15296180 TI - Thermal stability enhancements of Candida rugosa lipase in ionic liquids. AB - Thermostability of Candida rugosa lipase in organic solvents and ionic liquids was studied. During our experiments the aim was to determine if the conversion degree and enzyme half-life are changed, i.e. how to tolerate the enzyme the elevated temperature in various solvents. It was found that the enzyme kept its activity and enantioselectivity much better in ionic liquids than in the traditional organic solvents. PMID- 15296181 TI - Combined enzymatic hydrolysis and HPAEC method for simultaneous analysis of galacturonic acid and neutral sugars of pectin. AB - Pectic substances are heteropolysaccharides from plant cell walls, which mainly consist of a homogalacturonan backbone of predominantly alpha-(1-->4) linked galacturonic acid (GalA) residues. This chain is interrupted by ramified rhamnogalacturonan regions with a certain amount of neutral sugars (rhamnose, arabinose, galactose, glucose, xylose and mannose) present as side-chains. The GalA residues can be methyl-esterified at the carboxyl group. Usually uronic acids of pectin are determined as anhydrogalacturonic acid by spectrophotometry, using the metahydrodiphenyl method (Thibault, 1979), while the monosaccharides are determined by High-Performance Liquid Chromatography (HPLC) after a methanolysis (Quemener and Thibault, 1990) or by Gaz Liquid Chromatography (GLC) as alditol acetates according to Blakeney et al. (1983). In this paper, a combined enzymatic hydrolysis and HPLC method is validated for simultaneous analysis of galacturonic acid and neutral sugars without any derivatisation. PMID- 15296182 TI - Regulation of cellulase and hemicellulase synthesis in the fungus Chrysosporium sp. PMID- 15296183 TI - Dextransucrase mutants of Leuconostoc mesenteroides BI-08 strain. AB - We have isolated mutants constitutive for dextransucrase from Leuconostoc mesenteroides--BI-08 using ethyl methane sulfonate. The mutants produced mainly extracellular dexrtansucrase on glucose media with higher activity (3.5-4 times) than what the parental strain produced on glucose and 2 times more than what the parental strain produced on sucrose. Based on endo-dextranase from Penicillium hydrolysis, mutant BI-08 dextransucrases produced slightly different dextrans when they were elaborated on a glucose medium and on a sucrose medium Differences in viscosity, water solubility, susceptibility to endo-dextranase hydrolysis dextrans elaborated by different mutants grown on glucose media and sucrose media were found. SDS-PAGE analysis shows that the molecular masses of the dextransucrases from the studing mutants were 130-180 kDa. PMID- 15296184 TI - Stereoselective oxidation of aliphatic diols and reduction of hydroxy-ketones with galactitol dehydrogenase from Rhodobacter sphaeroides D. AB - From the Rhodobacter sphaeroides mutant D a galactitol dehydrogenase (GDH) was isolated and characterized in an earlier investigation (1). The enzyme expressed activity with a wide spread substrate spectrum, like sugars, sugar alcohols, secondary alcohols or the corresponding ketones and it can be used for the production of the rare sugar L-tagatose by regioselective oxidation of galactitol (2). This study focuses on the preparation of optically pure aliphatic diols by oxidation of one enantiomer or stereospecific reduction of keto-alcohols and diketones. The oxidation of 1,2-propanediol, 1,2-butanediol, 1,2-pentanediol and 1,2-hexanediol occurred highly specific with the S-enantiomer leaving the R enantiomer of the diols in the reaction vessel. Also (S)-1,2,6-hexanetriol was oxidized by GDH to 1,6-dihydroxy-2-hexanone. The Km values of these reactions decreased with increasing length of the carbon chain. Reduction of hydroxyacetone or 1-hydroxy-2-butanone resulted in an excess of 93% (S)-1,2-propanediol and more than 98% of (S)-1,2-butanediol, respectively. The diketone 2,3-hexanedione was only reduced to (2R,3S)-2,3-hexanediol, one of the possible four configurations. The wide substrate spectrum on one hand and the selectivity in the reaction on the other hand make GDH a very interesting enzyme for the production of optically pure building blocks in the chemical synthesis of bioactive compounds. PMID- 15296185 TI - Physiological properties and enzymatic activities of Rhizopus oligosporus in solid state fermentations. AB - The physiology of fungus Rhizopus oligosporus in the solid state fermentation of Pisum sativum was investigated by means of digital analysis of microscopic image. The correlations between the hyphal fractions within physiological zones, the release of glucose and soluble proteins, and enzymatic activities of the examined strain were also estimated. PMID- 15296186 TI - The application of fungal endoxylanase in bread-making. AB - The endoxylanase from Aspergillus niger IBT-90 with the specific activity of 21.3 U/mg protein was used for improvement of wheat-rye and whole-meal bread. The addition of this xylanase in the range 500-1000 U/kg flour resulted in improvement of kneading and increase of loaf volume in the case of kinds of bread. This dough supplementation caused better crumb porosity and higher moisture in bread and finally the shelf life is extended. No increase of total acidity was resulted from the enzyme addition. The experimental results have confirmed the applicability of this xylanase in bakery. PMID- 15296187 TI - Catalytic activity of zymomonas mobilis extracellular "levan-levansucrase" complex in sucrose medium. AB - The fructan biosynthesis by ethanol sedimented "levan-levansucrase" complex from Zymomonas mobilis fermentation broth as well as purified levansucrase was investigated. The fructooligosaccharide (FOS) producing activity of "levan levamsucrase" sediment was investigated in 55% sucrose syrup at 45 degrees C. It was shown that FOS in the syrup were presented by 1-kestose, 6-kestose, neokestose and nystose. The increase of gluconic acid concentration was observed in the reaction mixture during the incubation suggesting about presence of glucose/fructose oxidoreductase in "levan-levansucrase" sediment. The influence of ethanol, glycerol and NaCl on levan and fructooligosaccharide formation by "levan-levansucrase" complex and purified levansucrase was studied and the changes in the ratio between different activities of levansucrase (sucrose hydrolysis, levan biosynthesis and FOS formation) were observed. Ethanol increases the FOS biosynthesis part in total activity of purified levansucrase. The technology of the production of prebiotics containing food product--fructan syrup by "levan-levansucrase " sediment as biocatalyst was developed. PMID- 15296188 TI - Microbial biosynthesis and making of pigment melanin. AB - The efficient strain and the technology of biosynthesis and extraction process from a biomass received in process cultivation of new yeast strain Saccharomyces neoformans var. nigricans are developed. The influence of a carbon substrates (glucose, spirit, milk whey) and conditions of fermentation process on an output and chromatically received melanin are investigated. It is is established, that for the given strain the fermentation time makes 36-40 hours. PMID- 15296189 TI - Enzymatic conversion of the clavan exopolysaccharide by Streptomyces sp. YSDL-20. AB - A screening programme was set up to isolate microorganisms able to hydrolyse the complex biopolymer clavan produced by Clavibacter michiganensis subsp. michiganensis LMG 5604. This valuable exopolysaccharide is very rich in L-fucose (37.5% w/w), a rare sugar, used in the medical field (Vanhooren, 1999). A microorganism capable of depolymerizing the polymer may decrease the high viscosity during clavan batch fermentations and remove the limitations of the oxygen transfer and consequently increase the clavan yield. It could also release free L-fucose or L-fucose rich oligosaccharides. An actinomycete, designated YSDL 20, isolated from a soil sample, was able to depolymerize this biopolymer. Based on its morphology and molecular characteristics, this strain could only be identified as Streptomyces sp.. On clavan, this strain displays good growth (17.5 g DCW/l after 96 h of cultivation) characterized by filamentous growth during the earlier days of cultivation followed by sporulation after 4 days. The flow behaviour of the Clavibacter broth was characterized, the fermentation culture broth behaves as a pseudoplastic fluid. The viscosity of the culture broth as well as of the purified clavan EPS, decreases when lyophilised supernatant of Streptomyces sp. YSDL-20 was added, indicating clavanase action. The viscosity decreases by 26% when the Clavibacter culture broth was incubated during 18 h with the crude Streptomyces enzyme source, whereas a 82% viscosity drop was observed, when the purified clavan EPS (10 g/l) was incubated with the lyophilised Streptomyces supernatant for 5 h. PMID- 15296190 TI - Nurse-patient ratios revisited. PMID- 15296191 TI - Teen camp: a unique approach to recruit future nurses. AB - TOPIC: A collaborative and unique approach to interest high school students in nursing. PURPOSE: To inform educators and nursing departments about an innovative approach to recruit future nurses. SOURCES: Professional literature and authors' experience. CONCLUSIONS: All students related positive experiences. The initial camp evaluation produced innovative input from the students, and each camp met its goal of creating career interest in the nursing profession. PMID- 15296192 TI - Risk: a concept analysis. AB - TOPIC: Concept analysis of risk. PURPOSE: To analyze the concept risk and provide a new definition of risk. SOURCES: Published literature. CONCLUSIONS: A new definition of risk that emerged from this concept analysis can provide clarity and direction for future research. Nurse researchers can look to this definition to expand what is known about health-seeking behaviors as opposed to "risk" behaviors and seek to further our understanding of the cognitive and experiential process of risk identification. PMID- 15296193 TI - Perceived risk of heart attack: a function of gender? AB - TOPIC: The influence of gender on women's risk beliefs for heart attack. PURPOSE: To inform healthcare providers how women's beliefs and attitudes contribute to treatment-seeking delay in the event of a heart attack, and to establish the importance of risk beliefs in women. SOURCES: Published literature in MEDLINE and CINAHL computerized databases, reference lists of obtained articles. CONCLUSIONS: Women's perceived risk beliefs for heart attack are influenced by the effects of media, cultural, and gender roles and the modeling of bias in health care. There is a need for healthcare providers to change their focus from acting on (etic) a patient to interacting within (emic) the belief systems of their patients to optimize positive outcomes. PMID- 15296194 TI - Concept mapping: an educational strategy for advancing nursing education. AB - TOPIC: Application of concept mapping as a tool in nursing education. PURPOSE: To highlight the use of concept mapping as a method for advanced learning in nursing education. SOURCES OF INFORMATION: Literature from nursing and general education, instructor and student experiences, and opinions from using concept maps as a method of teaching/learning. CONCLUSIONS: Instructors and students reported satisfaction from use of concept maps in the educational process. Teaching with the aid of concept maps has been incorporated as an innovative and viable teaching method in nursing education. PMID- 15296195 TI - Regarding abortion: informed consent or selective disclosure? PMID- 15296196 TI - Stories: caring about caring. PMID- 15296197 TI - Invest in yourself: where has "patient care" gone? PMID- 15296198 TI - Implementing capsule representation in a total hip dislocation finite element model. AB - Previously validated hardware-only finite element models of THA dislocation h ave clarifiedhow various component design and surgical placement variables contribute to resisting the propensity for implant dislocation. This body of work has now been enhanced with the incorporation of experimentally based capsule representation, and with anatomic bone structures. The current form of this finite element model provides for large deformation multi-body contact (including capsule wrap-around on bone and/or implant), large displacement interfacial sliding, and large deformation (hyperelastic) capsule representation. In addition, the modular nature of this model now allows for rapid incorporation of current or future total hip implant designs, accepts complex multi-axial physiologic motion inputs, and outputs case-specific component/bone/soft-tissue impingement events. This soft-tissue-augmented finite element model is being used to investigate the performance of various implant designs for a range of clinically-representative soft tissue integrities and surgical techniques. Preliminary results show that capsule enhancement makes a substantial difference in stability, compared to an otherwise identical hardware-only model. This model is intended to help put implant design and surgical technique decisions on a firmer scientific basis, in terms of reducing the likelihood of dislocation. PMID- 15296199 TI - Professionalism for medicine: opportunities and obligations. AB - Physicians' dual roles--as healer and professional--are linked by codes of ethics governing behaviour and are empowered by science. Being part of a profession entails a societal contract. The profession is granted a monopoly over the use of a body of knowledge and the privilege of self-regulation and, in return, guarantees society professional competence, integrity and the provision of altruistic service. Societal attitudes to professionalism have changed from supportive to increasingly critical--with physicians being criticised for pursuing their own financial interests, and failing to self-regulate in a way that guarantees competence. Professional values are also threatened by many other factors. The most important are the changes in healthcare delivery in the developed world, with control shifting from the profession to the State and/or the corporate sector. For the ideal of professionalism to survive, physicians must understand it and its role in the social contract. They must meet the obligations necessary to sustain professionalism and ensure that healthcare systems support, rather than subvert, behaviour that is compatible with professionalism's values. PMID- 15296200 TI - Oxygen effects on senescence in chondrocytes and mesenchymal stem cells: consequences for tissue engineering. AB - Primary isolates of chondrocytes and mesenchymal stem cells are often insufficient for cell-based autologous grafting procedures, necessitating in vitro expansion of cell populations. However, the potential for expansion is limited by cellular senescence, a form of irreversible cell cycle arrest regulated by intrinsic and extrinsic factors. Intrinsic mechanisms common to most somatic cells enforce senescence at the so-called "Hayflick limit" of 60 population doublings. Termed "replicative senescence", this mechanism prevents cellular immortalization and suppresses oncogenesis. Although it is possible to overcome the Hayflick limit by genetically modifying cells, such manipulations are regarded as prohibitively dangerous in the context of tissue engineering. On the other hand, senescence associated with extrinsic factors, often called "stress-induced" senescence, can be avoided simply by modifying culture conditions. Because stress-induced senescence is "premature" in the sense that it can halt growth well before the Hayflick limit is reached, growth potential can be significantly enhanced by minimizing culture related stress. Standard culture techniques were originally developed to optimize the growth of fibroblasts but these conditions are inherently stressful to many other cell types. In particular, the 21% oxygen levels used in standard incubators, though well tolerated by fibroblasts, appear to induce oxidative stress in other cells. We reasoned that chondrocytes and MSCs, which are adapted to relatively low oxygen levels in vivo, might be sensitive to this form of stress. To test this hypothesis we compared the growth of MSC and chondrocyte strains in 21% and 5% oxygen. We found that incubation in 21% oxygen significantly attenuated growth and was associated with increased oxidant production. These findings indicated that sub-optimal standard culture conditions sharply limited the expansion of MSC and chondrocyte populations and suggest that cultures for grafting purposes should be maintained in a low-oxygen environment. PMID- 15296201 TI - Adaptive meshing technique applied to an orthopaedic finite element contact problem. AB - Finite element methods have been applied extensively and with much success in the analysis of orthopaedic implants. Recently a growing interest has developed, in the orthopaedic biomechanics community, in how numerical models can be constructed for the optimal solution of problems in contact mechanics. New developments in this area are of paramount importance in the design of improved implants for orthopaedic surgery. Finite element and other computational techniques are widely applied in the analysis and design of hip and knee implants, with additional joints (ankle, shoulder, wrist) attracting increased attention. The objective of this investigation was to develop a simplified adaptive meshing scheme to facilitate the finite element analysis of a dual curvature total wrist implant. Using currently available software, the analyst has great flexibility in mesh generation, but must prescribe element sizes and refinement schemes throughout the domain of interest. Unfortunately, it is often difficult to predict in advance a mesh spacing that will give acceptable results. Adaptive finite-element mesh capabilities operate to continuously refine the mesh to improve accuracy where it is required, with minimal intervention by the analyst. Such mesh adaptation generally means that in certain areas of the analysis domain, the size of the elements is decreased (or increased) and/or the order of the elements may be increased (or decreased). In concept, mesh adaptation is very appealing. Although there have been several previous applications of adaptive meshing for in-house FE codes, we have coupled an adaptive mesh formulation with the pre-existing commercial programs PATRAN (MacNeal-Schwendler Corp., USA) and ABAQUS (Hibbit Karlson and Sorensen, Pawtucket, RI). In doing so, we have retained several attributes of the commercial software, which are very attractive for orthopaedic implant applications. PMID- 15296202 TI - Brucella osteomyelitis of the proximal tibia: a case report. AB - Brucellosis is a disease of domestic and wild animals that is transmittable to humans. Although endemic in some parts of the world, brucellosis is an uncommon human pathogen in the United States. The clinical presentation of brucellosis is nonspecific, and brucella osteomyelitis can produce lytic lesions on radiographs that resemble neoplasm. Diagnosis can therefore be difficult unless a high index of suspicion is maintained. We present a case of brucella osteomyelitis of the proximal tibia that demonstrates these features. PMID- 15296203 TI - Heterogeneity in growth properties of the rat Swarm chondrosarcoma. AB - Chondrosarcoma remains one of the most difficult clinical conundrums of orthopaedic pathology, with wide variation in clinical course. The natural history of chondrosarcoma ranges from slow indolent growth without metastasis over years to rapid proliferation and lethal metastasis. Molecular regulatory events in the growth of these neoplasms are poorly understood. Of the Swarm rat chondrosarcoma, originating from a single neoplasm in a Sprague-Dawley rat more than thirty-five years ago, two populations were identified with different growth properties. These two Swarm chondrosarcoma lines were characterized for growth properties, histomorphometric and ultrastructural integrity, and the ability for proteoglycans to form aggregates with hyaluronan. After careful comparison, no obvious clues to the variation in growth rate were noted. Further molecular analyses may lead to better understanding of the differential growth properties of these cell lines. Understanding the mechanisms involved in differential growth rates may lead to clinically applicable clues to predict clinical behavior of chondrosarcomas in humans. PMID- 15296204 TI - Effect of chemotherapy on segmental bone healing enhanced by rhBMP-2. AB - Segmental bone defects are challenging clinical problems, and current surgical solutions are associated with high complication rates. In oncologic reconstructive surgery, bone healing will occur coincidently with the administration of chemotherapy to treat the underlying disease. Effective methods of graft modification or bone graft alternatives can be of great help clinically. A series of osteoinductive proteins (bone morphogenetic proteins or BMPs) has been described and shown to enhance bone formation in animal models. This study was designed to evaluate the effect of chemotherapy on bone healing enhanced by rhBMP-2. We used a critical-sized bone-defect rabbit model. Histological and radiological analysis showed that chemotherapy affects both the quantity and the quality of the bone enhanced by the addition of rhBMP-2. These results suggest that the effect of chemotherapy on bone formation could be related to inhibition in a specific pathway stimulated by the rhBMP-2. PMID- 15296205 TI - Medial translation of the hip joint center associated with the Bernese periacetabular osteotomy. AB - This study assessed medial translation of the hip joint achieved by the Bernese periacetabular osteotomy (PAO) in correcting residual acetabular dysplasia deformities. 86 hips in 75 patients with an average age of 25 years (range, 12 50) were treated for symptomatic acetabular dysplasia with a periacetabular osteotomy. Radiographic analysis was performed to assess correction of the acetabular deformity with specific attention to the horizontal position of the hip joint center. All hips were followed until bony union of the iliac osteotomy and the average follow-up was 28 months. The lateral center edge angle improved an average 31.6 degrees (-0.4 degrees preoperative, 31.2 degrees at follow-up). The anterior center edge angle improved 39.3 degrees (-4.5 degrees to 34.8 degrees). The acetabular roof obliquity improved an average 21.8 degrees (25.1 degrees to 3.3 degrees). Preoperatively, the average distance from the medial aspect of the femoral head to the ilioischial line was 17.6 mm. This distance was decreased to an average 7.8 mm postoperatively. This change resulted in an average medial translation of the hip joint center of 9.8 mm, (range -6 to 31mm). Overall, some degree of medial translation of the hip joint center was obtained in 79 (92%) of the hips. Four (5%) were maintained in the same horizontal position, and 3 (3%) had slight lateral repositioning. For the hips translated medially, the average change was 10.0 mm, and 72% of all hips had an optimal correction with the distance between the medial aspect of the femoral head and the ilioischial line being between 0 and 10 mm. This study demonstrates that in addition to optimizing femoral head coverage, a major and distinct advantage of the periacetabular osteotomy is reproducible and consistent medial translation of the hip joint center. PMID- 15296206 TI - Ulnohumeral arthroplasty. AB - Seven patients underwent 9 ulnohumeral arthroplasties for degenerative arthritis of the elbow. At mean follow-up of 26 months, 5 elbows were pain free; two continued to cause mild pain and one to cause moderate pain. Extension improved from 22 degrees+/-8 degrees preoperatively to 12 degrees+/-9 degrees postoperatively (p=0.02); the average correction was 10 degrees+/-10 degrees. Flexion improved from 122 degrees+/-8 degrees to 133 degrees +/- 8 degrees (p=0.02); the average correction was 11 degrees+/-11 degrees. One patient had a late supracondylar humerus fracture which healed well with open reduction and internal fixation. Overall, we believe that ulnohumeral arthroplasty is relatively safe and easy to perform. Our patients did have modest improvements in range of motion, but complete relief of pain occurred in only about two thirds of the patients. PMID- 15296207 TI - Complications of treating distal radius fractures with external fixation: a community experience. AB - OBJECTIVE: To analyze the immediate postoperative complications associated with treating distal radius fractures with external fixation. DESIGN: A retrospective chart review of data obtained from 24 consecutive patients who were treated with small AO external fixators in 1997. SETTING: Two community medical centers. INTERVENTION: Preoperative and postoperative radiograph measurements were taken of radial inclination, radial tilt, and radial length, and fractures were classified according to the AO system. Patient charts were reviewed to document demographics, type of fixator used, open or percutaneous technique for pin placement, use of augmentation, additional operations, and complications. MAIN OUTCOME MEASUREMENTS: Complications associated with treating distal radius fractures with one type of external fixator. RESULTS: Sixteen of the 24 patients had complications: 5 with neuropathies of the median or superficial radial nerve, 9 with pin track infections, 2 with pin loosening, one with a nonunion, 2 with malunion, and 4 patients each with radial shortening, loss of radial tilt, collapse of ulnar border or volar intercalated segment instability (VISI) of the lunate and rotatory subluxation of the scaphoid. CONCLUSIONS: Postoperative complications following distal radius fractures treated with external fixation are common. Their effect, however, on long term functional results and patient satisfaction is negligible, with the exception of those patients with complications intrinsic to the fracture itself, i.e., nonunion, malunion or carpal malalignment. PMID- 15296208 TI - Leg length discrepancy in unilateral congenital clubfoot following surgical treatment. AB - Length discrepancy secondary to limb hypoplasia has been described as an associated finding in patients with unilateral clubfoot. In this manuscript we bring attention to limb length discrepancy as a result of surgical treatment in unilateral clubfoot. Three patients who underwent extensive posterior, medial and lateral release were noted to have an average discrepancy in foot height of 2.1 centimeters (range, 2.0-2.3 centimeters). A decrease in foot height in addition to baseline limb hypoplasia may lead to a significant discrepancy that may justify surgical treatment. In this manuscript we point out that length discrepancy in such cases may not be adequately quantified on standard anteroposterior scanograms. Standing lateral foot radiographs will document loss in foot height as a possible factor in length discrepancy in surgically treated clubfoot patients. PMID- 15296209 TI - Spine height and disc height changes as the effect of hyperextension using stadiometry and MRI. AB - STUDY DESIGN: In vivo biomechanical design using stadiometry and MRI to measure the height change due to (hyper)extension. SUMMARY OF BACKGROUND DATA: Spine height is decreased under loads such as lifting, whole body vibration and sitting. Extension including increased lumbar lordosis reduces the load on the spine. METHODS: The aim was to assess the effects of a supine hyperextended posture as a means of restoring the intervertebral disc height after loading and allowing rehydration of the discs. Ten healthy male subjects were tested. A hyperextension intervention was achieved by the means of an inflatable cushion placed under the lumbar spine. The spine height was measured using a stadiometer and MRI was used to assess disc height changes. RESULTS: The spine height gain after 10 minutes of a supine hyperextended posture differed significantly between individuals but everybody gained height. MRI images of the lumbar spine were used to measure the disc height. All but one subjects gained height during the hyperextension. Images of the spine during hyperextended posture showed increased lumbar curve and an increased anterior height of each disc compared with the dimensions of the disc with the spine in neutral posture. CONCLUSIONS: All subjects lost height during sitting. Both methods demonstrated a recovery of height due to hyperextension. Hyperextension could be considered as a prophylaxis against the height loss in occupational loading. PMID- 15296210 TI - Plantar foot surface temperatures with use of insoles. AB - PURPOSE: Patients with diabetes are often prescribed foot orthoses to help prevent foot ulcer formation. Orthotics are used to redistribute normal and shear stress. Shear stresses are not easily measurable and considered to be responsible for skin breakdown. Local elevation of skin temperature has been implicated as an early sign of impending ulceration especially in regions of high shear stress. The purpose of this study was to measure the effects of commonly prescribed insole materials on local changes in plantar foot temperature during normal gait. METHODS: Six commonly used foot orthosis materials were tested using the Thermo Trace infrared thermometer to measure foot temperature. Ten healthy adult volunteers without any history of diabetes or abnormal sensation participated in the study. During each trial the subject walked on a treadmill with the test material in the dominant foot's shoe, for six minutes at a speed of four miles per hour and rested for six minutes between trials. Four locations on the foot (hallux, first and fifth metatarsal heads, and heel) and the contralateral bicep temperatures were measured at 0, 1, 3, 5 minutes during the rest period. The order of material and skin location testing was randomized. RESULTS: Significant differences were found between baseline temperatures and foot temperatures for all materials. However, no differences were found between materials for any location on the foot. CONCLUSION: Previous studies have attempted to characterize materials based on laboratory and clinical testing, while other studies have attempted to characterize the effect of pressure on skin temperature. However, no study has previously attempted to characterize foot orthosis materials based on foot temperatures. This study compared foot temperatures of healthy adults based on the material tested. Although this study was unable to distinguish between materials based on foot temperatures, it was able to show a rise in foot temperature with any material used. This study demonstrates a need to a larger study on a population with diabetes. PMID- 15296211 TI - Patient and parent perspectives on treatment for adolescent idiopathic scoliosis. PMID- 15296212 TI - Autosomal dominant transmission of accessory navicular. AB - The accessory navicular bone is one of the most symptomatic bones of the foot. Although it has been reported to be present in various members of the same family, there is a lack of knowledge about its inheritance pattern. We report two large pedigrees in which accessory navicular is inherited in an autosomal dominant fashion with incomplete penetrance. PMID- 15296213 TI - Surgical treatment of solitary plasmocytoma of the spine: case series. AB - The mean survival of patients with skeletal solitary plasmocytoma is 75% at 5 year follow-up. This highly osteolytic tumor may compromise spinal stability. Radiotherapy is effective in local control of the disease, however, it is not effective in restoring spinal stability. Fracture risk and progressive vertebral collapse persist. For this reason, we must consider the need to establish the probability of progressive vertebral collapse, based on the degree of involvement of the vertebral body at the time of diagnosis. We used parameters described by Taneishi and Kaneda, as well as those of Heller and Boden to predict progressive vertebral collapse. Three cases are presented and their treatment is described. PMID- 15296214 TI - Operative treatment of cervical spondylotic myelopathy and radiculopathy. A comparison of laminectomy and laminoplasty at five year average follow-up. AB - BACKGROUND: The natural history of cervical spondylotic myelopathy is frequently one of slow, progressive neurological deterioration. The operative treatment for patients with moderate to severe involvement is decompression of the spinal cord. Laminectomy has been a traditional approach and laminoplasty has developed as an attractive alternative. The purpose of this study was to examine and compare the outcomes of these two procedures in similar groups of patients at a five year average follow-up. METHODS: A consecutive series of twenty patients who underwent open-door laminoplasty for multi-level cervical spondylotic myelopathy or radiculopathy was compared to a similar group of 22 matched patients who underwent multi-level laminectomies. Patients were similar in age, gender, number of operative levels, and length of follow-up. At the latest examination, each patient underwent a comprehensive neurological evaluation. A modification of the Nurick classification was used to assess the degree of myelopathy. Radiographs at latest follow-up were assessed for instability, and measurements of the space available-for-the-cord and Pavlov ratio were made at involved levels. RESULTS: Myelopathy, as determined by our modified Nurick scale, improved from a preoperative average of 2.44 to 1.48 in laminoplasty patients and from an average of 3.09 to 2.50 in laminectomy patients. Pain improved 57 percent and 8 percent in laminoplasty and laminectomy groups, respectively. Subjective neck stiffness was not significantly different based on the numbers available, although laminoplasty patients demonstrated some loss of range of motion on examination. The only variable that predicted the postoperative degree of myelopathy in both groups was the preoperative degree of myelopathy. CONCLUSIONS: Laminectomy and laminoplasty patients demonstrated improvements in gait, strength, sensation, pain, and degree of myelopathy. Laminoplasty was associated with fewer late complications. Based on this analysis, we believe that laminoplasty is an effective alternative to laminectomy in patients with multi-level cervical spondylotic myelopathy or radiculopathy. PMID- 15296215 TI - High-grade sarcomas mimicking traumatic intramuscular hematomas: a report of three cases. AB - We reported on three patients with high-grade soft-tissue sarcomas mimicking traumatic intramuscular hematomas. Patients had an episode of trauma to the extremity, and after initial clinical and imaging evaluations they were considered to have muscular hematomas. The lesions increased in size over time, leading to further evaluations that demonstrated the actual diagnosis. We conducted a retrospective review of the clinical findings, magnetic resonance images, and computed tomography scans to assess characteristics that will help in the differential diagnosis. We conclude that intramuscular hematomas following trauma should be approached with a high degree of clinical suspicion. MRI analysis can be used as an important diagnostic tool, but the results must be seen in the context of the clinical history. MRI is not sensitive or specific enough to rule out malignancy. The diagnosis of a high-grade sarcoma must be considered in these patients and any doubt should be resolved with a biopsy. PMID- 15296216 TI - Particulate debris osteolysis simulating malignant tumor. AB - Osteolysis induced by particulate debris from total joint implants is typically confined to bone and benign in radiographic appearance even when extensive. However, they can extend well beyond bone in which case they can simulate malignancies owing either to mass effects and pressure on adjacent tissues or owing to the radiographic appearance. We report two cases which presented as possible malignancy, and review the literature on extensive osteolysis. Recognition of this possibility may aid in interpretation of the clinical presentation and imaging studies. PMID- 15296217 TI - Use of beta-2-transferrin to diagnose CSF leakage following spinal surgery: a case report. PMID- 15296218 TI - Transitory inferior dislocation of the shoulder in a child after shoulder injury: a case report and treatment results. PMID- 15296220 TI - "Keeping up with the Joneses"--the story of Sir Robert Jones and Sir Reginald Watson-Jones. PMID- 15296219 TI - Bone and brain: a review of neural, hormonal, and musculoskeletal connections. AB - Nerves have been identified in bone. Their function has recently become the focus of intense study. Metabolic control of bone is influenced by the nervous system. Potential transmitters of this influence include glutamate, calcitonin gene related protein (CGRP), substance P, vasoactive intestinal peptide (VIP), pituitary adenylate cyclase activating polypeptide (PACAP), leptin, and catecholamines. Disorders of nerves - central or peripheral--can have substantial influence on bone health and repair. Specifically considered are the potential neural influences at work in such conditions as osteoporosis, fracture healing, Charcot osteoarthropathy, musculoskeletal pain syndromes, heterotopic ossification, skeletal growth and development, and obesity-related increased bone density. In this article, we review the current state of experimental and clinical evidence implicating the role of nervous tissue in regulating bone biology and discuss the current understanding of molecular signaling between nervous and osseus tissue in the homeostatic maintenance of the skeleton. PMID- 15296221 TI - Gas-phase production of single-walled carbon nanotubes from carbon monoxide: a review of the hipco process. AB - The latest process for producing large quantities of single-walled carbon nanotubes (SWNTs) to emerge from the Rice University, dubbed HiPco, is living up to its promise. The current production rates approach 450 mg/h (or 10 g/day), and nanotubes typically have no more than 7 mol % of iron impurities. Second generation HiPco apparatus can run continuously for 7-10 days at a time. In the HiPco process nanotubes grow in high-pressure, high-temperature flowing CO on catalytic clusters of iron. Catalyst is formed in situ by thermal decomposition of iron pentacarbonyl, which is delivered intact within a cold CO flow and then rapidly mixed with hot CO in the reaction zone. Upon heating, the Fe(CO)5 decomposes into atoms that condense into larger clusters. SWNTs nucleate and grow on these particles in the gas phase via CO disproportionation: CO + CO --> CO2 + C (SWNT), catalyzed by the Fe surface. The concentration of CO2 produced in this reaction is equal to that of carbon and can therefore serve as a useful real-time feedback parameter. It was used to study and optimize SWNT production as a function of temperature, pressure, and Fe(CO)5 concentration. The results of the parametric study are in agreement with current understanding of the nanotube formation mechanism. PMID- 15296222 TI - Laser ablation process for single-walled carbon nanotube production. AB - Different types of lasers are now routinely used to prepare single-walled carbon nanotubes. The original method developed by researchers at Rice University used a "double-pulse laser oven" process. Several researchers have used variations of the lasers to include one-laser pulse (green or infrared), different pulse widths (ns to micros as well as continuous wave), and different laser wavelengths (e.g., CO2, or free electron lasers in the near to far infrared). Some of these variations are tried with different combinations and concentrations of metal catalysts, buffer gases (e.g., helium), oven temperatures, flow conditions, and even different porosities of the graphite targets. This article is an attempt to cover all these variations and their relative merits. Possible growth mechanisms under these different conditions will also be discussed. PMID- 15296223 TI - On the growth mechanism of single-walled carbon nanotubes by catalytic carbon vapor deposition on supported metal catalysts. AB - A comprehensive kinetic study was performed to throw light on the formation mechanism of single walled carbon nanotubes (SWNTs) in chemical vapor deposition processes. SWNTs were synthesized by catalytic decomposition of methane or ethylene on supported transition metal catalysts. Kinetic curves (the amount of SWNT as a function of time) were obtained as a function of the nature and the preparation of the supported catalysts, temperature, the fluxes of the gases (the reagent hydrocarbon and the carrying gas), and the partial pressure of the hydrocarbon. The final products were characterized by transmission electron microscopy, Raman spectroscopy, chemical analysis, and thermogravimetric measurements. The fundamental factors determining the SWNT formation are discussed in detail, taking into consideration several observations from the literature as well. PMID- 15296224 TI - Nucleation and growth of single-walled nanotubes: the role of metallic catalysts. AB - We present a review of experimental and theoretical results on the nucleation and growth of single-walled nanotubes, with particular emphasis on the growth of nanotube bundles emerging from catalyst particles obtained from evaporation-based elaboration techniques. General results are first discussed. Experiments strongly suggest a root-growth process in which carbon, dissolved at high temperatures in catalytic particles, segregates at the surface at lower temperatures to form tube embryos and finally nanotubes through a nucleation and growth process. A theoretical analysis of the reasons carbon does not always form graphene sheets to wrap the particles suggests analogies with other surface or interface instabilities, in particular, with those found in epitaxial growth. In the second part, detailed experimental results for nickel-rare earth metal catalysts are presented. By using various electron microscopy techniques, it is shown that carbon and the rare earth metal co-segregate at the surface of the particle and form carbide platelets, providing nucleation sites for nanotubes growing in directions perpendicular to the surface. A simple theoretical model is then presented in which the role of the rare earth metal is just to transfer electrons from metal to carbon. The graphene sheet is shown to become unstable; pentagons and heptagons are favored, which can explain the occurrence of local curvatures and of tube embryos. Finally, a brief discussion of some recent atomistic models is given. PMID- 15296225 TI - Generation of single-walled carbon nanotubes from alcohol and generation mechanism by molecular dynamics simulations. AB - Recent advances in high-purity and high-yield catalytic chemical vapor deposition (CVD) generation of single-walled carbon nanotubes (SWNTs) from alcohol are comprehensively presented and discussed on the basis of results obtained from both experimental and numerical investigations. We have uniquely adopted alcohol as a carbon feedstock, and this has resulted in high-quality, low-temperature synthesis of SWNTs. This technique can produce SWNTs even at a very low temperature of 550 degrees C, which is about 300 degrees C lower than the conventional CVD methods in which methane or acetylene is typically used. We demonstrate the excellence of the proposed alcohol catalytic CVD method for high yield production of SWNTs when Fe-Co on USY-zeolite powder was used as a catalyst. At optimum CVD conditions, a SWNT yield of more than 40 wt % was achieved over the weight of the catalytic powder within the reaction time of 120 min. In addition to the advantages for mass production, this method is also suitable for the direct synthesis of high-quality SWNTs on Si and quartz substrates when combined with the newly developed liquid-based "dip-coat" technique to mount catalytic metals on the surface of substrates. This method allows easy and costless loading of catalytic metals without the need for any support or underlayer materials that were usually required in previous studies for the generation of a sufficient quantity of SWNTs on an Si surface. Finally, the result of molecular dynamics simulation for the SWNT growth process is presented to obtain a fundamental insight into the initial growth mechanism on the catalytic particles. PMID- 15296226 TI - Chemical models for simulating single-walled nanotube production in arc vaporization and laser ablation processes. AB - Chemical kinetic models for the nucleation and growth of clusters and single walled carbon nanotube (SWNT) growth are developed for numerical simulations of the production of SWNTs. Two models that involve evaporation and condensation of carbon and metal catalysts, a full model involving all carbon clusters up to C80, and a reduced model are discussed. The full model is based on a fullerene model, but nickel and carbon/nickel cluster reactions are added to form SWNTs from soot and fullerenes. The full model has a large number of species--so large that to incorporate them into a flow field computation for simulating laser ablation and arc processes requires that they be simplified. The model is reduced by defining large clusters that represent many various sized clusters. Comparisons are given between these models for cases that may be applicable to arc and laser ablation production. Solutions to the system of chemical rate equations of these models for a ramped temperature profile show that production of various species, including SWNTs, agree to within about 50% for a fast ramp, and within 10% for a slower temperature decay time. PMID- 15296227 TI - Arc process parameters for single-walled carbon nanotube growth and production: experiments and modeling. AB - Collarets rich in single-walled carbon nanotubes (SWCNTs) have been grown using a direct current arc method. Arc process parameters such as current, pressure, and anode to cathode distance were varied experimentally and by modeling to provide an optimal working window. The best collaret yields were obtained when helium was used as a buffer gas. Mixing helium with argon in the buffer permits controlling nanotube diameters. In addition to an experimental study, a modeling approach was developed assuming local thermal equilibrium and homogenous and heterogeneous neutral chemistry. The gas-phase chemical model involves 81 neutral carbon species (C1, C2, . . ., C79, C60F, C70F) and 554 reactions with rates taken from data of Krestinin and Moravsky. Axial profiles of temperature, C atom, C2 radical, and fullerene distributions in the reactor are predicted as a function of process parameters. Carbon nanotube growth is considered by a set of surface reactions simulating open nanotube growth. Because nanotube surface chemistry is controlled by the local terminated bond and not by the bulk nanotube bond, a mechanistic approach based on the formal resemblance between the bonding and the structure of open nanotube and other carbon surfaces is proposed to explain nanotube growth. Predicted growth rates are in the range of 100 to 1000 microm/min. PMID- 15296228 TI - Endofullerenes with metal atoms inside as precursors of nuclei of single-walled carbon nanotubes. AB - Thermodynamic estimations have been made which show that amorphous carbon cannot be the source of single-walled carbon nanotube growth during carbon/metal vapor condensation. Thus the source of carbon material for nanotube growth seems to be small carbon clusters into gas phase or clusters in the adsorbed state on the nanotube surface. Kinetic analysis of carbon/metal vapor condensation in the arc process was performed and showed that metal vapor becomes supersaturated at temperatures that are close to the temperature of the highest rate of fullerene shell formation. A new model of the nucleation of single-walled carbon nanotubes is proposed. In the model an endometallofullerene serves as the precursor of a nanotube nucleus, and the nucleus itself forms as an adduct arising after metal atoms attach to the endofullerene shell. The relative efficiencies of La/Ni, Gd/Ni, Ce/Ni, and Pr/Ni catalysts, in comparison with Y/Ni catalyst, were measured, and their high efficiency in buckytube formation was demonstrated. This fact was explained by the double action of a metal catalyst in buckytube nucleus formation. First, metal takes part in the formation of endometallofullerenes. Second, the metal atoms attach to the endofullerene shell to transform an endofullerene into a nanotube nucleus. The high performance of bimetallic catalysts lies in adjusting the values of two metal concentrations separately for each of these purposes. In accordance with observations, the proposed model predicts increasing size of endofullerenes with increasing concentration of the metal that controls endofullerene formation. PMID- 15296229 TI - Decomposition of carbon-containing compounds on solid catalysts for single-walled nanotube production. AB - Different catalyst formulations and reaction conditions have been used to test the validity of a hypothesis that tries to elucidate the mechanism of single walled nanotube (SWNT) formation by CO disproportionation over a highly selective cobalt-molybdenum catalyst. This model proposed an intrinsic dependence between the selectivity of the catalysts toward SWNT and the stabilization of Co species in a nonmetallic state, which in turn results from an interaction with Mo. The series of tests performed to examine this model include the doping of this highly selective catalyst with sodium, the substitution of molybdenum by tungsten in the original catalyst formulation, the variation on reaction temperature, and the introduction of hydrogen in the gas feedstock. All these modifications were carried out to modify the growth conditions in which the SWNTs are formed. The results are consistent with the hypothesis proposed by the authors. PMID- 15296230 TI - Self-organization of carbide superlattice and nucleation of carbon nanotubes. AB - In metal-carbon systems with known stable compounds, carbide nanocrystals self organize epitaxially on metal surfaces to form two-dimensional arrays during carbon deposition. The process is energetically driven by the competition between the strain and surface energies, and it appears to play an important role in the nucleation of single-walled carbon nanotubes. Interplay between energetics and kinetics controls carbon precipitation from the superlattice, such that the length scale of the carbide and superlattice appears to control the size and morphology of the precipitates. Furthermore, carbon precipitates appear to be "seedlings" of carbon nanotubes grown on top of the carbide nanocrystals. These findings reveal that the nucleation of carbon nanotubes is a nonequilibrium process and that a stable carbide superlattice can be used as an ordered template of carbon saturated "roots" for nucleating nanotube bundles with controlled diameter, spacing, and perhaps chirality. PMID- 15296231 TI - Multiscale simulations of carbon nanotube nucleation and growth: electronic structure calculations. AB - Several first-principles surface and bulk electronic structure calculations relating to the nucleation and growth of single-wall carbon nanotubes are described. Density-functional theory in various forms is used throughout. In the surface-related calculations, a 38-atom Ni cluster and several low-index Ni surfaces are investigated using pseudopotentials and plane-wave expansions. The energetic ordering of the sites for C atom adsorption is found to be the same, with the Ni(100) facet favored. The bulk diffusion coefficient of C in Ni as a function of cluster size and temperature is calculated from various molecular dynamics approaches. In another group of bulk-related calculations, Gaussian orbital basis sets are used to study a cluster or "flake" containing 14 C atoms. The flake is a segment of three hexagons from an "unrolled" carbon nanotube, with an armchair termination. The binding energies of C, Ni, Co, Fe, Cu, and Au atoms to it were calculated in an effort to gain insight into the mechanism for the high catalytic activity of Ni, Co, and Fe and the lack of it in Cu and Au. The binding energies of Cu and Au are about 1 eV less than those of the three catalytic elements. Similar methods are used to study the initial stages of nanotube growth within the context of classical nucleation theory. Finally, issues relating to the establishment of a fundamental catalytic mechanism are addressed. PMID- 15296232 TI - Voltammetric determination of tryptophan at a single-wall carbon nanotubes modified electrode. AB - A single-wall carbon nanotubes (SWNT)-film coated glassy carbon electrode (GCE) was described for the determination of tryptophan. In pH 2.5 Na2HPO4-citric acid buffer, tryptophan yields a well-defined and very sensitive oxidation peak at about 1.08 V at the SWNT-film coated GCE. The oxidation peak current increases greatly and the peak potential shifts toward more negative direction at the SWNT modified GCE in contrast to that at the bare GCE. Under optimized conditions, the oxidation peak current is proportional to the concentration of tryptophan over the range from 4 x 10(-8) to 1 x 10(-5) mol/L. The detection limit is 1 x 10(-8) mol/L at 3 min of accumulation. Using the proposed method, tryptophan in the human's blood serum samples was determined. PMID- 15296233 TI - Fe-sapphire and C-Fe-sapphire interactions and their effect on the growth of single-walled carbon nanotubes by chemical vapor deposition. AB - We previously reported that the quantity of single-walled carbon nanotubes grown on Fe-coated sapphire by chemical vapor deposition depended on the crystallographic faces of sapphires. In this report, we show that the interaction of Fe, sapphire, and carbon depended on the sapphire faces. We deduce that the quantity of Fe available to catalyze the growth of single-walled carbon nanotubes was suppressed by the formation of Fe-Al alloys and whether the Fe-Al alloys were formed on Fe-coated sapphire or not depended on the sapphire-surface structure. PMID- 15296234 TI - Single-walled carbon nanotube diameter. AB - Two different single-walled carbon nanotube (SWNT) growth modes (cap growth mode and circumference growth mode) are shown to exist. General SWNT diameter windows are derivable from catalyst particle size considerations. In addition, an almost complete picture of nanotube diameter dependencies for the cap growth mode is drawn from experiment. The nanotube diameter always scales linear with temperature, but the degree of dependence varies with the catalyst element. The nanotube diameter scales logarithmically with the gas pressure and catalyst composition. Very few or exactly one atom of a catalyst additive is sufficient to induce SWNT diameter changes. The experimental data allow the conclusion that the observed nanotube diameter is based on materials properties of sp2-bonded carbon/graphene sheets, on individual properties of the catalyst elements, and on additional kinetic components from temperature and pressure changes. Indications are found for a specific and maybe decisive role of adsorbate atoms at the surface of a catalyst particle on the nanotube diameter and therefore on the process of nanotube nucleation and growth. PMID- 15296235 TI - Computational fluid dynamics simulation of laser-ablated carbon plume propagation in varying background gases for single-walled nanotube synthesis. AB - A computational fluid dynamics study was conducted to model the plume resulting from the laser ablation of a carbon target in a laser ablation oven for the production of carbon single-walled nanotubes (SWNTs). The goal is to gain understanding into the fluid dynamics and thermodynamics of the plume to ultimately improve SWNT production techniques. The simulations were carried out with a 12-species, 14-reaction chemistry model that included carbon species from C to C6. Metal catalysts in the carbon target were ignored for these simulations. Simulation times ranged from immediate ablation onset to 8 ms past the initial onset of ablation. A secondary goal of the study was to compare computational results with experimental results for three different background gases in the laser ablation oven-argon, helium, and nitrogen. Computational results indicated that lighter carbon species were more quickly diffused into the background gas for helium and nitrogen, resulting in lower localized mass fractions of carbon nanotube "feedstock." The expectation is that this effect will reduce the production of carbon nanotubes, which has been confirmed by experimental evidence. From this investigation, a possible "indicator species" for the production of SWNT appears to be C5. PMID- 15296236 TI - In-situ optical analysis of the gas phase during the formation of carbon nanotubes. AB - A reactor has been developed at ONERA to investigate the gas phase during carbon nanotube formation by laser-induced fluorescence (LIF), Laser-induced incandescence (LII), coherent anti-Stokes Raman Scattering (CARS), and emission spectroscopy. Continuous vaporization is achieved with a continuous wave CO2 laser. Optimized conditions are used for single-walled nanotube growth, that is, a graphite target doped with 2 atom % Ni and 2 atom % Co, helium as buffer gas at a flow rate of 50 ml/s, and a pressure of 300 hPa. Temperature profiles are measured by CARS on H2, and soot images are obtained by LII in the hot carbonaceous flow. LIF and spontaneous emission of the C2 radical and Ni and Co atoms are presented. Spectral investigations are conducted at 3100 and 3200 K to have an optimized pair of excitation/detection wavelengths. Spatial investigations of the relative concentrations in the hot carbonaceous flow are performed up to 3500 K. The concentrations are measured as a function of target temperature. Two regimes of vaporization are observed. Vaporization is slow up to 3350 K and becomes much faster above this temperature. The fast regime in the 3350-3500 K range corresponds to the observed spatial extent of the metal vapors region. At 3500 K, the C2 profiles obtained with and without catalysts are very different as a result of carbon coalescence as well as carbon dissolution into the metal nanoparticles when these are present in the gas phase. The shape of the C2 profile can be related to nanotube formation and growth at a target temperature of 3500 K. PMID- 15296237 TI - Multiscale simulations of carbon nanotube nucleation and growth: mesoscopic continuum calculations. AB - As part of a focused computational effort on the multiscale simulations of carbon nanotube nucleation and growth, we have developed computer programs for coupled heat and mass flow in one and two dimensions. In the tip-growth mode, the sample is divided into three main regions, each of which can be further subdivided as required. In region 1, carbon is supplied to the catalytic particle from an ambient gas of carbon-containing compounds. The chemistry and thermodynamics of the decomposition of these compounds can be included in region 1, but the capability has not yet been implemented. The carbon diffuses through the catalytic particle in region 2 under concentration and temperature gradients and with a diffusion coefficient that can depend on both concentration and temperature. Region 3 consists of the interfacial region between the catalytic particle and the growing nanotube. Results to date demonstrate the key roles played by the size and shape of the catalytic particle in conjunction with the concentration and temperature gradients at the gas/solid interface and in region 2. Results also suggest how the growth of a single wall may interfere with, but not necessarily prevent, the growth of additional walls in a multi-walled nanotube. Again, the carbon concentration profile in the catalytic particle at the different growth sites is a key factor. PMID- 15296238 TI - Forensic science in the trial of Sally Clark. PMID- 15296239 TI - Inference with legal evidence: common sense is necessary, but not sufficient. AB - Recent cases have highlighted the issue of faulty probabilistic reasoning by expert witnesses in courts of law. While concern about potential miscarriages of justice is clearly well-placed, the consequences of such faulty reasoning do not seem to be fully appreciated. These are often counter-intuitive, as we show with two examples: the Interrogator's Fallacy and the Prosecutor's Fallacy. Both demonstrate the danger of relying solely on 'common sense' when drawing inferences from legal evidence. PMID- 15296241 TI - Changes in blood viscosity with heavy and light exercise. AB - To clarify the relationship of the intensity of acute exercise to sudden cardiac death, we examined the effects of short-term heavy and light exercise on whole blood viscosity. Nine healthy sedentary male volunteers performed ten minutes of heavy (more than 95% of maximum oxygen consumption) or light (60% to 65% of maximum oxygen consumption) exercise. Blood samples were obtained before, immediately after, and one hour after exercise. The whole blood viscosity was immediately examined with an oscillation-type viscometer and was found to increase significantly after exercise and subsequently return to baseline levels within one hour after exercise. The whole blood viscosity increased by a similar degree after heavy or light exercise. Therefore, our results suggest that there is a similar risk of sudden cardiac death, due to increased whole blood viscosity, after short-term heavy or light exercise. PMID- 15296240 TI - An evaluation of psychiatric support to magistrates' courts. AB - A Court Protocol was introduced to Hampshire Magistrates' Courts on 1 January 2003 which enabled magistrates to seek information on mentally disordered defendants from local mental health workers in place of requesting a full psychiatric report. The scheme was evaluated by examining 48 case files of mentally-disordered defendants who were referred between 1 January 2003 and 31 December 2003. It was found that its use had significantly reduced the time taken from first appearance to disposal and significantly reduced inappropriate referrals to consultant psychiatrists. In many cases, defendants were spared repeated bail adjournments and lengthy remands in prison. PMID- 15296242 TI - Victims of falls from a height surviving to hospital admission in two Singapore hospitals. AB - Falls from a height are a common problem in Singapore. However, there has been no analysis of the injury patterns and outcomes of patients who survived initial resuscitation after falls from a height in Singapore. One hundred and thirty-nine patients admitted over a two-and-a-half year period after falls of greater than one storey were studied. The higher the fall, the greater the likelihood of hypotension and neurological injury. The length of stay in hospital, the number of operative procedures and mortality also correlated with the height of the fall. The overall mortality rate was 11% but rose to 47% among patients who were hypotensive on admission. About half of the deaths were due to head injury with haemorrhage being the second most common cause. The majority of patients who fell from five or more storeys did so intentionally, and had more severe torso and extremity injuries compared with those who fell accidentally. These findings suggest that efforts in injury prevention and aggressive evaluation and treatment of fall victims arriving alive at the hospital continue to be important. PMID- 15296243 TI - Childlessness in elderly suicide: an analysis of coroner's inquests of 200 cases of elderly suicide in Cheshire 1989-2001. AB - Child bearing has been reported to be protective for women in terms of suicide risk. However, it is not clear whether the protective effect of having children diminishes in old age. A literature search has not found studies which examined childlessness as a possible risk factor in elderly suicide. In this study we attempt to explore whether childlessness has any significant association with elderly suicide and whether this differs between women and men. Data was extracted from the records of coroners' inquests into all unexpected deaths of persons aged 60 and over, in Cheshire, over a period of 13 years from 1989-2001. The study found no significant gender difference in childlessness in elderly suicide victims (P>.05). Significantly fewer widowed men who committed suicide were childless (OR 0.1, 95% CI 0.02-0.3 P<.001). A history of deliberate self harm (DSH) and being previously known to services were found to be significantly lower in childless female victims compared to elderly mothers (OR 0.4, 95% CI 0.1 0.9 P<.05). Leaving suicide notes and the method of suicide did not appear to be significantly influenced by a childless status in either of the sexes (P>.05). The protective effect of having children appears to diminish in old age. Childless females appear to be at a higher risk of succeeding in their first suicide attempt and more importantly, can easily escape the attention of services. PMID- 15296244 TI - The effect of clozapine on the social behaviour schedule in patients attending a forensic psychiatry day hospital. AB - This study examined the difference in the functional capacity of patients receiving clozapine, compared with those prescribed other antipsychotic medication, who attend the forensic psychiatry day hospital at Murray Royal Hospital, Perth. (This facility caters for 34 patients. It provides a number of advantages such as access to therapies and supervision of mental state and medication at this critical time in the care pathway.) This study was cross sectional in nature and examined the patients' demographic details, diagnosis and medication. A functional assessment was carried out using the social behaviour schedule (SBS) for each patient. We discovered that there was a marked difference in the hostility scores between clozapine patients and non-clozapine patients. Only 10.5% of clozapine patients had a severe score (more than two) on the SBS whereas 61.5% of those not receiving clozapine had a severe score. The non clozapine group had more socially unacceptable habits and were more destructive than patients receiving clozapine, as scored by the SBS. The slowness and under activity items were more severe in the clozapine patients. The non-clozapine group scored higher in the category of other behaviours that might impede progress, particularly drug-taking. PMID- 15296245 TI - Life after clozapine. AB - We consider the reasons underlying clozapine withdrawal in a group of hospitalised schizophrenic inpatients and review clinical outcome following clozapine withdrawal. A retrospective case note study was undertaken of all patients (n=41) registered at the State Hospital, Carstairs, with the Clozaril Patient Monitoring Service between 1990 and 2001 whose clozapine therapy had either been withdrawn or never initiated following registration. Twelve patients refused to commence clozapine following registration: twenty-nine patients were subsequently studied. Thirteen patients stopped clozapine owing to non compliance, whereas seven had clozapine withdrawn due to a clinically inadequate response. Eleven patients were withdrawn primarily because of an unacceptable side-effect burden, although only four had a 'red alert'. In twenty patients, abrupt clozapine withdrawal resulted in a marked, immediate deterioration in their mental state. Twenty-three patients remained at the State Hospital at a mean follow-up period of 75 months. The study highlights the relevance of psychoeducation for clozapine patients in the maintenance phase of treatment, the importance of minimising the side-effect burden by clozapine monotherapy, the effectiveness of clozapine serum level monitoring and the evidence base for adjuvant treatment. PMID- 15296246 TI - A study of p53 expression in thermal burns of human skin for determination of wound age. AB - Early expression of p53 protein in thermal burns of guinea pig skin has been reported. This study sought to determine if expression occurred in thermal burns of human skin and if immunohistochemical demonstration of p53 protein could be utilised to distinguish ante-mortem from post-mortem injuries as well as indicating the age of a lesion in the living subject. Biopsy samples were obtained from live patients and post-mortem examinations. Immunohistochemistry was used to demonstrate the presence of p53 protein. Staining was assessed by field counting of epithelial cell nuclei. In live subjects there was a tendency for early (six hour to five day) expression, with peak levels occurring around one to two days. Late samples (13 to 23 days) demonstrated minimal or no expression. In contrast, burn wounds from post-mortem examination demonstrated greater staining for p53 protein in the late (28 to 77 day) samples than in the early ones. It appears that expression of p53 protein may assist in the ageing of ante-mortem, but not post-mortem, burn wounds. This implies that results obtained from live subjects may not be applicable to post-mortem material and that any putative method for determining the age of a wound should be tested in both situations. PMID- 15296247 TI - Section 5(2) of the Mental Health Act: local and national use over a twelve-year period. AB - The pattern of use of Section 5(2) of the Mental Health Act 1983 was examined over a 12-year period at one psychiatric hospital. Although there was a national trend for Section 5(2) to be used more frequently, the local pattern was of a fluctuating number of patients being placed on this Section, but without there being any upward trend towards its increased use. As has been seen in other studies, Section 5(2), however, tended to be used for a younger group of patient who had often only been admitted within the previous 24 hours. PMID- 15296248 TI - Psychotropic prescriptions and elderly suicide rates. AB - Suicide rates in the elderly have declined in recent years in several countries including England and Wales. There have also been major changes in prescribing patterns of psychotropics in many countries during recent years. This study examined the association between a trend of decline in suicide rates in the elderly in England and Wales and changes in the prescribing of psychotropics for the period 1985 to 1996. Data on suicide rates was ascertained from the annual mortality statistics for the years 1985 to 1996. Data on the prescriptions of psychotropics was ascertained from the Statistic Division (SD1E) at the Department of Health. Spearman's rank correlation coefficient (rho) was used to examine the relationship between the prescription of psychotropics and suicide rates. The main findings of this study were: (i) a highly significant negative correlation between the prescription of tricyclic antidepressants or selective serotonin reuptake inhibitors (SSRIs) and elderly suicide rates; (ii) a significant positive correlation between the prescription of monoamine oxidase inhibitors (MAOIs) and elderly suicide rates; (iii) a highly significant negative correlation between the prescription of antimanics and antipsychotics and elderly suicide rates; (iv) a highly significant positive correlation between the prescription of hypnotics, anxiolytics and barbiturates and elderly suicide rates; and, (v) a significant negative correlation between the prescription of analgesics and elderly suicide rates. A trend of decline in suicide rates in the elderly in England and Wales for 1985 to 1996 and changes in the prescribing of psychotropics were significantly associated. The results of this study suggest that prescription patterns of psychotropics may be an important factor contributing to the recent decline in suicide rates in the elderly in England and Wales but this effect should be viewed in the context of other health and social factors. PMID- 15296249 TI - Organ transplantation programme: an overview of the present situation. AB - An ever-increasing demand for organs for transplantation has failed to keep pace with their availability. Social, religious and legal restrictions on the one hand and technological developments on the other have further worsened the existing shortage of organs. A correct definition of the 'moment of death', a more humane approach in breaking the news of death to relatives and de-linking the request for donation from the announcement of death may all go a long way in enhancing organ donation and procurement for needy patients. In India, the Transplantation of Human Organs Act was passed in 1994 to regulate the removal of organs from the living as well as the dead. However, it has certain fallacies. This paper reviews the various criteria in use to define the 'moment of death' around the world and with reference both to the Transplantation of Human Organs Act 1994 and the urgent need for organ retrieval in the present day paucity of donors. PMID- 15296250 TI - A fatal case of hepatic failure possibly induced by nitrosofenfluramine: a case report. AB - A 42-year-old female developed fulminant hepatic failure after having ingested an undetermined quantity of a herbal product over a period of approximately four months prior to the onset of her illness. Clinically, the cause of liver failure was assessed to be drug-induced and she eventually underwent total hepatectomy, with porto-caval shunting, in anticipation of a living-unrelated liver transplant. Unfortunately, her condition deteriorated and she died less than 48 hours post-operatively, approximately three weeks post-admission. An autopsy showed that the subject was deeply jaundiced and severely obese (BMI: 47.1 kg m( 2)), with evidence of diffuse haemorrhage, including the presence of 1.35 l of blood in the peritoneal cavity. The liver had been removed and was later recovered as a formalin-fixed specimen which was markedly contracted, comprising multiple micronodules interspersed with extensive areas of dense fibrotic tissue. Histologically, there was massive necrosis of the hepatic parenchyma, such that the residual hepatocytes were disposed as nodules displaying variable cellular regeneration and ballooning degeneration, attended by florid ductal proliferation and mixed inflammatory infiltrates. Infective, autoimmune, metabolic, vascular, neoplastic and most other natural causes of massive hepatocellular necrosis were effectively excluded. Analysis of the post-mortem blood samples yielded fluconazole, metronidazole, frusemide, lignocaine and tramadol, (therapeutic agents administered to the patient during her last illness). Subsequent analysis of the residual capsules revealed that they were adulterated by fenfluramine, N nitrosofenfluramine (1.3-1.6 mg per capsule), nicotinamide (13.3-15.6 mg per capsule) and thyroid extract. None of the herbal ingredients is currently known to be hepatotoxic and much the same applies to fenfluramine, nicotinamide (except when taken in mega-doses) and thyroid extract. However, as nitrosamines are known to be variably hepatotoxic, it would be reasonable to surmise that, in the absence of a more plausible cause of liver damage, N-nitrosofenfluramine was the likely cause of massive hepatocellular necrosis in this instance. PMID- 15296251 TI - An autopsy case of fatal anaphylactic shock following fluorescein angiography: a case report. AB - We present a rare autopsy case of fatal anaphylactic shock following fluorescein angiography. A 71-year-old Japanese woman undergoing retinal angiography to evaluate diabetic retinopathy died immediately after an injection of sodium fluorescein. Forensic autopsy and post-mortem biochemical analyses revealed an elevated serum level of tryptase which, in the absence of morphologic changes suggesting injury or disease, confirmed the diagnosis of fatal anaphylactic shock. Although serious adverse effects are rare after fluorescein angiography, patients should be observed, with appropriate resuscitation equipment available, for several hours after the administration of fluorescein. PMID- 15296252 TI - A fatal case of poisoning by lidocaine overdosage--analysis of lidocaine in formalin-fixed tissues: a case report. AB - The death of a 76-year-old man with heart disease as a result of the injection of an excessive dose of lidocaine is presented. The patient was given 5 ml of 10% lidocaine hydrochloride (500 mg) intravenously instead of 2.5 ml of 2% lidocaine hydrochloride (50mg) in order to treat repeated paroxysmal ventricular arrhythmia. Immediately following the injection the patient had tonic clonic seizures and complete cardiopulmonary arrest followed. Although resuscitation attempts once successfully restarted his pulse and spontaneous respiration, the patient died on the eighth day after the injection. Toxicological examinations were carried out on the tissues obtained at the time of autopsy and which had been fixed in formalin solution for 40 days, and lidocaine was detected in each tissue examined. The concentrations were (ng/g or ml): parietal lobe, 308.0; occipital lobe, 208.7; temporal lobe, 318.0; frontal lobe, 223.2; cerebellum 200.9; pons 285.7; liver, 109.5; kidney 52.2; skeletal muscle 127.0; and formalin solution 8.4. In an experiment on rats we determined the concentration changes of lidocaine in formalin fixed tissues. The concentrations of lidocaine in these tissues significantly decreased to 1/3-1/4 from the original. This data shows that the cause of death was poisoning by lidocaine overdose. PMID- 15296253 TI - Milk aspiration in an infant during supine bottle feeding: a case report. AB - A two-month-old infant died during bottle feeding in the supine position in the caregiver's absence. Scene investigations and autopsy examinations, including alpha-lactalbumin immunohistochemistry of the lungs, revealed the cause of death to be asphyxia due to aspiration of milk pooled in the naso-oral cavity, as a result of unsupervised supine feeding. This case emphasizes the need for an investigation into feeding positions and immunohistochemical examinations for the diagnosis of asphyxia due to milk aspiration. PMID- 15296254 TI - Radon in Ireland and beyond. PMID- 15296255 TI - High radon concentrations in a house near Castleisland, County Kerry (Ireland)- identification, remediation and post-remediation. AB - In July 2003, a passive radon measurement carried out over a 3-month period in a house near Castleisland in County Kerry (South-West of Ireland) identified a seasonally adjusted annual average concentration of approximately 49 000 Bq m( 3). This is the highest radon concentration ever recorded in a house in Ireland. It is almost 250 times higher than the national reference level of 200 Bq m(-3) for homes and it gives rise to an estimated annual radiation dose of approximately 1.2 Sv to the occupants. This paper describes the identification of the 'Castleisland house' and gives information on the local geology, the levels of natural background radiation in the area and the follow-up actions taken to remediate the house as well as the efforts made to heighten awareness in the locality of the hazards from radon. PMID- 15296256 TI - Current status of programmes to measure and reduce radon exposure in Irish workplaces. AB - National legislation, which implements European Council Directive 96/29/EURATOM in Ireland, sets a reference level of 400 Bq m(-3) averaged over any 3 month period for radon exposure in the workplace and also empowers the Radiological Protection Institute of Ireland to direct employers to have radon measurements carried out. This legislation came into effect in May 2000. Radon measurements have already been completed in show caves and other underground workplaces. Between 1998 and 2001, over 33 800 individual radon measurements were carried out in all ground floor offices and classrooms in 3444 schools nationwide as part of a programme undertaken jointly with the Department of Education and Science. Where the average indoor radon concentration in one or more rooms exceeded 200 Bq m(-3), remedial measures were implemented. For concentrations up to 400 Bq m(-3) this involved increased ventilation while for higher concentrations an active sump was normally installed. The results of the survey, as well as the effectiveness of the different remedial strategies, are discussed. In the case of other above ground workplaces, different approaches have been adopted. As a first step, workplaces in two known high radon areas were directed to have radon measurements carried out. This programme had limited success because of problems in obtaining accurate workplace databases and a general lack of awareness on the part of employers of the issues involved. From a sample of 2610 employers directed to measure radon, only 408 actually completed measurements and 37 workplaces were identified as having average 3 month average radon concentrations above 400 Bq m(-3). A total of 1356 employers ignored all correspondence, some of which was sent by registered post and signed for on receipt. Current initiatives are focused on the provision of information and include newspaper advertising as well as publications aimed specifically at both employer and employee representative groups. The ability to provide accurate information that encourages both measurement and remediation is seen as central to an effective radon workplace programme. PMID- 15296257 TI - Uncertainty in estimating probability of causation in a cross-sectional study: joint effects of radiation and hepatitis-C virus on chronic liver disease. AB - Exposure to other risk factors is an important consideration in assessing the role played by radiation in producing disease. A cross-sectional study of atomic bomb survivors suggested an interaction between whole-body radiation exposure and chronic hepatitis-C viral (HCV) infection in the etiology of chronic liver disease (chronic hepatitis and cirrhosis), but did not allow determination of the joint-effect mechanism. Different estimates of probability of causation (POC) conditional on HCV status resulted from additive and multiplicative models. We therefore estimated the risk for radiation conditional on HCV status using a more general, mixture model that does not require choosing between additivity or multiplicativity, or deciding whether there is interaction, in the face of the large uncertainty. The results support the conclusion that POC increases with radiation dose in persons without HCV infection, but are inconclusive regarding individuals with HCV infection, the lower confidence bound on estimated POC for radiation with HCV infection being zero over the entire dose range. Although the mixture model may not reflect the true joint-effect mechanism, it avoids restrictive model assumptions that cannot be validated using the available data yet have a profound influence on estimated POC. These considerations apply more generally, given that the additive and multiplicative models are often used in POC related work. We therefore consider that an empirical approach may be preferable to assuming a specific mechanistic model for estimating POC in epidemiological studies where the joint-effect mechanism is in doubt. PMID- 15296258 TI - An integrative radiation protection control system based on a CAN bus for the HT 7U tokamak fusion device. AB - A radiation protection control system has been designed, based on distributed computers and consideration of the features of the radiation source of the HT-7U fusion experimental device, for protecting the workers and the public against neutron and photon radiation, and especially for ensuring that workers cannot unexpectedly enter an area of high radiation level in any case. A multisubsystem (irradiation monitoring subsystem, access control subsystem, safety interlock subsystem and other related facilities) integration concept is proposed for the design. This system has been implemented on the basis of the up-to-date industrial field bus CAN, featuring simplicity and flexibility of installation and maintenance, capability for real-time long distance communication and multi master protocol. PMID- 15296259 TI - Performance characteristics of LiF thermoluminescent dosemeters employed in the National Personnel Radiation Dose Services in Tanzania. AB - Thermoluminescent dosemeters (TLDs) have been used to estimate the personal dose equivalent for external occupationally exposed workers in Tanzania. The reliability and precision of dose measurements and the accuracy of dose evaluation are important factors for the improvement and achievement of individual monitoring objectives. In this piece of work we describe a study of the major characteristics of TLDs (linearity of response, photon energy response, batch homogeneity and uniformity, calibration of TLDs and TLD systems in terms of operational quantities, fading, etc) in order to initiate a routine performance testing and quality assurance programme to be undertaken by central personal dosimetry services. The energy response of the dosemeters relative to that measured using 137Cs gamma rays was found to vary between 1.0 and 1.3 over the range of 33-1250 keV (60Co). The linearity of the response for the TL dose dependence in the dose range of 0.1-30 mGy for 48 keV and 137Cs (662 keV) radiation protection qualities varied from 15% to 7% respectively, for both energies. The minimum detectable dose, the batch homogeneity and uniformity, and the batch reproducibility were found to be 0.1 mGy, 16%, and 9% respectively while the fading characteristic of doped LiF after a received dose of 3 mGy was found to be 6.3% over a one-month period. These results are discussed in order to demonstrate the degree of accuracy achieved and the need for its improvement where necessary. PMID- 15296260 TI - Practical procedures for a radon etched track dosimetry service. AB - Etched track detectors are widely used for the detection of radon and its decay products. They have many desirable attributes: they are small, cheap, simple, non toxic and non-hazardous. Etched track detectors provide adequate accuracy for most radiological protection purposes provided stringent quality assurance is maintained. The UK validation scheme provides an important component of QA but continuous monitoring of conditions and results is also needed. If these conditions are observed, these detectors provide an entirely adequate tool for large-scale use in assessing levels of radon in houses. Accurate estimates of long-term average radon levels require a measurement over several months because of the short-term fluctuations in radon concentrations. PMID- 15296261 TI - Effect of reader and oven annealing on the glow curve structure and fading of a LiF:Mg,Cu,P TL dosimeter. AB - The effect of pre-irradiation annealing in an oven at 240 degrees C/10 min (oven anneal) and in a TLD reader at 6 degrees C s(-1), 10 s dwell time at a maximum temperature 240 degrees C (reader anneal), on the glow curve structure of a home made sample of LiF:Mg,Cu,P powder has been investigated. It is shown that on oven annealing the glow peak areas of peaks 2 and 3 increase and that of peak 4 decreases, while on reader annealing the area of peak 4 increases and those of peaks 2 and 3 effectively decrease. Values of fading during a post-irradiation storage of 6 months were measured to be 26% and 12% for samples undergoing oven and reader annealing respectively. To reduce the effect of fading several pre heats at temperatures of 100, 110, 120 and 130 degrees C for 10 min were investigated and 120 degrees C/10 min was chosen as the optimal condition for pre heat. It is also observed that when imposing pre-heat thermal treatment on this dosimeter or storing the irradiated dosimeter the height of peak 4 effectively increases while the integral of TL signals remains constant. PMID- 15296262 TI - Solid radioactive waste: a confused mixture of responsibilities and discussions. PMID- 15296263 TI - Exposures to non-ionising radiation. PMID- 15296264 TI - SRP meeting on Protecting Biota from Ionising Radiation. Edinburgh, 23 January 2004. PMID- 15296265 TI - DisTec 2004. Berlin, 26-28 April 2004. PMID- 15296266 TI - [Meningococcal meningitis: individual and collective approach]. PMID- 15296267 TI - [Macrophage activation syndrome]. AB - Macrophage activation syndrome is due to macrophage stimulation secondary to excessive cytokine secretion. Infectious (mainly viral) and neoplastic (lymphomas primarily) diseases are the mainly triggering circumstances of this syndrome associated with immunodepression. Signs are fever, hepatosplenomegaly, lymphadenopathy, cytopenia, deranged liver function increases level of lactate dehydrogenase, triglyceridemia and serum ferritin. Diagnosis is established by the presence of cellular phagocytosis by macrophage on medullary or hepatic biopsy. It is a potentially life threatening condition. Treatment of the triggering circumstance associated with corticosteroids, chemotherapy (etoposide) or intravenous immunoglobulins, according to etiology, are necessary. PMID- 15296268 TI - [Laparoscopic surgery]. PMID- 15296269 TI - [Bacterial meningitis of adults and children. Reduce the incidence, improve the prognosis]. PMID- 15296270 TI - [Epidemiology of bacterial meningitis in France in 2002]. AB - In France, two sources of data, the mandatory notification and the hospital laboratory network EPIBAC, allow the health authorities to monitor the incidence of bacterial meningitis. In 2002 around 1,500 cases were estimated in France. The more common microorganisms were: Streptococcus pneumoniae 49% (incidence 1.19/100,000), Neisseria meningitidis 33% (incidence 0.83) and Streptococcus agalactiae (Streptococcus B) 11% (incidence 0.27). Listeria monocytogenes accounted for 4% of the cases (incidence 0.11) and Haemophilus influenzae accounted for 3% (incidence 0.08). Trends in incidence showed an increase of meningococci meningitis since 1996, a steady decrease of H. influenzae and Listeria meningitis since 1992, and a quasi stable number of pneumococci meningitis, streptococci B meningitis and tuberculosis meningitis in the last 10 years. The impact of preventive measures on the incidence of Listeria meningitis and H. influenzae b meningitis has been clearly demonstrated through the dramatic decrease of meningitis due to these microorganisms. PMID- 15296271 TI - [Diagnosis and management of bacterial meningitis in the adult]. AB - Bacterial meningitis is a medical emergency requiring early diagnosis and therapy in order to reduce mortality and morbidity. Although fever is the most sensitive sign, occuring in a majority of patients, it may be absent, especially in oldest patients. Most patients have alterations in mental status but coma is more frequent in meningitis caused by S. pneumoniae. Focal neurologic signs are present in about 25% of the cases and are again much more frequent in the setting of pneumococcal meningitis. In adults with suspected meningitis, mass effect on CT-scan is unfrequent and as a consequence, the risk of lumbar punction is negligible. Very early administration of antibiotics, even before hospital admission in case of suspected meningococcal infection may result in a decreased abilty to identify the etiologic agent by cultures. The use of new techniques for detection of bacterial antigens and the development of rapid PCR assays may be particularly helpful in patients who had received antibiotics before lumbar puncture. PMID- 15296272 TI - [Meningococcal purpura fulminans in children]. AB - In France, the incidence of meningococcal infections is increasing and the most severe presentation, called purpura fulminans, has still a death rate of 20-25%. Diagnosis of invasive meningococcal infection must be evoked in any child presenting with febrile purpura (vasculitic rash not disappearing with "tumbler test"); a fulminating form must be suspected in the presence of only one ecchymosis and signs of infection, remembering that recognition of shock is difficult in children. The Health Authority recommend to administer a third generation cephalosporin promptly for any child with signs of infection and an ecchymotic purpura (>3 mm of diameter), and then to refer the patient to the hospital. Children with purpura fulminans should be referred to a paediatric intensive care unit. Management includes antibiotics, steroids, fluid resuscitation and catecholamines (be aware of hypoglycaemia, particularly in infants, and hypocalcaemia). Treatment of cutaneous necrosis and distal ischemia is difficult and still controversial: antithrombin, protein C, tissue plasminogen activator and vasodilator infusion have no proved efficacy. Cases must be rapidly notified to the Public Health Service who will institute chemoprophylaxis for close contacts. Given the predominance of serogroup B in France, we hope that an efficient vaccine will soon become available. PMID- 15296273 TI - [Antibiotic treatment of bacterial meningitis]. AB - Community-acquired bacterial meningitis are due, in most of the cases, to S. pneumoniae and N. meningitidis. These two pathogenes are concerned by antibiotic resistance phenomenons. S. pneumoniae is developing resistance in some countries, including France. The recommended empirical treatment is nowadays a third generation cephalosporin, combined with vancomycin if there is a risk of Penicillin resistant pneumococcus. Non-antibiotic drugs are now validated and very useful adjunctive treatment. PMID- 15296274 TI - [Corticosteroid treatment in bacterial meningitis of adults and children]. AB - The neurologic deleterious effect of bacterial meningitis are the consequences of an inflammatory local response suggesting that an adjunctive anti-inflammatory therapy is able to favoured a better prognostic. Many clinical trials indicate that dexamethasone significantly reduce auditive sequella in children with meningitis due to Haemophilus influenzae, reduce mortality and morbidity of meningitis due to S. pneumoniae in adults but has few effect on Neisseria meningitidis. Corticotherapy should be initiated just before or at the time of first antibiotherapy and prolonged during 2 to 4 days. Major concern is a potential decrease of antibiotics concentration in cerebrospinal fluid that may be detrimental in patients with meningitis caused by S. pneumoniae strains that are highly resistant to penicillin or cephalosporins. PMID- 15296276 TI - [Patient's card: meningococcal meningitis]. PMID- 15296275 TI - [Chemoprophylaxis and vaccine for prevention of bacterial meningitis in children]. AB - Given the devastating nature of Neisseria meningitidis disease and emergence of resistant strains prevention through chemoprophylaxis and meningococcal vaccine remains the best approach to control this serious infection. Chemoprophylaxis may limited strictly to the contact subjects. Polysaccharide meningococcal serogroups A, C, Y and W135 should be given less than 10 days to patients with prolonged contact with the index case. Meningococcal C conjugate vaccine constitutes an additional advantage in the prevention of meningococcal meningitis in children < 2 years. High Haemophilus serotype B coverage level led to near-disappearance of H. influenzae serotype b meningitis but chemoprophylaxis remains indicated. PMID- 15296277 TI - [Specialty choice]. PMID- 15296278 TI - [Menopause and andropause]. PMID- 15296279 TI - [Sore throat and pharyngitis in children and in adults]. PMID- 15296280 TI - [Obesity in children and adults]. PMID- 15296281 TI - [Accidental exposure to blood]. PMID- 15296282 TI - [Evaluation of the severity, and search of early complications of burns]. PMID- 15296283 TI - [Monoclonal gammopathies]. PMID- 15296284 TI - [Labour, childbirth and normal third stage]. PMID- 15296285 TI - [A masterpiece in peril? The anatomy museums of Delmas-Orfila-Rouviere]. PMID- 15296286 TI - Top 10 stupid environmental policies. PMID- 15296287 TI - Bugs boost phytoremediation. PMID- 15296288 TI - Livestock flood the environment with estrogen. PMID- 15296289 TI - Americans receive wake-up call on oceans. PMID- 15296290 TI - U.S. rules are in step with other governments'. PMID- 15296291 TI - Desalination, desalination everywhere. PMID- 15296292 TI - Analytical challenges hamper perfluoroalkyl research. PMID- 15296293 TI - Brief survey of EPA standard-setting and health assessment. AB - The Environmental Protection Agency (EPA) promulgates standards for air pollutants and drinking water contaminants, as part of its mandate to protect public health and welfare. The Agency also assesses the health risks associated with hundreds of chemical substances, often developing quantitative toxicity and cancer potency benchmarks. This article compares EPA standards and benchmark values to those of other countries and other agencies. This includes the national ambient air quality standards (NAAQS), the national primary drinking water regulations (NPDWR), and benchmark values from the Integrated Risk Information System (IRIS). Results show that the NAAQS are generally comparable to or less strict than the air quality standards of other countries and international organizations. The NPDWR tend to be less strict than the water quality standards of other countries for inorganic chemicals, and they are more strict about as often as they are less strict for organic chemicals. Reference values for toxicity and cancer potency derived in EPA health assessments posted in the IRIS database are less stringentthan those of other agencies about as often as they are more stringent, and they are often identical. Revisions to these values more often than not made them less stringent. These results suggest that EPA's standards and quantitative health assessments are not out of line with those of other agencies and other countries. PMID- 15296294 TI - Life cycle assessment for sustainable metropolitan water systems planning. AB - Life Cycle Assessment (LCA) is useful as an information tool for the examination of alternative future scenarios for strategic planning. Developing a life cycle assessment for a large water and wastewater system involves making methodological decisions about the level of detail which is retained through different stages of the process. In this article we discuss a methodology tailored to strategic planning needs which retains a high degree of model segmentation in order to enhance modeling of a large, complex system. This is illustrated by a case study of Sydney Water, which is Australia's largest water service provider. A prospective LCA was carried out to examine the potential environmental impacts of Sydney Water's total operations in the year 2021. To our knowledge this is the first study to create an LCA model of an integrated water and wastewater system with this degree of complexity. A "base case" system model was constructed to represent current operating assets as augmented and upgraded to 2021. The base case results provided a basis for the comparison of alternative future scenarios and for conclusions to be drawn regarding potential environmental improvements. The scenarios can be roughly classified in two categories: (1) options which improve the environmental performance across all impact categories and (2) options which improve one indicator and worsen others. Overall environmental improvements are achieved in all categories by the scenarios examining increased demand management, energy efficiency, energy generation, and additional energy recovery from biosolids. The scenarios which examined desalination of seawater and the upgrades of major coastal sewage treatment plants to secondary and tertiary treatment produced an improvement in one environmental indicator but deteriorations in all the other impact categories, indicating the environmental tradeoffs within the system. The desalination scenario produced a significant increase in greenhouse gas emissions due to coal-fired electricity generation for a small increase in water supply. Assessment of a greenfield scenario incorporating water demand management, on-site treatment, local irrigation, and centralized biosolids treatment indicates significant environmental improvements are possible relative to the assessment of a conventional system of corresponding scale. PMID- 15296295 TI - Co-control of urban air pollutants and greenhouse gases in Mexico City. AB - This study addresses the synergies of mitigation measures to control urban air pollutant and greenhouse gas (GHG) emissions, in developing integrated "co control" strategies for Mexico City. First, existing studies of emissions reduction measures--PROAIRE (the air quality plan for Mexico City) and separate GHG studies--are used to construct a harmonized database of options. Second, linear programming (LP) is developed and applied as a decision-support tool to analyze least-cost strategies for meeting co-control targets for multiple pollutants. We estimate that implementing PROAIRE measures as planned will reduce 3.1% of the 2010 metropolitan CO2 emissions, in addition to substantial local air pollutant reductions. Applying the LP, PROAIRE emissions reductions can be met at a 20% lower cost, using only the PROAIRE measures, by adjusting investments toward the more cost-effective measures; lower net costs are possible by including cost-saving GHG mitigation measures, but with increased investment. When CO2 emission reduction targets are added to PROAIRE targets, the most cost effective solutions use PROAIRE measures for the majority of local pollutant reductions, and GHG measures for additional CO2 control. Because of synergies, the integrated planning of urban-global co-control can be beneficial, but we estimate that for Mexico City these benefits are often small. PMID- 15296296 TI - A 2.5-year genotoxicity profile for a partially restored polluted river. AB - This study evaluates water genotoxicity of the Kishon River, the most polluted river in Israel that is under restoration. Water samples were collected every other month (January 2001-May 2003) from five sites, and genotoxicity was assayed by the alkaline comet assay using a fish hepatoma cell line (RTH-149). Genotoxicity in the Kishon River was reduced during 2002 as compared to the previous year. The results further revealed fluctuations in genotoxicity levels at all sites throughout the studied period with variations for the same month during consecutive years and seasonality. In general, summer samples were more genotoxic than winter samples. In the vast majority of the 75 water samples, all four parameters for genotoxicity that were employed revealed significant higher genotoxic levels than the controls. Comet percentage values in Kishon River samples were, on average, 1.8-2.4 times higher than controls. Damage score, comet tail length, and cumulative tail length values were 2.2-3.1, 2.4-3.7, and 2.4-3.7 times higher than controls, respectively. The Histadrut Bridge and Haifa fishing harbor (3.0 m depth) emerged as the most polluted sites, whereas Kiryat Haroshet was the least contaminated. Results call for a long-term genotoxicity follow-up plan at the Kishon River in order to assess the possibly evolving chronic genotoxicity state. PMID- 15296297 TI - Chlordanes in the mid-Atlantic atmosphere: New Jersey 1997-1999. AB - To characterize the atmospheric dynamics and behavior of chlordane compounds in the mid-Atlantic region of the United States, atmospheric concentrations were measured in 1997-1999 at three New Jersey locations as part of the New Jersey Atmospheric Deposition Network (NJADN) project. Observed concentrations of sigma chlordanes (cischlordane + trans-chlordane + cis-nonachlor + transnonachlor) are log-normally distributed, with a geometric mean concentration of 77.1 pg m(-3) and range from 6.1 to 481 pg m(-3). Gas-phase species comprised 83% (+/-23%) of the sigma-chlordanes species across all samples at all location. Gas-phase sigma chlordane concentrations are inversely proportional to temperature, with higher concentrations during periods of warmer air temperatures. Observed concentrations do not correlate with wind direction or air mass history, which suggests that observed concentrations in the New Jersey atmosphere are due to volatilization from soils and surfaces on the regional scale. PMID- 15296298 TI - Sediment core record of global fallout and Bikini close-in fallout Pu in Sagami Bay, Western Northwest Pacific margin. AB - The total 239-240Pu activity and 240Pu/239Pu atom ratio in the sediments in Sagami Bay of the western Northwest Pacific margin were investigated using ICP-MS with a shield torch system. 239+240Pu inventories in the examined sediment cores were found to be much higher than those predicted from atmospheric global fallout (42 MBq/km2) at the same latitude. In addition, elevated 240Pu/239Pu atom ratios ranging from 0.22 to 0.28 were observed in the sediment samples. On the basis of the vertical profiles of 239+240Pu and characterized 240Pu/239Pu atom ratios in a sediment core collected in the center of Sagami Bay, we identified two distinct sources of fallout Pu in the bay: the global stratospheric fallout with characteristic 240Pu/239Pu ratio of 0.18 and the transported close-in fallout derived from Bikini and Enewetak surface nuclear weapon test series in the 1950s. We propose that the Pu transportation was mainly due to oceanic processes (for example, through the North Equatorial Current and the Kuroshio Current). Using a two fallout end-member model, we find that the contribution of Bikini close-in fallout Pu ranged from 44 to 59% in Sagami Bay sediments. To the best of our knowledge, this is the first report that Pu contamination, which originated from Bikini and Enewetak nuclear weapon test series in the 1950s, has extended westwards as far as the Japanese coast. PMID- 15296299 TI - Toxicokinetics of PCDD, PCDF, and coplanar PCB congeners in Baikal seals, Pusa sibirica: age-related accumulation, maternal transfer, and hepatic sequestration. AB - To assess the toxicokinetic behavior and potential toxicity of polychlorinated dibenzo-p-dioxins (PCDDs), polychlorinated dibenzofurans (PCDFs), and coplanar polychlorinated biphenyls (PCBs) in Baikal seals, congener-specific levels and tissue distribution were evaluated in the liver and blubber, and the effects of biological factors including sex and growth were assessed. Total 2,3,7,8-TCDD toxic equivalents (TEQs) were in the range of 210-920 pgTEQ/g fat wt (180-800 pgTEQ/g wet wt) in the blubber and 290-7800 pgTEQ/g fat wt (10-570 pgTEQ/wet wt) in the liver. Non-ortho coplanar PCB126 was the most TEQ-contributed congener accounting for 37-59% of the total TEQs in the liver. From the unique congener profiles, weak metabolic properties of Baikal seals for 2,3,7,8-TCDF and 1,2,3,7,8-P5CDF are suggested. Concentrations of most congeners linearly increased with age in male seals, whereas in adult females the levels revealed an age-related decline. The increasing and declining rates were congener-specific. Maternal transfer rates of 5 representative congeners from adult female to pup through lactation, which was estimated from male-female differences in the body burden, was 1.1 ngTEQ/kg/day for the first pup and decreased with every lactational epoch. The liver-blubber distribution of 1,2,3,4,7,8-H6CDD, 1,2,3,6,7,8-H6CDD, PCB81, PCB126, and PCB169 was dependent on the hepatic total TEQ, indicating hepatic sequestration by induced cytochrome P450 (CYP). These results indicate that congener profile in Baikal seals is governed by complex factors including sex, tissue concentration, binding to CYP, and rates of absorption and metabolism/excretion. PMID- 15296300 TI - Semivolatile organic compounds in window films from lower Manhattan after the September 11th World Trade Center attacks. AB - The September 11th World Trade Center (WTC) terrorist attacks resulted in the large-scale release of contaminants that were deposited on the environment of New York City (NYC). Six weeks after the attacks, samples of an organic film on window surfaces were collected and analyzed for polybrominated diphenyl ethers (PBDE), polychlorinated biphenyls (PCB), polychlorinated naphthalenes (PCN), polycyclic aromatic hydrocarbons (PAH), and organochlorine pesticides (OCPs). Concentrations dropped by an order of magnitude within 1 km of the WTC and reached background concentrations by 3.5 km. Concentrations within 1 km of the WTC averaged 3280 ng/m2 for sigmaPBDE, 900 ng/m2 for sigmaPCB, 33 ng/m2 for sigmaPCN, and 77100 ng/m2 for sigmaPAH. Congener profiles of the sites nearest the WTC suggested a combination of combustion and evaporative sources of all compounds, whereas the background sites exhibited profiles consistent with evaporative sources. PBDE profiles showed enrichment in lower molecular weight congeners near the WTC, suggesting that these congeners were formed as a result of the combustion conditions. Homologue fractions of PCN combustion markers were approximately 2-9 times greater at near WTC sites compared to background NYC. Gas phase air concentrations were back-calculated from measured film concentrations using the film-air partition coefficient (KFA), and calculated air concentrations followed spatial trends observed in films. PMID- 15296301 TI - Atmospheric semivolatile organochlorine compounds in European high-mountain areas (central Pyrenees and high Tatras). AB - Atmospheric samples from two European high-mountain areas showed similar composition of semivolatile organochlorine compounds (SOC), such as polychlorobiphenyls (PCBs), DDTs, endosulfans, hexachlorobenzene (HCB), and hexachlorocyclohexanes (HCHs). Nearly all compounds were predominantly found in the gas phase and only the less volatile such as some PCBs (e.g., 149, 118, 153, 138, and 180) were found in higher abundance in the particulate phase. HCB, 49-85 pg m(-3), is the dominant SOC. This compound is only found in the gas phase exhibiting uniform concentrations irrespective of season and air mass origin. SOC of present use, like HCHs and endosulfans, were found in higher concentrations in the warm periods, 32-46 and 4-10 pg m(-3) in the gas + particulate phases, respectively, reflecting their seasonal pattern of use in many European countries. PCB and 4,4'-DDE, 39-42 and 4-6 pg m(-3) in the gas + particulate phases, respectively, also showed a seasonal trend despite neither the former nor the precursor of the latter (4,4'-DDT) being manufactured with their use drastically restricted since the 1980s. The seasonal differences are mainly due to a higher occurrence of air masses with strong continental inputs in the warm than in the cold periods. In this respect, samples whose air masses traveled at the high troposphere (backward air mass trajectories >6000 m) have been observed to carry considerably smaller PCB and 4,4'-DDE loads (9.3 +/- 2.8 and 0.4 +/- 0.05 pg m(-3), respectively) than overall average. PMID- 15296302 TI - Occurrence of antimicrobials in the final effluents of wastewater treatment plants in Canada. AB - To investigate the occurrence of antimicrobials in the final effluents from wastewater treatment plants (WWTPs) in Canada, analytical methods were developed or modified from previously described methods using solid-phase extraction followed by liquid chromatography-electrospray ionization tandem mass spectrometry. Thirty-one antimicrobials from the macrolide, quinolone, quinoxaline dioxide, sulfonamide, and tetracycline classes were investigated in the final (treated) effluents from eight WWTPs, located in five Canadian cities. Ciprofloxacin, clarithromycin, erythromycin-H20, ofloxacin, sulfamethoxazole, sulfapyridine, and tetracycline were frequently detected in the effluents. The detection of sulfapyridine in effluents is the first report of this compound in environmental samples. Antimicrobials used exclusively for veterinary applications or treatment of livestock, such as carbadox, olaquindox, and chlortetracycline were not detected in the WWTP final effluents. There appear to be differences in the relative concentrations of antimicrobials detected in WWTP final effluents in Canada relative to concentrations reported previously in northern Europe, particularly for quinolone and sulfonamide compounds. These data may reflect differences in prescription patterns in Canada and northern Europe. The antimicrobials frequently detected in WWTP effluents appear to be those prescribed heavily in Canada for medical applications, and these compounds should be considered priority compounds for monitoring in surface water near WWTP discharges. The concentrations of antimicrobials detected in WWTP final effluents did not exceed 1 microg/L; levels that are unlikely to affect the growth and survival of aquatic organisms. PMID- 15296303 TI - Chemical characterization and sorption capacity measurements of degraded newsprint from a landfill. AB - Newsprint samples collected from 12-16 ft (top layer (TNP)), 20-24 ft (middle layer (MNP)), and 32-36 ft (bottom layer (BNP)) below the surface of the Norman Landfill (NLF) were characterized by infrared (IR) spectroscopy, cross polarization, magic-angle spinning 13C nuclear magnetic resonance (CP-MAS 13C NMR) spectroscopy, and tetramethylammonium hydroxide (TMAH) thermochemolysis gas chromatography/mass spectrometry (GC/MS). The extent of NLF newsprint degradation was evaluated by comparing the chemical composition of NLF newsprint to that of fresh newsprint (FNP) and newsprint degraded in the laboratory under methanogenic conditions (DNP). The O-alkyl/alkyl, cellulose/lignin, and lignin/resin acid ratios showed that BNP was the most degraded, and that all three NLF newsprint samples were more degraded than DNP. 13C NMR and TMAH thermochemolysis data demonstrated selective enrichment of lignin over cellulose, and TMAH thermochemolysis further exhibited selective enrichment of resin acids over lignin. In addition, the crystallinity of cellulose in NLF newsprint samples was significantly lower relative to that of FNP and DNP as shown by 13C NMR spectra. The yield of lignin monomers from TMAH thermochemolysis suggested that hydroxyl groups were removed from the propyl side chain of lignin during the anaerobic decomposition of newsprint in the NLF. Moreover, the vanillyl acid/aldehyde ratio, which successfully describes aerobic lignin degradation, was not a good indicator of the anaerobic degradation of lignin on the basis of the TMAH data. The toluene sorption capacity increased as the degree of newsprint degradation increased or as the O-alkyl/alkyl ratio of newsprint decreased. The results of this study further verified that the sorbent O-alkyl/ alkyl ratio is useful for predicting sorption capacities of natural organic materials for hydrophobic organic contaminants. PMID- 15296304 TI - Persistent organic pollutants at the base of the Antarctic marine food web. AB - Various organochlorine pesticides and brominated diphenyl ethers (BDE-47, -99, and -100) were measured in sea ice algae, water column plankton, and juvenile and adult krill collected in the Palmer Long-Term Ecological Research (LTER) region west of the Antarctic Peninsula during late austral winter and midsummer, 2001 2002. BDEs were 100-1000 times higher in ice algae and 2-10 times higher in phytoplankton than the most abundant organochlorine pesticide, hexachlorobenzene (HCB), reflecting the current production and use of BDEs versus organochlorine pesticides. However, concentrations of HCB and BDEs were significantly lower in summer plankton than in ice algae indicating lower atmospheric inputs, removal from the water column, and/or biodilution of persistent organic pollutants at the base of the food web during summer. Concentrations of HCB (juvenile and adult krill) and BDEs (juvenile krill) were not significantly different from their primary food source (ice algae, phytoplankton), and BDEs were significantly lower in adult krill versus phytoplankton, indicating no biomagnification of HCB or BDEs during transfer from plankton to krill. The high concentrations of BDEs and HCB in ice algae and associated juvenile krill illustrate the importance of sea ice as a vector for entry of POPs into the Antarctic marine ecosystem. PMID- 15296305 TI - Groundwater discharge: potential association with fecal indicator bacteria in the surf zone. AB - Short-lived radium isotopes (223Ra and 224Ra) are used to investigate the potential association between groundwater discharge and microbial pollution at Huntington Beach, CA. We establish the tidally driven exchange of groundwater from the surficial beach aquifer across the beach face. Groundwater is found to be a source of nutrients (silica, inorganic nitrogen, and orthophosphate) to the surf zone, and these nutrients could possibly provide an environment for enhanced growth or increased persistence of fecal indicator bacteria (FIB). Ammonium and ortho-phosphate explain up to 12-20% of the variance in FIB levels in the surf zone. Elevated levels of FIB were only found in 1 of the 26 groundwater samples. However, FIB in the surf zone covary with radium at fortnightly, diurnal, and semi-diurnal tidal periods. In addition, radium accounts for up to 38% of the variance in log-FIB levels in the surf zone. A column experiment illustrates that Enterococcus suspended in Huntington Beach saline groundwater is not significantly filtered by sand collected from the field. This work establishes a mechanism for the subterranean delivery of FIB pollution to the surf zone from the surficial aquifer and presents evidence that supports an association between groundwater discharge and FIB. PMID- 15296306 TI - Estrogen content of dairy and swine wastes. AB - Naturally occurring estrogens in animal wastes may cause negative environmental impacts, yet their abundance in animal waste treatment and storage structures is poorly documented. To better quantify estrogen concentrations in animal wastes, multiple waste samples were collected from treatment and storage structures at dairy and swine facilities and analyzed for concentrations of 17beta-estradiol (E2), estrone (E1), and 17alpha-estradiol by gas chromatography-mass spectroscopy and by enzyme linked immunosorbent assay (E2 only). Mass ratios of each estrogen to the macronutrients nitrogen, phosphorus, and potassium were also determined. Because manure application rates are typically macronutrient-based, estrogen to macronutrient ratios are proportional to areal mass application rates of estrogen to fields. Swine farrowing waste (from farrowing sows and piglets) had the highest ratios of E2 to macronutrients. Mean ratios in swine farrowing waste were roughly twice those in swine finishing waste (from growing male and nonpregnant female animals) and more than four times higher than those in dairy waste (from lactating cows in various stages of their reproductive cycles); these differences were statistically significant (alpha = 0.05). Estrone followed a similar trend. In contrast, ratios of 17alpha-estradiol to macronutrients were highest in dairy operations. These results can be used to better predict estrogen loading rates on fields receiving swine and dairy wastes. PMID- 15296307 TI - Effect of sorbate planarity on environmental black carbon sorption. AB - Soot and charcoal, collectively termed "black carbon" or BC, can exhibit extremely strong sorption of many hydrophobic organic compounds. In order to include BC sorption in fate models, it is important to know BC nanopore surface areas. In addition, it is useful to know for which compounds BC sorption can be expected to be important. By nitrogen adsorption measurements at ultralow pressures on sediment that was strongly enriched in BC by HF treatment and/or chemothermal oxidation at 375 degrees C, we found that environmental BC has nanoporosity in the <4-10 A size range. The nanopore surface area (<50 A) of BC in Lake Ketelmeer (The Netherlands) sediment was approximately 58 m2/g. We measured sorption isotherms over a wide concentration interval for four compounds with the same Kow (10(46+/-0.1): planar anthracene (ANT), phenanthrene (PHE), and 4-chlorobiphenyl (4-PCB) along with nonplanar 2,2'-dichlorobiphenyl (2,2'-PCB). The environmental BC sorption coefficients of these iso-Kow compounds decreased in the order ANT > PHE approximately 4-PCB >> 2,2'-PCB and spanned a factor of 50 200, depending on concentration. Nonplanar 2,2'-PCB showed much more linear BC sorption (nF = 0.92) than the planar compounds (nF = 0.54-0.70). This shows that steric hindrance strongly attenuates BC-sorbate interactions for a nonplanar PCB. Thus, BC is more important for environmental sorption of planar compounds (>50% sorbed to BC in the nanogram per liter range) than for nonplanar ones (<10-20%). Using the measured BC nanopore surface area, a close agreement between modeled and measured BC sorption data could be found. PMID- 15296308 TI - Selenium biotransformations in an insect ecosystem: effects of insects on phytoremediation. AB - Phytoremediation of selenium-contaminated soils may be influenced by higher trophic levels including insects. We examined how selenium affects the behavior, survival, and development of the wasp parasitoid Cotesia marginiventris, parasitizing its natural host, the beet armyworm Spodoptera exigua, feeding on alfalfa, Medicago sativa, irrigated with water containing selenate. X-ray absorption spectroscopy was used to quantify the selenium chemical forms in each trophic level. Alfalfa partially transformed selenate to organoselenium. S. exigua contained only organoselenium, both directly absorbed from M. sativa and transformed from selenate. C. marginiventris cocoons collected shortly after larval emergence contained only organoselenium derived from the host. The surprising finding of trimethylselenonium-like species in adult parasitoids and the cocoons from which they emerged suggests that adults and pharates can detoxify excess selenium through methylation and volatilization. Adult parasitoids do not discriminate against selenium-containing alfalfa, even though alfalfa generates selenium volatiles. Parasitoids raised on selenium-fed larvae emerged later and pupae weighed less than their selenium-free counterparts. We conclude therefore that C. marginiventris can be used to control S. exigua damage to M. sativa being used to remove selenium from soils. Moreover, the presence of such insects may improve phytoremediation by increasing biotransformation of inorganic selenium and release of volatile selenium species. PMID- 15296309 TI - Redox speciation of copper in rainwater: temporal variability and atmospheric deposition. AB - Copper speciation was determined in 68 rainwater samples collected in Wilmington, NC, from August 25, 2000, to September 24, 2002. Volume-weighted average concentrations of Cu(total), dissolved Cu(II), and dissolved Cu(I) were 5.3, 3.2, and 1.4 nM, respectively, with a significantly higher ratio of Cu(II)/Cu(I) in summer relative to winter events. The concentrations of all Cu species were higher in storms of continental origin relative to marine-dominated events, suggesting anthropogenic and/or terrestrial sources are important contributors of Cu in precipitation. Concentrations of strong Cu-complexing ligands were consistently lower than dissolved Cu concentrations, indicating a significant portion, but not all, of the dissolved Cu in rainwater is strongly complexed. A portion of these ligands, in addition to the sulfite and chloride in precipitation, may be Cu(I)-complexing ligands, which may explain the resistance of Cu(I) against oxidation in rainwater. Using our rainwater concentration data along with other published rainwater Cu concentrations and an estimate for total global annual rain, the total global flux of Cu removed from the atmosphere via wet deposition is 150 x 106 kg yr(-1). This represents complete removal of the estimated Cu input into the troposphere and indicates essentially all Cu released into the global atmosphere is removed by rain. PMID- 15296310 TI - Sorption nonlinearity for organic contaminants with diesel soot: method development and isotherm interpretation. AB - An experimentally practical and precise flocculation-based method was developed, tested, and applied to determine phenanthrene and 1,2,4-trichlorobenzene sorption with NIST SRM 2975 diesel particulate matter. Following an initial equilibration period, polyaluminum chloride (PACI) solution was added to the sorption tubes in order to facilitate the formation of flocculated aggregates of soot particles. After separation of the solids through centrifugation, supernatant concentrations were determined as with conventional batch methods. The flocculation-based method was tested on three kinds of soot and then used to evaluate sorption kinetics and equilibrium with SRM 2975. Kinetic results showed that wetting of the soot required more than 20 days, but that 60 days was sufficient to achieve equilibration with both water and phenanthrene. Sixty-day isotherms for both phenanthrene and 1,2,4-trichlorobenzene were strongly nonlinear. At approximate 10(-3) of solubility, carbon-normalized distribution coefficients (Koc) were 10 20 times higher than those for absorption to sediment organic matter. Measurements at closer to solubility indicated much lower Koc, suggesting a total sorption capacity at aqueous solubility that is of similar magnitude to that in sediment organic matter. Independent analysis of extractable hydrocarbons suggests that absorption into a native hydrocarbon phase was not a major component of sorption. PMID- 15296311 TI - Using chromium stable isotope ratios to quantify Cr(VI) reduction: lack of sorption effects. AB - Chromium stable isotope values can be effectively used to monitor reduction of Cr(VI) in natural waters. We investigate effects of sorption during transport of Cr(VI) which may also shift Cr isotopes values, complicating efforts to quantify reduction. This study shows that Cr stable isotope fractionation caused by sorption is negligible. Equilibrium fractionation of Cr stable isotopes between dissolved Cr(VI) and Cr(VI) adsorbed onto gamma-Al2O3 and goethite is less than 0.04 per thousand (53Cr/52Cr) under environmentally relevant pH conditions. Batch experiments at pH 4.0 and pH 6.0 were conducted in series to sequentially magnify small isotope fractionations. A simple transport model suggests that adsorption may cause amplification of a small isotope fractionation along extreme fringes of a plume, leading to shifts in 53Cr/52Cr values. We therefore suggest that isotope values at extreme fringes of Cr plumes be critically evaluated for sorption effects. A kinetic effect was observed in experiments with goethite at pH 4 where apparently lighter isotopes diffuse into goethite clumps at a faster rate before eventually reaching equilibrium. This observed kinetic effect may be important in a natural system that has not attained equilibrium and is in need of further study. Cr isotope fractionation caused by speciation of Cr(VI) between HCrO4- and CrO4(2-) was also examined, and we conclude that it is not measurable. In the absence of isotope fractionation caused by equilibrium speciation and sorption, most of the variation in delta53Cr values may be attributed to reduction, and reliable estimates of Cr reduction can be made. PMID- 15296312 TI - Kinetics of Zn release in soils and prediction of Zn concentration in plants using diffusive gradients in thin films. AB - Effective concentrations (CE) of Zn measured by the technique of DGT (diffusive gradients in thin films) were compared, along with total concentrations of Zn and the concentrations of Zn in soil solutions, to Zn concentrations in plants. Soils variously contaminated with Zn were collected in the vicinity of two galvanized electrical transmission towers (pylons) and two motorway crash barriers. Lepidium sativum was grown in each soil and in corresponding control soils amended with ZnCl2 to similar total Zn concentrations. CE, concentrations in soil solution, and total Zn were measured in all soils, and total Zn was measured in the plant shoots. The CE values, soil solution Zn, and shoot Zn concentrations were all larger in ZnCl2 amended soils than in field contaminated soils at corresponding total Zn. Correlations between the concentration of Zn in the plants and the measured soil parameter followed the order CE > soil solution > total Zn. The low scatter in the plot of log plant concentration versus log CE revealed a relationship with two distinct features. Plant Zn was between 100 and 300 mg/kg up to an effective Zn concentration of about 2 mg/L, above which plant Zn increased steadily with increasing CE. Use of a dynamic model to interpret the DGT measurement suggests that the intrinsic rate of release of Zn from solid phase to solution, expressed as a rate constant, is much higher for soils that receive fresh supplies of Zn. This finding provides a mechanistic basis for reconciling laboratory experiments, where metal is freshly amended, to data obtained in the field. The potential of DGT as a surrogate for metal availability to plants is further confirmed by this work. PMID- 15296313 TI - Inter-individual differences in the ability of human milk-fat extracts to enhance the genotoxic potential of the procarcinogen benzo[a]pyrene in MCF-7 breast cells. AB - Environmental factors are believed to play an important role in cancer aetiology. Whether environmental pollutants act in isolation or in combination within mixtures remains unclear. Four human milk-fat extracts (from resident U.K. women) were screened for levels of organochlorinated and brominated compounds prior to being tested (1-50 mg-equiv) for micronucleus (MN)-forming activity in MCF-7 cells. Using the cytokinesis-block micronucleus assay, micronuclei (MNi) were scored in 1000 binucleate cells per treatment. Cell viability (% plating efficiency) and immunohistochemical detection of p53 induction were also measured. The effects of treatment with 1 mg-equiv of extract in combination with benzo[a]pyrene (BP) were also examined. BP-DNA adducts were detected and quantified by 32P-postlabeling analysis. Dose-related increases in MNi independent of pollutant concentrations were induced by milk-fat extracts. All four extracts elevated the percentage of p53 positive cells, although not always in a dose-related fashion. Some combinations resulted in profound low-dose induced increases in MNi and significant elevations in the percentage of p53 positive cells, which occurred without further reduction in cell viability or mitotic rate. When one particular extract was combined with BP, a 100-fold increase in BP-DNA adducts was detected as compared with the levels induced by BP alone; an effect not induced by other extracts. This adduct-enhancing extract was fractionated into 14 fractions that were subsequently tested (1 mg-equiv of original extract) in combination with 0.01 microM BP. Fraction 1, into which nonpolar pollutants mostly eluted, enhanced MN-forming activity with BP. Surprisingly, the more polar and less likely to contain fat-soluble pollutants fractions 5 and 8 also enhanced MN-forming activity. No identifiable pollutants were present in these fractions. The results suggest that different environmental pollutants present in human tissue may influence the susceptibility of target cells to initiating events. PMID- 15296314 TI - Deriving soil critical limits for Cu, Zn, Cd, and Pb: a method based on free ion concentrations. AB - We present a method to calculate critical limits of cationic heavy metals accounting for variations in soil chemistry. We assume the free metal ion concentration (Mfree) to be the most appropriate indicator of toxicity, combined with a protective effect of soil cations (e.g., H+, Ca2+). Because soil metal cations tend to covary with pH, the concentration of Mfree exerting a given level of toxic effect (Mfree,toxic) can be expressed as a function of pH alone. We use linear regression equations to derive Mfree,toxic in toxicity experiments from soil pH, organic matter content, and endpoint soil metal. Chronic toxicity data from the literature, for plants, invertebrates, microbial processes, and fungi are interpreted in terms of an average log Mfree,toxic together with distributions of species sensitivity. This leads to critical limit functions to protect 95% of species, of the form log Mfree,CRIT = (pH + gamma. Appreciable effects of soil pH upon log Mfree,CRIT are found, with alpha = -1.21 (Cu), -0.34 (Zn), -0.43 (Cd), and -0.83 (Pb). Critical limit functions in terms of the geochemically active soil metal (Msoil,CRIT), that pool of metal which controls the free ion concentration, have also been derived, with soil pH and organic matter content as variables. The pH effect on Msoil,CRIT is relatively small, with slopes of 0.05 (Cu), 0.19 (Zn), 0.16 (Cd), and 0.20 (Pb), since the effect of pH on Mfree,CRIT is countered by the variation of Mfree with pH. PMID- 15296315 TI - Relations between environmental black carbon sorption and geochemical sorbent characteristics. AB - Pyrogenic carbon particles in sediments (soot and charcoal, collectively termed "black carbon" or BC) appear to be efficient sorbents of many hydrophobic organic compounds, so they may play an important role in the fate and toxicity of these substances. To properly model toxicant sorption behavior, it is important to (i) quantify the magnitude of the role of BC in sorption and (ii) elucidate which geochemical BC characteristics determine the strength of environmental BC sorption. Sorption isotherms of d10-phenanthrene (d10-PHE) were determined over a wide concentration range (0.0003-20 microg/L), for five sediments with widely varying characteristics. From the sorption isotherms, we determined Freundlich coefficients of environmental BC sorption, K(F,BCenv. These varied from 10(4.7) to 10(5.5). From the data, it could be deduced that BC was responsible for 49-85% of the total d10-PHE sorption at a concentration of 1 ng/L. At higher concentrations, the importance of BC for the sorption process diminished to <20% at 1 microg/L and 0-1% at 1 mg/L. There were no significant relationships between BC sorption strength and the tested geochemical BC characteristics [the fraction of small (<38 microm) BC particles, the BC resistance to high-temperature oxidation, the fraction of biomass-derived BC, the native polycyclic aromatic hydrocarbon and total organic carbon contents]. Because of the limited variation in BC sorption strength with widely varying BC characteristics, the presented BC sorption coefficients may putatively be used as generic starting points for environmental modeling purposes. PMID- 15296316 TI - Predominance and mineral stability diagrams revisited. AB - Predominance and mineral stability diagrams are most useful if they are based on a full chemical speciation. The diagrams can then include a wide range of reactions of importance in environmental chemistry including adsorption and ion exchange reactions. They can also take into account the possible interactions between a large number of components and are able to model changes at a fixed total concentration of each component rather than under the commonly used but less realistic assumption of a fixed activity. A "hunt-and-track" algorithm is described that enables predominance and mineral stability diagrams to be constructed using any general-purpose speciation program. Examples are given using PHREEQC and Orchestra. PMID- 15296317 TI - A model to estimate influent and effluent concentrations of estradiol, estrone, and ethinylestradiol at sewage treatment works. AB - To predict sewage influent and effluent concentrations of the steroid estrogens 17beta-estradiol, estrone, and 17alpha-ethinylestradiol, a review of human excretion was carried out. This included conjugation and metabolism of the natural and synthetic steroid estrogens within the body, together with quantities excreted in the urine and feces by different members of the population. This has been combined with fate and behavior information for conjugated and unconjugated estrogens in the sewage treatment system to enable sewage works influent and effluent concentration predictions to be made. The model has proved to be reasonably accurate when tested against recent measurements of these steroid estrogens in the influent and effluent of sewage treatment works. The model may be used with river dilution ratios to predict which sewage treatment works are most likely to cause the greatest endocrine disruption due to steroid estrogens. PMID- 15296318 TI - Toxic ratio as an indicator of the intrinsic toxicity in the assessment of persistent, bioaccumulative, and toxic chemicals. AB - Persistence, bioconcentration, and toxicity (PBT) are important hazardous properties of organic chemicals. In PBT assessments, it is desirable that the three criteria P, B, and T are independent. However, this requirement is not fulfilled if an aqueous lethal concentration (LC50) is used as T indicator because LC50 includes both bioconcentration and intrinsic toxicity. Indicators for intrinsic toxicity such asthe internal lethal concentration (ILC) are independent of a chemical's bioconcentration potential. However, ILC50 data are scarce and difficult to measure. Therefore, the toxic ratio (TR) is proposed here as an alternative. TR is defined as the ratio of a chemical's LC50 estimated from a QSAR for baseline toxicity and the experimental LC50 value. TR can also be interpreted as a measure of the ILC relative to the ILC for baseline toxicity. A TR of 10 separates specifically toxic chemicals from baseline toxicants. With some 800 chemicals, the practicability of classifying chemicals in terms of TR is demonstrated. Employing TR as toxicity indicator leads to different T scores for 30% of the chemicals studied. The baseline toxicity of hydrophobic compounds with TR < 10 does not receive a high T score but is still indicated by a high B score. The toxicity of specifically toxic hydrophilic substances is given additional emphasis by high TR values. These classification changes require that the interpretation of the B and T dimensions in PBT assessments is redefined. PMID- 15296319 TI - Adsorption of organic vapors to air-dry soils: model predictions and experimental validation. AB - Soil/air equilibrium partitioning has an important impact on the environmental distribution and fate of many organic chemicals. Modeling approaches that cover this process commonly assume that sorption in soil only occurs in soil organic matter. However, many researchers have already shown thatthis is not even correctfor nonpolar compounds in air-dry soils. Here, we extend the existing data set on sorption in air-dry soils by using a large and very diverse set of organic compounds covering many different functional groups for two standard soils and for relative humidity between 50 and 90%. The experimental data presented here as well as those from the literature are then used to examine two different modeling approaches: one that only considers absorption in soil organic matter and one that also considers adsorption to mineral surfaces. The results clearly show that sorption in air-dry soil cannot be explained when adsorption to mineral surfaces is ignored. Only the model that considers both sorbing phases, organic matter and mineral surfaces, gives good agreement for all experimental data. Our model for predicting adsorption to mineral surfaces does not require any further fitting with experimental data; thus, it can readily be incorporated into existing fate models in order to improve the description of the soil/air partition process. PMID- 15296320 TI - Electro-oxidation and amperometric detection of chlorinated phenols at boron doped diamond electrodes: a comparison of microcrystalline and nanocrystalline thin films. AB - We report on the electro-oxidation and amperometric detection of phenol and chlorinated phenols, the latter coupled with flow injection analysis (FIA) and high performance liquid chromatography (HPLC), using boron-doped microcrystalline and nanocrystalline diamond thin-film electrodes. The low background current, good response without extensive pretreatment, and low susceptibility to fouling are properties that make diamond an attractive new electrode for monitoring this class of pollutants. Cyclic voltammetric studies were performed to evaluate the redox response of phenol, 2-chlorophenol, 3-chlorophenol,4-chlorophenol, and pentachlorophenol (PCP) in phosphate buffer, pH 3.5, as a function of the potential scan rate and cycle number. The diamond electrode performance for the amperometric detection of these contaminants in FIA-EC and HPLC-EC was evaluated in terms of the linear dynamic range, limit of quantitation, sensitivity, response precision, and response stability. Both diamond types yielded low mass limits of quantitation of 100-1000 pg for all the phenolic compounds in FIA-EC, except PCP which was 3 ng, and 100-600 pg for all the compounds in HPLC-EC. In all cases, the S/N was 3 or greater. Both electrode types also exhibited good sensitivity, excellent response reproducibility (av 2.7% for FIA-EC and av 4.2% for HPLC-EC), and superb response stability for all the analytes. The electrodes could be used from days to weeks in the measurement with only a periodic soak in distilled 2-propanol required to maintain optimum performance. Both types of diamond outperformed glassy carbon, which exhibited short-lived responsiveness as a consequence of fouling by reaction products and potential-dependent changes in the electrode's physiochemical properties. The use of the HPLC-EC assay for the determination of 2-chlorophenol in a contaminated soil sample is also demonstrated. PMID- 15296321 TI - Tedlar bag sampling technique for vertical profiling of carbon dioxide through the atmospheric boundary layer with high precision and accuracy. AB - Carbon dioxide is the most important greenhouse gas other than water vapor, and its modulation by the biosphere is of fundamental importance to our understanding of global climate change. We have developed a new technique for vertical profiling of CO2 and meteorological parameters through the atmospheric boundary layer and well into the free troposphere. Vertical profiling of CO2 mixing ratios allows estimates of landscape-scale fluxes characteristic of approximately100 km2 of an ecosystem. The method makes use of a powered parachute as a platform and a new Tedlar bag air sampling technique. Air samples are returned to the ground where measurements of CO2 mixing ratios are made with high precision (< or =0.1%) and accuracy (< or =0.1%) using a conventional nondispersive infrared analyzer. Laboratory studies are described that characterize the accuracy and precision of the bag sampling technique and that measure the diffusion coefficient of CO2 through the Tedlar bag wall. The technique has been applied in field studies in the proximity of two AmeriFlux sites, and results are compared with tower measurements of CO2. PMID- 15296322 TI - Compound-specific carbon and hydrogen isotope analysis of sub-parts per billion level waterborne petroleum hydrocarbons. AB - Compound-specific carbon and hydrogen isotope analysis (CSCIA and CSHIA) has been increasingly used to study the source, transport, and bioremediation of organic contaminants such as petroleum hydrocarbons. In natural aquatic systems, dissolved contaminants represent the bioavailable fraction that generally is of the greatest toxicological significance. However, determining the isotopic ratios of waterborne hydrophobic contaminants in natural waters is very challenging because of their extremely low concentrations (often at sub-parts ber billion, or even lower). To acquire sufficient quantities of polycyclic aromatic hydrocarbons with 10 ng/L concentration for CSHIA, more than 1000 L of water must be extracted. Conventional liquid/liquid or solid-phase extraction is not suitable for such large volume extractions. We have developed a new approach that is capable of efficiently sampling sub-parts per billion level waterborne petroleum hydrocarbons for CSIA. We use semipermeable membrane devices (SPMDs) to accumulate hydrophobic contaminants from polluted waters and then recover the compounds in the laboratory for CSIA. In this study, we demonstrate, under a variety of experimental conditions (different concentrations, temperatures, and turbulence levels), that SPMD-associated processes do not induce C and H isotopic fractionations. The applicability of SPMD-CSIA technology to natural systems is further demonstrated by determining the delta13C and deltaD values of petroleum hydrocarbons present in the Pawtuxet River, RI. Our results show that the combined SPMD-CSIA is an effective tool to investigate the source and fate of hydrophobic contaminants in the aquatic environments. PMID- 15296323 TI - Automated solid-phase extraction and measurement of perfluorinated organic acids and amides in human serum and milk. AB - Organic fluorochemicals are used in multiple commercial applications including surfactants, lubricants, paints, polishes, food packaging, and fire-retarding foams. Recent scientific findings suggest that several perfluorochemicals (PFCs), a group of organic fluorochemicals, are ubiquitous contaminants in humans and animals world wide. Furthermore, concern has increased about the toxicity of these compounds. Therefore, monitoring human exposure to PFCs is important. We have developed a high-throughput method for measuring trace levels of 13 PFCs (2 perfluorosulfonates, 8 perfluorocarboxylates, and 3 perfluorosulfonamides) in serum and milk using an automated solid-phase extraction (SPE) cleanup followed by high-performance liquid chromatography-tandem mass spectrometry. The method is sensitive, with limits of detection between 0.1 and 1 ng in 1 mL of serum or milk, is not labor intensive, involves minimal manual sample preparation, and uses a commercially available automated SPE system. Our method is suitable for large epidemiologic studies to assess exposure to PFCs. We measured the serum levels of these 13 PFCs in 20 adults nonoccupationally exposed to these compounds. Nine of the PFCs were detected in at least 75% of the subjects. Perfluorooctanesulfonate (PFOS), perfluorohexanesulfonate (PFHxS), 2-(N methylperfluorooctane-sulfonamido)acetate (Me-PFOSA-AcOH), perfluorooctanoate (PFOA), and perfluorononanoate (PFNA) were found in all of the samples. The concentration order and measured levels of PFOS, PFOA, Me-PFOSA-AcOH, and PFHxS compared well with human serum levels previously reported. Although no human data are available for the perfluorocarboxylates (except PFOA), the high frequency of detection of PFNA and other carboxylates in our study suggests that human exposure to long-alkyl-chain perfluorocarboxylates may be widespread. We also found PFOS in the serum and milk of rats administered PFOS by gavage, but not in the milk of rats not dosed with PFOS. Furthermore, we did not detect most PFCs in two human milk samples. These findings suggest that PFCs may not be as prevalent in human milk as they are in serum. Additional studies are needed to determine whether environmental exposure to PFCs can result in PFCs partitioning into milk. Large epidemiological studies to determine the levels of PFCs among the U.S. general population are warranted. PMID- 15296324 TI - Radical generation by the interaction of transition metals with common oxidants. AB - Nine transition metals were tested for the activation of three oxidants and the generation of inorganic radical species such as sulfate, peroxymonosulfate, and hydroxyl radicals. From the 27 combinations, 14 M/Ox couples demonstrated significant reactivity toward transforming a model organic substrate such as 2,4 dichlorophenol and are further discussed here. It was found that Co(II) and Ru(III) are the best metal catalysts for the activation of peroxymonosulfate. As expected on the basis of the Fenton reagent, Fe(III) and Fe(II) were the most efficient transition metals for the activation of hydrogen peroxide. Finally, Ag(I) showed the best results toward activating persulfate. Quenching studies with specific alcohols (tert-butyl alcohol and ethanol) were also performed to identify the primary radical species formed from the reactive M/Ox interactions. The determination of these transient species allowed us to postulate the rate determining step of the redox reactions taking place when a metal is coupled with an oxidant in aqueous solution. It was found that when Co(II), Ru(III), and Fe(II) interact with peroxymonosulfate, freely diffusible sulfate radicals are the primary species formed. The same was proven for the interaction of Ag(I) with persulfate, but in this case caged or bound to the metal sulfate radicals might be formed as well. The conjunction of Ce(III), Mn(II), and Ni(II) with peroxymonosulfate showed also to generate caged or bound to the metal sulfate radicals. A combination of sulfate and hydroxyl radicals was formed from the conjunction of V(III) with peroxymonosulfate and from Fe(II) with persulfate. Finally, the conjunction of Fe(III), Fe(II), and Ru(III) with hydrogen peroxide led primarily to the generation of hydroxyl radicals. It is also suggested here that the redox behavior of a particular metal in solution cannot be predicted based exclusively on its size and charge. Additional phenomena such as metal hydrolysis as well as complexation with other counterions present in solution might affect the thermodynamics of the overall process and are further discussed here. PMID- 15296325 TI - Vulnerability of water distribution systems to pathogen intrusion: how effective is a disinfectant residual? AB - Can the spread of infectious disease through water distribution systems be halted by a disinfectant residual? This question is overdue for an answer. Regulatory agencies and water utilities have long been concerned about accidental intrusions of pathogens into distribution system pipelines (i.e., cross-connections) and are increasingly concerned about deliberate pathogen contamination. Here, a simulation framework is developed and used to assess the vulnerability of a water system to microbiological contamination. The risk of delivering contaminated water to consumers is quantified by a network water quality model that includes disinfectant decay and disinfection kinetics. The framework is applied to two example networks under a worst-case deliberate intrusion scenario. Results show that the risk of consumer exposure is affected by the residual maintenance strategy employed. The common regulation that demands a "detectable" disinfectant residual may not provide effective consumer protection against microbial contamination. A chloramine residual, instead of free chlorine, may significantly weaken this final barrier against pathogen intrusions. Moreover, the addition of a booster station at storage tanks may improve consumer protection without requiring excessive disinfectant. PMID- 15296326 TI - Reduction of nitroglycerin with elemental iron: pathway, kinetics, and mechanisms. AB - Nitroglycerin (NG) is a nitrate ester used in dynamites, propellants, and medicines and is therefore a common constituent in propellant-manufacturing and pharmaceutical wastewaters. In this study we investigated the reduction of NG with cast iron as a potential treatment method. NG was reduced stepwise to glycerol via 1,2- and 1,3-dinitroglycerins (DNGs) and 1- and 2-mononitroglycerins (MNGs). Nitrite was released in each reduction step and was further reduced to NH4+. Adsorption of NG and its reduction products to cast iron was minimal. A reaction pathway and a kinetic model for NG reduction with cast iron were proposed. The estimated surface area-normalized reaction rate constants for NG and NO2- were (1.65 +/- 0.30) x 10(-2) (L x m(-2) x h(-1)) and (0.78 +/- 0.09) x 10(-2) (L x m(-2) x h(-1)), respectively. Experiments using dialysis cell with iron and a graphite sheet showed that reduction of NG to glycerol can be mediated by graphite. However, reduction of NO2- mediated by graphite was very slow. NG and NO2- were also found to reduce to glycerol and NH4+ by Fe2+ in the presence of magnetite but not by aqueous Fe2+ or magnetite alone. These results indicate that in a cast iron-water system NG may be reduced via multiple mechanisms involving different reaction sites, whereas nitrite is reduced mainly by iron and/ or adsorbed Fe2+. The study demonstrates that iron can rapidly reduce NG to innocuous and biodegradable end products and represents a new approach to treat NG-containing wastewaters. PMID- 15296327 TI - Effect of cellulose/hemicellulose and lignin on the bioavailability of toluene sorbed to waste paper. AB - Paper constitutes about 38% of municipal solid waste, much of which is disposed of in landfills. Sorption to such lignocellulosic materials may limit the bioavailability of organic contaminants in landfills. The objective of this study was to identify the effect of individual biopolymers in paper on toluene sorption and bioavailability by subjecting fresh and anaerobically degraded office paper and newsprint to enzymatic hydrolysis and acid hydrolysis. Enzymatic degradation of cellulose and hemicellulose had no effect on toluene bioavailability. In contrast, acid-insoluble lignin controlled toluene sorption and bioavailability for both fresh and degraded newsprint. Acid-insoluble lignin could explain only 54% of the toluene sorption capacity of degraded office paper however, suggesting that crude protein and/or lipophilic organic matter were also important sorbent phases. Toluene sorbed to degraded office paper was also less bioavailable than toluene sorbed to an equivalent mass of lignin extracted from this sorbent. The latter result suggests that a fraction of toluene sorbed to degraded office paper may have been sequestered by lipophilic organic matter. The sorption and bioavailability data indicate that the preferential decomposition of cellulose and hemicellulose relative to lignin in landfills should not decrease the overall toluene sorption capacity of paperwaste or increase the bioavailability of sorbed toluene. PMID- 15296328 TI - Utilization of solar energy in the photodegradation of gasoline in water and of oil-field-produced water. AB - The photo-Fenton process utilizes ferrous ions (Fe2+), hydrogen peroxide (H2O2), and ultraviolet (UV) irradiation as a source of hydroxyl radicals for the oxidation of organic matter present in aqueous effluents. The cost associated with the use of artificial irradiation sources has hindered industrial application of this process. In this work, the applicability of solar radiation for the photodegradation of raw gasoline in water has been studied. The photo Fenton process was also applied to a real effluent, i.e., oil-field-produced water, and the experimental results demonstrate the feasibility of employing solar irradiation to degrade this complex saturated-hydrocarbon-containing system. PMID- 15296329 TI - Role of surface alteration in determining the mobility of U(VI) in the presence of citrate: implications for extraction of U(VI) from soils. AB - In the present study, the adsorption of U(VI) by a natural iron-rich sand in the presence of citrate was studied over a range of citrate concentrations and pH values. Adsorption of U(VI) on the iron-rich sand decreased in the presence of increasing concentrations of citrate. Adsorption of citrate to the sand was weak under most conditions studied. Several explanations for the adsorption behavior of U(VI) and citrate were investigated, including aqueous complexation of U(VI) by citrate, competition of U(VI) and citrate for adsorption sites, and extraction of Fe and Al from the sorbent surface by citrate (surface alteration). Although aqueous complexation of U(VI) by citrate may still play a significant role, both competitive adsorption and aqueous complexation proved to be inadequate explanations of the adsorption behavior. Both physical surface alteration (i.e., loss of surface area) and chemical surface alteration (i.e., change in the chemical composition of the sand surface) were investigated, with chemical surface alteration controlling the bulk of U(VI) adsorption. Considering these results, remediation schemes that involve organic complexing agents should address the possibility of surface alteration affecting radionuclide adsorption and mobility. PMID- 15296330 TI - Speciation of zinc in municipal solid waste incineration fly ash after heat treatment: an X-ray absorption spectroscopy study. AB - Fly ash is commonly deposited in special landfills as it contains toxic concentrations of heavy metals, such as Zn, Pb, Cd, and Cu. This study was inspired by our efforts to detoxify fly ash from municipal solid waste incineration by thermal treatment to produce secondary raw materials suited for reprocessing. The potential of the thermal treatment was studied by monitoring the evaporation rate of zinc from a certified fly ash (BCR176) during heating between 300 and 950 degrees C under different carrier gas compositions. Samples were quenched at different temperatures for subsequent investigation with X-ray absorption spectroscopy (XAS). The XAS spectra were analyzed using principal component analysis (PCA), target transformation (TT), and linear combination fitting (LCF) to analyze the major Zn compounds in the fly ash as a function of the temperature. The original fly ash comprised about 60% zinc oxides mainly in the form of hydrozincite (Zn5(OH)6(CO3)2) and 40% inerts like willemite (Zn2SiO4) and gahnite (ZnAl2O4) in a weight ratio of about 3:1. At intermediate temperatures (550-750 degrees C) the speciation underlines the competition between indigenous S and Cl with solid zinc oxides to form either volatile ZnCl2 or solid ZnS. ZnS then transformed into volatile species at about 200 degrees C higher temperatures. The inhibiting influence of S was found absent when oxygen was introduced to the inert carrier gas stream or chloride-donating alkali salt was added to the fly ash. PMID- 15296331 TI - Expanding exergy analysis to account for ecosystem products and services. AB - Exergy analysis is a thermodynamic approach used for analyzing and improving the efficiency of chemical and thermal processes. It has also been extended for life cycle assessment and sustainability evaluation of industrial products and processes. Although these extensions recognize the importance of capital and labor inputs and environmental impact, most of them ignore the crucial role that ecosystems play in sustaining all industrial activity. Decisions based on approaches that take nature for granted continue to cause significant deterioration in the ability of ecosystems to provide goods and services that are essential for every human activity. Accounting for nature's contribution is also important for determining the impact and sustainablility of industrial activity. In contrast, emergy analysis, a thermodynamic method from systems ecology, does account for ecosystems, but has encountered a lot of resistance and criticism, particularly from economists, physicists, and engineers. This paper expands the engineering concept of Cumulative Exergy Consumption (CEC) analysis to include the contribution of ecosystems, which leads to the concept of Ecological Cumulative Exergy Consumption (ECEC). Practical challenges in computing ECEC for industrial processes are identified and a formal algorithm based on network algebra is proposed. ECEC is shown to be closely related to emergy, and both concepts become equivalent if the analysis boundary, allocation method, and approach for combining global energy inputs are identical. This insight permits combination of the best features of emergy and exergy analysis, and shows that most of the controversial aspects of emergy analysis need not hinder its use for including the exergetic contribution of ecosystems. Examples illustrate the approach and highlight the potential benefits of accounting for nature's contribution to industrial activity. PMID- 15296332 TI - Origin of carbon in polychlorinated dioxins and furans formed during sooting combustion. AB - The importance of solid- and gas-phase carbon precursors for the formation of polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDDs/Fs) during sooting combustion was investigated in an entrained flow reactor (EFR). Experiments were performed at various methane (CH4) flame equivalence ratios with or without gas phase chlorine (Cl2) and fly ash, to provide a realistic environment for carbon reactions and PCDD/DF formation. Selected experiments were conducted with labeled 13CH4 and 37Cl2 to investigate the relative roles of different carbon and chlorine species for the formation of PCDD/DF. The presence of soot and ash were the two major factors controlling the PCDD/DF yields. The 16 PCDD/DF homologues as well as other analyzed chlorinated aromatics were formed by reaction pathways that varied with degree of chlorination. The mono- and dichlorinated homologues were formed by gas-phase, catalytic, or noncatalytic flame product reactions, occurring during soot formation in the near flame zone and/or at lower reaction temperatures (<650 degrees C) in the postcombustion zone. Meanwhile, the higher (tri- to octa-) chlorinated homologues were mainly formed in the postcombustion zone (<650 degrees C) by fly ash-catalyzed de novo synthesis of the soot. Of these, the PCDD/DFs were formed from high carbon number (>C12) fragments in the solid soot structure, while the PCDDs, at least in part, were also formed by reaction of two C6 fragments. The tri- to hexachlorinated DD/DF homologues were formed via a relatively fast de novo synthesis occurring during the first minutes of reactions on the continuously formed soot particles, whereas de novo synthesis on an aged soot matrix was the major pathway for the hepta- and octachlorinated congeners. PMID- 15296333 TI - Construction of a low-pressure microwave plasma reactor and its application in the treatment of volatile organic compounds. AB - Volatile organic compounds (VOCs) containing gases exhausted from industrial processes are highly toxic to human health; therefore, their safe elimination is extremely important. This paper describes how a novel prototype microwave plasma reactor was sequentially improved. The design concepts for each modification are also carefully described. For testing the reactor's performance, ethanol was selected as the target VOC. The decomposing and removal efficiency (DRE) value was used to evaluate the reactor's performance. The final stage of the revamping reactor was evaluated to meet the requirements of both stability and long term, high volume operation. When studying the VOC treatment operations, oxygen, air, and water vapor were used as carrier gases or additives to improve the efficiency of the DRE value and to facilitate the ethanol treatment. A DRE value higher than 99% was obtained under these optimal conditions: an ethanol vapor flow rate of 1730 cm3/min (67 mbar, 150 degrees C), power input higher than 1.5 kW, a frequency of 200 Hz, an air flow rate of 100 cm3/min (4 atm, 25 degrees C), and a liquid water addition rate higher than 0.21 mL/min (1 atm, 25 degrees C). With these conditions, the initial concentration of ethanol vapor mixed with the carrier gas was about 7-10%. Because of high processing efficiency and capacity of this system, it could be a potential alternative tool for treating industrial VOCs. PMID- 15296334 TI - [Benign desmoplastic fibroblastoma of the lower leg]. PMID- 15296335 TI - Nodular fasciitis: diagnosis by fine needle aspiration biopsy. AB - OBJECTIVE: To describe the cytomorphologic features of nodular fasciitis that differentiate it from schwannoma. STUDY DESIGN: The cytomorphologic features of 10 cases of nodular fasciitis were compared to those of 4 cases of biopsy-proven schwannoma. Aspirate smears were evaluated for cellular cohesion, cell type and stroma. Immunoperoxidase stains were utilized in select cases. RESULTS: The cases of nodular fasciitis exhibited cohesive clusters of epithelioid to spindle-shaped cells in a background of single, intact mesenchymal cells; inflammatory cells; and myxoid stroma. In contrast, schwannomas lacked single, intact cells and inflammation. Schwannoma stroma was also myxoid but appeared more finely fibrillar, and cell clusters were notable for alternating areas of hypercellularity and hypocellularity. Immunoperoxidase stains demonstrated smooth muscle actin reactivity in 5 cases of nodular fasciitis and S-100 in 2 cases of schwannoma. CONCLUSION: Nodular fasciitis can be distinguished from schwannomas on the basis of cytomorphologic features and immunocytochemical profile. Cytologic diagnosis of nodular fasciitis is important since it obviates the need for surgical excision. PMID- 15296336 TI - Fine needle aspiration cytology of seromas of the breast from irradiated lumpectomy sites. AB - OBJECTIVE: To study the cytologic features of cells in breast aspirates of seromas in women who had undergone irradiation at lumpectomy sites. STUDY DESIGN: Cytologic material from 38 females with an age range of 39-72 years who had undergone aspiration of seromas were retrieved to determine tumor recurrence or other atypia. The material was obtained using the conventional method of needle aspiration with a 22-gauge needle and 10-mL syringe, maintaining negative pressure. The syringe and needle contents were washed in a cytology container containing 30% ethyl alcohol in physiologic saline. From half the washings filter preparations were made and stained by the Papanicolaou method, while from the other half cell blocks were made and sections cut after processing and stained with hematoxylineosin. RESULTS: In 2 of 38 patients, filter preparations and cell blocks showed malignant cells of breast carcinoma that were similar to the primary. This was confirmed on core biopsy and subsequent mastectomy. In 21 of 38 cases mildly atypical cells with degenerative effects, mild hyperchromasia, liberal cytoplasm, macrophages, inflammatory cells and a few apocrine cells were seen, while in 11 of 38 patients scanty debris, few inflammatory cells, rare apocrine cells and a few macrophages were found. In 4 of 38 cases highly atypical cells with hyperchromatic nuclei and nucleoli were noted. However, a core biopsy in all 4 patients revealed no evidence of residual tumor. CONCLUSION: Cell atypia in aspirates from seromas should be interpreted with caution to avoid unnecessary, invasive surgery, keeping in mind that radiation can cause misleading changes. However, patients with clinical indications of recurrent tumor or with severe atypia should undergo biopsy to permit appropriate management. PMID- 15296337 TI - Criteria for the diagnosis of fibroepithelial lesions of the breast with liquid based cytology. AB - OBJECTIVE: To establish diagnostic criteria for diagnosing and differentiating fibroepithelial lesions of the breast on ThinPrep (Cytyc Corp., Boxborough, Massachusetts, U.S.A.). STUDY DESIGN: Eighty-four fibroepithelial lesions were sampled by ultrasound-guided aspiration biopsy. Based on smears and histologic correlates, there were 55 fibroadenomas, 26 papillary neoplasms and 3 phyllodes tumors. The ThinPrep slides for each sample were reviewed retrospectively and evaluated for specific morphologic and cytologic features. RESULTS: On ThinPrep slides, 95% of the fibroadenomas had a predominance of single myoepithelial nuclei, 89% had staghorn clusters, and 47% had myxoid stroma. Among the papillary neoplasms, 8% had a predominance of single columnar ductal cells, 31% had papillary groups, 23% had vessels, and 27% had collagenous spherulosis. The ThinPrep preparations of the phyllodes tumors showed that 67% had single myoepithelial nuclei, 33% had a predominance of single ductal cells, 67% had staghorn clusters, and 0% had myxoid stroma. A majority of the fibroadenomas and the papillary neoplasms showed mild to moderate ductal epithelial hyperplasia. A majority of the phyllodes tumors showed moderate ductal epithelial hyperplasia. CONCLUSION: Fibroepithelial lesions of the breast can be accurately differentiated using ThinPrep samples based on the evaluation of specific cytologic and morphologic features, including the presence of staghorn clusters, fibromyxoid stroma, vessels, collagenous spherulosis, papillary clusters and predominance of myoepithelial nuclei or columnar cells in the background. However, the degree of ductal epithelial hyperplasia does not aid in the diagnosis. PMID- 15296338 TI - Cytologic diagnosis of intraocular lymphoma in vitreous aspirates. AB - OBJECTIVE: To evaluate the cytologic findings of vitreous fluids with atypical, suspicious for malignancy or malignant lymphoid cells to assess cytologic parameters that may help in reaching the diagnosis of intraoclular lymphoma. STUDY DESIGN: Vitreous aspirates with a malignant, suspicious for malignancy or atypical lymphoid population were identified from the files of Barnes-Jewish Hospital during the previous 11 years. Cytologic preparations were reviewed. Pertinent clinical information was obtained from medical records. RESULTS: Thirteen vitreous aspirates from 12 patients were included. The chief complaints included floaters, blurred vision and decreased visual aculity. Bilateral ocular involvement was present in 8 (67%) patients. Three patients had a history of an extraocular lymphoid malignancy. All patients underwent pars plana vitrectomy and collection of the vitreous aspirate. Cytologic diagnoses included: malignant lymphoma (9 of 13), suspicious for malignant lymphoma (3 of 13) and atypical lymphoid population (1 of 13). Most samples had high cellularity (11 of 13) and necrosis (9 of 13). Abnormal lymphoid cells were large (2-4 times the size of a lymphocyte) and had a high nuclear/cytoplasmic ratio, prominent nucleoli, irregular nuclear contours and a fine to coarse chromatin pattern. All cases with malignant cytology had abundant abnormal lymphoid cells; inconclusive cases had few. Immunocytochemistry for CD20 and CD45RO was performed on 9 of 13 samples and was conclusive in 6 of 9. CONCLUSION: Cytologic analysis of vitreous aspirates can be useful in diagnosing intraocular involvement by malignant lymphoma. Sparse cellularity is the main factor leading to inconclusive diagnoses. Immunostaining can be useful in confirming the lymphoid nature of the malignant cells. PMID- 15296339 TI - Unusual cytologic findings in low grade papillary transitional cell carcinoma. AB - OBJECTIVE: To describe cases of low grade papillary transitional cell carcinoma (LG-pTCC) with a low nuclear cytoplasmic (N/C) ratio and unusual cytologic patterns with many isolated, single neoplastic cells. STUDY DESIGN: We defined the following unusual cytologic findings as "isolated, single cell pattern": (1) numerous single cells sometimes with a few flat cell clusters; (2) very low N/C ratio; (3) angulation of cytoplasmic contour; (4) pale, homogeneous cytoplasm; (5) hyperchromatic nuclei with an uneven contour; (6) monotonous cytologic appearance; and (7) clear background. We studied 2,956 cytologic specimens of voided urine from 114 LG-pTCC patients at our university hospital during a 10 year period. RESULTS: Thirty-six specimens had the isolated, single cell pattern. The isolated, single cell pattern showed less celllular atypia than does the typical pattern of LG-pTCC. On histology the cases with the isolated, single cell pattern showed a papillary structure with an erosive surface and were composed of mildly atypical neoplastic cells with very low N/C ratios. CONCLUSION: Some LG pTCCs show many single, atypical transitional cells. PMID- 15296340 TI - Reactive type II pneumocytes in bronchoalveolar lavage fluid. AB - OBJECTIVE: To evaluate the prevalence of reactive type II pneumocytes (RPII) in bronchoalveolar lavage (BAL) fluid samples obtained from patients with various pulmonary disorders. STUDY DESIGN: Consecutive BAL fluid samples were screened for the presence of RPII on May-Grunwald-Giemsa-stained cytocentrifuge preparations. BAL fluid samples with and without RPII were compared with regard to prevalence, associated clinical diagnoses and cytologic findings. RESULTS: RPII were generally large cells with a high nuclear:cytoplasmic ratio and deeply blue-stained, vacuolated cytoplasm. Most RPII occurred in cohesive cell groups, and the vacuoles tended to be confluent. Cytologic findings associated with RPII were foamy alveolar macrophages, activated lymphocytes and plasma cells. RPII were present in 94 (21.7%) of 433 included BAL fluid samples. The highest prevalences were noted in patients with systemic inflammatory response syndrome and alveolar hemorrhage. In addition, RPII tended to occur more frequently in ventilator-associated pneumonia, Pneumocystis carinii pneumonia, extrinsic allergic alveolitis and drug-induced pulmonary disorders. In contrast, RPII were not observed in BAL fluid samples obtained from patients with sarcoidosis. CONCLUSION: RPII were prevalent in about 20% of BAL fluid specimens. They were associated mainly with conditions of acute lung injury and not observed in sarcoidosis. PMID- 15296341 TI - Analysis of nuclear chromatin distribution in cervical glandular abnormalities. AB - OBJECTIVE: To measure the intensity of hematoxylin staining for the analysis of chromatin distribution and to define a clear set of standards. STUDY DESIGN: Cervical smears obtained from 12 patients with glandular lesions, 5 with squamous lesions and 3 without cervical lesions were used for NIH image analysis (public domain NIH image program developed at the U.S. National Institute of Health, available through the Internet by anonymous ftp from zippy.nimh.nih.gov or on floppy disk from the National Technical Information Service, Springfield, Virginia). In addition, the same cervical smears were restained with propidium iodide, and the DNA content in the nuclei was compared with that with hematoxylin staining. RESULTS: Chromatin distribution was categorized into 3 patterns. Pattern A was that for which the highest staining density was localized in the periphery of the nucleus, while in pattern C it was localized in the center of the nucleus. Pattern B was the intermediate type between patterns A and C. In patients with adenocarcinoma, pattern B was predominant; pattern C was also relatively frequent in this group. In atypical glandular cells observed in patients with squamous lesions, patterns A and B were predominant and pattern C rarely seen. Analysis of DNA content in the nucleus revealed that nuclei showing pattern B contained significantly higher quantities of DNA than those showing pattern A. CONCLUSION: Nuclear chromatin distribution seems to correlate well with DNA content, and analysis of the chromatin distribution pattern is helpful for the diagnosis of cervical glandular neoplasia. PMID- 15296342 TI - Hybrid capture II and polymerase chain reaction for identifying HPV infections in samples collected in a new collection medium: a comparison. AB - OBJECTIVE: To compare the performance of human papillomavirus (HPV) DNA detection by polymerase chain reaction (PCR) and Hybrid Capture II (HCII) test (Digene, Gaithersburg, Maryland, U.S.A.) in residual cells left in the collection vials of the DNACitoliq system (Digene Brasil, Sao Paulo, Brazil). STUDY DESIGN: A series of 263 cervical samples collected for liquid-based cytology with the DNACitoliq system was tested for oncogenic HPV types first with HCII and subsequently with PCR. After DNA purification with GFX Genomic Blood DNA Purification Kit (Amersham, Piscataway, New Jersey, U.S.A.), PCR was performed using AmpliTaq Gold DNA polymerase (Applied Biosystems). PGMY09/11 L1 consensus primers and GH20/PCO4 primers for human beta-globin target were coamplified. RESULTS: Altogether, 260 samples were positive for beta-globin, and 3 negative ones were excluded from the analysis. PCR and HCII yielded concordant results in 199 cases (76.5%) (102 positive and 97 negative), with Cohen's kappa of .577 (95% CI .477-.677) and weighted kappa of .733 (95% CI .659-.791). HPV prevalence in different categories of cytologic abnormalities was practically identical with HCII and PCR assays (P=.989). Among the 61 (23.5%) discrepant cases, 28 samples were HCII+/PCR- cases. Of these, 27 of 28 samples showed a low viral load, and 1 had an intermediate viral load. CONCLUSION: The data suggest that residual material from the DNACitoliq system adequately preserves HPV DNA for detection by HCII and PCR, with performance similar to that of specimen transport medium. PMID- 15296343 TI - Nongynecologic cytology practice guideline. PMID- 15296344 TI - Cytopathologic and genetic diagnosis of pulmonary amebiasis: a case report. AB - BACKGROUND: Amebiasis is a parasitic infection with Entamoeba histolytica. Pulmonary amebiasis is rare since the infection is commonly manifested as amebic colitis or liver abscess. Most pleuropulmonary amebiasis is seen in patients with amebic liver abscesses. A pulmonary amebic lesion without either a liver abscess or amebic colitis is extremely rare. Thus, reported cases of sputum cytologic diagnosis of a pulmonary amebic lesion from a patient without a liver abscess are also very rare. CASE: A 53-year-old man presented with a dry cough and mild fever. Chest radiography revealed an abnormal solitary mass lesion in the right upper lung field. The clinical diagnosis was a bacterial lung abscess. Sputum cytologic examination demonstrated many trophozoites of E. histolytica. Following sputum cytodiagnosis, serologic tests revealed a slightly high but almost normal titer of IgG antibodies to E. histolytica, indicating the possible presence of the pathogen. Polymerase chain reaction (PCR) using E. histolytica-specific primers for DNA extracted from the sputum sample revealed specific DNA product. CONCLUSION: Pulmonary amebiasis without either a liver abscess or amebic colitis must be distinguished from bacterial abscesses and neoplastic disease. A sputum cytologic examination combined with PCR for DNA extracted from a sputum sample is a good approach to the diagnosis of a pulmonary amebic abscess. PMID- 15296345 TI - Fine needle aspiration cytology of a sebaceous lymphadenoma: a case report. AB - BACKGROUND: Sebaceous lymphadenoma is a rare, benign neoplasm, histologically characterized by proliferating islands of epithelium with sebaceous glandular differentiation in a dense, lymphocytic background. The parotid gland is the most common site, and the patient usually presents with a well-circumscribed, enlarging and painless mass. Primary sebaceous lesions of the salivary glands are very rare entities and must be differentiated from more common, potentially malignant tumors. CASE: A 75-year-old male presented with a 6-month history of a mass in the tail of the parotid gland. The mass was not fixed or tender to palpation, was well delineated and measured 4 cm in greatest dimension. Fine needle aspiration (FNA) revealed a mixed population of large and small lymphocytes, including plasma cells and occasional tingible body macrophages. Scattered among the lymphocytes were 3-dimensional, cohesive aggregates of epithelial cells, many demonstrating the characteristic cytoplasmic vacuolization of sebocytes, surrounded by layers of basaloid cells. No mitoses or cellular pleomorphism was identified. These findings suggested a sebaceous lymphadenoma, confirmed on biopsy. CONCLUSION: Although sebaceous lymphadenoma is encountered infrequently, FNA findings can result in its accurate diagnosis. PMID- 15296346 TI - Epithelioid hemangioendothelioma of the lung diagnosed by transesophageal endoscopic ultrasound-guided fine needle aspiration: a case report. AB - BACKGROUND: Epithelioid hemangioendothelioma is a rare vascular tumor of the lung and is also known as intravascular sclerosing bronchoalveolar tumor. Although it has relatively low malignant potential, extensive pulmonary involvement and systemic metastasis have been described. The cytologic features of these tumors are not very well defined, with only few case reports describing the cytologic findings of epithelioid hemangioendothelioma of the lung on fine needle aspiration. CASE: Endoscopic ultrasound-guided fine needle aspiration of a hilar mass was performed on a 25-year-old female. The cytology showed loosely cohesive sheets and clusters of epithelioid cells. The cellular features included large, irregular nuclei with nucleoli and a moderate amount of vacuolated cytoplasm. Rare cells had a suggestion of cytoplasmic lumen formation. Histologic examination of tissue fragments on the cell block revealed a tumor composed of rounded to spindled epithelioid cells in a background of light blue stroma. The endothelial differentiation was evidenced by cytoplasmic vacuoles and lumens, some of which contained erythrocytes. The endothelial nature of these cells was confirmed by positive staining with factor VIII and CD34. CONCLUSION: The cytomorphologic features of epithelioid hemangioendothelioma described in the literature and observed in our case are distinctive and can help with the interpretation of cytologic smears and prevent misdiagnosis. PMID- 15296347 TI - Fine needle aspiration cytology of clear cell carcinoma of the gallbladder with hepatic infiltration: a case report. AB - BACKGROUND: Clear cell carcinoma of the gallbladder (CCG) is an unusual histologic variant recognized in the World Health Organ ization classification of tumors of the gallbladder and extrahepatic bile ducts. Although the clinicopathologic features have been documented in a few reports, to our knowledge the cytologic findings have not been described before. We report the fine needle aspiration cytology (FNAC) findings in a case of CCG with hepatic infiltration. CASE: A 72-year-old woman presented with right upper quadrant pain and hepatomegaly. Serum levels of CA19-9 and alpha-fetoprotein were elevated. Computed tomography revealed several hepatic nodules, the larger of which was a mass in contact with the gallbladder, which had a thickened wall. FNAC showed loose sheets and disassociated cells with abundant, clear, finely vacuolated cytoplasm. Atypical bare nuclei, binucleated cells and some multinucleated cells were also found. A simultaneous trucut biopsy from the main hepatic mass confirmed the diagnosis. CONCLUSION: CCG is a clear cell neoplasm that should be considered when clear changes are observed on FNAC. Recognition of the cytologic features, together with adequate clinicoradiologic study, may be sufficient to establish the diagnosis. PMID- 15296348 TI - Intraoperative cytologic diagnosis of unsuspected cardiac myxoma: a case report. AB - BACKGROUND: Atrial myxoma is the most common primary intracardiac tumor. The diagnosis is generally primary intracardiac tumor, based on classical clinical findings coupled with echocardiographic or magnetic resonance image findings demonstrating a cardiac mass. CASE: Unsuspected atrial myxoma was found in a woman who had been diagnosed with invasive lobular carcinoma of the breast. The echocardiographic findings in the presence of fever favored a clinical working diagnosis of infective endocarditis complicating a suspected cardiac metastasis. While intraoperative frozen section examination could not rule out metastatic invasive lobular carcinoma, cytologic touch imprint findings were diagnostic of myxoma. This appears to be the first report of concurrent breast carcinoma and atrial myxoma. To our knowledge, this is also the first report of intraoperative cytologic diagnosis of cardiac myxoma. CONCLUSION: In myxoma cases with a complicated clinical setting in which frozen section examination may be equivocal, intraoperative cytologic examination may be a useful diagnostic tool. PMID- 15296349 TI - Sclerosing polycystic adenosis of the left parotid gland: report of a case with fine needle aspiration cytology. AB - BACKGROUND: Sclerosing polycystic adenosis (SPCA) of major salivary glands is a rare recently described entity. We report a case of SPCA of the left parotid gland, including the cytologic and histopathologic findings. CASE: A 20-year-old man presented with a left parotid mass that had been growing slowly for 3 years. Fine needle aspiration cytology showed many syncytial cell clusters of variable size and some ductal structures with an inflammatory background. The cells forming syncytial clusters were large and polygonal, with abundant, eosinophilic, granular or lacelike cytoplasm. Apocrine differentiation with decapitation secretion was commonly seen. The ductal cells had a relatively high nuclear/cytoplasmic ratio, with granular cytoplasm. Grossly, the 5-cm lesion was a discrete, pale, cystic nodule embedded within the parotid gland parenchyma. Microscopically, the lesion was a nonencapsulated, circumscribed mass of sclerotic and hyalinized, collagenous tissue with lymphoplasmacytic infiltration. Sclerosing adenosis and cystic ducts with frequent apocrinelike cells were commonly seen. Some acinar cells contained eosinophilic, intracytoplasmic granules of various sizes. CONCLUSION: The presence of syncytial clusters with apocrine metaplasia and ductal structures in a lymphoplasmacytic background should suggest a diagnosis of SPCA of a major salivary gland. PMID- 15296350 TI - Inflammatory pseudotumor of the parotid gland: report of a case with fine needle aspiration cytology. AB - BACKGROUND: Inflammatory pseudotumor is a rare lesion of the parotid gland. It usually presents as a mass lesion; thus, the clinical and radiologicfeatures often suggest malignancy. To the best of our knowledge, fine needle aspiration cytologic findings in parotid inflammatory pseudotumor have not been reported previously. CASE: A 59-year-old male presented with a palpable right parotid mass. Computed tomography revealed a mass measuring 2.5 cm in diameter. Fine needle aspiration cytology showed inflammatory cells, foamy histiocytes and groups of spindle-shaped cells without cytologic atypia. A diagnosis of inflammatory pseudotumor was suggested and was confirmed on histology. CONCLUSION: In the presence of a clinically evident mass in the parotid gland and fine needle aspiration cytologic features of inflammatory cells with sheets of spindle cells, the diagnosis of inflammatory pseudotumor should be suspected. The differential diagnosis of this unusual parotid gland lesion principally includes sialadenitis and myoepithelioma. PMID- 15296351 TI - Unsuspected peritoneal leishmaniasis in an HIV-positive woman with ovarian cancer. PMID- 15296352 TI - Fine needle aspiration in a case of gastric teratoma. PMID- 15296353 TI - Metastatic Merkel cell carcinoma diagnosed on a Pap smear. PMID- 15296354 TI - [All fungi are edible...]. PMID- 15296355 TI - [Folic acid antagonists pemetrexed]. PMID- 15296356 TI - [Galantamine, an acetylcholinesterase inhibitor with various actions]. PMID- 15296357 TI - [Laxatives]. PMID- 15296358 TI - [Pollen associated natural allergies. Cross reactions of allergens]. PMID- 15296359 TI - [Hormone therapy with climacteric--what remains?]. PMID- 15296360 TI - [Which sun blockers are allowed for small children?]. PMID- 15296361 TI - Malignant tumours of female genital tract in North Eastern Nigeria. AB - OBJECTIVE: To ascertain the pattern and frequency of malignant tumours of female genital tract in North Eastern Nigeria. DESIGN: A retrospective analysis of surgical biopsy materials. SETTING: University of Maiduguri Teaching Hospital, the only teaching hospital in the North Eastern region of Nigeria. SUBJECTS: Three hundred and eighty-two cases of female genital malignancies histologically confirmed between January 1st 1991 and December 31st 2000. RESULTS: The age range of patients whose specimens were received during the ten year period was three to eighty years. Mean age of presentation was 44.2 years, (SD +/- 13). Cancer of the uterine cervix was the most common (70.5%), followed by ovarian cancer (16.3%), then cancer involving the uterus (8.5%). There was a steep rise in reported cases within the period of study especially for cancer of the cervix. Ovarian tumours were the most common tumours in the teenage group. CONCLUSION: The high incidence of cancer of the uterine cervix and the early mean age of presentation of all malignancies underlies the importance of screening programmes and awareness campaign in our community. The study also provides the basis for further analysis of female genital malignancies. PMID- 15296362 TI - MMR controversy raises questions about publication ethics. PMID- 15296363 TI - Terminologia Anatomica: revised anatomical terminology. PMID- 15296364 TI - The effect of sesamoid mobilization, flexor hallucis strengthening, and gait training on reducing pain and restoring function in individuals with hallux limitus: a clinical trial. AB - STUDY DESIGN: Clinical trial. OBJECTIVE: To determine the effect of 2 conservative intervention approaches for functional hallux limitus. BACKGROUND: Metatarsophalangeal joint (MPJ) sprains are common and can result in long-term sequelae such as persistent pain and loss of range of motion (ROM) secondary to bony proliferation and articular degeneration. It is important to determine the most effective intervention for functional hallux limitus to decrease pain and restore function. METHODS AND MEASURES: Twenty individuals with first MPJ pain, loss of motion, and weakness participated in the study. All patients received whirlpool, ultrasound, first MPJ mobilizations, calf and hamstring stretching, marble pick-up exercise, cold packs, and electrical stimulation. Ten of the 20 patients (experimental group) also received sesamoid mobilizations, flexor hallucis strengthening exercises, and gait training. Treatment was provided 3 times a week for 4 weeks. Measurements of first MPJ extension ROM, flexor hallucis strength, and subjective pain level were performed on the first and last visits. RESULTS: Following the 12 therapy sessions, the experimental group achieved significantly greater MPJ extension ROM and flexor hallucis strength and had significantly lower pain levels as compared to the control group (P<.001). CONCLUSIONS: These results suggest that sesamoid mobilization, flexor hallucis strengthening, and gait training should be included in the plan of care when treating an individual with functional hallux limitus. PMID- 15296365 TI - Effect of heat modalities on hamstring length: a comparison of pneumatherm, moist heat pack, and a control. AB - STUDY DESIGN: Prospective, researcher-blinded, repeated-measures, randomized complete block design. OBJECTIVES: To compare the effects of a single treatment of Pneumatherm, moist heat pack, and a control treatment on hamstring muscle length. BACKGROUND: Traditionally, heating modalities have been used to facilitate increases in tissue length. The Pneumatherm has been developed over the past 20 years for use in the clinical treatment of a variety of musculoskeletal pathologies. However, there is no published evidence supporting the use of Pneumatherm for improving muscle length. SUBJECTS: Participants consisted of 30 healthy, college-age males taken from a convenience sampling from the University of Indianapolis student population. METHODS AND MEASURES: Participants received a 3-treatment sequence on consecutive days. Treatments involved applying the determined modality to the posterior thigh using standard treatment protocols. A hand-held dynamometer was used to establish a consistent passive measurement force to measure hamstring muscle length. RESULTS: A mixed model analysis of variance with pretest-posttest (3 pretest and 3 posttest measures) and treatment sequence of the modalities (6 sequences of Pneumatherm, moist heat, and control) was completed. The only significant effect was for pretest-posttest measures. Post hoc comparisons revealed that the Pneumatherm posttest value was significantly different from all other measures. There were no differences found between pretest scores and the moist heat and control posttest scores. CONCLUSIONS: Our results demonstrate that the Pneumatherm modality is an effective agent for increasing hamstring muscle length following a single 20 minute treatment. In this study, a significant gain in hamstring muscle length was not found following a 1-time treatment with moist heat. The Pneumatherm may be a good option when heat is used to assist in gaining flexibility of the hamstring musculature. PMID- 15296366 TI - Electromyographic analysis of the rotator cuff and deltoid musculature during common shoulder external rotation exercises. AB - STUDY DESIGN: Prospective single-group repeated-measures design. OBJECTIVES: To quantify electromyographic (EMG) muscle activity of the infraspinatus, teres minor, supraspinatus, posterior deltoid, and middle deltoid during exercises commonly used to strengthen the shoulder external rotators. BACKGROUND: Exercises to strengthen the external rotators are commonly prescribed in rehabilitation, but the amount of EMG activity of the infraspinatus, teres minor, supraspinatus, and deltoid during these exercises has not been thoroughly studied to determine which exercises would be most effective to achieve strength gains. METHODS AND MEASURES: EMG measured using intramuscular electrodes were analyzed in 10 healthy subjects during 7 shoulder exercises: prone horizontal abduction at 100 degrees of abduction and full external rotation (ER), prone ER at 90 degrees of abduction, standing ER at 90 degrees of abduction, standing ER in the scapular plane (45 degrees abduction, 30 degrees horizontal adduction), standing ER at 0 degrees of abduction, standing ER at 0 degrees of abduction with a towel roll, and sidelying ER at 0 degrees of abduction. The peak percentage of maximal voluntary isometric contraction (MVIC) for each muscle was compared among exercises using a 1-way repeated-measures analysis of variance (P<.05). RESULTS: EMG activity varied significantly among the 7 exercises. Sidelying ER produced the greatest amount of EMG activity for the infraspinatus (62% MVIC) and teres minor (67% MVIC). The greatest amount of activity of the supraspinatus (82% MVIC), middle deltoid (87% MVIC), and posterior deltoid (88% MVIC) was observed during prone horizontal abduction at 100 degrees with full ER. CONCLUSIONS: Results from this study provide initial information to develop rehabilitation programs. It also provides information helpful for the design and conduct of future studies. PMID- 15296367 TI - A comparison of human muscle temperature increases during 3-MHz continuous and pulsed ultrasound with equivalent temporal average intensities. AB - STUDY DESIGN: A repeated-measure crossover design was used. The independent variable was the type of ultrasound (pulsed or continuous) and the dependent variable was intramuscular temperature. OBJECTIVE: To compare changes in intramuscular temperature resulting from the use of pulsed ultrasound versus continuous ultrasound with an equivalent spatial average temporal average (SATA) intensity. BACKGROUND: There is a lack of research on the heat-generating capabilities of pulsed ultrasound within human muscle. METHODS AND MEASURES: The subjects were 16 healthy volunteers (mean age +/- SD, 21.3 +/- 2.5 years). Each subject was treated with pulsed ultrasound (3 MHz, 1.0 W/cm2, 50% duty cycle, for 10 minutes) and continuous ultrasound (3 MHz, 0.5 W/cm2, for 10 minutes) during a single testing session. Tissue temperature returned to baseline and stabilized between treatments and treatment order was randomized. Tissue temperature was measured every 30 seconds using a 26-gauge needle microprobe inserted at a depth of 2 cm in the left medial gastrocnemius muscle. Data were analyzed using a linear mixed model. RESULTS: Treatment with continuous ultrasound produced a mean (+/-SD) temperature increase of 2.8 degrees C +/- 0.8 degrees C above baseline. Treatment with pulsed ultrasound produced a mean (+/-SD) temperature increase of 2.8 degrees C +/- 0.7 degrees C above baseline. Statistical analysis revealed no significant differences in either the extent or rate of temperature increases between the 2 modes of ultrasound application. CONCLUSION: Pulsed ultrasound (3 MHz, 1.0 W/cm2, 50% duty cycle, for 10 minutes) produces similar intramuscular temperature increases as continuous ultrasound (3 MHz, 0.5 W/cm2, for 10 minutes) at a 2-cm depth in the human gastrocnemius. Spatial average temporal average intensity is an important consideration when selecting pulsed ultrasound parameters intended to deliver nonthermal effects. PMID- 15296368 TI - Shoulder strength and range of motion in female amateur-league tennis players. AB - STUDY DESIGN: Cross sectional. OBJECTIVES: To compare the strength and range of motion of the dominant and nondominant shoulders of adult female amateur tennis players. The secondary purpose of the study was to examine whether there were any differences in the observed relationships between women with a past history of shoulder pain and those with no history of shoulder pathology. BACKGROUND: Information on characteristics of the shoulder in amateur female tennis players is scarce, as research has concentrated on highly skilled or male players, despite the cumulative prevalence of shoulder pain in this group. METHODS AND MEASURES: Fifty-one female competitive, amateur tennis players (average age, 45 years) were tested bilaterally for shoulder internal/external rotation passive range of motion and isokinetic concentric strength. RESULTS: Shoulder range of motion and strength ratios were comparable between sides. In the dominant arm, the total rotational range of motion was 221 degrees, with an internal to external rotator peak torque ratio of 1.05. External rotator strength was significantly greater in the dominant arm of individuals with no history of pain. CONCLUSIONS: Range of motion and strength adaptations widely reported in highly skilled tennis players were not apparent in amateur female players. In assessment and management, clinicians should regard the amateur female tennis player as a separate entity from the highly skilled player. PMID- 15296369 TI - More on writing good case reports. PMID- 15296370 TI - I'll just take my toys and go home. PMID- 15296371 TI - Over-the-counter medication labels: are we ignoring the needs of blind consumers? PMID- 15296372 TI - Difficulty in designing unbiased studies of high-tech devices. PMID- 15296373 TI - Further response to Dr. Sheedy's article on PALs. PMID- 15296374 TI - So sue me: doctors without insurance. PMID- 15296375 TI - Pediatric use of topical ophthalmic drugs. AB - BACKGROUND: Changes in the policies of the Food and Drug Administration (FDA) have resulted in a rapid increase in the number of drugs now labeled for pediatric use. In topical ophthalmic drugs, there are pharmaceuticals approved for use in children in the treatment of allergy, inflammation, and bacterial and viral infection. Clinicians should anticipate that this trend will continue into the future. OVERVIEW: This article reviews the history of FDA oversight of drugs used to treat children. Recent shifts in policies and approach are emphasized. Pediatric issues associated with ophthalmic drug use are discussed, including important differences regarding dosage, instillation, distribution, systemic toxicity, and metabolism. A summary of topical ophthalmic drugs approved for pediatric use, including age considerations, is provided. CONCLUSIONS: Clinicians should be aware of the increasing number of topical ophthalmic drugs that are approved for pediatric use. Social, regulatory, and economic factors suggest this trend will continue. When prescribing these drugs for use in children, clinicians should remember to refer to details of labeling for Pediatric Use, including the age of the child for whom the drug is approved. PMID- 15296376 TI - Anterior scleritis: three case reports and a review of the literature. AB - BACKGROUND: Scleritis is a rare ocular condition involving vasculitis. It can result in severe ocular morbidity and has a high association with underlying systemic disease and mortality. Management can be challenging and under-diagnosis is a concern. Treatment usually requires systemic medications and co-management with a medical specialist in the appropriate field to manage any underlying systemic etiology. METHODS: Three cases are presented: idiopathic nodular scleritis with complete recovery; bilateral, diffuse anterior scleritis with rheumatoid arthritis; and scleromalacia perforans treated with chemotherapeutic agents and co-managed with a rheumatologist. A search of the English language literature is reported, also. RESULTS: The reported cases exemplify much of our knowledge regarding scleritis. The literature review focuses on the challenges of an accurate diagnosis and management. Management is challenging and, although standards exist, controversy remains. The literature discusses the relationship of scleritis to underlying systemic disease and the significant implication of this association. CONCLUSION: Knowledge of scleritis may aid in determining a timely and accurate diagnosis and treatment of both the ocular and any underlying systemic conditions, thus decreasing morbidity and mortality. PMID- 15296378 TI - High-tech optometry. PMID- 15296377 TI - Educating the educators: increasing grade-school teachers' ability to detect vision problems. AB - BACKGROUND: The State University of New York College of Optometry/University Optometric Center conducts vision screenings in various public schools in New York City. A study was devised to determine whether teachers' abilities to detect vision problems in their students could be enhanced. METHODS: Students of two different grade schools were screened, in 1998-1999 and again two years later. In 1998-1999, the week before each group of students was to be screened, teachers were asked to indicate which children were felt to have vision problems. In 2000 2001, prior to the screenings, a lecture on vision and its relationship to learning was given to the same teachers who had responded two years earlier, and handouts and brochures delineating vision problems were also given to each teacher. A comparison was then made of the teachers' ability to detect vision problems before and after the teacher education. RESULTS: In the first year, 111 of 377 of children screened (29%) were referred. Teachers correctly identified 39% of the acuity failures and 29% of the functional failures. Two years later, a total of 31% of children were referred. The same teachers correctly identified 68% of the acuity failures and 67% of the functional failures. CONCLUSIONS: There was a statistically significant increase in the ability of teachers to correctly identify children with learning problems based on education. Both the accuracy of identifying children with acuity problems, as well as identifying children with functional visual problems, increased. Based on these results, it is recommended that an in-service lecture be given to school teachers to heighten their awareness of vision problems that may impact learning performance. PMID- 15296379 TI - Ownership of patient records. PMID- 15296380 TI - Healthy eyes healthy people as a practice builder. PMID- 15296381 TI - Make sure to note correct place of service on claims. PMID- 15296382 TI - High performance solid-phase analytical derivatization of phenols for gas chromatography-mass spectrometry. AB - The solid-phase analytical derivatization of phenols with pentafluoropyridine is performed. Fourteen phenols including chlorophenols and alkylphenols, could be efficiently adsorbed on a strong anion-exchange solid phase, Oasis MAX. The phenols adsorbed on Oasis MAX as phenolate ions were desorbed after derivatization with pentafluoropyridine. After optimization of the adsorption and derivatization, we established a procedure for the determination of the phenols in water samples by means of GC-MS. Under the optimized conditions, calibration curves were linear in the range of 10-1000 ng/l for the alkylphenols (100-10000 ng/l for nonylphenol) and 50-1000 ng/l for the others. By processing 100 ml samples, the method detection limits (MDLs) were in the range of 0.45-2.3 ng/l for the alkylphenols (8.5 ng/l for nonylphenol) and 2.4-16 ng/l for the others. Compared with the biphasic reaction system, the signal-to-noise ratios obtained by the solid-phase analytical derivatization were significantly higher. This is ascribed to the fact that coexisting neutral and acidic compounds are efficiently removed from the sample solution by this solid-phase analytical derivatization system. PMID- 15296383 TI - Application of solid-phase microextraction for determining phenylurea herbicides and their homologous anilines from vegetables. AB - Residues of metobromuron, monolinuron and linuron herbicides and their aniline homologous were analyzed in carrots, onions and potatoes by solid-phase microextraction (SPME) performed with a polyacrylate fiber. A juice was obtained from food samples that were further diluted, and an aliquot was extracted after sodium chloride (14%) addition and pH control. At pH 4 only the phenylureas were extracted. A new extraction at pH 11 allowed the extraction of phenylureas plus homologous aniline metabolites. Determination was carried out by gas chromatography with nitrogen-phosporus detection (NPD) the identity of the determined compounds was studied by gas chromatography-mass spectrometry. Limits of quantification (LOQs) obtained with NPD and MS (selected-ion monitoring) were in the microg/kg order allowing determination of maximum residue levels (MRLs) established in the Spanish regulations. MRLs ranged from 0.02 to 0.1 mg/kg depending on the kind of food and herbicide. Under the proposed conditions matrix effects were low enough to permit calibration with samples proceeding from ecological (non-pesticide treated) crops. Twelve commercial samples of each carrots, onions and potatoes were analyzed and only three samples of potatoes contained residues of linuron at levels below MRLs. PMID- 15296384 TI - Direct zonal liquid chromatographic method for the kinetic study of actinomycin DNA binding. AB - The binding of an anticancer drug (actinomycin D or ACTD) to double-stranded DNA (dsDNA) was studied by means of high-performance liquid chromatography (HPLC). ACTD is an antitumor antibiotic containing one chromophore group and two pentapeptidic lactone cycles that binds dsDNA. Incubations of ACTD with DNA were performed at physiological pH. The complexed and free ligand concentrations of the mixture were quantified at 440 nm from their separation on a size-exclusion chromatographic (SEC) column using the same buffer for the elution and the sample incubation. The DNA and the ACTD-DNA complexes were eluted at the column exclusion volume while the ligand was retained on the support. An apparent binding curve was obtained by plotting the amount emerging at the exclusion column volume against that eluted at free ACTD retention volume. A dissociating effect was evidenced and the binding parameters were significantly different from those obtained at equilibrium by visible absorbance titration. The equilibrium binding parameters determined by absorption spectroscopy were used as starting data in the numerical simulations of the chromatographic process. The results showed a strong dependency of the apparent binding parameters on the reaction kinetics. Finally the comparison of the apparent binding curve obtained from the HPLC experiments and from the numerical simulations permitted an evaluation of the dissociation rate constant (kd = 0.004 s(-1)). PMID- 15296385 TI - Effect of temperature on the chromatographic retention of ionizable compounds. I. Methanol-water mobile phases. AB - The retention mechanism of acids and bases in reversed-phase liquid chromatography (RPLC) has been experimentally studied by examining the temperature dependence of retention, with emphasis on the role of the buffer ionization equilibria in the retention and selectivity. Retention factors of several ionizable compounds in a typical octadecylsilica column and using buffers dissolved in 50% (w/w) methanol as eluents at three temperatures in the range of 25-50 degrees C were measured. Two pairs of buffer solutions were prepared by a close adjusting of their pH at 25 degrees C; differences in their ionization enthalpies determined a different degree of ionization when temperature was raised and, as a consequence, a different shift in the eluent pH. Predictive equations of retention that take into account the temperature effect on both the transfer and the ionization processes are proposed. This study demonstrates the significant role that the selected buffer would have in retention and selectivity in RPLC at temperatures higher than 25 degrees C, particularly for co-eluted solutes. PMID- 15296386 TI - Column selection for liquid chromatographic estimation of the k'w hydrophobicity parameter. AB - Newer reversed-phase column technologies that incorporate polar groups either by an endcapping procedure or by embedding them into the stationary phase ligand have been receiving much attention in the literature for their robustness when highly aqueous conditions are used. We investigated their ability to accurately determine the chromatographic hydrophobicity value log k'w. The non-linear deviations of retention data as mobile phase conditions approach zero percent modifier are a large source of error when extrapolating to log k'w values using the linear solvent strength model. Here, we compare a conventional reversed-phase stationary phase with others that have incorporated either polar embedded or polar endcapped phases, along with a hybrid-based particle derivatized with a polar embedded ligand. Our results show that polar endcapped phases perform very similarly to the conventional phase and do not show any improved ability for determining log k'w, but polar embedded phases have reduced curvature in the data, and therefore result in less error in extrapolation. We also investigated the solubility parameter model and the [ET(30)] model for their extrapolation efficiency, and have concluded that the [ET(30)] model shows the least error when extrapolating the data. PMID- 15296387 TI - Thermally-treated clay as a stationary phase in liquid chromatography. AB - Spray-dried, spherical synthetic hectorite particles have been thermally-treated at 500 degrees C for 16 h and used as adsorbent materials in reversed-phase liquid chromatography. The retention of a 22 mono and disubstituted aromatic compounds was evaluated to study the retention mechanisms on the clay mineral. The retention of solutes on the thermally-treated clays was markedly different than that measured on octadecylsilica (ODS) columns under identical conditions, but remarkably similar to retention characteristics of the same solutes on porous graphitic carbon columns. The clay columns exhibit an enhanced selectivity over the ODS column in separation of nitroaromatic positional isomers. Under identical mobile phase compositions, a selectivity, alpha, of 7.15 between ortho- and para dinitrobenzene isomers was measured on the clay column compared to a alpha of 1.04 on the ODS column. PMID- 15296388 TI - High-performance liquid chromatographic enantioseparations on monolithic silica columns containing a covalently attached 3,5-dimethylphenylcarbamate derivative of cellulose. AB - Covalent immobilization of 3,5-dimethylphenylcarbamate derivative of cellulose was performed in situ onto native silica monoliths cladded in a 50 mm x 4.6 mm polyether ether ketone high-performance liquid chromatographic (HPLC) column. The covalent attachment of cellulose derivative in the range of 16-19% (w/w) was performed via an epoxide moiety. The column obtained by this technique combines the high enantiomer-resolving ability of the polysaccharide derivative with favourable dynamic properties of monolithic HPLC columns. The covalent attachment of the cellulose derivative enables this column to be used in combination with the mobile phases which are incompatible with coated-type polysaccharide columns due to solubility of chiral selector in some organic solvents. PMID- 15296389 TI - Study of short polystyrene monolith-fritted micro-liquid chromatography columns for analysis of neutral and basic compounds. AB - This study details the effects of poly(styrene-divinylbenzene) (PS-DVB) frits in micro-HPLC columns for the separation of neutral and basic compounds. The procedure comprised the optimization of separations with only monolith or conventional fritted columns followed by method transference to short monolith fritted columns. It was observed that a superior separation was achieved with the new columns compared to silica-fritted-packed columns once triethylamine (TEA) was added in small percentages. The separation of basic and neutral compounds was achieved in fast analysis times in the isocratic mode. PMID- 15296390 TI - Selection of reversed-phase liquid chromatographic columns with diverse selectivity towards the potential separation of impurities in drugs. AB - To select appropriate stationary phases from the continuously expanding supply of potentially suitable HPLC columns, the properties of 28 frequently applied stationary phases were determined by measuring several chromatographic parameters. From these results, based on chromatographic expertise, eight stationary phases with different properties and selectivities were selected. The aim of this study is to apply chemometric tools to evaluate the initially selected set of columns, i.e. a more systematic approach for making such a selection is examined. Starting from the information obtained on the 28 stationary phases, the re-evaluation was performed independently based on the chemometric techniques Pareto-optimality, principal component analysis (PCA), and Derringer's desirability functions. The aim was to select a set of efficient columns exhibiting large selectivity differences. The chemometrically selected stationary phases were divided in groups based on hydrophobicity, a critical retention-determining property in reversed-phase chromatography. This allowed to further reducing the selection to three columns. It is demonstrated that the selection by the chemometric approaches in general is fairly comparable with the initial selection. PMID- 15296391 TI - Enantioseparation of amino acid derivatives on an immobilized network polymer derived from L-tartaric acid. AB - Seven structurally related amino acid derivatives were successfully enantioseparated by HPLC with a commercially available column containing a chiral immobilized network polymer derived from L-tartaric acid. The experiments were carried out under normal-phase conditions. All the solutes could be baseline separated using n-hexane/2-propanol (95/5) as eluent at a flow rate of 1 ml/min at 25 degrees C, with reasonable retention time (<12 min). The effects of the polar alcohol modifier (type and content) in the mobile phase and the column temperature on the enantioseparation were studied. Apparent thermodynamic parameters were also calculated from the plots of ln alpha or ln k' versus 1/T. Some mechanistic aspects of chiral recognition were discussed with respect to the structures of the solutes. It was found that the enantioseparations are all enthalpy driven, and the N-acyl groups of the solutes have significant influence on the chiral recognition. PMID- 15296392 TI - Determination of alkylbenzene metabolites in groundwater by solid-phase extraction and liquid chromatography-tandem mass spectrometry. AB - Benzylsuccinate (BSA), methylbenzylsuccinate (methylBSA), and ethylbenzylsuccinate (ethylBSA) are unambiguous anaerobic biotransformation products from toluene, xylenes, and ethylbenzene decay, respectively, and may be used to indicate intrinsic bioremediation is occurring at hydrocarbon contaminated sites. In order to improve upon current methods that detect and quantify anaerobic hydrocarbon metabolites in field samples, solid-phase extraction (SPE) and direct sample injection methods coupled with liquid chromatography-tandem mass spectrometry (LC-MS-MS) were evaluated. In laboratory studies, recoveries of authentic standards of non-deuterated or deuterated benzylsuccinates and toluates ranged from 80 to 106% with relative standard errors ranging from 2 to 4%. The method detection limits for these analytes using SPE-LC-MS-MS ranged from 0.006 to 0.029 microg/L whereas those for direct injection-LC-MS-MS ranged from 0.61 to 1.5 microg/L. Given the increased sensitivity of using SPE coupled with LC-MS-MS, this technique was then used to analyze for the presence of putative anaerobic alkylbenzene metabolites in groundwater from a hydrocarbon-contaminated site where single-well push-pull tests were conducted using deuterated aromatic hydrocarbons. Both deuterated and non-deuterated benzylsuccinates and toluates were successfully detected and quantified in field samples using this method. PMID- 15296393 TI - High-performance liquid chromatography-ultrasonic nebulizer high-power nitrogen microwave-induced plasma mass spectrometry, real-time on-line coupling for selenium speciation analysis. AB - The coupling of a high-power nitrogen (N2) microwave-induced plasma (MIP) mass spectrometry--(MS) (1.3 kW) with high-performance liquid chromatography, connected with concentric nebulizer (CN), ultrasonic nebulizer (USN) and a hydride generation (HG) systems, for the optimization and determination of selenium compounds, has been carried out. The MIP-MS system fulfils the ideal requirement being an on-line real-time chromatographic detector for Se speciation analysis. Interchanging of MIP-MS system fabricated nebulizer (concentric) with an ultrasonic nebulizer increases about 3.4-12 (peak height) and 6.5-10 (peak area) times ion signals for the selenium compounds. The detection limits for selenate, selenite, trimethylselenonium ion (TmSe), selenomethionine (Semet) and selenoethionine (Seet) (in Milli-Q-water) obtained with the optimized HPLC-USN N2MIP-MS system are 0.11, 0.14, 0.09, 0.14 and 0.10 microg L(-1), respectively, about 12-48 times lower than the HPLC-CN-MIP-MS and 1.5-4.4 (peak height) times lower compared to the HPLC-CN-inductively coupled plasma (ICP)-MS coupling. Considering peak area, the repeatability (R.S.D. for three successive analyses) and intermediate precision (R.S.D. for three successive analyses performed on three different days), achieved for five Se compounds are 0.8-5.6, and 1.1-5.9%, comparable with the HPLC-CN-ICP-MS, HPLC-HG-MIP-MS and HPLC-CN-MIP-MS systems. The combined HPLC-USN-N2MIP-MS has been adequately applied for the determination of Se compounds in certified National Institute for Environmental studies human urine CRM No. 18. The results reasonably agree with the HPLC-CN-ICP-MS values. This encouraging combination may be an alternative ion source of mass spectrometry for coming generation in regard to the selenium speciation analysis. PMID- 15296394 TI - Application of ion-exchange cartridge clean-up in food analysis. VI. Determination of six penicillins in bovine tissues by liquid chromatography electrospray ionization tandem mass spectrometry. AB - A multiresidue analytical method was developed for the quantification of benzylpenicillin (PCG), phenoxymethylpenicillin (PCV), oxacillin (MPIPC), cloxacillin (MCIPC), nafcillin (NFPC) and dicloxacillin (MDIPC) in bovine tissues using liquid chromatography- electrospray ionization tandem mass spectrometry (LC ESI MS/MS) with a multiple reaction monitoring technique. Using the deuterated PCG and NFPC as internal standard was effective for improvement of repeatability and accuracy. We chose [M-H-141]- as a monitor ion of MRM analysis and [M-H]- as a precursor ion for each penicillin. Combination of an ion-exchange cartridge clean-up and ion-pair LC enable us to determine the residual penicillins using the standard curves made from standard solutions without the influence of sample matrix on the MS. The average recoveries of PCG, PCV, MPIPC, MCIPC, NFPC and MDIPC from bovine liver, kidney and muscle at the same concentrations as the tolerance levels of PCG (50 microg/kg) ranged from 77 to 101% with the coefficients of variation ranging from 0.7 to 4.2% (n = 5). The limits of quantification for the six penicillins were 2-10 microg/kg in bovine muscle, liver and kidney (S/N ratio >10). PMID- 15296395 TI - Simultaneous determination of fluoroquinolone, sulfonamide, and trimethoprim antibiotics in wastewater using tandem solid phase extraction and liquid chromatography-electrospray mass spectrometry. AB - A robust and sensitive method for the detection of fluoroquinolones, sulfonamides and trimethoprim has been developed. Wastewater samples were acidified and extracted through an anion-exchange cartridge in tandem with a hydrophilic lipophilic balance (HLB) cartridge, a procedure that reduced interferences from wastewater organic matter. The extracted antibiotics were analyzed using liquid chromatography electrospray mass spectrometry and selected ion monitoring. Quantification of antibiotics was assessed both by internal standard and standard addition methods. Average recoveries for a range of wastewater matrices were 37 to 129% for a 1 microg/L spiking concentration. The method detection limits (MDLs) of antibiotics in deionized water, final and secondary effluent ranged from 2 to 7 ng/L, from 20 to 50 ng/L, and from 30 to 90 ng/L, respectively. Assessment of matrix interference shows that signal suppression and MDL increases with higher amounts of organic matter in the sample. Analyses of samples from two municipal wastewater treatment plants indicate that ciprofloxacin, ofloxacin, sulfamethoxazole and trimethoprim are present in the secondary effluents at median concentrations of 100-160, 205-305, 395-575, and 40-705 ng/L, respectively. PMID- 15296396 TI - Development and in-house validation of a liquid chromatography-electrospray tandem mass spectrometry method for the simultaneous determination of Sudan I, Sudan II, Sudan III and Sudan IV in hot chilli products. AB - An accurate method based on the use of reversed-phase (RP) liquid chromatography tandem mass spectrometry interfaced with electrospray (LC-ESI-MS/MS) was devised for the determination of Sudan I, Sudan II, Sudan III and Sudan IV in hot chilli food samples. A simple sample treatment procedure entailing the use of an extraction step with acetone without clean-up was developed. A C18 column with an aqueous formic acid/methanol mixture as the mobile phase was used under isocratic conditions. Mass spectral acquisition was done in positive ion mode by applying selected reaction monitoring of three fragmentation transitions per compound to provide a high degree of selectivity. The method was in-house validated in terms of detection limits (LOD), quantitation limits (LOQ), linearity, sensitivity, accuracy, recovery, and selectivity on two kinds of hot chilli sauces. Good results in the low ng/g level were obtained for LOD and LOQ of all analytes in matrices. Under both intra-day repeatability (R.S.D. between 1 and 13%) and intermediate precision (about 5-15% R.S.D. for both chilli sauce matrices) conditions, precision proved to be typical of determinations based on electrospray LC-MS and acceptable for routine monitoring purposes. Extraction recoveries for all four azo-dyes in chilli tomato sauce ranged from 92 to 103% at a spiking level of 5 microg/kg, whereas values between 72 and 97% were calculated in chilli tomato and cheese sauce at the same concentration level. The applicability of the method to the determination of Sudan azo-dyes in hot chilli products was demonstrated. PMID- 15296397 TI - Determination of iodide in seawater and edible salt by microcolumn liquid chromatography with poly(ethylene glycol) stationary phase. AB - An ion chromatography method for rapid and direct determination of iodide in seawater and edible salt is reported. Separation was achieved using a laboratory made C30 packed column (100 mm x 0.32 mm i.d.) modified with poly(ethylene glycol) (PEG). Effects of eluent composition on retention behavior of inorganic anions have been investigated. Both cation and anion of the eluent affected the retention of analyte anions. The retention time of anions increased with increasing eluent concentration when lithium chloride, sodium chloride, potassium chloride, sodium sulfate, magnesium sulfate were used as the eluent, while it decreased with increasing eluent concentration when ammonium sulfate was used as the eluent. The detection limit for iodide obtained by injecting 0.2 microl of sample was 9 microg/l (S/N = 3). The present method was successfully applied to the rapid and direct determination of iodide in seawater and edible salt samples. Partition may be involved in the present separation mode. PMID- 15296398 TI - Effect of flow development region and fringing magnetic force field on annular split-flow thin fractionation. AB - Split-flow thin (SPLITT) fractionation devices have been widely used to separate macromolecules, colloids, cells and particles. Recently, the quadrupole magnetic flow sorter (QMS) has been reported in the literature as another family of SPLITT fractionation device. However, the separation performance observed in the experimental measurements is generally found to deviate from the ideal behaviour. Possible causes such as hydrodynamic lift force, high particle concentration and imperfect geometries have been extensively examined. However, the effects of flow development regions and fringing magnetic force field at the separation channel inlet and outlet, which are ignored by the theory, have not been investigated. The error introduced by ignoring these effects need to be rigorously studied so that the theory can be used to optimise operation flow rates with confidence. Indeed, we find in this paper that these ignored effects are responsible to the discrepancy between the experimental data and the theoretical predictions. A new theory has been proposed for optimisation of device operation. PMID- 15296399 TI - Tetramethyl-p,p'-sildiphenylene ether-dimethyl, diphenylsiloxane copolymers as stationary phases in gas chromatography. AB - A 33% tetramethyl-p,p'-sildiphenylene ether (SDPE)-67% dimethylsiloxane copolymer was prepared and characterized by 1H and 29Si NMR spectroscopy. The random copolymer was coated on fused-silica capillary columns and used as stationary phase in GC. Highly deactivated capillary columns with high separation efficiency and a working range from -10 to 400 degrees C were obtained. For comparison, a commercially available capillary column with an SDPE-diphenyl, dimethylsiloxane terpolymer was tested. The selectivity of SDPE phases is best described by the assumption that an SDPE unit is equivalent to two dimethylsiloxy and one diphenylsiloxy group. Both capillary columns exhibited low column bleed in combination with seriously increased elution temperatures. PMID- 15296400 TI - Determination of perfluorocarboxylic acids in aqueous matrices by ion-pair solid phase microextraction-in-port derivatization-gas chromatography-negative ion chemical ionization mass spectrometry. AB - A rapid, selective and simple analytical procedure using tetrabutylammonium as ion pair in conjunction with solid-phase microextraction followed by in-port derivatization-GC-negative ion chemical ionization mass spectrometry was developed. The procedure allows an accurate determination of perfluoroalkylcarboxylic acids in aqueous samples at ng L(-1) levels (i.e. method detection limit 20 ng L(-1) forperfluorodecanoic acid) improving previous GC methods in terms of analysis time and sensitivity. Ammonia as reagent gas in the negative ion chemical ionization mass spectrometry increased the sensitivity at least 3-fold compared to methane for perfluorocarboxylic acid butyl esters. The developed procedure was successfully applied to effluents from wastewater treatment plants (i.e. 0.05-8.2 microg L(-1)) and harbor seawaters. PMID- 15296401 TI - Two-phase partition chromatography using soybean oil eliminates pesticide residues in aqueous ginseng extract. AB - This study was carried out to develop a cost-, labor- and efficiency-effective elimination method of pesticide residues in ginseng extract. A two-phase partition method between soybean oil and distilled water or aqueous ginseng extract was employed for the elimination of pesticide residues. Content of the pesticides was determined by gas chromatography with electron capture or nitrogen phosphorus detector. A total of 15 pesticides representing four categories (organochlorine, organophosphorus, carbamate, pyrethroid) were spiked (ca. 2 ppm) to 2 ml soybean oil in a test tube and the oil was mixed with 6 ml distilled water or 10% aqueous ginseng extract. The test tubes were then vortexed (2 min) and centrifuged at 3000 rpm for 15 min to separate the oil and aqueous layers. Each layer was harvested and subjected to quantitative analysis of pesticides. The average distribution ratio of the pesticides to the oil layer was 94.4 +/- 6.7% in the mixture of the oil and distilled water, and 105.5 +/- 6.6% in the mixture of the oil and ginseng extract. No significant qualitative and quantitative change of ginsenosides, the active ingredients of Panax ginseng, was observed in the ginseng extract before and after the oil treatment. These results suggest that two-phase partition chromatography between soybean oil and the aqueous phase is a cost-, labor- and efficiency-effective reliable method for the elimination of pesticide residues in ginseng extract. PMID- 15296402 TI - Hold-and-flush, a novel fraction collection method in semi-preparative subcritical and supercritical fluid chromatography. AB - In supercritical fluid chromatography, the partial phase separation that occurs in the dead volume between automatic backpressure regulator and collection vessels causes significant peak tailing and delayed arrival of compounds to the collectors. As a result, when two peaks are barely baseline separated, it becomes very difficult to correctly set fraction collection triggers, which in turn results in fractions being collected with lower purity and lower yield. The problem can be solved with a simple addition of a four-port, two-position switching valve between UV detector and automatic back pressure regulator. The valve acts as a timed gate to release each peak out of the back pressure regulator and into the collection vessel, while at the same time it holds later peaks inside the UV detector and the column in a closed loop. PMID- 15296403 TI - Packed column supercritical fluid chromatography of a peroxysome proliferator activating receptor agonist drug. Achiral and chiral purity of substance, formulation assay and its enantiomeric purity. AB - This paper describes packed column supercritical fluid chromatography (SFC) for the analysis of a peroxysome proliferator-activating gamma-receptor agonist that is a carboxylic acid. Evaluation of conditions for the separation of this candidate drug and related compounds in bulk substance is described. A Chiralcel OD column was used for this purpose due to its high selective retention of related substances and relative inertness, though the enantioselectivity was negligible, with methanol as polar modifier. A high enantioselectivity was obtained on Chiralpak AD and it was possible to determine the enantiomeric purity within 10 min on a 5 cm short column. Both the achiral and the chiral systems were run without acid additive in the mobile phase and the level of detection of impurities by area was about 0.1%. For the analysis of samples dissolved in water, without any isolation step, 2-propanol was used as modifier. Due to the column surface activity, evidently generated by injected water, citric acid 1 mM was included as additive in the 2-propanol in order to maintain symmetric and undistorted peak shape. The detection limit for the assay was 21 microg mL(-1) (50 nmol mL(-1)) for 5 microL injected (R.S.D. 6.4%, n = 8). A 5 cm short Chiralcel OD column was used. Determination of enantiomeric purity of the drug in aqueous samples required increased sensitivity. The sample was acidified and extracted into a small volume of 1-pentanol, out of which 25 microl was analyzed by SFC. The minor enantiomer at the 3% (w/w) level added could be confirmed. Its ratio remained constant during the procedure as measured relative to a reference solution in organic media. PMID- 15296404 TI - Novel variable volume injector for performing sample introduction in a miniaturised isotachophoresis device. AB - A microdevice design furnished with a novel sample injector, capable of delivering variable volume samples, for miniaturised isotachophoretic separations is presented. Micromachining by direct milling was used to realise two flow channel network designs on poly(methyl methacrylate) chips. Both designs comprised a wide bore sample channel interfaced, via a short connection channel, to a narrow bore separation channel. Superior injection performance was observed with a connection channel angled at 45 degrees to the separation channel compared to a device using a channel angled at 90 degrees. Automated delivery of electrolytes to the microdevice was demonstrated with both hydrostatic pumping and syringe pumps; both gave reproducible sample injection. A range of different sampling strategies were investigated. Isotachophoretic separations of model analytes (metal ions and an anionic dye) demonstrated the potential of the device. Separations of ten metal cations were achieved in under 475 s. PMID- 15296405 TI - Investigation of the novel mixed-mode stationary phase for capillary electrochromatography. I. Preparation and characterization of sulfonated naphthalimido-modified silyl silica gel. AB - A novel packing material, 3-(4-sulfo-1,8-naphthalimido)propyl-modified silyl silica gel (SNAIP), was prepared for the use as a stationary phase of capillary electrochromatography (CEC). The sulfonic acid groups on SNAIP stationary phase contributed to the generation of electroosmotic flow (EOF) at low pH and served as a strong cation-exchanger. In CEC with SNAIP, a mixed-mode separation was predicted, comprising hydrophobic and electrostatic interactions as well as electrophoretic migration process. In order to understand the retention mechanism on SNAIP, effects of buffer pH, concentration, and mobile phase composition on EOF mobility and the retention factors of barbiturates and benzodiazepines were systematically investigated. Moreover, the retention behavior of barbiturates on SNAIP was investigated and compared with those on octadecyl silica (ODS), phenyl bonded silica, and 3-(1,8-naphthalimido)propyl-modified silyl silica gel to confirm the presence of pi-pi interaction on its retention mechanism. It was observed that a column efficiency was more than 85,000 N/m for retained compounds and the relative standard deviations for the retention times of EOF marker, thiourea, and five barbiturates were below 2.5% (n = 4). Under an applied voltage of 20 kV and a mobile phase consisted of 5 mM phosphate (pH 3.8) and 40% methanol, the baseline separation of five barbiturates was achieved within 3 min. PMID- 15296406 TI - Micellar electrokinetic chromatography at low pH with polyelectrolyte-coated capillaries. AB - The performance of capillaries coated with a poly(diallyldimethylammonium) (PDADMA) monolayer or poly(diallyldimethylammonium)-poly(styrenesulfonate) bilayer was investigated and compared under micellar electrokinetic chromatographic (MEKC) conditions. Both monolayer (positively charged) and bilayer (negatively charged) coatings with micellar (sodium dodecyl sulfate) electrolyte generated very stable and pH-independent cathodal electroosmotic flow (EOF). From the results obtained, it can be concluded that in a doubly coated capillary the second poly(styrenesulfonate) layer is replaced by sodium dodecyl sulfate micelles during flushing with micellar electrolyte. Consequently, in order to obtain a stable and pH-independent cathodal electroosmotic flow for the MEKC separations, the capillary coating with the second polyanion layer is not necessary. The importance of the PDADMA coating was illustrated by comparing MEKC separations of the common developing agents (hydroquinone, phenidone, pyrocatechol, pyrogallol and quinone) on a bare uncoated capillary with the coated capillary. The coating provides reproducible MEKC separations at low pH (pH 3.0) with relative standard deviation (R.S.D.) values for migration times and peak areas lower than 0.45 and 3.3%, respectively. Good linearities in the range from 5 x 10(-5) to 2 x 10(-3) mol l(-1) were obtained for all five compounds, with correlation coefficients higher than 0.998. The detection limits were in the range from 5 x 10(-6) mol l(-1) for pyrocatechol to 2 x 10(-5) mol l(-1) for quinone. The proposed MEKC system was applied to the determination of hydroquinone and phenidone in X-ray photographic developer solutions. PMID- 15296407 TI - Determination of liquiritigenin and isoliquiritigenin in Glycyrrhiza uralensis and its medicinal preparations by capillary electrophoresis with electrochemical detection. AB - A simple, reliable, reproducible and sensitive method, based on capillary electrophoresis with electrochemical detection (ED), for the determination of liquiritigenin and isoliquiritigenin in Glycyrrhiza uralensis and its medicinal preparations was described. Operated in a wall-jet configuration, a 300 microm diameter carbon-disk electrode was used as the working electrode, which exhibits good responses at + 1000 mV (versus SCE) for the two analytes. Under the optimum conditions, the analytes were base-line separated within 8 min, and excellent linearity was obtained in the concentration range from 5.0 x 10(-4) to 1.0 x 10( 6) mol/l. The detection limit (S/N = 3) was 4.7 x 10(-7) and 2.9 x 10(-7) mol/l for liquiritigenin and isoliquiritigenin, respectively. This work provides a useful method for the analysis of traditional Chinese medicines. PMID- 15296408 TI - Trace analysis of methyl tert-butyl ether in water samples using headspace solvent microextraction and gas chromatography-flame ionization detection. AB - In this study, a simple, rapid and efficient method for the extraction and determination of MTBE in water samples by the headspace solvent microextraction (HSME) and gas chromatography at sub (microg/l) level is described. Some significant variables such as type of solvent, extraction time, salt concentration, sample and microdrop volumes, stirring rate, sample and microsyringe needle temperatures were optimized. Using optimum extraction conditions (benzyl alcohol as extracting solvent, 4 M NaCl, sample temperature 35 degrees C, sample volume 6 ml, stirring rate 1000 rpm, microsyring needle temperature -6 degrees C, extraction time 7.5 min and micro drop volume of 2 microl) a detection limit of 0.06 microg/l and a good linearity (R2 > 0.999) in a calibration range of 0.1-500 microg/l were achieved. This HSME method was applied to the analysis of MTBE in tap, well and spring waters and a groundwater sample contaminated by leaking gasoline from an underground storage tank (LUST) in a gasoline service station. PMID- 15296409 TI - Selective determination of trimethylamine in air by liquid chromatography using solid phase extraction cartridges for sampling. AB - The selective determination of trimethylamine (TMA) in air by liquid chromatography is reported. Sampling is effected by flushing air through C18 packed solid-phase extraction (SPE) cartridges at a flow rate of 15 mL/min for 15 min. Next, TMA is desorbed from the cartridges and injected into the chromatographic system. The analyte is then selectively retained on a precolumn (20 mm x 2.1 mm i.d., packed with 30 microm, Hypersil C18 phase), and derivatized on-line by injecting 9-fluorenylmethyl chloroformate (FMOC). Finally, the TMA FMOC derivative is transferred to the analytical column (125 mm x 4 mm i.d., LiChrospher 100 RP18, 5 microm), and monitored at 262 nm. The method was applied to the measurement of TMA in air in the 0.25-2.5 microg interval (equivalent to concentrations of TMA of 1.1-11 mg/m3), providing good linearity, reproducibility and accuracy. The mean recovery of TMA was (96 +/- 7%) (n = 12), and the limit of detection was 0.05 microg. The proposed procedure allows the selective determination of TMA in the presence of other primary and secondary short-chain aliphatic amines. PMID- 15296410 TI - Reserach on newly emerging and reemerging infectious diseases. PMID- 15296411 TI - Highland fringe malaria and challenges in its control: the lesson from Akaki town. AB - Malaria is a major public health problem and of considerable socio-economic burden in most parts of Ethiopia. The country has witnessed recurrent epidemics of the disease, resulting in grave consequences including in areas designated as highland fringe. A study was undertaken to grossly assess the magnitude of the problems, the effectiveness of the control options and to explore the challenges encountered and the experiences gained during the 1998 malaria epidemic in Akaki Town and its environs. Health facility clinical records of individual patients and weekly surveillance and epidemic control reports were utilized as principal sources of data. The information revealed that the epidemic was very alarming affecting a sizable part of Akaki and the surrounding areas, with the total number of cases amounting to 622. The epidemic was controlled by case detection and treatment as well as by intensive vector control activities. The control endeavor, however, posed great difficulties due to the absence of systematic malaria control program, owing to underestimation of the threat from highland malaria. The contribution from the adjacent Oromia Malaria Control Program and The Federal Ministry of Health to control the epidemic (mainly vector control) was reckoned to be substantial. Thus, capacity building targeted to early detection, prevention and control of malaria epidemics, and preparedness is deemed to be of paramount importance. A viable Integrated Disease Surveillance and Response at health facilities could ensure early containment of the otherwise devastating epidemics. PMID- 15296412 TI - Severe malaria in the under-fives--clinical featrues, management and outcome in a district hospital. AB - A prospective hospital based study on severe malaria in under-five children was carried out over a period of one year in a district hospital of Ethiopia to determine the case fatality rate (CFR), factors contributing for high mortality, health seeking behaviour of the care takers, direct cost and feasibility of operating the WHO treatment guidelines at a district level. One hundred and one children aged between 6 and 59 months fulfilling the criteria of severe malaria have been recruited in the study and treated according to WHO guidelines. The most frequently encountered clinical manifestations of severe malaria were prostration and hypoglycaemia, the prevalence being 28.7% respectively. The over all case fatality rate of severe malaria was 11.9%. Cerebral malaria was the only isolated form of severe malaria with high CFR (OR=5.06, 95%CI 1.01-25.1). The hospital CFR of severe anaemia was 16.7%, which could have been reduced by provision of safe blood transfusion. Most of the children (80.2%) presented to the hospital in more than 24 hours after the onset of the illness. Forty seven percent of children received drugs at home and in 96% of the cases it was antimalarial drugs. The antimalarial drug treatment (dose/duration) was adequate in 71%. Children receiving appropriate anti-malaria treatment at home show a tendency towards a lower CFR. The total direct cost per disease episode ranged from USD 13.75 to 27.5. Eighty five percent of the direct cost was due to expenditures in the hospital. Implementation of the management protocol based on the WHO guidelines has required substantial input of resources. Major constraints noted were related to availability of safe blood andfollow-up after discharge. The study provides useful information for improved case management of severe malaria thereby reduce mortality of the under-five children due to severe malaria. PMID- 15296413 TI - Tuberculous meningitis in a district hospital from Southern Ethiopia. AB - In order to describe the clinical presentation, immediate outcome and risk factors associated with Tuberculous meningitis (TBM), 28 children with TBM were analyzed. The male to female ratio was 1.5:1. The mean age of the cases was 8 years (range 5 months-14 years). Nineteen (67%) of the patients were not vaccinated for Tuberculosis (TB). Nineteen (67%) patients had exposure to adults with pulmonary TB of which 14 (50%) were family members. Among these were four children who were vaccinated and their age range was from 7 month to 8 years. Thirteen (40%) were seen by health professionals with in three months before symptoms related to TBM, the mean duration of symptoms before seeking medical advice was 3.2 months. Using the weight height percentage of median, sixteen (57%) had malnutrition. Twenty (71%) patients were in stage three of TBM at presentation. Eleven (38%) had positive reaction to Mantoux test and 25 (89%) had abnormal chest radiography the most common finding being hilar lymphadenopathy. CSF (Cerebro Spinal Fluid) total cell count showed pleocytosis of < 200 in all but two cases (71%), and raised protein level on quantitative determination. Acid Fast Bacilli (AFB) test done in seven patients, was positive in two (29%) cases. One (4%) direct sputum smear and gastric aspirate culture were positive. Thirteen 13 (46%) patients died despite treatment and 9 (64%) had severe neurological complications. Delay at presentation advanced stage of TBM and unvaccinated state for tuberculosis were closely associated with poor outcome (P<0.05). Hence health workers who treat children should maintain high index of suspicion at all times and evaluate for TBM. The value of prophylaxis for children who have close contact with infectious cases should be evaluated in Ethiopia. PMID- 15296414 TI - Analysis and reporting of meningococcal meningitis epidemic in north Gondar 2001 2002. AB - Severe epidemic meningococcal meningitis occurs in countries of the meningitis belt of Sub-Saharan Africa, including Ethiopia. Many epidemics occurred in this country in the past decade, the recent being in 2000 in Addis Ababa unusually during the wet and rainy season. The study was a cross-sectional design. Data were collected with prepared questionnaires and the line listing. CSF (Cerebro Spinal Fluid) culture and antibiotic sensitivities were done for a limited number of patients. The objective of the study was to assess the progress and management outcome of the epidemic. During the out breaks of epidemic meningitis in north Gondar zone of the Amhara regional state in 2001 and 2002 children and young adults were most affected. There were 384 cases and 26 deaths in 2001 and 1235 cases and 128 deaths, in 2002, making a total of 1619 cases and 154 deaths with a case fatality rate of 9.5%. The etiologic agent was sero-group A. The most affected age group was 15-30 years. About 80% of the cases were in the age 30 years and below. Surveillance, epidemic preparedness, interventions and response were found to be inadequate. Selective vaccination was not effective in handling the epidemic. Efficient surveillance, local processing and use of data, regional laboratory support, multisectoral approach and mass vaccination were recommended to appropriately and timely handle such epidemics. PMID- 15296415 TI - Human helminthiasis in Wondo Genet, southern Ethiopia, with emphasis on geohelminthiasis. AB - A parasitological survey was made in 1999 and 2002 to determine the magnitude of geohelminthiasis (soil-transmitted helminthiasis) and generate reference baseline helminthological data for Wondo Genet area, southern Ethiopia. Stool specimens were collected using Kato technique from 3167 schoolchildren, 92 schoolteachers, and 1160 residents of two Peasant Associations (PAs) and microscopically examined. The prevalence of infection for Ascaris lumbricoides and Trichuris trichiura among schoolchildren was 83.4% and 86.4%, respectively, and the respective intensity of infection was 7343 eggs per gram of stool (EPG) and 461 EPG. The prevalence of infection for Ascaris lumbricoides, Trichuris trichiura, and Schistosoma mansoni among schoolteachers was 33.7%, 35.9% and 17.4%, respectively, with the respective intensity of 1089 EPG, 194 EPG and 89 EPG. Similarly, the prevalence of infection among the residents of Shesha Kekele and Wondo Wosha Peasant Associations for Ascaris lumbricoides, Trichuris trichiura, and Schistosoma mansoni was 77%, 88% and 30%, respectively, with the respective intensity of 4673 EPG, 452 EPG and 125 EPG. Both the prevalence and intensity of infection were low among the schoolteachers possibly due to a better personal hygiene. The most prevalent soil-transmitted helminth in the area was Trichuris trichiura. Other rare helminths encountered were Taenia species, hookworms, Enterobius vermicularis and Hymenolepis nana, all occurring in less than 4% prevalence of infection. The heavy helminth burden among the schoolchildren calls for immediate intervention to reduce morbidity and transmission of helminthiasis in Wondo Genet. PMID- 15296416 TI - Evaluation of antiretroviral treatment in two private medical centers in Addis, Ethiopia. AB - Ethiopia is one of the countries affected severely by the HIV/AIDS epidemic in sub-Saharan Africa. The disease is gradually fatal without antiretroviral therapy (ARV). The experience with antiretroviral therapy is limited in this country. This study is the first of its kind that has evaluated the status and implication of the clinical use of ARV drugs in Ethiopia. The guidelines for the initiation of treatment, the strengths and weaknesses of such treatment and the implications of similar intervention in a wider scale were assessed. Clinical records of 33 patients under follow-up and treatment with ARV in Hayat Hospital and Bethezatha Medical Center were identified. Data were collected from the medical records using a tool designed for the purpose. All 33 patients were receiving ARV for a mean duration of about 4 months. Twenty- three (69.7%) were males and 10 (30.3%) were females and the mean age was 38 years. Fifteen (45.5%) were married, fourteen (40.7%) were single, one (3%) divorced and there was no record for three (9.1%). Fifteen (45.5%) were businessmen, seven (21.2%) were private employees, four (12.1%) civil servants and in the remaining seven (21.2%) their occupation was not recorded. The decision to initiate ARV therapy was entirely based on the presence of symptomatic HIV disease. CD4+ lymphocyte count was done in 24 (72.7%) and basic hematological tests were done for all. However, biochemical investigations were incomplete in all of them. Triple therapy was started for all patients and the cost of the drugs was covered by their family in eleven (33.3%), by the patients in twenty-two (66.7%). Side effects of the drugs were noted in ten (30.3%) that resulted in change of the regimen in two and drug interruption in one. Response to treatment was entirely based on clinical parameters only. Twenty-three patients (69.7%) were noted to improve while nine (27.3%) remained the same and death occurred in one (3%). Standard triple therapy was used in all patients in this study and symptomatic HIV disease was the reason for the initiation of treatment. The improvement observed was not substantiated by immunological or virological parameters. Thus, the impacts of ARVs cannot be measured and the response documented in this study can be due to the effective treatment of opportunistic infections. Hence a standard protocol should be developed based on the minimum set-up required for the initiation of ARV therapy. PMID- 15296417 TI - Iatrogenic bile duct strictures: a review of 22 cases. AB - The incidence of iatrogenic bile duct strictures in Ethiopia appears to be increasing. Of 27 patients that sustained bile duct injuries at open cholecystectomy, admitted during May 1996 to December 2002, 22 cases of bile duct strictures are presented to evaluate outcome of treatment. The mean age was 40 years, 15 females. Twenty-one were referrals. The usual presenting features were biliary peritonitis and jaundice. The average time lapse between the original surgery and admission to hospital was eight months. About 73% had Bismuth grade III-IV strictures and all patients underwent Roux-en-Y hepatico-jejunostomy. Postoperatively, biliary-cutaneous fistula, recurrent ascending cholangitis and wound infection were observed frequently. The overall mortality rate was 13.6%. Bile duct injuries and strictures occur in young productive age groups. Prevention of the occurrence of bile duct injury and its progression to a devastating stricture reduces morbidity and mortality. PMID- 15296418 TI - Pseudo-Meig's syndrome: parasitic leiomyoma with ascites in a 52-year old lady. AB - There is a possibility that a pedunculated subserous leiomyoma loses its vascular connection with the uterus and gets new blood vessel supply from other pelvic or nearby structure and continues its parasitic growth. A 52-year-old lady presented with abdominal distension of 12 years duration and laparotomy was done with preoperative diagnosis of possible malignant ovarian tumour. Intraoperative finding was very big non-specific but well demarcated and easily excisable mass located partly in the utero-vesical pouch with blood supply entirely from the greater omentum and anterior abdominal wall. There was about 3600 ml clear ascitic fluid in the paracolic gutters. The mass was sent for histo-pathologic examination and the report was compatible with leiomyoma of uterine origin. Patient's recovery was quiet dramatic and subsequent out patient follow-ups were uneventful. The presence of ascites of different amount doesn't necessarily signify malignancy of any visceral primary or secondary origin. Therefore, benign tumours of ovarian or uterine origin should be included in the differential diagnosis of pelvic or abdominal mass with ascites to come up with scientifically sound deduction and intervention that may bring about remarkable impact on ameliorating mortality and morbidity. PMID- 15296419 TI - Cyclopia. AB - A deformity consisting of a single orbital fossa with absent globes with proboscis like structure on forehead is described in a 1200 gram female neonate who died immediately after birth. This neonate was born to a 35 years old Para VIII mother at term. Observations from clinical and post mortem examination are presented and literature review made. Because of its rare occurrence and tentative evidence for both genetic and environmental etiologies, reporting of cases of Cyclopia is encouraged. PMID- 15296420 TI - New location, military fighter pilots headline 2004 GHA summer meeting. PMID- 15296421 TI - Transforming Medicare and keeping the promise. PMID- 15296422 TI - IRS ruling provides guidance on joint ventures with for-profits. PMID- 15296423 TI - A new era begins on the Emerald Coast. PMID- 15296424 TI - Considerations for the recovery of recombinant proteins from plants. AB - The past 5 years have seen the commercialization of two recombinant protein products from transgenic plants, and many recombinant therapeutic proteins produced in plants are currently undergoing development. The emergence of plants as an alternative production host has brought new challenges and opportunities to downstream processing efforts. Plant hosts contain a unique set of matrix contaminants (proteins, oils, phenolic compounds, etc.) that must be removed during purification of the target protein. Furthermore, plant solids, which require early removal after extraction, are generally in higher concentration, wider in size range, and denser than traditional bacterial and mammalian cell culture debris. At the same time, there remains the desire to incorporate highly selective and integrative separation technologies (those capable of performing multiple tasks) during the purification process from plant material. The general plant processing and purification scheme consists of isolation of the plant tissue containing the recombinant protein, fractionation of the tissue along with particle size reduction, extraction of the target protein into an aqueous medium, clarification of the crude extract, and finally purification of the product. Each of these areas will be discussed here, focusing on what has been learned and where potential concerns remain. We also present details of how the choice of plant host, along with location within the plant for targeting the recombinant protein, can play an important role in the ultimate ease of recovery and the emergence of regulations governing plant hosts. Major emphasis is placed on three crops, canola, corn, and soy, with brief discussions of tobacco and rice. PMID- 15296425 TI - Metabolic carbon fluxes and biosynthesis of polyhydroxyalkanoates in Ralstonia eutropha on short chain fatty acids. AB - Short chain fatty acids such as acetic, propionic, and butyric acids can be synthesized into polyhydroxyalkanoates (PHAs) by Ralstonia eutropha. Metabolic carbon fluxes of the acids in living cells have significant effect on the yield, composition, and thermomechanical properties of PHA bioplastics. Based on the general knowledge of central metabolism pathways and the unusual metabolic pathways in R. eutropha, a metabolic network of 41 bioreactions is constructed to analyze the carbon fluxes on utilization of the short chain fatty acids. In fed batch cultures with constant feeding of acid media, carbon metabolism and distribution in R. eutropha were measured involving CO2, PHA biopolymers, and residual cell mass. As the cells underwent unsteady state metabolism and PHA biosynthesis under nitrogen-limited conditions, accumulative carbon balance was applied for pseudo-steady-state analysis of the metabolic carbon fluxes. Cofactor NADP/NADPH balanced between PHA synthesis and the C3/C4 pathway provided an independent constraint for solution of the underdetermined metabolic network. A major portion of propionyl-CoA was directed to pyruvate via the 2-methylcitrate cycle and further decarboxylated to acetyl-CoA. Only a small amount of propionate carbon (<15% carbon) was directly condensed with acetyl-CoA for 3 hydroxyvalerate. The ratio of glyoxylate shunt to TCA cycle varies from 0 to 0.25, depending on the intracellular acetyl-CoA level and acetic acid in the medium. Malate is the node of the C3/C4 pathway and TCA cycle and its decarboxylation to dehydrogenation ranges from 0.33 to 1.28 in response to the demands on NADPH and oxaloacetate for short chain fatty acids utilization. PMID- 15296426 TI - An innovative application of the "flexible" GRID/PCA computational method: study of differences in selectivity between PGAs from Escherichia coli and a Providentia rettgeri mutant. AB - The original GRID/PCA technique was adapted for the development of a tool potentially useful for the plan of a research strategy in rational enzyme design. The use of the MOVE directive of GRID made it possible to partially take into account protein flexibility, and the multivariate analysis was used as an instrument for focusing only on relevant information related to the differences in enzyme substrate selectivities. The comparison of two different penicillin G acylases, from Escherichia coli and from Providentia rettgeri, was used as a case study; these enzymes are very similar and their reported selectivities differ only for a couple of mutations around the active site. The "flexible" GRID/PCA method was able to correctly predict the observed selectivity differences caused not only by mutations of residues of the active site but also by long range effects on substrate selectivity due to sequence mutations on residues not directly involved in substrate recognition. PMID- 15296427 TI - Optimal process synthesis for the production of multiple recombinant proteins. AB - This paper presents a novel solution strategy for the synthesis of multiproduct and multihost protein production processes. There are several possible hosts that may express each of the products, and different downstream processing separation and purification tasks are needed, which in part depend on the host selection. Moreover, alternative unit operations may be available for some of these separation tasks. Finally, these processing units may be arranged in different configurations. A single mixed-integer optimization model represents the different decisions involved in synthesizing a plant for producing multiple proteins. The mathematical model optimizes the profit of the multiproduct plant and allows the decisions to be made simultaneously, namely, the choice of hosts, downstream operations, the configuration and size of units, as well as their scheduling. An example is solved for a plant that must produce four proteins for which there are alternative hosts for their expression (Escherichia coli, Chinese hamster ovary cells, and yeast that, depending on the product, may express it as an extracellular or intracellular protein) that require 15 stages with choices of unit operations as well as in or out of phase operations. Given the very large quantity of novel recombinant proteins for a number of novel therapeutic uses presently being approved or "in the pipeline", multiproduct and multihost recombinant protein production plants have recently been or are being built for the manufacture of these products. The strategy presented in this paper is of crucial value for the optimal utilization of such plants. PMID- 15296428 TI - Production of structured lipids by acidolysis of an EPA-enriched fish oil and caprylic acid in a packed bed reactor: analysis of three different operation modes. AB - Structured triacylglycerols (ST) enriched in eicosapentaenoic acid (EPA) in position 2 of the triacylglycerol (TAG) backbone were synthesized by acidolysis of a commercially available EPA-rich oil (EPAX4510, 40% EPA) and caprylic acid (CA), catalyzed by the 1,3-specific immobilized lipase Lipozyme IM. The reaction was carried out in a packed bed reactor (PBR) operating in two ways: (1) by recirculating the reaction mixture from the exit of the bed to the substrate reservoir (discontinuous mode) and (2) in continuous mode, directing the product mixture leaving the PBR to a product reservoir. By operating in these two ways and using a simple kinetic model, representative values for the apparent kinetic constants (kX) for each fatty acid (native, Li or odd, M) were obtained. The kinetic model assumes that the rate of incorporation of a fatty acid into TAG per amount of enzyme, rX (mole/(h g lipase)) is proportional to the extent of the deviation from the equilibrium for each fatty acid (i.e., the difference of concentration between the fatty acid in the triacylglycerol and the concentration of the same fatty acid in the triacylglycerol once the equilibrium of the acidolysis reaction is reached). The model allows comparing the two operating modes through the processing intensity, defined as mLt/(V[TG]0) and mL/(q[TG]0), for the discontinuous and continuous operation modes, respectively. In discontinuous mode, ST with 59.5% CA and 9.6% EPA were obtained. In contrast, a ST with 51% CA and 19.6% EPA were obtained when using the continuous operation mode. To enhance the CA incorporation when operating in continuous mode, a two step acidolysis reaction was performed (third operation mode). This continuous two-step process yields a ST with a 64% CA and a 15% EPA. Finally, after purifying the above ST in a preparative silica gel column, impregnated with boric acid, a ST with 66.9% CA and 19.6% EPA was obtained. The analysis by reverse phase and Ag+ liquid chromatography of the EPA-enriched ST demonstrated that the CA was placed in positions 1 and 3 and the EPA was occupying position 2 of the final ST. PMID- 15296429 TI - Effects of substrate pretreatment and water activity on lipase-catalyzed cellulose acetylation in organic media. AB - Lipase-catalyzed acetylation of cellulose solubilized in the dimethyl sulfoxide/paraformaldehyde organic solvent system was conducted with lipase A12 from Aspergillus niger. The accompanying side cellulase activity of the A. niger lipase partly accounted for the enhanced acetylation mediated by the enzyme, via facilitating the partial degradation of cellulose substrate as evidenced by high performance size exclusion chromatograph analysis. The enzymatic cellulose acetylation was improved by substrate pretreatment with cellulase or ultrasound by 18 and 14%, respectively, as a result of the reduced substrate molecular size. Additionally, the ultrasound-pretreated cellulose as the starting substrate was beneficial for the cellulose solution preparation due to the increased accessible surface of cellulose as evidenced by its increased sedimentation volume and SEM micrographs. The effect of thermodynamic water activity (aw) on lipase catalytic activity in organic media was also investigated. The maximum acetylation extent (nearly 11 wt %) occurred at aw = 0.52, which was improved by 51% relative to the enzymatic reaction with no control of water activity. The much larger extent to which the lipase-catalyzed cellulose acetylation was enhanced by water activity optimization than by substrate pretreatment further supported the predominant role played by the major lipase activity of the A. niger lipase over its side cellulase activity in catalyzing cellulose ester synthesis in organic media. PMID- 15296430 TI - Effect of oxygen limitation and medium composition on Escherichia coli fermentation in shake-flask cultures. AB - Shake-flask cultures are widely used for screening of high producing strains. To select suitable strains for production scale, cultivation parameters should be applied that provide optimal growth conditions. A novel method of measuring respiratory activity in shake-flask cultures was employed to analyze Escherichia coli fermentation under laboratory conditions. Our results suggest that the length of fermentation, choice of medium, and aeration do not normally satisfy the requirements for unlimited growth in shake flasks. Using glycerol rather than glucose as a carbon source greatly reduced the accumulation of overflow and fermentative metabolites when oxygen supply was unlimited. A rich buffered medium, Terrific Broth (TB), yielded 5 times more biomass compared to LB medium but also caused oxygen limitation in standard shake-flask cultures at shaking frequencies below 400 rpm. These results were used to optimize the production of benzoylformate decarboxylase from Pseudomonas putida in E. coli SG13009, resulting in a 10-fold increase in volumetric enzyme production. This example demonstrates how variation of medium composition and oxygen supply can be evaluated by the measurement of the respiratory activity. This can help to efficiently optimize screening conditions for E. coli. PMID- 15296431 TI - Biokinetic evaluation and modeling of continuous thiocyanate biodegradation by Klebsiella sp. AB - Biokinetics for autotrophic degradation of thiocyanate using batch culture of Klebsiella sp. were evaluated both analytically and numerically. A sequential approach with an analytical method followed by a numerical approximation was used to evaluate and to ensure the accuracy of the parameter estimation. The nonlinear least-squares method with a 95% confidence interval was employed. The growth conditions were maintained at pH 7 and 38 degrees C for all experiments. With an automated incubation and turbidity reader, a total of 16 different initial thiocyanate concentrations, ranging from 10 to 300 mg L(-1), were used to develop a kinetic expression of specific growth rate as a function of substrate concentration. The biodegradation of thiocyanate with Klebsiella sp. followed a substrate inhibition pattern. Three identical automated bioreactors with working volumes of 1.5 L, equipped with sterilizable sampling ports, were also used for the numerical approximation of the biokinetic parameters in batch mode. A fourth order Runge-Kutta method was used to approximate the substrate inhibition kinetics of the Klebsiella sp. utilizing thiocyanate. Although the kinetic coefficients estimated by analytical and numerical methods were not statistically different at a 0.05 alpha level, model responses of numerical approximation generated a better prediction of changes in thiocyanate and cell mass concentrations. The hypothetical maximum growth rate, micro m, half saturation coefficient, Ks, microbial yield coefficient, Y, cell mass decay rate coefficient, kd, and substrate inhibition coefficient, Ksi, were evaluated as being 0.62 +/- 0.05 d(-1), 85 +/- 8 mg SCN- L(-1), 0.076 +/- 0.011 mg cell mass (mg SCN)(-1), 0.03 +/- 0.002 d(-1), and 131 +/- 22 mg SCN- L(-1), respectively. The calculated maximal substrate concentration, Sm, and apparent maximum specific growth rate, micro'm, were 105.5 +/- 8.7 mg SCN- L(-1) and 0.24 +/- 0.01 d(-1), respectively. Using these estimated parameters, the theoretical performance of the continuous operation was also illustrated, which depicts the residual thiocyanate and Klebsiella sp. concentrations in the non-steady and steady states at different hydraulic retention times (HRTs). Assuming the influent concentration of 250 mg SCN- L(-1), the expected treatment efficiency ranged from 94.9% to 69.4% between 20 and 5 days HRT, respectively. Klebsiella sp. was expected to be washed out at 4.8 days HRT, thus resulting in no treatment of thiocyanate. PMID- 15296432 TI - Scale-up of centrifugal impeller bioreactor for hyperproduction of ginseng saponin and polysaccharide by high-density cultivation of panax notoginseng cells. AB - Scale-up of a novel centrifugal impeller bioreactor (CIB) was demonstrated for production of valuable plant-specific secondary metabolites by high-density cell cultures. Initial kLa was identified to be a key factor affecting cell growth and production of ginseng saponin and polysaccharide by high-density cultivation of Panax notoginseng cells in a 3-L CIB. A high level of ginseng saponin and polysaccharide production was obtained at an initial kLa value of 30.2 h(-1). A maximum dry cell weight (DW) and production titer of ginseng saponin and polysaccharide reached 22.0 +/- 0.3, 1.5 +/- 0.1, and 2.7 +/- 0.2 g/L on day 15 with their corresponding productivity of 1140 +/- 42, 81 +/- 8, and 150 +/- 17 mg/(L.d), respectively. Based on initial kLa level, the CIB high-cell-density cultivation process was successfully scaled up from 3 L to 30 L. A maximum DW and production titer of ginseng saponin and polysaccharide in a 30-L CIB reached 25.5 +/- 0.5, 1.7 +/- 0.1, and 2.9 +/- 0.1 g/L (on day 15) at an initial kLa value of 28.7 h(-1), respectively, and their corresponding productivity was 1340 +/- 56, 91 +/- 9, and 164 +/- 15 mg/(L.d). Furthermore, by adopting a fed-batch cultivation strategy, a maximum DW and concentrations of total saponin and polysaccharide in the 30-L CIB were enhanced to 30.3 +/- 1.0, 2.1 +/- 0.1, and 3.5 +/- 0.2 g/L with their corresponding productivity of 1467 +/- 87, 102 +/- 13, and 179 +/- 18 mg/(L.d), respectively. The work suggests that the CIB may have great potential in large-scale high-density plant cell cultures for efficient production of useful secondary metabolites. PMID- 15296433 TI - Characteristics of a methanotrophic culture in a membrane-aerated biofilm reactor. AB - The membrane-aerated biofilm reactor (MABR) shows considerable potential as a bioprocess that can exploit methanotrophic biodegradation and offers several advantages over both conventional biofilm reactors and suspended-cell processes. This work seeks primarily to investigate the oxidation efficiency in a methanotrophic MABR. A mixed methanotrophic biofilm was immobilized on an oxygen permeable silicone membrane in a single tube hollow fiber configuration. Under the conditions used the maximum oxygen uptake rate reached values of 16 g/m2.d, and the rate of biofilm growth achieved was 300 microm/d. Both indicators reflect a very high metabolic rate. It was shown that the biofilm was predominantly in a dual-substrate limitation regime but below about 250 microm was fully penetrated by both substrates. Oxygen limitation was not observed. Analysis indicated that microbial activity stratification was evident and the location of stratified layers of oxygen-consuming components of the consortium could be manipulated via the intramembrane oxygen pressure. The results confirm that an MABR can be employed to minimize substrate diffusion limitations in thick biofilms. PMID- 15296434 TI - Development of a novel whey beverage by fermentation with kefir granules. Effect of various treatments. AB - The development of a novel whey-based beverage with acceptable organoleptic properties is reported, where various treatments were studied. Kefir yeast immobilized on delignified cellulosic materials (DCM) or gluten pellets proved to accelerate whey fermentation significantly, with the latter support being not so preferable. Kefir granules seemed to achieve similar fermentation times as DCM. The final pH of the product is suggested to be 4.1 since the profile of the volatile byproducts was higher than other pH values tested. The addition of fructose seemed to be beneficial on the volatile content of the product, although its acceptability as determined by a preference panel was similar to that of the control. Finally, black raisin extract appeared to promote fermentation without any positive effect on the preference of the evaluators. PMID- 15296435 TI - A software tool to assist business-process decision-making in the biopharmaceutical industry. AB - Conventionally, software tools for the design of bioprocesses have provided only limited business-related information for decision-making. There is an industrial need to investigate manufacturing options and to gauge the impact of various decisions from economic as well as process perspectives. This paper describes the development and use of a tool to provide an assessment of whole flowsheets by capturing both process and business aspects. The tool is demonstrated by considering the issues concerned when making decisions between two potential flowsheets for a common product. A case study approach is used to compare the process and business benefits of a conventional process route employing packed chromatography beds and an alternative that uses expanded bed adsorption (EBA). The tool allows direct evaluation of the benefits of capital cost reduction and increased yield offered by EBA against penalties of using potentially more expensive EBA matrix with lower lifetimes. Furthermore, the tool provides the ability to gauge the process robustness of each flowsheet option. PMID- 15296436 TI - Separation of human serum albumin and human immunoglobulins using carrier phase ultrafiltration. AB - The fractionation of the plasma proteins human serum albumin (HSA) and human immunoglobulins (HIgG) using the combination of two newly developed techniques, pulsed sample injection technique and carrier phase ultrafiltration (CPUF), is discussed in this paper. The effects of pH and ionic strength on the transmission of a single protein (i.e., either HSA or HIgG) through 100 and 300 kDa MWCO polyethersulfone (PES) membranes were quantified using the pulsed sample injection technique. The experimental results thus obtained suggested that it would be possible to fractionate these proteins by optimizing the solution pH and ionic strength. With 100 and 300 kDa PES membranes, effective separation of HSA and HIgG was achieved by CPUF using suitable conditions, i.e., pH 4.7 and low salt concentration. The fractionation of HSA and HIgG by "reverse selectivity" using 300 kDa membranes was also examined. PMID- 15296437 TI - Effect of freezing rates and excipients on the infectivity of a live viral vaccine during lyophilization. AB - Lyophilization is the most popular method for achieving improved stability of labile biopharmaceuticals, but a significant fraction of product activity can be lost during processing due to stresses that occur in both the freezing and the drying stages. The effect of the freezing rate on the recovery of herpes simplex virus 2 (HSV-2) infectivity in the presence of varying concentrations of cryoprotectant excipients is reported here. The freezing conditions investigated were shelf cooling (223 K), quenching into slush nitrogen (SN2), and plunging into melting propane cooled in liquid nitrogen (LN2). The corresponding freezing rates were measured, and the ice crystal sizes formed within the samples were determined using scanning electron microscopy (SEM). The viral activity assay demonstrated the highest viral titer recovery for nitrogen cooling in the presence of low (0.25% w/v sucrose) excipient concentration. The loss of viral titer in the sample cooled by melting propane was consistently the highest among those results from the alternative cooling methods. However, this loss could be minimized by lyophilization at lower temperature and higher vacuum conditions. We suggest that this is due to a higher ratio of ice recrystallization for the sample cooled by melting propane during warming to the temperature at which freeze-drying was carried out, as smaller ice crystals readily enlarge during warming. Under the same freezing condition, a higher viral titer recovery was obtained with a formulation containing a higher concentration of sugar excipients. The reason was thought to be twofold. First, sugars stabilize membranes and proteins by hydrogen bonding to the polar residues of the biomolecules, working as a water substitute. Second, the concentrated sugar solution lowers the nucleation temperature of the water inside the virus membrane and prevents large ice crystal formation within both the virus and the external medium. PMID- 15296438 TI - Mass transfer studies of cell permeabilization and recovery of alkaline phosphatase from Escherichia coli by reverse micellar solutions. AB - Transfer of alkaline phosphatase (AP) directly from Escherichia coli cells into reverse micellar solutions (RMS) was studied by varying the water content, pH, and ionic strength of RMS. Prior to the mass transfer studies, the optimum conditions for the activity of alkaline phosphatase in reverse micellar solutions were determined. The maximum enzyme activity could be detected at higher pH and water content, indicating the ionization of p-nitrophenol to be crucial in the enzyme activity. The transfer of AP from E. coli followed a two-step process with most of the recovery being achieved in the first 3-10 min beyond which it slowed. A model suggesting separate locations for the enzyme in the cell wall has been proposed. PMID- 15296439 TI - High temperature increases the refolding yield of reduced lysozyme: implication for the productive process for folding. AB - Misfolding poses a serious problem in the biotechnological field in obtaining the active protein from inclusion bodies. Here we show that high temperature increases the refolding yield of reduced lyosyzme by a simple dilution method. The refolding yields at 98 degrees C were three times higher than those at 20 degrees C in the solutions tested, which is related to the fact that the thermally unfolded state of lysozyme is a more productive form for folding than the denaturant-induced fully unfolded state. The thermal-assisted refolding could be used for various reduced and denatured proteins as a result of its simplicity and low cost. PMID- 15296440 TI - Reversible and strong immobilization of proteins by ionic exchange on supports coated with sulfate-dextran. AB - New and strong ionic exchange resins have been prepared by the simple and rapid ionic adsorption of anionic polymers (sulfate-dextran) on porous supports activated with the opposite ionic group (DEAE/MANAE). Ionic exchange properties of such composites were strongly dependent on the size of the ionic polymers as well as on the conditions of the ionic coating of the solids with the ionic polymers (optimal conditions were 400 mg of sulfate-dextran 5000 kDa per gram of support). Around 80% of the proteins contained in crude extracts from Escherichia coli and Acetobacter turbidans could be adsorbed on these porous composites even at pH 7. This interaction was stronger than that using conventional carboxymethyl cellulose (CMC) and even others such as supports coated with aspartic-dextran polymer. By means of the sequential use of the new supports and supports coated with polyethyleneimine (PEI), all proteins from crude extracts could be immobilized. In fact, a large percentage (over 50%) could be immobilized on both supports. Finally, some industrially relevant enzymes (beta-galactosidases from Aspergillus oryzae, Kluyveromyces lactis, and Thermussp. strain T2, lipases from Candida antarctica A and B, Candida rugosa, Rhizomucor miehei, and Rhyzopus oryzae and bovine pancreas trypsin and chymotrypsin) have been immobilized on these supports with very high activity recoveries and immobilization rates. After enzyme inactivation, the protein could be fully desorbed from the support, and then the support could be reused for several cycles. Moreover, in some instances the enzyme stability was significantly improved, mainly in the presence of organic solvents, perhaps as a consequence of the highly hydrophilic microenvironment of the support. PMID- 15296441 TI - Shear-induced inactivation of alpha-amylase in a plain shear field. AB - A newly developed shearing device was used to study shear-induced inactivation of thermostable alpha-amylase in a plain shear field, under conditions comparable to extrusion. The results show that the inactivation can be described well with a first-order process, in which the inactivation energy largely depends on the shear stress, instead of specific mechanical energy or strain history. The resulting dependency of the rate of inactivation on the shear stress is very strong and nonlinear, which leads to the conclusion that in many cases the maximally applied shear stress determines the inactivation. Quantification of the inactivation rates gives design criteria for the application of enzymes in more viscous systems than conventionally used, provided that the reactor is designed such that no peak shear stresses occur. PMID- 15296442 TI - Toward a robust model of packing and scale-up for chromatographic beds. 1. Mechanical compression. AB - The packing of compressible biochromatographic resins at large scale suffers from a poor understanding of how column packing method, resin properties, and column geometry impact column performance. To improve understanding, we develop and evaluate a one-dimensional, continuum mechanics model of column packing by mechanical compression. We show that the model can quantitatively predict the change in bed height, applied stress, and internal axial porosity profile without adjustable parameters when the modulus and wall friction coefficients are determined independently. The model possesses theoretical relationships for wall support and resin rigidity that should enable it to describe the mechanical compression of any biochromatographic resin for any column diameter. Moreover, this framework could provide a path to analogous models for flow packing and dynamic axial compression. PMID- 15296443 TI - Toward a robust model of packing and scale-up for chromatographic beds. 2. Flow packing. AB - We developed and evaluated a model for predicting the flow packing of nonrigid chromatographic resins. The model is based on elasticity theory and accounts for resin rigidity and column diameter. When a modulus determined from a standard mechanical compression (consolidation) test is used, the model captures the primary phenomena of the scale-up process. However, moduli determined from flow packing experiments improve the accuracy of the predictions and show that the apparent rigidity of chromatographic resins is lower for flow packing than for mechanical compression. Using a modulus from flow-packing experiments provided quantitative scale-up predictions of flow packing carried out in columns with diameters between 200 and 450 mm at different locations and by different operators. PMID- 15296444 TI - A novel magnetic adsorbent for immunoglobulin-g purification in a magnetically stabilized fluidized bed. AB - A novel magnetic poly(ethylene glycol dimethacrylate-N-methacryloly-L histidinemethylester) [m-poly(EGDMA-(MAH)] support was prepared for purification of immunoglobulin G (IgG) in a magnetically stabilized fluidized bed by suspension polymerization. Elemental analysis of the magnetic beads for nitrogen was estimated as 70 micromol MAH/g polymer. Magnetic poly(EGDMA-MAH) beads were used in the separation of immunoglobulin-G (IgG) from aqueous solutions and/or human plasma in a magnetically stabilized fluidized bed system. IgG adsorption capacity of the beads decreased with an increase in the flow rate. The maximum IgG adsorption was observed at pH 6.0 for MES buffer. IgG adsorption onto the m poly(EGDMA) was negligible. Higher adsorption values (up to 262 mg/g) were obtained in which the m-poly(EGDMA-MAH) sorbents were used from aqueous solutions. Higher amounts of IgG were adsorbed from human plasma (up to 320 mg/g) with a purity of 87%. IgG molecules could be repeatedly adsorbed and desorbed with these sorbents without noticeable loss in their IgG adsorption capacity. PMID- 15296445 TI - Novel isoelectric precipitation of proteins in a pressurized carbon dioxide-water ethanol system. AB - A novel isoelectric precipitation of proteins in a pressurized carbon dioxide water-ethanol system was developed where carbon dioxide was used as a volatile acid. The pH-pressure curves of the system with the absence and presence of proteins were investigated. By introducing the pressurized carbon dioxide to a solution containing protein, the pH value in the solution was decreased to the isoelectric region of the model protein BSA. Addition of ethanol could lower the buffer capacity of the protein, which made the precipitation concentration of protein go beyond the limits in a system without ethanol and well exploited the application field of the technique. In addition, ethanol in solution played the role of aiding precipitation in the process. Another model protein, hen egg white lysozyme, was also studied but could not be precipitated in the above system. All of these phenomena prove that isoelectric precipitation is the key point in the pressurized carbon dioxide-water-ethanol system. PMID- 15296446 TI - Quantitative studies of cell-bubble interactions and cell damage at different pluronic F-68 and cell concentrations. AB - Pluronic F-68 (PF-68) is routinely used as a shear-protection additive in mammalian cell cultures. However, most previous studies of its shear protection mechanisms have typically been qualitative in nature and have not covered a wide range of PF-68 and cell concentrations. In this study, interactions between air bubbles along with the associated cell damage were investigated using the novel adenovirus-producing cell line PER.C6, a human embryonic retinoblast transfected with the adenovirus type 5 E1 gene. A wide range of PF-68 and cell concentrations (approximately 3 orders of magnitude) were used in these studies. At low PF-68 concentrations (0.001 g/L), cells had a very high affinity for bubbles, indicated by a more than 10-fold increase in cell concentration in the foam layer liquid versus the bulk liquid. At high PF-68 concentrations ( approximately 3 g/L), however, the cell concentration in the foam layer liquid was only approximately 40% of that in the bulk cell suspension. The number of cells associated with each bubble decreased from approximately 1000 cells at 0.001 g/L PF-68 to approximately 120 cells at 3 g/L PF-68. Despite the lower cell affinity for bubbles at a high PF-68 concentration, at high cell concentrations (10(7) cells/mL and 1 g/L PF-68) significant cell entrapment occurred in the foam layer, on the order of 1000 cells/bubble. For the cells carried by the bubbles, quantitative cell damage data revealed that the probability of cell death from bubble rupture was independent of bulk cell concentration but was affected by PF 68 concentration. These quantitative studies further indicated that even at a low PF-68 concentration of 0.03 g/L, approximately 30% of the attached cells were killed during the bubble rupture process. At the same time, at low PF-68 concentration (<0.1 g/L), significant cell death occurred prior to bubble rupture. On average, a bubble disrupted more cells in the bulk liquid and/or foam layer than during rupture. For both mechanisms, the number of cells damaged by each bubble increased with decreasing PF-68 concentration and increasing bulk cell concentration. PMID- 15296447 TI - Expression of functional human transferrin in stably transfected Drosophila S2 cells. AB - Human transferrin (hTf) is a serum glycoprotein involved in Fe3+ transport. Here, a plasmid encoding the hTf gene fused with a hexahistidine (His6) epitope tag under Drosophila metallothionein promoter (pMT) was stably transfected into Drosophila melanogaster S2 cells as a nonlytic plasmid-based system. Following 3 days of copper sulfate induction, transfected S2 cells were found to secrete hTf into serum-free culture medium at a competitively high expression level of 40.8 microg/mL, producing 6.8 microg/mL/day in a 150-mL spinner flask culture. Purification of secreted recombinant hTf using immobilized metal affinity chromatography (IMAC) yielded 95.5% pure recombinant hTf with a recovery of 32%. According to MALDI-TOF mass spectrometry analysis, purified S2 cell-derived His6 tagged recombinant hTf had a molecular weight (76.4 kDa) smaller than that of native apo-hTf (78.0 kDa). 2-Dimensional gel electrophoresis patterns showed recombinant hTf had a simpler and less acidic profile compared to that of native hTf. These data suggest recombinant hTf was incompletely (noncomplex) glycosylated and lacked sialic acids on N-glycans. However, this difference in N glycan structure compared to native hTf had no effect on the iron-binding activity of recombinant hTf. The present data show that a plasmid-based stable transfection S2 cell system can be successfully employed as an alternative for producing secreted functional recombinant hTf. PMID- 15296448 TI - Clinical grade vector production: analysis of yield, stability, and storage of gmp-produced retroviral vectors for gene therapy. AB - Retroviral vector gene transfer of a therapeutic gene to correct or modify a disease process is a promising strategy for many inherited and acquired diseases. A major obstacle in this process is the large-scale production of the gene transfer vector under good manufacturing practice (GMP) conditions. We have used the CellCube bioreactor system to produce five batches of GMP-grade vector. The production batches were of 10-20 L each, and the titers were around 2 x 10(6) IU/mL. We find that this particular vector is relatively stable with a half-life of about 8 h at 37 degrees C, 40 h at 20 degrees C, and 14 days at 4 degrees C. The half-life during storage at -80 degrees C is around 18 months. The supernatant may be frozen and thawed up to five times without any significant loss of titer. We have also made a comparison between the CellCube bioreactor and the automated roller bottle system RollerCell 40 (RC 40). The yields from the two systems were comparable. PMID- 15296449 TI - In vitro assessment of encapsulated C3A hepatocytes functions in a fluidized bed bioreactor. AB - In the present in vitro model, the authors intended to assess viability and functionality of hepatocytes encapsulated into alginate beads and submitted to a fluidized bed motion in a bioreactor. Human immortalized C3A line was chosen as cell model. Two controls consisting of (1) cells cultured on flasks and (2) cells encapsulated in alginate beads under static conditions were implemented. The cell functions studied were total protein, albumin, urea, and ammonia synthesis, as well as ammonia removal in the case of overdose. The comparison among the three cases studied showed that the three-dimensional structure of alginate offered a suitable environment for cell functions. In addition, the fluidized bed bioreactor enhanced the mass transfer and thus increased the amount of species released out of the beads, as compared with the static case. Ammonia detoxification only appeared reduced by encapsulation. The concept of a fluidized bed bioartificial liver was thus validated by this in vitro model, which indicated that cell functions could be efficiently retained. In addition, as far as urea and protein synthesis and release were concerned, the use of the C3A cell line, in combination with encapsulation and fluidization technology, offered a real potentiality for the purpose of extracorporeal liver supply. PMID- 15296450 TI - Modeling and simulation of fed-batch protein refolding process. AB - The simplified kinetic model that assumes competition between first-order folding and third-order aggregation was used to model the fed-batch refolding of denatured-reduced lysozyme. It was found that the model was able to describe the process at limited concentration ranges, i.e., 1-2 and 5-7 mg mL(-)(1), respectively, at extensive guanidinium chloride (GdmCl) concentrations and controlled concentrations of oxidizing and reducing agents. The folding or aggregation rate constant was different at the two protein concentration ranges and strongly dependent on the denaturant concentration. As a result, both rate constants at the two concentration ranges were expressed as functions of GdmCl concentration. The rate constants determined by fed-batch experiments could be employed for the prediction of the fed-batch process but were not able to be extended to a batch refolding by direct dilution. Computer simulations show that the denaturant concentration and fed-batch flow rate are important factors influencing the refolding yield. Prolonged fed-batch time is beneficial to keep the transient intermediate concentration at a low level and to increase the yield of correctly folded protein. This is of importance when the denaturant concentration in refolding buffer solution is low. Thus, at a low denaturant concentration, fed-batch time should be sufficiently long, whereas at an appropriately high GdmCl concentration, a short fed-batch time or a high feed rate of the denatured protein is effective to give a high refolding yield. PMID- 15296451 TI - Stability of enzymes and proteins in dried glassy systems: effect of simulated sunlight conditions. AB - The purpose of the present work was to study the effects of simulated sunlight conditions on enzyme inactivation and structural damage in dehydrated glassy systems. Freeze-dried samples containing different enzymes (lactase, invertase, lysozyme and amyloglucosidase) were exposed to light using a medium-pressure metal halide HPA 400 W lamp. After 1 h of light exposure, the samples showed a significant reduction (more than 50%) in the denaturation peak area as analyzed by DSC, and this could be attributed to protein denaturation. For most of the pure enzymes, the loss of enzymic activity after 1 h of light exposure was around 50%. In the case of enzymes included in anhydrous model systems (trehalose, raffinose, maltodextrin, and dextran), the remaining activity also decreased dramatically during the light treatment. We showed that the light exposure in dehydrated systems generated both the loss of enzymic activity and structural changes such as denaturation (observed by DSC) and protein fragmentation and aggregation (observed by electrophoresis). Overall, we can conclude that a short exposure to the light produces dramatic changes in the enzymic activity in dehydrated systems with or without protective matrices. PMID- 15296452 TI - Effect of glutaraldehyde concentration on the physical properties of polymerized hemoglobin-based oxygen carriers. AB - Artificial blood substitutes based on glutaraldehyde cross-linked hemoglobin (PolyHb) are currently being developed for use in human subjects needing blood transfusions. Despite the commercial development of PolyHb dispersions, a systematic study of the effect of varying the glutaraldehyde to hemoglobin (G-Hb) molar ratio on the resulting PolyHb physical properties (molecular weight distribution and oxygen binding parameters) has not been conducted to date. The results of this study show that increasing the G-Hb molar ratio elicits a general decrease in the P50 (partial pressure of oxygen at which Hb is half saturated with oxygen) and cooperativity and a simultaneous increase in the weight averaged molecular weight (Mw) of the PolyHb dispersion and methemoglobin (MetHb) level. Three PolyHb dispersions (20:1, 30:1, and 40:1 G-Hb molar ratios) displayed potential as artificial blood substitutes. The 20:1 PolyHb dispersion resulted in the presence of more intramolecularly cross-linked and non-cross-linked tetramers versus cross-linked species that were larger than a tetramer ( approximately 75% tetrameric and approximately 25% higher-order species), lower MetHb level (8%), and P50 (20.1 mmHg) similar in magnitude to that of non-cross-linked Hb. The 30:1 PolyHb dispersion consisted of more higher-order species ( approximately 76%), higher MetHb level (28%), and lower P50 (13.3 mmHg). The 40:1 PolyHb dispersion resulted in a similar P50 of 13.0 mmHg and similar MetHb level (30%); however, this PolyHb dispersion only consisted of species larger than a tetramer. The molecular weight distribution of PolyHb dispersions was determined using asymmetric flow field-flow fractionator (AFFF) coupled with multiangle static light scattering (MASLS). This is the first time that AFFF-MASLS has been used to characterize the molecular weight distribution of PolyHb dispersions. PMID- 15296453 TI - Study of the attachment of Pseudomonas aeruginosa on gold and modified gold surfaces using surface plasmon resonance. AB - This paper describes how the technique of surface plasmon resonance (SPR) can be utilized to follow (in real time) the attachment of Pseudomonas aeruginosa bacteria on bare gold and gold modified with a self-assembled monolayer (SAM) of mercaptounadecanoic acid. We show that SPR is able to discriminate between the adsorption of live versus dead (thermally shocked) bacteria. Moreover, the SPR distinguishes between the adsorption of wild-type versus mutant bacteria (single gene knockouts), the concentration of the bacterial suspension, and between bacteria adsorbing on SAM-modified and bare gold. SPR is able to measure bacterial adsorption within seconds of the bacterial suspension being introduced. Finally, a qualitative correlation between results from SPR with a crystal violet staining assay for different mutant bacteria was observed. PMID- 15296454 TI - A metal-chelating piezoelectric sensor chip for direct detection and oriented immobilization of polyHis-tagged proteins. AB - A metal-chelating piezoelectric (PZ) chip for direct detection and controlled immobilization of polyHis-tagged proteins has been demonstrated. The chip was prepared by covalently binding a hydrogel matrix complex of oxidized dextran and nitrilotriacetic acid (NTA) ligand onto an activated alkanethiol-modified PZ crystal. The resulting chip effectively captured Ni2+ ions onto its NTA surface, as disclosed by the resonant frequency shift of the crystal and an X-ray photoelectron spectroscopy analysis. The real-time frequency analysis revealed that the bare NTA chip was nonfouling, regenerable, and highly reusable during continuous repetitive injections of ion solutions and binding proteins. In addition, the chip displayed good long-term reusability and storage stability. The individual binding studies of a polyHis-tagged glutathione-S-transferase and its native untagged form on various metal-charged chips revealed that Co2+, Cu2+, and Ni2+ ions each had different immobilization ability on the NTA surface, as well as their binding ability and selectivity with the tagged protein. As a result, the tagged protein immobilized on the Ni2+-charged chip can actively be bound with its antibody and substrate. Further, the quantitative analyses of the tagged protein in crude cell lysate with a single Ni2+-charged chip and of its substrate with a protein-coated chip were also successfully demonstrated. Therefore, this study initiates the possibilities of oriented, reversible, and universal immobilization of any polyHis-tagged protein and its functional study using a real-time PZ biosensor. PMID- 15296455 TI - Growth behavior in plant cell cultures based on emissions detected by a multisensor array. AB - The use of a multisensor array based on chemical gas sensors to monitor plant cell cultures is described. The multisensor array, also referred to as an electronic nose, consisted of 19 different metal oxide semiconductor sensors and one carbon dioxide sensor. The device was used to continuously monitor the off gas from two plant cell suspension cultures, Morinda citrifolia and Nicotiana tabacum, cultivated under batch conditions. By analyzing the multiarray responses using two pattern recognition methods, principal component analysis and artificial neural networks, it was possible to monitor the course of the cultivations and, in turn, to predict (1) the biomass concentration in both systems and (2) the formation of the secondary metabolite, antraquinone, by M. citrifolia. The results identify the multisensor array method as a potentially useful analytical tool for monitoring plant process variables that are otherwise difficult to analyze on-line. PMID- 15296456 TI - Substrate desolvation as a governing factor in enzymatic transformations of PAHs in aqueous-acetonitrile mixtures. AB - Conversion of hydrophobic substrates such as polyaromatic hydrocarbons (PAHs) was studied in aqueous-organic media using transformation of pyrene by cytochrome c. The experiments were conducted in pure solvents and aqueous-acetonitrile mixtures. The reaction rates dropped gradually as the solvent concentration was increased and were negligible in pure solvents. Thermodynamic calculations show that substrate desolvation was the governing factor in this reaction. The decrease in free energy of activation upon increasing the solvent concentration was found to be proportional to the substrate transfer free energy. This suggests that in order to make such reactions feasible in organic media, it will be necessary to improve the binding between the proteins and the substrates. PMID- 15296457 TI - Affinity precipitation and macroaffinity ligand facilitated three-phase partitioning for refolding and simultaneous purification of urea-denatured pectinase. AB - Protein refolding is an integral step in the recovery of protein activity from inclusion bodies. It is shown that affinity precipitation and macroaffinity ligand facilitated three-phase partitioning (MLFTPP) led to refolding of urea denatured pectinase present in a commercial preparation, with simultaneous purification. Affinity precipitation consists of precipitation of the desired enzyme by complexing it with a suitable stimulus-sensitive macroaffinity ligand. This ligand in this case was alginate/esterified alginate. The complex of the polymer-pectinase could be precipitated by adding calcium ions. In MLFTPP (carried out by adding tertiary butanol and ammonium sulfate to the aqueous solution of crude enzyme and the polymer), the polymer or its complex with the enzyme form an interfacial precipitate between tert-butyl alcohol phase and aqueous phase. It is believed that in both processes, while molecular recognition of alginate/esterified alginate to pectinase facilitates their selective binding to the enzyme, the correct refolding is facilitated by preventing molecular aggregation of unfolded enzyme molecules. Three-phase partitioning with esterified alginate as the macroaffinity ligand gave 100% recovery with 4-fold purification. Affinity precipitation with 1% alginate gave 52% yield with 18-fold purification. On the other hand, use of 0.5% esterified alginate gave only 7-fold purification but with 75% recovery of activity. PMID- 15296458 TI - Immobilization of lactase from Kluyveromyces lactis greatly reduces the inhibition promoted by glucose. full hydrolysis of lactose in milk. AB - The kinetic constants (Km, Vmax, and inhibition constants for the different products) of soluble and different immobilized preparations of beta-galactosidase from Kluyveromyces lactis were determined. For the soluble enzyme, the Km was 3.6 mM, while the competitive inhibition constant by galactose was 45 mM and the noncompetitive one by glucose was 758 mM. The immobilized preparations conserved similar values of Km and competitive inhibition, but in some instances much higher values for the noncompetitive inhibition constants were obtained. Thus, when glyoxyl or glutaraldehyde supports were used to immobilize the enzyme, the noncompetitive inhibition was greatly reduced (Ki approximately 15,000 and >40,000 mM, respectively), whereas when using sugar chains to immobilize the enzyme the behavior had an effect very similar to the soluble enzyme. These results presented a great practical relevance. While using the soluble enzyme or the enzyme immobilized via the sugar chain as biocatalysts in the hydrolysis of lactose in milk only around 90% of the substrate was hydrolyzed, by using of these the enzyme immobilized via the glyoxyl or the glutaraldehyde groups, more than 99% of the lactose in milk was hydrolyzed. PMID- 15296459 TI - Recombinant protein production in high cell density cultures of Escherichia coli with galactose as a gratuitous inducer. AB - A new expression system was developed by introducing two major modifications into the genome of Escherichia coli: a deletion in the gal operon (DeltagalEKT) to allow the use of the inexpensive compound galactose as a gratuitous inducer and the introduction of the gal P2 promoter driving the expression of the T7 RNA polymerase. The novel JRR10 strain containing these two features gives high-level expression of a reporter gene cloned under the T7 phi10 promoter in high cell density cultures. The cost of the induction of this novel system is more than 30 times lower than that of the IPTG-induced system of the widely used BL21 strain. PMID- 15296460 TI - Enzymatic hydrolysis of gelatin layers on used lith film using thermostable alkaline protease for recovery of silver and PET film. AB - To develop a new efficient and potential industrial enzymatic process for the recovery of silver and poly(ethylene terephthalate) (PET) from used lith film for printing, which has not been recycled at all, enzymatic hydrolysis of gelatin layers on lith film was investigated using the thermostabilized mutant enzyme of the alkaline protease from alkaliphilic Bacillus sp. B21-2. The rate of gelatin hydrolysis of lith film in a stirred-tank reactor increased with the temperature and enzyme concentration. The time required to complete the hydrolysis of gelatin on lith film was longer than that on X-ray film because of the tightly cross linked structure of the gelatin layers of lith film. The time required to complete the hydrolysis by using the mutant enzyme was less than that using the wild-type enzyme. The gelatin hydrolysis of lith film was well explained by a model that took into consideration a number of physical processes in addition to the chemical process. PMID- 15296461 TI - Biochemical pulping of bagasse. AB - This study deals with pretreatment of wheat straw with lignin-degrading fungi and its effect on chemical pulping. Ceriporiopsis subvermispora strains, which preferentially attack the lignin, were used for biochemical pulping of bagasse. Treatment of depithed bagasse with different strains of C. subvermispora reduced the kappa number by 10-15% and increased unbleached pulp brightness by 1.1-2.0 ISO points on chemical pulping at the same alkali charge. Bleaching of biopulps at the same chemical charge increased final brightness by 4.7-5.6 ISO points and whiteness by 10.2-11.4 ISO points. Fungal treatment did not result in any adverse effect on the strength properties of pulp. PMID- 15296462 TI - Glycosylation profiles of the human colorectal cancer A33 antigen naturally expressed in the human colorectal cancer cell line SW1222 and expressed as recombinant protein in different insect cell lines. AB - The A33 antigen is a cell surface glycoprotein expressed in human gastrointestinal epithelium and in 95% of colorectal cancers. We have compared the N-linked glycosylation profile of A33 antigen naturally expressed in a human colorectal cancer cell line with recombinant human A33 antigen (rA33) produced in insect cell culture using the baculovirus expression vector. N-Linked glycans were enzymatically released from the protein, and glycan composition was analyzed by HPLC. In three insect cell lines tested (Sf-21, Tn5B1-4, and Tn-4s), glycosylation of rA33 was dominated by high mannose structures (M5Gn2 to M9Gn2; 78-95% of total N-linked glycans), with M8Gn2 being the single most abundant glycoform. A33 antigen naturally expressed in the SW1222 human colon cancer cell line (A33) also possessed a high abundance of high mannose glycans (72%). No complex glycosylation was detected on rA33 expressed in insect cells. Natural A33 was galactosylated to a small extent (6%). These results illustrate a case of similar glycosylation of a glycoprotein between a recombinant version produced in insect cell culture and its counterpart naturally expressed in human cell culture. PMID- 15296463 TI - Solid-state fermentation for production of griseofulvin on rice bran using Penicillium griseofulvum. AB - Griseofulvin is a secondary metabolite produced from fungal species that have morphology suitable for solid-state fermentation (SSF). Reports on production of griseofulvin by SSF are scarce. The present work investigates SSF for griseofulvin production, optimization of its process parameters vis-a-vis the conventional submerged fermentation and its downstream processing from the same. Rice bran adjusted to an initial moisture content (IMC) of 50% (v/w) inoculated with 1 mL of a suspension of 10(6) spores/mL under agitation at 250 rpm containing the modified Czapek-Dox medium and additional 0.1% choline chloride as a precursor gave a yield of griseofulvin in 9 days that was comparable to submerged fermentation after 28 days. The yield of griseofulvin (microg/g dry biomass) was comparable in SSF and submerged fermentation. The biomass was estimated by estimation of chitin. Discussions on the effect of each parameter in SSF have also been included. PMID- 15296464 TI - Optimization of batch and fed-batch bioreactors using simulated annealing. AB - In order for biobased industrial products to compete economically with petroleum derived products, significant reduction in their processing cost is necessary. Since most bioprocesses are operated in batch or fed-batch mode, their optimization involves theoretical and computational challenges. Simulated annealing (SA), a stochastic optimization algorithm, is used in this study to solve a number of challenging optimization problems related to the design and operation of bioreactors. Two well-known case studies are considered in which the robustness and efficiency of the SA algorithm is demonstrated. More specifically, in the first case study it is shown that the global optimal solution located by SA achieves significant improved productivity when compared with the results of previous investigations. In the second case study a realistic objective function is considered where the economic performance of a bioprocess is optimized. SA exhibits impeccable performance and robustness and was able to locate the global optimal solution irrespective of the initial point selected. PMID- 15296465 TI - Quantitative analysis of indigo and indigo precursors in leaves of Isatis spp. and Polygonum tinctorium. AB - Analysis of extracts from two woad species (Isatis tinctoria and Isatis indigotica) and Polygonum tinctorium revealed that only one indigo precursor (indican) was present in Polygonum, but two precursors were found in Isatis spp. This was done using high performance liquid chromatography (HPLC), coupled to an evaporative light scattering detector (ELSD). In Isatis spp., the indigo precursors indican and a fraction representing isatan B were identified. The proportion of indican and isatan B was different between the two Isatis spp. tested. For the first time, it was possible to quantify the precursors in woad plant species, and the results were found to be in good agreement with those made from total indigo quantification using two different spectrophotometric methods or a derivatization technique. PMID- 15296466 TI - Effect of simultaneous application of stressful culture conditions on specific productivity and heterogeneity of erythropoietin in Chinese hamster ovary cells. AB - A single stressful culture condition induced by hypoosmotic stress (210 mOsm kg( 1)), low culture temperature (32 degrees C), or NaBu addition (1 mM) resulted in a 1.8- to 2.2-fold enhancement of specific erythropoietin (EPO) productivity (qEPO) of recombinant Chinese hamster ovary (rCHO) cells compared to normal culture condition (37 degrees C and 310 mOsm kg(-1)). Simultaneous application of these stressful conditions further enhanced qEPO up to approximately 5-fold. However, the quality of EPO was affected by stressful culture conditions. The proportion of acidic isoforms of EPO under a single stressful condition was 2.8 13.8% lower than that under normal culture condition. Simultaneous application of the stressful conditions further decreased the portion of acidic isoforms but not significantly. Despite 5-fold enhancement of q(EPO), the portion of acidic isoforms under the simultaneous application of stressful culture conditions was 12.9-21.6% lower than that under normal culture condition. Taken together, these results suggest the potential of simultaneous application of different stressful culture conditions to the production phase of two-stage culture, where cell growth and production phases are separated, for improved EPO production. PMID- 15296467 TI - Reversible immobilization of glucoamylase by ionic adsorption on sepabeads coated with polyethyleneimine. AB - Glucoamylase (GA) from Aspergillus niger was immobilized via ionic adsorption onto DEAE-agarose, Q1A-Sepabeads, and Sepabeads EC-EP3 supports coated with polyethyleneimine (PEI). After optimization of the immobilization conditions (pH, polymer size), it was observed that the adsorption strength was much higher in PEI-Sepabeads than in Q1A-Sepabeads or DEAE-supports, requiring very high ionic strength to remove glucoamylase from the PEI-supports (e.g., 1 M NaCl at pH 5.5). Thermal stability and optimal temperature was marginally improved by this immobilization. Recovered activity depended on the substrate used, maltose or starch, except when very low loading was used. The optimization of the loading allowed the preparation of derivatives with 750 IU/g in the hydrolysis of starch, preserving a high percentage of immobilized activity (around 50%). PMID- 15296468 TI - Perinatal programming of appetite control by leptin? PMID- 15296469 TI - Is growth hormone deficiency contributing to heart failure in patients with beta thalassemia major? AB - A 21-year-old woman with beta-thalassemia major (beta-TM) and GH deficiency developed end-stage heart failure, New York Heart Association (NYHA) functional class IV, within 3 months after withdrawal of recombinant human growth hormone (GH). A myocardial biopsy excluded myocarditis and showed moderate iron deposit in the heart. Before her admission, intensified treatments with digoxin, angiotensin-converting enzyme inhibitor, diuretics and extra chelation therapy (desferrioxamine (DFO)) had not improved her progressive heart failure. At admission, GH was reinstituted together with intensified treatment of cardiac drugs and low doses of DFO, and her heart failure reversed. Four months later, NYHA functional class II was reached and within 1 year her cardiac function was normalised. We suggest that GH deficiency due to iron-induced damage to the hypothalamic-pituitary axis can contribute to heart failure in adult patients with beta-TM. PMID- 15296470 TI - Serum parathyroid hormone level is associated with body mass index. The 5th Tromso study. AB - OBJECTIVE: To study whether serum parathyroid hormone (PTH) and serum calcium are associated with body mass index (BMI), and their predicting role in obesity. DESIGN: Population based, cross-sectional study. METHODS: In 2001 a population based health survey was held in Tromso, North Norway. Questionnaires on medical history and life-style factors were completed and anthropometric data were collected. Calcium and vitamin D intakes and a physical activity score were calculated. Serum calcium and PTH were measured in a subset of 3447 men and 4507 women. Pearson correlation and linear regression were used to evaluate associations between BMI, PTH and serum calcium, and logistic regression was used to test PTH and serum calcium as predictors of obesity and to calculate odds ratio. Relative risk was calculated using frequency tables. RESULTS: For serum calcium and PTH there was a significant positive relation to BMI in both genders (P<0.001), which to our knowledge has not previously been reported on the basis of a large epidemiological study. Age, low calcium and vitamin D intakes were explanatory variables for serum PTH. The highest quartile of serum PTH (>4.20 pmol/l) was a significant predictor for obesity (P<0.001) in both genders, adjusted for age, physical activity and serum calcium. Obesity rates were higher in those with PTH levels in the highest quartile compared with those in the lower quartiles, which resulted in a relative risk of 1.40 (95% confidence interval (C.I.) 1.20-1.60) for men and 1.48 (95% C.I. 1.31-1.67) for women. CONCLUSIONS: Serum PTH, adjusted for age, physical activity and serum calcium, is positively associated with BMI in both sexes, and serum PTH is an independent predictor of obesity in our statistical model. PMID- 15296471 TI - Effects of chronic administration of PPAR-gamma ligand rosiglitazone in Cushing's disease. AB - OBJECTIVE: Rosiglitazone, a thiazolidinedione compound with peroxisome proliferator-activated receptor-gamma (PPAR-gamma)-binding affinity, is able to suppress adrenocorticotropic hormone (ACTH) secretion in treated mice and in AtT20 pituitary tumor cells. These observations suggested that thiazolidinediones may be effective as therapy for Cushing's disease (CD). PATIENTS AND METHODS: Rosiglitazone (8 mg/day) was administered to 14 patients with active CD (13 women, one man, 18-68 years). Plasma ACTH, serum cortisol (F) and urinary free cortisol (UFC) levels were measured before and then monthly during rosiglitazone administration. RESULTS: In six patients a reduction of ACTH and F levels and a normalization of UFC were observed 30-60 days after the beginning of rosiglitazone administration: there was a significant difference between basal and post-treatment values for UFC (1238+/-211 vs 154+/-40 nmol/24 h, P<0.03), but not for ACTH (15.9+/-3.7 vs 7.9+/-0.9 pmol/l) and F levels (531+/-73 vs 344+/-58 nmol/l). Two of six cases, followed up for 7 months, showed a mild clinical improvement. Eight patients were nonresponders after 30-60 days of rosiglitazone treatment: their ACTH, F and UFC levels did not differ before and during drug administration. Immunohistochemical analysis of pituitary tumors removed from two responder and two nonresponder patients showed a similar intense immunoreactivity for PPAR-gamma in about 50% of cells. CONCLUSIONS: The administration of rosiglitazone seems able to normalize cortisol secretion in some patients with CD, at least for short periods. Whether the activation of PPAR-gamma by rosiglitazone might be effective as chronic pharmacologic treatment of CD needs a more extensive investigation through a randomized and controlled study. PMID- 15296472 TI - Evaluation of cardiac structure by echoreflectivity analysis in acromegaly: effects of treatment. AB - OBJECTIVES: Cardiac echoreflectivity is a noninvasive tool for evaluating cardiac fibrosis. The present paper aimed to study the modifications of cardiac echoreflectivity in a group of acromegalic patients before and after therapy, and to assess possible correlations with serum levels of procollagen III (PIIINP), a peripheral index of collagen synthesis. DESIGN AND METHODS: Cardiac echoreflectivity (as assessed by analyzing 2-D echocardiograms digitized off-line onto a personal computer) and PIIINP levels were evaluated in 16 acromegalic patients of new diagnosis not affected by arterial hypertension (10 males, six females, age+/-s.d.: 38+/-10 years), and in a group of 16 sex- and age-matched healthy subjects. All the patients were re-evaluated after surgical and/or medical therapy for acromegaly. The echo patterns were analyzed by software that supplies the derived collagen volume fraction (dCVF), an index of fibrosis. RESULTS: At baseline, acromegalic patients showed significantly higher dCVF values and PIIINP levels than healthy controls (3.1+/-0.5% vs 1.6+/-0.3%, P<0.01 and 8.7+/-2.2 vs 3.1+/-1.1 ng/ml, P<0.05, respectively, by unpaired Student's t test). After therapy, dCVF and PIIINP levels normalized in the six controlled patients (that is, GH of <2.5 microg/l and IGF-I within normal range) (dCVF from 2.8+/-0.4% to 1.4+/-0.2%, P<0.001; PIIINP from 8+/-2.7 to 3.3+/-1.9 ng/ml, P<0.05), while no significant changes were found in noncontrolled patients (dCVF from 3.3+/-0.6% to 2.9+/-1.2% and PIIINP from 9.1+/-1.9 to 7.9+/-3.5 ng/ml, P=NS). A positive correlation between dCVF and PIIINP (r=0.75, P<0.001) and between IGF-I and both dCVF and PIIINP (r=0.65 and 0.61 respectively, P<0.05) was found in acromegalic patients. CONCLUSIONS: Cardiac echoreflectivity, which may be a reflection of heart collagen content, is increased in patients with active acromegaly and correlates with PIIINP concentrations. After cure or adequate control of the disease, both parameters revert to normal. Echoreflectivity analysis could be a useful adjuvant parameter in the assessment of the activity of acromegalic disease. PMID- 15296473 TI - Effect of hormone therapy, tibolone and raloxifene on circulating vascular endothelial growth factor in Greek postmenopausal women. AB - OBJECTIVE: To evaluate the effect of continuous combined hormone therapy (HT), tibolone and raloxifene on circulating vascular endothelial growth factor (VEGF) in postmenopausal women. DESIGN: One-year prospective intervention study. METHODS: One hundred and forty-six postmenopausal women with a mean age of 51.8+/ 4.1 (s.d.) years received 0.625 mg conjugated equine estrogen (CEE) plus 5 mg medroxyprogesterone acetate (MPA) (CEE/MPA, n=34), 2.5 mg tibolone (n=37), 60 mg raloxifene (n=40) or no active treatment (control group, n=35). Plasma VEGF was estimated at baseline and at 6 and 12 months. RESULTS: In both the CEE/MPA treated and the tibolone-treated groups plasma VEGF increased significantly at month 6 and remained elevated at month 12 (CEE/MPA baseline: 268.1+/-187.8 pg/ml, month 6: 320.0+/-175.3 pg/ml, month 12: 321.1+/-181.8 pg/ml, P=0.01; tibolone baseline: 240.6+/-165.8 pg/ml, month 6: 271.4+/-172.7 pg/ml, month 12: 274.8+/ 183.1 pg/ml, P=0.03). These changes were significantly different from the respective changes in the control group after adjusting for T-score in bone densitometry (CEE/MPA: P=0.02, tibolone: P=0.04). The effect of HT or tibolone on plasma VEGF was mainly evident in women with low baseline VEGF levels (<243.2 pg/ml, median for whole sample). On the contrary, VEGF levels in the raloxifene treated or the control group did not change throughout the study. CONCLUSION: Both continuous combined HT and tibolone increased circulating VEGF in postmenopausal women, while raloxifene had no effect. Further research is needed to clarify the clinical relevance of these findings with respect to cardiovascular risk in postmenopausal women. PMID- 15296474 TI - A promoter polymorphism of the CYP27B1 gene is associated with Addison's disease, Hashimoto's thyroiditis, Graves' disease and type 1 diabetes mellitus in Germans. AB - BACKGROUND: CYP27B1 hydroxylase catalyzes the conversion of 25 hydroxyvitamin D(3) (25OHD(3)) to 1,25(OH)(2)D(3), the most active natural vitamin D metabolite, which plays a role in the regulation of immunity and cell proliferation. We therefore investigated two single nucleotide polymorphisms in the CYP27B1 hydroxylase gene for an association with Addison's disease, Hashimoto's thyroiditis, Graves' disease and type 1 diabetes mellitus. METHODS: Patients with Addison's disease (n=124), Hashimoto's thyroiditis (n=139), Graves' disease (n=334), type 1 diabetes mellitus (n=252) and healthy controls (n=320) were genotyped for the promoter (-1260) C/A polymorphism and for the intron 6 (+2838) C/T polymorphism of the CYP27B1 gene. Patients and controls were compared using genotype-wise and allele-wise X(2) testing. RESULTS: A significant association was found between allelic variation of the promoter (-1260) C/A polymorphism and Addison's disease, Hashimoto's thyroiditis, Graves' disease and type 1 diabetes mellitus (P=0.0062, P=0.0173, P=0.0094 and P=0.0028 respectively). Significant differences were also observed for the intron 6 (+2838) C/T polymorphism (P=0.0058) in Hashimoto's thyroiditis but not for the other autoimmune endocrine diseases. CONCLUSIONS: The CYP27B1 promoter (-1260) C/A polymorphism appears to be associated with endocrine autoimmune diseases but the CYP27B1 intron 6 (+2838) C/T polymorphism appears to be associated only with Hashimoto's thyroiditis. These results imply a regulatory difference of the CYP27B1 hydroxylase to predispose to endocrine autoimmunity. PMID- 15296475 TI - Type 2 diabetes and impaired glucose tolerance in European children and adolescents with obesity -- a problem that is no longer restricted to minority groups. AB - BACKGROUND: The incidence of childhood obesity and type 2 diabetes is an increasing problem in Europe. We determined the prevalence of impaired glucose regulation in a predominantly Caucasian cohort of 491 children and adolescents with obesity. METHODS: Fasting glucose and insulin levels were determined in all 491 subjects. Patients with an abnormal fasting glucose or with additional risk factors (positive family history of type 2 diabetes, acanthosis nigricans, hyperlipidemia; n=102) underwent an oral glucose tolerance test (OGTT; 1.75 g glucose/kg body weight). Homeostasis model assessment was used to estimate insulin resistance in all subjects. The insulin sensitivity index was determined in those subjects who underwent an OGTT. Screening for mutations in the melanocortin 4 receptor (MC4R) gene and the coding region of the brain-derived neutrophic factor (BDNF) in 37 patients with an impaired glucose tolerance was performed by WAVE analysis. RESULTS: Out of the total of 491 patients, 12 had an abnormal fasting glucose level. Of the 102 patients who underwent an OGTT, 37 had impaired glucose tolerance; 6 out of the 102 patients were diagnosed with type 2 diabetes. Eighty-eight per cent of patients with abnormal glucose tolerance and 66% of patients with type 2 diabetes were Caucasian. Insulin resistance indices correlated well with the degree of abnormal glucose tolerance. Using the screening algorithm for type 2 diabetes as advocated by the American Diabetes Association, 68% of patients with impaired glucose tolerance and 66% of patients with type 2 diabetes would have been missed. No abnormalities in the MC4R and BDNF genes were detected. CONCLUSIONS: Impaired glucose tolerance and type 2 diabetes are far more common in obese European children of Caucasian origin than previously thought. Using fasting glucose levels as the main screening tool appears to be insufficient in detecting these children. PMID- 15296476 TI - The role of peripheral benzodiazepine receptors on the function and survival of isolated human pancreatic islets. AB - OBJECTIVE: Peripheral benzodiazepine receptors (PBRs) are part of the mitochondrial permeability transition pore, and their activation may induce cell death. PBRs are expressed in human pancreatic islets, and cytokine-induced damage is accompanied by changes in their properties. We hypothesized that PBRs can have a role in human islet physiopathology, and evaluated the effects of prolonged exposure to two specific PBR ligands, PK11195 and Ro5-4864 on the function and survival of isolated human islets. DESIGN: Isolated human islets were prepared from the pancreas of 25 multiorgan cadaveric donors and incubated for 12 h in the presence of PK11195 or Ro5-4864. Insulin secretion studies and apoptosis experiments were then performed, together with assessment of intracellular pathways involved in islet cell function and survival. METHODS: Islets were prepared by enzymatic digestion and density gradient purification. Insulin secretion was assessed by the batch incubation method, and glucose oxidation was evaluated by the use of D-[U-(14)C]glucose. Apoptosis was studied using the TUNEL technique, ELISA methods, and electron microscopy evaluation. PCR experiments were performed by the use of specific primers. RESULTS: Glucose-stimulated insulin release was significantly lower after exposure to PK11195 than after exposure to Ro5-4864. This was accompanied by reduced glucose oxidation and no major change of insulin or GLUT-1 mRNA expression. Apoptosis was higher in PK11195-exposed islets, and electron microscopy demonstrated the involvement of beta-cells. The apoptotic effects were prevented by bongkrekic acid and low-dose cyclosporin A, which stabilize the mitochondrial membrane, and were associated with no evident change of inducible nitric oxide synthase (iNOS), B-cell leukemia/lymphoma-2 (Bcl-2) or Bcl-2-associated X protein (Bax) expression. Caspase inhibition markedly reduced the amount of apoptosis, and the role of these proteases was confirmed by the increased activity of caspase-3 and caspase 9. CONCLUSIONS: Prolonged binding to PBRs may cause human beta-cells functional damage and apoptosis, a phenomenon which is prevented by stabilizing the mitochondrial membrane; occurs without changes of iNOS, Bax and Bcl-2 mRNA expression; and involves caspase activation. These results suggest an involvement of PBRs in human pancreatic beta-cell function and survival. PMID- 15296477 TI - Plasma chromogranin A in incidental non-functioning, benign, solid adrenocortical tumors. AB - OBJECTIVE: To evaluate whether adenomas arising from the adrenal cortex, a tissue of epithelial origin, are associated with high chromogranin A (CgA) levels and whether such tumors may express and release this protein. In addition, to investigate whether high CgA levels imply a neuroendocrine differentiation of the adrenocortical adenomas and, therefore, represent a humoral marker of malignant transformation of these tumors. DESIGN: Plasma CgA of 80 patients with non functioning, benign adrenocortical adenomas was compared with that of 137 tumor free subjects. In 15 patients, the masses were surgically removed and CgA was measured 2 months later. The other 65 patients with adrenocortical adenomas underwent clinical and radiological follow-up (range 24-36 months). METHODS: CgA was evaluated by immunoradiometric assay in peripheral blood and by immunohistochemistry in adrenal tissue specimens. RESULTS: An increase in plasma CgA (P<0.001) was observed in patients with adrenocortical adenomas (83.4+/-7.5 ng/ml) in comparison with tumor-free subjects (43.1+/-1.5 ng/ml). The prevalence of high CgA levels was 25% in the former and 0.7% in the latter. By multiple regression analysis, an increase (49%) in the expected median CgA value was estimated for adrenocortical adenomas (P<0.001). Receiver operating characteristic analysis showed a good diagnostic performance of CgA in identifying patients with adrenocortical adenomas (pure accuracy=0.78, 95% CI=0.71-0.84). In the operated patients, CgA levels did not change before (80.6+/ 16.5 ng/ml) and after (74.3+/-16.3 ng/ml) surgery and in no case was CgA immunoreactivity found in adenoma tissues. The non-operated patients did not develop signs or symptoms of disease and showed no features of malignant transformation of the masses. CONCLUSIONS: Our data showed a strong association between adrenocortical adenomas and high CgA levels. CgA hypersecretion was not due to adenoma tissue, which did not show immunoreactivity for CgA. Finally, elevated CgA levels did not represent a humoral marker of malignant transformation of cortical adenomas. PMID- 15296478 TI - Effects of insulin--like growth factor-I treatment on the endocrine pancreas of hypophysectomized rats: comparison with growth hormone replacement. AB - BACKGROUND: In GH-deficient humans, GH and IGF-I treatment cause opposite effects on serum insulin concentrations and insulin sensitivity. This finding contrasts with the somatomedin hypothesis that IGF-I mediates GH action, as postulated for skeletal growth, and raises the question whether GH-induced IGF-I acts on the endocrine pancreas in the same way as administered IGF-I. OBJECTIVE: To compare the effects of the two hormones on the endocrine pancreas of hypophysectomized rats. METHODS: Animals were infused for 2 days, via miniosmotic pumps, with IGF-I (300 microg/day), GH (200 mU/day) or vehicle. We measured (i) glucose, IGF-I, insulin, C-peptide and glucagon in serum and (ii) IGF-I, insulin and glucagon mRNAs and peptides in the pancreas by radioimmunoassay, immunohistochemistry and northern analysis. RESULTS: Both GH and IGF-I treatment increased serum and pancreatic IGF-I but, unlike GH, IGF-I treatment strongly reduced serum insulin and C-peptide (and, to a lesser extent, serum glucagon). Nevertheless, the animals did not become hyperglycaemic. GH, but not IGF-I, increased pancreatic insulin and glucagon content, as also indicated by immunohistochemistry, and increased IGF-I mRNA. Neither GH nor IGF-I caused significant changes in insulin and glucagon mRNA. CONCLUSIONS: The decrease in serum insulin and C-peptide by IGF-I treatment without significant changes in insulin gene expression and pancreatic insulin content suggests inhibition of insulin secretion. Within this setting, the absence of hyperglycaemia points to enhanced insulin sensitivity, although an insulin-like action of infused IGF-I may have partially compensated for the decreased insulin concentrations. GH-induced circulating or pancreatic IGF-I, or both, does not mimic the pancreatic effects of infused IGF-I in the absence of GH, suggesting that GH may counteract the action of GH-induced IGF-I on the endocrine pancreas. PMID- 15296479 TI - Ghrelin exerts a proliferative effect on a rat pituitary somatotroph cell line via the mitogen-activated protein kinase pathway. AB - OBJECTIVES: Ghrelin is a brain-gut peptide with GH-releasing and appetite inducing activities and a widespread tissue distribution. Ghrelin is the endogenous ligand of the GH secretagogue receptor type 1a (GHS-R1a), and both ghrelin and the GHS-R1a are expressed in the pituitary. There are conflicting data regarding the effects of ghrelin on cell proliferation. A positive effect on proliferation and activation of the mitogen-activated protein kinase (MAPK) pathway has been found in hepatoma, adipose, cardiomyocyte and prostate cell lines. However, ghrelin has also been shown to have anti-proliferative effects on breast, lung and thyroid cell lines. We therefore examined the effect of ghrelin on the rat pituitary cell line GH3. METHODS: RT-PCR was used for the detection of GHS-R1a and pre-proghrelin mRNA expression in GH3 cells. The effect of ghrelin on cell proliferation was studied using [(3)H]thymidine incorporation; cell counting and the activation of the MAPK pathway were studied using immunoblotting and inhibitors of the extracellular signal-regulated kinase 1 and 2 (ERK 1/2), protein kinase C (PKC) and tyrosine phosphatase pathways. RESULTS: GHS-R1a and ghrelin mRNA expression were detected in GH3 cells. Ghrelin, at 10(-10) to 10(-6) M concentrations, significantly increased [(3)H]thymidine incorporation (at 10( 9) M, 183+/-13% (means+/-s.e.m.) compared with untreated controls), while 12 phorbol 13-myristate acetate (PMA) at 10(-7) M (used as a positive control) caused a 212+/-14% increase. A reproducible stimulatory effect of desoctanoyl ghrelin was also observed on [(3)H]thymidine incorporation (135+/-5%; P<0.01 at 10(-9) M compared with control), as well as on the cell count (control 6.8 x 10(4)+/-8.7 x 10(3) cells/ml vs desoctanoyl ghrelin (10(-9) M) 1.04 x 10(5)+/-7.5 x 10(3) cells/ml; P<0.01). Ghrelin caused a significant increase in phosphorylated ERK 1/2 in immunoblotting, while desoctanoyl ghrelin showed a smaller but also significant stimulatory effect. The positive effect of ghrelin and desoctanoyl ghrelin on [(3)H]thymidine incorporation was abolished by the MAPK kinase inhibitor U0126, the PKC inhibitor GF109203X and the tyrosine kinase inhibitor tyrphostin 23, suggesting that the ghrelin-induced cell proliferation of GH3 cells is mediated both via a PKC-MAPK-dependent pathway and via a tyrosine kinase-dependent pathway. This could also be clearly demonstrated by Western blot analysis, where a transient increase in ERK 1/2 phosphorylation by ghrelin was attenuated by all three inhibitors. CONCLUSION: We have shown a novel role for ghrelin in stimulating the proliferation of a somatotroph pituitary tumour cell line, suggesting that ERK activation is involved in mediating the effects of ghrelin on cell proliferation. Desoctanoyl ghrelin showed a similar effect. As ghrelin has been shown to be expressed in both normal and adenomatous pituitary tissue, locally produced ghrelin may play a role in pituitary tumorigenesis via an autocrine/paracrine pathway. PMID- 15296480 TI - A mathematical comparison of techniques to predict biologically available testosterone in a cohort of 1072 men. AB - OBJECTIVE: In the absence of widely available measures of determining free and/or bioavailable testosterone (BioT) physicians may use formulae such as the free androgen index (FAI) to estimate free testosterone. We compared the efficacy of calculated markers of androgen status in predicting serum BioT and hypogonadism. DESIGN: Total testosterone (TT), sex hormone binding globulin (SHBG) and BioT were determined in a large cohort of men. Comparison of calculated androgen levels was performed following endocrine assessment. METHODS: TT and SHBG were determined by ELISA, and BioT was determined by ammonium sulphate precipitation. From these data we calculated FAI and free testosterone using two other published formulae - FTnw (free testosterone as calculated by the method of Nanjeee and Wheeler) and FTv (free testosterone as calculated by the method of Vermeulen). A novel formula was derived to calculate BioT from given levels of TT and SHBG (BTcalculated). The ability of the methods (FAI, FTnw, FTv, BTcalc) to predict BioT were compared using regression analysis. The ability of these markers of androgen status to predict biochemical hypogonadism was compared using area under receiver operator curve (auROC). RESULTS: The equation derived from our data was the best predictor of BioT (R(2)=0.73, P<0.0001) although TT was also a good marker (R(2)=0.68, P=0.0001). In the determination of hypogonadism, of all currently available formulae none were better that the TT (auROC: TT=0.93, FAI=0.72, FTnw=0.91, FTv=0.88) although when TT is borderline (7.5 2 mmol/L, and (2) those with co-oximetry measured saturations (SaO(2)) < 90% and no evidence of shock. Measurements of SaO(2) of whole blood were compared to simultaneous pulse oximetry saturations (SpO(2)). Data were analyzed to detect significant differences in SpO(2) readout failures between oximeters and average SpO(2) - SaO(2) +/- 1 SD for each oximeter. A total of 122 SaO(2) measurements were recorded; the median SaO(2) was 83% (57 - 100%). SpO(2) failures after cardiopulmonary bypass were 41% (25/61) for N versus 10% (6/61) for M (P < .001). There was a significant difference in bias (ie, average SpO(2) - SaO(2)) and precision (+/- 1 SD) between oximeters (N, 1.1 +/- 3.3 vs M, -0.2 +/- 4.1; P < .001) in the postcardiopulmonary bypass group but no significant difference in bias and precision between oximeters in the cyanotic congenital heart disease group (N, 2.9 +/- 4.6 vs M, 2.8 +/- 6.2; P = .848). The Nellcor N-395 pulse oximeter failed more often immediately after cardiopulmonary bypass than did the Masimo SET Radical pulse oximeter. SpO2 measured with both oximeters overestimated SaO2 in the presence of persistent hypoxemia. PMID- 15296627 TI - Environmental health promotion: bridging traditional environmental health and health promotion. AB - This article highlights the juncture between environmental health and health promotion and underscores the need for health promotion involvement in environmental health practice. It begins with a synopsis of current issues in environmental public health and deficiencies in environmental public health practice that could be partly ameliorated by an increased focus on environmental health promotion. Environmental health promotion lies at the intersection between the two disciplines and can be defined as any planned process employing comprehensive health promotion approaches to assess, correct, control, and prevent those factors in the environment that can potentially harm the health and quality of life of present and future generations. An introduction is also provided to the six articles contained in this special issue focused on environmental health promotion, and a brief discussion of crosscutting themes and issues is presented. PMID- 15296628 TI - Understanding wicked problems: a key to advancing environmental health promotion. AB - Complex environmental health problems--like air and water pollution, hazardous waste sites, and lead poisoning--are in reality a constellation of linked problems embedded in the fabric of the communities in which they occur. These kinds of complex problems have been characterized by some as "wicked problems" wherein stakeholders may have conflicting interpretations of the problem and the science behind it, as well as different values, goals, and life experiences. Accordingly, policy makers, public health professionals, and other stakeholders who grapple with these problems cannot expect to effectively resolve them by relying solely on expert-driven approaches to problem solving. Rather, they need to acknowledge that wicked environmental health problems are most likely to yield to (1) the application of effective community health promotion skills, (2) a sustained commitment to sound toxicological and epidemiological science, (3) the application of systems thinking, and (4) transparent communication among all stakeholders. PMID- 15296629 TI - Social determinants of health: implications for environmental health promotion. AB - In this article, the authors draw on the disciplines of sociology and environmental and social epidemiology to further understanding of mechanisms through which social factors contribute to disparate environmental exposures and health inequalities. They propose a conceptual framework for environmental health promotion that considers dynamic social processes through which social and environmental inequalities--and associated health disparities--are produced, reproduced, and potentially transformed. Using empirical evidence from the published literature, as well as their own practical experiences in conducting community-based participatory research in Detroit and Harlem, the authors examine health promotion interventions at various levels (community-wide, regional, and national) that aim to improve population health by addressing various aspects of social processes and/or physical environments. Finally, they recommend moving beyond environmental remediation strategies toward environmental health promotion efforts that are sustainable and explicitly designed to reduce social, environmental, and health inequalities. PMID- 15296630 TI - Community capacity for environmental health promotion: determinants and implications for practice. AB - The human response to an environmental hazard can either reduce or exacerbate its impact on health. This article reviews determinants of community-level responses to environmental health hazards. The aim is to identify factors that can enhance a community's capacity to protect itself and to suggest public health strategies that can increase such capacity. Four case histories of community environmental health action are presented to test a theoretical model for understanding the determinants of community capacity to promote environmental health. Specific actions public health professionals can take to strengthen community capacity include increasing access to accurate science, building strong relationships between communities and local health departments, and supporting political reforms that level the playing field for communities that seek to challenge corporate or government practices. PMID- 15296631 TI - Application of health promotion theories and models for environmental health. AB - The field of environmental health promotion gained new prominence in recent years as awareness of physical environmental stressors and exposures increased in communities across the country and the world. Although many theories and conceptual models are used routinely to guide health promotion and health education interventions, they are rarely applied to environmental health issues. This article examine show health promotion theories and models can be applied in designing interventions to reduce exposure to environmental health hazards. Using the Community Action Against Asthma (CAAA) project as an example, this article describes the application of these theories and models to an intervention aimed at reducing environmental triggers for childhood asthma. Drawing on the multiple theories and models described, a composite ecological stress process model is presented, and its implications for environmental health promotion discussed. PMID- 15296632 TI - Environmental health promotion interventions: considerations for preparation and practice. AB - Interventions to address current, future, and potential public health dilemmas, such as air pollution, urban sprawl, brown field reclamation, and threats of intentional toxic exposures would benefit from a synergy between the disciplines of environmental health and health education. A comparison between the Protocol for Assessing Community Excellence in Environmental Health and the PRECEDE PROCEED model used in health education illustrates some similarities and differences in terminology, assessment procedures, intervention design, and types of evidence used by the two disciplines. Promising intervention strategies draw on the expertise of both fields and include social action, policy and media advocacy, coalition building, organizational change, lay health advisers, risk communication, and tailored educational messages. Appropriate targets of change can range from the equitable distribution of resources to individual behavior change. Significant interdisciplinary evaluation research is necessary to accelerate the identification of successful models for reducing the burden of environmental health problems in communities. PMID- 15296633 TI - Environmental health promotion: progress and future opportunities. PMID- 15296636 TI - Development of a fluorescence polarization assay for the molecular chaperone Hsp90. AB - Heat shock protein 90 (Hsp90) is a molecular chaperone with essential functions in maintaining transformation, and there is increasing interest in developing Hsp90 inhibitors as cancer therapeutics. In this study, the authors describe the development and optimization of a novel assay for the identification of Hsp90 inhibitors using fluorescence polarization. The assay is based on the competition of fluorescently (BODIPY) labeled geldanamycin (GM) for binding to purified recombinant Hsp90alpha (GM is a natural product that binds to the ATP/ADP pocket in the amino terminal of Hsp90). The authors show that GM-BODIPY binds Hsp90alpha with high affinity. Even at low Hsp90alpha concentrations (30 nM), the measured polarization value is close to the maximum assay range of 160 mP, making measurements very sensitive. Its performance, as judged by signal-to-noise ratios (> 10) and Z and Z' values (> 0.5), suggests that this is a robust and reliable assay. GM, PU24FCl, ADP, and ATP, all known to bind to the Hsp90 pocket, compete with GM-BODIPY for binding to Hsp90alpha with EC(50)s in agreement with reported values. These data demonstrate that the Hsp90-FP-based assay can be used for high throughput screening in aiding the identification of novel Hsp90 inhibitors. PMID- 15296637 TI - Establishment and application of in vitro membrane potential assays in cell lines with endogenous or recombinant expression of ATP-sensitive potassium channels (Kir6.2/SUR1) using a fluorescent probe kit. AB - The flow of current through the adenosine triphosphate (ATP)-sensitive potassium channel (K(ATP)) of the isoform Kir6.2/SUR1 regulates the resting membrane potential in the pancreatic beta-cell. In combination with the cellular glucose metabolism, it is an important minute-to-minute regulator of insulin secretion and whole-body glucose homeostasis. The same K(ATP) isoform is further reported to be present in glucagon-secreting alpha-cells, intestinal L-cells, and glucose responsive neurons in the hypothalamus. All in all, this makes Kir6.2/SUR1 an interesting drug target. Using a commercially available fluorescent membrane potential probe kit and a conventional 96-well fluorescence plate reader, the authors have developed and established qualitative membrane potential assays used to screen for potassium channel closers (KCCs) and openers (KCOs) in insulin- and glucagon-secreting cell lines as well as in cells with recombinant expression of the human Kir6.2/SUR1 channel complex. Both glucose- and KCC-induced depolarization could be demonstrated. The magnitudes of these responses and KCO induced repolarization at high glucose displayed some variation between the different cell lines but a similar rank order of test compounds. Some cell types required the presence of a KCC, such as tolbutamide, to display significant effects of KCOs. The authors find that robust and reliable functional in vitro assays compatible with medium-throughput screening and high-throughput screening can be developed as a base for finding new, more potent, and isoform-selective KCCs and KCOs. PMID- 15296638 TI - High-throughput screening assay for identification of small molecule inhibitors of Aurora2/STK15 kinase. AB - STK15/Aurora2 is a centrosome-associated serine/threonine kinase, the protein levels and kinase activity of which rise during G2 and mitosis. STK15 overexpression induces tumorigenesis and is amplified in various human cancers and tumor cell lines. Thus, STK15 represents an important therapeutic target for small molecule inhibitors that would disrupt its activity and block cell proliferation. The availability of a robust and selective small molecule inhibitor would also provide a useful tool for identification of the potential role of STK15 in cell cycle regulation and tumor development. The authors report the development of a novel, fast, simple microplate assay for STK15 activity suitable for high-throughput screening. In the assay, gamma-(33)P-ATP and STK15 were incubated in a myelin basic protein (MBP)-coated FlashPlate(R) to generate a scintillation signal. The assay was reproducible, the signal-to-noise ratio was high (11) and the Z' factor was 0.69. The assay was easily adapted to a robotic system for drug discovery programs targeting STK15. The authors also demonstrate that STK15 is regulated by phosphorylation and the N-amino terminal domain of the protein. Treatment with phosphatase inhibitors (okadaic acid) or deletion of the N-amino terminal domain results in a significant increase in the enzymatic activity. PMID- 15296639 TI - Beta galactosidase enzyme fragment complementation as a high-throughput screening protease technology. AB - The authors describe a homogeneous, high-throughput screening (HTS) assay for measuring protease activity and detection of inhibitors. The assay comprises a cyclic beta-galactosidase (beta-gal) enzyme donor peptide (ED) containing a protease-selective cleavage sequence. Alone, the cyclic peptide is inactive, but when linearized following protease cleavage, ED complements with beta-gal enzyme acceptor forming active beta-gal enzyme. This then catalyzes the formation of either fluorescent or chemiluminescent products, with beta-gal turnover providing a highly amplified signal, and thus an assay technology of high sensitivity. To demonstrate the utility of the technology, an EFC assay was developed to measure the activity of 2, caspase 3 and beta-secretase. Using a cyclic ED containing the caspase 3 substrate sequence, DEVD, the EFC assay signal was linear with respect to caspase 3 concentration. The assay was very sensitive, being able to detect activity at low picogram amounts of caspase 3. For the beta-secretase (BACE) EFC assay, a cyclic ED containing the Swedish mutant cleavage site of amyloid precursor protein (APP), SEVNLDAEFK, was used. In a similar fashion to the caspase 3 assay, the signal induced by BACE activity was linear with respect to enzyme concentration and was highly sensitive, being able to detect nanogram quantities of BACE. The assay was also more sensitive than a commercially available FRET-based assay of BACE activity. It is concluded that the EFC protease assay is a simple, flexible, and sensitive technology for HTS of proteases. PMID- 15296640 TI - Two simple and generic antibody-independent kinase assays: comparison of a bioluminescent and a microfluidic assay format. AB - In this study, the authors have compared the performance of 2 high-throughput screening assays for a serin/threonine kinase: a microplate-based, bioluminescent assay that uses the luciferin/luciferase system to monitor ATP consumption, and a microfluidic assay that measures the change in mobility in an electric field of a fluorescently labeled peptide upon phosphorylation. Both assays are homogeneous, nonradioactive, antibody independent and could be miniaturized to a reaction volume of 4 microl. The robustness of both formats was demonstrated by Z' values > 0.8. Screening of a small library (2133 compounds) showed that the results obtained with both technologies correlate very well. Although the threshold for hits was set to a comparably low value-22.2% and 13.7% inhibition for the ATP consumption and microfluidic assay, respectively, corresponding to mean plus 3 standard deviations-the overlap of active compounds identified with the 2 assay formats was greater than 94%. Thus, both assays allow the identification of even low potency inhibitors with a high level of confidence. PMID- 15296641 TI - Miniaturization of intracellular calcium functional assays to 1536-well plate format using a fluorometric imaging plate reader. AB - The measurement of intracellular calcium response transients in living mammalian cells is a popular functional assay for identification of agonists and antagonists to receptors or channels of pharmacological interest. In recent years, advances in fluorescence-based detection techniques and automation technologies have facilitated the adaptation of this assay to 384-well microplate format high-throughput screening (HTS) assays. However, the cost and time required performing the intracellular calcium HTS assays in the 384-well format can be prohibitive for HTS campaigns of greater than 1 x 10(6) wells. For these reasons, it is attractive to miniaturize intracellular calcium functional assays to the 1536-well microplate format, where assay volumes and plate throughput can be decreased by several fold. The focus of the research described in this article is the miniaturization of an intracellular calcium assay to 1536-well plate format. This was accomplished by modifying the hardware and software of a fluorometric imaging plate reader (FLIPR) to enable transfer of nanoliters of test compound directly to a 1536-well assay plate, and measure the resulting calcium response from all 1536 wells simultaneously. An intracellular calcium functional assay against the rat muscarinic acetylcholine receptor subtype 1 (rmAchR1) G-protein coupled receptor (GPCR) was miniaturized and executed on this modified instrument. In experiments measuring the activity of known muscarinic receptor agonists and antagonists, the miniaturized FLIPR assay gave EC(50) and IC(50) values and rank order potency comparable to the 384-well format assays. Calculated Z' factors for the miniaturized agonist and antagonist assays were, respectively, 0.56 +/- 0.21 and 0.53 +/- 0.22, which were slightly higher (Z'(agonist) = 0.55 +/- 0.33) and lower (Z'(antagonist) = 0.70 +/- 0.18) than the corresponding values in the 384-well assays. A mock agonist HTS campaign against the muscarinic receptor in miniaturized format was able to identify all wells spiked with the rmAchR1 agonist carbachol. PMID- 15296642 TI - Identification of novel small-molecule inhibitors for human transketolase by high throughput screening with fluorescent intensity (FLINT) assay. AB - The metabolic enzyme transketolase (TK) plays a crucial role in tumor cell nucleic acid synthesis, using glucose through the elevated nonoxidative pentose phosphate pathway (PPP). Identification of inhibitors specifically targeting TK and preventing the nonoxidative PPP from generating the RNA ribose precursor, ribose-5-phosphate, provides a novel approach for developing effective anticancer therapeutic agents. The full-length human transketolase gene was cloned and expressed in Escherichia coli and the recombinant human transketolase protein purified to homogeneity. A fluorescent intensity (FLINT) assay was developed and optimized. Library compounds were screened in a high-throughput screening (HTS) campaign using the FLINT assay. Fifty-four initial hits were identified. Among them, 2 scaffolds with high selectivity, ideal physiochemical properties, and low molecular weight were selected for lead optimization studies. These compounds specifically inhibited in vitro TK enzyme activity and suppressed tumor cell proliferation in at least 3 cancer cell lines: SW620, LS174T, and MIA PaCa-2. Identification of these active scaffolds represents a good starting point for development of drugs specifically targeting TK and the nonoxidative PPP for cancer therapy. PMID- 15296643 TI - Screening assay for the identification of deoxyhypusine synthase inhibitors. AB - The 1st step in the posttranslational hypusine [N(epsilon)-(4-amino-2 hydroxybutyl)lysine] modification of eukaryotic translation initiation factor 5A (eIF5A) is catalyzed by deoxyhypusine synthase (DHS). The eIF5A intermediate is subsequently hydroxylated by deoxyhypusine hydroxylase (DHH), thereby converting the eIF5A precursor into a biologically active protein. Depletion of eIF5A causes inhibition of cell growth, and the identification of eIF5A as a cofactor of the HIV Rev protein turns this host protein and therefore DHS into an interesting target for drugs against abnormal cell growth and/or HIV replication. The authors developed a 96-well format DHS assay applicable for the screening of DHS inhibitors. Using this assay, they demonstrate DHS inhibition by AXD455 (Semapimod, CNI-1493). This assay represents a powerful tool for the identification of new DHS inhibitors with potency against cancer and HIV. PMID- 15296644 TI - High-throughput screening assays for the assessment of CYP2C9*1, CYP2C9*2, and CYP2C9*3 metabolism using fluorogenic Vivid substrates. AB - CYP2C9 is a genetically polymorphic human cytochrome P450 isozyme involved in the oxidative metabolism of many drugs, including nonsteroidal anti-inflammatory compounds. Individuals genotyped heterozygous or homozygous for CYP2C9 allelic variants have demonstrated altered metabolism of some drugs primarily metabolized by CYP2C9. The ability to expand screening of CYP2C9 allelic variants to a larger set of drugs and pharmaceutical agents would contribute to a better understanding of the significance of CYP2C9 polymorphisms in the population and to predictions of possible outcomes. The authors report the development of an in vitro fluorescence-based assay employing recombinant CYP2C9 variants (CYP2C9*1, CYP2C9*2, and CYP2C9*3) and fluorogenic Vivid(R) CYP2C9 substrates to explore the effects of CYP2C9 polymorphisms on drug metabolism, using drugs primarily metabolized by CYP2C9. Several chemically diverse fluorogenic substrates (Vivid(R) CYP2C9 blue, green, and red substrates) were used as prototypic probes to obtain in vitro CYP2C9 metabolic rates and kinetic parameters, such as apparent K(m), V(max), and V(max)/K(m) ratios for each allelic variant. In addition, a diverse panel of drugs was screened as assay modifiers with CYP2C9*1, CYP2C9*2, CYP2C9*3, and the fluorogenic Vivid(R) CYP2C9 substrates. The inhibitory potential of this large group of chemically diverse drugs and compounds has been assessed on the basis of their ability to compete with Vivid(R) CYP2C9 substrates in fluorescent reporter assays, thus providing a sensitive and quick assessment of polymorphism-dependent changes in CYP2C9 metabolism. PMID- 15296648 TI - Intracellular IFN-gamma production and IL-12 serum levels in latent autoimmune diabetes of adults (LADA) and in type 2 diabetes. AB - Th1 cytokines, such as interleukin-2 (IL-2) and interferon-gamma (IFN-gamma), and Th1-inducing cytokines, such as IL-12, are involved in the pathogenesis of various organ-specific autoimmune diseases, including autoimmune diabetes. In this study, we investigated intracellular IFN-gamma release by T lymphocytes and IL-12 serum levels in 48 type 2 and 36 latent autoimmune diabetes of adults (LADA) diabetics and 25 control subjects in an attempt to evaluate their role in the pathogenesis of these clinical entities. Ionomycin (ION) and phorbol-12 myristate-13-acetate (PMA)-activated peripheral blood mononuclear cells (PBMCs) were stained with anti-CD4-FITC or anti-CD8-FITC and anti-IFN-gamma phycoerythrin (PE) monoclonal antibodies (mAbs) and analyzed by flow cytometry. IL-12 serum levels were determined by enzyme-linked immunosorbent assay (ELISA). In all study groups, IFN-gamma content of CD4(+) and CD8(+) lymphocytes was significantly upregulated by stimulation. Furthermore, it was observed that CD4(+) and CD8(+) lymphocytes from type 2 diabetics produced significantly lower levels of IFN gamma compared with LADA patients and controls. However, the percentages of CD4(+)/IFN-gamma(+) and CD8(+)/IFN-gamma(+) cells from type 2 diabetics were significantly higher compared with controls. The flow cytometric picture of intracellular IFN-gamma release in LADA patients did not differ from that observed in controls. However, IL-12 serum levels in type 2 and LADA diabetics were lower than in controls. Because Th1 cytokines have been associated with the pathogenesis of autoimmune diabetes, these results preclude Th1 involvement in the autoimmune phenomena observed in LADA patients. In contrast, the low IFN gamma levels observed in type 2 diabetics in combination with the low IL-12 serum levels might be a contributing factor in the frequently observed chronic complications in these patients. PMID- 15296649 TI - Potent inhibition of SARS-associated coronavirus (SCOV) infection and replication by type I interferons (IFN-alpha/beta) but not by type II interferon (IFN-gamma). AB - We sought to investigate the anti-severe acute respiratory syndrome (SARS) associated coronavirus (SCoV) activities of type I (alpha and beta) and type II (gamma) interferons (IFN) in vitro. Type I IFNs protected cells from cytopathic effects (CPE) induced by SCoV, and inhibited viral genomic RNA replication in FRhk-4 cells (measured by quantitative RT-PCR) in a dose-dependent manner. Intracellular viral RNA copies were reduced 50% by IFN-alpha at a concentration of 25 U/ml and by IFN-beta at a concentration of 14 U/ml. IFN-gamma had fewer effects on inhibition of viral infection and replication. The type I IFN receptor signaling pathway in host cells is mainly involved in the inhibition of SCoV infection and replication. Type I IFNs could be used as potential agents for anti SARS treatment. PMID- 15296650 TI - Effects of the hepatitis C virus core protein on innate cellular defense pathways. AB - The hepatitis C virus (HCV) core protein is thought to contribute to HCV pathogenesis through its interaction with various signal transduction pathways. In this study, we explored the interaction of the core protein with innate defense pathways (interferon [IFN] regulatory factor [IRF], Jak-Stat, and inducible nitric oxide synthase [iNOS]) in HeLa and Huh7 human cell lines. Expression of a patient-derived genotype 1b core protein activated human IRF-1 and guanylate-binding protein-2 (GBP-2) promoters, induced IRF-1 mRNA, but failed to induce IRF-3 phosphorylation. HCV core protein caused dose-dependent induction of the IFN-beta promoter and IFN-beta mRNA but not the IFN-alpha1 and IFN-alpha4 promoters. In the presence of IFN-alpha, core expression was associated with increased IFN-stimulated gene factor 3 (ISGF3) binding to the IFN-stimulated response element (ISRE) and tyrosine phosphorylation of Stat1. Core expression resulted in dose-dependent activation of the ISRE and gamma activated sequence (GAS) promoters, in both the absence and the presence of either IFN-alpha or IFN gamma. Core stimulated the human iNOS promoter and induced iNOS protein. The data indicate that HCV core can modulate IRF, Jak-Stat, and iNOS pathways and suggest mechanisms by which core could affect HCV persistence and pathogenesis. PMID- 15296651 TI - MIP-1alpha induces differential MAP kinase activation and IkappaB gene expression in human B lymphocytes. AB - The chemokine macrophage inflammatory protein-1alpha (MIP-1alpha) stimulates migration of B cells and affects B cell immunoglobulin production. However, the molecular mechanisms by which MIP-1alpha modulates these biologic effects have not been completely defined. Previously, we demonstrated that treatment of B cells with MIP-1alpha induced the transcription factor, nuclear factor (NF) kappaB, to bind to DNA, concomitant with the degradation of IkappaBalpha, a cytoplasmic inhibitor of NF-kappaB activation. Here, we report that MIP-1alpha treatment of tonsil B cells induced IkappaB gene expression that was dependent on MIP-1alpha-mediated activation of a pathway(s) involving NF-kappaB and phosphatidylinositol-3 kinase (PI3K). The NF-kappaB pathway is understood to be controlled in an autoregulatory fashion, so expression of IkappaB is thought to provide a means by which B cells modulate this pathway after stimulation with MIP 1alpha. Although the idea of NF-kappaB autoregulation is not novel, this is the first report to suggest the regulation of B cell gene expression by MIP-1alpha. In addition, we observed the activation of Jun N-terminal kinase (JNK) and p38 mitogenic-activated protein kinase (MAPK), but not extracellular signal-related kinase (ERK) in response to MIP-1alpha. Although p38 and NF-kappaB activity were both necessary for B cell migration, IkappaB gene expression was not affected by p38 inhibition, suggesting that p38 is involved in a separate MIP-1alpha-mediated signal transduction pathway. PMID- 15296652 TI - Serum activity of prolyl endopeptidase, but not of dipeptidyl peptidase IV, is decreased by immunotherapy with IFN-alpha in high-risk melanoma patients. AB - Immunotherapy with interferon-alpha (IFN-alpha) induces neuropsychiatric side effects, most notably depression. In hepatitis patients treated with IFN-alpha, severity of depression correlates with a decrease in serum activity of dipeptidyl peptidase IV (DPP-IV, EC 3.4.14.5), a membrane-bound protease involved in the cleavage of cytokines and neuroactive peptides. Abnormal serum activity of the cytosolic peptidase prolyl endopeptidase (PEP, EC 3.4.21.26, postprolyl cleaving enzyme, prolyl oligopeptidase) has been documented in patients with a variety of psychiatric disorders, most consistently in mood disorders. The serum activity of PEP and DPP-IV was measured before and after 4 weeks of high-dose induction treatment with IFN-alpha in 18 patients with high-risk melanoma. In this exploratory study, we show a clear decrease in the serum activity of PEP after 4 weeks of treatment with IFN-alpha. This decrease was not related to changes in hematologic parameters. In contrast, serum activity of DPP-IV did not change. Further studies focusing on a possible role of PEP in the pathophysiology of IFN alpha-induced depression are warranted. PMID- 15296653 TI - Colony-stimulating factor-1 promotes clonogenic growth of normal murine colonic crypt epithelial cells in vitro. AB - The intestinal epithelium is a continuously renewing tissue. In the colon, stem cells are maintained at the base of highly organized crypts, where they undergo asymmetric division and give rise to daughter cells that proliferate and migrate up the crypt as they differentiate, then become senescent and are finally shed into the intestinal lumen. The growth factor requirements of fetal and prenatal colon cells for colony formation and that influence the establishment of cell lines from Immorto-mouse (Charles River, Wilmington, MA) transgenic embryos were explored. Single cell suspensions were isolated and cultured in a large range of growth factor combinations and conditions to determine their growth properties in soft agar. We report an important advance in the culture of mouse colonocytes by using macrophage colony-stimulating factor (CSF-1) and granulocyte-macrophage colony-stimulating factor (GM-CSF). A substantial proportion of colonies grown under low oxygen tension in the presence of CSF-1 and GM-CSF express intestinal epithelial A33 antigen, have the expected gene expression profile, including c fms and transcription factor c-myb, and show an appropriate epithelial cell morphology and undetectable CD45. Confocal microscopy on isolated crypts displays basolateral expression of c-Fms and E-cadherin on most epithelial cells. Fetal colon cultures from the Immorto-mouse with CSF-1 produced rapid outgrowth and readily established cell lines, in contrast to cultures without CSF-1. These observations have implications for the understanding of colon epithelial development and recovery following cytotoxic damage as well as providing a basis for the observation that some colon (and other epithelial) tumor cells respond to CSF-1 and GM-CSF. PMID- 15296655 TI - Deaf culture: when "positive" is a good thing. PMID- 15296656 TI - Perceptions of HIV/AIDS by deaf gay men. AB - Deaf gay men represent a subpopulation of the gay male community at particularly high risk for HIV/AIDS due to numerous barriers including language, stigma, and inequitable access to health services. The participants in this exploratory pilot study (N = 5) struggled with the ongoing threat of HIV infection and the pervasive nature of AIDS-related debilitation, death, and grief. Whether HIV infected or not, they described living at the intersection of multiple communities--the deaf, gay, and hearing--each characterized by unique communication styles, cultural expectations, and a propensity to marginalize outsiders. Health care providers were perceived as lacking compassion and largely ignorant to the needs of deaf persons, in general, and deaf gay men, in particular. Printed HIV materials were considered culturally inappropriate, incomprehensible, and ineffective. These findings suggest an extraordinary risk for adverse mental and physical health outcomes if care is not appropriately designed for this vulnerable population. PMID- 15296657 TI - Illness representation and smoking behavior: a focus group study of HIV-positive men. AB - Focus group interviews were conducted with 13 men living with HIV (mean age = 44.7) to explore their beliefs about cigarette smoking and HIV. Interview data were audiotaped, transcribed verbatim, and systematically analyzed using inductive techniques. Participants believed cigarette smoking provides a number of benefits to persons living with HIV. Although participants acknowledged that smoking has disadvantages, smokers generally discounted health risks, noting how it improves their sense of well-being and reasoning that they would not live long enough to suffer its consequences. Although smoking is a risk factor for HIV related morbidity and mortality, rates of smoking are high among men living with HIV. Research completed with other population groups finds beliefs are significant in explaining variance in smoking behavior change. Smoking-cessation programs targeting HIV-positive men may be more successful if illness-specific belief systems are taken into account. Additional study is warranted to substantiate the effectiveness of this approach. PMID- 15296658 TI - Successful techniques for retaining a cohort of infants and children born to HIV infected women: the prospective P2C2 HIV study. AB - Retaining subjects from disadvantaged populations in long-term studies is necessary to obtain high-quality data. This article presents cumulative retention rates from a 5-year prospective cohort study, the Pediatric Pulmonary and Cardiovascular Complications of Vertically Transmitted HIV Infection study. It also presents results of a cross-sectional qualitative survey about factors that induced caregivers to stay in the study. Although the repeated study visits were long and uncomfortable, cumulative retention among the 298 HIV-infected children was 80%. Incentives considered important by the caregivers included phone contact with nurse coordinators, nurse coordinators accompanying the caregiver and child during visits, phone reminders for appointments, help with scheduling, meals and transportation, access to health care, and relationships with staff. Thus, the high follow-up rate was in part due to nurses' efforts to reduce the study's burden on the families, provide tangible and intangible incentives, and establish personal relationships with families. PMID- 15296659 TI - Linking HIV/AIDS clients' self-care with outcomes. AB - The self-care practice of HIV/AIDS patients has become an important topic to help HIV/AIDS patients maintain their maximum level of well-being in chronic illness management. This article presents a self-care outcomes model that is applicable to HIV/AIDS nursing practice and research, and it identifies attributes and outcomes related to HIV/AIDS patients'self-care. The self-care outcomes model was developed based on the Outcomes Model for Health Care Research and literature review. Key variables related to HIV/AIDS self-care were summarized and discussed based on nine dimensions: client inputs, client processes, client outcomes, provider inputs, provider processes, provider outcomes, setting inputs, setting processes, and setting outcomes. This article reveals that self-care in HIV/AIDS is complex and may be influenced by many factors relating to individual, family, and health care system. More research with advanced multivariate statistical models and randomized controlled trial design will help determine the effectiveness of self-care strategies and interventions. PMID- 15296660 TI - Physical and leisure activity, body composition, and life satisfaction in HIV positive Hispanics in Puerto Rico. AB - Hispanics represent 13% of the U.S. population but account for 19% of the new AIDS cases reported in 2000. The antiretroviral drug therapy used for the treatment of HIV/AIDS may cause lipodystrophy and insulin resistance, among other effects. Physical and leisure activities reduce these effects and improve the emotional and physical well-being of HIV-positive persons. This study describes physical and leisure activities, life satisfaction, depression, and body composition of HIV-positive Hispanics in Puerto Rico and compares body composition, CD4 counts, depression, leisure time, and life satisfaction of participants classified as physically active or inactive. Sixty-eight individuals were evaluated using questionnaires and biophysical measurements. Descriptive statistics and independent t tests were used for data analysis. Physically active participants had higher life satisfaction scores and healthier body composition as compared to those physically inactive. Health professionals must encourage the promotion of a physically active lifestyle among HIV-positive Hispanics. PMID- 15296661 TI - Alternative modes of representation: there are no shortcuts. PMID- 15296662 TI - The significance of control models: intentional and unintentional effects. AB - Organization and control research has largely focused on the reform work and changes in the public sector. Many studies have focused on the reasons for reforms rather than on their significance. In this article, the authors deal with a control model applied in maternity health and based on remuneration for performance. At the end of the article, the authors note that formal control models are significant. Staff members have adapted to the model, but the model has also been adjusted to suit reality. This change has meant intentional effects, in the sense that the work has been adapted to suit the conditions given, but also unintentional effects in the form of attempts to outwit the system. PMID- 15296663 TI - Cultural models of illness and recovery in breast cancer support groups. AB - In an ethnographic study of breast cancer support groups for white women, the authors describe the cultural model of illness and recovery espoused by the groups and examine contested areas that might influence participation. Through analysis of interviews, observation at meetings, and program documents, they develop a model of group culture that includes five components: recovery narrative, group metaphors, perceived benefits, group processes, and contested domains. The recovery narrative focuses on optimism and personal growth, and members invoke metaphors of family and sisterhood to describe their relationship to the group. Linkages are made between perceived benefits and group processes. Contested domains challenge dominant features of the recovery narrative. The authors discuss the value of cultural studies of illness support groups for understanding member-group fit. PMID- 15296664 TI - Work and family as constituents of sense of coherence. AB - In this article, the authors focus on the intersection of the sense of coherence (SOC) and professional career in the light of qualitative data. They analyze the career descriptions written by young physicians in response to an open question in a survey questionnaire. In these narrative accounts, respondents who scored high on the SOC scale did not always orientate to work in a predictable way. Visiting narrative theoretization and feminist critique, the authors suggest a reflective survey style that seeks in the research process not so much an endeavor to reach a preexisting truth but, rather, a dialogue aimed at explaining the phenomenon at hand in a meaningful, agency-enabling way. PMID- 15296665 TI - Defining social support in context: a necessary step in improving research, intervention, and practice. AB - A substantial body of work on the concept of social support has resulted in many definitions, but none have been accepted as definitive. The lack of consensus about the definition of social support has resulted in a lack of consistency and comparability among studies. More important, the validity of any study attempting to measure or influence social support is undermined by the use of generic definitions, which lack contextual sensitivity. In this article concept analysis is used to evaluate definitions of social support to ascertain their utility for research. The authors argue that a contextualized approach to the definition of social support is necessary to improve clarity in research, and results in interventions or practices that are useful. They also assert that the development of a contextualized definition of social support requires qualitative methods to explore the meaning of social support with groups of people for whom intervention research is ultimately intended. PMID- 15296666 TI - Is quality of life a healthy concept? Measuring and understanding life experiences of older people. AB - The concept of quality of life has received considerable attention as an inclusive notion of health and as a basis for health interventions. The authors' argument in this article is that notwithstanding this attention, little consensus exists as to definition of the term. In addition, a focus on measurement has led to the neglect of wider aspects of quality of life. Such difficulties are particularly relevant to the study of quality of life of older people. Analysis of interview data suggests that older people's understandings of quality of life are not readily measurable and should be viewed in terms of phenomenological experience. The authors discuss the implications for studying quality of life of this group and difficulties for the concept itself. PMID- 15296667 TI - Premises, principles, and practices in qualitative research: revisiting the foundations. AB - In this keynote address, the author focuses on what we bring to qualitative inquiry and how we conduct our research. What we do, why we do it, and how we do it remain contested issues. She proposes that we look at our methodological premises anew, revisit our principles, and revise our practices. Throughout this address, she draws on Goffman's methodological insights to provide a foundation for reassessing qualitative inquiry. She argues that researchers can build on Goffman's ideas to strengthen their methodological practices and research products. Last, she counters current institutional scrutiny of qualitative inquiry and suggests unacknowledged benefits of this work. PMID- 15296668 TI - Issues of representation within qualitative inquiry. AB - Although qualitative inquiry has developed into a popularized and very useful way of conducting research within the health sciences, there has been a relatively disproportionate amount of literature devoted to "who" is represented in such inquiries. It is most often assumed that the end text should present an objective, value-free, and accurate representation of the participants and therefore exclude by all means the researcher's presence from the study. Although this might hold grounds for some inquiries, it is not necessarily the norm of a qualitative research. In this article, the author argues that the representation of the researcher in qualitative inquiries is inevitable, and the exclusion, or not, of the researcher from the text is a mere conventional agreement founded on a paradigmatic consensus. He concludes with the notion that there is a correlation between issues of representation and the researcher's stated epistemological and ontological assumptions. PMID- 15296669 TI - Qualitative methods and the development of clinical assessment tools. AB - Practitioners are under increased pressure to demonstrate effectiveness and to do so in as short a time as possible. In addition, evidence-based practice calls for practitioners to take into consideration not only best research evidence but also the wants and preferences of patients, and their own clinical expertise. In light of these demands, standardized assessment tools are likely to become increasingly central in clinical practice in a variety of health care fields, such as nursing, social work, psychiatry, psychology, and medicine. In this article, the author uses two case studies to show how qualitative methods can contribute to the development and evaluation of clinical assessment tools that are responsive to contemporary pressures. She also shows that evaluations of clinical instruments are incomplete if they do not include a qualitative component. PMID- 15296670 TI - Using drawings as a research method with adults. PMID- 15296671 TI - Cardiac-specific gene expression facilitated by an enhanced myosin light chain promoter. AB - BACKGROUND: Adenoviral gene transfer has been shown to be effective in cardiac myocytes in vitro and in vivo. A major limitation of myocardial gene therapy is the extracardiac transgene expression. METHODS: To minimize extracardiac gene expression, we have constructed a tissue-specific promoter for cardiac gene transfer, namely, the 250-bp fragment of the myosin light chain-2v (MLC-2v) gene, which is known to be expressed in a tissue-specific manner in ventricular myocardium followed by a luciferase (luc) reporter gene (Ad.4 x MLC250.Luc). Rat cardiomyocytes, liver and kidney cells were infected with Ad.4 x MLC.Luc or control vectors. For in vivo testing, Ad.4 x MLC250.Luc was injected into the myocardium or in the liver of rats. Kinetics of promoter activity were monitored over 8 days using a cooled CCD camera. RESULTS: In vitro: By infecting hepatic versus cardiomyocyte cells, we found that the promoter specificity ratio (luc activity in cardiomyocytes per liver cells) was 20.4 versus 0.9 (Ad.4 x MLC250.Luc vs. Ad.CMV). In vivo: Ad.4 x MLC250.Luc significantly reduced luc activity in liver (38.4-fold), lung (16.1-fold), and kidney (21.8-fold) versus Ad.CMV (p =.01); whereas activity in the heart was only 3.8-fold decreased. The gene expression rate of cardiomyocytes versus hepatocytes was 7:1 (Ad.4 x MLC.Luc) versus 1:1.4 (Ad.CMV.Luc). DISCUSSION: This new vector may be useful to validate therapeutic approaches in animal disease models and offers the perspective for selective expression of therapeutic genes in the diseased heart. PMID- 15296672 TI - Direct comparison of radiolabeled probes FMAU, FHBG, and FHPG as PET imaging agents for HSV1-tk expression in a human breast cancer model. AB - 2'-Deoxy-2'-fluoro-5-methyl-1-beta-D-arabinofuranosyluracil (FMAU), 9-(4-fluoro-3 hydroxy-methyl-butyl)guanine (FHBG) and 9-[(3-fluoro-1-hydroxy-2-propoxy)methyl] guanine (FHPG) have been evaluated in a human breast cancer model as potential radiotracers for PET imaging of HSV1-tk gene expression. In vitro accumulation of [14C]FMAU, [18F]FHBG, and [18F]FHPG in HSV1-tk-expressing cells was 14- to 16 fold (p <.001), 9- to 13-fold (p <.001), and 2- to 3-fold (p <.05) higher than tk negative control cells, respectively, between 30 and 240 min. Accumulation of FMAU and FHBG in vector-transduced cells was 10- to 14-fold and 6- to 10-fold higher than wild-type cells, respectively. At 2 hr, uptake of [(14)C]FMAU in tkpositive cells was 6.3-fold and 60-fold higher than [18F]FHBG and [18F]FHPG, respectively. In vivo, tumor uptake of [14C]FMAU in HSV1-tk-expressing cells was 3.7-fold and 5.5-fold (p <.001) higher than tk-negative control cells at 1 and 2 hr, respectively. Tumor uptake of [18F]FHBG was 4.2-fold and 12.6-fold higher (p <.001) than tk-negative cells at the same time points. Incorporation of [14C]FMAU in tk-positive tumor was 18-fold and 24-fold higher (p <.001) than [18F]FHBG at 1 and 2 hr, respectively. Micro-PET images support the biodistribution results and indicate that both [18F]FMAU and [18F]FHBG are useful for imaging HSV1-tk expression in breast cancer. Although FMAU demonstrates higher total incorporation (%dose/g) in tumor tissue compared with the other tracers, FHBG is superior in terms of specific accumulation in transfected cells at later time points. PMID- 15296673 TI - Cellular imaging at 1.5 T: detecting cells in neuroinflammation using active labeling with superparamagnetic iron oxide. AB - The ability to visualize cell infiltration in experimental auto-immune encephalomyelitis (EAE), a well-known animal model for multiple sclerosis in humans, was investigated using a clinical 1.5-T magnetic resonance imaging (MRI) scanner, a custom-built, high-strength gradient coil insert, a 3-D fast imaging employing steady-state acquisition (FIESTA) imaging sequence and a superparamagnetic iron oxide (SPIO) contrast agent. An "active labeling" approach was used with SPIO administered intravenously during inflammation in EAE. Our results show that small, discrete regions of signal void corresponding to iron accumulation in EAE brain can be detected using FIESTA at 1.5 T. This work provides early evidence that cellular abnormalities that are the basis of diseases can be probed using cellular MRI and supports our earlier work which indicates that tracking of iron-labeled cells will be possible using clinical MR scanners. PMID- 15296674 TI - Micro-PET imaging of alphavbeta3-integrin expression with 18F-labeled dimeric RGD peptide. AB - The alphav integrins, which act as cell adhesion molecules, are closely involved with tumor invasion and angiogenesis. In particular, alphavbeta3 integrin, which is specifically expressed on proliferating endothelial cells and tumor cells, is a logical target for development of a radiotracer method to assess angiogenesis and anti-angiogenic therapy. In this study, a dimeric cyclic RGD peptide E[c(RGDyK)]2 was labeled with 18F (t(1/2) = 109.7 min) by using a prosthetic 4 [18F]fluorobenzoyl moiety to the amino group of the glutamate. The resulting [18F]FB-E[c(RGDyK)]2, with high specific activity (200-250 GBq/micromol at the end of synthesis), was administered to subcutaneous U87MG glioblastoma xenograft models for micro-PET and autoradiographic imaging as well as direct tissue sampling to assess tumor targeting efficacy and in vivo kinetics of this PET tracer. The dimeric RGD peptide demonstrated significantly higher tumor uptake and prolonged tumor retention in comparison with a monomeric RGD peptide analog [18F]FB-c(RGDyK). The dimeric RGD peptide had predominant renal excretion, whereas the monomeric analog was excreted primarily through the biliary route. Micro-PET imaging 1 hr after injection of the dimeric RGD peptide exhibited tumor to contralateral background ratio of 9.5 +/- 0.8. The synergistic effect of polyvalency and improved pharmacokinetics may be responsible for the superior imaging characteristics of [18F]FB-E[c(RGDyK)]2. PMID- 15296675 TI - Fluorescently labeled adenovirus with pIX-EGFP for vector detection. AB - Adenoviruses are extensively studied in terms of their use as gene therapy vectors and pathogenesis. These vectors have been targeted on both transcriptional and transductional levels to achieve cell-specific gene delivery. Current detection strategies, including reporter gene expression, viral component detection, and vector labeling with fluorophores, have been applied to analyze adenoviral vectors; however, these methods are inadequate for assessing transductional targeting. As an alternative to conventional vector detection techniques, we developed a specific genetic labeling system whereby an adenoviral vector incorporates a fusion between capsid protein IX and EGFP. DNA packaging and thermostability were marginally hampered by the modification while DNA replication, cytopathic effect, and CAR-dependent binding were not affected. The fluorescent label was associated with the virus capsid and conferred a fluorescent property useful in detecting adenoviral particles in flow cytometry, tracking, and tissue sections. We believe our genetic adenovirus labeling system has important implications for vector development, detecting adenovirus vectors in targeting schemes, and studying adenovirus biology. In addition, this technique has potential utility for dynamic monitoring of adenovirus replication and spread. PMID- 15296676 TI - Validating bioluminescence imaging as a high-throughput, quantitative modality for assessing tumor burden. AB - Bioluminescence imaging (BLI) is a highly sensitive tool for visualizing tumors, neoplastic development, metastatic spread, and response to therapy. Although BLI has engendered much excitement due to its apparent simplicity and ease of implementation, few rigorous studies have been presented to validate the measurements. Here, we characterize the nature of bioluminescence output from mice bearing subcutaneous luciferase-expressing tumors over a 4-week period. Following intraperitoneal or direct intratumoral administration of luciferin substrate, there was a highly dynamic kinetic profile of light emission. Although bioluminescence was subject to variability, strong correlations (r >.8, p <.001) between caliper measured tumor volumes and peak light signal, area under light signal curve and light emission at specific time points were determined. Moreover, the profile of tumor growth, as monitored with bioluminescence, closely resembled that for caliper measurements. The study shows that despite the dynamic and variable nature of bioluminescence, where appropriate experimental precautions are taken, single time point BLI may be useful for noninvasive, high throughput, quantitative assessment of tumor burden. PMID- 15296678 TI - Measuring fatigue in sarcoidosis: the Fatigue Assessment Scale (FAS). AB - Fatigue is a major problem in a wide range of diseases including sarcoidosis. However, there is no standard measure for assessing fatigue. Therefore, the aim of the present study was to evaluate the usefulness of the Fatigue Assessment Scale (FAS) in two samples of sarcoidosis patients. Sample 1 included 1 046 members of the Dutch Sarcoidosis Society and Sample 2 consisted of 80 sarcoidosis patients of the outpatient clinic of the Sarcoidosis Management Centre Maastricht, the Netherlands. All patients completed the FAS as well as the 'energy and fatigue' subscale of the WHOQOL-100. Additionally, the participants of Sample 1 filled in the Beck Depression Inventory (BDI). In addition, 241 patients of Sample 1 completed the FAS for the second time after a one-week interval. The FAS appeared to be a unidimensional scale. The content validity, construct validity and internal consistency of the FAS were good. The test - retest reliability was.89. Four FAS items appeared to have a gender bias: three items were uniformly biased and one item non-uniformly biased. Correction for gender bias in the calculation of the FAS total score is not indicated. In conclusion, the FAS is a promising measure for assessing fatigue in sarcoidosis patients. PMID- 15296679 TI - Conscientiousness, emotional stability, perceived control and the frequency, recency, rate and years of blood donor behaviour. AB - OBJECTIVES: Personality factors were used to explore aspects of blood donor behaviour (a form of prosocial volunteer behaviour). Based on the Big 5 model of personality, a number of mediation (via perceived control) and moderation hypotheses linking personality to aspects of blood donor behaviour (frequency, recency, rate and years of past behaviour) were explored. DESIGN: A cross-section quasi-experimental survey of 630 blood donors. METHOD: Prior to donation, 630 donors completed measures of the Big 5 personality domains and donor history (frequency, recency, rate and years of past behaviour); after donation, they indicated levels of perceived procedural and emotional control. RESULTS: There was no evidence that perceived control mediates the relationship between personality and behaviour. However, donors were able to express control in a low control context. For males, the relationship between years as a donor and both the frequency and the rate of donation were stronger for those higher in conscientiousness (C), whereas for females, emotional stability (ES) acted as the moderator. Recency and frequency of past behaviour were orthogonal. DISCUSSION: The results extend previous work by showing that C (for males) and ES (for females) help to sustain behaviour over time. The recency and frequency of past behaviour assess qualitatively different aspects of past behaviour and this should be explored in future studies. These results have important practical implications for designing donor recruitment and retention interventions. PMID- 15296677 TI - Ultrasonic analysis of peptide- and antibody-targeted microbubble contrast agents for molecular imaging of alphavbeta3-expressing cells. AB - The goal of targeted ultrasound contrast agents is to significantly and selectively enhance the detection of a targeted vascular site. In this manuscript, three distinct contrast agents targeted to the alphavbeta3 integrin are examined. The alphavbeta3 integrin has been shown to be highly expressed on metastatic tumors and endothelial cells during neovascularization, and its expression has been shown to correlate with tumor grade. Specific adhesion of these contrast agents to alphavbeta3-expressing cell monolayers is demonstrated in vitro, and compared with that of nontargeted agents. Acoustic studies illustrate a backscatter amplitude increase from monolayers exposed to the targeted contrast agents of up to 13-fold (22 dB) relative to enhancement due to control bubbles. A linear dependence between the echo amplitude and bubble concentration was observed for bound agents. The decorrelation of the echo from adherent targeted agents is observed over successive pulses as a function of acoustic pressure and bubble density. Frequency-domain analysis demonstrates that adherent targeted bubbles exhibit high-amplitude narrowband echo components, in contrast to the primarily wideband response from free microbubbles. Results suggest that adherent targeted contrast agents are differentiable from free floating microbubbles, that targeted contrast agents provide higher sensitivity in the detection of angiogenesis, and that conventional ultrasound imaging techniques such as signal subtraction or decorrelation detection can be used to detect integrin-expressing vasculature with sufficient signal-to-noise. PMID- 15296680 TI - Constructing an integrated model of the antecedents of adolescent smoking. AB - OBJECTIVE: Reviews have called for integrative, theoretically informed models of the 'hundreds of associations' (Miller & Slap, 1989, p. 131) between psychosocial measures and adolescent smoking (e. g. Tyas & Pederson, 1998). Such a model was tested. METHOD: A prospective, classroom-based survey measuring previously identified correlates of smoking allowed comparison of the strength of relationships between antecedents and smoking status six months later. The prospective sample included 225 13 to 14-year-olds. Measures of behaviour specific cognitions derived from the Theory of Planned Behaviour (TPB)-as well as personality, self-esteem, parental support and parental control, sociodemographic factors, and descriptive norms-were included. Relationships between antecedents were explored using path analyses. RESULTS: High initial rates of smoking were observed. Of the variance in smoking six months later, 56% was explained by seven direct predictors: intention, perceived ease of smoking, estimated number of friends smoking, percentage of older brothers smoking, self-esteem, extraversion and car access. DISCUSSION: Results emphasize the importance of behaviour specific cognitions specified by the TPB but suggest that other factors, including extraversion and self-esteem, need to be included in models of the antecedents of smoking. The findings also imply that some antecedents, such as parental support, may indirectly influence adolescent smoking through their effect on other variables. PMID- 15296681 TI - The readiness to change questionnaire: reliability and validity of a Swedish version and a comparison of scoring methods. AB - OBJECTIVES: The aim of this study was to investigate the relative merits of three methods of scoring the Swedish version of the Readiness to Change Questionnaire (RTCQ), either by assigning a stage in the Prochaska and DiClemente (1986) Stages of Change Model or by treating the scores as a continuous readiness to change variable. Assigning a stage of change was achieved with both the quick method and the refined method. DESIGN AND METHODS: Out of 563 patients screened at an emergency surgical ward for risky alcohol consumption, 165 met risk criteria and responded to the RTCQ. The three scoring methods were examined with regard to internal consistency, test - retest reliability, construct and predictive validity. RESULTS: All three methods of treating the RTCQ scores had satisfactory reliability. Since stages of change (quick method) were significantly but modestly correlated to alcohol consumption and to change-related behaviours at intervention, and moderately correlated to alcohol problems, the quick method had reasonable construct validity. The refined method had higher construct validity; however, this method left 32% of the patients without a stage assignment. The continuous readiness scale had higher construct validity than the quick method, but was not in par with the refined method. No scoring method was found to have predictive validity. CONCLUSIONS: The RTCQ scores treated as a continuous readiness scale were a viable alternative to the original ways of assigning a stage of change to a patient. The Swedish RTCQ is reliable and has reasonable construct validity, but its predictive validity needs further investigation. PMID- 15296682 TI - Evaluation of a model of adjustment to an iatrogenic hepatitis C virus infection. AB - OBJECTIVE: The aim of this study was to evaluate a model of adjustment to an iatrogenic hepatitis C(HCV) infection in a cohort of women. PARTICIPANTS: Eighty three women diagnosed with an iatrogenic HCV infection were recruited; 49 women had chronic infection (PCR positive) and the remaining 34 women were considered to have a self-limiting HCV infection (PCR negative). MEASURES: The Hepatitis C Survey Questionnaire (HCSQ; Coughlan, Sheehan, Carr, & Crowe, unpublished) and the General Health Questionnaire (GHQ30; Goldberg & Williams, 1988) were used in this study. RESULTS: Structural equation modelling (SEM) was carried out to evaluate and modify a recursive path model using Moos and Schaefer's (1984) model of coping with illness as the basis for developing a multivariate model of adjustment to an iatrogenic HCV infection. The final model fit, chi(2)(30) = 21.9 p =.86, CFI = 1.00, RMSEA = 0.000, was judged to be theoretically acceptable, indicating that positive illness appraisal, ability to work and negative behaviour as a consequence of feelings of anger and blame are directly related to adjustment. CONCLUSION: This model has provided support for the following general relationships, namely that, adjustment to an iatrogenic HCV infection is related to: (1) illness and social factors; (2) cognitive appraisals; (3) adaptive tasks; and (4) coping skills thus emphasizing the need to develop a biopsychosocial model of treatment. PMID- 15296683 TI - Effects of a tailored lifestyle self-management intervention in patients with type 2 diabetes. AB - OBJECTIVES: The aim of the present study was to develop, implement and evaluate a brief intervention to improve adherence to the recommended lifestyle changes for patients with Type 2 diabetes, in particular to help patients to reduce the total amount of fat consumed and to increase lifestyle physical activity levels. DESIGN AND METHOD: A brief, tailored lifestyle self-management intervention for patients with Type 2 diabetes was evaluated in a randomized controlled trial. One hundred participants (aged 40 - 70 yrs) completed assessments at three time points- baseline, three months and one year. Participants were allocated to either an intervention group who received the brief tailored intervention including follow up telephone calls, or a usual care control group. RESULTS: Results indicate that the intervention was successful in helping patients to reduce fat intake and, to a lesser extent, increase lifestyle physical activity levels. These self-reported changes in behaviour were reflected in the objective data with weight maintenance in the intervention group compared to the control group, together with a significant reduction (2 cm) in waist circumference. CONCLUSIONS: These results provide further evidence of the effectiveness of tailored interventions for lifestyle change. PMID- 15296684 TI - Cognitive-behavioural rehabilitation programme for patients with an implanted cardioverter defibrillator: a pilot study. AB - OBJECTIVES: The effectiveness of a comprehensive 12-week CR programme for ICD patients was evaluated. DESIGN: All surviving and suitable ICD patients being cared for by a regional implantation centre were invited to attend a 12-week cognitive-behavioural cardiac rehabilitation programme that had been modified to meet the needs of this group. Patients assenting were randomized to either an immediate treatment or a waiting treatment group. Measures were taken prior to randomization, at the end of the treatment or waiting period, at the end of the second treatment group for that group only and at three months post-treatment for both groups. OUTCOME MEASURES: The Hospital Anxiety and Depression Scale, the Total Concerns Questionnaire, the Quality of Life after Myocardial Infarction Questionnaire, the EuroQual (subjective health rating scale), the Shuttle Test and a number of ICD shocks and ATP episodes were used in this study. RESULTS: For those patients willing and able to attend, the cognitive-behavioural CR programme produced significant benefits in terms of psychological and functional adaptation to living with the device. CONCLUSIONS: A comprehensive 12-week CR programme that incorporated both psychological and exercise-based components significantly reduced anxiety and depression and improved quality of life of ICD patients. It is not clear if these benefits are sustained. PMID- 15296685 TI - Music can facilitate blood pressure recovery from stress. AB - OBJECTIVES: Interventions that reduce the magnitude of cardiovascular responses to stress are justified, at least in part, by the notion that exaggerated responses to stress can damage the cardiovascular system. Recent data suggest that it is worthwhile to explore, in addition to the magnitude of the cardiovascular responses during stress (reactivity), the factors that affect the return to baseline levels after the stressor has ended (recovery). This experiment examined the effect of listening to music on cardiovascular recovery. DESIGN AND METHOD: Participants (N = 75) performed a challenging three-minute mental arithmetic task and then were assigned randomly to sit in silence or to listen to one of several styles of music: classical, jazz or pop. RESULTS: Participants who listened to classical music had significantly lower post-task systolic blood pressure levels (M = 2.1 mmHg above pre-stress baseline) than did participants who heard no music (M = 10.8 mmHg). Other musical styles did not produce significantly better recovery than silence. CONCLUSIONS: The data suggest that listening to music may serve to improve cardiovascular recovery from stress, although not all music selections are effective. PMID- 15296686 TI - Investigation of the interactive effects of gender and psychological factors on pain response. AB - OBJECTIVES: There is growing evidence to suggest that certain psychological modulators of pain sensitivity are dependent on gender. The aim of the present study was to examine further whether cognitive-affective factors (with specific focus on situational anxiety) shown to modulate pain report and behaviour have differential effects on men's and women's response to experimentally induced pain. METHOD: A sample of 80 healthy university students (40 women, 40 men) was assessed on subjective measures of anxiety (situational and dispositional), anxiety-sensitivity and attitudes toward the experimental pain procedure prior to being exposed to a cold pressor test (constant temperature + 1 degrees C; +/- 1 degrees C, cut-off limit 240 s). RESULTS: The present study produced three main findings: (1) No effect of gender was found on any of the pain measures; however, an interactive effect of anxiety and gender was found on measures of pain tolerance with low anxiety men displaying significantly higher pain tolerance than both low and high anxiety women. (2) Men classified as low-anxious tolerated cold pressor pain significantly longer than men classified as high-anxious, while no such effect of anxiety on pain response was observed in women. (3) Gender specific associations were observed between other psychological variables (including anxiety-sensitivity and attitudes toward pain) and thermal pain response. CONCLUSION: Results from the present study suggest that there are important differences in the way cognitive-affective factors impact on the pain response of men and women. Further research is needed to explore potential psychosocial and physiological mechanisms that may underlie such differences. PMID- 15296687 TI - Approaching the salutogenesis of sense of coherence: the role of 'active' self esteem and coping. AB - OBJECTIVES: The aim of this work was to investigate the health relevant status of sense of coherence (SOC). First, its relation to general health was studied. Secondly, the importance of self-esteem (SE) type, locus of control, 'fighting spirit' and coping style for SOC was explored. It was predicted that SOC has relevance for good health and that active types of self-esteem, together with the other adaptive dispositions, play a role in SOC. DESIGN: A cross-sectional, retrospective study was used. METHODS: The personality and health questionnaires were administered to 409 undergraduates. The data were analysed using correlations and hierarchical multiple regression methods. RESULTS: The correlational and regression analyses indicated that SOC has more importance for health than the other more specific adaptive styles. They further revealed that, although negative affect was a main predictor of SOC and health, an active type of SE (a combination of high basic and high earning SE) and locus of control were associated with a high SOC. CONCLUSIONS: It is concluded that SOC has a unique relation to general health, and although SOC strongly reflects affective traits, the concept seems also to mirror more dynamic dispositions, referring to an active self-esteem structure and self-determination. PMID- 15296688 TI - Prevention research in women's health: studies from the field--introduction. PMID- 15296689 TI - The impact of a promotora on increasing routine chronic disease prevention among women aged 40 and older at the U.S.-Mexico border. AB - A randomized controlled intervention tested the effectiveness of a community health worker (CHW) program in increasing compliance with annual preventive exams among uninsured Hispanic women living in a rural U.S.-Mexico border area. During 1999-2000, household surveys were administered to women aged 40 and older. Uninsured women not receiving routine comprehensive preventive care were invited to participate in a free comprehensive clinical exam. Participants in the initial exam were eligible to participate in the CHW (promotora) intervention. Women were randomized to one of two intervention arms. One arm received a post-card reminder for an annual preventive exam, the other a postcard reminder and follow-up visit by a promotora. Receiving the promotora intervention was associated with a 35% increase in rescreening over the postcard-only reminder (risk ratio [RR] = 1.35, 95% confidence interval 0.95-1.92). Using promotoras to increase compliance with routine screening exams is an effective strategy for reaching this female population. PMID- 15296690 TI - Stepped-care, community clinic interventions to promote mammography use among low income rural African American women. AB - Few studies have investigated community clinic-based interventions to promote mammography screening among rural African American women. This study randomized older low-income rural African American women who had not participated in screening in the previous 2 years to a theory-based, personalized letter or usual care; no group differences in mammography rate were evident at 6-month follow-up. Women who had not obtained a mammogram were then randomized to a tailored call delivered by community health care workers or a tailored letter. There were no group differences in mammography rates after the second 6-month follow-up. However, among women who had never had a mammogram, the tailored call was more effective in promoting mammography use. Tailored counseling may be an effective screening promotion strategy for hard-to-reach rural African American women with no history of screening. Further research into this strategy may facilitate efforts to reduce health disparities in underserved low-income rural African American populations. PMID- 15296691 TI - Walking patterns in a sample of African American, Native American, and Caucasian women: the cross-cultural activity participation study. AB - This analysis describes walking patterns among African American, Native American, and Caucasian women from South Carolina and New Mexico. Walking was assessed using pedometer and physical activity (PA) record data based on 4 consecutive days on either three (Study Phase 1) or two (Study Phase 2) occasions. Participants walked 5,429 +/- 2,959 steps per day and recorded 159 +/- 59 minutes per day of total walking in the PA record. Most daily walking was accumulated during household (46%), transportation (26%), occupation (16%), and exercise related (10%) walking. There was a modest correlation between steps per day and minutes per day. Steps per day were higher with education and household size, and lower with increasing age and body mass index. These findings have implications for developing PA surveys and for planning interventions related to walking patterns among women. PMID- 15296692 TI - Sociodemographic, behavioral, and psychological correlates of current overweight and obesity in older, urban african american women. AB - To better understand obesity and overweight among urban African American women, the authors examined sociodemographic, behavioral, and psychological factors within body mass index (BMI) categories. A total of 496 women were recruited for cardiovascular risk factor screening from 20 urban African American churches. Study participants had a mean age of 52.8 years, 13.5 years of education, and an average BMI of 32 kg/m2. Bivariate analyses showed increased overall energy intake and decreased physical performance on a walk test, and general well-being declined as the BMI class increased; obese women had the lowest physical performance and well-being levels and the highest energy intake levels. There was no difference by BMI category, however, in social variables such as educational attainment, employment, marital status, or household income. This study suggests that although women with increasing BMI have some physical and well-being concerns, the major social variables are not differentially distributed by BMI in this sample of women. PMID- 15296693 TI - Disaster down East: using participatory action research to explore intimate partner violence in eastern North Carolina. AB - In the aftermath of Hurricane Floyd in 1999, a Community Advisory Committee requested assistance from its university partners (University of North Carolina) to address stress and increased risk for intimate partner violence (IPV). Collected from 12 study work sites, baseline data indicated that IPV rates were higher among blue-collar women in eastern North Carolina than national population based rates suggest. IPV victims reported higher levels of perceived stress, psychological distress, somatic complaints, and post-traumatic stress disorder (PTSD) symptoms than did nonvictimized coworkers. As for the relationship of the flood to IPV, no significant increase in IPV incidence occurred after the flood. Regardless of their flood experience, however, IPV victims consistently reported greater stress, PTSD symptoms, and somatic and psychological problems. Moreover, IPV victims may be at higher risk for stress-mediated chronic illnesses and for using negative coping behaviors. This study uses an established trusting relationship between researchers and community members to explore community needs and inform intervention design. PMID- 15296694 TI - Relationship characteristics and sexual practices of African American adolescent girls who desire pregnancy. AB - This study examined associations between African American adolescent girls' desire to become pregnant and their sexual and relationship practices. Odds ratios and 95% confidence intervals were used to detect significant associations between pregnancy desire and the assessed correlates. Of 522 participants (14 to 18 years old), 67 (12.8%) were pregnant and were thus excluded from this analysis. Of the remaining 455 adolescents, 107 (23.6%) expressed some desire to be pregnant at the time of assessment. Adolescents who desired pregnancy were significantly more likely to report having had sex with a casual partner and to use contraception inconsistently. Factors involving an adolescent girl's relationship with her partner (e.g., being in a relationship, length of relationship, time spent with boyfriend, or satisfaction with boyfriend) were not significantly associated with the desire for pregnancy. Effective pregnancy and sexually transmitted disease prevention programs for female adolescents should address their level of pregnancy desire. PMID- 15296697 TI - Therapeutic reduction of coronary atheromatous plaque burden using bioengineered apoA-I Milano. PMID- 15296698 TI - Lipoprotein lipase and its role in regulation of plasma lipoproteins and cardiac risk. AB - For over 50 years, biologists and clinicians have studied lipoprotein lipase (LPL) and learned about its structure, function, cellular production, physiology, and human genetics. LPL is the principal enzyme that removes triglyceride from the bloodstream. It also determines plasma levels of high-density lipoprotein. Surprisingly, within the past several years, a number of new and unexpected proteins have been discovered that regulate the actions of LPL. These include the very low-density lipoprotein receptor, angiopoetin-like protein 3, and apolipoprotein A-V. In addition, mouse genetic studies have confirmed tissue culture findings of nonenzymatic roles of LPL both in lipid metabolism and atherogenesis. These basic observations are now being related to new information on human genetic polymorphism in this gene that is likely to affect clinical evaluation of lipoprotein disorders and cardiac risk. PMID- 15296699 TI - Hepatic lipase: friend or foe and under what circumstances? AB - Hepatic lipase (HL) plays a role in the metabolism of chylomicron and very low density lipoprotein remnants, low-density lipoproteins (LDL), and high-density lipoproteins (HDL), which are all implicated in atherosclerosis. Considering the effects of HL on these lipoproteins, it appears that HL has pro- as well as antiatherogenic potential. In line with clinical observations, most effects of HL on lipoprotein metabolism during hypertriglyceridemia may be interpreted as promoting atherosclerosis (formation of small, dense LDL, lowering of HDL levels), whereas most effects during hypercholesterolemia seem to be potentially antiatherogenic (stimulation of reverse cholesterol transport, clearing of intermediate-density lipoprotein). The potential modulation of pro- or antiatherogenics effect of HL by other factors, such as LDL receptor, cholesterol ester transfer protein, lipoprotein lipase, and ATP-binding cassette A-1 activity, is discussed. PMID- 15296700 TI - Low-density lipoprotein reduction in high-risk patients: how low do you go? AB - Cardiovascular disease and its clinical sequelae remain the leading causes of morbidity and mortality in many regions of the world. Dyslipidemia is a critical risk factor to intercept in both the primary and secondary prevention of acute cardiovascular events. The prospective, placebo-controlled clinical trials conducted with statins over the course of the past 15 years have conclusively demonstrated that these drugs significantly reduce risk for fatal and nonfatal myocardial infarction, ischemic stroke, unstable angina, and frequency of myocardial ischemia, as well as cardiovascular and all-cause mortality. Of considerable interest is the fact that, even under the exquisitely controlled circumstances of a clinical trial, endpoint reductions in these trials typically occur in the range of 20% to 35%. Understandably, much attention is now being focused on deriving the pharmacologic means by which to further increase the magnitude of endpoint reduction. Epidemiologic investigation has demonstrated that the relationship between cholesterol and risk for atherosclerotic disease is a continuous one. Consequently, it is reasonable to assume that more aggressive reductions of low-density lipoprotein (LDL) cholesterol might result in even greater reductions of cardiovascular event rates and atheromatous plaque progression than heretofore observed. Two recent clinical trials, Reversal of Atherosclerosis with Aggressive Lipid Lowering (REVERSAL) and Pravastatin or Atorvastatin Evaluation and Infection Therapy (PROVE IT), prospectively tested and confirmed the validity of more aggressive LDL cholesterol lowering in high risk patients with established coronary artery disease. PMID- 15296701 TI - ASCOT-LLA and the primary prevention of coronary artery disease in hypertensive patients. AB - Although each revision of the US National Cholesterol Education Program guidelines has made increasing provision for the use of global risk assessment in determining need for and intensity of therapy, the guidelines' continued focus on low-density lipoprotein (LDL) cholesterol may result in inadequate or no treatment for individuals at high risk for coronary artery disease (CAD) who do not have substantially elevated LDL cholesterol. However, recent clinical trial evidence has shown that high-risk patients benefit from lipid-regulating therapy regardless of LDL cholesterol level. In the Anglo-Scandinavian Cardiac Outcomes Trial--Lipid Lowering Arm, high-risk hypertensive patients had reductions in clinical events despite not having substantially elevated LDL cholesterol at baseline. These results suggest that all hypertensive patients with additional risk factors should receive lipid-regulating statin therapy to prevent CAD events. PMID- 15296702 TI - High-density lipoprotein subfractions and risk of coronary artery disease. AB - Numerous studies have shown that levels of high-density lipoprotein (HDL) cholesterol are inversely related to coronary artery disease risk. The HDL subfractions, however, seem to differ in their capacity to confer protection, with the large HDL2 subfraction appearing to be more important than the small HDL3 subfraction. Lipid-modifying drugs differ in their HDL-raising efficacy, and they also differ in how they affect HDL subfractions. Clinical trials show that raising total HDL cholesterol improves clinical and angiographic outcomes. It remains to be determined, however, whether a shift in distribution of HDL particles provides greater benefit than just an increase in total HDL. PMID- 15296703 TI - Lessons learned from the prospective pravastatin pooling project. AB - The Prospective Pravastatin Pooling (PPP) Project was a prospectively defined collaboration of three randomized, placebo-controlled, long-term, monotherapy (40 mg/d) trials of the lipid-lowering agent pravastatin. A pooled database of 19,768 participants with a mean of 5.2 years of follow-up was created, providing the investigators with over 100,000 patient-years of experience to address questions on total and cause-specific mortality, coronary incidence, stroke, and safety. One trial (West of Scotland Coronary Prevention Study) was primary prevention and two (Cholesterol and Recurrent Events and Long-term Intervention with Pravastatin in Ischemic Disease) were secondary prevention. Pravastatin was shown to safely reduce all-cause mortality, fatal and nonfatal coronary events, and stroke events in patients with a broad range of patient characteristics. PMID- 15296704 TI - ALLHAT-LLT: questions, questions, and more questions (and some answers). AB - The Antihypertensive and Lipid-Lowering Treatment to Prevent Heart Attack Trial- Lipid Lowering Trial (ALLHAT-LLT) compared 40 mg/d of pravastatin with usual care among 10,355 men and women aged 55 years or older with stage 1 or 2 hypertension, at least one additional coronary heart disease risk factor, and low-density lipoprotein (LDL) cholesterol levels of 120 to 189 mg/dL. After a mean of 4.8 years of treatment and follow-up, the difference in total cholesterol between the two arms was 9.6%, whereas in a small, nonrandomized subsample, the LDL cholesterol differential was 16.7%. No differences were observed between the pravastatin and usual-care groups with respect to all-cause mortality, cardiovascular deaths, noncardiovascular deaths, and a composite endpoint of fatal coronary heart disease plus nonfatal myocardial infarction. Despite these null findings, the results of ALLHAT-LLT are not inconsistent with previous trials because of the very small lipid differences in the two arms. This indirectly supports the hypothesis that LDL cholesterol lowering is central to the cardiovascular benefits associated with statin therapy, with greater clinical impacts observed when there are greater differences between treatment and control arms. ALLHAT-LLT underscores the difficulty of conducting an open-label trial in an era of rapidly changing professional and public understanding of the possible benefits of lipid-lowering therapy and highlights the substantial gap between actual care in clinical practice and optimal care based on the best knowledge from randomized clinical trials. PMID- 15296705 TI - Low-density lipoprotein particle number and risk for cardiovascular disease. AB - The key role played by low-density lipoprotein (LDL) particles in the pathogenesis of coronary heart disease (CHD) is well accepted, as is the benefit of lowering LDL in high-risk patients. What remains controversial is whether we are using the best measure(s) of LDL to identify all individuals who would benefit from therapy. Many studies have shown that, at a given level of LDL cholesterol, individuals with predominantly small LDL particles (pattern B) experience greater CHD risk than those with larger-size LDL. However, it is not clear from this observation that small LDL particles are inherently more atherogenic than large ones because, at a given level of LDL cholesterol, individuals with small LDL have more LDL particles in total. The phenotype of small LDL particle size co-segregates with a cluster of metabolic factors, including elevated triglycerides and reduced HDL cholesterol, and in multivariate analyses has generally been found not to be independently associated with CHD risk. In contrast, LDL particle number measured by nuclear magnetic resonance has consistently been shown to be a strong, independent predictor of CHD. PMID- 15296706 TI - Evidence-based treatment of lipids in the elderly. AB - The elderly (men aged 65 years and older and women aged 75 years and older) constitute a population at high absolute risk for the morbidity and mortality of atherosclerotic cardiovascular disease. Statins have been shown in multiple large trials to reduce the burden of atherosclerotic disease in both middle-aged and elderly patients at elevated risk for coronary events, stroke, and death. We reviewed the major statin trials with particular emphasis on the significant number of elderly subjects. The impact of statins on the elderly, both positive and negative, is tabulated. In addition, we briefly discuss risk assessment in the elderly because selection of elderly patients for intensive low-density lipoprotein cholesterol reduction with statins requires clinical judgment that must weigh the need for subclinical measures of atherosclerosis. We also consider negative aspects, risks, and costs of such therapy. PMID- 15296707 TI - Inhibition of cholesteryl ester transfer protein activity: a new therapeutic approach to raising high-density lipoprotein. AB - High-density lipoprotein (HDL) cholesterol levels are inversely associated with risk of atherosclerotic cardiovascular disease (ASCVD), leading to the concept that pharmacologic therapy to raise HDL cholesterol levels may reduce ASCVD risk. There is substantial interest in the concept of inhibition of the cholesteryl ester transfer protein (CETP) as a novel strategy for raising HDL cholesterol levels, as well as reducing levels of atherogenic lipoproteins. This article reviews the physiology of CETP in lipoprotein metabolism and the data in animals and humans that are relevant to the question of whether CETP inhibition may some day be part of the clinical armamentarium for treating dyslipidemia and atherosclerotic vascular disease. PMID- 15296708 TI - The chromosomal passenger complex takes center stage during mitosis. AB - The chromosomal passenger complex plays important roles in key mitotic events, including chromosome bi-orientation, the spindle assembly checkpoint, and cytokinesis. Two groups now report the identification of a novel component of the Incenp/survivin/auroraB complex (Gassmann et al., 2004; Sampath et al., 2004) and show that different subcomplexes may exist during mitosis. Exciting data support the involvement of the passenger complex in yet another key event, the assembly of the mitotic spindle. PMID- 15296709 TI - Chk1 activation and the nuclear/cytoplasmic ratio. AB - ATR and Chk1 are important components of a cell cycle checkpoint pathway. In this issue of Developmental Cell, Conn et al. shed a novel light on the molecular mechanism of Chk1 activation and raise the possibility of a developmental checkpoint that regulates Chk1 in response to the nuclear/cytoplasmic ratio. PMID- 15296710 TI - Chewing the fat; regulating autophagy in Drosophila. AB - Autophagy is the major cellular process responsible for bulk cytoplasmic degradation. Two reports in this issue of Developmental Cell describe how both PI3 kinase and TOR signaling in Drosophila are critical for controlling autophagy in response to developmental and environmental cues. PMID- 15296711 TI - Signaling mucins in the (S)limelight. AB - Mucins may be the ugly ducklings of molecular biology. Their large size, repetitive nature, and unglamorous biological activities have not favored their study. However, integral membrane mucins have conserved intracellular C termini that may influence intracellular signaling. In a recent issue of Genes & Development, Cullen et al. show that the C terminus of membrane mucin-like Msb2 activates a CDC42/MAPK cascade to control filamentous growth of baker's yeast. PMID- 15296712 TI - SSD: sterol-sensing direct. AB - Cholesterol homeostasis is established by a complex of three proteins, one of which contains a hydrophobic domain previously termed a sterol-sensing domain. New biochemical studies of this domain demonstrate direct high-affinity binding of the sterol-sensing domain to sterol. PMID- 15296713 TI - Emerging asymmetry and embryonic patterning in early mouse development. AB - Recent studies have revealed asymmetries in the mouse zygote and preimplantation embryo, well before the establishment of anterior-posterior polarity after implantation. Whether these asymmetries are causally related to embryonic patterning or are coincidental outcomes of the topology of normal development remains uncertain. PMID- 15296714 TI - Role and regulation of starvation-induced autophagy in the Drosophila fat body. AB - In response to starvation, eukaryotic cells recover nutrients through autophagy, a lysosomal-mediated process of cytoplasmic degradation. Autophagy is known to be inhibited by TOR signaling, but the mechanisms of autophagy regulation and its role in TOR-mediated cell growth are unclear. Here, we show that signaling through TOR and its upstream regulators PI3K and Rheb is necessary and sufficient to suppress starvation-induced autophagy in the Drosophila fat body. In contrast, TOR's downstream effector S6K promotes rather than suppresses autophagy, suggesting S6K downregulation may limit autophagy during extended starvation. Despite the catabolic potential of autophagy, disruption of conserved components of the autophagic machinery, including ATG1 and ATG5, does not restore growth to TOR mutant cells. Instead, inhibition of autophagy enhances TOR mutant phenotypes, including reduced cell size, growth rate, and survival. Thus, in cells lacking TOR, autophagy plays a protective role that is dominant over its potential role as a growth suppressor. PMID- 15296715 TI - Programmed autophagy in the Drosophila fat body is induced by ecdysone through regulation of the PI3K pathway. AB - Eukaryotic cells catabolize their own cytoplasm by autophagy in response to amino acid starvation and inductive signals during programmed tissue remodeling and cell death. The Tor and PI3K signaling pathways have been shown to negatively control autophagy in eukaryotes, but the mechanisms that link these effectors to overall animal development and nutritional status in multicellular organisms remain poorly understood. Here, we reveal a complex regulation of programmed and starvation-induced autophagy in the Drosophila fat body. Gain-of-function genetic analysis indicated that ecdysone receptor signaling induces programmed autophagy whereas PI3K signaling represses programmed autophagy. Genetic interaction studies showed that ecdysone signaling downregulates PI3K signaling and that this represents the effector mechanism for induction of programmed autophagy. Hence, these studies link hormonal induction of autophagy to the regulatory function of the PI3K signaling pathway in vivo. PMID- 15296716 TI - Convergence of signaling pathways in the control of differential cell growth in Arabidopsis. AB - Seedling apical hook development involves a complex interplay of hormones and light in the regulation of differential cell growth. However, the underlying molecular mechanisms that integrate these diverse signals to control bending of the embryonic stem are poorly understood. The Arabidopsis ethylene-regulated HOOKLESS1 (HLS1) gene is essential for apical hook formation. Herein, we identify two auxin response regulators that act downstream of HLS1 to control cell elongation in the hypocotyl. Extragenic suppressors of hls1 were identified as mutations in AUXIN RESPONSE FACTOR 2 (ARF2). The level of ARF2 protein was decreased by ethylene, and this response required HLS1. Exposure to light decreased HLS1 protein levels and evoked a concomitant increase in ARF2 accumulation. These studies demonstrate that both ethylene and light signals affect differential cell growth by acting through HLS1 to modulate the auxin response factors, pinpointing HLS1 as a key integrator of the signaling pathways that control hypocotyl bending. PMID- 15296717 TI - Regulation of dendritic maintenance and growth by a mammalian 7-pass transmembrane cadherin. AB - Drosophila Flamingo is a 7-pass transmembrane cadherin that is necessary for dendritic patterning and axon guidance. How it works at the molecular level and whether homologs of Flamingo play similar roles in mammalian neurons or not have been unanswered questions. Here, we performed loss-of-function analysis using an RNAi system and organotypic brain slice cultures to address the role of a mammalian Flamingo homolog, Celsr2. Knocking down Celsr2 resulted in prominent simplification of dendritic arbors of cortical pyramidal neurons and Purkinje neurons, and this phenotype seemed to be due to branch retraction. Cadherin domain-mediated homophilic interaction appears to be required for the maintenance of dendritic branches. Furthermore, expression of various Celsr2 forms elicited distinct responses that were dependent on an extracellular subregion outside the cadherin domains and on a portion within the carboxyl intracellular tail. Based on these findings, we discuss how Celsr2 may regulate dendritic maintenance and growth. PMID- 15296718 TI - EspFU is a translocated EHEC effector that interacts with Tir and N-WASP and promotes Nck-independent actin assembly. AB - Several microbial pathogens including enteropathogenic E. coli (EPEC) exploit mammalian tyrosine-kinase signaling cascades to recruit Nck adaptor proteins and activate N-WASP-Arp2/3-mediated actin assembly. To promote localized actin "pedestal formation," EPEC translocates the bacterial effector protein Tir into the plasma membrane, where it is tyrosine-phosphorylated and binds Nck. Enterohemorrhagic E. coli (EHEC) also generates Tir-dependent pedestals, but in the absence of phosphotyrosines and Nck recruitment. To identify additional EHEC effectors that stimulate phosphotyrosine-independent actin assembly, we systematically generated EHEC mutants containing specific deletions in putative pathogenicity-islands. Among 0.33 Mb of deleted sequences, only one ORF was critical for pedestal formation. It lies within prophage-U, and encodes a protein similar to the known effector EspF. This proline-rich protein, EspFU, is the only EHEC effector of actin assembly absent from EPEC. Whereas EHEC Tir cannot efficiently recruit N-WASP or trigger actin polymerization, EspFU associates with Tir, binds N-WASP, and potently stimulates Nck-independent actin assembly. PMID- 15296719 TI - A simple molecular complex mediates widespread BMP-induced repression during Drosophila development. AB - The spatial and temporal control of gene expression during the development of multicellular organisms is regulated to a large degree by cell-cell signaling. We have uncovered a simple mechanism through which Dpp, a TGFbeta/BMP superfamily member in Drosophila, represses many key developmental genes in different tissues. A short DNA sequence, a Dpp-dependent silencer element, is sufficient to confer repression of gene transcription upon Dpp receptor activation and nuclear translocation of Mad and Medea. Transcriptional repression does not require the cooperative action of cell type-specific transcription factors but relies solely on the capacity of the silencer element to interact with Mad and Medea and to subsequently recruit the zinc finger-containing repressor protein Schnurri. Our findings demonstrate how the Dpp pathway can repress key targets in a simple and tissue-unrestricted manner in vivo and hence provide a paradigm for the inherent capacity of a signaling system to repress transcription upon pathway activation. PMID- 15296720 TI - Recruitment of ectodermal attachment cells via an EGFR-dependent mechanism during the organogenesis of Drosophila proprioceptors. AB - Drosophila proprioceptors (chordotonal organs) are structured as a linear array of four lineage-related cells: a neuron, a glial cell, and two accessory cells, called cap and ligament, between which the neuron is stretched. To function properly as stretch receptors, chordotonal organs must be stably anchored at both edges. The cap cells are anchored to the cuticle through specialized lineage related attachment cells. However, the mechanism by which the ligament cells at the other edge of the organ attach is not known. Here, we report the identification of specialized attachment cells that anchor the ligament cells of pentascolopidial chordotonal organs (lch5) to the cuticle. The ligament attachment cells are recruited by the approaching ligament cells upon reaching their attachment site, through an EGFR-dependent mechanism. Molecular characterization of lch5 attachment cells demonstrated that they share significant properties with Drosophila tendon cells and with mammalian proprioceptive organs. PMID- 15296721 TI - A crucial interaction between embryonic red blood cell progenitors and paraxial mesoderm revealed in spadetail embryos. AB - Zebrafish embryonic red blood cells (RBCs) develop in trunk intermediate mesoderm (IM), and early macrophages develop in the head, suggesting that local microenvironmental cues regulate differentiation of these two blood lineages. spadetail (spt) mutant embryos, which lack trunk paraxial mesoderm (PM) due to a cell-autonomous defect in tbx16, fail to produce embryonic RBCs but retain head macrophage development. In spt mutants, initial hematopoietic gene expression is absent in trunk IM, although endothelial and pronephric expression is retained, suggesting that early blood progenitor development is specifically disrupted. Using cell transplantation, we reveal that spt is required cell autonomously for early hematopoietic gene expression in trunk IM. Further, we uncover an interaction between embryonic trunk PM and blood progenitors that is essential for RBC development. Importantly, our data identify a hematopoietic microenvironment that allows embryonic RBC production in the zebrafish. PMID- 15296722 TI - Cdc28/Cdk1 regulates spindle pole body duplication through phosphorylation of Spc42 and Mps1. AB - Duplication of the Saccharomyces cerevisiae spindle pole body (SPB) once per cell cycle is essential for bipolar spindle formation and accurate chromosome segregation during mitosis. We have investigated the role that the major yeast cyclin-dependent kinase Cdc28/Cdk1 plays in assembly of a core SPB component, Spc42, to better understand how SPB duplication is coordinated with cell cycle progression. Cdc28 is required for SPB duplication and Spc42 assembly, and we found that Cdc28 directly phosphorylates Spc42 to promote its assembly into the SPB. The Mps1 kinase, previously shown to regulate Spc42 phosphorylation and assembly, is also a Cdc28 substrate, and Cdc28 phosphorylation of Mps1 is needed to maintain wild-type levels of Mps1 in cells. Analysis of nonphosphorylatable mutants in SPC42 and MPS1 indicates that direct Spc42 phosphorylation and indirect regulation of Spc42 through Mps1 are two overlapping pathways by which Cdc28 regulates Spc42 assembly and SPB duplication during the cell cycle. PMID- 15296723 TI - The DNA damage checkpoint in embryonic cell cycles is dependent on the DNA-to cytoplasmic ratio. AB - In Xenopus, cell cycle checkpoints monitoring DNA damage, DNA replication, and spindle assembly do not appear until after the midblastula transition (MBT; 4000 cells). We show that a DNA damage checkpoint can slow the cell cycle even in 2 cell embryos when the DNA content is increased. Slowing follows caffeine sensitive activation of the checkpoint kinase, Chk1; degradation of the cell cycle phosphatase, Cdc25A; and inhibitory phosphorylation of Cdc25C and cyclin dependent kinases (Cdks). Alterations in the DNA-to-cytoplasmic ratio elicit a dose-dependent DNA damage checkpoint, and the ratio required to activate Chk1 for the damage response is lower than that associated with "developmental" activation of Chk1 shortly after the MBT. Our results indicate that a maternal damage response, independent of zygotic transcription, is present even in very early embryos, and requires both double-stranded DNA ends and a threshold DNA-to cytoplasmic ratio to significantly affect the cell cycle. PMID- 15296724 TI - NAD+ specificity of bacterial DNA ligase revealed. PMID- 15296725 TI - Complete three-dimensional structures of picornaviral RNA-dependent RNA polymerases. PMID- 15296726 TI - The evolving role of 3D domain swapping in proteins. PMID- 15296727 TI - Computational studies of membrane channels. AB - The determination of the structure of several members of the K+ channel and aquaporin family represents a unique opportunity to explain the mechanism of these biomolecular systems. With their ability to go beyond static structures, molecular dynamics simulations offer a unique route for relating functional properties to membrane channel structure. The recent progress in this area is reviewed. PMID- 15296729 TI - Optimization of protein production in mammalian cells with a coexpressed fluorescent marker. AB - The expression of mammalian proteins in sufficient abundance and quality for structural studies often presents formidable challenges. Many express poorly in bacterial systems, whereas it can be time consuming and expensive to produce them from cells of higher organisms. Here we describe a procedure for the direct selection of stable mammalian cell lines that express proteins of interest in high yield. Coexpression of a marker protein, such as green fluorescent protein, is linked to that of the desired protein through an internal ribosome entry site in the vector that is transfected into cells in culture. The coexpressed marker is used to select for highly expressing clonal cell lines. Applications are described to a membrane protein, the 5HT2c serotonin receptor, and to a secreted cysteine-rich protein, resistin. Besides providing an expeditious means for producing mammalian proteins for structural work, the resulting cell lines also readily support tests of functional properties and structure-inspired hypotheses. PMID- 15296730 TI - Crystal structures of a ligand-free and malonate-bound human caspase-1: implications for the mechanism of substrate binding. AB - Caspase-1, a mediator of the posttranslational processing of IL-1beta and IL-18, requires an aspartic acid in the P1 position of its substrates. The mechanisms of caspase-1 activation remain poorly understood despite numerous structures of the enzyme complexed with aspartate-based inhibitors. Here we report a crystal structure of ligand-free caspase-1 that displays dramatic rearrangements of loops defining the active site to generate a closed conformation that is incompatible with substrate binding. A structure of the enzyme complexed with malonate shows the protein in its open (active-site ligand-bound) conformation in which malonate reproduces the hydrogen bonding network observed in structures with covalent inhibitors. These results illustrate the essential function of the obligatory aspartate recognition element that opens the active site of caspase-1 to substrates and may be the determinant responsible for the conformational changes between ligand-free and -bound forms of the enzyme, and suggest a new approach for identifying novel aspartic acid mimetics. PMID- 15296731 TI - Crystal structure of UAP56, a DExD/H-box protein involved in pre-mRNA splicing and mRNA export. AB - UAP56 is an essential eukaryotic pre-mRNA splicing factor and mRNA export factor. The mechanisms of its functions are not well understood. We determined the crystal structures of the N- and C-terminal domains of human UAP56 (comprising 90% of the full-length UAP56) at 1.9 A resolution. The two domains each have a RecA-like fold and are connected by a flexible linker. The overall fold of each domain is highly similar to the corresponding domains of eIF4A (a prototypic DExD/H-box protein), with differences at the loops and termini. This structural similarity suggests that UAP56 is likely to possess ATPase and helicase activity similar to eIF4A. The NTP binding pocket of UAP56 is occupied by a citrate ion, mimicking the phosphates of NTP and retaining the P loop in an open conformation. The crystal structure of the N-terminal domain of UAP56 also reveals a dimer interface that is potentially important for UAP56's function. PMID- 15296732 TI - Structure of Escherichia coli AMP nucleosidase reveals similarity to nucleoside phosphorylases. AB - AMP nucleosidase (AMN) catalyzes the hydrolysis of AMP to form adenine and ribose 5-phosphate. The enzyme is found only in prokaryotes, where it plays a role in purine nucleoside salvage and intracellular AMP level regulation. Enzyme activity is stimulated by ATP and suppressed by phosphate. The structure of unliganded AMN was determined at 2.7 A resolution, and structures of the complexes with either formycin 5'-monophosphate or inorganic phosphate were determined at 2.6 A and 3.0 A resolution, respectively. AMN is a biological homohexamer, and each monomer is composed of two domains: a catalytic domain and a putative regulatory domain. The overall topology of the catalytic domain and some features of the substrate binding site resemble those of the nucleoside phosphorylases, demonstrating that AMN is a new member of the family. The structure of the regulatory domain consists of a long helix and a four-stranded sheet and has a novel topology. PMID- 15296733 TI - Crystal structure of the catalytic core of human DNA polymerase kappa. AB - We present the crystal structure of the catalytic core of human DNA polymerase kappa (hPolkappa), the first structure of a human Y-family polymerase. hPolkappa is implicated in the proficient extension of mispaired primer termini on undamaged DNAs, and in the extension step of lesion bypass. The structure reveals a stubby "fingers" subdomain, which despite its small size appears to be tightly restrained with respect to a putative templating base. The structure also reveals a novel "thumb" subdomain that provides a basis for the importance of the N terminal extension unique to hPolkappa. And, most surprisingly, the structure reveals the polymerase-associated domain (PAD) juxtaposed on the dorsal side of the "palm" subdomain, as opposed to the fingers subdomain. Together, these properties suggest that the hPolkappa active site is constrained at the site of the templating base and incoming nucleotide, but the polymerase is less constrained following translocation of the lesion. PMID- 15296734 TI - Finding functional sites in structural genomics proteins. AB - Assigning function to structures is an important aspect of structural genomics projects, since they frequently provide structures for uncharacterized proteins. Similarities uncovered by structure alignment can suggest a similar function, even in the absence of sequence similarity. For proteins adopting novel folds or those with many functions, this strategy can fail, but functional clues can still come from comparison of local functional sites involving a few key residues. Here we assess the general applicability of functional site comparison through the study of 157 proteins solved by structural genomics initiatives. For 17, the method bolsters confidence in predictions made based on overall fold similarity. For another 12 with new folds, it suggests functions, including a putative phosphotyrosine binding site in the Archaeal protein Mth1187 and an active site for a ribose isomerase. The approach is applied weekly to all new structures, providing a resource for those interested in using structure to infer function. PMID- 15296735 TI - Crystal structures of CTP synthetase reveal ATP, UTP, and glutamine binding sites. AB - CTP synthetase (CTPs) catalyzes the last step in CTP biosynthesis, in which ammonia generated at the glutaminase domain reacts with the ATP-phosphorylated UTP at the synthetase domain to give CTP. Glutamine hydrolysis is active in the presence of ATP and UTP and is stimulated by the addition of GTP. We report the crystal structures of Thermus thermophilus HB8 CTPs alone, CTPs with 3SO4(2-), and CTPs with glutamine. The enzyme is folded into a homotetramer with a cross shaped structure. Based on the binding mode of sulfate anions to the synthetase site, ATP and UTP are computer modeled into CTPs with a geometry favorable for the reaction. Glutamine bound to the glutaminase domain is situated next to the triad of Glu-His-Cys as a catalyst and a water molecule. Structural information provides an insight into the conformational changes associated with the binding of ATP and UTP and the formation of the GTP binding site. PMID- 15296736 TI - Active site geometry and substrate recognition of the molybdenum hydroxylase quinoline 2-oxidoreductase. AB - The soil bacterium Pseudomonas putida 86 uses quinoline as a sole source of carbon and energy. Quinoline 2-oxidoreductase (Qor) catalyzes the first metabolic step converting quinoline to 2-oxo-1,2-dihydroquinoline. Qor is a member of the molybdenum hydroxylases. The molybdenum ion is coordinated by two ene-dithiolate sulfur atoms, two oxo-ligands, and a catalytically crucial sulfido-ligand, whose position in the active site was controversial. The 1.8 A resolution crystal structure of Qor indicates that the sulfido-ligand occupies the equatorial position at the molybdenum ion. The structural comparison of Qor with the allopurinol-inhibited xanthine dehydrogenase from Rhodobacter capsulatus allows direct insight into the mechanism of substrate recognition and the identification of putative catalytic residues. The active site protein variants QorE743V and QorE743D were analyzed to assess the catalytic role of E743. PMID- 15296737 TI - The combinatorial extension method reveals a sphingolipid binding domain on pancreatic bile salt-dependent lipase: role in secretion. AB - Structure similarity searches using a combinatorial extension approach revealed that a protein fold structurally related to the sphingolipid binding domain (SBD) of HIV-1 gp120 (V3 loop) is present on pancreatic bile salt-dependent lipase (BSDL). A synthetic peptide derived from the predicted V3-like domain of BSDL interacted with reconstituted monolayers of sphingolipids such as GalCer and GlcCer. Using Chinese hamster ovary cells stably transfected with the cDNA encoding the rat BSDL (CHO-3B clone) or pancreatic SOJ-6 cells expressing the human BSDL as models, we showed that the enzyme cofractionates with caveolin-1. The secretion of BSDL by CHO-3B cells was inhibited by permeable drugs affecting rafts structure (D609, PDMP, and filipin). Data suggest that the functional interaction between the BSDL SBD and lipid rafts is physiologically relevant and could be essential for sensing the BSDL folding prior to secretion. A tentative model accounting for the phosphorylation-induced dissociation of BSDL from rafts is presented. PMID- 15296738 TI - Structural rearrangement accompanying NAD+ synthesis within a bacterial DNA ligase crystal. AB - DNA ligase is an enzyme important for DNA repair and replication. Eukaryotic genomes encode ligases requiring ATP as the cofactor; bacterial genomes encode NAD(+)-dependent ligase. This difference in substrate specificities and the essentiality of NAD(+)-dependent ligase for bacterial survival make NAD(+) dependent ligase a good target for designing highly specific anti-infectives. Any such structure-guided effort would require the knowledge of the precise mechanism of NAD+ recognition by the enzyme. We report the principles of NAD+ recognition by presenting the synthesis of NAD+ from nicotinamide mononucleotide (NMN) and AMP, catalyzed by Enterococcus faecalis ligase within the crystal lattice. Unprecedented conformational change, required to reorient the two subdomains of the protein for the condensation to occur and to recognize NAD+, is captured in two structures obtained using the same protein crystal. Structural data and sequence analysis presented here confirms and extends prior functional studies of the ligase adenylation reaction. PMID- 15296739 TI - Systematic analysis of added-value in simple comparative models of protein structure. AB - Added-value is the additional information that a model carries with respect to the template structure used for model building. Thousands of single-template models, corresponding to proteins of known structure, were analyzed. The accuracy of structure-derived properties, such as residue accessibility, surface area, electrostatic potential, and others, was determined as a function of template:target sequence identity by comparing the models with their corresponding experimental structures. Added-value was determined by comparing the accuracy in models with that from templates. Geometry-dependent properties such as neighborhood of buried residues and accessible surface area showed low added-value. Properties that also depend on the protein sequence, such as presence of polar areas and electrostatic potential, showed high added-value. In general added-value increases when template:target sequence identity decreases, but it is also affected by alignment errors. This study justifies the use of models instead of the use of templates to estimate structure-derived properties of a target protein. PMID- 15296740 TI - Crystal structure of the native chaperonin complex from Thermus thermophilus revealed unexpected asymmetry at the cis-cavity. AB - The chaperonins GroEL and GroES are essential mediators of protein folding. GroEL binds nonnative protein, ATP, and GroES, generating a ternary complex in which protein folding occurs within the cavity capped by GroES (cis-cavity). We determined the crystal structure of the native GroEL-GroES-ADP homolog from Thermus thermophilus, with substrate proteins in the cis-cavity, at 2.8 A resolution. Twenty-four in vivo substrate proteins within the cis-cavity were identified from the crystals. The structure around the cis-cavity, which encapsulates substrate proteins, shows significant differences from that observed for the substrate-free Escherichia coli GroEL-GroES complex. The apical domain around the cis-cavity of the Thermus GroEL-GroES complex exhibits a large deviation from the 7-fold symmetry. As a result, the GroEL-GroES interface differs considerably from the previously reported E. coli GroEL-GroES complex, including a previously unknown contact between GroEL and GroES. PMID- 15296741 TI - Crystal structure of an acylpeptide hydrolase/esterase from Aeropyrum pernix K1. AB - Acylpeptide hydrolases (APH; also known as acylamino acid releasing enzyme) catalyze the removal of an N-acylated amino acid from blocked peptides. The crystal structure of an APH from the thermophilic archaeon Aeropyrum pernix K1 to 2.1 A resolution confirms it to be a member of the prolyl oligopeptidase family of serine proteases. The structure of apAPH is a symmetric homodimer with each subunit comprised of two domains. The N-terminal domain is a regular seven-bladed beta-propeller, while the C-terminal domain has a canonical alpha/beta hydrolase fold and includes the active site and a conserved Ser445-Asp524-His556 catalytic triad. The complex structure of apAPH with an organophosphorus substrate, p nitrophenyl phosphate, has also been determined. The complex structure unambiguously maps out the substrate binding pocket and provides a basis for substrate recognition by apAPH. A conserved mechanism for protein degradation from archaea to mammals is suggested by the structural features of apAPH. PMID- 15296742 TI - Snapshot of protein structure evolution reveals conservation of functional dimerization through intertwined folding. AB - Protein-protein interactions govern a wide range of cellular processes. Molecular recognition responsible for homodimerization and heterodimerization in the rel/NF kappaB family of eukaryotic transcription factors relies on a small cluster of hydrophobic residues. We have carried out a structural analysis of six NF-kappaB p50 dimer interface mutants; one of them revealed a remarkable alteration. One or possibly both its mutations cause a switch into an intertwined dimer, in which the molecular partners exchange nearly half of their fold. In spite of the extensive swapping of secondary structure elements, the topology within each counterpart is preserved, with a very similar overall structure and minimal changes at the interface. Thus intertwining rescues structure and function from a destabilizing mutation. Since the mutants originate from a directed evolution experiment and are functional, the data provide an evolutionary snapshot of how a protein structure can respond to mutations while maintaining a functional molecular architecture. PMID- 15296743 TI - Structural basis for interactions between tenascins and lectican C-type lectin domains: evidence for a crosslinking role for tenascins. AB - The C-terminal G3 domains of lecticans mediate crosslinking to diverse extracellular matrix (ECM) proteins during ECM assembly, through their C-type lectin (CLD) subdomains. The structure of the rat aggrecan CLD in a Ca(2+) dependent complex with fibronectin type III repeats 3-5 of rat tenascin-R provides detailed support for such crosslinking. The CLD loops bind Ca2+ like other CLDs, but no carbohydrate binding is observed or possible. This is thus the first example of a direct Ca(2+)-dependent protein-protein interaction of a CLD. Surprisingly, tenascin-R does not coordinate the Ca2+ ions directly. Electron microscopy confirms that full-length tenascin-R and tenascin-C crosslink hyaluronan-aggrecan complexes. The results are significant for the binding of all lectican CLDs to tenascin-R and tenascin-C. Comparison of the protein interaction surface with that of P-selectin in complex with the PGSL-1 peptide suggests that direct protein-protein interactions of Ca(2+)-binding CLDs may be more widespread than previously appreciated. PMID- 15296744 TI - 10 residue folded peptide designed by segment statistics. AB - We have designed a peptide termed chignolin, consisting of only 10 amino acid residues (GYDPETGTWG), on the basis of statistics derived from more than 10,000 protein segments. The peptide folds into a unique structure in water and shows a cooperative thermal transition, both of which may be hallmarks of a protein. Also, the experimentally determined beta-hairpin structure was very close to what we had targeted. The performance of the short peptide not only implies that the methodology employed here can contribute toward development of novel techniques for protein design, but it also yields insights into the raison d'etre of an autonomous element involved in a natural protein. This is of interest for the pursuit of folding mechanisms and evolutionary processes of proteins. PMID- 15296745 TI - A basis for SUMO protease specificity provided by analysis of human Senp2 and a Senp2-SUMO complex. AB - Modification of cellular proteins by the ubiquitin-like protein SUMO is essential for nuclear metabolism and cell cycle progression in yeast. X-ray structures of the human Senp2 catalytic protease domain and of a covalent thiohemiacetal transition-state complex obtained between the Senp2 catalytic domain and SUMO-1 revealed details of the respective protease and substrate surfaces utilized in interactions between these two proteins. Comparative biochemical and structural analysis between Senp2 and the yeast SUMO protease Ulp1 revealed differential abilities to process SUMO-1, SUMO-2, and SUMO-3 in maturation and deconjugation reactions. Further biochemical characterization of the three SUMO isoforms into which an additional Gly-Gly di-peptide was inserted, or whereby the respective SUMO tails from the three isoforms were swapped, suggests a strict dependence for SUMO isopeptidase activity on residues C-terminal to the conserved Gly-Gly motif and preferred cleavage site for SUMO proteases. PMID- 15296746 TI - The crystal structure of the RNA-dependent RNA polymerase from human rhinovirus: a dual function target for common cold antiviral therapy. AB - Human rhinoviruses (HRV), the predominant members of the Picornaviridae family of positive-strand RNA viruses, are the major causative agents of the common cold. Given the lack of effective treatments for rhinoviral infections, virally encoded proteins have become attractive therapeutic targets. The HRV genome encodes an RNA-dependent RNA polymerase (RdRp) denoted 3Dpol, which is responsible for replicating the viral genome and for synthesizing a protein primer used in the replication. Here the crystal structures for three viral serotypes (1B, 14, and 16) of HRV 3Dpol have been determined. The three structures are very similar to one another, and to the closely related poliovirus (PV) 3Dpol enzyme. Because the reported PV crystal structure shows significant disorder, HRV 3Dpol provides the first complete view of a picornaviral RdRp. The folding topology of HRV 3Dpol also resembles that of RdRps from hepatitis C virus (HCV) and rabbit hemorrhagic disease virus (RHDV) despite very low sequence homology. PMID- 15296747 TI - Nocturnal vision and landmark orientation in a tropical halictid bee. AB - BACKGROUND: Some bees and wasps have evolved nocturnal behavior, presumably to exploit night-flowering plants or avoid predators. Like their day-active relatives, they have apposition compound eyes, a design usually found in diurnal insects. The insensitive optics of apposition eyes are not well suited for nocturnal vision. How well then do nocturnal bees and wasps see? What optical and neural adaptations have they evolved for nocturnal vision? RESULTS: We studied female tropical nocturnal sweat bees (Megalopta genalis) and discovered that they are able to learn landmarks around their nest entrance prior to nocturnal foraging trips and to use them to locate the nest upon return. The morphology and optics of the eye, and the physiological properties of the photoreceptors, have evolved to give Megalopta's eyes almost 30 times greater sensitivity to light than the eyes of diurnal worker honeybees, but this alone does not explain their nocturnal visual behavior. This implies that sensitivity is improved by a strategy of photon summation in time and in space, the latter of which requires the presence of specialized cells that laterally connect ommatidia into groups. First-order interneurons, with significantly wider lateral branching than those found in diurnal bees, have been identified in the first optic ganglion (the lamina ganglionaris) of Megalopta's optic lobe. We believe that these cells have the potential to mediate spatial summation. CONCLUSIONS: Despite the scarcity of photons, Megalopta is able to visually orient to landmarks at night in a dark forest understory, an ability permitted by unusually sensitive apposition eyes and neural photon summation. PMID- 15296748 TI - Roundabout 2 regulates migration of sensory neurons by signaling in trans. AB - BACKGROUND: Roundabout (Robo) receptors and their ligand Slit are important regulators of axon guidance and cell migration. The development of Drosophila embryonic sense organs provides a neuronal migration paradigm where the in vivo roles of Slit and Robo can be assayed using genetics. RESULTS: Here we show that Slit-Robo signaling controls migration of Drosophila larval sensory neurons that are part of the Chordotonal (Cho) stretch receptor organs. We used live imaging to show that abdominal Cho organs normally migrate ventrally during development, whereas thoracic Cho organs do not. Robo2 overexpression in cis (in the sensory neurons) or in trans (on neighboring visceral mesoderm) transforms abdominal organs to a thoracic morphology and position by blocking migration, while loss of Slit-Robo signaling produces a reverse transformation in which thoracic organs migrate ectopically. Rescue and tissue-specific knockout experiments indicate that trans signaling by Robo2 contributes to the normal positioning of the thoracic Cho organs. The differential positioning of Cho organs between the thorax and abdomen is known to be regulated by Hox genes, and we show that the essential Hox cofactor Homothorax, represses Robo2 expression in the abdominal visceral mesoderm. CONCLUSIONS: Our results suggest that segment-specific neuronal migration patterns are directed through a novel signaling complex (the "Slit sandwich") in which Robo2 on the thoracic visceral mesoderm binds to Slit and presents it to Robo receptors on Cho neurons. The differential positioning of Cho organs between thorax and abdomen may be determined by Hox gene-mediated repression of robo2. PMID- 15296749 TI - Laser microsurgery in fission yeast; role of the mitotic spindle midzone in anaphase B. AB - INTRODUCTION: During anaphase B in mitosis, polymerization and sliding of overlapping spindle microtubules (MTs) contribute to the outward movement the spindle pole bodies (SPBs). To probe the mechanism of spindle elongation, we combine fluorescence microscopy, photobleaching, and laser microsurgery in the fission yeast Schizosaccharomyces pombe. RESULTS: We demonstrate that a green laser cuts intracellular structures in yeast cells with high spatial specificity. By using laser microsurgery, we cut mitotic spindles labeled with GFP-tubulin at various stages of anaphase B. Although cutting generally caused early anaphase spindles to disassemble, midanaphase spindle fragments continued to elongate. In particular, when the spindle was cut near a SPB, the larger spindle fragment continued to elongate in the direction of the cut. Photobleach marks showed that sliding of overlapping midzone MTs was responsible for the elongation of the spindle fragment. Spindle midzone fragments not connected to either of the two spindle poles also elongated. Equatorial microtubule organizing center (eMTOC) activity was not affected in cells with one detached pole but was delayed or absent in cells with two detached poles. CONCLUSIONS: These studies reveal that the spindle midzone is necessary and sufficient for the stabilization of MT ends and for spindle elongation. By contrast, SPBs are not required for elongation, but they contribute to the attachment of the nuclear envelope and chromosomes to the spindle, and to cell cycle progression. Laser microsurgery provides a means by which to dissect the mechanics of the spindle in yeast. PMID- 15296750 TI - ATM is required for telomere maintenance and chromosome stability during Drosophila development. AB - ATM is a large, multifunctional protein kinase that regulates responses required for surviving DNA damage: including DNA repair, apoptosis, and cell cycle checkpoints. Here, we show that Drosophila ATM function is essential for normal adult development. Extensive, inappropriate apoptosis occurs in proliferating atm mutant tissues, and in clonally derived atm mutant embryos, frequent mitotic defects were seen. At a cellular level, spontaneous telomere fusions and other chromosomal abnormalities are common in atm larval neuroblasts, suggesting a conserved and essential role for dATM in the maintenance of normal telomeres and chromosome stability. Evidence from other systems supports the idea that DNA double-strand break (DSB) repair functions of ATM kinases promote telomere maintenance by inhibition of illegitimate recombination or fusion events between the legitimate ends of chromosomes and spontaneous DSBs. Drosophila will be an excellent model system for investigating how these ATM-dependent chromosome structural maintenance functions are deployed during development. Because neurons appear to be particularly sensitive to loss of ATM in both flies and humans, this system should be particularly useful for identifying cell-specific factors that influence sensitivity to loss of dATM and are relevant for understanding the human disease, ataxia-telangiectasia. PMID- 15296751 TI - Telomere protection without a telomerase; the role of ATM and Mre11 in Drosophila telomere maintenance. AB - The conserved ATM checkpoint kinase and the Mre11 DNA repair complex play essential and overlapping roles in maintaining genomic integrity. We conducted genetic and cytological studies on Drosophila atm and mre11 knockout mutants and discovered a telomere defect that was more severe than in any of the non Drosophila systems studied. In mutant mitotic cells, an average of 30% of the chromosome ends engaged in telomere fusions. These fusions led to the formation and sometimes breakage of dicentric chromosomes, thus starting a devastating breakage-fusion-bridge cycle. Some of the fusions depended on DNA ligase IV, which suggested that they occurred by a nonhomologous end-joining (NHEJ) mechanism. Epistasis analyses results suggest that ATM and Mre11 might also act in the same telomere maintenance pathway in metazoans. Since Drosophila telomeres are not added by a telomerase, our findings support an additional role for both ATM and Mre11 in telomere maintenance that is independent of telomerase regulation. PMID- 15296752 TI - The Drosophila ATM ortholog, dATM, mediates the response to ionizing radiation and to spontaneous DNA damage during development. AB - Cells of metazoan organisms respond to DNA damage by arresting their cell cycle to repair DNA, or they undergo apoptosis. Two protein kinases, ataxia telangiectasia mutated (ATM) and ATM and Rad-3 related (ATR), are sensors for DNA damage. In humans, ATM is mutated in patients with ataxia-telangiectasia (A-T), resulting in hypersensitivity to ionizing radiation (IR) and increased cancer susceptibility. Cells from A-T patients exhibit chromosome aberrations and excessive spontaneous apoptosis. We used Drosophila as a model system to study ATM function. Previous studies suggest that mei-41 corresponds to ATM in Drosophila; however, it appears that mei-41 is probably the ATR ortholog. Unlike mei-41 mutants, flies deficient for the true ATM ortholog, dATM, die as pupae or eclose with eye and wing abnormalities. Developing larval discs exhibit substantially increased spontaneous chromosomal telomere fusions and p53 dependent apoptosis. These developmental phenotypes are unique to dATM, and both dATM and mei-41 have temporally distinct roles in G2 arrest after IR. Thus, ATM and ATR orthologs are required for different functions in Drosophila; the developmental defects resulting from absence of dATM suggest an important role in mediating a protective checkpoint against DNA damage arising during normal cell proliferation and differentiation. PMID- 15296753 TI - The Drosophila Mre11/Rad50 complex is required to prevent both telomeric fusion and chromosome breakage. AB - The MRN complex consists of the two evolutionarily conserved components Mre11 and Rad50 and the third less-conserved component Nbs1/Xrs2. This complex mediates telomere maintenance in addition to a variety of functions in response to DNA double-strand breaks, including homologous recombination, nonhomologous end joining (NHEJ), and activation of DNA damage checkpoints. Mutations in the Mre11 gene cause the human ataxia-telangiectasia-like disorder (ATDL). Here, we show that null mutations in the Drosophila mre11 and rad50 genes cause both telomeric fusion and chromosome breakage. Moreover, we demonstrate that these mutations are in the same epistasis group required for telomere capping and mitotic chromosome integrity. Using an antibody against Rad50, we show that this protein is uniformly distributed along mitotic chromosomes, and that Rad50 is unstable in the absence of its binding partner Mre11. To define the roles of rad50 and mre11 in telomere protection, mutant chromosome preparations were immunostained for both HP1 and HOAP, two proteins that protect Drosophila telomeres from fusion. Cytological analysis revealed that mutations in rad50 and mre11 drastically reduce accumulation of HOAP and HP1 at telomeres. This suggests that the MRN complex protects Drosophila telomeres by facilitating recruitment of HOAP and HP1 at chromosome ends. PMID- 15296754 TI - Circadian clock mutation disrupts estrous cyclicity and maintenance of pregnancy. AB - Classic experiments have shown that ovulation and estrous cyclicity are under circadian control and that surgical ablation of the suprachiasmatic nuclei (SCN) results in estrous acyclicity in rats. Here, we characterized reproductive function in the circadian Clock mutant mouse and found that the circadian Clock mutation both disrupts estrous cyclicity and interferes with the maintenance of pregnancy. Clock mutant females have extended, irregular estrous cycles, lack a coordinated luteinizing hormone (LH) surge on the day of proestrus, exhibit increased fetal reabsorption during pregnancy, and have a high rate of full-term pregnancy failure. Clock mutants also show an unexpected decline in progesterone levels at midpregnancy and a shortened duration of pseudopregnancy, suggesting that maternal prolactin release may be abnormal. In a second set of experiments, we interrogated the function of each level of the hypothalamic-pituitary-gonadal (HPG) axis in order to determine how the Clock mutation disrupts estrous cyclicity. We report that Clock mutants fail to show an LH surge following estradiol priming in spite of the fact that hypothalamic levels of gonadotropin releasing hormone (GnRH), pituitary release of LH, and serum levels of estradiol and progesterone are all normal in Clock/Clock females. These data suggest that Clock mutants lack an appropriate circadian daily-timing signal required to coordinate hypothalamic hormone secretion. Defining the mechanisms by which the Clock mutation disrupts reproductive function offers a model for understanding how circadian genes affect complex physiological systems. PMID- 15296755 TI - Latrotoxin receptor signaling engages the UNC-13-dependent vesicle-priming pathway in C. elegans. AB - alpha-latrotoxin (LTX), a 120 kDa protein in black widow spider venom, triggers massive neurotransmitter exocytosis. Previous studies have highlighted a role for both intrinsic pore-forming activity and receptor binding in the action of this toxin. Intriguingly, activation of a presynaptic G protein-coupled receptor, latrophilin, may trigger release independent of pore-formation. Here we have utilized a previously identified ligand of nematode latrophilin, emodepside, to define a latrophilin-dependent pathway for neurotransmitter release in C. elegans. In the pharyngeal nervous system of this animal, emodepside (100 nM) stimulates exocytosis and elicits pharyngeal paralysis. The pharynxes of animals with latrophilin (lat-1) gene knockouts are resistant to emodepside, indicating that emodepside exerts its high-affinity paralytic effect through LAT-1. The expression pattern of lat-1 supports the hypothesis that emodepside exerts its effect on the pharynx primarily via neuronal latrophilin. We build on these observations to show that pharynxes from animals with either reduction or loss of function mutations in Gq, phospholipaseC-beta, and UNC-13 are resistant to emodepside. The latter is a key priming molecule essential for synaptic vesicle mediated release of neurotransmitter. We conclude that the small molecule ligand emodepside triggers latrophilin-mediated exocytosis via a pathway that engages UNC-13-dependent vesicle priming. PMID- 15296756 TI - ARAP3 is a PI3K- and rap-regulated GAP for RhoA. AB - Rho and Arf family small GTPases are well-known regulators of cellular actin dynamics. We recently identified ARAP3, a member of the ARAP family of dual GTPase activating proteins (GAPs) for Arf and Rho family GTPases, in a screen for PtdIns(3,4,5)P(3) binding proteins. PtdIns(3,4,5)P(3) is the lipid product of class I phosphoinositide 3OH-kinases (PI3Ks) and is a signaling molecule used by growth factor receptors and integrins in the regulation of cell dynamics. We report here that as a Rho GAP, ARAP3 prefers RhoA as a substrate and that it can be activated in vitro by the direct binding of Rap proteins to a neighbouring Ras binding domain (RBD). This activation by Rap is GTP dependent and specific for Rap versus other Ras family members. We found no evidence for direct regulation of ARAP3's Rho GAP activity by PtdIns(3,4,5)P(3) in vitro, but PI3K activity was required for activation by Rap in a cellular context, suggesting that PtdIns(3,4,5)P(3)-dependent translocation of ARAP3 to the plasma membrane may be required for further activation by Rap. Our results indicate that ARAP3 is a Rap effector that plays an important role in mediating PI3K-dependent crosstalk between Ras, Rho, and Arf family small GTPases. PMID- 15296757 TI - APOBEC3F properties and hypermutation preferences indicate activity against HIV-1 in vivo. AB - APOBEC3G (CEM15 ) deaminates cytosine to uracil in nascent retroviral cDNA. The potency of this cellular defense is evidenced by a dramatic reduction in viral infectivity and the occurrence of high frequencies of retroviral genomic-strand G --> A transition mutations. The overwhelming dinucleotide hypermutation preference of APOBEC3G acting upon a variety of model retroviral substrates is 5' GG --> -AG. However, a distinct 5'-GA --> -AA bias, which is difficult to attribute to APOBEC3G alone, prevails in HIV-1 sequences derived from infected individuals (e.g., ). Here, we show that APOBEC3F is also a potent retroviral restrictor but that its activity, unlike that of APOBEC3G, is partially resistant to HIV-1 Vif and results in a clear 5'-GA --> -AA retroviral hypermutation preference. This bias is also apparent in a bacterial mutation assay, suggesting that it is an intrinsic APOBEC3F property. Moreover, APOBEC3F and APOBEC3G appear to be coordinately expressed in a wide range of human tissues and are independently able to inhibit retroviral infection. Thus, APOBEC3F and APOBEC3G are likely to function alongside one another in the provision of an innate immune defense, with APOBEC3F functioning as the major contributor to HIV-1 hypermutation in vivo. PMID- 15296758 TI - Cytidine deamination of retroviral DNA by diverse APOBEC proteins. AB - The human cytidine deaminase APOBEC3G edits both nascent human immunodeficiency virus (HIV) and murine leukemia virus (MLV) reverse transcripts, resulting in loss of infectivity. The HIV Vif protein is able to protect both viruses from this innate restriction to infection. Here, we demonstrate that a number of other APOBEC family members from both humans and rodents can mediate anti-HIV effects, through cytidine deamination. Three of these, rat APOBEC1, mouse APOBEC3, and human APOBEC3B, are able to inhibit HIV infectivity even in the presence of Vif. Like APOBEC3G, human APOBEC3F preferentially restricts vif-deficient virus. Indeed, the mutation spectra and expression profile found for APOBEC3F indicate that this enzyme, together with APOBEC3G, accounts for the G to A hypermutation of proviruses described in HIV-infected individuals. Surprisingly, although MLV infectivity is acutely reduced by APOBEC3G, no other family member tested here had this effect. It is especially interesting that although both rodent APOBECs markedly diminish wild-type HIV infectivity, MLV is resistant to these proteins. This implies that MLV may have evolved to avoid deamination by mouse APOBEC3. Overall, our findings show that although APOBEC family members are highly related, they exhibit significantly distinct antiviral characteristics that may provide new insights into host-pathogen interactions. PMID- 15296759 TI - High coding density on the largest Paramecium tetraurelia somatic chromosome. AB - Paramecium, like other ciliates, remodels its entire germline genome at each sexual generation to produce a somatic genome stripped of transposons and other multicopy elements. The germline chromosomes are fragmented by a DNA elimination process that targets heterochromatin to give a reproducible set of some 200 linear molecules 50 kb to 1 Mb in size. These chromosomes are maintained at a ploidy of 800n in the somatic macronucleus and assure all gene expression. We isolated and sequenced the largest megabase somatic chromosome in order to explore its organization and gene content. The AT-rich (72%) chromosome is compact, with very small introns (average size 25 nt), short intergenic regions (median size 202 nt), and a coding density of at least 74%, higher than that reported for budding yeast (70%) or any other free-living eukaryote. Similarity to known proteins could be detected for 57% of the 460 potential protein coding genes. Thirty-two of the proteins are shared with vertebrates but absent from yeast, consistent with the morphogenetic complexity of Paramecium, a long standing model for differentiated functions shared with metazoans but often absent from simpler eukaryotes. Extrapolation to the whole genome suggests that Paramecium has at least 30,000 genes. PMID- 15296760 TI - Arabidopsis GNARLED encodes a NAP125 homolog that positively regulates ARP2/3. AB - In migrating cells, the actin filament nucleation activity of ARP2/3 is an essential component of dynamic cell shape change and motility. In response to signals from the small GTPase Rac1, alterations in the composition and/or subcellular localization of the WAVE complex lead to ARP2/3 activation. The human WAVE complex subunit, WAVE1/SCAR1, was first identified in Dictyostelium and is a direct ARP2/3 activator. In the absence of an intact WAVE complex, SCAR/WAVE protein is destabilized. Although the composition of the five-subunit WAVE complex is well characterized, the means by which individual subunits and fully assembled WAVE complexes regulate ARP2/3 in vivo are unclear. The molecular genetics of trichome distortion in Arabidopsis is a powerful system to understand how signaling pathways and ARP2/3 control multicellular development. In this paper we prove that the GNARLED gene encodes a homolog of the WAVE subunit NAP125. Despite the moderate level of amino acid identity between Arabidopsis and human NAP125, both homologs were functionally interchangeable in vivo and interacted physically with the putative Arabidopsis WAVE subunit ATSRA1. gnarled trichomes had nearly identical cell shape and actin cytoskeleton phenotypes when compared to ARP2/3 subunit mutants, suggesting that GRL positively regulates ARP2/3. PMID- 15296761 TI - Arabidopsis NAP1 is essential for Arp2/3-dependent trichome morphogenesis. AB - The dynamic nature of the eukaryotic actin cytoskeleton is essential for the locomotion of animal cells and the morphogenesis of plant and fungal cells. The F actin nucleating/branching activity of the Arp2/3 complex is a key function for all of these processes. The SCAR/WAVE family represents a group of Arp2/3 activators that are associated with lamellipodia formation. A protein complex of PIR121, NAP1, ABI, and HSPC300 is required for SCAR regulation by cell signaling pathways, but the exact nature of this interaction is controversial and represents a continually evolving model. The mechanism originally proposed was of a SCAR trans repressing complex supported by evidence from in vitro experiments. This model was reinforced by genetic studies in the Drosophila central nervous system and Dictyostelium, where the knockout of certain SCAR-complex components leads to excessive SCAR-mediated actin polymerization. Conflicting data have steadily accumulated from animal tissue culture experiments suggesting that the complex activates rather than represses in vivo SCAR activity. Recent biochemical evidence supports the SCAR-complex activator model. Here, we show that genetic observations in Arabidopsis are compatible with an activation model and provide one potential mechanism for the regulation of the newly identified Arabidopsis Arp2/3 complex. PMID- 15296763 TI - Framework bolsters stem cell progress. AB - With many of the leading science nations still stuck in debates on the use of embryonic stem cells, Britain, with a regulatory framework in place, is well positioned to take the lead. Michael Gross reports. PMID- 15296764 TI - 'Designer' babies. PMID- 15296765 TI - Early birds. AB - Concern is growing about the impact of climate change on organisms but there's much uncertainty about its effects. A new European study of two bird species does not provide good news. PMID- 15296766 TI - Philip Cohen. AB - Philip Cohen trained at University College London and, after postdoctoral research at the University of Washington, joined the University of Dundee Scotland, in 1971, where he has worked ever since. He is a Royal Society Research Professor and Director of the Medical Research Council Protein Phosphorylation Unit. His main contributions have been in the area of protein phosphorylation and its role in cell regulation and human disease. In 1998, he was knighted for his contributions to biochemistry and the development of Life Sciences at Dundee. PMID- 15296767 TI - Volvox. PMID- 15296769 TI - Computational biology: a propagating wave of interest. PMID- 15296771 TI - The quiet revolution. AB - Sixty years have passed since Avery, MacLeod and McCarty published their landmark paper revealing DNA as the genetic material, writes Heather Dawes. PMID- 15296772 TI - Rags before the riches: Friedrich Miescher and the discovery of DNA. PMID- 15296773 TI - We are most aware of our place in the world when about to fall. PMID- 15296774 TI - More than one way to build a flagellum: comparative genomics of parasitic protozoa. PMID- 15296775 TI - Telomeres: not all breaks are equal. AB - ATM, Rad50 and Mre11 have been shown to prevent telomere fusion in Drosophila, thereby extending the protective role of DNA damage checkpoint proteins to non canonical telomeres formed without telomerase. How do these proteins help chromosomal termini escape fusion through 'repair' while promoting repair of induced DNA breaks? PMID- 15296776 TI - Nocturnal vision: bees in the dark. AB - Some eyes work better in the dark than others. The apposition type of compound eye that bees and other diurnal insects possess is usually of little use after nightfall. Nevertheless some tropical sweat bees have pushed the limits of this unfavourable design so far that they can navigate using landmarks that are too dim for humans to make out. PMID- 15296777 TI - Invertebrate learning: what can't a worm learn? AB - The nematode worm Caenorhabditis elegans can learn and remember the stimuli it encounters, the environment it is in, and its own physiological state. Analyses of mutations in C. elegans that affect different aspects of experience are beginning to address the nature of learning. PMID- 15296778 TI - Visual development: experience puts the colour in life. AB - Recent findings show that colour processing, like most other sensory attributes, is shaped by experience. While such studies can reveal the mechanisms of development, can they also help uncover the mechanisms of perception? PMID- 15296779 TI - Molecular biology: what ubiquitin can do for transcription. AB - Ubiquitin, the peptide 'tag' that targets eukaryotic proteins for degradation by the proteasome, has also been implicated in transcriptional activation. The mechanism of gene activation might include recruitment of a transcriptional elongation factor by ubiquitinated activators. PMID- 15296780 TI - Neuronal diseases: small heat shock proteins calm your nerves. AB - Mutations in HSPB1 and HSPB8, members of the small heat shock protein family, have recently been shown to cause some distal motor neuropathies. Their function in motor neurones is now under scrutiny. PMID- 15296781 TI - Ecology: compensating for extinction. AB - Food web interactions allow communities to compensate for the loss of species. Compensation of this kind may reshuffle communities so that today's resilient species are tomorrow's vulnerable species, creating a false impression of ecosystem stability following the first wave of extinction. PMID- 15296782 TI - Homologous recombination: down to the wire. AB - Exchange of strands between homologous DNA molecules is catalyzed by evolutionarily conserved recombinases. These proteins can occur in different quaternary arrangements: rings or helical filaments. Recent results reveal that recombinase function follows from the filamentous form. PMID- 15296783 TI - Axon guidance: mice and men need Rig and Robo. AB - For many growing axons, navigating across the midline of the nervous system is a crucial stage of their development. New studies on mice and humans show that the axon guidance receptor Robo3/Rig1 is indispensable for axons to accomplish this task. PMID- 15296784 TI - Techniques of signal generation required for electropermeabilization. Survey of electropermeabilization devices. AB - Electropermeabilization is a phenomenon that transiently increases permeability of the cell plasma membrane. In the state of high permeability, the plasma membrane allows ions, small and large molecules to be introduced into the cytoplasm, although the cell plasma membrane represents a considerable barrier for them in its normal state. Besides introduction of various substances to cell cytoplasm, permeabilized cell membrane allows cell fusion or insertion of proteins to the cell membrane. Efficiency of all these applications strongly depends on parameters of electric pulses that are delivered to the treated object using specially developed electrodes and electronic devices--electroporators. In this paper we present and compare most commonly used techniques of signal generation required for electropermeabilization. In addition, we present an overview of commercially available electroporators and electroporation systems that were described in accessible literature. PMID- 15296785 TI - Electrochemical investigation of the dynamics of Mycobacterium smegmatis cells' transformation to dormant, nonculturable form. AB - Dynamics of transformation of Mycobacterium smegmatis cells by cultivation under nonoptimal conditions (partial starvation) to dormant, nonculturable form has been studied. For this aim, an electrochemical method was developed to detect both viable and 'viable but nonculturable' (VBNC) cells. The current produced by bacteria placed at the electrode surface was measured in the presence of 2,6 dichlorophenol indophenol (DCIP) at the applied potential of 350 mV. It has been established that electrochemical activity changes parallel with the growth of biomass. The transition of M. smegmatis to a dormant, nonculturable state goes along with the decrease of the detection current up to 20% of the maximum level. This means that nonculturable cells have rather high rest metabolic activity. The course of the CFU values has a complicated character during bacterial growth. The placement of the bacterial culture on the solid medium appears to cause a new stress that stops proliferation and stimulates aggregation. Both processes distort CFU measurement results. The quantitative estimation of the viable but nonculturable cells by counting colonies, measuring optical density and current produced by bacteria has been discussed. PMID- 15296786 TI - Electrochemical study of quercetin-DNA interactions: part I. Analysis in incubated solutions. AB - The present study aims to investigate the quercetin-deoxyribonucleic acid (DNA) interaction occurring in bulk solution either electrochemically using differential pulse voltammetry or spectrophotometrically, in order to explain the possible DNA-damaging activity of quercetin. A very weak interaction between quercetin and DNA in solution was found to take place. However, since extensive quercetin-induced DNA damage via reaction with Cu(II) has been reported, an electrochemical study of the DNA-Cu(II)-quercetin system in solution was undertaken. The product of DNA interaction with quercetin-Cu(II) complex was observed. Damages to DNA were electrochemically recognized via the increasing of the anodic peaks corresponding to the oxidation of guanosine and adenosine bases and spectrophotometrically via increasing of the 260 nm adsorption band. It was also observed that dsDNA damage produced by the quercetin-Cu(II) complex occurred with time. Control experiments with different mixtures of Cu(II), quercetin, ssDNA, dsDNA or poly[A] were carried out in order to establish a possible mechanism of interaction between DNA and quercetin via Cu(II). PMID- 15296787 TI - Electrochemical study of quercetin-DNA interactions: part II. In situ sensing with DNA biosensors. AB - Quercetin interaction with dsDNA was investigated electrochemically using two types of DNA biosensor in order to evaluate the occurrence of DNA damage caused by oxidized quercetin. The results showed that quercetin binds to dsDNA where it can undergo oxidation. The radicals formed during quercetin oxidation cause breaks of the hydrogen bonds in the dsDNA finally giving rise to 8-oxoguanine since the DNA guanosine and adenosine nucleotides in contact with the electrode surface can easily be oxidized. A mechanism for oxidized quercetin-induced damage to dsDNA immobilized onto a glassy carbon electrode surface is proposed and the formation of 8-oxoguanine is explained. The importance of DNA-electrochemical biosensors in the determination of the interaction mechanism between DNA and quercetin is clearly demonstrated. PMID- 15296788 TI - Static and 50 Hz magnetic fields of 0.35 and 2.45 mT have no effect on the growth of Saccharomyces cerevisiae. AB - The present work reports the growth effects induced by static and sinusoidal 50 Hz magnetic fields (MF) on the haploid yeast strain Saccharomyces cerevisiae WS8105-1C. Magnetic fields were generated by a pair of Helmholtz coils (40 cm in diameter) with 154 turns of copper wire in each and separated 20 cm. The experiments were performed at 0.35 and 2.45 mT, and yeasts were exposed to MF during 24 and 72 h in the homogeneous field area. Growth was monitored by measuring the optical density at 600 nm. The data presented in the current report indicate that static and sinusoidal 50 Hz MF (0.35 and 2.45 mT) do not induce alterations in the growth of S. cerevisiae. PMID- 15296789 TI - A comparison between the use of a redox mediator in solution and of surface modified electrodes in the electrocatalytic oxidation of nicotinamide adenine dinucleotide. AB - Cyclic voltammetry was successfully applied to study the oxidation of nicotinamide adenine dinucleotide (NADH) both in homogeneous and heterogeneous phase. The first case was realized with a solution containing p-methylamino phenolsulphate (MAP) as redox mediator and the diaphorase (DI) from Clostridium kluveri as enzyme while the second one by using both a glassy carbon (GC) and a carbon nanotube paste (CNTP) electrode modified with electrodeposited films derived from 3,4-dihydroxybenzaldehyde (3,4-DHB). Such systems were successively coupled with glucose dehydrogenase (GDH) reaction to realize the redox chain present in glucose biosensors. A critical comparison of the two systems was also reported. PMID- 15296790 TI - Artificial autonomic reflexes: using functional electrical stimulation to mimic bladder reflexes after injury or disease. AB - Autonomic reflexes controlling bladder storage (continence) and emptying (micturition) involve spinal and supraspinal nerve pathways, with complex mechanisms coordinating smooth muscle activity of the lower urinary tract with voluntary muscle activity of the external urethral sphincter (EUS). These reflexes can be severely disrupted by various diseases and by neurotrauma, particularly spinal cord injury (SCI). Functional electrical stimulation (FES) refers to a group of techniques that involve application of low levels of electrical current to artificially induce or modify nerve activation or muscle contraction, in order to restore function, improve health or rectify physiological dysfunction. Various types of FES have been developed specifically for improving bladder function and while successful for many urological patients, still require substantial refinement for use after spinal cord injury. Improved knowledge of the neural circuitry and physiology of human bladder reflexes, and the mechanisms by which various types of FES alter spinal outflow, is urgently required. Following spinal cord injury, physical and chemical changes occur within peripheral, spinal and supraspinal components of bladder reflex circuitry. Better understanding of this plasticity may determine the most suitable methods of FES at particular times after injury, or may lead to new FES approaches that exploit this remodeling or perhaps even influence the plasticity. Advances in studies of the neuroanatomy, neurophysiology and plasticity of lumbosacral nerve circuits will provide many further opportunities to improve FES approaches, and will provide "artificial autonomic reflexes" that much more closely resemble the original, healthy neuronal regulatory mechanisms. PMID- 15296791 TI - New monoclonal antibody (AIC) identifies interstitial cells of Cajal in the musculature of the mouse gastrointestinal tract. AB - Interstitial cells of Cajal (ICC) are pacemaker cells for the spontaneous muscular contractions and neuromodulators that mediate neurotransmission from enteric neurons to smooth muscle cells in the gastrointestinal (GI) tract. They express c-Kit, and the antibody for c-Kit (especially ACK2) has been a useful tool for functional and morphological studies. ACK2, however, does not work on tissues fixed with paraformaldehyde, and not all ICC express c-Kit in human. Therefore, in order to find a new marker of ICC and/or new antibody resisting aldehyde fixation, we produced a new monoclonal antibody that identifies ICC and then investigated the properties of its antigen. Isolated ICC were used for immunization. Hybridomas fused with myeloma SP2 were screened by immunohistochemistry. ACK2 and each antibody were applied on serial sections, and the clone producing anti-ICC antibody (AIC) that stains ICC was established. The distribution of AIC immunopositive cells was examined in other organs and also GI muscles of W/Wv mice. The biochemical properties were studied using dot blot analysis. AIC recognized ICC; however, distribution of immunopositive cells in W/Wv mice and other organs was different from that of c-Kit. The immunoreactivity was stable for paraformaldehyde but was blocked by either Triton X-100 or SDS. In conclusion, new antibody AIC recognized ICC but the antigen was not c-Kit, which confirms the existence of good markers of ICC besides c-Kit. Although the antigen has not been isolated, AIC is suitable for morphological study and is useful for investigation of ICC in c-Kit mutants. PMID- 15296792 TI - Cholinergic stimulation with pyridostigmine increases heart rate variability and baroreflex sensitivity in rats. AB - OBJECTIVE: Impaired parasympathetic modulation increases the risk for sudden death in patients with heart diseases. Therefore, cholinergic stimulation may have a potential protective role. The aim of this study was to verify the effects of pyridostigmine bromide, a reversible cholinesterase inhibitor, on heart rate (HR), blood pressure (BP), HR and BP variability, and baroreflex sensitivity (BS). METHODS: Male Wistar rats were divided in two groups: (1) treated with pyridostigmine in drinking water (7 days, n=10; PYR) and (2) a control group (n=12; CTR). BP was recorded in freely moving rats, and HR and BP variability were quantified by the standard deviation (S.D.) of the mean values during a 30 min period and by spectral analysis. BS was assessed by the ratio between pulse interval and BP power spectra (spontaneous BS) and also by the changes on HR produced by phenylephrine and sodium nitroprusside-induced BP changes. RESULTS: Treated rats had a PYR intake of 7.91+/-1.90 mg/day (approximately 31 mg/kg/day). There were no differences between groups concerning resting HR (P=0.158), systolic BP (P=0.481), and BP variability (P=0.201). On the other hand, treatment with PYR increased HR variability on the time domain (S.D.-PYR: 13.5+/-5.3 ms vs. CTR: 9.9+/-3.6 ms; P=0.034) and frequency domain (Total power--PYR: 208.3+/-157.7 ms(2) vs. CTR: 109.2+/-65.6 ms(2); P=0.030). BS was also augmented with PYR for both the spontaneous method (High frequency band--PYR: 2.55+/-1.06 ms/mm Hg vs. CTR: 1.85+/-0.60 ms/mm Hg; P=0.033) and the drug-induced reflex bradycardia (PYR: 2.48+/-1.02 bpm/mm Hg vs. CTR: 1.54+/-0.58 bpm/mm Hg; P=0.024) and reflex tachycardia (PYR: 4.08+/-1.04 bpm/mm Hg vs. CTR: 2.95+/-1.30 bpm/mm Hg; P=0.037). CONCLUSIONS: In conclusion, treatment with pyridostigmine increased HR variability and BS in normal rats with no modifications on basal hemodynamic parameters. Considering that reduced HR variability and baroreflex sensitivity are independent risk factors in heart disease, the present results support the concept that cholinergic stimulation with pyridostigmine may become a therapeutic option for vagal dysfunction. PMID- 15296793 TI - Nicotinic acetylcholine receptor subtypes in nociceptive dorsal root ganglion neurons of the adult rat. AB - Stimulation of nicotinic acetylcholine receptors (nAChR) excites peripheral sensory nerve fibres, but also exert antinociceptive effects. The differences in these nAChR-mediated effects could be related to the expression of different nAChR subtypes located on nociceptive neurons. In the present study, we focused on the recently described alpha 10-nAChR subunit, and on alpha 4 and alpha 7 subunits, which are the most abundant subunits in the central nervous system. In nociceptive neurons from thoracic and lumbar dorsal root ganglia (DRG), nAChR subunits were found at transcriptional (RT-PCR), translational (immunohistochemistry) and functional levels. Cultured DRG neurons express mRNA for the subunits alpha 2-7 and alpha 10. The alpha-subunit proteins 4, 7 and 10 were colocalised in virtually all nociceptive neurons that were identified by immunoreactivity for the vanilloid receptor TRPV-1. These findings were corroborated by current recordings and calcium measurements, which revealed excitatory inward currents and calcium responses in capsaicin sensitive neurons. PMID- 15296794 TI - Sequence of forebrain activation induced by intraventricular injection of hypertonic NaCl detected by Mn2+ contrasted T1-weighted MRI. AB - In order to define the sequence of forebrain activation involved in osmoregulation, central activation in response to intracerebroventricular injection of NaCl solution (10 microl of 0.15, 0.5, or 1.5 M) was detected using manganese-contrasted magnetic resonance imaging (MRI) in anesthetized rats. Changes in renal sympathetic nerve activity (RNA) were also measured, and the time courses of forebrain activation and RNA changes compared. NaCl injection resulted in rapid activation of the subfornical organ (SFO), organum vasculosum lamina terminalis (OVLT), and periventricular regions and the lateral hypothalamic area (LHA), then of the paraventricular hypothalamic nucleus (PVN) and supraoptic nucleus (SON). The delay in activation in the PVN and SON showed a wide variation from 0 to 5.78 min, and the average delay in the PVN (2.88+/-0.34 min) and SON (2.90+/-0.39 min) was significantly greater than that in the SFO (0.40+/-0.10 min) and OVLT (0.74+/-0.13 min). NaCl (1.5 M) injection elicited a rapid, large increase in RNA, which consisted of two components, an early rapid increase at 99 s after injection (160+/-27%) and a slower increase at 9 min after injection (209+/-34%). These results suggest that the PVN and SON are activated not only by the afferent input from the SFO and OVLT but also by diffusion of the hypertonic stimulus to these regions and probably by their intrinsic osmosensitivity. The PVN might be responsible for the second slower component of the RNA response, but cannot be responsible for the first component. PMID- 15296795 TI - Use of time-frequency analysis to investigate temporal patterns of cardiac autonomic response during head-up tilt in chronic fatigue syndrome. AB - Although a number of studies have reported alterations in cardiac autonomic nervous system function in chronic fatigue syndrome (CFS), the results are not consistent across studies. Reasons for these discrepancies include (1) the use of a heterogeneous patient sample that included those with orthostatic postural tachycardia (POTS), a condition with an autonomic changes, and (2) the use of frequency domain techniques which require a stationary signal and averaging data across relatively long epochs. To deal with these shortcomings, we used the smoothed pseudo-Wigner-Ville transform (SPWVT) to analyze heart rate variability (HRV) and blood pressure variability (BPV) during head-up tilt (HUT) by separating CFS patients into those with and without POTS. SPWVT has the advantage of providing instantaneous information about autonomic function under nonstable physiological conditions. We studied 18 CFS patients without POTS, eight CFS patients with POTS and 25 sedentary healthy controls during supine rest and during the first 10 min after HUT. While we found significant effects of postural change in both groups for all autonomic variables, there were significant group x time interactions between CFS without POTS and controls for only instant center frequency (ICF) within the low frequency region both from HRV (p=0.02) and from BPV (p=0.01). Although the physiological meaning of ICF still remains unknown, the data suggest that even CFS patients without POTS may have a subtle underlying disturbance in autonomic function. PMID- 15296796 TI - Effect of vitamin E on carotid artery elasticity and baroreflex gain in young, healthy adults. AB - In this study we tested the hypothesis that dietary vitamin E supplementation can improve carotid artery elasticity and cardio-vagal baroreflex gain in young, healthy individuals. A total of 20 subjects were studied in a double-blind, placebo-controlled, randomized study. Subjects in the active treatment group received 700 IU/day vitamin E for 1 month. Each subject was studied three times: before, during and 1 month after treatment. Plasma vitamin E levels were determined using high-performance liquid chromatography. Carotid artery diameter was measured by ultrasound and radial artery pressure by tonometry. Baroreflex function was assessed by time and frequency domain spontaneous indices. Plasma vitamin E levels increased by 123%, which was associated with a 20% increase in carotid artery compliance and a 30-60% increase in baroreflex indices. All these changes regressed 1 month after cessation of vitamin E supplementation. Significant correlations were observed across conditions (control, treatment and recovery), among plasma vitamin E concentrations, carotid artery compliance and distensibility values and two of the baroreflex gain indices in the treatment group. Our results demonstrate that vitamin E supplementation can increase carotid artery compliance and baroreflex gain in young, apparently healthy adults. PMID- 15296797 TI - Age-related changes in the morphology of the myenteric plexus of the human colon. AB - Aging is believed to affect the structure and function of the enteric nervous system, but little specific information on this topic is available, particularly in humans. The aim of this study was to investigate the effect of age on the structure of myenteric ganglia in the human colon. We examined myenteric ganglia in colonic specimens obtained from 168 patients aged 10 days to 91 years. Nerves were stained in whole mount preparations using the vital fluorescent dye 4-(4 dimethylaminostyryl)-methylpyridinium iodide (4-Di-2-ASP) and other staining methods. Human myenteric ganglia were classified into three types: normal, those containing empty spaces ('cavities') and those containing large nerve fiber bundles. We found a statistically significant increase with age in the proportion of ganglia with cavities. Conversely, there was a decrease with age in the proportion of normal ganglia. The proportion of fiber-containing ganglia did not change with age. These findings indicate that there is an increase with age in the number of abnormally appearing myenteric ganglia in the human colon, which may contribute to the disturbed colonic motility in the aging population. PMID- 15296798 TI - Pressor effect of water instilled via a gastrostomy tube in pure autonomic failure. AB - BACKGROUND: The oral ingestion of water increases seated blood pressure in chronic autonomic failure although the mechanisms of this effect remain unclear. Recent studies in normal subjects suggest that oropharyngeal stimulation during swallowing may be of greater importance in causing a rise in blood pressure (BP) than the gastric effects of water. We therefore assessed the haemodynamic effects of water instilled directly into the stomach via a gastrostomy tube in pure autonomic failure (PAF). METHODS: The subject had longstanding (>20 years) PAF. A gastrostomy tube had been previously placed because of dysphagia. Distilled water (480 ml) was instilled in the seated position with BP and heart rate (HR) measured over the following 40 min while the subject remained seated. Systolic and diastolic BP (SBP and DBP) and heart rate (HR) were recorded intermittently every 3 min with a Dinamap automated syphgmomanometer and continuously using a Portapres device. Subsequent model flow analysis of the Portapres data provided beat-to-beat estimates of cardiac output, stroke volume, and total peripheral resistance (TPR). Subjective orthostatic symptoms were recorded before and after water. RESULTS: Seated SBP and DBP increased after water instillation with increases first noted between 5 and 8 min after the water had been instilled. The BP remained elevated until 35 min post water increase over baseline being +36.5 mm Hg SBP and +24.3 mm Hg DBP. HR, cardiac output, and stroke volume remained unchanged during the study. Total peripheral resistance (TPR) increased post water. These results are similar to those reported in a recent study involving oral ingestion of 480 ml of water in PAF subjects. CONCLUSIONS: Instilling water directly into the stomach in a patient with PAF resulted in similar haemodynamic responses to those seen when water is taken orally. Thus, oropharyngeal factors and swallowing do not appear to be essential in the generation of the water pressor effect in autonomic failure. PMID- 15296809 TI - Special populations recruitment for the Women's Health Initiative: successes and limitations. AB - The Women's Health Initiative (WHI) is a study designed to examine the major causes of death and disability in women. This multi-arm, randomized, controlled trial of over 160,000 post-menopausal women of varying ethnic and socioeconomic backgrounds and a goal of 20% of the study participants from minority populations is perhaps one of the most challenging recruitment efforts ever undertaken. Of the two main study arms, the Clinical Trial (CT) and the Observational Study (OS), the CT arm recruitment goal was to randomize 64,500 postmenopausal women 50 79 years of age. Women enrolled in the study will be followed for a period of 8 12 years. Ten clinical centers, out of a total of 40 throughout the United States, were selected as minority recruitment centers on the basis of their history of interaction with and access to large numbers of women from certain population subgroups. WHI enrollment began in September 1993 and ended in December 1998, resulting in the randomization and enrollment of a total of 161,856 (17.5% minority) women participants (68,135 (18.5% minority) in the CT and 93,721 (16.7%) in the OS). Within the CT arm, WHI achieved 101.7% of the goal of 48,000 participants in the Dietary Modification (DM) component, and 99.4% of the goal of 27,500 in the hormone-replacement component (HRT), with 11.8% overlap between DM and HRT. Of those who expressed initial interest in WHI, African Americans had the highest randomization yields in the DM component and Hispanics had the highest in the HRT component (15.2% and 10.2%, respectively). Overall, mass mailing was the greatest source of randomized participants. In addition, minority clinics found community outreach, personal referrals, and culturally appropriate recruitment materials particularly effective recruitment tools. For minority recruitment, our findings suggest that the key to high yield is reaching the target population through appropriate recruitment strategies and study information that get their attention. Also, once minority subjects are reached, they tend to participate. PMID- 15296810 TI - Issues in the design of a clinical trial with a behavioral intervention--the Zambia exclusive breast-feeding study. AB - PURPOSE: We present the rationale and design of the Zambian Exclusive Breast feeding Study (ZEBS), a randomized trial evaluating the efficacy of short duration exclusive breast-feeding (EBF) as a strategy to reduce postnatal human immunodeficiency virus (HIV) transmission while preserving the other health benefits of this important mode of infant feeding. METHODS: One thousand two hundred HIV-positive pregnant women were recruited in Lusaka, Zambia, and followed with their infants for 24 months. In addition to Nevirapine (NVP), all women received intensive and frequent clinic- and home-based counseling to support exclusive breast-feeding. When the infant was 1 week of age, half of the women were randomly assigned to a group encouraged to abruptly (<24 h) cease all breast-feeding at 4 months. The primary outcome of the experimental (randomized) comparison is HIV-free survival at 24 months. The design is also observational and will compare HIV transmission rates between those who do and do not adhere to the counseling intervention promoting exclusive breast-feeding. CONCLUSION: Our study aims to quantify the benefit-risk ratio of early cessation of exclusive breast-feeding to interrupt mother-to-child transmission of HIV with an intensive behavioral intervention and has both observational and experimental analytic approaches. Our study design assesses efficacy and also has a prominent applied component that if the intervention is effective, it will permit rapid and sustainable adoption within low-resource communities. PMID- 15296811 TI - Investigator and site selection and performing GCP clinical studies in India. AB - The optimum site and investigator selection process remains a closely guarded confidential matter and an essential part of development expertise of big pharmaceutical companies and CROs. The right and careful selection and evaluation of investigators and site is critical for successful completion of the trial within budget, timelines and generation of high quality data. The criteria for site and investigator selection in India for Good Clinical Practices (GCP) clinical trials are described for a start up company/CRO and can be applied to any country in Asia and Africa. Foreign sponsors doing clinical studies in India should pay close attention to site and investigator selection. The first GCP study in India was done only in 1995. At the dawn of 21st century, India is at the take off stage in clinical trials now. GCP studies can be done in India, as the quality of data is good, costs are lower and patient enrolment is much faster resulting in early completion of studies. PMID- 15296812 TI - Enrollment in clinical trials according to patients race: experience from the VA Cooperative Studies Program (1975-2000). AB - BACKGROUND: Racial distribution of clinical trial participants is important because results from these studies serve to define evidence-based practice. This report summarizes the experience of the VA Cooperative Studies Program (CSP) in enrolling white, black and Hispanic patients. METHODS: An analysis of enrollment in randomized controlled trials conducted by VA CSP between 1975 and 2000. A standardized enrollment ratio for each trial was calculated by dividing the observed number of enrolled white patients in the trial by the expected number of eligible white patients based on the proportion of white patients hospitalized at the enrolling VA Medical Centers. RESULTS: 138 VA CSP clinical trials were initiated between 1975 and 2000, 83 contained information on race for 71,463 patients. Overall, 76% of enrolled patients were white, 20% were black, and 4% were Hispanic. Based on standardized enrollment ratios, 60 of the 83 trials had 95% confidence intervals that excluded 1.0. Of these, 32 studies enrolled more white patients than expected and 28 enrolled more Black and/or Hispanic patients than expected based on the racial distribution of patients hospitalized at sites involved in the trials. When trials were separated by intervention type, 13 of the 19 trials that had an invasive arm enrolled fewer minority patients than expected. In trials that targeted diseases that affect minority populations to a greater degree than whites (diabetes, hypertension and end stage renal disease), 11 of the 14 trials enrolled more minority patients than expected. CONCLUSIONS: There were several trials that enrolled either more or less minority patients than expected based on patients hospitalized at study sites. Trials that included an invasive arm enrolled fewer minority participants than expected. Trials that involve invasive therapies may wish to adopt special recruitment strategies to reach minority populations. PMID- 15296813 TI - When free condoms and spermicide are not enough: barriers and solutions to participant recruitment to community-based trials. AB - While randomised controlled trials remain the accepted 'gold standard' in medical research, participant recruitment is often problematic, particularly with primary care trials or those requiring healthy volunteers. Such difficulties can jeopardise the trial, leading to early abandonment, reduced statistical power or timetable and budget overruns. Substantial changes in recruitment plans may reduce the generalisability of the research. In order to overcome some of the more common recruitment difficulties, it is important that researchers share their recruitment strategy successes and failures. We report our experience of recruiting healthy volunteers to a condom trial, based within primary care and community populations. This was an RCT of the effect that using an additional spermicidal lubricant has on condom failure. We originally aimed to recruit entirely from Family Planning Clinics, but eventually required a wide variety of strategies. Targeted mailings, newspaper coverage and electronic 'posters' were among the most successful we used to bolster clinic recruitment. Concerned at our slow recruitment rates, we conducted a questionnaire survey investigating the reasons for participation and non-participation in the research completed by 101 trial participants, 112 decliners and 90 controls (total 303). The most important reasons given for taking part included 'considering the research to be important' (85%), 'wanting to help the researchers' (70%), 'having time to help' (62%) and 'getting free condoms and lubricant' (56%). The most popular reasons for declining were 'not wanting to use condoms' (38%), 'partner's unwillingness to take part' (29%), 'not wanting to alter usual contraceptive practice' (27%), 'not having time' (21%). Contrary to expectations, embarrassment and fears about confidentiality were relatively unimportant factors in this decision. In conclusion, the key to attaining recruitment targets was the core research team taking an active part, working closely with clinic staff and maintaining tight control of the process. Altruism remains a powerful motivation for participants, supported by incentives and procedural details to minimise personal inconvenience. Even for intimate research topics, these general factors outweigh specific issues. PMID- 15296814 TI - Surveillance of the eye and vision in a clinical trial of MART1-transformed dendritic cells for metastatic melanoma. AB - PURPOSE: To report the protocol for surveillance of the eye and vision in a clinical trial of MART1-transduced dendritic cells for metastatic melanoma. METHODS: In a phase I/II clinical trial of dendritic cell-based genetic immunotherapy for metastatic cutaneous melanoma, ophthalmic evaluation is performed prior to immunization (Baseline Evaluation), 56+/-7 days after first vaccination (mid-study evaluation), when dendritic cell injections are complete 112+/-7 days after first vaccination (end-study evaluation) and 168+/-7 days after first vaccination (post-study evaluation). RESULTS: The protocol for baseline, mid-study and end-study evaluations of the eye and vision includes ophthalmic history, comprehensive ophthalmic examination, psychophysical and electrophysiological visual function assessment, fundus photography and fluorescein angiography. Post-study evaluation consists of the 25-item visual functioning questionnaire augmented to elicit autoimmune manifestation with complete ophthalmic evaluation if vision-related symptoms or abnormalities are noted during or after the vaccination. CONCLUSION: Limited adverse effects on the eye and vision have been reported in melanoma immunotherapy trials, although this novel mode of therapy has the potential to induce melanoma paraneoplastic syndromes known to severely impair vision. Therefore, surveillance of the eye and vision should be considered in melanoma immunotherapy trials. PMID- 15296815 TI - Adjustment for baseline measurement error in randomized controlled trials induces bias. AB - When estimating the treatment effect in a randomized controlled trial, it is common to have a continuous outcome which is also observed at baseline. These observations are often prone to measurement error, for example due to within patient variability. Controversy exists in the literature about whether baseline measurement error should be adjusted for in this context. Computer simulations were used to compare the biases in the estimated treatment effect, with and without adjusting for measurement error, and for different levels of observed baseline imbalance. The impacts of sample size (30 per group and 300 per group) and reliability coefficient (0.6, 0.8 and 1) were also assessed. The results show that in randomized controlled trials, the ordinary least squares (OLS) estimator without adjusting for measurement error is unbiased. On the contrary, adjusting for measurement error leads to bias, especially when sample sizes are small and/or measurement error is large. The treatment effect adjusting for measurement error is on average overestimated when the baseline mean of the control group is larger than that of the treated group. It is underestimated when the control group has a smaller baseline mean. PMID- 15296816 TI - Incorporating the sampling variation of the disease prevalence when calculating the sample size in a study to determine the diagnostic accuracy of a test. AB - During the design stage of a study to assess the population sensitivity (P(S)) (or specificity) of a diagnostic test, the number of subjects (N) who will be administered both a gold standard test and a new test needs to be calculated. A common approach is to calculate the number of cases (n) with a specific disease or condition as diagnosed by the gold standard test first, and then to determine N based on the prevalence or incidence rate of the disease (P(P)) in the population, calculated as N=n/P(P). Due to sampling variation, given the sample size N, the number of cases having the disease identified by the gold standard test could be less than N x P(P). In this case, the study would be under-powered and may fail to produce an unbiased and precise estimate. In this study, we investigated this possibility for a situation where the required sample size is calculated using the confidence interval approach. When the sampling variation is considered, the variance of the sample sensitivity is slightly inflated, but its confidence interval width becomes widely dispersed. In order to reach the originally designed precision, adjustment in the sample size, N, is needed and suggested in this paper. PMID- 15296817 TI - Alcohol-related expectancies are associated with the D2 dopamine receptor and GABAA receptor beta3 subunit genes. AB - Molecular genetic research has identified promising markers of alcohol dependence, including alleles of the D2 dopamine receptor (DRD2) and the GABAA receptor beta3 subunit (GABRB3) genes. Whether such genetic risk manifests itself in stronger alcohol-related outcome expectancies, or in difficulty resisting alcohol, is unknown. In the present study, A1+ (A1A1 and A1A2 genotypes) and A1- (A2A2 genotype) alleles of the DRD2 and G1+ (G1G1 and G1 non-G1 genotypes) and G1 (non-G1 non-G1 genotype) alleles of the GABRB3 gene were determined in a group of 56 medically ill patients diagnosed with alcohol dependence. Mood-related alcohol expectancy (AE) and drinking refusal self-efficacy (DRSE) were assessed using the Drinking Expectancy Profile (Manual for the Drinking Expectancy Profile, Behaviour Research and Therapy Centre, Brisbane, 1996). Patients with the DRD2 A1+ allele, compared with those with the DRD2 A1- allele, reported significantly lower DRSE in situations of social pressure. Similarly, lower DRSE was reported under social pressure by patients with the GABRB3 G1+ allele when compared to those with the GABRB3 G1- alleles. Patients with the GABRB3 G1+ allele also revealed reduced DRSE in situations characterized by negative affect than those with the GABRB3 G1- alleles. Patients carrying the GABRB3 G1+ allele showed stronger AE relating to negative affective change (for example, increased depression) than their GABRB3 G1- counterparts. Biological influence in the development of some classes of cognitions is hypothesized. The clinical implications, particularly with regard to patient-treatment matching and the development of an integrated psychological and pharmacogenetic approach, are discussed. PMID- 15296818 TI - Sex hormones and biogenic amine turnover of sex offenders in relation to their temperament and character dimensions. AB - Relationships between Cloninger's temperament and character dimensions and plasma sex hormone levels and biogenic amine turnover were studied in male prison inmates convicted of rape (n=61) or child molestation (n=24) and normal male controls (n=25). The participants completed the Temperament and Character Inventory (TCI), which includes the temperament dimensions Novelty Seeking, Harm Avoidance, Reward Dependence and Persistence as well as the character dimensions Self-Directedness, Cooperativeness and Self-Transcendence. Plasma levels of testosterone, dihydrotestosterone, sex hormone binding globulin, luteinizing hormone (LH) and follicle-stimulating hormone were estimated in plasma samples and 5-hydroxyindoleacetic acid (5-HIAA), homovanillic acid (HVA), and 3-methoxy-4 hydroxyphenylglycol (MHPG) in urine samples. Both sex offender groups had higher Novelty, Seeking and lower Reward Dependence, Self-Directedness and Cooperativeness scores compared with the controls. Plasma levels of testosterone and dihydrotestosterone were significantly higher in rapists than in controls. Novelty Seeking scores were positively correlated with LH levels in rapists, and with testosterone levels in child molesters. Harm Avoidance scores were negatively correlated with 5-HIAA levels in rapists and with HVA levels in child molesters. In rapists, the calculated free androgen index showed a negative correlation with 5-HIAA. For the sex offender sample as a whole, the subgroup with high testosterone levels had higher Harm Avoidance scores, the subgroup with low HVA levels had lower Cooperativeness scores, and the subgroups with high 5HIAA or MHPG levels had lower Persistence scores. The results indicate that Novelty Seeking behavior in the group of rapists is associated with a hyperactive hypothalamic-pituitary-gonadal axis. In addition, low serotonin turnover and low dopamine turnover seem to be associated with a passive-avoidant behavioral style in rapists and child molesters, respectively. PMID- 15296819 TI - Modulating sensorimotor gating in healthy volunteers: the effects of desipramine and haloperidol. AB - In schizophrenia both an involvement of a reduced prefrontal dopaminergic activity and an enhanced noradrenergic activity have been suggested. In addition, patients suffering from schizophrenia show reduced sensorimotor gating and reduced habituation. If there is a causality between these neurotransmitters and these processes, then either a reduction in dopaminergic activity or an enhanced noradrenergic activity in healthy volunteers would result in reduced sensorimotor gating and reduced habituation. In the present study, a group of 12 healthy male volunteers was tested four times in a prepulse inhibition (PPI) paradigm 2.5 h following administration of placebo/placebo, placebo/desipramine (50 mg), placebo/haloperidol (2 mg) and desipramine (50 mg)/haloperidol (2 mg). A significant reduction of percentage PPI was found in all active treatments compared with placebo/placebo, while no treatment effects on habituation were found. Furthermore, a significant increase in heart rate was found in both desipramine treatments, from 120 min following oral intake onwards. Both desipramine and haloperidol reduced PPI, which suggests that an enhanced noradrenergic activity and a reduced dopaminergic activity lead to a reduction in sensorimotor gating. Since reduced sensorimotor gating is found in schizophrenia, these results supply further evidence for a reduced prefrontal dopaminergic activity and an enhanced noradrenergic activity in schizophrenia. Furthermore, the combination of haloperidol and desipramine did not have a synergistic effect on PPI, which indicates an interaction between the compounds. The site for this interaction is most likely located in the prefrontal cortex, since evidence is accumulating that extracellular dopamine concentration is regulated by noradrenergic terminals, particularly in the frontal areas of the brain. Since no effects on habituation were found, this suggests that neither enhanced noradrenergic nor decreased dopaminergic activity is involved in this process. PMID- 15296820 TI - Parametric study of accuracy and response time in schizophrenic persons making visual or auditory discriminations. AB - The inability to modulate processing time in conjunction with varying difficulty levels may be a core component of schizophrenia's cognitive deficit. In this study we used a parametric design to demonstrate this group's inability to increase and decrease response times in association with varying levels of task demand during auditory and visual recognition tasks. Unlike participants with schizophrenia, healthy volunteers responded to increasing levels of difficulty and high error by robustly increasing their average response times. In the group with schizophrenia, the greater the correlation between a subject's Response-Time and error rate the better was the subject in his/her overall discrimination accuracy. The higher their correlations the better they performed across all levels of difficulty in both modalities. The schizophrenia group's tendency to process high and low error conditions with similar behavioral resources may reflect a relatively static, non-dynamic cognitive repertoire. PMID- 15296821 TI - Platelet serotonin and plasma prolactin and cortisol in healthy, depressed and schizophrenic women. AB - Serotonin (5-hydroxytryptamine, 5-HT) is involved in the regulation of hypothalamic-pituitary-adrenal axis (HPA) activity and prolactin (PRL) secretion. The present study examined the relationship between platelet 5-HT and plasma cortisol and PRL concentrations in 20 schizophrenic, 25 depressed, and 25 healthy women. At the time of blood sampling, the schizophrenic and depressed patients had been drug-free for at least 7 days. Platelet 5-HT, plasma cortisol and PRL concentrations were determined by spectrofluorimetric, radioimmunoassay and immunoradiometric methods, respectively. Platelet 5-HT concentration was significantly higher in schizophrenic patients than in depressed patients or in healthy controls, while it was significantly lower in depressed patients than in healthy controls or in schizophrenic patients. Plasma cortisol levels were significantly increased both in schizophrenic and in depressed patients compared with values in healthy controls. Values of plasma PRL were similar across groups. A significant correlation was found between platelet 5-HT and plasma cortisol, and platelet 5-HT and plasma PRL concentrations in healthy controls, but not in schizophrenic or depressed patients. There was no significant relationship between plasma PRL and cortisol levels in any of the groups. Our data, although obtained on peripheral biochemical markers, indicate that depression and schizophrenia are characterized by disturbed 5-HT transmission and dysregulated HPA axis activity. PMID- 15296822 TI - Serum prolactin levels in unmedicated first-episode and recurrent schizophrenia patients: a possible marker for the disease's subtypes. AB - Various studies indicate that we must consider schizophrenia not as a single disease but as several distinct etiological processes that give rise to characteristic symptoms. In the current study, we aimed to examine prolactin serum levels in unmedicated first-episode and recurrent schizophrenic patients. The prolactin levels were compared among the different schizophrenia subtypes, i.e. paranoid, schizoaffective and disorganized. Prolactin serum samples were assessed on the morning after the admission in 48 first-episode and 38 recurrent unmedicated hospitalized schizophrenia patients. Two psychiatrists made the diagnosis without knowledge of laboratory results and completed the rating scales. Despite all prolactin levels being within or close to the normal range, we found significant differences in prolactin serum levels among schizophrenia subtype patients: the lowest values were for the paranoid type, intermediate for the schizoaffective and the highest for the disorganized patients. The results seem to indicate a pronounced hyperdopaminergic activity in paranoid schizophrenia, suggesting differences in dopaminergic tone between the schizophrenia subtypes, and support the clinical and the neuropsychological individuality of disease subtypes. There were no significant differences in prolactin serum levels of the schizophrenia subtypes between the first-episode and the recurrent patients. It appears that there are constant patterns of dopamine bioactivity in acutely psychotic unmedicated schizophrenia patients, whether the patients are first admitted or recurrent. PMID- 15296823 TI - Potential psychosocial mechanisms linking depression to immune function in elderly subjects. AB - Although depression and immune changes in elderly subjects constitute a considerable health risk, mechanisms underlying the association between depression and immune function are unclear. The question of whether personality and social support can explain the variation in immune function during depression was addressed in 21 elderly depressive and 23 control subjects. The following variables were studied: neuroticism, extraversion, received social support, depression-related immune parameters [i.e. numbers of lymphocytes, lymphocyte subsets CD3+, CD8+, natural killer-like T cells (NKT), CD4/CD8 ratio, and interleukin-6 (Il-6)]. We found that neuroticism reduced the association between depression and Il-6 (from 62 to 22.4%) and between depression and CD3+ (from 27.6 to 21.6%), and was also directly related to Il-6 (i.e. adjusted for age and depression). Social support reduced the association between depression and NKT cells from 25 to 18%, while it was also directly related to NKT cells. Extraversion, adjusted for age and depression, was negatively related to CD4/CD8 ratio. Subjects with high extraversion and high social support had more NKT cells. We concluded that changes in immune function during depression can partly be explained by neuroticism and received social support, whereas immune function is also directly related to these psychosocial variables. Neuroticism may exert its contribution to the risk for depression partly via Il-6 production. PMID- 15296824 TI - Mixed states in bipolar II disorder: should full hypomania always be required? AB - The aim of the study was to test whether the definition of depressive mixed states (DMX) in bipolar II disorder should require satisfaction of full criteria for hypomania or if only a few hypomanic symptoms should be required. Consecutive outpatients with bipolar II major depressive episode (MDE) (n=260) were assessed with the Structured Clinical Interview for DSM-IV. Presence of hypomanic symptoms during MDE was systematically assessed, and symptoms were graded by rating scale. The following three definitions of DMX were compared: (1) MDE plus full criteria for hypomania, (2) MDE plus three or more hypomanic symptoms (DMX3), and (3) MDE plus one or two hypomanic symptoms (DMX1-2). DMX definitions were compared on variables typically associated with bipolar disorders (young age of onset, many recurrences, atypical features of depression, and bipolar family history). The distributions of hypomanic symptom scores, age, and age of onset were studied by Kernel density estimation curves and by histograms. Bimodality would support distinct disorders, whereas lack of bimodality would support continuity among the different DMX definitions. The frequency of DMX+full hypomania was 12.3%, that of DMX3 was 46.9%, and that of of DMX1-2 was 38.8%. Comparisons among the groups on bipolar validators found that most differences were not significant. Kernel density estimation curves and histograms did not show bimodality, and had near normal distribution shapes. The findings do not support a categorical definition of bipolar II DMX like that of DSM-IV for bipolar I mixed state but are consistent with a dimensional definition of bipolar II DMX. The high frequency of DMX in bipolar II MDE supports the need for controlled studies to test the effects of antidepressants on depressive mixed state (as clinical observations suggest possible negative effects). PMID- 15296825 TI - Neuropsychological investigation of decision-making in anorexia nervosa. AB - Anorexia nervosa (AN) could be considered a form of obsessive-compulsive disorder in which an impairment of the cognitive domain related to decision-making was found. We explored this function in AN patients, as well as possible differences between restricting type and binge/purge type, with the aim of examining the hypothesis that AN is part of the obsessive-compulsive spectrum. Decision-making was assessed in 59 inpatients with AN and 82 control subjects using the Gambling task, which simulates real-life decision-making by assessing the ability to balance immediate rewards against long-term negative consequences. We confirmed the supposed deficit of decision-making in AN. However, restricting and binge eating/purge subtypes showed different patterns of decision-making impairment. Poor performance on the Gambling task is not a mere consequence of starvation and does not appear to be related to illness severity. The decision-making deficiency that some AN patients show is linked to those individual features that contribute to the phenomenological expression of the disorder. PMID- 15296826 TI - Perceived parental rearing style in obsessive-compulsive disorder: relation to symptom dimensions. AB - Obsessive-compulsive disorder (OCD) runs in families, but the specific contribution of genetic and environmental factors to its development is not well understood. The aim of this study was to assess whether there are differences in perceived parental child-rearing practices between OCD patients and healthy controls, and whether any relationship exists between parental characteristics, depressive symptoms and the expression of particular OCD symptom dimensions. A group of 40 OCD outpatients and 40 matched healthy controls received the EMBU (Own Memories of Parental Rearing Experiences in Childhood), a self-report measure of perceived parental child-rearing style. The Yale-Brown Obsessive Compulsive Scale (Y-BOCS) and the Hamilton Depression Rating Scale (HDRS) were used to assess the severity of obsessive-compulsive and depressive symptoms. The Y-BOCS Symptom Checklist was used to assess the nature of obsessive-compulsive symptoms, considering the following five symptom dimensions: contamination/cleaning, aggressive/checking, symmetry/ordering, sexual/religious and hoarding. Logistic and multiple linear regression analyses were conducted to study the relationship between parental style of upbringing, depressive symptoms and OCD symptom dimensions. Severe OCD (Y-BOCS: 27.0+/-7.4) and mild to moderate depressive symptoms (HDRS: 14.0+/-5.4) were detected in our sample. Compared with healthy controls, OCD patients perceived higher levels of rejection from their fathers. No differences between the groups with respect to perceived levels of overprotection were detected. The seventy of depressive symptoms could not be predicted by scores on any perceived parental characteristics. Hoarding was the only OCD symptom dimension that could be partially predicted by parental traits, specifically low parental emotional warmth. Social/cultural variables such as parental child-rearing patterns, in interaction with biological and genetic factors, may contribute to the expression of the OCD phenotype. PMID- 15296827 TI - Monocyte chemoattractant protein-1 (MCP1) promoter -2518 polymorphism may confer a susceptibility to major depressive disorder in the Korean population. AB - We conducted a case-control association study of the monocyte chemoattractant protein-1 (MCP1) gene -2518 polymorphism in 90 patients with major depressive disorder. Genotyping was performed by polymerase chain reaction methods. We found significant differences in genotype and allele frequencies. The present study suggests that this polymorphism may confer a susceptibility to major depressive disorder in the Korean population. PMID- 15296828 TI - Capacity of the 10-item Depression in the Medically Ill screening measure to detect depression 'caseness' in psychiatric out-patients. AB - The 10-item Depression in the Medically Ill (DMI-10) screening measures has been demonstrated to be useful in medically ill and general practice patients. Its usefulness as a screening or monitoring measure in depressed psychiatric out patients is now reported. One hundred subjects-currently depressed or recovered from a recent episode-completed the measure, with scores for those 69 currently meeting DSM-IV depression caseness criteria compared with 31 non-depressed subjects. A cut-off score of 10 or more had high sensitivity and specificity for discriminating between 'cases' and 'non-cases'. The discriminating capacity of each item was also quantified. We conclude that the DMI-10 is brief, gender non specific and less intrusive than many depression screening measures in clinical practice with depressed patients, with the currently established cut-off score similar to that established in medically ill samples. Analyses suggest useful items for clinicians to determine depression caseness status. PMID- 15296829 TI - Rise and fall of minocycline in neuroprotection: need to promote publication of negative results. AB - Initial studies conducted on the neuroprotective effects of minocycline, a second generation tetracycline, in experimental models of neurodegeneration gave promising results. However, more recently, minocycline has clearly been shown to have variable and even contradictory (beneficial or detrimental) effects in different species and models of neurological disorders, and its "neuroprotective" mechanisms remain to be clarified. Although its anti-inflammatory properties are likely to contribute to its neuroprotective effects observed in several animal models, a body of recent evidence indicates that our community should proceed with caution in the clinical use of minocycline for central nervous system disorders. PMID- 15296830 TI - Calcium channel blockers ameliorate disease in a mouse model of multiple sclerosis. AB - Multiple sclerosis (MS) and experimental autoimmune encephalomyelitis (EAE), an animal model of MS, are inflammatory demyelinating diseases of the central nervous system. The inflammatory attacks lead to glial dysfunction and death, axonal damage, and neurological deficits. Numerous studies in rat suggest that extracellular calcium influx, via voltage-gated calcium channels (VGCC), contributes to white matter damage in acute spinal cord injury and stroke. Our immunohistochemical finding that mouse spinal cord axons display subunits of L type VGCC also supports this hypothesis. Furthermore, we hypothesized that VGCC also play a role in EAE, and possibly, MS. In our study, administration of the calcium channel blockers (CCB) bepridil and nitrendipine significantly ameliorated EAE in mice, compared with vehicle-treated controls. Spinal cord samples showed reduced inflammation and axonal pathology in bepridil-treated animals. Our data support the hypothesis that calcium influx via VGCC plays a significant role in the development of neurological disability and white matter damage in EAE and MS. PMID- 15296831 TI - Regulation of ischemic cell death by the lipoic acid-palladium complex, Poly MVA, in gerbils. AB - Modulation of ischemic cell death can be accomplished via a multitude of mechanisms, such as quenching radical species, providing alternative energy sources, or altering glutamate excitation. Transient cerebral ischemia will induce apoptotic cell death selectively to hippocampal cornus ammon's field 1 of the hippocampus (CA1) pyramidal cells, while neighboring CA3 and dentate neurons are spared. Poly MVA is a dietary supplement based on the nontoxic chemotherapeutic lipoic acid-palladium complex (LAPd). LAPd is a liquid crystal that works in cancer cells by transferring excess electrons from membrane fatty acids to DNA via the mitochondria. Therefore, by its structural nature and action as a redox shuttle, it can both quench radicals as well as provide energy to the mitochondria. To understand the role of LAPd in regulating ischemic cell death, we studied Poly MVA. Male Mongolian gerbils were subjected to 5 min of bilateral carotid artery occlusion under a controlled temperature environment (37.0-38.0 degrees C). Animals were injected with physiological saline or either 30, 50, or 70 mg/kg of Poly MVA every 24 h beginning immediately after the occlusion until being sacrificed on experimental day 4. Damage was evaluated by analyzing nesting behavior and conducting blinded measures of viable CA1 lengths. All Poly MVA treatment dosages significantly (p < 0.05) reduced hippocampal CA1 damage by 72 h. Nesting scores were significantly improved after 30 and 50 mg/kg treatment but not 70 mg/kg. While nesting is usually a very accurate indicator of morphological damage, the 70 mg/kg-treated animals demonstrated excessive energy, thus ignoring the nesting material. While numerous routes offer varying degrees of CA1 neuronal survival after transient global ischemia, only the LAPd complex, which quenches radicals and provides energy to stabilize the mitochondria, offers such significant protection. Thus, the administration of Poly MVA may be a potent neuroprotective agent for victims of transient ischemic attack (TIA), cardiac arrest, anesthetic accidents, or drowning. PMID- 15296832 TI - Targeted disruption of PSD-93 gene reduces platelet-activating factor-induced neurotoxicity in cultured cortical neurons. AB - PSD-93, a molecular adaptive protein, binds to and clusters the N-methyl-D aspartate (NMDA) receptor and assembles a specific set of signaling proteins (for example neuronal nitric oxide synthase, nNOS) around the NMDA receptor at synapses in the central nervous system. This suggests that PSD-93 might mediate many NMDA receptor-dependent physiological and pathophysiological functions. We report here that PSD-93 colocalizes and interacts with the NMDA receptor and neuronal nitric oxide synthase in cultured cortical neurons. Targeted disruption of PSD-93 gene significantly prevented NMDA receptor-nitric oxide signaling dependent neurotoxicity triggered via platelet-activating factor (PAF) receptor activation. In addition, the deficiency of PSD-93 markedly attenuated platelet activating factor-induced increase in cyclic guanosine 3',5'-monophosphate (cGMP) and prevented platelet-activating factor-promoted formation of NMDA receptor neuronal nitric oxide synthase complex. These findings indicate that PSD-93 is involved in the NMDA receptor--nitric oxide-mediated pathological processing of neuronal damage triggered via platelet--activating factor receptor activation. Since platelet-activating factor is a potent neuronal injury mediator during the development of brain trauma, seizures, and ischemia, the present work suggests that PSD-93 might contribute to molecular mechanisms of neuronal damage in these brain disorders. PMID- 15296833 TI - The role of endogenous versus exogenous tPA on edema formation in murine ICH. AB - To minimize the neurotoxic injury by clot-derived substances after intracerebral hemorrhage (ICH) on the surrounding brain tissue, minimally invasive neurosurgical protocols have evolved evacuating the hematoma by stereotaxic injection of a fibrinolytic agent such as recombinant tissue plasminogen activator (rtPA), followed by aspiration of the lysed clot. However, the possible contribution of the presence of exogenous tPA itself to the toxic effects of hematoma-derived factors complicates the rationale and efficacy of this therapeutic approach. To clarify the role of exogenous rtPA on edema development, we examined the extent of edema formation in a murine model of collagenase induced ICH, which included tPA-deficient (tPA-/-) and wild-type (wt) mice. In 16 (7 tPA-/- and 9 wt mice) out of 32 mice, 1 mg/kg rtPA was injected into the hematoma 5 h after ICH induction followed by aspiration of the liquefied clot 20 min later. In the control group (8 tPA-/- and 8 wt mice), only collagenase was injected. The edema volume was quantified using SPOT software on Luxol Fast Blue and Cresyl violet-stained cross-sections 24 h, 3, and 7 days post surgery. Twenty four hours after ICH induction, tPA-/- mice had a significantly smaller edema volume (P< 0.01), even when rtPA was administered. Between days 3 and 7 after ICH, exogenous rtPA exerts its edema-promoting effect irrespective of the underlying genotype and exhibits an extensive microglial activation adjacent to the clot. In conclusion, the role of the endogenous tPA appears to be limited to the early phase of edema formation, whereas exogenous rtPA is edema-promoting between days 3 and 7 after ICH. PMID- 15296834 TI - Treatment of spinal cord injury by transplantation of fetal neural precursor cells engineered to express BMP inhibitor. AB - Spontaneous recovery after spinal cord injury is limited. Transplantation of neural precursor cells (NPCs) into lesioned adult rat spinal cord results in only partial functional recovery, and most transplanted cells tend to differentiate predominantly into astrocytes. In order to improve functional recovery after transplantation, it is important that transplanted neural precursor cells appropriately differentiate into cell lineages required for spinal cord regeneration. In order to modulate the fate of transplanted cells, we advocate transplanting gene-modified neural precursor cells. We demonstrate that gene modification to inhibit bone morphogenetic protein (BMP) signaling by noggin expression promoted differentiation of neural precursor cells into neurons and oligodendrocytes, in addition to astrocytes after transplantation. Furthermore, functional recovery of the recipient mice with spinal cord injury was observed when noggin-expressing neural precursor cells were transplanted. These observations suggest that gene-modified neural precursor cells that express molecules involved in cell fate modulation could improve central nervous system (CNS) regeneration. PMID- 15296835 TI - Rubrospinal neurons fail to respond to brain-derived neurotrophic factor applied to the spinal cord injury site 2 months after cervical axotomy. AB - Numerous experimental therapies to promote axonal regeneration have shown promise in animal models of acute spinal cord injury, but their effectiveness is often found to diminish with a delay in administration. We evaluated whether brain derived neurotrophic factor (BDNF) application to the spinal cord injury site 2 months after cervical axotomy could promote a regenerative response in chronically axotomized rubrospinal neurons. BDNF was applied to the spinal cord in three different concentrations 2 months after cervical axotomy of the rubrospinal tract. The red nucleus was examined for reversal of neuronal atrophy, GAP43 and Talpha1 tubulin mRNA expression, and trkB receptor immunoreactivity. A peripheral nerve transplant paradigm was used to measure axonal regeneration into peripheral nerve transplants. Rubrospinal axons were anterogradely traced and trkB receptor immunohistochemistry performed on the injured spinal cord. We found that BDNF treatment did not reverse rubrospinal neuronal atrophy, nor promote GAP 43 and Talpha1 tubulin mRNA expression, nor promote axonal regeneration into peripheral nerve transplants. TrkB receptor immunohistochemistry demonstrated immunoreactivity on the neuronal cell bodies, but not on anterogradely labeled rubrospinal axons at the injury site. These findings suggest that the poor response of rubrospinal neurons to BDNF applied to the spinal cord injury site 2 months after cervical axotomy is not related to the dose of BDNF administered, but rather to the loss of trkB receptors on the injured axons over time. Such obstacles to axonal regeneration will be important to identify in the development of therapeutic strategies for chronically injured individuals. PMID- 15296836 TI - Minocycline worsens hypoxic-ischemic brain injury in a neonatal mouse model. AB - Hypoxic-ischemic encephalopathy (HIE) is a leading cause of mortality and morbidity during the perinatal period, and currently no therapeutic drug is available. Minocycline, an antibiotic, has recently been shown to have neuroprotective effects distinct from its antimicrobial effect in several neurological disorders including ischemic brain injury. We examined the effect of minocycline on neonatal hypoxic-ischemic brain injury by using histologic scoring in both mouse and rat models. Mouse (C57Bl/6) and rat (SD) pups were exposed to a unilateral hypoxic-ischemic insult at 8 and 7 days of age, respectively. Minocycline hydrochloride was administered according to protocols that were reported to provide neuroprotection in adult or neonatal rats. Seven days after the insult, we examined brain injury in Nissl stained sections. Although minocycline ameliorated brain injury in the developing rat, it increased injury in the developing mouse. This detrimental effect in the mouse was consistent across different regions (cortex, striatum, and thalamus), with both single and multiple injection protocols and with both moderate and high-dose treatment (P < 0.05). The mechanism of the contrasting effects in mouse and rat is not clear and remains to be elucidated. Minocycline has been used as an antibiotic in the clinical setting for decades; therefore, it may be considered for use in infants with hypoxic-ischemic brain damage, based on prior reports of neuroprotection in the rat. However, it is important to examine this drug carefully before clinical use in human infants, taking our data in the mouse model into consideration. PMID- 15296837 TI - Valacyclovir treatment ameliorates the persistently increased pentylenetetrazol induced seizure susceptibility in mice with herpes simplex virus type 1 infection. AB - Herpes simplex virus type 1 (HSV-1) is an important pathogen related to epilepsy. We have shown previously that corneal inoculation of mice with HSV-1 causes acute spontaneous behavioral and electrophysiological seizures and increases hippocampal excitability and kainite-induced seizure susceptibility. In this study, we aimed to determine whether early-life HSV-1 infection in mice might cause short- and long-term enhanced susceptibility to pentylenetetrazol (PTZ) induced seizures and to evaluate whether early antiviral drug therapy was effectively ameliorating this deficit. Seizure threshold was calculated by the latency of onset of the myoclonic jerk, generalized clonus, and maximal tonic clonic convulsion. We demonstrate that the localization of viral antigens was predominantly within the bilateral temporal areas (amygdala, piriform, and entorhinal cortex) of HSV-1-infected mice. We also present evidence that mice of all HSV-1-infected groups had a shorter latency and higher severity to PTZ induced seizures than in age-matched, mock-infected controls. Treatment of HSV-1 infected mice with valacyclovir, a potent inhibitor of HSV-1 replication, produced a dose-dependent decrease in the signs of neurological deficits, pathological damages, and PTZ-induced seizure severity. Our results are consistent with the hypothesis that early-life HSV-1 infection leads to persistent enhancement of neuronal excitability in limbic circuits, which could result in an overall increased propensity to induce seizures later in life. Additionally, prompt optimal antiviral therapy effectively decreases seizure susceptibility in HSV-1-infected mice by limiting the level of viral replication and inflammatory response induced by virus. The present study provides not only experimental evidence, but also a new therapeutic strategy in HSV-1-associated human epilepsy. PMID- 15296838 TI - Sparing of behavior and basal extracellular dopamine after 6-hydroxydopamine lesions of the nigrostriatal pathway in rats exposed to a prelesion sensitizing regimen of amphetamine. AB - Repeated administration of amphetamine leads to enduring augmentation of its behavioral-activating effects, enhanced dopamine (DA) release in striatal regions, and morphological changes in DA target neurons. Here we show that exposure to a 2-week escalating-dose regimen of amphetamine prevents behavioral asymmetries of forelimb use and spontaneous (drug-independent) turning behavior following unilateral 6-hydroxydopamine (6-OHDA) lesions of the nigrostriatal pathway made 7-14 days after termination of amphetamine treatment (Experiments 1 3). Exposure to three nonescalating injections of amphetamine 7 days before 6 OHDA lesions had no effect (Experiment 2). Prelesion amphetamine treatment led to normalization of basal extracellular levels of striatal DA as measured by microdialysis on days 11-14 and 25-28 after lesioning (Experiment 3). However, there were no significant differences between treatment groups in postmortem tissue levels of DA and its metabolites, indicating a dissociation between the DA depletion and the extracellular levels of DA as measured by microdialysis. Finally, rats exposed to the escalating amphetamine regimen had reduced lesion induced loss of TH-IR cells in the ipsilateral DA cell body regions (Experiment 3). Thus, prelesion exposure to the escalating doses of amphetamine may render the cells resistant to the consequences of damage after subsequent 6-OHDA lesions, possibly by accelerating the development of compensatory changes in the DA neurons that typically accompany behavioral recovery. The potential role of amphetamine-induced endogenous neurotrophic factors in the behavioral sparing and normalization of basal extracellular DA levels observed after subsequent 6-OHDA lesions is discussed. PMID- 15296839 TI - Sex-related differences in the regeneration of sensory axons and recovery of nociception after peripheral nerve crush in the rat. AB - Sex-related differences regarding the regeneration of nociceptive axons and the recovery of nociception after sural nerve crush injury were examined in rats. The elongation rate of the fastest regenerating sensory axons in females started to increase after the first 6 days. This resulted in about 15% greater axon elongation distance at 8 days after crush in female than in male rats as determined by the nerve pinch test. The number of regenerating sensory axons in female and male rats, however, was not different. The recovery of nociception in the instep started earlier and was more extensive in females than in males during the entire 24-week recovery period, so that the pain sensitive area was finally about 20% larger in females than in males. Although ovariectomy significantly reduced plasma estradiol concentration in female rats, it did not change the elongation distance of regenerating nociceptive axons, which remained significantly greater than in male rats. Elimination of the cells in the distal nerve segment by freezing revealed that a more effective cell support in the distal nerve segment is probably responsible for faster regeneration of nociceptive axons in females than in males, rather than the circulating female sex hormones. PMID- 15296840 TI - A comparison of thyroxine- and polyamine-mediated enhancement of rat facial nerve regeneration. AB - Thyroid hormones and spermidine, a motor neuron trophic polyamine (PA), have been shown to enhance peripheral motor nerve regeneration; however, the mechanism by which these treatment modalities exert their effect is unknown. Similarities in treatment outcome suggest that these molecules may be working via a common mechanism. Such an explanation is plausible since thyroid hormone is a potent inducer of ornithine decarboxylase (ODC), which is the rate-limiting enzyme involved in polyamine synthesis. This study was designed to morphologically evaluate the effects of exogenous thyroxine and spermidine on the regeneration of the rat facial nerve. Myelinated fiber density, axonal size, and degree of myelination were assayed by light and electron microscopy 21 days following facial nerve crush. Strikingly, the two treatment modalities had identical effects on all parameters tested. Each significantly enhanced the density of myelinated axons in regenerating nerves relative to the vehicle control. In addition, relative to the control treatment, both thyroxine and spermidine significantly increased the cross-sectional area of regenerating axons (P < 0.05). Interestingly, neither of the drug treatments had any effect on remyelination at the position where this parameter was analyzed. The concurrent administration of both thyroxine and spermidine did not synergistically enhance motor neuron regeneration. These data support the hypothesis that thyroxine and spermidine enhance neural regeneration by a common mechanism. PMID- 15296841 TI - Myosin II activity is required for severing-induced axon retraction in vitro. AB - Understanding the mechanistic basis of the response of neurons to injury is directly relevant to the development of effective therapeutic approaches aimed at the amelioration of nervous system damage. Axons retract in response to severing. We investigated the mechanism of axon retraction in response to severing in vitro, testing the hypothesis that actomyosin contractility drives severing induced axon retraction. Axon retraction commenced within 5 min following severing and correlated with actin filament accumulation at the site of severing. Depolymerization of actin filaments prevented retraction, demonstrating that actin filaments are required for severing-induced axon retraction. Direct inhibition of myosin II, using blebbistatin, minimized axon retraction in response to severing. Blocking RhoA-kinase (ROCK), a modulator of myosin II activity, inhibited axon retraction. Similarly, inhibiting myosin light chain kinase (MLCK) with a cell-permeable pseudo-substrate peptide also inhibited axon retraction. These data demonstrate that myosin II activity is required for severing-induced axon retraction in vitro, and suggest myosin II as a target for therapeutic interventions aimed at minimizing retraction following severing in vivo. PMID- 15296842 TI - Agmatine reduces infarct area in a mouse model of transient focal cerebral ischemia and protects cultured neurons from ischemia-like injury. AB - Agmatine is a primary amine formed by the decarboxylation of L-arginine synthesized in mammalian brain. In this study, we investigated the neuroprotective effect of agmatine on ischemic and ischemia-like insults. Primary cortical neuronal cultures were subjected to oxygen-glucose deprivation (OGD), a model of ischemia-like injury, and treated with agmatine before or at the start of OGD, or upon reperfusion. Neuronal death was reduced when agmatine was present during OGD, and this protection was associated with a reduction of nitric oxide (NO) and neuronal nitric oxide synthase (nNOS), but not inducible NOS (iNOS). Protection by agmatine was also studied at the in vivo level using a model of middle cerebral artery occlusion (MCAO) in mice. Mice were subjected to 2 h MCAO. Agmatine was administered either 30 min before ischemia, at the start of MCAO, at the start of reperfusion, or 2 or 5 h into reperfusion. Agmatine markedly reduced infarct area in all treatment groups except when treatment was delayed 5 h. The number of nNOS immunopositive cells was correlated with neuroprotection. Interestingly, immunoreactivity for iNOS was reduced only when agmatine was administered before and at the onset of MCAO. Our study suggests that agmatine may be a novel therapeutic strategy to reduce cerebral ischemic injury, and may act by inhibiting the detrimental effects of nNOS. PMID- 15296843 TI - The acetylcholine fiber density of the neocortex is altered by isolated rearing and early methamphetamine intoxication in rodents. AB - Alterations in the cholinergic physiology of the brain were the first to be observed when research on environmental influences on postnatal brain development began 35 years ago. Since then, the effects of isolated rearing (IR) or early pharmacological insults have been shown not only on the physiology, but also the anatomy of a variety of transmitter systems. The cholinergic fiber density, however, still remained to be assessed. We therefore used a histochemical procedure to stain cholinergic fibers in the brains of young adult gerbils reared either in groups in enriched environments or isolated in standard makrolon cages. Half of the animals from each rearing condition had received a single high dose of methamphetamine on postnatal day 14. Fiber densities were measured by computerized image analysis in the medial and orbital prefrontal cortex (PFC), dysgranular and granular insular cortex, sensorimotor cortices, and the entorhinal cortex of both hemispheres. Isolation rearing increased the cholinergic fiber densities in the prefrontal cortices of the left hemisphere and in the entorhinal cortex of the right hemisphere by about 10%, with no effect in the respective contralateral side. The early methamphetamine intoxication showed no influence in prefrontal and entorhinal cortices, but diminished the acetylcholine (ACh) innervation of the forelimb area of cortex in both hemispheres in IR gerbils and of the left hemisphere in ER gerbils, and reduced the acetylcholine innervation in the hindlimb area in both sides in both rearing groups. These results demonstrate that (a) cholinergic fiber density is differentially regulated in different cortical areas and (b) the plasticity of the cholinergic system can only be understood in the interplay with other neuromodulatory innervations. PMID- 15296844 TI - Neonatal hypoxia suppresses oligodendrocyte Nogo-A and increases axonal sprouting in a rodent model for human prematurity. AB - Premature human infants frequently suffer from periventricular leukomalacia (PVL) characterized by the loss of central myelinated tracts in the brain [Neuropathology, 22 (2002) 193]. Rodent chronic sublethal hypoxia (CSH) from P3 to 33 (postnatal day 3-33) provides a model for PVL characterized by cerebral ventriculomegaly and reductions in cerebral white matter volume [Brain Res. Dev. Brain Res. 111 (1998) 197; Proc. Natl. Acad. Sci. USA 100 (2003) 11718]. Here, we demonstrate that mice exposed to CSH from P3 to P33 followed by normoxia from P33 to P75 continue to exhibit a locomotor hyperactivity that resembles behavioral changes observed in some human children with very low birth weights. Because periventricular white matter is specifically lost in PVL, we examined the expression of oligodendrocyte proteins. Hypoxic rearing dramatically decreases the level of the axon outgrowth inhibitor Nogo-A in oligodendrocytes of CNS white matter at P12. The Nogo-A decrease exceeds the moderate decrease in another myelin protein, myelin associated glycoprotein (MAG). Although myelin protein expression returns to normal by maturity (P75), persistent abnormalities in axonal trajectories are detectable. Anterograde axonal tracing from motor cortex demonstrates ectopic corticofugal fibers in the corticospinal tract (CST), corpus callosum, and caudate nucleus of adult animals reared in CSH. Thus, hypoxia induced reduction in myelin-derived axon outgrowth inhibitors appears to contribute axonal misconnection to the pathology of very low birth weight infants. PMID- 15296845 TI - Transient exposure of rat pups to hyperoxia at normobaric and hyperbaric pressures does not cause retinopathy of prematurity. AB - We have shown that hyperoxia reduces brain damage in a rat model of hypoxia ischemia. The purpose of this study was to examine the possibility of hyperoxia in inducing vision-threatening retinopathy. Two different experiments were conducted in this study. PART 1: seven-day-old rat pups were subjected to unilateral carotid artery ligation followed by 2 h of hypoxia (8% O2 at 37 degrees C). Pups were treated with 100% oxygen at 1 ATA, 1.5 ATA, and 3.0 ATA for a duration of 1 h. PART 2: Newborn rat pups were exposed to 100% oxygen at 1, 1.5, or 3.0 ATA for 1 h, the same treatment protocol used for brain protection after hypoxia-ischemia. Retinopathy was evaluated by the degree of neovascularization (measuring retinal vascular density), by the structural abnormalities (histology) in the retina, and by the expression of hypoxia hyperoxia sensitive proteins including hypoxia-inducible factor-1alpha (HIF 1alpha) and vascular endothelial growth factor (VEGF) at 24 h, 1, 2, and 10 weeks after hyperoxia exposure. Hyperoxic treatment at all pressures administered significantly reduced the hypoxia-ischemic-induced reduction in brain weight. Retinal vascular density measurements revealed no signs of neovascularization after hyperoxia exposure. There were also no abnormalities in the structure of the retina and no changes in the protein expression of HIF-1alpha and VEGF following hyperoxia exposure. Exposure to hyperoxia for 1 h at normobaric or hyperbaric pressures did not result in the structural changes or abnormal vascularization that is associated with retinopathy of prematurity, suggesting that hyperoxia is a safe treatment for hypoxic newborn infants. PMID- 15296846 TI - Cognitive and neurological deficits induced by early and prolonged basal forebrain cholinergic hypofunction in rats. AB - In the present study we examined the long-term effects of neonatal lesion of basal forebrain cholinergic neurons induced by intracerebroventricular injections of the immunotoxin 192 IgG saporin. Animals were then characterised behaviourally, electrophysiologically and molecularly. Cognitive effects were evaluated in the social transmission of food preferences, a non-spatial associative memory task. Electrophysiological effects were assessed by recording of cortical electroencephalographic (EEG) patterns. In addition, we measured the levels of proteins whose abnormal expression has been associated with neurodegeneration such as amyloid precursor protein (APP), presenilin 1 and 2 (PS 1, PS-2), and cyclooxygenases (COX-1 and COX-2). In animals lesioned on postnatal day 7 and tested 6 months thereafter, memory impairment in the social transmission of food preferences was evident, as well as a significant reduction of choline acetyltransferase activity in hippocampus and neocortex. Furthermore, similar to what observed in Alzheimer-like dementia, EEG cortical patterns in lesioned rats presented changes in alpha, beta and delta activities. Levels of APP protein and mRNA were not affected by the treatment. Levels of hippocampal COX-2 protein and mRNA were significantly decreased whereas COX-1 remained unaltered. PS-1 and PS-2 transcripts were reduced in hippocampus and neocortex. These findings indicate that neonatal and permanent basal forebrain cholinergic hypofunction is sufficient to induce behavioural and neuropathological abnormalities. This animal model could represent a valid tool to evaluate the role played by abnormal cholinergic maturation in later vulnerability to neuropathological processes associated with cognitive decline and, possibly, to Alzheimer-like dementia. PMID- 15296847 TI - Repeated episodic exposure to ethanol affects neurotrophin content in the forebrain of the mature rat. AB - Chronic exposure to ethanol can cause deficits in learning and memory. It has been suggested that withdrawal is potentially more damaging than the ethanol exposure per se. Therefore, we explored the effect of repeated episodic exposure to ethanol on key regulators of cortical activity, the neurotrophins. Rats were exposed to ethanol via a liquid diet for 3 days per week for 6-24 weeks. Control rats were pair-fed an isocaloric liquid diet or ad libitum fed chow and water. The concentrations of nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and neurotrophin-3 (NT-3) were determined using enzyme-linked immunosorbant assays (ELISAs). Five telencephalic structures were examined: parietal cortex, entorhinal cortex, hippocampus, the basal nucleus, and the septal nuclei. All five areas expressed each of the three neurotrophins; BDNF was most abundant and NGF the least. The parietal cortex was susceptible to ethanol exposure, NGF and BDNF content increased, and NT-3 content fell, whereas no changes were detectable in the entorhinal cortex. In the hippocampus, the amount all three neurotrophins increased following episodic ethanol exposure. Neurotrophin content in the two segments of the basal forebrain was affected; NGF and NT-3 content in the basal forebrain was reduced and NGF and BDNF content in the septal nuclei was increased by ethanol exposure. In many cases where ethanol had an effect, the change was transient so that by 24 weeks of episodic exposure, no significant changes were evident. Thus, the effects of ethanol are site- and time-dependent. This pattern differs from changes caused by chronic ethanol exposure, hence, neurotrophins must be vulnerable to the effects of withdrawal. Furthermore, the ethanol-induced changes do not appear to fit a model consistent with retrograde regulation, rather they suggest that neurotrophins act through autocrine/paracrine systems. PMID- 15296848 TI - Blockade of adenosine A2A receptors antagonizes parkinsonian tremor in the rat tacrine model by an action on specific striatal regions. AB - Acute administration of the acetylcholinesterase inhibitor tacrine to rats induces tremulous jaw movements which can be used as a valuable model of parkinsonian tremor. In the present study, the number of tremor episodes and jaw movements were evaluated to assess the effects of the selective A2A antagonists SCH 58261 and SCH BT2 on tremorgenesis. SCH 58261 dose-dependently, and maximally at 5 mg/kg, reduced the number of both tremor episodes (-35%) and jaw movements ( 50%), induced in rats by tacrine (2.5 mg/kg ip). Since adenosine A2A receptors are largely expressed throughout the striatum, chronic cannulae were implanted in the rat dorsomedial (DMS) and ventrolateral striatum (VLS) to investigate whether A2A antagonists could act at this level. Infusion of SCH BT2 (5 microg/microl), a water-soluble analogue of SCH 58261, in VLS antagonized both tremor episodes ( 68%) and jaw movements (-76%) elicited by tacrine (2.5 mg/kg ip), whereas SCH BT2 infusion in DMS was less effective in blocking jaw movements (-50%) and did not significantly affect the number of tremor episodes. Taken together, the results of this study indicate that A2A antagonists effectively reduce the magnitude of tremulous jaw movements induced in rats by acute tacrine, mainly by an action in VLS and suggest that A2A antagonists might be used as specific agents against parkinsonian tremor. PMID- 15296849 TI - Electro-acupuncture stimulation protects dopaminergic neurons from inflammation mediated damage in medial forebrain bundle-transected rats. AB - Through producing a variety of cytotoxic factors upon activation, microglia are believed to participate in the mediation of neurodegeneration. Intervention against microglial activation may therefore exert a neuroprotective effect. Our previous study has shown that the electro-acupuncture (EA) stimulation at 100 Hz can protect axotomized dopaminergic neurons from degeneration. To explore the underlying mechanism, the effects of 100 Hz EA stimulation on medial forebrain bundle (MFB) axotomy-induced microglial activation were investigated. Complement receptor 3 (CR3) immunohistochemical staining revealed that 24 sessions of 100 Hz EA stimulation (28 days after MFB transection) significantly inhibited the activation of microglia in the substantia nigra pars compacta (SNpc) induced by MFB transection. Moreover, 100 Hz EA stimulation obviously inhibited the upregulation of the levels of tumor necrosis factor (TNF)-alpha and interleukin (IL)-1beta mRNA in the ventral midbrains in MFB-transected rats, as revealed by reverse transcriptase polymerase chain reaction (RT-PCR). ED1 immunohistochemical staining showed that a large number of macrophages appeared in the substantia nigra (SN) 14 days after MFB transection. The number of macrophages decreased by 47% in the rats that received 12 sessions of EA simulation after MFB transection. These data indicate that the neuroprotective role of 100 Hz EA stimulation on dopaminergic neurons in MFB-transected rats is likely to be mediated by suppressing axotomy-induced inflammatory responses. Taken together with our previous results, this study suggests that the neuroprotective effect of EA on the dopaminergic neurons may stem from the collaboration of its anti-inflammatory and neurotrophic actions. PMID- 15296850 TI - A new model of ischemic preconditioning using young adult hippocampal slice cultures. AB - In ischemic preconditioning (IPC), brief sublethal ischemia protects neurons from a subsequent lethal ischemia. In vivo models faithfully display preconditioning, yet, these models are technically challenging, time-consuming and expensive. In vitro models of preconditioning have also been developed that are technically easier and less expensive. A drawback of pre-existing in vitro models is that since susceptibility to ischemic injury is age-dependent; neuroprotection is being studied in neurons that have intrinsic resistance to oxygen-glucose deprivation (OGD). This study introduces a new in vitro model of ischemic preconditioning in hippocampal slice cultures isolated from 20-30-day-old rats. Slice cultures show a high susceptibility and sharp thresholds toward ischemia that is comparable to that found in vivo. A 5-min OGD treatment was not neurotoxic to young adult slice cultures, while a 10-min OGD treatment was neurotoxic. In addition, the sublethal 5-min OGD treatment protected against a 10 min OGD treatment that was delivered 24 h later. Neuroprotection was seen in preconditioned slice cultures stained with propidium iodide (PI) or with antisera against the neuron-specific antigen NeuN. Energy failure is hypothesized to trigger ischemic preconditioning and a 5-min OGD treatment induced transient energy failure in young adult slice cultures. This model may assist in the search for new therapeutics for the prevention and/or treatment of stroke. PMID- 15296851 TI - Quantification of sPLA2-induced early and late apoptosis changes in neuronal cell cultures using combined TUNEL and DAPI staining. AB - The terminal deoxynucleotidyl transferase (TdT) dUTP nick end labeling (TUNEL) stain is in wide use for measuring apoptosis in neurons, as well as in other cell types. TUNEL may give false positive results due to variations in labeling technique as well as staining of cells that have undergone non-apoptotic DNA strand breaks. Therefore, in isolation, TUNEL is not a certain indicator of apoptosis. Recently, we have demonstrated the potent apoptotic effect of secreted phospholipase A2 from group III (sPLA2-III) on primary cortical neurons from rat. Here we describe a computer-assisted method for quantifying TUNEL-positive neurons after sPLA2-III induced apoptosis. Extent of TUNEL is normalized to total nuclear content using 4',6-diamidino-2-phenylindole (DAPI) staining. Furthermore, DAPI counterstaining allows for determination of a nuclear morphology indicator, based on nuclear size and roundness, which we call the nuclear area factor. We found that the nuclear area factor is an early indicator of cell death (significant after 4 h post treatment), while TUNEL staining is significant at later times (26 h). Thus, the independent staining techniques using TUNEL and DAPI complement each other, and with commercially available image analysis software, may be used to indicate early as well as delayed cell injury processes. PMID- 15296852 TI - Grooming analysis algorithm for neurobehavioural stress research. AB - Since rodent self-grooming behaviours are elicited by both comfort and stressful conditions, traditional measures such as duration, latency of onset and the number of bouts may be not suitable to dissociate between these opposite conditions. The aim of the current study was to improve and optimize ethological measurement of self-grooming in neurobehavioural stress research enabling differentiation between stress and no-stress situations. This protocol assists in the correct interpretation of animal grooming behaviours and detection of stress by measuring alterations in grooming microstructure in different test situations. While a general pattern of self-grooming uninterrupted cephalocaudal progression is normally observed in no-stress (comfort) conditions in mice and other rodents, the percentage of "incorrect" transitions between different stages and the percentage of interrupted grooming bouts may be used as behavioural marker of stress. The protocol can be a useful tool in neurobehavioural stress research including modelling stress-evoked states, pharmacological screening of potential antistress drugs or behavioural phenotyping of genetically modified animals. PMID- 15296853 TI - A new method for the assessment of spatial orientation and spatial anxiety in mice. AB - The implication of integrated functional sensory relations of the body to space in anxiety disorders is a very important issue which encourages the development of animal models, in particular, for pharmacological perspectives and for the functional assessment of the deficits induced by genetic manipulation in the mouse or the rat. A new experimental device is presented here: It is comprised of a rotating tunnel and a rotating-beam controlled by computer which can be used for multiple visuo-idiothetic and kinesthetic sensory conflict situations during active locomotor behaviour by mice. The system is linked to a digital video system, Video-Track trade mark, designed to track and record the movements of the animals. Anxious BALB/cByJ mice were compared to non-anxious C57BL/6J mice and were seen to display highly disturbed locomotor behaviour in a sensory conflict situation. The model highlights the advantages of video-digital analysis for animal behavioural sciences. PMID- 15296854 TI - In vivo tracking of bone marrow stromal cells transplanted into mice cerebral infarct by fluorescence optical imaging. AB - Recent experimental studies have indicated that bone marrow stromal cells (BMSC) improve neurological deficits when transplanted into the animal models of various neurological disorders, although precise mechanism still remains unclear. In this study, we developed a new in vivo fluorescence optical imaging protocol to sequentially track the transplanted into the brain of the living animals subjected to cerebral infarct. Mice BMSC were harvested from transgenic mice expressing green fluorescent protein (BMSC-GFP). They were stereotactically transplanted into the ipsilateral striatum of mice subjected to permanent middle cerebral artery occlusion after 7 days of ischemia (n=12). During 12 weeks after transplantation, the skull was exposed and the green fluorescence emitted from the brain surface was sequentially observed, using in vivo fluorescence optical microscopy. As the results, regional green fluorescence was detected in the ipsilateral parietal region 4-12 weeks after transplantation in all animals and became more apparent over the time. The images obtained through the skull were very similar to those acquired by thinning or removing the skull. Immunohistochemistry evaluation revealed that the transplanted cells migrated towards the ischemic boundary zone and expressed the neuronal or astrocytic marker, supporting the findings on fluorescence optical images. Sequential visualization of the BMSC transplanted into the brain of living animals would be valuable for monitoring the migration, growth and differentiation of the transplanted cells to explore the fate and safety of stem cell transplantation for various neurological disorders. PMID- 15296855 TI - A fluorescence-based technique for screening compounds that protect against damage to brain mitochondria. AB - Mitochondrial failure to generate ATP can be due to damage to their membranes, which leads to release of solutes, e.g., pyridine nucleotides, from the mitochondrial matrix. We developed a highly sensitive fluorescence assay for detecting a pathologic increase in mitochondrial membrane permeability. The assay is based on coupled enzymatic reactions that produce hydrogen peroxide in the presence of the reduced or oxidized form of nicotinamide adenine dinucleotide (NADH/NAD). The hydrogen peroxide is a substrate for horseradish peroxidase that converts Amplex Red into highly fluorescent Resorufin. The assay is able to detect nanomolar levels of pyridine nucleotides in the medium. Calcium additions to isolated rat brain or liver mitochondria incubated in a potassium-based medium with added enzymes caused osmotic swelling, as detected with light scattering, and production of Resorufin, due to release of NADH/NAD. These events were blocked by cyclosporin A (CsA) or Bongkrekic acid (BKA), inhibitors of the mitochondrial permeability transition (MPT). These results indicate that the NADH/NAD release assay is a simple, reliable, and sensitive method for detecting mitochondrial damage and for screening of compounds that protect mitochondria from injury. PMID- 15296856 TI - Long-lived retrograde fluorescent labeling of corticospinal neurons in the living animal. AB - For pathophysiological studies, it is advantageous to label specific neuronal populations in living animals. This study aimed to establish a method for stable and long-lasting fluorescent labeling of corticospinal neurons in the living animal. The two fluorescent dyes Fluoro-Red and Fluoro-Green were injected in the cervical spinal cord of anesthetized newborn rats. After a recovery period, treated rats were returned to the mother. After 24 h and 14 days, fixed brain sections revealed wide-spread fluorescence in elongated or pyramidal-shaped cell profiles in a discrete internal cortical layer, consistent with layer V pyramidal cells. Labeled neurons displayed spontaneous synaptic activity using the slice patch clamp method. These results suggest that these dyes are effective tools for pathophysiological and slice patch clamp studies focused on specific neuron groups. PMID- 15296857 TI - Krypton laser-induced photothrombotic distal middle cerebral artery occlusion without craniectomy in mice. AB - Recent advances in genetical engineering of the mouse have highlighted the importance of reproducible and less invasive models of cerebral ischemia in mice. In this paper, we developed minimally invasive and reproducible model of distal middle cerebral artery (MCA) occlusion in mice using krypton (Kr) laser-induced photothrombosis. C57BL/6 or BALB mice (n=8 each) were anesthetized with halothane. The skin was cut, the temporal muscle was retracted, and the right distal MCA was observed through the skull. A Kr laser beam of wavelength 568 nm was focused onto the MCA over the intact skull. Upon laser irradiation, intravenous administration of a rose bengal solution was begun. After 4 min of irradiation, the laser beam was refocused on the MCA just proximal to the first spot, and another 4-min irradiation was performed. Then, the right common carotid artery (CCA) was ligated. Three days later, the brain was removed, and infarct volume was determined. Infarction confined almost solely to the cortical area was produced in each mouse. Mean infarct volume in C57BL/6 mice was 25.2+/-13.7 mm3. The BALB mice group showed significantly larger and more reproducible infarction (44.1+/-5.2 mm3; the coefficient of variation was 12%) than did C57BL/6 mice (P<0.005). Our photothrombosis model of stroke in mice can be performed without craniectomy, and its reproducibility is satisfactory when using BALB mice. PMID- 15296858 TI - The emergence of HIV resistance and new antiretrovirals: are we winning? PMID- 15296860 TI - Antifungal pharmacokinetics and pharmacodynamics: understanding the implications for antifungal drug resistance. AB - Pharmacodynamics (PDs) describe the relationship between drug exposure and outcome. The drug exposures in these analyses are most commonly expressed in a variety of pharmacokinetic terms. The outcome of interest with anti-infective therapy is either microbiologic resolution or a clinical surrogate of treatment efficacy. An in vitro measure of drug potency, such as the minimum inhibitory concentration (MIC) is also frequently considered in this relationship. Examination of the relationships among drug pharmacokinetics, MIC, and efficacy has provided a framework for choice of antifungal drug and dose. These analyses provide a PD target for drug class/organism combinations. The PD target can be useful for defining the upper MIC limit for a drug-dosing regimen that would be expected to result in treatment efficacy. The PD target can be used to optimize dosing regimens and to aid in defining susceptibility breakpoints. PMID- 15296859 TI - Multifaceted roles of cyclooxygenase-2 in lung cancer. AB - Lung cancer is the leading cause of cancer death in the United States. Although the low 5-year survival rate (under 15%) has changed minimally in the last 25 years, new agents and combinations of agents that target tumor proliferation, invasion, and survival may lead to improvement in patient outcomes. There is evidence that cyclooxygenase-2 (COX-2) is overexpressed in lung cancer and promotes tumor proliferation, invasion, angiogenesis, and resistance to apoptosis. COX-2 inhibitors have been found to inhibit tumor growth in animal models and have demonstrated responses when combined with conventional therapy in phase II clinical trials. Further understanding of the mechanisms involved in COX 2-mediated tumorigenesis and its interaction with other molecules in lung cancer may lead to improved therapeutic strategies for this disease. In addition, delineation of how COX-2-dependent genes modulate the malignant phenotype will provide novel insights in lung cancer pathogenesis. PMID- 15296861 TI - Chemically modified tetracyclines as inhibitors of matrix metalloproteinases. AB - Matrix metalloproteinases belong to a diverse group of enzymes that are not only involved in restructuring the extracellular matrix, but also play a major role in various pathophysiological conditions by virtue of their complicated expression, activation, and regulation processes. They have been widely implicated to function as major contenders in cancer progression, frequently due to their role in invasion, proliferation and metastasis. MMP inhibitors have been specifically designed to target these altered activities of MMPs, mostly by means of inhibiting their function and by diminishing their increased expression in various disease states, particularly cancer. Tetracyclines and chemically modified tetracyclines (CMTs) have been rationally designed to inhibit the activity of MMPs and thus decrease the potential risk of spread of tumor cells to distant sites by invasion and metastasis. Pre-clinical and early clinical data for one of these CMTs, COL-3 (formerly CMT-3) indicate considerable potential for this group of anticancer agents. Further testing and rational modifications of these CMT analogues might lead to new anticancer agents. PMID- 15296863 TI - Molecular ecotoxicology of plants. AB - Plant molecular exotoxicology investigates ecological implications of genetic and molecular responses to toxins, herbicides, pollutants and natural stress factors. Plant fitness is analysed by examining the relationships between plant genotype and ecological phenotype, enabling regulatory networks formed by second messenger molecules and transcriptional as well as post-transcriptional events to be elucidated. This general approach is illustrated here by specific case studies: detoxification by glucosyl transfer or binding to cell wall macromolecules; roles of the multifunctional formaldehyde dehydrogenase; and abiotic induction of plant immunity through reactive oxygen species. As a practical application of molecular ecotoxicology, the interaction of commercialized transgenic crop plants with potential environmental selection factors is discussed. PMID- 15296862 TI - Customizing chemotherapy for colon cancer: the potential of gene expression profiling. AB - The value of gene expression profiling, or microarray analysis, for the classification and prognosis of multiple forms of cancer is now clearly established. For colon cancer, expression profiling can readily discriminate between normal and tumor tissue, and to some extent between tumors of different histopathological stage and prognosis. While a definitive in vivo study demonstrating the potential of this methodology for predicting response to chemotherapy is presently lacking, the ability of microarrays to distinguish other subtleties of colon cancer phenotype, as well as recent in vitro proof-of principle experiments utilizing colon cancer cell lines, illustrate the potential of this methodology for predicting the probability of response to specific chemotherapeutic agents. This review discusses some of the recent advances in the use of microarray analysis for understanding and distinguishing colon cancer subtypes, and attempts to identify challenges that need to be overcome in order to achieve the goal of using gene expression profiling for customizing chemotherapy in colon cancer. PMID- 15296864 TI - Tumors of pet birds. AB - Neoplastic diseases are becoming more than a postmortem diagnosis due to the increasing knowledge base and improving quality of avian medicine. The expectation for better health care demands a diagnosis, prognosis, and therapy options. However, the published information regarding prognosis and therapy of specific neoplasms remains limited in avian medicine. With each case report or study that provides this information, there is an improvement in the level of care we can offer our companion avian species. This review will cover some basic information about specific tumor types and will reference the more recent reports in the avian literature. It is not intended to be all encompassing. PMID- 15296865 TI - Spontaneous neoplasms of lagomorphs. AB - Many of the same neoplastic processes reported in companion animal medicine have been documented in lagomorphs. With only a few reports in the literature regarding rabbits and hares, the behavior of many of these tumors cannot be predicted. Further, based on limited current knowledge, the practitioner is left to make decisions for diagnostics and therapeutics in these cases based upon current recommendations for companion animals with adjustments for lagomorph physiology. We hope that this review of lagomorph neoplasia will inform the general practitioner of the occurrence of these tumors, promote further documentation and research of these processes in rabbits, and encourage the practitioner of rabbit medicine along with rabbit owners to consult with veterinary oncologists regarding neoplastic processes in rabbits rather than just giving a guarded prognosis. PMID- 15296866 TI - Ferret oncology: diseases, diagnostics, and therapeutics. AB - Many standard diagnostic and chemotherapeutic protocols can be adapted for use in ferrets. Unique anatomic and clinical features dictate modification of protocols, but should not prohibit diagnosis or treatment. Ferrets may be the easiest of nontraditional species to treat with chemotherapeutics. We can provide more options for our patients, with improved quality of life and longer survival times than ever before. Although clients are never happy to hear the diagnosis of "cancer," it is no longer a word that condemns their beloved pet. PMID- 15296867 TI - Spontaneous tumors of small mammals. AB - The most common tumor of guinea pigs is bronchogenic papillary adenoma; of hedgehogs is mammary gland adenocarcinoma; of hamsters is adrenal cortical adenoma; of gerbils is ovarian granulosa cell and theca cell tumors; of mice is pulmonary carcinoma; and of rats is mammary fibroadenoma. A relatively low incidence of tumors is described for chinchillas and hamsters, whereas the incidence of tumors is high for gerbils, hedgehogs, mice, and rats. Limited literature regarding neoplasia exists for prairie dogs, sugar gliders, and chinchillas. PMID- 15296868 TI - Reptile neoplasia: a retrospective study of case submissions to a specialty diagnostic service. AB - This retrospective study appears to be the largest publication to date regarding the prevalence of neoplasia in reptiles. As in previous publications, neoplasia is most common in snakes, followed by lizards, chelonians, and crocodilians. Several interesting trends were documented in this study, some of which appear to be previously unidentified, and some that support the findings of previous publications. PMID- 15296869 TI - Amphibian oncology. AB - Spontaneous neoplasia is rare in all three orders of Amphibia. Tumors are documented in most major organ systems, and some have various underlying etiologies, including viral infection, environmental contaminants, and genetic predisposition. Currently,treatment options are limited to removal of the predisposing condition(s), palliative care, surgical excision, and, when necessary,humane euthanasia may be elected. Neoplasia must be distinguished from common infectious, nonneoplastic conditions that can negatively impact population health. This article is a review of the more common types of neoplasia in amphibians, and includes clinically relevant information, such as biologic behavior,anatomy, associated etiologies, major differential diagnoses, and clinical management. PMID- 15296870 TI - Oncology of invertebrates. AB - In this article, "Invertebrates" are considered to be all those animals that are not in the five main groups of vertebrates (Mammalia,Aves, Reptilia, Amphibia, and three classes of fish), and the term neoplasm is used in its traditional sense of "new, uncontrolled, growth". The history of study of such lesions is reviewed and methods of investigation and control are outlined. PMID- 15296871 TI - Neoplasia in fishes. AB - Similar to higher vertebrates, neoplasia is not an uncommon disease in fishes, which are the largest group of vertebrates. However,neoplasia in fishes is generally a benign condition with relatively few exceptions of malignant disease. The objective of this discussion is to provide an overview of neoplasia and the various neoplastic disease conditions in fishes according to organ system,including the few neoplasms of species that are familiar to the aquatic animal or exotic animal practitioner. The discussion also considers the various nonneoplastic lesions in fishes that may be confused with neoplasms, and treatment of neoplastic disease in fishes that is generally restricted to surgical intervention. PMID- 15296872 TI - Current therapies in exotic animal oncology. AB - The majority of information on oncology therapies has been reported in humans, canine, and feline patients, and laboratory animals with experimentally induced tumors. A variety of treatments,including radiation therapy, chemotherapy, photodynamic therapy, and others have been used with exotic animals. There are many species of exotic pets, and anatomic differences, as well as husbandry and nutritional requirements, must be taken into account to provide optimal care. By providing a broad overview of therapies and considerations for treatment, this article is intended to provide the practitioner with an overview of approach and options when addressing oncology cases in exotic animals. PMID- 15296873 TI - Surgical oncology of exotic animals. AB - This article reviews the initial management of exotic patients with amass. The important principles of the initial workup, including how to get a definitive diagnosis and how to stage the extent of the disease are covered. There are many biopsy instruments currently available, and the advantages and disadvantages of these are presented. Principles of tumor excision and marking the tissue sampled are also discussed. It is important to submit all of the tissue excised and evaluate the surgical margins for the presence of neoplastic cells, even if the tumor seems too small to put in the sample vial. Follow-up for patients that have had cancer surgery is also reviewed, along with the effects of adjunct therapies on wound healing. PMID- 15296874 TI - The use of chemotherapy in exotic animals. AB - Although there is little published data on successful treatment of neoplasia with chemotherapy drugs in exotic animal species, there has not been adequate research done, or even large case series on how these drugs should be dosed and which drugs are most appropriate to use for which tumor types. As people demand better treatment for their pets, hopefully more information will become available, and the reader is advised to keep track of the literature for new information as it becomes available. PMID- 15296875 TI - Current research in avian chemotherapy. AB - Chemotherapy as a treatment modality is increasingly being used in avian oncology. Currently, most chemotherapeutic agents can only be used empirically, as pharmacokinetic data in birds are lacking.Recently, the pharmacokinetic profile of the platinum analogs,cisplatin, and carboplatin has been reported in Sulfur-crested cockatoos,paving the way for clinical and toxicity trials. PMID- 15296877 TI - The cardiothoracic surgeon and the basic scientist. PMID- 15296878 TI - A new computer model of mitral valve hemodynamics during ventricular filling. AB - OBJECTIVE: Quantitative description of left ventricular diastolic filling and mitral valve function remains difficult despite advances in echocardiography. The purpose of the present study was to develop a lumped parameter model of left ventricular filling and validate it in porcine trials under physiological conditions and after valve replacement. METHODS: Six animals were instrumented with aortic flow meter, left atrial pressure catheter and combined left ventricular pressure-conductance catheter. The model simulates ventricular and arterial pressures and flows during diastolic filling. Input parameters include maximum mitral valve area, blood viscosity and density, atrial compliance, left ventricular active relaxation characteristics and initial pressure and flow values. The outputs of the model are atrial and ventricular pressure as well as transmitral flow as a function of time. The model primarily consists of a system of four first-order, non-linear ordinary differential equations which were solved with MATLAB software. RESULTS: Left atrial and ventricular pressure data and model flow curves were nearly identical under baseline conditions, during rapid preload reduction by vena caval occlusion and after prosthetic valve replacement. Measured and model based calculation of early diastolic filling volume (E-wave), showed an excellent correlation under all three conditions (r = 0.998, P < 0.0001; r = 0.997, P < 0.0001; r = 0.974, P < 0.0001, respectively) with a mean difference less then two percent. CONCLUSION: The new lumped parameter model of left ventricular filling allows for the first time a detailed simulation of pressure and flow curves in the left heart including transmitral hemodynamics. PMID- 15296879 TI - Computational simulation of intracoronary flow based on real coronary geometry. AB - OBJECTIVE: To assess the feasibility of computationally simulating intracoronary blood flow based on real coronary artery geometry and to graphically depict various mechanical characteristics of this flow. METHODS: Explanted fresh pig hearts were fixed using a continuous perfusion of 4% formaldehyde at physiological pressures. Omnipaque dye added to lead rubber solution was titrated to an optimum proportion of 1:25, to cast the coronary arterial tree. The heart was stabilized in a phantom model so as to suspend the base and the apex without causing external deformation. High resolution computerized tomography scans of this model were utilized to reconstruct the three-dimensional coronary artery geometry, which in turn was used to generate several volumetric tetrahedral meshes of sufficient density needed for numerical accuracy. The transient equations of momentum and mass conservation were numerically solved by employing methods of computational fluid dynamics under realistic pulsatile inflow boundary conditions. RESULTS: The simulations have yielded graphic distributions of intracoronary flow stream lines, static pressure drop, wall shear stress, bifurcation mass flow ratios and velocity profiles. The variability of these quantities within the cardiac cycle has been investigated at a temporal resolution of 1/100th of a second and a spatial resolution of about 10 microm. The areas of amplified variations in wall shear stress, mostly evident in the neighborhoods of arterial branching, seem to correlate well with clinically observed increased atherogenesis. The intracoronary flow lines showed stasis and extreme vorticity during the phase of minimum coronary flow in contrast to streamlined undisturbed flow during the phase of maximum flow. CONCLUSIONS: Computational tools of this kind along with a state-of-the-art multislice computerized tomography or magnetic resonance-based non-invasive coronary imaging, could enable realistic, repetitive, non-invasive and multidimensional quantifications of the effects of stenosis on distal hemodynamics, and thus help in precise surgical/interventional planning. It could also add insights into coronary and bypass graft atherogenesis. PMID- 15296880 TI - Geometry assessment of coronary artery anastomoses with construction errors by epicardial ultrasound. AB - OBJECTIVE: There is concern about the quality of the distal anastomosis in off pump coronary artery bypass grafting. We investigated the impact of specific construction errors on anastomotic geometry using epicardial ultrasound. METHODS: Twelve ex vivo pressure perfused porcine and five isolated post-mortem human hearts were used to construct 35 internal mammary artery to coronary artery anastomoses, either without (n = 7) or with a standardized construction error (oversutured toe, oversutured heel, cross-over or purse string; each error, n = 7). The anastomotic geometry was visualized and measured by a 13 MHz ultrasound mini-transducer. Impression cast material was used to validate anastomotic geometry. RESULTS: All 28 errors were visualized properly. Two unintended construction abnormalities were observed. In the porcine heart, the ratio of anastomotic orifice area and outflow corner area was 1.3+/-0.2 (mean+/-standard deviation) in the control group and reduced in the error groups: oversutured toe, 0.6+/-0.2 (P = 0.001 oversutured heel, 0.9+/-0.2 (P = 0.037); cross-over, 0.4+/ 0.2 (P < 0.001); purse string, 0.3+/-0.2 (P < 0.001). None of the errors reduced the area of the inflow or outflow corner itself compared to the recipient coronary artery. In the human heart, all construction errors as well as wall plaque were visualized properly. In all anastomoses, ultrasound geometry corresponded to cast geometry. CONCLUSIONS: Ex vivo, epicardial 13 MHz ultrasound enabled accurate visualization and assessment of four different construction errors in the coronary anastomosis. All errors reduced the area of the anastomotic orifice, but not the inflow or outflow corner. PMID- 15296881 TI - Inhibition of inducible nitric oxide synthase and superoxide production reduces matrix metalloproteinase-9 activity and restores coronary vasomotor function in rat cardiac allografts. AB - OBJECTIVE: Oxidants such as nitric oxide (NO) and superoxide are involved in coronary endothelial dysfunction, an early event in the process of allograft coronary atherogenesis, possibly by activation of matrix metalloproteinases (MMPs) and extracellular matrix proteins. We investigated the contribution of inducible nitric oxide synthase (iNOS) derived NO and superoxide on (MMP)-9 activity and to changes in coronary vasomotor function in rat cardiac allografts. METHODS AND RESULTS: An allogenic (Brown Norway to Lewis rats) heterotopic cardiac transplantation model was used to study the effect of continuous treatment with a selective iNOS inhibitor; N-(3-(aminomethyl) benzyl) acetamidine (1400W), and polyethylene glycol conjugated superoxide dismutase (SOD) either alone or in combination on coronary vasomotor dysfunction. 1400W or SOD 24 h alone or their combination improved endothelium-dependent (bradykinin) and independent (sodium nitroprusside) coronary flow reserve and inhibited enhanced MMP-9 protein and activity. In addition, histopathological study revealed that either 1400W or SOD or their combination reduced superoxide production and nitrotyrosine protein. CONCLUSION: The present study demonstrates for the first time that selective iNOS inhibition or SOD treatment reduces enhanced MMP-9 protein and activity associated with improvement of both, endothelium-dependent and -independent coronary vasomotor function in rat cardiac allografts. This is accompanied by reduction of nitrotyrosine and superoxide production. This suggests that the proteolytic enzyme MMP-9 is an effector molecule of oxidant mediated coronary vasomotor dysfunction. PMID- 15296882 TI - Prevention of myocardial reperfusion injury by poly(ADP-ribose) synthetase inhibitor, 3-aminobenzamide, in cardioplegic solution: in vitro study of isolated rat heart model. AB - OBJECTIVE: Cardioplegic arrest remains the method of choice for myocardial protection in cardiac surgery. Poly(adenosine 5'-diphosphate-ribose) synthetase (PARS) inhibitor has been suggested to attenuate the ischemia-reperfusion injury in myocardial infarction by preventing energy depletion associated with oxidative stress. We investigated the efficacy of a cardioplegic solution containing a PARS inhibitor, 3-aminobenzamide (3-AB), for myocardial protection against ischemia reperfusion injury caused by cardioplegic arrest. METHODS: Isolated hearts were set on a Langendorff apparatus and perfused. The hearts were arrested for 90 min with a cardioplegic solution given at 30-min intervals and then reperfused for 20 min. The hearts of rat in the 3-AB(-) group (n = 8) were perfused with a standard cardioplegic solution and terminal warm cardoplegia, whereas the 3-AB(+) group (n = 8) received these solutions supplemented with 3-AB (100 microM). Left ventricular function and release of cardiac enzymes were monitored before and after cardioplegic arrest. After reperfusion, NAD+ (nicotinamide-adenine dinucleotide) levels were assessed, and the tissues were examined immunohistochemically for oxidative stress and apoptosis. RESULTS: During reperfusion, the 3-AB(+) group showed significantly higher (P = 0.005)dp/dt and lower creatine phosphokinase (CPK) level and glucotamic-oxaloacetic transaminase (GOT) in the effluent (CPK; P = 0.003 GOT; P < 0.001) The cardiomyocytes of the 3 AB(+) group also preserved a higher NAD+ level (P < 0.001). Immunohistochemical study of oxidative stress revealed a lesser extent (P = 0.007) of nuclear staining and a lower fraction of apoptosis in the 3-AB(+) group. CONCLUSION: Cardioplegic solution supplemented with 3-AB provides efficient myocardial protection in cardioplegic ischemic reperfusion by suppressing oxidative stress and overactivation of PARS. PMID- 15296883 TI - Peroxynitrite, a product between nitric oxide and superoxide anion, plays a cytotoxic role in the development of post-bypass systemic inflammatory response. AB - OBJECTIVE: Cardiopulmonary bypass (CPB) is known to induce post-bypass systemic inflammatory response. Peroxynitrite (ONOO-) is a potent oxidant formed by a rapid reaction between nitric oxide (NO) and superoxide anion. We hypothesized that ONOO- plays a role in the development of post-bypass systemic inflammatory response and examined the efficacy of ONOO- scavenger in a rat-CPB model. METHODS: Adult Sprague-Dawley rats underwent 60 min of CPB (100 ml/kg per min, 34 degrees C). Group-P (n = 10) received 50 mg/kg of ONOO- scavenger, quercetin, intraperitoneally 24 h before the initiation of CPB, and Group-C (n = 10) served as controls. RESULTS: There were significant time-dependent changes in plasma nitrate+nitrite (NOx), the percentage ratio of nitrotyrosine to tyrosine (%NO2 Tyr: an indicator of ONOO- formation), interleukin (IL)-6, IL-8, and respiratory index (RI). There were significant differences in %NO2-Tyr between the groups both at CPB termination (Group-P vs C; 0.26+/-0.07 vs 0.55+/-0.11%, P < 0.01) and 3 h after CPB termination (0.65+/-0.14 vs 1.46+/-0.25%, P < 0.01); whereas there were no significant differences in NOx between the groups at any sampling point ((at CPB termination) Group-P vs C; 31.6+/-4.3 vs 32.7+/-4.1 micromol/l, (3 h after CPB termination) Group-P vs C; 47.8+/-4.9 vs 51.7+/-5.3 micromol/l). Group P showed significantly lower plasma IL-6 (176.8+/-44.3 vs 302.4+/-78.1 pg/ml, P < 0.01), IL-8 (9.45+/-1.78 vs 16.42+/-2.53 ng/ml, P < 0.01) and RI (1.07+/-0.19 vs 1.54+/-0.25, P < 0.01) 3 h after CPB termination, though there were no significant differences between the groups at CPB termination. CONCLUSIONS: These results suggest that ONOO- plays a crucial role in the development of post-bypass systemic inflammatory response and the pretreatment with quercetin has a potential benefit to avoid deleterious effects of ONOO-. PMID- 15296884 TI - Insulin potentiates expression of myocardial heat shock protein 70. AB - OBJECTIVE: Since insulin stimulates nitric oxide (NO) production and an increase in NO following heat shock is required for myocardial heat shock protein 70 (Hsp70) synthesis, we hypothesized that insulin would enhance myocardial Hsp70 synthesis by augmenting NO signaling. We examined whether a physiologic dose of insulin increased myocardial Hsp70 in unstressed and heat shock treated rats. METHODS: Adult male Sprague-Dawley rats were assigned to groups: (1) control, (2) insulin injected (200 microU/gm body weight), (3) heat shock treated (core body temperature 42 degrees C for 15 min), (4) heat shock and insulin treated, (5) L nitroarginine methyl ester (L-NAME) and heat shock and insulin treated, (6) sodium nitroprusside (SNP) and heat shock and insulin treated. Six hours later, myocardial Hsp70 content and localization was analyzed. RESULTS: Hsp70 was increased in heat shock treated hearts (120.6+/-16.8 ng/mg protein, P < 0.001) vs. control (12.9+/-2.0 ng/mg protein), or insulin treated hearts (15.5+/-0.83 ng/mg protein). In addition, Hsp70 was increased in the heat shock and insulin treated hearts (164.4+/-7.53 ng/mg protein) compared to control, insulin only (P = 0.001) or heat shock only treated hearts (P = 0.01). L-NAME did not abolish the insulin induced increase in Hsp70 in heat shocked hearts (195.2+/-13.4 ng/mg protein, P = 0.21) and SNP did not further enhance Hsp70 in the insulin and heat shocked group (188.9+/-8.2 ng/mg protein, P = 0.71). Western analysis and confocal microscopy revealed a lowlevel expression of myocardial Hsp70 in response to insulin. Hsp70 was localized primarily in blood vessels after insulin or heat shock treatments. CONCLUSIONS: Insulin caused a low-level expression of myocardial Hsp70 and potentiated Hsp70 synthesis in response to heat shock. The ability of insulin to potentiate Hsp70 after heat shock is independent of NO signaling as it was not altered by either LNAME or SNP pretreatment. Blood vessels appear to be the primary site of Hsp70 after insulin or heat shock treatment. PMID- 15296885 TI - Efficacy of FK633, an ultra-short acting glycoprotein IIb/IIIa antagonist on platelet preservation during and after cardiopulmonary bypass. AB - OBJECTIVE: Temporary pharmacologic inhibition of platelet function during and after cardiopulmonary bypass (CPB) (platelet anesthesia) is an attractive strategy for preserving platelets during CPB. We examined the efficacy of FK633, an ultra-short acting glycoprotein IIb/IIIa antagonist. METHODS: The study was carried out in six mongrel dogs that received an intravenous bolus of 0.1 mg/kg of FK633 at the time of administration of heparin (group F), and six control dogs (group C). All animals underwent 60 min of normothermic CPB followed by a 2-h observation period. Blood samples for platelet count, platelet aggregation to adenosine diphosphate and parameters concerning the coagulation system were obtained at eight time points. Hemodynamics, bleeding time, and postoperative blood loss were assessed serially. Scanning electron micrograph of the oxygenator's membrane was investigated. RESULTS: FK633 significantly protected platelet number (group F, 59+/-10% versus group C, 38+/-15% of the pre-CPB value; P < 0.01), and inhibited platelet aggregation to adenosine diphosphate (group F, 13+/-12% versus group C, 35+/-9% of the pre-CPB value; P < 0.01) during CPB. Postoperative blood loss did not significantly differ between the two groups, but there was a tendency of less bleeding in group F (group F, 73+/-23 ml versus group C, 111+/-44 ml; P = 0.09). In group F, scanning electron micrograph of the oxygenator's membrane showed that its surface was free from platelets. There were no significant differences between the groups in hemodynamics. CONCLUSIONS: An ultra-short acting glycoprotein IIb/IIIa antagonist, FK633, is effective in preventing both platelet aggregation and thrombocytopenia during CPB, and may be effective for minimizing postoperative bleeding. PMID- 15296886 TI - The effect of leucodepletion on leucocyte activation, pulmonary inflammation and respiratory index in surgery for coronary revascularisation: a prospective randomised study. AB - OBJECTIVE: Leucocyte activation is central to end-organ damage that occurs during cardiac surgery under cardiopulmonary bypass (CPB). Exhaled nitric oxide (NO) increases in inflammatory lung conditions and has been proposed as a marker of pulmonary inflammation during CPB. This study examined the effect of leucodepletion on leucocyte activation, pulmonary inflammation and oxygenation in patients undergoing coronary revascularisation. METHODS: Fifty low-risk patients undergoing first time coronary artery bypass graft (CABG) were randomised to two groups. Twenty-five patients had an arterial line leucocyte-depleting filter and 25 controls had a standard filter. Arterial blood samples were taken before CPB, 5 and 30 min on CPB, 5 min after aortic clamp removal and 6 h post-operatively. Activated leucocytes were identified with Nitroblue Tetrazolium staining. NO was sampled via an endotracheal teflon tube 15 min after median sternotomy before CPB and 30 min after discontinuation of CPB using a real-time chemiluminescense analyser. Respiratory index (alveolar-arterial oxygenation index, AaOI) was calculated before CPB, 1, 2, 4, 8 and 18 h post-operatively. Clinical outcome end points were also recorded. RESULTS: Total and activated leucocyte counts were significantly lower following leucodepletion during CPB (P < 0.0001). Exhaled NO rose significantly after CPB in the control group (3.8+/-1 ppb/s before CPB vs 5.6+/-2 ppb/s after CPB (P = 0.003) but not in the leucodepleted group (3.7+/-1 ppb/s before CPB vs 3.9+/-1 ppb/s after CPB (P = 0.051). AaOIs were consistently lower after leucodepletion (anova, P = 0.001). The duration of mechanical ventilation, the intensive care and hospital stay and the frequency of cardiac and respiratory complications were similar in the two groups. CONCLUSIONS: Leucodepletion reduces the numbers of circulating activated leucocytes and the pulmonary inflammation during CPB. This appears to limit lung injury and improve oxygenation in low-risk patients undergoing CABG surgery. Larger numbers of patients are required to evaluate the effect of continuous arterial line leucodepletion on the clinical outcome. PMID- 15296887 TI - Aqueous oxygen: the solution to relief hypoxic pulmonary hypertension. AB - OBJECTIVE: To evaluate the effects of hyperbaric oxygen solution on hypoxic pulmonary hypertension. METHODS: Eleven calves, 2-month-old, 71+/-6 kg, underwent general anaesthesia, mechanical ventilation and median sternotomy. Catheters for continuous pressure and blood gas measurements were inserted in carotid and femoral arteries, left atrium, right atrium and pulmonary artery (PA), and a flow probe placed around the PA. After baseline measurements 30 min hypoxic ventilation reduced the mean arterial PO2 from 285+/-115 to 46+/-11 mmHg (P < 0.0001). At this point, without changes in hypoxic ventilation (mean arterial PO2 maintained at 50+/-5 mmHg), 3 ml/min of hyperbaric aqueous oxygen (AO, oxygen diluted in saline solution) was infused directly into the PA for 30 min, with continuous reading of the monitored parameters. RESULTS: Hypoxic ventilation raised significantly (P < 0.005) the values of systolic (36+/-7 vs 22+/-6 mmHg), diastolic (16+/-3 vs 9+/-4 mmHg) and mean (24+/-4 vs 14+/-4 mmHg) PA pressure, PA/systemic pressure ratio for systolic (0.47+/-0.09 vs 0.24+/-0.06) and mean (0.49+/-0.13 vs 0.23+/-0.08) pressures and Pulmonary Vascular Resistance (PVR) (6.89+/-0.87 vs 2.67+/-0.38 U), while the Pulmonary Blood Flow (PBF) decreased (2.7+/-0.4 vs 3.7+/-0.4 l/min). AO infusion reduced significantly (P < 0.005) the values obtained with hypoxic ventilation with systolic (26+/-6 vs 36+/-7 mmHg), diastolic (11+/-4 vs 16+/-3 mmHg) and mean (16+/-4 vs 24+/-4 mmHg) PA pressure, PA/systemic pressure ratio for systolic (0.27+/-0.07 vs 0.47+/-0.09) and mean (0.27+/-0.08 vs 0.49+/-0.13) pressures and PVR (3.42+/-0.31 vs 6.89+/-0.87 U), while the PBF increased (3.6+/-0.4 vs 2.7+/-0.4 l/min). CONCLUSIONS: Acute infusion of hyperbaric AO solution into the PA completely reverses the negative effects of acute hypoxia on pulmonary circulation. PMID- 15296888 TI - Hypophosphatemia following open heart surgery: incidence and consequences. AB - OBJECTIVE: Significant hypophosphatemia (SH) is common after major surgery and may be associated with considerable morbidity, including respiratory and cardiac failure. The contribution of SH to these complications after cardiac surgery is not well defined. METHODS: In this prospective study, levels of serum phosphorus and other electrolytes (potassium, magnesium and calcium) were measured in 566 consecutive patients (395 men, 182 women; mean age 65.5+/-11.1 years) undergoing elective cardiac surgery at three time points: prior to surgery, immediately on admission to the ICU, and on the first postoperative day. Preoperative (type of surgery, Bernstein-Parsonnet risk estimate), intraoperative (duration of bypass and cross-clamp, intraoperative fluid and blood product use) and postoperative data (duration of ventilation, duration of ICU and hospital stay, requirement for cardioactive drug support, development of atrial fibrillation, and mortality) were collected. Patients were divided into two groups according to the immediate postoperative phosphate level: SH, phosphate <0.48 mmol/l (mean phosphate 0.28+/ 0.13 mmol/l, n = 194), and a control group (mean phosphate value 0.84+/-0.08 mmol/l, n = 372). Patients with SH received treatment with sodium or potassium phosphate (0.8 mmol/kg body weight over 6-12 h). RESULTS: SH was present in 34.3% of patients. There were no differences in the baseline characteristics between the two groups. Patients with SH received more intraoperative blood product transfusions. The postoperative course of patients with SH was characterized by prolonged ventilation (2.1+/-1.7 versus 1.1+/-0.9 days, P = 0.05), more patients requiring cardioactive drugs (12-24 h 16 versus 10.9%, P = 0.05 and >24 h 23.5 versus 13.8%, P = 0.05); and a prolonged hospital stay (7.8+/-3.4 versus 5.6+/ 2.5 days, P = 0.05). CONCLUSIONS: SH was common after open-heart surgery and was associated with an increased incidence of important complications. We suggest that phosphate levels be routinely measured immediately after surgery and appropriate therapy instituted. PMID- 15296889 TI - Prophylactic tranexamic acid in elective, primary coronary artery bypass surgery using cardiopulmonary bypass. AB - OBJECTIVE: Perioperative use of tranexamic acid (TA), a synthetic antifibrinolytic drug, decreases perioperative blood loss, and the proportion of patients receiving blood transfusion in cardiac surgery, but the results may vary in different clinical settings. The primary objective of the present study was to determine the efficacy of TA to decrease chest tube drainage and the proportion of patients requiring perioperative allogeneic transfusions following primary, elective, on-pump coronary artery bypass grafting (CABG) in patients with a low baseline risk of postoperative bleeding. METHODS: In a double-blinded, prospective, placebo-controlled study, 46 patients were randomized into two groups. One group received TA 1.5 g as a bolus, followed by a constant infusion of 200 mg/h until 1.5 g. The other group received placebo (0.9% saline). Among exclusion criteria were treatment with acetylsalicylic acid, non-steroidal anti inflammatory drugs or other platelet inhibitors within 7 days before surgery. RESULTS: Preoperative demographics, biochemical and surgical characteristics were comparable between groups. At 6 h postoperatively, there was a trend towards a greater blood loss (median and interquartile range) in the placebo group (710 and 460-950 ml) compared to the TA group (400 and 350-550 ml), but the difference did not reach statistical significance. Neither were transfusion rates and the amount of autotransfused shed mediastinal blood different between the groups postoperatively. Postoperative d-dimer concentrations were significantly higher in the placebo group compared to the TA group (P < 0.001) This difference could not be explained by differences in the amount of autotransfused shed mediastinal blood alone. Plasma concentrations of beta-thromboglobulin and platelet factor 4 were significantly increased postoperatively in both groups, but without any intergroup differences. Seven patients (15%), one in the TA group and six in the placebo group, were reoperated due to excessive bleeding. Surgical correctable bleeding was found in all except two patients from the placebo group. CONCLUSIONS: An antifibrinolytic effect following prophylactic use of TA in elective, primary CABG among patients with a low risk of postoperative bleeding, did not result in any significant decrease in postoperative bleeding compared to a placebo group. PMID- 15296890 TI - Limitations of additive EuroSCORE for measuring risk stratified mortality in combined coronary and valve surgery. AB - OBJECTIVE: To study the use of the additive and logistic European System for Cardiac Operative Risk Evaluation (EuroSCORE) to predict mortality following adult combined coronary artery bypass grafting (CABG) and valve surgery. METHODS: Data were collected prospectively, from all four centres providing adult cardiac surgery in the north west of England, on 1769 consecutive patients undergoing combined CABG and valve surgery between April 1997 and March 2002. Observed in hospital mortality was compared to predicted mortality as determined by both additive and logistic EuroSCORE. RESULTS: Observed mortality for simultaneous CABG and valve surgery was 8.7%, compared to 6.7% (additive) and 9.4% (logistic). Sixty-five percent of patients were classified as high-risk (additive EuroSCORE >5); the observed mortality was 11.5%, compared to 8.1% (additive) and 12.8% (logistic). Discrimination was similar in both systems as measured by the C statistic (additive 0.73, logistic 0.73). CONCLUSIONS: The logistic EuroSCORE is more accurate at predicting mortality in simultaneous CABG and valve surgery, as the additive EuroSCORE significantly under-predicts in this high-risk group. PMID- 15296891 TI - Surgical ablation of post-infarction ventricular tachycardia guided by mapping in sinus rhythm: long term results. AB - OBJECTIVE: Some patients after myocardial infarction have an increased risk of malignant ventricular tachyarrhythmias (VTA) or sudden cardiac death. The aim of the study was to evaluate long-term results of surgical ablation of an arrhythmogenic substrate guided by simplified intraoperative mapping of pathological ventricular electrograms during sinus rhythm. METHODS: The study population consisted of 77 patients (9 women; mean age 62.4+/-8.5 years) with previous Q-wave myocardial infarction and at least one documented episode of sustained VT/VF more than one month after the last infarction. The left ventricular ejection fraction was 31.3+/-8.8%. All but eight patients had clinical indication for concomitant coronary artery bypass surgery. All underwent preoperative electrophysiologic study. Intraoperative epicardial and endocardial mapping during sinus rhythm was performed using a multielectrode with 16 bipolar electrodes in combination with a multichannel recording system. Myocardial regions revealing fractionated, low amplitude signals lasting > or =130 ms were surgically excised or cryoablated. All surviving patients were restudied within one to two weeks after surgery using identical programmed electrical stimulation protocol. RESULTS: Five (6.5%) patients died in the perioperative (30-days) period. In the remaining cohort, inducibility of any sustained VTA after surgical procedure was observed in 21 subjects (29.2%). An implantable cardioverter defibrillator (ICD) was implanted in these patients. Recurrence of sustained VTA was documented during follow-up period in two patients who were noninducible after the surgery (at the month 10 and 22, respectively), and both received ICD as well. No patient died of sudden cardiac death. In 14 ICD patients, no significant VTA was documented during the mean follow-up of 37.3+/-23.2 months. Altogether, 61 from the 72 patients surviving the surgery (84.7%) remained free of spontaneous recurrences of VTA during the follow-up. CONCLUSIONS: Surgical ablation of an arrhythmogenic substrate guided by simplified intraoperative mapping in normothermic heart during sinus rhythm appears to be both safe and efficacious procedure that prevents recurrences of VTA in a substantial proportion of patients. PMID- 15296892 TI - Perioperative outcome and long-term survival of surgery for acute post-infarction mitral regurgitation. AB - OBJECTIVE: To determine factors predictive of mortality in patients undergoing emergency mitral valve surgery in the setting of severe post-infarction regurgitation. METHODS: Patients admitted for an acute myocardial infarction who required urgent mitral valve surgery for severe regurgitation were studied. Factors predictive of outcome were analysed. RESULTS: Fifty-five consecutive patients (mean 65+/-10 years, 37 males) were included. The infarct was inferior in 31 patients, posterior in 10, anterior in 9 and lateral in 5. Thirty-four patients (62%) were in Killip class IV. Peroperative findings confirmed total papillary muscle rupture in 25 patients (posteromedial in 21, anterolateral in 4), and partial rupture in 12 patients (posteromedial in 10, anterolateral in 2). Papillary muscle dysfunction without rupture was responsible for regurgitation in 18 patients (posteromedial in 15, anterolateral in 3). The mitral valve was replaced by a prosthesis in all but 4 patients, who had valvuloplasty. Coronary angiography was done in 32 patients, of whom 18 underwent concomitant coronary artery bypass grafting and 2 balloon angioplasty. Surgery was performed on average 7 days after infarction. Thirteen patients (24%) died during the perioperative period. Absence of coronary revascularisation was significantly associated with increased perioperative mortality (34% vs. 9%, P = 0.02). Of the 42 surviving patients, there were 5 deaths during a mean follow-up of 4.0+/-3.7 years. CONCLUSION: In patients with acute post-infarction mitral regurgitation, perioperative mortality is high, but can be improved with concomitant CABG in addition to valve surgery. Long-term outcome of survivors is favourable. PMID- 15296893 TI - Excellent results for atrial fibrillation surgery in the presence of giant left atrium and mitral valve disease. AB - OBJECTIVE: The incidence of sinus conversion in the enlarged left atrium after atrial fibrillation surgery is reported to be low. The purpose of the current study was to investigate the effects of atrial fibrillation surgery on mitral valve disease associated with a giant left atrium (GLA). METHOD: From July of 1997 to February of 2002, 188 patients received mitral valve and atrial fibrillation surgery. The patients were placed in either GLA group (n = 94), or NGLA group (n = 94), based on LA size. The presence and onset of sinus rhythm and the incidence and velocity of transmitral A waves were monitored during the early postoperative period and throughout the follow up period of 42 months. RESULTS: The onset of postoperative sinus rhythm was slightly earlier in the NGLA group than in the GLA group at 1.3+/-0.4 days versus 3.1+/-1.2 days, respectively, (P = 0.008). The sinus conversion rates in the GLA and the NGLA groups were 91.5 and 97.9% in the early postoperative period, and 94.7 and 95.7% at 6 months after surgery, respectively. A wave appearance rates in the early postoperative period in the GLA and the NGLA groups were 62.2 and 71.7%, and continued to improve over time to 94 and 95% by 36 months, respectively. Peak A wave velocities in the early postoperative period in GLA and NGLA groups were 67.4+/-34.0 and 61.1+/ 29.5 cm/s without significant change during the follow up. CONCLUSION: The results suggest that atrial fibrillation surgery is effective at inducing sinus rhythm and restoring left atrial contractile function after concomitant mitral valve surgery regardless of LA size. PMID- 15296894 TI - Impact of surgical era on outcomes of patients undergoing elective atherosclerotic ascending aortic aneurysm operations. AB - OBJECTIVE: This retrospective study evaluates, if recent refinements in peri operative management, have an impact on clinical outcome of patients undergoing elective repair of their ascending thoracic aorta. METHODS: One hundred sixty five (n = 165) consecutive patients were operated during a 7 year period at our department. The cohort was divided in an early group I (from Jan 1997 to Dec 1999, n = 75) and a late group II (from Jan 2000 to Jan 2003, n = 90). The mean age was 60.9+/-13.1 years in group I versus 58.1+/-13.6 years in group II. In group I 50 patients (66.6%) underwent replacement of the ascending thoracic aorta alone, 17 patients (22.6%) received a composite graft, 8 patients (10.6%) had an additional aortic valve replacement and 14 patients (18.6%) needed concomitant coronary artery bypass grafting. In group II the procedures were as follows: interposition graft alone in 58 patients (64.4%), composite graft in 26 patients (28.8%), aortic valve replacement in 6 (6.6%) and CABG in 11 patients (12.2%). RESULTS: Overall hospital mortality for the entire cohort was 6.6% (11/165) with no significant differences between the early and late group with 6.6% (5/75) and 6.6% (6/90), respectively, P = 0.985. Causes were multi organ failure in 63.3% (n = 7), stroke in 9% (n = 1) myocardial infarction in 18.1% (n = 2) and refractory bleeding in 9% (n = 1). Concomitant CABG, repair of the aortic valve and composite graft, emerged as independent risk factor for mortality in multivariate logistic regression analysis with P = 0.001. Differences, became apparent in ICU as well as hospital stay with a median ICU stay in group I of 7.1+/-12.9 days versus 4.4+/-6.8 days in group II, and median hospital stay of 16.7+/-5.3 days versus 9.5+/-8.4 days for group I and II, P < 0.05, respectively. Furthermore through the implementation of blood conservation techniques, a substantial reduction of transfusion requirements could be achieved (PRBC from 3.2+/-4 to 1.1+/-1.7 units, FFP 5.2+/-3 to 2.3+/-0.5 units, Platelets from 1.3+/-2 to 0.3+/ 0.07 units). CONCLUSIONS: Even with the implementation of various refinements in surgical and anaesthetic techniques, the current risk of mortality for ascending aortic aneurysm repair has not changed in the last 7 years. However, shortened ICU and hospital stays as well as diminished usage of blood derivates are mainly the result of a more aggressive and improved peri- and post-operative management due to economic considerations. PMID- 15296895 TI - Changes in coagulation condition, cytokine, adhesion molecule after repair of type A aortic dissection. AB - OBJECTIVE: Because residual dissection often exists even after the repair of a type A dissection, we evaluated coagulation conditions, cytokine levels, and adhesion molecule levels in mid-term follow up after repair of type A dissections. METHODS: Thrombin-antithrombin III complex (TAT), D-dimer, soluble interleukin-2 receptor (sIL-2R), soluble intercellular adhesion molecule (sICAM) 1, and type III procollagen peptide (PIIIP) were measured in 12 patients (mean age=63 years) following the repair of a type A aortic dissection at 6-82 months after repair (median=33 months). RESULTS: In the chronic phase, TAT and D-dimer were significantly higher in patients following the repair of a type A dissection compared to healthy controls (TAT; 12+/-8 vs. 2.5+/-1.2 ng/ml, P = 0.0001, D dimer; 779+/-1384 vs. 104+/-46 U/ml, P = 0.0001). Cytokine was significantly higher in the affected patients (sIL-2R; 556+/-205 vs. 398+/-132 U/ml, P = 0.003, sICAM-1; 255+/-131 vs. 211+/-48 ng/ml, P = 0.136). Collagen turnover (PIIIP) showed a significantly higher value in the affected patients (0.80+/-0.32, vs. 0.58+/-0.13 U/ml, P = 0.002). sIL-2R, sICAM-1 and PIIIP showed a negative correlation with the follow-up period (sIL-2R; r = -0.733, P = 0.0067, sICAM-1; r = -0.61, P = 0.035, PIIIP; r = -0.692, P = 0.0126). We found a positive correlation between aortic size and TAT (r = 0.644, P = 0.0238, n = 12) as well as with D-dimer (r = -0.7831, P = 0.0106, n = 12) and TAT showed significantly higher values in the residual dissection group compared to those without residual dissection (16.6+/-7.9 vs. 7.45+/-4.75 ng/ml, P = 0.035). CONCLUSION: Hypercoagulation conditions continued even after repair. Both TAT and D-dimer would be good indices for following up patients having repaired aortic dissections. Furthermore, cytokine, adhesion molecules, and collagen turnover would return to a stable state unless impairment and expansion of the vessel wall occurred. PMID- 15296896 TI - Repair of the descending thoracic aorta: impact of open distal anastomosis technique on spinal cord perfusion, neurological outcome and spinal cord histopathology. AB - OBJECTIVES: We investigated the impact of equilibrating distal aortic pressure with atmospheric pressure (open distal anastomosis) on spinal cord perfusion, neurological outcome and spinal cord histopathology in a rat model of descending thoracic aortic surgery. METHODS: Proximal thoracic aortic occlusion was obtained in Sprague-Dawley rats by inflating the balloon of a 2F Fogarty catheter introduced through the left femoral artery. Rats were separated into three groups: sham-operation (n = 5) without balloon inflation, control (n = 15) with inflation of the balloon, and open distal (n = 15) with inflation of the balloon combined with incision of the right femoral artery to allow free drainage of distal aortic blood. Balloon inflation was maintained for 15 min. Rectal temperature, arterial blood gases and pH, distal arterial blood pressure (DABP) and lumbar spinal cord blood flow (SCBFl) were recorded throughout the procedure. Neurobehavioral status was assessed daily using a 0-5 scale and rats were sacrificed after 48 h of reperfusion and their spinal cord harvested for histopathology and immunohistochemistry for microtubule-associated protein-2 (MAP 2). RESULTS: DABP and SCBFl values were lower during thoracic aortic occlusion in the open distal group, compared to the control group (P < 0.001). Paraplegia and mortality rates were dramatically increased in the open distal group (87.7 and 46.6%, respectively) compared to the control group (0 and 6.6%, respectively, (P < 0.001 and 0.02). Severe metabolic acidosis and bowel infarct were also more frequent in the open distal group (P < 0.001). Sham-operated and control rats had virtually normal spinal cords, whereas rats in the open distal group had severe ischemic injury throughout gray matter. CONCLUSIONS: Equilibrating distal arterial pressure with atmospheric pressure during thoracic aortic occlusion decreased spinal cord blood flow, increased mortality and worsened spinal cord injury in rats. These results suggest that the open distal anastomosis technique should be used with caution in patients undergoing repair of the descending thoracic or thoracoabdominal aorta. PMID- 15296897 TI - Effects of the patent false lumen on the long-term outcome of type B acute aortic dissection. AB - OBJECTIVE: To determine the most effective treatment, we performed a detailed comparative study of the clinical course of patients with type B aortic dissection with a patent or thrombosed false lumen who did not undergo surgery in the acute period. We examined the effect of patency of the false lumen on outcome. METHODS: Computed tomography scans of 138 patients with type B acute aortic dissection were reviewed. Of 138 patients, 110 were medically treated and survived the acute period. We focused on the outcome of these 110 patients, 62 with medically treated thrombosed false lumen (thrombosed group) and 48 with medically treated patent false lumen (patent group). We investigated factors influencing outcome among the 110 patients. The follow-up period was up to 10 years after the onset of aortic dissection. The three study endpoints were death from any cause, dissection-related death (aortic rupture, perioperative death, or death due to organ ischemia), and a dissection-related event (aortic rupture or surgery). In the patent group, we investigated factors influencing long-term outcome. RESULTS: Patency of the false lumen was an independent risk factor for dissection-related death (P = 0.038, hazard ratio=5.6, confidence interval=1.1 28) and for a dissection-related event (P = 0.000, hazard ratio=7.6, confidence interval=2.7-22) but not for death from any cause (P = 0.769, hazard ratio=1.2, confidence interval=0.45-2.91). In the patent group, location of the most dilated aortic segment at the distal arch was an independent risk factor for dissection related death (P = 0.026, hazard ratio=13.6, confidence interval=1.4-135) and for a dissection-related event (P = 0.048, hazard ratio=2.6, confidence interval=1.0 6.9). CONCLUSIONS: Patency of the false lumen is a strong independent prognostic factor for type B aortic dissection. Location of the most dilated aortic segment at the distal arch is a significant risk factor in the patients with a patent false lumen. PMID- 15296898 TI - Primary sternal plating in high-risk patients prevents mediastinitis. AB - OBJECTIVE: Sternal wound infection leading to post-operative mediastinitis is a devastating complication of cardiac surgery carrying nearly a 15% mortality rate despite current treatment methods. Instability of bone fragments pre-disposes a patient to have non-union, mal-union and can subsequently lead to deep sternal wound infections progressing to mediastinitis. Rigid plate fixation has been utilized for acquired and surgically created fractures of virtually every bone in the body to prevent instability. However, the current standard for sternotomy closure remains the method of wire-circlage. Application of rigid plate fixation for sternal osteotomies affords greater stability of the sternum. We report on our preliminary experience with this technique in high-risk patients. METHODS: From July of 2000 to December 2001, rigid plate fixation was applied to 45 patients designated as having high risk for sternal dehiscence and subsequent mediastinitis. High risk was defined as patients having 3 or more established historical risk factors, including: COPD, Re-Operative Surgery, Renal Failure, Diabetes, Chronic Steroid Use, Morbid Obesity, Concurrent Infection and Acquired or Iatrogenic Immunosuppression. Intra-operative risk factors included off midline sternotomy, osteoporosis, long cardio-pulmonary bypass runs (>2 h), transverse fractures of the sternum. Rigid plate fixation was performed using a combination of plates secured by bi-cortical screws, after the cardiac surgical procedure was complete and hemostasis was secured. RESULTS: Rigid plate fixation was performed on 26 males and 19 females. The average age of patients was 63 (43 88) years. The average follow-up was 15 weeks (range 8-41 weeks). While there were 4 peri-operative deaths unrelated to sternal closure: one from aspiration pneumonia (post-operative day 9), one from a pulmonary embolus (post-operative day 29), one from overwhelming sepsis from pre-existing endocarditis (post operative day 15), and one for primary respiratory failure (post-operative day 12). All others healed successfully. One patient who had a sterile dehiscence subsequently underwent successful re-operative rigid fixation. Comparing the cohort of patients who received rigid plate fixation to a matched population of high-risk patients during a similar time period who received wire closure, revealed a significant difference in the incidence of post-operative mediastinitis. The wire closed group (n = 207) had 18 deaths unrelated to sternal closure and had 28 patients who developed mediastinitis (14.8%). The rigid plate fixation group had no mediastinitis (Fisher's exact test, P = 0.006). The total incidence of post-operative mediastinitis during the designated study period was 4.2%. CONCLUSION: Patients who benefited from sternal closure with rigid plate fixation showed a significant decrease in the incidence of post-operative mediastinitis when compared to similar population of patients whose sterna were closed with wire. PMID- 15296899 TI - Management of 150 flail chest injuries: analysis of risk factors affecting outcome. AB - OBJECTIVE: Flail chest continues to be an important injury with significant complications. The records of 150 patients presenting with flail chest injury were reviewed to determine risk factors affecting morbidity and mortality. MATERIAL AND METHOD: During a 7-year period 150 patients with a flail chest injury were admitted to our trauma center. There were 111 men (74%) and 39 women (26%) ranging in age from 18 to 88 years with a mean age of 56.9. Only 66 (44%) had an isolated flail chest injury on admission. The majority of patients were older than 55 years (n = 89, 59.3%), 80 (53.3%) presented with an hemo-, or/and pneumothorax, 36 (24%) sustained a head injury and 25 (16.7%) needed ICU monitoring. The mean ISS score was 38. Age, concomitant diseases, presence of pneumothorax and/or hemothorax, Severity Score (ISS), the need for mechanical support, length of stay and deaths were evaluated by using the t-test and chi2 test where appropriate. RESULTS: Sixty-seven patients (44.6%) were conservatively treated, while 80 (53.3%) needed thoracic drainage. Only in 6 cases (4%) thoracotomy was required, while in 9 (6%) laparotomy was performed. Mortality rate reached 5.3%. The main factors correlated with an adverse outcome were: ISS and the presence of associated injuries, while age, hemopneumothorax and mechanical support affected the length of hospitalization but not the mortality. CONCLUSIONS: (1) Age and hemopneumothorax did not affect mortality. (2) ISS was found to a strong predictor on outcome concerning morbidity and prolonged hospitalization but did not influence mortality rate. (3) Mechanical support was not considered a necessity for the treatment of flail chest. PMID- 15296900 TI - Emergency thoracotomy in the pre-hospital setting: a procedure requiring clarification. AB - OBJECTIVE: The aim of this study is to investigate the influence of Emergency Thoracotomy (ET) on mortality in a group of patients suffering from severe thoracic trauma requiring Helicopter Emergency Medical Service (HEMS) transfer to hospital. This is not clearly defined especially when thoracotomy takes place in the pre-hospital setting. METHODS: A retrospective review of 670 consecutive patients with severe thoracic trauma, transferred to The Royal London Hospital by HEMS between November 1994 and December 2002. ET (on scene, in the Accident and Emergency (A&E) department or in the operating theatre) was performed in 53 patients (7.7%). Both univariate and multivariate analyses were performed to evaluate ET as an independent predictor of mortality. RESULTS: There were 510 males and 160 females with a mean Injury Severity Score (ISS) of 35.12+/-17.5. Univariate analysis identified ET to be a predictor of mortality (OR=0.15, 95% CI=0.07-0.30). However, with multivariate analysis, ET was not found to be an independent predictor of mortality (OR=1.93, 95% CI=0.61-6.1). The independent predictors of mortality identified were: age>60 years (OR 5.57, 95% CI 2.19 14.16), Glasgow Coma Score <8 at the scene (OR=7.4, 95% CI=3.15-17.46), ISS>25 (OR 5.3, 95% CI=1.64-17.11), need for intubation at the scene (OR=2.80, 95% CI=1.022-7.69), oxygen saturation in A&E (<89%) (OR=2.39, 95% CI=1.13-5.05), haemothorax (OR=3.30, 95% CI=1.53-7.13) and bilateral injury (OR=3.1, 95% CI=1.51 6.61). CONCLUSIONS: Our study has shown that when confounding variables are accounted for, ET is not a predictor of mortality following severe chest trauma. This implies that in a well-selected group of patients it may be a significant and life-saving procedure. PMID- 15296902 TI - Blunt thoracic trauma in children: review of 137 cases. AB - OBJECTIVE: Thoracic injuries are uncommon in children and few report present on blunt ones. METHODS: Between 1994 and 2003, 137 children with blunt thoracic injury were reviewed. RESULTS: The mean age of children was 6.9+/-7.3 (1-16) years. Etiology was falls in 46.7%, traffical accidents in 51% and abuse in 2.2%. Average height in fallen-down cases was 6.4+/-2 (range: 3-11) m. Calculated mean kinetic energy transfer to body was 1923+/-1056 J. When first seen, 70% (82/117) of the patients had vital signs that were within normal limits. Forty-two (35.9%) children had isolated thoracic injury. Associated injuries were present in 75 (64.1%) children. Head injury was the most common associated injury present in 33 (28.2%). Pulmonary contusion was the most common thoracic injury with 68 (49.6%). Seventeen (12.4%) required surgery, 11 (8%) of them were thoracic (4 for diaphragmatic tear, 2 for flail chest, 2 for tracheobronchial injuries, 2 for laceration, 1 for esophageal rupture). Surgical group had higher ISS (26.8 vs 36.2, P = 0.001). Fifteen were lost (10.9%): There were lethal injuries in 7; chest tube treatment in 3; intensive care unit management in 2; mechanical support in 2 and observation in 1 patient. No death occurred for operations. Mortality rate was the lowest at injuries to chest alone and the highest for multi-system injuries (P < 0.05). The hospital length of stay for averaged 13.4+/ 8.8 (range: 4-49) days. CONCLUSION: Associated injury is the most important mortality factor. Thoracic operations can be performed with minimal morbidity and without mortality in children with blunt thoracic trauma. PMID- 15296903 TI - A prospective audit evaluating the role of video-assisted cervical mediastinoscopy (VAM) as a training tool. AB - OBJECTIVE: Cervical mediastinoscopy is an important diagnostic and staging technique. Limited operative field and visibility have traditionally made it a difficult procedure to learn and supervise. Video-assisted techniques can aid training in the procedure. We designed a prospective study to assess the usefulness of video-assisted mediastinoscopy (VAM) as a training tool. METHODS: 43 patients were operated upon by two trainees during their initial formation in general thoracic surgery (25 patients in 15 months, and 18 patients in 9 months, respectively). INDICATIONS: staging (n = 23), diagnosis of enlarged mediastinal nodes (n = 14) and diagnosis/staging (n = 6). End-points of the study: operative time, need of consultant assistance during procedures, and ability of the trainee to identify all nodal stations independently. RESULTS: There were no complications. The mean operative time was 29 (range 18-51) min. Valid histological samples were obtained in all cases. There were no false negative results in the 13 patients who underwent subsequent lung resection (sensitivity 100%). Operative time (R2 = 0.83 and 0.77), need for consultant assistance (R2 = 0.98 and 0.94), and failure to independently reach all nodal stations (R2 = 0.95 and 0.94) significantly decreased with experience in both trainees' cases (cubic curve fit; P < 0.001 throughout). DISCUSSION: VAM permits a rapid learning and adequate supervision of the technique without compromising safety, operative time or completeness of the procedure. The main advantages are: increased visual field, image magnification, adequate light source and the ability to use two instruments simultaneously. VAM should be the technique of choice in thoracic surgical teaching units. PMID- 15296904 TI - Comparative analysis of T2 selective division of rami-communicantes (ramicotomy) with T2 sympathetic clipping in the treatment of craniofacial hyperhidrosis. AB - OBJECTIVE: The main cause of dissatisfaction after sympathetic trunk blocking surgery (T2 sympathectomy, sympathetic clipping) for craniofacial hyperhidrosis is compensatory sweating. Preserving sympathetic trunk may decrease the incidence of compensatory sweating, and we introduce T2 ramicotomy, which may better preserve the sympathetic nerve trunk in order to reduce compensatory sweating. METHODS: From January 2000 to November 2002, video-assisted thoracoscopic (VAT) T2 sympathetic clipping and VAT ramicotomy were performed in 44 patients suffering from craniofacial hyperhidrosis. Twenty-two patients underwent T2 sympathetic clipping (group 1), and 22 underwent division of T2 rami communicantes (group 2). We retrospectively analyzed the rate of satisfaction, dryness of face, and grade of compensatory sweating. RESULTS: Both groups were similar with respect to facial dryness (P = 0.099). Group 1: excessive dry 5 patients (22.7%), dry 17 patients (77.3%); group 2: excessive dry 3 patients (13.6%), dry 15 patients (68.1%), and persistent sweating 4 patients (18.3%). The rate of satisfaction was 77.3% in group 1, and 63.6% in group 2 with no significant difference (P > 0.05). The rate of compensatory sweating in group 2 (72.7%) was significantly lower than in group 1 (95.4%) (P < 0.039). The chance of embarrassing and disabling compensatory sweating was lower in group 2 than in group 1; 76.5% (embarrassing in 8 patients, disabling in 9) in group 1, and 36.4% (embarrassing in 7 patients, disabling in 1) in group 2 which was statistically significant (P < 0.006). CONCLUSIONS: T2 ramicotomy for craniofacial hyperhidrosis lowers the rate of compensatory sweating and excessive dryness of face compared to T2 clipping. PMID- 15296905 TI - Thymic carcinoma. Clinical institutional experience with 15 patients. AB - OBJECTIVE: We retrospectively evaluated 15 patients with thymic carcinoma treated with various modalities and investigated overall management of this disease. METHODS: From 1983 to 2003, we treated 15 patients with thymic carcinoma (12 squamous cell carcinomas, 2 undifferentiated carcinomas and one adenocarcinoma). According to Masaoka's staging system, they consisted of 2 at stage II, 5 at stage III, 4 at stage IVa and 4 at stage IVb. RESULTS: Ten patients were histologically diagnosed preoperatively, and 5 patients underwent an exploratory procedure under the diagnosis of thymoma or benign teratoma. Complete resection was performed in 9 patients (2 stage II, 5 stage III and 2 stage IVa), which included 4 patients who received induction therapy, 4 who received postoperative radiation therapy, and 1 who received postoperative chemotherapy. Six patients with unresectable tumors were treated by irradiation (40-60 Gy) with or without chemotherapy. The median survival was 13 months for patients without resection, and 57 months for patients with a complete resection. Total 3-year and 5-year survival rates were 51.9 and 39.0%, respectively. CONCLUSIONS: We concluded that a complete resection is mainstay of therapy when possible, but chemoradiation therapy being potential benefit in the management of thymic carcinoma. However, considering the high prevalence of advanced stage patients, to establish the effective regimen of induction therapy in the additional multicenter trials should be mandatory. PMID- 15296906 TI - Safety for preoperative use of steroids for transsternal thymectomy in myasthenia gravis. AB - OBJECTIVE: Effects of preoperative steroids on morbidity and mortality after transsternal thymectomy are analyzed. METHODS: There were 620 transsternal thymectomies for myasthenia gravis performed in the period 1973-2002. There were 547 patients with nonthymomatous myasthenia, including 17 patients submitted to repeated thymectomy (rethymectomy) and 46 patients with thymoma. There were 522 upper median sternotomies performed in the period 1973-1997 and 98 complete median sternotomies performed in the period 1998-2002 (74 patients without thymoma, 7 thymomas and 17 rethymectomies). One hundred and seventeen of these 620 patients were taking steroids preoperatively. The impact of steroids use on postoperative respiratory insufficiency (ventilator), need for tracheostomy, superficial wound dehiscence, superficial wound suppuration, sternal dehiscence and mortality was analyzed. RESULTS: The rate of morbidity and mortality in the steroids and the no-steroids groups included respiratory insufficiency necessitating the use of a ventilator in 46/503 patients from the no-steroids group (9.1%) and 8/117 patients from the steroids group (6.8%; P = 0.4260), need for tracheostomy in 14/503 patients from the no-steroids group (2.8%) and 3/117 patients from the steroids group (2.6%; P = 9054). Complications with wound healing (including sternal dehiscence, superficial wound dehiscence and superficial wound suppuration) occurred in 47/503 (9.3%) patients from the no steroids group and in 1/117 (0.9%) patient from the steroids group (P = 0.0023) The overall complications rate was 30.6% (154/503) in the no-steroids group and 11.1% (13/117) in the steroids group (P = 0.0001). Mortality rate in the no steroids group was 0.4% (2/503 patients) and none in the steroids group. Morbidity in patients with complete sternotomy was low, respiratory insufficiency in three patients (3.1%), and superficial wound suppuration in one patient (1.0%), no other morbidity and no mortality. Changing morbidity in periods 1973 1980, 1981-1990 and 1991-2002 is analyzed. CONCLUSIONS: Preoperative use of steroids for transsternal thymectomy in patients with myasthenia gravis had no negative impact on morbidity and mortality, conversely, the results in patients taking steroids were better, with significant difference of results regarding the overall complication rates and the overall complication rates of wound healing. PMID- 15296907 TI - Long-term survival and prognostic factors in thymic epithelial tumours. AB - OBJECTIVE: The aim of this study is to analyze long-term survival and the prognostic significance of some factors after surgical resection of thymic epithelial tumours. METHODS: We performed a retrospective analysis of clinical and histopathological data on 132 patients operated on for thymic tumours, from 1970 and 2001. Histologic diagnosis based on the new WHO classification system was made by a single pathologist. A univariate and multivariate analysis of prognostic factors predicting survival was carried out. RESULTS: There were: 108 complete resections (81.8%), 12 partial resections (9.1%) and 12 biopsies (9.1%). Overall 5, 10 and 15-year survival rate was 72, 61 and 52.5%, respectively. The Masaoka staging system showed 44 stage I, 18 stage II, 52 stage III and 18 stage IV. Histologic results were: 14 subtype A, 31 AB, 20 B1, 28 B2, 29 B3 and 10 C; the respective proportions of invasive tumour (stage II-IV) was 28.6, 58.1, 50, 75, 86.2 and 100%. There were 16 tumour recurrences (14.8%) of 108 radically resected thymomas, 10 were treated with radical re-resection. In univariate analysis, four prognostic factors were statistically significant: radical resection, Masaoka clinical staging, WHO histologic subtype and resectable tumour recurrence. In multivariate analysis, the independent factors predicting long term survival were WHO histology and Masaoka stage. CONCLUSIONS: The WHO histologic classification seems to be the most significant prognostic factor reflecting the invasiveness of the thymic tumour. Completeness of resection and Masaoka stage I and II assure a better survival. Unresectable recurrence of thymic tumour predicted a worse prognosis. PMID- 15296908 TI - Prognostic significance of nm23-H1 expression in esophageal squamous cell carcinoma. AB - OBJECTIVES: Tumor recurrence and metastasis are major causes of treatment failure in esophageal squamous cell carcinoma (ESCC). Recently, nm23, originally considered to be an anti-metastatic gene, has been reported to associate with various roles in different human cancers. We therefore investigated the clinical significance of nm23-H1 expression in ESCC. METHODS: Pathological sections were immunohistochemically stained with monoclonal antibody that was specific to nm23 H1. Expression of positive nm23-H1 staining was further confirmed by Western blot and reverse transcription-polymerase chain reaction (RT-PCR). The relationship between nm23-H1 expression and clinicopathological variables was examined by statistical analysis. Except for 11 (7%) surgical morality, the remaining 145 patients entered the prognostic analysis. The cisplatin-based chemotherapy was established for the patients with tumor stages at or beyond IIb, or with tumor recurrence. Survival difference between groups was compared by log rank test. RESULTS: Immunohistochemically, nm23-H1 expression was detected in 39.3% (57/145) of the pathological sections. It was positively correlated with tumor stage (P = 0.002), evident lymphovascular invasion (P < 0.001) and tumor recurrence (P = 0.02). Survival of nm23-H1 positive group was statistically superior to nm23-H1 negative group (P < 0.0001) By multivariate survival analysis, tumor stage, the number of lymph node metastasis and expression of nm23-H1 were the independent prognostic factors for ESCC patients. CONCLUSIONS: Our study demonstrated that nm23-H1 expression was associated with disease progression in ESCC. However, survival of nm23-H1 positive group was superior to nm23-H1 negative group. This paradoxical result could suppose that nm23-H1 expression might increase cisplatin chemosensitivity and hence improve survival. Screening for nm23-H1 expression in tumor cells may be a potential therapeutic strategy in ESCC patients. PMID- 15296909 TI - Correlation between telomerase expression and terminal restriction fragment length ratio in non-small cell lung cancer--an adjusted measurement and its clinical significance. AB - OBJECTIVE: To establish a new model for analyzing the correlation between the terminal restriction fragment (TRF) length and telomerase activity in non-small cell lung cancer (NSCLC) patients due to inconsistent results in previous reports. METHODS: Between January 1999 and December 1999, 79 NSCLC patients were studied. The activity of telomerase was measured by telomeric repeat amplification protocol, and the telomere restriction fragment (TRF) length was measured by Telomere Length Assay Kit and Southern blotting. The correlation between expression of telomerase activity and the TRF length ratio (TRFLR) using the tumor-to-normal TRFLR (t/n TRFLR) was examined. RESULTS: Positive telomerase activity was observed in 48 (60.8%) of the tumor tissue samples and in 5 (6.3%) of the normal tissue samples (P < 0.0001). The mean TRF lengths were 5.32+/-1.53 kb in normal tissue samples and 3.95+/-1.92 kb in tumor tissue samples, respectively (P = 0.0001). The 4-year cumulative survival rates of lower t/n TRFLR (< or = 75%) and higher t/n TRFLR (>75%) patients were 69.2 and 41.2%, respectively (P = 0.0227). Independent prognostic factors include t/n TRFLR (P = 0.014), T-status (P = 0.027), and N-status (P = 0.020) of the tumor. CONCLUSIONS: Our data suggest that there is a good correlation between the t/n TRFLR and expression of telomerase activity. A higher t/n TRFLR may indicate that the tumor regains its ability to repair the telomere lost and escapes the apoptosis scenario, which is subsequently reflected in poorer patient outcomes. PMID- 15296910 TI - Impact of pleural effusion pH on the efficacy of thoracoscopic mechanical pleurodesis in patients with breast carcinoma. AB - OBJECTIVE: A prospective randomised study was conducted to compare the efficacy of treating malignant pleural effusions (MPE) in patients with breast carcinoma by thoracoscopic mechanical pleurodesis (TMP) as a new palliative treatment and talc pleurodesis (TP) at various pleural fluid pH levels and to determine whether at low pH values, when the success of TP is reduced, TMP is more successful. METHODS: 87 female patients with breast carcinoma and a resulting MPE resistant to systemic therapy were divided into two groups: TMP and TP groups. In the TMP group 24 patients with pH levels above 7.3 and 21 patients with pH levels below 7.3 underwent thoracoscopic parietal and visceral pleural abrasion utilising general anaesthesia. In the TP group, 22 patients with pH levels above 7.3 and 20 patients with pH levels below 7.3 were administered 5 g of sterile talc, dissolved in 100 ml of physiological solution, via a chest tube, utilising local anaesthesia. Postoperative follow-up was performed to determine a possible recurrence of MPE with periodic radiographs, the duration of chest tube drainage and hospitalisation, occurrence of complications, and perioperative mortality. The following was used for statistical analysis: t-test for odd samples, chi2 test, logistic regression, and multiple linear regression. RESULTS: TMP and TP were equally successful (92 and 91%) in patients with pH levels above 7.3. Differences occurred in patients with pH below 7.3 (81 and 55%) (P = 0.07). The lowest pH value at which TMP proved successful was 7.06, while for TP this value was 7.25. In TMP group the average duration of chest tube drainage amounted to 3.8 days and hospitalisation to 5.5 days, while in TP group it was 5.6 and 7.5 days, respectively. Differences were statistically significant (P < 0.05). 16% of easily treatable complications and no case of perioperative mortality were identified in TMP group, while 26% of complications and four cases of perioperative mortality were noted in TP group. CONCLUSIONS: TMP is a safe palliative treatment for MPE in breast carcinoma, with a minimal number of complications and a short hospital stay; it is more successful than TP in patients with pH of MPE below 7.3. PMID- 15296911 TI - Giant lipoma of chest wall. PMID- 15296912 TI - Idiopathic left ventricular aneurysm. PMID- 15296913 TI - Three-dimensional demonstration of the Adamkiewicz artery and its collateral supply with contrast-enhanced magnetic resonance angiography. PMID- 15296914 TI - Instent restenosis after carotid stenting necessitating open carotid surgical repair. PMID- 15296915 TI - Left ventricular assist device (LVAD) enables survival during 7 h of sustained ventricular fibrillation. AB - We describe the case of a patient implanted with a DeBakey left ventricular assist device (LVAD) as bridge to transplant who survived 7 h of ventricular fibrillation. He was successfully converted into a stable sinus rhythm. PMID- 15296916 TI - Successful emergency surgery on triple-vessel spontaneous coronary artery dissection. AB - Although there is increasing number of reported spontaneous coronary artery dissection (SCAD) recently, this is the first successful operative case of myocardial revascularization on triple-vessel SCAD. Emergency coronary artery bypass grafting was performed on a 42-year-old female presented with acute myocardial infarction, who had failed thrombolysis with on-going angina and acute heart failure. PMID- 15296917 TI - Totally endoscopic robotic-guided pulmonary veins ablation: an alternative method for the treatment of atrial fibrillation. AB - In patients affected by isolated atrial fibrillation, epicardial pulmonary veins ablation can be performed with minimally invasive robotic-guided cardiac surgery techniques. This approach might become a feasible alternative to percutaneous transcatheter procedures. In this case report, we present a totally endoscopic robotic-guided pulmonary veins microwave ablation, on beating heart. A 64-year old male patient affected by paroxysmal atrial fibrillation was scheduled for an epicardial ablation procedure. Through three 1 cm-length port accesses, the "da Vinci" robotic system's camera and arms were introduced in the patient's chest. The pericardial reflections along the superior and inferior vena cava, as well as the transverse sinus, were dissected. Through an additional 0.5 cm-length port, a guidewire was advanced gradually across the transverse sinus, the diaphragmatic surface of the heart and the oblique sinus, finally surfacing outside the thoracic wall through the same port. Once tied to the microwave probe, the guidewire was pulled out carrying the probe inside the chest up to encircle the pulmonary veins. Once in place, a box lesion of the pulmonary veins was produced by microwave. At the 3-month follow up the patient is in sinus rhythm and so far did not longer experienced paroxysmal arrhythmic episodes. PMID- 15296918 TI - Fibrosing mediastinitis as a cause of superior vena cava syndrome. AB - Fibrosing mediastinitis is a rare, chronic inflammatory process that can cause superior vena cava syndrome, and can mimic malignancy. We present two cases of this disease where surgical resection was not possible and review the treatment options. PMID- 15296919 TI - Unilateral hyperhydrosis in Pourfour du Petit syndrome. AB - Upper limp hyperhydrosis is an idiopathic disease with bilateral involvement. However, Pourfour du Petit syndrome, the opposite of Horner syndrome, may result in unilateral upper limb hyperhydrosis. It occurs following hyperactivity of the sympathetic cervical chain as a consequence of irritation secondary to trauma. We report herein two cases with Pourfour du Petit syndrome showing unilateral upper limb hyperhydrosis. The patients presented with right-sided mydriasis and ipsilateral hemifacial hyperhydrosis. The onset of disease was followed by a trauma in both patients. They underwent upper thoracic sympathectomy with favorable outcome. A history of an antecedent trauma in patients with unilateral upper limb hyperhydrosis and anisocoria may imply a possible diagnosis of Pourfour du Petit syndrome. PMID- 15296920 TI - Upper trachea sleeve resection and anastomosis for invasive thyroid carcinoma. PMID- 15296921 TI - Cervical ultrasound in operable non-small cell lung cancer. PMID- 15296923 TI - Systolic ventricular filling. PMID- 15296925 TI - Novel biocatalysis by database mining. AB - Broad-based adoption of biocatalytic methods will require widely available database tools, analogous to previous efforts compiling information for the facilitation of chemical synthesis. The analog to chemical reagents are enzymes. The analog to chemical synthetic routes are metabolic pathways. The free on-line database BRENDA exemplifies efforts to compile relevant information on enzymes for biocatalytic purposes. Likewise, the University of Minnesota Biocatalysis/Biodegradation Database focuses on novel enzymes and metabolic pathways useful in environmental and industrial biotechnology. The development of biocatalytic protocols will be facilitated by the increasing availability of well curated database information on enzymatic enantioselectivity and capabilities for transforming disparate chemical functional groups. PMID- 15296926 TI - Prospecting for biocatalysts and drugs in the genomes of non-cultured microorganisms. AB - Modern biotechnology has a steadily increasing demand for vitamins, antibiotics and, in particular, novel biocatalysts for use in the production of flavors, agrochemicals, pharmaceuticals and high-value fine chemicals. Novel experimental approaches are being developed in attempts to identify such molecules. However, it is known that up to 99.8% of the microbes present in many environments are not readily culturable; hence, they cannot be exploited for biotechnology. The 'metagenome technology' offers a solution to this problem by developing culture independent methods to isolate, clone and express environmental DNA. So far, metagenome-based approaches have led to the isolation of many novel biocatalysts and a variety of other molecules with a high potential for downstream applications. PMID- 15296927 TI - Novel methods for directed evolution of enzymes: quality, not quantity. AB - In the past decade methods of directed molecular evolution have proven revolutionary in protein engineering. An increasing number of powerful new combinatorial techniques have joined rational design methods as effective tools for the manipulation and tailoring of biocatalysts. More and more, research in this maturing field is focusing on the quality and comprehensiveness of library construction and analysis. Additionally, in-depth studies have begun to highlight the underlying evolutionary mechanisms, limitations, and consequences of the various methodologies. Together, these investigations are creating a framework for future engineering projects. PMID- 15296928 TI - Dealing with complexity: evolutionary engineering and genome shuffling. AB - Comparative analysis of the growing number of microbial genome sequences has shown a high plasticity of genomes and several mechanisms for the adaptation of microbial cells to changing environmental conditions have been discovered. By contrast, the underlying metabolic networks of microorganisms are under strict control and relatively rigid, which poses a significant challenge for rational metabolic engineering approaches. Recursive shuffling of whole genomes has recently been demonstrated as an effective new evolutionary whole-cell engineering approach for the rapid improvement of industrially important microbial phenotypes. PMID- 15296929 TI - Ultra-high-throughput screening based on cell-surface display and fluorescence activated cell sorting for the identification of novel biocatalysts. AB - Enzyme libraries displayed on the surface of microbial cells or microbeads can be screened with fluorogenic substrates that provide a physical linkage of the reaction product to the corresponding enzyme. Libraries exceeding 10(9) different variants can be quantitatively analysed and screened by flow cytometry at a rate of 30 000 cells/second. The promise of screening methods based on fluorescence activated cell sorting for directed enzyme evolution is being realized and significantly improved enzymes have been reported recently. PMID- 15296930 TI - Detergent proteases. AB - Over the past 20 years, the development of subtilisins as typical detergent proteases has employed all the tools of enzyme technology, resulting in a constant flow of new and improved enzymes. The number of molecules identified and characterized, however, is in clear opposition to the number of molecules that are entering the market. Will the next-generation detergent proteases be based on new backbones different from subtilisins, or will the use of all available technologies (rational design, directed evolution and exploitation of natural diversity) yield improved subtilisins, ending the current era dominated by high alkaline subtilisins? These questions will have to be answered not only by the performance of the molecules themselves, but also by their yield in fermentation and their compatibility with existing production technologies. PMID- 15296931 TI - A new perspective on thiamine catalysis. AB - Our knowledge of thiamine-catalyzed ligase and lyase reactions has entered a new dimension. Significant achievements have been made in the field of enzymatic catalysis with the detection of hitherto unknown reaction types - extending the synthetic potential of known thiamine diphosphate (ThDP)-dependent enzymes - and the identification and characterization of new enzymes. As we learn more about ThDP-dependent enzymes, we find an ever-expanding range of reactions that they are able to catalyze and see increased amino acid sequence heterogeneity. By contrast, the three-dimensional structures of these enzymes, so far, seem to be highly similar. Non-enzymatic thiazolium and triazolium catalysts have also been developed, enhancing the scope of acyl anion chemistry. PMID- 15296932 TI - Prokaryotic expression of antibodies and affibodies. AB - Recent advances have been made in the development of systems for the display and expression of recombinant antibodies and affibodies in filamentous phages, Escherichia coli and other prokaryotic cells. Emphasis has been placed on improving phage and phagemid vectors, alternative systems for expression in different cellular compartments (e.g. the outer membrane, periplasm, cytoplasm and extracellular secretion) and novel multimerization systems for generating bivalent or multivalent binding molecules. PMID- 15296933 TI - Biocatalytic degradation of pollutants. AB - Microbial reactions play key roles in biocatalysis and biodegradation. The recent genome sequencing of environmentally relevant bacteria has revealed previously unsuspected metabolic potential that could be exploited for useful purposes. For example, oxygenases and other biodegradative enzymes are benign catalysts that can be used for the production of industrially useful compounds. In conjunction with their biodegradative capacities, bacterial chemotaxis towards pollutants might contribute to the ability of bacteria to compete with other organisms in the environment and to be efficient agents for bioremediation. In addition to the bacterial biomineralization of organic pollutants, certain bacteria are also capable of immobilizing toxic heavy metals in contaminated aquifers, further illustrating the potential of microorganisms for the removal of pollutants. PMID- 15296934 TI - From the editor's desk. PMID- 15296935 TI - A novel basal lamina matrix of the stratified epithelium of the ovarian follicle. AB - Basal laminas are important sheets of specialized extracellular matrix that underlie and surround groups of cells, such as epithelia or endothelia, enabling the cells to orientate their basal/apical polarity and creating a microenvironment for them. Basal laminas can also individually encapsulate whole cells, such as muscle cells, thereby forming a microenvironment but not polarizing the enclosed cells. Other mesenchymal or stromal cells exist with no basal lamina. In the course of studying the bovine follicular basal lamina which underlies the multilayered epithelium of the ovarian follicle, we identified a developmentally regulated novel extracellular matrix (which we call focimatrix for focal intra-epithelial matrix). Focimatrix is composed of basal lamina-like material deposited as plaques or aggregates between the multilayers of the epithelial granulosa cells. The focimatrix does not encapsulate individual or groups of cells and therefore does not form a microenvironment for them. Focimatrix contains collagen type IV subunits alpha1 and alpha2 (but not alpha3 alpha6), and laminin chains alpha1, beta2 and gamma1 (but not alpha2 or beta1), and nidogen-1 and perlecan (but not versican). The amount of focimatrix increases with increasing follicular size, and its appearance precedes the expression by granulosa cells of the enzymes for steroid hormone synthesis, cholesterol side chain cleavage cytochrome P450 (SCC) and 3beta-hydroxysteroid dehydrogenase (3beta-HSD), in the days preceding ovulation. The expression in granulosa cells of two components examined, nidogen-1 and perlecan, also increases substantially when follicles enlarge to a sufficient size capable of ovulating. Following ovulation the follicular basal lamina is degraded, and presumably focimatrix is too since it is not detected in corpora lutea that develop from the ovulating follicles. During this development the granulosa cells undergo an epithelial mesenchymal transition (EMT) into luteal cells following ovulation, and substantially increase their expression of steroidogenic enzymes in the process. During EMT epithelial cells lose polarity. Since focimatrix exists on more than one side of the granulosa cells, we propose that it disrupts the polarity induced by the follicular basal lamina in the lead up to ovulation. Hence focimatrix maybe a key part of the follicular/luteal EMT. PMID- 15296936 TI - ADAMTS-8 exhibits aggrecanase activity and is expressed in human articular cartilage. AB - Members of the ADAMTS (a disintegrin and metalloprotease with thrombospondin motifs) family share common structural features including a disintegrin domain, a zinc metalloprotease domain, and at least one thrombospondin motif. Aberrant expression of several of these proteins has led to an understanding of their role in human disease; however, a link to function for many has not yet been made. One such uncharacterized family member, ADAMTS-8, shares significant protein sequence homology with a subgroup of ADAMTSs that includes ADAMTS-1, ADAMTS-4, ADAMTS-5, and ADAMTS-15. Each of these proteases has been shown to cleave 'aggrecanase susceptible' site(s) within the extracellular matrix (ECM) proteoglycan aggrecan, and ADAMTS-4 and ADAMTS-5 have been postulated to play a role in the depletion of articular cartilage in osteoarthritic disease. Based on sequence relationships, in the present study we examined the ability of ADAMTS-8 to exhibit 'aggrecanase' activity. A neoepitope monoclonal antibody (MAb; AGG-C1; anti-NITEGE373) was developed and used to demonstrate the ability of ADAMTS-8 to cleave aggrecan at the aggrecanase-susceptible Glu373-Ala374 peptide bond. In addition, expression analyses demonstrated the presence of ADAMTS-8 mRNA transcripts in normal and osteoarthritic human cartilage. PMID- 15296937 TI - Fibroblast growth factor-2 in serum-free medium is a potent mitogen and reduces dedifferentiation of human ear chondrocytes in monolayer culture. AB - The loss of the differentiated phenotype (dedifferentiation) during the expansion culture of donor chondrocytes remains a large problem in cartilage tissue engineering. Dedifferentiated chondrocytes produce other matrix components and therefore the tissue produced will be of less suitable quality. Previously, the addition of fibroblast growth factor-2 (FGF2) to a serum-containing medium (SCM) during expansion culture was shown to have positive effects on the phenotype of articular chondrocytes. In the present study, we focused on a more defined, serum free medium (SFM), to expand chondrocytes in monolayer culture for the purpose of cartilage tissue engineering. Adult human ear chondrocytes were expanded in serum free medium supplemented with 100 ng/ml FGF2. Expansion culture in a conventional serum-containing medium (10% FCS) served as control. The cell yield during expansion culture in serum-free medium with FGF2 was significantly higher compared to serum-containing medium. In addition, chondrocytes expanded in the serum-free medium with FGF2 expressed a more differentiated phenotype at the end of monolayer culture, as indicated by higher gene expression ratios of collagen type II to collagen type I and aggrecan to versican. Also, a higher gene expression of Sox9 was found. Next, suspension in alginate and subsequent culture in vitro or subcutaneous implantation in nude mice was used to evaluate the capacity of the chondrocytes, expanded in either medium, to re-express the differentiated phenotype (redifferentiation) and to form cartilage. The observed beneficial effects of the serum-free medium with FGF2 on the chondrocyte phenotype at the end of monolayer culture were sustained on both transcriptional and extracellular level throughout both redifferentiation methods. PMID- 15296938 TI - Wound splinting modulates granulation tissue proliferation. AB - Attachment of the extracellular matrix to a substratum is important for fibroblast survival and proliferation in three-dimensional in vitro culture systems. We hypothesized that wound matrix attachment in a wound splinting model would modulate wound cell proliferation in vivo. Male rats were excisionally wounded on the dorsum, and a splint was sutured to the wound edge. In one experiment (N = 12), 6 rats were desplinted on day 5, and then all were sacrificed 24 h later, 6 h after 5-bromo-2'-deoxyuridine (BrdU) injection. In the second experiment (N = 18), 6 rats each were desplinted, desplinted with wound edge release, or not disturbed, followed by BrdU injection and sacrifice 24 h later. BrdU-labeled nuclei were quantified on frozen sections of granulation tissue, cut at three different levels. In the first experiment, the percentage of BrdU-positive nuclei per high power field (hpf) in the splinted vs. desplinted animals was 6.15 +/- 2.45 (S.D.) vs. 3.03 +/- 1.58%* p<0.001, ANOVA. In the second experiment, the number of BrdU-positive per hpf was 33.1 +/- 17.4 vs. 14.5 +/- 17.1 vs. 10.2 +/- 9.1* (splinted vs. desplinted vs. desplinted/released); *p<0.001 [analysis of variance (ANOVA)]. Removal of the wound splint decreased the rate of BrdU-labeled cells in the granulation tissue by approximately 50%; complete disruption of wound matrix attachment may have decreased this rate even further. Wound cell proliferation is modulated by lateral attachment of the wound matrix. PMID- 15296939 TI - Increased formation of pyridinoline cross-links due to higher telopeptide lysyl hydroxylase levels is a general fibrotic phenomenon. AB - Fibrosis is characterized by an excessive accumulation of collagen which contains increased levels of pyridinoline cross-links. The occurrence of pyridinolines in the matrix is an important criterion in assessing the irreversibility of fibrosis, which suggests that collagen containing pyridinoline cross-links significantly contributes to the unwanted collagen accumulation. Pyridinoline cross-links are derived from hydroxylated lysine residues located within the collagen telopeptides (hydroxyallysine pathway). Here, we have investigated whether the increase in hydroxyallysine-derived cross-links in fibrotic conditions can be ascribed to an increased expression of one of the lysyl hydroxylases (LH1, LH2 with its splice variants LH2a and LH2b, or LH3) and/or to an increased expression of lysyl oxidase (LOX). In fibroblast cultures of hypertrophic scars, keloid and palmar fascia of Dupuytren's patients, as well as in activated hepatic stellate cells, increased levels of LH2b mRNA expression were observed. Only minor amounts of LH2a were present. In addition, no consistent increase in the mRNA expression levels of LH1, LH3 and LOX could be detected, suggesting that LH2b is responsible for the overhydroxylation of the collagen telopeptides and the concomitant formation of pyridinolines as found in the collagen matrix deposited in long-term cultures by the same fibrotic cells. This is consistent with our previous observation that LH2b is a telopeptide lysyl hydroxylase. We conclude that the increased expression of LH2b, leading to the increased formation of pyridinoline cross-links, is present in a wide variety of fibrotic disorders and thus represents a general fibrotic phenomenon. PMID- 15296940 TI - Enhanced procollagen processing in skeletal muscle after a single bout of eccentric loading in humans. AB - Increases in procollagen processing within skeletal muscle have previously been reported in small animal models only. While indirect measurements in humans have suggested an increase procollagen processing, no intra-skeletal muscle measurements have confirmed these findings. In this study, eight young healthy male subjects performed a single bout of unaccustomed high intensity eccentric exercise on one leg, with the contralateral leg being the control. A significant increase in the muscle interstitial concentration of the N-terminal propeptide of procollagen type I (PINP) was observed (day 0: 1.96 +/- 0.44 ng ml(-1), day 2: 1.94 +/- 0.32 ng ml(-1), day 4: 3.90 +/- 1.03 ng ml(-1), day 8: 7.23 +/- 2.34 ng ml(-1)*, *P < 0.05 vs. basal and control) with no change being noted in the control leg. By day 2 post-exercise, an increase in the histological immunoreactivity of PINP and the N-terminal propeptide of procollagen type III (PIIINP) was also shown in the exercising leg only. Further, from day 2 post exercise, immunoreactivity for tenascin C and reactive macrophages (CD68+ cells) was seen within the perimysial and endomysial connective tissue of the exercising leg only, indicating a high mechanical load and inflammation. This study shows that following a single bout of high intensity eccentric exercise there is an increase in procollagen processing within skeletal muscle in humans. PMID- 15296942 TI - Perlecan and its immunoglobulin like domain IV are abundant in vitreous and serum of the chick embryo. AB - Perlecan is a highly conserved heparan sulfate proteoglycan in cartilage and basement membranes. We identified chick perlecan and a 90 KD perlecan fragment in vivo using a newly generated monoclonal antibody. Chick perlecan is, like its human and mouse homologue, a hybrid heparan sulfate/chondroitin sulfate proteoglycan with a core protein of 400 KD. Analysis of the 90 KD fragment by Matrix Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry (MALDI-TOF MS) and Capillary LC nano Electrospray Ionization tandem MS (LC nano ESI MS/MS) showed that it belonged to domain IV of the perlecan core protein. We found that full-length perlecan and its domain IV fragment are abundant in embryonic vitreous body and serum. Their expression in vitreous and serum is greatly down-regulated shortly after hatching of the chick. We speculate that the abundance of perlecan in the embryonic circulation and vitreous reflects the ongoing formation of new BMs in the expanding vascular system and the growing retina. In addition, we found that perlecan as a substrate does not support, rather inhibits neurite outgrowth. PMID- 15296943 TI - Expression of collagen XVIII and MMP-20 in developing teeth and odontogenic tumors. AB - Collagen XVIII is a basement membrane (BM) component, whereas MMP-20 (enamelysin) is a matrix metalloproteinase predominantly expressed in teeth. Since MMP-20 was found to degrade collagen XVIII, we studied the co-expression of these proteins in dental tissues. Collagen XVIII surrounded the developing tooth during early and late bell stages and was also present in developing enamel. Western blotting indicated that developing enamel contains collagen XVIII N-terminal fragments of the frizzled variant. Enamelysin was co-localized with collagen XVIII in the developing enamel matrix and stratum intermedium. Electron microscope analysis showed that total mineral, calcium and phosphorus contents of enamel were slightly increased in collagen XVIII null mice but the analysis revealed no visible defects in the enamel or dentin structures. In odontogenic tumors MMP-20 and collagen XVIII were co-localized in the enamel-like tumor matrix. Our results show that collagen XVIII is present in developing teeth, but its absence seems not to be critical for the development of the teeth. PMID- 15296944 TI - Inhibition of interleukin-1beta-stimulated collagenase and stromelysin expression in human tendon fibroblasts by epigallocatechin gallate ester. AB - The medicinal benefits of green tea (Camellia sinensis) consumption have been attributed to bioavailable polyphenols, notably epigallocatechin gallate (EGCG). We have assessed the effects of EGCG and its non-esterified counterpart EGC on the expression of the collagenases, matrix metalloproteinases (MMP)-1 and -13, and the stromelysin, MMP-3, in human tendon-derived fibroblasts. Interleukin (IL) 1beta increased MMP-1, -3 and -13 mRNA and output at least 30-fold. EGCG reduced this stimulation, by 20-30% at 2.5 microM and more than 80% at 25 microM, and had a smaller effect on MMP-2 mRNA expression, which was not stimulated by IL-1beta. In all experiments EGCG was at least 10-fold more potent than EGC. EGCG reduced the stimulation of p54 JNK/SAPK phosphorylation by IL-1beta but did not affect p38 MAPK phosphorylation, the degradation of IkappaB or the activating phosphorylation of NFkappaB. We conclude that EGCG reduces the IL-1-stimulated expression of both collagenase and stromelysin mRNA species, an effect which may be mediated by inhibition of the JNK/SAPK pathway. Taken together with previous reports of EGCG effects on the expression and/or activity of gelatinases and aggrecanases, our results underline the importance of extracellular matrix breakdown as a potential target for the actions of green tea polyphenols. PMID- 15296945 TI - Contributions of the MMP-2 collagen binding domain to gelatin cleavage. Substrate binding via the collagen binding domain is required for hydrolysis of gelatin but not short peptides. AB - Two matrix metalloproteinases, MMP-2 and MMP-9, contain each three fibronectin type II-like modules, which form their collagen binding domains (CBDs). The contributions of CBD substrate interactions to the catalytic activities of these gelatinases have attracted special interest. Recombinant (r) CBDs retain collagen binding properties and deletions of CBDs in these MMPs reduce activities on collagen and elastin. We have characterized further the requirement of the CBD for MMP-2 cleavage of gelatin. The analyses used intact rMMP-2 and rCBD to eliminate any confounding effects that might result from structural perturbations in rMMP-2 induced by deletion of the approximately 20 kDa internal CBD. In protein-protein binding assays, 2% DMSO disrupted gelatin interactions of both rCBD and rMMP-2. At this concentration, DMSO also reduced the gelatinolytic activity by approximately 70%, pointing to a central role of CBD-substrate interactions during MMP-2 cleavage of gelatin. Subsequently, soluble rCBD was determined to competitively inhibit gelatin binding of unmodified rMMP-2 to gelatin by 73% and to reduce the MMP-2 degradation of gelatin by 70-80%. The residual gelatin cleavage that was not inhibited even by molar excess rCBD could be accounted for by degradation of short substrate molecules. Indeed, rCBD inhibited rMMP-2 cleavage of an 11 amino acid collagen-like peptide substrate (NFF-1) by less than 10%. These observations were confirmed with enzyme extracts from experimental tumors in mice. In the presence of rCBD, approximately 65% of the MMP-derived gelatinolytic activity was eliminated. Together, these results demonstrate that the CBD is absolutely required for MMP-2 cleavage of full-length collagen alpha-chains, but not for short protein fragments such as those generated by hydrolysis of gelatin. PMID- 15296946 TI - Reticulated hyaluronan hydrogels: a model for examining cancer cell invasion in 3D. AB - The extracellular polysaccharide hyaluronan (HA) controls cell migration, differentiation and proliferation, and contributes to the invasiveness of human cancers. The roles of HA cell surface receptors and hyaluronidases (HAses) in this process are still controversial. In order to investigate their involvement in cancer pathogenesis, we developed a reticulated HA hydrogel, a three dimensional matrix in which cells can invade and grow. We have studied thirteen cell lines, from primary tumors or metastases, that migrated into the HA hydrogel and proliferated giving rise to clusters and colonies. The number of colonies, which reflects tumor cell invasiveness, ranged from 7 to 193 after 5 days of culture. Invasion was dependent on the production of HAse as well as other factors. Optimal colonization occurred when cells released HAse, lacked HA binding sites and did not secrete HA. Moreover, we describe for the first time a HAse activity at physiological pH that may be responding to the confinement of the enzyme in a three-dimensional structure. We show here that this reticulated matrix provides a three-dimensional model for investigating mechanisms involved in malignant invasion. PMID- 15296947 TI - Mice lacking the extracellular matrix adaptor protein matrilin-2 develop without obvious abnormalities. AB - Matrilins are putative adaptor proteins of the extracellular matrix (ECM) which can form both collagen-dependent and collagen-independent filamentous networks. While all known matrilins (matrilin-1, -2, -3, and -4) are expressed in cartilage, only matrilin-2 and matrilin-4 are abundant in non-skeletal tissues. To clarify the biological role of matrilin-2, we have developed a matrilin-2 deficient mouse strain. Matrilin-2 null mice show no gross abnormalities during embryonic or adult development, are fertile, and have a normal lifespan. Histological and ultrastructural analyses indicate apparently normal structure of all organs and tissues where matrilin-2 is expressed. Although matrilin-2 co localizes with matrilin-4 in many tissues, Northern hybridization, semiquantitative RT-PCR, immunohistochemistry and biochemical analysis reveal no significant alteration in the steady-state level of matrilin-4 expression in homozygous mutant mice. Immunostaining of wild-type and mutant skin samples indicate no detectable differences in the expression and deposition of matrilin-2 binding partners including collagen I, laminin-nidogen complexes, fibrillin-2 and fibronectin. In addition, electron microscopy reveals an intact basement membrane at the epidermal-dermal junction and normal organization of the dermal collagen fibrils in mutant skin. These data suggest that either matrilin-2 and matrilin-2 mediated matrix-matrix interactions are dispensable for proper ECM assembly and function, or that they are efficiently compensated by other matrix components including wild-type levels of matrilin-4. PMID- 15296948 TI - Second-generation biopharmaceuticals. AB - The majority of first generation biopharmaceuticals are unengineered murine monoclonal antibodies or simple replacement proteins displaying an identical amino acid sequence to a native human protein. While some such products continue to be approved, an increasing number of modern biopharmaceuticals are engineered, second-generation products. Engineering can entail alteration of amino acid sequence, alteration of the glycocomponent of a glycosylated protein, or the covalent attachment of chemical moieties such as polyethylene glycol. Engineering has been applied in order to alter a protein's immunological or pharmacokinetic profile, or in order to generate novel fusion products. Better understanding of the links between protein structure and function will underpin the development of an increasing number of engineered biopharmaceuticals in the future. PMID- 15296949 TI - Localized delivery of growth factors for bone repair. AB - Delivery of growth factors for tissue (e.g. bone, cartilage) or cell repair (e.g. nerves) is about to gain important potential as a future therapeutic tool. Depending on the targeted cell type and its state of differentiation, growth factors can activate or regulate a variety of cellular functions. Therefore, strictly localized delivery regimens at well-defined kinetics appear to be logical prerequisites to assure safe and efficacious therapeutic use of such factors and avoid unwanted side effects and toxicity, a major hurdle in the clinical development of growth factor therapies so far. This review summarizes various approaches for localized growth factor delivery as focused on bone repair. Similar considerations may apply to other growth factors and therapeutic indications. Considering the vast number of preclinical studies reported in the area of growth factor-assisted bone repair, it surprises though that only two medical products for bone repair have so far been commercialized, both consisting of a collagen matrix impregnated with a bone morphogenetic protein. The marked diversity of the reported growth factors, delivery concepts and not yet standardized animal models adds to the complexity to learn from past preclinical studies presented in the literature. Nonetheless, it is now firmly established from the available information that the type, dose and delivery kinetics of growth factors all play a decisive role for the therapeutic success of any such approach. Very likely, all of these parameters have to be adapted and optimized for each animal model or clinical case. In the future, systems for localized growth factor delivery thus need to be designed in such a way that their modular components are readily adaptable to the individual pathology. To make such customized systems feasible, close cooperative networks of biomedical and biomaterials engineers, pharmaceutical scientists, chemists, biologists and clinicians need to be established. PMID- 15296950 TI - Chances and pitfalls of cell penetrating peptides for cellular drug delivery. AB - Over the past decade, several classes and/or prototypes of cell penetrating peptides (CPP) have been identified and investigated in multiple aspects. CPP represent peptides, which show the ability to cross the plasma membrane of mammalian cells, and may thus give rise to the intracellular delivery of problematic therapeutic cargos, such as peptides, proteins, oligonucleotides, plasmids and even nanometer-sized particles, which otherwise cannot cross the plasma membrane. Most of the currently recognized CPP are of cationic nature and derived from viral, insect or mammalian proteins endowed with membrane translocation properties. The exact mechanisms underlying the translocation of CPP across the cellular membrane are still poorly understood. However, several similarities in translocation can be found. Early studies on CPP translocation mechanisms tended to suggest that the internalization of these peptides was neither significantly inhibited by low temperature, depletion of the cellular adenosine triphosphate (ATP) pool, nor by inhibitors of endocytosis. Moreover, chemical modification of the peptide sequence, such as the synthesis of retro-, enantio- or retroenantio-analogs, appeared not to affect the internalization properties. Therefore, translocation was concluded to result from direct, physical transfer through the lipid bilayer of the cell membrane. Later studies, however, showed convincing evidence for the involvement of endocytosis as the dominating mechanism for cellular internalization. In addition to describing the general properties of the commonly recognized classes of CPP, in this review we will also point out some limitations and typical pitfalls of CPP as carriers for therapeutics. In particular we will comment on emerging discrepancies with the current dogma, on cell-to-cell variability, biological barrier permeability, metabolic fate, toxicity and immunogenicity of CPP. PMID- 15296951 TI - Trimethylated chitosan as polymeric absorption enhancer for improved peroral delivery of peptide drugs. AB - The absorption enhancing effects of chitosan and its derivatives have been intensively studied in recent years. It has been shown that these compounds are potent absorption enhancers. Chitosan is only soluble in acidic environments and is therefore incapable of enhancing absorption in the small intestine, the main absorption area in the gastrointestinal tract. Special emphasis has been placed on the absorption enhancing properties of N-trimethyl chitosan chloride (TMC), a partially quaternised derivative of chitosan, due to its solubility in neutral and basic environments. TMC is prepared by the reductive methylation of chitosan. The degree of quaternisation can be altered by increasing the number of reaction steps or by increasing the reaction time. Although the molecular weight of the polymer increases with addition of the methyl groups, a net decrease in the molecular weight is observed due to a decrease in the chain length of the polymer. TMC, like chitosan, possesses mucoadhesive properties. In vitro studies performed on Caco-2 cell monolayers showed a pronounced reduction in the transepithelial electrical resistance (TEER). TMC is also able to increase the permeation of hydrophilic compounds such as [14C]-mannitol and [14C] polyethylene glycol 4000 ([14C] PEG 4000, MW4000) across the cell monolayers. It was also shown that the degree of quaternisation of the polymer plays an important role on its absorption enhancing properties, especially in neutral environments where chitosan is ineffective as an absorption enhancer. The reduction in TEER is an indication of the opening of the tight junctions located between epithelial cells. Opening of the tight junctions will result in enhancement of absorption via the paracellular route. Confocal laser scanning microscopy confirmed transport of large hydrophilic compounds via the paracellular route as well as the mechanism of action of the polymer in which redistribution of the cytoskeletal F-actin is provoked, which leads to the opening of the tight junctions. Various in vivo studies in different animal models confirmed the ability of TMC to increase the absorption of the peptide drugs buserelin and octreotide after intraduodenal or -jejunal administration. However, TMC has always been administered as a solution in these studies. The impracticality of administering a solution, as well as the fact that most peptides are unstable in the presence of water, have led to the need for a solid oral dosage form with which TMC can be administered together with peptide drugs. Recent studies have focused on the development and in vivo evaluation of solid oral dosage forms. PMID- 15296952 TI - Drug delivery of oligonucleotides by peptides. AB - Oligonucleotides are promising tools for in vitro studies where specific downregulation of proteins is required. In addition, antisense oligonucleotides have been studied in vivo and have entered clinical trials as new chemical entities with various therapeutic targets such as antiviral drugs or for tumour treatments. The formulation of these substances were widely studied in the past. With this review we will focus on peptides used as drug delivery vehicles for oligonucleotides. Different strategies are summarised. Cationically charged peptides from different origins were used e.g. as cellular penetration enhancers or nuclear localisation tool. Examples are given for Poly-L-lysine alone or in combination with receptor specific targeting ligands such as asialoglycoprotein, galactose, growth factors or transferrin. Another large group of peptides are those with membrane translocating properties. Fusogenic peptides rich in lysine or arginine are reviewed. They have been used for DNA complexation and condensation to form transport vehicles. Some of them, additionally, have so called nuclear localisation properties. Here, DNA sequences, which facilitate intracellular trafficking of macromolecules to the nucleus were explored. Summarizing the present literature, peptides are interesting pharmaceutical excipients and it seems to be feasible to combine the specific properties of peptides to improve drug delivery devices for oligonucleotides in the future. PMID- 15296953 TI - Thiomers: potential excipients for non-invasive peptide delivery systems. AB - In recent years thiolated polymers or so-called thiomers have appeared as a promising alternative in the arena of non-invasive peptide delivery. Thiomers are generated by the immobilisation of thiol-bearing ligands to mucoadhesive polymeric excipients. By formation of disulfide bonds with mucus glycoproteins, the mucoadhesive properties of these polymers are improved up to 130-fold. Due to formation of inter- and intramolecular disulfide bonds within the thiomer itself, dosage forms such as tablets or microparticles display strong cohesive properties resulting in comparatively higher stability, prolonged disintegration times and a more controlled release of the embedded peptide drug. The permeation of peptide drugs through mucosa can be improved by the use of thiolated polymers. Additionally some thiomers exhibit improved inhibitory properties towards peptidases. The efficacy of thiomers in non-invasive peptide delivery could be demonstrated by various in vivo studies. Tablets comprising a thiomer and pegylated insulin, for instance, resulted in a pharmacological efficacy of 7% after oral application to diabetic mice. Furthermore, a pharmacological efficacy of 1.3% was achieved in rats by oral administration of calcitonin tablets comprising a thiomer. Human growth hormone in a thiomer-gel was applied nasally to rats and led to a bioavailability of 2.75%. In all these studies, formulations comprising the corresponding unmodified polymer had only a marginal or no effect. According to these results drug carrier systems based on thiomers seem to be a promising tool for non-invasive peptide drug delivery. PMID- 15296954 TI - Classification of orally administered drugs on the World Health Organization Model list of Essential Medicines according to the biopharmaceutics classification system. AB - Since its inception in 1995, the biopharmaceutical classification system (BCS) has become an increasingly important tool for regulation of drug products world wide. Until now, application of the BCS has been partially hindered by the lack of a freely available and accurate database summarising solubility and permeability characteristics of drug substances. In this report, orally administered drugs on the Model list of Essential Medicines of the World Health Organization (WHO) are assigned BCS classifications on the basis of data available in the public domain. Of the 130 orally administered drugs on the WHO list, 61 could be classified with certainty. Twenty-one (84%) of these belong to class I (highly soluble, highly permeable), 10 (17%) to class II (poorly soluble, highly permeable), 24 (39%) to class III (highly soluble, poorly permeable) and 6 (10%) to class IV (poorly soluble, poorly permeable). A further 28 drugs could be provisionally assigned, while for 41 drugs insufficient or conflicting data precluded assignment to a specific BCS class. A total of 32 class I drugs (either certain or provisional classification) were identified. These drugs can be further considered for biowaiver status (drug product approval based on dissolution tests rather than bioequivalence studies in humans). PMID- 15296955 TI - The use of polymers for dermal and transdermal delivery. AB - The use of polymers for skin preparations is manifold. Requirements of such polymers are dependent on the formulation types. The most applied polymers on skin belong to various classes, for example to cellulose derivatives, chitosan, carageenan, polyacrylates, polyvinylalcohol, polyvinylpyrrolidone and silicones. They are gelating agents, matrices in patches and wound dressings, anti-nucleants and penetration enhancers. Correlations between commercially available products and results of new scientific investigations are often difficult or not possible, because of the lack of comparative data especially for transdermal patches. Finally, two promising future trends of polymeric systems, gene delivery and tissue engineering, are discussed. PMID- 15296956 TI - Skin deep. AB - Over the past 30 or so years there has been a considerable advance in our knowledge of the mechanisms of skin permeation. This has largely been brought about by the development of sophisticated biophysical techniques and increased computing powers. The advanced technology has clearly provided indications, at a molecular level, about routes of permeation and how the barrier function can be modulated by excipients with which actives are formulated. This publication reviews some of the advances that have been made and mathematical models that have been constructed to predict percutaneous penetration and transdermal delivery. The models also indicate the various enhancement strategies that can be used in dermal penetration. In the past, it has been difficult to identify precise mechanisms of action of the different classes of enhancer but a combination of appropriate biophysical techniques, mathematical modelling and chemometric analysis can help identify the contributing processes. The models can also be used to indicate rate control in transdermal delivery, whether it is in the applied delivery device or in the skin. PMID- 15296957 TI - Visualization of skin penetration using confocal laser scanning microscopy. AB - The use of skin as an alternative route for administering systemically active drugs has attracted considerable interest in recent years. However, the skin provides an excellent barrier, which limits the number of drug molecules suitable for transdermal delivery. Thus, in order to improve cutaneous delivery, it is necessary to adopt an enhancement method, either (i) passively using novel formulations, e.g. microemulsions, liposomes, and colloidal polymeric suspensions, or more conventional skin permeation enhancers, or (ii) with a physical approach, such as, iontophoresis, sonophoresis or electroporation. Although there has been much progress, the precise modes of action of the different techniques used are far from well-understood. The objective of this review, therefore, is to evaluate how confocal laser scanning microscopy may contribute to the determination of the mechanisms of diverse skin penetration enhancement strategies. PMID- 15296958 TI - Roll compaction/dry granulation: pharmaceutical applications. AB - Roll compaction/dry granulation (RCDG) is an agglomeration process of growing importance. New machine generations and improvements in instrumentation and process control have resulted in an increasing number of pharmaceutical applications of RCDG. This literature review illustrates the progress and the use of RCDG in the production of directly compressible excipients, the compaction of drugs and drug formulations, the granulation of inorganic materials, the granulation of dry herbal material and the production of immediate/sustained release formulations. PMID- 15296959 TI - Polysaccharide-decorated nanoparticles. AB - Surface modified colloidal carriers such as nanoparticles are able to modulate the biodistribution of the loaded drug when given intravenously, but also to control the absorption of drugs administered by other routes. This review presents the different strategies to coat the surface of polymeric as well as inorganic nanoparticles with polysaccharides. Various physicochemical and biological methods have been described to demonstrate such surface modification. The medical applications, mainly in imaging cancer, of polysaccharide-coated nanoparticles are presented, including their abilities to increase the blood circulation time and to target specific tumoral tissues. It has been shown that these coatings allow also to improve drug absorption via nasal or ocular pathways, due the mucoadhesive and/or permeability enhancer properties of the polysaccharides. Finally, the ability of polysaccharide-coated nanoparticles to deliver DNA or oligonucleotides will be discussed. PMID- 15296960 TI - Physicochemical characterization of colloidal drug delivery systems such as reverse micelles, vesicles, liquid crystals and nanoparticles for topical administration. AB - Topical administration of cosmetics and pharmaceuticals involves a variety of different formulations of which colloidal drug carrier systems are currently of particular interest. After a short introduction of reverse micellar solutions, liquid crystals, vesicles and nanoparticles, appropriate methods of physicochemical characterization are introduced including X-ray diffraction, laser light scattering, electron microscopy, and differential scanning calorimetry. Emphasis is laid on topical applications of the colloidal drug delivery systems (DDS) covered, with the main objective of both sustained drug release and improved stability of DDS. PMID- 15296961 TI - The potential of lipid emulsion for ocular delivery of lipophilic drugs. AB - For nearly a decade, oil-in-water lipid emulsions containing either anionic or cationic droplets have been recognized as an interesting and promising ocular topical delivery vehicle for lipophilic drugs. The aim of this review is to present the potential of lipid emulsions for ocular delivery of lipophilic drugs. The review covers an update on the state of the art of incorporating the lipophilic drugs, a brief description concerning the components and the classification of lipid emulsions. The ocular fate following topical instillation, safety evaluation experiments and the applications of lipid emulsions are thoroughly discussed. PMID- 15296962 TI - The use of asymmetrical flow field-flow fractionation in pharmaceutics and biopharmaceutics. AB - Field-flow fractionation (FFF) is a family of flexible analytical fractionating techniques which have the advantage that the separation of analytes is achieved, solely through the interaction of the sample with an external, perpendicular physical field, rather than by the interaction with a stationary phase. The rapid progress in pharmaceutical biotechnology goes along with an increasing demand in potent, high-efficient analytical methods. Thus, FFF techniques are gaining increasing attention for their ability to separate and characterize populations of polymers, colloids and particles of up to about 100 microm in size. It is the intention of this review to provide an overview on common FFF techniques, to summarize inherent advantages and limitations and to introduce both established and challenging applications in the (bio)pharmaceutical field. Thereby, asymmetrical flow FFF is addressed predominantly, since it is the most versatile applicable FFF technique. PMID- 15296963 TI - Biomimetic polymers in pharmaceutical and biomedical sciences. AB - This review describes recent developments in the emerging field of biomimetic polymeric biomaterials, which signal to cells via biologically active entities. The described biological effects are, in contrast to many other known interactions, receptor mediated and therefore very specific for certain cell types. As an introduction into this field, first some biological principles are illustrated such as cell attachment, cytokine signaling and endocytosis, which are some of the mechanisms used to control cells with biomimetic polymers. The next topics are then the basic design rules for the creation of biomimetic materials. Here, the major emphasis is on polymers that are assembled in separate building blocks, meaning that the biologically active entity is attached to the polymer in a separate chemical reaction. In that respect, first individual chemical standard reactions that may be used for this step are briefly reviewed. In the following chapter, the emphasis is on polymer types that have been used for the development of several biomimetic materials. There is, thereby, a delineation made between materials that are processed to devices exceeding cellular dimensions and materials predominantly used for the assembly of nanostructures. Finally, we give a few current examples for applications in which biomimetic polymers have been applied to achieve a better biomaterial performance. PMID- 15296964 TI - In situ-forming hydrogels--review of temperature-sensitive systems. AB - In the past few years, an increasing number of in situ-forming systems have been reported in the literature for various biomedical applications, including drug delivery, cell encapsulation, and tissue repair. There are several possible mechanisms that lead to in situ gel formation: solvent exchange, UV-irradiation, ionic cross-linkage, pH change, and temperature modulation. The thermosensitive approach can be advantageous for particular applications as it does not require organic solvents, co-polymerization agents, or an externally applied trigger for gelation. In the last 2 decades, several thermosensitive formulations have been proposed. This manuscript focuses on aqueous polymeric solutions that form implants in situ in response to temperature change, generally from ambient to body temperature. It mainly reviews the characterization and use of polysaccharides, N-isopropylacrylamide copolymers, poly(ethylene oxide-b propylene oxide-b-ethylene oxide) (poloxamer) and its copolymers, poly(ethylene oxide)/(D,L-lactic acid-co-glycolic acid) copolymers, and thermosensitive liposome-based systems. PMID- 15296965 TI - Benefits of die-wall instrumentation for research and development in tabletting. AB - Instrumented presses used in tabletting research and development are normally equipped to measure punch force and displacement. Die-wall monitoring is rare, probably because instrumentation and calibration are quite difficult. The authors critically examine the tenets of radial pressure measurement in compression physics. The theoretical background concerning axial to radial stress transmission during the different phases of the compression cycle is presented. The literature reporting on the use of radial stress measurement to assess the self-lubricating properties of materials or the effect of lubricants is reviewed. Examples of interpretation of radial pressure cycles to define the basic material behaviour are given. The influence of particle size and shape as well as that of process and formulation variables on die-wall response are also discussed. Substantial inconsistencies can be seen in the literature with respect to the interpretation of experimental data, often because of the poor reliability of results and mostly because powders are essentially not solid, isotropic bodies. There is also a distinct lack of complementary tabletting parameters that would help understanding their comparative benefits. For this reason, original data on 13 model compounds are presented together with a classification of the materials encountered in pharmaceutical tabletting, based on selected parameters. In conclusion, none of the determined parameters, including those derived from radial pressure measurement, is able, alone, to predict the material behaviour under compression. Although die-wall instrumentation contributes little to the development of improved tablet formulations, it is valuable for characterising the mechanical properties of the materials. This is particularly advantageous given that the mechanical properties account for variations in tabletting performance to a much greater extent than the magnitude of the interparticulate attractions. Nevertheless, because of the peculiar nature of powders compared to solid, isotropic bodies, there is a need to develop new models for analysing their behaviour and to put more emphasis on examination of time-dependent deformation in the later stage of the compression cycle. PMID- 15296966 TI - In situ forming parenteral drug delivery systems: an overview. AB - Biodegradable injectable in situ forming drug delivery systems represent an attractive alternative to microspheres and implants as parenteral depot systems. Their importance will grow as numerous proteins will lose their patent protection in the near future. These devices may offer attractive opportunities for protein delivery and could possibly extend the patent life of protein drugs. The controlled release of bioactive macromolecules via (semi-) solid in situ forming systems has a number of advantages, such as ease of administration, less complicated fabrication, and less stressful manufacturing conditions for sensitive drug molecules. For these reasons, a number of polymeric drug delivery systems with the ability to form a drug reservoir at the injection site are under investigation. Here, we review various strategies used for the preparation of in situ forming parenteral drug depots and their potential benefits/draw-backs, especially with regard to the delivery of protein drug candidates. PMID- 15296967 TI - Much more from the chicken's egg than breakfast--a wonderful model system. AB - The chick embryo has played a major role in the advance of our understanding of embryonic development. It has been particularly helpful in relation to amniotes like the mouse, for unlike the mouse in the early embryo the epiblast is beautifully flat, while in the mouse it is all curled up. Here I briefly describe some history and a few of the major achievements that came from the chick embryo. PMID- 15296968 TI - The contribution of chicken embryology to the understanding of vertebrate limb development. AB - The chicken is an excellent model organism for studying vertebrate limb development, mainly because of the ease of manipulating the developing limb in vivo. Classical chicken embryology has provided fate maps and elucidated the cell cell interactions that specify limb pattern. The first defined chemical that can mimic one of these interactions was discovered by experiments on developing chick limbs and, over the last 15 years or so, the role of an increasing number of developmentally important genes has been uncovered. The principles that underlie limb development in chickens are applicable to other vertebrates and there are growing links with clinical genetics. The sequence of the chicken genome, together with other recently assembled chicken genomic resources, will present new opportunities for exploiting the ease of manipulating the limb. PMID- 15296969 TI - The acquisition of neural fate in the chick. AB - Neural development in the chick embryo is now understood in great detail on a cellular and a molecular level. It begins already before gastrulation, when a separation of neural and epidermal cell fates occurs under the control of FGF and BMP/Wnt signalling, respectively. This early specification becomes further refined around the tip of the primitive streak, until finally the anterior posterior level of the neuroectoderm becomes established through progressive caudalization. In this review we focus on processes in the chick embryo and put classical and more recent molecular data into a coherent scenario. PMID- 15296970 TI - Unveiling the establishment of left-right asymmetry in the chick embryo. AB - Vertebrates display striking left-right asymmetries in the placement of internal organs, which are concealed by a seemingly bilaterally symmetric body plan. The establishment of asymmetries about the left-right axis occurs early during embryo development and requires the concerted and sequential action of several epigenetic, genetic and cellular mechanisms. Experiments in the chick embryo model have contributed crucially to our current understanding of such mechanisms and are reviewed here. Particular emphasis is given to the elucidation of a genetic network that conveys left-right information from Hensen's node to the organ primordia, characterized to a significant degree of detail in the chick embryo. We also point out a number of early and late events in the determination of left-right asymmetries that are currently poorly understood and for whose study the chick embryo model presents several advantages. We anticipate that the availability of the chick genome sequence will be combined with multidisciplinary approaches from experimental embryology, biophysics, live-cell imaging, and mathematical modeling to boost up our knowledge of left-right organ asymmetry in the near future. PMID- 15296971 TI - Somite polarity and segmental patterning of the peripheral nervous system. AB - The analysis of the outgrowth pattern of spinal axons in the chick embryo has shown that somites are polarized into anterior and posterior halves. This polarity dictates the segmental development of the peripheral nervous system: migrating neural crest cells and outgrowing spinal axons traverse exclusively the anterior halves of the somite-derived sclerotomes, ensuring a proper register between spinal axons, their ganglia and the segmented vertebral column. Much progress has been made recently in understanding the molecular basis for somite polarization, and its linkage with Notch/Delta, Wnt and Fgf signalling. Contact repulsive molecules expressed by posterior half-sclerotome cells provide critical guidance cues for axons and neural crest cells along the anterior-posterior axis. Diffusible repellents from surrounding tissues, particularly the dermomyotome and notochord, orient outgrowing spinal axons in the dorso-ventral axis ('surround repulsion'). Repulsive forces therefore guide axons in three dimensions. Although several molecular systems have been identified that may guide neural crest cells and axons in the sclerotome, it remains unclear whether these operate together with considerable overall redundancy, or whether any one system predominates in vivo. PMID- 15296972 TI - The chick embryo: a leading model in somitogenesis studies. AB - The vertebrate body is built on a metameric organization which consists of a repetition of functionally equivalent units, each comprising a vertebra, its associated muscles, peripheral nerves and blood vessels. This periodic pattern is established during embryogenesis by the somitogenesis process. Somites are generated in a rhythmic fashion from the presomitic mesoderm and they subsequently differentiate to give rise to the vertebrae and skeletal muscles of the body. Somitogenesis has been very actively studied in the chick embryo since the 19th century and many of the landmark experiments that led to our current understanding of the vertebrate segmentation process have been performed in this organism. Somite formation involves an oscillator, the segmentation clock whose periodic signal is converted into the periodic array of somite boundaries by a spacing mechanism relying on a traveling threshold of FGF signaling regressing in concert with body axis extension. PMID- 15296973 TI - Segmentation and compartition in the early avian hindbrain. AB - For the comparative embryologists of the early 20th century, the segment-like bulges that appear transiently during the early stages of vertebrate hindbrain development were both the object of fascination and the subject of vigorous dispute. Conflicting views were held as to the significance of these 'rhombomeres' to brain development and their more general relevance to head evolution. Whether rhombomeres are inconsequential bumps in the embryonic brain or true segments-iterative or metameric units-has only recently been resolved. A number of studies using more modern techniques (such as immunohistochemistry, in situ hybridisation, axonal tracing, single cell labelling, heterotopic and orthotopic grafting, and the manipulation of gene expression by electroporation) have shown that the hindbrain has a truly metameric cellular organisation. The avian embryo has played a particularly prominent role in such studies by virtue of its large size and accessibility, its amenability to microsurgery, and its well-described anatomy. Furthermore, electrophysiological studies, also on avian embryos, have shown that segmentation of the parent neuroepithelium into rhombomeres plays a crucial part in establishing the functional organization of the hindbrain. Segmentation suggests the early allocation of defined sets of precursor cells and is therefore presumed to allow a specific identity for each successive segment to emerge from a common ground plan. This short review will focus on the contribution the avian embryo has made to our understanding of this fly-like region of the vertebrate brain, in respect of its morphology and neuronal architecture, the cellular and molecular mechanisms involved in establishing and maintaining the segments, and the molecular controls of segmental identity. PMID- 15296974 TI - The avian embryo as a model to study the development of the neural crest: a long and still ongoing story. AB - The aim of this review is to evoke briefly the progress that has been made in our knowledge about the contribution of the neural crest to the vertebrate body since it was discovered by Wilhelm His in 1868. Although first studied essentially in amphibian embryos, a large amount of what is known on this very special structure was gained by experimental work carried out on the avian embryo. The making of chimeras between quail and chick has permitted not only to analyse the normal course of neural crest cell migration and differentiation but also to reveal some of the cellular interactions that regulate these events. Looking to the future, we can foresee that the novel methods, which now allow to manipulate gene activities in definite groups of cells and at elected times in the developing embryo, will make the avian model even more instrumental than ever to approach the developmental problems raised by neural crest cell differentiation. PMID- 15296975 TI - The generation and diversification of spinal motor neurons: signals and responses. AB - Motor neurons are probably the best characterised neuronal class in the vertebrate central nervous system and have become a paradigm for understanding the mechanisms that control the development of vertebrate neurons. For many investigators working on this problem the chick embryo is the model system of choice and from these studies a picture of the steps involved in motor neuron generation has begun to emerge. These findings suggest that motor neuron generation is shaped by extracellular signals that regulate intrinsic, cell autonomous determinants at sequential steps during development. The chick embryo has played a prominent role in identifying the sources of these signals, defining their molecular identities and determining the cell intrinsic programs they regulate. PMID- 15296976 TI - The chicken genome and the developmental biologist. AB - Recently the initial draft sequence of the chicken genome was released. The reasons for sequencing the chicken were to boost research and applications in agriculture and medicine, through its use as a model of vertebrate development. In addition, the sequence of the chicken would provide an important anchor species in the phylogenetic study of genome evolution. The chicken genome project has its roots in a decade of map building by genetic and physical mapping methods. Chicken genetic markers for map building have generally depended on labour intensive screening procedures. In recent years this has all changed with the availability of over 450,000 EST sequences, a draft sequence of the entire chicken genome and a map of over 1 million SNPs. Clearly, the future for the chicken genome and developmental biology is an exciting one. Through the integration of these resources, it will be possible to solve challenging scientific questions exploiting the power of a chicken model. In this paper we review progress in chicken genomics and discuss how the new tools and information on the chicken genome can help the developmental biologists now and in the future. PMID- 15296977 TI - Gain- and loss-of-function in chick embryos by electroporation. AB - It remained very difficult to manipulate gene expression in chick embryos until the advent of in ovo electroporation which enabled the induction of both gain-of function, and recently loss-of-function, of a gene of interest at a specific developmental stage. Gain-of-function by electroporation is so effective that it has become widely adopted in developmental studies in the chick. Recently, it became possible to induce loss-of-function by introducing an siRNA expression vector by electroporation. In this review, the methods of electroporation for gain-of-function and for loss-of-function by siRNA are discussed. PMID- 15296978 TI - Efficient identification of regulatory sequences in the chicken genome by a powerful combination of embryo electroporation and genome comparison. AB - Recently expanded knowledge of gene regulation clearly indicates that the regulatory sequences of a gene, usually identified as enhancers, are widely distributed in the gene locus, revising the classical view that they are clustered in the vicinity of genes. To identify regulatory sequences for Sox2 expression governing early neurogenesis, we scanned the 50-kb region of the chicken Sox2 locus for enhancer activity utilizing embryo electroporation, resulting in identification of a number of enhancers scattered throughout the analyzed genomic span. The 'pan-neural' Sox2 expression in early embryos is actually brought about by the composite activities of five separate enhancers with distinct spatio-temporal specificities. These and other functionally defined enhancers exactly correspond to extragenic sequence blocks that are conspicuously conserved between the chicken and mammalian genomes and that are embedded in sequences with a wide range of sequence conservation between humans and mice. The sequences conserved between amniotes and teleosts correspond to subregions of the enhancer subsets which presumably represent core motifs of the enhancers, and the limited conservation partly reflects divergent expression patterns of the gene. The phylogenic distance between the chicken and mammals appears optimal for identifying a battery of genetic regulatory elements as conserved sequence blocks, and chicken embryo electroporation facilitates functional characterization of these elements. PMID- 15296979 TI - Avian pluripotent stem cells. AB - Pluripotent embryonic stem cells are undifferentiated cells capable of proliferation and self-renewal and have the capacity to differentiate into all somatic cell types and the germ line. They provide an in vitro model of early embryonic differentiation and are a useful means for targeted manipulation of the genome. Pluripotent stem cells in the chick have been derived from stage X blastoderms and 5.5 day gonadal primordial germ cells (PGCs). Blastoderm-derived embryonic stem cells (ESCs) have the capacity for in vitro differentiation into embryoid bodies and derivatives of the three primary germ layers. When grafted onto the chorioallantoic membrane, the ESCs formed a variety of differentiated cell types and attempted to organize into complex structures. In addition, when injected into the unincubated stage X blastoderm, the ESCs can be found in numerous somatic tissues and the germ line. The potential give rise to somatic and germ line chimeras is highly dependent upon the culture conditions and decreases with passage. Likewise, PGC-derived embryonic germ cells (EGCs) can give rise to simple embryoid bodies and can undergo some differentiation in vitro. Interestingly, chicken EG cells contribute to somatic lineages when injected into the stage X blastoderm, but only germ line chimeras have resulted from EGCs injected into the vasculature of the stage 16 embryo. To date, no lines of transgenic chickens have been generated using ESCs or EGCs. Nevertheless, progress towards the culture of avian pluripotent stem cells has been significant. In the future, the answers to fundamental questions regarding segregation of the avian germ line and the molecular basis of pluripotency should foster the full use of avian pluripotent stem cells. PMID- 15296980 TI - Genetic variants for chick biology research: from breeds to mutants. AB - The availability of the draft sequence of the chicken genome will undoubtedly propel an already important vertebrate research model, the domestic chicken, to a new level. This review describes aspects of chicken natural history and cross disciplinary biological value. The diversity of extant genetic variants available to researchers is reviewed along with institutional stock locations for North America. An overview of the problem of lack of long-term stability for these resources is presented. PMID- 15296981 TI - Prospects for transgenesis in the chick. AB - Research to develop a useful method for genetic modification of the chick has been on-going since the first demonstrations in the mouse in the 1980s that genetic modification is an invaluable tool for the study of gene function. Manipulation of the chick zygote is possible but inefficient. Considerable progress has been made in developing potentially pluripotent embryo stem cells and their contribution to somatic chimeric birds well-established. Germ line transmission of gametes derived from genetically modified embryo cells has not been described. Transfer of primordial germ cells from a donor embryo to a recipient and production of functional gametes from the donor-derived cells is possible. Genetic modification of primordial germ cells before transfer and their recovery through the germ line has not been achieved. The first transgenic birds described were generated using retroviral vectors. The use of lentiviral vectors may make this approach a feasible method for transgenic production, although there are limitations to the applications of these vectors. It is likely that a method will be developed in the next few years that will enable the use of transgenesis as a tool in the study of development in the chick and for many other applications in basic research and biotechnology. PMID- 15296982 TI - Overdiagnosis of breast cancer in screening. PMID- 15296983 TI - The sentinel node procedure in patients with melanoma. PMID- 15296984 TI - Axillary recurrence after sentinel lymph node biopsy. AB - Sentinel lymph node biopsy (SLNB) without further axillary dissection in patients with sentinel node-negative breast carcinoma appears to be a safe procedure to ensure locoregional control. During a median follow-up of 35 months the false negative rate was 1% in our study population of 185 patients. BACKGROUND: The objective of this prospective study is to provide data on follow-up of patients with primary operable breast carcinoma staged with SLNB without axillary lymph node dissection (ALND) if the sentinel lymph nodes (SLNs) were tumour-negative. METHODS: One hundred and eighty-five patients were enrolled. Preoperative dynamic and static lymphoscintigraphy were performed; both a vital blue dye and a gamma detection probe were used intraoperatively. Patients with tumour-positive SLNs received completion ALND or if no SLNs could be identified. All patients were monitored according to regional follow-up protocols. RESULTS: The SLNs were identified in 179 out of the 185 patients. In 73 patients the SLNs were tumour positive and in 106 patients tumour-negative. The median follow-up was 35 months (range 17-59). In one SLN-negative patient an axillary recurrence occurred 26 months after the SLNB (false-negative rate: 1%). CONCLUSIONS: SLNB without ALND appears to be a safe procedure to ensure locoregional control in SLN-negative breast carcinoma, if carried out by an experienced team. PMID- 15296985 TI - Operable breast cancer patients with diagnostic delay--oncological and emotional characteristics. AB - AIMS: Delayed diagnosis in cancer patients often implies presentation with advanced disease with poorer prognosis as a consequence. The aim of the present study was to gain more insight into mechanisms which determine patient delay in the diagnosis of operable breast cancer, stages I and II. METHODS: Patient delay was related to socio-demographic, psychological and clinical-oncological variables in 96 consecutive patients investigated one day before surgery. RESULTS: Patients with a diagnostic delay for one month or more (N=29) reported increased emotional control compared to patients without delay (N=67) (Mean score on Courtauld Emotional Control scale (CEC) 54.5 vs 46.4; p=0.003) and more often grade I tumour (17 out of 29 vs 16 out of 67 patients; p=0.002). Diagnostic delay was predicted independently by tumour differentiation (hazard ratio=5.0; p<0.01 (95% CI: 1.7-14.8)) and emotional control (hazard ratio=5.1; p<0.01 (95% CI: 1.6 16.1)). Multivariate survival analysis with tumour grading and patient delay as covariates showed significant survival effect for tumour differentiation only (hazard ratio=4.4; p<0.05 (95% CI: 1.3-15.4)). CONCLUSION: There seems to be an association between aggressiveness of tumour growth, diagnostic delay and emotional control in patients with early stage breast cancer. Clinical implications of these findings are discussed. PMID- 15296986 TI - Quality of life after sentinel lymph node biopsy in early breast cancer. AB - AIM: This study assessed the effects of multiple therapeutic factors on quality of life (QOL) in the treatment of breast cancer. METHODS: We surveyed 179 recurrence-free women with early breast cancer who had undergone a sentinel lymph node procedure, between January 1999 and June 2001. Age, tumour size, breast and axillary procedure, nodal status, chemotherapy, supra-clavicular fossa radiotherapy, and hormone therapy were tested as possible factors associated with poor QOL. RESULTS: Information on QOL was obtained for 148 out of 179 patients. Age less than 55 years and chemotherapy were factors associated with impairment of physical well-being. Tumour size was associated with poor socio-familial well being. Factors associated with altered arm subscale scores were age <55, axillary procedure, nodal status, chemotherapy and supra-clavicular fossa radiotherapy. Unexpectedly, sentinel lymph node (SLN) procedure delayed the onset of chemotherapy if the metastatic status of SLN was not diagnosed intra-operatively. CONCLUSION: Efforts are needed to improve the QOL of young patients. Axillary procedure affects only QOL related to arm morbidity. PMID- 15296987 TI - Patients' and surgeons' perspectives on axillary surgery for breast cancer. AB - AIMS: The objectives of this study were to compare the postoperative morbidity of Sentinel lymph node biopsy (SLNB) and axillary lymph node dissection (ALND) and to compare the views of surgeons and patients regarding postoperative morbidity. METHODS: A prospective and comparative study was initiated to evaluate, 1 year after surgery, morbidity and sequelae after SLNB in 231 patients. Group I (n=141) underwent SLNB without ALND, group II (n=90) underwent SLNB followed by ALND when SLN where involved. Morbidity analysis was performed, respectively, by surgeons and patients. RESULTS: One hundred and eighty-five patients (80.5%) completed the questionnaire including 113 with SLNB alone, and 72 with ALND. One year after surgery, SLNB produced less morbidity than ALND for symptoms and function. There were significantly different assessments between surgeons and patients for pain, arm mobility and sensitiveness. CONCLUSIONS: One-year postoperative morbidity after SLNB is significantly lower than after ALND but views of surgeons and patients appears to be significantly different. Additional data are required to assess late consequences of axillary surgery. PMID- 15296988 TI - A comparative study of envelope mastectomy and immediate reconstruction (EMIR) with standard latissimus dorsi immediate breast reconstruction. AB - BACKGROUND: Latissimus dorsi breast reconstruction has problems with scars at the donor site and on the reconstructed breast. We report the feasibility and aesthetic results of Envelope Mastectomy and Immediate Reconstruction (EMIR), which utilises a single lateral mammary fold incision. PATIENTS AND METHODS: Between 2001 and 2002, 20 EMIRs were performed in 19 patients, one as a staged bilateral procedure. Twenty consecutive patients, matched for body habitus, who had undergone standard latissimus dorsi breast reconstruction by the same surgeon from 1996 to 2000 were used as controls. Patient satisfaction was assessed using a validated Body Image Scale. Standard post-operative photographs were scored by three independent observers. RESULTS: Length of stay and complication rates were equivalent between both groups. Cosmetic self-assessment scores on the Body Image Scale and scores by the independent observers were satisfactory for both groups but no statistically significant difference was observed between groups. CONCLUSIONS: EMIR is a technically feasible and cosmetically acceptable method of immediate breast reconstruction. PMID- 15296989 TI - Preoperative radio/chemo-radiotherapy in combination with intraoperative radiotherapy for T3-4Nx rectal cancer. AB - AIMS: To analyse the results of a single institution experience of combined preoperative radio/chemo-radiotherapy and intraoperative electron-radiation therapy (IORT) for locally advanced rectal cancer and to compare the results with surgery alone retrospectively. METHODS: The study cohort comprised 99 patients with clinical T3-4NxM0 adenocarcinoma of the rectum who had received preoperative radio/chemo-radiotherapy, radical surgery, and IORT [Group I]. Until 1998, 67 patients were treated with radiation only [Group Ia], and after 1999, 32 patients were concurrently given tegafur and uracil (UFT) [Group Ib]. 68 patients with clinical T3-4NxM0 rectal cancer were treated with surgery alone [Group II]. RESULTS: The median follow-up was 67 months in Group I and 83 months in Group II. Local recurrence rate was 2% in Group I, which was significantly lower than 16% in Group II (p=0.002) Both disease-free survival and overall survival in Group I were significantly better than those in Group II (p=0.04, p=0.02, respectively). Sphincter preservation was possible in 78% in Group Ib, which was significantly more than 42% in Group Ia (p=0.002). CONCLUSIONS: The combined preoperative radio/chemo-radiotherapy and IORT for clinical T3-4Nx rectal cancer significantly reduces local recurrence and improves prognosis. Combination of preoperative radiotherapy and oral UFT improves the feasibility of sphincter-preservation. PMID- 15296990 TI - Two-stage liver resection and chemotherapy for bilobar colorectal liver metastases. AB - BACKGROUND: A significant number of patients with colorectal metastatic disease confined to the liver are inoperable at assessment. For these patients, the outlook is poor. Chemotherapy can 'down-stage' some tumours and render them operable. The authors present a series of patients with inoperable disease despite down-staging with chemotherapy, who underwent a two-stage resection to clear their metastatic disease. METHODS: The case-notes of 11 patients who were found to have inoperable hepatic metastatic disease were identified using computerised medical records and mean hospital stay, survival and long-term follow-up data was noted. RESULTS: The mean follow-up from initial resection was 13.5 months (range of 5-20 months). Three deaths were recorded in the follow-up interval. Causes of death included recurrence of hepatic disease following completion of two-stage resection, progression of original hepatic disease leading to inoperability at second stage operation and recurrence of original primary colorectal tumour. The mean survival in the patients who died was 17 months (range of 15-19 months). The remaining patients are alive to date with six patients showing no evidence of hepatic recurrence, follow-up period of 13 months (range of 8-20 months). One patient developed de novo prostate cancer and is awaiting his second liver resection, and one patient has stable hepatic disease with no evidence of progression. CONCLUSION: Two-stage liver resection can prolong survival when compared to chemotherapy alone, with a recurrence rate equivalent to ablation techniques. Longer-term studies are needed for further evaluation. PMID- 15296991 TI - The Japanese integrated staging score using liver damage grade for hepatocellular carcinoma in patients after hepatectomy. AB - AIMS: The new Japanese staging system for hepatocellular carcinoma (HCC), the Japan integrated staging (JIS) score, accounts for both Child-Pugh classification and Japan tumour node metastasis (TNM) staging. However, in HCC patients who undergo hepatectomy, liver function is relatively good and a better prognostic classification of hepatic function is necessary. METHODS: The present study was designed to analyse the modified JIS score using liver damage grade by the Liver Cancer Study Group of Japan instead of the Child-Pugh classification (using the category indocyanine green retention rate at 15 min [ICG(R15)] instead of encephalopathy), and to compare the Japan TNM stage in 101 patients who underwent resection of HCC. RESULTS: The liver damage grade showed significantly better discrimination of disease-free and overall survival than did the Child-Pugh classification. The modified JIS score system showed significant differences of disease-free and overall survivals in each score and this system was superior for discriminating survivals compared with the TNM staging. CONCLUSIONS: The combined staging system of hepatic function, particularly ICG(R15), and tumour stage provides a better prediction of prognosis. The JIS score using the liver damage grade was a useful predictor of prognosis of HCC patients who underwent hepatic resection. PMID- 15296992 TI - Totally implantable femoral vein catheters in cancer patients. AB - INTRODUCTION: Totally implantable devices are increasingly being utilized for chemotherapy treatment of oncological patients. When it is impossible to implant the reservoir on the anterior wall of the thorax, or when there is an obstruction of the superior vena cava system, alternative access routes must be sought. Of these, the femoral vein is the most utilized. Few studies have been performed to analyse the results obtained from the implantation and utilization of such catheters in the femoral vein. The goal of this work was to prospectively study the results obtained from the implantation of 20 TIC in femoral veins in a large sized cancer hospital with its own dedicated vascular clinical team. MATERIAL AND METHODS: Twenty femoral TIC were inserted in 20 patients out of a group of 560 cancer patients submitted to TIC implantation for chemotherapy. Evaluations were made of the early and late-stage complications and patient evolution until removal of the device, death or the end of the treatment. RESULTS: The prospective analysis showed a mean duration of 215 days for the catheters. There were 16 patients with no complications. There were no early complications. Among the late complications, three were infections, representing 0.69/1000 days of catheter use, and one was a deep vein thrombosis (0.23/1000 days of catheter use). One catheter was removed due to primary bacteremia and one due to subcutaneous pocket infection. Fourteen patients died while the catheter was functioning and four patients are still making use of the catheter. CONCLUSION: The low rate of complications implying catheter loss in this study confirms the safety and convenience of the use of femoral TIC in patients who cannot be submitted to implantation in the superior vena cava system. PMID- 15296993 TI - Experience with video-assisted surgery for suspected mediastinal tumours. AB - AIM: To assess the therapeutic feasibility of video-assisted thoracoscopic surgery (VATS) in the excision of suspected mediastinal tumours. METHODS: The case notes of 24 consecutive patients referred to a single surgeon between 1997 and 2002 for excision of suspected mediastinal tumours were reviewed. The operative, post-operative and pathological characteristics of patients treated thoracoscopically and by open procedure were analysed. RESULTS: Thirteen of 24 patients underwent thoracoscopic excision. The mean age of the two groups was similar as was the mean operating time and duration of chest drainage. However, patients in the thoracoscopic group had less chest drainage, less pain and a shorter hospital stay. CONCLUSIONS: Video-assisted thoracoscopic excision of mediastinal tumours is a safe and technically feasible procedure and may offer significant post-operative advantages over open procedures. PMID- 15296994 TI - The results of therapy for bilateral multiple primary lung cancers: 30 years experience in a single centre. AB - AIMS: This study reviews our 30 years experience in the clinical assessment and surgical management of bilateral multiple primary lung cancer (BMPLC). METHODS: Between January 1973 and December 2001, 1906 patients with primary lung cancer underwent surgical resection in Kanazawa University Hospital. Thirty-seven patients (1.9%) who had developed a BMPLC using the criteria of Martini and Antakli. RESULTS: Eighteen patients had synchronous lesions, and 18 patients had metachronous lesions. One patient had synchronous and metachronous lesions. Overall 10-year survival was 56%. The actuarial 5-year survival for bilateral synchronous cancers was 69%, median survival (MST) 90 months (range 8-153 months), and 10-year survival was 47%. The actuarial 5-year survival for second metachronous cancers was 51%, with an MST of 114 months (range 6-192 months). CONCLUSION: Aggressive surgical therapy is effective in patients with a bilateral MPLC if they satisfy the usual criteria of operability. The surgical methods that preserve healthy lung tissue such as sleeve resection and limited resection must be selected in compliance with cancer characters. PMID- 15296995 TI - The meaning and predictivity of Hurthle cells in fine needle aspiration cytology for thyroid nodular disease. AB - AIMS: The significance of Hurthle cells in thyroid nodule fine needle aspiration cytology (FNAC) samples remains uncertain. This study aims to clarify the meaning and the predictivity of this kind of cells. METHODS: One hundred and ten patients with Hurthle cells in FNAC of thyroid nodules were reviewed. Histopathology was correlated with cytological findings. RESULTS: The density of Hurthle cells in FNACs ranged from 20 to 100%. Eighty-nine patients had benign nodular disease (Hurthle cell or follicular adenoma), and 21 patients had malignant tumours. The presence of more than 50% Hurthle cells in FNAC correlated with benign or malignant Hurthle cell neoplasm. Hurthle cell carcinomas displayed more than 90% Hurthle cells in FNAC. CONCLUSIONS: Surgery is indicated for all nodular lesions with more than 50% Hurthle cells in FNAC. PMID- 15296996 TI - Results of surgical treatment of sporadic medullary thyroid carcinoma following routine measurement of serum calcitonin. AB - AIM: The aim of this study was to evaluate the results of the surgical management of medullary thyroid carcinoma (MTC), following the introduction of systematic calcitonin measurement in patients referred for thyroid diseases. METHOD: We included all the patients with elevated calcitonin and MTC from January 1993 to March 2001. RESULTS: Among 8497 patients, MTC was diagnosed in 52 with a mean age of 56.1 years. Thirty-two fine needle biopsies led to diagnose MTC in 19 cases. The median basal pre-operative calcitonin level was 245 pg/ml. Elevated calcitonin serum was the only indicator of MTC in 31 patients. Fifty-one patients underwent total thyroidectomies, with lymphadenectomy in 45. Thirteen patients had lymph node involvement. Post-operatively, 40 (77%) had normal basal and pentagastrin (Pg) stimulated calcitonin serum levels, and remained normal at a mean follow-up of 5.16 years (1.8-8). CONCLUSION: Routine pre-operative measurement of calcitonin should be performed because it is often the only indicator of MTC at an early stage. This could lead to an improved MTC cure rate. PMID- 15296997 TI - Mononucleotide markers of microsatellite instability in carcinomas of the urinary bladder. AB - AIMS: To determine the presence of microsatellite instability (MSI) and to assess the expression of the human mismatch repair (MMR) gene products hMLH1 and hMSH2 in primary transitional cell carcinomas (TCCs) of the urinary bladder in relation to clinico-pathological parameters. METHODS: Seventy-two cases of primary TCC were screened for the presence of alterations in MSI markers by molecular techniques and evaluated immunohistochemically for the expression of hMLH1 and hMSH2 proteins. Clinical data were available in 70 cases. The percentage of MSI rose to 16.6%. RESULTS: Reduced (<20%) hMLH1 expression was closely related to the presence of MSI (p=0.0004). Neither MMR proteins nor MSI was associated with grade, stage, papillary status. Clinical outcome analysed as a function of MSI did not show significant differences in terms of both disease-free and overall survival. Reduced hMLH1 expression was a significant predictor of shorter disease free survival in univariate and multivariate analysis. CONCLUSIONS: The presence of MSI is not related to classical clinico-pathological parameters in TCCs, nor does it appear to be of prognostic significance. hMLH1 was an important indicator for recurrence. PMID- 15296998 TI - Marginal mandibulectomy for lateral sulcus tumours. AB - OBJECTIVE: To report a retrospective series of marginal mandibulectomy for cancers of oral cavity, with special reference to squamous cancers of gingival buccal complex. METHODS: Retrospective record review of 107 patients who underwent marginal mandibulectomy between 1994 and 2001. RESULTS: Eighty-three marginal mandibulectomies were done for gingivo-buccal complex cancers. Local failure rate was 16%. The 2-year and 5-year disease free survival rates were 69 and 60%, respectively. The local recurrence free survival at the end of 2 and 5 years were 79 and 70%, respectively. CONCLUSION: In carefully selected patients, marginal mandibulectomy is an oncologically safe procedure to achieve good local control. PMID- 15296999 TI - Comparison of driving simulator performance and neuropsychological testing in narcolepsy. AB - Daytime sleepiness and cataplexy can increase automobile accident rates in narcolepsy. Several countries have produced guidelines for issuing a driving license. The aim of the study was to compare driving simulator performance and neuropsychological test results in narcolepsy in order to evaluate their predictive value regarding driving ability. Thirteen patients with narcolepsy (age: 41.5+/-12.9 years) and 10 healthy control patients (age: 55.1+/-7.8 years) were investigated. By computer-assisted neuropsychological testing, vigilance, alertness and divided attention were assessed. In a driving simulator patients and controls had to drive on a highway for 60 min (mean speed of 100 km/h). Different weather and daytime conditions and obstacles were presented. Epworth Sleepiness Scale-Scores were significantly raised (narcolepsy patients: 16.7+/ 5.1, controls: 6.6+/-3.6, P < or = 0.001). The accident rate of the control patients increased (3.2+/-1.8 versus 1.3+/-1.5, P < or = 0.01). Significant differences in concentration lapses (e.g. tracking errors and deviation from speed limit) could not be revealed (9.8+/-3.5 versus 7.1+/-3.2, pns). Follow-up investigation in five patients after an optimising therapy could demonstrate the decrease in accidents due to concentration lapses (P < or = 0.05). Neuropsychological testing (expressed as percentage compared to a standardised control population) revealed deficits in alertness (32.3+/-28.6). Mean percentage scores of divided attention (56.9+/-25.4) and vigilance (58.7+/-26.8) were in a normal range. There was, however, a high inter-individual difference. There was no correlation between driving performance and neuropsychological test results or ESS Score. Neuropsychological test results did not significantly change in the follow-up. The difficulties encountered by the narcolepsy patient in remaining alert may account for sleep-related motor vehicle accidents. Driving simulator investigations are closely related to real traffic situations than isolated neuropsychological tests. At the present time the driving simulator seems to be a useful instrument judging driving ability especially in cases with ambiguous neuropsychological results. PMID- 15297000 TI - Thalamic deep brain stimulation for posttraumatic action tremor. AB - We report a case of thalamic deep brain stimulation (DBS) for treatment of posttraumatic tremor. An 18-year-old right-handed man developed a disabling and medically refractory action tremor in the right upper extremity 9 months after sustaining diffuse axonal injury in a motor vehicle collision. DBS of the left ventral intermediate nucleus of the thalamus (Vim) suppressed the tremor without complication and should be considered as an option for the management of intractable posttraumatic tremor. PMID- 15297001 TI - The cerebrovascular dilatation effects of olprinone, a phosphodiesterase III inhibitor, in comparison with acetazolamide--a pilot study. AB - To examine the effects of olprinone, a phosphodiesterase III inhibitor, on cerebral blood flow (CBF), we compared the effects of olprinone on CBF to that of acetazolamide. Using technetium-99m-ethyl cysteinate dimer (99mTc-ECD) brain SPECT, we measured regional CBF (rCBF) at 33 sites, including 16 right and left pairs of non-infarct cerebral cortexes, in seven stroke patients (66.0+/-3.2 years) in a resting state and 15 min after the administration of acetazolamide. Within 1 week, rCBF at each site was measured 15 min after the initiation of olprinone infusion. Resting rCBF showed a significant negative correlation with the change in rCBF (DeltaCBF) during olprinone infusion (r = -0.43, P=0.013), but no significant correlation was seen following acetazolamide administration. The difference in rCBF between the right and left cortex increased more following acetazolamide administration (14.1+/-10.9 ml/(min 100 g)) than during olprinone infusion (5.4+/-4.8 ml/(min 100 g), P=0.013). The rCBF at four regions of interest (ROI) with low-resting CBF (< 49 ml/(min 100 g)) further decreased following the administration of acetazolamide. The vasodilatory effects of olprinone are dependent on resting CBF instead of on the intracerebral steal phenomenon that occurs with acetazolamide. PMID- 15297002 TI - Cerebrospinal fluid vascular endothelial growth factor in patients with amyotrophic lateral sclerosis. AB - Oxidative stress and glutamate-mediated toxicity may play an important role in the etiopathogenesis of amyotrophic lateral sclerosis (ALS). The vascular endothelial growth factor (VEGF) is a neuroprotective cytokine activated by hypoxia. The aim of this study was to measure VEGF levels in the cerebrospinal fluid (CSF) of ALS patients. The study concerned 30 ALS patients and 30 control subjects. The VEGF was measured by the enzyme-linked immunosorbent assay. The results have shown that CSF VEGF levels are significantly increased in patients with long duration of ALS and in patients with limb-onset of the disease compared with controls (P < 0.05). Moreover, the type of ALS patients' subgroup significantly influences CSF VEGF levels (P = 0.05). The CSF VEGF levels were significantly increased in patients with limb-onset compared to patients with bulbar-onset of ALS, and in patients with long duration of ALS compared to patients with its short duration (P < 0.05). There was a significant correlation between CSF VEGF levels and duration of ALS (P < 0.05). It seems that a significant increase in CSF VEGF levels in patients with limb-onset of ALS and in patients with long duration of the disease may have a protective role against glutamate-mediated toxicity and oxidative damage of motor neurons. However, the conclusions are limited due to relatively small subgroups of ALS patients and by lack of a control group consisting of healthy persons. Further investigations could help to confirm the results from this preliminary report. PMID- 15297003 TI - Individual risk factors for carpal tunnel syndrome: an evaluation of body mass index, wrist index and hand anthropometric measurements. AB - In this study we aimed to identify the role of the body mass index (BMI), wrist index and hand anthropometric measures as risk factors for carpal tunnel syndrome (CTS) in both genders. Based on clinical and electrophysiologic diagnostic criteria, 154 female and 44 male CTS patients, as well as 150 female and 44 male age-matched control subjects, were selected. BMI, wrist index, hand shape index, digit index and hand length/height ratio were compared between the CTS patients and the control subjects for each gender separately. Mean BMI was found to be a significant risk factor for CTS in both genders. The wrist index was found to be higher in female (P < 0.001) and in male (P = 0.034) CTS groups than in the respective control groups. Logistic regression analysis revealed the wrist index to be an independent risk factor in females, but not in males. Shape and digit indices were significantly higher in female CTS patients than in corresponding control subjects, and regression analysis showed the shape and digit indices to be independent risk factors for CTS. In the male CTS group, the shape and digit indices did not significantly differ from their controls. Differences in the hand length/height ratio were not statistically significant in female and male CTS patients compared to their controls and it was not found to be an independent risk factor for CTS. Our study confirmed BMI as an independent risk factor for CTS in both genders. Hand and wrist anthropometrics were found to be independent risk factors for CTS in females, but not in males. PMID- 15297004 TI - Silent radiological imaging time in patients with brain metastasis. AB - Cerebral metastasis is a common finding in patients with systemic carcinoma and is an indication for progress of the disease. When brain metastases occur, they lead to a considerable decrease in both survival and the quality of life, in patients who otherwise might be functional. Furthermore, the location, size and number of such lesions, play a decisive role in management and prognosis. Even though early diagnosis and treatment is curative in rare cases, it may lead to a useful remission of the central nervous system (CNS) symptoms, enhance the patient's quality of life and prolong survival. The radiological exams established in the diagnosis of this condition, include computed tomography (CT) scan or magnetic resonance imaging (MRI). In cases of "micrometastatic" disease though, these exams may be pronounced as normal. This retrospective study was performed in patients with advanced systemic disease, who presented with neurological findings of intracranial mass lesion, in the absence of radiological evidence. Early-occurring symptoms were evaluated in accordance to location of the primary disease and follow-up with repetitive MRI scans was performed, in an attempt to confirm the diagnosis and facilitate prompt and appropriate treatment. PMID- 15297005 TI - Occipital stroke shortly after cannabis consumption. AB - There are several indications that imply cannabis consumption triggers cerebrovascular events. A right occipital ischemic stroke occurred in a 37-year old Albanese man with a previously uneventful medical history, 15 min after having smoked a cigarette with approximately 250 mg of marijuana. Clinical manifestations of the stroke were left-sided hemiparesis, hemi-hypesthesia and blurred vision, which vanished spontaneously and almost completely after 3 days. He has been smoking joints regularly from the age of 27, with a frequency of 2-3 joints per week during the 6 months that preceded his stroke. Except for cigarette smoking and slight dyslipidaemia, classical risk factors for stroke/embolism were absent. Therefore, as the family history for cerebrovascular events, blood pressure, clotting tests, examinations for thrombophilia, vasculitis, extracranial and intracranial arteries and cardiac investigations were normal or respectively negative, the stroke was attributed to the chronic cannabis consumption. PMID- 15297006 TI - Clinical and electrophysiological features in Chinese patients with Kennedy's disease. AB - Kennedy's disease is a X-linked neuromuscular disorder caused by an expanded trinucleotide repeat in the androgen receptor gene. To ascertain the clinical diagnosis of Kennedy's disease in a Chinese population, we used a rapid, accurate PCR-based sizing method for the CAG repeat allelotype. The clinical and electrophysiological features of affected patients are described. The CAG repeats ranged from 43 to 53 and were inversely correlated with the age of onset (r = 0.63; P < 0.005). PMID- 15297007 TI - Do neurosurgeons subscribe to the guideline lumbosacral radicular syndrome? AB - BACKGROUND: This study presents a survey of the opinion of neurosurgeons on the multidisciplinary clinical guideline 'lumbosacral radicular syndrome'. The aim was to describe to what extent neurosurgeons in the Netherlands endorse the content of this guideline. The guideline was issued in 1996 by the Netherlands Institute of Quality Health Care and this is the first attempt to evaluate the implementation of this guideline. METHODS: All active neurosurgeons (n=92) in the Netherlands were invited to complete a questionnaire investigating to what extent they agree with the 26 recommendations in the guideline 'lumbosacral radicular syndrome'. The results are represented in frequencies (%) in order to express the magnitude of their consent or dissent with the recommendations. RESULTS: Overall, 75% of the neurosurgeons responded and, of these, 94% agreed (at least partially) with the content of the guideline. Of the 26 recommendations in the guideline, seven were not fully endorsed by the neurosurgeons. Three of these seven recommendations may need revision based on newly published data. CONCLUSION: This survey shows that almost all neurosurgeons subscribed (at least partially) to the multidisciplinary LRS guideline. Therefore, one important aspect of the implementation process has been fulfilled, i.e. acceptance of the content of the guideline. PMID- 15297008 TI - Electrophysiological effects and clinical results of direct brain stimulation for intractable epilepsy. AB - OBJECTIVE: Epilepsy can be considered as a result of the imbalance of the excitatory and inhibitory processes. Therefore, the artificial enhancement of the activity of brain inhibitory mechanisms might lead to a beneficial therapeutic effect for intractable epilepsy patients. MATERIAL AND METHODS: Studies of the inhibitory effects of electrical stimulation of the head of the caudate nucleus (HCN), cerebellar dentate nucleus (CDN), thalamic centromedian nucleus (CM), and neocortical and temporal lobe mesiobasal epileptic foci were performed on 150 patients with implanted intracerebral electrodes. Chronic brain stimulation with implanted neurostimulators was performed on 54 patients. Sixteen were followed up to 1.5 years (mean 1.2 years). RESULTS: The study demonstrated that 4-8 Hz HCN and 50-100 Hz CDN stimulation suppressed the subclinical epileptic discharges and reduced the frequency of generalized, complex partial, and secondary generalized seizures. CM stimulation (20-130 Hz) desynchronized the EEG and suppressed partial motor seizures. Direct subthreshold 1-3 Hz stimulation of the epileptic focus may suppress rhythmic afterdischarges (ADs). Seizures were eliminated for 26 of 54 patients (48%), worthwhile improvement was achieved for 23 of 54 patients (43%), and no improvement was observed in 5 of 54 patients (9%). CONCLUSION: The artificial increase of the activity of brain inhibitory system may suppress the activity of epileptic foci, and, in long run, stabilize this epileptic foci activity at a lower, perhaps normal, level. Therapeutic direct brain stimulation, therefore, might serve as a useful tool in the treatment of intractable and multifocal epilepsy, and might be combined with ablative surgical methods. PMID- 15297009 TI - A compensatory mechanism in unilateral akinetic-rigid syndrome: an fMRI study. AB - The motor mechanisms of a patient with unilateral hand clumsiness in the early stages of akinetic-rigid syndrome were assessed by functional magnetic resonance imaging (fMRI). Movements of the unaffected hand produced activation in the contralateral sensorimotor cortex (SMC) and ipsilateral SMC and superior parietal lobule (SPL). The affected hand activated the bilateral SMCs, supplementary motor areas, and SPLs. We speculated that the bilateral activation indicated recruitment of a pre-existing bilaterally organized large-scale neural network to perform the motor task. PMID- 15297010 TI - Sudden progression of a glioblastoma in partial remission? AB - There are only sparse data on viral CNS infections in patients with malignant glioma. We report a case of fatal herpes encephalitis in a patient with glioblastoma in partial remission and provide a short review of the literature. PMID- 15297011 TI - Aphemia-like syndrome from a right supplementary motor area lesion. AB - Lesions in the left supplementary motor area (SMA) can result in a transcortical motor aphasia with nonfluent spontaneous verbal output and relatively preserved repetition. Reading and writing are proportionally affected. We report a patient with an ischemic lesion in the right SMA. He had impaired articulation and normal repetition plus preserved reading and writing, consistent with an aphemia. This patient supports the dissociation of articulatory fluency and linguistic fluency and suggests that both SMAs affect the initiating of articulatory movements required to produce words whereas the left SMA also affects linguistic aspects of speech. PMID- 15297012 TI - Scalp malignant peripheral nerve sheath tumor (MPNST) with bony involvement and new bone formation: case report. AB - Malignant peripheral nerve sheath tumors (MPNSTs) are rare neoplasms, usually arising from somatic soft tissues or peripheral nerves. Primary MPNST of the scalp is extremely rare, with only a single case reported so far. Here, we describe an unusual case of scalp MPNST in a 50-year-old male. The tumor was associated with bony projection, intracranial extension and underlying bone destruction. The tumor was treated with complete surgical excision followed by adjuvant radiotherapy. Histologically, the tumor showed malignant spindle cells with focal S-100 positivity on immunohistochemistry and a diagnosis of MPNST was made. This case is being reported for its rarity and presence of associated bony projection, which to the best of our knowledge, has not been described before in soft tissue sarcomas. PMID- 15297013 TI - Bithalamic infarcts: embolism of the top of basilar artery or deep cerebral venous thrombosis? AB - Bithalamic infarcts are usually attributed to thromboembolism of the top of the basilar artery. However, in some cases, deep cerebral venous thrombosis and thrombosis of cerebral venous sinuses was proved to be the cause. The case of a 47-year-old female with ischemic thalamic and mesencephalic lesions is reported, that was attributed to thrombosis of internal cerebral veins. In cases of bithalamic infarcts, apart from the top of the basilar artery syndrome, deep cerebral venous thrombosis should be taken into consideration. Neuroimaging findings such as generalized cerebral edema, multiple infarcts or hemorrhages, hyperdense appearance of cerebral sinuses or veins and filling defects in the cerebral venous sinuses in contrast-CCT, can lead to the proper diagnosis. PMID- 15297014 TI - Protective effect of naringin, a bioflavonoid on ferric nitrilotriacetate-induced oxidative renal damage in rat kidney. AB - An iron chelate, ferric nitrilotriacetate (Fe-NTA), induces acute proximal tubular necrosis as a consequence of lipid peroxidation and oxidative tissue damage that eventually leads to high incidence of renal adenocarcinomas in rodents. This study was designed to investigate the effect of Naringin, a bioflavonoid with anti-oxidant potential, on Fe-NTA-induced nephrotoxicity in rats. One hour after a single intra-peritoneal (i.p.) injection of Fe-NTA (8 mg iron/kg body weight), a marked deterioration of renal architecture and renal function was observed. Fe-NTA induced a significant renal oxidative stress, demonstrated by elevated thiobarbituric acid reacting substances (TBARS) and reduction in activities of renal catalase, superoxide dismutase, and glutathione reductase. Pre-treatment of animals with Naringin, 60 min before Fe-NTA administration, markedly attenuated renal dysfunction, morphological alterations, reduced elevated TBARS, and restored the depleted renal anti-oxidant enzymes. These results clearly demonstrate the role of oxidative stress and its relation to renal dysfunction and suggest a protective effect of Naringin on Fe-NTA induced nephrotoxicity in rats. PMID- 15297015 TI - Beryllium-specific immune response in primary cells from healthy individuals. AB - The effect of beryllium (Be) exposure has been extensively studied in patients with chronic beryllium disease (CBD). CBD patients carry mutated MHC class II alleles and show a hyperproliferation of T cells upon Be exposure. The exact mechanism of Be-induced T-cell proliferation in these patients is not clearly understood. It is also not known how the inflammatory and suppressive cytokines maintain a balance in healthy individuals and how this balance is lost in CBD patients. To address these issues, we have initiated cellular and biochemical studies to identify Be-responsive cytokines and other cellular markers that help maintain a balance in healthy individuals. We have established an immune cell model derived from a mixture of peripheral blood mononuclear cells (PBMCs) and dendritic cells (DCs). In this article, we demonstrate that pro-inflammatory cytokine IL6 shows decreased release whereas suppressive cytokine IL10 shows enhanced release after 5-10 h of Be treatment. Furthermore, the Be-specific pattern of IL6 and IL10 release is dependent upon induction of threonine phosphorylation of a 45 kDa cytosolic protein (p45), as early as 90 min after Be treatment. Pharmacological inhibition of phosphatidylinositol 3' kinase (PI3'K) by wortmannin and p38 mitogen-activated protein kinase (MAPK) by SB203580 reveal that PI3'K mediates Be-specific p45 phosphorylation and IL6 release, whereas p38 MAPK regulates the release of IL6 and IL10 and the phosphorylation of p45 independent of metal-salt treatment. While the IL10 and IL6 release pathways are uncoupled in these cells, they are linked to phosphorylation of p45. These findings suggest that the balance between IL10 and IL6 release and the correlated p45 phosphorylation are important components of the Be-mediated immune response in healthy individuals. PMID- 15297016 TI - Toxicity of nutritionally available selenium compounds in primary and transformed hepatocytes. AB - The essential trace element selenium is also toxic at low doses. Since supplementation of selenium is discussed as cancer prophylaxis, we investigated whether or not bioavailable selenium compounds are selectively toxic on malignant cells by comparing primary and transformed liver cells as to the extent and mode of cell death. Sodium selenite and selenate exclusively induced necrosis in a concentration-dependent manner in all cell types investigated. In primary murine hepatocytes, the EC50 was 20 microM for selenite, 270 microM for selenate, and 30 microM for Se-methionine. In the human carcinoma cell line HepG2, the EC50 for selenite was 40 microM, and for selenate 1.1 mM, whereas Se-methionine was essentially non-toxic up to 10 mM. Similar results were found in murine Hepa1-6 cells. Exposure of primary murine cells to selenate or selenite resulted in increased lipid peroxidation. Toxicity was inhibited by superoxide dismutase plus catalase, indicating an important role for reactive oxygen intermediates. In primary hepatocytes, metabolical depletion of intracellular ATP by the ketohexose tagatose, significantly decreased the cytotoxicity of Se-methionine, while the one of selenite was increased. These data do not provide any in vitro evidence that bioavailable selenium compounds induce preferentially apoptotic cell death or selectively kill transformed hepatocytes. PMID- 15297017 TI - Involvement of cytochrome c release and caspase activation in toosendanin-induced PC12 cell apoptosis. AB - Our previous study showed that toosendanin, a triterpenoid derivative isolated from a Chinese traditional medicine, could induce apoptosis in PC12 cells. In this study we confirmed the apoptosis-inducing effect of toosendanin in PC12 cells with new evidences in morphology and biochemistry: the shrinkage of cytosol, the condensation and fragmentation of nuclei and the formation of DNA ladder. It was also demonstrated that toosendanin decreased the PC12 cell viability in a time- and concentration-dependent manner. To elucidate the pathway linked with the toosendanin-induced apoptosis, the cytochrome c in the cytosol and the cleavage of poly(ADP-ribose) polymerase (PARP) were examined. The obtained results showed that toosendanin caused the release of cytochrome c from mitochondria into the cytosol and then led to the activation of caspase, indicating that the cytochrome c release and caspase activation were involved in the toosendanin-induced apoptosis process. These results suggested the possibility that toosendanin could serve as a candidate for anti-cancer drug. PMID- 15297018 TI - Tachykinin substance P depletion by capsaicin exacerbates inflammatory response to sidestream cigarette smoke in rats. AB - To evaluate the role of substance P (SP)-containing C-fiber nerves in the development of the inflammatory responses to sidestream cigarette smoke (SSCS), female Fischer 344 rats were randomly assigned into vehicle and capsaicin groups, respectively. Then, half the number in each group (N = 24) was nose-only exposed to air or 0.4 mg/m3 total particulate matter of SSCS for 4 h/day for 7 days. Exposure of the vehicle rats to SSCS induced obvious pulmonary neurogenic inflammation as indicated by elevations in plasma extravasation and proinflammatory cytokine secretions [interieukin (IL)-1beta and IL-12]. In addition, except for SP release, SSCS exposure significantly induced the tachykininergic toxicities at the gene level: upregulation of beta preprotachykinin-I (beta-PPT-I) mRNA. However, neither SSCS exposure nor capsaicin pretreatment affects the immunolabeling density of neurokinin-1 receptor (NK-1R) in airway epithelium. SSCS also significantly inactivated pulmonary neutral endopeptidase (NEP) in lung tissue. Moreover, pretreatment with capsaicin significantly exacerbated the SSCS-induced inflammatory responses mentioned above as well as the release of plasma protein. Considering that capsaicin did not affect the normal control baselines of these parameters except for a decrease in NK-1R mRNA, we conclude that the degree of SSCS-induced inflammatory response was exacerbated because of the depletion of stored SP and/or inactivation of capsaicin-sensitive C-fiber nerves. Our data suggest the loss of afferent tachykinin SP signaling may lead to dysfunction of the sensory C fiber nerve reflexes during exposure to SSCS, suggesting that SP serves a protective role. PMID- 15297019 TI - N-nitroso metabolite of carbofuran induces apoptosis in CHL cells by cytochrome c mediated activation of caspases. AB - Carbofuran is an anti-acetylcholinesterase insecticide regarded as a relatively safe chemical based on extensive toxicological data. However, the N-nitroso metabolite of carbofuran has been reported to be genotoxic. We previously observed that N-nitrosocarbofuran (NOCF) induces apoptosis and cell cycle arrest in Chinese hamster lung (CHL) fibroblasts. To extend our initial observations, we investigated the molecular mechanism of NOCF-induced apoptosis. Treatment of cells with NOCF caused dose-dependent upregulation of cytosolic factors, such as Bax and Bid, and release of cytochrome c, which were accompanied by activation of caspase-9. We also observed activation of caspase-8 and caspase-3, and subsequent cleavage of poly(ADP-ribose) polymerase. A broad-spectrum caspase inhibitor and a caspase-8-specific inhibitor completely blocked caspase-3 activation and cell death induced by NOCF. These results suggest that the mitochondrial pathway is primarily involved in the NOCF-induced apoptosis of CHL cells. PMID- 15297020 TI - Effect of oral administration of terephthalic acid on testicular functions of rats. AB - To investigate the toxic effect of terephthalic acid (TPA) on testicular functions of rats, male Sprague-Dawley rats were orally administered TPA in diet at the levels 0 (control), 0.2, 1 and 5% for 90 days. Testicular functions were assessed by histopathology, testicular sperm head counts, daily sperm production, sperm motility (measured by computer-assisted sperm analysis, CASA), biochemical indices (marker testicular enzymes), and serum testosterone. Oral feeding with terephthalic acid did not cause body and testes weight loss in TPA-treated groups. Histopathologically, damages of spermatogenic cells and Sertoli cells were observed by electron microscope, testicular sperm head counts, daily sperm production, and activities of sorbitol dehydrogenase (SDH) were decreased significantly in the 5% TPA group. The motility of spermatozoa was reduced significantly in all treated groups, which was correlated with administration doses. Serum testosterone concentrations were not declined in treated groups. In conclusion, TPA can cause impairment of testicular functions. The primary sites of action may be spermatogenic cells and Sertoli cells. The results of the present study provide first information of TPA on testicular functions in male rats. PMID- 15297021 TI - Disruption of sphingolipid homeostasis by myriocin, a mycotoxin, reduces thymic and splenic T-lymphocyte populations. AB - Myriocin is a naturally occurring fungal metabolite possessing potent immunosuppressive properties. The biochemical mechanism of action of this compound is inhibition of serine palmitoyltransferase (SPT), the key rate limiting enzyme in sphingolipid biosynthesis, intermediates of which are important mediators of immune signaling. Previous studies have shown that myriocin strongly suppressed immune function with T-lymphocyte functions being most sensitive. To further our understanding of the mechanisms of this effect, we investigated the impact of subacute treatment with myriocin on lymphocyte populations in the thymus and spleen of male BALB/c mice following intraperitoneal injection of myriocin at 0, 0.1, 0.3, and 1.0 mg/kg daily for 5 consecutive days. Cellular analysis of the thymus demonstrated that total cellularity was dose-dependently reduced and the reduction was significant in mice treated with 1.0 mg/kg myriocin. Phenotyping showed that CD4+ and CD4+/CD8+ double positive lymphocyte populations were sensitive to myriocin. No change in total cellularity of the spleen was noted but there was a significant reduction in the CD4+ lymphocyte population in mice treated with 1.0 mg/kg myriocin. There was a strong positive correlation between total CD4+ lymphocytes in the thymus and those in the spleen. Analysis of sphingolipid levels showed a dose-dependent reduction of sphinganine in the thymus, which were positively correlated with all reductions in lymphocyte populations. These results suggest that the immunosuppressive properties of myriocin may be due to diminished T-lymphocyte populations likely related to inhibition of SPT and disruption of sphingolipid homeostasis. PMID- 15297022 TI - Phthalate treatment does not influence levels of IgE or Th2 cytokines in B6C3F1 mice. AB - Bronchial asthma is mediated, in part, by the immunoregulatory cytokines interleukins 4 and 13 (IL-4 and IL-13). These cytokines stimulate IgE synthesis that in turn is associated with airway hyper-responsiveness. Compounds that stimulate IgE synthesis and elicit bronchial reactivity are generally considered to be respiratory sensitizers. Recently, it has been hypothesized that exposure to phthalates may contribute to childhood asthma. To address this question, di-(2 ethylhexyl) phthalate (DEHP) was tested using a protocol adapted from work by Dearman that involves topical application (and challenge) of test substances to mice followed by measurements of total serum IgE. In addition, auricular lymph nodes were harvested for measurement of IL-4 and IL-13 proteins and their corresponding messenger RNAs. Because skin absorption of high molecular weight phthalates is limited, liver weight increase, a measure of peroxisomal proliferation, was monitored to assure that internal dosing had been achieved. ELISA and RNAse protection assays demonstrated that DEHP treatment did not significantly affect IgE, IL-4, or IL-13 levels. Similarly, IL-4 and IL-13 mRNA levels were not elevated. In contrast, all of these were significantly elevated by trimellitic anhydride (TMA), a respiratory sensitizer used as the positive control in this assay. Liver weights were significantly elevated by DEHP, providing evidence of sufficient percutaneous absorption to induce physiological responses. To extend these observations, three other commercial phthalate ester plasticizers, di-isononyl phthalate (DINP), di-isohexyl phthalate (DIHP), and butyl benzyl phthalate (BBP), were assessed using the same protocol. As above, ELISA and RNAse protection assays showed that IgE, IL-4, and IL-13 proteins, and IL-4 and IL-13 mRNAs in the phthalate-treated animals were all at levels similar to that of control values. The positive control, TMA, produced large, statistically significant increases in all parameters, demonstrating responsiveness of the assay. Another control, dinitrochlorobenzene (DNCB), a contact sensitizer, also responded as expected, producing smaller but statistically significant increases in IgE and in mRNA for IL-4 and IL-13 but not in the levels of these cytokines. In summary, treatment with DEHP, DINP, DIHP, and BBP did not result in significant elevations in total serum IgE, IL-4, or IL 13. As such it is unlikely that these substances would produce antibody-mediated respiratory allergy. PMID- 15297023 TI - Assessment of the potential irritation and photoirritation of novel amino acid based surfactants by in vitro methods as alternative to the animal tests. AB - The ultraviolet-A radiation damage effects on skin and eyes will be increased by phototoxic compounds which could be present in pharmaceutical or cosmetic formulations. Great efforts have been made in the last years to find surfactants to replace those with phototoxic potential in commercial use. Series of different in vitro models for phototoxicity, included to validated neutral red uptake (NRU) 3T3 phototoxicity assay are useful screening tools. The phototoxic effects of a novel family of glycerol amino acid-based surfactant compounds were examined via these assays. Human red blood cells and two immortalised cell lines, murine fibroblast cell line 3T3, and one human keratinocyte cell line, HaCaT, were the in vitro models employed to predict potential photoirritation. The phototoxic end points assessed were hemolysis (human red blood cell test) and resazurin transformation to resorufin and NRU in cell culture methods. The results suggest that no phototoxic effects by any new amino acid derived-surfactants, could be identified. PMID- 15297024 TI - Role of nitric oxide in NAG-ST induced store-operated calcium entry in rat intestinal epithelial cells. AB - This study was undertaken to find out the mechanism of non-agglutinable Vibrio cholerae heat-stable enterotoxin (NAG-ST)-induced calcium influx across the plasma membrane. Adriamycin, an inhibitor of IP3-specific 3-kinase, could not inhibit NAG-ST-induced calcium influx in rat intestinal epithelial cells, which suggested that inositol 1,3,4,5-tetrakisphosphate (IP4) had no role in NAG-ST induced calcium influx. NAG-ST increased intracellular nitric oxide level of rat enterocytes as measured by a fluorimetric method using a fluoroprobe 4,5 diaminofluorescein-2-diacetate (DAF-2DA). N-Nitro-L-arginine, an inhibitor of nitric oxide synthase, inhibited NAG-ST-induced rise in nitric oxide level and also calcium influx. Inhibition of inositol trisphosphate (IP3)-mediated intracellular calcium mobilization by Dantrolene could also inhibit NAG-ST induced rise in intracellular nitric oxide level. Moreover, inhibition of soluble guanylate cyclase by inhibitors (ODQ, LY83583) could inhibit the NAG-ST-induced rise in cyclic guanosine-3',5'-monophosphate (cGMP) level and calcium influx. From this study, it is evident that NAG-ST causes IP3-mediated calcium release from intracellular calcium store, which then stimulates nitric oxide production by activating nitric oxide synthase and the nitric oxide through cGMP activates calcium influx. PMID- 15297025 TI - Analysis of changes in energy and redox states in HepG2 hepatoma and C6 glioma cells upon exposure to cadmium. AB - The energy and redox states of the HepG2 hepatoma and the C6 glioma cells were studied by quantifying the levels of ATP, ADP, AMP, GSH, and GSSG. These values were used to calculate the energy charge potential (ECP = [ATP + 0.5ADP]/TAN), total adenosine nucleotides (TAN = ATP + ADP + AMP), total glutathione (TG = [GSH + GSSG]/TAN), and the redox state (GSH/GSSG ratio). For comparison between cell types, the level of each energy metabolite (ATP, ADP, and AMP) was normalized against TAN of the respective cell. The results showed that ATP:ADP:AMP ratio was 0.76:0.11:0.13 for the HepG2 cells and 0.80:0.11:0.09 for the C6 glioma cells. ECP was 0.81 +/- 0.01 and 0.85 +/- 0.01 for the HepG2 and the C6 glioma cells, respectively. GSH/GSSG ratio was 2.66 +/- 0.16 and 3.63 +/- 0.48 for HepG2 and C6 glioma cells, respectively. TG was 3.2 +/- 0.54 for the HepG2 cells and 2.43 +/- 0.18 for the C6 glioma cells, indicating that the level of total glutathione is more than two to three times higher than the total energy metabolites in these cell lines. Following a 3-h incubation in medium containing different concentrations of Cd, there was a dose-dependent decrease in cell viability. The 3-h LC50 for the HepG2 cells was 0.5 mM and that for the C6 glioma cells was 0.4 mM. Cellular TAN decreased with a decrease in cell viability. Upon careful analysis of the energy state, there was a significant increase in relative amount of ATP and decrease in ADP and AMP in both cells as Cd concentration increased from 0 to 0.1, 0.2, and 0.6 mM. However, ECP in both cell lines increased, which indicated that the level of high energy phosphate was adequate. There was also a significant increase in TG and a significant decrease in GSH/GSSG in the C6 glioma cells when cells were exposed to as low as 0.1 mM Cd, which suggested that the cellular redox state was compromised. The HepG2 cells, on the other hand, showed no significant change in both TG and GSH/GSSG level until Cd concentration reached 0.6 mM. Results of the present study also showed that there were differences between the two cells in response to the same level of Cd exposure. The C6 glioma cells were more sensitive to Cd-induced injuries. Although there was a decrease in total amount of high energy phosphate as cell viability decreased, the surviving cells were not devoid of high energy phosphates. The relative abundance of ATP amongst the adenosine nucleotide pool and the increase in ECP could be interpreted as a way the cells signal the conservation of energy utilization in response to the damaged mitochondrial function. This move for energy conservation might be the cause of eventual cell death through the process of apoptosis. PMID- 15297026 TI - Synergistic effects of fumonisin B1 and ochratoxin A: are in vitro cytotoxicity data predictive of in vivo acute toxicity? AB - Contamination of food and feeds by mycotoxins is a major problem of human and animals health concern which is also extremely detrimental to economy. Mycotoxins producing moulds may produce a diversity of toxins such as aflatoxins, ochratoxins, trichothecenes, zearalenone, fumonisins, tremorgenic toxins and ergot alkaloids. Although toxicological, environmental and epidemiological studies have addressed the problem of these toxins one by one, more than one mycotoxin are found usually in the same contaminated commodities. That rises the incommensurable problem of multi-toxicosis in which the respective metabolites are also involved. These mycotoxins bear potential toxicity leading to acute and chronic effects in humans and animals, depending on species. The mechanisms that lead to toxic effects, such as immune toxicity, and carcinogenicity are complexe. The risk assessment for humans potentially exposed to multi-mycotoxins suffers very much from the lack of adequate food consumption data. Furthermore, for a given mycotoxin synergism and antagonism with other mycotoxins found in the same food commodities are not taken into account. The case of combination of ochratoxin A (OTA) and fumonisin B1 (FB1) has been addressed in the present paper with the purpose of predicting the in vivo toxicity using a simple in vitro test, i.e. neutral red uptake, in three different cell-lines, C6 glioma cells, Caco-2 cells and Vero cells. Using the equation of [ATLA 27 (1999) 957], in vivo toxicity (LD50) is in adequation with the in vitro data, (IC50 values) for both toxins as well as for the combination of 10 microM OTA and variable concentrations of FB1 (10-50 microM). A synergistic effect is prouved in vitro that is in line with some in vivo data from the literature. Such simple in vitro test may thus help predicting in vivo toxicity of combinations of mycotoxins naturally occurring in foodstuffs. PMID- 15297027 TI - Styrene 7,8-oxide induces mitochondrial damage and oxidative stress in neurons. AB - Styrene 7,8-oxide (SO) is the main metabolite of styrene, a neurotoxic compound used industrially. Neurons exposed to SO undergo apoptosis with characteristic features including chromatin rearrangements and caspase activation. We report that the execution phase of apoptosis induced by SO (0.3 mM) in SK-N-MC neurons is triggered by translocation of apoptogenic factors (e.g., cytochrome c) into the cytosol. In addition, mitochondria exhibit lower Ca2+ capacity and loss of mitochondrial membrane potential (DeltaPsi). Lipid peroxidation, measured as thiobarbituric acid reactive substances (TBARS), is increased after 12 h. Pre treatment with the antioxidant MnTBAP (100 microM) prevents the decrease of Ca2+ capacity, cytochrome c release, activation of caspases, exposure of phosphatidylserine and cell death. Hence, the neurotoxic effects of SO are related to mitochondrial damage and oxidative stress. PMID- 15297028 TI - Protective effect of (+)-catechin against 5-fluorouracil-induced myelosuppression in mice. AB - The object of this study was to investigate the efficacy of (+)-catechin, which was isolated from Actinidia arguta Planch (Actinidiaceae), as a bone marrow cell proliferation-promoting compound against the hematotoxicity of 5-fluorouracil (5 FU) in mice. Intraperitoneally injected (+)-catechin (1 and 10 mg/kg per day) accelerated the recovery of the number of white blood cells (WBC) and platelets (PLT) but did not affect the number of circulating red blood cells (RBC). (+) Catechin also augmented the number of myelocytes and splenocytes. Dual color flow cytometric analysis revealed that (+)-catechin reversed the reduction of the population of leukocytes (CD11b+ monocytes, Gr-1+ granulocytes and CD3+ T and CD45RA+ B lymphocytes) in whole blood, spleen and bone marrow caused by 5-FU. (+) Catechin (1 and 10 mg/kg per day) showed remarkable recovery of Gr-1+ cells in all three types of tissues and of CD11b+ cells in the bone marrow cells. These findings suggest that (+)-catechin selectively enhances the recovery of the population of granulocytes reduced by 5-FU in mice. PMID- 15297029 TI - Protective effect of naringin, a bioflavonoid on glycerol-induced acute renal failure in rat kidney. AB - Rhabdomyolysis-induced myoglobinuric acute renal failure accounts for about 10 40% of all cases of acute renal failure (ARF). Reactive oxygen intermediates have been demonstrated to play an etiological role in myoglobinuric renal failure. This study was designed to investigate the effect of naringin, a bioflavonoid with antioxidant potential, in glycerol-induced ARF in rats. Five groups of rats were employed in this study, group I served as control, group II was given 50% glycerol (8 ml/kg, intramuscularly), group III, IV, and V were given glycerol plus naringin 100, 200, and 400mg/kg p.o. route, respectively) 60 min prior to the glycerol injection. Renal injury was assessed by measuring plasma creatinine, blood urea nitrogen, creatinine, and urea clearance. The oxidative stress was measured by renal malondialdehyde levels, reduced glutathione levels, and by enzymatic activity of catalase, glutathione reductase, and superoxide dismutase. Glycerol treatment resulted in a marked renal oxidative stress and significantly deranged the renal functions. Pretreatment of animals with naringin 60 min prior to glycerol injection markedly attenuated renal dysfunction, morphological alterations, reduced elevated thiobarbituric acid reacting substances (TBARS), and restored the depleted renal antioxidant enzymes. These results clearly demonstrate the role of oxidative stress and its relation to renal dysfunction, and suggest a protective effect of naringin in glycerol-induced renal failure in rats. PMID- 15297030 TI - Differential effects of mercury, lead and copper on the constitutive and inducible expression of aryl hydrocarbon receptor (AHR)-regulated genes in cultured hepatoma Hepa 1c1c7 cells. AB - Both simultaneous and sequential exposure to heavy metals and aryl hydrocarbon receptor (AHR)-ligands potentially occur in human populations, yet there have been relatively few studies of combined effects of heavy metals and AHR-ligands on AHR-regulated genes. To investigate the effects of heavy metals on AHR regulated genes; cytochrome P450 1a1 (cyp1a1), NAD(P)H:quinone oxidoreductase (QOR) and glutathione S-transferase Ya (GST Ya), murine hepatoma Hepa 1c1c7 cells were incubated with increasing concentrations of Hg2+ (2.5-10 microM), Pb2+ (10 100 microM), and Cu2+ (1-100 microM) alone or with the AHR-ligand, 2,3,7,8 tetrachlorodibenzo-p-dioxin (0.1 nM), 3-methylcholanthrene (0.25 microM), beta naphthoflavone (10 microM), or benzo[a]pyrene (1 microM). The results clearly showed that metals alone did not significantly alter the cyp1a1 activity and protein levels but increased its mRNA expression, whereas a significant reduction in AHR ligand-mediated induction of cyp1a1 activity was observed by all metals. The decrease in cyp1a1 activity was associated with an increase, no change, or decrease in cyp1a1 mRNA and protein levels by Hg2+, Pb2+ and Cu2+ respectively, suggesting pre- and post-transcription mechanisms are involved. With respect to QOR, the activity and mRNA levels were increased by all metals in the absence or presence of an AHR-ligand, with the exception of Cu2+ which significantly decreased the induction of QOR. Differently, GST Ya activity was significantly increased by Cu2+ and Pb2+ and inhibited by Hg2+, while its mRNA was increased by Hg2+ and Pb2+ and decreased by Cu2+. All metals significantly increased the expression of heme oxygenase-1, which coincided with the changes in the phase I and phase II enzyme activities. These results demonstrate that heavy metals differentially modulate the constitutive and the inducible expression of AHR regulated genes. PMID- 15297031 TI - Abrogation of potassium bromate-induced renal oxidative stress and subsequent cell proliferation response by soy isoflavones in Wistar rats. AB - Potassium bromate (KBrO3) is a potent nephrotoxic agent. In this study, we show the modulatory effect of soy isoflavones on KBrO3-mediated renal oxidative stress and subsequent cell proliferation response in Wistar rats. KBrO3 (125 mg/kg body weight, intraperitoneally) caused reduction in renal glutathione content, activities of renal anti-oxidant enzymes, viz., glutathione peroxidase, glutathione reductase, catalase, glucose-6-phosphate dehydrogenase and phase-II metabolising enzymes such as glutathione-S-transferase and quinone reductase with enhancement in xanthine oxidase, lipid peroxidation, gamma-glutamyl transpeptidase and hydrogen peroxide (H2O2). KBrO3 treatment also induced blood urea nitrogen, serum creatinine and tumor promotion markers, viz., ornithine decarboxylase (ODC) activity and thymidine [3H] incorporation into renal DNA. Treatment of rats orally with soy isoflavones (5 mg/kg body weight and 10 mg/kg body weight) resulted in a significant decrease in xanthine oxidase (P < 0.05), lipid peroxidation, gamma-glutamyl transpeptidase, H2O2 generation, blood urea nitrogen, serum creatinine, renal ODC activity and DNA synthesis (P < 0.001). There was also significant recovery of renal glutathione content (P < 0.01), anti oxidant enzymes and phase-II metabolising enzymes (P < 0.001). Thus, our results show that soy isoflavones acts as potent chemopreventive agent against KBrO3 mediated renal oxidative stress, toxicity and subsequent cell proliferation response in Wistar rats. PMID- 15297032 TI - An examination of quinone toxicity using the yeast Saccharomyces cerevisiae model system. AB - The toxicity of quinones is generally thought to occur by two mechanisms: the formation of covalent bonds with biological molecules by Michael addition chemistry and the catalytic reduction of oxygen to superoxide and other reactive oxygen species (ROS) (redox cycling). In an effort to distinguish between these general mechanisms of toxicity, we have examined the toxicity of five quinones to yeast cells as measured by their ability to reduce growth rate. Yeast cells can grow in the presence and absence of oxygen and this feature was used to evaluate the role of redox cycling in the toxicity of each quinone. Furthermore, yeast mutants deficient in superoxide dismutase (SOD) activity were used to assess the role of this antioxidant enzyme in protecting cells against quinone-induced reactive oxygen toxicity. The effects of different quinones under different conditions of exposure were compared using IC50 values (the concentration of quinone required to inhibit growth rate by 50%). For the most part, the results are consistent with the chemical properties of each quinone with the exception of 9,10-phenanthrenequinone (9,10-PQ). This quinone, which is not an electrophile, exhibited an unexpected toxicity under anaerobic conditions. Further examination revealed a potent induction of cell viability loss which poorly correlated with decreases in the GSH/2GSSG ratio but highly correlated (r2 > 0.7) with inhibition of the enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH), suggesting disruption of glycolysis by this quinone. Together, these observations suggest an unexpected oxygen-independent mechanism in the toxicity of 9,10 phenanthrenequinone. PMID- 15297033 TI - How does peripheral lipopolysaccharide induce gene expression in the brain of rats? AB - Lipopolysaccharide (LPS), the principal cell-wall component of gram-negative bacteria, is responsible for alterations in the central and peripheral tissues associated with gram-negative infections. However, the mechanism by which peripheral LPS cause central effects is not fully known. This study showed that peripheral LPS sequentially increased IL-1beta and iNOS mRNA levels, NO2 level, and CRF mRNA level in the hypothalamic PVN, and corticosterone concentration in blood. Brain-endothelium, but not hypothalamic PVN samples, from LPS injected rats contained ions for LPS lipids, bound BODIPY-LPS (bLPS), and expressed TLR-4, TLP-2 and CD14 mRNAs. This suggests that (1) LPS does not cross the blood-brain barrier, and (2) brain-endothelial cells contain LPS binding sites, TLR-4, TLR-2 and CD14. Systemic LPS injection increased [14C]sucrose uptake, but did not affect [14C]dextran uptake into the brain. Thus, when injected systemically, LPS binds to its receptor and enter the endothelial cells where it increase BBB permeation in a mass-selective manner and triggers a series of signaling events leading to the development of inflammatory response in the brain. PMID- 15297034 TI - Elevated frequency of sister chromatid exchanges of lymphocytes in sarin-exposed victims of the Tokyo sarin disaster 3 years after the event. AB - We previously reported that the frequency of sister chromatid exchanges (SCEs) among victims of the Tokyo subway sarin disaster was significantly higher than that of controls 2-3 months after the disaster. It has been reported that the victims were also exposed to the by-products generated during sarin synthesis, i.e., diisopropyl methylphosphonate (DIMP), diethyl methylphosphonate (DEMP) and N,N-diethylaniline (DEA) during the disaster and we previously found that DIMP, DEMP and DEA induced a significant SCE increase in human lymphocytes in vitro. To monitor the genetic aftereffects of the sarin exposure, SCEs of peripheral blood lymphocytes were measured in fire fighters and police officers involved in the disaster 3 years after the event. We found that the frequency of SCEs was still significantly higher in the exposed subjects than the controls, suggesting a risk of the genetic aftereffects of the sarin exposure. We further found a significant positive correlation between the frequency of SCEs and the inhibition of serum cholinesterase activity in the exposed subjects, suggesting that the elevated frequency of SCEs is related to the sarin exposure. On the other hand, there was no significant difference in natural killer activity between the exposed and the controls. PMID- 15297035 TI - Cytotoxic and oxidative effects induced by man-made vitreous fibers (MMVFs) in a human mesothelial cell line. AB - The introduction of man-made vitreous fibers (MMVFs) as a substitute for asbestos in industrial and residential applications raises concerns about their potential health hazards. The aim of our study was to assess cytotoxic and oxidative effects induced on a human mesothelial cell line (MeT-5A) by exposure to glass wool (GW), rock wool (RW) and refractory ceramic fibers (RCF) in comparison with crocidolite asbestos (CR). MeT-5A cells were exposed for 24 h to 2, 5 and 10 microg/cm2 of MMVF and crocidolite fibers and analysed by scanning electron microscope (SEM) for cell surface alterations. Cells were exposed for 2 h to 1, 2, 5 and 10 microg/cm2 of the same fibers and analysed by enzyme Fpg-modified comet test for direct and oxidative DNA damage. SEM revealed loss of microvilli in cells exposed to RCF and numerous blebs in cells exposed to higher doses of RW. Comet test showed significant direct DNA damage in cells exposed to RCF even at the lowest dose. Comet test with Fpg, that permits the detection of oxided DNA bases, showed significant oxidative DNA damage in cells exposed to higher doses of RW. The presence of DNA damage and alterations of cell surface induced by low doses of RCF and the presence of oxidative DNA damage and blebs on cell surface in cells exposed to higher dose of RW suggest possible cytotoxic, oxidative and genotoxic effects for these MMVFs. PMID- 15297036 TI - D-serine-induced nephrotoxicity: possible interaction with tyrosine metabolism. AB - D-serine selectively damages renal proximal tubule cells in rats by a mechanism that is not fully understood. Recent proteomic analysis identified that D-serine elevated plasma fumarylacetoacetate hydrolase (FAH). FAH is involved in tyrosine catabolism; hence, this pathway may be involved in mediating the toxicity. This work examines whether 2-(2-nitro-4-trifluoromethylbenzoyl)-cyclohexane-1,3-dione (NTBC), a potent inhibitor of the enzyme 4-hydroxyphenylpyruvate dioxygenase (HPPD) located upstream of FAH, modulates D-serine-induced nephrotoxicity. Rats were pretreated with NTBC (0.5 mg/kg p.o.) or corn oil and then 30 min later given either D-serine (250 mg/kg i.p.) or water. Urine was collected every 12 h until termination (48 h) and analysed by 1H NMR spectroscopy and principal component analysis (PCA). Markers of proximal tubule injury were evident in urine following treatment with D-serine and NTBC + D-serine. PCA could not distinguish between these urine samples suggesting that NTBC does not effect the development of nephrotoxicity. Clinical chemistry analysis of urine and terminal plasma samples and histopathological examination of the kidneys confirmed this. NTBC alone caused a marked increase in the excretion of 4-hydroxyphenylpyruvate (HPPA) and 4-hydroxyphenyllactate (HPLA); however, HPPA and HPLA excretion was minimal following NTBC + D-serine. Instead marked tyrosinuria was observed suggesting that D-serine-induced renal damage markedly affects the handling of increased levels of HPPA and HPLA resulting from the inhibition of HPPD. PMID- 15297037 TI - S-adenosylmethionine protects against intrabiliary glutathione degradation induced by long-term administration of cyclosporin A in the rat. AB - We investigate the ability of S-adenosylmethionine (SAMe) to antagonize the cyclosporine A (CyA)-induced inhibition of biliary glutathione efflux induced by long-term administration of CyA (10 mg/kg per day-CyA10 or 20 mg/kg per day-CyA20 for 4 weeks) in rats. CyA treatment reduced the liver content of total glutathione and caused a significant increase in the oxidized-to-reduced glutathione ratio and the thiobarbituric acid-reactive substances (TBARS) concentration. When the rats were concurrently treated with SAMe (10 mg/kg twice daily) and CyA, all these parameters did not significantly differ from control values. Treatment with CyA induced a significant increase in liver GGT activity that was attenuated by coadministration of SAMe. Biliary efflux of total glutathione was significantly reduced in animals treated with CyA. These changes were abolished by SAMe administration. Following inhibition of the intrabiliary catabolism of the tripeptide by acivicin, glutathione efflux rates increased to a lesser extent in animals cotreated with SAMe when compared to those receiving only CyA. The significant decrease in biliary efflux of oxidized glutathione induced by CyA was totally (S + CyA10) or partially (S + CyA20) prevented by coadministration of SAMe. Our observations confirm that SAMe cotreatment in rats antagonizes CyA-induced inhibition in the biliary efflux of glutathione and suggest that protection against intrabiliary glutathione degradation plays a major role in this protective effect. PMID- 15297038 TI - Uranium mill tailings: nuclear waste and natural laboratory for geochemical and radioecological investigations. AB - Uranium mill tailings (UMT) are a high volume, low specific activity radioactive waste typically disposed in surface impoundments. This review focuses on research on UMT and related earth materials during the past decade relevant to the assessment of: (1) mineral hosts of radionuclides; (2) the use of soil analogs in predicting long-term fate of radionuclides; (3) microbial and diagenetic processes that may alter radionuclide mobility in the surficial environment; (4) waste-management technologies to limit radionuclide migration; and (5) the impact of UMT on biota. PMID- 15297039 TI - Pre-assessment of the speciation of 60Co, 125Sb, 137Cs and 241Am in a contaminated aquifer. AB - A sample of contaminated groundwater was analyzed using a combination of wet techniques to obtain geochemical information on the mobile species of 60Co, 125Sb, 137Cs and 241Am. The techniques were combined in a scheme to determine the predominant character of the radionuclides in negative or positive fractions, size separation by ultrafiltration, and their association with natural organic matter (NOM). The analyses indicated that the radionuclides of interest were predominantly in the negative fraction. Most of the 60Co and 125Sb were in the small size fraction (<5000 Da), and 137Cs and 241Am were found with the larger, colloidal-sized material. Antimony-125 and 60Co were predominantly in the hydrophilic fraction, while 137Cs and 241Am were found in hydrophobic fractions. Our analysis indicated that 137Cs is found in the same fraction as the large sized colloidal (hydrophobic) material, suggesting an association with NOM. The results suggested that 60Co and 241Am were associated with NOM, in different size fractions, suggesting that these two nuclides are bound to different sites. Finally, the 125Sb results were inconclusive, whether this nuclide is associated with NOM, or it is inorganic. PMID- 15297040 TI - Erosion of atmospherically deposited radionuclides as affected by soil disaggregation mechanisms. AB - The interactions of soil disaggregation with radionuclide erosion were studied under controlled conditions in the laboratory on samples from a loamy silty-sandy soil. The fate of 134Cs and 85Sr was monitored on soil aggregates and on small plots, with time resolution ranging from minutes to hours after contamination. Analytical experiments reproducing disaggregation mechanisms on aggregates showed that disaggregation controls both erosion and sorption. Compared to differential swelling, air explosion mobilized the most by producing finer particles and increasing five-fold sorption. For all the mechanisms studied, a significant part of the contamination was still unsorbed on the aggregates after an hour. Global experiments on contaminated sloping plots submitted to artificial rainfalls showed radionuclide erosion fluctuations and their origin. Wet radionuclide deposition increased short-term erosion by 50% compared to dry deposition. A developed soil crust when contaminated decreased radionuclide erosion by a factor 2 compared to other initial soil states. These erosion fluctuations were more significant for 134Cs than 85Sr, known to have better affinity to soil matrix. These findings confirm the role of disaggregation on radionuclide erosion. Our data support a conceptual model of radionuclide erosion at the small plot scale in two steps: (1) radionuclide non-equilibrium sorption on mobile particles, resulting from simultaneous sorption and disaggregation during wet deposition and (2) later radionuclide transport by runoff with suspended matter. PMID- 15297041 TI - Lake fish as the main contributor of internal dose to lakeshore residents in the Chernobyl contaminated area. AB - Two field expeditions in 1996 studied 137Cs intake patterns and its content in the bodies of adult residents from the village Kozhany in the Bryansk region, Russia, located on the shore of a drainless peat lake in an area subjected to significant radioactive contamination after the 1986 Chernobyl accident. The 137Cs contents in lake water and fish were two orders of magnitude greater than in local rivers and flow-through lakes, 10 years after Chernobyl radioactive contamination, and remain stable. The 137Cs content in lake fish and a mixture of forest mushrooms was between approximately 10-20 kBq/kg, which exceeded the temporary Russian permissible levels for these products by a factor of 20-40. Consumption of lake fish gave the main contribution to internal doses (40-50%) for Kozhany village inhabitants Simple countermeasures, such as Prussian blue doses for dairy cows and pre-boiling mushrooms and fish before cooking, halved the 137Cs internal dose to inhabitants, even 10 years after the radioactive fallout. PMID- 15297042 TI - Health risk assessment for uranium in Korean groundwater. AB - We analyzed the radiological and chemical risks of uranium in groundwater. The total sample number over 4 years was 498. There were several use patterns of groundwater in Korea, but we considered the risk only for drinking water. The geometric mean of uranium concentration in 10 areas in Korea was 0.17 microg x l( 1). The excess cancer risks were in the 10(-7) level in the radiological risk aspect and the hazard quotient was 0.005 in the chemical risk aspect. Therefore, we could conclude that an adverse health risk is unlikely to be posed due to exposure to uranium. However, the concentration of uranium must be monitored periodically and adequate action taken in the few and small areas that contain high uranium levels in groundwater. PMID- 15297043 TI - Distribution of fallout radionuclides (7Be, 137Cs, 210Pb and 239,240Pu) in soils of Taiwan. AB - Depth profiles and cumulative deposition of four fallout radionuclides (7Be, 137Cs, 210Pb and 239,240Pu) were determined in presumably undisturbed soils in Taiwan. Inventories of these radionuclides in different areas correlate significantly with each other (except 7Be) and with mean annual rainfall, providing a necessary condition for the development of soil erosion studies in Taiwan. However, the data show very large spatial variability between and within landscape units, reflecting the steep topographic and meteorological gradients in the island. Thus, the application of fallout radionuclides to study soil conservation in Taiwan is expected to be a demanding task; it will call for dense sampling even at undisturbed reference sites. PMID- 15297044 TI - Systematic review of the effects of pertussis vaccines in children. PMID- 15297045 TI - Therapeutic immunization with an inactivated HIV-1 Immunogen plus antiretrovirals versus antiretroviral therapy alone in asymptomatic HIV-infected subjects. AB - To determine whether the addition of an inactivated-gp120-depleted HIV-1 Immunogen to antiretrovirals (ARTs) conferred a beneficial effect on delaying time to virologic failure relative to that obtained by ARTs alone, a phase II clinical trial was performed in 243 asymptomatic, ART naive, HIV-1 seropositive adults. The Cox model showed that HIV-1 Immunogen treatment was associated with a 34% decrease in the risk of virologic failure (P = 0.056). When the analysis incorporated baseline HIV-RNA stratification the risk of virologic failure in the HIV-1 Immunogen Arm was significantly reduced a 37% compared to the IFA placebo Arm (P = 0.034). The data suggest that therapeutic immunization plus ARTs could influence virologic control. PMID- 15297046 TI - Vaccination of high-risk patients against influenza: impact on primary care contact rates during epidemics. Analysis of routinely collected data. AB - A general practice (GP) based retrospective cohort study was conducted to assess the effects of influenza vaccination on the primary care contact rate during influenza epidemics. Given the rising workload of family physicians, particularly due to ageing of the population, it is very relevant to know to whether influenza vaccination of high-risk patients reduces the contact rate during epidemics. No effect of vaccination was found on the contact rate of GP during a mild epidemic period. During a 'normal' influenza epidemic, the workload was reduced through fewer contacts by patients with cardiovascular or diabetic diseases. Epidemic periods severe enough to show contact rate reduction occurred approximately every other year. PMID- 15297047 TI - Preclinical study of influenza virus A M2 peptide conjugate vaccines in mice, ferrets, and rhesus monkeys. AB - A universal influenza virus vaccine that does not require frequent updates and/or annual immunizations will offer significant advantages over current seasonal flu vaccines. The highly conserved influenza virus A M2 membrane protein has been previously suggested as a potential antigen target for such a vaccine. Here, we report systematic evaluation of M2 peptide conjugate vaccines (synthetic peptides of M2 extracellular domain conjugated to keyhole limpet hemocyanin (KLH) or Neisseria meningitidis outer membrane protein complex (OMPC)) in mice, ferrets, and rhesus monkeys. The conjugate vaccines were highly immunogenic in all species tested and were able to confer both protection against lethal challenge of either H1N1 or H3N1 virus in mice and reduce viral shedding in the lower respiratory tracts of mice and ferrets. The protection against lethal challenge in mice could also be achieved by passive transfer of monkey sera containing high M2 antibody titers. In addition, we showed that M2 antisera were cross reactive with M2 peptides derived from a wide range of human influenza A strains, but they failed to react with M2 peptides of the pathogenic H5N1 virus (A/Hong Kong/97). The data presented here will permit better understanding of the potential of an M2-based vaccine approach. PMID- 15297048 TI - A phase I study to evaluate a human papillomavirus (HPV) type 18 L1 VLP vaccine. AB - Human papillomavirus (HPV) infection can cause genital warts and cervical cancer. HPV types 6 and 11 cause >90% of genital wart cases; HPV16 and 18 cause 70% of cervical cancers. A prophylactic HPV (types 6, 11, 16, 18) L1 virus-like particle (VLP) vaccine may substantially reduce the incidence of these lesions. This report describes the results of a phase I study of the HPV18 component of such a vaccine. Forty women were randomized to receive either HPV18 L1 VLP vaccine or placebo. Anti-HPV18 responses were measured using a competitive radioimmunoassay (cRIA). Tolerability was evaluated using vaccination report cards (VRC). The study showed that the HPV18 L1 VLP vaccine was generally well-tolerated and highly immunogenic. Peak anti-HPV18 geometric mean titers (GMT) in vaccines were 60-fold greater than those observed in women following natural HPV18 infection. Further studies of a multivalent HPV L1 VLP vaccines are warranted. PMID- 15297049 TI - PorA-specific differences in antibody avidity after vaccination with a hexavalent Men B outer membrane vesicle vaccine in toddlers and school children. AB - A clinical phase II trial with an experimental hexavalent outer membrane vesicle (OMV) vaccine (HexaMen) containing six different porin A (PorAs) was carried out in toddlers (2-3 years) and schoolchildren (7-8 years) in The Netherlands. HexaMen exists of two OMVs each containing three different PorA types. The serum bactericidal activity (SBA) after vaccination against the six PorAs was significantly different and was higher in toddlers than in schoolchildren. After vaccination the SBA against P1.5-2,10 was 4-6 times higher than against P1.7-2,4. The aim of this study was to test whether the differences in SBA could be explained by a difference in subtype-specific antibody avidity maturation. The avidity index (AI) of antibodies against three subtypes (PorA types P1.5-2,10; P1.12-1,13 and P1.7-2,4) was measured by ELISA and evaluated in relation to SBA. A significant avidity maturation for the 3 PorA subtypes was found. This maturation was most pronounced for P1.5-2,10 (mean AI = 72%), correlating with the highest SBA titres. Generally, the avidity titre correlated best with SBA. No differences in avidity indices against the three tested PorAs were found between toddlers and school children indicating that avidity maturation induced by this vaccine is not age-dependent. PMID- 15297050 TI - Oral polio vaccination and low case fatality at the paediatric ward in Bissau, Guinea-Bissau. AB - Oral polio vaccine (OPV) and diphtheria-tetanus-pertussis (DTP) vaccines are given simultaneously in routine immunisation programmes in developing countries. It is therefore difficult to determine the separate effects of these vaccines on survival. We used the shortage of DTP vaccine in Bissau to examine the impact of OPV on the case fatality at the paediatric ward in Bissau. For 719 children less than 5 years of age whose vaccination card had been seen at admission and who had not yet received measles vaccine, having received OPV only was associated with a case fatality of 6% compared with 15% for children having received combined DTP and OPV vaccinations, the case fatality ratio (CFR) being 0.29 (95% confidence interval (CI) 0.11-0.77). Even if children fleeing the hospital were assumed to have died shortly after leaving the hospital, the case fatality would still be lower for children having received OPV only (CFR = 0.41; (95% CI 0.20-0.81)). The tendency was similar for children hospitalised with pneumonia, diarrhoea, and presumptive malaria. Control for background factors had no impact on the estimate. In areas with high mortality, OPV administered alone may have non specific beneficial effects or DTP may have a negative effect for children who had received both DTP and OPV. PMID- 15297051 TI - Interfering vaccine: a novel antiviral that converts a potentially virulent infection into one that is subclinical and immunizing. AB - Infection with human influenza A virus can reach cataclysmic levels, with 40 or more million deaths arising from the 1918 pandemic. Preventative and therapeutic measures have improved since that time, but new approaches are needed. Here, we describe one such new approach--the interfering vaccine, which has two activities -it prophylactically prevents influenza, and at the same time converts an otherwise lethal infection into one that is avirulent and immunizing. Mice treated in this way develop a solid immunity that protects them against a subsequent challenge with homologous virus, and to a lesser extent from challenge with heterologous influenza A viruses. The interfering vaccine comprises non infectious, defective interfering (DI) influenza A virus. Prophylaxis is mediated directly by DI RNA, and results from interference with the replication of the infecting virus. However, interference is incomplete, and there is sufficient wild-type virus multiplication to stimulate a virus-specific natural immunity. As the replication mechanism is common to all influenza A viruses, an interfering vaccine should protect from, and permit immunity to be developed to all influenza A viruses. Indeed, we demonstrate protection against two viruses with antigenically unrelated HA and NA proteins. Thus an interfering vaccine, unlike the conventional vaccine, is independent of the antigenicity of the infecting virus. In principle, interfering vaccines derived from other virus systems could also be developed. PMID- 15297052 TI - Characteristics of Vibrio cholerae proteinases: potential, candidate vaccine antigens. AB - Vibrio cholerae extracellular proteinases (proteases) have been studied as potential candidate antigens for acellular cholera vaccines. Proteinases from V. cholerae NCTC 10732 were prepared from batch culture either by ammonium sulphate precipitation and G100 Sephadex gel filtration or by isoelectric focusing (IEF). Proteinase activity was at a maximum level after 24 h, coincident with the late exponential phase and early stationary phase. Three major IEF peaks of activity were resolved with specific activities in the range 17.2-195 EU ml(-1 )mg(-1). Sodium dodecyl sulphate-polyacrylamide gel electrophoreses (SDS-PAGE) of these fractions revealed 42, 45, 57 and 75 kDa bands in which proteinase activity was demonstrable. Peptide digest analysis suggested different catalytic specificities for each proteinase fraction. Metalloproteinase and serine proteinase inhibitors, alpha(2)-macroglobulin (alpha(2)-M), the thiol proteinase inhibitor and N ethylmaleimide inhibited the proteinases. The proteinases nicked Escherichia coli heat-labile toxin to yield catalytically active sub-units, confirmed by the measurement of intrinsic ADP-ribosylation activity. The possible value of these putative V. cholerae antigens in an acellular vaccine is discussed. PMID- 15297053 TI - An adenoviral type 5 vector carrying a type 35 fiber as a vaccine vehicle: DC targeting, cross neutralization, and immunogenicity. AB - Substituting the coat proteins of adenoviral vector serotype 5 (Ad5) can alter vector tropism and circumvent vector neutralization. Here we report that an Ad5 vector carrying a part of the fiber molecule of human subgroup B adenovirus serotype 35 (Ad5.Fib35) transduces cultured human dendritic cells (DC) and circulating myeloid derived DC with approximately 10-fold greater efficiency than Ad5 in vitro. The improved DC transduction results in increased T-cell activation ex vivo. In vivo however, immunogenicity of the vectors in mice and non-human primates did not correlate with in vitro DC tropism. Ad5.Fib35 was less immunogenic in monkeys than Ad5, despite the improved primate DC tropism of Ad5.Fib35. In mice with high Ad5 vector-specific immunity, Ad5.Fib35 showed no significant difference in anti-insert immunity over Ad5 indicating that fiber exchange alone does not evade pre-existing Ad5 immunity. We thus conclude that, for ex vivo vaccination, Ad5.Fib35 shows promise as vector for loading of DC but is unable to circumvent anti-Ad5 immunity limiting its in vivo utility. PMID- 15297054 TI - Very small size proteoliposomes derived from Neisseria meningitidis: an effective adjuvant for Th1 induction and dendritic cell activation. AB - Recent findings about pathogens and innate immune system interactions have opened new opportunities for adjuvants designs. We have elaborated a new approach, in which gangliosides are incorporated into the outer membrane complex of Neisseria meningitidis (Nm) to form very small size proteoliposomes (VSSP). VSSP, used as monotherapy, demonstrated a unique ability to render immunogenic highly tolerated gangliosides. These results drove our attention to the immunopotentiating properties of VSSP. Here, we examined the VSSP adjuvant effect on the humoral and cellular responses, dendritic cell (DC) activation, and differentiation of Th cells. Also, the role of LPS in VSSP effect was dissected. This study reveals that VSSP is a potent adjuvant for dendritic cells activation and Th1 differentiation. PMID- 15297055 TI - Immunostimulatory oligodeoxynucleotides are potent enhancers of protective immunity in mice immunized with recombinant ORFF leishmanial antigen. AB - Unmethylated CpG dinucleotides in bacterial DNA or synthetic oligonucleotides (ODN) have proved as promising adjuvants for promotion of T helper 1 (Th1) type immune response. The potent Th1 like immune activation by CpG-ODNs suggests a possible utility for vaccination against leishmaniasis. We therefore investigated the effect of ODN containing immunostimulatory CG motifs as adjuvant with recombinant ORFF (rORFF) leishmanial antigen. BALB/c mice were vaccinated with the rORFF with or without CpG-ODN as adjuvant and then challenged with Leishmania donovani metacyclic promastigotes. Administration of CpG-ODN alone resulted in partial protection against challenge with L. donovani in BALB/c mice. Combination of rORFF and CpG-ODN showed enhanced reduction in parasite load (84%) when compared to rORFF (56%) vaccinated mice. Immunization with rORFF alone did not induce the typical Th response whereas co-administration of rORFF with CpG-ODN resulted in enhanced production of immunoglobulin G2a and interferon gamma. Our results further demonstrate that CpG-ODN alone or in combination with rORFF resulted in a dose dependent increase of nitric oxide production in activated macrophages. These studies suggest that CpG-ODN are promising immune enhancers for vaccination against visceral leishmaniasis. PMID- 15297056 TI - Recombinant cholera toxin B subunit (rCTB) as a mucosal adjuvant enhances induction of diphtheria and tetanus antitoxin antibodies in mice by intranasal administration with diphtheria-pertussis-tetanus (DPT) combination vaccine. AB - Recombinant cholera toxin B subunit (rCTB) which is produced by Bacillus brevis carrying pNU212-CTB acts as a mucosal adjuvant capable of enhancing host immune responses specific to unrelated, mucosally co-administered vaccine antigens. When mice were administered intranasally with diphtheria-pertussis-tetanus (DPT) combination vaccine consisting of diphtheria toxoid (DTd), tetanus toxoid (TTd), pertussis toxoid (PTd), and formalin-treated filamentous hemagglutinin (fFHA), the presence of rCTB elevated constantly high values of DTd- and TTd-specific serum ELISA IgG antibody titres, and protective levels of diphtheria and tetanus toxin-neutralizing antibodies but the absence of rCTB did not. Moreover, the addition of rCTB protected all mice against tetanic symptoms and deaths. DPT combination vaccine raised high levels of serum anti-PT IgG antibody titres regardless of rCTB and protected mice from Bordetella pertussis challenge. These results suggest that co-administration of rCTB as an adjuvant is necessary for induction of diphtheria and tetanus antitoxin antibodies on the occasion of intranasal administration of DPT combination vaccine. PMID- 15297057 TI - A murine model for the study of immune memory in response to pneumococcal conjugate vaccination. AB - We developed a murine model for assessment of immunological memory and antibody induced protection to nasopharyngeal (NP) challenges. BALB/c female mice (n = 10 mice per study parameter) were immunized with two priming doses of the licensed 7 valent pneumococcal (Pnc) conjugate vaccine and immune responses [antibody immunoglobulin G (IgG) levels, avidity and opsonophagocytic activity] were monitored for 26 weeks until IgG levels decreased to nearly baseline. A booster dose of either 2 microg conjugate or 5 microg polysaccharide vaccine was given at week 26. The ability of these two treatments to recall immune memory established by the conjugate vaccine was determined for types 4 and 14 for up to 63 days post booster. The ability of challenge with pneumococcal type 14 to recall the immune response was also evaluated, as well as, the number of antibody secreting cells (ASC) specific to polysaccharide (Ps) 4, 6B, and 14. A higher dose of conjugate vaccine (2 microg) was necessary to elicit a significant increase in IgG levels after priming with one dose. Priming with lower doses (0.5 and 1.0 microg) only elicited modest increases in IgG levels. Recall of the immune response was found with either conjugate or Ps vaccines. NP challenge with type 14 at week 26 did not recall the immune response, although reduction in NP Pnc load was seen post primary immunization at 5, 10 and 26 weeks. ASCs were detected in response to either conjugate or Ps booster doses. This model allows for the screening and determination of potential alternative vaccination regimens and the study of immunological markers of memory following Pnc vaccination. PMID- 15297058 TI - A candidate vaccine based on the hepatitis C E1 protein: tolerability and immunogenicity in healthy volunteers. AB - The tolerability and immunogenicity of the hepatitis C virus E1 protein as a candidate vaccine was examined in a Phase I, single-arm study. Twenty healthy male volunteers were injected in the deltoid muscle at weeks 0, 3 and 6 with 20 microg recombinant E1 adsorbed on alum. A fourth (booster) dose was administered to 19 subjects at week 26. The candidate therapeutic vaccine was well tolerated. Three vaccine doses induced a clear humoral anti-E1 response that was boosted by a fourth dose. A strong, specific cellular immune response towards E1 was elicited in all vaccine recipients, which included a clear Th1 type response in all but one of the subjects. PMID- 15297059 TI - Reactions after pneumococcal vaccine alone or in combination with influenza vaccine. AB - We studied adverse reactions to immunisation in 541 individuals receiving simultaneous pneumococcal and influenza vaccination, and in 320 recipients of pneumococcal vaccine alone. Five days after immunisation, the participants completed a questionnaire covering systemic and local reactions to vaccination. Adverse effects were rated as mild if they did not interfere with the participant's daily activities, and moderate or serious if they moderately or markedly restricted these activities. There were no differences between the groups regarding general malaise, headache, myalgias and elevated body temperature. Redness at the injection site, but not soreness or swelling, occurred more frequently in individuals immunised simultaneously with both vaccines. Except for fever and local swelling, adverse reactions were significantly more frequent in women than in men in study groups. The rate of adverse reactions was higher in individuals less then 65 years of age than in older participants. Local reactions were reported by 358 (41.6%) participants, but they were mild and soon subsided. No serious reactions were reported. pneumococcal and influenza vaccine can be safely administered simultaneously. PMID- 15297060 TI - Expression kinetics of the interleukin-2/immunoglobulin (IL-2/Ig) plasmid cytokine adjuvant. AB - The development of strategies to augment the immunogenicity of plasmid DNA vaccines is critical for improving their clinical utility. One such strategy involves the coadministration of plasmid cytokine adjuvants with DNA vaccines. Although a large number of plasmid cytokines have shown promise as adjuvants in preclinical animal models, little is known about their expression kinetics and mechanism of action. We have previously shown that administration of a plasmid encoding the interleukin-2/immunoglobulin (IL-2/Ig) cytokine fusion protein durably augmented DNA vaccine-elicited immune responses in rhesus monkeys for over 10 months. We sought to determine whether persistent cytokine expression from this plasmid accounted for these long-lasting effects. In fact, we found that expression from plasmid IL-2/Ig was transient with an extinction half-life in vivo of approximately 2 days. We next assessed whether the generation of anti cytokine antibodies may have accounted for these transient expression kinetics. Importantly, both mice and rhesus monkeys inoculated with this plasmid cytokine did not develop detectable antibody responses against IL-2. These data suggest that the durable augmentation of DNA vaccine-elicited cellular immune responses afforded by this plasmid cytokine was likely due to enhanced initial priming of memory T lymphocytes rather than chronic cytokine expression. PMID- 15297061 TI - Intravaginal immunization with viral subunit protein plus CpG oligodeoxynucleotides induces protective immunity against HSV-2. AB - Although the genital tract has been considered a poor inductive site for immunization with non-replicating antigens, genital immunization may be important for protection against sexually transmitted infections. Recently, we and others showed that CpG oligodeoxynucleotides (ODNs) serve as potent adjuvants for mucosal immunization. The purpose of this study was to determine whether intravaginal (IVAG) immunization with recombinant glycoprotein B (rgB) of herpes simplex virus type 2 (HSV-2) plus CpG ODN can induce specific immunity and protect against genital HSV-2 challenge. C57BL/6 mice were immunized IVAG with rgB plus CpG ODN, rgB plus non-CpG ODN, or rgB alone and challenged IVAG with HSV 2. Mice immunized with rgB + CpG had higher levels of anti-gB IgA and IgG in the vaginal washes and serum compared to mice immunized with rgB alone. Mice immunized with rgB + CpG also had the highest levels of gB-specific IgG in the nasal washes, however no specific IgA was detected in the nasal washes of any group. Mice immunized IVAG with rgB + CpG showed higher survival and lower pathology scores following genital HSV-2 challenge than mice immunized with rgB + non-CpG ODN or rgB alone. Additionally, vaginal viral titers were lower in the rgB + CpG group after infection. These results clearly show that the genital tract is capable of generating a protective immune response after local intravaginal immunization and that a non-replicating antigen is able to induce such a response when administered with an appropriate adjuvant. PMID- 15297062 TI - Full-length HIV-1 Tat protein necessary for a vaccine. AB - AIDS vaccines now use a truncated version of 86 residues of the Tat protein related to the HIV-1 HXB2 strain predominant in Europe and North America. We compared antibodies raised in rabbits using a B subtype short Tat HXB2(86) and a full-length Tat HXB2(100). Serum against HXB2(86) recognizes only B and D subtypes while serum against HXB2(100) recognizes B, D, and C subtype variants. Conformational epitopes appear to be involved in the capacity of anti-Tat HXB2 sera to recognized non-homologous Tat variants. A linear B-epitope identified in sequence 71-81 in HXB2(86) disappears in HXB2(100), which has a new linear B epitope identified at the C-terminus. Anti-HXB2(100) serum has a higher titer in neutralizing antibody against homologous and non-homologous variants compared to anti-HXB2(86) serum. We suggest that a Tat vaccine should contain a Tat variant with regular size, up to 99-101 residues now found in the field. PMID- 15297063 TI - Intranasal immunization with outer membrane protein P6 and cholera toxin induces specific sinus mucosal immunity and enhances sinus clearance of nontypeable Haemophilus influenzae. AB - Nontypeable Haemophilus influenzae (NTHi) is one of the leading pathogens in sinusitis. One of the outer membrane proteins of NTHi, P6, is a common antigen to all strains and is an attractive candidate for a subunit bacterial vaccine. In this study, we characterized normal sinus mucosa (SM) and investigated the potential of intranasal immunization with P6 and cholera toxin (CT) for induction of mucosal protective immunity against NTHi in the maxillary sinuses of rats. Intranasal immunization induced P6-specific sinus mucosal and systemic immunological responses, mainly of the IgA and IgG isotype. The protective effect of intranasal immunization was demonstrated by enhancement of sinus clearance of NTHi. The present study showed that unilateral intranasal immunization has a capacity to induce protective immunity against NTHi in the bilateral maxillary sinuses. Systemic administration of the vaccine did not affect sinus clearance of NTHi. These findings suggest that a nasal vaccine might be useful for preventing sinusitis. PMID- 15297064 TI - Ethical principles for collective immunisation programmes. AB - Ethical issues arise in discussion of both the content and implementation of collective immunisation programmes. In this paper we propose and discuss seven principles that may guide reflection and debate in this controversial area. Whilst this paper is not intended to be a final and complete account of the relevant principles for collective immunisation programmes we hope that it can help stimulate more active discussion of these issues. Debate about these principles may help to make moral conflicts more explicit and open up the possibility of resolution. We argue that analysis and discussion of the ethical issues should be part of any justification of collective vaccination programmes. PMID- 15297065 TI - Aluminum hydroxide adjuvant induces macrophage differentiation towards a specialized antigen-presenting cell type. AB - Aluminum hydroxide (AlOOH) has been used for many years as a vaccine adjuvant, but little is known about its mechanism of action. We investigated in this study the in vitro effect of aluminum hydroxide adjuvant on isolated macrophages. We showed that AlOOH-stimulated macrophages contain large and persistent intracellular crystalline inclusions, a characteristic property of muscle infiltrated macrophages described in animal models of vaccine injection, as well as in the recently described macrophagic myofasciitis (MMF) histological reaction in humans. AlOOH-loaded macrophages exhibited phenotypical and functional modifications, as they expressed the classical markers of myeloid dendritic cells (HLA-DR(high)/CD86(high)/CD83(+)/CD1a(-)/CD14(-)) and displayed potent ability to induce MHC-II-restricted antigen specific memory responses, but kept a macrophage morphology. This suggests a key role of macrophages, in the reaction to AlOOH adjuvanted vaccines and these mature antigen-presenting macrophages may therefore be of particular importance in the establishment of memory responses and in vaccination mechanisms leading to long-lasting protection. PMID- 15297066 TI - Safety and immunogenicity of CPG 7909 injection as an adjuvant to Fluarix influenza vaccine. AB - CPG 7909, a 24-mer B-Class CpG oligodeoxynucleotide (ODN), was tested for safety, tolerability and its ability to augment the immunogenicity of a commercial trivalent killed split influenza vaccine (Fluarix containing A/Beijing/262/95, A/Sydney/5/97 and B/Harbin/7/94; SmithKline Beecham) in a phase Ib blinded, randomized, controlled clinical trial. Sixty healthy volunteers were recruited in two consecutive cohorts of 30 subjects, who were randomly assigned to receive Fluarix plus 1mg CPG 7909 or Fluarix plus saline control (15 subjects each). Vaccines were administered by intramuscular injection on a single occasion with subjects in the first cohort receiving a 1/10th dose of Fluarix and those in the second cohort receiving the full-dose. All safety measures including physical evaluation, laboratory blood assays, and assays for DNA autoimmunity were within normal values except for transient and clinically inconsequential decreases in total white blood cell counts in groups receiving CPG 7909. All vaccines were found to be generally well tolerated with similar frequency and intensity for most adverse reactions for groups receiving CPG 7909 as controls. Exceptions were injection site pain and headache, which were reduced in frequency in subjects receiving the 1/10th Fluarix dose without CpG, compared to the frequency in all other groups. There was a lack of pre-existing immunity, defined as hemagglutinin inhibition (HI) activity < or =20, for all subjects to the influenza strains A/Beijing/262/95 and B/Harbin/7/94 and for some subjects to A/Sydney/5/97. Post vaccination humoral immune responses, as determined 2 and 4 weeks later by assay of HI activity and ELISA to detect antibodies against hemagglutinin (anti-HA) were similar for both full and reduced Fluarix doses but the cellular immune responses (measured as PBMC antigen-specific IFN-gamma secretion) were reduced in the 1/10th Fluarix dose group. Humoral responses were not significantly enhanced by the addition of CPG 7909, except in individuals with pre-existing immunity to A/Sydney/5/97 strain (baseline HI activity titre >20), where there was a trend to higher HI activity with CPG 7909 (P = 0.06). The addition of CPG 7909 to the 1/10th dose of Fluarix did however result in significantly higher levels of IFN gamma secretion from peripheral blood mononuclear cells recovered at 4 weeks and restimulated ex vivo with A/Beijing/262/95 (P = 0.048) and B/Harbin/7/94 (P = 0.0057), restoring these to the level seen with full-dose vaccine. These results suggest that addition of CPG 7909 to Fluarix may allow the use of reduced vaccine doses without reduced immunogenicity. PMID- 15297067 TI - Active immunization against murine TNFalpha peptides in mice: generation of endogenous antibodies cross-reacting with the native cytokine and in vivo protection. AB - New lines of treatment targeting cytokines have been successfully developed recently and are now widely used in therapy. They are based on passive administration of cytokine inhibitors either soluble receptors or mAbs and the major example is TNFalpha in rheumatoid arthritis (RA). Since a few years, our group has developed a novel alternative approach targeting cytokines by using active immunization against biologically inactive but immunogenic cytokine derivatives. In the present work, we present a new aspect of this research, based on immunization against specific cytokine peptides chosen by molecular modelling. We could elicit a significant humoral response against four TNFalpha peptides by active immunization, and show that the Abs generated cross-reacted with the native cytokine with good titers as determined by ELISA. Interestingly, during coimmunization experiments with couples of peptides, one showed a clear immunodominant effect over the other. Overall, we could not show the neutralization of TNFalpha biological activity in vitro by the immunized sera, but it seems that it is not a prerequisite to observe clinical efficacy. Indeed, using the LPS/galactosamine-induced shock, we could demonstrate that one of the four peptides tested conferred a clinical protection. These results validate the feasibility and efficacy of active immunization against cytokine peptides, and confirm that active immunization against cytokines could represent in the future an alternative to passive immunization in many diseases. PMID- 15297068 TI - Adolescent and adult pertussis vaccination: computer simulations of five new strategies. AB - Approximately one million adult pertussis cases occur annually in the US, and infants still die from pertussis. Computer simulations were used to predict the impact of vaccination of children, adults and/or adolescents, and household members of newborns (cocoon strategy). Childhood vaccination greatly reduced cases in children, but increased the incidence in adolescents and adults. Routine adolescent and adult vaccination had a large direct effect, whereas the cocoon strategy had a predominantly indirect effect on young infants. The number needed to vaccinate (NNV) to prevent a case of typical pertussis in the entire population was lowest for the adolescent strategy. The cocoon strategy had the lowest NNV to prevent a case of typical pertussis in young infants. The current vaccination schedule, local epidemiological data, age-specific cost of pertussis cases, and accessibility of the target population will determine which strategy has the highest likelihood of success in achieving the public health goal. PMID- 15297069 TI - A preliminary experiment of absorption of antinuclear antibodies by the hepatitis B vaccine components, in a case of neurolupus. AB - The aim of this work is to propose a method to search for a possible antigenic community between hepatitis B vaccine components and some auto-antibodies produced in rare patients with an autoimmune disease discovered few weeks or months after a hepatitis B vaccine injection. A 12-year-old girl showed symptoms of transverse myelitis 2 months after a hepatitis B vaccine booster injection. Later, high titers of antinuclear antibodies were found and a diagnosis of neurolupus was established. One serum was then mixed with various concentrations of hepatitis B vaccine. The antinuclear antibodies were totally absorbed at the highest concentration of vaccine, but not by the lowest. Anti-HBs specific antibodies were absorbed by all concentrations of vaccine. The results of this single preliminary experiment (which as to be repeated for other patients, and with large control studies) cannot establish a cross relationship. But it seems that this approach requires to be explored with some improvements. It could also incite us to use lower dosages of HBs antigen in the vaccine and to use the subcutaneous route of injection (to get lower titers of antibodies and to allow a slower diffusion of the antigen). PMID- 15297070 TI - Incidence of severe Plasmodium falciparum malaria as a primary endpoint for vaccine efficacy trials in Bandiagara, Mali. AB - Potential endpoints for blood stage malaria vaccine efficacy trials include uncomplicated malaria disease, which is hard to differentiate from other febrile illnesses, and mortality, which requires prohibitively large sample sizes. Strictly defined severe malaria predicts malaria-associated mortality where case fatality rates are known. To assess the suitability of severe malaria as a trial endpoint, we conducted a census in 1999 and measured the incidence of severe malaria from 1999 to 2001 in Bandiagara, Mali. The annual incidence of severe malaria in children <6 years of age was 2.3% (n = 2,284) yielding an estimated sample size of 4,580 for a vaccine trial designed to detect 50% efficacy with 80% power at P = 0.05 with 5% loss to follow-up. A trial using severe malaria as an endpoint in this setting would thus require expanding the study population or the length of the trial. This approach may be useful in assessing the suitability of potential sites for malaria vaccine trials. PMID- 15297071 TI - Generation of reassortant influenza vaccines by reverse genetics that allows utilization of a DIVA (Differentiating Infected from Vaccinated Animals) strategy for the control of avian influenza. AB - Vaccination of poultry with inactivated influenza vaccine can be an effective tool in the control of avian influenza (AI). One major concern of using inactivated vaccine is vaccine-induced antibody interference with serologic surveillance and epidemiology. In the United States, low pathogenicity H5 and H7 subtype AI viruses have caused serious economic losses in the poultry industry. Most of these viruses also have the accompanying N2 subtype and no H5N1 or H7N8 subtype AI viruses have been identified in poultry in the US. In order to allow the Differentiation of Infected from Vaccinated Animals (DIVA) while maintaining maximum efficacy of the vaccine, we generated reassortant viruses by reverse genetics that contained the same H5 and H7 hemagglutinin (HA) gene as the challenge virus, but a heterologous N1 or N8 neuraminidase (NA) gene. In vaccination-challenge experiments in 2-week-old specific pathogen free chickens, reassortant influenza vaccines (rH5N1 and rH7N8) demonstrated similar antibody profiles and comparable protection rates as vaccines prepared with parent H5N2 and H7N2 viruses. Further, we were able to differentiate the sera from infected and vaccinated birds by neuraminidase inhibition test and indirect immunofluorescent antibody assay on the basis of different antibodies elicited by their NA proteins. These results demonstrate the usefulness of a reverse genetics system for the rapid generation of reassortant AI virus that allows utilization of the DIVA strategy for the control of AI infections in poultry. PMID- 15297072 TI - Safety and immunogenicity of intranasal murine parainfluenza virus type 1 (Sendai virus) in healthy human adults. AB - Human parainfluenza virus-type 1 (hPIV-1) is the most common cause of pediatric laryngotracheobronchitis (croup) and results in close to 30,000 US hospitalizations each year. No effective vaccine is available. We examined murine PIV-1 (Sendai virus, SeV) as a live, xenotropic vaccine for the closely related human PIV-1 in a phase I, dose escalation study in healthy adults. Intranasal Sendai virus was uniformly well-tolerated and showed evidence of immunogenicity in three of nine vaccinees despite pre-existing, cross-reactive immunity presumably induced by previous exposure to human PIV-1. Results encourage future trials to evaluate the efficacy of Sendai virus in preventing human PIV-1 infection in infants and children. PMID- 15297073 TI - Long-lasting specific antibodies against CETP induced by subcutaneous and mucosal administration of a 26-amino acid CETP epitope carried by heat shock protein 65 kDa in the absence of adjuvants. AB - The heat shock protein 65 kDa (Hsp65) of Mycobacterium tuberculosis var. bovis was fused with the linear polypeptide epitope of cholesteryl ester transfer protein C-terminal fragment (CETPC) and expressed as soluble protein in Escherichia coli. The fusion protein Hsp65-CETPC was purified by anion exchange column and eluted at 100-130 mM NaCl in 10mM phosphate buffer (pH 8.0), and then used to immunize mice via subcutaneous injection or intranasal delivery in the absence of adjuvants. Antibodies against CETPC were detected in immunized mice sera by enzyme-linked immunosorbent assay (ELISA) and verified by Western blot analysis. Specific antibodies were successfully induced and lasted for more than 12 weeks in animals immunized with the fusion protein via both subcutaneous and intranasal routes even in the absence of adjuvants. Results showed that Hsp65 could be used as a convenient carrier molecule for presenting foreign polypeptide epitopes, such as CETPC, to the immune system in vivo. Antibodies induced by Hsp65-CETPC could partially inhibit the excessive activity of CETP to normal level. Therefore, Hsp65-CETPC might be further developed to a vaccine against atherosclerosis. PMID- 15297074 TI - Prediction of CTL epitopes using QM, SVM and ANN techniques. AB - Cytotoxic T lymphocyte (CTL) epitopes are potential candidates for subunit vaccine design for various diseases. Most of the existing T cell epitope prediction methods are indirect methods that predict MHC class I binders instead of CTL epitopes. In this study, a systematic attempt has been made to develop a direct method for predicting CTL epitopes from an antigenic sequence. This method is based on quantitative matrix (QM) and machine learning techniques such as Support Vector Machine (SVM) and Artificial Neural Network (ANN). This method has been trained and tested on non-redundant dataset of T cell epitopes and non epitopes that includes 1137 experimentally proven MHC class I restricted T cell epitopes. The accuracy of QM-, ANN- and SVM-based methods was 70.0, 72.2 and 75.2%, respectively. The performance of these methods has been evaluated through Leave One Out Cross-Validation (LOOCV) at a cutoff score where sensitivity and specificity was nearly equal. Finally, both machine-learning methods were used for consensus and combined prediction of CTL epitopes. The performances of these methods were evaluated on blind dataset where machine learning-based methods perform better than QM-based method. We also demonstrated through subgroup analysis that our methods can discriminate between T-cell epitopes and MHC binders (non-epitopes). In brief this method allows prediction of CTL epitopes using QM, SVM, ANN approaches. The method also facilitates prediction of MHC restriction in predicted T cell epitopes. PMID- 15297075 TI - Potent immunogenicity of DNA vaccines encoding Plasmodium vivax transmission blocking vaccine candidates Pvs25 and Pvs28-evaluation of homologous and heterologous antigen-delivery prime-boost strategy. AB - Transmission-blocking vaccines target the sexual stages of the malaria parasite and prevent further development within the mosquito vector halting the transmission of the parasite. Zygote/ookinetes are potential targets of antibodies inhibiting oocyst development in the mosquito midgut and rendering mosquitoes non-infectious. DNA vaccine constructs were developed expressing Pvs25 and Pvs28 (Plasmodium vivax zygote/ookinete surface proteins) fused at the amino terminus with tissue plasminogen activator signal peptide. Antibodies produced in mice after immunization with three doses recognized respective antigens in the parasites and in an ELISA, and these antibodies when tested in membrane feeding assay were potent blockers of P. vivax transmission. Co-immunization with Pvs25 and Pvs28 DNA vaccine constructs did not affect the antigen specific antibody responses against individual antigens, and the antibodies remained effective in blocking parasite transmission demonstrating 91-99% reduction in oocyst number in the mosquito midgut. Several combinations of homologous and heterologous antigen delivery prime boost strategy were also evaluated and the results suggested that antibody titers and transmission-blocking activities by the three prime-boost strategies (DNA prime/DNA boost, DNA prime/protein boost, and protein prime/protein boost) were comparable with slightly better immunogenicity of heterologous antigen-delivery prime/boost as compared to DNA/DNA alone. These results demonstrate potent immunogenicity of DNA vaccines encoding Pvs25 and Pvs28 and warrant further evaluation in non-human primates. PMID- 15297076 TI - The effectiveness of pneumococcal polysaccharide vaccines in adults: a systematic review of observational studies and comparison with results from randomised controlled trials. AB - The use of pneumococcal polysaccharide vaccine has remained controversial since licensure, especially in the elderly. Observational studies form much of the evidence base. We conducted a systematic review of observational studies and compared results with those obtained from an earlier review of randomised controlled trials (RCTs). Estimates of protection against invasive disease from observational studies were consistent, homogenous and compatible with sparse information obtained from RCTs. Studies were of moderate quality. From 13 observational studies the estimate of vaccine efficacy against invasive disease was 53% (46-59%) compared with 38% (-4 to 63%) from nine RCTs. Estimates of protection against all-cause pneumonia were based on fewer, heterogeneous studies that were not consistent with the findings from RCTs for this outcome. From five studies combined efficacy was 32% (7-50%) compared with 3% (-16 to 19%) from 13 RCTs. PMID- 15297077 TI - Stable isotope tracing and triglyceride kinetics. PMID- 15297078 TI - Synbiotics to strengthen gut barrier function and reduce morbidity in critically ill patients. PMID- 15297079 TI - Dietary fat, insulin sensitivity and the metabolic syndrome. AB - Insulin resistance is the pathogenetic link underlying the different metabolic abnormalities clustering in the metabolic syndrome. It can be induced by different environmental factors, including dietary habits. Consumption of energy dense/high fat diets is strongly and positively associated with overweight that, in turn, deteriorates insulin sensitivity, particularly when the excess of body fat is located in abdominal region. Nevertheless the link between fat intake and overweight is not limited to the high-energy content of fatty foods; the ability to oxidize dietary fat is impaired in some individuals genetically predisposed to obesity. Insulin sensitivity is also affected by the quality of dietary fat, independently of its effects on body weight. Epidemiological evidence and intervention studies clearly show that in humans saturated fat significantly worsen insulin-resistance, while monounsaturated and polyunsaturated fatty acids improve it through modifications in the composition of cell membranes which reflect at least in part dietary fat composition. A recent multicenter study (KANWU) has shown that shifting from a diet rich in saturated fatty acids to one rich in monounsaturated fat improves insulin sensitivity in healthy people while a moderate alpha-3 fatty acids supplementation does not affect insulin sensitivity. There are also other features of the metabolic syndrome that are influenced by different types of fat, particularly blood pressure and plasma lipid levels. Most studies show that alpha-3 fatty acids reduce blood pressure in hypertensive but not in normotensive subjects while shifting from saturated to monounsaturated fat intake reduces diastolic blood pressure. In relation to lipid abnormalities alpha-3 fatty acids reduce plasma triglyceride levels but in parallel, increase LDL cholesterol. Substitution of unsaturated fat for saturated fat not only reduces LDL cholesterol but contributes also to reduce plasma triglycerides in insulin resistant individuals. In conclusion, there is evidence available in humans indicating that dietary fat quality influences insulin sensitivity and associated metabolic abnormalities. Therefore, prevention of the metabolic syndrome has to be targeted: (1) to correct overweight by reducing the energy density of the habitual diet (i.e., fat intake) and (2) to improve insulin sensitivity and associated metabolic abnormalities through a reduction of dietary saturated fat, partially replaced, when appropriate, by monounsaturated and polyunsaturated fats. PMID- 15297080 TI - Stable isotope tracer dilution for quantifying very low-density lipoprotein triacylglycerol kinetics in man. AB - BACKGROUND & AIM: A number of approaches have been employed in the past to measure very low-density lipoprotein (VLDL) triacylglycerol (TG) kinetics in humans in vivo, varying in the selection of tracer and mode of administration. All, however, make use of labeled TG precursors and more or less complicated mathematical models to derive the kinetic parameters of interest. The aim of the present study was to develop a conceptually straightforward method, based on the traditional tracer infusion technique, for quantifying VLDL-TG production rates in man using stable isotopes. METHOD: Our approach involves ingestion of [U 13C3]glycerol to endogenously label the glycerol in VLDL-TG, plasmapheresis, isolation of the newly 13C-labeled VLDL from plasma, and administration within the next 2-3 days via a primed constant autologous reinfusion. This procedure produces enough tracer for a priming dose plus 2-3 h of infusion. In the physiological conditions examined (basal and hyperglycemic states, fat- and carbohydrate-rich diets), with almost 3-fold ranging VLDL-TG pool sizes, a steady state in plasma VLDL-TG glycerol tracer-to-tracee ratio was readily achieved within 2 h. Consequently, calculations are made according to the isotope dilution principle, thus avoiding assumptions implicit in more complicated models. CONCLUSION: The stable isotope VLDL-TG tracer dilution method offers an alternative and reliable tool for the determination of endogenous VLDL-TG kinetics in man under a variety of metabolic states. PMID- 15297081 TI - Influence of synbiotic containing Lactobacillus acidophilus La5, Bifidobacterium lactis Bb 12, Streptococcus thermophilus, Lactobacillus bulgaricus and oligofructose on gut barrier function and sepsis in critically ill patients: a randomised controlled trial. AB - BACKGROUND & AIMS: Infective complications are a common cause of mortality and morbidity in critically ill patients. Many factors affect sepsis, one of which is gut barrier function. The aim of this study was to determine whether the oral administration of a synbiotic preparation could alter gut barrier function in critically ill patients and thus reduce sepsis. METHODS: A total of 90 patients admitted to an intensive care unit (ICU) were randomised to receive either synbiotic or placebo preparations (45 into each group). The synbiotic preparation consisted of Lactobacillus acidophilus La5, Bifidobacterium lactis Bb 12, Streptococcus thermophilus and Lactobacillus bulgaricus (probiotics) with oligofructose (prebiotic). Gut barrier function was assessed by measurement of intestinal permeability (lactulose/rhamnose test) and culture of nasogastric aspirate on days 1 and 8. All septic complications and mortality were recorded. RESULTS: There were no differences between the groups in terms of age, sex, APACHE II or POSSUM scores. After 1 week of therapy, patients in the synbiotic group had a significantly lower incidence of potentially pathogenic bacteria (43% versus 75%, P = 0.05) and multiple organisms (39% versus 75%, P = 0.01) in their nasogastric aspirates than controls. There were no significant differences between the groups in terms of intestinal permeability, septic complications or mortality. CONCLUSIONS: The administration of synbiotic in critically ill patients favourably altered the microbial composition of the upper gastrointestinal tract but had no effect on intestinal permeability and was not associated with measurable clinical benefit. PMID- 15297082 TI - Prospective evaluation of nutritional status related to body mass indices and outcomes after modified D2 gastrectomy for carcinoma. AB - AIMS: The aim of this study was to examine the perioperative nutritional status, body mass indices (BMI) and nutritional intakes of patients undergoing a modified D2 gastrectomy (preserving pancreas and spleen) for carcinoma to determine whether a relationship exists between the above and outcomes. METHODS: Fifty consecutive patients [median age 71 years, 38 male] with gastric adenocarcinoma were studied prospectively. RESULTS: Seven patients (14%) were obese (BMI > 30 kg/m2), 16 patients (32%) were overweight (BMI > 25 kg/m2), 21 patients (42%) were of normal weight (BMI 20-25 kg/m2), and six patients (12%) were underweight (BMI < 20 kg/m2). Operative morbidity was commoner in underweight patients (33%) when compared with overweight patients (17%, P = 0.391) and patients of normal weight (14%, P = 0.289). Fatal complications, however (two patients, 4%) were confined to overweight patients (P = 0.118). Preoperative serum albumin levels were significantly higher in overweight patients (43 g/dl) compared to underweight patients (34.5 g/dl; P = 0.003), though no correlation was found between patients' serum albumin levels and postoperative morbidity (r = -0.023, P = 0.877). Overweight patients were significantly less likely to achieve their protein requirements postoperatively than underweight patients (P = 0.037). Early enteral feeding contributed to 56% of the median energy requirements and 45% of the median protein requirements on the seventh postoperative day. CONCLUSION: BMI alone is a poor indicator of outcomes after modified D2 gastrectomy for carcinoma. The role of early enteral nutrition in patients undergoing gastrectomy for cancer deserves further evaluation. PMID- 15297083 TI - Ornithine alpha-ketoglutarate supplementation influences motor activity in healthy rats. AB - BACKGROUND: Ornithine alpha-ketoglutarate (OKG) improves nutritional status in malnourished patients. Published and unpublished data suggest OKG may have effects on the central nervous system that may contribute to its action. OBJECTIVE: We investigated the effect of an OKG-enriched diet on behaviour in healthy rats. DESIGN: Thirty male Wistar rats were randomised in three groups: the OKG group was fed for 5 days (D0-D5) at 90% of spontaneous food intake with an OKG-enriched diet (5 g/kg/d). The non-essential amino acids (NEAA) group was fed similarly with a regimen enriched with NEAA (glycine, alanine, histidine and serine) to be isonitrogenous to OKG group. The ad libitum (AL) group had no treatment and was fed ad libitum with a standard regimen throughout. Rats were tested at D4 for motor activity by actimetry, and at D5 first for spontaneous alternation behaviour measured in the Y-maze, and then for exploratory behaviour measured using the open-field test (stressful environment). RESULTS: We found that OKG supplementation enhanced global motricity by actimetry (AL 772 +/- 55, NEAA 811 +/- 54 vs. OKG 966 +/- 24 arbitrary units, P < 0.05) and total numbers of arms visited in the Y-maze (AL 26 +/- 2, NEAA 30 +/- 3 vs. OKG 38 +/- 3, P < 0.05). The lack of any effect of the OKG-enriched diet in the open-field test shows that the enhancement of locomotion activity was most probably not due to an increase in anxiety or fear in the rats. CONCLUSION: An OKG-enriched diet can induce beneficial stimulant effects that may be involved in the mechanism of action of OKG. PMID- 15297084 TI - Clinical outcome and atherothrombogenic risk profile after prolonged wash-out following long-term treatment with high doses of n-3 PUFAs in patients with an acute myocardial infarction. AB - BACKGROUND: Sustained effects following withdrawal of n-3 PUFAs are unknown. METHODS: Clinical outcome [cardiac death, resuscitation, recurrent myocardial infarction (MI) or unstable angina pectoris] was assessed after prolonged wash out following randomised treatment with high-dosed n-3 PUFAs or corn oil for 12 24 months in 300 acute MI patients. Atherothrombogenic risk markers, serum glucose and markers of lipid peroxidation and inflammation were evaluated in 89 out of the 100 last included patients. RESULTS: After a total median observation period of 45 (range 0-53) months no intergroup difference in prognosis was observed for any of the cardiac events. Favourable effects on serum triglycerides and HDL-cholesterol by n-3 PUFAs were lost after washout, but triglycerides decreased in the corn oil as compared to the n-3 group, P < 0.001. The decline in total cholesterol after withdrawal was similar in both groups. No intergroup difference in the change in thiobarbituric acid-malondialdehyde, a marker of lipid peroxidation, ultrasensitive C-reactive protein, homocysteine, glucose or blood platelets was noted at sustained follow-up. CONCLUSION: Clinical outcome was similar in both patient groups, and the atherothrombogenic risk improvement by n-3 PUFAs was lost after prolonged wash-out. Withdrawal did not affect homocysteine, glucose or markers of lipid peroxidation or inflammation. PMID- 15297085 TI - The nasal loop provides an alternative to percutaneous endoscopic gastrostomy in high-risk dysphagic stroke patients. AB - INTRODUCTION: In the management of dysphagic stroke patients, percutaneous endoscopic gastrostomies (PEGs) are frequently sited early due to the failure of nasogastric tube (NGT) feeding, with NGTs becoming displaced in over 58% of cases. PEG insertion is a procedure with significant mortality and morbidity. We adapted a novel technique of securing NGTs (a nasal loop) which is non-invasive, allows successful NG feeding and may avoid the need for PEG placement. AIMS: To show that nasal loops result in improved delivery of enteral nutrition. To compare the outcome and complication rate of nasal loop fed patients with those undergoing PEG feeding. METHODS: A 6 month prospective audit of dysphagic stroke patients who were referred for PEG. All patients who were referred with failed NG feeding within 28 days of presentation were offered a nasal loop. Patients who were 28 days post-stroke had a PEG placed if appropriate. The daily feed intake was monitored before and after nasal loop placement. Complication rates and patient outcomes were documented at 2 week and 3 month follow-up. RESULTS: Nasal loop group: 14 patients had a nasal loop for a median of 15 days. The median daily feed provided was 0% before nasal loop and 100% after. Four patients went on to recover normal swallowing, 4 patients died and 6 later proceeded to PEG. PEG group: Seven patients proceeded direct to PEG, 1 died and 6 were alive and PEG fed at 3 months. There were 6 complications from PEG insertion. No patients recovered normal swallowing. CONCLUSIONS: Nasal loops are safe, well tolerated, and effective at delivering full enteral nutrition. Nasal loops allow time for patients who may recover normal swallowing to do so, and thus avoid a PEG. Nasal loops avoid unnecessary PEG insertion in those with a poor prognosis who will not ultimately survive their initial stroke. PMID- 15297086 TI - Oxidative stress in mothers who have conceived fetus with neural tube defects: the role of aminothiols and selenium. AB - BACKGROUND & AIMS: Methionine metabolic impairment and selenium deficit have been associated to neural tube defects. The relationship between thiol metabolism and selenium is not well known. We assessed the status of aminothiols and selenium, as well as thiolic status and the amino acids involved in arginine synthesis in the case of selenium depletion and repletion, studying their relationship to neural tube defects. METHODS: We studied 44 women of 37 +/- 8 years (mean +/- SD) who had conceived fetuses with neural tube defects as cases; and 181 women of 39 +/- 7 years (mean +/- SD) with healthy children as controls. We determined selenium, vitamin B12, serum folates, plasma thiol compounds and amino acids. Homocysteine transsulfuration was assessed using total cysteine/total homocysteine ratio (tCys/tHcy), and selenium repletion cut-off value was 1.06 micromol/l (84 microg/l). RESULTS: Cases showed significantly lower levels (median) than controls of total homocysteine (P = 0.001), total cysteinylglycine (P < 0.001), selenium (P < 0.001) and tryptophane (P = 0.002); and higher tCys/tHcy levels (P < 0.001), glutathione (P = 0.008) and L-arginine (P = 0.001). Cases with selenium depletion (selenium < or = 1.06 micromol/l) had significantly higher levels than controls of cysteine (P = 0.010), glutathione (P = 0.005), tCys/tHcy (P < 0.001), and arginine (P = 0.004), but significantly lower levels than controls of tryptophane (P = 0.027), cysteinylglycine (P < 0.001) and folates (P < 0.001). Only cysteinylglycine was lower than controls (P < 0.001) when selenium > 1.06 micromol/l. Methionine levels were higher in cases with selenium depletion than in repletion (P = 0.029). CONCLUSIONS: According to our data, a diet deficient in selenium and folates or their absorption impairment, and/or other mechanisms related to polyamines and nitric oxide can lead to oxidant/antioxidant imbalance and to a higher occurrence of these malformations. PMID- 15297087 TI - Methylprednisolone sodium succinate in pediatric parenteral nutrition: influence of vehicle injection. AB - Many drugs can be administered in parenteral nutrient mixtures, but no work on the delivery of corticoids by such means is reported. We studied the influence of pediatric parenteral nutrient mixtures on the kinetic parameters of the corticoid methylprednisolone injected in an intravenous bolus in rabbits as its sodium succinate ester. Four groups of six male New Zealand rabbits were used. After extraction, the plasma drug concentrations were measured by high performance liquid chromatography. The distribution volume of the ester and the clearance rates of the two entities (ester and methylprednisolone) were lowered in the presence of a lipid emulsion. The description of these pharmacokinetic parameters provides a basis for a preclinical study of 24h administration of methylprednisolone in a parenteral nutrient mixture. PMID- 15297088 TI - Is early enteral nutrition a risk factor for gastric intolerance and pneumonia? AB - BACKGROUND: Early enteral nutrition (EN) after injury reduces septic complications, but upper digestive intolerance (UDI) occurring immediately post trauma is a risk factor for pneumonia. Our study aimed to determine whether early intragastric feeding may lead to gastric intolerance and subsequent pneumonia in ventilated multiply injured patients. METHODS: This prospective study involved two groups of patients randomized either to immediate intragastric EN, or to delayed intragastric EN started later than 24 h after admission. UDI was diagnosed when gastric residual volume, measured with a 50-ml syringe after stopping the feeding for 2 h, exceeded 200 ml at least at two consecutive measurements, and/or when vomiting occurred. RESULTS: Out of 52 patients, 27 were included in the early EN group, and 25 in the delayed-EN group. On day 4, the early EN group received a greater amount of feeding because of intolerance problems occurring in the delayed-EN group (1175 +/- 485 ml vs. 803 +/- 545 ml). Twenty-five subjects--33% of the early EN patients and 64% of the delayed-EN patients--met the criteria for pneumonia (P = 0.050). On average, patients with pneumonia were older, more severely injured, and therefore required more ventilator days and a longer stay in the intensive care unit than patients without pneumonia. CONCLUSIONS: If properly administered, early enteral nutrition can decrease the incidence of upper intestinal intolerance and nosocomial pneumonia in patients with multiple injuries. PMID- 15297089 TI - Visual contrast enhances food and liquid intake in advanced Alzheimer's disease. AB - BACKGROUND & AIMS: Patients with severe Alzheimer's disease (AD) in long-term care have deficient contrast sensitivity and poor food and liquid intake. The present study examined how contrast manipulations affect these intake levels. METHODS: Participants were nine men with advanced AD. Independent variables were meal type (lunch and supper) and condition (baseline, intervention, and post intervention). Dependent variables were amount of food (grams) and liquid (ounces). Data were collected for 30 days (10 days per condition) for two meals per day. White tableware was used for the baseline and post-intervention conditions, and high-contrast red tableware for the intervention condition. In a follow-up study 1 year later, other contrast conditions were examined (high contrast blue, low-contrast red and low-contrast blue). RESULTS: Mean percent increase was 25% for food and 84% for liquid for the high-contrast intervention (red) versus baseline (white) condition, with 8 of 9 participants exhibiting increased intake. In the follow-up study, the high-contrast intervention (blue) resulted in significant increases in food and liquid intake; the low-contrast red and low-contrast blue interventions were ineffectual. CONCLUSIONS: Simple environmental manipulations, such as contrast enhancement, can significantly increase food and liquid intake in frail demented patients with AD. PMID- 15297090 TI - Effect of nutritional support on clinical outcome in patients at nutritional risk. AB - BACKGROUND & AIMS: Undernourished patients have an increased risk of complications and a prolonged hospital stay, compared to those who are not undernourished. The aim of this study was to evaluate the effect of nutritional intervention in a random sample of hospitalized patients at nutritional risk. METHODS: A randomized, controlled trial of nutritional intervention in 212 patients. Intervention consisted of a specialized nutritional team (nurse and dietician) who attended patients and staff for motivation, detailed a nutritional plan, assured delivery of prescribed food and gave advice on enteral or parenteral nutrition when appropriate. The control group received the standard regime used in the department. The primary endpoint was the part of the length of stay (LOS) that was considered to be sensitive to nutritional support, designated LOSNDI. The nutritional discharge index (NDI) consists of three objective criteria: (1) the patient is able to manage toilet visits without assistance, reflecting mobilization; (2) the patient is without fever (tp < 38 degrees C), reflecting absence of infection; and (3) the patient has no intravenous access, reflecting absence of complications in general. On the day when all three criteria were fulfilled, hospital stay was no longer considered to be sensitive to nutritional support. Actual LOS is also reported. Incidence and severity of complications were recorded to explain LOSNDI findings. As a secondary endpoint, quality of life was evaluated by the Short Form 36 (SF-36) questionnaire. RESULTS: Intervention led to an intake of > or = 75% of requirements in 62% of the intervention patients, as compared to 36% of the control patients. Rates of complications, mean LOSNDI and LOS were not significantly different between the two study groups. However, among patients with complications a difference in LOSNDI between intervention patients (14 +/- 2 days, mean +/- SE) and control patients (20 +/- 2 days) was statistically significant (P = 0.015). In the same patients, LOS was 17 +/- 2 days in the intervention group and 22 +/- 2 days in the control group (P = 0.028). The SF-36 questionnaire did not show a significant effect of treatment. CONCLUSIONS: Protein and energy intake of nutritionally at risk patients was increased which resulted in shortening of the part of the length of stay that was considered to be sensitive to nutritional support (LOSNDI) and shorter length of stay (LOS) among patients with complications. PMID- 15297091 TI - Value of body mass index in the detection of severe malnutrition: influence of the pathology and changes in anthropometric parameters. AB - BACKGROUND & AIMS: We have estimated the prevalence of severe malnutrition in groups of patients hospitalized for different medical causes and assessed the sensitivity of BMI in the diagnosis of severe malnutrition. DESIGN: A prospective study enrolled 1052 patients: 396 patients with liver cirrhosis including 165 non ascitic patients (NAP), 124 patients with mild ascites (MAP), 107 patients with tense ascites (TAP), 251 patients after cardiac surgery (SCP), 81 patients with cardiac diseases (MCP), 85 patients with stroke (SP), 36 patients with degenerative neurological diseases (DNP), 68 patients after surgery of a hip fracture (HFP), 91 patients with palliative care for cancer (CP) and 44 elderly patients with medical affections (EP). BMI, mid-arm muscular circumference (MAMC) and triceps skinfold thickness (TST) were measured within 48 h after admission. Patients with MAMC and TST below the 5th percentile of a reference population when aged < or = 74 or the 10th percentile when aged > or = 75 were defined as severely malnourished. Sensitivity of BMI < 20 to detect malnutrition was assessed. RESULTS: The prevalence of severe malnutrition was the highest in TAP (39.1%) HFP (25.6%) and MAP (24.3%) and the lowest in SCP (4%), SP (4.8%), DNP (5.7%) and MCP (7.4%) (P < 10(-4)). In multivariate analysis, low TST was associated with female gender (P < 10(-4)) mild and tense ascites (P = 0.038, P = 0.0004), low MAMC with male gender (P < 10(-4)), low BMI with female gender (P = 0.0082), hip fracture (P = 0.0407) and cancer (P = 0.0059). The sensitivity of BMI to detect severe malnutrition was the highest in HFP, CP and EP (100%, 80% and 100% respectively) and the lowest in TAP, MCP and SP (40%, 33.3% and 50% respectively). After exclusion of TAP, sensitivity of BMI to detect malnutrition correlated significantly with the coefficient of correlation between MAMC and TST observed in each group (r = 0.821, P = 0.0066). CONCLUSION: Ascitic cirrhotic patients and elderly patients after surgery of hip fracture had the highest prevalence of severe malnutrition. BMI had the highest sensitivity when both TST and MAMC were damaged to the same extent. BMI < 20 has a high sensitivity in the diagnosis of severe malnutrition in elderly and cancer patients but not in cirrhotic patients with tense ascites, cardiovascular and neurological patients. PMID- 15297092 TI - Effects of arginine supplementation on mucosal immunity in rats with septic peritonitis. AB - BACKGROUND: Supplemental Arginine (Arg) has been demonstrated to improve the immunologic response and reduce mortality in rodents with sepsis. However, the effects of Arg on gut-associated lymphoid tissue function after infection and sepsis are not clear. The aim of this study was to study the effect of Arg supplemented diets before and Arg-enriched total parenteral nutrition (TPN) after sepsis or both on the intestinal immunity of rats with septic peritonitis. METHODS: Rats were assigned to four groups. Groups 1 and 2 were fed a semipurified diet, while in the diets of groups 3 and 4, part of the casein was replaced with Arg. After feeding the experimental diets for 10 days, sepsis was induced by cecal ligation and puncture (CLP); at the same time, the internal jugular vein was cannulated. All rats were maintained on TPN for 3 days. Groups 1 and 3 were infused with conventional TPN, while groups 2 and 4 were given a TPN solution supplemented with Arg, which replaced 10% of the total amino acids. All rats were sacrificed 3 days after CLP. Intestinal immunoglobin (Ig) A levels, total lymphocyte yields, and lymphocyte subpopulations in Peyer's patches were analyzed. In vitro cytokine secretion by splenocytes and Peyer's patch lymphocytes were also measured. RESULTS: Total lymphocyte yields in Peyer's patches, and small intestinal immunoglobulin A (IgA) secretion in group 4 were significantly higher than the groups 1 and 2. No differences were observed between groups 3 and 4. There were no differences in the interleukin (IL)-2 and interferon- gamma levels among all groups when splenocytes were stimulated with mitogen. However, in vitro splenocyte IL-10 production in group 4 was significantly higher than those of groups 1 and 2, and had no difference from group 3. There were no differences in the ratios of B and T lymphocyte subpopulations in Peyer's patches among all groups. CONCLUSIONS: Enteral Arg supplementation before sepsis tended to enhance total lymphocyte yields in Peyer's patches and intestinal IgA secretion. Arg administered both before and after CLP had a synergistic effect on improving intestinal immunity, possibly by enhancing systemic IL-10 secretion. However, intravenous Arg administration after CLP had no favorable effects on mucosal immunity in rats with septic peritonitis. PMID- 15297093 TI - Basal and postprandial substrate oxidation rates in obese women receiving two test meals with different protein content. AB - BACKGROUND & AIMS: Fuel utilisation and storage in lean and obese subjects seem to be differently affected by the macronutrient distribution intake. The aim of this intervention study was to explore the extent to which the fat mass status and the macronutrient composition of an acute dietary intake influence substrate oxidation rates. METHODS: Fuel utilisation in 26 women, 14 obese (BMI = 37.1 +/- 1.1 kg/m2) and 12 lean (BMI = 20.6 +/- 0.5 kg/m2) was measured over 6 h to compare the metabolic effect of a single balanced protein (HC) meal and a high protein (HP) single meal, which were designed to supply similar energy contents (1672 kJ). The macronutrient composition as a percentage of energy of the HC meal was 55% carbohydrate, 15% protein and 30% fat, while the HP meal contained 40% carbohydrate, 30% protein and 30% fat. Nutrient oxidation rates and energy expenditure were calculated by indirect calorimetry (hood system), whereas exogenous amino acid oxidation was estimated from the 13C isotopic enrichment of breath after oral administration of L[1-13C]leucine. RESULTS: Fasting lipid oxidation was higher in the obese than in the lean women (P < 0.05), and it was significantly correlated with body fatness (P < 0.01). A single HP meal consumption produced higher postprandial fat oxidation as compared with HC meal intake (P < 0.02), in both obese and lean subjects, with no apparent changes in glucose oxidation rates. Furthermore, postprandial fat utilisation after the test meal intake was higher in obese than in the lean women (P < 0.01). The time course of 13CO2 in breath followed a similar pattern in both dietary groups, although a non-statistically significant higher trend in protein and 13C-leucine oxidation was found in the HP group. CONCLUSIONS: Net lipid oxidation depends on both short-term dietary composition intake and fat body mass, being significantly higher after a relatively high protein meal as compared to a balanced diet intake and in obese women as compared to lean controls. PMID- 15297094 TI - Glutamine and CXC chemokines IL-8, Mig, IP-10 and I-TAC in human intestinal epithelial cells. AB - BACKGROUNDS & AIMS: Chemokines are a family of small proteins involved in immune and inflammatory responses. Human intestinal epithelial cells act as a sentinel in the immune response and produce CXC chemokines such as IL-8, Mig, IP-10 and I TAC. Glutamine has various effects on immuno-inflammatory response in human intestine. METHODS: The present study aimed to determine the effect of glutamine on the IL-8, Mig, IP-10 and I-TAC production by ELISA and their mRNA level by RT PCR (expressed as % gapdh) in two human intestinal epithelial cell lines Caco 2/TC7 and HCT-8 under basal conditions or during stimulation with combined cytokines. RESULTS: Under basal conditions, studied chemokines were not influenced by glutamine. When intestinal epithelial cells were stimulated with cytokines, increasing concentrations of glutamine from 2 to 10 mM in HCT-8 cells significantly decreased I-TAC and IP-10 mRNA level (respectively 219 to 182%; P < 0.01; 257 to 176%; P < 0.05) and I-TAC and IP-10 production (respectively 21.2 to 13.0; P < 0.05; 696 to 548 ng/prot mg; P < 0.01). Glutamine also reduced IP-10 mRNA level (186 to 135%, P < 0.05) in cytokines-stimulated Caco-2/TC7 cells. CONCLUSIONS: Down-regulation of CXC chemokines by glutamine could contribute to its therapeutic potential in intestinal inflammation and during critical illness. PMID- 15297095 TI - Effects of protein-rich supplementation and nandrolone in lean elderly women with femoral neck fractures. AB - AIM: To evaluate the effects of a protein-rich liquid supplementation, alone or in combination with the anabolic steroid nandrolone decanoate, on body composition, activities of daily living (ADL) status and the health-related quality of life (HRQoL) after a femoral neck fracture. METHODS: Sixty women, aged 83 +/- 5 years (mean +/- SD), BMI < 24 kg/m2 (20.4 +/- 2 kg/m2 ) and capable of co-operating, with a femoral neck fracture treated with internal fixation, were randomised to open treatment during 6 months with a protein-rich liquid formula alone (PR, Fortimel, 200 ml/day, 20 g protein/day) or in combination with nandrolone decanoate (PR/N, Deca-Durabol 25 mg i.m./3 weeks) or to a control group (C). The patients were re-examined after 6 and 12 months regarding body weight (BW), lean body mass (LBM, DXA), ADL status according to Katz, HRQoL according to EQ 5-D and fracture healing. RESULTS: LBM decreased in the C (-1.2 +/- 2 kg) and PR groups (-1.2 +/- 1 kg) but remained the same in the PR/N group (0.3 +/- 1 kg) (P < 0.05 between groups). ADL remained at a high level in the two intervention groups but declined significantly in the C group (P < 0.005 between groups). The decline in HRQoL was least pronounced in the PR/N group at 6 months (P < 0.05 between groups). Patients with fracture healing complications lost more BW (P < 0.05) and LBM (P < 0.01) than patients with uneventful fracture healing. CONCLUSION: Protein-rich liquid supplementation in combination with nandrolone given for 6 months to lean elderly women after a femoral neck fracture may positively affect LBM, ADL and HRQoL. PMID- 15297096 TI - Renal effects of parenteral fish oil administered to heart-beating organ donors and renal-transplant recipients: a tolerance study. AB - BACKGROUND & AIMS: Nutrition can interfere with organ function during the different stages of transplantation. Oral fish oil supplementation to kidney transplant recipients has been found to improve renal function. The aim of the present study was to determine the safety and tolerance of intravenous administration of fish-oil emulsion to heart-beating brain-dead donors and, subsequently, to the kidney recipients, and to assess its effects on renal function. METHODS: A lipid emulsion enriched with omega-3 fatty acids (MLF 541) was given intravenously to 8 heart-beating, brain-dead organ donors for up to 4 h before organ harvesting and to the kidney recipients for 5 days postoperatively. Hemodynamic, biochemistry and hematological parameters were measured before and at the end of lipid administration in the donors and on posttransplantation days 1, 5, 30 and 180 in the recipients. Findings in the recipients were compared with a concurrent control group. RESULTS: There were no significant changes in hemodynamic or laboratory parameters during the MLF infusion in the donors or the 5 days of MLF administration in the recipients. Blood urea nitrogen and serum creatinine levels decreased over time in both the study and control recipients (P < 0.05 for both), with no significant between-group difference at any of the time points studied. CONCLUSIONS: Administration of MLF 541 is safe in organ donors and in kidney recipients. Further studies involving nutrients as pharmacological agents in organ transplantation are warranted. PMID- 15297097 TI - Health effect of vegetable-based diet: lettuce consumption improves cholesterol metabolism and antioxidant status in the rat. AB - BACKGROUND & AIMS: It is often assumed that fruits and vegetables contribute to protect against degenerative pathologies such as cardiovascular diseases. Besides epidemiological observations, scientific evidences for their mechanism of action are scarce. In the present study, we investigated the mean term and post-prandial effects of lettuce ingestion on lipid metabolism and antioxidant protection in the rat. RESULTS: Feeding rats a 20% lettuce diet for 3 weeks resulted in a decrease cholesterol LDL/HDL ratio and a marked decrease of liver cholesterol levels (-41%). Concurrently, fecal total steroid excretion increased (+44%) and apparent absorption of dietary cholesterol was significantly depressed (-37%) by the lettuce diet. Lettuce diet also displayed an improvement of vitamin E/TG ratio in plasma and limited lipid peroxidation in heart as evidenced by TBARS. In post-prandial experiment, lettuce intake significantly increased both ascorbic acid and alpha-tocopherol plasma levels which contribute to improve plasma antioxidant capacity within 2 h of consumption. Other lipid-soluble antioxidants (lutein and vitamin E) may also improve the plasma antioxidant capacity. CONCLUSION: Lettuce consumption increases the total cholesterol end-products excretion and improves antioxidant status due to the richness in antioxidants (vitamins C, E and carotenoids). In our model, lettuce clearly shows a beneficial effect on lipid metabolism and on tissue oxidation. Therefore regular consumption of lettuce should contribute to improve protection against cardiovascular diseases. PMID- 15297098 TI - Improved haemorrheological properties by Ginkgo biloba extract (Egb 761) in type 2 diabetes mellitus complicated with retinopathy. AB - BACKGROUND & AIMS: Abnormal haemorrheological property changes in erythrocyte deformability, plasma and blood viscosity, and blood viscoelasticity may play very important roles in the development of microangiopathies in diabetes mellitus (DM). In this study, we demonstrate the improvement in abnormal haemorrheological parameters in DM with ingestion of Ginkgo biloba extract 761 (Egb 761). METHODS: Haemorrheological parameters before and 3 months after Egb 761 oral ingestion were determined in 25 type 2 DM patients with retinopathy. These parameters included lipid peroxidation stress of erythrocytes, erythrocyte deformability, plasma and blood viscosity, blood viscoelasticity, and retinal capillary blood flow velocity. RESULTS: After taking Egb 761 orally for 3 months, the blood viscosity was significantly reduced at different shear rates, by 0.44 +/- 0.10 (gamma = 400), 0.52 +/- 0.09 (gamma = 150) and 2.88 +/- 0.57 (gamma = 5). Viscoelasticity was significantly reduced in diabetic patients by 3.08 +/- 0.78 (0.1 Hz). The level of erythrocyte malondialdehyde (MDA) was reduced by 30%; however, the deformability of erythrocyte was increased by 20%. And lastly, retinal capillary blood flow rate was increased from 3.23 +/- 0.12 to 3.67 +/- 0.24 cm min(-1). CONCLUSION: In this preliminary clinical study, 3 months of oral administration of Egb 761 significantly reduced MDA levels of erythrocytes membranes, decreased fibrinogen levels, promoted erythrocytes deformability, and improved blood viscosity and viscoelasticity, which may facilitate blood perfusion. Furthermore, it effectively improved retinal capillary blood flow rate in type 2 diabetic patients with retinopathy. PMID- 15297099 TI - Lipid effects on neutrophil calcium signaling induced by opsonized particles: platelet activating factor is only part of the story. AB - BACKGROUND & METHODS: Total parenteral nutrition is frequently used in clinical practice to improve the nutritional status of patients. However, the risk for infectious complications remains a drawback in which immune-modulating effects of the lipid component may play a role. To characterize these lipid effects we investigated neutrophil activation by opsonized yeast particles under influence of lipid emulsions derived from fish oil (VLCT), olive oil (LCT-MUFA), soybean oil (LCT), and a physical mixture of coconut and soybean oil (LCT-MCT). RESULTS: Serum-treated zymosan (STZ) evoked a biphasic increase in cytosolic Ca2+ concentration ([Ca2+]c) with an initial slow rise that turned into a second fast rise until a plateau was reached. LCT-MCT (5 mM) pretreatment markedly increased the rate of [Ca2+]c rise during the initial phase, abolished the second phase and lowered the plateau. These effects of LCT-MCT were mimicked by the protein kinase C (PKC) activating phorbol ester PMA. LCT, LCT-MUFA and VLCT, on the other hand, decreased the rate of [Ca2+]c rise during both phases and lowered the plateau. The platelet-activating factor (PAF) receptor antagonist WEB 2086 inhibited the second phase, demonstrating that PAF acts as an intercellular messenger in STZ induced Ca2+ mobilization, but did not interfere with the stimulatory effect of LCT-MCT or PMA on the initial rate of [Ca2+]c rise. CONCLUSIONS: Structurally different lipids act only in part through PAF to distinctively modulate neutrophil calcium signaling in response to activation by opsonized particles. PMID- 15297100 TI - Glycaemic responses after ingestion of 3 local carbohydrate-based foods in West Indian patients with type-2 diabetes mellitus. AB - BACKGROUND & AIM: Previous studies suggest that inadequate glycaemic control in diabetic patients might be related to the type of carbohydrates the patients consume regularly. Thus, we aimed to assess glucose and insulin responses after diabetic and non-diabetic subjects ingested 3 commonly consumed carbohydrate based foods. METHODS: Thirty-eight type-2 diabetic and 27 non-diabetic subjects were studied in 3 different occasions of 7 days apart. On each day of the study, anthropometric indices were measured and after collecting fasting blood samples, subjects randomly consumed bread, roti or rice within 10 min. Subsequently 7 ml of venous blood samples were collected at 60, 90, 120 and 150 min for determination of glucose and insulin responses. RESULTS: Although the diabetic patients were older than the healthy subjects (P < 0.05), both subjects had similar weight, body mass index and waist and hip circumferences (P > 0.05). The mean fasting and post meal plasma glucose concentrations for the 3 test foods were higher in diabetic patients than the corresponding values for the healthy subjects (all; P < 0.001). Generally, roti elicited the highest total incremental glucose responses in the diabetic patients irrespective of ethnic group (P < 0.05). CONCLUSION: There were variations in glucose and insulin responses to the 3 test foods. However, roti elicited the highest postprandial hyperglycaemia and should therefore be discouraged in regular dietary plan of diabetic patients. PMID- 15297101 TI - Dietary protein precipitation properties have effects on gastric emptying in healthy volunteers. AB - BACKGROUND & AIMS: Strategies that reduce the size of particles in the stomach accelerate gastric emptying. Partial dephosphorylation of casein reduces the size of protein precipitates (curds) in acid conditions and facilitates peptic digestion. We hypothesized that changing the precipitation properties of casein by partial dephosphorylation would accelerate gastric emptying. METHODS: Eight healthy male volunteers entered a prospective, double blind, randomized study with crossover design. Gastric emptying of milk based formula containing either unmodified or dephosphorylated casein was assessed by scintigraphy. Gastric pH measurements were acquired concurrently. RESULTS: A trend to faster gastric emptying was observed for the unmodified preparation, with lower median half time (unmodified 133; dephosphorylated 214 min, P = 0.09) and area under the curve (unmodified 8425 min%; dephosphorylated 9135 min%, P = 0.08). A positive correlation was found between half time for the dephosphorylated preparation and the treatment effect (r2 = 0.81, P < 0.02). Gastric pH was unaffected. CONCLUSIONS: The study hypothesis was rejected; indeed gastric emptying tended to be faster for the unmodified than the dephosphorylated protein. This effect was more pronounced in subjects with slow gastric emptying on the dephosphorylated preparation. Properties other than the size of protein precipitates determine the rate of gastric emptying for milk based formula. PMID- 15297102 TI - Peripheral blood mononuclear cell fatty acid composition and inflammatory mediator production in adult Crohn's disease. AB - BACKGROUND & AIMS: Crohn's disease (CD) is associated with nutritional deficiencies, altered plasma concentrations of polyunsaturated fatty acids (PUFA) and an anti-inflammatory response to fish oil that contains n-3 PUFA. This suggests that, in CD, immune cells may have altered n-3 PUFA composition with functional consequences. The aim of this study is to investigate n-3 and n-6 PUFA composition and synthetic function of peripheral blood mononuclear cells (PBMC) in the basal state. METHODS: A case control study of 52 adult CD patients and healthy, age- and sex-matched controls. Composition of PBMC and plasma phospholipids were measured by gas chromatography and production of tumour necrosis factor-alpha, prostaglandin E2 (PGE2) and interferon-gamma (IFN-gamma) by PBMC were measured by ELISA. RESULTS: CD was associated with higher concentrations of eicosapentaenoic acid and other n-3 PUFA, and lower arachidonic acid (AA) (n-6 PUFA) in PBMC. This was not explained by differences in dietary fat intake. Lower rates of production of PGE2 and IFN-gamma by PBMC were noted in quiescent and active CD, respectively, compared to controls. CONCLUSIONS: CD is associated with a greater availability, and not a deficiency, of n-3 PUFA in PBMC, but lower concentrations of AA, and lower rates of production of PGE2 and IFN-gamma, compared to healthy controls. PMID- 15297103 TI - Simple and accurate assessment of energy expenditure in ventilated paediatric intensive care patients. AB - AIMS: To assess validity and reliability of energy expenditure measurements with a short Douglas bag protocol compared to the standard metabolic monitor in a paediatric intensive care setting. METHODS: 51 paired measurements were performed in 14 ventilated patients (age 0-18 years) with sepsis, trauma or following major surgery. Measured data were compared mutually and compared to Schofield equations using Bland-Altman analysis. RESULTS: Comparing Douglas bag (3.21 +/- 1.43 MJ/day) and metabolic monitor (3.15 +/- 1.49 MJ/day) we found bias in energy expenditure of -0.06 (equal to -2%, NS) with limits of agreement of -0.5 to 0.4 MJ/day (equal to -16% to +13%). Intra-measurement variability (coefficient of variation) was within 10% for both methods. Both the metabolic monitor and Douglas bag showed significant bias compared to Schofield equations (3.39 +/-1.64 MJ/day) of -7% (P < 0.01) and -5% (P < 0.05), respectively, with wide limits of agreement: metabolic monitor vs. Schofield: -37% to +22%, Douglas bag vs. Schofield: -37% to +26%. CONCLUSIONS: The Douglas bag method compared favourably to the metabolic monitor where Schofield equations failed to predict individual energy expenditure. Considering its low cost, this renders the short and simple Douglas bag method a robust measure and a routinely applicable instrument for tailored nutritional assessment in critically ill children. PMID- 15297104 TI - Metabolism of defined structured triglyceride particles compared to mixtures of medium and long chain triglycerides intravenously infused in dogs. AB - The present study aimed to determine whether including medium-chain fatty acids (MCFA) in specifically designed structured triglycerides (STG) with a MCFA in sn 1 and sn-3 positions and a long-chain (LC) FA in sn-2 position (MLM) would lead to different effects on plasma lipids and FA distribution into plasma and tissue lipids by comparison to a mixture of separate MCT and LCT molecules (MMM/LLL). The fatty acid (FA) composition was comparable in both lipid emulsions. Lipids were infused over 9h daily, in 2 groups of dogs (n = 6 each), for 28 days as a major component (55% of the non-protein energy intake) of total parenteral nutrition (TPN). Blood samples were obtained on specific days, before starting and just before stopping TPN. The concentration of plasma lipids was measured before starting and before stopping TPN on days 1, 2, 3, 4, 5, 8, 10, 12, 16 and 28. Biopsies were obtained from liver, muscle and adipose tissue 15 days before starting, and again on the day following cessation of TPN. In addition, the spleen was removed after the TPN period. FA composition in plasma and tissue lipids was analysed by gas liquid chromatography in different lipid components of plasma and tissues. No differences in either safety or tolerance parameters were detected between both lipid preparations. A lower rise of plasma TG (P < 0.05) was observed during MLM infusion, indicating a faster elimination rate of MLM vs MMM/LLL emulsion. In spite of the differences of TG molecules which would be assumed to affect the site of FA delivery and metabolic fate, FA distribution in phospholipids (PL) of hepatic and extrahepatic tissues did not substantially differ between both emulsions. PMID- 15297105 TI - Sunflower oil does not protect against LDL oxidation as virgin olive oil does in patients with peripheral vascular disease. AB - BACKGROUND & AIMS: The aim of this study was to compare the in vivo effects of a diet rich in virgin olive oil or sunflower oil on the lipid profile and on LDL susceptibility to oxidative modification in free-living Spanish male patients with peripheral vascular disease. METHODS: A total of 20 Spanish male subjects diagnosed with peripheral vascular disease were randomly divided into two groups (n = 10) receiving different supplements, virgin olive oil and sunflower oil for 4 months. RESULTS: The adaptation of patients to the experimental supplements was demonstrated since plasma and LDL fatty acids composition reflected dietary fatty acids. No differences in triglycerides, total cholesterol, LDL-cholesterol or HDL cholesterol concentrations were found between the groups of patients. A significantly higher LDL susceptibility to oxidation was observed after sunflower oil intake in comparison with virgin olive oil, in spite of an increase in LDL alpha-tocopherol concentration in sunflower oil group. CONCLUSIONS: The results of the present study provide further evidence that sunflower-oil-enriched diets does not protect LDL against oxidation as virgin olive oil does in patients with peripheral vascular disease. PMID- 15297106 TI - Oral supplements differing in fat and carbohydrate content: effect on the appetite and food intake of undernourished elderly patients. AB - BACKGROUND & AIMS: Since fat, relative to other macronutrients, has low satiety and high energy density, it may have therapeutic application for supplementing energy intake. This study compared the effect of isoenergetic (1050 kJ) high fat or high carbohydrate oral supplements, given at breakfast, on the short-term appetite and energy intake in undernourished elderly subjects. METHODS: Sixteen hospitalised, undernourished (body mass index: 20 +/- 3 kg/m2), elderly (77 +/- 8 yr) people were randomly allocated to a control or 1 of 2 supplement groups [fat: carbohydrate: protein (% energy) was 70:25:5 or 25:70:5]. In each group, energy intake (24-h food consumption) and appetite (visual analogue scales) were assessed over 3 consecutive days. RESULTS: Mean energy intake significantly (P = 0.0035) increased following supplementation: high fat 6973 kJ/d, high carbohydrate 6906 kJ/d vs. control 6079 kJ/d but mean voluntary 24-h energy intake remained unaffected. Compared to controls, supplemented subjects experienced reduced hunger (P = 0.07) between breakfast and lunch, but showed no difference over the whole day (P = 0.55). CONCLUSIONS: Under these study conditions a 1050 kJ oral supplement, irrespective of macronutrient composition, does not cause voluntary short-term energy intake compensation in undernourished elderly people. PMID- 15297107 TI - A prospective cohort study of feeding needle catheter jejunostomy in an upper gastrointestinal surgical unit. AB - BACKGROUND & AIM: Feeding jejunostomy is recommended to facilitate early enteral nutrition after major upper gastrointestinal surgery. We aimed to determine the benefits and risks associated with routine practice of feeding needle catheter jejunostomy (NCJ) in high-risk upper gastrointestinal surgery. METHOD: This is a prospective consecutive cohort study of 84 patients underwent feeding NCJ over a 3 years period in an Upper Gastrointestinal Surgical Unit. RESULTS: Feeding NCJ was placed after two-stage oesophago-gastrectomy in 24 patients (28.6%), after gastrectomy in 29 patients (34.5%), after liver resections in 7 patients (8.3%), pancreatic resection in 6 patients (7.1%), bile duct reconstruction in 8 patients (9.5%) and other operations in 10 patients (12%). The mean (SE) estimated nutritional requirement per 24 h was 1791 (31)kcal. Eighty-two patients (98%) started enteral feed on day 1 after surgery. Fifty-seven patients (68%) achieved the target nutritional requirements in 3 days. Four patients were discharged home on jejunal feed whilst only two patients required parenteral nutrition support. The rest tolerated full oral diet. There was no procedure related mortality. The morbidity related to feeding tube and feeding were 12.9% and 20%, respectively. CONCLUSIONS: Routine practice of feeding NCJ is safe. Their benefits outweigh the risks in a specialist centre. PMID- 15297108 TI - Safe and efficacious prolonged use of an olive oil-based lipid emulsion (ClinOleic) in chronic intestinal failure. AB - BACKGROUND & AIMS: Injectable lipid emulsion is an important component of parenteral nutrition. ClinOleic is a lipid emulsion composed of olive oil (80%) and soybean oil (20%). This study evaluated the efficacy and safety of ClinOleic in adults already receiving parenteral nutrition, comparing it to their usual lipid (soybean-oil-based). METHODS: Thirteen adults dependent on home parenteral nutrition were recruited from a single hospital. ClinOleic was administered for 6 months. Two-monthly assessments were made. In addition, clinical and adverse events were recorded for 6-month periods before, during and after the study. RESULTS: Total numbers of important complications for the 6 months before, during and after the study were 13, 9 and 9, respectively. There were, respectively, 5, 3 and 2 line infections, and 2, 0 and 5 thrombotic episodes in the 3 periods. The numbers of unplanned admissions were, respectively, 8, 5 and 7, with in-patient days accounting for 3.4%, 1.5%, and 2.6% of feeding days, respectively. One patient died (pneumonia). One new case of cholecystolithiasis appeared. CONCLUSION: ClinOleic may be used as a safe alternative to standard soybean-oil based lipid emulsions. PMID- 15297109 TI - Impact of enteral versus parenteral nutrition on the incidence of fungal infections: a retrospective study in ICU patients on mechanical ventilation with selective digestive decontamination. AB - BACKGROUND: Infections are an important cause of morbidity and mortality in patients in intensive care units (ICUs). Fungal infections have increased substantially over recent years and fungi have become one of the important pathogens in ICU patients. This study tests the hypothesis that the incidence of fungal infections is lower in critically ill patients under mechanical ventilation receiving enteral rather than total parenteral nutrition. METHODS: By using a prospectively built database, we analyzed retrospectively the charts of 110 critically ill, intubated patients hospitalized in surgical and medical ICUs and receiving selective digestive topical decontamination (SDD), which consisted of administering non-absorbable antibiotics. Patients without contraindications, and expected to be intubated for more than 72 h, received enteral nutrition within 24 h after intubation. Patients with contraindications for enteral nutrition received total parenteral nutrition, which was discontinued when the criteria for enteral nutrition were met. The incidence of fungal infections in both subgroups of patients was compared. RESULTS: Seventy-nine patients received enteral nutrition and 31 total parenteral (10 patients did not meet the inclusion criteria). Both subgroups were similar with regard to their APACHE II score, age, sex distribution and comorbidities at the time of study entry. No difference was observed in the rate of fungal infections between enteral nutrition (5/29) and total parenteral nutrition (7/71) patient groups. CONCLUSION: No significant decrease in the incidence of fungal infections in critically ill patients receiving SDD was observed between those receiving enteral and total parenteral nutrition. PMID- 15297110 TI - Insulinogenic index at 15 min as a marker of stable eating behavior in bulimia nervosa. AB - BACKGROUND & AIMS: The aim of this study was to determine the relationship between insulinogenic index at 15 min (II15 min), body weight maintenance, and the presence of vomiting in patients with bulimia nervosa. METHODS: Forty-eight bulimic inpatients and 14 controls underwent an oral glucose tolerance test on the seventh hospital day. We calculated II15 min and other biological markers, including serum amylase concentrations. During the first week after admission, we monitored the frequency of vomiting and calculated changes in body weight. Patients were divided into 4 subgroups according to the presence of vomiting and weight loss. RESULTS: Two-factor analysis of variance of the II15 min value revealed significant main effects of vomiting and body weight change (P < 0.001 for both). The II15 min values for controls and bulimic patients with weight loss and no vomiting were lower than those of other bulimic groups. The II15 min values were positively correlated with serum amylase concentrations (r = 0.37, P < 0.01), body weight change (r = 0.35, P < 0.05), and frequencies of vomiting (r = 0.49, P < 0.05). CONCLUSIONS: These findings suggest that II15 min values may be a useful marker for assessing the stability of eating behavior in patients with bulimia nervosa. PMID- 15297111 TI - Comparative toxicity of oleic acid and linoleic acid on Jurkat cells. AB - BACKGROUND: Lipid emulsions for parenteral nutrition commercially available are mainly composed of long-chain triacylglycerol containing a high proportion of alpha-6 polyunsaturated fatty acids or alpha-9 monounsaturated fatty acids. The immunological impact of such therapy is particularly important because parenteral and enteral diets are often administered to critical ill patients. The comparative toxicity of oleic acid and linoleic acid on Jurkat cells, a human T lymphocyte cell line, and the type of cell death induced by these fatty acids were determined. METHODS: Cell death was investigated by cytometry: decrease in cell volume, increase of granularity, DNA fragmentation, phosphatidylserine externalization, mitochondrial depolarization, lipid accumulation; by fluorescence microscopy: chromatin condensation and acridine orange/ethidium bromide assay; and by RT-PCR: mRNA expression of apoptotic genes. RESULTS: Evidence is presented herein that oleic acid is much less toxic to Jurkat cells than linoleic acid. Both fatty acids promote apoptosis and necrosis of these cells. The mechanism of cell death induced by these fatty acids seem to involve with mitochondrial depolarization, lipid accumulation and the levels of C-MYC and P53 mRNA expression. CONCLUSION: Therefore, oleic acid may offer an immunological less harmful alternative to linoleic acid for parenteral and enteral diets preparation. PMID- 15297112 TI - Preoperative oral carbohydrate treatment attenuates endogenous glucose release 3 days after surgery. AB - BACKGROUND & AIMS: Postoperative metabolism is characterised by insulin resistance and a negative whole-body nitrogen balance. Preoperative carbohydrate treatment reduces insulin resistance in the first day after surgery. We hypothesised that preoperative oral carbohydrate treatment attenuates insulin resistance and improves whole-body nitrogen balance 3 days after surgery. METHODS: Fourteen patients undergoing total hip replacement were double-blindly randomised to preoperative oral carbohydrate treatment (12.5%, 800 + 400 ml, n = 8) or placebo (n = 6). Glucose kinetics (6,6-D2-glucose), substrate utilisation (indirect calorimetry) and insulin sensitivity (hyperinsulinaemic-euglycaemic clamp) were measured preoperatively and on the third day after surgery. Nitrogen losses were monitored for 3 days after surgery. Values are mean (SEM). Analysis of variance (ANOVA) statistics were used. RESULTS: Endogenous glucose release during insulin infusion increased after surgery in the placebo group. Preoperative carbohydrate treatment, as compared to placebo, significantly attenuated postoperative endogenous glucose release (0.69 (0.07) vs. 1.21 (0.13)mg kg(-1) x min(-1), P < 0.01), while whole-body glucose disposal and nitrogen balance were similar between groups. CONCLUSIONS: While insulin resistance in the first day after surgery has previously been characterised by reduced glucose disposal, enhanced endogenous glucose release was the main component of postoperative insulin resistance on the third postoperative day. Preoperative carbohydrate treatment attenuated endogenous glucose release on the third postoperative day. PMID- 15297113 TI - Insulin stimulated glucose disposal in peripheral tissues studied with microdialysis and stable isotope tracers. AB - BACKGROUND & AIMS: Methods to study glucose kinetics in vivo in specific tissues or tissue beds in humans are often not feasible due to invasiveness or costs of equipment needed. Here we investigate whether the loss (fractional extraction) of 2H7-glucose infused via a microdialysis catheter can be used to study glucose disposal in skeletal muscle and subcutaneous adipose tissue. METHODS AND RESULTS: A perfusion period of 2 h was needed to ensure an isotopic steady state in the microdialysis catheters in skeletal muscle and adipose tissue. In six healthy volunteers the fractional extraction increased during a hyperinsulinemic euglycemic clamp in both skeletal muscle and adipose tissue. Following 48 h of starvation in the same subjects, insulin was not able to increase the fractional extraction of 2H7-glucose from the microdialysis in comparison with a baseline measurement. CONCLUSIONS: In response to insulin infusion, the fractional extraction of 2H7-glucose from a microdialysis catheter increases in skeletal muscle and subcutaneous adipose tissue and this increase is blunted during insulin resistance induced by starvation. These results validate that the fractional extraction of a glucose tracers infused via microdialysis can be used as an index of glucose disposal in peripheral tissues or tissue beds. PMID- 15297114 TI - [Oxygen uptake and maximal oxygen uptake: interests and limits of their measurements]. AB - The paper presents a review of the interests and limits of oxygen uptake measurement in the functional testing of athletes and disabled people. The validity of the oxygen uptake as an estimation of the oxygen consumption and aerobic synthesis of ATP is discussed in the introduction of the review. The author discusses the interests of oxygen uptake measurements for the study of energy cost in addition to maximal oxygen uptake. The limits of the study of oxygen uptake kinetics at the beginning of exercise are discussed. The methodology of oxygen measurement is mainly focused on the characteristics of the different ergometers and the choice of an exercise protocol. The review ends with short statements related to the current knowledge on maximal oxygen uptake, its limiting factors and the effects of age, gender, body mass, active muscle mass and training. PMID- 15297115 TI - [Posture and movement analysis and sports medicine]. AB - The use of posture and movement analysis methods has developed during the past 15 years. These methods are of special interest in the field of sport sciences and have allowed to improve the understanding of physiology of posture and movement in athletes. More recently these methods have been used in the field of sport medicine. In some cases, they have helped to identify abnormalities which cannot be seen on standard clinical examination and to understand the mechanism of lesions occurring during sport activities. For the future these methods should provide useful information for understanding the physiopathology of lesions, for developing prevention of pathologies related to sport and for elaborating and assessing new treatment protocols in the field of sport medicine. PMID- 15297116 TI - [Medicolegal aspects of doping in sports]. AB - OBJECTIVES: To investigate the medico-legal aspects of national and international procedures for monitoring prescription drug use by competing athletes. METHODS: We studied the French law No. 99-223 of March 23, 1999, relating to the protection of the health of athletes? We also studied annual statistics from the Ministry of Sports concerning anti-doping controls, substances detected by the National Doping Control Laboratory and penalties applied since 2000, as well as the World Anti-Doping Code, which came into effect on January 1, 2004, and should be universally applied by 2006. RESULTS: Athletes registered with a federation or unregistered athletes taking part in competitions approved by sporting federations can use prescription drugs but must follow strict rules. Athletes under investigation for drug use must declare all drugs or products recently taken. The use of prescription drugs not on the list of the prohibited substances is allowed, but evidence of the use of such drugs is the responsibility of the prescriber. A medical practitioner in France who considers it essential to prescribe prohibited drugs or drugs under certain restrictions must systematically inform the athlete about the regulations by providing various certificates and forms. For international athletes, a form authorizing therapeutic use must be submitted to the validation committee of the applicable international federation. Disciplinary, ordinal and penal sanctions are also described. CONCLUSION: Prescription drug use by an athlete is never a light matter and always engages the responsibility of the doctor. Anti-doping controls and sanctions encourage physicians to comply scrupulously with the medico-legal rules set forth by the public health code and the world anti-doping code. PMID- 15297117 TI - [Isokinetic thigh muscle strength in sports: a review]. AB - There is growing evidence that isokinetic muscle strength, is one of the most common testing method why muscle strength is thought to be a major factor in athletic success and rehabilitation. A lot of publications during the last 20 years concerned the peak torque, the concentric ratio flexor/extensor, with comparison between males and females, sport specialties, young and old people. Isokinetic is also used for evaluation of knee disorders. The results are very useful after knee ligament surgery, less for femoro-patellar disorders and arthrosis. More recently some authors proposed the functional concept (eccentric flexors/concentric extensors ratio) as a predictive method for preventive muscle injuries or ACL lesions. They demonstrated more discomfort after muscle disorders with isokinetic eccentric testing, and proposed rehabilitation programs for prevention. However apart from a few situations, isokinetic testing does not fully predict functional measurements. It must be used with other techniques of evaluation (clinical methods and imagery). PMID- 15297118 TI - [Eccentric muscular contraction: implications in treatment of athletes]. AB - OBJECTIVES: To review the research on the effects and indications of eccentric muscular contraction in the athlete. METHODS: We searched the Medline database using the key words eccentric and negative work for reports of eccentric muscular contraction published from 1993 to 2004. RESULTS: The physiologic features of eccentric contraction remain unknown but significantly differ from concentric contraction. Eccentric contraction has a specific effect: during sports practice, it permits the muscular and tendinous complex to support strain and strengthens the muscular action of articular stabilization. But eccentric contraction produces injuries: delayed-onset muscle soreness, and muscle rupture and tendinous injuries such as luxation, rupture, and tendinitis. CONCLUSION: Treatment and prevention must have controlled protocols. In therapy, eccentric contraction must be realized at progressive speeds and resistances. Prevention of tendinous injury necessitates quantifying eccentric activity during training. PMID- 15297119 TI - [Delayed post effort muscle soreness]. AB - Muscle intolerance to exercise may result from different processes. Diagnosis involves confirming first the source of pain, then potential pathological myalgia. Delayed-onset muscle soreness (DOMS), commonly referred as tiredness, occurs frequently in sport. DOMS usually develops 12-48 h after intensive and/or unusual eccentric muscle action. Symptoms usually involve the quadriceps muscle group but may also affect the hamstring and triceps surae groups. The muscles are sensitive to palpation, contraction and passive stretch. Acidosis, muscle spasm and microlesions in both connective and muscle tissues may explain the symptoms. However, inflammation appears to be the most common explanation. Interestingly, there is strong evidence that the progression of the exercise-induced muscle injury proceeds no further in the absence of inflammation. Even though unpleasant, DOMS should not be considered as an indicator of muscle damage but, rather, a sign of the regenerative process, which is well known to contribute to the increased muscle mass. DOMS can be associated with decreased proprioception and range of motion, as well as maximal force and activation. DOMS disappears 2 10 days before complete functional recovery. This painless period is ripe for additional joint injuries. Similarly, if some treatments are well known to attenuate DOMS, none has been demonstrated to accelerate either structural or functional recovery. In terms of the role of the inflammatory process, these treatments might even delay overall recovery. PMID- 15297120 TI - [Pain after anterior cruciate ligament reconstruction: detail and treatment]. AB - OBJECTIVE: Several factors can influence the occurrence of pain after anterior cruciate ligament (ACL) ligamentoplasty, namely, factors linked to the selection of graft, the surgical technique and rehabilitation program. The aim of this research was to review the literature for different types of pain experienced after ACL reconstruction to illuminate better prevention and treatment. METHOD: We reviewed reports by searching the PUBMED* research engine using the key words pain, complication, and anterior cruciate ligament. RESULTS: We found an increasing incidence of anterior pain after use of the patellar tendon for ACL reconstruction. Some anterior pain can be mistakenly interpreted as donor site morbidity or patello-femoral problems, but it is in fact linked to graft impingement caused by bad placement of tunnels and/or proliferation of a fibrovascular nodule on the ACL graft ("cyclops syndrome"). Some posterior external pain, sometimes associated, can occur under the same circumstances. Cartilage, ligament or meniscal lesions or saphenous nerve injuries can explain some occurrences of pain. Pain linked to loco-regional factors such as reflex sympathetic dystrophy appear to be rare with rehabilitation and surgery. CONCLUSION: The treatment of pain after ACL reconstruction lies first in the establishment of an etiologic diagnosis. Prevention, by controlling tendon muscular and joint stress during rehabilitation, is essential and requires sound knowledge of biomechanical data. PMID- 15297121 TI - [Rating scores for ACL ligamentoplasty]. AB - INTRODUCTION: Recently, results of surgery for anterior cruciate ligament (ACL) have been systematically assessed. Several scales have been developed, but for most rigorous validation is lacking. METHODOLOGY: We reviewed reports of published scales for ACL surgery and compared their psychometric properties. We searched the MedLine and Cochrane databases with the key words anterior cruciate ligament, surgery, and rating score. A scale was reviewed if its reliability, validity, and responsiveness were reported at least once. RESULTS: We reviewed four scales (Lysholm and Tegner, Cincinnati, IKDC, and Koos). Test-retest reliability was good, except for the IKDC. For all scales, construct validity could not been ascertained. Responsiveness was acceptable and of the same magnitude for the Lysholm and Tegner, and Cincinnati scales. CONCLUSION: None of the scales had sufficient psychometric properties and all seemed too complicated for routine use. Validation of a simple scale is needed. Psychometric properties of the last version of the IKDC (IKDC 2000) and the Cincinnati scales should be studied. PMID- 15297122 TI - [Sprained ankle in children]. AB - OBJECTIVE: To review the literature on sprained ankle in children. We can describe our clinical experience but not statistical analysis because of the few cases. MATERIALS AND METHODS: We searched MedLine with sprain, ankle, children, talo-fibular ligament, ligament injury as key words. RESULTS: We found few and no results from a prospective or randomized trial. We analyzed three surgical publications: Vahvanen (1983, 1984), and Chaumien (1986). Vahvanen's studies concerned 50 ankles treated nonoperatively and 40 treated operatively, and Chaumien described 19 patients. Vahvanen's results suggest that sprained ankle is surprisingly common in children and that primary repair of the ligament will resolve symptoms and lead to a stable ankle joint. Chaumien's description revealed severe lesions defined by radiographic criteria. Sixteen of the 18 patients who underwent surgery healed without any complications. The surgical repair of acute ankle sprain in children has generated much controversy. CONCLUSION: Ankle sprain is a common injury in children and probably underestimated. Clinical evaluation and pain on pressure at the anterior talo fibular ligament are used in diagnosis. Radiographic studies are necessary to eliminate fractures or an avulsion fragment. Ultrasound can be helpful. Most practitioners and orthopedic surgeons prefer to treat ankle sprains in children nonoperatively. PMID- 15297123 TI - [Role of orthoses in ligament injuries of the knee]. AB - OBJECTIVE: To review the literature for the benefits of the three different knee braces (prophylactic, rehabilitation, functional) that can be used to treat ligament injuries of the knee. What is their influence on stability? Do they have adverse effects? Is their use justified? METHODS: We searched the Medline and Embase databases with use of the keywords knee, orthoses, brace, proprioception, stability, rehabilitation, physical therapy, and anterior cruciate ligament for reports published between 1980 and 2003 and selected 93 articles, expert reports or literature reviews. RESULTS: Anterior tibial displacement can be controlled with mechanical strains (150 N), which are lower than physiological restraints? (400 N). The control of joint position is improved by increasing proprioception. Soft tissue stiffness influences the control of anterior tibial displacement. The beneficial effects observed are mainly subjective. Our analysis of the literature showed limitations and variations in study methodologies. CONCLUSION: We did not find any justification for use of either prophylactic or rehabilitation braces. Functional braces can improve stability, as reported by the patient, and may be used in some situations. PMID- 15297124 TI - [Exertional compartment syndrome]. AB - OBJECTIVES: To review the literature on chronic exertional compartment syndrome. METHODS: We searched the Medline database with use of the keys words compartment syndrome, exertional, chronic, pressure, and fasciotomy. RESULTS: Exertional compartment syndrome is characterized by pain on exertion, which recedes at rest, and by excessive increase in compartment intramuscular pressure. Intramuscular pressure measurement is the reference diagnostic tool, but it has not been standardized or evaluated. Pressure observed during the first 5 min after exertion stops is more often used in diagnosis. The first studies of noninvasive investigations (magnetic resonance imaging, thallium single-photon emission tomographic imaging, near infrared spectroscopy) revealed their inadequate diagnostic value. The pathophysiological features of exertional compartment syndrome remain unclear: increased muscle bulk, fascia thickness and stiffness, stimulation of fascial sensory stretch-receptors, poor venous return, micromuscular injuries, and small clinical myopathic abnormalities. Treatment includes decreased sport activity or fasciotomy with partial fasciectomy. Several authors have used endoscopically assisted fasciotomy, which retrospective studies have shown to be successful. Long-term outcome studies could investigate the persistence of exertional minor pain and recurrence of the compartment syndrome with this treatment. CONCLUSION: Further studies are required to understand the physiopathology, standardize the intramuscular pressure test and evaluate the pressure threshold values, evaluate noninvasive investigations and specify the long-term outcome of fasciotomy. PMID- 15297125 TI - [Nonsurgical treatment of tennis elbow]. AB - OBJECTIVE: To review the literature on nonsurgical treatment of tennis elbow. METHODS: We searched Medline for all randomized controlled trials (RCTs), controlled clinical trials (CCTs) and literature reviews published from 1966 to December 2003 on nonsurgical treatment of tennis elbow. We used the keys words controlled clinical trial, tennis elbow on lateral epicondylitis, and treatment. We found 46 reports of RCTs and CCTs on 14 nonsurgical treatments and 11 literature reviews. RESULTS: Corticosteroid injection is the best treatment option for the short term. However, beneficial effects persisted only for a short time, and the long-term outcome could be poor. For the long term, physiotherapy (pulsed ultrasound, deep friction massage and exercise programme) was the best option but was not significantly different from the "wait-and-see" approach. Some support is offered for the use of topical nonsteroid anti-inflammatory drugs, at least for the short term. There is insufficient evidence to support or refute the use of acupuncture, extracorporeal shock wave therapy, manipulation, orthoses, low-energy laser, glycosaminoglycan polysulfate injection, botulinum toxin injection, or topical nitric oxide application. CONCLUSION: Further trials, with use of appropriate methods and adequate sample sizes, are needed before conclusions can be drawn about the effects of many of the treatments for tennis elbow and their ability to change the condition's natural course. PMID- 15297127 TI - [Stress fracture in athletes and prevalence of extensive rehabilitation]. AB - OBJECTIVE: The aim of this work was to review the literature for the place of extensive rehabilitation for athletes with stress fractures. METHOD: We searched the Medline and Embase databases using the keywords stress fracture, sports, rehabilitation, management and treatment. Only French and English articles were included, and articles about bone physiology, animal models, and spine and chest localisations were excluded. From 468 scientific articles, 62 were chosen because they corresponded to literary reviews or to therapeutic evaluations. RESULTS: Treatment of stress fracture is justified according to risk factors, stress fracture complications, the precocity of diagnosis, the therapeutic method and when the athlete needs to return to the sport. The most common treatment is discontinuing the sport, followed by rest. The progressive resumption of sport is rarely described but must take into account mechanical constraints that can be controlled by the use of shoes adapted to a supple ground. Some treatments such as immobilization by pneumatic splint surgery or use of electric fields are controversial. CONCLUSION: Extensive treatment of stress fractures is mainly dedicated to high-level athletes who need to regain previous physical capacities as soon as possible. PMID- 15297126 TI - [Pubalgia: from diagnosis to return to the sports field]. AB - OBJECTIVES: To review reports of the diagnosis and treatment of groin pain (pubalgia) on the basis of anatomical considerations, epidemiology and pathogenicity. METHODS: We searched the Medline database using the key words groin injury, groin pain, and symphisis syndrome for information on groin pain. RESULTS: Despite the limitations of this study, athletic pubalgia appears to be a real diagnosis, with a long duration of symptoms and a therapeutic challenge. The pathophysiologic processes of this lower abdominal pain resulting from over use is unclear, but muscular imbalance might be involved in the pathogenicity. There is no consensus on the diagnostic criteria and the role of imaging (magnetic resonance imaging). Physicians should eliminate the diagnosis of hip and groin injuries in athletes. Specific rehabilitation should include eliminating the pain triggering factors, increasing the limited flexibility, and strengthening the abdominal muscles and adductor muscles. The multidisciplinary team's goal is to restore function and prevent recurrence. Successful surgical repair is predictable in well-selected patients. CONCLUSION: Further studies are required for better assessment of incidence, the natural course of groin pain, and optimal clinical evaluation in screening patients. Overall, a large prospective randomized study of athletes with groin pain would help determine optimal treatment. PMID- 15297128 TI - [Evolution in prostheses for sprinters with lower-limb amputation]. AB - For about 15 years, technical advances in prosthetic treatment have been the main factor in the increased performance of athletes with lower-limb amputation. For trans-tibial amputation, the prosthesis for sprinting is composed of a gel liner and a socket joined by a locking or virtual vacuum liner. Because of these dynamic properties, the carbon prosthetic foot equipped with tacks ensures outstanding performance. For trans-femoral amputation, a hydraulic swing and a stance control unit are added to the same prosthesis. In comparison with the able bodied runner, athletes with amputation have smaller loading times in the prosthetic limb and larger ones in the sound limb. The length of the energy storing prosthetic foot is determined by the "up-on-the-toes" running gait. The sprinting gait with trans-tibial amputation is almost symmetrical. The hip extensor effort is the main compensation of propulsion reduction with lower-limb amputation. With trans-femoral amputation, the lack of knee increases the asymmetry. The total prosthetic knee extension (early in late-swing phase and lasting during total stance phase) compensates with extension of both hips, especially the opposite one. The amputation and sound limb load transfer with lumbar hyperlordosis concern the pelvis, trunk and shoulders. Because of athletes with amputation, research in prosthetic treatment has progressed. The development of orthotics and prostheses for such athletes has benefited non-athletes with amputation. PMID- 15297129 TI - [Hip and knee replacement and sport]. AB - INTRODUCTION: Knee or hip arthroplasty in the sportsman raise many problems including the possibility or to come back to sports activities, and the implants longevity. Unicompartimental prosthesis of the knee is a relatively preserving surgery whose results in the long run in particular for the sporting subject can be for some authors sometimes disappointing. Total knee arthroplasty remains more invasive but with functional results always compatible with some sports activities. The functional result depends much on the postoperative, plurifactorial flexion. Concerning the hip, the sportsman, more active and younger than the average population suffering from osteoarthritis suffer statically more of osteoarthritis. The contribution of the total hip prostheses without cement, anatomical even custom, and of the new couples of friction seems to allow a better longevity. In addition, the contribution of the mini-invasive techniques and the computer-assisted surgery seem techniques with a promising future. METHODS: We present our experience with the unicompartimental arthroplasty, the postoperative flexion in the total knee prostheses and in the hip arthroplasty for the younger patient and the relations between these interventions and sports. RESULTS: Many authors studied the possibilities of sporting recovery (standard of sport and level of practice). The majority of the activities can be taken again, by excluding the sports of team, of ball and the jogging. The contribution of the mini-invasive techniques seems to allow a faster recovery and the uncemented hip prostheses give good long-term results in the young and active patient. PMID- 15297130 TI - [Measurement of shoulder disability in the athlete: a systematic review]. AB - OBJECTIVES: To identify all available shoulder disability questionnaires and to examine those that could be used for athlete. METHODS: We systematically reviewed the literature in Medline using the keywords shoulder, function, scale, index, score, questionnaire, disability, quality of life, assessment, and evaluation. We searched for scales used for athletes with the keywords scale name AND (sport OR athlete). Data were completed by using the "Guide des Outils de Mesure et d'Evaluation en Medecine Physique et de Readaptation" textbook. Analysis took into account the clinimetric quality of the instruments and the number of items specifically related to sports. RESULTS: A total of 37 instruments have been developed to measure disease-, shoulder-specific or upper extremity specific outcome. Older instruments were developed before the advent of modern measurement methods. They usually combined objective and subjective measures. Recent instruments were designed with use of more advanced methods. Most are self administered questionnaires. Fourteen scales included items assessing sport activity. Four of these scales have been used to assess shoulder disability in athlete. Six scales have been used to assess such disability but do not have specific items related to sports. CONCLUSION: There is no gold standard for assessing shoulder outcome in the general population and no validated outcome instruments specifically for athletes. We suggest the use of ASES, WOSI and WORC scales for evaluating shoulder function in the recreational athletes. The DASH scale should be evaluated in this population. The principal criterion in evaluating shoulder function in the high level athlete is a return to the same level of sport performance. Further studies are required to identify measurement tools for shoulder disability that have a high predictive value for return to sport. PMID- 15297131 TI - Recurrences after conformal parotid-sparing radiotherapy for head and neck cancer. AB - BACKGROUND AND PURPOSE: Evaluation of loco-regional failure patterns and survival after parotid-sparing three-dimensional conformal and intensity modulated radiotherapy (IMRT) for head and neck cancer. PATIENTS AND METHODS: From June 1999 to July 2002, seventy-two patients with lateralised head and neck tumours, excluding nasopharyngeal tumours and patients with bilateral or contralateral neck disease, were irradiated with a parotid-sparing technique. Three-dimensional conformal planning was used in 68 patients, 4 patients were treated with dynamic IMRT. Bilateral neck node irradiation was performed in all patients, the junctional (or high level II) nodes, contralateral to the tumour, however, were excluded from the clinical target volume to spare the adjacent parotid from irradiation. In 20 patients with persistent or recurrent loco-regional disease, the localisation and volume of the treatment failure, as determined by computed tomography (CT), was copied on the pre-treatment CT-study used for treatment planning. Minimum, mean and maximum doses administered to the region of the failure were calculated and dose--volume histograms were computed of each failure. The failures were divided in three groups depending on the percentage of their volume receiving 95% of the prescribed dose. Recurrences were defined to be in-field (IF) if >95% of their volume received 95% of the prescribed dose and out field (OF) if <20% of their volume received 95% of the prescribed dose. When 20 95% of the volume of the recurrence received 95% of the prescribed dose, this recurrence was defined as extending outside the field (EOF). RESULTS: With a median follow-up time of 19 months, the 2-year loco-regional control rate was 69% with primary radiotherapy and 63.5% with surgery followed by irradiation (P = 0.77). The 2-year overall survival rate for the entire patient population was 67.4%. At the time of analysis, 20 of the 72 patients had developed a loco regional failure; 2 patients (2/20) presented with a loco-regional relapse combined with distant metastasis. Fifteen of the 20 loco-regional failures (15/20) occurred within the high dose region (IF). Five patients (5/20) developed a failure of which the bulky tumour mass was located within the high dose region but extending outside the treatment volume (EOF). No relapses were seen out-field (OF) and no patients relapsed in the spared junctional area contralateral to the tumour. CONCLUSIONS: The selection of patients treated with parotid-sparing radiotherapy, by omitting irradiation to the junctional nodes contralateral to the tumour, proved to be safe in our hands, since no recurrences developed in the spared area. As this parotid-sparing technique reduces significantly the dose to the contralateral parotid and is easy to perform, it should be considered for all selected patients. PMID- 15297132 TI - Potential outcomes of modalities and techniques in radiotherapy for patients with hypopharyngeal carcinoma. AB - BACKGROUND AND PURPOSE: To determine potential improvements in treatment outcome for patients with hypopharyngeal carcinoma, T4N0M0, using proton and intensity modulated photon radiotherapy (IMRT) compared to a standard 3D conformal radiotherapy treatment (3D-CRT) in terms of local tumour control probability, TCP, and normal tissue complication probability (NTCP) for the spinal cord and the parotid glands using. PATIENTS AND METHODS: Using the three-dimensional treatment-planning system, Helax-TMS, 5 patients were planned with protons, IMRT, and 3D-CRT plans. The prescribed dose used was 30 fractions x 2.39 Gy for the protons and IMRT and 35 fractions x 2.00 Gy for 3D-CRT. The treatment plans were evaluated using dose volume data and dose response models were used to calculate TCP and NTCP. The target volumes were delineated to spare the parotid glands. A dose escalation was made for protons and IMRT using NTCP constraints to the spinal cord. RESULTS: On average, protons and IMRT increase TCP by 17% compared to 3D-CRT. For the spinal cord NTCP values are zero for all methods and patients. Average NTCP values for the parotid glands were >90% for 3D-CRT and significantly lower for protons and IMRT varying from 43-65%. The average parotid gland dose was 33 Gy for the protons, 38 Gy for IMRT and 48 Gy for 3D-CRT. CONCLUSIONS: Protons and IMRT gave a significant TCP increase compared to 3D-CRT while no significant difference between protons and IMRT was found. Protons generally show lower non-target tissue doses, which indicates a possibility for further dose escalation. Large individual dose differences between protons and IMRT for parotid glands indicate that some patients may benefit more from protons and others from IMRT. PMID- 15297133 TI - Long-term bladder, colorectal, and sexual functions after radical radiotherapy for urinary bladder cancer. AB - PURPOSE: To describe bladder, colorectal, and sexual dysfunctions among long time survivors after radical radiotherapy for urinary bladder cancer, and compare the results with a healthy control group. MATERIALS AND METHODS: We identified 261 patients who had received radical radiotherapy for bladder cancer in the period 1994-2001. Patients were treated with a CT-based three-field technique with 60 Gy in 2 Gy fractions, 5 fractions/week. Sixty-two patients were alive and candidates for the study. For comparison, 185 controls were selected from the Danish National Register. Information was collected in an interview based on the LENT SOMA tables and questions concerning changes in daily life following radiotherapy. RESULTS: Fifty-three patients (85%), median age 77 years (range 51 84) entered the study. Median follow-up time was 29 months (range 18-103 months). There were 63 controls (34%). Fourteen percent of the patients reported that radiotherapy had moderate to severe impact on their present bladder function. Compared with the control group, significantly more patients had dysuria, and urethral stenosis, and were using bladder catheter. Twenty-nine percent of the patients reported moderate to severe impact on their present bowel function. Significantly more patients had diarrhoea, fecal urgency and fecal incontinence, and were using antidiarrhoea medication and sanitary pads. Twenty-five percent of the patients reported moderate to severe impact on their present sexual function. Impotence and lack of sexual desire were significantly higher among the male patients. CONCLUSIONS: Following radical radiotherapy, most patients had a well functioning bladder, whereas 14% reported moderate to severe bladder dysfunctions. Due to the presence of bowel in the treatment field, radiotherapy is associated with considerable long-term intestinal side effects. Moreover, radiotherapy may result in sexual dysfunctions. PMID- 15297134 TI - The immunohistochemical assessment of hypoxia, vascularity and proliferation in bladder carcinoma. AB - BACKGROUND AND PURPOSE: Hypoxia and proliferation are important determinants of radiation responsiveness; prospective measures of these before radiotherapy may enable individualisation of treatment schedules. Immunohistochemical techniques offer a potential means of achieving this in routine biopsy material. MATERIAL AND METHODS: Cellular hypoxia as measured by pimonidazole fixation and immunohistochemistry has been evaluated in a series of human bladder cancers with dual staining of sections for pimonidazole and either the vascular markers, CD31/34, or proliferation markers, Ki-67 or cyclin A. Twenty one tumour specimens were examined suitable for the double staining technique. RESULTS: The median hypoxic fraction was 9% (range 0-38). Seven tumours did not stain for pimonidazole and 11 exhibited necrosis. The mean vascular density ranged from 16.7 to 160.6 vessels per mm2. The median hot spot count was 30 (range 16-43). There was a statistically significant increase in vessel density in hypoxic compared to oxic regions measured by both vessel density (P = 0.02) and hot spot count (P = 0.004). Proliferation indices decreased from oxic to hypoxic areas close to blood vessels. CONCLUSIONS: We have demonstrated that bladder cancer exhibits a range of hypoxia, proliferation and vascular density which may be used to form the basis for patient selection for hypoxia modification, accelerated radiotherapy and vascular targeting agents. PMID- 15297135 TI - Early improvements in vision after fractionated stereotactic radiotherapy for primary optic nerve sheath meningioma. AB - BACKGROUND AND PURPOSE: To report on the efficacy and follow-up of 23 patients with primary optic nerve sheath meningioma (ONSM) with fractionated stereotactic conformal radiotherapy (SCRT). PATIENTS AND METHODS: Between 1996 and 2003, 23 patients ( = 23 eyes) with ONSM were treated. Indications for primary stereotactic radiotherapy were tumour progression documented by imaging or symptoms (loss of vision, pain). All patients received SCRT with a median dose of 50.4Gy in 6 weeks. RESULTS: After a median follow-up of 20 months (1-68 months) a 95% (21 of 22) visual control was seen: vision improved in 16 patients and remained stable in 5. For 13/16 patients improvement was documented already within 1-3 months after SCRT. Vision became worse in one patient. An improvement of pain was observed after radiotherapy in 6 patients as well as of proptosis in 1 patient. For 1 patient pain was persistent after SCRT. In one patient 4 years after SCRT a radiation retinitis and vitreous haemorrhage was seen. CONCLUSIONS: Fractionated stereotactic radiotherapy improves vision, often shortly after treatment, and is thus a viable treatment option for this tumour entity. PMID- 15297136 TI - Time without symptoms and toxicity (TWIST) analysis of adjuvant radiation therapy for endometrial cancer. AB - BACKGROUND AND PURPOSE: Postoperative radiotherapy in endometrial cancer reduces the risk of local relapse but is also associated with substantial acute and late reactions. The aim of our study was to evaluate time without tumor symptoms and toxicity (TWIST) in a consecutive series of 317 endometrial cancer patients administered postoperative irradiation. PATIENTS AND METHODS: Both low-dose rate brachytherapy (BRT) and external beam irradiation (EBRT) were applied in 247 patients (78%), only BRT--in 49 (15%) and only EBRT--in 21 patients (7%). Median follow-up was 7.3 years (range, 4-21 years). TWIST analysis based on actuarial freedom from recurrent disease and from late radiotherapy effects was performed with the use of Kaplan-Meier method. The impact of patient- and treatment-related factors on TWIST was assessed with uni- and multivariate tests. RESULTS: Five year overall survival was 78%, and five-year disease free survival--75%. Recurrence occurred in 70 patients (22%), of whom in 11 (3.5%)--exclusively in the pelvis. Acute and late reactions of any grade occurred in 268 (85%) and 158 patients (51%), respectively. Late bowel effects of any grade were observed in 41% of patients. Severe late effects occurred in 35 patients (11%). Actuarial probability of two- and five-year survival free of disease and severe (grades 3 or 4) late effects (TWIST) was 84% and 71%, respectively (median TWIST, 16.2 years). When all-grade late effects were considered, two- and five-year TWIST probability was 50 and 30%, respectively, and median TWIST was only 2.0 years. When both acute and late reactions were taken into account, median TWIST was 22 months. In unifactorial test, higher age ( P = 0.013) FIGO stage ( P < 0.001) total radiotherapy dose ( P < 0.001) normalized total dose based on linear quadratic model ( P = 0.001) EBRT fraction dose ( P < 0.001) and use of cesium BRT ( P = 0.042) were correlated with shorter TWIST. In multifactorial analysis, higher age ( P = 0.001) FIGO stage ( P = 0.001) and total radiotherapy dose ( P < 0.001) were independent factors correlated with shorter TWIST. CONCLUSIONS: Endometrial cancer patients treated with postoperative irradiation have a long time interval without relapse and severe late toxicity. However, when any late normal tissue injury is considered, the median time without relapse and late toxicity is significantly shorter. The impact of mild late radiotherapy complications on the quality of life should be further investigated. TWIST calculation should be attempted in future prospective studies evaluating the role of postoperative radiotherapy. PMID- 15297137 TI - Psychosocial distress and need for psychotherapeutic treatment in cancer patients undergoing radiotherapy. AB - BACKGROUND AND PURPOSE: Empirical investigations examining the need for psychotherapeutic treatment of inpatients in radio-oncology departments are rare. We, therefore, assessed the degree of psychosocial distress of radio-oncology inpatients and their need for psychotherapeutic interventions. PATIENTS AND METHODS: Eighty-seven patients from two radio-oncology wards underwent psychodiagnostic interviews and completed self-rating instruments to determine mental disorders, psychosocial distress, coping strategies and quality of life. Need for psychotherapeutic interventions and treatment motivation were estimated by patients and by a professional. RESULTS: Using ICD-10 criteria, mental and behavioral disorders were diagnosed in 51% of the patients, most of them adjustment disorders. The professional regarded 32.2% of patients as needing psychotherapeutic treatment, compared with 43% of the patients who were motivated to accept at least one of the psychotherapeutic treatments offered. There was a marked discrepancy between the points of view of the patient and the professional, in that, while the professional regarded anxiety as a highly significant predictor of the need for treatment, psychosocial distress played no role in the patients' estimation. CONCLUSIONS: In determining the indication for psychotherapeutic treatment, both objectified psychosocial distress and a patients' subjective treatment expectations should be considered. There is a need to develop screening instruments that integrate both aspects. PMID- 15297138 TI - Implementing the UK Medical Research Council (MRC) RT01 trial (ISRCTN 47772397): methods and practicalities of a randomised controlled trial of conformal radiotherapy in men with localised prostate cancer. AB - BACKGROUND AND PURPOSE: Radiotherapy is the most frequently used treatment for men with localised prostate cancer. Conformal radiotherapy (CFRT) is a relatively new development. MRC RT01 was set-up to explore optimum CFRT dose. PATIENTS AND METHODS: RT01 was an international multi-centre randomised controlled trial for men with T1b-T3a, N0, M0 prostate cancer that evolved from a single-centre pilot trial of similar design. All men received at least 3 months of pre-radiotherapy hormone treatment, before randomisation to standard (64 Gy) or high dose (74 Gy) radical CFRT. Accrual was completed in December 2001 with 843 men randomised from 25 centres in less than 4 years. RT01 has been a catalyst for implementing CFRT across UK. In addition to the Trial Management Group, independent Data Monitoring and Ethics Committee and independent Trial Steering Committee, a Quality of Life and Health Economics (QL/HE) group, a radiotherapy Quality Assurance (QA) Group and a Radiography Trial Implementation Group were set up. The QL/HE group ensured implementation, compliance, analysis and interpretation of the QL and HE data in the trial. The inauguration of QA and Radiography groups facilitated inter-centre collaboration. The QA Group ensured procedures were in place before and during trial participation, and monitored quality and consistency with systems including a physics questionnaire, a clinical examples exercise, a standard operating procedure document, designing and building a phantom, and convening a complications modelling subgroup. The Radiography group agreed and implemented technique improvements. RESULTS: More centres participated than initially predicted, enabling recruitment better than scheduled. The trial expedited the implementation of CFRT in many UK radiotherapy centres. Additionally, the QA and Radiography groups helped ensure smooth initiation and established consistency in planning, dosimetry and delivery of prostate CFRT services at participating UK centres. Considerable data has been collected; a series of papers will be produced, although mature clinical trial results are not anticipated until 2006 2008. PMID- 15297139 TI - Idealized line source configuration for permanent 125I prostate implants. AB - BACKGROUND AND PURPOSE: To validate the use of idealized seed orientations in conjunction with the line source formalism for post-implant dosimetry of permanent 125I prostate implants. PATIENTS AND METHODS: Post-implant, a CT scan and three fluoroscopic images were obtained for 32 patients having undergone permanent implants. From these images, the seed positions and orientations (phi,theta) were determined (1625 individual seeds). Two different dosimetric calculations were done: one using real orientations and one using idealized orientations (seeds along the axis of implantation). Dose volume histograms (DVHs) and key dosimetric parameters were compiled for the prostate, urethra, rectum, bladder and penile bulb, to evaluate the difference between the two approximations. RESULTS: The phi angle distribution (phi = 1.1 degrees; sigma phi = 22.9 degrees) and the theta angle distribution (theta= -4.29 degrees; sigma theta = 27.1 degrees) were found to be similar to the first order except for the pronounced peak of the phi angle distribution. The DVHs comparison and dosimetric parameters study reveal no significant difference between the two approximations. The difference in D90 for the prostate was only 0.02% (sigma = 0.91%) The differences were slightly higher in the case of the organs at risk, as expected from the dosimetric characteristics of the seed model used. CONCLUSIONS: The angular distributions (phi,theta) of individual seeds were determined. The dosimetric evaluation shows that line source formalism can be used in conjunction with an idealized seed configuration presented here to report prostate and organs at risk dose coverage. PMID- 15297140 TI - Effect of VEGF receptor-2 antibody on vascular function and oxygenation in spontaneous and transplanted tumors. AB - BACKGROUND AND PURPOSE: The primary objectives of this study were to address two major questions. (1) Does VEGF receptor-2 antibody (DC101) produce detrimental effects on tumor vascular function and oxygenation that could compromise adjuvant therapies? (2) Is pathophysiological response to such antiangiogenic strategies different in transplanted versus primary spontaneous tumors? MATERIALS AND METHODS: The effects of early and late initiation DC101 treatment were evaluated using spontaneous murine mammary carcinomas and two markedly different transplanted mammary tumors, MCa-35 and MCa-4. Mice were administered DC101 or saline, tumors were frozen, and immunohistochemical staining was quantified using image analysis of multiply-stained frozen sections. Total blood vessels were identified using antibodies to CD31 or panendothelial antigen, perfused vessels via i.v. injection of fluorescent DiOC7, and tumor hypoxia by hypoxia marker (EF5) uptake. RESULTS: Tumor growth was significantly inhibited following DC101 administration in all tumor models. In general, early initiation DC101 treatment reduced perfused vessel counts and increased tumor hypoxia, while late initiation treatment had no significant impact on either. Results indicate that DC101 slows tumor growth through a decrease in vascular function, leading to increased tumor cell apoptosis and necrosis at sites distant from perfused blood vessels, and suggest that DC101 accelerates the rate at which tumor cells outgrow their functional vascular supply. CONCLUSIONS: Although highly variable among individual spontaneous tumors, the overall effects of DC101 on tumor hypoxia were quite similar between spontaneous and transplanted tumors. Since reductions in tumor oxygenation due to antiangiogenic treatment were transient, initial pathophysiological deficiencies that could compromise conventional therapies over the short-term may be of less relevance when administered over more extended treatment schedules. PMID- 15297141 TI - Increased expression of pro-inflammatory cytokines as a cause of lung toxicity after combined treatment with gemcitabine and thoracic irradiation. AB - BACKGROUND AND PURPOSE: Preclinical evidence suggesting gemcitabine potentiates the anti-tumor effects of irradiation has resulted in clinical trials to evaluate the treatment efficacy of gemcitabine and concurrent thoracic irradiation in non small-cell lung cancer (NSCLC). Although these studies demonstrated favorable tumor response, this combined treatment modality was accompanied by severe treatment-related toxicities predominantly of the lung. In an attempt to elucidate the determinants of lung toxicity for gemcitabine, we analyzed the expression of the pro-inflammatory cytokines TNF-alpha, IL-1alpha and IL-6 in the lung tissue of mice treated with gemcitabine and concurrent thoracic irradiation. MATERIALS AND METHODS: Four study groups were defined: C57BL/6J mice that received neither irradiation nor gemcitabine (NT-group), those that received gemcitabine (120 mg/kg intraperitoneal, i.p.) but no irradiation (GEM-group), those that underwent thoracic irradiation (12 Gy) without gemcitabine (XRT group), and those that received both gemcitabine (120 mg/kg i.p., 2 h before irradiation) and thoracic irradiation (GEM/XRT-group). The mice were sacrificed at 1 h, 1 and 3 days, 1, 2 and 4 weeks post-treatment (p.t.). The mRNA expression of TNF-alpha, IL-1alpha and IL-6 in the lung tissue was quantified by competitive RT-PCR. The cellular origin of the cytokine expression was identified by immunohistochemistry. The cytokine expression was correlated with histopathological alterations. RESULTS: The TNF-alpha, IL-1alpha and IL-6 expression in the lung tissue of the GEM/XRT mice was clearly higher at all assessment time points compared to the NT mice (statistically significant at 1 h, 1 and 3 days, 1, 2 and 4 weeks p.t.), XRT mice (statistically significant at 1 week p.t.) or GEM mice (statistically significant at 1 h, 1 and 2 weeks p.t.). Maximal treatment-induced cytokine expression in the lung tissue of the GEM/XRT mice occurred already at 1 week p.t. (TNF-alpha: 30.9 +/- 5.3/IL-1alpha: 28.3 +/- 5.0/IL-6: 4.9 +/- 0.1 times basal level), and coincides with pathohistologically discernable interstitial pneumonitis. The elevated levels of TNF-alpha and IL 1alpha have been found to correlate with immunohistochemical staining of the bronchiolar epithelium and predominantly of inflammatory cells. CONCLUSIONS: Our data provide evidence that the increased expression of pro-inflammatory cytokines and the induction of a cytokine-triggered inflammatory response may be a determinant of the observed elevated lung toxicity after concurrent treatment with gemcitabine and thoracic irradiation. PMID- 15297142 TI - Identification of key elements that are responsible for heme-mediated induction of the avian heme oxygenase-1 gene. AB - Heme oxygenase (HO) catalyzes the conversion of heme to biliverdin with the release of iron and carbon monoxide. HO-1 is highly inducible by a large number of physical and chemical factors. In recent work, we had identified a metalloporphyrin-responsive element (MPRE) that localized at -3.7 kb upstream of the transcription start site of the chick HO-1 gene. Here, we identify four additional heme-responsive elements (HeREs), which are "expanded" AP-1 sites, in the 5'-flanking region of the chick HO-1 gene. These sites, located at -4675, 4599, -3660, and -3625 bp from the transcription start site of the gene, were necessary and sufficient for up-regulation of luciferase reporter gene expression in the presence of heme and cobalt protoporphyrin (CoPP), but not several other metalloporphyrins. Site-directed mutagenesis was carried out using pcHO7.1Luc or pcHO7.1-4.6Luc as templates. Single and multiple mutations of HeREs and MPRE significantly abrogated the heme- and CoPP-dependent up-regulation of reporter gene expression in transient or stable transfection experiments. CONCLUSIONS: The chick HO-1 promoter region contains "expanded" AP-1 sites that are important for up-regulation of the gene by heme and CoPP, but not other metalloporphyrins. These key regulatory elements consist of consensus AP-1 binding sites that have been extended by three base pairs. PMID- 15297143 TI - Mutation hotspots in the p53 gene in tumors of different origin: correlation with evolutionary conservation and signs of positive selection. AB - We present a classification analysis of the mutation spectra of the p53 gene and construct maps of hotspots for the germline (Li-Fraumein syndrome), different types of tumors and their derived cell lines. While spectra from solid tumors share common hotspots with the germline spectrum, they also contain unique sets of somatic hotspots that are not observed in the germline. All these hotspots correspond to amino acid replacements in the DNA-binding interface of p53. The mutation spectra of lymphomas and cell lines derived from lymphomas and lung cancers contained few hotspots compared to solid tumors. Thus, the distribution of hotspots in the p53 gene appears to depend on the tumor type and cell growth conditions; this specificity is missed by the bulk hotspot analysis. A negative correlation was detected between the amino acid replacement propensity in tumors and evolutionary variability: the hotspots are located in the positions that are highly conserved in p53 and its paralogs, p63 and p73. In all the mutation spectra, substitutions leading to amino acid replacements strongly dominate over silent substitutions, indicating that functional sites evolving under strong purifying selection are subject to intensive positive selection in p53-dependent tumors. These results are compatible with the gain-of-function concept of the role of p53 in tumorigenesis. PMID- 15297144 TI - Genomic structure and multiple alternative transcripts of the mouse TRAD/RAD51L3/RAD51D gene, a member of the recA/RAD51 gene family. AB - The RecA/RAD51 family plays a central role in DNA recombinational repair. The targeted disruption of mouse RAD51L3/TRAD is lethal during embryogenesis, suggesting that this protein is essential for development. Recently, we reported multiple alternative splice variants of human RAD51L3/TRAD transcripts. In this study, we have identified multiple mouse transcript variants. Complete sequence analysis of the genomic and cDNA clones has confirmed that the exon-intron structures obey the GT/AG splicing rule, and that the multiplicity of the transcripts is due to alternative splicing. In addition, we have determined the transcription initiation site by rapid amplification of cDNA 5'-end (5'-RACE). These results show that the mouse gene structure is very similar to that of humans. PMID- 15297145 TI - Novel alternative splicing of mRNAs encoding poly(A) polymerases in Arabidopsis. AB - The Arabidopsis thaliana genome possesses four genes whose predicted products are similar to eukaryotic poly(A) polymerases from yeasts and animals. These genes are all expressed, as indicated by RT/PCR and Northern blot analysis. The four Arabidopsis PAPs share a conserved N-terminal catalytic core with other eukaryotic enzymes, but differ substantially in their C-termini. Moreover, one of the four Arabidopsis enzymes is significantly shorter than the other three, and is more divergent even within the conserved core of the protein. Nonetheless, the protein encoded by this gene, when produced in and purified from E. coli, possesses nonspecific poly(A) polymerase activity. Genes encoding these Arabidopsis PAPs give rise to a number of alternatively spliced mRNAs. While the specific nature of the alternative splicing varied amongst these three genes, mRNAs from the three "larger" genes could be alternatively spliced in the vicinity of the 5th and 6th introns of each gene. Interestingly, the patterns of alternative splicing vary in different tissues. The ubiquity of alternative splicing in this gene family, as well as the differences in specific mechanisms of alternative processing in the different genes, suggests an important function for alternatively spliced PAP mRNAs in Arabidopsis. PMID- 15297146 TI - Effects of single-stranded DNA binding proteins on primer extension by telomerase. AB - We present a biochemical analysis of the effects of three single-stranded DNA binding proteins on extension of oligonucleotide primers by the Tetrahymena telomerase. One of them, a human protein designated translin, which was shown to specifically bind the G-rich Tetrahymena and human telomeric repeats, slightly stimulated the primer extension reactions at molar ratios of translin/primer of <1:2. At higher molar ratios, it inhibited the reactions by up to 80%. The inhibition was caused by binding of translin to the primers, rather than by a direct interaction of this protein with telomerase. A second protein, the general human single-stranded DNA binding protein Replication Protein A (RPA), similarly affected the primer extension by telomerase, even though its mode of binding to DNA differs from that of translin. A third protein, the E. coli single-stranded DNA binding protein (SSB), whose binding to DNA is highly cooperative, caused more substantial stimulation and inhibition at the lower and the higher molar ratios of SSB/primer, respectively. Both telomere-specific and general single stranded DNA binding proteins are found in living cells in telomeric complexes. Based on our data, we propose that these proteins may exert either stimulatory or inhibitory effects on intracellular telomerases, depending on their local concentrations. PMID- 15297147 TI - Murine glypican-4 gene structure and expression; Sp1 and Sp3 play a major role in glypican-4 expression in 3T3-F442A cells. AB - In this report we describe the genomic organization of the mouse glypican-4 (Gpc4), an analysis of its promoter and its transcriptional regulation in the 3T3 F442A adipocyte cell line. The Gpc4 gene consists of nine exons separated by eight introns. A series of deletion mutants and 4391 bp of the 5'-flanking region were cloned into pGL3-BASIC upstream of the luciferase reporter gene and transfected into 3T3-F442A adipocytes. Analysis of a 4.3-kb DNA fragment at the 5'-flanking region of this gene revealed that the Gpc4 promoter is a TATA-less promoter with a large cluster of GC boxes. Competitive electrophoretic mobility shift and supershift assays identified a cluster of nine functional GC boxes binding Sp1 and Sp3 in this region. Transactivation experiments in insect cells showed that both Sp1 and Sp3 are major activators of the Gpc4 promoter. Gpc4 is expressed in adipocytes where its expression is highest in confluent 3T3-F442A adipoblasts and decreases dramatically as cells differentiate. Sp protein analyses demonstrated a major decrease in Sp3 protein in differentiated adipocytes as compared to undifferentiated adipoblasts. These experiments show that Gpc4 is developmentally regulated in 3T3-F442A adipocytes and suggest that Sp transcription factors play a significant role in the regulated expression of Gpc4. PMID- 15297148 TI - Mutagenesis of the cysteine residues in the transcription factor NtcA from Anabaena PCC 7120 and its effects on DNA binding in vitro. AB - NtcA is a transcription factor found in a wide variety of cyanobacteria. It is a key component in the control of the nitrogen metabolism, and regulates genes involved in ammonia assimilation, heterocyst differentiation and nitrogen fixation. NtcA expression is subject to nitrogen control, but there is also evidence that the binding of NtcA to DNA can be regulated by a redox mechanism involving the two cysteine residues in the NtcA protein from Anabaena PCC 7120. In order to investigate this further, the two cysteine residues in NtcA were mutated into alanine to give four variants of the protein: wild-type NtcA, the point-mutated variants Cys157Ala and Cys164Ala, as well as the double mutant Cys157Ala/Cys164Ala. The binding of a DNA probe containing a palindromic NtcA binding motif was investigated by gel mobility shift analysis under non-reducing and reducing conditions. The experiments show that the DNA binding in vitro is stronger in the presence of the reducing agent DTT than in its absence. However, this effect is not due to breaking of a disulfide bond between the cysteine residues, since the double mutant containing no cysteines was also affected by DTT. A molecular model of a monomer of NtcA, based on the homologous cAMP receptor protein structure, was created in order to locate the positions of the cysteine residues. The NtcA model suggested that the positions of the sulfur atoms are not compatible with formation of a bond between them. PMID- 15297149 TI - Peroxisomal proliferator activated receptor gamma coactivator (PGC-1alpha) stimulates carnitine palmitoyltransferase I (CPT-Ialpha) through the first intron. AB - Peroxisomal proliferator activated receptor gamma coactivator-1 (PGC-1alpha) is a transcriptional coactivator that promotes mitochondrial biogenesis and energy metabolism in brown fat, skeletal muscle and heart. Previous studies demonstrated that PGC-1alpha is present at low levels in the liver but that the hepatic abundance of PGC-1alpha is elevated in diabetic and fasted animals. Elevated PGC 1alpha expression is associated with increased fatty acid oxidation and hepatic glucose production. Carnitine palmitoyltransferase-I (CPT-I) is a rate controlling step in the mitochondrial oxidation of long chain fatty acids. CPT-I transfers the acyl moiety from fatty acyl-CoA to carnitine for the translocation of long chain fatty acids across the mitochondrial membrane. There are two isoforms of CPT-I including a liver isoform CPT-Ialpha and a muscle isoform CPT Ibeta. Here, we characterized the regulation of CPT-Ialpha isoform by PGC-1alpha. PGC-1alpha stimulates CPT-Ialpha primarily through multiple sites in the first intron. We found that PGC-1alpha can induce CPT-Ialpha gene expression in cardiac myocytes and primary hepatocytes. Our results indicate that PGC-1alpha elevates the expression of CPT-Ialpha via a unique mechanism that utilizes elements within the intron. PMID- 15297150 TI - Characterization of a myostatin-like gene from the bay scallop, Argopecten irradians. AB - A complete cDNA was cloned from the bay scallop (Argopecten irradians) that codes for a 382-amino-acid myostatin-like protein (sMSTN). The sMSTN sequence is most similar to mammalian myostatin (MSTN), containing a conserved proteolytic cleavage site (RXXR) and conserved cysteine residues in the C-terminus. Based on quantitative RT-PCR, the sMSTN gene is predominantly expressed in the adductor muscle, with limited expression in other tissues. Using the sMSTN sequence, a Ciona MSTN-like gene was also identified from the Ciona intestinalis genome. These findings indicate that the MSTN gene has been conserved throughout evolution and suggests that MSTN could play a major role in muscle growth and development in invertebrates, as it does in mammals. PMID- 15297151 TI - Promoter variant-dependent expression of the STAT5A gene in bovine liver. AB - Using PCR-heteroduplex and sequencing we identified A/G substitution at position 488 in the promoter region of the bovine STAT5A gene. With reverse transcription PCR (RT-PCR) and other means we showed that the STAT5A expression level in the liver was higher in cattle with AA than GG genotype. Electrophoretic mobility shift assay (EMSA) showed that the A-->G transition at position -488 increased the STAT5A gene promoter binding capacity for liver nuclear proteins [possibly hepatic nuclear factor (HNF)-3]. PMID- 15297152 TI - The thalamic reticular nucleus: structure, function and concept. AB - On the basis of theoretical, anatomical, psychological and physiological considerations, Francis Crick (1984) proposed that, during selective attention, the thalamic reticular nucleus (TRN) controls the internal attentional searchlight that simultaneously highlights all the neural circuits called on by the object of attention. In other words, he submitted that during either perception, or the preparation and execution of any cognitive and/or motor task, the TRN sets all the corresponding thalamocortical (TC) circuits in motion. Over the last two decades, behavioural, electrophysiological, anatomical and neurochemical findings have been accumulating, supporting the complex nature of the TRN and raising questions about the validity of this speculative hypothesis. Indeed, our knowledge of the actual functioning of the TRN is still sprinkled with unresolved questions. Therefore, the time has come to join forces and discuss some recent cellular and network findings concerning this diencephalic GABAergic structure, which plays important roles during various states of consciousness. On the whole, the present critical survey emphasizes the TRN's complexity, and provides arguments combining anatomy, physiology and cognitive psychology. PMID- 15297153 TI - Why study transport of peptides and proteins at the neurovascular interface. AB - The blood-brain barrier (BBB) is an immense neurovascular interface. In neurodegenerative, ischemic, and traumatic disorders of the central nervous system (CNS), the BBB may hinder the delivery of many therapeutic peptides and proteins to the brain and spinal cord. Fortunately, the mistaken dogma that peptides and proteins do not cross the BBB has been corrected during the past two decades by the accumulating evidence that peptides and proteins in the periphery exert potent effects in the CNS. Not only can peptides and proteins serve as carriers for selective therapeutic agents, but they themselves may directly cross the BBB after delivery into the bloodstream. Their passage may be mediated by simple diffusion or specific transport, both of which can be affected by interactions in the blood compartment (outside the BBB) and within the endothelial cells (at the BBB level). Although the majority of current delivery strategies focuses on modification of the molecule to be delivered, understanding the mechanisms of transport will eventually facilitate regulation of the BBB directly. We review the different aspects of interactions and discuss recent advances in the cell biology of peptide/protein transport across the BBB. Better understanding of the nature and regulation of the transport systems at the BBB will provide a new direction to enhance the interactions of peripheral peptides and proteins with the CNS. PMID- 15297154 TI - Genetic contributions to Parkinson's disease. AB - Sporadic Parkinson's disease (PD) is a common neurodegenerative disorder, characterized by the loss of midbrain dopamine neurons and Lewy body inclusions. It is thought to result from a complex interaction between multiple predisposing genes and environmental influences, although these interactions are still poorly understood. Several causative genes have been identified in different families. Mutations in two genes [alpha-synuclein and nuclear receptor-related 1 (Nurr1)] cause the same pathology, and a third locus on chromosome 2 also causes this pathology. Other familial PD mutations have identified genes involved in the ubiquitin-proteasome system [parkin and ubiquitin C-terminal hydroxylase L1 (UCHL1)], although such cases do not produce Lewy bodies. These studies highlight critical cellular proteins and mechanisms for dopamine neuron survival as disrupted in Parkinson's disease. Understanding the genetic variations impacting on dopamine neurons may illuminate other molecular mechanisms involved. Additional candidate genes involved in dopamine cell survival, dopamine synthesis, metabolism and function, energy supply, oxidative stress, and cellular detoxification have been indicated by transgenic animal models and/or screened in human populations with differing results. Genetic variation in genes known to produce different patterns and types of neurodegeneration that may impact on the function of dopamine neurons are also reviewed. These studies suggest that environment and genetic background are likely to have a significant influence on susceptibility to Parkinson's disease. The identification of multiple genes predisposing to Parkinson's disease will assist in determining the cellular pathway/s leading to the neurodegeneration observed in this disease. PMID- 15297155 TI - Regulation of affect by the lateral septum: implications for neuropsychiatry. AB - Substantial evidence indicates that the lateral septum (LS) plays a critical role in regulating processes related to mood and motivation. This review presents findings from the basic neuroscience literature and from some clinically oriented research, drawing from behavioral, neuroanatomical, electrophysiological, and molecular studies in support of such a role, and articulates models and hypotheses intended to advance our understanding of these functions. Neuroanatomically, the LS is connected with numerous regions known to regulate affect, such as the hippocampus, amygdala, and hypothalamus. Through its connections with the mesocorticolimbic dopamine system, the LS regulates motivation, both by stimulating the activity of midbrain dopamine neurons and regulating the consequences of this activity on the ventral striatum. Evidence that LS function could impact processes related to schizophrenia and other psychotic spectrum disorders, such as alterations in LS function following administration of antipsychotics and psychotomimetics in animals, will also be presented. The LS can also diminish or enable fear responding when its neural activity is stimulated or inhibited, respectively, perhaps through its projections to the hypothalamus. It also regulates behavioral manifestations of depression, with antidepressants stimulating the activity of LS neurons, and depression-like phenotypes corresponding to blunted activity of LS neurons; serotonin likely plays a key role in modulating these functions by influencing the responsiveness of the LS to hippocampal input. In conclusion, a better understanding of the LS may provide important and useful information in the pursuit of better treatments for a wide range of psychiatric conditions typified by disregulation of affective functions. PMID- 15297156 TI - Amyloid-beta: phylogenesis of a chameleon. PMID- 15297157 TI - FIGO Stage IB2 cervix cancer and putting all your eggs in one basket. PMID- 15297158 TI - A thousand several tongues. PMID- 15297159 TI - Surgical-pathological predictors of disease-free survival and risk groupings for IB2 cervical cancer: do the traditional models still apply? AB - OBJECTIVE: To evaluate how the independent predictors of recurrence for stage IB2 cervical cancers treated with up-front radical hysterectomy apply to established risk models. METHODS: Patients with IB2 cervical cancers diagnosed from 1990 to 2000 were identified from tumor registries of two institutions. Patients were classified into risk groups: high-risk (HR) (positive nodes, parametria, or margins), intermediate-risk (IR) (positive lymph vascular space involvement (LVSI) with any cervical stromal invasion (CSI), or (-) LVSI and > middle- CSI), or low-risk (LR) (absence of HR or IR characteristics). Disease-free survival (DFS) was estimated by Kaplan-Meier method and comparisons between subgroups were studied by log rank. A Cox proportional hazards model was used to determine independent predictors of recurrence. RESULTS: We identified 86 patients with stage IB2 tumors treated by RH. We found 34% of patients to be HR, 60% IR, and 6% LR. Of the 52 IR patients, 28 had (+) LVSI with superficial, middle, and outer 1/3 CSI, and 24 had (-) LVSI with middle or outer 1/3 invasion. Overall, postoperative adjuvant radiation (PRT) was used in 52% of the 86 patients, including 0/5 LR, 16/52 IR, and 29/29 HR patients. Univariate predictors of recurrence were pelvic nodal disease, (+) LVSI, (+) parametria, outer 1/3 CSI, and tumor size > 6 cm. Age, grade, histology, and the use of postoperative radiation were not associated with recurrence. Multivariate analysis identified LVSI as the only independent predictor of recurrence (RR 5.2, P = 0.03). Two-year DFS for LR, IR, and HR patients was 100%, 83%, and 60%, respectively. Only 4/24 (17%) IR patients with (-) LVSI got PRT compared with 12/28 (43%) of IR patients with (+) LVSI. The 2-year DFS for IR patients with (-) LVSI was 96%. IR (+) patients recurred more frequently with a 2-year DFS of 71%. CONCLUSIONS: Overall, 66% of patients with IB2 disease were classified as having low or intermediate risk disease. IR patients with (-) LVSI and all LR patients did well with surgery alone. This study defines the independent importance of LVSI and questions the utility of published IR models when applied to stage IB2 cervical cancer. PMID- 15297160 TI - Endometrial pathologies associated with postmenopausal tamoxifen treatment. AB - OBJECTIVE: To evaluate various endometrial pathologies described in association with postmenopausal tamoxifen treatment, as well as the clinical aspects of these endometrial pathologies. METHODS: A search was made in PUB MED for all studies published in English, up to the end of 2003, reporting on endometrial pathologies in association with postmenopausal tamoxifen treatment. Overall 106 studies were available, and all are included in this review. The types of studies included were mostly randomized clinical trials, non-randomized cohort studies, prospective and retrospective case controlled studies. RESULTS: Endometrial polyps represent the most common endometrial pathology associated with postmenopausal tamoxifen exposure. A high rate of malignancy was reported in these polyps. Endometrial hyperplasia, endometrial polyps, endometrial cancer and malignant mixed mesodermal tumors and sarcoma are more commonly diagnosed in postmenopausal breast cancer tamoxifen-treated patients as compared to non treated patients. Long-term tamoxifen users are more likely to succumb to endometrial cancer and endometrial sarcomas than non-users, due to the unfavorable histology of the endometrial malignancy, and an advanced stage of diagnosis. CONCLUSIONS: The clinician should be alerted to these pathologies, which, in some cases, may potentially increase the mortality of these patients. Consequently, it is suggested that their supervision is of importance, especially if the patients experience any gynecological symptoms, including pelvic pain or pressure. PMID- 15297161 TI - Brain metastases of epithelial ovarian carcinoma responding to cisplatin and gemcitabine combination chemotherapy: a case report and review of the literature. AB - BACKGROUND: Brain represents a rare site of metastasis in patients with epithelial ovarian carcinoma (EOC). CASE REPORT: We observed a case of multiple brain metastases in an EOC patient after complete response of a pelvic recurrence to platinum/paclitaxel chemotherapy. Complete response of brain metastases was observed after whole brain radiotherapy and subsequent chemotherapy by combination of cisplatin and gemcitabine. Three subsequent recurrences of brain metastases were controlled by re-treatment by the combination of 5-fluorouracil, cisplatin and gemcitabine. METHODS: Because of limited information on the outcome of EOC brain metastases in reported case series, a pooled analysis of the published reports in patients with EOC brain metastases was performed. Data were extracted from 46 reports that contained sufficient details on 189 individual patients. The survival was analyzed by the Kaplan-Meier method. Univariate and multivariate analyses were performed by the log-rank test and Cox method, respectively. RESULTS: The most favorable outcome was observed in patients treated by surgery combined with radiotherapy and/or chemotherapy. The survival was significantly better in reports describing only one or two cases, in patients diagnosed after 1992, in patients who received therapy in addition to symptomatic treatment, in patients treated by radiotherapy, chemotherapy and surgery, in patients without extracranial metastases and with single brain metastases. On multivariate analysis, the absence of extracranial metastases, treatment by chemotherapy, surgery and radiotherapy were independent positive predictors of survival. CONCLUSIONS: EOC brain metastases are responsive to chemotherapy. An aggressive multidisciplinary therapeutic approach including chemotherapy may lead to prolonged survival. PMID- 15297162 TI - Rectovaginal radiation fistula repair using an obturator fasciocutaneous thigh flap. AB - BACKGROUND: Rectovaginal fistulae are a known complication of pelvic radiotherapy utilizing locally applied isotope implants. Most often, either permanent colostomy or reconstruction with a well-vascularized flap is necessary. Traditional techniques for fistula repair utilize bulky muscle flaps, disfiguring pudendal artery flaps or may require laparotomy. CASE: We describe the management of a 26-year-old woman with a large radiation-induced rectovaginal fistula. A fasciocutaneous medial thigh flap based on terminal branches of the obturator artery and vein was used without colostomy and resulted in pain-free sexual function and minimal vulva disfigurement. CONCLUSION: A medial thigh fasciocutaneous flap without muscle can be transferred into the vagina on the obturator vessels and may become the preferred method for managing large rectovaginal fistulas. PMID- 15297163 TI - The serine protease stratum corneum chymotryptic enzyme (kallikrein 7) is highly overexpressed in squamous cervical cancer cells. AB - OBJECTIVE: To determine whether the Stratum Corneum Chymotryptic Enzyme (SCCE), a novel serine protease known to contribute to the cell shedding process by catalyzing the degradation of intercellular cohesive structures at the skin surface, is overexpressed in human cervical tumors. METHODS: SCCE expression was evaluated in 18 cervical cancer cell lines (i.e., 10 primary and 8 established cell lines) as well as in 8 normal cervical keratinocyte cultures by RT-PCR. In addition, SCCE expression was evaluated by immunohistochemistry on paraffin embedded tumor tissue. RESULTS: Normal cervical keratinocytes did not express SCCE. In contrast, 50% of the primary and 50% of the established cervical cancer cell lines expressed SCCE by RT-PCR. Eighty percent (i.e., four of five) of primary squamous cervical tumors and 20% (i.e., one of five) of primary adenocarcinomas expressed SCCE. Five out of five (100%) of the patients harboring SCCE-positive tumors were found to have metastatic involvement of the pelvic tumor draining lymph nodes. Immunohistochemistry staining of paraffin-embedded cervical cancer specimens confirmed SCCE expression in tumor cells and its absence on normal cervical epithelial cells. CONCLUSION: Squamous cervical cancer expressed high levels of SCCE, suggesting that this protease may play an important role in invasion and metastasis. Because SCCE appears only in abundance in tumor tissue and contains a secretion signal sequence, suggesting that SCCE is secreted, it may prove to be a useful diagnostic/prognostic tool for the detection of metastatic or recurrent disease or as a novel molecular target for cervical cancer therapy. PMID- 15297164 TI - BAG-1 expression in normal and neoplastic endometrium. AB - OBJECTIVE: BAG-1 has anti-apoptotic actions and is known to bind BCL-2 and steroid receptors. High levels of BAG-1 have been implicated as a prognostic indicator in breast cancer. Whether this observation can be generalized to endometrial cancer remains unknown. METHODS: IRB permission was obtained for use of human discarded tissue. Immunohistochemical analyses were performed on: proliferative endometrium (PEM, 6), secretory endometrium (SEM, 28), "low-grade" neoplastic lesions (complex atypical hyperplasia and grade 1 endometrial adenocarcinomas) (19), and "high-grade" cancers (grade 2 and 3 endometrial adenocarcinomas) (13). The level of total BAG-1 and its isoforms was evaluated by Western blot in lysates from Ishikawa cells (grade 1), MFE 296 cells (grade 2), and SK-UT(2) cells (grade 3). RESULTS: The proportion of "high-grade" cancers with positive cytoplasmic staining for BAG-1 was higher than that of secretory endometrium (P = 0.006). Additionally, the proportion of specimens with positive staining for nuclear BAG-1 expression was significantly higher among high-grade carcinoma specimens compared to secretory specimens (P = 0.009). A high proportion (91%) of all specimens were positive for BCL-2, limiting the ability to subcategorize the other variables analyzed. There was no relationship between positive nuclear BAG-1 expression and either estrogen receptor (ER) or progesterone receptor (PR) expression. BAG-1 was expressed in the three cell lines evaluated and total BAG-1 level was not different among the different cell lines. CONCLUSION: BAG-1 is expressed in the endometrium. High-grade cancers stain more frequently than secretory endometrium for both cytoplasmic and nuclear BAG-1 expression, perhaps indicating an association between expression of BAG-1 and prognosis. PMID- 15297165 TI - Quantitative histopathology and chromosome 9 polysomy in a clinical trial of 4 HPR. AB - OBJECTIVE: This trial examined the use of 4-hydroxyphenyl-retinamide (4-HPR), demonstrated to be a potent inhibitor of carcinogenesis in vitro and in animal models, in patients with cervical intraepithelial neoplasia (CIN) grades 2 to 3. Quantitative pathology and chromosome 9 polysomy were used to understand the biology and quantify the clinical histopathologic changes observed. METHODS: Patients were randomized to 4-HPR or placebo for 6 months and followed for six more months. Cervical biopsies were obtained at baseline, 6 months, and 12 months; the biopsies were read blinded three times by the study pathologist. Feulgen-stained sections were also obtained and analyzed using computer-assisted image cytometry. Chromosome 9 polysomy was performed on tissue slices using in situ hybridization and measured quantitatively. Statistical analyses were carried out in S-Plus (Insightful Corporation, Seattle, WA) and R. RESULTS: The interim analysis, planned for 40 patients, was carried out on 39. The 6- and 12-month analyses showed a statistically significant difference between the two study arms. When code was broken, the 4-HPR-treatment arm was found to have fared less well than placebo. Analyses of Feulgen-stained sections provided a quantitative measure of the increase of DNA content and texture features. Chromosome 9 polysomy was also measured using image analysis. The changes observed were consistent with those of cells displaying cancerous changes, indicating a lack of response. CONCLUSION: 4-HPR is not active at 200 mg/day. The interim analysis was helpful in directing the study; and, in this case, ending it. The intermediate endpoint biomarkers of quantitative histomorphometry and chromosome 9 polysomy yielded quantitative and repeatable results consistent with the findings of the clinical pathologist. PMID- 15297166 TI - The benefit of video-assisted thoracoscopic surgery before planned abdominal exploration in patients with suspected advanced ovarian cancer and moderate to large pleural effusions. AB - OBJECTIVE: To analyze the findings and impact on the management of video-assisted thoracoscopic surgery (VATS) before planned abdominal exploration in patients with suspected advanced ovarian cancer and moderate to large pleural effusions. METHODS: We reviewed the charts of all patients with suspected advanced ovarian cancer and moderate to large pleural effusions who underwent VATS from 10/01 to 7/03. VATS was performed under double lumen endotracheal anesthesia. A 2-cm chest wall incision was made in the fifth intercostal space on the side of the effusion. The thoracoscope was introduced and biopsies of suspicious lesions were performed through the single incision. After VATS, all patients had a chest tube placed through the incision, and those with malignant effusions underwent talc pleurodesis either intraoperatively or postoperatively. RESULTS: Twelve patients underwent VATS during the study period. Median operative time for VATS was 31 min (range: 20-49 min) with no complications attributable to the procedure. The median amount of pleural fluid drained was 1000 ml (range: 500-2000 ml). Solid, pleural-based tumor was found in six cases (50%), with nodules >1 cm noted in four patients (33%) and nodules <1 cm noted in two patients (17%). Of the six cases with no grossly visible pleural tumor, the pleural fluid was positive for malignant cells in two patients (17%) and negative in four patients (33%). Further initial patient management included the following: laparotomy with optimal cytoreduction, 6 (50%); diagnostic laparoscopy, 3 (25%); and no abdominal exploration, 3 (25%). Final diagnosis of primary disease site was as follows: ovary, 9 (75%); fallopian tube, 1 (8%); endometrium, 1 (8%); and lymphoma, 1 (8%). Based on the findings during VATS, laparotomy and attempted cytoreduction were avoided in four patients (33%), and the cytoreductive procedure was modified in one patient (8%). CONCLUSION: Fifty percent of patients with suspected advanced ovarian cancer and moderate to large pleural effusions who underwent VATS had solid pleural-based tumor identified, and in 33% of cases the tumor nodules were >1 cm in diameter. VATS should be considered in these cases to delineate the extent of disease, treat the effusion, and to potentially select patients for either intrathoracic cytoreduction or a neoadjuvant chemotherapy approach. PMID- 15297167 TI - Prognostic value of EphA2 and EphrinA-1 in squamous cell cervical carcinoma. AB - OBJECTIVES: To analyze the expressions of the protein and mRNA of EphA2 and EphrinA-1 in squamous cell cervical carcinomas and explore their prognostic value in cervical carcinoma. METHODS: Immunohistochemistry was used to assess the protein expressions of EphA2 and EphrinA-1 in 206 patients with squamous cervical carcinoma FIGO stage Ia-IVb. Frozen tissues from 20 cases in which the tumors showed variable EphA2 and EphrinA-1protein expressions were used for laser capture microdissection (LCM). About 50 cancer cells in each frozen section were captured with the LCM method and processed for RT-PCR detection of EphA2 and EphrinA-1 mRNA. RESULTS: Among the 206 squamous cervical carcinomas, 23 (11.2%) showed negative, 94(45.6%) weakly positive, 72 (35.0%) moderately positive, and 17 (8.3%) strongly positive for EphA2 immunostaining. For EphrinA-1 protein expression, 17 tumors (8.3%) showed negative, 95 (46.1%) weakly positive, 71(34.5%) moderately positive, and 23 (11.2%) strongly positive. EphA2 and EphrinA-1 often colocalized in the same tumor areas and vascular endothelial cells. Variable amount of mRNA expressions of EphA2 and EphrinA-1 were observed in the 20 tumors analyzed. There was no significant correlation between the overexpressions of EphA2/EphrinA-1 and age and FIGO stage. High level of EphA2 was significantly associated with overall survival in univariate and multivariate analysis. Moderate to high EphrinA-1 protein expression was significantly associated with overall survival in multivariate analysis. The combined high level of expression of EhpA2 and moderate to high level of expression of EphrinA 1 was a strong predictor of overall survival. CONCLUSIONS: High levels of EphA2 together with moderate to high level of EphrinA-1 protein expressions in squamous cervical carcinoma were predictive for a shorter overall survival and these proteins may be valuable prognostic markers. PMID- 15297168 TI - Oral dexamethasone attenuates Doxil-induced palmar-plantar erythrodysesthesias in patients with recurrent gynecologic malignancies. AB - OBJECTIVE: The aim of this study is to evaluate the effect of oral dexamethasone in attenuating palmar-plantar erythrodysesthesias (PPE) in Doxil-treated patients for gynecologic malignancies. METHODS: An IRB-approved prospective case study was conducted in patients with recurrent gynecologic malignancies who were treated with Doxil (50 mg/m(2)) on a 28-day cycle. Patients experiencing grades II-IV PPE were delayed until resolution then retreated without dose reduction and with a tapering oral dexamethasone regimen (8 mg BID days -1 to 4; 4 mg BID day 5; 4 mg day 6). Standard treatment for grades II-IV PPE in those not receiving dexamethasone was weekly dose delay until resolution of symptoms up to 2 weeks. If resolution occurred within 3 weeks of delay, a 25% dose reduction was made. Persistent grades III/IV PPE resulted in withdrawal of Doxil. RESULTS: Twenty three patients (ovarian-16, uterine-7) were treated between January 1998 and December 2000. The median number of cycles administered was 5 (range 1-20). Nine patients (39%) developed grades II-IV PPE. All nine patients received more than five cycles of Doxil. The median time to PPE was 3 cycles (range 2-5). Six out of nine PPE patients received scheduled dose dexamethasone. All six had complete or near complete resolution of PPE and all continued treatment without subsequent dose modification. All three of the nine PPE patients not receiving dexamethasone required treatment delays and were dose reduced. CONCLUSION: Oral dexamethasone is effective in attenuating or eliminating Doxil-induced PPE. The use of the dexamethasone regimen prevents treatment delay and dose reduction. PMID- 15297169 TI - Effects of intraperitoneal hyperthermic chemotherapy in ovarian cancer. AB - OBJECTIVES: To evaluate the clinical effect of intraperitoneal hyperthermic chemotherapy (IPHC) in ovarian cancer patients. PATIENTS AND METHODS: We retrospectively reviewed 117 stages Ic-III ovarian cancer patients, who were diagnosed at the Gynecology Department of Kangnam St. Mary's Hospital between January 1994 and January 2000. Of these, 57 patients underwent cytoreductive surgery (conventional treatment) with IPHC and 60 patients (control group) underwent conventional treatment only. IPHC consisted of administering a mixture of 350 mg/m(2) of carboplatin and 5,000,000 IU/m(2) of interferon-alpha, and maintaining the intraperitoneal temperature at 43-44 degrees C during surgery. RESULTS: The overall 5-year survival rate was 58.6%; that of the IPHC group was 63.4% vs. 52.8% in the control group, with significantly higher survival in the IPHC group (P = 0.0078). Considering stage III ovarian cancer patients only (n = 74), the survival rate was 53.8% in the IPHC group (n = 35) and 33.3% in the control group (n = 39) and was significantly higher in the IPHC group (P = 0.0015). For stage III ovarian cancer patients whose tumor was reduced to less than 1 cm during a second procedure (n = 53), the 5-year survival rate was 65.6% in patients who underwent IPHC (n = 26) and 40.7% in the control patients (n = 27) (P = 0.0046). IPHC was an independent prognostic factor that was not affected by surgical staging, tumor size after second surgery, or patient age, according to a multivariate analysis (Hazard ratio = 0.496, P = 0.0176). CONCLUSION: Our study suggests that IPHC is a promising new treatment modality in ovarian cancer. PMID- 15297170 TI - Adjuvant chemotherapy as treatment of high-risk stage I and II endometrial cancer. AB - OBJECTIVE: This study was performed to define the subgroups of patients who benefit from postoperative adjuvant chemotherapy in stage I and II endometrial carcinoma. METHODS: A retrospective review of 170 International Federation of Gynecology and Obstetrics (FIGO) stage I and II endometrial carcinoma patients treated between 1988 and 2000 at Niigata University Hospital was performed. All patients underwent surgery, of which 41 patients underwent adjuvant chemotherapy, consisting of intravenous cisplatin, doxorubicin, and cyclophosphamide. Multivariate analysis was performed for the prognostic factors and actuarial techniques were used for the survival and recurrence rates. RESULTS: The patients were divided into low-risk and high-risk groups based on the number of prognostic factors (tumor grade G3, outer half myometrial invasion, lymph-vascular space involvement (LVSI), and cervical invasion). The 5-year disease-free survival and the 5-year overall survival for the low-risk group were 97.4%, and 100%, respectively, which were significantly better than 77.4% and 88.1% for the high risk group (P < 0.0001, P < 0.0001), respectively. Among high-risk group patients, the 5-year disease-free survival and the 5-year overall survival were 88.5% and 95.2% in 26 patients treated with adjuvant chemotherapy, and 50.0% and 62.5% in eight cases who underwent only surgery (P = 0.0150, P = 0.0226). Disease recurrence occurred in 7 (20.6%) of 34 high-risk group patients. Four of seven recurrences occurred in patients who did not receive postoperative chemotherapy, in which all four were distant failure. In the remaining three patients who were in the CAP group, two had vaginal wall recurrence and only one had pulmonary recurrence. Three recurrences were also observed in the 133 low-risk group patients. Only isolated vaginal wall recurrence occurred in three patients without adjuvant chemotherapy after the initial surgery. CONCLUSIONS: There is possibility that postoperative adjuvant CAP may be omitted in surgical stage I or II endometrial cancer patients with 0 or 1 prognostic factor. The high-risk group of patients should be treated with postoperative adjuvant CAP to decrease distant failure and improve prognosis. PMID- 15297171 TI - CA125- and tumor-specific T-cell responses correlate with prolonged survival in oregovomab-treated recurrent ovarian cancer patients. AB - OBJECTIVE: To evaluate immune responses and clinical outcomes for combined oregovomab and chemotherapy treatment of patients with recurrent ovarian cancer. METHODS: Patients with advanced recurrent ovarian cancer were administered oregovomab over 12 weeks before chemotherapy, then optionally concurrent with chemotherapy x 2. Antibody responses, including human anti-mouse antibody (HAMA), anti-idiotypic antibody (Ab2) and anti-CA125, were assessed by ELISA; T-cell responses to CA125, autologous tumor and oregovomab by interferon (IFN)-gamma enzyme-linked immunoSPOT (ELISPOT) were also evaluated. Clinical outcomes were recorded. RESULTS: Twenty patients were enrolled; median follow-up was 15.8 months. Oregovomab was well tolerated and did not produce drug-related serious adverse reactions. In 15/19 (79%) patients, robust treatment-emergent humoral responses were observed to the constant (HAMA) and variable region (Ab2) of oregovomab, and 2/19 (11%) patients developed anti-CA125 antibodies. Significant increases in T-cell responses were measured in 7/18 (39%) patients in response to CA125, in 5/8 (63%) patients in response to autologous tumor and in 9/18 (50%) patients in response to oregovomab. Immune responses appeared by week 12 (four doses) and were generally maintained or augmented in patients continuing combined treatment with oregovomab and chemotherapy. Median survival was 70.4 weeks (4.6 141.6 weeks), and the median progression-free interval was 11 weeks (2.6-114.6 weeks). Patients who mounted a T-cell response to CA125 and/or autologous tumor showed significantly improved survival (median not reached vs. 51.9 weeks, P = 0.002) compared to patients who did not. CONCLUSIONS: Oregovomab was well tolerated and induced multiple antigen-specific immune responses, maintained during concomitant chemotherapy. A significant survival benefit was observed in patients mounting a T-cell response to CA125 and/or autologous tumor. PMID- 15297172 TI - Infectivity enhanced adenoviral-mediated mda-7/IL-24 gene therapy for ovarian carcinoma. AB - OBJECTIVE: Melanoma differentiation associated gene-7 [mda-7/Interleukin (IL)-24] has been identified as a novel anti-cancer agent, which specifically induces apoptosis in cancer cells but not in normal epithelial, endothelial and fibroblast cells. The objective of this study was to evaluate the anti-tumor effect of adenovirus-mediated mda-7/IL-24 (Ad.mda-7) gene therapy in ovarian carcinoma and further improve anti-tumor effect by enhancing infectivity of Ad.mda-7. METHODS: A panel of human ovarian carcinoma cells, OV-4, HEY, SKOV3, SKOV3.ip1 and control normal human mesothelial cells, were infected by a replication deficient recombinant adenovirus encoding mda-7/IL-24 and control virus Ad.CMV.Luc. After 72 h, apoptosis was evaluated by TUNEL and Hoechst staining and further quantified by fluorescent activated cell sorter (FACS) analysis. Infectivity of Ad.mda-7 was enhanced by retargeting it to CD40 or EGF receptors overexpressed on ovarian cancer cells. Subsequently, enhancement in apoptosis of CD40- or epidermal growth factor receptor (EGFR)-retargeted Ad.mda-7 was evaluated. RESULTS: Adenoviral-mediated delivery of mda-7 induces apoptosis ranging from 10-23% in human ovarian cancer cells tested with the highest percentage of apoptosis noted in SKOV3 cells. Minimal apoptosis was noted in normal mesothelial cells. CD40- or EGFR-retargeted Ad.mda-7 increased apoptosis by 10-32% when compared to that achieved with untargeted Ad.mda-7. CONCLUSION: Ad.mda-7 exhibits ovarian cancer-specific apoptosis, but does not affect normal human mesothelial cells. Infectivity enhanced CD40- and EGFR-retargeted Ad.mda-7 augments apoptosis induction, thus increasing the therapeutic index and translational potential of Ad.mda-7 gene therapy. PMID- 15297173 TI - The role of the preoperative serum carcinoembryonic antigen level in early-stage adenocarcinoma of the uterine cervix. AB - OBJECTIVE: The purpose of the study was to identify the relationship between preoperative serum levels of carcinoembryonic antigen (CEA) and clinicopathological variables in early-stage adenocarcinoma of the uterine cervix. METHODS: From February 1990 to August 2002, 117 patients with surgically treated early-stage cervical adenocarcinoma that had had preoperative serum CEA evaluations were retrospectively reviewed. The cut-off value for CEA, based on the manufacturer's recommendations, was 5 ng/ml. For an evaluation of the relationship between the clinicopathological factors and increased levels of serum tumor markers, the Chi-Square/Fisher's exact test and logistic regression were used for univariate and multivariate analysis, respectively. RESULTS: The mean age of the patients was 46 years (range, 21-78). Of the 117 patients, 28 had preoperative serum CEA levels greater than 5 ng/ml. In a univariate analysis, the increased marker was associated with a larger tumor size, presence of lymphovascular invasion, and deeper cervical wall invasion. However, in a multivariate analysis, the preoperative CEA level had a significant impact on the determination of the depth of stromal invasion (OR 4.12, 95% CI 1.97-8.68, P < 0.001). CONCLUSION: In early-stage cervical adenocarcinoma, preoperative serum CEA levels seem to be useful in estimating the depth of cervical stromal invasion. Assessment of tumor antigen CEA levels should be integrated with the routine examination in the work-up of patients with adenocarcinoma of the uterine cervix. PMID- 15297174 TI - An assessment of age and other factors influencing protocol versus alternative treatments for patients with epithelial ovarian cancer referred to member institutions: a Gynecologic Oncology Group study. AB - OBJECTIVE: To assess the frequency with which patients with epithelial ovarian cancer are enrolled in prospective Gynecologic Oncology Group (GOG) treatment studies and to assess whether enrollment is influenced by patient age or other factors. METHODS: The study was open to patients with primary, previously untreated epithelial ovarian cancer referred to member institutions. Eligible patients provided written informed consent to have demographic and medical records data submitted to the protocol database for analysis. Pathologic diagnosis was confirmed by central review. Demographic and clinical data, including coexisting medical conditions, tumor stage, grade, and histology, surgical procedures, planned postoperative therapy, and reason/s (if applicable) a patient was not treated per GOG protocol, were collected. RESULTS: Nine hundred and forty-eight patients were initially eligible. Subsequently, 137 (15%) patients were excluded based on pathology (low malignant potential tumors), as were 10 whose invasive disease was unstaged. Among 801 eligible patients, 36% were > or =65 years of age, 52% had papillary serous tumors, and 73% had stage III/IV disease. In patients aged <65 years, 67% were stage III/IV compared to 82% of patients > or =65 years of age. The mean age was 5.5 years greater for patients with stage III/IV versus stage I/II disease. Compared to their younger counterparts, older patients with stage III/IV disease were less likely to enter into a GOG treatment protocol. Most common reasons were patient ineligibility (33%), refusal (29%), and investigator decision (20%). CONCLUSION: Age appears to be an important factor influencing treatment selection among patients with stage III/IV ovarian cancer. In addition to reviewing eligibility criteria, practitioners' attitudes should be monitored to assure that elderly patients are not inappropriately denied participation in GOG clinical trials. PMID- 15297175 TI - Down-regulation and promoter methylation of tissue inhibitor of metalloproteinase 3 in choriocarcinoma. AB - OBJECTIVE: To assess the differential gene expression in neoplastic and normal trophoblastic cells and evaluate the effect of methylation on tissue inhibitor of metalloproteinase 3 (TIMP3) expression in choriocarcinoma (CCA) cells. METHODS: The Atlas Human Cancer 1.2 Array (Clontech) was used to compare differential gene expression in a trophoblastic cell line (B6) established from first term placenta with two choriocarcinoma cell lines (JAR and JEG-3). The differentially expressed candidate genes in the placental and malignant trophoblastic cells in these cell lines were confirmed by real-time polymerase chain reaction (PCR), Western blotting and immunohistochemistry. Differential expression of a specific gene, TIMP3, was confirmed by immunohistochemistry using clinical specimens of choriocarcinoma and placenta. The involvement of promoter methylation in suppression of TIMP3 expression in choriocarcinoma was examined using methylation specific PCR (MSP) and demethylation treatment. RESULTS: Differential expression of 23 genes was observed in choriocarcinoma cell lines compared to the placental trophoblastic cells using the cDNA array analysis (Atlas Human Cancer Array, Clontech). Among these differentially expressed genes, down-regulation of TIMP3, PLAB, IGFBP3 and up-regulation of CCNB1 were confirmed by real-time PCR determination. Reduced expression of TIMP3 was further confirmed in clinical samples of choriocarcinoma by immunohistochemical staining. Methylation of TIMP3 promoter was detected in choriocarcinoma cell lines and clinical samples of choriocarcinoma. Treatment with a demethylation drug, 5-aza-2'-deoxycytidine, in choriocarcinoma cell lines restored TIMP3 expression. CONCLUSION: Down-regulation of TIMP3, PLAB, IGFBP3 and up-regulation of CCNB1 were observed in choriocarcinoma cells compared to placental trophoblasts. Down-regulation of TIMP3 expression was confirmed in clinical specimens of choriocarcinoma and may play a role in its pathogenesis. Promoter methylation of the TIMP3 is involved in suppression of TIMP3 expression. Differentiation expression of TIMP3 in choriocarcinoma may have potential application in clinical diagnosis and patient treatment. PMID- 15297176 TI - High-dose rate brachytherapy for Stage I/II papillary serous or clear cell endometrial cancer. AB - OBJECTIVE: To determine the efficacy of high-dose rate brachytherapy as adjuvant treatment for Stage I/II papillary serous or clear cell endometrial cancer. METHODS: A retrospective study of all patients with Stage I/II papillary serous or clear cell endometrial cancer treated with high-dose rate brachytherapy between 1995 and 2001 was performed. Following surgical staging, which included hysterectomy with pelvic and aortic lymphadenectomy, all patients without extrauterine disease were treated with high-dose rate brachytherapy and followed for recurrence. The locations of recurrences were noted and were classified as local or distant. RESULTS: Three (13%) recurrences occurred among 24 patients with Stage I/II papillary serous or clear cell carcinoma. The risk of recurrence was similar for papillary serous and clear cell cancer (12% vs. 12%). Local control was achieved in 96%. The risk of recurrence for those with no myometrial invasion, less than 1/2, or more than 1/2 myometrial invasion was 0%, 10%, and 50%, respectively (P < 0.04). Two of the three recurrences were distant and all patients with recurrence died despite additional treatment. CONCLUSIONS: High dose rate brachytherapy (HDR) as the sole adjuvant treatment of Stage I/II papillary serous or clear cell carcinoma is associated with a 13% risk of recurrence. Although local control with HDR is excellent, the risk of distant recurrence is increased with deep myometrial invasion. High-dose rate brachytherapy is adequate for Stage IA cases, but more aggressive treatment combining chemotherapy with HDR should be evaluated for more advanced Stage I/II cases. PMID- 15297177 TI - Are borderline tumors of the ovary safely treated by laparoscopy? AB - OBJECTIVES: To evaluate the risk of the laparoscopic approach to patients with borderline ovarian tumors compared to the laparotomic management. METHODS: We treated or followed in our institution 479 women with borderline ovarian tumor. Sixty-two patients had fertility-sparing surgery followed by restaging or follow up intervention: 30 operated by laparoscopy, 32 by laparotomy. Restaging surgery was performed in five cases and second-look surgery in 57. RESULTS: The diameter of the cyst is significantly lower in patients treated by laparoscopy, especially in women who underwent cystectomy (4.7 cm) compared to oophorectomy (10 cm, P = 0.008). Rupture of the cyst and stage IC were more frequent in the laparoscopic group. After a median follow-up of 61 months for the laparoscopic group and 77 months for the laparotomic group, we observed 11 patients (37%) with persistent disease after primary laparoscopy (adnexa, five cases; peritoneal implants, three cases; both patterns, three cases). After primary laparotomy, no patients showed early persistence of tumor, and ovarian relapses were diagnosed in seven women (22%) 33-138 months after laparotomy. The univariate analysis for the risk of neoplastic persistence after primary laparoscopy shows that patients with cysts greater than 5 cm have a higher risk (odds ratio 9.7, P = 0.02) compared to smaller cysts. No other factors proved significant, but the odds ratios for patients with serous tumor (5.8), stage IC (2.0), and those undergoing cystectomy (1.9) suggest a relationship to the probability of persistence. CONCLUSION: Laparoscopic treatment in borderline ovarian tumors should be reserved to masses not greater than 5 cm. When conservative therapy is desired, the entire affected ovary should be removed. If the neoplasia is bilateral, cystectomy could be allowed in women who wish to preserve fertility, although they are at high risk of relapse. PMID- 15297178 TI - Triplet combination of gemcitabine, carboplatin, and paclitaxel in previously treated, relapsed ovarian and peritoneal carcinoma: an experience in Taiwan. AB - OBJECTIVE: The aim of this phase II study was to evaluate the efficacy and toxicity of gemcitabine, carboplatin, and paclitaxel (GCP) combination as salvage therapy in patients with relapsed ovarian or peritoneal cancer who had previously received platinum-based chemotherapy. PATIENTS AND METHODS: Patients with progressive ovarian or peritoneal carcinoma who had previously received platinum based chemotherapy were enrolled. Gemcitabine was administered at 800 mg/m(2) as a 30-min intravenous infusion on days 1 and 8; carboplatin (AUC of 5) and paclitaxel (175 mg/m(2)) were administered as 60-min and 3-h intravenous infusions, respectively, on day 1. Treatment cycles were repeated every 3 weeks for a maximum of nine cycles. RESULTS: Twenty patients (ovarian carcinoma, 19; peritoneal carcinoma, 1) received this triplet regimen as salvage therapy. All the patients had previously received at least one platinum-based regimen for chemotherapy and 17 of them had received platinum plus paclitaxel. The median number of previous regimens was 2 (range, 1-4), and the median platinum-free interval was 9 months (range, 1-18). A total of 130 cycles were administered with a median of six cycles per patient (range, 3-9). The overall response rate was 75%, including 12 complete responses (60%; 95% confidence interval [CI], 36.1 80.9) and three partial responses (15%; 95% CI, 3.2-37.9). The other five patients showed stable disease (25%; 95% CI, 8.7-49.1). The median duration of the progression-free survival was 6.5 months (range, 3-20). Myelosuppression was the main toxicity, with leukopenia being the most prominent (grade 3/4 toxicity in 35% patients), followed by thrombocytopenia in 20% patients. In addition, 35% patients had grade 3 anemia. All the toxicities were manageable and the patients recovered fully. Among non-hematological toxicities, the only notable one was grades 2 and 3 hepatic toxicity seen in two and one patients, respectively, necessitating a decrease in the paclitaxel dose in two patients. CONCLUSIONS: GCP combination is an effective salvage chemotherapy in patients with heavily pretreated and relapsed ovarian and peritoneal cancer. The significant side effects of myelosuppression and hepatic toxicity were of moderate severity and manageable. PMID- 15297179 TI - The impact of treatment for genital cancer on quality of life and body image- results of a prospective longitudinal 10-year study. AB - OBJECTIVE: To evaluate the impact of treatment for genital cancer on quality of life and body image to determine patients' therapy-related needs for quality improvement of medical care before and after surgery. METHODS: We started to evaluate women with cervical cancer planned for pelvic exenteration in 1993 and integrated women planned for a Wertheim-Meigs surgery in 1995 before surgery, 4 and 12 months after surgery. Thanks to funding since 1999, more than 400 patients with a diagnosis of genital (n = 185) or breast (n = 217) cancer participated in this prospective study until July 2003. In this paper, we will focus on n = 129 women with cervical cancer. The assessment protocol included objective questionnaires for quality of life and body image (CARES; EORTC; Body image by Strauss and Appelt). The evaluation of quality of life incorporated five dimensions: physical and psychosocial health, marital and sexual status, and medical interaction. RESULTS: Before surgery, women with a Wertheim's procedure indicated significantly less problems concerning the quality of life global score (P = 0.002) and several subscales compared to women with a pelvic exenteration. After surgery, both groups indicated their sexual problems to be the greatest restriction in terms of quality of life, especially in women with non reconstructive surgery as well as in women with adjuvant radio and/or chemotherapy. Concerning body image, attractiveness or self-confidence was significantly reduced postoperatively compared to the preoperative status for both groups (P = 0.000), and also worsened with the extent of treatment. Worries about the patient's family persisted over time and represented the most important item about all questions concerning quality of life as well as the fear of recurrence. CONCLUSION: This on-going study demonstrates the interferences between the treatment modality and the patient's quality of life, especially about sexuality and body image. Our results suggest not only to provide reconstructive surgery if possible, but also to integrate psychosocial information aspects on future quality of life outcome before surgery as well as to offer psychosocial support related to the extent of treatment modality after surgery. PMID- 15297180 TI - Phase 2 trial of carboplatin plus tamoxifen in platinum-resistant ovarian cancer and primary carcinoma of the peritoneum. AB - OBJECTIVES: A previously reported phase 2 trial suggested substantial clinical activity associated with the combination of a platinum agent and tamoxifen in the treatment of platinum-resistant ovarian cancer. We wished to confirm or refute this observation in a patient population with well-characterized platinum resistant disease. METHODS: Patients with ovarian or fallopian tube cancers or primary carcinoma of the peritoneum whose disease had either failed to respond to a platinum-based regimen or had responded but experienced a "treatment-free interval (TFI)" of < or =3 months, or if the TFI was >3 months they had been retreated and failed a platinum-based program, were eligible for entry into this phase 2 single institution protocol. Carboplatin (AUC 5) was delivered on a q-21 day cycle. Tamoxifen was administered at a dose of 80 mg/day for the first cycle, and then reduced to 40 mg/day. Treatment was to be continued until evidence of disease progression or unacceptable toxicity. RESULTS: Fourteen patients were treated on this phase 2 trial. In addition to being platinum-resistant, 10 patients had cancers that were also documented to be taxane-resistant (similar criteria to that defined above for platinum). While treatment was generally well tolerated, there were no objective (measurable disease or CA-125 response criteria) or subjective responses to this treatment program. CONCLUSION: In this phase 2 trial, we have been unable to confirm a meaningful level of clinical activity for the combination of carboplatin plus tamoxifen in a patient population with well-characterized platinum-resistant ovarian cancer. PMID- 15297181 TI - Is stabilization of disease a useful indicator for survival in second-line treatment of ovarian carcinoma pre-treated with Paclitaxel-Platinum? AB - OBJECTIVE: Recurrent ovarian carcinoma is considered an incurable disease and second-line chemotherapy is administered for extension of survival and palliation. The impact of continued antineoplastic treatment in patients with stable disease without a demonstrable response is uncertain. The aim of this analysis was to assess the value of a stabilization of the tumor size in second line chemotherapy as an indicator of survival. METHODS: Retrospective, single institution study of 487 consecutive patients with primary epithelial ovarian carcinoma. INCLUSION CRITERIA: (1) FIGO stage IC-IV epithelial ovarian carcinoma; (2) first-line chemotherapy with Paclitaxel and a Platinum-compound; (3) refractory, persistent, or recurrent disease diagnosed by imaging methods; and (4) intravenous second-line chemotherapy with single Topotecan or Paclitaxel Carboplatin. Univariate and multivariate analyses of survival with the World Health Organization (WHO) tumor response parameter included as a time-dependent variable were performed. RESULTS: The response rates were (N = 100): complete response (CR) 27%, partial response (PR) 14%, stable disease (SD) 41% and progressive disease (PD) 18%. In a multivariate Cox regression analysis of survival, SD was found to be an independent prognostic factor for survival and the death hazard ratio was 0.37 (SD versus PD; 95% CI: 0.16-0.86; P = 0.02). There was no statistically significant difference in survival between patients with PR and SD (P = 0.09). CONCLUSION: In second-line chemotherapy of ovarian cancer, patients demonstrating SD have a survival benefit compared to patients with PD measured by the WHO tumor response criteria. PMID- 15297182 TI - Marked allelic imbalance on chromosome 5q31 does not explain alpha-catenin expression in epithelial ovarian cancer. AB - OBJECTIVE: Human alpha-catenin gene (CTNNA1) on chromosome 5q31 is aberrantly expressed in various types of cancer including epithelial ovarian tumors. Allelic imbalance on this region has also been described in several malignant diseases. In the present work, the role of CTNNA1 as a candidate tumor suppressor gene was studied by comparing protein expression with allelic imbalance in human epithelial ovarian tumors. METHODS: Alpha-catenin protein expression was determined from two areas of 41 tumors, and tissues from these areas were microdissected. After DNA extraction, AI analysis was carried out with eight microsatellite markers. RESULTS: Altogether, 93% of the tumors (38 of 41) showed allelic imbalance at one or more loci. Two distinct common regions of allelic imbalance were identified, one comprising markers D5S2002 and D5S1995 and the other markers D5S393 and D5S476. Loss of the CTNNA1 gene did not appear to be involved in down-regulation of alpha-catenin in ovarian tumors, since allelic imbalance with a variety of markers, including CTNNA1 associated marker D5S476, was found in tumor samples independently of alpha-catenin expression. Furthermore, allelic imbalance analyses of two different samples from the same tumor revealed genetic heterogeneity. CONCLUSIONS: High allelic imbalance frequency indicates that chromosomal region 5q31 is functionally important in epithelial ovarian cancer. Allelic imbalance occurs at two distinct regions of which one includes the CTNNA1 gene. However, this gene is likely to be inactivated by mechanisms other than allelic imbalance. In addition, genetic heterogeneity observed in these tumors demonstrates the multiclonal nature of epithelial ovarian tumors. PMID- 15297183 TI - Hypoxia up-regulates the effects of prostaglandin E2 on tumor angiogenesis in ovarian cancer cells. AB - OBJECTIVE: Emerging evidence supports a role for prostaglandins (PG) and their synthesizing enzyme, cyclooxygenase (COX), in tumor angiogenesis. The objective of this study was to investigate the effects of prostaglandin E(2) (PGE(2)) on the expression of vascular endothelial growth factor (VEGF) and hypoxia-inducible transcription factor-1 alpha (HIF-1alpha) genes in epithelial ovarian cancer (EOC) cells. METHODS: Two human EOC cell lines, MDAH-2774 and SKOV-3, were treated with exogenous dimethyl prostaglandin E(2) (dmPGE(2)) at two doses of 10 and 50 microg/ml and cultured for 24 h under both normoxic and hypoxic conditions. Total RNA was extracted from EOC cells with the use of a monophasic solution of phenol and GITC/Trizol method. The levels of COX-2, VEGF, and HIF 1alpha mRNA were measured by quantitative reverse transcription PCR (RT-PCR). RESULTS: Under normoxic conditions, treatment of both ovarian cancer cell lines with dmPGE(2) resulted in a significant increase in VEGF expression but had no effect on HIF-1alpha. Culturing the cells under hypoxic conditions resulted in an increase in HIF-1alpha and VEGF mRNAs. The combination of hypoxia and dmPGE(2) treatment resulted in the highest levels of VEGF and HIF-1alpha when compared to either individual treatment. CONCLUSION: PGE(2) is a potent stimulator of VEGF expression in ovarian cancer cells. This effect of PG is further potentiated under hypoxic conditions where it is also associated with a significant increase in HIF-1alpha expression. PMID- 15297184 TI - Attitudes to chemotherapy in patients with ovarian cancer. AB - BACKGROUND: Recurrent ovarian cancer (OVCA) has become the model for cancer as a chronic disease, yet little is known about what motivates patients and physicians in treatment choices. METHODS: We investigated the attitudes of patients with epithelial OVCA and staff towards palliative chemotherapy for recurrent OVCA with a cross-sectional questionnaire study. RESULTS: Instruments were developed and piloted in 15 patients. This exploratory study reflects substantial bias in the sample populations. One hundred twenty-two patients and 37 staff were enrolled in the US and 39 patients and 25 staff were enrolled in the UK. UK patients had a lower educational status (P = 0.001), lower stage disease (P = 0.025), and less prior lines of chemotherapy (P < 0.001). 61% of patients had recurrent OVCA and 67% of staff were physicians. Seventy-three percent of patients recalled a discussion about prognosis and 74% wanted to know details of the prognosis for a typical patient (US = UK). Most patients (48%) thought that their physician was realistic, and 57% of staff felt that they were optimistic. The vast majority of both staff and patients thought that patients positively reinterpreted what they were told. Five percent of staff thought that palliative care was "incompatible" when considering chemotherapy as an option for their second recurrence of OVCA, compared with 36% of US patients, significantly more than the 12% of UK patients (P = 0.007). Patients thought that standard chemotherapy for a second recurrence of OVCA produced remission in 50% and cure in 15% of patients. Staff reported 20% and 0%, respectively. Fifty percent of patients and 57% of staff would want chemotherapy as an asymptomatic patient with a normal CT and a rising CA-125. Patients generally appear to be very tolerant of grade II chemotherapy-induced toxicity with staff being less tolerant than patients of nausea, anorexia, diarrhea, and rash. Staff rated life prolongation by 3 months to 1 year very much less acceptable than patients (P < 0.001). Although possibly allowing comprehensive collection of sensitive data, the questionnaire was too distressing for some patients and made 11% of patients feel uncomfortably anxious. CONCLUSIONS: Patients are optimistic and in the US, may be more reluctant than staff to see the Palliative Care Team. These data challenge the assertion that the use of palliative chemotherapy is physician-driven. PMID- 15297185 TI - Panniculectomy: improving lymph node yield in morbidly obese patients with endometrial neoplasms. AB - OBJECTIVE: Panniculectomy has been used to facilitate pelvic surgery in obese women. The goal of this study was to determine the effect of panniculectomy on staging adequacy and lymph node yield in obese women with endometrial carcinoma undergoing staging laparotomy. METHODS: A retrospective review of patients with endometrial neoplasms who underwent panniculectomy at the time of hysterectomy was performed. For each subject, two control patients were matched by body mass index (BMI). RESULTS: Twenty-seven endometrial cancer patients who underwent panniculectomy at the time of staging were identified. Panniculectomy was successfully performed in all 27 patients. While the mean number of pelvic nodes was statistically similar between the two groups (16.2 vs. 13.7) (P = 0.199), the paraaortic node count was higher in patients who underwent panniculectomy (4.3 vs. 2.9) (P = 0.032). A paraaortic node dissection was not feasible in 3 (11.1%) of the panniculectomy patients and in 11 (20.4%) of the controls (P = 0.365). There were no differences in intraoperative or postoperative complications or in survival between the two groups. CONCLUSION: Among obese women with endometrial cancer, panniculectomy is well tolerated, feasible, and associated with acceptable morbidity. While the clinical significance of an increased paraaortic node count is uncertain, our findings suggest that panniculectomy may enhance operative exposure and facilitate endometrial cancer staging. PMID- 15297186 TI - A phase I study of Adp53 (INGN 201; ADVEXIN) for patients with platinum- and paclitaxel-resistant epithelial ovarian cancer. AB - PURPOSE: Adenoviral p53 (Adp53) is a replication-deficient adenovirus containing human p53 cDNA. This phase I study was designed as a toxicity study of multiple dosing of Adp53 administered by intraperitoneal (IP) delivery to patients with ovarian cancer. EXPERIMENTAL DESIGN: Eligibility criteria included patients with platinum- and paclitaxel-resistant metastatic epithelial ovarian cancer; a Zubrod performance status of 0, 1, or 2; and adequate bone marrow, liver, and renal function. Patients underwent laparoscopy, washings, biopsies, and placement of an IP catheter within 10 days of Adp53 administration. Adp53 was given daily for 5 days every 3 weeks at one of the following four dose levels: 3 x 10(10), 3 x 10(11), 1 x 10(12), or 3 x 10(12) viral particles (vp). RESULTS: Seventeen patients were enrolled in the trial. Fifteen (88%) patients are evaluable for toxicity. The mean age of the study group was 51 years (range 32-67). All but one patient received two or more chemotherapy regimens before study entry. No dose limiting toxicities (DLT) were observed. Grade 3 toxicities included fatigue (six patients), fever (two patients), chills (one patient), abdominal pain (three patients), nausea (two patients), and sinus congestion (one patient). One patient had Grade 3 edema and headache. There were no hematologic toxicities. Eleven patients (65%) are evaluable for response. Two of 17 patients (12%) had a mixed response. Four patients (24%) had stable disease for up to four courses. Five patients (29%) had progressive disease after one to two courses. CONCLUSIONS: Multiple dosing of IP Adp53 was well tolerated in this group of heavily pretreated patients; however, the dosing schedule and the amount cannot be concluded from this study. With a negative randomized trial of ovarian cancer in front-line treatment that included an adenovirus p53 plus chemotherapy, we feel that further refinement of gene therapy is required before additional trials are undertaken. OVERVIEW SUMMARY: Ovarian cancer is the most lethal of the gynecologic malignancies. It also tends to recur and progress within the abdominal cavity. Because of this, regional intraperitoneal therapy for ovarian cancer is attractive. Mutation and/or deletion of the p53 gene are common in advanced ovarian cancer. In this study, we have tested the safety and practicality of using an adenovirus-mediated delivery of the p53 gene to patients with chemo-refractory ovarian cancer via an intraperitoneal catheter. Fifteen patients were treated. Common toxicities were abdominal pain, fever, and chills. Several patients also had catheter infections. One patient had prolonged decrease in CA125 and stable disease. The best mechanism of delivery of gene therapy for patients is unclear, however, no severe toxicities were found using an adenovirus mediated p53 gene in this group heavily pretreated patients with recurrent ovarian cancer. PMID- 15297187 TI - Wilms Tumor Gene (WT1) and p53 expression in endometrial carcinomas: a study of 130 cases using a tissue microarray. AB - OBJECTIVE: With the exception of ovarian serous carcinoma, Wilms tumor suppressor gene (WT1) expression in common gynecologic carcinomas has not been described in detail. We studied a large number of endometrial carcinomas to determine the range of tumors that express WT1; this could have prognostic and therapeutic significance. METHODS: We studied the immunohistochemical expression of WT1 and p53 in 130 primary human endometrial carcinomas of various histological subtypes, grades, and stages using a tissue microarray. The clinical data were retrieved from the medical records. RESULTS: WT1 was expressed in a wide variety of endometrial cancers and was most marked in malignant mixed Mullerian tumors (MMMTs) (70% positive). WT1 expression was significantly correlated with high histological grade, and there was a trend toward a worse clinical outcome for patients whose tumors expressed WT1. An association between expression of WT1 and p53 and between these and outcome was noted in a univariate analysis, but only stage and p53 status remained prognostically significant independent variables. CONCLUSION: WT1 is expressed in appreciable numbers of endometrial cancers, particularly MMMTs. These findings support further investigation of WT1 as a possible therapeutic target in gynecologic malignancies. PMID- 15297188 TI - Synchronous primary cancers of the endometrium and ovary: a single institution review of 84 cases. AB - OBJECTIVES: Synchronous primary cancers of the endometrium and ovary are found in 10% of women with ovarian cancer and 5% of women with endometrial cancer. The purpose of this study was to characterize patients diagnosed with synchronous primary cancers of the endometrium and ovary with an emphasis on risk factors. METHODS: Between 1989 and 2002, 84 patients with synchronous primary cancers of the endometrium and ovary were identified. Patients with uterine papillary serous carcinoma were excluded. Clinical and pathologic information was obtained from medical records. Parametric methods were used to compare clinical and pathologic features. Kaplan-Meier survival analyses were performed and compared using the log rank test. RESULTS: Median age at diagnosis was 50 years. Median body mass index (BMI) was 28 kg/m(2). Fifty-one percent (43/84) of the women were premenopausal and 33% (28/84) were nulliparous. The most common presenting symptom was abnormal vaginal bleeding; in those women with abnormal vaginal bleeding, 69% had stage I ovarian cancer. Ovarian cancer was an incidental finding in 48% of these patients. Sixty-eight percent of patients (57/84) had endometrioid histology of both their endometrial and ovarian cancers. Patients with early stage ovarian cancer tended to have a more favorable prognosis than those with advanced stage disease (median survival not reached in stage I and II versus 66 months in stage III and IV, P = 0.06). Patients with concordant endometrioid histology had a favorable prognosis (median survival 119 versus 48 months in all other groups, P = 0.02). CONCLUSIONS: In this large series of patients, women with synchronous primary cancers of the endometrium and ovary were young, obese, nulliparous, and premenopausal. Patients with concordant endometrioid tumors of the endometrium and ovary had a favorable prognosis, with median survival approaching 10 years. PMID- 15297189 TI - Identification of a novel mechanism of NF-kappaB inactivation by progesterone through progesterone receptors in Hec50co poorly differentiated endometrial cancer cells: induction of A20 and ABIN-2. AB - OBJECTIVE: Nuclear factor kappa B (NFkappaB) is a strong anti-apoptotic factor, which is constitutively active in human endometrial cancer cells. Progesterone is the principal growth inhibitory hormone in the endometrial epithelium and promotes apoptosis. To identify the pathways through which progesterone controls NFkappaB function, we explored its genomic and non-genomic effects in endometrial cancer cells. METHODS: PR-negative Hec50co endometrial cancer cells were engineered to express high levels of the A or B isoform of PR (PRA or PRB) by adenoviral infection. Cells were treated with progesterone or vehicle alone, and RNA was isolated. Affymetrix microarrays were performed and transcriptional control of the genes of highest interest was confirmed by semi-quantitative RT PCR. To assess the non-genomic effects of PR on inflammation associated with NF kappaB, electromobility shift assays (EMSAs) were performed. RESULTS: Expression analysis demonstrated a significant effect of progesterone after 12- and 24-h treatment on several genes; in particular, A20 and ABIN-2 were induced through PRB. These factors bind in a complex and inhibit NFkappaB transcriptional activity. In addition, EMSAs revealed the complete inhibition of NFkappaB dimer binding to DNA by both PRA and PRB. CONCLUSIONS: Progesterone is the principal differentiating hormone in the endometrium. We have now identified several down stream pathways of action, one of which is the control of genes involved in NFkappaB activity. The tumorigenic inflammatory and anti-apoptotic effects of NFkappaB are inhibited by progesterone/PRB through the transcriptional control of binding proteins A20 and ABIN-2. This pathway offers interesting targets for future therapeutic development. PMID- 15297190 TI - Phase II trial of 3-h infusion of paclitaxel in patients with adenocarcinoma of endometrium: Japanese Multicenter Study Group. AB - OBJECTIVE: We assessed the antineoplastic effect and adverse reactions of paclitaxel monotherapy with paclitaxel 210 mg/m(2) given every 3 weeks by 3-h infusion on patients with endometrial cancer given as a 3-h infusion. METHODS: This study was a multi-center, open-label phase II clinical trial of paclitaxel 210 mg/m(2) given every 3 weeks by 3-h infusion. Patients with advanced or recurrent endometrial cancer were enrolled. The primary endpoint for efficacy was tumor response rate. The secondary endpoints were duration of response and adverse drug reactions. RESULTS: Among 23 patients evaluated for efficacy, partial remission (PR) was achieved in 7, no change (NC) in 10, progressive disease (PD) in 5, and not estimable (NE) in 1. The overall response rate was 30.4% (7/23 cases). In seven PR cases, median duration of response was 130 days (100-245 days). Subjective or objective symptoms > or =grade 3 included febrile neutropenia and constipation in 8.7% (2/23 cases) each; and nausea, vomiting, fatigue, pain, urinary tract infection, lowered oxygen saturation, anorexia, arthralgia, myalgia, neuropathy, weight loss, dyspnea, and need for red cell transfusion in 4.3% (1/23) each. Laboratory test abnormalities > or =grade 3 included neutropenia (78.3%, 18/23), leucopenia (47.8%, 11/23), lowered hemoglobin (13.0%, 3/23), decreased potassium (8.7%, 2/23), and decreased sodium (4.3%, 1/23). All adverse reactions were successfully managed by prolonging treatment interval, dose reduction, interrupting administration, discontinuation, and administration of G-CSF. CONCLUSION: Three-hour intravenous infusion of paclitaxel 210 mg/m(2) is useful for endometrial cancer. Antineoplastic effect was achieved and adverse reactions were clinically manageable. PMID- 15297191 TI - Survival benefit of metastasectomy for Krukenberg tumors from gastric cancer. AB - OBJECTIVE: An optimal treatment strategy for ovarian metastases of gastric cancer has not been clearly established. The aim of this study was to examine the role of a metastasectomy in the management of metachronous Krukenberg tumors after curative surgery for gastric cancer. METHODS: Among 1235 female patients who had undergone a curative gastric resection for stomach cancer between 1987 and 1998, 54 (4.4%) developed Krukenberg tumors as a first recurrence without evidence of a distant metastasis. Of these 54 patients, 33 underwent a metastasectomy while 21 did not. The survival duration between the two groups was analyzed and compared. RESULTS: The clinicopathological features of Krukenberg tumors as well as those of the primary cancers in the two groups were similar. All 33 patients in the resection group underwent subsequent adjuvant chemotherapy, including the 7 who received intraperitoneal chemotherapy. The 21 patients in the non-resection group were managed by either systemic chemotherapy (n = 16) or supportive care (n = 5) alone. The median survival duration of all the patients was 9 months (95% confidence interval, 3-15 months). The median survival time in the resection group was 17 months (95% confidence interval, 10-24 months), which was significantly longer than that in the non-resection group, 3 months (95% confidence interval, 2-4 months) (P < 0.001). CONCLUSION: Our results suggest that a metastasectomy was associated with an improved survival in patients with metachronous Krukenberg tumors from gastric cancer. These data offer a strong argument in favor of performing metastasectomy for Krukenberg tumors in the absence of an obvious distant metastasis. PMID- 15297192 TI - Cisplatin and irinotecan in squamous cell carcinoma of the cervix: a phase II study of the Gynecologic Oncology Group. AB - OBJECTIVE: To evaluate the combination of cisplatin and irinotecan as first-line treatment of patients with advanced, persistent, or recurrent squamous cell carcinoma of the cervix. METHODS: Patients with no prior treatment for metastatic disease, presence of measurable tumors, performance status of 0 or 1, and adequate bone marrow, renal, and hepatic functions were potentially eligible for this trial. Cisplatin and irinotecan were given weekly at starting doses of 25 and 65 mg/m(2), respectively, for three consecutive weeks. Cycles were to be repeated every 28 days with dose adjustments as required. Patient accrual was based on a two-stage design with at least seven responses out of 28 patients in the first stage required to proceed to a second stage of accrual seeking a response rate of 40% or better. RESULTS: Of 34 patients entered onto the study, 31 were eligible and 27 were evaluable for response. Ten had received prior chemoradiation containing cisplatin. Among the five (two complete and three partial) observed responses, two were in the subset of patients who had received prior chemoradiation. This level of activity was deemed insufficient to warrant a second stage of accrual. Predominant toxicities were myelosuppression and gastrointestinal symptoms, although six patients experienced none of these adverse effects. CONCLUSION: At these doses, weekly cisplatin and irinotecan failed to demonstrate sufficient activity to undertake a phase III study. Although not apparent in this study, prior chemoradiation may affect response to platinum-based combinations and its impact should be considered in the design of future trials. PMID- 15297193 TI - Human papillomavirus type 16 E6, E7, and L1 variants in cervical cancer in Indonesia, Suriname, and The Netherlands. AB - OBJECTIVE: Human papillomavirus type 16 (HPV 16) has several intratypic variants, and some are associated with enhanced oncogenic potential. For risk determination as well as for future vaccine development, knowledge about variants is important. Regarding the geographical distribution of HPV variants and the lack of data from Indonesia and Suriname, we studied the prevalence of HPV 16 variants in cervical cancer in these high incidence countries. Data were compared with The Netherlands, a low-risk country. METHODS: DNA samples from 74 formalin-fixed paraffin-embedded HPV 16-positive cervical carcinomas from Indonesia (Java, N = 22), Suriname (N = 25), and The Netherlands (N = 27) were amplified using primers specific for the E6, E7, and part of the L1 regions. Products were sequenced and analyzed. RESULTS: A specific Javanese variant, with mutations 666A in E7 and 6826T in L1, was found in 73% of the Indonesian samples, 56% having an additional mutation in the E6 open reading frame (ORF; 276G), giving the predicted amino acid change N58S. This Javanese variant was also found in three Surinamese samples, which reflects what could be expected from migration of Javanese people to Surinam. Other non-European variants were identified in Indonesian, Surinamese, and Dutch samples in 14%, 28%, and 19%, respectively. CONCLUSION: The majority of the HPV 16-positive cervical cancers in Indonesia are caused by a specific intratypic variant that was rarely found before in other countries. PMID- 15297194 TI - Association of hemoglobin level with survival in cervical carcinoma patients treated with concurrent cisplatin and radiotherapy: a Gynecologic Oncology Group Study. AB - OBJECTIVES: To determine if there is an association of hemoglobin level before or during concurrent cisplatin and radiotherapy (RT) with disease outcome in women with locally advanced cervical cancer, and to assess if the association is particularly significant at a specific interval or time during treatment. METHODS: A retrospective review of 494 patients treated on two consecutive prospective Gynecologic Oncology Group (GOG) trials was conducted. Demographic data, pathologic information, treatment-related factors, and hemoglobin values at baseline and during each week of therapy were collected. Cox proportional hazards model was performed to evaluate the impact of hemoglobin level on progression free survival (PFS). RESULTS: Of the combined patients, 278 (56%) and 216 (44%) were diagnosed with Stage II and Stage III/IV disease, respectively. Controlling for age, race, performance status, disease stage, tumor size, cell type, and duration of radiotherapy, mean hemoglobin values during treatment were predictive of disease progression (P < 0.0001). The pretreatment level was not significant when hemoglobin levels during treatment were included in the multivariate analysis. When the 6-week treatment course was divided into 2-week periods (early, middle, and late), analysis revealed hemoglobin values during the late period were the most predictive of disease progression (P = 0.0289). CONCLUSIONS: Hemoglobin levels during combined radiotherapy and cisplatin were independent predictors of treatment outcome in advanced cervical carcinoma. The pretreatment level was not a significant predictor of outcome when hemoglobin levels during treatment were included in the multivariate regression model. Levels in the last part of treatment were the most predictive of disease recurrence and survival. PMID- 15297195 TI - Phase II trial of Oxaliplatin and 5-Fluorouracil/Leucovorin combination in epithelial ovarian carcinoma relapsing within 2 years of platinum-based therapy. AB - OBJECTIVE: To evaluate the efficacy and toxicity of Oxaliplatin and 5 Fluorouracil (5-FU)/Leucovorin (LV) combination in ovarian cancer relapsing within 2 years of prior platinum-based chemotherapy in a phase II trial. METHODS: Eligible patients had at least one prior platinum-based chemotherapy regimen, elevated CA-125 > or = 60 IU/l, radiological evidence of disease progression and adequate hepatic, renal and bone marrow function. Patients with raised CA-125 levels alone as marker of disease relapse were not eligible. Oxaliplatin (85 mg/m(2)) was given on day 1, and 5-Fluorouracil (370 mg/m(2)) and Leucovorin (30 mg) was given on days 1 and 8 of a 14-day cycle. RESULTS: Twenty-seven patients were enrolled. The median age was 57 years (range 42-74 years). The median platinum-free interval (PFI) was 5 months (range 0-17 months) with only 30% of patients being platinum sensitive (PFI > 6 months). Six patients (22%) had two prior regimens of chemotherapy. A total of 191 cycles were administered (median 7; range 2-12). All patients were evaluable for toxicity. The following grade 3/4 toxicities were noted: anemia 4%; neutropenia 15%; thrombocytopenia 11%; neurotoxicity 8%; lethargy 4%; diarrhea 4%; hypokalemia 11%; hypomagnesemia 11%. Among 27 enrolled patients, 20 patients were evaluable for response by WHO criteria and 25 patients were evaluable by Rustin's CA-125 criteria. The overall response rate (RR) by WHO criteria was 30% (95% CI: 15- 52) [three complete responses (CRs) and three partial responses (PRs)]. The CA-125 response rate was 56% (95% CI: 37-73). Significantly, a 25% (95% CI: 9-53) radiological and a 50% (95% CI: 28-72) CA-125 response rate were noted in platinum resistant patients (PFI < 6 months). The median response duration was 4 months (range 3-12) and the median overall survival was 10 months. CONCLUSION: Oxaliplatin and 5 Fluorouracil/Leucovorin combination has a good safety profile and is active in platinum-pretreated advanced epithelial ovarian cancer. PMID- 15297196 TI - Dominant human papillomavirus 16 infection in cervical neoplasia in young Japanese women; study of 881 outpatients. AB - OBJECTIVE: Human papillomavirus (HPV) infection is reported to be related to carcinogenesis in the uterine cervix. In Japan, screening for cervical cancer by cytology is performed in women over 30 years old. The purpose of this study was to determine whether there is an association between patient age and cervical neoplasia or HPV infection in Japanese women. METHODS: Specimens from 881 randomly selected patients who came to our clinic were tested for HPV DNA by using Hybrid Capture II, whereas specimens from a 204-patient randomly selected subset diagnosed with cervical neoplasia were tested for HPV DNA by using polymerase chain reaction (PCR). HPV typing was performed in all the PCR-positive cases. RESULTS: The HPV-positive rate in the 20- to 29-year-old patients (29.0% in the normal cytology/histology group and 85.5% in the abnormal group) was higher than in the 30- to 59-year-old patients, and the rate declined until age 60 when age increased. While HPV 18, HPV 52, other HPV types, and HPV types as a whole were frequently detected in 30- to 49-year-old patients, HPV 16 was detected more frequently in the younger group than the other HPV types (P = 0.03). Among the HPV 16-positive patients with cervical neoplasia, the proportion of cervical intraepithelial neoplasia (CIN) 3 cases was high (44%) in the 20- to 29-year-old group. CONCLUSIONS: Screening for cervical neoplasia by cytology should also be performed in women under 30 years old in Japan. The HPV typing could be a tool to strictly follow-up younger women who were diagnosed with CIN. PMID- 15297197 TI - Consequences of inadvertent, suboptimal primary surgery in carcinoma of the uterine cervix. AB - OBJECTIVES: Invasive cervical cancer that is discovered only after simple hysterectomy remains a problem. Little is known about the best management of this group since there are no relevant outcome studies. This study aimed to quantify the benefits of guideline-based treatment by comparing outcome data in patients treated by inappropriate simple hysterectomy and adjuvant radiotherapy with data in patients treated with primary radical surgery, radiotherapy, or radiochemotherapy. METHODS: Records of 288 patients who had undergone radical hysterectomy with pelvic lymphadenectomy or simple hysterectomy were extracted and divided into three groups-radical hysterectomy alone (n = 89), radical hysterectomy and adjuvant radiotherapy (n = 119), and simple hysterectomy with adjuvant radiotherapy (n = 80). Disease-free and overall survival were calculated using Kaplan-Meier analyses. RESULTS: There was a trend towards better overall survival in the radical hysterectomy group. Disease-free survival was significantly better in patients treated by radical hysterectomy, followed by simple hysterectomy plus radiotherapy, and then radical hysterectomy plus radiotherapy (P(log rank DFS) < 0.002). When the two radical surgery groups were combined and compared with the suboptimally treated group, no significant differences were seen for overall survival. CONCLUSION: Postoperative radiotherapy is a good treatment for patients with cervical cancer who have undergone suboptimal simple hysterectomy. Appropriate selection criteria for further surgery remain to be defined. PMID- 15297198 TI - E-cadherin and catenins in early squamous cervical carcinoma. AB - OBJECTIVES: To examine the prognostic significance of the protein expression of E cadherin, alpha-, beta-, and gamma-catenin in early squamous cervical carcinoma (SCC). METHODS: We studied 219 patients who underwent radical hysterectomy and bilateral lymphadenectomy at our institution for stage IB SCC between 1987 and 1993. Immunohistochemistry using monoclonal antibodies against E-cadherin, alpha , beta-, and gamma-catenin was used to examine protein expression. Ten patients who underwent hysterectomy for uterine prolapse served as controls. RESULTS: Membrane expression for E-cadherin, alpha-, beta-, and gamma-catenin was decreased and low expression (< or =50% positive cells) was found in 198/219 (90%), 154/219 (70%), 157/219 (72%), and 181/219 (83%) tumors, respectively, and high (>50% positive cells) in 21/219 (10%), 65/219 (30%), 62/219 (28%), and 38/219 (17%) tumors, respectively. In univariate analysis, all classical clinicopathological parameters but none of the investigated proteins were associated with prognosis. In multivariate analysis, only deep stromal invasion was independently related to survival. CONCLUSION: E-cadherin, alpha-, beta-, and gamma-catenin were not independently associated with prognosis in stage IB SCC. PMID- 15297199 TI - Sentinel lymph node detection in early cervical cancer with combination 99mTc phytate and patent blue. AB - OBJECTIVE: The purpose of this study was to examine sentinel lymph node (SLN) detection in patients with early stage cervical cancer using (99m)Tc phytate and patent blue dye and to compare our method with published findings utilizing other radioisotopic tracers. PATIENTS AND METHODS: A total of 20 consecutive patients with cervical cancer scheduled for radical hysterectomy and total pelvic lymphadenectomy at our hospital underwent SLN detection study. The day before surgery, lymphoscintigraphy was performed with injection of 99m-technetium ((99m)Tc)-labeled phytate into the uterine cervix. At surgery, patients underwent lymphatic mapping with a gamma-detecting probe and patent blue injected into the same points as the phytate solution. RESULTS: At least one positive node was detected in 18 patients (90%). A total of 46 sentinel nodes were detected (mean, 2.3; range, 1-5). Most sentinel nodes were in one of the following sites: external iliac (21 nodes), obturator (15 nodes), and parametrial (7 nodes). Eleven (24%) sentinel nodes were detected only through radioactivity and two (4%) were detected only with blue dye. The sensitivity, specificity, and negative predictive value for SLN detection were all 100%. Nine published studies involving 295 patients had a summarized detection rate of 85%. Summarized sensitivity, specificity, and negative predictive value were 93%, 100%, and 99%, respectively. CONCLUSION: Combination of (99m)Tc phytate and patent blue is effective in SLN detection in early stage cervical cancer. PMID- 15297200 TI - Phase II study of sequential doublets: topotecan and carboplatin, followed by paclitaxel and carboplatin, in patients with newly diagnosed advanced ovarian cancer. AB - OBJECTIVE: Incorporating topotecan into standard platinum/taxane chemotherapy for advanced ovarian cancer has been complicated by myelosuppression. This study evaluated sequential doublets of topotecan and carboplatin, followed by paclitaxel and carboplatin, in newly diagnosed advanced ovarian cancer patients. METHODS: Forty-five patients (median age, 56 years; range, 38-77 years) with stage III/IV disease and GOG performance status <2 were enrolled and received four cycles of topotecan (1.0 mg/m(2)/day on days 1 to 3) and carboplatin (AUC 4 on day 1), followed by four cycles of paclitaxel (175 mg/m(2) via 3-h IV infusion on day 1) and carboplatin (AUC 5 on day 1). All cycles were 21 days. Antitumor response was assessed after four and eight cycles; patients with clinical complete response (CR) underwent second-look laparotomy for determination of pathologic CR (PCR). Dose reductions were instituted for grade 4 neutropenia and thrombocytopenia, and for grade 3/4 nonhematologic toxicity. RESULTS: Among 41 CA 125 evaluable patients, complete and partial responses were observed in 29 (70.7%) and 11 (26.8%) patients, respectively. Of the 12 clinical CRs (43%) in 28 evaluable patients, 10 patients underwent second-look laparotomy, with 3 PCRs (30%). Median time to progression was 14 months and actuarial survival was 23 months. Neutropenia was the primary toxicity and cause of dose adjustments and delays, including two deaths. CONCLUSION: The antitumor activity observed is comparable with other series, although neutropenic complications were increased. Progression-free and actuarial survivals were slightly inferior. A Phase III trial (GOG 182) of sequential doublets in the reverse sequence is ongoing. PMID- 15297201 TI - Effectiveness of platinum-based chemotherapy among elderly patients with advanced ovarian cancer. AB - BACKGROUND: Platinum-based chemotherapy is the standard of care for women with advanced ovarian cancer based on the results of randomized trials. We previously showed that only about half of women over the age of 65 years with this disease received platinum-based chemotherapy, and that the likelihood of receiving it decreases with age. METHODS: We used the Surveillance, Epidemiology, and End Results (SEER)-Medicare database to identify women diagnosed from 1/1/92 to 12/31/96 with stage III or IV ovarian cancer who survived > or =120 days beyond diagnosis, and were > or =65 years of age. Cox proportional hazards models and propensity scores were used to control for known predictors of receiving treatment and to estimate the relative effectiveness of different platinum-based regimens. RESULTS: Of the 1759 patients in the sample who met our eligibility criteria, 53% received platinum-based therapy. For this sample, the Cox proportional hazard ratio was 0.72 (95% CI, 0.62-0.91) for mortality associated with the use of any platinum-based therapy, and 0.59 (95% CI, 0.45-0.76) for combination platinum/paclitaxel therapy. Similar results were obtained using propensity score modeling. CONCLUSIONS: In this population-based study, we found that only about half of women with advanced ovarian cancer over age 65 were treated with platinum-based chemotherapy; however, survival improved by 38% in treated women, similar to the benefits described in randomized controlled trials among younger patients, and were greatest when platinum was combined with paclitaxel. An effort to increase the utilization of platinum combination therapy among older patients with advanced ovarian cancer is justified. PMID- 15297202 TI - Defective mismatch repair and the development of recurrent endometrial carcinoma. AB - OBJECTIVE: To investigate whether defective DNA mismatch repair (MMR) defines a subgroup at risk for recurrence in sporadic endometrial carcinoma patients. METHODS: Primary tumors from 44 patients with recurrent stage I endometrial carcinoma were compared after matching, with tumors of 44 patients being free of recurrence for minimal 3 years. Paraffin-embedded primary tumors (n = 88) and recurrent tumors (n = 32) were subjected to immunohistochemical analysis for hMSH2 and hMLH1 expression. Subsequently, a staining index (SI = 0-9) was calculated based on staining intensity and quantity. DNA was extracted from paraffin-embedded tissues, and promoter methylation of hMLH1 was determined by nested methylation-specific PCR (MSP). Microsatellite instability (MSI) was assessed by BAT-26 or BAT-25. RESULTS: Low hMSH2 expression was observed in 2% of primary tumors of control patients without recurrence, 14% of primary tumors of patients with recurrence, and 0% of recurrent tumors. Low hMLH1 expression was observed in 32%, 19%, and 22%, respectively. hMLH1 gene promoter methylation was detected in 50%, 47%, and 32%, and MSI was found in 16%, 14%, and 30%, respectively. No significant differences were found between primary tumors of patients with and without recurrence with respect to hMSH2 and hMLH1 expression, hMLH1 promoter methylation, and MSI. When primary and recurrent tumors were compared, there was an increased correlation of hMLH1 methylation with low hMLH1 expression and MSI in recurrent tumors. CONCLUSION: MSI, hMLH1 promoter methylation, and the expression of hMLH1 and hMSH2 are not predictive for the development of recurrent stage I endometrial carcinoma. In the progression of tumor, "de novo" hMLH1 methylation rarely occurs, instead there is further derailment of the MMR pathway in affected tumors. PMID- 15297203 TI - The overlay autogenous tissue (OAT) patch to control major intraoperative vascular injury in an ovine model. AB - OBJECTIVE: Iatrogenic surgical injury to large blood vessels in the abdomen or pelvis is a rare adverse event but may be lethal. We present a new technique to repair serious vascular injury using a free graft of omentum or fascia as an overlay autogenous tissue (OAT) patch. METHODS: Repair to venous and arterial defects was made using an OAT patch of omentum or rectus abdominis sheath sutured through the patch and the adventitia of the injured vessel at four sites. The technique was tested in 33 repairs in an experimental sheep model. RESULTS: Thirty-three patch repairs were attempted and all were successfully completed. Twenty-one (64%) patches were controlled by the patch alone, and adjuvant local pressure was required in 12 (36%) cases. CONCLUSION: We have demonstrated that an OAT patch can be used to control vascular injury in the experimental sheep model. This technique may be useful to all surgeons especially those operating in the pelvis where there may be restricted access to repair vascular injury. PMID- 15297204 TI - The control of severe intraoperative bleeding using an overlay autogenous tissue (OAT) patch: case reports. AB - BACKGROUND: Severe intraoperative bleeding cannot always be controlled by standard surgical techniques. We recently reported a new technique to repair serious vascular injury using a free graft of omentum or rectus abdominus fascia as an overlay autogenous tissue (OAT) patch in the experimental sheep model. We now describe the successful clinical use of this patch in three patients. CASE REPORTS: Radical surgery was performed on three patients with pelvic malignancy with resulting uncontrollable bleeding from the internal iliac vein, pelvic side wall and paravaginal venous plexuses, respectively. Hemostasis was secured using an OAT patch made of abdominus rectus fascia in two cases and appendix epiploicae as an omental substitute in the other. DISCUSSION: The utility of the OAT patch is described in three different clinical situations. It is suggested that this technique may be especially useful to gynecologic oncologists when standard surgical techniques fail to control bleeding or there is limited access to the site of injury. PMID- 15297205 TI - Recurrence of prolactin-producing endometrial stromal sarcoma with sex-cord stromal component treated with progestin and aromatase inhibitor. AB - INTRODUCTION: Endometrial stromal tumors with sex-cord-like elements are relatively rare. We report a case of this neoplasm with prolactin as a tumor marker for recurrent disease. We also report response of recurrent disease to progesterone and aromatase inhibitor. CASE REPORT: A 48-year-old woman was diagnosed with Stage I endometrial stroma sarcoma with sex-cord component at the time of hysterectomy for presumed fibroid uterus. One and a half years later, she presented with recurrent disease in the abdomen associated with breast tenderness, galactorrhea, and an elevated prolactin level. She received three cycles of BEP (Bleomycin, Etoposide, Cisplatin) with partial response and followed by an optimal debulking procedure. Two out of a six additional planned cycles of BEP were administered with complete tumor response and normalized prolactin level. Second recurrence, 9 months later, again presented with galactorrhea and rising prolactin. Disease was progressive through three cycles of Docetaxel and Gemcitabine therapy, but had an objective response to treatment with anastrozole and megestrol acetate. Prolactin level normalized. Two years later there is stable disease and the patient remains symptom-free. DISCUSSION: Endometrial stromal sarcoma with sex-cord stromal component may be hormonally functional. Similarly to pure endometrial stromal sarcomas, they may respond to hormonal treatment, and further study is warranted. PMID- 15297206 TI - Lobular carcinoma in situ of the breast within a fibroadenoma. A case report. AB - BACKGROUND: Fibroadenoma is a benign breast tumor that is effectively treated by local excision. Although uncommon, breast fibroadenoma may harbor breast carcinoma, either in situ or invasive. CASE: We present a 27-year-old woman who presented for the management of an apparently benign mass in the right breast. The clinician and ultrasonographic characters of the mass were indicative of a breast fibroadenoma. A local excision of the mass was performed. Histology showed the presence of LCIS within the fibroadenoma. After a detailed discussion with the patient about the available management options, the conservative approach (which includes local excision/biopsy only and regular follow-up) was preferred. CONCLUSION: LCIS within a breast fibroadenoma is a rare entity. Carcinomas arising within a fibroadenoma have the same biological behavior as those arising independently and, therefore, their management should be the same. Close surveillance following local excision/biopsy is the preferred management for LCIS within a fibroadenoma. PMID- 15297207 TI - Pregnancy after trachelectomy: a high-risk condition of preterm delivery. Report of a case and review of the literature. AB - BACKGROUND: Trachelectomy is a conservative but locally radical procedure associated with a high risk of preterm delivery. CASE: A 28-year-old patient with cervical cancer FIGO stage IB1 was treated with laparoscopic pelvic lymphadenectomy followed by trachelectomy. Three years later, she conceived spontaneously. In consideration of the high risk of preterm delivery, the cervical status was evaluated by transvaginal ultrasonography. At 16 weeks' gestation, we observed the cerclage suture correctly placed at the level of the internal cervical os and a "neo-cervical" segment length of 1.5 cm. Thereafter, serial ultrasound measurements showed preservation of the cervical competence. The patient achieved an uneventful pregnancy and delivered by elective cesarean section at 37 weeks. CONCLUSION: Transvaginal scans to evaluate the competence of the "neo-cervix" may contribute to the management and counseling of patients after trachelectomy. PMID- 15297208 TI - Grade 3 liposomal-doxorubicin-induced skin toxicity in a patient following complete resolution of moderately severe sunburn. AB - PURPOSE: Palmar-plantar erythrodysesthesia (PPE) is a potentially serious toxicity of a number of cytotoxic chemotherapeutic agents, including liposomal doxorubicin. Activities that may increase the risk of this toxicity should be avoided. CASE REPORT: A patient with platinum-resistant ovarian cancer, responded to, and tolerated (including no skin rash) an initial cycle of liposomal doxorubicin (40 mg/m(2)). Unfortunately, several days before her next scheduled cycle, she developed significant sunburn (intense erythema without blistering). Despite an additional 1-week delay (total of 5 weeks from the prior liposomal doxorubicin), and complete visual recovery from the effects of the sunburn, the patient developed severe (grade 3) PPE involving both hands (pain, pronounced erythema, blistering, without ulceration), and a slightly less extensive reaction of both feet, following subsequent treatment with a 25% reduced dose of the agent (30 mg/m(2)). CONCLUSION: Caution is advised when considering the administration of liposomal doxorubicin following a major sunburn, despite total resolution of the visible effects on the skin. A delay of several weeks may be appropriate to avoid exacerbation by the chemotherapeutic agent of persistent subclinical damage to normal epithelial cells. PMID- 15297209 TI - The use of taxanes in choriocarcinoma; a case report and review of the literature. AB - BACKGROUND: Both preclinical data and case reports support the use of taxanes for high-risk metastatic choriocarcinoma. CASE: We report the case of a 31-year-old with metastatic choriocarcinoma who required 3rd-line treatment with a paclitaxel cisplatin-based regimen. She achieved a complete response and remains relapse free 21 months after her last dose of chemotherapy. CONCLUSION: The literature suggests that paclitaxel contributes significantly to the treatment of choriocarcinoma and its use should be explored further. This is the first case report formally reporting its combination with cisplatin. PMID- 15297210 TI - Hematogenous rectal metastasis 20 years after removal of epithelial ovarian cancer. AB - BACKGROUND: Rectal tumors are rarely metastatic. Moreover, hematogenous spread is rare in ovarian cancer whose dissemination frequently occurs through peritoneal or lymphatic ways. CASE: A 55-year-old female presented with a rectal metastasis that appeared 20 years after the treatment of a primary clear cell carcinoma of the ovary. The cytokeratin 7 positive/cytokeratin 20 negative immunophenotype assessed the ovarian origin of the rectal tumor. Because of the integrity of the rectal serosa and the uninvolved mesenteric lymph nodes, we hypothesize our rectal metastasis to come from the hematogenous way. CONCLUSION: This is the first reported case of hematogenous rectal metastasis in epithelial ovarian carcinoma and that, with so late delay. PMID- 15297211 TI - Cecal pelvic transposition following total pelvic exenteration. AB - BACKGROUND: Multiple techniques have been utilized in an attempt to prevent small bowel obstructions following total pelvic exenteration. Pelvic transposition of the cecum may be an effective way to reduce the incidence of this serious complication. CASES: We present three women who underwent total pelvic exenteration and cecal pelvic transposition to exclude the small bowel from the "empty pelvis". All three patients did well without any postoperative small bowel complications. CONCLUSION: Cecal pelvic transposition precludes the small bowel from entering the pelvis. This procedure can be performed relatively easily by a gynecologic oncologist experienced with bowel surgery. A larger patient series is needed to assess the efficacy of this previously unpublished procedure. PMID- 15297212 TI - Angiomyolipoma of the uterus associated with tuberous sclerosis: case report and review of the literature. AB - BACKGROUND: Angiomyolipoma (AML) is a rare benign tumor that occurs most commonly in the kidney. Only a few cases have been described in the uterus. CASE: We describe a 32-year-old woman with tuberous sclerosis who presented with irregular bleeding. Preoperatively, the lesions were confused with leiomyoma on ultrasound. At laparotomy, multiple circumscribed subserosal and intramural purplish lesions on the corpus uteri were noted leading to a diagnosis of uterine angiomyolipoma with the histological and immunohistochemical findings. CONCLUSION: In contrast to other reported uterine angiomyolipoma cases, HMB-45 melanoma-specific antibody immunoreactivity is demonstrated in the present case. The histopathologic diagnosis as well as immunohistochemical analysis is discussed here with a review of the literature. PMID- 15297213 TI - Gastrointestinal adenocarcinoma arising in a mature cystic teratoma of the ovary. AB - BACKGROUND: Mature cystic teratoma of the ovary (MCTO) is the most common ovarian germ cell neoplasm and is usually diagnosed in early adulthood. Malignant transformation is rare, occurring in approximately 2% of all cases. Though malignant transformation can occur from any of the embryonic germ layers, the most common malignancy arising in these otherwise benign tumors is squamous cell carcinoma. CASE: We present a patient with a MCTO where malignant transformation of gastrointestinal epithelium resulted in moderately differentiated adenocarcinoma. After 3 years of follow-up, she remains free of disease. CONCLUSION: Although gastrointestinal epithelium is often found in MCTOs, adenocarcinoma arising from this cell type is uncommon. This is the third reported case of adenocarcinoma arising in gastrointestinal epithelium of a MCTO. PMID- 15297214 TI - Preservation of pregnancy in a patient with a stage IIIB ovarian epithelial carcinoma diagnosed at 22 weeks of gestation and treated with initial chemotherapy: case report and literature review. AB - BACKGROUND: To report a case of successful management of a FIGO stage III endometrioid carcinoma of the ovary diagnosed during pregnancy at 22 weeks of gestation and treated with initial chemotherapy while preserving the pregnancy. CASE: The patient underwent a planned cesarean section at 34 weeks after two courses of carboplatin. She delivered a healthy baby. At the same time, a radical hysterectomy, omentectomy, pelvic and paraaortic lymphadenectomies and peritonectomies were carried out. The surgical resection was complete (no macroscopic residual disease). During histologic examination, traces of persistent disease were found. The patient underwent seven postoperative courses of chemotherapy (carboplatin + paclitaxel regimen) after radical surgery. After 18 months of follow-up posttreatment, the patient remains in complete remission and the child's development is normal. CONCLUSION: Chemotherapy during pregnancy with preservation of the fetus could be considered and should be discussed in case of epithelial ovarian cancer (EOC) diagnosed during the second trimester of the pregnancy. PMID- 15297215 TI - Commenting on centralizing surgery for gynecologic oncology: a strategy assuring better quality treatment? (89:4-8) by Karsten Munstedt, et al. PMID- 15297217 TI - Early ovarian cancer--time for a rethink on stage? PMID- 15297219 TI - [Laparoscopic coloproctectomy with ileoanal anastomosis: a foreseeable victory?]. PMID- 15297220 TI - [Preoperative screening and nutritional support of nutritional deficiencies]. AB - Nutritional deficiencies have to be considered as an independent risk factor for postoperative morbidity. Peri-operative nutritional support reduce this risk in elective abdominal surgery for cancer and cardiac surgery. Preoperative nutritional support for 7-10 days reduce postoperative complications in undernourished patients by 10% but is not operant when administered after surgery. Enteral route is as effective. Recent studies using immunonutrients conclude that a short preoperative oral intake is able to reduce complications even in well-nourished patients. Then, a preoperative nutritional screening must be routinely performed leading to a nutritional programme. PMID- 15297221 TI - [Two stage videoassisted restorative proctocolectomy. Early experience of 12 cases]. AB - AIM OF THE STUDY: This study reports our early experience in two-stage video assisted restorative proctocolectomy (RPC). PATIENTS AND METHODS: From May 1999 to May 2003, 12 video assisted RPCs were performed (mucosal ulcerative colitis: n = 11; familial adenomatous polyposis: n = 1). These patients were matched for age, gender, body mass index and indication for surgery, with 12 patients who underwent RPC by laparotomy (open group). RESULTS: Median operative time was significantly longer in the video assisted RPC group (400 min; range: 360-490) vs open group (300 min; range: 210-390) (P = 0.003). A conversion in midline laparotomy (under the umbilicus) was necessary in 3/12 patients (25%) in the video assisted RPC group. Return to bowel function and oral intake occurred two days earlier after video assisted RPC (respectively, P = 0.009 and P = 0.0001) but length of stay was not significantly shorter in this group. A complication occurred in 3/12 patients (25%) in both groups, which lead to a reoperation in one patient in the open group (ns). CONCLUSION: Two-stage videoassisted RPC is feasible at the cost of a lengthening of operative time, Nevertheless postoperative results after video assisted RPC are comparable to those obtained after RPC by laparotomy. PMID- 15297222 TI - [Long-term followup of patients with alveolar echinococcosis]. AB - INTRODUCTION: Alveolar Echinicoccosis is a severe parasitic disease: its natural evolution is comparable to a slowly progressive malignant liver tumour. There is no definitive medical therapy. Surgery remains the only option to assure a cure. This report is our surgical experience for the care of this affection. PATIENTS AND METHOD: Between 1980 and 2002, 12 patients were operated for an hepatic alveolar echinococcosis. For 11 of them, the affection was primary. Lesions were located in the right liver lobe (n = 9), in the left lobe (n = 1), in both lobes (n = 1) and close to the hilar region (n = 1). Twice there was a diaphragmatic infiltration and once a pleural infiltration. Resections consisted in: segmentectomies in the right lobe (n = 4), right hepatectomy (n = 5, associated once with a partial I and IV segmentectomy), left hepatectomy (n = 1), one liver transplantation (n = 1), one drainage of a parasitic cavity (n = 1). One pleurectomy and a partial diaphragmatic resection were made. All patients were treated postoperatively with benzimidazole chemotherapy. RESULTS: Three patients presented some complications: segmental necrosis with biloma (n = 1), biliary fistula tract (n = 1), subdiaphragmatic hematoma (n = 1), cholangitis (n = 1). 10 patients are alive (median follow-up of 10 years). Two patients still present some parasitic lesions; the situation remains doubtful for one of them. Two patients died (one of them in the context of a disease progression). CONCLUSION: Surgical treatment, associated with medical therapy, assured a control of the parasitic lesions or a definitive cure in most cases. When the disease is limited to the liver with no possibility for partial hepatectomy, a transplantation is an option. PMID- 15297223 TI - [Trevira: a new polyester implant for the treatment of incisional hernia. Results of an experimental study]. AB - OBJECTIVE: The synthetic implant meshes in clinical use for the treatment of abdominal hernias are generally made of polyester in France and of polypropylene in Germany. Serving as an implant material for the replacement of the cruciate ligament, the Trevira is another polymer of polyester in clinical use with excellent results. This animal trial was performed to ascertain whether it offers any advantages over polypropylene for abdominal incisional hernia repair. MATERIAL AND METHODS: [corrected] In 12 pigs 10 x 10 cm of the abdominal wall preserving the peritoneum was resected and subsequently implanted a 15 x 15 cm synthetic mesh of polyethylene terephthalate (Trevira) in half of them and of polypropylene (Prolene) in the other half using a sublay technique. After two and six month the implant size was measured and the extend of the foreign body reaction determined by the microscopically presence of foreign body giant cells. RESULTS: No significant differences concerning the implant size were shown between the two groups at any of the time periods. The acute inflammatory reaction observed was significantly higher at the polypropylene than at the polyethylene terephthalate implant (number of giant cells after 2 month: Prolene 2.2 +/-0.4, Trevira: 0.8 +/-0.2, after six month: Prolene: 4.6 +/-1.3, Trevira: 1.1 +/-0.5). In contrast to the polyethylene terephthalate all polypropylene samples showed calcification areas after six month. CONCLUSION: In this animal trial Trevira mesh showed a high biocompatibility with a low foreign body reaction. It appears to be a promising new implant for the treatment of hernia. PMID- 15297224 TI - [Treatment of pancreatic pseudocysts by laparoscopic cystogastrostomy]. AB - AIM: To evaluate the clinical results of laparoscopic cystogastrostomy and to determine the potential advantages of this new therapeutic option. PATIENTS AND METHODS: This study concerned 12 patients presenting with pancreatic pseudocyst and operated on by laparoscopic cystogastrostomy between 1997 and 2002. There were five men and seven women with a median age of 46 years (range: 30-72). In ten patients, the pseudocyst developed after acute pancreatitis and the median delay between the acute onset and surgery was 7 months (range: 2-24). In two patients, the pseudocyst was associated with chronic pancreatitis. All the patients had a single cyst bulging into the posterior wall of the stomach and the median cyst diameter was 9 cm (range: 5-14). RESULTS: Endoluminal gastric laparoscopy was used in six patients and intraperitoneal transgastric laparoscopy in six patients. Conversion to open surgery was required in one patient because the cyst could not be correctly localised by laparoscopy. The median size of the cystogastrostomy was 3 cm (range: 2-5). In eight patients, necrotic debris were still present within the cyst. The median operative time was 90 min (range: 60 140) and the median postoperative hospital stay was 6 days (range: 4-24). No mortality was recorded and postoperative morbidity was limited to one haematoma of the rectus sheath on a port site. One patient was readmitted on the 20th postoperative day because of cyst infection due to partial closure of the cystogastrostomy and was treated by endoscopic placement of a stent. One patient was lost for follow-up 2 months after surgery. With a median clinical and radiological follow-up of 12 months (range: 6-36), no recurrence of pancreatic pseudocyst was observed. CONCLUSIONS: In this series, laparoscopic cystogastrostomy is associated with a low postoperative morbidity and an effective permanent result. Laparoscopy has two main advantages: an excellent control of haemostasis and the creation of a wide communication with debridement of the cyst contents thus minimizing the risk of infection or recurrence of the pseudocyst. PMID- 15297225 TI - [Colostomy vs self-expanding metallic stents: comparison of the two techniques in acute tumoral left colonic obstruction]. AB - Self-expanding metallic stents is an alternative treatment to colostomy that is the treatment of choice in acute tumoral left colonic obstruction. AIM OF THE STUDY: To compare morbidity, mortality, length of hospital stay and treatment performed after desobstruction using the two methods. PATIENTS AND METHODS: Thirty-three patients admitted for acute obstruction of the left colon were retrospectively separated in two groups depending on the type of intervention performed to treat the obstruction ("colostomy" group: 17 patients and "self expanding stent group": 16 patients). We studied complications after desobstruction, hospital courses and surgical strategy performed after the acute phase. RESULTS: Time between desobstruction and colectomy was shorter in the "self-expanding stent group" than in the "colostomy group" (18.5 days versus 73 days). Age superior than 75 years and colostomy were the two main factors predicting the risk of definitive colostomy (P < 0.05). Global mean hospital stay was longer in the colostomy group (32.7 days versus 19.3 days, P = 0.02). Two perforations and one local recurrence occurred in the "self-expanding stent group". CONCLUSIONS: Self-expanding metallic stent can decrease the permanent colostomy rate and the number of interventions. The recurrence rate seems to be theoretically increased with the stenting method. Then, colostomy must be done for patients in curative situation. The self-expanding metallic stent should be used as a palliative care. PMID- 15297226 TI - [Differentiated thyroid carcinoma in children and adolescents: therapeutic strategy according to clinic presentation]. AB - AIM OF THE STUDY: The aim of this retrospective study was to propose a therapeutic strategy according to clinic and pathologic presentations, in differentiated thyroid carcinoma in patients less than 20 years of age. PATIENTS AND METHODS: From 1995 to 2002, 74 patients less than 20 years of age were operated on for a differentiated thyroid carcinoma. Patients were divided in two groups according to the presence of lymph node (LN) detected before operation (19 "with LN" and 55 "without LN"). RESULTS: Surgery consisted of total thyroidectomy and lymph node dissection in the group "with LN". In the second group "without LN", total thyroidectomy was performed in 19 patients (associated with lymph node dissection in seven) and loboisthmectomy in 29 patients. Post operative radio iodine ablation was performed in 25 patients (16 "with LN" and nine "without LN"). During the follow up (median 71 months), 9/19 patients "with LN" underwent surgery for lymph node recurrence. Four patients of the "without LN" group were reoperated for recurrence in the controlateral thyroid lobe after initial loboisthmectomy. Initial presence of lymph nodes was a risk factor for reoperation. Survival without reoperation at 5 and 10 years was 58% and 38% for the "with LN" group and 96% and 91% for the "without LN" group respectively (P < 0.001). At the time of analysis, all patients were alive. Ninety percent of patients were in remission (68% of "with LN" and 98% of "without LN" patients). CONCLUSION: The risk of reoperation for lymph node recurrence during first post operative year is important in the "with LN" group. Therapeutic strategy consists of total thyroidectomy, complete lymph node dissection, radio iodine administration and l-thyroxine treatment. In "without LN" patients risk of lymph node recurrence is low. Treatment consist of total thyroidectomy, lymph node dissection and radio iodine administration are not systematic. PMID- 15297227 TI - [Which surgical technique for an uncomplicated umbilical hernia?]. PMID- 15297228 TI - [Gallbladder cancer revealed by a jaundice caused by an endobiliary tumor thrombus]. AB - Main bile duct neoplasic thrombosis is a rare cause of jaundice in case of gallbladder cancer. We report the case of 27-year-old woman in whom the endoluminal biopsy of biliary thrombus confirmed the suspected diagnosis of gallbladder cancer. An initial laparoscopic exploration found a localized peritoneal carcinomatosis. However, in this exceptional situation with an unknown prognostic, a surgical procedure has been performed including hepatectomy IV-V with biliary principal bile duct removal, hepatico-jejunal anastomosis (Roux-en Y), with complete resection of localized peritoneal carcinomatosis. Post operative course were uneventful and this patient was asymptomatic under chemotherapy with a six month follow-up. PMID- 15297229 TI - [Hydatic acute pancreatitis: a case report]. AB - Acute pancreatitis caused by hydatic cyst of the liver was rarely reported. The authors report a new case of hydatic pancreatitis characterized by visualization of hydatid membranes in the biliary tract without any biliary stone or alcoholic consumption. PMID- 15297230 TI - [Malignant tumours arising in extraovarian endometriosis: three case reports and review of the literature]. AB - In its extraovarian form, co-existence of carcinoma and endometriosis is a sufficient argument used in favour of the malignant transformation of endometric lesions. Estrogen as well as the loss of 5q chromosome heterozygosity are considerate as initiators of that type of carcinogenesis. Endometrioid histological type is the most frequent and is revealed usually by abdominal pain. The incidence of carcinoma arising in endometriosis is about 0.8% and 5-year survival rate of pelvic endometrioid form is about a 100% after surgery and radiotherapy. PMID- 15297231 TI - [Technique of surgical ampullectomy]. AB - We describe the technique of surgical ampullectomy, which consists of complete resection of the papilla of Vater, including the sphincter, the distal part of common bile duct and Wirsung duct, and the duodenal wall around the papilla. Limits of resection are assessed by frozen section, particularly on both biliary and pancreatic ducts which are sutured together and reinserted on the duodenal wall. Surgical ampullectomy, combined with frozen section, is associated with a low morbidity, and represents a valid alternative to pancreaticoduodenectomy and endoscopic ampullectomy for presumed-benign ampullomas. PMID- 15297232 TI - [Clinical value of crystalluria study]. AB - Crystalluria is a marker of urine supersaturation present in both normal and pathological conditions. Indeed, nature and characteristics of the spontaneous crystalluria are of clinical interest for detecting and following biological disorders involved in renal diseases. Method. Crystalluria examination should preferably be performed on first morning urine or fresh fasting voiding samples by polarised microscopy in a Malassez cell. Urine samples must be stored at 37 degrees C or at room temperature and examined within two hours following voiding. Results and discussion. Crystalluria should be interpreted according to various criteria: 1) chemical nature of crystals for abnormal crystals such as struvite, ammonium urate, cystine, dihydroxyadenine, xanthine or drugs; 2) crystalline phase of common chemical species as calcium oxalates, calcium phosphates and uric acids; 3) crystal morphology (calcium oxalates); 4) crystal size (calcium oxalates); 5) crystal abundance (calcium oxalates, calcium phosphates, uric acids, cystine); 6) crystal aggregation (calcium oxalates); 7) frequency of crystalluria assessed on serial first morning urine samples, a very useful tool for long-term surveillance of patients. Within calcium oxalate crystalluria, presence of whewellite is a marker of elevated oxalate concentration (urine oxalate > 0.3 mmol/L); a crystal number > 200/mm 3 is highly suggestive of heavy hyperoxaluria of genetic or absorptive origin. Predominant weddellite crystalluria is most often indicative of an excessive urine calcium concentration (> 3.8 mmol/L); a dodecahedric aspect of the crystals is a marker for heavy hypercalciuria (> 6 mmol/L) while an increased crystal size (>or= 35 microm) is indicative of simultaneous hypercalciuria and hyperoxaluria. Calculation of the global crystal volume, especially when applied to calcium oxalates or cystine, is a clinically useful tool for the monitoring of patients suffering from primary hyperoxaluria or cystinuria. Lastly, presence of crystalluria in more than 50% of serial first voided morning urine samples is in our experience the most reliable biological marker for detecting the risk of stone recurrence in lithiasic patients. Conclusion. Crystalluria examination is an essential laboratory test for detecting and following pathological conditions, which may induce renal stone disease or alter kidney function due to urine crystals. PMID- 15297233 TI - [Clinico-biological evaluation of malnutrition]. AB - Malnutrition is frequently observed at hospital, concerning 30 to 50% of hospitalised patients. Increased length of stay and cost of care has made of this problem a major economic stake. Indeed, malnutrition diagnosis and prevention has become, since 2001, one of major french government's priority. Malnutrition results from unbalanced nutritional requirement and in-take. It associates both weight, proteic and functional loss. Its diagnosis and evaluation of its gravity are first clinical and need body mass index determination. For biological diagnosis, nutritional markers have to be very sensitive to nutritional state. Nutritional profile can be made, associating two nutritional markers (albumin and transthyretin) and one inflammation protein (alpha 1-acid-glycoprotein). Various clinical, biological or clinico-biological indexes can also be calculated. Altogether, both indexes and nutrition profile are excellent tools for (1) diagnosis and classification of malnutrition state, moderate or severe, (2) prognostic and (3) evaluation of nutritional supplementation efficiency. PMID- 15297234 TI - [Molecular identification of mycobacteria and detection of antibiotic resistance]. AB - Mycobacteria are responsible for many human infections, especially species of tuberculosis complex, causative agents of tuberculosis. With nine millions new cases every year, this disease is responsible for more than two millions of deaths. Nontuberculous mycobacteria (e.g. Mycobacterium avium-intracellulare, Mycobacterium kansasii, Mycobacterium xenopi or Mycobacterium ulcerans) can cause infections too, usually in particular clinical settings. Standard diagnosis of mycobacterial infections relies on direct examination and culture. Although culture in liquid media allows the detection of mycobacterial growth at an earlier stage, isolation and phenotypic identification requires several weeks, as does antimicrobial susceptibility testing. Nowadays, molecular tools are available, allowing quicker accurate diagnosis. Detection of Mycobacterium tuberculosis complex by amplification-based tests can be performed directly from clinical samples, although most identifications are successfully after isolation. Several commercial techniques are now available but identification is limited to selected species, at a high cost. Sequencing of genomic targets (such as rrs, rpoB, gyrB, 16S-23S intergenic spacer or hsp65) allows accurate and quick identifications but requires access to a sequencer. Eventually, our better knowledge of the action mechanisms of the different drugs allows genotypic detection of most antibiotic resistances. Indeed, characterization of mutations in specific target genes (such as rpoB, katG, embB, pncA, gyrA or rrl) should be an effective tool for rapid detection of resistance, although this method has only been used so far for rifampin resistance detection. Nevertheless, this approach, limited to reference laboratories, should always be performed in conjunction with antibiogram. PMID- 15297235 TI - [Transferrine soluble receptors' contribution to the assessment of iron status in homozygous drepanocytic anemia]. AB - Sickle cell anemia does not cause martial deprivation per se, but may worsen when iron deficiency exists, notably in tropical zone where infectious diseases and malnutrition are endemic mainly during childhood. This study was aimed to assess iron deficiency prevalence among children with sickle cell disease (SCD) and to determine the best parameters for its diagnosis. In addition to classical parameters, we measured transferrine's soluble receptors which can reveal an iron deficiency, either isolated or associated to another condition since its level is not influenced by chronic anemia. Assays were carried out in 40 homozygous SCD patients, aged 3 to 18 years, having an hemoglobin level < 11 g/dL and in 30 age paired controls assumed to be healthy and having a negative Emmel test and an hemoglobin level < 11 g/dL. The results showed hyposideremia (serum iron < 60 microg/dL) in 17.5% of the patients. Ferritinemia, transferrinemia as well as total iron fixation capacity were in the normal range for the majority of SCD patients in spite of the frequency of hyposideremia and microcytic anemia (20%). Transferrine's saturation coefficient was low in 22.5% of patients, which can be due to martial deprivation or to inflammatory status. These results confirm the limitations of usual biochemical parameters in the diagnosis of iron deficiency in homozygous drepanocytosis. Soluble receptors' levels were increased in 60% of controls; that proves that iron deficiency prevalence is high in our countries. Higher levels were found in 97.5% of patients. However, receptors' levels are increased during haemolysis, thus it is difficult to ascertain the origin of the increase, but taking into account its index value can reduces misinterpretation. In addition, considering simultaneously microcytosis, hypochromia, transferrine's soluble receptor level and its index, we can speculate that martial deficiency occurs in 20% of SCD patients, a percentage close to the 17.1% obtained by other authors using only the combination of microcytosis and hypochromia. It results from this study that associating microcytosis and hypochromia could validly assess iron deficiency during drepanocytosis. PMID- 15297236 TI - [Analytic study of dot blotting for the detection of anti-Jo-1, anti-M2, anti ribosomes and anti-LKM]. AB - The Cyto-Dot 4 HM043 kit commercialised by BMD, has replaced the Cyto-Dot HM010 kit that allowed three auto-antibodies detection (anti-Jo-1, anti-M2 and anti ribosomal protein). Detection of anti-LKM1 auto-antibody was added. These four auto-antibodies have in common only the intracytoplasmic localisation of their respective antigen. The aim of our study was to evaluate this new kit using 104 sera and to compare our results with reference techniques (indirect immunofluorescence IF for anti-M2, anti-ribosomal protein and anti-LKM1, double immunodiffusion ID for anti-Jo-1 and anti-LKM1, western blotting WB for anti-M2) and with Cyto-Dot HM010. The one hundred and four sera were divided into five groups: Group I (n = 12) with anti-Jo-1 detected by ID; Group II (n = 28) with 26 anti-M2 positive by IF and WB, 2 anti-M2 positive only by WB; Group III (n = 10) with anti-ribosomal protein detected by IF 5 of which precipitated by ID; Group IV (n = 32) with anti-LKM1 by IF and ID divided into 18 AIH2 and 14 HCV; Group V (n = 22) consisting of 14 healthy individuals and 8 patients with hypergammaglobulinemia. Results of this study are similar to those of Cyto-Dot HM010 for the three auto-antibodies already in use. Cyto-Dot 4 is a very good anti-LKM1 confirmation method as it is ID. PMID- 15297237 TI - [Determination of anti-transglutaminase antibodies in the diagnosis of coeliac disease in children: results of a five year prospective study]. AB - AIM: To evaluate the interest of IgA antibodies to tissue transglutaminase in the diagnosis of children coeliac disease compared with anti-endomysium and anti gliadin antibodies. SUBJECTS AND METHODS: Seventy children with coeliac disease (mean age: 5 years and 8 months) and 99 disease controls (mean age: 4 years and 5 months). IgA anti-transglutaminase were tested by ELISA using a human recombinant tissue transglutaminase. IgA anti-endomysium were detected by indirect immunofluorescence on monkey oesophagus. RESULTS: The middle rate of IgA anti transglutaminase was 101.06 units in patients and only 0.47 unit in controls. IgA anti-transglutaminase and IgA anti-endomysium were in agreement in 98.8% of cases; only two cases were discordant (+/- and -/+). Globally, the two markers had the same sensitivity (90%), specificity (98%), negative (93.2%) and positive (96.9%) predictive values. For anti-gliadin antibodies, the IgG were more sensitive (88.6%) and the IgA more specific (93.9%). CONCLUSION: IgA anti-tissue transglutaminase can be used instead of IgA anti-endomysium as a serological marker of screening and diagnosis of coeliac disease in children after 3 years. PMID- 15297238 TI - [Proposal for a discriminant level of BNP in very elderly persons with heart failure]. AB - A diagnosis of heart failure (HF) in elderly patient can be difficult. The purpose of this study is to evaluate the cut-off of brain natriuretic peptide (BNP) Triage Assay (Biosite) in the diagnosis of HF in this population : 250 very older patients with age or = 2) A1 biopsies in the first 12 months after transplant. Bronchiolitis obliterans syndrome developed in 68% of patients with multiple A1 lesions at a mean of 599 +/- 435 days, compared with 43% of patients with one or less A1 lesions at a mean of 819 +/- 526 (p = 0.022). Eighteen patients experienced multiple A1 biopsies after transplant in the absence of high grade rejection episodes yet also developed earlier obliterative bronchiolitis (456 +/- 245 days, p = 0.020). We conclude that for A1 transbronchial lung biopsies, the conventional treatment of observation only is now challenged even in patients who are asymptomatic. Patients who experience multiple A1 lesions develop an earlier onset of obliterative bronchiolitis and may warrant alternative immunosuppressive strategies. PMID- 15297271 TI - Mechanical ventilation depresses protein synthesis in the rat diaphragm. AB - Prolonged mechanical ventilation results in diaphragmatic atrophy and contractile dysfunction in animals. We hypothesized that mechanical ventilation-induced diaphragmatic atrophy is associated with decreased synthesis of both mixed muscle protein and myosin heavy chain protein in the diaphragm. To test this postulate, adult rats were mechanically ventilated for 6, 12, or 18 hours and diaphragmatic protein synthesis was measured in vivo. Six hours of mechanical ventilation resulted in a 30% decrease (p < 0.05) in the rate of mixed muscle protein synthesis and a 65% decrease (p < 0.05) in the rate of myosin heavy chain protein synthesis; this depression in diaphragmatic protein synthesis persisted throughout 18 hours of mechanical ventilation. Real-time polymerase chain reaction analyses revealed that mechanical ventilation, in comparison with time matched controls, did not alter diaphragmatic levels of Type I and IIx myosin heavy chain messenger ribonucleic acid levels in the diaphragm. These data support the hypothesis that mechanical ventilation results in a decrease in both mixed muscle protein and myosin heavy chain protein synthesis in the diaphragm. Further, the decline in myosin heavy chain protein synthesis does not appear to be associated with a decrease in myosin heavy chain messenger ribonucleic acid. PMID- 15297272 TI - Permanent pacemakers and implantable defibrillators: considerations for intensivists. AB - Pacemakers and cardioverter-defibrillators are implanted in patients with cardiovascular disease for an ever-increasing array of indications. Intensivists provide care frequently for patients who have these devices, and thus, they must be familiar with common problems and nuances that may contribute to critical illness. Close collaboration of the critical care physician and cardiologist/electrophysiologist assures that pacemakers and defibrillators are tuned to optimize the hemodynamic milieu of critically ill patients. Many recent advances in the sophistication of implanted devices are reviewed herein. PMID- 15297273 TI - The Clara cell10 adenine38guanine polymorphism and sarcoidosis susceptibility in Dutch and Japanese subjects. AB - CC10 (CC16, uteroglobin) is a pulmonary protein postulated to play a counter regulatory role in sarcoidosis pathogenesis. The adenine38guanine (A38G) polymorphism of the encoding CC10 gene (SCGB1A1) is functional. Recently, an association between the low CC10 producing 38A allele and sarcoidosis susceptibility has been reported in Japanese patients from Hokkaido. The aim of the present study was to confirm this association in a clinically well characterized population of Dutch white and Kyoto Japanese patients with sarcoidosis and control subjects. No difference in genotype or allele frequency was found between patients with sarcoidosis and control subjects in either ethnic population. Remarkably, however, a significant difference was found between the control subjects from Kyoto and Hokkaido, but not between the Japanese groups of patients with sarcoidosis. Furthermore, review of previously published A38G genotyping results showed a consistent difference in CC10 A38G allele frequencies between whites and Japanese subjects. We conclude that the CC10 A38G polymorphism does not influence sarcoidosis susceptibility in Dutch whites or in Japanese subjects from Kyoto. This stresses the importance of studying the influence of polymorphisms on disease susceptibility in multiple ethnically and geographically distinct disease and control populations before reaching conclusions. PMID- 15297274 TI - Considering the role of four months of rifampin in the treatment of latent tuberculosis infection. PMID- 15297275 TI - Treated cytomegalovirus pneumonia is not associated with bronchiolitis obliterans syndrome. AB - The association of cytomegalovirus (CMV) infection with the development of bronchiolitis obliterans syndrome (BOS) is unclear. We studied 341 lung transplant recipients to assess whether histopathologically diagnosed CMV pneumonia treated with ganciclovir was a risk factor for development of BOS and patient survival. We also analyzed the relationship between CMV donor/recipient serologic status and BOS plus the temporal association between acute rejection and CMV pneumonia. Freedom from BOS for patients with (n = 151) and without (n = 190) CMV pneumonia was 83 and 90% (1 year), 52 and 56% (3 years), and 29 and 38% (5 years), respectively (p = 0.2660). Cumulative survival of patients with and without CMV pneumonia was 90 and 93% (1 year), 70 and 74% (3 years), and 58 and 63% (5 years), respectively (p = 0.1811). There were no significant differences in either development of BOS or patient survival with any combination of donor/recipient serostatus for CMV. Acute rejection occurred in the month preceding CMV pneumonia in 62 of 193 (32%) cases. Histopathologically confirmed CMV pneumonia treated with ganciclovir is not a risk factor for BOS or patient survival, nor is any particular CMV serologic donor/recipient group. CMV pneumonia often follows acute rejection, perhaps as a result of augmented immunosuppression. PMID- 15297276 TI - Unraveling the genetics of chronic kidney disease using animal models. AB - Identifying genes underlying common forms of kidney disease in humans has proven difficult, expensive, and time consuming. Quantitative trait loci (QTL) for several complex traits are concordant among mice, rats, and humans, suggesting that genetic findings from these animal models are relevant to human disease. Therefore, we reviewed the literature on genetic studies of kidney disease in rat and mouse and examined the concordance between kidney disease QTL across species. Fifteen genomic regions contribute to kidney disease in the rat, with 12 replicated either in a separate rat cross or in another species. Five loci found in humans were concordant to QTL found in the rat. Two of these were found by homology to a previously identified rat QTL on chromosome 1, demonstrating that kidney disease loci in animal models can predict the location of kidney disease loci in humans. In contrast to the rat, the mouse has been underutilized in the genetic analysis of polygenic kidney disease, although mutagenesis and QTL analysis in the mouse are likely to contribute new findings in the near future. Knowledge of kidney disease loci conserved between the mouse and rat will identify prime candidate loci to test for association with chronic kidney disease in humans. PMID- 15297277 TI - Segment-specific effects of cardiovascular risk factors on carotid artery intima medial thickness in women at midlife. AB - OBJECTIVE: We investigated associations between segment-specific carotid intima medial thickness (IMT) and cardiovascular risk factors collected before menopause for insight into mechanisms of atherosclerosis development. METHODS AND RESULTS: Participants were 453 healthy women (aged 46 to 58 years) enrolled in a dietary and physical activity randomized clinical trial. Ultrasound scan measures were taken approximately 2.7 years after baseline in the common carotid artery (CCA), bifurcation (bulb), and internal carotid artery (ICA) segments. When scanned, 84% remained premenopausal. In linear regression models adjusted for age, menopausal status, and intervention group, measures independently (P<0.05) and positively associated were as follows: baseline weight (beta=0.007 per 5 kg), systolic blood pressure (SBP; beta=0.008 per 10 mm Hg), and age (beta=0.02 per 5 years) with CCA IMT; smoking (beta=0.08), weight (beta=0.009), and SBP (beta=0.02) with bulb IMT; and apoprotein B (beta=0.01 per 0.1 g/L) with ICA IMT. Differential effects in a repeated measures model with all 3 IMT locations showed these risk factors to have segment-specific positive associations. The effect of weight was strongest in the CCA, smoking and SBP were specific to the bulb, and apoprotein B was strongest in the ICA segment. CONCLUSIONS: Analyses indicate that cardiovascular risk factors may differentially affect IMT in the CCA, bulb, and ICA segments of healthy middle-aged women. PMID- 15297278 TI - Gene expression phenotypes of atherosclerosis. AB - OBJECTIVE: Fulfilling the promise of personalized medicine by developing individualized diagnostic and therapeutic strategies for atherosclerosis will depend on a detailed understanding of the genes and gene variants that contribute to disease susceptibility and progression. To that end, our group has developed a nonbiased approach congruent with the multigenic concept of complex diseases by identifying gene expression patterns highly associated with disease states in human target tissues. METHODS AND RESULTS: We have analyzed a collection of human aorta samples with varying degrees of atherosclerosis to identify gene expression patterns that predict a disease state or potential susceptibility. We find gene expression signatures that relate to each of these disease measures and are reliable and robust in predicting the classification for new samples with >93% in each analysis. The genes that provide the predictive power include many previously suspected to play a role in atherosclerosis and additional genes without prior association with atherosclerosis. CONCLUSIONS: Hence, we are reporting a novel method for generating a molecular phenotype of disease and then identifying genes whose discriminatory capability strongly implicates their potential roles in human atherosclerosis. PMID- 15297279 TI - Infliximab in spondyloarthropathy associated with Crohn's disease: an open study on the efficacy of inducing and maintaining remission of musculoskeletal and gut manifestations. AB - OBJECTIVE: To evaluate the efficacy and tolerability of anti-tumour necrosis factor alpha (TNFalpha) monoclonal antibody (infliximab) in the treatment of spondyloarthropathy (SpA) associated with active and inactive Crohn's disease (CD). METHODS: Twenty four patients with SpA associated with active or inactive CD (16 active, 8 quiescent) were treated with anti-TNFalpha monoclonal antibody (infliximab) with repeated infusions for a period of 12-18 months. The treatment aimed at ameliorating the general musculoskeletal and spinal pain, controlling peripheral arthritis and enthesitis, decreasing the BASDAI score, modifying acute phase reactants, and reducing CD activity. RESULTS: Infliximab improved both gastrointestinal (p<0.01) and overall articular symptoms (BASDAI, p<0.01; general musculoskeletal and spinal pain, p<0.01; peripheral arthritis, p<0.01) in patients with active CD. Additionally, infliximab effectively controlled not only axial involvement and peripheral arthritis but also enthesitis (p<0.01) and prevented inflammatory bowel disease reactivation in patients with inactive CD and low inflammatory markers. Amelioration of gut and musculoskeletal involvement persisted for up to 12 months. CONCLUSION: Infliximab may act on the inflammation of entheses and of periarticular structures, which usually does not cause a change in the haematological markers that are the main indicators of pain and joint ankylosis in SpA. Infliximab induces and maintains remission of CD while at the same time treating active and severe SpA, suggesting that it should be the preferred drug for the treatment of active and severe SpA associated with active or quiescent CD. PMID- 15297280 TI - Scoring of radiographic progression in randomised clinical trials in ankylosing spondylitis: a preference for paired reading order. AB - OBJECTIVES: To describe the influence of the reading order (chronological v paired) on radiographic scoring results in ankylosing spondylitis. To investigate whether this method is sufficiently sensitive to change because paired reading is requested for establishing drug efficacy in clinical trials. METHODS: Films obtained from 166 patients (at baseline, 1 year, and 2 years) were scored by one observer, using the modified Stoke Ankylosing Spondylitis Spinal Score. Films were first scored chronologically, and were scored paired 6 months later. RESULTS: Chronological reading showed significantly more progression than paired reading both at 1 year (mean (SD) progression 1.3 (2.6) v 0.5 (2.4) units) and at 2 years (2.1 (3.9) v 1.0 (2.9) units); between-method difference: p<0.001 at 1 year, and p<0.001 at 2 years. After 1 year, progression (>0 units) was found in 35/166 (21%) patients after paired reading and in 55/166 (33%) after chronological reading. After 2 years, these figures were 50/166 (30%) and 68/166 (41%), respectively. Sample size calculations showed that 94 patients in each treatment arm are required in a randomised clinical trial (RCT) to provide sufficient statistical power to detect a difference in 2 year progression if films are scored paired. CONCLUSION: Reading with chronological time order is more sensitive to change than reading with paired time order, but paired reading is sufficiently sensitive to pick up change with a follow up of 2 years, resulting in an acceptable sample size for RCTs. PMID- 15297281 TI - Autoantibody formation in patients with rheumatoid arthritis treated with anti TNF alpha. AB - BACKGROUND: Research on autoantibody formation in patients treated with TNF alpha inhibitors has produced contradictory results. OBJECTIVE: To study the prevalence of autoantibodies in patients with rheumatoid arthritis treated with the TNF alpha inhibitor infliximab. METHODS: 53 patients (48 female, 11 male) treated with infliximab for rheumatoid arthritis were followed for autoantibody production before treatment and after 14, 30, and 54 weeks. Six patients treated with etanercept were studied for comparison. The analyses included antibodies against nuclear antigens (ANA), extractable nuclear antigens, double stranded (ds)DNA (by ELISA, IIF on Crithidia luciliae for IgM and IgG, and Farr assay), nucleosomes, cardiolipin, smooth muscle, mitochondria, proteinase 3, and myeloperoxidase antigens. RESULTS: The number of patients treated with infliximab who developed antibodies against dsDNA of both IgG and IgM class (tested by IIF) increased significantly. The prevalence of patients positive for IgG class increased to 66% at 30 weeks and 45% at 54 weeks, and of IgM class to 85% and 70%, respectively. The titre and number of patients expressing antibodies against nucleosomes and ANA also increased significantly. The number of rheumatoid factor or anticardiolipin positive patients was stable and there was no increase in antibodies against the other antigens. A lupus-like syndrome was seen in one patient. No patient treated with etanercept developed any of these autoantibodies. CONCLUSIONS: Patients treated with infliximab may develop anti dsDNA antibodies of both IgM and IgG class, anti-nucleosome antibodies, and ANA, with a gradual increase until 30 weeks. PMID- 15297282 TI - First clinical evaluation of sagittal laser optical tomography for detection of synovitis in arthritic finger joints. AB - OBJECTIVE: To identify classifiers in images obtained with sagittal laser optical tomography (SLOT) that can be used to distinguish between joints affected and not affected by synovitis. METHODS: 78 SLOT images of proximal interphalangeal joints II-IV from 13 patients with rheumatoid arthritis were compared with ultrasound (US) images and clinical examination (CE). SLOT images showing the spatial distribution of scattering and absorption coefficients within the joint cavity were generated. The means and standard errors for seven different classifiers (operator score and six quantitative measurements) were determined from SLOT images using CE and US as diagnostic references. For classifiers showing significant differences between affected and non-affected joints, sensitivities and specificities for various cut off parameters were obtained by receiver operating characteristic (ROC) analysis. RESULTS: For five classifiers used to characterise SLOT images the mean between affected and unaffected joints was statistically significant using US as diagnostic reference, but statistically significant for only one classifier with CE as reference. In general, high absorption and scattering coefficients in and around the joint cavity are indicative of synovitis. ROC analysis showed that the minimal absorption classifier yields the largest area under the curve (0.777; sensitivity and specificity 0.705 each) with US as diagnostic reference. CONCLUSION: Classifiers in SLOT images have been identified that show statistically significant differences between joints with and without synovitis. It is possible to classify a joint as inflamed with SLOT, without the need for a reference measurement. Furthermore, SLOT based diagnosis of synovitis agrees better with US diagnosis than CE. PMID- 15297283 TI - Tumour necrosis factor alpha blockade in treatment resistant pigmented villonodular synovitis. AB - BACKGROUND: Pigmented villonodular synovitis (PVNS) is considered to be a neoplastic-like disorder of the synovium histologically characterised by villonodular hyperplasia, resulting in dense fibrosis and haemosiderin deposition. The pathogenesis of the disease is still unknown. CASE REPORT: A patient presented with severe treatment resistant PVNS of the right knee joint. Several conventional treatment regimens, including open surgical synovectomy and intra-articular injections of yttrium-90 ((90)Y) failed to control the disease. After finding marked tumour necrosis factor alpha (TNF alpha) expression in arthroscopic synovial tissue samples, treatment with an anti-TNF alpha monoclonal antibody (infliximab) at a dose of 5 mg/kg was started. Additional courses with the same dose given 2, 6, 14, and 20 weeks later, and bimonthly thereafter up to 54 weeks, controlled the signs and symptoms. Immunohistological analysis at follow up identified a marked reduction in macrophage numbers and TNF alpha expression in the synovium. DISCUSSION: This is probably the first case which describes treatment with TNF alpha blockade of PVNS in a patient who is refractory to conventional treatment. It provides the rationale for larger controlled studies to elucidate further the efficacy of TNFalpha blockade treatment in refractory PVNS. PMID- 15297284 TI - A summary of implications of recent clinical trials for the National Cholesterol Education Program Adult Treatment Panel III guidelines. PMID- 15297285 TI - Aortic valve: turning over a new leaf(let) in endothelial phenotypic heterogeneity. PMID- 15297286 TI - How does mutant proprotein convertase neural apoptosis-regulated convertase 1 induce autosomal dominant hypercholesterolemia? PMID- 15297287 TI - The mystery of PCSK9. PMID- 15297288 TI - ATVB in focus: Diabetic vascular disease: pathophysiological mechanisms in the diabetic milieu and therapeutic implications. PMID- 15297289 TI - Common polymorphism in the MTP promoter attenuates the dyslipidemic and proatherogenic effects of excess body weight. PMID- 15297290 TI - Associations among plasma lipoprotein subfractions as characterized by analytical capillary isotachophoresis, apolipoprotein E phenotype, Alzheimer disease, and mild cognitive impairment. PMID- 15297291 TI - Is there really a power shortage in clinical trials testing the "homocysteine hypothesis?". PMID- 15297292 TI - Implications of recent clinical trials for the National Cholesterol Education Program Adult Treatment Panel III guidelines. AB - The Adult Treatment Panel III (ATP III) of the National Cholesterol Education Program issued an evidence-based set of guidelines on cholesterol management in 2001. Since the publication of ATP III, 5 major clinical trials of statin therapy with clinical end points have been published. These trials addressed issues that were not examined in previous clinical trials of cholesterol-lowering therapy. The present document reviews the results of these recent trials and assesses their implications for cholesterol management. Therapeutic lifestyle changes (TLC) remain an essential modality in clinical management. The trials confirm the benefit of cholesterol-lowering therapy in high-risk patients and support the ATP III treatment goal of low-density lipoprotein cholesterol (LDL-C) <100 mg/dL. They support the inclusion of patients with diabetes in the high-risk category and confirm the benefits of LDL-lowering therapy in these patients. They further confirm that older persons benefit from therapeutic lowering of LDL-C. The major recommendations for modifications to footnote the ATP III treatment algorithm are the following. In high-risk persons, the recommended LDL-C goal is <100 mg/dL, but when risk is very high, an LDL-C goal of <70 mg/dL is a therapeutic option, ie, a reasonable clinical strategy, on the basis of available clinical trial evidence. This therapeutic option extends also to patients at very high risk who have a baseline LDL-C <100 mg/dL. Moreover, when a high-risk patient has high triglycerides or low high-density lipoprotein cholesterol (HDL-C), consideration can be given to combining a fibrate or nicotinic acid with an LDL-lowering drug. For moderately high-risk persons (2+ risk factors and 10-year risk 10% to 20%), the recommended LDL-C goal is <130 mg/dL, but an LDL-C goal <100 mg/dL is a therapeutic option on the basis of recent trial evidence. The latter option extends also to moderately high-risk persons with a baseline LDL-C of 100 to 129 mg/dL. When LDL-lowering drug therapy is employed in high-risk or moderately high risk persons, it is advised that intensity of therapy be sufficient to achieve at least a 30% to 40% reduction in LDL-C levels. Moreover, any person at high risk or moderately high risk who has lifestyle-related risk factors (eg, obesity, physical inactivity, elevated triglycerides, low HDL-C, or metabolic syndrome) is a candidate for TLC to modify these risk factors regardless of LDL-C level. Finally, for people in lower-risk categories, recent clinical trials do not modify the goals and cutpoints of therapy. PMID- 15297293 TI - PIML: the Pathogen Information Markup Language. AB - MOTIVATION: A vast amount of information about human, animal and plant pathogens has been acquired, stored and displayed in varied formats through different resources, both electronically and otherwise. However, there is no community standard format for organizing this information or agreement on machine-readable format(s) for data exchange, thereby hampering interoperation efforts across information systems harboring such infectious disease data. RESULTS: The Pathogen Information Markup Language (PIML) is a free, open, XML-based format for representing pathogen information. XSLT-based visual presentations of valid PIML documents were developed and can be accessed through the PathInfo website or as part of the interoperable web services federation known as ToolBus/PathPort. Currently, detailed PIML documents are available for 21 pathogens deemed of high priority with regard to public health and national biological defense. A dynamic query system allows simple queries as well as comparisons among these pathogens. Continuing efforts are being taken to include other groups' supporting PIML and to develop more PIML documents. AVAILABILITY: All the PIML-related information is accessible from http://www.vbi.vt.edu/pathport/pathinfo/ PMID- 15297294 TI - Predicting GPCR-G-protein coupling using hidden Markov models. AB - MOTIVATION: Determining the coupling specificity of G-protein coupled receptors (GPCRs) is important for understanding the biology of this class of pharmacologically important proteins. Currently available in silico methods for predicting GPCR-G-protein coupling specificity have high error rate. METHOD: We introduce a new approach for creating hidden Markov models (HMMs) based on a first guess about the importance of various residues. We call these knowledge restricted HMMs to emphasize the fact that the state space of the HMM is restricted by the application of a priori knowledge. Specifically, we use only those amino acid residues of GPCRs which are likely to interact with G-proteins, namely those that are predicted to be in the intra-cellular loops. Furthermore, we concatenate these predicted loops into one sequence rather than considering them as four disparate units. This reduces the HMM state space by drastically decreasing the sequence length. RESULTS: Our knowledge restricted HMM based method to predict GPCR-G-protein coupling specificity has an error rate of <1%, when applied to a test set of GPCRs with known G-protein coupling specificity. AVAILABILITY: Academic users can get the data set mentioned herein and HMMs from the authors. PMID- 15297295 TI - Comparative analysis of methods for representing and searching for transcription factor binding sites. AB - MOTIVATION: An important step in unravelling the transcriptional regulatory network of an organism is to identify, for each transcription factor, all of its DNA binding sites. Several approaches are commonly used in searching for a transcription factor's binding sites, including consensus sequences and position specific scoring matrices. In addition, methods that compute the average number of nucleotide matches between a putative site and all known sites can be employed. Such basic approaches can all be naturally extended by incorporating pairwise nucleotide dependencies and per-position information content. In this paper, we evaluate the effectiveness of these basic approaches and their extensions in finding binding sites for a transcription factor of interest without erroneously identifying other genomic sequences. RESULTS: In cross validation testing on a dataset of Escherichia coli transcription factors and their binding sites, we show that there are statistically significant differences in how well various methods identify transcription factor binding sites. The use of per-position information content improves the performance of all basic approaches. Furthermore, including local pairwise nucleotide dependencies within binding site models results in statistically significant performance improvements for approaches based on nucleotide matches. Based on our analysis, the best results when searching for DNA binding sites of a particular transcription factor are obtained by methods that incorporate both information content and local pairwise correlations. AVAILABILITY: The software is available at http://compbio.cs.princeton.edu/bindsites. PMID- 15297296 TI - limmaGUI: a graphical user interface for linear modeling of microarray data. AB - SUMMARY: limmaGUI is a graphical user interface (GUI) based on R-Tcl/Tk for the exploration and linear modeling of data from two-color spotted microarray experiments, especially the assessment of differential expression in complex experiments. limmaGUI provides an interface to the statistical methods of the limma package for R, and is itself implemented as an R package. The software provides point and click access to a range of methods for background correction, graphical display, normalization, and analysis of microarray data. Arbitrarily complex microarray experiments involving multiple RNA sources can be accomodated using linear models and contrasts. Empirical Bayes shrinkage of the gene-wise residual variances is provided to ensure stable results even when the number of arrays is small. Integrated support is provided for quantitative spot quality weights, control spots, within-array replicate spots and multiple testing. limmaGUI is available for most platforms on the which R runs including Windows, Mac and most flavors of Unix. AVAILABILITY: http://bioinf.wehi.edu.au/limmaGUI. PMID- 15297297 TI - Training HMM structure with genetic algorithm for biological sequence analysis. AB - SUMMARY: Hidden Markov models (HMMs) are widely used for biological sequence analysis because of their ability to incorporate biological information in their structure. An automatic means of optimizing the structure of HMMs would be highly desirable. However, this raises two important issues; first, the new HMMs should be biologically interpretable, and second, we need to control the complexity of the HMM so that it has good generalization performance on unseen sequences. In this paper, we explore the possibility of using a genetic algorithm (GA) for optimizing the HMM structure. GAs are sufficiently flexible to allow incorporation of other techniques such as Baum-Welch training within their evolutionary cycle. Furthermore, operators that alter the structure of HMMs can be designed to favour interpretable and simple structures. In this paper, a training strategy using GAs is proposed, and it is tested on finding HMM structures for the promoter and coding region of the bacterium Campylobacter jejuni. The proposed GA for hidden Markov models (GA-HMM) allows, HMMs with different numbers of states to evolve. To prevent over-fitting, a separate dataset is used for comparing the performance of the HMMs to that used for the Baum-Welch training. The GA-HMM was capable of finding an HMM comparable to a hand-coded HMM designed for the same task, which has been published previously. PMID- 15297298 TI - Highly specific prediction of phosphorylation sites in proteins. AB - SUMMARY: The prediction of significant short functional protein sequences has inherent problems. In predicting phosphorylation sites, problems came from the shortness of phosphorylation sites, the difficulties in maintaining many different predefined models of binding sites, and the difficulties of obtaining highly sensitive predictions and of obtaining predictions with a constant sensitivity and specificity. The algorithm presented in this paper overcomes these problems. The proposed algorithm PHOSITE is based on the case-based sequence analysis. This enables the prediction of phosphorylation sites with constant specificity and sensitivity. Furthermore, this method leads not only to the prediction of phosphorylation sites in general but also predicts the most probable type of kinase involved. AVAILABILITY: The tool PHOSITE implementing the presented method can be evaluated under the website http://www.phosite.com. PMID- 15297299 TI - GO::TermFinder--open source software for accessing Gene Ontology information and finding significantly enriched Gene Ontology terms associated with a list of genes. AB - SUMMARY: GO::TermFinder comprises a set of object-oriented Perl modules for accessing Gene Ontology (GO) information and evaluating and visualizing the collective annotation of a list of genes to GO terms. It can be used to draw conclusions from microarray and other biological data, calculating the statistical significance of each annotation. GO::TermFinder can be used on any system on which Perl can be run, either as a command line application, in single or batch mode, or as a web-based CGI script. AVAILABILITY: The full source code and documentation for GO::TermFinder are freely available from http://search.cpan.org/dist/GO-TermFinder/. PMID- 15297300 TI - Haploview: analysis and visualization of LD and haplotype maps. AB - Research over the last few years has revealed significant haplotype structure in the human genome. The characterization of these patterns, particularly in the context of medical genetic association studies, is becoming a routine research activity. Haploview is a software package that provides computation of linkage disequilibrium statistics and population haplotype patterns from primary genotype data in a visually appealing and interactive interface. AVAILABILITY: http://www.broad.mit.edu/mpg/haploview/ CONTACT: jcbarret@broad.mit.edu PMID- 15297301 TI - Network structures and algorithms in Bioconductor. AB - In this paper, we review the central concepts and implementations of tools for working with network structures in Bioconductor. Interfaces to open source resources for visualization (AT&T Graphviz) and network algorithms (Boost) have been developed to support analysis of graphical structures in genomics and computational biology. AVAILABILITY: Packages graph, Rgraphviz, RBGL of Bioconductor (www.bioconductor.org). PMID- 15297302 TI - Modular, scriptable and automated analysis tools for high-throughput peptide mass fingerprinting. AB - A set of new algorithms and software tools for automatic protein identification using peptide mass fingerprinting is presented. The software is automatic, fast and modular to suit different laboratory needs, and it can be operated either via a Java user interface or called from within scripts. The software modules do peak extraction, peak filtering and protein database matching, and communicate via XML. Individual modules can therefore easily be replaced with other software if desired, and all intermediate results are available to the user. The algorithms are designed to operate without human intervention and contain several novel approaches. The performance and capabilities of the software is illustrated on spectra from different mass spectrometer manufacturers, and the factors influencing successful identification are discussed and quantified. MOTIVATION: Protein identification with mass spectrometric methods is a key step in modern proteomics studies. Some tools are available today for doing different steps in the analysis. Only a few commercial systems integrate all the steps in the analysis, often for only one vendor's hardware, and the details of these systems are not public. RESULTS: A complete system for doing protein identification with peptide mass fingerprints is presented, including everything from peak picking to matching the database protein. The details of the different algorithms are disclosed so that academic researchers can have full control of their tools. AVAILABILITY: The described software tools are available from the Halmstad University website www.hh.se/staff/bioinf/ SUPPLEMENTARY INFORMATION: Details of the algorithms are described in supporting information available from the Halmstad University website www.hh.se/staff/bioinf/ PMID- 15297303 TI - How many samples are needed to build a classifier: a general sequential approach. AB - MOTIVATION: The standard paradigm for a classifier design is to obtain a sample of feature-label pairs and then to apply a classification rule to derive a classifier from the sample data. Typically in laboratory situations the sample size is limited by cost, time or availability of sample material. Thus, an investigator may wish to consider a sequential approach in which there is a sufficient number of patients to train a classifier in order to make a sound decision for diagnosis while at the same time keeping the number of patients as small as possible to make the studies affordable. RESULTS: A sequential classification procedure is studied via the martingale central limit theorem. It updates the classification rule at each step and provides stopping criteria to ensure with a certain confidence that at stopping a future subject will have misclassification probability smaller than a predetermined threshold. Simulation studies and applications to microarray data analysis are provided. The procedure possesses several attractive properties: (1) it updates the classification rule sequentially and thus does not rely on distributions of primary measurements from other studies; (2) it assesses the stopping criteria at each sequential step and thus can substantially reduce cost via early stopping; and (3) it is not restricted to any particular classification rule and therefore applies to any parametric or non-parametric method, including feature selection or extraction. AVAILABILITY: R-code for the sequential stopping rule is available at http://stat.tamu.edu/~wfu/microarray/sequential/R-code.html PMID- 15297304 TI - The fate and career destinations of doctors who qualified at Uganda's Makerere Medical School in 1984: retrospective cohort study. PMID- 15297305 TI - Effectiveness of a multiple intervention to reduce antibiotic prescribing for respiratory tract symptoms in primary care: randomised controlled trial. AB - OBJECTIVES: To assess the effectiveness of a multiple intervention aimed at reducing antibiotic prescription rates for symptoms of the respiratory tract in primary care. DESIGN: Randomised controlled trial. SUBJECTS: Twelve peer review groups including 100 general practitioners with their collaborating pharmacists in the region of Utrecht, Netherlands. INTERVENTION: The intervention consisted of group education meetings, with a consensus procedure on indication for and type of antibiotics and with training in communication skills; monitoring and feedback on prescribing behaviour; group education for assistants of general practitioners and pharmacists; and education material for patients. The control group did not receive any of these elements. MAIN OUTCOME MEASURES: Antibiotic prescription rates for acute symptoms of the respiratory tract and patients' satisfaction. RESULTS: 89 general practitioners completed the study (89%). At baseline, prescription rates for antibiotics for respiratory tract symptoms did not differ between intervention and control group (27% v 29%, respectively). After nine months, the prescription rates in the intervention group fell to 23%, whereas the control group's rose to 37% (mean difference in change -12%, 95% confidence interval -18.9% to -4.0%). Multilevel analysis confirmed the results of the unadjusted analysis (intervention effect -10.7%, -20.3% to -1.0%). Patients' satisfaction was high and did not differ in the two groups at baseline or after the intervention. CONCLUSIONS: A multiple intervention reduced prescribing rates of antibiotics for respiratory tract symptoms while maintaining a high degree of satisfaction among patients. Further research should focus on the sustainability and cost effectiveness of this intervention. PMID- 15297306 TI - The nucleotide transporter MRP4 (ABCC4) is highly expressed in human platelets and present in dense granules, indicating a role in mediator storage. AB - Platelet aggregation is initiated by the release of mediators as adenosine diphosphate (ADP) stored in platelet granules. Possible candidates for transport proteins mediating accumulation of these mediators in granules include multidrug resistance protein 4 (MRP4, ABCC4), a transport pump for cyclic nucleotides and nucleotide analogs. We investigated the expression of MRP4 in human platelets by immunoblotting, detecting a strong signal at 170 kDa. Immunofluorescence microscopy using 2 MRP4-specific antibodies revealed staining mainly in intracellular structures, which largely colocalized with the accumulation of mepacrine as marker for delta-granules and to a lower extent at the plasma membrane. Furthermore, an altered distribution of MRP4 was observed in platelets from a patient with Hermansky-Pudlak syndrome with defective delta-granules. Adenosine triphosphate (ATP)-dependent cyclic guanosine monophosphate (cGMP) transport codistributed with MRP4 detection in subcellular fractions, with highest activities in the dense granule and plasma membrane fractions. This transport was inhibited by dipyramidole, indomethacin, and MK571 with median inhibitory concentration (IC(50)) values of 12, 22, and 43 microM, and by ibuprofen. Transport studies with [(3)H]ADP indicated the presence of an orthovanadate-sensitive ADP transporting system, inhibited by dipyramidole, MK571, and cyclic nucleotides. The results indicate a function of MRP4 in platelet mediator storage and inhibition of MRP4 may represent a novel mechanism for inhibition of platelet function by some anti-inflammatory drugs. PMID- 15297307 TI - Differential regulation of P-selectin ligand expression in naive versus memory CD4+ T cells: evidence for epigenetic regulation of involved glycosyltransferase genes. AB - Lymphocytes are targeted to inflamed sites by specific "homing" and chemokine receptors. Most of them, including ligands for P- and E-selectin, are absent from naive CD4(+) T cells and become induced after activation and differentiation in effector/memory cells. Polarized effector cells are characterized by the rapid production of distinct cytokines upon restimulation. Their cytokine memory is in part controlled by epigenetic imprinting during differentiation. Here we ask whether a similar mechanism could regulate selectin ligand expression, mediating entry into inflamed sites, notably within the skin. We report that acquisition of selectin ligands by naive but not memory CD4(+) cells depends on progression through the G(1)/S phase of the cell cycle-a phase susceptible to modification of the chromatin structure. Cell-cycle arrest prevented transcriptional activation of glycosyltransferases involved in the generation of selectin ligands, suggesting that progression through the cell cycle is required to unlock their genes. Artificial DNA demethylation strongly increased the frequency of selectin ligand-expressing cells, suggesting that DNA methylation keeps transferase genes inaccessible in naive T cells. Due to these findings we propose that selectin dependent inflammation-seeking properties are imprinted by epigenetic modifications upon T-cell differentiation into effector cells. PMID- 15297308 TI - Nonhematopoietic mesenchymal stem cells can be mobilized and differentiate into cardiomyocytes after myocardial infarction. AB - Bone marrow (BM) cells are reported to contribute to the process of regeneration following myocardial infarction. However, the responsible BM cells have not been fully identified. Here, we used 2 independent clonal studies to determine the origin of bone marrow (BM)-derived cardiomyocytes. First, we transplanted single CD34(-) c-kit(+)Sca-1(+) lineage(-) side population (CD34(-)KSL-SP) cells or whole BM cells from mice ubiquitously expressing enhanced green fluorescent protein (EGFP) into lethally irradiated mice, induced myocardial infarction (MI), and treated the animals with granulocyte colony-stimulating factor (G-CSF) to mobilize stem cells to the damaged myocardium. At 8 weeks after MI, from 100 specimens we counted only 3 EGFP(+) actinin(+) cells in myocardium of CD34(-) KSL SP cells in mice that received transplants, but more than 5000 EGFP(+) actinin(+) cells in whole BM cell in mice that received transplants, suggesting that most of EGFP(+) actinin(+) cells were derived from nonhematopoietic BM cells. Next, clonally purified nonhematopoietic mesenchymal stem cells (MSCs), cardiomyogenic (CMG) cells, that expressed EGFP in the cardiomyocyte-specific manner were transplanted directly into BM of lethally irradiated mice, MI was induced, and they were treated with G-CSF. EGFP(+) actinin(+) cells were observed in the ischemic myocardium, indicating that CMG cells had been mobilized and differentiated into cardiomyocytes. Together, these results suggest that the origin of the vast majority of BM-derived cardiomyocytes is MSCs. PMID- 15297309 TI - Haploinsufficiency of AML1 results in a decrease in the number of LTR-HSCs while simultaneously inducing an increase in more mature progenitors. AB - The AML1/CBFbeta transcriptional complex is essential for the formation of definitive hematopoietic stem cells (HSCs). Moreover, development of the hematopoietic system is exquisitely sensitive to the level of this complex. To investigate the effect of AML1 dosage on adult hematopoiesis, we compared the hematopoietic systems of AML1+/- and AML1+/+ mice. Surprisingly, loss of a single AML1 allele resulted in a 50% reduction in long-term repopulating hematopoietic stem cells (LTR-HSCs). This decrease did not, however, extend to the next level of hematopoietic differentiation. Instead, AML1+/- mice had an increase in multilineage progenitors, an expansion that resulted in enhanced engraftment following transplantation. The expanded pool of AML1+/- progenitors remained responsive to homeostatic mechanisms and thus the number of mature cells in most lineages remained within normal limits. Two notable exceptions were a decrease in CD4(+) T cells, leading to an inversion of the CD4(+) to CD8(+) T-cell ratio and a decrease in circulating platelets. These data demonstrate a dosage-dependent role for AML1/CBFbeta in regulating the quantity of HSCs and their downstream committed progenitors, as well as a more restricted role in T cells and platelets. The latter defect mimics one of the key abnormalities in human patients with the familial platelet disorder resulting from AML1 haploinsufficiency. PMID- 15297310 TI - Combined disruption of both the MEK/ERK and the IL-6R/STAT3 pathways is required to induce apoptosis of multiple myeloma cells in the presence of bone marrow stromal cells. AB - The interleukin-6 receptor (IL-6R)/signal transducer and activator of transcription 3 (STAT3) pathway contributes to the pathogenesis of multiple myeloma (MM) and protects MM cells from apoptosis. However, MM cells survive the IL-6R blockade if they are cocultured with bone marrow stromal cells (BMSCs), suggesting that the BM microenvironment stimulates IL-6-independent pathways that exert a pro-survival effect. The goal of this study was to investigate the underlying mechanism. Detailed pathway analysis revealed that BMSCs stimulate STAT3 via the IL-6R, and mitogen-activated protein (MAP) kinases via IL-6R independent mechanisms. Abolition of MEK1,2 activity with PD98059, or ERK1,2 small interfering RNA knockdown, was insufficient to induce apoptosis. However, the combined disruption of the IL-6R/STAT3 and MEK1,2/ERK1,2 pathways led to strong induction of apoptosis even in the presence of BMSCs. This effect was observed with MM cell lines and with primary MM cells, suggesting that the BMSC induced activation of MEK1,2/ERK1,2 renders MM cells IL-6R/STAT3 independent. Therefore, in the presence of cells from the BM micro-environment, combined targeting of different (and independently activated) pathways is required to efficiently induce apoptosis of MM cells. This might have direct implications for the development of future therapeutic strategies for MM. PMID- 15297311 TI - Global regulation of erythroid gene expression by transcription factor GATA-1. AB - Transcription factor GATA-1 is required for erythropoiesis, yet its full actions are unknown. We performed transcriptome analysis of G1E-ER4 cells, a GATA-1-null erythroblast line that undergoes synchronous erythroid maturation when GATA-1 activity is restored. We interrogated more than 9000 transcripts at 6 time points representing the transition from late burst forming unit-erythroid (BFU-E) to basophilic erythroblast stages. Our findings illuminate several new aspects of GATA-1 function. First, the large number of genes responding quickly to restoration of GATA-1 extends the repertoire of its potential targets. Second, many transcripts were rapidly down-regulated, highlighting the importance of GATA 1 in gene repression. Third, up-regulation of some known GATA-1 targets was delayed, suggesting that auxiliary factors are required. For example, induction of the direct GATA-1 target gene beta major globin was late and, surprisingly, required new protein synthesis. In contrast, the gene encoding Fog1, which cooperates with GATA-1 in beta globin transcription, was rapidly induced independently of protein synthesis. Guided by bioinformatic analysis, we demonstrated that selected regions of the Fog1 gene exhibit enhancer activity and in vivo occupancy by GATA-1. These findings define a regulatory loop for beta globin expression and, more generally, demonstrate how transcriptome analysis can be used to generate testable hypotheses regarding transcriptional networks. PMID- 15297312 TI - Establishment of the CD4+ T-cell pool in healthy children and untreated children infected with HIV-1. AB - Current understanding of how the T-cell pool is established in children and how this is affected by HIV infection is limited. It is widely believed that the thymus is the main source for T cells during childhood. Here we show, however, that healthy children had an age-related increase in total body numbers of naive and memory T cells, whereas absolute numbers of T-cell receptor excision circles (TRECs) did not increase. This suggests that expansion of the naive T-cell pool after birth is more dependent on T-cell proliferation than was previously recognized. Indeed, the proportion of dividing naive T cells was high, especially in younger children, which is consistent with expansion through proliferation, in addition to antigen-mediated naive T-cell activation leading to formation of the memory T-cell pool. In untreated children infected with HIV-1, total body numbers of T cells and TRECs were low and stable, whereas T-cell division levels were significantly higher than in healthy children. We postulate that in children infected with HIV, similar to adults infected with HIV, continuous activation of naive T cells leads to erosion of the naive T-cell pool and may be a major factor in lowering CD4(+) T-cell numbers. PMID- 15297313 TI - Infant acute lymphoblastic leukemia with MLL gene rearrangements: outcome following intensive chemotherapy and hematopoietic stem cell transplantation. AB - Forty-four infants with acute lymphoblastic leukemia (ALL) characterized by MLL gene rearrangements were treated on a protocol of intensive chemotherapy followed by hematopoietic stem cell transplantation (HSCT) between November 1998 and June 2002. The remission induction rate was 91.0%, and the 3-year overall survival and event-free survival (EFS) rates, with 95% confidence intervals, were 58.2% (43.5% 72.9%) and 43.6% (28.5%-58.7%), respectively. Univariate analysis of EFS by presenting features indicated a poorer outcome in patients younger than 6 months of age with high white blood cell counts (>/= 100 x 10(9)/L; EFS rate, 9.4% versus 55.1% for all others, P = .0036) and in those with central nervous system invasion (EFS rate, 10.0% versus 56.9% for all others, P = .0073). The 3-year posttransplantation EFS rate for the 29 patients who underwent HSCT in first remission was 64.4% (46.4%-82.4%). In this subgroup, only the timing of HSCT (first remission versus others) was a significant risk factor by multivariate analysis (P < .0001). These results suggest that early introduction of HSCT, possibly with a less toxic conditioning regimen, may improve the prognosis for infants with MLL(+) ALL. Identification of subgroups or patients who respond well to intensified chemotherapy alone should have a high priority in future investigations. PMID- 15297314 TI - Stimulation of endothelial cell proliferation by FGF-2 in the presence of fibrinogen requires alphavbeta3. AB - We have shown previously that fibrin(ogen) binding potentiates the capacity of fibroblast growth factor 2 (FGF-2) to stimulate endothelial cell (EC) proliferation. We have now investigated the receptor requirement for EC proliferation by fibrinogen-bound FGF-2. ECs were cultured with 25 ng/mL FGF-2 with or without 10 microg/mL fibrinogen, and proliferation was measured as (3)H thymidine incorporation. Proliferation was increased 2.4 +/- 0.5-fold over medium alone with FGF-2 and increased significantly more to 4.0 +/- 0.7-fold with fibrinogen and FGF-2 (P < .005). Addition of 7E3 or LM609, antibodies to alpha(v)beta(3), inhibited EC proliferation with fibrinogen-bound FGF-2 by 80% +/ 8% (P < .001) or 67% +/- 14% (P < .002), respectively, to levels significantly less than that observed with FGF-2 alone (P < .001). Neither LM609 nor 7E3 exhibited any inhibition of activity with FGF-2 alone. Peptide GRGDS caused dose dependent inhibition of proliferation by fibrinogen-bound FGF-2 of 31% +/- 8%, 45% +/- 9%, and 68% +/- 11% at 0.25, 0.5, and 1 mM, respectively. Coimmunoprecipitation and immunofluorescence studies demonstrated a direct specific association between alpha(v)beta(3) and FGF receptor 1 (FGFR1) in ECs and fibroblasts when exposed to both FGF-2 and fibrinogen but not with vitronectin. We conclude that fibrinogen binding of FGF-2 enhances EC proliferation through the coordinated effects of colocalized alpha(v)beta(3) and FGFR1. PMID- 15297315 TI - A shortened activated partial thromboplastin time is associated with the risk of venous thromboembolism. AB - Hypercoagulability due to high coagulation factors XI, VIII, IX, II, and fibrinogen is recognized as a risk factor of venous thromboembolism (VTE). These factors are cumulatively explored by the activated partial thromboplastin time (APTT). To test the hypothesis that a short APTT increases the risk of VTE, a case-control study was carried out in 605 patients referred for thrombophilia testing after documented VTE and in 1290 controls. Median APTT ratio (coagulation time of test-to-reference plasma) values were 0.97 (range: 0.75-1.41) for patients and 1.00 (range: 0.72-1.33) for controls (P < .001). In patients who had an APTT ratio smaller than the fifth percentile of the distribution in controls, the odds ratio (OR) for VTE was 2.4 (95% confidence interval [CI]: 1.7-3.6) and was independent of inherited thrombophilic abnormalities. Further statistical analyses in 193 patients and 259 controls for whom factor VIII (FVIII) levels were available showed a decrease of the OR from 2.7 (95% CI: 1.4-5.3) to 2.1 (95% CI: 1.0-4.2), indicating that the risk was only partially mediated by high FVIII levels. In conclusion, hypercoagulability detected by a shortened APTT is independently associated with VTE. This inexpensive and simple test should be considered in the evaluation of the risk of VTE. PMID- 15297316 TI - Anaplastic large cell lymphomas lack the expression of T-cell receptor molecules or molecules of proximal T-cell receptor signaling. AB - Anaplastic large cell lymphoma (ALCL) designates a heterogeneous group of CD30(+) (systemic or primary cutaneous) peripheral T-cell lymphomas (PTCLs). A subgroup of systemic ALCL is transformed by anaplastic lymphoma kinase (ALK). We compared 24 ALK(+), 15 ALK(-) systemic, and 7 cutaneous ALCLs with 29 nonanaplastic PTCLs in terms of T-cell receptor (TCR) rearrangements, expression of TCRs and TCR associated molecules (CD3, ZAP-70 [zeta-associated protein 70]). Despite their frequent clonal rearrangement for TCRbeta, only 2 (4%) of 47 ALCLs expressed TCRbeta protein, whereas TCRs were detected on 27 of 29 nonanaplastic PTCLs. Moreover, both TCRbeta(+) ALCLs lacked CD3 and ZAP-70 (ie, molecules indispensable for the transduction of cognate TCR signals). Defective expression of TCRs is a common characteristic of all types of ALCL, which may contribute to the dysregulation of intracellular signaling pathways controlling T-cell activation and survival. This molecular hallmark of ALCL is analogous to defective immunoglobulin expression distinguishing Hodgkin lymphoma from other B cell lymphomas. PMID- 15297317 TI - Antiproliferative activity of a humanized anti-CD74 monoclonal antibody, hLL1, on B-cell malignancies. AB - The humanized anti-CD74 monoclonal antibody (mAb) hLL1 is under evaluation as a therapeutic agent. The effects of hLL1-at times in comparison with the CD20 mAb rituximab-were assessed on non-Hodgkin lymphoma (NHL) and multiple myeloma (MM) cell lines and in tumor-bearing SCID mice. In vitro, hLL1 caused growth inhibition and induction of apoptosis in B-cell lines when cross-linked with an antihuman immunoglobulin G (IgG) second antibody. The sensitivity profile of the cell lines was different for hLL1 and rituximab, and antiproliferative activity was augmented when the 2 mAbs were combined. Unlike rituximab, hLL1 did not induce antibody-dependent cellular cytotoxicity or complement-mediated cytotoxicity. In xenograft models of NHL and MM, treatment with hLL1 yielded significant survival benefits without cross-linking agents. Efficacy was greater in the MM model, in which median survival time was increased more than 4.5-fold. Thus, hLL1 has therapeutic potential as a naked mAb for B-cell malignancies because of high antigen expression on malignant cells, specifically MM, with limited expression on normal tissue, and because of its antiproliferative activity. Further, hLL1 may be a therapeutic candidate for rituximab-resistant disease because the 2 antibodies apparently act through distinct mechanisms and exhibit different expression and sensitivity profiles, and activity can be augmented when the mAbs are combined. PMID- 15297318 TI - Absence of clinical GVHD and the in vivo induction of regulatory T cells after transplantation of facilitating cells. AB - Graft-versus-host disease (GVHD) and failure of engraftment limit clinical bone marrow transplantation (BMT) to patients with closely matched donors. Engraftment failure of purified allogeneic hematopoietic stem cells (HSCs) has been decreased in various BMT models by including donor BM-derived CD8(+)/alphabetagammadeltaTCR(-) facilitating cells (FCs) or CD8(+)/alphabetaTCR(+) T cells in the BM inoculum. To aggressively investigate the GVHD potential of these donor CD8(+) populations, a purified cell model of lethal GVHD was established in a murine semiallogeneic parent --> F(1) combination. Lethally irradiated recipients were reconstituted with purified donor HSCs alone or in combination with splenic T cells (T(SP)), BM-derived T cells (T(BM)), or the FC population. In marked contrast to the lethal GVHD present in recipients of HSCs plus T(SP) or CD8(+) T(BM), recipients of donor HSC+FC inocula did not exhibit significant clinical or histologic evidence of GVHD. Instead, HSC+FC recipients were characterized by increased splenocyte expression of transforming growth factor-beta (TGF-beta) and the induction of the regulatory T-cell genes CTLA4, GITR, and FoxP3. These findings suggest that the FCs, which express a unique FCp33-TCRbeta heterodimer in place of alphabetaTCR, permits HSC alloengraftment and prevents GVHD through the novel approach of regulatory T-cell induction in vivo. PMID- 15297319 TI - CLEVER-1 mediates lymphocyte transmigration through vascular and lymphatic endothelium. AB - Common lymphatic endothelial and vascular endothelial receptor-1 (CLEVER-1; also known as stabilin-1 or FEEL-1) is a large multifunctional glycoprotein implicated in scavenging, angiogenesis, and cell adhesion. Here we studied the function of human CLEVER-1 in leukocyte trafficking. Lymphatic vessels expressed CLEVER-1 constitutively in skin in vivo, whereas on vascular endothelium it appeared only upon inflammation. On isolated vascular endothelial cells, CLEVER-1 supported rolling and transmigration of peripheral blood mononuclear cells (PBMCs) under physiologically relevant laminar shear stress. Intriguingly, CLEVER-1 also mediated transmigration of leukocytes through cultured lymphatic endothelium under static conditions. Thus, synthesis of CLEVER-1 is differentially regulated on the 2 anatomically distinct vascular beds, and CLEVER-1 mediates the transmigration step of the leukocyte traffic in both of them. Notably, CLEVER-1 is the first adhesion molecule shown to be involved in the PBMC transmigration through the lymphatic arm of the immune system. PMID- 15297320 TI - How protective is the working time directive? PMID- 15297321 TI - How many eggs? PMID- 15297322 TI - Policing access to primary care with identity cards. PMID- 15297323 TI - Topical NSAIDs in osteoarthritis. PMID- 15297324 TI - Preventing malaria in UK travellers. PMID- 15297325 TI - Secret US report surfaces on antidepressants in children. PMID- 15297328 TI - Chief medical officer warns that many blood transfusions are unnecessary. PMID- 15297329 TI - Man wins battle to keep receiving life support. PMID- 15297331 TI - Sudanese army shuns UN resolution as humanitarian crisis continues. PMID- 15297332 TI - Consumers' Association voices concern at over the counter statins. PMID- 15297333 TI - Baby milk manufacturers agree out of court settlement. PMID- 15297334 TI - Over the limit? PMID- 15297336 TI - Australia's free trade deal with US hangs in balance on drugs. PMID- 15297338 TI - Acquired haemophilia A may be associated with clopidogrel. PMID- 15297339 TI - New perspectives--approaches to medical education at four new UK medical schools. PMID- 15297340 TI - Promises and delivery--a research imperative for new approaches to medical education. PMID- 15297341 TI - The ethics of medical education. PMID- 15297342 TI - Patients in medical education and research. PMID- 15297344 TI - Neurocardiogenic syncope. PMID- 15297346 TI - Rehabilitation after burn injury. PMID- 15297347 TI - Eligibility of overseas visitors and people of uncertain residential status for NHS treatment. PMID- 15297349 TI - Investigation into GPs with high patient mortality: monitoring death rates will become increasingly complex. PMID- 15297350 TI - Investigation into GPs with high patient mortality: situation is even more complicated. PMID- 15297353 TI - Guidance has high priority in interventional procedures: objective evidence needs to be provided. PMID- 15297354 TI - Scientific articles have hardly changed in 50 years: plain English and minimal Latin may explain readability of 1950s paper... PMID- 15297355 TI - Scientific articles have hardly changed in 50 years: ...and "scientific rigour" may be to blame for current dullness... PMID- 15297356 TI - Scientific articles have hardly changed in 50 years: ...but Asher was asking why medical journals were so dull back then. PMID- 15297357 TI - Japan Tobacco Incorporated has found a new way of promoting tobacco in motor sports. PMID- 15297358 TI - Role of poverty must be emphasised. PMID- 15297365 TI - Human cortical dysplasia and epilepsy: an ontogenetic hypothesis based on volumetric MRI and NeuN neuronal density and size measurements. AB - In epilepsy patients with cortical dysplasia (CD), this study determined the probable ontogenetic timing of pathogenesis based on the number, location and appearance of neurons. Magnetic resonance imaging (MRI) determined gray and white matter volumes of affected and non-affected cerebral hemispheres, and gray and white matter neuronal-nuclear protein (NeuN) densities and sizes were assessed in epilepsy surgery patients (0.2-38 years) with CD (n = 25) and non-CD etiologies (n = 14), and compared with autopsy cases (n = 13; 0-33 years). Pathology group, seizure type and age at surgery were compared against MRI and NeuN data. CD patients demonstrated increased MRI cerebral (3%) and gray matter (8%) volumes of the affected compared with non-affected cerebral hemisphere, and increased layer 1 (131%), upper cortical (9-23%) and white matter (28-77%) NeuN densities compared with autopsy cases. Non-CD cases showed decreased cerebral volumes of the affected hemisphere (14-18%) without changes in NeuN densities. Compared with autopsy cases, in CD and non-CD patients, cortical neurons were hypertrophied. Patients with a history of infantile spasms had a 40% increase in the size of layer 1 neurons compared with cases without spasms. By age, regardless of pathology group, there were logarithmic increases in MRI cerebral and white matter volumes, logarithmic increases in the size of lower gray and superficial white matter neurons, and logarithmic decreases in gray and white matter neuronal densities. These results support the concept that there were more neurons than expected in layer 1, gray, and white matter of CD patients compared with non-CD and autopsy cases. In addition, the location and appearance of neurons are consistent with the hypothesis that CD is the consequence of abnormalities occurring late in corticoneurogenesis that involve excessive neurogenesis with retention of pre-plate cells in the molecular layer and subplate regions. PMID- 15297366 TI - Developmental changes in human cerebral functional organization for word generation. AB - A fundamental issue in cognitive neuroscience is the nature of developmental changes in human cerebral functional organization for higher cognitive functions. Event-related functional magnetic resonance imaging was used to measure developmental changes in the functional neuroanatomy subserving controlled lexical association. First, brain regions showing significant differences in activity between school-age children and young adults, despite equivalent task performance, were identified. Then, activity in these regions was more fully characterized in individuals spanning the ages of 7-32 years old. Cross-sectional and regression analyses showed systematic increases and decreases in levels of activity over age, by region. Age-related increases in activity were primarily newly recruited, later-stage processing regions, such as in left frontal and left parietal cortex. Decreases, on the other hand, were all positive activations that attenuated with age and were found across a wider neuroanatomical range, including earlier processing regions such as bilateral extrastriate cortex. The hemodynamic magnitude, neuroanatomical location and maturational timecourse of these progressive and regressive changes have implications for models of the developing specialization in human cerebral functional organization. PMID- 15297367 TI - Representation of interaural temporal information from left and right auditory space in the human planum temporale and inferior parietal lobe. AB - The localization of low-frequency sounds mainly relies on the processing of microsecond temporal disparities between the ears, since low frequencies produce little or no interaural energy differences. The overall auditory cortical response to low-frequency sounds is largely symmetrical between the two hemispheres, even when the sounds are lateralized. However, the effects of unilateral lesions in the superior temporal cortex suggest that the spatial information mediated by lateralized sounds is distributed asymmetrically across the hemispheres. This paper describes a functional magnetic resonance imaging experiment, which shows that the interaural temporal processing of lateralized sounds produces an enhanced response in the contralateral planum temporale (PT). The response is stronger and extends further into adjacent regions of the inferior parietal lobe (IPL) when the sound is moving than when it is stationary. This suggests that the interaural temporal information mediated by lateralized sounds is projected along a posterior pathway comprising the PT and IPL of the respective contralateral hemisphere. The differential responses to moving sounds further revealed that the left hemisphere responded predominantly to sound movement within the right hemifield, whereas the right hemisphere responded to sound movement in both hemifields. This rightward asymmetry parallels the asymmetry associated with the allocation of visuo-spatial attention and may underlie unilateral auditory neglect phenomena. PMID- 15297368 TI - Anti-angiogenic activity of inositol hexaphosphate (IP6). AB - A significant anticancer activity of the naturally occurring carbohydrate inositol hexaphosphate (IP(6)) has been reported against numerous cancer models. Since tumors require angiogenesis for growth and metastasis, we hypothesize that IP(6) reduces tumor growth by inhibiting angiogenesis. Because angiogenesis depends on the interaction between endothelial and tumor cells, we investigated the effect of IP(6) on both. IP(6) inhibited the proliferation and induced the differentiation of endothelial cells in vitro; the growth of bovine aortic endothelial cells (BAECs) evaluated by MTT proliferation assay was inhibited in a dose-dependent manner (IC(50) = 0.74 mM). The combination of IP(6) and vasostatin, a calreticulin fragment with anti-angiogenic activity, was synergistically superior in growth inhibition than either compound. IP(6) inhibited human umbilical vein endothelial cell (HUVEC) tube formation (in vitro capillary differentiation) on a reconstituted extracellular matrix, Matrigel, and disrupted pre-formed tubes. IP(6) significantly reduced basic fibroblast growth factor (bFGF)-induced vessel formation (P < 0.01) in vivo in Matrigel plug assay. Exposure of HepG2, a human hepatoma cell line, to IP(6) for 8 h, resulted in a dose-dependent decrease in the mRNA levels of vascular endothelial growth factor (VEGF), as assessed by RT-PCR. IP(6) treatment of HepG2 cells for 24 h also significantly reduced the VEGF protein levels in conditioned medium, in a concentration-dependent manner (P = 0.012). Thus, IP(6) has an inhibitory effect on induced angiogenesis. PMID- 15297369 TI - Azoxymethane-induced pre-adipocyte factor 1 (Pref-1) functions as a differentiation inhibitor in colonic epithelial cells. AB - Inbred mice differ dramatically in their sensitivity to the colon carcinogen, azoxymethane (AOM). Identifying genes associated with this differential susceptibility in mice may ultimately reveal molecular mechanisms responsible for colon carcinogenesis. A cDNA array approach was taken to study gene expression changes induced by AOM in the colons of sensitive (A/J) and resistant (AKR) mice. Among the genes represented on the array, pre-adipocyte factor 1 (Pref-1), associated previously with suppression of adipocyte differentiation, was induced specifically by AOM in the distal colons of sensitive A/J mice (5.4-fold). Reverse transcription-PCR followed by sequence analysis revealed the presence of four alternative splice variants of Pref-1 mRNA in the colon. The potential significance of Pref-1 in colon tumorigenesis was explored in colon cancer cells infected with a retroviral construct containing the major splice variant. Over expression of Pref-1A in HT-29 cells led to a marked resistance to butyrate induced differentiation and growth inhibition. Our data indicate that Pref-1, a protein that suppresses differentiation and promotes colonocyte growth, may account in part for the sensitivity of A/J mice to AOM-induced carcinogenesis. In addition, detection of Pref-1 in a human colon tumor cell line suggests that it may also participate in human colon tumorigenesis. PMID- 15297370 TI - Tobacco smoke induces CYP1B1 in the aerodigestive tract. AB - Several members of the P450 family, including cytochrome P450 1B1 (CYP1B1), can convert tobacco smoke (TS) procarcinogens, including benzo[a]pyrene (B[a]P), to carcinogenic intermediates. In this study we investigated the effects of TS condensate and B[a]P on the expression of CYP1B1 in vitro and in vivo. CYP1B1 mRNA and protein were induced by both TS condensate and B[a]P in cell lines derived from the human aerodigestive tract. Treatment with TS condensate stimulated binding of the aryl hydrocarbon receptor (AhR) to an oligonucleotide containing a canonical xenobiotic response element (XRE) site and induced XRE luciferase activity. These findings are consistent with prior evidence that polycyclic aromatic hydrocarbons, known ligands of the AhR, stimulate CYP1B1 transcription by an XRE-dependent mechanism. To determine whether these in vitro findings applied in vivo, both murine and human studies were carried out. Short term exposure to TS induced CYP1B1 in the tongue, esophagus, lung and colon of experimental mice. In contrast, CYP1B1 was not induced by TS in the aorta of these mice. Levels of CYP1B1 mRNA were also elevated in the bronchial mucosa of human tobacco smokers versus never smokers (P < 0.05). Taken together, these results support a role for CYP1B1 in TS-induced carcinogenesis in the aerodigestive tract. PMID- 15297371 TI - FTY720 induces apoptosis of human hepatoma cell lines through PI3-K-mediated Akt dephosphorylation. AB - Our aim was to study the anticancer effect of the novel immunomodulator FTY720 in vitro and in vivo by investigation of cell cycle entry, cell cycle regulation, cell survival and apoptosis pathways. Three hepatoma cell lines with different p53 statuses (HepG2, Huh-7 and Hep3B) and one non-tumorigenic immortalized liver cell line (MIHA) were used for an in vitro study. The in vivo effects of FTY720 were evaluated in a nude mouse tumor model. Cell cycle distribution and cell cycle regulator proteins p27(Kip1) and cyclin D1, together with the PI3-K/Akt pathway, mitogen-activated protein kinases and cleaved caspase-3 and caspase-9, were evaluated. FTY720 selectively induced cell apoptosis in hepatoma cell lines with overexpression of cleaved caspase-3 and caspase-9, but the same phenomena were not found in MIHA cells. FTY720 induced Akt dephosphorylation at Ser473 mediated by phosphoinositide 3-kinase (PI3-K) inhibition. Dephosphorylation led to down-regulation of p42/p44 and dephosphorylation of Forkhead transcription factor and GSK-3beta and, subsequently, up-regulation of p27(Kip1) and down regulation of cyclin D1. In our in vivo model FTY720 induced apoptosis of tumor cells by down-regulation of the Akt pathway. FTY720 suppressed tumor growth without notable side-effects in normal liver. In conclusion, FTY720 is a novel anticancer agent that induces apoptosis of hepatoma cell lines both in vitro and in vivo through PI3-K-mediated Akt dephosphorylation in a p53-independent manner. PMID- 15297372 TI - Effects of a combination of docosahexaenoic acid and 1,4-phenylene bis(methylene) selenocyanate on cyclooxygenase 2, inducible nitric oxide synthase and beta catenin pathways in colon cancer cells. AB - Epidemiological and preclinical studies suggest that diets that are rich in n-3 polyunsaturated fatty acids (PUFAs) and selenium (Se) reduce the risk of colon cancer. Studies conducted in our laboratory have indicated that synthetic organoselenium 1,4-phenylene bis(methylene) selenocyanate (p-XSC) is less toxic and more effective than inorganic Se and selenomethionine, the major Se compound in natural selenium yeast. Through cDNA microarray analysis, we have demonstrated earlier that the n-3 PUFA docosahexaenoic acid (DHA), modulated more than one signaling pathway by altering several genes involved in colon cancer growth. There is increasing interest in the use of combinations of low doses of chemopreventive agents that differ in their specific modes of action as this approach can minimize toxicity and increase efficacy in model assays. In the present study we assessed the efficacy of DHA and p-XSC individually and in combination at low doses in CaCo-2 colon cancer cells, using cell growth inhibition and apoptosis as measures of chemopreventive efficacy. On the basis of western blot and RT-PCR analysis, we also determined the effects of DHA and p-XSC on the levels of expression of cyclooxygenase-2, inducible nitric oxide synthase, cyclin D1, beta-catenin and nuclear factor kappaB, all of which presumably participate in colon carcinogenesis. A 48 h incubation of CaCo-2 cells with 5 microM each DHA or p-XSC induced cell growth inhibition and apoptosis and altered the expression of the above molecular parameters. Interestingly, the modulation of these cellular and molecular parameters was more pronounced in cells treated with low doses of DHA and p-XSC (2.5 microM each) in combination than in cells treated with these agents individually at higher concentrations (5.0 microM each). These findings are viewed as highly significant since they will provide the basis for the development of combinations of low dose regimens of DHA and p XSC in preclinical models against colon carcinogenesis and, ultimately, in human clinical trials. PMID- 15297373 TI - AMP-activated protein kinase activity is required for vanadate-induced hypoxia inducible factor 1alpha expression in DU145 cells. AB - Hypoxia-inducible factor 1 (HIF-1), a pivotal transcription factor composed of HIF-1alpha and HIF-1beta subunits, plays a major role in tumor progression by activating a number of genes critically involved in adaptation to hypoxia. HIF-1 is also induced by several carcinogenic metals. Vanadate, an environmental toxic metal, is considered as a potent inducer of tumors in animals and is reported to activate HIF-1 activity. However, the involved mechanisms are poorly understood. In the present study, we have examined the biochemical mechanisms of the vanadate induced HIF-1 activation in cancer cells by primarily focusing on the role of AMP activated protein kinase (AMPK), which plays an essential role as an energy sensor under ATP-deprived conditions. We demonstrate that AMPK was rapidly activated in response to vanadate in DU145 human prostate carcinoma, and that its activation preceded HIF-1alpha expression. Under this condition, inhibition of AMPK by a pharmacological and molecular approach dramatically abolished the vanadate-induced HIF-1alpha expression as well as HIF-1-mediated physiological responses. Phosphatidylinositol-3 kinase/Akt/mammalian target of rapamycin signaling was also involved in vanadate-induced HIF-1alpha expression, but it was independent of AMPK signaling pathway. Moreover, we demonstrate a role of reactive oxygen species as an upstream signal for these two pathways. These results suggest that AMPK is a novel and critical component of HIF-1 regulation, further implying its involvement in vanadate-induced carcinogenesis. PMID- 15297374 TI - Association of human connexin40 gene polymorphisms with atrial vulnerability as a risk factor for idiopathic atrial fibrillation. AB - Alterations in distribution, density, and properties of cardiac gap junctions, which mediate electrical coupling of cardiomyocytes, are considered potentially arrhythmogenic. We recently reported 2 linked polymorphisms within regulatory regions of the gene for the atrial gap junction protein connexin40 (Cx40) at nucleotides -44 (G-->A) and +71 (A-->G), which were associated with familial atrial standstill. The present study examined whether these Cx40 polymorphisms were associated with increased atrial vulnerability in vivo and arrhythmia susceptibility. In 30 subjects without structural heart disease, of whom 14 had documented sporadic paroxysmal atrial fibrillation (AF) and 16 had no AF history, inducibility of AF was assessed using an increasingly aggressive atrial stimulation protocol. Coefficient of spatial dispersion of refractoriness (CD) was calculated. CD was defined as the SD of 12 local mean fibrillatory intervals recorded at right atrial sites, expressed as a percentage of the overall mean fibrillatory interval. Cx40 genotypes were determined by direct DNA sequencing. Subjects were stratified according to normal or increased CD with a cutoff value of 3.0, because CD >3.0 was previously shown to be strongly associated with enhanced atrial vulnerability. The prevalence of the minor Cx40 allele (-44A) and -44AA genotype was significantly higher in subjects with increased dispersion (n=13) compared with those with CD < or =3.0 (n=17; P=0.00046 and P=0.025; odds ratios of 6.7 and 7.4) and a control population (n=253; P=0.00002 and P=3.90x10( 7)). Carriers of -44AA genotype had a significantly higher CD compared with those with -44GG genotype (6.37+/-1.21 versus 2.38+/-0.39, P=0.018), whereas heterozygotes had intermediate values (3.95+/-1.38, NS). All subjects with increased CD had a history of idiopathic AF compared with only 1 subject with normal CD. The -44A allele and -44AA genotype were significantly more frequent in subjects with prior AF than in those without (P=0.0019 and P=0.031; odds ratios 5.3 and 6.2). This study provides strong evidence linking Cx40 polymorphisms to enhanced atrial vulnerability and increased risk of AF. The full text of this article is available online at http://circres.ahajournals.org. PMID- 15297375 TI - Ets-1 stimulates platelet-derived growth factor A-chain gene transcription and vascular smooth muscle cell growth via cooperative interactions with Sp1. AB - The platelet-derived growth factor (PDGF) family of ligands (composed of A-, B-, C-, and D-chains), potent mitogens, and chemoattractants for cells of mesenchymal origin has been implicated in numerous vascular pathologies involving smooth muscle cell (SMC) hyperplasia. Understanding the molecular mechanisms mediating PDGF transcription would provide new insights into strategies to control PDGF dependent pathophysiologic processes. We demonstrated previously that PDGF-A expression is under the positive regulatory influence of Sp1, Sp3, and Egr-1 and is negatively controlled by GCF2, NF-1(X), and WT-1. In this article, we demonstrate that Ets-1 induces PDGF-A expression in primary rat aortic SMCs at the level of transcription and mRNA expression. Electrophoretic mobility shift, supershift, and mutational analyses revealed a functional role for the ( 555)TTCC(-552) motif in the PDGF-A promoter that binds endogenous Ets-1. Chromatin immunoprecipitation analysis showed the interaction of endogenous and exogenous Ets-1 or glutathione S-transferase-tagged Ets-1, bearing only the DNA binding domain with the authentic PDGF-A promoter. Conversely, dominant-negative mutant of Ets-1 blocked the promoter interaction of endogenous Ets-1. Overexpression of Ets-1 but not the mutant form of Ets-1 activates the PDGF-A promoter cooperatively with Sp1. Sp1, which interacts with Ets-1, failed to induce PDGF-A promoter-dependent expression if the promoter contained a site specific mutation in this novel Ets-binding site. Small interfering RNA to Ets-1 and Sp1 blocked PDGF-BB- and serum-inducible PDGF-A expression. SMC growth was stimulated by Ets-1 and Sp1 separately and further increased by both factors together. Ets-1-inducible mitogenesis is blocked by antibodies neutralizing PDGF A and involves activation of the PDGF alpha-receptor, which binds PDGF-A. These findings identify a functional cis-acting element for Ets-1 in the PDGF-A promoter and demonstrate that Sp1 and Ets-1 cooperatively activate PDGF-A transcription in vascular SMCs. PMID- 15297376 TI - Targeted disruption of hesr2 results in atrioventricular valve anomalies that lead to heart dysfunction. AB - Genes involved in the Notch signaling pathway have been shown to be critical regulators of cardiovascular development. In vitro studies have revealed that the Notch signaling pathway directly regulates transcription of hairy and enhancer of split-related (hesr) genes, encoding basic helix-loop-helix transcription factors. To assess the functional role of hesr genes in cardiovascular development, we generated mice with a targeted disruption of the hesr2 gene and used echocardiography to analyze heart function of the mutant mice. In the early postnatal period, a majority of hesr2 homozygous mice die as a result of congestive heart failure accompanied by pronounced heart enlargement. Transthoracic echocardiography on 5-day-old homozygous mice revealed tricuspid and mitral valve regurgitation and a dilated left ventricular chamber with markedly diminished fractional shortening of the left ventricle. The hemodynamic anomalies were accompanied by morphological changes, such as dysplastic atrioventricular (AV) valves, a perimembranous ventricular septal defect, and a secundum atrial septal defect. AV valve regurgitations attributable to dysplasia of the AV valves were most likely responsible for the heart dysfunction in hesr2 homozygous mice. These observations indicate that the Notch signaling target hesr2 plays an important role in the formation and function of the AV valves. In addition, hesr2 activity may be important for proper development of cardiomyocytes, thereby assuring normal left ventricular contractility. Because of the unique spectrum of cardiac anomalies expressed by hesr2-null mice, they represent a useful model system for elucidating the genetic basis of heart dysfunction. PMID- 15297377 TI - Overexpression of urokinase by macrophages or deficiency of plasminogen activator inhibitor type 1 causes cardiac fibrosis in mice. AB - Several studies implicate elevated matrix metalloproteinase activity as a cause of cardiac fibrosis. However, it is unknown whether other proteases can also initiate cardiac fibrosis. Because absence of urokinase plasminogen activator (uPA) prevents development of cardiac fibrosis after experimental myocardial infarction in mice, we hypothesized that elevated activity of uPA or deficiency of the uPA inhibitor plasminogen activator inhibitor-1 (PAI-1) might cause cardiac fibrosis. We used mice with scavenger-receptor (SR)-directed, macrophage targeted uPA overexpression (SR-uPA+/0 mice) and PAI-1 null mice to test these hypotheses. Our studies revealed that SR-uPA+/0 mice developed cardiac fibrosis beginning between 5 and 10 weeks of age. Fibrosis was preceded by cardiac macrophage accumulation, implicating uPA-secreting macrophages as important contributors to development of fibrosis. A key role for uPA-secreting macrophages in development of cardiac fibrosis was supported by experiments in which recipients of bone marrow transplants from SR-uPA+/0 donors but not nontransgenic donors developed cardiac macrophage accumulation and fibrosis. SR-uPA+/0 mice and recipients of SR-uPA+/0 bone marrow had neither macrophage accumulation nor fibrosis in other major organs despite the presence of higher levels of uPA in these organs than in hearts. PAI-1 null mice but not congenic, age-matched controls also developed macrophage accumulation and fibrosis in hearts but not in other organs. We conclude: (1) either elevated macrophage uPA expression or PAI-1 deficiency is sufficient to cause cardiac macrophage accumulation and fibrosis; (2) macrophages are important contributors to the development of cardiac fibrosis; and (3) the heart is particularly sensitive to the effects of excess uPA activity. PMID- 15297378 TI - Rho kinase-induced nuclear translocation of ERK1/ERK2 in smooth muscle cell mitogenesis caused by serotonin. AB - There is now considerable evidence supporting a mitogenic action of serotonin (5 HT) on vascular smooth muscle cells (SMC) that might participate in pulmonary hypertension (PH). Our previous studies have demonstrated that 5-HT-induced proliferation depends on the generation of reactive oxygen species and activation of extracellular signal-regulated kinase (ERK) 1/ERK2. Activation of Rho kinase (ROCK) in SMC also may be important in PH. We undertook the present study to assess the role of Rho A/ROCK and its possible relation to ERK1/ERK2 in 5-HT induced pulmonary artery SMC proliferation. We found that this stimulation of SMC proliferation requires Rho A/ROCK as inhibition with Y27632, a ROCK inhibitor, or dominant negative (DN) mutant Rho A blocks 5-HT-induced proliferation, cyclin D1 expression, phosphorylation of Elk, and the DNA binding of transcription factors, Egr-1 and GATA-4. 5-HT activated ROCK, and the activation was blocked by GR 55562 and GR127935, 5-HT 1B/1D receptor antagonists, but not by serotonin transport (SERT) inhibitors. Activation of Rho kinase by 5-HT was independent of activation of ERK1/ERK2, and 5-HT activated ERK1/ERK2 independently of ROCK. Treatment of SMC with Y27632 and expression of DNRho A in cells blocked translocation of ERK1/ERK2 to the cellular nucleus. Depolymerization of actin with cytochalasin D (CD) and latrunculin B (latB) failed to block the translocation of ERK, suggesting that the actin cytoskeleton does not participate in the translocation. The studies show for the first time to our knowledge combinational action of SERT and a 5-HT receptor in SMC growth and Rho A/ROCK participation in 5-HT receptor 1B/1D-mediated mitogenesis of vascular SMCs through an effect on cytoplasmic to nuclear translocation of ERK1/ERK2. PMID- 15297379 TI - Lineage and morphogenetic analysis of the cardiac valves. AB - We used a genetic lineage-labeling system to establish the material contributions of the progeny of 3 specific cell types to the cardiac valves. Thus, we labeled irreversibly the myocardial (alphaMHC-Cre+), endocardial (Tie2-Cre+), and neural crest (Wnt1-Cre+) cells during development and assessed their eventual contribution to the definitive valvar complexes. The leaflets and tendinous cords of the mitral and tricuspid valves, the atrioventricular fibrous continuity, and the leaflets of the outflow tract valves were all found to be generated from mesenchyme derived from the endocardium, with no substantial contribution from cells of the myocardial and neural crest lineages. Analysis of chicken-quail chimeras revealed absence of any substantial contribution from proepicardially derived cells. Molecular and morphogenetic analysis revealed several new aspects of atrioventricular valvar formation. Marked similarities are seen during the formation of the mural leaflets of the mitral and tricuspid valves. These leaflets form by protrusion and growth of a sheet of atrioventricular myocardium into the ventricular lumen, with subsequent formation of valvar mesenchyme on its surface rather than by delamination of lateral cushions from the ventricular myocardial wall. The myocardial layer is subsequently removed by the process of apoptosis. In contrast, the aortic leaflet of the mitral valve, the septal leaflet of the tricuspid valve, and the atrioventricular fibrous continuity between these valves develop from the mesenchyme of the inferior and superior atrioventricular cushions. The tricuspid septal leaflet then delaminates from the muscular ventricular septum late in development. PMID- 15297380 TI - Critical roles for the Fas/Fas ligand system in postinfarction ventricular remodeling and heart failure. AB - In myocardial infarction (MI), granulation tissue cells disappear via apoptosis to complete a final scarring with scanty cells. Blockade of this apoptosis was reported to improve post-MI ventricular remodeling and heart failure. However, the molecular biological mechanisms for the apoptosis are unknown. Fas and Fas ligand were overexpressed in the granulation tissue at the subacute stage of MI (1 week after MI) in mice, where apoptosis frequently occurred. In mice lacking functioning Fas (lpr strain) and in those lacking Fas ligand (gld strain), apoptotic rate of granulation tissue cells was significantly fewer compared with that of genetically controlled mice, and post-MI ventricular remodeling and dysfunction were greatly attenuated. Mice were transfected with adenovirus encoding soluble Fas (sFas), a competitive inhibitor of Fas ligand, on the third day of MI. The treatment resulted in suppression of granulation tissue cell apoptosis and produced a thick, cell-rich infarct scar containing rich vessels and bundles of smooth muscle cells with a contractile phenotype at the chronic stage (4 weeks after MI). This accompanied not only alleviation of heart failure but also survival improvement. However, the sFas gene delivery during scar tissue phase was ineffective, suggesting that beneficial effects of the sFas gene therapy owes to inhibition of granulation tissue cell apoptosis. The Fas/Fas ligand interaction plays a critical role for granulation tissue cell apoptosis after MI. Blockade of this apoptosis by interfering with the Fas/Fas ligand interaction may become one of the therapeutic strategies against chronic heart failure after large MI. PMID- 15297381 TI - The small heart mutation reveals novel roles of Na+/K+-ATPase in maintaining ventricular cardiomyocyte morphology and viability in zebrafish. AB - Forward genetic screens in zebrafish have been used to identify mutations in genes with important roles in organogenesis. One of these mutants, small heart, develops a diminutive and severely malformed heart and multiple developmental defects of the brain, ears, eyes, and kidneys. Using a positional cloning approach, we identify that the mutant gene encodes the zebrafish Na+/K+-ATPase alpha1B1 protein. Disruption of Na+/K+-ATPase alpha1B1 function via morpholino "knockdown" or pharmacological inhibition with ouabain phenocopies the mutant phenotype, in a dose-dependent manner. Heterozygosity for the mutation sensitizes embryos to ouabain treatment. Our findings present novel genetic and morphological details on the function of the Na+/K+-ATPase alpha1B1 in early cardiac morphogenesis and the pathogenesis of the small heart malformation. We demonstrate that the reduced size of the mutant heart is caused by dysmorphic ventricular cardiomyocytes and an increase in ventricular cardiomyocyte apoptosis. This study provides a new insight that Na+/K+-ATPase alpha1B1 is required for maintaining ventricular cardiomyocyte morphology and viability. PMID- 15297382 TI - Generation of functional culture-derived platelets from CD34+ progenitor cells to study transgenes in the platelet environment. AB - The possibility of evaluating the function of transgenes in platelets requires the generation of platelets from nucleated progenitor cells in vitro. In this article, we provide effective culture conditions for generating functional culture-derived (CD) human and mouse platelets from CD34(+) progenitor cells that allow expression of any foreign protein of interest. We have evolved an effective cytokine cocktail (thrombopoietin, stem cell factor, interleukin [IL]-1beta, IL 6) that induces a high yield of CD platelets and optimal shedding from cultivated megakaryocytes generated from CD34(+) progenitor cells. CD platelets showed similar functional and morphological characteristics compared with isolated blood platelets, including surface expression of platelet antigens (CD41, CD42, CD62P), aggregation, release of granule constituents (P-selectin, platelet factor 4, serotonin). Moreover, transmission electron microscopy revealed the presence of typical alpha- and dense granules and dense tubular system in CD platelets. Additionally, we showed that stable transgene expression in CD platelets can be performed through infection of CD34(+) progenitor cells using adenoviral vectors. Thus, we describe a methodology that enables studying functional consequences of transgenes of interest in the natural environment of platelets that may impose substantial impact on potential future platelet research and therapeutic target evaluation. The full text of this article is available online at http://circres.ahajournals.org. PMID- 15297383 TI - Is modulation of sodium-calcium exchange a therapeutic option in heart failure? PMID- 15297384 TI - A new slice of pie? Estrogen regulation of plasminogen activator inhibitor-1. PMID- 15297385 TI - Protein glycation: a firm link to endothelial cell dysfunction. AB - The advanced glycation end products (AGEs) are a heterogeneous class of molecules, including the following main subgroups: bis(lysyl)imidazolium cross links, hydroimidazolones, 3-deoxyglucosone derivatives, and monolysyl adducts. AGEs are increased in diabetes, renal failure, and aging. Microvascular lesions correlate with the accumulation of AGEs, as demonstrated in diabetic retinopathy or renal glomerulosclerosis. On endothelial cells, ligation of receptor for AGE (RAGE) by AGEs induces the expression of cell adhesion molecules, tissue factor, cytokines such as interleukin-6, and monocyte chemoattractant protein-1. A chief means by which AGEs via RAGE exert their effects is by generation of reactive oxygen species, at least in part via stimulation of NADPH oxidase. Diabetes associated vascular dysfunction in vivo can be prevented by blockade of RAGE. Thus, agents that limit AGE formation, increase the catabolism of these species, or antagonize their binding to RAGE may provide new targets for vascular protection in diabetes. PMID- 15297386 TI - Examining intracellular organelle function using fluorescent probes: from animalcules to quantum dots. AB - Fluorescence microscopy imaging has become one of the most useful techniques to assess the activity of individual cells, subcellular trafficking of signals to and between organelles, and to appreciate how organelle function is regulated. The past 2 decades have seen a tremendous advance in the rational design and development in the nature and selectivity of probes to serve as reporters of the intracellular environment in live cells. These probes range from small organic fluorescent molecules to fluorescent biomolecules and photoproteins ingeniously engineered to follow signaling traffic, sense ionic and nonionic second messengers, and report various kinase activities. These probes, together with recent advances in imaging technology, have enabled significantly enhanced spatial and temporal resolution. This review summarizes some of these developments and their applications to assess intracellular organelle function. PMID- 15297387 TI - How do red blood cells dilate blood vessels?--Reply. PMID- 15297388 TI - High lactate dehydrogenase isoenzyme 1 in a patient with malignant germ cell tumor is attributable to aberrant methylation of the LDHA gene. PMID- 15297389 TI - The potential role of neutrophils in promoting the metastatic phenotype of tumors releasing interleukin-8. AB - In the last decade, several groups have shown a direct correlation between the inappropriate or ectopic release of interleukin (IL)-8 by tumor cells in vitro and their growth and metastatic potential using in vivo models of tumor growth. IL-8 is a potent neutrophil chemoattractant. Neutrophils, as "early responders" to wounds and infections, release enzymes to remodel the extracellular matrix of the tissues through which they migrate to reach the site of the wound or infection. It is proposed that the host's cellular response to IL-8 released by tumor cells enhances angiogenesis and contributes to tumor growth and progression. The activities released by the responding neutrophils could serve as enablers of tumor cell migration through the extracellular matrix, helping them enter the vasculature and journey to new, metastatic sites. The reactive oxygen species produced by neutrophilic oxidases to kill invading organisms have the potential to interact with tumor cells to attenuate their apoptotic cascade and increase their mutational rate. It is proposed that the increase in metastatic potential of tumors ectopically releasing IL-8 is, in part, attributable to their ability to attract neutrophils. Discussed here are possible mechanisms by which the neutrophils responding to ectopic IL-8 contribute to the in vivo growth, progression, and metastatic potential of tumor cells. Possible targets are also presented for the development of therapies to attenuate the effects of the ectopic IL-8 release by tumor cells. PMID- 15297390 TI - Biological relevance of adduct detection to the chemoprevention of cancer. AB - Adducts arise from the chemical modification of bases in DNA or amino acids in proteins by toxic chemicals. Many chemicals known to be carcinogenic in humans have been shown to form adducts or to cause oxidative damage to genomic DNA in model systems. Biomarkers of carcinogenesis reflect biological events that take place between exposure to external or endogenous carcinogens and the subsequent development of cancer. Therapeutic intervention for the purpose of cancer chemoprevention may modify these biomarkers. In this article, the potential efficacy of DNA adducts as biomarkers of carcinogenesis and chemoprevention is discussed using criteria defined for phases of biomarker development. The sensitivity of adduct detection in histologically normal tissue offers opportunities for the early detection of carcinogenesis. Extensive evidence for aflatoxin B(1) adducts as biomarkers of risk and progression of hepatic carcinogenesis and for oxidative DNA adducts as biomarkers of the development of prostate carcinogenesis is reviewed together with the clinical trials measuring these adducts as biomarkers of the efficacy of chemoprevention. Favorable modification of oxidative DNA adducts by dietary intervention and chemoprevention has been demonstrated in preclinical and clinical studies. Protein adducts and DNA adducts in blood constituents or urine may act as useful surrogates for the target organ. Additional information regarding reliability, reproducibility, specificity, and confounding variables are required at the clinical level to validate adducts as suitable biomarkers of chemoprevention. "We do not administer antihypertensive drugs to patients in clinical trials without checking their blood pressure, so why should we give antioxidants without checking that they have decreased oxidant status. PMID- 15297391 TI - Phase I and pharmacokinetic study of the oral fluoropyrimidine S-1 on a once daily-for-28-day schedule in patients with advanced malignancies. AB - PURPOSE: The oral fluoropyrimidine S-1, which consists of a mixture of a 5 fluorouracil (5-FU) prodrug (tegafur), a dihydropyrimidine dehydrogenase inhibitor [5-chloro-2,4-dihydroxypyrimidine (CDHP)], and an inhibitor of orotate phosphoribosyltransferase [potassium oxonate (oxonic acid)], was developed to increase the feasibility and therapeutic index of 5-FU administered orally. The principal objective of this study was to assess the feasibility of administering S-1 on a once-daily-for-28-day schedule every 5 weeks, determine the maximum tolerated dose, characterize the pharmacokinetics of S-1, and seek evidence of anticancer activity. EXPERIMENTAL DESIGN: Patients with advanced solid malignancies were treated with escalating doses of S-1 on a once-daily oral schedule for 28 days every 5 weeks. The maximum tolerated dose was defined as the highest dose in which fewer than two of the first six new patients experienced dose-limiting toxicity. The pharmacokinetic profiles of the tegafur, CDHP, and oxonic acid constituents were characterized. RESULTS: Twenty patients were treated with 72 courses of S-1 at three dose levels ranging from 50 to 70 mg/m(2)/day. Diarrhea, which was often associated with abdominal discomfort and cramping, was the principal dose-limiting toxicity of S-1 on this protracted schedule. Nausea, vomiting, mucositis, fatigue, and cutaneous effects were also observed but were rarely severe. Myelosuppression was modest and uncommon. A partial response and a 49% reduction in tumor size were observed in patients with fluoropyrimidine- and irinotecan-resistant colorectal carcinoma. The pharmacokinetic data suggested potent inhibition of 5-FU clearance by CHDP, with resultant 5-FU exposure at least 10-fold higher than that reported from equitoxic doses of tegafur modulated by uracil in the oral fluoropyrimidine UFT. CONCLUSIONS: The recommended dose for Phase II studies of S-1 administered once daily for 28 consecutive days every 5 weeks is 50 mg/m(2)/day. The pharmacokinetic data indicate substantial modulation of 5-FU clearance by CDHP. Based on these pharmacokinetic data, the predictable toxicity profile of S-1, and the low incidence of severe adverse effects at the recommended Phase II dose, evaluations of S-1 on this schedule are warranted in malignancies that are sensitive to the fluoropyrimidines. PMID- 15297392 TI - Phase I study of concomitant chemoradiotherapy with paclitaxel, fluorouracil, gemcitabine, and twice-daily radiation in patients with poor-prognosis cancer of the head and neck. AB - PURPOSE: We previously demonstrated high locoregional control, in patients with poor-prognosis head and neck cancer (HNC), using paclitaxel, 5-fluorouracil, hydroxyurea, and concomitant hyperfractionated radiotherapy. In the present phase I trial, gemcitabine, a novel antimetabolite with strong radiation-enhancing activity, replaces hydroxyurea. We sought to determine the recommended phase II dose and clinical efficacy in poor-prognosis HNC patients. EXPERIMENTAL DESIGN: Seventy-two patients enrolled. Eligibility criteria included recurrent or second primary HNC, metastases or expected 2-year survival <20%. Chemoradiotherapy consisted of 5-fluorouracil, 600 mg/m(2)/d, for 5 days; paclitaxel, 100 mg/m(2) on Day 1; and concurrent 1.5 Gy twice-daily radiation for 5 days. Gemcitabine was dose escalated, 50-300 mg/m(2) on day 1. Cycles repeated every 14 days until the completion of chemoradiation. Dose-limiting toxicities (DLTs) included: neutropenic fever; grade > or =4 neutropenia or thrombocytopenia for >4 days; grade > or =4 mucositis or dermatitis for >7 days; or grade 3 toxicity necessitating chemotherapy dose reductions. Non-DLT dose reductions in 5 fluorouracil and/or paclitaxel were allowed. RESULTS: Seventy-nine percent of assessable patients experienced a clinical response. Five-year actuarial survival is 33.0%, and locoregional control is 61.4%. The recommended phase II dose of gemcitabine in this regimen is 100 mg/m(2) during cycles 1-5 (1 of 7 patients with DLT) or 200 mg/m(2) delivered only during cycles 3-5 (3 of 19 with DLT). Grades 3 and 4 mucositis (56 and 21%, respectively) and dermatitis (25 and 21%, respectively) were common. CONCLUSIONS: Gemcitabine, 5-fluorouracil, paclitaxel, and twice-daily radiation, delivered on alternating weeks, is active in patients with poor-prognosis HNC, although severe mucositis limits the clinical applicability of this regimen. Refinements in radiotherapy, including intensity modulated radiation therapy, may improve the tolerance for this regimen. PMID- 15297393 TI - CpG island hypermethylation of the DNA repair enzyme methyltransferase predicts response to temozolomide in primary gliomas. AB - PURPOSE: The DNA repair enzyme O(6)-methylguanine DNA methyltransferase (MGMT) inhibits the killing of tumor cells by alkylating agents, and its loss in cancer cells is associated with hypermethylation of the MGMT CpG island. Thus, methylation of MGMT has been correlated with the clinical response to 1,3-bis(2 chloroethyl)-1-nitrosourea (BCNU) in primary gliomas. Here, we investigate whether the presence of MGMT methylation in gliomas is also a good predictor of response to another emergent alkylating agent, temozolomide. EXPERIMENTAL DESIGN: Using a methylation-specific PCR approach, we assessed the methylation status of the CpG island of MGMT in 92 glioma patients who received temozolomide as first line chemotherapy or as treatment for relapses. RESULTS: Methylation of the MGMT promoter positively correlated with the clinical response in the glioma patients receiving temozolomide as first-line chemotherapy (n = 40). Eight of 12 patients with MGMT-methylated tumors (66.7%) had a partial or complete response, compared with 7 of 28 patients with unmethylated tumors (25.0%; P = 0.030). We also found a positive association between MGMT methylation and clinical response in those patients receiving BCNU (n = 35, P = 0.041) or procarbazine/1-(2-chloroethyl)-3 cyclohexyl-1-nitrosourea (n = 17, P = 0.043) as first-line chemotherapy. Overall, if we analyze the clinical response of all of the first-line chemotherapy treatments with temozolomide, BCNU, and procarbazine/1-(2-chloroethyl)-3 cyclohexyl-1-nitrosourea as a group in relation to the MGMT methylation status, MGMT hypermethylation was strongly associated with the presence of partial or complete clinical response (P < 0.001). Finally, the MGMT methylation status determined in the initial glioma tumor did not correlate with the clinical response to temozolomide when this drug was administered as treatment for relapses (P = 0.729). CONCLUSIONS: MGMT methylation predicts the clinical response of primary gliomas to first-line chemotherapy with the alkylating agent temozolomide. These results may open up possibilities for more customized treatments of human brain tumors. PMID- 15297394 TI - Excision repair cross-complementation group 1 polymorphism predicts overall survival in advanced non-small cell lung cancer patients treated with platinum based chemotherapy. AB - DNA repair is a critical mechanism of resistance to platinum-based chemotherapy. Excision repair cross-complementation group 1 (ERCC1) is the lead enzyme in the nucleotide excision repair process. Increased ERCC1 mRNA levels are related directly to platinum resistance in various cancers. We examined the association between two polymorphisms of ERCC1, codon 118 C/T and C8092A, which are associated with altered ERCC1 mRNA levels and mRNA stability, and overall survival (OS) in 128 advanced non-small cell lung cancer patients treated with platinum-based chemotherapy. The two polymorphisms were in linkage disequilibrium. There was a statistically significant association between the C8092A polymorphism and OS (P = 0.006, by log-rank test), with median survival times of 22.3 (C/C) and 13.4 (C/A or A/A) months, respectively, suggesting that any copies of the A allele were associated with poor outcome. No statistically significant association was found for the codon 118 polymorphism and OS (P = 0.41, by log-rank test), with median survival times of 19.9 (T/T), 16.1 (C/T), and 13.3 (C/C) months, respectively. In conclusion, the ERCC1 C8092A polymorphism may be a useful predictor of OS in advanced non-small cell lung cancer patients treated with platinum-based chemotherapy. PMID- 15297395 TI - Global gene expression profile of nasopharyngeal carcinoma by laser capture microdissection and complementary DNA microarrays. AB - A number of genetic and epigenetic changes underlying the development of nasopharyngeal carcinomas have recently been identified. However, there is still limited information on the nature of the genes and gene products whose aberrant expression and activity promote the malignant conversion of nasopharyngeal epithelium. Here, we have performed a genome-wide transcriptome analysis by probing cDNA microarrays with fluorescent-labeled amplified RNA derived from laser capture microdissected cells procured from normal nasopharyngeal epithelium and areas of metaplasia-dysplasia and carcinoma from EBV-associated nasopharyngeal carcinomas. This approach enabled the identification of genes differentially expressed in each cell population, as well as numerous genes whose expression can help explain the aggressive clinical nature of this tumor type. For example, genes indicating cell cycle aberrations (cyclin D2, cyclin B1, activator of S-phase kinase, and the cell cycle checkpoint kinase, CHK1) and invasive-metastatic potential (matrix metalloproteinase 11, v-Ral, and integrin beta(4)) were highly expressed in tumor cells. In contrast, genes underexpressed in tumors included genes involved in apoptosis (B-cell CLL/lymphoma 6, secretory leukocyte protease inhibitor, and calpastatin), cell structure (keratin 7 and carcinoembryonic antigen-related cell adhesion molecule 6), and putative tumor suppressor genes (H-Ras-like suppressor 3, retinoic acid receptor responder 1, and growth arrested specific 8) among others. Gene expression patterns also suggested alterations in the Wnt/beta-catenin and transforming growth factor beta pathways in nasopharyngeal carcinoma. Thus, expression profiles indicate that aberrant expression of growth, survival, and invasion-promoting genes may contribute to the molecular pathogenesis of nasopharyngeal carcinoma. Ultimately, this approach may facilitate the identification of clinical useful markers of disease progression and novel potential therapeutic targets for nasopharyngeal carcinoma. PMID- 15297396 TI - Imiquimod treatment induces expression of opioid growth factor receptor: a novel tumor antigen induced by interferon-alpha? AB - PURPOSE: Imiquimod represents a synthetic local immune response modifier that has demonstrated efficacy in clearing basal cell carcinoma. Via interaction with Toll like receptor 7 on immune cells, imiquimod induces local production of cytokines, such as interferon (IFN)-alpha. EXPERIMENTAL DESIGN: To more closely define and elucidate mechanisms leading to basal cell carcinoma clearance in vivo, we examined gene expression profiles of skin basal cell carcinoma before and after treatment with 5% imiquimod cream (Aldara) by using high-density oligonucleotide arrays. RESULTS: We show that imiquimod predominantly induces genes involved in different aspects of immune response. In addition to effects on immunity, imiquimod treatment modulates the expression of genes involved in the control of apoptosis and oncogenesis. Array data indicated that imiquimod treatment induces expression of opioid growth factor receptor, a molecule recently reported to be a target for antitumor antibody responses. Immunohistochemistry revealed in vivo up regulation of opioid growth factor receptor protein on tumor and on infiltrating cells after treatment. By using basal cell carcinoma cell lines treated with IFN alpha or imiquimod, we show that opioid growth factor receptor up-regulation is IFN-alpha-mediated, rather then directly imiquimod-mediated. By using tissue microarray containing 52 basal cell carcinomas, we demonstrate opioid growth factor receptor expression in almost half of the cases. Expression of opioid growth factor receptor correlated with a longer recurrence-free period in basal cell carcinoma that recurred after radiotherapy (Kaplan-Meier analysis, P = 0.041). CONCLUSIONS: In addition to its immunomodulatory and antiproliferative activity, opioid growth factor receptor seems to have a prognostic significance in basal cell carcinoma patients. Our data add to the growing list of basal cell carcinoma-associated tumor antigens. PMID- 15297397 TI - Expression profiling of T-cell lymphomas differentiates peripheral and lymphoblastic lymphomas and defines survival related genes. AB - PURPOSE: T-Cell lymphomas constitute heterogeneous and aggressive tumors in which pathogenic alterations remain largely unknown. Expression profiling has demonstrated to be a useful tool for molecular classification of tumors. EXPERIMENTAL DESIGN: Using DNA microarrays (CNIO-OncoChip) containing 6386 cancer related genes, we established the expression profiling of T-cell lymphomas and compared them to normal lymphocytes and lymph nodes. RESULTS: We found significant differences between the peripheral and lymphoblastic T-cell lymphomas, which include a deregulation of nuclear factor-kappaB signaling pathway. We also identify differentially expressed genes between peripheral T cell lymphoma tumors and normal T lymphocytes or reactive lymph nodes, which could represent candidate tumor markers of these lymphomas. Additionally, a close relationship between genes associated to survival and those that differentiate among the stages of disease and responses to therapy was found. CONCLUSIONS: Our results reflect the value of gene expression profiling to gain insight about the molecular alterations involved in the pathogenesis of T-cell lymphomas. PMID- 15297398 TI - Antitumor activity of systemically delivered ribozymes targeting murine telomerase RNA. AB - PURPOSE: To test ribozymes targeting mouse telomerase RNA (mTER) for suppression of the progression of B16-F10 murine melanoma metastases in vivo. EXPERIMENTAL DESIGN: Hammerhead ribozymes were designed to target mTER. The ribozyme sequences were cloned into a plasmid expression vector containing EBV genomic elements that substantially prolong expression of genes delivered in vivo. The activity of various antitelomerase ribozymes or control constructs was examined after i.v. injection of cationic liposome:DNA complexes containing control or ribozyme constructs. Expression of ribozymes and mTER at various time points were evaluated by quantitative real-time PCR. Telomerase activity was examined using the telomeric repeat amplification protocol. RESULTS: Systemic administration of cationic liposome:DNA complexes containing a plasmid-expressed ribozyme specifically targeting a cleavage site at mTER nucleotide 180 significantly reduced the metastatic progression of B16-F10 murine melanoma. The antitumor activity of the anti-TER 180 ribozyme in mice was abolished by a single inactivating base mutation in the ribozyme catalytic core. The EBV-based expression plasmid produced sustained levels of ribozyme expression for the full duration of the antitumor studies. In addition to antitumor activity, cationic liposome:DNA complex-based ribozyme treatment also produced reductions in both TER levels and telomerase enzymatic activity in tumor-bearing mice. CONCLUSIONS: Systemic, plasmid-based ribozymes specifically targeting TER can reduce both telomerase activity and metastatic progression in tumor-bearing hosts. The work reported here demonstrates the potential utility of plasmid-based anti-TER ribozymes in the therapy of melanoma metastasis. PMID- 15297399 TI - Superior activity of the combination of histone deacetylase inhibitor LAQ824 and the FLT-3 kinase inhibitor PKC412 against human acute myelogenous leukemia cells with mutant FLT-3. AB - PURPOSE: Mutant FLT-3 receptor tyrosine kinase is a client protein of the molecular chaperone heat shock protein 90 and is commonly present and contributes to the leukemia phenotype in acute myelogenous leukemia (AML). LAQ824, a cinnamyl hydroxamate histone deacetylase inhibitor, is known to induce acetylation and inhibition of heat shock protein 90. Here, we determined the effects of LAQ824 and/or PKC412 (a FLT-3 kinase inhibitor) on the levels of mutant FLT-3 and its downstream signaling, as well as growth arrest and cell-death of cultured and primary human AML cells. EXPERIMENTAL DESIGN: The effect of LAQ824 and/or PKC412 treatment was determined on the levels of FLT-3 and phosphorylated (p)-FLT-3, on downstream pro-growth and pro-survival effectors, e.g., p-STAT5, p-AKT, and p extracellular signal-regulated kinase (ERK) 1/2, and on the cell cycle status and apoptosis in the cultured MV4-11 and primary AML cells with mutant FLT-3. RESULTS: Treatment with LAQ824 promoted proteasomal degradation and attenuation of the levels of FLT-3 and p-FLT-3, associated with cell cycle G(1)-phase accumulation and apoptosis of MV4-11 cells. This was accompanied by attenuation of p-STAT5, p-AKT, and p-ERK1/2 levels. STAT-5 DNA-binding activity and the levels of c-Myc and oncostatin M were also down-regulated. Cotreatment with LAQ824 and PKC412 synergistically induced apoptosis of MV4-11 cells and induced more apoptosis of the primary AML cells expressing mutant FLT-3. This was also associated with more attenuation of p-FLT-3, p-AKT, p-ERK1/2, and p-STAT5. CONCLUSIONS: The combination of LAQ824 and PKC412 is highly active against human AML cells with mutant FLT-3, which merits in vivo studies of the combination against human AML. PMID- 15297400 TI - The effects of standard anthracycline-based chemotherapy on soluble ICAM-1 and vascular endothelial growth factor levels in breast cancer. AB - PURPOSE: The circulating soluble form of intercellular adhesion molecule-1 (sICAM 1) and vascular endothelial growth factor (VEGF) are elevated in women with breast cancer and associated with tumor progression and poor prognosis. This study examined the effects of anthracycline-based chemotherapy on plasma sICAM-1 and VEGF, as well as soluble P-selectin, von Willebrand factor, and interleukin-6 levels. EXPERIMENTAL DESIGN: Twenty-six women diagnosed with stage I-IIIA breast cancer (mean age, 48.4 +/- 10.4 years; range, 34-79 years) were studied before (week 1) and at weeks 2 and 3 of cycles 1 and 4 of chemotherapy. RESULTS: The initial effect of chemotherapy was to reduce sICAM-1 levels; compared with pretreatment, sICAM-1 levels were decreased at week 2 of both cycles (P values < 0.01). sICAM-1 levels were elevated, however, at the start of cycle 4 as compared with pretreatment (P < 0.01). Chemotherapy led to an increase in sICAM-1 levels in node-positive but not node-negative patients (P < 0.01). VEGF levels were decreased at week 2 of cycle 4 (P = 0.001) and remained so at week 3. Similar to sICAM-1, VEGF levels were elevated at the start of cycle 4 as compared with pretreatment (P < 0.006). Soluble P-selectin levels decreased during week 2 of cycle 4 (P = 0.026). Neither interleukin-6 or von Willebrand factor were significantly changed in response to chemotherapy. CONCLUSIONS: The findings support prior studies suggesting that sICAM-1 levels derive from sources other than endothelial cells. In addition, whereas the more immediate effect of chemotherapy is to reduce sICAM-1 and VEGF, continued treatment may lead to significant elevations. PMID- 15297401 TI - Immune responses to a class II helper peptide epitope in patients with stage III/IV resected melanoma. AB - The importance of CD8(+) cytolytic T cells for protection from viral infection and in the generation of immune responses against tumors has been well established. In contrast, the role of CD4(+) T-helper cells in human infection and in cancer immunity has yet to be clearly defined. In this pilot study, we show that immunization of three resected, high-risk metastatic melanoma patients with a T-helper epitope derived from the melanoma differentiation antigen, melanoma antigen recognized by T cells-1, results in CD4(+) T-cell immune responses. Immune reactivity to that epitope was detected by DR4-peptide tetramer staining, and enzyme-linked immunospot assay of fresh and restimulated CD4(+) T cells from patients over the course of the 12-month vaccine regimen. The postvaccine CD4(+) T cells exhibited a mixed T-helper 1/T-helper 2 phenotype, proliferated in response to the antigen and promiscuously recognized the peptide epitope bound to different human leukocyte antigen-DRbeta alleles. For 1 DRbeta1*0401(+) patient, antigen-specific CD4(+) T cells recognized human leukocyte antigen-matched antigen-expressing tumor cells, secreted granzyme B, and also exhibited cytolysis that was MHC class II-restricted. These data establish the immunogenicity of a class II epitope derived from a melanoma associated antigen and support the inclusion of class II peptides in future melanoma vaccine therapies. PMID- 15297402 TI - Pilot trial evaluating an 123I-labeled 80-kilodalton engineered anticarcinoembryonic antigen antibody fragment (cT84.66 minibody) in patients with colorectal cancer. AB - PURPOSE: The chimeric T84.66 (cT84.66) minibody is a novel engineered antibody construct (V(L)-linker-V(H)-C(H)3; 80 kDa) that demonstrates bivalent and high affinity (4 x 10(10) m(-1)) binding to carcinoembryonic antigen (CEA). The variable regions (V(L) and V(H)) assemble to form the antigen-combining sites, and the protein forms dimers through self-association of the C(H)3 domains. In animal models, the minibody demonstrated high tumor uptake, approaching that of some intact antibodies, substantially faster clearance than intact chimeric T84.66, and superior tumor-to-blood ratios compared with the cT84.66 F(ab')(2) fragment, making it attractive for further evaluation as an imaging and therapy agent. The purpose of this pilot clinical study was to determine whether (123)I cT84.66 minibody demonstrated tumor targeting and was well tolerated as well as to begin to evaluate its biodistribution, pharmacokinetics, and immunogenicity in patients with colorectal cancer. EXPERIMENTAL DESIGN: Ten patients with biopsy proven colorectal cancer (6 newly diagnosed, 1 pelvic recurrence, 3 limited metastatic disease) were entered on this study. Each received 5-10 mCi (1 mg) of (123)I-labeled minibody i.v. followed by serial nuclear scans and blood and urine sampling over the next 48-72 h. Surgery was performed immediately after the last nuclear scan. RESULTS: Tumor imaging was observed with (123)I-labeled minibody in seven of the eight patients who did not receive neoadjuvant therapy before surgery. Two patients received neoadjuvant radiation and chemotherapy, which significantly reduced tumor size before surgery and minibody infusion. At surgery, no tumor was detected in one patient and only a 2-mm focus was seen in the second patient. (123)I-labeled minibody tumor targeting was not seen in either of these pretreated patients. Mean serum residence time of the minibody was 29.8 h (range, 10.9-65.4 h). No drug-related adverse reactions were observed. All 10 patients were evaluated for immune responses to the minibody, with no significant responses observed. CONCLUSION: This pilot study represents one of the first clinical efforts to evaluate an engineered intermediate-molecular-mass radiolabeled antibody construct directed against CEA. cT84.66 minibody demonstrates tumor targeting to colorectal cancer and a faster clearance in comparison with intact antibodies, making it appropriate for further evaluation as an imaging and therapy agent. The mean residence time of the minibody in patients is longer than predicted from murine models. We therefore plan to further evaluate its biodistribution and pharmacokinetic properties with minibody labeled with a longer-lived radionuclide, such as (111)In. PMID- 15297403 TI - Randomized phase II trial of sequential chemotherapy in advanced non-small cell lung cancer (SWOG 9806): carboplatin/gemcitabine followed by paclitaxel or cisplatin/vinorelbine followed by docetaxel. AB - PURPOSE: Improving chemotherapeutic efficacy in non-small cell lung cancer (NSCLC) will require the development of new drugs or new strategies to better use currently available agents. Sequential administration offers an opportunity to increase drug diversity while maintaining dose intensity. On the basis of the data indicating the activity of taxanes as second-line therapy and the lack of efficacy for more than three cycles of platinum-based therapy, this randomized Phase II study tested the concept of planned sequential chemotherapy in advanced NSCLC. EXPERIMENTAL DESIGN: Patients with selected stage IIIb (pleural effusion)/stage IV NSCLC, performance status of 0-1 and normal organ function were eligible. THERAPY: arm 1, carboplatin (area under the curve = 5.5 mg/ml x min day 1) and gemcitabine (1000 mg/m(2) days 1 and 8 every 21 days x 3) followed by paclitaxel (225 mg/m(2) every 21 days x 3) or arm 2, cisplatin (100 mg/m(2) day 1), vinorelbine (25 mg/m(2) days 1 and 8 every 21 d x 3) followed by docetaxel (75-100 mg/m(2) every 21 days x 3). RESULTS: Two-hundred four patients were accrued, of whom, 178 were eligible and evaluable. Eighty percent of patients were stage IV on arm 1 and 85% on arm 2. Response rates were 21 and 28% on arms 1 and 2, respectively. Median, 1-year and 2-year survivals were 9 months, 34 and 13%, and 9 months, 36 and 8%, on arms 1 and 2, respectively. CONCLUSIONS: Sequential therapy, as used in this study, resulted in comparable efficacy to previous Southwest Oncology Group trials of two drug combinations in this population; however, it failed to meet criteria for further study. PMID- 15297404 TI - A phase I study of interleukin 12 with trastuzumab in patients with human epidermal growth factor receptor-2-overexpressing malignancies: analysis of sustained interferon gamma production in a subset of patients. AB - PURPOSE: On the basis of preclinical studies, we hypothesized that interleukin (IL)12 would potentiate the antitumor actions of an antihuman epidermal growth factor receptor-2 (HER2) monoclonal antibody (trastuzumab). We conducted a Phase I trial to determine the safety and optimal biological dose of IL-12 when given in combination with trastuzumab. PATIENTS AND METHODS: Patients with metastatic HER2-positive malignancies received trastuzumab on day 1 of each weekly cycle. Beginning in week 3, patients also received intravenous injections of IL-12 on days 2 and 5. The IL-12 component was dose-escalated within cohorts of 3 patients (30, 100, 300, or 500 ng/kg). Correlative assays were conducted using serum samples and peripheral blood cells obtained during the course of therapy. RESULTS: Fifteen patients were treated, including 12 with HER2 2+ or 3+ breast cancer. The regimen was well tolerated with IL-12-induced grade 1 nausea and grade 2 fatigue predominating. Evaluation of dose-limiting toxicity and biological end points suggested that the 300 ng/kg dose was both the maximally tolerated dose and the optimal biological dose of IL-12 for use in combination with trastuzumab. Two patients with HER2 3+ breast cancer within the 500 ng/kg dose level experienced grade 1 asymptomatic decreases in left ventricular ejection fraction of 12% and 19% after 3 and 10 months of therapy, respectively. There was one complete response in a patient with HER2 3+ breast cancer metastatic to the axillary, mediastinal, and supraclavicular nodes, and 2 patients with stabilization of bone disease lasting 10 months and >12 months, respectively. Correlative assays showed sustained production of interferon (IFN)gamma by natural killer cells only in those patients experiencing a clinical response or stabilization of disease. Elevated serum levels of macrophage inflammatory protein-1alpha, tumor necrosis factor-alpha, and the antiangiogenic factors IFN-gamma inducible protein-10 and monokine induced by gamma were also observed in these patients. Patient genotyping suggested that a specific IFN gamma gene polymorphism might have been associated with increased IFN-gamma production. The ability of patient peripheral blood cells to conduct antibody dependent cellular cytotoxicity against tumor targets in vitro did not correlate with clinical response or dose of IL-12. CONCLUSIONS: The addition of IL-12 to trastuzumab therapy did not appear to enhance the efficacy of this antibody treatment. Sustained production of IFN-gamma and other cytokines were observed in three patients: One who exhibited a complete response and two others who had stabilization of disease lasting over 6 months. Given the small sample size and heterogeneity of the patient population, the effects of IL-12 on the innate immune response to trastuzumab therapy should be further explored in the context of a larger clinical trial. PMID- 15297405 TI - Phase I trial of the cyclin-dependent kinase inhibitor flavopiridol in combination with docetaxel in patients with metastatic breast cancer. AB - PURPOSE: The purpose of this study was to determine the toxicities and characterize the pharmacokinetics of docetaxel and flavopiridol in patients with metastatic breast cancer. EXPERIMENTAL DESIGN: Docetaxel was administered at an initial dose of 60 mg/m(2) followed in 24 hours by a 72-hour infusion of flavopiridol at 50 mg/m(2)/d every 3 weeks. Because dose-limiting myelosuppression occurred, the schedule was amended to docetaxel, 50 mg/m(2), followed by escalating doses of flavopiridol (starting dose, 26 mg/m(2)/d) as a 1 hour infusion daily for 3 days. Pharmacokinetic studies were performed. Ki67, p53, and phosphorylated retinoblastoma protein (phospho-Rb) in paired tumor and buccal mucosa biopsies (obtained pre- and posttreatment) were examined by immunohistochemistry. RESULTS: Eleven patients were enrolled. Five patients received docetaxel and 72-hour flavopiridol. Dose-limiting toxicity was grade 4 neutropenia. Six patients received docetaxel and 1-hour flavopiridol, and the dose-limiting toxicity was grade 3 hypotension. Pharmacokinetics of flavopiridol and docetaxel were consistent with historical data. Nuclear staining with p53 increased and phospho-Rb decreased in 10 pairs of buccal mucosa biopsies posttreatment (P = 0.002 and P = 0.04, respectively). No significant changes in Ki67, p53, or phospho-Rb were detected in six paired tumors. Two patients sustained stable disease for >3 months (72-hour flavopiridol), and one partial response was observed (1-hour flavopiridol). CONCLUSIONS: Docetaxel combined with 72-hour flavopiridol was not feasible because of dose-limiting neutropenia. Dose escalation of a 1-hour infusion of flavopiridol with docetaxel was also not possible. The changes in p53 and phospho-Rb in buccal mucosa suggest that a biological effect with flavopiridol was achieved. PMID- 15297406 TI - A Phase I pharmacokinetic and biological correlative study of oblimersen sodium (genasense, g3139), an antisense oligonucleotide to the bcl-2 mRNA, and of docetaxel in patients with hormone-refractory prostate cancer. AB - PURPOSE: To assess the feasibility of administering oblimersen sodium, a phosphorothioate antisense oligonucleotide directed to the Bcl-2 mRNA, with docetaxel to patients with hormone-refractory prostate cancer; to characterize the pertinent pharmacokinetic parameters, Bcl-2 protein inhibition in peripheral blood mononuclear cell(s) (PBMC) and tumor; and to seek preliminary evidence of antitumor activity. EXPERIMENTAL DESIGN: Patients were treated with increasing doses of oblimersen sodium administered by continuous i.v. infusion on days 1 to 6 and docetaxel administered i.v. over 1 h on day 6 every 3 weeks. Plasma was sampled to characterize the pharmacokinetic parameters of both oblimersen and docetaxel, and Bcl-2 protein expression was measured from paired collections of PBMCs pretreatment and post-treatment. RESULTS: Twenty patients received 124 courses of the oblimersen and docetaxel combination at doses ranging from 5 to 7 mg/kg/day oblimersen and 60 to 100 mg/m(2) docetaxel. The rate of severe fatigue accompanied by severe neutropenia was unacceptably high at doses exceeding 7 mg/kg/day oblimersen and 75 mg/m(2) docetaxel. Nausea, vomiting, and fever were common, but rarely severe. Oblimersen mean steady-state concentrations were 3.44 +/- 1.31 and 5.32 +/- 2.34 at the 5- and 7-mg/kg dose levels, respectively. Prostate-specific antigen responses were observed in 7 of 12 taxane-naive patients, but in taxane-refractory patients no responses were observed. Preliminary evaluation of Bcl-2 expression in diagnostic tumor specimens was not predictive of response to this therapy. CONCLUSIONS: The recommended Phase II doses for oblimersen and docetaxel on this schedule are 7 mg/kg/day continuous i.v. infusion days 1 to 6, and 75 mg/m(2) i.v. day 6, respectively, once every 3 weeks. The absence of severe toxicities at this recommended dose, evidence of Bcl 2 protein inhibition in PBMC and tumor tissue, and encouraging antitumor activity in HPRC patients warrant further clinical evaluation of this combination. PMID- 15297407 TI - Phase I and pharmacologic study of intermittently administered 9 nitrocamptothecin in patients with advanced solid tumors. AB - PURPOSE: 9-Nitrocamptothecin (9NC) is an oral camptothecin analogue currently administered at 1.5 mg/m(2)/day x 5 days/week in Phase III studies for pancreatic carcinoma. In an effort to increase the dose administered per day and determine whether the daily dose or number of days of treatment influence toxicity, we performed a Phase I study of 9NC using intermittent schedules of administration. EXPERIMENTAL DESIGN: On schedule A, 9NC was administered orally daily x 5 days for 2 weeks every 4 weeks (one cycle). On schedule B, 9NC was administered orally daily x 14 days every 4 weeks (one cycle). Dose levels were determined by adaptive dose finding. Serial blood samples were obtained on day 1 of each schedule for pharmacokinetic studies of 9NC and its 9-aminocamptothecin (9AC) metabolite, and lactone forms were measured by high-performance liquid chromatography. RESULTS: The recommended Phase II doses for schedules A and B were 2.43 and 1.70 mg/m(2)/day, respectively, each providing the same dose intensity (i.e., 24 mg/m(2)/cycle). The primary toxicities on schedules A and B were neutropenia, thrombocytopenia, and diarrhea. On schedule A, two patients with gastric cancer and two patients with pancreatic cancer had stable disease for more than six cycles. On schedule B, one patient with pancreatic cancer had stable disease for more than six cycles, and a patient with pancreatic cancer had a partial response. There was significant interpatient variability in the disposition of 9NC and 9AC. Most of the drug remained in the 9NC form with a ratio of 9NC to 9AC of approximately 4 to 1. CONCLUSIONS: These studies suggest that 9NC administered on an intermittent schedule is tolerable and may be an active regimen in patients with gastric or pancreatic cancers. Dosing 9NC on a mg/m(2) basis does not reduce pharmacokinetic variability. PMID- 15297408 TI - Extended follow-up of patients treated with imatinib mesylate (gleevec) for chronic myelogenous leukemia relapse after allogeneic transplantation: durable cytogenetic remission and conversion to complete donor chimerism without graft versus-host disease. AB - PURPOSE: Over the last several years, donor lymphocyte infusions have become the standard approach for patients with chronic myelogenous leukemia (CML) who relapse after allogeneic stem cell transplantation (SCT). Recent reports indicate that imatinib mesylate (Gleevec) can induce remissions in these patients as well. Less is known about the extent and durability of these responses. EXPERIMENTAL DESIGN: We studied 15 patients treated with imatinib for recurrent CML after SCT, 10 patients with stable phase CML (SP-CML), 1 with accelerated phase (AP-CML), and 4 with blast phase (BP-CML). The dose of imatinib was 600 mg (n = 10) or 400 mg (n = 5) daily. Patients were followed for hematological, cytogenetic, and molecular response. A susbset of responders was followed for changes in donor derived hematopoietic chimerism. RESULTS: Of the 10 patients with SP-CML, all achieved a hematological response. Within 3 months, five of these patients had achieved a complete cytogenetic response (CCR). By six months, 9 of 10 patients had achieved CCR. The BCR-ABL transcript could not be detected by reverse transcription-PCR in 7 of these 9 patients. Patients who achieved CCR showed evidence of conversion to complete donor chimerism. No patient developed graft versus-host disease. With a median follow up of 25 months, all patients are alive and 9 of 10 patients remain in CCR. Only one of the 5 patients with AP/BP-CML achieved a complete cytogenetic response. CONCLUSIONS: We find that imatinib is well tolerated in patients with SP-CML who relapse after SCT. Responses were rapid, durable, and associated with conversion to full donor chimerism without graft-versus-host disease. Imantinib was far less effective in patients who relapsed with AP/BP-CML. Imatinib should be evaluated as either an alternative or as an adjunct to donor lymphocyte infusions for patients with SP-CML who relapse after SCT. PMID- 15297409 TI - Pharmacokinetics of O(6)-benzylguanine in pediatric patients with central nervous system tumors: a pediatric oncology group study. AB - PURPOSE: To report the results of the first pharmacokinetic study in pediatric patients of O(6)-benzylguanine (O(6)BG), which irreversibly inactivates the DNA repair protein alkylguanine-alkyltransferase, thus enhancing the cytotoxicity of nitrosoureas. EXPERIMENTAL DESIGN: As part of a Pediatric Oncology Group Phase I study, 120 mg/m(2) of O(6)BG was administered i.v. over 1 h, before 1,3-bis(2 chloroethyl)-1-nitrosourea administration in children with recurrent or refractory brain tumors. Serial blood samples for plasma pharmacokinetic studies were obtained. Concentrations of O(6)BG and its active metabolite O(6)-benzyl-8 oxoguanine (8-oxo-O(6)BG) were measured by high-performance liquid chromatography. A pharmacokinetic model and additional first-order elimination rate constants for each compound were developed. RESULTS: O(6)BG concentration versus time data were evaluated for 25 patients. The peak concentration of O(6)BG (mean +/- SD) was 11 +/- 4 microm, and the peak concentration of its active metabolite, 8-oxo-O(6)BG, was 35 +/- 10 microm. O(6)BG was rapidly eliminated with a half-life of 85 +/- 140 min, area under the curve of 795 +/- 320 microm. min and clearance of 760 +/- 400 ml/min/m(2). The area under the curve of 8-oxo O(6)BG when extrapolated to infinity was 22,700 +/- 11,800 microm. min. The clearance and terminal half-life of 8-oxo-O(6)BG were 30 +/- 15 ml/min/m(2) and 360 +/- 220 min, respectively. CONCLUSIONS: There is rapid elimination of O(6)BG after i.v. administration over 1 h. In contrast, the terminal half-life for the active metabolite, 8-oxo-O(6)BG, is 4-fold longer. The pharmacokinetic parameters for O(6)BG and 8-oxo-O(6)BG are similar to those reported previously in adults. PMID- 15297410 TI - Prognostic analysis of early lymphocyte recovery in patients with advanced breast cancer receiving high-dose chemotherapy with an autologous hematopoietic progenitor cell transplant. AB - PURPOSE: The purpose of this study was to evaluate the prognostic effect of early posttransplant lymphocyte recovery in patients with advanced breast cancer receiving high-dose chemotherapy with autologous hematopoietic progenitor cell transplantation. EXPERIMENTAL DESIGN: We analyzed the effect of the absolute lymphocyte count on day +15 posttransplant on freedom from relapse and overall survival in patients with high-risk primary breast cancer or metastatic breast cancer, enrolled between 1990 and 2001 in prospective high-dose chemotherapy trials, using a uniform regimen of cyclophosphamide, cisplatin, and 1,3-bis(2 chloroethyl)-1-nitrosourea. RESULTS: Four hundred and seventy-six patients (264 high-risk primary breast cancer and 212 metastatic breast cancer patients) were evaluated at median follow-up of 8 years (range, 1.5-11 years). The disease-free survival and overall survival rates in the high-risk primary breast cancer group were 67% and 70%, respectively. Patients with metastatic breast cancer patients had 21.8% disease-free survival and 31.5% overall survival rates. Day +15 absolute lymphocyte count correlated with freedom from relapse (P = 0.007) and overall survival (P = 0.04) in the metastatic breast cancer group, but not in the high-risk primary breast cancer group (P = 0.5 and 0.8, respectively). The prognostic effect of absolute lymphocyte count in metastatic breast cancer was restricted to those patients receiving unmanipulated peripheral blood progenitor cells (P = 0.04). In contrast, absolute lymphocyte count had no significant effect in those metastatic breast cancer patients receiving bone marrow or a CD34 selected product. In multivariate analyses, the prognostic effect of day +15 absolute lymphocyte count in metastatic breast cancer was independent of other predictors, such as disease status, pre-high-dose chemotherapy treatment, number of tumor sites, or HER2. CONCLUSIONS: Early lymphocyte recovery is an independent outcome predictor in metastatic breast cancer patients receiving high-dose chemotherapy and an autologous peripheral blood progenitor cell transplant. These observations suggest that immune strategies targeting minimal posttransplant residual disease may prove worthwhile. PMID- 15297411 TI - Methylation of p16 CpG islands associated with malignant transformation of gastric dysplasia in a population-based study. AB - PURPOSE: Inactivation of p16 by aberrant methylation of CpG islands is a frequent event in carcinomas and precancerous lesions of various organs, including the stomach. The aim of this study is to investigate the relationship between p16 methylation and malignant transformation of human gastric dysplasia (DYS) based on follow-up endoscopic screening in a high-risk population. EXPERIMENTAL DESIGN: Genomic DNA samples were extracted from paraffin blocks of gastric mucosal biopsies that were histopathologically diagnosed as low-grade DYS from patients who developed gastric carcinomas [GCs (n = 21)] and those that did not do so (n = 21) during 5 years of follow-up. The methylation status of p16 CpG islands of each sample was detected by methylation-specific PCR, denatured high-performance liquid chromatography, and sequencing. RESULTS: Aberrant p16 methylation was observed in 5 of 21 samples of DYS that progressed to GC but in 0 of 21 samples that did not progress to GC (P = 0.048, two-sided). Sequencing results confirmed that all CpG sites were methylated in the analyzed sequence from these five p16 methylated cases. Furthermore, p16 methylation was also observed in the five subsequent GCs. Unmethylated p16 CpG islands were detected in all of the samples without p16 methylation. CONCLUSIONS: Our findings suggest p16 methylation is correlated with the malignant transformation of gastric DYS, and p16 methylation might be a useful biomarker for prediction of malignant potential of gastric DYS. PMID- 15297412 TI - B7-H1 expression on non-small cell lung cancer cells and its relationship with tumor-infiltrating lymphocytes and their PD-1 expression. AB - PURPOSE: B7-H1/PD-L1 (B7-H1) and B7-DC/PD-L2 (B7-DC) are ligands for the receptor PD-1, which is known to negatively regulate T-cell activation. In the present study, we investigated the expression of B7-H1 and B7-DC in tumor specimens of non-small cell lung cancer and their relationships with clinicopathological variables and postoperative survival. Furthermore, we examined the correlation between B7-H1 expression on tumor cells and the number of tumor-infiltrating lymphocytes (TILs) or PD-1 expression on TILs. EXPERIMENTAL DESIGN: The expression of B7-H1 and B7-DC in 52 surgically resected specimens of non-small cell lung cancer was evaluated immunohistochemically. RESULTS: Expression of B7 H1 and B7-DC was focally observed in all non-small cell lung cancer tumor specimens. No relationship was found between the expression of B7-H1 or B7-DC and clinicopathological variables or postoperative survival. However, in the same sections evaluated, significantly fewer TILs were identified in B7-H1-positive tumor regions than in B7-H1-negative tumor regions in a subset of five patients (P = 0.01). Moreover, the percentage of TILs expressing PD-1 was significantly lower in B7-H1-positive tumor regions than in B7-H1-negative tumor regions (P = 0.02). CONCLUSIONS: The expression of B7-H1 on tumor cells in local areas reciprocally correlated with the number of TILs, and this may contribute to negative regulation in antitumor immune responses in non-small cell lung cancer. PMID- 15297413 TI - Expression of pro- and antiapoptotic proteins in circulating CD8+ T cells of patients with squamous cell carcinoma of the head and neck. AB - OBJECTIVE: Apoptosis of T lymphocytes in the circulation of patients with squamous cell carcinoma of the head and neck (SCCHN) was shown to target effector CD8+ rather than CD4+ T cells. This study evaluates the contribution of pro- and antiapoptotic components of the mitochondria-dependent pathway to apoptosis of circulating CD8+ T cells in these patients. EXPERIMENTAL DESIGN: Blood samples were obtained from 77 patients with SCCHN and 51 normal control(s) (NC). Percentages of CD8+Annexin V+ (ANX+) and CD8+CD95+ cells, changes in mitochondrial membrane potential and levels of expression of Bcl-2, Bcl-XL, and Bax in CD8+ T lymphocytes were measured by quantitative flow cytometry. RESULTS: Elevated percentages (P < 0.001) of early apo-ptotic (CD8+ANX+ CD95+) T cells in the circulation distinguish SCCHN patients from NCs but not patients with no evidence of disease (NED) from those with active disease (AD). Circulating CD8+ but not CD4+ T cells in patients were found to contain higher levels of proapoptotic Bax and antiapoptotic Bcl-XL (P < 0.01) than NC cells. The Bax/Bcl-2 ratio was elevated in CD8+ T cells of patients relative to NCs (P < 0.01), and it correlated with the percentage of ANX+CD8+ T cells (P = 0.007). The Bax/Bcl-XL ratio discriminated AD from NED patients. CONCLUSION: Apoptosis of circulating CD8+T cells is found in SCCHN patients with AD or NED. Up-regulated Bax and Bcl XL expression, the elevated Bax/Bcl-2 ratio and its association with ANX binding implicate the mitochondrial pathway in death of CD8+ T cells of patients with SCCHN. Understanding of molecular mechanisms of T-cell death and survival is essential for the development of more effective biotherapies for SCCHN. PMID- 15297414 TI - Primary acute lymphoblastic leukemia cells use a novel promoter and 5'noncoding exon for the human reduced folate carrier that encodes a modified carrier translated from an upstream translational start. AB - The human reduced folate carrier (hRFC) is reported to be regulated by up to seven alternatively spliced noncoding exons (A1, A2, A, B, C, D, and E). Noncoding exon and promoter usage was analyzed in RNAs from 27 childhood acute lymphoblastic leukemia (ALL) specimens by real-time PCR and/or 5' rapid amplification of cDNA ends (5' RACE) assay. By real-time PCR, total hRFC transcripts in ALL spanned a 289-fold range. Over 90% of hRFC transcripts were transcribed with A1, A2, and B 5' untranslated regions (UTRs). Analysis of 5' RACE clones showed that the A1 + A2 5'UTRs contained A1 sequence alone or a fusion of A1 and A2, implying the existence of a single, alternatively spliced 1021-bp A1/A2 noncoding region. High frequency sequence polymorphisms (AGG deletion, C/T transition) identified in the A1/A2 region by 5'RACE were confirmed in normal DNAs. By reporter assays in HepG2 hepatoma and Jurkat leukemia cells, A1/A2 promoter activity was localized to a 134-bp minimal region. Translation from an upstream AUG in the A1/A2 noncoding region in-frame with the normal translation start resulted in synthesis of a larger ( approximately 7 kDa) hRFC protein with transport properties altered from those for wild-type hRFC. Although there was no effect on transcript or protein stabilities, in vitro translation from A1/A2 transcripts was decreased compared with those with the B 5'UTR. Our results document the importance of the hRFC A1/A2 upstream region in childhood ALL and an intricate transcriptional and posttranscriptional regulation of hRFC A1/A2 mRNAs. Furthermore, they suggest that use of the A1/A2 5'UTR may confer a transport phenotype distinct from the other 5'UTRs due to altered translation efficiency and transport properties. PMID- 15297415 TI - Elevated S-phase kinase-associated protein 2 protein expression in acute myelogenous leukemia: its association with constitutive phosphorylation of phosphatase and tensin homologue protein and poor prognosis. AB - PURPOSE: The F-box protein S-phase kinase-associated protein 2 (Skp2) positively regulates the G(1)-S phase transition by controlling the stability of several G(1) regulators, such as p27Kip1. However, the clinical significance of Skp2 in patients with acute myelogenous leukemia (AML) remains unknown. EXPERIMENTAL DESIGN: We examined the clinical and biological significance of Skp2 expression in AML and evaluated the relationship between Skp2 and p27Kip1 expression and phosphatase and tensin homologue (PTEN) phosphorylation. RESULTS: Western blot analysis showed that high Skp2 expression was observed in 57 (57.6%) cases and significantly correlated with unfavorable cytogenetics (P = 0.035) but not with age, white blood cell count, serum lactic dehydrogenase level, and the French American-British subtype. An inverse correlation was not observed between Skp2 and p27Kip1 expression. However, p27Kip1 protein was preferentially localized to cytoplasm in the high-Skp2-expression group. The cytoplasmic to nuclear ratio of p27Kip1 expression was significantly correlated with the levels of Skp2 expression (P < 0.001). The frequency of PTEN phosphorylation was significantly higher in the high-Skp2-expression group compared with the low- Skp2-expression group (P = 0.035). The Skp2 overexpression was significantly associated with shorter disease-free survival and overall survival (P = 0.0386 and P = 0.0369, respectively). Multivariate analysis showed that Skp2 expression was an independent prognostic factor both in the disease-free survival and overall survival. CONCLUSION: These findings suggest that Skp2 expression is an independent marker for a poor prognosis in AML. The presence of a positive correlation between Skp2 and phosphorylated PTEN suggests that an aberration in the PTEN/Skp2 signaling pathway might be operating in AML. PMID- 15297416 TI - Promoter hypermethylation of resected bronchial margins: a field defect of changes? AB - PURPOSE: Histologically positive bronchial margins after resection for non-small cell lung cancer are associated with shortened patient survival due to local recurrence. We hypothesized that DNA promoter hypermethylation changes at bronchial margins could be detected in patients with no histological evidence of malignancy and that they would reflect epigenetic events in the primary tumor. EXPERIMENTAL DESIGN: Bronchial margins, primary tumor, bronchoalveolar fluid, and paired nonmalignant lung were obtained from 20 non-small cell lung cancer patients who underwent a lobectomy or greater resection. Disease-specific recurrence was the primary end point. The methylation status of p16, MGMT, DAPK, SOCS1, RASSF1A, COX2, and RARbeta was examined using methylation-specific polymerase chain reaction. RESULTS: All malignancies had methylation in at least one locus. Concordance of one gene with an identical epigenetic change in the tumor or bronchial margin was observed in 85% of patients. Only one patient had methylation at the bronchial margin for a gene that was not methylated in the corresponding tumor. Median time to recurrence was 37 months (range, 5-71 months). There were two local recurrences and five metastases. There were no significant correlations between DNA methylation in tumor, margins, or bronchoalveolar fluid specimens and either regional recurrence or distant metastases. CONCLUSIONS: Histologically negative bronchial margins of resected non-small cell lung cancer exhibit frequent hypermethylation changes in multiple genes. These hyper-methylation abnormalities are also present in the primary tumor and thus may represent a field defect of preneoplastic changes that occurs early in carcinogenesis. PMID- 15297417 TI - Expression of vascular endothelial growth factor receptor-3 by lymphatic endothelial cells is associated with lymph node metastasis in prostate cancer. AB - PURPOSE: The molecular mechanisms underlying lymph node metastasis are poorly understood, despite the well-established clinical importance of lymph node status in many human cancers. Recently, vascular endothelial growth factor (VEGF)-C and VEGF-D have been implicated in the regulation of tumor lymphangiogenesis and enhancement of lymphatic invasion via activation of VEGF receptor-3. The purpose of this study was to determine the expression pattern of the VEGF-C/VEGF-D/VEGF receptor-3 axis in prostate cancer and its relationship with lymph node metastasis. EXPERIMENTAL DESIGN: The expression pattern of VEGF-C, VEGF-D, and VEGF receptor-3 in localized prostate cancer specimens (n = 37) was determined using immunohistochemistry. RESULTS: Widespread, heterogeneous staining for VEGF C and VEGF-D was observed in all cancer specimens. Intensity of VEGF-C staining was lower in benign prostate epithelium than in adjacent carcinoma, whereas no difference between benign epithelium and carcinoma was observed for VEGF-D staining. VEGF receptor-3 immunostaining was detected in endothelial cells of lymphatic vessels in 18 of 37 tissue samples. The presence of VEGF receptor-3 positive vessels was associated with lymph node metastasis (P = 0.0002), Gleason grade (P < 0.0001), extracapsular extension (P = 0.0382), and surgical margin status (P = 0.0069). In addition, VEGF receptor-3 staining highlighted lymphatic invasion by VEGF-C-positive/VEGF-D-positive carcinoma cells. CONCLUSIONS: Together, these results suggest that paracrine activation of lymphatic endothelial cell VEGF receptor-3 by VEGF-C and/or VEGF-D may be involved in lymphatic metastasis. Thus the VEGF-C/VEGF-D/VEGF receptor-3 signaling pathway may provide a target for antilymphangiogenic therapy in prostate cancer. PMID- 15297418 TI - EphA2 expression is associated with aggressive features in ovarian carcinoma. AB - PURPOSE: EphA2 (epithelial cell kinase) is a transmembrane receptor tyrosine kinase that has been implicated in oncogenesis. There are no published data regarding the role of EphA2 in ovarian carcinoma, which is the focus of the present study. EXPERIMENTAL DESIGN: Nontransformed (HIO-180) and ovarian cancer (EG, 222, SKOV3, and A2780-PAR) cell lines were evaluated for EphA2 by Western blot analysis. Five benign ovarian masses, 10 ovarian tumors of low malignant potential, and 79 invasive ovarian carcinomas were also evaluated for EphA2 expression by immunohistochemistry. All samples were scored in a blinded fashion. Univariate and multivariate analyses were used to determine significant associations between EphA2 expression and clinicopathological variables. RESULTS: By Western blot analysis, EG, 222, and SKOV3 cell lines overexpressed EphA2, whereas A2780-PAR and HIO-180 had low to absent EphA2 expression. All of the benign tumors had low or absent EphA2 expression. Among the invasive ovarian carcinomas examined (mean age of patients was 59.2 years), 60 (75.9%) tumors overexpressed EphA2 and the other 19 tumors had negative or minimal EphA2 expression. There was no association of EphA2 overexpression with ascites, likelihood of nodal positivity, pathological subtype, and optimum surgical cytoreduction (residual tumor <1 cm). However, EphA2 overexpression was significantly associated with higher tumor grade (P = 0.02) and advanced stage of disease (P = 0.001). The median survival for patients with tumor EphA2 overexpression was significantly shorter (median, 3.1 years; P = 0.004); the median survival for patients with low or absent EphA2 tumor expression was at least 12 years and has not yet been reached. In multivariate analysis using the Cox proportional hazards model, only volume of residual disease (P < 0.04) and EphA2 overexpression (P < 0.01) were significant and independent predictors of survival. CONCLUSIONS: EphA2 overexpression is predictive of aggressive ovarian cancer behavior and may be an important therapeutic target. PMID- 15297419 TI - Relevance of different UGT1A1 polymorphisms in irinotecan-induced toxicity: a molecular and clinical study of 75 patients. AB - PURPOSE: We wanted to assess polymorphisms in the uridine diphosphoglucuronosyl transferase 1A1 (UGT 1A1) gene: the TATA box polymorphism and UGT 1A1 G71R and Y486D mutations in the coding sequence, the main mutations characterizing Gilbert's syndrome, as predictors of severe toxic event occurrence after irinotecan (CPT-11) administration. Therefore, we set up a rapid, sensitive, and reliable technique in routine practice to detect before CPT-11 treatment, the at risk patients. EXPERIMENTAL DESIGN: Seventy-five patients with advanced colorectal cancer and treated with CPT-11 and 5-fluorouracil, entered the study. We used the Pyrosequencing technology a real-time sequencing method, to detect the UGT 1A1 TATA box polymorphisms and mutations in the coding regions. Patients were also assessed for both biochemical and clinical evaluation and tolerance to treatment. RESULTS: No G71R and Y486D mutations were found in our population. Frequencies for UGT 1A1 TATA box polymorphisms were 41, 47, and 9% for wild-type 6/6, heterozygous 6/7, and Gilbert's syndrome 7/7, respectively. Tolerance to treatment decreased with increased number of TA repeat with 71% of the patients in 7/7 group who experienced grade 3/4 toxicity. CONCLUSIONS: The method we set up is suitable for the detection of UGT 1A1 polymorphism in routine practice before irinotecan treatment. It could help to detect the patients homozygous or heterozygous for Gilbert's syndrome, at-risk of CPT 11-induced toxicity, and thus could help to individualize the dose to optimize efficacy and limit toxicity. PMID- 15297420 TI - Patterns of chromosomal alterations in breast ductal carcinoma in situ. AB - PURPOSE: Ductal carcinoma in situ (DCIS) is thought to be a nonobligate precursor of invasive cancer. Genomic changes specific to pure DCIS versus invasive cancer, as well as alterations unique to individual DCIS subtypes, have not been fully defined. EXPERIMENTAL DESIGN: Chromosomal copy number alterations were examined by comparative genomic hybridization in 34 cases of pure DCIS and compared with 12 cases of paired synchronous DCIS and invasive ductal cancer, as well as to 146 additional cases of invasive breast cancer of ductal or lobular histology. Genomic differences between high-grade and low/intermediate-grade DCIS, as well as between pure DCIS and invasive cancer, were identified. RESULTS: Pure DCIS showed almost the same degree of chromosomal instability as invasive ductal cancers. A higher proportion of low/intermediate-grade versus high-grade DCIS had loss of 16q (65 versus 12%, respectively; P = 0.002). When compared with lower grade DCIS, high-grade DCIS exhibited more frequent gain of 17q (65 versus 41%; P = 0.15) and higher frequency loss of 8p (77 versus 41%; P = 0.04). Chromosomal alterations in those cases with synchronous DCIS and invasive ductal cancer showed a high degree of shared changes within the two components. CONCLUSIONS: DCIS is genetically advanced, showing a similar degree of chromosomal alterations as invasive ductal cancer. The pattern of alterations differed between high- and low/intermediate-grade DCIS, supporting a model in which different histological grades of DCIS are associated with distinct genomic changes. These regions of chromosomal alterations may be potential targets for treatment and/or markers of prognosis. PMID- 15297421 TI - Cyclin D1, p53, and p21Waf1/Cip1 expression is predictive of poor clinical outcome in serous epithelial ovarian cancer. AB - PURPOSE: Dysregulation of cell cycle control, in particular G(1)-S-phase transition, is implicated in the pathogenesis of most human cancers, including epithelial ovarian cancer (EOC). However, the prognostic significance of aberrant cell cycle gene expression in EOC remains unclear. EXPERIMENTAL DESIGN: The expression of selected genes from the pRb pathway that regulates G(1)-S-phase progression, including cyclin D1, p16(Ink4a), cyclin E, p27(Kip1), p21(Waf1/Cip1), and p53, was examined in a consecutive series of 134 serous EOC using immunohistochemistry and the results correlated to disease outcome. RESULTS: Molecular markers predictive of reduced overall survival in univariate analysis were overexpression of cyclin D1 (P = 0.03) and p53 (P = 0.03) and reduced expression of p27(Kip1) (P = 0.05) and p21(Waf1/Cip1) (P = 0.02), with the latter three also being prognostic for a shorter progression-free interval. In addition, patients displaying overexpression of p53 with concurrent loss of p21(Waf1/Cip1) had a significantly shorter overall (P = 0.0008) and progression free survival (P = 0.0001). On multivariate analysis, overexpression of cyclin D1 and combined loss of p21(Waf1/Cip1) in the presence of p53 overexpression were independent predictors of overall survival. Similarly, the combination of p21(Waf1/Cip1) loss and p53 overexpression was independently predictive of a shorter progression-free interval. Overexpression of p53 and cyclin E and reduced expression of p27(Kip1) and p21(Waf1/Cip1) were significantly associated with increasing tumor grade. CONCLUSIONS: This study confirms that dysregulation of cell cycle genes is common in EOC, and that aberrant expression of critical cell cycle regulatory proteins can predict patient outcome in serous EOC. PMID- 15297422 TI - Distribution and clinical significance of heparan sulfate proteoglycans in ovarian cancer. AB - PURPOSE: Heparan sulfate proteoglycans have been implicated in cancer cell growth, invasion, metastasis, and angiogenesis. This study was designed to compare their expression in normal ovary and ovarian tumors and then to examine their prognostic significance in ovarian cancer. EXPERIMENTAL DESIGN: The expression of syndecan-1, -2, -3, and -4, glypican-1, and perlecan was assessed by immunohistochemistry in 147 biopsies that included normal ovary and benign, borderline, and malignant ovarian tumors. Clinical data, including tumor stage, performance status, treatment, and survival, were collected. Univariate and multivariate analyses were performed to evaluate prognostic significance. RESULTS: The expression patterns of syndecan-1 and perlecan were altered in ovarian tumors compared with normal ovary. Syndecan-1 was not detected in normal ovary but was present in the epithelial and stromal cells of benign and borderline tumors and in ovarian adenocarcinomas. Perlecan expression was decreased in basement membranes that were disrupted by cancer cells but maintained in the basement membranes of blood vessels. Syndecan-2, -3, and -4, and glypican-1 were expressed in normal ovary and benign and malignant ovarian tumors. Stromal expression of syndecan-1 and glypican-1 were poor prognostic factors for survival in univariate analysis. CONCLUSION: We report for the first time distinct patterns of expression of cell surface and extracellular matrix heparan sulfate proteoglycans in normal ovary compared with ovarian tumors. These data reinforce the role of the tumor stroma in ovarian adenocarcinoma and suggest that stromal induction of syndecan-1 contributes to the pathogenesis of this malignancy. PMID- 15297423 TI - Noscapine crosses the blood-brain barrier and inhibits glioblastoma growth. AB - The opium alkaloid noscapine is a commonly used antitussive agent available in Europe, Asia, and South America. Although the mechanism by which it suppresses coughing is currently unknown, it is presumed to involve the central nervous system. In addition to its antitussive action, noscapine also binds to tubulin and alters microtubule dynamics in vitro and in vivo. In this study, we show that noscapine inhibits the proliferation of rat C6 glioma cells in vitro (IC(50) = 100 microm) and effectively crosses the blood-brain barrier at rates similar to the ones found for agents such as morphine and [Met]enkephalin that have potent central nervous system activity (P < or = 0.05). Daily oral noscapine treatment (300 mg/kg) administered to immunodeficient mice having stereotactically implanted rat C6 glioblasoma into the striatum revealed a significant reduction of tumor volume (P < or = 0.05). This was achieved with no identifiable toxicity to the duodenum, spleen, liver, or hematopoietic cells as determined by pathological microscopic examination of these tissues and flow cytometry. Furthermore, noscapine treatment resulted in little evidence of toxicity to dorsal root ganglia cultures as measured by inhibition of neurite outgrowth and yielded no evidence of peripheral neuropathy in animals. However, evidence of vasodilation was observed in noscapine-treated brain tissue. These unique properties of noscapine, including its ability to cross the blood-brain barrier, interfere with microtubule dynamics, arrest tumor cell division, reduce tumor growth, and minimally affect other dividing tissues and peripheral nerves, warrant additional investigation of its therapeutic potential. PMID- 15297424 TI - CD95-mediated apoptosis is impaired at receptor level by cellular FLICE inhibitory protein (long form) in wild-type p53 human ovarian carcinoma. AB - PURPOSE: Ovarian carcinoma is a highly lethal malignancy that often becomes resistant to chemotherapy. Alterations in apoptotic signals and p53 status contribute to drug resistance, and CD95-mediated apoptosis is also deficient in resistant cells. We analyzed the mechanism of resistance to CD95-mediated apoptosis in ovarian carcinoma cell lines differing in p53 status. EXPERIMENTAL DESIGN: CD95-mediated apoptosis was induced by agonistic anti-CD95 antibody, and the apoptotic cascade was monitored with biochemical and functional assays. RESULTS: CD95-mediated apoptosis was blocked in human ovarian cancer cells. In cell lines with wild-type p53, treatment with the protein synthesis inhibitor cycloheximide (CHX) together with anti-CD95 overcame the resistance, suggesting the presence of a labile inhibiting protein. Indeed, the labile protein cellular FLICE-inhibitory protein long form (c-FLIP(L)) was found to block caspase-8 recruitment to the death-inducing signaling complex (DISC), and sensitization of cells by CHX was due to c-FLIP(L) down-modulation at the DISC level. Down regulation of c-FLIP(L) with antisense oligonucleotides increased CD95-mediated apoptosis as in cells sensitized by CHX, demonstrating the direct involvement of c-FLIP(L) in apoptosis resistance. Removal of c-FLIP(L) block at DISC level allowed full activation of the mitochondrial pathway and, eventually, apoptosis in wild-type p53 cells, whereas in cells with mutated p53, c-FLIP(L) involvement in CD95-mediated apoptosis resistance appeared to be irrelevant. Immunohistochemical analysis of an ovarian tumor tissue array revealed c-FLIP(L) expression in samples with no p53 accumulation (P = 0.034), and a significant (P = 0.037) inverse relationship between c-FLIP(L) and p53 expression levels was also observed in 27 epithelial ovarian cancer specimens with known p53 status. CONCLUSION: The inhibitory protein c-FLIP(L) is involved in resistance to CD95 mediated apoptosis in ovarian carcinoma cells with wild-type p53. PMID- 15297425 TI - Mifepristone induces growth arrest, caspase activation, and apoptosis of estrogen receptor-expressing, antiestrogen-resistant breast cancer cells. AB - PURPOSE: A major clinical problem in the treatment of breast cancer is the inherent and acquired resistance to antiestrogen therapy. In this study, we sought to determine whether antiprogestin treatment, used as a monotherapy or in combination with antiestrogen therapy, induced growth arrest and active cell death in antiestrogen-resistant breast cancer cells. EXPERIMENTAL DESIGN: MCF-7 sublines were established from independent clonal isolations performed in the absence of drug selection and tested for their response to the antiestrogens 4 hydroxytamoxifen (4-OHT) and ICI 182,780 (fulvestrant), and the antiprogestin mifepristone (MIF). The cytostatic (growth arrest) effects of the hormones were assessed with proliferation assays, cell counting, flow cytometry, and a determination of the phosphorylation status of the retinoblastoma protein. The cytotoxic (apoptotic) effects were analyzed by assessing increases in caspase activity and cleavage of poly(ADP-ribose) polymerase. RESULTS: All of the clonally derived MCF-7 sublines expressed estrogen receptor and progesterone receptor but showed a wide range of antiestrogen sensitivity, including resistance to physiological levels of 4-OHT. Importantly, all of the clones were sensitive to the antiprogestin MIF, whether used as a monotherapy or in combination with 4-OHT. MIF induced retinoblastoma activation, G(1) arrest, and apoptosis preceded by caspase activation. CONCLUSIONS: We demonstrate that: (a) estrogen receptor(+)progesterone receptor(+), 4-OHT-resistant clonal variants can be isolated from an MCF-7 cell line in the absence of antiestrogen selection; and (b) MIF and MIF plus 4-OHT combination therapy induces growth arrest and active cell death of the antiestrogen-resistant breast cancer cells. These preclinical findings show potential for a combined hormonal regimen of an antiestrogen and an antiprogestin to combat the emergence of antiestrogen-resistant breast cancer cells and, ultimately, improve the therapeutic index of antiestrogen therapy. PMID- 15297426 TI - The effect of megestrol acetate on growth of HepG2 cells in vitro and in vivo. AB - PURPOSE: Hepatocellular carcinoma (HCC) is generally considered as a sex hormone dependent tumor, and hormonal therapy has been proposed as a strategy for the treatment of HCC. The aim of the study is to investigate the effect of megestrol acetate, a synthetic progesteronal agent, on growth of HepG2 cells in vitro and in vivo. EXPERIMENTAL DESIGN: Cell growth in vitro was assessed by a colormetric method, and cell growth in vivo was assessed by tumor volumetrics. RESULTS: Megestrol acetate was shown to inhibit the growth of HepG2 cells in vitro in dose and time-dependent manners with an IC (50) of 260 microm (24-h incubation). The growth of HepG2 cell-transplanted tumors in nude mice was also inhibited by i.p. injection of megestrol acetate (10 mg/kg/day). The tumor volumes of the megestrol acetate-treated group regressed to 59% of controls by week 6 and to 41% of controls by week 13. Apoptosis following G(1) arrest was observed in megestrol acetate-treated cells and may be a mechanism through which megestrol acetate inhibits HepG2 cells. Megestrol acetate was also demonstrated to have a beneficial effect on the weight gain of tumor-bearing nude mice, and the mean weight of the megestrol acetate-treated animals was higher than that of controls from week 4 of the treatment period, and the differences were statistically significant in week 5 and 6 (P < 0.05, compared with controls). No significant survival advantage was, however, demonstrated in the treatment group. CONCLUSIONS: This study showed that megestrol acetate inhibited the growth of HepG2 cells grown in vitro and in vivo. These data provide useful information for clinical study of megestrol acetate for the treatment of HCC. PMID- 15297427 TI - Pharmacokinetics and tissue disposition of indole-3-carbinol and its acid condensation products after oral administration to mice. AB - Indole-3-carbinol (I3C) and 3,3'-diindolylmethane (DIM) are promising cancer chemopreventive agents in rodent models, but there is a paucity of data on their pharmacokinetics and tissue disposition. The disposition of I3C and its acid condensation products, DIM, [2-(indol-3-ylmethyl)-indol-3-yl]indol-3-ylmethane (LTr(1)), indolo[3,2b]carbazole (ICZ) and 1-(3-hydroxymethyl)-indolyl-3 indolylmethane (HI-IM) was studied, after oral administration of I3C (250 mg/kg) to female CD-1 mice. Blood, liver, kidney, lung, heart, and brain were collected between 0.25 and 24 h after administration and the plasma and tissue concentrations of I3C and its derivatives determined by high-performance liquid chromotography. I3C was rapidly absorbed, distributed, and eliminated from plasma and tissues, falling below the limit of detection by 1 h. Highest concentrations of I3C were detected in the liver where levels were approximately 6-fold higher than those in the plasma. Levels of DIM, LTr(1), and HI-IM were much lower, although they persisted in plasma and tissues for considerably longer. DIM and HI IM were still present in the liver 24 h after I3C administration. Tissue levels of DIM and LTr(1) were found to be in equilibrium with plasma at almost every time point measured. In addition to acid condensation products of I3C, a major oxidative metabolite (indole-3-carboxylic acid) and a minor oxidative metabolite (indole-3-carboxaldehyde) were detected in plasma of mice after oral administration of I3C. ICZ was also tentatively identified in the liver of these mice. This study shows for the first time that, after oral administration to mice, I3C, in addition to its acid condensation products, is absorbed from the gut and distributed systemically into a number of well-perfused tissues, thus allowing the possibility for some pharmacological activity of the parent compound in vivo. PMID- 15297428 TI - In vitro combination treatment with perifosine and UCN-01 demonstrates synergism against prostate (PC-3) and lung (A549) epithelial adenocarcinoma cell lines. AB - PURPOSE: Antineoplastic agents often achieve antitumor activity at the expense of close to unacceptable toxicity. One potential avenue to improve therapeutic index might combine agents targeting distinct components of the same growth regulatory pathway. This might lead to more complete modulation of the target pathway at concentrations lower than those associated with limiting adventitious toxicities from either agent alone. The protein kinase antagonist UCN-01 is currently used in Phase I/II trials and has recently been demonstrated to inhibit potently PDK1. We have recently documented that the alkylphospholipid perifosine potently also inhibits Akt kinase (PKB) activation by interfering with membrane localization of Akt. This leads to the hypothesis that these two agents might act synergistically through distinct mechanisms in the PI3K/Akt proliferation and survival-related signaling pathway. EXPERIMENTAL DESIGN: The synergistic effects of UCN-01 and perifosine, on two cell lines (A-549 and PC-3), were examined using various long term in vitro assays for cell growth, cell cycle distribution, clonogenicity, survival morphology, and apoptosis. Along with Western blotting experiments were performed to determine whether this synergistic combination of two drugs has significant effect on their downstream targets and on biochemical markers of apoptosis. RESULTS: After 72 h, perifosine at concentrations of 1.5 and 10 microM UCN-01 at 40 and 250 nM did not significantly affect the growth of PC-3 and A459 cells, respectively. However, in combination at the same respective individual concentrations (1.5 microM and 40 nM of perifosine and UCN-01, respectively, in PC-3 cells and 10 microM perifosine and 0.25 microM UCN-01 in the somewhat more resistant A549 cells), virtually complete growth inhibition of both the cell lines resulted. Supra-additive inhibition of growth was also demonstrated in independent clonogenic assays. Mechanistic studies in cell culture models suggest enhanced depletion of the S-phase population in cells treated by the combination. This correlated with enhanced inactivation of Akt along with activation of caspases 3 and 9 and poly(ADP-ribose) polymerase cleavage. Evidence of synergy was formally demonstrated and occurred across a wide range of drug concentrations and was largely independent of the order or sequence of drug addition. CONCLUSIONS: As the concentrations of UCN-01 and perifosine causing synergistic inhibition of cell growth are clinically achievable without prominent toxicity, these data support the development of clinical studies with this combination. PMID- 15297429 TI - trans-3,4,5'-Trihydroxystibene inhibits hypoxia-inducible factor 1alpha and vascular endothelial growth factor expression in human ovarian cancer cells. AB - trans-3,4,5'-Trihydroxystibene (resveratrol) is a natural product commonly found in the human diet and has been shown recently to have anticancer effects on various human cancer cells. However, the molecular basis for its anticancer action remains to be elucidated. In this study, we investigated the effect of resveratrol on hypoxia-inducible factor 1alpha (HIF-1alpha) and vascular endothelial growth factor (VEGF) expression in human ovarian cancer cells A2780/CP70 and OVCAR-3. We found that although resveratrol did not affect HIF 1alpha mRNA levels, it did dramatically inhibit both basal-level and growth factor-induced HIF-1alpha protein expression in the cells. Resveratrol also greatly inhibited VEGF expression. Mechanistically, we demonstrated that resveratrol inhibited HIF-1alpha and VEGF expression through multiple mechanisms. First, resveratrol inhibited AKT and mitogen-activated protein kinase activation, which played a partial role in the down-regulation of HIF-1alpha expression. Second, resveratrol inhibited insulin-like growth factor 1-induced HIF-1alpha expression through the inhibition of protein translational regulators, including M(r) 70,000 ribosomal protein S6 kinase 1, S6 ribosomal protein, eukaryotic initiation factor 4E-binding protein 1, and eukaryotic initiation factor 4E. Finally, we showed that resveratrol substantially induced HIF-1alpha protein degradation through the proteasome pathway. Our data suggested that resveratrol may inhibit human ovarian cancer progression and angiogenesis by inhibiting HIF 1alpha and VEGF expression and thus provide a novel potential mechanism for the anticancer action of resveratrol. PMID- 15297430 TI - Nitric oxide accelerates interleukin-13 cytotoxin-mediated regression in head and neck cancer animal model. AB - Receptors for interleukin-13 (IL-13R) are overexpressed on several types of solid cancers including gliobastoma, renal cell carcinoma, AIDS Kaposi's sarcoma, and head and neck cancer. Recombinant fusion proteins IL-13 cytotoxin (IL13-PE38QQR or IL13-PE38) have been developed to directly target IL-13R-expressing cancer cells. Although it has been found that IL-13 cytotoxin has a direct potent antitumor activity in vivo in nude mice models of human cancers, the involvement of indirect antitumor effecter molecules such as nitric oxide (NO) is unknown. To address this issue, we assessed the effect of NO inhibiter N(omega)-monomethyl-l arginine on IL-13 cytotoxin-mediated cytotoxicity and NO2/NO3 production in HN12 head and neck cancer cells. In addition, antitumor effects and NO levels in HN12 and KCCT873 head and neck tumors xenografted s.c. in nude mice when treated with IL-13 cytotoxin were evaluated by tumor measurement, Western blot, and immunohistochemistry analyses. Pretreatment of animals with N(omega)-monomethyl-l arginine significantly decreased the NO levels and IL-13 cytotoxin-mediated antitumor effects. In addition, depletion of macrophages, known to produce NO, also decreased antitumor activity of IL-13 cytotoxin. Based on these studies, we concluded that NO accelerates antitumor effect of IL-13 cytotoxin on head and neck tumor cells. Because IL-13 cytotoxin is currently being tested in the clinic for the treatment of patients with recurrent glioblastoma maltiforme, our current findings suggest maintaining macrophage and NO-producing cellular function for optimal therapeutic effect of this targeted agent. PMID- 15297431 TI - Class I histone deacetylase-selective novel synthetic inhibitors potently inhibit human tumor proliferation. AB - We have developed previously a class of synthetic hybrid histone deacetylase (HDAC) inhibitors, which were built from hydroxamic acid of trichostatin A and pyridyl ring of MS-275. In this study we evaluated the antitumor effects of these novel hybrid synthetic HDAC inhibitors, SK-7041 and SK-7068, on human cancer cells. Both SK-7041 and SK-7068 effectively inhibited cellular HDAC activity at nanomolar concentrations and induced the time-dependent hyperacetylation of histones H3 and H4. These HDAC inhibitors preferentially inhibited the enzymatic activities of HDAC1 and HDAC2, as compared with the other HDAC isotypes, indicating that class I HDAC is the major target of SK-7041 and SK-7068. We found that these compounds exhibited potent antiproliferative activity against various human cancer cells in vitro. Growth inhibition effect of SK-7041 and SK-7068 was related with the induction of aberrant mitosis and apoptosis in human gastric cancer cells. Both compounds induced the accumulation of cells at mitosis after 6 h of treatment, which was demonstrated by accumulation of tetraploid cells, lack of G(2) cyclin/cyclin-dependent kinase inactivation, and higher mitotic index. After 12 h of treatment, apoptotic cells were increased through mitochondrial and caspase-mediated pathway. Finally, in vivo experiment showed that SK-7041 or SK 7068 was found to reduce the growth of implanted human tumors in nude mice. Therefore, based on isotype specificity and antitumor activity, SK-7041 and SK 7068 HDAC inhibitors are expected to be promising anticancer therapeutic agents and need additional clinical development. PMID- 15297432 TI - A concentrated aglycone isoflavone preparation (GCP) that demonstrates potent anti-prostate cancer activity in vitro and in vivo. AB - PURPOSE: Isoflavones have anticancer activities, but naturally occurring isoflavones are predominantly glycosylated and poorly absorbed. Genistein combined polysaccharide (GCP; Amino Up Chemical Co., Sapporo, Japan), is a fermentation product of soy extract and basidiomycetes mycillae that is enriched in biologically active aglycone isoflavones. This study analyzes GCP in vitro and in vivo for potential utility as a prostate cancer chemopreventative agent. EXPERIMENTAL DESIGN: Androgen-sensitive LNCaP and androgen-independent PC-3 cells were grown with various concentrations of GCP. In vitro cell growth was analyzed by the WST-1 assay, and apoptosis was assessed by fluorescence-activated cell sorting and detection of poly(ADP-ribose) polymerase cleavage using Western blot techniques. Effects of GCP on expression of cell cycle-regulatory proteins p53 (LNCaP only), p21, and p27 and the protein kinase Akt were considered using Western blot techniques. An in vivo LNCaP xenograft model was used to study the effects of a 2% GCP-supplemented diet on tumor growth in comparison with a control diet. RESULTS: GCP significantly suppressed LNCaP and PC-3 cell growth over 72 h (89% and 78% in LNCaP and PC-3, respectively, at 10 microg/ml; P < 0.0001). This reduction was associated with apoptosis in LNCaP cells, but not in PC-3 cells. GCP induced p27 and p53 (LNCaP only) protein expression within 6 h and suppressed phosphorylated Akt in both cell lines. The 2% GCP-supplemented diet significantly slowed LNCaP tumor growth, increasing apoptosis (P < 0.001), and decreasing proliferation (P < 0.001) over 4 weeks. CONCLUSIONS: GCP has potent growth-inhibitory effects against prostate cancer cell lines in vitro and in vivo. These data suggest GCP has potential as an effective chemopreventive agent against prostate cancer cell growth. PMID- 15297433 TI - Identification of serum amyloid a protein as a potentially useful biomarker for nasopharyngeal carcinoma. PMID- 15297434 TI - Testosterone replacement therapy: current trends and future directions. AB - Male hypogonadism is characterized by abnormally low serum testosterone levels associated with typical symptoms, including mood disturbance, sexual dysfunction, decreased muscle mass and strength, and decreased bone mineral density. By restoring serum testosterone levels to the normal range using testosterone replacement therapy, many of these symptoms can be relieved. For many years, injectable testosterone esters or surgically implanted testosterone pellets have been the preferred treatment for male hypogonadism. Recently, newer treatment modalities have been introduced, including transdermal patches and gels. The development of a mucoadhesive sustained-release buccal tablet is the latest innovation, which will provide patients with an additional option. The availability of new treatment modalities has helped to renew interest in the management of male hypogonadism, highlighting the need to address a number of important but previously neglected questions in testosterone replacement therapy. These include the risks and benefits of treatment in different patient populations (e.g. the elderly) and the need for evidence-based diagnosis and treatment monitoring guidelines. While some recommendations have been developed in individual countries, up-to-date, internationally accepted evidence-based guidelines that take into account national differences in clinical practice and healthcare delivery would optimize patient care universally. PMID- 15297435 TI - Androgen insufficiency in women: diagnostic and therapeutic implications. AB - The proposed key symptoms of the female androgen insufficiency syndrome (FAIS) include reduced libido, diminished well being and lowered mood. The diagnosis of FAIS is made on the basis of these symptoms in the setting of a low serum free testosterone level. However, there is currently no readily available inexpensive assay which reliably measures free testosterone levels in the female range. The diagnosis of FAIS is further complicated by the lack of data demonstrating a minimum serum free testosterone level which, if below this, correlates with the symptoms of FAIS. Despite the complexities involved with defining FAIS, the symptoms have been reported to respond well to testosterone replacement. There is a need for formulations of testosterone therapy specifically designed for use in women, along with clear guidelines regarding optimal therapeutic doses and long term safety data. PMID- 15297436 TI - Triiodothyronine stimulates food intake via the hypothalamic ventromedial nucleus independent of changes in energy expenditure. AB - Increased food intake is characteristic of hyperthyroidism, although this is presumed to compensate for a state of negative energy balance. However, here we show that the thyroid hormone T(3) directly stimulates feeding at the level of the hypothalamus. Peripheral administration of T(3) doubled food intake in ad libitum-fed rats over 2 h and induced expression of the immediate early gene, early growth response-1, in the hypothalamic ventromedial nucleus (VMN), whereas maintaining plasma-free T(3) levels within the normal range. T(3)-induced feeding occurred without altering energy expenditure or locomotion. Injection of T(3) directly into the VMN produced a 4-fold increase in food intake in the first hour. The majority of T(3) in the brain is reported to be produced by tissue specific conversion of T(4) to T(3) by the enzyme type 2 iodothyronine deiodinase (D2). Hypothalamic D2 mRNA expression showed a diurnal variation, with a peak in the nocturnal feeding phase. Hypothalamic D2 mRNA levels also increased after a 12- and 24-h fast, suggesting that local production of T(3) may play a role in this T(3) feeding circuit. Thus, we propose a novel hypothalamic feeding circuit in which T(3), from the peripheral circulation or produced by local conversion, stimulates food intake via the VMN. PMID- 15297437 TI - Mechanisms of hemorrhage-induced hepatic insulin resistance: role of tumor necrosis factor-alpha. AB - Hemorrhage, sepsis, burn injury, surgical trauma and critical illness all induce insulin resistance. Recently we found that trauma and hemorrhage acutely induced hepatic insulin resistance in the rat. However, the mechanisms of this hemorrhage induced acute hepatic insulin resistance are unknown. Here we report on the mechanisms of this hepatic insulin resistance. Protein levels and phosphorylation of the insulin receptor and insulin receptor substrate-1/2 (IRS-1/2) were measured, as was the association between IRS-1/2 and phosphatidylinositol 3 kinase (PI3K). Also examined were the hepatic expression of TNFalpha and TNFalpha induced serine phosphorylation of IRS-1. Insulin receptor and IRS-1/2 protein levels and insulin-induced tyrosine phosphorylation of the insulin receptor were unaltered. In contrast, insulin-induced tyrosine phosphorylation of IRS-1/2 and association between IRS-1/2 and PI3K were dramatically reduced after hemorrhage. Hepatic levels of TNFalpha mRNA and protein were increased as was phosphorylation of IRS-1 serine 307 after hemorrhage. Our data provide the first evidence that compromised IRS-1/2 tyrosine phosphorylation and their association with PI3K contribute to hemorrhage-induced acute hepatic insulin resistance. Increased local TNFalpha may play a role in inducing this hepatic insulin resistance after trauma and hemorrhage. PMID- 15297438 TI - Free fatty acids and cytokines induce pancreatic beta-cell apoptosis by different mechanisms: role of nuclear factor-kappaB and endoplasmic reticulum stress. AB - Apoptosis is probably the main form of beta-cell death in both type 1 diabetes mellitus (T1DM) and T2DM. In T1DM, cytokines contribute to beta-cell destruction through nuclear factor-kappaB (NF-kappaB) activation. Previous studies suggested that in T2DM high glucose and free fatty acids (FFAs) are beta-cell toxic also via NF-kappaB activation. The aims of this study were to clarify whether common mechanisms are involved in FFA- and cytokine-induced beta-cell apoptosis and determine whether TNFalpha, an adipocyte-derived cytokine, potentiates FFA toxicity through enhanced NF-kappaB activation. Apoptosis was induced in insulinoma (INS)-1E cells, rat islets, and fluorescence-activated cell sorting purified beta-cells by oleate, palmitate, and/or cytokines (IL-1beta, interferon gamma, TNFalpha). Palmitate and IL-1beta induced a similar percentage of apoptosis in INS-1E cells, whereas oleate was less toxic. TNFalpha did not potentiate FFA toxicity in primary beta-cells. The NF-kappaB-dependent genes inducible nitric oxide synthase and monocyte chemoattractant protein-1 were induced by IL-1beta but not by FFAs. Cytokines activated NF-kappaB in INS-1E and beta-cells, but FFAs did not. Moreover, FFAs did not enhance NF-kappaB activation by TNFalpha. Palmitate and oleate induced C/EBP homologous protein, activating transcription factor-4, and immunoglobulin heavy chain binding protein mRNAs, X box binding protein-1 alternative splicing, and activation of the activating transcription factor-6 promoter in INS-1E cells, suggesting that FFAs trigger an endoplasmic reticulum (ER) stress response. We conclude that apoptosis is the main mode of FFA- and cytokine-induced beta-cell death but the mechanisms involved are different. Whereas cytokines induce NF-kappaB activation and ER stress (secondary to nitric oxide formation), FFAs activate an ER stress response via an NF-kappaB- and nitric oxide-independent mechanism. Our results argue against a unifying hypothesis for the mechanisms of beta-cell death in T1DM and T2DM. PMID- 15297439 TI - Development and application of a rat ovarian gene expression database. AB - The pituitary gonadotropins play a key role in follicular development and ovulation through the induction of specific genes. To identify these genes, we have constructed a genome-wide rat ovarian gene expression database (rOGED). The database was constructed from total RNA isolated from intact ovaries, granulosa cells, or residual ovarian tissues collected from immature pregnant mare serum gonadotropin (PMSG)/human chorionic gonadotropin-treated rats at 0 h (no PMSG), 12 h, and 48 h post PMSG, as well as 6 and 12 h post human chorionic gonadotropin. The total RNA was used for DNA microarray analysis using Affymetrix Rat Expression Arrays 230A and 230B (Affymetrix, Santa Clara, CA). The microarray data were compiled and used for display of individual gene expression profiles through specially developed software. The final rOGED provides immediate analysis of temporal gene expression profiles for over 28,000 genes in intact ovaries, granulosa cells, and residual ovarian tissue during follicular growth and the preovulatory period. The accuracy of the rOGED was validated against the gene profiles for over 20 known genes. The utility of the rOGED was demonstrated by identifying six genes that have not been described in the rat periovulatory ovary. The mRNA expression patterns and cellular localization for each of these six genes (estrogen sulfotransferase, synaptosomal-associated protein 25 kDa, runt-related transcription factor, calgranulin B, alpha1-macroglobulin, and MAPK phosphotase-3) were confirmed by Northern blot analyses and in situ hybridization, respectively. The current findings demonstrate that the rOGED can be used as an instant reference for ovarian gene expression profiles, as well as a reliable resource for identifying important yet, to date, unknown ovarian genes. PMID- 15297440 TI - Inhibition of hepatic gluconeogenesis and enhanced glucose uptake contribute to the development of hypoglycemia in mice bearing interleukin-1beta- secreting tumor. AB - Mice bearing IL-1beta-secreting tumor were used to study the chronic effect of IL 1beta on glucose metabolism. Mice were injected with syngeneic tumor cells transduced with the human IL-1beta gene. Serum IL-1beta levels increased exponentially with time. Secretion of IL-1beta from the developed tumors was associated with decreased food consumption, reduced body weight, and reduced blood glucose levels. Body composition analysis revealed that IL-1beta caused a significant loss in fat tissue without affecting lean body mass and water content. Hepatic phosphoenolpyruvate carboxykinase and glucose-6-phosphatase activities and mRNA levels of these enzymes were reduced, and 2-deoxy-glucose uptake by peripheral tissues was enhanced. mRNA levels of glucose transporters (Gluts) in the liver were determined by real-time PCR analysis. Glut-3 mRNA levels were up-regulated by IL-1beta. Glut-1 and Glut-4 mRNA levels in IL-1beta mice were similar to mRNA levels in pair-fed mice bearing nonsecreting tumor. mRNA level of Glut-2, the major Glut of the liver, was down-regulated by IL 1beta. We concluded that both decreased glucose production by the liver and enhanced glucose disposal lead to the development of hypoglycemia in mice bearing IL-1beta-secreting tumor. The observed changes in expression of hepatic Gluts that are not dependent on insulin may contribute to the increased glucose uptake. PMID- 15297441 TI - Regulation of bone mass and bone turnover by neuronal nitric oxide synthase. AB - Nitric oxide (NO) is produced by NO synthase (NOS) and plays an important role in the regulation of bone cell function. The endothelial NOS isoform is essential for normal osteoblast function, whereas the inducible NOS isoform acts as a mediator of cytokine effects in bone. The role of the neuronal isoform of NOS (nNOS) in bone has been studied little thus far. Therefore, we investigated the role of nNOS in bone metabolism by studying mice with targeted inactivation of the nNOS gene. Bone mineral density (BMD) was significantly higher in nNOS knockout (KO) mice compared with wild-type controls, particularly the trabecular BMD (P < 0.01). The difference in BMD between nNOS KO and control mice was confirmed by histomorphometric analysis, which showed a 67% increase in trabecular bone volume in nNOS KO mice when compared with controls (P < 0.001). This was accompanied by reduced bone remodeling, with a significant reduction in osteoblast numbers and bone formation surfaces and a reduction in osteoclast numbers and bone resorption surfaces. Osteoblasts from nNOS KO mice, however, showed increased levels of alkaline phosphatase and no defects in proliferation or bone nodule formation in vitro, whereas osteoclastogenesis was increased in nNOS KO bone marrow cultures. These studies indicate that nNOS plays a hitherto unrecognized but important physiological role as a stimulator of bone turnover. The low level of nNOS expression in bone and the in vitro behavior of nNOS KO bone cells indicate that these actions are indirect and possibly mediated by a neurogenic relay. PMID- 15297442 TI - Identification and characterization of the zebrafish and fugu genes encoding tuberoinfundibular peptide 39. AB - Although the PTH type 2 receptor (PTH2R) has been isolated from mammals and zebrafish, only its mammalian agonist, tuberoinfundibular peptide 39 (TIP39), has been characterized thus far. To determine whether zebrafish TIP39 (zTIP39) functions similarly with the zebrafish PTHR (zPTH2R) and human PTH2Rs and to determine its tissue-specific expression, fugu (Takifugu rubripes) and zebrafish (Danio rerio) genomic databases were screened with human TIP39 (hTIP39) sequences. A single TIP39 gene was identified for each fish species, which showed significant homology to mammalian TIP39. Using standard molecular techniques, we isolated cDNA sequences encoding zTIP39. The fugu TIP39 precursor was encoded by a gene comprising at least three exons. It contained a hydrophobic signal sequence and a predicted prosequence with a dibasic cleavage site, similar to that found in mammalian TIP39 ligands. Phylogenetic analyses suggested that TIP39 forms the basal group from which PTH and PTHrP have been derived. Functionally, subtle differences in potency could be discerned between hTIP39 and zTIP39. The human PTH2R and zPTH2R were stimulated slightly better by both hTIP39 and zTIP39, whereas zTIP39 had a higher potency at a previously isolated zPTH2R splice variant. Whole-mount in situ hybridization of zebrafish revealed strong zTIP39 expression in the region of the hypothalamus and in the heart of 24- and 48-h-old embryos. Similarly, zPTH2R expression was highly expressed throughout the brain of 48- and 72-h-old embryos. Because the mammalian PTH2R was also most abundantly expressed in these tissues, the TIP39-PTH2R system may serve conserved physiological roles in mammals and fishes. PMID- 15297443 TI - Deletion of the pleckstrin and phosphotyrosine binding domains of insulin receptor substrate-2 does not impair its ability to regulate cell proliferation in myeloid cells. AB - 32D IGF-I receptor (IR) cells are IL-3-dependent myeloid cells that can be induced to differentiate into granulocytes by IGF-I. Like the parental 32D cells, 32D IGF-IR cells do not express the insulin receptor substrate (IRS)-1 or IRS-2. We investigated the effect of ectopic expression of IRS-2 in 32D IGF-IR cells. Expression in these cells of a wild-type IRS-2 inhibits IGF-I-induced differentiation, and the cells grow indefinitely in the absence of IL-3. We also investigated the effect of a mutant IRS-2 lacking both the pleckstrin (PH) and the phosphotyrosine-binding (PTB) domains, which are known to bind to the IR. The partial differentialPHPTB IRS-2 is fully as capable as the wild-type IRS-2 (and wild-type IRS-1) to stimulate the growth and inhibit the differentiation of 32D IGF-IR cells. In contrast, an IRS-1 protein lacking the same PH and PTB domains is completely inactive in blocking differentiation and stimulating IL-3 independent growth of 32D IGF-IR cells. The partial differentialPHPTB IRS-2 protein is dependent for its effect on an activated IGF-IR, is cytoplasmic, binds to the beta-subunit of the IGF-IR, and requires for its action the presence of phosphatidylinositol 3-kinase binding sequences. These experiments show that the PH and PTB domains of IRS-2 (but not IRS-1) are dispensable for the IGF-I/IRS-2 mediated growth of 32D myeloid cells. Our results also indicate that IRS-2 (either wild type or partial differentialPHPTB) is capable of inhibiting the differentiation of 32D cells. PMID- 15297444 TI - Adrenergic regulation and diurnal rhythm of p38 mitogen-activated protein kinase phosphorylation in the rat pineal gland. AB - In this study, we investigated adrenergic and photoneural regulation of p38MAPK phosphorylation in the rat pineal gland. Norepinephrine (NE), the endogenous neurotransmitter, dose-dependently increased the levels of phosphorylated MAPK kinase 3/6 (MKK3/6) and p38MAPK in rat pinealocytes. Time-course studies showed a gradual increase in MKK3/6 and p38MAPK phosphorylation that peaked between 1 and 2 h and persisted for 4 h post NE stimulation. In cells treated with NE for 2 and 4 h, the inclusion of prazosin or propranolol reduced NE-induced MKK3/6 and p38MAPK phosphorylation, indicating involvement of both alpha- and beta adrenergic receptors for the sustained response. Whereas treatment with dibutyryl cAMP or ionomycin mimicked the NE-induced MKK3/6 and p38MAPK phosphorylation, neither dibutyryl cGMP nor 4beta-phorbol 12-myristate 13-acetate had an effect. The NE-induced increase in MKK3/6 and p38MAPK phosphorylation was blocked by KT5720 (a protein kinase A inhibitor) and KN93 (a Ca(2+)/calmodulin-dependent kinase inhibitor), but not by KT5823 (a protein kinase G inhibitor) or calphostin C (a protein kinase C inhibitor). In animals housed under a lighting regimen with 12 h of light, MKK3/6 and p38MAPK phosphorylation increased in the rat pineal gland at zeitgeber time 18. The nocturnal increase in p38MAPK phosphorylation was blocked by exposing the animal to constant light and reduced by treatment with propranolol, a beta-adrenergic blocker. Together, our results indicate that activation of p38MAPK is under photoneural control in the rat pineal gland and that protein kinase A and intracellular Ca(2+) signaling pathways are involved in NE regulation of p38MAPK. PMID- 15297445 TI - Dissecting linear and conformational epitopes on the native thyrotropin receptor. AB - The TSH receptor (TSHR) is the primary antigen in Graves' disease. In this condition, autoantibodies to the TSHR that have intrinsic thyroid-stimulating activity develop. We studied the epitopes on the native TSHR using polyclonal antisera and monoclonal antibodies (mAbs) derived from an Armenian hamster model of Graves' disease. Of 14 hamster mAbs analyzed, five were shown to bind to conformational epitopes including one mAb with potent thyroid-stimulating activity. Overlapping conformational epitopes were determined by cell-binding competition assays using fluorescently labeled mAbs. We identified two distinct conformational epitopes: epitope A for both stimulating and blocking mAbs and epitope B for only blocking mAbs. Examination of an additional three mouse derived stimulating TSHR-mAbs also showed exclusive binding to epitope A. The remaining nine hamster-derived mAbs were neutral or low-affinity blocking antibodies that recognized linear epitopes within the TSHR cleaved region (residues 316-366) (epitope C). Serum from the immunized hamsters also recognized conformational epitopes A and B but, in addition, also contained high levels of TSHR-Abs interacting within the linear epitope C region. In summary, these studies indicated that the natively conformed TSHR had a restricted set of epitopes recognized by TSHR-mAbs and that the binding site for stimulating TSHR Abs was highly conserved. However, high-affinity TSHR-blocking antibodies recognized two conformational epitopes, one of which was indistinguishable from the thyroid-stimulating epitope. Hence, TSHR-stimulating and blocking antibodies cannot be distinguished purely on the basis of their conformational epitope recognition. PMID- 15297446 TI - Differential regulation of the cell cycle by alpha1-adrenergic receptor subtypes. AB - Alpha(1)-Adrenergic receptors have been implicated in growth-promoting pathways. A microarray study of individual alpha(1)-adrenergic receptor subtypes (alpha(1A), alpha(1B), and alpha(1D)) expressed in Rat-1 fibroblasts revealed that epinephrine altered the transcription of several cell cycle regulatory genes in a direction consistent with the alpha(1A)- and alpha(1D)-adrenergic receptors mediating G(1)-S cell cycle arrest and the alpha(1B-)mediating cell-cycle progression. A time course indicated that in alpha(1A) cells, epinephrine stimulated a G(1)-S arrest, which began after 8 h of stimulation and maximized at 16 h, at which point was completely blocked with cycloheximide. The alpha(1B) adrenergic receptor profile also showed unchecked cell cycle progression, even under low serum conditions and induced foci formation. The G(1)-S arrest induced by alpha(1A)- and alpha(1D)-adrenergic receptors was associated with decreased cyclin-dependent kinase-6 and cyclin E-associated kinase activities and increased expression of the cyclin-dependent kinase inhibitor p27(Kip1), all of which were blocked by prazosin. There were no differences in kinase activities and/or expression of p27(Kip1) in epinephrine alpha(1B)-AR fibroblasts, although the microarray did indicate differences in p27(Kip1) RNA levels. Cell counts proved the antimitotic effect of epinephrine in alpha(1A) and alpha(1D) cells and indicated that alpha(1B)-adrenergic receptor subtype expression was sufficient to cause proliferation of Rat-1 fibroblasts independent of agonist stimulation. Analysis in transfected PC12 cells also confirmed the alpha(1A)- and alpha(1B) adrenergic receptor effect. The alpha(1B)-subtype native to DDT1-MF2 cells, a smooth muscle cell line, caused progression of the cell cycle. These results indicate that the alpha(1A)- and alpha(1D)-adrenergic receptors mediate G(1)-S cell-cycle arrest, whereas alpha(1B)-adrenergic receptor expression causes a cell cycle progression and may induce transformation in sensitive cell lines. PMID- 15297447 TI - Dehydration-induced synaptic plasticity in magnocellular neurons of the hypothalamic supraoptic nucleus. AB - Norepinephrine plays a critical role in the regulation of hypothalamic neuroendocrine function, in large part through modulation of synaptic glutamate and gamma-aminobutyric acid (GABA) release. Hypothalamic magnocellular neuroendocrine cells undergo dramatic changes in synaptic organization under conditions of increased hormone release, including increased numbers of glutamatergic, GABAergic and noradrenergic synapses. We studied the functional plasticity of magnocellular neurons of the rat supraoptic nucleus induced by chronic dehydration using whole-cell recordings in hypothalamic slices. Dehydrated rats showed increases in glutamate and GABA release onto magnocellular neurons, as evidenced by an increase in the frequency of spontaneous excitatory (29%) and inhibitory (33%) postsynaptic currents. The change in glutamate release was likely due to increased numbers of release sites because paired-pulse facilitation analysis did not reveal a change in the probability of transmitter release. In untreated rats, norepinephrine facilitates glutamate release and attenuates GABA release onto magnocellular neurons. Dehydration resulted in a marked enhancement of norepinephrine's actions, doubling both the norepinephrine induced increase in glutamate release and decrease in GABA release. The norepinephrine dose-response curve was shifted to the left with dehydration, revealing an increase in norepinephrine sensitivity. Thus, dehydration leads to an increase in glutamate and GABA release onto supraoptic magnocellular neurons as well as a marked enhancement of the facilitatory effect of norepinephrine on glutamate release and inhibitory effect on GABA release. This synaptic plasticity would be expected to increase the excitability of the magnocellular neurons and support the enhanced bursting capacity and facilitated hormone secretion observed in vivo with chronic dehydration. PMID- 15297448 TI - Small-molecule insulin mimetic reduces hyperglycemia and obesity in a nongenetic mouse model of type 2 diabetes. AB - Adiposity positively correlates with insulin resistance and is a major risk factor of type 2 diabetes. Administration of exogenous insulin, which acts as an anabolic factor, facilitates adipogenesis. Recently nonpeptidal insulin receptor (IR) activators have been discovered. Here we evaluate the effects of the orally bioavailable small-molecule IR activator (Compound-2) on metabolic abnormalities associated with type 2 diabetes using a nongenetic mouse model in comparison with the effects of a novel non-thiazolidinedione (nTZD) peroxisome proliferator activated receptor-gamma agonist. Both Compound-2 and nTZD alleviated fasting and postprandial hyperglycemia; accelerated glucose clearance rate; and normalized plasma levels of nonesterified fatty acids, triglycerides, and leptin. Unlike nTZD, which increased body weight gain, and total fat mass, which is a common feature for PPARgamma agonists, Compound-2 prevented body weight gain and hypertrophy of brown, and white adipose tissue depots and the development of hepatic steatosis in the mouse model of type 2 diabetes. The effect of the two compounds on proximal steps in insulin signal transduction pathway was analyzed in tissues. Compound-2 enhanced insulin-stimulated phosphorylation of IR tyrosine and/or Akt in the liver, skeletal muscle, and white adipose tissue, whereas nTZD potentiated the phosphorylation of IR and Akt in the adipose tissue only. In conclusion, small-molecule IR activators have unique features as insulin sensitizers and hold potential utility in the treatment of type 2 diabetes and obesity. PMID- 15297449 TI - Goldfish calmodulin: molecular cloning, tissue distribution, and regulation of transcript expression in goldfish pituitary cells. AB - Calmodulin (CaM) is a Ca(2+)-binding protein essential for biological functions mediated through Ca(2+)-dependent mechanisms. In the goldfish, CaM is involved in the signaling events mediating pituitary hormone secretion induced by hypothalamic factors. However, the structural identity of goldfish CaM has not been established, and the neuroendocrine mechanisms regulating CaM gene expression at the pituitary level are still unknown. Here we cloned the goldfish CaM and tested the hypothesis that pituitary expression of CaM transcripts can be the target of modulation by hypothalamic factors. Three goldfish CaM cDNAs, namely CaM-a, CaM-bS, and CaM-bL, were isolated by library screening. These cDNAs carry a 450-bp open reading frame encoding the same 149-amino acid CaM protein, the amino acid sequence of which is identical with that of mammals, birds, and amphibians and is highly homologous (>/=90%) to that in invertebrates. In goldfish pituitary cells, activation of cAMP- or PKC-dependent pathways increased CaM mRNA levels, whereas the opposite was true for induction of Ca(2+) entry. Basal levels of CaM mRNA was accentuated by GnRH and pituitary adenylate cyclase activating polypeptide but suppressed by dopaminergic stimulation. Pharmacological studies using D1 and D2 analogs revealed that dopaminergic inhibition of CaM mRNA expression was mediated through pituitary D2 receptors. At the pituitary level, D2 activation was also effective in blocking GnRH- and pituitary adenylate cyclase-activating polypeptide-stimulated CaM mRNA expression. As a whole, the present study has confirmed that the molecular structure of CaM is highly conserved, and its mRNA expression at the pituitary level can be regulated by interactions among hypothalamic factors. PMID- 15297450 TI - ATP increases the affinity between MutS ATPase domains. Implications for ATP hydrolysis and conformational changes. AB - MutS is the key protein of the Escherichia coli DNA mismatch repair system. It recognizes mispaired and unpaired bases and has intrinsic ATPase activity. ATP binding after mismatch recognition by MutS serves as a switch that enables MutL binding and the subsequent initiation of mismatch repair. However, the mechanism of this switch is poorly understood. We have investigated the effects of ATP binding on the MutS structure. Crystallographic studies of ATP-soaked crystals of MutS show a trapped intermediate, with ATP in the nucleotide-binding site. Local rearrangements of several residues around the nucleotide-binding site suggest a movement of the two ATPase domains of the MutS dimer toward each other. Analytical ultracentrifugation experiments confirm such a rearrangement, showing increased affinity between the ATPase domains upon ATP binding and decreased affinity in the presence of ADP. Mutations of specific residues in the nucleotide binding domain reduce the dimer affinity of the ATPase domains. In addition, ATP induced release of DNA is strongly reduced in these mutants, suggesting that the two activities are coupled. Hence, it seems plausible that modulation of the affinity between ATPase domains is the driving force for conformational changes in the MutS dimer. These changes are driven by distinct amino acids in the nucleotide-binding site and form the basis for long-range interactions between the ATPase domains and DNA-binding domains and subsequent binding of MutL and initiation of mismatch repair. PMID- 15297451 TI - A two-component flavin-dependent monooxygenase involved in actinorhodin biosynthesis in Streptomyces coelicolor. AB - The two-component flavin-dependent monooxygenases belong to an emerging class of enzymes involved in oxidation reactions in a number of metabolic and biosynthetic pathways in microorganisms. One component is a NAD(P)H:flavin oxidoreductase, which provides a reduced flavin to the second component, the proper monooxygenase. There, the reduced flavin activates molecular oxygen for substrate oxidation. Here, we study the flavin reductase ActVB and ActVA-ORF5 gene product, both reported to be involved in the last step of biosynthesis of the natural antibiotic actinorhodin in Streptomyces coelicolor. For the first time we show that ActVA-ORF5 is a FMN-dependent monooxygenase that together with the help of the flavin reductase ActVB catalyzes the oxidation reaction. The mechanism of the transfer of reduced FMN between ActVB and ActVA-ORF5 has been investigated. Dissociation constant values for oxidized and reduced flavin (FMNox and FMNred) with regard to ActVB and ActVA-ORF5 have been determined. The data clearly demonstrate a thermodynamic transfer of FMNred from ActVB to ActVA-ORF5 without involving a particular interaction between the two protein components. In full agreement with these data, we propose a reaction mechanism in which FMNox binds to ActVB, where it is reduced, and the resulting FMNred moves to ActVA-ORF5, where it reacts with O2 to generate a flavinperoxide intermediate. A direct spectroscopic evidence for the formation of such species within ActVA-ORF5 is reported. PMID- 15297452 TI - Transcriptional regulation of APOBEC3G, a cytidine deaminase that hypermutates human immunodeficiency virus. AB - Apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like 3G (APOBEC3G) is an antiretroviral deoxycytidine deaminase that lethally hypermutates human immunodeficiency virus type 1 (HIV-1) but is itself neutralized by the HIV-1 encoded viral infectivity factor. Accordingly, APOBEC3G occurs specifically in human T lymphocytic cell lines that contain this antiviral defense, including H9. Since the substrate specificities of related cytidine deaminases are strongly influenced by their intracellular quantities, we analyzed the factors that control APOBEC3G expression. The levels of APOBEC3G mRNA and protein were unaffected by treatment of proliferating H9 cells with interferons or tumor necrosis factor-alpha but were enhanced up to 20-fold by phorbol myristate acetate. This induction was mediated at the transcriptional level by a pathway that required activation of the protein kinase Calpha/betaI isozyme (PKC), mitogen-activated protein kinase kinase (MEK) 1 and 2, and extracellular signal regulated kinase (ERK). Correspondingly, induction of APOBEC3G was blocked by multiple inhibitors that act at diverse steps of this pathway. The PKCalpha/betaI/MEK/ERK pathway also controlled basal levels of APOBEC3G mRNA and protein, which consequently declined when cells were treated with these inhibitors or arrested in the G(0) state of the cell cycle by serum starvation. We conclude that expression of the antiviral APOBEC3G editing enzyme is dynamically controlled by the PKCalpha/betaI/MEK/ERK protein kinase cascade in human T lymphocytes. PMID- 15297453 TI - Human heme oxygenase oxidation of 5- and 15-phenylhemes. AB - Human heme oxygenase-1 (hHO-1) catalyzes the O2-dependent oxidation of heme to biliverdin, CO, and free iron. Previous work indicated that electrophilic addition of the terminal oxygen of the ferric hydroperoxo complex to the alpha meso-carbon gives 5-hydroxyheme. Earlier efforts to block this reaction with a 5 methyl substituent failed, as the reaction still gave biliverdin IXalpha. Surprisingly, a 15-methyl substituent caused exclusive cleavage at the gamma-meso rather than at the normal, unsubstituted alpha-meso-carbon. No CO was formed in these reactions, but the fragment cleaved from the porphyrin eluded identification. We report here that hHO-1 cleaves 5-phenylheme to biliverdin IXalpha and oxidizes 15-phenylheme at the alpha-meso position to give 10 phenylbiliverdin IXalpha. The fragment extruded in the oxidation of 5-phenylheme is benzoic acid, one oxygen of which comes from O2 and the other from water. The 2.29- and 2.11-A crystal structures of the hHO-1 complexes with 1- and 15 phenylheme, respectively, show clear electron density for both the 5- and 15 phenyl rings in both molecules of the asymmetric unit. The overall structure of 15-phenylheme-hHO-1 is similar to that of heme-hHO-1 except for small changes in distal residues 141-150 and in the proximal Lys18 and Lys22. In the 5-phenylheme hHO-1 structure, the phenyl-substituted heme occupies the same position as heme in the heme-HO-1 complex but the 5-phenyl substituent disrupts the rigid hydrophobic wall of residues Met34, Phe214, and residues 26-42 near the alpha meso carbon. The results provide independent support for an electrophilic oxidation mechanism and support a role for stereochemical control of the reaction regiospecificity. PMID- 15297454 TI - Ubiquitylation of nascent globin chains in a cell-free system. AB - The ubiquitin/proteasome pathway for degradation of completed and nascent globin chains was evaluated using a cell-free in vitro coupled transcription/translation assay. No decrease in radiolabeled globin chains was observed when ubiquitin, energy regenerating source (or ATP), and E1 and E2 enzymes were added 30 min after the start of translation when globin chain synthesis had plateaued. In contrast, the addition of these components prior to the start of translation resulted in no radiolabeled globin chains after 30 min. The loss of radiolabeled globin chains was dependent on ATP concentration; the higher the concentration, the less the radiolabeled globin chains formed. Prior to the initiation of transcription/translation, cell extract was preincubated with the proteasomal inhibitor MG132 in the absence of globin chain expression vector after which ubiquitin-protein isopeptidase inhibitor, Ubal, and expression vector were added in the presence of 1.5 mm ATP. Thereafter, radiolabeled monoubiquitylated and multiubiquitylated globin chains with few unmodified globin chains were formed. Our results suggest that polyubiquitylated globin chains are localized to the polysomal fractions. These results suggest that nascent globin chains are potential targets for ubiquitylation and deubiquitylation during or soon after translation and that ATP levels play a role in the balance between polypeptide synthesis and degradation. PMID- 15297455 TI - A TRPC1/TRPC3-mediated increase in store-operated calcium entry is required for differentiation of H19-7 hippocampal neuronal cells. AB - Store-operated calcium entry (SOCE) and TRPC protein expression were investigated in the rat-derived hippocampal H19-7 cell line. Thapsigargin-stimulated Ba2+ entry and the expression of TRPC1, TRPC3, TRPC4, TRPC5, TRPC6, and TRPC7 mRNA and protein were observed in proliferating H19-7 cells. When cells were placed under differentiating conditions, a change in TRPC homolog expression profile occurred. The expression of TRPC1 and TRPC3 mRNA and protein dramatically increased, while the expression of TRPC4 and TRPC7 mRNA and protein dramatically decreased; in parallel a 3.4-fold increase in the level of thapsigargin-stimulated Ba2+ entry was observed and found to be inhibited by 2-aminoethoxydiphenylborane. The selective suppression of TRPC protein levels by small interfering RNA (siRNA) approaches indicated that TRPC1 and TRPC3 are involved in mediating SOCE in proliferating H19-7 cells. Although TRPC4 and TRPC7 are expressed at much higher levels than TRPC1 and TRPC3 in proliferating cells, they do not appear to mediate SOCE. The co-expression of siRNA specific for TRPC1 and TRPC3 in proliferating cells inhibited approximately the same amount of SOCE as observed with expression of either siRNA alone, suggesting that TRPC1 and TRPC3 work in tandem to mediate SOCE. Under differentiating conditions, co-expression of siRNA for TRPC1 and TRPC3 blocked the normal 3.4-fold increase in SOCE and in turn blocked the differentiation of H19-7 cells. This study suggests that placing H19-7 cells under differentiating conditions significantly alters TRPC gene expression and increases the level of SOCE and that this increase in SOCE is necessary for cell differentiation. PMID- 15297456 TI - Suppression of uracil-DNA glycosylase induces neuronal apoptosis. AB - A chronic imbalance in DNA precursors, caused by one-carbon metabolism impairment, can result in a deficiency of DNA repair and increased DNA damage. Although indirect evidence suggests that DNA damage plays a role in neuronal apoptosis and in the pathogenesis of neurodegenerative disorders, the underlying mechanisms are poorly understood. In particular, very little is known about the role of base excision repair of misincorporated uracil in neuronal survival. To test the hypothesis that repair of DNA damage associated with uracil misincorporation is critical for neuronal survival, we employed an antisense (AS) oligonucleotide directed against uracil-DNA glycosylase encoded by the UNG gene to deplete UNG in cultured rat hippocampal neurons. AS, but not a scrambled control oligonucleotide, induced apoptosis, which was associated with DNA damage analyzed by comet assay and up-regulation of p53. UNG mRNA and protein levels were decreased within 30 min and were undetectable within 6-9 h of exposure to the UNG AS oligonucleotide. Whereas UNG expression is significantly higher in proliferating as compared with nonproliferating cells, such as neurons, the levels of UNG mRNA were increased in brains of cystathionine beta-synthase knockout mice, a model for hyperhomocysteinemia, suggesting that one-carbon metabolism impairment and uracil misincorporation can induce the up-regulation of UNG expression. PMID- 15297457 TI - On the slowing of S phase in response to DNA damage in fission yeast. AB - Eukaryotic cells slow their progression through S phase upon DNA damage. The mechanism that leads to this slowing is called the intra-S-phase checkpoint. Previous studies demonstrated that in the fission yeast Schizosaccharomyces pombe this checkpoint is mediated by a pathway that includes Rad3 (similar to human ATR and ATM) and Cds1 (similar to human Chk1 and Chk2). Here we present evidence that a major downstream target of this pathway is the cyclin-dependent kinase, Cdc2. We also present evidence suggesting that the intra-S-phase checkpoint makes a relatively minor contribution to the survival of cells with damaged DNA. PMID- 15297458 TI - Suppression of the human parathyroid hormone promoter by vitamin D involves displacement of NF-Y binding to the vitamin D response element. AB - An earlier report in the literature indicated the vitamin D response element (VDRE) in the human parathyroid hormone (hPTH) promoter could be specifically bound by an unidentified transcription factor in addition to the vitamin D receptor (VDR) complex. We confirmed that OK and HeLa cell nuclear extracts formed a specific complex with the hPTH VDRE that was insensitive to competition with other VDRE sequences. However, this factor could be competed for by a consensus NF-Y DNA-binding site, and an anti-NF-Y antibody was able to supershift the bound band. Mutational analysis indicated that the NF-Y-binding site partially overlapped the 3' portion of the VDRE. Transfection studies using an hPTH promoter construct in Drosophila SL2 cells demonstrated strong synergistic transactivation by NF-Y interactions with both the VDRE site and a previously described distal NF-Y-binding site. Finally, mobility shift studies indicated that the VDR heterodimer competed with NF-Y for binding to the VDRE sequence, and NF-Y-stimulated activity of the hPTH promoter could be suppressed in a hormone dependent manner when the VDR heterodimer complex was coexpressed in SL2 cells. In summary, these findings establish the presence of a proximal NF-Y-binding site in the hPTH promoter and highlight the potential for synergism between distal and proximal NF-Y DNA elements to strongly enhance transcription. Furthermore, findings suggest that the repressive effects of vitamin D on hPTH gene transcription may involve displacement of NF-Y binding to the proximal site by the VDR heterodimer, which subsequently attenuates synergistic transactivation. PMID- 15297459 TI - Neuronal roles of the integrin-associated protein (IAP/CD47) in developing cortical neurons. AB - Little is known about the role of the integrin-associated protein (IAP, or CD47) in neuronal development and its function in the central nervous system. We investigated neuronal responses in IAP-overexpressing cortical neurons using a virus-gene transfer system. We found that dendritic outgrowth was significantly enhanced in IAP (form 4)-transfected neurons. Furthermore, synaptic proteins including synaptotagmin, syntaxin, synapsin I, and SNAP25 (25-kDa synaptosomal associated protein) were up-regulated. In accordance with this finding, the release of the excitatory transmitter glutamate and the frequencies of Ca2+ oscillations (glutamate-mediated synaptic transmission) were increased. Interestingly, the overexpression of IAP activated mitogen-activated protein kinase (MAPK), and this activation was required for the IAP-dependent biological effects. After down-regulation of the endogenous IAP by small interfering RNA, MAPK activity, synaptic protein levels, and glutamate release decreased. These observations suggest that the IAP plays important roles in dendritic outgrowth and synaptic transmission in developing cortical neurons through the activation of MAPK. PMID- 15297460 TI - Increased site 1 affinity improves biopotency of porcine growth hormone. Evidence against diffusion dependent receptor dimerization. AB - Based on phage display optimization studies with human growth hormone (GH), it is thought that the biopotency of GH cannot be increased. This is proposed to be a result of the affinity of the first receptor for hormone far exceeding that which is required to trap the hormone long enough to allow diffusion of the second receptor to form the ternary complex, which initiates signaling. We report here that despite similar site 1 kinetics to the hGH/hGH receptor interaction, the potency of porcine GH for its receptor can be increased up to 5-fold by substituting hGH residues involved in site 1 binding into pGH. Based on extensive mutations and BIAcore studies, we show that the higher potency and site 1 affinity of hGH for the pGHR is primarily a result of a decreased off-rate associated with residues in the extended loop between helices 1 and 2 that interact with the two key tryptophans Trp104 and Trp169 in the receptor binding hot spot. Our mutagenic analysis has also identified a second determinant (Lys165), which in addition to His169, restricts the ability of non-primate hormones to activate hGH receptor. The increased biopotency of GH that we observe can be explained by a model for GH receptor activation where subunit alignment is critical for effective signaling. PMID- 15297461 TI - Local control of AMPA receptor trafficking at the postsynaptic terminal by a small GTPase of the Rab family. AB - The delivery of neurotransmitter receptors into the synaptic membrane is essential for synaptic function and plasticity. However, the molecular mechanisms of these specialized trafficking events and their integration with the intracellular membrane transport machinery are virtually unknown. Here, we have investigated the role of the Rab family of membrane sorting proteins in the late stages of receptor trafficking into the postsynaptic membrane. We have identified Rab8, a vesicular transport protein associated with trans-Golgi network membranes, as a critical component of the cellular machinery that delivers AMPA type glutamatergic receptors (AMPARs) into synapses. Using electron microscopic techniques, we have found that Rab8 is localized in close proximity to the synaptic membrane, including the postsynaptic density. Electrophysiological studies indicated that Rab8 is necessary for the synaptic delivery of AMPARs during plasticity (long-term potentiation) and during constitutive receptor cycling. In addition, Rab8 is required for AMPAR delivery into the spine surface, but not for receptor transport from the dendritic shaft into the spine compartment or for delivery into the dendritic surface. Therefore, Rab8 specifically drives the local delivery of AMPARs into synapses. These results demonstrate a new role for the cellular secretory machinery in the control of synaptic function and plasticity directly at the postsynaptic membrane. PMID- 15297462 TI - Regulation of casein kinase-2 (CK2) activity by inositol phosphates. AB - Casein kinase 2 (CK2) was one of the first protein kinases to be discovered and has been suggested to be responsible for as much as one-fifth of the eukaryotic phosphoproteome. Despite being responsible for the phosphorylation of a vast array of proteins central to numerous dynamic cellular processes, the activity of CK2 appears to be unregulated. In the current study, we identified a protein kinase activity in rat liver supernatant that is up-regulated by inositol 1,3,4,5 tetrakisphosphate (IP4) and inositol hexakisphosphate (IP6). The substrate for the inositol phosphate-regulated protein kinase was identified as a phosphatidylcholine transfer protein-like protein. Using the phosphorylation of this substrate in an assay, we purified the inositol phosphate-regulated protein kinase and determined it to be CK2. Bacterially expressed recombinant CK2, however, showed very high basal activity and was only modestly activated by IP6 and not regulated by IP. We found that an endogenous component present in rat liver supernatant was able to inhibit both recombinant and liver-purified CK2 basal activity. Under these conditions, recombinant CK2 catalytic activity could be increased substantially by IP4, inositol 1,3,4,5,6-pentakisphosphate (IP5), and IP6. We concluded that, contrary to the previously held view, CK2 can exist in a state of low constitutive activity allowing for its regulation by inositol phosphates. The ability of the higher inositol phosphates to directly stimulate CK2 catalytic activity provides the first evidence that these signaling molecules can operate via a direct control of protein phosphorylation. PMID- 15297463 TI - A neutralizing anti-Nogo66 receptor monoclonal antibody reverses inhibition of neurite outgrowth by central nervous system myelin. AB - The Nogo66 receptor (NgR1) is a neuronal, leucine-rich repeat (LRR) protein that binds three central nervous system (CNS) myelin proteins, Nogo, myelin-associated glycoprotein, and oligodendrocyte myelin glycoprotein, and mediates their inhibitory effects on neurite growth. Although the LRR domains on NgR1 are necessary for binding to the myelin proteins, the exact epitope(s) involved in ligand binding is unclear. Here we report the generation and detailed characterization of an anti-NgR1 monoclonal antibody, 7E11. The 7E11 monoclonal antibody blocks Nogo, myelin-associated glycoprotein, and oligodendrocyte myelin glycoprotein binding to NgR1 with IC50 values of 120, 14, and 4.5 nm, respectively, and effectively promotes neurite outgrowth of P3 rat dorsal root ganglia neurons cultured on a CNS myelin substrate. Further, we have defined the molecular epitope of 7E11 to be DNAQLR located in the third LRR domain of rat NgR1. Our data demonstrate that anti-NgR1 antibodies recognizing this epitope, such as 7E11, can neutralize CNS myelin-dependent inhibition of neurite outgrowth. Thus, specific anti-NgR1 antibodies may represent a useful therapeutic approach for promoting CNS repair after injury. PMID- 15297464 TI - A juxtamembrane tyrosine in the colony stimulating factor-1 receptor regulates ligand-induced Src association, receptor kinase function, and down-regulation. AB - Recent literature implicates a regulatory function of the juxtamembrane domain (JMD) in receptor tyrosine kinases. Mutations in the JMD of c-Kit and Flt3 are associated with gastrointestinal stromal tumors and acute myeloid leukemias, respectively. Additionally, autophosphorylated Tyr559 in the JMD of the colony stimulating factor-1 (CSF-1) receptor (CSF-1R) binds to Src family kinases (SFKs). To investigate SFK function in CSF-1 signaling we established stable 32D myeloid cell lines expressing CSF-1Rs with mutated SFK binding sites (Tyr559 TFI). Whereas binding to I562S was not significantly perturbed, Y559F and Y559D exhibited markedly decreased CSF-1-dependent SFK association. All JMD mutants retained intrinsic kinase activity, but Y559F, and less so Y559D, showed dramatically reduced CSF-1-induced autophosphorylation. CSF-1-mediated wild-type (WT)-CSF-1R phosphorylation was not markedly affected by SFK inhibition, indicating that lack of SFK binding is not responsible for diminished Y559F phosphorylation. Unexpectedly, cells expressing Y559F were hyperproliferative in response to CSF-1. Hyperproliferation correlated with prolonged activation of Akt, ERK, and Stat5 in the Y559F mutant. Consistent with a defect in receptor negative regulation, c-Cbl tyrosine phosphorylation and CSF-1R/c-Cbl co association were almost undetectable in the Y559F mutant. Furthermore, Y559F underwent reduced multiubiquitination and delayed receptor internalization and degradation. In conclusion, we propose that Tyr559 is a switch residue that functions in kinase regulation, signal transduction and, indirectly, receptor down-regulation. These findings may have implications for the oncogenic conversion of c-Kit and Flt3 with JMD mutations. PMID- 15297465 TI - New class of blue animal pigments based on Frizzled and Kringle protein domains. AB - The nature of coloration in many marine animals remains poorly investigated. Here we studied the blue pigment of a scyfoid jellyfish Rhizostoma pulmo and determined it to be a soluble extracellular 30-kDa chromoprotein with a complex absorption spectrum peaking at 420, 588, and 624 nm. Furthermore, we cloned the corresponding cDNA and confirmed its identity by immunoblotting and mass spectrometry experiments. The chromoprotein, named rpulFKz1, consists of two domains, a Frizzled cysteine-rich domain and a Kringle domain, inserted into one another. Generally, Frizzleds are members of a basic Wnt signal transduction pathway investigated intensely with regard to development and cancerogenesis. Kringles are autonomous structural domains found throughout the blood clotting and fibrinolytic proteins. Neither Frizzled and Kringle domains association with any type of coloration nor Kringle intrusion into Frizzled sequence was ever observed. Thus, rpulFKz1 represents a new class of animal pigments, whose chromogenic group remains undetermined. The striking homology between a chromoprotein and members of the signal transduction pathway provides a novel node in the evolution track of growth factor-mediated morphogenesis compounds. PMID- 15297466 TI - Functional analysis of the alpha-defensin disulfide array in mouse cryptdin-4. AB - The alpha-defensin antimicrobial peptide family is defined by a unique tridisulfide array. To test whether this invariant structural feature determines alpha-defensin bactericidal activity, mouse cryptdin-4 (Crp4) tertiary structure was disrupted by pairs of site-directed Ala for Cys substitutions. In a series of Crp4 disulfide variants whose cysteine connectivities were confirmed using NMR spectroscopy and mass spectrometry, mutagenesis did not induce loss of function. To the contrary, the in vitro bactericidal activities of several Crp4 disulfide variants were equivalent to or greater than those of native Crp4. Mouse Paneth cell alpha-defensins require the proteolytic activation of precursors by matrix metalloproteinase-7 (MMP-7), prompting an analysis of the relative sensitivities of native and mutant Crp4 and pro-Crp4 molecules to degradation by MMP-7. Although native Crp4 and the alpha-defensin moiety of proCrp4 resisted proteolysis completely, all disulfide variants were degraded extensively by MMP 7. Crp4 bactericidal activity was eliminated by MMP-7 cleavage. Thus, rather than determining alpha-defensin bactericidal activity, the Crp4 disulfide arrangement confers essential protection from degradation by this critical activating proteinase. PMID- 15297467 TI - Live cell imaging of Gs and the beta2-adrenergic receptor demonstrates that both alphas and beta1gamma7 internalize upon stimulation and exhibit similar trafficking patterns that differ from that of the beta2-adrenergic receptor. AB - To visualize and investigate the regulation of the localization patterns of Gs and an associated receptor during cell signaling, we produced functional fluorescent fusion proteins and imaged them in HEK-293 cells. alphas-CFP, with cyan fluorescent protein (CFP) inserted into an internal loop of alphas, localized to the plasma membrane and exhibited similar receptor-mediated activity to that of alphas. Functional fluorescent beta1gamma7 dimers were produced by fusing an amino-terminal yellow fluorescent protein (YFP) fragment to beta1 (YFP N-beta1) and a carboxyl-terminal YFP fragment to gamma7 (YFP-C-gamma7). When expressed together, YFP-N-beta1 and YFP-C-gamma7 produced fluorescent signals in the plasma membrane that were not seen when the subunits were expressed separately. Isoproterenol stimulation of cells co-expressing alphas-CFP, YFP-N beta1/YFP-C-gamma7, and the beta2-adrenergic receptor (beta2AR) resulted in internalization of both fluorescent signals from the plasma membrane. Initially, alphas-CFP and YFP-N-beta1/YFP-C-gamma7 stained the cytoplasm diffusely, and subsequently they co-localized on vesicles that exhibited minimal overlap with beta2AR-labeled vesicles. Moreover, internalization of beta2AR-GFP, but not alphas-CFP or YFP-N-beta1/YFP-C-gamma7, was inhibited by a fluorescent dominant negative dynamin 1 mutant, Dyn1(K44A)-mRFP, indicating that the Gs subunits and beta2AR utilize different internalization mechanisms. Subsequent trafficking of the Gs subunits and beta2AR also differed in that vesicles labeled with the Gs subunits exhibited less overlap with RhoB-labeled endosomes and greater overlap with Rab11-labeled endosomes. Because Rab11 regulates traffic through recycling endosomes, co-localization of alphas and beta1gamma7 on these endosomes may indicate a means of recycling specific alphasbetagamma combinations to the plasma membrane. PMID- 15297468 TI - Corticotropin-releasing factor in brain: a role in activation, arousal, and affect regulation. AB - Organisms exposed to challenging stimuli that alter the status quo inside or outside of the body are required for survival purposes to generate appropriate coping responses that counteract departures from homeostasis. Identification of an executive control mechanism within the brain capable of coordinating the multitude of endocrine, physiological, and functional coping responses has high utility for understanding the response of the organism to stressor exposure under normal or pathological conditions. The corticotropin-releasing factor (CRF)/urocortin family of neuropeptides and receptors constitutes an affective regulatory system due to the integral role it plays in controlling neural substrates of arousal, emotionality, and aversive processes. In particular, available evidence from pharmacological intervention in multiple species and phenotyping of mutant mice shows that CRF/urocortin systems mediate motor and psychic activation, stimulus avoidance, and threat recognition responses to aversive stimulus exposure. It is suggested that affective regulation is exerted by CRF/urocortin systems within the brain based upon the sensitivity of local brain sites to CRF/urocortin ligand administration and the appearance of hypothalamo-pituitary-adrenocortical activation following stressor exposure. Moreover, these same stress neuropeptides may constitute a mechanism for learning to avoid noxious stimuli by facilitating the formation of so-called emotional memories. A conceptual framework is provided for extrapolation of animal model findings to humans and for viewing CRF/urocortin activation as a continuum measure linking normal and pathological states. PMID- 15297469 TI - Volatile anesthetics inhibit calcitonin gene-related peptide receptor-mediated responses in pithed rats and human neuroblastoma cells. AB - Calcitonin gene-related peptide (CGRP) has a potent vasodilatory effect that is mediated by specific receptors predominantly coupled to the activation of adenylate cyclase. The effects of volatile anesthetics on CGRP-induced vasodilation are unclear. We studied the effects of sevoflurane and isoflurane on CGRP-induced vasodilation in pithed rats and CGRP receptor-mediated responses in SK-N-MC cells, which are used as a model system to study the CGRP receptor and its downstream pathways. Male Wistar rats were pithed by inserting a stainless steel rod into the spinal cord. Mean arterial pressure (MAP) and cardiac output were maintained at approximately 100 mmHg and 50 ml.min(-1), respectively, with continuous infusion of noradrenaline. After 30 min of inhalation of anesthetics, CGRP (0.1, 0.3, 1.0, and 3.0 microg/kg) was administered intravenously. In SK-N MC cells, CGRP-, forskolin-, or cholera toxin-induced cAMP production was measured with or without anesthetics using radioimmunoassays. CGRP receptor binding density and affinity for the agonist were determined with (2 [125I]iodohystidyl10) CGRP with or without the anesthetics. Sevoflurane (4%) and isoflurane (2%) significantly inhibited the decrease in MAP and systemic vascular resistance. Furthermore, both anesthetics significantly inhibited CGRP- but not forskolin-induced cAMP production. Sevoflurane (4%) and isoflurane (4%) significantly inhibited cholera toxin-induced cAMP production. Both anesthetics did not affect ligand binding. These data suggest that sevoflurane and isoflurane inhibit CGRP-induced vasodilation at the site between the CGRP receptor and adenylate cyclase activation. The inhibitory site of volatile anesthetics on the CGRP receptor-mediated response involves Gs protein. PMID- 15297470 TI - Nitric oxide (NO)-releasing naproxen (HCT-3012 [(S)-6-methoxy-alpha-methyl-2 naphthaleneacetic Acid 4-(nitrooxy)butyl ester]) interactions with aspirin in gastric mucosa of arthritic rats reveal a role for aspirin-triggered lipoxin, prostaglandins, and NO in gastric protection. AB - Administration of selective and nonselective cyclooxygenase (COX)-2 inhibitors to rheumatoid arthritis patients taking low doses of acetylsalicylic acid (ASA) for cardiovascular prevention associates with increased risk of gastrointestinal bleeding. The present study was undertaken to investigate whether administration of HCT-3012 [(S)-6-methoxy-alpha-methyl-2-naphthaleneacetic acid 4 (nitrooxy)butyl ester], a nitric oxide (NO)-releasing derivative of naproxen, exacerbates gastric mucosal injury in arthritic rats administered low doses of ASA. Our results demonstrated that while treating arthritic rats with a dose of 30 mg/kg/day ASA causes detectable mucosal injury, but had no effect on arthritis score and interleukin-6 plasma levels, coadministration of naproxen (10 mg/kg/day) and celecoxib (30 mg/kg/day), in combination with ASA from day 7 to day 21, attenuates arthritis development (P <0.01 versus arthritis alone), but markedly enhanced gastric mucosal damage caused by ASA (P <0.01 versus ASA alone). In contrast, coadministration of HCT-3012 (15 mg/kg/day) significantly attenuated arthritis development, because HCT-3012 was equally or more effective than naproxen and celecoxib in attenuating local and systemic inflammation (P >0.001 versus arthritis) without exacerbating gastric mucosal injury caused by ASA. Arthritis development associates with gastric COX-2 induction, mRNA and protein, and enhanced gastric prostaglandin E2 (PGE2) synthesis (P <0.01 versus control rats). Although all treatments, including celecoxib, were effective in reducing gastric PGE2 synthesis, administering arthritic rats with ASA resulted in a significant increase in gastric content of aspirin-triggered lipoxin (ATL), a COX-2-derived lipid mediator that regulates proinflammatory responses at the neutrophils/endothelial interface. Administering arthritic rats with naproxen and celecoxib abrogates ATL formation induced by ASA although enhanced neutrophils accumulate into the gastric mucosa (P <0.01 versus ASA alone). In contrast, whereas HCT-3012 inhibited ATL formation, it did not increase neutrophil recruitment into the gastric microcirculation. Collectively, these data indicate that HCT-3012 derived from NO has the potential to compensate for inhibition of PGE2 and ATL and to protect the gastric mucosa by limiting the recruitment of neutrophils. These data suggest that HCT-3012 might be a safer alternative to nonsteroidal anti-inflammatory drugs and coxibs in rheumatic patients that take low doses of ASA. PMID- 15297471 TI - Physiological capacity of the reticuloendothelial system for the degradation of hemoglobin vesicles (artificial oxygen carriers) after massive intravenous doses by daily repeated infusions for 14 days. AB - A hemoglobin vesicle (HbV; diameter 252 +/- 53 nm) or liposome-encapsulated Hb is an artificial oxygen carrier developed for use as a transfusion alternative, and its oxygen-transporting capacity has been well characterized, although critical physiological compartments for the Hb degradation after a massive infusion of HbV and the safety outcome remain unknown. In this study, we aimed to examine the compartments for its degradation by daily repeated infusions (DRI) of HbV, focusing on its influence on the reticuloendothelial system (RES). Male Wistar rats intravenously received the HbV suspension at 10 ml/kg/day for 14 consecutive days. The cumulative infusion volume (140 ml/kg) was equal to 2.5 times the whole blood volume (56 ml/kg). The animals tolerated the DRI well and survived, and body weights continuously increased. One day after DRI, hepatosplenomegaly occurred significantly through the accumulation of large amounts of HbV. Plasma clinical chemistry was overall normal, except for a transient elevation of lipid components derived from HbV. These symptoms subsided 14 days after DRI. Hemosiderin deposition and up-regulation of heme oxygenase-1 coincided in the liver and spleen but were not evident in the parenchyma of these organs. Furthermore, the plasma iron and bilirubin levels remained unchanged, suggesting that the heme-degrading capacity of the RES did not surpass the ability to eliminate bilirubin. In conclusion, phospholipid vesicles for the encapsulation of Hb would be beneficial for heme detoxification through their preferential delivery to the RES, a physiological compartment for degradation of senescent RBCs, even at doses greater than putative clinical doses. PMID- 15297472 TI - Routine Epstein-Barr virus diagnostics from the laboratory perspective: still challenging after 35 years. PMID- 15297473 TI - Molecular serotyping of Klebsiella species isolates by restriction of the amplified capsular antigen gene cluster. AB - The objective of the present work was to develop a molecular method that would enable determination of the capsular serotypes of Klebsiella isolates without the use of antiserum. PCR amplification of the capsular antigen gene cluster (cps) was followed by digestion with the restriction enzyme HincII (cps PCR-restriction fragment length polymorphism [RFLP] analysis). The profiles (C patterns) obtained for 224 strains representing the 77 known K serotypes showed 3 to 13 fragments ranging in size from 0.2 to 4.4 kb. A total of 97 distinct C patterns were obtained; 100% of 61 pairs of samples tested twice showed reproducible C patterns. The C patterns were K-type specific; i.e., the C pattern(s) of any K serotype was distinct from the C patterns of all other K serotypes, with the only exceptions being serotypes K22 and K37, which are known to cross-react. For 12 of 17 K types for which at least two strains were included, C-pattern variations were found among strains with the same K serotype. Therefore, cps PCR-RFLP analysis has a higher discriminatory power than classical K serotyping. C-pattern identity was observed among strains with a given K type that were collected many years apart and from distinct sources, indicating C-pattern stability. Only 4.5% of the strains were nontypeable, because of unsuccessful PCR amplification (whereas 8 to 23% are nontypeable by classical K serotyping). Three of four noncapsulated strains analyzed showed recognizable C patterns. The K serotypes of 18 (82%) of 22 recent Klebsiella pneumoniae clinical isolates could be deduced from their C patterns. In conclusion, cps PCR-RFLP analysis allows determination of the K serotype, while it is easier to perform and more discriminatory than classical serotyping. PMID- 15297474 TI - Phenotypic and genotypic changes in Salmonella enterica subsp. enterica serotype typhimurium during passage in intestines of broiler chickens fed on diets that included ionophore anticoccidial supplements. AB - The effect of continuous in-feed administration of anticoccidial agents on antimicrobial sensitivity and the level of bacterial shedding in poultry experimentally infected with Salmonella enterica subsp. enterica serotype Typhimurium definitive type 104 (DT104) were investigated. On day 0, 1,200 1-day old Salmonella-free broiler chicks were placed into 50 pens, and the pens were randomly allocated to one of five treatments: nonsupplemented (negative control; T1), monensin at 120 mg/kg of diet (T2), salinomycin at 60 mg/kg of diet (T3), semduramicin at 20 mg/kg of diet (T4), or semduramicin at 25 mg/kg of diet (T5). Each bird was inoculated with a well-characterized strain of serotype Typhimurium DT104 on day 10. On day 49, the birds were euthanatized humanely. Bulk fecal samples were collected on days 13, 43, and 48 and were examined for organisms which had acquired resistance. The genetic basis of acquired resistance was determined from representative samples of isolates. Of 784 Salmonella-selective plates supplemented with antimicrobial agents, only 33 showed growth. These isolates came from all treatment regimens, including the nonsupplemented control. A number of phenotypic changes were observed; these included changes in motility, phage type, and agglutination properties. Supplementation of the diet with an anticoccidial drug does not appear to affect antimicrobial resistance or the level of excretion of salmonellae. Most of the changes observed do not seem to be related to the presence of a supplement in feed. Salmonellae appear to be capable of acquiring antimicrobial resistance and phenotypic changes independently of specific antimicrobial selection pressures. PMID- 15297475 TI - High genetic diversity of the attachment (G) protein of human metapneumovirus. AB - Complete genes encoding the predicted nucleoprotein (N), phosphoprotein (P), matrix protein (M), fusion protein (F), M2-1protein, M2-2protein, small hydrophobic protein (SH), and attachmentprotein (G) of seven newly isolated human metapneumoviruses (hMPVs) were analyzed and compared with previously published data for hMPV genes. Phylogenetic analysis of the nucleotide sequences indicated that there were two genetic groups, tentatively named groups 1 and 2, similar to the grouping of human respiratory syncytial virus. Although the predicted amino acid sequences of N, P, M, F, and M2 were highly conserved between the two groups (amino acid identities, 96% for N, 85% for P, 97% for M, 94% for F, 95% for M2-1, and 90% for M2-2), the amino acid identities of the SH and G proteins were low (SH, 58%; G, 33%). Furthermore, each group could be subdivided into two subgroups by phylogenetic analysis, tentatively named subgroups 1A and 1B and subgroups 2A and 2B. The predicted amino acid sequences of G within members of each subgroup were highly conserved (amino acid identities, 88% for group 1A, 93% for group 1B, and 96% for group 2B). The G of hMPV is thought to be the major antigenic determinant and to play an important role in the production of neutralizing antibodies. Clarification of the antigenic diversity of G is important for epidemiological analysis and for establishment of strategies to prevent hMPV infection. PMID- 15297476 TI - Analysis of clonal composition of Mycobacterium tuberculosis isolates in primary infections in children. AB - The assumption that Mycobacterium tuberculosis infections should be considered clonally homogeneous has been weakened in the last few years. Recent studies have shown (i) the isolation of different M. tuberculosis strains from sequential episodes, (ii) mixed infections by two M. tuberculosis strains, and (iii) genetic variations in M. tuberculosis subpopulations due to microevolution events. Nevertheless, it is unknown whether clonal heterogeneity could be found in the initial steps of M. tuberculosis infection, i.e., the primary infection. In the present study we analyzed the clonal composition of the M. tuberculosis isolates causing primary infections in children. Cultures were clonally homogeneous in most cases (11 of 12). In 1 of the 12 cases (8.3%), clonal heterogeneity among the M. tuberculosis isolates was found by spoligotyping and IS6110-based restriction fragment length polymorphism analysis. This case occurred in a 2-year old child in whom microevolution events were unlikely and who had no risk factors for overexposure to M. tuberculosis. Clonal heterogeneity should also be considered in primary M. tuberculosis infections, including circumstances in which it is usually unexpected. PMID- 15297477 TI - Sex pheromone response, clumping, and slime production in enterococcal strains isolated from occluded biliary stents. AB - Bile-resistant bacteria, particularly gram-positive Enterococcus faecalis and Enterococcus faecium, play an important role in biliary stent occlusion, because their sessile mode of growth protects them against host defenses and antimicrobial agents. Twelve E. faecalis and seven E. faecium strains isolated from occluded biliary stents have been investigated for slime production, presence of aggregation substance genes, and ability to adhere to Caco-2 cells. Ten isolates were strong producers of slime, and seven isolates produced clumps when exposed to pheromones of E. faecalis JH2-2 and/or OG1RF. The small E. faecium clumps differed from the large clumps of E. faecalis and were similar to those of E. faecium LS10(pBRG1) carrying a pheromone response plasmid. After induction with pheromones, the adhesion to Caco-2 cells of clumping-positive strains was found to increase from two- to fourfold. Amplicons of the expected size were detected in three clumping-positive and three clumping-negative E. faecalis isolates by using primers (agg) internal to a highly conserved region of the E. faecalis pheromone response plasmids pAD1, pPD1, and pCF10 and primers internal to prgB of the E. faecalis plasmid pCF10. The agg/prgB-positive E. faecalis strains were also positive in Southern hybridization experiments with a prgB-specific probe. No PCR products were obtained with the same primers from four clumping-positive isolates (one E. faecalis and three E. faecium strains), which were also Southern hybridization negative. Our results demonstrate that slime production and pheromone response are both present in isolated enterococci, suggesting that clinical strains with these features might have a selective advantage in colonizing biliary stents. PMID- 15297478 TI - Suicide PCR on skin biopsy specimens for diagnosis of rickettsioses. AB - As rickettsioses may be severe diseases and Rickettsia prowazekii is a potential agent of bioterrorism, highly efficient diagnostic techniques are required to detect rickettsiae in patients. We developed a nested PCR assay using single-use primers targeting single-use gene fragments present in the genomes of both Rickettsia conorii and R. prowazekii. We used this "suicide" PCR with DNA from 103 skin biopsy specimens from patients who definitely had a rickettiosis, 109 skin biopsy specimens from patients who possibly had a rickettsiosis, and 50 skin biopsy specimens from patients with nonrickettsial diseases. The suicide PCR detected "R. conorii conorii" in 38 biopsy specimens, R. africae in 28 biopsy specimens, R. slovaca in 12 biopsy specimens, "R. sibirica mongolotimonae" in 5 biopsy specimens, R. aeschlimannii in 2 biopsy specimens, and "R. conorii caspia" and "R. sibirica sibirica" in 1 biopsy specimen each. The technique had a specificity of 100% and a sensitivity of 68%. It was 2.2 times more sensitive than culture (P < 10(-2)) and 1.5 times more sensitive than regular PCR (P < 10( 2)). The efficacy of the suicide PCR was reduced by antibiotic therapy prior to biopsy (P < 10(-2)) and was increased when it was performed with eschar biopsy specimens (P = 0.03). We propose the use of the suicide PCR as a sensitive, specific, and versatile technique for improving the diagnosis of rickettsioses, especially when it is used on eschar biopsy specimens taken prior to antibiotic therapy. PMID- 15297479 TI - Evaluation of immunoglobulin M (IgM) and IgG rapid cassette test kits for diagnosis of melioidosis in an area of endemicity. AB - An enzyme-linked immunosorbent assay-based rapid cassette immunoglobulin G (IgG) and IgM immunochromogenic test kit was compared to the indirect hemagglutination test (IHA) for the diagnosis of acute melioidosis in northeastern Thailand. Admission sera from 70 culture-confirmed septicemic melioidosis patients and 30 patients with localized infections were tested. As a control group, 80 patients with other acute febrile illnesses (other bacterial infections, leptospirosis, or scrub typhus) and 119 healthy individuals were tested. The diagnostic sensitivity of the IgG and IgM tests and the IHA test were 79, 67, and 72%, respectively, with corresponding specificities of 90, 80, and 68%. This kit represents an improvement over IHA for the diagnosis of melioidosis an area of endemicity although, as with other serological tests, it has reduced diagnostic utility in a population with high background seropositivity. PMID- 15297480 TI - Real-time PCR assay using molecular beacon for quantitation of hepatitis B virus DNA. AB - Levels of hepatitis B virus (HBV) DNA in the blood serve as an important marker in monitoring the disease progression and treatment efficacy of chronic HBV infection. Several commercial assays are available for accurate measurement of HBV genomic DNA, but many of them are hampered by relatively low sensitivity and limited dynamic range. The aim of this study was to develop a sensitive and accurate assay for measuring HBV genomic DNA using real-time PCR with a molecular beacon (HBV beacon assay). The performance of this assay was validated by testing serial dilutions of the two EUROHEP HBV DNA standards (ad and ay subtypes) of known concentrations. The assay showed low intra-assay (<7%) and interassay (<5%) variations for both subtypes. Its dynamic range was found to be 10(1) to 10(7) copies per reaction (1.0 x 10(2) to 1.0 x 10(9) copies ml(-1)). The assay was further evaluated clinically using serum samples from 175 individuals with chronic hepatitis B. The HBV DNA level measured by this assay showed good correlation with that measured by the commercially available COBAS AMPLICOR HBV Monitor test (r = 0.901; P < 0.001). The higher sensitivity and broader dynamic range of this assay compared to the existing commercial assays will provide an ideal tool for monitoring disease progression and treatment efficacy in HBV infected patients, in particular for those with low levels of HBV viremia. PMID- 15297481 TI - High-resolution genotyping of Campylobacter upsaliensis strains originating from three continents. AB - Ninety-six Campylobacter upsaliensis strains that originated from Australia, Canada, and Europe (Germany) and that were isolated from humans, dogs, and cats were serotyped for their heat-stable surface antigens. All of them were genotyped by enterobacterial repetitive intergenic consensus sequence PCR (ERIC-PCR) profiling, and 83 strains were genotyped by macrorestriction analysis with the endonuclease XhoI. Eighty-four percent of the strains belonged to five different serotypes (serotypes OI, OII, OIII, OIV, and OVI), with the proportions of strains in each serotype being comparable among the groups of strains from all three continents. Two serotypes, OIII and OIV, were prevalent at rates of 35 to 40%. Serotypes OI, OII, and OVI were detected at rates of 1.5 to 15%. Between 10 and 17.7% of the strains did not react with the available antisera. Analysis of the ERIC-PCR profiles revealed two distinct genotypic clusters, which represented the German and the non-European strains, respectively. XhoI macrorestriction yielded two genotypic clusters; one of them contained 80.2% of the German strains and 34.6% of the non-European strains, and the second cluster consisted of 65.4% of the non-European strains and 19.8% of the German strains. Fourteen strains from all three continents were analyzed for their 16S rRNA gene sequences. Only two minor variations were detected in four of the strains. In conclusion, C. upsaliensis has undergone diverging processes of genome arrangement on different continents during evolution without segregating into different subspecies. PMID- 15297482 TI - Molecular typing and distribution of Staphylococcus aureus isolates in Eastern Canadian dairy herds. AB - Macrorestriction analysis of SmaI-digested chromosomal DNA, using pulsed field gel electrophoresis (PFGE) was performed to type and estimate genetic relationships among 288 Staphylococcus aureus isolates recovered from 58 Eastern Canadian dairy herds. In addition, a subset of the collection was phage typed and evaluated for sensitivity to 10 antimicrobial compounds. Of 288 isolates recovered, 29 distinct PFGE types were identified. Based on estimates of genetic relationships, the PFGE types were assigned to six lineage groups, designated A through F. Of all of the isolates, ca. 93% were assigned to lineage groups A, D, or F. In 58.6% of herds, only a single PFGE type was recovered, while the remainder had two to four types. Of the 212 isolates evaluated for antimicrobial resistance, 24.5% were resistant to one or more antimicrobials. Resistance to penicillin (9.9%) was most common, followed by resistance to sulfadimethoxine (7.5%). Isolates resistant to multiple antibiotics were rare. A total of 63% of isolates responded to phages from groups 1 and 3, and 32.8% could not be typed with any of the phage strains used. The other 4.1% belonged to a variety of phage types. Most of the PFGE lineage group A and F isolates corresponded to phage groups 3 and 1, respectively, and most group D isolates were not typeable. PFGE typing had better discriminatory power than phage typing in defining the relatedness of the S. aureus isolates. Distribution of PFGE types and phage types was independent across regions and within herds. PMID- 15297483 TI - Clinical and epidemiological aspects of infections caused by fusarium species: a collaborative study from Israel. AB - Fusarium infections are an important problem worldwide, commonly affecting immunocompromised individuals. We conducted a retrospective study in two Israeli tertiary medical centers of factors predisposing to infection by Fusarium spp. and their influence on the epidemiology and clinical outcome of this infection. Fusarium spp. were isolated from 89 patients with a median age of 57 years. Sixty eight patients were considered immunocompetent. Seven patients had disseminated disease, 34 had locally invasive disease, and 48 had superficial infection. Most infections were limited and occurred mainly in lower limbs. Factors associated with in-hospital mortality were chronic renal failure, hematological malignancy, immunosuppression, disseminated infection, and positive blood culture. Multivariate analysis showed that chronic renal failure, hematological malignancy, burns, and disseminated infection were independently associated with mortality. A surge in the frequency of infections was noticed during the summer for patients from rural areas, involving mainly the eyes and lungs. At one of the hospitals (in a mountainous area), there was an increase in the isolation rate over time. PMID- 15297484 TI - Bartonella koehlerae, a new cat-associated agent of culture-negative human endocarditis. AB - Bartonella koehlerae is reported for the first time to be a human pathogen that causes culture-negative endocarditis. It is also shown that this species, isolated twice before from domestic cats, can be recovered as well from a stray cat population in Israel. This work follows a recent report of the same case in which the causative agent was misidentified as B. henselae, based on serology and PCR-restriction fragment length polymorphism (RFLP) analysis (A. Schattner, O. Zimhony, B. Avidor, and M. Gilad, Lancet 361:1786, 2003). B. koehlerae was identified in the valvular tissue of an endocarditis patient by DNA sequencing of the PCR products of two Bartonella genes: the genes for citrate synthase (gltA) and riboflavin synthase (ribC). The commonly used PCR-RFLP analysis of the TaqI digested gltA PCR product did not distinguish between B. koehlerae and B. quintana or between B. elizabethae and B. clarridgeiae. PmlI digestion of the gltA amplification product failed to differentiate between B. quintana, B. clarridgeiae, and B. elizabethae. RFLP analysis of the heat shock protein (htrA) gene by TaqI digestion misidentified B. koehlerae as B. henselae. However, RFLP analysis of the ribC PCR product, digested with TaqI, was able to distinguish between the human endocarditis-associated Bartonella species tested, B. henselae, B. quintana, B. elizabethae, and B. koehlerae, as well as between the cat associated Bartonella species, B. henselae and B. clarridgeiae. Given the expanding number of Bartonella species emerging as human pathogens, it is suggested that PCR-RFLP analysis for the diagnosis of Bartonella infections target several genes and be coupled with DNA sequencing to avoid species identification. PMID- 15297485 TI - Selected pool of peptides from ESAT-6 and CFP-10 proteins for detection of Mycobacterium tuberculosis infection. AB - We have validated a new test for detecting Mycobacterium tuberculosis infection. A pool of synthetic peptides derived from ESAT-6 and CFP-10 proteins was used to detect the number of specific gamma interferon-producing T cells by means of an enzyme-linked immunospot assay. Sixty-eight individuals positive for M. tuberculosis infection, either human immunodeficiency virus-seropositive or seronegative, were studied. The test results were highly specific (87.5%) and sensitive (93.1%), more so than a classical lymphoproliferative assay (specificity and sensitivity of 77.27%), opening new possibilities for diagnosis and screening of tuberculosis. Moreover, the test allowed us to distinguish individuals infected with M. tuberculosis from those vaccinated with BCG. PMID- 15297486 TI - Interlaboratory comparison of results of susceptibility testing with caspofungin against Candida and Aspergillus species. AB - Seventeen laboratories participated in a study of interlaboratory reproducibility with caspofungin microdilution susceptibility testing against panels comprising 30 isolates of Candida spp. and 20 isolates of Aspergillus spp. The laboratories used materials supplied from a single source to determine the influence of growth medium (RPMI 1640 with or without glucose additions and antibiotic medium 3 [AM3]), the same incubation times (24 h and 48 h), and the same end point definition (partial or complete inhibition of growth) for the MIC of caspofungin. All tests were run in duplicate, and end points were determined both spectrophotometrically and visually. The results from almost all of the laboratories for quality control and reference Candida and Aspergillus isolates tested with fluconazole and itraconazole matched the NCCLS published values. However, considerable interlaboratory variability was seen in the results of the caspofungin tests. For Candida spp. the most consistent MIC data were generated with visual "prominent growth reduction" (MIC(2)) end points measured at 24 h in RPMI 1640, where 73.3% of results for the 30 isolates tested fell within a mode +/- one dilution range across all 17 laboratories. MIC(2) at 24 h in RPMI 1640 or AM3 also gave the best interlaboratory separation of Candida isolates of known high and low susceptibility to caspofungin. Reproducibility of MIC data was problematic for caspofungin tests with Aspergillus spp. under all conditions, but the minimal effective concentration end point, defined as the lowest caspofungin concentration yielding conspicuously aberrant hyphal growth, gave excellent reproducibility for data from 14 of the 17 participating laboratories. PMID- 15297487 TI - Characterization of multiple-antimicrobial-resistant Escherichia coli isolates from diseased chickens and swine in China. AB - Escherichia coli isolates from diseased piglets (n = 89) and chickens (n = 71) in China were characterized for O serogroups, virulence genes, antimicrobial susceptibility, class 1 integrons, and mechanisms of fluoroquinolone resistance. O78 was the most common serogroup identified (63%) among the chicken E. coli isolates. Most isolates were PCR positive for the increased serum survival gene (iss; 97%) and the temperature-sensitive hemagglutinin gene (tsh; 93%). The O serogroups of swine E. coli were not those typically associated with pathogenic strains, nor did they posses common characteristic virulence factors. Twenty three serogroups were identified among the swine isolates; however, 38% were O nontypeable. Overall, isolates displayed resistance to nalidixic acid (100%), tetracycline (98%), sulfamethoxazole (84%), ampicillin (79%), streptomycin (77%), and trimethoprim-sulfamethoxazole (76%). Among the fluoroquinolones, resistance ranged between 64% to levofloxacin, 79% to ciprofloxacin, and 95% to difloxacin. DNA sequencing of gyrA, gyrB, parC, and parE quinolone resistance-determining regions of 39 nalidixic acid-resistant E. coli isolates revealed that a single gyrA mutation was found in all of the isolates; mutations in parC together with double gyrA mutations conferred high-level resistance to fluoroquinolones (ciprofloxacin MIC, >/=8 microg/ml). Class 1 integrons were identified in 17 (19%) isolates from swine and 42 (47%) from chickens. The majority of integrons possessed genes conferring resistance to streptomycin and trimethoprim. These findings suggest that multiple-antimicrobial-resistant E. coli isolates, including fluoroquinolone-resistant variants, are commonly present among diseased swine and chickens in China, and they also suggest the need for the introduction of surveillance programs in China to monitor antimicrobial resistance in pathogenic bacteria that can be potentially transmitted to humans from food animals. PMID- 15297488 TI - Evaluation of DNA extraction and PCR methods for detection of Enterocytozoon bienuesi in stool specimens. AB - An evaluation of the sensitivities of three DNA extraction methods, i.e., FTA filter paper, a QIAamp stool mini kit, and a conventional phenol-chloroform method, by using specimens with known concentrations of Enterocytozoon bieneusi spores was performed. FTA filter paper and the QIAamp stool mini kit were the most sensitive methods, which could detect E. bieneusi in specimens with a concentration of 800 spores/ml. We also compared five previously described PCR methods that use five different primer pairs for the detection of E. bieneusi and showed that MSP3-MSP4B and EBIEF1-EBIER1 were the most sensitive primers. Although both sets of primers showed the same sensitivity, using the MSP3-MSP4B primers can directly provide genotypic information by sequencing. A blinded diagnostic test to compare PCR and light microscopy methods for the detection of E. bieneusi in stool specimens was also conducted. The use of FTA filter paper for DNA extraction together with the PCR method using the primer pair MSP3-MSP4B showed 100% sensitivity and 100% specificity for the detection of E. bieneusi in stool specimens, while the light microscopy method gave a sensitivity of 86.7% and a specificity of 100%. PMID- 15297489 TI - Rapid differentiation of Aspergillus species from other medically important opportunistic molds and yeasts by PCR-enzyme immunoassay. AB - We developed a PCR-based assay to differentiate medically important species of Aspergillus from one another and from other opportunistic molds and yeasts by employing universal, fungus-specific primers and DNA probes in an enzyme immunoassay format (PCR-EIA). Oligonucleotide probes, directed to the internal transcribed spacer 2 region of ribosomal DNA from Aspergillus flavus, Aspergillus fumigatus, Aspergillus nidulans, Aspergillus niger, Aspergillus terreus, Aspergillus ustus, and Aspergillus versicolor, differentiated 41 isolates (3 to 9 each of the respective species; P < 0.001) in a PCR-EIA detection matrix and gave no false-positive reactions with 33 species of Acremonium, Exophiala, Candida, Fusarium, Mucor, Paecilomyces, Penicillium, Rhizopus, Scedosporium, Sporothrix, or other aspergilli tested. A single DNA probe to detect all seven of the most medically important Aspergillus species (A. flavus, A. fumigatus, A. nidulans, A. niger, A. terreus, A. ustus, and A. versicolor) was also designed. Identification of Aspergillus species was accomplished within a single day by the PCR-EIA, and as little as 0.5 pg of fungal DNA could be detected by this system. In addition, fungal DNA extracted from tissues of experimentally infected rabbits was successfully amplified and identified using the PCR-EIA system. This method is simple, rapid, and sensitive for the identification of medically important Aspergillus species and for their differentiation from other opportunistic fungi. PMID- 15297490 TI - Species of the family Helicobacteraceae detected in an Australian sea lion (Neophoca cinerea) with chronic gastritis. AB - We describe the first case of gastritis in a male Australian sea lion (Neophoca cinerea) in which members of the family Helicobacteraceae, particularly the genus Wolinella, were detected. The sea lion exhibited clinical signs of gastrointestinal disease, including abdominal pain, lack of appetite, and lethargy. Examination of one ileal and five gastric biopsy specimens collected over a 10-year period revealed persistent fibrosis and/or superficial focal erosion and ulceration of the lamina propria. Spiral-shaped organisms 5 to 12 microm long were observed in two of the gut biopsy specimens. While Helicobacter species were detected by PCR in one of the gastric biopsy specimens, Wolinella species were detected in four of the five gastric specimens, including those in which spiral-shaped organisms were observed. Comparisons of biopsy specimen ribosomal DNA sequences with those obtained from the feces of this animal, the gastric tissue of a clinically healthy individual, and the feces of several other cohoused sea lions and fur seals revealed a separate and possibly novel gastric Helicobacter species. A possibly novel Wolinella species, along with Wolinella succinogenes, was also identified. These findings highlight the pathogenic potential of other members of this family in the etiopathogenesis of gastric disease in these animals. PMID- 15297491 TI - Clinical evaluation of the digene hybrid capture II test and the COBAS AMPLICOR monitor test for determination of hepatitis B virus DNA levels. AB - The measurement of hepatitis B virus (HBV) DNA is important for the assessment of liver disease and treatment efficacy. Most commercially available assays for the determination of HBV DNA levels have limited linear ranges. This study was performed to evaluate the clinical performance of the Digene Hybrid Capture II (Digene HC II assay) and the COBAS AMPLICOR Monitor test (COBAS-AM assay), with special emphasis on anti-HBV e antigen (HBeAg)-positive patients with low HBV DNA levels. A total of 425 Chinese patients with chronic hepatitis B were recruited. A total of 107 patients were HBeAg positive, and 318 patients were HBeAg negative. The Digene HC II assay and the COBAS-AM assay had similar intra-assay and interassay variabilities. A total of 264 patients (62.1%) had HBV DNA levels undetectable by the Digene HC II assay, and 47 patients (11.1%) had HBV DNA levels undetectable by the COBAS-AM assay (P < 0.001). For the 161 patients with HBV DNA levels detectable by the Digene HC II assay, the HBV DNA levels obtained by the Digene HC II assay and by the COBAS-AM assay showed an excellent correlation (r = 0.95; P < 0.001). The linear ranges of the Digene HC II assay and the COBAS-AM assay marginally overlapped. Before HBV DNA levels could be determined by the COBAS-AM assay, predilution had to be performed for 158 of 161 patients (98.1%) with HBV DNA levels detectable by the Digene HC II assay and for 10 of 264 patients (3.8%) with HBV DNA levels undetectable by the Digene HC II assay. The cost for assaying each serum sample by using different strategies was calculated. The COBAS-AM assay was more sensitive than the Digene HC II assay and more suitable for monitoring low levels of HBV viremia. PMID- 15297492 TI - Alanine-threonine polymorphism of Helicobacter pylori RpoB is correlated with differential induction of interleukin-8 in MKN45 cells. AB - Geographical differences in the genetic diversity of Helicobacter pylori isolates were examined by analyzing rpoB sequences. An extremely high level of allelic diversity among H. pylori strains was found. The rpoB sequences of Asian and non Asian (North and South American, European, and South African) strains were found to differ. An amino acid polymorphism (alanine and threonine RpoB types) was found at the 497th residue by deduced amino acid analysis. RpoB with a threonine residue (RpoB(Thr)) was uniquely present in East Asian countries, and two-thirds of the H. pylori isolate population in this region was RpoB(Thr); however, this type was rare or absent in Western countries, where RpoB(Ala) predominated. RpoB(Thr) strains induced a much larger amount of interleukin-8, a chemokine that plays an important role in chronic inflammation, than RpoB(Ala) strains in cultured MKN45 cells. PMID- 15297493 TI - Capillary electrophoretic restriction fragment length polymorphism patterns for the Mycobacterial hsp65 gene. AB - PCR-restriction fragment length polymorphism (RFLP) analysis is a nonprobe method for the rapid identification of Mycobacterium species. We demonstrate the separation of DNA or restriction fragments digested from the mycobacterial gene encoding the 65-kDa heat shock protein (hsp65) by capillary electrophoresis (CE). By using a pair of unlabeled primers, Tb11 and Tb12, and only one restriction enzyme, HaeIII, we investigated a total of 52 reference and clinical strains encompassing 12 Mycobacterium species. The electrophoretic separation of high resolution CE required <20 min and was capable of identifying fragments as small as 12 bp. A good agreement of measurement was observed between the sizes of restriction fragments resolved by CE, and the real sizes were deduced from the sequence analysis. Distinct differentiations were also well demonstrated between some species and subspecies by an extra HaeIII digestion site. With the advantage of the complete RFLP pattern available from CE, it appears to be more convenient to use an electropherogram rather than performing the cumbersome slab gel electrophoresis plus diagnostic algorithm to identify Mycobacterium species. Beyond the agarose and polyacrylamide gel electrophoresis, high-resolution CE provides an alternative for rapid identification of Mycobacterium species that is feasible for automation and routine use without the need for costly probes. PMID- 15297494 TI - Genetic variability of the G glycoprotein gene of human metapneumovirus. AB - Human metapneumovirus (hMPV) has been associated with respiratory illnesses like those caused by human respiratory syncytial virus (HRSV) infection. Similar to other pneumoviruses, genetic diversity has been reported for hMPV. Little information is currently available on the genetic variability of the G glycoprotein (G), which is the most variable gene in RSV and avian pneumovirus. The complete nucleotide sequences of the G open reading frame (ORF) of 24 Canadian hMPV isolates were determined. Phylogenetic analysis showed the existence of two major groups or clusters (1 and 2). All but one of the hMPV isolates that we examined belonged to cluster 1. Additional genetic variability was observed in cluster 1, which separated into two genetic subclusters. Within cluster 1 the nucleotide sequence identity for the G ORF was 74.2 to 100%, and the identity for the predicted amino acid sequence was 61.4 to 100%. The G genes of cluster 1 isolates were more divergent from the cluster 2 isolates, with 45.6 to 50.5% and 34.2 to 37.2% identity levels for the nucleotide and amino acid sequences, respectively. Sequence analysis also revealed changes in stop codon usage, resulting in G proteins of different lengths (217, 219, 228, and 236 residues). Western blot analysis with the use of hMPV-specific polyclonal antisera to each hMPV cluster showed significant antigenic divergence between the G proteins of clusters 1 and 2. These results suggest that the G protein of hMPV is continuously evolving and that the genetic diversity observed for the hMPV genes is reflected in the antigenic variability, similar to HRSV. PMID- 15297495 TI - Characterization of a trinucleotide repeat sequence (CGG)5 and potential use in restriction fragment length polymorphism typing of Mycobacterium tuberculosis. AB - The genomes of 28 bacterial strains, including mycobacterial species Mycobacterium tuberculosis and Mycobacterium bovis, were analyzed for the presence of a special class of microsatellite, that of trinucleotide repeat sequences (TRS). Results of a search of all 10 possible TRS motifs (i.e., CCT, CGG, CTG, GAA, GAT, GTA, GTC, GTG, GTT, and TAT) with five or more repeating units showed that (CGG)(5) was highly represented within the genomic DNA of M. tuberculosis and M. bovis. Most of the (CGG)(5) repeats in the genome were within the open reading frames of two large gene families encoding PE_PGRS and PPE proteins that have the motifs Pro-Glu (PE) and Pro-Pro-Glu (PPE). (CGG)(5)-probed Southern hybridization showed that some mycobacterial species, such as Mycobacterium marinum, Mycobacterium kansasii, and Mycobacterium szulgai, possess many copies of (CGG)(5) in their genomes. Analysis of clinical isolates obtained from Tokyo and Warsaw with both IS6110 and (CGG)(5) probes showed that there is an association between the fingerprinting patterns and the geographic origin of the isolates and that (CGG)(5) fingerprinting patterns were relatively more stable than IS6110 patterns. The (CGG)(5) repeat is a unique sequence for some mycobacterial species, and (CGG)(5) fingerprinting can be used as an epidemiologic method for these species as well as IS6110 fingerprinting can. If these two fingerprinting methods are used together, the precise analysis of M. tuberculosis isolates will be accomplished. (CGG)(5)-based fingerprinting is particularly useful for M. tuberculosis isolates with few or no insertion elements and for the identification of other mycobacterial species when informative probes are lacking. PMID- 15297497 TI - Use of a genus- and species-specific multiplex PCR for identification of enterococci. AB - The 16S rRNA gene has previously been used to develop genus-specific PCR primers for identification of enterococci. In addition, the superoxide dismutase gene (sodA) has been identified as a potential target for species differentiation of enterococci. In this study, Enterococcus genus-specific primers developed by Deasy et al. (E1/E2) were incorporated with species-specific primers based upon the superoxide dismutase (sodA) gene for development of a multiplex PCR. This assay provides simultaneous genus and species identification of 23 species of enterococci using seven different reaction mixtures. Accuracy of identification of the multiplex PCR was determined by comparisons to standard biochemical testing, the BBL Crystal kit, VITEK, and API Rapid ID 32 Strep. Isolates from swine feces, poultry carcasses, environmental sources, and retail food were evaluated and, overall, results for 90% of the isolates tested by PCR agreed with results obtained using standard biochemical testing and VITEK. Eighty-five percent and 82% of PCR results agreed with results from the API Rapid ID 32 Strep and BBL Crystal tests, respectively. With the exception of concurrence between identification using standard biochemical testing and VITEK (85%) and between BBL Crystal and VITEK (83%), the percent agreement for PCR was higher than or equal to any other pairwise comparison. Multiplex PCR for genus and species determination of enterococci provides an improved, rapid method for identification of this group of bacteria. PMID- 15297496 TI - Use of 16S ribosomal DNA PCR and denaturing gradient gel electrophoresis for analysis of the microfloras of healing and nonhealing chronic venous leg ulcers. AB - The bacterial microfloras of 8 healing and 10 nonhealing chronic venous leg ulcers were compared by using a combination of cultural analysis and denaturing gradient gel electrophoresis (DGGE) of PCR-amplified 16S rRNA gene products. Cultural analysis of the microflora revealed that the majority of both wound types carried the aerobes Staphylococcus and Pseudomonas spp. (89 and 80%, respectively). Sequencing of 16S ribosomal DNAs selected on the basis of DGGE profiling allowed the identification of strains not detected by cultural means. Of considerable interest was the finding that more than 40% of the sequences represented organisms not cultured from the wound from which they were amplified. DGGE profiles also revealed that all of the wounds possessed one apparently common band, identified by sequencing as Pseudomonas sp. The intensity of this PCR signal suggested that the bacterial load of nonhealing wounds was much higher for pseudomonads compared to healing wounds and that it may have been significantly underestimated by cultural analysis. Hence, the present study shows that DGGE could give valuable additional information about chronic wound microflora that is not apparent from cultural analysis alone. PMID- 15297498 TI - Comparison of the BBL CHROMagar Staph aureus agar medium to conventional media for detection of Staphylococcus aureus in respiratory samples. AB - Screening for Staphylococcus aureus has become routine in certain patient populations. This study is the first clinical evaluation of the BBL CHROMagar Staph aureus agar (CSA) medium (BD Diagnostics, Sparks, Md.) for detection of S. aureus in nasal surveillance cultures and in respiratory samples from cystic fibrosis (CF) patients. S. aureus colonies appear mauve on CSA. Other organisms are inhibited or produce a distinctly different colony color. S. aureus was identified from all media by slide coagulase, exogenous DNase, and mannitol fermentation assays. Susceptibility testing was performed using the agar dilution method. A total of 679 samples were evaluated. All samples were inoculated onto CSA. Nasal surveillance cultures were inoculated onto sheep blood agar (SBA) (BD Diagnostics), and samples from CF patients were inoculated onto mannitol salt agar (MSA) (BD Diagnostics). Of the 679 samples cultured, 200 organisms produced a mauve color on CSA (suspicious for S. aureus) and 180 were positive for S. aureus on SBA or MSA. Of 200 CSA-positive samples 191 were identified as S. aureus. Nine mauve colonies were slide coagulase negative and were subsequently identified as Staphylococcus lugdunensis (one), Staphylococcus epidermidis (three), Staphylococcus haemolyticus (one), and Corynebacterium species (four). CSA improved the ability to detect S. aureus by recovering 12 S. aureus isolates missed by conventional media. Of the 192 S. aureus isolates recovered, 122 were methicillin susceptible and 70 were methicillin resistant. Overall, the sensitivity and specificity of CSA in this study were 99.5 and 98%, respectively. There was no difference in the performance of the slide coagulase test or in susceptibility testing performed on S. aureus recovered from CSA compared to SBA or MSA. Our data support the use of CSA in place of standard culture media for detection of S. aureus in heavily contaminated respiratory samples. PMID- 15297499 TI - Characterization and prevalence of MefA, MefE, and the associated msr(D) gene in Streptococcus pneumoniae clinical isolates. AB - Recent work has shown that the efflux genes in Streptococcus pneumoniae that are responsible for acquired macrolide resistance can be distinguished as either mef(E) or mef(A). The genetic elements on which mef(A) and mef(E) are found also carry an open reading frame (ORF) that is 56% homologous to msr(A) in Staphylococcus. The prevalence of mef(A/E) and of the msr-like ORF [msr(D)] was evaluated in 153 mef(+) S. pneumoniae clinical isolates collected in North America, Europe, Africa, and Asia from 1997 to 2002. Clinical isolates were screened with PCR primers specific for either mef(A) or mef(E) and for msr(D). mef(A), mef(E), and msr(D) were cloned from mef(+) strains and transformed into a susceptible, competent strain of S. pneumoniae. The transformants were tested for antimicrobial susceptibilities and efflux pump induction. The results of this work demonstrated that mef(A) is more often isolated in parts of Europe, with some incidence in Canada, and that the msr-like gene alone can confer the efflux phenotype. PMID- 15297500 TI - Use of fluorescent amplified fragment length polymorphism for molecular epidemiology of leptospirosis in India. AB - Nineteen isolates of leptospires recovered from patients during three epidemics that occurred at different places and different times in the Andaman Islands and eight isolates from sporadic cases were characterized using serological and molecular genetic techniques. Group sera and monoclonal antibodies were used for antigenic characterization, whereas fluorescent amplified fragment length polymorphism (FAFLP) was used for genotyping. Of the 27 isolates, 19 were identified as belonging to serogroup Grippotyphosa, 3 belonged to serogroup Australis, 2 belonged to serogroup Icterohaemorrhagiae, and 1 each belonged to serogroups Hebdomadis, Canicola, and Sejroe. Analysis of FAFLP data grouped these 27 isolates into two main clusters of genotypes. One of the clusters, populated by 19 isolates, included 16 outbreak isolates. Seven of these 19 isolates belonged to serovar Ratnapura, 10 belonged to serovar Valbuzzi, and 1 each belonged to serovar Grippotyphosa and serovar Saxkoebing. Of the 27 patients from whom isolates were obtained, 9 had severe illness, and 6 of these 9 patients had pulmonary involvement, 1 had pulmonary and hepatorenal involvement, and the remaining 2 had hepatorenal involvement alone. Two patients out of the nine severe cases died subsequently. The isolates from sporadic cases showed great genetic diversity and were also diverse antigenically. Perhaps the strains belonging to a dominant genotype (the outbreak-associated cluster) possessed epidemic potential and higher virulence with a greater predilection to cause pulmonary complications than strains belonging to other genetic backgrounds. PMID- 15297501 TI - Verification of an assay for quantification of hepatitis C virus RNA by use of an analyte-specific reagent and two different extraction methods. AB - Protocols were designed for quantification and detection of hepatitis C virus (HCV) RNA by the use of an analyte-specific reagent (ASR) (Roche COBAS TaqMan48 [CTM48] HCV) after manual and automated RNA extraction. The purposes were to determine (i) assay performance characteristics using manual and automated RNA extraction methods, (ii) whether measurable range and limit of detection (LOD) of the ASR assay were influenced by genotype, and (iii) correlation of quantification by CTM48 HCV ASR and COBAS Monitor HCV v. 2.0. For HCV genotype 1 (Gt1), the lower limits of quantification after manual extraction were slightly lower than those for automated extraction (1.0 versus 1.5 log(10) IU/ml). Results were linear up to the highest concentration tested after extraction by both methods (manual, 6.1 log(10); automated, 6.4 log(10)). Similar results were obtained for Gt2 (1.8 to 6.8 log(10) IU/ml) and Gt3 (1.6 to 6.8 log(10) IU/ml) after automated extraction. The LOD of Gt1 virus was 10 IU/ml after manual extraction and between 25 and 37.5 IU/ml after automated extraction. Results with Gt2 and Gt3 viruses were similar after automated extraction (Gt2, between 25 and 50 IU/ml; Gt3, 25 IU/ml). Variability (intrarun and interrun, at concentrations throughout the range of quantification) was /=16 microg/ml) against M. furfur, M. restricta, M. globosa, and M. slooffiae strains, which were AM3 confirmed. Agreement of the two methods was 84 to 97%, and intraclass correlation coefficients were statistically significant (P < 0.001). Because of higher amphotericin B MICs provided by Etest for strains also displaying high BMD MICs (>/=1 microg/ml), agreement was poorer. The proposed media are used for the first time and can support optimum growth of eight Malassezia species for recording concordant BMD and Etest MICs. PMID- 15297503 TI - Genomic analysis distinguishes Mycobacterium africanum. AB - Mycobacterium africanum is thought to comprise a unique species within the Mycobacterium tuberculosis complex. M. africanum has traditionally been identified by phenotypic criteria, occupying an intermediate position between M. tuberculosis and M. bovis according to biochemical characteristics. Although M. africanum isolates present near-identical sequence homology to other species of the M. tuberculosis complex, several studies have uncovered large genomic regions variably deleted from certain M. africanum isolates. To further investigate the genomic characteristics of organisms characterized as M. africanum, the DNA content of 12 isolates was interrogated by using Affymetrix GeneChip. Analysis revealed genomic regions of M. tuberculosis deleted from all isolates of putative diagnostic and biological consequence. The distribution of deleted sequences suggests that M. africanum subtype II isolates are situated among strains of "modern" M. tuberculosis. In contrast, other M. africanum isolates (subtype I) constitute two distinct evolutionary branches within the M. tuberculosis complex. To test for an association between deleted sequences and biochemical attributes used for speciation, a phenotypically diverse panel of "M. africanum-like" isolates from Guinea-Bissau was tested for these deletions. These isolates clustered together within one of the M. africanum subtype I branches, irrespective of phenotype. These results indicate that convergent biochemical profiles can be independently obtained for M. tuberculosis complex members, challenging the traditional approach to M. tuberculosis complex speciation. Furthermore, the genomic results suggest a rational framework for defining M. africanum and provide tools to accurately assess its prevalence in clinical specimens. PMID- 15297504 TI - Amplified fragment length polymorphism reveals genomic variability among Mycobacterium avium subsp. paratuberculosis isolates. AB - Ninety-six primer sets were used for amplified fragment length polymorphism (AFLP) to characterize the genomes of 20 Mycobacterium avium subsp. paratuberculosis field isolates, 1 American Type Culture Collection (ATCC) M. avium subsp. paratuberculosis isolate (ATCC 19698), and 2 M. avium subsp. avium isolates (ATCC 35716 and Mac 104). AFLP analysis revealed a high degree of genomic polymorphism among M. avium subsp. paratuberculosis isolates that may be used to establish diagnostic patterns useful for the epidemiological tracking of M. avium subsp. paratuberculosis isolates. Four M. avium subsp. paratuberculosis polymorphic regions revealed by AFLP were cloned and sequenced. Primers were generated internal to these regions for use in PCR analysis and applied to the M. avium subsp. paratuberculosis field isolates. An appropriate PCR product was obtained in 79 of 80 reactions, while the M. avium subsp. avium isolates failed to act as templates for PCR amplification in seven of eight reactions. This work revealed the presence of extensive polymorphisms in the genomes of M. avium subsp. paratuberculosis and M. avium subsp. avium, many of which are based on deletions. Of the M. avium subsp. paratuberculosis-specific sequences studied, one revealed a 5,145-bp region with no homologue in the M. avium subsp. avium genome. Within this region are genes responsible for integrase-recombinase function. Three additional M. avium subsp. paratuberculosis-polymorphic regions were cloned, revealing a number of housekeeping genes; all were evaluated for their diagnostic and epidemiological value. PMID- 15297505 TI - Comparison of results of fluconazole disk diffusion testing for Candida species with results from a central reference laboratory in the ARTEMIS global antifungal surveillance program. AB - The accuracy of antifungal susceptibility tests is important for accurate resistance surveillance and for the clinical management of patients with serious infections. Our main objective was to compare the results of fluconazole disk diffusion testing of Candida spp. performed by ARTEMIS participating centers with disk diffusion and MIC results obtained by the central reference laboratory. A total of 2,949 isolates of Candida spp. were tested by NCCLS disk diffusion and reference broth microdilution methods in the central reference laboratory. These results were compared to the results of disk diffusion testing performed in the 54 participating centers. All tests were performed and interpreted following NCCLS recommendations. Overall categorical agreement between participant disk diffusion test results and reference laboratory MIC results was 87.4%, with 0.2% very major errors (VME) and 3.3% major errors (ME). The categorical agreement between the disk diffusion test results obtained in the reference laboratory with the MIC test results was similar: 92.8%. Likewise, good agreement was observed between participant disk diffusion test results and reference laboratory disk diffusion test results: 90.4%, 0.4% VME, and 3.4% ME. The disk diffusion test was especially reliable in detecting those isolates of Candida spp. that were characterized as resistant by reference MIC testing. External quality assurance data obtained by surveillance programs such as the ARTEMIS Global Antifungal Surveillance Program ensure the generation of useful surveillance data and result in the continued improvement of antifungal susceptibility testing practices. PMID- 15297506 TI - Development of two PCR-based techniques for detecting helical and coccoid forms of Helicobacter pylori. AB - The primary mode of transmission of Helicobacter pylori, a human pathogen carried by more than half the population worldwide, is still unresolved. Some epidemiological data suggest water as a possible transmission route. H. pylori in the environment transforms into a nonculturable, coccoid form, which frequently results in the failure to detect this bacterium in environmental samples by conventional culture techniques. To overcome limitations associated with culturing, molecular approaches based on DNA amplification by PCR have been developed and used for the detection of H. pylori in clinical and environmental samples. Our results showed the glmM gene as the most promising target for detection of H. pylori by PCR amplification. Under optimal amplification conditions, glmM-specific primers generated PCR-amplified products that were specific for H. pylori and some other Helicobacter species. Genome sequence analysis revealed the existence of a conserved region linked to a hypervariable region upstream of the 16S rRNA gene of H. pylori. Selective PCR primer sets targeting this sequence were evaluated for the specific detection of H. pylori. One primer set, Cluster2 and B1J99, were shown to be highly specific for H. pylori strains and did not produce any PCR products when other Helicobacter species and other bacterial species were analyzed. In tests with 32 strains of H. pylori, 6 strains of other Helicobacter species, 8 strains of Campylobacter jejuni, and 21 strains belonging to different genera, the primers for glmM were selective for the Helicobacter genus and the primers containing the region flanking the 16S rRNA gene were selective for H. pylori species only. The combination of two sensitive PCR-based methods, one targeting the glmM gene and the other targeting a hypervariable flanking region upstream of the 16S rRNA gene, are complementary to each other. Whereas the glmM-specific primers provide a rapid, sensitive presumptive assay for the presence of H. pylori and closely related Helicobacter spp., the primers for sequences flanking the 16S rRNA gene can confirm the presence of H. pylori and locate the potential source of this bacterium. PMID- 15297507 TI - Comparison of two urinary antigen tests for establishment of pneumococcal etiology of adult community-acquired pneumonia. AB - The Binax NOW immunochromatographic test (ICT) detecting the pneumococcal C polysaccharide and a serotype-specific latex agglutination (LA) test detecting 23 pneumococcal capsular antigens were evaluated for establishing pneumococcal etiology in community-acquired pneumonia (CAP) by use of nonconcentrated urine. ICT was considered to be strongly positive for result lines at least as intense as the control line and weakly positive for less intense result lines. When 215 adult CAP patients were tested, strong ICT, weak ICT, and LA positivity were found in 28, 24, and 16 patients, respectively; of these patients, 13 (46%), 6 (25%), and 13 (81%), respectively, had pneumococcal bacteremia and 27 (96%), 17 (71%), and 15 (94%), respectively, had Streptococcus pneumoniae isolated from blood, sputum, and/or nasopharynx. Among 108 controls tested, 2 (1.9%) were weakly ICT positive. When weak positivity was considered negative, the sensitivity of ICT decreased from 79% (19 of 24) to 54% (13 of 24), while the specificity increased from 83% (158 of 191) to 92% (176 of 191); no controls were false positive. The sensitivity and specificity of LA were 54% (13 of 24) and 98% (188 of 191), respectively. Eight of nine LA serotypes corresponded to culture serotypes. In conclusion, using nonconcentrated urine and dividing ICT-positive results into strongly and weakly positive results is a suitable way of performing ICT. While weak ICT positivity should be interpreted with caution, strong ICT positivity and LA positivity should be considered supportive of pneumococcal etiology in adult CAP. As such, these assays might have implications for antibiotic use in CAP. LA has promising potential for pneumococcal serotyping, although further evaluation is required. PMID- 15297508 TI - MICs of selected antibiotics for Bacillus anthracis, Bacillus cereus, Bacillus thuringiensis, and Bacillus mycoides from a range of clinical and environmental sources as determined by the Etest. AB - This paper presents Etest determinations of MICs of selected antimicrobial agents for 76 isolates of Bacillus anthracis chosen for their diverse histories and 67, 12, and 4 cultures, respectively, of its close relatives B. cereus, B. thuringiensis, and B. mycoides derived from a range of clinical and environmental sources. NCCLS breakpoints are now available for B. anthracis and ciprofloxacin, penicillin, and tetracycline; based on these breakpoints, the B. anthracis isolates were all fully susceptible to ciprofloxacin and tetracycline, and all except four cultures, three of which had a known history of penicillin resistance and were thought to originate from the same original parent, were susceptible to penicillin. Based on NCCLS interpretive standards for gram-positive and/or aerobic bacteria, all cultures were susceptible to amoxicillin-clavulanic acid and gentamicin and 99% (one with intermediate sensitivity) of cultures were susceptible to vancomycin. No group trends were apparent among the different categories of B. cereus (isolates from food poisoning incidents and nongastrointestinal infections and food and environmental specimens not associated with illness). Differences between B. anthracis and the other species were as expected for amoxicillin and penicillin, with all B. anthracis cultures, apart from the four referred to above, being susceptible versus high proportions of resistant isolates for the other three species. Four of the B. cereus and one of the B. thuringiensis cultures were resistant to tetracycline and a further six B. cereus and one B. thuringiensis cultures fell into the intermediate category. There was a slightly higher resistance to azithromycin among the B. anthracis strains than for the other species. The proportion of B. anthracis strains fully susceptible to erythromycin was also substantially lower than for the other species, although just a single B. cereus strain was fully resistant. The Etest compared favorably with agar dilution in a subsidiary test set up to test the readings, and it compared with other published studies utilizing a variety of test methods. PMID- 15297509 TI - Molecular epidemiology of hospital-associated and community-acquired Clostridium difficile infection in a Swedish county. AB - All episodes of Clostridium difficile associated diarrhea (CDAD) diagnosed in a defined population of 274,000 including one tertiary and two primary hospitals and their catchment areas were studied during 12 months. The annual CDAD incidence in the county was 97 primary episodes per 100,000, and 78% of all episodes were classified as hospital associated with a mean incidence of 5.3 (range, 1.4 to 6.5) primary episodes per 1,000 admissions. The incidence among hospitalized individuals was 1,300-fold higher than that in the community (33,700 versus 25 primary episodes per 100,000 persons per year), reflecting a 37-fold difference in antibiotic consumption (477 versus 13 defined daily doses [DDD]/1,000 persons/day) and other risk factors. Three tertiary hospital wards with the highest incidence (13 to 36 per 1,000) had CDAD patients of high age (median age of 80 years versus 70 years for other wards, P < 0.001), long hospital stay (up to 25 days versus 4 days), or a high antibiotic consumption rate (up to 2,427 versus 421 DDD/1,000 bed days). PCR ribotyping of C. difficile isolates available from 330 of 372 CDAD episodes indicated nosocomial acquisition of the strain in 17 to 27% of hospital-associated cases, depending on the time interval between index and secondary cases allowed (2 months or up to 12 months), and only 10% of recurrences were due to a new strain of C. difficile (apparent reinfection). In other words, most primary and recurring episodes were apparently caused by the patient's endogenous strain rather than by one of hospital origin. Typing also indicated that a majority of C. difficile strains belonged to international serotypes, and the distribution of types was similar within and outside hospitals and in primary and relapsing CDAD. However, type SE17 was an exception, comprising 22% of hospital isolates compared to 6% of community isolates (P = 0.008) and causing many minor clusters and a silent nosocomial outbreak including 36 to 44% of the CDAD episodes in the three high-incidence wards. PMID- 15297510 TI - New genome type of adenovirus serotype 4 caused nosocomial infections associated with epidemic conjunctivitis in Japan. AB - Human adenovirus type 4 is one of the major serotypes isolated from patients with adenoviral conjunctivitis. In 2001 we encountered nosocomial infections with epidemic conjunctivitis in the ophthalmology ward of one hospital in Sapporo, which is in the northern part of Japan. Adenoviruses were isolated from the patients with this nosocomial infection and identified as adenovirus type 4 (AdV 4) by a neutralization test with serotype-specific antiserum. When the cleavage patterns of the isolates were compared with the full viral genome with BamHI and SmaI, the cleavage patterns of the isolates were shown to be different from those of AdV-4p and other previously known AdV-4 variants. The nucleotide sequences of the fiber gene of the isolates showed the highest homologies (94.3%) with AdV-4 among the nucleotide sequences available from GenBank and formed a monophyletic cluster along with the prototype strain of AdV-4. The isolates, however, were located in a different lineage from those of AdV-4p and the AdV-4 variant from the sporadic infections. We conclude that the nosocomial infection that appeared in 2001 was caused by a new genome type of AdV-4, which was designated AdV-4c. PMID- 15297511 TI - Use of 16S rRNA gene sequencing for rapid confirmatory identification of Brucella isolates. AB - Members of the genus Brucella are categorized as biothreat agents and pose a hazard for both humans and animals. Current identification methods rely on biochemical tests that may require up to 7 days for results. We sequenced the 16S rRNA genes of 65 Brucella strains along with 17 related strains likely to present a differential diagnostic challenge. All Brucella 16S rRNA gene sequences were determined to be identical and were clearly different from the 17 related strains, suggesting that 16S rRNA gene sequencing is a reliable tool for rapid genus-level identification of Brucella spp. and their differentiation from closely related organisms. PMID- 15297512 TI - Rapid identification of Nocardia farcinica clinical isolates by a PCR assay targeting a 314-base-pair species-specific DNA fragment. AB - Nocardia farcinica is the most clinically significant species within the Nocardia asteroides complex. Differentiation of N. farcinica from other members of N. asteroides complex is important because this species characteristically demonstrates resistance to several extended-spectrum antimicrobial agents. Traditional phenotypic characterization of this species is time- and labor intensive and often leads to misidentification in the clinical microbiology laboratory. We previously observed a 409-bp product for all strains of N. farcinica by using randomly amplified polymorphic DNA analysis with the primer DKU49. In this investigation, the 409-bp fragment was sequenced and then used to design a specific primer pair, Nf1 (16-mer) and Nf2 (16-mer), complementary to the 409-bp fragment. PCR amplification of genomic DNA from 28 N. farcinica isolates with Nf1 and Nf2 generated a single intense 314-bp fragment. The specificity of the assay with these primers was verified, since there were no PCR amplification products observed from heterologous nocardial species (n = 59) or other related bacterial genera (n = 41). Restriction enzyme digestion using CfoI and direct sequencing of the 314-bp fragment further confirmed the specificity of the assay for N. farcinica. This highly sensitive and specific PCR assay provides a rapid (within 1 day of obtaining DNA) method for identification of this medically important emerging pathogen. Rapid diagnosis of N. farcinica infection may allow for earlier initiation of effective therapy, thus improving patient outcome. PMID- 15297513 TI - Comparison of a new lateral-flow chromatographic membrane immunoassay to viral culture for rapid detection and differentiation of influenza A and B viruses in respiratory specimens. AB - The performance of a new rapid lateral-flow chromatographic membrane immunoassay test kit for detection of influenza virus was evaluated and compared to that of viral culture in respiratory secretions collected from 400 adults and children seen at three large university hospitals during the recent 2003 influenza season. The rapid test provided results in 15 min, with excellent overall performance statistics (sensitivity, 94.4%; specificity, 100%; positive predictive value, 100%; negative predictive value, 97.5%). Both influenza A and B type viruses were reliably detected, with no significant difference in performance statistics noted by influenza virus type or by the center performing the test. PMID- 15297514 TI - Recommended test panel for differentiation of Klebsiella species on the basis of a trilateral interlaboratory evaluation of 18 biochemical tests. AB - Klebsiella pneumoniae and Klebsiella oxytoca are the two most frequently encountered Klebsiella species giving rise to infections in humans, but other Klebsiella species can also be found in clinical specimens: Klebsiella ozaenae, Klebsiella rhinoscleromatis, Klebsiella terrigena, Klebsiella planticola, Klebsiella ornithinolytica, and Enterobacter aerogenes (Klebsiella mobilis). However, many of these species are indistinguishable by the conventional methods employed routinely in the clinical microbiological laboratory. Several investigators have suggested various additional tests, but as yet there is no standardized test panel for identifying all Klebsiella species and subspecies. In the present study, performed in three national Klebsiella reference laboratories, we have evaluated a test panel consisting of 18 biochemical tests on 242 strains comprising all Klebsiella species and subspecies. The test panel was designed to identify organisms preliminarily identified as belonging to the genus Klebsiella on the basis of conventional methods or automated identification systems. With the described test panel it is possible to find one or more positive test results differentiating any Klebsiella species, except Klebsiella rhinoscleromatis, from its closest relative. PMID- 15297515 TI - Comparison of oligonucleotide ligation assay and consensus sequencing for detection of drug-resistant mutants of human immunodeficiency virus type 1 in peripheral blood mononuclear cells and plasma. AB - Drug-resistant mutants of human immunodeficiency virus type 1 (HIV-1) recede below the limit of detection of most assays applied to plasma when selective pressure is altered due to changes in antiretroviral treatment (ART). Viral variants with different mutations are selected by the new ART when replication is not suppressed or wild-type variants with greater replication fitness outgrow mutants following the cessation of ART. Mutants selected by past ART appear to persist in reservoirs even when not detected in the plasma, and when conferring cross-resistance they can compromise the efficacy of novel ART. Oligonucleotide ligation assay (OLA) of virus in plasma and peripheral blood mononuclear cells (PBMC) was compared to consensus sequence dideoxynucleotide chain terminator sequencing for detection of 91 drug resistance mutations that had receded below the limit of detection by sequencing of plasma. OLA of plasma virus detected 27.5% (95% confidence interval [CI], 19 to 39%) of mutant genotypes; consensus sequencing of the PBMC amplicon from the same specimen detected 23.1% (95% CI, 14 to 34%); and OLA of PBMC detected 53.8% (95% CI, 44 to 64%). These data suggest that concentrations of drug-resistant mutants were greater in PBMC than in plasma after changes in ART and indicate that the OLA was more sensitive than consensus sequencing in detecting low levels of select drug-resistant mutants. PMID- 15297516 TI - Population genetic analysis of Bartonella bacilliformis isolates from areas of peru where Carrion's disease is endemic and epidemic. AB - Carrion's disease is caused by infection with the alpha-proteobacterium Bartonella bacilliformis. Distribution of the disease is considered coincident with the distribution of its known vector, the sand fly Lutzomyia verrucarum. Recent epidemics of B. bacilliformis infections associated with atypical symptomatology in nonendemic regions have raised questions regarding the historic and present distribution of this bacterium and the scope of disease that infection causes. Phylogenetic relationships and genomic diversity of 18 B. bacilliformis isolates (10 isolates from a region where Carrion's disease is epidemic, Cuzco, Peru, and 8 isolates from a region where Carrion's disease is endemic, Caraz, Peru) were assessed using genomic data generated by infrequent restriction site PCR and gene sequence analysis of the flagellin gltA and ialB genes. A population genetic analysis of the genomic diversity suggests that what was once considered an epidemic region of Peru did not result from the recent introduction of B. bacilliformis. PMID- 15297517 TI - Use of fluorescence resonance energy transfer hybridization probes to evaluate quantitative real-time PCR for diagnosis of ocular toxoplasmosis. AB - Toxoplasma gondii infection is an important cause of chorioretinitis in Europe and the United States. Ophthalmological examination and a good clinical response to adequate therapy mainly support ocular toxoplasmosis diagnosis. However, clinical diagnostic may be difficult in some atypical cases. In these cases, laboratory confirmation, based on detection of local specific antibodies and parasite DNA by conventional PCR, is therefore important to confirm the disease etiology. More recently, real-time PCR has been developed to improve prenatal congenital toxoplasmosis diagnosis. We therefore examined the diagnostic value of quantitative real-time PCR for the detection of T. gondii in aqueous humor samples, associated with quantification of human beta-globin to control sample quantitative quality, by using a double fluorescence resonance energy transfer hybridization probes system with a double fluorescence reading. Of the 23 the clinically toxoplasmosis suspect patients, 22 showed serological evidence of exposure to Toxoplasma; one had a serological profile indicative of active infection. The analysis of paired aqueous humor and serum samples revealed an intraocular antibody production in 9 of 23 cases (39.1%). The quantitative real time PCR revealed positive and high parasite numbers and high Toxoplasma/human genome ratios in three cases. Furthermore, PCR was the only positive confirmatory test in two cases (11.1%). None of the patients included in the control group (n = 7) had evidence of either local specific antibody production or T. gondii DNA detection, suggesting a good relative assay specificity. On the whole, quantitative real-time PCR appears to be useful for diagnosing atypical ocular toxoplasmosis presentations. PMID- 15297518 TI - Use of tuf sequences for genus-specific PCR detection and phylogenetic analysis of 28 streptococcal species. AB - A 761-bp portion of the tuf gene (encoding the elongation factor Tu) from 28 clinically relevant streptococcal species was obtained by sequencing amplicons generated using broad-range PCR primers. These tuf sequences were used to select Streptococcus-specific PCR primers and to perform phylogenetic analysis. The specificity of the PCR assay was verified using 102 different bacterial species, including the 28 streptococcal species. Genomic DNA purified from all streptococcal species was efficiently detected, whereas there was no amplification with DNA from 72 of the 74 nonstreptococcal bacterial species tested. There was cross-amplification with DNAs from Enterococcus durans and Lactococcus lactis. However, the 15 to 31% nucleotide sequence divergence in the 761-bp tuf portion of these two species compared to any streptococcal tuf sequence provides ample sequence divergence to allow the development of internal probes specific to streptococci. The Streptococcus-specific assay was highly sensitive for all 28 streptococcal species tested (i.e., detection limit of 1 to 10 genome copies per PCR). The tuf sequence data was also used to perform extensive phylogenetic analysis, which was generally in agreement with phylogeny determined on the basis of 16S rRNA gene data. However, the tuf gene provided a better discrimination at the streptococcal species level that should be particularly useful for the identification of very closely related species. In conclusion, tuf appears more suitable than the 16S ribosomal RNA gene for the development of diagnostic assays for the detection and identification of streptococcal species because of its higher level of species-specific genetic divergence. PMID- 15297519 TI - High-throughput detection of pathogenic yeasts of the genus trichosporon. AB - The need for a rapid and accurate method for the detection of fungal pathogens has become imperative as the incidence of fungal infections has increased dramatically. Herein, we tested the Luminex 100, a novel flow cytometer, for the detection of the medically important genus Trichosporon. This genus was selected as our proof-of-concept model due to the close phylogenetic relationship between the species. The method, which is based on a nucleotide hybridization assay, consists of a combination of different sets of fluorescent beads covalently bound to species-specific capture probes. Upon hybridization, the beads bearing the target amplicons are classified by their spectral addresses with a 635-nm laser. Quantitation of the hybridized biotinylated amplicon is based on fluorescence detection with a 532-nm laser. We tested in various multiplex formats 48 species specific and group-specific capture probes designed in the D1/D2 region of ribosomal DNA, internal transcribed spacer regions, and intergenic spacer region. Species-specific biotinylated amplicons were generated with three sets of primers to yield fragments from the three regions. The assay was specific and fast, as it discriminated species differing by 1 nucleotide and required less than 50 min following amplification to process a 96-well plate. The sensitivity of the assay allowed the detection of 10(2) genome molecules in PCRs and 10(7) to 10(8) molecules of biotinylated amplification product. This technology provided a rapid means of detection of Trichosporon species with the flexibility to identify species in a multiplex format by combining different sets of beads. PMID- 15297520 TI - Comparison of the directigen flu A+B membrane enzyme immunoassay with viral culture for rapid detection of influenza A and B viruses in respiratory specimens. AB - The performance of a commercially available, rapid membrane enzyme immunoassay for influenza A and B virus detection was compared to that of viral culture in 4,092 respiratory specimens collected from patients presenting with respiratory symptoms during the 2002-2003 influenza season. The test's overall sensitivity was 43.83%, lower than previously reported but similar for detection of both influenza A and B viruses (42.98 versus 44.76%). However, specificity, 99.74%, was excellent for both influenza A and B viruses (99.82 versus 99.92%). These values make this test a very good confirmatory test when clinical suspicion is high, but a less accurate screening test for large populations. PMID- 15297521 TI - Use of 16S rRNA, 23S rRNA, and gyrB gene sequence analysis to determine phylogenetic relationships of Bacillus cereus group microorganisms. AB - In order to determine if variations in rRNA sequence could be used for discrimination of the members of the Bacillus cereus group, we analyzed 183 16S rRNA and 74 23S rRNA sequences for all species in the B. cereus group. We also analyzed 30 gyrB sequences for B. cereus group strains with published 16S rRNA sequences. Our findings indicated that the three most common species of the B. cereus group, B. cereus, Bacillus thuringiensis, and Bacillus mycoides, were each heterogeneous in all three gene sequences, while all analyzed strains of Bacillus anthracis were found to be homogeneous. Based on analysis of 16S and 23S rRNA sequence variations, the microorganisms within the B. cereus group were divided into seven subgroups, Anthracis, Cereus A and B, Thuringiensis A and B, and Mycoides A and B, and these seven subgroups were further organized into two distinct clusters. This classification of the B. cereus group conflicts with current taxonomic groupings, which are based on phenotypic traits. The presence of B. cereus strains in six of the seven subgroups and the presence of B. thuringiensis strains in three of the subgroups do not support the proposed unification of B. cereus and B. thuringiensis into one species. Analysis of the available phenotypic data for the strains included in this study revealed phenotypic traits that may be characteristic of several of the subgroups. Finally, our results demonstrated that rRNA and gyrB sequences may be used for discriminating B. anthracis from other microorganisms in the B. cereus group. PMID- 15297522 TI - Evaluation of the QuickLab RSV test, a new rapid lateral-flow immunoassay for detection of respiratory syncytial virus antigen. AB - Rapid respiratory syncytial virus (RSV) diagnosis is vital to the prevention of nosocomial RSV infections. We evaluated a new rapid lateral-flow RSV immunoassay, the QuickLab RSV test, that requires use of only one reagent. We compared QuickLab to the Directigen RSV (DIR) assay, which requires six reagents, and direct fluorescent antibody (DFA) testing. DFA results were considered the "gold standard." For 133 nasopharyngeal aspirates tested, DFA results were 77 (57.8%) positive, 47 (35.3%) negative, and 9 (6.8%) indeterminate. The sensitivities, specificities, positive predictive values, and negative predictive values of QuickLab and DIR tests were 93.3% (70 of 75) and 80.8% (59 of 73), 95.6% (43 of 45) and 100.0% (46 of 46), 97.2% (70 of 72) and 100.0% (59 of 59), and 89.6% (43 of 48) and 76.7% (46 of 60), respectively. QuickLab was significantly (P = 0.02) more sensitive than DIR; the difference in specificities was not significant. DFA was more sensitive than DIR (P < 0.001) but not more sensitive than QuickLab (P = 0.45). The results of DIR testing were initially uninterpretable and required retesting with 15% of the specimens compared to 3% of QL results (P < 0.001). We conclude that the QuickLab RSV test has sensitivity similar to that of the DFA assay and better than that of the DIR assay. QuickLab testing is also simpler to perform and interpret than both DFA and DIR testing. PMID- 15297523 TI - Use of positive blood cultures for direct identification and susceptibility testing with the vitek 2 system. AB - In order to further decrease the time lapse between initial inoculation of blood culture media and the reporting of results of identification and antimicrobial susceptibility tests for microorganisms causing bacteremia, we performed a prospective study in which specially processed fluid from positive blood culture bottles from Bactec 9240 (Becton Dickinson, Cockeysville, Md.) containing aerobic media were directly inoculated into Vitek 2 system cards (bio-Merieux, France). Organism identification and susceptibility results were compared with those obtained from cards inoculated with a standardized bacterial suspension obtained following subculture to agar; 100 consecutive positive monomicrobic blood cultures, consisting of 50 gram-negative rods and 50 gram-positive cocci, were included in the study. For gram-negative organisms, 31 of the 50 (62%) showed complete agreement with the standard method for species identification, while none of the 50 gram-positive cocci were correctly identified by the direct method. For gram-negative rods, there were 50% categorical agreements between the direct and standard methods for all drugs tested. The very major error rate was 2.4%, and the major error rate was 0.6%. The overall error rate for gram negatives was 6.6%. Complete agreement in clinical categories of all antimicrobial agents evaluated was obtained for 19 of 50 (38%) gram-positive cocci evaluated; the overall error rate was 8.4%, with 2.8% minor errors, 2.4% major errors, and 3.2% very major errors. These findings suggest that the Vitek 2 cards inoculated directly from positive Bactec 9240 bottles do not provide acceptable bacterial identification or susceptibility testing in comparison with corresponding cards tested by a standard method. PMID- 15297524 TI - Performance characteristics of a quantitative TaqMan hepatitis C virus RNA analyte-specific reagent. AB - We determined the dynamic range, reproducibility, accuracy, genotype bias, and sensitivity of the TaqMan hepatitis C virus (HCV) analyte-specific reagent (ASR). Serum samples were processed using the MagNA Pure LC instrument and run on the COBAS TaqMan 48 analyzer. The performance characteristics of the ASR were also compared with those of the qualitative AMPLICOR and quantitative AMPLICOR MONITOR HCV tests. The ASR exhibited a >/=6-log(10) linear dynamic range and excellent reproducibility, with a mean coefficient of variation of 14%. HCV RNA concentration measured with the ASR agreed within an average of 0.42 log(10) (2.6 fold) of the labeled concentration with members of a standard reference panel. HCV genotypes 1 to 4 were amplified with similar efficiencies with the ASR. The ASR and AMPLICOR MONITOR viral load results were significantly correlated (r = 0.8898; P < 0.01), but the agreement was poor (mean difference, 0.45 +/- 0.35 log(10)) for 72 HCV RNA-positive clinical samples. However, 98.9% agreement between the ASR and qualitative AMPLICOR test results was found with 60 positive and 29 negative samples. Limiting-dilution experiments demonstrated that the limits of detection for ASR and AMPLICOR tests were 84 and 26 IU/ml, respectively. The performance characteristics of the TaqMan HCV ASR are appropriate for all clinical applications of HCV RNA testing. PMID- 15297525 TI - Multicenter evaluation of use of penicillin and ampicillin as surrogates for in vitro testing of susceptibility of enterococci to imipenem. AB - Imipenem is approved by the U.S. Food and Drug Administration (FDA) for treatment of infections caused by Enterococcus faecalis. However, there are no NCCLS guidelines for testing susceptibility of enterococci against imipenem. To assess whether or not ampicillin or penicillin could be used as a surrogate for broth microdilution (BMD) testing of imipenem versus Enterococcus species, 633 strains of E. faecalis, E. faecium, and other enterococci isolated from blood cultures of patients at three geographically distinct university hospitals were tested by the NCCLS BMD and disk diffusion (DD) methods. Using FDA susceptibility breakpoints for imipenem and NCCLS breakpoints for penicillin and ampicillin, categorical agreement (CA) for penicillin-imipenem and ampicillin-imipenem tested with E. faecalis and E. faecium by BMD was >/=94% but was /=98% and was 92% for other enterococci; CA for penicillin-imipenem was 91% for E. faecalis, 98% for E. faecium, and 87% for other enterococci. Further analysis showed that testing E. faecalis with ampicillin resulted in no false-susceptible (FS) or false-resistant (FR) results by BMD, no FS results by DD, and a single FR result by DD (0.2%), whereas testing with penicillin resulted in no FS results by BMD or DD and two FR results by BMD (0.4%). For E. faecium and other enterococci, the combination of FS and FR results was such that surrogate testing with penicillin or ampicillin appears not to be sufficiently reliable to be used clinically. We conclude that ampicillin is an accurate predictor of the in vitro activity of imipenem against E. faecalis. PMID- 15297526 TI - Molecular characteristics of nosocomial and Native American community-associated methicillin-resistant Staphylococcus aureus clones from rural Wisconsin. AB - In central and northern Wisconsin methicillin-resistant Staphylococcus aureus (MRSA) was first detected in 1989. Over the next 10-year period, 581 MRSA isolates were collected, 17.2% of which came from patients who were treated at five Native American clinics. These isolates were typed by SmaI-macrorestricted pulsed-field gel electrophoresis (PFGE). The PFGE patterns clustered the isolates into six major clonal groups (MCGs), i.e., MCGs 1 to 6, and 19 minor clonal groups (mCGs). The 25 clonal groups were represented by 109 unique PFGE types. Sixty-five percent of the MCG-2 isolates were recovered from patients who were treated at Native American clinics. Ninety-four percent of the MCG-2 isolates harbored the staphylococcal cassette chromosome mec (SCCmec) IVa. These isolates also had PFGE profiles that were clonally related to the midwestern community associated MRSA (CA-MRSA) strain, MW2. The representative isolates from MCG-2 had the multilocus sequence type allelic profile 1-1-1-1-1-1-1 and contained pvl genes. They were also susceptible to various antibiotics, a finding consistent with the CA-MRSA phenotype. SCCmec IV was also present in other mCGs. Unlike MCG 2, isolates from the remaining five MCGs harbored SCCmec II and were resistant to multiple antibiotics, suggesting their nosocomial origin. The 19 mCGs were represented by diverse SCCmec types and three putative new variants referred to as SCCmec Ib, IIa, and IIb. PMID- 15297527 TI - Comparison of real-time PCR signal-amplified in situ hybridization and conventional PCR for detection and quantification of human papillomavirus in archival cervical cancer tissue. AB - Archival paraffin-embedded tumor specimens offer a wealth of information for both cancer research and for routine clinical applications. However, the use of formalin-fixed, paraffin-embedded specimens for quantitative real-time PCR is not yet a standard diagnostic method in many laboratories, in particular for the quantification of human papillomavirus (HPV). Particularly high-risk HPV types are involved in almost 100% of the carcinogenesis of cervical cancer. We compared the diagnostic applicability and sensitivity of real-time PCR to that of chromogenic tyramide-signal-amplified in situ hybridization and conventional PCR for the detection of HPV from archival tissue in 164 cases of carcinoma in situ and cervical cancer. Furthermore, we examined whether the viral load of HPV is of prognostic relevance. Our findings indicate that patients in tumor stage I with a lower viral load of HPV type 16 (HPV16; up to 1,000 copies/ng of DNA) had a significantly better survival than HPV 16-negative patients (P = 0.037). We observed a greater sensitivity of both real-time PCR and conventional PCR for the detection of HPV16 and -18 compared to signal amplified in situ hybridization. We found a considerable concordance between HPV16 (kappa = 0.661) and HPV18 (kappa = 0.781) status as measured by real-time PCR and conventional PCR, indicating similar sensitivities. We recognized an inhibitory effect of formalin fixation and paraffin embedding on the evaluation of real-time PCR quantification. PMID- 15297528 TI - Use of DNA microarrays for rapid genotyping of TEM beta-lactamases that confer resistance. AB - Standard clinical procedures for pathogen resistance identification are laborious and usually require 2 days of cultivation before the resistance can be determined unequivocally. In contrast, clinicians and patients face increasing threats from antibiotic-resistant pathogenic bacteria in terms of their frequencies and levels of resistance. A major class of microbial resistance stems from the occurrence of beta-lactamases, which, if mutated, can cause the severe extended-spectrum beta lactamase (ESBL) or inhibitor-resistant TEM (IRT) phenotype, which cause resistance to extended-spectrum cephalosporins, monobactams, and beta-lactamase inhibitors. We describe an oligonucleotide microarray for identification of the single nucleotide polymorphisms (SNPs) of 96% of the TEM beta-lactamase variants described to date which are related to the ESBL and/or IRT phenotype. The target DNA, originating from Escherichia coli, Enterobacter cloacae, and Klebsiella pneumoniae cells isolated from clinical samples, was amplified and fluorescently labeled by PCR with consensus primers in the presence of cyanine 5-labeled nucleotides. The total assay, including PCR, hybridization, and image analysis, could be performed in 3.5 h. The microarray results were validated by standard clinical procedures. The microarray outperformed the standard procedures in terms of assay time and the depth of information provided. In conclusion, this array offers an attractive option for the identification and epidemiologic monitoring of TEM beta-lactamases in the routine clinical diagnostic laboratory. PMID- 15297529 TI - Concurrent infections with vector-borne pathogens associated with fatal hemolytic anemia in a cattle herd in Switzerland. AB - Bovine anaplasmosis is a vector-borne disease that results in substantial economic losses in other parts of the world but so far not in northern Europe. In August 2002, a fatal disease outbreak was reported in a large dairy herd in the Swiss canton of Grisons. Diseased animals experienced fever, anorexia, agalactia, and depression. Anemia, ectoparasite infestation, and, occasionally, hemoglobinuria were observed. To determine the roles of vector-borne pathogens and to characterize the disease, blood samples were collected from all 286 animals: 50% of the cows were anemic. Upon microscopic examination of red blood cells, Anaplasma marginale inclusion bodies were found in 47% of the cows. The infection was confirmed serologically and by molecular methods. Interestingly, we also found evidence of infections with Anaplasma phagocytophilum, large Babesia and Theileria spp., and Mycoplasma wenyonii. The last two species had not previously been described in Switzerland. Anemia was significantly associated with the presence of the infectious agents detected, with the exception of A. phagocytophilum. Remarkably, concurrent infections with up to five infectious vector-borne agents were detected in 90% of the ill animals tested by PCR. We concluded that A. marginale was the major cause of the hemolytic anemia, while coinfections with other agents exacerbated the disease. This was the first severe disease outbreak associated with concurrent infections with vector-borne pathogens in alpine Switzerland; it was presumably curtailed by culling of the entire herd. It remains to be seen whether similar disease outbreaks will have to be anticipated in northern Europe in the future. PMID- 15297530 TI - High prevalence of Helicobacter Species detected in laboratory mouse strains by multiplex PCR-denaturing gradient gel electrophoresis and pyrosequencing. AB - Rodent models have been developed to study the pathogenesis of diseases caused by Helicobacter pylori, as well as by other gastric and intestinal Helicobacter spp., but some murine enteric Helicobacter spp. cause hepatobiliary and intestinal tract diseases in specific inbred strains of laboratory mice. To identify these murine Helicobacter spp., we developed an assay based on PCR denaturing gradient gel electrophoresis and pyrosequencing. Nine strains of mice, maintained in four conventional laboratory animal houses, were assessed for Helicobacter sp. carriage. Tissue samples from the liver, stomach, and small intestine, as well as feces and blood, were collected; and all specimens (n = 210) were screened by a Helicobacter genus-specific PCR. Positive samples were identified to the species level by multiplex denaturing gradient gel electrophoresis, pyrosequencing, and a H. ganmani-specific PCR assay. Histologic examination of 30 tissue samples from 18 animals was performed. All mice of eight of the nine strains tested were Helicobacter genus positive; H. bilis, H. hepaticus, H. typhlonius, H. ganmani, H. rodentium, and a Helicobacter sp. flexispira-like organism were identified. Helicobacter DNA was common in fecal (86%) and gastric tissue (55%) specimens, whereas samples of liver tissue (21%), small intestine tissue (17%), and blood (14%) were less commonly positive. Several mouse strains were colonized with more than one Helicobacter spp. Most tissue specimens analyzed showed no signs of inflammation; however, in one strain of mice, hepatitis was diagnosed in livers positive for H. hepaticus, and in another strain, gastric colonization by H. typhlonius was associated with gastritis. The diagnostic setup developed was efficient at identifying most murine Helicobacter spp. PMID- 15297531 TI - Scytalidium dimidiatum causing recalcitrant subcutaneous lesions produces melanin. AB - Scytalidium dimidiatum is a pigmented dematiaceous coelomycete that typically causes chronic superficial skin diseases and onychomycosis, as well as deeper infections, such as subcutaneous abscesses, mycetoma, and even fungemia in immunocompromised patients. A second species, Scytalidium hyalinum, has hyaline hyphae and arthroconidia and is considered by some authors to be an albino mutant of S. dimidiatum. This study aimed to confirm the presence of melanin or melanin like compounds (which have been previously implicated in the virulence of other fungal pathogens) in S. dimidiatum from a patient with multiple subcutaneous nodules. Treatment of the hyphae and arthroconidia with proteolytic enzymes, denaturant, and concentrated hot acid yielded dark particles, which were stable free radicals, consistent with their identification as melanins. Extracted melanin particles from S. dimidiatum cultures were labeled by melanin-binding monoclonal antibodies (MAbs) from Sporothrix schenckii, Aspergillus fumigatus, and Cryptococcus neoformans. Lesional skin from the patient infected with S. dimidiatum contained fungal cells that were labeled by melanin-binding MAbs, and digestion of the tissue yielded dark particles that were also reactive. S. hyalinum was also subjected to the melanin extraction protocol, but no dark particles were yielded. PMID- 15297532 TI - Assessment of a new selective chromogenic Bacillus cereus group plating medium and use of enterobacterial autoinducer of growth for cultural identification of Bacillus species. AB - A new chromogenic Bacillus cereus group plating medium permits differentiation of pathogenic Bacillus species by colony morphology and color. Probiotic B. cereus mutants were distinguished from wild-type strains by their susceptibilities to penicillin G or cefazolin. The enterobacterial autoinducer increased the sensitivity and the speed of enrichment of B. cereus and B. anthracis spores in serum-supplemented minimal salts medium (based on the standard American Petroleum Institute medium) and buffered peptone water. PMID- 15297533 TI - Identification of Escherichia coli O114 O-antigen gene cluster and development of an O114 serogroup-specific PCR assay. AB - Screening for the Escherichia coli O serotype is the traditional test for identification of E. coli clones. The O-antigen gene cluster of the E. coli O114 type strain was sequenced, and 12 open reading frames were assigned functions on the basis of homology. By screening against all 186 E. coli and Shigella O serotypes, five genes specific to E. coli O114 were identified. A PCR assay based on the O-antigen-specific genes was developed and tested on 41 clinical isolates of E. coli O114. The PCR assay was shown to be highly specific and sensitive. When tested with pork and water samples, as few as 0.12 CFU of E. coli O114 g(-1) were detected. Thus, the PCR assays established in this study can be used to reliably identify E. coli O114 strains and may also be used to detect E. coli O114 strains in food, water, and other environmental samples. PMID- 15297534 TI - Molecular epidemiology of Enterobacteriaceae isolates producing extended-spectrum beta-lactamases in a French hospital. AB - In 2002, 80 isolates of Enterobacteriaceae producing extended-spectrum beta lactamases (ESBLs) were collected from infected patients in our hospital. Enterobacter aerogenes was the most common bacterium isolated from all specimens (36.5%). The ESBLs were predominantly (90%) TEM derivatives (TEM-24, TEM-3). Pulsed-field gel electrophoresis highlighted that E. aerogenes, Klebsiella pneumoniae, and Citrobacter koseri had a clonal propagation. PMID- 15297535 TI - Comparison of sequence analysis and a novel discriminatory real-time PCR assay for detection and quantification of Lamivudine-resistant hepatitis B virus strains. AB - We report a rapid and accurate real-time PCR-based method to quantify wild-type and lamivudine-resistant hepatitis B virus by using a common forward primer paired with different reverse primers. Excellent concordance was demonstrated between sequencing and the discriminatory real-time assay; however, a mixture of quasispecies was more frequently detected by discriminatory real-time PCR. PMID- 15297536 TI - Detection of Haemophilus influenzae type b by real-time PCR. AB - A real-time PCR assay targeting the capsulation locus of Haemophilus influenzae type b (Hib) was developed. The linear detection range was from 1 to 10(6) microorganisms per reaction mixture. No H. influenzae other than Hib or any other control bacteria typically found in the upper respiratory tract was detected. PMID- 15297537 TI - Isolation of Bartonella rattimassiliensis sp. nov. and Bartonella phoceensis sp. nov. from European Rattus norvegicus. AB - Thirty-three isolates of Bartonella spp., including 11 isolates not belonging to previously known species, were isolated from 66 Rattus norvegicus subjects trapped in the city of Marseille, France. Based on seven different gene sequences, the 11 isolates were assigned to Bartonella rattimassiliensis sp. nov. and Bartonella phoceensis sp. nov. PMID- 15297538 TI - Differentiation of the major Listeria monocytogenes serovars by multiplex PCR. AB - A new multiplex PCR assay was developed to separate the four major Listeria monocytogenes serovars isolated from food and patients (1/2a, 1/2b, 1/2c, and 4b) into distinct groups. The PCR test, which constitutes a rapid and practical alternative to laborious classical serotyping, was successfully evaluated with 222 Listeria strains. PMID- 15297539 TI - Analysis of p51, groESL, and the major antigen P51 in various species of Neorickettsia, an obligatory intracellular bacterium that infects trematodes and mammals. AB - The p51 gene that encodes the major antigenic 51-kDa protein in Neorickettsia risticii was identified in strains of Neorickettsia sennetsu and the Stellantchasmus falcatus agent but not in Neorickettsia helminthoeca, suggesting that p51-based diagnosis would be useful to distinguish among them. groESL sequencing results delineated the phylogenic relationships among Neorickettsia spp. PMID- 15297540 TI - Real-time TaqMan PCR for quantifying oral bacteria during biofilm formation. AB - A TaqMan PCR was developed for quantifying early colonizer microorganisms in dental biofilms. To design species-specific primers and TaqMan probes, genomic subtractive hybridization was used. This quantitative assay in combination with subtractive hybridization may be of value in the study of microbial ecosystems consisting of related species that are involved in the formation and etiology of biofilms. PMID- 15297541 TI - In vivo acquisition of high-level resistance to imipenem in Escherichia coli. AB - Four clonally related Escherichia coli strains were isolated successively from bile duct of a girl suffering from sclerosing cholangitis. One of them, selected after an imipenem-containing regimen, was resistant to carbapenems and to broad spectrum cephalosporins due to a plasmid-mediated cephalosporinase, CMY-2, and the lack of outer membrane proteins OmpF and OmpC. PMID- 15297542 TI - Evaluation of Granada agar plate for detection of Streptococcus agalactiae in urine specimens from pregnant women. AB - The Granada agar plate (GAP; Biomedics SL, Madrid, Spain) was evaluated for the detection of group B streptococci (GBS) in urine specimens from pregnant women submitted for testing for asymptomatic bacteriuria and was compared with blood agar (BA [Columbia agar with 5% sheep blood]; bioMerieux, Marcy l'Etoile, France). The GAP detected 103 out of 105 GBS, whereas BA detected only 50. Use of the GAP could be a good method for the detection of GBS in urine specimens from pregnant women. PMID- 15297543 TI - Multicenter evaluation of a new screening test that detects Clostridium difficile in fecal specimens. AB - Clostridium difficile causes approximately 25% of nosocomial antibiotic associated diarrheas and most cases of pseudomembranous colitis. We evaluated C. DIFF CHEK, a new screening test that detects glutamate dehydrogenase of C. difficile. Our results showed that this test was comparable to PCR in sensitivity and specificity and outperformed bacterial culture. PMID- 15297544 TI - Rapid and specific detection of the O15:K52:H1 clonal group of Escherichia coli by gene-specific PCR. AB - Primers specific for Escherichia coli O15:K52:H1 were devised based on a novel single-nucleotide polymorphism identified within the housekeeping gene fumC, i.e., G594A. In experiments comparing various reference typing methods, the new primers provided 100% sensitivity and specificity for the O15:K52:H1 clonal group, including 162 diverse clinical and reference E. coli isolates. PMID- 15297545 TI - Clonal spread of emm type 28 isolates of Streptococcus pyogenes that are multiresistant to antibiotics. AB - Fifty-three pharyngitis-related and invasive isolates of Streptococcus pyogenes that are resistant to bacitracin were collected. They were also resistant to streptomycin, kanamycin, macrolides, lincosamides, and streptogramin B. These multiresistant isolates were of emm type 28 and clonally related as shown by pulsed-field gel electrophoresis. PMID- 15297546 TI - Persistence of human immunodeficiency virus type 1 subtype B DNA in dried-blood samples on FTA filter paper. AB - The stability of human immunodeficiency virus type 1 (HIV-1) DNA in whole blood collected on filter paper (FTA Card) was evaluated. After >4 years of storage at room temperature in the dark our qualitative assay detected virus at a rate similar to that of our initial test (58 of 60, 97%; P = 0.16), suggesting long term HIV-1 DNA stability. PMID- 15297547 TI - Use of rapid and conventional testing technologies for human immunodeficiency virus type 1 serologic screening in a rural Kenyan reference laboratory. AB - We report a prospective comparison of human immunodeficiency virus type 1 testing by enzyme immunoassay and Western blot with four rapid tests of 486 subjects performed in rural Kenya. Rapid test sensitivity was 100%. Specificity ranged from 99.1 to 100%. Combined use of two Food and Drug Administration-approved rapid tests yielded a single false-positive result. PMID- 15297548 TI - Evaluation of Xenostrip-Tv, a rapid diagnostic test for Trichomonas vaginalis infection. AB - An immunochromatographic strip test, Xenostrip-Tv, was compared to wet mount and PCR for the diagnosis of Trichomonas vaginalis infection in women. Of 428 specimens tested, 54 (12.6%) were positive by an "expanded gold standard," defined as either a positive wet mount and PCR test with primers TVK3 and TVK7 and/or a positive PCR test confirmed by a second PCR assay with primers TVA5-1 and TVA6; 26 (6%) were positive by wet mount, and 36 (8.4%) were positive by Xenostrip-Tv test. Since the Xenostrip-Tv test is rapid and easy to perform and proved to be more sensitive than wet mount, it should be considered as an alternative to wet mount for point-of-care diagnosis of trichomoniasis, especially in settings where microscopy is impractical. PMID- 15297549 TI - V-antigen genotype and phenotype analyses of clinical isolates of Pseudomonas aeruginosa. AB - The pcrV genotype was analyzed in clinical isolates of Pseudomonas aeruginosa which showed a negative phenotype for secretion of V-antigen PcrV. The suppression of PcrV secretion in these isolates was due not to a lack of the pcrV gene but rather to suppression of PcrV expression. PMID- 15297550 TI - Detection of human papillomavirus DNA in cervical samples: analysis of the new PGMY-PCR compared to the hybrid capture II and MY-PCR assays and a two-step nested PCR assay. AB - The PGMY-PCR for human papillomavirus (HPV) was evaluated, in parallel with nested PCR (nPCR), in samples with noted Hybrid Capture II (HCII) and MY-PCR results. PGMY-PCR detected HPV DNA in 2.5% of HCII-negative-MY-PCR-negative samples and in 71.7% of HCII-positive-MY-PCR-negative samples; also, it detected the MY-PCR-negative-nPCR-negative types HPV-42, HPV-44, HPV-51, HPV-87, and HPV 89. PMID- 15297551 TI - Cryptococcus infection in tropical Australia. AB - Eighteen cases of disease caused by the saprophytic fungi Cryptococcus neoformans and Cryptococcus bacillisporus are described from the Northern Territory of Australia. The majority of infections were with Cryptococcus bacillisporus and in the rural Aboriginal population, often causing pulmonary mass lesions. PMID- 15297552 TI - Modified multiplex PCR method for detection of pyrogenic exotoxin genes in staphylococcal isolates. AB - A modified multiplex PCR method for detection of nine Staphylococcus aureus enterotoxin genes (sea, seb, sec, sed, see, seg, seh, sei, and sej) and one form of immunoreactive toxic shock syndrome toxin based on a previously published method (S. R. Monday and G. A. Bohach, J. Clin. Microbiol. 37:3411-3414, 1999) has been developed. The modified PCR protocol seems robust and gives reliable results. PMID- 15297553 TI - Distribution of Porphyromonas gingivalis biotypes defined by alleles of the kgp (Lys-gingipain) gene. AB - Paired subgingival plaque samples representing the most-diseased and least diseased sites were collected from 34 adult patients with diagnosed chronic periodontitis. The percentage of Porphyromonas gingivalis relative to the total anaerobic and gram-negative bacterial load at each site was determined by real time PCR. Based on variations in the noncatalytic C terminus of the Lys-gingipain (Kgp), it was reasoned that DNA sequence variation in the 3'-coding region of the kgp gene might determine functional biotypes. Perusal of the available sequence information in GenBank indicated three such forms of the kgp gene corresponding to P. gingivalis strains HG66, 381, and W83. Analysis of patient samples revealed the presence of a fourth genotype (W83v) that showed duplication of a sequence recognized by the W83 reverse primer. The four biotypes, HG66, 381, W83, and W83v, were present in the study group in the ratio 8:11:6:5, respectively. Each subject was colonized by one predominant biotype, and only three patients were colonized by a trace amount of a second biotype. PMID- 15297554 TI - Surveillance of methicillin-resistant Staphylococcus aureus in a pediatric hospital in Mexico City during a 7-year period (1997 to 2003): clonal evolution and impact of infection control. AB - Between 1997 and 2000 a single multidrug-susceptible methicillin-resistant Staphylococcus aureus clone, M (sequence type 30 [ST30]-staphylococcal cassette chromosome mec [SCCmec] type IV), was present in a pediatric hospital in Mexico City, Mexico. In 2001 the international multidrug-resistant New York-Japan clone (ST5-SCCmec type II) was introduced into the hospital, completely replacing clone M by 2002. PMID- 15297555 TI - Evaluation of a new automated, standardized generic nucleic acid extraction method (total nucleic acid isolation kit) used in combination with cytomegalovirus DNA quantification by COBAS AMPLICOR CMV MONITOR. AB - The new generic Total Nucleic Acid Isolation kit improved the detection limit of a cytomegalovirus (CMV) PCR from 400 to 200 copies/ml. Sensitivity and specificity in 30 CMV-positive and 100 negative samples were 100%. Analytical performance was excellent. The kit provides a reliable, standardized, and time saving tool for DNA extraction. PMID- 15297556 TI - Infection of a Japanese patient by genotype 4 hepatitis e virus while traveling in Vietnam. AB - Cases of imported hepatitis E in industrialized countries infected with a genotype 1 hepatitis E virus (HEV) have been identified. We report a 56-year-old Japanese man who acquired infection with a genotype 4 HEV with 98.8% identity to a Vietnamese isolate after ingestion of uncooked shellfish while traveling in Vietnam. PMID- 15297557 TI - Molecular characterization of Trichomonas tenax causing pulmonary infection. PMID- 15297558 TI - Mucoid nitrate-negative Moraxella nonliquefaciens from three patients with chronic lung disease. AB - Mucoid strains of Moraxella nonliquefaciens were recovered from the sputa of three indigenous Australians with chronic lung disease. These atypical strains failed to reduce nitrate, and one strain produced beta-lactamase. While the mucoid phenotype of M. nonliquefaciens has rarely been reported, the mucoid nitrate-negative biovar has never been previously reported. PMID- 15297559 TI - Solitary neurocysticercosis case caused by Asian genotype of Taenia solium confirmed by mitochondrial DNA analysis. AB - A Japanese woman presenting with neurologic symptoms was presumptively diagnosed with neurocysticercosis based on imaging findings. Hooklets in the scolex of the resected lesion were not confirmed through histopathological observation. However, the illness was confirmed by mitochondrial DNA analysis to be a solitary neurocysticercosis case caused by the Asian genotype of Taenia solium. PMID- 15297560 TI - Mycobacterium lentiflavum as an emerging causative agent of cervical lymphadenitis. AB - A lymph node excision was performed on a 45-year-old woman with left cervical swelling. The disorder which developed after the patient had undergone oral surgery for a severe periodontal disease failed to respond to antimicrobial chemotherapy. A mycobacterial strain subsequently identified by high-performance liquid chromatography analysis of cell wall mycolic acids as Mycobacterium lentiflavum grew from the excised specimen. This case and previously published reports highlight the relevance of M. lentiflavum as an emerging causative agent of mycobacterial cervical lymphadenitis. PMID- 15297561 TI - Reemerging threat of epidemic typhus in Algeria. AB - We report a case of epidemic typhus in a patient from the Batna region of Algeria, who presented with generalized febrile exanthema. The clinical diagnosis was confirmed by serological cross-adsorption followed by Western blotting. Our report emphasizes the threat of epidemic typhus in the highlands of Algeria. PMID- 15297562 TI - Intrafamilial spread of the same clarithromycin-resistant Helicobacter pylori infection confirmed by molecular analysis. AB - Clarithromycin-resistant Helicobacter pylori (CRHP) is increasing worldwide, especially in children. We report a family case in which both the mother and child were infected with CRHP. DNA analysis revealed that all of the mother's and daughter's isolates were indistinguishable, suggesting that the same CRHP strain spread between the family members. The spread of CRHP within families may be increasing. PMID- 15297563 TI - Two novel clinical presentations of Burkholderia cepacia infection. AB - We report two cases of multidrug-resistant Burkholderia cepacia (B. cepacia genomovar I) and Burkholderia multivorans causing multiple liver abscesses in a patient with bronchial asthma (case 1) and peritonitis in a patient with cirrhosis and hepatitis C virus disease (case 2), respectively. Both patients were treated successfully. PMID- 15297564 TI - Are extracorporeal membrane oxygenation circuits that are primed with plasmalyte and stored a likely source of infection? PMID- 15297565 TI - Increased sensitivity of a latex agglutination method for serotyping group B streptococcus. PMID- 15297566 TI - Genotyping primers for fully automated multilocus variable-number tandem repeat analysis of Escherichia coli O157:H7. PMID- 15297567 TI - Etiology of acute diarrhea in adults in southwestern Nigeria. PMID- 15297568 TI - Contraction augments L-type Ca2+ currents in adherent guinea-pig cardiomyocytes. AB - As integrins are thought to function as mechanoreceptors, we studied whether they could mediate mechanical modulation of the L-type Ca2+ channel current (ICa) in guinea-pig cardiac ventricular myocytes (CVMs). CVMs were voltage clamped with 280 ms pulses from -45 to 0 mV at 0.5 Hz (1.8 mM [Ca2+]o, 22 degrees C). Five minutes after whole-cell access (designated as 0 min) peak ICa was determined from a current-voltage (I-V) curve. Additional recordings were made after 5, 10 and 15 min. At control, ICa was not stable, but ran down during these periods. This run-down of ICa was attenuated by soluble fibronectin (FN) and was changed to an enhancement of ICa when CVMs were attached to FN-coated coverslips. Soluble peptide containing the integrin binding sequence of FN, Arg-Gly-Asp (RGD motif), did not modulate ICa; however, ICa increased in stimulated CVMs attached to RGD peptide-coated coverslips. The effect was not specific to integrins, because attachment to poly-D-lysine-coated coverslips also augmented ICa in stimulated CVMs. Augmentation of ICa by immobilized FN required rhythmical contraction of attached CVMs, because it was attenuated without electrical stimulation and after cell dialysis with the calcium chelator BAPTA. Furthermore, contraction-induced augmentation of ICa in FN-attached CVMs was sensitive to inhibition of protein kinase C (PKC; by Ro-31-8220), inhibition of tyrosine kinase activity (herbimycin A) and cytoskeletal depolymerization (cytochalasin D or colchicine). We attribute augmentation of ICa to the activation of signalling cascades by shear forces that are generated when CVMs contract against attachment; in vivo similar signals may occur when CVMs contract against attachment of integrins to the extracellular matrix. PMID- 15297569 TI - Recapture after exocytosis causes differential retention of protein in granules of bovine chromaffin cells. AB - After exocytosis, chromaffin granules release essentially all their catecholamines in small fractions of a second, but it is unknown how fast they release stored peptides and proteins. Here we compare the exocytic release of fluorescently labelled neuropeptide Y (NPY) and tissue plasminogen activator from single granules. Exocytosis was tracked by measuring the membrane capacitance, and single granules in live cells were imaged by evanescent field microscopy. Neuropeptide Y left most granules in small fractions of a second, while tissue plasminogen activator remained in open granules for minutes. Taking advantage of the dependence on pH of the fluorescence of green fluorescent protein, we used rhythmic external acidification to determine whether and when granules re-sealed. One-third of them re-sealed within 100 s and retained significant levels of tissue plasminogen activator. Re-sealing accounts for only a fraction of the endocytosis monitored in capacitance measurements. When external [Ca2+] was raised, even neuropeptide Y remained in open granules until they re-sealed. It is concluded that a significant fraction of chromaffin granules re-seal after exocytosis, and retain those proteins that leave granules slowly. We suggest that granules vary the stoichiometry of release by varying both granule re-sealing and the association of proteins with the granule matrix. PMID- 15297570 TI - External Ca(2+)-dependent excitation--contraction coupling in a population of ageing mouse skeletal muscle fibres. AB - In the present work, we investigate whether changes in excitation-contraction (EC) coupling mode occur in skeletal muscles from ageing mammals by examining the dependence of EC coupling on extracellular Ca(2+). Single intact muscle fibres from flexor digitorum brevis muscles from young (2-6 months) and old (23-30 months) mice were subjected to tetanic contractile protocols in the presence and absence of external Ca(2+). Contractile experiments in the absence of external Ca(2+) show that about half of muscle fibres from old mice are dependent upon external Ca(2+) for maintaining maximal tetanic force output, while young fibres are not. Decreased force in the absence of external Ca(2+) was not due to changes in charge movement as revealed by whole-cell patch-clamp experiments. Ca(2+) transients, measured by fluo-4 fluorescence, declined in voltage-clamped fibres from old mice in the absence of external Ca(2+). Similarly, Ca(2+) transients declined in parallel with tetanic contractile force in single intact fibres. Examination of inward Ca(2+) current and of mRNA and protein assays suggest that these changes in EC coupling mode are not due to shifts in dihydropyridine receptor (DHPR) and/or ryanodine receptor (RyR) isoforms. These results indicate that a change in EC coupling mode occurs in a population of fibres in ageing skeletal muscle, and is responsible for the age-related dependence on extracellular Ca(2+). PMID- 15297571 TI - Tetanic depression is overcome by tonic adenosine A(2A) receptor facilitation of L-type Ca(2+) influx into rat motor nerve terminals. AB - Motor nerve terminals possess multiple voltage-sensitive calcium channels operating acetylcholine (ACh) release. In this study, we investigated whether facilitation of neuromuscular transmission by adenosine generated during neuronal firing was operated by Ca(2+) influx via 'prevalent' P-type or via the recruitment of 'silent' L-type channels. The release of [(3)H]ACh from rat phrenic nerve endings decreased upon increasing the stimulation frequency of the trains (750 pulses) from 5 Hz (83 +/- 4 x 10(3) disintegrations per minute per gram (d.p.m. g(-1)); n = 11) to 50 Hz (30 +/- 3 x 10(3) d.p.m. g(-1); n = 5). The P-type Ca(2+) channel blocker, omega-agatoxin IVA (100 nm) reduced (by 40 +/- 10%; n = 6) the release of [(3)H]ACh evoked by 50-Hz trains, while nifedipine (1 microM, an L-type blocker) was inactive. Tetanic depression was overcome (88 +/- 6 x 10(3) d.p.m. g(-1); n = 12) by stimulating the phrenic nerve with 50-Hz bursts (five bursts of 150 pulses, 20 s interburst interval). In these conditions, omega-agatoxin IVA (100 nM) failed to affect transmitter release, but nifedipine (1 microM) decreased [(3)H]ACh release by 21 +/- 7% (n = 4). Inactivation of endogenous adenosine with adenosine deaminase (ADA, 0.5 U ml(-1)) reduced (by 54 +/- 8%, n = 5) the release of [(3)H]ACh evoked with 50-Hz bursts. This effect was opposite to the excitatory actions of adenosine (0.5 mm), S-(p nitrobenzyl)-6-thioinosine (5 microM, an adenosine uptake blocker) and CGS 21680C (3 nM, a selective A(2A) receptor agonist); as the A(1) receptor agonist R-N(6) phenylisopropyl adenosine (R-PIA, 300 nM) failed to affect the release of [(3)H]ACh, the results indicate that adenosine generated during 50-Hz bursts exerts an A(2A)-receptor-mediated tonus. The effects of ADA (0.5 U ml(-1)) and CGS 21680C (3 nm) were prevented by nifedipine (1 microM). Blocking tonic A(2A) receptor activation, with ADA (0.5 U ml(-1)) or 3,7-dimethyl-1-propargyl xanthine (10 microM, an A(2A) antagonist), recovered omega-agatoxin IVA (100 nM) inhibition and caused the loss of function of nifedipine (1 microM). Data indicate that, in addition to the predominant P-type Ca(2+) current triggering ACh release during brief tetanic trains, motoneurones possess L-type channels that may be recruited to facilitate transmitter release during high-frequency bursts. The fine-tuning control of Ca(2+) influx through P- or L-type channels is likely to be mediated by endogenous adenosine. Therefore, tonic activation of presynaptic A(2A) receptors operating Ca(2+) influx via L-type channels may contribute to overcome tetanic depression during neuronal firing. PMID- 15297572 TI - Albumin transcytosis across the epithelium of the lactating mouse mammary gland. AB - Murine milk contains 18 mg ml(-1) serum albumin, a concentration equal to that in the serum of the lactating mouse. We examined cellular transport using in vivo methods in the mouse. At steady state the specific activity of (125)I-albumin injected into the blood stream was equal in plasma and whey, confirming that milk albumin is extra-mammary in origin. Fluorescent albumin crossed the gland from basolateral surface to lumen via cytoplasmic vesicles, but was not transported in the apical to basal direction. Albumin was segregated from transferrin at the basal surface of the epithelial cells and did not colocalize with either caveolin 1 or -2. Vesicular transport was not disrupted by filipin providing additional evidence that, unlike the vascular endothelium, caveoli are not involved. Cytoplasmic albumin was localized to vesicles containing IgA and transport was disrupted by agents that interfere with clathrin-mediated endocytosis. Together, these findings provide evidence that albumin is transported across the mammary epithelium by the same pathway as immunoglobulin. The possibility that the massive transfer of albumin into mouse milk is mediated by fluid phase transport is considered. PMID- 15297573 TI - Electrophysiological mapping of the nociceptive inputs to the substantia gelatinosa in rat horizontal spinal cord slices. AB - To study the functional projection patterns of the primary afferents in the spinal cord, the postsynaptic responses of substantia gelatinosa (SG) neurones evoked by L5 dorsal root stimulation (DRS) were examined from the neurones located at L2 to S1 in horizontal slices of the adult rat spinal cord using a blind whole-cell patch-clamp technique. In the voltage-clamp mode, the L5 DRS evoked the Adelta- and C-afferent-mediated excitatory postsynaptic currents (EPSCs) in more than 70% of the neurones tested at the L5 level. Both Adelta- and C-afferent EPSCs were also recorded in more than 50% of the neurones at L4. At L3 and L6, the number of neurones receiving the C-afferent EPSCs (> 40%) was significantly greater than that of Adelta-afferent EPSCs (< 20%). On the other hand, the Adelta- and C-afferent-mediated inhibitory postsynaptic currents (IPSCs) elicited by L5 DRS were almost equally observed from L2 to S1. In the current-clamp mode, L5 DRS evoked Adelta- and C-afferent-mediated EPSPs, some of which initiated action potentials (APs). Most of the Adelta-afferent-mediated APs were limited at the L5 level, while C-afferent-mediated APs were observed at L5 and L4. As the L2 DRS-evoked APs in the L2 SG neurones were suppressed by L5 DRS, the widespread distribution of the inhibitory inputs was considered to be functional. These findings suggest that the excitatory projection of the C afferents to the SG neurones was thus spread more rostrocaudally than that of the Adelta afferents, thereby contributing to more diffuse pain transmission. In addition, the widespread distribution of the inhibitory inputs may thus play a role as a lateral inhibitory network and thereby prevent the expansion of the excitatory inputs of noxious stimuli. PMID- 15297574 TI - Mechanisms of hyperpolarization in regenerated mature motor axons in cat. AB - We found persistent abnormalities in the recovery of membrane excitability in long-term regenerated motor nerve fibres in the cat as indicated in the companion paper. These abnormalities could partly be explained by membrane hyperpolarization. To further investigate this possibility, we compared the changes in excitability in control nerves and long-term regenerated cat nerves (3 5 years after tibial nerve crush) during manoeuvres known to alter axonal membrane Na(+)-K(+) pump function: polarization, cooling to 20 degrees C, reperfusion after 10 min ischaemia, and up to 60 s of repetitive stimulation at 200 Hz. The abnormalities in excitability of regenerated nerves were reduced by depolarization and cooling and increased by hyperpolarization and during postischaemia. Moreover, the time course of recovery of excitability from repetitive stimulation and ischaemia was prolonged in regenerated nerves. Our data are consistent with an increased demand for electrogenic Na(+)-K(+) pumping in regenerated nerves leading to membrane hyperpolarization. Such persistent hyperpolarization may influence the ability of the axon to compensate for changes in membrane potential following normal repetitive activity. PMID- 15297575 TI - Persistent abnormalities of membrane excitability in regenerated mature motor axons in cat. AB - The purpose of our study was to assess by threshold tracking internodal and nodal membrane excitability during the maturation process after tibial nerve crush in cat. Various excitability indices (EI) were computed non-invasively by comparing the threshold of a submaximal compound motor potential at different stimulation durations (strength-duration relationship), after a conditioning nerve impulse (recovery of excitability), or during the application of a polarizing current (threshold electrotonus). Four months after the lesion, regenerated nerves showed a higher rheobase, shorter chronaxie, shorter refractory period and higher than normal threshold variations during threshold electrotonus (TE). A partial recovery was observed during the first 2 years of maturation. The recovery to depolarizing TE seemed complete but all other EI remained abnormal even after 5 years of regeneration, the most pronounced being the 157 +/- 8% (mean +/- S.E.M.) increase in threshold during hyperpolarizing TE compared with 94 +/- 4% in controls. These EI abnormalities are consistent with increased input impedance. Nevertheless, the time course of maturation and incomplete recovery of EI could only be partially explained by changes in fibre morphology. The highly abnormal response to hyperpolarizing but not to depolarizing TE suggests that voltage dependent membrane function also remained abnormal, possibly due to membrane hyperpolarization. PMID- 15297576 TI - Presynaptic modulation by neuromedin U of sensory synaptic transmission in rat spinal dorsal horn neurones. AB - Neuromedin U (NMU) is a brain-gut peptide first isolated from the spinal cord. Recent studies on NMU and its receptors have suggested a role of NMU in sensory transmission. Here we report on the localization of NMU in sensory neurones, and the actions of NMU in the substantia gelatinosa (SG) and the deep layer of the dorsal horn (laminae III-V) in adult rat spinal cord slices using the patch-clamp technique. An immunohistochemical study revealed that NMU peptide was present in most of the dorsal root ganglion neurones. In the spinal cord, NMU-immunoreactive neurones were located in the deep layer (laminae III-V), but not in the SG. However, NMU-positive axon terminals were observed in the SG as well as the deep layer. Bath-applied NMU (10 microm) increased the frequency, but not amplitude, of miniature excitatory postsynaptic currents (mEPSCs) in the SG and deep layer neurones by 146 +/- 14% (P < 0.01, n = 17) and 174 +/- 21% (P < 0.01, n = 6), respectively, without inducing any postsynaptic membrane currents recorded in tetrodotoxin. On the other hand, NMU did not affect miniature inhibitory postsynaptic currents recorded in tetrodotoxin. These findings, taken together, suggest that NMU acts on the presynaptic terminals of the primary afferent fibres working as an autocrine/paracrine neuromodulator to increase mEPSC frequency of the SG and deep layer neurones. This may account for the spinal mechanisms of the NMU-induced hyperalgesia reported previously. PMID- 15297577 TI - Differential effect of bicycling exercise intensity on activity and phosphorylation of atypical protein kinase C and extracellular signal-regulated protein kinase in skeletal muscle. AB - Atypical protein kinase C (aPKC) and extracellular signal-regulated kinase (ERK) are emerging as important signalling molecules in the regulation of metabolism and gene expression in skeletal muscle. Exercise is known to increase activity of aPKC and ERK in skeletal muscle but the effect of exercise intensity hereon has not been studied. Furthermore, the relationship between activity and phosphorylation of the two enzymes during exercise is unknown. Nine healthy young men exercised for 30 min on a bicycle ergometer on two occasions. One occasion consisted of three consecutive 10 min bouts of 35, 60 and 85% of peak pulmonary oxygen uptake V(O(2 peak)) and the second of one 30 min bout at 35% of V(O(2 peak)). Both trials also included 30 min recovery. Muscle biopsies were obtained from the vastus lateralis muscle before and after each exercise bout. Exercise increased muscle aPKC activity at 35% V(O(2 peak)), whereupon no further increase was observed at higher exercise intensities. Activation of aPKC was not accompanied by increased phosphorylation of aPKC Thr(410/403). ERK1/2 activity increased in a similar pattern to aPKC, reaching maximal activity at 35% V(O(2 peak)), whereas ERK1 Thr(202)/Tyr(204) and ERK2 Thr(183)/Tyr(185) phosphorylation increased with increasing exercise intensity. Thus, aPKC and ERK1/2 activity in muscle during exercise did not correspond to phosphorylation of sites on aPKC or ERK1/2, respectively, which are considered important for their activation. It is concluded that assessment of aPKC and ERK1/2 activity in muscle using phosphospecific antibodies did not reflect direct activity measurements on immunoprecipitated enzyme in vitro. Thus, estimation of enzyme activity during exercise by use of phosphospecific antibodies should not be performed uncritically. In addition, increase in muscle activity of aPKC or ERK1/2 during exercise is not closely related to energy demands of the muscle but may serve other regulatory or permissive functions in muscle. PMID- 15297578 TI - Spontaneous, synchronous electrical activity in neonatal mouse cortical neurones. AB - Spontaneous [Ca2+]i transients were measured in the mouse neocortex from embryonic day 16 (E16) to postnatal day 6 (P6). On the day of birth (P0), cortical neurones generated widespread, highly synchronous [Ca2+]i transients over large areas. On average, 52% of neurones participated in these transients, and in 20% of slices, an average of 80% participated. These transients were blocked by TTX and nifedipine, indicating that they resulted from Ca2+ influx during electrical activity, and occurred at a mean frequency of 0.91 min(-1). The occurrence of this activity was highly centred at P0: at E16 and P2 an average of only 15% and 24% of neurones, respectively, participated in synchronous transients, and they occurred at much lower frequencies at both E16 and P2 than at P0. The overall frequency of [Ca2+]i transients in individual cells did not change between E16 and P2, just the degree of their synchronicity. The onset of this spontaneous, synchronous activity correlated with a large increase in Na+ current density that occurred just before P0, and its cessation with a large decrease in resting resistance that occurred just after P2. This widespread, synchronous activity may serve a variety of functions in the neonatal nervous system. PMID- 15297579 TI - Inhibitory regulation of constitutive transient receptor potential-like cation channels in rabbit ear artery myocytes. AB - In the present study we have investigated an inhibitory pathway regulating a constitutively active Ca(2+)-permeable non-selective cation conductance (I(cat)) in rabbit ear artery smooth muscle cells. Constitutive single channel activity of I(cat) was recorded in cell-attached and inside-out patches with similar unitary conductance values. In inside-out patches with relatively high constitutive activity the G-protein activator GTPgammaS inhibited channel activity which was reversed by the protein kinase C (PKC) inhibitor chelerythrine indicating a G protein pathway inhibits channel activity via PKC. Spontaneous channel activity was also suppressed by the G-protein inhibitor GDPbetaS suggesting a G-protein is also involved in initiation of constitutive channel activity. Bath application of antibodies to G(alphaq)/G(alpha11) enhanced channel activity whereas anti G(alpha1-3)/G(alphao) antibodies decreased basal channel activity which suggests that G(alphaq)/G(alpha11) and G(alphaiota)/G(alphao) proteins initiate, respectively, the inhibitory and excitatory cascades. The phospholipase C (PLC) inhibitor U73122 increased spontaneous activity which implies a role for PLC in the inhibitory pathway. Bath application of the diacylycerol (DAG) analogue 1 oeoyl-2-acetyl-sn-glycerol (OAG) decreased the probability of channel opening (NP(o)) and this was reversed by chelerythrine. Application of the PKC activator phorbol 12, 13-dibutyrate (PDBu) and chelerythrine, respectively, decreased and increased NP(o). These data indicate that spontaneously active cation channels are inhibited by a tonic inhibitory pathway involving G(alphaq)/G(alpha11) mediated stimulation of PLC to generate DAG which activates PKC to inhibit channel opening. There were some patches with relatively low NP(o) and it was evident that the inhibitory pathway was particularly marked in these cases. Moreover in the latter patches GTPgammaS and OAG caused marked increases in NP(o). Together with inhibitory effects of GDPbetaS and anti-G(alpha1 3)/G(alphao) antibodies the results suggest that there is constitutive G(alphai)/G(alphao) protein activity leading to channel opening via a DAG mediated but PKC-independent mechanism. Finally, with whole-cell recording it is shown that noradrenaline increases I(cat) and the noradrenaline-evoked response is markedly potentiated by PKC inhibition. This latter observation shows that PKC also limits agonist-evoked I(cat) in these arterial myocytes. PMID- 15297580 TI - Stress and glucocorticoid inhibit apical GLUT2-trafficking and intestinal glucose absorption in rat small intestine. AB - We have proposed a new model of rat intestinal sugar absorption in which high glucose concentrations promote rapid insertion of GLUT2 into the apical membrane, so that absorptive capacity is precisely regulated to match dietary intake. Construction and building work during expansion and refurbishment of our department permitted opportunistic experiments on the effects of building-induced stress on the GLUT2 component of absorption. In fed rats perfused with 75 mM glucose in vivo, stress rapidly inhibited glucose absorption 36.4 +/- 3.0% compared with control rats. Selective inhibition of the GLUT2 component with phloretin demonstrated that stress inhibited the GLUT2 component by 42.8 +/- 3.8%, which correlated with a corresponding diminution in apical GLUT2 levels: the SGLT1 component and its level were unaltered by stress. Effects of stress were reversed by the administration in drinking water of metyrapone, which inhibits 11-beta-hydroxylase. Injection of dexamethasone into control rats 60 min before perfusion resulted in absorption and transporter properties indistinguishable from stressed rats. Our data are consistent with the view that stress activates the hypothalamus-pituitary-adrenal (HPA) axis, causing release of glucocorticoid. The ensuing inhibition of GLUT2 trafficking and absorption seems necessary to prevent enhanced intestinal delivery of glucose to the circulation from antagonizing the essential stress response of glucorticoid in mobilizing peripheral energy stores for emergency purposes. PMID- 15297581 TI - Polyvinyl alcohol particles versus tris-acryl gelatin microspheres for uterine artery embolization for leiomyomas. PMID- 15297582 TI - Polyvinyl alcohol particles and tris-acryl gelatin microspheres for uterine artery embolization for leiomyomas: results of a randomized comparative study. AB - PURPOSE: To determine if the type of embolic material used for uterine artery embolization (UAE) for leiomyomas has an impact on short-term recovery or the effectiveness of embolization. MATERIALS AND METHODS: One hundred patients were randomly assigned to undergo UAE with polyvinyl alcohol (PVA) particles or tris acryl gelatin microspheres. Short-term, in-hospital medication use and pain levels were recorded. After discharge, symptom severity, temperature, and medications used were recorded daily for 1 week and symptom levels were measured for weeks 2-4. Three months after embolization, contrast material-enhanced magnetic resonance imaging examinations were evaluated blindly to determine the extent of leiomyoma infarction. Symptom and quality of life (QOL) status was determined with use of questionnaires. Analysis was completed with use of chi(2) analysis, Fisher exact tests, Student t tests, and analysis of variance as appropriate. Regression analysis was used to analyze the impact on outcome of baseline factors (other than type of embolic agent). RESULTS: No significant differences were noted at baseline between the two treatment groups. On average, there were significantly higher volumes of tris-acryl microspheres used (9.0 mL vs 3.0 mL; P =.0001), whereas microcatheter occlusion was more common with PVA (28% vs 4%, P =.001). There were no differences in pain severity, other postprocedural symptoms, or medication use between the two treatment groups. There were also no differences in the frequency of incompletely infarcted leiomyomas, degree of improvement in symptom score, patient satisfaction, or QOL. CONCLUSION: No substantive differences were detected between outcomes of embolization with PVA particles or tris-acryl gelatin microspheres. PMID- 15297583 TI - Recovery after uterine artery embolization for leiomyomas: a detailed analysis of its duration and severity. AB - PURPOSE: To determine the duration and severity of recovery after uterine artery embolization (UAE) for leiomyomas. MATERIALS AND METHODS: As part of a study comparing different embolic materials used for UAE, detailed data on the severity of postprocedural recovery were gathered in 99 patients. These data included patient-controlled analgesia records, visual analogue scale (VAS) pain scores of daily peak pain levels for 7 days, medication use in the first week, and severity of constitutional symptoms experienced over the course of the first month after the procedure. The VAS scale assesses acute pain severity on a 10-cm linear scale and yields a continuous measure from 1 to 10. The constitutional symptoms were scored based on a questionnaire. The data were analyzed with use of summary statistics, and linear regression analysis was used to determine the impact of various baseline factors on the severity of recovery. RESULTS: The mean peak VAS score for the first 24 hours after UAE was 3.03 (SD, 0.26) and the mean maximum score in the first week was 4.89 (SD, 0.26). Only 11 patients had an in-hospital VAS score greater than 7, and 19 had a VAS score of greater than 7 on any of the first 7 days after discharge. The mean number of oral narcotic tablets used per patient was 10.8 in the first week. Although 33 patients had a temperature higher than normal sometime in the first postprocedural week, high temperature (>38.5 degrees C) occurred in only two patients. There were no differences detected in the measured parameters based on the type of embolic material used. CONCLUSION: Despite the reputation of UAE to the contrary, when current techniques are used, recovery after UAE for fibroids is relatively mild, with few instances of severe pain, high fever, or severe constitutional symptoms. PMID- 15297584 TI - Power injection of contrast media via peripherally inserted central catheters for CT. AB - PURPOSE: To evaluate patient safety, catheter rupture rates, and computed tomography (CT) image quality when using peripherally inserted central catheters (PICCs) in vivo for the power injection of CT contrast media at standard injection rates. MATERIALS AND METHODS: A prospective study to evaluate the safety and effectiveness of power injection of contrast media via indwelling PICCs was performed. Single-lumen and double-lumen polyurethane PICCs (5 F) were injected in vivo with contrast media for clinical CT examinations at injection rates ranging from 1 mL/sec to 4 mL/sec. Data collected included PICC rupture rate, patient complications, injection rate, peak injection pressure, PICC length, PICC age, and quality of contrast enhancement on the CT images. RESULTS: One hundred ten power injections of PICCs for CT examinations were performed. There were 12 injections of single-lumen PICCs and 98 injections of double-lumen PICCs. The most common injection rate was 2 mL/sec, accounting for 89 of the 110 injections (81%). Two PICCs ruptured during power injection, both as a result of operator error. One of the PICCs that ruptured was clamped at the time of injection and the other one was kinked at its venous entry site. One additional PICC showed evidence of dysfunction; it ballooned without actually rupturing. No significant patient complications occurred. Contrast enhancement of the CT images was subjectively rated as average or above average in 95% of cases. CONCLUSIONS: Contrast media can be power-injected via PICCs for routine CT examinations at a rate of 2 mL/sec, yielding satisfactory image quality without exposing patients to significant additional risk. Power injection rates greater than 2 mL/sec, as are typically used in CT angiography applications, were not fully evaluated by this study. PMID- 15297585 TI - Transcatheter arterial chemoembolization for hepatocellular carcinoma fed by the reconstructed inferior phrenic artery: anatomical and technical analysis. AB - PURPOSE: To evaluate reconstructed patterns of occluded inferior phrenic artery (IPA) and determine the technical success rate and complications of transcatheter arterial chemoembolization (TACE) for hepatocellular carcinoma (HCC) fed by the occluded IPA through the anastomosing branch. MATERIALS AND METHODS: In 19 patients, 24 IPAs, including two that had been previously embolized, were demonstrated through collateral pathways. The incidence of each collateral circulation was evaluated. Thirteen IPAs in 12 patients fed the tumor and TACE was attempted. TACE was performed only if the catheter could be advanced into the anastomosing branch so that the nontarget branches were avoided. RESULTS: A reconstructed unilateral IPA was observed in 14 patients (11 right IPAs and three left IPAs) and two reconstructed IPAs were observed in five. The IPA was demonstrated through the dorsal pancreatic artery (n = 13), inferior or middle adrenal artery (n = 7), left gastric artery (n = 2), contralateral IPA (n = 2), lumbar artery (n = 1), and small branch derived from the celiac trunk (n = 1). Five IPAs (21%) were demonstrated through more than two separate arteries, including two demonstrated through both dorsal pancreatic arteries arising from the celiac and superior mesenteric artery. The IPA opacified through the lumbar artery had been previously embolized. TACE of the reconstructed IPA was possible in 10 of 13 IPAs (77%). Complications related to the procedure were a small amount of pleural effusion (n = 4) and basal atelectasis (n = 2). CONCLUSION: The IPA is reconstructed mainly through the retroperitoneal anastomosing branch in the upper abdomen. TACE of the reconstructed IPA can be performed with a high success rate without major complications. PMID- 15297586 TI - Hepatic arterial port systems for treatment of liver metastases: factors affecting patency and adverse events. AB - PURPOSE: To assess the outcome of interventional hepatic arterial port placement in a prospective phase II trial. MATERIALS AND METHODS: One-hundred five consecutive patients were included in this study. Primary endpoint was port patency; secondary endpoints were complications, toxicity, response, and progression free and overall survival. Seventy-eight patients presented with liver metastasis only, 6 patients had additional minor extrahepatic disease, and 21 patients had no evidence of disease after liver resection, laser-induced thermotherapy, or computed tomography (CT)-guided interstitial brachytherapy of liver metastasis. Exclusive access route was the femoral artery. Subgroup analysis compared either 4-F catheters (n = 58) to 2.2-F (n = 33) and 2.7-F (n = 20) microcatheters or different strategies in anatomic variants of the celiac branch: neglect (n = 10) or embolization of minor hepatic feeders (n = 11), splenic arterial port (n = 8), double port (n = 7). RESULTS: Technical success was 99%. Assisted port patency after 6 months was 93%. Complications demanding port revisions were significantly lower in patients receiving 4-F versus 2.2-F and 2.7-F systems (P <.001), with disconnection as the major problem with use of microcatheters. Hepatic artery thrombosis occurred in 10 patients (9%), with successful lysis in two patients. With use of 4-F and 2.2-F catheters, there was no difference with respect to catheter occlusion or hepatic thrombosis. No differences were noted in complications or outcome applying four different strategies in celiac branch variants. In a subgroup of patients receiving folinic acid/5-fluorouracil (170 mg/600 mg; 10% dose escalation per cycle) for 5 days every 4 weeks only 15% experienced Grade 3 toxicity. Patients with liver metastasis and salvage therapy demonstrated progression-free survival of 63% after 6 months and a median survival of 16 months. CONCLUSION: Interventional placement of hepatic arterial port systems may overcome frequent hepatic arterial chemotherapy failures as encountered in all published major trials on hepatic arterial infusion. PMID- 15297587 TI - Hepatic arterial infusion chemotherapy with use of an implanted port system in patients with advanced hepatocellular carcinoma: prognostic factors. AB - PURPOSE: This study was retrospectively undertaken to identify prognostic factors in patients with advanced hepatocellular carcinoma (HCC) treated by hepatic arterial infusion chemotherapy with a percutaneously implantable port system inserted. MATERIALS AND METHODS: Eighty-eight patients underwent arterial infusion chemotherapy for portal venous invasion (n = 39), severe liver dysfunction (n = 6), or tumor regrowth after chemoembolization, percutaneous ethanol injection therapy, and surgery (n = 77). Twenty-five variables representing patients' characteristics, previous treatments, tumor characteristics, liver profiles, various staging systems, and therapeutic effect were analyzed with univariate and multivariate analyses. RESULTS: The 1- and 3 year survival rates were 55% and 24%, respectively, with a mean survival period of 19.5 months +/- 1.9 in all patients. Cancer of the Liver Italian Program (CLIP) score, Okuda stage, therapeutic effect, tumor extension, alkaline phosphatase and aspartate aminotransferase levels, ascites, and portal venous invasion were identified as significant prognostic factors by univariate analysis. Multivariate analysis identified CLIP score, Okuda stage, and therapeutic effect as significant independent prognostic factors. CONCLUSION: Although our results should be confirmed in future prospective studies, the prognostic factors identified in the present study should prove helpful in classifying patients with advanced HCC who are treated by arterial infusion chemotherapy and should serve as useful guidelines on arterial infusion chemotherapy in clinical practice. PMID- 15297588 TI - Radiopacity of current endovascular stents: evaluation in a multiple reader phantom study. AB - PURPOSE: To evaluate the radiopacity of endovascular stents based on the fluoroscopy mode in a phantom of the human pelvis. MATERIALS AND METHODS: The following stents were included in this study: Medtronic AVE Bridge, Medtronic AVE Bridge X, Cordis Covered Nitinol (Covent), Guidant Dynalink, Luminexx, Guidant Megalink, Memotherm Flexx, Palmaz Medium, Palmaz-Schatz Long-Medium, Palmaz Corinthian PQ394Q and PQ294Q, SelfX, SMART without markers, SMART with radiopaque markers, Easy Wallstent. To evaluate radiopacity, images of the stents placed in four different positions (lumbosacral junction left and right, iliosacral joint left and right) of a pelvic phantom were taken at the following modes: spotfilm, continuous fluoroscopy, 15 pulses per second, 7.5 pulses per second, and 3 pulses per second. Images were presented at random to four independent readers and radiopacity scores were assessed: 0 = not visible, 1 = poor visibility, 2 = average visibility, 3 = good visibility, and 4 = very good visibility. RESULTS: The Covent stent had the highest overall radiopacity score (3.25), followed by the Luminexx (3.04) and the Medtronic AVE Bridge X (2.74) stents. At the spotfilm mode, the best visible stents were the Medtronic AVE Bridge X, the Covent and the Easy Wallstent stents and at the continuous fluoroscopy mode, the Covent, the Luminexx, and the Medtronic AVE Bridge X stents. Decreasing the fluoroscopy mode went hand in hand with a reduction of the radiopacity scores of all stents. At the standard fluoroscopy mode of 7.5 pulses per second, the Covent stent was seen well or very well in 96.9%, followed by the Luminexx (76.9%), and the Medtronic AVE Bridge X (41.25%) stents. CONCLUSIONS: Stent radiopacity directly depends on the fluoroscopy mode; if the pulse frequency decreased, detecting the stents became more difficult. Stent mass correlates with stent radiopacity (e.g., Cordis Covered Nitinol, Bridge X). Radiopaque markers may improve stent radiopacity dramatically (e.g., Luminexx vs Memotherm Flexx). PMID- 15297589 TI - Endovascular repair of spontaneous or traumatic iliac vein rupture. AB - Spontaneous rupture of the iliac vein and rupture resulting from blunt trauma are both very unusual. Herein one case of each are reported and were managed by emergent endovascular repair with use of covered stents. Favorable outcomes were achieved in both cases. PMID- 15297590 TI - Pancreatitis caused by rheolytic thrombolysis: an unexpected complication. AB - Two patients developed acute pancreatitis after mechanical thrombolysis with use of the AngioJet system. Patient 1 had undergone a remote complex revascularization of the lower extremities and presented with acute ischemia after thrombosis of his composite distal bypass. Patient 2 presented with superior vena cava (SVC) syndrome and had thrombosis of the SVC and innominate veins. Despite dissimilar presentations, both patients had renal insufficiency, were treated with mechanical and chemical thrombolysis, and had extensive thrombus burden. The pathophysiology of acute pancreatitis in this setting is believed to be secondary to massive hemolysis in the presence of chronic renal insufficiency. This phenomenon should be considered in patients whom develop abdominal pain after mechanical thrombolysis. PMID- 15297591 TI - Endovascular repair of an aortopulmonary artery fistula with use of controlled release coils. AB - Aortopulmonary artery fistula is traditionally treated surgically. The present case report describes endovascular repair of an aortopulmonary artery fistula in a patient in whom two thoracotomies had been performed. The fistula occurred at the site of a pseudoaneurysm from the proximal anastomosis of a graft placed to treat a type B aortic dissection. Two controlled-release endovascular coils were positioned across the fistula, resulting in immediate closure. The fistula remains closed with resolution of the pseudoaneurysm after more than 3 years of follow-up. PMID- 15297592 TI - Cutaneous thermal injury after endovenous laser ablation of the great saphenous vein. AB - Herein a case of cutaneous thermal injury in the leg of a patient who underwent endovenous laser (EVL) ablation of an incompetent great saphenous vein (GSV) is reported. Follow-up ultrasonography (US) of the site of skin burn showed that the burn was directly over thrombosed superficial tributaries originating from the GSV, but medial to the treated GSV. At the level of skin burn, the thrombosed GSV was 22 mm deep, but the tributaries were 1 mm deep. In addition, US showed echogenic fat surrounding and conforming to the superficial thrombosed tributaries. Based on the clinical scenario and follow-up US findings, it was concluded that the cutaneous thermal injury resulted from heated blood traveling from the 22-mm-deep GSV to the superficial tributaries directly beneath the site of skin burn. PMID- 15297593 TI - Percutaneous pediculoplasty in osteoporotic compression fractures. AB - Percutaneous pediculoplasty is a vertebroplasty-complementary technique that can be carried out with one needle for each single approach. This report describes five cases of osteoporotic vertebral and pedicular compression fractures that were treated with percutaneous vertebroplasty and bilateral pediculoplasty with use of polymethylmethacrylate and high-quality fluoroscopic guidance. All patients reported complete pain relief. This is a safe, fast, and effective treatment for osteoporotic compression fractures with pedicle compromise. PMID- 15297594 TI - Portosplenic blood flow separation in a patient with portosystemic encephalopathy and a spontaneous splenorenal shunt. AB - A patient with portosystemic encephalopathy, hyperammonemia, and a spontaneous splenorenal shunt was admitted to the authors' institution after a failed attempt at transvenous retrograde shunt obliteration. As an alternative approach, the authors separated splenic and portal flows by embolizing only the proximal splenic vein while leaving the shunt intact. Thus, the splenic flow could escape into the systemic circulation and an extreme increase in portal pressure was avoided. The procedure could provide rapid decreases in blood ammonia levels and a fast resolution of symptoms, but repeated interventions were required. PMID- 15297595 TI - Microcatheter placement through a side hole created in a 5-F catheter into proximal subclavian arterial branches causing hemoptysis. AB - Five patients with moderate to massive hemoptysis who had a bronchial artery of anomalous origin or a nonbronchial systemic artery originating from the proximal subclavian artery underwent microcatheter placement through a created side hole of a 5-F catheter. All patients had pulmonary tuberculosis and had undergone bronchial artery embolization for hemoptysis. The side holes were made in the lesser (n = 2) or greater curvature sides (n = 3) of 5-F nonbraided Headhunter catheters. A microcatheter was passed through the side hole of the 5-F catheter into the target artery for embolization. Polyvinyl alcohol particles were used as the embolic material. The technical success rate was 100%, and immediate control of hemoptysis was achieved in all patients without complication. PMID- 15297596 TI - Emergent transcatheter arterial embolization of ruptured inferior phrenic artery aneurysm with N-butyl cyanoacrylate. PMID- 15297597 TI - Re: Fatal pulmonary complications after arterial embolization with 40-120-microm tris-acryl gelatin microspheres. PMID- 15297598 TI - Microsatellite variation and evolutionary history of PCDHX/Y gene pair within the Xq21.3/Yp11.2 hominid-specific homology block. AB - To better understand the evolutionary dynamics of repetitive sequences in human sex chromosomes, we have analyzed seven new X/Y homologous microsatellites located within PCDHX/Y, one of the two recently described gene pairs in the Xq21.3/Yp11.2 hominid-specific homology block, in samples from Portugal and Mozambique. Sharp differences were observed on X/Y allele distributions, concerning both the presence of private alleles and a different modal repeat length for X-linked and Y-linked markers, and this difference was statistically significant. Higher diversity was found in X-linked microsatellites than in their Y chromosome counterparts; when comparing populations, Mozambicans showed more allele diversity for the X chromosome, but the contrary was true for the Y chromosome microsatellites. Evolutionary patterns, relying on intragenic PCDHX/Y SNPs, also revealed distinct scenarios for X and Y chromosomes. Greater microsatellite diversity was displayed by African X chromosomes within the most common haplotypes shared by both populations, whereas higher microsatellite diversity was found in Portugal for the ancestral Y chromosome haplotype. The most frequent PCDHY haplotype in Portuguese was the derived one, and it was not found in Mozambicans. TMRCA estimated by the rho parameter resulted in 13,700 years (7,500-20,000 years), which is consistent with a recent, post-Out-of-Africa origin for this haplotype. In conclusion, the newly described microsatellite loci generally displayed greater X-linked to Y-linked diversity and this pattern was also detected with slower evolving markers, with a remarkable differentiation between populations observed for Y chromosome haplotypes and, thus, greater divergence among Y chromosomes in human populations. PMID- 15297599 TI - Molecular claims of Gondwanan age for Australian agamid lizards are untenable. AB - A recent mtDNA study proposes a surprisingly deep (approximately 150 MYA) divergence between SE Asian and Australasian agamid lizards, consistent with ancient Gondwanan vicariance rather than dispersal across the Indonesian Archipelago. However, the analysis contains a fundamental error: use of rates of molecular evolution inferred from uncorrected sequence divergence to put a time frame on a tree with branch lengths greatly elongated by complex likelihood and rate-smoothing models. Furthermore, this date implies that basal splits within agamids occurred implausibly early, at least 300 MYA (100 Myr before the first fossil lizards and coincident with the earliest fossil reptiles). Analyses of the mtDNA data using more appropriate methods and new information from nuclear (c mos) sequences suggest a much more recent divergence between SE Asian and Australian agamids (around 30 MYA). Using two fossil boundary dates, bootstrapping the c-mos data gives a 95% confidence interval for this divergence time that is sufficiently recent (14-41 MYA) to exclude an ancient Gondwanan vicariance and is more consistent with Miocene over-water dispersal. As with the mtDNA, the c-mos data implies implausibly old basal divergences among agamids if a Gondwanan age is assumed for the Australasian clade. The analyses also highlight how methods for creating ultrametric trees (especially nonparametric rate smoothing) can greatly modify branch lengths and, thus, always require internal calibrations. The errors associated with inferred dates in the previous study (inferred through parametric bootstrapping) were also unjustifiably low, as this method only considers stochasticity in the substitution model and ignores much larger sources of uncertainty, such as variation in character sampling, tree topology, and calibration accuracy. PMID- 15297600 TI - Predicting mammalian SINE subfamily activity from A-tail length. AB - Based on previous observations that newly inserted LINEs and SINEs have particularly long 3' A-tails, which shorten rapidly during evolutionary time, we have analyzed the rat and mouse genomes for evidence of recently inserted SINEs and LINEs. We find that the youngest predicted subfamilies of rodent identifier (ID) elements, a rodent-specific SINE derived from tRNA(Ala), are preferentially associated with A-tails over 50 bases in the rat genome, as predicted. Furthermore, these studies detected a subfamily of ID elements that has made over 15,000 copies that is younger than any previously reported ID subfamily. We use PCR analysis of genomic loci to demonstrate that all subfamily members tested inserted after the divergence of Rattus norvegicus from Rattus rattus. We also found evidence that the rodent B1 family of elements is much more active currently in mouse than in rat. These data provide useful estimates of recent activity from all of the mammalian retrotransposons, as well as allowing identification of the most recent insertions for use as population and speciation markers in those species. Both the current rat ID and mouse B1 elements that are active have small, specific interruptions in their 3' A-tail sequences. We suggest that these interruptions stabilize the length of the A-tails and contribute to the activity of these subfamilies. We present a model in which the dynamics of the 3' A-tail may be a central controlling factor in SINE activity. PMID- 15297601 TI - The evolutionary repertoires of the eukaryotic-type ABC transporters in terms of the phylogeny of ATP-binding domains in eukaryotes and prokaryotes. AB - ABC (ATP-binding cassette) transporters play an important role in the communication of various substrates across cell membranes. They are ubiquitous in prokaryotes and eukaryotes, and eukaryotic types (EK-types) are distinguished from prokaryotic types (PK-types) in terms of their genes and domain organizations. The EK-types and PK-types mainly consist of exporters and importers, respectively. Prokaryotes have both the EK-types and the PK-types. The EK-types in prokaryotes are usually called "bacterial multidrug ABC transporters," but they are not well characterized in comparison with the multidrug ABC transporters in eukaryotes. Thus, an exhaustive search of the EK types among diverse organisms and detailed sequence classification and analysis would elucidate the evolutionary history of EK-types. It would also help shed some light on the fundamental repertoires of the wide variety of substrates through which multidrug ABC transporters in eukaryotes communicate. In this work, we have identified the EK-type ABC transporters in 126 prokaryotes using the profiles of the ATP-binding domain (NBD) of the EK-type ABC transporters from 12 eukaryotes. As a result, 11 clusters were identified from 1,046 EK-types ABC transporters. In particular, two large novel clusters emerged, corresponding to the bacterial multidrug ABC transporters related to the ABCB and ABCC families in eukaryotes, respectively. In the genomic context, most of these genes are located alone or adjacent to genes from the same clusters. Additionally, to detect functional divergences in the NBDs, the Kullback-Leibler divergence was measured among these bacterial multidrug transporters. As a result, several putative functional regions were identified, some corresponding to the predicted secondary structures. We also analyzed a phylogeny of the EK-type ABC transporters in both prokaryotes and eukaryotes, which revealed that the EK-type ABC transporters in prokaryotes have certain repertoires corresponding to the conventional ABC protein groups in eukaryotes. On the basis of these findings, we propose an updated evolutionary hypothesis in which the EK-type ABC transporters in both eukaryotes and prokaryotes consisted of several kinds of ABC transporters in putative ancestor cells before the divergence of eukaryotic and prokaryotic cells. PMID- 15297602 TI - Long-term conservation of six duplicated structural genes in cephalopod mitochondrial genomes. AB - The complete nucleotide sequences of the mitochondrial (mt) genomes of three cephalopods, Octopus vulgaris (Octopodiformes, Octopoda, Incirrata), Todarodes pacificus (Decapodiformes, Oegopsida, Ommastrephidae), and Watasenia scintillans (Decapodiformes, Oegopsida, Enoploteuthidae), were determined. These three mt genomes encode the standard set of metazoan mt genes. However, W. scintillans and T. pacificus mt genomes share duplications of the longest noncoding region, three cytochrome oxidase subunit genes and two ATP synthase subunit genes, and the tRNA(Asp) gene. Southern hybridization analysis of the W. scintillans mt genome shows that this single genome carries both duplicated regions. The near-identical sequence of the duplicates suggests that there are certain concerted evolutionary mechanisms, at least in cephalopod mitochondria. Molecular phylogenetic analyses of mt protein genes are suggestive, although not statistically significantly so, of a monophyletic relationship between W. scintillans and T. pacificus. PMID- 15297603 TI - Mitogenic activity of estrogens in human breast cancer cells does not rely on direct induction of mitogen-activated protein kinase/extracellularly regulated kinase or phosphatidylinositol 3-kinase. AB - We have addressed the question of rapid, nongenomic mechanisms that may be involved in the mitogenic action of estrogens in hormone-dependent breast cancer cells. In quiescent, estrogen-deprived MCF-7 cells, estradiol did not induce a rapid activation of either the MAPK/ERK or phosphatidylinositol-3 kinase (PI 3K)/Akt pathway, whereas the entry into the cell cycle was documented by the successive inductions of cyclin D1 expression, hyperphosphorylation of the retinoblastoma protein (Rb), activity of the promoter of the cyclin A gene, and DNA synthesis. However, pharmacological inhibitors of the src family kinases, 4 amino-5-(4-methylphenyl)-7-(t-butyl) pyrazolo[3,4-d] pyrimidine (PP1) or of the PI-3K (LY294002) did prevent the entry of the cells into the cell cycle and inhibited the late G1 phase progression, whereas the inhibitor of MAPK/ERK activation (U0126) had only a partial inhibitory effect in the early G1 phase. In agreement with these results, small interfering RNA targeting Akt strongly inhibited the estradiolinduced cell cycle progression monitored by the activation of the promoter of the cyclin A gene. The expression of small interfering RNA targeting MAPK 1 and 2 also had a clear inhibitory effect on the estradiol induced activation of the cyclin A promoter and also antagonized the estradiol induced transcription directed by the estrogen response element. Finally, transfection of the estrogen receptor into NIH3T3 fibroblasts did not confer to the cells sensitivity to a mitogenic action of estradiol. We conclude that the induction of the cell cycle by estradiol does not require a direct activation of MAPK/ERK or PI-3K signaling protein kinase cascades, but that these kinases appear to have a permissive role in the cell cycle progression. PMID- 15297604 TI - Follicle-stimulating hormone induced changes in gene expression of murine testis. AB - Even though FSH is not required for qualitatively normal spermatogenesis, it plays an important role in the spermatogenic capacity of the testis. Although the actions of FSH are well documented, most of these studies were done in vitro, and the molecular targets of FSH in vivo remain largely unverified. To understand the complete mechanism of FSH actions in spermatogenesis, it is important to identify the genes that are involved in its signaling, and know how these genes are affected by FSH. We have used hypogonadal (hpg) mouse that lacks circulating FSH as an in vivo model in conjunction with the Affymetrix murine GeneChip U74A (12,488 genes) to monitor changes in testicular gene expression as a result of FSH signaling. Hpg mice were injected with 10 IU ovine FSH, killed 4, 8, 12, or 24 h post treatment, and their testicular gene expression was compared with that of untreated control hpg mice. The abundance of a large number of mRNAs was affected by the FSH treatment. The primary effect of FSH resulted in increased steady-state levels of many mRNAs in testes of hpg mice. Several transcripts were identified whose abundance was decreased as well. We have used real-time PCR to confirm the changes in levels of transcripts such as renin-1, Kruppel-like factor 4, Mad4 (max-interacting protein repressor), Nur-related protein 1, and hairy/enhancer of splits gene 1 that were found to be regulated by FSH in testes of hpg mice. PMID- 15297605 TI - Pancreatic islet progenitor cells in neurogenin 3-yellow fluorescent protein knock-add-on mice. AB - The basic helix-loop-helix transcription factor Neurogenin 3 (NGN3) controls endocrine cell fate specification in uncommitted pancreatic progenitor cells. Ngn3-deficient mice do not develop any islet cells and are diabetic. All the major islet cell types, including insulin-producing beta-cells, derive from Ngn3 positive endocrine progenitor cells. Therefore, the characterization of this population of immature cells is of particular interest for the development of novel strategies for cell replacement therapies in type 1 diabetes. To explore further the biology of islet progenitor cells we have generated a mouse in which Ngn3-expressing cells are labeled with the enhanced yellow fluorescent protein (EYFP) using a knock-add-on strategy. In this approach, the EYFP cDNA is introduced into the 3'-untranslated region of the proendocrine transcription factor, Neurogenin 3, without deleting any endogenous coding or regulatory sequences. In Ngn3(EYFP/+) and Ngn3(EYFP/EYFP) mice, the EYFP protein is targeted to Ngn3-expressing progenitors in the developing pancreas, and islets develop normally. Islet progenitors can be purified from whole embryonic pancreas by fluorescence-activated cell sorting from Ngn3(EYFP/+) mice and their development can be monitored in real time in pancreas explant cultures. These experiments showed that endocrine progenitors can form de novo and expand, in vitro, in the absence of signals from the surrounding mesenchyme, suggesting that endocrine commitment is a default pathway. The Ngn3(EYFP) mice represent a valuable tool to study islet cell development and neogenesis in normal and diabetic animals as well as for the determination of the conditions to generate beta-cells in vitro. PMID- 15297606 TI - Regulation of signal transducer and activator of transcription 1 (STAT1) and STAT1-dependent genes by RET/PTC (rearranged in transformation/papillary thyroid carcinoma) oncogenic tyrosine kinases. AB - Chimeric RET/PTC (rearranged in transformation/papillary thyroid carcinoma) oncoproteins are constitutively active tyrosine kinases found in thyroid papillary carcinoma and nonneoplastic Hashimoto's thyroiditis. Although several proteins have been identified to be substrates of RET/PTC kinases, the pathogenic roles played by RET/PTC in malignant and benign thyroid diseases and the molecular mechanisms that are involved are not fully understood. We found that RET/PTC expression phosphorylates the Y701 residue of STAT1, a type II interferon (IFN)-responsive protein. RET/PTC-mediated signal transducer and activator of transcription 1 (STAT1) phosphorylation requires RET/PTC kinase activity to be intact but other tyrosine kinases, such as Janus kinases or c-Src, are not involved. RET/PTC-induced STAT1 transcriptional activation was not inhibited by suppressor of cytokine signaling-1 or -3, or protein inhibitors of activated STAT3 [(protein inhibitor of activated STAT (PIAS3)], but PIAS1 strongly repressed the RET/PTC-induced transcriptional activity of STAT1. RET/PTC-induced STAT1 activation caused IFN regulatory factor-1 expression. We found that STAT1 and IFN regulatory factor-1 cooperated to significantly increase transcription from type IV IFN-gamma responsive promoters of class II transactivator genes. Significantly, cells stably expressing RET/PTC expressed class II transactivator and showed enhanced de novo membrane expression of major histocompatibility complex (MHC) class II proteins. Furthermore, RET/PTC1-bearing papillary thyroid carcinoma cells strongly expressed MHC class II (human leukocyte-associated antigen-DR alpha) genes, whereas the surrounding normal tissues did not. Thus, RET/PTC is able to phosphorylate and activate STAT1. This may lead to enhanced MHC class II expression, which may explain why the tissues surrounding RET/PTC positive cancers are infiltrated with lymphocytes. Such immune response-promoting activity of RET/PTC may also relate to the development of Hashimoto's thyroiditis. PMID- 15297607 TI - The expression level of the orphan nuclear receptor GCNF (germ cell nuclear factor) is critical for neuronal differentiation. AB - The germ cell nuclear factor (GCNF) is essential for normal embryonic development and gametogenesis. To test the prediction that GCNF is additionally required for neuronal differentiation, we used the mouse embryonal carcinoma cell line PCC7 Mz1, which represents an advantageous model to study neuronal cells from the stage of fate choice until the acquirement of functional competence. We generated stable transfectants that express gcnf sense or antisense RNA under the control of a tetracycline-regulated promoter. After retinoic acid-induced withdrawal from the cell cycle, sense clones developed a neuron network with changed properties, and the time course of neuron maturation was shortened. Consistent with these data, differentiation of neuronal precursor cells was impaired in antisense cultures. This involved a delay in 1) the down-regulation of nestin, a marker for undifferentiated neuroepithelial cells and stem cells of the central nervous system, and 2) up-regulation of the somatodendritic protein microtubule associated protein 2 and the synaptic vesicle protein synaptophysin. Neuronal cells in the antisense cultures acquired functional competence, although with a significant delay. Our data propose that the level of GCNF is critical for differentiation and maturation of neuronal precursor cells. PMID- 15297608 TI - Strategies and mechanisms for host and pathogen survival in acute and persistent viral infections. AB - Persistent viral infections causing serious diseases derive, primarily, from altered function of the immune system. Knowledge of the very complex composition and function of the innate and adaptive branches of the immune system is essential to understanding persistent infection. The best solution to the problem of persistent infection is by prevention using prophylactic vaccines. Hit and run viruses evade immune destruction by infecting new hosts and rarely persist. Hit and stay viruses evade immune control by sequestration, blockade of antigen presentation, cytokine escape, evasion of natural killer cell activities, escape from apoptosis, and antigenic change. Twelve prophylactic vaccines against hit and run agents exist, and there are only three vaccines against hit and stay viruses, all of which are of DNA composition. Several new vaccines against hit and stay viruses are feasible, but protective vaccines against RNA HIV and hepatitis C agents are highly unlikely, short of a major breakthrough. Therapeutic vaccines are very improbable without a magnitude of favorable new discoveries. In the meantime, antiviral chemotherapy, chemotherapy/prophylactic vaccination, and short interfering RNA silencing are worthy of intense investigation. PMID- 15297609 TI - SspB delivery of substrates for ClpXP proteolysis probed by the design of improved degradation tags. AB - The ssrA-degradation tag sequence contains contiguous binding sites for the SspB adaptor and the ClpX component of the ClpXP protease. Although SspB normally enhances ClpXP degradation of ssrA-tagged substrates, it inhibits proteolysis under conditions that prevent tethering to ClpX. By increasing the spacing between the protease and adaptor-binding determinants in the ssrA tag, substrates were obtained that displayed improved SspB-mediated binding to and degradation by ClpXP. These extended-tag substrates also showed significantly reduced conditional inhibition but bound SspB normally. Both wild-type and mutant tags showed highly dynamic SspB interactions. Together, these results strongly support delivery models in which SspB and ClpX bind concurrently to the ssrA tag, but also suggest that clashes between SspB and ClpX weaken simultaneous binding. During substrate delivery, this signal masking is overcome by tethering SspB to ClpX, which ensures local concentrations high enough to drive tag engagement. This obstruct-then-stimulate mechanism may have evolved to allow additional levels of regulation and could be a common trait of adaptor-mediated protein degradation. PMID- 15297610 TI - Autocatalytic self-propagation of misfolded prion protein. AB - Prions are thought to replicate in an autocatalytic process that converts cellular prion protein (PrP(C)) to the disease-associated misfolded PrP isoform (PrP(Sc)). Our study scrutinizes this hypothesis by in vitro protein misfolding cyclic amplification (PMCA). In serial transmission PMCA experiments, PrP(Sc) was inoculated into healthy hamster brain homogenate containing PrP(C). Misfolded PrP was amplified by rounds of sonication and incubation and reinoculated into fresh brain homogenate every 10 PMCA rounds. The amplification depended on PrP(C) substrate and could be inhibited by recombinant hamster PrP. In serial dilution experiments, newly formed misfolded and proteinase K-resistant PrP (PrPres) catalyzed the structural conversion of PrP(C) as efficiently as PrP(Sc) from brain of scrapie (263K)-infected hamsters, yielding an approximately 300-fold total amplification of PrPres after 100 rounds, which confirms an autocatalytic PrP-misfolding cascade as postulated by the prion hypothesis. PrPres formation was not paralleled by replication of biological infectivity, which appears to require factors additional to PrP-misfolding autocatalysis. PMID- 15297611 TI - Highly pathogenic SHIVs and SIVs target different CD4+ T cell subsets in rhesus monkeys, explaining their divergent clinical courses. AB - In contrast to simian immunodeficiency viruses (SIVs), which induce immunodeficiency over a 1- to 3-year period, highly pathogenic simian-human immunodeficiency viruses (SHIVs) cause a complete, irreversible, and systemic depletion of CD4(+) T lymphocytes in rhesus monkeys within weeks of infection. By using small-molecule competitors specific for CCR5 and CXCR4 in ex vivo assays, we found that highly pathogenic SHIV(DH12R) exclusively uses CXCR4 for infection of rhesus peripheral blood mononuclear cells, whereas SIV(mac239) and SIV(smE543) use CCR5 for entry into the same cells. During the period of peak virus production in SHIV(DH12R)- or SHIV(89.6P)-infected rhesus monkeys, massive elimination of CXCR4(+) naive CD4(+) T cells occurred. In contrast, circulating CCR5(+) memory CD4(+) T cells were selectively depleted in rapidly progressing SIV-infected monkeys. At the time of their death, two SIV rapid progressors had experienced a nearly complete loss of the memory CD4(+) T cell subset from the blood and mesenteric lymph nodes. Thus, pathogenic SHIVs and SIVs target different subsets of CD4(+) T cells in vivo, with the pattern of CD4(+) T lymphocyte depletion being inextricably linked to chemokine receptor use. In the context of developing an effective prophylactic vaccine, which must potently control virus replication during the primary infection, regimens that suppress SHIVs might not protect monkeys against SIV or humans against HIV-1. PMID- 15297612 TI - Sulfur pollution suppression of the wetland methane source in the 20th and 21st centuries. AB - Natural wetlands form the largest source of methane (CH(4)) to the atmosphere. Emission of this powerful greenhouse gas from wetlands is known to depend on climate, with increasing temperature and rainfall both expected to increase methane emissions. This study, combining our field and controlled environment manipulation studies in Europe and North America, reveals an additional control: an emergent pattern of increasing suppression of methane (CH(4)) emission from peatlands with increasing sulfate (SO(4)(2-)-S) deposition, within the range of global acid deposition. We apply a model of this relationship to demonstrate the potential effect of changes in global sulfate deposition from 1960 to 2080 on both northern peatland and global wetland CH(4) emissions. We estimate that sulfur pollution may currently counteract climate-induced growth in the wetland source, reducing CH(4) emissions by approximately 15 Tg or 8% smaller than it would be in the absence of global acid deposition. Our findings suggest that by 2030 sulfur pollution may be sufficient to reduce CH(4) emissions by 26 Tg or 15% of the total wetland source, a proportion as large as other components of the CH(4) budget that have until now received far greater attention. We conclude that documented increases in atmospheric CH(4) concentration since the late 19th century are likely due to factors other than the global warming of wetlands. PMID- 15297613 TI - Quasiclassical computation. AB - The chemical kinetic description of time evolution where the phase is random but the states are discrete is discussed as a basis for a computational approach. This proposed scheme uses numbers in the entire range of 0 to 1 to represent Boolean propositions. In the implementation by chemical kinetics these numbers are the mole fractions of different species. Vibrational relaxation in a mixture of HCl and DCI is the physical system that is used to illustrate the approach. Energy exchange in such a mixture corresponds to two strongly coupled two-level systems. A search problem, previously discussed in the quantum computing literature, is solved as an example. The solution requires the same number of function evaluations as in the quantal case. The action of the oracle is described in detail. PMID- 15297614 TI - CisModule: de novo discovery of cis-regulatory modules by hierarchical mixture modeling. AB - The regulatory information for a eukaryotic gene is encoded in cis-regulatory modules. The binding sites for a set of interacting transcription factors have the tendency to colocalize to the same modules. Current de novo motif discovery methods do not take advantage of this knowledge. We propose a hierarchical mixture approach to model the cis-regulatory module structure. Based on the model, a new de novo motif-module discovery algorithm, CisModule, is developed for the Bayesian inference of module locations and within-module motif sites. Dynamic programming-like recursions are developed to reduce the computational complexity from exponential to linear in sequence length. By using both simulated and real data sets, we demonstrate that CisModule is not only accurate in predicting modules but also more sensitive in detecting motif patterns and binding sites than standard motif discovery methods are. PMID- 15297615 TI - Potent block of Cx36 and Cx50 gap junction channels by mefloquine. AB - Recently, great interest has been shown in understanding the functional roles of specific gap junction proteins (connexins) in brain, lens, retina, and elsewhere. Some progress has been made by studying knockout mice with targeted connexin deletions. For example, such studies have implicated the gap junction protein Cx36 in synchronizing rhythmic activity of neurons in several brain regions. Although knockout strategies are informative, they can be problematic, because compensatory changes sometimes occur during development. Therefore, it would be extremely useful to have pharmacological agents that block specific connexins, without major effects on other gap junctions or membrane channels. We show that mefloquine, an antimalarial drug, is one such agent. It blocked Cx36 channels, expressed in transfected N2A neuroblastoma cells, at low concentrations (IC(50) approximately 300 nM). Mefloquine also blocked channels formed by the lens gap junction protein, Cx50 (IC(50) approximately 1.1 microM). However, other gap junctions (e.g., Cx43, Cx32, and Cx26) were only affected at concentrations 10- to 100-fold higher. To further examine the utility and specificity of this compound, we characterized its effects in acute brain slices. Mefloquine, at 25 microM, blocked gap junctional coupling between interneurons in neocortical slices, with minimal nonspecific actions. At this concentration, the only major side effect was an increase in spontaneous synaptic activity. Mefloquine (25 microM) caused no significant change in evoked excitatory or inhibitory postsynaptic potentials, and intrinsic cellular properties were also mostly unaffected. Thus, mefloquine is expected to be a useful tool to study the functional roles of Cx36 and Cx50. PMID- 15297616 TI - MED16 and MED23 of Mediator are coactivators of lipopolysaccharide- and heat shock-induced transcriptional activators. AB - Transcriptional activators interact with diverse proteins and recruit transcriptional machinery to the activated promoter. Recruitment of the Mediator complex by transcriptional activators is usually the key step in transcriptional activation. However, it is unclear how Mediator recognizes different types of activator proteins. To systematically identify the subunits responsible for the signal- and activator-specific functions of Mediator in Drosophila melanogaster, each Mediator subunit was depleted by RNA interference, and its effect on transcriptional activation of endogenous as well as synthetic promoters was examined. The depletion of some Mediator gene products caused general transcriptional defects, whereas depletion of others caused defects specifically related to activation. In particular, MED16 and MED23 were required for lipopolysaccharide- and heat-shock-specific gene expression, respectively, and their activator-specific functions appeared to result from interaction with specific activators. The corequirement of MED16 for other forms of differentiation-inducing factor-induced transcription was confirmed by microarray analysis of differentiation-inducing factor (DIF)- and MED16-depleted cells individually. These results suggest that distinct Mediator subunits interact with specific activators to coordinate and transfer activator-specific signals to the transcriptional machinery. PMID- 15297617 TI - Efficient system-wide coordination in noisy environments. AB - Many natural and social systems display global organization and coordination without centralized control. The origin of this global coordination is a topic of great current interest. Here we investigate a density-classification task as a model system for coordination and information processing in decentralized systems. We show that sophisticated strategies, selected under idealized conditions, are not robust to environmental changes. We also demonstrate that a simple heuristic is able to successfully complete the classification task under a broad range of environmental conditions. Our findings hint at the possibility that complex networks and ecologically efficient rules coevolve over time. PMID- 15297618 TI - Goal-directed whisking increases phase-locking between vibrissa movement and electrical activity in primary sensory cortex in rat. AB - We tested the hypothesis that behavioral context modulates phase-locking between rhythmic motor activity and concomitant electrical activity induced in primary sensory (S1) cortex. We used exploratory whisking by rat as a model system and recorded two measures: (i) the mystacial electromyogram ( nabla EMG) as a surrogate of vibrissa position, and (ii) the field potential ( nabla LFP) in S1 cortex as an indicator of electrical activity. The degree to which the nabla EMG and nabla LFP were phase-locked was compared for three categories of rhythmic whisking: (i) searching for an object with the vibrissae for a food reward, (ii) whisking in air for the goal of returning to the home cage, and (iii) whisking with no reward. We observed that the magnitude of phase-locking was nearly tripled for the two rewarded conditions compared to unrewarded whisking. Critically, increased locking was not accompanied by an increase in the amplitude of the cortical nabla LFP for the rewarded tasks. Additional experiments showed that there was no significant relation between the amplitude of a sensory-evoked response in S1 cortex and the magnitude of the locking between the nabla EMG and the nabla LFP during whisking. We conclude that the behavioral context of a whisking task can increase the modulation of S1 cortical activity by motor output without a concomitant increase in the magnitude of activity. PMID- 15297619 TI - Immunotherapy for Alzheimer's disease. AB - The utility of vaccine strategies to treat neurodegenerative diseases such as Alzheimer's disease (AD) may still hold promise. Both active and passive immunization strategies reduced AD-like pathology and restored cognitive deficits in transgenic mice. These results were initially met with considerable optimism; however, phase IIa clinical trials were halted because of a small but significant occurrence of meningoencephalitis. Knowledge gained from studies on amyloid-beta peptide (A beta) immunotherapy will allow optimization of new-generation vaccines, targeting highly specific epitopes while reducing undesired side effects. In harnessing and steering the immune system, an effective response can be generated against A beta. If this proves successful, A beta vaccination could provide the first definitive treatment for AD. PMID- 15297620 TI - Therapeutic cancer vaccines: using unique antigens. AB - A decade ago, it seemed rational that our rapidly increasing knowledge of the molecular identities of tumor antigens and a deeper understanding of basic immunology would point the way to an effective therapeutic cancer vaccine. Significant progress has been made, but we do not yet have a cancer vaccine that can reliably and consistently induce tumor destruction or improve patient survival. Random mutations in cancer cells generate unique antigens in each individual, and this may be important in terms of generating a therapeutic immune response. Autologous heat shock protein-peptide complexes produced from each patient's tumor is a logical personalized approach that may obviate the need to identify the unique antigens contained in the individual vaccine. Heat shock proteins elicit adaptive and innate immune responses and have been tested in a variety of animal models and different human cancers. Activity has been seen in several animal studies. Early-phase human studies have also suggested some activity in certain cancers. Large, randomized phase 3 studies are ongoing, and these will effectively answer the question of efficacy regarding this approach to therapeutic vaccination. There are sufficient data to support the notion that cancer vaccines can induce anti-tumor immune responses in humans with cancer. How best to translate this increase in immune responsiveness to consistently and reproducibly induce objective cancer regression or increased survival remains unclear at this time. PMID- 15297621 TI - Energetic clues to pathways to biomineralization: precursors, clusters, and nanoparticles. AB - Nanoparticle and nanocluster precursors may play a major role in biomineralization. The small differences in enthalpy and free energy among metastable nanoscale phases offer controlled thermodynamic and mechanistic pathways. Clusters and nanoparticles offer concentration and controlled transport of reactants. Control of polymorphism, surface energy, and surface charge on nanoparticles can lead to morphological control and appropriate growth rates of biominerals. Rather than conventional nucleation and growth, assembly of nanoparticles may provide alternative mechanisms for crystal growth. The Ostwald step rule, based on a thermodynamic view of nucleation and growth, is supported by the observation that more metastable phases tend to have lower surface energies. Examples from nonbiological systems, stressing the interplay of thermodynamic and kinetic factors, illustrate features potentially important to biomineralization. PMID- 15297622 TI - Essential role for virus-neutralizing antibodies in sterilizing immunity against Friend retrovirus infection. AB - The current experiments use the Friend retrovirus model to demonstrate that vaccine-primed B cells are essential for sterilizing immunity, and the results indicate that the requisite function of these cells is the production of virus neutralizing antibodies rather than priming or reactivation of T cells. B cell deficient mice were poorly protected by vaccination, but adoptive transfer experiments showed that the T cells from B cell-deficient mice were primed as well as those from wild-type mice. Furthermore, passive transfer of virus neutralizing antibodies completely compensated for B cell deficiency. The presence of virus-neutralizing antibodies at the time of infection was crucial for vaccine efficacy. Interestingly, virus-neutralizing antibodies worked synergistically with vaccine-primed T cells to provide a level of protection many orders of magnitude greater than either antibodies or immune T cells alone. Nonneutralizing antibodies also contributed to protection and acted cooperatively with neutralizing antibodies to reduce infection levels. These results emphasize the importance of inducing both T cell responses and virus-neutralizing antibody responses for effective retroviral vaccine protection. PMID- 15297623 TI - Evidence for deep magma injection beneath Lake Tahoe, Nevada-California. AB - A deep earthquake swarm in late 2003 at Lake Tahoe, California (Richter magnitude < 2.2; depth of 29 to 33 kilometers), was coeval with a transient displacement of 6 millimeters horizontally outward from the swarm and 8 millimeters upward measured at global positioning system station Slide Mountain (SLID) 18 kilometers to the northeast. During the first 23 days of the swarm, hypocentral depths migrated at a rate of 2.4 millimeters per second up-dip along a 40-square kilometer structure striking north 30 degrees west and dipping 50 degrees to the northeast. SLID's transient velocity of 20 millimeters per year implies a lower bound of 200 nanostrains per year (parts per billion per year) on local strain rates, an order of magnitude greater than the 1996 to 2003 regional rate. The geodetic displacement is too large to be explained by the elastic strain from the cumulative seismic moment of the sequence, suggesting an aseismic forcing mechanism. Aspects of the swarm and SLID displacements are consistent with lower crustal magma injection under Lake Tahoe. PMID- 15297624 TI - Small interfering RNA-induced transcriptional gene silencing in human cells. AB - Small interfering RNA (siRNA) and microRNA silence genes at the transcriptional, posttranscriptional, and/or translational level. Using human tissue culture cells, we show that promoter-directed siRNA inhibits transcription of an integrated, proviral, elongation factor 1alpha (EF1A) promoter-green fluorescent protein reporter gene and of endogenous EF1A. Silencing was associated with DNA methylation of the targeted sequence, and it required either active transport of siRNA into the nucleus or permeabilization of the nuclear envelope by lentiviral transduction. These results demonstrate that siRNA-directed transcriptional silencing is conserved in mammals, providing a means to inhibit mammalian gene function. PMID- 15297625 TI - SRC mediates a switch from microtubule- to actin-based motility of vaccinia virus. AB - The cascade of events that leads to vaccinia-induced actin polymerization requires Src-dependent tyrosine phosphorylation of the viral membrane protein A36R. We found that a localized outside-in signaling cascade induced by the viral membrane protein B5R is required to potently activate Src and induce A36R phosphorylation at the plasma membrane. In addition, Src-mediated phosphorylation of A36R regulated the ability of virus particles to recruit and release conventional kinesin. Thus, Src activity regulates the transition between cytoplasmic microtubule transport and actin-based motility at the plasma membrane. PMID- 15297626 TI - Mitochondrial fusion intermediates revealed in vitro. AB - The events that occur during the fusion of double-membraned mitochondria are unknown. As an essential step toward determining the mechanism of mitochondrial fusion, we have captured this event in vitro. Mitochondrial outer and inner membrane fusion events were separable and mechanistically distinct, but both required guanosine 5'-triphosphate hydrolysis. Homotypic trans interactions of the ancient outer transmembrane guanosine triphosphatase, Fzo1, were required to promote the fusion of mitochondrial outer membranes, whereas electrical potential was also required for fusion of inner membranes. Our conclusions provide fundamental insights into the molecular events driving mitochondrial fusion and advance our understanding of the evolution of mitochondrial fusion in eukaryotic cells. PMID- 15297627 TI - Linked expression of Ah receptor, ARNT, CYP1A1, and CYP1B1 in rat mammary epithelia, in vitro, is each substantially elevated by specific extracellular matrix interactions that precede branching morphogenesis. AB - Cytochrome P4501B1 (CYP1B1), the major constitutively expressed CYP in the rat mammary gland, is induced by Ah-receptor (AhR) ligands, while CYP1A1 is predominantly expressed only after induction. These CYPs contribute to carcinogenic activation of polycyclic aromatic hydrocarbons (PAHs). AhR, ARNT, and CYP1B1 were only weakly expressed, even after 2,3,7,8-tetrachlorodibenzo-p dioxin induction, when rat mammary epithelial cells (RMEC) were cultured on plastic. RMEC cultured on the extracellular matrix (ECM), Matrigel, or on a floating gel of collagen I demonstrated branching morphogenesis and substantially increased basal CYP1B1 and induced CYP1A1 expression, in parallel with large increases in AhR and ARNT expression. Branching was more pronounced in the Wistar Kyoto than in the Wistar Furth rat strain. Although EGF enhanced branching, neither strain nor growth factor treatment substantially impacted CYP expression. Increased AhR and ARNT expression is observed within 24 h of dispersal on Matrigel, substantially prior to branch formation. Culture on thin layers of collagen I, collagen IV, and laminin, respectively, failed to reproduce the branching morphogenesis or increases in AhR, ARNT, or CYP expression. However, adherent, gelled collagen I recapitulated the increased protein expression, without supporting branching. This increased protein expression was closely paralleled by enhanced expression of beta-catenin and E-cadherin, components of cell-cell adhesion complexes. A synthetic peptide that selectively antagonizes integrin-ECM interactions reduced branch formation, without diminishing AhR, ARNT, and CYP expression. These data demonstrate that early ECM surface adhesion interactions mediate AhR and ARNT expression, which enhances CYP expression, independent of branching morphogenesis. PMID- 15297628 TI - Serum S100B predicts a malignant course of infarction in patients with acute middle cerebral artery occlusion. AB - BACKGROUND AND PURPOSE: Early predictors of infarct volume may improve therapeutic decisions in patients with acute cerebral ischemia. We investigated whether measurements of serum astroglial protein S100B can predict a malignant course of infarction in acute middle cerebral artery (MCA) occlusion. METHODS: We included 51 patients (24 women, mean age 69.1+/-12.4 years) admitted within 6 hours after stroke symptom onset caused by proximal MCA occlusion, as shown by magnetic resonance angiography (n=39), intra-arterial angiography (n=4), or transcranial duplex sonography (n=8). Blood samples were drawn at hospital admission and 8, 12, 16, 20, and 24 hours after symptom onset. Serum S100B concentrations were determined using a fully automated immunoluminometric assay. A malignant course of infarction was defined as the occurrence of clinical signs of cerebral herniation within the first 7 days of treatment or the clinical decision to perform decompressive hemicraniectomy caused by critical space occupying swelling as detected by repeated neuroimaging. RESULTS: Sixteen patients developed malignant infarction (31%). Beginning with the 12-hour value, mean S100B serum concentrations were significantly higher in patients with a malignant course compared with those without (12 hours 1.23+/-1.24 versus 0.29+/ 0.45 microg/L; 16 hours 1.80+/-1.65 versus 0.38+/-0.53 microg/L; 20 hours 1.90+/ 1.53 versus 0.44+/-0.48 microg/L; and 24 hours 2.41+/-1.59 versus 0.57+/-0.66 microg/L; all P<0.001). A 12-hour S100B value >0.35 microg/L predicted malignant infarction with 0.75 sensitivity and 0.80 specificity. A 24-hour value >1.03 microg/L provided 0.94 sensitivity and 0.83 specificity. CONCLUSIONS: The serum marker S100B can predict a malignant course of infarction in proximal MCA occlusion. This finding may improve the identification and monitoring of patients at particularly high risk for herniation. PMID- 15297629 TI - Functional MRI follow-up study of language processes in healthy subjects and during recovery in a case of aphasia. AB - BACKGROUND AND PURPOSE: The goal of this study was to develop a functional MRI (fMRI) paradigm robust and reproducible enough in healthy subjects to be adapted for a follow-up study aiming at evaluating the anatomical substratum of recovery in poststroke aphasia. METHODS: Ten right-handed subjects were studied longitudinally using fMRI (7 of them being scanned twice) and compared with a patient with conduction aphasia during the first year of stroke recovery. RESULTS: Controls exhibited reproducible activation patterns between subjects and between sessions during language tasks. In contrast, the patient exhibited dynamic changes in brain activation pattern, particularly in the phonological task, during the 2 fMRI sessions. At 1 month after stroke, language homotopic right areas were recruited, whereas large perilesional left involvement occurred later (12 months). CONCLUSIONS: We first demonstrate intersubject robustness and intrasubject reproducibility of our paradigm in 10 healthy subjects and thus its validity in a patient follow-up study over a stroke recovery time course. Indeed, results suggest a spatiotemporal poststroke brain reorganization involving both hemispheres during the recovery course, with an early implication of a new contralateral functional neural network and a later implication of an ipsilateral one. PMID- 15297630 TI - Association of polymorphisms and haplotypes in the elastin gene in Dutch patients with sporadic aneurysmal subarachnoid hemorrhage. AB - BACKGROUND AND PURPOSE: A locus containing the elastin gene has been linked to familial intracranial aneurysms in 2 distinct populations. We investigated the association of single-nucleotide polymorphisms (SNPs) and haplotypes of SNPs in the elastin gene with the occurrence of subarachnoid hemorrhage (SAH) from sporadic aneurysms in the Netherlands. METHODS: We genotyped 167 SAH patients and 167 matching controls for 18 exonic and intronic SNPs in the elastin gene. A Bonferroni correction was applied for multiple comparisons with all novel associations, with a correction factor derived from the number of SNPs tested (P value after Bonferroni correction [P(corr)]). RESULTS: SAH was statistically significant associated with an SNP in exon 22 of the elastin gene (minor allele frequency was 0.000 in patients and 0.028 in controls; odds ratio [OR], 0.0; 95% CI, 0.0 to 0.7; P=0.004; P(corr)=0.05) and possibly with an SNP in intron 5 (minor allele frequency was 0.062 in patients and 0.128 in controls; OR, 0.5; 95% CI, 0.2 to 0.8; P=0.007; P(corr)=0.08). Haplotypes of intron 5/exon 22 (P(corr)=0.002), intron 4/exon 22 (P(corr)=0.02), and intron 4/intron 5/exon 22 (P=9.0x10(-9)) were also associated with aneurysmal SAH. CONCLUSIONS: Variants and haplotypes within the elastin gene are associated with the risk of sporadic SAH in Dutch patients. Gradual increase of statistical power with the inclusion of 2 or 3 SNPs in the studied haplotypes supports the validity of our conclusion that the elastin gene is a susceptibility locus for SAH. PMID- 15297632 TI - European research momentum. PMID- 15297631 TI - Increased expression of transforming growth factor-beta1 as a stabilizing factor in human atherosclerotic plaques. AB - BACKGROUND AND PURPOSE: Transforming growth factor-beta (TGF-beta) is a growth factor/cytokine involved in vascular remodeling and atherogenesis. Recent studies in apolipoprotein E-deficient mice have demonstrated a pivotal role of TGF-beta in the maintenance of the balance between inflammation and fibrosis in atherosclerotic plaques. Furthermore, inhibition of TGF-beta signaling has been shown to accelerate plaque formation and its progression toward an unstable phenotype in mice. However, if this mechanism is operative also in humans is still unknown. The aim of this study was to characterize the expression of TGF beta1 in human carotid plaque and to correlate it with the extent of inflammatory infiltration and collagen content with the clinical signs of plaque instability. METHODS: Plaques were obtained from patients undergoing carotid endoarterectomy and divided into symptomatic and asymptomatic according to clinical evidence of recent transient ischemic attack or stroke. Plaques were analyzed for TGF-beta1 expression by Immunocytochemistry, Western, and Northern blotting analysis. Immunocytochemistry was used to identify CD68+ macrophages, CD3 T lymphocytes, HLA-DR+ cells, and alpha-smooth muscle cells. Procollagen and interstitial collagen content were analyzed by immunohistochemistry and Sirius Red staining, respectively. RESULTS: Plaque TGF-beta1 mRNA was increased up to 3-fold in asymptomatic as compared with symptomatic plaques. Plaques from asymptomatic group had fewer (P<0.0001) macrophages and T lymphocytes compared with symptomatic plaques. TGF-beta1 gene was transcriptionally active as demonstrated by increased (P<0.0001) TGF-beta1 protein expression in asymptomatic plaques. Immunohistochemistry showed that TGF-beta was mainly expressed in plaque shoulder and was associated with a comparable increase (P<0.0001) in plaque procollagen and collagen content. CONCLUSIONS: In conclusion, this study demonstrates the higher expression of TGF-beta1 in human asymptomatic lesions and provides evidence that TGF-beta1 may play an important role in the process of plaque stabilization. PMID- 15297633 TI - Election 2004. The calculus of making stem cells a campaign issue. PMID- 15297634 TI - Research protests. Britain unveils a plan to curb animal-rights 'extremists'. PMID- 15297635 TI - Neurobiology. Untangling Alzheimer's by paring plaques bolsters amyloid theory. PMID- 15297636 TI - Nanotechnology. Yellow light for nanotech. PMID- 15297637 TI - Astrophysics. Dark-matter sighting ends in shock. PMID- 15297638 TI - X-ray scan shows oldest known bird had a bird brain. PMID- 15297639 TI - Scientific publishing. Seeking advice on 'open access,' NIH gets an earful. PMID- 15297640 TI - In memoriam. Philip Hauge Abelson, 1913-2004. PMID- 15297641 TI - Evolutionary biology. The birth of the nucleus. PMID- 15297642 TI - Edward Hammond profile. Activist throws a bright light on institutes' biosafety panels. PMID- 15297643 TI - Planetary science. Rainbow of Martian minerals paints picture of degradation. PMID- 15297644 TI - Basic and clinical immunology meeting. An old favorite is resurrected: regulatory T cells take the stage. PMID- 15297645 TI - Basic and clinical immunology meeting. And action! Dendritic cells go live. PMID- 15297646 TI - Basic and clinical immunology meeting. Genes crisscross disease lines. PMID- 15297647 TI - Creating a European Research Council. PMID- 15297648 TI - Predators and prey in the Channel Islands. PMID- 15297649 TI - Comment on "Observation of the inverse Doppler effect". PMID- 15297651 TI - Neuroscience. Probing the neural basis of body ownership. PMID- 15297652 TI - Chemistry. The modest undressing of a silicon center. PMID- 15297653 TI - Physics. Only skin deep. PMID- 15297654 TI - Immunology. NK cells lose their inhibition. PMID- 15297655 TI - Metamaterials and negative refractive index. AB - Recently, artificially constructed metamaterials have become of considerable interest, because these materials can exhibit electromagnetic characteristics unlike those of any conventional materials. Artificial magnetism and negative refractive index are two specific types of behavior that have been demonstrated over the past few years, illustrating the new physics and new applications possible when we expand our view as to what constitutes a material. In this review, we describe recent advances in metamaterials research and discuss the potential that these materials may hold for realizing new and seemingly exotic electromagnetic phenomena. PMID- 15297657 TI - The Spirit Rover's Athena science investigation at Gusev Crater, Mars. AB - The Mars Exploration Rover Spirit and its Athena science payload have been used to investigate a landing site in Gusev crater. Gusev is hypothesized to be the site of a former lake, but no clear evidence for lacustrine sedimentation has been found to date. Instead, the dominant lithology is basalt, and the dominant geologic processes are impact events and eolian transport. Many rocks exhibit coatings and other characteristics that may be evidence for minor aqueous alteration. Any lacustrine sediments that may exist at this location within Gusev apparently have been buried by lavas that have undergone subsequent impact disruption. PMID- 15297658 TI - Pancam multispectral imaging results from the Spirit Rover at Gusev Crater. AB - Panoramic Camera images at Gusev crater reveal a rock-strewn surface interspersed with high- to moderate-albedo fine-grained deposits occurring in part as drifts or in small circular swales or hollows. Optically thick coatings of fine-grained ferric iron-rich dust dominate most bright soil and rock surfaces. Spectra of some darker rock surfaces and rock regions exposed by brushing or grinding show near-infrared spectral signatures consistent with the presence of mafic silicates such as pyroxene or olivine. Atmospheric observations show a steady decline in dust opacity during the mission, and astronomical observations captured solar transits by the martian moons, Phobos and Deimos, as well as a view of Earth from the martian surface. PMID- 15297659 TI - Surficial deposits at Gusev Crater along Spirit Rover traverses. AB - The Mars Exploration Rover Spirit has traversed a fairly flat, rock-strewn terrain whose surface is shaped primarily by impact events, although some of the landscape has been altered by eolian processes. Impacts ejected basaltic rocks that probably were part of locally formed lava flows from at least 10 meters depth. Some rocks have been textured and/or partially buried by windblown sediments less than 2 millimeters in diameter that concentrate within shallow, partially filled, circular impact depressions referred to as hollows. The terrain traversed during the 90-sol (martian solar day) nominal mission shows no evidence for an ancient lake in Gusev crater. PMID- 15297660 TI - Wind-related processes detected by the Spirit Rover at Gusev Crater, Mars. AB - Wind-abraded rocks, ripples, drifts, and other deposits of windblown sediments are seen at the Columbia Memorial Station where the Spirit rover landed. Orientations of these features suggest formative winds from the north-northwest, consistent with predictions from atmospheric models of afternoon winds in Gusev Crater. Cuttings from the rover Rock Abrasion Tool are asymmetrically distributed toward the south-southeast, suggesting active winds from the north-northwest at the time (midday) of the abrasion operations. Characteristics of some rocks, such as a two-toned appearance, suggest that they were possibly buried and exhumed on the order of 5 to 60 centimeters by wind deflation, depending on location. PMID- 15297661 TI - Spirit at Gusev Crater: plates. PMID- 15297662 TI - Localization and physical properties experiments conducted by Spirit at Gusev Crater. AB - The precise location and relative elevation of Spirit during its traverses from the Columbia Memorial station to Bonneville crater were determined with bundle adjusted retrievals from rover wheel turns, suspension and tilt angles, and overlapping images. Physical properties experiments show a decrease of 0.2% per Mars solar day in solar cell output resulting from deposition of airborne dust, cohesive soil-like deposits in plains and hollows, bright and dark rock coatings, and relatively weak volcanic rocks of basaltic composition. Volcanic, impact, aeolian, and water-related processes produced the encountered landforms and materials. PMID- 15297663 TI - Textures of the soils and rocks at Gusev Crater from Spirit's Microscopic Imager. AB - The Microscopic Imager on the Spirit rover analyzed the textures of the soil and rocks at Gusev crater on Mars at a resolution of 100 micrometers. Weakly bound agglomerates of dust are present in the soil near the Columbia Memorial Station. Some of the brushed or abraded rock surfaces show igneous textures and evidence for alteration rinds, coatings, and veins consistent with secondary mineralization. The rock textures are consistent with a volcanic origin and subsequent alteration and/or weathering by impact events, wind, and possibly water. PMID- 15297664 TI - Magnetic properties experiments on the Mars exploration Rover Spirit at Gusev Crater. AB - The magnetic properties experiments are designed to help identify the magnetic minerals in the dust and rocks on Mars-and to determine whether liquid water was involved in the formation and alteration of these magnetic minerals. Almost all of the dust particles suspended in the martian atmosphere must contain ferrimagnetic minerals (such as maghemite or magnetite) in an amount of approximately 2% by weight. The most magnetic fraction of the dust appears darker than the average dust. Magnetite was detected in the first two rocks ground by Spirit. PMID- 15297665 TI - Chemistry of rocks and soils in Gusev Crater from the alpha particle x-ray spectrometer. AB - The alpha particle x-ray spectrometer on the Spirit rover determined major and minor elements of soils and rocks in Gusev crater in order to unravel the crustal evolution of planet Mars. The composition of soils is similar to those at previous landing sites, as a result of global mixing and distribution by dust storms. Rocks (fresh surfaces exposed by the rock abrasion tool) resemble volcanic rocks of primitive basaltic composition with low intrinsic potassium contents. High abundance of bromine (up to 170 parts per million) in rocks may indicate the alteration of surfaces formed during a past period of aqueous activity in Gusev crater. PMID- 15297666 TI - Mineralogy at Gusev Crater from the Mossbauer spectrometer on the Spirit Rover. AB - Mossbauer spectra measured on Mars by the Spirit rover during the primary mission are characterized by two ferrous iron doublets (olivine and probably pyroxene) and a ferric iron doublet (tentatively associated to nanophase ferric iron oxide). Two sextets resulting from nonstoichiometric magnetite are also present, except for a coating on the rock Mazatzal, where a hematite-like sextet is present. Greater proportions of ferric-bearing phases are associated with undisturbed soils and rock surfaces as compared to fresh rock surfaces exposed by grinding. The ubiquitous presence of olivine in soil suggests that physical rather than chemical weathering processes currently dominate at Gusev crater. PMID- 15297667 TI - Initial results from the Mini-TES experiment in Gusev Crater from the Spirit Rover. AB - The Miniature Thermal Emission Spectrometer (Mini-TES) on Spirit has studied the mineralogy and thermophysical properties at Gusev crater. Undisturbed soil spectra show evidence for minor carbonates and bound water. Rocks are olivinerich basalts with varying degrees of dust and other coatings. Dark-toned soils observed on disturbed surfaces may be derived from rocks and have derived mineralogy (+/-5 to 10%) of 45% pyroxene (20% Ca-rich pyroxene and 25% pigeonite), 40% sodic to intermediate plagioclase, and 15% olivine (forsterite 45% +/-5 to 10). Two spectrally distinct coatings are observed on rocks, a possible indicator of the interaction of water, rock, and airfall dust. Diurnal temperature data indicate particle sizes from 40 to 80 microm in hollows to approximately 0.5 to 3 mm in soils. PMID- 15297668 TI - Basaltic rocks analyzed by the Spirit Rover in Gusev Crater. AB - The Spirit landing site in Gusev Crater on Mars contains dark, fine-grained, vesicular rocks interpreted as lavas. Pancam and Mini-Thermal Emission Spectrometer (Mini-TES) spectra suggest that all of these rocks are similar but have variable coatings and dust mantles. Magnified images of brushed and abraded rock surfaces show alteration rinds and veins. Rock interiors contain 95th percentile). Similar findings were obtained in an independent population, and biochemical studies indicated that most sequence variants in the low HDL-C group were functionally important. Thus, rare alleles with major phenotypic effects contribute significantly to low plasma HDL-C levels in the general population. PMID- 15297677 TI - Brood parasitic cowbird nestlings use host young to procure resources. AB - Young brood parasites that tolerate the company of host offspring challenge the existing evolutionary view of family life. In theory, all parasitic nestlings should be ruthlessly self-interested and should kill host offspring soon after hatching. Yet many species allow host young to live, even though they are rivals for host resources. Here we show that the tolerance of host nestlings by the parasitic brown-headed cowbird Molothrus ater is adaptive. Host young procure the cowbird a higher provisioning rate, so it grows more rapidly. The cowbird's unexpected altruism toward host offspring simply promotes its selfish interests in exploiting host parents. PMID- 15297676 TI - HLA and NK cell inhibitory receptor genes in resolving hepatitis C virus infection. AB - Natural killer (NK) cells provide a central defense against viral infection by using inhibitory and activation receptors for major histocompatibility complex class I molecules as a means of controlling their activity. We show that genes encoding the inhibitory NK cell receptor KIR2DL3 and its human leukocyte antigen C group 1 (HLA-C1) ligand directly influence resolution of hepatitis C virus (HCV) infection. This effect was observed in Caucasians and African Americans with expected low infectious doses of HCV but not in those with high-dose exposure, in whom the innate immune response is likely overwhelmed. The data strongly suggest that inhibitory NK cell interactions are important in determining antiviral immunity and that diminished inhibitory responses confer protection against HCV. PMID- 15297679 TI - Adoption. PMID- 15297680 TI - The challenge of neonatal mortality in India. PMID- 15297681 TI - Risk factors of threshold retinopathy of prematurity. AB - OBJECTIVE: To determine the risk factors which predispose to the development of threshold retinopathy of prematurity among patients of retinopathy of prematurity. METHODS: The ROP clinic records of a 3 year period were retrospectively studied to identify babies with threshold ROP (T-ROP) and sub threshold ROP (ST-ROP). Various antenatal and perinatal risk factors, neonatal morbidity and therapeutic interventions were compared between the 2 groups. RESULTS: Of the total of 108 babies, 55 had T-ROP and 53 had ST-ROP. On univariate analysis, packed cell transfusions for anemia, double volume exchange transfusions (DVET), number of DVET, ventilation, gestational age 25% in 8 (50%) patients with a decrease of >40% documented in 3 of these. No perceptible adverse effects were recognized. PMID- 15297688 TI - Bartsocas-Papas syndrome in a Pakistani family from Kuwait. AB - We report a rare case of Bartsocas Papas Syndrome, a lethal autosomal recessive type of Popliteal Pterigium syndrome, from a consaguineous Pakistani family who had typical anomalies of face, limbs and genitalia with additional peripheral pulmonary stenosis. Antenatal diagnosis and option for termination of pregnancy is advised. PMID- 15297689 TI - Malignant hypertension in a child with solitary functioning hydronephrotic kidney. AB - A child with solitary functioning hydronephrotic kidney and hypertensive encephalopathy is described. Removal of the contralateral dysplastic kidney failed to normalize his blood pressure. PMID- 15297690 TI - Hot water epilepsy - a report of three cases. AB - Hot water epilepsy (HWE) is a rare form of reflex epilepsy caused by bathing with hot water. In this paper, we describe three cases with hot water epilepsy. It occurs generally in children with normal psychomotor development and children continue to develop normally after seizure. HWE has usually a favorable prognosis by first avoiding lukewarm water and secondly using either intermittent oral prophylaxis with benzodiazepines or conventional AEDs. PMID- 15297691 TI - Griscelli syndrome - a case report. AB - Griscelli syndrome is a rare autosomal recessive disorder characterized by partial albinism with variable immunodeficiency. Silvery gray hair with large, clumped melanosomes on microscopy of hair shafts are diagnostic. The commonest complication leading to mortality includes lymphohistiocytic proliferation in various organs, including the brain. We present a child with classic clinical features and confirmatory findings of clumped melanosomes on microscopy of hair shaft. PMID- 15297692 TI - Juvenile idiopathic osteoporosis. AB - Juvenile idiopathic osteoporosis (JIO) is a very rare condition of primary bone demineralisation that presents in childhood. Most pediatricians are unfamiliar with this condition owing to the difficulty in recognition and a long list of differential diagnosis. We report an 8-year-old girl who presented with generalized severe osteoporosis. Diagnosis of JIO was made by excluding other common causes of osteoporosis in this age. PMID- 15297693 TI - Lymphomatous malformation of the chest wall. PMID- 15297694 TI - Townes-Brocks syndrome. PMID- 15297695 TI - Kayser-Fleischer ring. PMID- 15297696 TI - Achalasia with growth retardation. PMID- 15297697 TI - Congenital myotonic dystrophy: a rare cause of polyhydramnios. PMID- 15297699 TI - Diagnostic dilemma of cecal duplication. PMID- 15297698 TI - Laparoscopy in suspected Meckels diverticulum: negative nuclear scan notwithstanding. PMID- 15297700 TI - Cold chain system in Chandigarh during intensified pulse polio immunization 2001 2002. PMID- 15297701 TI - Juvenile dermatomyositis presenting with anasarca. PMID- 15297702 TI - Fine localization of Nefl and Nef3 and its exclusion as candidate gene for lens rupture 2(lr2). AB - Cataract causing lr2 gene is found in the CXSD mouse, which is a recombinant inbred strain of BALB/c and STS mice. For the process of positional cloning of lr2, several candidate genes were selected in the middle region of chromosome 14, but most of them were excluded by combination of recombination and homozygosity mapping. Components of neurofilament proteins, neurofilament light polypeptide (Nefl) and neurofilament3 medium (Nef3), were linked to D14Mit87 which was not separated from the lr2 locus in the homozygosity mapping. When the expression levels of Nefl and Nef3 in eyes were compared in CXSD and BALB/c mice, there were no differences in expression levels. The cDNA sequences of the two genes from CXSD, BALB/c and STS mice were subsequently compared. Several nucleotide differences in cDNA sequences were detected between the mice strains but the majority of the changes were silent mutations that did not alter the amino acids. The sole amino acid difference, E567K in the glutamate rich region of Nfm, between BALB/c and CXSD was found to be a simple genetic polymorphism because the same substitution existed in STS, a non-cataract mouse strain. Therefore we excluded Nefl and Nef3 from the candidate genes for lr2 based on expression and mutation analyses. PMID- 15297703 TI - Effects of pair housing on diurnal rhythms of heart rate and heart rate variability in miniature swine. AB - This study investigated the effects of pair housing on diurnal rhythms of heart rate and autonomic nervous activity in miniature swine. For this purpose, six adult Gottingen miniature swine were initially housed individually in an animal cage. Then, two of each swine were housed in a large cage together for 3 weeks. After that swine were separated into individual cages again. During this experimental procedure, electrocardiogram (ECG) was recorded with a Holter ECG recorder. Autonomic nervous activity was evaluated by power spectral analysis of heart rate variability. Heart rate and autonomic nervous activity clearly showed a diurnal rhythm in miniature swine housed in individual cages. When two swine were housed together, heart rate was significantly increased throughout the day and diurnal rhythm disappeared. Although these changes gradually recovered to basal levels, these parameters had not completely returned to basal levels even after 2 weeks. Heart rate was still higher than the initial level just after swine were re-housed in their own individual cages. Heart rate and autonomic nervous activity returned to basal levels about 2 weeks after re-housing. Further, heart rate in some swine decreased below their initial levels. These results suggest that it takes miniature swine at least 2 weeks to adapt to different circumstances. Furthermore, the power spectral analysis of heart rate variability can be used as a useful method in a study for answering controversial issues related to stress response. PMID- 15297704 TI - Establishment of an efficient BAC transgenesis protocol and its application to functional characterization of the mouse Brachyury locus. AB - Transgenesis using large DNA such as YAC or BAC has extended the range of applications in functional genomics. Here we describe an efficient BAC transgenesis protocol using a simple BAC DNA preparation method adopted from YAC DNA purification methods. This method allowed us to isolate BAC DNA from small scale culture of BAC-containing cells in sufficient quantity and purity for microinjection. More than 40 founders have been produced with linearized BAC DNA prepared by this method, and 85% of them contained intact BAC transgenes. In contrast, when circular BAC DNA was injected, an approximately three-fold reduction of transgene integration rate was observed and fewer intact transgene integrations were obtained. A line of transgenic mice carrying a 170-kb BAC clone generated in this way successfully rescued tail and embryonic lethality phenotypes of the mouse Brachyury (T) mutants, further demonstrating the utility of this method in functional analysis of the mouse genome. PMID- 15297705 TI - The normalization of guinea pig leukocyte fractions and lymphocyte subsets in blood and lymphoid tissues using a flow cytometric procedure. AB - Many hematological and immunological parameters remain unclear in the study of the guinea pig. In this study, we established the mean values of blood counts, the percentage of leukocyte fractions and lymphocyte subsets in blood and various lymphoid tissues of the guinea pig with a flow cytometric procedure using MIL4/SSC. The mean counts of WBC and RBC in the blood were lower, and MCV and MCH were higher than those of other rodents, resembling those of humans. Furthermore, the mean percentages of blood lymphocytes were smaller and that of granulocyte was larger than those of other rodents, resembling those of humans. We further established a flow cytometric procedure for lymphocyte subsets and clarified the mean percentages of T- and B-cells, CD4(+)-, CD8(+)- and MHC Class II(+)- T cells, and CD4(-)CD8 (-) T-cells. The latter were morphologically larger in cell size and cytoplasm than CD4(+)- plus CD8(+) T-cells, and this subset had a significantly higher percentage in newborn animals. Furthermore, the appearance of the MHC Class II(+) T-cell subset was suggested to be a marker of hyper activation of T-cells in BCG-immunized animals. Thus, both the novel flow cytometric procedure for leukocyte fractions and lymphocyte subsets, and the established normal values will be useful tools in studying guinea pigs as models of various diseases and biological phenomena. PMID- 15297706 TI - Standardized data and relationship between bone growth and bone metabolism in female Gottingen minipigs. AB - Minipigs have been studied as a model of osteoporosis. However, little information is available regarding their bone physiology. We established standardized bone data and investigated the relationship between bone growth and bone metabolism in female minipigs. Blood and urine samples were obtained from 53 female Gottingen minipigs, 3-76 months of age, for measurement of bone biomarkers (i.e., BAP, OC, NTX, and DPD). The lumbar vertebra and femur were excised to determine the growth plate condition, bone length, bone mineral content (BMC), and bone mineral density (BMD). High levels of bone biomarkers were observed during the initial period after birth, decreasing thereafter with age. Bone biomarkers were confirmed to be highly correlated with age (R(2) > 0.7). The growth plates of the lumbar vertebra and the femur began to close at 21 and 25 months of age, respectively, and closed completely at 42 months of age. Bone length increased rapidly before growth plate closure, and reached a peak at 21 and 28 months of age in the lumbar vertebra and the femur, respectively. The levels of BMC and BMD increased rapidly before growth plate closure, and continued to increase slowly until 76 months of age. A high negative correlation (-0.855 < r < -0.711, p<0.001) was confirmed between the bone biomarkers and the bone measurement data. These results indicate that the bone turnover velocity is consistent with the bone growth velocity in female Gottingen minipigs. PMID- 15297707 TI - Progression of coronary atherosclerosis relates to the onset of myocardial infarction in an animal model of spontaneous myocardial infarction (WHHLMI rabbits). AB - Recently, we developed myocardial infarction-prone WHHLMI rabbits from coronary atherosclerosis-prone WHHL rabbits (WHHLCA rabbits) by selective breeding. In this study, we examined the relation of atherosclerotic plaques to the onset of myocardial infarction. We examined myocardial lesions of 378 WHHL rabbits born between 1992 and 2000, and atherosclerosis lesions of 93 WHHLCA and 82 WHHLMI rabbits. The aortic lesions were evaluated as percent surface lesion area. The coronary lesions were evaluated as cross sectional narrowing using sections prepared at 500 or 1,000 microm intervals. Serum lipid levels were assayed with enzymatic methods. The cumulative incidence of fatal myocardial infarction between 11 and 35 months old was 90% in WHHLMI rabbits and 21% in WHHLCA rabbits, respectively. Selective breeding increased the serum cholesterol levels by about 200 mg/dl despite there being no changes in triglyceride levels. Aortic and coronary atherosclerosis progressed markedly in WHHLMI rabbits compared to WHHLCA rabbits. Especially, WHHLMI rabbits over 15 months old showed more than 90% cross sectional narrowing of the left circumflex arteries, main stem of the left coronary artery, and the origin portion of the right coronary artery. In addition, there were no gender differences in atherosclerotic lesions of both aortas and coronary arteries. In conclusion, the present study showed that marked progression of coronary atherosclerosis was probably associated with spontaneous development of myocardial infarction in WHHLMI rabbits. PMID- 15297708 TI - Response of cortical and cancellous bones to mild calcium deficiency in young growing female rats: a bone histomorphometry study. AB - The purpose of the present study was to clarify the differences in the alterations of cellular activities of osteoblasts and osteoclasts, mineralization, and bone mass in cortical and cancellous bones of young growing rats with mild calcium deficiency. Twenty female Sprague-Dawley rats, 6 weeks of age, were randomized by the stratified method into two groups with 10 rats in each group: 0.5% (normal) calcium diet group and 0.1% (low) calcium diet group. After 10 weeks of feeding, bone histomorphometric analysis was performed on cancellous bone of the proximal tibia as well as cortical bone of the tibial shaft. Calcium deficiency increased eroded surface (ES/bone surface [BS]) and the number of osteoclast (N.Oc/BS) with an increase in osteoblast surface (ObS/BS), but decreased bone formation rate (BFR/BS) in cancellous bone. However, cancellous bone volume was preserved, while cortical bone area was decreased as a result of decreased periosteal bone gain and enlargement of the marrow cavity. These results suggest that short-term mild calcium deficiency in young growing female rats increased bone resorption by increasing osteoclastic recruitment, and suppressed mineralization followed by increased osteoblastic recruitment in cancellous bone, but cancellous bone loss was counteracted through redistribution of calcium from cortical bone to cancellous bone. PMID- 15297709 TI - Zona-float method for separating mouse eggs from other cells. AB - We have developed a new method for separating mouse eggs from other cells, such as cumulus cells, using centrifugation with Percoll. Solutions of 45, 22.5, 11.3, and 5.6% Percoll were tested. With the 22.5% solution, 99% of whole eggs obtained by in vitro fertilization were collected from the upper part of the Percoll solution, and 98% of 2-cell embryos collected from these eggs developed to the blastocyst stage. Offspring were obtained after transfer of collected embryos to female mice. The greatest advantage of this method is that undamaged eggs are separated from other cells in one simple operation, regardless of the number of eggs. PMID- 15297710 TI - Effect of sugar alcohols on gut function and body composition in normal and cecectomized rats. AB - The effects of two sugar alcohols on feed utilization, digesta retention, gut fermentation and serum lipid profiles were compared in normal and cecectomized rats to examine the possibility of the cecectomized rat as an experimental animal with relevance to humans. Semi-purified diets containing no sugar alcohol, 7% sorbitol or 7% lactitol were fed to normal and cecectomized rats for 16 days. The digestibility of the crude fat and the compositions of the carcass dry matter and crude fat were significantly decreased by feeding sugar alcohols in both groups, but the effects were relatively higher in the cecectomized rats than in the normal rats. Diarrhea, faster transit times and shorter retention times of digesta were noted in the cecectomized rats fed sugar alcohols, while the inverse results were observed in the normal rats fed similar diets. The concentration of cecal organic acids was increased in the normal rats, whereas the concentration of colonic organic acids was decreased in the cecectomized rats fed sugar alcohols, compared with their corresponding control groups. The concentration of serum total cholesterol was decreased in both the normal and cecectomized rats fed diets containing sugar alcohols. The tendencies for diarrhea, faster digesta transit and reduced body fat induced by the fermentable materials in the cecectomized rat have good relevance to the parallel effects of fermentable materials in humans, suggesting the possibility of using the cecectomized rat as a model to study some of the physiological effects of sugar alcohols in humans. PMID- 15297711 TI - Effects of psychological stress on autonomic control of heart in rats. AB - The aim of this study was to examine the effects of psychological stress on autonomic control of the heart in rats. For this purpose, we evoked anxiety-like or fear-like states in rats by means of classical conditioning and examined changes in autonomic nervous activity using an implanted telemetry system and power spectral analysis of heart rate variability. Anxiety-like states resulted in a significant increase in heart rate (HR), low frequency (LF) power, and LF/HF ratio, with no change in high frequency (HF) power. Fear-like states resulted in a significant increase in HR and a significant decrease in HF power with no significant change in both LF power and LF/HF ratio, although LF/HF ratio increased slightly. These results suggest that autonomic balance becomes predominant in sympathetic nervous activity in both anxiety-like and fear-like states. These changes in rats correspond to changes which are relevant to cardiovascular diseases in humans under many kinds of psychological stress. Therefore, the experimental design of this study is a useful experimental model for investigating the effects of psychological stress on autonomic control of the heart in humans. PMID- 15297712 TI - Linkage mapping of the locus responsible for male pseudohermaphroditism (mp) on rat chromosome 7. AB - TF is a mutant rat strain showing male pseudohermaphroditism controlled by an autosomal single recessive gene (mp). The affected rats show lack of Leydig cells and androgen deficiency. In this study, we performed linkage analysis using F(2) progeny of crosses between TF and BN strains to determine the chromosomal localization of the mp locus. The mp locus was mapped in a 4 cM region of the distal region of rat chromosome 7 between D7Rat3 and D7Rat115 or D7Rat94. Comparison of the linkage map with corresponding regions of the published rat genome sequence revealed several candidate genes for the mp mutation, including the Dhh, Tegt, Gdp3, and Amhr2 genes. PMID- 15297713 TI - Successful retrograde transport of fluorescent latex nanospheres in the cerebral cortex of the macaque monkey. AB - Retrograde axonal transport of latex nanospheres offers a means of delivering chemical agents to a targeted region of the central nervous system (CNS). In this study we performed microinjections of latex nanospheres into the cerebral cortex of cynomolgus monkeys and observed successful retrograde labeling of neurons in the contralateral region. Our data indicate the successful use of this delivery system, reported in studies using other animals, may also be achievable with primates as well. PMID- 15297714 TI - Renin, cyclooxygenase-2 and neuronal nitric oxide synthase in the kidneys of transgenic Tsukuba hypertensive mouse. AB - The transgenic Tsukuba hypertensive mouse (THM), which expresses the human renin and angiotensinogen genes, develops hypertension secondary to increased renin angiotensin system activity. The aim of the present study was to assess expression of the renin, cyclooxygenase-2 (COX-2), and neuronal nitric oxide synthase (nNOS) proteins in THM kidneys by immunohistochemical stainings. Renin expression was decreased in the THM kidneys when compared to kidneys from heterozygotes or control mice. Although no differences were observed in nNOS expression, overexpression of the COX-2 protein was observed in the macula densa cells in THM kidneys. PMID- 15297715 TI - Comparative study on toxicokinetics of bisphenol A in F344 rats, monkeys (Macaca fascicularis), and chimpanzees (Pan troglodytes). AB - We compared the toxicokinetics of bisphenol A (BPA) among three animal species: rats, cynomolgus monkeys and chimpanzees. Rats and monkeys were administered BPA orally or subcutaneously at 10 or 100 mg/kg body weight, while chimpanzees were administered only 10 mg/kg of BPA. BPA in serum was measured by ELISA. In oral administration of BPA at 10 mg/kg, both C(max) and AUC were rats < chimpanzee < monkeys. In oral administration of BPA at 100 mg/kg, both C(max) and AUC were rats < monkeys. Subcutaneous BPA administrations also revealed similar results, although the values of toxicokinetic parameters in subcutaneous administration were higher than those in oral administration. These results suggest that orally or subcutaneously administered BPA in primates is more easily absorbed than that in rats. We conclude that there are considerable differences in distribution, metabolism, and excretion of BPA between rodents and primates. PMID- 15297716 TI - Refined porcine follicle stimulating hormone promotes the responsiveness of rabbits to multiple-ovulation treatment. AB - We investigated whether refined follicle stimulating hormone (FSH) with only a little contaminating LH can promote the responsiveness of rabbits to multiple ovulation treatment. One group of female rabbits was stimulated with refined porcine FSH (pFSH), an FSH source with low LH activity, and another group was treated with pFSH. The mean number of eggs recovered from donors stimulated with refined pFSH (27 +/- 3) was significantly greater (P<0.05) than that with pFSH (20 +/- 2). Furthermore, the mean number of remaining follicles of donors stimulated with refined pFSH (19 +/- 4) was significantly greater (P<0.05) than that with pFSH (12 +/- 1). To decrease the number of remaining follicles in donors treated with refined pFSH, the dose of human chorionic gonadotropin (hCG) was increased from 75 to 150. However, there were no differences in the numbers of eggs and remaining follicles. The results of the present study suggest that refined pFSH with little contaminating LH promotes the responsiveness of rabbits to multiple-ovulation treatment compared with pFSH. PMID- 15297717 TI - [Metabolites produced by actinomycetes--antiviral antibiotics and enzyme inhibitors]. AB - In 1995, we discovered new antiherpetic antibiotics, called fattiviracins. The producing organism was classified as a strain belonging to Streptomyces microflavus. The strain produced at least 13 fattiviracin derivatives (FV-1 to FV 13). Fattiviracins were obtained as a white amorphous powder, and their molecular weights are in the range of 1400 to 1500. They are readily soluble in water, methanol, pyridine, and DMSO, but insoluble in other organic solvents. Fattiviracins have macrocyclic diesters formed by the binding of two trihydroxy fatty acids and two D-glucose residues in the molecule, and they can be divided into five families according to the length of the fatty acid moiety. Fattiviracins have potent activity against enveloped DNA viruses such as the herpes family, HSV-1, and VZV and enveloped RNA viruses such as influenza A and B viruses, and three strains of HIV-1, with EC(50) values on the order of a few micrograms per milliliter. The biosynthetic pathway of fattiviracins is also becoming clearer. Using bacitracin-resistant strains, enhanced and astringent production of fattiviracin was achieved. Fattiviracin FV-13, which has the longest fatty acid chains in the molecule, was dramatically enhanced by a C(55) isoprenyl phosphate metabolism. In addition, we have screened various inhibitors of enzymes such as alkaline protease, glucosyltransferase, glucuronidase, phospholipase, deoxyribonuclease, DNA methyltransferase, and DNA topoisomerase. All the inhibitors we discovered are briefly summarized in this paper. PMID- 15297718 TI - [Studying chemistry of heterocyclic compounds--some rearrangement reactions of indoles and the related compounds]. AB - Indole alkaloids are one of the most interesting research fields. The chemistry of monoterpenoid indole alkaloids, in particular, offers many skeletal rearrangement reactions including terpenoid rearrangements. The author describes those compounds encountered or connected with in his research, including iridoids, indole alkaloids, adamantanes found in nature, and oxyindoles that are correlated with rearrangements. PMID- 15297720 TI - [A novel deprotonative functionalization of aromatics with phosphazene base]. AB - A novel type of deprotonative functionalization of aromatics was accomplished with a phosphazene base (t-Bu-P4 base). For various nitrogen heteroaromatics and benzene derivatives, deprotonative 1,2-additions proceeded with the t-Bu-P4 base and ZnI(2) as an additive in the presence of carbonyl compounds. The t-Bu-P4 base has both extremely strong Bronsted basicity and less nucleophilicity due to its huge, widely conjugated structure, and highly chemoselective deprotonations were achieved. In addition the nonmetallic t-Bu-P4 base should not function as a Lewis acid. Therefore the deprotonation with the t-Bu-P4 base is considered to proceed via a different pathway from traditional deprotonative metalation of aromatics with metallic bases. Some reactions with unique regioselectivities were observed. PMID- 15297719 TI - [Serotonin and anticancer drug-induced emesis]. AB - Cytotoxic drug-induced nausea and vomiting are the side effects most feared by cancer patients. Emesis is an instinctive defense reaction caused by the somato autonomic nerve reflex, which is integrated in the medulla oblongata. Emesis caused by anticancer drugs is associated with an increase in the concentration of serotonin (5-HT) (5-HT) in the intestinal mucosa and brainstem. 5-HT released from the enterochromaffin (EC) cells, which synthesize and secrete 5-HT, stimulates the 5-HT receptors on the adjacent vagal afferent nerves. The depolarization of the vagal afferent nerves stimulates the vomiting center in the brainstem and eventually induces a vomiting reflex. 5-HT released from EC cells appears to mediate the cisplatin-induced emesis sensitive to 5-HT(3) receptor antagonists. The precise role of 5-HT in the occurrence of vomiting has not been fully elucidated. The present review describes the role of 5-HT in anticancer drug-induced emesis from the viewpoint of 5-HT release and afferent vagal nerve activity. Various models and methods for predicting emesis are also evaluated. PMID- 15297721 TI - [Inhibitory effects of some traditional medicines on proliferation of HIV-1 and its protease]. AB - In attempts to discover anti-HIV agents from natural sources, various traditional medicine extracts were tested for their inhibitory effects on HIV-1 proliferation and its protease. An extract of the seeds of Croton tiglium showed potent inhibitory effects on the proliferation of HIV-1. The active principle was determined to be phorbol esters. Several derivatives of phorbol ester were evaluated for inhibition of proliferation as well as activation of protein kinase C (PKC). Of these compounds, 12-O-acetylphorbol 13-decanoate (6) showed the most potent inhibition of proliferation without activating PKC. Some triterpenes from the stems of Cynomorium songaricum and the woody part of Xanthoceras sorbifolia showed inhibitory activity against HIV-1 protease. Various derivatives of oleanolic acid were synthesized and evaluated for their inhibitory activity against HIV-1 protease. Their inhibitory mechanism was also investigated. PMID- 15297722 TI - [Creation of functional muteins using phage libraries for pharmacoproteomic-based drug discovery and development of DDS]. AB - Tumor necrosis factor-alpha (TNF-alpha) has been expected to be a promising new antitumor agent, but toxic side effects by the systemic administration of TNF alpha limit its clinical application. In this study, we attempted to improve the therapeutic potency of TNF-alpha by using our protein-drug innovation systems. Among phage libraries displaying various mutant TNF-alphas, we isolated some lysine-deficient super mutant TNF-alphas, typified by mTNF-alpha-K90R, with higher TNF-receptor affinities and stronger bioactivity in vitro, in spite of the importance of lysine residues for trimer formation and receptor binding. mTNF alpha-K90R showed more than 10 times stronger in vivo antitumor effects and 1.3 times less toxicity than wild-type TNF-alpha (wTNF-alpha). Site-specifically mono PEGylated mTNF-alpha-K90R (sp-PEG-mTNF-alpha-K90R) at N-terminus showed higher in vitro bioactivity than unmodified wTNF-alpha, whereas randomly mono-PEGylated wTNF-alpha at a lysine residue (ran-PEG-wTNF-alpha) had less than 6% of the bioactivity of wTNF-alpha. The antitumor therapeutic window of sp-PEG-mTNF-alpha K90R was extended by about 5 times, 60 times and 18 times compared with those of mTNF-alpha-K90R, wTNF-alpha and ran-PEG-wTNF-alpha, respectively. sp-PEG-mTNF alpha-K90R may, thus, be a potential systemic anti-tumor therapeutic agent. These data suggested that our fusion protein-drug innovation system composed of a creation system of functional mutant proteins based on phage display technique and a site-specific PEGylation system may open up a new avenue to the optimal protein therapy. PMID- 15297723 TI - [Development of drug delivery system for intrathecal administration and its therapeutic effect on cerebral vasospasm and ischemia]. AB - To date, the pharmacologic approach to cerebral vasospasm and ischemia has been hampered in part by an inability to attain sufficiently high concentrations of drugs in the cerebrospinal fluid (CSF). To overcome this limitation of current drug therapy, we have developed a sustained-release preparation of the protein kinase inhibitor fasudil. Experimental cerebral vasospasm in rats and dogs was induced by double injection of autologous arterial blood into the cisterna magna. Focal cerebral ischemia in rats was induced by middle cerebral artery occlusion using an intraluminal suture technique. A single intrathecal injection of liposomal fasudil can maintain a therapeutic the drug concentration in the CSF due to the sustained-release property of liposomes, significantly decreasing intact size of acute ischemia and decreasing vasoconstriction of the basilar artery in cerebral vasospasm. This novel approach for the treatment of cerebral vasospasm and ischemia may have significant potential for use in the clinical setting. PMID- 15297724 TI - [Development of a new analgesic based on metabolism of endomorphin, an endogenous opioid peptide]. AB - Endomorphin-2 (Tyr-Pro-Phe-PheNH(2)) was discovered as an endogenous ligand for the mu-opioid receptor. The physiologic function of endomorphin-2 as a neurotransmitter or neuromodulator may cease after rapid enzymatic processing in the synapses of the brain, like other neuropeptides. The present study was conducted to examine the metabolism of endomorphin-2 by synaptic membranes prepared from mouse brain. Major metabolites were free tyrosine, free phenylalanine, Tyr-Pro, and PheNH(2). Both the degradation of endomorphin-2 and the accumulation of major metabolites were inhibited by specific inhibitors of dipeptidyl peptidase IV, such as diprotin A and B. On the other hand, the accumulation of Phe-PheNH(2) and Pro-Phe-PheNH(2) was increased in the presence of bestatin, an aminopeptidase inhibitor, whereas that of free phenylalanine and PheNH(2) was decreased. Furthermore, purified dipeptidyl peptidase IV hydrolyzed endomorphin-2 at the cleavage site, the Pro(2)-Phe(3) bond. Thus degradation of endomorphin-2 by brain synaptic membranes appears to occur mainly through the cleavage of the Pro(2)-Phe(3) bond by dipeptidyl peptidase IV, followed by the release of free phenylalanine and PheNH(2) from the liberated fragment, Phe PheNH(2), by aminopeptidase. We have also examined the effects of diprotin A on the antinociception induced by intracerebroventricularly administered endomorphin 2 in the mouse paw withdrawal test. Diprotin A simultaneously injected with endomorphin-2 enhanced endomorphin-2-induced antinociception. These results indicate that dipeptidyl peptidase IV may be an important peptidase responsible for terminating endomorphin-2-induced antinociception at the supraspinal level in mice. These findings also suggest that selective dipeptidyl peptidase IV inhibitors or dipeptidyl peptidase IV-resistant endomorphin-2 analogues have the potential for the clinical use as analgesics. PMID- 15297725 TI - [A systematic review of the clinical effectiveness of azathioprine in patients with ulcerative colitis]. AB - To clarify the effectiveness and safety of azathioprine (AZA) and 6 mercaptopurine (6MP) in the induction and maintenance of remission in ulcerative colitis (UC) by using a systematic review of published studies. Studies were searched for from within the 1966 to March 2003 MEDLINE database, Cochrane Library 2003 issue 1, and the 1981 to March 2003 Japana Centra Revuo Medicina database. References from published studies and reviews were also obtained. Randomized, placebo-controlled trials of oral AZA or 6MP therapy in adult patients with active or quiescent UC were included. Ratios for the induction and maintenance of remission, the steroid-sparing effect, and the incidence of adverse drug reactions (ADRs) were compared and evaluated between the two study arms and expressed by the odds ratio (OR) specific for the individual studies and the meta-analytic summary for the OR. We could find no randomized controlled trial for 6MP therapy. However, four clinical trials for AZA therapy were included in this meta-analysis. For the induction of remission, the pooled OR of the response to AZA therapy compared with placebo in active UC was 1.45 (95% Confidence Interval (CI): 0.68 to 3.08). For the maintenance of remission, the pooled OR for AZA therapy was 2.26 (95% CI: 1.27 to 4.01). The number needed to treat (NNT) to prevent one recurrence was 6 patients. The pooled OR for AZA therapy's ADRs compared with placebo was 2.11 (95% CI: 0.92 to 4.84). From the viewpoint of effectiveness and safety, this meta-analysis suggests that AZA might be useful in the maintenance of remission in UC patients. PMID- 15297726 TI - [The problems of multiple-chemical sensitivity patients in using medicinal drugs]. AB - Multiple-chemical sensitivity (MCS) patients are presumed to be compelled to lead inconvenient and difficult lives, because unpleasant and multiorgan symptoms are caused by very small amounts of various chemicals in the living environment. Therefore we conducted a questionnaire survey of MCS patients who are members of support groups to elucidate the problems of MCS patients in using medicinal drugs. In this report, we selected 205 persons who stated that they had been "diagnosed with MCS by a physician" or "a physician suspected a diagnosis of MCS" on the questionnaire as the reason they judged themselves to have MCS. The questionnaire results showed that about 60% of MCS patients have difficulty in using medicinal drugs and that the difficulties are more likely to occur in women, in people 40-59 years old, and in patients who developed MCS in reaction to pesticides or medicinal drugs. The prescribed drugs and OTC drugs noted as usable or unusable by patients in the questionnaire were analyzed from the viewpoint of their medicinal constituents. The results indicated that lidocaine is likely to be unusable by MCS patients. In addition, caffeine, aspirin, chlorphenylamine maleate, minocycline hydrochloride, levofloxacin, etc. were also likely to be unusable by MCS patients. Many patients who recorded drugs containing the above-mentioned remedies as unusable had a past history of allergy, suggesting that allergy is involved in the difficulties of MCS patients in using medicinal drugs. PMID- 15297727 TI - Protective effects of barley and its hydrolysates on gastric stress ulcer in rats. AB - This research intends to clarify the protective effect of barley and its hydrolysates with respect to a water immersion stress-induced ulcer in the rat model. The beta-(1-->3)-glucan content of barley, and specifically beta-(1- >4),(1-->3)-glucan content was determined and then gastric stress ulcerogenesis induced by water immersion was conducted using five-week-old male Sprague-Dawley rats (7 rats in one group). The barley diet group was fed 10% barley flour that was substituted with sucrose in the control diet. For the 3 groups fed on soluble dietary fiber (SDF), the diets were supplemented with 0.46 g of SDF, equivalent to 100 g of the control diet; 0.46 g of SDF is equivalent to 10 g of barley flour. The rats were housed in a stress-cage and immersed in a water bath (23 degrees C) up to their necks for 21 h. The content of SDF and beta-(1-->3)-glucan content in barley flour were 4.6% and 3.4%, respectively. Although strongly anti ulcer activities were observed in the barley (10%), SDF isolated and beta-(1-->3) glucan fraction (Hydrolysate I) prepared from barley flour after treatment with lichenase, in other words, beta-(1-->4),(1-->3)-glucan itself, its hydrolysate (Hydrolysate II) with beta-(1-->3)-glucosidase did not display any anti-ulcer activity. This finding suggests that the beta-(1-->3)-glucosyl-linkage on beta-(1 ->3)-glucan is an important part of the active principle for anti-ulcerogenesis. PMID- 15297728 TI - Understanding glial differentiation in vertebrate nervous system development. AB - Recent progresses in molecular and developmental biology provide us a good idea how differentiation of glial cells in the vertebrate nervous system is regulated. Combinations of positional cues such as secreted proteins and cell-intrinsic mechanisms such as transcription factors are essential for the regulation of astrocyte and oligodendrocyte differentiation from the neural epithelium. In contrast, regulatory mechanisms of glial differentiation from neural crest derived cells in the peripheral nervous system are less understood. However, recent studies suggest that, at least in part, the peripheral gliogenesis is regulated by mechanisms, such as Notch signaling, that are also important for the gliogenesis in the developing central nervous system. PMID- 15297729 TI - Exercise-related time course of pulsatility index in brachial artery following forearm exercise assessed by Doppler ultrasound. AB - At rest, vascular reactivity assessed by the changes in pulsatility index (PI) is one indicator of vessel stenosis in some clinical/basic science research. However, all types of vessel stenosis do not show an alteration in the PI, because flow perfusion may be maintained by the development of collateral vessels such as in severe arterial stenosis or non-severe arterial stenosis. Therefore at rest, changes in the PI may not always be a precise indicator of vessel stenosis. However, a few studies have used the PI following exercise, which may provide additional information on hemodynamics. The purpose of the present study was to examine the exercise-related time course of the PI in the brachial artery after ischemic or non-ischemic isometric handgrip exercise (IHE) using Doppler ultrasound, and to determine the potential use of this parameter as an indicator of vascular disease. Ten healthy young male subjects performed IHE at 10% and 30% of maximum voluntary contraction (MVC) for 2-minutes (min) with or without arterial occlusion (AO), or 2-min of AO alone. Following each 2-min session, PI was determined during the 5-min recovery period. A significant difference in the recovery PI was observed between IHE, ischemic IHE, as well as AO alone. Exercise with AO significantly increased the reduction in the PI compared to exercise alone, or AO alone, at both 10% and 30%MVC. These results suggest, exercise induced changes in the time course of the PI during recovery may potentially be a useful diagnostic tool. Exercise-induced ischemic state may potentially be a useful indicator for detecting arteriovascular disease, even if it is not detected by AO alone. PMID- 15297730 TI - The effects of hyperbaric oxygen treatment on oxidant and antioxidants levels during liver regeneration in rats. AB - The effects of hyperbaric oxygen (HBO) therapy on oxidant/antioxidant metabolism are controversial and its effects on hepatic regeneration are not known. In this study, we investigated a possible beneficial effect of HBO therapy on oxidant and antioxidants levels during liver regeneration. To conduct this study, seventy percent hepatectomy was performed on forty-eight Spraggue-Dawley rats and the rats were divided into two equal groups: HBO-treated group and untreated group (non-HBO group). We determined the levels of malondialdehyde (MDA), an oxidative stress marker, and the levels of antioxidant enzymes/reagents, including glutathione (GSH), superoxide dismutase (SOD) activity, copper (Cu) and zinc (Zn), in the remnant liver samples. We also measured mitotic index (MI) and proliferating cell nuclear antigen (PCNA) levels to assess the degree of liver regeneration. HBO treatment significantly decreased MDA levels, whereas it increased SOD activity, GSH and Zn levels. In contrast, Cu levels were lower in the HBO-treated livers than the levels in the untreated remnant livers. The effect of HBO treatment may be mediated by the suppression of certain enzymes that are responsible for lipid peroxidation. In addition, HBO treatment may induce the production of antioxidant enzymes/reagents by remnant liver tissues. The HBO-treated rats maintained their body weights but the untreated rats lost body weights. HBO treatment also increased MI and PCNA levels, indicating HBO treatment enhances liver regeneration. These results indicate that HBO treatment has beneficial effects on liver regeneration by decreasing MDA and by increasing antioxidant activities. We therefore suggest that HBO therapy may be useful after liver resection. PMID- 15297731 TI - Lung injury after aortic occlusion-reperfusion in rats: the role of gadolinium chloride. AB - Aortic ischemia-reperfusion (AIR) induced lung injury has already been documented. Kupffer cell blockage (KCB) with gadolinium chloride (GdCl3) has also been shown to attenuate remote organ damage caused by ischemia reperfusion. The present study was designed to examine the effect of GdCl3 in lung ischemia reperfusion injury induced by aortic occlusion. Thirty-two rats were randomly allocated to four groups as follows: SHAM (Sham Laparotomy), SHAM+KCB, AIR, and AIR+KCB. An atraumatic microvascular clamp was placed across the infrarenal abdominal aorta just after its origin from the aorta for 30 minutes. The microvascular clamp on the infrarenal abdominal aorta was removed and reperfused for 60 minutes. GdCl3 was given 24 hours prior to the experiment. Malondialdehyde (MDA) level and myeloperoxidase (MPO) activity were assayed in lung tissues. MDA level and MPO activity in the AIR group were significantly higher than those in the other groups. When compared to AIR group, KCB with GdCl3 significantly decreased MDA level and MPO activity in the AIR+KCB group. These results suggest that GdCl3 attenuates the lung injury caused by AIR. The effects of GdCl3 on reduced lung damage may be mediated through significant decreases in both MDA level and MPO activity. PMID- 15297732 TI - Effects of life away from home and physical exercise on nutrient intake and blood/serum parameters among girl students in Japan. AB - This study was initiated to examine if the life away from home and participation in sport activities affect nutritional health among girl university students. For this purpose, anthropometric data, peripheral blood and spot urine samples, 24 hour food duplicate samples, and answers to questionnaires were collected from 71 girl students at 19 to 23 years of ages who provided informed consent to participate in the study. Of the 71 participants, 29 and 42 participants lived in their homes or outside, respectively, and 23 subjects participated in sport activities whereas 48 subjects did not. Hematology, serum biochemistry and nutrient intakes were evaluated in comparison with the life conditions (home vs. dormitory, boarding house, etc.) and participation in sport activities. The population studied had insufficient intake of energy, protein, and minerals such as Ca and Fe. Those who lived in home or participated in sport activities took more energy and protein (although not the two minerals) than others. Skipping of breakfast was more common among those who lived away from home and had no sport activity. Thus, two social factors of life in home and participation in sport clubs contribute favorably for better food habits, but not necessarily improved intakes of Ca and Fe. PMID- 15297733 TI - Chymase is activated in the pulmonary inflammation and fibrosis induced by paraquat in hamsters. AB - The involvement of chymase has been implicated in fibrotic response to tissue injuries. Besides its direct action, chymase indirectly promotes fibrotic response by generating angiotensin (Ang) II from Ang I. In the present study, we examined whether chymase and angiotensin converting enzyme (ACE), that also generates Ang II, were activated in the pulmonary inflammation and fibrosis induced by paraquat (PQ) in hamsters. In an acute PQ intoxication group, PQ was administered subcutaneously and the lungs were excised four days after administration. In a chronic PQ intoxication group, PQ was administered at the same dose once a week for six weeks. The lungs were excised five weeks after the last administration. On dissection, alveolar hemorrhage and capillary stasis were found in the acute PQ intoxication group while interstitial pulmonary fibrosis was found in the chronic PQ intoxication group. The pulmonary tissue chymase activity was elevated in both intoxication groups when compared with a respective age-matched, vehicle (saline)-administered group. Pulmonary tissue ACE activity, on the other hand, decreased in both intoxication groups. These data suggested that activated chymase may be involved in the establishment of PQ-induced pulmonary fibrosis in hamsters. PMID- 15297734 TI - Characterization of a Shiga toxin 1-neutralizing recombinant Fab fragment isolated by phage display system. AB - Shiga toxin (Stx)-producing Escherichia coli (STEC) causes bloody diarrhea, hemorrhagic colitis, and hemolytic uremic syndrome. Molecular structural analysis of Stx-neutralizing monoclonal antibody (mAb) will be helpful for development of therapeutics and prophylactics for STEC infection. In this study, we cloned the genes of Stx 1-neutralizing mAb, termed 5-5B from hybridoma cells by phage display system and characterized its recombinant Fab (rFab) fragment. 5-5B rFab fragment reacted with Stx1, but not with Stx2 and bovine serum albumin (BSA). It also showed the neutralizing activity against the cytotoxicity of Stx1. These results imply that 5-5B rFab fragment is functionally identical to parent mAb. The variable heavy (VH) and light domains were found to be highly homologous with the derived germ line sequences. As for VH domain, the complementarity determining regions (CDRs) showed higher replacement/substitution mutation ratio than that in the frame work regions. Among the regions, CDR2 showed the most frequent nucleotide and amino acid substitutions. These results suggest that heavy chain CDR2 may mainly be associated with the 5-5B function, that is neutralizing cytotoxicity of Stx1. PMID- 15297735 TI - An instrument capable of grading visual function: results from patients with retinitis pigmentosa. AB - There was no device to grade visual function in patients with retinitis pigmentosa (RP). We have therefore developed an instrument capable of measuring and quantifying the visual capabilities, and here present the results from patients with RP. In total, 118 eyes of 59 patients, 26 men and 33 women, with RP were studied. Seven eyes had hand movement (HM) and eight had light perception (LP) vision, and the others had better visual acuity. The Low Vision Evaluator (LoVE) consists of a pair of goggles with white, light-emitting diodes as the stimulus, a control box, an on-off button to signal the detection of the stimulus, and a printer for permanent records. There are 15 luminance levels of stimuli (combination of 5 intensities and 3 durations). The stimuli are delivered in a random sequence with an audio signal presented 0.3 seconds prior to the light stimulus. Each eye was tested separately, and each stimulus magnitude (intensity x duration) was presented 3 times for a total of 27 stimuli per eye. With 6 catch trials (audio signal without a light stimulus), a total of 60 trials were examined in a full examination. The conventional visual acuity and kinetic visual fields were determined. 59 patients had different visual acuities that ranged from no light perception (NLP) to 1.5 vision, and visual field sizes that ranged from 0.0001 to 3.96 steradians. The visual acuity and visual field size were significantly correlated with the LoVE score (r=0.58 and 0.64, respectively; p<0.01). These results indicate that the LoVE is capable of grading the visual function of RP patients with various visual acuities and visual fields. The testing procedures are simple for the patient and examiner, and this instrument can be used to assess the effectiveness of medical and surgical therapy. PMID- 15297736 TI - Upregulation of retinoic acid-inducible gene-I in T24 urinary bladder carcinoma cells stimulated with interferon-gamma. AB - Urinary bladder epithelial cells play an important role in the host defense against urinary tract infections. Interferon-gamma (IFN-gamma) is a potent cytokine that regulates immune responses by inducing multiple genes in many types of cells including urinary bladder epithelial cells. Retinoic acid-inducible gene I (RIG-I) is a member of the DExH-box family, which is involved in various reactions related to RNA metabolism, and is induced in leukemic cells by retinoic acid or in endothelial cells by lipopolysaccharide. We have studied the expression of RIG-I in T24 cells, a cell line derived from human urinary bladder epithelial carcinoma cells. IFN-gamma stimulated T24 cells to express RIG-I mRNA and protein in concentration- and time-dependent manners. Immunohistochemical analysis revealed the expression of RIG-I in the urinary bladder epithelium from a patient with chronic urinary tract infection and in a bladder epithelial carcinoma. We conclude that RIG-I may play some role in inflammatory reactions in the urinary tract epithelium. PMID- 15297737 TI - Metabolism of pyrogallol to purpurogallin by human erythrocytic hemoglobin. AB - The aim of this study was to investigate the oxido-reductive reactions of human hemoglobin with pyrogallol and the metabolism of pyrogallol by the protein, which contains a protoporphyrin IX like cytochrome P-450. Pyrogallol, having three hydroxy groups at the adjacent positions in the benzene ring, oxidized human oxyhemoglobin to methemoglobin and reduced human methemoglobin to oxyhemoglobin. Since superoxide dismutase and catalase inhibited these reactions extensively, active oxygens such as superoxide and hydrogen peroxide were considered to be involved in the oxido-reductive reaction of human hemoglobin by pyrogallol. It was also found that the metabolism of pyrogallol to purpurogallin occurred quickly in human erythrocytes, i.e., when pyrogallol was added to human erythrocyte suspension, it oxidized intracellular hemoglobin and produced purpurogallin. The metabolism of pyrogallol to purpurogallin was explained by the pyrogallol oxidation with superoxide and hydrogen peroxide produced during the oxido-reductive reactions of human hemoglobin with pyrogallol. The present results show that human erythrocytes can metabolize pyrogallol, suggesting that the cells may be involved in the metabolism of some drugs in the human body. PMID- 15297738 TI - Development and construct validation of the Korean competence scale (KCS). AB - A comprehensive Korean competence scale (KCS) designed for Korean older adults, was developed and its construct validity evaluated. KCS assesses their higher levels of functional competence than activities of daily livings (ADL), necessary for independence in the home and community. The internal structure of the scale was tested using an exploratory and confirmatory factor analysis with two elderly Korean sample groups. As expected, the exploratory factor analysis result demonstrated a three-factor solution comprised of the following three domains: instrumental self-maintenance, intellectual activity, and social role. The confirmatory factor analysis results showed that an acceptable solution could be estimated for a second-order factor model comprised of these factors. These findings provide evidence for the construct validity of the instrument and have implications for future research on factorial invariance. PMID- 15297739 TI - Relationship between human cytomegalovirus glycoprotein B genotype and serum alanine aminotransferase elevation in infants. AB - The glycoprotein B (gB) region of the human cytomegalovirus (HCMV) is a major envelope glycoprotein that is a principal target of neutralizing antibodies and is known to stimulate the immune response of cytotoxic T lymphocytes. HCMV is currently classified into four genotypes on the basis of the nucleotide sequence of the gB region. The presence of HCMV in patients under 3 years of age was determined by subjecting urine samples taken from the patients to polymerase chain reaction (PCR) analysis. Analysis by direct sequencing of the gB region was carried out in 90 cases. These cases were grouped into the gB genotype 1 and gB genotype 3. Of 28 cases with a peak serum alanine aminotransferase (ALT) level(> or =100 IU/l), the duration of observed serum ALT elevation in the gB genotype 1 patients (166.7+/-126.7 days [mean+/-S.D.] [19 cases]) was significantly longer than that in the gB genotype 3 patients (39.7+/-31.7 days [9 cases]) (p<0.01). In the 54 cases with a serum ALT level(> or =50 IU/l), similar tendency was seen (p<0.05). These findings suggest that when serum ALT elevation is confirmed in young children infected with HCMV, analysis of the gB region is helpful for prediction of the duration of serum ALT elevation in the early stage of infection. PMID- 15297740 TI - Primary cutaneous mucinous carcinoma initially diagnosed as metastatic adenocarcinoma. AB - The authors report a rare case of primary mucinous carcinoma of the skin initially diagnosed as a metastatic adenocarcinoma. The tumor occurred in the right axilla in a 75-year-old man. Initial pathological diagnosis was metastatic adenocarcinoma. However, no primary focus was found in the body. The revised diagnosis by the authors was primary cutaneous mucinous carcinoma. The tumor (1.5 cm) was characterized by proliferation of atypical epithelial cells arranged in cell nests with many pseudolumens resembling adenoid cystic carcinoma. It was also characterized by much mucinous stroma or pool around tumor cells. No apparent eccrine or apocrine differentiation was noted histologically and immunohistochemically. The present case suggests that primary cutaneous mucinous carcinoma may be misdiagnosed as metastatic adenocarcinoma, and that it may resemble adenoid cystic carcinoma. PMID- 15297741 TI - Maternal plasma hypoxanthine levels in nonpreeclamptic twin pregnancies. AB - We have observed that the elevated plasma adenosine levels are associated with hyperuricemia in nonpreeclamptic twin pregnancies. In animal models, extracellular adenosine is taken up by cells to form adenine nucleotides or is degraded to other purine metabolites such as hypoxanthine, which is further metabolized to xanthine and uric acid. In this study, we measured plasma hypoxanthine levels to evaluate the role of adenosine in hyperuricemia among women with twin pregnancies. Maternal blood samples were taken in 13 twin and 20 singleton pregnancies at 35-36 weeks' gestation. The average maternal plasma hypoxanthine level in twin pregnancies was significantly higher than that in singleton pregnancies. In addition, the plasma hypoxanthine levels have positive correlations both with plasma adenosine and serum uric acid levels. Our results support that an increased adenosine is the main factor contributing to hyperuricemia in twin pregnancies. PMID- 15297742 TI - Supplementation of antioxidants prevents oxidative stress during a deep saturation dive. AB - Conflicting views exist at the present regarding the influences of a deep saturation dive on liver function in divers. Therefore, we first reevaluated whether a deep saturation dive (400 msw) induces a hepatic disturbance. As the result, plasma activities of both transaminases (aspartate aminotransferase [AST] and alanine aminotransferase [ALT]) increased significantly, whereas cholinesterase (Ch-E) activity decreased markedly, being highly suggestive of liver dysfunction. Assuming that the liver dysfunction was attributable to oxidative stress, we next examined the effects of supplementation of antioxidants (600 mg of vitamin C, 150 mg of alpha-tocopherol, and 600 mg of tea catechins per day) on liver function in saturation divers. As was anticipated, the antioxidants taken appeared to prevent a hepatic disturbance, indicating that a deep saturation dive provokes liver dysfunction probably due to oxidative stress. Thus, we recommend that saturation divers should take supplements of antioxidants. PMID- 15297743 TI - Genetic and phylogenetic analysis of glycoprotein of rabies virus isolated from several species in Brazil. AB - Genetic and phylogenetic analyses of the region containing the glycoprotein (G) gene, which is related to pathogenicity and antigenicity, and the G-L intergenic region were carried out in 14 Brazilian rabies virus isolates. The isolates were classified as dog-related rabies virus (DRRV) or vampire bat-related rabies virus (VRRV), by nucleoprotein (N) analysis. The nucleotide and amino acid (AA) homologies of the area containing the G protein gene and G-L intergenic region were generally lower than those of the ectodomain. In both regions, nucleotide and deduced AA homologies were lower among VRRVs than among DRRVs. There were AA differences between DRRV and VRRV at 3 antigenic sites and epitopes (IIa, WB+ and III), suggesting that DRRV and VRRV can be distinguished by differences of antigenicity. In a comparison of phylogenetic trees between the ectodomain and the area containing the G protein gene and G-L intergenic region, the branching patterns of the chiropteran and carnivoran rabies virus groups differed, whereas there were clear similarities in patterns within the DRRV and VRRV groups. Additionally, the VRRV isolates were more closely related to chiropteran strains isolated from Latin America than to Brazilian DRRV. These results indicate that Brazilian rabies virus isolates can be classified as DRRV or VRRV by analysis of the G gene and the G-L intergenic region, as well as by N gene analysis. PMID- 15297744 TI - Age-related changes in bone mineral density, cross-sectional area and the strength of long bones in the hind limbs and first lumbar vertebra in female Wistar rats. AB - Age-related changes in bone mineral density (BMD) and cross-sectional area and bone strength index (SSI) of the femur, tibia, humerus, and first lumbar vertebra in female Wistar (WM/MsNrs) rats were examined by a quantitative computed tomography (pQCT) method. One hundred and sixteen virgin female Wistar (WM/MsNrs) rats aged 2-33 months were used. The data indicate that the total BMD values of metaphyses and diaphyses of long bones increased until 12 months, then decreased to a varying degree depending on the bone after 15-24 months, but the values of cortical and trabecular BMD with age were not always similar to the total BMD value. Nevertheless, the values for cross-sectional area and SSI in the long bones increased regardless of the total BMD decrease with age, indicating that this increase might have been due to a characteristic of the modeling pattern in rats. The total and cortical BMD values in the first lumbar vertebra decreased after 18 months, and SSI did after 15 months. The data obtained in this study were compared with those obtained from males in a previous study. In conclusion, it was indicated that in this strain the rats over 12 months with the highest total BMD values in the femur and tibia, and before the onset of various tumors, are useful as a model animal for osteoporosis experiments and observation of senile bone change. PMID- 15297745 TI - Comparative anatomical study on the relationships between the vestigial pelvic bones and the surrounding structures of finless porpoises (Neophocaena phocaenoides). AB - Morphology of the modern cetaceans represents the results of adaptation of the ancestral terrestrial mammals to aquatic life through their evolutional processes. Some of the primitive fossil cetaceans are known to have both fore and hind limbs, whereas the pelvic bones of modern cetaceans are, in general, a pair of slender rod-like structures within the abdominal wall muscles just anterior to the anus with no articulations to the axial skeleton in both sexes. It is interesting and important to consider the causes and processes of how the hind limbs were lost and how the pelvis was reduced during the process of adaptation. In the present study, we tried to evaluate the topography and function of rudimentary pelvic bones of the finless porpoise (Neophocaena phocaenoides), one of the members of the odontocete cetaceans, with special references to the structures around the pelvic bones. Some soft tissues such as M. ischiocavernosus relating to the pelvic bone are transformed following the drastic reduction of the pelvis. This transformation tells us that the cetaceans adapted to the aquatic life during evolutional processes chose the tail flukes driven by the powerful trunk muscles for locomotion, instead of modifying the hind limbs into hind flippers as seen in pinnipeds. On the other hand, it is evident that a function of the pelvic bones of the male finless porpoise was supporting the penis as those of terrestrial mammals. It is noteworthy that the morphological features of the ancestral terrestrial mammals can be traced when they are carefully compared with those of the finless porpoise. PMID- 15297746 TI - Morphological variations in brachial plexus of beagle dogs: evaluation of utility as sources of allogeneic nerve grafts. AB - Basic studies were carried out to apply frozen allogeneic nerve grafts in dogs after wide-ranging defects of the brachial plexus due to surgical resection of tumor. In this study, morphological variations in branching patterns of the brachial plexus were examined in ten beagle dogs, to evaluate whether the brachial plexus might represent a useful source of allogeneic nerve grafts. Spatial relationships between the axillary lymph node, which had the possibility of carcinomatous metastasis, and the musculocutaneous (MC) nerve, which was important for the function of the forelimbs, were also investigated. In all ten cases examined, the brachial plexus received ventral roots from the fifth cervical nerve to the first thoracic nerve. No significant variation in the branching pattern was found in any nerve except the phrenic, MC and dorsal thoracic nerves. Four communicating branches were observed and had some morphological variations which might be negligible for nerve grafting. Considering previous physiological and anatomical reports, the most important nerve to be reunited in graft operations for functional recovery is the radial nerve. The MC nerve and median or ulnar nerve should also be considered as possibilities for reuniting. Distances between the axillary lymph nodes and the MC nerve ranged from 11.2 mm to 21 mm (mean +/- SD: 16.1 +/- 2.3 mm). In conclusion, it was suggested that morphological variations in the brachial plexus were technically acceptable to apply allogeneic nerve grafts at least in beagle dogs. PMID- 15297747 TI - A possibility of application of the 105-kilodaltons protein of Brachyspira alvinipulli cross-reacting with antisera to five species in the genus Brachyspira to diagnosis. AB - The antigenic properties of Brachyspira (B.) alvinipulli ATCC 51933 and strain C2 were analyzed and compared with those of B. hyodysenteriae ATCC 27164 and ATCC 31212, B. pilosicoli ATCC 51139, B. innocens ATCC 29796 and B. aalborgi NCTC 11492. In gel immunodiffusion tests, a protein in B. alvinipulli ATCC 51933 reacted strongly with anti-B. alvinipulli ATCC 51933-serum and formed two precipitin lines. Furthermore, by an immunoblotting technique, the 105 kilodaltons (kDa) protein in B. alvinipulli ATCC 51933 reacted strongly with each of the antisera to B. hyodysenteriae, B. pilosicoli, B. innocens and B. aalborgi. Therefore, the 105-kDa protein could be applied to diagnosis of chicken infection by B. alvinipulli and B. pilosicoli. But the 105-kDa protein reacting with the anti-B. alvinipulli ATCC 51933-serum was not confirmed in B. hyodysenteriae, B. pilosicoli, B. innocens and B. aalborgi. The N-terminal amino acid sequence of the 105-kDa protein isolated from B. alvinipulli ATCC 51933 was Met-Lys-Lys-Met Val-Tyr-Phe-Phe-Gly-Asn. The amino acid alignment of this protein possessed 50% homology with the periplasmic-iron-binding protein BitC in B. hyodysenteriae. PMID- 15297748 TI - A three-year study of enterohemorrhagic Escherichia coli O157 on a farm in Japan. AB - A long-term study was performed on the prevalence of enterohemorrhagic Escherichia coli (EHEC) O157 in bovine faeces. The present study was conducted on heifers raised on a farm showing a high isolation rate of EHEC O157 in previous years. The prevalence of EHEC O157 isolated from faecal samples was 10.6% (222/2104), 5.6% (181/3225), and 5.6% (153/2744) from 1998 to 2000, respectively. The numbers of EHEC O157-positive heifers for the same 3 years were 46.3% (185/400), 36.8% (147/399), and 31.7% (130/410), respectively. The seasonal prevalence of EHEC O157 varied according to the year. Most positive heifers excreted the EHEC O157 only once during the survey, though it was excreted 2 or 3 times by some heifers. The results obtained in the present study showed that the farm examined was heavily contaminated with EHEC O157. It is assumed that EHEC O157 does not remain in individual cattle long-term, but does exist long-term on farms due to repeated infection. PMID- 15297749 TI - Epizootiologic survey for Babesia microti among small wild mammals in northeastern Eurasia and a geographic diversity in the beta-tubulin gene sequences. AB - We previously reported that small wild rodents in Japan harbor two types of novel Babesia microti-like parasites (designated as Hobetsu and Kobe types), but not the type commonly found in the northeastern United States (U.S. type) where human babesiosis is endemic. To determine whether these new types of parasites are distributed in places surrounding Japan, an epizootiologic survey was undertaken in three geographically distant areas in northeastern Eurasia; South Korea, Vladivostok in Russia, and Xinjiang in China. Blood samples were collected from a total of 387 animals comprising 24 species. DNAs extracted from the samples were tested by nested PCR targeting babesial nuclear small-subunit rRNA gene (rDNA), which revealed that small rodents harboring B. microti exist in all three survey areas. Sequence analysis showed that all PCR-positive samples had rDNA sequences virtually identical to that of U.S.-type B. microti. However, when beta-tubulin gene sequences were compared, evident geographic variations were seen. By use of primers specific for each of the beta-tubulin genes of Kobe-, Hobetsu-, and U.S. type parasites, a type-specific PCR was developed. Parasite with Hobetsu- or Kobe type sequence was not detected from any of the three survey areas. These findings suggest that U.S.-type B. microti is widely distributed among small wild mammals in temperate zones of not only North America, but also Eurasia, whereas that Hobetsu- and Kobe-type parasites may be uniquely distributed in Japan. PMID- 15297750 TI - Measurement of polynuclear aromatic hydrocarbons in canine lung after alkaline decomposition. AB - An alkaline decomposition method employing a KOH/alcohol solution was studied, and polynuclear aromatic hydrocarbons (PAHs) contained in particles remaining in canine lung were measured. As a result, BaA, BkF, BaP, and BghiP were found. By this method, PAHs extracted from the lungs of 32 dogs were 13.0-166.0 ng (mean, 63.0 ng) for BaA, 6.6-90.2 ng (mean, 27.4 ng) for BkF, 9.8-167.4 ng (mean 47.2 ng) for BaP, and 10.8-206.0 ng (mean, 61.8 ng) for BghiP. The results showed no correlation between the age and the concentration of PAHs in the lung, but some correlation was found between the age and the lung weight (p<0.01). There were significant correlations among the concentrations of the compounds in the lung (p<0.01). These results suggest that dogs, like humans, are affected by automobile exhaust and other common generation sources of such substances. PMID- 15297751 TI - Centrosome amplification and chromosomal instability in feline lymphoma cell lines. AB - To evaluate the presence of centrosome amplification and the resulting chromosomal instability in cat tumors, a newly established feline lymphoma cell line and four already established feline lymphoma cell lines were examined using immunohistochemical analysis of centrosomes. The number of chromosomes were subsequently counted by metaphase spread. Moreover, to explore whether mutational inactivation of the p53 gene or inactivation of the P53 protein caused by mdm2 gene overexpression, occurred in the feline lymphoma cell lines, mutational analysis of the feline p53 gene was carried out. The expression of feline mdm2 mRNA was evaluated by reverse transcriptase-polymerase chain reaction (RT-PCR). Centrosome amplification and chromosomal instability was observed in three out of the five feline lymphoma cell lines. Of these three feline lymphoma cell lines, one had aberrations in the P53 amino-acid sequence, whereas the others had none. There was no significant difference in the expression of mdm2 mRNA between peripheral blood mononuclear cells (PBMC) obtained from a normal cat and that of the five feline lymphoma cell lines. These findings indicate that centrosome amplification also occurs in cat tumors and is strongly correlated with chromosomal instability, suggesting that the immunostaining of centrosomes could be an alternative method for the examination of the chromosomal instability. Furthermore, this study suggests the presence of unknown mechanism that leads to the centrosome amplification in feline lymphomas. PMID- 15297752 TI - Establishment of a chicken monoclonal antibody panel against mammalian prion protein. AB - A panel of chicken monoclonal antibodies (mAbs) was developed against prion protein (PrP), the sequence of which is a highly conserved molecule among mammals. A portion of the splenocytes from chickens immunized with recombinant mouse PrP was fused with the chicken B cell line, MuH1. The remaining splenocytes were used to generate the recombinant mAbs by phage display. A total of 36 anti PrP mAbs, 2 from cell fusion and 34 from phage display were established. The specificity of these mAbs was determined by Western blot and ELISA using various PrP antigens including recombinant PrPs, synthetic PrP peptides and PrPs from brains or scrapie-infected neuroblastoma cell line. These mAbs were classified into three main groups, protease K (PK)-sensitive (Group I), PK cleavage site proximal (Group II) and PK-resistant (Group III), based on their abilities to recognize PrP following PK-treatment. Some mAbs were found to selectively recognize different glycoforms of PrP as well as the metabolic fragments of PrP. Furthermore, we found that PrP recognition by chickens differed from that by PrP knockout mouse. These results indicate that these newly generated PrP antibodies from chickens will help to research the PrP and to establish the diagnosis of prion disease. PMID- 15297754 TI - Inhibition by sulfatide of 21-kDa protein phosphorylation by protein kinase C in cow mammary gland and its reversal by phosphatidylserine. AB - The effect of sulfatide, a sulfated sphingolipid, on phosphorylation of endogenous proteins by protein kinase C (PKC) was examined in cow mammary gland. Several proteins, including 21-kDa, 43-kDa and 56-kDa proteins in the cytosolic fraction, were found to be substrates for PKC by phosphorylation in the absence or presence of the cofactors 1-oleoyl-2-acetyl-sn-glycerol (OAG), phosphatidylserine (PS) and Ca2+. Sulfatide inhibited the 21-kDa phosphorylation, whereas it enhanced the 56-kDa and 43-kDa phosphorylation. Experiments were then conducted to examine whether other sphingolipids, including sphingosine, dihydrosphingosine, ceramides, galactocerebrosides, psychosine and sphingomyelin, modulated phosphorylation of the PKC substrates. Sphingosine, dihydrosphingosine and psychosine did not inhibit the 21-kDa phosphorylation; however, they enhanced the 56-kDa and 43-kDa phosphorylation. Ceramides, galactocerebrosides and sphingomyelin did not inhibit the 21-kDa or enhance the 56-kDa and 43-kDa phosphorylation. The inhibition by sulfatide of the 21-kDa phosphorylation was reversed by excess addition of PS, but not by OAG or Ca2+; whereas the enhancement by sulfatide, as well as sphingosine, dihydrosphingosine and psychosine, of 56-kDa and 43-kDa phosphorylation was not affected by PS, OAG or Ca2+. It is suggested that sulfatide is involved in the regulation of PKC dependent phosphorylation by modulating the association of PKC substrates, in particular the 21-kDa protein, with membrane phospholipids in cow mammary gland. PMID- 15297753 TI - Allele frequency distribution of the canine dopamine receptor D4 gene exon III and I in 23 breeds. AB - Various canine breeds are remarkably different from each other not only in their sizes and shapes but also in behavioral traits, suggesting that some of them are under genetic control. Although dopaminergic neurotransmission system is considered to affect animal behavior, little is known about related genes in canine. Relations between specific alleles in polymorphic regions of the dopamine receptor D4 gene (DRD4) and personality or psychiatric disorders have been reported in humans, and we first found polymorphism in exon III region of the gene in 4 canine breeds. In this study we surveyed allele frequency distribution in 23 breeds including a total of 1,535 unrelated individuals. In exon III, 8 alleles including a novel allele were identified. A group of breeds in which the alleles 447b, 498 and 549 were frequent tended toward high scores in aggression related behavioral traits than that with frequent alleles 435 and 447a. Moreover, a polymorphism based on 24 bp insertion/deletion was found in exon I region for the first time in dogs. This information may be of use for candidate gene studies of behavioral variation in dogs. PMID- 15297755 TI - Usefulness of myelography with multiple views in diagnosis of circumferential location of disc material in dogs with thoracolumbar intervertebral disc herniation. AB - The usefulness of myelography with multiple views (lateral, ventrodorsal, left and right oblique view) in the diagnosis of the exact circumferential location of herniated disc material around the spinal cord in 80 dogs diagnosed with thoracolumbar intervertebral disc herniation at surgery was assessed by comparison of clinical and surgical findings. The circumferential location of the compressing mass was diagnosed in 94% of dogs on myelography. The oblique view was of more benefit than the ventrodorsal view in diagnosing the circumferential distribution of the compressing mass. Only the oblique view contributed to a diagnosis of lateralization of the compressing mass in 45% of dogs. Fourteen percent of dogs had clinical lateralization contralateral to myelographic lateralization. The myelographic localization agreed with the surgical localization in 97% of dogs with regard to the exact location of herniated disc material. The presence of clinical lateralization contralateral to myelographic lateralization and a high proportion of agreement of myelographic and surgical localization documents that myelography with multiple views is useful and essential to accurately determine the circumferential location of disc material around the spinal cord. PMID- 15297756 TI - Nephropathogenesis of chickens experimentally infected with various strains of infectious bronchitis virus. AB - Four-day-old specific-pathogen-free chickens were inoculated by eyedrop with four different strains (Gray, JMK, CV56b, and Wolgemuth) of infectious bronchitis virus (IBV). Birds were monitored clinically and euthanatized at 1, 4, 7, and 14 days postinfection and tissues were collected for virus isolation, histopathologic examination, in situ hybridization (ISH), and immunohistochemistry (IHC). Clinical disease was severe in chickens infected with Wolgemuth, but no overt disease was observed with the other strains. Virus was isolated from the kidneys of chickens infected with the Gray-, CV56b-, and Wolgemuth-strains of IBV. Histologically, interstitial nephritis was evident in chickens infected with these same 3 strains. However, viral nucleic acid and antigen were detected only with Wolgemuth-infected kidneys by ISH and IHC. These results indicate that the pathological changes in kidneys from chickens infected with Gray and CV56b may not have resulted from the cytolytic action of the virus. PMID- 15297757 TI - Development of polymerase chain reaction and comparison with in situ hybridization for the detection of Haemophilus parasuis in formalin-fixed, paraffin-embedded tissues. AB - DNA extraction and nested polymerase chain reaction (PCR) were developed for the detection of Haemophilus parasuis from formalin-fixed, paraffin-embedded tissues. The results for nested PCR were compared with those determined by in situ hybridization. The optimal results obtained show that use of xylene deparaffinization, digestion with proteinase K followed by nested PCR is a reliable detection method. A distinct positive signal was detected in 20 pigs naturally infected with H. parasuis by in situ hybridization. The rate of agreement between nested PCR and in situ hybridization for the detection of H. parasuis in formalin-fixed, paraffin-embedded tissues was 100%. The nested PCR could be applied successfully to formalin-fixed, paraffin-embedded tissues for the detection of H. parasuis with bacterial isolation. PMID- 15297759 TI - Examination of meat components in commercial dog and cat feed by using polymerase chain reaction-restriction fragment length polymorphisms (PCR-RFLPs) technique. AB - It has been shown that certain slow neurological diseases such as bovine spongiform encephalopathy (also known as "mad cow" disease) could be transmitted through contaminated food intake by animals; therefore, the examination of meat components in commercial feeds is important for the control of the disease in public health. The combination of polymerase chain reaction-restriction fragment length polymorphisms (PCR-RFLPs) technique was applied to examine the meat components in dog and cat commercial feeds. The partial nucleotide sequence (359 bp) of animal mitochondrial cytochrome b (cytb, CYT) gene was amplified by PCR and then digested with restriction enzyme Alu I or Mbo I. In this work, eight brands of commercial dog and cat feeds available in Taiwan were examined. All brands of dog feeds that were tested contained meat from four different animals (cattle, pig, goat and chicken). In cat feeds, the chicken meat was found in five out of eight brands. PMID- 15297758 TI - Reproductive disorders in pubertal and adult phase of the male rats exposed to vinclozolin during puberty. AB - Vinclozolin (VCZ) is a systemic dicarboximide fungicide with antiandrogenic activity. Reproductive toxicity of VCZ was investigated in male rats exposed to VCZ during puberty. Sprague-Dawley male rats aged with 35 days were assigned to six different groups; negative control, positive control receiving flutamide (100 mg/kg), VCZ (100, 200 and 400 mg/kg), and a combination of VCZ (200 mg/kg) + methyltestosterone (100 mg/kg). The animals were treated with test compounds by oral gavage daily during 35 to 44 days of age. In pubertal rats sacrificed on the next day after final treatment, VCZ or flutamide-treated group showed a decrease in weights of prostate, epididymis, and seminal vesicle, hypertrophy of Leydig cells in the testis, detached debris and sloughed cells in the tubules of the caput epididymis, and an increase in serum testosterone levels. On the other hand, combined treatment of VCZ + methyltestosterone decreased testicular weight, increased seminal vesicle weight, and induced degeneration of spermatocytes. In adult rats sacrificed at five weeks after final treatment, flutamide decreased testicular sperm counts, and VCZ, flutamide and VCZ + methyltestosterone also decreased epididymal sperm counts. In addition, treatment of VCZ (400 mg) or VCZ + methyltestosterone decreased some motion kinematic parameters of sperms including curvilinear velocity, mean angular displacement and lateral head displacement. Flutamide treatment also decreased lateral head displacement. These results indicate that VCZ exposure during pubertal period in male rats causes reproductive disorders in puberty and adulthood. PMID- 15297760 TI - Ovarian teratoma with a formed lens and nonsuppurative inflammation in an old dog. AB - A 9-year, 7-month-old female German shepherd weighing 26.6 kg was admitted to the hospital for pica and diarrhea. A large mass was found in the right ovary and removed, and cross section of the mass revealed a multilobular tumor consisting of several cystic cavities which contained tufts of dark hair in thick creamy white sebaceous fluid. Histologically, the tumor consisted of adipose tissue, central nervous tissue, crystalline lens, cartilage and bone. In the central nervous tissue, lens and lesions like nonsuppurative inflammation comprizing of accumulation of glial cells and lymphocytic perivascular cuffing were observed. The tumor was diagnosed as a mature cystic teratoma. PMID- 15297761 TI - Detection of bovine lactoferrin binding protein on Jurkat human lymphoblastic T cell line. AB - Lactoferrin (Lf), a member of the transferrin family protein, is an iron-binding protein that is known to interact with mammalian cells through a specific receptor. We examined binding of Lf to Jurkat human lymphoblastic T cell line (Jurkat cells) by far Western blotting, and found that bovine Lf and human Lf bound to the same protein components of Jurkat cells, and that pepsin digestion of Lf disrupts the sites responsible for binding to cellular proteins. We also found that the sugar chains of bovine Lf are not involved in binding between bovine Lf and Jurkat cells. Bovine Lf, bovine transferrin and ovotransferrin bound to the same proteins of Jurkat cells, which had molecular weights of about 35 kDa. PMID- 15297762 TI - Effects of Central Kalimantan plant extracts on intraerythrocytic Babesia gibsoni in culture. AB - The inhibitory effects of 45 plant extracts selected from Central Kalimantan, Indonesia on Babesia gibsoni in vitro and their acute toxicity to mice were evaluated. Of these plant extracts studied, Arcangelisia flava, Curcuma zedoaria, Garcinia benthamiana, Lansium domesticum and Peronema canescens were found to have appreciable antibabesial activity with IC50 values from 5.3 to 49.3 microg/ml without acute toxicity in mice at the intraperitoneal dose of 0.7 g/kg of body weight. PMID- 15297763 TI - Brachyspira pilosicoli isolated from pigs in Japan. AB - Two of four weak beta-hemolytic isolates of intestinal spirochetes isolated from pigs in Japan possessed a unique base alignment of TTTTTT on the 16S ribosomal DNA of Brachyspira pilosicoli and were identified as B. pilosicoli. The other two isolates were not identified by this technique. The identified isolates were 4.2 to 11 microm in length and 0.2 to 0.3 microm in diameter, 4 periplasmic flagella at each end were observed dominantly. The isolates were hippurate positive but indole negative. This is the first report on the isolation of B. pilosicoli from pigs in Japan. PMID- 15297764 TI - Identification of the Cryptosporidium isolate from chickens in Japan by sequence analyses. AB - The Cryptsosporidium isolate from chickens in Japan by Itakura et al. is not yet accurately identified because of several discrepancies in phenotypic features. We attempted to identify this isolate by analyzing the partial sequences of the 18S rRNA, COWP and HSP70 genes. The chicken isolate showed nearly 100% homology in each gene with C. baileyi, but less than 91% homology with C. meleagridis. In addition, these genes were classified into the same cluster with C. baileyi other than C. meleagridis by phylogenetic analysis. From these results, the Cryptosporidium isolate from chickens in Japan is considered to be one of the strains of C. baileyi. PMID- 15297765 TI - Improvement of anemia associated with chronic renal failure by recombinant human erythropoietin treatment in ICR-derived glomerulonephritis (ICGN) mice. AB - The ICR-derived glomerulonephritis (ICGN) mouse, a novel inbred mouse strain with a hereditary nephrotic syndrome, develops severe anemia associated with chronic renal failure. To reveal the pathogenic mechanism of anemia in ICGN mice, we subcutaneously administered recombinant human erythropoietin (rhEPO; 5 IU/mouse/day) or saline for 5 days to ICGN mice. In terminal-stage ICGN mice with severe anemia, rhEPO significantly increased hematocrit (Ht), red blood cells (RBC) and hemoglobin levels. Endogenous EPO levels in peripheral blood were reduced by rhEPO injection. No histopathological changes in bone marrow and kidneys were induced by rhEPO injection. Insufficiency of EPO may cause anemia in ICGN mice. PMID- 15297766 TI - Detection of a bovine group C rotavirus from adult cows with diarrhea and reduced milk production. AB - Only two strains (Shintoku and porcine-like WD534tc) of group C rotavirus (GCR) from cattle have been reported to date. A GCR designated the Yamagata strain was the only pathogen detected in an outbreak of adult cow diarrhea accompanied by a decrease in milk production. The nucleotide sequences of the VP6 and VP7 genes from strain Yamagata were determined. Comparative sequence analysis showed that the sequence identities between strains Yamagata and Shintoku were markedly high in both VP6 gene (98.1%) and VP7 gene (93.5%), and that these strains belonged to the same clusters which were distinguished from GCRs from different host species in phylogenetic trees of these genes. These results suggested strongly that cattle species is one of the natural hosts of GCR infection, and that GCRs are a cause of adult cow diarrhea. PMID- 15297767 TI - Migration of warble fly larvae in the yak and optimum timing of ivermectin treatment. AB - Sixty yaks were autopsied to determine the migration pattern of warble fly larvae. In August, first instars were observed in the body of yak for the first time. These larvae peaked in number in October. From November to February, second instars were detected and their number peaked in January. Third instars appeared in January and peaked in March. Forty-five yaks were administered with ivermectin: 15 animals in September, 15 in October and 15 in November. Between December and June, the number of warbles was checked by palpation. Although some warbles were observed in the September- and November-treated groups, no warbles were detected in the October-treated group. Treatment of yaks with ivermectin was most effective for warble fly in October. PMID- 15297768 TI - Involvement of peripheral mechanism in the verapamil-induced potentiation of morphine analgesia in mice. AB - Morphine's analgesic actions are thought to be mediated through both the central and peripheral nervous systems. L-type calcium channel blockers have been reported to potentiate the analgesic effects of morphine, but the locus of this interaction is not known. In this experiment, we examined the site of verapamil induced potentiation of morphine analgesia in mice using the quaternary opioid receptor antagonist naloxone-methiodide (NLX-M). Subcutaneous injections of morphine increased locomotor activity and serum corticosterone level, which are mediated by the central nervous system. These central effects were not antagonized by 0.1 mg/kg of NLX-M, whereas this dose of NLX-M partially antagonized the analgesic effect of morphine. Treatment with verapamil potentiated morphine analgesia in a dose-dependent manner. The verapamil-induced potentiation of morphine analgesia was abolished by pretreatment with NLX-M (0.1 and 1 mg/kg). These findings suggest that peripheral mechanisms partially contribute to morphine analgesia and mediate the potentiation of morphine analgesia by verapamil. PMID- 15297770 TI - Acetylcholine-induced translocation of RhoA in freshly isolated single smooth muscle cells of rat bronchi. AB - By using immunofluorostaining and confocal laser microscopy, acetylcholine induced translocation of RhoA was visualized in freshly isolated bronchial smooth muscle cells of the rat. The cellular distribution of RhoA at rest was observed uniformly in the cytosolic space with no staining in the nucleus, whereas acetylcholine stimulation induced a relocalization of RhoA to the cell periphery. From the results of line scans and surface plots, the peripheral to cytosolic ratio of RhoA was significantly increased by acetylcholine stimulation. Thus, the present study clearly demonstrated an acetylcholine-induced translocation of RhoA to the plasma membrane in single bronchial smooth muscle cells of the rat. PMID- 15297769 TI - Soybean isoflavones eliminate nifedipine-induced flushing of tail skin in ovariectomized mice. AB - Hot flushes are one of the most frequent symptoms in menopausal women. We investigated effect of soybean isoflavones (Soyaflavone HG) on nifedipine-induced flushing in ovariectomized mice. Ovariectomy markedly aggravated nifedipine induced increase in tail skin temperature. Soyaflavone HG (10 mg/kg, p.o., once a day for 5 days) inhibited nifedipine-induced flushing in ovariectomized mice. The inhibitory effect of Soyaflavone HG was significantly reversed by an estrogen receptor antagonist, ICI 182,780, suggesting that Soyaflavone HG prevents nifedipine-induced flushing partially through estrogen receptors. We presented the experimental evidence suggesting that soybean isoflavones including Soyaflavone HG have the benefits for menopausal hot flushes. PMID- 15297771 TI - Antioxidant effects of stereoisomers of N-acetylcysteine (NAC), L-NAC and D-NAC, on angiotensin II-stimulated MAP kinase activation and vascular smooth muscle cell proliferation. AB - We examined the effects of the stereoisomers of N-acetylcysteine (NAC), L-NAC and D-NAC, on cellular glutathione (GSH) concentration and whether NAC-regulated cellular GSH levels are directly associated with angiotensin II (Ang II)-induced intracellular signaling events in vascular smooth muscle cells (VSMC). Both L-NAC and D-NAC similarly increased intracellular GSH concentration. We found that L NAC and D-NAC both inhibited Ang II-induced c-Jun N-terminal kinase and p38 mitogen-activated protein kinase activation and [(3)H]-thymidine incorporation in VSMC. Our present study indicates the comparable effects of NAC stereoisomers in regulating intracellular GSH and the redox-dependent intracellular signaling mechanisms in VSMC. PMID- 15297772 TI - Safety of infliximab in patients suffering from inflammatory bowel disease. PMID- 15297773 TI - Safety aspects of infliximab in inflammatory bowel disease patients. A retrospective cohort study in 100 patients of a German University Hospital. AB - BACKGROUND: Infliximab, a chimeric anti-tumour necrosis factor monoclonal antibody with potent anti-inflammatory effects, represents an effective treatment option in patients with severe inflammatory bowel disease (IBD). Serious side effects of such an immunomodulating therapy are speculated and therefore we reviewed our clinical experience in a retrospective safety study looking upon a single cohort of 100 IBD patients from a large German University Hospital. METHODS: 100 patients with severe Crohn's disease (n = 92), ulcerative colitis (n = 7) or indeterminate colitis (n = 1) treated with infliximab (5 mg/kg) from January 2000 to December 2003 were retrospectively analysed for acute and subacute adverse events by chart review. RESULTS: Overall, infliximab therapy was generally well tolerated. No fatal complications, malignancies, autoimmune diseases, neurologic or cardiovascular complications were observed in the cohort during the study period. Overall, adverse events were observed in 10 patients: 2 patients showed an acute infusion reaction, 1 patient a serum sickness-like reaction, in 4 patients a bacterial or viral infection occurred, in 1 patient pancytopenia and 2 patients developed surgical complications. Only 6 patients with adverse events required admission to hospital. A case of tuberculosis after infliximab was not found. The lack of adverse side-effects was associated with young median age and infrequent comorbidities of the cohort. CONCLUSION: Regarding its strong immunomodulating capacity, infliximab appears to be an efficient and relatively safe therapeutic option for patients with severe IBD. However, the use of infliximab requires careful screening and close patient monitoring to identify patients at risk and the infrequent, but sometimes serious complications of infliximab. PMID- 15297774 TI - Mutually distinctive gradients of three types of intestinal alkaline phosphatase along the longitudinal axis of the rat intestine. AB - BACKGROUND/AIMS: A descending gradient of alkaline phosphatase (AP) activity is well known to exist in the rat intestine. To understand the physiological role(s) of intestinal AP (IAP), we examined in detail the distribution of IAP subtypes along the longitudinal axis of the rat intestine. METHODS: The consecutive 2-cm segments of the upper intestine were analyzed by sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE) followed by staining for IAP activity. The reverse transcription-polymerase chain reaction was conducted to assess the relative levels of tissue mRNA for the 2 distinct genes of rat IAP, rIAP-I and rIAP-II. RESULTS: A descending gradient of IAP activity was found along the upper small intestine, where 3 bands of the activity, termed types 1, 2 and 3 from the anode were located on SDS-PAGE gels. Along the longitudinal axis, types 1 and 3 showed ascending and descending gradients, respectively; whereas type 2 had a broad uniform distribution. The mRNA level for rIAP-I showed an ascending distribution, and that of rIAP-II, a descending one. Type 3 IAP was drastically diminished in duodenum of cysteamine-ulcer rats, but type 1 remained constant. CONCLUSION: These distinctive gradients of IAP subtypes may provide a clue to scrutinize the physiological significance of AP in the rat intestine, which is a flow system with polarity. PMID- 15297775 TI - Patients with subjective food hypersensitivity: the value of analyzing intestinal permeability and inflammation markers in gut lavage fluid. AB - BACKGROUND/AIM: Subjective food hypersensitivity is prevalent in the general population. The aim of this study was to seek objective evidence of food hypersensitivity by analyzing intestinal permeability and inflammation markers in gut lavage fluid. METHODS: Fifty-two patients with abdominal complaints self attributed to food hypersensitivity were examined by skin prick test, serum IgE analysis, double-blind, placebo-controlled food challenge (DBPCFC), and intestinal lavage. The results were compared with those of 44 patients without food hypersensitivity. Neither the patients nor the controls had organic diseases that could explain their symptoms. Intestinal lavage was performed by administering 2 liters of isotonic polyethylene glycol (molecular weight 3,350 daltons) solution containing 50 microCi of [51Cr]EDTA through a nasoduodenal tube. The first clear fluid passed per rectum was collected and analyzed for histamine, eosinophilic cationic protein (ECP), tryptase, and calprotectin. The intestinal permeability was assessed by determining the 5-hour urinary excretion of [51Cr]EDTA. Calprotectin was also analyzed in native faecal samples. RESULTS: The ECP concentration in gut lavage fluid was significantly higher in the patients than in the controls (p = 0.007), but the overlap between groups was large. Food hypersensitivity was confirmed by positive DBPCFC in only 4 patients. On average, histamine and ECP concentrations were high in these patients. Tryptase, intestinal permeability, and faecal and lavage calprotectin levels were normal. CONCLUSIONS: Very few patients had objective evidence of food hypersensitivity. Analyzing intestinal permeability and inflammation markers in gut lavage fluid did not contribute to the diagnosis, but further studies on histamine and ECP are warranted. PMID- 15297776 TI - Adsorptive granulocyte and monocyte apheresis versus prednisolone in patients with corticosteroid-dependent moderately severe ulcerative colitis. AB - BACKGROUND/AIM: Active ulcerative colitis (UC) is often associated with increased peripheral granulocytes and monocytes/macrophages which show activation behavior and prolonged survival time. Further, mucosal granulocyte level parallels intestinal inflammation and can predict UC relapse. Accordingly, our aim was to see if adsorptive granulocyte/monocyte apheresis (GMA) can promote remission and spare steroid in patients with steroid-dependent (SD) UC. METHODS: 69 SD patients, at the time of relapse, were randomly assigned to groups I (n = 46) and II (n = 23). The mean dose of prednisolone (PSL) was 12 mg/day/patient, CAI (clinical activity index) 9.2 in both groups. Group I patients were given up to 11 GMA sessions over 10 weeks with Adacolumn; in group II, the mean dose of PSL was increased to 30 mg/day/patient. RESULTS: At week 12, 83% of group I and 65% of group II patients were in remission, CAI in group I was 1.7 (p < 0.001) and in group II, 2.5 (p < 0.001). Further, during the 12 weeks of treatment, the cumulative amount of PSL received per patient was 1,157 mg in group I and 1,938 mg in group II (p = 0.001). CONCLUSIONS: GMA appeared to be an effective adjunct to standard drug therapy of moderately severe UC by promoting remission and sparing steroids. PMID- 15297777 TI - Gallbladder involvement of Henoch-Schonlein purpura mimicking acute acalculous cholecystitis. AB - BACKGROUND/AIMS: We report the case of a 53-year-old man with fever and severe abdominal pain following an upper respiratory tract infection. METHODS/RESULTS: The clinical signs, the laboratory markers, and the ultrasound findings were consistent with acute acalculous cholecystitis and he underwent cholecystectomy. Histologically the gallbladder showed vasculitis and the patient developed postoperatively a purpuric rash of the legs with transient ankle arthritis. Gastroscopy revealed prepyloric ulcers consistent with vasculitis. The patient was diagnosed as having Henoch-Schonlein purpura. Gastric ulcers as well as arthritis disappeared upon treatment with corticosteroids and proton pump inhibitors. CONCLUSION: Taken together, Henoch-Schonlein purpura can mimic acute cholecystitis and should be considered as a rare differential diagnosis of acute cholecystitis. PMID- 15297778 TI - Gastric emptying of a solid meal starts during meal ingestion: combined study using 13C-octanoic acid breath test and Doppler ultrasonography. Absence of a lag phase in 13C-octanoic acid breath test. AB - Scintigraphy and the 13C-octanoic acid breath test are both applied to assess gastric emptying. Using the 13C-octanoic acid breath test, excretion curves show 13C excretion immediately after ingestion of a solid egg meal, in contrast with scintigraphy where gastric emptying is observed after a lag phase. The aim of our study was to investigate whether transpyloric flow occurs during and directly after meal ingestion. Therefore, transpyloric flow was measured during and after ingestion of an egg meal labeled with 13C-octanoic acid, using Doppler ultrasonography. The breath test was performed simultaneously, with samples taken at regular intervals. The first emptying episode was observed 6.9 (3.9-16.2) min after start of meal ingestion. A significant relation between recovery of 13C and total duration of gastric emptying during the first 20 min was observed (partial correlation coefficient r = 0.80, p < 0.001). In conclusion, transpyloric flow starts during ingestion of a solid egg meal and results in detectable excretion of 13C. PMID- 15297779 TI - Efficacy and safety of pantoprazole versus ranitidine in the treatment of patients with symptomatic gastroesophageal reflux disease. AB - BACKGROUND/AIM: Gastroesophageal reflux disease (GERD) is a prevalent disease associated with a high symptom burden and a reduced quality of life. This multicenter, randomized, double-blind study compared relief from key GERD symptoms (heartburn, acid eructation, and pain on swallowing) and from other gastrointestinal symptoms (epigastric pain, vomiting, nausea, flatulence, retching, and retrosternal feeling of tightness) and safety profiles of the proton pump inhibitor pantoprazole and the H2 antagonist ranitidine in patients suffering from symptomatic GERD. METHODS: The patients [338 intention-to-treat (ITT) population; 284 per-protocol (PP) population] received 20 mg pantoprazole (once daily in the morning) plus ranitidine placebo (once daily in the evening; ITT n = 167, PP n = 136) or pantoprazole placebo (once daily in the morning) plus 300 mg ranitidine (once daily in the evening; ITT n = 171, PP n = 148) for 28 days. The primary efficacy criterion (ITT and PP populations) was relief from key GERD symptoms (heartburn, acid eructation, and pain on swallowing) after 28 days of treatment. Secondary criteria (PP) included relief from key GERD symptoms on day 14, relief from all gastrointestinal symptoms on days 14 and 28, and relief from key GERD symptoms on days 14 and 28. Safety evaluations included adverse events and laboratory assessments. RESULTS: Significantly more pantoprazole treated patients were free from key GERD symptoms at day 28 (68.3%, n = 114) as compared with ranitidine-treated patients (43.3%, n = 74; 95% confidence interval for odds ratio 1.84-4.51). Pantoprazole was also significantly more efficacious in controlling all gastrointestinal symptoms of GERD. By day 28, 51.5% (n = 70) of the pantoprazole-treated patients were completely symptom free versus 31.1% (n = 46) of the ranitidine-treated patients (95% confidence interval for odds ratio 1.45-3.83). Both treatments were well tolerated. CONCLUSION: Pantoprazole is significantly superior to ranitidine in the treatment of key and associated gastrointestinal symptoms of GERD and is well tolerated. PMID- 15297780 TI - Properties of human trabecular bone cells from elderly women: implications for cell-based bone engraftment. AB - Tissue-engineered bone regeneration offers new therapeutic options in the treatment of patients with fractures. Due to the changes in the hormone levels, elderly women are most affected by bone fractures and thus constitute one of the future target groups of cell-based bone therapy. For designing cell-based therapy approaches, a better understanding of individual-dependent variations in bone derived cells of the host is required. In this study, a simple, high-yield procedure is described for the collection of cells from bone tissue of a high number of elderly women. The cultured cells display stem cell characteristics indicated by the presence of a CD13+, CD44+, CD90+, CD147+, CD14-, CD34-, CD45- and CD144- cell populations and by a stable undifferentiated phenotype as well as by the ability to proliferate extensively while retaining the potential to differentiate along the osteoblastic lineage even after 27 cell doublings. A high variability in the number of cell-forming units (CFUs) within a donor population of 34 samples, in the morphology within 50 donors, in the expression of alkaline phosphatase within 15 samples and in the responsiveness to BMP-2 was evident, but no age-related correlation could be found. In conclusion, the data indicate that individual variations in cell number, cell morphology and in the osteogenic potential of progenitor cells of the patient may be relevant for a successful treatment of bone fractures in the elderly by cell-based therapy approaches. PMID- 15297781 TI - Evaluation of mineralized collagen and alpha-tricalcium phosphate as scaffolds for tissue engineering of bone using human mesenchymal stem cells. AB - Owing to their plasticity and high proliferation capacity in vitro, mesenchymal stem cells (MSC) isolated from human bone marrow are promising candidates for use in tissue engineering approaches for the repair or replacement of mesenchymal tissues such as bone, cartilage or tendon. In keeping with the tissue engineering concept, these cells are cultivated on three-dimensional (3D) scaffolds to replace 3D tissue defects. Among the scaffolds tested for tissue engineering of bone, those containing phosphorus and calcium, as natural bone does, are the most promising candidates for this purpose. In this study, MSC from five patients were isolated from bone marrow. After in vitro expansion, cells were cultivated and differentiated towards the osteogenic lineage on mineralized collagen sponges and alpha-tricalcium phosphate (alpha-TCP). To analyze how appropriate these scaffolds would be for tissue engineering purposes, we established an in vitro characterization system to describe seeding efficiency, cell distribution and proliferation behavior on each scaffold. Real-time reverse transcriptase polymerase chain reaction quantification of important genes involved in osteogenic differentiation [e.g. bone sialoprotein (BSP), bone morphogenic protein 2, alkaline phosphatase and osteocalcin] was used to monitor the differentiation process of cells seeded on mineralized collagen and alpha-TCP. Using this in vitro characterization, we were able to demonstrate effective 3D growth of MSC on both scaffolds investigated. Improved osteogenic differentiation was observed on the scaffolds as compared to control monolayers. Of the two matrices, mineralized collagen was superior to alpha-TCP with regard to seeding efficacy (98 vs. 67%, p = 0.0003), increase in osteogenic marker genes (BSP expression on day 24, Pcollagen vs. TCP = 0.046) and 3D cell alignment (cell infiltration up to 500 vs. 200 microm). In conclusion, our data suggest that mineralized collagen is a promising candidate for use as a scaffold in tissue engineering of bone. PMID- 15297782 TI - Developing mouse Sertoli cells in vitro: effects on developing ovaries in co culture and production of anti-Mullerian hormone. AB - Previous work in our laboratory showed that pre-Sertoli cells adopt an epithelial phenotype when cultured in the presence of reconstituted basement membrane (RBM), and so cultures were established with and without this substrate. Biological activity of isolated developing mouse Sertoli cells maintained in vitro was assessed in the current study by utilising a co-culture approach, to determine whether the cells were capable of affecting ovarian differentiation. Developing Sertoli cells isolated at embryonic day (E) 12.5 exerted a deleterious effect on E12.5 ovaries in co-culture, inducing a loss of germ cells. However, when cells were isolated a day later and co-cultured with E13.5 ovaries (after entry to meiosis has begun), germ cells survived and showed evidence of meiosis, although ovigerous cords in co-cultures were masculinized compared to those of control cultured ovaries. Thus, both stages examined showed biological effects; cultured pre-Sertoli cells explanted at E12.5 showed a negative effect on female germ cells, whereas those explanted at E13.5 masculinized ovigerous cords. The functional status of isolated developing mouse Sertoli cells in vitro was further assessed by immunocytochemistry to investigate the expression of anti-Mullerian hormone, an early product of pre-Sertoli cells. Positive immunostaining was seen in developing Sertoli cells in vitro, particularly where cells had been explanted to an RBM substrate, demonstrating that good epithelial morphology is associated with improved function. Our culture system is therefore well suited for investigating factors produced by developing Sertoli cells, their role in influencing testicular morphogenesis and their potential to perturb ovarian differentiation. We believe that this in vitro approach provides a more physiological assessment compared with the knockout mouse model, where global effects of genes with housekeeping functions can compromise overall development. PMID- 15297783 TI - Precedence of cell-zona adhesion over cell-cell adhesion during marsupial blastocyst formation prohibits morula formation and ensures that both the pluriblast and trophoblast are superficial. AB - This study, based on 38 samples taken between the 16-cell stage on day 2.5 of gestation and the expanded 1.0-mm-diameter unilaminar blastocyst on day 6, describes the ultrastructural changes that occur in the conceptus of the marsupial Sminthopsis macroura in relation to cell-zonal adhesion initiated at the zygote stage and cell-cell adhesion initiated at the 16-cell stage, lineage allocation, extracellular matrix (ECM) secretion and embryo coat changes. In S. macroura, rather flattened pluriblast and rounded trophoblast cells appear as different cell types during the fourth division when nucleolar reticulation suggests activation of the zygotic genome in both cell types. The differences disappear nearly completely in blastocysts of 0.6-0.8 mm in diameter, but the two cell types then reappear as two distinct populations. The ECM varies depending on its location within the conceptus up to the stage of the expanding blastocyst. It is of rather granular appearance between the cell lining and zona pellucida and consists of patches of homogeneous material embedded in an electron-lucent substance in the cleavage cavity. Homogeneous ECM coats trophoblast but not pluriblast cells on blastocoelic surfaces. Transient structures such as 'myosin like' fibrillar arrays, probably associated with exocytosis of ECM, and pearl string-like whorls are still present, but both disappear during further expansion of the 0.6- to 0.8-mm blastocyst. During blastocyst expansion, the patchy homogeneous ECM in the blastocoel changes structure and appears flocculent, while the continuous ECM coating trophoblast cells disappears. Pluriblast cells and yolk mass identify the embryonic pole and hemisphere, and the opposite hemisphere becomes abembryonic and is eventually fully lined by trophoblast cells. An increase in endocytotic, mainly coated vesicles at the apical, zona-orientated surface of both cell types is noticed and is probably responsible for uptake of the mucoid coat. In 1-mm blastocysts, numerous vesicles contain rod-shaped crystalline inclusions. PMID- 15297784 TI - Wallerian degeneration in the optic nerve of the rabbit. AB - Progressive anterograde axonal degeneration is known to follow after transection of the axon from the soma, which to some extent correlates with the passage of time after the lesion. However, the minimum time required for such changes to begin remains unresolved. In this study, 20 young adult rabbits of either sex underwent experimental monocular enucleation (left eye) under general anaesthesia. Left optic nerves from such animals were treated as experimental and those from either side of non-operated animals served as controls. Animals were sacrificed postoperatively at periods ranging from 12 h to 3 months. Brains were fixed with 10% formalin and Karnovsky fixatives by an intracardiac perfusion method. Light microscopy of 8-microm paraffin sections and 0.5-microm araldite sections from the optic nerves did not reveal any changes at 12 h. At 24 h, focal minute cavities appeared across the optic nerves. Those nerves from late postoperative stages revealed such cavities with increasing dimensions, disarray of fascicular organization, fragmentation, ovoid formation and finally dissolution of the myelin sheaths. There was an appreciable increase in the number, size and aggregation of glia cells. The debris of degeneration remained prominent even 3 months after enucleation. Electron microscopy revealed splitting of myelin, intramyelinic and periaxonal oedema and occurrence of amorphous and electron-dense materials in the degenerating nerve fibres. It was concluded that while the optic nerve showed degenerative changes as early as 24 h after enucleation, debris of degeneration was only partly removed even after 3 months. PMID- 15297785 TI - Changes of the cardiac architectures and functions for chronic hemodialysis patients with dry weight determined by echocardiography. AB - BACKGROUND/AIMS: Left ventricular hypertrophy (LVH) has long been known as an independent risk factor for cardiovascular deaths, in both dialysis and general populations. Numerous factors influence the pathophysiology of LVH. However, extracellular fluid may have a particularly important influence on this impact. Inferior vena cava diameter (IVCD) estimation is a non-invasive and relatively convenient method for obtaining a good correlation with the intravascular fluid status, and may obtain an optimal dry weight (DW) for chronic hemodialysis patients. This study estimates the DW of end-stage renal disease (ESRD) patients by echocardiographic measurement of the IVCD to observe changes in cardiac morphology and function. METHODS: A total of 88 patients, ranging from 26 to 90 (59.4 +/- 13.3) years of age, were involved in this study. The patients were divided into study (n = 48) and control (n = 40) groups. All patients received IVCD assessment via echocardiography bi-monthly for 1 year. In the study group patients, DW was adjusted according to the IVCD by echocardiography. Meanwhile, in the control group patients, DW was adjusted based on traditional clinical parameters. All patients underwent cardiac examinations and measurements, including left ventricular mass (LVM), wall thickness, chamber size and left ventricular systolic function by echocardiography, at the beginning and end of the study. RESULTS: Both groups displayed comparable clinical and biochemical parameters. The IVCD index correlated well with the cardiac parameters estimated by echocardiography. The LVM and left ventricular mass index (LVMI) was reduced significantly in the study group patients (from 200 +/- 64.2 to 187 +/- 63.2 g, p = 0.021; from 132 +/- 37.6 to 123 +/- 37.3 g/m(2), p = 0.014, respectively). Furthermore, the study group patients with fluid overload, named study subgroup A, displayed significant differences not only in LVM and LVMI, but also in septal wall thickness, left ventricular end-diastolic dimension and left atrial dimension. In contrast, the control group displayed no changes in these cardiac architectures during the study period. CONCLUSION: Adjusting DW via the IVCD measured by echocardiography for hemodialysis patients may prevent the progression of chamber dilatation and LVH, especially for patients with fluid overload. PMID- 15297786 TI - Dialysate leukocytes, sICAM-1, hyaluronan and IL-6: predictors of outcome of peritonitis? AB - BACKGROUND/AIMS: Despite effective antibiotic therapy, peritonitis still remains a major problem in peritoneal dialysis (PD). The aim of the present study was to investigate changes of CRP, dialysate leukocytes and IL-6, hyaluronan (HA) and sICAM-1 in dialysate during and after peritonitis and their association to the outcome of peritonitis. METHODS: Dialysate IL-6, HA and sICAM-1 were measured at the onset and on day 4, at the end of the treatment and 2 months after onset of peritonitis. Furthermore, CRP and dialysate leukocytes were measured on days 1-4. RESULTS: All measured soluble factors were higher on the first and fourth day than at the end of the treatment. sICAM-1 and HA were lower at the end of the treatment in patients who later had a relapse/re-infection. IL-6 remained higher 2 months after clinically cured peritonitis. CRP and dialysate leukocytes were higher on day 4 in patients with poor outcome. CONCLUSIONS: Peritonitis causes increased excretion of soluble factors. Low concentrations of sICAM-1 and HA at the end of the treatment were negative prognostic indicators. Higher IL-6 levels after peritonitis could be a sign of ongoing inflammation in the peritoneal membrane. Delayed decrease in CRP and dialysate leukocytes may indicate poor outcome. PMID- 15297787 TI - Fractional solute removal and KT/V in different modalities of renal replacement therapy. AB - The efficacy of solute removal by renal replacement therapy can be assessed by the commonly used index of KT/V (the fraction of the volume cleared from a solute). Fractional solute removal (FSR, the fraction of the total amount of the solute that was removed) is an alternative index that may be more appropriate than KT/V for comparison of the efficacy of different treatment modalities. To elucidate the relationship between these two indexes, we propose to discriminate between two notions of clearance: (1) instantaneous clearance K = (solute removal rate)/C(B), where C(B) is solute concentration in blood, and (2) treatment clearance K(T) = (average rate of solute removal per treatment)/C(B), where C(B) is C(B) at the beginning of the treatment. K is the clearance of the purification device (glomeruli, hemodialyzer or hemofilter) and the diffusive mass transport parameter (K(BD), MTAC) for continuous ambulatory peritoneal dialysis (CAPD). For all modalities of renal replacement therapy: FSR = K(T)T/V, and K(T) generally decreases with the treatment time. For purification of a single compartment with a constant volume, V, using an open loop system (i.e. with no recirculation or dwelling of dialysis fluid, as in hemodialysis (HD), hemofiltration (HF) or in the native kidney), FSR is a function of only one lumped, nondimensional parameter, KT/V(B), where V(B) is the distribution volume of the solute within the body. In contrast, if closed loop systems are applied, as for example in HD with recirculation of dialysis fluid (RD) or in peritoneal dialysis, FSR depends on two lumped, nondimensional parameters: KT/V(B) and KT/V(D), where V(D) is the volume of dialysis fluid. It is necessary to discriminate between K and K(T) for analysis of dialysis dose. For HD and HF, FSR is a function of KT/V, whereas KT/V alone does not allow calculation of FSR for CAPD and RD. The current practice of using K(T)T/V for CAPD but KT/V for HD and HF leads to confusion because of the inconsistency in the interpretation of the quantitative prescription of dialysis dose. The application of FSR, instead of KT/V, for all treatment modalities may solve this dilemma. Furthermore, K(T)T/V (currently used only for CAPD) is equal to FSR for all treatment modalities. Both FSR and K(T) may be generalized to describe the total solute removal per treatment cycle composed from a few treatment sessions. A few different definitions of the adequacy parameters for the treatment cycle are formulated and discussed. PMID- 15297789 TI - Characteristics of heart period variability in intubated very low birth weight infants with respiratory disease. AB - BACKGROUND: Heart period variability provides a measure of balance between the sympathetic nervous system (SNS) and parasympathetic nervous system (PNS). Since the PNS develops during the final weeks of gestation, premature infants have an overriding SNS. Spectral power analysis of heart period variability reveals two main frequency regions, the low frequency region (LF) representing primarily SNS activity and the high frequency region (HF) representing PNS activity. OBJECTIVES: To identify the characteristics of heart period power in the LF and HF regions in very low birth weight (VLBW) infants in the neonatal intensive care unit across gestational age groups and between sleep and awake states. METHODS: Data were collected from 16 intubated and mechanically ventilated VLBW infants with respiratory disease. Using spectral analysis techniques, heart period power in the two main frequency regions was extracted. RESULTS: HF power did not improve with gestational age as expected. LF power did increase with age, albeit nonsignificantly. LF and HF power were not significantly different between awake and sleep states. CONCLUSIONS: The results of this preliminary study suggest that PNS tone does not improve with gestational age in VLBW infants with respiratory disease. The intensive care environment may stimulate a sympathetic response in these infants and disrupt normal PNS development. PMID- 15297790 TI - N-acetylcysteine administration during the first week of life does not improve lung function in extremely low birth weight infants. AB - Oxygen toxicity is thought to be an important factor involved in development of bronchopulmonary dysplasia (BPD) in the very preterm infant. Glutathione (GSH) plays a major role in the antioxidant defense system in the preterm lung and there are theoretical implications that N-acetylcysteine (NAC) treatment could improve its function. The purpose of this study was to investigate whether NAC treatment during the first week of life to preterm infants improved neonatal lung function as a measure of lung injury. The study was part of a multi-center Nordic controlled trial with prophylactic intravenous NAC treatment (16-32 mg/kg/day) for 6 days in newborn infants with birth weights 500-999 g. Lung mechanics, with calculations of compliance and resistance of the respiratory system, together with measurements of functional residual capacity and indices of gas mixing efficiency in the lung, were performed in 33 preterm infants (18 received NAC and 15 placebo) before discharge from the NICU. Median (range) gestational age was 25 (24-28) weeks in the NAC-treated infants and 25 (24-29) in the placebo group. Corresponding mean (SD) birth weights were 0.774 (0.11) and 0.761 (0.12) kg respectively. Lung function measurements did not show any significant differences between NAC-treated infants compared to placebo when examined before discharge from the NICU. We conclude that prophylactic NAC treatment to extremely low birth weight infants during the first week of life does not improve lung function at term. PMID- 15297791 TI - Prevalence of neurological disorders in Bangalore, India: a community-based study with a comparison between urban and rural areas. AB - A population-based neuroepidemiological survey of 102,557 individuals in urban and rural Bangalore in Southern India was conducted to determine the prevalence and pattern of neurological disorders. The study population included subjects from urban (51,502) and rural (51,055) areas, identified through a two-stage stratified random sampling method. Trained social workers administered the screening questionnaire, which had been tested and validated in an earlier pilot study and a neurologist examined the individuals who screened positive. Adults, children (<15 years) and elderly adults (>60 years) constituted 61, 34 and 5% of the study group, respectively. There was a distinct difference in education, occupation and income levels between urban and rural areas with all these parameters being lower in the rural population. In the surveyed population, 3,206 individuals with neurological disorders were detected resulting in crude and age adjusted prevalence rates of 3,126 and 3,355 per 100,000 population, respectively. The prevalence rate among children, middle-aged (31-40 years) and elderly adults was 2,653, 3,932 and 5,012 per 100,000 population, respectively. The prevalence of neurological disorders among women (3,617) was higher compared with men (2,657). The prevalence rate in urban and rural populations was 2,190 and 4,070/1,00,000, respectively, implying that neurological disorders were twice as frequent in rural areas as in urban areas. The prevalence rates per 100,000 population of the most frequent disorders in the descending order of frequency were: headache (1,119), epilepsy (883), febrile convulsions (330), cerebrovascular disorder (150), and mental retardation (142). This large-scale population-based survey provides data that will be crucial for developing hospital and community-based neurological services in India and other developing countries. PMID- 15297792 TI - Relation between motorcycle helmet use and cervical spinal cord injury. AB - A case-control study was conducted to determine the relationship between motorcycle helmets, including type (i.e. partial or full coverage) and fastening status (i.e. loose or firm), and cervical spinal cord injuries (SCIs) in Taiwan. Based on a nationwide registry consisting of 396 motorcycle riders with traumatic SCIs during the 4-year period from July 1992 to June 1996, 229 subjects injured at cervical levels were defined as cases and 167 injured at other spinal levels as controls. Furthermore, additional information on the type and fastening status of the helmet was collected by telephone interview, and 147 subjects, including 79 with cervical and 68 with noncervical lesions, responded. Results of the logistic regression model show that neither different helmet types nor fastening status increased the occurrence of cervical SCIs when motorcycle riders were involved in severe crashes. PMID- 15297793 TI - Prevalence of inherited ataxias in the province of Padua, Italy. AB - Few population studies are available on epidemiological indexes of hereditary ataxias. An investigation on the prevalence rate of these movement disorders is in progress for the Veneto region, the main area of northeast Italy with a population of 4,490,586 inhabitants. The first results of this epidemiological survey concern the province of Padua, which numbers 845,203 residents (January 1, 2002). The prevalence rate of inherited ataxias has been estimated at 93.3 cases per million inhabitants. The most common types appeared to be the autosomal dominant forms, namely spinocerebellar ataxia type 1 and 2, with a prevalence of 24 per 1,000,000. In the same population, with a prevalence rate of 6 per 1,000,000, Friedreich's ataxia was defined as the prominent recessive autosomal form. There were very rare cases of ataxia telangiectasia, ataxia with vitamin E deficiency and cerebellar ataxia with congenital muscular dystrophy, a recently identified autosomal recessive disease. PMID- 15297794 TI - Incidence of myasthenia gravis in the province of Ferrara: a community-based study. AB - BACKGROUND: The reported annual incidence of myasthenia gravis (MG) ranges from 0.25 to 15 per million. The sex- and age-related pattern of disease incidence is still debated. METHODS: An intensive descriptive study was performed in the province of Ferrara (mean population 360,950 people) over the period 1985 through 2000. RESULTS: The average crude annual incidence rate was 2 per 100,000. We confirm a female preponderance in the total population, particularly in the youngest age groups. CONCLUSIONS: We observed an early increase in incidence in females, partly due to thymoma-associated MG, while MG without thymoma showed increasing incidence with age nonsignificantly different in the two sexes. PMID- 15297795 TI - Childhood infections as risk factors for multiple sclerosis: Belgrade case control study. AB - The aim of this case-control study was to analyze the role of childhood infections and vaccinations in patients with MS in the Belgrade population. The study group comprised 110 cases with definite MS according to Poser's criteria, in whom onset symptoms occurred up to 2 years prior to the interview. An equal number of controls, individually matched by sex, age and area of residence, was recruited from patients with various nonautoimmune neurological disorders. Measles (OR = 2.6, 95%CI 1.4-5.0), chickenpox (OR = 3.0, 95%CI 1.5-6.0), rubella (OR = 2.4, 95%CI 1.2-4.7), whooping cough (OR = 1.9, 95%CI 0.8-4.4), and mumps (OR = 1.8, 95%CI 0.8-4.5), at age < or = 7 years, were more frequently reported by MS cases. The total number of childhood viral infections (including measles, rubella, chickenpox, and mumps) at age < or = 7 years was significantly higher in MS cases than in controls (OR = 1.8, 95%CI 1.4-2.5). Concerning vaccinations, no statistically significant differences were found between groups. According to multivariate analysis, rubella (OR = 2.5, 95%CI 1.4-4.4, p = 0.001) and measles (OR = 2.4, 95%CI 1.3-4.3, p = 0.003) at age < or = 7 years were significantly related to MS. PMID- 15297796 TI - Buffalo metropolitan area and Erie County stroke study: rationale, design, and methods. AB - OBJECTIVES: The primary objective of this study was to define the incidence, disability, and death associated with stroke in the Buffalo metropolitan area and Erie County. This area has the highest stroke rate in New York State and therefore represents an ideal site to develop a successful model for prevention and management of stroke. DESIGN: A cross-sectional design to study all new and recurrent strokes that occurred in the calendar year 2000 in the geographical location of Buffalo metropolitan area and Erie County. PATIENTS AND DATA COLLECTED: A retrospective review of an estimated 5,000 patients with new stroke will be performed at regional hospitals and the coroner's office to determine the stroke subtypes, cerebrovascular risk factors, diagnostic investigations, treatment provided, and outcome. The total population residing in Buffalo in the year 2000 is available through the recent census. The study will also evaluate the quality of care provided for stroke patients including effectiveness of primary and secondary stroke prevention measures within this geographical region. CONCLUSIONS: We believe that this information will assist in allocation of resources and implementation of steps to improve stroke prevention and treatment. PMID- 15297797 TI - Racial disparities in subarachnoid hemorrhage mortality: Los Angeles County, California, 1985-1998. AB - We examined the racial distribution of subarachnoid hemorrhage (SAH) mortality in a unique multiracial community. Mortality rates for SAH among the residents of Los Angeles County were calculated from death certificate data (1985-1998). Residential postal zones were classified into three strata as a measure of socioeconomic status. The number of SAH deaths was 2,897. The age-adjusted SAH mortality rate was 1.9 in whites, 2.7 in Hispanics, 3.0 in Asians and 3.7 in blacks. In those younger than 70 years of age, the SAH mortality rate among blacks was 2.2 times that of whites and 1.8 times that of Hispanics and Asians. The SAH mortality rate declines after age 70 in blacks. The SAH mortality rate was higher in women than in men in all races and it was highest in elderly Asian women (23.5 per 100,000). An inverse relationship was observed between income and SAH mortality rates in all racial groups except whites. PMID- 15297799 TI - Door-to-door survey of major neurological diseases in rural Ecuador--the Atahualpa Project: methodological aspects. AB - We carried out a door-to-door survey to assess the prevalence, incidence, risk factors and possible etiology of major neurological diseases in Atahualpa, a rural village located in the coastal region of Ecuador. A total of 2,548 individuals were interviewed with standardized screening questionnaires to detect suspected cases of epilepsy, stroke and dementia, and they underwent a serum immunoblot test for the detection of anticysticercal antibodies (phase I). Positive individuals and a random sample of negative subjects were examined by neurologists (phase II). Then, patients with clinically confirmed disease and a control group of healthy subjects were further evaluated with complementary examinations (phase III). Here, we describe the methodology of this field study that may be used as a model for the evaluation of neurological disorders in rural communities of developing countries. PMID- 15297798 TI - Multiple sclerosis in southern Europe: Monreale City, Italy. A twenty-year follow up incidence and prevalence study. AB - BACKGROUND: Several follow-up studies showed increasing prevalence and incidence rates for multiple sclerosis (MS). OBJECTIVE: To ascertain, throughout a follow up study, the incidence and prevalence of MS in the city of Monreale, Sicily, southern Italy. METHODS: We calculated crude and age- and sex-specific prevalence rates on December 31, 2000, and determined incidence rates for the period January 1, 1992 to December 31, 2000. RESULTS: The prevalence of MS was 71.2 per 100,000 population (48,5/100,000 in men; 93,0/100,000 in women). The incidence rate of MS for the period 1992-2000 was 4.0/100,000 per year. CONCLUSION: This study showed a nonsignificant increase in MS incidence rates in Monreale city for 1992-2000 compared to 1981-1991. Prevalence rates were similar to those of the previous follow-up study. Intervals between onset of symptoms and diagnosis seemed shorter than in prior studies. There is no evidence that the high prevalence and incidence rates have changed in this interval but numbers are too small for firm statements. These findings indicate that in Monreale city MS prevalence is stable and confirm Sicily as a high-risk area for MS. PMID- 15297800 TI - Health and psychosocial status of patients with turner syndrome after transition to adulthood: the Belgian experience. AB - BACKGROUND: Most girls with Turner syndrome (TS) are intensively followed by paediatricians, but are lost to follow-up when they reach adulthood. To gain insight into the adult medical and psychosocial situation, we performed a survey in young adult TS patients. PATIENTS AND METHODS: A questionnaire concerning current health status, education, occupation and living situation was sent to 160 young adult TS women, all treated during childhood with GH and oestrogen if needed. RESULTS: We received 102 completed questionnaires. Mean +/- SD age at reception of the questionnaire was 23.4 +/- 3.3 years, height 153.3 +/- 5.2 cm, body mass index 23.7 +/- 4.9 kg/m(2). Age and auxological parameters were comparable between responders and non-responders. Thirteen (12.7%) responders were not under regular medical care; 15 (14.7%) were seen by a general practitioner, while 28 (27.4%) needed several specialists. Forty-one (40.2%) patients reported health problems. The most frequently reported problem was hypertension (10.7%), followed by hypothyroidism (5.8%) and back problems (4.9%). Twenty-four (23.5%) of the 41 patients were taking medication for the indicated health problems. Twenty-six (25.5%) women had undergone spontaneous puberty; 16 of them reported spontaneous menstruations while 10 received oestrogen replacement therapy. Of the 76 women with induced puberty, 11 (14.5%) were not taking any oestrogen anymore. Compared with the general population, more TS women attended university and more obtained higher education. Forty-six women (45.1%) were working full-time, 7 (6.9%) were unemployed, and 4 (3.9%) received an allocation. Seventy (68.6%) patients were still living with their parents, while 18 (17.6%) were living together or married, and 14 (13.7%) were living alone. CONCLUSIONS: The transition of adolescents with TS to adult medical care is not optimal in Belgium. Although 40.2% of these young women reported health problems, 12.7% did not consult any physician. Many TS women did not take oestrogen replacement therapy. A specialized multidisciplinary approach for adults with TS is needed in order to optimize health and psychosocial status in these women. PMID- 15297801 TI - Hypoglycemia-dependent beta2-adrenoceptor downregulation: a contributing factor to hypoglycemia unawareness in patients with Type-1 diabetes? AB - To evaluate the influence of the incidence and unawareness of hypoglycemia on lymphocyte beta2-adrenoceptor densities, we measured beta2-adrenoceptor density using [125I]-iodocyanopindolol and CGP 12177 before and after 1 week of treatment optimization in 33 adults with type-1 diabetes mellitus. Diabetes treatment of all patients was modified to improve their glycemic control. During this week, all patients had to complete a protocol with 7 daily glucose measurements, one of which was at night. The subjective symptoms were evaluated in case of hypoglycemia. A significant correlation between a hypoglycemia incidence below (but not above) the threshold of 2.75 mmol/l (50 mg/dl) and beta2-adrenoceptor densities on lymphocytes was found after the study week (r = -0.72, p < 0.00001). Nine patients suffering from hypoglycemia unawareness had a significantly higher incidence of hypoglycemia (p < 0.002) and lower beta2-adrenoceptor densities on lymphocytes compared to 24 patients who recognized all of their hypoglycemic episodes (p < 0.004). We conclude that downregulation of beta2-adrenoceptor densities on lymphocytes occurs as a result of recurrent hypoglycemia defined as glucose levels of < 2.75 mmol/l. Beta2-adrenoceptor densities are decreased in patients with subjective hypoglycemia unawareness and might contribute to the reduced beta-adrenergic sensitivity in this subgroup of patients. PMID- 15297802 TI - Assessment of hepatic glucose metabolism by indirect calorimetry in combination with a non-invasive technique using naturally enriched 13C glucose in healthy children and adolescents. AB - The metabolic fate of hepatic glucose can be best studied using invasive techniques such as tracer infusions and frequent blood sampling which have been revealed to be impractical in the pediatric age group. The aim of this study was to develop a non-invasive method based on indirect calorimetry and expired 13CO2 monitoring in order to gain insight into the mechanisms leading to impaired glucose tolerance in children and teenagers. As a first step, net glucose oxidation (NGO) and energy expenditure (EE) were measured in 47 subjects (range 7.5-17.3 years) of whom 18 were prepubertal (P1), 11 in early puberty (P2-P3) and 18 in late puberty (P4-P5) after 3-hourly loads of 180 mg/kg of oral maize glucose containing naturally enriched 13C. Isotope analysis allowed to calculate exogenous and endogenous glucose oxidation (EXGO, ENGO) and, hence, to derive TGS and NGS, that is glycogen turnover. NGO and EE decreased significantly with pubertal progression, reflecting higher metabolism at younger ages, whereas EXGO remained constant. TGS did not change significantly whereas NGS showed a significant negative correlation with pubertal progression: this can be explained by the fact that glycogenolysis exceeded glycogen synthesis in this experimental setting. This non-invasive method appears to be a promising tool to study the fate of hepatic glucose and therefore glycogen turnover in children at risk of developing glucose intolerance and/or type 2 diabetes. PMID- 15297803 TI - Compound heterozygous and homozygous mutations of the TSHbeta gene as a cause of congenital central hypothyroidism in Europe. AB - BACKGROUND: Thyroid hormones are crucial for normal growth and central nervous system development. In recent years, germline variants of the TSHbeta subunit gene have been identified as a cause of congenital TSH deficiency. METHODS: We performed a genetic and clinical study in children from four European countries diagnosed with congenital isolated central hypothyroidism. RESULTS: TSHbeta gene analysis revealed compound heterozygosity for 145C-->T (Q49X) and 313delT (C105Vfs114X) in 1 infant and homozygous mutation 313delT (C105Vfs114X) in 5 patients. Although all presented with typical symptoms of hypothyroidism, diagnosis and treatment was delayed until 3-5 months in 5 of 6 patients. In a longitudinal sibpair analysis, thyroxine substitution initiated immediately after birth was effective to prevent developmental delay and growth retardation. CONCLUSION: Clinical awareness is required to detect hypothyroidism due to TSHbeta mutations, which is not identified by TSH-based newborn screening. TSHbeta variants C105Vfs114X and Q49X are the most frequent cause of this severe disorder in Europe, now for the first time observed in compound heterozygous state. PMID- 15297804 TI - Do birth variable data predict melatonin production in 8- to 9-year-old children? Analysis of excreted 6-sulfatoxymelatonin. AB - HYPOTHESES: A cross-sectional study on urinary excretion of 6-sulfatoxymelatonin (aMT.6S) in young adults suggests a relation between melatonin production and body size at birth. As individual melatonin production remains stable during childhood and adolescence, this melatonin-birth size relation should also exist in children. METHODS: Daily urinary output of aMT.6S of 147 healthy white children (78 boys, 69 girls), 8 or 9 years of age, was quantified by ELISA and related to birth variable data. RESULTS: Contrary to expectation, aMT.6S output was not related to the ponderal index at birth but a moderate positive association with body mass index at the age of 8-9 years was seen. CONCLUSION: This study in children contradicts previous findings in adults. As no obvious reason can be identified for this discrepancy, further research (particularly a longitudinal study) is recommended to clarify whether birth variable data may predict melatonin production in certain circumstances during or after puberty. PMID- 15297805 TI - The reliability of haplotyping inference in nuclear families: misassignment rates for SNPs and microsatellites. AB - Single nucleotide polymorphisms (SNPs) are widely used when investigators try to map complex disease genes. Although biallelic SNP markers are less informative than microsatellite markers, one can increase their information content by using haplotypes. However, assigning haplotypes (i.e., assigning phase) correctly can be problematic in the presence of SNP heterozygosity. For example, a doubly heterozygous individual, with genotype 12, 12, could have haplotypes 1-1/2-2 or 1 2/2-1 with equal probability; in the absence of additional information, there is no way to determine which haplotype is correct. Thus an algorithm that assigns haplotypes to such an individual will assign the wrong one 50% of the time. We have studied the frequency of haplotype misassignments, i.e., haplotypes that are misassigned solely because of inherent marker ambiguity (not because of errors in genotyping or calculation). We examined both SNPs and microsatellite markers. We used the computer programs GENEHUNTER and SIMWALK to assign the haplotypes. We simulated (a) families with 1-5 children, (b) haplotypes involving different numbers of marker loci (3, 5, 7 and 10 loci, all in linkage equilibrium), and (c) different allele frequencies. Misassignment rates are highest (a) in small families, (b) with many SNP loci, and (c) for loci with the greatest heterozygosity (i.e., where both alleles have frequency 0.5). For example, for triads (i.e., one-child families with both parents genotyped), misassignment rates for SNPs can reach almost 50%. Family sizes of 4-5 children are required in order to ensure a misassignment frequency of < or = 5% for ten-SNP haplotypes with allele frequencies of 0.25-0.5. For microsatellites, a family size of at least 2-3 children is necessary to keep haplotyping misassignments < or = 5%. Finally, we point out that it is misleading for a computer program to yield haplotype assignments without indicating that they may have been misassigned, and we discuss the implications of these misassignments for association and linkage analysis. PMID- 15297806 TI - Association analysis of the plasminogen activator inhibitor-1 4G/5G polymorphism in Hispanics and African Americans: the IRAS family study. AB - OBJECTIVE: Plasminogen activator inhibitor type-1 (PAI-1) plays a central role in fibrolysis and has recently been hypothesized to influence components of the insulin resistance syndrome. We consider whether the 4G/5G polymorphism influences components of insulin resistance and obesity solely through PAI-1 protein levels or also though a secondary pathway. In addition, we explore whether transforming growth factor (TGF-beta1), a key regulator of PAI-1 expression, modifies the influence of the PAI-1 4G/5G polymorphism on these traits. METHODS AND RESULTS: The Insulin Resistance and Atherosclerosis (IRAS) Family Study genotyped 287 African American (18 pedigrees) and 811 Hispanic American (45 pedigrees) individuals for the 4G/5G PAI-1 and two TGF-beta1 polymorphisms (R25P, C-509T). Individuals were recruited from three clinical centers located in San Antonio (urban Hispanic), San Luis Valley (rural Hispanic) and Los Angeles (African American). The presence of the 4G PAI-1 allele was positively associated with PAI-1 protein level (combined sample p < 0.0001). Hispanic Americans average 65% higher PAI-1 protein levels than African Americans (p < 0.0001). Consistently across ethnic groups, increased PAI-1 protein levels were associated with increased insulin resistance and overall and central obesity (p value < 0.0001, combined sample). Adjusting for PAI-1 protein levels, there was evidence of an association of PAI-1 genotype (4G) with insulin sensitivity (p < 0.002) and subcutaneous fat (p < 0.01). These associations were not influenced by TGF-beta1 genotypes. CONCLUSIONS: PAI-1 protein is a strong correlate of insulin resistance (IR) and obesity in Hispanics and African Americans. However, PAI-1 4G/5G polymorphism appears to influence insulin resistance and obesity beyond its direct influence on serum PAI-1 protein levels. PMID- 15297807 TI - Comparative in vitro expression study of four Fabry disease causing mutations at glutamine 279 of the alpha-galactosidase A protein. AB - Fabry disease is an X-linked lysosomal storage disorder caused by the deficiency of alpha-galactosidase A that results in the accumulation of neutral sphingolipids. We report a novel point mutation in exon 6, Q279K, carried by an asymptomatic child with a family history of classic Fabry disease. Moreover, we comparatively study the in vitro expression and enzyme activity of Q279K and three other already described mutants in glutamine 279. The Q279K, Q279H and Q279R mutants transfected in COS-1 cells expressed no activity while the residual enzyme activity of the Q279E mutant represented 10% of wild type value. Western blot analysis demonstrated a differential behavior of the mutant proteins: Q279K and Q279H persisted as the inactive 50-kD precursor, indicating that these mutations may affect the normal processing of the enzyme, while the Q279R mutant was not detected probably due to an unstable protein which is rapidly degraded. The in vitro expression studies of the novel Q279K mutation were confirmed by Western blot analysis performed in the patient's lymphocytes which revealed the alpha-galactosidase A precursor of 50 kD but not the processed form. PMID- 15297808 TI - Evidence of admixture from haplotyping in an epidemiological study of UK Caucasian males: implications for association analyses. AB - OBJECTIVE: Cohort and case-control genetic association studies offer the greatest power to detect small genotypic influences on disease phenotypes, relative to family-based designs. However, genetic subdivisions could confound studies involving unrelated individuals, but the topic has been little investigated. We examined geographical and interallelic association of SNP and microsatellite haplotypes of the Y chromosome, of regions of chromosome 11, and of autosomal SNP genotypes relevant to cardiovascular risk traits in a UK-wide epidemiological survey. RESULTS: We show evidence (p = 0.00001) of the Danelaw history of the UK, marked by a two-fold excess of a Viking Y haplotype in central England. We also found evidence for a (different) single-centre geographical over-representation of one haplotype, both for APOC3-A4-A5 and for IGF2. The basis of this remains obscure but neither reflect genotyping error nor correlate with the phenotypic associations by centre of these markers. A panel of SNPs relevant to cardiovascular risks traits showed neither association with geographical location nor with Y haplotypes. CONCLUSION: Combinations of Y haplotyping, autosomal haplotyping, and genome-wide SNP typing, taken together with phenotypic2 associations, should improve epidemiological recognition and interpretation of possible confounding by genetic subdivision. PMID- 15297809 TI - Selecting tagging SNPs for association studies using power calculations from genotype data. AB - Recent studies have indicated that linkage disequilibrium (LD) between single nucleotide polymorphism (SNP) markers can be used to derive a reduced set of tagging SNPs (tSNPs) for genetic association studies. Previous strategies for identifying tSNPs have focused on LD measures or haplotype diversity, but the statistical power to detect disease-associated variants using tSNPs in genetic studies has not been fully characterized. We propose a new approach of selecting tSNPs based on determining the set of SNPs with the highest power to detect association. Two-locus genotype frequencies are used in the power calculations. To show utility, we applied this power method to a large number of SNPs that had been genotyped in Caucasian samples. We demonstrate that a significant reduction in genotyping efforts can be achieved although the reduction depends on genotypic relative risk, inheritance mode and the prevalence of disease in the human population. The tSNP sets identified by our method are remarkably robust to changes in the disease model when small relative risk and additive mode of inheritance are employed. We have also evaluated the ability of the method to detect unidentified SNPs. Our findings have important implications in applying tSNPs from different data sources in association studies. PMID- 15297810 TI - Human amniotic fluid glycoproteins expressing sialyl Lewis carbohydrate antigens stimulate progesterone production in human trophoblasts in vitro. AB - BACKGROUND: Progesterone is thought to mediate immune modulator effects by regulating uterine responsiveness. The aim of the study was to clarify the effect of transferrin and glycodelin A (former name PP14) as sialyl Lewis X-expressing glycoproteins on the release of progesterone by trophoblast cells in vitro. METHODS: Cytotrophoblast cells were prepared from human term placentas by standard dispersion of villous tissue followed by a Percoll gradient centrifugation step. Trophoblasts were incubated with varying concentrations (50 300 microg/ml) of human amniotic fluid- and serum-transferrin as well as with glycodelin A. Culture supernatants were assayed for progesterone, human chorionic gonadotropin (hCG) and cortisol by enzyme immunometric methods. RESULTS: The release of progesterone is increased in amniotic fluid transferrin- and glycodelin A-treated trophoblast cell cultures compared to untreated trophoblast cells. There is no relation between transferrin and the hCG or cortisol production of trophoblast cells. CONCLUSION: The results suggest that sialyl Lewis carbohydrate antigen-expressing amniotic fluid glycoproteins modulate the endocrine function of trophoblasts in culture by upregulating progesterone production. PMID- 15297811 TI - Withdrawal of article by the FDA after objection from Medtronic. PMID- 15297812 TI - Impact of endovascular repair on early outcomes of ruptured abdominal aortic aneurysms. AB - OBJECTIVE: Our purpose was to examine the impact of the introduction of endovascular treatment on the early outcomes of ruptured abdominal aortic aneurysms (AAAs) during 2 consecutive time periods at a single institution. METHODS: The hospital records of a single tertiary care center from 1997 to 2004 were retrospectively reviewed, and 36 consecutive patients who underwent treatment for acute ruptured AAA were identified. They were divided into 19 (53%) patients who were all treated with conventional open surgery from 1997 to 2001 (early) and 17 (47%) patients who were treated either with open (n = 4, 24%) or endovascular (n = 13, 76%) methods from 2002 to 2004 (late). All endovascular repairs were performed with commercially available bifurcated devices. Outcome measures included death, major complications, disposition at discharge (home or extended care facility), procedure time, blood loss, and hospital length of stay. RESULTS: Age, sex, and AAA size were similar between the 2 groups. Perioperative mortality in the early and late periods were 37% versus 12%, respectively (P =.13); rates of major complications were 84% versus 65%, respectively (P =.26); and discharge to home rather than extended care facility was 32% versus 59%, respectively (P =.18). Median procedure times (275 vs 149 minutes, P <.01), blood loss (3800 vs 138 mL, P <.0001), and length of stay (18 vs 6 days, P <.05) were all higher during the early period than in the late period. CONCLUSIONS: This preliminary study suggests that introduction of endovascular therapies may be potentially beneficial in the overall treatment scheme of patients with ruptured AAAs. However, longer follow-up and larger cohorts are needed to better establish its feasibility and efficacy compared with conventional open surgical repair. PMID- 15297813 TI - Effect of improved endograft design on outcome of endovascular aneurysm repair. AB - OBJECTIVE: This study was undertaken to identify factors that lead to improvements in the results of endovascular aneurysm repair, with particular focus on new endograft design. METHODS: We analyzed data for patients enrolled in the European Collaborators on Stent Graft Techniques for Abdominal Aortic Aneurysm Repair (EUROSTAR) registry, and compared those for endografts now withdrawn from the market with those for endografts currently in use. Patients in whom a variety of endograft types were used in small numbers were excluded. Postoperative and long-term outcomes were initially compared with univariate analyses, and subsequently multivariate tests were used to adjust for baseline differences between the 2 groups. The main outcome measures were freedom from a variety of secondary interventions, aneurysm rupture, and death. RESULTS: Some 1224 patients received "withdrawn" endografts, and 2768 patients received "current" endografts. The 2 groups were generally similar, but patients with current devices were more often men, significantly older, more frequently unfit for open surgery, and had larger aneurysms with wider necks. Of no surprise, current endografts were also more often used by experienced (>60 previous cases) surgical teams (44% vs 20%; P <.0001). Thirty-day clinical outcomes were comparable in the 2 groups, although patients with withdrawn devices were less likely to have type II endoleak (9.2% vs 5.5%; P <.0001), and those with current devices had a shorter mean hospital stay (5.4 vs 6.8 days; P <.0001). At 3 years more patients with current devices were free from secondary transfemoral intervention (88.4% vs 76%; P <.0001) and conversion to open repair (95.4% vs 93.4%; P =.007). Aneurysm-related mortality at 3 years, defined as death due to aneurysm rupture or within 30 days of a secondary intervention, was also less frequent with current endografts (2.7% vs 4.4%; P =.02). Aneurysm rupture at 3 years was infrequent (0.8% vs 1.8%; P =.07). At multivariate analysis the use of current devices was a protective factor against late conversion to open repair (hazard ratio, 0.49; 95% confidence interval, 0.28-0.86; P =.014) and aneurysm related death (hazard ratio, 0.51, 95% confidence interval, 0.34-0.75; P =.0008). Larger aneurysm or neck diameter and shorter neck length were also associated with late conversion to open repair; larger aneurysm diameter, older age, and unfitness for open surgery were predictive of aneurysm-related death. CONCLUSION: Modern endograft design has improved the results of endovascular aneurysm repair. PMID- 15297814 TI - Decreased use of iliac extensions and reduced graft junctions with software assisted centerline measurements in selection of endograft components for endovascular aneurysm repair. AB - OBJECTIVE: The purpose of this study was to determine the impact of using computerized software-assisted centerline measurements for extensions and graft junctions during the selection of endograft components for modular aortic endografts in endovascular repair of abdominal aortic aneurysms. METHODS: From April 1998 to December 2002, 289 modular aortic endografts were implanted at our institution. These included 248 grafts (prior to 2002, group 1) with components selected on the basis of manual caliper measurements from combined contrast computed tomography (CT) and marker-catheter arteriography data, and 41 grafts (2002, group 2) with components selected with the use of computerized software that allowed for centerline measurements on 3-dimensional reconstructions based on CT data. These 2 groups were compared for the number and type of extensions required per case. Seventeen other relevant variables were analyzed for their potential influence on selection of endograft components. These variables included age, gender, maximum aneurysm size, level of distal fixation, length and diameter at the fixation points, endograft manufacturer (make), and configuration. The significance of the observed differences was analyzed with a multivariate regression model, adjusting for potentially confounding preoperative measures. RESULTS: Multivariate analysis demonstrated that the number of right iliac extensions, left iliac extensions, total extensions, and total graft junctions was significantly reduced by the use of computerized software-assisted centerline measurements (group 2) compared with caliper measurements (group 1), independent of all other 17 preoperative variables. Notably, the mean number of required right iliac extensions was double in group 1 versus group 2. CONCLUSIONS: Centerline software-assisted measurements can significantly reduce the need for iliac extensions and, concomitantly, the number of required endograft junctions. On average, twice as many extensions were required for right iliac fixation when the manual caliper measurements were used compared with software-assisted measurements. These findings are highly relevant to issues of total endograft cost and long-term endograft integrity and focus attention on the tools that may need to be considered standards of care rather than optional for selection of endograft components. PMID- 15297815 TI - Complications of endovascular repair of high-risk and emergent descending thoracic aortic aneurysms and dissections. AB - PURPOSE: The advent of endovascular prostheses to treat descending thoracic aortic lesions offers an alternative approach in patients who are poor candidates for surgery. The development of this approach includes complications that are common to the endovascular treatment of abdominal aortic aneurysms and some that are unique to thoracic endografting. METHODS: We conducted a retrospective review of 60 emergent and high-risk patients with thoracic aortic aneurysms (TAAs) and dissections treated with endovascular prostheses over 4 years under existing investigational protocols or on an emergent compassionate use basis. RESULTS: Fifty-nine of the 60 patients received treatment, with one access failure. Thirty five patients received treatment of TAAs. Four of these procedures were performed emergently because of active hemorrhage. Twenty-four patients with aortic dissections (16 acute, 8 chronic) also received treatment. Eight of the patients with acute dissection had active hemorrhage at the time of treatment. Three devices were used: AneuRx (Medtronic; n = 31), Talent (Medtronic; n = 27), and Excluder (Gore; n = 1). Nineteen secondary endovascular procedures were performed in 14 patients. Most were secondary to endoleak (14 of 19), most commonly caused by modular separation of overlapping devices (n = 8). Other endoleaks included 4 proximal or distal type I leaks and 2 undefined endoleaks. The remaining secondary procedures were performed to treat recurrent dissection (n = 1), pseudoaneurysm enlargement (n = 3), and endovascular abdominal aortic aneurysm repair (n = 1). One patient underwent surgical repair of a retrograde ascending aortic dissection after endograft placement. Procedure-related mortality was 17% in the TAA group and 13% in the dissection group, including 2 acute retrograde dissections that resulted in death from cardiac tamponade. Overall mortality was 28% at 2-year follow-up. CONCLUSION: Although significant morbidity and mortality remain, endovascular repair of descending TAAs and dissections in patients at high-risk patients can be accomplished with acceptable outcomes compared with traditional open repair. The major cause for repeat intervention in these patients was endoleak, most commonly caused by device separation. Improved understanding of these complications may result in a decrease in secondary procedures, morbidity, and mortality in these patients. The need for secondary interventions in a significant number of patients underscores the necessity for continued surveillance. PMID- 15297816 TI - Twenty-year experience with acute distal thoracic aortic dissections. AB - BACKGROUND: There are few large studies in the literature that document the clinical outcome of an acute dissection of the distal thoracic aorta (ADDA), particularly since the advent of percutaneous techniques for therapeutic and prophylactic treatment of complications of ADDA. The goal of this study was to evaluate the outcome of ADDA with respect to medical, surgical, and percutaneous treatment over a 20-year period, and to use this information to estimate the benefit that future prophylactic therapy may yield. METHODS: The hospital records of all patients admitted with ADDA during the period of the study were reviewed retrospectively. RESULTS: There were 119 patients who fit the criteria of ADDA. Medical management was performed in 92 patients, with an overall mortality in this group of 13% (12/92 patients). Major morbidity occurred in 34 of the 83 surviving patients managed nonoperatively. Percutaneous interventions consisting of aortic fenestration and branch vessel stenting in 5 patients had a mortality rate of 40% and was only effective in the treatment of isolated renal artery malperfusion. Twenty-two patients underwent aortic surgery for complications or risk of impending rupture. Postoperative mortality was 18% (4/22 patients). Significant risk factors for death were rupture, acute renal failure, mesenteric ischemia, and age >70. No patient who had surgical fenestration required reoperation on the tailored segment. On the basis of clinical outcomes, we estimate that a maximum of 37% of patients could benefit acutely from prophylactic treatment of ADDA with aortic stent grafts, and an additional 13% could benefit chronically from such prophylactic treatment. CONCLUSIONS: ADDA remains a challenging clinical problem with many failures of medical, surgical, and percutaneous therapy. Surgery remains an effective therapeutic option in the treatment of complications of acute dissection of the distal thoracic aorta, and surgical aortic fenestration is a durable treatment for malperfusion. A minority of patients may benefit from prophylactic treatment of ADDA with thoracic stent grafts. PMID- 15297817 TI - Surgical management of peripancreatic arterial aneurysms. AB - Peripancreatic artery aneurysms--gastroduodenal (GDA) and pancreaticoduodenal (PDA)--are highly unusual. We report 4 such aneurysms and have collated reports of true peripancreatic artery aneurysms based on an extensive review of the English literature. From this review, patient characteristics, clinical behavior, outcome and management strategies are assessed. PMID- 15297818 TI - Radiotherapy-induced supra-aortic trunk disease: early and long-term results of surgical and endovascular reconstruction. AB - PURPOSE: Few articles have dealt specifically with management of radiotherapy induced supra-aortic trunk disease. We investigated the results of surgical and endovascular treatment of these lesions, and present our findings in a large series of patients. METHODS: The study was conducted at 11 centers. Over 10 years 64 patients with radiotherapy-induced supra-aortic trunk disease underwent surgical or endovascular treatment. Data were collected retrospectively in a consecutive cohort of patients, and were analyzed with the Kaplan-Meier method. RESULTS: Mean patient age was 64.4 years. The indications for radiotherapy included breast cancer (30%), head and neck malignancies (50%), and lymphomas (19%). The mean interval between irradiation and arterial revascularization was 15.2 years. Thirteen of the 64 patients (20%) had asymptomatic disease, and 51 patients (80%) had symptomatic disease. Ninety-two stenotic or occlusive lesions were observed, which involved the common carotid artery (n = 62), the subclavian artery (n = 26), or the innominate artery (n = 4). Twenty-three patients (36%) had multiple supra-aortic trunk lesions, but only 8 patients underwent reconstruction of multiple supra-aortic trunks. Five patients (8%) underwent sternotomy for revascularization from the ascending aorta. Forty-seven patients required revascularization of a common carotid artery; procedures included bypass grafting (n = 30), angioplasty with stent placement (n = 13), carotid-carotid transposition (n = 2), and endarterectomy (n = 2). Fifteen patients underwent restoration of a subclavian artery. One patient died on postoperative day 5, of stroke after early occlusion of an intercarotid crossover bypass graft. Mean follow-up was 37 months (range, 2-120 months). Ten late deaths occurred during follow-up. The probability of survival at 4 years was 78.1% +/- 8.6%. During follow-up, 6 patients had stroke, 4 bypass occlusions occurred and 3 stenoses occurred in the revascularized arteries. At 4 years the probability of freedom from stroke was 85% +/- 8.8%. At 4 years the primary patency rate was 79.3% +/- 8.5% and the secondary patency rate was 87.9% +/- 7.2%. CONCLUSIONS: In light of the context, the results of arterial revascularization to treat radiation-induced arterial lesions of the supra-aortic trunk are satisfactory. PMID- 15297819 TI - Degree of carotid plaque calcification in relation to symptomatic outcome and plaque inflammation. AB - OBJECTIVE: We undertook this study to quantitate differences in the degree of calcification between symptomatic and asymptomatic plaques removed at carotid endarterectomy (CEA) and to determine associated extent of plaque macrophage infiltration, a histopathologic feature of plaque instability. METHODS: CEA plaques (n = 48) were imaged at 1.25-mm intervals with spiral computed tomography (CT; 10-15 images per plaque). Indications for CEA were transient ischemic attack (n = 16), stroke (n = 5), amaurosis (n = 4), and critical asymptomatic stenosis (n = 23). The percent area calcification for each plaque was determined in spiral CT serial sections and averaged for each plaque. In 31 of 48 plaques macrophage infiltration was quantitated in corresponding histologic sections with immunohistochemical techniques. RESULTS: The mean (+/- SD) age of patients with symptomatic and asymptomatic plaques was 66 +/- 7 years vs 71 +/- 7 years, respectively, and degree of stenosis was 76% versus 82%, respectively (P =.05). Atherosclerosis risk factors were similar between groups. Percent plaque area calcification was twofold greater in asymptomatic versus symptomatic plaques (48% +/- 19% vs 24% +/- 20%, respectively; P <.05). At receiver operating characteristic curve analysis, 80% of symptomatic plaques were below and 87% of asymptomatic plaques were above a cutoff point of 30% plaque area calcification. Macrophage burden was greater in the symptomatic plaques than in the asymptomatic plaques (52% vs 23%; P <.03). A strong inverse relationship between the degree of plaque calcification and macrophage infiltration was found in critical carotid stenoses (r = -0.87; P <.001). CONCLUSIONS: Symptomatic plaques are less calcified and more inflamed than asymptomatic plaques. Regardless of clinical outcome, a strong inverse correlation was found between the extent of carotid plaque calcification and the intensity of plaque fibrous cap inflammation as determined by the degree of macrophage infiltration. Carotid plaque calcification is associated with plaque stability, and is a potential spiral CT in vivo quantitative marker for cerebrovascular ischemic event risk. PMID- 15297820 TI - Durability of carotid endarterectomy for treatment of symptomatic and asymptomatic stenoses. AB - PURPOSE: Although many studies have well established that carotid endarterectomy (CEA) is beneficial in selected patients with severe carotid disease, only a few large studies have focused on the durability of the surgical procedure. Carotid artery angioplasty and stenting (CAS) has recently been proposed as a potential alternative to CEA. We analyzed the incidence of late occlusion and recurrent stenosis after CEA. METHODS: Over 13 years 1000 patients underwent 1150 CEA procedures to treat symptomatic and asymptomatic high-grade carotid stenosis. CEA procedures involving either traditional CEA with patching (n = 302) or eversion CEA (n = 848) were all performed by the same surgeon, with patients under deep general anesthesia and cerebral protection involving continuous electroencephalographic monitoring for selective shunting. All patients underwent postoperative duplex ultrasound scanning and clinical follow-up at 1, 6, and 12 months, and yearly thereafter. New neurologic events, late occlusions, and recurrent stenoses 50% or greater were recorded. Complete follow-up (mean, 6.2 years; range, 6-156 months) was obtained in 95% of patients (949 of 1000), for an overall average of 95% of procedures (1092 of 1150). Survival analysis was performed with the Kaplan-Meier life table method. RESULTS: Perioperative (30 day) mortality rate was 0.3% (3 of 1000), and stroke rate was 0.9% (11 of 1150), with a combined mortality and stroke rate of 1.2%. The incidence of late occlusion and recurrent stenosis 70% or greater was 0.6% and 0.5%, respectively, with a combined occlusion and restenosis rate of 1.1%. Kaplan-Meier analysis showed that the rate of freedom from occlusion, restenosis 70% or greater, and combined occlusion and restenosis 70% or greater at 12 years was 99,4%, 99.5%, and 98.8%, respectively. Occlusion and restenosis developed asymptomatically. CONCLUSIONS: CEA is a low-risk procedure for treating severe symptomatic and asymptomatic carotid disease, with excellent long-term durability. Proponents of CAS should bear this in mind before considering CAS as a routine alternative to CEA. PMID- 15297821 TI - Ocular findings as predictors of carotid artery occlusive disease: is carotid imaging justified? AB - OBJECTIVES: Hemispheric neurologic symptoms, amaurosis fugax, and Hollenhorst plaques at eye examination are standard indications for carotid imaging to identify carotid artery occlusive disease (CAOD). Previous reports have suggested that other ocular findings, such as retinal artery occlusion and anterior ischemic optic neuropathy, are associated with CAOD. However, the predictive value of ocular findings for the presence of CAOD is controversial. The purpose of this study was to define the predictive value of ocular symptoms and ophthalmologic examination in identifying significant CAOD. METHODS: Over 3 years 145 patients were referred for carotid imaging on the basis of ocular indications in 160 eyes. Forty patients were excluded because of concurrent non-ocular indications for carotid imaging, leaving 105 patients referred exclusively for ocular indications to evaluate. Ophthalmologic history and eye examination were correlated with carotid duplex ultrasound findings. RESULTS: Amaurosis fugax was associated with a positive scan in 20.0% of carotid arteries (P =.022). Hollenhorst plaques at fundoscopic examination were associated with a positive scan in 18.2% of carotid arteries (P =.02). Ocular findings exclusive of Hollenhorst plaques were particularly poor predictors of CAOD, inasmuch as only 1 of 64 arteries (1.6%) had significant ipsilateral internal carotid artery stenosis (P =.022). Venous stasis retinopathy was the only ocular finding other than Hollenhorst plaques with any predictive value (1 of 5 scans positive; positive predictive value, 20.0%). CONCLUSIONS: Ocular symptoms and findings are poor predictors of CAOD. Amaurosis fugax, Hollenhorst plaques, and venous stasis retinopathy demonstrated moderate predictive value, whereas all other ocular findings demonstrated no predictive value in identifying CAOD. PMID- 15297822 TI - The use of the Angioseal device for femoral artery closure. AB - BACKGROUND: As vascular surgeons perform increasing numbers of percutaneous catheter-based procedures, they will need to become familiar with varying methods of femoral artery closure. Few studies on closure devices have included significant numbers of patients with peripheral arterial disease. It is the purpose of this study to determine whether the Angioseal device (St. Jude Medical) is a satisfactory method of achieving femoral artery puncture site hemostasis in these patients. METHODS: The records of all patients undergoing Angioseal closure of femoral artery puncture by a single vascular surgeon were reviewed. Indication, type of intervention, and size of the vascular sheath employed were recorded. Times to mobilization and discharge were determined. Patients were seen before discharge and 1, 4, and 12 weeks after the procedure, and at 3- month intervals thereafter. Complications including hemorrhage, pseudoaneurysm, infection, and vessel occlusion were noted. RESULTS: Between February 1, 2002, and August 31, 2003, 220 Angioseal collagen plugs were deployed in 188 patients. Attempts were made to deploy Angioseal devices in 92% of patients undergoing percutaneous procedures during this time period. One hundred forty-four procedures were diagnostic and 74 procedures included 76 therapeutic interventions. One hundred forty-four 5F sheaths, 47 6F sheaths, and 29 7F sheaths were employed during the procedures. Time to mobilization and discharge was 58 +/- 19 minutes and 102 +/- 31 minutes in patients undergoing diagnostic studies, 68 +/- 22 minutes and 146 +/- 42 minutes following interventions using 6F sheaths, and 127 +/- 18 minutes and 219 +/- 37 minutes when interventions were performed using 7F sheaths. No patient developed a major hematoma or infection following the use of an Angioseal. There were 2 complications related to device deployment. One patient developed a 1.4-cm false aneurysm at the femoral artery puncture site that resolved spontaneously. A second patient required operation for vessel occlusion when an Angioseal was deployed in a markedly diseased femoral artery. These adverse events occurred early in our experience. CONCLUSIONS: The Angioseal provides a secure method of achieving hemostasis following femoral artery puncture. It is easy to deploy, has a high level of patient satisfaction, and allows for early ambulation and hospital discharge. When simple guidelines are observed, the device can be safely used in the great majority of patients with peripheral vascular disease. It offers considerable advantages over the traditional method of manual compression. PMID- 15297823 TI - Acute arterial thrombosis associated with cocaine abuse. AB - PURPOSE: Cocaine-induced arterial thrombosis is uncommon, and most reported cases involved small-diameter vessels such as the cerebral and coronary arteries. This study was undertaken to review our experience with peripheral arterial thrombosis presumed caused by cocaine abuse. METHODS: Hospital records were reviewed for all patients admitted over 10 years with acute arterial occlusion involving the peripheral arterial system. Patients with confirmation of cocaine use or of its derivative, crack cocaine, within 24 hours of hospital admission formed the basis of this study. Symptoms at presentation, management, and outcome in these patients were reviewed. RESULTS: Three hundred eighty-two patients with acute peripheral arterial occlusion were identified during the study period. The presumptive diagnosis of cocaine-induced arterial occlusion was made in 5 patients (4 men, mean age 38 years). Cocaine use was achieved via intranasal inhalation in 2 patients (40%), whereas the 3 remaining patients smoked crack cocaine (60%). The mean time between cocaine use and onset of arterial thrombosis was 9.2 hours (range, 2-20 hours). Symptoms at presentation included acute limb ischemia without pedal Doppler signals (3 patients, 60%) and abdominal pain without femoral pulses (2 patients, 40%). Arterial occlusion was confirmed on angiograms in all patients, which revealed aortic thrombosis in 1 patient (20%), iliac thrombosis in 2 patients (40%), superficial femoral artery thrombosis in 1 patient (20%), and popliteal artery occlusion in 1 patient (20%). Surgical thrombectomy was successfully performed in 4 patients (80%), and 1 patient (20%) underwent successful thrombolytic therapy for femoropopliteal artery occlusion. There was no perioperative mortality. All 5 patients who were discharged were available for follow-up (mean, 36 months; range, 6-75 months). There was 1 late death from myocardial infarction. In 1 patient recurrent lower extremity arterial thrombosis developed after 28 months, which was successfully treated with thrombolytic therapy. CONCLUSIONS: Our study underscores cocaine abuse as a potential cause of acute arterial thrombosis. Cocaine-induced arterial thrombosis should be suspected in patients with recent history of cocaine abuse with acute limb ischemia without an identifiable source or overt cardiovascular risk factors. Prompt angiography with operative or endovascular intervention should be performed to avert arterial ischemic sequelae. PMID- 15297825 TI - Neovascularization and recurrent varicose veins: more histologic and ultrasound evidence. AB - BACKGROUND: The recurrence of varicose veins is a common and costly consequence of varicose vein surgery. Despite the long history and vast experience of varicose vein surgery, the exact cause of recurrence is still unknown. This study aims to investigate the cause of recurrence further by correlating findings from duplex ultrasound scans, resin casts, and histologic investigation at the recurrence of the saphenofemoral junction. In particular, frequency and neovascularization are evaluated. METHOD: Forty-nine saphenofemoral junctions (SFJs) from 42 patients who presented for re-operation on their varicose veins were examined with duplex ultrasound and physiologic air plethysmography tests before surgery. All patients had reflux at the groin for which surgery was carried out. Specimens taken during surgery were sectioned and stained for conventional histology and immunohistology, and 5 specimens were infused with resin to form a cast of the venous vasculature. RESULTS: All but 3 re-operation specimens (94%) showed multiple vessels at the stump site of the previous SFJ ligation. Neovascular channels of variable size, number, and tortuosity accounted for the ultrasound appearances and reflux to recurrent varicosities in the vast majority of specimens. These new vessels connected to the common femoral vein at the site of the previous SFJ. In 2 incompetent junctions without femoral vein involvement, while small vessels were seen surrounding the femoral stump scar, ultrasound and histology confirmed both neovascular and residual (enlarged collateral) connections from epigastric and pudendal vessels into the thigh. CONCLUSION: Neovascularisation is the major cause for ultrasound-confirmed recurrence of reflux in the groin following varicose vein surgery. PMID- 15297826 TI - Hemodynamic and clinical impact of ultrasound-derived venous reflux parameters. AB - PURPOSE: This study was undertaken to assess which ultrasound-derived parameter was superior for measuring venous reflux quantitatively and to evaluate the importance of popliteal vein valve reflux. PATIENTS AND METHODS: A retrospective analysis was performed of 244 refluxive limbs in 182 patients who underwent ultrasound scanning, venous pressure measurement, air plethysmography, and clinical classification of severity according to the CEAP score. Reflux time (RT, s), peak reflux velocity (PRV, m/s), time of average rate of reflux (TAF, mL/min), absolute displaced volume retrogradely (ADV, mL) were compared to clinical class, ambulatory venous pressure (% drop), venous filling time (s), and venous filling index (mL/s) using nonparametric statistical tests. A P value of <.05 was considered significant. Limbs were divided into 3 groups: (A) axial great saphenous vein reflux only (n = 68); (B) axial deep reflux including popliteal vein incompetence with or without concomitant gastrocnemius or great or small saphenous vein reflux (all ultrasound reflux parameters of each refluxive vein added at the knee level) (n = 79); and (C) all limbs with popliteal vein reflux (the ultrasound data of the refluxive popliteal vein exclusively was used in comparison regardless of concomitant associated reflux) (n = 103). Limbs were also stratified into limbs with skin changes and ulcer (C-class 4-6) and those without (C-class 1-3) and subsequently compared. RESULTS: No meaningful significant correlation was found between RT and the clinical and hemodynamic results in groups A and B. The PRV and TAF correlated significantly with the hemodynamic parameters. The PRV and TAF and clinical severity trended towards correlation in group A (P =.0554 and P =.0998, respectively), but was significantly correlated in group B. The poor hemodynamic condition in the subset of C-class 4-6 limbs in groups A and B was reflected in a greater PRV, TAF, and ADV in this subset as compared with the limbs in C-class 1-3. RT was not significantly different in the subsets of limbs, further suggesting that RT is not related to hemodynamic or clinical state of the limbs. No meaningful correlations were found in group C. Although the hemodynamic data were significantly poorer in the subset of limbs with C-class 4-6 than in C-class 1-3, the ultrasound-derived parameters were not significantly different. CONCLUSION: The duration of valve reflux time (or valve closure time) cannot be used to quantify severity of reflux and is purely a qualitative measurement. The PRV and the rate of reflux appeared to better reflect the magnitude of venous incompetence. In the presence of axial reflux, it appeared logical and physiologically correct to sum up these reflux parameters for each venous segment crossing the knee. The popliteal valve reflux (the "gatekeeper" function) was not in itself an important determinant of venous hemodynamics and clinical severity. Additional reflux in other venous segments must be taken into account. PMID- 15297827 TI - Outcome after autogenous brachial-axillary translocated superficial femoropopliteal vein hemodialysis access. AB - OBJECTIVE: The optimal configuration for patients with "complex" or "tertiary" hemodialysis access needs remains undefined. This study was designed to examine the utility of the autogenous brachial-axillary translocated superficial femoropopliteal vein access (SFV ACCESS) in this subset of patients. METHODS: Patients presenting for permanent hemodialysis access without a suitable upper extremity vein for autogenous access identified by duplex ultrasound mapping and those with repeated prosthetic access failures were considered candidates for SFV ACCESS. Ankle-brachial indices were obtained, and duplex scanning of the superficial femoropopliteal and saphenous veins was performed. Patients deemed candidates for SFV ACCESS also underwent preoperative upper extremity arteriography and venography. A retrospective review of the complete medical record was performed, and a follow-up telephone or personal interview was conducted. RESULTS: Thirty patients (mean age +/- SD, 54 +/- 15 years; male, 33%; white, 37%; with diabetes, 50%; obese, 21%) underwent SFV ACCESS among approximately 650 access-related open surgical procedures during the study period. The patients had been receiving dialysis for 4 +/- 5 years (range, 0-24 years), and had 3 +/- 3 (range, 0-17) prior permanent accesses, whereas 90% were actively dialyzed through tunneled catheters. In-hospital 30-day mortality was 3%, and the hospital length of stay was 7 +/- 7 days. Fifty-seven percent of the patients experienced some type of perioperative complication, and 38% required a remedial surgical procedure. Hand ischemia developed in 43% of the patients (severity grade: 1, 10%; 2, 7%; 3, 27%), and a distal revascularization, interval ligation was performed in all those with grade 3 ischemia. Thigh wound complications or hematomas developed in 23% of the patients, and arm wound complications or hematomas developed in 17%. The incidence of thigh wound complications was significantly greater (57% vs 9%; P =.03) in obese patients, but the other perioperative complications analyzed could not be predicted on the basis of age, gender, or comorbid conditions. The SFV ACCESS was cannulated 7 +/- 1 weeks postoperatively. The primary, primary assisted, and secondary patency rates were 96% +/- 4%, 100% +/- 0%, and 100% +/- 0%, respectively, at 6 months; 79% +/- 8%, 91% +/- 6%, and 100% +/- 0%, respectively, at 12 months; and 67% +/- 13%, 86% +/- 9%, and 100% +/- 0%, respectively, at 18 months (life table analysis; % +/- SE). CONCLUSIONS: The intermediate term functional patency rate after SFV ACCESS is excellent, although the magnitude of the procedure and the complication rate are significant. SFV ACCESS should only be considered in patients with limited access options. PMID- 15297828 TI - Randomized comparison of 6-mm straight grafts versus 6- to 8-mm tapered grafts for brachial-axillary dialysis access. AB - OBJECTIVE: This report presents the results of a prospective randomized study that compared 2 grafts of different diameter: 6 mm, and 8 mm tapered to 6 mm at the arterial site, placed in the upper arm for hemodialysis in a selected population of patients younger than 71 years without diabetes. METHODS: Seventy consecutive patients younger than 71 years without diabetes who required an upper arm graft between January 1997 and January 2002 and without previous access in the same limb were randomly allocated to receive either a 6-mm graft or 6- to 8 mm graft. Graft flow was measured every 3 months with the Doppler dilution technique. When access flow was less than 600 mL/min, fistulography was performed, and any stenosis was surgically treated with venous outflow replacement. Thrombectomy and associated stenosis treatment in the same stage was performed in all cases immediately after detection of thrombosis. Complication rate, and primary, assisted primary, and secondary patency rates were compared between the two groups with the Student t test and life table analysis. RESULTS: Mean access flow was 975 mL/min for 6-mm grafts (range, 600-1500 mL/min; 95% confidence interval [CI], 889-1070), and for 6- to 8-mm grafts was 1397 mL/min (range, 1122-2700 mL/min; 95% CI, 1122-1672). This difference was significant (P <.01). Complication rate was 0.45 episodes per graft-year in 6-mm grafts, and 0.19 episodes per graft-year in 6- to 8-mm grafts (P <.01). At 1, 2, and 3 years, primary patency rates were 62%, 58%, and 44%, respectively, for 6-mm grafts, and 85%, 78%, and 73% for 6- to 8-mm grafts; log-rank comparison between curves was P =.0259. At 1, 2, and 3 years, secondary patency rates were 85%, 85%, and 85%, respectively, for 6-mm grafts, and 90%, 90%, and 90% for 6- to 8-mm grafts; log rank comparison between curves was not significant, at P =.0603. At 1, 2, and 3 years, assisted primary patency rates were 84%, 79%, and 76%, respectively, for 6 mm grafts, and 90% for 6- to 8-mm grafts; log-rank comparison was P =.0414. CONCLUSIONS: The results of this study show an advantage in terms of primary and assisted primary patency rates, and complication rate for upper arm grafts with diameter 6 mm to 8 mm over grafts with 6-mm diameter in a patient population younger than 70 years without diabetes. The finding of a similar secondary patency rate in both groups is probably due to the surveillance program with sequential measurement of access flow and prompt surgical treatment of stenosis. However, we needed twice the number of rescue procedures in 6-mm grafts to achieve a similar patency rate as with large-bore grafts. These study results must be carefully evaluated, taking into consideration the small number of patients and the selected patient population. PMID- 15297829 TI - Activation of fibrinolytic pathways is associated with duration of supraceliac aortic cross-clamping. AB - PURPOSE: The cause of the coagulopathy seen with supraceliac aortic cross clamping (SC AXC) is unclear. SC AXC for 30 minutes results in both clotting factor consumption and activation of fibrinolytic pathways. This study was undertaken to define the hemostatic alterations that occur with longer intervals of SC AXC. METHODS: Seven pigs underwent SC AXC for 60 minutes. Five pigs that underwent infrarenal aortic cross-clamping (IR AXC) for 60 minutes and 11 pigs that underwent SC AXC for 30 minutes served as controls. No heparin was used. Blood samples were drawn at baseline, 5 minutes before release of the aortic clamp, and 5, 30, and 60 minutes after unclamping. Prothrombin time, partial thromboplastin time, platelet count, and fibrinogen concentration were measured as basic tests of hemostatic function. Thrombin-antithrombin complexes were used to detect the presence of intravascular thrombosis. Fibrinolytic pathway activation was assessed with levels of tissue plasminogen activator antigen and tissue plasminogen activator activity, plasminogen activator inhibitor-1 activity, and alpha2-antiplasmin activity. Statistical analysis was performed with the Student t test and repeated measures of analysis of variance. RESULTS: Prothrombin time, partial thromboplastin time, and platelet count did not differ between groups at any time. Fibrinogen concentration decreased 5 minutes (P =.005) and 30 minutes (P =.006) after unclamping in both SC AXC groups, but did not change in the IR AXC group. Thrombin-antithrombin complexes increased in both SC AXC groups, but were not significantly greater than in the IR AXC group. SC AXC for both 30 and 60 minutes produced a significant increase in tissue plasminogen activator antigen during clamping and 5 minutes after clamping. This increase persisted for 30 and 60 minutes after clamp release in the 60-minute SC AXC group. Tissue plasminogen activator activity, however, increased only in the 60-min SC AXC group during clamping (P =.02), and 5 minutes (P =.05) and 30 minutes (P =.06) after unclamping, compared with both control groups. CONCLUSIONS: Thirty and 60 minutes of SC AXC results in similar degrees of intravascular thrombosis and fibrinogen depletion. Although SC AXC for both 30 and 60 minutes leads to activation of fibrinolytic pathways, only 60 minutes of SC AXC actually induces a fibrinolytic state. Fibrinolysis appears to be an important component of the coagulopathy associated with SC AXC, and is related to the duration of aortic clamping. CLINICAL RELEVANCE: The coagulopathy frequently associated with thoracoabdominal aortic aneurysm repair is thought to revolt visceral ischemia-reperfusion. The nature of this coagulopathy is controversial. The current study demonstrates that the major hemostatic alteration associated with supraceliac aortic cross-clamping is activation of fibrinolytic pathways. The magnitude of this fibrinolytic response is directly related to the duration of supraceliac aortic occlusion. Future efforts to treat this coagulopathy may well include judicious use of autofibrinolytic agents. PMID- 15297830 TI - Rapamycin suppresses experimental aortic aneurysm growth. AB - OBJECTIVE: Inflammatory modulators are important in the pathogenesis of aneurysmal disease. Gene expression profiling of experimental abdominal aortic aneurysm (AAA) tissue demonstrated upregulation of the FK506BP12 (rapamycin binding protein) gene product. Rapamycin is a potent immunosuppressor that prevents recurrent stenosis. However, its effect on aneurysm formation has not been studied. We therefore examined the effect of rapamycin in an experimental rat AAA model. METHODS: Adult male Wistar rats underwent elastase infusion into isolated infrarenal aortas to create experimental aneurysms. Rats were randomized to receive either rapamycin or placebo via gastric lavage daily starting on the day of surgery. On postoperative day 7 the aneurysm was measured, the infrarenal aorta was harvested, and the rats were euthanized. NF kappa B was measured with Western blotting as a marker of inflammation. Matrix metalloproteinase (MMP)-9 protein levels were measured. Hematoxylin-eosin and elastin staining were used to examine tissue inflammation and elastin preservation. RESULTS: Aneurysms were significantly smaller in diameter in the rapamycin-treated group (3.3 +/- 0.7 mm vs 4.5 +/- 0.5 mm; P <.0001). NF kappa B levels were significantly reduced by 64% +/- 14% in rapamycin-treated aortas (P =.023). MMP-9 was reduced in rapamycin treated aortas by 54% +/- 22% (P =.043). Hematoxylin-eosin and elastin staining showed no changes in inflammatory infiltrate or degradation of elastin fibers in elastase-infused aortic segments in rapamycin-treated rats. CONCLUSION: Rapamycin significantly reduces the rate of aneurysm expansion in the experimental AAA rat model by 40%. Biochemical evidence suggests that this is related to suppression of inflammatory signaling and decreased MMP-9 levels. Rapamycin could provide a new treatment for small aneurysms. CLINICAL RELEVANCE: Human aortic aneurysms are characterized histologically by an inflammatory infiltrate with severe proteolytic destruction. Rapamycin is an immunosuppressive agent commonly used to control transplant rejection and intimal hyperplasia by modulating the inflammatory cascade. In this experimental model rapamycin suppressed aneurysm expansion, decreased NF kappa B activation (a marker of inflammation), and decreased matrix metalloproteinase-9 levels. It is hoped that rapamycin or other similar anti-inflammatory drugs will one day be able to control aneurysm expansion in patients PMID- 15297831 TI - Type II endoleak in porcine model of abdominal aortic aneurysm. AB - PURPOSE: The purpose of this study was to develop a reliable in vivo porcine model of type II endoleak resulting from endovascular aortic aneurysm repair (EVAR), for the study and treatment of type II endoleak. METHODS: Eight pigs underwent creation of an infrarenal aortic aneurysm, with a Dacron patch with preservation of lumbar branches. An indwelling pressure transducer was placed in the aneurysm sac. After 1 week the animals underwent EVAR with a custom-made Talent endograft. After another week the animals underwent laparoscopic lumbar artery ligation. Abdominal and pelvic computed tomography was performed after each procedure. Aneurysm sac pressure was measured in sedated and awake animals. RESULTS: All eight animals underwent successful creation of an aortic aneurysm and EVAR resulting in exclusion of the aneurysm sac. After creation of the aneurysm the sac mean arterial pressure (MAP) was 72.5 +/- 6.1 mm Hg and the sac pulse pressure was 44.8 +/- 8.7 mm Hg. Postoperative computed tomography scans demonstrated a type II endoleak from the lumbar branches in all animals. While aneurysm sac MAP (56.5 +/- 7.9 mm Hg; P <.01) and pulse pressure (13.6 +/- 4.1 mm Hg; P <.01) decreased after EVAR, sac pulse pressure remained, with type II endoleak. All animals underwent laparoscopic lumbar artery ligation, which resulted in further reduction in the sac MAP (38.3 +/- 4.6 mm Hg; P <.02) and immediate absence of sac pulse pressure (0 mm Hg; P <.01). Necropsy confirmed the absence of collateral flow in the aneurysm sac, with fresh thrombus formation in all animals. CONCLUSION: We present a reliable and clinically relevant in vivo large animal model of type II endoleak. CLINICAL RELEVANCE: We set out to show that aortic aneurysm sac pressurization caused by lumbar arterial flow in the setting of type II endoleak can be reproduced in an in vivo porcine model of endovascular aortic aneurysm repair. Indeed, in this model the aneurysm sac pulse pressure was a sensitive indicator of type II endoleak, correlating well with findings at computed tomography, and lumbar artery ligation eliminated the endoleak, as demonstrated on computed tomography scans and sac pressure measurement. Therefore we believe this in vivo large animal model can be instrumental in the study of many aspects of the physiologic features of type II endoleak. PMID- 15297832 TI - Wall shear modulation of cytokines in early vein grafts. AB - OBJECTIVE: Pro-inflammatory cytokine-driven mechanisms have been implicated in vein graft failure, though little is known about the effect of hemodynamic factors and anti-inflammatory counter-regulatory mechanisms. We hypothesized that early temporal expression of the pro-inflammatory cytokine interleukin (IL)-1 beta and the anti-inflammatory cytokine IL-10 proceeds by way of wall shear stress-dependent pathways in the arterializing vein graft. METHODS: Rabbits (n = 27) underwent bilateral jugular vein carotid interposition grafts, and simultaneous unilateral distal carotid branch ligation, to produce both low-flow and high-flow grafts in the same animal. Vein grafts were harvested at 1, 3, 7, 14, and 28 days and were assessed for architecture, wall shear stress, and cytokine messenger RNA levels (quantitative real-time two-step reverse transcription polymerase chain reaction). RESULTS: The model resulted in an immediate 90% flow reduction (P <.001, paired t test) in the vein graft on the ligated side, and a 36% increase (P =.01) in contralateral graft flow. This persisted as approximately 15-fold flow differential throughout the 28-day period. The construction yielded a 15-fold differential in wall shear stress between low-flow and high-flow vein grafts (P <.001, two-way repeated measures analysis of variance). Intimal hyperplasia began by day 3, and was 6-fold more in low wall shear grafts by 28 days (230.6 +/- 35.4 microm intimal thickness vs 36.1 +/- 17.6 microm for low shear versus high shear grafts; P =.001). For both cytokines time independently affected mRNA expression (P <.001, global analysis of variance). Exposure of vein grafts to the arterial circulation markedly up regulated IL-1 beta at 1 day, with significantly more induction in the low shear setting (P =.002). IL-1 beta protein localized to the developing neointima at days 1 and 3. Conversely, IL-10 slowly increased until day 14, with significantly more expression in the high shear grafts (P <.001). CONCLUSIONS: Vein graft adaptation induces early pro-inflammatory cytokine IL-1 beta expression and delayed protective IL-10 expression (most notable under high shear conditions), both of which are modulated by wall shear. These differential temporal windows offer strategies for appropriately timed pro-inflammatory or anti-inflammatory therapies to interrupt pathologic vein graft adaptations. CLINICAL RELEVANCE: Neointimal hyperplasia continues to limit the durability of vein bypass grafts. Emerging evidence suggests that inflammatory mechanisms drive the neointimal hyperplasic response. This study demonstrates that specific hemodynamic forces (altered wall shear stress) differentially affect early pro-inflammatory interleukin (IL)-1 beta and delayed anti-inflammatory IL-10 signaling. These distinct temporal windows for IL-1 beta and IL-10 cytokine expression offer strategies for appropriately timed pro-inflammatory and anti-inflammatory therapies to interrupt pathologic vein graft adaptations. PMID- 15297833 TI - Intracellular calcium transients are necessary for platelet-derived growth factor but not extracellular matrix protein-induced vascular smooth muscle cell migration. AB - PURPOSE: Vascular smooth muscle cell (SMC) migration is a critical component of the hyperplastic response that leads to recurrent stenosis after interventions to treat arterial occlusive disease. We investigated the relationship between intracellular calcium ([Ca(2+)](i)) and migration of vascular SMCs in response to platelet-derived growth factor (PDGF) and extracellular matrix (ECM) proteins. METHODS: Human saphenous vein SMCs were used for all experiments. SMC migration in response to agonists was measured with a microchemotaxis assay. A standard fluorimetric assay was used to assess changes in [Ca(2+)](i) in response to the various combinations of growth factors and ECM proteins. RESULTS: The calcium ionophore A23187 produced a rapid rise in [Ca(2+)](i) and a corresponding 60% increase in SMC migration, whereas chelation of [Ca(2+)](i) with BAPTA (1,2-bis [aminophenoxy] ethane-N,N,N',N'-tetraacetic acid) produced a fivefold decrease in PDGF-induced chemotaxis, suggesting that [Ca(2+)](i) is both sufficient and necessary for SMC migration. Stimulation of SMCs with PDGF produced an early peak followed by a late plateau in [Ca(2+)](i). To establish a relationship between temporal fluctuations in [Ca(2+)](i) and SMC migration, SMCs were pretreated with caffeine and ryanadine, which eliminated the initial peak but not the late plateau in [Ca(2+)](i), and had no effect on chemotaxis in response to PDGF. Incubation of SMCs with nickel chloride eliminated the late plateau, but had no effect on the initial peak in [Ca(2+)](i), and reduced PDGF-stimulated migration by fivefold. We then evaluated the role of calcium in SMC migration induced by ECM proteins such as laminin, fibronectin, and collagen types I and IV. All four matrix proteins stimulated SMC migration, but none produced an elevation in [Ca(2+)](i). Moreover, preincubation of SMCs with caffeine and ryanadine or nickel chloride had no effect on ECM protein-induced chemotaxis. CONCLUSION: [Ca(2+)](i) transients are necessary for PDGF but not ECM protein-induced SMC chemotaxis. Moreover, the ability of PDGF to stimulate vascular SMC migration appears dependent on influx of extracellular calcium through membrane channels. CLINICAL RELEVANCE: Recurrent stenosis after angioplasty or surgical bypass remains a significant challenge in treating vascular occlusive disease. In addition to growth factors, extracellular matrix (ECM) proteins may be potent agonists of this process. In this study we show that the influx of extracellular calcium is an important mechanism for platelet-derived growth factor-induced smooth muscle cell migration but not ECM-induced migration. Of note, in clinical trials calcium channel blockers failed to inhibit recurrent stenosis. Our data provide mechanistic insight to help explain this negative outcome in that therapies designed to inhibit restenosis depend on the effects of both growth factors and ECM proteins. PMID- 15297834 TI - Genomic, serologic, and clinical case-control study of Chlamydia pneumoniae and peripheral artery occlusive disease. AB - OBJECTIVES: Chlamydia pneumoniae has been related to atherosclerotic disease in both seroepidemiologic and genomic studies. We performed a case-control study to determine seropositivity and DNA detection in arteries of patients with peripheral artery occlusive disease and of healthy subjects. METHODS: The study included 64 patients with peripheral artery occlusive disease, and 50 control subjects who underwent varicose vein surgery, matched to the patient group for age, sex, and tobacco use. The fibrinogen level in all study subjects was measured as a marker of inflammation. Blood samples were taken from all subjects for determination of immunoglobulin (Ig) G elementary bodies (EB) against C pneumoniae with microimmunofluorescence (MIF) and enzyme-linked immunosorbent assay (ELISA), and of IgA EB with ELISA. The cutoff titers were 1:32 for MIF and 1.1 for ELISA. Biopsy specimens of arterial atheromatous plaque were obtained from patients, and of pudendal artery and saphenous vein from control subjects, and were studied with hemi-nested polymerase chain reaction. RESULTS: There were no differences in fibrinogen level between patients and controls. The prevalence of IgG anti-EB with MIF was 78% in patients and 24% in control subjects (P =.0001; odds ratio [OR], 11.3; 95% confidence interval [CI], 4.7-27.2). Prevalence of IgG anti-EB with ELISA was 75% in patients and 16% in control subjects (P =.0001; OR, 15.7; 95% CI, 6.1-40). There were no differences in IgA anti-EB titers. Bacterial DNA was detected in 67% of atheromatous plaques versus 12% of pudendal arteries (P =.0001) and 4% of saphenous veins. A weak correlation was found between seropositivity and the presence of intravascular DNA. CONCLUSIONS: Our results support the hypothesis that C pneumoniae is related to the pathogenesis of atherosclerotic peripheral artery occlusive disease. CLINICAL RELEVANCE: This study explored the infectious hypothesis in the context of the pathogenesis of atherosclerosis. This hypothesis has been supported by findings that certain infectious agents can cause or accelerate the course of diseases in which the possibility of a microbial cause was not previously proposed, as in the case of peptic ulcer and spongiform encephalopathy. The present study demonstrated the presence of Chlamydia pneumoniae and seropositivity in atheromatous plaques in patients with peripheral artery occlusive disease. These results contribute to a body of research that is opening up the possibility of treating atherosclerotic disease with antibiotic agents, and preventing it with immunization. PMID- 15297835 TI - Endovascular aneurysm repair: Treatment of choice for abdominal aortic aneurysm coincident with horseshoe kidney? Three case reports and review of literature. AB - There is still controversy as to which surgical method is the most suitable for repair of abdominal aortic aneurysm with concomitant horseshoe kidney (AAA-HSK). We report three cases of AAA-HSK treated with endovascular aneurysm repair. In one of these patients we sacrificed the accessory renal artery by applying coils before the operation. Renal infarction, hypertension, or elevated serum creatinine level was not observed in any of our patients. If the blood supply to the kidneys is taken into consideration, endovascular aneurysm repair is our preferred surgical method for repair of AAA-HSK when anatomic conditions are suitable for stent-graft application and kidney function is normal. PMID- 15297836 TI - Iatrogenic subclavian artery pseudoaneurysm causing airway compromise: treatment with percutaneous thrombin injection. AB - Central venous cannulation is an increasingly common diagnostic and therapeutic procedure in modern medical practise. Inadvertent iatrogenic injury to an adjacent major artery during attempted central venous cannulation is an uncommon but potentially lethal complication. If injury leads to pseudoaneurysm formation, complications may arise from mass effect, embolism, or even rupture with continued hemorrhage. Traditional surgical repair of these lesions is often difficult, especially if access within the thorax is required in patients at high risk. We present a case report of successful management of a large subclavian artery pseudoaneurysm with the new approach of percutaneous thrombin injection. PMID- 15297837 TI - Extracranial carotid aneurysm related to pregnancy. AB - Carotid aneurysms are rare, and pregnancy-related carotid aneurysms are rarer still. We report the case of an extracranial carotid aneurysm related to pregnancy. It was noted in the 20th week of gestation, and was uncomplicated. Surgery was successfully performed urgently to preempt embolism or rupture. The patient had a full-term normal delivery. The graft is patent, and the patient has no symptoms at 28-month follow-up. To the best of our knowledge, this case represents the first successful repair of an extracranial carotid aneurysm during pregnancy. PMID- 15297838 TI - Axillorenal arteriovenous graft: a new approach for dialysis access. AB - As the population requiring hemodialysis grows, it becomes increasingly common to encounter patients with limited options for vascular access. Because inability to secure vascular access is a life-threatening problem, it is important to consider all possible options in each patient. We report a new arteriovenous grafting procedure in which the left renal vein is used for outflow in a patient with multiple venous occlusions. Patency of the graft continues 18 months after placement. This graft carries acceptable morbidity, and can be revised. Consideration of this graft is appropriate in selected patients. PMID- 15297839 TI - Floating thoracic aortic thrombus in "protein S" deficient patient. PMID- 15297840 TI - Chronobiology of rupture and dissection of aortic aneurysms. AB - A growing body of evidence suggests that the occurrence of cardiovascular events is not evenly distributed over time, but shows peculiar temporal patterns that vary with time of day, day of the week, and month (season) of the year. These patterns coincide with the temporal variation in the pathophysiologic mechanisms that trigger cardiovascular events and the physiologic changes in body rhythms. These two factors in combination contribute to the periodicity in susceptibility to acute cardiovascular events. The classic assumption of epidemiologic studies that there is a constancy in risk for disease during the various time domains has now been challenged by the emerging new concept of chronorisk. In the last two decades temporal patterns (circadian, weekly, seasonal) have been identified for several acute cardiovascular diseases, such as acute myocardial infarction, sudden death, pulmonary embolism, and stroke, with peak incidence for most in the morning and during winter. One of the most life-threatening cardiovascular emergencies, aortic aneurysm rupture or dissection, also demonstrates periodicity, characterized by a similar temporal distribution, which suggests a common pathophysiologic mechanism or triggers similar to other cardiovascular acute emergencies. We review the data on chronobiology of acute aortic rupture or dissection, and discuss various pathophysiologic mechanisms that account for this variability. It is likely that identification of consistent recurring patterns in the underlying risk mechanisms could provide potential new insights for more precise diagnosis and efficacious therapeutic intervention. PMID- 15297841 TI - Nostrums, quackery, and ethics in vascular surgery: How to remain true to the path of Hippocrates and still feed our families. PMID- 15297842 TI - Medical device adverse event reports and the Food and Drug Administration. PMID- 15297843 TI - The ethics of personal advertising in surgery. PMID- 15297844 TI - Iron-chelation therapy: an update. AB - Chronically transfused patients develop iron overload that leads to organ damage and ultimately to death. The introduction of the iron-chelating agent, desferrioxamine mesylate, dramatically improved the life expectancy of these patients. However, the very demanding nature of this treatment (subcutaneous continuous infusion via a battery-operated portable pump) has been the motivation for attempts to develop alternative forms of treatment that would facilitate the patients' compliance. In this review, we describe the most important advances in iron-chelating therapy. In particular, we analyze a new method of administering desferrioxamine mesylate (twice daily subcutaneous bolus injections) and a novel, orally active iron chelator (ICL670A). We also present a meta-analysis of the largest trials on the oral iron chelator deferiprone and the results of combined therapy (deferiprone and desferrioxamine). PMID- 15297845 TI - Pain syndromes in haematological malignancies: an overview. AB - Several pain syndromes, which may be related to the diagnostic procedures, to the treatments, or to disease itself, may be recorded during the disease course of most haematological malignancies. So far, the painful complication occurring in this setting has been poorly investigated. Pain arising from skeletal and bone marrow (BM) involvement represents the most frequent disease-related painful states observed in this setting, while patients undergoing treatments with curative intent, such as BM transplantation, usually experienced painful stomatitis. Additionally, more than one pathologic process may coexist simultaneously in one patient and the pathophysiology of pain and hypersensitivity may change over time. An accurate diagnostic assessment and the identification of the underlying pathogenetic mechanism may dictate the treatment approach. For most patients in pain, the World Health Organisation's three-step analgesic scale provides adequate relief with oral options. Pain left unrelieved may induce an aberrant peripheral activity and central functional alterations, generating chronic neuropathic pain. In the aim to summarize the current knowledge on this topic, the pertinent literature and the current guidelines for the pain management were reviewed by a group of haematologists, experienced in palliative care and by a skilled algologist, involved as consultant in this clinical setting. PMID- 15297846 TI - Myeloablative radiochemotherapy followed by autologous peripheral blood stem cell transplantation as first-line therapy in peripheral T-cell lymphomas: first results of a prospective multicenter study. AB - Peripheral T-cell lymphomas (PTCL) show a poor outcome following conventional chemotherapy. However, the role of high-dose therapy is still unclear. We initiated a prospective multicenter phase II study to evaluate the feasibility and the efficacy of myeloablative radiochemotherapy followed by autologous peripheral blood stem cell transplantation (APBSCT) as first-line treatment in PTCL. After 4-6 courses CHOP (cyclophosphamide, doxorubicine, vincristine, and prednisone) chemotherapy, an induction therapy with either the DexaBEAM (dexamethasone, carmustine, melphalan, etoposide, and cytarabine) or the ESHAP (etoposide, methylprednisolone, cytarabine, and cisplatin) protocol was administered. If a complete (CR) or a partial remission (PR) was achieved, myeloablative radiochemotherapy (hyperfractionated total body irradiation and high-dose cyclophosphamide) with APBSCT was performed. From June 2000 to November 2002, 30 patients (pts) were enrolled into the study. The two main PTCL subgroups included were PTCL not specified and angioimmunoblastic T-cell lymphoma (each n=12). In all, 21 of the 30 pts (70%) were transplanted. The main toxicities were myelosuppression and infections. The overall response rate after myeloablative therapy was 100% (20 CR, one PR). In total, 16 of 21 transplanted pts (76%) remained in CR at a median follow-up of 15 months (range, 6-32 months) from treatment start. Nine pts received no APBSCT mainly due to progressive disease. In conclusion, this first prospective PTCL-restricted study of frontline myeloablative radiochemotherapy with APBSCT shows feasibility and high efficacy. Our data suggest a substantial impact on outcome, however, longer follow-up is necessary to confirm survival benefit. PMID- 15297847 TI - Thalidomide plus oral melphalan compared with thalidomide alone for advanced multiple myeloma. AB - Thalidomide, the prototype of a new class of agents active against multiple myeloma (MM), exerts synergistic/additive effects when combined with other drugs. The aim of this study was to compare the toxicity and efficacy of thalidomide alone and in combination with oral melphalan. Patients with advanced MM received 100 mg/day oral thalidomide escalated weekly up to 600 mg/day (n=23; T group), alone or with 0.20 oral mg/kg/die melphalan administered monthly for four consecutive days (n=27; TM group). A>/=50% paraprotein reduction was observed in 59% of TM compared with 26% of T patients (P=0.009); three TM patients were found to have an absence of paraprotein by immunofixation. After a median follow-up of 13 months (range 6-32), progression-free survival (PFS) at 2 years was significantly longer in the TM group (61 versus 45%; P=0.0376), whereas overall survival did not differ significantly. Toxicity was not significantly greater with the combination therapy; although DVT was more frequent (11 versus 4%), as was grade 3 leukopenia (30 versus 13%; P=0.073), there were no cases of severe infection. Thalidomide administered with oral melphalan improved response rates and PFS in patients with advanced MM without significantly increasing severe toxicity. PMID- 15297848 TI - Efficacy of low-dose thalidomide and dexamethasone as first salvage regimen in multiple myeloma. AB - PURPOSE: The efficacy of low-dose thalidomide (THAL) plus dexamethasone (DEX) has been evaluated in myeloma. The clinical outcome of patients treated with THAL-DEX was compared with that of a control group treated with conventional chemotherapy (CC). EXPERIMENTAL DESIGN: A total of 120 relapsed/refractory patients to one (52%), or two or more(48%) lines of chemotherapy were treated with THAL 100mg/day (continuous) and DEX 40 mg (days 1-4 of each month). Their clinical outcome was compared to a control group of 120 patients frequency matched for serum beta2 microglobulin levels and Durie and Salmon clinical stage. Clinical characteristics were homogeneous in the two groups. RESULTS: In patients treated after one line of chemotherapy, THAL-DEX significantly improved outcome. Median progression-free survival (PFS) was superior in THAL-DEX group versus CC group (17 months versus 11 months, P = 0.0024). The median survival for THAL-DEX patients has not to been reached, but the probabilities of survival at 3 years were 60% after THAL-DEX and 26% after CC (P = 0.0016). The clinical outcome of patients receiving THAL-DEX or CC after two or more lines of chemotherapy, was similar. In the THAL-DEX group, the medianPFS was 11 months compared to 9 months in the CC group (P = NS). No differences in overall survival (OS) were observed (median OS 19 months for both THAL-DEX and CC). CONCLUSIONS: As first salvage regimen, THAL-DEX was superior to CC, as second or third salvage regimen, it was equivalent to CC. THAL-DEX is not myelotoxic. It postpones the delivery of effective salvage chemotherapy. This might explain the survival benefit. PMID- 15297849 TI - Frequency of hepatitis B virus mutant in asymptomatic hepatitis B virus carriers receiving prophylactic lamivudine during chemotherapy for hematologic malignancies. AB - Hepatitis B virus (HBV) reactivation is a potentially fatal complication of chemotherapy in asymptomatic HBV carriers. Prophylactic lamivudine has proven effective for its prevention, but the potential emergence of lamivudine-resistant HBV YMMD mutants, as shown in patients treated for chronic hepatitis, may limit its use. To evaluate the frequency of HBV YMMD mutant and its clinical significance, we have analysed 32 courses of primary lamivudine prophylaxis given to HBV carriers with haematologic malignancies, from the start until 1-5 months after the end of chemotherapy. Lamivudine was used for a median of 6 months (range 2-24+) and median follow-up was 19.5 months (range 5-40). Four episodes of HBV reactivation with mild hepatitis and no evidence of mutant strain occurred after chemotherapy completion and after lamivudine withdrawal. At follow-up YMMD mutant was detected in one patient with normal transaminase levels, who had been on continuous lamivudine for 20 months. In conclusion, among HBV carriers treated with chemotherapy for haematologic malignancies, the emergence of HBV YMMD mutant occurred in 3.1% of prophylactic lamivudine courses and was of little clinical relevance. PMID- 15297850 TI - Impaired T-cell activation and cytokine productivity after transplantation of positively selected CD34+ allogeneic hematopoietic stem cells. AB - Transplantation of positively selected, CD34(+) peripheral blood stem cells from alternative donors frequently results in delayed immune reconstitution. A shift towards a type 2 cytokine production might be a major contributing factor. We therefore decided to measure IFN-gamma, IL-2, IL-4, and IL-10 after stimulation of peripheral mononuclear cells with PMA/ionomycin and on a single cell level by intracellular cytokine staining during different stages of immune reconstitution. Immediately after transplantation, secretion of all selected cytokines was substantially diminished, and remained subnormal compared to controls until the end of the first year despite normalizing T-cell levels. IL-2 was predominantly produced by CD4(+)CD45RA(+) naive, whereas IFN-gamma originated mainly from CD8(+)CD45RO(+) memory T cells. Secretion of IL-2 was correlated with the numbers of naive CD4(+) T cells, whereas IFN-gamma secretion correlated with total CD3(+) T-cell counts. IL-4 and IL-10 were produced by CD4(+) and CD8(+) memory T cells; secretion of these cytokines was low, however, and did not increase during follow up. Therefore, a shift towards a preferential production of type 2 cytokines could not be observed. Analysis of CD69 upregulation upon stimulation revealed a deficiency in patient T-cell activation, which unexpectedly comprised both naive and memory T-cell subpopulations. Therefore, we suggest that a defect in T-cell activation intrinsic to the host and not graft-versus-host disease, post transplant immunosuppression or a shift towards a type 2 cytokine pattern contributes to the impaired production of cytokines post-transplant. Further studies will focus on the elimination of host factors that may adversely affect T cell function after transplantation. PMID- 15297851 TI - Antiphospholipid antibodies in lymphoma: prevalence and clinical significance. AB - To evaluate whether the presence of antiphospholipid antibodies in lymphoma patients influences their response to treatment, and their rate of thromboembolic complications, we followed up 100 consecutive patients with different lymphomas, who underwent measurement of lupus anticoagulants and anticardiolipin antibodies at diagnosis. In all, 27 patients had lupus anticoagulants and/or anticardiolipin antibodies. This prevalence was significantly higher than in a group of 100 age- and sex-matched normal control subjects (8%; P=0.0008, odds ratio 4.25, 95% confidence interval, 1.82-9.92). At diagnosis, antiphospholipid-positive and negative patients were similar with respect to age, sex, type and staging of lymphomas. During follow-up, the rate of thrombosis was significantly higher in patients with (5.1% patients/year) than without (0.75% patients/year) antiphospholipid antibodies. The two groups were similar with respect to relapse and death rate. In conclusion, antiphospholipid antibodies are associated with lymphomas. Their determination is useful to identify patients at high risk to develop thrombotic complications, but not to predict treatment outcome or disease prognosis. PMID- 15297852 TI - G-CSF induced progenitor mobilization in mice with PIGA- blood cells. AB - OBJECTIVE: In patients with paroxysmal nocturnal hemoglobinuria (PNH) a proportion of blood cells are deficient in glycosyl phosphatidylinositol (GPI) anchored proteins due to a mutation in the PIGA gene. Previous studies showed that in PNH the majority of circulating early progenitor cells were normal but after G-CSF were mainly, of the PNH phenotype. This suggested that GPI-linked proteins contribute to the regulation of progenitor trafficking from bone marrow to peripheral blood. METHODS: To test this hypothesis we studied progenitor cells in bone marrow, spleen, and peripheral blood in response to G-CSF in mice genetically engineered to have a proportion of blood cells deficient in GPI linked proteins (LF mice). RESULTS: In contrast to humans, LF and wild-type mice have comparable numbers of progenitor cells in bone marrow, spleen, and peripheral blood. Similarly, in LF mice the proportion of PIGA- progenitor cells in peripheral blood corresponds the proportion of PIGA- progenitor cells measured in bone marrow and spleen. After G-CSF the number of circulating progenitors significantly increased but the proportion of PIGA- cells remained the same in peripheral blood,bone marrow, and spleen. CONCLUSIONS: Our data indicate that under basal laboratory conditions the lack of GPI-linked protein does not cause a retention of progenitor cells in the bone marrow. This implies that the preferential circulation of normal progenitor cells in patients with PNH requires an additional component that most likely is provided by the altered microenvironment of the underlying bone marrow failure. PMID- 15297853 TI - Autocrine pathway of angiopoietins-Tie2 system in AML cells: association with phosphatidyl-inositol 3 kinase. AB - Hematopoietic cells and endothelial cells are mutually correlated in their development and growth. Various angiogenic factors, such as vascular endothelial growth factor (VEGF) and angiopoietins (Angs), are thought to be associated with leukemia cell growth. In this study, we examined if the Angs-Tie2 autocrine pathway works in primary AML cells or not by using soluble Tie2-Fc, which inhibits Angs from binding to Tie2 receptor. After 48 h of culture with Tie2-Fc, nine AML cells from 19 examined samples were not influenced by Tie2-Fc (group A), while AML cells from remaining 10 patients demonstrated remarkable reduction of cell number by Tie2-Fc treatment (group B). Tie2 receptor, upon binding to Angs, are known to activate phosphatidyl-inositol 3 kinase (PI3 kinase). Then, we examined the effect of LY294002, a potent PI3 kinase inhibitor, on primary AML cells. Cell number reduction effect by the treatment of LY294002 was much more prominent in cells of group B than of group A. In addition, extent of cell number reduction by Tie2-Fc and LY294002 was quite well correlated. These observations demonstrated that cells from a part of AML were dependent on autocrine Angs-Tie2 pathway. This notion was further supported by the study of two AML cell lines, KG 1 and HL-60: the growth of KG-1 was suppressed by Tie2-Fc, and also by anti-Tie2 antibody, which inhibits receptor-ligand interaction, while that of HL-60 was not suppressed by Tie2-Fc or anti-Tie2 antibody. Our results will help to explore the angiogenesis-oriented or endothelial cell-mediated therapy for leukemia. PMID- 15297854 TI - Effective treatment of primary plasma cell leukemia with thalidomide and dexamethasone - a case report. AB - Plasma cell leukemia (PCL) is an aggressive disease with poor prognosis and a median survival of only 2-6 months. Currently, no standard therapy is available, but intensive polychemotherapy appears to be more effective than the conventional melphalan plus prednisone. However, the efficacy of thalidomide in PCL has not yet been widely evaluated. Recently, treatment with thalidomide has been reported to yield promising results in refractory multiple myeloma. Here, we report on a patient with primary PCL in whom first-line treatment with thalidomide/dexamethasone resulted in a rapid response and achievement of a very good partial remission. PMID- 15297856 TI - Congenital methaemoglobinaemia Type I in a Turkish infant due to a novel mutation, Pro144Ser, in NADH-cytochrome b5 reductase. AB - A baby centrally cyanosed from birth was investigated for a congenital cardiac defect. Echocardiography and angiography revealed patent foramen ovale without any other cardiac abnormality. Congenital methaemoglobinaemia was considered as the methaemoglobin level was 27%, suggesting either Hb M or a deficiency of the NADH-cytochrome b5 reductase (cytb5r) enzyme. Measurement of the cytb5r enzyme activity of this patient indicated a reduced level of 7.3 IU/g Hb (normal range 11.5-26.9 IU/g Hb). Sequencing the DIA 1 gene that encodes cytb5r revealed a novel C403T base change, predicting a proline to serine change at codon 144. This amino-acid change is not located in the enzyme's active site and does not cause loss of function. Instead it results in reduced stability of the enzyme and development of the less severe or Type I form of recessive congenital methaemoglobinaemia. The infant was started on daily ascorbic acid treatment. She has very mild cyanosis and normal growth and developmental parameters on follow up at 10 months of age. PMID- 15297855 TI - Chronic hepatitis C associated with Coombs-positive hemolytic anemia. AB - Hepatitis C virus (HCV) is a recognized cause of significant extrahepatic disease. Induction of autoimmune hemolytic anemia (AIHA) has been reported, either during or after interferon (IFN) treatment of HCV infection. We herein report a 56-year-old patient with HCV infection who developed severe Coombs positive AIHA in the absence of treatment with IFN. Prednisone therapy was initiated, but intravenous immunoglobulins were added because of persistent hemolysis. Clinical course was complicated by rapid deterioration and the development of Creutzfeldt-Jakob disease. Having discarded other possible causes of AIHA, we suggest a possible association between AIHA and infection by HCV. PMID- 15297857 TI - Steady-state plasma lactoferrin levels in relation to infections and complications of sickle cell disease. PMID- 15297858 TI - Angiogenic and lymphangiogenic microvessel density in breast carcinoma: correlation with clinicopathologic parameters and VEGF-family gene expression. AB - Angiogenesis and lymphangiogenesis are essential for breast cancer progression and are regulated by vascular endothelial growth factors (VEGF). To determine clinical and molecular correlates of these processes, we measured blood and lymphatic vascular microvessel density in 29 invasive carcinomas (22 ductal, six lobular, one papillary), using the vascular marker CD31 and the novel lymphatic marker D2-40. Microvessel density was assessed microscopically and by image cytometry, and was compared with tumor histology, grade, stage, lymph node metastasis, hormone receptors, HER2/neu status, and expression of VEGF, VEGF-C and VEGF-D by immunohistochemistry or quantitative RT-PCR. Strong correlation was observed between visual and image cytometric microvessel density using D2-40 but not CD31 (P=0.016 and 0.1521, respectively). Image cytometric CD31 microvessel density correlated with tumor size, grade, stage and lymph node metastasis (P=0.0001, 0.0107, 0.0035 and 0.0395, respectively). D2-40 microvessel density correlated with tumor stage (P=0.0123 by image cytometry) and lymph node metastasis (P=0.0558 by microscopy). Immunohistochemical VEGF signal in peritumoral blood vessels correlated with image cytometric CD31 and D2-40 microvessel density (P=0.022 and 0.0012, respectively), consistent with the role of VEGF in blood and lymphatic vascular growth. Intratumoral VEGF-C and VEGF-D expression by quantitative RT-PCR correlated with D2-40 (P=0.0291 by image cytometry) but not with CD31 microvessel density, which could suggest a selective role of VEGF-C and VEGF-D in lymphangiogenesis. CD31 and D2-40 microvessel density correlated significantly with several prognostic factors, including lymph node metastasis. Thus, measurements of angiogenesis and lymphangiogenesis may have utility for breast cancer pathology, particularly for estimation of metastatic risk. PMID- 15297859 TI - Successful combined cataract surgery and drainage of a needling-induced chronic ciliochoroidal detachment. PMID- 15297860 TI - Primary placement of a titanium sleeve in hydroxyapatite orbital implants. AB - BACKGROUND: To study a new surgical option of primary placement of a titanium sleeve into hydroxyapatite implants during enucleation or evisceration. METHODS: A standard enucleation or cornea preserved evisceration was performed, followed by preplacement of a titanium sleeve into the hydroxyapatite implant by a hand drill sleeve driver. Care must be taken to ensure that the titanium sleeve is positioned centrally when the implant is put inside the orbital socket or eviscerated shell. The Tenon capsule and conjunctiva were meticulously closed with minimal tension. Complications such as sleeve exposure, coralline exposure, and infection of the titanium sleeve were closely observed. RESULTS: In all, 30 patients were treated in the above fashion with 10 enucleation and 20 evisceration procedures. The follow-up period ranged from 9 to 24 months. Three of the sleeves were found to have exposed spontaneously at 5 and 7 weeks following original surgery. They had no further complication except one sleeve loosening. The remaining 27 sleeves that did not spontaneously expose pursued secondary exposure of the titanium sleeve and peg insertion by conjunctival cutdown procedure 3 months after original surgery. Two sleeves were found to be oblique positioned after the conjunctival cutdown procedure. Fortunately, all the 30 patients were successfully fit with a peg-coupled prosthesis with good motility. CONCLUSION: Primary placement of a titanium sleeve into hydroxyapatite implants has several advantages, including high patient acceptance, technical simplicity, and office-based conjunctival cutdown pegging procedure. By avoiding the expense of postoperative imaging study and additional prosthetic modification, a more rapid and efficient rehabilitation is possible. PMID- 15297861 TI - Zonular disinsertion five years after implantation of a plate haptic silicone intraocular lens. PMID- 15297862 TI - The Bristol cataract listing survey: profile of listed patients with visual acuity 6/12 or better. AB - INTRODUCTION: The aim of this study was to assess the vision-related quality of life (VR-QOL) in patients with good distance Snellen visual acuity (VA) who are listed for cataract surgery. METHODS: An observational cross-sectional prospective study of patients listed for cataract surgery. VA and VR-QOL data using the VCMI questionnaire were collected on patients attending preoperative assessment during June 2002. RESULTS: A total of 397 cataract patients were listed during this month. Following exclusions there were 378 eligible individuals, 210 (56%) of whom had a VA of 6/12 or better in the eye scheduled for surgery. Of these, 40% patients had only mild VR-QOL impairment. More than half of the patients with good VA (6/12 or better) in the surgery eye and mild VR QOL impairment described their vision as poor in this eye. However, most of these patients were not dissatisfied with their overall level of vision. CONCLUSION: A significant number of patients listed for cataract surgery with VA of 6/12 or better had only mild VR-QOL impairment and were not dissatisfied with their overall level of vision. The decision to list a patient for surgery may have been based on the patient's perception of monocular vision rather than their quality of life. PMID- 15297863 TI - Prospective survey of adverse reactions to topical antiglaucoma medications in a hospital population. AB - AIM: To identify the relative incidence and profile of adverse drug reaction (ADR) to various topical ocular hypotensives in a hospital setting. METHODS: All the patients presenting in outpatients clinic and accident and emergency with an ADR to topical hypotensive agent from August 2000 to January 2001 were included in the study. Details regarding the type date of commencing the treatment, the date of developing ADR, time to resolution of the ADR were noted. RESULTS: Over the period of 6 months, 66 patients presented with 73 ADRs. Brimonidine was the most frequent offending agent. In total, 23 (34.8%) presented with ADR after being commenced on treatment for more than 12 months. In all, 16 (24%) patients had IOP > 21 on presentation, eight (12%) patients underwent filtration surgery following the development of ADR. CONCLUSION: Adverse drug reaction to ocular hypotensive agents is not uncommon and can have a major impact on glaucoma management. Delayed presentation and association with raised intraocular pressure presentation emphasise the need for effective patient education to encourage prompt reporting of ADR. PMID- 15297864 TI - Listing of cataract patients by optometrists. PMID- 15297865 TI - Reference: community refinement of glaucoma referrals. PMID- 15297866 TI - Costs of shared care. PMID- 15297867 TI - Re Diagnostic effectiveness of noncontact slitlamp examination in the identification of retinal tears. PMID- 15297869 TI - Diagnostic effectiveness of noncontact slit-lamp examination in the identifying of retinal tears. PMID- 15297871 TI - Evaluation of risk factors for advanced glaucoma in Ghanaian patients. AB - PURPOSE: This study was to determine factors associated with individuals presenting late with advanced glaucomatous optic nerve damage. METHODS: A case control study recruiting 123 patients with early features of primary open angle glaucoma (control) and 93 patients with advanced glaucoma (cases) was carried out for risk-factor analysis. Exposures of interest included those already established as major risk factors for glaucoma. These were initial intraocular pressure (IOP), age, and family history. In addition, occupation, ethnic origin, history of diagnosis of diabetes mellitus, hypertension, sickle cell disease, and previous eye examination were of interest. RESULTS: Univariate analysis showed that initial IOP>31 mmHg, age of > 60 years, absence of family history of glaucoma, occupational grouping, ethnicity, and male sex were associated with advanced glaucoma at presentation. Adjusted odds ratio or by multiple logistic regression model showed that initial IOP>31 mmHg in a patient was more likely to present with advanced glaucoma (OR 2.66, 95% confidence interval (CI) 1.45, 4.91; P-value 0.0017) than lower pressures. Patients aged 60-69 years (OR 2.53, 95% CI 1.01, 6.31; P-value 0.0473) and 70-90 years (OR 5.16, 95% CI 1.97, 13.51; P-value 0.0008) were more likely to present with advanced glaucoma than younger ones CONCLUSIONS: Subjects with initial IOP>31 mmHg were nearly three times more likely to present with advanced glaucoma than those with IOP<32 mmHg. Subjects over the age of 60 years were more than two times likely to present with advanced glaucoma than younger subjects. PMID- 15297872 TI - A stop-EGFP transgenic mouse to detect clonal cell lineages generated by mutation. AB - The investigation of cell lineages and clonal organization in tissues is facilitated by techniques that allow labelling of clonal cell lineages. Here, we describe a novel transgenic mouse that allows clonal cell lineages to be traced in virtually any tissue. A green fluorescent cell lineage is generated by a random mutation at an enhanced green fluorescent protein gene that carries a premature stop codon, ensuring clonality. The transgenic system allows efficient detection of mutations and stem-cell fate mapping in the epidermis using live mice, as well as in the kidney and liver post-mortem. Cell lineages that descended from single epidermal stem cells were found to be capable of generating three adjacent corneocytes using the system, providing evidence for horizontal migration of epidermal cells between epidermal proliferative units (EPUs), in contrast to the classical EPU model. The transgenic mouse system is expected to provide a novel tool for stem-cell lineage studies. PMID- 15297873 TI - Paraxial protocadherin coordinates cell polarity during convergent extension via Rho A and JNK. AB - Convergent extension movements occur ubiquitously in animal development. This special type of cell movement is controlled by the Wnt/planar cell polarity (PCP) pathway. Here we show that Xenopus paraxial protocadherin (XPAPC) functionally interacts with the Wnt/PCP pathway in the control of convergence and extension (CE) movements in Xenopus laevis. XPAPC functions as a signalling molecule that coordinates cell polarity of the involuting mesoderm in mediolateral orientation and thus selectively promotes convergence in CE movements. XPAPC signals through the small GTPases Rho A and Rac 1 and c-jun N-terminal kinase (JNK). Loss of XPAPC function blocks Rho A-mediated JNK activation. Despite common downstream components, XPAPC and Wnt/PCP signalling are not redundant, and the activity of both, XPAPC and PCP signalling, is required to coordinate CE movements. PMID- 15297874 TI - A dedicated translation factor controls the synthesis of the global regulator Fis. AB - BipA is a highly conserved protein with global regulatory properties in Escherichia coli. We show here that it functions as a translation factor that is required specifically for the expression of the transcriptional modulator Fis. BipA binds to ribosomes at a site that coincides with that of elongation factor G and has a GTPase activity that is sensitive to high GDP:GTP ratios and stimulated by 70S ribosomes programmed with mRNA and aminoacylated tRNAs. The growth rate dependent induction of BipA allows the efficient expression of Fis, thereby modulating a range of downstream processes, including DNA metabolism and type III secretion. We propose a model in which BipA destabilizes unusually strong interactions between the 5' untranslated region of fis mRNA and the ribosome. Since BipA spans phylogenetic domains, transcript-selective translational control for the 'fast-track' expression of specific mRNAs may have wider significance. PMID- 15297875 TI - Essential roles of KIF4 and its binding partner PRC1 in organized central spindle midzone formation. AB - A number of proteins accumulate in the anaphase spindle midzone, but the interaction and precise role of these proteins in midzone organization remain obscure. Here, we found that the microtubule-bundling protein PRC1 bound separately to the three motor proteins, KIF4, MKLP1 and CENP-E, but not to the chromosomal passenger proteins. In KIF4-deficient cells, the central spindle was disorganized, and all midzone-associated proteins including PRC1 failed to concentrate at the midline, instead being dispersed along the loosened microtubule bundles of the central spindle. This suggests that KIF4 is essential for the organization of central spindles and for midzone formation. In PRC1 deficient cells, no midzone was formed, KIF4 and CENP-E did not localize to the disconnected half-spindle, and MKLP1 and chromosomal passenger proteins localized to discrete subdomains near microtubule plus ends in the half-spindle. Thus, PRC1 is required for interaction of the two half-spindles and for localization of KIF4 and CENP-E. These results suggest that KIF4 and its binding partner PRC1 play essential roles in the organization of central spindles and midzone formation. PMID- 15297876 TI - Suppression of retroviral infection by the RAD52 DNA repair protein. AB - Reverse transcription of retroviral RNA into linear double-stranded DNA and its integration into the host cell genome are essential steps in the retroviral life cycle. The nonhomologous end-joining (NHEJ) DNA repair pathway has been implicated in protecting cells from retrovirus-induced apoptosis caused by strand breaks in host cell DNA or unintegrated linear viral DNA. In eukaryotes, both the NHEJ and homologous recombination (HR) pathways play important roles in repairing DNA double-strand breaks. Here we show that the HR repair protein RAD52 modulates the outcome of recombinant HIV-l vector infection by markedly reducing the efficiency of productive integration events. Increased retroviral integration is the first major phenotype described for a RAD52 deficiency in mammalian cells. Mutations in other HR proteins (XRCC2, XRCC3 and BRCA2) do not markedly affect retroviral transduction rates, suggesting that the HR repair pathway per se does not influence retroviral infection. Instead, the mechanism of attenuation of retroviral infection by RAD52 appears to be based upon competition between the RAD52 protein and active integration complexes for the retroviral cDNA genome. PMID- 15297877 TI - Crystal structure of the nuclear effector of Notch signaling, CSL, bound to DNA. AB - Notch signaling is a conserved pathway of communication between neighboring cells that results in cell fate specification, and CSL is the universal transcriptional effector of Notch signaling. The Notch intracellular domain translocates to the nucleus after proteolytic release upon Notch extracellular engagement, and there it displaces corepressors from DNA-bound CSL and recruits activators of Notch target genes. Here we report the 2.85 A crystal structure of CSL with a target DNA. CSL comprises three structurally integrated domains: its amino (NTD)- and carboxy (CTD)-terminal domains are strikingly similar to those of Rel transcription factors, but a surprising beta-trefoil domain (BTD) is inserted between them. CSL-bound DNA is recognized specifically by conserved residues from NTD and BTD. A hydrophobic pocket on BTD is identified as the likely site of Notch interaction with CSL, which has functional implications for the mechanism of Notch signaling. PMID- 15297878 TI - Vertical collapse of a cytolysin prepore moves its transmembrane beta-hairpins to the membrane. AB - Perfringolysin O (PFO) is a prototype of the large family of pore-forming cholesterol-dependent cytolysins (CDCs). A central enigma of the cytolytic mechanism of the CDCs is that their membrane-spanning beta-hairpins (the transmembrane amphipathic beta-hairpins (TMHs)) appear to be approximately 40 A too far above the membrane surface to cross the bilayer and form the pore. We now present evidence, using atomic force microscopy (AFM), of a significant difference in the height by which the prepore and pore protrude from the membrane surface: 113+/-5 A for the prepore but only 73+/-5 A for the pore. Time-lapse AFM micrographs show this change in height in real time. Moreover, the monomers in both complexes exhibit nearly identical surface features and these results in combination with those of spectrofluorimetric analyses indicate that the monomers remain in a perpendicular orientation to the bilayer plane during this transition. Therefore, the PFO undergoes a vertical collapse that brings its TMHs to the membrane surface so that they can extend across the bilayer to form the beta-barrel pore. PMID- 15297879 TI - Activation of cardiac Cdk9 represses PGC-1 and confers a predisposition to heart failure. AB - Hypertrophy allows the heart to adapt to workload but culminates in later pump failure; how it is achieved remains uncertain. Previously, we showed that hypertrophy is accompanied by activation of cyclin T/Cdk9, which phosphorylates the C-terminal domain of the large subunit of RNA polymerase II, stimulating transcription elongation and pre-mRNA processing; Cdk9 activity was required for hypertrophy in culture, whereas heart-specific activation of Cdk9 by cyclin T1 provoked hypertrophy in mice. Here, we report that alphaMHC-cyclin T1 mice appear normal at baseline yet suffer fulminant apoptotic cardiomyopathy when challenged by mechanical stress or signaling by the G-protein Gq. At pathophysiological levels, Cdk9 activity suppresses many genes for mitochondrial proteins including master regulators of mitochondrial function (peroxisome proliferator-activated receptor gamma coactivator 1 (PGC-1), nuclear respiratory factor-1). In culture, cyclin T1/Cdk9 suppresses PGC-1, decreases mitochondrial membrane potential, and sensitizes cardiomyocytes to apoptosis, effects rescued by exogenous PGC-1. Cyclin T1/Cdk9 inhibits PGC-1 promoter activity and preinitiation complex assembly. Thus, chronic activation of Cdk9 causes not only cardiomyocyte enlargement but also defective mitochondrial function, via diminished PGC-1 transcription, and a resulting susceptibility to apoptotic cardiomyopathy. PMID- 15297880 TI - Regulation of human SRY subcellular distribution by its acetylation/deacetylation. AB - SRY, a Y chromosome-encoded DNA-binding protein, is required for testis organogenesis in mammals. Expression of the SRY gene in the genital ridge is followed by diverse early cell events leading to Sertoli cell determination/differentiation and subsequent sex cord formation. Little is known about SRY regulation and its mode of action during testis development, and direct gene targets for SRY are still lacking. In this study, we demonstrate that interaction of the human SRY with histone acetyltransferase p300 induces the acetylation of SRY both in vitro and in vivo at a single conserved lysine residue. We show that acetylation participates in the nuclear localisation of SRY by increasing SRY interaction with importin beta, while specific deacetylation by HDAC3 induces a cytoplasmic delocalisation of SRY. Finally, by analysing p300 and HDAC3 expression profiles during both human or mouse gonadal development, we suggest that acetylation and deacetylation of SRY may be important mechanisms for regulating SRY activity during mammalian sex determination. PMID- 15297881 TI - Mutation of the mouse Rad17 gene leads to embryonic lethality and reveals a role in DNA damage-dependent recombination. AB - Genetic defects in DNA repair mechanisms and cell cycle checkpoint (CCC) genes result in increased genomic instability and cancer predisposition. Discovery of mammalian homologs of yeast CCC genes suggests conservation of checkpoint mechanisms between yeast and mammals. However, the role of many CCC genes in higher eukaryotes remains elusive. Here, we report that targeted deletion of an N terminal part of mRad17, the mouse homolog of the Schizosaccharomyces pombe Rad17 checkpoint clamp-loader component, resulted in embryonic lethality during early/mid-gestation. In contrast to mouse embryos, embryonic stem (ES) cells, isolated from mRad17(5'Delta/5'Delta) embryos, produced truncated mRad17 and were viable. These cells displayed hypersensitivity to various DNA-damaging agents. Surprisingly, mRad17(5'Delta/5'Delta) ES cells were able to arrest cell cycle progression upon induction of DNA damage. However, they displayed impaired homologous recombination as evidenced by a strongly reduced gene targeting efficiency. In addition to a possible role in DNA damage-induced CCC, based on sequence homology, our results indicate that mRad17 has a function in DNA damage dependent recombination that may be responsible for the sensitivity to DNA damaging agents. PMID- 15297882 TI - Structural basis for the dual coding potential of 8-oxoguanosine by a high fidelity DNA polymerase. AB - Accurate DNA replication involves polymerases with high nucleotide selectivity and proofreading activity. We show here why both fidelity mechanisms fail when normally accurate T7 DNA polymerase bypasses the common oxidative lesion 8-oxo-7, 8-dihydro-2'-deoxyguanosine (8oG). The crystal structure of the polymerase with 8oG templating dC insertion shows that the O8 oxygen is tolerated by strong kinking of the DNA template. A model of a corresponding structure with dATP predicts steric and electrostatic clashes that would reduce but not eliminate insertion of dA. The structure of a postinsertional complex shows 8oG(syn).dA (anti) in a Hoogsteen-like base pair at the 3' terminus, and polymerase interactions with the minor groove surface of the mismatch that mimic those with undamaged, matched base pairs. This explains why translesion synthesis is permitted without proofreading of an 8oG.dA mismatch, thus providing insight into the high mutagenic potential of 8oG. PMID- 15297885 TI - The biochemical mechanism of caspase-2 activation. AB - A unified model for initiator caspase activation has previously been proposed based on the biochemical analysis of caspase-8 and -9. Caspase-2 is structurally related to caspase-9, but its mechanism of activation is not known. Using an uncleavable mutant of caspase-2, we show that dimerization (and not processing) is the key event that drives initial procaspase-2 activation. Following dimerization, caspase-2 undergoes autocatalytic cleavage that promotes its stable dimerization and further enhances the catalytic activity of caspase-2. Although the caspase-2 zymogen does not require cleavage for the initial acquisition of activity, intersubunit cleavage is required to generate levels of activity required to induce cell death by overexpression. We also provide evidence that the reported disulfide bond linkage between two caspase-2 monomers is dispensable for caspase-2 dimerization. As caspase-2 does not require cleavage for its initial activation, our findings confirm caspase-2 to be a bona fide initiator caspase. PMID- 15297883 TI - Galectin-1 induces nuclear translocation of endonuclease G in caspase- and cytochrome c-independent T cell death. AB - Galectin-1, a mammalian lectin expressed in many tissues, induces death of diverse cell types, including lymphocytes and tumor cells. The galectin-1 T cell death pathway is novel and distinct from other death pathways, including those initiated by Fas and corticosteroids. We have found that galectin-1 binding to human T cell lines triggered rapid translocation of endonuclease G from mitochondria to nuclei. However, endonuclease G nuclear translocation occurred without cytochrome c release from mitochondria, without nuclear translocation of apoptosis-inducing factor, and prior to loss of mitochondrial membrane potential. Galectin-1 treatment did not result in caspase activation, nor was death blocked by caspase inhibitors. However, galectin-1 cell death was inhibited by intracellular expression of galectin-3, and galectin-3 expression inhibited the eventual loss of mitochondrial membrane potential. Galectin-1-induced cell death proceeds via a caspase-independent pathway that involves a unique pattern of mitochondrial events, and different galectin family members can coordinately regulate susceptibility to cell death. PMID- 15297884 TI - Serum bioactive lysophospholipids prevent TRAIL-induced apoptosis via PI3K/Akt dependent cFLIP expression and Bad phosphorylation. AB - Serum contains a variety of biomolecules, which play an important role in cell proliferation and survival. We sought to identify the serum factor responsible for mitigating tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induced apoptosis and to investigate its molecular mechanism. TRAIL induced effective apoptosis without serum, whereas bovine serum decreased apoptosis by suppressing cytochrome c release and caspase activation. Indeed, albumin-bound lysophosphatidic acid (LPA) and sphingosine-1-phosphate (S1P) inhibited TRAIL induced apoptosis by suppressing caspase activation and cytochrome c release. LPA increased phosphatidylinositol 3-kinase (PI3K)-dependent Akt activation, cellular FLICE-inhibitory protein (cFLIP) expression, and Bad phosphorylation, resulting in inhibition of caspase-8 activation and Bad translocation to mitochondria. The antiapoptotic effect of LPA was abrogated by PI3K inhibitor, transfection with dominant-negative Akt, and specific downregulation of cFLIP expression using siRNA and further increased by siRNA-mediated suppression of Bad expression. Moreover, sera from ovarian cancer patients showed more protective effect against TRAIL-induced apoptosis than those from healthy donors, and this protection was suppressed by PI3K inhibitor. Our results indicate that albumin-bound LPA and S1P prevent TRAIL-induced apoptosis by upregulation of cFLIP expression and in part by Bad phosphorylation, through the activation of PI3K/Akt pathway. PMID- 15297886 TI - Cathepsin B-independent abrogation of cell death by CA-074-OMe upstream of lysosomal breakdown. PMID- 15297887 TI - Cystic fibrosis as a cause of infertility. AB - Cystic fibrosis (CF) is one of the autosomal recessive diseases, caused by mutations in a gene known as cystic fibrosis transmembrane regulator (CFTR). The majority of adult males with CF (99%) is characterized by congenital bilateral absence of vas deferens (CBAVD). CBAVD is encountered in 1-2% of infertile males without CF. Females with CF are found to be less fertile than normal healthy women. In females with CF, delayed puberty and amenorrhoea are common due to malnutrition. CFTR mutations are also associated with congenital absence of the uterus and vagina (CAUV). The National Institutes of Health recommend genetic counseling for any couple seeking assisted reproductive techniques with a CF male or obstructive azoospermia which is positive for a CF mutation. PMID- 15297888 TI - Trophoblastic cDNA cloning of porcine Pregnancy-Associated Glycoprotein genes (pPAG) and in silico analysis of coded polypeptide precursors. AB - Multiple Pregnancy-Associated Glycoprotein (PAG) genes, coding various chorionic products that belong to the aspartic proteinase family (AP), have been discovered in various eutherians (Placentalia). This paper presents cloning of five novel porcine PAG (pPAG) cDNAs: pPAG3, pPAG4, pPAG5, pPAG6, pPAG8 and pPAG10. All cDNAs have been identified by trophoblastic library screening on 13-17 days post coitum (dpc). These cloned and sequenced pPAG cDNAs have been deposited in GenBank database of National Institute of Heath/ National Center of Biotechnology Information (NIH/NCBI) with various accession numbers: AF315377, AF272734, AY188554, AF272735, AY373029 and one during submission. The open reading frames (ORF) of cloned cDNA sequences allowed for identification and in silico analysis of polypeptide pPAG precursors, including their identity content and various amino acid substitutions, which caused hypothetical structure changes of the pPAG precursors. In conclusion, our results concerning transcriptomal cloning and in silico analyses of the novel identified trophoblastic cDNA of the pPAG genes provided contributive data regarding porcine genome identification and physiology of the gestation in this species. PMID- 15297889 TI - The involvement of luteinizing hormone (LH) and Pregnancy-Associated Glycoprotein family (PAG) in pregnancy maintenance in the pig. AB - The paper presents the effect of in vivo immuno-neutralization of porcine luteinizing hormone (pLH) by species-homologous porcine antiserum (anti-pLH) administrations on pregnancy maintenance and immunodetection of the PAG proteins in precipitated plasma proteins of pregnant gilts. Pregnant gilts were passively immunized with 100 ml of porcine anti-pLH (titer 1:10 000) by multiple intravenous infusions performed from 37(th) to 42(nd) day post coitum (dpc; 12-h intervals). Blood samples of pregnant gilts were taken 12 times daily from 35 until 50 dpc. Concentrations of progesterone (P(4)) and pLH were determined by radioimmunoassays in systemic blood plasma of treated gilts and control pregnant gilts. The immuno-neutralization of peripheral pLH with the use of homologous anti-pLH serum resulted in a significant reduction (p<0.001) of plasma P(4) concentrations in two out of six treated gilts only, but abortion did not occur. In the remaining four passively immunized pregnant gilts, plasma P(4) concentration was increased (p<0.001) and the abortion occurred (47 dpc) only in one of the gilts. In addition, various anti-pPAG sera were purified by sequential adsorptions with endometrial proteins of cyclic gilts. Western blotting demonstrated the expression of the PAG proteins in precipitated plasma proteins of pregnant gilts. In conclusion, the passive immuno-neutralization of porcine LH by species-homologous antiserum (anti-pLH) did not affect the pregnancy maintenance. Thus, the maintenance of mid-pregnancy in gilts may depend also on other than LH luteotrophic factors. In addition, Western analysis of precipitated plasma proteins of pregnant pigs suggests a role of the PAG family during pregnancy in the pig. PMID- 15297890 TI - The effect of cyproterone acetate on the antler cycle in red deer (Cervus elaphus L.). AB - The aim of the study was to test the effect of antiandrogen, cyproterone acetate (CA) on the antler cycle in the red deer (Cervus elaphus). CA was administered to three adult red deer stags (Edward, Fuks and Gacek) in weekly intervals. Edward and Fuks were given 600 mg + 600 mg of CA, whereas Gacek was given 600 mg + 300 mg. CA was injected during the hard antler phase: in mid-October (Edward), at the end of November (Fuks) and at the end of January (Gacek). CA caused the antler casting 17 to 22 days after the first injection. In all stags, the casting of antlers was followed by a period of intensive growth of new antlers. Edward was given CA at the end of October. This treatment was responsible for occurrence of the two antler cycles in the year of the experiment. When CA was administered during the middle of the hard antler phase an additional short antler cycle occurs followed by new antler growth. CA treatment in the later part of hard antler phase may cause a prolonged antler cycle. PMID- 15297891 TI - Oxytocin stimulates prostaglandin F2alpha secretion and prostaglandin F synthase protein expression in porcine myometrial tissue. AB - The possibility of PGF(2)alpha production and presence of prostaglandin F synthase (PGFS; PGD(2) 11-ketoreductase) was studied in control and oxytocin (OT) stimulated myometrial slices isolated from cyclic (Days 14-16) and early pregnant (Days 14-16) sows. Oxytocin (10(-7) M) stimulated (p<0.01) PGF(2)alpha production in both cycling and early pregnant myometrial slices. Prostaglandin F(2)alpha release was higher (p<0.01) in control as well as OT-treated myometrium of early pregnant sows in comparison to cycling myometrium. Prostaglandin F synthase expression at protein level was evident in myometrial slices of cyclic as well as early pregnant sows. The signals of PGFS was stronger (p<0.05) in cycling myometrium exposed to OT compared to that of control. There were no significant differences (p>0.05) in PGFS protein expression between control and OT-stimulated myometrial tissue of early-pregnant sows. The results of this study indicate the local PGF(2)alpha synthesis and the presence of PGFS in porcine cycling and early pregnant myometrial tissue. In addition, OT increased PGD(2) 11-ketoreductase protein expression in myometrium harvested during the porcine estrous cycle. However, the OT-stimulated PGF(2)alpha myometrial secretion was observed in both, cycling and pregnant gilts. PMID- 15297892 TI - Genistein affects testosterone secretion by Leydig cells in roosters (Gallus gallus domesticus). AB - Genistein is one of non-steroidal phytoestrogens present in soya and soybean products as well as in other legumes. Phytoestrogens possess estrogen-like biological activity and may influence human and animal reproduction. The aim of this study was to examine the effect of genistein on testosterone (T) secretion by isolated Leydig cells in roosters. Genistein (5-50 microM) inhibited (p<0.05) in vitro basal and LH-stimulated T secretion by Leydig cells in a dose dependent manner. No significant effect of lavendustin C (inhibitor of PTK, a non phytoestrogen) on the T production was observed. In conclusion, genistein, present in commercial poultry feeds, may influence testicular steroidogenesis but its effect on reproductive performance of roosters requires further examinations. PMID- 15297893 TI - Luteinizing hormone and prolactin are not retrograde transferred in perihypophyseal vascular complex in ewes. AB - The objective of the study was to determine whether luteinizing hormone (LH) and prolactin (PRL) can access the brain by way of transfer from the venous blood of the cavernous sinus to the arterial blood supplying the brain and hypophysis. Studies were performed on heads of 22 mature sheep isolated during different phases of the estrous cycle and perfused with autologous blood. We were not able to demonstrate any transfer of LH and PRL in the investigated periods. This suggests that molecular weight of hormone may be a main factor determining the permeation and transfer of hormones in the perihypophyseal vascular complex. PMID- 15297894 TI - The influence of virtual reality on playfulness in children with cerebral palsy: a pilot study. AB - The purpose of this paper was to examine the effects of virtual play intervention on the level of playfulness of children with cerebral palsy. Thirteen children aged 8-13 years comprised the study group. Children attended eight one-hour virtual reality play sessions in which they were immersed and interacted with virtual reality. The Test of Playfulness (TOP) was used as the measure to assess playfulness. Participants were videotaped while they played during 12 different environments over the course of their intervention time. Three randomly selected virtual reality play sessions were chosen to score three different virtual reality environments within each session yielding a total of nine trials (environments) for each participant. The types of virtual environments varied across participants. Overall, the different virtual reality play environments produced varying levels of playfulness according to the TOP's four different subscale scores. Motivation ranged from 1.50 to 2.25, internal control ranged from 1.00 to 1.88, suspension of reality ranged from 0 to 0.26, and framing ranged from 1.33 to 1.78. The three environments producing the highest playfulness ratings were called Paint, Trip and Island Sounds. These environments allowed creativity, persistence with the task, pleasure, and a certain degree of control. Two environments did not appear to foster playfulness. A possible reason was that these environments were too unpredictable and frustrating for participants. These results will be useful for creating new virtual reality software applications that will encourage playfulness in children with disabilities. PMID- 15297895 TI - The effects of guided imagery relaxation in people with COPD. AB - The aim of this study was to explore the effects of guided imagery relaxation in people with chronic obstructive pulmonary disease (COPD) using a randomized controlled design. Half of 26 participants were allocated to the treatment group in which six practice sessions on guided imagery were conducted, while the control group was instructed to take rest quietly during the six sessions. At the seventh session, physiological changes: partial percentage of oxygen saturation; heart rate; upper thoracic surface electromyography; skin conductance; and peripheral skin temperature were recorded during a 30-minute session with a sampling frequency of one minute. Repeated measures analysis of variance was used to explore the changes of the parameters between the groups. Mann-Whitney U test was used to compare the change of perceived dyspnoea between the groups. Results showed there was a statistically significant (p < 0.05) increase in partial percentage of oxygen saturation in the treatment group, but no significant effects on the other physiological parameters. Further study exploring the psychological effects of guided imagery is suggested. PMID- 15297896 TI - Comparison of clinical reasoning skills in occupational therapy students in the USA and Scotland. AB - Problem-based learning (PBL) in occupational therapy (OT) education has become increasingly important. However, the efficacy in identifying clinical reasoning skills has not been well researched. The purpose of this study was to evaluate the utilization of clinical reasoning by students in the USA and Scotland. Thirty OT students were videotaped during a PBL case. Responses were transcribed and compiled into 122 statements. The two primary investigators independently coded each of the responses into one of five categories: Procedural, Interactive, Conditional reasoning, N/A, or Other. Intraclass correlation coefficients reflected strong reliability between the two raters (ICC = 0.914). Results of this study indicated that the predominant form of clinical reasoning was procedural in nature (61%) followed by conditional reasoning (27%) and interactive reasoning (12%). Students in Scotland tended to use interactive reasoning more than the students in the USA. Further research is needed to explore educational models to foster clinical reasoning skills in occupational therapy students. PMID- 15297897 TI - Time use of people with schizophrenia living in the community. AB - The objective of this study was to describe the time use of people with schizophrenia living in the community in Japan. Time use of participants with schizophrenia was compared with people without schizophrenia. The time use of participants without a work-related routine was also compared with those with a work-related routine. As has been reported in other studies, persons with schizophrenia spend a significantly greater amount of time sleeping and resting than people without schizophrenia. However, participants with schizophrenia chose to participate not only in passive occupational categories but also in active occupational categories by selecting work, co-operative workshops, play, socialization and homemaking. Results also showed that having a work-related routine influenced time use. People with a work-related routine spent much time engaged in the work-related routines and in sleeping, and people without the routine spent much time sleeping, listening to music, watching TV, and reading magazines. The present study contributes to developing knowledge that supports occupational therapy practice focused on occupational choice including routine occupations to build a healthy life for people with schizophrenia living in the community. To explore further the relationship between time use and health, further research would be needed to study individual experiences in occupations. PMID- 15297898 TI - Centring in regression analyses: a strategy to prevent errors in statistical inference. AB - Regression analyses are perhaps the most widely used statistical tools in medical research. Centring in regression analyses seldom appears to be covered in training and is not commonly reported in research papers. Centring is the process of selecting a reference value for each predictor and coding the data based on that reference value so that each regression coefficient that is estimated and tested is relevant to the research question. Using non-centred data in regression analysis, which refers to the common practice of entering predictors in their original score format, often leads to inconsistent and misleading results. There is very little cost to unnecessary centring, but the costs of not centring when it is necessary can be major. Thus, it would be better always to centre in regression analyses. We propose a simple default centring strategy: (1) code all binary independent variables +1/2; (2) code all ordinal independent variables as deviations from their median; (3) code all 'dummy variables' for categorical independent variables having m possible responses as 1 - 1/m and -1/m instead of 1 and 0; (4) compute interaction terms from centred predictors. Using this default strategy when there is no compelling evidence to centre protects against most errors in statistical inference and its routine use sensitizes users to centring issues. PMID- 15297899 TI - Multilevel investigation of variation in HoNOS ratings by mental health professionals: a naturalistic study of consecutive referrals. AB - Episodes of mental healthcare in specialist psychiatric services often begin with the assessment of clinical and psychosocial needs of patients by healthcare professionals. Particularly for patients with complex needs or severe problems, ratings of clinical and social functioning at the start of each episode of care may serve as a baseline against which subsequent measures can be compared. Currently, little is known about service variations in such assessments on referrals from primary care. We set out to quantify variability in initial assessments performed by healthcare professionals in three CMHTs in Bristol (UK) using the Health of the Nation Outcome Scales (HoNOS). We tested the hypothesis that variations in HoNOS total and sub-scale scores are related to referral source (general practices), healthcare assessor (in CMHTs) and the assessor's professional group. Statistical analysis was performed using multilevel variance components models with cross-classified random effects. We found that variation due to assessor substantially exceeded that due to referral source (general practices). Furthermore, patient variance differed by assessor profession for the HoNOS--Impairment scores. Assessor variance differed by assessor profession for the HoNOS--Social scores. As HoNOS total and subscale scores show much larger variation by assessor than by referral source, investigations of HoNOS scores must take assessors into account. Services should implement and evaluate interdisciplinary training to improve consistency in use of rating thresholds; such initiatives could be evaluated using these extensions of multilevel models. Future research should aim to integrate routine diagnostic data with continuous outcomes to address selection effects (of patients to assessors) better. PMID- 15297900 TI - Self-reported use of mental health services versus administrative records: care to recall? AB - Estimates of the level of unmet need for mental health treatment often rely on self-reported use of mental health services. However, depressed persons may over report their use in relation to administrative records if they are highly distressed. This study seeks to replicate and explicate the finding that persons at a high level of distress report more mental health service use than recorded in their healthcare records. The study sample, N = 36,892, 12 years and older, was drawn from the 1996/97 Ontario portion of the Canadian National Population Health Survey. Respondents were individually linked to their administrative mental healthcare records 12 months backward in time. Of these, 96.5% agreed to the link and 23,063 (62.5%) were linked. Almost two-thirds of those who were depressed in the past year were currently at a high level of distress. Differential reporting of use for highly distressed persons in excess of 100% remained in the use of different types of physician providers after adjustments for other potential determinants of use. Telescoping was also not an explanation. The patterns of differential reporting between groups expected to diverge and converge in their recall ability were consistent with a recall bias. As this study was not able to rule out a recall bias, it further accentuates concerns about the impact of bias in the measurement of mental health-service use and inferences made concerning the determinants of use. PMID- 15297901 TI - Continuation and long-term maintenance treatment with Hypericum extract WS 5570 after successful acute treatment of mild to moderate depression--rationale and study design. AB - Unipolar major depression is often a chronic disease that may require lifelong prophylaxis. Recovery from an acute episode is followed by 4-6 months of relapse prevention. After that, long-term maintenance treatment is administered to avoid recurrence. We present the rationale and design of an ongoing double-blind, randomized, placebo-controlled trial investigating the efficacy of Hypericum extract WS 5570 in relapse prevention in recurrent unipolar depression. An estimated sample of 425 adults with recurrent, mild to moderate major depression (ICD-10 and DSM-IV criteria), > or = 3 previous episodes (last 5 years) and a total score > or = 20 points on the 17-item Hamilton Rating Scale for Depression (HAMD) will be included. After a one-week wash out patients receive 3 x 300 mg/day WS 5570 single-blind for 6 weeks. Responders are randomized to 26 weeks of double-blind continuation treatment with 3 x 300 mg/day WS 5570 or placebo. Patients completing continuation treatment without relapse enter 52 weeks of doubleblind maintenance treatment, where those treated with WS 5570 are re randomized to 3 x 300 mg/day WS 5570 or placebo. The primary outcome measure is the time to relapse during continuation treatment (HAMD > or = 16, clinical diagnosis of depression, or premature treatment termination for inefficacy). Hypericum extract, with its favourable tolerability profile, could be an interesting option for long-term prophylaxis. The trial was designed according to current consensus and guidance. Notably, it includes long-term prophylactic treatment with the same drug and the same therapeutic dose applied during acute treatment, uses well-defined outcome measures and provides a clear distinction between relapse and recurrence. PMID- 15297902 TI - The Homeless Supplement to the Diagnostic Interview Schedule: test-retest analyses. AB - This study sought to extend previous work on reliability of self-reported residential history in a homeless population with high rates of drug abuse. The latest version of the Homeless Supplement to the Diagnostic Interview Schedule (DIS/HS) was used to achieve reliability on homelessness experience, use of shelters, transience, and recent residential patterns. Homeless study volunteers were recruited for a test-retest study from a drop-in day centre for mentally ill homeless people (N = 25) and a substance abuse day programme (N = 30). They were administered the instrument approximately one to two days apart. Kappa and intraclass correlation analyses were performed to assess reliability. Overall, the reliabilities of most variables were acceptable, ranging from fair to excellent. Six items were reconstructed to achieve reliability and two were dropped. Substance dependence and adult antisocial behaviour patterns did not affect reliability on most items. This study has developed a reliable self report instrument for measuring residential history that can be used with homeless and drug abusing populations. Replication is needed in larger, more representative samples and comparison of reliability with other psychiatric and cognitive characteristics. PMID- 15297904 TI - The National Comorbidity Survey Replication (NCS-R): background and aims. AB - The National Comorbidity Survey Replication (NCS-R) is a new nationally representative community household survey of the prevalence and correlates of mental disorders in the US. The NCS-R was carried out a decade after the original NCS. The NCS-R repeats many of the questions from the NCS and also expands the NCS questioning to include assessments based on the more recent Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition (DSM-IV) diagnostics system (American Psychiatric Association, 1994). The NCS-R was designed to (1) investigate time trends and their correlates over the decade of the 1990s and (2) expand the assessment of the prevalence and correlates of mental disorders beyond the assessment in the baseline NCS in order to address a number of important substantive and methodological issues that were raised by the NCS. This paper presents a brief review of these aims. PMID- 15297905 TI - The US National Comorbidity Survey Replication (NCS-R): design and field procedures. AB - The National Comorbidity Survey Replication (NCS-R) is a survey of the prevalence and correlates of mental disorders in the US that was carried out between February 2001 and April 2003. Interviews were administered face-to-face in the homes of respondents, who were selected from a nationally representative multi stage clustered area probability sample of households. A total of 9,282 interviews were completed in the main survey and an additional 554 short non response interviews were completed with initial non-respondents. This paper describes the main features of the NCS-R design and field procedures, including information on fieldwork organization and procedures, sample design, weighting and considerations in the use of design-based versus model-based estimation. Empirical information is presented on non-response bias, design effect, and the trade-off between bias and efficiency in minimizing total mean-squared error of estimates by trimming weights. PMID- 15297906 TI - The World Mental Health (WMH) Survey Initiative Version of the World Health Organization (WHO) Composite International Diagnostic Interview (CIDI). AB - This paper presents an overview of the World Mental Health (WMH) Survey Initiative version of the World Health Organization (WHO) Composite International Diagnostic Interview (CIDI) and a discussion of the methodological research on which the development of the instrument was based. The WMH-CIDI includes a screening module and 40 sections that focus on diagnoses (22 sections), functioning (four sections), treatment (two sections), risk factors (four sections), socio-demographic correlates (seven sections), and methodological factors (two sections). Innovations compared to earlier versions of the CIDI include expansion of the diagnostic sections, a focus on 12-month as well as lifetime disorders in the same interview, detailed assessment of clinical severity, and inclusion of information on treatment, risk factors, and consequences. A computer-assisted version of the interview is available along with a direct data entry software system that can be used to keypunch responses to the paper-and-pencil version of the interview. Computer programs that generate diagnoses are also available based on both ICD-10 and DSM-IV criteria. Elaborate CD-ROM-based training materials are available to teach interviewers how to administer the interview as well as to teach supervisors how to monitor the quality of data collection. PMID- 15297908 TI - European Head and Neck Society-EHNS. PMID- 15297907 TI - Clinical calibration of DSM-IV diagnoses in the World Mental Health (WMH) version of the World Health Organization (WHO) Composite International Diagnostic Interview (WMHCIDI). AB - An overview is presented of the rationale, design, and analysis plan for the WMH CIDI clinical calibration studies. As no clinical gold standard assessment is available for the DSM-IV disorders assessed in the WMH-CIDI, we adopted the goal of calibration rather than validation; that is, we asked whether WMH-CIDI diagnoses are 'consistent' with diagnoses based on a state-of-the-art clinical research diagnostic interview (SCID; Structured Clinical Interview for DSM-IV) rather than whether they are 'correct'. Consistency is evaluated both at the aggregate level (consistency of WMH-CIDI and SCID prevalence estimates) and at the individual level (consistency of WMH-CIDI and SCID diagnostic classifications). Although conventional statistics (sensitivity, specificity, Cohen's kappa) are used to describe diagnostic consistency, an argument is made for considering the area under the receiver operator curve (AUC) to be a more useful general-purpose measure of consistency. In addition, more detailed analyses are used to evaluate consistency on a substantive level. These analyses begin by estimating prediction equations in a clinical calibration subsample, with WMH-CIDI symptom-level data used to predict SCID diagnoses, and using the coefficients from these equations to assign predicted probabilities of SCID diagnoses to each respondent in the remainder of the sample. Substantive analyses then investigate whether estimates of prevalence and associations when based on WMH-CIDI diagnoses are consistent with those based on predicted SCID diagnoses. Multiple imputation is used to adjust estimated standard errors for the imprecision introduced by SCID diagnoses being imputed under a model rather than measured directly. A brief illustration of this approach is presented in comparing the precision of SCID and predicted SCID estimates of prevalence and correlates under varying sample designs. PMID- 15297909 TI - Anaerobic 2-propanol degradation in anoxic paddy soil and the possible role of methanogens in its degradation. AB - The anaerobic degradation of 2-propanol in anoxic paddy soil was studied with soil cultures and a 2-propanol-utilizing methanogen. Acetone was the first and the major intermediate involved in the methanogenic degradation of 2-propanol. Analyses with a methanogenesis inhibitor, bacteria antibiotics, and the addition of H2 to the gas phase revealed that 2-propanol oxidation to acetone directly occurred using 2-propanol-utilizing methanogens, but not with H2-producing syntrophic bacteria, for which the removal of acetone is required for complete 2 propanol oxidation. The 2-propanol-utilizing strain IIE1, which is phylogenetically closely related to Methanoculleus palmolei, was isolated from paddy soil, and the potential role of the strain in 2-propanol degradation was investigated. 2-Propanol is one of the representative fermentation intermediates in anaerobic environments. This is the first report on the anaerobic 2-propanol degradation process. PMID- 15297910 TI - Effect of different temperature downshifts on protein synthesis by Aeromonas hydrophila. AB - The psychrotrophic bacterium Aeromonas hydrophila 7966 was subjected to cold shocks from 30 degrees C to 20 degrees C, 15 degrees C, 10 degrees C, or 5 degrees C, or were incubated at low temperature to determine its adaptative response. The cell protein patterns analyzed by two-dimensional electrophoresis revealed that only a few proteins were underexpressed, whereas numerous new proteins appeared with the decrease of temperature, and some others were overexpressed. Among them, a few constituted cold shock proteins because they were transiently induced, whereas others belong to the acclimatation family proteins. Two cold shock proteins of 11 kDa were synthesized at low level because they were visualized only after radiolabeling or silver staining. Moreover, under our experimental conditions, no major cold shock protein of a molecular mass similar to that of E. coli (7.4 kDa) could be identified. PMID- 15297911 TI - Cloning and characterization of a mosquito larvicidal toxin produced during vegetative stage of Bacillus sphaericus 2297. AB - The mosquitocidal toxin 1 (mtx1) gene from genomic DNA of B. sphaericus strain 2297 was cloned and expressed in E. coli. DNA sequencing analysis of the cloned gene revealed a single open reading frame encoding an 870-amino acid polypeptide. Expression level of the full-length gene in E. coli was very low even though strong promoter was used or the gene was expressed as a fusion protein. Expression level was highly improved after the putative leader sequence was deleted, and the truncated gene was expressed as a fusion protein with glutathione S-transferase (GST-tMtx1). E. coli cells expressing GST-tMtx1 was highly toxic to Culex quinquefasciatus larvae and showed lower toxicity against Anopheles dirus and Aedes aegypti larvae. Enterobacter amnigenus An11, a mosquito larval gut colonizable bacteria, transformed with the cloned gene exhibited mosquito larvicidal activity. Result suggested that there is a potential to develop this protein to be used as an alternative mosquito control agent. PMID- 15297912 TI - Temperature control of a 3,4-dihydroxybenzoate (protocatechuate)-based siderophore in Bacillus anthracis. AB - Bacillus anthracis Sterne produced a catecholate siderophore named anthrachelin that was based on 3,4-dihydroxybenzoic acid (3,4-DHB, or protocatechuic acid), a catechol moiety previously unreported as a siderophore component. During iron restriction, both anthrachelin and free 3,4-DHB were excreted. Growth at 37 degrees C (as compared with 23 degrees C) decreased excretion of anthrachelin but not its precursor 3,4-DHB, suggesting that anthrachelin assembly is temperature regulated. A plasmidless strain also produced anthrachelin in an iron- and temperature-regulated fashion, indicating that anthrachelin genes are chromosomal. In addition to anthrachelin-mediated iron delivery, B. anthracis also used heme, hemoproteins, iron-transferrin, and certain heterologous siderophores (xenosiderophores) produced by other microorganisms as iron sources. Downregulation of anthrachelin production at the temperature of the mammalian host (which triggers toxin production in this pathogen) may focus the B. anthracis iron acquisition systems to exploit the iron sources prevailing in the infected host. PMID- 15297913 TI - Catabolite repression and virulence gene expression in Listeria monocytogenes. AB - Previous studies have suggested that carbohydrates may affect expression of virulence genes in Listeria monocytogenes. Which carbohydrates influence virulence gene expression and how carbohydrates mediate expression, however, is not clear. The goal of this work was to examine how carbohydrates affect virulence gene expression in L. monocytogenes 10403S. Growth studies were conducted in medium containing glucose and various sugars. Metabolism of arbutin, arabitol, cellobiose, mannose, maltose, trehalose, and salicin were repressed in the presence of glucose. Only when glucose was consumed were these sugars fermented, indicating that catabolite repression by glucose had occurred. To determine whether virulence gene expression was also influenced by catabolite repression, we performed primer extension experiments, using primers for hly and prfA, which encode for a hemolysin and the regulator protein PrfA, respectively. In the presence of cellobiose and arbutin, transcription of hemolysin was reduced. However, none of the sugars affected transcription of prfA. The results demonstrate that catabolite repression occurs in L. monocytogenes and suggests that, at least in strain 10403S, cellobiose and arbutin repress expression of hemolysin. PMID- 15297914 TI - Identification and functional analysis of dTDP-glucose-4,6-dehydratase gene and its linked gene cluster in an aminoglycoside antibiotics producer of Streptomyces tenebrarius H6. AB - Streptomyces tenebrarius H6 produces a variety of aminoglycoside antibiotics, such as apramycin, tobramycin, and kanamycin B. Primers were designed according to the highly conserved sequences of the dTDP-glucose-4,6-dehydratase genes, and a 0.6-kb PCR product was obtained from S. tenebrarius H6 genomic DNA. With the 0.6-kb PCR product as a probe, a BamHI 7.0-kb fragment was isolated. DNA sequence analysis of the 7.0-kb fragment revealed four ORFs and an incomplete ORF. In search of databases, the deduced product of one ORF (orfE) showed 62% identity to the dTDP-glucose-4,6-dehydratase, StrE of S. griseus. Three other ORFs (orfG1, orfG2, and orfGM) showed 55%, 62%, and 42% similarities, respectively, to glycosyltransferase from Clostridium acetobutylicum and mannosyltransferase from Xanthomonas axonopodis pv. citri str. 306 and glycosyltransferase from Pseudomonas putida KT2440. Upstream of the orfE was an incomplete ORF, and the deduced product showed 56% similarity to dTDP-4-dehydrorhamnose, StrL from S. griseus. The function of the orfE gene was studied by targeted gene disruption. The resulting mutant failed to produce tobramycin and kanamycin B, but still produced apramycin, suggesting that the orfE gene and linked gene cluster are essential for the biosynthesis of tobramycin and kanamycin B in S. tenebrarius H6. PMID- 15297915 TI - Isolation, identification, and characterization of a novel, oil-degrading bacterium, Pseudomonas aeruginosa T1. AB - A novel, oil-degrading bacterium (strain T1) was isolated from a hot spring in Hokkaido, Japan. It efficiently degrades different types of fats and oils, including edible oil waste. When grown in a mineral salt medium containing 1% triacylglycerol (as salad oil), hydrolysis products were 1,3- and 1,2 diacylglycerols, monoacylglycerol, and free fatty acid. However, these products were almost completely consumed during cultivation at 30 degrees C for 5 days, indicating that extracellular lipase acts randomly at different sn-positions of acylglycerols and that strain T1 has a high capacity to utilize free fatty acids. Secreted lipase activity was induced by salad oil and oleic acid. This strain was a Gram-negative straight rod shaped, aerobic, with a polar flagellum, capable of growing in temperature ranges between 15 degrees C and 55 degrees C. The 16S rRNA gene sequence analysis and DNA-DNA hybridization revealed it as a new strain of Pseudomonas aeruginosa. The type strain was T1. PMID- 15297916 TI - Influence of dipeptidyl peptidase inhibitors on growth, peptidase activity, and ammonia production by ruminal microorganisms. AB - The aim was to investigate known and potential new inhibitiors of dipeptidyl peptidases (DPP) for their effects on ruminal microorganisms. Gly-Phe diazomethylketone (GPD), Ala-Ala chloromethylketone (AAC), benserazide (DL-serine 2-(2,3,4- trihydroxybenzyl) hydrazide), and diprotin A (Ile-Pro-Ile) inhibited DPP activities of Prevotella albensis, P. ruminicola, P. bryantii, P. brevis, and mixed ruminal microorganisms, though incompletely and, except for diprotin A, without absolute specificity for any of the peptidases. Leucine aminopeptidase activity of Streptococcus bovis was also inhibited by GPD and benserazide. The inhibitors had no effect on the growth of the bacteria, except for GPD, which inhibited growth of P. albensis when only peptides were available for growth. Benserazide had some inhibitory effects on the growth of Megasphaera elsdenii and Prevotella spp., even in the absence of peptides. The predatory activity of ciliate protozoa on bacteria was unaffected by DPP inhibitors. Ammonia production from casein by mixed ruminal microorganisms was inhibited significantly (P < 0.05) by AAC (29% inhibition) and benserazide (33%). It was concluded that DPP inhibitors can influence the rate of NH3 production in the rumen and may form the basis for developing protein-sparing feed additives for ruminants. PMID- 15297917 TI - A molecular method for detection of Aspergillus carbonarius in coffee beans. AB - Ochratoxin A (OTA) is a carcinogenic and nephrotoxic mycotoxin that has been detected in a variety of food products, including green coffee beans. About 80% of Aspergillus carbonarius strains collected from coffee beans are able to produce OTA on this substrate. The rapid identification of this fungal species would be desirable. RAPD assays were applied to identify amplification products specific for A. carbonarius. One selected fragment, denoted OPX7809, was cloned and sequenced. Based on the nucleotide sequence obtained, specific oligonucleotides (OPX7F809 and OPX7R809) were designed and used as primers for DNA amplification. One amplified band of 809 bp was obtained from A. carbonarius genomic DNA, whereas no amplified fragment from DNA of other Aspergillus species was detected. This PCR analysis was also successfully employed to detect A. carbonarius in coffee beans. This PCR assay could contribute to the early and rapid detection of the potential presence of OTA in coffee samples. PMID- 15297918 TI - Cloning, function, and expression of sanS: a gene essential for nikkomycin biosynthesis of Streptomyces ansochromogenes. AB - A 2-kb SmaI DNA fragment was cloned from the cosmid library of Streptomyces ansochromogenes. This DNA fragment contains a complete open reading frame which is 1275 bp in length, designated sanS (GenBank accession no. AF322179). The deduced SanS protein consists of 424 amino acids and belongs to a superfamily of enzymes with an unusual ATP-grasp fold. The disruption and complementation of sanS indicated that sanS is essential for nikkomycin biosynthesis in Streptomyces ansochromogenes. The sanS gene was subcloned into expression vector pET23b and overexpressed in E. coli BL21 (DE3). The protein was then purified and showed ATPase activity. PMID- 15297919 TI - Enantioselective hydrolysis of butyl 2-ethylhexanoate by a strain of Nocardia corynebacteroides. AB - The results of a screen for microbial esterases that have enantioselective activity for the hydrolysis of butyl 2-ethylhexanoate are described. The preliminary screen determined that a nocardioform bacterial strain, NRRL 21057, exhibited significant activity in preferentially hydrolyzing the S enantiomer of butyl 2-ethylhexanoate. Molecular systematics methods identified NRRL 21057 as a strain of Nocardia corynebacteroides. A survey of phylogenetically related species in the genera Gordonia, Rhodococcus, and Nocardia strains demonstrated that N. corynebacteroides NRRL 21057 is the most active strain known for the specific hydrolysis of the R-isomer of butyl 2-ethylhexanoate and that it provides the S-isomer of 2-ethylhexanoate in 86% enantiomeric excess within 22 h. PMID- 15297920 TI - Intracellular location and survival of Mycoplasma penetrans within HeLa cells. AB - Mycoplasma penetrans invades HeLa cells and survives within them for prolonged periods of time. The intracellular distribution of M. penetrans within HeLa cells was studied utilizing the acidotropic dye LysoTracker (green), which permeates cell membranes and upon protonation remains trapped in acidic compartments. The excitation and emission spectra of the green LysoTracker are suitable for colocalization studies with rabbit anti- M. penetrans antibodies and red Cy5 goat anti-rabbit IgG. The images collected by confocal laser scanning microscopy revealed that in the infected HeLa cells almost all Cy5 fluorescent foci (red) were located within the LysoTrack-labelled intracellular compartments, apparently endosomes. Viable mycoplasmas were detected within endosomes for prolonged periods of time, apparently due to a potent antioxidant activity detected in M. penetrans. PMID- 15297922 TI - Molecular characteristics of phosphoenolpyruvate: mannose phosphotransferase system in Streptococcus bovis. AB - To elucidate the regulatory mechanism of catabolite control in Streptococcus bovis, we investigated the molecular properties and gene expression of the mannose-specific phosphoenolpyruvate (PEP)-dependent sugar: phosphotransferase system (PTS). The mannose PTS gene cluster (man) was found to comprise a gene encoding enzyme (E) II AB (manL) and genes encoding EIIC (manM), EIID (manN), and a putative regulator (manO). The gene cluster (man operon) was transcribed from one transcriptional start site, which was located 40 bp upstream of the manL start codon. However, two transcriptional start sites were found between manN and manO in primer extension analysis, and the manO may be transcribed independently from the man operon. The man operon and manO were constitutively transcribed without being affected by culture conditions, such as the sugar supplied (glucose, galactose, fructose, maltose, lactose, sucrose, or mannose), growth rate, or pH. PMID- 15297923 TI - Wolbachia replication and host cell division in Aedes albopictus. AB - Wolbachia pipientis is an obligate intracellular endosymbiont of a range of arthropod species. The microbe is best known for its manipulations of host reproduction that include inducing cytoplasmic incompatibility, parthenogenesis, feminization, and male-killing. Like other vertically transmitted intracellular symbionts, Wolbachia's replication rate must not outpace that of its host cells if it is to remain benign. The mosquito Aedes albopictus is naturally infected both singly and doubly with different strains of Wolbachia pipientis. During diapause in mosquito eggs, no host cell division is believed to occur. Further development is triggered only by subsequent exposure of the egg to water. This study uses diapause in Wolbachia-infected Aedes albopictus eggs to determine whether symbiont replication slows or stops when host cell division ceases or whether it continues at a low but constant rate. We have shown that Wolbachia densities in eggs are greatest during embryonation and then decline throughout diapause, suggesting that Wolbachia replication is dependent on host cell replication. PMID- 15297924 TI - Cell surface display of the chlamydial glycolipid exoantigen (GLXA) demonstrated by antibody-dependent complement-mediated cytotoxicity. AB - The chlamydial species are Gram-negative bacterial pathogens critical to human health. Their developmental cycle is associated with the formation and release of the broadly conserved glycolipid exoantigen (GLXA), which has been implicated in the chlamydial elementary body-host cell interaction. This study examines the potential surface display of this glycolipid by chlamydiae-infected cells and the ability of the GLXA they secrete to associate with the plasma membranes of uninfected cells, a prerequisite for exerting influence on them. The sequential incubation of anti-GLXA antibody and complement with Chlamydia trachomatis serovar K or C. pneumoniae AR-39-infected HeLa 229 or macrophage cells resulted in significant cellular cytotoxicity, which preceded the formation of mature elementary bodies. For uninfected cells, co-incubation of GLXA, purified from supernatants of either C. trachomatis or C. pneumoniae-infected HeLa 229 cells, followed by the successive addition of mouse anti-GLXA antibody and complement, yielded similar levels of cellular cytotoxicity. Thus, GLXA indeed is displayed on the surface of infected cells and, therefore, if antibody of appropriate specificity were present, this GLXA could serve to target these infected cells for elimination. Furthermore, released GLXA can associate with uninfected cells and therefore would be positioned to influence their behavior, especially in the context of infection. PMID- 15297925 TI - Competition of Bacillus thuringiensis Cry1 toxins for midgut binding sites: a basis for the development and management of transgenic tropical maize resistant to several stemborers. AB - Binding and competition of five Bacillus thuringiensis toxins--Cry1Ab, Cry1Ac, Cry1Ba, Cry1Ca, and Cry1Ea--for midgut binding sites from three pests, Spodoptera frugiperda, Diatraea saccharalis, and Diatraea grandiosella, were investigated as part of a strategy to develop tropical transgenic maize resistant to several stemborers. On S. frugiperda, Cry1Ab and Cry1Ac compete for the same binding site; Cry1Ba and Cry1Ca compete for a second binding site. Cry1Ea recognizes a third specific binding site in S. frugiperda and does not compete with any of the other toxins. On D. grandiosella and D. saccharalis, Cry1Ac competes with Cry1Ab and not with Cry1Ba and Cry1Ca. Cry1Ba and Cry1Ca recognize each a specific binding site and do not compete with any of the other four toxins. Cry1Ea does not recognize any binding site on Diatraea species. Combinations of toxins are proposed to develop transgenic maize resistant to the three stemborers while allowing resistance management. PMID- 15297926 TI - Ca2+ dependence and inhibitory effects of trifluoperazine on plasma membrane ATPase of Thermoactinomyces vulgaris. AB - Ca2+ enhanced the plasma membrane Ca(2+)-ATPase (PMCA) specific activities in wild-type strain 1227 and mutant strains 1278, 1286, and 1261 of Thermoactinomyces vulgaris. The Ca(2+)-ATPase specific activities showed marked increase with increasing concentrations of Ca2+ added in the form of CaCl2 in the culture medium and reached the optimum values at 0.6 mM in strains 1227, 1278, and 1286 and at 0.7 mM in strain 1261 of T. vulgaris. Trifluoperazine, a specific blocker of calmodulin, when added in vivo at concentrations of 2 microM and 8 microM along with the respective optimal concentrations of Ca2+, decreased the PMCA-specific activities to a low level in a dose-dependent manner. The results of the present investigation suggest the presence of a Ca(2+)-dependent protein activator (CaDPA) in the microenvironment constituting this enzyme; and such Ca(2+)-modulated protein has been assigned to play an important role in the enhancement of PMCA levels in this aerobic, spore-forming, thermophilic actinomycete. PMID- 15297927 TI - The UV response in Saccharomyces cerevisiae involves the mitogen-activated protein kinase Slt2p. AB - Exposure to UV causes a response in yeast and mammalian cells, which is distinct from the response to DNA damage. We report that the mitogen-activated protein kinase Slt2p is involved in this response in Saccharomyces cerevisiae. Thus, budding yeast and mammalian cells respond to UV by using very similar signal transduction pathways. PMID- 15297928 TI - Polysaccharides of pseudomonas pathovar strains that infect pea, tomato, and soya bean. AB - In order to understand the mode of action of taxonomically related Pseudomonas syringae pathovar strains that infect pea, tomato, and soya bean, we examined their extracellular polysaccharides (EPS). Maximum production of polysaccharide in shake culture of these pathogens was observed between 24 and 60 h. P. syringae pv. pisi 519, the bacterial blight pathogen of pea, produced a higher amount of polysaccharide (34.87 microg/mL) at 60 h compared with 32.67 microg/mL produced by P. syringae pv. glycinea NCPPB 1783, the bacterial blight pathogen of soya bean, and 30.03 microg/mL produced by P. syringae pv. tomato NCPPB 269, the bacterial speck pathogen of tomato. EPS produced by P. syringae pv. pisi 519, P. syringae pv. tomato NCPPB 269, and P. syringae pv. glycinea NCPPB 1783 was characterized with infrared (FTIR), nuclear magnetic resonance (NMR), high performance thin layer chromatography, (HPTLC), and matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry. HPTLC profiles revealed the presence of glucose and glucuronic acid in all bacteria and mannose only in P. syringae pv. tomato. Molecular mass of EPS of P. syringae pv. pisi (m/z 933.8), P. syringae pv. tomato (m/z 950.4), and P. syringae pv. glycinea (m/z 933.5) was confirmed by MALDI-TOF mass spectrometry. PMID- 15297929 TI - Respirometric activities of heterotrophic and nitrifying populations in aerobic granules developed at different substrate N/COD ratios. AB - Aerobic granules were successfully developed at substrate N/COD ratios ranging from 5/100 to 30/100 by weight. By measuring respective respirometric activities of heterotrophic, ammonia-oxidizing, and nitrite-oxidizing bacteria, it was found that the relative abundance of nitrifying bacteria over heterotrophs in aerobic granules was closely related to the substrate N/COD ratios. Results further showed that the populations of both ammonia and nitrite oxidizers were significantly enriched with the increase of the substrate N/COD ratio, while a decreasing trend of heterotrophic population was observed in the aerobic granules. These seem to indicate that high substrate N/COD ratio favors the selection of nitrifying bacteria in the aerobic granules, while the relative activity of nitrifying population against heterotrophic population evolved until a balance between two populations was reached in the aerobic granular sludge community. Moreover, cell elemental composition was correlated with the shift in microbial populations, e.g., the enriched nitrifying population in the aerobic granules resulted in a high cell nitrogen content normalized to cell carbon content. This study provides a good insight into microbial interaction in aerobic granules. PMID- 15297930 TI - Characterization of the DNA adenine 5'-GATC-3' methylase HpyIIIM from Helicobacter pylori. AB - The effect of inactivation of the 5'-GATC-3' methylase HpyIIIM in Helicobacter pylori (H. pylori) on mismatch repair, adherence, and in vitro fitness was examined. Chromosomal DNA from 90 H. pylori strains was isolated, and restriction enzyme digestion indicated all strains examined possess HpyIIIM. Wild-type H. pylori and a strain with an inactive HpyIIIM were found to have rifampicin mutation frequencies of 2.93 x 10(-7) and 1.05 x 10(-7) (p > 0.05), respectively, indicating that HpyIIIM does not appear to be important in mismatch repair. Adherence of H. pylori in an in vitro model cell system was also unaffected by inactivation of HpyIIIM. Inactivation of HpyIIIM did not result in a decrease in fitness, as determined by liquid in vitro competition experiments. PMID- 15297931 TI - Metalloproteases secreted by Actinobacillus suis. AB - Actinobacillus suis secretes metalloproteases into its medium. These secreted proteins, when concentrated by precipitation with 70% (NH4)2SO4 or methanol, displayed proteolytic activity at >200 kDa molecular mass bands in 10% polyacrylamide gels copolymerized with bovine casein (1%). They showed activity in a broad pH range (from pH 5 to pH 10) and were inhibited by 20 mM EDTA or EGTA, but could be reactivated by calcium. They were found heat stable at 40 degrees C, 50 degrees C, 60 degrees C, and 70 degrees C, but their activity diminished at 80 degrees C or higher. They degraded pig and bovine IgG and cross reacted with a polyclonal serum against a high molecular mass secreted protease from A. pleuropneumoniae. Extracellular proteases could play a role in diseases caused by A. suis. PMID- 15297932 TI - Isolation and some properties of fimbriae of oral Streptococcus intermedius. AB - Streptococcus intermedius 1208-1 carried linear fiber-like fimbriae that extended radially from the cell surface. The fimbriae were isolated by pipetting and sonication and were purified by ammonium sulfate precipitation followed by a column chromatography series. Heat treatment in the presence of sodium dodecyl sulfate resulted in the dissociation into smaller molecules. Rabbit antiserum raised against the purified protein reacted with fimbriae on the surface of bacteria under immunogold staining. Serotype g or g-related strains produced the fimbriae and aggregated in human saliva. The aggregation was inhibited by the anti-fimbriae immunoglobulin Fab fragment or the purified fimbriae. PMID- 15297933 TI - The Metarhizium anisopliae trp1 gene: cloning and regulatory analysis. AB - The trp1 gene from the entomopathogenic fungus Metarhizium anisopliae, cloned by heterologous hybridization with the plasmid carrying the trpC gene from Aspergillus nidulans, was sequence characterized. The predicted translation product has the conserved catalytic domains of glutamine amidotransferase (G domain), indoleglycerolphosphate synthase (C domain), and phosphoribosyl anthranilate isomerase (F domain) organized as NH2-G-C-F-COOH. The ORF is interrupted by a single intron of 60 nt that is position conserved in relation to trp genes from Ascomycetes and length conserved in relation to Basidiomycetes species. RT-PCR analysis suggests constitutive expression of trp1 gene in M. anisopliae. PMID- 15297934 TI - Attention deficit hyperactivity disorder: fine mapping supports linkage to 5p13, 6q12, 16p13, and 17p11. AB - We completed fine mapping of nine positional candidate regions for attention deficit/hyperactivity disorder (ADHD) in an extended population sample of 308 affected sibling pairs (ASPs), constituting the largest linkage sample of families with ADHD published to date. The candidate chromosomal regions were selected from all three published genomewide scans for ADHD, and fine mapping was done to comprehensively validate these positional candidate regions in our sample. Multipoint maximum LOD score (MLS) analysis yielded significant evidence of linkage on 6q12 (MLS 3.30; empiric P=.024) and 17p11 (MLS 3.63; empiric P=.015), as well as suggestive evidence on 5p13 (MLS 2.55; empiric P=.091). In conjunction with the previously reported significant linkage on the basis of fine mapping 16p13 in the same sample as this report, the analyses presented here indicate that four chromosomal regions--5p13, 6q12, 16p13, and 17p11--are likely to harbor susceptibility genes for ADHD. The refinement of linkage within each of these regions lays the foundation for subsequent investigations using association methods to detect risk genes of moderate effect size. PMID- 15297935 TI - Linkage disequilibrium and association of MAPT H1 in Parkinson disease. AB - The MAPT H1 haplotype has been associated with four-repeat (4R) tauopathies, including progressive supranuclear palsy, corticobasal degeneration, and argyrophilic grain disease. More controversial is that the same haplotype has been associated with Parkinson disease (PD). Using H1-specific single-nucleotide polymorphisms, we demonstrate that MAPT H1 is a misnomer and consists of a family of recombining H1 alleles. Population genetics, linkage disequilibrium, and association analyses have shown that specific MAPT H1 subhaplotypes are preferentially associated with Parkinson disease. Using a sliding scale of MAPT H1-specific haplotypes--in age/sex-matched PD cases and controls from central Norway--we have refined the disease association to within an approximately 90-kb interval of the 5' end of the MAPT locus. PMID- 15297937 TI - Are cancers of the esophagus, gastroesophageal junction, and cardia one disease, two, or several? AB - Recent epidemiological shifts have led to controversy about the etiology and treatment of gastroesophageal (GE) junction adenocarcinomas. The Siewert classification conveniently describes the anatomical location of these tumors relative to the GE junction. However, available data suggest that there may be epidemiological, clinical, and molecular differences among GE junction adenocarcinomas, depending on whether they are predominantly esophageal or gastric in location. Much more investigation is required to confirm or refute these apparent differences, which can affect clinical treatment. PMID- 15297938 TI - Epidemiology of upper gastrointestinal malignancies. AB - The demographics of esophageal and gastric cancer have been changing dramatically in the United States over the past several decades. While incidence rates for esophageal squamous cell carcinoma and distal gastric carcinoma have been declining, the trends for adenocarcinoma of the esophagus and proximal stomach have been rising rapidly, particularly among white males. The incidence of these upper gastrointestinal (GI) malignancies varies widely based on geographic location, race, and socioeconomic status. The primary causes of squamous cell carcinoma of the esophagus are tobacco use and alcohol consumption, whereas the main risk factors for adenocarcinoma of the esophagus are gastroesophageal reflux disease and obesity. Dietary factors and Helicobacter pylori infection play an important role in the development of gastric cancer. Understanding the epidemiology and etiologies of esophageal and gastric carcinomas will lead to the development of interventions for screening and prevention in high-risk populations. PMID- 15297939 TI - Pathology of upper gastrointestinal malignancies. AB - This review describes the most important macroscopic, histological, and molecular biological features of carcinomas of the esophagus, the gastroesophageal junction (GEJ), and the stomach. The three most frequent histological subtypes of upper GI tract carcinoma, ie, adenocarcinoma, squamous cell carcinoma, and small cell carcinoma, are considered. Important histological classifications of upper GI tract adenocarcinomas such as Lauren's classification and the World Health Organization (WHO) classification, as well as rare histological variants of esophageal and gastric cancer, are described. Routes of tumor spread and principles of tumor classification are outlined. Furthermore, descriptions of the immediate precursor lesions of upper GI tract carcinomas, ie, dysplasias, are included. Finally, somatic genetic and epigenetic changes, which are associated with tumor development in the upper GI tract, are discussed. PMID- 15297940 TI - Molecular biology of upper gastrointestinal malignancies. AB - While cancers of the upper gastrointestinal tract are related by location, they have distinct clinical and molecular characteristics. This review will focus on the molecular biology of esophageal adenocarcinoma (EAC) of which alterations in p53, overexpression of cyclin D1, loss of p16, and aneuploidy have been best characterized. Key similarities and differences, when compared to esophageal squamous cell carcinoma (ESCC), gastric adenocarcinoma (GC), and adenocarcinoma of the gastric cardia (AGC), will also be highlighted. Currently, malignancies of the upper gastrointestinal tract are often diagnosed at an advanced stage and are generally associated with a poor patient prognosis. With an improved understanding of the molecular biology of these tumors, there is hope that new targets for diagnosis, chemoprevention, and therapy will be developed. PMID- 15297941 TI - Screening for upper gastrointestinal tract malignancies. AB - The screening for malignancies involving the upper gastrointestinal (GI) tract has taken on new importance as the incidence of cancers such as adenocarcinoma of the esophagus has risen. The availability of new screening technology and molecular biological techniques has dramatically improved our ability to detect early lesions. We have also developed a detailed understanding of the natural history of many upper GI malignancies. This new knowledge has led to significant improvement in population-based screening programs. In this review we will detail the evidence supporting our current screening methods and provide a concise overview of current screening recommendations for select upper GI malignancies. PMID- 15297942 TI - Premalignant lesions of the esophagogastric mucosa. AB - Premalignant esophagogastric (EG) lesions develop against a background of chronic inflammation, called a premalignant condition. For esophageal squamous cell cancer, causal factors include alcohol, tobacco, hot beverages, oral consumption of opioids, and probably infectious agents. For adenocarcinoma in the Barrett's esophagus (BE), gastroesophageal reflux disease (GERD) is the principal causal factor. At the EG junction, adenocarcinoma arises either from the esophagus or from the proximal stomach (cardia). In the distal stomach, chronic gastritis with atrophy is the premalignant condition related to Helicobacter pylori infection. A high intake of salt and low intake of antioxidants also play a role. The histopathology of EG premalignant lesions is now included in the groups low-grade and high-grade intraepithelial neoplasia (IEN) of the revised Vienna classification. Endoscopy is the gold standard for detection of the lesions at the preclinical stage and their appearance is described in subtypes of the type 0 of the Japanese classification, with a distinction between protruding and nonprotruding lesions. There is a priority for primary prevention of causal factors rather than for mass screening, which is justified only in Japan for the prevention of stomach cancer. The trend to early detection of premalignant lesions justifies the development of mini-invasive endoscopic procedures of treatment. PMID- 15297943 TI - Staging and preoperative evaluation of upper gastrointestinal malignancies. AB - Esophageal and gastric cancers are distinct carcinomas of the upper gastrointestinal tract, although the distinction between them becomes less clear at the gastroesophageal junction (GEJ). Increasingly accurate staging is possible based on newer radiographic and surgical techniques such as positron emission tomography (PET), laparoscopy and thoracoscopy, laparoscopic ultrasound, and endoscopic ultrasound (EUS). For both cancer types, tumor classification is determined by depth of penetration of the primary tumor into the gastric or esophageal wall. For esophageal cancer, primary tumor anatomic position-upper, mid, and lower esophagus-is used to define the local nodal basin. Metastases in nodes outside the local basin are considered to be distant (M) rather than regional (N). In gastric cancer, the region of nodal metastasis has been abandoned in favor of the number of lymph nodes containing metastasis, which predicts outcome more accurately-patients with more than 15 positive lymph nodes have an outcome comparable to those with M disease. Increasing consideration is being given to the subclassification of tumors near the GEJ into types based on anatomical position, although this staging scheme ("Adenocarcinoma of the EsophagoGastric junction" or AEG type) has not yet been universally adopted. We review the current pathologic staging systems for esophageal and gastric cancers, the clinical staging approaches for these diseases, and the controversy surrounding classification of tumors of the GEJ. PMID- 15297944 TI - Imaging of esophageal and gastric cancer. AB - The three major aims of imaging in esophageal and gastric cancer are to distinguish between locoregional and systemic disease (M stage), to determine local tumor extension (T and N stages), and to assess response to chemotherapy or chemoradiotherapy. Depending on the applied standard of reference, the sensitivity of computed tomography (CT) for detection of distant metastases ranges between less than 50% to greater than 90%. In esophageal cancer, positron emission tomography with the glucose analog fluorodeoxyglucose (FDG-PET) has been shown to detect metastatic disease in approximately 20% of the patients who were considered to have only locoregional disease with CT. In clinical studies, endoscopic ultrasound (EUS) has been shown to differentiate between tumor stages T1/T2 and stages T3/T4 with an accuracy of more than 90%. Accuracy of EUS for differentiating individual tumor stages in routine clinical use has been reported to be markedly lower. Assessment of tumor response by FDG-PET has been shown to correlate with histopathologic tumor regression and patient survival. Furthermore, quantitative measurements of tumor FDG uptake may predict histopathologic tumor response and patient outcome as early as 2 weeks after initiation of preoperative chemotherapy. PMID- 15297945 TI - Surgery for upper gastrointestinal malignancies. AB - The incidence, diagnostic modalities, and therapeutic options for gastric and esophageal cancers have undergone important changes in the last decades. Distal gastric cancers are seen less often compared to proximal tumors. Early detection and improved perioperative care have increased their resectability. Partial stomach resection with a proper albeit nonradical lymph node dissection seems adequate treatment, and adjuvant treatment is not required for most patients. Tumors located in and around the gastroesophageal junction have become the most frequently seen upper gastrointestinal malignancies. Their anatomical relations and their early tendency to spread to thoracic and abdominal lymph nodes make these tumors among the most difficult to treat. The value and extent of surgery and the impact of additional (neo-)adjuvant therapy are less clear. It seems beneficial to treat these patients in high-volume centers and it is mandatory to increase the number of patients included in multimodality therapy trials. PMID- 15297946 TI - Treatment of localized esophageal cancer. AB - The treatment of localized esophageal cancer (LEC) is controversial. The approaches that are used in daily practice include surgery or radiation alone, preoperative or postoperative radiation, preoperative or postoperative chemotherapy, and definitive or preoperative chemoradiation. The varied modalities used to treat LEC reflect both the lack of randomized trial data and the suboptimal results with current therapy. Nonetheless, the available data suggest that surgery and definitive concurrent chemoradiation represent two standard treatment options. The use of preoperative chemoradiation remains investigational, with phase III trials failing to demonstrate a consistent benefit for this option over chemoradiation or surgery alone. This review will examine the available data on the current treatment approaches in LEC. PMID- 15297947 TI - Treatment of localized gastric cancer. AB - The curative management of gastric adenocarcinoma depends upon complete resection of the primary tumor. In patients with lymph node metastases in the resected specimen, the relapse and death rates from recurrent cancer are at least 70% to 80%. There is continued debate over whether more extensive lymph node dissection (D2) improves survival when compared to less extensive operations. Until recently, attempts at preventing recurrence have employed adjuvant chemotherapy and have been ineffective. A large US Intergroup study (INT-0116) demonstrated that combined chemoradiation following complete gastric resection improves median time to relapse (30 v 19 months, P <.0001) and overall survival (35 months v 28 months, P =.01). The improvements in disease-free and overall survival created by postoperative chemoradiation have defined a new standard of care. Also the publication of a large phase III neoadjuvant chemotherapy clinical trial using epirubicin, cisplatin, and 5-fluorouracil (5-FU) suggested that this technique may downstage tumors and increase resectability. Future advances in the therapy of resectable gastric cancer may come from studies of preoperative neoadjuvant chemoradiation and the application of targeted therapies such as growth receptor antagonists and antiangiogenesis agents. PMID- 15297948 TI - Treatment of metastatic esophagus and gastric cancer. AB - Cancers of the upper gastrointestinal tract include cancers of the esophagus, gastroesophageal junction, and stomach. Metastatic or unresectable upper gastrointestinal malignancies are incurable but do benefit from palliative chemotherapy. Several agents have been examined in the treatment of these diseases, with modest single-agent activity. In combination, these chemotherapeutic agents demonstrate improved antitumor activity with acceptable toxicity. Several combination therapies have been developed and have been examined in recent large phase III randomized clinical trials. These studies, for the most part, have failed to demonstrate a survival advantage over the reference arm. However, with the examination of newer cytotoxic agents, as well as with the application of molecularly targeted approaches to upper gastrointestinal malignancies, there remains hope for improved therapies for these diseases in the future. PMID- 15297950 TI - Improving pain management through leadership and interdisciplinary collaboration. PMID- 15297951 TI - The use of "as-needed" range orders for opioid analgesics in the management of acute pain: a consensus statement of the American Society for Pain Management Nursing and the American Pain Society. AB - The use of "as needed" or "PRN" range orders for opioid analgesics in the management of acute pain is a common clinical practice. This approach provides flexibility in dosing to meet individual patients' unique analgesic requirements. Range orders enable necessary and safe dose adjustments based on an individual's response to treatment. The purpose of this paper is to present the consensus statement of the American Society for Pain Management Nursing and the American Pain Society on the use of "as-needed" range orders for opioid analgesics in the management of acute pain. The implementation of this statement should promote quality pain management through safe medication practices and the appropriate use of range orders for opioid analgesics in acute pain management. PMID- 15297952 TI - Foot and hand massage as an intervention for postoperative pain. AB - Physiological responses to pain create harmful effects that prolong the body's recovery after surgery. Patients routinely report mild to moderate pain even though pain medications have been administered. Complementary strategies based on sound research findings are needed to supplement postoperative pain relief using pharmacologic management. Foot and hand massage has the potential to assist in pain relief. Massaging the feet and hands stimulates the mechanoreceptors that activate the "nonpainful" nerve fibers, preventing pain transmission from reaching consciousness. The purpose of this pretest-posttest design study was to investigate whether a 20-minute foot and hand massage (5 minutes to each extremity), which was provided 1 to 4 hours after a dose of pain medication, would reduce pain perception and sympathetic responses among postoperative patients. A convenience sample of 18 patients rated pain intensity and pain distress using a 0 to 10 numeric rating scale. They reported decreases in pain intensity from 4.65 to 2.35 (t = 8.154, p <.001) and in pain distress from 4.00 to 1.88 (t = 5.683, p <.001). Statistically significant decreases in sympathetic responses to pain (i.e., heart rate and respiratory rate) were observed although blood pressure remained unchanged. The changes in heart rate and respiratory rate were not clinically significant. The patients experienced moderate pain after they received pain medications. This pain was reduced by the intervention, thus supporting the effectiveness of massage in postoperative pain management. Foot and hand massage appears to be an effective, inexpensive, low-risk, flexible, and easily applied strategy for postoperative pain management. PMID- 15297953 TI - Health-related quality of life and pain beliefs among people suffering from chronic pain. AB - Chronic pain, when not effectively treated and relieved, may have a harmful effect on all aspects of health-related quality of life (HRQL). Furthermore, pain beliefs are considered an important mediating psychological factor in chronic pain. The present study focused on HRQL as measured by the Medical Outcomes Survey-Short Form (SF-36) and addressed possible relationships between pain beliefs as measured by the Pain Beliefs and Perceptions Inventory (PBAPI). The possible impact of background variables such as age, gender, social support, pain intensity, pain duration, and analgesics on HRQL were controlled for in the analyses. The study sample consisted of 81 people who were recruited from a multidisciplinary pain management program. Data were collected as the first part of a routine pretreatment evaluation. The chronic-pain patients reported lower scores on all dimensions of HRQL compared to normal controls and other patient groups. No significant association was found between pain beliefs and the physical health dimension of HRQL whereas gender, pain duration, and pain intensity were significant predictors of that dimension. In addition, one of the dimensions of pain beliefs (i.e., mystery) was found to be predictive of the mental health dimension of HRQL. Social support made an additional contribution to the explained variance in mental health. The implications of these results for assessing HRQL and pain beliefs in chronic pain are discussed. PMID- 15297958 TI - Borderline Ovarian Tumor Workshop, Bethesda, Maryland, August 27-28, 2003. PMID- 15297954 TI - The cognitive effects of opioids. AB - Successful opioid therapy often depends on achieving a balance between analgesic effectiveness and side effects. The risk of opioid-induced cognitive impairment often hinders clinicians and patients from initiating or optimizing opioid therapy. Despite subjective experiences of mental dullness and sedation, objective tests of cognitive functioning do not always demonstrate marked changes following opioid administration. To guide clinical practice, as well as patient and family teaching, pain management nurses should be familiar with literature regarding this topic. The purpose of this article is to review the empiric literature on opioids and cognitive functioning, including the relationships among pain, cognition, delirium, and opioids. In general, research reflects minimal to no significant impairments in cognitive functioning. If impairment does occur, it is most often associated with parenteral opioids administered to opioid-naive individuals. Some evidence suggests that opioids may actually enhance cognitive function and decrease delirium in some patient populations. This article describes this research and explores the clinical implications of the research in this area. PMID- 15297959 TI - Borderline ovarian tumors: key points and workshop summary. AB - This article documents major points of agreement and disagreement among experts invited to participate in a Borderline Ovarian Tumor Workshop held in Bethesda, MD, on August 27-28, 2003. It is suggested that controversies related to the diagnosis and management of these tumors are often related to lack of data in the literature (small numbers of cases, unreported or unclear criteria for diagnosis and follow-up, insufficient length of follow-up, etc), and specific recommendations are made for further investigation and for reporting of data in future studies. PMID- 15297960 TI - Borderline ovarian tumors: diverse contemporary viewpoints on terminology and diagnostic criteria with illustrative images. AB - The National Cancer Institute sponsored a Borderline Ovarian Tumor Workshop held in August 2003 in Bethesda, MD. This report was developed from discussions at the Workshop. The participants acknowledged several areas of disagreement on basic terminology issues and agreed that a glossary with example images would help clarify many commonly misunderstood issues. This report defines terminology used in the pathological description of borderline tumors and their variants, and illustrates examples of each of the most common entities. It also addresses controversial aspects of the definitions and issues involving specimen handling and reporting. For those issues where there is disagreement, the terminology and diagnostic approaches reflecting the differing views are presented. PMID- 15297961 TI - Serous borderline (low malignant potential, atypical proliferative) ovarian tumors: workshop perspectives. AB - Although the category of serous borderline ovarian tumor (S-BOT) was established more than 30 years ago, the nomenclature and prognostic significance of various histological features of these neoplasms continues to engender controversy. The Borderline Ovarian Tumor Workshop was held in Bethesda, MD, in August 2003 in an attempt to examine the existing data, establish areas of agreement, and identify areas needing further investigation. This report addresses 6 areas of controversy regarding S-BOT: (1) tumors with and without a micropapillary architecture (typical vs micropapillary type), (2) peritoneal implants, (3) stromal microinvasion, (4) ovarian surface involvement, (5) lymph node involvement, and (6) recurrent tumors. Each of these issues is addressed by summarizing the data in the literature on which the discussions were based, areas of agreement that emerged, divergent opinions and the reasoning behind them, and the conclusions of the participants with recommended nomenclature. PMID- 15297962 TI - Mucinous borderline ovarian tumors: points of general agreement and persistent controversies regarding nomenclature, diagnostic criteria, and behavior. AB - This report focuses on the borderline category of ovarian mucinous tumors and summarizes the points of general agreement and persistent controversies identified by experts in the field who participated in the Borderline Ovarian Tumor Workshop held in Bethesda, MD, in August 2003. Points of agreement and persistent controversies regarding nomenclature, diagnostic criteria, and behavior are addressed for the following ovarian mucinous tumor categories: mucinous borderline ovarian tumor (M-BOT; synonymously referred to as atypical proliferative mucinous tumor of ovary or mucinous ovarian tumor of low malignant potential), M-BOT with intraepithelial carcinoma, and M-BOT with microinvasion. The morphologic spectrum of M-BOTs with regard to distinction from mucinous cystadenoma and the confluent glandular/expansile type of invasive mucinous carcinoma is also addressed. Non-ovarian mucinous tumors, including the secondary ovarian mucinous tumors associated with pseudomyxoma peritonei and metastatic mucinous carcinomas with a deceptive pattern of invasion, are recognized as tumors that can simulate primary M-BOTs. Improved classification of these mucinous tumors has clarifed the behavior of true M-BOTs by excluding these simulators from the M-BOT category. PMID- 15297963 TI - Current challenges and opportunities for research on borderline ovarian tumors. AB - This article summarizes key issues for future research on borderline ovarian tumors that emerged at a National Cancer Institute-sponsored Borderline Ovarian Tumor Workshop held in August 2003 in Bethesda, MD. Limitations in existing research and opportunities for future advances have been highlighted. The application of new molecular techniques in combination with improved study designs holds promise for elucidating the pathogenesis of these tumors and revealing the source of the extra-ovarian lesions ("implants") with which they are frequently associated. Clarification of the etiology of borderline tumors and the pathogenesis of their associated implants is critical for improving pathological diagnosis, revising the classification system of ovarian neoplasms, and developing optimal, evidence-based clinical management algorithms. PMID- 15297964 TI - Vaccination to prevent and treat cervical cancer. AB - Human papillomaviruses (HPVs) are the primary etiologic agents of cervical cancer. Thus, cervical cancer and other HPV-associated malignancies might be prevented or treated by HPV vaccines. Transmission of papillomavirus may be prevented by the generation of antibodies to capsid proteins L1 and L2 that neutralize viral infection. However, because the capsid proteins are not expressed at detectable levels by infected basal keratinocytes or in HPV transformed cells, therapeutic vaccines generally target nonstructural early viral antigens. Two HPV oncogenic proteins, E6 and E7, are critical to the induction and maintenance of cellular transformation and are coexpressed in the majority of HPV-containing carcinomas. Thus, therapeutic vaccines targeting E6 and E7 may provide the best option for controlling HPV-associated malignancies. Various candidate therapeutic HPV vaccines are currently being tested whereby E6 and/or E7 are administered in live vectors, as peptides or protein, in nucleic acid form, as components of chimeric virus-like particles, or in cell-based vaccines. Encouraging results from experimental vaccination systems in animal models have led to several prophylactic and therapeutic vaccine clinical trials. If these preventive and therapeutic HPV vaccines prove successful in patients, as they have in animal models, then oncogenic HPV infection and its associated malignancies may be controllable by vaccination. PMID- 15297965 TI - Clinicopathologic study and laboratory diagnosis of 23 cases with West Nile virus encephalomyelitis. AB - The differences in pathologic findings of fatal cases of West Nile virus (WNV) encephalitis in the context of underlying conditions and illness duration are not well known. During 2002, we studied central nervous system (CNS) tissue samples from 23 patients who had serologic and immunohistochemical (IHC) evidence of a recent WNV infection. Fifteen patients had underlying medical conditions (5 malignancies, 3 renal transplants, 3 with diabetes or on dialysis, 2 with AIDS, and 2 receiving steroids). WNV serology was positive for 18 patients, negative for 2, and not available for 3. Perivascular lymphocytic infiltrates, microglial nodules, and loss of neurons were predominantly observed in the brainstem and anterior horns in the spinal cord. IHC using antibodies against flaviviruses and WNV showed viral antigens in 12 (52%) of 23 patients. Viral antigens were found inside neurons and neuronal processes predominantly in the brainstem and anterior horns. In general, the antigens were focal and sparse; however, in 4 severely immunosuppressed patients, extensive viral antigens were seen throughout the CNS. Positive IHC staining was observed in tissues of 7 of 8 patients who died within 1 week after illness onset, compared with 4 of 14 with more than 2 weeks' illness duration. WNV causes an encephalomyelitis by primarily affecting brainstem and spinal cord. Differences in the amount of viral antigen may be related to underlying medical conditions and length of survival. IHC can be an important diagnostic method, particularly during the 1st week of illness, when antigen levels are high. PMID- 15297966 TI - Structural organization of subgemmal neurogenous plaques in foliate papillae of tongue. AB - Little is known about subgemmal neurogenous plaques in the foliate papillae of tongue, which prompted the present investigation. The plaques were immunohistochemically studied in biopsies from 16 adults with the use of neural, stromal, basement membrane, and cell-cycle markers. They displayed a zonal pattern of organization. The neurofibroma-like superficial zone expanded lamina propria and was contiguous to the epithelium covering the papillae. It consisted of tangled composites of S-100 protein-positive spindled cells and fibrils stained for protein gene product 9.5 and neurofilament protein. The composites also expressed the CD56 antigen, could be traced into overlying taste buds, were associated with abundant laminin, and were intermingled with scattered dendritic cells expressing factor XIIIalpha and with mast cells decorated on staining for CD117. Similar composites, but arranged in fascicles and invested by epithelial membrane antigen-positive and collagen IV-positive sheaths, characterized the deep zone of the plaques. Intrafascicular CD57-positive myelin-like annuli and CD34-positive spindled cells were also features of this zone. The sheaths and, most often, the CD57-positive annuli and CD34-positive cells were progressively spread apart toward the intermediate zone of the plaques and were lost superficially. Ki67 and Bcl-1 were not expressed in the plaques. The results suggest that composites of Schwann cells and unmyelinated axons make up the superficial bulk of the plaques, whereas perineurial cells, endoneurial fibroblasts, and myelinated axons are present more deeply. It is possible that the composites achieve neuroeffector relationships and are not neoplastic. Trophic influences from gustatory nerve fibers could play a role in the development of the plaques. PMID- 15297967 TI - Expression of glutaredoxin is highly cell specific in human lung and is decreased by transforming growth factor-beta in vitro and in interstitial lung diseases in vivo. AB - Glutaredoxins (Grx) are thiol-disulfide oxidoreductases with antioxidant capacity and catalytic functions closely associated with glutathione, an antioxidant abundantly present in human lung. The present study investigated the expression of both human glutaredoxins in cultured human lung cells and lung homogenates by reverse-transcription polymerase chain reaction and Western blotting. Immunohistochemical studies were conducted with 38 human lung specimens, including healthy lung, parenchymal sarcoidosis, extrinsic allergic alveolitis, and usual interstitial pneumonia (UIP). The ultrastructural localization of Grx1 was assessed by immunoelectron microscopy. In addition, cultured airway epithelial cells were exposed to tumor necrosis factor (TNF)-alpha and transforming growth factor (TGF)-beta. Both Grx1 and Grx2 could be detected at the mRNA and protein level in cultured human lung cells, but only Grx1 was prominently expressed in lung homogenates and alveolar macrophages. Immunohistochemically, Grx1 was highly concentrated to alveolar macrophages and weakly positive in the bronchial epithelium. Grx1 was ultrastructurally localized to the plasma membrane, cytoplasmic vacuoles, and nucleus. The expression of Grx1 decreased in alveolar macrophages of sarcoidosis and allergic alveolitis compared with the case for controls (P < 0.001), and bronchial epithelium of these diseases revealed no Grx1 immunoreactivity. Fibroblast foci and other fibrotic areas in UIP were mainly negative. In A549 cells, Grx1 was down-regulated by TGF beta, whereas TNF-alpha caused no clear effect. Overall, high expression of Grx1 in alveolar macrophages suggests its importance in the primary defense of human lung. Decreased expression of Grx1 further suggests the impairment of this system both in inflammatory and fibrotic lung diseases, consistent with the down regulation of Grx1 by TGF-beta in vitro. PMID- 15297968 TI - Analysis of alpha-methylacyl-CoA racemase (P504S) expression in high-grade prostatic intraepithelial neoplasia. AB - Alpha-methylacyl-CoA racemase (AMACR), also known as P504S, is a recently identified molecular marker for prostate cancer. The expression of AMACR/P504S has also been observed in high-grade prostatic intraepithelial neoplasia (PIN), a precursor lesion of prostate cancer. However, a detailed study focusing on the analysis of AMACR/P504S expression in high-grade PIN has not been performed. In this study, we analyzed AMACR/P504S expression by immunohistochemistry in 3954 prostatic ducts and acini with high-grade PIN from 140 prostatectomy specimens. AMACR/P504S immunoreactivity was measured as negative (0), weakly positive (+1), moderately positive (+2), and strongly positive (+3). AMACR/P504S immunoreactivity was detected in 90.0% (126/140) of high-grade PIN cases, although only 41.5% (1642/3954) of prostatic glands involved by PIN showed AMACR/P504S immunoreactivity. A significantly higher AMACR/P504S-positive rate (56.0%) was found in isolated high-grade PIN glands adjacent to cancer (distance less than 5 mm) compared with those away from cancer (distance more than 5 mm; 14%, P < 0.0001). High-grade PIN glands adjacent to cancer also showed a higher (P < 0.0004) AMACR/P504S intensity (1.62) than did those away from cancer (1.11). Our results suggest that PIN strongly positive for AMACR/P504S might be more closely associated with cancer than PIN negative or weakly positive for AMACR/P504S. This study provides additional evidence to link high-grade PIN as a precursor lesion to prostatic adenocarcinoma. PMID- 15297969 TI - Expression of candidate tumor markers in ovarian carcinoma and benign ovary: evidence for a link between epithelial phenotype and neoplasia. AB - EpCAM, epithelial membrane antigen (EMA)-mucin 1 (MUC1), mesothelin, and CD9 have been reported to be overexpressed at the RNA level in ovarian carcinomas. By using immunohistochemistry, we profiled the protein expression of these gene products in ovarian carcinoma tissues and compared them with benign ovarian surface epithelium (OSE) and cortical inclusion cysts (CICs). Immunoreactivity for EMA and calretinin were used to define epithelial and mesothelial differentiation in nontumor tissues, respectively. Papillary serous (n = 16) and endometrioid (n = 10) tumors were immunopositive for EMA/MUC1 (100%), mesothelin (75% and 30%, respectively), CD9 (88% and 90%, respectively), and EpCAM (100%). All ovarian carcinomas and carcinoma cell lines tested were negative for calretinin. In nonneoplastic ovary, both OSE and CICs ranged from flat-to cuboidal to stratified and ciliated in appearance. OSE with a cuboidal morphology had a similar immunoreactivity as omental peritoneum, expressing calretinin, mesothelin, and CD9. In contrast, CICs with stratified and ciliated epithelium show expression patterns similar to those in fallopian tubes. They frequently expressed EMA, EpCAM, mesothelin, and CD9. This immunophenotype is preserved in ovarian carcinomas, suggesting that Mullerian metaplasia signals the acquisition of these markers and that their expression is maintained in ovarian carcinomas that originate from this epithelium. PMID- 15297970 TI - XIAP expression is an independent prognostic marker in clear-cell renal carcinomas. AB - Deregulation of apoptosis plays an important role in carcinogenesis, tumor progression, and resistance to chemotherapy. XIAP is considered to be the most potent caspase inhibitor of all known IAP (inhibitor of apoptosis) family members. To explore the relevance of XIAP for progression and prognosis in renal cell carcinomas (RCCs) of the clear-cell type, we analyzed XIAP protein expression in formalin-fixed tissue from 145 clear-cell RCCs by immunohistochemistry. XIAP protein expression was found in 95% of clear-cell RCCs. A significant increase of XIAP expression became evident from well (G1) to poorly (G3) differentiated clear-cell RCCs (P < 0.0001) and from low (pT1) to advanced (pT3) tumor stages (P = 0.0016). Log-rank test showed a significant inverse correlation (P = 0.0174) between XIAP expression and tumor aggressiveness as indicated by patients' survival. Most important, multivariate Cox regression analysis revealed that XIAP expression is an independent prognostic parameter (P = 0.018) in clear-cell RCCs. Our results suggest an important role for XIAP mediated inhibition of apoptosis during progression of clear-cell RCCs and introduce XIAP expression as a new independent prognostic marker in this tumor type. PMID- 15297971 TI - Inverse correlation between p16INK4A expression and NF-kappaB activation in melanoma progression. AB - Expression of p16INK4A, the product of the melanoma susceptibility gene CDKN2A, has been shown to decrease in correlation with tumor progression. P16INK4A is a key regulator of cell-cycle function, and likely interacts with a variety of targets alongside cyclin-dependent kinases (CDKs). One such target is nuclear factor KB (NF-kappaB), a pleiotropic transcription factor that plays a crucial role in apoptosis, oncogenesis and cell cycle control. NF-kappaB p65 has been shown to be activated in melanoma cell lines but few studies decribe its expression in the tissue. In the present study we focused on synchronous expression of p16INK4A and NF-kappaB p65 and their functional activation in melanoma cell lines and biopsy tissue. Activation of NF-kappaB p65, as observed by electrophoretic mobility shift assay in cell lines, was correlated with expression and cellular localization of the active and inactive forms of its inhibitor, IkappaB-alpha. In melanocytic lesions, p16INK4A and NF-kappaB p65 expression were inversely correlated with levels of the nuclear component of NF kappaB p65 increasing from nevi to primary melanomas and metastases. PMID- 15297972 TI - Heterogeneity of genomic breakpoints in MSN-ALK translocations in anaplastic large cell lymphoma. AB - Anaplastic large cell lymphomas (ALCL) are associated with the t(2;5)(p23;q35) translocation involving the anaplastic lymphoma kinase (ALK) and the nucleophosmin (NPM) genes. However, genes other than NPM may fuse to ALK in these tumors. In this study we have identified an ALCL with a distinctive cell membrane restrictive ALK immunostaining in which the molecular characterization showed a new fusion gene between moesin (MSN) and ALK with different breakpoints than previously recognized. The ALK breakpoint occurred in an exonic sequence, and the chimeric gene included an intronic sequence of MSN. Identification of the genomic breakpoint in the derivative chromosome 2 revealed a 72-base pair deletion involving both MSN and ALK sequences. These findings provide further evidence of the breakpoint heterogeneity in ALK translocations and highlight the importance of ALK immunostaining in the diagnosis of ALCL and the identification of the underlying genetic abnormalities in this lymphoma. PMID- 15297973 TI - AA-type amyloidosis associated with non-Hodgkin's lymphoma: a case report. AB - Amyloid-associated protein (AA)-type systemic amyloidosis has been referred to as secondary amyloidosis because it is secondary to an associated inflammatory condition. It is extremely rare in patients with non-Hodgkin's lymphoma (NHL). Here we report an autopsy case of follicular small cleaved cell lymphoma with focal large B-cell lymphoma transformation in association with systemic AA-type amyloidosis. Formalin-fixed, paraffin-embedded tissues from autopsy and the patient's previous surgical specimen were studied by Congo red stain; electron microscopy; and immunostaining with antibodies against AA protein, P component, and kappa and lambda light chains. There was a marked AA amyloid deposition in the glomeruli of both kidneys, the retroperitoneal lymphoma mass, the blood vessels, the adrenal glands, and the adipose tissues. The patient's previous surgical specimens were negative for amyloid. We propose that this patient's systemic AA-type amyloidosis developed along the course of his NHL. PMID- 15297979 TI - Abstracts of the World Federation of Societies of Biological Psychiatry 2004 WFSBP Asia-Pacific Congress. July 9-11, 2004. Seoul, Korea. PMID- 15297974 TI - Tuberculous tenosynovitis. AB - Tuberculous tenosynovitis is rare and may be overlooked as a cause of chronic tenosynovitis. This report presents a case of a young woman with tuberculosis tenosynovitis of the wrist, and highlights the clinical, imaging, histological, and laboratory features most commonly seen in this disease. PMID- 15297980 TI - Deriving haplotypes through recombination and gene conversion pathways. AB - Retracing the trajectories of past genetic events is crucial to understand the structure of the genome, both in individuals and across populations. A haplotype describes a string of polymorphic sites along a DNA segment. Haplotype diversity is due to mutations creating new variants, and to recombinations and gene conversions that mix and redistribute these variants among individual chromosomes in populations. A number of studies have revealed a relatively simple pattern of haplotype diversity in the human genome, dominated by a few common haplotypes representing founder ancestral ones. New haplotypes are usually rare and have a limited geographic distribution. We propose a method to derive a new haplotype from a set of putative ancestral haplotypes, once mutations in place, through minimal recombination and gene conversion pathways. We describe classes of pathways that represent the whole set of minimal pathways leading to a new haplotype. We show that obtaining this set of pathways can be represented as a problem of finding "secondary structures" of minimum energy. We present a polynomial algorithm solving this folding problem. PMID- 15297981 TI - Ancestral maximum likelihood of evolutionary trees is hard. AB - Maximum likelihood (ML) (Neyman, 1971) is an increasingly popular optimality criterion for selecting evolutionary trees. Finding optimal ML trees appears to be a very hard computational task--in particular, algorithms and heuristics for ML take longer to run than algorithms and heuristics for maximum parsimony (MP). However, while MP has been known to be NP-complete for over 20 years, no such hardness result has been obtained so far for ML. In this work we make a first step in this direction by proving that ancestral maximum likelihood (AML) is NP complete. The input to this problem is a set of aligned sequences of equal length and the goal is to find a tree and an assignment of ancestral sequences for all of that tree's internal vertices such that the likelihood of generating both the ancestral and contemporary sequences is maximized. Our NP-hardness proof follows that for MP given in (Day, Johnson and Sankoff, 1986) in that we use the same reduction from Vertex Cover; however, the proof of correctness for this reduction relative to AML is different and substantially more involved. PMID- 15297982 TI - Efficiently finding regulatory elements using correlation with gene expression. AB - We present an efficient algorithm for detecting putative regulatory elements in the upstream DNA sequences of genes, using gene expression information obtained from microarray experiments. Based on a generalized suffix tree, our algorithm looks for motif patterns whose appearance in the upstream region is most correlated with the expression levels of the genes. We are able to find the optimal pattern, in time linear in the total length of the upstream sequences. We implement and apply our algorithm to publicly available microarray gene expression data, and show that our method is able to discover biologically significant motifs, including various motifs which have been reported previously using the same data set. We further discuss applications for which the efficiency of the method is essential, as well as possible extensions to our algorithm. PMID- 15297983 TI - Binding matrix: a novel approach for binding site recognition. AB - Recognition of protein-DNA binding sites in genomic sequences is a crucial step for discovering biological functions of genomic sequences. Explosive growth in availability of sequence information has resulted in a demand for binding site detection methods with high specificity. The motivation of the work presented here is to address this demand by a systematic approach based on Maximum Likelihood Estimation. A general framework is developed in which a large class of binding site detection methods can be described in a uniform and consistent way. Protein-DNA binding is determined by binding energy, which is an approximately linear function within the space of sequence words. All matrix based binding word detectors can be regarded as different linear classifiers which attempt to estimate the linear separation implied by the binding energy function. The standard approaches of consensus sequences and profile matrices are described using this framework. A maximum likelihood approach for determining this linear separation leads to a novel matrix type, called the binding matrix. The binding matrix is the most specific matrix based classifier which is consistent with the input set of known binding words. It achieves significant improvements in specificity compared to other matrices. This is demonstrated using 95 sets of experimentally determined binding words provided by the TRANSFAC database. PMID- 15297984 TI - Characterization of the splice sites in GT-AG and GC-AG introns in higher eukaryotes using full-length cDNAs. AB - For the purpose of analyzing the relation between the splice sites and the order of introns, we conducted the following analysis for the GT-AG and GC-AG splice site groups. First, the pre-mRNAs of H. sapiens, M. musculus, D. melanogaster, A. thaliana and O. sativa were sampled by mapping the full-length cDNA to the genomes. Next, the consensus sequences at different regions of pre-mRNAs were analyzed in the five species. We also investigated the mononucleotide and dinucleotide frequencies in the extensive regions around the 5' splice sites (5'ss) and 3' splice sites (3'ss). As a result, differential frequencies of nucleotides at the first 5'ss in both the GT-AG and GC-AG splice site groups were observed in A. thaliana and O. sativa pre-mRNAs. The trend, which indicates that GC 5'ss possess strong consensus sequences, was observed not only in mammalian pre-mRNAs but also in the pre-mRNAs of D. melanogaster, A. thaliana and O. sativa. Furthermore, we examined the consensus sequences of the constitutive and alternative splice sites. It was suggested that in the case of the alternative GC AG introns, the tendency to have a weak consensus sequence at 5'ss is different between H. sapiens and M. musculus pre-mRNAs. PMID- 15297985 TI - Clustering of amino acids for protein secondary structure prediction. AB - Simple hidden Markov models are proposed for predicting secondary structure of a protein from its amino acid sequence. Since the length of protein conformation segments varies in a narrow range, we ignore the duration effect of length distribution, and focus on inclusion of short range correlations of residues and of conformation states in the models. Conformation-independent and -dependent amino acid coarse-graining schemes are designed for the models by means of proper mutual information. We compare models of different level of complexity, and establish a practical model with a high prediction accuracy. PMID- 15297986 TI - Prediction of protein secondary structure based on residue pairs. AB - The GOR program for predicting protein secondary structure is extended to include triple correlation. A score system for a residue pair to be at certain conformation state is derived from the conditional weight matrix describing amino acid frequencies at each position of a window flanking the pair under the condition for the pair to be at the fixed state. A program using this score system to predict protein secondary structure is established. After training the model with a learning set created from PDB_SELECT, the program is tested with two test sets. As a method using single sequence for predicting secondary structures, the approach achieves a high accuracy near 70%. PMID- 15297987 TI - Multivariate entropy distance method for prokaryotic gene identification. AB - A new simple method is found for efficient and accurate identification of coding sequences in prokaryotic genome. The method employs a Shannon description of artificial language for DNA sequences. It consists in translating a DNA sequence into a pseudo-amino acid sequence with 20 fundamental words according to the universal genetic code. With an entropy-density profile (EDP), the method maps a sequence of finite length to a vector and then analyzes its position in the 20 dimensional phase space depending on its nature. It is found that the ratio of the relative distance to an averaged coding and non-coding EDP over a small number (up to one) of open reading frames (ORFs) can serve as a good coding potential. An iterative algorithm is designed for finding a set of "root" sequences using this coding potential. A multivariate entropy distance (MED) algorithm is then proposed for the identification of prokaryotic genes; it has a feature to combine the use of a coding potential and an EDP-based sequence similarity analysis. The current version of MED is unsupervised, parameter-free and simple to implement. It is demonstrated to be able to detect 95-99% genes with 10-30% of additional genes when tested against the RefSeq database of NCBI and to detect 97.5-99.8% of confirmed genes with known functions. It is also shown to be able to find a set of (functionally known) genes that are missed by other well-known gene finding algorithms. All measurements show that the MED algorithm reaches a similar performance level as the algorithms like GeneMark and Glimmer for prokaryotic gene prediction. PMID- 15297988 TI - Techniques for optimization of queries on integrated biological resources. AB - Today, scientific data are inevitably digitized, stored in a wide variety of formats, and are accessible over the Internet. Scientific discovery increasingly involves accessing multiple heterogeneous data sources, integrating the results of complex queries, and applying further analysis and visualization applications in order to collect datasets of interest. Building a scientific integration platform to support these critical tasks requires accessing and manipulating data extracted from flat files or databases, documents retrieved from the Web, as well as data that are locally materialized in warehouses or generated by software. The lack of efficiency of existing approaches can significantly affect the process with lengthy delays while accessing critical resources or with the failure of the system to report any results. Some queries take so much time to be answered that their results are returned via email, making their integration with other results a tedious task. This paper presents several issues that need to be addressed to provide seamless and efficient integration of biomolecular data. Identified challenges include: capturing and representing various domain specific computational capabilities supported by a source including sequence or text search engines and traditional query processing; developing a methodology to acquire and represent semantic knowledge and metadata about source contents, overlap in source contents, and access costs; developing cost and semantics based decision support tools to select sources and capabilities, and to generate efficient query evaluation plans. PMID- 15297989 TI - Searching for transcription factor binding site clusters: how true are true positives? AB - The computational detection of functional transcription factor binding sites in genomic sequence is one of the challenges of the post-genomic era. Several groups have approached this problem from different directions and have demonstrated considerable success. The purpose of this communication, however, is to point out an imperfection in the way computational results are commonly reported that may lead to a distorted picture of the performance of existing algorithms. PMID- 15297990 TI - Evidence of airborne transmission of SARS. PMID- 15297991 TI - Evidence of airborne transmission of SARS. PMID- 15297992 TI - Registries and informed consent. PMID- 15297993 TI - Registries and informed consent. PMID- 15297994 TI - [Nosebleed and its treatment]. PMID- 15297995 TI - [[1]Benzothieno[3,2-b]pyridin-4-yl-amine--synthesis and investigation of activity against malaria]. AB - The ethyl 4-chlorobenzothieno[3,2-b]pyridine-3-carboxylate (2) reacted with the hydrochlorides of the mono- and bis-phenol Mannich bases 6 to yield the amodiaquine and pyronaridine analogues 9. The chloroquine analogue 10 was formed by melting 2 with the novaldiamine base (7) in phenol. The stability of the 4 aminophenols 9 was investigated by anodic oxidation using the rotating platinum electrode by means of difference pulse voltammetry. The half wave potentials were measured giving E(1/2) approximately 1.05 V. Compound 9g displayed the highest activity against the growth of the malaria parasite Plasmodium falciparum. Testing against the chloroquine sensitive 3D7 and the chloroquine resistant Dd2 strain resulted in IC50 values of 150 nM and 210 nM, respectively. Surprisingly, the 3-carbinol 4 and the 3-chloromethyl derivative 5, synthesized from the 3 carboxylic acid ester 2, reacted with the phenol Mannich base 6a and the novaldiamine base (7), respectively, to yield the 4-pyridone 8. PMID- 15297997 TI - Shaping the intimate: influences on the experience of everyday nerves. AB - Before 1980, most people experiencing common nervous problems and who sought medical help complained of anxiety and were treated for anxiety. Similar experiences increasingly led to complaints of or treatment for panic attacks in the late 1980s and early 1990s, and to complaints of or treatment for mood disorders by the mid-1990s. Today, such patients seem once again increasingly likely to complain of and be treated for anxiety. This paper reviews a series of mechanisms whereby company marketing can both transform the perceptions of physicians and shape the experiences of those seeking treatment and the self understanding of those not in treatment. These include the standard ploys of company sales departments to increase demand for products, including celebrity endorsements, the sponsoring of educational events and a host of reminders. The portfolio of marketing manoeuvres has grown, though, by translating educational events and celebrity events into the arena of scientific research: clinical trials have increasing become part of the marketing of disorders and their treatments; ghost-written scientific papers are authored by celebrity researchers. The portfolio of marketing manoeuvres has also grown to encompass new ways of creating fashion through medical activism, by setting up patient groups and disease awareness campaigns. The result is a transformation and growth in disorders tailor-made to fit ever more visible drugs. PMID- 15297996 TI - Modeling of an annular photocatalytic reactor for water purification: oxidation of pesticides. AB - Photocatalytic oxidation (PCO) over titanium dioxide (TiO2) is a "green" sustainable process for the treatment and purification of water and wastewater. However, the application of PCO for wastewater treatment on an industrial scale is currently hindered by a lack of simple mathematical models that can be readily applied to reactor design. Current models are either too simplistic or too rigorous to be useful in photocatalytic reactor design, scale-up, and optimization. In this paper a simple mathematical model is presented for slurry, annular, photocatalytic reactors that still retains the essential elements of a rigorous approach while providing simple solutions. The model extends the applicability of the thin-film model of photocatalytic reactors previously presented to include the case of geometrically thick photoreactors (i.e., those reactors in which the thickness of the annular zone is significant as compared to the outer radius of the reactor). The model uses a novel six-flux absorption scattering model to represent the radiation field in the reaction space, which assumes that scattered photons follow the route of the six directions of the Cartesian coordinates. The model was successfully validated with experimental results from the photocatalytic oxidation of the pesticide isoproturon in an experimental reactor. The mathematical model presented may be used as a tool for the design, scale-up, and optimization of annular photocatalytic reactors for water treatment and purification. PMID- 15297998 TI - The anxieties of globalization: antidepressant sales and economic crisis in Argentina. AB - This paper describes the role of market research firms in shaping the actions of key players in the pharmaceutical arena. It focuses on strategies for marketing novel antidepressants (selective serotonin reuptake inhibitors, SSRIs) to doctors in Buenos Aires during the Argentine financial crisis of 2001, posing the question of whether increased antidepressant sales were due to the social situation or to promotional practices. This case demonstrates how 'pharmaceutical relations' - interactions between doctors and pharmaceutical companies - are structured by a gift economy whose effects are monitored through the sales numbers produced by database firms. It suggests that the use of these numbers takes on special importance given the distinctiveness of both the Argentine context and the antidepressant market. More generally, the case points to the interpretive flexibility of psychotropic medication. In the Argentine setting, doctors' prescription of SSRIs was dependent neither on a diagnosis of depression nor on a biological understanding of mental disorder. These drugs found a different means of entering the professionally mediated marketplace: doctors understood and used SSRIs as a treatment not for a lack of serotonin in the brain, but for the suffering caused by the social situation - the sense of insecurity and vulnerability that the economic and political crisis had wrought. PMID- 15297999 TI - Prepubertal midface growth in unilateral cleft lip and palate following alveolar molding and gingivoperiosteoplasty. AB - OBJECTIVES: To examine the long-term effect of nasoalveolar molding and gingivoperiosteoplasty (modified Millard type) on midface growth at prepuberty. PROCEDURES: In this retrospective study, 20 consecutive patients with a history of complete unilateral cleft lip and palate were evaluated. Ten patients had nasoalveolar molding and gingivoperiosteoplasty performed at lip closure; 10 control patients had nasoalveolar molding but no gingivoperiosteoplasty because of late start in treatment or poor compliance. A single surgeon (C.B.C.) performed all surgical procedures. Standardized lateral cephalometric radiographs were evaluated at two time periods: T1 at pre-bone-grafting age and T2 at prepuberty age. Superimposition and cephalometric analysis were undertaken to investigate the two groups. Two cephalometric reference planes, sella-nasion and basion-nasion, were used to assess the vertical and sagittal relations of the midface (ANS-PNS). The reference landmarks were procrustes fitted. The mean location and variance of ANS and PNS landmarks were computed. All results were analyzed by permutation test. RESULTS: No significant difference in mean location or variance of ANS-PNS in both vertical and sagittal planes at both T1 and T2 periods were found between the two groups (p > .05). CONCLUSIONS: The results suggested that midface growth in sagittal or vertical planes (up to the age of 9 to 13 years) were not affected by presurgical alveolar molding and gingivoperiosteoplasty (Millard type). PMID- 15298000 TI - A new approach for accessing retroperitoneal space using a 5-mm [corrected] visual access cannula. AB - Retroperitoneal lymph node evaluation is a very important step in planning cancer therapy. Traditional staging laparotomy is too invasive and causes considerable morbidity. However, laparoscopic access of the retroperitoneal space offers the onco-endoscopist a unique, noninvasive, and less dangerous method for assessing and managing women with gynecologic malignancies. The retroperitoneal approach is noninvasive and less dangerous way of accessing the paraaortic space. A new minimally invasive approach of accessing the retroperitoneal space using a 5-mm visual access cannula is introduced. A complete paraaortic and pelvic lymphadenectomy was possible for 60 patients with varied gynecologic malignancies using our retroperitoneal laparoscopic approach. Less blood loss and rapid recovery was observed, and no bowel complication was encountered. The use of the 5-mm visual access cannula offers a less invasive surgical procedure and provides cosmetic advantages. PMID- 15298001 TI - Attention to 'details': etiquette and the pharmaceutical salesman in postwar American. AB - This paper provides a sketch of the emerging role of the pharmaceutical salesman, or 'detail man', in the growth years of the American post-World War II pharmaceutical industry. Using training manuals, trade literature, in-house company newsletters, memoirs, and a variety of other published sources, the paper follows the delicate tactics employed by salesmen and their managers in their attempts to recast drug salesmanship as a 'professional service' fulfilling vital functions within medical education. As they worked to legitimate their presence in the nation's hospitals and clinics, particular emphasis was given to precise management of the etiquette of doctor-salesman interaction. Ultimately, the techniques employed by mid-century salesmen and their managers were to prove successful in generating a widespread acceptance of the industry representative within the clinical spaces of hospital and clinic. Indeed, many of the practices of market research and market strategy employed across the pharmaceutical industry today have their origins in the practices of the individual detail man. Exploration of the postwar pharmaceutical salesman as an overlooked historical 'type' provides significant insights into the intersection of medicine and the consumer marketplace during the later 20th century. PMID- 15298003 TI - The ethics of medical misadventure. PMID- 15298002 TI - Three-dimensional morphology of the palate in subjects with unilateral complete cleft lip and palate at the stage of permanent dentition. AB - OBJECTIVE: Three-dimensional analysis of palate size and shape in patients with complete unilateral cleft lip and palate (UCLP) at the stage of permanent dentition. SUBJECTS: Thirty randomly selected dental casts of boys approximately 15 years old with complete UCLP and 28 dental casts of normal boys of the same age. INTERVENTIONS: All patients underwent lip repair according to Tennison with primary periosteoplasty (mean age 8.5 months) and palate repair by pushback and pharyngeal flap surgery (mean age 4.9 years). MAIN OUTCOME MEASURES: Data on the palate height in 210 defined locations. RESULTS: The palate in patients with UCLP was narrower throughout its whole extent, more anteriorly than posteriorly. From the canines posteriorly, it was also lower, and the difference as compared with controls increased in a posterior direction up to the level of second premolars (up to 30%) and then slightly diminished (to 21% between the first molars). The reduction of area of transverse sections reached 45% between premolars and 39% between first molars. The palate in the anterior portion was highest on the cleft side and in a posterior direction the maximum height of the palate shifted toward the midline and even beyond that line toward the noncleft side. Palatal height did not depend on dentoalveolar arch width. CONCLUSION: The smaller width and height of the palate confirm the substantially reduced space for the tongue in patients with UCLP. The reduction is only slightly larger than in previously examined patients with isolated cleft palate. Palatal vault is asymmetrical, highest anteriorly on the cleft side and posteriorly on the noncleft side. PMID- 15298004 TI - The code and the guide--practical instruments for practical people: a reply to Cannold. PMID- 15298005 TI - Confidential Health Care for Adolescents: position paper for the society for adolescent medicine. PMID- 15298006 TI - Oxidative stress in neurodegeneration: cause or consequence? AB - Oxidative stress has long been linked to the neuronal cell death that is associated with certain neurodegenerative conditions. Whether it is a primary cause or merely a downstream consequence of the neurodegenerative process is still an open question, however. The advent of a growing number of in vitro and in vivo models that emulate human disease pathology is aiding scientists in deciphering just where oxidative stress intersects with other cellular events in the emerging roadmap leading to neurodegeneration. Here I review the evidence for oxidative stress in neurodegeneration and how this relates to other cellular events. PMID- 15298007 TI - Advances in the early detection of Alzheimer's disease. AB - The combination of an aging population and the promise, possibly in the near future, of disease-modifying therapies have made the characterization of the early stages of Alzheimer's disease (AD) a topic of major research interest. In this article we review recent progress in our understanding of the evolution of early AD with particular reference to the symptomatic pre-dementia stage designated 'mild cognitive impairment', emphasizing work on the early cognitive profile and associated neuroimaging studies. PMID- 15298008 TI - Back to the future: the 'old-fashioned' way to new medications for neurodegeneration. AB - Despite the increasing prevalence of Alzheimer's disease, Parkinson's disease and less common neurodegenerative diseases-and despite the large amount of primary research that has been carried out into the causes and pathogenic features of these conditions-progress toward effective treatments has been remarkably slow. Why is this, and what can be done to accelerate it? There are a number of obstacles to effective drug discovery for neurodegeneration, but by considering these problems it is possible to identify lessons for the future. PMID- 15298009 TI - Pharmaceutical formularies: the right formula for cost and utilization? PMID- 15298010 TI - Limited-benefit policies: public and private-sector experiences. PMID- 15298011 TI - ERISA update: the Supreme Court Texas decision and other recent developments. AB - This issue brief is part of a continuing series of policy papers published by The Robert Wood Johnson Foundation's State Coverage Initiatives program, housed at AcademyHealth, and the National Academy for State Health Policy on the state health policy implications of ERISA's preemption clause.1 The purpose of the brief is to explore the U.S. Supreme Court's June 2004 decision that ERISA preempts the Texas HMO liability law and its effects on other state health plan liability laws. The brief also examines implications of ERISA preemption for state health insurance regulation, "pay or play" health coverage laws, and premium assistance programs. PMID- 15298012 TI - State regulation of pharmaceutical clinical trials. PMID- 15298014 TI - The errors of error testing: potential liability issues for medication error testing of pharmaceutical trademarks under U.S. law. PMID- 15298013 TI - A brief history of 180-day exclusivity under the Hatch-Waxman Amendments to the Federal Food, Drug, and Cosmetic Act. AB - This article summarizes the history of the 180-day exclusivity provision on the Hatch-Waxman Amendments to the Federal Food, Drug, and Cosmetic Act (FDCA). Part II presents the statutory language, as amended in the Medicare Prescription Drug, Improvement, and Modernization Act of 2003 (MMA), and summarizes the law that applies to new abbreviated new drug applications (ANDAs) (those filed after December 8, 2003, provided there was no paragraph IV certification to the listed drug prior to December 8), as well as the law that applies to all other ("old") ANDAs. Part III describes the legislative history of the original 1984 provision and traces its judicial and administrative history through the present. Part IV describes the history of the 2003 amendments and describes the key changes made in 2003. While Congress addressed in 2003 a number of the interpretive issues that had arisen since 1984, the new law is intricate and undoubtedly will give rise to new interpretive questions in the months and years ahead. PMID- 15298015 TI - The Food and Drug Administration and patent law at a crossroads: the listing of polymorph patents as a barrier to generic drug entry. PMID- 15298016 TI - Public health, hypocrisy, and brown-skinned people. PMID- 15298017 TI - Disease outbreaks in Sudan worsen ongoing crisis. PMID- 15298018 TI - Global fund switches to artemisinin. PMID- 15298019 TI - WHO regroups efforts against polio resurgence in Africa. PMID- 15298020 TI - India leads meningitis A vaccine development. PMID- 15298021 TI - India expands mass chemotherapy to eradicate filariasis. PMID- 15298022 TI - Mapping the evolution of influenza A. PMID- 15298023 TI - Malaria vaccination could drive parasite evolution. PMID- 15298024 TI - SARS virus in tears? PMID- 15298025 TI - ESBLs: the next challenge in infection control. PMID- 15298026 TI - Watchdog slams UK government's AIDS policy. PMID- 15298028 TI - AIDS in Cambodia. PMID- 15298029 TI - Infectious titer assay for adeno-associated virus vectors with sensitivity sufficient to detect single infectious events. AB - A highly sensitive assay for determination of infectious titers of recombinant adeno-associated virus (AAV) by limiting dilution analysis is described. This assay is capable of detecting single infectious events and can therefore provide an absolute rather than relative measure of infectivity. The assay utilizes a HeLa-derived AAV2 Rep/Cap-expressing cell line, D7-4, grown in 96-well plates and infected with replicate 10-fold serial dilutions of AAV2 vectors in the presence of adenovirus type 5. Forty-eight hours after infection, vector genome replication is determined by quantitative PCR (Q-PCR). A linear relationship between vector genome input and replicated copy number (slope = 2670 copies per vector genome) was determined, enabling detection of one infectious event per well by Q-PCR. The observed binomial distribution of the end-point data confirmed that single infectious events could be detected, and allowed calculation of infectious titers by the Karber method. Analysis of an AAV2 reference vector, AAV hFIX16, in 21 independent determinations gave an average ratio of AAV vector genomes (VG) to infectious units (IU) of 8.3 +/- 4.2 VG/IU, a value close to the theoretical limit. No significant differences in vector particle-to-infectious unit ratios were observed between vectors purified by column chromatography (9.3 +/- 5.0 VG/IU, n = 7) and cesium chloride gradient ultracentrifugation (6.4 +/- 3.2 VG/IU, n = 7). PMID- 15298030 TI - Prepreparation of succinylcholine. PMID- 15298031 TI - Prepreparation of succinylcholine. PMID- 15298032 TI - Two new healthcare standards to aid in data capture. PMID- 15298033 TI - Images in cardiology: "Bleeding heart": cardiac complications in thrombotic thrombocytopenic purpura. M. A. Sheikh: Clin Cardiol 2004:27:70. PMID- 15298034 TI - Refractory chronic stable angina--now what? PMID- 15298035 TI - Spontaneous coronary artery dissection. AB - Spontaneous coronary artery dissection (SCAD) is an unusual cause of acute myocardial ischemia with complex pathophysiology. This paper reviews the major diagnostic and therapeutic issues of this rare but important disease. The diagnosis of SCAD should be strongly considered in any patient who presents with symptoms suggestive of acute myocardial ischemia, particularly in young subjects without traditional risk factors for coronary artery disease (especially in young women during the peripartum period or in association with oral contraceptive use). Urgent coronary angiography is indicated to establish the diagnosis and to determine the appropriate therapeutic approach. The decision to pursue medical management, percutaneous coronary intervention, or surgical revascularization is based primarily on the clinical presentation, extent of dissection, and amount of ischemic myocardium at risk. PMID- 15298036 TI - Considerations in combination therapy: fibrinolytics plus glycoprotein IIb/IIIa receptor inhibitors in acute myocardial infarction. AB - The combined use of a fibrinolytic and a platelet glycoprotein (GP) IIb/IIIa receptor inhibitor to target the fibrin and platelet components of occlusive thrombi offers the potential for more rapid and complete reperfusion in patients with acute myocardial infarction (MI), although there have been concerns about the safety of this combination therapy. Data from the recent GUSTO-V and the ASSENT-3 trials support the use of this regimen in that the 30-day death or nonfatal reinfarction rate (7 days) in GUSTO-V and death or in-hospital reinfarction or in-hospital refractory ischemia rate in ASSENT-3 were reduced (p = 0.001 and p = 0.0001, respectively). The need for revascularization in both these trials was also reduced significantly. There was no increased risk of intracranial hemorrhage or stroke with the combination therapy, but an increased rate of nonintracranial severe or major bleeding was observed. At present, patients aged > 75 years should not receive combination therapy. Further studies in subgroup patient populations are warranted. PMID- 15298037 TI - Vascular age: integrating carotid intima-media thickness measurements with global coronary risk assessment. AB - BACKGROUND: An imaging test that quantifies atherosclerotic burden and that can be integrated with existing risk stratification paradigms would be a very useful clinical tool. HYPOTHESIS: Measurement of carotid intima-media thickness (CIMT) is feasible in a clinical setting. Such measurements can be integrated into coronary risk assessment models. METHODS: Carotid intima-media thickness was measured by B-mode ultrasound in 82 consecutive patients without manifest atherosclerotic vascular disease. The values were used to determine "vascular age" (VA) based on nomograms from the Atherosclerosis Risk in Communities study. Vascular age was substituted for chronological age and standard and vascular age adjusted 10-year coronary heart disease (CHD) risk estimates were compared. RESULTS: The mean chronological age was 55.8 +/- 9.0 years. The mean VA using CIMT was 65.5 +/- 18.9 years (p < 0.001). The Framingham 10-year hard CHD risk estimate was 6.5 +/- 4.9%. Substituting CIMT-derived VA for chronological age increased the 10-year CHD risk estimate to 8.0 +/- 6.8% (p < 0.001). Of 14 subjects initially at intermediate risk, 5 (35.7%) were reclassified as higher risk and 2 (14.3%) were reclassified as lower risk. Significant predictors of reclassification were tobacco use, high-density lipoprotein cholesterol, systolic blood pressure, and low-density lipoprotein cholesterol. CONCLUSIONS: Measurement of CIMT, a noninvasive estimate of current atherosclerotic burden, is feasible in a clinical setting and can be integrated into CHD risk assessment models. Determining VA using CIMT values may help individualize the age component of population-based CHD risk estimates. This strategy should be tested in a large trial with hard clinical endpoints. PMID- 15298038 TI - Bail-out stenting for left main coronary artery dissection during catheter-based procedure: acute and long-term results. AB - BACKGROUND: The optimal treatment of patients with left main coronary artery (LMCA) dissection during catheter-based procedure remains uncertain. HYPOTHESIS: In cases with significant LMCA dissection occurring during catheter-based procedure, prompt stent implantation may be safe and associated with favorable clinical outcome. METHODS: We evaluated the acute and long-term results of bail out stenting for LMCA dissection occurring during a catheter-based procedure in 10 patients. RESULTS: Initially, there was no significant stenosis of LMCA segments in these patients. Catheter-induced dissection occurred in eight patients (during diagnostic angiography in three patients and during guiding catheter manipulation in five patients). Two patients suffered dissection in the setting of stent deployment in other vessels. Therefore, bail-out stenting for LMCA dissection was performed in a total of 10 patients. In four patients, hypotension developed and an intra-aortic balloon pump was placed during the procedure. Stents were successfully deployed in all patients; there was no in hospital mortality. Six-month angiographic follow-up was performed in eight patients. No angiographic restenosis (diameter stenosis > or = 50%) was observed in any patient at follow-up study. During a mean follow-up of 31 +/- 25 months after hospital discharge, there was no major adverse cardiac event (death, myocardial infarction, and target lesion revascularization). CONCLUSIONS: Bail out LMCA stenting is technically feasible and showed good acute and long-term results in a small series of patients. PMID- 15298039 TI - Atrial electrophysiologic abnormalities in patients with Wolff-Parkinson-White syndrome but without paroxysmal atrial fibrillation. AB - BACKGROUND: Paroxysmal atrial fibrillation (PAF) frequently occurs in patients with Wolff-Parkinson-White (WPW) syndrome. HYPOTHESIS: The purpose of this study was to analyze the atrial electrophysiologic abnormalities and vulnerability to develop atrial fibrillation (AF) in patients with WPW syndrome but with no previous history of PAF. METHODS: We investigated atrial electrophysiologic abnormalities and vulnerability to AF in patients with WPW syndrome but without PAF. An electrophysiologic study was performed in 28 patients with WPW syndrome, 23 with atrioventricular nodal reentrant tachycardia (AVNRT) and 25 with other arrhythmias (control), all of whom had no history of PAF. The following atrial excitability parameters were assessed: effective refractory period (ERP), spontaneous or paced (A1) and extrastimulated (A2) atrial electrogram widths, percent maximum atrial fragmentation (%MAF; A2/A1 x 100), wavelength index (WLI; ERP/A2), and inducibility of AF. RESULTS: The ERP tended to be shorter in patients with WPW syndrome and in those with AVNRT than in the control group. The %MAF increased (154 +/- 33 vs. 137 +/- 23%, p < 0.05) and WLI decreased (2.7 +/- 0.8 vs. 3.4 +/- 1.0, p < 0.05) significantly in patients with WPW syndrome compared with the control group; however, these parameters in patients with AVNRT showed intermediate values. Atrial fibrillation was more inducible in patients with WPW syndrome (4/28 [14.3%]) than in those with AVNRT (4.3% [1/23]) and the control group (0/25 [0%]). With respect to patients with WPW syndrome and with and without inducible AF, the %MAF increased (195 +/- 23 vs. 148 +/- 30%, p < 0.01) and the WLI decreased (2.2 +/- 0.3 vs. 2.9 +/- 0.9, p < 0.05) in subjects with inducible AF. CONCLUSIONS: Atrial electrophysiologic abnormalities, especially atrial conduction delays, are more prominent in patients with WPW syndrome, even if they had no previous history of PAF. These abnormalities may play an important role in determining the vulnerability to AF. PMID- 15298040 TI - Hemodynamic and tissue oxygenation responses to exercise and beta-adrenergic blockade in patients with hyperthyroidism. AB - BACKGROUND: Exercise-induced dyspnea is a frequent feature in patients with hyperthyroidism. HYPOTHESIS: Data from clinical studies to elucidate the origin of this symptom are lacking. In the current study, we examined the hemodynamic and oxygenation responses to exercise and beta-adrenergic blockade in patients with hyperthyroidism and their relationship with dyspnea. METHODS: Hemodynamic studies were performed under resting conditions and after isotonic exercise in 15 patients with hyperthyroidism and 11 control subjects. Exercise was applied using a bicycle ergometer, with progressive loads. In the hyperthyroid group, measurements were repeated at rest and during supine exercise after administering 15 mg of intravenous metoprolol. RESULTS: End-diastolic pulmonary artery pressure and cardiac index were higher in the hyperthyroid group than in controls (18.6 +/ 5.3 vs. 11.2 +/- 4.9 mmHg; p = 0.02, and 6.0 +/- 1.7 vs. 2.8 +/- 0.5 l/min/m2; p = 0.0001, respectively). After exercise, there was an increase in end-diastolic pulmonary artery pressure in the hyperthyroid group (18.6 +/- 5.3 to 25.5 +/- 9.9 mmHg; p = 0.02), revealing impaired cardiocirculatory reserve. Pulmonary arteriolar resistance increased significantly in parallel with end-diastolic pulmonary artery pressure after drug administration, suggesting an inadequate cardiovascular response after beta blockade in patients with hyperthyroidism. CONCLUSION: We observed that functional left ventricular reserve is impaired in patients with hyperthyroidism, suggesting an explanation for the frequent symptom of dyspnea and impaired exercise tolerance. Moreover, we also suggest that beta adrenergic blockade may adversely affect cardiovascular function in patients with hyperthyroidism. PMID- 15298041 TI - Elevation of the soluble thrombomodulin levels is associated with inflammation after percutaneous coronary interventions. AB - BACKGROUND: Thrombomodulin (TM) is an endothelial cell surface thrombin-binding protein with anticoagulation ability by thrombin-mediated activation of protein C. An increase of plasma soluble TM level is reported to be associated with severity and worse outcome of coronary artery disease. HYPOTHESIS: This prospective study investigated the relation of the elevated levels of plasma soluble TM and inflammatory and myonecrotic markers in patients undergoing percutaneous coronary intervention (PCI). METHODS: Plasma levels of soluble TM, C reactive protein (CRP), and creatine kinase and its MB isoenzyme were measured before and after PCI in 100 patients undergoing PCIs. RESULTS: Peak TM levels after PCIs were significantly higher than baseline (3.39 +/- 1.63 vs. 2.90 +/- 1.57 ng/ml, p < 0.001). The peak TM levels after PCIs correlated significantly with the peak CRP and MB levels, and the maximal inflation duration (r = 0.423, p < 0.001; r = 0.212, p = 0.034; r = 0.307, p= 0.002, respectively). CONCLUSIONS: Soluble TM levels increase significantly after PCI. The elevation of the soluble TM after PCI shows better correlation with inflammation than myocardial injury, indicating an endothelial origin. Measurement of soluble TM could be useful and calls for further studies on the prognostic effects of this marker in this clinical condition. PMID- 15298042 TI - Images in cardiology: Pericardial yolk sac tumor presenting as cardiac tamponade in a 21-month-old child. PMID- 15298043 TI - Images in cardiology: Percutaneous coronary angioplasty of a single coronary artery originating above the left sinus of Valsalva. PMID- 15298044 TI - Transthoracic echocardiography for precardioversion screening during atrial flutter/fibrillation in young patients. AB - BACKGROUND: Transthoracic echocardiography (TTE) is reliable for detection of thrombi in the left ventricle and right atrium, but not in the left atrial appendage. Therefore, transesophageal echocardiography (TEE) is routinely performed in adults prior to electric cardioversion for atrial flutter/fibrillation (AFF). Whether young survivors of congenital heart disease repair with AFF need routine TEE prior to electric cardioversion is unknown. HYPOTHESIS: Electric cardioversion for AFF is safe in survivors of congenital heart disease repair/palliation if an intracardiac thrombus is not suspected on TTE imaging. METHODS: This study reports the outcome of patients in a pediatric tertiary care cardiac unit where electric cardioversion was performed if no intracardiac thrombus was suspected on TTE. We performed a retrospective chart review of all patients treated with electric cardioversion for AFF at Children's Hospital of Michigan during 1997-2002. RESULTS: Of 35 patients who presented with 110 episodes of AFF requiring electric cardioversion during the study duration, 32 (age 3 months-49 years, median age 20.5 years, 104 AFF episodes) had previously undergone palliative surgery or repair of their congenital heart disease. Of these 32 patients, 18 were survivors of a Fontan palliation (for a single-ventricle variant) and the remaining 14 were survivors of other defects and repairs (septal defects, valve replacements, and tetralogy of Fallot). During 81% of the episodes, patients were receiving aspirin, warfarin, or heparin for anticoagulation at presentation. Transthoracic echocardiography was performed in 74 AFF episodes; of these, 10 TTE studies were suspicious for atrial thrombi. Transesophageal echocardiography confirmed the presence of a thrombus in 3 of these 10 patients. These patients received warfarin for 2 weeks and then underwent electric cardioversion. No thromboembolic events occurred immediately after or on follow-up in any patient. CONCLUSIONS: These findings suggest that TTE may be an effective imaging tool for precardioversion screening in young patients with AFF. PMID- 15298046 TI - Endothelial dysfunction in patients with exaggerated blood pressure response during treadmill test. AB - BACKGROUND: The diagnostic and prognostic importance of exaggerated blood pressure response to exercise is controversial. Endothelial dysfunction has been demonstrated in patients with atherosclerosis and risk factors for coronary artery disease, but there is a paucity of information on patients with exercise induced hypertension. HYPOTHESIS: We designed the study to evaluate endothelial function in patients with exaggerated blood pressure response during exercise. METHODS: Exercise-induced hypertension was defined as systolic blood pressure > or = 210 mmHg in men and > or = 190 mmHg in women during the treadmill test. Using a high-resolution ultrasound technique, endothelial function of the brachial artery in patients with exercise-induced hypertension (n = 25) and control subjects (n = 25) was investigated. RESULTS: Endothelium-dependent vasodilation was impaired in patients with exercise-induced hypertension compared with controls (7.77 +/- 5.14 vs. 2.81 +/- 2.29%, p < 0.05). On univariate analysis, the extent of vasodilation correlated negatively with age (r = -0.43, p < 0.05) and delta systolic blood pressure (r = -0.39, p < 0.05). Even after adjustment for factors known to affect endothelial function, endothelium dependent vasodilation was decreased in patients with exercise-induced hypertension (beta = 5.375, p = 0.02). CONCLUSION: Patients with exercise-induced hypertension have impaired endothelium-dependent vasodilation. This study also supports the concept that endothelial dysfunction may play an important role in exercise-induced hypertension. PMID- 15298045 TI - Relation of circulating interleukin-6 to left ventricular remodeling in patients with reperfused anterior myocardial infarction. AB - BACKGROUND: During the remodeling process after myocardial infarction (MI), the expression of proinflammatory cytokines is enhanced in the myocardium. However, only a few clinical studies have been conducted on cytokine involvement in left ventricular (LV) remodeling after MI. HYPOTHESIS: Circulating proinflammatory cytokines may be involved in LV remodeling in patients with reperfused MI. METHODS: We studied 25 patients with acute anterior MI who had undergone coronary reperfusion therapy, and 10 normal control subjects with no cardiac disease. In all patients, LV ejection fraction, end-diastolic volume index (EDVI), and end systolic volume index (ESVI) were determined using left ventriculography at the acute phase and 6 months after onset. The delta EDVI and delta ESVI were calculated as the value of LV volume reduction, suggesting LV reverse remodeling. Serum levels of interleukin (IL)-6 and tumor necrosis factor (TNF)-alpha were measured using enzyme-linked immunosorbent assay. RESULTS: Serum levels of IL-6 and TNF-alpha at the acute phase were significantly higher in patients with MI than in control subjects (both p < 0.05). The IL-6 levels correlated well negatively with delta EDVI (r = 0.779, p = 0.039), whereas no correlation was found for TNF-alpha. According to multivariate analysis, IL-6 at the acute phase was a significant independent predictor for LV remodeling after reperfused MI (p = 0.007). CONCLUSIONS: Circulating IL-6 levels correlated closely with LV geometric changes during the remodeling process in patients with reperfused MI. Our study addresses the usefulness of another marker for LV remodeling after MI. PMID- 15298047 TI - Association between paraoxonase-1 activity and lipid peroxidation indicator levels in people living in the Antalya region with angiographically documented coronary artery disease. AB - BACKGROUND: Paraoxonase-1 (PON1) is a high-density lipoprotein (HDL)-associated enzyme capable of hydrolyzing lipid peroxides. Thus, PON1 plays a preventing role in atherosclerosis by protecting against lipid peroxidation. HYPOTHESIS: The incidence of coronary artery disease (CAD) is high in the Turkish population, and many risk factors have been studied as determinants of CAD. In Turkish people living in the Antalya region, we aimed to determine serum PON1 activity and its relation to lipoproteins and lipid peroxidation markers. METHODS: We measured the activity of serum PON1 together with concentrations of a variety of lipid constituents--total cholesterol (TC), low-density lipoprotein cholesterol (LDL C), very low-density lipoprotein cholesterol (VLDL-C), HDL cholesterol (HDL-C), triglycerides (TG), apolipoprotein (apo) A-I, apoB, and lipid peroxidation indicators (conjugated diene [CD] and thiobarbituric acid-reactive substances [TBARS])--in 108 patients with CAD and 64 healthy subjects (controls). RESULTS: We found that the PON1 activity was significantly reduced in patients with CAD (222.37 +/- 11.31 IU/l) compared with controls (331.75 +/- 20.98 IU/l). These patients had significantly lower HDL-C, PON1/HDL-C, apoA-I, PON1/ApoA-I, and ApoA I/ApoB, and higher LDL-C, TC/HDL-C, LDL-C/HDL-C, apoB, CD and TBARS than did controls. Total cholesterol and apoA-I concentrations were significantly higher in women than in men in both groups. After multiple logistic regression analysis, TBARS (odds ratio [OR] 568.87; p = 0.000), age (OR 1.10; p = 0.000), gender (OR 4.58; p = 0.008), apoA-I/apoB (OR 0.046; p = 0.003), and PON1/apoA-I (OR 0.58; p = 0.007) were independently indicative of the presence of CAD. CONCLUSIONS: This is the first report of decreased serum PON1 activity and increased lipid peroxidation indicators (CD and TBARS) of patients with CAD living in Antalya, Turkey. Our results indicate that TBARS levels, age, gender, apoA-1/ApoB, and PON1/apoA-I ratios are important markers of CAD. PMID- 15298048 TI - Mario R. Garcia-Palmieri. PMID- 15298049 TI - William Harvey Trophy and international cardiology. PMID- 15298050 TI - [Proposals of the working group "Antibiotic resistance" for the configuration of microtitre plates to be used in routine antimicrobial susceptibility testing of bacterial pathogens from infections of large food-producing animals and mastitis cases]. AB - Two layouts for microtitre plates, which should serve for in-vitro susceptibility testing in routine diagnostics, have been set up by the working group "Antibiotic resistance" of the German Society for Veterinary Medicine. One of these layouts was designed for the testing of bacteria from cases of mastitis and the other for bacteria from infections in large food-producing animals. The choice of the antimicrobial agents and their concentrations to be included in these layouts were based on (1) the bacteria frequently associated with the respective diseases/animals, (2) the antimicrobial agents licensed for therapeutic use in these diseases/animals, (3) the currently available breakpoints, and (4) cross resistances between the antimicrobial agerts so far known to occur in the respective bacteria. PMID- 15298051 TI - [The first answer to viral infections: type I interferon]. AB - The interferon system is part of the innate immune system in vertebrates. It represents the first line of host defence against viral infections. Virus entry triggers intracellular signalling pathways which lead to the secretion of soluble factors such as interferons and other cytokines. Interferons signal to neighbouring cells that a viral infection has occurred and induce an "antiviral state" resulting in inhibition of virus replication. The first recombinant interferons were produced in the 1980ies and were considered to be a major breakthrough. At present, interferons are routinely used in the therapy of certain viral and autoimmune diseases as well as for neoplastic disorders in man. In 2001 the first interferon preparation for veterinary use was licensed in the European Union. This review summarises the molecular mechanisms of the interferon system and the viral counteractions. The current type I interferon therapies in humans are described and an overview of recent clinical studies in veterinary medicine, including cat, dog, horse, cow, sheep, pig, and poultry, is given. We review the potential application of interferons and arguments in favor or against its therapeutic use in veterinary medicine. PMID- 15298052 TI - Response of Streptococcus suis to iron-restricted growth conditions at high and low oxygen availability. AB - Streptococcus suis (S. suis) is an important pathogen in pigs and has to overcome strict iron limitations in its host environment. Here, we studied iron-restricted growth of a highly virulent S. suis strain in vitro at aerobic and CO2-enriched growth conditions. At both conditions, depleting of iron in the culture medium with nitrilotriacetic acid (NTA) resulted in decreased growth rates and down regulation of several proteins. Sensitivity to NTA was significantly higher at aerobic versus CO2-enriched conditions. Growth could not be restored by addition of host iron sources such as ferritin, hemin, hemoglobin, lactoferrin or transferrin. Accordingly, S. suis was not able to produce detectable amounts of siderophores. On the other hand, growth at iron-restricted conditions was fully restored by addition of Mn2+ (at aerobic and CO2-enriched conditions) or Mg2+ (only at CO2-enriched conditions). In conclusion our results suggest that, unlike many other bacteria, S. suis adapts to iron restricted conditions by a change in its metabolism in order to replace Fe2+ by Mn2+ or Mg2+ rather than by expressing specific iron uptake systems. PMID- 15298053 TI - Prevalence of Haemophilus parasuis serotypes in large outdoor and indoor pig units in Hungary/Romania/Serbia. AB - Gross postmortem pathologic-anatomical and bacteriological examination, and serotyping for Haemophilus parasuis were performed in 401 weaned pigs with the clinical symptoms of Glasser's disease, originating from indoor (204 piglets out of 12 units) and outdoor (197 piglets out of 9 units) units in the country triangle Hungary/Romania/Serbia. The majority of the isolates in indoor units included Haemophilus parasuis serotypes 2, 5 and non typable, while in outdoor units, a high diversity (serotypes 1, 2, 4, 5, 11, 12, 14, 15, and non typable) of Haemophilus parasuis serotypes was found. PMID- 15298055 TI - [New strategies for drug development]. AB - The strategy how new drugs are being developed has dramatically changed during the last decade. Until then a rational, target-aimed design of drugs was not possible, because the molecular basis of disease was largely unexplored. The progress in basic science has revealed that many diseases are caused by the malfunction of proteins. These proteins are targets for drugs, which should bind to the protein and inhibit its function. Knowledge of the three dimensional structure of the target protein is helpful to find binding pockets for the drug on the molecular surface. The strategy of this knowledge-based or structure-based drug-design is explained with the help of several examples for recently approved drugs, antivirals against influenza and AIDS, and a drug against a rare cancer, chronic myeloic leukemia. PMID- 15298056 TI - [The efficacy of different oral dosage forms of furazolidone for E. coli infections in carrier pigeons]. AB - The clinical efficiacy of furazolidon for treatment of E. coli-induced gastro intestinal infections in racing pigeons was investigated. 36 adult pigeons were treated with 2 different oral modes of application (capsule/drinking water) with a daily therapeutic dosage of 12.5 mg furazolidon/pigeon. The pigeons used for this study (Columba livia f. domestica) originated from conventional breeders and were housed in 3 different groups (control-, capsule- and powder-group) in different stables. After infection with an E. coli-strain (O150:H8) that proved to be pathogenic for pigeons, the animals developed clinical signs of disease within 2 days. After onset of disease the treatment with furazolidon for 5 days started. This phase was followed by an adspectory phase for 6 days. The negative identification of the E. coli O150:H8 was determined as main parameter for the clinical efficiacy of the treatment with furazolidon. This parameter showed a highly significant (p = 0.0001) difference between both groups treated with furazolidon and the control group. In both groups treated with furazolidon the E. coli strain could not be isolated after the end of the treatment. An improvement of clinical signs was seen 24 hours after treatment via capsule and 48 hours after treatment via drinking water formulation. The time difference might be caused by the high concentration of furazolidon in the capsules due to the single daily application. Considering the inaccurate dosing via drinking water that results from the varying drinking water intake in pigeons, the application by capsule should be prefered. Both furazolidon preparations proved to be effective in treating gastro-intestinal E. coli-infections in racing pigeons in a dosage of 12.5 mg/pigeon for 5 days, however, best results were obtained by application via capsule. PMID- 15298054 TI - Toxoplasma gondii antibodies in the white stork Ciconia ciconia. AB - The prevalence of Toxoplasma gondii in chicks of wild birds and captive individuals was studied in the Poznan environs and in the Poznan Zoological Garden in the years 2002-2003. Bird blood was tested for T. gondii antibodies by an indirect fluorescent antibody test. T. gondii antibodies were detected from 5.8% of 205 analysed white stork chicks and 13.6% of 44 analysed adult storks in the zoo. Because toxoplasmosis is one of the more common parasitic zoonoses worldwide, we briefly discuss the potential epidemiological importance of stork toxoplasmosis to humans. PMID- 15298057 TI - [Clinical comparison of isoflurane and sevoflurane anaesthesia in the gerbil (Meriones unguiculatus)]. AB - The objective of this study was a comparison of the volatile anaesthetics isoflurane and sevoflurane in terms of their clinical effects in gerbils (Meriones unguiculatus) (n=12 each). Induction of anaesthesia was performed in a body chamber with an anaesthetic concentration of 4.0 Vol.% at an oxygen flow of 500 ml/min for isoflurane and 8.0 Vol.% at an oxygen flow of 1000 ml/min for sevoflurane, respectively. Anaesthesia was maintained via nose cone with an anaesthetic concentration of 2.8 to 3.2 Vol.% at an oxygen flow of 200 ml/min for isoflurane and 5.0 to 5.2 Vol.% at an oxygen flow of 400 ml/min for sevoflurane. Those anaesthetic concentrations ensured reflex status conform with surgical tolerance. In spite of its higher blood-gas coefficient induction time was slightly faster for isoflurane. Recovery time was significantly longer in the isoflurane group than it was in the sevoflurane group. Both inhalants caused respiratory depression. Respiratory rate was lower in sevoflurane animals compared to isoflurane. The animals were positioned on a heating pad immediately after induction, thus a decrease of the body temperature could be prevented. Both inhalants can be recommended for usage in gerbils. Sevoflurane showed no clinical benefit compared to isoflurane. PMID- 15298058 TI - Clinical efficacy of meloxicam (Metacam) and flunixin (Finadyne) as adjuncts to antibacterial treatment of respiratory disease in fattening cattle. AB - The clinical efficacy of two non-steroidal anti-inflammatory drugs (NSAIDs), meloxicam (Metacam 20 mg/ml) and flunixin meglumine (Finadyne), as adjuncts to antibacterial therapy in the treatment of acute febrile respiratory disease in cattle was compared. The randomised blind, positive controlled study was conducted under feedlot conditions in Mexico. Overall, 201 female cattle (weighing 220-250 kg) diagnosed with bronchopneumonia at the feedlot were recruited into the study. On Day 0 all animals were treated with 20 mg oxytetracycline/kg body-weight (Bivatop 200) by subcutaneous injection, in conjunction with either meloxicam (0.5 mg/kg subcutaneously, Metacam 20 mg/ml, n = 100), or flunixin meglumine (2.2 mg/kg intravenously, Finadyne, n = 101). According to label instructions, meloxicam was administered as a single dose, whereas flunixin meglumine could be administered daily for up to 3 consecutive days depending on the rectal temperature (with re-administration, if rectal temperature > or = 40.0 degrees C). Rectal temperature, respiratory rate, appetite, dyspnoea, coughing, nasal discharge and general condition were recorded on Days 0 (prior to treatment), 1, 2, 3 and 7 using a weighted numerical score. Scores were summed to generate a 'Clinical Sum Score' (CSS, range 7 to 24 points). Individual animal body weights were measured on Days 0 and 7. Nasal swabs were collected from 10 animals per treatment group on Day 0 for microbiological culture. Clinical parameters and the mean CSS showed no significant differences between treatment groups with mean CSS on Days 0 and 7 of 16.18 and 10.55 in the meloxicam group and 16.41 and 10.88 in the flunixin meglumine group. However, a significantly lower mean rectal temperature was measured in the meloxicam group on Day 2 (p < or = 0.01). No significant differences in mean body weights were found between groups. Repeated administration of flunixin meglumine was performed in 45% of the animals. No suspected adverse drug events related to treatments were reported. It is concluded that a single subcutaneous dose of meloxicam was as clinically effective as up to 3 consecutive daily intravenous doses of flunixin meglumine when used as an adjunctive therapy to antibacterial therapy in the treatment of acute febrile respiratory disease in feedlot cattle. PMID- 15298059 TI - Effects of a dietary chitosan and calcium supplement on Ca and P metabolism in cats. AB - The aim of the present study in cats was to investigate the potential effects of a calcium carbonate and chitosan supplement on blood parameters in aged cats with moderate chronic renal failure and on the mineral balance in adult healthy cats. For the trials, 10 neutered cats 2-4 years of age were fed for 21 days and six neutered cats (2 male and 4 female), 14 years of age, with elevated urea and phosphorus level in the plasma were fed for 35 days with a supplement. The apparent digestibility of phosphorus was (p < 0.05) reduced in the treatment period. Plasma urea inorganic phosphate decreased significantly (p < 0.05) in the old cats after 35 days of treatment. The treatment had a significant effect on the phosphorus, gross energy, dry matter, crude ash, crude fiber and crude protein digestibility in adult healthy cats. The practical implication could be an alternative treatment option for cats refusing to ingest veterinary renal diets. PMID- 15298060 TI - [Body and claw measurements and pressure distribution under the claws in calves of different cattle breeds]. AB - In 58 female calves of the three breeds German Holstein (GH), German Brown (GB) and German Red (GR) the body weight and height were determined at the age of 79 to 188 days. At the right front limb and the left hind limb the following claw measurements were taken: dorsal border length, diagonal length, heel length and height, angle of the dorsal border and hardness of the claw horn. Furthermore, the punctual pressure under the medial and the lateral claw of the right front limb and the left hind limb were determined using an electronic measuring system. The GH calves weighing 136.8 kg were heavier than GB, which was the tallest breed with 106.2 cm height at the sacral bone. GR calves were the smallest (99.1 cm) and weighed 121.1 kg. The length of the diagonal and the angle of the dorsal border were smaller at the hind limb of all breeds which resulted in a smaller area of ground surface for the claw of this limb. The GR calves had the longest and flattest claws. The GB showed the highest pressures per cm2 at the front limb with 25.6 N/cm2 under both claws as well as at the hind limb with 26.7 N/cm2. The AR had the smallest pressure load with 20.8 N/cm2 at the front limb and 20.2 N/cm2 at the hind limb. The animals showed a larger relative weight load and area of ground surface on the medial claws of the front and hind limbs than on the lateral claws. The highest pressures were found underneath the rear part of the medial claw in all breeds. GB calves showed the highest average pressures (33.3 N/cm2 at the front limb) while GR calves had the lowest (26.3 N/cm2 at the hind limb). GR calves had the highest claw hardness at all measuring positions. PMID- 15298061 TI - [The incidence of agglutination and its influence on sperm quality and fertility of boar semen]. AB - The aim of this study was to examine the incidence of sperm agglutinations and their relationships with sperm quality and fertility. Semen samples of 40 boars of an AI-station were investigated. Nineteen of the 40 investigated boars showed a constantly low (< 10% agglutinated sperm), 3 an intermediate (10-20%) and 6 boars a high level (> 20%) of agglutination in raw semen. The degree of agglutination in sperm samples of 12 boars varied distinctly during the investigation period. During summer more (P < 0.05) agglutinated sperm were observed (11.0 +/- 11.6%) than during winter (6.2 +/- 7.3%). There was no association between bacterial contamination and incidence of agglutinations (P > 0.05). After dilution in extender the percentage of agglutinated sperm decreased from 6.2 +/- 7.3% to 1.1 +/- 1.4% (P < 0.0001). Twenty-four hours after dilution the percentage of progressively motile sperm was 7.4% lower (P < 0.05) in ejaculates with an initially high degree of agglutination (> 20% agglutinated sperm) compared to samples with an initially low degree of agglutinated sperm (< 10%). Plasma membrane integrity, mitochondrial membrane potential, acrosome reaction and chromatin structure were independent (P > 0.05) from the level of agglutination. Fertility data did not differ (P > 0.05) between boars with low and high numbers of agglutinated sperm in raw semen. The results show that there are individual, ejaculatory and seasonal variations in the incidence and degree of agglutination. Agglutinations have a negative effect on motility of sperm and disappear to a large extent after dilution in sperm extender. They have no negative consequences on fertility. PMID- 15298062 TI - [Cauda equina compression syndrome (CECS): retrospective study of surgical treatment with partial dorsal laminectomy in 86 dogs with lumbosacral stenosis]. AB - We report our results of partial lumbosacral laminectomy for treatment of canine Cauda equina Compression Syndrome due to a lumbosacral stenosis. Opposite to conventional techniques of dorsal laminectomy, only widening of the Spatium interarcuale is performed. This is achieved by exstirpation of the Lig. flavum and partial dorsal laminectomy of the first sacral segment. The Proc. spinosi and integrity of facet joints are fully maintained by this technique. In 96.5% of 86 dogs treated with this method relief of dorsal pressure and permanent rapid regression of clinical symptoms was achieved. In two cases recurrence of clinical symptoms was observed during follow up and one case showed no improvement at all. In conclusion partial dorsal laminectomy is a minimal invasive technique for treatment of Cauda equina compression syndrome expressed by pain reaction accompanied by minor neurological deficits caused by lumbosacral stenosis. Maintained spinal stability allows short reconvalescence and the unrestricted use of dogs immediately post operation. PMID- 15298063 TI - [Cultivation and expansion of canine Schwann cells using reexplantation]. AB - Despite the numerous available possibilities for the surgical treatment of peripheral nerve lesions found in the dog, the success of these treatments is often unsatisfactory. It has been proven that Schwann cells (SC) have a positive influence on the regeneration of nerve stumps. Implanting a guidance channel seeded with autologous SC at the lesion site could be a new therapeutic approach. The aim of this research was to investigate the in vitro cultivation and expansion of canine SC as the main requirement for the treatment referred to above. Biopsies were carried out on 17 nerve samples originating from dogs of different breed, age, gender and condition. The reexplantation method was employed, followed by dissociation using hyaluronidase, collagenase and trypsin and further expansion. The samples were divided into six groups which were treated with a varying combination of mitogens (forskolin, bovine PEX, choleratoxin, heregulin). To obtain the quantities of SC, the specimens were immunostained by a p75-antibody. By employing a growing number of agents it was possible to obtain an increase in both the quantity of cells and purity of cultures. A maximum of 16x10(5) cells per millilitre of suspension was achieved. The largest SC purity measured 27.1%. The maximum SC quantity achieved was 43.3x10(4) SC per millilitre. PMID- 15298064 TI - Stem cell therapy: opportunities and challenges. PMID- 15298065 TI - Inflammation and autoimmunity as a central theme in neurodegenerative disorders: fact or fiction? AB - Irrespective of the initiating stimuli, neurodegenerative disorders including multiple sclerosis (MS), Alzheimer's disease, Parkinson's disease and stroke share many characteristics of inflammation and autoimmunity. This review summarizes and correlates the information relating to the role of cytokines and chemokines in initiating and propagating the inflammatory/immune response in these pathologies. For example, in MS there is a continuous realignment in the inflammatory and immune response. However, due to the redundancy in the cytokine/chemokine response, it is extremely unlikely that any one therapy will be successful in treating neurodegenerative diseases. This review attempts to highlight specific targets for therapeutic intervention. PMID- 15298066 TI - Therapeutic potential of stem cells in central nervous system regeneration. AB - Neurodegenerative disorders and traumatic brain injury result in the loss of specific neuronal populations. Stem cells are self-renewing, multi- or pluripotent cells capable of differentiating into a wide range of cell types, properties which make stem cells a potentially invaluable source of transplantable cells. Recent experimental studies have indicated that several stem cell populations have the ability to replace lost neurons and to repair the damaged nervous system following transplantation. This review evaluates the potential of various stem cell populations in the treatment of human neurodegenerative conditions and traumatic brain injury. PMID- 15298067 TI - Drugs in development for Parkinson's disease. AB - Pharmacological treatment of Parkinson's disease (PD) is entering a new and exciting era. Real promise now exists for the clinical application of a large range of molecules in development that will combat different aspects and stages of the condition. These include methyl- and ethyl-esterified forms of L-dopa (etilevodopa and melevodopa), inhibitors of enzymes such as monoamine oxidase type-B (eg, rasagiline), catechol-O-methyl transferase (eg, BIA-3202) and the monoamine re-uptake mechanism (eg, brasofensine). In addition, a range of full and partial dopamine agonists (eg, sumanirole, piribedil and BP-897) and their new formulations, for example, patch delivery systems (eg, rotigotine) are being developed. We also highlight non-dopaminergic treatments that will have wide ranging applications in the treatment of PD and L-dopa-induced dyskinesia. These include alpha2 adrenergic receptor antagonists (eg, fipamezole), adenosine A2A receptor antagonists (eg, istradefylline), AMPA receptor antagonists (eg, talampanel), neuronal synchronization modulators (eg, levetiracetam) and agents that interact with serotonergic systems such as 5-hydroxytryptamine (5-HT)1A agonists (eg, sarizotan) and 5-HT2A antagonists (eg, quetiapine). Lastly, we examine a growing number of neuroprotective agents that seek to halt or even reverse disease progression. These include anti-apoptotic kinase inhibitors (eg, CEP-1347), modulators of mitochondrial function (eg, creatine), growth factors (eg, leteprinim), neuroimmunophilins (eg, V-10367), estrogens (eg, MITO-4509), c synuclein oligomerization inhibitors (eg, PAN-408) and sonic hedgehog ligands. PMID- 15298068 TI - Medicinal cannabis extracts for the treatment of multiple sclerosis. AB - Prior to 2002, few clinical data were available to indicate whether cannabis extracts may be beneficial. However, in the last two years, results of several placebo-controlled clinical trials of orally administered compounds have been published, and these cast doubt on the efficacy of delta9-tetrahydrocannabinol (delta9-THC) in objectively reducing spasticity in MS. By contrast, it has been claimed that sublingually administered cannabis extracts that contain approximately equal concentrations of delta9-THC and cannabidiol, a natural cannabinoid that does not act on the CB1 receptor, can produce a statistically and clinically significant reduction in spasticity, although this claim has yet to be thoroughly validated. Nonetheless, results of preclinical trials also lend support to the hypothesis that the endogenous cannabinoid system may be involved in the regulation of spasticity and pain. A better indication of the clinical potential of the different cannabis extracts will have to await the publication of the most recent clinical trial data. This review critically evaluates the most recent evidence available on the potential use of medicinal extracts of cannabis to relieve pain and spasticity in multiple sclerosis. PMID- 15298069 TI - CGRP receptor antagonists: a new choice for acute treatment of migraine? AB - The neuropeptide calcitonin gene-related peptide (CGRP) is believed to play a central role in the underlying pathology of migraine. Serum levels of CGRP, elevated during a migraine attack, return to normal as pain alleviates. Recently, a causative role for CGRP in migraine has been suggested. Based on these findings, it was proposed that blockade of postsynaptic CGRP receptors, and hence the physiological effects of CGRP, should effectively abort a migraine attack. This review will discuss the therapeutic potential of olcegepant, the first non peptide CGRP receptor antagonist available for human studies, within the context of current neurovascular theories on migraine pathology. PMID- 15298070 TI - Antidepressants in the treatment of irritable bowel syndrome and visceral pain syndromes. AB - Irritable bowel syndrome (IBS) is characterized by abdominal pain associated with disordered defecation, which may include urgency and altered stool frequency. Visceral pain syndromes, including IBS, may be effectively treated by a variety of therapies that modulate the interactions between the central and enteric nervous systems. Clinical observations and preliminary data suggest that antidepressants may be efficacious for the treatment of these syndromes. The tricyclic antidepressants (TCAs) have been utilized most extensively in this area, but there is a need for more rigorous efficacy data. Serotonin, an important neurotransmitter in both the central and enteric nervous systems, modifies both motility and sensation in the gut. Recognition of the importance of serotonin in digestive motility and sensation has sparked interest in the use of agents that modify serotonergic transmission in visceral pain syndromes. Pharmacological therapeutics that modulate the biological amines (serotonin, norepinephrine, dopamine and catecholamines) both peripherally and within the central nervous system may offer more effective therapies for these disorders. The selective serotonin reuptake inhibitors are commonly used in clinical practice, but more rigorous, controlled studies are needed to determine their effects beyond the treatment of psychiatric comorbidity. The newer generation antidepressants may provide additional insight into the pathophysiology of the brain-gut interactions and their relationship to functional bowel disorders, providing new therapeutic interventions. PMID- 15298071 TI - Utility of selective serotonin reuptake inhibitors in premature ejaculation. AB - The introduction of selective serotonin reuptake inhibitors (SSRIs) has revolutionized our understanding of the treatment of premature ejaculation. Lifelong premature ejaculation may be a neurobiological phenomenon, namely part of a biological variability of the intravaginal ejaculation latency time in men. Animal studies support this view, and an animal model for premature and delayed ejaculation has recently been developed. It is proposed that drug treatment of premature ejaculation should consist of 5-hydroxytryptamine (5-HT)2c receptor stimulation and/or 5-HT1A receptor inhibition. A meta-analysis of 35 daily treatment studies with selective serotonin reuptake inhibitors (SSRIs) and clomipramine demonstrated comparable efficacy of clomipramine with the SSRIs sertraline and fluoxetine in delaying ejaculation, whereas the efficacy of the SSRI paroxetine was greater than all other SSRIs and clomipramine. It is postulated that acute treatment with SSRIs, including those with short half lives, will not produce an ejaculation delay equivalent to that induced by daily treatment of SSRIs. PMID- 15298072 TI - GW-1000. GW Pharmaceuticals. AB - GW Pharmaceuticals is developing GW-1000 (Sativex), a narrow ratio delta9 tetrahydrocannabinol:cannabidiol product for the potential treatment of multiple sclerosis, spinal cord injury, neurogenic pain and peripheral neuropathy. In March 2003, the company filed for approval for the treatment of MS with the UK Medicines Control Agency, and in May 2004, filed for new drug submission with Health Canada. PMID- 15298073 TI - NCX-701. NicOx. AB - NicOx is developing NCX-701, a nitric oxide-releasing derivative of paracetamol (acetaminophen), for the potential treatment of inflammation and pain. The compound is currently undergoing phase III clinical trials. PMID- 15298074 TI - Z-338. Zeria/Yamanouchi. AB - Zeria and Yamanouchi are co-developing Z-338, a thiazole-4-carboxamide derivative with prokinetic activities, as a potential treatment for dyspepsia and gastric motility disorders. This compound is currently in phase II clinical trials. PMID- 15298075 TI - Gaboxadol. Lundbeck/Merck. AB - H Lundbeck A/S, in collaboration with Merck & Co Inc, is developing gaboxadol, a GABAA agonist, for the potential treatment of sleep disorders. The compound is currently undergoing phase III clinical trials. PMID- 15298076 TI - Compassion fatigue following the September 11 terrorist attacks: a study of secondary trauma among New York City social workers. AB - Experience suggests that individuals working in the caring and psychotherapeutic professions are among those to provide mental health services to disaster victims suffering from psychological trauma following catastrophic events. Yet, few studies have focused on the emotional exhaustion from working with such clients, referred to as compassion fatigue (CF) in this study, and how CF differs from other occupational hazards, such as secondary trauma (ST) and job burnout. In the present study, we used recently validated scales to predict ST and job burnout related to providing services to those affected by the World Trade Center (WTC) attacks. Our study data were based on a random survey of 236 social workers living in New York City (NYC), over 80% of which reported being involved in post WTC disaster counseling efforts. Our analyses indicated that controlling for demographic factors, years of counseling, and personal trauma history, ST was positively associated with WTC recovery involvement (p <. 001) and negatively associated with having a supportive work environment (p < . 01). In contrast, job burnout was negatively associated with having a supportive work environment (p < .01), but not associated with WTC involvement or WTC counseling efforts. We discuss these results in light of future conceptual and empirical research needs. PMID- 15298077 TI - Objective assessment of secondary trauma. AB - This study reports on the continued development of the Secondary Trauma Scale (STS) and the establishment of cutoff scores. Cutoff scores are unavailable for existing secondary trauma scales. Participants were 118 young adult volunteers who reported having had close and continued exposure to a person or persons who had been traumatized. Sub-samples were drawn from the initial pool, and these sub samples showed mild to severe anxiety and depression along with problematic levels of intrusion and avoidance. STS scores of 38 or higher were associated with mild to severe anxiety and depression and with problematic intrusion and avoidance symptoms. It was suggested that at STS scores of 45 or higher, clinicians and researchers should be particularly concerned. The STS demonstrated strong psychometric characteristics and was significantly correlated with known measures of emotional distress and dissociation. PMID- 15298078 TI - Police trauma encounters: precursors of compassion fatigue. AB - Given frequent assignments of responding to critical situations, police officers are a high-risk population for exposure to traumatic stress. It was hypothesized that types and increased frequencies of certain traumas lead to increased risk for PTSD symptoms and eventually to a state of compassion fatigue through secondary processes. Compassion fatigue was conceptualized as the cost of caring without reward or result. Results indicated that the homicide of another officer in the line of duty and dealing with victims of serious crime resulted in the greatest increased risk of trauma symptoms. Gender differences were found in trauma risk, with women officers experiencing higher risk from dealing with abused children. and male officers experiencing higher risk due to shooting incidents where officers were involved. It was concluded that increased frequency and type of traumas, especially those occurring to other co-workers and those associated with gender, may eventually lead to a secondary process of emotional compassion fatigue. Available strategies for prevention are discussed PMID- 15298079 TI - Issues and controversies in the understanding and diagnosis of compassion fatigue, vicarious traumatization, and secondary traumatic stress disorder. AB - Understanding the effects of prolonged contact, in a professional role, with trauma victims has led to conceptualizations of helper stress. Various terms such as compassion fatigue, vicarious traumatization, secondary traumatic stress reactions, empathic strains, burn out, and Type land Type II countertransference have been proposed These terms required conceptual classification to make a proper diagnosis and classification of their impact on the helping process. It is proposed that Traumatoid States is a more inclusive and accurate term to define sub-types of occupationally-related stress response syndromes (OSRS). PMID- 15298080 TI - Myokimia and potassium channel gene Kv1.1. PMID- 15298081 TI - Respiratory insufficiency as a presenting symptom of LGMD2D in adulthood. AB - Several forms of recessive limb girdle muscular dystrophy (LGMD2C-F) are due to mutations in genes coding for sarcoglycans. Clinically, most sarcoglycanopathies present in childhood with skeletal muscle wasting and early loss of ambulation; respiratory insufficiency is rare. However, some cases of LGMD2D with a late onset and a milder course have been reported. In this study, two adult brothers, compound heterozygous for two missense mutations of the SGCA gene (Arg77Cys, Val247Met), presented with respiratory insufficiency while they were still ambulatory. PMID- 15298082 TI - Mutations in the Myelin Protein Zero result in a spectrum of Charcot-Marie-Tooth phenotypes. AB - Initially the Myelin Protein Zero gene was shown to be mutated in the demyelinating form of Charcot-Marie-Tooth disease (CMT1). The vast majority of the mutations in the Myelin Protein Zero gene have been detected in the Charcot Marie-Tooth (1B) disease, however, some of them were found in patients suffering from congenital hypomyelinating neuropathy and axonal type Charcot-Marie-Tooth disease. In this study, a Charcot-Marie-Tooth disease phenotype diversity associated with different mutations in the MPZ gene, is described. PMID- 15298083 TI - Oral antibiotics and breast cancer. PMID- 15298084 TI - Walking in peace. PMID- 15298085 TI - Dr. S. Y. Tan's thoughts on lifelong learning and CME. PMID- 15298086 TI - The Prepaid Health Care Act. PMID- 15298087 TI - Physicians and complementary-alternative medicine: training, attitudes, and practices in Hawaii. AB - INTRODUCTION: There were only few studies addressing the physicians' training, attitudes, and utilization of complementary and alternative medicine (CAM) therapies, compared to the well-documented escalating use of CAM among consumers. Patients who use CAM, however often do not disclose their utilization to their physicians. This study thus surveyed knowledge, attitudes, and practices of complementary and alternative medicine among physicians on the island of Oahu in Hawaii. The Hawaii Medical Service Association (HMSA) provided physicians' names and contact information. METHOD: This is a descriptive study with a questionnaire that was mailed to all physicians on the list. A total of 299 physicians responded to this survey. Response rate was 17.45%. RESULTS: Physicians reported having moderate acknowledge in acupuncture, massage, prayer/spiritualty chiropractic, hypnosis and meditation. They recognized the value of these treatments and refer their patients to have these treatments. On the other hand, homeopathy naturopathy, electromagnetic therapies, therapeutic/healing touch, and nutriceuticals were consistently rated as having no role in conventional medicine, strongly opposed to in practice, or would not refer patients to. Respondents felt CAM could be most effective for pain, musculoskeletal, psychological conditions and smoking cessation. DISCUSSIONS: Results indicated that the respondents had knowledge about prayer/spirituality, massage, chiropractic, meditation, hypnosis, and acupuncture, and also rated them as playing a role in conventional medicine, and would refer or have referred patients to. When they had little knowledge about naturopathy, electromagnetic therapies, nutriceuticals, and homeopathy, they rated these therapies as having no role in conventional medicine, and were strongly opposed to in practice or would not refer patients to. Respondents reported the least know abut the Aryurveda and Native American medicine. However, there was no significant correlation between negative attitude and practice patterns. Similarly therapeutic touch and chiropractic were perceived as therapies, but no role in conventional medicine although the respondents reported having some knowledge of these therapies. Thus, knowledge may not be necessarily associated with negative attitudes and practice patterns. PMID- 15298088 TI - Are immune responses pivotal to cancer patient's long term survival? Two clinical case-study reports on the effects of Morinda citrifolia (Noni). AB - In the State of Hawaii, there are abundant claims of benefit from cancer patients' use of the fruit juice of Morinda citrifolia (Noni). There is no well documented clinical report in peer review journals. The author here studiously examined 2 such claims through interview, review of the medical records and pathology slides. The author concludes that these cases are valuable experiences and hope to stimulate interest in Noni research as an important part of adjuvant immunotherapy for cancer PMID- 15298089 TI - Water fluoridation information found on the World Wide Web. AB - OBJECTIVE: There is much controversy over the fluoridation of community water supplies. This study surveyed the informational content of internet sites on the World Wide Web (WWW) regarding water fluoridation. METHODS: One hundred websites were identified and 59 were evaluated with a 6 point scoring system using predetermined criteria. RESULTS: Of these 59 sites, 54% recommend water fluoridation as compared to 31% that oppose it. CONCLUSIONS: The informational content of WWW sites may range from factual, to unsubstantiated opinions, to frank fraud. However, this information is presented to the public indiscriminately. The informational content of the WWW contains varied views and recommendations for community water supply fluoridation. Public health officials and practicing physicians should be aware that the internet WWW may influence public opinion. PMID- 15298090 TI - Experiencing life as a rural health professional. PMID- 15298091 TI - AANCART comes to Hawaii. AB - There are 176,707 foreign-born Asian Americans living in Hawaii, according to the 2000 Census. Data from the Hawaii Tumor Registry show that foreign-born Asian Americans in Hawaii are diagnosed with cancer at a later stage than the white population in Hawaii. It is the intent of AANCART at the Cancer Research Center of Hawaii to address this disparity of cancer status of the foreign-born Asian American population in Hawaii. PMID- 15298092 TI - Rural minority and multicultural preventive care, primary care, and mental health issues: challenges and opportunities. PMID- 15298094 TI - Mental health and substance abuse services among rural minorities. AB - This paper provides a brief overview of current conditions and prospects for increased access to mental health and substance abuse services among rural minorities. First, it addresses challenges in ensuring rural minorities access to needed services. Second, it considers steps to increase rural minority participation in the mental health and substance abuse workforce. The dual emphasis is on (1) reaching now the isolated, rural, and frontier minority populations in need of these services and (2) building a rural health workforce that is reflective of rural minority cultures and offers continually higher quality and sustainable services to rural and minority populations. PMID- 15298093 TI - Disparities in access to care among rural working-age adults. AB - CONTEXT: Nationally, minority population disparities in health and in the receipt of health services are well documented but are infrequently examined within rural populations. PURPOSE: The purpose of this study is to provide a national picture of health insurance coverage and access to care among rural minorities. METHODS: A cross-sectional analysis using the 1999-2000 National Health Interview Surveys examined insurance status and receipt of ambulatory care during the past year. Multiple logistic regression was used to measure factors influencing the odds of insurance coverage and a provider visit. FINDINGS: Among rural minority adults, 32% of blacks, 35% of "other" race persons, and 45% of Hispanics were uninsured compared to 18% of whites. Differences in insurance status were not significant for rural blacks and Hispanics after resources such as education, income, and employment were held constant. Examining use, 37% of rural Hispanics and 27% of blacks, versus 20% of whites and 19% of persons of other race, had not made a health care visit in the past year. When resources were held constant, blacks and persons of other race/ethnicity no longer differed from whites, but differences among Hispanics persisted. CONCLUSIONS: A comprehensive approach to the health needs of rural working age adults must consider the unique characteristics of rural communities and populations, requiring cultural as well as financial creativity in the design of health delivery systems. The importance of resources such as education and employment points to the need to link health problems to area-specific rural economic development. PMID- 15298095 TI - Rural community-dwelling elders' reports of access to care: are there Hispanic versus non-Hispanic white disparities? AB - CONTEXT: Consumer reports can provide useful information about the dimensions of access in need of improvement for particular population subgroups. PURPOSE: To determine if there are Hispanic versus non-Hispanic white disparities in rural elders' reports of their health care access. METHODS: A telephone survey was conducted among 2,097 rural community-dwelling elders in West Texas. Dependent variables included reports of the ability to obtain care (see personal doctor/nurse, see specialist, obtain help over phone, and obtain transportation to the clinic) and reports of the ability to obtain care without a long wait (get help over the phone without a long wait, see provider for illness/injury when wanted, see provider for routine care when wanted, and have short office waiting times). Independent variables included predisposing, enabling, and need factors. Univariate and multivariate logistic regression analyses were conducted. FINDINGS: In univariate logistic analyses, Hispanics had worse reports of their ability to always/ usually see their personal doctor, see a specialist, obtain transportation to the clinic, see a doctor for illness/injury when wanted, and see a doctor for routine care when wanted. When adding enabling factors to the models, only reports of the ability to see a doctor for illness or injury and for routine care when wanted remained significant. CONCLUSIONS: Though the rural medical care system may need to target directly Hispanics to improve their timely access to acute and routine care, the enhancement of health insurance coverage may lead to improved access to personal doctors and specialists among all rural elders. PMID- 15298096 TI - Factors influencing the retention and attrition of community health aides/practitioners in Alaska. AB - CONTEXT: The Community Health Aide Program (CHAP) is a unique program employing local, indigenous peoples as primary care nonphysician providers in extremely remote frontier, tribal Alaskan communities. With attrition rates up to 20%, recommendations for improving retention are necessary to maintain access to health services for Alaska Natives in these communities. PURPOSE: The purpose of this study was to identify factors contributing to retention in Alaska's CHAP program. METHODS: Key informant interviews were conducted with 41 community health aides/practitioners (CHA/Ps) in 15 villages statewide. Efforts were made to ensure the sample included a mix of villages with high retention of health aides and villages with lower retention. Geographic and ethnic diversity were also considered. Transcripts were coded using NUD*IST software, and data were analyzed for differences between high retention and low retention villages and between more experienced and less experienced CHA/Ps. FINDINGS: Five fundamental needs of health aides were identified as crucial for retention of personnel. These needs include strong co-worker support, access to basic training, a fully staffed clinic, good community support, and supportive families. CONCLUSIONS: For 35 years, the CHAP program has worked to diminish health disparities for Alaska Natives. Though unique challenges associated with the job have factored into low retention of CHA/Ps, improved retention is possible with easier access to basic training, increased support from colleagues and community, enhanced team-building skills, and better on-call schedules. PMID- 15298097 TI - Health differences among Lumbee Indians using public and private sources of care. AB - CONTEXT: Of 2.4 million American Indians, approximately 60% are eligible to receive Indian Health Service (IHS) benefits, leaving many to seek care elsewhere. It is unknown if their quality of care, health behaviors, and health status vary by source of care, as demonstrated for other populations. PURPOSE: The purpose of this study was to determine whether preventive services, health behaviors, and number of health conditions vary as a function of having non-IHS public versus private physicians as sources of usual care. METHODS: 1,177 Lumbee Indians, who are ineligible to receive IHS services, completed a telephone interview that included information on receipt of preventive measures, tobacco use, physical activity, breast self-examination, and medical conditions. Frequencies, chi-squares, t tests, odds ratios, and confidence intervals were used to compare variables by source of care. FINDINGS: 939 respondents (80%) had a private and 210 (18%) a public health clinic physician as their usual source of care; 28 (2%) reported having neither. Logistic regression analyses, restricted to the 1,149 participants who reported either a private or public source of care, revealed no differences in receipt of preventive services or health status by usual source of care. Smokeless tobacco use was less common among persons using private than public providers. CONCLUSIONS: Lumbees whose usual source of care was a public clinic physician did not differ in receipt of preventive services or in health status compared to their counterparts who received care from a private physician. More targeted research into health similarities and differences arising from access to public and private sources of care is warranted. PMID- 15298099 TI - Access to health care among Latinos of Mexican descent in colonias in two Texas counties. AB - CONTEXT: Critical to resolving the problem of health disparities among Latinos is examining the needs within ethnic subpopulations. This paper focused on the unique challenges encountered by one ethnic subpopulation-Latinos of Mexican descent living in colonias. Findings reaffirm the importance of looking within ethnic subpopulations to understand the complexities of health disparities. PURPOSE: This paper reports on data collected measuring access to health care among Latinos of Mexican descent living in several colonias in two southern Texas counties. METHOD: Observations are based on data gathered from a non-probability sample obtained through a face to face questionnaire focusing on health care access. Persons living in two Texas counties near the US border were included in this sample, and demographic data were compared to county, state and national statistics to examine the comparability of the sample to similar populations. FINDINGS: Findings from this snowball sample, N=271, suggested lower rates of health insurance coverage compared to the Latino population nationwide, decreased patterns of preventive screenings for blood pressure, diabetes, cholesterol, cancers (ie, breast, cervical, and prostate), and regular access to health care in Mexico by slightly more than half the sample. CONCLUSIONS: Seeking care in Mexico may be a viable solution for many indigent people of Mexican descent living in close geographic proximity to the border because it surmounts the political, cultural, linguistic, or economic barriers to health care services in the United States. Nonetheless, there are longer term questions regarding quality of care and health and wellness for this group of people. PMID- 15298098 TI - Improving access to primary care for a growing Latino population: the role of safety net providers in the rural Midwest. AB - CONTEXT: Many rural Midwestern communities are experiencing rapid growth in Latino populations with low rates of health insurance coverage, limited financial resources, language and cultural differences, and special health care needs. PURPOSE: We report on 2-day site visits conducted in 2001 and 2002 in 3 communities (Marshalltown, Iowa; Great Bend, Kansas; and Norfolk, Nebraska) to document successful strategies to meet Latino health care needs. METHODS: We interviewed key informants to identify successful community strategies for dealing with health care access challenges facing the growing Latino population in the Midwest. FINDINGS: Interventions have been developed to meet new demands including (1) use of free clinics, (2) school health programs, (3) outreach by public health, social services and religious organizations, and (4) health care providers' efforts to communicate with patients in Spanish. Strain on safety net services for Latinos is due in part to a complicated and unstable mix of public and private funds, a large but overtaxed volunteer provider base, the dependence on a limited number of community leaders, and limited time for coordination and documentation of activities. CONCLUSIONS: We suggest the development of a Rural Safety Net Support System to provide targeted funding to rural areas with growing immigrant populations. Federal community health center support could be redirected to new and existing safety net providers to support the development of a safety net monitoring system. PMID- 15298100 TI - Cost analysis in telemedicine: empirical evidence from sites in Arizona. AB - CONTEXT: Support of telemedicine for largely rural and ethnically diverse populations is premised on expectations that it increases opportunities for appropriate and timely medical services, and that it improves cost-effective service delivery. PURPOSE: To understand the cost-effectiveness of telemedicine in 8 small and/or rural sites in Arizona. METHODS: A cost analysis framework was used to measure the efficacy of telemedicine in the selected sites from May 1, 2000, to April 30, 2001. FINDINGS: The costs for telemedicine services in half the study sites were more than the costs for conventional face-to-face diagnosis when the volume of telemedicine services used at a site was relatively low. This result persisted even when the opportunity cost for the patients in accessing more traditional types of care was included in the cost estimates. CONCLUSIONS: These preliminary findings suggest that telemedicine in some instances may not be cost-effective for providing medical specialists for underserved communities, particularly if these networks are underutilized by the patient population. Further analyses are needed to assess factors influencing utilization patterns of telemedicine services by underserved and ethnic communities before implementing these programs at clinical sites. PMID- 15298101 TI - A program to improve access to health care among Mexican immigrants in rural Colorado. AB - CONTEXT: Migration to the United States from Mexico is increasing every year. Mexican immigrants tend to be poor, uninsured, monolingual Spanish speakers without adequate access to appropriate medical care. As a further barrier, many are also undocumented. PURPOSE: This article describes a program developed to improve access to health care among Mexican immigrants in northern Colorado. METHODS: The program was implemented by a migrant/community health center in rural northern Colorado based on findings from an in-depth health needs survey of the target population. The program consists of community outreach services vertically integrated into the main medical clinics, which comprise Salud Family Health Centers. A mobile unit went to nontraditional areas identified by community workers as gathering places for Mexican immigrants. Services provided included preventive health care (screening for diabetes, hypertension, mental health problems, dental problems, and HIV); education; and primary care for acute problems. Patients were referred to a health care home for ongoing care. RESULTS: In the first 6 months, 1,553 Mexican immigrants were seen on the mobile unit. Hypertension and psychosocial problems were the most common problems in this population. Thirty-five percent of patients who received consultation in the mobile unit have visited any of the clinics for follow-up within the following year. CONCLUSIONS: A community-based mobile outreach program targeted toward Mexican immigrants can be effective in uncovering medical and mental illness and in directing patients to a health care home. This is an important first step in eliminating health disparities among this population. PMID- 15298102 TI - Meeting the health care needs of a rural Hispanic migrant population with diabetes. AB - CONTEXT: There is a need for models of health care that provide accessible, culturally appropriate, quality services to the population of Hispanic migrant farmworkers at risk for or diagnosed with diabetes. PURPOSES: The purposes of this study were to describe the Migrant Health Service, Inc (MHSI), Diabetes Program, the conceptual model on which it is based, and 4 types of outcomes achieved over a 3-year period. METHODS: Types and amounts of medical services and education were studied. Qualitative data obtained from program records and documents were analyzed to determine the nature of the program. Quantitative data were used to measure outcomes of the program. FINDINGS: The multiplecomponent MHSI Diabetes Program is addressing economic, cultural, and language barriers experienced by the target population. The program provides a continuum of health services and education that meet American Diabetes Association (ADA) Clinical Practice Recommendations on diabetes. The program exposes regional health care professionals and university students from numerous academic disciplines to Hispanic farmworker culture. CONCLUSIONS: Evidence-based program management, patient care, and program evaluation are traits of this program, which offers accessible, culturally appropriate, quality health services and education to Hispanic farmworkers. The multicomponent program model has high potential for positively impacting the health of the target population. PMID- 15298103 TI - Stress among migrant and seasonal farmworkers in rural southeast North Carolina. AB - CONTEXT: Although funding to enhance the delivery of health care among migrant farmworkers has primarily focused on urgent care issues within this population, the etiology of mental health risks and perceived stress is poorly understood. PURPOSE: To identify the type and severity of stress perceived by migrant and seasonal farmworkers in rural southeast North Carolina. METHODS: During the pre agricultural season in 2002, 151 migrant and seasonal farmworkers completed the 39-item Migrant Farmworker Stress Inventory (MFWSI) in either English or Spanish. FINDINGS: Fifty-one percent (n = 77) of the respondents perceived themselves at a high level of stress (mean score above 80 of "caseness") that may subsequently put them at greater risk for experiencing psychological difficulties. The stressors highly ranked (mean>2.5 in a maximum of 4, with "extremely stressful" in a 5-point Likert scale 0 to 4) were related to their mobile lifestyle, language barriers, insecure job and legal status, financial restraint, and long working hours. Also, drug and alcohol use in the migrant community was found to be one of the significant sources of stressors. Variables influencing high levels of stress include education, social support, religion, marital status, and age. Despite a relatively high level of perceived stress, the majority of respondents (71.5%) viewed their physical health as either "good" or "excellent." CONCLUSIONS: Findings from the study suggest the availability of social support systems may provide significant insight into developing appropriate health services for migrant and seasonal farmworkers and their families. PMID- 15298104 TI - Rural Mexican-American adolescent sexual risk behavior. AB - CONTEXT: There is a need for community-based, culturally sensitive, cognitive behavioral interventions to reduce sexual risk behavior among minority adolescents. Studies of adolescent risk and protective behaviors have focused on identifying modifiable psychosocial variables that predict differential outcomes for subsequent intervention efforts. Research has been scarce in studies of rural minority adolescent women. PURPOSE: To examine the protective and risk behaviors of these rural Mexican-American adolescent women and their relationship to physical or sexual abuse. METHODS: Mexican-American adolescent women aged 14-19 years were recruited through a rural health clinic and administered a self-report assessment for protective and risk behavior and sexual, physical, and psychological abuse. FINDINGS: Rural minority adolescent women endured high levels of psychological distress and many risk behaviors yet experienced few protective behaviors. Barriers to health care included access and confidentiality. Physically or sexually abused adolescents endured relatively greater risk and fewer protective behaviors than nonabused. CONCLUSIONS: Rural Mexican-American adolescent women may benefit from confidential identification and assessment of abuse history and risk and protective behaviors so that appropriate psychological treatment can accompany accessible medical treatment. The prevalence of risk behaviors and abuse among these women presents a need for development of behavioral interventions for risk reduction and promotion of health protective behaviors. PMID- 15298105 TI - "A 28-day program ain't helping the crack smoker"--perceptions of effective drug abuse prevention interventions by north central Florida African Americans who use cocaine. AB - CONTEXT: Cocaine is a major problem in the rural South, but knowledge is limited regarding the impact on African American populations. PURPOSE: This study of 18 39-year-old black drug users assessed perceptions of contributing factors to drug use and possible interventions. METHODS: The study design was qualitative descriptive, utilizing 4 focus groups with 5 rural women and 14 small-city residents. FINDINGS: Some respondents perceived that drug use initiation and continuation were due to themes of (1) loss, (2) peer pressure, (3) personal problems and dealing with pain, (4) desire for fun or to "feel good," and (5) drugs and drug-related messages within their environments. Common themes of effective strategies to stop drug use were (1) the necessity of wanting to quit, (2) the importance of help or support from family and friends, and (3) the need for resources, such as a job, car, and housing. Some respondents agreed on 3 human resources: (1) family, (2) ex-users, and (3) churches. Strategies to increase attendance at drug prevention programs included (1) making the program fun/enjoyable, (2) having mixed gender programs, (3) providing food/money, and (4) having the programs in their community. Recurrent themes were the lack of drug prevention intervention programs available to respondents and the failure of traditional programs of the majority culture to adequately meet their needs. CONCLUSION: Effective drug prevention programs for southern African Americans who use cocaine must be community based, personalized, and culturally relevant. PMID- 15298106 TI - Correlates of physician visits among children and adolescents in west Texas: effects of hyperglycemia symptoms. AB - CONTEXT: Health care services use by children varies tremendously. Because of the increasing prevalence of diabetes in children and adolscents, one of the major concerns is access to physician care among children with diabetes and diabetes symptoms. PURPOSE: This population-based cross-sectional study examines correlates of physician visit among children and adolescents living in west Texas. METHODS: A telephone survey was administered in 2002 to a random sample of households in 106 counties of West Texas. The sample included 5,462 respondents with children aged between 3 and 18 years. Proportional odds ordered logistic regression analysis was used to determine correlates of physician visits in the previous 12 months. FINDINGS: Hispanic children were less likely than non Hispanic whites to have a recent physician visit; there were no significant rural urban differences. Children with insurance (adjusted odds ratio= 2.21, 95% CI = 1.89-2.59) were more likely to visit physicians. Almost 16% of children in this study did not have any health insurance coverage. Children reporting 3 or more hyperglycemia symptoms and those with a family history of diabetes had 1.81 times and 1.20 times the odds of visiting the physician. CONCLUSIONS: Presence of health insurance and increasing symptoms of diabetes were found to influence the utilization of physician services. Since most of the cases of diabetes that have recently been diagnosed among Texas youth are type 2 diabetes, it is important that adolescents and their parents are educated about the risk factors and how to recognize them. PMID- 15298107 TI - The green gene revolution. PMID- 15298108 TI - The darkening earth. Less Sun at the Earth's surface complicates climate models. PMID- 15298110 TI - Anonymous trust. Making trusted computing work with privacy. PMID- 15298109 TI - Bumpy flying. Scalloped flippers of whales could reshape wings. PMID- 15298111 TI - Sloshing in space. Analyzing how liquids affect the motion of ships. PMID- 15298113 TI - A plan for water. A welcome federal strategy of ocean care has some worried nonetheless. PMID- 15298112 TI - Outsourcing drug work. Pharmaceuticals ship R&D and clinical trials to India. PMID- 15298114 TI - Middle of the country. As farming declines, rural America adapts to survive. PMID- 15298116 TI - Miracle on probability street. The law of large numbers guarantees that one-in-a million miracles happen 295 times a day in America. PMID- 15298115 TI - Penny-wise smart labels. If smart tags cost only one cent apiece, they would be everywhere. PMID- 15298117 TI - From finish to start. Was the grand challenge robot race in March the fiasco it appeared to be? Hardly, argues William "Red" Whittaker. The annual event is pushing mobile robotics to get real. PMID- 15298118 TI - Back to the future of cereals. Genomic studies of the world's major grain crops, together with a technology called marker-assisted breeding, could yield a new green revolution. PMID- 15298119 TI - Electrodynamic tethers in space. PMID- 15298120 TI - Virtual-reality therapy. PMID- 15298121 TI - Nuclear bunker buster bombs. PMID- 15298122 TI - Next stretch for plastic electronics. PMID- 15298123 TI - Questions that plague physics. Lawrence M. Krauss speaks about unfinished business. PMID- 15298124 TI - Arsenic crisis in Bangladesh. PMID- 15298125 TI - Medical imaging. Seeing inside. PMID- 15298126 TI - Crippled but not crashed. Neural networks can help pilots land damaged planes. PMID- 15298127 TI - What causes hiccups? PMID- 15298128 TI - How do sunless tanners work? PMID- 15298129 TI - An original appliance to correct posterior cross-bite. PMID- 15298130 TI - Facial changes in Puertorican patients as a result of 4 first bicuspid extraction orthodontic treatment using fixed mechanics. PMID- 15298131 TI - Dr. John W Witzig, father of functional orthodontics in North America. PMID- 15298132 TI - Simplicity in orthodontic treatment--part II. PMID- 15298133 TI - Opening the dental and skeletal bite in the straightwire appliance. PMID- 15298134 TI - Instrument care. Troubleshooting for the dental staff. PMID- 15298135 TI - Automated external defibrillators (AEDs). AB - Automated external defibrillators, or AEDs, will automatically analyze a patient's ECG and, if needed, deliver a defibrillating shock to the heart. The basic function of an AED is similar to that of a traditional defibrillator/monitor, but AEDs are much easier to use. As such, these devices can be used by a variety of medical and nonmedical personnel after only minimal training. The use of AEDs by nonmedical personnel (e.g., security guards, flight attendants) is often referred to as public access defibrillation, or PAD. In this Evaluation, we present our findings for eight newly evaluated models from five suppliers. We also summarize our findings for three previously evaluated models that are still on the market. We rate the models for first-responder use and for PAD use. Our ratings apply to the use of these devices both within the hospital and in the prehospital environment. PMID- 15298136 TI - Fatal air embolism caused by the misconnection of medical device hoses to needleless Luer ports on IV administration sets. PMID- 15298138 TI - Circuit-board failure prevents automated positioning of Medline (MC healthcare) electric bed. PMID- 15298137 TI - Uptime guarantees for PACS--calculating allowable downtime. PMID- 15298139 TI - Cystic fibrosis. AB - On a daily basis, pathologists examine the fundamental basis of human diseases using morphologic, immunologic, and molecular techniques. Cystic fibrosis (CF), as a clinically heterogeneous disease, exemplifies the complex challenges of genetic diseases for the pathologist who attempts to explain the mechanisms of disease and provide rationale for clinical management. This review includes an overview of CF and a discussion of pathophysiologic features and practical components of clinical and anatomic pathology, and concludes with a review of molecular diagnostics. PMID- 15298140 TI - New developments in prenatal screening for Down syndrome. AB - Since the introduction of the triple screen, the emergence of additional analytes found to be associated with trisomy 21 pregnancies has challenged the clinical pathology and obstetric communities to determine which combination provides the best performance characteristics as a screening test. The cost, practicality, and psychosocial and ethical dimensions of different screening regimens further complicate the issue. This review attempts to give an overview of the various markers, current methods of screening, and most promising recent developments in antenatal screening for Down syndrome, with an emphasis on performance in prospective studies that have emerged during the last few years, rather than on estimates from statistical modeling. PMID- 15298141 TI - Myelodysplastic syndromes. AB - Myelodysplastic syndromes are a group of clonal hematopoietic stem cell disorders in which there is ineffective hematopoiesis. Affected patients are primarily elderly people, but also at risk are certain subsets of patients who have been exposed to antineoplastic chemotherapy and select patients with primary bone marrow failure syndromes such as Fanconi anemia or aplastic anemia. The diagnosis of myelodysplastic syndrome implies a certain probability of leukemic evolution, but the magnitude of this probability varies greatly among patients. Over the years there have been several diagnostic and prognostic classifications of these disorders in an attempt to achieve diagnostic accuracy that is biologically and clinically meaningful. With the aging of the population and with improved survival among patients with cancer, pathologists are likely to confront myelodysplastic syndromes more frequently in their practice. This review discusses the clinical and pathologic manifestations of this group of disorders, major diagnostic and prognostic classification systems, pathogenesis, and select issues that might be clinically relevant in the future. PMID- 15298142 TI - The role of the clinical laboratory in the diagnosis of Cushing syndrome. AB - Endogenous Cushing syndrome (CS) is an uncommon disease in which patients often present with nonspecific initial symptoms that may affect many organ systems, with considerable morbidity and mortality. In the diagnosis of CS, the use of appropriate laboratory tests for the initial documentation of hypercortisolism as well as for distinguishing between adrenocorticotropic hormone dependent and independent forms are important for identifying the source and localizing the lesion. These tests are also essential for guiding the clinician to the correct surgical procedure to potentially cure the patient. PMID- 15298143 TI - Prion diseases: recent developments toward diagnostic tests. AB - Concerns about the large-scale European crisis of bovine spongiform encephalopathy, the finding of 1 infected cow in Alberta, Canada, in 2003, and the theoretical concern of prion transmissibility in blood have renewed interest in the development of rapid, minimally invasive diagnostic assays for prion diseases. Herein we review the pathologic features, clinical manifestations, diagnostic criteria, and recent developments that may lead to new diagnostic screening assays for prion diseases. PMID- 15298144 TI - HER-2/neu testing in breast cancer. AB - The testing of newly diagnosed breast cancer specimens for HER-2/neu status has achieved "standard of practice" status for the management of breast cancer in the United States. The discussion as to the best method to determine HER-2/neu status in these samples continues, with the fluorescence in situ hybridization method gaining popularity owing to the recent evidence that it, in comparison with immunohistochemical analysis, might more accurately predict clinical responses to trastuzumab-based therapies. With trastuzumab achieving excellent results in the treatment of HER-2/neu-positive advanced disease and under extensive evaluation in major clinical trials for its potential efficacy when used at earlier clinical stages, the potential role(s) for HER-2/neu testing as a predictor of response to other therapies being resolved by large prospective clinical outcome studies, and the more convenient gene-based chromogenic in situ hybridization technique "waiting in the wings," the saga of HER-2/neu testing in breast cancer will continue to unfold over the next several years. PMID- 15298145 TI - DNA ploidy and cell cycle analysis in breast cancer. AB - During the past 10 years there has been considerable interest in the application of new technologies to identify human malignancy and predict disease outcome. Markers of cell proliferation and the technologies of flow cytometry and image analysis for the determination of DNA total content in human tumor cells have been studied in breast cancer for 20 years. In this review, the uses and limitations of these technologies for the determination of ploidy status are discussed. This review also considers the prognostic significance and potential clinical utility of ploidy measurements, S phase calculation, and individual cell cycle regulatory biomarker expression levels. PMID- 15298146 TI - Prognostic factors in prostate cancer. AB - The ability of traditional and newer molecular-based prognostic factors to predict the outcome of prostate cancer is of considerable interest to urologists, pathologists, and patients. In this review, a series of traditional and newer molecular-based prognostic factors are considered, including those that have achieved widespread use, newer tests that are beginning to be used in clinical practice, and emerging molecular markers that have yet to be widely validated in the published literature or clinical trials. PMID- 15298147 TI - Malignant melanoma 2003: predisposition, diagnosis, prognosis, and staging. AB - The incidence and mortality of melanoma has seemed to level off for certain groups after a steady increase during the last 50 years. This trend is suspected to be secondary to education efforts aimed at prevention and better detection and removal of thin, biologically benign melanomas. Since no effective systemic therapies exist for metastatic melanoma, early detection and removal of thin melanomas offer the best chance of cure. For thicker melanomas, sentinel lymph node biopsy has improved the accuracy of staging and prognostic evaluation. However, approximately one third of patients diagnosed with metastatic melanomas present without previous regional lymph node metastases. As the genomic understanding of melanoma's pathogenesis grows, new methods likely will be developed to more accurately identify the people at risk for melanoma, those who have high-risk melanomas, and those who have disseminated disease. We review current and potential biomarkers useful for the screening for and prevention, diagnosis, staging, and prognosis of melanoma. PMID- 15298148 TI - A practical approach to selected problematic melanocytic lesions. AB - Pigmented skin lesions are among the most common skin lesions. A majority can be diagnosed with ease; however, a minority of the cases are difficult and have the potential for error with catastrophic consequences for patients and the potential for litigation. Melanoma-related claims are second only to breast biopsy-related claims in surgical pathology. This review tries to highlight the problematic areas and diagnostic pitfalls in melanocytic lesions with emphasis on melanomas that might be underdiagnosed and benign melanocytic lesions that might be overdiagnosed and on practical approaches to deal with these lesions. Scanning power examination only is insufficient to avoid the potential diagnostic pitfalls in melanocytic lesions. We recommend examination of every melanocytic lesion in at least several high-power fields for cytologic atypia and mitotic activity. When dealing with problematic cases, it is important to see the entire lesion to determine whether lateral circumscription, nevoid maturation, and deep mitotic activity are present. Immunohistochemical analysis is useful, but it cannot replace careful examination of H&E-stained sections. PMID- 15298149 TI - HER-2/neu evaluation in breast cancer are we there yet? AB - The HER-2/neu oncogene is located on chromosome 17q and encodes a transmembrane glycoprotein with intracellular tyrosine kinase activity. Several studies have shown an association of HER-2 gene amplification or protein overexpression with prognosis and predictor of therapeutic response. Most important, the presence of amplification or overexpression is the basis of eligibility for trastuzumab therapy. However, there are several methods of determining HER-2 status, each measuring a different aspect such as DNA content, gene copy number, protein expression, expression of RNA, and circulating HER-2 extracellular domain protein. There is no consensus with regard to the optimal test for HER-2 assessment. This review examines the various methods used in an attempt to define the most accurate and reliable test. The most widely used assays are immunohistochemical analysis and fluorescence in situ hybridization (FISH), which measure protein expression and gene amplification, respectively. Based on current data, FISH is the most accurate and reproducible test with a better correlation with prognosis and response to therapy. PMID- 15298150 TI - Current concepts on Riedel thyroiditis. AB - Riedel thyroiditis (RT) is an uncommon form of chronic thyroiditis in which the thyroid gland is replaced by fibrous tissue. The etiologic mechanisms underlying RT are unclear: the prevailing view is that it is part of a generalized fibroinflammatory process also involving other organs. The clinical manifestations of RT are protean, often resembling malignancy owing to goiter of remarkably hard consistency. Physical examination, laboratory analysis, cytology, and imaging features are not useful for differentiating between RT and neoplastic diseases or the fibrous variant of Hashimoto thyroiditis in the presurgical evaluation of patients. Histologic examination is necessary to establish the final diagnosis of RT. The present article reviews the most recent concepts about etiologic mechanisms, pathogenesis, diagnosis, and management of RT PMID- 15298151 TI - Iron and brain disorders. AB - Iron is the most important element in the body, essential for almost all types of cells, including brain cells. The role of iron in the brain has been known for years. Iron deficiency and iron excess have been associated with pathophysiology of different brain disorders. Iron deficiency has been reported to have a role in brain development and the pathophysiology of restless legs syndrome. Iron accumulation has been related to some neurologic disorders such as Alzheimer disease, Parkinson disease, type I neurodegeneration with brain iron accumulation, and other disorders. Despite years of investigation, the reason for iron imbalance in the brain is not known. It also is not known whether the accumulation of iron in the brain is primary or secondary to development of neurodegenerative disorders. This review summarizes the present knowledge on the role of iron in human brain disorders. PMID- 15298152 TI - Cat-scratch disease: historic, clinical, and pathologic perspectives. AB - Cat-scratch disease (CSD) initially was described in 1931, but the etiologic agent (Bartonella henselae) was not elucidated until decades later. This disease is the most common cause of chronic lymphadenopathy among children and adolescents, characteristically manifesting as subacute regional lymphadenitis with an associated inoculation site due to a cat scratch or bite, often accompanied by fever. The hallmark histologic lesion is granulomatous inflammation with a central stellate microabscess. Numerous atypical manifestations of CSD have been described, and these often lack the characteristic superficial lymphadenopathy and inoculation site papule. These atypical forms may be misdiagnosed initially as other infectious processes or neoplasms. We present a review of the history and epidemiologic features of CSD, describe common and unusual clinicopathologic manifestations, and discuss current diagnostic modalities. PMID- 15298153 TI - Hemolytic uremic syndrome revisited: Shiga toxin, factor H, and fibrin generation. AB - The hemolytic uremic syndrome (HUS) is a disease characterized by microangiopathic hemolytic anemia, thrombocytopenia, and renal failure. These features reflect the underlying histopathologic lesion: fibrin-rich thrombi that predominate in the renal microvasculature. HUS most commonly affects children younger than 5 years and is associated with Shiga toxin-producing enteric bacteria, the most important of which is Escherichia coli O157:H7. In this setting, HUS is epidemic and also might affect adults, particularly elderly people. Sporadic cases of HUS more commonly occur in adults and are associated with a wide variety of inciting agents and conditions. Although the disease manifestations might be similar and endothelial activation or injury likely represents a common etiologic event, differing responses to therapy suggest different pathogenic mechanisms. As more is understood about the underlying pathogenesis of the diseases that we now lump together as HUS, more efficacious and rational treatment and prevention strategies are likely to follow. PMID- 15298155 TI - Haptoglobin phenotypes in health and disorders. AB - Haptoglobin is a positive acute phase protein that binds free hemoglobin and removes it from the circulation to prevent kidney injury, and iron lossfollowing hemolysis. Also, by bindingfree hemoglobin, haptoglobin functions as an antioxidant. In addition, haptoglobin acts as a potent immunosuppressor of lymphocyte function and modulates the helper T-cell type 1 and type 2 (Th1/Th2) balance within the body. Three major haptoglobin phenotypes are known to exist (Hp 1-1, Hp 2-1, and Hp 2-2). Hp 1-1 is biologically the most effective in binding free hemoglobin and suppressing inflammatory responses associated with free hemoglobin. Hp 2-2 is biologically the least active, and Hp 2-1 is moderately active. The possible association of allelic polymorphism of haptoglobin with various pathologic conditions such as coronary artery disease has been studied. This article reviews the known functions of haptoglobin and the present understanding of a possible association of haptoglobin phenotypes with pathogenesis of a number of human disorders. PMID- 15298156 TI - Elevated serum potassium values: the role of preanalytic variables. AB - Potassium is one of the most frequently tested analytes in the clinical laboratory. Because of its critical role in body homeostasis, laboratory errors that cause inaccurate potassium results can significantly affect patient safety. This review is limited to the spurious increase of serum potassium levels (pseudohyperkalemia), ie, instances of elevated potassium results that cannot be explained clinically and do not correspond to the status of the patient; when these specimens are recollected, potassium values usually drop substantially without clinical intervention. Because the workup of falsely elevated potassium levels consumes valuable health care resources and can result in patient care delays, it is essential to identify all variables that can cause pseudohyperkalemia, understand the mechanisms by which these variables affect serum potassium levels, and define corrective actions to ameliorate the problem. In most cases, increases in serum potassium are due to factors in the preanalytic phase of the testing cycle. The effect of patient-specific variables and variables related to specimen acquisition and processing, handling, and transport are discussed. PMID- 15298154 TI - Thrombotic thrombocytopenic purpura: from platelet aggregates to plasma. AB - Thrombotic thrombocytopenic purpura (TTP) is a syndrome of severe thrombocytopenia and microangiopathic hemolytic anemia without an alternative explanation. Although some patients also have a combination of fever and neurologic and/or renal manifestations, these are not required for the diagnosis. Thus, plasmapheresis should start as soon as TTP is placed high in the differential diagnosis to prevent significant mortality. Histopathologically, TTP is characterized by widespread platelet thrombi in the microcirculation. Ultralarge von Willebrand factor (vWf) multimers found in the patient's plasma are the basis for the platelet thrombi. Recent evidence has linked the abnormal fragments of vWf with deficiency of a plasma enzyme named vWf-cleaving protease, or ADAMTS-13. While a small percentage of patients with TTP have a constitutional defect in this enzyme, many with the acute idiopathic form have an antibody to ADAMTS-13, affecting its ability to cleave vWf. The determination of the enzyme activity and the presence of its inhibitor have emerged as a potential tool in the diagnosis and prognosis of TTP. Furthermore, it helps to differentiate TTP from the hemolytic uremic syndrome, in which the level of ADAMTS-13 is expected to be normal or only slightly decreased. PMID- 15298157 TI - Clinical laboratory monitoring of coenzyme Q10 use in neurologic and muscular diseases. AB - Coenzyme Q10 (Q10) is available as an over-the-counter dietary supplement in the United States. While its use could be considered a form of alternative therapy, the medical profession has embraced the use of Q10 in specific disease states, including a series of neurologic and muscular diseases. Clinical laboratory monitoring is available for measurement of total Q10 in plasma and tissue and for measurement of redox status, ie, the ratio of reduced and oxidized forms of Q10. Many published studies have been anecdotal, in part owing to the rarity of some diseases involved. Unfortunately, many studies do not report Q10 levels, and, thus, the relationship of clinical response to Q10 concentration in plasma frequently is not discernible. Consistent laboratory monitoring of patients treated with this compound would help ease interpretation of the results of the treatment, especially because so many formulations of Q10 exist in the marketplace, each with its own bioavailability characteristics. Q10 has an enviable safety profile and, thus, is ideal to study as an adjunct to more conventional therapy. Defining patient subpopulations and characteristics that predict benefit from exogenous Q10 and defining therapeutic ranges for those particular applications are major challenges in this field. PMID- 15298158 TI - World report on violence and health. AB - Violence is not an intractable social problem or an inevitable part of the human condition. We can do much to address and prevent it. The world has not yet fully measured the size of the task and does not yet have all the tools to carry it out. But the global knowledge base is growing and much useful experience has already been gained. The World Report on Violence and Health attempts to contribute to that knowledge base. It is hoped that the report will inspire and facilitate increased cooperation, innovation and commitment to preventing violence around the world. These are the recommendations of the World Report on Violence and Health: Create, implement and monitor a national action plan for violence prevention. Enhance capacity for collecting data on violence. Define priorities for, and support research on the causes, consequences, costs and prevention of violence. Promote primary prevention responses. Strengthen responses for victims of violence. Integrate violence prevention into social and educational policies, and thereby promote gender and social equality. Increase collaboration and exchange of information on violence prevention. Promote and monitor adherence to international treaties, laws and other mechanisms to protect human rights. Seek practical, internationally-agreed responses to the global drugs trade and the global arms trade. PMID- 15298159 TI - The current status of hepatitis B in Lebanon. AB - The current status of hepatitis B infection in Lebanon is unknown due to the scarcity of published studies on the subject. This study was conducted to summarize the available information on hepatitis B in Lebanon since 1966, as well as to determine the current status of the problem, by analyzing the prevalence of positive hepatitis B surface antigen (HBsAg) reported from different laboratories of major hospitals covering the six districts of Lebanon in the year 2000. The overall HBsAg carrier rate among 61,271 tested individuals was 2.2%, being 13% among 30,809 blood donors, and 3.6% among 13,669 tested individuals in serology laboratories. There were marked geographical variations in the HBsAg carrier rate being 0.8% in Mount Lebanon, 1.9% in each of Bekaa and Greater Beirut, 2.2% in North Lebanon, 2.4% in Nabatiyeh, reaching up to 4.7% in South Lebanon. These findings are comparable to the previously reported studies on pregnant women and children, thus confirming that Lebanon is moderately endemic for hepatitis B. Such information stresses the urgent need for efficient national public health surveillance campaigns, and mass vaccination programs. In addition, the universal screening of pregnant women for HBsAg, and the implementation of universal newborn vaccination against hepatitis B virus (HBV) should be the standard of medical care for the control and eradication of HBV in Lebanon. PMID- 15298160 TI - Symptomatic antiepileptic drug associated vitamin D deficiency in noninstitutionalized patients: an under-diagnosed disorder. AB - Four noninstitutionalized patients, 4 months - 51 years old, presented out of 421 patients with epilepsy seen within a period of 2 years with serious symptoms of vitamin D deficiency secondary to chronic antiepileptic drug therapy. Presenting symptoms included exacerbation of seizure activity, status epilepticus, carpopedal spasms, fractures, osteomalacia, and rickets. All had low serum calcium and low vitamin D levels. Our experience supports the practice of screening patients on chronic antiepileptic drug therapy for vitamin D abnormalities. PMID- 15298161 TI - Methacholine challenge test: correlation with symptoms and atopy. AB - Methacholine challenge test (MCT) is a diagnostic test to assess airway hyperresponsiveness, one feature of asthma. Our study presents the results of MCT, done in our pulmonary function laboratory, correlated with patients' symptoms and allergic status. This is a cross-sectional study, where 134 patients were included : 60 patients had a PC-20 < or = 32 mg/ml (MCT+ group), while 74 had a PC-20 > 32 mg/ml (MCT- group). Dyspnea and wheezing were correlated to positive methacholine reactivity while cough was inversely correlated to it. In the 66 patients presenting isolated cough, 21 were MCT+ (32%). Methacholine reactivity was correlated to positive prick test, particularly for dust mite allergy (p = 0.006). There was a positive trend towards a higher incidence of dyspnea and wheezing with lower methacholine concentrations (p < 0.01), and lower incidence of cough with lower methacholine concentrations (p = 0.004). MCT was valuable in excluding the diagnosis of asthma in more than 50% of the cases and when positive, a good tool in guiding therapeutic choices. It showed a high correlation with the atopic status of the patient. It has definitely its place in diagnostic approach of respiratory symptoms. PMID- 15298162 TI - [Indications for therapeutic activity of cyclosporine]. AB - Ciclosporine was launched in therapeutic use in 1978. It marked the transplantation era due to the success of organ transplantation. Ciclosporine is actually used in bone marrow transplantation and in the transplantation of slid organs such as kidney, liver, heart, lungs, lung-heart, pancreas and bowel. It was proven also efficient in the treatment of auto-immune diseases especially in the treatment of psoriasis and nephrotic syndrom. Pharmacokinetic and drug interactions of ciclosporine have to be taken in consideration in order to allow an optimal use of the medication. However, the drug presents a long list of side effects: hypertension, increased level of serum creatinine, hypomagnesemia, severe gingivitis, maxillary modification, which can restrict the therapeutic use of the drug. PMID- 15298163 TI - [Prenatal diagnostic interpretations of complex/severe congenital cardiac malformations]. AB - Cardiac malformations occur the most within congenital anomalies. These malformations remain the leading cause of death in infancy related to severe and complex forms that appear earlier after birth. Actually, fetal echocardiography represents the best method for diagnosis of severe cardiac malformations. Depending upon several studies about congenital heart defects, the advantage is therefore the decrease of morbidity and mortality owing to pre- and postnatal interventions. PMID- 15298164 TI - Imaging of hepatic cystic lesions. AB - Cystic lesions of the liver are a common finding and the differential diagnosis is wider than often considered. Awareness of the radiologic key features allows a correct presumptive diagnosis in a lot of cases. Diagnostic accuracy is increased when combining the characteristic radiologic features to the clinical history and laboratory findings as summarized in Table I. Further useful information can be obtained in the most difficult cases with atypical findings by means of fine needle aspiration performed under imaging guidance. In endemic regions, hepatic echinococcal cyst should be always kept in mind and excluded from the differential diagnosis of HCL. PMID- 15298165 TI - Granulocytic sarcoma with spinal cord compression in chronic myelogenous leukemia: a case report. AB - We report on a 19-year-old man with a spinal cord compression secondary to granulocytic sarcoma (GS) as the initial presentation of a chronic myelogenous leukemia (CML). Blastic crisis developed two months later. According to our case report and to the literature, the diagnosis of GS could predict a rapid progression to blastic phase. PMID- 15298166 TI - Reverse genetics of mononegavirales. AB - "Reverse genetics" or de novo synthesis of nonsegmented negative-sense RNA viruses (Mononegavirales) from cloned cDNA has become a reliable technique to study this group of medically important viruses. Since the first generation of a negative-sense RNA virus entirely from cDNA in 1994, reverse genetics systems have been established for members of most genera of the Rhabdo-, Paramyxo-, and Filoviridae families. These systems are based on intracellular transcription of viral full-length RNAs and simultaneous expression of viral proteins required to form the typical viral ribonucleoprotein complex (RNP). These systems are powerful tools to study all aspects of the virus life cycle as well as the roles of virus proteins in virus-host interplay and pathogenicity. In addition, recombinant viruses can be designed to have specific properties that make them attractive as biotechnological tools and live vaccines. PMID- 15298167 TI - Reverse genetics systems for the generation of segmented negative-sense RNA viruses entirely from cloned cDNA. AB - Reverse genetics is defined as the generation of virus entirely from cloned cDNA. For negative-sense RNA viruses, whose genomes are complementary to mRNA in their orientation, the viral RNA(s) and the viral proteins required for replication and translation must be provided to initiate the viral replication cycle. Segmented negative-sense RNA viruses were refractory to genetic manipulation until 1989. In this chapter, we review developments in the reverse genetics of segmented negative-sense RNA viruses, beginning with the in vitro reconstitution of viral polymerase complexes in the late 1980s and culminating in the generation of Bunyamwera and influenza virus entirely from plasmid DNA almost a decade later. PMID- 15298168 TI - Transcription and replication of nonsegmented negative-strand RNA viruses. AB - The nonsegmented negative-strand (NNS) RNA viruses of the order Mononegavirales include a wide variety of human, animal, and plant pathogens. The NNS RNA genomes of these viruses are templates for two distinct RNA synthetic processes: transcription to generate mRNAs and replication of the genome via production of a positive-sense antigenome that acts as template to generate progeny negative strand genomes. The four virus families within the Mononegavirales all express the information encoded in their genomes by transcription of discrete subgenomic mRNAs. The key feature of transcriptional control in the NNS RNA viruses is entry of the virus-encoded RNA-dependent RNA polymerase at a single 3' proximal site followed by obligatory sequential transcription of the linear array of genes. Levels of gene expression are primarily regulated by position of each gene relative to the single promoter and also by cis-acting sequences located at the beginning and end of each gene and at the intergenic junctions. Obligatory sequential transcription dictates that termination of each upstream gene is required for initiation of downstream genes. Therefore, termination is a means to regulate expression of individual genes within the framework of a single transcriptional promoter. By engineering either whole virus genomes or subgenomic replicon derivatives, elements important for signaling transcript initiation, 5' end modification, 3' end polyadenylation, and transcription termination have been identified. Although the diverse families of NNS RNA virus use different sequences to control these processes, transcriptional termination is a common theme in controlling gene expression and overall transcriptional regulation is key in controlling the outcome of viral infection. The latest models for control of replication and transcription are discussed. PMID- 15298169 TI - Orthomyxovirus replication, transcription, and polyadenylation. AB - Efficient in vitro and in vivo systems are now in place to study the role of viral proteins in replication and/or transcription, the regulation of these processes, polyadenylation of viral mRNAs, the viral promoter structures, or the significance of noncoding regions for virus replication. In this chapter, we review the status of current knowledge of the orthomyxovirus RNA synthesis. PMID- 15298170 TI - Escaping from the cell: assembly and budding of negative-strand RNA viruses. AB - Negative-strand RNA virus particles are formed by a process that includes the assembly of viral components at the plasma membranes of infected cells and the subsequent release of particles by budding. Here, we review recent progress that has been made in understanding the mechanisms of negative-strand RNA virus assembly and bud- ding. Important topics for discussion include the key role played by the viral matrix proteins in assembly of viruses and viruslike particles, as well as roles played by additional viral components such as the viral glycoproteins. Various interactions that contribute to virus assembly are discussed, including interactions between matrix proteins and membranes, interactions between matrix proteins and glycoproteins, interactions between matrix proteins and nucleocapsids, and interactions that lead to matrix protein self-assembly. Selection of specific sites on plasma membranes to be used for virus assembly and budding is described, including the asymmetric budding of some viruses in polarized epithelial cells and assembly of viral components in lipid raft microdomains. Evidence for the involvement of cellular proteins in the late stages of rhabdovirus and filovirus budding is discussed as well as the possible involvement of similar host factors in the late stages of budding of other negative-strand RNA viruses. PMID- 15298171 TI - Accessory genes of the paramyxoviridae, a large family of nonsegmented negative strand RNA viruses, as a focus of active investigation by reverse genetics. AB - The Paramyxoviridae, a large family of nonsegmented negative-strand RNA viruses, comprises several genera each containing important human and animal pathogens. They possess in common six basal genes essential for viral replication and, in addition, a subset of accessory genes that are largely unique to each genus. These accessory genes are either encoded in one or more alternative overlapping frames of a basal gene, which are accessed transcriptionally or translationally, or inserted before or between the basal genes as one or more extra genes. However, the question of how the individual accessory genes contribute to actual viral replication and pathogenesis remained unanswered. It was not even established whether they are dispensable or indispensable for the viral life cycle. The plasmid-based reverse genetics of the full-length viral genome has now come into wide use to demonstrate that most, if not all, of these putative accessory genes can be disrupted without destroying viral infectivity, conclusively defining them as indeed dispensable accessory genes. Studies on the phenotypes of the resulting gene knockout viruses have revealed that the individual accessory genes greatly contribute specifically and additively to the overall viral fitness both in vitro and in vivo. PMID- 15298172 TI - Identification and characterization of viral antagonists of type I interferon in negative-strand RNA viruses. AB - Interferons are cytokines secreted in response to viral infections with potent antiviral activity, and they represent a critical component of the innate immune response against viruses. It has now become apparent that many viruses have evolved different mechanisms to counteract the interferon response, allowing their efficient replication and propagation in their hosts. This review discusses how the development of reverse genetics techniques and the increase in our knowledge of the interferon response have led to the discovery of interferon antagonistic functions of different genes of viruses belonging to the negative strand RNA virus group. In many cases, these viral genes encode accessory pro- teins that are not required for viral infectivity but are critical for optimal replication and for virulence in the host. PMID- 15298173 TI - Toward novel vaccines and therapies based on negative-strand RNA viruses. AB - The study of negative-strand RNA viruses has suggested new strategies to produce more attenuated viruses. Reverse genetics has allowed the implementation of the strategies, and new or improved monovalent vaccines are being developed. In addition, recombinant viruses expressing foreign proteins or epitopes have been produced with the aim of developing multivalent vaccines capable of stimulating humoral and cellular immune responses against more than one pathogen. Finally, recombinant viruses that selectively enter cells expressing tumor markers or the HIV envelope protein have been engineered and shown to lyse target cells. Preclinical and clinical trials of improved and multivalent vaccines and therapeutic (oncolytic) viruses are ongoing. PMID- 15298174 TI - Influenza vaccines generated by reverse genetics. AB - Influenza viruses cause annual epidemics and occasional pandemics of acute respiratory disease. Vaccination is the primary means to prevent and control the disease. However, influenza viruses undergo continual antigenic variation, which requires the annual reformulation of trivalent influenza vaccines, making influenza unique among pathogens for which vaccines have been developed. The segmented nature of the influenza virus genome allows for the traditional reassortment between two viruses in a coinfected cell. This technique has long been used to generate strains for the preparation of either inactivated or live attenuated influenza vaccines. Recent advancements in reverse genetics techniques now make it possible to generate influenza viruses entirely from cloned plasmid DNA by cotransfection of appropriate cells with 8 or 12 plasmids encoding the influenza virion sense RNA and/or mRNA. Once regulatory issues have been addressed, this technology will enable the routine and rapid generation of strains for either inactivated or live attenuated influenza vaccine. In addition, the technology offers the potential for new vaccine strategies based on the generation of genetically engineered donors attenuated through directed mutation of one or more internal genes. Reverse genetics techniques are also proving to be important for the development of pandemic influenza vaccines, because the technology provides a means to modify genes to remove virulence determinants found in highly pathogenic avian strains. The future of influenza prevention and control lies in the application of this powerful technology for the generation of safe and more effective influenza vaccines. PMID- 15298175 TI - The life and work of Josef Rudinger. AB - The life and work of Josef Rudinger (1924- 1975), a key figure in the history of peptide science, are outlined. PMID- 15298176 TI - Structure-activity relationship of an antibacterial peptide, maculatin 1.1, from the skin glands of the tree frog, Litoria genimaculata. AB - Maculatin 1.1 (Mac) is a cationic antibacterial peptide isolated from the dorsal glands of the tree frog, Litoria genimaculata, and has a sequence of GLFGVLAKVAAHVVPAIAEHF-NH2. A short peptide lacking the N-terminal two residues of Mac was reported to have no activity. To investigate the structure-activity relationship in detail, several analogs and related short peptides of Mac were synthesized. CD measurement showed that all the peptides took more or less an alpha-helical structure in the presence of anionic lipid vesicles. Analogs which are more basic than Mac had strong antibacterial and hemolytic activities, while short peptides lacking one or two terminal residues exhibited weak or no activity. Outer and inner membrane permeabilization activities of the peptides were also reduced with shortening of the peptide chain. These results indicate that the entire chain length of Mac is necessary for full activity, and the basicity of the peptides greatly affects the activity. PMID- 15298177 TI - Tat cell-penetrating peptide has the characteristics of a poly(proline) II helix in aqueous solution and in SDS micelles. AB - Tat cell-penetrating peptide (GRKKRRQRRRPPQG) is able to translocate and carry molecules across cell membranes. Using CD spectroscopy the conformation of this synthetic peptide was studied in aqueous and membrane-mimicking, micellar SDS solutions at different temperatures. The CD spectrum of the Tat cell-penetrating peptide in SDS micellar solution was virtually unchanged from that in aqueous solution, and at low temperature it was close to that of a poly(proline) II helix. PMID- 15298178 TI - Cell-physiological effects of elastin derived (VGVAPG)n oligomers in a unicellular model system. AB - Elastin is one of the most significant components of the extracellular matrix, which supports the stretchiness of the blood vessels via its helical structure and cross-links. Enzymatic decomposition of this protein could induce chemotactic responses of cell populations in the surrounding tissues by several peptide sequences, e.g. XGXXPG. In our present work the VGVAPG variant and its oligomers were studied. The objective of the experiments was to learn (i) whether the chemotactic effect of these peptides is general in different levels of phylogeny: (ii) whether increasing the number of monomer units influences the chemotactic behaviour of the cell? The trimer had the strongest chemoattractant effect in a wide concentration range (10(-12) - 10(-7) M), while the monomer and the pentamer were chemorepellent. All tri-, tetra-, penta- and hexamers could chemotactically select subpopulations with a high chemotactic responsiveness to the identical peptide, in the long term. With regard to its repellent effect, the pentamer had a negative effect on phagocytosis. All six oligomers had a growth-promoter effect in Tetrahymena. The characteristic cell-physiological effects of VGVAPG oligomers signal that molecules of the extracellular matrix can induce identical responses even in lower levels of phylogeny, e.g. in the Ciliates. PMID- 15298179 TI - Conformational and biochemical analysis of the cyclic peptides which modulate serine protease activity. AB - Prostate-specific antigen (PSA), a member of the kallikrein sub-group of the trypsin serine protease family, is a widely used marker for prostate cancer. Several sequences with specific binding to PSA have been identified by using phage display peptide libraries. The GST-fusion proteins of the characterized sequences have been shown to increase the enzyme activity of PSA to a synthetic substrate. The corresponding three cyclic synthetic analogues CVFTSNYAFC (A-1), CVFAHNYNYLVC (B-2) and CVAYCIEHHCWTC (C-4) have similar PSA promoting activity. Despite differences in the amino acid sequences, all three peptides bind to the same region of PSA. The conformation of the peptides was investigated by proton NMR spectroscopy. In addition, alanine replacement was used to characterize the prerequisites for binding. It is proposed that interactions with PSA are based on the aromatic and hydrophobic features of the amino acid side chains. Furthermore, it is suggested that peptides form beta-turn structures forced by cysteine bridges directing important aromatic side chains to the same side of the turn structure. PMID- 15298180 TI - Geometrical and conformational preferences of the 9-fluorenylmethoxycarbonyl amino moiety. AB - Structural parameters, originating from x-ray crystallographic data, have been compiled for 13 derivatives of amino acids, peptides and related compounds, which contain a total of 14 Fmoc-NH- moieties. For these moieties, molecular geometries and conformations--described by the omegao, theta1, theta2 and theta3' torsion angles--were analysed and compared with the corresponding parameters for the Z-NH and Boc-NH-moieties (290 and 553, respectively). To gain a deeper insight into the conformational features of the Fmoc-NH- moiety, ab initio free molecule calculations were performed for fully relaxed minima. Also the potential energy surface as a function of the torsion angles (theta3', theta2) was generated. The conformational features of the Fmoc-NH- moiety: (i) two possible values for the angle omegao (approximately 180 degrees or, rarely, approximately theta degrees) and (ii) the angle theta1 = 180 degrees +/- 15 degrees, are common to the Z-NH- and Boc-NH- systems. By contrast, the theta2 and theta3 angles in the Fmoc, Z and Boc groups differ essentially. In the Fmoc groups theta2 mostly has values of 180 degrees +/- 30 degrees and values up [115 degrees] seem to be forbidden, whereas fewer than half of the Z groups adopt theta2 approximately 180 degrees and the remainder have theta2 in the range of [90 degrees +/- 20 degrees]. On the other hand, the Boc methyl groups are staggered. The theta3 values observed for Fmoc are limited to the regions of 180 degrees +/- 20 degrees and 160 degrees +/- 20 degrees], while for the Z group a variety of theta3 occurs. The orientation of the fluorenyl vs the urethane function is mostly trans. Our results suggest a lower conformational flexibility for the Fmoc group compared with that of the Z group. Our calculations confirm that the observed conformational features for the Fmoc-NH- moiety are inherent properties. The Fmoc-NH-moiety in crystals involves the participation of its O=C-NH functionality in hydrogen bonds. PMID- 15298181 TI - Plant peptide hormone phytosulfokine (PSK-alpha): synthesis of new analogues and their biological evaluation. AB - Phytosulfokine-alpha (PSK-alpha), a sulfated growth factor (H-Tyr(SO3H)-Ile Tyr(SO3H)-Thr-Gln-OH) universally found in both monocotyledons and dicotyledons, strongly promotes proliferation of plant cells in culture. In our studies on structure/activity relationship in PSK-alpha the synthesis of a series of analogues was performed: [H-D-Tyr(SO3H)1]- (9), [H-Phe(4-SO3H)1]- (10), [H-D Phe(4-SO3H)1]- (11), [H-Phg(4-SO3H)1]- (12), [H-D-Phg(4-SO3H)1]- (13), H-Phe(4 NHSO2CH3)1]- (14), [H-D-Phe(4-NHSO2CH3)1]- (15), [H-Phe(4-NO2)1]- (16), [H-D Phe(4-NO2)1]- (17), [H-Phg(4-NO2)1]- (18), [H-D-Phg(4-NO2)1]- (19), [H-Hph(4 NO2)1]- (20), [H-Phg(4-OSO3H)1]- (21), [Phe(4-NO2)3]- (22), [Phg(4-NO2)3]- (23), [Hph(4-NO2)3]- (24), [H-Phe(4-SO3H)1, Phe(4-SO3H)3]- (25) [H-Phe(4-NO2)1, Phe(4 NO2)3]- (26), [H-Phg(4-NO2)1, Phg(4-NO2)3]- (27), [H-Hph(4-NO2)1, Hph(4-NO2)3]- (28) and [Val3]- PSK-alpha (29). For modification of the PSK-alpha peptide chain the novel amino acids and their derivatives were synthesized, such as: H-L-Phg(4 SO3H)-OH (1), H-D-Phg(4-SO3H)-OH (2), Fmoc-Phg(4-SO3H)-OH (3), Fmoc-D-Phg(4-SO3H) OH (4), Boc-Phg(4-NHSO2CH3)-OH (5), Boc-D-Phg(4-NHSO2CH3)-OH (6) Boc-Phe(4 NHSO2CH3)-OH (7), and Boc-D-Phe(4-NHSO2CH3)-OH (8). Peptides were synthesized by a solid phase method according to the Fmoc procedure on a Wang-resin. Free peptides were released from the resin by 95% TFA in the presence of EDT. All peptides were tested by competitive binding assay to the carrot membrane using 3H labelled PSK according to the Matsubayashi et al. test. PMID- 15298182 TI - Preparation and characterization of two LysB29 specifically labelled fluorescent derivatives of human insulin. AB - The preparation and characterization of two novel LysB29 selectively labelled fluorescent derivatives of human insulin are described. Two probes were chosen: 4 chloro-7-nitrobenz-2-oxa-1,3-diazole (NBD) and 7-methoxycoumarin-4-acetic acid (MCA), which have a relatively small, compact structure and are able to react with amino groups to form highly fluorescent derivatives. The combination of solid phase peptide synthesis and enzymatic semisynthesis was chosen for preparation of these fluorescent derivatives. Using two different protocols of solid-phase peptide synthesis, two fluorescent octapeptides were prepared corresponding to the position B23-B30 of human insulin, each with a different fluorescent label, NBD or MCA, on the epsilon-amino group of lysine. Then, the fluorescent octapeptides were coupled to desoctapeptide-(B23-B30)-insulin by a trypsin catalysed reaction. The receptor binding affinities of two novel fluorescent derivatives of human insulin with NBD and MCA (HI-NBD and HI-MCA) were determined on rat adipose tissue plasma membranes. Both fluorescent insulins, HI-NBD and HI-MCA, had only slightly reduced binding affinity and will be used for studying the interaction of insulin with its receptor. PMID- 15298183 TI - Small fish for healthy babies. PMID- 15298184 TI - Chromium and sediment toxicity. PMID- 15298185 TI - Adsorptive and absorptive contributions to the gas-particle partitioning of polycyclic aromatic hydrocarbons: state of knowledge and recommended parametrization for modeling. AB - Four contrasting descriptions of the gas-particle partitioning of SOCs are currently used: the Junge-Pankow adsorption model, the empirical Finizio organic matter (OM) absorption relationship, the Harner-Bidleman OM absorption model, and a dual black carbon (BC) adsorption and OM absorption model. Use of these four descriptions in a box model resulted in very different global fates, particularly for PAHs such as chrysene and benzo[a]pyrene. By reviewing published gas-particle distributions of PAHs, we found evidence for both absorptive and adsorptive contributions. Based on results from laboratory and controlled field studies we suggest that on average, octanol-air partitioning (Koa) is a good approximation for the OM absorption of PAHs. However, higher concentrations in particles than could be explained by OM absorption were found in selected gas-particle partitioning field studies, which were corrected for gaseous adsorption to the filter. We argue that adsorption onto BC is responsible for most of the additional sorption. Apparent adsorption coefficients to BC, K(BC-air), were derived from field studies and showed good agreement with those predicted by adsorption onto diesel soot. For atmospheric long-range transport models we suggestthe use of a dual OM absorption and BC adsorption model, with BC properties being approximated by diesel soot: Kp = 10(-12) (f(om) 1/rho(oct) Koa + fBC/rho(BC) K(soot-air) a(atm-BC)/a(soot)). We hypothesize that kinetic constraints related to shell-like particle structures might lead to deviations from sorption equilibrium and higher particle-borne fractions of PAHs in particular at remote sites. PMID- 15298186 TI - Polybrominated diphenyl ethers and organochlorines in archived northern fur seal samples from the Pacific coast of Japan, 1972-1998. AB - The present study clearly indicated possible prolonged and chronic marine pollution by polybrominated diphenyl ethers (PBDEs) unless the international regulatory measures are reinforced substantially. Fat tissues of female northern fur seals (Callorhinus ursinus), collected at 10 time periods between 1972 and 1998 from the Pacific coast of northern Japan, were analyzed for PBDEs and organochlorine compounds (OCs). PCBs and DDTs were the predominant contaminants in the fur seal samples. PBDEs were detected in all the samples analyzed, the values ranging from 0.33 to 100 ng/g lipid wt. with predominance of BDE-47. The lowest PBDE levels were in the fur seals collected in 1972, with the peak concentration around 1991-1994 and then decreased to about 50% in 1997-1998. Compositions of higher brominated congeners increased since 1972, while some lower brominated congeners decreased, implying a change in the pattern of use, particularly the increased use of highly brominated diphenyl ethers during 1972 1998. In the meantime, PCB compositions in fur seals showed no temporal variation, suggesting a continuous input of PCBs into the marine environment in significant quantities. As peak concentrations of PBDEs occurred later than OCs, it is essential to follow up the patterns of PBDEs pollution that may be of great concern in the future. To our knowledge, this is the first report on the contamination by PBDEs in marine mammals from the Asia-Pacific waters. PMID- 15298187 TI - Physical and kinetic speciation of copper and zinc in three geochemically contrasting marine estuaries. AB - The physical and kinetic speciation of Cu and Zn in three impacted marine estuaries was examined. Contrasts in sources of metal-binding ligands, solution chemistry, and hydrologic forcing between and withinthethree study systems (Cape Fear River Estuary, North Carolina; Norfolk-Hampton Roads-Elizabeth River, Virginia; San Diego Bay, California) were exploited to enhance our understanding of Cu and Zn speciation. Trace metal-optimized tangential-flow ultrafiltration at 1 kDa nominal molecular weight limit (NMWL) was used to fractionate <0.4 microm species into colloidal and "dissolved" pools. Colloidal species of dissolved organic matter (DOM) and copper were significant and often the dominant pools in each of the three study systems. Characteristic colloidal fractions of both DOM and Cu ranged from near 70% of <0.4 microm concentrations in Cape Fear to 50% in San Diego Bay. Colloidal Cu and DOM were strongly coupled, and variability in observed <0.4 microm Cu concentrations was closely related to the concentrations of colloidal-associated metal. Colloidal fractions were much smaller for Zn than that of Cu; ranging from 10-30% in Cape Fear to less than 5% in San Diego Bay, and no relationship to DOM was observed. Kinetic separations on Chelex resin revealed the presence of large nonlabile pools of Cu in each of the study systems, with the highest fractions (70-100%) in Cape Fear and Norfolk and lowest (30-50%) in San Diego Bay. A close relationship was observed between colloidal and nonlabile Cu species, implying slow reactivity of colloidal-bound Cu. The fraction of filterable Zn labile to Chelex averaged 97%, 85%, and 60% in San Diego, Norfolk, and Cape Fear, respectively. Anthropogenic Zn appeared almost exclusively in the <1 kDa fraction, while anthropogenic Cu was distributed between dissolved and colloidal pools. Copper particle-partition coefficients (Kd) followed the trend: San Diego >> Norfolk > Cape Fear and were inversely correlated with DOC concentrations. Colloid-based partition coefficients were significantly greater, in many cases an order of magnitude greater, than particle based partition coefficients. The partitioning data suggest the presence of metal enriched bacterial-derived exudates and/or discrete metal phases in colloidal sized particles in impacted regions of these estuaries. The strong relationships observed between Cu and DOC indicate that Cu partitioning behavior over a range of estuarine environments may be modeled effectively with a limited set of coefficients. Our measurements of metal lability and size distribution imply that the fraction of <0.4 microm Zn that is likely to be bioavailable is greater than that for Cu, especially in impacted regions of the study systems. PMID- 15298188 TI - Influence of an industrial waste incinerator as assessed by the levels and congener patterns of polychlorinated dibenzo-p-dioxins and polychlorinated dibenzofurans. AB - To assess the spatial change in the influence of an industrial waste incinerator, a total of 47 soil samples (in continuous manner with distance) and 65 human blood samples (40 within 5 km and 20 at 7 and 12 km) were analyzed for polychlorinated dibenzo-p-dioxins/polychlorinated dibenzofurans (PCDD/Fs). The influence was not clearly observed both on the soil and blood levels of PCDD/Fs as the levels in the near zone (within 5 km) were not statistically different from those in the far zones at 7 and 12 km. Assessment was conducted on the congener patterns by using principal component analysis and by characterizing the congener fractions as a function of distance. In soil, the congener fractions of 2,3,4,7,8-PeCDF, 1,2,3,4,7,8-HxCDF, 1,2,3,6,7,8-HxCDF, 1,2,3,4,6,7,8-HpCDF, and OCDF decreased progressively with the distance. Blood was consistent with soil in that each congener fraction of these PCDFs (except OCDF) was statistically greater in the near zone than the far zones. The decreases in these PCDFs were balanced by OCDD in both soil and blood. It was concluded that although not obviously observed in the contamination levels, the influence of the incinerator was clearly shown by the congener patterns of PCDD/Fs in both soil and blood that changed with distance. PMID- 15298189 TI - Simultaneous assessment of sources, processes, and factors influencing herbicide losses to surface waters in a small agricultural catchment. AB - To take appropriate measures to minimize agricultural herbicide inputs into surface waters, detailed knowledge is required about all the factors that control the losses of a given compound from point sources (i.e., farmyards) as well as from the diffuse sources (i.e., the fields) within a given catchment. In this and in a companion paper, we present the results of a comprehensive field study, in which the temporal and spatial variability of the losses of three herbicides (atrazine, dimethenamid, and metolachlor) into the surface waters within a small catchment (2.1 km2) were investigated on different scales (i.e., field scale to whole catchment) after a controlled application of the compounds. In this paper, we discuss the loss dynamics of the three herbicides (and some of their metabolites) from the whole catchment over a period of 67 d after application. An identical mixture of the three herbicides was applied on 13 cornfields within 12 h, allowing for a comparison of their losses under identical meteorological conditions. Thanks to a high temporal sampling resolution, it was possible to distinguish between losses from a farmyard and losses from the fields. Farmyard losses contributed less than 20% to the total loads but caused the highest concentrations. The major herbicide losses from the agricultural fields occurred during the first two rain events after application that led to significant surface runoff and preferential flow into tile drains. In the soils of all fields, dimethenamid declined somewhat faster than atrazine and metolachlor, whereas atrazine was mobilized most effectively to runoff water. Relative losses of the three compounds did not vary by more than a factor of 3 (0.82, 0.27, and 0.41% of the mass applied for atrazine, dimethenamid, and metolachlor, respectively). Highest peak concentrations at the outlet of the catchment were found for atrazine (i.e., approximately 8 microg L(-1) for a short period (<2 h) due to point source losses and between 1 and 3.5 microg L(-1) during more than 24 h due to diffuse losses). PMID- 15298190 TI - Variability of herbicide losses from 13 fields to surface water within a small catchment after a controlled herbicide application. AB - Diffuse losses from agricultural fields are a major input source for herbicides in surface waters. In this and in a companion paper, we present the results of a comprehensive field study aimed at assessing the overall loss dynamics of three model herbicides (i.e., atrazine, dimethenamid, and metolachlor) from a small agricultural catchment (2.1 km2) and evaluating the relative contributions of various fields having different soil and topographical characteristics. An identical mixture of the three model herbicides as well as an additional pesticide for identification of a given field were applied within 12 h on 13 cornfields (total area approximately 12 ha), thus ensuring that the herbicides were exposed to identical meteorological conditions. After the simultaneous application, the concentrations of the compounds were monitored in the soils and at the outlets of three subcatchments containing between 4 and 5 cornfields each. Particular emphasis was placed on the two rain events that led to the major losses of the herbicides. The rank orders of herbicide dissipation in the soils and of the compound-specific mobilization into runoff were the same in all three subcatchments and were independent of the field characteristics. In contrast, the field properties caused the relative losses from two subcatchments to differ by up to a factor of 56 during the most important event, whereas compound-specific differences of the three neutral herbicides caused the losses to vary only by a factor of 2 during the same event. The enormous spatial variability was mainly caused by factors influencing the fraction of rain that was lost to surface water by fast transport mechanisms. Thus, the key factors determining the spatially variable herbicide losses were the permeability of the soils, the topography, and the location of subsurface drainage systems. These results illustrate the large potential to reduce herbicide losses by avoiding application on risk areas. PMID- 15298191 TI - Characterizing dependence of pesticide load in surface water on precipitation and pesticide use for the Sacramento River watershed. AB - Transport of pesticides by surface runoff during rainfall events is a major process contributing to pesticide contamination in rivers. This study presents an empirical regression model that relates pesticide loading over time in the Sacramento River with the precipitation and pesticide use in the Sacramento River watershed. The model closely simulated loading dynamics of diazinon, simazine, and diuron during 1991-1994 and 1997-2000 winter storm seasons. The coefficients of determination for regression ranged from 0.168 to 0.907, all were significant at <0.001. The results of this study provide strong evidence that precipitation and pesticide use are the two major environmental variables dictating the dynamics of pesticide transport into surface water in a watershed. The capability of the statistical model to provide time-series estimates on pesticide loading in rivers is unique and may be useful fortotal maximum daily load (TMDL) assessments. PMID- 15298192 TI - Levels and toxicokinetic behaviors of PCDD, PCDF, and coplanar PCB congeners in common cormorants from Lake Biwa, Japan. AB - Concentrations of polychlorinated dibenzo-p-dioxins (PCDDs), polychlorinated dibenzofurans (PCDFs), and coplanar polychlorinated biphenyls (Co-PCBs) were determined in the liver and pectoral muscle of common cormorants (Phalacrocorax carbo) collected from Lake Biwa, Japan. To clarify the toxicokinetic behaviors and potential toxicities of these chemicals, the present study addresses life stage- and tissue-specific accumulation of the congeners. Total 2,3,7,8 tetrachlorodibenzo-p-dioxin toxic equivalents (TEQs) were in the range of 360 to 50,000 pg/g lipid weight in the liver and 310 to 12,000 pg/g lipid weight in the pectoral muscle. Among congeners, for which toxic equivalency factors were assigned, PCB126, 2,3,4,7,8-P5CDF, and 1,2,3,7,8-P5CDD made a greater contribution to total TEQs in the liver. Hepatic concentrations of T4- to H6CDDs, P5- and H6CDFs, and Co-PCBs (except PCB77) significantly increased with growth of cormorants, leading to life-stage-related compositional changes. The concentration ratios of liver to pectoral muscle revealed preferential accumulation of higher chlorinated congeners in hepatic tissue. For most congeners, concentration ratios significantly increased with an increase in hepatic total TEQs, suggesting their concentration-dependent hepatic sequestration. These results imply the presence of hepatic binding protein(s) such as cytochrome P450, inducible by these chemicals, which mayfunction as a binding species different from aryl hydrocarbon receptor. On the basis of these results, we conclude that the toxicokinetic behavior of each congener is life stage-, tissue-, and concentration-dependent. TEQs in wildlife populations exposed to multiple congeners with varying concentrations should be used with caution for risk assessment, even within a species. PMID- 15298193 TI - Maternal and fetal mercury and n-3 polyunsaturated fatty acids as a risk and benefit of fish consumption to fetus. AB - Maternal fish consumption brings both risks and benefits to the fetus from the standpoint of methylmercury (MeHg) and n-3 PUFA (polyunsaturated fatty acids). MeHg is one of the most risky substances to come through fish consumption, and mercury concentrations in red blood cells (RBC-Hg) are the best biomarker of MeHg exposure. Docosahexaenoic acid (DHA, C22:6n-3), which is one of the most important fatty acids for normal brain development and function, is also derived from fish consumption. Our objective in this study was to examine the relationships between RBC-Hg and plasma fatty acid composition in mother and fetus at parturition. Venous blood samples were collected from 63 pairs of mothers and fetuses (umbilical cord blood) at delivery. In all cases, fetal RBC Hg levels were higher than maternal RBC-Hg levels. The geometric mean of fetal RBC-Hg was 13.4 ng/g, which was significantly (p < 0.01) higher than that of maternal RBC-Hg (8.41 ng/g). While the average fetal/maternal RBC-Hg ratio was 1.6, the individual ratios varied from 1.08 to 2.19, suggesting considerable individual differences in MeHg concentrations between maternal and fetal circulations at delivery. A significant correlation was observed between maternal and fetal DHA concentrations (r = 0.37, p < 0.01). Further, a significant correlation was observed between RBC-Hg and plasma DHA in fetus (r = 0.35, p < 0.01). These results confirm that both MeHg and DHA which originated from fish consumption transferred from maternal to fetal circulation and existed in the fetal circulation with a positive correlation. Pregnant women in particular need not give up eating fish to obtain such benefits. However, they would do well to at least consume smaller fish, which contain less MeHg, thereby balancing the risks and benefits from fish comsumption. PMID- 15298194 TI - Quantification of bacterial chemotaxis in porous media using magnetic resonance imaging. AB - Bacterial chemotaxis has the potential to enhance biodegradation of organic contaminants in polluted groundwater systems. However, studies of bacterial chemotaxis in porous media are scarce. In this study we use magnetic resonance imaging (MRI) for the noninvasive measurement of changes in bacterial-density distributions in a packed column at a spatial resolution of 330 microm as a function of time. We analyze both the diffusive and the chemotactic behavior of Pseudomonas putida F1 in the presence of the chemical stimulus trichloroethylene (TCE). The migration of motile bacteria in experiments without TCE was described using an effective motility coefficient, whereas the presence of TCE required addition of a nonzero chemotactic sensitivity coefficient, indicating a significant response to TCE. The need for a chemotactic sensitivity term was justified by a test for statistical significance. This study represents the first quantification of bacterial chemotactic parameters within a packed column. For conditions under which chemotaxis occurs in porous media, it may potentially be exploited to significantly improve rates of in situ pollutant biodegradation in the subsurface environment, particularlyfor pollutants dissolved in water trapped in low-permeability formations or lenses. PMID- 15298195 TI - Fractionation of stable isotope-labeled organic pollutants as a potential tracer of atmospheric transport processes. AB - To test the potential for using stable isotope fractionation to examine the atmospheric transport of semivolatile organic compounds (SOCs), we conducted simplified distillation experiments in the laboratory and a tracer-release experiment using mixtures of stable isotope-labeled (D and 13C) and unlabeled SOCs. Perdeuterated phenanthrene and alpha-hexachlorocyclohexane were transported more slowly via air-water gas exchange in our laboratory experiments, resulting in significant isotope fractionation of perdeuterated/unlabeled compound mixtures. In contrast, isotope fractionation of 13C6-labeled SOCs was much lower. A field tracer-release study was then conducted by spiking a seawater retention pond with a mixture of D10-labeled, 13C2-labeled, and unlabeled phenanthrene and examining isotope fractionation of the mixture after air-water gas exchange. No preferential fractionation of D10-vs 13C2-labeled phenanthrene was observed in the pond water; however, greater fractionation of D10-vs 13C2-labeled phenanthrene was observed in air samples collected within a 1-100 m radius of the pond. Thus, stable isotope tracers may provide a means of examining the atmospheric transport and air-earth exchange rates of POPs in an environmental realistic setting. PMID- 15298196 TI - Role of speciation in organotin toxicity to the yeast Candida maltosa. AB - Assessment of organometal pollution requires an understanding of the various processes that influence the bioavailability and toxicity of the contaminant. Organotins may exist as both cationic species and neutral hydroxides in aqueous solution, with the formation of chloride species in the presence of Cl-. Although these species have different chemical properties, there is very little information on the influence of speciation on organotin and microbial cell interactions. Tributyltin (TBT) and triphenyltin (TPT) interactions with the yeast Candida maltosa were investigated between pH 3.5 and 7.5 and in up to 0.5 M NaCl at pH 5.5. Toxicity increased with both pH and NaCl concentration and the mechanisms of interaction depended on the species present in solution. TBT and TPT interacted by different mechanisms, as evidenced by action on membrane fluidity. Furthermore, there was a strong correlation between toxicity and overall octanol-water distribution ratio (D(OW)) of organotin compounds. Triorganotin cations are less toxic than triorganotin hydroxides, which are in turn less toxic than triorganotin chlorides. These findings underline the importance of speciation effects on organotin interactions in the environment. PMID- 15298197 TI - On the origin of the optical properties of humic substances. AB - Absorption and fluorescence spectroscopy and laser photobleaching experiments were employed to probe the origins of the optical properties of humic substances (HS). Luminescence quantum yields and the wavelengths of maximum emission were acquired for Suwannee River humic acid (SRHA) and fulvic acid (SRFA) at an extensive series of excitation wavelengths across the ultraviolet and visible. Laser irradiation at a series wavelength across the ultraviolet and visible was further employed to destroy selectively chromophores absorbing at specific wavelengths, using absorption spectroscopy to follow the absorption losses with irradiation time. The results provide unequivocal evidence that the absorption and emission spectra of these materials cannot result solely from a simple linear superposition of the spectra of numerous independent chromophores. Instead, the long wavelength absorption tail of HS (>350 nm) appears to arise from a continuum of coupled states. We propose that this behavior results from intramolecular charge-transfer interactions between hydroxy-aromatic donors and quinoid acceptors formed by the partial oxidation of lignin precursors. We further propose that these donor-acceptor interactions may be a common phenomenon, occurring within all natural hydroxy- or polyhydroxy-aromatic polymers that form appropriate acceptors upon partial oxidation. Examples of such species include lignin, polyphenols, tannins, and melanins. PMID- 15298198 TI - Exchange of polychlorinated biphenyls (PCBs) and polychlorinated naphthalenes (PCNs) between air and a mixed pasture sward. AB - To improve understanding of air-to-vegetation transfer of persistent organic pollutants (POPs), uptake and depuration of polychlorinated biphenyls (PCBs) and polychlorinated naphthalenes (PCNs) between grass sward and air was investigated. Pasture swards were placed in fanned (2 m s(-1) wind speed) and unfanned conditions for a period of 20 days and sampled at intervals. Depuration was carried out after a short (4 days) and a long (14 days) exposure period. Prior to contamination, a mixed pasture sward at a semi-rural location contained sigmaPCN concentrations 15-20% of the sigmaPCB concentration. Uptake of both PCBs and PCNs was broadly linear in fanned and unfanned conditions over the 20-day period, i.e., the pasture did not reach equilibrium with the air. Uptake rates (fluxes) were greater under the fanned conditions. The difference in uptake rates between fanned and unfanned conditions increased with degree of chlorination for both PCBs and PCNs, ranging between a factor of 2 for tri-chlorinated PCBs and PCNs and a factor 5 for octa-chlorinated PCBs. Depuration results over the first hours were very scattered, showing an initial period of loss, followed by an increase in concentrations, possibly as a result of re-volatilization of PCBs from the soil in the trays, with consequent recapture by the overlying sward. Rapid clearance was observed over the following days, but depuration of PCBs and PCNs was still incomplete after 14 days, with 20% of the initial concentration of the sigmaPCBs and 10% of the sigmaPCNs retained by the sward. There was no difference in the proportion of POPs retained in the sward between the 4- and 14-day contamination treatments. POP-specific differences in the amount of compound "trapped" in leaves after contamination were observed. The results show that, although changes in the rate of air movement around a pasture have an effect on the uptake rate of POPs into the vegetation, plant-side resistance controls both the air-to-pasture and pasture-to-air exchange of gas-phase PCBs and PCNs; i.e., differences between plant species in cuticle composition and/or structure affecting the permeability of the cuticle are of greater importance than differences in leaf morphology affecting aerodynamic roughness. PMID- 15298199 TI - Rapid release of mercury from intertidal sediments exposed to solar radiation: a field experiment. AB - There is increasing evidence of the primary importance of photochemical reactions and transfer of gaseous mercury to the atmosphere. Although mercury in aquatic sediments is efficiently retained, resuspension and bioturbation in intertidal sediments may expose temporarily anoxic sediments to solar radiation. Field experiments were performed to investigate these processes. Anoxic sediments from two areas in the Tagus estuary with different degrees of Hg contamination (experiments I and II) were homogenized and distributed into two sets of 36 uncovered Petri dishes. The samples were placed on the intertidal sediments and exposed to direct solar radiation and kept under dark (control) for 6-8 h. The decrease rates of acid volatile sulfides (abrupt in the first 3 h) and of pyrite (linear) were the same in sediments under solar radiation and dark. The total Hg concentrations were relatively constant in sediments kept in dark, but decreased from 17.6 to 7.65 and 3.45 to 1.35 nmol g(-1) in experiments I and II, respectively. In those exposed to solar radiation during the period of higher UV intensity. Similar evolutions were found in nonreactive Hg in pore waters (3.00 2.59 and 0.725-0.105 nM). On the contrary, reactive Hg was higher in pore waters of the sediments exposed to solar radiation and increased with time, from 424 to 845 pM and 53 to 193 pM. These results indicate that most mercury released in pore waters was photochemically reduced in a short period of time and escaped rapidly to the atmosphere. Episodes of bottom resuspension and bioturbation in the intertidal sediments enhance the transfer of gaseous mercury to the atmosphere. PMID- 15298200 TI - Role of dissolved organic matter, nitrate, and bicarbonate in the photolysis of aqueous fipronil. AB - A multivariate kinetic model of aqueous fipronil photodegradation was developed as a function of dissolved organic matter (DOM), bicarbonate, and nitrate at concentrations that bracketthose commonly observed in natural waters (ca. 0-10 mg/L). Several pathways were available for fipronil photodegradation in this system, including direct photolysis and indirect photooxidation by species produced during the illumination of natural waters (e.g., 3NOM*, 1O2*, *OH, *CO3(1-), *OOR, *OOH, e(aq)-, O2(*-)). Product studies indicated thatfipronil was quantitatively converted to fipronil desulfinyl, a product that is associated with direct photolysis alone. DOM was the only variable that affected fipronil degradation; it decreased the rate of fipronil photodegradation primarily through competitive light absorption (i.e., attenuation) and the quenching of fipronil*. The addition of sodium chloride (30 percent per thousand) resulted in a more rapid rate (approximately 20%) of fipronil loss in comparison to equivalent experiments performed without sodium chloride, implying that fipronil may be more photolabile in marine environments. PMID- 15298201 TI - Experimental visualization of solute transport and mass transfer processes in two dimensional conductivity fields with connected regions of high conductivity. AB - Solute transport displaying mass transfer behavior (i.e., tailing) occurs in many aquifers and soils. Spatial patterns of hydraulic conductivity may play a role because of both advection and diffusion through isolated low conductivity areas. We demonstrated such processes in laboratory experiments designed to visualize solute transport through a thin chamber (40 cm x 20 cm x 0.64 cm thick) packed with glass beads and containing circular emplacements of smaller glass beads with lower conductivity. The experiments used three different contrasts of conductivity between the large-bead matrix and the emplacements, targeting three different regimes of solute transport: low contrast, targeting macrodispersion; intermediate contrast, targeting advection-dominated mass transfer between the high-conductivity regions and the emplacements; and high contrast, targeting diffusion-dominated mass transfer. Use of a strong light source, a high resolution CCD camera, and a colorimetric dye produced images with a spatial resolution of about 400 microm and a concentration range of approximately 2 orders of magnitude. These images confirm the existence of the three different regimes, and we observed tailing driven by both advection and diffusion. Outflow concentration measured by spectrophotometer achieved 3 orders of magnitude in concentration range and showed good agreement with known models in the case of dispersion and diffusive mass transfer, with estimated parameters close to a priori predictions. Existing models for diffusive mass transfer did notfitthe breakthrough curves from the intermediate-contrast chamber, but a model of slow advection through cylinders did. Thus, both breakthrough curves and chamber images confirm that different contrasts in small-scale K lead to different regimes of solute transport and thus require different models of upscaled solute transport. PMID- 15298202 TI - Contribution of natural organic matter to copper leaching from municipal solid waste incinerator bottom ash. AB - The leaching of heavy metals, such as copper, from municipal solid waste incinerator (MSWI) bottom ash is a concern in many countries and may inhibit the beneficial reuse of this secondary material. The enhanced leaching of copper from three MSWI bottom ash samples by dissolved organic carbon (DOC) was investigated with specific attention for the nature of the organic ligands. A competitive ligand exchange-solvent extraction (CLE-SE) method was used to measure Cu binding to DOC. Two types of binding sites for Cu were identified and geochemical modeling showed that the organically bound fraction varied from 82% to 100% between pH 6.6 and 10.6. Model calculations showed that complexation by previously identified aliphatic and aromatic acids was unable to explain the enhanced Cu leaching from the MSWI residues. High-performance size-exclusion chromatography (HPSEC) and the standard extraction procedure to isolate and purify natural organic matter revealed that about 0.5% of DOC consists of humic acids and 14.3-25.6% consists of fulvic acids. Calculated Cu binding isotherms based on these natural organic compounds, and the nonideal competitive adsorption Donnan (NICA-Donnan) model, provide an adequate description of the organic Cu complexation in the bottom ash leachates. The results show that fulvic acid-type components exist in MSWI bottom ash leachates and are likely responsible for the generally observed enhanced Cu leaching from these residues. These findings enable the use of geochemical speciation programs, which include models and intrinsic parameters for metal binding to natural organic matter, to predict Cu leaching from this widely produced waste material under variable environmental conditions (e.g., pH, ionic strength, and concentrations of competing metals). The identified role of fulvic acids in the leaching of Cu and possibly other heavy metals can also be used in the development of techniques to improve the environmental quality of MSWI bottom ash. PMID- 15298203 TI - Photochemical fate of sulfa drugs in the aquatic environment: sulfa drugs containing five-membered heterocyclic groups. AB - The photochemical fate of five sulfa drugs with varying five-membered heterocyclic substituents (sulfamethoxazole, sulfisoxazole, sulfamethizole, sulfathiazole, and sulfamoxole) was investigated in aqueous solution. The rate of direct photolysis of these compounds is dependent upon the identity of the heterocyclic R group as well as the pH of the solution. Matrix deconvolution methods were employed to determine the absorption spectrum and photolysis rate of each protonation state (cationic, neutral, and anionic). From these data, quantum yields for direct photodegradation were calculated for each protonation state of the sulfa drugs. The quantum yields calculated range from <0.005 for the neutral state of sulfamethizole to 0.7 +/- 0.3 for the protonated state of sulfisoxazole. The protonation state that is most photoreactive varies among the sulfa drugs and cannot be attributed to the rate of photon absorption. Products arising from the direct photolysis of the sulfa drugs were also investigated. For all the compounds, sulfanilic acid was observed as a common product. The singlet oxygen quenching rates of the sulfa drugs were determined by laser flash photolysis and range from (2 +/- 1) x 10(4) M(-1) s(-1) for sulfamethoxazole to (3.0 +/- 0.7) x 10(8) M(-1) s(-1) for sulfamoxole. Reaction of the sulfa drugs with hydroxyl radical is not modulated by the R group, and the rate constants are all near the bimolecular diffusion-controlled limit of 10(10) M(-1) s(-1). The photodegradation of the sulfa drugs in natural water samples of Lake Josephine (St. Paul, MN) and Lake Superior was attributed solely to direct photolysis. This study indicates thatthese similarly structured antibiotics will be subject to a wide range of photodegradation rates with sulfathiazole degrading relatively quickly, sulfisoxazole and sulfamethizole degrading moderately, and sulfamethoxazole degrading much more slowly. PMID- 15298204 TI - Thermodynamic properties of multifunctional oxygenates in atmospheric aerosols from quantum mechanics and molecular dynamics: dicarboxylic acids. AB - Ambient particulate matter contains polar multifunctional oxygenates that partition between the vapor and aerosol phases. Vapor pressure predictions are required to determine the gas-particle partitioning of such organic compounds. We present here a method based on atomistic simulations combined with the Clausius Clapeyron equation to predict the liquid vapor pressure, enthalpies of vaporization, and heats of sublimation of atmospheric organic compounds. The resulting temperature-dependent vapor pressure equation is a function of the heat of vaporization at the normal boiling point [deltaHvap(Tb)], normal boiling point (Tb), and the change in heat capacity (liquid to gas) of the compound upon phase change [deltaCp(Tb)]. We show that heats of vaporization can be estimated from calculated cohesive energy densities (CED) of the pure compound obtained from multiple sampling molecular dynamics. The simulation method (CED) uses a generic force field (Dreiding) and molecular models with atomic charges determined from quantum mechanics. The heats of vaporization of five dicarboxylic acids [malonic (C3), succinic (C4), glutaric (C5), adipic (C6), and pimelic (C7)] are calculated at 500 K. Results are in agreement with experimental values with an averaged error of about 4%. The corresponding heats of sublimation at 298 K are also predicted using molecular simulations. Vapor pressures of the five dicarboxylic acids are also predicted using the derived Clausius-Clapeyron equation. Predicted liquid vapor pressures agree well with available literature data with an averaged error of 29%, while the predicted solid vapor pressures at ambient temperature differ considerably from a recent study by Bilde et al. (Environ. Sci. Technol. 2003, 37, 1371-1378) (an average of 70%). The difference is attributed to the linear dependence assumption thatwe used in the derived Clausius-Clapeyron equation. PMID- 15298205 TI - Estimation of diffusion coefficient of chromium in colloidal silica using digital photography. AB - In order to study the effectiveness of using colloidal silica, NYACOL DP5110, to stabilize chromium-contaminated soil, the diffusion of chromium in colloidal silica gel was estimated from laboratory experiments. To measure diffusion coefficients of chromium in the colloidal silica gel, a new measurement method based on digital photography was introduced. A series of experiments were designed and conducted to validate this new method and to estimate the diffusion coefficients of chromium in the colloidal silica gel. Accuracy of the proposed method was evaluated by several differentways. It was found that the apparent diffusion coefficient of chromium in colloidal silica gel ranged from 1.76 to 8.48 x 10(-10) m2/s depending mainly on the concentration of silica in the gel with chromium concentration less than 10(-2) M. Higher silica concentrations yielded lower diffusion coefficients due to the obstruction to the free movement of chromium. The adsorption isotherm of chromate to colloidal silica gel was found to be linear at pH 7; the partition coefficient was calculated to be 0.549 L/g. Mass balance calculations were performed to evaluate the accuracy of the proposed method and found that the measuring error was less than 6.5%. Based on the test data, the estimation of diffusion coefficients for chromium in colloidal silica gel using digital photography seems to be accurate and precise. This method is suitable for analyzing colored chemicals inside clear/white gels. From the results, it can be concluded that the gel behaves as a porous material with silica network forming continuous solid phase and its pore space saturated with water. The chromium ions diffuse in porous silica gel on a tortuous path. Therefore, the bulk diffusion dominates. Thus, the silica can be represented as a fix and impenetrable immersion in the solution. The presence of these motionless silica chains leads to an increase in the mean path of the diffusing molecules between two points in the system. On the basis of the test results, it can also be concluded that colloidal silica, NYACOL DP5110, for in-situ treatment of chromium-contaminated soils seems to be ineffective. Further research of more realistic simulation of diffusion and refined gel formulation with the capacity to convert the chromium to an immobile form is recommended. PMID- 15298206 TI - Optimizing detection limits for the analysis of petroleum hydrocarbons in complex environmental samples. AB - To evaluate the sources, transport, bioremediation, fate, and effects of spilled petroleum and petroleum products, environmental studies often measure parent and alkylated polycyclic aromatic hydrocarbons (PAH), alkanes, and chemical biomarkers (e.g., triterpanes). Accurate data for low analyte concentrations are required when environmental samples contain hydrocarbons from multiple sources that need to be resolved and quantified. The accuracy and usefulness of the analyses can be improved by lowering the method detection limits (MDLs) for these compounds. Misidentification of hydrocarbon source can result when the MDLs are too high. Modifications to standard analytical methods (i.e., U.S. Environmental Protection Agency Method 8270) can lower MDLs by factors ranging from 10 to 1000. This reduction has important implications for ecological-risk assessments. Modifications having the greatest impact on the MDL include GCMS analysis in the selected-ion-monitoring mode (SIM), increased sample size, column cleanup of the extract, and decreased preinjection volume (volume of final extract prior to injection into instrument). In one study in which a benthic sediment sample was spiked with low levels of topped (heated to remove more volatile PAH that are naturally enriched in crude oil) Alaska North Slope crude, MDLs for individual PAH analytes and biomarkers were determined to be less than 0.5 ng/g (ppb) dry weight and less than 5 ppb dry weightfor individual alkanes. Similar results were obtained when the sediment was spiked with the 16 EPA priority pollutants. In addition, a method has been developed to estimate MDLs for source-specific alkylated PAH analytes and chemical biomarker compounds for which standards are not commercially available or are prohibitively expensive. These improved analytical techniques have been used to identify and quantify low levels of hydrocarbons, derived from both natural and anthropogenic sources, found in the benthic sediments of Prince William Sound, AK. PMID- 15298207 TI - Potential contamination of shipboard air samples by diffusive emissions of PCBs and other organic pollutants: implications and solutions. AB - Air samples were taken onboard the RRS Bransfield on an Atlantic cruise from the United Kingdom to Halley, Antarctica, from October to December 1998, with the aim of establishing PCB oceanic background air concentrations and assessing their latitudinal distribution. Great care was taken to minimize pre- and post collection contamination of the samples, which was validated through stringent QA/QC procedures. However, there is evidence that onboard contamination of the air samples occurred,following insidious, diffusive emissions on the ship. Other data (for PCBs and other persistent organic pollutants (POPs)) and examples of shipboard contamination are presented. The implications of these findings for past and future studies of global POPs distribution are discussed. Recommendations are made to help critically appraise and minimize the problems of insidious/diffusive shipboard contamination. PMID- 15298208 TI - Aesthetics of simulated soiling patterns on architecture. AB - Two desk-top exercises investigated public acceptability of idealized soiling patterns on buildings, using methodologies typical of the psychology of art. The exercises used a range of simulated soiling patterns around a simple architectural element, a pedimented window. In the first experiment respondents were asked to arrange the images from the "most acceptable" pattern to the "least acceptable". Results hinted at the importance of certain soiling features in driving the ranking. The second exercise explored the characteristics of soiling patterns that most affected their acceptability. In this experiment the images were organized in pairs. People were requested to choose the pattern they found more acceptable in each pair. Uniform patterns and those which created shadowing effects proved more acceptable. Patterns with non-integer fractal dimension that obscured architectural forms were less acceptable. There was usually a preference for images showing less soiling, whereas vertical features and lumpiness were not as acceptable. Results gave an insight into spatial factors that might influence the acceptability of soiling on real buildings. Thus suggesting it is necessary to consider both the level and the distribution of soiling when trying to gauge public reaction. PMID- 15298209 TI - Comparison between acetic acid and landfill leachates for the leaching of Pb(II), Cd(II), As(V), and Cr(VI) from cementitious wastes. AB - The Toxicity Characteristic Leaching Procedure (TCLP) has been widely used to characterize the suitability of solid wastes for disposal in landfills. However, the widespread application of this test for the assessment of wastes disposed in different landfill types is often questionable. This paper investigates the leaching profiles of cement-stabilized heavy metal ions, namely, Pb (II), Cd (II), As(V), and Cr(VI), using acetic acid and leachates from municipal and nonputrescible Australian landfill sites. The leaching profiles of Pb, Cd, As, and Cr using acetic acid were found to be similar to the nonputrescible landfill leachate and differed markedly from the municipal solid waste (MSW) leachate. The additional presence of high amounts of organic and inorganic compounds in the municipal landfill leachate influenced the leaching profiles of these metal ions as compared to the acetic acid and the nonputrescible systems. It is postulated that the organic compounds present in the municipal landfill leachate formed complexes with the Pb and Cd, increasing the mobility of these ions. Moreover, the organic compounds in the municipal landfill leachate induced a reducing environment in the leachate, causing the reduction of Cr(VI) to Cr(III). It was also found that the presence of carbonates in the municipal landfill leachate affected the stability of calcium arsenate, with the carbonate competing with arsenate for calcium at high pH, forcing arsenate into the solution. PMID- 15298210 TI - Formate ion decomposition in water under UV irradiation at 253.7 nm. AB - Formate ion (HCO2-) occurs in natural waters as a result of photooxidation of humic substances. Under UV irradiation, as applied in water purification (253.7 nm), formate ion decomposed following split-rate pseudo-zero-order kinetics (k1 and k2 are initial and final rate constants, respectively). In the presence of dissolved oxygen (DO), it was found that (a) k1 < k2, (b) k1 and k2 increased with initial formate ion concentration ([HCO2-]0 = (1.73-38.3) x 10(-5) mol L( 1)) and absorbed UV intensity (Ia = (1.38-3.99) x 10(-6) mol quanta L(-1) s(-1)), and (c) k1 and k2 were relatively insensitive to initial pH (pHo = 5.41-8.97) in buffer-free solutions. Both rate constants decreased with increasing carbonate alkalinity ((0-1.0) x 10(-3) mol L(-1)) and k1 was virtually unchanged in phosphate buffer at pH0 between 5.25 and 9.92. Carbonate buffer lowered the rate of formate ion decay, possibly due to scavenging of OH* radicals. Initial rate constant k1 slightly increased with temperature (15-35 degrees C), while k2 remained unchanged. The reaction pH increased rapidly during irradiation of buffer-free NaHCO2 solution to approach an equilibrium level as [HCO2-] reached the method detection level (MDL). The pH profile of buffer-free formate ion decay was estimated using closed-system equilibrium analysis. DO utilization during UV irradiation was 0.5 mol of O2/mol of HCO2-, while nonpurgeable organic carbon (NPOC) measurements on kinetic samples closely followed the HCO2- profile, thus strongly suggesting the transformation of HCO2- -C to CO2 in the presence of DO. In DO-free water, k1 > k2 was observed. Furthermore, k(1,DO FREE) > k(1,DO) (k(1,DO) = k1) and k(2,DO FREE) < k(2,DO) (k(2,DO) = k2). The effect of dual acid solutions on HCO2- decay was examined in a mixture of NaHCO2 and sodium oxalate (Na2C2O4). HCO2- decomposed readily until [HCO2-] approximately equal to MDL but at a lower rate than in buffer-free HCO2- solutions, while C2O4(2-) remained virtually unchanged. C2O4(2-) decay commenced following near complete conversion of HCO2-. PMID- 15298211 TI - Radiation chemistry of methyl tert-butyl ether in aqueous solution. AB - The chemical kinetics of the free-radical-induced degradation of the gasoline oxygenate methyl tert-butyl ether (MTBE) in water have been investigated. Rate constants for the reaction of MTBE with the hydroxyl radical, hydrated electron, and hydrogen atom were determined in aqueous solution at room temperature, using electron pulse radiolysis and absorption spectroscopy (*OH and e- aq) and EPR free induction decay attenuation (*H) measurements. The rate constant for hydroxyl radical reaction of (1.71 +/- 0.02) x 10(9) M(-1) s(-1) showed that the oxidative process was the dominant pathway, relative to MTBE reaction with hydrogen atoms, (3.49 +/- 0.06) x 10(6) M(-1) s(-1), or hydrated electrons, <8.0 x 10(6) M(-1) s(-1). The hydroxyl radical reaction gives a transient carbon centered radical which subsequently reacts with dissolved oxygen to form peroxyl radicals, the rate constant for this reaction was (2.17 +/- 0.06) x 10(9) M(-1) s(-1). The second-order decay of the MTBE peroxyl radical was 2k = (6.0 +/- 0.3) x 10(8) M(-1) s(-1). These rate constants, along with preliminary MTBE degradation product distribution measurements, were incorporated into a kinetic model that compared the predicted MTBE removal from water against experimental measurements performed under large-scale electron beam treatment conditions. PMID- 15298212 TI - In situ metal precipitation in a zinc-contaminated, aerobic sandy aquifer by means of biological sulfate reduction. AB - The applicability of in situ metal precipitation (ISMP) based on bacterial sulfate reduction (BSR) with molasses as carbon source was tested for the immobilization of a zinc plume in an aquifer with highly unsuitable initial conditions (high Eh, low pH, low organic matter content, and low sulfate concentrations), using deep wells for substrate injection. Batch experiments revealed an optimal molasses concentration range of 1-5 g/L and demonstrated the necessity of adding a specific growth medium to the groundwater. Without this growth medium, even sulfate, nitrogen, phosphorus, and potassium addition combined with pH optimization could not trigger biological sulfate reduction. In column experiments, precipitation of ZnS(s) was induced biologically as well as chemically (by adding Na2S). In both systems, zinc concentrations dropped from about 30 mg/L to below 0.02 mg/L. After termination of substrate addition the biological system showed continuation of BSR for at least 2 months, suggesting the insensitivity of the sulfate reducing system for short stagnations of nutrient supply, whereas in the chemical system an immediate increase of Zn concentrations was observed. A pilot experiment conducted in situ at the zinc contaminated site showed a reduction of zinc concentrations from around 40 mg/L to below 0.01 mg/L. Termination of substrate supply did not result in an immediate stagnation of the BSR process, but continuation of BSR was observed for at least 5 weeks. PMID- 15298213 TI - Removal of chlorophenols from wastewater using red mud: an aluminum industry waste. AB - Removal of toxic substances from wastewaters using low-cost alternatives to activated carbon is an important area in environmental sciences. Efforts have been made to convert red mud, an aluminum industry waste, into a low-cost potential adsorbent, and the final material has been used for the removal of phenol, 2-chlorophenol, 4-chlorophenol, and 2,4-dichlorophenol from wastewater. It is pertinent to mention that phenol and chlorophenols are highly carcinogenic and an priority class of pollutants which needs to be removed from effluents before discharge into water bodies. 2,4-Dichlorophenol and 4-chlorophenol are sorbed by the developed adsorbent up to 94-97%, while the removal of 2 chlorophenol and phenol was up to 50-81%. The removal of phenols and its derivatives was achieved up to 98% by column experiments at a flow rate of 0.5 mL/min. The order of removal was 2,4-dichlorophenol > 4-chlorophenol > 2 chlorophenol > phenol, and the removal takes place through a particle diffusion mechanism. The adsorption was found to be endothermic in nature and follows both Langmuir and Freundlich models. Estimation of the phenols was carried out by capillary electrophoresis, and the adsorbent has been successfully tried for the removal of chlorophenols from a wastewater. The developed process is very useful, economic, rapid, and reproducible for the removal of phenols. PMID- 15298214 TI - Transformation of aromatic ether- and amine-containing pharmaceuticals during chlorine disinfection. AB - Many of the human pharmaceuticals detected in municipal wastewater effluent, surface water, and groundwater contain functional groups that could undergo transformation reactions during chlorine disinfection. To assess the potential importance of these reactions to the environmental fate of pharmaceuticals, the rate of transformation of a group of compounds was measured over a pH range of 5 10. Several of the pharmaceuticals reacted rapidly with free chlorine (i.e., HOCl/OCl-) and would be expected to undergo transformation under the conditions typically encountered in many chlorine disinfection systems. For compounds containing aromatic ether functional groups, the rate of transformation was strongly affected bythe other substituents on the ring. The amine-containing pharmaceuticals underwent a rapid reaction with hypochlorous acid to form chlorinated amines, which could be converted back into the parent compound by reaction with thiosulfate. In the absence of thiosulfate, the chlorinated amines slowly decomposed to form species that could not be converted back into the parent compound. The reaction rates of the pharmaceuticals with combined chlorine (i.e., chloramines) were significantly slower, and transformation of the compounds would not be expected under the conditions encountered during chloramination. PMID- 15298215 TI - Effect of platinum deposits on TiO2 on the anoxic photocatalytic degradation pathways of alkylamines in water: dealkylation and N-alkyation. AB - Most photocatalytic degradation (PCD) reactions of aquatic pollutants require the presence of dissolved oxygen and hence do not occur in anoxic suspensions. We investigated the PCD reactions of alkylamines in anoxic water using TiO2 deposited with Pt nanoparticles. Unlike typical PCD reactions, the absence of dissolved oxygen increases the PCD rates of alkylamines on Pt/TiO2 and generates products that are different from those formed on pure TiO2. In particular, N alkylated amines (e.g., (CH3)3N produced from (CH3)2NH) as well as dealkylated amines are generated in a deaerated Pt/TiO2 suspension. This anoxic N-alkylation pathway is enabled only in the presence of Pt deposits on TiO2 and is applicable only to neutral alkylamines and not to alkylammonium cations. The Pt surface appears to interactwith the lone-pair electron on the N atom and catalyze the anoxic degradation of alkylamines mainly through a radical mechanism. Methyl radicals generated on Pt participate in the N-methylation reaction. The presence of intermediate methyl radicals on the Pt surface was verified by the detection of CH4 and CH3CH3 gases evolved during the PCD of (CH3)3N in an anoxic Pt/TiO2 suspension, whereas no such products were observed in a pure TiO2 suspension. The anoxic PCD of N-methylethylamine on Pt/TiO2 also produces both N-ethylated and N methylated amines as byproducts, which indicates that both methyl and ethyl radicals are generated during the anoxic degradation process. From a practical point of view, the present finding that undesirable alkylated amines can be produced on Pt/TiO2 in anoxic conditions indicates that caution is necessary when applying Pt/TiO2 photocatalyst to the treatment of water that contains amines. PMID- 15298216 TI - Green chemistry methods in sulfur dyeing: application of various reducing D sugars and analysis of the importance of optimum redox potential. AB - The importance of sulfur dyeing of cellulosic fibers, particularly cotton, is realized economically throughout the dyeing industry. At the present time, dyeing with sulfur dyes requires the use of various auxiliaries, many of which have adverse effects on the environment. The most damaging of these is the reducing agent sodium sulfide, required to reduce the dye molecules to a water-soluble leuco form to enable adsorption and diffusion into the fiber. In this study, attempts have been made to replace the sodium sulfide used within the sulfur dyeing process with a variety of environmentally friendly reducing sugars. The redox potential of various hexose and pentose monosaccharides and reducing disaccharides was recorded and compared. Subsequently, cotton was dyed with the world's most important sulfur dye, C. I. Sulfur Black 1, using the reducing sugars under alkaline conditions, and compared to dyeings secured by employing commercial sulfide reducing agents. It was observed that reducing sugars gave comparable, and in many cases superior, color strength and wash fastness results, with respect to the commercial sulfide-based reducing agents, which still account for the vast majority of sulfur dyeing processes and that pose significant environmental concern. Employment of reducing sugars in sulfur dyeing could provide a sustainable, nontoxic, biodegradable, cost-effective alternative to sodium polysulfide and sodium hydrogen sulfide. Comparison of the redox potential of reducing sugars against the color strength of the dyeings secured demonstrated that there was an optimum redox potential of around -650 mV for maximum color strength achieved. The same redox potential also conferred the lowest color loss upon washing. These observations were attributed to reduction of the polymeric dye molecules to an optimum size for fiber affinity and diffusion into the fiber, but which would also confer maximum wash fastness upon oxidation. PMID- 15298217 TI - Electricity generation using an air-cathode single chamber microbial fuel cell in the presence and absence of a proton exchange membrane. AB - Microbial fuel cells (MFCs) are typically designed as a two-chamber system with the bacteria in the anode chamber separated from the cathode chamber by a polymeric proton exchange membrane (PEM). Most MFCs use aqueous cathodes where water is bubbled with air to provide dissolved oxygen to electrode. To increase energy output and reduce the cost of MFCs, we examined power generation in an air cathode MFC containing carbon electrodes in the presence and absence of a polymeric proton exchange membrane (PEM). Bacteria present in domestic wastewater were used as the biocatalyst, and glucose and wastewater were tested as substrates. Power density was found to be much greater than typically reported for aqueous-cathode MFCs, reaching a maximum of 262 +/- 10 mW/m2 (6.6 +/- 0.3 mW/L; liquid volume) using glucose. Removing the PEM increased the maximum power density to 494 +/- 21 mW/m2 (12.5 +/- 0.5 mW/L). Coulombic efficiency was 40-55% with the PEM and 9-12% with the PEM removed, indicating substantial oxygen diffusion into the anode chamber in the absence of the PEM. Power output increased with glucose concentration according to saturation-type kinetics, with a half saturation constant of 79 mg/L with the PEM-MFC and 103 mg/L in the MFC without a PEM (1000 omega resistor). Similar results on the effect of the PEM on power density were found using wastewater, where 28 +/- 3 mW/m2 (0.7 +/- 0.1 mW/L) (28% Coulombic efficiency) was produced with the PEM, and 146 +/- 8 mW/m2 (3.7 +/- 0.2 mW/L) (20% Coulombic efficiency) was produced when the PEM was removed. The increase in power output when a PEM was removed was attributed to a higher cathode potential as shown by an increase in the open circuit potential. An analysis based on available anode surface area and maximum bacterial growth rates suggests that mediatorless MFCs may have an upper order-of-magnitude limit in power density of 10(3) mW/m2. A cost-effective approach to achieving power densities in this range will likely require systems that do not contain a polymeric PEM in the MFC and systems based on direct oxygen transfer to a carbon cathode. PMID- 15298218 TI - Comment on "sources and variations of mercury in tuna". PMID- 15298219 TI - Creating healthier societies. PMID- 15298220 TI - Registering clinical trials is necessary for ethical, scientific and economic reasons. PMID- 15298221 TI - Working across sectors for public health. PMID- 15298222 TI - Signs of illness in Kenyan infants aged less than 60 days. AB - OBJECTIVE: Little data has been published on the presenting symptoms and signs among ill infants aged <60 days from developing countries. We aimed to describe and evaluate the potential of simple clinical features to identify severe illness among young infants who present to rural district hospitals in Kenya. METHODS: Standardized assessment tools were designed to record clinical symptoms and signs. Data were collected prospectively on all infants aged <60 days who weighed > or = 1.5 kg and were admitted over an 18-month period. The same data were collected, prospectively from infants recruited to a contemporaneous hospital birth cohort who became ill and were assessed and treated as outpatients at the same hospital. FINDINGS: Data on 467 outpatient consultations and 769 inpatient episodes were available for analysis. These data highlighted the importance of findings in the history, particularly breathing difficulties, abnormal feeding, and abnormal behaviour, as well as clinical signs in the evaluation of young infants. They indicated possible important differences in the panel of signs useful for detecting severe illness in infants aged 0-6 days and those aged 7-59 days. They also showed that some simplification of current guidelines that still preserved the sensitivity and specificity for detecting very severe disease might be possible. CONCLUSION: Simple clinical features may allow distinction between severe and non-severe illness to be made with reasonable confidence. Prospective studies on an adequate scale are needed urgently to provide current integrated management of childhood illness guidelines for young infants with an adequate evidence base. PMID- 15298223 TI - Sexual violence against intimate partners in Cape Town: prevalence and risk factors reported by men. AB - OBJECTIVE: To describe the prevalence of and risk factors for the perpetration of sexual violence by men against female intimate partners. METHODS: A cross sectional study was conducted on 1368 randomly selected men working in three Cape Town municipalities. The men were interviewed with the aid of a questionnaire on current sexual partners in the preceding 10 years, personal and relationship characteristics and the use of violence against their partners. RESULTS: The perpetration of sexual violence against intimate partners in the past 10 years was reported by 15.3% of the men. After adjustment for sociodemographic circumstances, the factors associated with such violence were involvement in physical conflict outside the home, problematic alcohol use, having more than one current partner and abusing partners verbally. While having frequent conflict with partners was important for the risk of sexual violence, only two types of conflict sources were significantly associated with this risk, namely conflict over sexual refusal and conflict when men perceived their authority to be undermined. CONCLUSION: Sexual violence in intimate relations was common. The risk of being sexually violent was associated with the use of violence to solve problems in other settings, having more than one current partner, alcohol abuse and verbally abusing a partner. It was also associated with particular types of conflict stemming from ideas of male sexual entitlement and dominance. Prevention programmes that focus on gender relations and non-violent conflict resolution for men and youths may be useful in combating such sexual violence. PMID- 15298224 TI - Cost-effectiveness analysis of cataract surgery: a global and regional analysis. AB - OBJECTIVE: To estimate the population health effects, costs and cost effectiveness of selected cataract surgery interventions in areas of the world with different epidemiological profiles. METHODS: Effectiveness estimates are based on a review of the literature taking into account factors such as operative failure, complications and patient non-compliance. A population model was applied to follow the lifelong impact on individuals having cataract surgery. Costing estimates are based on primary data collected in 14 epidemiological subregions by regional costing teams and on a literature review. Costings were estimated for different geographical coverage levels using non-linear cost functions. FINDINGS: Intra- and extra-capsular cataract surgeries are cost-effective ways to reduce the impact of cataract-blindness. Extra-capsular cataract surgery is more cost effective than intra-capsular surgery in all regions considered. Providing extra capsular cataract surgery to 95% of those who need it (95% coverage level) would avert over 3.5 million disability-adjusted life years (DALYs) per year globally. The cost-effectiveness ranges from 57 International dollars (1 dollar) per DALY in the WHO South-East Asia Region where there is high overall child and adult mortality to 1 dollar 2307 per DALY in the WHO Western Pacific Region where there is low overall child and adult mortality. CONCLUSION: Extra-capsular surgery for cataracts at a high level of coverage is the most cost-effective way of restoring sight in all epidemiological subregions considered. Analysts from countries within a region are encouraged to further contextualize the results based on their own country's specific parameters. PMID- 15298225 TI - The global burden of typhoid fever. AB - OBJECTIVE: To use new data to make a revised estimate of the global burden of typhoid fever, an accurate understanding of which is necessary to guide public health decisions for disease control and prevention efforts. METHODS: Population based studies using confirmation by blood culture of typhoid fever cases were sought by computer search of the multilingual scientific literature. Where there were no eligible studies, data were extrapolated from neighbouring countries and regions. Age-incidence curves were used to model rates measured among narrow age cohorts to the general population. One-way sensitivity analysis was performed to explore the sensitivity of the estimate to the assumptions. The burden of paratyphoid fever was derived by a proportional method. FINDINGS: A total of 22 eligible studies were identified. Regions with high incidence of typhoid fever (>100/100,000 cases/year) include south-central Asia and south-eastAsia. Regions of medium incidence (10-100/100,000 cases/year) include the rest of Asia, Africa, Latin America and the Caribbean, and Oceania, except for Australia and New Zealand. Europe, North America, and the rest of the developed world have low incidence of typhoid fever (<10/100,000 cases/year). We estimate that typhoid fever caused 21,650,974 illnesses and 216,510 deaths during 2000 and that paratyphoid fever caused 5,412,744 illnesses. CONCLUSION: New data and improved understanding of typhoid fever epidemiology enabled us to refine the global typhoid burden estimate, which remains considerable. More detailed incidence studies in selected countries and regions, particularly Africa, are needed to further improve the estimate. PMID- 15298226 TI - Cotrimoxazole prophylaxis reduces mortality in human immunodeficiency virus positive tuberculosis patients in Karonga District, Malawi. AB - OBJECTIVE: To estimate the impact of cotrimoxazole prophylaxis on the survival of human immunodeficiency virus (HIV)-positive tuberculosis (TB) patients. METHODS: A cohort study with a historical comparison group was conducted. End-of-treatment outcomes and 18-month survival were compared between TB patients registered in 1999 and patients registered in 2000 in Karonga District, Malawi. Case ascertainment, treatment and outpatient follow-up were identical in the two years except that in 2000 cotrimoxazole prophylaxis was offered to HIV-positive patients in addition to routine care. The prophylaxis was provided from the time a patient was identified as HIV-positive until 12 months after registration. Analyses were carried out on an intention-to-treat basis for all TB patients, and also separately by HIV status, TB type and certainty of diagnosis. FINDINGS: 355 and 362 TB patients were registered in 1999 and 2000, respectively; 70% were HIV positive. The overall case fatality rate fell from 37% to 29%, i.e. for every 12.5 TB patients treated, one death was averted. Case fatality rates were unchanged between the two years in HIV-negative patients, but fell in HIV positive patients from 43% to 24%. The improved survival became apparent after the first 2 months and was maintained beyond the end of treatment. The improvement was most marked in patients with smear-positive TB and others with confirmed TB diagnoses. CONCLUSION: Survival of HIV-positive TB patients improved dramatically with the addition of cotrimoxazole prophylaxis to the treatment regimen. The improvement can be attributed to cotrimoxazole because other factors were unchanged and the survival of HIV-negative patients was not improved. Cotrimoxazole prophylaxis should therefore be added to the routine care of HIV positive TB patients. PMID- 15298227 TI - A simplified screening strategy for thalassaemia and haemoglobin E in rural communities in south-east Asia. AB - OBJECTIVE: To evaluate a simple screening strategy for thalassaemia and haemoglobin (Hb) E in a prevention and control programme for thalassaemia in rural communities with limited resources. METHODS: Blood samples from 301 Thai Khmer participants were screened for thalassaemia and Hb E using a combined modified one-tube osmotic fragility (OF) test and a modified dichlorophenolindophenol (DCIP) precipitation test. Results were evaluated with standard haematological analyses including erythrocyte indices, Hb typing and quantification and polymerase chain reaction (PCR) analysis of alpha-globin and beta-globin genes. FINDINGS: Participants were divided into four groups according to the results of the combined tests. Altogether, 104 of 301 participants (34.6%) had negative results on both tests; 48 (15.9%) were positive on the OF test but not the DCIP test; 40 (13.3%) were negative on the OF test but positive on DCIP test; and 109 (36.2%) were positive on both tests. No carrier of clinically significant forms of thalassaemia (alpha(o)-thalassaemia, beta-thalassaemia) or Hb E was found among the group that had negative results for both tests. All participants with Hb E had positive DCIP tests. Carriers of alpha+-thalassaemia or Hb Constant Spring could generate either positive or negative OF test results but they all had negative DCIP tests. Using both tests as a preliminary screening for the three important groups of carriers gave a sensitivity of 100% and a specificity of 69.8%. The positive predictive value of the combined test was 77.2%. The negative predictive value was 100%. Further evaluation of the screening system by local staff at three community hospitals found a sensitivity of 98.1-100% and a specificity of 65.4-88.4% with positive predictive values of 75.0-86.9% and negative predictive values of 98.1-100%. CONCLUSION: A combined test using OF and DCIP could be used as an effective preliminary screening alternative to an electronic blood cell count for identifying carriers with alpha(o)-thalassaemia, beta-thalassaemia and Hb E. The strategy should prove useful for population screening in prevention and control programmes in rural communities in south-east Asia where laboratory facilities and economic resources are limited. PMID- 15298228 TI - The future incidence of leprosy: a scenario analysis. AB - OBJECTIVE: To investigate the impact of the current strategy for the elimination of leprosy on its incidence and to assess the consequences of failure to sustain this strategy. METHODS: Scenarios for assessing the impact of the elimination strategy were implemented in a computer simulation program. The scenarios reflected the assumptions made regarding contagiousness, transmission and bacille Calmette-Guerin (BCG) vaccination. The trend in case detection rate for the main countries in which leprosy was endemic during 1985-98 was fitted, and incidence up to 2020 was projected. FINDINGS: Owing to the gradual shortening of delays in detection up to 1998, and because of the low relapse rate that occurs with multidrug treatment MDT, incidence is predicted to decrease beyond 2000 in all scenarios. The annual decline was a few per cent higher when favourable assumptions were made about protection and coverage of BCG vaccination. Overall, the predicted annual decline in incidences ranged from 2% to 12%. CONCLUSION: The elimination strategy reduces transmission, but the decline may be slow. Relaxation of control after 2005 is unjustified given the uncertainty about the rate of decline and the adverse effects of longer delays in detection. A long term strategy for leprosy control should be adopted. PMID- 15298229 TI - Averting a malaria disaster in Africa--where does the buck stop? AB - The serious threat posed by the spread of drug-resistant malaria in Africa has been widely acknowledged. Chloroquine resistance is now almost universal, and resistance to the successor drug, sulfadoxine-pyrimethamine (SP), is growing rapidly. Combination therapy has been suggested as being an available and potentially lasting solution to this impending crisis. However, the current cost of combination therapy, and especially that of artemisinin combination therapy (ACT), is potentially a serious drawback, even if a significant part of its cost is passed on to the end-user. If the question of cost is not successfully addressed this could lead to adverse results from the deployment of combination therapy as first-line treatment. These adverse effects range from an increase in potentially fatal delays in infected individuals presenting to medical services, to exclusion of the poorest malaria sufferers from receiving treatment altogether. Urgent steps are needed to reduce the cost of combination therapy to the end-user in a sustainable way if it is to be usable, and some possible approaches are discussed. PMID- 15298230 TI - Genomics knowledge and equity: a global public goods perspective of the patent system. AB - Genomics, the comprehensive examination of an organism's entire set of genes and their interactions, will have a major impact on the way disease is diagnosed, prevented and treated in the new millennium. Despite the tremendous potential it holds for improving global health, genomics challenges policy-makers to ensure that its benefits are harnessed equitably across populations and nations. The classification of genomics as a global public good and the inequity encountered in the development and application of genomics knowledge are outlined in this paper, We examine the effect of the current patent system on the distribution of costs and benefits relating to genomics knowledge between countries of different economic strength. The global public goods concept provides a normative economic rationale for the modification of certain aspects of the current patent system and for the creation of complementary mechanisms to respond to the health needs of low-income and middle-income countries. PMID- 15298231 TI - New generation of non-profit initiatives tackles world's "neglected" diseases. PMID- 15298232 TI - Improving quality of cardiovascular care in the real world: how can we remove the barriers? PMID- 15298233 TI - Can we use automated data to assess quality of hypertension care? AB - OBJECTIVE: To determine whether extractable blood pressure (BP) information available in a computerized patient record system (CPRS) could be used to assess quality of hypertension care independently of clinicians' notes. STUDY DESIGN: Retrospective cohort study of a random sample of hypertensive patients from 10 Department of Veterans Affairs (VA) sites across the country. METHODS: We abstracted BPs from electronic clinicians' notes for all medical visits of 981 hypertensive patients in 1999. We compared these with BP measurements available in a separate vitals signs file in the CPRS. We also evaluated whether assessments of performance varied by source by using patients' last documented BP reading. RESULTS: When the vital signs file and notes were combined, a BP measurement was taken for 71% of 6097 medical visits; 60% had a BP measurement only in the vital signs file. Combining sources, 43% of patients had a BP reading of less than 140/90 mm Hg; by site this varied (34%-51%). Vital signs file data alone yielded similar findings; site rankings by rates of BP control changed minimally. CONCLUSIONS: Current performance review programs collect clinical data from both clinicians' notes and automated sources as available. However, we found that notes contribute little information with respect to BP values beyond automated data alone. The VA's vital signs file is a prototypical automated data system that could make assessment of hypertension care more efficient in many settings. PMID- 15298234 TI - Hypertension management: the care gap between clinical guidelines and clinical practice. AB - OBJECTIVE: To evaluate how well hypertension is managed in HMO patients and to assess opportunities for improvement. STUDY DESIGN: Retrospective cohort study. PATIENTS AND METHODS: The study population included HMO members (age 45-84 years) who had at least 1 ambulatory encounter with an ICD-9-CM diagnosis code of essential hypertension during the first 6 months of 1999. Medical records were reviewed to obtain information on blood pressure measurements, sex, age, coexisting medical conditions, smoking status, and changes made to the antihypertensive drug regimen. RESULTS: We identified 681 members with 3347 encounters related to hypertension management during 1999. Overall, 74 (11%) patients were at target blood pressure for all visits and 260 (38%) were at target blood pressure for at least 50% of the visits; 222 (33%) patients were not at target blood pressure for any visit. A history of coronary artery disease or cerebrovascular disease was associated with better blood pressure control (defined as being at goal levels during at least 50% of visits), while being older (age > or = 75) or having diabetes mellitus was associated with poorer control. Medication regimen intensifications occurred in 10% of visits with systolic blood pressure levels of 140-149 mm Hg, compared with 45% of visits with levels of > or = 180 mm Hg. Medication regimen intensifications occurred in 21% of visits with diastolic blood pressure levels of 90-99 mm Hg and 43% of visits with levels of > or = 100 mm Hg. CONCLUSION: Efforts are required to reduce "therapeutic inertia," particularly in patients with modestly elevated systolic blood pressure levels. PMID- 15298235 TI - The potential role of community-based registries to complement the limited applicability of clinical trial results to the community setting: heart failure as an example. AB - BACKGROUND: Clinical trials do not represent community settings, making widespread implementation of evidence-based medicine problematic. New heart failure treatments are an example, as results comparable to those of clinical trials have not been observed in the community. Alternatives to clinical trials could provide useful complementary information. OBJECTIVES AND METHODS: To review the clinical trials and community experiences in heart failure management by searching Pubmed with key words "observational studies," "clinical trials," and "heart failure," to present the preliminary results of a community-based heart failure registry as a complementary database, and to assess the potential value and limitations of the registry approach. RESULTS: Recent advances in the treatment of heart failure led to guidelines using clinical trial evidence as the rationale for transferring newer therapeutic technologies to the community practice setting. Implementation of such guidelines is slow, reflecting concerns over applicability of clinical trial results to the community setting. A community-based registry of beta-blocker treatment for heart failure showed outcomes comparable to those of clinical trials, despite significant differences between physicians and their patients in these settings. CONCLUSION: Registries can complement clinical trials to expedite technology transfer to the community setting. PMID- 15298236 TI - Impact of an educational intervention for secondary prevention of myocardial infarction on Medicaid drug use and cost. AB - OBJECTIVES: The objectives of this drug utilization review program were (1) to increase beta-blocker prescribing to fee-for-service post-acute myocardial infarction (AMI) Medicaid patients; (2) to improve compliance among patients who were prescribed beta-blockers post-AMI; and (3) to evaluate the economic implications of increased beta-blocker prescribing. STUDY DESIGN: Pre-post nonequivalent group design. PATIENTS AND METHODS: The intervention targeted physicians of Pennsylvania Medicaid recipients who had an AMI between November 1, 1998, and November 1, 1999. Educational materials were sent to the physicians of post-AMI patients not receiving beta-blockers. Preintervention and postintervention rates of beta-blocker prescribing in the Medicaid program within 7 and 30 days of discharge after an AMI hospitalization were compared. Similarly, pre- and postintervention compliance rates were compared for AMI patients who were prescribed beta-blockers. Cost savings and number of avoided deaths also were calculated. RESULTS: There was a 5.5%, to 6.9% increase in beta-blocker prescribing after the intervention, depending on the follow-up period. Postintervention AMI patients were 16% more likely to be prescribed a beta blocker. There was an 8.3% increase in patient compliance with beta-blocker therapy from preintervention to postintervention. In the first 2 years of the intervention, the estimated cost savings to the Pennsylvania Medicaid program ranged from 71,970 dollars to 76,678 dollars, respectively. An estimated 3 deaths were avoided. CONCLUSIONS: The intervention resulted in increased appropriate prescribing and compliance with beta-blockers among post-AMI patients. There also were estimated cost savings to Pennsylvania Medicaid as a result of reduced hospitalization, and fewer deaths. PMID- 15298237 TI - Aligning financial incentives with "Get With The Guidelines" to improve cardiovascular care. AB - OBJECTIVE: To describe the impact of a commercial insurer's financial incentives to hospitals in conjunction with collaboration with the American Heart Association (AHA) to accelerate implementation of Get With The Guidelines Coronary Artery Disease (GWTG-CAD), a quality improvement program to rapidly improve cardiovascular secondary prevention in hospitalized patients. STUDY DESIGN: Observational assessment of quality improvement program participation and implementation in response to financial incentives. METHODS: The study population included all hospitals that participated with the Hawaii Medical Service Association (HMSA, Blue Cross Blue Shield of Hawaii) Hospital Quality and Service Recognition Program and had more than 30 annual admissions for acute coronary artery disease. These 13 hospitals were given encouragement and financial incentives to implement GWTG-CAD. Financial incentives were determined by a prorated amount of the total HMSA hospital reimbursement for all acute services, as part of a more comprehensive hospital "pay for performance" program. RESULTS: Incentives to 10 of 13 eligible hospitals included reimbursement for half the annual cost of the AHA Patient Management Tool. In addition, HMSA's pay for performance program--the Hospital Quality and Service Recognition Program- distributed monetary awards totaling 354,883 dollars, based on points awarded for GWTG-CAD workshop attendance documentation (10 hospitals), recognition by the AHA as a GWTG-CAD hospital, and attainment of 85% adherence to the GWTG-CAD performance measures (4 hospitals). CONCLUSIONS: Community-based promotion of GWTG-CAD and financial incentives provided by a commercial insurer resulted in the rapid implementation of a secondary prevention program for coronary artery disease in most hospitals in the State of Hawaii within a single year. PMID- 15298238 TI - The effect of increases in HMO penetration and changes in payer mix on in hospital mortality and treatment patterns for acute myocardial infarction. AB - OBJECTIVE: To determine whether changes in health maintenance organization (HMO) penetration or payer mix affected in-hospital mortality and treatment patterns of patients with acute myocardial infarction (AMI). STUDY DESIGN: Observational study using patient-level logistic regression analysis and hospital and year fixed effects of data from the Agency for Healthcare Research and Quality's Healthcare Cost and Utilization Project Nationwide Inpatient Sample, a geographically diverse sample of 20% of the hospitalized patients in the United States. PATIENTS AND METHODS: Discharges of patients (n = 340,064) with a primary diagnosis of acute myocardial infarction who were treated in general medical or surgical hospitals that contributed at least 2 years of data to the HealthCare Cost and Utilization Project Nationwide Inpatient Sample from 1989 to 1996. In hospital mortality and rates of cardiac catheterization, angioplasty, or coronary artery bypass grafting for Medicare patients or non-Medicare patients were the main outcome measures. RESULTS: Among Medicare patients, increases in HMO penetration were associated with reduced odds of receiving cardiac catheterization, angioplasty, or coronary artery bypass grafting of 3% to 16%, but were not associated with any change in mortality risk. Increases in the number of HMOs within a metropolitan statistical area, our measure of HMO competition, were associated with small but significant increases in the odds of cardiac catheterization and angioplasty of about 2%. There was no pattern of changes in cardiac procedure rates or in-hospital mortality among non-Medicare patients. CONCLUSION: Increases in HMO penetration reduced cardiac procedure rates by statistically significant but small amounts among Medicare patients with AMI, without affecting mortality rates. PMID- 15298239 TI - [Let's teach the methods for doctor-patient communication during anesthesiology super-rotation]. PMID- 15298240 TI - [Effects of perfusion pressure on cerebral blood flow and oxygenation during normothermic cardiopulmonary bypass]. AB - BACKGROUND: Central nervous system dysfunction after cardiopulmonary bypass (CPB) is an important cause of morbidity and mortality after cardiac surgery. Perfusion pressure (PP) during CPB could be one of the important determinants of cerebral blood flow (CBF). The objective of the present study was to determine the effect of PP on CBF and cerebral oxgenation during normothermic CPB. METHODS: Twelve adult patients undergoing coronary artery bypass graft surgery were randomly assigned to one of two groups based on PP (High and Low group). Patients in High group received phenylephrine immediately after the onset of CPB to maintain PP between 60 and 80 mmHg. Oxyhemoglobin (O2Hb), deoxyhemoglobin (HHb), tissue oxygenation index (TOI), and oxidized cytochrome aa3 (CtOx) were measured by near infrared spectroscopy, and internal jugular venous bulb blood oxygen saturation (SjvO2) was measured simultaneously. S-100 beta protein concentrations were also measured before and after CPB. RESULTS: SjvO2 in High group increased significantly during CPB. CtOx in Low group decreased significantly during CPB, whereas TOI was unchanged. Although S-100 beta increased significantly at the end of CPB, there was no difference between the groups. CONCLUSIONS: These results suggest that maintaining high PP is benefical for CBF during normothermic CPB. PMID- 15298241 TI - [Effect of flumazenil on hypoglossal and phrenic nerves activities in rabbits]. AB - BACKGROUND: Upper airway obstruction and inadequate ventilation often arise during sedation and anesthesia by benzodiazepines (Bz). Flumazenil antagonizes these effects of active benzodiazepines on the central nervous system. To estimate the influence of flumazenil on the endogenous Bz system related respiratory control, we studied the effect of flumazenil and diazepam on the neural activity and the respiratory response caused by a brief (60 sec) respiratory arrest (RA) manifested in the hypoglossal nerve (HG) and the phrenic nerve (PH) activities in rabbits. METHODS: Experiments were performed on adult rabbits which were vagotomized, paralyzed and artificially ventilated with 50% N2O, 50% oxygen and 0.5% sevoflurane. We evaluated and compared the effects of the sequential administrations of flumazenil and diazepam on the peak amplitude (AMP) as well as the root mean square (RMS) of HG and PH, and respiratory cycle (Tc). RESULTS: Flumazenil by itself increased HG activity more than PH activity with no influence on Tc. But it was not dose-related. Previous administration of flumazenil in total dose of 0.25 mg x kg(-1) could not prevent the anticipated respiratory depression caused by diazepam 2.0 mg x kg(-1). These depressions are greater in HG activity than in PH activity. Additional flumazenil 0.15 mg x kg( 1) following the administration of diazepam promptly reversed these inhibitory effects on HG activity beyond the control level. The same dose of flumazenil, however, did not reverse PH activity sufficiently. RA response was characterized by raised AMPs and augmented RMSs (deltaAMPs, deltaRMSs) with marked prolongation in Tc (deltaTc). Flumazenil and diazepam did not seem to have any influence upon these RA responses. There was a significant change in cardiovascular parameters with the tested dosages of flumazenil and diazepam, but the change was in the normal physiological range. CONCLUSIONS: These results suggest the possibility that the endogenous benzodiazepine system is likely to play an inhibitory role in the regulation of respiration, especially in the maintenance of upper airway patency but the system is unrelated to the chemosensitive-respiratory control. PMID- 15298242 TI - [The effects of propofol on left ventricular systolic and diastolic function during induction of anesthesia--a transthoracic echocardiographic study]. AB - BACKGROUND: The effects of clinical doses of propofol on left ventricular (LV) systolic function remain controversial and LV diastolic function has not been evaluated during induction of anesthesia with propofol. We assessed the effects of propofol on LV systolic and diastolic function during induction of anesthesia in adult patients with transthoracic echocardiography. METHODS: Twenty-three patients, ASA 1-2 and age < 70 y.o., received propofol 2 mg x kg(-1) for induction of anesthesia. LV systolic function was evaluated by fractional shortening (FS), ejection fraction (EF), rate-corrected mean velocity of circumferential fiber shortening (Vcfc) and a contractile index, LV end-systolic wall stress (ESWS) versus Vcfc relation (ESWS-Vcfc relation). LV diastolic functions were assessed by analysis of transmitral flow velocity, peak early diastolic and late diastolic filling velocities (E wave and A wave), E/A ratio and E wave deceleration time (DT). RESULTS: After induction, propofol preserved FS, EF, Vcfc and ESWS-Vcfc relation and caused a significant decrease in E wave and A wave, and a significant increase in E/A ratio and maintained DT. CONCLUSIONS: During induction of anesthesia in adult patients, propofol preserved LV systolic and diastolic functions. PMID- 15298243 TI - [Changes in intracuff pressure of endotracheal tubes permeable or resistant to nitrous oxide and incidence of postoperative sore throat]. AB - BACKGROUND: We assessed the nitrous oxide (N2O) gas-barrier properties of a new endotracheal tube cuff, the Profile Soft-Seal Cuff (Resistant: R) (Sims Portex, Kent, UK). METHODS: The tracheas of randomly selected patients were intubated with the Profile Cuff (Permeable: P) (Sims Portex) tuble or with Portex Soft-Seal Cuff (R) (n=20 each) endotracheal tube. Cuffs were inflated with air, and intracuff pressure was measured during anesthesia using 67% N2O. Postoperative sore throat was assessed. In addition, the volume-pressure relationship of the cuff was determined in vitro. RESULTS: Cuff pressure increased gradually during anesthesia in both groups. The mean cuff pressure of the group R was significantly lower than that of the group P from 10 minutes to 230 minutes. The inflated gas and the deflated gas were not significantly different in both groups. The incidence of postoperative sore throat was not significantly different between the two groups. In vitro, the mean cuff pressure of the group R was significantly lower than that of the group P. CONCLUSIONS: The difference of cuff pressure is considered due to the difference in cuff compliance. PMID- 15298244 TI - [Efficacy of oral balance gel for dry mouth in preoperative patients]. AB - BACKGROUND: Following anticholinergic premedication and preoperative fasting, preoperative patients with a potential xerostomia have complaints associated with oral dryness. Xerostomia may lead to risk of mucosal burning and secondary infection. The purpose of this prospective study was to assess the effect of oral balance gel on dryness of the mouth in preoperative patients. METHODS: Thirty nine patients scheduled for elective surgery were randomly assigned to either of the group with or without using the oral balance gel. Severity of the dry mouth was assesed using a 4-point scale (0=none, 1=mild, 2=moderate, 3=severe) and diadochokinesis test was performed on the day before surgery and on arrival at the OR. RESULTS: Comparing results of the two stages, we found that patients with no treatment had significantly deteriorated state of dry mouth, but patients who had received the oral balance gel had no significantly worse dry mouth compared with the preoperative state. CONCLUSIONS: In this study, patients without the oral balance gel frequently reported oral symptons and oral dysfunction associated with xerostomia. We conclude that the use of oral balance gel in preoperative patients is effective for the prevention of dryness of the dry mouth. PMID- 15298245 TI - [Successful management of a patient for cardiac surgery with difficulty in weaning from cardiopulmonary bypass by using both isosorbide dinitrate and olprinone hydrochloride]. AB - A 57-year-old man with mitral stenosis underwent mitral valve plasty under general anesthesia. He had a history of cerebral infarction. Although he was with atrial fibrillation, his left ventricular function was good. Preoperative coronary angiography revealed no significant coronary stenosis. Induction of anesthesia and the surgical procedure had been uneventful, but the patient had difficulty to wean the patient from cardiopulmonary bypass because of unexpected low cardiac output syndrome. O1-prinone hydrochloride, a newly developed phosphodiesterase III inhibitor, was initiated in addition to high doses of dopamine and dobutamine. This increased the amplitude of the electrocardiogram and caused ST elevation of the lead II. A full dose of isosorbide dinitrate was administered intravenously to differentiate coronary artery spasm from coronary air embolism. This drastically improved the ventricular function and mixed venous oxygen saturation, and weaning from CPB was finally accomplished. The heart showed hypercontraction and inotropes were tapered gradually without further cardiac events. Although there are various etiologies for low cardiac output syndrome after CPB, the possibility of myocardial ischemia must be the first consideration. Full pharmacological support must be tried before initiating a mechanical assist modality. Coronary dilators, nitrates in particular, and phosphodiesterase III inhibitors are promising agents in such cases. PMID- 15298246 TI - [Anesthetic management of a child with Schwartz-Jampel syndrome]. AB - A 6-year-old child with Schwartz-Jampel syndrome (SJS) underwent tenotomy of bilateral lower limbs under general anesthesia. Patient with SJS has problems such as difficulty of intubation owing to microstomia and jaw muscle rigidity, and is susceptible to malignant hyperthermia by using volatile inhalation anesthetics. In this case, we used a laryngeal mask for airway management and anesthesia was maintained with inhalation of nitrous oxide and continuous i.v. infusion of propofol with caudal block, and his clinical course was uneventful. PMID- 15298247 TI - [Perioperative management of a patient with polycythemia who developed massive hemorrhage]. AB - A 34-year-old man, 170 cm in height and 70 kg in weight, was scheduled for emergency operation because of gastric perforation due to gastric cancer under general anesthesia. His preoperative blood analysis showed 5.2 x 10(3) mm(-3) of red blood cell, 18 g x dl(-1) of hemoglobin and 48% of hematocrit. Based on this and other data, he was diagnosed as having polycythemia caused by stress. Anesthesia was induced with thiopental and maintained with O2-N2O-sevoflurane. The intraoperative blood loss reached approximately 7,000 ml. Although we administered only 4 units of fresh-frozen plasma (FFP) and 9,150 ml of fluid with no red cell concentrated, his hemodynamic state was stable during surgery. After the surgery, we administered the minimum amount of FFP according to his blood analysis. Although red cell concentrated was not administered in the perioperative period, his general condition remained stable. PMID- 15298248 TI - [The use of a Parker Flex-Tip tube in a case of large laryngeal polyps]. AB - A 58-year-old man was scheduled for laryngomicrosurgery for treatment of large laryngeal polyps. Although awake induction was initially attempted to prevent airway obstruction, his vocal cords could not be visualized. We therefore tried intubation with a Parker Flex-Tip (PFT) tube under the guidance of a bronchial fiberscope (BF). After inserting the BF, his trachea was intubated easily. The operation was performed without any complications. We conclude that a PFT tube is useful for endotracheal intubation in a patient with large laryngeal polyps. PMID- 15298249 TI - [Withdrawal syndrome in a critically ill child after sedation with midazolam and fentanyl]. AB - A 7-year-old girl suffered from withdrawal syndrome with systemic convulsion after sedation with midazolam and fentanyl. She had a history of severe accidental alkaline esophagitis, and under went polysurgeries. This time, she was scheduled to receive reconstruction of the esophagus with small intestine in order to resolve esophageal stenosis. Operation and anesthesia lasted for 14 hours, and 17 hours, respectively. In the postoperative period, she was under heavy sedation with midazolam and fentanyl in order to keep neck position immobile. Her sedation persisted for 14 postoperative days, and the total doses of midazolam and fentanyl were more than 100 mg x kg(-1), and 6.4 mg x kg(-1), respectively. Thereafter, her sedation was tapered and discontinued within about 24 hours. After 12 hours, she suddenly developed systemic convulsion with loss of consciousness. There was no evidence of obvious organic central nervous system abnormality. We suspected withdrawal syndrome, and gradual decrease of midazolam and fentanyl prevented her from going into withdrawal syndrome. We have to pay attention to withdrawal syndrome when heavy and long sedation with midazolam and fentanyl was employed and the drugs were then tapered and discontinued. PMID- 15298250 TI - [Recovery from fatal ventricular fibrillation after immediate application of percutaneous cardiopulmonary support]. AB - We present a patient who recovered from refractory ventricular fibrillation after immediate application of percutaneous cardiopulmonary support (PCPS). On the postoperative day (POD) 3 after the Y-grafting surgery for abdominal aortic aneurysm, circulation collapsed due to sudden onset of ventricular fibrillation. Because ventricular fibrillation had persisted in spite of medical treatment and defibrillation, we established PCPS and his circulation recovered. Although an emergent coronary angiography revealed no new lesions, we performed an emergent percutaneous catheter intervention to deny the possibility that ischemic changes had contributed to the arrhythmia. Soon after percutaneous transluminal coronary angioplasty, we successfully weaned him from PCPS, and extubated his trachea on the POD 5 without any neurological deficits. On the POD 8, ventricular fibrillation occurred again and defibrillation was effective at this time. We suspected cardiac ischemia, prolonged QT interval, and electrical remodeling due to hypertrophic heart as possible causes of refractory ventricular fibrillation. Therefore, we performed percutaneous transluminal coronary angioplasty, terminated famotidine administration, maintained normal electrolytes level, started administration of beta-blocker, and implanted an cardioverter defibrillator. On the POD 16, he was discharged from the ICU with no neurological deficits. PMID- 15298251 TI - [Intra-aortic balloon pumping is effective for hemodynamic management of catecholamine resistant ampulla (Takotsubo) cardiomyopathy]. AB - Ampulla (Takotsubo) cardiomyopathy resembles acute myocardial infarction, with absent coronary stenosis. We report a case of catecholamine resistant ampulla cardiomyopathy, successfully managed using intra-aortic balloon pumping (IABP). After laparotomy, the patient developed circulatory collapse. T wave inversion was observed on electrocardiogram. In spite of circulatory support using high dose cathecholamines, the hemodynamics of the patient showed no improvement. Upon starting IABP, the patient's hemodynamics were improved and the dose of catecholamines could be reduced. Because the ampulla cardiomyopathy is considered a subtype of 'stunned myocardium' and the hemodynamic disorder is temporary and reversible, IABP appears effective for hemodynamic management. PMID- 15298252 TI - [The usefulness of intraoperative TEE monitoring in a patient with renal cell carcinoma]. AB - A 55-year-old man was admitted to our hospital for treatment of renal cell carcinoma. Preoperative MRI showed infradiaphragmatic extension of the tumor thrombus. However, intraoperative transesophageal echocardiography (TEE) revealed intracardiac extension of the thrombus. Therefore, the tumor thrombus was extirpated under cardiopulmonary bypass. The patient recovered without any complications. Intraoperative TEE monitoring is useful not only for the evaluation of cardiac functions but also for intraoperative diagnosis of a tumor thrombus during the operation for renal cell carcinoma. PMID- 15298253 TI - [The successful anesthetic management of a patient with pheochromocytoma using prostaglandin E1 and a novel, short-acting beta-adrenergic blocker: landiolol hydrochloride]. AB - A 48-year-old male with a history of hypertension was scheduled to undergo resection of a tumor in the upper region of the left kidney. However, his operation was postponed once because pheochromocytoma was suspected from the tumor location, sweating, and insomnia in addition to hypertension. The measurement of plasma catecholamines confirmed the presence of pheochromocytoma. Anesthesia was induced with thiopental and fentanyl, while ventilating with 5% sevoflurane in oxygen, followed by tracheal intubation facilitated with vecuronium. Anesthesia was maintained with 33% nitrous oxide and 0.6-3% sevoflurane in oxygen, in conjunction with fentanyl and 1% mepivacaine through an epidural catheter (T11-12). An arterial catheter and a pulmonary artery catheter were inserted. From the beginning of the operation, prostaglandin E1 and landiolol were administered continuously. Systolic blood pressure and heart rate were controlled between 90-140 mmHg and 80-105 beats x min(-1), respectively. Systemic vascular resistance was stable between 700-900 dyn x s x cm(-5) throughout the procedure. The operation was completed uneventfully. The patient was transferred to the general ward, extubated, and was in a stable condition. Various combinations of vasodilating and antihypertensive drugs have been used intraoperatively during the resection of pheochromocytoma. Of these, prostaglandin E1 and landiolol hydrochloride are very promising for maintaining stable hemodynamics. PMID- 15298254 TI - [Bispectral index monitoring during anesthesia with propofol for electroconvulsive therapy]. AB - We studied the changes in the bispectral index (BIS) of 22 patients during electroconvulsive therapy (ECT) under propofol and succinylcholine anesthesia. BIS values (mean+/-SD) were 94+/-4 before anesthesia, 51+/-15 before ECT, 49+/-20 after ECT, and 60+/-17 at complete re-awakening. The BIS values at re-awakening were significantly different from those before anesthesia (P<0.05). In conclusion, our study suggests that the BIS values following ECT might not reliably correlate with the patients' clinical level of consciousness. PMID- 15298255 TI - [Anesthetic management for cleft palate plasty in a patient with Pierre-Robin syndrome]. AB - A girl (15 months-old) with Pierre-Robin Syndrome was scheduled for cleft palate plasty. She had a past history of difficulty feeding, mild airway obstruction during sleeping and mental retardation. After induction of anesthesia with an inhalational anesthetic technique, conventional tracheal intubation was impossible. We introduced a laryngeal mask airway (LMA) and successfully intubated through the LMA. After extubation of the tracheal tube, she developed upper airway obstruction with arterial desaturation. We ventilated her lungs in the lateral position with an inhalation of epinephrine and injection of methylprednisolone. Airway obstruction then improved gradually. In this case, LMA was a valuable device as a guide for the tracheal intubation. Because airway obstruction after extubation is a common complication in a patient with Pierre Robin syndrome, we need to observe the patient closely. PMID- 15298256 TI - [Subcutaneous emphysema during endoscopic resection of the gastric mucosa]. AB - A 69-year-old male underwent endoscopic mucosal resection for an early gastric cancer under general anesthesia with sevoflurane and nitrous oxide. Because of gastro-intestinal gas insufflation for endoscopic visualization, abdominal distension was found remarkable and the peak inspiratory pressure increased from 20 cmH2O to 30 cmH2O. About 2.5 hours after initiation of the procedure, iatrogenic perforation was identified endoscopically and emphysema became apparent not only in the scrotum, but also in the subcutaneous tissues of the chest and neck. In order to ameliorate this complication, an emergent distal gastrectomy was performed involving laparotomy. Nitrous oxide should be avoided in endoscopic mucosal resection under general anesthesia to prevent intestinal distension and attention should be paid to subcutaneous emphysema as a sign of perforation of the gastro-intestinal tract. PMID- 15298257 TI - [Epidural subarachnoid space and placement of epidural catheters]. PMID- 15298258 TI - [The transintervertebral disc approach for educational practice of the neurolytic celiac plexus block]. AB - BACKGROUND: The transintervertebral disc approach was proposed recently for percutaneous neurolytic celiac plexus block (NCPB). Its superior simplicity, reliability, as well as safety potentially overcome the technical hurdles of NCPB that may interfere with the practical use of this validated analgesic intervention for abdominal cancer pain. The present study was conducted to evaluate the effectiveness of the use of this approach in a resident education program for NCPB. METHODS: The clinical results of NCPBs conducted from January 2001 to September 2002 were examined comparing that performed by institutional residents with that by specialized physicians authorized by the Japanese Society of Pain Clinicians. The transintervertebral disc approach was used in all cases. Each resident completed NCPB under close supervision of the specialists. RESULTS: Twenty-four patients received NCPB during the study period. Seven residents randomly completed 12 procedures and 4 specialists did others. The duration of fluoroscopy to complete the procedure was 256+/-109 sec in the resident group and 392+/-194 sec in the specialist group (ns). Significant pain reduction was obtained immediately after NCPB in all patients without any intergroup difference. No critical complication was observed in each group. CONCLUSIONS: The transintervertebral disc approach can be used effectively and safely in educational practice of NCPB for less-trained physicians. PMID- 15298260 TI - [Progress and problems related to transfusion therapy]. PMID- 15298259 TI - [A trouble in air conditioning in the operating area]. AB - We experienced malfunction of air conditioning system in the operating area. Rust inside the circulating pipe to the operating area was an obstacle to inflow of cold and hot water. Installing an additional air conditioning system and treatment with chemicals to remove the dust made it possible to adjust room temperature appropriately. Anesthesiologists should be interested and understand equipments used in the operation area such as air conditioning system. PMID- 15298261 TI - [Problems and an overview of organizations dealing with blood and blood products]. PMID- 15298262 TI - [Legal basis of operations dealing with blood and blood products and blood transfusion treatment: Promulgation of a new law dealing with blood and significance of the revised drug law]. PMID- 15298263 TI - [Proper blood component transfusion. 1. Erythrocyte preparations]. PMID- 15298264 TI - [Proper blood component transfusion. 2. Platelet preparations]. PMID- 15298265 TI - [Proper blood component transfusion: 3. Fresh frozen plasma and plasma fraction preparations other than albumin and coagulation factor components]. PMID- 15298267 TI - [Proper blood component transfusion. 5. Albumin preparations]. PMID- 15298266 TI - [Proper blood component transfusion. 4. Coagulation factor preparations]. PMID- 15298268 TI - [Adverse effects of blood transfusion. 1. Posttransfusion hepatitis]. PMID- 15298269 TI - [Adverse effects of blood transfusion. 2. Infections other than hepatitis caused by transfusion]. PMID- 15298270 TI - [Adverse effects of blood transfusion. 3. Post-transfusion GVHD]. PMID- 15298271 TI - [Adverse effects of blood transfusion. 4. Hemolytic effects]. PMID- 15298272 TI - [Adverse effects of blood transfusion. 5. Non-hemolytic adverse effects]. PMID- 15298273 TI - [New policies to prevent transfusion-related adverse effects. 1. The current status and future prospect of preoperative autologous blood donation]. PMID- 15298274 TI - [New methods to manage transfusion-related problems. 2. Leukocyte removal and inactivation of pathogens before blood preservation]. PMID- 15298276 TI - [Cell therapy. 1. Donor lymphocyte infusion therapy]. PMID- 15298275 TI - [The current status and policies to counteract accidents related to blood transfusion]. PMID- 15298277 TI - [Cell therapy. 2. Vascular regeneration or vasculogenesis]. PMID- 15298278 TI - [Cell therapy. 3. Cell therapy, regeneration therapy and translational research- in an effort to update the blood transfusion department]. PMID- 15298279 TI - [Progress and problems facing transfusion therapy--with special reference to the domain of internal medicine: a discussion]. PMID- 15298280 TI - [Grave's disease associated with Wilson disease]. PMID- 15298281 TI - [Markedly improving sarcoid cervical myelopathy after the diagnostic treatment of prednisolone, associated with highly spondylotic spines]. PMID- 15298282 TI - [Marginal zone B-cell lymphoma of MALT with improvement of panhypopituitarism by chemotherapy]. PMID- 15298283 TI - [Acute myocardial infarction produced by methylergometrine maleate-induced coronary vasospasm]. PMID- 15298284 TI - [Serial brain MRI findings of a patient with thrombotic thrombocytopenic purpura complicated by malignant hypertension and reversible posterior leukoencephalopathy syndrome]. PMID- 15298285 TI - [Thiazide-induced hypercalcemic nephropathy in a patient with secondary hypoparathyroidism]. PMID- 15298286 TI - [GIST (gastrointestinal stromal tumor)]. PMID- 15298287 TI - [Palliative therapy: the current status in Japan]. PMID- 15298288 TI - [Development of the treatment for bulbospinal muscular atrophy]. PMID- 15298289 TI - [Ongoing myocardial damage in chronic heart failure]. PMID- 15298290 TI - More than a hill of beans' difference. PMID- 15298291 TI - Back to basics managing IT projects. AB - To minimize the likelihood of IT project failure, it's important to focus on five foundations of sound project management: roles, committees, charters, plans, status reporting. PMID- 15298292 TI - Medicare implications of discounts to the uninsured. AB - Discount policies are likely to face increased scrutiny in light of congressional inquiries on Medicare outlier payments and recent media attention regarding pricing for the uninsured. If you're considering making changes to your hospital's policy, you'll need to understand the current regulatory environment, recognize potential concerns, and develop a plan of action that complies with recent HHS guidance. PMID- 15298293 TI - How are hospitals financing the future? Core competencies in capital planning. AB - Financing the Future is a yearlong project to help hospitals take advantage of growth opportunities. Led by HFMA in partnership with GE Healthcare Financial Services, the project provides information, insights, strategies, and tools designed to help hospitals finance their future. The findings of Financing the Future are based on research conducted by HFMA and PricewaterhouseCoopers. To access the first four Financing the Future reports, visit www.financingthefuture.org. PMID- 15298294 TI - Making the tough choices for cost control. AB - To prioritize cost savings, providers can benefit from following a fundamental approach that involves using easily attainable small-scale initiatives (less than 50,000 dollars) to support larger-scale projects (typically over 250,000 dollars) that can realize improvements in six months. Increases in capital that result from these efforts can then be used to focus on equally important opportunities where the organization has less control, or payoff may take longer. PMID- 15298295 TI - Sustaining a market-based healthcare system. AB - Purchasers of health care are not holding the healthcare system accountable for quality and cost. Employers need to: Offer their employees a wide range of choices in health coverage. Earmark for employees' purchase a fixed dollar amount for health care set at or below the price of the low-priced plan. Insist that carriers and providers report the quality of care delivered. PMID- 15298296 TI - Balancing physician and cost-containment demands. AB - For clinicians, it's tempting to draw conclusions that new medications are easier to use and monitor, are safer, and therefore result in better outcomes. Similarly, for CFOs, it's tempting to view an older medication with a lower cost as the best choice. Both physicians and CFOs need reliable data that measure the total cost of care of patients in their hospital to select the appropriate medications. PMID- 15298297 TI - Election 2004: implications for providers. AB - It may be difficult to believe, but President Bush and Senator Kerry do agree on a few important issues such as the need for improved healthcare information technology. But in regard to medical liability reform, health care for small businesses, and Medicare managed care, there are significant differences between their proposals. No matter who wins the presidential election, healthcare providers are going to feel the impact-especially with more seniors in HMOs and more low-income families seeking health care. PMID- 15298298 TI - Hospital-physician joint ventures: threat-or opportunity? AB - Physicians are increasingly drifting away from their hospitals to form ventures that compete with those very hospitals. Healthcare executives are being pressed to think of creative ways to maintain relationships with their physicians and "stay in the swim" of the revenue stream generated by ancillary services. Among these arrangements are integrated "centers ol excellence," ancillary services joint ventures, physician medical director-management relationships, and exclusive specialty services contracts. PMID- 15298299 TI - The drug discount card: will it cut drug prices for seniors? AB - Medicare's drug discount card program is expected to help participants save an average of 10 to 15 percent on the cost of outpatient prescription drugs. Many of the beneficiaries signing up for the program will also qualify for financial assistance. In 2006, the program will be replaced by Medicare's outpatient drug benefit. Data from the card program should help federal actuaries estimate the cost of the drug benefit. PMID- 15298300 TI - Conquering capacity. AB - Recognizing existing capacity constraints and the accompanying lost revenue and market share, Lucile Packard Children's Hospital at Stanford implemented an improvement initiative, beginning with an assessment of its patient flow processes. Key success factors included: Measuring to improve patient flow, increasing care and service coordination, redefining staffing and job functions, creating and sustaining culture change. PMID- 15298301 TI - Strategic price setting: ensuring your financial viability through price modeling. PMID- 15298302 TI - Delivering bad news. PMID- 15298304 TI - Outpatient procedures are not always the cheapest. PMID- 15298303 TI - The CFO as marketing's champion. PMID- 15298305 TI - The world around us. PMID- 15298306 TI - A 4-year-old girl with fever and "appendicitis-like" abdominal pain. PMID- 15298307 TI - Environmental risks in childhood. AB - Differences between children and adults with respect to exposure rates, absorption of chemicals, metabolism, and organ development make children uniquely vulnerable to environmental hazards. Moreover, biology does not exist outside of the social life of the child. At least two important implications flow from these findings. First, pediatricians must pay closer attention to the conditions of childhood and the specific details of a child's life in attempting a complete understanding of childhood diseases, particularly their sources in environmental and social conditions. At a minimum, pediatricians must inquire more carefully about environmental exposures and children's complaints in order to make accurate diagnoses. Perhaps equally critical, pediatricians must engage in more aggressive prevention efforts. While they often try to prevent exposure by educating parents about keeping household chemicals away from young children, they might likewise consider that the information they possess about children's special vulnerability to environmental risks could usefully inform political and social decisions to protect children from those risks. This includes, for example, supporting air pollution standards that are protective of children and advocating for stricter controls for certain chemicals to reduce health risks for this vulnerable group. PMID- 15298308 TI - Mercury in the environment: sources, toxicities, and prevention of exposure. AB - Acute and chronic exposure to mercury can significantly affect the health of a population, specifically the children. Methylmercury may pose the highest threat, as it is ubiquitous in the environment and it is a potent neurotoxicant. Methylmercury easily passes through the placenta to the developing fetus. Elemental mercury, or quicksilver, also poses a threat to children because it may be found readily in schools, hospitals, and medicine cabinets, and its intriguing liquid nature may be enticing to children. Pediatricians must be diligent in informing patients of possible exposure sources, and alerting them to new government advisories and recommendations. They should also be knowledgeable regarding classic clinical presentations of mercury toxicity. It is only in cases involving a knowledge of mercury that appropriate historical information is obtained and correct diagnoses are made. Preventing mercury exposure and consequent toxicity is of importance because therapies are controversial and long term consequences may be significant. PMID- 15298309 TI - Does anything beat DEET? AB - In comparison trials, DEET is more effective than any other insect repellent. Despite some reports of serious adverse events, when comparing the thousands of other reports of exposure and millions of past users, DEET has a good safety record. The appropriate and safest concentration to use on children remains unclear, however. Due to potential absorption through the skin, prudence would dictate that the lowest effective concentration for the time period of exposure be used. Because research has shown that solvents with less skin permeation may be used as an alternative to the ethanol used in some commercial DEET preparations, manufacturers could develop products that are less likely to be absorbed. Pediatricians should be familiar with the duration of action of various formulations of DEET and the efficacy (and in some cases lack of efficacy) of other products in order to advise patients on safe but effective methods of insect control. PMID- 15298310 TI - Bronchiolitis poses significant public health burden. AB - Bronchiolitis is the leading cause of infant morbidity, and hospitalization rates are rising. The effect of this disease is not limited to the acute illness episode. Approximately 40% to 50% of children diagnosed with bronchiolitis suffer from subsequent wheezing and airway reactivity or asthma. Attempts to address the burden of this disease via vaccine development have been largely unsuccessful, and treatment is purely supportive rather than curative. As such, primary prevention is paramount. If outdoor air pollution exacerbates this disease, as has been found for other pediatric respiratory diseases, actions to ensure that regulatory standards protect this vulnerable population will be paramount. Increased anatomic and physiologic susceptibility to the pro-inflammatory effects of air pollutants, coupled with the pro-inflammatory response in bronchiolitis, underlies the concern that infants exposed to higher levels of ambient air pollutants may be at increased risk for developing more severe bronchiolitis requiring hospitalization. PMID- 15298311 TI - Consequences of acute and chronic exposure to arsenic in children. AB - Arsenic is a toxic chemical and may cause adverse health effects in children and adults. It is known to affect the nervous, gastrointestinal, and hematological systems and cause skin and internal cancers in people exposed to levels greater than 300 ppb in their drinking water. For most people, the major exposure to arsenic comes from food (8 to 14 microg inorganic arsenic per day), but when the arsenic level in water is elevated, drinking water becomes the predominant source of exposure. Because it is very difficult to limit arsenic exposure from food, it would be wise to limit arsenic exposure from those more controllable sources. Pediatricians should ascertain the levels of arsenic in drinking water of patients with high arsenic levels, using the supplier or, in the case of private wells, a professional water-testing laboratory assay. The Safe Drinking Water Act does not cover private wells or those water systems with less than 15 hook-ups or those that serve less than 25 people. Pediatricians should instruct parents to use prepared baby formulas or prepare them using water with the arsenic removed and to curtail playing time for younger children in places that have sand containing large amounts of arsenic. Such procedures will limit arsenic exposure to a minimum. PMID- 15298312 TI - Indoor air pollution: a health concern. PMID- 15298313 TI - The effects of housing interventions on child health. AB - Housing hazards contribute to considerable morbidity and mortality among millions of children each year in the US, but few interventions are proven to control asthma and lead poisoning. Moreover, there is little evidence that many of the current recommendations to control residential hazards are safe and efficacious. The only interventions that have been found to work consistently are home visitation programs and home modification, such as installment of window guards and carpet removal. Altering the environment to protect the health of children requires pediatrician intervention. New models of cooperation between pediatricians and public health agencies must deal with residential hazards in an integrated manner and cannot be focused on one disease process or one method at a time. With research in more effective environmental interventions and pediatric public-health partnerships, primary and secondary prevention of diseases from residential hazards may become a reality in the future. PMID- 15298314 TI - Community psychiatry in unusual and remote settings. PMID- 15298315 TI - Community psychiatry in the sub-arctic. Experiences with the shift from hospital based to community-based psychiatric services in Northern Norway. PMID- 15298316 TI - Remoteness and issues in mental health care: experience from rural Australia. PMID- 15298317 TI - [Mental health in Basilicata]. PMID- 15298318 TI - Contemporary thinking about the role of genes and environment in eating disorders. AB - OBJECTIVE: To review recent literature documenting how family, twin and molecular genetic studies of eating disorders have revolutionized our conceptualizations of anorexia and bulimia nervosa. METHODS: We summarized extant literature on genetic epidemiology of eating disorders. RESULTS: Results of extant studies highlight the underlying biological vulnerabilities associated with these conditions. Genetic research has also opened up new avenues and approaches for exploring how the environment exerts its influence on risk. CONCLUSIONS: We discuss state-of the-science findings in the genetics of eating disorders, explore various mechanisms of gene-environment interplay, and discuss implications of this research for science, practice, families and individuals with eating disorders. PMID- 15298319 TI - Schizophrenia: still Kraepelin's dementia praecox? AB - Asking whether E. Kraepelin's early dementia praecox and disease concepts (1896) are still valid today, we condensed his early theory into four theses: 1) schizophrenia is a disease entity, distinguishable from affective psychosis. 2) It is caused by a specific neuropathology. 3) It usually manifests itself in adolescence or early adulthood. 4) Underlying schizophrenia is a progressive disease process that leads to defects and dementia. Having tested whether Kraepelin's dementia praecox and modern schizophrenia are actually comparable, we studied 1) how schizophrenia and depression are linked or separable in terms of symptoms, risk factors and illness course from onset until five years after first treatment contact. The analyses are based on a population-based sample of 130 first admissions because of schizophrenia, 130 age- and sex-matched first admissions because of unipolar depressive disorder and 130 "healthy" population controls from the study area. 2) Results will be presented that, though not very specific, confirm Kraepelin's farsighted hypothesis of a neuropathological basis of the disorder. In this context it will be discussed whether the brain changes are developmental or degenerative in origin. 3) The distribution of age of onset extends far into old age. In a sample of 1109 consecutive first admissions because of nonaffective psychosis from the total age range it was shown that age dependent developmental factors modify certain components of symptomatology linearly and significantly. The main risk factors, too, significantly change with age. 4) Long-term course was examined in three studies of epidemiologically recruited first-episode samples: Study 1 included five cross sections over 5 years, Study 2 was a prospective pre-post-comparison over 12 years supplemented by a retrospective assessment of the illness course (IRAOS) and Study 3 encompassed 10 cross sections over fifteen years. Finally, the disease concept of schizophrenia, as it presents itself in the light of current knowledge, will be outlined and compared with Kraepelin's earlier and later view of the disorder. PMID- 15298320 TI - Cannabis use and risk of psychosis: an etiological link? AB - OBJECTIVE: The nature of the link between cannabis use and psychosis remains to be clarified. METHOD: The paper reviews the evidence suggesting that cannabis may be a risk factor for psychosis onset. RESULTS: Cross-sectional and retrospective epidemiological studies show that individuals with psychosis use cannabis more often than other individuals in the general population. It has long been considered that this association is explained by the self-medication hypothesis, postulating that cannabis is used to self-medicate psychotic symptoms. This hypothesis has been recently challenged by several prospective studies carried out in population-based samples, showing that cannabis exposure is associated with an increased risk of psychosis, possibly by interacting with a pre-existing vulnerability for these disorders. A dose-response relationship was found between cannabis exposure and risk of psychosis, and this association was independent from potential confounding factors such as exposure to other drugs and pre existence of psychotic symptoms. However, the diagnostic specificity is weak, as cannabis exposure may be a risk factor for the occurrence of a large spectrum of psychiatric disorders, ranging from schizophrenia to mood and anxiety disorders. CONCLUSION: Considering the growing number of adolescents exposed to cannabis, the impact of this substance on the population mental health should be further explored. PMID- 15298321 TI - Health status, resource consumption, and costs of dysthymic patients in Italian primary care. AB - AIMS: To describe the health status, resource consumption and costs of patients with dysthymic disorder in the Italian primary care setting. METHODS: A total of 79 general practitioners (GPs) participated the study. Diagnosis was based on each GP's clinical assessment. At entry the Mini-International Neuropsychiatric Interview (MINI) was used as a supporting diagnostic aid. Health status was measured with the SF-36 questionnaire. Resource consumption and costs regarded the six months before enrolment. RESULTS: Out of 598 patients enrolled by GPs according to their clinical assessment, 503 fulfilled the MINI criteria and 95 did not. The latter had a better perception of their health than the former. Resource consumption was similar in the two groups; and the total per patient six month costs did not differ significantly. CONCLUSIONS: The study confirms there may be a gap between standardised criteria for defining dysthymia and everyday clinical practice. All dysthymic patients diagnosed by GPs might be considered together from a health policy perspective. PMID- 15298322 TI - [Mental health department accreditation. Classification and assessment criteria]. AB - AIMS: To compare different accreditation categories in Italy and to suggest some criteria to assess them. METHODS: The A. compare specific characteristics of different model of normative and professional accreditation and suggest some criteria to distinguish different type of accreditation programme and to assess quality and usefulness of an accreditation programme. RESULTS AND CONCLUSIONS: Different types of accreditation are attributed to some basic criteria: voluntary/by law, high/low standard, assessment object. Criteria to assess the usefulness are: check list accuracy, support to quality improvement, documental burden, services impact. PMID- 15298323 TI - Sponge left in pt.: did required surgery delay treatment & cause death? PMID- 15298324 TI - Failure to monitor local anesthesia pt. before discharge. Case on point: Messer v. Martin, 2004 WL 1171736 N.W.2d -WI(2004). PMID- 15298325 TI - NC: Home Health Agency's CNA applies for comp.: how did 'illiterate' & 'retarded' become CNA? PMID- 15298326 TI - MO: nurse's aide's knee 'pops' climbing stairs: work-related stair climbing injury compensable. PMID- 15298327 TI - Caveat when drawing blood on DUI suspects. Case on point: City of Las Vegas v. Walsh, 2004 WL 1293893 P.2d -NV. PMID- 15298328 TI - Interaction between phonological and grammatical processing in single word production in Kiswahili. AB - Grammatical priming of picture naming was investigated in Kiswahili, which has a complex grammatical noun class system (a system like grammatical gender), with up to 15 noun classes that have obligatory agreements on adjectives, verbs, pronouns and other parts of speech. Participants heard a grammatically agreeing (concordant), nonagreeing (discordant) or neutral prime before seeing a picture of a common object and being asked to name the object. Priming was found, with naming following concordant primes being faster than naming following the neutral prime ('say'). However, more interestingly, effects were found such that where two noun classes share a prefix, the grammatical prime from each of these two noun classes also primed words that have the same prefix but are not in the same noun class, and hence for which the prime was not grammatical. It is concluded that the prime appears to be facilitating the phonological form of the prefix rather than the syntacto-semantic group of words that are known as a noun class, and that the phonological form associated with a grammatical entity may be more significant in its processing than has previously been supposed. PMID- 15298329 TI - The smooth signal redundancy hypothesis: a functional explanation for relationships between redundancy, prosodic prominence, and duration in spontaneous speech. AB - This paper explores two related factors which influence variation in duration, prosodic structure and redundancy in spontaneous speech. We argue that the constraint of producing robust communication while efficiently expending articulatory effort leads to an inverse relationship between language redundancy and duration. The inverse relationship improves communication robustness by spreading information more evenly across the speech signal, yielding a smoother signal redundancy profile. We argue that prosodic prominence is a linguistic means of achieving smooth signal redundancy. Prosodic prominence increases syllable duration and coincides to a large extent with unpredictable sections of speech, and thus leads to a smoother signal redundancy. The results of linear regressions carried out between measures of redundancy, syllable duration and prosodic structure in a large corpus of spontaneous speech confirm: (1) an inverse relationship between language redundancy and duration, and (2) a strong relationship between prosodic prominence and duration. The fact that a large proportion of the variance predicted by language redundancy and prosodic prominence is nonunique suggests that, in English, prosodic prominence structure is the means with which constraints caused by a robust signal requirement are expressed in spontaneous speech. PMID- 15298330 TI - Syllable onset intervals as an indicator of discourse and syntactic boundaries in Taiwan Mandarin. AB - This study looks at the syllable onset interval (SOI) patterning in Taiwan Mandarin spontaneous speech and its relationship to discourse and syntactic units. Monologs were elicited by asking readers to tell stories depicted in comic strips and were transcribed and segmented into Discourse Segment Units (Grosz & Sidner, 1986), clauses, and phrases. Results showed that the degree of final lengthening was modulated by boundary types. Lengthening before discourse boundaries was longer than that before clausal boundaries, which was in turn longer than that before phrasal boundaries. Final SOI lengthening also seemed to reflect cognitive load. At the discourse and clausal levels, the degree of lengthening is modulated by narration order. First narrations tended to have longer final SOIs than second narrations. In addition, there was also a mild lengthening effect in nonfinal SOIs, as was evidenced by the length differences in initial and medial SOIs and the differential lengthening effect regarding the positioning of phrases in a clause. PMID- 15298331 TI - Probability in the grammar of German and Dutch: interfixation in triconstituent compounds. AB - This study addresses the possibility that interfixes in multiconstituent nominal compounds in German and Dutch are functional as markers of immediate constituent structure. We report a lexical statistical survey of interfixation in the lexicons of German and Dutch which shows that all interfixes of German and one interfix of Dutch are significantly more likely to appear at the major constituent boundary than expected under chance conditions. A series of experiments provides evidence that speakers of German and Dutch are sensitive to the probabilistic cues to constituent structure provided by the interfixes. Thus, our data provide evidence that probability is part and parcel of grammatical competence. PMID- 15298332 TI - Tearing down the walls between O.R. and S.P.D. AB - The focus of this article is the relationship between the Operating Room (O.R.) and the Sterile Processing Department (S.P.D.). The goal is to help readers gain insights into the driving forces which impact on both departments and to help them establish realistic expectations for improving the performance and quality of this relationship. It is important to discuss ways to solve conflict and enhance teamwork between the O.R. and S.P.D. Both departments need to establish a common understanding and ensure that everyone "talks the same language" and "tears down the walls" between them. For the purpose of this article the short form of S.P.D. will be used for Sterile Processing Department (sometimes known as Central Processing), the department where instrument processing takes place and the distribution center for the entire hospital's supplies. PMID- 15298333 TI - Leaders' role in infection prevention and control: fighting the invisible and invincible... if only... PMID- 15298334 TI - Choosing a reprocessing method. AB - The choice of a reprocessing method depends on many factors and there is no one perfect method that suits all situations. The needs of patients, staff and the healthcare facility, as well as the types of products available at any given time, will all influence the decision. And that decision will not be final. Products change as do consumer needs. As a result the reprocessing decision will need to be reviewed, on a regular basis, in order to ensure that it remains current. PMID- 15298335 TI - Custom procedure packs--the Regina experience. PMID- 15298337 TI - Seasonality in clinical onset of type 1 diabetes in belgian patients above the age of 10 is restricted to HLA-DQ2/DQ8-negative males, which explains the male to female excess in incidence. AB - AIMS/HYPOTHESIS: Type 1 diabetes arises from an interplay between environmental and genetic factors. The reported seasonality at diagnosis supports the hypothesis that currently unknown external triggers play a role in the onset of the disease. We investigated whether a seasonal pattern is observed at diagnosis in Belgian Type 1 diabetic patients, and if so whether seasonality varies according to age, sex and genetic risk, all known to affect the incidence of Type 1 diabetes. METHODS: The seasonal pattern at clinical diagnosis was assessed in 2176 islet antibody-positive diabetic patients aged 0 to 39 years diagnosed between 1989 and 2000. Additional stratification was performed for age, sex and HLA-DQ genotype. RESULTS: Overall, a significant seasonal pattern at clinical diagnosis of diabetes was observed (p<0.001). More subjects were diagnosed in the period of November to February (n=829) than during the period of June to September (n=619) characterised by higher averages of maximal daily temperature and daily hours of sunshine. However, the seasonal pattern was restricted to patients diagnosed above the age of 10 (0-9 years: p=0.398; 10-19 years: p<0.001; 20-29 years: p=0.003; 30-39 years: p=0.015). Since older age at diagnosis is associated with a male to female excess and a lower prevalence of the genetic accelerator HLA-DQ2/DQ8, we further stratified the patients aged 10 to 39 years (n=1675) according to HLA-DQ genotype and sex, and we found that the seasonal pattern was largely restricted to male subjects lacking DQ2/DQ8 (n=748; p<0.00 vs all others: n=927; p=0.031). CONCLUSIONS/INTERPRETATION: In a subgroup of male patients diagnosed over the age of 10, the later stages of the subclinical disease process may be more driven by sex- and season-dependent external factors than in younger, female and genetically more susceptible subjects. These factors may explain the male to female excess in diabetes diagnosed in early adulthood. PMID- 15298336 TI - Transcriptional networks controlling pancreatic development and beta cell function. AB - Transcription factors provide the genetic instructions that drive pancreatic development and enable mature beta cells to function properly. To understand fully how this is accomplished, it is necessary to unravel the regulatory networks formed by transcription factors acting on their genomic targets. This article discusses recent advances in our understanding of how transcriptional networks control early pancreas organogenesis, embryonic endocrine cell formation and the differentiated function of adult beta cells. We discuss how mutations in several transcription factor genes involved in such networks cause Maturity onset diabetes of the young (MODY). Finally, we propose that pancreatic gene programs might be manipulated to generate beta cells or to enhance the function of existing beta cells, thereby providing a possible treatment of different forms of diabetes. PMID- 15298338 TI - Insulin analogues (insulin detemir and insulin aspart) versus traditional human insulins (NPH insulin and regular human insulin) in basal-bolus therapy for patients with type 1 diabetes. AB - AIMS/HYPOTHESIS: The aim of the trial was to compare the efficacy and tolerability of two types of basal-bolus therapy, using either the soluble long acting basal insulin analogue, insulin detemir, in combination with the rapid acting analogue, insulin aspart, or NPH insulin in combination with mealtime regular human insulin. METHODS: In this 18-week, 1:1 randomised, open-labelled, parallel trial, 595 patients with Type 1 diabetes mellitus received insulin detemir or NPH insulin in the morning and at bedtime in combination with mealtime insulin aspart or regular human insulin respectively. RESULTS: Glycaemic control with insulin detemir/insulin aspart was improved in comparison with NPH insulin/regular human insulin (HbA1c: 7.88% vs 8.11%; mean difference: -0.22% point [95% CI: -0.34 to -0.10]; p<0.001). Self-measured 8-point plasma glucose profiles differed between the groups (p<0.001), with lower postprandial plasma glucose levels in the insulin detemir/insulin aspart group. Within-person day-to day variation in plasma glucose was lower with insulin detemir/insulin aspart than with NPH insulin/regular human insulin (SD: 2.88 vs 3.12 mmol/l; p<0.001). Risk of overall and nocturnal hypoglycaemia (23.00-06.00 hours) was, respectively, 21% (p=0.036) and 55% (p<0.001) lower in the insulin detemir/insulin aspart group than in the NPH insulin/regular human insulin group. Body weight (adjusted for baseline and change in HbA1c) was 1 kg lower with insulin detemir/insulin aspart than with NPH insulin/regular human insulin (p<0.001). CONCLUSIONS/INTERPRETATION: Basal-bolus therapy using insulin detemir/insulin aspart offers a better balance of control and tolerability than with NPH insulin/regular human insulin. The low variability and more physiological action profiles generated with these insulin analogues resulted in improved glycaemic control with lower risk of hypoglycaemia and no concomitant body weight increase. PMID- 15298339 TI - Diet but not aerobic exercise training reduces skeletal muscle TNF-alpha in overweight humans. AB - AIMS/HYPOTHESIS: Our aim was to test the hypothesis that TNF-alpha protein levels in skeletal muscle are important in mediating the improvements in glucose homeostasis that are associated with diet and exercise regimens intended to reduce cardiovascular risk. METHODS: We recruited 20 people with a body mass index of 32.1 +/- 1.2 kg/m2 (mean +/- SEM) and one other component of the metabolic syndrome. The average age was 51.2 +/- 8.1 years (mean +/- SD). Of the 20 subjects, 6 were men and 14 were women. All subjects completed an 8-week control period, followed by randomisation to 8 weeks of moderate cycling exercise (30 min, three times per week) or to a diet with the following characteristics: low in saturated fat, high in fibre, low glycaemic index, rich in complex carbohydrates. RESULTS: Diet induced a small reduction in body mass index (3.0 +/ 0.7%, p<0.05), although weight loss was not intended. Exercise training increased maximum oxygen consumption by 12 +/- 6% (p<0.05). Both interventions reduced fasting plasma insulin levels by about 20%. Diet reduced skeletal muscle TNF-alpha protein by 54 +/- 10% (p<0.05), an effect that was independent (p=0.94 in covariate analysis) of the small concurrent weight loss (-2.8 +/- 0.7 kg). Levels of GLUT4 protein were unchanged in the diet group. In contrast, exercise training did not significantly change TNF-alpha protein expression, but GLUT4 protein expression increased by 105 +/- 37% (p<0.05). CONCLUSIONS/INTERPRETATION: These data indicate that the metabolic benefits of a diet aimed at cardiovascular risk reduction are associated with a decrease in skeletal muscle TNF-alpha protein. PMID- 15298340 TI - Genes for systemic vascular complications are differentially expressed in the livers of type 2 diabetic patients. AB - AIMS/HYPOTHESIS: Type 2 diabetes is characterised by excessive hepatic glucose production and frequently leads to systemic vascular complications. We therefore analysed the relationship between the gene expression profile in the liver and the pathophysiology of Type 2 diabetes. METHODS: Liver biopsy samples were obtained from twelve patients with Type 2 diabetes and from nine non-diabetic patients. To assay gene expression globally in the livers of both groups, we made complementary DNA (cDNA) microarrays consisting of 1080 human cDNAs. Relative expression ratios of individual genes were obtained by comparing cyanine 5 labelled cDNA from the patients with cyanine 3-labelled cDNA from reference RNA from the liver of a non-diabetic patient. RESULTS: On assessing the similarities of differentially expressed genes, the gene expression profiles of the twelve diabetic patients formed a separate cluster from those of the non-diabetic patients. Of the 1080 genes assayed, 105 (9.7%) were up-regulated and 134 (12%) were down-regulated in the diabetic livers (p<0.005). The genes up-regulated in the diabetic patients included those encoding angiogenic factors such as vascular endothelial growth factor, endothelin and platelet-derived growth factor. They also included TGF superfamily genes such as TGFA and TGFB1 as well as bone morphogenetic proteins. Among the down-regulated genes in the diabetic patients were molecules defending against stress, e.g. flavin-containing monooxygenase and superoxide dismutase. CONCLUSIONS/INTERPRETATION: These findings suggest that livers of patients with Type 2 diabetes have gene expression profiles indicative of an increased risk of systemic vascular complications. PMID- 15298341 TI - Brain energy metabolism during hypoglycaemia in healthy and type 1 diabetic subjects. AB - AIMS/HYPOTHESIS: This study aimed to examine brain energy metabolism during moderate insulin-induced hypoglycaemia in Type 1 diabetic patients and healthy volunteers. METHODS: Type 1 diabetic patients (mean diabetes duration 13 +/- 2.5 years; HbA1c 6.8 +/- 0.3%) and matched controls were studied before, during (0 120 min) and after (120-240 min) hypoglycaemic (approximately 3.0 mmol/l) hyperinsulinaemic (1.5 mU x kg(-1) min(-1)) clamp tests. Brain energy metabolism was assessed by in vivo 31P nuclear magnetic resonance spectroscopy of the occipital lobe (3 Tesla, 10-cm surface coil). RESULTS: During hypoglycaemia, the diabetic patients showed blunted endocrine counter-regulation. Throughout the study, the phosphocreatine:gamma-ATP ratios were lower in the diabetic patients (baseline: controls 3.08 +/- 0.29 vs diabetic patients 2.65 +/- 0.43, p<0.01; hypoglycaemia: 2.97 +/- 0.38 vs 2.60 +/- 0.35, p<0.05; recovery: 3.01 +/- 0.28 vs 2.60 +/- 0.35, p<0.01). Intracellular pH increased in both groups, being higher in diabetic patients (7.096 +/- 0.010 vs. 7.107 +/- 0.015, p<0.04), whereas intracellular magnesium concentrations decreased in both groups (controls: 377 +/ 33 vs 321 +/- 39; diabetic patients: 388 +/- 47 vs 336 +/- 68 micromol/l; p<0.05). CONCLUSIONS/INTERPRETATION: Despite a lower cerebral phosphocreatine:gamma-ATP ratio in Type 1 diabetic patients at baseline, this ratio does not change in control or diabetic patients during modest hypoglycaemia. However, both groups exhibit subtle changes in intracellular pH and intracellular magnesium concentrations. PMID- 15298342 TI - The effect of exercise on regional adipose tissue and splanchnic lipid metabolism in overweight type 2 diabetic subjects. AB - AIMS/HYPOTHESIS: To test the hypothesis that adipose tissue lipolysis is enhanced in patients with Type 2 diabetes mellitus, we examined the effect of exercise on regional adipose tissue lipolysis and fatty acid mobilisation and measured the acute effects of exercise on the co-ordination of adipose tissue and splanchnic lipid metabolism. METHODS: Abdominal, subcutaneous adipose tissue and splanchnic lipid metabolism were studied by conducting measurements of arterio-venous concentrations and regional blood flow in six overweight Type 2 diabetic subjects before, during and after exercise. RESULTS: Exercise induced an increase in adipose tissue lipolysis and fatty acid release. However, the increase in adipose tissue blood flow was small, limiting fatty acid mobilisation from this tissue. Some of the fatty acids were released in excess in the post-exercise phase. The splanchnic fatty acid uptake was unchanged during the experiment but splanchnic ketogenesis increased in the post-exercise phase. The arterial glucose concentration decreased during exercise and continued to decrease afterwards, indicating an imbalance between splanchnic glucose production and whole-body glucose utilisation. CONCLUSIONS/INTERPRETATION: Regional subcutaneous, abdominal adipose tissue lipolysis is no higher in patients with Type 2 diabetes than in young, healthy subjects. Exercise stimulates adipose tissue lipolysis, but due to an insufficient increase in blood flow, a high fraction of the fatty acids liberated by lipolysis cannot be released to the blood. Splanchnic glucose release is smaller than whole-body glucose utilisation during exercise and post exercise recovery. PMID- 15298343 TI - CD40 expression on human pancreatic duct cells: role in nuclear factor-kappa B activation and production of pro-inflammatory cytokines. AB - AIMS/HYPOTHESIS: Human pancreatic duct cells are closely associated with islet beta cells, and contaminate islet suspensions transplanted in Type 1 diabetes mellitus patients. Activated duct cells produce cytotoxic mediators and possibly contribute to the pathogenesis of Type 1 diabetes mellitus or islet graft rejection. As CD40 transduces activation signals involved in inflammatory and immune disorders, we investigated CD40 expression on duct cells and their response to CD40 engagement. METHODS: CD40 expression on human pancreatic duct cells was analysed by flow cytometry and immunohistochemical analyses. To assess the function of CD40 expression on duct cells, activation of the transcription factor nuclear factor-kappa B was determined using electrophoretic mobility shift assay and ELISA. Cytokine mRNA levels were quantified by real-time RT-PCR, and protein levels by Luminex technology. RESULTS: Isolated human pancreatic duct cells and Capan-2 cell lines were found to express constitutively CD40. The expression of CD40 on duct cells was confirmed in vivo on human normal and pathological pancreatic specimens. CD40 ligation on Capan-2 cells induced rapid nuclear factor-kappa B activation, and supershift assays demonstrated that p50/p65 heterodimers and p50/p50 homodimers were present in the activated complexes in the nucleus. This activation was accompanied by tumour necrosis factor-a and interleukin-1beta mRNA accumulation. Tumour necrosis factor-alpha protein secretion was confirmed in CD40-activated Capan-2 cells and in isolated human pancreatic duct cells. CONCLUSIONS/INTERPRETATION: Interaction between activated T lymphocytes expressing CD40 ligand and duct cells expressing CD40 may contribute to the immune responses involved in Type 1 diabetes mellitus and islet graft rejection. Interfering with CD40-mediated duct cell activation could alleviate beta cell damage of immune origin. PMID- 15298344 TI - Effect of maternal low-protein diet and taurine on the vulnerability of adult Wistar rat islets to cytokines. AB - AIMS/HYPOTHESIS: A maternal low-protein diet has been shown to induce an increased susceptibility of fetal islets to cytokines, but this effect can be avoided by maternal taurine supplementation. Here, we question whether these effects persist until adulthood in the offspring, despite the animal having a normal diet after weaning. METHODS: Pregnant Wistar rats received a diet of either 20% or 8% protein (control [C group] and recuperated [R group] respectively), which was or was not supplemented with taurine (control treated with taurine [CT group] and recuperated treated with taurine [RT group] respectively) during gestation and lactation. When the female offspring reached adulthood, an OGTT was performed. In a second stage, islets were isolated from these offspring, then pretreated or not with taurine, and subsequently treated with cytokines. RESULTS: Fasting glycaemia was higher (p<0.05) and insulinaemia was lower (p<0.01) in the R group than in the C group. Taurine supplementation decreased insulinaemia in the CT group and tended to increase it in the RT group. After the OGTT, glycaemia in R animals was not different from that in the C group, despite a blunted insulin response (p<0.05) which was restored by taurine. Supplementation in C-group mothers led to a weak glucose intolerance. In vitro, more apoptotic cells were observed in R islets after cytokines treatment (p<0.01). The addition of taurine to the culture medium in the R and C groups protected the islets from the cytokines (p<0.01). Maternal taurine supplementation decreased the sensitivity of islets in the RT group (p<0.01), but increased sensitivity in the CT group (p<0.01). CONCLUSIONS/INTERPRETATION: The increased vulnerability of islets to cytokines due to a restriction of protein during fetal development was still evident when the offspring reached adulthood. The low-protein diet also induced hyperglycaemia in the presence of lower insulinaemia. Taurine supplementation protected adult islets of the R group from cytokine toxicity and restored the insulinaemia. However, unnecessary supplementation of taurine could have detrimental effects. PMID- 15298345 TI - Generation of hydrogen peroxide and failure of antioxidative responses in pancreatic islets of male C57BL/6 mice are associated with diabetes induced by multiple low doses of streptozotocin. AB - AIMS/HYPOTHESIS: We studied the impact of the reactive oxygen species hydrogen peroxide (H2O2) and antioxidative enzymes on the pathogenesis of diabetes induced by multiple low doses of streptozotocin (MLD-STZ). METHODS: We isolated the islets of C57BL/6 mice. For ex vivo analyses, mice had been injected with MLD STZ. For in vitro analyses, islets were incubated with different concentrations of STZ, with either of the two moieties of STZ, methylnitrosourea and D-glucose, with H2O2 or with alloxan. Levels of H2O2 generation were measured by the scopoletin method. We assessed mRNA expression of Cu/Zn and Mn superoxide dismutase, catalase, and glutathione peroxidase (GPX) by semiquantitative polymerase chain reaction. GPX activity was measured spectrophotometrically. In vitro, beta cell function was assayed by measuring basal and D-glucose-stimulated release of immunoreactive insulin using an ELISA kit. RESULTS: Ex vivo, MLD-STZ significantly increased H2O2 generation in male but not in female mice. It also increased GPX activity and mRNA expression of catalase, Cu/Zn and Mn superoxide dismutase, and GPX in female but not in male mice. In vitro, STZ significantly stimulated H2O2 generation in islets of male mice only. In male islets, alloxan increased H202 generation at a highly toxic concentration, but D-glucose and methylnitrosourea did not. Both STZ and H2O2 dose-dependently inhibited the release of immunoreactive insulin after a D-glucose challenge. CONCLUSIONS/INTERPRETATION: The results indicate that H2O2 participates in the pathogenesis of MLD-STZ diabetes in male C57BL/6 mice, which do not up-regulate antioxidative enzymes in islets. Conversely, female mice are protected, probably due to an increment of several enzymes with the potential to detoxify H2O2. PMID- 15298346 TI - Pancreatic islets from cyclin-dependent kinase 4/R24C (Cdk4) knockin mice have significantly increased beta cell mass and are physiologically functional, indicating that Cdk4 is a potential target for pancreatic beta cell mass regeneration in Type 1 diabetes. AB - AIMS/HYPOTHESIS: Cyclin-dependent kinase 4 (Cdk4) is crucial for beta cell development. A mutation in the gene encoding for Cdk4, Cdk4R24C, causes this kinase to be insensitive to INK4 cell cycle inhibitors and induces beta cell hyperplasia in Cdk4R24C knockin mice. We aimed to determine whether this Cdk4R24C mutation also affects proper islet function, and whether it promotes proliferation in human islets lentivirally transduced with Cdk4R24C cDNA. METHODS: Our study was conducted on wild-type and Cdk4R24C knockin mice. Pancreases were morphometrically analysed. Intraperitoneal glucose tolerance tests and intravenous insulin tolerance tests were performed on wild-type and Cdk4R24C mice. We also did in vitro islet perifusion studies and islet metabolic labelling analysis. Human islets were transduced with Cdk4R24C cDNA. RESULTS: Pancreatic islets from Cdk4R24C knockin mice exhibit a larger insulin-producing beta cell area and a higher insulin content than islets from wild-type littermates. Insulin secretion in response to glucose is faster and reaches a higher peak in Cdk4R24C mice without leading to hypoglycaemia. Conversion of pro insulin into insulin and its intermediates is similar in Cdk4R24C and wild-type mice. Glucose utilisation and oxidation measured per islet were similar in both experimental groups. Insulin secretion was faster and enhanced in Cdk4R24C islets perifused with 16.7 mmol/l glucose, with slower decay kinetics when glucose returned to 2.8 mmol/l. Moreover, human islets expressing Cdk4R24C cDNA exhibited higher beta cell proliferation. CONCLUSIONS/INTERPRETATION: Despite their hyperplastic growth, Cdk4R24C insulin-producing islet cells behave like differentiated beta cells with regard to insulin production, insulin secretion in response to glucose, and islet glucose metabolism. Therefore Cdk4 could possibly be used to engineer a source of beta cell mass for islet transplantation. PMID- 15298347 TI - Vascular endothelial cadherin and beta-catenin in human fetoplacental vessels of pregnancies complicated by Type 1 diabetes: associations with angiogenesis and perturbed barrier function. AB - AIMS/HYPOTHESIS: Increased angiogenesis of fetoplacental vessels is a feature of pregnancies complicated by Type 1 diabetes mellitus, but the underlying molecular mechanisms are unknown. This investigation tests whether the diabetic maternal environment alters the phenotypic expression of placental vascular endothelial cadherin and beta-catenin, which have been implicated as key molecules in barrier formation and angiogenesis in the endothelium. METHODS: Term placental microvessels from normal pregnancies (n=8) and from those complicated by Type 1 diabetes (n=8) were perfused with 76-Mr dextran tracers (1 mg/ml) and subjected to immunocytochemistry, immunoblotting and microscopy. Junctional integrity, localisation and phosphorylation were investigated along with total protein levels of vascular endothelial cadherin, beta-catenin and vascular endothelial growth factor. Stereological sampling and estimation tools were used to quantify aspects of angiogenesis and endothelial proliferation. RESULTS: In the Type 1 diabetic placentae, junctional localisations of vascular endothelial cadherin and beta-catenin altered significantly, with more than 50% of microvessels showing complete loss of immunoreactivity and with no overall loss of total protein. Tracer leakage was associated with these vessels. There was a two- to three-fold increase in vessels showing junctional phospho-tyrosine immunoreactivity and hyperphosphorylated beta-catenin. Vascular endothelial growth factor levels were higher in these placentae. A four-fold increase in endothelial proliferation was observed, along with an increase in total length of capillaries without any change in luminal diameter. CONCLUSIONS/INTERPRETATION: Molecular perturbations of vascular endothelial cadherin and beta-catenin occur in fetoplacental vessels of pregnancies complicated by Type 1 diabetes. Phosphorylation and loss of these molecules from the adherens junctional domains may be influenced in part by the elevated levels of vascular endothelial growth factor in the placenta. Perturbations of the junctional proteins may explain the observed breach in barrier integrity and may contribute to the mechanisms that drive proliferation and increases in capillary length. PMID- 15298348 TI - Evaluation of orally active poly(ADP-ribose) polymerase inhibitor in streptozotocin-diabetic rat model of early peripheral neuropathy. AB - AIMS/HYPOTHESIS: Poly(ADP-ribose) polymerase activation depletes NAD+ and high energy phosphates, activates protein kinase C, and affects gene expression in various tissues. This study was designed to characterise the effects of the potent, orally active poly(ADP-ribose) polymerase inhibitor PJ34 in the Wistar rat model of early diabetic neuropathy. METHODS: Control and streptozotocin diabetic rats were maintained with or without PJ34 treatment (30 mg x kg(-1) x day(-1)) for two weeks, after two weeks without treatment. Endoneurial blood flow was assessed by hydrogen clearance; metabolites and high-energy phosphates were assayed by enzymatic spectrofluorometric methods; and poly(ADP-ribose) was detected by immunohistochemistry. RESULTS: Blood glucose concentrations were increased to a similar extent in untreated and PJ34-treated diabetic rats compared with controls. Intense poly(ADP-ribose) immunostaining was observed in the sciatic nerve of diabetic rats, but not in other groups. Final sciatic motor nerve conduction velocity and digital sensory nerve conduction velocity were reduced by 24% and 22% respectively in diabetic rats compared with controls (p<0.01 for both), and both were 98% corrected by PJ34 (p<0.01 vs diabetic group for both). In contrast, with PJ34 treatment, nerve blood flow showed a modest (17%) increase, and vascular conductance showed a tendency to increase. Free mitochondrial and cytosolic NAD+:NADH ratios, assessed from the glutamate and lactate dehydrogenase systems, phosphocreatine concentrations, and phosphocreatine:creatine ratios were decreased in diabetic rats and essentially normalised by PJ34. In both untreated and PJ34-treated diabetic rats, nerve glucose, sorbitol and fructose were increased to a similar extent. PJ34 did not affect any variables in control rats. CONCLUSIONS/INTERPRETATION: Short-term poly(ADP-ribose) polymerase inhibitor treatment reverses functional and metabolic abnormalities of early diabetic neuropathy. Complete normalisation of nerve blood flow is not required for correction of motor or sensory nerve conduction velocities, provided that a therapeutic agent can restore nerve energy state via direct action on Schwann cells. PMID- 15298349 TI - Prevention of sensory disorders in diabetic Sprague-Dawley rats by aldose reductase inhibition or treatment with ciliary neurotrophic factor. AB - AIMS/HYPOTHESIS: Sensory neuropathy in diabetic patients frequently presents itself as progressive loss of thermal perception, while some patients describe concurrent spontaneous pain, allodynia or hyperalgesia. Diabetic rats develop thermal hypoalgesia and tactile allodynia by unknown mechanisms. We investigated whether sensory disorders in rats were related to glucose metabolism by aldose reductase. We also explored the therapeutic potential of exogenous neurotrophic factors. METHODS: Behavioural assessments of thermal and tactile sensitivity were performed in normal rats and in rats with streptozotocin-induced diabetes. Some of the rats were treated with insulin, aldose reductase inhibitors, ciliary neurotrophic factor or brain-derived neurotrophic factor. RESULTS: Thermal hypoalgesia was present after 8 weeks of diabetes and was prevented by insulin treatment, which maintained normoglycaemia, by the aldose reductase inhibitor Statil or by ciliary neurotrophic factor. Brain-derived neurotrophic factor did not have an effect. When diabetic rats were tested after shorter durations of diabetes, they showed transient thermal hyperalgesia after 4 weeks which progressed to thermal hypoalgesia after 8 weeks. The aldose reductase inhibitor IDD 676 (Lidorestat), given from the onset of diabetes, prevented the development of thermal hyperalgesia and also stopped progression to thermal hypoalgesia when delivered in the last 4 weeks of an 8-week period of diabetes. Tactile allodynia was not prevented by neurotrophic factor or aldose reductase inhibitor treatment. CONCLUSIONS/INTERPRETATION: Transient thermal hyperalgesia and subsequent progressive thermal hypoalgesia occur in diabetic rats secondary to exaggerated flux through the polyol pathway. A depletion of ciliary neurotrophic factor mediated by the polyol pathway may be involved in the aetiology of thermal hypoalgesia. PMID- 15298350 TI - Diabetic background retinopathy is associated with impaired coronary vasoreactivity in people with Type 1 diabetes. AB - AIMS/HYPOTHESIS: We examined whether diabetic background retinopathy is associated with reduced coronary vasoreactivity in people with Type 1 diabetes. METHODS: A total of 21 men with Type 1 diabetes were investigated, including 9 men with background retinopathy and 12 men without retinopathy. In addition, 12 non-diabetic, age-matched subjects were studied. All subjects were non-smokers, otherwise healthy and had no other diabetic complications. Resting myocardial blood flow and hyperaemic dipyridamole-stimulated flow (dipyridamole, 0.56 mg/kg during a 4-min period), a measure of coronary vasoreactivity, were measured during euglycaemic hyperinsulinaemic clamp (1 mU x kg(-1) x min(-1)) using positron emission tomography and oxygen-15-labelled water. RESULTS: Resting myocardial blood flow (0.82 +/- 0.13 vs 0.96 +/- 0.23 vs 0.88 +/- 0.25 ml x g(-1) x min(-1), with vs without retinopathy vs non-diabetic subjects) and coronary vascular resistance (111.2 +/- 23.4 vs 95.5 +/- 15.8 vs 101.9 +/- 31.5 mmHg x min x g x ml(-1) respectively) were not significantly different between the groups. Dipyridamole infusion induced an increase in blood flow and a decrease in coronary vascular resistance in all study subjects (p<0.001). However, dipyridamole-stimulated flow and coronary vascular resistance were blunted in diabetic patients with retinopathy (2.9 +/- 0.9 ml x g(-1) x min(-1) and 34.1 +/- 11.3 mmHg x min x g x ml(-1)) when compared to diabetic patients without retinopathy (4.0 +/- 1.3 ml x g(-1) x min(-1), p=0.04 and 24.6 +/- 7.5 mmHg x min x g x ml(-1), p=0.03) or non-diabetic subjects (4.5 +/- 1.4 ml x g(-1) x min(-1) p=0.008 and 22.2 +/- 8.7 mmHg x min x g x ml(-1), p=0.01). Myocardial flow reserve was impaired in diabetic patients with retinopathy (3.6 +/- 1.0) when compared to non-diabetic subjects (5.3 +/- 1.9, p=0.02) but not significantly reduced when compared to diabetic patients without retinopathy (4.2 +/- 1.4, p=0.2). CONCLUSIONS/INTERPRETATION: Diabetic background retinopathy appears to be associated with impaired coronary vasoreactivity in young people with Type 1 diabetes. PMID- 15298351 TI - Heritability estimates for beta cell function and features of the insulin resistance syndrome in UK families with an increased susceptibility to type 2 diabetes. AB - AIMS/HYPOTHESIS: The aim of this study was to measure the heritability estimates for metabolic traits and the features of the insulin resistance syndrome in families with an increased genetic susceptibility to Type 2 diabetes. METHODS: A total of 811 non-diabetic relatives from 278 pedigrees of northern European extraction in which there was a sib-pair with Type 2 diabetes were recruited and studied at the six Diabetes UK Warren Type 2 diabetes centres. Heritability estimates were calculated, allowing for key covariates (age, sex, BMI and recruitment centre). Values greater than 0.10 were considered statistically significant in comparison to zero. RESULTS: Fasting glucose concentration and homeostasis model assessment of pancreatic beta cell function (HOMA %B) had the highest heritability estimates of 0.72 and 0.78 respectively. Heritability estimates for the features of the insulin resistance syndrome (BMI, WHR, systolic and diastolic blood pressure, serum lipids and homeostasis model assessment of insulin sensitivity [HOMA %S]) were also high. The heritability estimate for fasting glucose was markedly higher in the present study (0.77 vs 0.21 adjusted for age and sex; p<0.001) than in a comparable study of families from the same background population but with no increased susceptibility to diabetes. However, the estimates for the features of the insulin resistance syndrome were similar in the two studies. CONCLUSIONS/INTERPRETATION: In families with a high risk of Type 2 diabetes, the heritability estimates for fasting glucose, pancreatic beta cell function and the features of the insulin resistance syndrome were all high. The higher heritability estimate for pancreatic beta cell function suggests that this resource may be most effective when investigating genetic susceptibility to beta cell dysfunction. PMID- 15298352 TI - Allelic variation in class I K gene as candidate for a second component of MHC linked susceptibility to type 1 diabetes in non-obese diabetic mice. AB - AIMS/HYPOTHESIS: Recent studies have revealed that MHC-linked susceptibility to Type 1 diabetes is determined by multiple components. In the non-obese diabetic (NOD) mouse, a second component (Idd16) has been mapped to a region adjacent to, but distinct from Idd1 in the class II region. In this study, we investigated the class I K gene as a candidate gene for Idd16. METHODS: We determined the genomic sequences of the class I K gene as well as the reactivity of K molecules with monoclonal antibodies in the NOD mouse, the Cataract Shionogi (CTS) mouse, and the NOD.CTS-H-2 congenic strain, which possesses a resistance allele to Type 1 diabetes at the Idd16 on the NOD genetic background genes. RESULTS: While the K sequence of the NOD mouse was identical to that of Kd type, ten nucleotide substitutions were identified in the CTS mouse compared with the NOD mouse. Of these, three were in exon 4, giving two amino acid substitutions, which were identical to those seen in KK type. These characteristics were retained in the NOD.CTS-H-2 congenic strain, which had a lower incidence and delayed onset of Type 1 diabetes owing to a resistance allele at Idd16. Lymphocytes from NOD.CTS H2 congenic mice reacted with anti-Kd and anti-Kk monoclonal antibodies, reflecting the unique sequence of the K gene. The nucleotide sequence of the K gene in the non-obese non-diabetic (NON) mouse was also unique, consisting of a combination of Kk- and Kb-like sequences. CONCLUSIONS/INTERPRETATION: These data suggest that H2-K is unique in CTS and NON mice, and that allelic variation of the class I K gene may be responsible for Idd16. PMID- 15298353 TI - Paradoxical effects of insulin on cardiac L-type calcium current and on contraction at physiological temperature. AB - AIMS/HYPOTHESIS: L-type calcium current (I(Ca,L)) is a major determinant of mammalian cardiac contraction, and data from studies performed at room temperature suggest that this current is stimulated by insulin. This investigation aimed to determine whether or not insulin stimulates cardiac I(Ca,L) at 37 degrees C. METHODS: Isolated guinea pig ventricular myocytes were studied at room temperature and at 37 degrees C. Myocytes were either field stimulated or whole-cell voltage clamped, and cell shortening was measured using video edge detection. RESULTS: Insulin stimulated I(Ca,L) at ambient temperature. However, at 37 degrees C the effect of insulin was to decrease rather than to increase I(Ca,L). This action was concentration dependent and was not associated with voltage shifts in steady-state activation or inactivation properties of I(Ca,L). At 37 degrees C, insulin increased the extent of myocyte contraction despite producing a significant decrease in I(Ca,L) amplitude. CONCLUSIONS/INTERPRETATION: The findings of this study indicate that temperature is a key experimental variable in the study of the physiological actions of insulin. Furthermore, the increase in cardiac cell contraction by insulin at physiological temperature is not due to an increase in I(Ca,L), but is probably due to stimulation of excitation-contraction coupling downstream of I(Ca,L). PMID- 15298354 TI - Familial partial lipodystrophy associated with compound heterozygosity for novel mutations in the LMNA gene. PMID- 15298355 TI - Lack of association between metabolic traits and the -512 polymorphism in FOXC2 in german people with normal glucose tolerance. PMID- 15298356 TI - Who are more insulin resistant, people with IFG or people with IGT? PMID- 15298357 TI - The Gly1057Asp polymorphism in IRS-2 interacts with obesity to affect beta cell function. PMID- 15298358 TI - -to: Tschoepe D, Menart B, Ferber P et al. (2003) genetic variation of the platelet surface integrin GPIIb-IIIa (PIA1/A2-SNP) shows a high association with Type 2 diabetes mellitus. Diabetologia 46:984-989. PMID- 15298359 TI - -to: Gardiner TA, Anderson HR, Degenhardt T et al. (2003) prevention of retinal capillary basement membrane thickening in diabetic dogs by a non-steroidal anti inflammatory drug. Diabetologia 46:1269-1275. PMID- 15298360 TI - "Prediction and early detection of diabetic complications: common pathways and unique needs". Keble College, Oxford Austust 1-4, 2003. PMID- 15298361 TI - Using guidelines to improve care of patients with chronic kidney disease. PMID- 15298362 TI - Cost impacts of illness on families and society. PMID- 15298363 TI - Potential application of the National Kidney Foundation's chronic kidney disease guidelines in a managed care setting. AB - Chronic kidney disease (CKD) is a public health problem in the United States with rising incidence and prevalence. The disease may progress to end-stage renal disease, which is associated with significant morbidity and mortality. The National Kidney Foundation recently established clinical practice guidelines to aid in the identification and stratification of CKD. Central to these guidelines is the use of glomerular filtration rate estimation equations. Integrated healthcare systems can use the guidelines to identify patients with CKD early in its course. Early identification allows for timely nephrology referral, institution of measures to slow the progression of CKD, and treatment of CKD specific complications. Administration of these efforts early in the course of CKD improves subsequent morbidity and mortality of affected patients. By identifying CKD early in its course, patients can be referred to a coordinated case management team that can help administer care. The team can follow up a patient from the early diagnosis of CKD to end-stage renal disease and renal replacement therapy. Finally, continuous quality improvement can be instituted to assist in improving patient care. PMID- 15298364 TI - Genetic testing and pharmacogenomics: issues for determining the impact to healthcare delivery and costs. AB - OBJECTIVES: To determine the potential impact of genetic testing and pharmacogenomics on healthcare delivery and costs. STUDY DESIGN: Literature review. METHODS: We examined 3 examples: (1) BRCA1/2 testing for breast cancer risk, (2) HER2/neu overexpression testing to guide drug treatment in women with breast cancer, and (3) CYP2C9 testing before the use of the anticoagulant warfarin. We discussed each genetic testing example from the perspective of the patient, provider, insurer, industry, government, and society. RESULTS: The expanded use of genetic information offers many potential clinical benefits, but also many economic challenges. One of those challenges will be managing the impact of genetic testing on healthcare delivery and costs. CONCLUSIONS: Systematic, evidence-based technology assessments and economic evaluations will have to be used to guide the incorporation of genomics into clinical practice. More research also will be needed to assess patient preferences and willingness to pay for genomic technologies; how providers can assess and use genomic technologies; and how the industry, insurers, and government can best balance the relevant costs and benefits. PMID- 15298365 TI - Prescribing proton pump inhibitors for initial treatment of acid-related gastrointestinal diseases in a managed care population. AB - OBJECTIVE: To determine whether health services use by patients with selected acid-related gastrointestinal disorders (peptic ulcer disease, gastroesophageal reflux disease, and gastritis or dyspepsia) is lower after initial treatment with proton pump inhibitors (PPIs) than with histamine2 receptor antagonists (H2RAs). STUDY DESIGN: Retrospective, 2-year longitudinal study. PATIENTS AND METHODS: Among continuous enrollees from December 1, 1996, to June 1, 2002, in a group model health maintenance organization, 13,971 members were electronically selected who began receiving antisecretory therapy during that period and who had no previous drug therapy, endoscopy, or hospitalization for gastrointestinal disease. Adjusted medical costs and healthcare use related to gastrointestinal disease (measured by office visits, endoscopy, or imaging and hospital admissions) and factors associated with initial and subsequent drug therapy were analyzed using a 2-stage model. This method adjusted for unobservable confounders, primarily drug selection bias, an inherent limitation of retrospective database studies. RESULTS: Drug costs were more than 4-fold higher (P < .001) when PPIs rather than H2RAs were prescribed initially, but non-drug costs and health services use showed no decrease. A history of physicians' prescribing PPIs in the prior 12 months was associated with prescribing PPIs as initial therapy (odds ratio, 4.29; 95% confidence interval, 3.74-4.90) and with step-up therapy (change from H2RAs to PPIs). A history of physicians' prescribing H2RAs in the prior 12 months was associated with step-down therapy (change from PPIs to H2RAs). CONCLUSION: Prescribing PPI compared with H2RA therapy as initial therapy for acid-related gastrointestinal disease produced no decrease in nondrug costs or health services use. PMID- 15298366 TI - Medical management of osteoporosis. AB - Osteoporosis is a cause of considerable morbidity and mortality in men and women. Medical intervention can reduce the progression of osteoporosis and decrease the fracture risk associated with low bone mineral density. In this article, we review the evidence for medical therapies for osteoporosis, including estrogen, calcitonin, bisphosphonates, selective estrogen receptor modulators, and the newest approved agent, recombinant human parathyroid hormone. We also discuss several controversial areas in osteoporosis treatment, including the management of men with osteoporosis, approach to monitoring the effects of osteoporosis therapy, and need for cost-effective strategies for osteoporosis treatment. PMID- 15298367 TI - Plan-sponsor savings and member experience with point-of-service prescription step therapy. AB - OBJECTIVE: To examine the effect of prescription step-therapy programs in terms of plan-sponsor savings and member experience at the point of service. STUDY DESIGN: Plan-sponsor savings were measured using a quasi-experimental, case control design. Member experience with step therapy was measured using a self administered mailed survey. METHODS: A 20,000-member plan implemented 3 step therapy programs in September 2002: proton pump inhibitors, selective serotonin reuptake inhibitors, and nonsteroidal anti-inflammatory drugs. Pharmacy claims from September 1, 2001, through June 30, 2003, were examined to compare changes in per-member-per-month (PMPM) net cost between the intervention group and a random sample of members from commercial plans without the step therapy programs. A mailed, self-administered survey was sent to members with a step edit from September 1, 2002 to December 31, 2002. RESULTS: The employer experienced a decrease of 0.83 dollars in net cost after implementing step therapy, while the comparison group had an upward trend of 0.10 dollars PMPM for these therapy classes. Member-reported outcomes indicated that approximately 30% of patients received a generic, 23% were granted a medical exception for the brand, 17% received no medication, and 16% paid the full retail price for the brand. If the pharmacist vs the patient contacted the physician, members were 8 times more likely to receive a medication covered by the health plan (OR, 8.10; 95% CI, 2.94 22.33 vs OR, 8.23; 95% CI, 3.11-21.93). Compared with those who received first line therapy, those who paid out of pocket for the brand medication vs those who did not receive any medication were less likely to be satisfied with their pharmacy benefit (OR, 0.25; 95% CI, 0.08-0.80 vs OR, 0.12; 95% CI, 0.04-0.41). CONCLUSIONS: Step therapy produces significant drug savings. However, there appear to be opportunities to further members' and providers' understanding of these programs. PMID- 15298368 TI - Sick, and tired of the endless paperwork. PMID- 15298369 TI - As the shadows fell. The story of Ronald Reagan's last decade is at once grim and tender. The personal history of how Nancy coped with hie Alzheimer's. PMID- 15298370 TI - 'God has a plan,' my dad always said. PMID- 15298371 TI - Nancy's next campaign. The former First Lady's passion for stem-cell research has fueled a political battle. Where does the science stand? PMID- 15298372 TI - The keys to caregiving. PMID- 15298373 TI - Reagan's last political gift. PMID- 15298374 TI - A tortured debate. Amid feuding and turf battles, lawyers in the White House discussed specific terror-interrogation techniques like 'water-boarding' and 'mock burials'. PMID- 15298375 TI - A high dose of tech. PMID- 15298377 TI - Vanishing minds. PMID- 15298376 TI - 2004: A medical odyssey. PMID- 15298378 TI - Type H superconductors and the vortex lattice. AB - Superconducting material is used, for example, in magnetic resonance imaging for medical examinations and particle accelerators in physics. Knowledge about superfluid liquids can give us deeper insight into the ways in which matter behaves in its lowest and most ordered state. Work by the author on superconduction in liquid helium established the existence of type II superconductors and proved that vortex lattices exist in superfluid helium, in the presence of magnetic fields. He showed that the Ginzburg-Landau theory could be extended to include this "new" type of superconductors, which today are in common use. His work on phase transitions of these superconductors under the influence of magnetic fields was groundbreaking, although he has worked in many other areas since then. He was awarded the Nobel prize in 2003 "for pioneering contributions to the theory of superconductors and superfluids" with V L. Ginzburg and A. J. Leggett. PMID- 15298379 TI - On superconductivity and superfluidity (what I have and have not managed to do), as well as on the 'physical minimum' at the beginning of the 21 st century. AB - The question of thermeolectric effects in superconductors is still a particular problem, which evidently emerges only in the presence of a temperature gradient. The well-known Londons theory yielded much, and is widley employed under certain conditions even nowadays, but is absolutely insufficient and has to be generalized. This problem was solved in the psi-theory of superconductivity by V.L. Ginzburg and L.D. Landau. Together they developed a phenomenological theory of superconductivity in the late 1940s. This theory proposes that those electrons that contribute to superconduction form a superfluid. The superconductor is described by a complex function psi called the order parameter, and /psi/ indicates the fraction of electrons that has condensed into a superfluid. In his Nobel lecture V. L. Ginzburg also gives a 'list' of top problems in contemporary physics. Acquaintance with all subjects included in this 'list' is what he calls the 'physical minimum'. PMID- 15298380 TI - 2003 Nobel Prize in physics for theoretical work on superfluid 3He. AB - The element helium comes in two (stable) forms, 4He and 3He; at low temperatures and pressures both form liquids rather than solids. The liquid phase of the common isotope, 4He, was realized nearly a century ago, and since 1938 has been known to show, at temperatures below about 2K, the property of superfluidity--the ability to flow through the narrowest capillaries without apparent friction. The light isotope, 3He, is believed to be of quite a different nature; however, because of its similarity to the electrons in metals, which at low temperatures sometimes form "Cooper pairs" and thereby become superconducting, theorists in the 1960s and early 1970s had speculated that something similar might happen in liquid 3He, which would then also show superfluidity though for reasons rather different than 4He. In 1972 nuclear magnetic resonance (NMR) experiments at Cornell University revealed the existence, below 3 millidegrees, if two new phases, one of which displayed extraordinary NMR properties. Anthony Leggett is one of the theorists who succeeded in fitting the experimental properties into the "Cooper-pairing" scenario; in particular, he explained the NMR behavior and predicted further novel NMR phenomena which were subsequently found. PMID- 15298381 TI - Direct measurement of dynamic frequency shift induced by cross-correlations in 15N-enriched proteins. AB - We describe a new NMR experimental scheme that allows the direct determination of the dynamic frequency shift induced by chemical shift anisotropy/dipolar interaction (CSA/DD) cross-correlations in 15N-enriched proteins. Its principle consists of comparing two rates of polarisation transfer between the amide proton and nitrogen. The first rate, which is independent of the dynamic frequency shift, is based on a selective Hartmann-Hahn coherence transfer. The second rate, which depends on the dynamic frequency shift, is based on a free evolution of the transverse magnetisation. We report experimental validation of this approach by measuring the average dynamic frequency shift due to CSA/DD cross-correlations in the calcium-binding protein D9k. The method may also be applicable to the measurement of dynamic frequency shift induced by cross-correlations between the Curie spin and dipolar interactions. PMID- 15298382 TI - Two-dimensional double-quantum 2H NMR spectroscopy in the solid state under OMAS conditions: correlating 2H chemical shifts with quasistatic line shapes. AB - Solid-state 2H NMR spectroscopy is a well-established and versatile method to study molecular orientation and dynamics in selectively deuterated samples. Herein, we introduce a 2D 2H double-quantum (DQ) NMR experiment performed under fast magic-angle spinning with a slight offset of the magic angle (OMAS). The experiment combines 2H chemical-shift resolution with DQ-filtered quasistatic 2H line shapes. In this way, it is possible to separate 2H resonances and to independently determine 2H quadrupole couplings at multiple sites. While 2H chemical shifts are resolved in the 2H DQ dimension, the quadrupole parameters can be obtained from characteristic line shapes which are reintroduced in the second dimension by the magic-angle offset. The 2D 2H DQ OMAS experiment is demonstrated on L-histidine which was deuterated at multiple sites by recrystallisation from D2O. PMID- 15298383 TI - On the bond-stretch isomerism in the benzo[1,2:4,5]dicyclobutadiene system--an ab initio MR-AQCC study. AB - Bond-stretch isomerism in benzo[1,2:4,5]dicyclobutadienle (BDCB) has been investigated using the MR-AQCC/6-31G(d) method, a high-level multireference ab initio approach including size-extensivity corrections. The applied theoretical approach includes both nondynamical and dynamical electron correlation effects. Full MR-AQCC geometry optimizations of localized (1) and delocalized (3) isomers as well as the transition structure (TS) have been determined using D2h, symmetry restriction. The calculations show that both isomers are approximately of equal stability separated by a barrier with a height of about 5 kcal mol(-1). Thus, the present results strongly indicate that benzof[1,3:4,5]dicyclobutadiene is a very good candidate for an organic compound exhibiting bond-stretch isomerism, since isomers 1 and 3 correspond to true minima on the double-well potential energy surface, which are separated by a sufficiently high barrier. It is particularly important to emphasize that isomer 3 represents a realization of the highly elusive quasi-[10]annulene. PMID- 15298384 TI - Two-photon absorption in linear bis-dioxaborine compounds-the impact of correlation-induced oscillator-strength redistribution. AB - Quantum-chemical calculations of the two-photon absorption (TPA) cross-sections are used to determine the characteristics of the electronic excited states responsible for the observed peaks in the TPA spectra of two bis-dioxaborine substituted biphenyl derivatives. We find two distinct TPA-active states with very different TPA cross-sections: the difference is explained on the basis of electron correlation. These effects, on the one hand, lead to TPA cross-sections of up to 500 x 10(-50) cm4 s photon(-1) for the state favored by correlation; on the other hand, they limit the overall cross-sections achievable in this class of materials. PMID- 15298385 TI - Scanning force microscopy based rapid force curve acquisition on supported lipid bilayers: experiments and simulations using pulsed force mode. AB - In situ pulsed force mode scanning force microscopy (PFM-SFM) images of phase separated solid-supported lipid bilayers are discussed with the help of computer simulations. Simultaneous imaging of material properties and topography in a liquid environment by means of PFM-SFM is severely hampered by hydrodynamic damping of the cantilever. Stiffness and adhesion images of solid-supported membranes consisting of cholesterol, sphingomyelin, and 1,2-dioleyl phosphatidylcholine obtained in aqueous solution exhibit contrast inversion of adhesion and stiff. ness images depending on parameters such as driving frequency, amplitude, and trigger setting. Simulations using a simple harmonic oscillator model explain experimental findings and give a deeper insight into the way PFM-SFM experiments have to be performed in order to obtain interpretable results and hence pave the way for reliable material contrast imaging at high speed. PMID- 15298386 TI - Surfactant-directed polypyrrole/CNT nanocables: synthesis, characterization, and enhanced electrical properties. AB - We describe here a new approach to the synthesis of size-controllable polypyrrole/carbon nanotube (CNT) nanocables by in situ chemical oxidative polymerization directed by the cationic surfactant cetyltrimethylammonium bromide (CTAB) or the nonionic surfactant polyethylene glycol mono-p-nonylphenyl ether (Opi-10). When carbon nanotubes are dispersed in a solution containing a certain concentration of CTAB or Opi-10, the surfactant molecules are adsorbed and arranged regularly on the CNT surfaces. On addition of pyrrole, some of the monomer is adsorbed at the surface of CNTs and/or wedged between the arranged CTAB or Opi-10 molecules. When ammonium persulfate (APS) is added, pyrrole is polymerized in situ at the surfaces of the CNTs (core layer) and ultimately forms the outer shell of the nanocables. Such polypyrrole/CNT nanocables show enhanced electrical properties; a negative temperature coefficient of resistance at 77-300 K and a negative magnetoresistance at 10-200 K were observed. PMID- 15298387 TI - Amorphous silica as a versatile supermolecular ligand for Ni(II) amine complexes: toward interfacial molecular recognition. AB - Selective adsorption of Ni(II) amine complexes used as precursors for supported catalysts was studied on amorphous silica surfaces. The nature of the adsorption sites was probed by [Ni(en)(dien) (H2O)]2+, [Ni(en)2(H2O)2]2+, and [Ni(dien)(H2O)3]2+ (en = ethylenediamine, dien = diethylenetriamine), which respectively contain one, two, and three labile aqua ligands. The silica surface acts as a mono- or polydentate ligand that can substitute the aqua ligands of the Ni(II) complexes in an inner-sphere adsorption mechanism. Room-temperature adsorption isotherms indicate that each nickel complex selects a limited number of adsorption sites; different sites are recognised by the three complexes, even though they have the same charge and comparable sizes. Several spectroscopic techniques (UV/Vis/NIR, EXAFS, and 29Si NMR) were used to confirm the selective character of the interaction of Ni(II) amine complexes with the silica surface. The specific sites include both silanol/silanolate groups in the same number as the original labile ligands and other surface groups that probably act as hydrogen-bond acceptors. These two types of groups cooperate to result in interfacial molecular-recognition phenomena with interactional complementarity. PMID- 15298388 TI - Microsolvation of phthalocyanines in superfluid helium droplets. AB - Experimental and theoretical investigations of the spectroscopy of molecules in superfluid helium droplets provide evidence for the key role of the first helium layer surrounding the dopant molecule in determining the molecule's spectroscopic features. Recent investigations of emission spectra of phthalocyanine in helium droplets revealed a doubling of all transitions. Herein, we present the emission spectra of Mg-phthalocyanine and of phthalocyanine-argon clusters in helium droplets, which confirm the splitting as a general effect of the helium environment. A scheme of levels is deduced from the emission spectra and attributed to quantized states of the first helium layer surrounding the dopant molecule. PMID- 15298389 TI - Photochromism of novel molybdate/alkylamine composite thin films. AB - Novel inorganic/organic composite films of molybdates with photochromic properties have been prepared by self-assembly using alkylammonium ions as a supramolecular template. Both 1-hexadecylammonium/polyoxomolybdate (C16-Mo) and 1 octadecylammonium/polyoxomolybdate (C18-Mo) composite films have been successfully fabricated. The elemental analysis and thermal gravimetric analysis show that the main product in the C16-Mo film was (C16H33NH3)4Mo8O26. The X-ray diffraction (XRD) results indicate that the composite films were lamellar in nature. The IR, Raman and X-ray photoelectron spectroscopy (XPS) results show that the polyoxomolybdate anions present as MoO6 octahedra and that the Mo species exists as Mo6+ in the freshly prepared films. The alkyl chains in the 1 hexadecylammonium chains were linear and the alkyl groups are an all-trans configuration. Upon UV irradiation of the C16-Mo films, some Mo6+ was reduced to Mo5+, some -NH3+ became -NH2 with a concomitant increase in the concentration of OH groups on the molybdate moieties, and the films were colored. Thus, the photochromism of the films involves the reduction of Mo6+ to Mo5+, coupled with a proton transfer from 1-hexadecylammonium ions to an oxygen atom at the Mo site. In contrast to thin films of transition-metal oxides, which all show photochromism in the blue region of the electromagnetic spectrum, these composite films show photochromism in the violet region with the greatest absorbance change at 472 nm. PMID- 15298390 TI - Morphology and nanomechanics of conducting plastic crystals. AB - We present a temperature-controlled scanning probe microscopy characterisation of morphology and mechanical behaviour of conducting N,N'-cyclized pyrazolium salts. These salts are plastic crystals belonging to the pyrazolium trifluoromethanesulfonimide family. Going from a five-membered to a seven membered ring, and adding a methyl group in alpha to either the five-membered or six-membered rings, allows modulation of the temperature of the phase transitions. Before and after the transitions, the materials' Young moduli, hardnesses and surface roughnesses change. We attribute these macroscopic modifications to specific states: brittle, elastoplastic and viscoplastic, corresponding to variations in the extent of the dislocations present in the crystal lattice planes of the compounds. PMID- 15298391 TI - Evaporation of water microdroplets on self-assembled monolayers: from pinning to shrinking. PMID- 15298392 TI - Generation and spectroscopic characterization of methylnitrene diradicals adsorbed on the Cu(110) surface. PMID- 15298393 TI - System-size biresonance for intracellular calcium signaling. PMID- 15298395 TI - Molecular simulation study of interactions of carbon dioxide and water with ionic liquids. PMID- 15298394 TI - Pseudorotation of natural and chemically modified biological phosphoranes: implications for RNA catalysis. PMID- 15298396 TI - Resolving rotational spectra of hydrogen adsorbed on a single-walled carbon nanotube substrate. PMID- 15298397 TI - The impact of phase changes, alloying and segregation in supported rhpd catalysts during selective NO reduction by H2. PMID- 15298399 TI - Performance of a compact adaptive-optics system. AB - The design of an adaptive-optics system for correction of a beam propagating through high-speed, unpredictable optical turbulence required the use of a robust controller rather than a conventional least-squares controller. We describe the 37-channel, 50-Hz adaptive-optical system and its performance (lambda/75 rms). PMID- 15298398 TI - Photochemistry of charge-transfer complexes in a viologen periodic mesoporous organosilica: time evolution from femtoseconds to minutes. PMID- 15298400 TI - Layer-oriented simulation tool. AB - The Layer-Oriented Simulation Tool (LOST) is a numerical simulation code developed for analysis of the performance of multiconjugate adaptive optics modules following a layer-oriented approach. The LOST code computes the atmospheric layers in terms of phase screens and then propagates the phase delays introduced in the natural guide stars' wave fronts by using geometrical optics approximations. These wave fronts are combined in an optical or numerical way, including the effects of wave-front sensors on measurements in terms of phase noise. The LOST code is described, and two applications to layer-oriented modules are briefly presented. We have focus on the Multiconjugate adaptive optics demonstrator to be mounted upon the Very Large Telescope and on the Near-IR Visible Adaptive Interferometer for Astronomy (NIRVANA) interferometric system to be installed on the combined focus of the Large Binocular Telescope. PMID- 15298401 TI - Artificial apposition compound eye fabricated by micro-optics technology. AB - By exploring micro-optical design principles and technology, we have developed an artificial apposition compound eye. The overall thickness of the imaging system is only 320 microm, the diagonal field of view is 21 degrees, and the f-number is 2.6. The monolithic device consists of an UV-replicated microlens array upon a thin silica substrate with a pinhole array in a metal layer on the back side. The pitch of the pinholes differs from that of the lens array to provide individual viewing angle for each channel. Theoretical limitations of resolution and sensitivity are discussed as well as fabrication issues and compared with experimental results. A method to generate nontransparent walls between optical channels to prevent cross talk is proposed. PMID- 15298402 TI - High-resolution optical angle sensors: approaching the diffraction limit to the sensitivity. AB - We carry out a detailed analysis of angle-sensitive devices based on the critical angle effect. We consider their use in measuring small angular deflections of a laser beam. We establish the diffraction limit to the sensitivity for optical angle sensors based on reflection and transmission of a laser beam. We find that this limit is identical to that of the triangulation scheme when using a position sensitive detector or the autocollimation scheme. We analyze the main proposals to date of optical-angle sensors based on the critical-angle effect, focusing on their maximum sensitivity and their polarization dependence in practical conditions. We propose and analyze theoretically a novel and simple angle sensitive device for sensing optical-beam deflections with very low polarization dependence and a maximum sensitivity close to the diffraction limit when used with typical laser beams. We discuss the basic principles for designing this type of device, provide numerical results, and point out a convenient fabrication procedure. PMID- 15298403 TI - Annular pupils, radial polarization, and superresolution. AB - An annular pupil, which can be used to produce a Bessel beam, when combined with radially polarized illumination promises improvements in microscope resolution, increased packing density for optical storage, and finer optical lithography. When combined with a circular detection pupil in confocal microscopy a point spread function 112 nm wide results (lambda = 488 nm). Radially polarized annular illumination of a solid-immersion lens can yield a focal spot smaller than 100 nm for lambda = 488 nm. Use of radially polarized illumination with pupil masks is discussed. PMID- 15298404 TI - Parabola-doublet aplanat becomes anastigmatic when second doublet is inserted near focus. AB - A doublet of choice glasses may be located in the converging focal cone of the infinity-focused parabola to yield an aplanatic telescope or camera. The resulting angular field is limited by high astigmatism but is significantly larger than that of the coma-limited parabola. The spherical and chromatic aberrations are so well corrected and the coma so well balanced that the doublet may be used unaltered with a parabola of arbitrary focal length and speed with excellent results for the unvignetted rays. A second doublet nearer to the focus and designed independently of the first corrects the system's astigmatism while preserving its aplanaticism. It may also be designed for flattening the field. This arrangement may allow for greater flexibility in the placing of optical elements than does Wynne's triplet for modest-aperture systems. Equations are presented for choosing candidate glasses for the first doublet from the very limited manifold of solving glasses. PMID- 15298405 TI - Effects of different beacon wavelengths on atmospheric compensation in strong scintillation. AB - During strong scintillation, the number and location of branch points in a distorted optical field induced by atmospheric turbulence are closely related to the characteristic parameters of the turbulence effect, propagation distance, and wavelength. It is necessary to consider the effect of the beacon's wavelength on the adaptive optics system that is used to compensate for atmospheric turbulence. Our analytical results show that the performance of adaptive optics can be improved by nearly a factor of 2 when the beacon's wavelength is chosen slightly longer than the wavelength of the main laser in the branch points considered. PMID- 15298406 TI - Temporal filtering by double diffraction. AB - We present a theoretical analysis of the temporal behavior of double-diffraction setups. It applies, in particular, to Talbot and Montgomery interferometers, whose operation is based on the self-imaging effect. The use of both types of interferometer as temporal filters for optical and terahertz applications was recently suggested. We show that double-diffraction setups can be modeled as communications channels with dispersive behavior caused by diffraction. We develop mathematical expressions for the phase delay, the group velocity, and the group-velocity dispersion for both quasi-monochromatic and polychromatic case. Based on these results, the temporal impulse response of a double-diffraction setup is derived. Finally, a general description of its practical implementation are presented. PMID- 15298407 TI - Restoration of images captured by a staggered time delay and integration camera in the presence of mechanical vibrations. AB - Staggered time delay and integration (TDI) scanning image acquisition systems are usually employed in low signal-to-noise situations such as thermal imaging. Analysis and restoration of images acquired by thermal staggered TDI sensors in the presence of mechanical vibrations that may cause space-variant image distortions (severe geometric warps and blur) are studied. The relative motion at each location in the degraded image is identified from the image when a differential technique is used. This information is then used to reconstruct the image by a technique of projection onto convex sets. The main novelty is the implementation of such methods to scanned images (columnwise). Restorations are performed with simulated and real mechanically degraded thermal images. PMID- 15298408 TI - Phase demodulation from a single fringe pattern based on a correlation technique. AB - We present a method for determining the demodulated phase from a single fringe pattern. This method, based on a correlation technique, searches in a zone of interest for the degree of similarity between a real fringe pattern and a mathematical model. This method, named modulated phase correlation, is tested with different examples. PMID- 15298409 TI - Interferometric ellipsometer with wavelength-modulated laser diode source. AB - An interferometric ellipsometer, with no moving parts and an inexpensive laser diode source, is demonstrated. Temporal fringes are produced by a small modulation of the laser diode bias current and unbalanced arms in the interferometer. Fringe analysis algorithms are developed, and accurate measurements of the optical properties of a number of samples are made. Temperature tuning the laser diode center wavelength allows the frequency dependence of the optical properties to be determined over a wavelength range of approximately 1 nm. PMID- 15298410 TI - Amorphization induced by subnanosecond laser pulses in phase-change optical recording media. AB - We have investigated the dynamics of amorphization induced in phase-change optical recording media by focused laser pulses of subnanosecond duration. We initiated localized amorphism by using a focused laser beam to melt the phase change material and completed the change by rapid cooling by means of thermal diffusion. These studies were conducted by use of real-time reflectivity measurements with a pump-and-probe technique in which both pump and probe pulses had a duration of approximately 510 ps. Our transient-reflectivity measurements indicate that the process that leads to amorphism has three distinct stages, namely, rapid melting, solidification, and slow relaxation. PMID- 15298411 TI - Holographic creation of photonic crystals. AB - We describe the creation of general photonic crystals by means of holography with an experimental demonstration. The recordings of periodic variations of amplitude and phase by the interference of coherent laser beams offer a natural means for the creation of one- two- or three-dimensional photonic crystals. Based on the principle of the interference of four noncoplanar beams, we present a comparative analysis of two different approaches for creating photonic crystals and use numerical simulated lattice structures to illustrate the differences between these two approaches. We then use a specific symmetrical optical architecture and select the proper approach to create holographic photonic crystals. The advantages and constraints of this holographic method are discussed. PMID- 15298412 TI - Real-time phase tracking in single-photon interferometers. AB - A new technique for phase tracking in quantum cryptography systems is proposed that adjusts phase in an optimal way, using only as many photon counts as necessary. We derive an upper bound on the number of photons that need to be registered during phase adjustment to achieve a given phase accuracy. It turns out that most quantum cryptosystems can successfully track phase on a single photon level, entirely with software, without any additional hardware components or extensive phase-stabilization measures. The technique is tested experimentally on a quantum cryptosystem. PMID- 15298413 TI - Polarization mode coupling in circularly birefringent gratings. AB - Polarization mode coupling in circularly birefringent gratings is analyzed. It is numerically found that efficient LP01x-LP02y mode coupling (where LP is linear polarization) is possible in a 50-cm-long circularly birefringent fiber grating formed in a terbium-doped borosilicate glass fiber and that complete LP01x-LP02y and LP01x-LP03y mode couplings result after a few-centimeter-long circularly birefringent grating that is formed in a bismuth-substitute iron garnet waveguide. Various parameters of polarization mode coupling in a number of circularly birefringent gratings are also computed. PMID- 15298414 TI - Laser phase noise reduction for industrial interferometric applications. AB - Laser ultrasound is a technique used for the ultrasonic inspection of composites during manufacturing of advanced jet fighters. With this technique laser interferometry is used to detect ultrasonic displacements generated by a laser. In theory, the signal-to-noise ratio is proportional to the square root of the collected detection light. In practice, laser phase noise limits the signal-to noise ratio above a certain collected light level. Two techniques are presented to decrease effects due to laser noise. In one technique the dual-cavity Fabry Perot currently used is replaced by an interferometer based on a photorefractive crystal. The other technique has a high-finesse Sagnac cavity that filters the phase noise from the detection laser. Experimental results demonstrate that these two techniques significantly reduce limitations due to laser noise. PMID- 15298415 TI - Local one-dimensional approximation for fast simulation of Z-scan measurements with an arbitrary beam. AB - We apply a finite-difference algorithm that combines the local one-dimensional approximation and the Crank-Nicolson algorithms to solve the three-dimensional nonlinear Schrodinger equation. This scheme is unconditionally stable and accurate to second order. Therefore it offers a simple and accurate means to study a two-dimensional Z scan for arbitrary beam shape and medium length. As an example, we analyze the characteristics of a Z scan by utilizing an elliptic Gaussian beam for a thick nonlinear medium. The effects of ellipticity and waist separation of the elliptic beam on the normalized transmittance of the closed aperture and open-aperture Z scan are demonstrated. PMID- 15298416 TI - Retrieval of profile information from airborne multiaxis UV-visible skylight absorption measurements. AB - A recent development in ground-based remote sensing of atmospheric constituents by UV-visible absorption measurements of scattered light is the simultaneous use of several horizon viewing directions in addition to the traditional zenith-sky pointing. The different light paths through the atmosphere enable the vertical distribution of some atmospheric absorbers, such as NO2, BrO, or O3, to be retrieved. This approach has recently been implemented on an airborne platform. This novel instrument, the airborne multiaxis differential optical absorption spectrometer (AMAXDOAS), has been flown for the first time. In this study, the amount of profile information that can be retrieved from such measurements is investigated for the trace gas NO2. Sensitivity studies on synthetic data are performed for a variety of representative measurement conditions including two wavelengths, one in the UV and one in the visible, two different surface spectral reflectances, various lines of sight (LOSs), and for two different flight altitudes. The results demonstrate that the AMAXDOAS measurements contain useful profile information, mainly at flight altitude and below the aircraft. Depending on wavelength and LOS used, the vertical resolution of the retrieved profiles is as good as 2 km near flight altitude. Above 14 km the profile information content of AMAXDOAS measurements is sparse. Airborne multiaxis measurements are thus a promising tool for atmospheric studies in the troposphere and the upper troposphere and lower stratosphere region. PMID- 15298417 TI - Effect of particle asphericity on single-scattering parameters: comparison between Platonic solids and spheres. AB - The single-scattering properties of the Platonic shapes, namely, the tetrahedron, hexahedron, octahedron, dodecahedron, and icosahedron, are investigated by use of the finite-difference time-domain method. These Platonic shapes have different extents of asphericity in terms of the ratios of their volumes (or surface areas) to those of their circumscribed spheres. We present the errors associated with four types of spherical equivalence that are defined on the basis of (a) the particle's geometric dimension (b) equal surface area (A), (c) equal volume (V), and (d) equal-volume-to-surface-area ratio (V/A). Numerical results show that the derivations of the scattering properties of a nonspherical particle from its spherical counterpart depend on the definition of spherical equivalence. For instance, when the Platonic and spherical particles have the same geometric dimension, the phase function for a dodecahedron is more similar than that for an icosahedron to the spherical result even though an icosahedron has more faces than a dodecahedron. However, when the nonspherical and spherical particles have the same volume, the phase function of the icosahedral particle essentially converges to the phase function of the sphere, whereas the result for the dodecahedron is quite different from its spherical counterpart. Furthermore, the present scattering calculation shows that the approximation of a Platonic solid with a sphere based on V/A leads to larger errors than the spherical equivalence based on either volume or projected area. PMID- 15298418 TI - Mars laser hygrometer. AB - We have designed and built a miniature near-IR tunable diode laser (TDL) spectrometer for measuring in situ the water vapor mixing ratio either in the Martian atmosphere or thermally evolved from Martian soil or ice samples. The laser hygrometer uses a thermoelectrically cooled single-mode distributed feedback TDL at 1.87 microm to scan over a selected vibration-rotation line of both H2O and CO2 near 5327.3 cm(-1). A working prototype that weighs only 230 g has been built and used to generate spectra whose analysis demonstrates precision sensitivities as fine as 1 part in 10(6) by volume in 1 s or 0.1 part in 10(6) in 10 s at Martian pressures and temperatures. Absolute uncertainties of approximately 5% are calculated. PMID- 15298419 TI - Experimental method based on wavelength-modulation spectroscopy for the characterization of semiconductor lasers under direct modulation. AB - An experimental method is presented for characterization of the combined intensity and frequency modulation produced when the injection current of a laser diode is modulated. The reported technique is based on the analysis of the harmonic signals produced when a modulated laser is used to probe a gas absorption line by the so-called wavelength-modulation spectroscopy method. Based on a theoretical model of this technique, we present two methods that facilitate the determination of (i) the deviation in laser frequency and (ii) the phase shift between intensity and frequency modulation. These methods are illustrated experimentally by measurement of the modulation parameters of a 2-microm distributed-feedback laser by use of a CO2 absorption line. The experimental results have been compared with those obtained with another traditional method and have shown full agreement in the frequency range (400 Hz-30 kHz) considered. PMID- 15298420 TI - The oxygen dissociation curve: quantifying the shift. AB - An oxyhaemoglobin dissociation curve (ODC) quantifies the most important function of red blood cells and that is the affinity for oxygen and its delivery to the tissues. Oxygen affinity for haemoglobin plays a critical role in the delivery of oxygen to the tissues and is changed by shifting to the left or right. A shift to the left implies an increased oxygen affinity and, hence, tighter binding due to the higher oxygen saturation in relation to the pO2. On the other hand, a shift to the right corresponds to a decreased oxygen affinity and easier release of oxygen to the tissues. It is well known that the ODC shifts in response to changes in pH, pCO2 and 2,3 diphosphoglycerate. However, how much the ODC shifts has never been quantified. Arterial and venous blood gases were taken during cardiopulmonary bypass and two indices were used to quantify the shift of the ODC; the p50 shift and the SO2 difference. Arterial blood shifted to the right by 4 +/- 0.1 mmHg at a pH of 7.24 and shifted to the left by -3.5 +/- 0.05 mmHg at a pH of 7.51. The change in arterial saturation was minimal, rising by 0.8% and dropping by -5% and did not correlate to p50 shifting and changes in pH, but demonstrated changes dependent on the concentration of dyshaemoglobins. The venous blood exhibited a greater range of p50 shifting at each pH value. At a pH of 7.24, the p50 shifted to the right by 4.8 +/- 2 mmHg and at a pH of 7.51 the p50 shifted to the left by -4 +/- 1.8 mmHg. Unlike the arterial blood, the change in saturation correlated well to p50 shifting. It is shown here for the first time how much the curve shifts with changes in pH and how this may be used to evaluate treatment strategies. PMID- 15298421 TI - Metabolic acidosis developing during cardiopulmonary bypass is related to a decrease in strong ion difference. AB - Metabolic acidosis is a frequent complication of cardiopulmonary bypass (CPB). Commonly, its cause is ascribed to hypoperfusion; however, iatrogenic causes, related to the composition and volume of intravascular fluids that are administered, are increasingly being recognized. The aim of this study was to determine if metabolic acidosis during CPB was associated with hypoperfusion, change in strong ion difference (SID) or haemodilution. Forty-nine patients undergoing cardiac surgery using CPB in the Royal Infirmary of Edinburgh (RIE) or the HCI, Clydebank were included in the study. Arterial blood samples were aspirated before induction of anaesthesia and the end of CPB. Samples were subjected to blood gas analysis and measurement of electrolytes and lactate. Changes in concentrations were then calculated. Change variables that were found to be significant (p < 0.1) univariate correlates of the change in hydrogen ion concentration were identified and entered into a multivariate regression model with hydrogen ion concentration at the end of CPB as the outcome variable (r2 = 0.65, p < 0.001). Change variance in hydrogen ion concentration was created by first entering the baseline hydrogen ion concentration into the model. Next, any variance resulting from the respiratory component of acidosis was removed by entering the change in arterial carbon dioxide tension (regression coefficient (beta)=0.67, p < 0.01). Change in SID (beta = -0.34, p < 0.01) and surgical institution (beta = 0.40, p < 0.01) were then found to be predictors of the remaining variance whilst change in concentration of lactate (beta in = 0.16, p = 0.07) and volume of intravascular fluid that was administered (beta = -0.07, p = 0.52) were rejected from the model. These findings suggest that the metabolic acidosis developing during CPB is partially the result of iatrogenic decrease in SID rather than hypoperfusion, as estimated by lactate concentration, or haemodilution. PMID- 15298422 TI - Liver blood flow during cardiac surgery. AB - OBJECTIVE: Impairment of liver blood flow and, therefore, potentially liver function, has important short-term consequences because of the liver's key metabolic importance and role in drug metabolism. The objective of this study was to quantify the effect of cardiac surgery on liver blood flow from before the induction of anaesthesia to 24 hours postoperatively. METHOD: Ten patients with no history of liver impairment, moderate or good left ventricular function, and undergoing routine hypothermic coronary artery bypass graft surgery, were entered into the study. Liver blood flow was determined by the clearance of indocyanine green (ICG), expressed as a percentage disappearance rate (PDR). RESULTS: The mean baseline percentage disappearence rate (PDR) of indocyanine green (ICG) was 19.84 +/- 4.47%/min. This increased marginally to 20.42 +/- 6.67%/min following the induction of anaesthesia, but after 15 min of cardiopulmonary bypass, the PDR fell to 13.51 +/- 3.69%/min; this was significantly lower than all other PDRs measured throughout the study. Prior to extubation, the PDR increased again to 20.01 +/- 3.72%/ min, and this level was maintained at 12 hours (PDR 20.32 +/- 3.53%min) and 24 hours (PDR 20.51 +/- 2.27%/min). CONCLUSION: The induction of anaesthesia and positive pressure ventilation do not affect liver blood flow. Cardiopulmonary bypass at 30 degrees C is associated with a significant reduction in liver blood flow, which returns to normal within 4-6 hours of surgery and remains normal for up to 24 hours after surgery. PMID- 15298423 TI - Atrial natriuretic factor in normothermic and hypothermic cardiopulmonary bypass. AB - BACKGROUND: To evaluate the plasmatic changes of atrial natriuretic factor (ANF) during and after cardiopulmonary bypass (CPB) in normothermia and hypothermia. METHODS: Twenty-three patients (n = 23) undergoing coronary artery bypass graft surgery were randomly assigned to two groups. In Group I (n = 11), the patients underwent operation in normothermia; in Group II (n = 12), the operation was performed in hypothermia (26 degrees C). RESULTS: Plasma ANF levels were determined after induction of anaesthesia, at the end of CPB and one hour postoperatively. There were no demographic differences between the two groups, diuresis (p = 0.90) and natriuresis (p = 0.95). Plasma levels of ANF were significantly elevated during and after CPB in both groups (p < 0.01). The groups differed significantly for plasma levels of ANF during CPB and postoperatively (p < 0.05), but did not differ prebypass (p = 0.08). There was no correlation in either group between ANF release and central venous pressure, natriuresis and diuresis. There was only a borderline relationship between ANF concentration and diuresis after CPB in Group I. CONCLUSION: CPB triggers the production and release of ANF. The present study demonstrates a significantly enhanced ANF release during hypothermia and reperfusion after ischaemia. Thus, these data suggest the protective role of ANF on the hypoxic myocardium, and they confirm that ANF does not play a role in diuresis and natriuresis during and after hypothermic CPB. PMID- 15298424 TI - Comparison of the effect of venovenous versus venoarterial extracorporeal membrane oxygenation on renal blood flow in newborn lambs. AB - Venovenous extracorporeal membrane oxygenation (VV ECMO) using double lumen catheters is an alternative to venoarterial (VA) ECMO and allows for total blood flow using the patient's cardiac output in comparison to partial blood flow provided during VA ECMO. OBJECTIVE: To compare the effects of VV versus VA ECMO on renal blood flow. DESIGN: Prospective study. SETTING: Research laboratory in a hospital. SUBJECT: Newborn lambs 1-7 days of age (n = 15). INTERVENTIONS: In anesthetized, ventilated lambs, femoral artery and vein were cannulated for monitoring and renal venous blood sampling. An ultrasonic flow probe was placed on the left renal artery for continuous renal blood flow measurements. Animals were randomly assigned to control (non-ECMO), VV ECMO and VA ECMO groups. After systemic heparinization, the animals were cannulated and studied at bypass flows of 120 mL/kg/min (partial bypass) for two hours in both ECMO groups and 200 mL/kg/min (full bypass) for an additional 30 min in the VA group. Changes in blood pressure and renal flow on ECMO and during ECMO bridge unclamping were recorded continuously. Plasma renin activity (PRA) levels were sequentially sampled. RESULTS: Systemic blood pressure was not different in VV or VA ECMO at partial bypass flow. However, systemic blood pressure increased significantly at maximal bypass flow in the VA ECMO group. There was no change in renal flow in either VV or VA ECMO groups. PRA levels did not correlate with bypass flow change. During unclamping of the ECMO bridge, blood pressure and renal flow drop significantly in the VA group, but not in the VV group. CONCLUSION: VV and VA ECMO at partial bypass flows had comparable effect on blood pressure, renal blood flow and PRA level in this short-term study. However, unclamping of the ECMO bridges did differentially affect blood pressure and renal blood flow between VV and VA groups. We speculate that this repeated acute change in long-run VA ECMO support may play a role in the persistent hypertension seen in some patients. PMID- 15298425 TI - Cerebral saturations trend with mixed venous saturations in patients undergoing extracorporeal life support. AB - Cerebral saturation (SCO2) monitors are noninvasive tools that continuously measure saturations in the cerebral cortex, a predominately venous bed. The purpose of this study was to see if a trend existed between measurements of SCO2 and mixed venous saturation values (SVO2) for patients on extracorporeal life support (ECLS). Six patients required ECLS for cardiac failure after congenital cardiac surgery, and one patient required ECLS for pulmonary failure. Patients were divided into two groups, those without systemic/pulmonary venous mixing (n = 3, Group I) and those with mixing due to an intraatrial shunt or left ventricular vent (n = 4, Group II). The age of patients was 0.4 +/- 0.5 years (mean +/- SD), weight was 5.2 +/- 2.3 kg, and time on ECLS was 8.3 +/- 4.8 days. No significant abnormalities were seen on head imaging. A total of 786 paired data points were collected. Mean values were different; however, there was a significant trend between SCO2 and SVO2 for the entire sample (R2 = 0.66, p < 0.001). Cerebral saturation trends follow mixed venous trends and, therefore, may be helpful in combination with other physical and laboratory findings in the care of the critically ill child. PMID- 15298426 TI - Comparison of a Duraflo II-coated cardiopulmonary bypass circuit and a trillium coated oxygenator during open-heart surgery. AB - BACKGROUND: Cardiopulmonary bypass (CPB) evokes a systemic inflammatory response. In attempting to improve the biocompatibility of the equipment, various methods to coat the inner surfaces of the CPB systems have been developed. The present study compares a Trillium Biopassive surface-coated Affinity oxygenator with a Duraflo II totally heparin-coated CPB system. METHODS: Low-risk patients admitted for primary coronary artery bypass grafting or aortic valve replacement were randomized to operation using the Trillium- or the Duraflo II-coated setups. Heparin concentration, complement activation (C3bc activation products and terminal complement complex (TCC)), platelet activation (platelet numbers and beta-thromboglobulin (BTG)), leukocyte activation (leukocyte numbers and myeloperoxidase (MPO)), coagulation (thrombin/antithrombin complexes (TAT)) and fibrinolytic activity (plasmin/alpha2-antiplasmin complexes (PAP)) were measured during CPB and two hours postoperatively. RESULTS: Platelet counts decreased during CPB, without significant intergroup differences. The median BTG concentration increased moderately in both groups and were slightly higher in the Trillium group during CPB (p < 0.05), but not postoperatively. Complement activation products (C3bc and TCC), leukocyte counts, MPO, TAT and PAP activity showed no differences between the two groups. CONCLUSIONS: There were small differences in the inflammatory response between the two extracorporeal circulation devices compared in this study. PMID- 15298427 TI - Methylprednisolone prevents inflammatory reaction occurring during cardiopulmonary bypass: effects on TNF-alpha, IL-6, IL-8, IL-10. AB - OBJECTIVE: This study examined the correlation between tumour necrosis factor alpha (TNF-alpha), interleukin (IL)-6 and IL-8, IL-10 and methylprednisolone pretreatment. METHODS: This is a prospective, randomized and double-blinded study. Sixty patients undergoing coronary artery bypass grafting (CABG) were randomized to receive either intravenous methylprednisolone (n = 30, Group M) or intravenous placebo (n = 30, Group S). The patients received intravenously either 30 mg/kg methylprednisolone (Group M) or placebo (Group S) 10 min before and after cardiopulmonary bypass (CPB). In an intensive care unit (ICU), four additional doses were given at 6-hourly intervals. Blood samples for the measurements of TNF-alpha, IL-6, IL-8 and IL-10 were obtained before induction of anaesthesia (T0 = control value), after induction (T1), before starting CPB (T2), after aortic declamping (T3), at the end of CPB (T4) and 6 hours (T5), 12 hours (T6) and 24 hours (T7) after skin closure. Creatine kinase (CK) and creatine kinase isoenzyme MB (CK-MB) were evaluated at the following intervals: T0, T5, T6 and T7. RESULTS: When compared with the control value, TNF-alpha, IL-6 and IL-8 significantly increased in Group S and Group M (p < 0.05), but these values were significantly greater in Group S than in Group M (p < 0.05). In comparison with the control value, IL-10 increased in both groups (p < 0.05), but was significantly greater in Group M than in Group S (p < 0.05). CK and CK-MB were increased in both groups in postoperative values compared to control values. In Group S, CK and CK-MB levels were significantly lower than in Group M (p < 0.05). CONCLUSION: In this study, we have found that preoperative administration of methylprednisolone has decreased TNF-alpha, IL-6 and IL-8 release, and increased the perfusing IL-10 levels after CPB. Thus, methylprednisolone may decrease the inflammatory response during the CPB procedure. PMID- 15298428 TI - Improved hydrodynamics of a new aortic cannula with a novel tip design. AB - BACKGROUND: Reduction of atheroembolic complications during cardiopulmonary bypass remains a major challenge in cardiac surgery. New cannula tip designs may help to attenuate this problem by improved hydrodynamics. METHODS: Pressure gradients and back pressures of a new aortic cannula tip design were measured and compared with the Medos X-Flow, Sarns Soft-Flow and Argyle THI cannulae at various flow rates in a mock circulation followed by flow visualization. RESULTS: Pressure gradients were the lowest for the new cannula. Back pressures of the new cannula were up to 84% lower than for the Argyle cannula. The back pressure profile and flow visualization of the new cannula showed broad centric flow dispersion with a transcannula increase of flow area from 38 mm2 to 139 mm2. CONCLUSIONS: The new design of an aortic cannula tip provides improved hydrodynamics, with low pressure gradients, low back pressures and a uniform central dispersion of flow, reducing the sandblasting effect. PMID- 15298429 TI - Adult stem cell driven genesis of human-shaped articular condyle. AB - Uniform design of synovial articulations across mammalian species is challenged by their common susceptibility to joint degeneration. The present study was designed to investigate the possibility of creating human-shaped articular condyles by rat bone marrow-derived mesenchymal stem cells (MSCs) encapsulated in a biocompatible poly(ethylene glycol)-based hydrogel. Rat MSCs were harvested, expanded in culture, and treated with either chondrogenic or osteogenic supplements. Rat MSC-derived chondrogenic and osteogenic cells were loaded in hydrogel suspensions in two stratified and yet integrated hydrogel layers that were sequentially photopolymerized in a human condylar mold. Harvested articular condyles from 4-week in vivo implantation demonstrated stratified layers of chondrogenesis and osteogenesis. Parallel in vitro experiments using goat and rat MSCs corroborated in vivo data by demonstrating the expression of chondrogenic and osteogenic markers by biochemical and mRNA analyses. Ex vivo incubated goat MSC-derived chondral constructs contained cartilage-related glycosaminoglycans and collagen. By contrast, goat MSC-derived osteogenic constructs expressed alkaline phosphatase and osteonectin genes, and showed escalating calcium content over time. Rat MSC-derived osteogenic constructs were stiffer than rat MSC derived chondrogenic constructs upon nanoindentation with atomic force microscopy. These findings may serve as a primitive proof of concept for ultimate tissue-engineered replacement of degenerated articular condyles via a single population of adult mesenchymal stem cells. PMID- 15298430 TI - Investigating tendon fascicle structure-function relationships in a transgenic age mouse model using multiple regression models. AB - Proper replacement or repair of damaged tendons or ligaments requires functionally engineered tissue that mimics their native mechanical properties. While tendon structure-function relationships are generally assumed, there exists little quantitative evidence of the roles of distinct tendon components in tendon function. Previous work has used linear correlations to assess the independent, univariate effects of one structural or one biochemical variable on mechanics. The current study's objective was to simultaneously and rigorously evaluate the relative contributions of seven different structural and compositional variables in predicting tissue mechanical properties through the use of multiple regression statistical models. Structural, biochemical, and mechanical analysis were all performed on tail tendon fascicles from different groups of transgenic mice, which provide a reproducible, noninvasive, in vivo model of changes in tendon structure and composition. Interestingly, glycosaminoglycan (GAG) content was observed to be the strongest predictor of mechanical properties. GAG content was also well correlated with collagen content and mean collagen fibril diameter. Collagen fibril area fraction was a significant predictor only of material properties. Therefore, in a large multivariate model, GAG content was the largest predictor of mechanical properties, perhaps both through direct influence and indirectly through its correlation with collagen content and fibril structure. PMID- 15298431 TI - Characterization of the atherosclerotic carotid bifurcation using MRI, finite element modeling, and histology. AB - Atherogenesis is known to be associated with the stresses that act on or within the arterial wall. Still, the uneven distribution of atherosclerotic lesions and the impact of vessel remodeling on disease progression are poorly understood. A methodology is proposed to study the correlations between fluid dynamic parameters and histological markers of atherosclerosis. Trends suggested by preliminary data from four patients with advanced carotid bifurcation arterial disease are examined and compared to hypotheses in the literature. Four patients were scanned using MRI and ultrasound, and subsequently underwent carotid endarterectomy. For each patient. a geometric model and a numerical mesh were constructed from MR data, and velocity boundary conditions established. Computations yield values for average wall shear stress (WSS), maximum wall shear stress temporal gradient (WSSTG), and Oscillatory Shear Index (OSI). Following surgery, the excised plaques were sectioned, stained for smooth muscle cells (SMC), macrophages (M phi), lipid (LIP), and collagen (COL), and analyzed quantitatively. Correlations attempted between the various fluid dynamic variables and the biological markers were interesting but inconclusive. Tendencies of WSSTG and WSS to correlate negatively with M phi and LIP, and positively with COL and SMC, as well as tendencies of OSI to correlate positively with Mphi and LIP and negatively with COL and SMC, were observed. These trends agree with hypotheses in the literature, which are based on ex vivo and in vitro experimental studies. PMID- 15298432 TI - 3D MRI-based multicomponent FSI models for atherosclerotic plaques. AB - A three-dimensional (3D) MRI-based computational model with multicomponent plaque structure and fluid-structure interactions (FSI) is introduced to perform mechanical analysis for human atherosclerotic plaques and identify critical flow and stress/strain conditions which may be related to plaque rupture. Three dimensional geometry of a human carotid plaque was reconstructed from 3D MR images and computational mesh was generated using Visualization Toolkit. Both the artery wall and the plaque components were assumed to be hyperelastic, isotropic, incompressible, and homogeneous. The flow was assumed to be laminar, Newtonian, viscous, and incompressible. The fully coupled fluid and structure models were solved by ADINA, a well-tested finite element package. Results from two dimensional (2D) and 3D models, based on ex vivo MRI and histological images (HI), with different component sizes and plaque cap thickness, under different pressure and axial stretch conditions, were obtained and compared. Our results indicate that large lipid pools and thin plaque caps are associated with both extreme maximum (stretch) and minimum (compression when negative) stress/strain levels. Large cyclic stress/strain variations in the plaque under pulsating pressure were observed which may lead to artery fatigue and possible plaque rupture. Large-scale patient studies are needed to validate the computational findings for possible plaque vulnerability assessment and rupture predictions. PMID- 15298433 TI - Shear stress-induced binding of large and unusually large von Willebrand factor to human platelet glycoprotein Ibalpha. AB - Platelet membrane glycoprotein (GP) Ib(alpha), a component of the GP Ib-IX-V complex, is a receptor for von Willebrand factor (VWF). A small quantity of large VWF multimers binds to platelets under high shear stress, and induces aggregation. We studied the shear-induced attachment of large and unusually large VWF multimers to the GPIb(alpha) extracellular domain (glycocalicin), human platelets, and GPIb(alpha) gxpressing Chinese hamster ovary (CHO) cells. Compared to binding in the presence of botrocetin and ristocetin, shear stress only induced low-level NVWF (normal plasma VWF multimers) binding. This shear stress induced interaction is also dependent on VWF multimeric size. Elevated binding levels of endothelial cell VWF (enriched in unusually large VWF multimers) to glycocalicin-coated beads were observed under low shear conditions, which did not result in the attachment of normal plasma VWF. PMID- 15298434 TI - Platelet receptor glycoprotein VI-mediated adhesion to type I collagen under hydrodynamic flow. AB - Glycoprotein VI (GPVI) is a platelet receptor that directly binds collagen. It has been shown by expressing GPVI in rat basophilic leukemia (RBL-2H3) cells that GPVI mediates adhesion to type I collagen under static conditions. However, the ability of GPVI to secure adhesion to collagen type I under flow has not been measured. We studied the interaction of GPVI and type I collagen under hydrodynamic flow using RBL-2H3 cells transfected with the GPVI receptor. We found that GPVI-expressing RBL-2H3 cells adhere to collagen under flow, significantly more so than non-GPVI-expressing RBL-2H3 cells. Inhibition of GPVI by the 11A12 anti-GPVI antibody significantly blocks adhesion to collagen, indicating that GPVI specifically interacts with collagen. Probing the role of signaling in GPVI binding to collagen, we used mutants of GPVI and observed that signal transduction did not inhibit adhesion. To test the correlation between receptor expression and adhesion, we tested three GPVI-expressing RBL-2H3 cell lines (A, B, and C) with different levels of receptor expression. At a single shear rate, the level of adhesion increases monotonically with surface expression. The results, using this model cell line, indicate that GPVI is capable of mediating adhesion to collagen under shear, in a density-dependent fashion that is independent of GPVI signaling. PMID- 15298435 TI - Relationship between collagen fibrils, glycosaminoglycans, and stress relaxation in mitral valve chordae tendineae. AB - The tensile properties of mitral valve chordae tendineae derive from their structural make-up. The objectives of this study were to compare the stress relaxation properties of different types of chordae and relate their variation to structural features. Fifty chordae from eight hearts were subjected to stress relaxation tests. The percent stress relaxation and the relaxation rates were found to increase in the order of marginal. basal, and strut chordae. The water content of the three types of chordae was the same (marginal 77.1+/-5.9%, basal 77.0+/-3.4%, strut 78.0+/-2.3% wet weight). The collagen, elastin, and glycosaminoglycan (GAG) content in chordae were quantified using hydroxyproline assay, fastin elastin assay, and fluorophore-assisted carbohydrate electrophoresis, respectively. Collagen content of marginal chordae was only slightly less than that of basal and strut chordae (marginal 56.6+/-8.2%, basal 61.4+/-5.6%, strut 63.8+/-3.9% dry weight). There was also no significant difference in elastin content between the chordae (marginal 5.3+/-3.2%, basal 5.4+/-2.7%, strut 4.6+/-1.7% dry weight). However, the concentrations of unsulfated chondroitin/dermatan sulfate, 6-sulfated chondroitin sulfate, and 4 sulfate chondroitin sulfate significantly decreased in the order of marginal, basal, and strut. The total GAG-content also decreased in the order of marginal, basal, and strut (p = 0.06). The greater amount of GAGs in marginal versus strut chordae is consistent with our previous observations that marginal chordae have a greater collagen fibril density and thus more GAG-mediated, fibril-to-fibril linkages. The greater number of proteoglycan linkages may prevent the slippage of fibrils with respect to each other, and thus reduce stress relaxation. The different viscoelastic properties of mitral valve chordae can thus be explained morphologically. PMID- 15298436 TI - Activation dynamics in anisotropic cardiac tissue via decoupling. AB - Bidomain theory for cardiac tissue assumes two interpenetrating anisotropic media -intracellular (i) and extracellular (e)--connected everywhere via a cell membrane; four local parameters sigma(i,e)(l,t) specify conductivities in the longitudinal (l) and transverse (t) directions with respect to cardiac muscle fibers. The full bidomain model for the propagation of electrical activation consists of coupled elliptic-parabolic partial differential equations for the transmembrane potential upsilon(m) and extracellular potential phi(e), together with quasistatic equations for the flow of current in the extracardiac regions. In this work we develop a preliminary assessment of the consequences of neglecting the effect of the passive extracardiac tissue and intracardiac blood masses on wave propagation in isolated whole heart models and describe a decoupling procedure, which requires no assumptions on the anisotropic conductivities and which yields a single reaction-diffusion equation for simulating the propagation of activation. This reduction to a decoupled model is justified in terms of the dimensionless parameter epsilon = (sigma(i)(l)sigma(e)(t) - sigma(i)(t)sigma(e)(l))/(sigma(i)(l) + sigma(e)(l))(sigma(i)(t) + sigma(e)(t)). Numerical simulations are generated which compare propagation in a sheet H of cardiac tissue using the full bidomain model, an isolated bidomain model, and the decoupled model. Preliminary results suggest that the decoupled model may be adequate for studying general properties of cardiac dynamics in isolated whole heart models. PMID- 15298437 TI - Effects of an artery/vascular graft compliance mismatch on protein transport: a numerical study. AB - Small-diameter vascular graft failure by intimal hyperplasia and thrombosis may result from flow disturbances and disruption of chemical transport in the fluid at the distal anastomosis, because of compliance mismatch between the graft and host artery. In previous studies. lower-than-normal wall shear stress (WSS), particle trapping, and high particle residence times were observed at the distal anastomosis due to a pulsatile tubular expansion effect caused by nonuniform radial deformations. This study was undertaken to examine effects of compliance and radius mismatch on the distribution of a model protein released at the graft fluid interface. Finite element simulations of end-to-end vascular grafting were performed under pulsatile flow, using fluid-structure coupling to give physiologic wall displacements. Results showed that protein is convected smoothly downstream in a uniform compliant tube. A compliance mismatch disturbed the transport, causing positive and negative gradients in the concentration profile at the distal anastomosis. This was seen when the graft and artery radii were matched at zero pressure and at mean arterial pressure; low WSSs were only observed in the former case. Thus the distal intimal hypertrophy seen in noncompliant grafts may be caused partly by decreased WSS, and partly by concentration gradients of dissolved chemicals affecting chemotaxis of cells. PMID- 15298438 TI - MUAP number estimates in surface EMG: template-matching methods and their performance boundaries. AB - Estimates of the number of motor unit action potential (MUAP)s appearing in the surface electromyogram (EMG) signal, which offers potentially valuable information about motor unit recruitment and firing rates, are likely to provide a more accurate reflection of the neural command to muscle than are current EMG quantification methods. In this paper, we show that the basic shapes of surface MUAPs recorded from the first dorsal interosseous (FDI) muscle can ideally be represented by a small number of waveforms. On the basis of this, we seek to estimate the number of MUAPs present in standard surface EMG records, using template-matching techniques to identify MUAP occurrences. Our simulation study indicates that the performance of template-matching methods for MUAP number estimation is mainly constrained by the MUAP superposition in the signal, and the maximum number of MUAPs allowed in the signal for a good estimation is determined by the duration of MUAPs. To further explore this from experimental surface EMG signals, we compare the recordings from a selective multiple concentric ring electrode against those derived from a standard differential EMG electrode situated over the same muscle. We conclude that the ring surface electrode only slightly reduces the MUAP duration and the less MUAP superposition rate contained in the signal is mainly achieved by reducing the pick up area of the electrode. Using a template-matching method, although the number of MUAPs can be approximately estimated based on a very selective surface EMG recording at low force levels, the maximum number of MUAPs correctly estimated from the surface EMG is constrained by the MUAP duration. PMID- 15298439 TI - Dynamic measurements of transverse optical trapping force in biological applications. AB - Optical tweezers present a technology for measurements of biological forces in the piconewton range. In such applications, one method of calibrating the transverse optical trapping force involves relating a known external force to the displacement of the trapped object from the trapping center. In this work we used Fourier analysis of the equation of motion to calculate the displacement of the trapped object from the trapping center under an external force induced by viscous drag. Triangular waveforms of different frequencies were used both in theoretical modeling and experiments to induce a force on a trapped object. We investigated the contribution of various factors including frequency of the external force, fluid viscosity, density, and dimensions of the trapped object, stiffness of the optical trap, and frequency response of the instruments used to control the motion of the viscous medium to the accuracy of the calibration. The developed model can be adopted for calibration of the transverse trapping force, analysis of the trapped object motion, and reconstruction of a force profile during measurements of dynamic biological forces. PMID- 15298440 TI - Insulin minimal model indexes and secretion: proper handling of uncertainty by a Bayesian approach. AB - The identification of the insulin minimal model (MM) for the estimation of insulin secretion rate (ISR) and physiological indexes (e.g. beta-cell sensitivity) requires the knowledge of C-peptide (CP) kinetics. The four parameters of the two-compartment model of CP kinetics in a given individual can be derived either from an additional bolus experiment or, more frequently, from a population model. However, in both situations, the CP kinetics is uncertain and, in MM identification, it should be treated as such. This paper shows how to handle CP kinetics uncertainty by using a Bayesian methodology. In seven subjects, MM indexes and ISR were estimated together with their confidence intervals, using either the bolus data or the population model to assess CP kinetics. The two main results that arise from the application of the new methodology are: (i) the use of the population model in place of the bolus data to determine CP kinetics does not affect, on average, the point estimates of ISR profile and MM parameters but only the confidence intervals which becomes wider (less than 50%); (ii) in both the bolus and population situation neglecting the uncertainty of CP kinetics, as done in MM literature so far, introduces no bias, on average, on point estimates of MM indexes but only an underestimation of confidence intervals. PMID- 15298441 TI - Recommended levels of folic acid and vitamin B12 fortification: a PAHO/MOD/CDC technical consultation. PMID- 15298442 TI - Physiology of folate and vitamin B12 in health and disease. AB - Folate is a water-soluble B-vitamin and enzymatic cofactor that is necessary for the synthesis of purine and thymidine nucleotides and for the synthesis of methionine from homocysteine. Impairment of folate-mediated one-carbon metabolic pathways can result from B-vitamin deficiencies and/or single nucleotide polymorphisms, and increases risk for pathologies, including cancer and cardiovascular disease, and developmental anomalies including neural tube defects. Although several well validated metabolic and genomic biomarkers for folate deficiency exist, our understanding of the biochemical and genetic mechanisms whereby impaired folate metabolism increases risk for developmental anomalies and disease is limited, as are the mechanisms whereby elevated folate intake protects against these pathologies. Therefore, current initiatives to increase folate intakes in human populations to ameliorate developmental anomalies and prevent disease, while effective, lack predictive value with respect to unintended adverse outcomes. PMID- 15298443 TI - Folate and vitamin B12 recommended intakes and status in the United States. AB - This paper presents a synopsis of the Institute of Medicine (IOM) Dietary Reference Intakes (DRIs) for folate and the recommended folic acid intake to reduce the risk of neural tube defects. Estimated intake of folic acid is presented and total folate intake is compared to the DRI. Vitamin B12 DRIs are highlighted in addition to the IOM recommendation for crystalline vitamin B12 intake specifically for individuals > or =51 years of age. Recently reported evidence of vitamin B12 inadequacy in the aging segment of the U.S. population will be addressed. PMID- 15298444 TI - Oral synthetic folic acid and vitamin B12 supplements work--if one consumes them. AB - Oral supplements of synthetic folic acid and vitamin B12 are very effective in increasing blood levels of the vitamins and are known to prevent birth defects and cardiovascular diseases. PMID- 15298445 TI - Folate and vitamin B12 status in the Americas. AB - There is growing interest in the potential for folic acid fortification in the Americas and recognition of the high prevalence of low plasma vitamin B12 concentrations reported in various studies. This review summarized available data on plasma vitamin B12 and folate concentrations in the Americas. At least 40% of individuals had deficient or marginal plasma vitamin B12 concentrations in almost all locations and across age groups. Low plasma folate concentrations were less common. It is hypothesized that vitamin B12 deficiency may result from a low intake of animal source foods, while a higher intake of refined flour may result in low plasma folate. PMID- 15298446 TI - Folic acid food fortification in Canada. AB - By January 1998, most Canadian cereal grains (e.g., white flour) were being fortified with folic acid, with a large percentage being fortified by mid-1997. This was in compliance with both American and Canadian mandatory fortification deadlines of January and November 1998, respectively. It was estimated that between 0.1 to 0.2 mg of additional synthetic folic acid per day would be provided through this initiative, the goal of which was to lower the rate of neural tube defects (NTD). The current report outlines some of the changes to the health status of Canadians in relation to its folic acid food fortification initiative. PMID- 15298447 TI - The Costa Rican experience: reduction of neural tube defects following food fortification programs. AB - Fortification of wheat flour in 1997 and corn flour in 1999 with folic acid among other micronutrients was implemented in Costa Rica by means of two decrees, resulting in an effective public health impact. A prevalence of 25% of folic acid serum levels deficiency detected in fertile women in 1996 decreased 87% in urban areas two years later, whereas in rural areas diminished by 63%. In addition, a significant reduction of neural tube defects at the national level has been reported, dropping from a rate of 9.7 per 1000 lb during the period 1996-1998 to 6.3 per 1000 lb in the period 1999-2000. Finally, there has been a reported 74% reduction in the number of Neural Tube Defects at Birth (NTB) at the National Children's Hospital, resulting in 105 cases treated in 1995 to 26 cases in 2001. PMID- 15298448 TI - Folic acid fortification of wheat flour: Chile. AB - Neural tube defects (open spina bifida, anencephaly, and encephalocele) represent the first congenital malformations to be preventable through public health measures such as supplementation and/or food fortification with folic acid. In Chile, starting in January 2000, the Chilean Ministry of Health legislated to add folic acid to wheat flour (2.2 mg/kg) to reduce the risk of NTDs. This policy resulted in an estimated mean additional supply of 427 microg/d in significant increases in serum folate and red cell folate of 3.8 and 2.4-fold, respectively, in women of fertile age, one year after fortification. The impact on the rate of NTDs is presently being studied in all births, both live births and still births, with birth weight >500 g in the city of Santiago. Preliminary results show a reduction of 40% in the rates on NTDs from the pre-fortification period (1999 2000) to post-fortification period (2001-June 2002). Fortification of wheat flour with folic acid in Chile is effective in preventing NTDs in Chile. PMID- 15298449 TI - Modeling the level of fortification and post-fortification assessments: U.S. experience. AB - Mandatory fortification of enriched cereal-grain products became effective in the United States on January 1, 1998. This fortification was undertaken to assist women of child-bearing age in increasing their intake of folic acid to reduce their risk of having a pregnancy affected by a neural tube birth defect. The process by which the Food and Drug Administration modeled the level of fortification with folic acid illustrates the complex issues and general principles that emerge when fortification of a nation's food supply is evaluated as a means of addressing a public health concern. The effectiveness of fortification for a target population and safety for the much larger general population impose conflicting challenges that must be considered concurrently when making decisions regarding fortification. Recent data show improved folate status and apparent decreases in risk of neural tube birth defects in the U.S. Much about the long-term effects of the fortification program remains unknown and careful monitoring over time will be necessary to ensure that the program functions as intended. PMID- 15298450 TI - Recommended levels of folic acid and vitamin B12 fortification: conclusions. PMID- 15298451 TI - NJ reforms fall short of MD hopes. PMID- 15298452 TI - Cerumen removal. PMID- 15298453 TI - The quest for an easy EMR. PMID- 15298454 TI - Malpractice mess. Is this the way out? PMID- 15298455 TI - Big shoes to fill. PMID- 15298456 TI - All this for 108 dollars? PMID- 15298457 TI - The scary truth about tail coverage. PMID- 15298458 TI - Caring for Chinese, Japanese, and Korean patients. PMID- 15298459 TI - Should your patient still be driving? PMID- 15298460 TI - Office spirometry: don't just blow it off. PMID- 15298461 TI - Improving collections. PMID- 15298462 TI - What shouldn't go in your policy manual. PMID- 15298463 TI - Does the modified Ballard method of assessing gestational age perform well in a Zimbabwean population? AB - OBJECTIVE: To evaluate the performance and the utility of using birthweight adjusted scores of the Ballard method of estimating gestational age in a Zimbabwean population. DESIGN: A validation study. SETTING: Harare Maternity Hospital, from October to December 1999. SUBJECTS: Three hundred and sixty four African newborn infants, with a known last menstrual period (LMP), within 56 hours of life. MAIN OUTCOME MEASURES: Ballard scores obtained by examining the newly born infants compared to gestational age calculated from the last menstrual period. RESULTS: The Ballard method was a good predictor of gestational age, useful in differentiating term from preterm infants in the Zimbabwean population. There was a strong correlation between total neurological criteria (Pearson coefficient = 0.79), total physical criteria (Pearson coefficient = 0.77), total scores (Pearson coefficient = 0.81), with the gestational age calculated from the last menstrual period. The error of prediction of one single observation was 1.89 weeks. Our regression line for predicting gestational age was Y(LMP gestational age) = 24.493 + 0.420*score. Addition of birthweight to the linear regression model improved estimation of gestational age; Y(LMP gestational age)= 24.002 + 0.292*score + 0.0016*grams. The variance explained r2 was 0.63 and improved to 0.67 after addition of birthweight. CONCLUSION: The Ballard method can be used to differentiate pre-term from term infants at birth in a Zimbabwean population. The introduction of birthweight into the maturity scale improves assessment of gestational age and corrects error caused by low birthweight. We, therefore, recommend the use of our birthweight-adjusted Ballard maturity scales for routine clinical practice. PMID- 15298464 TI - Neonatal hypothermia levels and risk factors for mortality in a tropical country. AB - OBJECTIVE: To determine the prevalence of hypothermia, factors associated with hypothermia and risk factors for mortality in hypothermic newborn infants. STUDY DESIGN: Cross sectional descriptive study. STUDY SITE: Harare Central Hospital Neonatal Unit (NNU). SUBJECT: Three hundred and thirteen consecutive newborn infants admitted to the NNU for care. STUDY FACTORS: Temperature on admission to the NNU, mode of delivery, time of delivery, age on admission to the NNU, birth weight, sex, birth before arrival, need for resuscitation. RESULTS: Prevalence of hypothermia on admission was 85% with a mean axillary temperature of 34.3 degrees C (SD= 1.6). Median age on admission was 120 minutes and there was a case fatality rate of 18.3%. The need for resuscitation, age at admission to NNU, time of delivery, birth weight, sex and being born before arrival were not significantly associated with being hypothermic. The only factors that were associated with mortality were babies being born before arrival and birth weight below 1 500 gms. Age at admission to NNU, sex, time of delivery and need for resuscitation were not significantly associated with mortality. CONCLUSION: Neonatal hypothermia on admission remains a major problem in our population. There is need to increase awareness among nursing staff and mothers about the serious consequences of hypothermia particularly in very low birth weight newborns. Training in this area is called for. PMID- 15298465 TI - Factors associated with tumour stage at presentation in invasive cervical cancer. AB - OBJECTIVE: The main objective of the study was to determine the demographic, social, clinical, laboratory and histologic factors associated with late stage presentation in cervical cancer. DESIGN: A cross sectional study. SETTING: Government tertiary referral institutions, Harare, Zimbabwe STUDY POPULATION: One hundred consecutive cases of histology proven cervical cancer that presented for treatment between November 2001 and April 2002. MAIN OUTCOME MEASURES: The patients were categorized as early invasive cancer (stage I and II) and late invasive cancer (stage III and IV). RESULTS: The median age of the patients was 48 years (Q1=39 and Q3= 60). Eighty percent presented with late stage disease. Squamous cell carcinoma was the commonest histology (96%) with adenocarcinoma constituting only 4% of all tumours. Poorly differentiated tumour histology and no history of prior cervical cancer screening were found to be significantly associated with late tumour stage at presentation. The odds of presenting with late stage disease in women with a poorly differentiated tumour were 12.97 (95% CI 2.03 to 82.55; p = .007), whilst the odds of late stage presentation in the absence of a history of screening were 11.13 (95% CI 1.33 to 93.21; p = .026). CONCLUSIONS: Intrinsic tumour characteristics were the most important in this population in determining late stage at diagnosis and the value of screening was also highlighted by the results. The odds ratios had wide 95% confidence intervals, thus limiting their usefulness as point estimates. PMID- 15298466 TI - When is it herbal intoxication? A retrospective study of children admitted with herbal intoxication at Umtata General Hospital, South Africa. AB - OBJECTIVE: To establish the profile and fate of children admitted with a "herbal intoxication" label. DESIGN: Retrospective cross sectional study of records for all children admitted with "herbal intoxication" for the years 1998 to 2001 inclusive. SETTING: The Paediatrics Department at Umtata General Hospital, the main teaching hospital for the University of Transkei Faculty of Health Sciences. MAIN OUTCOME MEASURES: Demographic profile, herbal medication history and clinical progress and fate of children admitted with "herbal intoxication". RESULTS: There were 156 case records for the period under review. The age range was a few days to 12 years, being neonates 30 (19.2%); one to six months, 77 (49.4%); seven months to one year 34 (21.8%); one to five years, 11 (7.0%); five years and older, four (2.6%). The reasons for giving the herbal preparations were stated in only 52 cases (33.3%), of which five were for diarrhoea and vomiting, four for constipation and abdominal pain, three to change stool colour, and 41 for "playte" (generic term given to all types of ill health). The preparations were administered orally (n=95), by enema (n=12), and both orally and by enema (n= 12), while route of administration was not stated in 37 of the cases. Sixty eight (43.6%) of the children died, 84 (53.8%) improved, while the fate of four (2.6%) was not indicated. Most of the deaths (63.2%) were among the zero to six months age group, and most deaths occurred within two days of admission. The average duration of stay (days) in hospital was 2.94 +/- 0.65 (range 1 to 39) for the group that succumbed and 8.88 +/- 0.73 (range 1 to 40) for the survivors (p < 0.001). CONCLUSION: There is a high mortality rate among children admitted with suspicion of herbal intoxication, and there is need to manage such children with vigilance. PMID- 15298467 TI - Polyostotic fibrous dysplasia with auditory impairment and delayed menarche in a Nigerian. PMID- 15298468 TI - Healthworker's participation in voluntary counselling and testing in three Districts of Mashonaland East Province, Zimbabwe. F Tarwireyi and F Majoko. Cent Afr J Med 2003;49(5/6):58- 62. PMID- 15298469 TI - [The ethics of physician smoking. Why counsel against smoking?]. PMID- 15298470 TI - Outcome of cancer patients admitted to the intensive care unit (ICU). AB - This is a retrospective study aiming to determine the cause, the survival and the factors influencing survival in cancer patients admitted to an ICU. Between January 1998 and June 1999, 181 cancer patients were admitted to the ICU of HDF hospital (a general academic hospital of 300 beds). One hundred fifteen patients were admitted after surgery and 66 for medical reasons. Among the non-surgical group, 37 had solid tumors and 29 had hematological malignancies. Most of non surgical patients were admitted for respiratory or infectious complications due to their disease or treatment-related (59 patients). Among the non-surgical group of patients, 44 required mechanical ventilation (MV). The mortality rate was 41% during ICU recovery, 62% during the hospitalization period, 73% at 2 months from discharge and 83% at last follow-up. The duration of stay in the ICU was the only factor affecting mortality. Age, disease type and MV did not influence the mortality rate in this population. PMID- 15298471 TI - Cancer patients and critical care medicine. PMID- 15298472 TI - [Surgical wound infection in cardiac surgery. A prospective multidisciplinary study]. AB - In an attempt to define the incidence of surgical wound complications following open heart surgery, 210 consecutive patients were included in a prospective monocentric study. All were adults with a mean age of 61 years. The different variables incriminated in pathogenesis of surgical wound infections were analyzed in order to evaluate their respective influence. No cases of mediastinitis were noted. The incidence of surgical wound infections was 5.5%. Body mass index > 25 and extensive surgical saphenous dissection in the lower extremities were the only two factors found to show as statistical significance. PMID- 15298473 TI - [Two-year follow-up study of postoperative rehabilitation, return to work, and athletic activities of 111 Lebanese patients after coronary bypass]. AB - The goal of this paper is to study the progress of the first phase of rehabilitation of a Lebanese population treated by coronary artery bypass surgery, as well as to follow long-term socioprofessional reinsertion and resumption of physical activity in these patients. 111 patients operated in our hospital of coronary artery bypass in 1997 have been studied retrospectively two years after the surgery. The analyzed parameters are socioprofessional demographic data, physical and sports activities before and after the bypass surgery as well as the applied rehabilitation program. Among the studied patients (mean age = 64 years, 77% of men), 30% only had a professional activity before the surgery. The majority of these patients returned to work in a mean delay of 45 days. 76% of the patients have regular physical activity two years after the surgery versus 50% before. On the 63 smoking patients before the bypass surgery, 41 continue to smoke 2 years later. The behavior of the Lebanese patients after a coronary artery bypass surgery is particular and different from what is reported in the literature: 1) The resumption of the initial work is more frequent and more precocious. 2) The patients start regular physical activity without surveillance or previous codification of the frequency and the intensity of the effort. 3) The majority of the smoking patients continue to smoke after surgery. 4) The functional rehabilitation is limited to the phase I. PMID- 15298474 TI - [Role of endoscopic gastric biopsies in the management of gastritis. A study of 250 consecutive cases]. AB - BACKGROUND: The practical role of gastric biopsy in the management of gastritis is controversial. AIM: To estimate the yield of endoscopic biopsies in the clinical, endoscopic and pathologic approach of gastritis. MATERIAL AND METHODS: Prospective study of 250 consecutive patients who underwent an upper G.I. endoscopy between July 1996 and January 1997, for upper G.I. symptoms, miscellaneous manifestations requiring an upper G.I. endoscopy or presenting a gastritis on EGD performed for other indications. Every patient had 6 biopsies: 2 in the antrum, 2 in the corpus, and 2 in an intermediate zone. RESULTS: After defining the abnormal elemental endoscopic and pathologic patterns, gastric mucosa was endoscopically normal in 57 cases (22.8%) and abnormal in the remaining of the 250 cases (77.2%). The pathologic findings were normal in 69 cases (27.6%) and abnormal in the remaining 181 cases of 250. H. pylori was found in 126 cases (50.4%), 10 cases of which (7.9%) had normal pathology. There was no significant correlation between clinical symptoms, endoscopy and pathology. There was a correlation between endoscopic abnormalities and tobacco use (P = 0.0073), NSAIDs use (P = 0.0001) and the presence of H. pylori (P < 0.0001). There was also a correlation between pathologic findings, tobacco use (P = 0.0015), NSAIDs use (P = 0.0022) and the presence of HP (P < 0.0001). On the other hand, there was a correlation between the presence of an inflammatory infiltrate in H. pylori gastritis (P = 0.0007) and its absence in NSAIDs use (P = 0.0003). The correlation between endoscopy and pathology existed only for certains patterns: erosion and ulcerations (P = 0.0002), purpuric (P = 0.033), congestion (P < 0.0001) and mosaic (0.0095). CONCLUSION: Gastric biopsy brings no important practical supplement to endoscopic examination except in revealing the presence of H. pylori. It adds nothing to endoscopy in helping explain the clinical symptoms. But it is obvious that it may reveal some serious pre-malignant dysplasia or malignant gastric lesions (maltoma, linitis plastica). This did not occur during our study. PMID- 15298475 TI - DNA-based subtypes and antimicrobial susceptibility profiles of Haemophilus influenzae and Haemophilus parainfluenzae isolated from different tonsillar sites of children undergoing tonsillectomy and/or adenoidectomy. AB - We did a comparative analysis between DNA-based subtypes and antimicrobial susceptibility profiles on Haemophilus influenzae and Haemophilus parainfluenzae, isolated from multiple tonsillar sites per individual from patients with chronic recurrent tonsillitis and/or tonsillar idiopathic hypertrophy and undergoing tonsillectomy and/or adenoidectomy. A total of eighty-eight Haemophilus isolates were obtained aseptically from the surface and core of tonsils and/or adenoids of 32 out of 60 patients and identified at the species level by the X and V factors and the API NH Kit. The H. influenzae and H. parainfluenzae isolates as well as ATCC strains were tested for antimicrobial susceptibility using a panel of antimicrobial agents. Random amplified polymorphic DNA (RAPD) was done on extracted DNA from all Haemophilus isolates and ATCC strains, using one 10 mer and one 18 mer primers to subtype the two species. Antimicrobial susceptibility testing data have shown a variation in generated susceptibility patterns to tested antimicrobial agents among H. influenzae and H. parainfluenzae isolates. This variation was demonstrated too among isolates obtained from different tonsillar sites (core and surface) in a single patient. RAPD analysis identified 58/88 (66%) different RAPD patterns. Variations in RAPD patterns among H. influenzae and H. parainfluenzae were also observed in isolates obtained from different tonsillar sites of the same individual. A correlation between RAPD patterns and antimicrobial susceptibility data, have shown: 1) the predominance of one strain (RAPD pattern) of either Haemophilus species among isolated organisms per patient, and exhibiting different antimicrobial susceptibility profiles or 2) the existence of multiple strains (RAPD patterns) of either Haemophilis species per patient, and showing either a single or multiple antimicrobial susceptibility profile(s). These observations question the validity of swab cultures obtained from a single tonsillar site per patient, for detection, identification and determination of antimicrobial profiles of the etiology of tonsillitis, since swab specimens taken from only one site may or may not reflect the etiology of infection. PMID- 15298476 TI - [Acute asthma attack in children presenting to the emergency department. Survey of 96 cases]. AB - Throughout a 2-year period, children who presented at Hotel-Dieu de France emergency department (ED) with acute asthma were analyzed prospectively and data on their environment, family and personal history as well as treatment were recorded. Treatment delivered at the ED, response and further outcome were analyzed. Out of 2024 children aged less than 15 years, 96 (5%) had acute asthma attack. Their median age was 4 years and M/F ratio was 2:1. Median age at onset of asthma was 2 years. Only 66 patients were recognized as asthmatics and 20% were given regular inhaled daily treatment. Current attack was mild in 45%, moderate in 45% and severe in 10% of cases. Home treatment before ED admission was often inadequate. Nine patients required hospital admission after failure of treatment at the ED. Within a median follow-up of 12 months, half of the patients experienced further attacks sometimes requiring ED care (27%) or hospital admission (8%). These data highlight the fact that asthma in our country is still largely under recognized and inadequately treated. PMID- 15298477 TI - Is MRI effective in detecting intraarticular abnormalities of the injured knee? AB - BACKGROUND DATA: Magnetic resonance imaging (MRI) is widely used for the diagnosis of intraarticular anomalies of the knee. Its reliability is controversial mainly in the pediatric population, and its results may vary from one center to the other. The purpose of this study was to assess the validity of MRI in detecting intraarticular abnormalities in patients who sustained an injury of their knee, by comparing its findings to those of arthroscopy. MATERIAL AND METHODS: Two hundred and seventeen traumatic knees underwent MRI followed by arthroscopy. Findings of MRI and arthroscopy were statistically compared. RESULTS: Accuracy of MRI was 95.39% and 99.08% for anterior and posterior cruciate ligament ruptures respectively, 85.25% and 90.78% for medial and lateral meniscal tears respectively, and 91.24% for injury of the articular cartilage. There was not a significant difference between the pediatric and the adult populations, except for a greater sensitivity to detect lateral meniscal tears and a better specificity and accuracy to identify chondral lesions in the first group. Variability between centers was significant for the diagnosis of chondral defects. CONCLUSION: The good MRI reliability in detecting intraarticular abnormalities as demonstrated in this report, underlines its importance in the assessment of the injured knee before arthroscopy. PMID- 15298478 TI - Hypoxic cells in tumors as a target for cancer therapy. AB - Hypoxic cells that are found in solid tumors are resistant to anticancer drugs and radiation therapy. Thus, for effective anticancer chemotherapy, it is important to identify drugs with selective toxicity towards hypoxic cells. The recent development of new drugs that are toxic only when activated in the hypoxic cell opens a new era of cancer treatment. Recently, we evaluated the hypoxia selective toxicity of four differently substituted quinoxaline 1,4-dioxides (QdNOs) in human cancer cells. These compounds were synthesized by the Beirut Reaction. The various QdNOs were found to exert potent hypoxic cytotoxic activities against human colon cancer cells (T-84) and to possess a 50-100 fold greater cytotoxicity under hypoxia compared to oxia. Interestingly, the hypoxia cytotoxicity ratio (HCR: ratio between drug concentration in air and in hypoxia to give 10% cell survival) of these compounds was found to depend on the nature of the substituents on the quinoxaline 1,4-dioxide heterocycle. Because of their differential hypoxic cytotoxicity, these drugs could provide useful therapeutic agents against solid tumors. Presently we are investigating the selective cytotoxicity of QdNOs for hypoxic cells in tumors in vivo and their ability to potentiate radiation-induced tumor cell killing. We will also study their in vitro anti-angiogenic activity and their mechanism of action at the molecular level. The deciphering of the mechanism of action of QdNOs may allow us to ultimately recommend their use as therapeutic agents against human tumors. PMID- 15298479 TI - [Difficulties of the new classification system for bladder cancer]. AB - Bladder cancer is a public health problem because of its increasing incidence. The histological grade being an important prognostic parameter in evaluating mainly superficial, non-infiltrative, urothelial tumors, it is of great interest to define a reproducible classification that can be made available to oncologists, urologists and family doctors as well. New classifications have been recently proposed. They will be discussed here. It will be argued that the current classification is still valid and should be maintained. PMID- 15298480 TI - [Percutaneous closure of a large ductus arteriosus with the Amplatzer device]. AB - We describe here the first Lebanese experience of percutaneous closure of a large patent ductus arteriosus in a 15-year-old female. The ductus arteriosus was successfully occluded using the new Amplatzer Duct Occluder system. The procedure time was 40 minutes and the young patient was discharged home the next day. This new device, conceived since 1997, gives very encouraging results according to the recent medical literature. PMID- 15298481 TI - [Peritoneal pseudomyxoma: two case reports and review of the literature]. AB - In this article, we present two cases of peritoneal pseudomyxoma suspected on abdominal CT Scan and then confirmed on pathologic examination. A mucinus carcinoma was the primary lesion in the two cases, appendicular in the first case and with an undeterminate origin in the second. The prognosis was bad in these cases despite the debulking surgery and the systemic and intraperitoneal chemotherapy. PMID- 15298482 TI - Sudden infant death syndrome in Canada: trends in rates and risk factors, 1985 1998. AB - In Canada, sudden infant death syndrome (SIDS) remains the leading cause of postneonatal death. However, SIDS rates have been declining in many countries, including Canada. This decline has been largely attributed to recommendations to avoid placing infants to sleep in the prone position. We examined the postneonatal rate of mortality due to SIDS and to other causes in relation to the initial risk reduction campaign. The postneonatal mortality rate due to SIDS decreased from 0.97 to 0.54 per 1,000 neonatal survivors between 1985-1989 and 1994-1998 (relative risk [RR] = 0.56, 95% confidence interval [CI] 0.51-0.62). The rate of postneonatal mortality due to other causes also decreased during the same period, though to a smaller extent, from 1.19 to 0.86 (RR = 0.72, 95% CI 0.66-0.78). With the exception of seasonality, established risk factors for SIDS remained essentially unchanged between the two time periods. The observed reduction in postneonatal SIDS is consistent with a positive impact of the initial recommendations regarding risk reduction. However, the lack of reliable risk factor data limits the extent to which the decline can be attributed directly to the campaign. PMID- 15298483 TI - A model for non-communicable disease surveillance in Canada: the prairie pilot diabetes surveillance system. AB - The Prairie Pilot Diabetes Surveillance Project was organized to design and test a prototype population-based surveillance system, using administrative data, for a chronic disease exemplar - diabetes mellitus. The Canadian model of a public health surveillance system for chronic conditions described here specifies a process by which administrative and claims data arising from provincial health insurance programs are merged into an annual person-level summary file (APLSF), yielding one summary record for each person insured within each province. The APLSF is the basis for a variety of estimates, including incidence, prevalence, mortality, complication rates and health services utilization. The model was used to produce comparable interprovincial estimates of several parameters with respect to diabetes for the entire population in the provinces of Alberta, Manitoba and Saskatchewan. All processing of identifiable health data occurred within the provinces where the data were generated. Combining results across provinces was based on further aggregation of the summary data from each province and not by pooling of identifiable person-level data. On the basis of preliminary outputs for diabetes mellitus, the model appears to provide coherent estimates of key diabetes parameters and reflects anticipated differences in health services and outcomes, by disease state. Three characteristics of the model recommend it as a resource for non-communicable disease surveillance in Canada: a) it maximizes the utility of existing data; b) it includes both those with and those without the disease in question; and c) it respects provincial legislation regarding personal health data, yet permits reporting of multi-provincial, population-based data. PMID- 15298484 TI - Refining the measurement of the economic burden of chronic diseases in Canada. AB - This article presents an analysis of the economic burden of a number of chronic diseases in Canada. In the analysis, we adjusted our measure of utilization of physician and hospital services for co-existing chronic diseases, which we found to be widely prevalent and to have an impact on resource use. Using data from the 1999 National Population Health Survey, we developed resource use rankings for several chronic conditions and decomposed these measures into prevalence and per person utilization components. Our results indicate that, for the diseases with the greatest impact, resource use measures are driven more by disease prevalence than intensity of resource use. The diseases with the highest overall degree of resource use are back pain, arthritis or rheumatism, high blood pressure and migraines for people under 60; and arthritis or rheumatism and high blood pressure for people over 60. Our methods can be used to forecast the overall relative impact of resource use due to disease prevalence and per-person resource use intensity for various conditions. PMID- 15298485 TI - Rates of claims for cumulative trauma disorder of the upper extremity in Ontario workers during 1997. AB - Surveillance of work-related cumulative trauma disorder of the upper extremity (CTDUE) requires valid and reliable claim extraction strategies and should examine for confounding and interaction. This research estimated crude and specific rates of CTDUE claims in Ontario workers during 1997 while acknowledging misclassification and testing for confounding and interaction. Lower and upper limit event estimates were obtained by means of an algorithm applied to the Ontario Workplace Safety and Insurance Board (OWSIB) database and were combined with "at-risk" estimates obtained from the Canadian Labour Force Survey (LFS). Poisson regression was used to evaluate confounding and interaction. The method used to identify CTDUE claims had a substantial impact on the magnitude of rates, female to male rate ratios, the most commonly affected part of the upper extremity and the highest risk occupational categories. Poisson regression identified sex interactions. It allowed rigorous evaluation of the data and indicated that rates should be examined separately for men and women. Researchers should clearly define extraction strategies and examine the impact of misclassification. PMID- 15298486 TI - Rates and external causes of blunt head trauma in Ontario: analysis and review of Ontario Trauma Registry datasets. AB - Contemporary studies of blunt head trauma and its determinants are important for prevention. It is also important to understand the strengths and limitations of the common sources of data used for the ongoing study of these injuries. Using the Ontario Trauma Registry, we described frequent patterns of blunt head trauma and identified priorities for prevention and research. A review of methodological issues that arose during the analysis of these trauma registry data is also provided. Blunt head trauma cases were identified within two data sets of the Ontario Trauma Registry. The Minimal Data Set is population-based and contains acute care injury hospitalizations, and the Comprehensive Data Set contains "major injuries" treated at a lead trauma hospital. Injury control priorities varied by age group, sex and data set and these are profiled in the manuscript. The results indicate the importance of examining multiple sources of surveillance data in establishing injury control priorities. The methodological review demonstrated the need to critically examine the completeness and accuracy of trauma registry data in arriving at decisions about priorities. PMID- 15298487 TI - The frequent expression of cancer/testis antigens provides opportunities for immunotherapeutic targeting of sarcoma. AB - Sarcomas are rare but aggressive malignant tumors associated with high mortality, for which the efficacy of standard therapies remains limited. In order to develop immunotherapeutic approaches for the treatment of sarcoma, we studied the relevance of cancer/testis antigens (CTAs), a group of antigens whose expression is developmentally regulated and that is specifically found in some tumor types, as sarcoma vaccine targets. CTA expression was assessed by PCR and/or immunohistochemistry (IHC) in sarcoma tumor samples that included different histological subtypes and sarcoma cell lines. Expression of HLA class I was assessed by IHC in tumor samples and by FACS analysis in cell lines. More than 70% of the tumor samples expressed at least one CTA. The majority of tumors and cell lines expressed normal levels of HLA class I. HLA class I expression in cell lines was enhanced upon treatment with IFN-gamma. CTA expression was enhanced or induced by treatment with the demethylating agent 5-aza-2'-deoxycytidine, resulting in recognition by specific CTLs. Interestingly, a spontaneous humoral and CD8+ T cellular response to the CTA NY-ESO-1 was detected in a synovial sarcoma patient. Together, these findings strongly support the implementation of CTA-based immunotherapy of sarcoma as a means to improve the efficacy of the standard therapy. PMID- 15298489 TI - Gamma-hydroxybutyric acid: neurobiology and toxicology of a recreational drug. AB - gamma-Hydroxybutyric acid (GHB) is a short-chain fatty acid that occurs naturally in mammalian brain where it is derived metabolically from gamma-aminobutyric acid (GABA), the primary inhibitory neurotransmitter in the brain. GHB was synthesised over 40 years ago and its presence in the brain and a number of aspects of its biological, pharmacological and toxicological properties have been elucidated over the last 20-30 years. However, widespread interest in this compound has arisen only in the past 5-10 years, primarily as a result of the emergence of GHB as a major recreational drug and public health problem in the US. There is considerable evidence that GHB may be a neuromodulator in the brain. GHB has multiple neuronal mechanisms including activation of both the gamma-aminobutyric acid type B (GABA(B)) receptor, and a separate GHB-specific receptor. This complex GHB-GABA(B) receptor interaction is probably responsible for the protean pharmacological, electroencephalographic, behavioural and toxicological effects of GHB, as well as the perturbations of learning and memory associated with supra physiological concentrations of GHB in the brain that result from the exogenous administration of this drug in the clinical context of GHB abuse, addiction and withdrawal. Investigation of the inborn error of metabolism succinic semialdehyde deficiency (SSADH) and the murine model of this disorder (SSADH knockout mice), in which GHB plays a major role, may help dissect out GHB- and GABA(B) receptor mediated mechanisms. In particular, the mechanisms that are operative in the molecular pathogenesis of GHB addiction and withdrawal as well as the absence seizures observed in the GHB-treated animals. PMID- 15298490 TI - Gamma-butyrolactone and 1,4-butanediol: abused analogues of gamma hydroxybutyrate. AB - gamma-Hydroxybutyrate (GHB) is a GABA-active CNS depressant, commonly used as a drug of abuse. In the early 1990s, the US Drug Enforcement Administration (DEA) warned against the use of GHB and restricted its sale. This diminished availability of GHB caused a shift toward GHB analogues such as gamma butyrolactone (GBL) and 1,4-butanediol (1,4-BD) as precursors and surrogates. Both GBL and 1,4-BD are metabolically converted to GHB. Furthermore, GBL is commonly used as a starting material for chemical conversion to GHB. As such, the clinical presentation and management of GBL and 1,4-BD intoxication shares a great deal of common ground with that for GHB. This similarity exists not only for acute intoxication but also for withdrawal in those patients with a history of extended high-dose abuse. This review examines the history of GHB analogue abuse as well as the clinical presentation and management of acute intoxication and withdrawal associated with abuse of these compounds. PMID- 15298491 TI - Gamma-hydroxybutyrate: bridging the clinical-analytical gap. AB - Laboratory detection of gamma-hydroxybutyrate (GHB) has been published as early as the 1960s. However, wide-scale use of GHB during the 1990s has led to the development of current analytic methods to test for GHB and related compounds. Detection of GHB and related compounds can be clinically useful in confirming the cause of coma in an overdose patient, determining its potential role in a postmortem victim, as well as evaluating its use in a drug-facilitated sexual assault victim. Analytical method sensitivity must be known in order to determine the usefulness and clinical application. Most laboratory cut-off levels are based on instrument sensitivity and will not establish endogenous versus exogenous GHB levels. Interpretation of GHB levels must include a knowledge base of endogenous GHB, metabolism of GHB and related compounds, as well as postmortem generation. Due to potential analytical limitations in various GHB methods, it is clinically relevant to specifically request for GHB as well as related GHB compounds if they are also in question. Various storage conditions (collection time, types of containers, use of preservatives, storage temperature) can also affect the analysis and interpretation of GHB and related compounds. PMID- 15298492 TI - The gamma-hydroxybutyrate withdrawal syndrome. AB - gamma-Hydroxybutyrate (GHB) is endogenous inhibitory transmitter that, when administered in pharmacological doses, has sedative-hypnotic properties. It is used in anaesthesia for the treatment of narcolepsy/catalepsy and in alcohol/opioid detoxification treatment regimens. Based on its purported anabolic effects, GHB use became established among bodybuilders. As the euphorigenic effects of GHB became publicised, attendees at dance clubs and rave parties began to use it alone or in combination with other psychoactive drugs. Following the ban of GHB in 1990, several precursor products (e.g. gamma-butyrolactone, butanediol) became widely used as replacement drugs until their ultimate proscription from lawful use in 2000. GHB and its precursors, like most sedative hypnotic agents, can induce tolerance and produce dependence. Although many GHB users will experience a mild withdrawal syndrome upon drug discontinuation, those with chronic heavy GHB use can experience severe withdrawal. This syndrome clinically resembles the withdrawal syndrome noted from alcohol and other sedative-hypnotic drugs (e.g. benzodiazepines). Distinct clinical features of GHB withdrawal are its relatively mild and brief autonomic instability with prolonged psychotic symptoms. Patients with fulminant GHB withdrawal require aggressive treatment with cross-tolerant sedative hypnotics, such as benzodiazepines. PMID- 15298493 TI - Hydrogen peroxide poisoning. AB - Hydrogen peroxide is an oxidising agent that is used in a number of household products, including general-purpose disinfectants, chlorine-free bleaches, fabric stain removers, contact lens disinfectants and hair dyes, and it is a component of some tooth whitening products. In industry, the principal use of hydrogen peroxide is as a bleaching agent in the manufacture of paper and pulp. Hydrogen peroxide has been employed medicinally for wound irrigation and for the sterilisation of ophthalmic and endoscopic instruments. Hydrogen peroxide causes toxicity via three main mechanisms: corrosive damage, oxygen gas formation and lipid peroxidation. Concentrated hydrogen peroxide is caustic and exposure may result in local tissue damage. Ingestion of concentrated (>35%) hydrogen peroxide can also result in the generation of substantial volumes of oxygen. Where the amount of oxygen evolved exceeds its maximum solubility in blood, venous or arterial gas embolism may occur. The mechanism of CNS damage is thought to be arterial gas embolisation with subsequent brain infarction. Rapid generation of oxygen in closed body cavities can also cause mechanical distension and there is potential for the rupture of the hollow viscus secondary to oxygen liberation. In addition, intravascular foaming following absorption can seriously impede right ventricular output and produce complete loss of cardiac output. Hydrogen peroxide can also exert a direct cytotoxic effect via lipid peroxidation. Ingestion of hydrogen peroxide may cause irritation of the gastrointestinal tract with nausea, vomiting, haematemesis and foaming at the mouth; the foam may obstruct the respiratory tract or result in pulmonary aspiration. Painful gastric distension and belching may be caused by the liberation of large volumes of oxygen in the stomach. Blistering of the mucosae and oropharyngeal burns are common following ingestion of concentrated solutions, and laryngospasm and haemorrhagic gastritis have been reported. Sinus tachycardia, lethargy, confusion, coma, convulsions, stridor, sub-epiglottic narrowing, apnoea, cyanosis and cardiorespiratory arrest may ensue within minutes of ingestion. Oxygen gas embolism may produce multiple cerebral infarctions. Although most inhalational exposures cause little more than coughing and transient dyspnoea, inhalation of highly concentrated solutions of hydrogen peroxide can cause severe irritation and inflammation of mucous membranes, with coughing and dyspnoea. Shock, coma and convulsions may ensue and pulmonary oedema may occur up to 24-72 hours post exposure. Severe toxicity has resulted from the use of hydrogen peroxide solutions to irrigate wounds within closed body cavities or under pressure as oxygen gas embolism has resulted. Inflammation, blistering and severe skin damage may follow dermal contact. Ocular exposure to 3% solutions may cause immediate stinging, irritation, lacrimation and blurred vision, but severe injury is unlikely. Exposure to more concentrated hydrogen peroxide solutions (>10%) may result in ulceration or perforation of the cornea. Gut decontamination is not indicated following ingestion, due to the rapid decomposition of hydrogen peroxide by catalase to oxygen and water. If gastric distension is painful, a gastric tube should be passed to release gas. Early aggressive airway management is critical in patients who have ingested concentrated hydrogen peroxide, as respiratory failure and arrest appear to be the proximate cause of death. Endoscopy should be considered if there is persistent vomiting, haematemesis, significant oral burns, severe abdominal pain, dysphagia or stridor. Corticosteroids in high dosage have been recommended if laryngeal and pulmonary oedema supervene, but their value is unproven. Endotracheal intubation, or rarely, tracheostomy may be required for life threatening laryngeal oedema. Contaminated skin should be washed with copious amounts of water. Skin lesions should be treated as thermal burns; surgery may be required for deep burns. In the case of eye exposure, the affected eye(s) shod eye(s) should be irrigated immediately and thoroughly with water or 0.9% saline for at least 10-15 minutes. Instillation of a local anaesthetic may reduce discomfort and assist more thorough decontamination. PMID- 15298494 TI - Pharmaceutical drug overdose case reports: from the world literature. PMID- 15298496 TI - Suitability of 7 aortic stent-graft models for MRI-based surveillance. AB - PURPOSE: To evaluate the magnetic resonance imaging (MRI) characteristics of commercially available stent-grafts used for abdominal aortic aneurysm repair. METHODS: Seven endovascular grafts (AneuRx, Lifepath, Talent, Excluder, Zenith, Quantum LP, and Ancure) were suspended in a water bath containing gadolinium and scanned using a 1.5-T clinical MRI scanner. Two different scan techniques (T(1) weighted spoiled gradient echo and spin echo) based upon a clinical MRI endograft surveillance protocol were used for each stent-graft. The scans were evaluated for susceptibility artifacts and radiofrequency (RF) shielding and caging artifacts. RESULTS: For most endografts, the lumen and structures surrounding the endograft were well visualized. However, the ferromagnetic properties of the Zenith and Lifepath devices resulted in large susceptibly artifacts that obliterated the endograft lumen as well as adjacent structures. All fully supported grafts showed some amount of signal loss from the graft lumen caused by RF caging. For the Ancure graft, evaluation around the attachment sites might be problematic. CONCLUSIONS: An MRI-based surveillance protocol appears to be a viable option for the AneuRx, Talent, Excluder, and Quantum LP stent-grafts. PMID- 15298497 TI - Measurement of aortic compliance in abdominal aortic aneurysms before and after open and endoluminal repair: preliminary results. AB - PURPOSE: To assess aortic wall compliance as a portent of rupture risk in patients with abdominal aortic aneurysms. METHODS: In this pilot study, 38 patients (32 men; median age 78 years, range 63-95) underwent an ultrasound scan: 20 pre-repair and 24 post-repair (18 endovascular [EVR] and 6 open). Six patients from the pre-repair group were included in a post repair study after EVR. Cine loop images were analyzed offsite using wall tracking software, which measured aortic diameter changes during cardiac cycles. Brachial blood pressure was measured, and elastic modulus (Ep) and stiffness (beta) were calculated. Preop Ep and beta were determined at the neck, inflection points (IP), and mid sac levels. Postop Ep and beta were calculated in mid sac only for technical reasons. RESULTS: Preoperative Ep and beta were significantly higher at IP compared with neck (median Ep 24.22 versus 12.95 N/cm(2), p<0.003; median beta 16.27 versus 8.65, p<0.003). At the mid sac, Ep and beta were also significantly higher compared with neck: Ep 26.41 versus 12.95 N/cm(2), p=0.001; beta 17.94 versus 8.65, p=0.001. The values for IP and mid sac were Ep 24.22 versus 26.41 N/cm(2), p=0.64; beta 16.27 versus 17.94, p=0.64. In the postop cases (n=24), Ep and beta in successful endovascular repair (n=12) were significantly higher than in open repair, respectively: median Ep 34.31 versus 12.33 N/cm(2), p<0.001; median beta 23.18 versus 8.24, p<0.001. Patients with endoleaks or endotension (n=6) had significantly elevated Ep and beta compared with those without endoleaks (n=12): median Ep 79.79 versus 34.31 N/ cm(2), p=0.002; median beta 51.52 versus 23.18, p<0.002. Six patients scanned before and after EVR showed a decrease of Ep and beta in 3, no change in 1, and an increase in 2. An increase greater than 2 fold was noted in a patient with a gross type II endoleak. CONCLUSIONS: This pilot study shows that estimates of aortic wall compliance agree well with known values for wall stress distribution. EVR leaves patients with greater wall stiffness than those undergoing open repair, a situation accentuated by endoleaks. Wall compliance and stiffness measurement promises to be useful for the evaluation of success of endovascular repair. PMID- 15298498 TI - Subintimal angioplasty as a primary modality in the management of critical limb ischemia: comparison to bypass grafting for aortoiliac and femoropopliteal occlusive disease. AB - PURPOSE: To compare the 30-day morbidity, mortality, length of hospital stay, and patency rates in patients with critically ischemic limbs treated with subintimal angioplasty (SA) versus standard bypass surgery. METHOD: Between October 2001 and August 2003, 137 patients (74 women; mean age 70 years, range 43-92) with critical limb ischemia underwent subintimal angioplasty (n=88) or bypass surgery (n=49) for superficial femoral artery (SFA) or aortoiliac lesions. All patients had lesions classified as C or D according to the TransAtlantic Inter-Society Consensus. Data was retrieved from hospital inpatient inquiry and VascuBase. Parallel group comparison was used in performing a prospective observational study. RESULTS: Primary technical success was 100% for both SA and bypass grafting. Thirty-day survival was 100% in the SFA-SA and aortoiliac bypass groups and 96% and 93%, respectively, in the SFA bypass and aortoiliac SA groups. Limb salvage was 97% and 82% in the SFA-SA and SFA bypass groups, respectively; at the aortoiliac levels, the rates were 100% and 86% for SA versus bypass. Subintimal angioplasty significantly reduced hospital stay (p<0.001). Primary patency was not statistically different in the SA versus bypass groups; however, secondary patency was higher in the SFA bypass group. CONCLUSIONS: Frequent clinical follow up and a duplex surveillance program are necessary to maintain patency in this cohort. Subintimal angioplasty is increasingly replacing bypass surgery in the management of critical limb ischemia without compromising primary patency, limb salvage, patient survival, or subsequent vascular intervention. PMID- 15298499 TI - Post-endograft abdominal aortic aneurysm shrinkage varies among hospitals: observations from multicenter trials. AB - PURPOSE: To evaluate differences in abdominal aortic aneurysm (AAA) shrinkage among hospitals following protocol-driven patient selection and using endografts from a single manufacturer. METHODS: Standardized inclusion criteria for the Talent endograft multicenter trials included AAA diameter >/=40 mm and proximal neck limits of length >/=5 mm, diameter 14 to 32 mm, and angle /=5-mm decrease in the AAA largest minor axis diameter. Trial sites with >10 complete study cases were selected for stepwise logistic regression analysis. In the 13 trial sites meeting this criterion, 323 patients (mean age 74; 93% men) were treated for aneurysms with a mean pretreatment diameter of 53 mm. RESULTS: At 12 months, significant AAA shrinkage occurred in 192 (59%) cases. The AAA shrinkage rate was 71% to 82% at 3 sites, 60% to 64% at 4 sites, 45% to 50% at 4 sites, and 35% and 27% at the 2 remaining sites. In the multivariate analysis, the hospital site showed a strong, independent association with aneurysm shrinkage (p<0.04). Neck and pretreatment AAA diameters were also found to be important factors (p<0.04). Age, gender, AAA classification, neck length, and angle were not significant correlates. Sixty four (20%) endoleaks (29 type I, 34 type II, and 1 type III) were observed. The incidence of proximal endoleak was significantly different among sites (p<0.001) and highest in the 3 sites with the lowest AAA shrinkage rate. CONCLUSIONS: AAA shrinkage rates vary significantly among hospitals using the same endograft and protocol-defined patient selection criteria. Site-specific factors appear to be an important variable leading to successful endograft repair, as defined by post endograft aneurysm shrinkage. PMID- 15298500 TI - Ultrasound navigation through the heart for off-pump aortic valved stent implantation: new tools for new goals. AB - PURPOSE: To validate the use of simultaneous intracardiac and intravascular ultrasound (IVUS) guidance for periprocedural dimension assessment, Residual Coronary/Sinus-Stent Index (RCSSI) estimation, target site identification, deployment monitoring, and quality control of off-pump aortic Valved Stent implantation. METHODS: Five pigs (56+/-5 kg) underwent off-pump orthotopic aortic valve implantation using a custom-made self-expanding Valved Stent. Intracardiac ultrasound (AcuNav) was introduced via the right femoral vein. After left-sided thoracotomy, pursestring sutures were placed on the left ventricular apex. Following heparinization, a guidewire was inserted through the apex and advanced over the aortic valve under fluoroscopy. A wire-guided IVUS catheter transducer (6-F, 12.5-MHz) was inserted and the aortic target site identified. IVUS probe location was tracked with AcuNav, and measures of the aortic root were taken by both. After removal of the IVUS, the Valved Stent delivery system was introduced over the guidewire under fluoroscopy and AcuNav monitoring; the Valved Stent was deployed over the native valves. In vivo assessment included leaflet motion, planimetric valve orifice and RCSSI (stent to aortic wall distance/coronary diameter) calculations, coronary blood flow characteristics, transvalvular gradient, regurgitation, and paravalvular leaking in combination with continuous cardiac output measures. Macroscopic analysis was performed at necropsy. RESULTS: IVUS dimensions of the aortic root were equal to AcuNav and necropsy dimensions. Both tools showed good valvular function, with full valvular opening and closing in 3 of 5 valves. At necropsy, the 3 aortic Valved Stents were safely anchored. One Valved Stent was placed supra-annularly; 2 dislodged into the left ventricle because of size mismatch. One Valved Stent showed a moderate to severe paravalvular leak. CONCLUSIONS: Simultaneous intracardiac and intravascular ultrasound-guided off-pump orthotopic aortic Valved Stent implantation via left sided thoracotomy is feasible in an animal model. IVUS and intracardiac ultrasound allow adequate aortic dimension assessment and Valved Stent delivery monitoring, as well as postimplantation coronary flow evaluation. PMID- 15298501 TI - Heterogeneity of tensile strength and matrix metalloproteinase activity in the wall of abdominal aortic aneurysms. AB - PURPOSE: To measure the tensile strength of the aneurysm wall and the matrix metalloproteinase (MMP) activity in similar samples of aortic tissue. METHODS: Detailed mechanical testing was conducted on 124 standardized specimens of aneurysm wall harvested from 24 patients undergoing elective aneurysm repair. The intrasac pressure required to cause aneurysm rupture was calculated based upon the Law of Laplace. In addition, MMP-2 and 9 were assayed from these specimens. Sixty specimens of nonaneurysmal aorta from 6 cadaveric organ donors served as controls. Intrasubject and intersubject variations were analyzed. RESULTS: In the aneurysm specimens, the Young's modulus was 1.80x10(6) N/m(2), the load at break was 6.36 N, the strain at break was 0.30, the ultimate strength was 0.53x10(6) N/ m(2), and the MMP activity was 312 for MMP-2 and 460 for MMP-9. In the controls, the circumferential measurements were a Young's modulus of 1.82x10(6) N/m(2), a load at break of 5.43 N, strain at break of 0.29, ultimate strength of 0.61x10(6) N/m(2), and MMP activity of 395 for MMP-2 and 2019 for MMP-9. Longitudinal measurements in controls were a Young's modulus of 1.38x10(6) N/m(2), a load at break of 11.39 N, a strain at break of 0.33, and ultimate strength of 1.30x10(6) N/m(2). Intra and intersubject variation of all parameters was very high. Based upon the lowest measured tensile strength for each aneurysm, the intrasac pressure required to cause rupture varied from 142 to 982 mmHg. CONCLUSIONS: Localized "hot spots" of MMP hyperactivity could lead to focal weakening of the aneurysm wall and rupture at relatively low levels of intraluminal pressure. These data suggest that tensile strength of the sac is just as important as intrasac tension in determining the risk of rupture. Moreover, these observations may explain why some small aneurysms rupture and larger aneurysms do not. Assessment of rupture risk based on computation or measurement of wall stress may be subject to error and inaccuracy due to variations in wall tensile strength. PMID- 15298502 TI - Relationship of matrix metalloproteinases and macrophages to embolization during endoluminal carotid interventions. AB - PURPOSE: To investigate if relationships exist among macrophage infiltration, plasma matrix metalloproteinase (MMP) levels, and the number of emboli generated during endoluminal carotid interventions. METHODS: Carotid endarterectomy specimens excised as intact cylinders (n=27) were subjected to a standardized angioplasty procedure under radiological guidance in an ex vivo pulsatile flow model. Emboli collected in distal filters were counted and sized using microscopy. Preoperative plasma gelatinase activity was determined by gelatin zymography and quantified using image analysis software. Levels of tissue inhibitors of metalloproteinases (TIMP) 1 and 2 were determined by ELISA. Macrophages within postangioplasty plaques were analyzed using immunohistochemical staining for CD68 antigen and graded by a blinded examiner. Statistical analysis was performed using Spearman's rank correlation. RESULTS: The median number of emboli recorded during angioplasty was 104 (interquartile range 33.75-242.5, absolute range 13-1090). Plasma MMP-9 and MMP-2 levels correlated with emboli number (r=0.544 [p=0.003] and r=0.412, [p=0.033], respectively), while TIMP-1 and TIMP-2 levels did not. Macrophage infiltration within the plaques correlated with emboli number (r=0.722, p<0.001) and the plasma MMP-9 level (r=0.489, p=0.010). CONCLUSIONS: These data indicate that plaque macrophage infiltration may play a role in the generation of emboli during endoluminal carotid intervention, possibly via modulation of protease activity. PMID- 15298503 TI - Durability and validity of a remote, miniaturized pressure sensor in an animal model of abdominal aortic aneurysm. AB - PURPOSE: To investigate whether a remote, miniaturized pressure sensor could maintain calibration and function through organized thrombus over an extended period in a porcine model of abdominal aortic aneurysm (AAA). METHODS: Six adult pigs had an AAA surgically created and excluded. A sensor zeroed to atmospheric pressure was placed within the aneurysm sac and another within the suprarenal aorta of each animal. Pressure measurements were taken at the initial operation and then on a weekly basis over 2 months. The aortic sensors were correlated to an intra-arterial pressure catheter at the initial operation and at the time of sacrifice. Back-table sensor correlation with atmospheric pressure was done at the time of explantation. RESULTS: Three animals died during the follow-up period. Five animals were available for 6-week follow-up, of which 3 survived for the complete 8-week protocol. Two of the surviving animals had an intra-aortic sensor. All 5 aneurysm sac sensors functioned throughout the experimental period. At the time of sacrifice, the sacs contained a large amount of organized thrombus in which the sac sensors were deeply embedded. The 3 aortic sensors also functioned throughout the course of the experiments. The pressures correlated within 5 mmHg to the catheter-based measurements taken at the initial operation and at the time of sacrifice. Comparison to atmospheric pressure revealed no calibration offset in any sensor. CONCLUSIONS: This chronic implantation study demonstrates the durability of a remote, miniaturized pressure sensor within a surgically created aneurysm sac as well as the suprarenal aorta of a porcine AAA model. There was no calibration offset in any of the sensors, and they remained valid at explantation. We believe that this is further evidence of the potential applicability of this sensor for clinical use. PMID- 15298504 TI - Safety of thrombolytic therapy with urokinase or recombinant tissue plasminogen activator for peripheral arterial occlusion: a comprehensive compilation of published work. AB - PURPOSE: To report a comprehensive literature review focused on comparing the risk of complications with urokinase versus recombinant tissue plasminogen activator (rtPA) for thrombolytic treatment of peripheral arterial occlusions. METHODS: The English-language literature between 1985 and 2002 was searched for studies that used tissue-derived urokinase or rtPA in the treatment of peripheral arterial occlusions. Forty-eight studies (22 urokinase, 22 rtPA, and 4 that included both treatments) were identified, encompassing 2226 urokinase-treated patients and 1927 rtPA-treated patients. The safety of each thrombolytic agent was assessed based on the incidence of major hemorrhage, intracerebral hemorrhage, major limb amputation, transfusions, and mortality. RESULTS: The review revealed a wide range of study protocols, patient conditions, ages of occlusions, dosages/delivery methods of lytic agents, and criteria for reporting complications. The incidence of major hemorrhage varied widely, but the overall rate was lower among urokinase-treated patients (6.2%) than for patients treated with rtPA (8.4%, p=0.007). The overall incidence of intracerebral hemorrhage was also significantly lower for urokinase (0.4% versus 1.1% for rtPA, p=0.020). The major amputation rate was similar for both treatments (urokinase 7.9%, rtPA 7.2%), but the mortality rate was significantly lower for urokinase (3.0% versus 5.6% for rtPA, p<0.001). The need for transfusions was less frequent with urokinase (11.1% versus 16.1%, p=0.002). CONCLUSIONS: These results from a large body of published literature suggest that urokinase may be associated with a lower incidence of complications than rtPA in the treatment of peripheral arterial occlusions. PMID- 15298505 TI - Stent repair of bilateral post-traumatic dissections of the internal carotid artery. AB - PURPOSE: To report endovascular repair of bilateral internal carotid arteries (ICAs) in a youth following an accident. CASE REPORT: A 17-year-old boy presented with dissection of both ICAs after a motorcycle accident. The lesions started 2 cm above the bifurcation and continued upward to the siphon, with normal flow into the circle of Willis. Surgical repair was excluded due to the critical clinical condition of the patient and owing to the anatomical extent of the dissection. Carotid Wallstent endoprostheses were deployed bilaterally to repair the dissected segments of the ICAs. After 13 months, the patient is in good health, with complete patency of both arteries. CONCLUSIONS: The good result we experienced in this case indicates that stents may be a safe and effective treatment for traumatic bilateral carotid dissections. However, the long-term durability of these devices in a young patient remains to be determined. PMID- 15298506 TI - Use of the Parodi Anti-Emboli System and transient subclavian steal for cerebral protection during emergent vertebral artery recanalization. AB - PURPOSE: To report the use of the Parodi Anti-Emboli System (PAES) for cerebral protection during emergent vertebral artery recanalization. CASE REPORT: A 56 year-old chimney sweep was referred with recurrent episodes of vertigo and gait ataxia. Left vertebral artery (LVA) flow was barely detectable on duplex Doppler, and brain computed tomography revealed a small infarct in the posterior inferior cerebellar artery territory. Angiography showed subtotal ostial stenosis of the LVA with poor distal flow and possible thrombus. Due to a high risk of distal embolization with percutaneous treatment, anticoagulation was initiated, and the lesion was to be re-evaluated in 2 to 3 weeks. However, 2 days later, the patient developed severe, aggravating headache, gait and left-limb ataxia, horizontal nystagmus, and vomiting. Emergent angiography showed a total ostial LVA occlusion. The PAES was employed to elicit a temporary subclavian steal during percutaneous LVA recanalization, thus protecting the brain from embolization. The ostial LVA was successfully recanalized and stented, with immediate symptom cessation. CONCLUSIONS: The PAES can be successfully applied in the subclavian artery to prevent distal embolization during emergent vertebral artery recanalization. Since a significant proportion of vertebral strokes are embolic, PAES may play a novel role in the treatment of acute cerebellar stroke. PMID- 15298507 TI - Screening carotid arteriography: why it still makes sense. PMID- 15298508 TI - Comparison of a percutaneous separate stent endograft and a conventional thoracic stent-graft for endovascular repair of type B aortic dissection. AB - PURPOSE: To compare the immediate and midterm outcomes of aortic dissection repair with a separate stent endograft (SSE) versus a conventionally constructed thoracic stent-graft. METHODS: The records of 35 patients treated for type B aortic dissection from September 1997 to April 2003 were reviewed. Seventeen patients (12 men; mean age 58.8+/-11.6 years) underwent endovascular repair with a separate stent endograft (SSE), a custom-made device with a reduced profile suitable for percutaneous introduction through a 12-F sheath. Eighteen patients (10 men; mean age 56.1+/-12.8 years) underwent treatment with a conventional custom-made stent-graft. RESULTS: Angiographic success was achieved in 13/17 (76.5%) of the SSE-treated patients and 12/18 (66.7%) for the conventional device group (p=0.521). Clinical success (complete obliteration/thrombosis of the false lumen) was achieved in 12/17 (70.6%) and 11/18 (61.1%), respectively (p=0.555). There were 2 cases of stent-graft movement during deployment and 2 access site complications in the conventional stent-graft group, whereas the SSE patients had no complications. Except for 2 conventional stent-graft patients who were lost to follow-up, all patients are alive at a mean 19.5+/-11.6 months for the SSE group and 34.2+/-21.5 months for the conventional stent-graft patients. CONCLUSIONS: The separate stent endograft can be deployed percutaneously without the need for blood pressure reduction, achieving accurate deployment without migration. In this small clinical experience, patients treated with the SSE had no access site complications and demonstrated midterm results comparable to the conventional stent-graft cohort, suggesting the possible usefulness of this device for the treatment of thoracic aortic dissection. PMID- 15298509 TI - Axillary artery access for interventional procedures. AB - PURPOSE: To evaluate axillary artery access for the interventional treatment of carotid or splanchnic arteries that have angulated takeoff or complex anatomy when larger catheters (up to 9 F) are needed. TECHNIQUE: The axillary artery approach was used to treat the left internal carotid artery (ICA) in 3 patients (2 angulated takeoffs and 1 bovine arch) and a celiac axis aneurysm. An 8-F, 45 cm-long introducer sheath was inserted for the carotid procedures, whereas a 9-F, 90-cm sheath was chosen for the celiac aneurysm. Cerebral protection and stenting were successfully performed in all carotid patients; an 8x40-mm stent-graft was implanted to exclude the celiac artery aneurysm. An 8-F vascular closure device was used in the axillary arteries; hemostasis was immediate, and no hematoma or other complications were recorded in follow-up. CONCLUSIONS: This preliminary experience revisits the axillary approach as an alternative access route for interventional procedures. In association with a vascular closure device, this approach should be considered as a useful and safe option for those interventional procedures in which larger sheaths or catheters are required to cope with difficult arterial anatomies. PMID- 15298510 TI - Thrombus aspiration as a bailout procedure during percutaneous renal angioplasty. AB - PURPOSE: To present a case in which thrombus aspiration, urokinase, and abciximab were used to recanalize a sudden acute thrombotic occlusion of the right renal artery during percutaneous renal angioplasty. CASE REPORT: A 72-year-old man with severe arterial hypertension, impaired renal function, and peripheral artery disease was referred for interventional renal revascularization of a proximal stenosis of the right renal artery. Predilation was unsuccessful, and stent placement was followed by immediate occlusion of the distal renal artery, probably due to dislocation of a mural thrombus. Since intra-arterial administration of urokinase (300,000 IU) was ineffective, thrombus aspiration was performed using the 7-F guiding catheter. After successful removal of the thrombus, abciximab was given intravenously. Control angiograms showed recanalization of the stented segment and patency of the distal renal arteries, an outcome confirmed 8 months later by duplex ultrasound. CONCLUSIONS: As demonstrated in our case, thromboembolic complications can be rapidly and successfully treated on the table by combined measures, such as catheter thrombus extraction and pharmacological strategies. PMID- 15298511 TI - Transradial approach to coil embolization of an intracranial aneurysm. AB - PURPOSE: To report the use of a transradial approach to coil embolization of an intracranial aneurysm in a morbidly obese patient. TECHNICAL NOTE: When the transfemoral approach was inaccessible in a morbidly obese patient with a ruptured intracranial aneurysm, coil embolization was performed via a 6-F sheath placed in the radial artery. Multiple platinum coils were delivered to exclude the 14-mm basilar tip aneurysm. Because heparin was not reversed, the sheath was left in the artery for 24 hours then removed. The radial artery was pulsatile, and blood supply to the hand was good. CONCLUSIONS: The radial artery appears to be a suitable route for access to the intracranial vessels when the femoral artery is not available. PMID- 15298512 TI - Continuous tenecteplase infusion combined with peri/postprocedural platelet glycoprotein IIb/IIIa inhibition in peripheral arterial thrombolysis: initial safety and feasibility experience. AB - PURPOSE: To evaluate a continuous-infusion protocol for peripheral arterial thrombolysis using tenecteplase (TNK), with regard to the technique, dosing, infusion times, and clinical outcomes. METHODS: Between November 1999 and July 2002, 48 patients (30 men; mean age 68.5+/-11.9 years) presented with acute limb ischemia (ALI) owing to iliofemoral arterial thrombosis, which was treated with continuous TNK infusion (either 0.50 mg/h [n=22, group A] or 0.25 mg/h [n=26, group B]). All patients received periprocedural heparin (500 U/h) and peri and postprocedural tirofiban for 6 to 12 hours. Follow-up included ankle-brachial index and duplex ultrasound at baseline, 1 month, and 6 months. The variables retrospectively analyzed included total infusion time, total TNK dose, fibrinogen analysis, clinical and thrombolysis outcomes, and complications. RESULTS: The overall clinical procedural success was 95.8%. Complete (>95%) lysis was observed in 35 (73%) patients; overall mean infusion time was 7.5 hours, and overall mean TNK dose was 4.8 mg. No deaths, intracranial bleeding, or embolic events occurred in either group. Of the 8 (16.7%) complications, 5 (10.4%) were major: 1 femoral repair (group A), 2 >5-cm nonsurgical hematomas (1 in each group), and 2 gastrointestinal hemorrhages (1 in each group). The 3 (6.3%) minor complications were minor hematomas (2 in group A and 1 in group B). The 30-day and 14-month mean limb salvage rates were 95.8% (46/ 48) and 89.6% (43/48), respectively. CONCLUSIONS: Continuous TNK infusion (0.25-0.50 mg/h) is a safe and feasible treatment for continuous pharmacological thrombolysis in ALI, potentially offering decreased infusion times and bleeding complications, as well as improved outcomes. PMID- 15298513 TI - Impact of age, height, and body mass index on arterial diameters in infants and children: a model for predicting femoral artery diameters prior to cardiovascular procedures. AB - PURPOSE: To measure common femoral artery (CFA) diameters in infants and children referred for cardiac catheterization and investigate if CFA diameters can be predicted upon the basis of age, body mass index (BMI), and height. METHODS: CFA diameters were measured in 84 infants and children (50 boys; age range 1- 220 months) referred for diagnostic or therapeutic cardiac interventions. Sonographic measurements were made in a supine position utilizing a 7.5-MHz linear transducer; diameters were defined as the intima to intima distance. Age was described in months and height in centimeters. The Spearman correlation coefficient (rho) was used to test the similarity of diameters between sides; the Pearson correlation coefficient (r) was used to analyze the influence of age, height, and BMI on CFA diameter. RESULTS: Diameters of the right and left CFA were similar (rho=0.951). Age and height were highly correlated (rho=0.956), but not BMI and height (rho=0.279). The best model was CFA diameter = -0.838 + 0.031 height + 0.046 BMI. Height was the most relevant determinant for CFA diameter (p<0.0001, 90% CI 0.027 to 0.036; BMI: p=0.093, 90% CI 0.001 to 0.090, and the intercept: p=0.032, 90% CI-1.475 to-0.200). CONCLUSIONS: Common femoral artery diameter can be sufficiently predicted from height and BMI of infants and children prior to femoral catheterization or surgical reconstruction. PMID- 15298514 TI - Clinical outcome of primary infrainguinal subintimal angioplasty in diabetic patients with critical lower limb ischemia. AB - PURPOSE: To evaluate the clinical outcome of subintimal angioplasty in diabetic patients with critical limb ischemia (CLI) compared to nondiabetics irrespective of the patency status of the treated arteries. METHODS: The records of 99 consecutive patients (53 men; median age 78.5 years, range 42-92) suffering from CLI who underwent primary infrainguinal subintimal angioplasty in 112 limbs within a 6-month period were studied retrospectively. A third of the patients (n=33) were diabetic. The technical success, perioperative morbidity/mortality, and clinical success were compared between the diabetic and nondiabetic patients. Kaplan-Meier life-table analysis was used to analyze clinical success, limb salvage, and survival for both groups. RESULTS: The overall technical success was 89% (81% in diabetics, 93% in nondiabetics, p=0.05). Perioperative morbidity was 8% (16.7% in diabetics, 3.9% in nondiabetics, p=0.03). The perioperative mortality was zero. The clinical success at 12, 24, and 36 months was 74%, 72%, and 65% in nondiabetics and 69%, 63%, and 54% in diabetics, respectively (p=0.17). The limb salvage rate at 36 months was 88% overall (90% in nondiabetics, 82% among diabetics, p=0.20). The 36-month survival rate was 61% in nondiabetics and 57% in diabetics (p=0.29). CONCLUSIONS: In terms of clinical outcome, infrainguinal subintimal angioplasty is almost equally effective in diabetics as in nondiabetics suffering from CLI. PMID- 15298515 TI - Biocompatibility studies of the Anaconda stent-graft and observations of nitinol corrosion resistance. AB - PURPOSE: To validate the deployment, in vivo performance, biostability, and healing capacity of the Anaconda self-expanding endoprosthesis in a canine aortic aneurysm model. METHODS: Aneurysms were surgically created in 12 dogs by sewing a woven polyester patch onto the anterior side of the thoracic or abdominal aorta. Anaconda prostheses were implanted transfemorally for prescheduled periods (1 or 3 months). Aneurysm exclusion and stent-graft patency were monitored angiographically. Healing was assessed with histological analysis and scanning electron microscopy (SEM). Textile analysis determined the physical and chemical stability of the woven polyester material, while the biostability of the nitinol wires was evaluated with SEM and spectroscopy. RESULTS: All prostheses were intact at explantation. After 1 month, endothelial-like cells were migrating in a discontinuous manner both proximally and distally over the internal collagenous pannus at the device-host boundary. After 3 months, endothelialization had reached the midsections of the devices, with a thicker collagenous internal capsule. Patches of endothelial-like cells were sharing the luminal surface with thrombotic deposits. However, the wall of the device at the level of the aneurysm was generally poorly healed, with multiple thrombi scattered irregularly over the luminal surface. The polyester fabric was intact except for some filaments that were ruptured adjacent to the sutures and some abrasion caused by the nitinol wires. No evidence of corrosion was found on the nitinol stents. CONCLUSIONS: This Anaconda stent-graft has demonstrated its ability to exclude arterial aneurysms. The device used in this study was an experimental prototype, and the manufacturer has incorporated new immobilization features into the model for clinical use. The constituent materials appear to be suitable in terms of biocompatibility, biofunctionality, and short-term durability. PMID- 15298516 TI - Stent-graft migration: a reappraisal of analysis methods and proposed revised definition. AB - PURPOSE: To demonstrate the need for a radiographic definition of migration that accurately describes a specific failure mode of an aortic stent-graft. METHODS: The diagnosis of endograft migration, as defined by the Society for Vascular Surgery/American Association for Vascular Surgery (SVS/AAVS) standards, requires a synthesis of clinical and/or radiographic observations. Radiographic studies and clinical reports of 704 consecutive patients treated over a 6-year period with abdominal aortic endografts were retrospectively reviewed. According to the current SVS/AAVS standards, 25 patients were identified as having endograft migration. Follow-up computed tomographic scans and radiographs available for 24 of these patients were scrutinized from discharge to the time of any observed proximal or distal fixation system movement based upon the discharge or 30-day CT scan. Proximal migration was defined with respect to the origin of the superior mesenteric artery (SMA) as movement in a caudal or cranial direction of >/=10 mm or >2 times the reconstructed resolution of the imaging study (whichever measurement was less). Distal migration was similarly defined using the aortic bifurcation and respective hypogastric artery origin as reference points. In an effort to assess the scan-to-scan variation, the distance between the SMA and lowest renal artery, which was expected to remain consistent, was measured. RESULTS: Film analysis with application of the revised migration definition confirmed fixation system failure with respect to the native arterial system in 12 (50%) of the 24 patients. Subjects judged to have endograft migration according to the reporting standards but not to have radiographic evidence of migration based on the modified criteria included 2 proximal endoleaks without evidence of device movement treated with proximal extensions, 1 procedure-related migration, 2 type III endoleaks treated with a second prosthesis implanted within the first, 1 distal endoleak treated with a limb extension, 1 rupture with presumed distal limb migration, and 2 cases of component separation. Three limb extensions were placed in the absence of leak or migration. These 12 patients all had radiographic evidence that the proximal and distal aspects of the originally implanted device did not move with respect to the native arterial vasculature, thus confirming stability of the respective fixation system. CONCLUSIONS: Adherence to this comprehensive definition of device migration, which differs from the published reporting standards, allows differentiation of the specific mechanisms of device failure. Data viewed in this context will aid in the understanding of device strengths and weaknesses, potentially improve patient assessment, and encourage design modifications to address specific aspects relating to fixation failure. PMID- 15298517 TI - Stent-graft migration: how do we know when we have it and what is its significance? PMID- 15298519 TI - Retreat and resilience: life experiences of older women with intellectual disabilities. AB - Older women with intellectual disabilities remain the least studied and understood members of the disability population, and yet they often live well into late adulthood. In this exploratory study we used extensive interviews to examine the demographics, economic and personal safety nets, health, social roles, and well-being of 29 Irish and American older women with intellectual disabilities. Results suggest that these women have very limited resources, social networks, and opportunities. All the women were poor and most lived in group residences, with paid staff as their main allies and careproviders. They reported that their health was good, though it often limited their activities. Despite their societal limitations, these women reported this is the happiest period of their lives. PMID- 15298520 TI - Status of end of life care in organizations providing services for older people with a developmental disability. AB - Volunteers of America conducted a national survey of directors of 500 organizations providing services to older people with an intellectual disability in order to establish a baseline for assessing current status of end of life care. The questionnaire contained 56 questions. The return rate was 32% (N = 160). Data analysis provided an overview of organizational needs, resource allocations, end of life care services currently provided, obstacles to care, methods for monitoring such care, and existing and recommended strategies for improving this care. Results underscored the needs for practical guidelines and resources, increased staffing and training, and policy-level reduction of obstacles to promote improved end of life care. PMID- 15298521 TI - Older adults with intellectual disability in residential care centers in Israel: health status and service utilization. AB - To determine their health status, we studied 2,282 Israeli adults with intellectual disability who were at least 40 years of age and lived in residential care. Results showed that age is a significant factor in health status. The frequency of different disease categories (e.g., cardiovascular disease, cancer, and sensory impairments) increased significantly with age for both genders. Cardiovascular disease in this population was less prevalent when compared to the general population, suggesting that underdiagnosis of some diseases or conditions may be prevalent in this population. The patterns of organ system morbidity with increasing age were similar to those in other studies conducted in several countries, suggesting that health status and outcomes could be independent of cultural factors. PMID- 15298522 TI - Deinstitutionalization for older adults with severe mental retardation: results from Australia. AB - A deinstitutionalization research project in which residents from the largest institution in Queensland, Australia, were relocated after a government decision to close the center was described. Outcomes of relocation into community living for adults with severe mental retardation, many of whom were older (over 40 years) and had been institutionalized for much of their lives, was addressed within the Australian context. Results from the relocation and outcomes for people aged 20 to 39 years (n = 37), 40 to 59 years (n = 39), and over 60 years (n = 24) were described. Group results focused on adaptive and maladaptive behavior, choice-making, and objective life quality were discussed together with the implications for service provision. PMID- 15298523 TI - Older parent caregivers' engagement with the service system. AB - Older parents of adults with intellectual disability are reported to be frequently isolated from the services designed to support their caregiving. The interaction between older parent caregivers' biographies and their involvement with the service system was examined. Parental status was predicted to be an explanatory mechanism for understanding the differential nature of their engagement with services. Analysis of interviews with 64 such caregivers of adults with intellectual disability revealed that their perceptions of their parental status were derived from a complex interaction of their values and beliefs, actions in relation to their adult child, constraints as well as resources available to them, and their relationships with service agencies. Implications for service delivery and directions for research are discussed. PMID- 15298524 TI - Patterns of service utilization by adults with a developmental disability: type of service makes a difference. AB - Patterns and correlates of service utilization by adults with a developmental disability were examined using data from 831 mothers of an adult child with a developmental disability. A modified Andersen model of health services was used to examine service utilization in seven domains. Multinomial logistic regression revealed that predictors of services received as well as predictors of unmet need for services varied by service. Findings emphasize the importance of considering predisposing characteristics, enabling resources, and need as well as service in order to understand patterns of service utilization. PMID- 15298525 TI - Controlled evaluation of support groups for grandparent caregivers of children with developmental disabilities and delays. AB - There have been growing reports of older women and men caring for their grandchildren and great grandchildren. Many of these grandparents are caring for children with developmental disabilities. To systematically examine the effectiveness of a support group intervention for such grandparents, we recruited 97 grandparents through three agencies in New York City and assigned them to treatment and wait list control conditions. Significant reductions in symptoms of depression and increases in sense of empowerment and caregiving mastery were found for the treatment group. Similar effects were found for the control subjects when they later received the intervention. PMID- 15298526 TI - Importance of access to fixed-imaging fluoroscopy: practice implications for the vascular surgeon. AB - PURPOSE: To examine the impact of unfettered access to high quality fixed-imaging fluoroscopy in a vascular surgery practice. METHODS: The case mix of 2 vascular surgeons was retrospectively examined for a 12-month period before (period A) and after (period B) routine access to fixed-imaging equipment was established. Operative and endovascular cases were identified by their CPT codes. Trends in procedure frequency and gross charges were assessed. RESULTS: Endovascular code usage increased 174% (p<0.001) following routine access to fixed imaging equipment. There was an overall 2.3-fold increase in angioplasty across all vascular beds (p<0.001), as well as a 2.1-fold increase in stent utilization (p<0.001). More complex diagnostic and interventional procedures were performed, as evidenced by a large increase in third-order catheterizations (p<0.001). Open surgical therapy decreased overall by 11.4% (p=0.051) in period B. Reductions in open surgery for peripheral arterial occlusive disease were most pronounced, decreasing 35.6% (p<0.001). Overall gross charges increased 6% in group B. Endovascular procedures accounted for 36.6% of gross charges in period B, doubling its contributions from period A (17.1%, p=0.01). Open major vascular case contributions to gross charges fell from 54.4% to 36.2%. CONCLUSIONS: A significant shift in case mix was observed after routine access to fixed imaging equipment was established, with a dramatic increase seen in percutaneous endovascular case volume and complexity. Corresponding contributions to gross charges for endovascular procedures became equivalent to that of all open major vascular cases combined. Routine access to fixed imaging fluoroscopy appeared to be the chronological fulcrum on which the balance of endovascular and open vascular cases has shifted, allowing the development of a fully integrated vascular and endovascular practice. PMID- 15298527 TI - Health plan member experience with point-of-service prescription step therapy. AB - OBJECTIVE: To better understand health plan member experience with point-of service prescription step-therapy edits and outcomes in terms of drug received. METHODS: Self-administered mailed surveys were sent to 1,000 members who experienced a step-therapy edit from September 1, 2002, through January 31, 2003, for proton pump inhibitors or nonsteroidal anti-inflammatory drugs. Based upon these findings, a second survey was conducted by telephone among 617 members who experienced a step-therapy edit from January through April 25, 2003, and who had no subsequent prescription claim for the drug therapy class associated with the edit. RESULTS: The mailed survey generated a 23% response rate, and the telephone survey generated a 33% response rate. Just over 66% of the mail survey respondents indicated that they contacted their physician directly to try to remedy the situation, while 40% indicated that their pharmacist contacted their physician. Forty-four (44%) percent of members indicated that they received a different medication than was originally prescribed, 15% obtained prior authorization for the brand medication, 11% received no medication, 11% paid full price for the branded medication, 8% got an over-the-counter medication, and 4% received samples from their physician. Approximately 7% sought other means of obtaining coverage (i.e., they used spouse.s insurance) or did not remember the outcome. Member and pharmacy contact with the physician significantly influenced whether the member obtained a medication covered by their health plan (odds ratio [OR] = 6.5; 95% confidence interval [95% CI], 2.76-15.12 and OR = 4.6; 95% CI, 1.96-10.60, respectively). In a separate survey conducted by telephone among a different group of members, insight into reasons why members did not obtain any medication was obtained. Using a closed-ended question, 12% (n = 25) of members indicated receiving no medication. Upon further questioning, however, 32% of those who indicated that they had not received a medication said that they had in fact received a medication to treat their condition some time after the step edit. The second most common reason for not receiving a medication included issues related to cost (i.e., willingness to pay) or affordability (16% and 28%, respectively). CONCLUSIONS: The results of this study suggest that a majority of members receive a medication covered by their health plan subsequent to rejection of a claim for a prescribed drug that is the target of a step-therapy edit. However, there are opportunities for better member and provider communication designed to increase the use of first-line drugs and reduce the number of members paying out-of-pocket or receiving no medication. PMID- 15298528 TI - The burden of illness of irritable bowel syndrome: current challenges and hope for the future. AB - OBJECTIVES: To review unmet medical needs associated with irritable bowel syndrome (IBS), to discuss factors that contribute to these unmet needs, and to provide an overview of advancements in IBS diagnosis and treatment options that may influence treatment strategies. SUMMARY: IBS is characterized by a multiple symptom complex of abdominal pain or discomfort and altered bowel habits (i.e., constipation, diarrhea, or both in alternation) and is associated with a large unmet medical need. IBS symptoms are chronic and bothersome, and they have a profound negative impact on patients. quality of life (i.e., affecting sleep, personal relationships, travel, diet, and sexual functioning). IBS imposes a substantial economic burden in direct medical costs and in indirect social costs such as absenteeism from work and school and lost productivity, along with the less-measurable costs of a decreased quality of life. The annual cost of IBS treatment in the United States has been estimated to be between $1.7 billion and $10 billion in direct medical costs (excluding prescription and over-the-counter [OTC] drug costs) and $20 billion for indirect costs. The goals of IBS therapy are to provide global relief of the multiple symptoms of IBS and to relieve single IBS symptoms. Although traditional IBS therapies (e.g., laxatives, antidepressants, antispasmodics, and bulking agents) are useful for some patients in relieving single IBS symptoms, patients generally are dissatisfied with their overall efficacy and tolerability. These agents have not been proven in randomized, controlled clinical trials to be more effective than placebo in providing global relief of the multiple symptoms of IBS. Over the past 2 decades, numerous advancements in the diagnosis and management of IBS have provided hope for the future, including research strides in our understanding of the underlying pathophysiology of IBS; new diagnostic and management recommendations based on a stepwise, symptom-based approach; and the development of novel pharmacologic agents. CONCLUSION: IBS imposes a high socioeconomic burden on its sufferers and on society. Research strides in the underlying pathophysiology of this disorder have enhanced our understanding of IBS, but many questions remain to be answered. Development of evidence-based guidelines on the stepwise, symptom-based approach to IBS diagnosis and the continuing efforts to develop unique pharmacologic classes targeted at the multiple symptoms of this disorder are steps in the right direction. Though cost-effectiveness data on specific pharmacologic classes are not yet available, these ongoing efforts may help promote timely IBS diagnosis and patient satisfaction with care, optimally decreasing the use of health care resources. PMID- 15298529 TI - Allergic rhinitis, asthma, and rhinosinusitis: diseases of the integrated airway. AB - OBJECTIVE: To review data supporting the integrated airway hypothesis. Allergic rhinitis, rhinosinusitis, and asthma are common conditions associated with significant morbidity and health care costs. A theory has been developed suggesting that these conditions may be manifestations of an inflammatory process within a continuous airway rather than fully separate diseases. Based on this theory, the presence of upper airway symptoms may negatively influence the natural course of lower airway disease. Controlling upper airway inflammation and symptoms among asthma patients may help improve health and economic outcomes. SUMMARY: Further clarifying and understanding the relationship between diseases of the upper and lower respiratory tracts is important because of the prevalence of allergic rhinitis, rhinosinusitis, and asthma and the resulting burden on patients and the health care system. Recent progress in understanding the biology of airway disease has identified inflammation as playing a critical and integrating role in these diseases; however, other important questions remain, including factors that determine the clinical phenotype in allergic airway disorders and optimal treatment approaches. CONCLUSIONS: Several recent studies have suggested that allergic rhinitis, rhinosinusitis, and asthma may be manifestations of a common underlying pathology, but there are many unanswered questions. More studies are needed to better define all the underlying pathologic mechanisms as well as treatments to optimize outcomes for patients with allergic rhinitis, rhinosinusitis, and asthma. PMID- 15298530 TI - Advancements in the treatment of psoriasis: role of biologic agents. AB - OBJECTIVE: To evaluate the role of biologic agents as antipsoriatic therapy. SUMMARY: Mild psoriasis can generally be managed with topical therapy. Moderate to-severe psoriasis has traditionally been treated with systemic therapies such as cyclosporine, methotrexate, retinoids, and phototherapy (ultraviolet B, psoralen plus ultraviolet A). The treatments for moderate-to-severe psoriasis often do not meet patient and physician expectations because of significant side effects (e.g., organ toxicity, skin cancer), lack of durable efficacy, and inconvenient administration schedules (e.g., daily dosing, multiple weekly exposures). The recognition of psoriasis as a T-cell.mediated disease has led to the development of biologic agents that more specifically target key steps in the pathologic process. A review of the literature was conducted to identify randomized controlled trials that have been published on the efficacy, safety, and quality-of-life effects of both approved and investigational biologics for the treatment of psoriasis. The first 2 biologic agents for the treatment of moderate-to-severe chronic plaque psoriasis were approved by the U.S. Food and Drug Administration (FDA) in 2003, alefacept in January and efalizumab in October. Both agents have demonstrated favorable safety profiles in clinical trials and significant benefits on patient quality of life. Head-to-head trials are lacking, but in placebo controlled trials, similar percentages of patients appear to respond to each of these 2 drugs. An advantage of alefacept is that it has been shown in clinical trials to provide durable off-treatment efficacy (approximately 7 months). Efalizumab has a relatively quick onset of antipsoriatic effect, but it needs to be administered once weekly continuously to maintain symptom control. Etanercept (approved by the FDA for treating moderate to-severe plaque psoriasis in May 2004) and infliximab (not FDA-approved for psoriasis treatment) have also shown promise in randomized controlled trials, although less data are available on these agents. Case reports and pilot studies suggest that other biologics under investigation may also prove useful for the treatment of psoriasis. Patient populations that may particularly benefit from biologic therapy are discussed. CONCLUSION: Biologic agents appear to offer a safe and effective alternative to conventional systemic therapies and phototherapy for the treatment of moderate-to-severe chronic plaque psoriasis. The biologics appear to be safer than traditional therapies, although long-term safety data still need to be established. PMID- 15298531 TI - A literature review of cardiovascular disease management programs in managed care populations. AB - OBJECTIVES: (1) To review the literature on cardiovascular disease management programs in managed care populations, (2) compare the rigor of the studies and their findings by disease state, and (3) posit directions for future research. SUMMARY: A total of 20 studies conducted in managed care populations were reviewed: 5 in patients with congestive heart failure (CHF), 9 in hypertensive patients, and 6 in hyperlipidemia and/or coronary artery disease (hyperlipidemia- CAD) patients. Management of CHF involved multifaceted programs that included the participation of multiple health care professionals, patient and physician education, promotion of intensive drug therapy and lifestyle modifications, and close patient monitoring. The most common CHF management strategies were case management and physician education, with an emphasis on close patient monitoring. Hypertension and hyperlipidemia-CAD intervention programs focused on chronic outpatient management and regular follow-up, with an emphasis on self-management skills. These programs were managed through regular and periodic interventions, including pharmacist-managed clinics and automated provider notices. Many of the studies employed "before-after" comparisons in the absence of a truly experimental design and posed significant limitations due to variations in the outcomes measured, lack of transparent disease severity stratification, and variation across types of managed care organizations. CONCLUSION: A number of cardiovascular disease management strategies in the literature reported promising results. Many of the multidisciplinary CHF disease management programs were more complex than were programs for hypertension and hyperlipidemia-CAD, due, at least in part, to the nature and severity of the disease. A lack of agreement on appropriate economic and clinical outcomes for evaluating the effectiveness of cardiovascular disease management strategies is readily apparent. PMID- 15298532 TI - Impact on drug costs and utilization of a clinical pharmacist in a multisite primary care medical group. AB - OBJECTIVES: To measure the cost and utilization outcomes of a pharmacist intervention in a primary care medical group operating under a financial risk contract with a health plan. METHODS: A prestudy-poststudy design using national drug utilization for the comparison was employed to assess the impact of physician-prescriber education using information derived from prescriber-specific drug cost and utilization analyses. Eight therapeutic areas involving 10 drug classes were targeted (antihistamines, nasal steroids, proton pump inhibitors, histamine-2 receptor antagonists, antidepressants, nonsteroidal anti-inflammatory drugs [NSAIDs, including celecoxib and rofecoxib], antibiotics, angiotensin converting enzyme [ACE] inhibitors, calcium channel blockers, and statins). The subject medical group was an independent, nonacademic, ambulatory, primary care medical practice of 65 primary care physicians consisting of 50 internists, 14 pediatricians, and 1 family practitioner. Drug costs were measured as net medical group costs per enrolled member per year (PMPY), the product of the average cost per prescription, and the number of prescriptions PMPY. RESULTS: The increase in drug cost PMPY for this medical group was 1.7% in a commercial contract in 1999 compared with an increase of 31.2% for health plans nationwide. Utilization as measured by prescriptions PMPY did not change in the national data comparison over the 2-year observation period and increased by 4.0% in the medical group. The national average cost per prescription claim increased 31.2% in 1999 compared with a decrease of 2.1% in the medical group. The decrease in average drug cost in this medical group was associated with an increase in the use of preferred drugs, including more generic drugs. CONCLUSION: A targeted educational program for physician-prescribers conducted by a clinical pharmacist working for a primary care medical group can reduce the expenditures for outpatient drug therapy by lowering the average cost per pharmacy claim. PMID- 15298533 TI - Clinical pharmacist intervention in a primary care medical group reduces financial losses. PMID- 15298534 TI - Step-therapy edits for PPIs and COX-2 drugs. What we do and do not know. PMID- 15298535 TI - Disease management opportunities for IBS. Placebo versus active therapy. PMID- 15298536 TI - The role of pharmacy benefit managers in formulary design: service providers or fiduciaries? PMID- 15298537 TI - Voluntary physical exercise-induced vascular effects in spontaneously hypertensive rats. AB - Forced training has been shown to have beneficial vascular effects in various animal exercise models. In the present study, we explored possible physiological and molecular effects of voluntary physical exercise on various vascular beds. SHR (spontaneously hypertensive rats) performed voluntary exercise for 5 weeks in a computerized wheel cage facility. Ex vivo myograph studies revealed an increased sensitivity of the ACh (acetylcholine)-mediated vasodilation in resistance arteries of the exercised animals (ED50=15.0+/-3.5 nmol/l) compared with the controls (ED50=37.0+/-8.8 nmol/l; P=0.05). The exercise/control difference was abolished after scavenging reactive oxygen radicals. In conduit arteries, ACh induced a similar vasodilatory response in both groups. The in vivo aortic wall stiffness, assessed by means of Doppler tissue echography, was significantly lower in the exercising animals than in controls. This was demonstrated by significantly increased peak systolic aortic wall velocity (P=0.03) and the velocity time integral (P=0.01) in exercising animals compared with controls. The relative gene expression of eNOS (endothelial nitric oxide synthase) was similar in both groups of animals, whereas Cu/ZnSOD (copper/zinc superoxide dismutase) gene expression was significantly increased (+111%; P=0.0007) in the exercising animal compared with controls. In conclusion, voluntary physical exercise differentially improves vascular function in various vascular beds. Increased vascular compliance and antioxidative capacity may contribute to the atheroprotective effects associated with physical exercise in conduit vessels. PMID- 15298538 TI - Functional biology of the primate fetal adrenal gland: advances in technology provide new insight. AB - 1. The main aim of the present review is to summarize recent experimental data from human and non-human primate models that have identified factors essential for adrenal development and other factors that may determine the regulation of the specific structural organization and function of the adrenal gland. 2. The fetal adrenal cortex has two morphologically distinct zones, with the outer definitive zone being comprised of tightly packed small cells, which appear to be steroidogenically quiescent until late gestation, and the inner fetal zone, which appears to be steroidogenically active throughout gestation. 3. In the primate fetus, growth of the adrenal gland involves hyperplasia, hypertrophy, migration and senescence. Cells appear to proliferate in the external portion of the definitive zone and then move centripetally and become non-proliferative in the fetal zone, where they acquire their steroidogenic capacities. 4. A variety of new technologies has been used to identify zonal-specific markers of the cortical zones within the developing human fetal adrenal gland. On microarray, 67 transcripts showed a minimum of a 2.5-fold difference between the fetal and adult adrenal gland. The vast majority of these genes had not been studied in relation to adrenal gland development or function. In combination with techniques such as laser capture microdissection, which has allowed the isolation of fairly pure zone-specific cell populations from the human fetal adrenal cortex, we can begin to unravel the complex interactions regulating adrenal growth and functional differentiation. PMID- 15298539 TI - Skeletal muscle function: role of ionic changes in fatigue, damage and disease. AB - 1. Repeated activity of skeletal muscle causes a variety of changes in its properties: muscles become weaker with intense use (fatigue), may feel sore and weak after repeated contractions involving stretch and can degenerate in some disease conditions. The present review considers the role of early ionic changes in the development of each of these conditions. 2. Single fibre preparations of mouse muscle were used to measure ionic changes following activity induced changes in function. Single fibres were dissected with intact tendons and stimulated to produce force. Fluorescent indicators were microinjected into the fibres to allow simultaneous ionic measurements with determination of mechanical performance. 3. One theory to explain muscle fatigue is that fatigue is caused by the accumulation of lactic acid, producing an intracellular acidosis that inhibits the myofibrillar proteins. In contrast, we found that during repeated tetani there was little or no pH change, but that failure of calcium release was a major contributor to fatigue. Currently, it is proposed that precipitation of calcium and phosphate in the sarcoplasmic reticulum contributes to the failure of calcium release. 4. Muscles can be used to shorten and produce force or they can be used to de-accelerate loads (stretched or eccentric contractions). One day after intense exercise involving stretched contractions, muscles are weak, sore and tender, and this damage can take a week to recover. In this condition, sarcomeres are disorganized and there are increases in resting intracellular Ca2+ and Na+. Recently, we demonstrated that the elevation of Na+ occurs through a stretch-activated channel that can be blocked by either gadolinium or streptomycin. Preventing the increase in [Na+]i with gadolinium also prevented part of the muscle weakness after stretched contractions. 5. Duchenne muscular dystrophy is a lethal degenerative disease of muscles in which the protein dystrophin is absent. Dystrophic muscles are more susceptible to stretch-induced muscle damage and the stretch-activated channel seems to be one pathway for the increases in intracellular Ca2+ and Na+ that are a feature of this disease. We have shown recently that blockers of the stretch-activated channel can minimize some of the short-term damage in muscles from the mdx mouse, which also lacks dystrophin. Currently, we are testing whether blockers of the stretch-activated channels given systemically to mdx mice can protect against some features of the disease. PMID- 15298540 TI - Effect of endothelin-1 on regional kidney blood flow and renal arteriole calibre in rabbits. AB - 1. Medullary blood flow (MBF) is important in the long-term control of arterial pressure. However, it is unclear which vascular elements regulate MBF. 2. Exogenous endothelin (ET)-1 decreases cortical more than medullary blood flow. We hypothesized that ET-1 would therefore constrict afferent (AA) and efferent arterioles (EA) of juxtamedullary glomeruli less than those of cortical glomeruli. 3. Mean arterial pressure, renal blood flow and cortical (CBF) and medullary (MBF) blood flow, via laser-Doppler flowmetry, were measured before and after intrarenal ET-1 (2 ng/kg per min; n = 6) or vehicle (n = 6) in anaesthetized rabbits. Kidneys were perfusion fixed, vascular casts formed, lumen diameters measured via scanning electron microscopy and relative resistance calculated. 4. Mean arterial pressure was not significantly affected by ET-1 infusion. Cortical glomerular arteriole lumen diameters were significantly reduced in the ET-1-infused group (AA approximately 30%, EA approximately 18%; PA < 0.01), compatible with the decrease in CBF (42 +/- 3%; PGT < 0.01). Juxtamedullary arteriole lumen diameters were also significantly reduced in the ET-1-infused group (AA approximately 34%, EA approximately 21%; PA < 0.01); however, MBF did not decrease. 5. In conclusion, our data suggest that juxtamedullary arterioles are not of primary importance in the regulation of MBF because, despite reductions in juxtamedullary arteriole diameters in response to ET-1, MBF was not decreased. PMID- 15298541 TI - Arterial structural changes in hypertensive rats induced by capsaicin and salt loading. AB - 1. The objective of the present study was to investigate the arterial structural changes in a salt-sensitive hypertensive rat model induced by treatment with capsaicin. 2. Newborn male Wistar rats were treated with 50 mg/kg capsaicin subcutaneously for 2 days. Control rats were treated with vehicle solution (5% ethanol and 5% Tween 80 in saline). After weaning at 3 weeks, rats were divided into four groups: (i) control with a normal salt diet (0.5% NaCl; CON + NS); (ii) control with a high-salt diet (4% NaCl; CON + HS); (iii) capsaicin plus normal salt diet (CAP + NS); and (iv) capsaicin plus a high-salt diet (CAP + HS). Treatment with different salt diets was initiated at 3 weeks of age and lasted for 18 weeks. Tail-cuff systolic blood pressure (BP) and bodyweight were examined. At the end of the treatment period, blood vessels were prepared by perfusion fixation. Heart weight and vascular dimensions were measured in the thoracic (artery) aorta, renal artery and mesenteric artery. 3. Two weeks after the initiation of the salt diet treatment, BP became significantly higher in the CAP + HS group than in any of the other groups and this difference was maintained until the end of the treatment period. 4. Beginning at 8 weeks after the initiation of the salt diet treatment (11 weeks of age), BP became higher in the CON + HS group than in the CON + NS and CAP + NS groups. Blood pressure was not significantly different between the CON + NS and CAP + NS groups. 5. Media thickness, media thickness to lumen ratio and cross-sectional area of the aorta, renal artery and mesenteric artery were significantly increased in the CAP + HS group compared with the other groups. Heart weight was also increased in the CAP + HS and CON + HS groups compared with the other groups. 6. Similar structural changes in the blood vessels and heart were also found in the CON + HS group compared with the CON + NS group. Lumen diameter was not altered by the treatments in any of the arteries studied. 7. We conclude that treatment with capsaicin increased the sensitivity of the BP of these rats to salt and this increase in BP is correlated with hypertrophy of the arteries (vascular remodelling) with no change in lumen size. A long-term high-sodium load induced hypertension in normal Wistar rats, which was accompanied by cardiovascular hypertrophy. PMID- 15298542 TI - Increase of anti-oxidation by exercise in the liver of obese Zucker rats. AB - 1. The effects of endurance training on the anti-oxidant status in diabetes were studied using obese Zucker rats. 2. We used a moderate exercise programme consisting of treadmill running at 20 m/min and 0% incline for 1 h/day, 7 days/week, for 8 weeks. At the end of the experimental period, changes in hepatic anti-oxidant enzymes in terms of protein content and mRNA levels were detected using western blotting analysis and northern blotting analysis, respectively. In addition, anti-oxidant enzyme activity was determined. 2. A significant reduction in mRNA levels and the protein content of hepatic Mn-superoxide dismutase (SOD) and glutathione peroxidase (GPx) were observed in non-exercise obese groups, but the mRNA and protein levels of these enzymes were markedly increased after exercise training. In addition, exercise training reversed the decreased enzyme activities of Mn-SOD and GPx in obese Zucker rats. 3. The diabetes-related lowering of the glutathione (GSH) concentration was elevated in exercised obese Zucker rats, indicating a marked effect of regular moderate exercise on the endogenous anti-oxidant system. 4. There were no marked changes in hepatic Cu/Zn SOD in terms of mRNA levels, protein content and activity in sedentary obese Zucker rats compared with their lean littermates. Endurance training did not modify the gene expression and activity of hepatic Cu/Zn-SOD. 5. The results of the present study suggest that regular moderate exercise could improve the anti oxidant defence function of Mn-SOD, GPx and GSH in obese Zucker rats. PMID- 15298543 TI - Transitory reduction in angiotensin AT2 receptor expression levels in postinfarct remodelling in rat myocardium. AB - 1. Myocardial infarction (MI) poses a significant risk for sudden cardiac death. The effectiveness of angiotensin-converting enzyme (ACE) inhibitors and AT1 receptor blockade in attenuating unfavourable post-MI outcomes indicates an important role for angiotensin (Ang) II signalling in the post-MI remodelling process. 2. AT1 and AT2 receptor expression is known to be altered during the early postinjury period and at the later failure stage in the infarcted heart. The aim of the present investigation was to characterize AngII receptor expression shifts in the intermediate, adaptive phases of post-MI hypertrophic remodelling. 3. The present study investigated relative cardiac AT1 and AT2 receptor expression levels using semiquantitative reverse transcription polymerase chain reaction (GAPDH normalized) in rats at 4 and 20 weeks after ligation of the left anterior descending coronary artery. 4. Heart weight and normalized heart weight were significantly higher in the MI group than in the sham group 4 weeks post-MI, with significant hypertrophy of the left ventricle, left atrium and right ventricle in MI rats. At 20 weeks post-MI, left ventricular hypertrophy remained significant, whereas the mass of the other cardiac tissues was not different to that of sham controls. 5. AT2 receptor expression was markedly reduced in both the non-infarct and infarcted areas of the left ventricular wall in the MI group compared with the sham-operated group 4 weeks after surgery. Expression levels were reduced to 8 and 13% of sham values in the viable and scar tissue regions, respectively. By 20 weeks post-MI, there was no evidence of AT2 receptor expression suppression in the left ventricle. No significant relative changes in AT1 receptor mRNA levels were observed at either 4 or 20 weeks post-MI. 6. The present study demonstrates, for the first time, a selective downregulation of left ventricular AT2 receptor expression in the intermediate phase of post-MI ventricular remodelling in the rat. This downregulation may provide an enhanced AT1 receptor-mediated compensatory progrowth signal in the early adaptive post-MI growth phase. A more detailed understanding of the time-course of differential AT1 and AT2 receptor expression regulation post-MI may potentially identify an optimal window for targeted pharmacological intervention in the treatment of MI. PMID- 15298544 TI - Central effect of histamine and peripheral effect of histidine on the formalin induced pain response in mice. AB - 1. The present study was designed to investigate the role of brain histamine in modulating pain transmission in mice. 2. In conscious mice implanted with an intracerebroventricular (i.c.v.) cannula, the effects of i.v.c. injections of normal saline (control) and low and high doses histamine (2 and 40 microg/mouse, respectively) were investigated on the duration of paw licking and biting induced by subcutaneous (s.c.) injection of formalin (20 microL; 5%) into the plantar surface of the left hindpaw. 3. To clarify the involvement of histidine in the pain response, the effects of intraperitoneal (i.p.) injections of low and high doses of histidine (50 and 1000 mg/kg, respectively) alone or before i.c.v. injection of histamine were also examined. 4. Intraplantar injection of formalin induced a biphasic pain response (first phase: 0-5 min after injection; second phase: 20-40 min after injection). 5. Histamine (2 microg/mouse, i.c.v.) had no effect on the first phase of the pain response, but suppressed the second phase. The higher dose of histamine (40 microg/mouse, i.c.v.) suppressed both phases of the pain response. 6. Histidine, at 50 mg/kg, i.p., had no effect on the pain response, but the higher dose (1000 mg/kg, i.p.) suppressed the both phases of the pain response. 7. Pretreatment with the low dose of histidine (50 mg/kg, i.p.) prior to administration of 2 microg/mouse, i.c.v., histamine did not change the antinociception induced by low-dose histamine. However, pretreatment with the high dose of histidine (1000 mg/kg, i.p.) prior to 2 microg/mouse, i.c.v., histamine produced antinociception that resembled that seen following administration of the high dose of either histidine or histamine. Pretreatment with the low dose of histidine (50 mg/kg, i.p.) prior to administration of 40 microg/mouse, i.c.v., histamine has no effect on the pain response following high dose histamine. Pretreatment with 1000 mg/kg, i.p., histidine prior to administration of 40 microg/mouse, i.c.v., histamine strongly suppressed both phases of the formalin-induced pain response, particularly the second phase. 8. The results of the present study indicate that: (i) activation of brain histamine produces antinociception in the mouse formalin test; (ii) peripheral loading with a high dose of histidine (1000 mg/kg, i.p.) alone exerts the same effect as that seen following 40 microg/mouse, i.c.v., histamine; and (iii) pretreatment with a high dose of histidine potentiates central histamine-induced antinociception. PMID- 15298545 TI - Protective effect of U74500A on phorbol myristate acetate-induced acute lung injury. AB - 1. The present study was designed to determine whether U74500A could ameliorate acute lung injury (ALI) induced by phorbol myristate acetate (PMA) in our rat isolated lung model compared with any amelioration induced by dimethylthiourea (DMTU), superoxide dismutase (SOD) and catalase. 2. Acute lung injury was induced successfully by PMA during 60 min of observation. At 2 microg/kg, PMA elicited a significant increase in microvascular permeability (measured using the capillary filtration coefficient Kfc), lung weight gain, the lung weight/bodyweight ratio, pulmonary arterial pressure and protein concentration of the bronchoalveolar lavage fluid. 3. Pretreatment with 1.5 mg/kg U74500A significantly attenuated ALI; there was no significant increase in any parameters measured, except for pulmonary arterial pressure. The protective effect of U74500A was approximately the same as that of 600 mg/kg DMTU. However, 6000 U/kg SOD, 50,000 U/kg catalase and 6000 U/kg SOD + 50,000 U/kg catalase had no protective effect. 4. These experimental data suggest that U74500A significantly ameliorates ALI induced by PMA in rats. PMID- 15298546 TI - (-)-Epigallocatechin gallate attenuates glutamate-induced cytotoxicity via intracellular Ca modulation in PC12 cells. AB - 1. The effects of (-)-epigallocatechin gallate (EGCG), a green tea polyphenol, on glutamate-induced increases in intracellular Ca2+ concentrations ([Ca2+]i) and cytotoxicity in PC12 cells were investigated. 2. Changes in [Ca2+]i were measured using Fura-2/AM calcium indicator dye and cellular viabilities were determined by a viable cell count and a 3-(4,4-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide reduction assay. 3. Glutamate increased [Ca2+]i in PC12 cells in a dose dependent manner. (-)-Epigallocatechin gallate attenuated this glutamate (30 mmol/L)-induced [Ca2+]i increase and EGCG (50 micromol/L) increased the viability of PC12 cells against glutamate-induced cytotoxicity. The EGCG effect was also found to be independent of its general anti-oxidant mechanism. In contrast, EGCG directly suppressed both N-methyl-D-aspartate (50 mmol/L)- and kainate (20 mmol/L)-mediated Ca2+ influx, but not metabotropic receptor-mediated Ca2+ release. 4. These results suggest that EGCG reduces the glutamate-induced [Ca2+]i increase by attenuating ionotropic Ca2+ influx and that this promotes the viability of PC12 cells. PMID- 15298547 TI - Identification of a zebrafish model of muscular dystrophy. AB - 1. Large-scale mutagenic screens of the zebrafish genome have identified a number of different classes of mutations that disrupt skeletal muscle formation. Of particular interest and relevance to human health is a class of recessive lethal mutations in which muscle differentiation occurs normally, but is followed by tissue-specific degeneration reminiscent of human muscular dystrophies. 2. We have shown that one member of this class of mutations, sapje (sap), results from mutations within the zebrafish orthologue of the human Duchenne muscular dystrophy (DMD) gene. Mutations in this locus cause Duchenne or Becker muscular dystrophies in human patients and are thought to result in a dystrophic pathology by disrupting the link between the actin cytoskeleton and the extracellular matrix in skeletal muscle cells. 3. We have found that the progressive muscle degeneration phenotype of sapje-mutant zebrafish embryos is caused by the failure of somitic muscle attachments at the embryonic myotendinous junction (MTJ). 4. Although a role for dystrophin at the MTJ has been postulated previously and MTJ structural abnormalities have been identified in the dystrophin-deficient mdx mouse model, in vivo evidence of pathology based on muscle attachment failure is thus far lacking. Therefore, the sapjre mutation may provide a model for a novel pathological mechanism of Duchenne muscular dystrophy and other muscle diseases. In the present review, we discuss this finding in light of previously postulated models of dystrophin function. PMID- 15298548 TI - Popping sarcomere hypothesis explains stretch-induced muscle damage. AB - 1. Exercise that involves stretching a muscle while active cause microscopic areas of damage, delayed onset muscle soreness and adaptation to withstand subsequent similar exercise. 2. Longer muscle lengths are associated with greater damage and recent animal experiments show that it is the length relative to optimum that determines the damage. 3. In humans, walking down stairs, taking two at a time, increases the length of the muscle during the lengthening and increases the delayed onset muscle soreness. 4. The observed pattern of damage is consistent with explanations based on sarcomere length instabilities. 5. The pattern of adaptation is consistent with the number of sarcomeres in series in a muscle being modulated by exercise, especially the range of muscle lengths over which eccentric exercise regularly occurs. PMID- 15298549 TI - Identifying athletes at risk of hamstring strains and how to protect them. AB - 1. One common soft-tissue injury in sports involving sprinting and kicking a ball is the hamstring strain. Strain injuries often occur while the contracting muscle is lengthened, an eccentric contraction. We have proposed that the microscopic damage to muscle fibres that routinely occurs after a period of unaccustomed eccentric exercise can lead to a more severe strain injury. 2. An indicator of susceptibility for the damage from eccentric exercise is the optimum angle for torque. When this is at a short muscle length, the muscle is more prone to eccentric damage. It is known that subjects most at risk of a hamstring strain have a previous history of hamstring strains. By means of isokinetic dynamometry, we have measured the optimum angle for torque for nine athletes with a history of unilateral hamstring strains. We also measured optimum angles for 18 athletes with no previous history of strain injuries. It was found that mean optimum angle in the previously injured muscles was at a significantly shorter length than for the uninjured muscles of the other leg and for muscles of both legs in the uninjured group. This result suggests that previously injured muscles are more prone to eccentric damage and, therefore, according to our hypothesis, more prone to strain injuries than uninjured muscles. 3. After a period of unaccustomed eccentric exercise, if the exercise is repeated 1 week later, there is much less evidence of damage because the muscle has undergone an adaptation process that protects it against further damage. We propose that for athletes considered at risk of a hamstring strain, as indicated by the optimum angle for torque, a regular programme of mild eccentric exercise should be undertaken. This approach seems to work because evidence from a group of athletes who have implemented such a programme shows a significant reduction in the incidence of hamstring strains. PMID- 15298550 TI - Stretch-activated channels in stretch-induced muscle damage: role in muscular dystrophy. AB - 1. Stretch-induced muscle injury results in the damage that causes reduced force and increased membrane permeability. This muscle damage is caused, in part, by ionic entry through stretch-activated channels and blocking these channels with Gd3+ or streptomycin reduces the force deficit associated with damage. 2. Dystrophin-deficient muscles are more susceptible to stretch-induced muscle injury and the recovery from injury can be incomplete. We have found that Na+ entry associated with stretch-induced injury is enhanced in dystrophin-deficient muscles and that blockers of stretch-activated channels are capable of preventing ionic entry and reducing muscle damage. 3. A model is presented that proposes links between stretch-induced injury, opening of stretch-activated channels, increased levels of intracellular ions and various forms of muscle damage. Although changes in Na+ accompany stretch-induced muscle injury, we believe that changes in Ca2+ probably have a more central role in the damage process. PMID- 15298551 TI - Role of contraction-induced injury in the mechanisms of muscle damage in muscular dystrophy. AB - 1. Duchenne muscular dystrophy (DMD) is a severe disease of skeletal muscle, characterized by an X-linked recessive inheritance and a lack of dystrophin in muscle fibres. It is associated with progressive and severe wasting and weakness of nearly all muscles and premature death by cardiorespiratory failure. 2. Studies investigating the susceptibility of dystrophic skeletal muscles to contraction-mediated damage, especially after lengthening actions where activated muscles are stretched forcibly, have concluded that dystrophin may confer protection to muscle fibres by providing a mechanical link between the contractile apparatus and the plasma membrane. In the absence of dystrophin, there is disruption to normal force transmission and greater stress placed upon myofibrillar and membrane proteins, leading to muscle damage. 3. Contraction protocols (involving activation and stretch of isolated muscles or muscle fibres) have been developed to assess the relative susceptibility of dystrophic (and otherwise healthy) muscles to contraction-induced injury. These protocols have been used successfully to determine the relative efficacy of different (gene, cell or pharmacological) interventions designed to ameliorate or cure the dystrophic pathology. More research is needed to develop specific 'contraction assays' that will assist in the evaluation of the clinical significance of different therapeutic strategies for muscular dystrophy. PMID- 15298553 TI - Unravelling synergistic immune interactions between respiratory virus infections and allergic airway inflammation. PMID- 15298554 TI - Cellular and molecular mechanisms in chronic obstructive pulmonary disease: an overview. AB - In the last decade, the analysis of bronchial biopsies and lung parenchyma obtained from chronic obstructive pulmonary disease (COPD) patients compared with those from smokers with normal lung function and non-smokers has provided new insights on the role of the different inflammatory and structural cells, their signalling pathways and mediators, contributing to a better knowledge of the pathogenesis of COPD. This review summarizes and discusses the lung pathology of COPD patients with emphasis on inflammatory cell phenotypes that predominate in different clinical conditions. In bronchial biopsies, a cascade of events takes place during progression from mild-to-severe disease. T lymphocytes, particularly CD8+ cells and macrophages are the prevalent inflammatory cells in the lung of healthy smokers and patients with mild COPD, while total and activated neutrophils predominate in severe COPD. The number of CD4+, CD8+ cells and macrophages expressing nuclear factor-kappa B (NF-kappaB), STAT-4 and IFN-gamma proteins as well as endothelial adhesion molecule-1 in endothelium is increased in mild/moderate disease. In contrast, activated neutrophils (MPO+ cells) and increased nitrotyrosine immunoreactivity develops in severe COPD. In bronchial biopsies obtained during COPD exacerbations, some studies have shown an increased T cell and granulocyte infiltration. Regular treatment with high doses of inhaled glucocorticoids does not significantly change the number of inflammatory cells in bronchial biopsies from patients with moderate COPD. The profile in lung parenchyma is similar to bronchial biopsies. 'Healthy' smokers and mild/moderate diseased patients show increased T lymphocyte infiltration in the peripheral airways. Pulmonary emphysema is associated with a general increase of inflammatory cells in the alveolar septa. The molecular mechanisms driving the lymphocyte and neutrophilic prevalence in mild and severe disease, respectively, needs to be extensively studied. Up-regulation of pro-inflammatory transcription factors NF-kappaB and STAT-4 in mild, activated epithelial and endothelial cells in the more severe disease may contribute to this differential prevalence of infiltrating cells. PMID- 15298555 TI - Monitoring response to treatment in asthma management: food for thought. AB - Asthma is a chronic inflammatory disorder of the airways that is characterized by episodic symptoms. In this regard, asthma management has classically involved periodic re-assessment by the health-care provider, during which therapy is altered mainly based on clinical and physiological parameters, such as assessment of symptoms, spirometry and peak expiratory flow monitoring. In this context, various markers of airway inflammation (e.g. eosinophils in the induced sputum, nitric oxide in the exhaled air) have been proposed to assess the severity of asthma and to adjust the therapy accordingly. The evaluation of airway hyper responsiveness with different stimuli has also been suggested as a new tool to monitor asthma. However, the lack of definite relationships between airway inflammation and asthmatic symptoms strongly limit the use of markers of asthma severity in the clinical setting. Therefore, the need of new tools to assess the severity of asthma is raised. The ideal measurement employed to establish the proper asthmatic therapy should be safe, non-invasive, easy to perform, reproducible and accurate, and have the capability to monitor the changes induced by the therapeutic interventions. A careful review of the available techniques, and the evaluation of their sensitivity and specificity in the clinical setting is warranted. PMID- 15298556 TI - Farming exposure in childhood, exposure to markers of infections and the development of atopy in rural subjects. AB - BACKGROUND: Within the context of the hygiene hypothesis, we aimed to study the potential association between farming-related risk factors and Toxoplasma gondii (T. gondii) as well as Helicobacter pylori (H. pylori) seropositivity. METHODS: The study included questionnaire data and serum samples of 321 young adults living in a rural environment. Serum samples were analysed for specific IgE to a common panel of aeroallergens (SX1) as well as IgG against T. gondii and H. pylori. RESULTS: Regular contact with animal stables before the age of 3 years (odds ratio (OR) (95% confidence interval): 2.0 [1.0; 4.0]) and unpasteurized milk consumption at age 6 years (1.8 [1.0; 3.3]) were the strongest risk factors for T. gondii infection. None of the farming-related factors were significantly associated with H. pylori infection. Current consumption of raw farm milk was not significantly associated with H. pylori infection (2.1 [0.8; 5.3]). Regular contact with animal houses before the age of 7 years was the strongest predictor for atopy (0.49 [0.26-0.96]). The reduction in risk could not be further decreased by any other factor under consideration. After adjustment for animal house contact, the OR for atopy was decreased by raw milk consumption and H. pylori infection in an additive manner. CONCLUSION: Exposure to farming environments in childhood might predict T. gondii seropositivity in rural subjects. Nevertheless, the strongest predictor for atopy in rural subjects seems to be regular contact with farm animals. Whether T. gondii infection is an intermediate factor in the association between farm contact and atopy needs to be confirmed in larger studies. PMID- 15298557 TI - Atopic disease and its determinants -- a focus on the potential role of childhood infection. AB - BACKGROUND: Atopic diseases develop on a genetic background and are modulated by environmental factors among which some infectious diseases are thought to have a protective influence. OBJECTIVE: The aim of this study was to determine the influence of infectious diseases in younger ages, bacterial and viral, on atopic diseases and sensitization to aero- and food-allergens in adults. METHODS: A population-based sample of 4262 subjects aged 25-74 years were interviewed concerning their history of infectious disease within the first 18 years of life. Information about allergic disease, including atopic eczema, allergic rhinitis (AR), and asthma was obtained. A blood sample was drawn and analysed for allergen specific IgE antibodies against food- and aero-allergens. RESULTS: Multiple logistic regression analyses identified viral infection to be associated with AR (adjusted odds ratio (OR) = 1.39; 95% confidence interval (95% CI): 1.13-1.72) and sensitization to aeroallergens (OR = 1.21; 95% CI: 1.05-1.41). Bacterial disease was a negative predictor for atopy development in the subgroup of patients sensitized to nutritional allergens with concomitant atopic eczema (OR = 0.34; 95% CI: 0.11-0.99), AR (OR = 0.67; 95% CI: 0.42-1.07), or asthma (OR = 0.41; 95% CI: 0.19-0.87). Influences of viral and bacterial infection on AR differed with regard to family history of atopic disease. CONCLUSION: In our study population, history of viral infection was consistently positively associated with AR. Our data suggests that bacterial infections might be preventive for specific subgroups of atopy. PMID- 15298558 TI - Polymorphisms in ADAM33 are associated with allergic rhinitis due to Japanese cedar pollen. AB - BACKGROUND: A recent report provided evidence that a disintegrin and metalloprotease domain 33 (ADAM33), a member of the ADAM family, is a novel susceptibility gene in asthma linked to bronchial hyper-responsiveness. However, there has been no investigation of the genetic role of ADAM33 variants in nasal allergy. OBJECTIVE: The purpose of this study was to test the association between ADAM33 polymorphisms and Japanese cedar pollinosis (JCPsis), a most common seasonal allergic rhinitis in Japan. METHODS: We conducted a case-control association study among a Japanese population, involving 95 adult individuals with JCPsis and 95 normal healthy controls. A total of 22 single-nucleotide polymorphisms (SNPs) in ADAM33 were genotyped using PCR-based molecular methods. RESULTS: Six SNPs of ADAM33 gene, three in introns (7575G/A, 9073G/A and 12540C/T) and three in the coding region (10918G/C, 12433T/C and 12462C/T), were strongly associated with JCPsis (P = 0.0002-0.022 for absolute allele frequencies) and most of the SNPs were in linkage disequilibrium with each other. A higher frequency of the common alleles of these SNPs was noted for the subjects with JCPsis in comparison with healthy controls. We also identified a haplotype associated with the disease susceptibility. In addition, associations were found between ADAM33 polymorphisms and various cedar pollinosis phenotypes including clinical severity, eosinophil counts in nasal secretion and allergen-specific IgE levels in sera, but not total serum IgE levels. CONCLUSION: These results indicate that polymorphisms in the ADAM33 gene are associated with susceptibility to allergic rhinitis due to Japanese cedar pollen, but the functional relationship still needs clarification. PMID- 15298559 TI - Gene-gene interaction between interleukin-4 and interleukin-4 receptor alpha in Korean children with asthma. AB - BACKGROUND: Interleukin-4 receptor alpha (IL-4Ralpha), which binds IL-4 and IL 13, is involved in signal transduction of those cytokines that lead to IgE production, and is also a key functional component of the Th2 lymphocyte phenotype. OBJECTIVE: To determine whether IL-4 and IL-4Ralpha polymorphisms are associated with susceptibility to asthma and whether there are gene-gene interactions between IL-4 and IL-4Ralpha polymorphisms. METHODS: We genotyped three groups of Korean children, consisting of 196 atopic asthmatics, 60 non atopic asthmatics, and 100 healthy children, for an IL-4 promoter polymorphism (C 590T) and three IL-4Ralpha polymorphisms (Ile50Val, Pro478Ser, and Arg551Gln) using PCR-RFLP (restriction fragment length polymorphism) assays. RESULTS: The allele frequencies of the IL-4 (C/T) polymorphism and the Ile50Val and Pro478Ser polymorphisms of IL-4Ralpha did not differ statistically among the three groups of children. For the Arg551Gln polymorphism, the combined genotype frequency of the Arg/Gln heterozygote and the Arg/Arg homozygote was significantly higher in atopic asthmatics (27.6%) than in healthy children (16.0%) (odds ratio (OR) = 1.97, 95% CI (confidence interval) = 1.07-3.71). The eosinophil fraction (%) and bronchial responsiveness were higher in children with the Arg/Gln and Arg/Arg genotype than in those with the Gln/Gln genotype (P = 0.036 and 0.024, respectively). In asthmatic children, combinations of the IL-4 CT/TT genotype and the IL-4Ralpha Arg/Gln and Arg/Arg genotypes were associated with significantly increased risk for development of asthma (OR = 3.70, 95% CI = 1.07-12.78, P = 0.038). CONCLUSIONS: In Korean children, the IL-4Ralpha Arg551 allele may play a role in susceptibility to atopic asthma and correlate with markers of asthma pathogenesis, including increased eosinophil fraction and enhanced bronchial hyper-responsiveness. In addition, a significant gene-gene interaction between the IL-4-590C and the IL-4Ralpha Arg551 allele significantly increases an individual's susceptibility to asthma. PMID- 15298560 TI - Induction of T 'regulatory' cells by standardized house dust mite immunotherapy: an increase in CD4+ CD25+ interleukin-10+ T cells expressing peripheral tissue trafficking markers. AB - BACKGROUND: Clinically effective subcutaneous allergen-specific immunotherapy (SIT) is associated with altered circulating T cell cytokine production and altered local cytokine responses with increased IL-10 following allergen challenge in target organs. OBJECTIVE: This study aimed to elucidate mechanisms for these T cell changes, by examining surface expression of markers for peripheral tissue trafficking on circulating cytokine-positive T cells following standardized house dust mite- (HDM-) SIT. METHODS: A randomized conventional HDM immunotherapy study was performed on a panel of 12 HDM-allergic subjects. Nine subjects received treatment with conventional HDM immunotherapy using a standardized extract and three subjects were treated by standard pharmacotherapy alone. Symptom and medication scores and allergen-induced cutaneous late-phase responses were assessed before and 9 months after institution of therapy. Before and at 3 and 9 months of SIT, peripheral blood mononuclear cells were cultured for 14 days with HDM extract and CD4+ and CD8+ T cell expression of CD62L, CD49d and CCR5 and production of IL-10, IFN-gamma and IL-4 were analysed by flow cytometry. Allergen-specific T cell proliferation was assessed by 3H-thymidine incorporation. RESULTS: At 9 months, all SIT-treated patients showed reduced symptom scores and late-phase cutaneous responses to HDM compared with baseline levels. The proportions of CD4+ T cells which were IL-10+ were increased (P < 0.01), and the proportions of CD4+ and CD8+ T cells which were IL-4+ decreased (P < 0.05) compared with baseline. CD4+ and CD8+ T cell IFN-gamma production, expression of surface markers for peripheral tissue trafficking and allergen specific proliferation remained unchanged during SIT treatment. However, increased proportions of CD4+CD62L(-), CD4+CD49d(hi), CD4+CCR5+ T cells expressing IL-10 were detected at 9 months of SIT compared with baseline (P < 0.05). IL-10 staining co-localized with CD4+CD25+ T cells. CONCLUSION: Clinically effective subcutaneous immunotherapy with a standardized HDM Dermatophagoides pteronyssinus preparation results in decreased numbers of IL-4+ T cells and expansion of CD4+IL-10+ T cells expressing a peripheral tissue trafficking phenotype. The co-localization of IL-10+ staining to CD4+CD25+ T cells is consistent with the induction of a T regulatory cell population by SIT. PMID- 15298561 TI - Effect of mite-impermeable mattress encasings and an educational package on the development of allergies in a multinational randomized, controlled birth-cohort study -- 24 months results of the Study of Prevention of Allergy in Children in Europe. AB - BACKGROUND: Sensitization to house dust mite (HDM) is an important risk factor for the development of asthma and allergic disease in childhood. Higher levels of HDM allergen are linked to increased sensitization to HDM. OBJECTIVE: To study the effect of mite-impermeable mattress encasings and an educational package on the development of allergies in a newborn cohort. METHODS: Six hundred and ninety six newborns at high risk of developing allergies were enrolled in three European countries (Germany, Austria, UK) in a prospective, randomized, controlled birth cohort study. Children were randomly assigned to an intervention and control group. Intervention measures included the use of mite-impermeable mattress encasings for the child's bed and a simple educational package on allergen avoidance. The control group received basic information about allergies. Children were followed up at age 6, 12, 18 and 24 months. RESULTS: 80.9% of the children were followed up to the age of 24 months. No difference in the prevalence of sensitization to HDM (control vs. intervention group: 8.4% vs. 6.1%, P=0.33) or the development of symptoms (recurrent wheezing 10.3% vs. 10.7%, nocturnal cough 12.5% vs. 12.5%) or allergic diseases (asthma 3.5% vs. 5.1%, eczema 20.0% vs. 19.6%, rhinitis 28.9% vs. 25.8%) could be found between the control and intervention group. CONCLUSION: In this study, HDM avoidance did not show a protective effect on the development of sensitization to HDM or symptomatic allergy in children at age 24 months. PMID- 15298562 TI - Bronchial hyper-responsiveness to hypertonic saline and blood eosinophilic markers in 8-13-year-old schoolchildren. AB - BACKGROUND: In adult asthma, bronchial hyper-responsiveness (BHR) to indirect stimuli reflects eosinophilic activation more closely than BHR to stimuli that directly cause smooth muscle contraction. AIM: To assess the relationship between BHR to the indirect stimulus hypertonic saline (HS), blood eosinophil numbers, and serum eosinophilic cationic protein (ECP) in children with and without current wheeze. METHODS: A cross-sectional survey among 8-13-year-old schoolchildren, using the International Study of Asthma and Allergic disease in Childhood questionnaire, bronchial challenge with HS, skin prick tests, serum IgE, blood eosinophil counts and ECP (in a subset). Based upon the presence of current wheeze (WHE) and BHR, we defined three case groups (WHE+BHR+, WHE-BHR+, WHE+BHR-) and the reference group (WHE-BHR-). By regression analyses, each case group was compared with the reference group for differences in atopic sensitization, blood eosinophil counts and serum ECP. RESULTS: Complete data were obtained for 470 children. BHR was present in 103 children (22%), 66 being asymptomatic and 37 symptomatic. Children of all three case groups were more often atopic. Sensitization to indoor allergens particularly occurred in children with BHR, irrespective of symptoms (P < 0.05). Children with WHE+BHR+ had highest values for blood eosinophils and serum ECP (P < 0.05). Children with WHE-BHR+ had less severe responsiveness. In atopic children with WHE-BHR+, serum ECP was higher than in children with WHE-BHR-(P < 0.05). CONCLUSIONS: BHR to HS is associated with blood markers of eosinophilic activation, particularly in atopic children. PMID- 15298563 TI - A case-control study of dietary and erythrocyte membrane fatty acids in asthma. AB - BACKGROUND: Epidemiological evidence suggests that increased dietary omega-6 and reduced omega-3 fatty acid intake, may have contributed to the rising prevalence of asthma, but these hypotheses have not been tested in studies comparing both dietary intake and objective measures of polyunsaturated fatty acids. OBJECTIVE: To assess whether a higher intake of omega-6 or a lower intake of omega-3 fatty acids increases the risk of asthma, by measuring dietary fatty acid intake by a food frequency questionnaire (FFQ) and erythrocyte membrane fatty acids, as an objective biomarker of intake. METHODS: We have compared individual fatty acid intake estimated by FFQ and by mass spectrometry of fasting erythrocyte cell membranes in 89 cases of asthma and 89 community-matched controls. RESULTS: The odds of asthma were increased in relation to intake of the omega-3 fatty acids eicosapentaenoic acid (odds ratio (OR) for difference between the 25th and 75th centiles of intake= 1.89, 95% CI 1.15-3.11) and docosahexaenoic acid (OR = 2.11, 95% CI 1.19-3.74). There was no evidence of any difference in erythrocyte membrane levels of omega-3 fatty acids, while the odds of asthma were reduced in relation to linoleic acid (omega-6) membrane levels (OR = 0.45, 95% CI 0.21 0.95). CONCLUSION: These findings suggest that dietary omega-3 fatty acids do not play a major role in protecting against asthma, and that higher levels of erythrocyte membrane linoleic acid are associated with a lower risk of asthma. PMID- 15298564 TI - The effect of supplementation with fish oil during pregnancy on breast milk immunoglobulin A, soluble CD14, cytokine levels and fatty acid composition. AB - BACKGROUND: Breast milk contains many immunomodulatory factors (soluble CD14 (sCD14), IgA and cytokines) with the potential to influence infant immune development. OBJECTIVE: To determine if changes in breast milk omega-3 polyunsaturated fatty acid (n-3 PUFA) composition as a result of maternal dietary fish oil supplementation during pregnancy can modify levels of these immunological parameters in breast milk. METHOD: In a randomized controlled trial, 83 atopic women received either 4 g fish oil capsules (containing 3.7 g n 3 PUFA) (n = 40) or 4 g olive oil capsules (n = 43) from 20 weeks gestation until delivery. Breast milk was collected 3 days post-partum and fatty acids were analysed by gas liquid chromatography and IgA, sCD14 and cytokines (IL-5, IL-6, IL-10, TNF-alpha and IFN-gamma) were quantitated by ELISA or time resolved fluorescence (TRF). RESULTS: Omega-3 docosahexaenoic acid (DHA; 22:6n-3) and eicosapentaenoic acid (EPA; 20:5n-3) levels were significantly higher (P < 0.001) in breast milk from women supplemented with fish oil (n = 33, DHA mean 1.15%, SD 0.47% and EPA mean 0.16%, SD 0.07%) than in samples from the control group (n = 40, DHA mean 0.50%, SD 0.17% and EPA mean 0.05%, SD 0.02%). Breast milk arachidonic acid (AA; 20:4n-6) levels were significantly lower (P = 0.045) in the fish oil group (mean 0.55%, SD 0.12%) compared with the control group (mean 0.61%, SD 0.14%). Breast milk IgA was positively correlated with DHA (P = 0.046) and 22:5n-3 (P = 0.003), but inversely correlated with linoleic acid (LA; 18:2n 6) (P=0.034). Levels of sCD14 were also positively correlated with 22:5n-3 (P=0.009). Cytokines involved in IgA synthesis (IL-10 and IL-6) were also significantly correlated with both IgA and n-3 PUFA levels, although there were no differences in the levels of breast milk IgA, sCD14 or cytokines between study groups. CONCLUSION: Supplementation with fish oil during pregnancy significantly alters early post-partum breast milk fatty acid composition. omega-3 PUFA levels were positively associated with IgA and sCD14 levels, suggesting a relationship between fatty acid status and mucosal immune function. PMID- 15298565 TI - Immunoglobulin G4 antibodies to rat urinary allergens, sensitization and symptomatic allergy in laboratory animal workers. AB - BACKGROUND AND OBJECTIVES: We have previously reported that high rat urinary allergen (RUA) exposure was not associated with increased risk of rat allergy in long-term-exposed laboratory animal (LA) workers. We aimed to assess whether strong allergen-specific IgG4 responses could explain the absence of a dose response in these subjects. We investigated whether IgG4 was associated with allergen exposure and prevalence of sensitization or respiratory symptoms to rats. The longitudinal relation between IgG4 and rat allergy was studied using data obtained during 2 years of follow-up. METHODS: Five hundred and twenty-nine LA workers answered a questionnaire on respiratory symptoms and occupational history and participated in skin prick testing. Blood samples were analysed for specific IgG4 and IgE to RUA. Exposure to RUA was estimated based on personal air samples. The relation between IgG4 and newly occurring sensitization or rat allergy was studied in workers who were not sensitized or did not report respiratory symptoms to rats. RESULTS: IgG4 titres were higher in atopic than in non-atopic subjects, and increased with higher allergen exposure. Titres were highest in subjects who were sensitized and reported respiratory symptoms to rats when compared with those who were not (geometric mean [geometric standard deviation] = 202 [5.7] vs. 8.4 [18.3] AU). The association between IgG4 and sensitization or symptomatic rat allergy was independent of estimated allergen exposure. IgG4 was a strong predictor of newly occurring sensitization and symptomatic rat allergy during follow-up in atopic and rat-sensitized subjects. CONCLUSION: High exposure to RUA is associated with a strong allergen-specific IgG4 antibody response. High anti-RUA IgG4 is a strong predictor of prevalent and incident sensitization and symptomatic rat allergy in atopic and rat-sensitized subjects. IgG4 can therefore not explain the absence of a dose response between allergen exposure and allergy in long-term-exposed workers. We consider anti-RUA IgG4 to be a marker that combines aspects of exposure and susceptibility. PMID- 15298566 TI - Sensitization to Ficus benjamina: relationship to natural rubber latex allergy and identification of foods implicated in the Ficus-fruit syndrome. AB - BACKGROUND: Ornamental Ficus benjamina (FB) has been recognized as a new indoor allergen. Little is known about the prevalence in moderately exposed subjects and the proposed association with fruit and Hevea latex hypersensitivity. OBJECTIVE: To study the prevalence of FB sensitization and the relationship with Hevea latex allergy, to identify cross-reacting fruits, and to characterize the responsible allergens. METHODS: A skin prick test solution prepared from FB latex (200 microg/mL) was included in our routine screening programme for suspect inhalant allergy. Patients reacting with the FB extract were further skin tested with exotic fruits by the prick-to-prick method. Inhibition of fig and FB CAP by FB latex, fig (Ficus carica), kiwi, the thiolproteases ficin and papain, Hevea latex and rHev b 6.02 (hevein) was performed in selected patients. RESULTS: Of 2662 patients with a positive skin test to any aeroallergen, 66 (2.5%) reacted with FB. Ten patients showed isolated sensitization to FB. Although FB-positive subjects were more often co-sensitized to Hevea latex than FB-negative (10.6% vs 3.8%, P< 0.01), nearly 90% tested negative for Hevea latex. Sensitization to FB was specifically associated with positive skin tests to fresh fig (83%), dried fig (37%), kiwi fruit (28%), papaya (22%), avocado (19%), banana (15%), and pineapple (10%) (n = 54). Clinical reactions were reported mainly from fresh and dried fig and kiwi (47%, 60%, and 64%, respectively, of skin test-positive patients), including seven patients with systemic reactions (urticaria, angiooedema, asthma). CAP to fig in 11 patients with clinical fruit allergy was inhibited on average by 87% by FB latex, 89% by fresh fig, 80% by dried fig, 38% by kiwi (100 microg/mL each), and by 59% and 44% by ficin and papain (50 microg/mL), respectively. No inhibition was obtained with Hevea latex and rHev b 6.02. CAP to FB was inhibited on average by 95% by FB, 60% by fresh fig, 41% by ficin, 29% by papain, and less than 7% by rubber latex allergens. CONCLUSIONS: Sensitization to FB latex is found in 2.5% of atopic individuals and mostly occurs independently of Hevea latex allergy. Sensitization is commonly associated with allergic reactions to figs and other tropical fruits ('Ficus-fruit syndrome'). This cross-reactivity is mediated at least in part by thiolproteases. PMID- 15298567 TI - Urinary leukotriene E4 in Henoch-Schonlein purpura. AB - BACKGROUND: Children with Henoch-Schonlein purpura (HSP) occasionally have allergic disease. We have previously shown that pranlukast hydrate was effective for purpura in HSP. Pranlukast hydrate is a leukotriene (LT) receptor antagonist; therefore, it is likely that LTs take part in the cause of HSP. Urinary leukotriene E4 (LTE4u) levels are a useful index of whole-body cysteinyl LT production in vivo. In this study, LTE4u was examined in children with HSP. OBJECTIVE: The purpose of this study was to examine the relation between the level of LTE4u and the cause of HSP. METHODS: Eighteen HSP children (six boys and 12 girls) and six healthy children were enrolled. RESULTS: LTE4u levels in patients with HSP were significantly higher (P< 0.05) at the onset than those in healthy children. Four weeks therapy with pranlukast hydrate lowers LTE4u levels in patients with HSP (P< 0.05). There were no differences in LTE4u between the group of HSP patients with purpura nephritis and the group of HSP patients without purpura nephritis. CONCLUSION: These results indicate that csyteinyl LTs may play a role in the pathophysiology of purpura in HSP. PMID- 15298568 TI - Increase in urinary leukotriene B4 glucuronide concentration in patients with aspirin-intolerant asthma after intravenous aspirin challenge. AB - BACKGROUND: Aspirin challenge of aspirin-intolerant asthma (AIA) patients causes a significant increase in leukotriene E4 (LTE4) concentration in urine. However, knowledge on leukotriene B4 (LTB4) generation in patients with AIA is insufficient. Recent research has demonstrated that exogenously administered LTB4 is excreted as glucuronide into the urine in human healthy subjects. OBJECTIVE: The purpose of this study is to estimate urinary LTB4 glucuronide (LTBG) concentration in the clinically stable condition in healthy subjects and asthmatic patients and to investigate changes in urinary LTBG concentration in patients with AIA after aspirin challenge. METHODS: A provocation test was performed by intravenous aspirin challenge. After urine was hydrolysed by beta glucuronidase, the fraction containing LTB4 was purified by high-performance liquid chromatography and LTB4 concentration was quantified by enzyme immunoassay. Urinary LTBG concentration was calculated as the difference between the concentration obtained with hydrolysis and that without hydrolysis. RESULTS: (1) After hydrolysis, the presence of urinary LTB4 was verified by gas chromatography-mass spectrometry-selected ion monitoring. (2) The urinary LTBG concentration was significantly higher in the asthmatic patients than in the healthy subjects (median, 5.37 pg/mg creatinine [range 1.2-13] vs. 3.32 pg/mg creatinine [range, 0.14-10.5], P = 0.0159). (3) The patients with AIA (n = 7), but not those with aspirin-tolerant asthma (n = 6), showed significant increases in LTBG and LTE4 excretions after aspirin challenge. (4) When the concentrations after aspirin challenge were analysed simultaneously, a significant linear correlation was observed between urinary LTBG concentration and urinary LTE4 concentration in patients with AIA (Spearman's rank correlation test, r = 0.817, P = 0.0003). CONCLUSION: LTBG is present in human urine, albeit at a concentration lower than urinary LTE4. In addition to a marked increase in cysteinyl-leukotriene production, aspirin challenge induced LTB4 production in AIA patients. PMID- 15298569 TI - Specific immunoglobulin E for staphylococcal enterotoxins in nasal polyps from patients with aspirin-intolerant asthma. AB - BACKGROUND: Nasal polyps infiltrated with eosinophils are commonly found in chronic asthmatic patients, more frequently in those with aspirin-intolerant asthma (AIA) than aspirin-tolerant asthma (ATA). Some studies have suggested a contribution of superantigens derived from Staphylococcus sp to nasal polyposis and eosinophilia, but their relative importance in AIA and ATA subjects is unknown. OBJECTIVE: We investigated whether local production of specific IgE to staphylococcal enterotoxins A and B (SEA and SEB) and relationships with markers of eosinophilic inflammation differ in the nasal polyps of AIA and ATA subjects. METHODS: Fifteen AIA subjects with positive responses to lysine-aspirin bronchoprovocation and 15 ATA subjects underwent polypectomy. Immunoassays were used to quantify eosinophil cationic protein (ECP), IL-5, mast cell tryptase, soluble IL-2 receptors (sIL-2R), total IgE, and specific IgE for SEA and SEB. RESULTS: ECP levels in nasal polyp homogenates were higher in AIA subjects than in ATA subjects (P < 0.02), with no significant differences in tryptase, IL-5 or sIL-2R. Total IgE, and specific IgE to both SEA and SEB, were detectable in some nasal polyps from both subject groups, but median levels were markedly higher in AIA subjects than in ATA subjects (P = 0.04, 0.01, 0.05, respectively). Levels of specific IgE to SEA and SEB correlated significantly with levels of ECP and IL-5, but not those of tryptase or sIL-2R. CONCLUSION: These findings suggest that staphylococcal superantigens may drive local eosinophilic inflammation in nasal polyp tissue, and that this is exacerbated in subjects with AIA. PMID- 15298570 TI - Interactions between eotaxin, histamine and mast cells in early microvascular events associated with eosinophil recruitment to the site of allergic skin reactions in humans. AB - BACKGROUND: The mechanism whereby allergen induces eotaxin expression at the site of allergic inflammation is incompletely understood. Structural cells, including endothelial cells, are a major source of eotaxin. OBJECTIVE: We have investigated, in vivo and in vitro, the relationship between mast cell activation and the expression of eotaxin (eotaxin 1) by endothelial cells. METHODS: The effects of intradermal allergen challenge and histamine injection on eotaxin mRNA and protein generation were studied in atopic subjects using immunofluorescence, immunohistochemistry and in situ hybridization. Histamine-induced expression of eotaxin mRNA and protein by endothelial cells was also measured, as was histamine induced eosinophil adhesion to cultured endothelial cells. RESULTS: A rapid increase in degranulating cutaneous mast cells, together with a concomitant increase in eosinophils, was observed 60 min after allergen challenge. This was accompanied by the appearance of immunoreactive eotaxin that peaked at 1 h around blood vessels and at 3 h within the tissue. Intradermal histamine injection produced an increase in the number of eotaxin+ cells in the tissues, which was maximal at the 3-h time-point. In vitro, endothelial cells produced eotaxin mRNA and protein product in a dose- and time-dependent fashion following incubation with histamine, an effect that was blocked by levocetirizine. Pre-incubation of endothelial cells with histamine also induced a significant increase in eosinophil adherence, an effect that was inhibited with an anti-eotaxin blocking monoclonal antibody. CONCLUSION: The antigen-induced expression of eotaxin by endothelial cells and the adherence and subsequent migration of eosinophils from the microvasculature to the tissues are rapid events partially under the control of histamine released from degranulating mast cells. PMID- 15298571 TI - Differential modulation of human basophil functions through prostaglandin D2 receptors DP and chemoattractant receptor-homologous molecule expressed on Th2 cells/DP2. AB - BACKGROUND: Both prostaglandin (PG) D receptor (DP) and CRTH2 (chemoattractant receptor-homologous molecule expressed on Th2 cells)/DP2 are high-affinity receptors for PGD2. Previous studies have demonstrated that PGD2 enhances releasability and induces CRTH2/DP2-mediated migration in human basophils, but the precise effects of PGD2 on basophils as well as receptor usage have not been fully clarified. OBJECTIVE: We comprehensively explored the roles of DP and CRTH2/DP2 in basophil functions by using selective agonists and antagonists for each receptor. METHODS: DP and CRTH2/DP2 transcripts were quantified by real-time PCR. We studied the effects of selective agonists (DP: BW245C; CRTH2/DP2: 13,14 dihydro-15-keto (DK)-PGD2) and/or antagonists (DP: BWA868C; CRTH2/DP2: ramatroban) on Ca2+ mobilization, migration, degranulation, CD11b expression and survival of human basophils. RESULTS: Basophils expressed transcripts of both DP and CRTH2/DP2, but the levels of CRTH2/DP2 transcripts were ca. 100-fold higher compared with DP transcripts. Ca2+ influx was induced in basophils by either PGD2 or DK-PGD2/CRTH2 agonist but not by BW245C/DP agonist. Basophils treated with PGD2 were completely desensitized to subsequent stimulation with DK-PGD2, but not vice versa. DK-PGD2 as well as PGD2 up-regulated CD11b expression, induced migration and enhanced degranulation, and those effects were completely antagonized by ramatroban/CRTH2 antagonist. In contrast, BW245C/DP agonist exhibited an inhibitory effect on basophil migration and IgE-mediated degranulation, and the migration inhibitory effect was effectively antagonized by BWA868C/DP antagonist. On the other hand, while PGD2 significantly shortened the basophil life-span, neither DK-PGD2/CRTH2 agonist nor BW245C/DP agonist did. CONCLUSION: CRTH2/DP2 is primarily responsible for the pro-inflammatory effects of PGD2 on human basophils, while DP introduces negative signals capable of antagonizing the effects of CRTH2/DP2 in these cells. The effects of PGD2 on longevity imply a mechanism(s) other than via DP or CRTH2/DP2. CRTH2/DP2 on basophils may afford opportunities for therapeutic targeting in allergic inflammation. PMID- 15298572 TI - Interleukin-4 increases murine airway response to kinins, via up-regulation of bradykinin B1-receptors and altered signalling along mitogen-activated protein kinase pathways. AB - BACKGROUND: IL-4 is believed to play a role in asthma and chronic obstructive pulmonary disease through promotion of eosinophilic inflammation and mucus hypersecretion. Whether IL-4 can induce a direct effect on airway smooth muscle remains unknown. OBJECTIVE: To investigate the effect of IL-4 on airway smooth muscle, focusing on the contractile response to des-Arg9-bradykinin and bradykinin. METHODS: Tracheal segments from murine airways were cultured for 1-8 days in the absence and presence of IL-4. The smooth muscle response induced by des-Arg9-bradykinin and bradykinin was investigated in myographs. Expression levels for the IL-4-, bradykinin B1- and B2-receptors were characterized using RT PCR. Specific inhibitors were used to study signal changes along the IL-4 receptor- (IL-4R-) coupled mitogen-activated protein (MAP) kinase (MAPK) pathways. RESULTS: IL-4 treatment increased the contractile response to des-Arg9 bradykinin and bradykinin in a concentration- and time-dependent manner. Dexamethasone and the transcriptional inhibitor actinomycin D blocked this effect. c-Jun N-terminal kinase inhibitor SP600125 also blocked the effect of both des-Arg9-bradykinin and bradykinin, whereas p38 inhibitor SB203580 blocked only the former and the MAPKK inhibitor PD098059, only the latter agonist responses. IL-4 treatment increased the mRNA levels representing bradykinin B1- but not B2-receptors. Levels of IL-4R were not altered during culture. CONCLUSION: Long-term exposure to IL-4 increases the contractile response induced by des-Arg9-bradykinin and bradykinin in cultured murine airways. This effect appears to be mediated via an up-regulation of B1-receptors and altered signalling along the MAPK pathways. PMID- 15298573 TI - Allergic airway inflammation is exacerbated during acute influenza infection and correlates with increased allergen presentation and recruitment of allergen specific T-helper type 2 cells. AB - BACKGROUND: Respiratory viral infections are a leading cause of the hospitalization of asthmatics, however, the cellular immunological interactions which underlie these two diseases remain elusive. OBJECTIVE: We sought to characterize the effect influenza viral infection has on allergic airway inflammation and to identify the cellular pathways involved. METHODS: We have used an ovalbumin (OVA) model of allergic airway inflammation, which involves sensitization of animals with OVA adsorbed in alum adjuvant followed by an intranasal challenge with OVA in phosphate-buffered saline. To study T cell recruitment into the lung, we adoptively transferred in vitro activated T cell receptor-transgenic T cells, which were subsequently identified by fluorescence activated cell sorting (FACS) analysis. In addition, to study in vivo dendritic cell (DC) migration, we administered fluorescently labelled dextran and identified DCs that had phagocytosed it by FACS analysis. RESULTS: We found that different stages of influenza infection had contrasting effects upon the outcome of OVA-induced allergic airway inflammation. The allergic response against OVA was exacerbated during the acute stage of influenza infection; however, mice were protected against the development of airway eosinophilia at late time-points following infection. We investigated the mechanisms responsible for the virus induced exacerbation and found that the response was partially independent of IL 4 and that there was increased delivery of inhaled allergens to the draining lymph node during the acute stage of the infection. In addition, virus-induced inflammation in the lung and draining lymph node resulted in the non-specific recruitment of circulating allergen-specific effector/memory cells. CONCLUSION: In addition to virus-mediated damage to the lung and airways, influenza viral infection can also enhance unrelated local allergic responses. PMID- 15298574 TI - Effects of primary and secondary low-grade respiratory syncytial virus infections in a murine model of asthma. AB - BACKGROUND: Respiratory syncytial virus (RSV) infection is known to develop and exacerbate asthma in young children. In adult, RSV causes recurrent but asymptomatic infections. However, the impact of asymptomatic RSV infection on adult asthma is yet to be determined. The present study is designed to determine the effects of primary and secondary low-grade RSV infections on allergic airway inflammation in a murine model of allergic asthma. METHODS: A low-grade RSV (2 x 10(3) plaque-forming units/mouse) was inoculated, and this caused neither pulmonary inflammation nor symptoms but induced significant IFN-gamma production in thoracic lymph nodes. To investigate interaction between low-grade virus and Dermatophagoides farinae (Df), airway hyper-responsiveness, lung inflammation and cytokine production from thoracic lymph nodes were compared after primary and secondary low-grade RSV infections in four groups of mice; control, Df allergen sensitized, RSV-infected and Df-sensitized RSV-infected mice. A direct comparison between low- and high-grade RSV infections was also performed in primary infection. To investigate the role of IL-5 during secondary RSV infection, anti IL-5 monoclonal antibody (anti-IL-5 mAb) was injected in mice and similar parameters were compared in four groups of mice. RESULTS: Primary high-grade RSV infection increased allergen-induced airway inflammation, while primary low-grade RSV infection attenuated allergen-induced airway inflammation concomitant with significant IFN-gamma production in lung-draining lymph nodes. In marked contrast, secondary low-grade RSV infection increased both IFN-gamma and IL-5 production, resulting in exacerbation of allergen-induced airway inflammation. Anti-IL-5 mAb treatment in secondary low-grade RSV infection and Df allergen sensitized mice attenuated virus and allergen-induced airway inflammation. CONCLUSIONS: Low-grade RSV infection per se does not cause pulmonary inflammation, whereas it induces a significant immunological response in the allergen-sensitized host. These results indicate that subclinical and recurrent RSV infection may play an important role in exacerbation and maintenance of asthma in adults, wherein IL-5 is critically involved. PMID- 15298575 TI - Resiquimod, a new immune response modifier from the family of imidazoquinolinamines, inhibits allergen-induced Th2 responses, airway inflammation and airway hyper-reactivity in mice. AB - BACKGROUND: Allergen-induced sensitization and airway disease are the results of adverse immune reactions against environmental antigens that may be prevented or inhibited by immune modifying strategies. OBJECTIVE: To investigate the effects of the novel immune response modifier resiquimod (R-848), from the family of imidazol-derivates, in a murine model of allergen-mediated Th2-immune responses and concomitant airway inflammation and airway hyper-reactivity. METHODS: BALB/c mice were systemically sensitized with ovalbumin (OVA) on days 1 and 14 and challenged with OVA aerosol on days 28 and 29. R-848 was applied intranasally to sensitized animals once prior to the first allergen airway challenge, on day 27. RESULTS: A single application of R-848 significantly reduced numbers of eosinophils and lymphocytes in bronchoalveolar lavage fluid and inhibited mucus gland hyperplasia, compared with sensitized and challenged controls. Associated with the decrease in airway inflammation, single intranasal treatment with R-848 abolished the development of airway hyper-reactivity after allergen sensitization and airway challenges. Additionally, Th2-cytokine production in lung tissues from sensitized and R-848-treated animals was reduced, whereas IL-12 and IFN-gamma production was increased, compared with non-treated sensitized mice. CONCLUSION: These data indicate that R-848 effectively inhibits allergen-induced airway inflammation and hyper-reactivity by modulation of increased Th2-immune responses. PMID- 15298576 TI - Enhanced airway inflammation and decreased subepithelial fibrosis in interleukin 6-deficient mice following chronic exposure to aerosolized antigen. AB - BACKGROUND: Airway inflammation and remodelling are characteristic features of chronic asthma. OBJECTIVE: To elucidate the role of interleukin (IL)-6 in airway responses to chronic antigen exposure. METHODS: We compared airway inflammation, subepithelial collagen deposition, cytokine mRNA expression, and airway responsiveness between IL-6-deficient and wild-type (WT) mice following sensitization and repeated exposure to ovalbumin (OVA) three times a week for 8 weeks. RESULTS: The repeated exposure to OVA induced infiltration of eosinophils, neutrophils, and lymphocytes into the airway, and caused thickening of the basement membrane and subepithelial fibrosis. IL-6-deficient mice exhibited more pronounced infiltration of these cells, a thinner basement membrane, and decreased subepithelial fibrosis, compared with WT mice. The repeated OVA exposure increased expression of IL-4, IL-13, eotaxin, monocyte chemoattractant protein-1 (MCP-1), and transforming growth factor-beta1 mRNA in WT mice. Among these factors, expression of IL-13 and MCP-1 mRNA was further enhanced in IL-6 deficient mice, compared with WT mice. However, both WT and IL-6-deficient mice exhibited similar levels of airway responsiveness to increasing doses of methacholine, even after repeated exposure to OVA. CONCLUSION: These results suggest that IL-6 has dual roles in the chronic phase of asthma: down-regulation of inflammatory cell infiltration and enhancement of airway remodelling. PMID- 15298579 TI - Assessing Cost-Effectiveness--Mental Health: introduction to the study and methods. AB - OBJECTIVE: The Assessing Cost-Effectiveness - Mental Health (ACE-MH) study aims to assess from a health sector perspective, whether there are options for change that could improve the effectiveness and efficiency of Australia's current mental health services by directing available resources toward "best practice" cost effective services. METHOD: The use of standardized evaluation methods addresses the reservations expressed by many economists about the simplistic use of League Tables based on economic studies confounded by differences in methods, context and setting. The cost-effectiveness ratio for each intervention is calculated using economic and epidemiological data. This includes systematic reviews and randomised controlled trials for efficacy, the Australian Surveys of Mental Health and Wellbeing for current practice and a combination of trials and longitudinal studies for adherence. The cost-effectiveness ratios are presented as cost (A$) per disability-adjusted life year (DALY) saved with a 95% uncertainty interval based on Monte Carlo simulation modelling. An assessment of interventions on "second filter" criteria ("equity", "strength of evidence", "feasibility" and "acceptability to stakeholders") allows broader concepts of 'benefit' to be taken into account, as well as factors that might influence policy judgements in addition to cost-effectiveness ratios. CONCLUSIONS: The main limitation of the study is in the translation of the effect size from trials into a change in the DALY disability weight, which required the use of newly developed methods. While comparisons within disorders are valid, comparisons across disorders should be made with caution. A series of articles is planned to present the results. PMID- 15298580 TI - Cost-effectiveness of cognitive behavioural therapy and selective serotonin reuptake inhibitors for major depression in children and adolescents. AB - OBJECTIVE: To assess from a health sector perspective the incremental cost effectiveness of cognitive behavioural therapy (CBT) and selective serotonin reuptake inhibitors (SSRIs) for the treatment of major depressive disorder (MDD) in children and adolescents, compared to "current practice". METHOD: The health benefit is measured as a reduction in disability-adjusted life years (DALYs), based on effect size calculations from meta-analysis of randomised controlled trials. An assessment on second stage filter criteria ("equity", "strength of evidence", "feasibility" and "acceptability to stakeholders") is also undertaken to incorporate additional factors that impact on resource allocation decisions. Costs and benefits are tracked for the duration of a new episode of MDD arising in eligible children (age 6-17 years) in the Australian population in the year 2000. Simulation-modelling techniques are used to present a 95% uncertainty interval (UI) around the cost-effectiveness ratios. RESULTS: Compared to current practice, CBT by public psychologists is the most cost-effective intervention for MDD in children and adolescents at A$9000 per DALY saved (95% UI A$3900 to A$24 000). SSRIs and CBT by other providers are less cost-effective but likely to be less than A$50 000 per DALY saved (> 80% chance). CBT is more effective than SSRIs in children and adolescents, resulting in a greater total health benefit (DALYs saved) than could be achieved with SSRIs. Issues that require attention for the CBT intervention include equity concerns, ensuring an adequate workforce, funding arrangements and acceptability to various stakeholders. CONCLUSIONS: Cognitive behavioural therapy provided by a public psychologist is the most effective and cost-effective option for the first-line treatment of MDD in children and adolescents. However, this option is not currently accessible by all patients and will require change in policy to allow more widespread uptake. It will also require "start-up" costs and attention to ensuring an adequate workforce. PMID- 15298581 TI - Cost-effectiveness of dexamphetamine and methylphenidate for the treatment of childhood attention deficit hyperactivity disorder. AB - OBJECTIVE: To analyze from a health sector perspective the cost-effectiveness of dexamphetamine (DEX) and methylphenidate (MPH) interventions to treat childhood attention deficit hyperactivity disorder (ADHD), compared to current practice. METHOD: Children eligible for the interventions are those aged between 4 and 17 years in 2000, who had ADHD and were seeking care for emotional or behavioural problems, but were not receiving stimulant medication. To determine health benefit, a meta-analysis of randomized controlled trials was performed for DEX and MPH, and the effect sizes were translated into utility values. An assessment on second stage filter criteria ("equity", "strength of evidence", "feasibility" and "acceptability to stakeholders") is also undertaken to incorporate additional factors that impact on resource allocation decisions. Simulation modelling techniques are used to present a 95% uncertainty interval (UI) around the incremental cost-effectiveness ratio (ICER), which is calculated in cost (in A$) per DALY averted. RESULTS: The ICER for DEX is A$4100/DALY saved (95% UI: negative to A$14 000) and for MPH is A$15 000/DALY saved (95% UI: A$9100-22 000). DEX is more costly than MPH for the government, but much less costly for the patient. CONCLUSIONS: MPH and DEX are cost-effective interventions for childhood ADHD. DEX is more cost-effective than MPH, although if MPH were listed at a lower price on the Pharmaceutical Benefits Scheme it would become more cost-effective. Increased uptake of stimulants for ADHD would require policy change. However, the medication of children and wider availability of stimulants may concern parents and the community. PMID- 15298583 TI - What determines compulsory community treatment? A logistic regression analysis using linked mental health and offender databases. AB - OBJECTIVE: Western Australia has one of the highest published rates of the use of compulsory treatment orders in the English-speaking world. Differences in patient characteristics, legislation and service setting may explain variations in the reported efficacy of compulsory community treatment. Our objective is to investigate predictors of Community Treatment Orders (CTO) placement in the first year of implementation in Western Australia and see if there were any differences in the type of patients placed on these orders compared to other studies. METHOD: A population-based record linkage study of Mental Health and Offender Databases comparing 265 patients on CTOs with a consecutive control group (CCG) of equal number matched on date of discharge from inpatient care or CTO placement. RESULTS: Previous health service use, after-care placement, mental disorder history including schizophrenic history, a positive forensic history of violence to others as well as patient's marital status were the significant predictors of CTO placement. CONCLUSIONS: Studies of compulsory community treatment appear to be of similar populations. In spite of the comparatively high rate of use, psychiatrists in Western Australia do not appear to be applying community treatment orders to different types of patient compared to elsewhere. We need further research to establish the relative contribution of patient characteristics, legislation and service setting toward the use and outcome of compulsory community treatment. PMID- 15298582 TI - Cost-effectiveness of psychological and pharmacological interventions for generalized anxiety disorder and panic disorder. AB - OBJECTIVE: To assess from a health sector perspective the incremental cost effectiveness of interventions for generalized anxiety disorder (cognitive behavioural therapy [CBT] and serotonin and noradrenaline reuptake inhibitors [SNRIs]) and panic disorder (CBT, selective serotonin reuptake inhibitors [SSRIs] and tricyclic antidepressants [TCAs]). METHOD: The health benefit is measured as a reduction in disability-adjusted life years (DALYs), based on effect size calculations from meta-analyses of randomised controlled trials. An assessment on second stage filter criteria ("equity", "strength of evidence", "feasibility" and "acceptability to stakeholders") is also undertaken to incorporate additional factors that impact on resource allocation decisions. Costs and benefits are calculated for a period of one year for the eligible population (prevalent cases of generalized anxiety disorder/panic disorder identified in the National Survey of Mental Health and Wellbeing, extrapolated to the Australian population in the year 2000 for those aged 18 years and older). Simulation modelling techniques are used to present 95% uncertainty intervals (UI) around the incremental cost effectiveness ratios (ICERs). RESULTS: Compared to current practice, CBT by a psychologist on a public salary is the most cost-effective intervention for both generalized anxiety disorder (A$6900/DALY saved; 95% UI A$4000 to A$12 000) and panic disorder (A$6800/DALY saved; 95% UI A$2900 to A$15 000). Cognitive behavioural therapy results in a greater total health benefit than the drug interventions for both anxiety disorders, although equity and feasibility concerns for CBT interventions are also greater. CONCLUSIONS: Cognitive behavioural therapy is the most effective and cost-effective intervention for generalized anxiety disorder and panic disorder. However, its implementation would require policy change to enable more widespread access to a sufficient number of trained therapists for the treatment of anxiety disorders. PMID- 15298584 TI - Factors predicting discharge of patients from community residential facilities: a longitudinal study from Italy. AB - OBJECTIVE: Community residential facilities for psychiatric patients have increased in Italy in the last years, but little information is available on their use, the patients they host, the interventions they deliver and the rate at which they discharge patients. To investigate these issues, we conducted a longitudinal study in 2000-2001 on all the community residential facilities in Lombardy, a large region in North Italy. METHOD: The study base comprised all the patients residing in the community residential facilities identified in Lombardy in 2000. Out of the 196 community residential facilities identified, 91% agreed to participate. The study sample consisted of all the patients living in the residential facilities on 15 November, 2000. A total of 1792 patients were recruited and described. RESULTS: In the study period, a total of 316 patients were discharged. Among these, 191 (11%) went to lower-protection settings or home and 49 (3%) to higher-protection settings. The probability of discharge to lower protection settings and home was higher for people in residential care centres, not coming from a psychiatric hospital, having shorter duration of the current admission, having work at the time of admission and with a low HoNOS score. Associations were found between discharge to higher-protection settings and old age, inadequate accommodation in staff opinion, and the public sector managing the facility. CONCLUSIONS: Turn-over of patients in the community residential facilities was limited. Discharges to higher-protection settings were related to need for specific care for older patients. Type of facility and duration of stay predicted discharge to lower-protection facilities and home independently from other patient characteristics. If a higher turn-over and a more extensive use of this resource must be achieved, roles of other psychiatric and social community based services should be taken into account. PMID- 15298585 TI - Rurality and mental health: the role of accessibility. AB - OBJECTIVE: The absence of an agreed definition of "rural" limits the utility of existing research into a possible relationship between rurality of residence and mental health. The present study investigates the bipolar dimension accessibility/remoteness as a possible correlate of mental health. METHOD: A continuous area of non-metropolitan Australia was selected to provide a range of scores on the Accessibility/Remoteness Index of Australia (ARIA). A questionnaire measuring demographics, the five-factor model of personality and three aspects of mental health (distress, disability and wellbeing) was mailed to 20 000 adults selected randomly from electoral rolls. RESULTS: Responses were received from 7615 individuals (response rate = 40.5%; 57.1% female). ARIA was not associated with either distress or disability measures, but a small negative association was found between accessibility and two measures of wellbeing. Individuals residing in locales with better access to services and opportunities for interaction reported higher levels of satisfaction with life (SWL) and positive affect (PA). Adjusting statistically for a range of demographic and personality correlates did not alter the effect of ARIA on SWL. The effect on PA remained significant after adjusting for demographics, but not once personality correlates entered the model. CONCLUSIONS: By sampling across a single proposed parameter of rurality, a novel profile of correlations was identified. In accord with existing data, accessibility was not associated with distress or disability. In contrast, accessibility was positively associated with the wellbeing aspect of mental health. Further attention to the measurement of rural place and the exploration of accessibility as a parameter with mental health relevance, is warranted. PMID- 15298586 TI - Is the Australian National Survey of Mental Health and Wellbeing a reliable guide for health planners? A methodological note on the prevalence of depression. AB - OBJECTIVE: To consider whether the prevalence of depression reported in the Australian National Survey of Mental Health and Wellbeing is a reliable guide for mental health planners. METHOD: A comparison of methodologies for the detection of depression in the Australian National Survey and a South Australian survey. RESULTS: The Australian National Survey using the Composite International Diagnostic Interview (CIDI) reported considerably less depression than a South Australian survey, which used the mood module of the PRIME-MD 1000 study. Although the PRIME-MD may over-diagnose depression, it is probable that the preclusion criteria of the CIDI result in an under-reporting of depression. CONCLUSIONS: It is probable that the Australian National Survey under-estimates the prevalence of depression in the community. This has implications not only in assessing the morbidity and economic burden of depression, but also for the planning of future mental health services. PMID- 15298587 TI - Comparing psychiatric diagnoses generated by the Strengths and Difficulties Questionnaire with diagnoses made by clinicians. AB - OBJECTIVE: To examine the level of agreement between clinical diagnoses by a community child and adolescent mental health service (CAMHS) and diagnoses generated by the Strengths and Difficulties Questionnaire (SDQ). METHOD: A computerized algorithm developed by Goodman et al. was used at a community CAMHS in Australia to predict child psychiatric diagnoses on the basis of the symptom and impact scores derived from the SDQ completed by parents (n = 130), teachers (n = 101) and young people (n = 38). These diagnoses were compared with the diagnoses made by clinicians in a multidisciplinary community outpatient team and an independent clinician that examined the case notes and was blind to the SDQ scores. RESULTS: The level of agreement between SDQ generated diagnoses and clinical team diagnoses was moderate to high, ranging from 0.39 to 0.56. Correlations between the SDQ and an independent clinician ranged from 0.26 to 0.43. All were statistically significant. CONCLUSION: The SDQ is a useful instrument to aid clinicians in diagnosis and could be used as part of the initial assessment process. PMID- 15298588 TI - Australian data and psychometric properties of the Strengths and Difficulties Questionnaire. AB - OBJECTIVE: We examine the Australian psychometric properties of the Strengths and Difficulties Questionnaire (SQD), a brief screening measure of behavioural and emotional problems in children and adolescents. METHOD: Using a large community sample (n = 1359) of young Australian children (4-9 years), we assessed the internal consistency, stability, and external validity of the parent-report SDQ. Normative data and cut-offs were also produced. RESULTS: Moderate to strong internal reliability was exhibited across all SDQ subscales, and support was found for the original five-factor structure of the measure. Adequate validity was evidenced in the relationship of these scales to one another, while correlations between the SDQ subscales, teacher ratings, and diagnostic interviews demonstrated sound external validity. SDQ total difficulties scores were associated with concurrent treatment status and scores over a 12-month period were stable. CONCLUSIONS: The current study of the SDQ with Australian children presents evidence of sound psychometric properties. Being the first study to empirically support the use of the SDQ in Australia, it is recommended that the youth and teacher-report forms of the measure receive similar attention in the future. PMID- 15298589 TI - Psychopathic spectrum disorders: is this a valid and useful construct? PMID- 15298590 TI - Type D personality, cortisol and cardiac disease. PMID- 15298591 TI - Repeated clinical episodes of Wernicke-Korsakoff syndrome. PMID- 15298598 TI - Helicobacter pylori infection in two areas in Japan with different risks for gastric cancer. AB - BACKGROUND: We evaluated the relationship between Helicobacter pylori and various factors associated with gastric cancer in two areas in Japan with different risks for mortality due to gastric cancer. METHODS: A total of 250 sera from Niigata and 209 from Okinawa were used. H. pylori antibody and CagA antibody were measured by antigen-specific ELISAs. Serum gastrin and pepsinogen levels were determined by RIA. RESULTS: Although there was no significant difference in H. pylori prevalence among the persons in Niigata (50%) and Okinawa (42%), CagA prevalence in these populations was significantly different, at 41% and 26%, respectively (OR = 1.98, 95%CI: 1.33-2.95, P < 0.01). Serum gastrin levels in Niigata were significantly lower than those in Okinawa in H. pylori-negative persons (P < 0.01). The serum pepsinogen I/II ratio in Niigata was significantly lower than that in Okinawa in H. pylori positive persons (P < 0.01), whereas there was no significant difference in H. pylori-negative persons. Among those positive for H. pylori, serum pepsinogen I/II ratio in Niigata was significantly lower than that in Okinawa in CagA-negative persons (P < 0.01), whereas no significant difference was observed in CagA-positive persons. CONCLUSIONS: These results suggest that the difference in the mortality ratio of gastric cancer between Niigata and Okinawa is mainly associated with the difference between areas in the prevalence of cagA-positive strains rather than that of H. pylori itself. PMID- 15298599 TI - Diversity of vacA and cagA genes of Helicobacter pylori in Japanese children. AB - BACKGROUND: Helicobacter pylori infection is generally acquired in childhood and persists as an asymptomatic infection for decades in most infected individuals. Only a minority develops a clinical outcome even in childhood, such as peptic ulcer. It has been reported that H. pylori infection with the type I strain, which expresses the VacA and CagA antigen, is associated with peptic ulcer. AIM: We examined the diversity of vacA and cagA genes in isolates obtained from Japanese paediatric patients with peptic ulcer or chronic gastritis to investigate the relationship between genetic diversity and clinical outcome. METHODS: The diversity of vacA and cagA genes was investigated by PCR and sequence analysis in 30 isolates obtained from Japanese paediatric patients with peptic ulcer (eight strains) or chronic gastritis (22 strains). RESULTS: All isolates from Japanese children were cagA-positive strains. Twenty-six strains (86.7%) had East Asian type CagA, and 4 (13.3%) had Western type CagA. The predominant vacA genotype was s1c/m1b (22/30, 73.3%). There was no significant association between the diversity of cagA and vacA genes and clinical outcome. All four children infected with Western CagA strain had a history of overseas travel or residence. CONCLUSION: The predominant genotype of H. pylori in Japanese children is East Asian CagA and vacA s1c/m1b genotype, regardless of clinical outcome. Japanese H. pylori strains are homogeneously of the East Asian type; however, Western strains can be introduced into Japan concomitant with host movement from foreign countries in childhood. PMID- 15298600 TI - Analysis of the cag pathogenicity island and IS605 of Helicobacter pylori strains isolated from patients with gastric cancer in Japan. AB - BACKGROUND: CagA protein is encoded by the cagA gene, which is part of the cag pathogenicity island (PAI) in Helicobacter pylori. Insertion sequence (IS) elements are a diverse set of specialized DNA segments that can move to new sites in bacterial genomes. AIM: To determine the role of cagPAI and IS605 in the development of gastric cancer, we analysed cagPAI from patients with gastric cancer and compared the results with the host's CagA antibody status. METHODS: H. pylori strains were isolated from 29 gastric cancer patients, and CagA status was determined by measuring serum antibody against CagA. The cagPAI region and IS605 were determined by PCR. RESULTS: CagA seropositivity tended to be higher in the IS605/PAI+ group (5/7, 71.4%) than in the IS605/PAI- group (9/22, 40.9%). Association with cag13 was more frequent in the IS605+ group (92.3%; 12/13) than in the IS605- group (25.0%; 4/16; P = 0.0005). CONCLUSIONS: cag13 may be associated with the presence of IS605 in gastric cancer patients. PMID- 15298601 TI - Sex differences in mucosal response to Helicobacter pylori infection in the stomach and variations in interleukin-8, COX-2 and trefoil factor family 1 gene expression. AB - BACKGROUND: Gastric cancer incidence in men is almost double that in women. We investigated mucosal responses in the stomach against Helicobacter pylori (H. pylori) infections to elucidate the interindividual or sex-related differences, which may in turn be associated with gastric cancer incidence, mucosal changes of stomach as measured by the Sydney System, and interleukin-8, cyclooxygenase-2 and trefoil factor family 1 (TFF1) gene expression. METHODS: An age-, sex-, H. pylori status- and disease-matched case-control study was performed in 574 H. pylori positive and 225 H. pylori-negative patients selected from 4125 patients with a diagnosis of benign disease of the stomach. Levels of acute and chronic inflammations, atrophy and intestinal metaplasia scored according to the Sydney System were compared by stomach site and by sex. Two biopsy specimens (antral and corpus gastric mucosa) from patients with benign gastric diseases (142 patients; 72 men, 70 women) were analysed for interleukin-8, cyclooxygenase-2 and TFF1 mRNA expression as measured by real-time PCR. RESULTS: Inflammation and activity scores in antrum with H. pylori infection were higher in men, but scores declined according to age. Atrophy and intestinal metaplasia scores in corpus with H. pylori infection appeared more severe in men than in women, especially in older patients. In women, atrophy score increased with increasing age, particularly in postmenopausal H. pylori-negative patients. Interleukin-8 mRNA induction was detected in both antrum and corpus mucosa in H. pylori infection, but sex differences were not found. Response of cyclooxygenase-2 mRNA expression against H. pylori infection in the mucosa was higher in men than women. In H. pylori negative patients, TFF1 mRNA levels in women were significantly higher than in men, and TFF1 mRNA was significantly lower in positive than negative women. CONCLUSIONS: Sex differences in mucosal responses to H. pylori infection in the stomach may be correlated with sex differences in the incidence of stomach cancer. PMID- 15298602 TI - Improvement in serum pepsinogens and gastrin in long-term monitoring after eradication of Helicobacter pylori: comparison with H. pylori-negative patients. AB - BACKGROUND: A decrease in pepsinogen and gastrin levels 1-3 months after Helicobacter pylori eradication is well known. However, few data are available on the long-term progression of these decreases beyond 1 year after eradication, and there has been no investigation into whether pepsinogen and gastrin levels return to normal levels as defined by data from H. pylori-negative patients with dyspepsia. AIM: We studied the effect of H. pylori eradication on pepsinogen and gastrin levels for more than 1 year, and compared levels to those in H. pylori negative patients with dyspepsia. We also investigated the effect of H. pylori eradication on the course of atrophic corpus gastritis as reflected by histology, and on PGI levels and PG I/II ratio. METHODS: We enrolled 172 H. pylori-positive patients with dyspepsia who had undergone successful eradication therapy of more than 1 year's duration and 101 non-treated H. pylori-negative patients with dyspepsia. H. pylori status was assessed at entry and at each endoscopy after eradication by culture, histological results, the rapid urease test and the urea breath test. In both groups, patients were evaluated for fasting serum pepsinogen I and II and gastrin using a radioimmunoassay technique, and underwent detailed histological assessment according to the updated Sydney System. RESULTS: In the H. pylori-negative patients, mean serum pepsinogen I and II, I/II ratio and gastrin levels were 52.6 +/- 20.8 ng/mL, 9.2 +/- 4.2 ng/mL, 6.0 +/- 1.7 and 53.5 +/- 29.2 pg/mL, respectively. In H. pylori-positive patients with long-term eradication, pepsinogen I and II, I/II ratio and gastrin levels were 81.3 +/- 46.6 ng/mL, 25.9 +/- 17.1 ng/mL, 3.4 +/- 1.3 and 131.9 +/- 130.8 pg/mL, respectively, before treatment. At 1-3 months after eradication, serum pepsinogen I and II levels in the H. pylori-positive patients decreased to levels similar to those in the negative patients, whereas pepsinogen I/II ratio and gastrin levels remained lower and higher, respectively, than in the negative patients. Serum pepsinogen I/II ratio and gastrin levels then became similar between the groups at 12-15 months after eradication. In histological findings, inflammation and neutrophil activity decreased by 1-3 months, and atrophy in the corpus and metaplasia in the antrum decreased by 12-15 months. CONCLUSION: The results suggest that atrophic corpus gastritis and superficial gastritis are reversible, as indicated by both histological and serological findings in a long-term follow up study. PMID- 15298603 TI - Diagnostic value of culture, histology and PCR for Helicobacter pylori in the remnant stomach after surgery. AB - BACKGROUND: Diagnosis of Helicobacter pylori infection in the remnant stomach has not been established. AIMS: To investigate the diagnostic value of culture, histology, PCR and serum IgG against H. pylori (ELISA) with and without eradication therapy in the remnant stomach, compared with the unoperated stomach. METHODS: Biopsy samples for bacterial culture and histological diagnosis of H. pylori were taken from the stoma and upper corpus of the remnant stomach and gastric juice was used for PCR assay. RESULTS: Bacterial culture-based diagnosis in the remnant stomach, sensitivity and specificity of culture were 95.1%, 100%; histology 89%, 92.3%; PCR 66%, 89.7%; and ELISA 100%, 50%, respectively, in cases without H. pylori eradication therapy. In assessment of the results of therapy for the remnant stomach, sensitivity and specificity of culture were 100%, 100%; histology 80%, 96.8%; PCR 80%, 91.7%; and ELISA 100%, 0%, respectively. CONCLUSION: Bacterial culture had the highest diagnostic value in the remnant stomach as well as unoperated stomach. Sensitivity by histology and PCR was lower in the remnant stomach than the unoperated stomach, but specificity values were equal. Serum ELISA assay was not suitable for the remnant stomach. PMID- 15298604 TI - DNA typing for Helicobacter pylori isolates from eradication-failed patients: comparison of the isolates before and after therapy. AB - BACKGROUND: Failure of Helicobacter pylori eradication occurs frequently despite use of multiple microbial agents. AIM: We aimed to study differences between H. pylori strains isolated before and after eradication failure. METHODS: We treated 87 patients with peptic ulcer using triple therapy consisting of omeprazole plus combinations of clarithromycin, amoxicillin, or metronidazole. We studied the status of cagA, vacA, and iceA by PCR, and examined the differences in H. pylori isolates by pulsed-field gel electrophoresis and arbitrary primer polymerase chain reaction. The minimum inhibitory concentration of clarithromycin, amoxicillin, or metronidazole was determined by an agar dilution method. RESULTS: Eradication therapy failed in 12 patients (14%); H. pylori isolates were obtained from all of these both before and after therapy. After eradication therapy, 10 patients were colonized with the same strain as before therapy, while the other two patients were colonized with different strains from those before therapy. In the former group, one isolate changed from metronidazole-sensitive to -resistant, one changed from clarithromycin- and metronidazole-sensitive to -resistant, and four were resistant to clarithromycin or metronidazole both before and after therapy. The other four isolates remained sensitive to clarithromycin and metronidazole after therapy. In the two patients who yielded apparently different isolates after therapy, they changed from clarithromycin- and metronidazole sensitive to -resistant. CONCLUSION: Eradication of H. pylori by first-line therapy is an important goal in the treatment of H. pylori-positive peptic ulcer, and that appropriate antimicrobial sensitivity testing should be conducted in patients with eradication failure. PMID- 15298605 TI - Hypoacidity combined with high gastric juice nitrite induced by Helicobacter pylori infection is associated with gastric cancer. AB - BACKGROUND: In patients with Helicobacter pylori infection, the concentration of nitrite in gastric juice is elevated. The degree of elevation correlates with that of inflammation and H. pylori density. AIM: The aim of this study was to examine hypoacidity and high nitrite levels related to H. pylori infection in patients with gastric cancer. METHODS: We studied 88 patients with more than one history of endoscopic mucosal resection (EMR) for early gastric cancer and 88 age matched controls. Concentration of nitrite in gastric juice was measured by Griess reaction, and serum pepsinogen levels were measured by RIA. RESULTS: Multiple malignant lesions were found in 20 of the 88 patients. Serum gastrin, gastric juice pH and nitrite levels in patients with gastric cancer were significantly higher and pepsinogen I and pepsinogen I/II significantly lower than in control subjects. Pepsinogen I level and I/II ratio were lower and gastric juice pH was higher in the protruded-type group than in the depressed type group. Pepsinogen I and pepsinogen I/II were lower and gastric juice pH was higher in multiple than in single cases. CONCLUSIONS: Hypoacidity combined with high gastric juice nitrite induced by H. pylori infection is associated with the intestinal type of gastric cancer, especially protruded lesions. PMID- 15298606 TI - Oesophageal adenocarcinoma or gastric cancer with or without eradication of Helicobacter pylori infection in chronic atrophic gastritis patients: a hypothetical opinion from a systematic review. AB - BACKGROUND: As chronic atrophic gastritis is a precancerous condition for gastric cancer and the eradication of Helicobacter pylori infection halts chronic gastritis, eradication of infection may prevent gastric cancer. However, as chronic atrophic gastritis is a risk factor for reflux oesophagitis after eradication of infection, the risk of oesophageal adenocarcinoma may also increase. METHODS: We systematically reviewed papers and estimated the expected annual incidence of oesophageal or gastric cancer with and without eradication of H. pylori infection in patients with chronic atrophic gastritis. RESULTS: The expected annual incidence of gastric cancer in patients with corpus atrophy with persistent infection was at least 5.8-fold higher than that for oesophageal adenocarcinoma after the eradication of infection at all ages. Even for patients with accompanying reflux oesophagitis or Barrett's oesophagus, the expected incidence of either gastric or oesophageal adenocarcinoma with persistent infection was higher than that of oesophageal adenocarcinoma after eradication of infection. CONCLUSION: If eradication of infection lowers the incidence of gastric cancer, it should be recommended for patients with corpus atrophy at all ages irrespective of the presence of reflux oesophagitis or Barrett's oesophagus, especially in populations having a high prevalence of gastric cancer. PMID- 15298607 TI - High prevalence rate of Helicobacter pylori resistance to clarithromycin during long-term multiple antibiotic therapy for chronic respiratory disease caused by non-tuberculous mycobacteria. AB - BACKGROUND: Helicobacter pylori resistance to clarithromycin, probably due to the frequent use of this antibiotic for the treatment of other diseases, is the greatest obstacle against its eradication. AIM: To clarify the prevalence of clarithromycin-resistant H. pylori in patients with non-tuberculous mycobacterial lung disease receiving multiple antibiotic treatment, including clarithromycin. METHODS: We enrolled 88 patients with non-tuberculous mycobacterial lung disease; 29 underwent upper gastrointestinal endoscopy for the diagnosis of H. pylori infection prior to treatment, and 60 underwent it during treatment. The diagnosis of H. pylori infection was confirmed by histological examination, urease test and microaerobic bacterial culture. The minimum inhibitory concentration of clarithromycin was determined and the DNA was analysed for each of the isolated H. pylori strains. RESULTS: Patients during the treatment had a high prevalence rate of clarithromycin-resistant H. pylori (100%). Analysis of DNA of the clarithromycin-resistant H. pylori isolates revealed point mutations at A2142G or A2143G. Moreover, a linear correlation was found between the total cumulative dose of clarithromycin and the minimum inhibitory concentration. CONCLUSION: All patients with non-tuberculous mycobacterial lung disease being treated long-term with multiple antibiotics, including clarithromycin, harboured clarithromycin resistant H. pylori in the stomach. Therefore, eradication of H. pylori before commencement of long-term therapy including clarithromycin should be recommended. PMID- 15298608 TI - Second-line treatment of Helicobacter pylori infection after dilution agar methods and PCR-RFLP analysis. AB - BACKGROUND: After unsuccessful first-line treatment of Helicobacter pylori infection, the percentage of clarithromycin-resistant strains has been reported as between 30% and 70% in Japan and other countries. A high prevalence of clarithromycin-resistant strains is reported to be associated with eradication failure. AIM: We examined antibiotic susceptibility testing using a combination of dilution agar methods with PCR-restriction fragment length polymorphism (RFLP) analysis. METHODS: We enrolled 41 patients in whom first-line treatment with LAC (lansoprazole, amoxycillin and clarithromycin) was unsuccessful. Endoscopic biopsied specimens were used to examine antibiotic susceptibility to clarithromycin by dilution agar methods. PCR-RFLP analysis was performed to determine the presence of point mutations, which are primarily responsible for resistance to clarithromycin. RESULTS: Clarithromycin-resistance rate after failure of the LAC regimen was 73.2%. Drug susceptibilities of three strains obtained by PCR-RFLP analysis were different from those by dilution agar methods. One strain with MIC values to clarithromycin of 0.05 micro g/mL had a point mutation, A2144G. This strain was not eradicated by repeating LAC, but was eradicated by substituting metronidazole for clarithromycin. CONCLUSIONS: Dilution agar methods should be combined with PCR-RFLP analysis before second line eradication to increase the accuracy of clarithromycin-susceptibility testing and to improve eradication efficacy. PMID- 15298609 TI - Helicobacter pylori eradication decreases blood neutrophil and monocyte counts. AB - BACKGROUND: The effect of Helicobacter pylori infection on systemic disorders is not well understood. AIM: The purpose of this study was to elucidate the systemic effects of H. pylori infection by comparing differential counts of leukocytes and platelets in peripheral blood before and after eradication of H. pylori. METHODS: A total of 164 H. pylori-positive patients underwent eradication therapy, and populations of peripheral blood leukocytes and platelets before and 0 (just after therapy), 1, 3 and 12 months after eradication were retrospectively analysed. RESULTS: In the eradicated group (n = 138), blood leukocytes, neutrophils and monocytes decreased significantly after eradication, but there was no significant change in eosinophils, basophils, lymphocytes or platelets. In the non-eradicated group (n = 26), there was no significant change in any studied parameter. With regard to smoking status, although leukocytes and neutrophils did not decrease after eradication in the smoking group, they significantly decreased after eradication in the nonsmoking group. CONCLUSIONS: These findings suggest that: (1) H. pylori infection increases neutrophil and monocyte counts in the peripheral blood, which indicates a significant role of H. pylori infection in systemic disorders; and (2) Smoking may mask the effect of H. pylori eradication on peripheral leukocytes, which would explain the controversy in previous reports concerning H. pylori infection and peripheral leukocytes. PMID- 15298610 TI - Helicobacter pylori-positive patients with pruritic skin diseases are at increased risk for gastric cancer. AB - BACKGROUND: Helicobacter pylori infection has been implicated as a possible cause of extraintestinal disorders such as skin diseases. A number of case reports describe patients with skin diseases, such as prurigo nodularis, that are associated with gastric cancer. AIM: The aim of this study was to examine the prevalence of H. pylori infection and the incidence of gastric cancer in patients with pruritic skin diseases. METHODS: The patients were examined for circulating specific IgG antibodies against H. pylori in sera using ELISA. H. pylori-positive patients who were more than 40 years old underwent endoscopic screening for gastric cancer. RESULTS: We examined 134 patients with pruritic skin diseases, including 55 cases of cutaneous pruritus, 21 cases of prurigo chronica multiforme, 15 cases of nummular dermatitis and 43 cases of chronic urticaria. Early gastric cancer was detected in 2/36 (5.6%) patients with cutaneous pruritus and 3/16 (18.8%) with prurigo chronica multiforme. The prevalence of early gastric cancer was 5.6%, which was much higher than that among patients undergoing general endoscopic screening for gastric cancer. CONCLUSIONS: These results suggest that H. pylori-positive patients with pruritic skin diseases may be at increased risk for development of gastric cancer, and endoscopic screening in such patients is recommended. PMID- 15298611 TI - Relationship between Helicobacter pylori infection and the prevalence, site and histological type of gastric cancer. AB - AIM: To evaluate the relationship between Helicobacter pylori infection and gastric cancer in Japan. METHODS: This was a multicentre study conducted in various regions in Japan. A total of 6578 individuals as controls and 2503 with histologically confirmed gastric cancer were enrolled. H. pylori status was determined by enzyme-linked immunosorbent assay (ELISA) for anti-H. pylori immunoglobulin G (IgG). RESULTS: The prevalence of H. pylori infection in gastric cancer patients was markedly high in all age groups. In contrast, the rate increased with age among control subjects. The overall prevalence of H. pylori infection in control subjects was 50.2% (3300/6578) vs. 82.8% in gastric cancer patients (2072/2503) (OR = 2.47; 95% CI: 2.19-2.79). The prevalence of H. pylori in early gastric cancer was significantly higher than that in advanced gastric cancer (86.5% vs. 75.7%; OR = 2.06; 95% CI: 1.66-2.55). There were no significant differences in the prevalence of H. pylori between the intestinal and diffuse types of gastric cancer or among gastric cancer in antrum, body and cardia. CONCLUSION: H. pylori infection is strongly associated with the development of gastric cancer. The difference in odds ratios among younger and older persons with gastric cancer likely reflects the decrease in prevalence of H. pylori in the population, and is more reflective of the actual risks associated with the infection. PMID- 15298612 TI - Helicobacter bilis infection in biliary tract cancer. AB - BACKGROUND: Biliary tract cancer is a highly fatal disease with poor prognosis, but the aetiology is poorly understood. AIM: We aimed to identify Helicobacter bilis infection in the gallbladder in patients with biliary tract disease. METHODS: Archival gallbladder specimens from 34 patients (14 males and 20 females) with an average age of 61.4 +/- 12.2 years (mean +/- SE) were retrieved, consisting of 11 cases of gallbladder cancer, three of bile duct cancer, 16 of cholecystolithiasis and four of pancreatic cancer. DNA was extracted and nested PCR using primers specific for 16S rRNA of H. bilis was performed. RESULTS: Amplification was observed in 3 of 11 gallbladder cancer cases (27.2%) and one of three cases with biliary duct cancer (33.3%). In total, four of 14 cases with biliary tract cancer were positive for H. bilis (28.6%). In addition, the presence of H. bilis was shown in two of 16 cases (12.5%) with cholecystolithiasis. Notably, although the number of cases examined was small, none of the four cases with pancreatic cancer showed the presence of H. bilis infection in the gallbladder without apparent abnormalities. CONCLUSION: H. bilis infection may play a role in biliary tract disease, particularly in biliary tract cancer. PMID- 15298613 TI - Anti-tumour effects of nobiletin, a citrus flavonoid, on gastric cancer include: antiproliferative effects, induction of apoptosis and cell cycle deregulation. AB - AIM: To demonstrate the antitumour effects of nobiletin (5,6,7,8,3',4' hexamethoxyflavone), a citrus flavonoid extracted from Citrus depressa Hayata, on human gastric cancer cell lines TMK-1, MKN-45, MKN-74 and KATO-III. MATERIALS AND METHODS: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, the TdT-mediated dUTP biotin nick-end labelling (TUNEL) method and cell cycle analysis revealed that nobiletin acted on these cells in several ways, namely by direct cytotoxicity, induction of apoptosis and modulation of cell cycle. The efficacy of combined treatment of nobiletin with a conventional anticancer drug, CDDP, was also examined. Treatment with nobiletin 24 h prior to CDDP administration showed a synergistic effect compared to the control. CONCLUSIONS: Although the effective dose and administration route of nobiletin require further investigation, our study represents a potential successful linking of this compound with the treatment of gastric cancer. PMID- 15298614 TI - Evaluation of treatment for gastro-oesophageal reflux disease with a proton pump inhibitor, and relationship between gastro-oesophageal reflux disease and Helicobacter pylori infection in Japan. AB - BACKGROUND: Effective therapy for gastro-oesophageal reflux disease (GERD) is associated with improvement in health-related quality of life. It remains unclear whether Helicobacter pylori infection protects against GERD. AIM: We evaluated the relationship between GERD and H. pylori, and whether the health-related quality of life score improved after medical treatment. METHODS: We enrolled 151 outpatients with upper abdominal symptoms; 81 patients received omeprazole 20 mg/day for 2 weeks. Health-related quality of life was assessed using the Gastrointestinal Symptom Rating Scale (GSRS) and the Psychological General Well Being (PGWB) index. H. pylori infection was diagnosed by serum antibody or endoscopy and the relationship between GERD and H. pylori was evaluated. RESULTS: In GERD patients, the mean GSRS score improved from 2.20 to 1.67 following treatment (P < 0.01). The mean GSRS reflux symptom score improved from 2.96 to 1.67 (P < 0.01). The mean PGWB score improved from 96.36 to 107.34 (P < 0.01). All scores in GERD patients significantly improved compared with non-GERD patients. The H. pylori-positive ratio was 66.15% in GERD patients and 65.21% in non-GERD patients (P = 0.94). CONCLUSIONS: Health-related quality of life is useful for evaluation of proton pump inhibitor treatment in GERD. The presence of H. pylori was not associated with the prevalence of GERD. PMID- 15298615 TI - Prevalence of symptomatic gastro-oesophageal reflux disease in Japanese patients with peptic ulcer disease after eradication of Helicobacter pylori infection. AB - BACKGROUND: The association between cure of Helicobacter pylori infection and the development of gastro-oesophageal reflux disease is controversial. AIM: To examine the prevalence of symptomatic GERD (sGERD) in Japanese patients with peptic ulcer disease after successful eradication and identify associated factors affecting sGERD development. METHODS: We retrospectively examined 72 patients (40 gastric ulcer and 32 duodenal ulcer) with successful eradication. Associated factors such as age, gender, drinking and smoking habits, body mass index, presence of gastric atrophy and hiatal hernia were analysed. RESULTS: Seven (9.7%) of 72 peptic ulcer patients newly developed sGERD. There were no differences in mean age, gender, smoking habit, drinking habit, body mass index, or presence of gastric atrophy and hiatal hernia between the sGERD and non-sGERD groups, while the proportion of subjects aged over 70 was significantly higher in the sGERD than the non-sGERD group. Six of 40 patients with gastric ulcer newly developed sGERD while only one of 32 patients with duodenal ulcer developed it. CONCLUSION: Approximately 10% of Japanese patients with peptic ulcer disease newly developed sGERD after cure of H. pylori infection. Age > 70 years was associated with development of sGERD. Eradication in patients in this age group should be carefully determined. PMID- 15298616 TI - Oesophageal hypersensitivity in Japanese patients with non-erosive gastro oesophageal reflux diseases. AB - BACKGROUND: Visceral hypersensitivity plays a major role in the pathogenesis of non-erosive oesophageal reflux disease (NERD). Prevalence of NERD differs according to the population and geographical region. Oesophageal hypersensitivity in NERD has not been well studied, especially in Japanese patients. AIM: To investigate oesophageal hypersensitivity in Japanese NERD patients. PATIENTS AND METHODS: We performed upper GI endoscopy and the modified acid perfusion test on 14 control subjects and 68 GERD patients, including 26 with NERD, 34 with erosive GERD, and six with Barrett's oesophagus. The stimulus-response function to acid was quantified by three parameters (lag time, intensity rating and the acid perfusion sensory score) and compared among four groups. RESULTS: The mean value of the lag time, intensity rating, and acid perfusion scores in NERD patients (4.6 +/- 3.4, 4.4 +/- 3.4, 27.8 +/- 26.7, respectively) were higher than in erosive GERD (3.2 +/- 3.3, 3.0 +/- 3.2, 18.2 +/- 24.8) and Barrett patients (2.5 +/- 4.0, 1.8 +/- 3.3, 15.0 +/- 28.8), and significantly higher than in the control group (1.7 +/- 2.7, 1.1 +/- 2.0, 5.4 +/- 11.8). The ratio of patients with higher sensory scores was also greater in the NERD group (57.7%) than in erosive GERD (32.3%) and Barrett group (16.7%), and significantly greater than in control group (6.7%). CONCLUSION: Our findings suggest that oesophageal sensitivity is likely to be enhanced especially in NERD patients also in Japanese population in comparison with erosive GERD, Barrett's oesophagus and controls. PMID- 15298617 TI - Impairment of gastrointestinal motility by nitrate administration: evaluation based on electrogastrographic changes and autonomic nerve activity. AB - BACKGROUND: Nitrates decrease the tone of the lower oesophageal sphincter, and may thus induce gastro-oesophageal reflux. AIM: In the present study, we evaluated electrogastrographic changes and heart-rate variability before and after the administration of nitrates. METHODS: In 15 patients with chest pain treated with nitrates, electrocardiography and percutaneous electrogastrography were performed before and after administration of nitrates. Autonomic nervous system function was evaluated by spectral analysis of heart-rate variability and serial changes in low frequency and high frequency power, and the low frequency/high frequency ratio were compared. Electrogastrograms were analysed by obtaining peak power amplitudes and their dominant frequencies. RESULTS: After the administration of nitrates (isosorbide dinitrate), high frequency power, an index of parasympathetic nervous activity, was significantly decreased, whereas the low frequency/high frequency ratio, an index of sympathetic nervous activity, was significantly increased. The mean peak amplitude of the electrogastrogram significantly increased postprandially both before and after treatment. After isosorbide dinitrate treatment, however, mean peak amplitudes after a meal were significantly lower than those obtained before treatment. The mean dominant frequency of the electrogastrogram did not vary before and after treatment. CONCLUSIONS: The present study suggests that nitrates inhibit gastrointestinal motility by decreasing autonomic nervous activity. PMID- 15298618 TI - Expression of leptin in two-layered culture of gastric mucous cells and fibroblasts: effect of Helicobacter pylori attachment. AB - BACKGROUND: Our recent histochemical studies have revealed an increase in myofibroblasts and in leptin and its receptor in endothelial cells, and myofibroblasts in Helicobacter pylori-infected human and Mongolian gerbil fundic mucosa. AIM: The present study was undertaken to clarify the H. pylori-induced interaction between leptin in cultured gastric surface mucous cells and fibroblasts. METHODS: GSM06 cells were incubated with an air- liquid interface on a collagen gel layer containing mouse fibroblast cell line L929. Medium containing H. pylori bacilli (ATCC43504) at 10-100 times higher concentration than the GSM06 cells was added from the luminal side and the localization of leptin was observed by immunohistochemistry. The transformation of L929 cells to myofibroblasts was detected by electron microscopy and PR 2D3 immunoreactivity. RESULTS: L929 cells in the control group showed a spindle shape with scarce cytoplasm. In the H. pylori-treated group, L929 cells showed features characteristic of myofibroblasts, and most GSM06 and L929 cells showed leptin immunoreactivity. In contrast, L929 cells incubated with H. pylori alone did not undergo this differentiation. CONCLUSIONS: Attachment of H. pylori to surface epithelial cells caused conversion of fibroblasts to myofibroblasts. We suggest that leptin plays a role in this transformation. PMID- 15298619 TI - DNA hypermethylation in gastric cancer. AB - BACKGROUND: Transcriptional silencing of tumour suppressor genes by DNA hypermethylation plays a crucial role in the progression of gastric cancer. Many genes involved in the regulation of cell cycle, tissue invasion, DNA repair and apoptosis have been shown to be inactivated by this type of epigenetic mechanism. RESULTS: Recent studies have demonstrated that DNA hypermethylation begins early in cancer progression, and in some cases, may precede the neoplastic process. Ageing is associated with DNA hypermethylation, and may provide a mechanistic link between ageing and cancer. Several reports have indicated that Epstein-Barr virus-related gastric cancer is associated with a high frequency of DNA hypermethylation, suggesting that viral oncogenesis might involve DNA hypermethylation with inactivation of tumour suppressor genes. Hypermethylation of hMLH1 with the resulting loss of its expression is known to cause microsatellite instability, which reflects genomic instability associated with defective DNA mismatch repair genes in the tumour. CONCLUSIONS: In conclusion, recent studies demonstrate that DNA hypermethylation is a crucial mechanism of inactivation of tumour suppressor genes in gastric cancer. A better understanding of DNA hypermethylation will provide us with new opportunities in the diagnosis and therapy of gastric cancer. PMID- 15298620 TI - Correlation of MAP kinases with COX-2 induction differs between MKN45 and HT29 cells. AB - BACKGROUND: Mitogen-activated protein (MAP) kinases, including extracellular signal-regulated kinases (ERK),c-Jun NH2-terminal kinases (JNK) and p38 MAP kinase (p38 MAPK) are important intermediates of the signal-transduction pathway from the cell surface to the nucleus. Expression of cyclooxygenase (COX)-2, associated with proliferation, apoptosis or both of gastrointestinal cancer cells, is mediated through MAP kinase families. However, the correlation between respective MAP kinase signals and COX-2 in the proliferation of gastric and colon cancer cells has not been well elucidated. AIM: We examined the effect of selective inhibitors of MAP kinases and COX-2 on serum-induced proliferation of gastric (MKN45) and colon (HT29) cancer cells. METHODS: After 24-h serum starvation, cancer cells were stimulated with 2% serum and COX-2 inhibitors (NS398 10 micromol/L, or etodolac 100 micromol/L) or 1 h after preincubation with inhibitors for ERK (PD98059 20 micromol/L) or p38 MAPK (SB203580 10 micromol/L). Phosphorylated MAP kinases and COX-2 protein were evaluated by Western blotting, and the proliferation of cancer cells was estimated by 3H-thymidine incorporation. Transcription factors nuclear factor-kappaB and CREB were assayed by an electorophoretic mobility shift assay. RESULTS: Serum increased the proliferation of MKN45 and HT29 cells by 280% and 200%, respectively, compared with the control levels (100%). In both cancer cells, phosphorylated MAP kinases were increased within 30 min after stimulation. PD98059 and SB203580 inhibited the serum-induced proliferation of MKN45 by 21% and 51% and of HT29 by 81% and 69%, respectively. NS398 and etodolac inhibited the proliferation of HT29 by 21% and 41%, respectively, but not that of MKN45. PD98059 and SB203580 also suppressed serum-induced expression of COX-2 protein in HT29 cells. In addition to the activation of MAP kinases and COX-2, activities of nuclear factor-kappaB and CREB were also increased during HT29 cell proliferation. CONCLUSIONS: These results suggest that the correlation of MAP kinases with COX-2 induction for cell proliferation differs between MKN45 and HT29 cells. PMID- 15298621 TI - Elemental diet modulates the growth of Clostridium difficile in the gut flora. AB - BACKGROUND AND AIMS: Tube feeding is regarded as a risk factor for Clostridium difficile-associated diarrhoea. Recently, we reported that C. difficile toxin was frequently found in patients receiving an elemental diet. The present study was conducted to clarify whether elemental diets are associated with the growth of C. difficile in the gut flora. METHODS: C. difficile was cultured for 72 h in various concentrations of elemental diet containing 3% thioglycollate, and the growth rate or activity of C. difficile was evaluated by Gram stain or by measuring optical density at 560 nm. Faecal samples from 10 healthy adults were cultured in elemental diet + 3% thioglycollate. RNA was extracted from faeces with glass powder, which can eliminate PCR inhibitors, and mRNA of C. difficile toxin B was measured by reverse transcription PCR. RESULTS: Maximum OD560 value during culture in thioglycollate-containing elemental diet was 2.4 times higher than that in thioglycollate alone (P = 0.0163). Viability of C. difficile was decreased in thioglycollate but not in thioglycollate-containing elemental diet. Toxin B mRNA was detected in five faecal samples (50%) before culture and in all samples after culture. CONCLUSIONS: Our results suggest that an elemental diet can modulate the growth of C. difficile in the gut flora. PMID- 15298622 TI - Helicobacter pylori eradication treatment modulates epithelial cell proliferation and tissue content of hepatocyte growth factor in the gastric mucosa. AB - BACKGROUND: Although Helicobacter pylori infection is now acknowledged as a major promoter of gastric cancer in humans, the carcinogenetic process of this effect has not been fully elucidated. Precancerous lesions such as intestinal metaplasia, enhanced proliferation of epithelial cells and elevated level of growth factors have been postulated to play a role. AIM: To analyse a relationship between gastric mucosal proliferation, mucosal content of hepatocyte growth factor and prevalence of intestinal metaplasia before and after successful H. pylori eradication therapy. METHODS: We evaluated 25 H. pylori-eradicated patients. At initial endoscopic examination, two biopsy tissue samples each were obtained from the antrum and great curvature of the corpus. Tissue content of hepatocyte growth factor and neutrophil myeloperoxidase were measured using an ELISA method, and histological assessment of intestinal metaplasia (haematoxylin and eosin) and proliferating cells (Ki-67 immunostaining) was performed. The patients were treated with a 1-week course of triple therapy. At 10 months after successful eradication, biochemical and histological assessments were repeated. RESULTS: Among all patients (n = 25), no intestinal metaplasia was detected in the corpus mucosa, but was observed in 10 patients (40%) in the antrum. This prevalence ratio was not changed after eradication. A slight decrease in HGF content was demonstrated in both sites, but the level of antral hepatocyte growth factor was significantly decreased in patients with intestinal metaplasia but not in those without. Proliferative index (Ki-67 positive cells/epithelium) was decreased after eradication therapy in both sites. An increase in proliferative index was observed in the antrum with intestinal metaplasia compared with that without, which significantly decreased after eradication therapy. CONCLUSIONS: H. pylori eradication therapy in the present study afforded an inhibitory effect on epithelial cell proliferation and on the mucosal content of HGF. PMID- 15298623 TI - Histological aspects and role of mast cells in Helicobacter pylori-infected gastritis. AB - BACKGROUND: Mast cells are one of the main pro-inflammatory cells, while their knowledge in Helicobacter pylori infection has not been summarized. METHODS: We reviewed studies on mast cells in H. pylori infection, and summarized the histological aspects and roles. RESULTS: The density of mast cells is greater in H. pylori-infected than in non-infected subjects. Increased mast cell density in infected gastritis significantly decreases after eradication. On electron microscopy, mast cells in infected gastric mucosa show degranulation. Some experimental studies demonstrate that mast cells are degranulated with H. pylori derived products. CONCLUSIONS: Mast cells are actively involved in the pathogenesis of H. pylori-infected gastritis. The possible roles are to initiate and promote the formation of oedema through degranulated and secreted mediators, and to release multiple chemotactic factors, which induce inflammatory cells to infiltrate to the site of oedema, showing acute inflammatory changes. Mast cells also stimulate the degradation of pericellular matrices and the growth of cells in their vicinity, and thereby promote tissue turnover. In chronic H. pylori infection, these reactions continue until the bacteria are eradicated. Mast cells may act both to maintain gastritis and to repair tissue damage in H. pylori infected chronic gastritis. PMID- 15298624 TI - Indometacin up-regulates TFF2 expression in gastric epithelial cells. AB - BACKGROUND: Trefoil factor family peptides are expressed in gastrointestinal epithelial cells and play a critical role in maintaining mucosal integrity. Although non-steroidal anti-inflammatory drugs (NSAIDs) are important causative agents of gastric mucosal lesions, few data are available about the effect of NSAIDs on trefoil family peptides in gastric mucosa. AIM: To examine whether indometacin, a widely used NSAID, affects trefoil factor family expression in gastric epithelial cells. METHODS: MKN45, a cell line derived from human gastric cancer, was used. TFF1, TFF2, and TFF3 mRNA expression was assessed by real-time quantitative reverse transcription-polymerase chain reaction (RT-PCR). TFF2 gene transcription was also examined by luciferase reporter gene assay. RESULTS: Relative expression level of TFF1, TFF2, TFF3 mRNA was 616: 12: 1 in unstimulated MKN45 cells. Although indometacin (1-250 micro mol/L) had no significant effect on the expression of TFF1 and TFF3 mRNA, it up-regulated TFF2 mRNA expression in a dose- and time-dependent manner. Luciferase reporter gene assay confirmed the up-regulation of TFF2 gene transcription by indometacin. Indometacin-induced up regulation of TFF2 expression was not antagonized by externally applied prostaglandin E2. CONCLUSION: These results suggest that indometacin up-regulates gastric epithelial cell TFF2 expression through a COX-independent mechanism. Since TFF peptides play an important role in gastric mucosal protection, indometacin-induced TFF2 may reduce the degree of gastric mucosal damage induced by indometacin. PMID- 15298625 TI - Heme oxygenase-1: a new therapeutic target for inflammatory bowel disease. AB - Heme oxygenase (HO) is the rate-limiting enzyme in the catabolism of heme, followed by production of biliverdin, free iron and carbon monoxide (CO). Three mammalian HO isozymes have been identified, one of which, HO-1, is a stress responsive protein induced by various oxidative agents. HO-2 and HO-3 genes are constitutively expressed. Recent studies demonstrate that the expression of HO-1 in response to different inflammatory mediators may contribute to the resolution of inflammation and have protective effects in several organs against oxidative injury. Although the mechanism underlying the anti-inflammatory actions of HO-1 remains poorly defined, both CO and biliverdin/bilirubin have been implicated in this response. In the intestinal tract, HO-1 is shown to be transcriptionally induced in response to oxidative stress, preconditioning and acute inflammation. Recent studies suggest that the induction of HO-1 expression plays a critical protective role in intestinal damage models induced by trinitrobenzene sulphonic acid or dextran sulphate sodium, indicating that activation of HO-1 may act as an endogenous defensive mechanism to reduce inflammation and tissue injury in the intestinal tract. These in vitro and in vivo data suggest that HO-1 may be a novel therapeutic target in patients with inflammatory bowel disease. PMID- 15298626 TI - Effect of dietary anti-Helicobacter pylori-urease immunoglobulin Y on Helicobacter pylori infection. AB - Recently, chicken egg yolk was recognized as an inexpensive antibody source, and the therapeutic usefulness of egg yolk immunoglobulin Y (IgY) in oral passive immunization has been investigated. Although multiple antibiotic treatments eradicate most Helicobacter pylori (H. pylori) infections, therapy fails in 10 15% of cases due to the development of drug resistance. Consequently, it is important that new, more broadly based therapies for the treatment of H. pylori infection should be identified. The present study evaluated the effect, on H. pylori infection, of IgY prepared from egg yolk of hens immunized with H. pylori urease (anti-HpU IgY). Seventeen asymptomatic volunteers diagnosed as H. pylori positive by the 13C-urea breath test (UBT) were orally administered anti-HpU IgY for 4 weeks. Four weeks later, UBT values were significantly decreased although no case showed H. pylori eradication. An H. pylori-positive 53-year-old female gastritis patient administered anti-HpU IgY plus lansoprazole for 8 weeks showed a decrease in serum pepsinogen (PG) I and UBT values as well as an increase in the PG I/II ratio. In conclusion, anti-HpU IgY may mitigate H. pylori-associated gastritis and partially attenuate gastric urease activity. Furthermore, anti-HpU IgY combined with antacids appears to ameliorate gastric inflammation. These encouraging results may represent a novel approach to the management of H. pylori associated gastroduodenal disease. PMID- 15298630 TI - Systematic review: the hepatotoxicity of non-steroidal anti-inflammatory drugs. AB - BACKGROUND: Non-steroidal anti-inflammatory drugs have been implicated in reports of liver injury. However, the precise risk of non-steroidal anti-inflammatory drugs for this rare complication is unknown. AIM: To review systematically the published literature of population-based epidemiological studies reporting the incidence or comparative risk of non-steroidal anti-inflammatory drugs for liver injury resulting in clinically significant events, defined as hospitalization or death. DATA EXTRACTION: Duplicate extraction of the methodological quality, design, source, population, years studied, particular non-steroidal anti inflammatory drugs studied, definitions, patient counts and follow-up, and the adjustment for confounders. RESULTS: Seven articles met inclusion criteria. The comparative risk of liver injury resulting in hospitalization for current non steroidal anti-inflammatory drug users compared with past non-steroidal anti inflammatory drug users ranged from 1.2 to 1.7, but none was statistically significant. The incidence of liver injury resulting in hospitalization ranged from 3.1 to 23.4/100,000 patient-years of current use of non-steroidal anti inflammatory drugs, with an excess risk compared with past non-steroidal anti inflammatory drugs users of 4.8-8.6/100,000 patient-years of exposure. There were zero deaths from liver injury associated with non-steroidal anti-inflammatory drugs use in over 396,392 patient-years of cumulative exposure. CONCLUSION: These findings allow for the possibility of a small increase in the risk of clinically relevant hepatotoxicity with non-steroidal anti-inflammatory drugs use, but do not document that such a risk occurs. PMID- 15298631 TI - Personal view: 'don't ask, don't tell'--the undesirable consequences of incidental test results in gastroenterology. AB - A gastroenterologist is frequently approached to perform an endoscopic procedure after an incidental test has resulted in an unexpected positive finding. Should the incidental test result be ignored or automatically followed by an endoscopic procedure? The present analysis strives to characterize the common pattern of such scenarios and resolve their underlying dilemma. A model of game theory is used as a mathematical tool to develop general management strategies. Three clinical scenarios are used as examples to demonstrate this approach. The model is based on how doctors rank the various outcomes with which they are confronted by the incidental test results. The ranks of different outcomes are listed in a decision matrix that is converted into a two-by-two, non-zero, ordinal game. All scenarios of incidental test results emerge as a similar type of game that is best played by both parties adhering to the same set of strategies: 'Don't do the test' and 'don't respond to whatever test result it yields'. These two strategies lead to a Nash equilibrium for a non-zero game, where neither party can improve its payoff any further by choosing a different strategy. Although the equilibrium does not provide the individual players with their best possible payoff, it yields the best overall outcome available to both parties given the economic and medical constraints of the situation. PMID- 15298632 TI - Personal view: rationale and proposed algorithms for symptom-based proton pump inhibitor therapy for gastro-oesophageal reflux disease. AB - Gastro-oesophageal reflux disease and non-erosive reflux disease are chronic, highly prevalent conditions requiring long-term treatment that is both effective and practical. On-demand therapy with a proton pump inhibitor may meet that need. It is becoming a mainstay of long-term treatment because it reduces the risk of over- and under-treatment, is cost-effective and user friendly. Epidemiological and clinical observations speak also in its favour. However, for the anticipated benefits of on-demand therapy to accrue in clinical practice, on-demand treatment algorithms are required. These algorithms must specify the initial evaluation and treatment of candidates, and follow-up protocols for an on-demand strategy. Our group has developed such algorithms, which are presented here. PMID- 15298633 TI - Intragastric acid suppression and pharmacokinetics of twice-daily esomeprazole: a randomized, three-way crossover study. AB - BACKGROUND: Patients with refractory gastro-oesophageal reflux disease, extra oesophageal reflux symptoms, Barrett's oesophagus, or Zollinger-Ellison syndrome may require greater acid suppression than that obtained with once-daily esomeprazole. AIM: To assess gastric acid suppression (determined by intragastric pH) and pharmacokinetics of twice-daily vs. once-daily esomeprazole. METHODS: In a randomized, double-blind, three-way crossover study, healthy subjects received esomeprazole 40 mg once daily, 20 mg twice daily, or 40 mg twice daily for five consecutive days. Twenty-four-hour continuous ambulatory intragastric pH was recorded on day 5. RESULTS: Esomeprazole 40 mg twice daily provided a mean of 19.2 h with intragastric pH > 4.0 (80.1% of a 24-h time period; 95% confidence interval 74.5-85.7%) vs. 14.2 h with 40 mg once daily (59.2%; 95% CI 53.7-64.7%) and 17.5 h with 20 mg twice daily (73.0%; 95% confidence interval 67.4-78.5%) in 25 subjects. Intragastric pH was maintained >4.0 for a similar percentage of time during active and sleeping periods for all doses. CONCLUSIONS: Esomeprazole 40 mg twice daily provides significantly greater acid suppression (number of hours in a 24-h period with pH > 4.0) than once-daily dosing and may be a reasonable consideration for patients requiring greater acid suppression for acid-related disease. PMID- 15298634 TI - A novel option in proton pump inhibitor dosing: lansoprazole orally disintegrating tablet dispersed in water and administered via nasogastric tube. AB - BACKGROUND: Lansoprazole orally disintegrating tablet, which rapidly disintegrates on the tongue or in water, provides a dosing alternative for patients with difficulty in swallowing. Gastric and nasogastric tubes are increasingly placed in patients with more severe swallowing disorders. AIM: This study assessed the pharmacokinetic profile of lansoprazole orally disintegrating tablet dispersed in a small volume of water and administered through a small-bore nasogastric tube. METHODS: Forty healthy adult men and women (18-43 years) received two single 30 mg lansoprazole orally disintegrating tablet doses (one administered directly onto the tongue without water, and one dispersed in water and administered via nasogastric tube) in a randomized, crossover fashion. RESULTS: The total plasma exposure to lansoprazole was comparable following both dosing regimens; mean AUC values for the lansoprazole orally disintegrating tablet nasogastric dispersion were < or =8.6% greater than those for the intact lansoprazole orally disintegrating tablet. Lansoprazole Cmax for the lansoprazole orally disintegrating tablet nasogastric dispersion was 20.9% greater than that for the intact lansoprazole orally disintegrating tablet, a difference of no clinical significance. CONCLUSIONS: Dispersion of the lansoprazole orally disintegrating tablet in a small volume of water and administering via nasogastric tube does not reduce the pharmacokinetic profile of the intact lansoprazole orally disintegrating tablet. This alternative dosing method may be useful in patients with nasogastric or gastric tubes. PMID- 15298635 TI - The role of acid suppression in patients with endoscopy-negative reflux disease: the effect of treatment with esomeprazole or omeprazole. AB - BACKGROUND: Patients with endoscopy-negative reflux disease have reflux symptoms, mainly heartburn, but not mucosal breaks characteristic of erosive oesophagitis. Standard-dose proton pump inhibitors can provide symptom relief in endoscopy negative reflux disease but the effect of greater acid suppression has not been studied. AIM: To test the hypothesis that esomeprazole produces heartburn resolution in a greater proportion of patients with ENRD than omeprazole. METHODS: Three multi-centre randomized, controlled, double-blind, 4-week acute treatment studies were conducted in endoscopy-negative reflux disease patients. In study A (n = 1282), patients received either esomeprazole 40 mg, esomeprazole 20 mg or omeprazole 20 mg daily; in studies B (n = 693) and C (n = 670) patients received either esomeprazole 40 mg or omeprazole 20 mg (B), and esomeprazole 20 mg or omeprazole 20 mg (C), respectively. RESULTS: Resolution of heartburn at 4 weeks (no heartburn symptoms during the last 7 days) was achieved in similar proportions of patients in each treatment arm in study A (esomeprazole 40 mg, 56.7%; esomeprazole 20 mg, 60.5%; omeprazole 20 mg, 58.1%), study B (esomeprazole 40 mg, 70.3%; omeprazole 20 mg, 67.9%) and study C (esomeprazole 20 mg, 61.9%; omeprazole 20 mg, 59.6%). There were no significant differences between treatment groups within each study. CONCLUSIONS: More than 60% of endoscopy-negative reflux disease patients reported heartburn resolution but, after 4 weeks of therapy, these proportions did not differ significantly between treatments. PMID- 15298636 TI - Identifying response to acid suppressive therapy in functional dyspepsia using a random starting day trial--is gastro-oesophageal reflux important? AB - BACKGROUND: Single subject trials offer an alternative approach to identify and characterize responders to a specific treatment. AIM: To test a new single subject trials design, called random starting day trial, to identify acid-related symptoms in dyspepsia. METHODS: A total of 119 patients with functional dyspepsia entered a 12-day, double-blind random starting day trial. All patients started on placebo and switched to omeprazole 80 mg/day at a randomized and blinded day between day 5 and day 9, with active treatment continuing for the rest of the trial. Based on changes of a daily symptom score, response was defined as a sustained > or =50% reduction of symptoms within 3 days of active treatment. RESULTS: Thirteen of 119 patients (11%) were classified as spontaneous responders because of complete symptom relief before switching to omeprazole. Of the remaining 106 patients, 15 (15.6%) were classified as responders. Five of six (83%) responders compared with 28 of 53 (53%) non-responders had pathological reflux. Multivariate testing identified symptoms suggestive of gastro-oesophageal reflux predictive of response. CONCLUSIONS: The random starting day trial design could identify a subset of dyspeptic patients with a uniform symptomatic response to acid-suppressive therapy. Response seems to be associated with gastro oesophageal reflux. The random starting day trial needs to be further validated to be considered as a reliable instrument in clinical research. PMID- 15298637 TI - Pneumatic balloon dilatation in achalasia: a prospective comparison of safety and efficacy with different balloon diameters. AB - BACKGROUND: Pneumatic dilatation is considered to be the first line therapy for achalasia, but long-term outcome studies are scarce and limited by their retrospective design. There is also no consensus on the optimal method for performing pneumatic dilation as regard to balloon diameter, amount and the rate inflation pressure. AIM: To address these questions in a large long-term prospective study. METHODS: Over a period of 10 years 262 achalasia patients referred to our centre were enrolled. All patients underwent a pre-treatment clinical evaluation and were followed every 6 months. The first 62 patients (group A) underwent dilatation with initial use of a 35 mm balloon with inflation pressure of 10 psi in 10 seconds (s). In group B (200 patients) we initially used a 30 mm balloon with inflation pressure of 10 psi in 30 s. Dilatation was repeated with incrementally larger balloons (35 and 40 mm) in case of relapse. We used rigiflex balloon and maintained pressure for 60 s after inflation in both groups. RESULTS: Three perforations occurred in group A whereas no perforation took place in Group B. The cumulative proportional remission rate with single dilatation in groups A and B decreased from 83 and 75% in 6 months to 60 and 57% after 30 months of therapy respectively, the differences did not reach statistical significance. In patients who had undergone further dilatations the probability of remaining in remission at 1 year after the first and the second dilatation was 38 and 88% in group A, 20 and 89% in group B respectively. The probability of remaining in remission for 2 years increased from 20% after the first dilatation to 70% after the second dilatation. CONCLUSION: Graded pneumatic balloon dilatation with 30 mm diameter and slower rate of balloon inflation is an effective and safe initial method of therapy for achalasia. The duration of remission can be extended by repeated dilatation with larger size balloon. PMID- 15298638 TI - Erythropoietin increases platelet reactivity and platelet counts in patients with alcoholic liver cirrhosis: a randomized, double-blind, placebo-controlled study. AB - BACKGROUND: Patients with liver cirrhosis have a complex haemostasis disturbance including thrombocytopenia and abnormal bleeding time. Erythropoietin is the primary stimulator for erythrocyte production and also induces megakaryocyte formation. In healthy men erythropoietin increased platelet count and platelet reactivity. AIM: As patients with liver cirrhosis often undergo invasive procedures, we were interested to study whether erythropoietin could improve platelet function in addition to thrombocytopenia. METHODS: In total, 22 thrombocytopenic (platelet counts < 120 g/L) patients with alcoholic liver cirrhosis received either 100 IE/kg erythropoietin or placebo on days 1, 3 and 5 in a 2:1 randomized, placebo-controlled double-blind fashion. Platelet counts and platelet reactivity (activator-stimulated expression of P-selectin on platelets measured by flow cytometry) were determined on study days 1, 3, 5 and 9. RESULTS: Median platelet count was 80 g/L which is borderline for major elective surgical interventions. Baseline values were not different between groups (P > 0.05). Treatment with erythropoietin increased platelet count by 25% (P = 0.01) and platelet reactivity twofold (P < 0.01) vs. baseline. The increase in platelet count vs. baseline was more pronounced in patients with platelet counts <80 g/L. No significant effect was observed in the placebo group. CONCLUSIONS: Treatment with erythropoietin significantly increased platelet counts and platelet reactivity in patients with alcoholic liver cirrhosis. Preoperative treatment with erythropoietin is therefore expected to yield higher platelet levels and better platelet function. PMID- 15298639 TI - The impact of immigration on the increasing incidence of hepatocellular carcinoma in the United States. AB - AIMS: To assess if the rising incidence of hepatocellular carcinoma in the United States can be accounted for by immigration and an ageing population, or is a true increase among the USA-born residents. METHODS: Design--A retrospective chart review. Setting--Urban, multiethnic hospital and specialty clinics in a large indigent health system in Houston, Texas. Subjects--Approximately 23,000 admissions and 143,000 out-patient clinic visits each year from 1992 through 2001 were assessed. A total of 494 patient records were selected and reviewed because of suspicion of hepatocellular carcinoma. Analysis- Hepatocellular carcinoma was confirmed by histopathology, alpha-fetoprotein level >400 ng/mL, and suggestive imaging studies. The age-adjusted incidence was determined and causative factors were identified. RESULTS: About 111 cases of confirmed hepatocellular carcinoma were found. The age-adjusted incidence rose from 3.44 per 100,000 hospital admissions during 1992-1996 (95% confidence interval: 2.86-4.02) to 5.19 during 1997-2001 (95% confidence interval: 4.41-5.97). The proportion of patients of non USA place of birth decreased between 1992-1996 and 1997-2001 (46-24%, respectively, P = 0.03). Fifty-two per cent and 68% were hepatitis C virus positive respectively; 37% and 34% were hepatitis B surface antigen-positive respectively; 46% and 59% had a history of alcohol abuse; and 22% and 11% had no identifiable risk factors. CONCLUSIONS: The incidence of hepatocellular carcinoma within the greater Houston area has increased during the past decade, rising by 51% from 1992-1996 to 1997-2001. This increase is not from immigration or population ageing but represents a true rise among the native born population. Hepatitis C and alcoholic cirrhosis are associated with a majority of cases, particularly in the latter half of the decade. PMID- 15298640 TI - Treatment of gastro-oesophageal reflux disease with rabeprazole in primary and secondary care: does Helicobacter pylori infection affect proton pump inhibitor effectiveness? AB - BACKGROUND: The presence of the gastric pathogen, Helicobacter pylori influences acid suppression by proton pump inhibitors and treatment outcome in patients with gastro-oesophageal reflux disease. AIM: To determine the influence of H. pylori infection on effectiveness of rabeprazole in primary and secondary care patients with gastro-oesophageal reflux disease. METHODS: Patients from primary and secondary care centres with uninvestigated gastro-oesophageal reflux disease (based on symptoms only) and investigated gastro-oesophageal reflux disease (endoscopically confirmed oesophagitis or endoscopy-negative reflux disease) were tested for H. pylori and treated with rabeprazole 20 mg once daily for 4-8 weeks in a non-randomized, multicentre, open-label study. Primary end-point for treatment effectiveness was complete resolution of both heartburn and acid regurgitation at 4-8 weeks; secondary end-point was quality of life as registered with the Psychological General Well-being Index. RESULTS: Data of 1787 patients could be analysed; mean duration of treatment was 36.3 days. At the evaluation visit 76.9% were heartburn-free, 77.7% regurgitation-free and 71% had complete symptom resolution. Overall Psychological General Well-being Index scores improved accordingly. Treatment was equally effective in patients with or without H. pylori infection, but more effective in patients with oesophagitis when compared with symptomatic gastro-oesophageal reflux disease. CONCLUSIONS: The effectiveness of rabeprazole in gastro-oesophageal reflux disease is not affected by the presence of H. pylori infection. PMID- 15298641 TI - The validity and accuracy of the Work Productivity and Activity Impairment questionnaire--irritable bowel syndrome version (WPAI:IBS). AB - BACKGROUND: Irritable bowel syndrome is a common chronic functional gastrointestinal disorder characterized by recurrent abdominal pain and discomfort associated with alterations in bowel habit. Irritable bowel syndrome affects patients' quality of life and increases productivity loss. AIM: To assess validity and accuracy of the Work Productivity and Activity Impairment questionnaire in irritable bowel syndrome as a tool for quantifying the effects of irritable bowel syndrome on productivity and daily activities. METHODS: Validity and accuracy were evaluated in 135 irritable bowel syndrome patients relative to three measures of irritable bowel syndrome disease severity; a debriefing questionnaire; retrospective diary; Work Limitations Questionnaire, and an activity impairment measure (Dimensions of Daily Activities). RESULTS: Symptom severity scores, diary scores, Work Limitations Questionnaire and Dimensions of Daily Activities were significant predictors of work productivity and activity impairment questionnaire in irritable bowel syndrome measures of work time missed, and work and activity productivity loss (P = 0.04 to < 0.0001). Impairment due to irritable bowel syndrome was estimated to be 2.9-4.3% for work time missed and 22-32% for impairment at work, the equivalent of 9.7 -14 h lost productivity per week. Activity impairment was 24-41%. CONCLUSIONS: Discriminative validity of the Work Productivity and Activity Impairment questionnaire in irritable bowel syndrome was established, making it the only validated tool for measuring the relative differences between disease severity groups and quantifying work productivity loss and activity impairment in irritable bowel syndrome patients. PMID- 15298642 TI - A rapid immunochromatographic assay for Helicobacter pylori in stool before and after treatment. AB - BACKGROUND: Current guidelines recommend non-invasive testing and treatment of young dyspeptic patients without alarm symptoms. AIM: To evaluate the accuracy of a new rapid immunochromatographic stool test to diagnose Helicobacter pylori infection before and after treatment compared with a gold standard. METHODS: Prospective, single-blind study, performed in a tertiary care hospital. A total of 303 consecutive dyspeptic patients underwent endoscopy with multiple biopsies. Infected patients were offered a treatment and invited to come back 4-6 weeks after the end of therapy to repeat the endoscopy. Patients were also asked to provide a stool sample before and after therapy. RESULTS: About 149 patients were H. pylori infected. The sensitivity and specificity before treatment were 91.3 and 93.5%; after treatment 92 and 100%. The likelihood ratios were robust enough to produce significant changes from pretest to post-test probability both in pre treatment (LR+ = 14, LR- = 0.093) and post-treatment (LR+ = 19.6, LR- = 0.095). CONCLUSIONS: The novel immunochromatographic stool test is fast, easy to perform and provides good differentiation between positive and negative results. It might become a rapid near patients test easily performed in the doctor office. PMID- 15298643 TI - Successful treatment of oesophageal candidiasis by micafungin: a novel systemic antifungal agent. AB - AIM: To determine the minimum effective dose and safety of micafungin in the treatment of HIV-related oesophageal candidiasis. METHOD: A total of 120 patients were enrolled in this open label study of the effects of daily 1 h infusions of micafungin on endoscopically proven fungal oesophagitis. Patients were randomly assigned to receive 12.5, 25, 50, 75 and 100 mg of micafungin daily. Response was evaluated clinically and endoscopically. RESULTS: The protocol defined minimum effective dose of micafungin was 12.5 mg. The percentage of patients experiencing clearing of physical signs and symptoms showed a dose-response relationship and reached 94.7% in the 100 mg dose group. All patients in the 50, 75 and 100 mg dose groups achieved an endoscopically verified improvement in oesophagitis. Adverse effects of micafungin were generally mild and not dose-related. No serious renal, hepatic or drug-related infusion reactions were encountered. CONCLUSION: Micafungin was found to be effective, well-tolerated and safe. The minimum effective dose was found to be 12.5 mg and a significant linear trend in the successful treatment of oesophageal candidiasis was observed across the doses used with 75 and 100 mg dose levels achieving high rates of clinical and endoscopic cure. PMID- 15298644 TI - Improvement in behavioral symptoms and advance of activity acrophase after short term bright light treatment in severe dementia. AB - Ten elderly subjects with severe dementia were given bright light (5000-8000 lux) for 45 min each morning for 4 weeks. Two rating scales of behavioral symptoms in dementia were used as outcome measures: Cohen-Mansfield Agitation Inventory (CMAI) and Behavior Pathology In Alzheimer's Disease Rating Scale (BEHAVE-AD), a scale for sleep-wake disturbances, and actigraphy to monitor activity rhythm. Behavioral symptoms improved with treatment. No changes in sleep-wake measures were found. There was an advance of the activity rhythm acrophase during treatment. These results suggest that short-time bright light improves behavioral symptoms and aspects of activity rhythm disturbances even in severely demented subjects. PMID- 15298645 TI - Obsessive-compulsive symptoms in parents of Tourette syndrome probands and autism spectrum disorder probands. AB - Obsessive-compulsive symptoms (OCS) frequently occur in patients with Tourette syndrome (TS) and autism spectrum disorders (ASD). It has been suggested that genetic factors play a role in the transmission of both TS and ASD and that obsessive-compulsive disorder (OCD) may have some genetic relationship with these disorders. The objective of this study was to explore whether the OCS associated with TS and ASD were found in the parents of TS and ASD probands by comparing them with normal controls. The subjects were parents of 13 TS and 16 ASD probands. All parents underwent an examination for tic symptoms and OCD, and completed the Maudsley Obsessional Compulsive Inventory (MOCI) and State-Trait Anxiety Inventory (STAI). No significant differences were observed in the MOCI and STAI scores among all three groups. However, the MOCI total score was higher in fathers of ASD probands than in male normal controls with a marginal significance. There was a significant tendency for the mean cleaning score of MOCI in fathers of ASD probands to be higher than that in male normal controls, and the mean checking score in fathers of ASD probands was fourfold higher than that in male normal controls, although there was no significant difference. No significant relationship was observed between OCS in TS or ASD probands and OCS of their parents. Further studies on OCD and OCS including a dimensional approach within ASD families are needed. PMID- 15298646 TI - Tourette's syndrome: psychopathology in adolescents. AB - The aim of this study is to foster an understanding of the clinical characteristics, in terms of general psychopathology, of Tourette syndrome (TS), as it manifests in adolescent patients, and to compare these results with those of previous studies conducted in Western countries. In total, 38 male and 5 female adolescent patients who visited a pediatric neurological specialty clinic for TS between 1 January and 1 March 2003, and who met the World Health Organisation criteria for TS, were studied. These patients were clinically interviewed and assessed with the Symptom-Checklist-90-R, Family APGAR and Tic Symptoms questionnaire, a self-report questionnaire based on DSM-IV TS diagnostic criteria. Among this population, the mean age of onset of motor tics was 9.65 +/- 3.7 years, and motor tics of the head and eyes were usually the first symptoms to manifest; coprolalia occurred in 44.1% (n = 19) of our patients. Many of our findings were similar to those reported in trials conducted in other regions of the world. TS symptoms showed stronger correlation with emotional distress in older patients. PMID- 15298647 TI - Lack of association between sigma receptor gene variants and schizophrenia. AB - Several pharmacological studies suggest the possible involvement of sigma(1) receptors in the pathogenesis of schizophrenia. An association has been reported between schizophrenia and two variants (GC-241-240TT and Gln2Pro) in the sigma(1) receptor gene (SIGMAR1). We also previously reported that, along with T-485 A, these two variants alter SIGMAR1 function. To investigate the role of SIGMAR1 in conveying susceptibility to schizophrenia, we performed a case-control study. We initially screened for polymorphisms in the SIGMAR1 coding region using PCR single strand conformation polymorphism analysis. The distribution of SIGMAR1 polymorphisms was analyzed in 100 schizophrenic and 104 control subjects. A novel G620A variant was detected in exon4. G620A was predicted to alter the amino acid represented by codon 211 from arginine to glutamine. Our case-control study showed no significant association between the T-485 A, GC-241-240TT, Gln2Pro, and G620A (Arg211Gln) variants and schizophrenia and clinical characteristics. These findings suggest that these SIGMAR1 variants may not affect susceptibility to schizophrenia. PMID- 15298648 TI - Olanzapine plus fluvoxamine and olanzapine alone for the treatment of an acute exacerbation of schizophrenia. AB - The objective of the present study was to compare the efficacy and adverse effects of olanzapine plus fluvoxamine and those of olanzapine alone, in schizophrenic patients with acute exacerbation. A randomized, placebo-controlled, 6-week trial was carried out at a University Hospital in Bangkok, Thailand. The efficacy and adverse effects were assessed biweekly by using the Brief Psychiatric Rating Scale (BPRS) and the Udvalg for Kliniske Undersogelser side effect scale, respectively. Twenty schizophrenic patients with acute exacerbation were randomly assigned to receive olanzapine plus fluvoxamine or olanzapine alone. The study found that the means of BPRS total and BPRS general psychopathology score changes were significantly larger in olanzapine plus fluvoxamine group (P = 0.037 and P = 0.045, respectively). The incidence of treatment adverse effects is comparable. In conclusion, the study findings suggest that fluvoxamine augmentation to olanzapine is well tolerated and more effective than olanzapine alone for short-term (6-week) treatment of an acute exacerbation of schizophrenia. Due to a number of limitations, further studies are warranted to confirm. PMID- 15298649 TI - Mood profile evaluation by families, young physicians and experienced psychiatrists in an actual clinical practice setting. AB - The purpose of this study was to explore differences in mood profile evaluation between raters with different backgrounds. Special emphasis was placed on expertise factor, influence of mood categories, and patient heterogeneity in a clinical setting. We administered Profile of Mood Scale (POMS) to patients' families, residents, and experienced psychiatrists as well as the patients, asking them to make an evaluation based on their image of the patients' subjective mood experience. Subscale scores and within-case across-item agreement indices among the raters were submitted for analysis. Analysis of variance of the subscale score showed significant effects of the rater by subscale-category and the rater by case components. The former was most influenced by the Vigor/Activity subscale in which the families and patients had higher scores than the physicians. As for rater-to-patient correlations of the score, psychiatrists were superior to residents in Anger/Hostility and Fatigue/Inertia, while families were superior to physicians in Vigor/Activity. These results suggest significance of expertise in mood evaluation as well as importance of time shared with the patients. The within-case across-item agreement data were subjected to cluster and canonical discriminant analysis. The analysis revealed four clusters which were best explained by two dimensions, one interpreted as general feasibility of evaluation and the other as characteristic emotional expression which caused separation of evaluation between families and physicians. Rater's expertise is a significant factor. Attention should also be paid to mood categories and patient heterogeneity. PMID- 15298650 TI - Patterns of self-cutting: a preliminary study on differences in clinical implications between wrist- and arm-cutting using a Japanese juvenile detention center sample. AB - The present study was aimed to clarify the differences in clinical implications between wrist- and arm-cutting. Subjects were 201 delinquent adolescents (178 males and 23 females) who had been admitted to a detention center from February to March 2003. A self-reporting questionnaire and the Adolescent Dissociative Experience Scale (ADES) were given. Traumatic events and other self-injurious behavior were compared among four groups. In total, 33 (16.4%) subjects reported wrist- and/or arm-cutting. Of the females, 60.9% (n = 14) had experienced self cutting behaviors compared to 10.7% of males (n = 19). Subjects were divided into four groups; 'non-cutting' (NC: n = 168, 83.6%), 'wrist-cutting' (WC: n = 5, 2.5%), 'arm-cutting' (AC: n = 19, 9.5%), and 'wrist- and arm-cutting' (WAC: n = 9, 4.5%). WC, AC, and WAC groups reported early separation, bulling in school, and histories of sexual/physical abuse more frequently than NC group. WC and WAC groups reported suicidal ideation and suicide attempts more frequently than NC and AC groups. The ADES scores in AC and WAC groups were significantly higher than in those in NC group (P < 0.001), while the scores in WC were not different from NC groups. WC and WAC groups self-cut due to suicide idea more frequently than the AC group, while AC group self-cut due to anger expression more frequently than WC or WAC groups. Self-cutters experienced early separation, bullying in school, and sexual/physical abuse more frequently than-non-self cutters. Arm-cutting behavior may predict dissociation, while wrist-cutting may involve with suicidality. PMID- 15298651 TI - Prevalence of post-traumatic stress disorder in incarcerated juvenile delinquents in Japan. AB - The present study investigated frequency of post-traumatic stress disorder (PTSD) in incarcerated juvenile delinquents in Japan. The presence of overwhelming traumatic experiences, which fulfilled the DSM-IV criterion A for PTSD, was evaluated using a self-report questionnaire in 251 delinquents (206 males and 45 females). The structured interview using the Clinician-administered PTSD Scale for DSM-IV (CAPS) was administered to those with the experiences. A substantial portion (36%) of the delinquents reported experiences of exposure to such overwhelming traumatic events, which fulfilled the criterion A. Among those who met the criterion A, 48 subjects (40 males and eight females) received the CAPS interview. Ten (21%) out of 48 were diagnosed with current or past history of full PTSD. Three subjects out of the 10 were currently diagnosed with full PTSD. Another 10 subjects out of 48 (21%) had fulfilled the criteria for current or past history of partial PTSD. Among the 10, one was diagnosed as currently suffering from partial PTSD. In females, frequencies of the past history were quite high (50% for full PTSD and 25% for partial PTSD in the eight subjects), while none was currently diagnosed with PTSD. PMID- 15298652 TI - Psychomotor retardation correlates with frontal hypoperfusion and the Modified Stroop Test in patients under 60-years-old with major depression. AB - Frontal hypoperfusion and frontal dysfunction have been reported in patients with major depression. It was also found that frontal hypoperfusion correlated with frontal dysfunction evaluated by neuropsychological tests in depressive patients aged 60 or over. These findings suggested that depression may cause frontal dysfunction and frontal hypoperfusion, and that these pathophysiological changes are manifested as psychomotor retardation. We performed single photon emission computed tomography and Modified Stroop Test on 35 patients with depression aged 25-83 to investigate association of depressive symptoms and psychological tests with cerebral blood perfusion. Additionally, we divided the patients into a younger (less than 60 years old) and an older (60 or over) group to examine the effect of age. Significant correlations were found between frontal perfusion, interference measure on Modified Stroop Test, and psychomotor retardation in all patients. These correlations were also found in the younger group. There was no significant difference on frontal perfusion, interference measure of the Modified Stroop Test, and psychomotor retardation between the two groups. The present findings suggest that frontal hypoperfusion, frontal dysfunction, and psychomotor retardation were associated with one another in not only the old but also the young patients with depression. PMID- 15298653 TI - Factors related to feelings of burden among caregivers looking after impaired elderly in Japan under the Long-Term Care insurance system. AB - Since the 1970s, the burden of caregiving has been the subject of rather intense study, a trend that will continue with the rapid graying of populations worldwide. Since the Long-Term Care insurance system began in 2000, few cross sectional studies have attempted to identify factors related to the feelings of burden among caregivers looking after the impaired elderly in Japan. In the present report, among 46 pairs of caregivers and impaired elderly, the elderly receiving regular nurses' visits in Kyoto Prefecture, Japan were assessed for problems with activities of daily living, the severity of dementia, the presence of behavioral disturbance, and cognitive impairment. The caregivers were asked to complete questionnaires in relation to their feelings of burden and caregiving situation. The results indicated that caregivers of impaired elderly with behavioral disturbances were more likely to feel a 'heavier burden.' Those temporarily relieved of caregiving three or more hours a day were less likely to experience 'heavier' caregiver burden than those who were not. Moreover, caregivers who found it 'inconvenient' to use care services tended to be more likely to feel a 'heavier' caregiver burden than those who did not. Recourse to respite services, which are ideally positioned to help, proved inconvenient because of their advance reservation system. More ready access to respite services in emergencies could do much to reduce caregiver burden. PMID- 15298654 TI - Obesity in schizophrenic outpatients receiving antipsychotics in Taiwan. AB - This investigation estimates and compares, for the first time, the distribution of body mass index (BMI: kg/m(2)) and the prevalence of obesity among Chinese outpatients with schizophrenia treated with antipsychotics. The BMI of 201 outpatients with schizophrenia-spectrum disorders was studied via a cross sectional naturalistic study. This investigation also compared the BMI of the subjects with a Taiwanese reference population. This investigation found no significant difference in the prevalence of obesity between male and female subjects. The prevalence of obesity among male and female patients in this investigation was, respectively, 2.74- and 2.51-fold greater than the Taiwanese reference population, and the prevalence of severe obesity among male and female patients was 4.66- and 3.53-fold greater than that in the Taiwanese reference population, respectively. The rate of severe obesity was especially high in patients treated with olanzapine. Atypical antipsychotics other than olanzapine did not seem to be more closely associated with obesity or severe obesity compared to typical antipsychotics. PMID- 15298655 TI - Effect of acute and chronic treatment with methamphetamine on mRNA expression of synaptotagmin IV and 25 KDa-synaptic-associated protein in the rat brain. AB - The effects of acute and chronic administration of methamphetamine (METH) on mRNA levels of synaptotagmin IV (SytIV) and an isoform of synaptic-associated protein of 25 KDa (SNAP25a) have been investigated in rat brain using in situ hybridization. Pretreatment with 0.5 mg/kg dopamine D1 receptor antagonist (SCH23390), but not 0.5 mg/kg N-methyl-D-aspartate (NMDA) receptor antagonist (MK 801), significantly attenuated the increased SytIV mRNA levels induced by acute METH administration in the striatum and the nucleus accumbens. Pretreatment with 0.5 mg/kg SCH23390, but not 0.5 mg/kg MK-801, significantly attenuated the increased SNAP25a mRNA levels induced by acute METH administration in the striatum and the dentate gyrus of the hippocampus. In the chronic treatment experiment, the SytIV mRNA levels of the group that received chronic treatment with METH followed by a METH challenge showed an increase similar to that seen after acute METH administration. In addition, those in the striatum, nucleus accumbens, and dentate gyrus were significantly higher than those of the group that received chronic treatment with saline followed by a METH challenge. The SNAP25a mRNA levels of the group that received chronic treatment with METH followed by a saline challenge were significantly higher than those of the group that received chronic treatment with saline followed by a saline challenge in the striatum and nucleus accumbens. The results of the present study suggest that SytIV may play an important role in the synaptic plasticity underlying METH induced neuroadaptive changes including behavioral sensitization. PMID- 15298656 TI - Factor analysis of Zung Scale scores in a Japanese general population. AB - The purpose of the present paper was to investigate the distribution of Zung Self rating Depression Scale (SDS) scores in a general population and its factor structure. Questionnaires on SDS items were sent to 7136 randomly selected residents aged 20-79 years who lived in districts in Japan with high rates of suicide. Valid responses were received from 5547 residents (response rate: 77.7%). Factor analysis of the SDS scores was conducted. The SDS scores of the male subjects were significantly lower than those of the female subjects in all age groups. A reverse-J-shaped relationship was found between age groups and mean SDS scores for the male and female subjects. The highest mean score was in the age group of 20-39 years, and the lowest mean score was in the age group of 60-69 years for the male and female subjects. In factor analysis, two factors consisting of 12 items were extracted, and 10 of those 12 items covered six Diagnostic and Statistical Manual of Mental Disorders (4th edn; DSM-IV) criteria describing psychological disturbances of depression. The distribution of SDS scores differed depending on the age group. Major components of SDS in the subjects covered the DSM-IV criteria for psychological disturbances of depression. PMID- 15298657 TI - Paroxetine in the treatment of obsessive-compulsive disorder: randomized, double blind, placebo-controlled study in Japanese patients. AB - The efficacy of paroxetine in the treatment of obsessive-compulsive disorder in Western populations is well established. The present study compares the efficacy and safety of paroxetine with placebo in the treatment of obsessive-compulsive disorder in Japanese patients. Patients aged 16 years or older who met Diagnostic and Statistical Manual of Mental Disorders (4th edn; DSM-IV) criteria for obsessive-compulsive disorder and had a Yale-Brown Obsessive-Compulsive Scale (Y BOCS) score of >/=16 were randomized to receive 12 weeks' therapy in a double blind manner. Paroxetine 20-50 mg/day or placebo was administered following a 1 week, placebo run-in phase. One hundred and ninety-one patients were randomized to either paroxetine or placebo, 188 patients were assessed as the full analysis set (FAS) and 144 patients completed the 12 week study. After adjustment for the Y-BOCS total score at baseline, reductions in obsessive-compulsive total score at week 6 and at the end of therapy were significantly greater in the paroxetine group than the placebo group. Most of the adverse events that occurred during the study were of mild to moderate intensity. Paroxetine is effective and well tolerated in Japanese adult patients with obsessive-compulsive disorder. PMID- 15298658 TI - Cigarette smoking among patients with schizophrenia and bipolar disorders. AB - The purpose of the present study was to investigate the ratio of smokers and the relationship of cigarette smoking to clinical features in patients with schizophrenia and bipolar disorders. One hundred and forty-four patients with schizophrenia or bipolar disorder along with 114 healthy controls were evaluated. A total of 57.5% of the patients with schizophrenia, 55.1% of the bipolar patients and 47.3% of the control group were smokers. Daily cigarette consumption among the patients with schizophrenia was higher than that for the bipolar patients, and control group. Among the patients with schizophrenia who were in acute psychotic episode, Scale for the Assessment of Positive Symptoms scores of the smokers were significantly higher. PMID- 15298659 TI - Association of the XBP1-116C/G polymorphism with schizophrenia in the Japanese population. AB - Schizophrenia and bipolar disorder share some clinical features and linkage studies have shown that several loci are common. Recently, the authors found that the -116C-->G substitution in the promotor region of XBP1, a pivotal gene in endoplasmic reticulum (ER) stress response, causes the impairment of ER stress response, and that the -116C/C genotype is a protective factor; in other words the presence of the G allele increases the risk for bipolar disorder. The gene is located on 22q12.1, which is also linked with schizophrenia. The polymorphisms were investigated in 234 schizophrenic patients as compared with controls. Significant difference of genotype distribution was observed, which suggested that the -116C/C genotype is a protective factor for both of the major mental disorders. PMID- 15298660 TI - Involvement of nucleus accumbens dopaminergic transmission in acoustic startle: observations concerning prepulse inhibition in rats with entorhinal cortex lesions. AB - The relationship between the entorhinal cortex and prepulse inhibition (PPI) as well as the nucleus accumbens dopaminergic participation in acoustic startle were examined in rats. After the entorhinal cortex was damaged bilaterally using ibotenic acid, a microdialysis probe was placed in the nucleus accumbens for detection of dopamine before, during and after acoustic startle stimuli. In rats with bilateral entorhinal cortex lesions PPI was reduced, and extracellular dopamine in the nucleus accumbens was elevated with or without acoustic stimuli. The entorhinal cortex and the sensorimotor gating system thus may be related via dopaminergic connections in the nucleus accumbens, even though dopamine release did not coincide completely with acoustic startle stimuli. PMID- 15298661 TI - Post-stroke mania in late life due to right temporoparietal infarction. PMID- 15298662 TI - Fluoxetine-induced mania in an Asian patient. PMID- 15298663 TI - Two cases of deep vein thrombosis associated with antipsychotic drug use. PMID- 15298664 TI - Effect of inhibiting melatonin biosynthesis on spatial memory retention and tau phosphorylation in rat. AB - We have found recently that melatonin protects SH-SY5Y neuroblastoma cells from calyculin A-induced neurofilament impairment and neurotoxicity. In the present study, we further investigated the in vivo effect of inhibiting melatonin biosynthesis on spatial memory retention and tau phosphorylation in rats and the potential underlying mechanisms by using haloperidol, a specific inhibitor of 5 hydroxyindole-O-methyltransferase, and a key enzyme in melatonin biosynthesis. We have found that injection of haloperidol into the lateral ventricle and into peritoneal cavity compromises spatial memory retention of rats and induces hyperphosphorylation of microtubule-associated protein tau at tau-1 (Ser199/Ser202) and PHF-1 (Ser396/Ser404) epitopes. At mean time, the activity of protein phosphatase-2A (PP-2A), a deficit phosphatase in the Alzheimer's disease brain and superoxide dismutase decreases with an elevated level of malondialdehyde. Supplementation with melatonin by prior injection for 1 wk and reinforcement during the haloperidol administration significantly improves memory retention deficits, arrests tau hyperphosphorylation and oxidative stress, and restores PP-2A activity. These results strongly support the involvement of decreased melatonin in Alzheimer-like spatial memory impairment and tau hyperphosphorylation, and PP-2A may play a role in mediating aberrant melatonin induced lesions. PMID- 15298665 TI - Melatonin reduces dimethylnitrosamine-induced liver fibrosis in rats. AB - Increased deposition of the extracellular matrix components, particularly collagen, is a central phenomenon in liver fibrosis. Stellate cells, the central mediators in the pathogenesis of fibrosis are activated by free radicals, and synthesize collagen. Melatonin is a potent physiological scavenger of hydroxyl radicals. Melatonin has also been shown to be involved in the inhibitory regulation of collagen content in tissues. At present, no effective treatment of liver fibrosis is available for clinical use. We aimed to test the effects of melatonin on dimethylnitrosamine (DMN)-induced liver damage in rats. Wistar albino rats were injected with DMN intraperitoneally. Following a single dose of 40 mg/kg DMN, either saline (DMN) or 100 mg/kg daily melatonin was administered for 14 days. In other rats, physiologic saline or melatonin were injected for 14 days, following a single injection of saline as control. Hepatic fibrotic changes were evaluated biochemically by measuring tissue hydroxyproline levels and histopathogical examination. Malondialdehyde (MDA), an end product of lipid peroxidation, and glutathione (GSH) and superoxide dismutase (SOD) levels were evaluated in blood and tissue homogenates. DMN caused hepatic fibrotic changes, whereas melatonin suppressed these changes in five of 14 rats (P < 0.05). DMN administration resulted in increased hydroxyproline and MDA levels, and decreased GSH and SOD levels, whereas melatonin reversed these effects. When melatonin was administered alone, no significant changes in biochemical parameters were noted. In conclusion, the present study suggests that melatonin functions as a potent fibrosuppressant and antioxidant, and may be a therapeutic choice. PMID- 15298666 TI - Pretreatment with melatonin exerts anti-inflammatory effects against ischemia/reperfusion injury in a rat middle cerebral artery occlusion stroke model. AB - Inflammatory response following cerebral ischemia/reperfusion plays a key pathogenic role in ischemic cerebral damage. Nitric oxide (NO), cyclooxygenase-2 (COX-2) and myeloperoxidase (MPO) are important inflammatory mediators. Neuronal NO synthase (nNOS) is a major initial source of excessive NO during ischemia/reperfusion. Induction of COX-2 and infiltration of polymorphonuclear cells expressing MPO are critical factors in delayed inflammatory damage. Previously, we demonstrated that administration of melatonin before ischemia significantly reduced the infarct volume in a rat middle cerebral artery occlusion (MCAO) stroke model. In this study, we examined the effect of pretreatment with melatonin at 5 mg/kg on the immunoreactivity (ir) for nNOS, COX 2, MPO, and glial fibrillary acidic protein (GFAP) at 24, 48, and 72 hr after right-sided endovascular MCAO for 1 hr in adult male Sprague-Dawley rats. Melatonin did not affect the hemodynamic parameters. When compared with rats with sham MCAO, ischemia/reperfusion led to an ipsilateral increase in cells with positive ir for nNOS (similar at all times) and in ir-GFAP (similar at all times). Ischemia/reperfusion led to appearance of cells with positive ir for COX 2 (greatest at 24 hr with a tendency to increase again at 72 hr) or MPO (greatest at 24 hr). A single dose of melatonin significantly lessened the ipsilateral increase in cells with positive ir for nNOS, COX-2 or MPO, but did not influence the ipsilateral change in ir-GFAP. Our results suggest that melatonin treatment mediates neuroprotection against ischemia/reperfusion injury partly via inhibition of the consequential inflammatory response. PMID- 15298667 TI - Attenuation of lipopolysaccharide-induced hyperreactivity of human internal mammary arteries by melatonin. AB - Reactive oxygen species (ROS) are thought to be important mediators in ischaemia/reperfusion injury following coronary vasospasm. The most ubiquitous action of melatonin is that of a free radical scavenger. Therefore, we investigated the action of melatonin by monitoring changes in the tone on ring preparations from human internal mammary arteries (IMA). In quiescent IMA rings melatonin (0.1 nm-10 microm) never elicited any change in baseline tension but 1 100 nm melatonin enhanced significantly maximal responses to noradrenaline (NA) in arteries with endothelial function. In NA (1 microm) precontracted arteries inhibition of nitric oxide (NO(*)) formation by N(G)-monomethyl-L-arginine (l NMMA, 100 and 400 microm) eliminated 43 +/- 7 and 61 +/- 7% of the acetylcholine (ACH) effect. Melatonin (100 and 400 nm) attenuated maximal endothelium-dependent relaxant responses to ACH slightly by 23 +/- 9 and 17 +/- 9% leaving responses to direct stimulation of soluble guanylate cyclase by sodium nitroprusside unchanged. Incubation of IMA for 20 hr at 37 degrees C with 1 microg/mL lipopolysaccharide (LPS) enhanced maximal NA effects to 147 +/- 18% (n = 22, P < 0.01) whereas 50 microg/mL LPS reduced the NA maxima to 68 +/- 9% (n = 10, P < 0.01) of the control effects. The LPS-induced potentiation was completely attenuated by coincubation with melatonin (400 nm) and significantly reduced by coincubation with the thromboxane synthase inhibitor dazoxiben (10 microm). It is suggested that the LPS-induced hyperreactivity of vascular smooth muscle is mediated through enhanced release of ROS and prostanoids and that melatonin inhibits the vascular hyperreactivity through selective scavenging of ROS. PMID- 15298668 TI - Melatonin stimulates calmodulin phosphorylation by protein kinase C. AB - Calmodulin (CaM)-dependent processes can be modulated by the availability of Ca(+2), the subcellular distribution of both CaM and its target proteins, CaM antagonism, and post-translational modifications such as CaM phosphorylation. Melatonin, the pineal secretory product synthesized during the dark phase of the photoperiod is an endogenous CaM antagonist. This indolamine causes CaM subcellular redistribution in epithelial MDCK and MCF-7 cells, and selectively activates protein kinase C alpha (PKC alpha) in neuronal N1E-115 cells. In the present work we have characterized the phosphorylation of CaM mediated by PKC alpha and its stimulation by melatonin in an in vitro reconstituted enzyme system. Additionally, the participation of MAPK and ERKs, downstream kinases of the PKC signaling pathway, was explored utilizing MDCK cell extracts as source of these kinases. Phosphorylation of CaM was characterized in the whole cells by MDCK cell metabolic labeling with [(32)P]-orthoposhospate, and CaM separation by sodium dodecyl sulphate-polyacrylamide gel electrophoresis, as well as by immunocolocalization of phosphorylated threonine/serine residues and CaM in cultured cells incubated with melatonin. Our results show that melatonin increased CaM phosphorylation by PKC alpha with an EC(50) of 10(-8) m in the presence of the phorbol ester, phorbol-12-myristate-13-acetate (PMA) in the in vitro reconstituted enzyme system. An increase in phosphorylated CaM was also observed in cells cultured with melatonin, or PMA for 2 hr, while, PKC, MAPK, or ERK inhibitors abolished CaM phosphorylation elicited by melatonin in MDCK cell extracts. Our data show that melatonin can stimulate phosphorylation of CaM by PKC alpha in the in vitro reconstituted system and suggest that in MDCK cells this phosphorylation is accomplished by PKC. Modification of CaM by melatonin can be another route to inhibit CaM interaction with its target enzymes. PMID- 15298669 TI - Melatonin suppresses reactive oxygen species induced by UV irradiation in leukocytes. AB - An investigation of the antioxidative UV protective effect of melatonin was performed in an in vitro irradiation model with leukocytes. Leukocytes were isolated from EDTA-treated whole blood and taken up in phosphate-buffered saline (PBS). Five of 10 aliquots were incubated with 2 mmol/L melatonin and 5 with PBS as a control. The samples were irradiated by UV light (280-360 nm, max: 310 nm) at doses between 75 and 300 mJ/cm(2) or left unirradiated. Radical formation was measured using the chemiluminescence technique. Staining with trypan blue was performed to assess cell viability. Melatonin significantly suppressed radical formation in cell solutions irradiated from 75 to 300 mJ/cm(2) (P /=1 mm ST segment elevation in V3R-V5R of right-sided ECG and 63 patients without RVMI constituted the control group. All patients underwent echocardiography within 24 hours of admission. Normal range of MPI for our laboratory was estimated from 50 age matched healthy subjects. RV MPI was elevated to a mean of 0.53 +/- 0.22 in RVMI (Normal MPI 0.20 +/- 0.05, P-value < 0.001). IMI without RVMI did not elevate MPI significantly (0.21 +/- 0.17, P-value NS). Repeat MPI estimation in 11 RVMI (7 thrombolyzed) patients after 5 days showed dramatic reduction (0.23 +/- 0.12, P value < 0.001). This reduction was noted irrespective of thrombolysis. RV MPI >/= 0.30 has high sensitivity (82%) and specificity (95%) for the diagnosis of RVMI in the presence of acute IMI. MPI can reliably diagnose RV infarction. It can be used to quantify right ventricular dysfunction and assess acute improvements in RV function. PMID- 15298684 TI - Microalbuminuria is associated with reduced cardiac cyclic variation of integrated backscatter signal in severe hypertension. AB - OBJECTIVE: Microalbuminuria (MA) as a marker of systemic vascular disease and left ventricular (LV) hypertrophy is associated with increased cardiovascular mortality and morbidity in patients with essential hypertension. The aim of this study was to investigate changes in cardiac cycle-dependent variation of integrated backscatter signals (CVIBS) in hypertensive patients with MA. METHODS: Randomly selected 60 hypertensive patients (mean age 51 +/- 8) with uncontrolled blood pressure (BP) (>/=130 mmHg systolic and/or 85 mmHg diastolic) were included. All patients underwent urinary albumin excretion (UAE) measurements, 24 hour ambulatory BP monitoring, and LV echocardiographic examination. UAE was measured in two separate 24-hour urine collection and mean of two values was taken into consideration. Normotensive 20 healthy subjects served as controls. CVIBS values were obtained from mid-anteroseptal, mid-posterolateral, and mid inferior areas at the papillary muscle level in the parasternal short-axis view. CVIBS was defined as the difference in integrated backscatter values between systole and diastole. CVIBS values in MA positive patients were compared with the values in MA negative patients and control subjects. RESULTS: Twelve patients had MA (UAE 30 to 300 mg/day) while 48 patients had normal UAE (<30 mg/day). The wall thickness (at septum and posterior) and left ventricular mass index (LVMI) values were all significantly higher in hypertensive patients with MA (P < 0.01). The CVIBS values in MA positive group were significantly lower than the CVIBS values both in MA negative hypertensive patients and control subjects (P < 0.01). CONCLUSION: This study demonstrates that in hypertensive patients a high LVMI is associated with reduced CVIBS values and MA appears to be a marker of hypertrophy. PMID- 15298685 TI - Improved detection of spontaneous echo contrast in the aorta with tissue Doppler imaging. AB - AIMS: Spontaneous echo contrast (SEC) within the cardiac chambers has been associated with increased risk of thromboembolism. We investigated the presence and severity of SEC in the aorta with tissue Doppler imaging (TDI) and compared these to the aortic flow velocity and to the clinical profile of the patients. METHODS AND RESULTS: Seventy patients (35 males, 35 females, mean age 64, 22-86 years) underwent TEE for standard indications. Spontaneous echo contrast was studied with conventional and TDI imaging. Aortic flow velocity was measured in the center and lateral part of the descending aorta. SEC of any grade was detected in 24 patients with conventional imaging and in 53 using TDI (P < 0.0001). The presence of swirling was associated with aortic atherosclerosis, older age, history of hypertension and coronary artery disease, atrial fibrillation, and previous embolic events. There was correlation between intraaortic swirling, larger descending aortic diameter (23.6 vs 17 mm, P < 0.00001) and lower peak aortic flow velocity (55 vs 68 cm/s, P = 0.038). CONCLUSION: Spontaneous echo contrast in the aorta is common in high-risk patients and is associated with increased clinical profile, larger aortic diameter, and lower peak aortic flow velocity. Tissue Doppler imaging is more sensitive in the detection of SEC than conventional imaging. PMID- 15298686 TI - Instantaneous quantitative video intensity heterogeneity: evaluation with low mechanical index contrast echocardiography. AB - BACKGROUND: Instantaneous video intensity of myocardium has been poorly characterized. Myocardial video intensity is usually displayed in the fitted curve from the exponential equation, y = a(1 - e (-bt)). However, information from the fitted curve will be as accurate as the original video intensity data from the perfusion image. Therefore, we sought to characterize the intramyocardial instantaneous video intensity from low mechanical index (MI) contrast echo imaging for variation. METHOD: Low-MI imaging using a nonlinear cancellation technique was performed on 10 subjects with normal myocardium. Quantitative video intensity was analyzed in five segments in the epicardium and subendocardium, as well as in systole and diastole. RESULTS: Video intensity varied between the epicardium and endocardium in each of the region that was analyzed, with the greatest variation in the inferior region (P < 0.0001). Diastolic and systolic differences were also present. CONCLUSION: Instantaneous video intensity is heterogeneous within the myocardium. Differences can result from attenuation, myocardial fiber structure, and even isotropic effects of the contrast agent, and should be taken into account when data are fitted into an exponential function. PMID- 15298688 TI - P wave dispersion and left atrial appendage function for predicting recurrence after conversion of atrial fibrillation and relation of p wave dispersion to appendage function. AB - BACKGROUND: We investigated P wave dispersion and left atrial appendage (LAA) function for predicting atrial fibrillation (AF) relapse, and the relationship between P wave dispersion and LAA function. METHODS: Sixty-four consecutive patients with AF lasting /=5 days, LA size >/=45 mm, maximum P wave duration >/=112 ms, P wave dispersion >/=47 ms, spontaneous echo contrast, minimum LAA area >/=166 mm(2), and LAA emptying velocity <36 cm/sec were univariate predictors of recurrence (each P < 0.05). By multivariate analysis, LA size (P = 0.02), P wave dispersion (P < 0.001), and LAA emptying flow (P = 0.01) identified patients with recurrent AF. Their positive predictive values were 91, 97, and 72%, respectively. CONCLUSION: The increased P wave dispersion in addition to the dilated LA and the depressed LAA emptying flow can identify patients at risk of recurrent AF after cardioversion. PMID- 15298687 TI - Age is not a predictor of patent foramen ovale with right-to-left shunt in patients with cerebral ischemic events. AB - BACKGROUND: Patent foramen ovale (PFO) with or without atrial septal aneurysm (ASA) is highly associated with cerebral ischemic events in young patients. The prevalence of PFO and ASA in elderly patients with cerebral ischemic events is not well described. OBJECTIVE: Our study is to evaluate the frequencies of PFO with right-to-left shunt (RLS) and ASA in elderly patients and to determine whether age is a predictor of flow-reversed PFO with RLS in cerebral ischemic events. METHODS: A prospective registry for all consecutive patients with cerebral ischemic events who were evaluated by transesophageal echocardiography (TEE) for the detection of possible cardiac source of embolization was established and maintained in a university hospital. Patients' demographics including age, gender, ethnic origin, cerebrovascular risk factors, and all positive TEE data were collected from July 2000 to August 2001 for statistical analysis. A univariate and multivariate stepwise logistic regression analysis was performed. RESULTS: In older patients the prevalence of PFO with RLS, PFO, and ASA was 25/118 (20%), 28/118 (24%), and 38/118 (32%), respectively, as opposed to younger patients, in whom it was 35/119 (30%), 39/119 (33%), and 38/119 (32%), respectively. Older patients had higher frequencies of hypertension (59; 69%), CAD (25; 21%), and prior history of stroke (23; 20%) as opposed to younger patients. Younger age (<60 years), gender, smoking history, hypertension, hyperlipidemia, CAD, and prior history of stroke were not associated with higher prevalence of PFO with RLS. Patent foramen ovale was associated with ASA (P < 0.001) and LVH (P < 0.019) in patients with TIA and stroke. In multivariate analysis only ASA (P < 0.001) remained significant with PFO, with RLS controlling for age, gender, and LVH. CONCLUSIONS: PFO with RLS and ASA are frequently present in elderly stroke and/or TIA patients and age is not a predictor for PFO. Transesophageal echocardiography should be considered for all stroke and/or TIA patients irrespective of their age. PMID- 15298689 TI - Extremely rapid formation of mitral valve ring abscess in infective endocarditis. AB - A patient with infective endocarditis (IE) due to methicillin-resistant staphylococcus aureus (MRSA) was found to have conversion of the hypoechoic region of the posterior mitral valve ring apparatus into a clearly delineated echolucent space by repeating transthoracic echocardiography at an interval of 1 week. Color Doppler showed features of blood entry into this space. Abscess formation in IE due to MRSA may be quick and repeated echocardiography may help detect the complications of IE. Semiurgent mitral valve plasty was performed for the associated prolapse of the posterior mitral leaflet using a hand-made, rolled, twisted autologous pericardial ring. PMID- 15298690 TI - Hypertrophic cardiomyopathy: a case of symptomatic Japanese type apical hypertrophic cardiomyopathy. AB - A 61-year-old male patient was hospitalized due to the exertional angina pectoris. A diagnosis of apical hypertrophic cardiomyopathy was made by ECG (electrocardiography), echocardiographic, and coronary angiographic findings. This case was reported and related literature was reviewed because of its similarity to Japanese type apical hypertrophic cardiomyopathy (AHCMP) cases rarely seen outside Asia. PMID- 15298691 TI - Retinal embolization of bicuspid aortic valve calcification. AB - We report the case of a 36-year-old man with calcified bicuspid aorta. Aortic disease was diagnosed after retinal embolism, which caused loss of vision affecting the left eye. Doppler examination did not identify carotid stenosis. Transthoracic echocardiography showed thickness and calcification of two leaflets, moderate aortic regurgitation, high normal left ventricular diastolic dimensions, and high normal interventricular septum thickness. Parasternal long axis view in the systolic frame showed dilated left ventricular outflow tract, dilated ascending aorta, and calcification and thickening of aortic leaflets, causing an echo-lucent area, in the diastolic frame. It also showed the eccentric position of the closed valve leaflets, short axis in the diastolic frame, and the absence of the typical "Y" letter sign, tracked by the three leaflets (septal, right coronary, and left coronary) in the normal tricuspid aortic valve. On long axis parasternal acquisition, we note high normal interventricular septum thickness and high normal diastolic diameter. Color Doppler analysis from the apical five-chamber view showed aortic regurgitation; regurgitant jet area was about 44% of the outflow tract, so aortic regurgitation was classified as moderate. Pulsed Doppler showed a high normal value of peak aortic velocity. Transesophageal echocardiography was performed, but no further findings were identified. We think that the likely retinal embolism sources were the aortic calcified leaflets, even if the aortic valve was not stenotic and with no signs of endocarditis. However, because we cannot exclude other potential causes, we put the patient on anticoagulant therapy. Furthermore, aortic valve replacement was not advised. PMID- 15298692 TI - Aortic ectasia with hemiazygos vein dilatation: a mimicker of aortic dissection. AB - Infrahepatic interruption of the inferior vena cava (IVC) is a rare but well documented finding. In this condition, the IVC between renal and hepatic vein is absent and the hepatic veins directly empty into the right atrium; because of the enlargement of the azygos-hemiazygos vein system, this condition could mimic aortic pathology. We will describe a case of aortic arch enlargement with dilatation of hemiazygos vein, which was initially misdiagnosed by two dimensional transesophageal echocardiography (TEE) as aortic dissection. TEE Doppler identified the real condition, which was confirmed by computed tomography. PMID- 15298693 TI - Esophageal duplication cyst: a challenging diagnosis of a paracardiac mass. AB - We present a case of esophageal duplication cyst, echocardiographically appearing as a mass above the roof of the left atrium and behind the right pulmonary artery. The differential diagnosis and the management of such disease are discussed. PMID- 15298694 TI - Precordial abscess inducing chest pain 20 years after surgical repair of a pentalogy of fallot. AB - A 25-year-old male asylum-seeker presented with chest pain, exertional dyspnea, and orthopnea 20 years after the surgical repair of a pentalogy of Fallot. An extracardiac mass compressing the right ventricle was subsequently detected and surgical decompression was performed to relieve the resulting right intraventricular hypertension. At operation, the mass proved to be a coagulase negative, staphylococcal abscess. In addition, the removal of the mass unmasked a previously nonrecognized pulmonary outflow stenosis that required balloon dilatation and beta-blocker therapy. While infections are known to occur after sternotomy, the formation of an abscess in the anterior mediastinum several years after the intervention appears to be exceptional; this diagnosis came to mind only after the more common complications had been considered, e.g., pseudoaneurysm or pericardial hematoma. To our knowledge, this is the first report of an abscess in the anterior mediastinum that had probably formed over many years following a sternotomy, compressed the right ventricle and masked a pulmonary stenosis. PMID- 15298695 TI - Live three-dimensional transthoracic echocardiographic assessment of anomalous origin of left coronary artery from the pulmonary artery. AB - We describe live three-dimensional transthoracic echocardiographic (3DTTE) findings in a 52-year-old female who had previously undergone an aortopulmonary tunnel operation for anomalous origin of the left coronary artery (ACA) from the pulmonary artery. Three-dimensional transthoracic echocardiography clearly delineated the origin of the ACA from the posterolateral aspect of the main pulmonary artery just above the pulmonary valve, the surgically created tunnel, as well as a small defect in the tunnel near the aortic end communicating with the pulmonary artery. PMID- 15298696 TI - Pseudo-left ventricle apical hypertrophy: bedside diagnosis with SonoVue contrast. PMID- 15298697 TI - Pulmonary stenosis caused by external compression of non-Hodgkin lymphoma. PMID- 15298698 TI - How to measure left atrial volume. PMID- 15298699 TI - Outcome after acute myocardial infarction: a comparison of patients seen by cardiologists and general physicians. AB - BACKGROUND: The management of acute myocardial infarction (AMI) has improved over the last 50 years with the more frequent use of effective medicines and procedures. The clinical benefit of the specialty of the attending physician is less clear. The United Kingdom National Service Framework for coronary heart disease (CHD) suggested that patients with CHD are likely to benefit from cardiological supervision. We set out to assess the effect of access to cardiologists on survival among AMI patients admitted in two UK hospitals. METHODS: The study was conducted in a university hospital and a district general hospital in England. Information was obtained on age, sex, ethnicity, Carstairs socioeconomic deprivation category derived from postcode of residence, comorbidity, distance from hospital and medication from all patients admitted with acute myocardial infarction in two acute trusts between July 1999 and June 2000. Record linkage to subsequent Hospital Episode Statistics and Registrar General's death records provided follow up information on procedures and mortality up to eighteen months after admission. Cox proportional hazard models were used to investigate the main hypothesis controlling for confounding. The main outcome measure was 18-month survival after myocardial infarction. RESULTS: Access to a cardiologist was univariately associated with improved survival (hazard ratio 0.16, 95% CI 0.10 to 0.25). This effect remained after controlling for the effect of patient characteristics (hazard ratio 0.22, 95% CI 0.14 to 0.25). The effect disappeared after controlling for access to effective medication (hazard ratio 0.70, 95% CI 0.33 to 1.46). CONCLUSIONS: Access to a cardiologist is associated with better survival compared to no access to a cardiologist among a cohort of patients already admitted with AMI. This effect is mainly due to the more frequent use of effective medicines by the group referred to cardiologists. Hospitals may improve survival by improving access to effective medicines and by coordinating care between cardiologists and general physicians. PMID- 15298700 TI - 3 dimensional modelling of early human brain development using optical projection tomography. AB - BACKGROUND: As development proceeds the human embryo attains an ever more complex three dimensional (3D) structure. Analyzing the gene expression patterns that underlie these changes and interpreting their significance depends on identifying the anatomical structures to which they map and following these patterns in developing 3D structures over time. The difficulty of this task greatly increases as more gene expression patterns are added, particularly in organs with complex 3D structures such as the brain. Optical Projection Tomography (OPT) is a new technology which has been developed for rapidly generating digital 3D models of intact specimens. We have assessed the resolution of unstained neuronal structures within a Carnegie Stage (CS)17 OPT model and tested its use as a framework onto which anatomical structures can be defined and gene expression data mapped. RESULTS: Resolution of the OPT models was assessed by comparison of digital sections with physical sections stained, either with haematoxylin and eosin (H&E) or by immunocytochemistry for GAP43 or PAX6, to identify specific anatomical features. Despite the 3D models being of unstained tissue, peripheral nervous system structures from the trigeminal ganglion (approximately 300 microm by approximately 150 microm) to the rootlets of cranial nerve XII (approximately 20 microm in diameter) were clearly identifiable, as were structures in the developing neural tube such as the zona limitans intrathalamica (core is approximately 30 microm thick). Fourteen anatomical domains have been identified and visualised within the CS17 model. Two 3D gene expression domains, known to be defined by Pax6 expression in the mouse, were clearly visible when PAX6 data from 2D sections were mapped to the CS17 model. The feasibility of applying the OPT technology to all stages from CS12 to CS23, which encompasses the major period of organogenesis for the human developing central nervous system, was successfully demonstrated. CONCLUSION: In the CS17 model considerable detail is visible within the developing nervous system at a minimum resolution of approximately 20 microm and 3D anatomical and gene expression domains can be defined and visualised successfully. The OPT models and accompanying technologies for manipulating them provide a powerful approach to visualising and analysing gene expression and morphology during early human brain development. PMID- 15298701 TI - The p68 and p72 DEAD box RNA helicases interact with HDAC1 and repress transcription in a promoter-specific manner. AB - BACKGROUND: p68 (Ddx5) and p72 (Ddx17) are highly related members of the DEAD box family and are established RNA helicases. They have been implicated in growth regulation and have been shown to be involved in both pre-mRNA and pre-rRNA processing. More recently, however, these proteins have been reported to act as transcriptional co-activators for estrogen-receptor alpha (ER alpha). Furthermore these proteins were shown to interact with co-activators p300/CBP and the RNA polymerase II holoenzyme. Taken together these reports suggest a role for p68 and p72 in transcriptional activation. RESULTS: In this report we show that p68 and p72 can, in some contexts, act as transcriptional repressors. Targeting of p68 or p72 to constitutive promoters leads to repression of transcription; this repression is promoter-specific. Moreover both p68 and p72 associate with histone deacetylase 1 (HDAC1), a well-established transcriptional repression protein. CONCLUSIONS: It is therefore clear that p68 and p72 are important transcriptional regulators, functioning as co-activators and/or co-repressors depending on the context of the promoter and the transcriptional complex in which they exist. PMID- 15298703 TI - Towards the development of a DNA-sequence based approach to serotyping of Salmonella enterica. AB - BACKGROUND: The fliC and fljB genes in Salmonella code for the phase 1 (H1) and phase 2 (H2) flagellin respectively, the rfb cluster encodes the majority of enzymes for polysaccharide (O) antigen biosynthesis, together they determine the antigenic profile by which Salmonella are identified. Sequencing and characterisation of fliC was performed in the development of a molecular serotyping technique. RESULTS: FliC sequencing of 106 strains revealed two groups; the g-complex included those exhibiting "g" or "m,t" antigenic factors, and the non-g strains which formed a second more diverse group. Variation in fliC was characterised and sero-specific motifs identified. Furthermore, it was possible to identify differences in certain H antigens that are not detected by traditional serotyping. A rapid short sequencing assay was developed to target serotype-specific sequence motifs in fliC. The assay was evaluated for identification of H1 antigens with a panel of 55 strains. CONCLUSION: FliC sequences were obtained for more than 100 strains comprising 29 different H1 alleles. Unique pyrosequencing profiles corresponding to the H1 component of the serotype were generated reproducibly for the 23 alleles represented in the evaluation panel. Short read sequence assays can now be used to identify fliC alleles in approximately 97% of the 50 medically most important Salmonella in England and Wales. Capability for high throughput testing and automation give these assays considerable advantages over traditional methods. PMID- 15298702 TI - Co-transcriptional folding is encoded within RNA genes. AB - BACKGROUND: Most of the existing RNA structure prediction programs fold a completely synthesized RNA molecule. However, within the cell, RNA molecules emerge sequentially during the directed process of transcription. Dedicated experiments with individual RNA molecules have shown that RNA folds while it is being transcribed and that its correct folding can also depend on the proper speed of transcription. METHODS: The main aim of this work is to study if and how co-transcriptional folding is encoded within the primary and secondary structure of RNA genes. In order to achieve this, we study the known primary and secondary structures of a comprehensive data set of 361 RNA genes as well as a set of 48 RNA sequences that are known to differ from the originally transcribed sequence units. We detect co-transcriptional folding by defining two measures of directedness which quantify the extend of asymmetry between alternative helices that lie 5' and those that lie 3' of the known helices with which they compete. RESULTS: We show with statistical significance that co-transcriptional folding strongly influences RNA sequences in two ways: (1) alternative helices that would compete with the formation of the functional structure during co-transcriptional folding are suppressed and (2) the formation of transient structures which may serve as guidelines for the co-transcriptional folding pathway is encouraged. CONCLUSIONS: These findings have a number of implications for RNA secondary structure prediction methods and the detection of RNA genes. PMID- 15298704 TI - Similarities in transcription factor IIIC subunits that bind to the posterior regions of internal promoters for RNA polymerase III. AB - BACKGROUND: In eukaryotes, RNA polymerase III (RNAP III) transcribes the genes for small RNAs like tRNAs, 5S rRNA, and several viral RNAs, and short interspersed repetitive elements (SINEs). The genes for these RNAs and SINEs have internal promoters that consist of two regions. These two regions are called the A and B blocks. The multisubunit transcription factor TFIIIC is required for transcription initiation of RNAP III; in transcription of tRNAs, the B-block binding subunit of TFIIIC recognizes a promoter. Although internal promoter sequences are conserved in eukaryotes, no evidence of homology between the B block binding subunits of vertebrates and yeasts has been reported previously. RESULTS: Here, I reported the results of PSI-BLAST searches using the B-block binding subunits of human and Shizosacchromyces pombe as queries, showing that the same Arabidopsis proteins were hit with low E-values in both searches. Comparison of the convergent iterative alignments obtained by these PSI-BLAST searches revealed that the vertebrate, yeast, and Arabidopsis proteins have similarities in their N-terminal one-third regions. In these regions, there were three domains with conserved sequence similarities, one located in the N-terminal end region. The N-terminal end region of the B-block binding subunit of Saccharomyces cerevisiae is tentatively identified as a HMG box, which is the DNA binding motif. Although I compared the alignment of the N-terminal end regions of the B-block binding subunits, and their homologs, with that of the HMG boxes, it is not clear whether they are related. CONCLUSION: Molecular phylogenetic analyses using the small subunit rRNA and ubiquitous proteins like actin and alpha-tubulin, show that fungi are more closely related to animals than either is to plants. Interestingly, the results obtained in this study show that, with respect to the B-block binding subunits of TFIIICs, animals appear to be evolutionarily closer to plants than to fungi. PMID- 15298705 TI - A single amino acid determines preference between phospholipids and reveals length restriction for activation of the S1P4 receptor. AB - BACKGROUND: Sphingosine-1-phosphate and lysophosphatidic acid (LPA) are ligands for two related families of G protein-coupled receptors, the S1P and LPA receptors, respectively. The lysophospholipid ligands of these receptors are structurally similar, however recognition of these lipids by these receptors is highly selective. A single residue present within the third transmembrane domain (TM) of S1P receptors is thought to determine ligand selectivity; replacement of the naturally occurring glutamic acid with glutamine (present at this position in the LPA receptors) has previously been shown to be sufficient to change the specificity of S1P1 from S1P to 18:1 LPA. RESULTS: We tested whether mutation of this "ligand selectivity" residue to glutamine could confer LPA-responsiveness to the related S1P receptor, S1P4. This mutation severely affected the response of S1P4 to S1P in a [35S]GTP gamma S binding assay, and imparted sensitivity to LPA species in the order 14:0 LPA > 16:0 LPA > 18:1 LPA. These results indicate a length restriction for activation of this receptor and demonstrate the utility of using LPA-responsive S1P receptor mutants to probe binding pocket length using readily available LPA species. Computational modelling of the interactions between these ligands and both wild type and mutant S1P4 receptors showed excellent agreement with experimental data, therefore confirming the fundamental role of this residue in ligand recognition by S1P receptors. CONCLUSIONS: Glutamic acid in the third transmembrane domain of the S1P receptors is a general selectivity switch regulating response to S1P over the closely related phospholipids, LPA. Mutation of this residue to glutamine confers LPA responsiveness with preference for short-chain species. The preference for short chain LPA species indicates a length restriction different from the closely related S1P1 receptor. PMID- 15298706 TI - Expression of PEG11 and PEG11AS transcripts in normal and callipyge sheep. AB - BACKGROUND: The callipyge mutation is located within an imprinted gene cluster on ovine chromosome 18. The callipyge trait exhibits polar overdominant inheritance due to the fact that only heterozygotes inheriting a mutant paternal allele (paternal heterozygotes) have a phenotype of muscle hypertrophy, reduced fat and a more compact skeleton. The mutation is a single A to G transition in an intergenic region that results in the increased expression of several genes within the imprinted cluster without changing their parent-of-origin allele specific expression. RESULTS: There was a significant effect of genotype (p < 0.0001) on the transcript abundance of DLK1, PEG11, and MEG8 in the muscles of lambs with the callipyge allele. DLK1 and PEG11 transcript levels were elevated in the hypertrophied muscles of paternal heterozygous animals relative to animals of the other three genotypes. The PEG11 locus produces a single 6.5 kb transcript and two smaller antisense strand transcripts, referred to as PEG11AS, in skeletal muscle. PEG11AS transcripts were detectable over a 5.5 kb region beginning 1.2 kb upstream of the PEG11 start codon and spanning the entire open reading frame. Analysis of PEG11 expression by quantitative PCR shows a 200-fold induction in the hypertrophied muscles of paternal heterozygous animals and a 13-fold induction in homozygous callipyge animals. PEG11 transcripts were 14-fold more abundant than PEG11AS transcripts in the gluteus medius of paternal heterozygous animals. PEG11AS transcripts were expressed at higher levels than PEG11 transcripts in the gluteus medius of animals of the other three genotypes. CONCLUSIONS: The effect of the callipyge mutation has been to alter the expression of DLK1, GTL2, PEG11 and MEG8 in the hypertrophied skeletal muscles. Transcript abundance of DLK1 and PEG11 was highest in paternal heterozygous animals and exhibited polar overdominant gene expression patterns; therefore, both genes are candidates for causing skeletal muscle hypertrophy. There was unique relationship of PEG11 and PEG11AS transcript abundance in the paternal heterozygous animals that suggests a RNA interference mechanism may have a role in PEG11 gene regulation and polar overdominance in callipyge sheep. PMID- 15298707 TI - Colorectal cancers with microsatellite instability display mRNA expression signatures characteristic of increased immunogenicity. AB - BACKGROUND: Colorectal cancers displaying high-degree microsatellite instability (MSI-H) have an improved prognosis compared to microsatellite stable (MSS) cancers. The observation of pronounced lymphocytic infiltrates suggests that MSI H cancers are inherently more immunogenic. We aimed to compare the gene expression profiles of MSI-H and MSS cancers to provide evidence for an activated immune response in the former. RESULTS: We analysed tissue from 133 colorectal cancer patients with full consent and Local Ethics Committee approval. Genomic DNA was analysed for microsatellite instability in BAT-26. High-quality RNA was used for microarray analysis on the Affymetrix HG-U133A chip. Data was analysed on GeneSpring software version 6.0. Confirmatory real-time RT-PCR was performed on 28 MSI-H and 26 MSS cancers. A comparison of 29 MSI-H and 104 MSS cancers identified 2070 genes that were differentially expressed between the two groups [P < 0.005]. Significantly, many key immunomodulatory genes were up-regulated in MSI-H cancers. These included antigen chaperone molecules (HSP-70, HSP-110, Calreticulin, gp96), pro-inflammatory cytokines (Interleukin (IL)-18, IL-15, IL 8, IL-24, IL-7) and cytotoxic mediators (Granulysin, Granzyme A). Quantitative RT PCR confirmed up-regulation of HSP-70 [P = 0.016], HSP-110 [P = 0.002], IL-18 [P = 0.004], IL-8 [0.002] and Granulysin [P < 0.0001]. CONCLUSIONS: The upregulation of a large number of genes implicated in immune response supports the theory that MSI-H cancers are immunogenic. The novel observation of Heat Shock Protein up regulation in MSI-H cancer is highly significant in light of the recognised roles of these proteins in innate and antigen-specific immunogenicity. Increased mRNA levels of pro-inflammatory cytokines and cytotoxic mediators also indicate an activated anti-tumour immune response. PMID- 15298709 TI - Mapping the distribution of Loa loa in Cameroon in support of the African Programme for Onchocerciasis Control. AB - BACKGROUND: Loa loa has recently emerged as a filarial worm of significant public health importance as a consequence of its impact on the African Programme for Onchocerciasis Control (APOC). Severe, sometimes fatal, encephalopathic reactions to ivermectin (the drug of choice for onchocerciasis control) have occurred in some individuals with high Loa loa microfilarial counts. Since high density of Loa loa microfilariae is known to be associated with high prevalence rates, a distribution map of the latter may determine areas where severe reactions might occur. The aim of the study was to identify variables which were significantly associated with the presence of a Loa microfilaraemia in the subjects examined, and to develop a spatial model predicting the prevalence of the Loa microfilaraemia. METHODS: Epidemiological data were collected from 14,225 individuals living in 94 villages in Cameroon, and analysed in conjunction with environmental data. A series of logistic regression models (multivariate analysis) was developed to describe variation in the prevalence of Loa loa microfilaraemia using individual level co-variates (age, sex, microl of blood taken for examination) and village level environmental co-variates (including altitude and satellite-derived vegetation indices). RESULTS: A spatial model of Loa loa prevalence was created within a geographical information system. The model was then validated using an independent data set on Loa loa distribution. When considering both data sets as a whole, and a prevalence threshold of 20%, the sensitivity and the specificity of the model were 81.7 and 69.4%, respectively. CONCLUSIONS: The model developed has proven very useful in defining the areas at risk of post-ivermectin Loa-related severe adverse events. It is now routinely used by APOC when projects of community-directed treatment with ivermectin are examined. PMID- 15298708 TI - Thermal modeling of lesion growth with radiofrequency ablation devices. AB - BACKGROUND: Temperature is a frequently used parameter to describe the predicted size of lesions computed by computational models. In many cases, however, temperature correlates poorly with lesion size. Although many studies have been conducted to characterize the relationship between time-temperature exposure of tissue heating to cell damage, to date these relationships have not been employed in a finite element model. METHODS: We present an axisymmetric two-dimensional finite element model that calculates cell damage in tissues and compare lesion sizes using common tissue damage and iso-temperature contour definitions. The model accounts for both temperature-dependent changes in the electrical conductivity of tissue as well as tissue damage-dependent changes in local tissue perfusion. The data is validated using excised porcine liver tissues. RESULTS: The data demonstrate the size of thermal lesions is grossly overestimated when calculated using traditional temperature isocontours of 42 degrees C and 47 degrees C. The computational model results predicted lesion dimensions that were within 5% of the experimental measurements. CONCLUSION: When modeling radiofrequency ablation problems, temperature isotherms may not be representative of actual tissue damage patterns. PMID- 15298710 TI - Medical Students' and Residents' preferred site characteristics and preceptor behaviours for learning in the ambulatory setting: a cross-sectional survey. AB - BACKGROUND: Medical training is increasingly occurring in the ambulatory setting for final year medical students and residents. This study looks to identify if gender, school, level of training, or specialty affects learner's (final year medical students and residents) preferred site characteristics and preceptor behaviours for learning in the ambulatory setting. METHODS: All final year medical students and residents at the five medical schools in Ontario (N = 3471) were surveyed about the site characteristics and preceptor behaviours most enhancing their learning in the ambulatory setting. Preferred site characteristics and preceptor behaviours were rank ordered. Factor analysis grouped the site characteristics and preceptor behaviours into themes which were then correlated with gender, school, level of training, and specialty. RESULTS: Having an adequate number and variety of patients while being supervised by enthusiastic preceptors who give feedback and are willing to discuss their reasoning processes and delegate responsibility are site characteristics and preceptor behaviours valued by almost all learners. Some teaching strategies recently suggested to improve efficiency in the ambulatory teaching setting, such as structuring the interview for the student and teaching and reviewing the case in front of the patient, were found not to be valued by learners. There was a striking degree of similarity in what was valued by all learners but there were also some educationally significant differences, particularly between learners at different levels and in different specialties. Key findings between the different levels include preceptor interaction being most important for medical students as opposed to residents who most value issues pertaining to patient logistics. Learning resources are less valued early and late in training. Teaching and having the case reviewed in front of the patient becomes increasingly less valued as learners advance in their training. As one approaches the end of ones' training office management instruction becomes increasingly valued. Differences between specialties pertain most to the type of practice residents will ultimately end up in (ie: office based specialties particularly valuing instruction in office management and health care system interaction). CONCLUSIONS: Preceptors need to be aware of, and make efforts to provide, teaching strategies such as feedback and discussing clinical reasoning, that learners have identified as being helpful for learning. If strategies identified as not being valued for learning, such as teaching in front of the patient, must continue it will be important to explore the barriers they present to learning. Although what all learners want from their preceptors and clinic settings to enhance their learning is remarkably similar, being aware of the educationally significant differences, particularly for learners at different levels and in different specialties, will enhance teaching in the ambulatory setting. PMID- 15298711 TI - Recovery of frog and lizard communities following primary habitat alteration in Mizoram, Northeast India. AB - BACKGROUND: Community recovery following primary habitat alteration can provide tests for various hypotheses in ecology and conservation biology. Prominent among these are questions related to the manner and rate of community assembly after habitat perturbation. Here we use space-for-time substitution to analyse frog and lizard community assembly along two gradients of habitat recovery following slash and burn agriculture (jhum) in Mizoram, Northeast India. One recovery gradient undergoes natural succession to mature tropical rainforest, while the other involves plantation of jhum fallows with teak Tectona grandis monoculture. RESULTS: Frog and lizard communities accumulated species steadily during natural succession, attaining characteristics similar to those from mature forest after 30 years of regeneration. Lizards showed higher turnover and lower augmentation of species relative to frogs. Niche based classification identified a number of guilds, some of which contained both frogs and lizards. Successional change in species richness was due to increase in the number of guilds as well as the number of species per guild. Phylogenetic structure increased with succession for some guilds. Communities along the teak plantation gradient on the other hand, did not show any sign of change with chronosere age. Factor analysis revealed sets of habitat variables that independently determined changes in community and guild composition during habitat recovery. CONCLUSIONS: The timescale of frog and lizard community recovery was comparable with that reported by previous studies on different faunal groups in other tropical regions. Both communities converged on primary habitat attributes during natural vegetation succession, the recovery being driven by deterministic, nonlinear changes in habitat characteristics. On the other hand, very little faunal recovery was seen even in relatively old teak plantation. In general, tree monocultures are unlikely to support recovery of natural forest communities and the combined effect of shortened jhum cultivation cycles and plantation forestry could result in landscapes without mature forest. Lack of source pools of genetic diversity will then lead to altered vegetation succession and faunal community reassembly. It is therefore important that the value of habitat mosaics containing even patches of primary forest and successional secondary habitats be taken into account. PMID- 15298712 TI - The National Women's Health Study: assembly and description of a population-based reproductive cohort. AB - BACKGROUND: Miscarriage is a common event but is remarkably difficult to measure in epidemiological studies. Few large-scale population-based studies have been conducted in the UK. METHODS: This was a population-based two-stage postal survey of reproductive histories of adult women living in the United Kingdom in 2001, sampled from the electronic electoral roll. In Stage 1 a short "screening" questionnaire was sent to over 60,000 randomly selected women in order to identify those aged 55 and under who had ever been pregnant or ever attempted to achieve a pregnancy, from whom a brief reproductive history was requested. Stage 2 involved a more lengthy questionnaire requesting detailed information on every pregnancy (and fertility problems), and questions relating to socio-demographic, behavioural and other factors for the most recent pregnancy in order to examine risk factors for miscarriage. Data on stillbirth, multiple birth and maternal age are compared to national data in order to assess response bias. RESULTS: The response rate was 49% for Stage 1 and 73% for the more targeted Stage 2. A total of 26,050 questionnaires were returned in Stage 1. Of the 17,748 women who were eligible on the grounds of age, 27% reported that they had never been pregnant and had never attempted to conceive a child. The remaining 13,035 women reported a total of 30,661 pregnancies. Comparison of key reproductive indicators (stillbirth and multiple birth rates and maternal age at first birth) with national statistics showed that the data look remarkably similar to the general population. CONCLUSIONS: This study has enabled the assembly of a large population-based dataset of women's reproductive histories which appears unbiased compared to the general UK population and which will enable investigation of hard to-measure outcomes such as miscarriage and infertility. PMID- 15298713 TI - Logistics of community smallpox control through contact tracing and ring vaccination: a stochastic network model. AB - BACKGROUND: Previous smallpox ring vaccination models based on contact tracing over a network suggest that ring vaccination would be effective, but have not explicitly included response logistics and limited numbers of vaccinators. METHODS: We developed a continuous-time stochastic simulation of smallpox transmission, including network structure, post-exposure vaccination, vaccination of contacts of contacts, limited response capacity, heterogeneity in symptoms and infectiousness, vaccination prior to the discontinuation of routine vaccination, more rapid diagnosis due to public awareness, surveillance of asymptomatic contacts, and isolation of cases. RESULTS: We found that even in cases of very rapidly spreading smallpox, ring vaccination (when coupled with surveillance) is sufficient in most cases to eliminate smallpox quickly, assuming that 95% of household contacts are traced, 80% of workplace or social contacts are traced, and no casual contacts are traced, and that in most cases the ability to trace 1 5 individuals per day per index case is sufficient. If smallpox is assumed to be transmitted very quickly to contacts, it may at times escape containment by ring vaccination, but could be controlled in these circumstances by mass vaccination. CONCLUSIONS: Small introductions of smallpox are likely to be easily contained by ring vaccination, provided contact tracing is feasible. Uncertainties in the nature of bioterrorist smallpox (infectiousness, vaccine efficacy) support continued planning for ring vaccination as well as mass vaccination. If initiated, ring vaccination should be conducted without delays in vaccination, should include contacts of contacts (whenever there is sufficient capacity) and should be accompanied by increased public awareness and surveillance. PMID- 15298714 TI - Treatment of retinopathy of prematurity with topical ketorolac tromethamine: a preliminary study. AB - BACKGROUND: Retinopathy of Prematurity (ROP) is a common retinal neovascular disorder of premature infants. It is of variable severity, usually heals with mild or no sequelae, but may progress to blindness from retinal detachments or severe retinal scar formation. This is a preliminary report of the effectiveness and safety of a new and original use of topical ketorolac in preterm newborn to prevent the progression of ROP to the more severe forms of this disease. METHODS: From January 2001 to December 2002, all fifty nine preterm newborns with birthweight less than 1250 grams or gestational age less than 30 weeks of gestational age admitted to neonatal intensive care were eligible for treatment with topical ketorolac (0.25 milligrams every 8 hours in each eye). The historical comparison group included all 53 preterm newborns, with the same inclusion criteria, admitted between January 1999 and December 2000. RESULTS: Groups were comparable in terms of weight distribution, Apgar score at 5 minutes, incidence of sepsis, intraventricular hemorrhage and necrotizing enterocolitis. The duration of oxygen therapy was significantly longer in the control group. In the ketorolac group, among 43 children that were alive at discharge, one (2.3%) developed threshold ROP and cryotherapy was necessary. In the comparison group 35 children survived, and six child (17%) needed cryotherapy (Relative Risk 0.14, 95%CI 0.00 to 0.80, p = 0.041). Adjusting by duration of oxygen therapy did not significantly change these results. Adverse effects attributable to ketorolac were not detected. CONCLUSIONS: This preliminary report suggests that ketorolac in the form of an ophthalmic solution can reduce the risk of developing severe ROP in very preterm newborns, without producing significant adverse side effects. These results, although promising, should be interpreted with caution because of the weakness of the study design. This is an inexpensive and simple intervention that might ameliorate the progression of a disease with devastating consequences for children and their families. We believe that next logical step would be to assess the effectiveness of this intervention in a randomized controlled trial of adequate sample size. PMID- 15298715 TI - Genome-wide array comparative genomic hybridization analysis reveals distinct amplifications in osteosarcoma. AB - BACKGROUND: Osteosarcoma is a highly malignant bone neoplasm of children and young adults. It is characterized by extremely complex karyotypes and high frequency of chromosomal amplifications. Currently, only the histological response (degree of necrosis) to therapy represent gold standard for predicting the outcome in a patient with non-metastatic osteosarcoma at the time of definitive surgery. Patients with lower degree of necrosis have a higher risk of relapse and poor outcome even after chemotherapy and complete resection of the primary tumor. Therefore, a better understanding of the underlying molecular genetic events leading to tumor initiation and progression could result in the identification of potential diagnostic and therapeutic targets. METHODS: We used a genome-wide screening method - array based comparative genomic hybridization (array-CGH) to identify DNA copy number changes in 48 patients with osteosarcoma. We applied fluorescence in situ hybridization (FISH) to validate some of amplified clones in this study. RESULTS: Clones showing gains (79%) were more frequent than losses (66%). High-level amplifications and homozygous deletions constitute 28.6% and 3.8% of tumor genome respectively. High-level amplifications were present in 238 clones, of which about 37% of them showed recurrent amplification. Most frequently amplified clones were mapped to 1p36.32 (PRDM16), 6p21.1 (CDC5L, HSPCB, NFKBIE), 8q24, 12q14.3 (IFNG), 16p13 (MGRN1), and 17p11.2 (PMP22 MYCD, SOX1,ELAC27). We validated some of the amplified clones by FISH from 6p12-p21, 8q23-q24, and 17p11.2 amplicons. Homozygous deletions were noted for 32 clones and only 7 clones showed in more than one case. These 7 clones were mapped to 1q25.1 (4 cases), 3p14.1 (4 cases), 13q12.2 (2 cases), 4p15.1 (2 cases), 6q12 (2 cases), 6q12 (2 cases) and 6q16.3 (2 cases). CONCLUSIONS: This study clearly demonstrates the utility of array CGH in defining high-resolution DNA copy number changes and refining amplifications. The resolution of array CGH technology combined with human genome database suggested the possible target genes present in the gained or lost clones. PMID- 15298716 TI - The oncogenic fusion protein RUNX1-CBFA2T1 supports proliferation and inhibits senescence in t(8;21)-positive leukaemic cells. AB - BACKGROUND: The fusion protein RUNX1-CBFA2T1 associated with t(8;21)-positive acute myeloid leukaemia is a potent inhibitor of haematopoetic differentiation. The role of RUNX1-CBFA2T1 in leukaemic cell proliferation is less clear. We examined the consequences of siRNA-mediated RUNX1-CBFA2T1 depletion regarding proliferation and clonogenicity of t(8;21)-positive cell lines. METHODS: The t(8;21)-positive cell line Kasumi-1 was electroporated with RUNX1-CBFA2T1 or control siRNAs followed by analysis of proliferation, colony formation, cell cycle distribution, apoptosis and senescence. RESULTS: Electroporation of Kasumi 1 cells with RUNX1-CBFA2T1 siRNAs, but not with control siRNAs, resulted in RUNX1 CBFA2T1 suppression which lasted for at least 5 days. A single electroporation with RUNX1-CBFA2T1 siRNA severely diminished the clonogenicity of Kasumi-1 cells. Prolonged RUNX1-CBFA2T1 depletion inhibited proliferation in suspension culture and G1-S transition during the cell cycle, diminished the number of apoptotic cells, but induced cellular senescence. The addition of haematopoetic growth factors could not rescue RUNX1-CBFA2T1-depleted cells from senescence, and could only partially restore their clonogenicity. CONCLUSIONS: RUNX1-CBFA2T1 supports the proliferation and expansion of t(8;21)-positive leukaemic cells by preventing cellular senescence. These findings suggest a central role of RUNX1-CBFA2T1 in the maintenance of the leukaemia. Therefore, RUNX1-CBFA2T1 is a promising and leukaemia-specific target for molecularly defined therapeutic approaches. PMID- 15298717 TI - The effect of hypertensive disorders in pregnancy on small for gestational age and stillbirth: a population based study. AB - BACKGROUND: Hypertensive disorders in pregnancy are leading causes of maternal, fetal and neonatal morbidity and mortality worldwide. However, studies attempting to quantify the effect of hypertension on adverse perinatal outcomes have been mostly conducted in tertiary centres. This population-based study explored the frequency of hypertensive disorders in pregnancy and the associated increase in small for gestational age (SGA) and stillbirth. METHODS: We used information on all pregnant women and births, in the Canadian province of Nova Scotia, between 1988 and 2000. Pregnancies were excluded if delivery occurred < 20 weeks, if birthweight was < 500 grams, if there was a high-order multiple pregnancy (greater than twin gestation), or a major fetal anomaly. RESULTS: The study population included 135,466 pregnancies. Of these, 7.7% had mild pregnancy induced hypertension (PIH), 1.3% had severe PIH, 0.2% had HELLP (hemolysis, elevated liver enzymes, low platelets), 0.02% had eclampsia, 0.6% had chronic hypertension, and 0.4% had chronic hypertension with superimposed PIH. Women with any hypertension in pregnancy were 1.6 (95% CI 1.5-1.6) times more likely to have a live birth with SGA and 1.4 (95% CI 1.1-1.8) times more likely to have a stillbirth as compared with normotensive women. Adjusted analyses showed that women with gestational hypertension without proteinuria (mild PIH) and with proteinuria (severe PIH, HELLP, or eclampsia) were more likely to have infants with SGA (RR 1.5, 95% CI 1.4-1.6 and RR 3.2, 95% CI 2.8-3.6, respectively). Women with pre-existing hypertension were also more likely to give birth to an infant with SGA (RR 2.5, 95% CI 2.2-3.0) or to have a stillbirth (RR 3.2, 95% CI 1.9 5.4). CONCLUSIONS: This large, population-based study confirms and quantifies the magnitude of the excess risk of small for gestational age and stillbirth among births to women with hypertensive disease in pregnancy. PMID- 15298718 TI - Misoprostol for treating postpartum haemorrhage: a randomized controlled trial [ISRCTN72263357]. AB - BACKGROUND: Postpartum haemorrhage remains an important cause of maternal death despite treatment with conventional therapy. Uncontrolled studies and one randomised comparison with conventional oxytocics have reported dramatic effects with high-dose misoprostol, usually given rectally, for treatment of postpartum haemorrhage, but this has not been evaluated in a placebo-controlled trial. METHODS: The study was conducted at East London Hospital Complex, Tembisa and Chris Hani Baragwanath Hospitals, South Africa. Routine active management of the third stage of labour was practised. Women with more than usual postpartum bleeding thought to be related to inadequate uterine contraction were invited to participate, and to sign informed consent. All routine treatment was given from a special 'Postpartum Haemorrhage Trolley'. In addition, participants who consented were enrolled by drawing the next in a series of randomised treatment packs containing either misoprostol 5 x 200 microg or similar placebo, which were given 1 orally, 2 sublingually and 2 rectally. RESULTS: With misoprostol there was a trend to reduced blood loss >/=500 ml in 1 hour after enrolment measured in a flat plastic 'fracture bedpan', the primary outcome (6/117 vs 11/120, relative risk 0.56; 95% confidence interval 0.21 to 1.46). There was no difference in mean blood loss or haemoglobin level on day 1 after birth < 6 g/dl or blood transfusion. Side-effects were increased, namely shivering (63/116 vs 30/118; 2.14, 1.50 to 3.04) and pyrexia > 38.5 degrees C (11/114 vs 2/118; 5.69, 1.29 to 25). In the misoprostol group 3 women underwent hysterectomy of whom 1 died, and there were 2 further maternal deaths. CONCLUSIONS: Because of a lower than expected incidence of the primary outcome in the placebo group, the study was underpowered. We could not confirm the dramatic effect of misoprostol reported in several unblinded studies, but the results do not exclude a clinically important effect. Larger studies are needed to assess substantive outcomes and risks before misoprostol enters routine use. PMID- 15298719 TI - Epidemiology of the human immunodeficiency virus in Saudi Arabia; 18-year surveillance results and prevention from an Islamic perspective. AB - BACKGROUND: data on HIV epidemiology and preventive measures in Islamic countries is limited. This study describes the results of 18-year of HIV surveillance in Saudi Arabia (SA) and the preventive measures implemented from an Islamic perspective. METHODS: surveillance for HIV has been underway in SA since 1984. Indications for HIV testing include clinical suspicion, screening of contacts of HIV-infected patients, and routine screening of blood and organ donors, prisoners, intravenous drug users, patients with other sexually transmitted infections, and expatriates pre-employment. This is a case series descriptive study of all confirmed HIV infections diagnosed in SA from 1984 through 2001. RESULTS: a total of 6046 HIV infections were diagnosed, of which 1285 (21.3%) cases were Saudi citizens. Over the 18-year surveillance period the number of HIV infections diagnosed annually among Saudi citizens gradually increased and, over the period 1997-2001, it reached to 84 to 142 cases per year. The number of cases per 100,000 population varied widely between regions with a maximum of 74 cases and a minimum of 2 cases. The infection was most common in the age group 20-40 years (74.6%) and predominantly affected men (71.6%). The modes of transmission among Saudi citizens and expatriates, respectively, were as follows: heterosexual contact, 487 (37.9%) and 1352 (28.4%) cases; blood transfusion, 322 (25.0%) and 186 (3.9%) cases; perinatal transmission, 83 (6.5%) and 19 (0.4%) cases; homosexual contact, 32 (2.5%) and 38 (0.8%) cases; intravenous drug use, 17 (1.3%) and 33 (0.7%) cases; bisexual contact, 10 (0.8%) and 14 (0.3%) cases; unknown, 334 (26.0%) and 3119 (65.5%) cases. The number of HIV infections transmitted by blood or blood products transfusion declined to zero by year 2001 and all such infections occurred due to transfusions administered before 1986. At HIV diagnosis, 4502/6046 (74.5%) patients had no symptoms, 787 (13.0%) patients had non-AIDS defining manifestations, and 757 (12.5%) patients had AIDS. A total of 514/1285 (40%) Saudi patients died by year 2001. CONCLUSIONS: the number of HIV cases in SA is limited with heterosexual contact being the main mode of transmission. From an Islamic perspective, preventive strategies include prevention of non-marital sex and intravenous drug use with encouragement of "safe sex" through legal marriage. PMID- 15298720 TI - Nitric oxide-an endogenous inhibitor of gastric acid secretion in isolated human gastric glands. AB - BACKGROUND: Endothelial nitric oxide synthase (eNOS) has previously been detected in the glandular part of the human gastric mucosa. Furthermore, nitric oxide (NO) has been shown to influence gastric secretion in various animal models. The present study was conducted to investigate the influence of exogenously and endogenously derived NO on histamine- and cAMP-stimulated gastric acid secretion in isolated human oxyntic glands. METHODS: Oxyntic glands were isolated from human gastric biopsies and were subsequently pre-treated with NO donors and nitric oxide synthase inhibitors and then exposed to histamine or dibutyryl-cAMP (db-cAMP). The secretory response of the glands was determined as accumulation of [14C]aminopyrine. RESULTS: The histamine- or db-cAMP-induced acid secretion was attenuated by L-arginine, a known source of endogenous NO, and also by the NO donors sodium nitroprusside (SNP) and S-nitroso-N-acetyl-penicillamine (SNAP). Pre-treatment with either of the NOS inhibitors NG-nitro-L-arginine methyl ester (L-NAME) or NG-nitro-L-arginine (L-NNA) enhanced the secretory response. CONCLUSION: Our results show that NO inhibits gastric acid secretion in isolated human gastric glands, and that there is endogenous formation of NO within the glandular epithelium in the vicinity of the parietal cells. PMID- 15298721 TI - Primary care follow up of patients discharged from the emergency department: a retrospective study. AB - BACKGROUND: The visit to the emergency department (ED) constitutes a brief, yet an important point in the continuum of medical care. The aim of our study was to evaluate the continuity of care of adult ED visitors. METHODS: We retrospectively reviewed all ED discharge summaries for over a month 's period. The ED chart, referral letter and the patient's primary care file were reviewed. Data collected included: age, gender, date and hour of ED visit, documentation of ED referral and ED discharge letter in the primary care file. RESULTS: 359 visits were eligible for the study. 192 (53.5%) of the patients were women, average age 54.1 +/- 18.7 years (mean +/- SD). 214 (59.6%) of the visits were during working hours of primary care clinics ("working hours"), while the rest were "out of hours" visits. Only 196 (54.6%) of patients had a referral letter, usually from their family physician. A third (71/214) of "working hours" visits were self referrals, the rate rose to 63.5% (92/145) of "out of hours" visits (p < 0.0001). The ED discharge letter was found in 50% (179/359) of the primary care files. A follow up visit was documented in only 31% (111/359). Neither follow up visit nor discharge letter were found in 43% of the files (153/359). CONCLUSIONS: We have found a high rate of ED self referrals throughout the day together with low documentation rates of ED visits in the primary care charts. Our findings point to a poor continuity of care of ED attendees. PMID- 15298722 TI - Monoclonal antibodies as effective therapeutic agents for solid tumors. AB - Monoclonal antibodies (mAbs) against growth factors or their receptors have been revealed to be effective therapeutic agents for solid tumors. Trastuzumab (humanized anti-HER2 mAb) is the first mAb approved for the treatment of a solid tumor, metastatic breast cancer. Large-scale phase III clinical trials are now ongoing to further evaluate the additive effects on chemotherapy and the efficacy as a maintenance monotherapy. Another anti-HER2 mAb CH401 that we developed also seems to have good potential. This chimeric mAb completely suppressed the growth of established human tumor xenografts in SCID mice after a single injection. Furthermore, CH401 characteristically showed much stronger induction of apoptosis in HER2-overexpressing gastric cancer cells compared to trastuzumab. Additional targets now being intensively evaluated are epidermal growth factor receptor (EGFR) and vascular endothelial growth factor (VEGF). Both cetuximab (chimeric anti-EGFR mAb) and bevacizumab (humanized anti-VEGF mAb) have recently been shown to be of clinical value for metastatic colorectal cancer. Anti-idiotype mAbs are unique as active immunotherapeutic agents, and survival benefits have been observed in clinical trials for solid tumors. PMID- 15298723 TI - ETS transcription factors: possible targets for cancer therapy. AB - Ets family (ETS) transcription factors, characterized by an evolutionally conserved Ets domain, play important roles in cell development, cell differentiation, cell proliferation, apoptosis and tissue remodeling. Most of them are downstream nuclear targets of Ras-MAP kinase signaling, and the deregulation of ETS genes results in the malignant transformation of cells. Several ETS genes are rearranged in human leukemia and Ewing tumors to produce chimeric oncoproteins. Furthermore, the aberrant expression of several ETS genes is often observed in various types of human malignant tumors. Considering that some ETS transcription factors are involved in malignant transformation and tumor progression, including invasion, metastasis and neo-angiogenesis through the activation of cancer-related genes, they could be potential molecular targets for selective cancer therapy. PMID- 15298724 TI - Detection of a mouse OGG1 fragment during caspase-dependent apoptosis: oxidative DNA damage and apoptosis. AB - We investigated the expression of mouse 8-oxoguanine DNA glycosylase 1 (mOGG1) in mouse non-parenchymal hepatocytes (NCTC) during etoposide- or mitomycin C (MMC) induced apoptosis. We observed mOGG1 fragmentation in apoptotic cells. The apoptosis accompanying the fragmentation of mOGG1 was caspase-dependent. The mOGG1 fragment existed in both the cytoplasm and nucleus of the etoposide-treated NCTC, indicating that the mOGG1 fragment could be transferred into the nucleus. In addition, 8-hydroxyguanine (8-OH-Gua, 7,8-dihydro-8-oxoguanine) accumulated in the DNA of NCTC treated with etoposide, suggesting that the mOGG1 fragment might not function as a normal repair enzyme in etoposide-treated NCTC. Although we have not clarified in detail the mechanism and the significance of the mOGG1 fragmentation, further study of the fragmentation of DNA repair enzymes might provide insights into the relationship between oxidative DNA damage and apoptosis. PMID- 15298725 TI - Tumorigenesis facilitated by Pten deficiency in the skin: evidence of p53-Pten complex formation on the initiation phase. AB - Pten, a tumor suppressor gene, is mutated in various human cancers and in hereditary cancer syndromes, such as Cowden disease. We have previously developed a knockout mouse in which Pten is specifically disrupted in the skin, resulting in hyperproliferation and spontaneous tumorigenesis of the skin keratinocytes. In this study, we further clarified the effects of Pten deficiency in tumorigenesis, by using a two-step model in intact skin of Pten knockout mouse. Although the conventional protocol requires serial exposures to DMBA and TPA, mice deficient for Pten developed skin papilloma within 6 weeks after a single exposure to DMBA, indicating that loss of Pten has a tumor-promoting effect. Serial exposure to DMBA-TPA ointments produced 10-fold more papillomas in the skin of knockout mice than in the wild-type counterpart, suggesting an increased rate of initiation. Therefore, we precisely examined the effect of DMBA. This treatment was highly apoptotic in wild-type mice, whereas the number of apoptotic cells was diminished in Pten-deficient skin. Moreover, primary keratinocytes isolated from Pten deficient mice were also resistant to the apoptotic effect of DMBA. The status of p53, Pten proteins and downstream targets of p53, such as p21, 14-3-3, and Reprimo, were also examined, and we found that accumulation of p53 protein and up regulation of p53 targets were delayed in Pten-knockout skin. These observations suggest that Pten is involved in rapid recruitment of p53 in the tumor initiation phase. PMID- 15298726 TI - Reactive oxygen species (ROS) control the expression of Bcl-2 family proteins by regulating their phosphorylation and ubiquitination. AB - We examined the influence of ROS on the phosphorylation and complex formation of Bcl-2 family proteins in Mn-superoxide dismutase (SOD) antisense-transfected squamous cell carcinoma cells, OSC-4 cells. The increase of intracellular ROS level induced by cis-diamminedichloroplatinum (CDDP) and gamma-ray treatment was greater in antisense-transfected cells than in control vector-transfected cells, and apoptosis was more extensively induced in the former. Antisense-transfected cells expressed high levels of Bax and Bak, but low levels of Bcl-2 and Bcl-XL when treated with CDDP, peplomycin, 5-fluorouracil or gamma-rays. After treatment with these agents, the phosphorylation of protein kinase A, Bcl-2 (Thr56) and Bad (Ser155) was increased, especially in antioxidant (N-acetylcysteine and pyrrolidine dithiocarbamate)-pretreated control cells, but the phosphorylation levels were very low in the antisense-transfected cells. Bcl-2 ubiquitination was increased, but ubiquitination of Bad and Bax was decreased in the antisense transfected cells, although their ubiquitination was increased by the antioxidants. These results reveal that ROS induce apoptosis by regulating the phosphorylation and ubiquitination of Bcl-2 family proteins, resulting in increased proapoptotic protein levels and decreased antiapoptotic protein expression. PMID- 15298727 TI - Alterations of the p16INK4a/p14ARF pathway in clear cell sarcoma. AB - Clear cell sarcoma (CCS) is a very rare soft tissue sarcoma with a poor prognosis. It has become apparent through immunohistochemical, ultrastructural, and microarray analyses that CCS is a soft tissue melanocytic neoplasm. Alterations in the p16INK4a/p14ARF gene are common in malignant melanoma, which is the prototypical melanocytic neoplasm. In the present study, we performed a clinicopathologic analysis and investigated p16 and cyclin D1 expression by immunohistochemistry in 14 cases. Furthermore, we investigated genetic changes of various tumor suppressor genes and an oncogene, including p16INK4a/p14ARF, p53, beta-catenin, and APC, in 11 cases. The 5-year overall survival rate in all the patients was 33.3%. A high mitotic rate was a significant adverse prognostic factor (P = 0.004). Decreased expression of p16 was observed in 4 (28.6%) of 14 cases. Overexpression of cyclin D1 was observed in 9 cases (64.3%). SSCP analysis followed by DNA direct sequencing revealed point mutations of the p16INK4a gene in 2 of 11 cases (18.2%). In addition, one case with the p14ARF mutation and 2 cases with the p53 mutation were observed. None of the cases harbored mutation of the beta-catenin or APC gene. Homozygous deletion of the p16INK4a/p14ARF gene was detected in one case. Methylation-specific PCR did not reveal hypermethylation of the p16INK4a/p14ARF promoter region in any of the cases. Three cases harbored genetic alterations of the p16INK4a/p14ARF gene (27.3%). All tumors with genetic alterations of the p16INK4a/p14ARF or p53 gene showed a high mitotic rate or tumor necrosis. These alterations were considered to be influential in the poor prognosis of CCS patients. PMID- 15298728 TI - Biophoton detection as a novel technique for cancer imaging. AB - Biophoton emission is defined as extremely weak light that is radiated from any living system due to its metabolic activities, without excitation or enhancement. We measured biophoton images of tumors transplanted in mice with a highly sensitive and ultra-low noise CCD camera system. Cell lines employed for this study were AH109A, TE4 and TE9. Biophoton images of each tumor were measured 1 week after carcinoma cell transplantation to estimate the tumor size at week 1 and the biophoton intensity. Some were also measured at 2 and 3 weeks to compare the biophoton distribution with histological findings. We achieved sequential biophoton imaging during tumor growth for the first time. Comparison of microscopic findings and biophoton intensity suggested that the intensity of biophoton emission reflects the viability of the tumor tissue. The size at week 1 differed between cell lines, and the biophoton intensity of the tumor was correlated with the tumor size at week 1 (correlation coefficient 0.73). This non invasive and simple technique has the potential to be used as an optical biopsy to detect tumor viability. PMID- 15298729 TI - Dominant-negative Stat5 inhibits growth and induces apoptosis in T47D-derived tumors in nude mice. AB - Signal transducer and activator of transcription 5 (Stat5) regulates growth, differentiation, and survival of mammary and hematopoietic cells. The role of Stat5 in breast cancer has not been established, although Stat5 is critical for some hematopoietic malignancies. In this study, we have analyzed the role of Stat5 in progression of the estrogen receptor-positive T47D human breast cancer cell line, in which Stat5b is constitutively activated. Expression of Stat5 regulated genes, such as cyclin D1 and bcl-xL, was strongly suppressed in T47D cells infected with the dominant-negative Stat5 adenovirus, AdStat5aDelta740. We also determined the phenotypic effects of introduction of dominant-negative Stat5 on T47D-derived tumors in nude mice. Tumors injected with AdStat5aDelta740 showed a 60% reduction in size, which was associated with the induction of apoptosis. Our results indicate the possibility of using dominant-negative Stat5 to induce apoptosis in certain Stat5-activated breast cancers. PMID- 15298730 TI - Involvement of cell cycle regulatory proteins and MAP kinase signaling pathway in growth inhibition and cell cycle arrest by a selective cyclooxygenase 2 inhibitor, etodolac, in human hepatocellular carcinoma cell lines. AB - Recent studies have shown that selective cyclooxygenase-2 (COX-2) inhibitors induce growth inhibition and cell cycle arrest in hepatocellular carcinoma (HCC) cell lines. However, the mechanism by which COX-2 inhibitors regulate the cell cycle and whether or not growth signal pathways are involved in the growth inhibition remain unclear. In this study, we investigated the mechanisms of growth inhibition and cell cycle arrest by etodolac, a selective COX-2 inhibitor, in HCC cell lines, HepG2 and PLC/PRF/5, by studying cell cycle regulatory proteins, and the MAP kinase and PDK1-PKB/AKT signaling pathways. Etodolac inhibited growth and PCNA expression and induced cell cycle arrest in both HCC cell lines. Etodolac induced p21WAF1/Cip1 and p27Kip1 expression and inhibited CDK2, CDK4, CDC2, cyclin A and cyclin B1 expression, but did not affect cyclin D1 or cyclin E. HGF and 10% FBS induced ERK phosphorylation, but phosphorylation of p38, JNK and AKT was down-regulated by etodolac. PD98059, a selective inhibitor of ERK phosphorylation, induced growth inhibition, the expression of p27Kip1 and cell cycle arrest. In conclusion, p21WAF1/Cip1, p27Kip1, CDK2, CDK4, CDC2, cyclin A, cyclin B1 and the MAP kinase signaling pathway are involved in growth inhibition and cell cycle arrest by a selective COX-2 inhibitor in HCC cell lines. PMID- 15298731 TI - Bowel movement frequency, medical history and the risk of gallbladder cancer death: a cohort study in Japan. AB - Few risk factors for gallbladder cancer have been identified with sufficient statistical power, because this cancer is rare. The present study was conducted to evaluate the association of bowel movement frequency and medical history with the risk of death from gallbladder cancer using the data set from a large-scale cohort study. A total of 113,394 participants (42.0% males), aged 40 to 89 years, were followed up for 11 years. Information on the medical history of selected diseases, history of blood transfusions, frequency of stools, and tendency toward diarrhea at baseline was collected through a self-administered questionnaire. The Cox proportional hazard model was used to estimate the hazard ratio (HR). During the follow-up period, a total of 116 deaths (46 males, 70 females) from gallbladder cancer were identified. After adjustments for age and gender, history of hepatic disease (HR: 2.28; 95% confidence intervals (95% CI): 1.24-4.21), frequency of stool, and tendency toward diarrhea (HR: 0.26; 95% CI: 0.08-0.83) were found to be significantly associated with the risk of death from gallbladder cancer. Compared with those who had a stool at least once a day, the HR was 2.06 (95% CI: 0.82-5.18) for those who had a stool less than once in 6 days (P for trend = 0.050). In this prospective study, constipation and a history of hepatic disease were found to elevate the risk of gallbladder cancer death, whereas a tendency toward diarrhea diminished it. PMID- 15298732 TI - Docetaxel enhances the cytotoxicity of cisplatin to gastric cancer cells by modification of intracellular platinum metabolism. AB - We have examined the combined anticancer effects of docetaxel (DOC) and cisplatin (CDDP) in vitro using the gastric cancer cell lines MKN-45, MKN-74, and TMK-1. Treatment of the cell lines with 30 microg/ml of DOC for 24 h followed by incubation with 3 or 10 microg/ml of CDDP for 24 h showed a clear synergistic effect. Sequence dependency of the agents was observed in these cell lines: DOC followed by CDDP (DC) showed a stronger antitumor effect than CDDP followed by DOC (CD) in all cell lines. To clarify the mechanism of action of the DC combination, total intracellular platinum (Pt) levels were evaluated after treatment with CDDP alone or combined with DC. For the MKN-45 and -74 cell lines, cells treated with DOC (10 microg/ml for 12 h) and then CDDP showed significantly increased intracellular Pt accumulation compared to cells treated with CDDP alone. We also investigated alterations in intracellular glutathione (GSH) concentration in response to DOC and CDDP. MKN-45 and -74 cells pretreated with DOC (10 microg/ml for 12 h) showed significantly increased intracellular GSH levels compared to cells administered CDDP only. To explain these findings, messenger RNA (mRNA) levels for multidrug resistance-associated protein-1 (MRP 1), the ATP-dependent pump for Pt-GSH complexes, were quantified in CDDP-treated MKN-45 cells with and without DOC pretreatment. While CDDP administration increased MRP-1 mRNA expression in MKN-45 cells, MRP-1 was not up-regulated after CDDP administration in DOC pretreated MKN-45 cells. Our results suggested that the enhanced CDDP toxicity due to DOC pretreatment may be related to the accumulation of intracellular Pt-GSH complexes, because DOC appears to suppress the MRP-1 up-regulation induced by CDDP exposure in gastric cancer cells. PMID- 15298733 TI - Antitumor activity of pyrvinium pamoate, 6-(dimethylamino)-2-[2-(2,5-dimethyl-1 phenyl-1H-pyrrol-3-yl)ethenyl]-1-methyl-quinolinium pamoate salt, showing preferential cytotoxicity during glucose starvation. AB - An anthelminthic, pyrvinium pamoate (PP), 6-(dimethylamino)-2-[2-(2,5-dimethyl-1 phenyl-1H-pyrrol-3-yl)ethenyl]-1-methyl-quinolinium pamoate salt, has been found to be extremely toxic to PANC-1 cells in glucose-free medium, but not to be toxic to the same cells cultured in ordinary medium, Dulbecco's modified Eagle's medium (DMEM). It showed the same preferential toxicity for various cancer cell lines during glucose starvation. When 0.1 microg/ml PP was added to the medium, spheroid growth of human colon cancer cell line WiDr was strongly inhibited to a diameter of 750 microm, and this finding is consistent with the concept of anti austerity. PP was also found to exert antitumor activity against human pancreatic cancer cell line PANC-1 in nude mice and SCID mice when it was administered subcutaneously or orally. Regarding the mechanism of PP action, inhibition of Akt phosphorylation, which has been found to be essential for the austerity mechanism, was observed in vitro and in vivo. These findings indicate that PP may be useful for anticancer therapy and that anti-austerity therapy could be a novel strategy for anticancer therapy. PMID- 15298734 TI - Phase I study of cisplatin, vinorelbine, and concurrent thoracic radiotherapy for unresectable stage III non-small cell lung cancer. AB - To determine the recommended phase II dose of vinorelbine in combination with cisplatin and thoracic radiotherapy (TRT) in patients with unresectable stage III non-small cell lung cancer (NSCLC), 18 patients received cisplatin (80 mg/m2) on day 1 and vinorelbine (20 mg/m2 in level 1, and 25 mg/m2 in level 2) on days 1 and 8 every 4 weeks for 4 cycles. TRT consisted of a single dose of 2 Gy once daily for 3 weeks followed by a rest of 4 days, and then the same TRT for 3 weeks to a total dose of 60 Gy. Fifteen (83%) patients received 60 Gy of TRT and 14 (78%) patients received 4 cycles of chemotherapy. Ten (77%) of 13 patients at level 1 and all 5 patients at level 2 developed grade 3-4 neutropenia. Four (31%) patients at level 1 and 3 (60%) patients at level 2 developed grade 3-4 infection. None developed > or = grade 3 esophagitis or lung toxicity. Dose limiting toxicity was noted in 33% of the patients in level 1 and in 60% of the patients in level 2. The overall response rate (95% confidence interval) was 83% (59-96%) with 15 partial responses. The median survival time was 30.4 months, and the 1-year, 2-year, and 3-year survival rates were 72%, 61%, and 50%, respectively. In conclusion, the recommended dose is the level 1 dose, and this regimen is feasible and promising in patients with stage III NSCLC. PMID- 15298735 TI - The Human Tissue Bill: good for consent but is it good for patients? PMID- 15298736 TI - The British Cardiac Society on the redefinition of myocardial infarction: a basis for discussion. PMID- 15298737 TI - The drugs don't work: pharmacogenomics--clinical biochemistry's future? PMID- 15298738 TI - British Cardiac Society Working Group on the definition of myocardial infarction. AB - The British Cardiac Society commissioned this report to help address inconsistencies in the terminology for acute coronary syndromes and wide variations in the threshold for the diagnosis of myocardial infarction (MI) depending on the assay performed, the precision, and the sensitivity. In addition, several publications have highlighted potential problems with the application of the European Society of Cardiology (ESC)/ American College of Cardiology (ACC) consensus document published in 2000. A revision process has been initiated under the guidance of the ESC, the ACC, and the American Heart Association (AHA). The purpose of this report is to help inform the next revision of the ESC/ACC/AHA guidelines for the diagnosis of MI. PMID- 15298739 TI - The measurement of vitamin D: analytical aspects. AB - In the past quarter of a century, our understanding of the metabolism and mechanism of action of vitamin D has been elucidated. During this period, many metabolites of vitamin D have been identified and a small proportion of these assayed in blood. The ability to assay these vitamin D metabolites has led to a better appreciation of the pathological role that altered vitamin D metabolism plays in the development of diseases of calcium homeostasis. However, for many physicians it is not clear which vitamin D metabolites should be quantitated and what the information gained tells us. Of the four major circulating vitamin D metabolites in blood, only two, namely 25-hydroxyvitamin D (25OHD) and 1,25 dihydroxyvitamin D [1,25(OH)(2)D], have warranted measurement. Of these, the need for assessing serum 1,25(OH)(2)D is actually quite limited and should therefore not be considered as part of the standard vitamin D testing regimen. 25OHD, on the other hand, provides us with the single best assessment of vitamin D nutritional status and should be the only vitamin D assay typically ordered for this reason. Which of the many methods that are available should a laboratory use for quantitating either of these vitamin D metabolites? Early methods required large volumes of blood, organic solvent extractions, and extensive purification of the vitamin D metabolites prior to assay. Today, these time-consuming and costly methods have given way to a range of radioimmunoassays and enzyme-linked immunosorbent assays that can accurately and conveniently provide important information concerning an individual's vitamin D status. This review will consider when vitamin D measurements should be undertaken and how best to perform such assays. PMID- 15298740 TI - The metabolic investigation of sudden infant death. AB - Inherited metabolic disorders account for a small but significant number of sudden unexplained deaths in neonates, infants and occasionally older children. In particular, inherited disorders of fatty acid oxidation may closely mimic sudden infant death syndrome. Post-mortem investigations offer the final opportunity to establish a diagnosis. Such diagnoses are of great importance to the families concerned and provide the opportunity for genetic counselling and antenatal diagnosis. Current advances in technology, particularly in the case of electrospray ionization tandem mass spectrometry, have revolutionized the investigation of metabolic disease at post-mortem, facilitating the identification of a wide range of metabolic diseases in tiny samples of blood, plasma and bile. Such analyses may provide vital clues to diagnosis, usually in the form of acylcarnitine profiles. Accurate diagnosis relies on the timely collection of appropriate samples and the subsequent selection of informative testing. In order to maximize the chances of a diagnosis, a collaborative approach between the various disciplines is vital. A brief description of the more frequently encountered inherited disorders, collection and processing of appropriate samples and available investigations that may lead to accurate diagnosis are clearly described. PMID- 15298741 TI - Thiopurine methyltransferase: should it be measured before commencing thiopurine drug therapy? AB - Thiopurines [azathioprine (AZA), 6-mercaptopurine (6-MP) and thioguanine (6-TG)] have a well-established role as immunosuppressive agents in a variety of chronic inflammatory conditions, haematological neoplasia and in transplant rejection. Despite good overall clinical response rates, particularly when used as steroid sparing agents, adverse effects are a limiting problem leading to withdrawal in up to a quarter of patients. Severe myelosuppression is the most serious toxicity occurring early or occasionally later during treatment. An understanding of the competing pathways involved in the metabolism of thiopurines has important implications for predicting some of the more severe toxicity seen with these drugs. Thiopurine methyl transferase (TPMT) is an enzyme catalysing the methylation of 6-MP, competing with xanthine oxidase (XO) and hypoxanthine guanine phosphoribosyl transferase (HGPRT) to determine the amount of 6-MP metabolised to cytotoxic thioguanine nucleotides. Allelic polymorphisms in the TPMT gene predict the activity of the enzyme such that 1 in 10 of the population are heterozygous and have approximately 50% of normal activity, whilst 1 in 300 are completely deficient. As a result, these individuals are at high risk of severe myelosuppression. Conversely, individuals with very high levels of TPMT activity are hyper-methylators in whom clinical response is less likely. Prior knowledge of TPMT status avoids exposure of individuals with zero TPMT to potentially fatal treatment with AZA or 6-MP and provides one of the best examples of predictive pharmacogenetics in therapeutics. This article reviews literature on the role of TPMT measurement prior to treatment with thiopurines and provides some guidance to the use of TPMT as a guide to tailoring thiopurine therapy. PMID- 15298742 TI - Reference intervals for thiopurine S-methyltransferase activity in red blood cells using 6-thioguanine as substrate and rapid non-extraction liquid chromatography. AB - BACKGROUND: Although widely used, thiopurine drugs have a narrow therapeutic index and treatment can result in life-threatening toxicity, the basis being pharmacogenetic variation in thiopurine metabolism by thiopurine S methyltransferase (TPMT). We recently developed a modified phenotyping assay to determine TPMT activity in red blood cells. Here we describe improvements to the method and establish reference intervals in a large prospective study. METHODS: A modified enzyme assay for TPMT activity is reported. It uses 6-thioguanine as substrate with heat treatment of the incubate to stop the reaction and precipitate protein prior to high-performance liquid chromatographic (HPLC) analysis. Measurement of the reaction product, 6-methylthioguanine (6-MTG), uses HPLC with fluorimetric detection. RESULTS: The assay shows excellent characteristics, with clear discrimination of patients who are deficient in TPMT activity (< 5 nmol 6-MTG per g Hb per h) from heterozygotes (5-24 nmol 6-MTG per g Hb per h) and patients with normal activity (>25 nmol 6-MTG per g Hb per h). CONCLUSION: A modified TPMT assay is described which is suited for routine analysis in a regional centre. The method overcomes the need for extraction and has speeded up the chromatographic determination of 6-MTG, enabling large numbers of samples to be analysed. A prospective study of 1000 individuals has established the distribution of TPMT activity using the assay. PMID- 15298743 TI - Characterization of the epitopes specific for the monoclonal antibody 9F5-3a and quantification of oxidized HDL in human plasma. AB - BACKGROUND: We previously isolated a monoclonal antibody, 9F5-3a, that is specific for HDL oxidized by CuSO(4). METHODS: We examined the characteristics of the 9F5-3a epitope by Western blot and measured the concentration of oxidized HDL in human plasma by enzyme-linked immunosorbent assay using this antibody. RESULTS: The monoclonal antibody specifically reacted with oxidized HDL in a mixture of HDL, LDL and modified lipoproteins. Oxidation of the HDL particles accelerated cross-linkage of apolipoproteins caused by lipid peroxidation, and the cross-linked apolipoprotein AI selectively reacted with the 9F5-3a antibody. Mean (standard deviation) plasma concentrations of oxidized HDL were 127 (50) microg/L in 23 healthy controls, 191 (65) microg/L in 30 patients with non insulin-dependent diabetes mellitus (P < 0.01 versus healthy controls) and 200 (87) microg/L in 25 patients with coronary artery disease (P < 0.01 versus healthy controls). The concentrations of oxidized HDL did not correlate with the concentrations of thiobarbituric acid-reactive substances. CONCLUSIONS: The results indicate that determination of oxidized HDL concentration may be useful for identifying patients with atherosclerotic disease. PMID- 15298744 TI - Paracetamol-associated interference in an HPLC-ECD assay for urinary free metadrenalines and catecholamines. AB - BACKGROUND: Some high-performance liquid chromatography techniques with electrochemical detection for urinary catecholamines and their metabolites can be subject to interference from drugs and their metabolites. Prime amongst these interferences are those due to paracetamol ingestion. METHODS: The prevalence of paracetamol contamination was determined by measuring the drug in all patient specimens submitted for catecholamine analysis over a 2-month period. These findings were then related to the proportion of unreportable results at each range of specimen paracetamol concentration. The apparent results from a small representative sample of the paracetamol-positive specimens are illustrated. RESULTS: Approximately one-third of urine specimens were found to contain paracetamol. Low-level contamination can produce apparent patterns of results which may easily be confused with those found in predominantly adrenaline-, or metadrenaline-secreting phaeochromocytomas. CONCLUSION: Despite the fact that significantly less than 5% of an oral dose of paracetamol is excreted as the free drug, its analysis provides a good surrogate marker for the likelihood of spurious results. It is useful to know prior to urinary free metadrenaline or catecholamine analysis which urine specimens are likely to contain potentially interfering paracetamol metabolites. PMID- 15298745 TI - Establishment of paediatric biochemical reference intervals. AB - BACKGROUND: The standard method for the determination of reference values in a population by testing a number of healthy volunteers is difficult within the paediatric age group; this study explores an alternative approach. METHODS: Biochemical blood test results collected by the Woman's and Children's Hospital over a 1 year period were used and the data selected to ensure, as far as possible, that the results were from patients who were "healthy". This was achieved using various selection criteria, such as the exclusion of patients with more than one test episode. Some of the data were skewed, making standard statistical approaches difficult. In such cases, transformations were used to ensure that the resulting information had a distribution that was approximately Gaussian. RESULTS: Data from different biochemical tests covering both genders and ages in the range 0-18 years were collected; this included more than 250 000 laboratory test results. After elimination of non-representative data and tests with insufficient results, there were approximately 23 500 results covering almost 3000 individuals and 16 different biochemical tests. There were no results from very young children (< 1 year) after the data selection process; hence the derivation of reference values from that age group was not possible. CONCLUSIONS: This approach permitted better delineation of reference intervals for common biochemical tests performed on paediatric patients than is currently readily available. In addition, some important benchmarks for transformation of medical data have been found. PMID- 15298746 TI - The response of serum apolipoprotein H to an oral fat load. AB - BACKGROUND: We tested the hypothesis that serum apolipoprotein H (apo H) concentration increases after an oral fat load. Such a study would give valuable insight into whether apo H was influenced by the postprandial state. METHODS: Ten male subjects aged 24-48 years were fed 62.5 g of total fat (saturates 12 g, monounsaturates 35.3 g, polyunsaturates 12.5 g). Venous blood was sampled hourly for 5 h post-oral fat load. RESULTS: No significant change in serum apo H concentration occurred following the oral fat load. However, serum apo H in the baseline samples correlated significantly with subject body mass index (r = 0.683, P < 0.05), body fat mass (r = 0.778, P < 0.01), lean body mass (r = 0.693, P < 0.05), serum triglyceride (r = 0.732, P < 0.02), serum insulin (r = 0.808, P < 0.01) and insulin resistance index (r = 0.794, P < 0.01). In stepwise multiple linear regression model, with serum apo A1, apo B, lipoprotein(a), total cholesterol, triglyceride, HDL-cholesterol, LDL-cholesterol, plasma glucose and insulin and apo H as the dependent variable, insulin remained in the model (r = 0.81, P < 0.01). Conversely, with body mass index, body fat mass, lean body mass and waist/hip ratio in the model and apo H as dependent variable, only body fat mass remained in the model (r = 0.78, P < 0.01). CONCLUSIONS: Serum apo H may be involved in insulin resistance and relates to various indices of adipose tissue, including body fat mass. However, serum apo H concentrations do not significantly change postprandially. PMID- 15298747 TI - Urinary 6beta-hydroxycortisol/17-hydroxycorticosteroids ratio as a measure of hepatic CYP3A4 capacity after enzyme induction. AB - BACKGROUND: The correlation between the urinary 6beta-hydroxycortisol/17 hydroxycorticosteroids (6beta-OHF/17-OHCS) ratio and the metabolic capacity of the most abundant form of hepatic cytochrome P450 (CYP3A4) after induction remains unclear. METHODS: Concentrations of 6beta-OHF and 17-OHCS in spot urine specimens obtained from 61 epileptic children receiving continuous carbamazepine therapy were measured by high-performance liquid chromatography. The relationship between the urinary 6beta-OHF/17-OHCS ratio and the serum carbamazepine concentration, corrected for dose and body weight, was examined. RESULTS: Serum carbamazepine was inversely associated with the urinary 6beta-OHF/17-OHCS ratio, and the hyperbolic relationship between the two parameters was statistically significant (P < 0.01). DISCUSSION: Carbamazepine is well known as a potent inducer and a substrate of hepatic CYP3A4. The present results suggest that measurement of the urinary 6betaOHF/17-OHCS ratio is helpful for assessing individuals' hepatic CYP3A4 capacity after enzyme induction. PMID- 15298748 TI - McArdle's disease diagnosed following statin-induced myositis. AB - We describe the case of a 69-year-old man with a history of muscular symptoms dating back to his childhood; McArdle's disease (glycogen-storage disease V) was diagnosed following an episode of myositis in which a statin and physical exertion appear to have been precipitating factors. This case demonstrates that the ischaemic lactate-ammonia test still has a place in screening patients with symptoms suggestive of McArdle's disease and emphasizes the importance of carrying out glycogen phosphorylase histochemistry on the skeletal muscle biopsy to confirm the diagnosis. In patients who develop a raised plasma creatine kinase level or muscular symptoms during lipid-lowering therapy, the clinician should be alert to the possibility of an underlying myopathy. PMID- 15298749 TI - Rhabdomyolysis in a patient with acute intermittent porphyria. AB - There are several conditions associated with rhabdomyolysis, such as direct muscle injury, viral infections, metabolic disorders and toxic effects from ingestion of drugs such as alcohol, opiates, cocaine or heroin. We report on a patient who was admitted to the accident and emergency unit with a clinical presentation of rhabdomyolysis and acute intermittent porphyria. PMID- 15298750 TI - Hypopituitarism presenting as a mixed hyperlipidaemia. AB - A case report is presented of a 37-year-old man who, largely by chance, was found to have a marked mixed hyperlipidaemia. As a teenager he had been treated for apparent idiopathic growth hormone deficiency, but had also never developed secondary sexual characteristics. Pituitary hormone measurement was now consistent with hypopituitarism and magnetic resonance imaging showed hypoplasia of his pituitary stalk as the likely cause. His hyperlipidaemia improved after appropriate hormone replacement. He thus appears to have had a mixed hyperlipidaemia secondary to hypopituitarism, which was secondary to a pituitary stalk abnormality, which in turn may have been associated to the trauma surrounding his normal breech delivery. PMID- 15298751 TI - Significance of serum troponin T in patients with kidney disease. PMID- 15298755 TI - Curcumin inhibits the growth of AGS human gastric carcinoma cells in vitro and shows synergism with 5-fluorouracil. AB - The inhibitory effect of curcumin and its synergism with 5-fluorouracil (5-FU) on the growth of the AGS human gastric carcinoma cell line was examined. Cell cycle analysis was used to elucidate the mechanisms for the inhibition by curcumin. Curcumin significantly inhibited the growth of AGS cells in a dose- and time dependent manner (P <.05). Curcumin caused a 34% decrease in AGS proliferation at 5 micromol/L, 51% at 10 micromol/L, and 92% at 25 micromol/L after 4 days of treatment. When curcumin (10 micromol/L) was removed after a 24-hour exposure, the growth pattern of curcumin-treated AGS cells was similar to that of control cells, suggesting reversibility of curcumin on the growth of AGS cells. Combining curcumin with 5-FU significantly increased growth inhibition of AGS cells compared with either curcumin or 5-FU alone (P <.05), suggesting synergistic actions of the two drugs. After 4 days of treatment with 10 micromol/L of curcumin, the G2/M phase fraction of cells was 60.5% compared with 22.0% of the control group, suggesting a G2/M block by curcumin treatment. Because the curcumin concentrations (5 micromol/L) used in our study were similar to steady state concentrations (1.77 +/- 1.87 micromol/L) in human serum of subjects receiving chronic administration of a commonly recommended dose (8 g/day), curcumin may be useful for the treatment of gastric carcinoma, especially in conjunction with 5-FU. PMID- 15298756 TI - Effect of chickpea aqueous extracts, organic extracts, and protein concentrates on cell proliferation. AB - Pulses should be part of a healthy diet, and it is also becoming clear that they have health-promoting effects. Nevertheless, most studies on the bioactive or health-promoting properties of pulses have been carried out using soybeans. We have studied cell growth-regulating properties, which may be responsible for anti cancer properties, in chickpea seeds. Chickpea seeds are a staple in the traditional diet of many Mediterranean, Asian, and South and Central American countries. In addition, chickpea seeds have industrial applications since they can be used for the preparation of protein concentrates and isolates. The cell lines Caco-2 (epithelial intestinal) and J774 (macrophages) have been exposed to chickpea seed extracts and protein preparations in order to screen the different chickpea fractions for effects on cell growth. Both cell growth-promoting and cell growth-inhibiting effects were found. Most interestingly, a fraction soluble in ethanol and acetone specifically and almost completely inhibited the growth of Caco-2 cells exhibiting a cancerous phenotype. It is concluded that chickpea seeds are a source of bioactive components and deserve further study for their possible anti-cancer effect. PMID- 15298757 TI - Sea tangle supplementation lowers blood glucose and supports antioxidant systems in streptozotocin-induced diabetic rats. AB - Diabetes mellitus is a chronic disease associated with serious complications that may be linked to increased lipid peroxidation. This study investigated the effects of dietary supplementation of sea tangle on lipid peroxidation and antioxidant systems in streptozotocin (STZ)-induced diabetes. Forty Sprague Dawley rats were divided into four groups (n = 10 each) fed AIN76-based diets with either sea tangle powder, water extract of sea tangle, or sodium alginate, or a control diet with no supplement. On day 21 after beginning the diets, rats received intramuscular injections of STZ (45 mg/kg of body weight) to induce diabetes. Experimental diet feeding was continued for 3 more weeks. Dietary supplementation with water extract of sea tangle resulted in lower plasma glucose compared with the control and sodium alginate groups. There was no significant difference in plasma and hepatic lipid peroxides among the groups. Sea tangle and sodium alginate did not affect activities of catalase and glutathione peroxidase; however, supplementation of water extract of sea tangle resulted in higher superoxide dismutase activity as compared with the control and sodium alginate groups. The plasma concentration of alpha-tocopherol increased in the sea tangle water extract group, but the hepatic concentration of alpha-tocopherol was not affected by dietary supplementation. Plasma retinol was not different among experimental groups. In conclusion, our results showed that water extract of sea tangle reduces plasma glucose and protects the antioxidant system in diabetic rats. These results suggest that water extract of sea tangle contains unknown physiologically active components, other than alginic acid, that may exert a protective effect against diabetes. PMID- 15298758 TI - The cholesterol-lowering effect of coconut flakes in humans with moderately raised serum cholesterol. AB - This study investigated the effect of coconut flakes on serum cholesterol levels of humans with moderately raised serum cholesterol in 21 subjects. The serum total cholesterol of subjects differed and ranged from 259 to 283 mg/dL. The study was conducted in a double-blind randomized crossover design on a 14-week period, consisting of four 2-week experimental periods, with each experimental period separated by a 2-week washout period. The test foods were as follows: corn flakes as the control food, oat bran flakes as the reference food, and corn flakes with 15% and 25% dietary fiber from coconut flakes (made from coconut flour production). Results showed a significant percent reduction in serum total and low-density lipoprotein (LDL) cholesterol (in mg/dL) for all test foods, except for corn flakes, as follows: oat bran flakes, 8.4 +/- 1.4 and 8.8 +/- 6.0, respectively; 15% coconut flakes, 6.9 +/- 1.1 and 11.0 +/- 4.0, respectively; and 25% coconut flakes, 10.8 +/- 1.3 and 9.2 +/- 5.4, respectively. Serum triglycerides were significantly reduced for all test foods: corn flakes, 14.5 +/ 6.3%; oat bran flakes, 22.7 +/- 2.9%; 15% coconut flakes, 19.3 +/- 5.7%; and 25% coconut flakes, 21.8 +/- 6.0%. Only 60% of the subjects were considered for serum triglycerides reduction (serum triglycerides >170 mg/dL). In conclusion, both 15% and 25% coconut flakes reduced serum total and LDL cholesterol and serum triglycerides of humans with moderately raised serum cholesterol levels. Coconut flour is a good source of both soluble and insoluble dietary fiber, and both types of fiber may have significant role in the reduction of the above lipid biomarker. To our knowledge, this is the first study conducted to show a relationship between dietary fiber from a coconut by-product and a lipid biomarker. Results from this study serves as a good basis in the development of coconut flakes/flour as a functional food, justifying the increased production of coconut and coconut by-products. PMID- 15298759 TI - Administration of a polysaccharide from Grifola frondosa stimulates immune function of normal mice. AB - We have reported that D-Fraction, a polysaccharide extracted from the edible maitake mushroom (Grifola frondosa), activates immunocompetent cells, thereby eliciting antitumor activity. To extend the application of D-Fraction as a nutritional supplement for healthy people as well as treatment for those with cancer, we investigated the effects of D-Fraction on the immune system in normal C3H/HeJ mice. Splenocytes from mice administered D-Fraction intraperitoneally for 17 consecutive days were cultured, and the culture supernatants were analyzed for nitric oxide (NO) and interleukin (IL)-12 production by antigen-presenting cells (APCs), including macrophages and dendritic cells, and also for the T helper (Th) 1 cytokine interferon (IFN)-gamma and the Th-2 cytokines IL-4 and IL-10. The level of IL-10 as well as those of NO and IFN-gamma were increased by D-Fraction as compared with the control, in which the serum immunoglobulin E level was increased. The results suggest that D-Fraction induced a Th-2 dominant response through the activation of macrophages, resulting in the enhancement of humoral immunity rather than cell-mediated immunity. Furthermore, an increase in the percentage ratio of CD69 and CD89 expression on major histocompatibility complex II(+) cells revealed activation of APCs 4 h after D-Fraction administration. These results indicate that D-Fraction enhances both the innate and adaptive arms of the immune response in normal mice. Therefore, its administration may enhance host defense against foreign pathogens and protect healthy individuals from infectious diseases. PMID- 15298760 TI - Effects of chlorella on activities of protein tyrosine phosphatases, matrix metalloproteinases, caspases, cytokine release, B and T cell proliferations, and phorbol ester receptor binding. AB - A Chlorella powder was screened using 52 in vitro assay systems for enzyme activity, receptor binding, cellular cytokine release, and B and T cell proliferation. The screening revealed a very potent inhibition of human protein tyrosine phosphatase (PTP) activity of CD45 and PTP1C with 50% inhibitory concentration (IC(50)) values of 0.678 and 1.56 microg/mL, respectively. It also showed a moderate inhibition of other PTPs, including PTP1B (IC(50) = 65.3 microg/mL) and T-cell-PTP (114 microg/mL). Other inhibitory activities and their IC(50) values included inhibition of the human matrix metalloproteinases (MMPs) MMP-1 (127 microg/mL), MMP-3 (185 microg/mL), MMP-7 (18.1 microg/mL), and MMP-9 (237 microg/mL) and the human peptidase caspases caspase 1 (300 microg/mL), caspase 3 (203 microg/mL), caspase 6 (301 microg/mL), caspase 7 (291 microg/mL), and caspase 8 (261 microg/mL), as well as release of the cytokines interleukin (IL)-1 (44.9 microg/mL), IL-2 (14.8 microg/mL), IL-4 (49.2 microg/mL), IL-6 (34.7 microg/mL), interferon-gamma (31.6 microg/mL), and tumor necrosis factor-alpha (11 microg/mL) from human peripheral blood mononuclear cells. Chlorella also inhibited B cell proliferation (16.6 microg/mL) in mouse splenocytes and T cell proliferation (54.2 microg/mL) in mouse thymocytes. The binding of a phorbol ester, phorbol 12,13-dibutyrate, to its receptors was also inhibited by Chlorella with an IC(50) of 152 microg/mL. These results reveal potential pharmacological activities that, if confirmed by in vivo studies, might be exploited for the prevention or treatment of several serious pathologies, including inflammatory disease and cancer. PMID- 15298761 TI - Supplementation of fenugreek leaves lower lipid profile in streptozotocin-induced diabetic rats. AB - The present study was undertaken to evaluate the lipid-lowering effect of fenugreek leaves in diabetes mellitus. Albino Wistar rats were randomly divided into six groups: normal untreated rats; streptozotocin (STZ)-induced diabetic rats; STZ-induced rats + fenugreek leaves (0.5 g/kg of body weight); STZ-induced rats + fenugreek leaves (1 g/kg of body weight); STZ-induced rats + glibenclamide (600 microg/kg of body weight); and STZ-induced rats + insulin (6 units/kg of body weight). Rats were made diabetic by STZ (40 mg/kg) injected intraperitoneally. Fenugreek leaves were supplemented in the diet daily to diabetic rats for 45 days, and food intake was recorded daily. Blood glucose, total cholesterol, triglycerides, and free fatty acids were determined in serum, liver, heart, and kidney. Our results show that blood glucose and serum and tissue lipids were elevated in STZ-induced diabetic rats. Supplementation of fenugreek leaves lowered the lipid profile in STZ-induced diabetic rats. PMID- 15298762 TI - Improvement of mouse memory by Myristica fragrans seeds. AB - Memory is one of the most complex functions of the brain and involves multiple neural pathways and neurotransmitter systems. The present study was undertaken to investigate the effect of Myristica fragrans (MF) seeds on learning and memory in mice. The n-hexane extract of MF was administered orally in three doses (5, 10, and 20 mg/kg p.o.) for 3 successive days to different groups of young and aged mice. The learning and memory parameters were assessed using elevated plus-maze and passive-avoidance apparatus. The effect of MF extract on scopolamine (0.4 mg/kg i.p.)- and diazepam (1 mg/kg i.p.)-induced impairment in learning and memory was also studied. MF extract at the lowest dose of 5 mg/kg p.o. administered for 3 successive days significantly improved learning and memory of young and aged mice. This extract also reversed scopolamine- and diazepam-induced impairment in learning and memory of young mice. MF extract enhanced learning and retention capacities of both young and aged mice. The exact mechanism of the memory-improving effect of MF extract was not explored in the present study. But, the observed memory-enhancing effect may be attributed to a variety of properties (individually or in combination) the plant is reported to possess, such as antioxidant, anti-inflammatory, or perhaps procholinergic activity. PMID- 15298763 TI - Saengshik, a formulated health food, decreases blood glucose and increases survival rate in streptozotocin-induced diabetic rats. AB - Saengshik is a Korean "non-cooked food" that is commercially produced and marketed. Ingredients in commercial Saengshik include grains, vegetables, fruits, mushrooms, sea plants, and various functional botanicals. This study investigated the effects of Saengshik on the survival rate of streptozotocin (STZ)-induced diabetic rats. Male Sprague-Dawley rats weighing around 190 g were randomly assigned to one of the three experimental groups: a healthy normal group (NC) and two groups with STZ-induced diabetes and fed either control diet (DC) or Saengshik diet (DS). Rats in all groups were supplied with a diet of equal energy. The animals were maintained on an experimental diet for 168 days in experiment I and for 42 days in experiment II. The body weight in the DS rats decreased less than in the DC rats in both experiments I and II. There was a trend for blood glucose level in the DS group to decrease during the experimental period in both experiments I and II. A survival rate of 50% was reached on day 49 in the DC group and on day 118 in the DS group. All rats in the DC group died by day 140, while 50% of the rats in the DS group were still alive on day 168, when experiment I was terminated. In experiment II, 50% of the DC group and 90% of the DS group survived at day 42. Saengshik did not have any influence on cholesterol levels, thiobarbituric acid-reactive substances, and activities of superoxide dismutase and glutathione peroxidase. These results suggest that blood glucose concentrations and the survival rate are positively affected by Saengshik feeding in diabetic rats. PMID- 15298764 TI - Kudzu root extract suppresses voluntary alcohol intake and alcohol withdrawal symptoms in P rats receiving free access to water and alcohol. AB - Alcohol-preferring (P) rats, given free choice to drink water or 15% alcohol, drank 7-10 g of alcohol/kg/day, giving blood alcohol values ranging from 16 to 24 mg/dL. Body weight and food and total fluid intake values in control and alcohol drinking P rats did not differ significantly, while water intake was inversely related to the amount of alcohol consumed. Alcohol withdrawal after 50 days of alcohol drinking caused withdrawal symptoms such as hypersensitivity, poor landing coordination, and tremors. A daily 0.5, 0.75, and 1.0 g/kg dose of kudzu root (KdR) did not affect body weight and food and water intake values in control (no alcohol) P rats. Subchronic feeding of relatively higher KdR doses (0.75 and 1.0 g/kg) caused a 25-30% reduction in weight gain. The 0.5 g/kg KdR dose caused a 50-60% reduction in alcohol consumption, abolished the development of alcohol withdrawal symptoms, but did not affect blood alcohol levels. The higher KdR doses did not further reduce alcohol consumption. Alcohol suppressed the weight reducing effects of KdR. The KdR extract used in this study contained 150 mg/g of puerarin, 13 mg/g of daidzin, 4 mg/g of daidzein, 3 mg/g of genistin, 0.2 mg/g of genistein, and 1 mg/g of glycetin. Blood and liver samples contained mostly puerarin and a trace amount of daidzein that may have been formed by the hydrolysis of daidzin by liver enzymes. An important observation was that brain samples obtained from KdR-fed or alcohol + KdR-fed rats did not contain any of the KdR isoflavones. Thus, KdR isoflavones suppressed alcohol drinking and withdrawal symptoms without entering the brain. PMID- 15298765 TI - Effects of purified puerarin on voluntary alcohol intake and alcohol withdrawal symptoms in P rats receiving free access to water and alcohol. AB - Alcohol preferring (P) rats, given "free choice" of water, exhibited daily intake of 60-75 g of water/kg of body weight. When given "free choice" of water and 15% ethanol, P rats consumed 7-13 g of alcohol/kg. Their water intake decreased proportionally to the alcohol intake, but total fluid intake did not differ significantly. Alcohol withdrawal after 50 days of alcohol drinking caused withdrawal symptoms such as hypersensitivity, poor coordination, and tremors. A daily 50 mg/kg dose of puerarin (PU) caused approximately 50% suppression in alcohol intake, but did not affect body weight and food and total fluid intake in P rats receiving "free choice" of water and 15% ethanol. Alcohol ingestion gradually returned to the control level despite consistent PU intake. However, alcohol intake following alcohol withdrawal was suppressed in PU-fed P rats. PU suppressed the severity of alcohol withdrawal symptoms. Thus, withdrawal symptoms do not occur in PU-fed rats even though their alcohol ingestion is comparable to that in control P rats. Brain, plasma, and liver samples were analyzed for the presence of kudzu root isoflavones, which are mostly PU (>90% of total isoflavones) and a trace amount of daidzin. Liver samples obtained from PU-fed P rats contained 20-30 microg/g of PU. An important observation was that plasma or brain samples obtained from PU-fed or alcohol + PU-fed rats did not contain PU. This study indicated that PU feeding transiently suppressed alcohol intake and abolished withdrawal symptoms at a time when alcohol intake had returned to the control level. The absence of PU in plasma and brain indicates the possibility that some nonspecific mechanism may be involved in the anti-alcoholism effects of PU in P rats. PMID- 15298766 TI - Anti-diabetic activity of Eugenia jambolana seed kernels on streptozotocin induced diabetic rats. AB - The present study evaluated the hypoglycemic activity of different parts of Eugenia jambolana seeds such as whole seed, kernel, and seed coat on streptozotocin-induced diabetic rats. Administration of the ethanolic extract of kernel at a concentration of 100 mg/kg of body weight significantly decreased the levels of blood glucose, blood urea, and cholesterol, increased glucose tolerance and levels of total proteins and liver glycogen, and decreased the activities of glutamate oxaloacetate transaminase and glutamate pyruvate transaminase in experimental diabetic rats. Whole seed showed a moderate hypoglycemic effect, and seed coat did not show any hypoglycemic effect. The hypoglycemic efficacy was compared with that of glibenclamide, a standard hypoglycemic drug. PMID- 15298767 TI - The effects of dietary bioflavonoid (rutin, quercetin, and naringin) supplementation on physiological changes in molar crestal alveolar bone-cemento enamel junction distance in young rats. AB - The objective of this study was to evaluate the influence of dietary bioflavonoid (rutin [R], quercetin [Q], and naringin [N]) supplementation on physiological molar crestal alveolar bone(CAB)-cemento-enamel junction (CEJ) distances in young male albino rats. The effects of diets supplemented with 0.57% R, Q, or N, at the expense of dextrose, were tested on 40 young rats, divided into four groups, for a period of 42 days. Rat skulls were defleshed, and CAB-CEJ distance was scored according to the modified method of Keyes and Gold (Oral Surg Oral Med Oral Pathol 1955;8:492). Data were analyzed using one-way analysis of variance, post hoc Tukey's test, and Spearman's (R(2)) correlation. P <.05 was used to reject the null hypothesis. The N group demonstrated the lowest CAB-CEJ distance, followed by the R and Q groups (P <.001-.05), except in the mandibular lingual region, where the Q group had a lower CAB-CEJ distance than the N and R groups (P <.05). The control group showed the largest CAB-CEJ distances. Dietary bioflavonoid supplementation was shown to significantly reduce molar CAB-CEJ distance (P <.001-.05) during alveolar development in male young rats. PMID- 15298768 TI - Differential effects of dietary fatty acids on the regulation of CYP2E1 and protein kinase C in human hepatoma HepG2 cells. AB - We investigated the effects of different fatty acids (FAs) or with different degrees of unsaturation on cytochrome P450 2E1 (CYP2E1) induction and protein kinase C (PKC) activity in human hepatoma HepG2 cells. As the degree of unsaturation increased, the cell survival rate decreased for FAs with 18 carbons, but linolenic acid (LNA) or docosahexaenoic acid (DHA) groups were similar even through they have different degrees of unsaturation. Treatment with palmitic acid (PA), oleic acid (OA), linoleic acid (LA), LNA, and DHA resulted in respective cellular FA concentrations of C16:0 (43.1%), C18:1 (18.5%), C18:2 (7.4%), LNA (2.85%), and C22:6 (3.13%), which was highest for the FA that was used as the treatment, indicating that their incorporation within the cell is directly proportional to treatment. After 2 hours of cultivation, the lipid peroxide (LPO) in the DHA group increased 600% compared with control, and was much higher than in the groups treated with the other FAs, with LNA > LA > OA > PA. CYP2E1 induction increased with greater effect as the degree of unsaturation of OA, LA, and DHA increased. PA did not affect PKC activity, but DHA treatment increased PKC activity the most. The effects of LNA and LA were similar, but less than that of DHA, and that of OA was lower still, indicating that activity of PKC is proportional to the degree of unsaturation, and not the configuration of the FA. Increased plasma membrane concentrations of n-3 FA, such as DHA, might exert regulatory effects on PKC by increasing membrane fluidity, causing changes in CYP2E1, elevating levels of LPO, or producing oxidative stress. PMID- 15298769 TI - Protective role of Phaseolus vulgaris on changes in the fatty acid composition in experimental diabetes. AB - The present investigation was carried out to evaluate the effect of Phaseolus vulgaris, an indigenous plant used in Unani and Ayurvedic medicine in India, on blood glucose, plasma insulin, cholesterol, triglycerides, free fatty acids, phospholipids, and fatty acid composition of total lipids in liver, kidney, and brain of normal and streptozotocin (STZ) diabetic rats. The results show that there was a significant increase in tissue cholesterol, triglycerides, free fatty acids, and phospholipids in STZ diabetic rats. The analysis of fatty acids showed that there was a significant increase in the concentrations of palmitic acid (16:1), stearic acid (18:0), and oleic acid (18:1) in liver, kidney, and brain, whereas the concentrations of linolenic acid (18:3) and arachidonic acid (20:4) were significantly decreased. Oral administration of the aqueous extract of P. vulgaris pods (200 mg/kg of body weight) for 45 days to diabetic rats decreased the concentrations of lipids and fatty acids, viz., palmitic, stearic, and oleic acids, whereas linolenic and arachidonic acids were elevated. Similarly, the administration of P. vulgaris pod extract (PPEt) to normal animals resulted in a significant hypolipidemic effect. These results suggest that PPEt exhibits hypoglycemic and hypolipidemic effects in STZ diabetic rats. It also prevents the fatty acid changes produced during diabetes. The effect of PPEt at 200 mg/kg of body weight was better than that of glibenclamide. PMID- 15298770 TI - Investigating the antimicrobial activity of natural honey and its effects on the pathogenic bacterial infections of surgical wounds and conjunctiva. AB - Antimicrobial activities of 10-100% (wt/vol) concentrations of new honey, stored honey, heated honey, ultraviolet-exposed honey, and heated stored honey were tested against common human pathogens, including Escherichia coli, Entrobacter cloacae, Pseudomonas aeruginosa, Shigella dysenteriae, Klebsiella sp., Haemophilus influenzae, Proteus sp., Staphylococcus aureus, Streptococcus hemolyticus group B, and Candida albicans. Antimicrobial activity of honey was tested in acidic, neutral, or alkaline media. These were compared with similar concentrations of glucose in nutrient broth. Surgical wounds were made on the dorsum of mice and infected with S. aureus or Klebsiella sp. The wounds were treated with local application of honey four times a day or appropriate antibiotics and compared with control values. Bacterial conjunctivitis due to E. coli, Proteus sp., S. aureus, Klebsiella sp., and P. aeruginosa was induced in rats. Conjunctival application of honey four times a day or appropriate antibiotics was used for treatment and compared with control values. Growth of all the isolates was completely inhibited by 30-100% honey concentrations. The most sensitive microbes were E. coli, P. aeruginosa, and H. influenzae. Glucose showed less antimicrobial activity than honey, and many microbes showed positive culture even in 100% glucose. Heating to 80 degrees C for 1 hour decreased antimicrobial activity of both new and stored honey. Storage of honey for 5 years decreased its antimicrobial activity, while ultraviolet light exposure increased its activity against some of the microorganisms. Antimicrobial activity of honey was stronger in acidic media than in neutral or alkaline media. Single doses of honey used to prepare the 60% concentration in nutrient broth were bacteriocidal for P. aeruginosa and bacteriostatic for S. aureus and Klebsiella sp. during certain periods. Local application of raw honey on infected wounds reduced redness, swelling, time for complete resolution of lesion, and time for eradication of bacterial infection due to S. aureus or Klebsiella sp. Its potency was comparable to that of local antibiotics. Honey application into infective conjunctivitis reduced redness, swelling, pus discharge, and time for eradication of bacterial infections due to all the isolates tested. PMID- 15298772 TI - Methionine-induced hyperhomocysteinemia promotes superoxide anion generation and NFkappaB activation in peritoneal macrophages of C57BL/6 mice. AB - It is well documented that hyperhomocysteinemia (HHcy) is an independent risk factor for atherosclerosis. This study was designed to investigate whether some of the atherosclerotic effects ascribed to HHcy are mediated by oxidative stress and nuclear factor kappa B (NFkappaB) activation in peritoneal macrophages of C57BL/6 mice fed a high (2%) methionine/low (1 mg/kg) folate diet for 12 weeks. Plasma homocysteine concentrations in mice fed methionine averaged 49 mol/L after 12 weeks of feeding, five times higher than that of controls. HHcy induced by methionine feeding significantly elevated oxidative stress, as measured by superoxide anion radical level (P <.05) in peritoneal macrophages. Furthermore, NFkappaB binding activities of peritoneal macrophages were higher in the methionine group than in the control group. These results suggest that HHcy induced by methionine may intensify disturbances in peroxidation and inflammatory mediator activation in peritoneal macrophages, and is a possible mechanism of its atherogenic effects. PMID- 15298771 TI - Nucleic acid-based diagnosis of bacterial vaginosis and improved management using probiotic lactobacilli. AB - Bacterial vaginosis (BV) is a common condition in women that represents an imbalance of the vaginal microflora, lactobacilli depletion, and excess growth of mainly anaerobic Gram-negative pathogens. Diagnosis is made using a series of tests or a Gram stain of a vaginal smear. Treatment with antibiotics is quite effective, but recurrences are common. A study of 55 vaginal samples from 11 postmenopausal women showed the presence of BV by the Gram stain-based Nugent scoring system, and polymerase chain reaction-denaturing gradient gel electrophoresis showed that Bacteroides or Prevotella species were the most common isolates recovered (24 of 25), with Escherichia coli, Staphylococcus aureus, and Streptococcus agalactiae also found in some samples. In one case, only Gardnerella vaginalis was found. These findings illustrate that BV remains common even among otherwise healthy women, but it is not caused solely by either Gardnerella or Mobiluncus. Use of a FemExam system (Cooper Surgical, Shelton, CT), based upon elevated pH and trimethylamine levels, to screen vaginal smears from 59 healthy women showed poor correlation with the Gram stain method. A randomized, placebo-controlled trial of these subjects showed that the lactobacilli-dominant microbiota was restored in subjects with BV but not in controls, following 2 months of daily oral intake of Lactobacillus rhamnosus GR-1 and Lactobacillus fermentum RC-14. These studies show that nucleic acid-based methods are effective at identifying bacteria responsible for BV. If such methods could be used to develop a commercially available, self-use kit, women would be much better placed to take control of their own health, for example, using medicinal food or dietary supplement products such as the clinically proven probiotic strains L. rhamnosus GR-1 and L. fermentum RC-14. PMID- 15298773 TI - Effect of Alstonia scholaris in enhancing the anticancer activity of berberine in the Ehrlich ascites carcinoma-bearing mice. AB - The chemomodulatory activity of Alstonia scholaris extract (ASE) was studied in combination with berberine hydrochloride (BCL), a topoisomerase inhibitor, in Ehrlich ascites carcinoma-bearing mice. The tumor-bearing animals were injected with various doses of ASE, and 8 mg/kg of BCL (one-fifth of the 50% lethal dose) was combined with different doses of ASE (60-240 mg/kg). The combination of 180 mg/kg of ASE with 8 mg/kg of BCL showed the greatest antitumor effect; the number of tumor-free survivors was more, and the median survival time and the average survival time increased up to 47 and 40.5 days, respectively, when compared with either treatment alone. Similarly, when 180 mg/kg of ASE was combined with different doses of BCL (2-12 mg/kg), a dose-dependent increase in the anticancer activity was observed up to 8 mg/kg of BCL. However, a further increase in the BCL dose to 10 and 12 mg/kg resulted in toxic side effects. The best effect was observed when 180 mg/kg of ASE was combined with 6 or 8 mg/kg of BCL, where an increase in the antineoplastic activity was reported. The efficacy of the combination of 180 mg/kg of ASE was also tested with 6 mg/kg body weight of BCL in various stages of tumorigenesis, and it was effective when given in the early stages, although the efficiency decreased with an increase in the tumor developmental stages. PMID- 15298774 TI - Evaluation of the protective efficacy of Asteracantha longifolia on acetaminophen induced liver damage in rats. AB - The effect of oral administration of methanolic extract of Asteracantha longifolia (AL) seeds on acetaminophen (APAP)-induced acute liver damage in rats was investigated. The activities of marker enzymes (aspartate transaminase, alanine transaminase, alkaline phosphatase, lactate dehydrogenase, and gamma glutamyl transferase) and bilirubin level in serum and the levels of cholesterol, triglycerides, and free fatty acids in both serum and liver were found to be increased when rats were challenged with APAP. This was also associated with a significant reduction of serum and tissue phospholipids. Pretreatment with AL extract prior to the administration of APAP prevented these alterations as evidenced by liver histopathology. Results indicated that the extract could offer protection against APAP-induced liver damage, suggesting its hepatoprotective activity. PMID- 15298775 TI - Antitumor activity of beta-D-glucan from Libyan dates. AB - Beta-glucan with antitumor activities was isolated from Libyan dates, and the structure of the purified glucan was characterized using methods such as methylation, periodate oxidation, and acetolysis. Glucans were found to exhibit potent antitumor activity; this activity could be correlated to their (1-->3) beta-D-glucan linkages. Such antitumor glucans have also been obtained from a number of other sources, such as yeast, fungi, bacteria, and plants. This is the first study to report antitumor activity for date glucan. PMID- 15298776 TI - Study on wound healing activity of Punica granatum peel. AB - The methanolic extract of dried pomegranate (Punica granatum) peels showed the presence of a high content of phenolic compounds (44.0%) along with other constituents. This extract was formulated as a 10% (wt/wt) water-soluble gel and was studied for its wound healing property against an excision wound on the skin of Wistar rats. The activity was compared with that of a commercial topical antibacterial applicant. The wound healing activity was assessed by measuring the percent contraction in skin and estimation of collagen content in terms of hydroxyproline content. Healed skin was also subjected to histopathological studies to examine the microscopic changes. The animals treated with 2.5% gel showed moderate healing (55.8% and 40.8% healing compared with negative and positive controls, respectively), whereas the group treated with 5.0% gel showed good healing (59.5% and 44.5% healing compared with negative and positive controls, respectively). The amount of hydroxyproline increased by twofold in the group treated with 5.0% gel. Histopathological studies also supported the wound healing on application of the gels. The group of rats that received 5.0% gel showed complete healing after 10 days, whereas in rats treated with 2.5% gel, healing was observed on day 12, in contrast to the positive control animals receiving the blank gel, which took 16-18 days for complete healing. The results of this study may be extended to different types of wounds so that the formulation could be exploited to develop it as a topical dermatological formulation. High-performance liquid chromatography analysis of the extract showed the presence of gallic acid and catechin as major components. PMID- 15298777 TI - D-003 and warfarin interaction on the bleeding time and venous thrombosis experimentally induced in rats. AB - D-003 is a mixture of higher aliphatic primary acids isolated and purified from sugarcane wax, the main component of which is octacosanoic acid. D-003 exhibits a cholesterol-lowering effect as well as antiplatelet and antithrombotic effects in experimental models. Warfarin is a coumarin derivative with anticoagulant activity that acts as a vitamin K antagonist. Since in clinical practice warfarin and D-003 could be administered together, the objective of this study was to evaluate the effects of the simultaneous administration of both drugs on the bleeding time and the venous thrombosis experimentally induced in rats. The combined therapy of minimally effective doses of D-003 and warfarin produced an antithrombotic effect significantly higher than those produced by each monotherapy. Likewise, the prolongation of bleeding time induced by warfarin was increased by the simultaneous administration with D-003, showing a synergistic effect between both drugs. PMID- 15298778 TI - The 3-D prefrontal cortex: Hemispheric asymmetries in prefrontal activity and their relation to memory retrieval processes. AB - Neuroimaging results have raised interest in characterizing hemispheric asymmetries in prefrontal activity during different types of memory retrieval tasks. In this issue, Dobbins et al. and Mitchell et al. report results suggesting that the two hemispheres of the prefrontal cortex may indeed make different contributions to memory retrieval. Here, I discuss these findings within the context of studies characterizing more general processing differences between the cerebral hemispheres and studies characterizing prefrontal organization along the dorsal-ventral and anterior-posterior dimensions. PMID- 15298779 TI - fMRI evidence for separable and lateralized prefrontal memory monitoring processes. AB - Source memory research suggests that attempting to remember specific contextual aspects surrounding prior stimulus encounters results in greater left prefrontal cortex (PFC) activity than simple item-based old/new recognition judgments. Here, we tested a complementary hypothesis that predicts increases in the right PFC with tasks requiring close monitoring of item familiarity. More specifically, we compared a judgment of frequency (JOF) task to an item memory task, in which the former required estimating the number of previous picture encounters and the latter required discriminating old from new exemplars of previously seen items. In comparison to standard old/new recognition, both source memory and the JOF task examined here require more precise mnemonic judgments. However, in contrast to source memory, cognitive models suggest the JOF task relies heavily upon item familiarity, not specific contextual recollections. Event-related fMRI demonstrated greater recruitment of right, not left, dorsolateral and frontopolar PFC regions during the JOF compared to item memory task. These data suggest a role for right PFC in the close monitoring of the familiarity of objects, which becomes critical when contextual recollection is ineffective in satisfying a memory demand. PMID- 15298780 TI - Prefrontal cortex activity associated with source monitoring in a working memory task. AB - Using functional magnetic resonance imaging (fMRI), we investigated prefrontal cortex (PFC) activity during remembering specific source information (format, location judgments) versus remembering that could be based on undifferentiated information, such as familiarity (old/new recognition [ON], recency judgments). A working memory (WM) paradigm with an immediate test yielded greater activation in the lateral PFC for format and location source memory (SM) tasks than ON recognition; this SM-related activity was left lateralized. The same regions of PFC were recruited in Experiment 2 when information was tested immediately and after a filled delay. Substituting recency for location judgments (Experiment 3) resulted in an overall shift in task context that produced greater right PFC activity associated with ON and recency tasks compared to the format task, in addition to left SM-related activity. These data extend to WM previous findings from long-term memory (LTM) indicating that the left and right PFC may be differentially involved in memory attributions depending on the specificity of information evaluated. The findings also provide evidence for the continuity of evaluative processes recruited in WM and LTM. PMID- 15298781 TI - Reaching out to see: arm position can attenuate human visual loss. AB - Electrophysiological recordings in monkeys have now revealed several brain regions that contain bimodal visuotactile neurons capable of responding to either tactile or visual stimuli placed on or near the hands, arms, and face. These cells have now been found in frontal, parietal, and subcortical areas of the monkey brain, suggesting a cortical network of neurons that preferentially represent near peripersonal space. The degree to which the visual responses of such cells rely on input from the primary visual cortex and the extent to which they may contribute to visual perception is not completely understood. Nonetheless, recent neuropsychological studies suggest that a similar representation of near space may be bimodally coded in humans as well. Given the accumulating evidence for specialized processing of visual stimuli placed near the hands and arms, we hypothesized that arm position may be capable of modulating human visual ability. Here we report the case of WM, who lost his ability to see in his left visual hemifield after sustaining damage to his right primary visual cortex. Interestingly, the placement of WM's left arm into his "blind" field resulted in significantly better detection of left visual field stimuli compared to when his hand was placed in his lap at midline. Moreover, we found this attenuation to be confined to stimuli presented within reaching distance (unless a tool that extended WM's reach was held while he performed the test). These findings are highly consistent with the characteristics of the bimodal visuo-tactile neurons that have been described in monkeys. Thus, it seems that arm position can modulate human visual ability, even after damage to the primary visual cortex. This study provides an exciting bridge between monkey neurophysiology and human visual capacity while also offering a novel approach for improving visual defects acquired via cortical injury. PMID- 15298782 TI - Neural correlates for the suppression of habitual behavior: a functional MRI study. AB - It has been suggested that inhibitory executive control of behavior is directed by the frontal lobes. We used functional magnetic resonance imaging to explore the brain regions that are involved in the inhibition of habitual manual responses. Fifteen right-handed subjects performed the rock-scissors-paper game against computer-simulated pictures of hands during the scanning procedure. The subjects were required to win, lose, or draw against the presented picture in a separate block. We considered that the situation in which subjects intentionally lost the game required the suppression of habitual behavior, because it is natural behavior for people to attempt to win the game. Compared with the WIN and DRAW conditions, the left premotor and sensorimotor areas were activated for both hand sessions with a positive correlation with error rates. Importantly, the LOSE condition in the case of the right hand yielded brain activation exclusively in the anterior part of the left inferior frontal gyrus, the activity which showed a negative correlation with error rates. Overall brain activations were predominant in the left hemisphere, irrespective of the hand used for the response. The results suggest that the anterior part of the left inferior frontal gyrus plays a critical role in the inhibition of habitual manual behavior, and that the left hemisphere is dominant for the selection of well-learned manual behavior. PMID- 15298783 TI - Behavioral and neuroimaging evidence for a contribution of color and texture information to scene classification in a patient with visual form agnosia. AB - A common notion is that object perception is a necessary precursor to scene perception. Behavioral evidence suggests, however, that scene perception can operate independently of object perception. Further, neuroimaging has revealed a specialized human cortical area for viewing scenes that is anatomically distinct from areas activated by viewing objects. Here we show that an individual with visual form agnosia, D.F., who has a profound deficit in object recognition but spared color and visual texture perception, could still classify scenes and that she was fastest when the scenes were presented in the appropriate color. When scenes were presented as black-and-white images, she made a large number of errors in classification. Functional magnetic resonance imaging revealed selective activation in the parahippocampal place area (PPA) when D.F. viewed scenes. Unlike control observers, D.F. demonstrated higher activation in the PPA for scenes presented in the appropriate color than for black-and-white versions. The results demonstrate that an individual with profound form vision deficits can still use visual texture and color to classify scenes-and that this intact ability is reflected in differential activation of the PPA with colored versions of scenes. PMID- 15298784 TI - Neural correlates of auditory repetition priming: reduced fMRI activation in the auditory cortex. AB - Repetition priming refers to enhanced or biased performance with repeatedly presented stimuli. Modality-specific perceptual repetition priming has been demonstrated behaviorally for both visually and auditorily presented stimuli. In functional neuroimaging studies, repetition of visual stimuli has resulted in reduced activation in the visual cortex, as well as in multimodal frontal and temporal regions. The reductions in sensory cortices are thought to reflect plasticity in modality-specific neocortex. Unexpectedly, repetition of auditory stimuli has resulted in reduced activation in multimodal and visual regions, but not in the auditory temporal lobe cortex. This finding puts the coupling of perceptual priming and modality-specific cortical plasticity into question. Here, functional magnetic resonance imaging was used with environmental sounds to reexamine whether auditory priming is associated with reduced activation in the auditory cortex. Participants heard environmental sounds (e.g., animals, machines, musical instruments, etc.) in blocks, alternating between initial and repeated presentations, and decided whether or not each sound was produced by an animal. Repeated versus initial presentations of sounds resulted in repetition priming (faster responses) and reduced activation in the right superior temporal gyrus, bilateral superior temporal sulci, and right inferior prefrontal cortex. The magnitude of behavioral priming correlated positively with reduced activation in these regions. This indicates that priming for environmental sounds is associated with modification of neural activation in modality-specific auditory cortex, as well as in multimodal areas. PMID- 15298785 TI - Modulation of motor excitability during speech perception: the role of Broca's area. AB - Studies in both human and nonhuman primates indicate that motor and premotor cortical regions participate in auditory and visual perception of actions. Previous studies, using transcranial magnetic stimulation (TMS), showed that perceiving visual and auditory speech increased the excitability of the orofacial motor system during speech perception. Such studies, however, cannot tell us which brain regions mediate this effect. In this study, we used the technique of combining positron emission tomography with TMS to identify the brain regions that modulate the excitability of the motor system during speech perception. Our results show that during auditory speech perception, there is increased excitability of motor system underlying speech production and that this increase is significantly correlated with activity in the posterior part of the left inferior frontal gyrus (Broca's area). We propose that this area "primes" the motor system in response to heard speech even when no speech output is required and, as such, operates at the interface of perception and action. PMID- 15298786 TI - How would you feel versus how do you think she would feel? A neuroimaging study of perspective-taking with social emotions. AB - Perspective-taking is a complex cognitive process involved in social cognition. This positron emission tomography (PET) study investigated by means of a factorial design the interaction between the emotional and the perspective factors. Participants were asked to adopt either their own (first person) perspective or the (third person) perspective of their mothers in response to situations involving social emotions or to neutral situations. The main effect of third-person versus first-person perspective resulted in hemodynamic increase in the medial part of the superior frontal gyrus, the left superior temporal sulcus, the left temporal pole, the posterior cingulate gyrus, and the right inferior parietal lobe. A cluster in the postcentral gyrus was detected in the reverse comparison. The amygdala was selectively activated when subjects were processing social emotions, both related to self and other. Interaction effects were identified in the left temporal pole and in the right postcentral gyrus. These results support our prediction that the frontopolar, the somatosensory cortex, and the right inferior parietal lobe are crucial in the process of self/other distinction. In addition, this study provides important building blocks in our understanding of social emotion processing and human empathy. PMID- 15298787 TI - Multisensory interaction in saccadic reaction time: a time-window-of-integration model. AB - Saccadic reaction time to visual targets tends to be faster when stimuli from another modality (in particular, audition and touch) are presented in close temporal or spatial proximity even when subjects are instructed to ignore the accessory input (focused attention task). Multisensory interaction effects measured in neural structures involved in saccade generation (in particular, the superior colliculus) have demonstrated a similar spatio-temporal dependence. Neural network models of multisensory spatial integration have been shown to generate convergence of the visual, auditory, and tactile reference frames and the sensorimotor coordinate transformations necessary for coordinated head and eye movements. However, because these models do not capture the temporal coincidences critical for multisensory integration to occur, they cannot easily predict multisensory effects observed in behavioral data such as saccadic reaction times. This article proposes a quantitative stochastic framework, the time-window-of-integration model, to account for the temporal rules of multisensory integration. Saccadic responses collected from a visual-tactile focused attention task are shown to be consistent with the time-window-of integration model predictions. PMID- 15298788 TI - Musical training enhances automatic encoding of melodic contour and interval structure. AB - In music, melodic information is thought to be encoded in two forms, a contour code (up/down pattern of pitch changes) and an interval code (pitch distances between successive notes). A recent study recording the mismatch negativity (MMN) evoked by pitch contour and interval deviations in simple melodies demonstrated that people with no formal music education process both contour and interval information in the auditory cortex automatically. However, it is still unclear whether musical experience enhances both strategies of melodic encoding. We designed stimuli to examine contour and interval information separately. In the contour condition there were eight different standard melodies (presented on 80% of trials), each consisting of five notes all ascending in pitch, and the corresponding deviant melodies (20%) were altered to descending on their final note. The interval condition used one five-note standard melody transposed to eight keys from trial to trial, and on deviant trials the last note was raised by one whole tone without changing the pitch contour. There was also a control condition, in which a standard tone (990.7 Hz) and a deviant tone (1111.0 Hz) were presented. The magnetic counterpart of the MMN (MMNm) from musicians and nonmusicians was obtained as the difference between the dipole moment in response to the standard and deviant trials recorded by magnetoencephalography. Significantly larger MMNm was present in musicians in both contour and interval conditions than in nonmusicians, whereas MMNm in the control condition was similar for both groups. The interval MMNm was larger than the contour MMNm in musicians. No hemispheric difference was found in either group. The results suggest that musical training enhances the ability to automatically register abstract changes in the relative pitch structure of melodies. PMID- 15298789 TI - Optimizing a linear algorithm for real-time robotic control using chronic cortical ensemble recordings in monkeys. AB - Previous work in our laboratory has demonstrated that a simple linear model can be used to translate cortical neuronal activity into real-time motor control commands that allow a robot arm to mimic the intended hand movements of trained primates. Here, we describe the results of a comprehensive analysis of the contribution of single cortical neurons to this linear model. Key to the operation of this model was the observation that a large percentage of cortical neurons located in both frontal and parietal cortical areas are tuned for hand position. In most neurons, hand position tuning was time-dependent, varying continuously during a 1-sec period before hand movement onset. The relevance of this physiological finding was demonstrated by showing that maximum contribution of individual neurons to the linear model was only achieved when optimal parameters for the impulse response functions describing time-varying neuronal position tuning were selected. Optimal parameters included impulse response functions with 1.0- to 1.4-sec time length and 50- to 100-msec bins. Although reliable generalization and long-term predictions (60-90 min) could be achieved after 10-min training sessions, we noticed that the model performance degraded over long periods. Part of this degradation was accounted by the observation that neuronal position tuning varied significantly throughout the duration (60-90 min) of a recording session. Altogether, these results indicate that the experimental paradigm described here may be useful not only to investigate aspects of neural population coding, but it may also provide a test bed for the development of clinically useful cortical prosthetic devices aimed at restoring motor functions in severely paralyzed patients. PMID- 15298790 TI - A role for cortical crosstalk in the binding problem: stimulus-driven correlations that link color, form, and motion. AB - The putative independence of cortical mechanisms for color, form, and motion raises the binding problem-how is neural activity coordinated to create unified and correctly segmented percepts? Binding could be guided by stimulus-driven correlations between mechanisms, but the nature of these correlations is largely unexplored and no one has (intentionally) studied effects on binding if this joint information is compromised. Here, we develop a theoretical framework which: (1) describes crosstalk-generated correlations between cortical mechanisms for color, achromatic form, and motion, which arise from retinogeniculate encoding; (2) shows how these correlations can facilitate synchronization, segmentation, and binding; (3) provides a basis for understanding perceptual oddities and binding failures that occur for equiluminant and stabilized images. These ideas can be tested by measuring both perceptual events and neural activity while achromatic border contrast or stabilized image velocity is manipulated. PMID- 15298791 TI - Dichoptic visual masking reveals that early binocular neurons exhibit weak interocular suppression: implications for binocular vision and visual awareness. AB - Visual masking effects are illusions in which a target is rendered invisible by a mask, which can either overlap or not overlap the target spatially and/or temporally. These illusions provide a powerful tool to study visibility and consciousness, object grouping, brightness perception, and much more. As such, the physiological mechanisms underlying the perception of masking are critically important to our understanding of visibility. Several models that require cortical circuits have been proposed previously to explain the mysterious spatial and timing effects associated with visual masking. Here we describe single-unit physiological experiments from the awake monkey that show that visual masking occurs in at least two separate and independent circuits, one that is binocular and one that is monocular (possibly even subcortical), without feedback from higher-level visual brain areas. These and other results together fail to support models of masking that require circuits found only in the cortex, but support our proposed model that suggests that simple ubiquitous lateral inhibition may itself be the fundamental mechanism that explains visual masking across multiple levels in the brain. We also show that area V1 neurons are dichoptic in terms of excitation, but monoptic in terms of inhibition. That is, responses within area V1 binocular neurons reveal that excitation to monocular targets is inhibited strongly only by masks presented to the same eye, and not by masks presented to the opposite eye. These results lead us to redefine the model for the first stage of binocular processing in the visual system, and may be crucial to interpreting the effects of other similar binocular and dichoptic stimulation paradigms, such as the binocular rivalry family of illusions. PMID- 15298792 TI - Timing of target discrimination in human frontal eye fields. AB - Frontal eye field (FEF) neurons discharge in response to behaviorally relevant stimuli that are potential targets for saccades. Distinct visual and motor processes have been dissociated in the FEF of macaque monkeys, but little is known about the visual processing capacity of FEF in humans. We used double-pulse transcranial magnetic stimulation [(d)TMS] to investigate the timing of target discrimination during visual conjunction search. We applied dual TMS pulses separated by 40 msec over the right FEF and vertex. These were applied in five timing conditions to sample separate time windows within the first 200 msec of visual processing. (d)TMS impaired search performance, reflected in reduced d' scores. This effect was limited to a time window between 40 and 80 msec after search array onset. These parameters correspond with single-cell activity in FEF that predicts monkeys' behavioral reports on hit, miss, false alarm, and correct rejection trials. Our findings demonstrate a crucial early role for human FEF in visual target discrimination that is independent of saccade programming. PMID- 15298793 TI - The influence of semantic and syntactic context constraints on lexical selection and integration in spoken-word comprehension as revealed by ERPs. AB - An event-related brain potential experiment was carried out to investigate the influence of semantic and syntactic context constraints on lexical selection and integration in spoken-word comprehension. Subjects were presented with constraining spoken sentences that contained a critical word that was either (a) congruent, (b) semantically and syntactically incongruent, but beginning with the same initial phonemes as the congruent critical word, or (c) semantically and syntactically incongruent, beginning with phonemes that differed from the congruent critical word. Relative to the congruent condition, an N200 effect reflecting difficulty in the lexical selection process was obtained in the semantically and syntactically incongruent condition where word onset differed from that of the congruent critical word. Both incongruent conditions elicited a large N400 followed by a left anterior negativity (LAN) time-locked to the moment of word category violation and a P600 effect. These results would best fit within a cascaded model of spoken-word processing, proclaiming an optimal use of contextual information during spoken-word identification by allowing for semantic and syntactic processing to take place in parallel after bottom-up activation of a set of candidates, and lexical integration to proceed with a limited number of candidates that still match the acoustic input. PMID- 15298794 TI - Cue validity and object-based attention. AB - In a previous study, Egly, Driver, and Rafal (1994) observed both space- and object-based components of visual selective attention. However, the mechanisms underlying these two components and the relationship between them are not well understood. In the present research, with a similar paradigm, these issues were addressed by manipulating cue validity. Behavioral results indicated the presence of both space- and object-based components under high cue validity, similar to the results of Egly et al.'s study. In addition, under low cue validity, the space-based component was absent, whereas the object-based component was maintained. Further event-related potential results demonstrated an object-based effect at a sensory level over the posterior areas of brain, and a space-based effect over the anterior region. The present data suggest that the space- and object-based components reflect mainly voluntary and reflexive mechanisms, respectively. PMID- 15298795 TI - Haloperidol impairs learning and error-related negativity in humans. AB - Humans are able to monitor their actions for behavioral conflicts and performance errors. Growing evidence suggests that the error-related negativity (ERN) of the event-related cortical brain potential (ERP) may index the functioning of this response monitoring system and that the ERN may depend on dopaminergic mechanisms. We examined the role of dopamine in ERN and behavioral indices of learning by administering either 3 mg of the dopamine antagonist (DA) haloperidol (n = 17); 25 mg of diphenhydramine (n = 16), which has a similar CNS profile but without DA properties; or placebo (n = 18) in a randomized, double-blind manner to healthy volunteers. Three hours after drug administration, participants performed a go/no-go Continuous Performance Task, the Eriksen Flanker Task, and a learning-dependent Time Estimation Task. Haloperidol significantly attenuated ERN amplitudes recorded during the flanker task, impaired learning of time intervals, and tended to cause more errors of commission, compared to placebo, which did not significantly differ from diphenhydramine. Drugs had no significant effects on the stimulus-locked P1 and N2 ERPs or on behavioral response latencies, but tended to affect post-error reaction time (RT) latencies in opposite ways (haloperidol decreased and diphenhydramine increased RTs). These findings support the hypothesis that the DA system is involved in learning and the generation of the ERN. PMID- 15298796 TI - [Application of fluid resuscitation and baso-active agents in the treatment of severe sepsis and septic shock]. PMID- 15298798 TI - [Effects of reconstructive human acidic fibroblast growth factor and wild type acidic fibroblast growth factor on skin cell proliferation]. AB - OBJECTIVE: To investigate the effects of reconstructive human acidic fibroblast growth factor (aFGF) and wild type aFGF on skin cell proliferation in rat. METHODS: Neonatal rat skin (area of 2 mmx2 mm) was cultured in Dulbecco's modification of Eagle's medium containing reconstructive human aFGF and wild type aFGF, respectively. The concentrations of aFGF were 1 microg/L, 10 microg/L, and 100 microg/L. After being cultured for 4 days, the area of skin was measured. RESULTS: After treatment with two different growth factors in three different concentrations (1 microg/L, 10 microg/L and 100 microg/L) for 4 days, the areas of skin in three reconstructive human aFGF groups were 1.4, 1.5 and 1.3 fold of that of control, respectively and the areas of three wild type aFGF groups were 1.5, 3.2 and 1.6 fold of that of control, respectively, while the area of skin in the control group was (2.96+/-1.12) mm(2). In comparison with those of other groups, the skin area of 10 microg/L wild type aFGF group was significantly increased (P<0.05). CONCLUSION: Reconstructive human aFGF confers less impact on cutaneous cell growth. The capability of wild type aFGF to induce cutaneous cell proliferation is much greater than that of reconstructive human aFGF. PMID- 15298797 TI - [Analysis of etiological factors of pseudo-epitheliomatous granuloma after cutaneous wound healing: a report of 11 cases of patients with this lesion]. AB - OBJECTIVE: To explore the pathogenetic mechanism, clinical and pathological characteristics, and prevention and treatment of pseudo-epitheliomatous granuloma after skin wound healing. METHODS: The clinical information and the treatment results of pseudo-epitheliomatous granuloma occurring in 11 patients (age 1-67 years) were reviewed and analyzed, their tissue specimens were used for microbial examination and histological observation. RESULTS: Some bacteria such as Staphylococcus aureus, Pseudomonas aeruginosa, Streptococcus hemolyticus B, and Streptococcus faecalis could be found in the culture of pseudo-epitheliomatous granuloma. The maJority of these bacteria was tolerant to celbenin but sensitive to vancomycin. The lesions were excised, and the wounds were covered with skin grafting or skin flap, supplemented by local of vancomycin to prevent recurrence of the lesion. Histological examination revealed in pseudo-epitheliomatous granuloma with long epithelial peduncle encapsulating granulation tissue like honeycomb, in which there were many capillaries, macrophages, lymphocytes and mast cells, with only a small amount of extracellular matrix. CONCLUSION: The main pathogenesis of pseudo-epitheliomatous granuloma formation might be improper treatment of wound in earlier period and infection of drug resistant bacteria. Surgical debridement followed by skin coverage and local inJection of vancomycin could be effective and curative. PMID- 15298799 TI - [Effects of acidic fibroblast growth factor on intestinal mucosal damage due to ischemia/reperfusion]. AB - OBJECTIVE: To investigate the effect of exogenous acidic fibroblast growth factor (aFGF) on ischemia/reperfusion (I/R) injury of the intestine in rat, and to explore the mechanism of its protective effect. METHODS: The model of rat intestinal I/R injury was replicated by clamping the superior mesenteric artery (SMA) for 45 minutes followed by reperfusion. Seventy-eight Wistar rats were divided randomly into sham-operation group, intestinal I/R control group and aFGF treated group. In sham-operation group, SMA were separated but not occluded. Animals were sacrificed, samples of blood and tissue from the intestine were obtained 45 minutes after ischemia in sham-operation group and 0.5, 1, 2, 6, 12, and 24 hours after reperfusion in the other groups. Plasma D-lactate level was measured and rat survival rate was noted. Histological changes in intestinal tissue were observed under light microscope. RESULTS: The results showed that rat survival rate in aFGF treatment group was higher than that of intestinal I/R control group (P<0.05), and plasma D-lactate levels were markedly lower in aFGF group than that of intestinal I/R control group (P<0.05). Improvement in pathological changes was observed at 1, 2, 6, 12, and 24 hours after the reperfusion in aFGF treatment group compared with intestinal I/R control group. CONCLUSION: These data suggest that aFGF has a protective effect on intestinal I/R damage, and it may be related to the mitogenic effects and some hormone-like activities of aFGF. PMID- 15298801 TI - [Analysis of diagnosis and misdiagnosis of pulmonary thromboembolism in emergency]. AB - OBJECTIVE: To investigate the differential diagnosis of pulmonary thromboembolism in emergency in order to increase the correct diagnosis rate of pulmonary thromboembolism. METHODS: The data of patients who were diagnosed pulmonary thromboembolism in Emergency Intensive Care Center of Anzhen Hospital from January 2000 to the end of August 2003 were analyzed. Patients were divided into two groups. One group of patients with pulmonary thromboembolism were diagnosed pneumonia, heart disease and cerebrovascular disease. The other group of patients were not pulmonary thromboembolism and were misdiagnosed. RESULTS: Five patients with pulmonary thromboembolism were diagnosed pneumonia, 7 and 3 patients were diagnosed heart disease and cerebrovascular disease, respectively. In the other group, 4 patients with lung or mediastinal tumors, 2 with pneumonia and 1 with bronchiectasis were diagnosed pulmonary thromboembolism. CONCLUSION: It is essential to correctly diagnose pulmonary thromboembolism, especially emergency physicians should master diagnosis method of pulmonary thromboembolism and reduce the rate missed of diagnosis. PMID- 15298802 TI - [Roles of hydroxyethyl starch solution in resuscitation for shock induced by ischemia/reperfusion injury of the intestine]. AB - OBJECTIVE: To compare the effects of high and low doses of 6% hydroxyethyl starch solution (HES) on resuscitation for shock induced by ischemia/reperfusion injury of the intestine in rabbits. METHODS: Thirty-two anesthetized rabbits were randomized into four groups of eight animals each. The animals in the control group received no fluid resuscitation, animals in group 2 received lactated Ringer's solution (LRS, 20 ml x kg(-1) x h(-1)), those in group 3 received LRS+HES (LRS 18 ml x kg(-1) x h(1)+HES 2 ml x kg(-1) x h(-1), low dosage of HES), and those in group 4 received HES only in high dosage of HES (20 ml x kg(-1) x h( 1)). All these rabbits underwent intestinal ischemia/reperfusion injury developed by occluding superior mesenteric artery (SMA) with a non-crushing vascular clamp for 60 minutes and then the clamp was loosened for 240 minutes. The fluid resuscitation began at the same time of reperfusion. Hemodynamic parameters including mean artery pressure, heart rate, aortic velocity (as cardiac output), and SMA blood flow were measured. Tissue oxygenation was assessed indirectly by measuring the tonometric parameters of the gut, including difference between partial pressure of carbon dioxide in intestinal intramucosal and partial pressure of carbon dioxide in arterial blood (Pt-aCO(2) gap), intestinal intramucosal pH (pHi), arterial blood lactate acid concentration and oxygen delivery. Mortality of the rabbits was counted at the end of the experiment. RESULTS: Hemodynamic parameters as measured in low and high doses of HES groups were significantly higher in values than LRS group (LRS) and control (all P<0.05). Low dose of HES was better in restoring hemodynamic parameters than high dose of HES (all P<0.05). Low dose of HES could greatly decrease lactate and Pt aCO(2) gap, significantly improve pHi compared with other three groups (all P<0.05), but high dose of HES did not do so, and oral and nasal bleeding even death of some animals were seen. Low dose and high dose of HES significantly improved oxygen delivery while LRS did not (all P<0.05). CONCLUSION: Low dose of HES together with LRS is more effective than only high dose of HES or LRS in the resuscitation for shock induced by ischemia/reperfusion injury of the intestine in rabbits, resulting in better hemodynamic parameters and tissue oxygenation. PMID- 15298805 TI - [Study on morphological change in intestinal mucosa from injury to repair after hemorrhagic shock]. AB - OBJECTIVE: To study on morphological changes in mucosa of the small and large intestine mucosa after resuscitation of hemorrhagic shock. METHODS: The morphological changes in intestinal mucosa were observed under light and electron microscope, including the histology of intestinal mucosa, determination of height of villi and evaluation of mucosa damage index in the different phases after traumatic-hemorrhagic shock. RESULTS: Mucosa epithelial injury was obvious in small intestine were even at 0 hour, becoming more serious in 1 hour up to 3 hours. The tissue repair began after 3 hours, though the injury was still serious. Most of the inJured mucosa began to repair after 6 hours, and completed in 24 hours. The condition of the large intestine was similar to that of the small intestine, but the injury was less severe. The mucosal thickness and the height of villi were diminished after 1 hour of shock, but there was no obvious change in the colon. CONCLUSION: In the early phase after hemorrhagic shock, intestinal mucosal barrier are subJected to damage, but it could repair and recover in a short time. Compare with small intestine, large intestine have stronger potentiality against injury. PMID- 15298803 TI - [Mechanism of better result of limited resuscitation in a model of uncontrolled hemorrhagic shock]. AB - OBJECTIVE: To investigate the mechanism of better result of limited resuscitation in a model of uncontrolled hemorrhagic shock. METHODS: Uncontrolled hemorrhagic shock was produced in 54 rats by a standardized massive splenic injury with transection of the middle branch of splenic artery (MSIA). The rats were randomly assigned to six groups (n=6) by maintenance of the level of mean arterial pressure (MAP): sham-operated group (SS), 40 (RS40), 50 (RS50), 60 (RS60), 80 (RS80) and 100 mm Hg (RS100, 1 mm Hg=0.133 kPa). When the MAP reached 40 mm Hg, resuscitation was begun. Ringer's solution was continued as needed to maintain the following desired endpoints for 45 minutes (T45 point): MAP of 40, 50, 60, 80 and 100 mm Hg. After the bleeding was controlled, resuscitation was continued with Ringer's solution and whole blood (2:1) to raise the MAP to 100 mm Hg for 120 minutes (T165 point), followed by a 240 minutes observation period (T405 point). All animals were observed for 240 minutes or till death. The blood samples were withdrawn from artery for hematocrit (Hct), blood lactate (BL), base excess(BE) at T0, T45, T165, T405 points. At the end of the experiment, a small amount of hepatic tissue was collected for measuring tissue blood perfusion, total antioxidative capacity (T-AOC), Na(+)-K(+)-ATPase, and malondialdehyde (MDA). RESULTS: At T45, T405 points, Hct in SS, RS50 and RS60 were significantly higher than in RS80 and RS100(P<0.05). At T405 point, BL and BE levels in RS80 and RS100 were significantly higher than that of the other groups (P<0.05). The contents of MDA in SS, RS40, and RS50 were significantly lower than in RS80 and RS100(P<0.05). T-AOC level, Na(+)-K(+)-ATPase were significantly lower in RS80 and RS100 than that in the other groups. Blood perfusion was significantly lower in RS80 and RS100 than that in SS, RS40, and RS50. CONCLUSION: In the setting of uncontrolled hemorrhagic shock, limited resuscitation could balance well the needs of organ perfusion and decrease lactate level. It might also exert a protective effect against ischemia/reperfusion injury to liver tissue. PMID- 15298807 TI - [Level of tissue plasminogen activator protein in pulmonary artery in acute pulmonary thromboembolism in rabbit]. AB - OBJECTIVE: To study the changes in tissue plasminogen activator(t-PA) protein in pulmonary artery and its clinical significance after acute pulmonary thromboembolism (PTE). METHODS: Thirty rabbits were randomly divided into four groups after replicating a model of acute PTE in rabbit by thrombi occlusion method. Specimens were obtained from both normal and morbid pulmonary artery 3, 8 and 24 hours after APE, and protein contents of t-PA were determined using immunohistochemical method. RESULTS: A few endothelial cells and smooth muscle cells of the normal pulmonary artery were positive for t-PA. After 3 hours of PTE, there was no significant changes in t-PA positive stain among embolismic, non-embolismic and normal pulmonary artery. After 8 and 24 hours of PTE, strong positive staining was found in the residual endothelial cells and a part of smooth muscle cells (all P<0.01). CONCLUSION: There is significantly strong positive staining for t-PA in the pulmonary artery wall after pulmonary embolism, implying that the local fibrinolysis activity was enhanced, and it might be helpful for lysis of the embolus. PMID- 15298809 TI - [Analysis of the status of diagnosis of acute pulmonary embolism and a study of further strategy to improve the diagnostic level]. AB - OBJECTIVE: To analyze the present status of diagnosis of pulmonary embolism (PE) by analyzing the clinical data of patients admitted during last 10 years to our hospital, in order to look for a significant diagnostic strategy to improve the diagnostic level. METHODS: The data of patients diagnosed to have PE in last 10 years were analyzed. In the last 5 months, the patients suspected to have PE were diagnosed by a comprehensive approach including clinical manifestations, lung scan and/or spirals computer tomography (SCT) in our emergency department. RESULTS: The diagnostic rate, the final diagnostic rate within 3 days and the diagnostic rate by lung scan or/and SCT were all higher in the last 3 months than ever. The proportions of angiograms and the overall 3-month PE risk were 2.8% and 0. CONCLUSION: The comprehensive approach which includes clinical manifestations, lung scan and/or SCT can obviously improve the diagnostic level of PE. PMID- 15298810 TI - [Therapeutic application of molecular adsorbent recirculation system in the treatment of multiple organ dysfunction syndrome patients]. AB - OBJECTIVE: To assess the effectiveness of molecular adsorbent recirculation system (MARS) to remove nitric oxide(NO) and cytokines in multiple organ dysfunction syndrome(MODS) in patients with severe liver failure. METHODS: Single MARS treatment were performed for 198 times with duration ranging from 6 to 24 hours on 61 MODS patients (42M/19F). The efficacy was evaluated by sequential organ failure assessment, biochemical parameters and the levels of pro inflammatory cytokines. RESULTS: The MARS therapy resulted in a significant removal of NO and certain cytokines such as tumor necrosis factor-alpha(TNF alpha), interleukin-2(IL-2), IL-6, IL-8, and lipopolysaccharide-binding protein(LBP), together with marked reduction of other non-water soluble albumin bound toxins and water soluble toxins. These were associated with an improvement of the patients' clinical conditions, including deranged hemodynamics, respiratory function, cardiovascular and renal functions, hepatic encephalopathy, thus resulting in a marked decrease of sequential organ failure assessment(SOFA) score and improved outcome. Twenty-five patients were able to be discharged from the hospital, and successful liver transplantation could be performed in 6 patients. The overall survival rate of 61 patients was 41.0%. CONCLUSION: MARS could be used for the treatment of MODS patients associated with elevated levels of NO and cytokines with satisfactory results. PMID- 15298813 TI - [The dynamic changes in cytokines and oxygen metabolism in critically ill patients]. PMID- 15298814 TI - [Dynamic changes in coagulation parameters and platelet counts in perioperative patients with in-situ liver transplantation]. PMID- 15298822 TI - Psychosocial aspects of genetic screening of pregnant women and newborns: a systematic review. AB - OBJECTIVES: To address five broad questions concerned with knowledge, anxiety, factors associated with participation/non-participation in screening programmes and the long-term sequelae of false-positive, true-positive in newborns and true negative results. DATA SOURCES: Five electronic databases, two journals and attempts were made to locate unpublished work. REVIEW METHODS: This review started from a substantial literature base that provided the basis for (a) scoping the literature, (b) informing search strategy terms and (c) identifying preliminary article inclusion and exclusion criteria. The main eligibility criteria were: any screening programme aimed at pregnant women or newborn babies that included a 'genetic' target condition, this included chromosomal anomalies; any study that reported psychosocial data collected directly from parents. The data extraction form developed for this study elicited data from the selected studies. The data were entered into a database, which provided a summary of the included papers. RESULTS: A total of 288 candidate publications were identified, 106 of which were eligible: 78 were concerned with antenatal screening and 28 with newborn screening. It was found that levels of knowledge adequate for decision-making were not being achieved despite information leaflets and videos having some effect. Studies that have succeeded in increasing knowledge have not observed a corresponding increase in anxiety, although some anxiety might be an appropriate response and may aid coping and decision-making. Anxiety is clearly raised in women receiving positive screening results, especially young women. However, evidence is lacking of a beneficial (i.e. reassuring) effect of receiving a screen-negative result. Anxiety in screen-positive women falls on receipt of subsequent reassuring results, but some residual anxiety may remain. A minority (perhaps up to 30%) of women receiving a screen-positive result in pregnancy expressed regret about their screening decision. Uptake of neonatal screening has been treated as a 'given' and not as a research topic. CONCLUSIONS: The results of this review have many implications for the work of the National Screening Committee. The most pressing of these, in order of priority, relate to: the inadequacy of current procedures for achieving informed consent; the cost of providing a satisfactory service; the unmet needs of 'false-positives', and the unmet needs of women's partners, particularly in carrier screening. It is suggested that research is conducted on the above four topics in order to fill gaps in the evidence base that relate to screening technologies which have been available for many years. In addition, future screening programmes will create a new list of research questions, based on the same main agenda but applied to new areas, for example, new conditions such as haemoglobinopathies and fragile X syndrome; new client groups such as young women and minority ethnic groups; and new testing modalities such as ultrasound. PMID- 15298825 TI - Effective and ineffective interventions for infant colic. PMID- 15298823 TI - The Social Support and Family Health Study: a randomised controlled trial and economic evaluation of two alternative forms of postnatal support for mothers living in disadvantaged inner-city areas. AB - OBJECTIVES: To determine whether increased postnatal support could influence maternal and child health outcomes. DESIGN: This was a randomised controlled trial comparing maternal and child health outcomes for women offered either of the support interventions with those for control women receiving standard services only. Outcome data were collected through questionnaires distributed 12 and 18 months postrandomisation. Process data were also collected. There was also an integral economic evaluation. SETTING AND PARTICIPANTS: Women living in deprived enumeration districts in selected London boroughs were eligible for the trial if they gave birth between 1 January and 30 September 1999. RESULTS: The 731 participants were found to be well matched in terms of socio-economic characteristics and health and support variables (14% of the participants were non-English speaking). Response rates at the two follow-up points were 90% and 82%. At both points there were no differences that could not be attributed to chance on the primary outcomes of maternal depression, child injury or maternal smoking. At the first follow-up, there was reduced use of general practitioners by support health visitor (SHV) children, but increased use of NHS health visitors and social workers by mothers. At the second follow-up, both community group support (CGS) and SHV mothers had less use of midwifery services (fewer were pregnant), and SHV mothers were less worried about their child's health and development. Uptake of the CGS intervention was low: 19%, compared with 94% for the SHV intervention. Satisfaction with the intervention among women in the SHV group was high. Based on the assumptions and conditions of the costing methods, the economic evaluation found no net economic cost or benefit of choosing either of the two interventions. CONCLUSIONS: There was no evidence of impact on the primary outcomes of either intervention. The SHV intervention was popular with women, and was associated with improvement in some of the secondary outcomes. This suggests that greater emphasis on the social support role of health visitors could improve some measures of family well-being. Possible areas for future research include a systematic review of social support and its effect on health; developing and testing other postnatal models of support that match more closely the age of the baby and the changing patterns of mothers' needs; evaluating other strategies for mobilising 'non-professional' support; developing and testing more culturally specific support interventions; developing more culturally appropriate standardised measures of health outcomes; providing longer term follow-up of social support interventions; and exploring the role of social support on the delay in subsequent pregnancy. PMID- 15298826 TI - Steroid injections effective for knee osteoarthritis. PMID- 15298827 TI - Peak expiratory flow rate does not predict asthma exacerbations. PMID- 15298828 TI - Open hernia repair better than laparoscopic. PMID- 15298829 TI - Petroleum jelly does not reduce recurrent pediatric epistaxis. PMID- 15298830 TI - Severe rash after dermatitis. PMID- 15298831 TI - Allowing spirituality into the healing process. PMID- 15298832 TI - Principles to make a spiritual assessment work in your practice. PMID- 15298833 TI - Is religious devotion relevant to the doctor-patient relationship? PMID- 15298834 TI - Isotonic saline or hypertonic saline: which is best for sinusitis? PMID- 15298835 TI - Aspirin for persons at higher risk for colorectal neoplasia. PMID- 15298836 TI - Diagnosis of melanoma. PMID- 15298837 TI - How should we manage newly diagnosed atrial fibrillation? PMID- 15298838 TI - Intention-to-treat analysis: protecting the integrity of randomization. PMID- 15298839 TI - Mad Cow disease: dealing sensibly with a new concern. PMID- 15298840 TI - Clinical inquiries. What is the best approach for managing recurrent bacterial vaginosis? PMID- 15298841 TI - Clinical inquiries. Should we screen women for hypothyroidism? PMID- 15298842 TI - Clinical inquiries. When should patients with stroke receive thrombolytics? PMID- 15298843 TI - Clinical inquiries. Is methylphenidate useful for treating adolescents with ADHD? PMID- 15298844 TI - Clinical inquiries. Does yoga speed healing for patients with low back pain? PMID- 15298845 TI - Clinical inquiries. Do inhaled beta-agonists control cough in URIs or acute bronchitis? PMID- 15298849 TI - Development and evaluation of a quantitative video-fluorescence imaging system and fluorescent tracer for measuring transfer of pesticide residues from surfaces to hands with repeated contacts. AB - A video imaging system and the associated quantification methods have been developed for measurement of the transfer of a fluorescent tracer from surfaces to hands. The highly fluorescent compound riboflavin (vitamin B2), which is also water soluble and non-toxic, was chosen as the tracer compound to simulate the transfer from surfaces to hands of pesticide residues deposited on carpeted and laminate surfaces of a residence. The system was designed around the unique properties of riboflavin. Excitation energy was centered near 440 nm (in the blue region of the visible spectrum); emitted energy was measured at 600 nm (in the red/orange region), well beyond the significant fluorescence peak maximum of natural skin. A video camera system with an image intensifier was interfaced to an image processing analysis software system. Quantification utilized chemometric techniques to account for the non-linearity of pixel detectivity and non-linear excitation strength. Method quantification and detection limits were approximately 0.1 and 0.02 micro g/cm(2), respectively. The relative error was approximately 100% at the quantification limit, but <20% at higher levels. Transfer of riboflavin to hands, resulting in dermal loadings in the range 0.1 2.0 micro g/cm(2), were measured with this system from surfaces whose loadings approximated the pesticide levels that occur in homes after broadcast application. PMID- 15298850 TI - The efficacy of local exhaust ventilation for controlling dust exposures during concrete surface grinding. AB - This study assessed the effectiveness of a commercially available local exhaust ventilation (LEV) system for controlling respirable dust and crystalline silica exposures during concrete grinding activities. Surface grinding was conducted at six commercial building construction sites in Seattle, WA, by cement masons. Time integrated filter samples and direct reading respirable dust concentrations were collected using a cyclone in line with a direct reading respirable dust monitor. Personal exposure levels were determined with and without LEV, one sample directly after the other. A total of 28 paired samples were collected in which three different dust collection shroud configurations were tested. Data obtained with a direct reading respirable dust monitor were adjusted to remove non-work task-associated dust exposures and was subsequently used to calculate the exposure reduction achieved. The application of LEV resulted in a reduction in the overall geometric mean respirable dust exposure from 4.5 to 0.14 mg/m(3), a mean exposure reduction of 92%. Despite the effective control of dust generated during surface grinding, 22 and 26% of the samples collected while LEV was being used were greater than the 8 h time-weighted average permissible exposure limit (Occupational Safety and Health Administration) and threshold limit value (American Congress of Governmental Industrial Hygienists) for respirable crystalline silica, respectively. PMID- 15298851 TI - RhoA and Rac1 are both required for efficient wound closure of airway epithelial cells. AB - Repair of the airway epithelium after injury is critical for restoring normal lung. The reepithelialization process involves spreading and migration followed later by cell proliferation. Rho-GTPases are key components of the wound healing process in many different types of tissues, but the specific roles for RhoA and Rac1 vary and have not been identified in lung epithelial cells. We investigated whether RhoA and Rac1 regulate wound closure of bronchial epithelial cells. RhoA and Rac1 proteins were efficiently expressed in a cell line of human bronchial epithelial cells (16HBE) by adenovirus-based gene transfer. We found that both constitutively active RhoA and dominant negative RhoA inhibited wound healing, suggesting that both activation and inhibition of RhoA interfere with normal wound healing. Overexpression of wild-type Rac1 induced upregulation of RhoA, disrupted intercellular junctions, and inhibited wound closure. Dominant negative Rac1 also inhibited wound closure. Inhibition of the downstream effector of RhoA, Rho-kinase, with Y-27632 suppressed actin stress fibers and focal adhesion formation, increased Rac1 activity, and stimulated wound closure. The activity of both RhoA and Rac1 are influenced by the polymerization state of microtubules, and cell migration involves coordinated action of actin and microtubules. Microtubule depolymerization upon nocodazole treatment led to an increase in focal adhesions and decreased wound closure. We conclude that coordination of both RhoA and Rac1 activity contributes to bronchial epithelial wound repair mechanisms in vitro, that inhibition of Rho-kinase accelerates wound closure, and that efficient repair involves intact microtubules. PMID- 15298852 TI - Therapeutic effect of in vivo transfection of transcription factor decoy to NF kappaB on septic lung in mice. AB - Nuclear factor-kappaB (NF-kappaB) plays a key role in regulating expression of several genes involved in the pathophysiology of endotoxic shock. We investigated whether in vivo introduction of synthetic double-stranded DNA with high affinity for the NF-kappaB binding site could block expression of genes mediating pulmonary vascular permeation and thereby provide effective therapy for septic lung failure. Endotoxic shock was induced by an intravenous injection of 10 mg/kg Escherichia coli endotoxin in mice. We introduced NF-kappaB decoy oligodeoxynucleotide (ODN) in vivo 1 h after endotoxic shock by using a gene transfer kit. At 10 h, blood samples were collected for measurement of histamine and for blood-gas analysis. Gene and protein expression levels of target molecules were determined by means of Northern and Western blot analyses, respectively. The transpulmonary flux of (125)I-labeled albumin was used as an index of lung vascular permeability. Administration of endotoxin caused marked increases in plasma histamine and gene and protein expressions of histidine decarboxylase, histamine H(1) receptors, and inducible nitric oxide synthase in lung tissues. Elevated lung vascular permeability was also found. Blood-gas analysis showed concurrent decreases in arterial Po(2), Pco(2), and pH. All of these events induced by endotoxin were significantly inhibited by transfection of NF-kappaB decoy ODN but not by its mutated (scrambled) form (used as a control). Our results indicate for the first time the potential usefulness of NF-kappaB decoy ODN for gene therapy of endotoxic shock. PMID- 15298853 TI - Airway injury in lung disease pathophysiology: selective depletion of airway stem and progenitor cell pools potentiates lung inflammation and alveolar dysfunction. AB - Identification of early events that contribute to the establishment of chronic lung disease has been complicated by the variable involvement of the airway and alveolar compartments in the complex physiology of end-stage disease. In particular, the impact of airway injury on alveolar integrity and function has not been addressed and would be facilitated by development of animal models of lung disease that specifically target a single cell type within the airway epithelium. We have previously demonstrated that ganciclovir treatment of CCtk transgenic mice, which express the herpes simplex thymidine kinase gene under regulation of the mouse Clara cell secretory protein (CCSP) promoter, results in elimination of the airway progenitor and stem cell pools and a consequent failure of airway regeneration that is associated with rapid morbidity and mortality. In this study, we used the CCtk model to test the hypothesis that selective airway injury initiates profound lung dysfunction through mechanisms that compromise alveolar integrity. Results demonstrate that elimination of the CCSP-expressing cell population results in secondary alveolar inflammation, edema, and depletion of the alveolar type II cell population. On the basis of these data we conclude that selective airway injury can serve as the inciting injury in diseases characterized by severely compromised alveolar function. PMID- 15298854 TI - Estrogen regulates pulmonary alveolar formation, loss, and regeneration in mice. AB - Lung tissue elastic recoil and the dimension and number of pulmonary gas-exchange units (alveoli) are major determinants of gas-exchange function. Loss of gas exchange function accelerates after menopause in the healthy aged and is progressively lost in individuals with chronic obstructive pulmonary disease (COPD). The latter, a disease of midlife and later, though more common in men than in women, is a disease to which women smokers and never smokers may be more susceptible than men; it is characterized by diminished lung tissue elastic recoil and presently irremediable alveolar loss. Ovariectomy in sexually immature rats diminishes the formation of alveoli, and estrogen prevents the diminution. In the present work, we found that estrogen receptor-alpha and estrogen receptor beta, the only recognized mammalian estrogen receptors, are required for the formation of a full complement of alveoli in female mice. However, only the absence of estrogen receptor-beta diminishes lung elastic tissue recoil. Furthermore, ovariectomy in adult mice results, within 3 wk, in loss of alveoli and of alveolar surface area without a change of lung volume. Estrogen replacement, after alveolar loss, induces alveolar regeneration, reversing the architectural effects of ovariectomy. These studies 1) reveal estrogen receptors regulate alveolar size and number in a nonredundant manner, 2) show estrogen is required for maintenance of already formed alveoli and induces alveolar regeneration after their loss in adult ovariectomized mice, and 3) offer the possibility estrogen can slow alveolar loss and induce alveolar regeneration in women with COPD. PMID- 15298855 TI - Ultrafine carbon black particles stimulate proliferation of human airway epithelium via EGF receptor-mediated signaling pathway. AB - Exposure to ambient ultrafine particles induces airway inflammatory reactions and tissue remodeling. In this experiment, to determine whether ultrafine carbon black (ufCB) affects proliferation of airway epithelium and, if so, what the mechanism of action is, we studied human primary bronchial epithelial cell cultures. Incubation of cells in the serum-free medium with ufCB increased incorporations of [(3)H]thymidine and [(3)H]leucine into cells in a time- and dose-dependent manner. This effect was attenuated by Cu- and Zn-containing superoxide dismutase (Cu/Zn SOD) and apocynin, an inhibitor of NADPH oxidase, and completely inhibited by pretreatment with the epidermal growth factor receptor (EGF-R) tyrosine kinase inhibitors AG-1478 and BIBX-1382, and the mitogen activated protein kinase kinase inhibitor PD-98059. Transfection of a dominant negative mutant of H-Ras likewise abolished the effect ufCB. Stimulation with ufCB also induced processing of membrane-anchored proheparin-binding (HB)-EGF, release of soluble HB-EGF into the medium, association of phosphorylated EGF-R and Shc with glutathione-S-transferase-Grb2 fusion protein, and phosphorylation of extracellular signal-regulated kinase (ERK). Pretreatment with AG-1478, [Glu(52)]Diphtheria toxin, a specific inhibitor of HB-EGF, neutralizing HB-EGF antibody, Cu/Zn SOD, and apocynin each inhibited ufCB-induced ERK activation. These results suggest that ufCB causes oxidative stress-mediated proliferation of airway epithelium, involving processing of HB-EGF and the concomitant activation of EGF-R and ERK cascade. PMID- 15298856 TI - Regulation of alveolar epithelial cell phenotypes in fetal sheep: roles of cortisol and lung expansion. AB - Our aim was to determine whether cortisol's effect on alveolar epithelial cell (AEC) phenotypes in the fetus is mediated via a sustained alteration in lung expansion. Chronically catheterized fetal sheep were exposed to 1) saline infusion, 2) cortisol infusion (122-131 days' gestation, 1.5-4.0 mg/day), 3) saline infusion plus reduced lung expansion, or 4) cortisol infusion plus reduced lung expansion. The proportions of type I and II AECs were determined by electron microscopy, and surfactant protein (SP)-A, -B, and -C mRNA levels were determined by Northern blot analysis. Cortisol infusions significantly increased type II AEC proportions (to 38.2 +/- 2.2%), compared with saline-infused fetuses (23.8 +/- 2.4%), and reduced type I AEC proportions (to 59.0 +/- 2.2%), compared with saline-infused fetuses (70.4 +/- 2.4%). Reduced lung expansion also increased type II AEC proportions (to 52.9 +/- 3.5%) and decreased type I AEC proportions (to 34.2 +/- 3.7%), compared with control, saline-infused fetuses. The infusion of cortisol into fetuses exposed to reduced lung expansion tended to further increase type II (to 60.3 +/- 2.1%, P = 0.066) and reduce type I AEC (to 26.6 +/- 2.3%, P = 0.07) proportions. SP-A, -B, and -C mRNA levels changed in parallel with the changes in type II AEC proportions. These results indicate that cortisol alters the proportion of type I and type II AECs via a mechanism unrelated to the degree of fetal lung expansion. However, reductions in fetal lung expansion appear to have a greater impact on the proportion of AECs than cortisol. PMID- 15298857 TI - Connective tissue growth factor expression and induction by transforming growth factor-beta is abrogated by simvastatin via a Rho signaling mechanism. AB - Connective tissue growth factor (CTGF), a potent profibrotic mediator, acts downstream and in concert with transforming growth factor (TGF)-beta to drive fibrogenesis. Significant upregulation of CTGF has been reported in fibrogenic diseases, including idiopathic pulmonary fibrosis (IPF), and is partly responsible for associated excessive fibroblast proliferation and extracellular matrix deposition, but no effective therapy exists for averting such fibrogeneic events. Simvastatin has reported putative antifibrotic actions in renal fibroblasts; this study explores such actions on human IPF-derived and normal lung fibroblasts and examines associated driving mechanisms. Simvastatin reduces basal CTGF gene and protein expression in all fibroblast lines, overriding TGF beta induction through inhibition of the cholesterol synthesis pathway. Signaling pathways driving simvastatin's effects on CTGF/TGF-beta interaction were evaluated using transient reporter transfection of a CTGF promoter construct. Inhibition of CTGF promoter activity by simvastatin was most marked at 10 muM concentration, reducing activity by 76.2 and 51.8% over TGF-beta-stimulated cultures in IPF and normal fibroblasts, respectively. We also show that geranylgeranylpyrophosphate (GGPP), but not farnesylpyrophosphate, induces CTGF promoter activity following simvastatin inhibition by 55.3 and 31.1% over GGPP negative cultures in IMR90 and IPF-derived fibroblasts, respectively, implicating small GTPase Rho involvement rather than Ras in these effects. Indeed, the specific Rho inhibitor C3 exotoxin significantly (P < 0.05) suppressed TGF-beta induced CTGF promoter activity in transfected lung fibroblasts, a finding further supported by transfection of dominant-negative and constitutively active RhoA constructs, thus demonstrating that simvastatin through a Rho signaling mechanism in lung fibroblasts can modulate CTGF expression and interaction with TGF-beta. PMID- 15298858 TI - How to review a manuscript: a "down-to-earth" approach. PMID- 15298859 TI - Biopsychosocial formulation: recognizing educational shortcomings. AB - OBJECTIVE: Since Engel introduced the biopsychosocial model, it has been extensively examined. The authors expect psychiatrists to formulate cases using the biopsychosocial model. However, resident psychiatrists' ability to generate formulations using this model has received little attention. METHODS: The authors evaluated resident biopsychosocial formulations using biopsychosocial scores from trained, blinded raters across four institutions. Second, the authors determined if an intervention could improve biopsychosocial formulation. DESIGN: This study included non-experimental and pre-post components using resident portfolio scores to measure biopsychosocial. PARTICIPANTS/SETTING: Residents from four postgraduate years (PGY) in four different programs participated. In one institution, faculty made a concerted effort to improve biopsychosocial formulation. There were 33 entries in 2000-2001 and 46 entries in 2001-2002. RESULTS: Using the combined data from all institutions, no PGY level averaged a rating of 3.0 (competent) in either year. In the institution implementing an intervention, a significant improvement was noted. CONCLUSION: This pilot study indicates that we can improve resident competency in this area. PMID- 15298860 TI - Residency training in emergency psychiatry: a model curriculum developed by the education committee of the american association for emergency psychiatry. AB - OBJECTIVE: Describe training goals, objectives and requirements in emergency psychiatry to assist residency programs in developing comprehensive training programs to ensure psychiatric residents acquire the necessary skills and knowledge to competently assess and manage patients with psychiatric emergencies. METHODS: The American Association for Emergency Psychiatry (AAEP) Education Committee developed these guidelines using a consensus-building process. CONCLUSION: These guidelines address all aspects of training including objectives, recommended training sites, rotation length, clinical supervision, curriculum content and evaluation. The objectives emphasize acute assessment and intervention skills. The AAEP Education Committee hopes that by implementing these guidelines, training programs will enable residents to become competent and comfortable working in a psychiatric emergency service. PMID- 15298861 TI - Impact of patient suicide on psychiatrists and psychiatric trainees. AB - OBJECTIVE: The authors investigated the impact of patient suicides on trainees and psychiatrists and their utilization of supports. METHODS: Graduates in practice and trainees of the residency program of the University of Toronto from 1980-1995 (N=495) were surveyed, retrospectively, with 239/495 responding (48%). Demographic and educational information, exposure to suicide, impact of the suicide(s), use of support systems, acute stress disorder and posttraumatic stress disorder symptoms, and impact of events and social relationship scores were collected. RESULTS: One-half of the respondents (120/239) experienced at least one suicide of a patient, 62% of them (74/120) during postgraduate training. Biologically oriented psychiatrists in practice were more at risk for patient suicide. An important minority (one-quarter) among those who experienced patient suicide had substantially higher (morbid) scores than the overall group. They also scored higher on an acute stress disorder and a posttraumatic stress disorder symptom checklist. The impact was more severe when the patient suicide occurred during training than after graduation and was inversely correlated with clinicians' perceived social integration into their relational professional network. CONCLUSIONS: The experience of patient suicide is common during training and in clinical practice. The majority of trainees and clinicians are able to cope normally with the trauma, but in an important minority the emotional impact approaches morbid levels. Training programs should prepare students for this occupational hazard and implement systematic protocols to support those trainees who are especially vulnerable to their patient's suicide and reduce their social isolation from their peer group. Formal and informal professional networks should heighten awareness of the impact of patient suicide on practicing colleagues and take adequate measures to support them. PMID- 15298862 TI - The good-enough mentoring relationship. AB - OBJECTIVE: This study aimed to describe the state of mentoring in today's academic environment. METHODS: Resident focus groups from across the nation discussed their opinions about mentoring and experiences with mentoring, and individual faculty members were videotaped discussing the same. RESULTS: Sixty six residents and five faculty members participated in the project. There was consensus among residents and faculty regarding important qualities of a mentoring relationship, obstacles to forming a mentorship, and methods to improve the mentorship experience in psychiatric training. CONCLUSION: Mentoring is still believed to be a vital component of a successful residency experience. Specific components that warrant further research include boundaries within the relationship, ethnicity and gender factors, and potential need for resident and faculty curricula on the topic of mentoring. PMID- 15298863 TI - A survey of american psychiatric residency programs concerning education in homelessness. AB - OBJECTIVES: This study aims to document how psychiatric residencies address homelessness and mental illness, to discover training barriers, and to identify educational recommendations. METHODS: The authors mailed a survey to 178 American psychiatric residency programs, requesting information about didactic and clinical offerings in homelessness. Programs without offerings were asked to provide reasons why. RESULTS: Of 106 responses, 60% had educational offerings. Concerning clinical experiences, most had fewer than 20% of residents rotating, and only 11% had mandatory rotations. Programs without offerings usually noted that training in this area was a low priority, and this was most frequently linked with perceived low community homelessness prevalence. CONCLUSION: Psychiatric residency programs have addressed education in mental illness and homelessness in various ways. That there were few residents in clinical rotations suggests a need to explore causes, including funding problems, and whether there is sufficient academic community psychiatry faculty. The findings also evoke the need for a model curriculum that enables clinical competency in this public health problem. PMID- 15298864 TI - Construct validity of an objective structured clinical examination (OSCE) in psychiatry: associations with the clinical skills examination and other indicators. AB - OBJECTIVE: The construct validity of checklist and global process scores for an objective structured clinical examination (OSCE) in psychiatry was assessed. Multiple regression analysis was used to predict psychiatry OSCE scores from the clinical skills examination, an obstetrics/gynecology (OB/GYN) OSCE, and the National Board of Medical Examiners (NBME) psychiatry subject examination. METHODS: Archival data from two successive classes of third-year medical students (1999-2000, N=142; 2000-2001, N=144) were aggregated and analyzed. RESULTS: The pattern and magnitude of convergence and discrimination were indicative of adequate construct validity for both the psychiatry checklist scores and the global process score. Clinical skills examination scores for history taking, interpersonal skills, and physical examination were related to psychiatry OSCE scores that reflected the same skill set. Construct validity was fairly higher for the global process rating. CONCLUSION: Evidence of construct validity of a psychiatry OSCE was obtained from multiple measures of performance, including the clinical skills examination. Findings lend support to the continued use of checklist and global process evaluations as part of OSCEs in psychiatry. PMID- 15298865 TI - Mission-based reporting in academic psychiatry. AB - OBJECTIVE: This article describes a data entry and analysis system called Mission Based Reporting (MBR) that is used to measure faculty and department activities related to specific academic missions and objectives. The purpose of MBR is to provide a reporting tool useful in evaluating faculty effort and in helping chairs 1) to better assess their department's performance in relation to other departments and their school as a whole, 2) to plan for the future, and 3) to reward individual faculty members. METHODS: Mission-Based Reporting summaries, generated for each faculty member and each department, illustrate contributions to each of four missions: research, teaching, clinical service, and administrative/public service. Data from MBR can be used to evaluate whether faculty scholarly contributions are appropriate to their rank and series. That report provides data from the Department of Psychiatry and Behavioral Sciences at the University of California and the University of California Davis School of Medicine (UC Davis). CONCLUSIONS: Mission-Based Reporting is a useful management tool for department and school administrators. Improvements in implementation are proposed. PMID- 15298866 TI - An educational intervention to improve residents' inpatient charting. AB - OBJECTIVE: This report describes an educational intervention designed to improve psychiatry residents' inpatient charting skills. METHODS: The residency training committee formed a multidisciplinary team to study the problem by using quality improvement principles. The team hypothesized that residents' charting would improve with education about the purpose of the medical record and instruction on the specific components to document. The team designed an educational intervention to train residents to record five items in the chart of every inpatient: an admission note, an off-service note, descriptions of medication changes, daily progress notes, and the name and discipline of the individual recording these items. Prior to the educational intervention, a chart abstractor determined the frequency, with which residents charted the five items. Additional chart audits were conducted 1 month and 6 months following education. RESULTS: Compliance in charting four of the five items improved significantly 1 month after education, and the improvement was maintained after 6 months. CONCLUSION: An educational intervention that is planned and implemented by a multidisciplinary team can enhance an area of resident performance that affects patient care. PMID- 15298867 TI - An electronic filing cabinet for "classic" articles and other teaching materials: something old, something new. AB - OBJECTIVE: "Classic" articles remain an important resource in teaching psychiatry. But, such materials are often buried in a faculty member's filing cabinet, effectively lost in today's world of instant access. We attempt to recover these materials through a marriage of traditional reading lists and contemporary technology. METHOD: The authors created a sharable electronic filing cabinet to store frequently used teaching materials. CONCLUSION: An electronic filing cabinet can organize teaching materials and increase efficiency in teaching. PMID- 15298868 TI - Media violence research and youth violence data: why do they conflict? AB - OBJECTIVE: Contrary to media headlines and public perceptions, there is little evidence of a substantial link between exposure to violent interactive games and serious real-life violence or crime. CONCLUSION: Further research is needed on whether violent games may affect less dramatic but real concerns such as bullying, fighting, or attitudes and beliefs that support aggression, as well as how effects may vary by child characteristics and types of games. There is also a need for research on the potential benefits of violent games for some children and adults. PMID- 15298869 TI - Small-group videotape training for psychotherapy skills development. AB - OBJECTIVE: Psychotherapy instructors have used video technology to train residents for over 40 years. Though it has met with some controversy, many will argue that videotape review is essential for self-directed learning and accurate psychotherapy supervision. METHODS: The author describes a technique of small group videotape training as provided in a psychiatry residency training program. RESULTS: He reviews the merits and limitations of this format and suggests simple and inexpensive technical approaches to augment this training. CONCLUSION: The author concludes that small-group videotape training is an efficient psychotherapy training format that encourages self-monitoring and the exchange of supportive peer feedback. PMID- 15298870 TI - Do we need more creative solutions to increase the number of second-year and beyond geropsychiatry fellows?: a postfellowship viewpoint. PMID- 15298871 TI - Orientation and pore-forming mechanism of a scorpion pore-forming peptide bound to magnetically oriented lipid bilayers. AB - The orientation and pore-forming mechanisms of pandinin 2 (pin2), an antimicrobial peptide isolated from venom of the African scorpion Pandinus imperator, bound to magnetically oriented lipid bilayers were examined by 31P and 13C solid-state, and 15N liquid-state NMR spectroscopy. 31P NMR measurements at various temperatures, under neutral and acidic conditions, showed that membrane lysis occurred only under acidic conditions, and at temperatures below the liquid crystal-gel phase transition of the lipid bilayers, after incubation for two days in the magnet. Differential scanning calorimetry measurements showed that pin2 induced negative curvature strain in lipid bilayers. The 13C chemical shift values of synthetic pin2 labeled at Gly3, Gly8, Leu12, Phe17, or Ser18 under static or slow magic-angle spinning conditions, indicate that pin2 penetrates the membrane with its average helical axis perpendicular to the membrane surface. Furthermore, amide H-D exchange experiments of 15N-Ala4, Gly8, and Ala9 triply labeled pin2 suggest that this peptide forms oligomers and confirms that the N terminal region creates membrane pores. PMID- 15298872 TI - Reversible aggregation plays a crucial role on the folding landscape of p53 core domain. AB - The role of tumor suppressor protein p53 in cell cycle control depends on its flexible and partially unstructured conformation, which makes it crucial to understand its folding landscape. Here we report an intermediate structure of the core domain of the tumor suppressor protein p53 (p53C) during equilibrium and kinetic folding/unfolding transitions induced by guanidinium chloride. This partially folded structure was undetectable when investigated by intrinsic fluorescence. Indeed, the fluorescence data showed a simple two-state transition. On the other hand, analysis of far ultraviolet circular dichroism in 1.0 M guanidinium chloride demonstrated a high content of secondary structure, and the use of an extrinsic fluorescent probe, 4,4'-dianilino-1,1' binaphthyl-5,5' disulfonic acid, indicated an increase in exposure of the hydrophobic core at 1 M guanidinium chloride. This partially folded conformation of p53C was plagued by aggregation, as suggested by one-dimensional NMR and demonstrated by light scattering and gel-filtration chromatography. Dissociation by high pressure of these aggregates reveals the reversibility of the process and that the aggregates have water-excluded cavities. Kinetic measurements show that the intermediate formed in a parallel reaction between unfolded and folded structures and that it is under fine energetic control. They are not only crucial to the folding pathway of p53C but may explain as well the vulnerability of p53C to undergo departure of the native to an inactive state, which makes the cell susceptible to malignant transformation. PMID- 15298873 TI - The assignment of the different infrared continuum absorbance changes observed in the 3000-1800-cm(-1) region during the bacteriorhodopsin photocycle. AB - The bleach continuum in the 1900-1800-cm(-1) region was reported during the photocycle of bacteriorhodopsin (bR) and was assigned to the dissociation of a polarizable proton chain during the proton release step. More recently, a broad band pass filter was used and additional infrared continua have been reported: a bleach at >2700 cm(-1), a bleach in the 2500-2150-cm(-1) region, and an absorptive behavior in the 2100-1800-cm(-1) region. To fully understand the importance of the hydrogen-bonded chains in the mechanism of the proton transport in bR, a detailed study is carried out here. Comparisons are made between the time-resolved Fourier transform infrared spectroscopy experiments on wild-type bR and its E204Q mutant (which has no early proton release), and between the changes in the continua observed in thermally or photothermally heated water (using visible light-absorbing dye) and those observed during the photocycle. The results strongly suggest that, except for the weak bleach in the 1900-1800-cm(-1) region and >2500 cm(-1), there are other infrared continua observed during the bR photocycle, which are inseparable from the changes in the absorption of the solvent water molecules that are photothermally excited via the nonradiative relaxation of the photoexcited retinal chromophore. A possible structure of the hydrogen-bonded system, giving rise to the observed bleach in the 1900-1800-cm( 1) region and the role of the polarizable proton in the proton transport is discussed. PMID- 15298874 TI - Thermal and sodium dodecylsulfate induced transitions of streptavidin. AB - The strong specific binding of streptavidin (SA) to biotin is utilized in numerous biotechnological applications. The SA tetramer is also known to exhibit significant stability, even in the presence of sodium dodecylsulfate (SDS). Despite its importance, relatively little is known about the nature of the thermal denaturation pathway for SA. This work uses a homogeneous SA preparation to expand on the data of previous literature reports, leading to the proposal of a model for temperature induced structural changes in SA. Temperature dependent data were obtained by SDS and native polyacrylamide gel electrophoresis (PAGE), differential scanning calorimetry (DSC), and fluorescence and ultraviolet (UV) visible spectroscopy in the presence and absence of SDS. In addition to the development of this model, it is found that the major thermal transition of SA in 1% SDS is reversible. Finally, although SA exhibits significant precipitation at elevated temperatures in aqueous solution, inclusion of SDS acts to prevent SA aggregation. PMID- 15298875 TI - Thermal injuries in three children caused by an electrical warming mattress. AB - Peroperative hypothermia is recognized to increase mortality and morbidity, and the paediatric anaesthetist faces specific challenges resulting from the increased body surface to volume ratio, particularly in smaller children. We describe three children who were consecutive patients on one operating list and sustained severe thermal injuries. These were due to a malfunctioning electrical heating mat, despite appropriate use and monitoring by the attending anaesthetist. It is rare for thermal warming devices to cause injury. We review the use of heating mats, and suggest modifications in their manufacture which may minimize the risks associated with heating devices. PMID- 15298876 TI - Comparison of the LMA-ProSeal and LMA-Classic in children. AB - BACKGROUND: The LMA-ProSeal is a new laryngeal mask airway with a rear cuff and drainage tube that allows a higher seal pressure than the LMA-Classic for the same intra-cuff pressure, and it permits drainage of gastric secretions and access to the alimentary tract. The LMA-ProSeal can be used in children but it does not have a rear cuff. This study compared the LMA-ProSeal and the LMA Classic in children for ease of insertion, airway sealing pressure and fibre optic visualization. METHODS: Sixty ASA I-II children undergoing herniorrhaphy, orchiopexy or myringotomy were included. The patients were randomly assigned to size 2 LMA-Classic trade mark or size 2 LMA-ProSeal groups for airway management. We assessed success rates at first attempt of insertion, airway sealing pressure, fibre-optic position, success rates of gastric tube placement and postoperative blood staining of the device, tongue-lip-dental trauma and hoarseness. RESULTS: There was no statistical difference between the two groups for the success rates at first attempt of insertion, airway sealing pressure and fibre-optic position. Gastric tube insertion was successful in 90% of cases in the LMA-ProSeal group. The LMA-Classic had a higher rate of postoperative blood staining, but there was no tongue-lip-dental trauma or hoarseness in either group. CONCLUSION: We conclude that ease of insertion and airway sealing pressure are similar between the LMA-ProSeal and the LMA-Classic in children. PMID- 15298877 TI - Comparison of manufacturers' specifications for 44 types of heat and moisture exchanging filters. AB - BACKGROUND: Although heat and moisture exchanging filters (HMEF) are recommended for use during anaesthesia, the criteria for choosing a filter are not clearly defined. Manufacturers offer many different types of HMEF with various technical characteristics. We compared the technical specifications provided by the manufacturers for different types of HMEF. METHODS: Filter manufacturers were asked to provide technical information. Additional information was obtained from websites. Information about 44 filters (16 mechanical and 28 electrostatic) was collated. RESULTS: Filter performances were estimated with different sizes of microorganism and durations of challenge. Twenty-eight filters had not been tested by independent laboratories. For 12 of the filters, information obtained from websites and from the manufacturers differed. Most filter specifications claimed high efficiency, particularly for filtration, microbial challenge number and test duration. Electrostatic filters used in anaesthesia were claimed to have high filtration efficiency, similar to the efficiency provided by mechanical filters. Excluding moisture output values did not alter the general conclusions. CONCLUSIONS: Technical aspects of the tests, international standards, and independent validation should be considered when a filter is chosen. PMID- 15298878 TI - Comparative effectiveness and safety of physician and nurse anaesthetists: a narrative systematic review. AB - BACKGROUND: Despite widespread debate on the merits of different models of anaesthesia care delivery, there are few published data on the relative safety and effectiveness of different anaesthesia providers. METHOD: We conducted a systematic search for, and critical appraisal of, primary research comparing safety and effectiveness of different anaesthetic providers. RESULTS: Our search of Medline, EMBASE, CINAHL, and HMIC for material published between 1990 and April 2003 yielded four articles of relevance to the question. The studies used a variety of methodologies and all had potential confounding factors limiting the validity of the results. CONCLUSIONS: In view of the paucity of high-level primary evidence in this area, it is not possible to draw a conclusion regarding differences in patient safety as a function of provider type. There are difficulties in classifying events as "anaesthesia-related", and also in the variable definitions of "supervision" and "anaesthesia care team". We suggest that existing attempts to show differences in outcome might usefully be complemented by studies examining measures of anaesthetic process. PMID- 15298879 TI - Contamination of anaesthetic gases with nitric oxide and its influence on oxygenation: study in patients undergoing open heart surgery. AB - BACKGROUND: Nitric oxide is important in vasomotor regulation. Contamination of anaesthetic gases with nitric oxide could affect gas exchange. METHODS: We measured oxygenation and nitric oxide concentrations in the inspiratory and expiratory limb of the ventilator circuit in patients about to have cardiac surgery. Measurements were made before surgery when the circulation and respiratory conditions were stable. Fi(o(2)) was set at 0.35. The breathing circuit was supplied with a fresh gas flow greater than the minute volume so that exhaled gas was not re-used. Three gas mixtures were given in sequence to each patient: oxygen and compressed air (AIRc), oxygen and nitrous oxide, and oxygen and synthetic air (AIRs) that was free from nitric oxide. All patients were given AIRs as the second gas and the other two gas mixtures (AIRc and nitrous oxide) were given randomly as the first and third gases. RESULTS: During ventilation with oxygen-AIRc, the median nitric oxide concentration was 5.6 ppb, during ventilation with oxygen-nitrous oxide it was 5.0 ppb and using oxygen-AIRs it was 1.5 ppb. When AIRc and nitrous oxide were used, Pa(o(2)) was greater and venous admixture was less than when AIRs was used. The different gas mixtures did not affect pulmonary vascular pressures or cardiac output. CONCLUSIONS: Compressed air and nitrous oxide contain very low concentrations of nitric oxide (<10 ppb). This can affect pulmonary oxygen transfer during anaesthesia. PMID- 15298880 TI - The learning curve. PMID- 15298881 TI - Improving mortality of coronary surgery over first four years of independent practice: retrospective examination of prospectively collected data from 15 surgeons. AB - OBJECTIVE: To study the "learning curve" associated with independent practice in coronary artery surgery. DESIGN: Retrospective analysis of prospectively collected data. SETTING: All NHS centres in north west England that carry out cardiac surgery in adults. PARTICIPANTS: 18 913 patients undergoing coronary artery surgery for the first time between April 1997 and March 2003, 5678 of whom were operated on by 15 surgeons in the first four years after their consultant appointment. MAIN OUTCOME MEASURES: Observed and predicted mortality (EuroSCORE) for surgeons in their first, second, third, and fourth years after appointment as a consultant compared with figures for established surgeons. RESULTS: Overall mortality decreased over the six years of study (P = 0.01). Of the patients operated on by established surgeons or newly appointed consultants, 265/13,235 (2.0%) and 109/5678 (1.9%), respectively, died (P = 0.71). There was a progressive decrease in observed mortality with time after appointment as a consultant from 2.2% in the first year to 1.2% in the fourth year (P = 0.049). This result remained significant after adjustment for time and case mix (P = 0.019). CONCLUSIONS: Mortality in patients operated on by newly appointed consultant surgeons is similar to mortality in patients operated on by established surgeons. There are significant decreases in crude and risk adjusted mortality in the four years after appointment. These findings should influence the nature of practice in newly appointed surgeons. PMID- 15298882 TI - Signaling in small subcellular volumes. I. Stochastic and diffusion effects on individual pathways. AB - Many cellular signaling events occur in small subcellular volumes and involve low abundance molecular species. This context introduces two major differences from mass-action analyses of nondiffusive signaling. First, reactions involving small numbers of molecules occur in a probabilistic manner which introduces scatter in chemical activities. Second, the timescale of diffusion of molecules between subcellular compartments and the rest of the cell is comparable to the timescale of many chemical reactions, altering the dynamics and outcomes of signaling reactions. This study examines both these effects on information flow through four protein kinase regulatory pathways. The analysis uses Monte Carlo simulations in a subcellular volume diffusively coupled to a bulk cellular volume. Diffusion constants and the volume of the subcellular compartment are systematically varied to account for a range of cellular conditions. Each pathway is characterized in terms of the probabilistic scatter in active kinase levels as a measure of "noise" on the pathway output. Under the conditions reported here, most signaling outcomes in a volume below one femtoliter are severely degraded. Diffusion and subcellular compartmentalization influence the signaling chemistry to give a diversity of signaling outcomes. These outcomes may include washout of the signal, reinforcement of signals, and conversion of steady responses to transients. PMID- 15298883 TI - Signaling in small subcellular volumes. II. Stochastic and diffusion effects on synaptic network properties. AB - The synaptic signaling network is capable of sophisticated cellular computations. These include the ability to respond selectively to different patterns of input, and to sustain changes in response over long periods. The small volume of the synapse complicates the analysis of signaling because the chemical environment is strongly affected by diffusion and stochasticity. This study is based on an updated version of a previously proposed synaptic signaling circuit (Bhalla and Iyengar, 1999) and analyzes three network computation properties in small volumes: bistability, thresholding, and pattern selectivity. Simulations show that although there are diffusive regimes in which bistability may persist, chemical noise at small volumes overwhelms bistability. In the deterministic situation, the network exhibits a sharp threshold for transition between lower and upper stable states. This transition is broadened and individual runs partition between lower and upper states, when stochasticity is considered. The third network property, pattern selectivity, is severely degraded at synaptic volumes. However, there are regimes in which a process similar to stochastic resonance operates and amplifies pattern selectivity. These results imply that simple scaling of signaling conditions to femtoliter volumes is unlikely, and microenvironments, such as reaction complex formation, may be essential for reliable small-volume signaling. PMID- 15298884 TI - Molecular Dynamics Simulations of Micelle Formation around Dimeric Glycophorin A Transmembrane Helices. AB - Insertion and formation of membrane proteins involves the interaction of protein helices with one another in lipid environments. Researchers have studied glycophorin A (GpA) transmembrane helices embedded in sodium dodecyl sulfate (SDS) micelles to identify contacts significant for helix dimerization. However, a detailed picture of the conformation and dynamics of the GpA-SDS system cannot be obtained solely through experiment. Molecular dynamics simulations of SDS and a GpA dimer can provide an atomic-level picture of SDS aggregation and helix association. We report 2.5-ns simulations of GpA wild-type and mutants in a preformed micelle as well as a 32-ns simulation showing the formation of a complete micelle around wild-type GpA from an initially random placement of SDS molecules in an aqueous environment. In the latter case, an initial instability of GpA helices in water is reversed after the helices become surrounded by SDS. The properties of the spontaneously formed micelle surrounding the GpA are indistinguishable from those of the preformed micelle surrounding the GpA dimer. PMID- 15298885 TI - Induced coalescence of cations through low-temperature Poisson-Boltzmann calculations. AB - The computational determination of preferred binding regions of divalent counterions to nucleic acids is either inaccurate (standard Poisson-Boltzmann approaches) or extremely time-consuming (Monte Carlo or molecular dynamics simulations). A novel "selective low-temperature" Poisson-Boltzmann method is introduced that, although approximate in nature, qualitatively accounts for ion correlation and charge-transfer effects and allows for the rapid determination of such regions through an "induced coalescence" of divalent ions. The method is illustrated here for the binding of Mg(2+) to a double-helical sequence of B-form DNA (CGCGAATTCGCG) but the technique is readily applicable to locating divalent cations in other systems such as DNA-endonuclease complexes and ribozymes. PMID- 15298886 TI - Free diffusion of steroid hormones across biomembranes: a simplex search with implicit solvent model calculations. AB - Steroid hormones such as progesterone, testosterone, and estradiol are derived from cholesterol, a major constituent of biomembranes. Although the hormones might be expected to associate with the bilayer in a fashion similar to that of cholesterol, their biological action in regulating transcription of target genes involves transbilayer transfer by free diffusion, which is not observed for cholesterol. We used a novel combination of a continuum-solvent model and the downhill simplex search method for the calculation of the free energy of interaction of these hormones with lipid membranes, and compared these values to that of cholesterol-membrane interaction. The hormones were represented in atomic detail and the membrane as a structureless hydrophobic slab embedded in implicit water. A deep free-energy minimum of approximately -15 kcal/mol was obtained for cholesterol at its most favorable location in the membrane, whereas the most favorable locations for the hormones were associated with shallower minima of 5.0 kcal/mol or higher. The free-energy difference, which is predominantly due to the substitution of cholesterol's hydrophobic tail with polar groups, explains the different manner in which cholesterol and the hormones interact with the membrane. Further calculations were conducted to estimate the rate of transfer of the hormones from the aqueous phase into hexane, and from hexane back into the aqueous phase. The calculated rates agreed reasonably well with measurements in closely related systems. Based on these calculations, we suggest putative pathways for the free diffusion of the hormones across biomembranes. Overall, the calculations imply that the hormones may rapidly cross biomembrane barriers. Implications for gastrointestinal absorption and transfer across the blood-brain barrier and for therapeutic uses are discussed. PMID- 15298887 TI - A molecular dynamics study of Ca(2+)-calmodulin: evidence of interdomain coupling and structural collapse on the nanosecond timescale. AB - A 20-ns molecular dynamics simulation of Ca(2+)-calmodulin (CaM) in explicit solvent is described. Within 5 ns, the extended crystal structure adopts a compact shape similar in dimension to complexes of CaM and target peptides but with a substantially different orientation between the N- and C-terminal domains. Significant interactions are observed between the terminal domains in this compact state, which are mediated through the same regions of CaM that bind to target peptides derived from protein kinases and most other target proteins. The process of compaction is driven by the loss of helical structure in two separate regions between residues 75-79 and 82-86, the latter being driven by unfavorable electrostatic interactions between acidic residues. In the first 5 ns of the simulation, a substantial number of contacts are observed between the first helix of the N-terminal domain and residues 74-77 of the central linker. These contacts are correlated with the closing of the second EF-hand, indicating a mechanism by which they can lower calcium affinity in the N-terminal domain. PMID- 15298888 TI - A model of the closed form of the nicotinic acetylcholine receptor m2 channel pore. AB - The nicotinic acetylcholine receptor is a neurotransmitter-gated ion channel in the postsynaptic membrane. It is composed of five homologous subunits, each of which contributes one transmembrane helix--the M2 helix--to create the channel pore. The M2 helix from the delta subunit is capable of forming a channel by itself. Although a model of the receptor was recently proposed based on a low resolution, cryo-electron microscopy density map, we found that the model does not explain much of the other available experimental data. Here we propose a new model of the M2 channel derived solely from helix packing and symmetry constraints. This model agrees well with experimental results from solid-state NMR, chemical reactivity, and mutagenesis experiments. The model depicts the channel pore, the channel gate, and the residues responsible for cation specificity. PMID- 15298889 TI - Exploring the counterion atmosphere around DNA: what can be learned from molecular dynamics simulations? AB - The counterion distribution around a DNA dodecamer (5'-CGCGAATTCGCG-3') is analyzed using both standard and novel techniques based on state of the art molecular dynamics simulations. Specifically, we have explored the population of Na(+) in the minor groove of DNA duplex, and whether or not a string of Na(+) can replace the spine of hydration in the narrow AATT minor groove. The results suggest that the insertion of Na(+) in the minor groove is a very rare event, but that when once the ion finds specific sites deep inside the groove it can reside there for very long periods of time. According to our simulation the presence of Na(+) inside the groove does not have a dramatic influence in the structure or dynamics of the duplex DNA. The ability of current MD simulations to obtain equilibrated pictures of the counterion atmosphere around DNA is critically discussed. PMID- 15298890 TI - Hydration of enzyme in nonaqueous media is consistent with solvent dependence of its activity. AB - Water plays an important role in enzyme structure and function in aqueous media. That role becomes even more important when one focuses on enzymes in low water media. Here we present results from molecular dynamics simulations of surfactant solubilized subtilisin BPN' in three organic solvents (octane, tetrahydrofuran, and acetonitrile) and in pure water. Trajectories from simulations are analyzed with a focus on enzyme structure, flexibility, and the details of enzyme hydration. The overall enzyme and backbone structures, as well as individual residue flexibility, do not show significant differences between water and the three organic solvents over a timescale of several nanoseconds currently accessible to large-scale molecular dynamics simulations. The key factor that distinguishes molecular-level details in different media is the partitioning of hydration water between the enzyme and the bulk solvent. The enzyme surface and the active site region are well hydrated in aqueous medium, whereas with increasing polarity of the organic solvent (octane --> tetrahydrofuran --> acetonitrile) the hydration water is stripped from the enzyme surface. Water stripping is accompanied by the penetration of tetrahydrofuran and acetonitrile molecules into crevices on the enzyme surface and especially into the active site. More polar organic solvents (tetrahydrofuran and acetonitrile) replace mobile and weakly bound water molecules in the active site and leave primarily the tightly bound water in that region. In contrast, the lack of water stripping in octane allows efficient hydration of the active site uniformly by mobile and weakly bound water and some structural water similar to that in aqueous solution. These differences in the active site hydration are consistent with the inverse dependence of enzymatic activity on organic solvent polarity and indicate that the behavior of hydration water on the enzyme surface and in the active site is an important determinant of biological function especially in low water media. PMID- 15298891 TI - Hill coefficient for estimating the magnitude of cooperativity in gating transitions of voltage-dependent ion channels. AB - A frequently used measure for the extent of cooperativity in ligand binding by an allosteric protein is the Hill coefficient, obtained by fitting data of initial reaction velocity (or fractional binding saturation) as a function of substrate concentration to the Hill equation. Here, it is demonstrated that the simple two state Boltzmann equation that is widely used to fit voltage-activation data of voltage-dependent ion channels is analogous to the Hill equation. A general empiric definition for a Hill coefficient (n(H)) for channel gating transitions that is analogous to the logarithmic potential sensitivity function of Almers is derived. This definition provides a novel framework for interpreting the meaning of the Hill coefficient. In considering three particular and simple gating schemes for a voltage-activated cation channel, the relation of the Hill coefficient to the magnitude and nature of cooperative interactions along the reaction coordinate of channel gating is demonstrated. A possible functional explanation for the low value of the Hill coefficient for gating transitions of the Shaker voltage-activated K(+) channel is suggested. The analogy between the Hill coefficients for ligand binding and for channel gating transitions further points to a unified conceptual framework in analyzing enzymes and channels behavior. PMID- 15298892 TI - Monitoring gramicidin conformations in membranes: a fluorescence approach. AB - We have monitored the membrane-bound channel and nonchannel conformations of gramicidin utilizing red-edge excitation shift (REES), and related fluorescence parameters. In particular, we have used fluorescence lifetime, polarization, quenching, chemical modification, and membrane penetration depth analysis in addition to REES measurements to distinguish these two conformations. Our results show that REES of gramicidin tryptophans can be effectively used to distinguish conformations of membrane-bound gramicidin. The interfacially localized tryptophans in the channel conformation display REES of 7 nm whereas the tryptophans in the nonchannel conformation exhibit REES of 2 nm which highlights the difference in their average environments in terms of localization in the membrane. This is supported by tryptophan penetration depth measurements using the parallax method and fluorescence lifetime and polarization measurements. Further differences in the average tryptophan microenvironments in the two conformations are brought out by fluorescence quenching experiments using acrylamide and chemical modification of the tryptophans by N-bromosuccinimide. In summary, we report novel fluorescence-based approaches to monitor conformations of this important ion channel peptide. Our results offer vital information on the organization and dynamics of the functionally important tryptophan residues in gramicidin. PMID- 15298893 TI - Voltage-gated rearrangements associated with differential beta-subunit modulation of the L-type Ca(2+) channel inactivation. AB - Auxiliary beta-subunits bound to the cytoplasmic alpha(1)-interaction domain of the pore-forming alpha(1C)-subunit are important modulators of voltage-gated Ca(2+) channels. The underlying mechanisms are not yet well understood. We investigated correlations between differential modulation of inactivation by beta(1a)- and beta(2)- subunits and structural responses of the channel to transition into distinct functional states. The NH(2)-termini of the alpha(1C)- and beta-subunits were fused with cyan or yellow fluorescent proteins, and functionally coexpressed in COS1 cells. Fluorescence resonance energy transfer (FRET) between them or with membrane-trapped probes was measured in live cells under voltage clamp. It was found that in the resting state, the tagged NH(2) termini of the alpha(1C)- and beta-subunit fluorophores are separated. Voltage dependent inactivation generates strong FRET between alpha(1C) and beta(1a) suggesting mutual reorientation of the NH(2)-termini, but their distance vis-a vis the plasma membrane is not appreciably changed. These voltage-gated rearrangements were substantially reduced when the beta(1a)-subunit was replaced by beta(2). Differential beta-subunit modulation of inactivation and of FRET between alpha(1C) and beta were eliminated by inhibition of the slow inactivation. Thus, differential beta-subunit modulation of inactivation correlates with the voltage-gated motion between the NH(2)-termini of alpha(1C)- and beta-subunits and targets the mechanism of slow voltage-dependent inactivation. PMID- 15298894 TI - Assembly of plant Shaker-like K(out) channels requires two distinct sites of the channel alpha-subunit. AB - SKOR and GORK are outward-rectifying plant potassium channels from Arabidopsis thaliana. They belong to the Shaker superfamily of voltage-dependent K(+) channels. Channels of this class are composed of four alpha-subunits and subunit assembly is a prerequisite for channel function. In this study the assembly mechanism of SKOR was investigated using the yeast two-hybrid system and functional assays in Xenopus oocytes and in yeast. We demonstrate that SKOR and GORK physically interact and assemble into heteromeric K(out) channels. Deletion mutants and chimeric proteins generated from SKOR and the K(in) channel alpha subunit KAT1 revealed that the cytoplasmic C-terminus of SKOR determines channel assembly. Two domains that are crucial for channel assembly were identified: i), a proximal interacting region comprising a putative cyclic nucleotide-binding domain together with 33 amino acids just upstream of this domain, and ii), a distal interacting region showing some resemblance to the K(T) domain of KAT1. Both regions contributed differently to channel assembly. Whereas the proximal interacting region was found to be active on its own, the distal interacting region required an intact proximal interacting region to be active. K(out) alpha subunits did not assemble with K(in) alpha-subunits because of the absence of interaction between their assembly sites. PMID- 15298895 TI - Effects of Kv1.2 intracellular regions on activation of Kv2.1 channels. AB - Depolarizing voltage steps activate voltage-dependent K(+) (Kv) channels by moving the voltage sensor, which triggers a coupling reaction leading to the opening of the pore. We constructed chimeric channels in which intracellular regions of slowly activating Kv2.1 channels were replaced by respective regions of rapidly activating Kv1.2 channels. Substitution of either the N-terminus, S4 S5 linker, or C-terminus generated chimeric Kv2.1/1.2 channels with a paradoxically slow and approximately exponential activation time course consisting of a fast and a slow component. Using combined chimeras, each of these Kv1.2 regions further slowed activation at the voltage of 0 mV, irrespective of the nature of the other two regions, whereas at the voltage of 40 mV both slowing and accelerating effects were observed. These results suggest voltage-dependent interactions of the three intracellular regions. This observation was quantified by double-mutant cycle analysis. It is concluded that interactions between N terminus, S4-S5 linker, and/or C-terminus modulate the activation time course of Kv2.1 channels and that part of these interactions is voltage dependent. PMID- 15298896 TI - Effect of ADP on Na(+)-Na(+) exchange reaction kinetics of Na,K-ATPase. AB - The whole-cell voltage-clamp technique was used in rat cardiac myocytes to investigate the kinetics of ADP binding to phosphorylated states of Na,K-ATPase and its effects on presteady-state Na(+)-dependent charge movements by this enzyme. Ouabain-sensitive transient currents generated by Na,K-ATPase functioning in electroneutral Na(+)-Na(+) exchange mode were measured at 23 degrees C with pipette ADP concentrations ([ADP]) of up to 4.3 mM and extracellular Na(+) concentrations ([Na](o)) between 36 and 145 mM at membrane potentials (V(M)) from -160 to +80 mV. Analysis of charge-V(M) curves showed that the midpoint potential of charge distribution was shifted toward more positive V(M) both by increasing [ADP] at constant Na(+)(o) and by increasing [Na](o) at constant ADP. The total quantity of mobile charge, on the other hand, was found to be independent of changes in [ADP] or [Na](o). The presence of ADP increased the apparent rate constant for current relaxation at hyperpolarizing V(M) but decreased it at depolarizing V(M) as compared to control (no added ADP), an indication that ADP binding facilitates backward reaction steps during Na(+)-Na(+) exchange while slowing forward reactions. Data analysis using a pseudo three-state model yielded an apparent K(d) of approximately 6 mM for ADP binding to and release from the Na,K-ATPase phosphoenzyme; a value of 130 s(-1) for k(2), a rate constant that groups Na(+) deocclusion/release and the enzyme conformational transition E(1) approximately P --> E(2)-P; a value of 162 s(-1)M(-1) for k(-2), a lumped second order V(M)-independent rate constant describing the reverse reactions; and a Hill coefficient of approximately 1 for Na(+)(o) binding to E(2)-P. The results are consistent with electroneutral release of ADP before Na(+) is deoccluded and released through an ion well. The same approach can be used to study additional charge-moving reactions and associated electrically silent steps of the Na,K-pump and other transporters. PMID- 15298897 TI - Conformational changes of the Ca(2+) regulatory site of the Na(+)-Ca(2+) exchanger detected by FRET. AB - The Na(+)-Ca(2+) exchanger is a plasma membrane protein expressed at high levels in cardiomyocytes. It extrudes 1 Ca(2+) for 3 Na(+) ions entering the cell, regulating intracellular Ca(2+) levels and thereby contractility. Na(+)-Ca(2+) exchanger activity is regulated by intracellular Ca(2+), which binds to a region (amino acids 371-508) within the large cytoplasmic loop between transmembrane segments 5 and 6. Regulatory Ca(2+) activates the exchanger and removes Na(+) dependent inactivation. The physiological role of intracellular Ca(2+) regulation of the exchanger is not yet established. Yellow (YFP) and cyan (CFP) fluorescent proteins were linked to the NH(2)- and CO(2)H-termini of the exchanger Ca(2+) binding domain (CBD) to generate a construct (YFP-CBD-CFP) capable of responding to changes in intracellular Ca(2+) concentrations by FRET efficiency measurements. The two fluorophores linked to the CBD are sufficiently close to generate FRET. FRET efficiency was reduced with increasing Ca(2+) concentrations. Titrations of Ca(2+) concentration versus FRET efficiency indicate a K(D) for Ca(2+) of approximately 140 nM, which increased to approximately 400 nM in the presence of 1 mM Mg(2+). Expression of YFP-CBD-CFP in myocytes, generated changes in FRET associated with contraction, suggesting that NCX is regulated by Ca(2+) on a beat-to-beat basis during excitation-contraction coupling. PMID- 15298898 TI - Homology-modeled structure of the yeast mitochondrial citrate transport protein. AB - We have used homology modeling to construct a three-dimensional model of the yeast mitochondrial citrate transport protein (CTP), based on the recently published x-ray crystal structure of another mitochondrial transport protein, the ADP/ATP carrier. Superposition of the backbone traces of the homology-modeled CTP onto the crystallographically determined ADP carrier structure indicates that the CTP transmembrane domains are well modeled (i.e., root mean square deviation of 0.94 A), whereas the loops facing the intermembrane space and the mitochondrial matrix are less certain (i.e., root mean square deviation values of 0.72-2.06 A). The homology-modeled CTP is consistent with our earlier de novo models of the transporter's transmembrane domains, with respect to residues which face into the transport path. Importantly, the resulting model is consistent with our previous experimental data obtained from measuring reactivity of 34 single cysteine mutants in transmembrane domains 3 and 4 with methanethiosulfonate reagents. The model also points to a likely dimer interface region. In conclusion, our data help to define the substrate translocation pathway in both the modeled CTP structure and the crystallographic ADP carrier structure. PMID- 15298899 TI - Determinants of gating polarity of a connexin 32 hemichannel. AB - There is good evidence supporting the view that the transjunctional voltage sensor (V(j)-sensor) of Cx32 and other Group 1 connexins is contained within a segment of the N-terminus that contributes to the formation of the channel pore. We have shown that the addition of negatively charged amino acid residues at several positions within the first 10 amino acid residues reverses the polarity of V(j)-gating and proposed that channel closure is initiated by the inward movement of this region. Here, we report that positive charge substitutions of the 2nd, 5th, and 8th residues maintain the negative polarity of V(j)-gating. These data are consistent with the original gating model. Surprisingly, some channels containing combinations of positive and/or negative charges at the 2nd and 5th positions display bipolar V(j)-gating. The appearance of bipolar gating does not correlate with relative orientation of charges at this position. However, the voltage sensitivity of bipolar channels correlates with the sign of the charge at the 2nd residue, suggesting that charges at this position may have a larger role in determining gating polarity. Taken together with previous findings, the results suggest that the polarity V(j)-gating is not determined by the sign of the charge lying closest to the cytoplasmic entry of the channel, nor is it likely to result from the reorientation of an electrical dipole contained in the N-terminus. We further explore the mechanism of polarity determination by utilizing the one-dimensional Poisson-Nernst-Plank model to determine the voltage profile of simple model channels containing regions of permanent charge within the channel pore. These considerations demonstrate how local variations in the electric field may influence the polarity and sensitivity of V(j)-gating but are unlikely to account for the appearance of bipolar V(j)-gating. PMID- 15298900 TI - Involvement of a heptad repeat in the carboxyl terminus of the dihydropyridine receptor beta1a subunit in the mechanism of excitation-contraction coupling in skeletal muscle. AB - Chimeras consisting of the homologous skeletal dihydropyridine receptor (DHPR) beta1a subunit and the heterologous cardiac/brain beta2a subunit were used to determine which regions of beta1a were responsible for the skeletal-type excitation-contraction (EC) coupling phenotype. Chimeras were transiently transfected in beta1 knockout myotubes and then voltage-clamped with simultaneous measurement of confocal fluo-4 fluorescence. All chimeras expressed a similar density of DHPR charge movements, indicating that the membrane density of DHPR voltage sensors was not a confounding factor in these studies. The data indicates that a beta1a-specific domain present in the carboxyl terminus, namely the D5 region comprising the last 47 residues (beta1a 478-524), is essential for expression of skeletal-type EC coupling. Furthermore, the location of beta1aD5 immediately downstream from conserved domain D4 is also critical. In contrast, chimeras in which beta1aD5 was swapped by the D5 region of beta2a expressed Ca(2+) transients triggered by the Ca(2+) current, or none at all. A hydrophobic heptad repeat is present in domain D5 of beta1a (L478, V485, V492). To determine the role of this motif, residues in the heptad repeat were mutated to alanines. The triple mutant beta1a(L478A/V485A/V492A) recovered weak skeletal-type EC coupling (DeltaF/F(max) = 0.4 +/- 0.1 vs. 2.7 +/- 0.5 for wild-type beta1a). However, a triple mutant with alanine substitutions at positions out of phase with the heptad repeat, beta1a(S481A/L488A/S495A), was normal (DeltaF/F(max) = 2.1 +/- 0.4). In summary, the presence of the beta1a-specific D5 domain, in its correct position after conserved domain D4, is essential for skeletal-type EC coupling. Furthermore, a heptad repeat in beta1aD5 controls the EC coupling activity. The carboxyl terminal heptad repeat of beta1a might be involved in protein-protein interactions with ryanodine receptor type 1 required for DHPR to ryanodine receptor type 1 signal transmission. PMID- 15298901 TI - Salting out the ionic selectivity of a wide channel: the asymmetry of OmpF. AB - Although the crystallographic structure of the bacterial porin OmpF has been known for a decade, the physical mechanisms of its ionic selectivity are still under investigation. We address this issue in a series of experiments with varied pH, salt concentrations, inverted salt gradient, and charged and uncharged lipids. Measuring reversal potential, we show that OmpF selectivity (traditionally regarded as slightly cationic) depends strongly on pH and salt concentration and is conditionally asymmetric, that is, the calculated selectivity is sensitive to the direction of salt concentration gradient. At neutral pH and subdecimolar salt concentrations the channel exhibits nearly ideal cation selectivity (t(G)(+)=0.98+/-0.01). Substituting neutral DPhPC with DPhPS, we demonstrate that the fixed charge of the host lipid has a small but measurable effect on the channel reversal potential. The available structural information allows for a qualitative explanation of our experimental findings. These findings now lead us to re-examine the ionization state of 102 titratable sites of the OmpF channel. Using standard methods of continuum electrostatics tailored to our particular purpose, we find the charge distribution in the channel as a function of solution acidity and relate the pH-dependent asymmetry in channel selectivity to the pH-dependent asymmetry in charge distribution. In an attempt to find a simple phenomenological description of our results, we also discuss different macroscopic models of electrodiffusion through large channels. PMID- 15298902 TI - The permeability of gap junction channels to probes of different size is dependent on connexin composition and permeant-pore affinities. AB - Gap junctions have traditionally been characterized as nonspecific pores between cells passing molecules up to 1 kDa in molecular mass. Nonetheless, it has become increasingly evident that different members of the connexin (Cx) family mediate quite distinct physiological processes and are often not interchangeable. Consistent with this observation, differences in permeability to natural metabolites have been reported for different connexins, although the physical basis for selectivity has not been established. Comparative studies of different members of the connexin family have provided evidence for ionic charge selectivity, but surprisingly little is known about how connexin composition affects the size of the pore. We have employed a series of Alexa dyes, which share similar structural characteristics but range in size from molecular weight 350 to 760, to probe the permeabilities and size limits of different connexin channels expressed in Xenopus oocytes. Correlated dye transfer and electrical measurements on each cell pair, in conjunction with a three-dimensional mathematical model of dye diffusion in the oocyte system, allowed us to obtain single channel permeabilities for all three dyes in six homotypic and four heterotypic channels. Cx43 and Cx32 channels passed all three dyes with similar efficiency, whereas Cx26, Cx40, and Cx45 channels showed a significant drop-off in permeability with the largest dye. Cx37 channels only showed significant permeability for the smaller two dyes, but at two- to sixfold lower levels than other connexins tested. In the heterotypic cases studied (Cx26/Cx32 and Cx43/Cx37), permeability characteristics were found to resemble the more restrictive parental homotypic channel. The most surprising finding of the study was that the absolute permeabilities calculated for all gap junctional channels in this study are, with one exception, at least 2 orders of magnitude greater than predicted purely on the basis of hindered pore diffusion. Consequently, affinity between the probes and the pore creating an energetically favorable in pore environment, which would elevate permeant concentration within the pore and hence the flux, is strongly implicated. PMID- 15298903 TI - Dynamics of membrane nanotubulation and DNA self-assembly. AB - A localized point-like force applied perpendicular to a vesicular membrane layer, using an optical tweezer, leads to membrane nanotubulation beyond a threshold force. Below the threshold, the force-extension curve shows an elastic response with a fine structure (serrations). Above the threshold the tubulation process exhibits a new reversible flow phase for the multilamellar membrane, which responds viscoelastically. Furthermore, with an oscillatory force applied during tubulation, broad but well-resolved resonances occur in the flow phase, presumably matching the time scales associated with the vesicle-nanotubule coupled system. These nanotubules, anchored to the optical tweezer also provide, for the first time, a direct probe of the real-time dynamics of DNA self-assembly on membranes. Our studies are a step in the direction of analyzing the dynamics of membrane self-assembly and artificial nanofluidic membrane networks. PMID- 15298904 TI - NMR Studies of lipid lateral diffusion in the DMPC/gramicidin D/water system: peptide aggregation and obstruction effects. AB - The PFG-NMR method has been used in macroscopically oriented bilayers to investigate the effect of the peptide gramicidin D on the lateral diffusion of dimyristoylphosphatidylcholine. By varying both the temperature (21-35 degrees C) and the gramicidin content (0-5 mol %) we have introduced solid obstacles into the lipid liquid crystalline bilayer. It was shown that the obstruction effect exerted by the peptide can be described with several different theoretical models, each based on different premises, and that the fit of the models to experimental data gave reasonable results. We found that each gramicidin molecule was surrounded by approximately one layer of bound lipids and that the obstruction from gel phase patches can be described as small solid obstacles. No evidence of linear aggregates of gramicidin, such as those reported by atomic force microscopy in the gel phase, was found. PMID- 15298905 TI - A kinetic study of the growth of fatty acid vesicles. AB - Membrane vesicles composed of fatty acids can be made to grow and divide under laboratory conditions, and thus provide a model system relevant to the emergence of cellular life. Fatty acid vesicles grow spontaneously when alkaline micelles are added to buffered vesicles. To investigate the mechanism of this process, we used stopped-flow kinetics to analyze the dilution of non-exchanging FRET probes incorporated into preformed vesicles during growth. Oleate vesicle growth occurs in two phases (fast and slow), indicating two pathways for the incorporation of fatty acid into preformed vesicles. We propose that the fast phase, which is stoichiometrically limited by the preformed vesicles, results from the formation of a "shell" of fatty acid around a vesicle, followed by rapid transfer of this fatty acid into the preformed vesicle. The slower phase may result from incorporation of fatty acid which had been trapped in an intermediate state. We provide independent evidence for the rapid transformation of micelles into an aggregated intermediate form after transfer from high to low pH. Our results show that the most efficient incorporation of added oleate into oleic acid/oleate vesicles occurs under conditions that avoid a large transient increase in the micelle/vesicle ratio. PMID- 15298906 TI - Orientation and interaction of oblique cylindrical inclusions embedded in a lipid monolayer: a theoretical model for viral fusion peptides. AB - We consider the elastic behavior of flat lipid monolayer embedding cylindrical inclusions oriented obliquely with respect to the monolayer plane. An oblique inclusion models a fusion peptide, a part of a specialized protein capable of inducing merger of biological membranes in the course of fundamental cellular processes. Although the crucial importance of the fusion peptides for membrane merger is well established, the molecular mechanism of their action remains unknown. This analysis is aimed at revealing mechanical deformations and stresses of lipid monolayers induced by the fusion peptides, which, potentially, can destabilize the monolayer structure and enhance membrane fusion. We calculate the deformation of a monolayer embedding a single oblique inclusion and subject to a lateral tension. We analyze the membrane-mediated interactions between two inclusions, taking into account bending of the monolayer and tilt of the hydrocarbon chains with respect to the surface normal. In contrast to a straightforward prediction that the oblique inclusions should induce tilt of the lipid chains, our analysis shows that the monolayer accommodates the oblique inclusion solely by bending. We find that the interaction between two inclusions varies nonmonotonically with the interinclusion distance and decays at large separations as square of the distance, similar to the electrostatic interaction between two electric dipoles in two dimensions. This long-range interaction is predicted to dominate the other interactions previously considered in the literature. PMID- 15298907 TI - The influence of short-chain alcohols on interfacial tension, mechanical properties, area/molecule, and permeability of fluid lipid bilayers. AB - We used micropipette aspiration to directly measure the area compressibility modulus, bending modulus, lysis tension, lysis strain, and area expansion of fluid phase 1-stearoyl, 2-oleoyl phosphatidylcholine (SOPC) lipid bilayers exposed to aqueous solutions of short-chain alcohols at alcohol concentrations ranging from 0.1 to 9.8 M. The order of effectiveness in decreasing mechanical properties and increasing area per molecule was butanol>propanol>ethanol>methanol, although the lysis strain was invariant to alcohol chain-length. Quantitatively, the trend in area compressibility modulus follows Traube's rule of interfacial tension reduction, i.e., for each additional alcohol CH(2) group, the concentration required to reach the same area compressibility modulus was reduced roughly by a factor of 3. We convert our area compressibility data into interfacial tension values to: confirm that Traube's rule is followed for bilayers; show that alcohols decrease the interfacial tension of bilayer-water interfaces less effectively than oil-water interfaces; determine the partition coefficients and standard Gibbs adsorption energy per CH(2) group for adsorption of alcohol into the lipid headgroup region; and predict the increase in area per headgroup as well as the critical radius and line tension of a membrane pore for each concentration and chain-length of alcohol. The area expansion predictions were confirmed by direct measurements of the area expansion of vesicles exposed to flowing alcohol solutions. These measurements were fitted to a membrane kinetic model to find membrane permeability coefficients of short-chain alcohols. Taken together, the evidence presented here supports a view that alcohol partitioning into the bilayer headgroup region, with enhanced partitioning as the chain-length of the alcohol increases, results in chain-length-dependent interfacial tension reduction with concomitant chain-length-dependent reduction in mechanical moduli and membrane thickness. PMID- 15298908 TI - Fluorescence correlation spectroscopy relates rafts in model and native membranes. AB - The lipid raft model has evoked a new perspective on membrane biology. Understanding the structure and dynamics of lipid domains could be a key to many crucial membrane-associated processes in cells. However, one shortcoming in the field is the lack of routinely applicable techniques to measure raft association without perturbation by detergents. We show that both in cell and in domain exhibiting model membranes, fluorescence correlation spectroscopy (FCS) can easily distinguish a raft marker (cholera toxin B subunit bound to ganglioside (GM1) and a nonraft marker (dialkylcarbocyanine dye diI)) by their decidedly different diffusional mobilities. In contrast, these markers exhibit only slightly different mobilities in a homogeneous artificial membrane. Performing cholesterol depletion with methyl-beta-cyclodextrin, which disrupts raft organization, we find an analogous effect of reduced mobility for the nonraft marker in domain-exhibiting artificial membranes and in cell membranes. In contrast, cholesterol depletion has differential effects on the raft marker, cholera toxin B subunit-GM1, rendering it more mobile in artificial domain exhibiting membranes but leaving it immobile in cell membranes, where cytoskeleton disruption is required to achieve higher mobility. Thus, fluorescence correlation spectroscopy promises to be a valuable tool to elucidate lipid raft associations in native cells and to gain deeper insight into the correspondence between model and natural membranes. PMID- 15298909 TI - Fluorescence correlation spectroscopy studies of Peptide and protein binding to phospholipid vesicles. AB - We used fluorescence correlation spectroscopy (FCS) to analyze the binding of fluorescently labeled peptides to lipid vesicles and compared the deduced binding constants to those obtained using other techniques. We used a well-characterized peptide corresponding to the basic effector domain of myristoylated alanine-rich C kinase substrate, MARCKS(151-175), that was fluorescently labeled with Alexa488, and measured its binding to large unilamellar vesicles (diameter approximately 100 nm) composed of phosphatidylcholine and phosphatidylserine or phosphatidylinositol 4,5-bisphosphate. Because the large unilamellar vesicles are significantly larger than the peptide, the correlation times for the free and bound peptide could be distinguished using single color autocorrelation measurements. The molar partition coefficients calculated from the FCS measurements were comparable to those obtained from binding measurements of radioactively labeled MARCKS(151-175) using a centrifugation technique. Moreover, FCS can measure binding of peptides present at very low concentrations (1-10 nmolar), which is difficult or impossible with most other techniques. Our data indicate FCS can be an accurate and valuable tool for studying the interaction of peptides and proteins with lipid membranes. PMID- 15298910 TI - DNA release from lipoplexes by anionic lipids: correlation with lipid mesomorphism, interfacial curvature, and membrane fusion. AB - DNA release from lipoplexes is an essential step during lipofection and is probably a result of charge neutralization by cellular anionic lipids. As a model system to test this possibility, fluorescence resonance energy transfer between DNA and lipid covalently labeled with Cy3 and BODIPY, respectively, was used to monitor the release of DNA from lipid surfaces induced by anionic liposomes. The separation of DNA from lipid measured this way was considerably slower and less complete than that estimated with noncovalently labeled DNA, and depends on the lipid composition of both lipoplexes and anionic liposomes. This result was confirmed by centrifugal separation of released DNA and lipid. X-ray diffraction revealed a clear correlation of the DNA release capacity of the anionic lipids with the interfacial curvature of the mesomorphic structures developed when the anionic and cationic liposomes were mixed. DNA release also correlated with the rate of fusion of anionic liposomes with lipoplexes. It is concluded that the tendency to fuse and the phase preference of the mixed lipid membranes are key factors for the rate and extent of DNA release. The approach presented emphasizes the importance of the lipid composition of both lipoplexes and target membranes and suggests optimal transfection may be obtained by tailoring lipoplex composition to the lipid composition of target cells. PMID- 15298912 TI - Lessons of slicing membranes: interplay of packing, free area, and lateral diffusion in phospholipid/cholesterol bilayers. AB - We employ 100-ns molecular dynamics simulations to study the influence of cholesterol on structural and dynamic properties of dipalmitoylphosphatidylcholine bilayers in the fluid phase. The effects of the cholesterol content on the bilayer structure are considered by varying the cholesterol concentration between 0 and 50%. We concentrate on the free area in the membrane and investigate quantities that are likely to be affected by changes in the free area and free volume properties. It is found that cholesterol has a strong impact on the free area properties of the bilayer. The changes in the amount of free area are shown to be intimately related to alterations in molecular packing, ordering of phospholipid tails, and behavior of compressibility moduli. Also the behavior of the lateral diffusion of both dipalmitoylphosphatidylcholine and cholesterol molecules with an increasing amount of cholesterol can in part be understood in terms of free area. Summarizing, our results highlight the central role of free area in comprehending the structural and dynamic properties of membranes containing cholesterol. PMID- 15298911 TI - Effect of nanomolar concentrations of sodium dodecyl sulfate, a catalytic inductor of alpha-helices, on human calcitonin incorporation and channel formation in planar lipid membranes. AB - Human Calcitonin (hCt) is a peptide hormone which has a regulatory action in calcium-phosphorus metabolism. It is currently used as a therapeutic tool in bone pathologies such as osteoporosis and Paget's disease. However, due to its amphiphilic property tends to form a gelatinous solution in water which consists of fibrils that limits its therapeutic use. Here we show that sodium dodecyl sulfate (SDS), an anionic detergent able to induce and stabilize alpha-helices in polypeptides, at a monomeric concentration ranging between 0.26 mM-5 pM (all concentrations are below the CMC), increases the rate and number of hCt channel formation in planar lipid membranes, at both high and low hCt concentrations, with a maximum increase at a molecular hCt/SDS ratio of 1000:1. This effect could be interpreted as a counteraction to the fibrillation process of hCt molecules by removing molecules available for aggregation from the fluid; furthermore, this action, independently of channel formation in the cell membrane, could improve the peptide-receptor interaction. The action of SDS could be attributable to the strength of the sulfate negative charge and the hydrophobic chain; in fact, a similar effect was obtained with lauryl sarcosine and not with a neutral detergent such as n-dodecyl-beta-D-maltoside. The very low molecular ratio between SDS and peptide is suggestive of a possible catalytic action of SDS that could induce alpha-helices, the appropriate structures for interacting with the membrane. Moreover, in the experimental conditions investigated, the addition of SDS does not modify the membrane's electrical properties and most of the channel properties. This finding may contribute to the knowledge of environment-folding diseases due to protein and peptides. PMID- 15298913 TI - Sphingomyelin-cholesterol domains in phospholipid membranes: atomistic simulation. AB - We have carried out an atomic-level molecular dynamics simulation of a system of nanoscopic size containing a domain of 18:0 sphingomyelin and cholesterol embedded in a fully hydrated dioleylposphatidylcholine (DOPC) bilayer. To analyze the interaction between the domain and the surrounding phospholipid, we calculate order parameters and area per molecule as a function of molecule type and proximity to the domain. We propose an algorithm based on Voronoi tessellation for the calculation of the area per molecule of various constituents in this ternary mixture. The calculated areas per sphingomyelin and cholesterol are in agreement with previous simulations. The simulation reveals that the presence of the liquid-ordered domain changes the packing properties of DOPC bilayer at a distance as large as approximately 8 nm. We calculate electron density profiles and also calculate the difference in the thickness between the domain and the surrounding DOPC bilayer. The calculated difference in thickness is consistent with data obtained in atomic force microscopy experiments. PMID- 15298914 TI - Ca-activation and stretch-activation in insect flight muscle. AB - Asynchronous insect flight muscle is specialized for myogenic oscillatory work, but can also produce isometric tetanic contraction. In skinned insect flight muscle fibers from Lethocerus, with sarcomere length monitored by a striation follower, we determined the relation between isometric force (F(0)) at serial increments of [Ca(2+)] and the additional active force recruited at each [Ca(2+)] by a stretch of approximately 12 nm per half-sarcomere (F(SA)). The isometric force-pCa relation shows that 1.5-2 units of pCa are necessary to raise isometric force from its threshold (pCa approximately 6.5) to its maximum (F(0,max)). The amplitude of F(SA) depends only on the preceding baseline level of isometric force, which must reach at least 0.05 F(0,max) to enable stretch-activation. F(SA) rises very steeply to its maximum as F(0) reaches approximately 0.2 F(0,max), then decreases as F(0) increases so as to produce a constant sum (F(0) + F(SA)) = F(max). Thus Ca- and stretch-activation are complementary pathways that trigger a common process of cross-bridge attachment and force production. We suggest that stretch-induced distortion of attached cross-bridges relieves the steric blocking by tropomyosin of additional binding sites on actin, thereby enabling maximum force even at low [Ca(2+)]. PMID- 15298915 TI - The elasticity of single titin molecules using a two-bead optical tweezers assay. AB - Titin is responsible for the passive elasticity of the muscle sarcomere. The mechanical properties of skeletal and cardiac muscle titin were characterized in single molecules using a novel dual optical tweezers assay. Antibody pairs were attached to beads and used to select the whole molecule, I-band, A-band, a tandem immunoglobulin (Ig) segment, and the PEVK region. A construct from the PEVK region expressing >25% of the full-length skeletal muscle isoform was chemically conjugated to beads and similarly characterized. By elucidating the elasticity of the different regions, we showed directly for the first time, to our knowledge, that two entropic components act in series in the skeletal muscle titin I-band (confirming previous speculations), one associated with tandem-immunoglobulin domains and the other with the PEVK region, with persistence lengths of 2.9 nm and 0.76 nm, respectively (150 mM ionic strength, 22 degrees C). Novel findings were: the persistence length of the PEVK component rose (0.4-2.7 nm) with an increase in ionic strength (15-300 mM) and fell (3.0-0.3 nm) with a temperature increase (10-60 degrees C); stress-relaxation in 10-12-nm steps was observed in the PEVK construct and hysteresis in the native PEVK region. The region may not be a pure random coil, as previously thought, but contains structured elements, possibly with hydrophobic interactions. PMID- 15298917 TI - Characterization of f-actin tryptophan phosphorescence in the presence and absence of tryptophan-free myosin motor domain. AB - The effect of binding the Trp-free motor domain mutant of Dictyostelium discoideum, rabbit skeletal muscle myosin S1, and tropomyosin on the dynamics and conformation of actin filaments was characterized by an analysis of steady-state tryptophan phosphorescence spectra and phosphorescence decay kinetics over a temperature range of 140-293 K. The binding of the Trp-free motor domain mutant of D. discoideum to actin caused red shifts in the phosphorescence spectrum of two internal Trp residues of actin and affected the intrinsic lifetime of each emitter, decreasing by roughly twofold the short phosphorescence lifetime components (tau(1) and tau(2)) and increasing by approximately 20% the longest component (tau(3)). The alteration of actin phosphorescence by the motor protein suggests that i), structural changes occur deep down in the core of actin and that ii), subtle changes in conformation appear also on the surface but in regions distant from the motor domain binding site. When actin formed complexes with skeletal S1, an extra phosphorescence lifetime component appeared (tau(4), twice as long as tau(3)) in the phosphorescence decay that is absent in the isolated proteins. The lack of this extra component in the analogous actin-Trp free motor domain mutant of D. discoideum complex suggests that it should be assigned to Trps in S1 that in the complex attain a more compact local structure. Our data indicated that the binding of tropomyosin to actin filaments had no effect on the structure or flexibility of actin observable by this technique. PMID- 15298916 TI - Formation and destabilization of actin filaments with tetramethylrhodamine modified actin. AB - Actin labeling at Cys(374) with tethramethylrhodamine derivatives (TMR-actin) has been widely used for direct observation of the in vitro filaments growth, branching, and treadmilling, as well as for the in vivo visualization of actin cytoskeleton. The advantage of TMR-actin is that it does not lock actin in filaments (as rhodamine-phalloidin does), possibly allowing for its use in investigating the dynamic assembly behavior of actin polymers. Although it is established that TMR-actin alone is polymerization incompetent, the impact of its copolymerization with unlabeled actin on filament structure and dynamics has not been tested yet. In this study, we show that TMR-actin perturbs the filaments structure when copolymerized with unlabeled actin; the resulting filaments are more fragile and shorter than the control filaments. Due to the increased severing of copolymer filaments, TMR-actin accelerates the polymerization of unlabeled actin in solution also at mole ratios lower than those used in most fluorescence microscopy experiments. The destabilizing and severing effect of TMR actin is countered by filament stabilizing factors, phalloidin, S1, and tropomyosin. These results point to an analogy between the effects of TMR-actin and severing proteins on F-actin, and imply that TMR-actin may be inappropriate for investigations of actin filaments dynamics. PMID- 15298918 TI - Porphyrin depth in lipid bilayers as determined by iodide and parallax fluorescence quenching methods and its effect on photosensitizing efficiency. AB - Photosensitization by porphyrins and other tetrapyrrole chromophores is used in biology and medicine to kill cells. This light-triggered generation of singlet oxygen is used to eradicate cancer cells in a process dubbed "photodynamic therapy," or PDT. Most photosensitizers are of amphiphilic character and they partition into cellular lipid membranes. The photodamage that they inflict to the host cell is mainly localized in membrane proteins. This photosensitized damage must occur in competition with the rapid diffusion of singlet oxygen through the lipid phase and its escape into the aqueous phase. In this article we show that the extent of damage can be modulated by employing modified hemato- and protoporphyrins, which have alkyl spacers of varying lengths between the tetrapyrrole ring and the carboxylate groups that are anchored at the lipid/water interface. The chromophore part of the molecule, and the point of generation of singlet oxygen, is thus located at a deeper position in the bilayer. The photosensitization efficiency was measured with 9,10-dimethylanthracene, a fluorescent chemical target for singlet oxygen. The vertical insertion of the sensitizers was assessed by two fluorescence-quenching techniques: by iodide ions that come from the aqueous phase; and by spin-probe-labeled phospholipids, that are incorporated into the bilayer, using the parallax method. These methods also show that temperature has a small effect on the depth when the membrane is in the liquid phase. However, when the bilayer undergoes a phase transition to the solid gel phase, the porphyrins are extruded toward the water interface as the temperature is lowered. These results, together with a previous publication in this journal, represent a unique and precedental case where the vertical location of a small molecule in a membrane has an effect on its membranal activity. PMID- 15298919 TI - Lamellar organization of pigments in chlorosomes, the light harvesting complexes of green photosynthetic bacteria. AB - Chlorosomes of green photosynthetic bacteria constitute the most efficient light harvesting complexes found in nature. In addition, the chlorosome is the only known photosynthetic system where the majority of pigments (BChl) is not organized in pigment-protein complexes but instead is assembled into aggregates. Because of the unusual organization, the chlorosome structure has not been resolved and only models, in which BChl pigments were organized into large rods, were proposed on the basis of freeze-fracture electron microscopy and spectroscopic constraints. We have obtained the first high-resolution images of chlorosomes from the green sulfur bacterium Chlorobium tepidum by cryoelectron microscopy. Cryoelectron microscopy images revealed dense striations approximately 20 A apart. X-ray scattering from chlorosomes exhibited a feature with the same approximately 20 A spacing. No evidence for the rod models was obtained. The observed spacing and tilt-series cryoelectron microscopy projections are compatible with a lamellar model, in which BChl molecules aggregate into semicrystalline lateral arrays. The diffraction data further indicate that arrays are built from BChl dimers. The arrays form undulating lamellae, which, in turn, are held together by interdigitated esterifying alcohol tails, carotenoids, and lipids. The lamellar model is consistent with earlier spectroscopic data and provides insight into chlorosome self-assembly. PMID- 15298920 TI - Small angle X-ray scattering studies and modeling of Eudistylia vancouverii chlorocruorin and Macrobdella decora hemoglobin. AB - Annelids possess giant extracellular oxygen carriers that exhibit a hexagonal bilayer appearance and have molecular masses of approximately 3.5 MDa. By small angle x-ray scattering (SAXS), Eudistylia vancouverii chlorocruorin and Macrobdella decora hemoglobin were investigated in solution. On the basis of the experimental SAXS data, three-dimensional models were established in a two-step approach (trial and error and averaging). The main differences between the complexes concern the structure of their central part and the subunit architecture. Usage of our SAXS models as templates for automated model generation (program DAMMIN) led to refined models that fit perfectly the experimental data. Special attention was paid to the inhomogeneous density distribution observed within the complexes. DAMMIN models without a priori information could not reproducibly locate low-density areas. The usage of templates, however, improved the results considerably, in particular by applying electron microscopy-based templates. Biologically relevant information on the presence of low-density areas and hints for their presumable location could be drawn from SAXS and sophisticated modeling approaches. Provided that different models are analyzed carefully, this obviously opens a way to gain additional biologically relevant structural information from SAXS data. PMID- 15298921 TI - Spectroscopic and interfacial properties of myoglobin/surfactant complexes. AB - The complexes of horse myoglobin (Mb) with the anionic surfactant sodium dodecyl sulfate (SDS), and with the cationic surfactants cetyltrimethylammonium chloride (CTAC) and decyltrimethylammonium bromide (DeTAB), have been studied by a combination of surface tension measurements and optical spectroscopy, including heme absorption and aromatic amino acid fluorescence. SDS interacts in a monomeric form with Mb, which suggests the existence of a specific binding site for SDS, and induces the formation of a hexacoordinated Mb heme, possibly involving the distal histidine. Fluorescence spectra display an increase of tryptophan emission. Both effects point to an increased protein flexibility. SDS micelles induce both the appearance of two more heme species, one of which has the features of free heme, and protein unfolding. Mb/CTAC complexes display a very different behavior. CTAC monomers have no effect on the absorption spectra, and only a slight effect on the fluorescence spectra, whereas the formation of CTAC aggregates on the protein strongly affects both absorption and fluorescence. Mb/DeTAB complexes behave in a very similar way as Mb/CTAC complexes. The surface activity of the different Mb/surfactant complexes, as well as the interactions between the surfactants and Mb, are discussed on the basis of their structural properties. PMID- 15298922 TI - Neuroglobin and other hexacoordinated hemoglobins show a weak temperature dependence of oxygen binding. AB - Mouse and human neuroglobins, as well as the hemoglobins from Drosophila melanogaster and Arabidopsis thaliana, were recombinantly expressed in Escherichia coli, and their ligand-binding properties were studied versus temperature. These globins have a common feature of being hexacoordinated (via the distal histidine) under deoxy conditions, as evidenced by a large amplitude for the alpha absorption band at 560 nm and the Soret band at 426 nm. The transition from the hexacoordinated form to the CO bound species is slow, as expected for a replacement reaction Fe-His --> Fe --> FeCO. The intrinsic binding rates would indicate a high oxygen affinity for the pentacoordinated form, due to rapid association and slow (100 ms-1 s) dissociation. However, the competing protein ligand results in a much lower affinity, on the order of magnitude of 1 torr. In addition to decreasing the affinity for external ligand, the competitive internal ligand leads to a weaker observed temperature dependence of the ligand affinity, since the difference in equilibrium energy for the two ligands is much lower than that of ligand binding to pentacoordinated hemoglobin. This effect could be of biological relevance for certain organisms, since it could provide a globin with an oxygen affinity that is nearly independent of temperature. PMID- 15298923 TI - (1)H/(15)N heteronuclear NMR spectroscopy shows four dynamic domains for phospholamban reconstituted in dodecylphosphocholine micelles. AB - We report the backbone dynamics of monomeric phospholamban in dodecylphosphocholine micelles using (1)H/(15)N heteronuclear NMR spectroscopy. Phospholamban is a 52-amino acid membrane protein that regulates Ca-ATPase in cardiac muscle. Phospholamban comprises three structural domains: a transmembrane domain from residues 22 to 52, a connecting loop from 17 to 21, and a cytoplasmic domain from 1 to 16 that is organized in an "L"-shaped structure where the transmembrane and the cytoplasmic domain form an angle of approximately 80 degrees (Zamoon et al., 2003; Mascioni et al., 2002). T(1), T(2), and (1)H/(15)N nuclear Overhauser effect values measured for the amide backbone resonances were interpreted using the model-free approach of Lipari and Szabo. The results point to the existence of four dynamic domains, revealing the overall plasticity of the cytoplasmic helix, the flexible loop, and part of the transmembrane domain (residues 22-30). In addition, using Carr-Purcell-Meiboom-Gill-based experiments, we have characterized phospholamban dynamics in the micros-ms timescale. We found that the majority of the residues in the cytoplasmic domain, the flexible loop, and the first ten residues of the transmembrane domain undergo dynamics in the micros-ms range, whereas minimal dynamics were detected for the transmembrane domain. Hydrogen/deuterium exchange factors measured at different temperatures support the existence of slow motion in both the loop and the cytoplasmic helix. We propose that these dynamic properties are critical factors in the biomolecular recognition of phospholamban by Ca-ATPase and other interacting proteins such as protein kinase A and protein phosphatase 1. PMID- 15298924 TI - Structure of human annexin a6 at the air-water interface and in a membrane-bound state. AB - We postulate the existence of a pH-sensitive domain in annexin A6 (AnxA6), on the basis of our observation of pH-dependent conformational and orientation changes of this protein and its N- (AnxA6a) and C-terminal (AnxA6b) halves in the presence of lipids. Brewster angle microscopy shows that AnxA6, AnxA6a, and AnxA6b in the absence of lipids accumulate at the air-water interface and form a stable, homogeneous layer at pH below 6.0. Under these conditions polarization modulation IR absorption spectroscopy reveals significant conformational changes of AnxA6a whereas AnxA6b preserves its alpha-helical structure. The orientation of protein alpha-helices is parallel with respect to the interface. In the presence of lipids, polarization modulation IR reflection absorption spectroscopy experiments suggest that AnxA6a incorporates into the lipid/air interface, whereas AnxA6b is adsorbed under the lipid monolayer. In this case AnxA6a regains its alpha-helical structures. At a higher pressure of the lipid monolayer the average orientation of the alpha-helices of AnxA6a changes from flat to tilted by 45 degrees with respect to normal to the membrane interface. For AnxA6b no such changes are detected, even at a high pressure of the lipid monolayer-suggesting that the putative pH-sensitive domain of AnxA6 is localized in the N-terminal half of the protein. PMID- 15298925 TI - The role of unstructured extensions in the rotational diffusion properties of a globular protein: the example of the titin i27 module. AB - The possibility of predicting the overall shape of a macromolecule in solution from its diffusional properties has gained increasing importance in the structural genomic era. Here we explore and quantify the influence that unstructured and flexible regions have on the motions of a globular protein, a situation that can occur from the presence of such regions in the natural sequence or from additional tags. I27, an immunoglobulin-like module from the muscle protein titin, whose structure and properties are well characterized, was selected for our studies. The backbone dynamics and the overall tumbling of three different constructs of I27 were investigated using (15)N NMR relaxation collected at two (15)N frequencies (60.8 and 81.1 MHz) and fluorescence depolarization spectroscopy after labeling of a reactive cysteine with an extrinsic fluorophore. Our data show that the presence of disordered tags clearly exerts a frictional drag that increases with the length of the tags, thus affecting the module tumbling in solution. We discuss the use and the limitations of current approaches to hydrodynamic calculations, especially when having to take into account local flexibility. PMID- 15298926 TI - Application of sparse NMR restraints to large-scale protein structure prediction. AB - The protein structure prediction algorithm TOUCHSTONEX that uses sparse distance restraints derived from NMR nuclear Overhauser enhancement (NOE) data to predict protein structures at low-to-medium resolution was evaluated as follows: First, a representative benchmark set of the Protein Data Bank library consisting of 1365 proteins up to 200 residues was employed. Using N/8 simulated long-range restraints, where N is the number of residues, 1023 (75%) proteins were folded to a C(alpha) root-mean-square deviation (RMSD) from native <6.5 A in one of the top five models. The average RMSD of the models for all 1365 proteins is 5.0 A. Using N/4 simulated restraints, 1206 (88%) proteins were folded to a RMSD <6.5 A and the average RMSD improved to 4.1 A. Then, 69 proteins with experimental NMR data were used. Using long-range NOE-derived restraints, 47 proteins were folded to a RMSD <6.5 A with N/8 restraints and 61 proteins were folded to a RMSD <6.5 A with N/4 restraints. Thus, TOUCHSTONEX can be a tool for NMR-based rapid structure determination, as well as used in other experimental methods that can provide tertiary restraint information. PMID- 15298927 TI - Self-assembly of the ionic peptide EAK16: the effect of charge distributions on self-assembly. AB - Amphiphilic peptides suspended in aqueous solution display a rich set of aggregation behavior. Molecular-level studies of relatively simple amphiphilic molecules under controlled conditions are an essential step toward a better understanding of self-assembly phenomena of naturally occurring peptides/proteins. Here, we study the influence of molecular architecture and interactions on the self-assembly of model peptides (EAK16s), using both experimental and theoretical approaches. Three different types of EAK16 were studied: EAK16-I, -II, and -IV, which have the same amino acid composition but different amino acid sequences. Atomic force microscopy confirms that EAK16-I and -II form fibrillar assemblies, whereas EAK16-IV forms globular structures. The Fourier transform infrared spectrum of EAK16-IV indicates the possible formation of a beta-turn structure, which is not found in EAK16-I and -II. Our theoretical and numerical studies suggest the underlying mechanism behind these observations. We show that the hairpin structure is energetically stable for EAK16-IV, whereas the chain entropy of EAK16-I and -II favors relatively stretched conformations. Our combined experimental and theoretical approaches provide a clear picture of the interplay between single-chain properties, as determined by peptide sequences (or charge distributions), and the emerging structure at the nano (or more coarse grained) level. PMID- 15298929 TI - Total internal reflection with fluorescence correlation spectroscopy: nonfluorescent competitors. AB - Total internal reflection with fluorescence correlation spectroscopy is a method for measuring the surface association/dissociation rate constants and absolute densities of fluorescent molecules at the interface of a planar substrate and solution. This method can also report the apparent diffusion coefficient and absolute concentration of fluorescent molecules very close to the surface. Theoretical expressions for the fluorescence fluctuation autocorrelation function when both surface association/dissociation kinetics and diffusion through the evanescent wave, in solution, contribute to the fluorescence fluctuations have been published previously. In the work described here, the nature of the autocorrelation function when both surface association/dissociation kinetics and diffusion through the evanescent wave contribute to the fluorescence fluctuations, and when fluorescent and nonfluorescent molecules compete for surface binding sites, is described. The autocorrelation function depends in general on the kinetic association and dissociation rate constants of the fluorescent and nonfluorescent molecules, the surface site density, the concentrations of fluorescent and nonfluorescent molecules in solution, the solution diffusion coefficients of the two chemical species, the depth of the evanescent field, and the size of the observed area on the surface. Both general and approximate expressions are presented. PMID- 15298928 TI - Spatial-temporal studies of membrane dynamics: scanning fluorescence correlation spectroscopy (SFCS). AB - Giant unilamellar vesicles (GUVs) have been widely used as a model membrane system to study membrane organization, dynamics, and protein-membrane interactions. Most recent studies have relied on imaging methods, which require good contrast for image resolution. Multiple sequential image processing only detects slow components of membrane dynamics. We have developed a new fluorescence correlation spectroscopy (FCS) technique, termed scanning FCS (i.e., SFCS), which performs multiple FCS measurements simultaneously by rapidly directing the excitation laser beam in a uniform (circular) scan across the bilayer of the GUVs in a repetitive fashion. The scan rate is fast compared to the diffusion of the membrane proteins and even small molecules in the GUVs. Scanning FCS outputs a "carpet" of timed fluorescence intensity fluctuations at specific points along the scan. In this study, GUVs were assembled from rat kidney brush border membranes, which included the integral membrane proteins. Scanning FCS measurements on GUVs allowed for a straightforward detection of spatial-temporal interactions between the protein and the membrane based on the diffusion rate of the protein. To test for protein incorporation into the bilayers of the GUVs, antibodies against one specific membrane protein (NaPi II cotransporter) were labeled with ALEXA-488. Fluorescence images of the GUVs in the presence of the labeled antibody showed marginal fluorescence enhancement on the GUV membrane bilayers (poor image contrast and resolution). With the application of scanning FCS, the binding of the antibody to the GUVs was detected directly from the analysis of diffusion rates of the fluorescent antibody. The diffusion coefficient of the antibody bound to NaPi II in the GUVs was approximately 200-fold smaller than that in solution. Scanning FCS provided a simple, quantitative, yet highly sensitive method to study protein-membrane interactions. PMID- 15298930 TI - Dynamic light scattering microscopy. A novel optical technique to image submicroscopic motions. I: theory. AB - The theoretical basis of an optical microscope technique to image dynamically scattered light fluctuation decay rates (dynamic light scattering microscopy) is developed. It is shown that relative motions between scattering centers even smaller than the optical resolution of the microscope are sufficient to produce significant phase variations resulting in interference intensity fluctuations in the image plane. The timescale and time dependence for the temporal autocorrelation function of these intensity fluctuations is derived. The spatial correlation distance, which reports the average distance between constructive and destructive interference in the image plane, is calculated and compared with the pixel size, and the distance dependence of the spatial correlation function is derived. The accompanying article in this issue describes an experimental implementation of dynamic light scattering microscopy. PMID- 15298931 TI - Dynamic light scattering microscopy. A novel optical technique to image submicroscopic motions. II: Experimental applications. AB - An experimental verification of an optical microscope technique to create spatial map images of dynamically scattered light fluctuation decay rates is presented. The dynamic light scattering microscopy technique is demonstrated on polystyrene beads and living macrophage cells. With a slow progressive scan charge-coupled device camera employed in a streak-like mode, rapid intensity fluctuations with timescales the order of milliseconds can be recorded from these samples. From such streak images, the autocorrelation function of these fluctuations can be computed at each location in the sample. The characteristic decay times of the autocorrelation functions report the rates of motion of scattering centers. These rates show reasonable agreement to theoretically expected values for known samples with good signal/noise ratio. The rates can be used to construct an image like spatial map of the rapidity of submicroscopic motions of scattering centers. PMID- 15298932 TI - Elastic light scattering from single cells: orientational dynamics in optical trap. AB - Light-scattering diagrams (phase functions) from single living cells and beads suspended in an optical trap were recorded with 30-ms time resolution. The intensity of the scattered light was recorded over an angular range of 0.5-179.5 degrees using an optical setup based on an elliptical mirror and rotating aperture. Experiments revealed that light-scattering diagrams from biological cells exhibit significant and complex time dependence. We have attributed this dependence to the cell's orientational dynamics within the trap. We have also used experimentally measured phase function information to calculate the time dependence of the optical radiation pressure force on the trapped particle and show how it changes depending on the orientation of the particle. Relevance of these experiments to potential improvement in the sensitivity of label-free flow cytometry is discussed. PMID- 15298933 TI - Evaluation of the ordering of membranes in multilayer stacks built on an ATR-FTIR germanium crystal with atomic force microscopy: the case of the H(+),K(+)-ATPase containing gastric tubulovesicle membranes. AB - Polarized attenuated total reflection Fourier transform infrared spectra were recorded on multilayer stacks of native gastric tubulovesicle membranes. The spectral intensity and linear dichroism were measured for average thicknesses ranging between 0 and 100 bilayers. Atomic force microscopy was used to investigate the orientation of the membranes at the top of the stack. Height profiles were obtained along randomly drawn lines and slopes were computed over various distances. Orientation distribution functions were obtained from the slopes and decomposed into Legendre polynomials. It was found that the second Legendre polynomials coefficient characterizing the membrane orientation was always larger than 0.9. It could therefore be concluded that the membrane tilt does not significantly contribute to the infrared dichroism, even for the largest thicknesses tested. PMID- 15298934 TI - Adenovirus type-5 entry and disassembly followed in living cells by FRET, fluorescence anisotropy, and FLIM. AB - We have used fluorescence resonance energy transfer (FRET) to follow the process of capsid disassembly for adenovirus (Ad) serotype 5 (Ad5) in living CHO-CAR cells. Ad5 were weakly labeled on their capsid proteins with FRET donor and acceptor fluorophores. A progressive decrease in FRET efficiency recorded during Ad5 uptake revealed that the time course of Ad5 capsid disassembly has two sequential protein dissociation rates with half-times of 3 and 60 min. Fluorescence anisotropy measurements of the segmental motions of fluorophores on Ad5 indicate that the first rate is linked to the detachment from the capsid of the protruding, flexible fiber proteins. The second rate was shown to report on the combined dissociation of protein IX, penton base, and hexons, which form the rigid icosahedral capsid shell. Fluorescence lifetime imaging microscopy measurements using a pH-sensitive probe provided information on the pH of the microenvironment of Ad5 particles during intracellular trafficking, and confirmed that the fast fiber dissociation step occurred at the onset of endocytosis. The slower dissociation phase was shown to coincide with the escape of Ad5 from endocytic compartments into the cytosol, and its arrival at the nuclear membrane. These results demonstrate a rapid, quantitative live-cell assay for the investigation of virus-cell interactions and capsid disassembly. PMID- 15298935 TI - Single-molecule three-color FRET. AB - Fluorescence resonance energy transfer (FRET) measured at the single-molecule level can reveal conformational changes of biomolecules and intermolecular interactions in physiologically relevant conditions. Thus far single-molecule FRET has been measured only between two fluorophores. However, for many complex systems, the ability to observe changes in more than one distance is desired and FRET measured between three spectrally distinct fluorophores can provide a more complete picture. We have extended the single-molecule FRET technique to three colors, using the DNA four-way (Holliday) junction as a model system that undergoes two-state conformational fluctuations. By labeling three arms of the junction with Cy3 (donor), Cy5 (acceptor 1), and Cy5.5 (acceptor 2), distance changes between the donor and acceptor 1, and between the donor and acceptor 2, can be measured simultaneously. Thus we are able to show that the acceptor 1 arm moves away from the donor arm at the same time as the acceptor 2 arm approaches the donor arm, and vice versa, marking the first example of observing correlated movements of two different segments of a single molecule. Our data further suggest that Holliday junction does not spend measurable time with any of the helices unstacked, and that the parallel conformations are not populated to a detectable degree. PMID- 15298936 TI - Microviscoelasticity of the apical cell surface of human umbilical vein endothelial cells (HUVEC) within confluent monolayers. AB - We studied the local viscoelasticity of the apical membrane of human umbilical vein endothelial cells within confluent layers by magnetic tweezers microrheometry. Magnetic beads are coupled to various integrins by coating with fibronectin or invasin. By analyzing the deflection of beads evoked by various force scenarios we demonstrate that the cell envelope behaves as a linear viscoelastic body if forces up to 2 nN are applied for short times (<20 s) but can respond in an adaptive way if stress pulses are applied longer (>30 s). The time-dependent shear relaxation modulus G(t) exhibits three time regimes: a fast response (t < 0.05 s) where the relaxation modulus G(t) obeys a power law G(t) approximately t(-0.82+/-0.02); a plateau-like behavior (at 0.05 s < t < 0.15 s); and a slow flow-like response which is, however, partially reversible. Strain field mapping experiments with colloidal probes show that local forces induce a strain field exhibiting a range of zeta = 10 +/- 1 microm, but which could only be observed if nonmagnetic beads were coupled to the cell surface by invasin. By application of the theory of elasticity of planar bodies we estimated a surface shear modulus of 2.5 x10(-4) N/m. By assuming a thickness of the actin cortex of approximately 0.5 microm we estimate a Young modulus micro approximately 400 Pa for the apical membrane. The value agrees with a plateau modulus of an entangled or weakly cross-linked actin network of an actin concentration of 100 microM (mesh size 0.2 microm). This result together with our observation of a strong reduction of the shear modulus by the actin destabilizing agent latrunculin A suggests that the shear modulus measured by our technique is determined by the actin cortex. The effect of two ligands inducing actin stress fiber formation and centripetal contraction of cells (associated with the formation of gaps in the confluent cell monolayer) on the viscoelastic responses were studied: histamine and lysophosphatidic acid (LPA). Histamine evoked a dramatic increase of the cell stiffness by >1 order of magnitude within <30 s, which is attributed to a transient rise of the intracellular Ca(2+) level, since DMSO exerted a similar effect. The stiffening is accompanied by a concomitant rounding of the cells as observed by microinterferometry and relaxes partially in the timescale of 5 min, whereas gaps between cells close after approximately 30 min. LPA did not exert a remarkable and reproducible effect other than an occasional very weak transient increase of the shear stiffness, which shows that the gap formation activated by LPA is mediated by a different mechanism than that induced by histamine. PMID- 15298937 TI - Imaging neuronal seal resistance on silicon chip using fluorescent voltage sensitive dye. AB - The electrical sheet resistance between living cells grown on planar electronic contacts of semiconductors or metals is a crucial parameter for bioelectronic devices. It determines the strength of electrical signal transduction from cells to chips and from chips to cells. We measured the sheet resistance by applying AC voltage to oxidized silicon chips and by imaging the voltage change across the attached cell membrane with a fluorescent voltage-sensitive dye. The phase map of voltage change was fitted with a planar core-coat conductor model using the sheet resistance as a free parameter. For nerve cells from rat brain on polylysine as well as for HEK293 cells and MDCK cells on fibronectin we find a similar sheet resistance of 10 MOmega. Taking into account the independently measured distance of 50 nm between chip and membrane for these cells, we obtain a specific resistance of 50 Omegacm that is indistinguishable from bulk electrolyte. On the other hand, the sheet resistance for erythrocytes on polylysine is far higher, at approximately 1.5 GOmega. Considering the distance of 10 nm, the specific resistance in the narrow cleft is enhanced to 1500 Omegacm. We find this novel optical method to be a convenient tool to optimize the interface between cells and chips for bioelectronic devices. PMID- 15298938 TI - Na/K pump-induced [Na](i) gradients in rat ventricular myocytes measured with two photon microscopy. AB - Via the Na/Ca and Na/H exchange, intracellular Na concentration ([Na](i)) is important in regulating cardiac Ca and contractility. Functional data suggest that [Na](i) might be heterogeneous in myocytes that are not in steady state, but little direct spatial information is available. Here we used two-photon microscopy of SBFI to spatially resolve [Na](i) in rat ventricular myocytes. In vivo calibration yielded an apparent K(d) of 27 +/- 2 mM Na. Similar resting [Na](i) was found using two-photon or single-photon ratiometric measurements with SBFI (10.8 +/- 0.7 vs. 11.1 +/- 0.7 mM). To assess longitudinal [Na](i) gradients, Na/K pumps were blocked at one end of the myocyte (locally pipette applied K-free extracellular solution) and active in the rest of the cell. This led to a marked increase in [Na](i) at sites downstream of the pipette (where Na enters the myocyte and Na/K pumps are blocked). [Na](i) rise was smaller at upstream sites. This resulted in sustained [Na](i) gradients (up to approximately 17 mM/120 microm cell length). This implies that Na diffusion in cardiac myocytes is slow with respect to trans-sarcolemmal Na transport rates, although the mechanisms responsible are unclear. A simple diffusion model indicated that such gradients require a Na diffusion coefficient of 10-12 microm(2)/s, significantly lower than in aqueous solutions. PMID- 15298939 TI - How does protein architecture facilitate the transduction of ATP chemical-bond energy into mechanical work? The cases of nitrogenase and ATP binding-cassette proteins. AB - Transduction of adenosine triphosphate (ATP) chemical-bond energy into work to drive large-scale conformational changes is common in proteins. Two specific examples of ATP-utilizing proteins are the nitrogenase iron protein and the ATP binding-cassette transporter protein, BtuCD. Nitrogenase catalyzes biological nitrogen fixation whereas BtuCD transports vitamin B(12) across membranes. Both proteins drive their reactions with ATP. To interpret how the mechanical force generated by ATP binding and hydrolysis is propagated in these proteins, a coarse grained elastic network model is employed. The analysis shows that subunits of the proteins move against each other in a concerted manner. The lowest-frequency modes of the nitrogenase iron protein and of the ATP binding-cassette transporter BtuCD protein are found to link the functionally critical domains, and these modes are suggested to be responsible for (at least the initial stages) large scale ATP-coupled conformational changes. PMID- 15298940 TI - Impedance analysis of the organ of corti with magnetically actuated probes. AB - An innovative method is presented to measure the mechanical driving point impedance of biological structures up to at least 40 kHz. The technique employs an atomic force cantilever with a ferromagnetic coating and an external magnetic field to apply a calibrated force to the cantilever. Measurement of the resulting cantilever velocity using a laser Doppler vibrometer yields the impedance. A key feature of the method is that it permits measurements for biological tissue in physiological solutions. The method was applied to measure the point impedance of the organ of Corti in situ, to elucidate the biophysical basis of cochlear amplification. The basilar membrane was mechanically clamped at its tympanic surface and the measurements conducted at different radial positions on the reticular lamina. The tectorial membrane was removed. The impedance was described by a generalized Voigt-Kelvin viscoelastic model, in which the stiffness was real valued and independent of frequency, but the viscosity was complex-valued with positive real part, which was dependent on frequency and negative imaginary part, which was independent of frequency. There was no evidence for an inertial component. The magnitude of the impedance was greatest at the tunnel of Corti, and decreased monotonically in each of the radial directions. In the absence of inertia, the mechanical load on the outer hair cells causes their electromotile displacement responses to be reduced by only 10-fold over the entire range of auditory frequencies. PMID- 15298941 TI - Single molecule elasticity measurements: a biophysical approach to bacterial nucleoid organization. PMID- 15298942 TI - Complementarities and convergence of results in bacteriorhodopsin trimer simulations. PMID- 15298944 TI - Fiber from fruit and colorectal neoplasia. PMID- 15298945 TI - XPD polymorphism and risk of subsequent cancer in individuals with nonmelanoma skin cancer. AB - BACKGROUND: Individuals with nonmelanoma skin cancer (NMSC) are at increased risk of developing subsequent cancers. Genetic predisposition to reduced DNA repair capacity may be an underlying susceptibility factor explaining the excess risk of malignancies. To test this hypothesis, a cohort study was conducted to examine the association between XPD Lys751Gln polymorphism and risk of a second primary cancer in individuals with NMSC. METHODS: A subgroup of 481 individuals with a history of NMSC who participated in the CLUE II community-based cohort was followed for the development of a second primary cancer. Blood specimens donated in 1989 were genotyped for the XPD Lys751Gln polymorphism using the 5' nuclease assay. Cox proportional regression with delayed entry was used to calculate the incidence rate ratio (IRR) and 95% confidence interval (95% CI) for risk of developing a second primary cancer according to XPD genotype. All statistical tests were two sided. RESULTS: Eighty individuals developed a second primary cancer. The most frequent occurring cancers were of the prostate (18%), lung (15%), and breast (15%). Persons with at least one Gln allele had an increased risk of a second primary cancer compared with the reference Lys/Lys genotype (adjusted IRR 2.22, 95% CI 1.30-3.76). When the reference category was limited to never smokers with the Lys/Lys genotype, the risk of developing a second primary cancer associated with having at least one Gln allele was increased >3-fold in both never smokers (IRR 3.93, 95% CI 1.36-11.36) and ever smokers (IRR 6.14, 95% CI 2.17-17.37). CONCLUSION: These findings suggest that individuals with NMSC who have the variant XPD Gln allele are at increased risk of developing a second primary cancer. PMID- 15298946 TI - Serum carotenoids and alpha-tocopherol and risk of nonmelanoma skin cancer. AB - BACKGROUND: Carotenoids and tocopherols have been hypothesized to protect against cancer. METHODS: We prospectively evaluated associations of several carotenoids and alpha-tocopherol with risk of nonmelanoma skin cancer using serum collected at baseline from 302 subjects in the Isotretinoin-Basal Cell Carcinoma Prevention Trial. All subjects had at least two BCCs in the 5 years prior to randomization. During 5 years of follow-up, 70 subjects did not develop a nonmelanoma skin cancer, 221 developed a BCC, and 85 developed a squamous cell carcinoma (SCC). Cox proportional hazards models were used to estimate risk ratios. Models were stratified by clinical center and gender and adjusted for age, solar damage, skin type, number of prior BCCs and/or SCCs, treatment group, body mass index, and serum low-density lipoprotein-cholesterol and high-density lipoprotein cholesterol. RESULTS: Risk of developing a subsequent BCC was not related to serum levels of any of the carotenoids measured or to alpha-tocopherol. Serum levels of alpha-carotene, beta-carotene, lycopene, and alpha-tocopherol also were not independently related to risk of a subsequent SCC. However, serum lutein, zeaxanthin, and beta-cryptoxanthin were positively related to SCC risk; risk ratios for subjects in the highest versus lowest tertiles of these micronutrients were 1.63 [95% confidence interval (95% CI) 0.88-3.01; P for trend = 0.01], 2.40 (95% CI 1.30-4.42; P for trend = 0.01), and 2.15 (95% CI 1.21-3.83; P for trend = 0.09), respectively. CONCLUSION: Additional research is needed on the relationship of carotenoids to SCC risk in the general population and in subsets of the population who are at increased risk. PMID- 15298947 TI - Carbohydrates and the risk of breast cancer among Mexican women. AB - OBJECTIVE: High carbohydrate intake has been hypothesized to be a risk factor for breast cancer, possibly mediated by elevated levels of free insulin, estrogens, and insulin-like growth factor-1. Therefore, we conducted a population-based case control study among a Mexican population characterized by relatively low fat and high carbohydrate intakes. METHODS: Women ages 20 to 75 years, identified through six hospitals in Mexico City (n = 475), were interviewed to obtain data relating to diet (using a food frequency questionnaire) and breast cancer risk factors. Controls (n = 1,391) were selected from the Mexico City population using a national sampling frame. RESULTS: Carbohydrate intake was positively associated with breast cancer risk. Compared with women in the lowest quartile of total carbohydrate intake, the relative risk of breast cancer for women in the highest quartile was 2.22 [95% confidence interval (95% CI) 1.63-3.04], adjusting for total energy and potential confounding variables (P for trend < 0.0001). This association was present in premenopausal and postmenopausal women (for highest versus lowest quartile, odds ratio 2.31, 95% CI 1.36-3.91 in premenopausal women and odds ratio 2.22, 95% CI 1.49-3.30 in postmenopausal women). Among carbohydrate components, the strongest associations were observed for sucrose and fructose. No association was observed with total fat intake. DISCUSSION: In this population, a high percentage of calories from carbohydrate, but not from fat, was associated with increased breast cancer risk. This relation deserves to be investigated further, particularly in populations highly susceptible to insulin resistance. PMID- 15298948 TI - Genetic polymorphisms in the IGFBP3 gene: association with breast cancer risk and blood IGFBP-3 protein levels among Chinese women. AB - Cumulative evidence suggests that insulin-like growth factors (IGF) play an important role in the etiology of breast cancer. The IGF binding proteins regulate the action of IGFs, and >90% of circulating IGFs are bound to IGFBP-3. We evaluated the associations of five (A-202C, G227C, C3804G, 5606InsA, and C5827T) genetic polymorphisms in the IGFBP3 gene with breast cancer risk and the blood IGFBP-3 protein level in a population-based, case-control study conducted among Chinese women in Shanghai. Genomic DNA samples from 1,193 incident breast cancer patients and 1,310 community controls were genotyped for IGFBP3 polymorphisms. Blood IGFBP-3 levels were determined for 390 controls. A 30% to 60% elevated risk of breast cancer was found to be associated with homozygosity for the variant allele in polymorphisms A-202C, G227C, 5606InsA, and C5827T. Carrying the variant allele in C3804G was also associated with an increased risk. About 13.5% of cases and 9.7% of controls had one or more of the above risk genotypes, resulting in odds ratio [OR; 95% confidence interval (95% CI)] of 1.4 (1.0-1.9). The ORs (95% CIs) were 1.3 (1.0-1.8) and 1.7 (1.1-2.5) for women with one to two and three to five risk genotypes, respectively (P for trend < 0.01). Four common haplotypes for the IGFBP3 gene were identified. Compared with the haplotype containing only the wild-type allele in the five loci, the haplotype with the variant allele in all sites was associated with an elevated risk of breast cancer (OR 1.4, 95% CI 1.0-1.9), particularly among younger women (OR 2.3, 95% CI 1.3-3.9). With the exception of C3804G, in which no homozygote was identified, the level of circulating IGFBP-3 was reduced in a dose-response manner with an increasing number of variant alleles in each of the other four polymorphic sites (P for trend < 0.05). These results indicated that IGFBP3 polymorphisms may be associated with the level of blood IGFBP-3 protein and an increased risk of breast cancer. PMID- 15298949 TI - Associations between reproductive and menstrual factors and postmenopausal sex hormone concentrations. AB - Reproductive and menstrual characteristics, as well as high circulating estrogen concentrations, are associated with risk of hormone-related cancers in postmenopausal women. To explore possible etiologic relationships between menstrual/reproductive characteristics and risk of hormone-related cancers, we examined associations between menstrual/reproductive factors and serum concentrations of free estradiol, total estradiol, estrone, sex hormone binding globulin (SHBG), and follicle stimulating hormone (FSH). This study was conducted in 173 postmenopausal women using data from the prerandomization visit of an exercise clinical trial. Participants were sedentary, overweight/obese, and not on hormone therapy. Women > or =20 years past menopause had 23% lower total estradiol and 30% lower free estradiol concentrations than women within 4 years of menopause (P for trend = 0.04 and 0.02, respectively). Nulliparous women had 19% higher FSH concentrations than parous women (P = 0.02). Among parous women, parity was positively associated with SHBG and negatively associated with free estradiol concentrations. Women with > or =4 children had 20% lower free estradiol and 38% higher SHBG concentrations compared with women with one birth (P for trend = 0.02 and 0.01, respectively). Total number of months spent breast feeding was modestly and inversely associated with serum FSH concentrations (P for trend = 0.07). Our results suggest that menstrual/reproductive characteristics may be associated with postmenopausal hormone concentrations; verification of these results in other studies may elucidate how these variables influence risk of hormone-related cancers. PMID- 15298950 TI - Use of oral contraceptives, alcohol, and risk for invasive breast cancer. AB - The aim of our study was to examine how the use of oral contraceptives (OCs) interact with alcohol on breast cancer risk within the large prospective follow up study, Norwegian Women and Cancer Study. Between 1991 and 1997, women aged 30 to 70 years were drawn at random from the central person register and mailed an invitation. Follow-up information was collected throughout 2001 by linkage to national registries. Only women (n = 86,948) with complete information on alcohol consumption and duration of OC use were included in the present analysis. A total of 1,130 invasive breast cancers were diagnosed during 618,638 person-years of follow-up. Consumption of > or =10.0 g/d alcohol was associated with a breast cancer relative risk (95% confidence interval) of 1.69 (1.32-2.15), consistent with a linear relationship (P for trend < 0.0001). Among alcohol consumers, an excess risk of breast cancer was observed for total duration of OC use only among women who consumed <5 g/d alcohol (P for trend = 0.0009). We observed a negative interaction between duration of OC use and alcohol consumption effects (P for interaction = 0.01). After stratification on menopausal status, the association between high alcohol intake and breast cancer was more prominent among postmenopausal women than among premenopausal women (P for heterogeneity = 0.01). No interaction between alcohol and duration of OC use were significant after stratification on menopausal status. Our findings in conjunction with biological data imply that alcohol and OCs have antagonistic effects on breast cancer risk through a common pathway. Whether the interactive effect differs according to menopausal status remains unclear and needs further investigations. PMID- 15298951 TI - Oral contraceptive use and breast cancer risk: modification by NAD(P)H:quinone oxoreductase (NQO1) genetic polymorphisms. AB - Despite intensive study, the relationship between oral contraception (OC) and breast cancer remains unclear. OCs contain a potent synthetic estrogen (ethinyl estradiol) but lower endogenous estradiol levels, and ethinyl estradiol is a weak progenitor of semiquinones, catechol estrogens capable of damaging DNA. NAD(P)H:quinone oxoreductase (NQO1) stabilizes semiquinones, thus potentially preventing genetic damage from catechol estrogens, and the NQO1 C609T polymorphism seems functionally relevant. Using data from the Shanghai Breast Cancer Study, a population-based case-control study, we investigated the relationships between OC use (20% ever using), breast cancer, and NQO1 (C/C 31% and C/T + T/T 69%) among 1,039 cases and 1,121 controls. Breast cancer was not significantly associated with NQO1 genotype. There was a significant protective association between OC after age 30 years and premenopausal breast cancer [odds ratio (OR) 0.51, 95% confidence interval (95% CI) 0.29-0.89] primarily with the NQO1 T allele (C/C OR 0.76, 95% CI 0.31-1.82; C/T + T/T OR 0.38, 95% CI 0.18 0.80; P for interaction = 0.19). The association between premenopausal breast cancer and OCs significantly differed with NQO1 genotype when using OCs for >18 months (C/C OR 2.34, 95% CI 0.92-5.99; C/T + T/T OR 0.69, 95% CI 0.38-1.25; P for interaction = 0.02). Among women with the C/C genotype, postmenopausal breast cancer was significantly associated with ever-using OCs (C/C OR 2.01, 95% CI 1.08 3.74; C/T + T/T OR 0.72, 95% CI 0.49-1.05; P for interaction < 0.01). This crossover was stronger with OC use prior to age 30 years (C/C OR 3.00, 95% CI 1.43-6.25; C/T or T/T OR 0.49, 95% CI 0.29-0.81; P for interaction < 0.01). Our results require confirmation but suggest that the OC and breast cancer association depends on the ability to invoke protection from catechol estrogens. PMID- 15298952 TI - Testicular, other genital, and breast cancers in first-degree relatives of testicular cancer patients and controls. AB - Previous studies showed an increased prevalence of testicular cancer among fathers and brothers of testicular cancer patients. We examined whether testicular, other genital, and breast cancers aggregate in parents and siblings of testicular cancer patients in a population-based case-control study, including males, ages 15 to 69 years at diagnosis, with primary malignant tumors of the testes or extragonadal germ cell tumors. Controls were ascertained through the mandatory registries of residents and frequency matched to the cases by age and region of residence. In a face-to-face interview, 269 cases and 797 controls provided health-related information on parents and siblings. We calculated odds ratios (OR) and corresponding 95% confidence intervals (95% CI) based on the generalized estimating equations technique, adjusting for the matching variables and relatives' age. Three (1.1%) fathers and eight (3.2%) brothers of cases were affected with testicular cancer compared with four (0.5%) fathers and two (0.2%) brothers of controls. The OR (95% CI) of familial testicular cancer was 6.6 (2.35 18.77). Only nonseminoma patients had fathers with testicular cancer, whereas the affected brothers were all related to seminoma patients. Overall, we found an increased risk for genital other than testicular cancers (OR 2.5, 95% CI 1.43 4.43). For breast cancer, we detected an increased risk in sisters (OR 9.5, 95% CI 2.01-45.16, adjusted for age of study participant and age of sister) but not in mothers. Our findings support the hypothesis that testicular and other genital cancers have a common familial component that may be due to genetic and shared exogenous factors such as estrogen exposure during fetal development. PMID- 15298953 TI - Risk of early-onset prostate cancer in relation to germ line polymorphisms of the vitamin D receptor. AB - Vitamin D inhibits prostate cancer cell growth, angiogenesis, and metastasis. These actions are mediated by the vitamin D receptor. We examined associations between prostate cancer risk and five polymorphisms in the VDR gene: four single nucleotide polymorphisms (FokI, BsmI, ApaI, and TaqI restriction sites) and the polyadenylic acid microsatellite. Specifically, we genotyped population-based samples of young African Americans (113 cases and 121 controls) and Whites (232 cases and 171 controls) and members of 98 predominantly White families with multiple cases of prostate cancer. Among Whites, there was no evidence for association between prostate cancer risk and alleles at any of the five polymorphic sites regardless of how the men were ascertained. Moreover, estimated five-locus haplotype frequencies were similar in White cases and controls. Among African Americans, prostate cancer risk was associated with homozygosity for the F allele at the FokI site (odds ratio 1.9, 95% confidence interval 1.0-3.3). In addition, estimated haplotype frequencies differed significantly (P < 0.01) between African American cases and controls. These findings need replication in other studies of African Americans. Homozygosity for the F allele at the FokI site is more prevalent in the African American population than in U.S. Whites. If the FokI association noted here were causal, this difference could account for some of the disease burden among African Americans and some of the excess risk in African Americans compared with Whites. PMID- 15298954 TI - Relationship between methylenetetrahydrofolate reductase C677T and A1298C genotypes and haplotypes and prostate cancer risk and aggressiveness. AB - Previous reports indicate that polymorphisms in the MTHFR gene play a role in cancer development, but their potential impact on prostate cancer has not been well studied. Here, we evaluate the association between two MTHFR polymorphisms, C677T and A1298C, and prostate cancer risk and aggressiveness in a moderately large family-based case-control study (439 cases and 479 sibling controls). Among all study subjects, we observed no association between the C677T variant and prostate cancer but a slight positive association between the A1298C variant and risk of this disease [odds ratio (OR) 1.41, 95% confidence interval (CI) 0.96 2.06; P = 0.08]. When stratifying the study population by disease aggressiveness at diagnosis, the C677T variant was positively associated with risk among men with less advanced disease (OR 1.86, 95% CI 1.00-3.46; P = 0.05). In contrast, when looking at men with more advanced disease, the C677T variant was inversely associated with risk (OR 0.51, 95% CI 0.32-0.82; P = 0.01), whereas the A1298C variant was positively associated with risk (OR 1.79, 95% CI 1.06-3.02; P = 0.03). Furthermore, the 677T-1298A haplotype was positively associated with prostate cancer among men with less advanced disease (OR 1.84, 95% CI 1.07-3.16; P = 0.03) and inversely associated with risk of more advanced disease (OR 0.47, 95% CI 0.29-0.76; P = 0.002). Our findings suggest that 677T and 1298A, or another variant on their haplotype, may be associated with a reduced risk of progression to more advanced prostate cancer. PMID- 15298955 TI - A population-based case-control study of the XRCC1 Arg399Gln polymorphism and susceptibility to bladder cancer. AB - Cigarette smoking is the major cause of bladder cancer. Constituents in tobacco smoke can induce oxidative DNA damage requiring base excision repair. The Arg399Gln polymorphism in the DNA base excision repair gene XRCC1 is associated with several phenotypic markers of reduced DNA repair capacity. Results from several epidemiologic studies suggest that the Arg399Gln polymorphism may influence susceptibility to several cancers including bladder cancer; however, data from large population-based studies are lacking. In a population-based case control study from New Hampshire, we observed a reduced risk among those homozygous for the Arg399Gln XRCC1 variant polymorphism compared with those with one or two wild-type alleles (odds ratio 0.6, 95% confidence interval 0.4-1.0). There was no indication of a gene-environment interaction between cigarette smoking and the variant genotype. Our data are consistent with a potential role of the XRCC1 Arg399Gln polymorphism in bladder cancer susceptibility and further suggest that there may be DNA lesions important in bladder carcinogenesis, repaired by the base excision repair mechanism, that are not directly associated with tobacco smoking. PMID- 15298956 TI - Benzo(a)pyrene diolepoxide (BPDE)-DNA adduct levels in leukocytes of smokers in relation to polymorphism of CYP1A1, GSTM1, GSTP1, GSTT1, and mEH. AB - Benzo(a)pyrene [B(a)P] diolepoxide (BPDE)-DNA adducts were measured in the leukocytes of 41 healthy smokers using high-performance liquid chromatography coupled with a fluorimetric detector. The correlation between exposure to B(a)P through smoking and BPDE-DNA adduct levels was poor (r = 0.31), although subjects in the high exposure group [B(a)P > 50 ng/d] had a slightly higher level of adducts compared with the less exposed group (mean +/- SE, 1.70 +/- 0.3 versus 1.09 +/- 0.1; P = 0.057). We studied the effect on BPDE-DNA adducts of individual variations in genes controlling B(a)P metabolism, classifying subjects in "low risk" and "high-risk" genotypes for smoking-related B(a)P DNA damage. The high risk group included subjects characterized by a combination of increased B(a)P activation [cytochrome P450 1A1 (CYP1A1) MspI and/or exon 7 Ile462Val allele variants and microsomal epoxide hydrolase (mEH) fast activity] and decreased deactivation ability [presence of glutathione S-transferase M1 (GSTM1) null allele and wild-type glutathione S-transferase P1 (GSTP1)]. The low-risk group included smokers with lower B(a)P activation (wild-type CYP1A1, low or intermediate mEH activity) and higher deactivation capacity (active GSTM1, GSTP1 Ile105Val allele). Subjects in the low-risk group had lower levels of BPDE-DNA adducts compared with subjects in the high-risk genotype group; this difference was significant using two markers (CYP1A1 and GSTM1, median +/- SD, 0.77 +/- 1.16 versus 1.89 +/- 0.39; P = 0.03) or three markers (CYP1A1, GSTM1, and GSTP1, median +/- SD, 0.66 +/- 0.93 versus 1.43 +/- 1.17; P = 0.013). The discrimination between groups was reduced when including mEH as an additional marker (P = 0.085). In conclusion, CYP1A1, GSTM1, and GSTP1 genotyping seems to be a risk predictor of BPDE-DNA adduct formation in leukocytes. PMID- 15298957 TI - Racial differences in enrolment in a cancer genetics registry. AB - BACKGROUND: Lower enrolment of minorities into research studies has been reported frequently. Most studies have little information about nonparticipants, making it difficult to identify characteristics associated with enrolment and how they might vary by race. METHODS: Women who had previously participated in a population-based, case-control study of breast cancer in North Carolina were invited to enroll in a cancer genetics registry. Detailed questionnaire data on sociodemographic characteristics and cancer risk factors were available for all women. We compared characteristics of women who agreed to be in the registry with those who were deceased, were unlocatable, or declined enrolment. Unconditional logistic regression analyses were done to identify predictors of enrolment. RESULTS: Enrolment rates were markedly lower among African Americans than Whites (15% and 36%, respectively) due to both lower contact rates (41% versus 63%) and lower enrolment rates among those contacted (37% versus 58%). Logistic regression models suggested that racial differences in enrolment were not due to socioeconomic characteristics or other cancer risk factors; race was the only significant predictor of enrolment in multivariable models (odds ratio 0.41, 95% confidence interval 0.23-0.72). CONCLUSIONS: Although all women had previously taken part in a research study, African American women were less likely to enroll in the cancer genetics registry than White women. A possible explanation of these findings is that studies of genetics may present particular concerns for African Americans. Further research is needed to identify attitudes and issues that present barriers to participation among minorities. PMID- 15298958 TI - Validation of p16INK4a as a marker of oncogenic human papillomavirus infection in cervical biopsies from a population-based cohort in Costa Rica. AB - Due to the high prevalence of cancer-associated types of human papillomavirus (HPV) and the poorly reproducible histologic classification of low-grade lesions, identifying infected women at highest risk for cancer prior to neoplastic progression remains a challenge. We therefore explored the utility of p16INK4a immunostaining as a potential diagnostic and prognostic biomarker for cervical neoplasia using paraffin-embedded tissue blocks (punch biopsies and loop electrosurgical excision procedures) obtained from women referred to colposcopy during the enrollment phase of the Guanacaste Project (1993 to 1994). All blocks from 292 women selected by HPV status (HPV negative, nononcogenic HPV positive, or oncogenic HPV positive) and representing the diagnostic spectrum of the population [normal to precancer: cervical intraepithelial neoplasia (CIN) 3] were immunostained for p16INK4a using the p16INK4a research kit based on the monoclonal antibody clone E6H4 (MTM Laboratories, Heidelberg, Germany). For CIN3, the sensitivity of diffuse p16INK4a immunostaining was 100% and the specificity was 95%. For CIN2, the sensitivity and specificity for diffuse staining were 81.1% and 95.4%, respectively. Generalized to the 10,000-woman cohort, this translated to positive predictive value and negative predictive value of 13.9% and 100% for CIN3, respectively, and 20.4% and 99.7% for CIN2 or CIN3, respectively. Of women with an initial diagnosis of less than CIN2 for whom follow-up data for up to 5 to 7 years were available, 44% with diffuse staining developed persistent infection (CIN2 or CIN3). Whereas our data support the diagnostic potential for p16INK4a, further prospective studies with detailed follow-up determining the prognostic capacity of this marker are needed. PMID- 15298959 TI - Childhood social environment and Hodgkin's lymphoma: new findings from a population-based case-control study. AB - BACKGROUND: Risk of Hodgkin's lymphoma in young adults has previously been associated with higher childhood socioeconomic status (SES) and other markers of delayed infection with common childhood pathogens, especially the Epstein-Barr virus. This study examines the current role of childhood social environment in the development of Hodgkin's lymphoma. METHODS: A population-based case-control study of 565 Hodgkin's lymphoma cases and 679 controls was conducted in the Boston, MA metropolitan area and the state of Connecticut to investigate the viral etiology of Hodgkin's lymphoma. RESULTS: A novel association was detected between attendance of nursery school or day care and reduced risk of Hodgkin's lymphoma among individuals ages 15 to 54 years. The odds ratio (95% confidence interval) for having attended preschool for at least 1 year was 0.64 (0.45-0.92). Risk of young-adult Hodgkin's lymphoma was also associated with family history of hematopoietic cancer, Jewish ethnicity, and cigarette smoking. Other indicators of childhood SES were not associated with young-adult Hodgkin's lymphoma. Among older adults ages 55 to 79 years, Hodgkin's lymphoma was associated with lower childhood SES but not with preschool attendance. CONCLUSIONS: Early exposure to other children at nursery school and day care seems to decrease the risk of Hodgkin's lymphoma in young adults, most likely by facilitating childhood exposure to common infections and promoting maturation of cellular immunity. This finding supports the delayed infection model of Hodgkin's lymphoma etiology in young adults while introducing a new major determinant of age at infection. Hodgkin's lymphoma seems to have a separate pathogenesis among older adults. PMID- 15298960 TI - Cytochrome P450 1A1 polymorphism and childhood leukemia: an analysis of matched pairs case-control genotype data. AB - The association between the genotypic frequencies of the cytochrome P450 1A1 polymorphism and the risk of childhood leukemia is explored with the data from a matched case-control study. The data are displayed in a 3 x 3 case-control array, and the discordant pair counts are assessed for quasi-independence, homogeneity, and symmetry. This statistical approach is contrasted to the more typical analysis of matched data based on a conditional logistic model and estimated odds ratios. The statistical analysis of 175 matched pairs (part of a large study of potential environmental/genetic influences on the risk of childhood leukemia) showed no evidence of an association between cytochrome P450 1A1 genotype frequencies and case-control status. PMID- 15298961 TI - Cancer prevention strategies that address the evolutionary dynamics of neoplastic cells: simulating benign cell boosters and selection for chemosensitivity. AB - Cells in neoplasms evolve by natural selection. Traditional cytotoxic chemotherapies add further selection pressure to the evolution of neoplastic cells, thereby selecting for cells resistant to the therapies. An alternative proposal is a benign cell booster. Rather than trying to kill the highly dysplastic or malignant cells directly, a benign cell booster increases the fitness of the more benign cells, which may be either normal or benign clones, so that they may outcompete more advanced or malignant cells in a neoplasm. In silico simulations of benign cell boosters in neoplasms with evolving clones show benign cell boosters to be effective at destroying advanced or malignant cells and preventing relapse even when applied late in progression. These results are conditional on the benign cell boosters giving a competitive advantage to the benign cells in the neoplasm. Furthermore, the benign cell boosters must be applied over a long period of time in order for the benign cells to drive the dysplastic cells to extinction or near extinction. Most importantly, benign cell boosters based on this strategy must target a characteristic of the benign cells that is causally related to the benign state to avoid relapse. Another promising strategy is to boost cells that are sensitive to a cytotoxin, thereby selecting for chemosensitive cells, and then apply the toxin. Effective therapeutic and prevention strategies will have to alter the competitive dynamics of a neoplasm to counter progression toward invasion, metastasis, and death. PMID- 15298962 TI - Serologic assessment of type 1 and type 2 immunity in healthy Japanese adults. AB - We assessed the informativeness of several serologic biomarkers of immune function using serum specimens collected in the Miyazaki Cohort Study from subjects who were seronegative for anti-human T-cell lymphotrophic virus I and anti-hepatitis C virus. To broadly characterize type 1 immune status, we measured EBV antibody titers, because titer profiles associated with cellular immune suppression are well described. We also tested for three type 2 biomarkers: total serum IgE, soluble CD23, and soluble CD30. Nonreactivity to a tuberculin purified protein derivative (PPD) skin test is indicative of diminished delayed-type hypersensitivity (type 1) responsiveness in the study population due to a history of tuberculosis exposure or Bacillus Calmette-Guerin vaccination. We therefore evaluated the serologic markers as predictors of PPD nonreactivity using logistic regression. Subjects whose EBV antibody profiles were consistent with deficient type 1 immunity were more than thrice as likely to be PPD nonreactive as persons with "normal" antibody titers. Elevated total IgE was also strongly associated with PPD nonreactivity (odds ratio 3.4, 95% confidence interval 1.2-9.9); elevated soluble CD23 had a weaker, but positive, odds ratio, whereas soluble CD30 levels were not predictive of PPD status. Therefore, PPD nonreactivity is associated, in this population, with a pattern of serum biomarkers that is indicative of diminished type 1 and elevated type 2 immunity. We conclude that, with the exception of soluble CD30, the serologic markers are informative for the characterization of type 1/type 2 immune status using archived sera from study populations of healthy adults. PMID- 15298963 TI - Diosgenin, a steroid saponin of Trigonella foenum graecum (Fenugreek), inhibits azoxymethane-induced aberrant crypt foci formation in F344 rats and induces apoptosis in HT-29 human colon cancer cells. AB - Trigonella foenum graecum (fenugreek) is traditionally used to treat disorders such as diabetes, high cholesterol, wounds, inflammation, and gastrointestinal ailments. Recent studies suggest that fenugreek and its active constituents may possess anticarcinogenic potential. We evaluated the preventive efficacy of dietary fenugreek seed and its major steroidal saponin constituent, diosgenin, on azoxymethane-induced rat colon carcinogenesis during initiation and promotion stages. Preneoplastic colonic lesions or aberrant crypt foci (ACF) were chosen as end points. In addition, we assessed the mechanism of tumor growth inhibition of diosgenin in HT-29 human colon cancer cells. To evaluate the effect of the test agent during the initiation and postinitiation stages, 7-week-old male F344 rats were fed experimental diets containing 0% or 1% fenugreek seed powder (FSP) or 0.05% or 0.1% diosgenin for 1 week and were injected with azoxymethane (15 mg/kg body weight). Effects during the promotional stage were studied by feeding 1% FSP or 0.1% diosgenin 4 weeks after the azoxymethane injections. Rats were sacrificed 8 weeks after azoxymethane injection, and their colons were evaluated for ACF. We found that, by comparison with control, continuous feeding of 1% FSP and 0.05% and 0.1% diosgenin suppressed total colonic ACF up to 32%, 24%, and 42%, respectively (P < or = 0.001 to 0.0001). Dietary FSP at 1% and diosgenin at 0.1% fed only during the promotional stage also inhibited total ACF up to 33% (P < or = 0.001) and 39% (P < or = 0.0001), respectively. Importantly, continuous feeding of 1% FSP or 0.05% or 0.1% diosgenin reduced the number of multicrypt foci by 38%, 20%, and 36% by comparison with the control assay (P < or = 0.001). In addition, 1% FSP or 0.1% diosgenin fed during the promotional stage caused a significant reduction (P < or = 0.001) of multicrypt foci compared with control. Dietary diosgenin at 0.1% and 0.05% inhibited total colonic ACF and multicrypt foci formation in a dose-dependent manner. Results from the in vitro experiments indicated that diosgenin inhibits cell growth and induces apoptosis in the HT-29 human colon cancer cell line in a dose-dependent manner. Furthermore, diosgenin induced apoptosis in HT-29 cells at least in part by inhibition of bcl-2 and by induction of caspase-3 protein expression. On the basis of these findings, the fenugreek constituent diosgenin seems to have potential as a novel colon cancer preventive agent. PMID- 15298964 TI - Stability of measurements of biomarkers of oxidative stress in blood over 36 hours. AB - Oxidative stress is hypothesized to play an important role in a variety of chronic diseases, but the short-term and long-term stability of measurements of biomarkers related to oxidative stress remains unclear. The objective of this study was to evaluate the stability of measurements of malondialdehyde (MDA), F2 isoprostanes, and fluorescent oxidation products in blood stored on ice within 36 hours until processing. Whole blood samples from six healthy women were processed at 0, 24, and 36 hours after being stored on ice. MDA was measured by the thiobarbituric acid-reactive substances assay with high-pressure liquid chromatography. F2-isoprostanes were measured by gas chromatography/mass spectrometry. The fluorescent oxidation products were measured by spectrofluorometry. Measurements of fluorescent oxidation products were very stable up to 36 hours. Intraclass correlation coefficients (ICC) were >0.95 for each time interval (0 to 24 and 0 to 36 hours). Measurements of MDA were the least stable. The median increased significantly from 0 to 24 hours and from 0 to 36 hours. The ICC for MDA for each time interval (0 to 24 and 0 to 36 hours) was <0.1. Finally, the median of F2-isoprostane measurements at each time point also increased significantly. ICCs were 0.45 for 0 to 24 hours and 0.09 for 0 to 36 hours. We conclude that measurements of fluorescent oxidation products in blood remain stable for up to 36 hours and may be used in large prospective epidemiologic studies of chronic diseases. PMID- 15298965 TI - Germ line BAX alterations are infrequent in Li-Fraumeni syndrome. AB - Multiple early-onset tumors, frequently associated with germ line TP53 mutations characterize the Li-Fraumeni familial cancer syndrome (LFS). LFS-like (LFS-L) families have lower rates of germ line TP53 alteration and do not meet the strict definition of LFS. This study examined 7 LFS cell lines and 30 LFS and 36 LFS-L primary leukocyte samples for mutations in the proapoptotic p53-regulated gene BAX. No germ line BAX mutations were found. A known BAX polymorphism was observed, yet there was no correlation between polymorphism frequency and TP53 status in either LFS or LFS-L. In summary, alterations of BAX are not responsible for cancers in TP53 wild-type LFS or LFS-L families. PMID- 15298966 TI - The CYP19 gene codon 39 Trp/Arg polymorphism increases breast cancer risk in subsets of premenopausal Japanese. AB - The production of estrogen from androgen via the estrogen biosynthesis pathway is catalyzed by aromatase P450 (CYP19). To assess the association between breast cancer risk and a polymorphism at codon 39 Trp/Arg of the encoding gene, a case control study was conducted at Aichi Cancer Center Hospital in Japan. Subjects were 248 histologically confirmed breast cancer patients and 603 hospital controls without cancer. Odds ratios (OR) and 95% confidence intervals (95% CI) were determined by logistic regression analysis. The allele frequency among controls was 3.8% for the C allele, and the OR (95% CI) of the polymorphism relative to TT genotype was 1.21 (0.69-2.14) for TC/CC genotypes combined. There was no association between CYP19 gene polymorphism and breast cancer risk in the study group as a whole, but homozygous and heterozygous carriers of the variant Arg allele showed a significantly increased risk of breast cancer among premenopausal women with a late age at first full-term pregnancy (OR 7.31, 95% CI 1.88-28.5) or a high body mass index (OR 2.77, 95% CI 1.12-6.87). Additional larger studies should be done to confirm that the rare CYP19 variant increases the risk of breast cancer among premenopausal Japanese women. PMID- 15298967 TI - No association between GPX Pro198Leu and risk of basal cell carcinoma. PMID- 15298968 TI - Breast cancer risk in overweight postmenopausal women. PMID- 15298969 TI - Assessment of welfare of the child in HIV positive couples. AB - BACKGROUND: Demand for assisted conception amongst HIV-infected couples is rising in parallel with increased efficacy of antiretroviral medication which has improved life expectancy and reduced vertical transmission risk. There are no published data on welfare of the child assessment in HIV positive couples undergoing assisted conception. METHODS: We assessed welfare of the child in 131 (i.e. total number seen, not treated) couples, where one or both partners were infected with HIV and referred to the infertility clinic at Chelsea and Westminster Hospital since 1999. In total, 59 couples received sperm washing treatment (male partner infected) resulting in 17 healthy babies, and 14 couples were treated in the female positive programme (5 concordant and 9 discordant for HIV) resulting in three healthy babies. RESULTS: Issues surrounding welfare of the child were commonly encountered in this series and were significant enough to withhold treatment in five cases. Many were relationship issues surrounding acquisition of infection, fear of infection in the negative partner or child (n = 1), poor prognosis (multiple drug resistance) (n=3) or disability related to infection (n = 1). CONCLUSIONS: Welfare of the child in HIV infected couples must be carefully considered in specialist centres with experienced counsellors. Issues surrounding treatment are complex and require close liaison with HIV specialists and involvement of the couple. PMID- 15298970 TI - Nuclear chromosomal localization in human preimplantation embryos: correlation with aneuploidy and embryo morphology. AB - BACKGROUND: Spatial organization of chromosomes is hypothesized to reflect transcriptional activity and regulatory protein function. Preimplantation genetic diagnosis allows assessment of the spatial relationship of chromosomes in human blastomeres. We thus examined the localization of chromosomes 13, 16, 18, 21, 22, X and Y in blastomeres from 6-8-cell stage embryos, correlating localization to aneuploidy and embryo morphology. METHODS: Following fluorescence in situ hybridization to enumerate chromosomes 13, 16, 18, 21, 22, X and Y, signal positions were localized within one of four concentric shells. Statistical analysis compared chromosome localization between euploid and aneuploid blastomeres as well as morphologically normal and abnormal embryos. RESULTS: Of 98 embryos, 109 blastomeres were evaluated. Within chromosomally normal blastomeres, no difference in the location of all seven chromosomes (P +10 mV relative to wild type), linker prolongation by duplicating the entire linker (Dup229-237) or by glutamine insertion (InsQ233Q, InsQQ233QQ and InsQQQ233QQQ, or Ins237QQQ) produced length-dependent progressive hyperpolarizing activation shifts (-35 mV < DeltaV1/2 < -4 mV). Based on these results, we conclude that only Met232 is prerequisite for channels to function, but the length and other constituents of the S3-S4 linker shape the ultimate activation phenotype. Our results also highlight several evolutionary similarities and differences between HCN and voltage-gated K+ channels. Manipulations of the S3-S4 linker length may provide a flexible approach to customize HCN gating for engineering electrically active cells (such as stem cell derived neuronal and cardiac pacemakers) for gene- and cell-based therapies. PMID- 15299005 TI - Negative regulation of MEKK1-induced signaling by glutathione S-transferase Mu. AB - Mitogen-activated protein kinase/extracellular signal-regulated kinase kinase kinase 1 (MEKK1) is an important component in the stress-activated protein kinase pathway. Glutathione S-transferase Mu 1-1 (GST M1-1) has now been shown to inhibit the stimulation of MEKK1 activity induced by cellular stresses such as UV and hydrogen peroxide. GST M1-1 inhibited MEKK1 activation in a manner independent of its glutathione-conjugating catalytic activity. In vitro binding and kinase assays revealed that GST M1-1 directly bound MEKK1 and inhibited its kinase activity. Co-immunoprecipitation analysis showed a physical association between endogenous GST M1-1 and endogenous MEKK1 in L929 cells. Overexpressed GST M1-1 interfered with the binding of MEKK1 to SEK1 in transfected HEK293 cells. Furthermore, GST M1-1 suppressed MEKK1-mediated apoptosis. Taken together, our results suggest that GST M1-1 functions as a negative regulator of MEKK1. PMID- 15299006 TI - Allosteric regulation and temperature dependence of oxygen binding in human neuroglobin and cytoglobin. Molecular mechanisms and physiological significance. AB - Two new globin proteins have recently been discovered in vertebrates, neuroglobin in neurons and cytoglobin in all tissues, both showing heme hexacoordination by the distal His(E7) in the absence of gaseous ligands. In analogy to hemoglobin and myoglobin, neuroglobin and cytoglobin are supposedly involved in O2 storage and delivery, although their physiological role remains to be solved. Here we report O2 equilibria of recombinant human neuroglobin (NGB) and cytoglobin (CYGB) measured under close to physiological conditions and at varying temperature and pH ranges. NGB shows both alkaline and acid Bohr effects (pH-dependent O2 affinity) and temperature-dependent enthalpy of oxygenation. O2 and CO binding equilibrium studies on neuroglobin mutants strongly suggest that the bound O2 is stabilized by interactions with His(E7) and that this residue functions as a major Bohr group in the presence of Lys(E10). As shown by the titration of free thiols with 4,4'-dithiodipyridine and by mass spectrometry, this mechanism of modulating O2 affinity is independent of formation of an internal disulfide bond under the experimental conditions used, which stabilize thiols in the reduced form. In CYGB, O2 binding is cooperative, consistent with its proposed dimeric structure. Similar to myoglobin but in contrast to NGB, O2 binding to CYGB is pH independent and exothermic throughout the temperature range investigated. Our data support the hypothesis that CYGB may be involved in O2-requiring metabolic processes. In contrast, the lower O2 affinity in NGB does not appear compatible with a physiological role involving mitochondrial O2 supply at the low O2 tensions found within neurons. PMID- 15299007 TI - Unique residues on the H2A.Z containing nucleosome surface are important for Xenopus laevis development. AB - Critical to vertebrate development is a complex program of events that establishes specialized tissues and organs from a single fertilized cell. Transitions in chromatin architecture, through alterations in its composition and modification markings, characterize early development. A variant of the H2A core histone, H2A.Z, is essential for development of both Drosophila and mice. We recently showed that H2A.Z is required for proper chromosome segregation. Whether H2A.Z has additional specific functions during early development remains unknown. Here we demonstrate that depletion of H2A.Z by RNA interference perturbs Xenopus laevis development at gastrulation leading to embryos with malformed, shortened trunks. Consistent with this result, whole embryo in situ hybridization indicates that endogenous expression of H2A.Z is highly enriched in the notochord. H2A.Z modifies the surface of a canonical nucleosome by creating an extended acidic patch and a metal ion-binding site stabilized by two histidine residues. To examine the significance of these specific surface regions in vivo, we investigated the consequences of overexpressing H2A.Z and mutant proteins during X. laevis development. Overexpression of H2A.Z slowed development following gastrulation. Altering the extended acidic patch of H2A.Z reversed this effect. Remarkably, modification of a single stabilizing histidine residue located on the exposed surface of an H2A.Z containing nucleosome was sufficient to disrupt normal trunk formation mimicking the effect observed by RNA interference. Taken together, these results argue that key determinants located on the surface of an H2A.Z nucleosome play an important specific role during embryonic patterning and provide a link between a chromatin structural modification and normal vertebrate development. PMID- 15299008 TI - Domain architectures and characterization of an RNA-binding protein, TLS. AB - Translocated in liposarcoma (TLS) is an important protein component of the heterogeneous nuclear ribonucleoprotein complex involved in the splicing of pre mRNA and the export of fully processed mRNA to the cytoplasm. We examined the domain organization of human TLS by a combined approach using limited proteolysis, matrix-assisted laser desorption ionization time-of-flight mass spectrometry, circular dichroism, inductively coupled plasma atomic emission spectroscopy, and NMR spectroscopy. We found that the RNA recognition motif (RRM) and zinc finger-like domains exclusively form protease-resistant core structures within the isolated TLS protein fragments, while the remaining regions, including the Arg-Gly-Gly repeats, appear to be completely unstructured. Thus, TLS contains the unstructured N-terminal half followed by the RRM and zinc finger-like domains, which are connected to each other by a flexible linker. We also carried out NMR analyses to obtain more detailed insights into the individual RRM and zinc finger-like domains. The 113Cd NMR analysis of the zinc finger-like domain verified that zinc is coordinated with four cysteines in the C4 type scheme. We also investigated the interaction of each domain with an oligo-RNA containing the GGUG sequence, which appears to be critical for the TLS function in splicing. The backbone amide NMR chemical shift perturbation analyses indicated that the zinc finger domain binds GGUG-containing RNA with a dissociation constant of about 1.0 x 10(-5) m, whereas the RRM domain showed no observable interaction with this RNA. This surprising result implies that the zinc finger domain plays a more predominant role in RNA recognition than the RRM domain. PMID- 15299009 TI - A disorder to order transition accompanies catalysis in retinaldehyde dehydrogenase type II. AB - Retinaldehyde dehydrogenase II (RalDH2) converts retinal to the transcriptional regulator retinoic acid in the developing embryo. The x-ray structure of the enzyme revealed an important structural difference between this protein and other aldehyde dehydrogenases of the same enzyme superfamily; a 20-amino acid span in the substrate access channel in retinaldehyde dehydrogenase II is disordered, whereas in other aldehyde dehydrogenases this region forms a well defined wall of the substrate access channel. We asked whether this disordered loop might order during the course of catalysis and provide a means for an enzyme that requires a large substrate access channel to restrict access to the catalytic machinery by smaller compounds that might potentially enter the active site and be metabolized. Our experiments, a combination of kinetic, spectroscopic, and crystallographic techniques, suggest that a disorder to order transition is linked to catalytic activity. PMID- 15299010 TI - In vivo bypass efficiencies and mutational signatures of the guanine oxidation products 2-aminoimidazolone and 5-guanidino-4-nitroimidazole. AB - The in vivo mutagenic properties of 2-aminoimidazolone and 5-guanidino-4 nitroimidazole, two products of peroxynitrite oxidation of guanine, are reported. Two oligodeoxynucleotides of identical sequence, but containing either 2 aminoimidazolone or 5-guanidino-4-nitroimidazole at a specific site, were ligated into single-stranded M13mp7L2 bacteriophage genomes. Wild-type AB1157 Escherichia coli cells were transformed with the site-specific 2-aminoimidazolone- and 5 guanidino-4-nitroimidazole-containing genomes, and analysis of the resulting progeny phage allowed determination of the in vivo bypass efficiencies and mutational signatures of the DNA lesions. 2-Aminoimidazolone was efficiently bypassed and 91% mutagenic, producing almost exclusively G to C transversion mutations. In contrast, 5-guanidino-4-nitroimidazole was a strong block to replication and 50% mutagenic, generating G to A, G to T, and to a lesser extent, G to C mutations. The G to A mutation elicited by 5-guanidino-4-nitroimidazole implicates this lesion as a novel source of peroxynitrite-induced transition mutations in vivo. For comparison, the error-prone bypass DNA polymerases were overexpressed in the cells by irradiation with UV light (SOS induction) prior to transformation. SOS induction caused little change in the efficiency of DNA polymerase bypass of 2-aminoimidazolone; however, bypass of 5-guanidino-4 nitroimidazole increased nearly 10-fold. Importantly, the mutation frequencies of both lesions decreased during replication in SOS-induced cells. These data suggest that 2-aminoimidazolone and 5-guanidino-4-nitroimidazole in DNA are substrates for one or more of the SOS-induced Y-family DNA polymerases and demonstrate that 2-aminoimidazolone and 5-guanidino-4-nitroimidazole are potent sources of mutations in vivo. PMID- 15299011 TI - The vitamin D response element in the distal osteocalcin promoter contributes to chromatin organization of the proximal regulatory domain. AB - Vitamin D receptor (VDR) and Runx2 are key regulators of tissue-specific gene transcription. Using the bone-related osteocalcin (OC) gene, we have previously shown that Runx2 is required for the extensive chromatin remodeling that accompanies gene activation. Here, we have addressed the direct contribution of the VDR to chromatin remodeling events necessary for regulation of OC transcription using mutational analysis. Our studies demonstrate that both the distal and proximal DNase I-hypersensitive sites characteristic of the transcriptionally active OC promoter are not enhanced in the absence of a functional vitamin D response element (VDRE). Furthermore, restriction enzyme accessibility studies reveal that nucleosomal reorganization of the proximal promoter occurs in response to vitamin D and this reorganization is abrogated by mutation of the VDRE. These findings indicate that binding of liganded VDR in the distal promoter directly impacts the chromatin structure of the proximal promoter. We find that, in the absence of functional Runx sites, the VDR cannot be recruited to the OC promoter and, therefore, the VDRE is not competent to mediate vitamin D responsiveness. On the other hand, chromatin immunoprecipitation assays show that Runx2 association with the OC promoter is not significantly impaired when the VDRE is mutated. Chromatin immunoprecipitation assays also demonstrate that basal levels of histone acetylation occur in the absence of Runx2 binding but that the VDRE and vitamin D are required for enhanced acetylation of histones H3 and H4 downstream of the VDRE. Together our results support a stepwise model for chromatin remodeling of the OC promoter and show that binding of the liganded VDR.retinoid X receptor directly impacts both the distal and proximal regulatory domains. PMID- 15299012 TI - Transformation of the mechanism of triple-helix peptide folding in the absence of a C-terminal nucleation domain and its implications for mutations in collagen disorders. AB - Folding abnormalities of the triple helix have been demonstrated in collagen diseases such as osteogenesis imperfecta in which the mutation leads to the substitution of a single Gly in the (Gly-X-Y)n sequence pattern by a larger residue. Model peptides can be used to clarify the details of normal collagen folding and the consequences of the interruption of that folding by a Gly substitution. NMR and CD studies show that placement of a (GPO)4 nucleation domain at the N terminus rather than the C terminus of a native collagen sequence allows the formation of a stable triple helix but alters the folding mechanism. Although C- to N-terminal directional folding occurs when the nucleation domain is at the C terminus, there is no preferential folding direction when the nucleation domain is at the N terminus. The lack of zipper-like directional folding does not interfere with triple-helix formation, and when a Gly residue is replaced by Ser to model an osteogenesis imperfecta mutation, the peptide with the N-terminal (GPO)4 domain can still form a good triple helix N-terminal to the mutation site. These peptide studies raise the possibility that mutant collagen could fold in a C to N direction in a zipper-like manner up to the mutation site and that completion of the triple helix N-terminal to the mutation would involve an alternative mechanism. PMID- 15299013 TI - The alpha1.alpha2 network of collagen IV. Reinforced stabilization of the noncollagenous domain-1 by noncovalent forces and the absence of Met-Lys cross links. AB - Collagen IV networks are present in all metazoa and underlie epithelia as a component of basement membranes. The networks are essential for tissue function and are defective in disease. They are assembled by the oligomerization of triple helical protomers that are linked end-to-end. At the C terminus, two protomers are linked head-to-head by interactions of their trimeric noncollagenous domains, forming a hexamer structure. This linkage in the alpha1.alpha2 network is stabilized by a putative covalent Met-Lys cross-link between the trimer-trimer interface (Than, M. E., Henrich, S., Huber, R., Ries, A., Mann, K., Kuhn, K., Timpl, R., Bourenkov, G. P., Bartunik, H. D., and Bode, W. (2002) Proc. Natl. Acad. Sci. U. S. A. 99, 6607-6612) forming a nonreducible dimer that connects the hexamer. In the present study, this cross-link was further investigated by: (a) comparing the 1.5-A resolution crystal structures of the alpha1.alpha2 hexamers from bovine placenta and lens capsule basement membranes, (b) mass spectrometric analysis of monomer and nonreducible dimer subunits of placenta basement membrane hexamers, and (c) hexamer dissociation/re-association studies. The findings rule out the novel Met-Lys cross-link, as well as other covalent cross-links, but establish that the nonreducible dimer is an inherent structural feature of a subpopulation of hexamers. The dimers reflect the reinforced stabilization, by noncovalent forces, of the connection between two adjoining protomers of a network. The reinforcement extends to other types of collagen IV networks, and it underlies the cryptic nature of a B-cell epitope of the alpha3.alpha4.alpha5 hexamer, implicating the stabilization event in the etiology and pathogenesis of Goodpasture autoimmune disease. PMID- 15299014 TI - Synchronized chemoenzymatic synthesis of monodisperse hyaluronan polymers. AB - The length of the hyaluronan (HA) polysaccharide chain dictates its biological effects in many cellular and tissue systems. Long and short HA polymers often appear to have antagonistic or inverse effects. However, no source of very defined, uniform HA polymers with sizes greater than 10 kDa is currently available. We present a method to produce synthetic HA with very narrow size distributions in the range of approximately 16 kDa to approximately 2 MDa. The Pasteurella HA synthase enzyme, pmHAS, catalyzes the synthesis of HA polymer utilizing monosaccharides from UDP-sugar precursors. Recombinant pmHAS will also elongate exogenously supplied HA oligosaccharide acceptors in vitro in a nonprocessive fashion. As a result of bypassing the slow initiation step in vitro, the elongation process is synchronized in the presence of acceptor; thus all of polymer products are very similar in length. In contrast, without the use of an acceptor, the final polymer size range is difficult to predict and the products are more polydisperse. HA polymers of a desired size are constructed by controlling the reaction stoichiometry (i.e. molar ratio of precursors and acceptor molecules). The use of modified acceptors allows the synthesis of HA polymers containing tags (e.g. fluorescent, radioactive). In this scheme, each molecule has a single foreign moiety at the reducing terminus. Alternatively, the use of radioactive UDP-sugar precursors allows the synthesis of uniformly labeled native HA polymers. Overall, synthetic HA reagents with monodisperse size distributions and defined structures should assist in the elucidation of the numerous roles of HA in health and disease. PMID- 15299015 TI - The COOH terminus of the amelogenin, LRAP, is oriented next to the hydroxyapatite surface. AB - The organic matrix in forming enamel consists largely of the amelogenin protein self-assembled into nanospheres that are necessary to guide the formation of the unusually long and highly ordered hydroxyapatite (HAP) crystallites that constitute enamel. Despite its ability to direct crystal growth, the interaction of the amelogenin protein with HAP is unknown. However, the demonstration of growth restricted to the c-axis suggests a specific protein-crystal interaction, and the charged COOH terminus is often implicated in this function. To elucidate whether the COOH terminus is important in the binding and orientation of amelogenin onto HAP, we have used solid state NMR to determine the orientation of the COOH terminus of an amelogenin splice variant, LRAP (leucine-rich amelogenin protein), which contains the charged COOH terminus of the full protein, on the HAP surface. These experiments demonstrate that the methyl 13C-labeled side chain of Ala46 is 8.0 A from the HAP surface under hydrated conditions, for the protein with and without phosphorylation. The experimental results provide direct evidence orienting the charged COOH-terminal region of the amelogenin protein on the HAP surface, optimized to exert control on developing enamel crystals. PMID- 15299016 TI - Dependence of site-2 protease cleavage of ATF6 on prior site-1 protease digestion is determined by the size of the luminal domain of ATF6. AB - ATF6 is an endoplasmic reticulum (ER) membrane-anchored transcription factor activated by regulated intramembrane proteolysis in the ER stress response. The release of the cytosolic transcription factor domain of ATF6 requires the sequential processing by the Golgi site-1 and site-2 proteases (S1P and S2P). It has been unclear why S2P proteolysis relies on previous site-1 cleavage. One possibility is that S2P localizes to a different cellular compartment than S1P; however, here we show that S2P localizes to the same compartment as S1P, the cis/medial-Golgi. In addition, we have re-localized S1P and S2P to the ER with brefeldin A and find that the sequential cleavage of ATF6 is reconstituted in the ER. The mapping of the region of ATF6 required for sequential S1P and S2P cleavage showed that short luminal domains resulted in S1P-independent S2P cleavage. The addition of artificial domains onto these short ATF6 luminal domains restored the S1P dependence of S2P cleavage, suggesting that it is the size rather than specific sequences in the luminal domain that determines the S1P dependence of S2P cleavage. These results suggest that the bulky ATF6 luminal domain blocks S2P cleavage and that the role of S1P is to reduce the size of the luminal domain to prepare ATF6 to be an optimal S2P substrate. PMID- 15299017 TI - Phosphorylation of fanconi anemia (FA) complementation group G protein, FANCG, at serine 7 is important for function of the FA pathway. AB - Fanconi anemia (FA) is an autosomal recessive disease of cancer susceptibility. FA cells exhibit a characteristic hypersensitivity to DNA cross-linking agents. The molecular mechanism for the disease is unknown as few of the FA proteins have functional motifs. Several post-translational modifications of the proteins have been described. We and others have reported that the FANCG protein (Fanconi complementation group G) is phosphorylated. We show that in an in vitro kinase reaction FANCG is radioactively labeled. Mass spectrometry analysis detected a peptide containing phosphorylation of serine 7. Using PCR-mediated site-directed mutagenesis we mutated serine 7 to alanine. Only wild-type FANCG cDNA fully corrected FA-G mutant cells. We also tested the effect of human wild-type FANCG in Chinese hamster ovary cells in which the FANCG homologue is mutant. Human FANCG complemented these cells, whereas human FANCG(S7A) did not. Unexpectedly, FANCG(S7A) bound to and stabilized the endogenous forms of the FANCA and FANCC proteins in the FA-G cells. FANCG(S7A) aberrantly localized to globules in chromatin and did not abrogate the internuclear bridges seen in the FA-G mutant cells. Phosphorylation of serine 7 in FANCG is functionally important in the FA pathway. PMID- 15299018 TI - Retinoic acid inhibition of chromatin remodeling at the human immunodeficiency virus type 1 promoter. Uncoupling of histone acetylation and chromatin remodeling. AB - All-trans retinoic acid (RA) represses HIV-1 transcription and replication in cultured monocytic cells and in primary monocyte-derived macrophages. Here we examine the role of histone acetylation and chromatin remodeling in RA-mediated repression. RA pretreatment of latently infected U1 promonocytes inhibits HIV-1 expression in response to the histone deacetylase (HDAC) inhibitor, trichostatin A (TSA). TSA is thought to activate HIV-1 transcription by inducing histone hyperacetylation within a regulatory nucleosome, nuc-1, positioned immediately downstream from the transcription start site. Acetylation of nuc-1 is thought to be a critical step in activation that precedes nuc-1 remodeling and, subsequently, transcriptional initiation. Here we demonstrate that TSA treatment induces H3 and H4 hyperacetylation and nuc-1 remodeling. Although RA pretreatment inhibits nuc-1 remodeling and HIV-1 transcription, it has no effect on histone acetylation. This suggests that acetylation and remodeling are not obligatorily coupled. We also show that growth of U1 cells in retinoid-deficient medium induces nuc-1 remodeling and HIV-1 expression but does not induce histone hyperacetylation. These findings suggest that remodeling, not histone hyperacetylation, is the limiting step in transcriptional activation in these cells. Together, these data suggest that RA signaling maintains the chromatin structure of the HIV-1 promoter in a transcriptionally non-permissive state that may contribute to the establishment of latency in monocyte/macrophages. PMID- 15299019 TI - Human tribbles, a protein family controlling mitogen-activated protein kinase cascades. AB - Control of mitogen-activated protein kinase (MAPK) cascades is central to regulation of many cellular responses. We describe here human tribbles homologues (Htrbs) that control MAPK activity. MAPK kinases interact with Trbs and regulate their steady state levels. Further, Trbs selectively regulate the activation of extracellular signal-regulated kinases, c-Jun NH2-terminal kinases, and p38 MAPK with different relative levels of activity for the three classes of MAPK observed depending on the level of Trb expression. These results suggest that Trbs control both the extent and the specificity of MAPK kinase activation of MAPK. PMID- 15299020 TI - Regulated membrane recruitment of dynamin-2 mediated by sorting nexin 9. AB - The endocytic proteins sorting nexin 9 (SNX9) and dynamin-2 (Dyn2) assemble in the cytosol as a resting complex, together with a 41-kDa protein. We show here that the complex can be activated for membrane binding of SNX9 and Dyn2 by incubation of cytosol in the presence of ATP. SNX9 was essential for Dyn2 recruitment, whereas the reverse was not the case. RNA interference experiments confirmed that SNX9 functions as a mediator of Dyn2 recruitment to membranes in cells. The 41-kDa component was identified as the glycolytic enzyme aldolase. Aldolase bound with high affinity to a tryptophan-containing acidic sequence in SNX9 located close to its Phox homology domain, thereby blocking the membrane binding activity of SNX9. Phosphorylation of SNX9 released aldolase from the native cytosolic complex and rendered SNX9 competent for membrane binding. The results suggest that SNX9-dependent recruitment of Dyn2 to the membrane is regulated by an interaction between SNX9 and aldolase. PMID- 15299021 TI - Quantifying the effects of molecular orientation and length on two-dimensional receptor-ligand binding kinetics. AB - Surface presentation of adhesion receptors influences cell adhesion, although the mechanisms underlying these effects are not well understood. We used a micropipette adhesion frequency assay to quantify how the molecular orientation and length of adhesion receptors on the cell membrane affected two-dimensional kinetic rates of interactions with surface ligands. Interactions of P-selectin, E selectin, and CD16A with their respective ligands or antibody were used to demonstrate such effects. Randomizing the orientation of the adhesion receptor or lowering its ligand- and antibody-binding domain above the cell membrane lowered two-dimensional affinities of the molecular interactions by reducing the forward rates but not the reverse rates. In contrast, the soluble antibody bound with similar three-dimensional affinities to cell-bound P-selectin constructs regardless of their orientation and length. These results demonstrate that the orientation and length of an adhesion receptor influences its rate of encountering and binding a surface ligand but does not subsequently affect the stability of binding. PMID- 15299022 TI - Transcript scanning reveals novel and extensive splice variations in human l-type voltage-gated calcium channel, Cav1.2 alpha1 subunit. AB - The L-type (Cav1.2) voltage-gated calcium channels play critical roles in membrane excitability, gene expression, and muscle contraction. The generation of splice variants by the alternative splicing of the poreforming Cav1.2 alpha1 subunit (alpha(1)1.2) may thereby provide potent means to enrich functional diversity. To date, however, no comprehensive scan of alpha(1)1.2 splice variation has been performed, particularly in the human context. Here we have undertaken such a screen, exploiting recently developed "transcript scanning" methods to probe the human gene. The degree of variation turns out to be surprisingly large; 19 of the 55 exons comprising the human alpha(1)1.2 gene were subjected to alternative splicing. Two of these are previously unrecognized exons and two others were not known to be spliced. Comparisons of fetal and adult heart and brain uncovered a large IVS3-S4 variability resulting from combinatorial utilization of exons 31-33. Electrophysiological characterization of such IVS3-S4 variation revealed unmistakable shifts in the voltage dependence of activation, according to an interesting correlation between increased IVS3-S4 linker length and activation at more depolarized potentials. Steady-state inactivation profiles remained unaltered. This systematic portrait of splice variation furnishes a reference library for comprehending combinatorial arrangements of Cav1.2 splice exons, especially as they impact development, physiology, and disease. PMID- 15299023 TI - cAMP-dependent protein kinase type I regulates ethanol-induced cAMP response element-mediated gene expression via activation of CREB-binding protein and inhibition of MAPK. AB - We have shown that the two types of cAMP-dependent protein kinase (PKA) in NG108 15 cells differentially mediate forskolin- and ethanol-induced cAMP response element (CRE)-binding protein (CREB) phosphorylation and CRE-mediated gene transcription. Activated type II PKA is translocated into the nucleus where it phosphorylates CREB. By contrast, activated type I PKA does not translocate to the nucleus but is required for CRE-mediated gene transcription by inducing the activation of other transcription cofactors such as CREB-binding protein (CBP). We show here that CBP is required for forskolin- and ethanol-induced CRE-mediated gene expression. Forskolin- and ethanol-induced CBP phosphorylation, demonstrable at 10 min, persists up to 24 h. CBP phosphorylation requires type I PKA but not type II PKA. In NG108-15 cells, ethanol and forskolin activation of type I PKA also inhibits several components of the MAPK pathway including B-Raf kinase, ERK1/2, and p90RSK phosphorylation. As a result, unphosphorylated p90RSK no longer binds to nor inhibits CBP. Moreover, MEK inhibition by PD98059 induces a significant increase of CRE-mediated gene activation. Taken together, our findings suggest that inhibition of the MAPK pathway enhances cAMP-dependent gene activation during exposure of NG108-15 cells to ethanol. This mechanism appears to involve type I PKA-dependent phosphorylation of CBP and inhibition of MEK dependent phosphorylation of p90RSK. Under these conditions p90RSK is no longer bound to CBP, thereby promoting CBP-dependent CREB-mediated gene expression. PMID- 15299024 TI - The cysteine-rich region of T1R3 determines responses to intensely sweet proteins. AB - A wide variety of chemically diverse compounds taste sweet, including natural sugars such as glucose, fructose, sucrose, and sugar alcohols, small molecule artificial sweeteners such as saccharin and acesulfame K, and proteins such as monellin and thaumatin. Brazzein, like monellin and thaumatin, is a naturally occurring plant protein that humans, apes, and Old World monkeys perceive as tasting sweet but that is not perceived as sweet by other species including New World monkeys, mouse, and rat. It has been shown that heterologous expression of T1R2 plus T1R3 together yields a receptor responsive to many of the above mentioned sweet tasting ligands. We have determined that the molecular basis for species-specific sensitivity to brazzein sweetness depends on a site within the cysteine-rich region of human T1R3. Other mutations in this region of T1R3 affected receptor activity toward monellin, and in some cases, overall efficacy to multiple sweet compounds, implicating this region as a previously unrecognized important determinant of sweet receptor function. PMID- 15299025 TI - Integrin-linked kinase regulates the nuclear entry of the c-Jun coactivator alpha NAC and its coactivation potency. AB - Overexpression of the integrin-linked kinase (ILK) was shown to increase c-Jun dependent transcription. We now show that this effect of ILK involves the c-Jun transcriptional coactivator, nascent polypeptide-associated complex and coactivator alpha (alpha-NAC). ILK phosphorylated alpha-NAC on residue Ser-43 upon adhesion of cells to fibronectin. Co-expression of constitutively active ILK with alpha-NAC led to the nuclear accumulation of the coactivator. Conversely, alpha-NAC remained in the cytoplasm of cells transfected with a dominant-negative ILK mutant, and a mutated alpha-NAC at phosphoacceptor position Ser-43 (S43A) also localized outside of the nucleus. The S43A alpha-NAC mutant could not potentiate the effect of ILK on c-Jun-dependent transcription. We conclude that ILK-dependent phosphorylation of alpha-NAC induced the nuclear accumulation of the coactivator and that phosphorylation of alpha-NAC by ILK is required for the potentiation of c-Jun-mediated responses by the kinase. The results represent one of the rare examples of a transcriptional coactivator shuttling between the cytosol and the nucleus. PMID- 15299026 TI - Characterization of the calcium-mediated response to alkaline stress in Saccharomyces cerevisiae. AB - Exposure of the yeast Saccharomyces cerevisiae to alkaline stress resulted in adaptive changes that involved remodeling the gene expression. Recent evidence suggested that the calcium-activated protein phosphatase calcineurin could play a role in alkaline stress signaling. By using an aequorin luminescence reporter, we showed that alkaline stress resulted in a sharp and transient rise in cytoplasmic calcium. This increase was largely abolished by addition of EGTA to the medium or in cells lacking Mid1 or Cch1, components of the high affinity cell membrane calcium channel. Under these circumstances, the alkaline response of different calcineurin-sensitive transcriptional promoters was also blocked. Therefore, exposure to alkali resulted in entry of calcium from the external medium, and this triggered a calcineurin-mediated response. The involvement of calcineurin and Crz1/Tcn1, the transcription factor activated by the phosphatase, in the transcriptional response triggered by alkalinization has been globally assessed by DNA microarray analysis in a time course experiment using calcineurin deficient (cnb1) and crz1 mutants. We found that exposure to pH 8.0 increased at least 2-fold the mRNA levels of 266 genes. In many cases (60%) the response was rather early (peak after 10 min). The transcriptional response of 27 induced genes (10%) was reduced or fully abolished in cnb1 cells. In general, the response of crz1 mutants was similar to that of calcineurin-deficient cells. By analysis of a systematic deletion library, we found 48 genes whose mutation resulted in increased sensitivity to the calcineurin inhibitor FK506. Twenty of these mutations (42%) also provoked alkaline pH sensitivity. In conclusion, our results demonstrated that calcium signaling and calcineurin activation represented a significant component of the yeast response to environmental alkalinization. PMID- 15299027 TI - Multiple functions of Jab1 are required for early embryonic development and growth potential in mice. AB - Jab1 interacts with a variety of signaling molecules and regulates their stability in mammalian cells. As the fifth component of the COP9 signalosome (CSN) complex, Jab1 (CSN5) plays a central role in the deneddylation of the cullin subunit of the Skp1-Cullin-F box protein ubiquitin ligase complex. In addition, a CSN-independent function of Jab1 is suggested but is less well characterized. To elucidate the function of Jab1, we targeted the Jab1 locus by homologous recombination in mouse embryonic stem cells. Jab1-null embryos died soon after implantation. Jab1-/- embryonic cells, which lacked other CSN components, expressed higher levels of p27, p53, and cyclin E, resulting in impaired proliferation and accelerated apoptosis. Jab1 heterozygous mice were healthy and fertile but smaller than their wild-type littermates. Jab1+/- mouse embryonic fibroblast cells, in which the amount of Jab1-containing small subcomplex, but not that of CSN, was selectively reduced, proliferated poorly, showed an inefficient down-regulation of p27 during G1, and was delayed in the progression from G0 to S phase by 3 h compared with the wild-type cells. Most interestingly, in Jab1+/- mouse embryonic fibroblasts, the levels of cyclin E and deneddylated Cul1 were unchanged, and p53 was not induced. Thus, Jab1 controls cell cycle progression and cell survival by regulating multiple cell cycle signaling pathways. PMID- 15299028 TI - Fos-related antigen 2 controls protein kinase A-induced CCAAT/enhancer-binding protein beta expression in osteoblasts. AB - Transcription factor CCAAT/enhancer-binding protein beta (C/EBPbeta) plays an important role in hormone-dependent gene expression. In osteoblasts C/EBPbeta can increase insulin-like growth factor I (IGF-I) transcription following treatment with hormones that activate protein kinase A, but little is known as yet about the expression of C/EBPbeta itself in these cells. We initially showed that prostaglandin E2 (PGE2) rapidly enhances C/EBPbeta mRNA and protein expression, and in this study we identified a 3'-proximal region of the C/EBPbeta promoter containing a 541-bp upstream sequence that could account for this effect. PGE2 dependent activation of C/EBPbeta was blocked by expression of a mutated regulatory subunit of protein kinase A or by mutation of two previously identified cAMP-sensitive cis-acting regulatory elements within the promoter between bp -111 and -61. Nuclear protein binding to these elements was induced by PGE2, required new protein synthesis, and was sensitive to antibody to the transcription factor termed Fos-related antigen 2 (Fra-2). Fra-2 cDNA generated from rat osteoblasts by reverse transcriptase PCR was 95% homologous to human Fra 2, and PGE2 rapidly induced Fra-2 mRNA and protein expression. Consistent with these findings, over-expression of Fra-2 significantly increased C/EBPbeta promoter activity in PGE2-induced osteoblasts, whereas expression of Fra-2 lacking its activation domain had a dominant negative inhibitory effect. Together, these results reveal a significant, hormone-dependent role for Fra-2 in osteoblast function, both directly, through its ability to increase new C/EBPbeta gene expression, and indirectly, through downstream C/EBP sensitive genes. PMID- 15299029 TI - Human intestinal epithelial cell survival and anoikis. Differentiation state distinct regulation and roles of protein kinase B/Akt isoforms. AB - We have shown previously that human intestinal epithelial cell survival and anoikis are distinctively regulated according to the state of differentiation. Here we analyzed the roles of protein kinase B/Akt isoforms in such differentiation state distinctions. Anoikis was induced in undifferentiated and differentiated enterocytes by inhibition of focal adhesion kinase (Fak; pharmacologic inhibition or overexpression of dominant-negative mutants) or beta1 integrins (antibody blocking) or by maintaining cells in suspension. Expression/activation parameters of Akt isoforms (Akt-1, Akt-2, and Akt-3) and Fak were analyzed. Activity of Akt isoforms was also blocked by inhibition of phosphatidylinositol 3-kinase or by overexpression of dominant-negative mutants. Here we report the following. 1) The expression/activation levels of Akt-1 increase overall during enterocytic differentiation, and those of Akt-2 decrease, whereas Akt-3 is not expressed. 2) Akt-1 activation is dependent on beta1 integrins/Fak signaling, regardless of the differentiation state. 3) Akt-2 activation is dependent on beta1 integrins/Fak signaling in undifferentiated cells only. 4) Activation of Akt-1 is phosphatidylinositol 3-kinase-dependent, whereas that of Akt-2 is not. 5) Akt-2 does not promote survival or apoptosis/anoikis. 6) Akt-1 is essential for survival. 7) Akt-2 cannot substitute for Akt-1 in the suppression of anoikis. Hence, the expression and regulation of Akt isoforms show differentiation state-specific distinctions that ultimately reflect upon their selective implication in the mediation of human intestinal epithelial cell survival. These data provide new insights into the synchronized regulation of cell survival/death that is required in the dynamic renewal process of tissues such as the intestinal epithelium. PMID- 15299030 TI - The Fanconi anemia proteins functionally interact with the protein kinase regulated by RNA (PKR). AB - Protein kinase regulated by RNA (PKR) plays critical roles in cell growth and apoptosis and is implicated as a potential pathogenic factor of Alzheimer's, Parkinson's, and Huntington's diseases. Here we report that this proapoptotic kinase is also involved in Fanconi anemia (FA), a disease characterized by bone marrow (BM) failure and leukemia. We have used a BM extract to show that three FA proteins, FANCA, FANCC, and FANCG, functionally interact with the PKR kinase, which in turn regulates translational control. By using a combined immunoprecipitation and reconstituted kinase assay, in which an active PKR kinase complex was captured from a normal cell extract, we demonstrated functional interactions between the FA proteins and the PKR kinase. In primary human BM cells, mutations in the FANCA, FANCC, and FANCG genes markedly increase the amount of PKR bound to FANCC, and this PKR accumulation is correlated with elevated PKR activation and hypersensitivity of BM progenitor cells to growth repression mediated by the inhibitory cytokines interferon-gamma and tumor necrosis factor-alpha. Specific inhibition of PKR by 2-aminopurine in these FA BM cells attenuates PKR activation and apoptosis induction. In lymphoblasts derived from an FA-C patient, overexpression of a dominant negative mutant PKR (PKRK296R) suppressed PKR activation and apoptosis induced by interferon-gamma and tumor necrosis factor-alpha. Furthermore, by using genetically matched wild-type and PKR-null cells, we demonstrated that forced expression of a patient-derived FA-C mutant (FANCCL554P) augmented double-stranded RNA-induced PKR activation and cell death. Thus, inappropriate activation of PKR as a consequence of certain FA mutations might play a role in bone marrow failure that frequently occurred in FA. PMID- 15299031 TI - Serine 18 phosphorylation of RAX, the PKR activator, is required for PKR activation and consequent translation inhibition. AB - It is now apparent that the double-stranded (ds)RNA-dependent protein kinase, PKR, is a regulator of diverse cellular responses to stress. Recently, the murine dsRNA-binding protein RAX and its human ortholog PACT were identified as cellular activators of PKR. Previous reports demonstrate that following stress, RAX/PACT associates with and activates PKR resulting in eIF2alpha phosphorylation, consequent translation inhibition, and cell death via apoptosis. Although RAX/PACT is phosphorylated during stress, any regulatory role for this post translational modification has been uncertain. Now we have discovered that RAX is phosphorylated on serine 18 in both human and mouse cells. The non phosphorylatable form of RAX, RAX(S18A), although still able to bind dsRNA and associate with PKR, fails to activate PKR following stress. Furthermore, stable expression of RAX(S18A) results in a dominant-negative effect characterized by deficiency of eukaryotic initiation factor 2 alpha subunit phosphorylation, delay of translation inhibition, and failure to undergo rapid apoptosis following removal of interleukin-3. We propose that the ability of RAX to activate PKR is regulated by a sequential mechanism featuring RAX association with PKR, RAX phosphorylation at serine 18, and activation of PKR. PMID- 15299032 TI - A novel adaptation of the integrin PSI domain revealed from its crystal structure. AB - Integrin beta-subunits contain an N-terminal PSI (for plexin-semaphorin-integrin) domain that contributes to integrin activation and harbors the PI(A) alloantigen associated with immune thrombocytopenias and susceptibility to sudden cardiac death. Here we report the crystal structure of PSI in the context of the crystallized alphaVbeta3 ectodomain. The integrin PSI forms a two-stranded antiparallel beta-sheet flanked by two short helices; its long interstrand loop houses Pl(A) and may face the EGF2 domain. The integrin PSI contains four cysteine pairs connected in a 1-4, 2-8, 3-6, 5-7 pattern. An unexpected feature of the structure is that the final, eighth cysteine is located C-terminal to the Ig-like hybrid domain and is thus separated by the hybrid domain from the other seven cysteines of PSI. This architecture may be relevant to the evolution of integrins and should help refine the current models of integrin activation. PMID- 15299033 TI - Role of trehalose phosphate synthase and trehalose during hypoxia: from flies to mammals. AB - Trehalose is a nonreducing disaccharide in which the two glucose units are linked in an alpha,alpha-1,1-glycosidic linkage. The best known and most widely distributed pathway of trehalose synthesis involves the transfer of glucose from UDP-glucose to glucose 6-phosphate to form trehalose-6-phosphate and UDP via the trehalose-6-phosphate synthase (TPS1). Trehalose-6-phosphate phosphatase (TPS2) then converts trehalose-6-phosphate to free trehalose. This sugar is present in a wide variety of organisms, including bacteria, yeast, fungi, insects, invertebrates and plants, and because of its particular physical features, trehalose is able to protect the integrity of cells against a variety of environmental stresses such as desiccation, dehydration, heat, cold and oxidation. Our current studies described here indicate that trehalose protects Drosophila and mammalian cells from hypoxic and anoxic injury. The mechanism of this protection is probably related to a decrease in protein denaturation through protein-trehalose interactions. PMID- 15299034 TI - Hypoxic survival strategies in two fishes: extreme anoxia tolerance in the North European crucian carp and natural hypoxic preconditioning in a coral-reef shark. AB - Especially in aquatic habitats, hypoxia can be an important evolutionary driving force resulting in both convergent and divergent physiological strategies for hypoxic survival. Examining adaptations to anoxic/hypoxic survival in hypoxia tolerant animals may offer fresh ideas for the treatment of hypoxia-related diseases. Here, we summarise our present knowledge of two fishes that have evolved to survive hypoxia under very different circumstances. The crucian carp (Carassius carassius) is of particular interest because of its extreme anoxia tolerance. During the long North European winter, it survives for months in completely oxygen-deprived freshwater habitats. The crucian carp also tolerates a few days of anoxia at room temperature and, unlike anoxia-tolerant freshwater turtles, it is still physically active in anoxia. Moreover, the crucian carp does not appear to reduce neuronal ion permeability during anoxia and may primarily rely on more subtle neuromodulatory mechanisms for anoxic metabolic depression. The epaulette shark (Hemiscyllium ocellatum) is a tropical marine vertebrate. It lives on shallow reef platforms that repeatedly become cut off from the ocean during periods of low tides. During nocturnal low tides, the water [O(2)] can fall by 80% due to respiration of the coral and associated organisms. Since the tides become lower and lower over a period of a few days, the hypoxic exposure during subsequent low tides will become progressively longer and more severe. Thus, this shark is under a natural hypoxic preconditioning regimen. Interestingly, hypoxic preconditioning lowers its metabolic rate and its critical P(O(2)). Moreover, repeated anoxia appears to stimulate metabolic depression in an adenosine-dependent way. PMID- 15299035 TI - Negotiating brain anoxia survival in the turtle. AB - The turtle brain's extraordinary ability to tolerate anoxia is based on constitutive and expressed factors. Constitutive factors that predispose for anoxia tolerance include enhanced levels of glycogen stores, increased densities of protective receptors, elevated antioxidant capacities and elevated heat shock protein. However, to survive an anoxic insult, three distinct phases must be negotiated successfully. (1) A coordinated downregulation of ATP demand processes to basal levels. This phase, which takes 1-2 h, includes a reduction in voltage gated K(+) (Kv) channel transcription and a substantial increase in Hsp72 and Hsc73 levels. During this period, adenosine and K(ATP) channels mediate several key events including channel arrest initiation and a reduction in the release of excitatory amino acids (EAAs). (2) Long-term survival (days) at basal levels of ATP expenditure. Neuronal network integrity is preserved through the continued operation of core activities. These include periodic electrical activity, an increased release of GABA and a continued release of glutamate and dopamine. Adenosine and GABA modulate the glutamate release. There is a further increase in Hsc73, indicating a 'housekeeping' role for this protein during this period. (3) A rapid upregulation of neuronal processes when oxygen becomes available to restore full function, together with the activation of protection mechanisms against reperfusion-generated reactive oxygen species. PMID- 15299036 TI - Hypoxia-ischemia in the immature brain. AB - The immature brain has long been considered to be resistant to the damaging effects of hypoxia and hypoxia-ischemia (H/I). However, it is now appreciated that there are specific periods of increased vulnerability, which relate to the developmental stage at the time of the insult. Although much of our knowledge of the pathophysiology of cerebral H/I is based on extensive experimental studies in adult animal models, it is important to appreciate the major differences in the immature brain that impact on its response to, and recovery from, H/I. Normal maturation of the mammalian brain is characterized by periods of limitations in glucose transport capacity and increased use of alternative cerebral metabolic fuels such as lactate and ketone bodies, all of which are important during H/I and influence the development of energy failure. Cell death following H/I is mediated by glutamate excitotoxicity and oxidative stress, as well as other events that lead to delayed apoptotic death. The immature brain differs from the adult in its sensitivity to all of these processes. Finally, the ultimate outcome of H/I in the immature brain is determined by the impact on the ensuing cerebral maturation. A hypoxic-ischemic insult of insufficient severity to result in rapid cell death and infarction can lead to prolonged evolution of tissue damage. PMID- 15299037 TI - Hypoxia tolerance in mammalian heterotherms. AB - Heterothermic mammals tolerate severe hypoxia, as well as a variety of central nervous system insults, better than homeothermic mammals. Tolerance to hypoxia may stem from adaptations associated with the ability to survive hibernation and periodic arousal thermogenesis. Here, we review evidence and mechanisms of hypoxia tolerance during hibernation, euthermy and arousal in heterothermic mammals and consider potential mechanisms for regenerative-like processes, such as synaptogenesis, observed within hours of hypoxic stress associated with arousal thermogenesis. PMID- 15299038 TI - Structural and functional adaptation to hypoxia in the rat brain. AB - Chronic exposure to a hypoxic environment leads to structural and functional adaptations in the rat brain. One significant adaptation is a decrease in intercapillary distances through a near doubling of the capillary density, which begins after about 1 week of hypoxic exposure and is completed by 3 weeks. Hypoxic angiogenesis is controlled by activation of downstream genes by Hypoxia Inducible Factor-1 and Angiopoietin-2. The processes that increase capillary density are reversible upon restoration of the ambient oxygen concentration. Capillary regression, which also occurs over a 3-week period, is accomplished through activation of apoptosis. The implication from these observations is that the brain naturally functions in a low, but controlled, oxygen environment. Acute imbalances in oxygen delivery and metabolic demand are addressed through changes in blood flow; persistent imbalances activate mechanisms that adjust capillary density. The mechanisms that control these processes decline with age. PMID- 15299039 TI - Cellular oxygen sensing need in CNS function: physiological and pathological implications. AB - Structural and functional integrity of brain function profoundly depends on a regular oxygen and glucose supply. Any disturbance of this supply becomes life threatening and may result in severe loss of brain function. In particular, reductions in oxygen availability (hypoxia) caused by systemic or local blood circulation irregularities cannot be tolerated for longer periods due to an insufficient energy supply to the brain by anaerobic glycolysis. Hypoxia has been implicated in central nervous system pathology in a number of disorders including stroke, head trauma, neoplasia and neurodegenerative disease. Complex cellular oxygen sensing systems have evolved for tight regulation of oxygen homeostasis in the brain. In response to variations in oxygen partial pressure (P(O(2))) these induce adaptive mechanisms to avoid or at least minimize brain damage. A significant advance in our understanding of the hypoxia response stems from the discovery of the hypoxia inducible factors (HIF), which act as key regulators of hypoxia-induced gene expression. Depending on the duration and severity of the oxygen deprivation, cellular oxygen-sensor responses activate a variety of short- and long-term energy saving and cellular protection mechanisms. Hypoxic adaptation encompasses an immediate depolarization block by changing potassium, sodium and chloride ion fluxes across the cellular membrane, a general inhibition of protein synthesis, and HIF-mediated upregulation of gene expression of enzymes or growth factors inducing angiogenesis, anaerobic glycolysis, cell survival or neural stem cell growth. However, sustained and prolonged activation of the HIF pathway may lead to a transition from neuroprotective to cell death responses. This is reflected by the dual features of the HIF system that include both anti- and proapoptotic components. These various responses might be based on a range of oxygen-sensing signal cascades, including an isoform of the neutrophil NADPH oxidase, different electron carrier units of the mitochondrial chain such as a specialized mitochondrial, low P(O(2)) affinity cytochrome c oxidase (aa(3)) and a subfamily of 2-oxoglutarate dependent dioxygenases termed HIF prolyl hydroxylase (PHD) and HIF asparaginyl hydroxylase, known as factor-inhibiting HIF (FIH-1). Thus specific oxygen-sensing cascades, by means of their different oxygen sensitivities, cell-specific and subcellular localization, may help to tailor various adaptive responses according to differences in tissue oxygen availability. PMID- 15299041 TI - Protective role of neuronal KATP channels in brain hypoxia. AB - During severe arterial hypoxia leading to brain anoxia, most mammalian neurons undergo a massive depolarisation terminating in cell death. However, some neurons of the adult brain and most immature nervous structures tolerate extended periods of hypoxia-anoxia. An understanding of the mechanisms underlying this tolerance to oxygen depletion is pivotal for developing strategies to protect the brain from consequences of hypoxic-ischemic insults. ATP-sensitive K(+) (K(ATP)) channels are good subjects for this study as they are activated by processes associated with energy deprivation and can counteract the terminal anoxic ischemic neuronal depolarisation. This review summarises in vitro analyses on the role of K(ATP) channels in hypoxia-anoxia in three distinct neuronal systems of rodents. In dorsal vagal neurons, blockade of K(ATP) channels with sulfonylureas abolishes the hypoxic-anoxic hyperpolarisation. However, this does not affect the extreme tolerance of these neurons to oxygen depletion as evidenced by a moderate and sustained increase of intracellular Ca(2+) (Ca(i)). By contrast, a sulfonylurea-induced block of K(ATP) channels shortens the delay of occurrence of a major Ca(i) rise in cerebellar Purkinje neurons. In neurons of the neonatal medullary respiratory network, K(ATP) channel blockers reverse the anoxic hyperpolarisation associated with slowing of respiratory frequency. This may constitute an adaptive mechanism for energy preservation. These studies demonstrate that K(ATP) channels are an ubiquituous feature of mammalian neurons and may, indeed, play a protective role in brain hypoxia. PMID- 15299040 TI - A unique pathway of cardiac myocyte death caused by hypoxia-acidosis. AB - Chronic hypoxia in the presence of high glucose leads to progressive acidosis of cardiac myocytes in culture. The condition parallels myocardial ischemia in vivo, where ischemic tissue becomes rapidly hypoxic and acidotic. Cardiac myocytes are resistant to chronic hypoxia at neutral pH but undergo extensive death when the extracellular pH (pH[o]) drops below 6.5. A microarray analysis of 20 000 genes (cDNAs and expressed sequence tags) screened with cDNAs from aerobic and hypoxic cardiac myocytes identified >100 genes that were induced by >2-fold and approximately 20 genes that were induced by >5-fold. One of the most strongly induced transcripts was identified as the gene encoding the pro-apoptotic Bcl-2 family member BNIP3. Northern and western blot analyses confirmed that BNIP3 was induced by 12-fold (mRNA) and 6-fold (protein) during 24 h of hypoxia. BNIP3 protein, but not the mRNA, accumulated 3.5-fold more rapidly under hypoxia acidosis. Cell fractionation experiments indicated that BNIP3 was loosely bound to mitochondria under conditions of neutral hypoxia but was translocated into the membrane when the myocytes were acidotic. Translocation of BNIP3 coincided with opening of the mitochondrial permeability pore (MPTP). Paradoxically, mitochondrial pore opening did not promote caspase activation, and broad-range caspase inhibitors do not block this cell death pathway. The pathway was blocked by antisense BNIP3 oligonucleotides and MPTP inhibitors. Therefore, cardiac myocyte death during hypoxia-acidosis involves two distinct steps: (1) hypoxia activates transcription of the death-promoting BNIP3 gene through a hypoxia inducible factor-1 (HIF-1) site in the promoter and (2) acidosis activates BNIP3 by promoting membrane translocation. This is an atypical programmed death pathway involving a combination of the features of apoptosis and necrosis. In this article, we will review the evidence for this unique pathway of cell death and discuss its relevance to ischemic heart disease. The article also contains new evidence that chronic hypoxia at neutral pH does not promote apoptosis or activate caspases in neonatal cardiac myocytes. PMID- 15299042 TI - Chaperones, protein aggregation, and brain protection from hypoxic/ischemic injury. AB - Chaperones, especially the stress inducible Hsp70, have been studied for their potential to protect the brain from ischemic injury. While they protect from both global and focal ischemia in vivo and cell culture models of ischemia/reperfusion injury in vitro, the mechanism of protection is not well understood. Protein aggregation is part of the etiology of chronic neurodegenerative diseases such as Huntington's and Alzheimer's, and recent data demonstrate protein aggregates in animal models of stroke. We now demonstrate that overexpression of Hsp70 in hippocampal CA1 neurons reduces evidence of protein aggregation under conditions where neuronal survival is increased. We have also demonstrated protection by the cochaperone Hdj-2 in vitro and demonstrated that this is associated with reduced protein aggregation identified by ubiquitin immunostaining. Hdj-2 can prevent protein aggregate formation by itself, but can only facilitate protein folding in conjunction with Hsp70. Pharmacological induction of Hsp70 was found to reduce both apoptotic and necrotic astrocyte death induced by glucose deprivation or oxygen glucose deprivation. Protection from ischemia and ischemia-like injury by chaperones thus involves at least anti-apoptotic, anti-necrotic and anti-protein aggregation mechanisms. PMID- 15299043 TI - Oxidants, antioxidants and the ischemic brain. AB - Despite numerous defenses, the brain is vulnerable to oxidative stress resulting from ischemia/reperfusion. Excitotoxic stimulation of superoxide and nitric oxide production leads to formation of highly reactive products, including peroxynitrite and hydroxyl radical, which are capable of damaging lipids, proteins and DNA. Use of transgenic mutants and selective pharmacological antioxidants has greatly increased understanding of the complex interplay between substrate deprivation and ischemic outcome. Recent evidence that reactive oxygen/nitrogen species play a critical role in initiation of apoptosis, mitochondrial permeability transition and poly(ADP-ribose) polymerase activation provides additional mechanisms for oxidative damage and new targets for post ischemic therapeutic intervention. Because oxidative stress involves multiple post-ischemic cascades leading to cell death, effective prevention/treatment of ischemic brain injury is likely to require intervention at multiple effect sites. PMID- 15299044 TI - Erythropoietin and the hypoxic brain. AB - Normal tissue function in mammals depends on adequate supply of oxygen through blood vessels. A discrepancy between oxygen supply and consumption (hypoxia) induces a variety of specific adaptation mechanisms at the cellular, local and systemic level. These mechanisms are in part governed by the activation of hypoxia-inducible transcription factors (HIF-1, HIF-2), which in turn modulate expression of hypoxically regulated genes such as those encoding vascular endothelial growth factor (VEGF) and erythropoietin (EPO). EPO is a glycoprotein that is produced mainly by interstitial fibroblasts in the kidneys of the adult and in hepatocytes in the foetus. Released into the circulation, EPO makes its way to the bone marrow, where it regulates red cell production by preventing apoptosis of erythroid progenitor cells. Recently, EPO has emerged as a multifunctional growth factor that plays a significant role in the nervous system. Both EPO and its receptor are expressed throughout the brain in glial cells, neurones and endothelial cells. Hypoxia and ischaemia have been recognised as important driving forces of EPO expression in the brain. EPO has potent neuroprotective properties in vivo and in vitro and appears to act in a dual way by directly protecting neurones from ischaemic damage and by stimulating endothelial cells and thus supporting the angiogenic effect of VEGF in the nervous system. Thus, hypoxia-induced gene products such as VEGF and EPO might be part of a self-regulated physiological protection mechanism to prevent neuronal injury, especially under conditions of chronically reduced blood flow (chronic ischaemia). In this review, I will briefly summarize the recent findings on the molecular mechanisms of hypoxia-regulated EPO expression in general and give an overview of its expression in the central nervous system, its action as a growth factor with non-haematopoietic functions and its potential clinical relevance in various brain pathologies. PMID- 15299045 TI - Clinical perspectives: neuroprotection lessons from hypoxia-tolerant organisms. AB - An effective treatment for brain ischemia is a pressing medical need. Research on brain ischemia has largely focused on understanding the mechanisms of neuron death as a way of identifying targets for therapy. An attractive alternative approach is to identify the survival strategies of hypoxia-tolerant neurons. The adaptation of vertebrate neurons to hypoxia occurs in at least three major ways: (1) as a constitutive property of neurons in anoxia-tolerant turtles and fish, (2) as a property of intra-uterine and early post-natal mammalian development, and (3) as part of a slower, chronic process, as in acclimitization to high altitude. Research on hypoxia-tolerant neurons has already revised several earlier concepts, including the role of calcium in cell death and survival, and the value of N-methyl-d-aspartate (NMDA) receptor antagonism. A broad and fundamental understanding of how neurons adapt to hypoxia is likely to help guide efforts to find new treatments for brain hypoxia and ischemia. PMID- 15299047 TI - Quantification of 18F-FDG uptake in non-small cell lung cancer: a feasible prognostic marker? PMID- 15299048 TI - PET imaging of GRP receptor expression in prostate cancer. PMID- 15299049 TI - Retrospective digital image fusion of multidetector CT and 18F-FDG PET: clinical value in pancreatic lesions--a prospective study with 104 patients. AB - Differential diagnosis of pancreatic lesions still remains a problem. Whereas CT provides high spatial resolution, PET detects malignant lesions with high sensitivity. The objective of this study was to evaluate the clinical benefit of PET/CT image fusion in the diagnostic workup of pancreatic cancer. METHODS: One hundred four patients with suspected pancreatic lesion underwent triple-phase multidetector CT and (18)F-FDG PET scanning. Voxel-based retrospective registration and fusion of CT and PET were performed with recently developed software. CT, PET, and fused images were assessed by 2 radiologists with regard to the detection of malignancies, possible infiltration of adjacent tissue or lymph nodes, or distant metastases. RESULTS: Fusion of CT and PET images was technically successful in 96.2%. In 2 cases, paraaortic lymph node infiltration was detected only by image fusion; in a further 8 cases, lymph node metastases were confirmed with improved localization. In 5 patients, additional pancreatic tumors or distant metastases only suspected during PET scanning were confirmed. Image fusion improved the sensitivity of malignancy detection from 76.6% (CT) and 84.4% (PET) to 89.1% (image fusion). Compared with CT alone, image fusion increased the sensitivity of detecting tissue infiltration to 68.2%, but at the cost of decreased specificity. CONCLUSION: The most important supplementary finding supplied by image fusion is a more precise correlation with focal tracer hot spots in PET. Image fusion improved the sensitivity of differentiating between benign and malignant pancreatic lesions with no significant change in specificity. All image modalities failed to stage lymph node involvement. PMID- 15299050 TI - The CT motion quantitation of lung lesions and its impact on PET-measured SUVs. AB - We previously reported that respiratory motion is a major source of error in quantitation of lesion activity using combined PET/CT units. CT acquisition of the lesion occurs in seconds, rather than the 4-6 min required for PET emission scans. Therefore, an incongruent lesion position during CT acquisition will bias activity estimates using PET. In this study, we systematically analyzed the range of activity concentration changes, hence SUV, for lung lesions. METHODS: Five lung cancer patients were scanned with PET/CT. In CT, data were acquired in correlation with the real-time positioning. CT images were acquired, in cine mode, at 0.45-s intervals for slightly longer (1 s) than a full respiratory cycle at each couch position. Other scanning parameters were a 0.5-s gantry rotation, 140 kVp, 175 mA, 10-mm couch increments, and a 2.5-mm slice thickness. PET data were acquired after intravenous injection of about 444-555 MBq of (18)F-FDG with a 1-h uptake period. The scanning time was 3 min per bed position for PET. Regularity in breathing was assisted by audio coaching. A commercial software program was then used to sort the acquired CT images into 10 phases, with 0% corresponding to end of inspiration (EI) and 50% corresponding to end of expiration (EE). Using the respiration-correlated CT data, images were rebinned to match the PET slice locations and thickness. RESULTS: We analyzed 8 lesions from 5 patients. Reconstructed PET emission data showed up to a 24% variation in the lesion maximum standardized uptake values (SUVs) between EI and EE phases. Examination of all the phases showed an SUV variation of up to 30%. Also, in some cases the lesion showed up to a 9-mm shift in location and up to a 21% reduction in size when measured from PET during the EI phase, compared with during the EE phase. CONCLUSION: Using respiration-correlated CT for attenuation correction, we were able to quantitate the fluctuations in PET SUVs. Because those changes may lead to estimates of lower SUVs, the respiratory phase during CT transmission scanning needs to be measured or lung motion has to be regulated for imaging lung cancer in routine clinical practice. PMID- 15299051 TI - Comparison of 18F-FDG and 11C-methionine for PET-guided stereotactic brain biopsy of gliomas. AB - We compared the contributions of the labeled tracers (11)C-methionine (Met) and (18)F-FDG for PET-guided stereotactic biopsy of brain gliomas. METHODS: In 32 patients with glioma, stereotactic Met PET and (18)F-FDG PET were integrated in the planning of stereotactic brain biopsy. PET images were analyzed to determine which tracer offered the best information for target definition. The stereotactic coregistration of PET images allowed accurate comparison of the level, distribution, and extent of uptake for both tracers according to tumor location and grade. RESULTS: A histologic diagnosis was obtained for all patients. All gliomas had an area of abnormal Met uptake, and 27 showed abnormal (18)F-FDG uptake. (18)F-FDG was used for target selection when its uptake was higher in tumor than in gray matter (14 gliomas). Seven were in the basal ganglia or brain stem. Met was used for target selection when there was no (18)F-FDG uptake or when (18)F-FDG uptake was equivalent to that in the gray matter (18 gliomas). Thirteen were in the cortex. Sixty-one of the 70 stereotactic trajectories obtained from the 32 patients were based on PET-defined targets and had an area of abnormal Met uptake. These 61 Met-positive trajectories always yielded a diagnosis of tumor. All nondiagnostic trajectories (n = 9) were obtained in areas with no increased uptake of Met. In all patients with increased uptake of both tracers, the focus of highest Met uptake corresponded to the focus of highest (18)F-FDG uptake. However, the extent of uptake of both tracers was variable. CONCLUSION: Distributions of highest Met and (18)F-FDG uptake are similar in brain gliomas. Because Met provides a more sensitive signal, it is the molecule of choice for single-tracer PET-guided neurosurgical procedures in gliomas. PMID- 15299052 TI - Perfusable tissue index as a potential marker of fibrosis in patients with idiopathic dilated cardiomyopathy. AB - A varying degree of interstitial and perivascular fibrosis is a common finding in idiopathic dilated cardiomyopathy (DCM). The perfusable tissue index (PTI), obtained with PET, is a noninvasive tool for assessing myocardial fibrosis on a regional level. Measurements of the PTI in DCM, however, have not been performed yet. This study was undertaken to test the hypothesis that the PTI is reduced in patients with DCM. METHODS: Fifteen patients with an advanced stage of DCM (New York Heart Association class III or IV and left ventricular ejection fraction [LVEF] < 35%) and 11 healthy control subjects were studied. PET was performed using H(2)(15)O and C(15)O to obtain the perfusable tissue fraction (PTF) and the anatomic tissue fraction (ATF), respectively. RESULTS: The PTI (=PTF/ATF) was reduced in DCM compared with control subjects (0.91 +/- 0.12 vs. 1.12 +/- 0.10; P < 0.01). Heterogeneity of the PTI, expressed as the coefficient of variation, was increased in DCM versus that of healthy control subjects (0.18 +/- 0.07 vs. 0.13 +/- 0.06; P < 0.05). There was no correlation between the PTI and echocardiographically derived LVEF in both groups. CONCLUSION: The PTI was reduced in patients with an advanced stage of DCM. Interstitial and perivascular fibrosis may be responsible for this reduction. Furthermore, the degree of the PTI reduction was variable in DCM patients, both on a regional level and between patients. Noninvasive assessment of fibrosis with the PTI offers the opportunity to evaluate the effects of fibrosis on regional myocardial function, correlate fibrosis with prognosis, and monitor pharmaceutical intervention. PMID- 15299053 TI - 123I-Metaiodobenzylguanidine myocardial scintigraphy in panic disorder. AB - Although autonomic function has been investigated in panic disorder, previous studies have not yet revealed a consistent autonomic change in this disease. The purpose of this study was to evaluate the cardiac sympathetic function in panic disorder using (123)I-metaiodobenzylguanidine ((123)I-MIBG). METHODS: Myocardial imaging using (123)I-MIBG was performed on 9 patients with panic disorder (7 men, 2 women; mean age, 37.4 +/- 13.2 y) and 11 control subjects (11 men; mean age, 37.6 +/- 9.3 y). Early (30 min) and delayed (4 h) planar images were taken after the injection of 111 MBq (123)I-MIBG. The mean counts in the whole heart and the mediastinum were obtained from the early and delayed images to calculate the heart-to-mediastinum count ratios (H/M ratios) and the myocardial washout rate. RESULTS: The (123)I-MIBG H/M ratios of the patients with panic disorder were 1.80 +/- 0.16 for the early images and 1.86 +/- 0.30 for the delayed images, which were significantly lower than those of the control subjects (2.15 +/- 0.15 [P = 0.001] and 2.26 +/- 0.21 [P = 0.009], respectively). The (123)I-MIBG washout rate from the heart in the patients with panic disorder (33.8% +/- 6.9%) was significantly higher than that in the control subjects (27.8% +/- 3.5%) (P = 0.02). CONCLUSION: (123)I-MIBG myocardial scintigraphy demonstrated impairment of cardiac sympathetic function in panic disorder. The results suggest that (123)I MIBG imaging could become a useful tool for analyzing the pathophysiology of panic disorder. PMID- 15299054 TI - Reducing bladder artifacts in clinical pelvic SPECT images. AB - SPECT imaging of the pelvis is hampered by the presence of bladder artifacts, which render up to 20% of the images unreadable. The artifacts are caused by the high level of activity in the bladder and by the change in activity level as the bladder fills during data acquisition. The changing activity, together with the inhomogeneous attenuation of the pelvis, leads to inconsistencies in the projections and consequently artifacts when the data are reconstructed with filtered backprojection (FBP). dSPECT is an iterative algorithm that permits the reconstruction of dynamic SPECT images from a single, slow-rotation SPECT data acquisition. The reconstruction algorithm incorporates attenuation correction (AC) and changing tracer distributions and has been shown to reduce bladder artifacts in simulated data. In this study, we showed that dSPECT is effective at removing bladder artifacts from clinically acquired pelvic bone SPECT images. METHODS: Data from 20 patient volunteers were reconstructed using FBP, rescaled block-iterative reconstruction (RBI) without AC, RBI with AC, and dSPECT. AC was based on patient-specific attenuation maps acquired with a (153)Gd scanning line source transmission system. For dSPECT, 16 time frames (4 projections/head/frame) were reconstructed and then summed to produce the final image. Artifact-to-bone contrast was compared, and image quality was subjectively assessed. RESULTS: Compared with FBP, RBI without AC significantly reduced (P = 0.008) the streak artifact. Both dSPECT and RBI with AC further significantly reduced (P < 0.001) the streak artifact and also improved the uniformity and symmetry of bone tracer uptake. RBI with AC and dSPECT produced equivalent images if the change in bladder activity during acquisition was modest; however, with large changes in the activity (>100%), RBI with AC did not completely remove the artifact. In that situation, dSPECT produced additional reductions in streak-to-bone contrast. CONCLUSION: Of the methods considered, dSPECT is the most effective at removing bladder artifacts in clinical pelvic SPECT. PMID- 15299055 TI - Radioimmunoscintigraphy for postprostatectomy radiotherapy: analysis of toxicity and biochemical control. AB - Our goal was to evaluate the role of radioimmunoscintigraphy (RIS) directed against prostate-specific membrane antigen (PSMA) in influencing postprostatectomy radiotherapy (RT) toxicity and biochemical control. METHODS: The records of 107 postprostatectomy RT patients were reviewed. The group for whom no RIS scan was obtained (group A, n = 54) was identified as was the group for whom a RIS scan was obtained (group B, n = 53). Group B was further subdivided into those who had a RIS and CT-scan correlation to aid in treatment planning (subgroup B1, n = 40) versus those who did not (subgroup B2, n = 13). Gastrointestinal (GI) and genitourinary (GU) toxicities were reviewed for each of these groups and subgroups and compared. Biochemical failures (defined as 2 successive PSA rises after a nadir of >or=0.2 ng/mL) were identified to generate biochemical failure-free survival (BFFS) curves for each of the groups and subgroups. RESULTS: No significant differences in late toxicity were observed between any group or subgroup. However, acute GI toxicity was higher in group B versus group A (P = 0.026), and acute GU toxicity was higher in subgroup B2 versus subgroup B1 (P = 0.050). Overall, most toxicity was grade 1 or 2; only one case of grade 3 toxicity and no cases of grade 4 or 5 toxicity were observed. Three-year BFFS was higher for group B versus group A (80.7% vs. 75.5%) and for subgroup B1 versus subgroup B2 (84.5% vs. 71.6%). On multivariate analysis of pretreatment (age, race), surgical/staging (stage, grade, margin status, extracapsular extension, lymph node status, seminal vesicle invasion, post radical retropubic prostatectomy [RRP] prostate-specific antigen [PSA] nadir, maximum post-RRP PSA, and RRP-to-RT interval), and treatment (hormone therapy, RT dose, RT technique, RIS scan, and RIS/CT correlation) factors on BFFS, the only covariate reaching significance was RIS/CT correlation (P = 0.042). CONCLUSION: A small BFFS advantage was observed in patients for whom RIS was used to guide RT decision making and treatment planning; however, this advantage only reached significance in this study for those for whom the RIS/CT correlation was used to guide target definition. The improved PSA control using RIS was achieved with a small increase in acute toxicity but with no observed change in late toxicity. These findings can serve as the basis for prospective studies in this area of investigation. PMID- 15299056 TI - Is 18F-FDG PET more accurate than standard diagnostic procedures in the detection of suspected recurrent melanoma? AB - The aim of this study was to determine the accuracy of (18)F-FDG PET in detecting recurrent melanoma. METHODS: PET findings were compared with those obtained by standard diagnostic clinical procedures (CP) to establish the role of PET in the management of patients with melanoma. From 156 patients with confirmed melanoma and recurrence suspected by clinical examination, 184 PET scans were retrospectively reviewed. Histology or clinical follow-up was used for final diagnosis. RESULTS: The sensitivity and specificity of PET for detecting lesions on an individual-patient basis were 74% and 86%, respectively, compared with respective values of 58% and 45% for CP alone. The overall accuracy for PET was 81%, compared with 52% for other methods. PET was more accurate (91% vs. 67%) than CP in detecting locoregional disease and distant metastases (85% vs. 55%), and PET results led to a change in the planned clinical management of 36% of patients included in this study. PET was more accurate than CT in detecting skin lesions, malignant lymph nodes, and metastases to the abdomen, liver, and bone. In the assessment of pulmonary disease, PET showed higher specificity (92% vs. 70%) than CT for the detection of lung parenchyma lesions; however, the sensitivity was better for CT (93%) than for PET (57%). CONCLUSION: PET is better than CP in detecting locoregional disease and distant metastases in all sites except the lung, where it appears to be a useful adjunct to CT. The use of PET as a routine clinical tool can lead to a substantial change in the clinical management of suspected recurrent melanoma. PMID- 15299057 TI - Simplified kinetic analysis of tumor 18F-FDG uptake: a dynamic approach. AB - Standardized uptake value (SUV) is often used to quantify (18)F-FDG tumor use. Although useful, SUV suffers from known quantitative inaccuracies. Simplified kinetic analysis (SKA) methods have been proposed to overcome the shortcomings of SUV. Most SKA methods rely on a single time point (SKA-S), not on tumor uptake rate. We describe a hybrid between Patlak analysis and existing SKA-S methods, using multiple time points (SKA-M) but reduced imaging time and without measurement of an input function. We compared SKA-M with a published SKA-S method and with Patlak analysis. METHODS: Twenty-seven dynamic (18)F-FDG scans were performed on 11 cancer patients. A population-based (18)F-FDG input function was generated from an independent patient population. SKA-M was calculated using this population input function and either a short, late, dynamic acquisition over the tumor (starting 25-35 min after injection and ending approximately 55 min after injection) or dynamic imaging 10 or 25 min to approximately 55 min after injection but using only every second or third time point, to permit a 2- or 3 field-of-view study. SKA-S was also calculated. Both SKA-M and SKA-S were compared with the gold standard, Patlak analysis. RESULTS: Both SKA-M (1 field of view) and SKA-S correlated well with Patlak slope (r > 0.99, P < 0.001, and r = 0.96, P < 0.001, respectively), as did multilevel SKA-M (r > 0.99 and P < 0.001 for both). Mean values of SKA-M (25-min start time) and SKA-S were statistically different from Patlak analysis (P < 0.001 and P < 0.04, respectively). One-level SKA-M differed from the Patlak influx constant by only -1.0% +/- 1.4%, whereas SKA-S differed by 15.1% +/- 3.9%. With 1-level SKA-M, only 2 of 27 studies differed from K(i) by more than 20%, whereas with SKA-S, 10 of 27 studies differed by more than 20% from K(i). CONCLUSION: Both SKA-M and SKA-S compared well with Patlak analysis. SKA-M (1 or multiple levels) had lower variability and bias than did SKA-S, compared with Patlak analysis. SKA-M may be preferred over SUV or SKA-S when a large unmetabolized (18)F-FDG fraction is expected and 1-3 fields of view are sufficient. PMID- 15299058 TI - In vitro proton magnetic resonance spectroscopic lactate and choline measurements, 18F-FDG uptake, and prognosis in patients with lung adenocarcinoma. AB - It has been reported that (18)F-FDG uptake, lactate concentration, and choline concentration are good indicators of malignant grade in several different kinds of tumors. In this study, we investigated the correlation between (18)F-FDG uptake in (18)F-FDG PET imaging, lactate concentration and choline concentration measured by in vitro (1)H magnetic resonance spectroscopy (MRS), and survival probabilities in human lung adenocarcinoma. METHODS: Nineteen patients with lung adenocarcinoma underwent (18)F-FDG PET before surgery. The (1)H MRS spectra were obtained in vitro from methanol-chloroform-water extracts of lung adenocarcinomas and normal lungs. The ratios of the lactate (R(lac)) or choline (R(cho)) concentration of lung adenocarcinoma to normal lung from the same patient were correlated with the mean standardized uptake value (SUV). The Kaplan-Meier life table method was used to analyze the relationship between (18)F-FDG uptake, R(lac), R(cho), and patient survival probabilities. RESULTS: There was no significant correlation between mean SUV and R(lac) or R(cho) in patients with lung adenocarcinoma. An SUV > 5 means poorer survival probabilities in patients with lung adenocarcinoma (P = 0.004). A higher R(lac) probably indicates a trend for patients with lung adenocarcinoma to have poorer survival probabilities; however, R(cho) is not an indicator of survival probability. (18)F-FDG uptake significantly correlated with cell differentiation (P = 0.007), whereas R(lac) and R(cho) had no correlation with it. CONCLUSION: No significant correlation was found between SUV and R(lac) or R(cho) in patients with lung adenocarcinoma. Compared with R(lac) and R(cho) measured by in vitro MRS, (18)F-FDG uptake is a better indicator of prognosis in patients with lung adenocarcinoma. PMID- 15299059 TI - Characterization of the normal adrenal gland with 18F-FDG PET/CT. AB - Prior studies have documented increased (18)F-FDG adrenal activity in both benign and malignant pathologic conditions. When whole-body PET imaging is performed without CT anatomic coregistration, however, the normal adrenal gland is difficult to recognize. The purpose of this study was to investigate the normal adrenal appearance and standardized uptake value (SUV) using (18)F-FDG PET/CT imaging. METHODS: Twenty patients with lymphoma with normal-appearing adrenal glands on prior CT examination (less than a 5% pretest likelihood of adrenal involvement) were studied. PET/CT imaging was performed 2 h after intravenous administration of (18)F-FDG. Unenhanced CT scans were acquired for attenuation correction and anatomic coregistration. PET images were reconstructed using an ordered-subsets expectation maximization algorithm and were corrected for body weight, dose, and radioactive decay. Ability to confirm visualization of the adrenal glands was determined for (18)F-FDG PET alone and for (18)F-FDG PET/CT by a consensus of 2 readers, and uptake of (18)F-FDG in the adrenal gland was compared with liver activity and scored visually (0 = no visualization, 1 = activity less than in liver, 2 = activity equal to liver activity, and 3 = activity greater than in liver). RESULTS: The 2 readers agreed on visualization of the adrenal glands with PET alone for 2 of 40 (5%) glands. With PET/CT, the readers agreed on visualization of 27 of 40 (68%) adrenal glands. Visual scores for normal adrenal activity ranged from 0 to 3, and maximum SUVs ranged from 0.95 to 2.46. Visual scoring of adrenal activity correlated well with both mean and maximal SUV (mean SUV vs. visual score: slope = 0.96, r = 0.88; maximum SUV vs. visual score: slope = 0.99, r = 0.87). CONCLUSION: PET/CT permits more reliable visualization of normal adrenal glands than does PET alone. Visual assessment of adrenal uptake correlates well with SUV measurement, and readers of PET/CT need to be aware of the wide range of normal adrenal uptake. PMID- 15299060 TI - Constant-infusion H(2)15O PET and acetazolamide challenge in the assessment of cerebral perfusion status. AB - Assessing the baseline perfusion and perfusion reserve after acetazolamide (ACZ) challenge is a common method for the evaluation of patients with cerebrovascular disease. Most previous studies using H(2)(15)O PET applied the bolus injection technique. There is considerable discrepancy regarding the optimal time point of imaging after ACZ injection. The purpose of this study was to continuously monitor cerebral blood flow (CBF) after ACZ using constant-infusion H(2)(15)O PET. METHODS: Four patients with stenoses of an internal carotid artery and 6 with moyamoya disease were studied. H(2)(15)O was continuously infused, and data were recorded in 1-min frames. After equilibration of H(2)(15)O, 5 min of baseline data were acquired, and then 1 g of ACZ was administered intravenously and data collection continued for 10-22 min. Arterial blood was continuously drawn for absolute quantification of CBF. RESULTS: The arterial (15)O concentration remained generally stable during scanning, and the cerebellar blood flow fluctuations of the 5 baseline scans were small. The scan-to-scan difference was 6% (difference of 2 successive scans/mean). In the nonpathologic areas, the increase in CBF started 1-2 min after administration of ACZ. The largest fraction of the increase occurred from 0 to 10 min. The ratio of CBF in pathologic areas to CBF in cerebellum showed an initial decrease that stabilized after 5 min. CONCLUSION: A continuous-infusion protocol is a viable alternative to single bolus injections for the assessment of cerebral perfusion status. Such a protocol is advantageous when the time course of CBF after an intervention is not known. With continuous monitoring, the optimal time point for evaluation of a certain parameter can be chosen post hoc. Furthermore, the time course of CBF itself may allow the definition of new parameters for evaluating perfusion status in cerebrovascular patients, both for assessment before a revascularization procedure and for follow-up. A limitation of the present study is the relatively small number of patients with each type of cerebrovascular disease and the lack of healthy subjects. PMID- 15299061 TI - Gamma-camera 18F-FDG PET in diagnosis and staging of patients presenting with suspected lung cancer and comparison with dedicated PET. AB - It is not clear whether high-quality coincidence gamma-PET (gPET) cameras can provide clinical data comparable with data obtained with dedicated PET (dPET) cameras in the primary diagnostic work-up of patients with suspected lung cancer. This study focuses on 2 main issues: direct comparison between foci resolved with the 2 different PET scanners and the diagnostic accuracy compared with final diagnosis determined by the combined information from all other investigations and clinical follow-up. METHODS: Eighty-six patients were recruited to this study through a routine diagnostic program. They all had changes on their chest radiographs, suggesting malignant lung tumor. In addition to the standard diagnostic program, each patient had 2 PET scans that were performed on the same day. After administration of 419 MBq (range = 305-547 MBq) (18)F-FDG, patients were scanned in a dedicated PET scanner about 1 h after FDG administration and in a dual-head coincidence gamma-camera about 3 h after tracer injection. Images from the 2 scans were evaluated in a blinded set-up and compared with the final outcome. RESULTS: Malignant intrathoracic disease was found in 52 patients, and 47 patients had primary lung cancers. dPET detected all patients as having malignancies (sensitivity, 100%; specificity, 50%), whereas gPET missed one patient (sensitivity, 98%; specificity, 56%). For evaluating regional lymph node involvement, sensitivity and specificity rates were 78% and 84% for dPET and 61% and 90% for gPET, respectively. When comparing the 2 PET techniques with clinical tumor stage (TNM), full agreement was obtained in 64% of the patients (Cohen's kappa = 0.56). Comparing categorization of the patients into clinical relevant stages (no malignancy/malignancy suitable for treatment with curative intent/nontreatable malignancy), resulted in full agreement in 81% (Cohen's kappa = 0.71) of patients. CONCLUSION: Comparing results from a recent generation of gPET cameras obtained about 2 h later than those of dPET, there was a fairly good agreement with regard to detecting primary lung tumors but slightly reduced sensitivity in detecting smaller malignant lesions such as lymph nodes. Depending on the population to be investigated, and if dPET is not available, gPET might provide significant diagnostic information in patients in whom lung cancer is suspected. PMID- 15299062 TI - Radiopharmaceutical therapy for palliation of bone pain from osseous metastases. AB - Bone metastasis occurs as a result of a complex pathophysiologic process between host and tumor cells leading to cellular invasion, migration adhesion, and stimulation of osteoclastic and osteoblastic activity. The process is mediated by parathyroid hormones, cytokines, and tumor-derived factors. Several sequelae occur as a result of osseous metastases and resulting bone pain can lead to significant debilitation. Pain associated with osseous metastasis is thought to be distinct from neuropathic or inflammatory pain. Several mechanisms-such as invasion of tumor cells, spinal cord astrogliosis, and sensitization of nervous system-have been postulated to cause pain. Pharmaceutical therapy of bone pain includes nonsteroidal analgesics and opiates. These drugs are associated with side effects, and tolerance to these agents necessitates treatment with other modalities. Bisphosphonates act by inhibiting osteoclast-mediated resorption and have been increasingly used in treatment of painful bone metastasis. While external beam radiation therapy remains the mainstay of pain palliation of solitary lesions, bone-seeking radiopharmaceuticals have entered the therapeutic armamentarium for the treatment of multiple painful osseous lesions. (32)P has been used for >3 decades in the treatment of multiple osseous metastases. The myelosuppression caused by this agent has led to the development of other bone seeking radiopharmaceuticals, including (89)SrCl, (153)Sm ethylenediaminetetramethylene phosphonic acid ((153)Sm-EDTMP), (179m)SnCl, and (166)Ho-Labeled 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetramethylenephosphonate ((166)Ho-DOTMP). (89)Sr is a bone-seeking radionuclide, whereas (153)Sm-EDTMP is a bone-seeking tetraphosphonate; both have been approved by the Food and Drug Administration for the treatment of painful osseous metastases. While both agents have been shown to have efficacy in the treatment of painful osseous metastases from prostate cancer, they may also have utility in the treatment of painful osseous metastases from breast cancer and perhaps from non-small cell lung cancer. This article illustrates the salient features of these radiopharmaceuticals, including the approved dose, method of administration, and indications for use. We conclude with recommended guidelines for therapy and follow-up. PMID- 15299063 TI - Patient-specific dosimetry for 131I thyroid cancer therapy using 124I PET and 3 dimensional-internal dosimetry (3D-ID) software. AB - Compared with conventional, whole-organ, S-factor-based dosimetry, 3-dimensional (3D), patient-specific dosimetry better accounts for radionuclide distribution and anatomic patient variability. Its accuracy, however, is limited by the quality of the cumulated activity information that is provided as input. This input has typically been obtained from SPECT and planar imaging studies. The objective was to implement and evaluate PET-based, patient-specific, 3D dosimetry for thyroid cancer patients. METHODS: Three to 4 PET imaging studies were obtained over a 7-d period in 15 patients with metastatic thyroid carcinoma after administration of (124)I-NaI. Subsequently, patients were treated with (131)I on the basis of established clinical parameters. Retrospective dosimetry was performed using registered (124)I PET images that were corrected for the half life difference between (124)I and (131)I. A voxel-by-voxel integration, over time, of the resulting (131)I-equivalent PET-derived images was performed to provide a single 3D dataset representing the spatial distribution of cumulated activity values for each patient. Image manipulation and registration were performed using Multiple Image Analysis Utility (MIAU), a software package developed previously. The software package, 3D-Internal Dosimetry (3D-ID), was used to obtain absorbed dose maps from the cumulated activity image sets. RESULTS: Spatial distributions of absorbed dose, isodose contours, dose-volume histograms (DVHs), and mean absorbed dose estimates were obtained for a total of 56 tumors. Mean absorbed dose values for individual tumors ranged from 1.2 to 540 Gy. The absorbed dose distribution within individual tumors was widely distributed ranging from a minimum of 0.3 to a maximum of 4,000 Gy. CONCLUSION: (124)I PET-based, patient-specific 3D dosimetry is feasible, and sequential PET can be used to obtain cumulated activity images for 3D dosimetry. PMID- 15299064 TI - Tumor imaging using a standardized radiolabeled adapter protein docked to vascular endothelial growth factor. AB - Direct radiolabeling of proteins can result in the loss of targeting activity, requires highly customized procedures, and yields heterogeneous products. Here we describe a novel imaging complex comprised of a standardized (99m)Tc-radiolabeled adapter protein noncovalently bound to a "Docking tag" fused to a "Targeting protein". The assembly of this complex is based on interactions between human 109 amino acid (HuS) and 15-amino acid (Hu-tag) fragments of ribonuclease I, which serve as an "Adapter protein" and a Docking tag, respectively. METHODS: HuS modified with hydrazinonicotinamide (HYNIC) was radiolabeled using (99m)Tc tricine to a specific activity of 3.4-7.4 MBq/microg. Protein complexes were then formed by mixing (99m)Tc-HuS with equimolar amounts of either Hu-tagged VEGF(121) (Hu-VEGF [vascular endothelial growth factor]) or Hu-tagged anti-VEGFR-2 single chain antibody (Hu-P4G7) and incubating on ice for 15 min. 4T1 luc/gfp luciferase expressing murine mammary adenocarcinoma cells (1 x 10(4)) were implanted subcutaneously or injected intravenously into BALB/c mice. Bioluminescent imaging (BLI) was performed 10 d later. Immediately after BLI visualization of tumor, 18.5-37 MBq of tracer (5-10 microg of protein) were injected via tail vein. One hour later planar or SPECT images were obtained, followed by killing the mice. RESULTS: There was significantly (P = 0.0128) increased uptake of (99m)Tc-HuS/Hu VEGF (n = 10) within subcutaneous tumor as compared with (99m)Tc-HuS/Hu-P4G7 (n = 5) at biodistribution assay (2.68 +/- 0.75 vs. 1.8 +/- 0.21; tumor-to subcutaneous tissue [ratio of specific activities], respectively), despite similar molecular weights. The focal (99m)Tc-HuS/Hu-VEGF uptake seen on planar images (3.44 +/- 1.16 [tumor to soft-tissue background]) corresponded directly to the locations of tumor observed by BLI. Region of interest analyses of SPECT images revealed a significant increase of (99m)Tc-HuS/Hu-VEGF (n = 5) within the lungs with BLI-detectable pulmonary tumor nodules as compared with controls (n = 4) (right: 4.47 +/- 2.07 vs. 1.79 +/- 0.56; left: 3.66 +/- 1.65 vs. 1.62 +/- 0.45, tumor lung [counts/pixel]/normal lung [counts/pixel], respectively). CONCLUSION: (99m)Tc-HuS/Hu-VEGF complex is stable for at least 1 h in vivo and can be effectively used to image mouse tumor neovasculature in lesions as small as several millimeters in soft tissue. We expect that a similar approach can be adapted for in vivo delivery of other targeting proteins of interest without affecting their bioactivity. PMID- 15299066 TI - microPET and autoradiographic imaging of GRP receptor expression with 64Cu-DOTA [Lys3]bombesin in human prostate adenocarcinoma xenografts. AB - Overexpression of gastrin-releasing peptide (GRP) receptor (GRPR) in both androgen-dependent (AD) and androgen-independent (AI) human neoplastic prostate tissues provides an attractive target for prostate cancer imaging and therapy. The goal of this study was to develop (64)Cu-radiolabeled GRP analogs for PET imaging of GRPR expression in prostate cancer xenografted mice. METHODS: [Lys(3)]bombesin ([Lys(3)]BBN) was conjugated with 1,4,7,10-tetraazadodecane N,N',N",N"'-tetraacetic acid (DOTA) and labeled with the positron-emitting isotope (64)Cu (half-life = 12.8 h, 19% beta(+)). Receptor binding of DOTA [Lys(3)]BBN and internalization of (64)Cu-DOTA-[Lys(3)]BBN by PC-3 prostate cancer cells were measured. Tissue biodistribution, microPET, and whole-body autoradiographic imaging of the radiotracer were also investigated in PC-3 (AI)/CRW22 (AD) prostate cancer tumor models. RESULTS: A competitive receptor- binding assay using (125)I-[Tyr(4)]BBN against PC-3 cells yielded a 50% inhibitory concentration value of 2.2 +/- 0.5 nmol/L for DOTA-[Lys(3)]BBN. Incubation of cells with the (64)Cu-labeled radiotracer showed temperature- and time-dependent internalization. At 37 degrees C, >60% of the tracer was internalized within the first 15 min and uptake remained constant for 2 h. Radiotracer uptake was higher in AI PC-3 tumor (5.62 +/- 0.08 %ID/g at 30 min after injection, where %ID/g is the percentage of injected dose per gram) than in AD CWR22 tumor (1.75 +/- 0.05 %ID/g at 30 min after injection). Significant accumulation of the activity in GRPR-positive pancreas was also observed (10.4 +/ 0.15 %ID/g at 30 min after injection). Coinjection of a blocking dose of [Lys(3)]BBN inhibited the activity accumulation in PC-3 tumor and pancreas but not in CWR22 tumor. microPET and autoradiographic imaging of (64)Cu-DOTA [Lys(3)]BBN in athymic nude mice bearing subcutaneous PC-3 and CWR22 tumors showed strong tumor-to-background contrast. CONCLUSION: This study demonstrates that PET imaging of (64)Cu-DOTA-[Lys(3)]BBN is able to detect GRPR-positive prostate cancer. PMID- 15299065 TI - PET imaging of oncogene overexpression using 64Cu-vasoactive intestinal peptide (VIP) analog: comparison with 99mTc-VIP analog. AB - The purpose of this study was to assess the feasibility of PET imaging of oncogene VPAC1 receptors overexpressed in human breast cancer cells. METHODS: Vasoactive intestinal peptide (VIP) analog (TP3982) was synthesized to harbor a carboxy-terminus lysine (Lys) residue separated from VIP-asparagine (Asn(28)) by 4-aminobutyric acid (Aba) as a spacer. Lys was derivatized with diaminopropionic acid coupled to a pair of dibenzoylthioglycolic acid residues as protecting groups. The analog was labeled with (64)Cu at pH 9 ((64)Cu-TP3982) and (99m)Tc at pH 12 ((99m)Tc-TP3982). (99m)Tc-TP3982 and VIP derivatized with Aba-GAGG and labeled with (99m)Tc ((99m)Tc-TP3654) were used as reference agents. Smooth muscle relaxivity assays performed with each derivative and compared with unaltered VIP(28) demonstrated functional integrity. In vitro stability of (64)Cu TP3982 was determined by challenging the complex with 100-mol excess of diethylenetriaminepentaacetic acid (DTPA), human serum albumin (HSA), and cysteine. In vivo stability was determined in urine and serum for up to 24 h. The mass of the Cu-TP3982 complex was determined by mass spectrometry. Human T47D breast tumor xenografts were grown in athymic nude mice. Planar scintigraphic imaging was performed at 4 and 24 h after the intravenous administration of (99m)Tc-TP3982 and (99m)Tc-TP3654 and PET imaging was performed using a small animal MOSAIC PET scanner, also at 4 and 24 h after injection of (64)Cu-TP3982. Tissue-distribution studies were also performed. In a separate experiment, receptors were blocked by intravenous injection of authentic VIP(28) 30 min before the administration of (64)Cu-TP3982 and tissue distribution was examined. RESULTS: (64)Cu-TP3982 labeling yields were 98% +/- 1.2% and those for (99m)Tc TP3982 and (99m)Tc-TP3654 were 98.2% +/- 1.1% and 97% +/- 1.6%, respectively. The biologic activity of both VIP analogs was uncompromised. When (64)Cu-TP3982 was challenged with 100-mol excess of DTPA, HSA, or cysteine, >98% radioactivity remained as (64)Cu-TP3982. In vivo, >98% of (64)Cu circulating in plasma remained as (64)Cu-TP3982. Of the (64)Cu excreted in urine 4, 20, and 24 h after injection, >98%, 89.9% +/- 0.9%, and 85% +/- 3%, respectively, were bound to TP3982. The mass of Cu-TP3982 as determined by surface-enhanced laser desorption/ionization time of flight (SELDI-TOF) was 4,049.7 Da. Four hours after receptor blocking with VIP(28), there was a significant reduction in uptake of all tissues except in the liver. With (64)Cu-TP3982, the 4-h postinjection tumor uptake was 10.8 +/- 2.1 %ID/g versus 0.5 +/- 0.02 %ID/g and 0.24 +/- 0.08 %ID/g for (99m)Tc-TP3982 and (99m)Tc-TP3654, respectively. Twenty-four hours after injection, the corresponding numbers were 17 +/- 0.7 %ID/g, 0.77 +/- 0.1 %ID/g, and 0.23 +/- 0.1 %ID/g. The severalfold greater uptake (21.2-74) of (64)Cu-TP3982 is attributable to the in vivo stability of the agent. CONCLUSION: The results suggest that the uncompromised biologic activity and the significantly greater tumor uptake of (64)Cu-TP3982, combined with the high sensitivity and enhanced resolution of PET imaging, make (64)Cu-TP3982 highly desirable for further studies in PET imaging of oncogene receptors overexpressed in breast and other types of cancers. PMID- 15299067 TI - Absolute quantification of regional cerebral glucose utilization in mice by 18F FDG small animal PET scanning and 2-14C-DG autoradiography. AB - The purpose of this study was to evaluate the feasibility of absolute quantification of regional cerebral glucose utilization (rCMR(glc)) in mice by use of (18)F-FDG and a small animal PET scanner. rCMR(glc) determined with (18)F FDG PET was compared with values determined simultaneously by the autoradiographic 2-(14)C-DG method. In addition, we compared the rCMR(glc) values under isoflurane, ketamine and xylazine anesthesia, and awake states. METHODS: Immediately after injection of (18)F-FDG and 2-(14)C-DG into mice, timed arterial samples were drawn over 45 min to determine the time courses of (18)F-FDG and 2 (14)C-DG. Animals were euthanized at 45 min and their brain was imaged with the PET scanner. The brains were then processed for 2-(14)C-DG autoradiography. Regions of interest were manually placed over cortical regions on corresponding coronal (18)F-FDG PET and 2-(14)C-DG autoradiographic images. rCMR(glc) values were calculated for both tracers by the autoradiographic 2-(14)C-DG method with modifications for the different rate and lumped constants for the 2 tracers. RESULTS: Average rCMR(glc) values in cerebral cortex with (18)F-FDG PET under normoglycemic conditions (isoflurane and awake) were generally lower (by 8.3%) but strongly correlated with those of 2-(14)C-DG (r(2) = 0.95). On the other hand, under hyperglycemic conditions (ketamine/xylazine) average cortical rCMR(glc) values with (18)F-FDG PET were higher (by 17.3%) than those with 2 (14)C-DG. Values for rCMR(glc) and uptake (percentage injected dose per gram [%ID/g]) with (18)F-FDG PET were significantly lower under both isoflurane and ketamine/xylazine anesthesia than in the awake mice. However, the reductions of rCMR(glc) were markedly greater under isoflurane (by 57%) than under ketamine and xylazine (by 19%), whereas more marked reductions of %ID/g were observed with ketamine/xylazine (by 54%) than with isoflurane (by 37%). These reverse differences between isoflurane and ketamine/xylazine may be due to competitive effect of (18)F-FDG and glucose uptake to the brain under hyperglycemia. CONCLUSION: We were able to obtain accurate absolute quantification of rCMR(glc) with mouse (18)F-FDG PET imaging as confirmed by concurrent use of the autoradiographic 2-(14)C-DG method. Underestimation of rCMR(glc) by (18)F-FDG in normoglycemic conditions may be due to partial-volume effects. Computation of rCMR(glc) from (18)F-FDG data in hyperglycemic animals may require, however, alternative rate and lumped constants for (18)F-FDG. PMID- 15299068 TI - Comparative 18F-FDG PET of experimental Staphylococcus aureus osteomyelitis and normal bone healing. AB - PET using (18)F-FDG is a promising imaging modality for bone infections, based on intensive consumption of glucose by mononuclear cells and granulocytes. The method may have limitations in distinguishing uncomplicated bone healing from osteomyelitis. Bone healing involves an inflammatory phase that represents a highly activated state of cell metabolism and glucose consumption, mimicking infection on PET images. This laboratory study of a standardized model was designed to compare the (18)F-FDG PET characteristics of normal bone healing with those of local osteomyelitis. METHODS: A localized osteomyelitis model of the rabbit tibia was created by modifying a previously reported canine model. In the osteomyelitic group (n = 8), a standardized metaphyseal defect of the proximal right tibia was surgically created and filled with a block of orthopedic bone cement, followed by injection of a predetermined amount (0.1 mL) of Staphylococcus aureus (strain 52/52A/80, 1 x 10(5)/mL) into the space around the cement. The control group of animals with normal bone healing (n = 8) underwent the same procedure, but the bacterial injection was replaced by a sterile saline injection. The bone cement was surgically removed during debridement at 2 wk. Osteomyelitis was confirmed with positive bacterial cultures during the debridement and 6 wk later at the time of sacrifice. (18)F-FDG PET and peripheral quantitative CT were performed 3 and 6 wk after the debridement. The presence of osteomyelitic bone changes on plain radiographs was classified according to a previously published system. RESULTS: Before surgery, the standardized uptake values of (18)F-FDG did not differ markedly between the right and left tibias. In the control animals, uncomplicated bone healing was associated with a temporary increase in (18)F-FDG uptake at 3 wk (P = 0.007), but it returned almost to normal by 6 wk. In the experimental animals, localized osteomyelitis resulted in an intense continuous uptake of (18)F-FDG, which was higher than that of healing and intact bones at 3 wk (P = 0.014 and P < 0.001, respectively) and at 6 wk (P < 0.001). CONCLUSION: (18)F-FDG PET seems to be an efficient tool in the differentiation of uneventful bone healing from bone healing complicated by localized osteomyelitis. PMID- 15299069 TI - Phase I clinical trial with fractionated radioimmunotherapy using 131I-labeled chimeric G250 in metastatic renal cancer. AB - This trial was performed to determine the maximum tolerated whole-body radiation absorbed dose of fractionated (131)I-cG250. METHODS: This was a phase 1 dose escalation trial. Dose escalation refers here to the escalation of average whole body absorbed dose. Fifteen patients with measurable metastatic renal cancer were studied. For each treatment cycle, patients initially received a "scout" administration consisting of 5 mg of cG250 antibody labeled with 185 MBq (5 mCi) of (131)I. Whole-body and serum activity was measured for 1 wk, and a simple pharmacokinetic model was fitted to the measured data. The pharmacokinetic model was used to calculate the required activities, administered in a fractionated pattern with 2-3 d between fractions, projected to deliver the prescribed whole body absorbed dose. The initial cohort of 3 patients was prescribed an average whole-body absorbed dose of 0.50 Gy. In subsequent cohorts this was increased in 0.25-Gy increments. The first fraction in each cycle was 1,110 MBq (30 mCi) of (131)I conjugated to 5 mg of antibody. Subsequent fractions consisted of variable activities depending on the patient-specific whole-body clearance rates and the times between fractions. Patients without evidence of disease progression were retreated after recovery from toxicity if there was no evidence of altered pharmacokinetics or serum human antichimeric antibody titers, for a total of no more than 3 treatments. RESULTS: For the initial treatment course, the pharmacokinetics of the scout dose accurately predicted the pharmacokinetics of fractionated (131)I-cG250 therapy. In 2 patients, altered clearance accurately predicted development of human antichimeric antibody. Targeting to known disease >or= 2 cm in diameter was noted in all patients. Dose-limiting toxicity was hematopoietic, and the maximum tolerated dose per cycle was 0.75 Gy. CONCLUSION: Measurements of whole-body and serum clearance of cG250 antibody can be used to accurately predict the clearance of subsequent administrations, thus enabling rational treatment planning. An additional practical benefit of real-time pharmacokinetic monitoring is that therapy can be altered dynamically to reduce toxic side effects. However, there was no evidence for fractionation-induced sparing of the hematopoietic system in this study. PMID- 15299070 TI - 18F-FDG uptake and breast density in women with normal breast tissue. PMID- 15299071 TI - PET tracers for osteosarcoma. PMID- 15299072 TI - p53-independent NOXA induction overcomes apoptotic resistance of malignant melanomas. AB - Once melanoma metastasizes, no effective treatment modalities prolong survival in most patients. This notorious refractoriness to therapy challenges investigators to identify agents that overcome melanoma resistance to apoptosis. Whereas many survival pathways contribute to the death-defying phenotype in melanoma, a defect in apoptotic machinery previously highlighted inactivation of Apaf-1, an apoptosome component engaged after mitochondrial damage. During studies involving Notch signaling in melanoma, we observed a gamma-secretase tripeptide inhibitor (GSI; z-Leu-Leu-Nle-CHO), selected from a group of compounds originally used in Alzheimer's disease, induced apoptosis in nine of nine melanoma lines. GSI only induced G2-M growth arrest (but not killing) in five of five normal melanocyte cultures tested. Effective killing of melanoma cells by GSI involved new protein synthesis and a mitochondrial-based pathway mediated by up-regulation of BH3-only members (Bim and NOXA). p53 activation was not necessary for up-regulation of NOXA in melanoma cells. Blocking GSI-induced NOXA using an antisense (but not control) oligonucleotide significantly reduced the apoptotic response. GSI also killed melanoma cell lines with low Apaf-1 levels. We conclude that GSI is highly effective in killing melanoma cells while sparing normal melanocytes. Direct enhancement of BH3-only proteins executes an apoptotic program overcoming resistance of this lethal tumor. Identification of a p53-independent apoptotic pathway in melanoma cells, including cells with low Apaf-1, bypasses an impediment to current cytotoxic therapy and provides new targets for future therapeutic trials involving chemoresistant tumors. PMID- 15299073 TI - Characterization of inhibitors of specific carboxylesterases: development of carboxylesterase inhibitors for translational application. AB - Carboxylesterases, expressed at high levels in human liver and intestine, are thought to detoxify xenobiotics. The anticancer prodrug 7-ethyl-10-[4-1 piperidino)-1-piperidino]carbonyloxycamptothecin (CPT-11) is also metabolized by carboxylesterases to produce the active drug 7-ethyl-10-hydroxycamptothecin. Activation of CPT-11 by human intestinal carboxylesterase (hiCE) in the human intestine may contribute to delayed onset diarrhea, a dose-limiting side effect of this drug. The goal of this study was to develop small molecule inhibitors selective for hiCE to circumvent or treat the toxic side effects of CPT-11. A secondary goal was to develop molecules that specifically inhibit activation of CPT-11 by a rabbit liver carboxylesterase (rCE). rCE is the most efficient CPT-11 activating enzyme thus far identified, and this enzyme is being developed for viral-directed enzyme prodrug therapy applications. Based on in vitro assays with partially purified hiCE and rCE proteins and on growth inhibition assays using U373MG human glioma cells transfected to express hiCE or rCE (U373pIREShiCE or U373pIRESrCE), we identified specific inhibitors of each enzyme. Lead compounds are derivatives of nitrophenol having 4-(furan-2-carbonyl)-piperazine-1 carboxylic acid or 4-[(4-chlorophenyl)-phenylmethyl]-piperazine-1-carboxylic acid substitutions in the p position. Kinetic analysis of each compound for hiCE compared with rCE showed that the Ki values of the most selective of these inhibitors differed by 6- to 10-fold. In growth inhibition assays, nontoxic, low micromolar concentrations of these inhibitors increased the EC50 of CPT-11 for U373pIREShiCE or U373pIRESrCE cells by 13- to >1,500-fold. The four compounds characterized in this study will serve as lead compounds for a series of inhibitors to be constructed using a combinatorial approach. PMID- 15299074 TI - The DNA double-stranded break repair protein endo-exonuclease as a therapeutic target for cancer. AB - DNA repair mechanisms are crucial for the maintenance of genomic stability and are emerging as potential therapeutic targets for cancer. In this study, we report that the endo-exonuclease, a protein involved in the recombination repair process of the DNA double-stranded break pathway, is overexpressed in a variety of cancer cells and could represent an effective target for developing anticancer drugs. We identify a dicationic diarylfuran, pentamidine, which has been used clinically to treat opportunistic infections and is an inhibitor of the endo exonuclease as determined by enzyme kinetic assay. In clonogenic and 3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays as well as in the in vivo Lewis lung carcinoma mouse tumor model, pentamidine is shown to possess the ability to selectively kill cancer cells. The LD50 of pentamidine on cancer cells maintained in vitro is correlated with the endo-exonuclease enzyme activity. Tumor cell that has been treated with pentamidine is reduced in the endo exonuclease as compared with the untreated control. Furthermore, pentamidine synergistically potentiates the cytotoxic effect of DNA strand break and cross link-inducing agents such as mitomycin C, etoposide, and cisplatin. In addition, we used the small interfering RNA for the mouse homologue of the endo-exonuclease to down-regulate the level of endo-exonuclease in the mouse myeloma cell line B16F10. Down-regulation of the endo-exonuclease sensitizes the cell to 5 fluorouracil. These studies suggested the endo-exonuclease enzyme as a novel potential therapeutic target for cancer. PMID- 15299075 TI - Preclinical validation of anti-TMEFF2-auristatin E-conjugated antibodies in the treatment of prostate cancer. AB - Current treatments for advanced stage, hormone-resistant prostate cancer are largely ineffective, leading to high patient mortality and morbidity. To fulfill this unmet medical need, we used global gene expression profiling to identify new potential antibody-drug conjugate (ADC) targets that showed maximal prostate cancer-specific expression. TMEFF2, a gene encoding a plasma membrane protein with two follistatin-like domains and one epidermal growth factor-like domain, had limited normal tissue distribution and was highly overexpressed in prostate cancer. Immunohistochemistry analysis using a specific monoclonal antibody (mAb) to human TMEFF2 showed significant protein expression in 74% of primary prostate cancers and 42% of metastatic lesions from lymph nodes and bone that represented both hormone-naive and hormone-resistant disease. To evaluate anti-TMEFF2 mAbs as potential ADCs, one mAb was conjugated to the cytotoxic agent auristatin E via a cathepsin B-sensitive valine-citrulline linker. This ADC, Pr1-vcMMAE, was used to treat male severe combined immunodeficient mice bearing xenografted LNCaP and CWR22 prostate cancers expressing TMEFF2. Doses of 3 to 10 mg/kg of this specific ADC resulted in significant and sustained tumor growth inhibition, whereas an isotype control ADC had no significant effect. Similar efficacy and specificity was shown with huPr1-vcMMAE, a humanized anti-TMEFF2 ADC. No overt in vivo toxicity was observed with either murine or human ADC, despite significant cross reactivity of anti-TMEFF2 mAb with the murine TMEFF2 protein, implying minimal toxicity to other body tissues. These data support the further evaluation and clinical testing of huPr1-vcMMAE as a novel therapeutic for the treatment of metastatic and hormone-resistant prostate cancer. PMID- 15299076 TI - Sanguinarine causes cell cycle blockade and apoptosis of human prostate carcinoma cells via modulation of cyclin kinase inhibitor-cyclin-cyclin-dependent kinase machinery. AB - Prostate cancer is the second leading cause of cancer-related deaths in males in the United States. This warrants the development of novel mechanism-based strategies for the prevention and/or treatment of prostate cancer. Several studies have shown that plant-derived alkaloids possess remarkable anticancer effects. Sanguinarine, an alkaloid derived from the bloodroot plant Sanguinaria canadensis, has been shown to possess antimicrobial, anti-inflammatory, and antioxidant properties. Previously, we have shown that sanguinarine possesses strong antiproliferative and proapoptotic properties against human epidermoid carcinoma A431 cells and immortalized human HaCaT keratinocytes. Here, employing androgen-responsive human prostate carcinoma LNCaP cells and androgen unresponsive human prostate carcinoma DU145 cells, we studied the antiproliferative properties of sanguinarine against prostate cancer. Sanguinarine (0.1-2 micromol/L) treatment of LNCaP and DU145 cells for 24 hours resulted in dose-dependent (1) inhibition of cell growth [as evaluated by 3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay], (2) arrest of cells in G0-G1 phase of the cell cycle (as assessed by DNA cell cycle analysis), and (3) induction of apoptosis (as evaluated by DNA ladder formation and flow cytometry). To define the mechanism of antiproliferative effects of sanguinarine against prostate cancer, we studied the effect of sanguinarine on critical molecular events known to regulate the cell cycle and the apoptotic machinery. Immunoblot analysis showed that sanguinarine treatment of both LNCaP and DU145 cells resulted in significant (1) induction of cyclin kinase inhibitors p21/WAF1 and p27/KIP1; (2) down-regulation of cyclin E, D1, and D2; and (3) down regulation of cyclin-dependent kinase 2, 4, and 6. A highlight of this study was the fact that sanguinarine induced growth inhibitory and antiproliferative effects in human prostate carcinoma cells irrespective of their androgen status. To our knowledge, this is the first study showing the involvement of cyclin kinase inhibitor-cyclin-cyclin-dependent kinase machinery during cell cycle arrest and apoptosis of prostate cancer cells by sanguinarine. These results suggest that sanguinarine may be developed as an agent for the management of prostate cancer. PMID- 15299077 TI - Polyomavirus middle T-induced mammary intraepithelial neoplasia outgrowths: single origin, divergent evolution, and multiple outcomes. AB - The development of models to investigate the pathobiology of premalignant breast lesions is a critical prerequisite for development of breast cancer prevention and early intervention strategies. Using tissue transplantation techniques, we modified the widely used polyomavirus middle T (PyV-mT) transgenic mouse model of breast cancer to study the premalignant stages of tumorigenesis. Premalignant atypical lesions were isolated from PyV-mT transgenic mice and used to generate two sets of three mammary intraepithelial neoplasia (MIN) outgrowth lines. Investigation of these six unique lines, each of which fulfills the criteria for MIN, has provided new information regarding the biology of PyV-mT-induced neoplasia. Although expression of the PyV-mT transgene was the primary initiating event for all lines, they exhibited different tumor latencies, metastatic potentials, and morphologies. Six distinguishable morphologic patterns of differentiation were identified within the premalignant outgrowths that are likely to represent several tumorigenic pathways. Further, several tumor phenotypes developed from each line and the tumors developing from the six lines had different metastatic potentials. These observations are consistent with the hypothesis that distinct pathways of PyV-mT-initiated neoplastic progression lead to different outcomes with respect to latency and metastasis. The MIN outgrowth lines share several characteristics with precursors of human breast cancer including the observation that gene expression profiles of tumors are more similar to those of the MIN outgrowth line outgrowth from which they developed than to other tumors. These lines provide an opportunity to study the full range of events occurring secondary to PyV-mT expression in the mammary gland. PMID- 15299078 TI - Transient adenoviral N-methylpurine DNA glycosylase overexpression imparts chemotherapeutic sensitivity to human breast cancer cells. AB - In an effort to improve the efficacy of cancer chemotherapy by intervening into the cellular responses to chemotherapeutic change, we have used adenoviral overexpression of N-methylpurine DNA glycosylase (MPG or ANPG/AAG) in breast cancer cells to study its ability to imbalance base excision repair (BER) and sensitize cancer cells to alkylating agents. Our results show that MPG overexpressing cells are significantly more sensitive to the alkylating agents methyl methanesulfonate, N-methyl-N'-nitro-N-nitrosoguanidine, methylnitrosourea, dimethyl sulfate, and the clinical chemotherapeutic temozolomide. Sensitivity is further increased through coadministration of the BER inhibitor methoxyamine, which covalently binds abasic or apurinic/apyrimidinic (AP) sites and makes them refractory to subsequent repair. Methoxyamine reduction of cell survival is significantly greater in cells overexpressing MPG than in control cells, suggesting a heightened production of AP sites that, if made persistent, results in increased cellular toxicity. We further explored the mechanism of MPG-induced sensitivity and found that sensitivity was associated with a significant increase in the number of AP sites and/or single-strand breaks in overexpressing cells, confirming a MPG-driven accumulation of toxic BER intermediates. These data establish transient MPG overexpression as a potential therapeutic approach for increasing cellular sensitivity to alkylating agent chemotherapy. PMID- 15299079 TI - Treatment of experimental breast cancer using interleukin-12 gene therapy combined with anti-vascular endothelial growth factor receptor-2 antibody. AB - We have shown previously that interleukin-12 (IL-12) gene therapy induced strong antitumor effects in several syngeneic murine tumor models including 4T1 mammary adenocarcinoma. Antiangiogenic treatment with a monoclonal antibody (mAb) directed against the vascular endothelial growth factor receptor-2 (VEGFR-2) is another promising treatment approach that can cause transient suppression of tumor growth. We hypothesized that the combination of IL-12 gene therapy and anti VEGFR-2 mAb will achieve better antitumor and antimetastatic effects against 4T1 adenocarcinoma than each treatment alone via implementation of different mechanisms. Administration of anti-VEGFR-2 mAb into BALB/c mice bearing s.c. 4T1 tumors induced significant suppression of tumor growth, as did intratumoral administration of naked IL-12 DNA. The combined treatment with anti-VEGFR-2 mAb and IL-12 DNA resulted in significantly enhanced inhibition of tumor growth as compared with each treatment alone. This combination was also effective against spontaneous lung metastases. In T-cell-deficient nude mice, both IL-12 DNA and anti-VEGFR-2 mAb were effective in suppressing tumor growth. In T-cell- and natural killer cell-deficient scid/beige mice, only anti-VEGFR-2 mAb was effective, suggesting that natural killer cells are involved in the antitumor effects induced by IL-12 DNA. In both types of immunodeficient mice, the combination of anti-VEGFR-2 mAb and IL-12 DNA was as effective in suppressing 4T1 tumor growth as anti-VEGFR-2 mAb alone. Antitumor effects of anti-VEGFR-2 mAb were associated with the inhibition of angiogenesis within the tumors, whereas the antiangiogenic effect of IL-12 gene therapy was not detected. Our results show a therapeutic benefit of combining IL-12 gene therapy and anti-VEGFR-2 mAb for cancer treatment. PMID- 15299080 TI - Antitumor activity of ZD6126, a novel vascular-targeting agent, is enhanced when combined with ZD1839, an epidermal growth factor receptor tyrosine kinase inhibitor, and potentiates the effects of radiation in a human non-small cell lung cancer xenograft model. AB - OBJECTIVE: Targeting the tumor vasculature may offer an alternative or complementary therapeutic approach to targeting growth factor signaling in lung cancer. The aim of these studies was to evaluate the antitumor effects in vivo of the combination of ZD6126, a tumor-selective vascular-targeting agent; ZD1839 (gefitinib, Iressa), an epidermal growth factor receptor tyrosine kinase inhibitor; and ionizing radiation in the treatment of non-small cell lung cancer xenograft model. METHODS: Athymic nude mice with established flank A549 human non small cell lung cancer xenograft model xenografts were treated with fractionated radiation therapy, ZD6126, ZD1839, or combinations of each treatment. ZD6126 (150 mg/kg) was given i.p. the day after each course of radiation. Animals treated with ZD1839 received 100 mg/kg per dose per animal, 5 or 7 days/wk for 2 weeks. Immunohistochemistry was done to evaluate the effects on tumor growth using an anti-Ki67 monoclonal antibody. Effects on tumor-induced vascularization were quantified using an anti-factor VIII-related antigen monoclonal antibody. RESULTS: ZD6126 attenuated the growth of human A549 flank xenografts compared with untreated animals. Marked antitumor effects were observed when animals were treated with a combination of ZD6126 and fractionated radiation therapy with protracted tumor regression. ZD6126 + ZD1839 resulted in a greater tumor growth delay than either agent alone. Similar additive effects were seen with ZD1839 + fractionated radiation. Finally, the addition of ZD6126 to ZD1839 and radiation therapy seemed to further improve tumor growth control, with a significant tumor growth delay compared with animals treated with single agent or with double combinations. Immunohistochemistry showed that ZD1839 induced a marked reduction in A549 tumor cell proliferation. Both ZD1839 and ZD6126 treatment substantially reduced tumor-induced angiogenesis. ZD6126 caused marked vessel destruction through loss of endothelial cells and thrombosis, substantially increasing the level of necrosis seen when combined with radiation therapy. The combination of radiation therapy, ZD6126, and ZD1839 induced the greatest effects on tumor growth and angiogenesis. CONCLUSION: This first report shows that a selective vascular-targeting agent (ZD6126) + an anti-epidermal growth factor receptor agent (ZD1839) and radiation have additive in vivo effects in a human cancer model. Targeting the tumor vasculature offers an excellent strategy to enhance radiation cytotoxicity. Polytargeted therapy with agents that interfere with both growth factor and angiogenic signaling warrants further investigation. PMID- 15299081 TI - Inhibition of constitutively activated nuclear factor-kappaB radiosensitizes human melanoma cells. AB - Melanoma tumors and cultured cell lines are relatively resistant to the cytotoxic effects of ionizing radiation, thereby limiting the use of radiotherapy for the clinical treatment of melanoma. New strategies for sensitizing melanoma cells therefore deserve examination. In an attempt to identify and target signaling pathways that contribute to radioresistance, we investigated the role of nuclear factor-kappaB (NF-kappaB), a transcription factor known to inhibit apoptosis induced by a variety of stimuli and promote radioresistance. Two human metastatic melanoma cell lines, A375 and MeWo, were used to examine the radiosensitizing effects of inhibitors of the NF-kappaB pathway. Nuclear extracts from these cell lines were tested for active NF-kappaB using the electrophoretic mobility shift assay. Both melanoma cell lines had constitutively activated NF-kappaB as observed by electrophoretic mobility shift assay. In an attempt to reverse NF kappaB activity, cells were treated either with vehicle alone (DMSO) or with a proteasome inhibitor Z-Leu-Leu-Leu-H (MG132; 10 micromol/L for 2 hours prior to irradiation) that inhibited both constitutive and radiation-induced NF-kappaB activity. The clonogenic cell survival assay showed that pretreatment with MG132 enhanced tumor cell radiosensitivity with the survival factor at 2 Gy being reduced from 48 +/- 0.8% and 48 +/- 1.6% in vehicle-treated cells to 27.7 +/- 0.32% and 34.3 +/- 0.7% in MG132-treated MeWo and A375 cells, respectively. To test the role of NF-kappaB in radioresistance more directly, MeWo cells were stably transfected with a dominant-negative mutant IkappaBalpha construct, which led to the inhibition of both constitutive and radiation-induced NF-kappaB activity. A modest restoration of radiosensitivity was also observed in the stably transfected MeWo cells with survival factor at 2 Gy values being reduced from 47 +/- 0.8% in parental MeWo cells to 32.9 +/- 0.7% in stable transfectants. Because constitutively activated mitogen-activated protein kinase kinase (MEK) pathway has been shown to lead to activated NF-kappaB, we wanted to determine the relative contribution of activated MEK in the human melanoma cells. To test this, MeWo and A375 melanoma cells were exposed to the MEK inhibitor PD184352. Treatment with PD184352 partially reversed NF-kappaB activity but did not impart radiation sensitivity to these cells. Our results indicate that activated NF kappaB may be one of the pathways responsible for the radioresistance of melanoma cells and that strategies for inhibiting its influence may be useful in restoring the radioresponse of melanomas. PMID- 15299082 TI - Transcriptome analysis of endometrial cancer identifies peroxisome proliferator activated receptors as potential therapeutic targets. AB - Endometrial cancer is the most common gynecologic malignancy, frequently arising in association with obesity and diabetes mellitus. To identify gene pathways contributing to endometrial cancer development, we studied the transcriptome of 20 endometrial cancers and 11 benign endometrial tissues using cDNA microarrays. Among the transcript changes identified in endometrial cancer were up-regulation of the nuclear hormone receptors peroxisome proliferator-activated receptors (PPAR) alpha and gamma, whereas retinoid X receptor beta was down-regulated. To clarify the contribution of PPARalpha to endometrial carcinogenesis, we did experiments on cultured endometrial carcinoma cells expressing this transcript. Treatment with fenofibrate, an activating ligand for PPARalpha, significantly reduced proliferation and increased cell death, suggesting that altered expression of nuclear hormone receptors involved with fatty acid metabolism leads to deregulated cellular proliferation and apoptosis. These results support further investigation of members of the PPAR/retinoid X receptor pathway as novel therapeutic targets in endometrial cancer. PMID- 15299083 TI - Telomerase inhibition by retinoids precedes cytodifferentiation of leukemia cells and may contribute to terminal differentiation. AB - Human promyelocytic leukemia HL60 cells display high telomerase activity, a phenotype related to their immortal status. All-trans retinoic acid (ATRA) is a clinically effective cytodifferentiating agent. To understand the mechanism underlying ATRA-induced cytodifferentiation, we did a kinetic analysis of the role of ATRA in inhibiting telomerase in HL60 cells. Our studies indicate that telomerase inhibition by ATRA occurred relatively early after treatment of HL60 cells due to a rapid decrease in hTERT gene expression. More importantly, however, we found through monitoring the expression of CD11b, a marker for granulocytic differentiation of HL60 cells, that down-regulation of telomerase preceded the differentiation of HL60 cells. These observations suggest that the hTERT gene may be a primary target of ATRA regulation of cellular differentiation and the antileukemia activity of ATRA may be mediated by its ability to induce the differentiation of the promyelocytic leukemia cells through down-regulation of the hTERT gene. PMID- 15299084 TI - Retinoid X receptor-gamma and peroxisome proliferator-activated receptor-gamma expression predicts thyroid carcinoma cell response to retinoid and thiazolidinedione treatment. AB - Poorly differentiated, metastatic thyroid cancer is difficult to treat. These tumors often do not concentrate radioactive iodine and may require chemotherapy, which is suboptimal and toxic. Nuclear hormone receptors peroxisome proliferator activated receptor gamma (PPARgamma) and retinoid X receptor (RXR) are variably expressed in thyroid carcinoma cell lines. Expression of these receptors may predict thyroid cancer cell response to treatment with rexinoids and thiazolidinediones. We studied three thyroid carcinoma cell lines: BHP 5-16 (PPARgamma-/RXRgamma+), BHP 2-7 (PPARgamma+/-/RXRgamma-), and DRO-90 (RXRgamma+/PPARgamma+). BHP 5-16 (RXRgamma+) cells treated with rexinoid had decreased proliferation to 69 +/- 6% growth compared with vehicle. BHP 2-7 (PPARgamma+) cells treated with thiazolidinedione had no decrease in cellular proliferation. DRO-90 (RXRgamma+ and PPARgamma+) cells had 36 +/- 10%, 15 +/- 3%, and 13 +/- 4% growth when treated with rexinoid, thiazolidinedione, or a combination, respectively. We next investigated the role of apoptosis in the ligand-responsive BHP 5-16 and DRO-90 cells. BHP 5-16 cells underwent no significant apoptosis with rexinoid (1 micromol/L). DRO-90 cells, however, had 3.6 +/- 1.3% apoptotic cells with vehicle, 13 +/- 3.5% with rexinoid (1 micromol/L), 18 +/- 4% with thiazolidinedione (1 micromol/L), and 28 +/- 6% with combination treatment (1 micromol/L), suggesting that apoptosis plays a major role in this anaplastic cell line and that the effects of the two ligands are additive. We conclude that receptor expression is necessary for inhibition of thyroid carcinoma growth with ligand treatment but may not be sufficient for response. Additionally, expression of both RXRgamma and PPARgamma may be necessary for maximal growth inhibition by ligands and may be required for the increased apoptosis. PMID- 15299085 TI - Altered Hsp90 function in cancer: a unique therapeutic opportunity. AB - Molecular chaperones or so-called heat shock proteins serve as central integrators of protein homeostasis within cells. In performing this function, they guide the folding, intracellular disposition, and proteolytic turnover of many key regulators of cell growth, differentiation, and survival. Recent data show essential roles for the chaperones in facilitating malignant transformation at the molecular level and support the concept that their altered utilization during oncogenesis is critical to the development of human cancers. The field is evolving rapidly, but it has become apparent that chaperones can serve as biochemical buffers at the phenotypic level for the genetic instability that is characteristic of many human cancers. Chaperone proteins thus allow tumor cells to tolerate the mutation of multiple critical signaling molecules that would otherwise be lethal. Much of the recent progress in understanding the complex role of heat shock proteins in tumorigenesis has been made possible by the discovery of several natural product antitumor antibiotics that selectively inhibit the function of the chaperone Hsp90. These agents have been used as probes to define the biological functions of Hsp90 at the molecular level and to validate it as a novel target for anticancer drug action. One of these agents, 17 allylamino,17-demethoxygeldanamycin (NSC 330507) has begun phase II clinical trials, and several second-generation compounds are now in late preclinical development. The best way to use Hsp90 inhibitors as anticancer agents remains to be defined. Trials accomplished to date, however, serve as proof of principle that Hsp90 function can be modulated pharmacologically without undue toxicity in humans. Given the redundancy and complexity of the signaling pathway abnormalities present in most cancers, the ability of Hsp90 inhibitors to alter the activity of multiple aberrant signaling molecules instead of just one or two (as most current-generation molecular therapeutics have been designed to do) may prove of unique therapeutic benefit. PMID- 15299086 TI - Prostaglandin EP receptors: targets for treatment and prevention of colorectal cancer? AB - The importance of the prostaglandin (PG) synthesis pathway, particularly the rate limiting enzymatic step catalyzed by cyclooxygenase, to colorectal carcinogenesis and development of novel anticolorectal cancer therapy is well established. The predominant PG species in benign and malignant colorectal tumors is PGE2. PGE2 acts via four EP receptors termed EP1 to EP4. Recently, EP receptors have been identified as potential targets for treatment and/or prevention of colorectal cancer. This review summarizes existing knowledge of the expression and function of the EP receptor subtypes in human and rodent intestine during tumorigenic progression and describes the current literature on targeting EP receptor signaling during intestinal tumorigenesis. PMID- 15299087 TI - Essential structural and functional determinants within the forkhead domain of FOXC1. AB - The forkhead domain (FHD)-containing developmental transcription factor FOXC1 is mutated in patients presenting with Axenfeld-Rieger malformations. In this paper, we report the introduction of positive, negative or neutral charged amino acids into critical positions within the forkhead domain of FOXC1 in an effort to better understand the essential structural and functional determinants within the FHD. We found that FOXC1 is intolerant of mutations at I87. Additionally, alterations of amino acids within alpha-helix 1 of the FOXC1 FHD affected both nuclear localization and transactivation. Amino acids within alpha-helix 3 were also found to be necessary for transactivation and can have roles in correct localization. Interestingly, changing amino acids within alpha-helix 3, particularly R127, resulted in altered DNA-binding specificity and granted FOXC1 the ability to bind to a novel DNA sequence. Given the limited topological variation of FHDs, due to the high conservation of residues, we anticipate that models of forkhead domain function derived from these data will be relevant to other members of the FOX family of transcription factors. PMID- 15299088 TI - hnRNP H binding at the 5' splice site correlates with the pathological effect of two intronic mutations in the NF-1 and TSHbeta genes. AB - We have recently reported a disease-causing substitution (+5G > C) at the donor site of NF-1 exon 3 that produces its skipping. We have now studied in detail the splicing mechanism involved in analyzing RNA-protein complexes at several 5' splice sites. Characteristic protein patterns were observed by pulldown and band shift/super-shift analysis. Here, we show that hnRNP H binds specifically to the wild-type GGGgu donor sequence of the NF-1 exon 3. Depletion analyses shows that this protein restricts the accessibility of U1 small nuclear ribonucleoprotein (U1snRNA) to the donor site. In this context, the +5G > C mutation abolishes both U1snRNP base pairing and the 5' splice site (5'ss) function. However, exon recognition in the mutant can be rescued by disrupting the binding of hnRNP H, demonstrating that this protein enhances the effects of the +5G > C substitution. Significantly, a similar situation was found for a second disease-causing +5G > A substitution in the 5'ss of TSHbeta exon 2, which harbors a GGgu donor sequence. Thus, the reason why similar nucleotide substitutions can be either neutral or very disruptive of splicing function can be explained by the presence of specific binding signatures depending on local contexts. PMID- 15299089 TI - Activator protein-1 in human male germ cell apoptosis. AB - Apoptosis limits germ cell number in the testis, and its dysregulation is associated with male infertility. Here, we evaluated the role of the transcription factor activator protein 1 (AP-1) in male germ cell apoptosis in a culture of human seminiferous tubules. AP-1 DNA-binding activity increased in cultured tubules within 2.5 h, which was earlier than the onset of apoptosis as detected by caspase 3 activation and apoptotic DNA fragmentation. The c-Jun, c Fos and JunD proteins were detected in the Sertoli cell nuclei, whereas apoptosis occurred in the germ cells. Follicle-stimulating hormone (FSH), whose receptors are expressed in the Sertoli cells, inhibited germ cell apoptosis and concomitantly suppressed AP-1 DNA-binding activity, but had no effect on nuclear factor kappaB (NF-kappaB) activation. These results suggest that AP-1 transcription factors are involved in the Sertoli cell-mediated control of germ cell apoptosis, and that inhibition of germ cell apoptosis by FSH appears to involve suppression of AP-1 activation. PMID- 15299090 TI - Glutathione S-transferase M1*null genotype but not myeloperoxidase promoter G 463A polymorphism is associated with higher susceptibility to endometriosis. AB - Glutathione S-transferase M1 (GSTM1), one member of the GST family, is responsible for metabolism of xenobiotics and carcinogens. Myeloperoxidase (MPO) plays an important role in the oxidation and activation of carcinogens and nitric oxide. Allelic variants of GSTM1 and MPO gene polymorphisms might impair detoxification function and increase the susceptibility to endometriosis. We aimed to investigate if these polymorphisms are useful markers for predicting endometriosis susceptibility. Women were divided into two groups: (i) endometriosis (n=150); (ii) non-endometriosis (n=159). Polymorphisms for GSTM1 and MPO were amplified by polymerase chain reaction and detected by electrophoresis after restriction digestion. The relative frequencies of the GSTM1*wild (+/+,+/0)/null (0/0) genotypes and MPO-463*G/A gene polymorphisms between both groups were compared. The distribution of GSTM1 polymorphisms was significantly different between the two groups. Proportions of GSTM1*wild/null alleles in both groups were: (i) 36.7/63.3%; (ii) 95/5% (P=0.001). In contrast, MPO-463 genotypes were not significantly different between the two groups. Proportions of MPO*A homozygote/heterozygote/G homozygote in both groups were: (i) 2.7/17.4/79.9% and (ii) 1.9/17/81.1% (P> 0.05). We conclude that the GSTM1*null genotype is associated with a higher risk of endometriosis development. MPO-463*G/A gene polymorphism is not related to the susceptibility of endometriosis. PMID- 15299091 TI - Ah receptor, CYP1A1, CYP1A2 and CYP1B1 gene polymorphisms are not involved in the risk of recurrent pregnancy loss. AB - The etiology of recurrent pregnancy loss (RPL) remains unclear, but it may be related to a possible genetic predisposition together with involvement of environmental factors. We examined the relation between RPL and polymorphisms in four genes, human aryl hydrocarbon (Ah) receptor, cytochrome P450 (CYP) 1A1, CYP1A2 and CYP1B1, which are involved in the metabolism of a wide range of environmental toxins and carcinogens. All cases and controls were women resident in Sapporo, Japan and the surrounding area. The Ah receptor, CYP1A1, CYP1A2 and CYP1B1 genotypes were assessed in 113 Japanese women with recurrent pregnancy loss (RPL) and 203 ethnically matched women experiencing at least one live birth and no spontaneous abortion (control). No significant differences in Ah receptor, CYP1A1, CYP1A2 and CYP1B1 genotype frequencies were found between the women with RPL and the controls [Ah receptor: Arg/Arg (reference); Arg/Lys and Lys/Lys, odds ratio (OR)=0.67; 95% confidence interval (CI)=0.40-1.11, CYP1A1: m1m1 (reference); m1m2 and m2m2, OR = 0.86; 95% CI = 0.53-1.40, CYP1A2: C/C and C/A (reference); A/A, OR = 1.16; 95% CI = 0.71-1.88, CYP1B1: Leu/Leu (reference); Leu/Val and Val/Val, OR = 1.18; 95% CI = 0.68-2.02]. The present study suggests that the Ah receptor, CYP1A1, CYP1A2 and CYP1B1 gene polymorphisms are not major genetic regulators in RPL. PMID- 15299092 TI - DNA microarray analysis of gene expression profiles in deep endometriosis using laser capture microdissection. AB - Endometriosis, a common gynecological disorder that causes infertility and pelvic pain, is defined as the presence of endometrial glands and stroma within extra uterine sites. However, despite extensive studies its etiology and pathogenesis are not completely understood. Differentially expressed genes were investigated in epithelial and stromal cells from deep endometriosis and matched eutopic endometrium using cDNA microarrays and laser capture microdissection. Validation of results of several up- and down-regulated genes was performed by quantitative real-time RT-PCR. Our data showed that platelet-derived growth factor receptor alpha (PDGFRA), protein kinase C beta1 (PKC beta1) and janus kinase 1 (JAK1) were upregulated, and Sprouty2 and mitogen-activated protein kinase kinase 7 (MKK7) were downregulated in endometriosis stromal cells, suggesting the involvement of the RAS/RAF/MAPK signaling pathway through PDGFRA in endometriosis pathophysiology. In addition, two potential negative regulators of aromatase expression, chicken ovalbumin upstream promoter transcription factor 2 (COUP-TF2) and prostaglandin E2 receptor subtype EP3 (PGE2EP3), were downregulated in endometriosis epithelial cells, which might result in increased local production of estrogen in endometriosis epithelial cells. Furthermore, three potential candidate genes that might be involved in endometriosis related pain were identified: tyrosine kinase receptor B (TRkB) in endometriosis epithelial cells, and serotonin transporter (5HTT) and mu opioid receptor (MOR) in endometriosis stromal cells were all upregulated. One of the candidate genes, MOR, may be involved in a defective immune system in endometriosis. This study has provided new insights into endometriosis pathophysiology. PMID- 15299093 TI - Absence of the genetic variant Val79Met in human chorionic gonadotropin-beta gene 5 in five European populations. AB - Chorionic gonadotropin (CG) is an essential signal in establishment and maintenance of pregnancy in humans and higher primates. A G-to-A transition in exon 3 of human CGbeta gene 5, changing the naturally occurring valine residue to methionine in codon 79 (Val(79)Met) has been reported at carrier frequency 4.2% in a random population from the Midwest of the United States. The biological activity of the variant hCG was similar to that of wild-type (WT) hCG. However, the Val(79)Met beta-subunit displayed impaired ability to assemble with alpha subunit, and the amount of hCG alpha/beta heterodimers formed and secreted by transfected cells was seriously impaired in the previous study. Because of these functional implications we found it important to study the occurrence of the Val(79)Met hCGbeta variant in other populations. By using a PCR-RFLP method, a search for the Val(79)Met hCGbeta variant was carried out on a total of 580 DNA samples from five European populations (Finland, Denmark, Greece, Germany and the UK). The results demonstrated an absence of the polymorphism in these populations. Hence, the naturally occurring variant (Val(79)Met) of the hCGbeta gene 5, found previously at high frequency in the US, is clearly less common, or absent, in the European populations studied. PMID- 15299094 TI - Hyperkalaemia: again. PMID- 15299095 TI - Fibrillary glomerulonephritis and immunotactoid glomerulopathy. PMID- 15299096 TI - The deadly risk of late referral. PMID- 15299097 TI - Endothelial nitric oxide synthase gene intron 4 polymorphism in patients with end stage renal disease. AB - BACKGROUND: Nitric oxide (NO) synthesized by endothelial cell NO synthase (ecNOS) is a potent regulator of intrarenal haemodynamics. A polymorphism in intron 4 of the ecNOS gene is a candidate gene in cardiovascular and renal diseases. We investigated a potential involvement of this polymorphism in chronic renal failure. METHODS: We performed a case-control study involving 706 patients with end-stage renal disease (ESRD) and 321 healthy controls. All subjects were genotyped for the ecNOS4 polymorphism by the polymerase chain reaction followed by agarose gel electrophoresis. RESULTS: The analysis revealed that the frequencies of the ecNOS4 genotypes were significantly different in ESRD patients, both diabetic and non-diabetic, than in controls. In all dialysis patients for aa, ab and bb genotypes the frequencies were, respectively, 6.5, 35 and 58.5% in the patient group, and 1, 25 and 74% in control subjects. The a allele carriers (aa + ab) were more frequent among ESRD patients than in controls (OR 1.95; 95% CI 1.13-3.4; P = 0.0031). No significant association was found when hypertensive ESRD patients were compared with normotensive patients. The distribution of genotypes was similar in both subgroups (P = 0.21). CONCLUSION: There was a significantly higher frequency of the ecNOS4a allele carriers among ESRD patients, both diabetic and non-diabetic, than in control subjects. This suggests that the ecNOS gene polymorphism may be associated with an increased risk of chronic renal failure. PMID- 15299099 TI - A strategy to achieve donor-specific hyporesponsiveness in cadaver renal allograft recipients by donor haematopoietic stem cell transplantation into the thymus and periphery. AB - INTRODUCTION: We designed a prospective, randomized clinical trial to evaluate the immune response to thymic and peripheral infusions of donor haematopoietic stem cells (HSCs) to create tolerance in recipients of cadaver renal allografts. METHOD: We divided 24 patients into two equal groups. For group A, 350 ml of unfractionated bone marrow (BM) was aspirated from the anterior iliac crests of donor cadavers. A 2 ml aliquot of concentrated marrow was infused into the thymus of the subject and 100 ml into the BM before surgery; the remaining 250 ml was infused peripherally post-transplantation. The mean nucleated cell count inoculated into the thymus was 3.3 x 10(4) cells/cm(3) and into the periphery 8.6 x 10(7) cells/kg body weight. Group B (controls) underwent renal transplantation directly. Recipients were lymphocytotoxicity cross-match negative in both groups. Group A received low dose prednisolone and cyclosporin; controls also received azathioprine. RESULTS: Over a mean follow-up of 703 days for both groups, group A had significantly better graft function with minimum acute rejection episodes or cytomegalovirus (CMV) infections, a mean serum creatinine (SCr) of 1.23 mg/dl and no graft or patient loss. Group B, with a mean SCr of 2.19 mg/dl had three patients with single acute rejection episodes, two of whom died following uncontrolled rejection-associated infections. The third patient maintained an SCr of 2.5 mg%. Actuarial graft survival was 87.5% in controls at the end of 2 years compared with group A with 100% graft survival at the end of 2 years. CONCLUSION: This novel approach of introducing unfractionated HSCs into the thymus and periphery to create tolerance is safe and efficacious and gives significantly better graft function, minimum acute rejection and no CMV disease with monotherapy. PMID- 15299098 TI - Vitamin C plasma level and response to erythropoietin in patients on maintenance haemodialysis. AB - BACKGROUND: Intravenous vitamin C supplementation to haemodialysis patients might ameliorate responsiveness to recombinant human erythropoietin (rHuEpo). This study was performed to analyse the relation between vitamin C plasma concentration and response to rHuEpo. METHODS: In a cross-sectional, single centre observational study including all haemodialysis patients, pre-dialysis plasma vitamin C concentrations were measured by high-performance liquid chromatography and response to rHuEpo (haemoglobin concentration/international units rHuEpo/kg/week) was recorded together with baseline laboratory data. RESULTS: Univariate analysis yielded a significant correlation between vitamin C plasma levels and response to rHuEpo (n = 130, r = 0.25, P = 0.004), which still persisted after adjustment for transferrin saturation, C-reactive protein, malondialdehyde, parathyroid hormone, route of rHuEpo administration, residual renal function and diabetes mellitus (adjusted r = 0.23, P = 0.014). Analysis per quartiles of vitamin C plasma level revealed a significantly lower response to rHuEpo with decreasing vitamin C values (P = 0.026). CONCLUSIONS: In unselected haemodialysis patients, vitamin C plasma levels account, at least partially, for the response to rHuEpo. Larger-sized interventional studies are needed to find out whether vitamin C plasma levels may or may not appropriately reflect the potential beneficial effect of vitamin C supplements on rHuEpo responsiveness. PMID- 15299100 TI - Long-term outcome of renal glucosuria type 0: the original patient and his natural history. PMID- 15299101 TI - Retroperitoneal fibrosis, sclerosing pancreatitis and bronchiolitis obliterans with organizing pneumonia. PMID- 15299103 TI - Acute transplant rejection induced by blood transfusion reaction to the Kidd blood group system. AB - Many renal transplant centres now try to avoid blood transfusion prior to renal transplantation, to avoid alloimmunization due to antibody production against donor antigens usually present on contaminating white cells. Post- or peri operative transfusions are usually not considered to present problems, since the patient is heavily immunosuppressed. We present a patient who suffered a rare transfusion reaction, that we believe may have initiated a severe vascular rejection of a kidney transplant, probably mediated by Kidd blood group antigens. PMID- 15299102 TI - A puzzling cause of persistent Pseudomonas aeruginosa septicaemia in a patient on maintenance haemodialysis. PMID- 15299104 TI - Orange-like skin lesion and hypertension--what is the link? PMID- 15299105 TI - Unexplained polyuria and non-obstructive hydronephrosis in a urological department. PMID- 15299106 TI - A rare cause of oedema. PMID- 15299107 TI - Iliac vein stenosis as a reversible cause of renal transplant dysfunction. PMID- 15299108 TI - The need for parathyroidectomy. PMID- 15299109 TI - Paralysis as first manifestation of primary aldosteronism. PMID- 15299110 TI - Lack of evidence for the 1484insG variant at the 3'-UTR of the protein tyrosine phosphatase 1B (PTP1B) gene as a genetic determinant of diabetic nephropathy development in type 1 diabetic patients. PMID- 15299111 TI - Renal involvement in Cogan's syndrome. PMID- 15299112 TI - When the kidney catches a cold: an unusual cause of acute renal failure. PMID- 15299113 TI - Erythropoietic proteins and antibody-mediated pure red cell aplasia: a potential role for micelles. PMID- 15299114 TI - Managing refractory uraemic pericarditis with colchicine. PMID- 15299115 TI - Corticosteroid and tamoxifen therapy in sclerosing encapsulating peritonitis in a patient on continuous ambulatory peritoneal dialysis. PMID- 15299116 TI - Emission of Plutella xylostella-induced compounds from cabbages grown at elevated CO2 and orientation behavior of the natural enemies. AB - Several plant species defend themselves indirectly from herbivores by producing herbivore-induced volatile compounds that attract the natural enemies of herbivores. Here we tested the effects of elevated atmospheric CO(2) (720 micromol mol(-1)) concentration on this indirect defense, physiological properties, and constitutive and induced emissions of white cabbage (Brassica oleracea ssp. capitata, cvs Lennox and Rinda). We monitored the orientation behavior of the generalist predator Podisus maculiventris (Heteroptera: Pentatomidae) and the specialist parasitoid Cotesia plutellae (Hymenoptera: Braconidae) to plants damaged by Plutella xylostella (Lepidoptera: Plutellidae) in the Y-tube olfactometer. Elevated CO(2) levels did not affect stomatal densities but reduced specific leaf area and increased leaf thickness in cv Lennox. In addition to enhanced constitutive monoterpene emission, P. xylostella damaged cabbages emitted homoterpene (E)-4,8-dimethyl-1,3,7-nonatriene, sesquiterpene (E,E)-alpha-farnesene, and (Z)-3-hexenyl acetate. Growth at elevated CO(2) had no significant effect on the emissions expressed per leaf area, while minor reduction in the emission of homoterpene (E)-4,8-dimethyl-1,3,7 nonatriene and (E,E)-alpha-farnesene was observed at elevated CO(2) in one of two experiments. The generalist predator P. maculiventris discriminated only between the odors of intact and P. xylostella-damaged cv Rinda plants grown at ambient CO(2) concentration, preferring the odor of the damaged plants. The specialist parasitoid C. plutellae preferred the odor of damaged plants of both cultivars grown at ambient CO(2) but did not detect damaged cv Lennox plants grown at elevated CO(2). The results suggest that elevated atmospheric CO(2) concentration could weaken the plant response induced by insect herbivore feeding and thereby lead to a disturbance of signaling to the third trophic level. PMID- 15299117 TI - Relocalization of nuclear ALY proteins to the cytoplasm by the tomato bushy stunt virus P19 pathogenicity protein. AB - The P19 protein of tomato bushy stunt virus (TBSV) is a multifunctional pathogenicity determinant involved in suppression of posttranscriptional gene silencing, virus movement, and symptom induction. Here, we report that P19 interacts with the conserved RNA-binding domain of an as yet uncharacterized family of plant ALY proteins that, in animals, are involved in export of RNAs from the nucleus and transcriptional coactivation. We show that the four ALY proteins encoded by the Arabidopsis genome and two ALY proteins from Nicotiana benthamiana are localized to the nucleus. Moreover, and in contrast to animal ALY, all but one of the proteins are also in the nucleolus, with distinct subnuclear localizations. Infection of plants by TBSV or expression of P19 from Agrobacterium results in relocation of three of the six ALY proteins from the nucleus to the cytoplasm demonstrating specific targeting of the ALY proteins by P19. The differential effects on subcellular localization indicate that, in plants, the various ALY proteins may have different functions. Interaction with and relocalization of ALY is prevented by mutation of P19 at residues previously shown to be important for P19 function in plants. Down-regulation of expression of two N. benthamiana ALY genes by virus-induced gene silencing did not interfere with posttranscriptional gene silencing. Targeting of ALY proteins during TBSV infection may therefore be related to functions of P19 in addition to its silencing suppression activity. PMID- 15299118 TI - Identification of syn-pimara-7,15-diene synthase reveals functional clustering of terpene synthases involved in rice phytoalexin/allelochemical biosynthesis. AB - Rice (Oryza sativa) produces momilactone diterpenoids as both phytoalexins and allelochemicals. Accordingly, the committed step in biosynthesis of these natural products is catalyzed by the class I terpene synthase that converts syn-copalyl diphosphate to the corresponding polycyclic hydrocarbon intermediate syn-pimara 7,15-diene. Here, a functional genomics approach was utilized to identify a syn copalyl diphosphate specific 9beta-pimara-7,15-diene synthase (OsDTS2). To our knowledge, this is the first identified terpene synthase with this particular substrate stereoselectivity and, by comparison with the previously described and closely related ent-copalyl diphosphate specific cassa-12,15-diene synthase (OsDTC1), provides a model system for investigating the enzymatic determinants underlying the observed difference in substrate specificity. Further, OsDTS2 mRNA in leaves is up-regulated by conditions that stimulate phytoalexin biosynthesis but is constitutively expressed in roots, where momilactones are constantly synthesized as allelochemicals. Therefore, transcription of OsDTS2 seems to be an important regulatory point for controlling production of these defensive compounds. Finally, the gene identified here as OsDTS2 has previously been mapped at 14.3 cM on chromosome 4. The class II terpene synthase producing syn-copalyl diphosphate from the universal diterpenoid precursor geranylgeranyl diphosphate was also mapped to this same region. These genes catalyze sequential cyclization steps in momilactone biosynthesis and seem to have been evolutionarily coupled by physical linkage and resulting cosegregation. Further, the observed correlation between physical proximity and common metabolic function indicates that other such class I and class II terpene synthase gene clusters may similarly catalyze consecutive reactions in shared biosynthetic pathways. PMID- 15299119 TI - Traffic lights in trichodesmium. Regulation of photosynthesis for nitrogen fixation studied by chlorophyll fluorescence kinetic microscopy. AB - We investigated interactions between photosynthesis and nitrogen fixation in the non-heterocystous marine cyanobacterium Trichodesmium IMS101 at the single-cell level by two-dimensional (imaging) microscopic measurements of chlorophyll fluorescence kinetics. Nitrogen fixation was closely associated with the appearance of cells with high basic fluorescence yield (F(0)), termed bright cells. In cultures aerated with normal air, both nitrogen fixation and bright cells appeared in the middle of the light phase. In cultures aerated with 5% oxygen, both processes occurred at a low level throughout most of the day. Under 50% oxygen, nitrogen fixation commenced at the beginning of the light phase but declined soon afterwards. Rapid reversible switches between fluorescence levels were observed, which indicated that the elevated F(0) of the bright cells originates from reversible uncoupling of the photosystem II (PSII) antenna from the PSII reaction center. Two physiologically distinct types of bright cells were observed. Type I had about double F(0) compared to the normal F(0) in the dark phase and a PSII activity, measured as variable fluorescence (F(v) = F(m) - F(0)), similar to normal non-diazotrophic cells. Correlation of type I cells with nitrogen fixation, oxygen concentration, and light suggests that this physiological state is connected to an up-regulation of the Mehler reaction, resulting in oxygen consumption despite functional PSII. Type II cells had more than three times the normal F(0) and hardly any PSII activity measurable by variable fluorescence. They did not occur under low-oxygen concentrations, but appeared under high-oxygen levels outside the diazotrophic period, suggesting that this state represents a reaction to oxidative stress not necessarily connected to nitrogen fixation. In addition to the two high-fluorescence states, cells were observed to reversibly enter a low-fluorescence state. This occurred mainly after a cell went through its bright phase and may represent a fluorescence-quenching recovery phase. PMID- 15299120 TI - The lys5 mutations of barley reveal the nature and importance of plastidial ADP Glc transporters for starch synthesis in cereal endosperm. AB - Much of the ADP-Glc required for starch synthesis in the plastids of cereal endosperm is synthesized in the cytosol and transported across the plastid envelope. To provide information on the nature and role of the plastidial ADP-Glc transporter in barley (Hordeum vulgare), we screened a collection of low-starch mutants for lines with abnormally high levels of ADP-Glc in the developing endosperm. Three independent mutants were discovered, all of which carried mutations at the lys5 locus. Plastids isolated from the lys5 mutants were able to synthesize starch at normal rates from Glc-1-P but not from ADP-Glc, suggesting a specific lesion in the transport of ADP-Glc across the plastid envelope. The major plastidial envelope protein was purified, and its sequence showed it to be homologous to the maize (Zea mays) ADP-Glc transporter BRITTLE1. The gene encoding this protein in barley, Hv.Nst1, was cloned, sequenced, and mapped. Like lys5, Hv.Nst1 lies on chromosome 6(6H), and all three of the lys5 alleles that were examined were shown to carry lesions in Hv.Nst1. Two of the identified mutations in Hv.Nst1 lead to amino acid substitutions in a domain that is conserved in all members of the family of carrier proteins to which Hv.NST1 belongs. This strongly suggests that Hv.Nst1 lies at the Lys5 locus and encodes a plastidial ADP-Glc transporter. The low-starch phenotype of the lys5 mutants shows that the ADP-Glc transporter is required for normal rates of starch synthesis. This work on Hv.NST1, together with the earlier work on BRITTLE1, suggests that homologous transporters are probably present in the endosperm of all cereals. PMID- 15299121 TI - Functional characterization of OsMADS18, a member of the AP1/SQUA subfamily of MADS box genes. AB - MADS box transcription factors controlling flower development have been isolated and studied in a wide variety of organisms. These studies have shown that homologous MADS box genes from different species often have similar functions. OsMADS18 from rice (Oryza sativa) belongs to the phylogenetically defined AP1/SQUA group. The MADS box genes of this group have functions in plant development, like controlling the transition from vegetative to reproductive growth, determination of floral organ identity, and regulation of fruit maturation. In this paper we report the functional analysis of OsMADS18. This rice MADS box gene is widely expressed in rice with its transcripts accumulated to higher levels in meristems. Overexpression of OsMADS18 in rice induced early flowering, and detailed histological analysis revealed that the formation of axillary shoot meristems was accelerated. Silencing of OsMADS18 using an RNA interference approach did not result in any visible phenotypic alteration, indicating that OsMADS18 is probably redundant with other MADS box transcription factors. Surprisingly, overexpression of OsMADS18 in Arabidopsis caused a phenotype closely resembling the ap1 mutant. We show that the ap1 phenotype is not caused by down-regulation of AP1 expression. Yeast two-hybrid experiments showed that some of the natural partners of AP1 interact with OsMADS18, suggesting that the OsMADS18 overexpression phenotype in Arabidopsis is likely to be due to the subtraction of AP1 partners from active transcription complexes. Thus, when compared to AP1, OsMADS18 during evolution seems to have conserved the mechanistic properties of protein-protein interactions, although it cannot complement the AP1 function. PMID- 15299122 TI - Novel regulation of aquaporins during osmotic stress. AB - Aquaporin protein regulation and redistribution in response to osmotic stress was investigated. Ice plant (Mesembryanthemum crystallinum) McTIP1;2 (McMIPF) mediated water flux when expressed in Xenopus leavis oocytes. Mannitol-induced water imbalance resulted in increased protein amounts in tonoplast fractions and a shift in protein distribution to other membrane fractions, suggesting aquaporin relocalization. Indirect immunofluorescence labeling also supports a change in membrane distribution for McTIP1;2 and the appearance of a unique compartment where McTIP1;2 is expressed. Mannitol-induced redistribution of McTIP1;2 was arrested by pretreatment with brefeldin A, wortmannin, and cytochalasin D, inhibitors of vesicle trafficking-related processes. Evidence suggests a role for glycosylation and involvement of a cAMP-dependent signaling pathway in McTIP1;2 redistribution. McTIP1;2 redistribution to endosomal compartments may be part of a homeostatic process to restore and maintain cellular osmolarity under osmotic stress conditions. PMID- 15299123 TI - Tobacco mosaic virus regulates the expression of its own resistance gene N. AB - The N gene of tobacco (Nicotiana tabacum) is a typical resistance (R) gene engendering localization of tobacco mosaic virus (TMV) infection and the elicitation of a hypersensitive necrotic response. The consensus model for R gene derived resistance is at the level of protein:protein interactions, in which proteins of the pathogen interact with already present receptor-like proteins produced by the plant's R genes. This article demonstrates, by quantitative real time reverse transcription-PCR analysis, that in tobacco carrying the dominant allele N, a basal level of transcription indeed occurs in noninfected plants. However, accumulation of N-mRNA in infected plants indicates that transcription is stimulated by TMV infection (up to 38-fold in locally infected leaves and up to 165-fold in upper, noninoculated leaves). Potato virus Y infection did not result in accumulation of N-mRNA, indicating a specific TMV-related phenomenon. The possible uncoupling of viral restriction from necrosis is discussed. PMID- 15299124 TI - Characterization of the complex locus of bean encoding polygalacturonase inhibiting proteins reveals subfunctionalization for defense against fungi and insects. AB - Polygalacturonase-inhibiting proteins (PGIPs) are extracellular plant inhibitors of fungal endopolygalacturonases (PGs) that belong to the superfamily of Leu-rich repeat proteins. We have characterized the full complement of pgip genes in the bean (Phaseolus vulgaris) genotype BAT93. This comprises four clustered members that span a 50-kb region and, based on their similarity, form two pairs (Pvpgip1/Pvpgip2 and Pvpgip3/Pvpgip4). Characterization of the encoded products revealed both partial redundancy and subfunctionalization against fungal-derived PGs. Notably, the pair PvPGIP3/PvPGIP4 also inhibited PGs of two mirid bugs (Lygus rugulipennis and Adelphocoris lineolatus). Characterization of Pvpgip genes of Pinto bean showed variations limited to single synonymous substitutions or small deletions. A three-amino acid deletion encompassing a residue previously identified as crucial for recognition of PG of Fusarium moniliforme was responsible for the inability of BAT93 PvPGIP2 to inhibit this enzyme. Consistent with the large variations observed in the promoter sequences, reverse transcription-PCR expression analysis revealed that the different family members differentially respond to elicitors, wounding, and salicylic acid. We conclude that both biochemical and regulatory redundancy and subfunctionalization of pgip genes are important for the adaptation of plants to pathogenic fungi and phytophagous insects. PMID- 15299126 TI - An in vivo analysis of the effect of season-long open-air elevation of ozone to anticipated 2050 levels on photosynthesis in soybean. AB - Rising atmospheric carbon dioxide concentration ([CO(2)]) is widely recognized, but less appreciated is a concomitant rise in tropospheric ozone concentration ([O(3)]). In industrialized countries, [O(3)] has risen by 0.5% to 2.5% per year. Tropospheric [O(3)] is predicted to reach a global mean of >60 nL L(-1) by 2050 with greater averages locally. Previous studies in enclosures suggest that this level of [O(3)] will decrease leaf photosynthesis, thereby limiting growth and yield of Glycine max L. Merr. SoyFACE (Soybean Free Air gas Concentration Enrichment) is the first facility to elevate atmospheric [O(3)] (approximately 1.2x current) in replicated plots under completely open-air conditions within an agricultural field. Measurements of gas exchange (assimilation versus light and assimilation versus intercellular [CO(2)]) were made on excised leaves from control and treatment plots (n = 4). In contrast to expectations from previous chamber studies, elevated [O(3)] did not alter light-saturated photosynthesis (A(sat), P = 0.09), carboxylation capacity (V(c,max), P = 0.82), or maximum electron transport (J(max), P = 0.66) for the topmost most recently fully expanded leaf at any stage of crop development. Leaves formed during the vegetative growth stage did not show a significant ozone-induced loss of photosynthetic capacity as they aged. Leaves formed during flowering did show a more rapid loss of photosynthetic capacity as they aged in elevated [O(3)]. A(sat), V(c,max), and J(max) (P = 0.04, 0.004, and 0.002, respectively) were decreased 20% to 30% by treatment with ozone. This is noteworthy since these leaves provide photosynthate to the developing grain. In conclusion, a small (approximately 20%) increase in tropospheric [O(3)] did not significantly alter photosynthetic capacity of newly expanded leaves, but as these leaves aged, losses in photosynthetic carbon assimilation occurred. PMID- 15299125 TI - Characterization of a root-specific Arabidopsis terpene synthase responsible for the formation of the volatile monoterpene 1,8-cineole. AB - Arabidopsis is emerging as a model system to study the biochemistry, biological functions, and evolution of plant terpene secondary metabolism. It was previously shown that the Arabidopsis genome contains over 30 genes potentially encoding terpene synthases (TPSs). Here we report the characterization of a monoterpene synthase encoded by two identical, closely linked genes, At3g25820 and At3g25830. Transcripts of these genes were detected almost exclusively in roots. An At3g25820/At3g25830 cDNA was expressed in Escherichia coli, and the protein thus produced was shown to catalyze the formation of 10 volatile monoterpenes from geranyl diphosphate, with 1,8-cineole predominating. This protein was therefore designated AtTPS-Cin. The purified recombinant AtTPS-Cin displayed similar biochemical properties to other known monoterpene synthases, except for a relatively low K(m) value for geranyl diphosphate of 0.2 microm. At3g25820/At3g25830 promoter activity, measured with a beta-glucuronidase (GUS) reporter gene, was primarily found in the epidermis, cortex, and stele of mature primary and lateral roots, but not in the root meristem or the elongation zone. Although the products of AtTPS-Cin were not detected by direct extraction of plant tissue, the recent report of 1,8-cineole as an Arabidopsis root volatile (Steeghs M, Bais HP, de Gouw J, Goldan P, Kuster W, Northway M, Fall R, Vivanco JM [2004] Plant Physiol 135: 47-58) suggests that the enzyme products may be released into the rhizosphere rather than accumulated. Among Arabidopsis TPSs, AtTPS-Cin is most similar to the TPS encoded by At3g25810, a closely linked gene previously shown to be exclusively expressed in flowers. At3g25810 TPS catalyzes the formation of a set of monoterpenes that is very similar to those produced by AtTPS-Cin, but its major products are myrcene and (E)-beta-ocimene, and it does not form 1,8-cineole. These data demonstrate that divergence of organ expression pattern and product specificity are ongoing processes within the Arabidopsis TPS family. PMID- 15299128 TI - Utility of different gene enrichment approaches toward identifying and sequencing the maize gene space. AB - Maize (Zea mays) possesses a large, highly repetitive genome, and subsequently a number of reduced-representation sequencing approaches have been used to try and enrich for gene space while eluding difficulties associated with repetitive DNA. This article documents the ability of publicly available maize expressed sequence tag and Genome Survey Sequences (GSSs; many of which were isolated through the use of reduced representation techniques) to recognize and provide coverage of 78 maize full-length cDNAs (FLCs). All 78 FLCs in the dataset were identified by at least three GSSs, indicating that the majority of maize genes have been identified by at least one currently available GSS. Both methyl-filtration and high-Cot enrichment methods provided a 7- to 8-fold increase in gene discovery rates as compared to random sequencing. The available maize GSSs aligned to 75% of the FLC nucleotides used to perform searches, while the expressed sequence tag sequences aligned to 73% of the nucleotides. Our data suggest that at least approximately 95% of maize genes have been tagged by at least one GSS. While the GSSs are very effective for gene identification, relatively few (18%) of the FLCs are completely represented by GSSs. Analysis of the overlap of coverage and bias due to position within a gene suggest that RescueMu, methyl-filtration, and high Cot methods are at least partially nonredundant. PMID- 15299127 TI - A novel auxin conjugate hydrolase from wheat with substrate specificity for longer side-chain auxin amide conjugates. AB - This study investigates how the ILR1-like indole acetic acid (IAA) amidohydrolase family of genes has functionally evolved in the monocotyledonous species wheat (Triticum aestivum). An ortholog for the Arabidopsis IAR3 auxin amidohydrolase gene has been isolated from wheat (TaIAR3). The TaIAR3 protein hydrolyzes negligible levels of IAA-Ala and no other IAA amino acid conjugates tested, unlike its ortholog IAR3. Instead, TaIAR3 has low specificity for the ester conjugates IAA-Glc and IAA-myoinositol and high specificity for the conjugates of indole-3-butyric acid (IBA-Ala and IBA-Gly) and indole-3-propionic-acid (IPA-Ala) so far tested. TaIAR3 did not convert the methyl esters of the IBA conjugates with Ala and Gly. IBA and IBA conjugates were detected in wheat seedlings by gas chromatography-mass spectrometry, where the conjugate of IBA with Ala may serve as a natural substrate for this enzyme. Endogenous IPA and IPA conjugates were not detected in the seedlings. Additionally, crude protein extracts of wheat seedlings possess auxin amidohydrolase activity. Temporal expression studies of TaIAR3 indicate that the transcript is initially expressed at day 1 after germination. Expression decreases through days 2, 5, 10, 15, and 20. Spatial expression studies found similar levels of expression throughout all wheat tissues examined. PMID- 15299129 TI - Transient release of oxygenated volatile organic compounds during light-dark transitions in Grey poplar leaves. AB - In this study, we investigated the prompt release of acetaldehyde and other oxygenated volatile organic compounds (VOCs) from leaves of Grey poplar [Populus x canescens (Aiton) Smith] following light-dark transitions. Mass scans utilizing the extremely fast and sensitive proton transfer reaction-mass spectrometry technique revealed the following temporal pattern after light-dark transitions: hexenal was emitted first, followed by acetaldehyde and other C(6)-VOCs. Under anoxic conditions, acetaldehyde was the only compound released after switching off the light. This clearly indicated that hexenal and other C(6)-VOCs were released from the lipoxygenase reaction taking place during light-dark transitions under aerobic conditions. Experiments with enzyme inhibitors that artificially increased cytosolic pyruvate demonstrated that the acetaldehyde burst after light-dark transition could not be explained by the recently suggested pyruvate overflow mechanism. The simulation of light fleck situations in the canopy by exposing leaves to alternating light-dark and dark-light transitions or fast changes from high to low photosynthetic photon flux density showed that this process is of minor importance for acetaldehyde emission into the Earth's atmosphere. PMID- 15299130 TI - Ovule abortion in Arabidopsis triggered by stress. AB - Environmental stresses frequently decrease plant fertility. In Arabidopsis, the effect of salt stress on reproduction was examined using plants grown in hydroponic medium. Salt stress inhibited microsporogenesis and stamen filament elongation. Because plants grown in hydroponic media can be rapidly and transiently stressed, the minimum inductive treatment to cause ovule abortion could be determined. Nearly 90% of the ovules aborted when roots were incubated for 12 h in a hydroponic medium supplemented with 200 mm NaCl. The anatomical effects of salt stress on maternal organs were distinct from those in the gametophyte. A fraction of cells in the chalaza and integuments underwent DNA fragmentation and programmed cell death. While three-fourths of the gametophytes aborted prior to fertilization, DNA fragmentation was not detected in these cells. Those gametophytes that survived were fertilized and formed embryos. However, very few of these developing embryos formed seeds; most senesced during seed development. Thus, during seed formation, there were multiple points where stress could prematurely terminate plant reproduction. These decreases in fecundity are discussed with respect to the hypothesis of serial adjustment of maternal investment. PMID- 15299131 TI - Brassinosteroids do not undergo long-distance transport in pea. Implications for the regulation of endogenous brassinosteroid levels. AB - It is widely accepted that brassinosteroids (BRs) are important regulators of plant growth and development. However, in comparison to the other classical plant hormones, such as auxin, relatively little is known about BR transport and its potential role in the regulation of endogenous BR levels in plants. Here, we show that end-pathway BRs in pea (Pisum sativum) occur in a wide range of plant tissues, with the greatest accumulation of these substances generally occurring in the young, actively growing tissues, such as the apical bud and young internodes. However, despite the widespread distribution of BRs throughout the plant, we found no evidence of long-distance transport of these substances between different plant tissues. For instance, we show that the maintenance of steady-state BR levels in the stem does not depend on their transport from the apical bud or mature leaves. Similarly, reciprocal grafting between the wild type and the BR-deficient lkb mutants demonstrates that the maintenance of steady state BR levels in whole shoots and roots does not depend on either basipetal or acropetal transport of BRs between these tissues. Together, with results from (3)H-BR feeding studies, these results demonstrate that BRs do not undergo long distance transport in pea. The widespread distribution of end-pathway BRs and the absence of long-distance BR transport between different plant tissues provide significant insight into the mechanisms that regulate BR homeostasis in plants. PMID- 15299132 TI - Roots, cycles and leaves. Expression of the phosphoenolpyruvate carboxylase kinase gene family in soybean. AB - Phosphorylation of phosphoenolpyruvate carboxylase (PEPc; EC 4.1.1.31) plays an important role in the control of central metabolism of higher plants. This phosphorylation is controlled largely at the level of expression of PEPc kinase (PPCK) genes. We have analyzed the expression of both PPCK genes and the PEPC genes that encode PEPc in soybean (Glycine max). Soybean contains at least four PPCK genes. We report the genomic and cDNA sequences of these genes and demonstrate the function of the gene products by in vitro expression and enzyme assays. For two of these genes, GmPPCK2 and GmPPCK3, transcript abundance is highest in nodules and is markedly influenced by supply of photosynthate from the shoots. One gene, GmPPCK4, is under robust circadian control in leaves but not in roots. Its transcript abundance peaks in the latter stages of subjective day, and its promoter contains a sequence very similar to the evening element found in Arabidopsis genes expressed at this time. We report the expression patterns of five PEPC genes, including one encoding a bacterial-type PEPc lacking the phosphorylation site of the plant-type PEPcs. The PEPc expression patterns do not match those of any of the PPCK genes, arguing against the existence of specific PEPc-PPCK expression partners. The PEPC and PPCK gene families in soybean are significantly more complex than previously understood. PMID- 15299133 TI - AtPng1p. The first plant transglutaminase. AB - Studies have revealed in plant chloroplasts, mitochondria, cell walls, and cytoplasm the existence of transglutaminase (TGase) activities, similar to those known in animals and prokaryotes having mainly structural roles, but no protein has been associated to this type of activity in plants. A recent computational analysis has shown in Arabidopsis the presence of a gene, AtPng1p, which encodes a putative N-glycanase. AtPng1p contains the Cys-His-Asp triad present in the TGase catalytic domain. AtPng1p is a single gene expressed ubiquitously in the plant but at low levels in all light-assayed conditions. The recombinant AtPng1p protein could be immuno-detected using animal TGase antibodies. Furthermore, western-blot analysis using antibodies raised against the recombinant AtPng1p protein have lead to its detection in microsomal fraction. The purified protein links polyamines-spermine (Spm) > spermidine (Spd) > putrescine (Put)-and biotin cadaverine to dimethylcasein in a calcium-dependent manner. Analyses of the gamma glutamyl-derivatives revealed that the formation of covalent linkages between proteins and polyamines occurs via the transamidation of gamma-glutamyl residues of the substrate, confirming that the AtPng1p gene product acts as a TGase. The Ca(2+)- and GTP-dependent cross-linking activity of the AtPng1p protein can be visualized by the polymerization of bovine serum albumine, obtained, like the commercial TGase, at basic pH and in the presence of dithiotreitol. To our knowledge, this is the first reported plant protein, characterized at molecular level, showing TGase activity, as all its parameters analyzed so far agree with those typically exhibited by the animal TGases. PMID- 15299134 TI - Proteome reference maps of vegetative tissues in pea. An investigation of nitrogen mobilization from leaves during seed filling. AB - A proteomic approach was used to analyze protein changes during nitrogen mobilization (N mobilization) from leaves to filling seeds in pea (Pisum sativum). First, proteome reference maps were established for mature leaves and stems. They displayed around 190 Coomassie Blue-stained spots with pIs from 4 to 7. A total of 130 spots were identified by mass spectrometry as corresponding to 80 different proteins implicated in a variety of cellular functions. Although the leaf proteome map contained more abundant spots, corresponding to proteins involved in energy/carbon metabolism, than the stem map, their comparison revealed a highly similar protein profile. Second, the leaf proteome map was used to analyze quantitative variations in leaf proteins during N mobilization. Forty percent of the spots showed significant changes in their relative abundance in the total protein extract. The results confirmed the importance of Rubisco as a source of mobilizable nitrogen, and suggested that in pea leaves the rate of degradation of Rubisco may vary throughout N mobilization. Correlated with the loss of Rubisco was an increase in relative abundance of chloroplastic protease regulatory subunits. Concomitantly, the relative abundance of some proteins related to the photosynthetic apparatus (Rubisco activase, Rubisco-binding proteins) and of several chaperones increased. A role for these proteins in the maintenance of a Rubisco activation state and in the PSII repair during the intense proteolytic activity within the chloroplasts was proposed. Finally, two 14-3-3-like proteins, with a potential regulatory role, displayed differential expression patterns during the massive remobilization of nitrogen. PMID- 15299135 TI - Types and frequencies of sequencing errors in methyl-filtered and high c0t maize genome survey sequences. AB - The Maize Genome Sequencing Consortium has deposited into GenBank more than 850,000 maize (Zea mays) genome survey sequences (GSSs) generated via two gene enrichment strategies, methylation filtration and high-C(0)t (HC) fractionation. These GSSs are a valuable resource for generating genome assemblies and the discovery of single nucleotide polymorphisms and nearly identical paralogs. Based on the rate of mismatches between 183 GSSs (105 methylation filtration + 78 HC) and 10 control genes, the rate of sequencing errors in these GSSs is 2.3 x 10( 3). As expected many of these errors were derived from insufficient vector trimming and base-calling errors. Surprisingly, however, some errors were due to cloning artifacts. These G.C to A.T transitions are restricted to HC clones; over 40% of HC clones contain at least one such artifact. Because it is not possible to distinguish the cloning artifacts from biologically relevant polymorphisms, HC sequences should be used with caution for the discovery of single nucleotide polymorphisms or paramorphisms. The average rate of sequencing errors was reduced 6-fold (to 3.6 x 10(-4)) by applying more stringent trimming parameters. This trimming resulted in the loss of only 11% of the bases (15,469/144,968). Due to redundancy among GSSs this more stringent trimming reduced coverage of promoters, exons, and introns by only 0%, 1%, and 4%, respectively. Hence, at the cost of a very modest loss of gene coverage, the quality of these maize GSSs can approach Bermuda standards, even prior to assembly. PMID- 15299136 TI - Differential antifungal and calcium channel-blocking activity among structurally related plant defensins. AB - Plant defensins are a family of small Cys-rich antifungal proteins that play important roles in plant defense against invading fungi. Structures of several plant defensins share a Cys-stabilized alpha/beta-motif. Structural determinants in plant defensins that govern their antifungal activity and the mechanisms by which they inhibit fungal growth remain unclear. Alfalfa (Medicago sativa) seed defensin, MsDef1, strongly inhibits the growth of Fusarium graminearum in vitro, and its antifungal activity is markedly reduced in the presence of Ca(2+). By contrast, MtDef2 from Medicago truncatula, which shares 65% amino acid sequence identity with MsDef1, lacks antifungal activity against F. graminearum. Characterization of the in vitro antifungal activity of the chimeras containing portions of the MsDef1 and MtDef2 proteins shows that the major determinants of antifungal activity reside in the carboxy-terminal region (amino acids 31-45) of MsDef1. We further define the active site by demonstrating that the Arg at position 38 of MsDef1 is critical for its antifungal activity. Furthermore, we have found for the first time, to our knowledge, that MsDef1 blocks the mammalian L-type Ca(2+) channel in a manner akin to a virally encoded and structurally unrelated antifungal toxin KP4 from Ustilago maydis, whereas structurally similar MtDef2 and the radish (Raphanus sativus) seed defensin Rs-AFP2 fail to block the L-type Ca(2+) channel. From these results, we speculate that the two unrelated antifungal proteins, KP4 and MsDef1, have evolutionarily converged upon the same molecular target, whereas the two structurally related antifungal plant defensins, MtDef2 and Rs-AFP2, have diverged to attack different targets in fungi. PMID- 15299137 TI - Graft transmission of a floral stimulant derived from CONSTANS. AB - Photoperiod in plants is perceived by leaves and in many species influences the transition to reproductive growth through long-distance signaling. CONSTANS (CO) is implicated as a mediator between photoperiod perception and the transition to flowering in Arabidopsis. To test the role of CO in long-distance signaling, CO was expressed from a promoter specific to the companion cells of the smallest veins of mature leaves. This expression in tissues at the inception of the phloem translocation stream was sufficient to accelerate flowering at the apical meristem under noninductive (short-day) conditions. Grafts that conjoined the vegetative stems of plants with different flower-timing phenotypes demonstrated that minor-vein expression of CO is able to substitute for photoperiod in generating a mobile flowering signal. Our results suggest that a CO-derived signal(s), or possibly CO itself, fits the definition of the hypothetical flowering stimulant, florigen. PMID- 15299138 TI - Disorganization of cortical microtubules stimulates tangential expansion and reduces the uniformity of cellulose microfibril alignment among cells in the root of Arabidopsis. AB - To test the role of cortical microtubules in aligning cellulose microfibrils and controlling anisotropic expansion, we exposed Arabidopsis thaliana roots to moderate levels of the microtubule inhibitor, oryzalin. After 2 d of treatment, roots grow at approximately steady state. At that time, the spatial profiles of relative expansion rate in length and diameter were quantified, and roots were cryofixed, freeze-substituted, embedded in plastic, and sectioned. The angular distribution of microtubules as a function of distance from the tip was quantified from antitubulin immunofluorescence images. In alternate sections, the overall amount of alignment among microfibrils and their mean orientation as a function of position was quantified with polarized-light microscopy. The spatial profiles of relative expansion show that the drug affects relative elongation and tangential expansion rates independently. The microtubule distributions averaged to transverse in the growth zone for all treatments, but on oryzalin the distributions became broad, indicating poorly organized arrays. At a subcellular scale, cellulose microfibrils in oryzalin-treated roots were as well aligned as in controls; however, the mean alignment direction, while consistently transverse in the controls, was increasingly variable with oryzalin concentration, meaning that microfibril orientation in one location tended to differ from that of a neighboring location. This conclusion was confirmed by direct observations of microfibrils with field-emission scanning electron microscopy. Taken together, these results suggest that cortical microtubules ensure microfibrils are aligned consistently across the organ, thereby endowing the organ with a uniform mechanical structure. PMID- 15299139 TI - Organ polarity in Arabidopsis. NOZZLE physically interacts with members of the YABBY family. AB - Plant lateral organs exhibit proximal-distal and adaxial-abaxial polarity. In Arabidopsis, abaxial cell fate is regulated in part by putative transcription factors of the YABBY family, such as FILAMENTOUS FLOWER (FIL) and INNER NO OUTER (INO), by a mechanism that currently is not fully understood. NOZZLE (NZZ) encodes a plant-specific nuclear protein. Genetic evidence has shown that NZZ is involved in the positive feedback regulation of INO, thereby acting both as a temporal and spatial repressor of INO transcription. This mechanism allows the ovule primordium to complete its proximal-distal organization, prior to the onset of adaxial-abaxial development in the chalaza. During our study, we isolated FIL in a yeast two-hybrid screen using NZZ as bait. In vitro pull-down experiments confirmed the NZZ-FIL interaction. NZZ also bound INO and YABBY3, suggesting that NZZ generally interacts with YABBY proteins in vitro. The polar-charged region of NZZ was necessary and sufficient to bind to the zinc finger of INO and to interact with its C terminus carrying the high mobility group-like domain. We suggest that NZZ coordinates proximal-distal patterning and adaxial-abaxial polarity establishment in the developing ovule by directly binding to INO. PMID- 15299140 TI - High genetic variability of herbivore-induced volatile emission within a broad range of maize inbred lines. AB - Maize plants (Zea mays) attacked by caterpillars release a mixture of odorous compounds that attract parasitic wasps, natural enemies of the herbivores. We assessed the genetic variability of these induced volatile emissions among 31 maize inbred lines representing a broad range of genetic diversity used by breeders in Europe and North America. Odors were collected from young plants that had been induced by injecting them with caterpillar regurgitant. Significant variation among lines was found for all 23 volatile compounds included in the analysis: the lines differed enormously in the total amount of volatiles emitted and showed highly variable odor profiles distinctive of each genotype. Principal component analysis performed on the relative quantities of particular compounds within the blend revealed clusters of highly correlated volatiles, which may share common metabolic pathways. European and American lines belonging to established heterotic groups were loosely separated from each other, with the most clear-cut difference in the typical release of (E)-beta-caryophyllene by European lines. There was no correlation between the distances among the lines based on their odor profiles and their respective genetic distances previously assessed by neutral RFLP markers. This most comprehensive study to date on intraspecific variation in induced odor emission by maize plants provides a further example of the remarkably high genetic diversity conserved within this important crop plant. A better understanding of the genetic control of induced odor emissions may help in the development of maize varieties particularly attractive to parasitoids and other biological control agents and perhaps more repellent for herbivores. PMID- 15299143 TI - A detailed picture of the origin of the Australian dingo, obtained from the study of mitochondrial DNA. AB - To determine the origin and time of arrival to Australia of the dingo, 582 bp of the mtDNA control region were analyzed in 211 Australian dingoes sampled in all states of Australia, 676 dogs from all continents, and 38 Eurasian wolves, and 263 bp were analyzed in 19 pre-European archaeological dog samples from Polynesia. We found that all mtDNA sequences among dingoes were either identical to or differing by a single substitution from a single mtDNA type, A29. This mtDNA type, which was present in >50% of the dingoes, was found also among domestic dogs, but only in dogs from East Asia and Arctic America, whereas 18 of the 19 other types were unique to dingoes. The mean genetic distance to A29 among the dingo mtDNA sequences indicates an origin approximately 5,000 years ago. From these results a detailed scenario of the origin and history of the dingo can be derived: dingoes have an origin from domesticated dogs coming from East Asia, possibly in connection with the Austronesian expansion into Island Southeast Asia. They were introduced from a small population of dogs, possibly at a single occasion, and have since lived isolated from other dog populations. PMID- 15299142 TI - Methyl jasmonate-induced ethylene production is responsible for conifer phloem defense responses and reprogramming of stem cambial zone for traumatic resin duct formation. AB - Conifer stem pest resistance includes constitutive defenses that discourage invasion and inducible defenses, including phenolic and terpenoid resin synthesis. Recently, methyl jasmonate (MJ) was shown to induce conifer resin and phenolic defenses; however, it is not known if MJ is the direct effector or if there is a downstream signal. Exogenous applications of MJ, methyl salicylate, and ethylene were used to assess inducible defense signaling mechanisms in conifer stems. MJ and ethylene but not methyl salicylate caused enhanced phenolic synthesis in polyphenolic parenchyma cells, early sclereid lignification, and reprogramming of the cambial zone to form traumatic resin ducts in Pseudotsuga menziesii and Sequoiadendron giganteum. Similar responses in internodes above and below treated internodes indicate transport of a signal giving a systemic response. Studies focusing on P. menziesii showed MJ induced ethylene production earlier and 77-fold higher than wounding. Ethylene production was also induced in internodes above the MJ-treated internode. Pretreatment of P. menziesii stems with the ethylene response inhibitor 1-methylcyclopropene inhibited MJ and wound responses. Wounding increased 1-aminocyclopropane-1-carboxylic acid (ACC) oxidase protein, but MJ treatment produced a higher and more rapid ACC oxidase increase. ACC oxidase was most abundant in ray parenchyma cells, followed by cambial zone cells and resin duct epithelia. The data show these MJ-induced defense responses are mediated by ethylene. The cambial zone xylem mother cells are reprogrammed to differentiate into resin-secreting epithelial cells by an MJ-induced ethylene burst, whereas polyphenolic parenchyma cells are activated to increase polyphenol production. The results also indicate a central role of ray parenchyma in ethylene-induced defense. PMID- 15299145 TI - Highly efficient gene replacements in Neurospora strains deficient for nonhomologous end-joining. AB - Gene disruption and overexpression play central roles in the analysis of gene function. Homologous recombination is, in principle, the most efficient method of disrupting, modifying, or replacing a target gene. Although homologous integration of exogenous DNA into the genome occurs readily in Saccharomyces cerevisiae, it is rare in many other organisms. We identified and disrupted Neurospora crassa genes homologous to human KU70 and KU80, which encode proteins that function in nonhomologous end-joining of double-stranded DNA breaks. The resulting mutants, named mus-51 and mus-52, showed higher sensitivity to methyl methanesulfonate, ethyl methanesulfonate, and bleomycin than wild type, but not to UV, 4-nitroquinoline 1-oxide, camptothecin, or hydroxyurea. Vegetative growth, conidiation, and ascospore production in homozygous crosses were normal. The frequency of integration of exogenous DNA into homologous sequences of the genome in the KU disruption strains of N. crassa was compared with that in wild type, mei-3, and mus-11. In mei-3 and mus-11, which are defective in homologous recombination, none or few homologous integration events were observed under any conditions. When mtr target DNA with approximately 2-kb 5' and 3' flanking regions was used for transformation of the KU disruption strains, 100% of transformants exhibited integration at the homologous site, compared to 10 to 30% for a wild-type recipient. Similar results were obtained when the ad-3A gene was targeted for disruption. These results indicate that KU disruption strains are efficient recipients for gene targeting. PMID- 15299144 TI - A mechanical basis for chromosome function. AB - We propose that chromosome function is governed by internal mechanical forces generated by programmed tendencies for expansion of the DNA/chromatin fiber against constraining features. PMID- 15299146 TI - Total synthesis of (+/-)-halichlorine, (+/-)-pinnaic acid, and (+/-)-tauropinnaic acid. AB - The related marine natural products halichlorine, pinnaic acid, and tauropinnaic acid have been synthesized. The described route provided access to all three compounds from a common, late-stage intermediate. The synthesis began with 1 pyrrolidino-1-cyclopentene from which an intermediate possessing the three contiguous stereocenters of the natural products was synthesized in just four steps. Olefin cross metathesis followed by a hydrogenation/hydrogenolysis reaction stereoselectively formed the piperidine ring. Use of a beta-lactam group provided internal protection for the highly congested nitrogen atom during side chain elaboration. The beta-lactam was subsequently reduced directly to an amino aldehyde, which after the Horner-Wadsworth-Emmons reaction was elaborated to pinnaic acid. The same amino aldehyde was also transformed into halichlorine after a thiol-mediated cyclization sequence to form the dehydroquinolizidine ring system. PMID- 15299141 TI - Coordinated genetic regulation of growth and lignin revealed by quantitative trait locus analysis of cDNA microarray data in an interspecific backcross of eucalyptus. AB - Phenotypic, genotypic, and transcript level (microarray) data from an interspecific backcross population of Eucalyptus grandis and Eucalyptus globulus were integrated to dissect the genetic and metabolic network underlying growth variation. Transcript abundance, measured for 2,608 genes in the differentiating xylem of a 91 (E. grandis x E. globulus) x E. grandis backcross progeny was correlated with diameter variation, revealing coordinated down-regulation of genes encoding enzymes of the lignin biosynthesis and associated methylation pathways in fast growing individuals. Lignin analysis of wood samples confirmed the content and quality predicted by the transcript levels measured on the microarrays. Quantitative trait locus (QTL) analysis of transcript levels of lignin-related genes showed that their mRNA abundance is regulated by two genetic loci, demonstrating coordinated genetic control over lignin biosynthesis. These two loci colocalize with QTLs for growth, suggesting that the same genomic regions are regulating growth, and lignin content and composition in the progeny. Genetic mapping of the lignin genes revealed that most of the key biosynthetic genes do not colocalize with growth and transcript level QTLs, with the exception of the locus encoding the enzyme S-adenosylmethionine synthase. This study illustrates the power of integrating quantitative analysis of gene expression data and genetic map information to discover genetic and metabolic networks regulating complex biological traits. (Sequence data for this article have been deposited with the EMBL/GenBank data libraries under accession numbers CB 967505 to CB 968059; CD 667988 to CD 670002; CD 670004; CD 670097; CD 670101 to CD 670112; and CD 670114 to CD 670137.) PMID- 15299147 TI - K+ channel interactions detected by a genetic system optimized for systematic studies of membrane protein interactions. AB - Organization of proteins into complexes is crucial for many cellular functions. However, most proteomic approaches primarily detect protein interactions for soluble proteins but are less suitable for membrane-associated complexes. Here we describe a mating-based split ubiquitin system (mbSUS) for systematic identification of interactions between membrane proteins as well as between membrane and soluble proteins. mbSUS allows in vivo cloning of PCR products into a vector set, detection of interactions via mating, regulated expression of baits, and improved selection of interacting proteins. Cloning is simplified by introduction of lambda attachment sites for GATEWAY. Homo- and heteromeric interactions between Arabidopsis K(+) channels KAT1, AKT1, and AKT2 were identified. Tests with deletion mutants demonstrate that the C terminus of KAT1 and AKT1 is necessary for physical assembly of complexes. Screening of a sorted collection of 84 plant proteins with K(+) channels as bait revealed differences in oligomerization between KAT1, AKT1, and AtKC1, and allowed detection of putative interacting partners of KAT1 and AtKC1. These results show that mbSUS is suited for systematic analysis of membrane protein interactions. PMID- 15299149 TI - Tissue-specific insulin resistance. PMID- 15299148 TI - Structure of the photolyase-like domain of cryptochrome 1 from Arabidopsis thaliana. AB - Signals generated by cryptochrome (CRY) blue-light photoreceptors are responsible for a variety of developmental and circadian responses in plants. The CRYs are also identified as circadian blue-light photoreceptors in Drosophila and components of the mammalian circadian clock. These flavoproteins all have an N terminal domain that is similar to photolyase, and most have an additional C terminal domain of variable length. We present here the crystal structure of the photolyase-like domain of CRY-1 from Arabidopsis thaliana. The structure reveals a fold that is very similar to photolyase, with a single molecule of FAD noncovalently bound to the protein. The surface features of the protein and the dissimilarity of a surface cavity to that of photolyase account for its lack of DNA-repair activity. Previous in vitro experiments established that the photolyase-like domain of CRY-1 can bind Mg.ATP, and we observe a single molecule of an ATP analog bound in the aforementioned surface cavity, near the bound FAD cofactor. The structure has implications for the signaling mechanism of CRY blue light photoreceptors. PMID- 15299150 TI - Organising an English journal club in the developing world. PMID- 15299151 TI - Repetitive strain injury. AB - Pain in the forearm is relatively common in the community. In the workplace forearm pain is associated with work involving frequent repetition, high forces, and prolonged abnormal postures. Nevertheless, other factors are involved in the presentation and the continuation of the pain. Notable among these factors are psychosocial issues and the workplace environment-the attitude to workers and their welfare, the physical conditions, and design of the job. Primary prevention may be effective but active surveillance is important with early intervention and an active management approach. Physical treatments have not been extensively evaluated. In the established case, management should be multidisciplinary, addressing physical aspects of the job but also addressing the "yellow, blue, and black flags" which should be viewed as obstacles to recovery. For the worker "on sick" a dialogue should be established between the worker, the primary care physician, and the workplace. Return to work should be encouraged and facilitated by medical interventions and light duty options. Rehabilitation programmes may be of use in chronic cases. PMID- 15299152 TI - Introduction to history taking and principles of sexual health. AB - Sexual health has a profound influence over an individual's physical and psychological wellbeing. There is an increasing need to develop effective strategies to reduce the incidence of sexually transmitted infections (STIs) globally. Many STIs are asymptomatic and yet, highly infectious. The sequelae of such infections may be catastrophic and hence, an integrated multidisciplinary and preventative approach to the control of STIs is clearly necessary. Sexual history taking holds the key to the practice of sexual health medicine. It provides the basis for treatment, prevention, education, and sexual health promotion. PMID- 15299153 TI - Probiotics and prebiotics in the elderly. AB - Probiotics (usually lactobacilli and bifidobacteria) and prebiotics (non digestible oligosaccharides) have been shown to be useful in preventing certain disease conditions as well as possibly promoting specific aspects of health. In the present review, the evidence from clinical trials for benefits from probiotics and prebiotics to elderly populations is presented and discussed, specifically in respect of three common conditions found in the elderly. Both probiotics and prebiotics may be helpful in malnutrition, particularly in lactose intolerance and calcium absorption, and in constipation. Probiotics have been shown clearly to boost immunity in the elderly, but the clinical significance of this remains to be clarified. These results are encouraging, and further large scale studies seem justified to establish the place of probiotic and prebiotic supplements in elderly subjects. PMID- 15299154 TI - Long term motor complications of levodopa: clinical features, mechanisms, and management strategies. AB - Levodopa is the most effective symptomatic treatment of Parkinson's disease. However, after an initial period of dramatic benefit, several limitations become apparent including, "dopa resistant" motor symptoms (postural abnormalities, freezing episodes, speech impairment), "dopa resistant" non-motor signs (autonomic dysfunction, mood and cognitive impairment, etc), and/or drug related side effects (especially psychosis, motor fluctuations, and dyskinesias). Motor complications include fluctuations, dyskinesias, and dystonias. They can be very disabling and difficult to treat. Therefore, strategies should ideally be developed to prevent them. Though mechanisms underlying motor complications are only partially understood, recent work has revealed the importance of pulsatile stimulation of postsynaptic dopamine receptors and the disease severity. As a result of intermittent stimulation there occurs a cascade of changes in cell signalling leading to upregulation of the N-methyl-D-aspartate subtype of gamma aminobutryric acid-ergic neurones. Modified preparations of levodopa (controlled release preparations, liquid levodopa), catecholamine-o-methyltransferase inhibitors, dopamine agonists, amantidine, and various neurosurgical approaches have been used in the prevention and/or treatment of motor complications. Current management of motor complications is less than satisfactory. With better understanding of the pathogenetic mechanisms, it is hoped that future therapeutic strategies will provide a safer and targeted treatment. PMID- 15299155 TI - Iliopsoas abscesses. AB - Iliopsoas abscess is a relatively uncommon condition that can present with vague clinical features. Its insidious onset and occult characteristics can cause diagnostic delays, resulting in high mortality and morbidity. The epidemiology, aetiology, clinical features, and management of iliopsoas abscess are discussed. PMID- 15299156 TI - History of medical screening: from concepts to action. AB - The objective of medical screening is to identify disease in its preclinical, and therefore hopefully still curable, phase. This may have been an old quest in medicine but it became historically possible when at least four conditions were met: the availability of simple, valid and acceptable forms of tests, the discovery of effective treatments, the establishment of a theory of screening, and the wide access to health care. Five selected examples that illustrate the history of medical screening are reviewed: screening for psychiatric disorders in the United States army as it is one of the oldest screening programmes; screening for syphilis as it used one of the earliest screening tests; screening for diabetes as one of the first modern forms of mass screening; screening for cervical cancer using the Pap test as one of the greatest successes of screening; and screening for breast cancer by mammography as this offers a good opportunity to discuss the development of modern evaluation of screening programmes. The evaluation of the impact of screening on human health slowly progressed, from obvious changes in the vital statistics such as the decline in incidence of syphilis, to less obvious changes such as the decline in mortality of cancer of the uterus, to finally more subtle changes, such as the impact of mammographic screening on breast cancer mortality. Methods of evaluation had therefore to adapt, evolving from simple surveys to case-control studies and randomised trials. The history of screening is short, but very rich and mostly still to be written. PMID- 15299157 TI - Patient and disease profile of emergency medical readmissions to an Irish teaching hospital. AB - OBJECTIVE: To determine whether there was a relationship between coded diseases at the time of hospital discharge, a pattern of ordering investigations, and hospital readmission in a major teaching hospital. DESIGN: Systematic review of data relating to emergency medical patients admitted to St James' Hospital Dublin between 1 January and 31 December 2002. DATA SOURCES AND METHODS: Data on discharges from hospital recorded in the Hospital In-Patient Enquiry (HIPE) system. The value of HIPE data in describing the relationship between the pattern of resource utilisation, diagnostic related groups, and hospital readmission has not previously been examined. RESULTS: Of 5038 episodes recorded among 4050 patients admitted, the number of readmissions was up to 15. Age and male gender were factors associated with readmission, and readmitted patients remained in hospital for longer. No particular test request predicted readmission, but computed tomography of the brain was associated with a reduced readmission rate. Discharge diagnostic related group coding at first discharge predicted readmission-codes related to heart failure, respiratory system, alcohol, malignancy, and anaemia. CONCLUSIONS: It was found that clinical coding using the HIPE database strongly predicted hospital readmission. It may be argued that early hospital readmission reflects unsatisfactory patient care, alternatively that many readmissions are not preventable, representing either new events in elderly patients with chronic illnesses and frequent co-morbidity or related to social factors. The utility of specific interventions, in patients at high risk for hospital readmission, could be explored. PMID- 15299159 TI - Evaluation of the role of a specialist tracheostomy service. From critical care to outreach and beyond. AB - The impact that a new specialist tracheostomy service, designed specifically for the care of patients with tracheostomies, was assessed in terms of type of tracheostomy tube used, time to first tube change, time to decannulation, and incidence of tracheostomy related complications in a teaching hospital with no on site ear, nose, and throat facility. A total of 170 patients were studied. After service implementation, fewer patients (17.6%, n = 21) were discharged from the intensive treatment unit to the wards with tracheostomy tubes compared with the first group (39%, n = 20) (p = 0.006), and the number of tracheostomy related complications on the wards were significantly reduced (p = 0.031). PMID- 15299158 TI - Value of routine duodenal biopsy in diagnosing coeliac disease in patients with iron deficiency anaemia. AB - BACKGROUND: Iron deficiency anaemia (IDA) is a recognised feature of coeliac disease in adults and can be its only presentation. OBJECTIVE: To determine the yield of routine distal duodenal biopsies in diagnosing coeliac disease in adult and elderly patients with IDA whose endoscopy revealed no upper gastrointestinal cause of iron deficiency. STUDY DESIGN: Prospective study in a teaching hospital endoscopy unit. METHOD: Altogether 504 consecutive patients with IDA, aged 16-80 years, attending for endoscopy were included in this study. At least two distal duodenal biopsies were taken if endoscopy revealed no cause of iron deficiency. RESULT: In nine (1.8%) patients duodenal biopsies revealed typical histological features of coeliac disease. Of these, five patients were above 65 years old. CONCLUSION: In adult and elderly patients undergoing endoscopy for IDA, the endoscopist should take distal duodenal biopsies to exclude coeliac disease if no upper gastrointestinal cause of anaemia is found. Coeliac disease is not an uncommon cause of IDA in patients >65 years of age and a history of chronic diarrhoea increases diagnostic yield in this age group. PMID- 15299160 TI - Observation and identification of lactate dehydrogenase anomaly in a postburn patient. AB - OBJECTIVE: Lactate dehydrogenase (LDH) anomaly is one of the macroenzymes. Macroenzymes are enzymes in serum that have formed high molecular mass complexes, either by self polymerisation or by association with other serum components. The aim of this study was to identify the properties of LDH anomaly and observe the changes from admission to discharge in a postburn patient with LDH anomaly in his serum. METHODS: LDH isoenzymes of the serum were electrophoretically fractionated with terylene cellulose acetate supporting media; LDH anomaly was identified by counter immunoelectrophoresis. RESULTS: An abnormal LDH-4 band and an extra band on the cathode of LDH-5 were observed in the serum of this patient and were found to be part of an LDH-IgG complex. As his symptoms improved, the patient's LDH anomaly gradually disappeared. The appearance and disappearance of the anomaly seemed to be related to the progression of the patient's burns. CONCLUSION: In clinical practice, it is important to keep in mind the possibility of an LDH anomaly in patients when the LDH level is abnormally high or does not seem to be related to the clinical state. Early discovery of an LDH anomaly in a patient's serum may be useful for diagnosis and treatment. PMID- 15299161 TI - A case of two adverse reactions. AB - Low molecular weight heparins are now widely prescribed in the treatment of thromboembolic disease and acute coronary syndromes. Anaphylaxis is a recognised but rare potentially life threatening side effect of heparin. Common clinical features of anaphylaxis are cardiovascular collapse, bronchospasm, cutaneous symptoms, angioedema, generalised oedema, or gastrointestinal symptoms. It is extremely rare, however, for patients to experience such dramatic and potentially life threatening consequences as seen in the case reported here. It has been shown that patients may be tolerant of certain low molecular weight heparins but sensitive to others. Adrenaline is regarded as the most important drug for any severe anaphylactic reaction. Administration by the intramuscular route produces significantly higher peak plasma concentrations compared with subcutaneous injection, which is clearly beneficial in the critically compromised patient. Current UK Resuscitation Council guidelines support the use of 0.5 ml of 1:1000 adrenaline to be administered intramuscularly. PMID- 15299162 TI - Life threatening hyperphosphataemia after administration of sodium phosphate in preparation for colonoscopy. AB - An elderly woman developed severe hyperphosphataemia, hypocalcaemia, and cardiac arrest after oral administration of sodium phosphate in preparation for colonoscopy. This is an unusual complication and is attributed to decreased phosphate excretion by the kidneys. At increased risk are patients with impaired renal function, age more than 65 years, and presenting with intestinal obstruction or decreased intestinal motility, increased intestinal permeability, liver cirrhosis, or congestive heart failure. Though there are no accepted guidelines for anticipation and prevention of this adverse effect, it may be desirable to check serum phosphate concentrations before choosing the method for colonic preparation and before giving the second oral dose of sodium phosphate in patients at risk. Hyperphosphataemia should be suspected if a patient develops hypotension or neuromuscular irritability after administration of sodium phosphate. Haemodialysis for direct removal of phosphate and intravenous calcium for treatment of symptomatic hypocalcaemia may be life saving. PMID- 15299164 TI - Pigmented sclera: a diagnostic challenge? PMID- 15299163 TI - IgA multiple myeloma presenting as non-obstructive jaundice. AB - Multiple myeloma can occasionally present with jaundice. The underlying process may be pancreatic head myeloma infiltration causing obstructive jaundice or hepatic amyloid deposition resulting in cholestatic jaundice. A rare case of myeloma presenting as jaundice due to hepatic myeloma infiltration is reported. PMID- 15299165 TI - Upper gastrointestinal haemorrhage. PMID- 15299166 TI - Hereditary spinocerebellar ataxias: number, prevalence, and treatment prospects. PMID- 15299167 TI - Chronic benign neutropenia among Chinese children. AB - OBJECTIVE: To delineate the clinical behaviour of chronic benign neutropenia in Chinese children in Hong Kong. DESIGN: Retrospective study. SETTING: University teaching hospital, Hong Kong. PATIENTS: All infants and children with absolute neutrophil count of 1.5 x 10(9) /L or lower for more than 3 months. MAIN OUTCOME MEASURES: Development of significant infection, and achievement of remission. RESULTS: Twenty-four children with chronic benign neutropenia were identified between 1992 and 2001. Their median age of diagnosis was 9 months. The mean (standard deviation) initial absolute neutrophil count was 0.28 x 10(9) /L (0.24 x 10(9) /L). Twenty-three patients presented with infection. Of the 19 patients tested, four (21%) were positive for anti-neutrophil antibodies. Bone marrow examination was performed in 17 patients: nine had normal results, but six showed evidence of peripheral consumption, one showed late maturation arrest at band stage, and one showed phagocytosis of myeloid cells by histiocytes. The overall hospitalised infection rate was 51.6 episodes per 1000 patient-months. Ten percent of cases were considered 'significant' infections and required hospital admission with either surgical intervention or intravenous therapy (antibiotics or fluid replacement). In the first year of diagnosis, more than 80% of patients had their lowest absolute neutrophil count (mean, 0.16 x 10(9) /L; standard deviation, 0.11 x 10(9) /L). Granulocyte-colony stimulating factor was used to treat three patients and induced transient elevation of absolute neutrophil count in all three. The projected remission rate was 55.4% at 3 years. Even for those with persistent disease, there was significant recovery in absolute neutrophil count to a mean of 0.5 x 10(9) /L (P<0.01). CONCLUSIONS: Patients with chronic benign neutropenia experienced a relatively benign clinical course regardless of their remission status. Only a small proportion of patients developed significant infections. A multi-centre prospective study may help identify predictive factors of remission. PMID- 15299168 TI - Treatment of early rectal tumours by transanal endoscopic microsurgery in Hong Kong: prospective study. AB - OBJECTIVE: To summarise the results of transanal endoscopic microsurgery for the treatment of rectal villous adenoma and early rectal tumours. DESIGN: Prospective study. SETTING: Regional hospital, Hong Kong. PATIENTS: Consecutive patients between November 1995 and January 2003. INTERVENTION: Transanal endoscopic microsurgery. MAIN OUTCOME MEASURES: Intra-operative morbidity and mortality, complication rate, operating time, postoperative morbidity and mortality, recurrence rate and correlation between preoperative ultrasonography staging and postoperative pathological staging. RESULTS: Thirty-two patients with rectal villous adenoma and early rectal carcinoma were registered, 31 of whom (14 men and 17 women) were included in the study. The median tumour size was 2.5 (range, 1-8) cm and the median operating time was 95 (45-220) minutes. The median follow up period was 23 (2-92) months, and there was no local recurrence. There was no operation-related mortality and the resection margins were all clear. Complications included temporary flatus incontinence (n=2), acute retention of urine (n=1), exacerbation of chronic obstructive airway disease (n=1), and secondary haemorrhage in a patient on aspirin. CONCLUSIONS: Transanal endoscopic microsurgery is a safe procedure and can achieve good local tumour control. It is ideal in the management of rectal villous adenomas at stages pT0 and pTis. Its application is now extended to the treatment of early rectal carcinoma at stage pT1 with curative intent. For tumours at stage pT2 or later, it can also serve as a good option for local palliation. PMID- 15299169 TI - Ethical attitudes of intensive care physicians in Hong Kong: questionnaire survey. AB - OBJECTIVES: To examine the practice and ethical attitudes of intensive care doctors in Hong Kong and to compare findings with those from European studies. DESIGN: Structured questionnaire survey, modified from a similar questionnaire used in Europe. SETTING: Eleven publicly funded intensive care units in Hong Kong. PARTICIPANTS: Ninety-five doctors practising in intensive care units. RESULTS: Of the sixty-five respondents, sizeable proportions indicated that the admission of patients to the intensive care unit is often (25%) or sometimes (51%) limited by bed availability. About 69% to 86% of doctors admit patients with limited prognosis or poor quality of life, although all felt that these admissions should be more restricted. 'Do-not-resuscitate' orders are applied by almost all respondents, and 52% and 89% of respondents would discuss such orders with the patient or with the family, respectively. The withholding and withdrawal of therapy from patients with no chance of recovery to a meaningful life is common in Hong Kong (99% and 89%, respectively). A total of 83% respondents involved patients or families in the decision to limit therapy, compared with less than half in Europe overall. When the family wanted aggressive life-support despite doctors' recommendations to limit therapy, 62% of the respondents would still withhold therapy while only 9% would withdraw therapy. More than 60% of doctors feel comfortable talking to patients' relatives about limitation of therapy. Approximately 75% felt that euthanasia is unacceptable. Most respondents (94%) reported that medical programmes should include more extensive discussion on ethical issues. CONCLUSION: The ethical attitudes of intensive care doctors in Hong Kong are similar to those of counterparts in Europe. However, Hong Kong doctors tend to involve families more often in the discussion of end-of-life issues. PMID- 15299170 TI - Correlation between serum level of neuron-specific enolase and long-term functional outcome after acute cerebral infarction: prospective study. AB - OBJECTIVE: To determine the value of measuring serum levels of neuron-specific enolase in predicting extent of disease and short- and long-term functional outcome after acute cerebral infarction. DESIGN: Prospective study. SETTING: Neurology departments at two university teaching hospitals, Shanghai. PATIENTS: Thirty-eight patients who presented for acute cerebral infarction between October 1998 and October 2000 were divided into two groups: those whose infarction extended to the cerebral cortex in the carotid artery region (cortical group) and those with an infarction in the subcortical carotid artery region (subcortical group). MAIN OUTCOME MEASURES: Using a solid-phase enzyme immunoassay, we measured serum levels of neuron-specific enolase on admission and on days 2, 3, and 15. Infarct volume was measured by computed tomography on day 5. The Activities of Daily Living scale was used to assess the clinical outcome at 1-, 3 , and 6-month follow-up after onset. RESULTS: Mean (standard deviation) serum neuron-specific enolase levels were significantly higher among patients with acute cerebral infarction than among controls (18.48 [16.61] ng/mL versus 9.00 [2.70] ng/mL; P<0.001). The neuron-specific enolase level was also higher in the cortical group than in the subcortical group (33.54 [29.71] ng/mL versus 15.97 [5.91] ng/mL; P<0.01). Levels peaked after 2.11 (0.86) days and correlated positively with the infarct volume (r=0.81; P<0.01) and negatively with clinical outcome at 1 month (r= -0.37; P<0.05), 3 months (r= -0.45; P<0.01), and 6 months (r= -0.65; P<0.001), as assessed on the Activities of Daily Living scale. CONCLUSION: Serum neuron-specific enolase levels after cerebral infarction may be a useful marker to predict infarct volume and short- or long-term functional outcome. PMID- 15299171 TI - Clinical features of hereditary spinocerebellar ataxia diagnosed by molecular genetic analysis. AB - OBJECTIVE: To assess the frequency and clinical features of different types of hereditary spinocerebellar ataxia in Hong Kong. DESIGN: Cross-sectional study using a questionnaire and clinical examination, with the majority of the information retrospectively collected. SETTING: Three regional hospitals, Hong Kong. PARTICIPANTS: All patients with spinocerebellar ataxia that was confirmed by molecular genetic tests between January 2001 and October 2003. MAIN OUTCOME MEASURES: History, latest physical examination results, clinical investigation results, and genetic profiles. RESULTS: A total of 16 Chinese patients had received diagnoses of spinocerebellar ataxia. These patients had spinocerebellar ataxia type 1 (n=3), spinocerebellar ataxia type 3 (Machado-Joseph disease; n=12), and dentatorubro-pallidoluysian atrophy (n=1). The most common manifestation was ataxia (15/16), followed by pyramidal signs (12/16). Other features such as bulbar dysfunction, ophthalmoplegia, neuropathy, and cognitive impairment were present but variable. CONCLUSIONS: The clinical manifestations of different types of spinocerebellar ataxia overlap, and genetic study is necessary to confirm the diagnosis. The frequency of spinocerebellar ataxia type 3 is greater than that of other types among these Chinese patients. The age of onset of this type may correlate inversely with the number of CAG repeats. PMID- 15299172 TI - Gout: a review of its aetiology and treatment. AB - OBJECTIVE: To review the current understanding of the causes and the management of gout. DATA SOURCES: Publications on all peer-review literature from MEDLINE from 1965 to January 2004. STUDY SELECTION: Selected and evaluated by the author. DATA EXTRACTION: Extracted and evaluated by the author. DATA SYNTHESIS: The underlying metabolic disorder in gout is hyperuricaemia. Most patients with hyperuricaemia remain asymptomatic throughout their lifetime. The phase of asymptomatic hyperuricaemia ends with the first attack of gouty arthritis or urolithiasis. The risk of gout and stone formation is increased with the degree and duration of hyperuricaemia. Drugs available for the treatment of acute gouty arthritis, such as non-steroidal anti-inflammatory drugs, selective cyclo oxygenase 2 inhibitors, systemic corticosteroids, or colchicine, are effective. For periods between attacks, prophylactic therapy, such as low-dose colchicine, is effective. In those with recurrent attacks of more than two to three times yearly, a uric acid-lowering agent as a long-term therapy should be considered to avoid recurrence and the development of tophaceous gout. CONCLUSIONS: Effective management of gout can be achieved through better understanding of the causes of the condition, preventive measures as well as drug treatment. PMID- 15299173 TI - Early magnetic resonance imaging of radiographically occult osteoporotic fractures of the femoral neck. AB - Osteoporosis is associated with thinning of cortical and trabecular bone, which reduces bone strength and predisposes individuals to fracture development. Femoral neck fractures in patients with osteoporosis may not be apparent on radiographs. Magnetic resonance imaging is useful at detecting these radiographically occult fractures; yet, the practice has not been widely adopted in Hong Kong. In this article, we review our experience of early magnetic resonance imaging in this clinical context--that is, imaging performed within 48 hours of presentation to hospital. Twenty-eight patients (age range, 69-93 years) over a 3-year period were studied. Magnetic resonance imaging revealed radiographically occult neck fractures in 14 (50%) cases (equivalent to 4% of all femoral neck fractures). These fractures were treated surgically (64%) or conservatively (36%) with good bone healing and clinical outcome. When no femoral neck fracture was present, magnetic resonance imaging revealed an alternative cause for symptoms in all 14 cases. We strongly endorse the use of early magnetic resonance imaging for patients with osteoporosis who have a clinically suspected femoral neck fracture that is not visible radiographically. PMID- 15299174 TI - A patient with an increased troponin level without evidence of ischaemic cardiac injury. AB - A 74-year-old man was admitted for chest infection with acute exacerbation of chronic obstructive pulmonary disease. He was incidentally found to have an increased serum level of cardiac troponin I, despite the absence of symptoms and electrocardiographic evidence of ischaemic heart disease. Troponin I became undetectable after the serum was treated with polyethylene glycol, which removed any interfering antibodies. Serum cardiac troponin T was also undetectable after this treatment. Interference of the cardiac troponin I assay by heterophilic antibodies was thus confirmed. Because of the possibility of false-positive results due to immunoassay interference, clinicians should be alerted whenever laboratory findings are incompatible with the clinical picture, and should be ready to perform additional laboratory tests. PMID- 15299175 TI - Combined atrial septostomy and oral sildenafil for severe right ventricular failure due to primary pulmonary hypertension. AB - Management of primary pulmonary hypertension is usually difficult because the disease is uncommon and the aetiology of the disease is not well understood. The disease is potentially lethal because it can lead to failure of the right ventricle, low cardiac output, and ensuing multiple organ failure. We report the successful treatment of a case of low-output syndrome due to primary pulmonary hypertension using combined drug therapy and atrial septostomy. Latest developments in the treatment of this disease are also discussed. PMID- 15299176 TI - A case of probable codeine poisoning in a young infant after the use of a proprietary cough and cold medicine. AB - We report a case of probable poisoning with codeine phosphate in a 3-month-old infant, which was associated with excessive dosing and concomitant use of antihistamines. Investigation into the patient's drug history identified the recent use of a proprietary cough and cold medicine containing codeine phosphate and dexchlorpheniramine. The prescribing information, available from a popular prescribing handbook, listed only one dosage for children, without any adjustment for age or size, and did not bear any warning for its use in young children. A review of the handbook identified seven additional remedies that were similarly listed. Medical practitioners and pharmacists should be aware of this prescribing pitfall. Improvements are needed in the prescribing information pertaining to the use of cough and cold formulas containing opioid or opioid-like antitussives among young children, and clear warnings should be included in drug inserts and formularies. PMID- 15299177 TI - A calcified lesion on abdominal X-ray. PMID- 15299178 TI - The evolution of complaint management in the Hong Kong Hospital Authority. Part 1: Complaints management--a tool for system change? PMID- 15299179 TI - Hospitalised care for patients with transient ischaemic attack. PMID- 15299181 TI - Concomitant radiochemotherapy vs radiotherapy alone in patients with head and neck cancer: a Hellenic Cooperative Oncology Group Phase III Study. AB - The primary objective of the present randomized phase III trial was to compare the 3-yr survival rate of patients treated with standard fractionated radiotherapy (RT) alone or with the same RT concomitantly with cisplatin (DDP) or carboplatin (Cb). From January 1995 until July 1999, 124 patients with histologically proven locally advanced non-nasopharyngeal head and neck cancer (HNC) were randomized to receive either RT monotherapy (70 Gy, Group A) or the same RT concomitantly with DDP (100 mg/m2 on d 2, 22, 42, Group B) or Cb (7 AUC on d 2, 22, 42, Group C). There were no significant differences in complete response rates between patients treated with RT alone or combined chemoradiotherapy. However, median time to progression (TTP) and overall survival (OS) were significantly longer in patients treated with concomitant chemoradiotherapy. Thus, median TTP was 6.3, 45.2, and 17.7 mo in groups A, B, and C respectively (p = 0.0002). Similarly, median OS was 12.2, 48.6, and 24.5 mo, respectively (p = 0.0003). At 3 yr follow-up, 17.5% of patients in group A were alive compared to 52% in group B and 42% in group C (p < 0.001). Patients treated with concomitant chemoradiotherapy experienced more frequently severe hematological toxicity. Also, severe nausea/vomiting was more pronounced in group B, as expected. The present study clearly demonstrated that concomitant chemoradiotherapy with platinum analogs significantly prolongs 3-yr survival and median OS in patients with locally advanced HNC compared to conventional RT alone. PMID- 15299182 TI - Lung cancer in patients with HIV Infection and review of the literature. AB - BACKGROUND: The improved survival of patients since the use of highly active antiretroviral treatments has lead to the reporting of non-AIDS defining tumors, such as lung cancer. METHODS: Analysis of the records of 22 HIV-infected patients with lung cancer (LC) diagnosed in three hospitals located in the Paris area (France). RESULTS: Twenty-one patients were smokers. The patients (86% male, 14% female) had a median age of 45 yr (range, 33-64 yr). Risk factors for HIV infection were intravenous drug use in 5 patients, homosexual transmission in 10 patients, and heterosexual transmission in 7 patients. At diagnosis of LC, seven patients had previously developed a CDC-defined AIDS manifestation, the median CD4 cell count was 364/mm3 (range 20-854/mm3) and median HIV1 RNA viral load was 3000 copies/mL. The most frequent histological subtype was squamous cell carcinoma (11 cases). A stage III-IV disease was observed in 75% of the patients. Only one patient had a small-cell lung carcinoma. Twenty-one patients received combined specific therapy, of which six patients underwent surgery for the LC. The median overall survival was 7 mo. No opportunistic infections occurred during LC therapy. CONCLUSIONS: LC occurs at a young age in HIV-infected smokers. LC is not associated with severe immunodeficiency. The prognosis is poor because of their initial extensive disease and a poor response to therapy. However, surgery appears to improve outcome in much the same way as in the general population. PMID- 15299183 TI - Effects of tamoxifen on bone mineral density and metabolism in postmenopausal women with early-stage breast cancer. AB - At the present time, tamoxifen is the most widely used anti-estrogen for adjuvant therapy and metastatic disease in postmenopausal women with breast cancer, a population at high risk for osteoporosis. This prospective study was designed to evaluate the effect of adjuvant tamoxifen on bone mineral density and all biochemical markers concomitantly in women with early-stage breast cancer in one study. Using dual-energy X-ray absorptiometry, prior to and 12 mo after tamoxifen treatment, bone mineral density in lumbar spine and femoral neck was measured in 44 women with T1-T2N0M0 estrogen-receptor-positive breast cancer receiving adjuvant treatment with tamoxifen 20 mg/d. Biomarkers that can affect bone mineral metabolism were measured before and after 3 and 12 mo of tamoxifen treatment. Bone mineral density was minimally increased in lumbar spine and femoral neck after 12 mo treatment with tamoxifen (p = 0.79 and 0.55, respectively). No differences were found in serum levels of calcium, phosphate, creatinine, ALAT, albumin, LDH, calcitonin, or estradiol. A significant decrease in osteocalcin levels was found after 3 and 12 mo (p < or = 0.01). TSH and PTH levels were increased (p < or = 0.05) after 3 mo, returning to baseline after 12 mo. In conclusion, tamoxifen has an estrogen-like effect on bone metabolism in postmenopausal women and is associated with preservation of bone mineral density in lumbar spine and femoral neck. Changes in serum concentration of biochemical markers may reflect decreased bone turnover or bone remodeling and add to the understanding of tamoxifen's effect on bone mineral density. PMID- 15299184 TI - The nuclear oxysterol receptor LXRalpha is expressed in the normal human breast and in breast cancer. AB - The liver X> or = receptor alpha (LXRalpha) is a nuclear receptor with a key role in bile acid biosynthesis and cholesterol metabolism. The present study investigated the expression and function of LXRalpha in the normal and malignant human breast. LXRalpha mRNA transcripts were detected by RT-PCR in nine breast carcinoma cell lines. The nucleotide sequence of the cloned PCR product was identical to the corresponding human LXRalpha cDNA sequence. Expression of LXRalpha protein was confirmed by immunoblot analysis of breast cancer cell lysates. LXRalpha mRNA was expressed in 14/15 (93%) of normal human breast mammoplasty specimens and in 11/15 (73%) of primary breast carcinomas. Oxysterol and nonsteroidal LXRalpha agonists at low micromolar concentrations inhibited proliferation of breast carcinoma cell lines in culture. The importance of LXRalpha signaling in cholesterol homeostasis and the observed expression of LXRalpha in normal breast tissue suggest that this nuclear oxysterol receptor has an important physiological function in the breast. LXRalpha gene expression is regulated by dietary fatty acids implicated in breast carcinogenesis and detection of LXRalpha expression in breast cancer cell lines and breast tumors in the present study indicates that LXRalpha may also be important in breast carcinogenesis. Inhibition of breast cancer cell proliferation suggests that pharmacological LXRalpha agonists may have potential preventive and/or therapeutic antitumor activity in breast cancer. PMID- 15299186 TI - Should mammographic screening be done in primary ovarian cancer?: A case control study in Turkish women. AB - Breast cancer is a significant global health problem. It is the most common malignancy in women. Mammographic screening is recommended for women older than 40 yr for early detection of breast cancer. The aim of this study is to evaluate the role of screening mammography in ovarian cancer independent of age. Eighty four patients with ovarian cancer were evaluated with bilateral mammography. Two hundred asymptomatic healthy controls with a similar age distribution were also imaged with screening mammography. Mammography results were classified according to the American College of Radiology criteria in five groups. The median age of the study group was 51.4 (range, 27-77) and 49.3 (range, 30-75) in the control group. Screening mammography detected four cases of malignancy (4.8%) in patients with ovarian cancer; two were the primary breast carcinomas(2.5%) and two were metastatic cancers from the ovary. Five subjects (2.5%) among healthy controls were also found to have breast cancer. Although the incidence of primary breast carcinoma was found to be similar in the two groups (2.5%), mammographic imaging detected metastatic disease to the breast from the ovaries. Mammography should therefore be considered in patients with ovarian cancer independent of age. PMID- 15299185 TI - Phase I trial of bi-weekly paclitaxel and gemcitabine as second-line therapy for patients with non-small-cell lung cancer previously treated with platinum-based chemotherapy. AB - A phase I study was conducted to evaluate the maximum tolerated dose, feasibility, and efficacy of bi-weekly-administered paclitaxel and gemcitabine in patients with non-small-cell lung cancer (NSCLC) who had previously been treated with platinum-based chemotherapy. In a dose-escalation study, 18 patients, under 75 yr old, with unresectable NSCLC that had relapsed or was resistant after platinum-containing chemotherapy with performance status of 0-2 were enrolled. Patients were treated with paclitaxel and gemcitabine bi-weekly. The dose escalation levels of paclitaxel (mg/m2) at a fixed dose of gemcitabine 1000 mg/m2 were 110 (level 1), 130 (level 2), 150 (level 3), and 170 (level 4), respectively. All patients were eligible for evaluation of toxicities. At level 4, one patient developed an infection with grade 3 neutropenia and two other patients developed severe neurotoxicity (over grade 3). Thus, the recommended dose for phase II was paclitaxel 150 mg/m2 and gemcitabine 1000 mg/m2 due to dose limiting toxicities including neutropenia and peripheral neurotoxicity. Partial response was seen in 4 cases of the 18 assessable patients, with an overall response of 22%. Bi-weekly paclitaxel and gemcitabine is feasible and appears to be an efficacious combination chemotherapy as second-line chemotherapy in refractory and recurrent patients with NSCLC who had been previously exposed to platinum-containing chemotherapy. PMID- 15299187 TI - Expression and initial promoter characterization of PCAN1 in retinal tissue and prostate cell lines. AB - Prostate cancer is the most frequently diagnosed neoplasia in men and one of the leading causes of cancer-related deaths in men over 60. In an effort to understand the molecular events leading to prostate cancer, we have identified PCAN1 (prostate cancer gene 1) (also known as GDEP), a gene that is highly expressed in prostate epithelial tissue and frequently mutated in prostate tumors. Here we demonstrate its expression in neural retina, and retinoblastoma cell culture but not retinal pigment epithelial cell culture. We further characterize PCAN1 expression in the prostate cell lines RWPE1, RWPE2, and LnCAP FGC. We demonstrate an increase in expression when the cells are grown in the presence of Matrigel, an artificial extracellular basement membrane. Expression was time dependent, with expression observed on d 6 and little or no expression on d 12. Testosterone was not found to increase PCAN1 expression in this culture system. In addition, normal prostate epithelial cells co-cultured with normal prostate stromal cells did not exhibit PCAN1 expression at any time. To definitively locate the transcription initiation sites, we performed restriction ligase-mediated 5' RACE, to selectively amplify only mRNA with a 5' cap. An initial characterization of the sequence upstream of the initiation sites determined six possible binding sites for the prostate specific regulatory protein NKX3.1 and four potential binding sites for the PPAR/RXR heterodimer that is involved in the control of cell differentiation and apoptosis. PMID- 15299188 TI - Allogeneic tumor-dendritic cell fusion vaccines for generation of broad prostate cancer T-cell responses. AB - Antigen-loaded dendritic cells (DC) are considered potent stimulators of protective immunity. In some studies, DC hybridized with tumor cells have shown promising antitumor responses in mice as well as in humans. A practical drawback of this approach is the limited availability of autologous tumor samples. We investigated the possibility of hybridizing allogeneic prostate cancer cells with human-monocyte-derived DC, and thereby combine the wide repertoire of antigens from the tumor cells with the costimulatory features of the autologous antigen presenting cells. Three tumor cell lines were used for hybridization using polyethylene glycol (PEG). We show that all three cell lines formed hybrids with DC to the same extent and without significant loss of viability. Restimulation of autologous T cells with these hybrids resulted in generation of tumor-specific IFNgamma-producing T cells with all three tumor cell lines. Similar tumor specificity could not be obtained if T cells were stimulated using a mixture of non-PEG-fused tumor cells and DC. Moreover, these T cells were capable of specific recognition of other tumor cells of prostate cancer origin and autologous DC pulsed with lysate from these prostate cancer cell lines, while a panel of unrelated EBV-transformed B cell lines were not recognized. These results are strongly indicative of the true tumor specificity of these T cells. Our results suggest that DC hybridized with allogeneic prostate cancer cell lines are potent stimulators of a broad prostate-tumor-specific response. PMID- 15299189 TI - Vincristine, cisplatin, teniposide, and cyclophosphamide combination in the treatment of recurrent or metastatic adrenocortical cancer. AB - The efficacy and tolerability of a combination of vincristine, cisplatin, teniposide, and cyclophosphamide (OPEC) in 11 patients (median age, 45 yr) with recurrent and/or metastatic adrenocortical cancer (ACC) (seven functional and four nonfunctional) were evaluated. All patients received this regimen after the failure of streptozocin and o,p'-DDD (SO) combination therapy. The regimen comprised cyclophosphamide, 600 mg/m2, and vincristine, 1.5 mg/m2, maximum dose 2.0 mg (d 1); cisplatin, 100 mg/m2 (d 2) and teniposide, 150 mg/m2 (d 4). Cycles were repeated every 4 wk. One to eight cycles (median, six cycles) of OPEC were administered to each patient. The median duration of treatment was 6 mo. The overall 2-yr survival rate was 82% and the median survival since diagnosis was 44 mo while it was 21 mo since start of OPEC therapy. Responses were obtained in nine patients: partial response in two patients, and stable disease in seven patients. The median duration of response was 6.75 mo. A total of 60 cycles of chemotherapy were given to all patients; grade 1-2 toxicity occurred in 57 cycles, while grade 3 toxicity was observed only in two cycles, according to NCI's Common Toxicity Criteria. We conclude that the OPEC regimen may be considered in recurrent or metastatic ACC as a second-line medical treatment. However, the combination is accompanied by considerable side effects and dose modifications are necessary in order to be able to recommend the treatment. This regimen needs further evaluation compared with SO therapy preferably in a randomized multicenter trial. PMID- 15299190 TI - The comparison of spontaneous LDH release activity from cultured PBMC with sera LDH activity in non-Hodgkin's lymphoma patients. AB - Based on the fact that lactate dehydrogenase (LDH) enzyme is a very sensitive indicator of the cellular metabolic state, aerobic or anaerobic direction of glycolysis, activation status, and malignant transformation, in this study we compared values of the spontaneous LDH release from circulating PBMC with sera LDH activity in 53 different subtypes of non-Hodgkin's lymphoma (NHL) patients. Results shows that serum LDH was significantly (p < 0.05) elevated in comparison to the range values only in the advance clinical stage (III and IV) in all investigated subtypes of NHL according to The Working and REAL classification. On the other hand, the spontaneous LDH release from cultures PBMC is significantly (p < 0.01) elevated in early and advanced stage in all investigated forms of NHL in comparison to healthy controls. Based on consideration that an increase in spontaneous LDH release appears before elevated sera LDH activity, we conclude that determination of spontaneous LDH release by microassay from cultured cells together with other findings may help in the diagnosis of NHL patients, especially in patients with early stage of disease. PMID- 15299191 TI - Enzyme activities controlling adenosine levels in normal and neoplastic tissues. AB - Adenosine is known to be associated with effects such as inhibition of immune response, coronary vasodilation, stimulation of angiogenesis, and inhibition of inflammatory reactions. Some authors suggest that adenosine may also have similar functions in tumor tissues. Tissue levels of adenosine are under close regulation by different enzymes acting at different levels. Adenosine is produced from AMP by the action of 5'-nucleotidase (5'-NT) and is converted back into AMP by adenosine kinase (AK) or into inosine by adenosine deaminase (ADA). Inosine is converted into purine catabolites by purine nucleoside phosphorylase (PNP), whereas AMP is converted into ADP and ATP by adenylate kinase (MK). The aim of this study was to analyze the activities of the above enzymes in fragments of neoplastic and apparently normal mucosa, obtained less than 5 cm and at least 10 cm from tumors, in 40 patients with colorectal cancer. The results showed much higher activities of ADA, AK, 5'-NT, and PNP in tumor tissue than in neighboring mucosa (p > 0.01 for ADA, AK, and PNP; p > 0.05 for 5'-NT), suggesting that the activities of purine metabolizing enzymes increase to cope with accelerated purine metabolism in cancerous tissue. The simultaneous increase in ADA and 5'-NT activities might be a physiological attempt by cancer cells to provide more substrate to accelerate salvage pathway activity. PMID- 15299192 TI - Erythropoietin against cisplatin-induced peripheral neurotoxicity in rats. AB - Cisplatin (CDDP) is a potent anticancer drug, and neurotoxicity is one of its most important dose-limiting toxicities. In this study we investigated the role of recombinant human erythropoietin (rhuEPO) for protection against CDDP-induced neurotoxicity. All experiments were conducted on female Wistar-albino rats. Animals were randomly assigned to three groups. Group A received only CDDP, group B received CDDP plus rhuEPO, and group C received only rhuEPO. Electroneurography (ENG) was done in the beginning and at the end of 7 wk, then the rats were sacrificed and the sciatic nerve was removed for histopathological examination. The mean initial latency was 2.7438 ms in group A, 2.4875 ms in group B, and 2.62 ms in group C. After 7 wk of treatment, the latency was 2.4938, 2.6313, and 2.3900 ms, respectively. The difference in latencies was not statistically significant. The amplitude of compound muscle action potential (CMAP) was 12.8125 mV, 14.3875 mV, and 14.5600 mV before the treatment and 8.4875, 12.8250, and, 13.0800 mV after treatment, respectively. Amplitude of CMAP was significantly greater in rhuEPO-treated groups (groups B and C) compared to cisplatin only Group A. The mean area of CMAP was 12.2625, 12.3500, and, 12.2800 mV s before the treatment and 5.7125, 10.6463, and 9.1600 mV s after the treatment, respectively. The area of CMAP was significantly larger in rhuEPO-treated groups. In histopathological studies thick, thin, and total number of nerve fibers were 4053, 5050, and 9103, in group A, 5100, 8231, and 13331, in group B, and 5264, 6010, and 11274, in group C respectively. In the microscopic examination active myelinization process was observed in rhuEPO-treated groups. We concluded that at the given dose and schedule CDDP-induced motor neuropathy and rhuEPO prevented this neuropathy by sparing the number of normal nerve fibers and by protecting the amplitude and area of CMAP. We concluded that rhuEPO may also play a role in active myelinization and it is an active agent in protection against CDDP-induced peripheral neuropathy, warranting further clinical studies. PMID- 15299193 TI - Rituximab-induced tumor progression: does it really happen? AB - Monoclonal antibodies have been gaining a wide role in the treatment of malignant diseases. A human chimeric anti-CD20 monoclonal antibody (Mab) rituximab (Rituxan in USA; Mabthera in Europe) was approved for the treatment of refractory or relapsed low-grade or follicular non-Hodgkin's lymphoma (NHL) in 1997 (1-3). Rituximab has efficacy in other refractory CD20+ NHLs, hairy cell leukemia, plasma cell dyscrasias, posttransplant lymphoproliferative syndrome, autoimmune phenomena such as refractory hemo lytic anemias, and immune thrombocytopenias (4 8). Its combination with standard chemotherapy protocols for NHL has been investigated thoroughly owing to its synergistic effect when combined with chemotherapeutic agents (3). Coiffier et al. recently published a randomized trial showing a statistically significant survival benefit of rituximab-CHOP combination over CHOP alone in elderly patients with diffuse large-B-cell lymphoma (9,10). In addition to these widening beneficial therapeutic effects, rituximab has well-known side effects, encountered especially during its first infusion, such as chills, fever, allergic reactions, cardiopulmonary syndrome, and tumor lysis syndrome. We would like to share our clinical observation in a patient with NHL, whose disease seemed to go into an accelerated progression phase after rituximab administration. PMID- 15299197 TI - Expression of cyclooxygenase-2 and thromboxane synthase in non-neoplastic and neoplastic thyroid lesions. AB - Cyclooxygenase-2 (COX-2) and thromboxane synthase (TBXAS) are important enzymes involved in the arachidonic acid pathway and synthesis of prostaglandins. We examined COX-2 and TBXAS immunoreactivity in 150 surgically resected thyroid specimens using immunohistochemistry to determine expression in benign and malignant thyroid lesions and to examine their roles in thyroid tumor progression. Papillary thyroid carcinomas and follicular carcinomas expressed higher levels of COX-2 compared to follicular adenomas and adenomatous nodules. We showed for the first time that TBXAS was expressed in thyroid tissues, with higher levels in papillary carcinomas compared to non-neoplastic and benign thyroid tissues. Western blot was performed on seven thyroid samples. These results indicate that both COX-2 and TBXAS are expressed in benign and malignant thyroid tissues. Although some malignant thyroid tumors showed higher levels of COX-2 expression, COX-2 and TBXAS are probably not useful in the immunohistochemical diagnosis of thyroid malignancies. However, the expression of both COX-2 and TBXAS by thyroid tissue may provide insight into the role of these enzymes in progression from benign to malignant thyroid tumors. PMID- 15299196 TI - Immunomodulatory functions of the diffuse neuroendocrine system: implications for bronchopulmonary dysplasia. AB - Pulmonary neuroendocrine (NE) cells are believed to be the precursor of NE lung carcinomas, including well-differentiated (carcinoids) and moderately/poorly differentiated (atypical carcinoids and small-cell carcinomas, SCLCs) subtypes. In early studies, we determined mechanisms by which NE cell-derived peptides such as bombesin-like peptide (BLP) promote normal fetal lung development. Postnatally, BLP may normally regulate perinatal adaptation of the pulmonary circulation. However, elevated BLP levels in premature infants shortly after birth predict which infants are at high risk for developing bronchopulmonary dysplasia (BPD, chronic lung disease of newborns). An anti-BLP blocking antibody abrogates clinical and pathological evidence of lung injury in two baboon models of BPD. These observations indicate that BLP mediates lung injury in BPD, supporting a role for BLP as pro-inflammatory cytokines. We have directly tested the effects of BLP on eliciting inflammatory cell infiltrates in vivo. Surprisingly, mast cells are the major responding cell population. These data suggest that the diffuse NE system may be a newly recognized component of innate immunity in multiple organ systems. We speculate that overproduction of NE cell derived peptides such as BLP may be responsible for a variety of chronic inflammatory disorders. PMID- 15299198 TI - Expression of adhesion molecules and cytokeratin 20 in merkel cell carcinomas. AB - Merkel cell carcinoma (MCC) is an aggressive neuroendocrine carcinoma of the skin. MCCs often show characteristic paranuclear dot-like immunopositivity for cytokeratin 20 (CK20), a globular aggregation of CK20 intermediate filaments. These aggregates typically form rhabdoid features and fibrous bodies and may be associated with a down-regulation in adhesion molecules (AMs). To date, the relationship between the expression of AMs and CK20 and clinicopathological findings in MCC has not been well examined. In this immunohistochemical study, we assessed the expression of AMs, CK20, and chromogranin A (CgA) on MCCs in 8 men and 23 women with this disease, and also characterized their clinicopathological features. This study is the largest of its kind that has been undertaken to date in Japanese patients. Compared to normal tissue, E-cadherin and alpha- and beta catenins showed reduced membranous expression in 95.7%, 46.7%, and 45.2% of MCCs, respectively. Nuclear E-cadherin localization was seen in four tumors, all of which predominantly showed a CK20 dot pattern. However, there was no significant relationship between the membranous expression of AMs and a CK20 dot pattern. E cadherin expression was significantly lower in tumors of > or =2 cm, and tumors negative for E-cadherin more frequently developed outside of the head and neck than within those regions. CgA was more intensely expressed in tumors with uniform nuclei and a dense lymphocytic infiltrate than in those that showed pleomorphisms and that had few, if any, infiltrating lymphocytes. These findings suggest that MCCs have a reduced expression of AMs and that down-regulation of E cadherin expression may correlate with increased tumor aggressiveness. The fact that no significant relationship was demonstrable between the membranous expression of AMs and the CK20 expression pattern suggests that the mechanism of aggregation of intermediate filaments may be different in different types of tumors. PMID- 15299199 TI - Correlation between tumor vascularity and clinical findings in patients with pituitary adenomas. AB - Angiogenesis generally plays an essential role in tumor growth and metastasis, and also influences the response to treatment in human malignant solid tumors. Even in nonmalignant tumors, angiogenesis is essential for tumor growth and invasion. In order to define the relationship between tumor vascularity and the clinical course in patients with pituitary adenomas, we quantified the vascularity in 47 pituitary adenomas and in 6 normal anterior pituitary glands obtained at autopsy using a computed image-analyzing system. We estimated two parameters, the vascular number and the area as the vascularity. Additionally, we calculated mean individual vessel size using the above two parameters. The relationships of tumor vascularity to clinical, endocrinological and histological findings was assessed. Factors considered included patient age and gender, preoperative medication, histological type, concentration of each hypersecreted pituitary hormone, maximum tumor size, cavernous sinus invasion, intratumoral hemorrhage, and immunohistological results of localization of vascular endothelial growth factor (VEGF). Vascularity was significantly higher in normal glands than in pituitary adenomas. However, there were no significant correlations between tumor vascularity and other clinical, endocrinological, or histological parameters, suggesting that increased angiogenesis is not essential for pituitary adenoma growth or invasiveness. PMID- 15299200 TI - Pancreatic endocrine pathology in von Hippel-Lindau disease: an expanding spectrum of lesions. AB - A 41-yr-old patient with a history of von Hippel-Lindau (VHL) disease with previously removed bilateral pheochromocytomas and renal cell carcinoma presented with progressive obstructive jaundice due to multiple lesions in the pancreas. The pancreatectomy specimen showed a range of endocrine lesions including islet hyperplasia, nesidioblastosis, microadenomas, and endocrine carcinoma. In addition, some of the non-tumorous islets displayed peliosis. The endocrine carcinoma showed a biphasic pattern composed of typical endocrine cells and oncocytes. The oncocytic component showed widespread lymphovascular invasion and lymph node metastasis. Immunohistochemistry and electron microscopy confirmed that the oncocytic cells were endocrine. Focal areas contained cells with foamy cytoplasm, a feature that is associated with pancreatic endocrine tumors in VHL. This case expands the spectrum of lesions seen in the pancreas of VHL patients. There is some overlap with lesions encountered in multiple endocrine neoplasia type I. In addition, the endocrine lesions were composed of two main cell types (typical and oncocytic cells) with the oncocytic component invading lymphatic channels and spreading to regional lymph nodes. PMID- 15299202 TI - Pituitary pathology in Erdheim-Chester disease. AB - Pituitary morphologic changes in patients with Erdheim-Chester disease have not been described in detail. We report here the histologic and immunohistochemical findings in the autopsy obtained pituitary of a 35-yr-old woman with extensively disseminated Erdheim-Chester disease. The posterior lobe was completely replaced by xanthogranulomatous infiltrates, providing an explanation for the patient's diabetes insipidus. The anterior lobe was intact and immunohistochemistry demonstrated expression of GH, TSH, FSH, LH, and alpha subunit within the normal range. A clinically observed decrease of anterior pituitary function was interpreted as hypothalamic in origin due to massive destruction of the hypophysial stalk and compression of the hypothalamus. Prolactin immunoreactive cells were numerous, consistent with the view that prolactin cell hyperplasia resulted from the loss of hypothalamic dopaminergic inhibition. Massive Crooke's hyalinization in the ACTH-producing cells was considered unrelated to Erdheim Chester disease and was the consequence of treatment with pharmacologic doses of glucocorticoid hormones. It can be concluded that prolactin cell hyperplasia may be the only finding in the adenohypophysis of patients with disseminated Erdheim Chester disease. It appears that in our patient the clinically apparent anterior hypopituitarism was not due to the lack of storage but rather to insufficient release of adenohypophysial hormones caused by the defect in hypothalamic regulation. PMID- 15299204 TI - Endogenous biotin staining as an artifact of antigen retrieval with automated immunostaining. PMID- 15299201 TI - Combined sellar fibrosarcoma and prolactinoma with neuronal metaplasia: report of a case unassociated with radiotherapy. AB - We report the occurrence of a primary pituitary fibrosarcoma causally unrelated to radiotherapy, admixed in association with a prolactin cell pituitary adenoma showing neuronal metaplasia. These unique findings were associated with multiple endocrine neoplasia type 1 (MEN 1). Primary fibrosarcoma involving the sella is a very rare tumor. The majority of cases have been associated with prior irradiation of either a pituitary adenoma or a craniopharyngioma. Pituitary adenoma with neuronal metaplasia is also rare and usually occurs in the setting of acromegaly. Despite the intimate association of both elements in our lesion, no transition of adenoma to sarcoma was demonstrable by immunohistochemistry or in situ hybridization studies. PMID- 15299203 TI - Malignant peripheral nerve sheath tumor of the thyroid: a clinicopathological and ultrastructural study of one case. AB - Thyroid malignant peripheral nerve sheath tumors (TMPNST) are very uncommon neoplasms that can be confused with anaplastic carcinoma, Riedel's thyroiditis, or other soft tissue tumors that may occur in the thyroid region. An example of TMPNST is presented in this report. The tumor occurred in a 56-yr-old woman. Fine needle aspiration did not provide adequate material. After thyroidectomy, the lesion posed important problems in differential diagnosis. Immunohistochemical, molecular, and electron microscopic features were taken into consideration to arrive at the correct diagnosis. Tumor cells were focally positive for keratins, a feature that has not been described in peripheral nerve sheath tumors of the thyroid, but that has been occasionally seen in tumors from other locations. After thyroidectomy, the patient received radiotherapy. She is well without evidence of recurrence 10 mo after surgery. PMID- 15299205 TI - Proliferative activity in pregnancy-related thyroid papillary carcinoma. PMID- 15299207 TI - Generation and use of transgenic mice as models of osteoarthritis. AB - The term osteoarthritis (OA) represents a group of diseases characterized by gradual degradation of articular cartilage and a number of associated degenerative processes within the joint. Consequently, no single animal model is likely to fulfil all the criteria of OA. The present chapter discusses the possibilities of using transgenic technologies for modification of the mouse genome to generate animal models of OA. After discussing the different approaches available, we provide an example of the generation of a traditional transgenic mouse strain and describe techniques and practical aspects of genotyping as well as the preparation of skeletal samples for radiological, histological, immunohistological, and molecular biologic analyses for phenotype characterization. We also present a histological grading system to evaluate the progression of OA lesions, with examples of other degenerative alterations in the knee joint structures. PMID- 15299208 TI - Development and clinical application in arthritis of a new immunoassay for serum type IIA procollagen NH2 propeptide. AB - Type II collagen, the most abundant protein of cartilage matrix, is synthesized as a procollagen molecule including the N-(PIINP) and C-(PIICP) propeptides at each end. Type II procollagen is produced in two forms as the result of alternative RNA splicing. One form (IIA) includes and the other form (IIB) excludes a 69-amino acid cysteine-rich globular domain encoded by exon 2 in PIINP. During the process of synthesis, these N-propeptides are removed by specific proteases and released in the circulation, and their levels are believed to reflect type II collagen synthesis. In this chapter we describe the development of a specific enzyme-linked immunosorbent assay (ELISA) for the measurement of the IIA form of PIINP (PIIANP) in serum based on a polyclonal antibody raised against recombinant human exon 2 fusion protein of type II procollagen. We show that this ELISA is highly specific for circulating PIIANP and has adequate technical precision. In patients with knee osteoarthritis and rheumatoid arthritis, serum PIIANP was decreased by 53% (p < 0.0001) and 35% (p < 0.001), respectively, suggesting that type IIA collagen synthesis is altered in these arthritic diseases. The measurement of serum PIIANP may be useful for the clinical investigation of patients with joint diseases. PMID- 15299209 TI - Histological and immunohistological studies on cartilage. AB - The material properties of dense avascular connective tissues such as cartilage present unique challenges to the development of any prospective histological procedures. This chapter documents some of the protocols we have developed in our laboratories for the histological and immunohistochemical evaluation of extracellular-matrix components in normal and pathological articular cartilage. Illustrative examples are provided of specimens from an ovine meniscectomy model of osteoarthritis. A major strength of this experimental animal model is that spatial and temporal changes in joint matrix components can be followed longitudinally in an animal population of well-defined pedigree that minimizes biological variation. Furthermore, a systematic experimental approach can be adopted with this model using cartilage specimens from early predegenerative stages to later advanced stages of joint degeneration. Significantly, the histological changes observed in this model closely parallel changes evident in joint tissues during osteoarthritis in humans and have provided significant insights into these degenerative processes. PMID- 15299210 TI - Histochemical visualization of the cartilage hyaladherins using a biotinylated hyaluronan oligosaccharide bioaffinity probe. AB - Hyaluronan (HA) binding proteins (HABPs) were localized in cartilaginous ovine tissues (articular cartilage, intervertebral disc) using a biotinylated HA (bHA) oligosaccharide bioaffinity probe. The bHA oligosaccharide probe was prepared by partial digestion of HA with ovine testicular hyaluronidase, and the oligosaccharides were labeled with biotin hydrazide and purified by a combination of aggrecan G1 domain and avidin affinity chromatography. Hyaladherins were prominently visualized in tissue sections using the bHA oligosaccharide probe as pericellular components in hypertrophic epiphyseal and vertebral growth plate chondrocytes and in the enlarged cells of the cartilaginous end plate of the intervertebral disc. Weaker extracellular staining was also evident in the matrix of the ovine newborn hip and knee joint cartilages. The bHA oligosaccharide probe also visualized intracellular HABPs (IHABPs) in the hypertrophic growth plate chondrocytes of the primary ossification centers. Monolayer cultures of ovine chondrocytes rapidly internalized the bHA oligosaccharide affinity probe to discrete cytoplasmic, nuclear, and perinuclear regions, which were visualized by indirect fluorescent microscopy. This bHA oligosaccharide affinity probe may be useful in future investigations designed to characterize these novel cartilage IHABPs and the role that HA endocytosis plays in cellular regulatory processes in cartilage homeostasis. PMID- 15299211 TI - Methods for cartilage and subchondral bone histomorphometry. AB - This chapter presents the histological assessment of cartilage and bone of tibial plateaus, by procedures that have been applied and validated in two animal models of osteoarthritis: meniscectomized rats and guinea pigs. It starts from bone sampling, followed by all the steps of sample preparation from embedding to sectioning (without prior decalcification), staining, and mounting. Depending on the cartilage or bone components to be visualized, two dyes are described: safranin O and Goldner's trichrome. On these stained sections, various histomorphometric parameters are then quantified using the dedicated programs of an image analyzer. The following parameters are evaluated at the medial side of the tibia and are described at the levels of both cartilage (cartilage thickness, fibrillation index, proteoglycan content ratio based on safranin-O staining intensities and chondrocyte density) and bone (subchondral bone plate thickness). PMID- 15299212 TI - Laser-mediated microdissection as a tool for molecular analysis in arthritis. AB - Most current approaches to the analysis of gene expression in arthritic tissue samples are based on RNA isolated either from cultured synovial cells or from synovial biopsies. However, this strategy does not distinguish between specific gene expression profiles of cells originating from separate tissue areas. Therefore, we established the combination of laser-mediated microdissection and RNA arbitrarily primed polymerase chain reaction (RAP-PCR) for differential display to analyze profiles of gene expression in histologically defined areas of arthritic tissue. Cryosections derived from synovial tissue were used to obtain cell samples from different tissue areas using a microbeam laser microscope. RNA was isolated and analyzed using nested RAP-PCR to generate a fingerprint of the expressed gene sequences. Differentially expressed bands were isolated, cloned, and sequenced. Differential expression of identified sequences was confirmed by in situ hybridization and immunohistochemistry. PMID- 15299213 TI - Analysis of protein distribution in cartilage using immunofluorescence and laser confocal scanning microscopy. AB - Protein localization in cartilage sections by antibodies that specifically bind to epitopes of a protein is one of the most powerful technologies in modern cartilage research. Studies using two or more primary antibodies that recognize different protein epitopes allow the colocalization of different gene products in one cartilage section. In addition, specific histochemical stains help to visualize nuclear DNA, mitochondria, and other subcellular compartments. By these immunohistological methods, the distribution of proteins can be analyzed throughout different zones of articular cartilage. In particular, with the use of laser scanning confocal microscopy, subcellular localization of proteins can also be determined (i.e, nuclear, cytoplasmic, membrane-associated, and extracellular). Overall, immunohistochemical methods are fairly simple to handle, and the reagents required are inexpensive, with the exception of basic technical equipment (fluorescence microscope or confocal microscope). However, as with many methodologies, technical knowledge and experience is important to avoid and/or interpret either false-positive or false-negative results. PMID- 15299214 TI - Molecular and biochemical assays of cartilage components. AB - The procedure described below is useful for extracting proteins, nucleic acids, and glycosaminoglycans from 5-40 mg of cartilage or tissue-engineered cartilage samples. This extraction method will generate samples compatible with Western blot, RNase protection, dimethyl methylene blue (DMB) assay for glycosaminoglycan, Hoechst DNA assay, and hydroxyproline assay. Most soluble matrix molecules can be extracted from pulverized samples using 4 M guanidine HCl, during a 30-min period of vortex agitation at 4 degrees C. Shorter agitation times can give inadequate solubilization. The guanidine HCl-insoluble pellet must be re-extracted with guanidine thiocyanate buffer, to solubilize RNA additionally. The final insoluble pellet can be rinsed with ethanol and digested with papain, to quantify collagen content as well as other insoluble or crosslinked material. Samples between 1 and 5 mg may be directly digested with a small volume of papain buffer for DMB, hydroxyproline, and Hoechst DNA assays. PMID- 15299215 TI - Mechanical characterization of native and tissue-engineered cartilage. AB - Cartilage functions as a low-friction, wear-resistant, load-bearing tissue. During a normal gait cycle, one cartilage surface rolls and slides against another, all the while being loaded and unloaded. The durability of the tissue also makes for an impressive material to study. However, when cartilage is damaged or diseased, the tissue has little capacity to repair itself. The goal of cell-based repair strategies to replace damaged or diseased tissue requires that the functional biomechanical properties of normal (developing or mature), diseased, and repair cartilage be restored. This chapter addresses some of the major methods used to assess the biomechanical properties of native and tissue engineered cartilage. First, the traditional methods of testing by compression, tension, shear, and indentation are reviewed. Next, additional methods to evaluate interfacial mechanics and lubrication are described. Thus, a variety of mechanical tests may be used to assess functional properties for normal, diseased, and tissue-engineered cartilage. PMID- 15299216 TI - Noninvasive study of human cartilage structure by MRI. AB - Magnetic resonance (MR) imaging is a 3D imaging technique that has recently begun to permit direct delineation of cartilage structure. This chapter summarizes current methodology for the morphological (e.g., volume, thickness) and compositional imaging of cartilage using quantitative MR as well as for semiquantitative scoring of cartilage disease. The chapter explains the relevance of MR in identifying disease status, in monitoring disease progression, and in identifying risk factors of osteoarthritis, as well as in evaluating treatment response to so-called structure-modifying osteoarthritis drugs. The practical methodological procedures presented involve the description of how to acquire MR images for structural analysis and the semiquantitative scoring of articular cartilage. We also present a description of image-analysis techniques for cartilage segmentation and characterization of cartilage structure (quantitative outcome parameters, such as volume and thickness) and guidelines for how to test the validity (accuracy), precision (reproducibility), and sensitivity to change of such methodologies inosteoarthritis. PMID- 15299217 TI - High-resolution MRI techniques for studies in small-animal models. AB - High-field magnetic resonance (MR), which allows high-resolution imaging, is a powerful research tool to examine and visualize hyaline cartilage of small joints noninvasively. Recent studies have shown that qualitative assessment of degenerative joint disease, derived from MR images, was reliable. The ability to show pathologic changes throughout the time-course of the disease from 3D datasets has also been demonstrated. Quantitative imaging for accurate determination of cartilage volume and thickness are still to be confirmed. This chapter presents a MR protocol for in vivo imaging of small-animal knee joints on a 7-T imager. Technical aspects and specifics of modality and field strength are discussed, as well as future developments and expected evolutions of techniques. PMID- 15299218 TI - High-resolution imaging of osteoarthritis using microcomputed tomography. AB - Three-dimensional imaging of osteoarthritis is so far limited to late stages of the disease. In this chapter we introduce microcomputed tomography (microCT) as a new imaging tool that offers exciting features for diagnosis of earlier disease stages and for disease monitoring. microCT provides spatial resolution better than 100 microm, but the size of the objects that can be scanned is restricted to several centimeters. The strength of X-ray-based techniques like micro CT is the excellent visualization of bone. Therefore, the main application of microCT in osteoarthritis (OA) will be the analysis of bone in small-animal models or of human bone biopsies. As an example, we will exemplarily describe the application of microCT for the examination of knee joints of male STR1N mice. This inbred strain spontaneously develops OA that carries many characteristics of the human disease. With microCT it is possible to monitor the prominent bony alterations such as osteophyte formation, trabecular remodeling, subchondral bone plate thickening, and subchondral sclerosis. We discuss sample preparation, scanning procedures, data processing, and analysis as well as implications and restrictions for in vivo and in vitro applications. PMID- 15299219 TI - High-resolution ultrasonography for analysis of age- and disease-related cartilage changes. AB - Because of their limited spatial resolution, current clinical noninvasive imaging modalities (radiography, computed tomography, conventional echography, and magnetic resonance imaging) are able to detect only the late stages of the cartilage degradation. To detect early lesions and follow their evolution in time with imaging, higher resolution is necessary. Recent work suggest that high frequency ultrasound may serve as a useful means for the investigation of cartilage matrix structural changes occurring under various experimental and clinical circumstances, like the growing process and osteoarthritis. In this chapter, an experimental 50-100-MHz ultrasound scanner is described for high resolution echographic imaging of articular cartilage. The procedures of data acquisition and signal processing are detailed for the quantitative evaluation of ultrasonic reflection and backscatter coefficients, which have been reported to be sensitive to subtle surface and internal disease-related alterations. Further technological developments and miniaturization of the echographic probes may lead to extension of this technique to the study of living small animals or to the clinical field in combination with conventional arthoscopy. PMID- 15299220 TI - Evaluation of cartilage composition and degradation by high-resolution magic angle spinning nuclear magnetic resonance. AB - Rheumatic diseases are accompanied by a progressive destruction of the cartilage layers of the joints. Although the number of patients suffering from rheumatic diseases is steadily increasing, degradation mechanisms of cartilage are not yet understood, and methods for early diagnosis are not available. Although some information on pathogenesis could be obtained from the nuclear magnetic resonance (NMR) spectra of degradation products in the supernatants of cartilage specimens incubated with degradation-causing agents, the most direct information on degradation processes would come from the native cartilage as such. To obtain highly resolved NMR spectra of cartilage, application of the recently developed high-resolution magic-angle spinning (HR-MAS) NMR technique is advisable to obtain small line-widths of individual cartilage resonances. This technique is nowadays commercially available for most NMR spectrometers and has the considerable advantage that the same pulse sequences as in high-resolution NMR can be applied. Except for a MAS spinning equipment, no solid-state NMR hardware is required. Therefore, this method can be easily implemented. Here, we describe the most important requirements that are necessary to record HR-MAS NMR spectra. The capabilities of the HR-MAS technique are discussed for the 1H and 13C NMR spectra of cartilage. PMID- 15299221 TI - Pulsed-field gradient-nuclear magnetic resonance (PFG NMR) to measure the diffusion of ions and polymers in cartilage: applications in joint diseases. AB - Since cartilage contains neither blood nor lymph vessels, diffusion is the most important transport process for the supply of cartilage with nutrients and for the removal of metabolic waste products. Therefore, diffusion measurements are of high interest in cartilage research. Different techniques of diffusion measurements exist. Here we describe methods based on pulsed-field gradient nuclear magnetic resonance (PFG NMR). This technique offers the considerable advantage that neither concentration gradients nor labeling of the diffusing species are required. In addition to the description of the fundamentals and the applicability of PFG NMR studies in cartilage research, emphasis is on the influence of the observation time, Delta, on the diffusion coefficient, D: at short times, diffusion is primarily determined by the water content of the sample, and great care is needed to keep this parameter constant. However, by varying the diffusion time, data on the internal structure of cartilage, e.g., the distance of the collagen fibrils, can also be obtained. In addition to classical water diffusion, the diffusion behavior of selected ions and polymers in cartilage is described. The capabilities, the limitations, and the clinical relevance of diffusion measurements for the assessment of joint diseases are discussed. PMID- 15299222 TI - Dynamics of collagen in articular cartilage studied by solid-state NMR methods. AB - Methods for studying the fast molecular dynamics of the rigid macromolecules in cartilage are described. The strong dipolar couplings and chemical shift anisotropies of these molecules necessitate application of solid-state nuclear magnetic resonance (NMR) techniques such as magic-angle spinning, cross polarization, and high-power dipolar decoupling to obtain resolved NMR spectra. The molecules in cartilage that are amenable to these techniques are collagen and the rigid portion of the glycosaminoglycans (mostly hyaluronan). Site-specific mobility information is obtained from scaled dipolar couplings measured in 2D NMR experiments. Motionally averaged dipolar couplings can be interpreted in terms of order parameters that provide information about the amplitudes of molecular motions. Qualitative dynamics information is obtained from the simple wideline separation experiment measuring 1H-1H widelines representing the strength of the 1H-1H dipolar coupling. Quantitative values for molecular order parameters are obtained from precise measurements of 1H-13C dipolar couplings along the C-H bond vector. Two experimental techniques, the Lee-Goldburg cross-polarization and the dipolar coupling/chemical shift experiment, are illustrated to measure these 1H 13C dipolar couplings. Unlike glycosaminoglycans in cartilage, the collagen moiety remains substantially ordered, undergoing fast small amplitude motions. As enzymes cleave the macromolecules in articular cartilage in the course of arthritis, solid-state NMR techniques are capable of characterizing the increased motions of their degradation products in diseased cartilage. PMID- 15299223 TI - Computerized protocol for anatomical and functional studies of joints. AB - This chapter describes a new methodology for the acquisition and computer elaboration of joint anatomy and motion data and the study of their correlation. The method uses a commercial industrial electrogoniometer, custom tools, and software designed and developed by the authors for interactive display of the anatomical structures during joint motion, numerical interpolations of the articular geometries, and kinematic analysis of motion. The original data acquisition protocol and computer elaboration software are described in detail, and a final subheading describes briefly previous studies and future developments. PMID- 15299224 TI - The vicissitudes of global eradication of polio. PMID- 15299225 TI - Current dietary trends in the management of diabetes. PMID- 15299226 TI - Medicinal plants in India. PMID- 15299227 TI - Thrombophilia in coronary artery disease: a double jeopardy. AB - Thrombophilia can be defined as an increased risk of thrombosis. The central event to the pathogenesis of any thrombotic episode is the perturbation of haemostasis, the cause of which may be genetic or environmental. The clinical manifestations of the chronic development of coronary artery atheroma are angina and acute myocardial infarction. In recent years literature is emerging on the role of different factors of blood coagulation in arterial thrombosis. Different coagulation factors, natural anticoagulants, platelet antigens and other factors such as homocysteine, lipoprotein (a), have been studied as risk factors for coronary artery disease (CAD). The results of many of these studies are contradictory. In India, there is an alarming rise in the number of young patients with myocardial infarction (MI) and an interesting feature is that a large majority of these patients lack the conventional risk factors. There have been scattered studies on the thrombophilia status among Indians. The management of thrombophilia can be done by a regimen of different drugs which has been evaluated in different clinical trials. Since the cost of thrombophilia investigations is quite phenomenal for a developing country like India, the selection of these investigations assumes an utmost importance. PMID- 15299228 TI - Antiviral activity of medicinal plants of Nilgiris. AB - BACKGROUND & OBJECTIVES: Medicinal plants have been traditionally used for different kinds of ailments including infectious diseases. There is an increasing need for substances with antiviral activity since the treatment of viral infections with the available antiviral drugs often leads to the problem of viral resistance. Herpes simplex virus (HSV) causes a variety of life threatening diseases. Since the chemotherapeutic agents available for HSV infections are either low in quality or limited in efficiency, there is a need to search for new and more effective antiviral agents for HSV infections. Therefore in the present study 18 plants with ethnomedical background from different families were screened for antiviral activity against HSV-1. METHODS: Different parts of the plants collected from in and around Ootacamund, Tamil Nadu were extracted with different solvents to obtain crude extracts. These extracts were screened for their cytotoxicity against Vero cell line by assay microculture tetrazolium (MTT) trypan blue dye exclusion, proteins estimation and 3H labeling. Antiviral properties of the plant extracts were determined by cytopathic effect inhibition assay and virus yield reduction assay. RESULTS: Three plant extracts Hypericum mysorense, Hypericum hookerianum and Usnea complanta exhibited significant antiviral activity at a concentration non toxic to the cell line used. The extracts of Melia dubia, Cryptostegia grandiflora and essential oil of Rosmarinus officinalis showed partial activity at higher concentrations. INTERPRETATION & CONCLUSION: Some of the medicinal plants have shown antiviral activity. Further research is needed to elucidate the active constituents of these plants which may be useful in the development of new and effective antiviral agents. PMID- 15299229 TI - Hepatoprotective effect of Epaltes divaricata extract on carbon tetrachloride induced hepatotoxicity in mice. AB - BACKGROUND & OBJECTIVES: Epaltes divaricata is widely used in Sri Lanka as an Ayurvedic medicine. In the present study the hepatoprotective and antioxidative effects of an aqueous extract of E. divaricata plant (Family-Compositae) were investigated against carbon tetrachloride induced hepatocellular injury in mice. METHODS: Healthy male mice (30-35 g body weight, 6-8 wk old) were used. A single dose of carbon tetrachloride (CCl4, 0.5 ml/kg in olive oil) was administered ip to induce hepatotoxicity and the plant extract at a dose of 0.9 g/kg was administered orally by gavage. Animals were sacrificed 24 h and 4 days after the administration of CCl4. Blood and liver tissue were collected for the assessment of serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP) and liver reduced glutathione level. The liver tissue was used for histopathological assessment of liver damage. RESULTS: Pre treatment of mice with the plant extract of Epaltes (0.9 g/kg) orally for 7 days significantly reduced serum levels of ALT (P<0.01), AST (P<0.01) and ALP (P<0.001) enzymes by 21.40, 47.36 and 71.12 per cent respectively and significantly increased (P<0.001) the liver reduced glutathione level by 42.32 per cent, 24 h after the administration of carbon tetrachloride. A marked improvement in the enzyme activities and the liver reduced glutathione level was observed in the Epaltes pre-treated mice 4 days after the administration of carbon tetrachloride. Histopathological studies provided supportive evidence for the biochemical analysis. INTERPRETATION & CONCLUSION: The results of the present study indicated that under the present experimental conditions, aqueous extract of Epaltes divaricata showed hepatoprotective abilities against carbon tetrachloride induced liver damage in mice. PMID- 15299230 TI - Presence of sopE gene & its phenotypic expression among different serovars of Salmonella isolated from man & animals. AB - BACKGROUND & OBJECTIVES: Salmonellae cause a spectrum of diseases in man and animals but their virulence factors responsible for induction of gastroenteritis and/or systematic infection are still poorly understood. Also, the different subspecies and serovars of Salmonella differ considerably in their virulence for man and animals. There is increasing evidence that Salmonella possesses a dedicated protein secretion system denoted type III secretion system (TTSS) that is involved in the early stage of Salmonella infection. One such TTSS is Salmonella outer protein E (SopE) that helps in the invasion of Salmonella by stimulating membrane ruffling. In the present study the presence of sopE gene and its phenotypic expression (SopE protein) among different serovars of Salmonella enterica isolated from man and animals in India was investigated. METHODS: A total of 50 isolates of S. enterica belonging to 11 serovars were tested for the presence of sopE gene by polymerase chain reaction. The in vitro phenotypic expression of SopE protein was detected by Western blotting using anti-SopE serum. RESULTS: Of the 50 isolates of S. enterica belonging to 11 serovars tested for the presence of sopE,14 belonging to three serovars viz., Enteritidis, Gallinarum and Virchow were found to carry the sopE gene. Similarly, 13 isolates belonging to same three serovars were found to express SopE protein phenotypically as detected by Western blotting using anti-SopE serum. INTERPRETATION & CONCLUSION: The results indicated that sopE gene appeared to be distributed and conserved among only a few serovars of Salmonella (Enteritidis, Gallinarum and Virchow) irrespective of their source of isolation. The presence of sopE gene in Salmonella provides an important pathogenic means to invade epithelial cells. PMID- 15299231 TI - Antimicrobial resistance in selected bacterial enteropathogens in north India. AB - BACKGROUND & OBJECTIVES: The resistance of enteropathogenic bacteria to commonly prescribed antibiotics is increasing both in developing as well as in developed countries. Resistance has emerged even to newer, more potent antimicrobial agents. The present study was therefore undertaken to report the current antibiotic resistance in common bacterial enteropathogens isolated in a tertiary care hospital in north India. METHODS: Faecal samples from 1802 patients were cultured for common bacterial enteropathogens and identified by standard methods. Antibiotic susceptibility was done by Stoke's disk diffusion method. The clinical and demographic profile of the patients was noted. RESULTS: Stool specimens from 119 (88 male, 31 female) patients yielded Shigella, Salmonella, Vibrio cholerae or Aeromonas. Fifty two per cent (62/119) of patients were children and 70 per cent were below the age of 5 yr. Twenty seven patients developed hospital acquired diarrhoea. Among all enteropathogens, Shigella spp. was the commonest followed by non-typhoidal Salmonella (27), V. cholerae O1 El tor serotype Ogawa (19), Aeromonas spp. (14), Salmonella Typhi and S. paratyphi A (2 isolates each). Resistance to antimicrobial agents was common among all pathogens. Among shigellae an overall resistance of 63.6, 58.1 and 16.3 per cent was observed for nalidixic acid, cotrimoxazole and furazolidone respectively. Seven isolates of Shigella were resistant to ciprofloxacin, (18.5%) of non-typhoidal salmonellae were resistant to ciprofloxacin. V. cholerae were generally susceptible to tetracycline (only 1 isolate out of 13 resistant) and other drugs except nalidixic acid (89.5% resistance) and cotrimoxazole (77.8% resistance). INTERPRETATION & CONCLUSION: Enteropathogens have developed high level resistance to first line agents used for empiric treatment of diarrhoea. Progressively increasing resistance to ciprofloxacin is a serious cause of concern. PMID- 15299232 TI - Ratio of middle cerebral to umbilical artery blood velocity in preeclamptic & hypertensive women in the prediction of poor perinatal outcome. AB - BACKGROUND & OBJECTIVES: Doppler velocimetry studies of placental and foetal circulation can provide important information regarding foetal well-being providing an opportunity to improve foetal outcome. The present study was undertaken to evaluate the role of middle cerebral to umbilical artery blood velocity waveform's systolic/diastolic ratio (MCA/UA) and biophysical profile as a predictor of perinatal outcome in hypertensive and preeclamptic pregnant women during the late third trimester. METHODS: Fifty preeclamptic pregnant women selected randomly in the last three weeks of the third trimester were stratified into two groups based on the MCA/UA ratio. All women were evaluated by foetal biophysical profile scoring. Thirty four women with foetal MCA/UA ratios > 1 and 16 with < or =1 were recruited in groups A and B respectively. The results of the ratio, and biophysical profile were evaluated with respect to the outcome of the infants and adverse perinatal outcome, defined as perinatal death, foetal cord blood gas analyses, cesarean delivery for foetal distress, admission to the neonatal intensive care unit, days in the neonatal intensive care unit (NICU) or low Apgar score. RESULTS: Rate of cesarean delivery was significantly (P<0.001) higher in group B than group A. There was a statistically significant increase in perinatal morbidity in B group. Apgar scores at 1 and 5 min were found to be lower in group B than group A. Umbilical cord blood partial oxygen pressure (pO2), partial carbon dioxide pressure (pCO2) was not different in the two groups; whereas, pH was lower in group B. In group A two neonates (5.9%) and in group B 12 neonates (75%) required admission in neonatal intensive care unit. Best cut-off levels of MCA, MCA/UA ratios were found to be 3 and 1, respectively. INTERPRETATION & CONCLUSION: The MCA/UA was valuable for predicting the outcome of preeclamptic and hypertensive pregnancies. When the ratio was <1, foetal prognosis was poor. PMID- 15299233 TI - Reliability & validity of the Malayalam Functional Assessment of Cancer Therapy for Head & Neck Cancer. AB - BACKGROUND & OBJECTIVES: The need for quality over quantity in cancer survival is increasingly being recognised in the developing countries, and the efforts to monitor quality of life (QOL) are increasing. However, the non-availability of a valid and reliable tool in the local language is a common problem. Cross culturally sensitive tools enable the researchers to compare different patient populations and identify cultural differences and variations. The present study was carried out to translate, validate and test for reliability a reliable QOL tool for the head and neck cancer patient population in a tertiary care hospital in south India. METHODS: The functional assessment of cancer therapy for head and neck cancer (FACT-H&N) was translated into the local language (Malayalam) and tested for reliability in 140 patients of head and neck cancer. RESULTS: The translated tool showed substantial psychometric sensitivity. The Cronbach's alpha for the total FACT-H&N was 0.94. The alpha scores for the five subscales ranged from 0.81-0.92. Significant correlations were observed amongst the total QOL and subscale scores and patient's demographic, disease and treatment variables. INTERPRETATION & CONCLUSION: The Malayalam translation of the FACT-H&N questionnaire was developed, tested and validated. It was found to satisfactorily measure QOL in head and neck cancer patients. PMID- 15299234 TI - Smouldering focus of kala-azar in Assam. PMID- 15299235 TI - A comparison of the torsional performance of stainless steel and titanium alloy tibial intramedullary nails: a clinically relevant approach. AB - In recent years there has been a tendency to design and manufacture intramedullary nails from titanium alloy rather than from stainless steel. The aim of this project was to compare the torsional performance of one manufacturers standard stainless steel and titanium alloy tibial intramedullary nails, using their distal locking screw holes and dedicated cross screws to secure each nail distally. A custom built test rig and materials testing machine were used to determine the torsional rigidity of the nails. Theory was used to calculate the torsional rigidity of the central parts of each nail. From the mechanical testing, the mean torsional rigidity of the titanium alloy nail system was 40.9 N m2 while that of the stainless steel nail system was 34.6 N m2, for all distal interlocking screw positions tested. Based on theoretical calculations the torsional rigidity of the central part of the nail was 83 N m2 for the stainless steel nail and 66 N m2 for the titanium alloy nail. This study shows the importance of using the distal locking screw holes and dedicated cross screws to secure intramedullary nails during mechanical testing so that clinically relevant results are obtained about the whole nail system and not just the nail. PMID- 15299236 TI - The proposal of noninvasive quantitative diagnostic method of the atherosclerosis and the clarification of organ correlation of atherosclerosis and oxygen metabolism. AB - The proposal of noninvasive diagnostic method of mechanical degradation of vascular wall is clinically useful and it will be correlated with noninvasive diagnostic method of atherosclerosis. Supersonic Doppler effect sensor has been used to measure blood flow velocity as a noninvasive measuring method. However, it is remain problem whether the output from the Doppler effect sensor really detects the pure blood flow velocity. Theoretically, when the Doppler effect sensor is set perpendicular to the blood flow direction, that is, perpendicular to the blood vessel, the output will correspond to the expansion velocity of blood vessel wall, because it detect the frequency of Doppler shifted supersonic scattered from vascular wall. Previously, on the basis of this concept, using Doppler effect sensor, we showed this method can really detect the deformation velocity of blood vessel wall and it correlates the degradation of elastic property of blood vessel. Furthermore, using this proposed measuring method, atherosclerosis is found to progress correspondingly with the visco-elastic degradation of vascular wall. In this paper, on the basis of our proposed method, the quantitative noninvasive estimation method of the degradation of vascular wall and the progressive degree of atherosclerosis by unique parameter has been proposed. Using this method, the degradation of vascular wall is correlated to the oxygen metabolic function of blood vessel corresponding to the function of oxygen transportation and progression of atherosclerosis. Furthermore, the organ correlation on the atherosclerosis between lower limb and carotid is investigated by this proposed method. PMID- 15299237 TI - Study of hardness and roughness modification in explanted joint components. AB - The significant wear of the UHMWPE bearings of explanted knee prostheses is produced mainly by micrometric debris ("third-body" wear) that diffuse toward the mobile metal-polymer interface. Here debris is crushed during the movement producing scratches in the metal and in the polymeric surfaces. Mechanical stress and the biological effects change the physical polymeric properties. In order to evaluate the area of UHMWE bearings submitted to high load stresses, in this work physical investigations are performed on the explanted knee prosthesis. Particularly, the roughness profile analysis (RPA) and the micro-hardness measurements (MHM) resulted suitable for the localisation of the mechanical and biological wear area. In the stressed zone, surface treatments could be applied in order to improve the mechanical resistance of the polymeric material. Particularly, the ion implantation with heavy ions is proposed to enhance the polymeric wear resistance. The Scanning Electron Microscopy (SEM), Energy Dispersive X-ray Spectrometry (EDX) and Infrared absorption spectroscopy (FTIR) techniques were also applied and results discussed. PMID- 15299238 TI - Wear properties of alumina/zirconia composite ceramics for joint prostheses measured with an end-face apparatus. AB - While only alumina is applied to all-ceramic joint prostheses at present, a stronger ceramic is required to prevent fracture and chipping due to impingement and stress concentration. Zirconia could be a potential substitute for alumina because it has high strength and fracture toughness. However, the wear of zirconia/zirconia combination is too high for clinical use. Although some investigations on composite ceramics revealed that mixing of different ceramics was able to improve the mechanical properties of ceramics, there are few reports about wear properties of composite ceramics for joint prosthesis. Since acetabular cup and femoral head of artificial hip joint are finished precisely, they indicate high geometric conformity. Therefore, wear test under flat contact was carried out with an end-face wear testing apparatus for four kinds of ceramics: alumina monolith, zirconia monolith, alumina-based composite ceramic, and zirconia based composite ceramic. Mean contact pressure was 10 MPa and sliding velocity was 40 mm/s. The wear test continued for 72 hours and total sliding distance was 10 km. After the test, the wear factor was calculated. Worn surfaces were observed with a scanning electron micrograph (SEM). The results of this wear test show that the wear factors of the both composite ceramics are similarly low and their mechanical properties are much better than those of the alumina monolith and the zirconia monolith. According to these results, it is predicted that joint prostheses of the composite ceramics are safer against break down and have longer lifetime compared with alumina/alumina joint prostheses. PMID- 15299239 TI - Active finite element analysis of skeletal muscle-tendon complex during isometric, shortening and lengthening contraction. AB - An active finite element model was developed to predict the mechanical behaviors of skeletal muscle-tendon complex during isometric, shortening and lengthening contraction. The active finite element was created through incorporation of a user-defined material property into ABAQUS finite element code. The active finite element is controlled by a motor element that is activated by a mathematical function. The nonlinear passive behavior of the muscle was defined by the viscoelastic elements and can be easily altered to other properties by using other elements in the material library without the need of re-defining the constitutive relation of the muscle. The isometric force-length relationship, force-strain relations of the muscle-tendon complex during both shortening and lengthening contraction and muscle relaxation response were predicted using the proposed finite element model. The predicted results were found to be in good agreement with available experimental data. In addition, the stress distribution in the muscle-tendon complex during isometric, shortening and lengthening contractions was simulated. The location of the maximum stress may provide useful information for studying muscle damage and fatigue in the future. PMID- 15299240 TI - Patterned layers of adsorbed extracellular matrix proteins: influence on mammalian cell adhesion. AB - Three patterned systems aiming at the control of mammalian cell behavior are presented. The determinant feature common to these systems is the spatial distribution of extracellular matrix (ECM) proteins (mainly collagen) on polymer substrates. This distribution differs from one system to another with respect to the scale at which it is affected, from the supracellular to the supramolecular scale, and with respect to the way it is produced. In the first system, the surface of polystyrene was oxidized selectively to form micrometer-scale patterns, using photolithography. Adsorption of ECM proteins in presence of a competitor was enhanced on the oxidized domains, allowing selective cell adhesion to be achieved. In the second system, electron beam lithography was used to engrave grooves (depth and width approximately 1 microm) on a poly(methyl methacrylate) (PMMA) substratum. No modification of the surface chemistry associated to the created topography could be detected. Cell orientation along the grooves was only observed when collagen was preadsorbed on the substratum. In the third system, collagen adsorbed on PMMA was dried in conditions ensuring the formation of a nanometer-scale pattern. Cell adhesion was enhanced on such patterned collagen layers compared to smooth collagen layers. PMID- 15299241 TI - Time-controlled pulse-drug release from dry-coated wax matrix tablets for colon drug delivery. AB - This study aimed to prepare a colon drug delivery system using dry-coated time controlled disintegration wax matrix tablets. Indomethacin was used as a model drug. Behenic acid and lactose were used as coating materials. The effects of lactose content and pH of the dissolution medium on drug release were investigated. The porosity and the tortuosity of the surface matrix layer were calculated. Four formulations of wax matrices containing different percentages of lactose in the surface layer, i.e. 70, 65, 60 and 55, were prepared. The lag times of indomethacin release from the matrices in 0.05 M phosphate buffer pH 7.4 were 50, 162, 294 and 539 minutes for formulations containing 70, 65, 60 and 55% lactose, respectively. The release of drug from formulations containing lactose in the range of 60-70% in different media, i.e. 0.05 M phosphate buffer pH 7.4, 0.05 M alkaline borate buffer pH 8.5 and in the case of pH changed media from pH 1.3 to pH 7.4, was not different (p=0.1). This implies that the different environment in the gastro-intestinal tract will not affect the release of this delivery device. The required lag time period can be met by varying the amount of lactose. PMID- 15299242 TI - The dependence between the strength and stiffness of cancellous and cortical bone tissue for tension and compression: extension of a unifying principle. AB - A strong positive correlation between the apparent ultimate strength and stiffness of bone tissue that can be expressed by a unified relationship has been observed for cortical bone in tension and low-density cancellous bone in compression. For practical purposes, the existence of a relationship between strength and stiffness is significant in that bone stiffness can be measured in vivo using non-invasive methods. It is generally accepted that bone strength is greater in compression than in tension whereas there is no substantial evidence that bone stiffness in compression is different from that in tension. This might suggest that compressive strength would relate to the stiffness, if at all, in a way that is different from tensile strength. In order to examine similarities and differences in the way strength is associated with stiffness between modes of loading and tissue type, we tested equine cortical bone and bovine cancellous bone in compression and examined these data together with previously reported data from compression testing of human cancellous bone as well as tensile testing of cortical bone from various sources. We have found for cortical bone that (i) the sensitivity of strength to stiffness is the same for tension and compression (p>0.75, ANCOVA), and (ii) the difference between the magnitudes of compressive and tensile strength for cortical bone is the result of an additive, rather than a multiplicative factor (52.1 MPa after adjusting to 1 microstrain/s, p<0.0001, ANOVA). High-density bovine tibial cancellous bone, on the other hand, has a steeper slope for its compressive strength-stiffness relationship than that for cortical bone and human cancellous bone, resulting in a transitional relationship between compressive strength and stiffness for a range of bone types and densities. Based on the current results and previous work, it is suggested that the offset strength in the compressive strength-stiffness relationship may be a direct manifestation of the difference between the compressive and tensile strengths of the bone material that constitutes the building blocks of the bone structure. Deviation of high-density cancellous bone compressive behavior from the other bone types and densities is attributed to stress distribution differences between the bone types. PMID- 15299243 TI - An assessment of stress analyses in the theory of abfraction. AB - Wedged-shaped lesions at the cemento-enamel junction of teeth have been attributed primarily to biomechanical loading forces that cause flexure and failure of enamel and dentin. This theory, termed abfraction, remains controversial. This review examined studies on mechanical properties of enamel and dentin and studies on bite forces and mastication as background information. Abfraction is based principally on a few early finite element analysis and photoelastic models showing stress concentration at the dental cervical area without actually showing enamel and dentin fracture. However, a review of more recent dental stress analyses has been contradictory. Particularly, analyses of the periodontal ligament and alveolar bone, not modeled in previous studies, have shown that those structures may dissipate occlusal loading forces from the cervical areas. In addition, some models may not fully represent intricate dental anatomy and complex occlusal function. Therefore, the key basis of the abfraction theory may be flawed. PMID- 15299244 TI - Electrochemical studies of the corrosion behaviour of titanium and the Ti-6Al-4V alloy using electrochemical impedance spectroscopy. AB - OBJECTIVES: The aim of this study was to compare the electrochemical behaviour of two materials used in oral implantology. METHODS: The resistance to corrosion of Ti grade 2 and the alloy Ti-6Al-4V was studied in an artificial saliva solution. It has been observed that the passivation of titanium by an oxidised layer can be shown both by cyclic voltametry and by electrochemical impedance spectroscopy. Moreover, this latter technique, rarely used in odontology, opens up interesting perspectives, enabling a more quantitative approach to the resistance of the passive layer to be adopted. RESULTS: Also, the impedance data recorded for Ti grade 2 and the alloy Ti-6Al-4V, in the artificial saliva solution were shown that Ti grade 2 has a higher resistance to corrosion and a thicker oxide layer than the alloy Ti-6Al-4V. SIGNIFICANCE: The fact that the electrochemical properties of Ti-6Al-4V are lower than that titanium's ones indicate than a release of ions aluminum and or vanadium ions in the body can occur. This is why we recommend to the dental practitioners to preferably use titanium in implantology. PMID- 15299245 TI - Tumor control probability for tumor therapy with beta radionuclide interstitial injection. PMID- 15299246 TI - Dosimetry of beta nuclide filled balloon catheter in endovascular brachytherapy. PMID- 15299248 TI - Adaptation concept, tissue remodeling, mechanobiology and tissue engineering: a survey. PMID- 15299249 TI - Biomechanical properties of human articular cartilage under compressive loads. AB - The function of articular cartilage is to support and distribute loads and to provide lubrication in the diarthrodial joints. Cartilage function is described by proper mechanical and rheological properties, strain and depth-dependent, which are not completely assessed. Unconfined and confined compression are commonly used to evaluate the Young's modulus (E) and the aggregate modulus (H(A)), respectively. The Poisson's ratio (nu) can be calculated indirectly from the equilibrium compression data, or using the biphasic indentation technique; it has recently been optically evaluated by using video microscopy during unconfined compression. The transient response of articular cartilage during confined compression depends on its permeability k; a constant value of k can be easily identified by a simple analytical model of confined compression tests, whereas more complex models or direct measurements (permeation tests) are needed to study the permeability dependence on deformation. A poroelastic finite element model of articular cartilage was developed for this purpose. The elastic parameters (E,nu) of the model were evaluated performing unconfined compression creep tests on human articular cartilage disks, whereas k was identified from the confined test response. Our combined experimental and computational method can be used to identify the parameters that define the permeability dependence on deformation, as a function of depth from articular surface. PMID- 15299250 TI - Mechano-acoustic determination of Young's modulus of articular cartilage. AB - The compressive stiffness of an elastic material is traditionally characterized by its Young's modulus. Young's modulus of articular cartilage can be directly measured using unconfined compression geometry by assuming the cartilage to be homogeneous and isotropic. In isotropic materials, Young's modulus can also be determined acoustically by the measurement of sound speed and density of the material. In the present study, acoustic and mechanical techniques, feasible for in vivo measurements, were investigated to quantify the static and dynamic compressive stiffness of bovine articular cartilage in situ. Ultrasound reflection from the cartilage surface, as well as the dynamic modulus were determined with the recently developed ultrasound indentation instrument and compared with the reference mechanical and ultrasound speed measurements in unconfined compression (n=72). In addition, the applicability of manual creep measurements with the ultrasound indentation instrument was evaluated both experimentally and numerically. Our experimental results indicated that the sound speed could predict 47% and 53% of the variation in the Young's modulus and dynamic modulus of cartilage, respectively. The dynamic modulus, as determined manually with the ultrasound indentation instrument, showed significant linear correlations with the reference Young's modulus (r(2)=0.445, p<0.01, n=70) and dynamic modulus (r(2)=0.779, p<0.01, n=70) of the cartilage. Numerical analyses indicated that the creep measurements, conducted manually with the ultrasound indentation instrument, were sensitive to changes in Young's modulus and permeability of the tissue, and were significantly influenced by the tissue thickness. We conclude that acoustic parameters, i.e. ultrasound speed and reflection, are indicative to the intrinsic mechanical properties of the articular cartilage. Ultrasound indentation instrument, when further developed, provides an applicable tool for the in vivo detection of cartilage mechano acoustic properties. These techniques could promote the diagnostics of osteoarthrosis. PMID- 15299251 TI - The role of viscoelasticity of collagen fibers in articular cartilage: theory and numerical formulation. AB - The relative importance of fluid-dependent and fluid-independent transient mechanical behavior in articular cartilage was examined for tensile and unconfined compression testing using a fibril reinforced model. The collagen matrix of articular cartilage was modeled as viscoelastic using a quasi-linear viscoelastic formulation with strain-dependent elastic modulus, while the proteoglycan matrix was considered as linearly elastic. The collagen viscoelastic properties were obtained by fitting experimental data from a tensile test. These properties were used to investigate unconfined compression testing, and the sensitivity of the properties was also explored. It was predicted that the stress relaxation observed in tensile tests was not caused by fluid pressurization at the macroscopic level. A multi-step tensile stress relaxation test could be approximated using a hereditary integral in which the elastic fibrillar modulus was taken to be a linear function of the fibrillar strain. Applying the same formulation to the radial fibers in unconfined compression, stress relaxation could not be simulated if fluid pressurization were absent. Collagen viscoelasticity was found to slightly weaken fluid pressurization in unconfined compression, and this effect was relatively more significant at moderate strain rates. Therefore, collagen viscoelasticity appears to play an import role in articular cartilage in tensile testing, while fluid pressurization dominates the transient mechanical behavior in compression. Collagen viscoelasticity plays a minor role in the mechanical response of cartilage in unconfined compression if significant fluid flow is present. PMID- 15299252 TI - Quantitative electrical impedance analysis of cartilage degradation. PMID- 15299253 TI - Zonal and directional variations in tensile properties of bovine articular cartilage with special reference to strain rate variation. AB - THE AIMS of this study were: (i) to investigate the variation in the tensile properties of articular cartilage with depth through cartilage thickness and fibre orientation; (ii) to determine the effect of strain rate on tensile properties of articular cartilage. MATERIALS AND METHOD: All experimental work was performed on cartilage specimens taken from two bovine knee joints. Osteochondral plugs 12 mm in diameter were harvested with a special reamer from the femur and the tibial plateaux of each knee. Slices (0.2 mm thick), of articular cartilage were cut from the plug with a microtome. The predominant orientation of the collagen fibres on the cartilage surface was determined using the pinpricking technique. Each specimen used for the tensile test was cut, so as to produce a dumbbell shape, with a gauge length of 6 mm. Uniaxial tensile tests were performed on each specimen in order to determine the tensile Young's modulus, and ultimate tensile strength (UTS). In this investigation, these tensile tests were carried out at different strain rate: 1, 20, 50 and 70%/sec. RESULTS: As regards the zonal properties, it was found that tensile stiffness was greater in the superficial layer than in deep layer. However, a few specimens from the deep layer displayed similar or greater stiffness compared to the superficial layer. With respect to the directional properties, the specimens oriented parallel to the predominant alignment of collagen, were stiffer than those, which were perpendicular to it in each layer. However, only the results regarding the deep layer can be considered statistically significant. In regard to the variation of modulus with the strain-rate, the results showed that there is no significant increase of the modulus with increasing strain rate from 20 to 50% per second. However, at 70% per second, articular cartilage stiffness considerably increased by up to one order of magnitude greater than that determined at lower strain rates in both the superficial and deep layer. Moreover, the UTS of cartilage specimens tested at 70% per second showed a significant rise, reaching values of four to five times that of those measured at 1, 20 or 50% per second. CONCLUSION: The steep increases in both the stiffness and ultimate tensile strength of cartilage at high strain rates point to the existence in cartilage of a mechanism for its protection from damage by stresses arising in trauma, which are usually applied at high rates. This mechanism needs to be elucidated. The reduced anisotropy found in the present study pointed out that collagen is likely to be less organized in bovine cartilage than in the human and therefore, a study of its ultra-structure would be appropriate. PMID- 15299254 TI - Macroscale modeling of cartilage: mixture theory versus homogenization. AB - The equations governing the electro-chemo-hydro-mechanical processes in cartilage are derived using the periodic homogenization technique. First the equations at the microscale are recalled. Then the homogenization technique is applied to propagate the physics to the macroscale. The results of this work and of the thermodynamical approach used by various authors are finally compared. PMID- 15299255 TI - Role of cell-associated matrix in the development of free-swelling and dynamically loaded chondrocyte-seeded agarose gels. AB - Chondrocytes embedded in agarose and subjected to dynamic deformational loading produce a functional matrix with time in culture, but there is usually a delay in the development of significant differences compared to free swelling. In this study, we hypothesized that the initial presence of a cell-associated matrix would expedite construct development in response to dynamic deformational loading. Seeded samples with enzymatically isolated chondrocytes and chondrons (the chondrocyte and its pericellular matrix) and examined the effects of seeding density and dynamic loading on the development of tissue properties. At 60 million/ml, dynamic loading significantly augmented the development of material properties in chondrocyte- and chondron-seeded constructs. Biochemical content and histological analysis indicated that the deposition of GAG, link protein and collagens are affected by the pre-existing cell-associated matrix of the chondron seeded samples. The pericellular matrix associated with the chondrons did not expedite the development of material properties in response to deformational loading, disproving our hypothesis. The relative concentration and distribution of matrix proteins may play a major role in the disparate responses observed for the chondrocyte- and chondron-seeded cultures. In further support of these findings, culturing chondrocytes in agarose for two weeks prior to the application of deformational loading also did not exhibit expedited construct development. PMID- 15299256 TI - Flexibility of type I collagen and mechanical property of connective tissue. AB - The hierarchical organization of the connective tissue, more specifically, the musculoskeletal soft tissue, has been extensively studied. With advancements in experimental methodology, investigation of the structure-function relationship has provided more insight into how the mechanical integrity of the tissue is created. Such information is essential in the linking the macroscopic loading environment of the tissue to the microscopic level of the tissue to be experienced by the cell. The flexibility and elastic modulus of gross connective tissue, the fascicle, the fiber and then the collagen molecule are compared based on the data available in the literature. PMID- 15299257 TI - Single molecule mechanical properties of type II collagen and hyaluronan measured by optical tweezers. AB - Type II collagen and hyaluronan are the two major components of extracellular molecules in cartilage and play an important role in mechanical functions of extracellular matrix. Currently, their mechanical properties have been investigated only at the gross-level. In this study, the mechanical properties of single type II collagen and hyaluronan molecules were directly measured using optical tweezers technique. The persistence length was found to be 11.2+/-8.4 nm in type II collagen and 4.5+/-1.2 nm in hyaluronan. This result suggested that type II collagen is stiffer than hyaluronan at the individual molecule level, which supports the general concept that collagen is responsible for resisting tensile force. The experimental system developed here also provides a powerful tool for quantifying mechanical properties of extracellular matrix at the single molecule level. PMID- 15299258 TI - Uniaxial tensile testing of canine annulus fibrosus tissue under changing salt concentrations. AB - Recent modelling efforts in the field of mechanics of the intervertebral disc, demonstrate that the deformation properties of intervertebral disc tissue are intimately linked to compositional changes. This paper presents uniaxial tensile relaxation experiments of canine annulus fibrosus tissue under stepwise changes of external salt concentration. PMID- 15299259 TI - Mathematical framework for modeling tissue growth. AB - A phenomenological continuum mechanics framework for modeling growth of living tissues is proposed. Tissue is considered as an open system where mass is not conserved. The momentum balance is completed with the full-scale mass balance. Constitutive equations define simple growing materials. 'Thermoelastic' formulation of a simple growing material is specified. Within this framework traction free growth of a cylinder is considered. It is shown that the theory accommodates the case where stresses are not generated in uniform volumetric growth. It is also found that surface growth corresponds to a boundary layer solution of the governing equations. PMID- 15299260 TI - NMDA receptor expression and roles in human articular chondrocyte mechanotransduction. AB - Mechanical forces influence articular cartilage structure by regulating chondrocyte activity. Mechanical stimulation results in activation of an alpha5beta1 integrin dependent intracellular signal cascade involving focal adhesion kinase and protein kinase C, triggering the release of interleukin-4 from the cell. In normal HAC the response to physiological mechanical stimulation is characterised by increased levels of aggrecan mRNA and a decrease in levels of mRNA for matrix metalloproteinase 3 (MMP-3), the net result of which would be to maintain and optimise cartilage structure and function. This protective/anabolic response is not seen when chondrocytes from osteoarthritic cartilage are subjected to an identical mechanical stimulation regime. Following the observation that the neurotransmitter substance P is involved in chondrocyte mechanotransduction the present study was undertaken to establish potential roles for glutamate receptors in the control of chondrocyte mechanical responses. Using immunohistochemistry and RTPCR normal and OA chondrocytes are shown to express NR1 and NR2a subunits of the NMDA receptor. Addition of NMDA receptor agonists to chondrocytes in primary culture resulted in changes in membrane potential consistent with expression of functional receptors. NMDA receptor antagonists inhibited the hyperpolarisation response of normal chondrocytes to mechanical stimulation but had no effect on the depolarisation response of osteoarthritic chondrocytes to mechanical stimulation. These studies indicate that at least one subset of the NMDA receptor family of molecules is expressed in cartilage and may have important modulatory effects on mechanotransduction and cellular responses following mechanical stimulation. Indeed the results suggest that there is an alteration of NMDA receptor signalling in OA chondrocytes, which may be critical in the abnormal response of OA chondrocytes to mechanical stimulation. Thus NMDA receptors appear to be involved in the regulation of human articular chondrocyte responses to mechanical stimulation, and in OA, mechanotransduction pathways may be modified as a result of altered activation and function of these receptors. PMID- 15299261 TI - Passage in monolayer influences the response of chondrocytes to dynamic compression. AB - This study tests the hypothesis that expansion by passage in monolayer influences the response of isolated articular chondrocytes to dynamic compression. Chondrocytes, isolated from bovine articular cartilage, were seeded in monolayer and passaged 4 times (P1-4). For assessment of chondrocytic and fibroblastic phenotype, freshly isolated and passaged cells were seeded on glass coverslips or in 2% alginate beads and cultured for 7 days in DMEM + 10% FCS. Samples were assayed for DNA and GAG content and stained for collagen types I and II. In separate experiments, freshly isolated or passaged chondrocytes were seeded at 10 x 10(6) cells.ml(-1) in 4% cylindrical agarose constructs and subjected to 15% dynamic compressive strain at 1 Hz for 24 hours. [(3)H]-thymidine incorporation, SO(4) incorporation and nitrite release were analysed. Immediately following isolation (P0), chondrocytes seeded in alginate expressed high levels of type II collagen, but did not stain for type I collagen. Following repeat passage the cells expressed enhanced levels of type I collagen, with an associated reduction in type II collagen staining. These data indicate a modulation to a fibroblastic phenotype during monolayer expansion which was not rapidly reversed by culture in a 3D hydrogel. Dynamic compression down-regulated SO(4) incorporation at P0, but did not affect [(3)H]-thymidine incorporation. By contrast the incorporation of both SO(4) and [(3)H]-thymidine was enhanced by dynamic compression at both P1 and to a lesser extent P2. SO(4) and [(3)H]-thymidine incorporation were inhibited at P3 and P4. Nitrite release was down-regulated by dynamic compression at all passages. These data demonstrate a clear modulation in the response of bovine articular chondrocytes to dynamic compression following passage in monolayer. PMID- 15299262 TI - The influence and interactions of hydrostatic and osmotic pressures on the intracellular milieu of chondrocytes. AB - The intracellular milieu of chondroctyes is regulated by an array of proteins in the cell membrane which operate as transport pathways, allowing ions and nutrients such as glucose and amino acids and metabolites such as lactate to cross the plasma membrane. Here we investigated the influence of hydrostatic pressure on intracellular calcium concentrations ([Ca(2+)](i)) in isolated bovine articular chondrocytes. We found that short applications of high hydrostatic pressures led to a significant increase in [Ca(2+)](i). The pressure-induced rise was abolished for long (240 sec) but not short (30 sec) pressure applications by removal of extracellular Ca(2+). The rise in pressure was also blocked by the inhibitors neomycin and thapsigargin confirming that pressure, by generating IP(3), led to an increase in [Ca(2+)](i) by mobilising the pool of Ca(2+) ions contained within intracellular stores. We also found that intracellular [Na(+)] was affected by a rise in osmotic pressure and further affected by application of hydrostatic pressure. The effect of hydrostatic pressure on sulphate incorporation depended strongly on extracellular osmolality. Since significant gradients in extracellular osmolality exist across intact cartilage, the results imply that responses of chondrocytes to the same pressure will vary depending on location in the joint. The results also indicate that hydrostatic pressures can affect several different transporter systems thus influencing the intracellular milieu and chondrocyte metabolism. PMID- 15299263 TI - Hydrostatic pressure-induced changes in cellular protein synthesis. AB - Hydrostatic pressure is a well-known effector of cellular protein synthesis. High continuous hydrostatic pressure inhibits protein synthesis in general. It has been known for a long time that 30S ribosomal subunit is associated with the effects of pressure on protein synthesis in prokaryotes, however, the mechanisms of action are still not completely understood. Our new data suggest that synthesis of eukaryotic elongation factor-2 (eEF-2) is decreased under 30 MPa continuous hydrostatic pressure. Thus, eEF-2 may have a role in the synthesis of pressure-regulated proteins in eukaryotic cells. The presence of pressure sensitive proteins indicate that hydrostatic pressure can induce very specific responses in stressed cells. Accumulation of heat shock protein 70 and 90 beta occurs under high pressure, independent of the general inhibition of protein synthesis, although this response appears clearly weaker than during heat stress. PMID- 15299264 TI - Cycle number and waveform of fluid flow affect bovine articular chondrocytes. AB - Many studies have shown that a loading-induced (bio)physical signal regulates chondrocyte behavior. In a recent study our group has demonstrated the shear stress level- and frequency-dependent effect of sinusoidal oscillatory fluid flow on bovine articular chondrocyte (BAC) cytosolic calcium concentration ([Ca(2+)](i)), neglecting the fact that chondrocytes are not likely to see these ideal waveform in vivo or in vitro. Furthermore, possible overload of articular cartilage or excessive shear stress in chondrocyte cultures are more likely to be of a short nature. Therefore, in this study we choose to investigate a saw-tooth waveform oscillating fluid flow at varying exposure times in comparison to the established sinusoidal oscillatory waveform. [Ca(2+)](i), as an early signaling molecule, was quantified using the fluorescent dye fura-2. BAC were exposed to 1 Hz sinusoidal or saw-tooth waveform oscillating fluid flow at 2.2 Pa flow rates in a parallel plate flow chamber for 8 different loading times. As little as 5 cycles of oscillatory fluid flow were sufficient to increase [Ca(2+)](i) significantly over baseline. The number of responding cells could not be increased any further after a sufficient number of cycles (11), regardless of the waveform. Furthermore, a saw-tooth waveform appeared to be more stimulatory than regular sinusoidal oscillating flow at higher cycle numbers. BAC appear to be able to respond to these biophysical stimuli in a differentiated manner. This ability might give every single chondrocyte the capability to maintain its territory autonomously, since chondrocytes distributed in articular cartilage without the possibility to interact, e.g., via cell processes. PMID- 15299265 TI - The influence of oxygen and hydrostatic pressure on articular chondrocytes and adherent bone marrow cells in vitro. AB - Tissue engineering of articular cartilage from chondrocytes or stem cells is considered to be a potential aspect in the treatment of cartilage defects. In order to optimize culture conditions the influence of low oxygen tension (5%) - single or in combination with intermittent hydrostatic pressure (HP: 30/2 min on/off loading; 0.2 MPa) - on the biosynthetic activity (sulfate and proline incorporation) of human osteoarthritic chondrocytes cultured on collagen I/III membranes was investigated. Additionally, chondrogenesis from high density or monolayer cultures of bovine adherent bone marrow cells (aBMC) with and without chondrogenic medium supplements (CM) was analyzed by RT-PCR (mRNA expression of aggrecan and collagen type II). We could show that low oxygen tension increases significantly the biosynthesis of collagen I/III membrane-associated chondrocytes and even higher under co-stimulation with HP. While there is no chondrogenesis in monolayer cultures, CM induces expression of cartilage matrix molecules in high density cultures of aBMC which is even increased under the influence of low oxygen tension. Both, low oxygen tension and HP without CM are alone not sufficient stimuli for chondrogenesis. It can be concluded that low oxygen tension and HP might be useful tools in cartilage tissue engineering and that these physico-chemical factors promote but do not induce chondrogenesis under the given conditions. PMID- 15299266 TI - Cyclic, mechanical compression enhances chondrogenesis of mesenchymal progenitor cells in tissue engineering scaffolds. AB - The effects of cyclic, mechanical compression on human bone marrow-derived mesenchymal progenitor cells undergoing chondrogenic differentiation were examined in this study. Mesenchymal progenitor cells were injected into cylindrical biodegradable scaffolds (hyaluronan-gelatin composites), cultured in a defined, serum-free chondrogenic medium and subjected to cyclic, mechanical compression. Scaffolds were loaded for 4 hours daily in the first 7 days of culture. At 1, 7, 14 and 21 days of culture, scaffolds were harvested for reverse transcriptase Polymerase Chain Reaction (RT-PCR), histology, quantitative DNA, proteoglycan and collagen analysis. Scaffolds loaded for 7 days showed a significant upregulation especially of chondrogenic markers (type II collagen, aggrecan; p<0.0001). No significant difference could be found for DNA content between loaded samples and unloaded controls. At day 1 in culture no significant differences in proteoglycan- and collagen contents could be detected between unloaded and loaded samples. After 21 days the proteoglycan (p<0.001) and collagen contents (p<0.0001) were significantly higher in the loaded samples compared to unloaded controls. By histological analysis (toluidine blue) a higher amount of proteoglycan-rich, extracellular matrix production throughout the matrix could be detected for loaded samples compared to unloaded controls. This study indicates that cyclic, mechanical compression enhances the expression of chondrogenic markers in mesenchymal progenitor cells differentiated in vitro resulting in an increased cartilaginous matrix formation, and suggests that mechanical forces may play an important role in cartilage repair. PMID- 15299267 TI - Biochemical and mechanical properties of subchondral bone in osteoarthritis. AB - The subchondral bone has long been known to thicken in osteoarthritis. However, recent evidence has demonstrated that the turnover of the bone is increased several fold, and further suggests that the thickening occurs prior to degradation of the articular cartilage, indicating that it plays a role in the pathogenesis of osteoarthritis. The mechanical and biochemical properties of the subchondral bone are therefore of particular interest in any attempt to determine the nature of the factors initiating osteoarthritis. We have shown that the subchondral bone collagen of the femoral head possessed a 20-fold increase in turnover, as assessed by procollagen rate of synthesis and metalloproteinase degradation, and a 25% decrease in mineralisation. This increased metabolism and high lysyl hydroxylation leads to narrower and weaker fibres. Additionally the phenotypic expression of the osteoblasts is modified to produce increasing proportions of type I homotrimer in addition to the normal type I heterotrimer, which further reduces the mechanical strength of the bone. Overall, the narrow immature collagen fibres, the reduction in pyrrole cross-linking, decreased mineralisation, and increased amounts of type I homotrimer, all contribute to a weakening of the mechanical properties of the subchondral bone. PMID- 15299268 TI - Adaptation of subchondral bone in osteoarthritis. AB - Osteoarthritis is a chronic joint disease with pathological changes in the articulating cartilage and all other tissues that occupy the joint. Radin and coworkers have suggested the involvement of subchondral bone in the disease process. However, evidence for an essential role in the etiology has never been proven. Recent studies showing reduced chemical and mechanical properties of subchondral bone in various stages of the disease have invigorated interest in the role of subchondral bone in the development and progression of the disease. The current study showed that the concept of bone adaptation might explain subchondral stiffening, a process where subchondral bone becomes typically sclerotic in osteoarthritis. In addition, we report reduced mechanical matrix tissue properties as well as an increase in denatured collagen content. In conclusion, although osteoarthritic bone tissue contains increased denatured collagen and has reduced matrix mechanical properties, the widely accepted concept of subchondral stiffening is compatible with the process of normal bone adaptation. PMID- 15299269 TI - Organ printing: fiction or science. AB - Aggregates of living cells (i.e. model tissue fragments) under appropriate conditions fuse like liquid drops. According to Steinberg's differential adhesion hypothesis (DAH), this may be understood by assuming that cells are motile and tissues made of such cells possess an effective surface tension. Here we show that based on these properties three-dimensional cellular structures of prescribed shape can be constructed by a novel method: cell aggregate printing. Spherical aggregates of similar size made of cells with known adhesive properties were prepared. Aggregates were embedded into biocompatible gels. When the cellular and gel properties, as well as the symmetry of the initial configuration were appropriately adjusted the contiguous aggregates fused into ring-like organ structures. To elucidate the driving force and optimal conditions for this pattern formation, Monte Carlo simulations based on a DAH motivated model were performed. The simulations reproduced the experimentally observed cellular arrangements and revealed that the control parameter of pattern evolution is the gel-tissue interfacial tension, an experimentally accessible parameter. PMID- 15299270 TI - The influence of cyclic tension amplitude on chondrocyte matrix synthesis: experimental and finite element analyses. AB - While not generally viewed as physiologically significant in articular cartilage, substantial tension can develop in fibrocartilage structures and in articular cartilage injuries. This study examined how different amplitudes of cyclic tension influence chondrocyte matrix synthesis. Bovine articular chondrocytes seeded in fibrin gels were loaded continuously for 48 hours at 1.0 Hz with displacements of 5%, 10%, or 20%. Protein and proteoglycan synthesis were measured by (3)H-proline and (35)S-sulfate incorporation, respectively. A poroelastic finite element model of the fibrin gel was developed to determine the strain distributions, hydrostatic pressures, and fluid velocities within the constructs at the various levels of displacement. Compared to unloaded controls, 10% and 20% displacements inhibited proteoglycan synthesis to the same extent, while 5% displacement had no effect. Tensile loading did not significantly affect protein synthesis. The finite element model predicted a wide range of strains and fluid velocities within the region of the gel analyzed for matrix synthesis, and the ranges overlapped for the different levels of displacement. These results indicate that the cyclic tension amplitude influences chondrocyte proteoglycan synthesis and that there may be a threshold in the response. PMID- 15299271 TI - Adipose-derived adult stem cells for cartilage tissue engineering. AB - Tissue engineering is a promising therapeutic approach that uses combinations of implanted cells, biomaterial scaffolds, and biologically active molecules to repair or regenerate damaged or diseased tissues. Many diverse and increasingly complex approaches are being developed to repair articular cartilage, with the underlying premise that cells introduced exogenously play a necessary role in the success of engineered tissue replacements. A major consideration that remains in this field is the identification and characterization of appropriate sources of cells for tissue-engineered repair of cartilage. In particular, there has been significant emphasis on the use of undifferentiated progenitor cells, or "stem" cells that can be expanded in culture and differentiated into a variety of different cell types. Recent studies have identified the presence of an abundant source of stem cells in subcutaneous adipose tissue. These cells, termed adipose derived adult stem (ADAS) cells, show characteristics of multipotent adult stem cells, similar to those of bone marrow derived mesenchymal stem cells (MSCs), and under appropriate culture conditions, synthesize cartilage-specific matrix proteins that are assembled in a cartilaginous extracellular matrix. The growth and chondrogenic differentiation of ADAS cells is strongly influenced by factors in the biochemical as well as biophysical environment of the cells. Furthermore, there is strong evidence that the interaction between the cells, the extracellular biomaterial substrate, and growth factors regulate ADAS cell differentiation and tissue growth. Overall, ADAS cells show significant promise for the development of functional tissue replacements for various tissues of the musculoskeletal system. PMID- 15299272 TI - The effect of media perfusion on three-dimensional cultures of human chondrocytes: integration of experimental and computational approaches. AB - This work examines the effect of perfusion on human mature articular chondrocytes cultured on synthetic biodegradable scaffolds (DegraPol). Human chondrocytes were isolated, seeded on the scaffolds and subjected to perfused culture at a flow rate of 0.5 ml/min, corresponding to an average inlet fluid velocity of 44 microm/s, with flow inversion every 1 minute. The flow was imposed at the construct surface in some constructs, it was forced through the construct thickness in other constructs and was absent in the static controls. We compared cell viability and morphology and we evaluated material properties of the constructs at 1 month of culture. Thickness-perfused constructs showed significantly higher material properties and roughly a two-fold cell viability, when compared both to surface-perfused constructs and to static controls. Chondrocytes maintained a phenotypic morphology in all experiments, probably favoured by a limited cell-scaffold interaction. Biosynthetic activity could be demonstrated only in the bioreactor-cultured constructs. In this experimental model, a bi-directional flow of culture medium was applied to the cells at a macroscopic level and computational modelling was used to quantify the fluid dynamic environment induced on the cells at a microscopic level. This method may be used to quantify the effects of fluid-dynamic shear on the growth modulation of tissue-engineered cartilage constructs, to potentially enhance tissue growth in vitro. PMID- 15299273 TI - Considerations on the use of ear chondrocytes as donor chondrocytes for cartilage tissue engineering. AB - Articular cartilage is often used for research on cartilage tissue engineering. However, ear cartilage is easier to harvest, with less donor-site morbidity. The aim of this study was to evaluate whether adult human ear chondrocytes were capable of producing cartilage after expansion in monolayer culture. Cell yield per gram of cartilage was twice as high for ear than for articular cartilage. Moreover, ear chondrocytes proliferated faster. Cell proliferation could be further stimulated by the use of serum-free medium with Fibroblast Growth Factor 2 (FGF2) in stead of medium with 10% serum. To evaluate chondrogenic capacity, multiplied chondrocytes were suspended in alginate and implanted subcutaneously in athymic mice. After 8 weeks the constructs demonstrated a proteoglycan-rich matrix that contained collagen type II. Constructs of ear chondrocytes showed a faint staining for elastin. Quantitative RT-PCR revealed that expression of collagen type II was 2-fold upregulated whereas expression of collagen type I was 2-fold down regulated in ear chondrocytes expanded in serum-free medium with FGF2 compared to serum-containing medium. Expression of alkaline phosphatase and collagen type X were low indicating the absence of terminal differentiation. We conclude that ear chondrocytes can be used as donor chondrocytes for cartilage tissue engineering. Furthermore, it may proof to be a promising alternative cell source to engineer cartilage for articular repair. PMID- 15299274 TI - Expression of adhesion molecules and collagen on rat chondrocyte seeded into alginate and hyaluronate based 3D biosystems. Influence of mechanical stresses. AB - Chondrocytes use mechanical signals, via interactions with their environment, to synthesize an extracellular matrix capable to withstanding high loads. Most chondrocyte-matrix interactions are mediated via transmembrane receptors such as integrins or non-integrins receptors (i.e. annexin V and CD44). The aim of this study was to analyze, by flow cytometry, the adhesion molecules (alpha5/beta1 integrins and CD44) on rat chondrocytes seeded into 3D biosystem made of alginate and hyaluronate. These biosystems were submitted to mechanical stress by knocking the biosystems between them for 48 hours. The expression of type I and type II collagen was also evaluated. The results of the current study showed that mechanical stress induced an increase of type II collagen production and weak variations of alpha5/beta1 receptors expression no matter what biosystems. Moreover, our results indicated that hyaluronan receptor CD44 expression depends on extracellular matrix modifications. Thus, these receptors were activated by signals resulted from cell environment variations (HA addition and modifications owing to mechanical stress). It suggested that this kind of receptor play a crucial role in extracellular matrix homeostasis. Finally, on day 24, no dedifferentiation of chondrocytes was noted either in biosystems or under mechanical stress. For all biosystems, the neosynthesized matrix contained an important level of collagen, which was type II, whatever biosystems. In conclusion, it appeared that the cells, under mechanical stress, maintained their phenotype. In addition, it seems that, on rat chondrocytes, alpha5/beta1 integrins did not act as the main mechanoreceptor (as described for human chondrocytes). In return, hyaluronan receptor CD44 seems to be in relation with matrix composition. PMID- 15299275 TI - Cartilage repair: surgical techniques and tissue engineering using polysaccharide and collagen-based biomaterials. AB - Lesions of articular cartilage have a large variety of causes among which traumatic damage, osteoarthritis and osteochondritis dissecans are the most frequent. Replacement of articular defects in joints has assumed greater importance in recent years. This interest results in large part because cartilage defects cannot adequately heal themselves. Many techniques have been suggested over the last 30 years, but none allows the regeneration of the damaged cartilage, i.e. its replacement by a strictly identical tissue. In the first generation of techniques, relief of pain was the main concern, which could be provided by techniques in which cartilage was replaced by fibrocartilage. Disappointing results led investigators to focus on more appropriate bioregenerative approaches using transplantation of autologous cells into the lesion. Unfortunately, none of these approaches has provided a perfect final solution to the problem. The latest generation of techniques, currently in the developmental or preclinical stages, involve biomaterials for the repair of chondral or osteochondral lesions. Many of these scaffolds are designed to be seeded with chondrocytes or progenitor cells. Among natural and synthetic polymers, collagen- and polysaccharide-based biomaterials have been extensively used. For both these supports, studies have shown that chondrocytes maintain their phenotype when cultured in three dimensions. In both types of culture, a glycosaminoglycan-rich deposit is formed on the surface and in the inner region of the cultured cartilage, and type II collagen synthesis is also observed. Dynamic conditions can also improve the composition of such three-dimensional constructs. Many improvements are still required, however, in a number of key aspects that so far have received only scant attention. These aspects include: adhesion/integration of the graft with the adjacent native cartilage, cell seeding with genetically-modified cell populations, biomaterials that can be implanted without open joint surgery and combined therapies, aimed at disease modification, pain relief and reduction of inflammation. PMID- 15299276 TI - Long-term culture of tissue engineered cartilage in a perfused chamber with mechanical stimulation. AB - One approach to functional tissue engineering of cartilage is to utilize bioreactors to provide environmental conditions that stimulate chondrogenesis in cells cultured on biomaterial scaffolds. We report the combined use of a three dimensional in vitro model and a novel bioreactor with perfusion of culture medium and mechanical stimulation in long-term studies of cartilage development and function. To engineer cartilage, scaffolds made of a non-woven mesh of polyglycolic acid (PGA) were seeded with bovine calf articular chondrocytes, cultured for an initial 30-day period under free swelling conditions, and cultured for an additional 37 day period in one of the three groups: (1) free swelling, (2) static compression (on 24 h/day, strain control, static offset 10%), and (3) dynamic compression (on 1 h/day; off 23 h/day; strain control, static offset 2%, dynamic strain amplitude 5%; frequency 0.3 Hz). Constructs were sampled at timed intervals and assessed with respect to structure, biochemical composition, and mechanical function. Mechanical simulation had little effect on the compositions, morphologies and on mechanical properties of construct interiors discs, but it resulted in distincly different properties of the peripheral rings and face sides. Contructs cultured with mechanical loading maintained their cylindrical shape with flat and parallel top and bottom surfaces, and retained larger amounts of GAG. The modular bioreactor system with medium perfusion and mechanical loading can be utilized to define the conditions of cultivation for functional tissue engineering of cartilage. PMID- 15299277 TI - Spectral and lifetime fluorescence imaging microscopies: new modalities of multiphoton microscopy applied to tissue or cell engineering. AB - Spectral and multiphoton imaging is the preferred approach for non-invasive study allowing deeper penetration to image molecular processes in living cells. But currently available fluorescence microscopic techniques based on fluorescence intensity, such as confocal or multiphoton excitation, cannot provide detailed quantitative information about the dynamic of complex cellular structure (molecular interaction). Due to the variation of the probe concentration, photostability, cross-talking, its effects cannot be distinguished in simple intensity images. Therefore, Time Resolved fluorescence image is required to investigate molecular interactions in biological systems. Fluorescence lifetimes are generally absolute, sensitive to environment, independent of the concentration of the probe and allow the use of probes with overlapping spectra but that not have the same fluorescence lifetime. In this work, we present the possibilities that are opened up by Fluorescence Lifetime Imaging Microscopy, firstly to collect images based on fluorescence lifetime contrast of GFP variants used as a reporter of gene expression in chondrocytes and secondly, to measure molecular proximity in erythrocyte (glycophorin/membrane) by Fluorescence Resonance Energy Transfer (FLIM-FRET). PMID- 15299278 TI - Human mesenchymal stem cells suppress induction of cytotoxic response to alloantigens. AB - Mesenchymal stem cells (MSC) fail to induce allogeneic responses in mixed lymphocyte reaction assays. Because MSC express HLA class I molecules, here we investigated whether they could be recognized as allogeneic targets by cytolytic T lymphocytes (CTL). With this aim, CTL precursor (CTLp) frequencies were measured following stimulation of T cells with either allogeneic mononuclear cells (MNC) or MSC originated from the same human bone marrow donor. Lysis of MSC was measured at day 10 of culture in standard chromium release assays. In addition, allogeneic PHA blast T cells or B-EBV lymphoblastoid cell lines (LCLs) generated from the same donor were used as positive controls of lysis. Our results showed that when allogeneic MNC were used to stimulate T cells, a high CTLp frequency was detected towards MSC targets. However, when MSC were used as stimulators, CTLp frequencies were markedly altered whatever the targets used, i.e.: MSC, PHA blast T cells or EBV-B LCLs. Moreover, when graded concentrations of MSC were added together with MNC upon stimulation of alloreactive T cells, we observed a dose-dependent decrease in CTLp frequencies towards MSC targets. This inhibition of MSC lysis was partially overcome by adding exogenous rh-IL-2 from the beginning of cultures. In addition, this suppressive effect was totally reproduced when, instead of MSC, supernatant harvested from MSC cultures was added to allogeneic MNC, upon stimulation of alloreactive T cells. In conclusion, our results demonstrate that MSC which can be recognized as targets by pre activated alloreactive CTLs, may be able to suppress differentiation of CTL precursors into CTL effectors through secretion of suppressive factors. PMID- 15299279 TI - Post-traumatic osteoarthritis: the role of accelerated chondrocyte senescence. AB - Joint injuries frequently lead to progressive joint degeneration that causes the clinical syndrome of post-traumatic osteoarthritis. The pathogenesis of osteoarthritis remains poorly understood, but patient age is a significant risk factor for progressive joint degeneration. We have found that articular cartilage chondrocytes show strong evidence of senescence with increasing age, including synthesis of smaller more irregular aggrecans; increased expression of lysosomal beta-galactosidase and telomere erosion; and decreased proteoglycan synthesis, response to the anabolic cytokine IGF-I, proliferative capacity, and mitochondrial function. These observations help explain the strong association between age and joint degeneration, but they do not explain how joint injury increases the risk of joint degeneration in younger individuals. We hypothesized that excessive loading of articular surfaces due to acute joint trauma or post traumatic joint instability, incongruity or mal-alignment increases release of reactive oxygen species, and that the increased oxidative stress on chondrocytes accelerates chondrocyte senescence thereby decreasing the ability of the cells to maintain or restore the tissue. To test this hypothesis, we exposed human articular cartilage chondrocytes from young adults to mechanical and oxidative stress. We found that shear stress applied to cartilage explants in a triaxial pressure vessel increased release of reactive oxygen species and oxidative stress induced chondrocyte senescence (as measured by expression of lysosomal beta galactosidase, nuclear and mitochondrial DNA damage and decreased mitochondrial function). These observations support the hypothesis that joint injury accelerates chondrocyte senescence and that this acceleration plays a role in the joint degeneration responsible for post-traumatic osteoarthritis. PMID- 15299280 TI - Hyaluronan injection affects neither osteoarthritis progression nor loading of the OA knee in dogs. AB - We previously reported that intraarticular injections of hyaluronan (HA), administered prophylactically to dogs in whom knee osteoarthritis had been induced by transection of the anterior cruicate ligament, did not significantly modify the intraarticular pathology but decreased the proteogylcan concentration of the articular cartilage by as much as 30%. Because the cartilage proteoglycan concentration is directly related to the stiffness of the tissue, these results raised the possibility that intraarticular HA therapy could exacerbate OA. In the present study, using a different HA formulation, with a longer interval between intraarticular HA injection and examination of joint tissues, we found that neither prophylactic nor therapeutic administration of HA had an effect on the severity of OA pathology, the magnitude of vertical ground reaction forces generated by the unstable hind limb (a surrogate for joint pain), or the cartilage proteoglycan concentration. The data suggest that the suppression of proteoglycan synthesis induced by HA is temporary and fully reversible and that HA injections do not result in overloading of the OA extremity. A significant correlation was noted between the severity of chondropathy and the magnitude of the vertical ground reaction forces generated by the unstable limb. PMID- 15299281 TI - Lubricin reduces cartilage--cartilage integration. AB - Cartilage integration in vivo does not occur, such that even cartilage fissures do not heal. This could be due not only to the limited access of chondrocytes to the wound, but also to exogenous factors. In this paper, we tested the hypothesis that lubricin, a lubricating protein physiologically present in the synovial fluid, reduces the integrative cartilage repair capacity. Disk/ring composites of bovine articular cartilage were prepared using concentric circular blades and cultured for 6 weeks with or without treatment with 250 microg/ml lubricin applied three times per week. Following culture, the percentage of contact area between the disks and the rings, as assessed by light microscopy, were equal in both groups. The adhesive strength of the integration interface, as assessed by push-out mechanical tests, was markedly and significantly lower in lubricin treated specimens (2.5 kPa) than in the controls (28.7 kPa). Histological observation of Safranin-O stained cross-sections confirmed the reduced integration in the lubricin treated composites. Our findings suggest that the synovial milieu, by providing lubrication of cartilage surfaces, impairs cartilage--cartilage integration. PMID- 15299282 TI - Short-term changes in cell and matrix damage following mechanical injury of articular cartilage explants and modelling of microphysical mediators. AB - The short-term responses of articular cartilage to mechanical injury have important implications for prevention and treatment of degenerative disease. Cell and matrix responses were monitored for 11 days following injurious compression of cartilage in osteochondral explants. Injury was applied as a single ramp compression to 14 MPa peak stress at one of three strain rates: 7 x 10(-1), 7 x 10(-3) or 7 x 10(-5) s(-1). Responses were quantified in terms of the appearance of macroscopic matrix cracks, changes in cell viability, and changes in cartilage wet weights. Loading at the highest strain rate resulted in acute cell death near the superficial zone in association with cracks, followed over the 11 days after compression by a gradual increase in cell death and loss of demarcation between matrix zones containing viable versus nonviable cells. In contrast, loading at the lowest strain rate resulted in more severe, nearly full-depth cell death acutely, but with no apparent worsening over the 11 days following compression. Between days 4 and 11, all mechanically injured explants significantly increased in wet weight, suggesting loss of matrix mechanical integrity independent of compression strain rate. Results demonstrate that short-term responses of cartilage depend upon the biomechanical characteristics of injurious loading, and suggest multiple independent pathways of mechanically-induced cell death and matrix degradation. Modifications to an existing fiber-reinforced poroelastic finite element model were introduced and the model was used for data interpretation and identification of microphysical events involved in cell and matrix injury. The model performed reasonably well at the slower strain rates and exhibited some capacity for anticipating the formation of superficial cracks during injurious loading. However, several improvements appear to be necessary before such a model could reliably be used to draw upon in vitro experimental results for prediction of injurious loading situations in vivo. PMID- 15299283 TI - Induction of heat shock protein 70 (Hsp70) by proteasome inhibitor MG 132 protects articular chondrocytes from cellular death in vitro and in vivo. AB - The aim of this work was to determine whether Hsp70 overexpression via proteasome inhibitor MG132 was able to protect chondrocytes towards mono-iodoacetate (MIA) cytotoxicity both in vitro and in vivo. In vitro, overexpression of Hsp70 via MG132 was significantly able to protect chondrocytes from MIA toxicity (MTT/LDH analyses). Hsp70 essentially mediated this chondroprotective effect as demonstrated by antisense strategy. In vivo, chondrocytic overexpression of Hsp70, after a preventive intra-articular injection of MG132 in rat knee, was sufficient to decrease the severity of OA-induced MIA lesions, as demonstrated histologically and biochemically. In conclusion, intracellular overexpression of Hsp70, through proteasome inhibition, could be an interesting tool in protecting chondrocytes from cellular injuries, either necrotic or apoptotic in nature, and thus might be a novel chondroprotective modality in rat experimental OA. PMID- 15299284 TI - Modulation of collagen synthesis in normal and osteoarthritic cartilage. AB - In osteoarthritic cartilage, chondrocytes are able to present heterogeneous cellular reactions with expression and synthesis of the (pro)collagen types characteristic of prechondrocytes (type IIA), hypertrophic chondrocytes (type X), as well as differentiated (types IIB, IX, XI, VI) and dedifferentiated (types I, III) chondrocytes. The expression of type IIA procollagen in human osteoarthritic cartilage support the assumption that OA chondrocytes reverse their phenotype towards a chondroprogenitor phenotype. Recently, we have shown that dedifferentiation of mouse chondrocytes induced by subculture was associated with the alternative splicing of type II procollagen pre-mRNA with a switch from the IIB to the IIA form. In this context, we demonstrated that BMP-2 favours expression of type IIB whereas TGF-beta1 potentiates expression of type IIA induced by subculture. These data reveal the specific capability of BMP-2 to reverse the program of chondrocyte dedifferentiation. This interesting feature needs to be tested with human chondrocytes since cell amplification is required for the currently used autologous chondrocyte transplantation. PMID- 15299285 TI - Type II collagen peptides for measuring cartilage degradation. AB - This paper describes two new immunoassays for a peptide of the triple helix of type II collagen (Coll 2-1) and its nitrated form (Coll 2-1 NO(2)). In healthy subjects aged between 20 and 65 years old, Coll 2-1 and Coll 2-1 NO(2) levels in serum were in means 125.13+/-3.71 and 0.16+/-0.08 nmol/l, respectively. These levels did not significantly vary with age. However, up to 45 years of age, Coll 2-1 NO(2) levels in women were significantly higher than in men. In patients with knee osteoarthritis (OA), Coll 2-1 in serum was found to be elevated compared to healthy controls (267.45+/-26.42 nmol/l vs 126.78+/-6.61 nmol/l). Further, we have demonstrated that an increase of the urinary levels of Coll 2-1 or Coll 2-1 NO(2) over 1 year was predictive of joint space narrowing progression in OA patients. In conclusion, these preliminary results indicate that Coll 2-1 could be a predictive marker of knee OA progression. PMID- 15299286 TI - Articular chondrocytes cultured in hypoxia: their response to interleukin-1beta and rhein, the active metabolite of diacerhein. AB - In the present report, we show that bovine articular chondrocytes cultured in low oxygen tension, i.e. in conditions mimicking their hypoxic in vivo environment, respond to IL-1beta (10 ng/ml) by an increased DNA binding activity of NF-kappaB and AP-1 transcription factors. Incubation of the cells with 10(-5) M rhein for 24 h was found to reduce this activity, particularly in the case of AP-1. Mitogen activated kinases (ERK-1 and ERK-2) were activated by exposure of the chondrocytes to 1-h treatment with IL-1beta. This effect was greater in hypoxia (3% O(2)) than in normoxia (21% O(2)). Rhein was capable of reducing the IL-1beta stimulated ERK1/ERK2 pathway whatever the tension of oxygen present in the environment. The mRNA steady-state levels of collagen type II (COL2A1) and aggrecan core protein were found to be significantly increased by a 24-h treatment with 10(-5) M rhein. This stimulating effect was also observed in the presence of IL-1beta, suggesting that the drug could prevent or reduce the IL 1beta-induced inhibition of extracellular matrix synthesis. IL-1-induced collagenase (MMP1) expression was significantly decreased by rhein in the same conditions. In conclusion, rhein can effectively inhibit the IL-1-activated MAPK pathway and the binding of NF-kappaB and AP-1 transcription factors, two key factors involved in the expression of several pro-inflammatory genes by chondrocytes. In addition, the drug can reduce the procatabolic effect of the cytokine, by reducing the MMP1 synthesis, and enhance the synthesis of matrix components, such as type II collagen and aggrecan. These results may explain the anti-osteoarthritic properties of rhein and its disease-modifying effects on OA cartilage, in spite of absence of inhibition at prostaglandin level. PMID- 15299287 TI - ATP in the mechanotransduction pathway of normal human chondrocytes. AB - Extracellular nucleotides have been shown to have diverse effects on chondrocyte function, generally acting via P2 purinoceptors. We have previously shown that mechanical stimulation at 0.33 Hz of normal human chondrocyte cultures causes cellular hyperpolarisation, while chondrocytes derived from osteoarthritic (OA) cartilage depolarise. Experiments have been undertaken to establish whether ATP is involved in the response of the chondrocyte to mechanical stimulation. Chondrocytes, isolated from normal and OA cartilage obtained, with consent, from human knee joints following surgery, were cultured in non-confluent monolayer. Cells were mechanically stimulated at 0.33 Hz for 20 minutes at 37 degrees C in the presence or absence of inhibitors of ATP signalling, or were stimulated by the addition of exogenous ATP or derivatives, and electrophysiological measurements recorded. Samples of medium bathing the cells were collected before and after mechanical stimulation, and the concentration of ATP in the cell medium was measured. Total RNA was extracted from cultured chondrocytes, reverse transcribed and used for RT-PCR with primers specific for P2Y2 purinoceptors. ATP, UTP 2-methylthioadenosine and alphabeta-methylene adenosine 5'-triphosphate all induced a hyperpolarisation response in normal human articular chondrocytes. No significant change was observed in the membrane potentials of chondrocytes isolated from OA cartilage following the addition of these nucleotides to the medium. In normal chondrocytes, the hyperpolarisation induced by ATP was blocked by the presence of apamin, indicating the involvement of small-conductance calcium-activated potassium channels. Following mechanical stimulation of normal chondrocytes, an increase was observed in ATP concentration in the cell culture medium bathing the cells. The presence within the culture medium of suramin or pyridoxal-phosphate-6-azophenyl-2',4'-disulphonic acid (PPADS) prior to and during the period of mechanical stimulation abolished the hyperpolarisation response in normal chondrocytes. The presence of mRNA for P2Y2 purinoceptors was demonstrated in both normal and OA chondrocytes by RT-PCR. These results suggest that ATP has a role in the response of normal chondrocytes to mechanical stimulation, via P2Y2 purinoceptors. This response appears to be different in chondrocytes derived from OA cartilage, and may be important in the progression of this disease. PMID- 15299288 TI - Functional tissue engineering of chondral and osteochondral constructs. AB - Due to the prevalence of osteoarthritis (OA) and damage to articular cartilage, coupled with the poor intrinsic healing capacity of this avascular connective tissue, there is a great demand for an articular cartilage substitute. As the bearing material of diarthrodial joints, articular cartilage has remarkable functional properties that have been difficult to reproduce in tissue-engineered constructs. We have previously demonstrated that by using a functional tissue engineering approach that incorporates mechanical loading into the long-term culture environment, one can enhance the development of mechanical properties in chondrocyte-seeded agarose constructs. As these gel constructs begin to achieve material properties similar to that of the native tissue, however, new challenges arise, including integration of the construct with the underlying native bone. To address this issue, we have developed a technique for producing gel constructs integrated into an underlying bony substrate. These osteochondral constructs develop cartilage-like extracellular matrix and material properties over time in free swelling culture. In this study, as a preliminary to loading such osteochondral constructs, finite element modeling (FEM) was used to predict the spatial and temporal stress, strain, and fluid flow fields within constructs subjected to dynamic deformational loading. The results of these models suggest that while chondral ("gel alone") constructs see a largely homogenous field of mechanical signals, osteochondral ("gel bone") constructs see a largely inhomogeneous distribution of mechanical signals. Such inhomogeneity in the mechanical environment may aid in the development of inhomogeneity in the engineered osteochondral constructs. Together with experimental observations, we anticipate that such modeling efforts will provide direction for our efforts aimed at the optimization of applied physical forces for the functional tissue engineering of an osteochondral articular cartilage substitute. PMID- 15299289 TI - Current perspectives on cartilage and chondrocyte mechanobiology. AB - It is well known that physiological forces are essential for the maintenance of normal composition and structure of articular cartilage. Although some of the mechanisms of mechanotransduction are known today, there are certainly many others left unrevealed. In order to understand the complicated systems present in articular cartilage, we have to bring together the data from all fields of cartilage mechanobiology. The 3rd Symposium on Mechanobiology of Cartilage and Chondrocyte was a good effort towards that goal. PMID- 15299291 TI - NMR - this other method for protein and nucleic acid structure determination. AB - For a quarter of a century X-ray diffraction in single crystals was unique in its ability to solve three-dimensional structures of proteins and nucleic acids at atomic resolution. The situation changed in 1984 with the completion of a protein structure determination by nuclear magnetic resonance (NMR) spectroscopy in solution, and today NMR is a second widely used method for biomacromolecular structure determination. This review describes the method of NMR structure determination of biological macromolecules, and attempts to place NMR structure determination in perspective with X-ray crystallography. NMR is most powerful for studies of relatively small systems with molecular weights up to about 30000, but these structures can be obtained in near-physiological milieus. The two techniques have widely different time scales which afford different insights into internal molecular mobility as well as different views of protein or nucleic acid molecular surfaces and hydration. Generally, in addition to information on the average three-dimensional structure, NMR provides information on a wide array of short-lived transient conformational states. Combining information from the two methods can yield a more detailed insight into the structural basis of protein and nucleic acid functions, and thus provide a more reliable platform for rational drug design and the engineering of novel protein functions. PMID- 15299292 TI - Use of glycerol, polyols and other protein structure stabilizing agents in protein crystallization. AB - A protein preparation to be used for crystallization should be homogeneous and should remain so throughout the course of a prolonged crystallization experiment. General methods for preparation of pure proteins and for prevention of their covalent modification (through proteolysis, sulfhydryl oxidation, etc.) during prolonged incubation are well known. Crystallographers are less aware of general methods for stabilization of proteins against non-covalent modifications (partial denaturation, heterogeneous aggregation) which can also introduce structural heterogeneity into a protein preparation. Related to this issue are methods to suppress protein conformational flexibility which can be a source of dynamic structural heterogeneity and which presents an entropic barrier to crystallization. However, for many years agents which stabilize protein structure have been described in the biochemical literature. Recently the most widely used of these structure-stabilizing agents, glycerol, was used to crystallize T7 RNA polymerase. The observation that this compound has general structure-stabilizing effects and that it was essential for crystallization of at least this one protein led to the suggestion that it might be generally useful in crystallizing flexible proteins and in inducing order in disordered segments of crystalline proteins. Subsequently, glycerol was used with good effect in the crystallization of a number of proteins. Other recent results suggest that soaking crystals in solutions containing glycerol can have 'structure-ordering' effects on the crystalline protein. These observations support the utility of glycerol in protein crystallization and suggest that the information in the biochemical literature on protein structure-stabilizing agents will find useful application in the field of protein crystal growth. PMID- 15299293 TI - Shaped protein single crystals. AB - The formation of protein single crystals grown with the shape controlled by the geometry of the capillary used as a growth cell is presented. The shaped crystals show strong birefringence under crossed nicols and diffract as single crystals up to 1.74 A. PMID- 15299294 TI - Use of iron anomalous scattering with multiple models and data sets to identify and refine a weak molecular replacement solution: structure analysis of cytochrome c' from two bacterial species. AB - The structure of cytochrome c' from two bacterial species, Alcaligenes sp and Alcaligenes denitrificans, have been determined from X-ray diffraction data to 3.0 A resolution using the anomalous scattering of the single Fe atom in each to identify and refine a weak molecular-replacement solution. Molecular-replacement studies, with the program AMORE, used two isomorphous data sets (from the two species), two independent search models (the cytochromes c' from Rhodospirillum molischianum and Rhodospirillum rubrum), both with and without side chains, and two different resolution ranges (10.0-4.0 and 15.0-3.5A) to generate a large number of potential solutions. No single solution stood out and none appeared consistently. The Fe-atom position in each structure was then determined from its anomalous-scattering contribution and all molecular- replacement solutions were discarded which did not (i) place the Fe atom correctly and (ii) orient the molecule such that a crystallographic twofold axis generated a dimer like those of the two search models. Finally, electron-density maps phased solely by the Fe atom anomalous scattering were calculated. As these were combined and subjected to solvent flattening and histogram matching (with the program SQUASH), correlation with the remaining molecular-replacement solutions identified one as correct and enabled it to be improved and subjected to preliminary refinement. The correctness of the solution is confirmed by parallel isomorphous-replacement studies. PMID- 15299295 TI - Structure of 6-phosphogluconate dehydrogenase refined at 2 A resolution. AB - The X-ray unliganded structure of 6-phosphogluconate dehydrogenase (E.C. 1.1.1.44) (6-PGDH) from sheep liver has been determined at 2A resolution and refined to a final R-factor of 19.8% for 35 031 unique reflections. The enzyme is dimeric, each subunit being comprised of an N-terminal coenzyme-binding domain with a Rossmann fold, a large all-helical domain and a small C-terminal tail. The model contains 473 residues, three sulfate ions and 346 water molecules; the two best defined sulfates are found in the active site. This structure, based on improved diffraction data, is an extension of the 2.5 A, resolution model reported earlier. It has good geometry with 92% of the residues falling in the most favoured areas of the Ramachandran plot. Several unusual features are discussed: the incorporation of an alanine in place of the second conserved glycine of the dinucleotide-binding fingerprint; a duplicated five-helix motif which is unique to this enzyme; an extended water network at the dimer interface and a C-terminal tail which is incorporated within the second subunit, forming not only a major part of the dimer interface but also part of the active site. PMID- 15299296 TI - Using sampling techniques in protein crystallization. AB - The crystallization of homogeneous or highly purified macromolecules depends on many variables such as precipitant, pH, choice of buffer, protein concentration, temperature, the participation of different mono- and divalent ions, as well as the presence of minute amounts of detergent and organic molecules. Finding the best combination among these many parameters is a multi-variable optimization problem. This kind of problem can be treated mathematically by sampling techniques. We have used this technique for protein crystallization. The iterative procedure starts with random sampling, followed by quantitative evaluation and cycles with weighted sampling. A simple procedure, derived from this concept and called MON48, has been successfully applied to many protein crystallization problems. PMID- 15299297 TI - Structure of a calcium-independent phospholipase-like myotoxic protein from Bothrops asper venom. AB - Myotoxin II, a myotoxic calcium-independent phospholipase-like protein isolated from the venom of Bothrops asper, possesses no detectable phospholipase activity. The crystal structure has been determined and refined at 2.8 A to an R-factor of 16.5% (F > 3sigma) with excellent stereochemistry. Amino-acid differences between catalytically active phospholipases and myotoxin II in the Ca(2+)-binding region, specifically the substitutions Tyr28-->Asn, Gly32-->Leu and Asp49-->Lys, result in an altered local conformation. The key difference is that the epsilon-amino group of Lys49 fills the site normally occupied by the calcium ion in catalytically active phospholipases. In contrast to the homologous monomeric Lys49 variant from Agkistrodon piscivorus piscivorus, myotoxin II is present as a dimer both in solution and in the crystalline state. The two molecules in the asymmetric unit are related by a nearly perfect twofold axis, yet the dimer is radically different from the dimer formed by the phospholipase from Crotalus atrox. Whereas in C. atrox the dimer interface occludes the active sites, in myotoxin II they are exposed to solvent. PMID- 15299298 TI - Structure of the trigonal form of recombinant oxidized flavodoxin from Anabaena 7120 at 1.40 A resolution. AB - The oxidized recombinant flavodoxin from the cyanobacterium Anabaena 7120 has been crystallized in a trigonal form. The recombinant protein has an identical primary structure to that purified directly from Anabaena, which functions as a substitute for ferredoxin in an iron-deficient environment for electron transfer from photosystem I to ferredoxin-NADP(+) reductase. X-ray data to 1.40 A were collected on a Siemens area detector. Of the 311 379 reflections collected, 36069 reflections were unique in space group P3(1)21 (a = 55.36, c = 102.59 A) with an R(merge) of 3.8%. The structure was solved by molecular replacement using coordinates from the wild-type monoclinic structure previously solved in this laboratory [Rao, Shaffie, Yu, Satyshur, Stockman & Markley (1992). Protein Sci. 1, 1413-1427]. The structure was refined with X-PLOR and SHELXL93 to a crystallographic R-factor of 13.9% for 32963 reflections with I> 2sigma(I). The final structure contains 2767 atoms including 31 flavin mononucleotide (FMN) atoms, 299 water molecules, and one sulfate ion. The protein is comprised of a central five-stranded beta-sheet surrounded by five helices and binds a single molecule of FMN at the C-terminus of the sheet. The trigonal protein structure and the crystal packing are compared with the monoclinic wild-type protein. Helix alpha3 in this structure is less distorted than in the monoclinic structure and shows additional hydrogen bonds in the N-terminal portion of the helix. The trigonal structure is extensively hydrogen bonded in three major areas with neighboring molecules compared with five regions in the monoclinic structure, but using significantly fewer hydrogen bonds to stabilize the lattice. There are several hydrogen bonds to the amide groups from water molecules several of which stabilize and extend the ends of the beta-sheet. PMID- 15299299 TI - A strategy for rapid and effective refinement applied to black swan lysozyme. AB - The crystal structure of a goose-type lysozyme from the egg white of black swan has been determined at 1.9 A resolution using a semi-automatic procedure based on the Calpha coordinates of the homologous goose protein. PMID- 15299300 TI - Overexpression of Crithidia fasciculata trypanothione reductase and crystallization using a novel geometry. AB - TR1, a previously cloned gene for Crithidia fasciculata trypanothione reductase (TR), has been overexpressed in Escherichia coli strain SG5 to produce about 20 mg enzyme 1(-l) of culture. Since natural C. fasciculata TR is heterogeneous, this expression system provides an important source of homogeneous C. fasciculata TR for use in structural studies and drug design. Steady-state kinetic constants of the purified recombinant enzyme are K(m) = 56 micro M and k(cat) = 10 500 min( 1). Four crystal forms of TR1 were grown using this preparation. Synchrotron radiation was crucial to discover the high level of order present in crystal form IV, which diffracts to about 1.4 A resolution. To optimize growth and handling of form IV crystals, a novel crystallization setup called the 'plug drop' was developed. PMID- 15299301 TI - Direct phasing of one-wavelength anomalous-scattering data of the protein core streptavidin. AB - The direct method [Fan, Hao, Gu, Qian, Zheng & Ke (1990). Acta Cryst. A46, 935 939] was used to break the phase ambiguity intrinsic to one-wavelength anomalous scattering data from a known protein of moderate size, core streptavidin, which was solved originally with three-wavelength anomalous diffraction data [Hendrickson, Pahler, Smith, Satow, Merritt & Phizackerley (1989). Proc. Natl Acad. Sci. USA, 86, 2190-2194]. Unlike that in the previous test with a small protein, the Fourier map calculated with the direct-method phases could not clearly reveal the moderate-sized protein structure. However, the phases can be improved step by step using Wang's solvent-flattening method, non crystallographic symmetry averaging and the skeletonization method. The final electron-density map clearly shows most Calpha positions and some side chains and it is traceable without prior knowledge of the structure. It is concluded that the direct method is capable of breaking the OAS phase ambiguity of a moderate sized protein at moderate resolution such as 3 A, while the combination of direct methods with macromolecular techniques may produce phases good enough for unknown protein structure to be traced. PMID- 15299302 TI - Reciprocal-space molecular-replacement averaging. AB - The molecular-replacement equations, in which electron-density averaging and skew averaging have been unified, were used in reciprocal space to refine and extend the resolution of phased reflections. A procedure has been developed for the treatment of molecular envelopes of general shape. The equations were successfully applied to the reflection data of bacteriophage phiX174 (60-fold redundancy). Truncation of the G diffraction function beyond the first few nodes did not have a significant effect on the quality of the molecular-replacement equations. Reciprocal-space molecular-replacement averaging should prove to be a useful alternative to real-space averaging. Strategies are discussed that are possible only in reciprocal space. PMID- 15299303 TI - Refined crystal structure of lysozyme from the rainbow trout (Oncorhynchus mykiss). AB - Lysozymes (E.C. 3.2.1.17) are well characterized ubiquitous enzymes that have an antibacterial effect. The lysozymes from rainbow trout (RBTL) (Oncorhynchus mykiss) could be particularly interesting in aquaculture since they show higher activity than egg-white lysozyme and lysozymes from other fish species against a variety of pathogenic bacteria. Two lysozymes, I and II, differing only in a single amino acid, were purified from the kidney of rainbow trout and shown to belong to the c-type class of lysozymes. The type II form was shown to be much more potent against a variety of bacteria than the type I enzyme. We have grown crystals from a mixture containing about 80% type I and 20% type II lysozyme from rainbow trout, and solved the X-ray crystal structure. The crystals are trigonal with a = 76.68, c = 54.46 A and space group P3(1)21. The phase problem was solved by the molecular-replacement method, and the structure was refined to an R-factor of 17.4% using data to 1.8 A resolution. The crystal structure shows that the three-dimensional structure of rainbow trout lysozyme is very similar to the previously solved structures of other c-type lysozymes. The single polypeptide of 129 amino acids is folded into two domains separated by a deep cleft which contains the active site. Secondary-structure elements, four alpha-helices and a three-stranded beta-sheet, are located in the same sequential positions as in the hen, turkey and human enzymes. The beta-sheet is found to be common for structures of both c- and g-type lysozymes. We suggest that differences in antibiotic activity of the two forms of RBTL are probably due to small differences in the hydophobicity of a small surface region. PMID- 15299304 TI - Structure of the photochemical reaction centre of a spheroidene-containing purple bacterium, Rhodobacter sphaeroides Y, at 3 A resolution. AB - The crystal structure of the photochemical reaction centre from Rhodobacter sphaeroides Y, a carotenoid-containing wild-type purple bacterium, has been determined at 3 A resolution. This membrane complex consists of three subunits (281, 307 and 260 residues, respectively) and ten cofactors. It was crystallized in presence of beta-D-octylglucoside. The crystals are orthorhombic with unit cell dimensions, a = 143.7, b = 139.8, c = 78.7 A, space group P2(1)2(1)2(1) with four molecules in the unit cell. Refinement of the structure by X-PLOR and manual reconstructions yielded an R value of 22.1% for 19630 reflections between 7 and 3 A. The secondary structure is highly homologous to those determined for Rhodopseudomonas viridis (Protein Data Bank entry 1PRC) and Rhodobacter sphaeroides R26 (Protein Data Bank entry 4RCR) reaction centres. In the latter two structures one Fe(2+) ion located between the two quinones is coordinated by four histidines and one glutamic acid. In the Rhodobacter sphaeroides Y structure, Mn(2+) occupies the same position with identical ligands and geometry. The carotenoid conformation which is a non-planar 15-15'-cis spheroidene molecule in our structure differs from the 13-14-cis 2,4-dihydroneusporene in the Rhodopseudomonas viridis structure. PMID- 15299305 TI - Structure of a monoclonal anti-ICAM-1 antibody R6.5 Fab fragment at 2.8 A resolution. AB - The specific binding of the monoclonal murine anti-intercellular adhesion molecule-1 (anti-ICAM-1) antibody, R6.5, inhibits the attachment of neutrophils to endothelium and prevents the attachment of major group human rhinovirus (HRV) to ICAM-1. This binding interferes with the host immune system and, as a result, the R6.5 antibody has been developed as a therapeutic anti-inflammatory and perhaps anti-HRV agent. The variable-region amino-acid sequence of R6.5 was determined from the anti-ICAM-1 cDNA. The crystallization conditions of the Fab fragment of R6.5 were established and the three-dimensional structure was determined by X-ray crystallography. The crystal space group is orthorhombic P2(1)2(1)2(1), a = 40.36, b = 137.76, c = 91.32 A, and the highest resolution of recorded reflections is 2.7 A. The molecular-replacement method using known Fab structures was employed to solve the R6.5 Fab structure. The final R-factor is 18.8% for a total of 3320 non-H protein atoms, 39 water molecules and 10 606 unique reflections. The protein exhibits the typical immunoglobulin fold. The surface contour of the antigen-combining site of the R6.5 antibody has a wide groove which resembles more the structure of an anti-polypeptide antibody than the structure of an anti-protein antibody. PMID- 15299306 TI - Crystallization of HMG-CoA reductase from Pseudomonas mevalonii. AB - Crystals of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase from Pseudomonas mevalonii have been grown by vapor diffusion in hanging drops at pH 6.7 using ammonium sulfate as the precipitant. Serial dilution seeding and manipulation of glycerol concentration were both used to obtain crystals larger than 1.0 mm. The crystals are cubic, space group I4(1)32, with a = 229.4 A. A V(m) value of 2.71 A(3) Da(-l) indicates 96 molecules per unit cell with two molecules in the asymmetric unit. These crystals diffract to 2.8 A with conventional X-ray sources, and beyond 2.4 A with synchrotron radiation. PMID- 15299307 TI - Characterization of lysozyme crystals with unusually low solvent content. AB - Studies on the low-humidity (88%) forms of tetragonal and monoclinic lysozyme, resulting from water-mediated transformations, have provided a wealth of information on the variability in protein hydration, its structural consequences and the water structure associated with proteins, in addition to facilitating the delineation of the rigid and the flexible regions in the protein molecule and the invariant features in its hydration shell. Surprisingly, monoclinic lysozyme continues to diffract even when the environmental humidity is drastically reduced, thus permitting the structural study of the enzyme at different levels of hydration. As part of a study in this direction, three very low humidity forms, two of them occurring at a nominal relative humidity of 38% and the other at 5% relative humidity, have been characterized. These have unprecedented low solvent contents of 16.9, 17.6 and 9.4%, respectively, as determined by the Matthews method. PMID- 15299308 TI - Crystallization and preliminary X-ray crystallographic studies of native elastase from North Atlantic salmon (Salmo salar). AB - Crystals of elastase from North Atlantic salmon have been grown from 2-methyl-2,4 pentanediol by the hanging-drop vapour-diffusion method at room temperature. They grow to dimensions of 0.7 x 0.4 x 0.3 mm in three weeks. The crystals belong to the tetragonal space group P4(1)2(1)2 or P4(3)2(1)2 with cell dimensions a = b = 68.0 A and c = 84.0 A. There are eight molecules in the unit cell. The crystals diffract to at least 1.6 A resolution and are suitable for a high-resolution crystal structure determination. PMID- 15299309 TI - Preliminary crystallographic analysis of an extremely thermostable glutamate dehydrogenase from the archaeon Pyrococcus woesei. AB - The extremely heat-stable glutamate dehydrogenase (GluDH) from Pyrococcus woesei was crystallized by the hanging-drop vapour-diffusion method. Crystals suitable for X-ray crystallographic investigations were obtained using polyethylene glycol (PEG) 4000 and ammonium acetate as precipitating agents. The crystals obtained diffract to a resolution of 2.8 A, have a prismatic shape and grow up to 0.5 mm in their maximal dimension. They belong to the triclinic system (space group P1; a = 90.9, b = 92.8, c = 107.1 A, alpha = 69.1, beta = 80.7, Vgamma = 65.0 degrees ) with a unit-cell volume of 765052 A(3) which accommodates one GluDH hexamer of 276 kDa. The averaged density of the crystal determined by Ficoll-gradient centrifugation is 1.15 g cm(-3), which corresponds to a molecular mass of 255 kDa in the unit cell. A native data set has been collected on an MAR image-plate system using Cu Kalpha radiation. The completeness of the data set in the range 316-3 A is 74%, and contains 64% of the data in the outer shell (3.4-2.8 A), with an average R(merge) value of 9%. Calculation of self-rotation functions revealed the 32 symmetry of the hexamer; 3 non-crystallographic twofold axes were found at a distance of 120 degrees in a plane perpendicular to the non-crystallographic threefold axis. PMID- 15299310 TI - New crystal forms of Escherichia coli and Saccharomyces cerevisiae soluble inorganic pyrophosphatases. AB - We have obtained new crystal forms of Escherichia coli and Saccharomyces cerevisiae soluble inorganic pyrophosphatase with and without substrate, competitive inhibitor and divalent cation. They diffract to higher resolution than any forms previously reported. The best E. coli crystals are in space group R32 with cell dimensions of 111.4 x 111.4 x 76.6 A and diffract to 2.0 A. The best S. cerevisiae crystals were grown from a mixture of PEG 1000 and 4000 in the presence of metal ions. They are in space group P2(1)2(1)2(1), have cell dimensions of 54.2 x 68.5 x 161.7 A and diffract to 1.8 A. PMID- 15299311 TI - Crystallization of a calcium-binding EGF-like domain. AB - Crystals of a calcium-binding epidermal growth factor (EGF)-like domain of human clotting factor IX suitable for X-ray diffraction analysis have been obtained by vapour diffusion (sitting drop) against 48% PEG 400. The crystals belong to the tetragonal space group P4(3)2(1)2, with unit-cell dimensions a = b = 40.3, c = 98.2 A. The crystals diffract beyond 1.5 A resolution and are relatively stable in the X-ray beam. This is the first reported crystallization of a calcium binding EGF-like domain. PMID- 15299312 TI - Coordinate-based cluster analysis. AB - A new approach to cluster analysis of structures based on collective superpositions rather than pairwise superpositions is presented. The method is fast and rigorous and is illustrated by application to 21 structures derived from NMR experiments. Source code, suitable for most laboratory machines, is available from the author, and a CCP4 version is in preparation. PMID- 15299313 TI - Do C-H...O hydrogen bonds contribute to the stability of nucleic acid base pairs? AB - The possible formation of inter-base C-H.O hydrogen bonds in A.T, A.U and certain non-Watson-Crick base pairs is examined. A geometrical analysis in conjunction with implications for the thermodynamic stability of the base pairs suggests that C-H.O hydrogen bonds could form in nucleic acid base pairs. They may alleviate destabilizing interactions that would arise if an unsatisfied hydrogen-bond acceptor were present and mediate secondary hydrogen-bonding effects in these base pairs. PMID- 15299314 TI - A probe molecule composed of seventeen percent of total diffracting matter gives correct solutions in molecular replacement. AB - It is often found in the crystallization of enzyme-inhibitor complexes that an inhibitor causes crystal packing which is different to that of native protein. This is the case for crystals of human non-pancreatic secreted phospholipase A(2) (124 residues) containing six molecules in the asymmetric unit when the protein is complexed with a potential acylamino analogue of a phospholid. The hexameric structure was determined by molecular replacement using the structure of monomeric native protein as a probe. As an extension to the experiment, it was tested whether a backbone polypeptide composed of 17% of a known monomeric structure could find its correct position on a target molecule in molecular replacement. A probe model composed of the backbone atoms of the N-terminal 77 residues of lysine-, arginine-, ornithine-binding protein (LAO, a total of 238 residues) liganded with lysine correctly finds its position on LAO liganded with histidine which crystallizes as a monomer in the asymmetric unit. The results indicate that as little as 17% of total diffracting matter can be used in molecular replacement to solve crystal structures or to obtain phase information which can be combined with phases obtained by the isomorphous-replacement method. PMID- 15299315 TI - The three-dimensional structure of the aspartate receptor from Escherichia coli. AB - The crystal structure of the periplasmic domain of the aspartate receptor from Escherichia coli has been solved and refined to an R-factor of 0.203 at 2.3 A, resolution. The dimeric protein is largely helical, with four helices from each monomer forming a four-helix bundle. The dimer interface is constructed from four helices, two from each subunit, also packed together in a four-helix bundle arrangement. A sulfate ion occupies the aspartate-binding site. All hydrogen bonds made to aspartate are substituted by direct or water-mediated hydrogen bonds to the sulfate. Comparison of the Escherichia coli aspartate-receptor structure with that of Salmonella typhimurium [Milburn, Prive, Milligan, Scott, Yeh, Jancarik, Koshland & Kim (1991). Science, 254, 1342-1347; Scott, Milligan, Milburn, Prive, Yeh, Koshland & Kim (1993). J. Mol. Biol. 232, 555-573] reveals strong conservation in the structure of the monomer, but more divergence in the orientation of the subunits with respect to one another. Mutations that render the Escherichia coli receptor incapable of responding to maltose are either located in spatially conserved sites or in regions of the structures that have high temperature factors and are therefore likely to be quite flexible. The inability of the receptor from Salmonella typhimurium to respond to maltose may, therefore, be because of differences in amino acids located on the binding surface rather than structural differences. PMID- 15299316 TI - Structure of the ferredoxin from Clostridium acidurici: model at 1.8 A resolution. AB - Ferredoxins (Fd) are electron-carrier proteins, the active sites of which are organized around clusters made of iron and inorganic sulfur. The Fd from Clostridium acidurici is 55 amino acids long and contains two [4Fe-4S] clusters. Crystals have been obtained in the space group P4(3)2(1)2, a = b = 34.441 (5), c = 74.778 (9) A. The structure was solved by molecular replacement using the Fd from Peptostreptcoccus asaccharolyticus as a search model, these two ferredoxins having 37 residues in common. Refinement using molecular-dynamics techniques was then initiated. Successive rounds of model building and refinement gave a structure that includes 45 water molecules with R = 15%. At this stage, the electron-density map clearly revealed discrepancies in the position of two amino acids in the published primary sequence. Refinement based on these modifications led to R = 14.3% for 3921 reflections up to 1.8 A, resolution. The geometry of the two clusters has been found to be in good agreement with that previously obtained at a lower resolution. Interactions of polypeptide chain with the [4Fe 4S] clusters, the cluster geometry as well as the hydrogen bonds involving S, Sgamma, N and water molecules are reported. PMID- 15299317 TI - Deconvolution of fully overlapped reflections from crystals of foot-and-mouth disease virus O1 G67. AB - Foot-and-mouth disease virus O(1) G67 forms crystals that appear similar to those of the closely related viruses O(1)K and O(1)BFS, both of which belong to space group I23. Statistical disorder in the O(1) G67 crystals means, however, that the measured diffraction data possess higher symmetry consistent with point group 432. It is shown that this is due to intimate twinning, with mosaic blocks randomly distributed between the two orientations. This results in a twofold loss of information due to the exact superimposition of non-identical reflections from the two orientations. A simple procedure has been devised to deconvolute these overlapped reflections by applying constraints in both real and reciprocal space. This procedure works well, providing interpretable electron-density maps for this virus. Other applications are discussed. PMID- 15299318 TI - Structure of the azurin mutant Phe114Ala from Pseudomonas aeruginosa at 2.6 A resolution. AB - The crystal structure of azurin mutant Phe114Ala from Pseudomonas aeruginosa has been solved by molecular replacement. The final crystallographic R value is 0.185 for 9832 reflections to a resolution of 2.6 A. The root-mean-square deviation for main-chain atom positions is 0.020 A between the four independent monomers in the asymmetric unit. The mutant Ala114 crystallized from PEG 4000 in a new crystal form and the crystals are monoclinic, P2(1), a= 51.0, b = 83.6, c= 66.4 A and beta = 110.5 degrees. The four molecules in the asymmetric unit are packed as a dimer of dimers and are related by an approximate twofold axis. The dimer packing and the dimer contact region are very similar to that of the Alcaligenes denitrificans azurin dimer. The mutation was performed at residue Phe114, which exhibits a pi-electron overlap with the copper ligand His117, to investigate its suggested role in the electron self-exchange reaction. Removal of steric constrains from the phenylalanine side chain created a somewhat different geometry around the copper site with an increased mobility of His117 resulting in an enlarged Cu-N length which may be responsible for the slight differences obtained in the spectral properties of the mutant versus the wild-type protein. PMID- 15299319 TI - Crystal structure of narbonin at 1.8 A resolution. AB - The three-dimensional structure of narbonin, a seed protein from Vicia narbonensis L, has been determined at 1.8 A resolution. Phase information was obtained by multiple isomorphous replacement and optimized anomalous dispersion. The narbonin structure was initially traced with only 17% amino-acid sequence information and preliminarily refined to a crystallographic R-factor of 16.5%. It is now refined to 15.9% using full sequence information derived from cDNA and after the addition of more solvent molecules. The monomeric molecule of narbonin is an eight-stranded parallel beta-barrel surrounded by alpha-helices in a beta/alpha-topology similar to that first observed in triose phosphate isomerase. Differences exist in the N-terminal part of the polypeptide chain, where the first helix is replaced by a loop and the second beta-strand is followed by an additional antiparallel alpha-sheet placed parallel on top of alpha-helices alpha3 and alpha4. Two short additional secondary structures are present. The first, an alpha-helix, is situated between the seventh beta-strand and the following helix, and the second, which is a 3(10) helix, between the eighth strand and the C-terminal helix. The most striking observation is the lack of a known enzymatic function for narbonin, because all TIM-like structures known so far are enzymes. PMID- 15299320 TI - X-ray structures of the B-DNA dodecamer d(CGCGTTAACGCG) with an inverted central tetranucleotide and its netropsin complex. AB - The crystal structures of the B-DNA dodecamer d(CGCGTTAACGCG) duplex (T2A2), with the inverted tetranucleotide core from the duplex d(CGCGAATTCGCG) [A2T2, Dickerson & Drew (1981). J. Mol. Biol. 149, 761-768], and its netropsin complex (T2A2-N) have been determined at 2.3 A resolution. The crystals are orthorhombic, space group P2(1)2(1)2(1), unit-cell dimensions of a = 25.7, b = 40.5 and c = 67.0 A, for T2A2 and a = 25.49, b = 40.87, c = 67.02 A for T2A2-N and are isomorphous with A2T2. The native T2A2 structure, with 70 water molecules had a final R value of 0.15 for 1522 reflections (F > 2sigma), while for the netropsin complex, with 87 water molecules, the R value was 0.16 for 2420 reflections. In T2A2, a discontinuous string of zig-zagging water molecules hydrate the narrow A.T minor groove. In T2A2-N, netropsin binds in one orientation in the minor groove, covering the TTAA central region, by displacing the string of waters, forming the majority of hydrogen bonds with DNA atoms in one strand, and causing very little perturbation of the native structure. The helical twist angle in T2A2 is largest at the duplex center, corresponding to the cleavage site by the restriction enzymes HpaI and HincII. The sequence inversion AATT-->TTAA of the tetranucleotide at the center of the molecule results in a different path for the local helix axis in T2A2 and A2T2 but the overall bending is similar in both cases. PMID- 15299321 TI - Structure of a new alkaline serine protease (M-protease) from Bacillus sp. KSM K16. AB - An alkaline serine protease, M-protease, from Bacillus sp. KSM-K16 has been crystallized. Two morphologically different crystal forms were obtained. Crystal data of form 1: space group P2(1)2(1)2(1), a = 47.3, b = 62.5, c = 75.6 A, V = 2.23 x 10(5) A(3), Z = 4 and V(m) = 2.09 A(3) Da(-1). Crystal data of form 2: space group P2(1)2(1)2(1), a = 75.82 (2), b = 57.79 (2), c = 54.19 (1) A, V = 2.29 (2) x 10(5) A(3), Z = 4 and V(m) = 2.15 A(3) Da(-1). The crystal structure of M-protease in form 2 has been solved by molecular replacement using the atomic model of subtilisin Carlsberg (SBC) which is 60% homologous with M-protease, and refined to the crystallographic R-factor of 0.189 for 7004 reflections with F(o)/sigma(F) > 3 between 7 and 2.4 A resolution. The final model of M-protease contains 1882 protein atoms, two calcium ions and 44 water molecules. The three dimensional structure of M-protease is essentially similar to other subtilisins of known structure. The 269 C(alpha) positions of M-protease have an r.m.s. difference of 1.06 A with the corresponding positions of SBC. The crystal data of form 2 are close to those of SBC, though the structure determination of form 2 made it clear that it is not isomorphous to the crystal structure of SBC. The deletions of amino acids occur at the residues 36' and 160'-163' compared with SBC (numerals with primes show the numbering for SBC). The deletion of the four residues (160'-163') may significantly affect the lack of isomorphism between M protease and SBC. PMID- 15299322 TI - Growth process of protein crystals revealed by laser Michelson interferometry investigation. AB - The elementary processes of protein crystal growth were investigated by means of laser Michelson interferometry on the example of the (101) face of tetragonal hen egg-white (HEW) lysozyme. The method allows real-time in situ observations of the morphology of the growing protein crystal surface, as well as simultaneous precise measurement of growth rate and step velocity on identified growth-layer sources. At the critical supersaturation of 1.6 the growth mechanism was shown to transform from dislocation-layer generation to surface nucleation. Measurements on different growth hillocks, with material of a different source and at a different temperature, all indicated that for supersaturations lower than approximately 1 growth is hindered by the competitive adsorption of (most probably) other protein species contained in HEW, although the material is pure by most analytical methods. At supersaturations sigma < 0.4 other impurities sometimes led to cessation of growth. However, at sigma in the range 0.9 < sigma < 1.6 growth processes are determined by the kinetics of pure lysozyme. This enabled us to measure the step kinetic coefficient beta for crystallization of a protein substance for the first time: beta = 2.8 micro m s(-1). This also means that by working in this supersaturation range we can eliminate the impurity effects. Other means to reduce influence of impurities is to use, if possible, a higher crystallization temperature. It is shown that slow crystallization of proteins is due primarily to impedance of the elementary act of entering the growth site and not to the low concentration of the solution. The value of beta does not depend on temperature, indicating the decisive role of entropy, not energy barriers, in the crystallization of biological macromolecules. PMID- 15299323 TI - Crystallographic analysis of Phe-->Leu substitution in the hydrophobic core of barnase. AB - The crystal structure of a barnase mutant, Phe-->Leu7 has been determined to 2.2 A resolution. No structural rearrangement is observed near the mutated residue. The F7L mutation is highly destabilizing and this is caused by the loss of extensive van der Waals contacts that wild-type Phe7 made with its neighbouring residues, and the exposure of a large hydrophobic pocket on the surface of the protein. The side-chain conformations of the mutated Leu7 residue have torsion angles chi(1) ranging from -138 degrees to -168 degrees and chi(2) ranging from +16 degrees to +70 degrees, for the three molecules in the asymmetric unit. These angles do not agree with the most frequently observed conformations in the protein side-chain rotamer library [Ponder & Richards (1987). J. Mol. Biol. 193, 775-791]. However, when compared to a more recent 'backbone-dependent' rotamer library [Dunbrack & Karplus (1993). J. Mol. Biol. 230, 543-574], the side-chain conformation of Leu7 agrees well with that of the most frequently observed rotamers. The side-chain conformation of Leu7 was found to be dictated by two factors: it has the lowest conformational energy and it buries the most hydrophobic surface area. PMID- 15299324 TI - Cytochrome c6 from the green alga Monoraphidium braunii. Crystallization and preminary diffraction studies. AB - Cytochrome c(6), a plastocyanin functionally interchangeable electron carrier between the chlorophyll molecule P700 of photosystem I and cytochrome f from cytochrome b(6)f complex, has been isolated from the green alga Monoraphidium braunii and crystallized by the vapour-diffusion technique in sodium citrate. Crystals belong to space group R3, with cell dimensions a = b = 51.93 (5) and c = 80.5 (1) A (hexagonal axes), with one molecule per asymmetric unit. They diffract beyond 1.9 A under a Cu Kalpha rotating-anode source, with an anomalous signal that allows the positioning of the heme Fe atom in the unit cell. PMID- 15299325 TI - A new crystal form of bovine heart ubiquinol: cytochrome c oxidoreductase: determination of space group and unit-cell parameters. AB - Ubiquinol:cytochrome c oxidoreductase, the middle segment of the mitochondrial respiratory chain, is a multi-subunit transmembrane redox enzyme. The purified protein from beef heart mitochondria has been crystallized by three groups in three different forms, but progress toward a structure has been hampered by the limited order (resolution) of the crystals. It has been found that under certain conditions the enzyme crystallizes in a new form suitable for X-ray diffraction studies. These crystals belong to space group C222(1) in the orthorhombic system. The cell dimensions are a = 384, b = 118 and c = 177 A. These new crystals at present diffract to 3.8 A at best. This is not significantly better than hexagonal [P6(1(5))22] crystals grown, but the new crystals have the advantage of less spot overlap because of face-centered packing which results in systematic extinctions. More importantly, the availability of the same enzyme in multiple crystal forms may allow phase refinement and extension by the method of molecular replacement. PMID- 15299326 TI - Crystallization of the NAD(P)-dependent glutamate dehydrogenase from the hyperthermophile Pyrococcus furiosus. AB - The NAD(P)-dependent glutamate dehydrogenase from Pyrococcus furiosus has been crystallized by the hanging-drop method of vapour diffusion using lithium sulfate as the precipitant. The crystals belong to the tetragonal system and are in space group P4(2)2(1)2 with unit-cell dimensions of a = b = 167.2, c = 172.9 A. Consideration of the values of V(m) and possible packing of the molecules within the cell suggest that the asymmetric unit contains a trimer. P. furiosus belongs to the family of Archaea and is one of the most thermostable organisms known, having an optimal growth temperature of 376 K. The glutamate dehydrogenase isolated from this organism has a half-life of 12 h at 373 K and, therefore, the determination of the structure of this enzyme will be important in advancing our understanding of how proteins are adapted to enable them to survive at such extreme temperatures. PMID- 15299327 TI - Crystallization and preliminary X-ray structure solution of Rhizomucor miehei aspartic proteinase. AB - Rhizomucor miehei aspartic proteinase (M(r) = 38701, 361 residues) has been crystallized in a form suitable for analysis by X-ray diffraction. The flattened rod-shaped crystals were grown from polyethylene glycol 8000 using vapour diffusion methods. The crystal form is in space group P2(1)2()12(1) [a = 41.67 (2), b = 51.21 (3) and c = 173.3 (2) A], with Z = 4 and one molecule in the asymmetric unit. Data were collected over the range 0 < h < 14, 0 < k < 16 and 0 < l < 62, resulting in 7032 unique reflections to give 72.1% completeness with a merging R of 0.067 to a resolution limit of 2.8 A. A molecular-replacement solution of the structure has been obtained using the aspartic proteinase from Rhizomucor pusillus as a model. Rigid-body refinement of the model and subsequent refinement using molecular dynamics were performed with X-PLOR, leading to a current R-factor of 20.1% for 2.8 A data. PMID- 15299328 TI - Crystallization and preliminary X-ray diffraction studies of Escherichia coli glyceraldehyde-3-phosphate dehydrogenase. AB - Phosphorylating glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a key enzyme in glycolysis. Single crystals of NAD-dependent GAPDH from Escherichia coli have been obtained by vapour diffusion at room temperature using trisodium citrate as precipitant. In almost the same crystallization conditions, two kinds of crystals were found to be suitable for X-ray diffraction. The crystals with only one half of a tetramer in the asymmetric unit were chosen for high- resolution analysis. They belonged to space group C222(1), with cell dimensions a = 79.1, b = 189.6 and c = 122.2 A. These crystals diffracted to 1.8 A resolution. PMID- 15299330 TI - Cryocrystallography of influenza virus hemagglutinin crystals. AB - X-ray diffraction data collected at cryogenic temperatures from flash-cooled crystals of influenza virus hemagglutinin show improvements in both resolution and quality relative to data collected at 277 K. These improvements are dramatic for flash-cooled hemagglutinin crystals irradiated with X-rays from a synchrotron source. At the Cornell High Energy Synchrotron Source flash-cooled hemagglutinin crystals diffracted at least 0.9 A farther than hemagglutinin crystals at ambient temperatures. Radiation damage in the flash-cooled crystals is reduced, making it possible to collect a complete data set from a single hemagglutinin crystal. However, radiation damage is not eliminated in the flash-cooled crystal. As a result the quality of X-ray data can be significantly degraded during long exposure times at a synchrotron source. PMID- 15299331 TI - A method for processing diffraction data from twinned protein crystals and its application in the structure determination of an FAD/NADH-binding fragment of nitrate reductase. AB - A general method to deconvolute oscillation data sets from twinned protein crystals to a corresponding single-crystal data set has been developed and applied to diffraction data measured from crystals of a fragment containing the FAD- and NADH-binding domains of nitrate reductase. The procedure allows straightforward processing of diffraction data from twinned crystals. Typically, R(merge) values of reduced data sets from the nitrate reductase crystals after deconvolution are about 0.06 compared to 0.13 and higher before deconvolution. Based on these deconvoluted data sets, the structure of the FAD- and NADH-binding domains of nitrate reductase could be solved successfully. The result indicates that crystal twinning does not necessarily prevent crystallographic structure determination. PMID- 15299332 TI - Modeling protein-substrate interactions in the heme domain of cytochrome P450(BM 3). AB - The crystal structure of heme domain of the fatty acid monooxygenase, cytochrome P450(BM-3), consisting of residues 1-455 has been independently solved to R = 0.18 at 2.0 A. The crystal form used, space group P2(1) with two molecules per asymmetric unit, is isomorphous with that form with residues 1-471 first described by Boddupalli et al. [Boddupalli, Hasemann, Ravinchandran, Lu, Goldsmith, Deisenhofer & Peterson (1992). Proc. Natl Acad. Sci. USA, 89, 5567 5571] and used by Ravichandran, Boddupalli, Hasemann, Peterson & Deisenhofer [(1993). Science, 261, 731-736] to determine the crystal structure. The substrate access channel consists of a large, hydrophobic cleft that appears to be the most likely route taken by fatty acid substrates. Attempts to soak crystals in mother liquor containing a variety of fatty acid substrates yielded featureless difference Fouriers even though fatty acid substrates are known to bind with dissociation constants in the micro M range. Modeling substrate-enzyme interactions reveals few contacts between the enzyme and substrate. More detailed modeling was carried out by subjecting both molecules in the asymmetric unit to extensive energy minimization. These studies reveal that the heme-domain active site cleft can undergo a large conformational change that closes the access channel thereby providing enhanced protein-substrate interactions. These conformational changes are prevented from occurring by intermolecular contacts in the crystal lattice which lock the protein in the 'open' conformation. PMID- 15299333 TI - Crambin: a direct solution for a 400-atom structure. AB - The crystal structure of crambin, a 46-residue protein containing the equivalent of approximately 400 fully occupied non-H-atom positions, was originally solved at 1.5 A by exploiting the anomalous scattering of its six S atoms at a single wavelength far removed from the absorption edge of sulfur. The crambin structure has now been resolved without the use of any anomalous-dispersion measurements. The technique employed was an ab initio 'shake-and-bake' method, consisting of a phase-refinement procedure based on the minimal function alternated with Fourier refinement. This method has successfully yielded solutions for a smaller molecule (28 atoms) using 1.2 A data, and a crambin solution was obtained at 1.1 A. PMID- 15299334 TI - Structure determination of OppA at 2.3 A resolution using multiple-wavelength anomalous dispersion methods. AB - OppA is a 58.8 kDa bacterial transport protein involved in the transport of peptides across the cytoplasmic membrane of Gram-negative bacteria. It binds peptides from two to five residues in length but with little sequence specificity. OppA from Salmonella typhimurium has been cloned and expressed in E. coli and the protein cocrystallized with uranyl acetate, producing two distinct crystal forms with different uranium sites. Multiple-wavelength data collected about the uranium L(III) edge have been collected at the Daresbury Synchrotron Radiation Source (SRS) to a nominal resolution limit of 2.3 A. Maximum-likelihood phasing methods have been used in phase determination from the multiple wavelength data giving a readily interpretable electron-density map, without any density modification. The electron-density map, calculated at 2.3 A resolution shows OppA to be a bilobal, principally beta-stranded, three-domain protein. The tri-lysine ligand molecule can be clearly seen in the peptide-binding site between the two lobes. PMID- 15299335 TI - Structure of octreotide, a somatostatin analogue. AB - Octreotide, a synthetic somatostatin analogue, is an octapeptide with one disulfide bridge. Crystals of octreotide are orthorhombic, space group P2(1)2(1)2(1), a = 18.458 (5), b = 30.009 (7), c = 39.705 (27) A, with three molecules of octapeptide, one ordered oxalate dianion and 52 water molecules in the asymmetric unit. Complete protonation of the NH(2) groups (as assumed in the refinement) would require three oxalate dianions in the asymmetric unit for charge neutrality; a chemical analysis indicated that four are present. In either case they are so disordered that they cannot be distinguished from the water molecules. The 18 951 unique reflections (R(sym) = 0.026) used for structure solution and refinement were recorded with the EMBL imaging-plate scanner using synchrotron radiation. The structure was solved by Patterson interpretation, locating the three disulfide bridges, followed by tangent phase expansion and E Fourier recycling. The anisotropic refinement against all F(2) data between 1.04 and 10.0 A resolution by blocked restrained full-matrix least-squares techniques converged to a conventional R index based on F of 0.084 [I > 2a(I) and 10.0 > d > 1.04 A] and wR2, the weighted R-index on F(2), of 0.246 (for all data). One peptide molecule adopts a flat beta-sheet structure; the other two possess different irregular backbone conformations, but are similar to each other. All three molecules have a distorted type II' beta-turn around the D-Trp-Lys region, but exhibit different side-chain conformations. The crystal structure is stabilized by a network of inter- and intramolecular hydrogen bonds. PMID- 15299336 TI - Comparison of different X-ray data-collection systems using the crystal structure of octreotide. AB - The octapeptide octreotide crystallizes with three peptide molecules and about 20% water in the asymmetric unit, and in many ways possesses diffraction properties similar to those of a 'mini-protein' consisting of 24 amino-acid residues. It diffracts to about 1.0 A but data in the range 1.4-1.0 A are weak. It provides a suitable test of different macromolecular X-ray data-collection techniques, especially of their ability to measure weak reflections accurately. In contrast to typical proteins it is possible to perform a full anisotropic refinement, that we believe provides a more objective test of the quality of the data than the internal consistency of equivalent reflections. We have collected a total of six data sets. The X-ray sources included synchrotron radiation, Cu Kalpha rotating anodes and Mo Kalpha sealed tubes; position-sensitive two dimensional detectors from four manufacturers and a four-circle diffractometer with scintillation counter were employed. Two of the six data sets were collected at low temperature. Reasonable anisotropic refinement was possible with all area detector data sets, although significant differences in the precision of the final model were observed. In addition we tested the ability of automated Patterson interpretation to solve the structure using the six independent data sets. The structure solution was only successful using the synchrotron or rotating-anode data sets, i.e. for the more intense sources. It appears that for structure solution the maximum resolution of the data is critical, whereas for refinement the accuracy of the data is more important. PMID- 15299337 TI - Well ordered crystals of a short-chain alcohol dehydrogenase from Drosophila lebanonensis: re-evaluation of the crystallographic data and rotation-function analysis. AB - Alcohol dehydrogenase prepared from Drosophila lebanonensis yields well ordered plate-like crystals which diffract to better than 2.3 A resolution. The crystals belong to space group P2(1) of the monoclinic system; the unit-cell dimensions are a = 65.25, b = 55.77, c = 70.02 A, alpha = 90, beta = 107.08, gamma = 90 degrees. The asymmetric unit of the crystal cell is most probably occupied by a dimer, corresponding to a packing density of 2.15 A(3) Da-L. The orientation of the non-crystallographic twofold symmetry axes is determined by analysis of a self-rotation function calculated with native intensity data. PMID- 15299338 TI - Structure of inhibited trypsin from Fusarium oxysporum at 1.55 A. AB - The structure of trypsin from the fungus Fusarium oxysporum has been refined at 1.55 A resolution by restrained least-squares minimization to an R-factor of 14.4%. The data were recorded from a single-crystal on the X31 beamline at EMBL, Hamburg, using a locally developed image-plate scanner. The final model consists of 1557 protein atoms, 400 water molecules, one molecule of isopropanol and one monoisopropyl phosphoryl inhibitor group covalently bound to the catalytic Ser195. Comparison of the structure with bovine trypsin reveals significant differences in the active site and suggests a possible explanation for the difference in substrate specificity between the two enzymes. In F. oxysporum trypsin the specificity pocket is larger than in bovine trypsin. This explains the preference of F. oxysporum trypsin for the bulkier arginine over lysine and the reverse preference in bovine trypsin. The binding cavity on the C-terminal side of the substrate is more restricted in F. oxysporum trypsin than in mammalian and Streptomyces griseus trypsins, which explains the relative inactivity of F. oxysporum trypsin towards peptide-pNA substrate analogues as an unfavourable steric interaction between the side of the binding cavity and the para-nitroanilino group of peptide-pNA. The observed restriction of the binding cavity does not lead to a reduced catalytic activity compared to other trypsins. PMID- 15299339 TI - Time-averaging crystallographic refinement: possibilities and limitations using alpha-cyclodextrin as a test system. AB - The method of time-averaging crystallographic refinement is assessed using a small molecule, alpha-cyclodextrin, as a test system. A total of 16 refinements are performed on simulated data. Three resolution ranges of the data are used, the memory relaxation time of the averaging is varied, and several overall temperature factors are used. The most critical factor in the reliable application of time-averaging is the resolution of the data. The ratio of data to molecular degrees of freedom should be large enough to avoid overfitting of the data by the time-averaging procedure. The use of a free R-factor can aid in determining whether time-averaging can be reliably applied. Good ensembles of structures are obtained using data up to 1.0 or 2.0 A resolution. Comparison of electron-density maps from time-averaging refinement and anisotropic temperature factor refinement indicates that the former technique yields a better representation of the exact data than the latter. PMID- 15299340 TI - Water molecules which apparently accept no hydrogen bonds are systematically involved in C-H...O interactions. AB - The hydrogen-bond acceptor functions are analyzed for 672 water molecules in crystal structures of hydrated small biological molecules. Of the 672 water molecules, 26 neither accept hydrogen bonds from O-H or N-H donors, nor coordinate to metal ions. As compensation, they satisfy their acceptor potential with up to four C-H.O hydrogen bonds. They are found in hydrated amino acids, purines and pyrimidines, and alkaloids, but not in carbohydrates, nucleosides, nucleotides and steroids. As for conventional hydrogen bonds, the preferred coordination geometry is tetrahedral. No example of a true non-accepting water molecule is found. A particularly strong C-H hydrogen-bond donor occurring in many biological molecules is C-H neighbouring protonated N. PMID- 15299341 TI - The influence of temperature on lysozyme crystals. Structure and dynamics of protein and water. AB - Lysozyme structures at six different temperatures in the range 95-295 K have been determined using X-ray crystallography at a resolution of 1.7 A. The crystals at lower temperatures had a 7.4% decrease in the unit-cell volume. The volume change was discontinuous with the volume being near 238 000 A(3) from 295 to 250 K and about 220 200 A(3) below 180 K. The thermal expansion of the protein has been analyzed and shows anisotropy, which is correlated with local atomic packing and secondary-structure elements. The lysozyme structure at low temperature is nearly the same as that at high temperature, with only small relative translations and rotations of structure elements including a hinge-bending rearrangement of two domains. Because of a considerable increase of lattice disorder at low temperature dynamical analysis of internal motion is difficult. The analysis of structural and dynamical properties of well ordered protein-bound water has been carried out. PMID- 15299342 TI - Correlation function method in protein crystallography. AB - A weighted correlation function as a method for computing electron-density maps is proposed to reduce the errors of the Fourier syntheses performed on inaccurate and/or incomplete data. The formulae are revised for the difference Patterson vector search, for multiple isomorphous replacement (MIR) and single isomorphous replacement (SIR) syntheses and for the difference Fourier synthesis. The examples show that the correlation-function approach has the potential to provide more reliable results than those obtained by conventional Fourier syntheses. PMID- 15299343 TI - Crystallization and preliminary analysis of two crystal forms of human clara cell 16 kDa protein (CC10). AB - The human Clara cell 16 kDa protein (CC10), isolated from lung lavage fluid, has been crystallized in two crystal forms. The first is in space group P1 and has cell parameters a = 43.04, b = 45.90, c = 51.29 A and alpha = 62.46, beta = 69.74, gamma = 69.43 degrees. Two molecules are present in the unit cell. The second form is in space group P222, with cell parameters a = 42.24, b = 84.06, c = 40.05 A and alpha = beta = gamma = 90 degrees, and four molecules per unit cell. Its diffraction pattern displays pseudo-body-centered symmetry. Both crystal forms diffract X-rays beyond 2.0 A. PMID- 15299344 TI - Crystallization and preliminary X-ray studies of recombinant horseradish peroxidase. AB - A non-glycosylated form of horseradish peroxidase c extracted from Escherichia coli inclusion bodies and refolded in the presence of haem and Ca(2+) ions has been used to grow protein crystals suitable for X-ray diffraction analysis. The crystals are prisms in the trigonal space group P3(1)12 or P3(2)12 with a = b = 158.9 and c = 114.3 A, and diffract to 1.9 A. There are four molecules, each of 34 kDa, in the asymmetric unit. The molecules of the asymmetric unit are related by approximate translational symmetry, resulting in pseudo-centerings. Data to approximately 15 A can thus be described by a lattice of a' = b' = 91.7 A and c' = 57.1 A, alpha = beta = 90 degrees and gamma = 120 degrees, including four molecules. PMID- 15299345 TI - Preliminary crystallographic data for an Fab to the melanoma-associated GD2 ganglioside, and the purification of a soluble form of this antigen. AB - An Fab fragment from a monoclonal antibody (ME36.1) to the melanoma-associated GD2 ganglioside has been purified and crystallized in space group P2(1) with unit cell dimensions a = 37.6, b = 94.1, c = 67.4 A, beta = 101.0 degrees. The crystals, which grow to a size of up to 0.6 x 0.5 x 0.3 mm, diffract to 2.5 A and native data have been collected to 2.8 A resolution. The crystal density is 1.22 g ml(-1) indicating one molecule of 48 kDa per asymmetric unit and a solvent content of 51%. A soluble form of the carbohydrate was obtained from the scarce GD2 glycolipid by enzymatic digestion with ceramide-glycanase. Small co-crystals of the Fab-GD2 complex have been obtained. As ME36.1 has been used in immunotherapy to treat malignant melanoma, knowledge of its interactions with the ganglioside could increase the efficacy of these treatments. PMID- 15299346 TI - Refined crystal structure of liver alcohol dehydrogenase-NADH complex at 1.8 A resolution. AB - The crystal structure of the ternary complex of horse liver alcohol dehydrogenase (LADH) with the coenzyme NADH and inhibitor dimethyl sulfoxide (DMSO) has been refined by simulated annealing with molecular dynamics and restrained positional refinement using the program X-PLOR. The two subunits of the enzyme were refined independently. The space group was P1 with cell dimensions a = 51.8, b = 44.5, c = 94.6 A, alpha = 104.8, beta = 102.3 and gamma = 70.6 degrees. The resulting crystallographic R factor is 17.3% for 62 440 unique reflections in the resolution range 10.0-1.8 A. A total of 472 ordered solvent molecules were localized in the structure. An analysis of secondary-structure elements, solvent content and NADH binding is presented. PMID- 15299347 TI - Acuracy of refined protein structures. II. Comparison of four independently refined models of human interleukin 1beta. AB - To assess the accuracy of refined structures, a comparison was made using independently determined structures of the same protein in the same crystal form. The models were re-refined against a common data set to minimize the effects of different data and different refinement protocols. The process did not converge to a single model. Rather the structures differed from each other by 0.84 A which was roughly three times that predicted by a Luzzati analysis [Luzzati (1952). Acta Cryst. 5, 802-810]. The individual structures are equally valid and at least partially independent as evidenced by a reduction of the R factor by 0.013 when a simple linear combination is used. Only 29 solvent molecules were common to all four models. PMID- 15299348 TI - Structures of three crystal forms of the sweet protein thaumatin. AB - Three crystal forms of the sweet-tasting protein thaumatin from the African berry Thaumatococcus daniellii have been grown. These include two naturally occurring isoforms, A and B, that differ by a single amino acid, and a recombinant form of isoform B expressed in yeast. The crystals are of space groups C2 with a = 117.7, b = 44.9, c = 38.0 A, and beta = 94.0 degrees, P2(1)2(1)2(1) with a = 44.3, b = 63.7 and c = 72.7 A, and a tetragonal form P4(1)2(1)2 with a = b = 58.6 and c = 151.8 A. The structures of all three crystals have been solved by molecular replacement and subsequently refined to R factors of 0.184 for the monoclinic at 2.6 A, 0.165 for the orthorhombic at 1.75 A, and 0.181 for the tetragonal, also at 1.75 A resolution. No solvent was included in the monoclinic crystal while 123 and 105 water molecules were included in the higher resolution orthorhombic and tetragonal structures, respectively. A bound tartrate molecule was also clearly visible in the tetragonal structure. The r.m.s. deviations between molecular structures in the three crystals range from 0.6 to 0.7 A for Calpha atoms, and 1.1 to 1.3 A for all atoms. This is comparable to the r.m.s. deviation between the three structures and the starting model. Nevertheless, several peptide loops show particularly large variations from the initial model. PMID- 15299349 TI - Refined crystal structure of Acinetobacter glutaminasificans glutaminase asparaginase. AB - The crystal structure of glutaminase-asparaginase from Acinetobacter glutaminasificans has been reinterpreted and refined to an R factor of 0.171 at 2.9 A resolution, using the same X-ray diffraction data that were used to build a preliminary model of this enzyme [Ammon, Weber, Wlodawer, Harrison, Gilliland, Murphy, Sjolin & Roberts (1988). J. Biol. Chem. 263, 150-156]. The current model, which does not include solvent, is based in part on the related structure of Escherichia coli asparaginase and is significantly different from the structure of the enzyme from A. glutaminasificans described previously. The reason for the discrepancies has been traced to insufficient phasing power of the original heavy atom derivative data, which could not be compensated for fully by electron density modification techniques. The corrected structure of A. glutaminasificans glutaminase-asparaginase is presented and compared with the preliminary model and with the structure of E. coli asparaginase. PMID- 15299350 TI - On the application of direct methods to oligonucleotide crystallography. AB - The direct methods program SAYTAN was applied to simulated data at various resolutions from three oligonucleotides. Success in solving the structures was found to depend more upon the resolution of the data than upon errors in the data or the complexity of the structure. Collecting the data at a reduced temperature has little effect, unless it alters the mosaicity of the crystal or changes the resolution of the data. The presence of a heavy atom dramatically improved the phase refinement, particularly at low resolution. PMID- 15299351 TI - On the application of phase relationships to complex structures. XXXIV. VFOM - a new figure of merit for protein phase sets at moderate resolution. AB - In recent years it has been shown that direct methods are capable of solving the structures of small proteins. Mukherjee & Woolfson [Acta Cryst. (1993), D49, 9 12] have shown that useful phase sets can be produced even at 3 A resolution but that the standard figures of merit could not distinguish the better phase sets from others. They found modified forms of the standard figures of merit that could pick out better phase sets for 2 A resolution or higher. Gilmore, Henderson & Bricogne [Acta Cryst. (1991), A47, 842-846] have shown that evaluation of the log-likelihood gain, coming from entropy-maximization procedures, is also very successful in picking out good protein phases sets. A new figure of merit is described, based on the expected charactistics of an electron-density map for a protein, and comparisons are made with the other figures of merit mentioned above. PMID- 15299352 TI - Refined structure of concanavalin A complexed with methyl alpha-D-mannopyranoside at 2.0 A resolution and comparison with the saccharide-free structure. AB - The three-dimensional structure of the complex between methyl alpha-D mannopyranoside and concanavalin A has been refined at 2.0 A resolution. Diffraction data were recorded from a single crystal (space group P2(1)2(1)2(1), a = 123.7, b = 128.6, c = 67.2 A) using synchrotron radiation at a wavelength of 1.488 A. The final model has good geometry and an R factor of 19.9% for 58 871 reflections (82% complete), within the resolution limits of 8 to 2 A, with F > 1.0sigma(F). The asymmetric unit contains four protein subunits arranged as a dimer of dimers with approximate 222 point symmetry. Each monomer binds one saccharide molecule. Each sugar is bound to the protein by hydrogen bonds and van der Waals contacts. Although the four subunits are not crystallographically equivalent, the protein-saccharide interactions are nearly identical in each of the four binding sites. The differences that do occur between the four sites are in the structure of the water network which surrounds each saccharide; these networks are involved in crystal packing. The structure of the complex is compared with a refined saccharide-free concanavalin A structure. The saccharide free structure is composed of crystallographically identical subunits, again assembled as a dimer of dimers, but with exact 222 symmetry. In the saccharide complex the tetramer association is different in that the monomers tend to separate resulting in fewer intersubunit interactions. The average temperature factor of the mannoside complex is considerably higher than that of the saccharide-free protein. The binding site in the saccharide-free structure is occupied by three ordered water molecules and the side chain of Asp71 from a neighbouring molecule in the crystal. These occupy positions similar to those of the four saccharide hydroxyls which are hydrogen bonded to the site. Superposition of the saccharide-binding site from each structure shows that the major changes on binding involve expulsion of these ordered solvents and the reorientation of the side chain of Tyrl00. Overall the surface accessibility of the saccharide decreases from 370 to 100 A(2) when it binds to the protein. This work builds upon the earlier studies of Derewenda et al. [Derewenda, Yariv, Helliwell, Kalb (Gilboa), Dodson, Papiz, Wan & Campbell (1989). EMBO J. 8, 2198 2193] at 2.9 A resolution, which was the first detailed study of lectin saccharide interactions. PMID- 15299353 TI - Structures of human and porcine aldehyde reductase: an enzyme implicated in diabetic complications. AB - The crystal structures of porcine and human aldehyde reductase, an enzyme implicated in complications of diabetes, have been determined by X-ray diffraction methods. The crystallographic R factor for the refined porcine aldehyde reductase model is 0.19 at 2.8 A resolution. There are two molecules in the asymmetric unit related by a local non-crystallographic twofold axis. The human aldehyde reductase model has been refined to an R factor of 0.21 at 2.48 A resolution. The amino-acid sequence of porcine aldehyde reductase revealed a remarkable homology with human aldehyde reductase. The coenzyme-binding site residues are conserved and adopt similar conformations in human and porcine aldehyde reductase apo-enzymes. The tertiary structures of aldhyde reductase and aldose reductase are similar and consist of a beta/alpha-barrel, with the coenzyme-binding site located at the carboxy-terminus end of the strands of the barrel. The crystal structure of porcine and human aldehyde reductase should allow in vitro mutagenesis to elucidate the mechanism of action for this enzyme and facilitate the effective design of specific inhibitors. PMID- 15299354 TI - Raster3D Version 2.0. A program for photorealistic molecular graphics. AB - Raster3D Version 2.0 is a program suite for the production of photorealistic molecular graphics images. The code is hardware independent, and is particularly suited for use in producing large raster images of macromolecules for output to a film recorder or high-quality color printer. The Raster3D suite contains programs for composing illustrations of space-filling models, ball-and-stick models and ribbon-and-cylinder representations. It may also be used to render figures composed using other graphics tools, notably the widely used program Molscript [Kraulis (1991). J. Appl. Cryst. 24, 946-950]. PMID- 15299355 TI - Radiation damage in protein crystals at low temperature. AB - This paper describes the study of the effects of radiation damage on the quality of data collected from a protein crystal at 100 K. It is shown that radiation damage causes measurable effects in the diffraction pattern. This implies that, even at liquid nitrogen temperatures, there is a limit to the size of a crystal from which a complete data set can be collected. PMID- 15299356 TI - Crystal structural analysis of tobacco necrosis virus at 5 A resolution. AB - X-ray diffraction intensities for tobacco necrosis virus crystals were collected at 5 A resolution using a Weissenberg camera with a large cassette of radius 430 mm. The synchrotron radiation source at the Photon Factory was used. The crystal structure of the virus was obtained by 91 cycles of the non-crystallographic symmetry averaging. Secondary structures such as alpha-helices and beta structures were clearly identified in the electron-density map at 5 A resolution. This virus resembles southern bean mosaic virus both in orientation of coat protein subunits and in their folding. Ordered and disordered parts of each subunit of tobacco necrosis virus are shorter and longer than the corresponding parts of the southern bean mosaic virus by 12 and 27 residues, respectively. PMID- 15299357 TI - Crystal structure of a monoclinic form of dihydropteridine reductase from rat liver. AB - A binary complex of dihydropteridine reductase and NADH crystallizes in the space group C2, with a = 222.2, b = 46.5, c = 95.3 A and beta = 101.1 degrees. There are two dimers in the asymmetric unit. The structure was solved by molecular replacement techniques and refined with 2.6 A data to a crystallographic R factor of 16.8%. Each dimer has twofold non-crystallographic symmetry and the four individual monomers in the asymmetric unit have the same overall molecular conformation. PMID- 15299358 TI - Comparison of homology models with the experimental structure of a novel serine protease. AB - A model structure of the human complement enzyme factor D was built based on homology with related serine proteases. A molecular-replacement solution of the factor D crystal structure employing the homology model refined without manual intervention to an R factor of 0.249 with 2.4 A native diffraction data. A multiple isomorphous replacement (MIR) electron-density map was subsequently produced, leading to a model refined at 2.0 A resolution to an R factor of 0.188. A homology model built with commercial modeling software was subjected to the same procedure. Comparisons of the homology models with the final refined MIR structure are presented. Major discrepancies were found in critical active-site regions. PMID- 15299359 TI - Error detection in crystallographic models. AB - A variety of criteria were tested for identifying errors in protein crystal coordinates. Statistical analysis was based on comparisons of a highly refined crystal structure and several preliminary models derived from molecular replacement. A protocol employing temperature factors, real-space fit residuals, geometric strains, dihedral angles and shifts from the previous refinement cycle is developed. These results are generally applicable to the detection of errors in partially refined protein crystal structures. PMID- 15299360 TI - Crystallization and preliminary X-ray crystallographic study of ribosome inactivating protein from barley seeds. AB - Ribosome-inactivating protein from barley seeds has been crystallized using polyethylene glycol as precipitant. The crystal belongs to the monoclinic space group C2, with unit-cell parameters a = 88.36, b = 62.59, c = 53.18 A and beta = 108.62 degrees. The asymmetric unit contains one molecule of ribosome inactivating protein with a corresponding crystal volume per protein mass (V(m)) of 2.32 A(3) Da(-1) and a solvent content of 47% by volume. The crystal diffracts to about 2.3 A with X-rays from a rotating-anode source and is very stable in the X-ray beam. X-ray data (nearly complete to 2.4 A Bragg spacing) have been collected from a native crystal. PMID- 15299361 TI - Crystallization and preliminary X-ray diffraction study of the flavoenzyme 2,4 pentadienoyl-CoA reductase from Clostridium aminovalericum. AB - The tetrameric flavoenzyme 2,4-pentadienoyl-CoA reductase has been crystallized from solutions containing polyethylene glycol as precipitant. The crystals grow in the monoclinic space group C2 with unit-cell dimensions a = 160.2, b = 120.2, c = 95.3 A, beta = 99.0 degrees. The packing parameter V(M) is 2.3 A(3) Da(-1) (Matthews parameter) for four monomers per asymmetric unit. Complete data sets to about 2.9 A resolution have been collected. PMID- 15299362 TI - Crystallization and preliminary X-ray analysis of anti-digoxin antibodies. AB - The Fab fragments of several monoclonal antibodies that bind digoxin and other cardiac glycosides have been screened for crystallization conditions. We have crystallized two of these in forms suitable for X-ray analysis. The anti-digoxin antibody 40-50 in complex with ouabain crystallizes with symmetry consistent with space group C2, with a = 92.6, b = 85.0, c = 73.0 A and beta = 131.7 degrees. This crystal form shows considerable non-isomorphism between crystals. A second anti-digoxin antibody, 49-10, crystallizes with symmetry consistent with space group P2(1)2(1)2, with a = 95.3, b = 147.1 and c = 76.2 A. PMID- 15299364 TI - Using genetic algorithms for solving heavy-atom sites. AB - A novel procedure has been developed for locating heavy-atom positions in crystals of macromolecules. This method used genetic algorithms (GA's) to search for heavy-atom sites that are consistent with an observed difference Patterson function. The procedure is straightforward to apply, space-group independent, and particularly powerful for cases involving non-crystallographic symmetry of multiple heavy atoms in the asymmetric unit. In this paper, we introduce how GA's are used for determining the heavy-atom positions and show how this method is more efficient than a sequential search. PMID- 15299365 TI - Enhancement of the method of molecular replacement by incorporation of known structural information. AB - Crystals of macromolecules often have two or more molecules per asymmetric unit, or contain domains of a macromolecule or a macromolecular complex that are structurally independent. In such cases the conventional molecular-replacement method attempts to determine the position of each structural unit independently. Typically, some parts of the structure can be determined more easily or more reliably than other parts. Methods are proposed whereby information from a part of a crystal structure that has been determined can be used to help determine the structure of the remainder. Two different strategies are discussed, 'subtraction' and 'addition'. With 'subtraction' strategy the Patterson function of the known part of the structure is subtracted from the 'observed' Patterson. This approach is found to be most effective in the context of the rotation function in that it eliminates peaks that are irrelevant to the desired solution. With 'addition' strategy the structure factors of the known component are added to those of the search model. This procedure is most effective in the context of the translation function because it brings the structure factors calculated from the search model closer to those observed. Methods of applying the fast Fourier transform to facilitate these calculations are described. A number of examples are provided including structures of mutants of T4 lysozyme that might not have been solved without recourse to the proposed methods. A method of including information from a heavy-atom derivative in a translation function is also developed and shown to be superior in some situations to the conventional translation function. PMID- 15299366 TI - Crystallization of wild-type and mutant ferricytochromes c at low ionic strength: seeding technique and X-ray diffraction analysis. AB - Ferricytochromes c were crystallized at low ionic strength by macroseeding techniques. Large crystals were grown by seed-induced self-nucleation which occurred anywhere in the drop, regardless of the location of the seed crystal. This unusual crystal-seeding method worked reproducibly in our hands, and X-ray quality crystals have been prepared of several ferricytochromes c: horse, rat (recombinant wild type), and two site-directed mutants of the latter, tyrosine 67 to phenylalanine (Y67F) and asparagine 52 to isoleucine (N52I). Crystals of any one of these four proteins could be used as seeds for the crystallization of any one of the others. All the crystals are of the same crystal form, with space group P2(1)2(1)2(1). There are two protein molecules per asymmetric unit. The crystals are stable in the X-ray beam and diffract to at least 2.0 A, resolution. Full crystallographic data sets have been collected from single crystals of all four proteins. PMID- 15299367 TI - Core tracing: depicting connections between features in electron density. AB - Core tracing is a threshold-independent method of determining connectivity (long chains of high-density values) in electron-density maps. It gives visually sparse pictures of large volumes which are useful for initial fitting and for molecular boundary determination. New methods for visual presentation of the traces are suggested by the way that the connectivity is parameterized in terms of local connections between maxima and the saddle (lowest) points along the connecting paths. The algorithm also partitions the density into small compact volumes containing the maxima. These volumes are useful for localization and statistical analysis. PMID- 15299368 TI - Accuracy and precision in protein crystal structure analysis: two independent refinements of the structure of poplar plastocyanin at 173 K. AB - The structure of the copper protein plastocyanin from poplar leaves (Populus nigra var. italica) at 173 K has been subjected to two independent refinements, using a single set of synchrotron X-ray data at 1.6 A resolution. Energy restrained refinement using the program EREF resulted in lower root-mean-square deviations from ideal geometry (e.g. 0.011 A for bond lengths) but a higher residual R (0.153) than restrained least-squares refinement using the program PROLSQ (0.014 A, 0.132). Electron-density difference maps in both refinements provided evidence for disorder at some side chains and solvent atoms, and the PROLSQ refinement made allowance for this disorder. The number of solvent sites identified at the 4sigma(rho) level was 171 in the EREF refinement and 189 in the PROLSQ refinement; 159 of the solvent sites are common to both refinements within 1 A. The root-mean-square differences between the atomic positions produced by the two refinements are 0.08 A for C(alpha) atoms, 0.08 A for backbone atoms and 0.12 A for all non-H atoms (excluding six obvious outliers) of the protein molecule. The two sets of Cu-ligand bond lengths differ by up to 0.07 A, and the ligand-Cu-ligand angles by up to 7 degrees. At 173 K the volume of the unit cell is 4.2% smaller than at 295 K. Greater order in the solvent region is indicated by the location of 79 more solvent sites, the identification of extensive networks of hydrogen-bonded rings of solvent molecules, and a general decrease in the thermal parameters. Within the unit cell, the protein molecules are significantly translated and rotated from their positions at ambient temperature. An important structural change at low temperature is a 180 degrees flip of the peptide group at Ser48-Gly49. Nearly all other significant differences between the structures of the protein at 173 and 295 K occur at exposed side chains. If the backbone atoms in the 173 and 295 K structures are superposed, excluding atoms involved in the peptide flip, the root-mean- square difference between the positions of 393 atoms is 0.25 A. Two internal water molecules, not included in previous descriptions of poplar plastocyanin, have been located. The plastocyanin Cu-site geometry at 173 K is not significantly different from that at 295 K. If plastocyanin undergoes a change in Cu-site geometry at low temperature, as has been suggested on the basis of resonance Raman spectroscopic evidence, then the change is not detected within the limits of precision of the present results. PMID- 15299369 TI - Differences in anionic inhibition of human carbonic anhydrase I revealed from the structures of iodide and gold cyanide inhibitor complexes. AB - The crystal structures of two anionic inhibitor complexes of human carbonic anhydrase I (HCAI), namely, HCAI-iodide and HCAI-Au(CN)(2)(-), have been refined by the restrained least-squares method at 2.2 and 2 A nominal resolution, respectively, with good stereochemistry for the final models. The R values have improved from 30.3 to 16.6% for HCAI-iodide and from 28.8 to 17.1% for HCAI Au(CN)(2)(-). The sites of inhibitor binding as elucidated are totally different in the two structures. The iodide anion replaces the zinc-bound H(2)O/OH(-) ligand and renders the enzyme inactive. This result confirms that the zinc-bound H(2)O/OH(-) is the activity-linked group in carbonic anhydrase enzymes. Au(CN)(2)(-) binds at a different and new site near the zinc ion, without liganding to the metal. The N atom of Au(CN)(2)(-) is within hydrogen-bonding distance of the zinc-bound H(2)O/OH(-) group which shifts by about 0.4 A away from the zinc ion in relation to its position in the native HCAI. It is proposed that the presence of the inhibitor Au(CN)(2)(-) results in a conformational reorientation of the activity-linked group, due to hydrogen-bond formation with the inhibitor, which in turn sterically hinders the binding of the substrate CO(2) molecule in the active site, leading to the inhibition of HCAI enzyme. PMID- 15299370 TI - A crystallographic study of haem binding to ferritin. AB - Ferritin, the iron-storage protein, binds porphyrins, metalloporphyrins and the fluorescent dyes ANS (8-anilino-1-naphthalenesulfonic acid) and TNS (2-p toluidinyl-6-naphthalenesulfonic acid), similarly to apo-myoglobin. Octahedral crystals of horse-spleen apo-ferritin (HSF; 174 amino acids) complexes prepared by the addition of haem, hematoporphyrin or Sn-protoporphyrin IX to a solution of apo-ferritin crystallize in space group F432 with cell parameter a = 184.0 A. X ray crystallographic analysis of single crystals prepared from a mixture containing haem or Sn-protoporphyrin IX shows that the haem-binding sites in these crystals are occupied by protoporphyrin IX, which is free of metal, rather than by the original metalloporphyrin. The present paper describes the structure of horse-spleen apo-ferritin cocrystallized with Sn-protoporphyrin IX. The 6797 reflections up to 2.6 A resolution used in the refinement were obtained from a data set recorded on a Nicolet/Xentronics area detector with Cu Kalpha radiation from a Rigaku RU 200 rotating anode. The final structure comprises 1613 non-H atoms, two Cd atoms and 170 solvent molecules. Four residues are described as disordered. The root-mean-square deviations from ideal bond lengths and angles are 0.013 A and 2.88 degrees, respectively. Protoporphyrins are observed in special positions on the twofold axes of the ferritin molecule with a stoichiometry of 0.4 per subunit. PMID- 15299371 TI - Preliminary crystallographic analysis of glyceraldehyde 3-phosphate dehydrogenase from the extreme thermophile Thermus aquaticus. AB - Crystals have been obtained of glyceraldehyde 3-phosphate dehydrogenase from the extreme thermophile, Thermus aquaticus. This enzyme is stable and active at 363 K, thus its three-dimensional structure should add insight into the structural basis of protein thermostability. Large high-quality crystals were grown using isopropanol and polyethylene glycol at pH 8.4. They crystallize in the orthorhombic space group P2(1)2(1)2(1) with cell dimensions a = 144.77 (6), b = 148.77 (5), c = 149.50 (7) A, and diffract to beyond 2.8 A. The volume of the unit cell and the packing observed in other GAPDH structures suggest that there are two tetramers per asymmetric unit. With 300 kDa/asymmetric unit expected in this form, its solution represents a challenging molecular replacement problem. A low-resolution data set has been recorded and used to carry out self-rotation, cross-rotation and Patterson-correlation refinement calculations. We found that the Q molecular axes of both tetramers are approximately coincident with the crystallographic a axis, and the non-crystallographic symmetry relating the two tetramers is approximately a rotation of 90 degrees about the a axis. PMID- 15299372 TI - High-resolution structures of single-metal-substituted concanavalin A: the Co,Ca protein at 1.6 A and the Ni,Ca-protein at 2.0 A. AB - The molecular structures of cobalt- and nickel-substituted concanavalin A have been refined at 1.6 and 2.0 A resolution, respectively. Both metal derivatives crystallize in space group I222 with approximate cell dimensions a = 89, b = 87 and c = 63 A and one monomer in the asymmetric unit. The final R factor for Co substituted concanavalin A is 17.8% for 29 211 reflections with F > 1.0sigma(F) between 8.0 and 1.6 A. For Ni-substituted concanavalin A the final R factor is 15.9% for 16 128 reflections with F > 1.0sigma(F) between 8.0 and 2.0 A resolution. Both structures contain a transition-metal binding site and a calcium binding site but, unlike Cd-substituted concanavalin A, do not have a third metal binding site. The Co-substituted concanavalin A structure diffracts to the highest resolution of any concanavalin A structure reported to date. A comparison of the structures of Ni-, Co-, Cd-substituted and native concanavalin A gives an indication of coordinate errors, which is a useful baseline for comparisons with saccharide complexes of concanavalin A described in other work. We also give a detailed account of multiple conformations which were found for five side-chain residues. PMID- 15299373 TI - Crystallization of hemoglobins II and III of the symbiont-harboring clam Lucina pectinata. AB - Diffraction data to 2.7 A resolution were measured on crystals of the homotetramers of components II and III of the cytoplasmic hemoglobin of the symbiont-harboring clam Lucina pectinata. Even though the crystallization conditions are different and the sequence homology of the two hemoglobins is only 63%, the crystals are isomorphous to each other and to the heterotetramer Hb II/III, implying that the residues primarily involved in the intermolecular interactions and responsible for crystal cohesion may be invariant. PMID- 15299374 TI - The CCP4 suite: programs for protein crystallography. AB - The CCP4 (Collaborative Computational Project, number 4) program suite is a collection of programs and associated data and subroutine libraries which can be used for macromolecular structure determination by X-ray crystallography. The suite is designed to be flexible, allowing users a number of methods of achieving their aims and so there may be more than one program to cover each function. The programs are written mainly in standard Fortran77. They are from a wide variety of sources but are connected by standard data file formats. The package has been ported to all the major platforms under both Unix and VMS. The suite is distributed by anonymous ftp from Daresbury Laboratory and is widely used throughout the world. PMID- 15299375 TI - Use of low-molecular-weight polyethylene glycol in the crystallization of RNA oligomers. AB - We have crystallized a variety of RNA oligonucleotides in a form suitable for X ray diffraction studies using polyethylene glycol with a low-molecular-weight distribution (PEG 400) as the precipitant. Crystallization experiments on a set of 26 RNA oligomers ranging from eight to 12 nucleotides in length resulted in eight diffraction-quality crystals. Of these eight RNA crystals, six utilized PEG 400 as the precipitating agent. We have also been able to obtain large single crystals of a DNA-RNA hybrid, transfer RNA (two different conditions) and a catalytic RNA from PEG 400 solutions. These results suggest that PEG 400 may be a generally useful alternative to 2-methyl-2,4-pentanediol (MPD) which has, thus far, been the most successful precipitant for DNA oligomers. PMID- 15299376 TI - Refined three-dimensional structure of the Fab fragment of a murine IgGl,lambda antibody. AB - We report the cDNA sequence determination and the crystal structure of the Fab fragment of a murine IgG1,lambda antibody (HC19), specific for an influenza virus hemagglutinin. The HC19 Fab-fragment structure has been refined; the crystallographic R-factor is 19.5% at 2.3 A resolution. We have compared the conformation of HC19 complementarity determining regions (CDRs) with those of CDR loops of Fab structures available from the Protein Data Bank. These loops were chosen based on the identity of key residues, following the canonical-structure approach; four CDRs have a main-chain conformation very similar to the canonical structure that had been identified. HC19 L1 CDR adopts a conformation clearly distinct from all L1 CDRs that belong to a chain of a different class or origin; this is determined by the nature of a few residues at positions in the sequence different from those of key residues in other light chains. This canonical structure should be representative of most murine lambda-class light chains, as inferred from the very high sequence homologies of these polypeptides. PMID- 15299377 TI - Crystallization and preliminary X-ray analysis of the river buffalo (Bubalus bubalis L.) BB phenotype carbonmonoxyhaemoglobin. AB - Ruminant haemoglobin (Hb) extracted from river buffalo (Bubalus bubalis) has been purified and crystallized. Two different Hb forms of the phenotype BB gave isomorphous crystals which diffracted to 2.8 A resolution and were not sensitive to radiation damage. Crystals of CO Hb have space group P2(1)2(1)2(1) with unit cell parameters a = 54.8, b = 64.0, c = 158.6 A, and contain one Hb molecule per asymmetric unit. PMID- 15299378 TI - Location of active sites of NiFe hydrogenase determined by the combination of multiple isomorphous replacement and multiwavelength anomalous-diffraction methods. AB - The active centers of NiFe hydrogenase from Desulfovibrio vulgaris Miyazaki F have been located in the electron-density map calculated at 4 A resolution. The electron-density map based on five heavy-atom derivatives showed four strong peaks which were clearly distinguished from the protein region. These strong densities have been successfully assigned to three iron-sulfur clusters and one Ni atom by a difference Fourier technique with coefficients of the best phases from the multiple isomorphous replacement (MIR) method and structure factors obtained at five wavelengths (1.040, 1.487, 1.730, 1.743 and 1.750 A) with the use of a synchrotron radiation source. Four active centers are approximately lined up at a distance of ca 13 A, which seems reasonable if they are connected with the electron-transfer chain. PMID- 15299379 TI - Crystallization of human methylamine-treated complement C3 and C3b. AB - Human methylamine-treated complement C3 (C3-MA) and C3b (C3b-MA) have been crystallized using ammonium sulfate as precipitant. The crystals of the two compounds are morphologically indistinguishable though they belong to different space groups. We show that only minor alterations in packing are responsible for the change in space group. Crystals of C3-MA are tetragonal [P4(1(3))22, a = b = 135, c = 610 A] with two molecules per asymmetric unit. Crystals of C3b-MA are also tetragonal [P4(1(3))2(1)2, a = b = 191, c = 610 A] with four molecules per asymmetric unit. The maximum diffraction observed is 7.7 A at cryogenic temperature using synchrotron radiation. PMID- 15299380 TI - Crystallization and preliminary X-ray diffraction studies of bacterial ribosomal protein L14. AB - Based on amino-acid sequence homology, it is predicted that ribosomal protein L14 is a member of a recently identified family of structurally related RNA-binding proteins. To verify this, the gene for Bacillus stearothermophilus L14 has been cloned, and the protein has been purified and crystallized. The crystals are in space group C2 with cell dimensions a = 67.0, b = 32.7, c = 49.4 A, and beta = 101.8 degrees, and there is one molecule in the asymmetric unit (V(m) = 2.0 A(3) Da(-1)). They are of high quality, and a native data set has been collected to a resolution of 1.6 A, with an R(merge) of 5.3%. PMID- 15299382 TI - Crystallogenesis of biological macromolecules: facts and perspectives. AB - This paper gives an overview of the science of crystals of biological macromolecules. The historical background of the field is outlined and the main achievements and open problems are discussed from both biological and physical chemical viewpoints. Selected results, including data from the authors, illustrate this overview. The perspectives of crystallogenesis for structural biology, but also more general trends, are presented. PMID- 15299383 TI - Production and crystallization of virus-like particles assembled in a heterologous protein expression system. AB - It is of considerable interest to separate the processes of viral infectivity and virion assembly. Until recently this has only been possible with viruses that could be disassembled and reassembled in vitro. Even in these cases it was difficult to establish the authenticity of reassembled capsid protein because of possible irreversible damage that may have occurred to the protein during disassembly. An ideal method for the study of virus assembly is a protein expression system in which conditions are appropriate for spontaneous particle formation from freshly synthesized polypeptides. The baculovirus expression system has proven to be an excellent means to this end. Recently, this approach has been used to study the T = 3 Flock House insect virus and it has been demonstrated that subunits with the wild-type protein sequence, and with site specific mutations that prevent particle maturation, will assemble and crystallize. This same approach has now been used at Purdue to study the T = 4 Nudaurelia omega capensis insect virus. There is no cell culture system currently available for the study of NomegaV, thus the expression system provides the first opportunity to study assembly under controlled conditions. PMID- 15299384 TI - Influence of polydispersity on protein crystallization: a quasi-elastic light scattering study applied to alpha-amylase. AB - The early stages of the crystallization process of porcine pancreatic alpha amylase were investigated by quasi-elastic light scattering. It is shown that at 288 and 293 K the diffusion coefficient does not monotonically change with increasing protein concentration but passes through a maximum at 10 mg ml(-1). In supersaturated solutions, prior to nucleation, the protein is strictly monodisperse. Nucleation induces the formation of aggregates and a polydispersity of, for example, 18% for an initial supersaturation C/C(e) = 5.8. Monodispersity is restored after the nuclei have grown and partially consumed the solute. On the other hand, polydispersity increases up to 20% at 298 K if the protein concentration decreases to 3-4 mg ml(-1), values at which the solutions are under saturated. When the protein concentration exceeds 5-6 mg ml(-1) the protein becomes monodisperse again. These results, confirmed by those of another system we are studying (bovine pancreatic trypsin inhibitor), are at variance with the statements that supersaturation is always at the origin of aggregation and polydispersity, and that in undersaturated solutions the diffusion coefficient should remain constant for obtaining crystals once the solutions are supersaturated. PMID- 15299385 TI - Predicting protein crystallization from a dilute solution property. AB - A dilute solution parameter obtained from static light-scattering measurements is proposed as a predictor for protein crystallization experiments. The osmotic second virial coefficients, B(22), have been measured for a variety of proteins in solvents that are known to promote crystallization and the values for B(22) were found to lie within a fairly narrow range which we refer to as a crystallization slot. Solution conditions which were known not to favor crystallization of the proteins resulted in B(22) values well outside the crystallization slot. PMID- 15299386 TI - Crystallization of previously desalted lysozyme in the presence of sulfate ions. AB - Lysozyme, which is known to crystallize readily in the presence of many salts, has never been crystallized by salting out with ammonium sulfate. In the present study, lysozyme was first completely desalted by treatment with strong cation- (H(+) form) and anion- (OH(-) form) exchange resins. This leads to a protein solution with only H(+) and OH(-) as counterions, corresponding to its isoionic point. Addition of 2.5-3 molar equivalents of H(2)SO(4) to isoionic lysozyme decreases the pH value to 9-8 and allows crystallization to take place. The space group was found to be P4(3)2(1)2, similar to the classical lysozyme crystals grown in the presence of NaCl at pH 4.5, with unit-cell dimensions a = b = 78.9, c = 38.5 A. Tentative explanation of the sulfate/lysozyme interaction was addressed by mass spectrometry, and shows non-covalent binding of the ions on the protein. PMID- 15299387 TI - Crystallization and preliminary X-ray analysis of recombinant bovine cellular retinoic acid-binding protein. AB - Crystals of bovine cellular retinoic acid-binding protein (CRABPI) have been grown from protein expressed in E. coli. Two different crystal forms were obtained. Crystals containing protein with the ligand all-trans retinoic acid belong to space group P4(1) or P4(3), a = b = 41.36, c = 202.71 A and diffract to 2.5 A. Crystals of CRABP with the synthetic retinoid analogue Am80 diffract to 1.9 A with space group P2(1) and cell dimensions a = 37.03, b = 105.93, c = 40.31 A, beta = 110.28 degrees. Considerations in the crystallization of proteins with light-sensitive ligands are discussed. PMID- 15299388 TI - Fusion proteins as tools for crystallization: the lactose permease from Escherichia coli. AB - A novel strategy is presented for the crystallization of membrane proteins or other proteins with low solubility and/or stability. The method is illustrated with the lactose permease from Escherichia coli, in which a fusion is constructed between the permease and a 'carrier' protein. The carrier is a soluble, stable protein with its C and N termini close together in space at the surface of the protein, so that the carrier can be introduced into an internal position of the target protein. The carrier is chosen with convenient spectral or enzymatic properties, making the fusion protein easier to handle than the native molecule. Data are presented for the successful construction, expression and purification of a fusion product between lactose permease and cytochrome b(562) from E. coli. The lactose transport activity of the fusion protein is similar to that of wild type lactose permease, and the fusion product has an absorption spectrum in the visible range which is essentially identical to that of cytochrome b(562). The fusion protein has a higher proportional polar surface area than wild-type permease, and should have better possibilities of forming the strong directional intermolecular contacts required of a crystal lattice. PMID- 15299389 TI - Enzymatic deglycosylation as a tool for crystallization of mammalian binding proteins. AB - Enzymatic deglycosylation has been used in attempts to crystallize several glycoproteins with the aim of overcoming the problems resulting from heterogeneity and flexibility of the attached glycan chains. An endoglycosidase preparation from Flavobacterium meningosepticum, comprising the enzymes endo F and PNGase-F, was used in experiments on the mammalian binding proteins lactoferrin and haemopexin. Significant differences were found in the susceptibility of different proteins to deglycosylation. For human lactoferrin (Lf) and its recombinant N-terminal half-molecule (Lf(N)), deglycosylation was rapid and complete, and was essential for obtaining high-quality crystals of both apo-Lf and Lf(N); for bovine Lf, however, complete deglycosylation did not occur. Similarly, for rabbit haemopexin the carbohydrate chain on the C-terminal domain was easily removed, but the three chains on the N-terminal domain proved more resistant and their removal led to some fragmentation of the protein. Nevertheless, this approach provided the only means of crystallizing the C terminal domain and is likely to be useful for other glycoproteins. PMID- 15299390 TI - Light-scattering investigations of nucleation processes and kinetics of crystallization in macromolecular systems. AB - Quasi-elastic light scattering (QELS) was used to investigate quantitatively the mechanisms of nucleation, postnucleation growth, and dissolution in ensembles of both crystalline and amorphous aggregates of satellite tobacco mosaic virus (STMV), ferritin, apoferritin and pumpkin seed globulin. At low supersaturation conditions, as described previously for small molecule crystallization, the metastable region was obtained. Under these conditions aggregation took place, but crystallization did not proceed and critical nuclei did not form over a long period of time. The critical solution supersaturation necessary to obtain crystals, sigma = ln(c/s) where c and s are concentration and solubility of protein, varied from approximately 0.1 for pumpkin seed globulin to approximately 0.9 for STMV. For higher supersaturation conditions when aggregation processes leading to formation of crystals are not established immediately but after a certain induction period, the supersaturation-dependent critical nuclear size, R(c), for different macromolecular systems was estimated from time-dependent size distribution analyses to be in the range of approximately 10(3) for proteins such as pumpkin globulin to approximately 10 for virus particles. From the same data, the molar interfacial free energy was deduced to be 3.3-9.2 kJ mol(-1). These are believed to be among the first estimates for macromolecular crystals. Under conditions of moderate supersaturation where induction periods preceded the appearance of critical nuclei, the potential barriers for formation were estimated to be in the range 8.3-50 kJ mol(-1). Growth and dissolution kinetics for pumpkin seed globulin were investigated. These experiments allowed determination of protein solubility versus solution temperature, protein and precipitant concentrations. Aggregation patterns which lead to crystal formation are distinctly different to those which produce an amorphous precipitate. The results provide additional evidence that QELS can be used to find general criteria that allow one to discriminate between conditions for a given protein system leading to crystalline or amorphous states at early stages of the aggregation process. PMID- 15299391 TI - Investigation of nucleating lysozyme solutions. AB - This laboratory has explored the potential of a combination of three analytical techniques to study the nucleation of chicken egg-white lysozyme. Collisional quenching of the fluorescent molecule SPQ [6-methoxy-N-(3 sulfopropyl)quinolinium] by chloride ions was used to determine the binding of the crystallizing agent to the protein at equilibrium and kinetically. Calorimetric measurements show that this binding generates an exothermic peak larger than the energy released during the early stages of nucleation. Light scattering intensity measurements were used to follow the aggregation kinetics. PMID- 15299392 TI - Evidence for a dimeric intermediate on the crystallization pathway of ribonuclease A. AB - Early steps in the crystallization process of pancreatic ribonuclease have been investigated by time-dependent fluorescence anisotropy, using a labeled protein as a fluorescent probe. Previous experiments have shown that steady-state fluorescence anisotropy is sensitive to protein-protein interactions and can be used to find new crystallization conditions. The present work describes an attempt, by means of time-resolved experiments, to detect and characterize species appearing in the early stages of the crystallization pathway. Fluorescence anisotropy decay was measured with synchrotron radiation as a light source under a variety of conditions where it is known that the solutions tend towards crystallization; the decay was analyzed by a maximum-entropy method that calculates a rotational correlation-time distribution. Fluorescence anisotropy originates in the Brownian rotatory motion of macromolecules and the values of the correlation times are related to the size and shape of different species present in the solution. In the presence of high salt concentrations, a bimodal distribution is always observed. Whereas a peak of protein monomer is still present, a second peak appears as a stable intermediate in the crystallization pathway. The correlation time of this new species varies between two and three times the correlation time of the monomer. The second peak is possibly the symmetrical dimer of the ribonuclease molecules commonly observed in all the high salt crystal forms. PMID- 15299393 TI - Improved crystals of the toxic protein MAP by protein engineering towards the host specificity. AB - Mirabilis anti-viral protein (MAP) is a ribosome-inactivating protein from Mirabilis jalapa L. Since MAP is effective over a broad spectrum of species, the protein is difficult to express in heterologous hosts such as Escherichia coli. Recently, we obtained a MAP mutant, Y72F which exhibits a lower (1/100) activity against E. coli ribosomes while retaining almost full activity against mammalian cells [Habuka, Miyano, Kataoka, Tsuge & Noma (1992). J. Biol. Chem. 267, 7758 7760]. For the crystallographic studies, the Y72F MAP expression vector with an OmpA leading sequence was constructed and expressed in E. coli. The Y72F MAP mutant was then isolated and purified from the cell culture medium. Crystals were grown using the crystallization conditions for the native MAP crystals [Miyano et al. (1992). J. Mol. Biol. 226, 281-283]: 50% ammonium sulfate containing 50 mM ammonium citrate and 2 mM adenine sulfate, pH 5.4. The crystals belong to space group P3(1)21 (or P3(2)21) with a = b = 104.1 and c = 134.3 A. The crystals are isomorphous with the wild-type crystals but diffract to higher resolution. Imaging-plate photographs of the Y72F mutant showed sharp intense spots without the streaking observed in the native crystals. PMID- 15299394 TI - Biological Macromolecule Crystallization Database, Version 3.0: new features, data and the NASA archive for protein crystal growth data. AB - Version 3.0 of the NIST/NASA/CARB Biological Macromolecule Crystallization Database (BMCD) includes crystal and crystallization data on all forms of biological macromolecules which have produced crystals suitable for X-ray diffraction studies. The data include summary information on each of the macromolecules, crystal data, crystallization conditions and comments about the crystallization procedure if it varies from the traditional methods employed for crystal growth. The database-management software maintains continuity with previous versions providing similar search procedures and displays. Version 3.0 of the BMCD includes protocols and results of crystallization experiments undertaken in space. These new data are comprised of both the NASA Protein Crystal Growth Archive, which includes information on all NASA-sponsored protein crystal growth experiments, and data describing other internationally sponsored microgravity macromolecule crystallization studies. The entries for the space growth crystallization experiments contain the crystallization protocols, apparatus descriptions, flight summary data, indication of success or failure of the experiments, references, etc. Other new features of the BMCD include the addition of crystallization procedures for small peptides and cross references to other structural biology databases. PMID- 15299395 TI - Screening and optimization strategies for macromolecular crystal growth. AB - Today the determination of successful crystallization conditions for a particular macromolecule remains a highly empirical process. Sparse-matrix and grid screening procedures are rapid and economical means to determine preliminary crystallization conditions. During optimization the variable set (pH, precipitant type and precipitant concentration) utilized in these procedures is screened in an attempt to determine appropriate conditions for the nucleation and growth of single crystals suitable for X-ray diffraction analysis. Unfortunately, in many cases this strategy will not produce single crystals suitable for X-ray diffraction analysis. We have explored, in an empirical sense, other tools for use during optimization. First, a new screening protocol is evaluated which employs less classical precipitating agents. Second, a set of 24 electrostatic crosslinking agents are evaluated for their ability to promote crystallization. Third, a panel of more than 30 detergents are evaluated for their ability to prevent sample aggregation and influence crystal growth. PMID- 15299396 TI - Orthorhombic lysozyme solubility. AB - The orthorhombic, or high-temperature, form of chicken egg-white lysozyme typically appears at temperatures >/=298 K. Solubility diagrams have been determined for this form of lysozyme from pH 4.0 to 5.4 in 0.2 pH increments using the micro-column technique. Data were collected in the 297-317 K temperature range which resulted in phase diagrams similar in overall shape to those obtained for the lower temperature tetragonal form. Specifically, the solubility increased with increasing temperature and decreased with increasing precipitant concentration. However, the solubility of the orthorhombic form is considerably less sensitive to temperature than the tetragonal form, resulting in a more flattened slope. On the other hand, pH effects on the high-temperature form were opposite to those on the low-temperature form. When holding the precipitant concentration constant, the solubility decreased with increasing pH for the orthorhombic form. Previous tetragonal data were incorporated with these orthorhombic data to produce intercept values. These values varied with both pH and precipitant concentration, but the general tendency of the slope was to decrease with increasing pH. PMID- 15299397 TI - Search designs for protein crystallization based on orthogonal arrays. AB - In protein crystallography, the initial experimental problem is the identification of physical and chemical conditions that will support nucleation and crystal growth. Ideally, experiments to search for such conditions would be based on a full-factorial structure, with variation in the temperature and solution composition. However, consideration of even a moderate number of possibilities for the composition of the system will result in factorial experiments which may be prohibitively large. In this paper it is proposed that search experiments for protein crystallization might be based on orthogonal arrays. These are subsets of full-factorial experiments which possess a great deal of symmetry, such that a uniform distribution of points throughout the experimental region is preserved. Such experiments have reasonable size, explore the proposed experimental region in a systematic fashion, and form a logical basis for a sequential approach to the search for crystallization conditions. Examples of such initial search experiments are given, and their application to some recent protein crystallization problems in this laboratory is described briefly. The relationship of this approach to other protein crystallization search procedures is also discussed. PMID- 15299398 TI - Predispensed gradient matrices - a new rapid method of finding crystallization conditions. AB - Predispensed gradient matrices allow the boundary between precipitate and clear solution to be located very rapidly for a particular protein and precipitant. In many cases crystals grow in the trials which were used to identify this boundary. The method involves dispensing a series of between 10 and 72 microbatch trials in which some parameter, such as precipitant concentration, is gradually changed. (Protein is not dispensed at this stage.) Protein is then added to selected trials using a predetermined algorithm, which takes into account the level of precipitation caused by previous additions. Thirteen crystal forms were obtained using the method with eight proteins and eight precipitants. Six forms were prisms or plates with maximum dimensions above 400 microm. PMID- 15299399 TI - Crystallization experiments with 2-enoyl-CoA hydratase, using an automated 'fast screening' crystallization protocol. AB - A convenient method for screening crystallization conditions using an automated fast-screen protocol has been implemented and tested on an enoyl-CoA hydratase. The crystallization solutions for the initial screening and subsequent optimizations are prepared using a crystallization robot. Enoyl-CoA hydratase (E.C. 4.2.1.17), purified from rat-liver mitochondria, is one of the enzymes from the beta-oxidation pathway of fatty-acid metabolism; it catalyzes the reversible hydration of 2-trans-enoyl-CoA's to L-3-hydroxy-acyl-CoA's. Different crystal forms, diffracting to 3.0 A, were obtained. PMID- 15299400 TI - Reverse screening. AB - Major emphasis has been placed in recent years on kits for screening crystallization conditions of macromolecules. Such approaches have undoubtedly speeded up the initial screening and, to a certain extent, helped in reducing the amount of protein required for the initial survey. Factorial screening techniques, either full-factorial or sparse-matrix approaches, have proved successful in the crystallization of many proteins. However, in cases where the amount of protein is limited a systematic approach based on an a priori choice of precipitants may be preferable to an extensive search. The approach described here targets such situations. The approach consists of the determination of the solubility characteristics of the macromolecule under study as a function of precipitant and macromolecule concentrations to define a working range for these parameters. Conditions under which the protein is highly supersaturated, and hence more conducive to nucleation, are established so as to favor the formation of an initial stable nucleus which can be one of the dominant problems that hinders successful crystallization of proteins. Later, changes in solubility as a function of pH and as a result of the introduction of additives are evaluated. In addition, when ligands are available for the formation of macromolecular complexes, screening of different complexes is used as a means to increase the probability of obtaining crystals. Solubility information derived from one, or more, complexes that have been screened can be used for comparison and to aid in the crystallization of other complexes. Cross-seeding between complexes is an intrinsic part of the method and provides an efficient way of obtaining crystals when spontaneous nucleation is hard to achieve. In the example presented here, reverse screening has enabled the production of crystals of several peptide complexes with an anti-malaria antibody. PMID- 15299401 TI - New developments of the IMPAX small-volume automated crystallization system. AB - Recent developments of the IMPAX system for automated crystallization are presented. A five-channel microtip has been introduced into the system thereby providing an extra degree of freedom for carrying out experiments. A new mouse driven program for screening has been introduced, which creates a much wider scope for designing and executing screens covering new conditions of crystallization. The hardware has been adapted so that the system can also be used to set up vapour-diffusion trials. A simple design of a vapour-diffusion vessel, suitable for sitting drops of 2-15 microl, using smaller reservoir volumes (up to 100 microl), facilitates large-scale systematic trials. PMID- 15299402 TI - Two distinct approaches to crystallization results-recording databases. AB - Two different systems for recording crystallization results are described. Both programs are written in Microsoft Excel. Results can be entered either as text or numerical scores. In each system, one can search or query on the results, macromolecule name, experimental parameters, etc. and correlate these to specific crystallization conditions or trends in the crystallizations. Such information is valuable when optimizing crystal growth conditions and determining new crystallization parameters to test. XTAL RES is a Macintosh program that is linked to PCRYSTAL (software used to enter details of the crystallization experiments) through an Oracle database on a VAX network. A second system, the Electronic Notebook, was designed as a 'stand-alone' Excel application running under Microsoft Windows. PMID- 15299403 TI - Poly(ethylene) glycol monomethyl ethers - an alternative to poly(ethylene) glycols in protein crystallization. AB - Poly(ethylene) glycol monomethyl ethers (Peg-mmes) are a series of methyl substituted poly(ethylene) glycols that have been used with some success in the crystallization of a number of hydrophobic proteins. Crystallization of a lipase from Humicola lanuginosa complexed with the C12 substrate analogue from Peg-mme 5000, an endoglucanase 1 and a 59 kDa fragment of human topoisomerase IIalpha crystallized from Peg-mme are described. The use of Peg-mme for improving the quality of crystals previously grown from normal poly(ethylene) glycol 8000 is also described. We suggest that these modified Peg-mmes should be regularly used in screening for crystallization. PMID- 15299404 TI - Crystallizing proteins - a rational approach? AB - The advances in recombinant DNA technology in recent years have had a dramatic effect on the area of protein crystallization. Large amounts of pure protein produced in various expression systems have made it possible to conduct experiments that would have been impossible with material from natural sources. With many more laboratories becoming involved in crystallizing proteins a great deal of new information has been generated on techniques to eliminate the so called 'bottleneck of crystallization' in determining a three-dimensional protein structure. More and more new and interesting proteins are being submitted to this laboratory for crystallization. Certain criteria may be set before crystallization trials are started, such as solubility, purity and aggregation tendencies. The introduction of robots now facilitates the screening of crystallization conditions. In cases where no crystals have been obtained after initial screening it can now be decided which possible modifications can be made to the protein itself to improve the chances of obtaining crystals. PMID- 15299405 TI - Time courses of equilibration for ammonium sulfate, sodium chloride and magnesium sulfate heptahydrate in the Z/3 crystallization plate. AB - Time courses of equilibration for three salts, sodium chloride, ammonium sulfate and magnesium sulfate heptahydrate have been measured in the Z/3 crystallization plate. It is shown that by varying both the diffusant and the reservoir depth the time taken to equilibrate can be as short as 200 or as long as 1400 h. Thus, the present design of the plate should accommodate a wide variety of desired crystallization kinetics. PMID- 15299406 TI - Crystallization of macromolecules in silica gels. AB - Procedures are described for the crystallization of proteins, nucleic acids and viruses in a silica-gel matrix using otherwise standard reagents and conditions. Methods are given based on both vapor diffusion in a sitting drop and liquid liquid diffusion. Using a variety of macromolecules our results suggest that the gel matrix suppresses nucleation, reduces the rate of growth, and generally leads to larger, higher-quality crystals of enhanced stability. Presumably these effects arise from the decreased mobility of the macromolecules and their flux at the crystal surface during growth. PMID- 15299407 TI - Investigations on protein crystal growth by the gel acupuncture method. AB - In this work we explore the possibilities of the gel-acupuncture technique, proposed previously for the growth of protein single crystals [Garcia-Ruiz, Moreno, Viedma & Coll (1993). Mater. Res. Bull. 28, 541-546]. The main advantage of the technique is that the crystals are obtained inside an X-ray capillary and, unlike classical microdiffusion techniques, it involves a very simple and accurate technical arrangement that permits the continuous monitoring of the crystals in their growth environment. In particular, we describe the growth of single crystals of lysozyme, concanavalin A and ribonuclease A. Different starting conditions have been used to grow single crystals of these proteins into different types of capillaries at several protein and precipitating-agent concentrations. It is demonstrated that the technique works for a wide range of precipitating agents commonly used in protein crystal growth, such as large polymers (PEG 4000 and PEG 6000), organic solvents (from methanol to butanol) and salts [NaCl, (NH(4))(2)SO(4)]. The range of inner diameter of the capillaries for which the technique works correctly has been also studied. The growth process and possible crystal movement was followed by video microscopy. Lysozyme crystals up to 3.1 mm were obtained but the average maximum linear crystal sizes were 2.0 mm for lysozyme, 0.4 mm for concanavalin A and 1.2 mm for ribonuclease, respectively. The waiting times to reach such a size, measured from the set-up of the experiments, were 72 h, 10 d and 5 d, respectively. Gels of tetramethoxysilane, tetraethoxysilane, sodium silicate, agar, high-strength agar and gel-gro have been tested in relation to their mechanical properties and their chemical interaction with the reactants. Finally, we discuss briefly the advantages of the gel-acupuncture technique and plausible applications other than crystal growth. PMID- 15299408 TI - The use of two novel methods to grow protein crystals by microdialysis and vapor diffusion in an agarose gel. AB - The crystals of most proteins are poorly ordered and diffract to lower resolutions than other crystals of simple and inorganic compounds. The use of two novel methods for gel protein crystal growth, utilizing liquid diffusion and vapor diffusion, are described for the growth of lysozyme and canavalin. Crystallization using gels has been demonstrated to improve crystal quality by reducing convective flow, sedimentation, nucleation and twinning. Preliminary X ray diffraction data are also presented. PMID- 15299409 TI - Study of nucleation-related phenomena in lysozyme solutions. Application to gel growth. AB - Two populations of aggregates are generally indentified in supersaturated solutions of biological macromolecules: small aggregates of a size which is less than 5 nm and large aggregates, the largest of which are at least one order of magnitude bigger. In order to understand the role played by the microporous network of a gel in the growth and behaviour of these different species in the prenucleation period, an in situ observation of nucleation has been carried out using either free solutions or solutions trapped in agarose gels. In a previous study, free solutions were investigated by small-angle neutron scattering (SANS) to identify the small aggregates. Optical observations, made under the same conditions, revealed the formation of an amorphous precipitate which disappeared at the end of the experiment. The sedimentation of this phase, which occurs in free solution but never occurs in gelled solution, depletes the solution bulk and this could explain why the nucleation density is higher in agarose gel than in free solution. The case of silica gel, the behaviour of which is completely different with respect to nucleation, will be discussed. PMID- 15299410 TI - A gel-mediated feeding technique for protein crystal growth from hanging drops. AB - A procedure which allows an investigator to supply a crystal with fresh mother material without inducing significant growth defects is described. This technique requires that the crystal is grown in a gelled hanging or sitting drop. An example concerning a model macromolecule, hen egg-white lysozyme, is given. Extension of this procedure to other macromolecules is discussed. PMID- 15299411 TI - Elimination of twinning in crystals of Sulfolobus sofataricus alcohol dehydrogenase holo-enzyme by growth in agarose gels. AB - Crystals of the binary complex of alcohol dehydrogenase from Sulfolobus solfataricus with NADH were shown to be twinned and not suitable for automated data collection. Several crystallization trials, performed with the aim of eliminating twinning, are described. Interestingly, crystals grown from agarose gel have been demonstrated to have a unique reciprocal lattice. These crystals are monoclinic, space group C2, with cell dimensions a = 134.47 (9), b = 85.26 (5), c = 71.76 (8) A, beta = 97.53 (4) degrees, and showed significant diffraction beyond 3.0 A resolution. PMID- 15299412 TI - Crystallographic studies on p21(H-ras) using the synchrotron Laue method: improvement of crystal quality and monitoring of the GTPase reaction at different time points. AB - The parameters affecting the crystal quality of complexes between p21(H-ras) and caged GTP have been investigated. The use of pure diastereomers of caged GTP complexed to the more stable p21(G12P)' mutant of p21 and the addition of n-octyl beta-D-glucopyranoside improved the reproducibility and decreased the mosaicity of the crystals significantly. Furthermore, the crystallization technique was changed from the batch method to the sitting-drop technique. With the availability of a larger yield of well ordered crystals, it was possible to extend the time-resolved crystallographic investigations on p21(H-ras). A structure of p21(G12P)':GTP could be obtained 2 min after photolytic removal of the cage group and led to the identification of a previously unidentified conformation for the so-called catalytically active loop L4. The refinement of five data sets collected within 2 min at different times (2-4, 11-13, 20-22, 30 32 and 90-92 min) after the initiation of the intrinsic GTPase reaction of the protein indicates that the synchrotron Laue method can be used to detect small structural changes and alternative conformations, but is presently limited in the analysis of larger rearrangements since these produce diffuse and broken electron density. PMID- 15299413 TI - Crystallization and preliminary X-ray structure analysis of thermally unstable p21(H-ras) guanosine complexes. AB - p21 is a small guanine nucleotide binding protein that is involved in intracellular signal transduction. Biochemical data suggest that the presence of the beta-phosphate is essential for strong binding of guanine nucleotides to the protein. Guanosine or GMP bind six orders of magnitude more weakly to p21 than GDP or GTP. Moreover, the thermal stability of the protein is dramatically reduced when bound to GMP or guanosine. We have crystallized C-terminally truncated forms of p21(H-ras), with guanosine or GMP bound, in the space groups P4(3)2(1)2, P2(1)2(1)2 and P2(1). The crystals diffract in the range 2.8-2.2 A. Details of the crystallization procedures, the characterization of the crystals and preliminary results of structure determination are described. An unexpected electron-density peak was found close to the position of the beta-phosphate in the phosphate-binding loop. PMID- 15299414 TI - Crystallization studies of the human immunodeficiency virus (HIV-1) Tat protein and its trans-activation response element (TAR) RNA. AB - Small single crystals are reported of a complex between a small peptide fragment of the HIV-1 Tat protein and a fragment of the RNA to which it binds. Tat is responsible for enhancing the level of expression of the human immunodeficiency virus type 1 (HIV-1) and is a logical target for AIDS therapy. Tat may function to increase the level of transcription initiation or to prevent premature termination of transcripts. In vitro, Tat binds through its basic domain (two Lys and six Arg in nine residues) to a three-nucleotide bulge of a stem-loop RNA structure called TAR. Complex formation between Tat and TAR is necessary for Tat activity. Peptides which contain the basic region of Tat also bind to TAR RNA. We have carried out crystallization experiments on a 27-nucleotide fragment of TAR RNA and on complexes between two Tat peptides and TAR. PMID- 15299415 TI - Crystallization, sequence and preliminary crystallographic data for transmission blocking anti-malaria Fab 4B7 with cyclic peptides from the Pfs25 protein of P. falciparum. AB - X-ray quality crystals of an Fab fragment from a transmission-blocking monoclonal antibody 4B7 (MAb 4B7) against a sexual stage protein Pfs25 of Plasmodium falciparum were grown as uncomplexed and peptide-complexed forms. Initially, the intact immunoglobulin was crystallized because cleavage with pepsin or papain did not produce a homogeneous product. Further proteolytic trials with elastase produced a suitable Fab fragment from which crystals have been obtained, both for the free Fab and in complex with cyclic peptides and in the presence of linear peptides. While linear peptides bind to MAb 4B7, cyclic peptides modeled on a predicted beta-hairpin loop of the third EGF-like domain of Pfs25 bind better and readily co-crystallize with the Fab. The genes for the variable domain of the Fab have been cloned, sequenced and the primary amino-acid sequence for the complete Fab deduced. This work explores the use of glycerol as an additive and the modification of the peptide sequence outside the epitope for improving in the crystallization. Data sets have been collected from crystals of several Fab peptide complexes and from uncomplexed Fab to resolutions ranging from 2.4 to 3.3 A. The packing arrangements of several crystal forms have been determined by molecular replacement, and refinement of their three-dimensional structures is in progress. The three-dimensional structure of this Fab complexed with the various peptides will aid in an understanding of the mode by which this antibody recognizes and prevents transmission of the malaria parasite. PMID- 15299416 TI - Crystallization of the complex between cyclophilin A and cyclosporin derivatives: the use of cross-seeding. AB - With ammonium sulfate as the precipitant, different crystal forms of the complex between cyclosporin A (CsA) and cyclophilin A (CypA) have been crystal- lized [Zurini, Kallen, Mikol, Pflugl, Jansonius & Walkinshaw (1990). FEBS Lett. 276, 63 66]. All have large unit cells and contain a pentamer or a decamer in the asymmetric unit. Using a more water soluble CsA analog, orthorhombic crystals containing only one molecule per asymmetric unit could be grown. They diffract to significantly higher resolution (2.1 A). In this crystal form, CsA has no packing interactions with neighbouring molecules and these crystals could be used to cross-seed other CypA/CsA analog complexes. Nine different CypA/ CsA analog complexes could be crystallized using this technique, most of them yielding highly diffracting crystals, quickly solvable by Fourier difference methods. PMID- 15299417 TI - Crystal growth of human estrogenic 17beta-hydroxysteroid dehydrogenase. AB - Estrogenic 17beta-hydroxysteroid dehydrogenase from human placenta, an enzyme of low solubility, has been crystallized in the complex form with its cofactor NADP(+). These are the first crystals with X-ray diffraction quality for structure analysis from any human steroid-converting enzyme. The crystals were grown by vapor diffusion in the presence of 0.06% beta-octylglucoside, using polyethylene glycol 4000 as the precipitant (27-28%) and one of several different salts at pH 7.5 and room temperature. Crystals grown with magnesium chloride diffract up to 2.4 A. The most important steps leading to the rapid success of the crystallization of this labile enzyme were the following: preparation of a highly active and homogeneous enzyme protein using a rapid procedure; the choice of a suitable enzyme buffer system and a detergent favorable to maintaining high activity and solubility for the enzyme; and a combined screening procedure. The present study could be useful for the successful crystal growth of other hydrophobic or membrane-bound proteins. PMID- 15299418 TI - Crystallization of an intact monoclonal antibody (4B7) against Plasmodium falciparum malaria with peptides from the Pfs25 protein antigen. AB - Monoclonal antibody 4B7 is a neutralizing antibody that binds the protein Pfs25 in the sexual stages of the malaria parasite Plasmodium falciparum and completely blocks transmission of the parasite from human serum to the mosquito host. Here we report the identification of the epitope on Pfs25 recognized by 4B7 and the crystallization of the intact murine monoclonal antibody with peptides corresponding to that epitope. This study highlights the importance of ligands in the crystallization of proteins. In this case peptides have been used to modulate the solubility of the peptide-IgG complex and may have provided different or additional crystal contacts to create or enhance a crystalline reticulum. Multiple crystal forms characterize this crystallization and the various peptides, differing both in length and sequence, have been used to investigate how such changes affect nucleation and crystal growth. PMID- 15299419 TI - Extension of the diffraction resolution of crystals. AB - The diffraction resolution of crystals of the guanine nucleotide-exchange factor complex, EF-Tu-Ts, has been extended from 5.0 to 2.5 A by lowering the solvent content in the crystals as well as the temperature of data collection. The common form of EF-Tu-Ts crystal belongs to space group P2(1)2(1)2(1) with a = 81.1, b = 109.9, c = 207.5 A and has a solvent content of 61%. The crystals diffract to a resolution of 5.0 A at 293 K and 4.0 A at 273 K. When cryoprotective agents are slowly diffused into the crystals, the cell constants shrink to a = 74.4, b = 109.9, c = 198.7 A and the solvent content falls to 55%. After the cryoprotective agent has been added, the crystals diffract to 2.7 A resolution at 293 or 273 K and 2.5 A at 250 K. X-ray diffraction data, collected before and after the transformation of individual EF-Tu-Ts crystals, demonstrate that a large percentage of the improvement in diffraction resolution is due solely to the addition of cryoprotective agents. The transfer procedures for the successful introduction of cryoprotective agents into EF-Tu-Ts crystals as well as the general applicability to other crystal systems will be discussed. PMID- 15299420 TI - Mesoscopic theories for protein crystal growth. AB - A computer-simulation method is proposed for studying the hydrodynamic interactions of rigid protein molecules. It is a combination of Stokes dynamics and continuum hydrodynamics. The Stokes equations of motion for the protein molecules, the creeping-flow equation for the solvent together with the no-slip boundary conditions give a complete representation of the system. The resulting three-dimensional boundary-value problem can be rewritten in a two-dimensional form (without any loss of information) considering the surfaces of the particles only. Then, by solving the equations on discrete surface elements, the so-called mobility matrix is determined in which all hydrodynamic interactions are included. Finally, after calculation of the conservative forces and the stochastic force, the new velocities of the protein molecules can be determined. The simulation method can be applied to arbitrary particle shapes. It can also handle arbitrary flow fields, and the effects of applying a flow field to the system can be studied. From analysis of the trajectories, information can be gained on the kinetics and thermodynamics in the early stages of the crystallization process. PMID- 15299421 TI - Quantitative analysis in the characterization and optimization of protein crystal growth. AB - Protein crystal growth often depends on the combination of many different factors. Some affect protein solubility directly; others may act indirectly by causing conformational changes. Systematic characterization of these factors can be important for generating good crystals. It can also provide useful insight into the biochemical behavior of the protein being crystallized. Here we focus on statistical methods to achieve these two objectives. (1) Characterization of a protein system by analyzing patterns of crystal polymorphism under different levels of biochemical parameters, such as ligands and pH. Tests of the reproducibility of crystal growth experiments indicate that quantitative scales of crystal quality can be statistically significant. Analysis of variance for a replicated, full-factorial design in which four factors were tested at two levels has been used to demonstrate highly significant, biochemically relevant, two factor interactions strongly implicating pH and ligand-dependent conformational changes. (2) Optimization of crystal growth via response-surface methods. 'Minimum predicted variance' designs provide for efficient response-surface experiments aimed at constructing quadratic models in several dimensions. We have used such models to improve crystal size and quality significantly for three forms of Bacillus stearothermophilus tryptophanyl-tRNA synthetase. In one case we can now avoid having to increase the size by repeated seeding, a difficult procedure that also produces unwanted growth of satellite crystals. Graphs of two dimensional level surfaces reveal a number of ridges, where the same result is obtained for many combinations of the factors usually varied when trying to improve crystals. An important inference is that it may be better to sample simultaneously for the effects of protein concentration and supersaturation. For a system involving only one crystallizing agent, supersaturation can be approximated as the product of protein and precipitant concentrations. Use of this search direction significantly improves the performance of response-surface experiments. Advantages of growing crystals at stationary points of their response surfaces include better crystals and higher reproducibility, since crystal growth at stationary points is insulated from the deleterious effects of experimental fluctuations. This arises because the derivatives of the response are by definition zero with respect to the experimental variables. Quantitative analysis of appropriately designed crystal growth experiments can thus be a powerful way to characterize complex and interacting biochemical dependencies in macromolecular systems and optimize parameters important to the crystallography. PMID- 15299422 TI - Internal symmetry of the molecular chaperone cpn60 (GroEL) determined by X-ray crystallography. AB - The Escherichia coli molecular chaperone cpn60 oligomer, [cpn60](14), also called GroEL, has been crystallized and examined by X-ray crystallography and self rotation function calculations. The crystals show unit-cell dimensions a = 143.3, b = 154.6 and c = 265 A, with alpha = 82, beta = 95 and gamma = 107 degrees. The space group is P1 and crystals diffract to 7 A. X-ray analysis shows that the oligomer has one sevenfold symmetry axis and seven twofold axes that are all perpendicular to the sevenfold. The symmetry suggests that [cpn60](24) consists of two heptamers, [cpn60](7), stacked on top of each other. The orientations of the symmetry axes of the two independent [cpn60](14) oligomers in the triclinic unit cell have been determined relative to the crystallographic axes. The two oligomers in the unit cell are arranged side-by- side, but the second oligomer is rotated 26 degrees around the sevenfold axis relative to the first oligomer. PMID- 15299423 TI - Crystallization and X-ray structure determination of cytochrome c2 from Rhodobacter sphaeroides in three crystal forms. AB - Cytochrome c(2) serves as the secondary electron donor that reduces the photo oxidized bacteriochlorophyll dimer in photosynthetic bacteria. Cytochrome c(2) from Rhodobacter sphaeroides has been crystallized in three different forms. At high ionic strength, crystals of a hexagonal space group (P6(1)22) were obtained, while at low ionic strength, triclinic (P1) and tetragonal (P4(1)2(1)2) crystals were formed. The three-dimensional structures of the cytochrome in all three crystal forms have been determined by X-ray diffraction at resolutions of 2.20 A (hexagonal), 1.95 A, (triclinic) and 1.53 A (tetragonal). The most significant difference observed was the binding of an imidazole molecule to the iron atom of the heme group in the hexagonal structure. This binding displaces the sulfur atom of Met l00, which forms the axial ligand in the triclinic and tetragonal structures. PMID- 15299424 TI - Observation of growth steps, spiral dislocations and molecular packing on the surface of lysozyme crystals with the atomic force microscope. AB - The (110) faces of lysozyme crystals in their mother liquor have been investigated using an atomic force microscope (AFM) in height mode. Crystal growth and dissolution steps, as well as simultaneous growth and dissolution in pits, have been observed. Screw dislocations were also observed but the fine structure has not yet been investigated. Images that may possess molecular resolution were obtained and compared with theoretical images based on the crystallographic structure and the effects of arbitrary tip profiles. Crystallographic periodicities of 38 and 112 A were observed. A recurring feature is a centered periodic array of minima that may be associated with one of the two nearly planar sheets of molecules present in the crystal that are parallel to the (110) faces. PMID- 15299425 TI - Studies on tetragonal lysozyme crystal growth rates. AB - A computer-controlled apparatus able to simultaneously follow the face growth rate of up to 40 crystals was developed. This apparatus was used to investigate the effects of solution pH on the (110) and (101) face growth rates of tetragonal lysozyme. Growth rates were measured at pH 4.0, 4.4, 4.8 and 5.2, in 0.1 M sodium acetate buffer with 5% NaCl, 295 K. Initial crystal sizes ranged from 10 to 40 microm. Plots of log supersaturation ratio (either C/C(sat) or C/C(sat) - 1) versus log(growth rate) are not linear, typically having a slope of approximately 8 at the lowest growth rates determined (10(-6) microm s(-1)), which falls off to a slope of approximately 2 at the highest growth rates (10(-2) microm s(-1)) measured. Ratios of C/C(sat) ranged from 4 to >20. The data show that lower solubility solutions require higher supersaturation ratios for equivalent growth rates. Data for the growth rate of the (101) face at pH 4.0 were widely scattered, especially at lower supersaturation ratios. Time-lapse video of crystals at low supersaturations shows that initially only the (110) faces grow, leaving 'notches' at the (110)-(110) corners. These corners then fill in and macro-steps appear on the (101) faces which rapidly move inward in the form of an octagon, restoring the crystal to a 'normal' appearance. This phenomenon has been observed for tetragonal crystals grown in either still or flowing solutions. Flowing solutions at lower supersaturations also gave cases where the corners did not fill in, with the (110) faces continuing to grow out until growth ceased. PMID- 15299426 TI - Acid pH crystallization of the basic protein lysin from the spermatozoa of red abalone (Haliotis rufescens). AB - A new crystal form of dimeric red lysin, a distinctly basic protein (M(r) = 16 070) from the red abalone (Haliotis rufescens), has been obtained using ammonium sulfate as precipitant with a sodium citrate-boric acid-citric acid buffer at pH 4.5. The acid pH crystal form resulted from a study aimed at developing conditions favorable to the sitting-drop vapor-diffusion crystallization of other abalone lysins which do not crystallize at neutral or basic pH conditions. The space group is P222(1) with cell dimensions a = 51.2, b = 47.0, c = 123.8 A and two molecules per asymmetric unit. PMID- 15299427 TI - Crystallization of seed globulins from legumes. AB - Seeds contain large quantities of proteins and are therefore main food sources. In the last century, protein extracts of legume seeds were dialysed against distilled water and in some cases small crystals of pure protein appeared. However, those crystals were generally of poor quality with respect to X-ray diffraction. Recently, the crystallization of some of them was improved and the structures of two 7S globulins, phaseolin from Phaseolus vulgaris and canavalin from Canavalia ensiformis, have been determined at 3.0 and 2.6 A resolution, respectively. Efforts to improve the quality of the phaseolin crystals resulted in three new crystal forms which will be discussed in this paper. The only high resolution X-ray analysis of a seed globulin from legumes is that of narbonin, a 2S protein from Vicia narbonensis. The crystal structure at 1.8 A shows a very compact packing in layers of molecules. The intermolecular contacts include salt bridges and hydrophobic clusters that might facilitate both the aggregation of the molecules and their crystallization. Because the seed globulins appear in large quantities in the protein bodies of the seeds, efficient packing of the molecules similar to the crystal packing can be assumed. PMID- 15299428 TI - In situ two-dimensional crystallization of a polytopic membrane protein: the cardiac gap junction channel. AB - In situ crystallization of rat ventricular gap junctions was accomplished by sequential dialysis of membranes against low concentrations of deoxycholate and dodecyl-beta-D-maltoside. Lipids are removed without solubilizing the protein, and the increased protein concentration in the membrane plane facilitates two dimensional crystallization in the native membrane environment. The two dimensional crystals have a nominal resolution of 16 A and display plane group symmetry p6 with a = b = 85 A and gamma = 120 degrees. Electron crystallography reveals that the cardiac gap junction membrane channel is formed by a hexameric cluster of protein subunits, and this hexameric design appears to be a recurring quaternary motif for the multigene family of gap junction proteins. Exposure of membranes to low concentrations of detergents may provide an approach for in situ two-dimensional crystallization of other connexins as well as other membrane proteins, especially those that are labile when solubilized as protein-detergent micelles. PMID- 15299429 TI - Phase diagram determination to elucidate the crystal growth of the photoreaction center from Rhodobacter sphaeroides. AB - Solubility curves (a phase diagram) of an amorphous precipitate and the orthorhombic crystals of a membrane protein, the photoreaction center from Rhodobacter sphaeroides, were determined. With the phase diagram, the process of crystal growth of the membrane protein was elucidated within a general physicochemical framework, and justification was provided of hitherto empirically selected crystallization conditions. PMID- 15299430 TI - Large single crystals of the Neurospora crassa plasma membrane H+-ATPase: an approach to the crystallization of integral membrane proteins. AB - Large single crystals of the dodecylmaltoside (DDM) complex of a polytopic integral membrane transport protein, the Neurospora plasma membrane H(+)-ATPase, have been obtained using an approach that attempts to take into account the possibly radically different physicochemical properties of the protein surfaces and the detergent micellar collar. The overall goal of the crystallization strategy employed was to identify conditions in which the protein surfaces of the DDM-ATPase complex are moderately insoluble and in which the DDM micellar collar is also near its solubility limit. The first step was to screen a variety of commonly used protein precipitants for those that were able to induce the aggregation of pure DDM micelles. The concentration at which any precipitant induced DDM micellar aggregation was hoped to be close to the concentration at which it might induce insolubility of the detergent micellar collar of the DDM ATPase complex. Of the nine precipitants tried, seven, all polyethylene glycols (PEGs), were able to induce DDM micelle insolubility. The seven PEGs were then tested for their effect on the solubility of the DDM-ATPase complex at a concentration slightly below that necessary to induce DDM micellar aggregation. Three of the PEGs caused extensive precipitation of the ATPase at this concentration and were, therefore, shelved. The other four PEGs did not induce precipitation at the concentration employed and were subsequently used at this concentration for crystallization trials in which the protein concentration was varied. Encouragingly, crystalline plates of the ATPase were obtained for each of the four PEGs tried, indicating that the overall approach may be valid. Unfortunately, the crystals obtained were visibly flawed, suggesting that the correct balance of protein surface and DDM micelle insolubility had not yet been reached. The ionic strength of the crystallization trials was then raised, which was known from other experiments to render the protein surfaces of the ATPase less soluble while having no effect on the DDM micellar aggregation point. For one of the PEGs, PEG 4000, this brought on a new, well formed hexagonal crystal habit. Subsequent optimization of the initial conditions has yielded large single hexagonal crystals of the H(+)-ATPase roughly 0.4 x 0.4 x 0.15 mm in size, holding promise for exploration of the structure of the ATPase by X-ray diffraction analysis. PMID- 15299431 TI - Packing selection of Bacillus lentus subtilisin and a site-specific variant. AB - The crystallization of a variant of Bacillus lentus subtilisin and the native enzyme was achieved using identical conditions. The variant B. lentus was found to grow in two crystal forms, form 1 and form 2, whereas the native B. lentus subtilisin enzyme crystallized in only one, form 1. Form 2 crystals, once obtained, were found to grow much more rapidly than form 1 crystals. The lattice contacts and structural changes giving both crystal forms have been examined. The results show that crystal form 2 has a more complex network of interactions. There is also a small surface conformational change in the form 2 structure relative to the native and variant form 1 crystals and at least two solvent molecules bound to the enzyme in crystal form 1 are displaced in crystal form 2. In addition, a site specific substitution in the variant at position 27 induces a 'short' lattice contact which does not exist in the native B. lentus or the form 2 variant B. lentus. These results suggest that in some circumstances engineered variants could be designed to crystallize more rapidly than the native enzyme. PMID- 15299432 TI - Intermolecular contacts in various crystal forms related to the open and closed conformational states of the catalytic subunit of cAMP-dependent protein kinase. AB - All hitherto solved crystal structures of the catalytic (C) subunit of cAMP dependent protein kinase can be classified into two groups, those with a closed and those with an open conformation of the ATP-binding lobe. The molecules with the closed conformation are all related by a crystallographic 2(1) axis that connects them into an infinite-chain motif. The motif has only one large contact region that involves many residues, several of them in the ATP-binding lobe, embedded in an extensive network of water molecules. The dominant feature of this region is the hydrophobic interaction between Trp196 and Arg133, Arg134. This motif has been found so far in three different crystal forms, two correspond to ternary enzyme-inhibitor-ATP complexes with mammalian and recombinant C, and one to a binary enzyme-inhibitor complex with recombinant C. The open conformation has been found in two closely related crystal structures, both of cubic symmetry, of the apoenzyme and a binary complex of the mammalian catalytic subunit. In this cubic structure of the binary complex, the hydrogen-bonded intramolecular contacts between Arg18 of the inhibitor and the ATP-binding lobe of the binary and ternary complexes of the recombinant enzyme are missing due to a strong hydrophobic intermolecular contact involving the diiodinated Tyr7. In solution, no crystal contacts prevent these hydrogen bonds involving Arg18 from forming so that it is likely that the binary complex with Tyr7 of the peptide inhibitor iodinated or not, can assume the closed conformation in solution. While the closed structure very likely represents a stable conformation in solution, there is no evidence to suggest that the open conformation represents a unique stable conformational state of the enzyme in solution. PMID- 15299433 TI - Water structure associated with proteins and its role in crystallization. AB - X-ray or neutron diffraction studies have shown, at the atomic level, that water molecules occupy well determined sites inside or at the surface of biological macromolecules. These water molecules are constitutive of biomolecules and play a crucial role in their structural and functional properties. Upon crystallization some water molecules are either desolvated or involved in crystal packing. The role of water in determining crystal packing has been experimentally confirmed by several X-ray analyses. PMID- 15299434 TI - An evaluation of the use of databases in protein structure refinement. AB - The speed of electron-density fitting during X-ray structure solution and refinement, and the quality of the protein model resulting, can both be enhanced by the use of databases of main- and side-chain conformations. Three structures are compared in this report, one refined at high resolution (1.7 A), and two at lower resolutions using either the database method (2.4 A resolution) or more traditional empirical electron-density fitting (1.9 A resolution). An analysis of peptide orientation was used as an aid in finding unusual portions of main-chain structure. The fit of side chains to known rotamer conformations was used to help determine the accuracy of these atomic positions. In addition, the use of an objective measure of the fit of structures to electron-density maps was evaluated, both alone and in combination with side-chain conformational information. PMID- 15299435 TI - X-ray structure of a monoclinic form of hen egg-white lysozyme crystallized at 313 K. Comparison of two independent molecules. AB - A monoclinic crystal of hen egg lysozyme (HEL, E.C. 3.2.1.17) was obtained at 313 K from a 10%(w/v) NaCl solution at pH 7.6 containing 5%(v/v) 1-propanol. Cell dimensions were a = 27.23, b = 63.66, c = 59.12 A and beta = 92.9 degrees, and the space group was P2(1). The unit cell contains four molecules (V(m) = 1.79 A(3) Da(-1)). The structure was solved by the isomorphous replacement method with anomalous scattering followed by phase improvement by the solvent-flattening method. The refinement of the structure was carried out by the simulated annealing method. The conventional R value was 0.187 for 18 260 reflections [|F(o)| > 3sigma(F)] in the resolution range 10-1.72 A. The r.m.s. deviations from the ideal bond distances and angles were 0.015 A and 3.0 degrees, respectively. The two molecules in the asymmetric unit are related by a translation of half a lattice unit along the a and c axes. The r.m.s. difference of equivalent C(alpha) atoms between the two molecules was 0.64 A and the largest difference was 3.57 A for Gly71. A significant structural change was observed in the regions of residues 45-50, 65-73 and 100-104. The residues 45-50, which connect two beta-strands, are shifted parallel to the beta-sheet plane between the two molecules. The residues 100-104 belong to the substrate-binding site (subsite A) and the high flexibility of this region may be responsible for the binding of the substrate and the release of reaction products. PMID- 15299436 TI - Solid-state phase transition in the crystal structure of ribulose 1,5 bisphosphate carboxylase/oxygenase. AB - The crystal structure is described of ribulose 1,5-bisphosphate carboxylase/oxygenase in a new crystal form. This new form (form V) was obtained from a previously known crystal form (form III) through a solid-state phase transition. The solid-state phase transition was brought about by transferring the crystal from a high-salt low-pH mother liquor to a low-salt high-pH synthetic mother liquor. The interplay of electrostatic repulsion and osmotic pressure induced a unit-cell shrinkage of 24 A along the c axis and expansion of 4 A along the a and b axes. The space group also changed from I422 to I4. The new crystal form was shown to be more resistant to X-ray radiation damage, which suggests the effect of crystal stabilization by non-penetrating molecules. The structure of ribulose 1,5-bisphosphate carboxylase/oxygenase in the new crystal form is compared with that of the old crystal form. PMID- 15299437 TI - Metal substitution in a blue-copper protein: the crystal structure of cadmium azurin at 1.8 A resolution. AB - Crystals of cadmium-substituted azurin have been prepared by diffusing Cd(II) into crystals of apo-azurin grown previously and their structure has been determined at high resolution by X-ray crystallography. Data to 1.8 A resolution were collected by Weissenberg photography (with image plates) using synchrotron radiation. These data were combined with a 2.2 A diffractometer data set to give 90% coverage to 1.8 A. An initial model was derived from the isomorphous Cu(II) azurin structure, and the cadmium and ligand positions added from 'omit' maps. Refinement was by restrained least squares (program PROLSQ), to a final R value of 0.168 for all data in the range 10.0-1.8 A (23 349 reflections). The final model of 1954 protein atoms, two Cd(II) ions (occupancy 0.75), four SO(4)(2-) ions and 239 water molecules has r.m.s. deviations of 0.015, 0.045 and 0.013 A from standard bond lengths, angle distances and planar groups. The protein structure is essentially the same as that of Cu(II)-azurin, with an r.m.s. deviation of 0.18 A for 97% of main-chain atoms after superposition of the two structures. The Cd atom is within 0.2 A of the equivalent copper position, displaced slightly away from the axial Met ligand towards the carbonyl O atom of Gly45. The latter has also moved slightly towards the metal, by a rotation of the peptide unit, to give a Cd-O bond of 2.76 A. The Cd-S(Cys) bond is lengthened to 2.39 A. The coordination geometry is slightly more tetrahedral than for Cu(II), and the cadmium-oxygen interaction is consistent with the presence of an oxygen ligand in the coordination sphere of stellacyanin. PMID- 15299438 TI - Application of an automatic molecular-replacement procedure to crystal structure analysis of cytochrome c2 from Rhodopseudomonas viridis. AB - An automatic molecular-replacement procedure has been applied to solve the crystal structure of cytochrome c(2) from Rhodopseudomonas viridis. The structure was solved on the basis of the structure of tuna cytochrome c as a search model using an automatic processing program system, AUTOMR. The refinements by molecular dynamics and restrained least-squares methods result in a current crystallographic R factor of 0.219 for diffraction data at 3 A resolution. PMID- 15299439 TI - Data collection at short wavelengths in protein crystallography. AB - The development of high-intensity X-ray sources and the use of insertion devices will make it possible to collect data routinely from protein crystals at very short wavelengths (lambda sigma(Fo)) between 10.0 and 1.8 A resolution. The final Rfactor was 17.0%, and the standard deviation of atomic position estimated by Luzzati plot [Luzzati (1952). Acta Cryst. 5, 802-810] was 0.2 A. The electron-density map was well defined for the two independent molecules except for the N-terminal residue and the three C-terminal residues. Equivalent Calpha atoms of two independent molecules in the asymmetric unit were superposed by the least-squares method with root-mean-square deviations of 0.26 A. Reasonable structural differences were observed at a polypeptide segment having few intramolecular interactions. Highly flexible regions of the molecule were assigned from the structural differences between the two independent molecules in the crystal and the distribution of temperature factors along the polypeptide chain. PMID- 15299455 TI - Crystallization of biosynthetic arginine decarboxylase from Escherichia coli. AB - Putrescine is the immediate precursor for the synthesis of polyamines and is normally generated by the action of ornithine decarboxylase. However, putrescine can also be produced by the conversion of arginine to agmatine by arginine decarboxylase (bADC) followed by the release of urea by agmatine ureohydrolase. Amino-acid sequence homology with the eukaryotic ornithine decarboxylases suggests that bADC may be a model for this group of decarboxylases. We report here the crystallization of arginine decarboxylase from E. coli. Crystals up to 1 mm in size are grown by vapor equilibration using Li(2)SO(4) and polyethylene glycols as precipitants. The crystals exhibit diffraction maxima beyond 3 A resolution and belong to space group P4(1(3))2(1)2 with a = 192.4 and c = 121.0 A. These unit-cell dimensions together with the estimated density of the crystals suggest the presence of one tetramer of bADC (71 kDa subunit(-1)) per asymmetric unit (V(m) = 2.0 A(3) Da(-1)). PMID- 15299456 TI - Detection, delineation, measurement and display of cavities in macromolecular structures. AB - A computer program, VOIDOO, is described which can be employed in the study of cavities such as they occur in macromolecular structures (in particular, in proteins). The program can be used to detect unknown cavities or to delineate known cavities, either of which may be connected to the outside of the molecule or molecular assembly under study. Optionally, output files can be requested that contain a description of the shape of the cavity which can be displayed by the crystallographic modelling program O. Additionally, VOIDOO can be used to calculate the volume of a molecule and to create a file containing data pertaining to the surface of the molecule which can also be displayed using O. Examples of the use of VOIDOO are given for P2 myelin protein, cellular retinol binding protein and cellobiohydrolase II. Finally, operational definitions to discern different types of cavity are introduced and guidelines for assessing the accuracy and improving the comparability of cavity calculations are given. PMID- 15299457 TI - Ta6Br14 is a useful cluster compound for isomorphous replacement in protein crystallography. AB - The metal cluster Ta(6)Br(14) has been used to prepare heavy-metal derivatives of two large proteins, ribulose-1,5-bisphosphate carboxylase/oxygenase and transketolase. In both cases, this cluster compound produced a single-site derivative for which a difference Patterson map, calculated to 5.5 A resolution, could be solved straightforwardly. Ta(6)Br(14) provided enough phase information to unambiguously locate the heavy-atom positions in other multiple-site derivatives. In transketolase, the heavy-metal complex binds at the surface of the protein in a dominantly hydrophobic pocket. In ribulose bisphosphate carboxylase/oxygenase, it binds between two molecules in the crystal lattice. There are negatively charged glutamic and/or aspartic acid residues in the vicinity of the bound clusters. Ta(6)Br(14) is useful over a wide range of pH. For large proteins and/or large unit cells, this compound should be included in the initial screening for heavy-metal derivatives. PMID- 15299458 TI - Space-group degeneracy in the packing of a non-selfcomplementary Z-DNA hexamer. AB - The X-ray diffraction pattern of the crystals of the non-selfcomplementary hexadeoxyribonucleotide d(CGCACG).d(CGTGCG) can be indexed in four different space groups: (i) P6(5) and P2(1), with cell parameters a = 17.75 (1), b = 17.76 (1), c = 42.77 (3) A, alpha = 90, beta = 90, gamma = 120 degrees, and (ii) P2(1)2(1)2(1) and C2, with cell parameters a = 17.75 (1), b = 30.74 (2), c = 42.77 (3) A, alpha = 90, beta = 90, gamma = 90 degrees. While the R(merge) for the equivalent reflections in the different space groups indicates that P2(1) is the correct choice in the present case, it is demonstrated that the near degeneracy of the space groups arises out of the fact that the DNA molecule is nearly cylindrical. A perfect cylinder would show perfect degeneracy. PMID- 15299459 TI - Initial crystallographic analysis of a recombinant human interleukin-1 receptor antagonist protein. AB - We report the crystallization of samples of a recombinant preparation of human interleukin-1 receptor antagonist protein (IRAP) and solution of the crystal structure by isomorphous replacement methods. Crystals were obtained by the hanging-drop vapor-diffusion method at 277 K from solutions of PEG 4000 containing sodium chloride, dithiothreitol and PIPES [sodium piperazione-N,N' bis(2-ethanesulfonate)] buffer at pH 7.0. Crystals appear within about a week and grow as truncated tetragonal bipyramids to 0.3-0.6 mm on an edge. X-ray diffraction data from these crystals specify space group P4(3)2(1)2 and unit-cell dimensions of a = b = 72.35(26), c = 114.7(8) A and Z = 16 (two molecules per asymmetric unit). Fresh crystals diffract to about 2.3 A resolution. The search for heavy-atom derivatives has produced two, potassium gold cyanide and trimethyl lead chloride, as same-site, single-site derivatives. Inspection of an electron density map at 4 A resolution calculated with these derivatives confirms that the IRAP molecule is a member of the interleukin-1 structural family. PMID- 15299460 TI - Structure of the ADP complex of the 3-phosphoglycerate kinase from Bacillus stearothermophilus at 1.65 A. AB - The structure of the ADP complex of the enzyme 3-phosphoglycerate kinase (PGK, E.C. 2.7.2.3) from Bacillus stearothermophilus NCA-1503 has been determined by the method of molecular replacement. The structure has been refined to an R factor of 0.16 for all data between 10.0 and 1.65 A resolution, using data collected on the Hendrix-Lentfer imaging plate at the EMBL outstation in Hamburg. The r.m.s. deviations from stereochemical ideality are 0.010 and 0.011 A for bonds and planes, respectively. Although crystallized in the presence of the nucleotide product MgATP, the high-resolution structure reveals the bound nucleotide to be MgADP reflecting the low intrinsic ATPase activity of PGK. Although the two domains of this enzyme are found to be some 4.5 degrees closer together than is found in the yeast and horse-muscle apo-enzyme structures, this structure represents the 'open' rather than the 'closed', catalytically competent form, of the enzyme. PMID- 15299461 TI - Analysis of diffuse scattering from yeast initiator tRNA crystals. AB - Yeast initiator tRNA crystals exhibit strong X-ray diffuse scattering. This scattering can be used to extract information about lattice-coupled and intramolecular motions in the crystals. The amplitudes and correlation distances of these motions can be estimated by calculating the diffuse scattering and comparing the results with the observed scattering. Results indicate that both anisotropic, lattice-coupled motions as well as short-range correlated local disorder in the anticodon arm contribute to the overall disorder in the crystals. These types of motions can be correlated with aspects of tRNA function. This additional information complements the results from analysis of crystallographic data and provides a more detailed picture of the structure and dynamics of the molecule. The degree to which the methodology presented here can account for the observed diffuse scattering from tRNA represents a significant step forward in the ability to use this conventionally discarded information, and encourages the ultimate extension of these ideas to a wide variety of macromolecular systems. PMID- 15299462 TI - New crystal forms of a micro-class glutathione S-transferase from rat liver. AB - Two new crystal forms of isoenzyme 3-3 of rat liver glutathione S-transferase (GST 3-3) have been obtained. They were grown under essentially the same crystallization conditions as those reported for the C2 crystal form [Fu, Rose, Chung, Tam & Wang (1991). Acta Cryst. B47, 813-814]. The new crystals belong to space group P2(1) with one form having cell dimensions a = 101.6, b = 69.5, c = 81.4 A, and beta = 113.6 degrees, and the other form having cell parameters a = 97.4, b = 81.1, c = 69.4 A and beta = 109.2 degrees. These new crystals diffract to at least 2.5 A, resolution. The molecular packing arrangements in these P2(1) crystals have been found by molecular replacement studies. PMID- 15299463 TI - Crystallization and preliminary X-ray analysis of a lipase from Chromobacterium viscosum. AB - Lipase from Chromobacterium viscosum has been purified to homogeneity and crystallized in a form suitable for X-ray diffraction analysis from 10-14% polyethylene glycol 4000 and 10-14% 2-methyl-2,4-pentane diol at pH 6.4 in the presence of 0.25%(w/v) n-octyl-beta-D-glucopyranoside. These crystals belong to space group P2(1)2(1)2 with refined lattice constants a = 41.1 A, b= 156.8, c = 43.6 A, indicating a cell content of one monomer per asymmetric unit of the crystal. The crystals diffract to a resolution of 2.2 A. PMID- 15299464 TI - A designed mutant of the enzyme glutathione reductase shortens the crystallization time by a factor of forty. AB - The packing of glutathione reductase from Escherichia coli in crystal form T showed a place where two molecules are at a distance of only 6 A between the closest atoms, i.e. where a contact is almost made. In order to form this contact with hydrogen bonds, two amino-acid residues were exchanged. This mutation had no effect on molecular packing or the resolution limit of the X-ray diffraction, but facilitated crystal nucleation dramatically and possibly increased the crystal growth rate and shortened the crystallization time. PMID- 15299465 TI - Crystallization and preliminary investigation of xylose isomerase from Bacillus coagulans. AB - Xylose isomerase from Bacillus coagulans has been crystallized in two different crystal forms. One crystal form is in space group P2(1)2(1)2, cell dimensions a = 462, b = 165, c = 82 A. The other is in space group I422, cell dimensions a = b = 113, c = 153 A. PMID- 15299466 TI - Crystallization and preliminary X-ray diffraction analysis of tetra-heme cytochrome c3 from sulfate- and nitrate-reducing Desulfovibrio desulfuricans ATCC 27774. AB - Crystals of the tetra-heme cytochrome c(3) (M(r) = 13 kDa, 107 residues, four heme groups) from sulfate- and nitrate-reducing Desulfovibrio desulfuricans ATCC 27774 have been obtained and crystallographically characterized. They belong to space group P6(1)22 with cell dimensions a = b = 61.84 (4) and c = 109.7 (2) A, and Z = 12. Intensity data were initially collected on a FAST system with a rotating-anode X-ray source leading to a total of 22 592 observations, from which only 4930 were unique, in the resolution range 20.0-2.4 A with an R(merge)(I) of 7.0%. Higher resolution data were measured on a FAST system at station 9.6 of the SRS (Daresbury, England), leading to 19 328 intensities, of which 11 179 were unique, in the resolution range 20.0-1.75 A and an R(merge)(I) of 5.5%. Cross rotation and translation functions were performed with ALMN and TFSGEN programs from the CCP4 suite. The packing of the molecules in the unit cell was checked with TOM/FRODO. Rigid-body refinement of the model and subsequent refinement using molecular dynamics were performed with X-PLOR, leading to a current R factor of 25.9%, for data up to 2.3 A. PMID- 15299470 TI - On the application of one-wavelength anomalous scattering. IV. The absolute configuration of the anomalous scatterers. AB - An essential first step in most techniques for using anomalous-scattering data for phase determination is to determine the positions of the anomalous scatterers. This is usually done by use of the anomalous differences, either as input to a direct-methods procedure or to produce a Patterson map. If the arrangement of anomalous scatterers is noncentrosymmetric then it is also necessary to find their absolute configuration and a process is described for doing this based on the properties of the P(s) function [Okaya, Saito & Pepinsky (1955). Phys. Rev. 98, 1857-1858]. If the arrangement of anomalous scatterers is centrosymmetric then the problem does not occur. PMID- 15299471 TI - MAD phasing: Bayesian estimates of F(A). AB - A Bayesian approach is applied to the calculation of Patterson functions and cross-Fourier maps in the analysis of multi-wavelength anomalous-diffraction (MAD) data. This procedure explicitly incorporates information available a priori on the likely magnitudes of partial structure factors (F(A)) corresponding to the anomalously scattering atoms, uses weighted-average estimates of F(A), and incorporates estimates of errors in the data that are not represented in the instrumental uncertainties. The method is demonstrated by application to MAD data collected on selenomethionine-containing gene V protein. PMID- 15299472 TI - MAD phasing: treatment of dispersive differences as isomorphous replacement information. AB - A framework analogous to that used for the analysis of data in the method of isomorphous replacement is applied to the analysis of multi-wavelength anomalous diffraction (MAD) data. The present method is advantageous in that refinement of parameters describing the anomalously scattering atoms in the structure can be carried out using nearly all the data, and in that phase information can be readily combined. The procedure described here involves an approximation in which it is assumed that the magnitude of the structure factor corresponding to anomalously scattering atoms in the unit cell is small compared to that corresponding to all other atoms in the structure. A model calculation is applied to a protein crystal with 682 non-H atoms in the asymmetric unit and two Se atoms as the anomalous scatterers. It is shown using this model calculation that the approximation used in this analysis does not substantially affect the accuracy of phase calculations for this MAD data. The method is demonstrated by application to MAD data collected on gene V protein. PMID- 15299473 TI - Cross-validation tests of time-averaged molecular dynamics refinements for determination of protein structures by X-ray crystallography. AB - Time-averaged structure-factor restraints have been used in two molecular dynamics refinement schemes to define ensembles of conformations for myoglobin that fit the experimentally measured Bragg scattering from P6 crystals. The geometries of the structures have been maintained to the same currently acceptable limits in all cases. Free R value analysis was used to assess the validity of the two approaches. In the first scheme, where atoms have no B values, the decrease in R value was found to be spurious as judged by a concomitant increase in the free R value. The other scheme, however, which retains individual B values, was found to yield both low R values and low free R values; thus, here the additional variables introduced by modeling the protein in terms of an evolving ensemble of states do not overfit the data. For comparison, refinements were also carried out on the system using several other techniques for isotropic and anisotropic crystallographic refinement. The time-averaged refinements with B values compare quite favorably with the standard methods, but yield additional information about substates of the system. Hence, correctly applied time-averaged refinements can yield accurate models for protein molecules; moreover, by essentially relaxing the harmonic approximation from the refinement process, these refinements allow a more detailed description of the motions of complex molecules, such as proteins, to be determined from X-ray crystallographic data. PMID- 15299474 TI - Crystallization and preliminary crystallographic data for the azurin mutant End 121 from Pseudomonas aeruginosa. AB - Pseudomonas aeruginosa azurin has been crystallized from a mutant where residues from Met 121 to Lys128 have been deleted from the protein. The crystals form pale blue well formed prisms in the orthorhombic space group P2(1)2(1)2(1), with cell dimensions a = 60.79 (5), b = 123.47 (5), c = 187.77 (5) A. The crystals diffract to 3.0 A and there are eight molecules in the asymmetric unit. PMID- 15299475 TI - Structure of chicken skeletal muscle troponin C at 1.78 A resolution. AB - The structure of chicken skeletal muscle troponin C (TnC) has been refined to an R value of 0.168, using 14 788 reflections, in the resolution range 8.0-1.78 A. Our earlier 2 A resolution structure [Satyshur, Rao, Pyzalska, Drendel, Greaser & Sundaralingam (1988). J. Biol. Chem. 263, 1628-1647] served as the starting model. The refined model includes atoms for all protein residues (1-162), 2 Ca(2+) ions, 169 water molecules and one sulfate ion. The high-resolution refinement shows more clearly the details of the protein and water structure. The side chains Glu63, Cysl01, Arg123, Aspl40 and Asp152 adopt two discretely ordered conformations. The long central helix is only slightly curved/bent (7.9 degrees ) and all the central helix NH.O=C hydrogen bonds are intact. Seven of the nine carbonyl O atoms of the mid segment of this helix, including the D/E linker region, are hydrogen bonded to water molecules which weakens the helix hydrogen bonds. In contrast, in each of the protected upper and lower thirds of the long central helix, only two carbonyl O atoms are hydrogen bonded to water molecules. The hydrogen-bonding patterns displayed by some of the carbonyl O atoms of NT and A helices of the N-terminal domain and the F and H helices of the C-terminal domain, which are on the exposed surface of the protein, are similar. The B helix of the calcium-free site I is kinked, with the local helix axes at either end making an angle of 39 degrees, by two inserted water molecules between N-H and O=C groups, breaking the adjacent helix hydrogen bonds. A sulfate ion from the crystallization buffer is also trapped in the B helix between the guanidinium group of Arg47 and these two inserted water molecules. The C helix of site II is devoid of similar hydration and is probably responsible for the different interhelical angles A/B at site I (134 degrees ) and C/D at site II (149 degrees ). Extensive interhelix hydrogen bonds occur between the side chains of the C and D helices of the 'apo' site II: Gln51-Asp89, Asn52-Asp89, Glu57-Gln85, Glu57 Glu88 and Glu64-Arg84, which apparently are disrupted upon Ca uptake and the resulting rearrangement of the helices expose the side chains, lining the palm of the N-(and C-) terminal domains, for interaction with specific peptide fragment of troponin I (Tnl) during muscle contraction. The dominant crystal packing motif involves a head-to-tail interaction between the N-terminal domain A helix of one molecule and the palm of the C-terminal domain of the 3(2)-related molecule, in a manner similar to that which can be expected for the TnC-TnI complex. Similar interactions may also be responsible for the dimerization of TnC at low pH. PMID- 15299476 TI - Structure of the recombinant Paramecium tetraurelia calmodulin at 1.68 A resolution. AB - The crystal structure of the recombinant calmodulin from Paramecium tetraurelia (rPCaM, M(r) = 16 700, 148 residues) has been determined at 1.68 A resolution. X ray intensity data were collected at 263 K using a Siemens-Nicolet area detector and Cu Kalpha radiation from a rotating-anode source. A total of 35 936 observations were processed with XENGEN1.3 and scaled to yield 16 255 unique reflections with R(symm)(I) of 4.1%. The crystals are triclinic, with unit-cell dimensions a = 29.89, b = 53.42, c = 25.35 A, alpha = 93.67, beta = 96.88, gamma = 89.24 degrees, space group P1, with one molecule in the unit cell. The atomic coordinates of the wild-type Paramecium calmodulin (PCaM) studied in our laboratory provided the starting model. Refinement of the structure by X-PLOR and refitting it into omit maps yielded an R value of 0.194 for 15 965 reflections greater than 3sigma(F) in the 6.0-1.68 A resolution range. The final model contained 1165 protein atoms for all of the 148 residues, four Ca(2+) ions, and 172 water molecules. The dumbbell structure has seven alpha-helices including a long 7.8 turn central helix connecting the two terminal domains each containing two EF-hand (helix-loop-helix motif) calcium-binding sites. The loops within each pair of EF-hand motifs in the N- and C-terminal domains are brought into juxtaposition to form a pair of hydrogen-bonded antiparallel beta-sheets which are extended at either ends by water bridges. The four calcium-binding EF-hands are superposable with r.m.s. deviations of 0.31-0.79 A. The best agreement is between site 1 and site 3 and the worst agreement is between site 1 and 4. The largest differences are in the ninth and tenth residues of the calcium-binding loops probably because of their involvement in the mini beta-sheets. The calcium coordination distances vary between 2.04 and 2.69 A, average 2.34 A. The rPCaM and wild-type PCaM have an r.m.s. deviation of 0.36 A for equivalent C(alpha) atoms. The side chains of Lys13 and Lys115 are more extended in rPCaM compared to the wild type where the post-translational modified di- and tri-methylated lysine residues are more folded. The sequence of PCaM differs from those of mammalian (MCaM) and Drosophila calmodulin (DCaM), but the overall structures are very similar, with r.m.s,. deviations of 0.44 and 1.68 A for equivalent C(alpha) atoms, respectively. However, in rPCaM, the first four N-terminal residues stretch out and make intermolecular crystal contacts, in contrast to those in recombinant Drosophila calmodulin (rDCaM), they stretch out in the opposite direction and towards the second calcium-binding site (see note below), while in MCaM and wild-type PCaM, the N-terminal residues are not visible. The central helix in rPCaM has all its backbone hydrogen bonds intact with no unusually long separation between the carbonyl and amide groups as found in MCaM and rDCaM. PMID- 15299477 TI - Crystallization of two monoclonal Fab fragments of similar amino-acid sequence bound to the same area of horse cytochrome c and interacting by potentially distinct mechanisms. AB - The mouse monoclonal antibodies (mAb), 2E5.G10 and 1F5.D1, are specific for horse cytochrome c and appear to bind the same epitope, since their heavy (H) and light (L) chains are functionally interchangeable. Comparison of the amino-acid sequences suggests that slightly different interactions may be involved in antigen recognition. In addition, the H chains differ at only a few amino-acid residues from the H chain of a rat cytochrome c-specific mAb suggesting that specificity for one protein over another may be determined by these amino-acid differences. To address these possibilities, the three-dimensional structures of the Fab portions of the mAb bound to cytochrome c are being determined by X-ray diffraction analysis. Here we describe the preparation and crystallization of the two complexes with horse cytochrome c. The complex of the Fab fragment of 2E5.G10 with horse cytochrome c yielded crystals of X-ray diffraction quality under two sets of conditions; in both the space group was P2(1). The corresponding complex of 1F5.D1 under one of these conditions crystallized in the P2(1)2(1)2(1) space group. Three-dimensional X-ray data for these two complexes have been collected with nominal resolutions of 2.86 and 2.48 A, respectively. PMID- 15299478 TI - Isomorphous binding of mercury-substituted thiosaccharides to pertussis toxin crystals yields crystallographic phases. AB - An isomorphous derivative of pertussis toxin crystals was prepared using a 2 alpha-mercuric analog of N-acetyl neuraminic acid in a method analogous to the use of inhibitors labelled with heavy atoms to solve crystal structures of enzymes. This derivative exploits the specific binding between pertussis toxin and terminal sialic acid residues on receptor glycoproteins. Difference Patterson maps yielded heavy-atom sites which refined with good statistics, indicating that the protein probably does not undergo a conformational change on receptor binding. Mercuric analogs of other monosaccharides should be easily obtainable using the same synthetic strategy, suggesting a general method for derivatizing crystals of carbohydrate-binding proteins. PMID- 15299479 TI - Crystal structure of chloromuconate cycloisomerase from Alcaligenes eutrophus JMP134 (pJP4) at 3 A resolution. AB - Chloromuconate cycloisomerase (E.C. 5.5.1.7) is an enzyme involved in the 2,4 dichlorophenoxyacetate degradation pathway of Alcaligenes eutrophus JMP134 (pJP4). The crystal structure of this protein was determined at 3 A resolution by molecular-replacement techniques using atomic coordinates from the reported crystal structure of the homologous muconate cycloisomerase (E.C. 5.5.1.1) from Pseudomonas putida as the search model (42% identical positions in the sequences). Structure refinement by simulated-annealing and restrained least squares techniques converged at R = 0.195. In the crystals studied, space group I4, the protein is present as two octamers per unit cell with two subunits per asymmetric unit. Each subunit consists of two globular domains, one of which forms an alpha/beta-barrel. Comparison of this structure with that of muconate cycloisomerase reveals the reasons for the altered substrate specificity of chloromuconate cycloisomerase. Marked differences are observed in polarity, accessibility and hydrogen-bonding potential in the channel leading into the active site as well as in the active center itself. PMID- 15299480 TI - Comparison of two crystal structures of TGF-beta2: the accuracy of refined protein structures. AB - Transforming growth factor-beta is a multifunctional cell-growth regulator and is a member of the TGF-beta superfamily of cytokines. Each monomer is 112 amino acids long and the mature active form is a 25 kDa homodimer. Recently, the crystal structure of TGF-beta2 has been determined independently in two laboratories [Daopin, Piez, Ogawa & Davies (1992). Science, 257, 369-373; Schlunegger & Grutter (1992). Nature (London), 358, 430-434] and subsequently refined to higher resolutions [Daopin, Li & Davies (1993). Proteins Struct. Funct. Genet. In the press; Schlunegger & Grutter (1993). J. Mol. Biol. In the press]. A detailed structural comparison shows that the two structures are nearly identical with the differences mostly located on the mobile regions of the molecule. The r.m.s. differences between the two structures are 0.10 A for 104 pairs of C(alpha) atoms, 0.15 A for 434 pairs of main-chain atoms, 0.33 A for 860 out of 890 pairs of protein atoms and a correlation of 90% between the temperature B factors of all protein atoms. Based on a comparison of the water molecules, a B value of 60.0 A(2) is recommended as the cut off for modeling new waters. The structural identity is striking because in one case the material was expressed in vivo in CHO cells whereas in the other case it was expressed in E. coli and had to be refolded in vitro. The overall coordinate errors are estimated to be 0.21 A from the Luzzati plot, 0.18 A from the sigma(A) plot, 0.24 A with Cruickshank's equations and 0.25 A using the empirical method of Perry & Stroud. These estimates are comparable to the r.m.s. structure superposition. The r.m.s. differences correlate very well with the crystallographic B values and the relation is best described with the Cruickshank formula. In addition to the estimation of an overall error, a new application of the Cruickshank formula is presented here to estimate the local errors. PMID- 15299481 TI - X-ray analysis of metal-substituted human carbonic anhydrase II derivatives. AB - Metal-substituted crystals of human carbonic anhydrase II belonging to space group P2(1) with cell dimensions a = 42.7, b = 41.7, c = 73.0 A and beta = 104.6 degrees were analyzed crystallographically. The resolution limit ranged from 1.82 to 1.92 A with high completeness (86.2-90.7%). Cobalt(II)-substituted carbonic anhydrase has a tetrahedral coordination around the metal both at pH 6 and pH 7.8, similar to the native zinc enzyme. In contrast, the catalytically inactive copper(II), nickel(II) and manganese(II) derivatives showed increased coordination number around the metal ion. Whereas the copper is best described as penta-coordinated, the nickel and manganese are best described as hexa coordinated. The results are briefly compared with spectroscopic observations and our current view on carbonic anhydrase catalysis. PMID- 15299482 TI - Wild-type and E106Q mutant carbonic anhydrase complexed with acetate. AB - The molecular structures of the acetate complexes of wild-type human carbonic anhydrase II (HCAII) and of E106Q mutant human carbonic anhydrase II were solved with high completeness (89-91%) to 2.1 and 1.9 A resolution, respectively. Both wild-type and mutant enzyme crystallize in space group P2(1) with cell dimensions a = 42.7, b = 41.7, c = 73.0 A and beta = 104.6 degrees. The altered active-site hydrogen-bond network caused by the mutation results in a different binding of the inhibitor in the two complexes. In the mutant, but not in the wild-type complex, a carboxylate O atom is within hydrogen-bond distance of Thr199 Ogamma1. In the wild-type enzyme ligand hydrogen bonding to this atom is normally only found for hydrogen-bond donors. The importance of this discrimination on catalysis by the enzyme is discussed briefly. PMID- 15299483 TI - Crystallization and preliminary X-ray diffraction studies of the bacteriophage Qbeta. AB - Crystals of bacteriophage Qbeta have been obtained by the vapor-diffusion technique. The crystals diffract to at least 3.5 A resolution. The crystal space group is C222(1) with the unit-cell parameters a = 478, b = 296, c = 477 A, alpha = beta = gamma = 90 degrees. The unit cell contains four virus particles. A pattern of systematic extinctions has been used to deduce the packing of the particles in the cell. A limited data set to 3.9 A resolution has been collected, and the predicted position has been confirmed by the self-rotation and the Patterson functions. PMID- 15299484 TI - Crystallization and preliminary X-ray studies on the mannose-specific lectin from Amaryllis bulbs. AB - Affinity-purified amaryllis lectin was used to grow single crystals using the hanging-drop method. The space group was found to be C2 with unit-cell dimensions a = 73.4 (1), b = 100.3 (1), c = 62.2 (1) A and beta = 137.3 (2) degrees. Data to 2.25 A resolution have been recorded and solution of the structure is currently underway by means of molecular-replacement techniques. PMID- 15299486 TI - Orientation of non-crystallographic symmetry axes in protein crystals. AB - A survey of 129 protein crystal structures with more than one molecule per asymmetric unit shows that local (non-crystallographic) symmetry axes are not randomly oriented. When compared to the crystal cell edges, face diagonals, body diagonal and reciprocal cell edges, 65% of the local symmetry axes are found to be parallel to one of the reference directions to within 15 degrees; another 18% are orthogonal to within 3 degrees; only 17% are in general orientations. In monoclinic, trigonal and hexagonal crystals, a majority of the local symmetry axes are orthogonal to the unique axis, while preferred orientations are parallel to the cell edges in orthorhombic crystals. PMID- 15299487 TI - Structure of recombinant bovine interferon-gamma at 3.0 A resolution. AB - The three-dimensional crystal structure of recombinant bovine interferon-gamma was determined using the multiple isomorphous replacement method at 3.0 A and refined to an R factor of 19.2%. This protein crystallizes in space group P2(1)2(1)2(1) with unit-cell parameters of a = 42.8, b = 79.9 and c = 85.4 A. There is one functional dimer in the asymmetric unit. The two polypeptide chains are related by a non-crystallographic twofold symmetry axis. The secondary structure is predominantly alpha-helical with extensive interdigitation of the alpha-helical segments of the polypeptide chains that make up the dimer. The secondary structure, tertiary structure and topology of this molecule are identical to the previously reported structures of recombinant rabbit interferon gamma and recombinant human interferon-gamma. The molecular topology is also similar to that of murine interferon-beta. These structural similarities strongly indicate the presence of a unique topological feature (fold) among gamma interferons from different species, and also among the different classes of interferons. PMID- 15299488 TI - Derivation by statistical methods of phase information from multiple-wavelength anomalous diffraction data. Basic questions, 'best' electron-density map, implementation and tests. AB - A probability distribution of the structure factor is established from the analysis of the effects of errors involved in the multiple-wavelength anomalous diffraction (MAD) method. This probability distribution, derived from those of the intensities, is two-dimensional for acentric reflections and uni-dimensional for centric reflections. It permits, using the centroid of the distribution, the calculation of the modulus and the phase of the 'best' structure factor. The procedure for extracting the phase and its figure of merit is presented. Tests performed on simulated data show the contribution of this method with respect to other methods which use a distribution of only the phase as a function of the error of closure. PMID- 15299489 TI - Mean phase error and the map-correlation coefficient. AB - In judging the effectiveness of methods of solving crystal structures, or in phase refinement and development, two criteria are commonly used. The first is the mean phase error, which may be weighted in some way, and the second is the map correlation coefficient which describes the similarity of a map with estimated phases to that with true phases. It is shown that these two measures are directly related and that given the individual phase errors the map correlation coefficient may be found without the need to calculate a map. Various aspects of this connection are examined, including the map correlation coefficient when weights are used for calculating maps and the conditions under which phase extension leads to maps with a higher map correlation coefficient - which involves a balance between the advantage of employing more data and the disadvantage that the extra data may have a higher average phase error. PMID- 15299490 TI - High-resolution structure of the complex between carboxypeptidase A and L-phenyl lactate. AB - The X-ray structures of native carboxypeptidase A and of the enzyme-inhibitor complex with L-phenyl lactate have been refined at 1.54 and 1.45 A resolution to R factors of 0.151 and 0.161, respectively. Crystals of the complex were isomorphous with the native crystals (space group P2(1), a = 51.60, b = 60.27, c = 47.25 A, beta = 97.27 degrees ). The high-resolution electron density allowed correction of many side-chain positions in the classical carboxypeptidase A model. This reflects the advantages of the high-quality complete synchrotron data collected with an imaging plate detector. The conformational changes in the active centre of the enzyme upon binding of the inhibitor are restricted to only two residues, Tyr248 and Arg145. L-Phenyl lactate is bound in the S1' pocket and forms hydrogen bonds to Arg145, Glu270 and to the zinc-bound water molecule. The present structure provides an explanation for the higher stability of the complexes with the products of esterolysis in comparison with those of amidolysis. This is consistent with the finding that product release is rate limiting for esters but not for peptides. PMID- 15299491 TI - Structure of the crystalline complex of cytidylic acid (2'-CMP) with ribonuclease at 1.6 A resolution. Conservation of solvent sites in RNase-A high-resolution structures. AB - The X-ray structure of the inhibitor complex of bovine ribonuclease A with cytidylic acid (2'-CMP) has been determined at 1.6 A resolution and refined by restrained least squares to R = 0.17 for 11 945 reflections. Binding of the inhibitor molecule to the protein is confirmed to be in the productive mode associated with enzyme activity. A study of conserved solvent sites amongst high resolution structures in the same crystal form reveals a stabilizing water cluster between the N and C termini. PMID- 15299492 TI - Structure determination and refinement of ribulose 1,5-bisphosphate carboxylase/oxygenase from Synechococcus PCC6301. AB - The structure of an activated quaternary complex of ribulose 1,5-bisphosphate carboxylase/oxygenase (rubisco) from Synechococcus PCC6301 has been solved by molecular replacement. The protein crystallizes in an orthorhombic P2(1)2(1)2(1) unit cell with a complete L(8)S(8) complex consisting of 4608 residues (37 680 non-hydrogen atoms) in the asymmetric unit. Data were collected both on film and image plate using synchrotron radiation; there were 218 276 unique reflections in the final 2.2 A data set. The eightfold non-crystallographic symmetry could be used both to improve map quality and to reduce the computing requirements of refinement. The coordinates were refined using strict non-crystallographic symmetry constraints. The stereochemistry of the final model is good, and the model has an R value of 20.0% for the reflections between 7 and 2.2 A. PMID- 15299493 TI - Refined structure of cadmium-substituted concanavalin A at 2.0 A resolution. AB - The three-dimensional structure of cadmium-substituted concanavalin A has been refined using X-PLOR. The R factor on all data between 8 and 2 A is 17.1%. The protein crystallizes in space group I222 with cell dimensions a = 88.7, b = 86.5 and c = 62.5 A and has one protein subunit per asymmetric unit. The final structure contains 237 amino acids, two Cd ions, one Ca ion and 144 water molecules. One Cd ion occupies the transition-metal binding site and the second occupies an additional site, the coordinates of which were first reported by Weinzierl & Kalb [FEBS Lett. (1971), 18, 268-270]. The additional Cd ion is bound with distorted octahedral symmetry and bridges the cleft between the two monomers which form the conventional dimer of concanavalin A. This study provides a detailed analysis of the refined structure of saccharide-free concanavalin A and is the basis for comparison with saccharide complexes reported elsewhere. PMID- 15299494 TI - Determination and refinement of the canine parvovirus empty-capsid structure. AB - The canine parvovirus (CPV) empty-capsid structure has been determined and refined to 3.0 A resolution in the tetragonal space group P4(3)2(1)2 with cell dimensions a = b = 254.5 and c = 795.0 A. The successful structure determination shows that reasonably good diffraction data were obtained in spite of the very long c axis. The structure was solved by molecular replacement using the electron density of CPV full particles in a monoclinic space group. The phases were refined by non-crystallographic symmetry averaging. The structure refinement was carried out by using the programs PROLSQ and X-PLOR. The final R factor for the structure that included 85 water molecules per icosahedral asymmetric unit was 21.1% for reflections between 6.0 and 3.0 A resolution with an r.m.s. deviation of bond lengths of 0.020 A from ideal values. The structure of CPV empty capsids showed conformational differences with respect to full capsids at a region where icosahedrally ordered DNA in full particles interacts with the capsid protein. It also confirmed the absence of density along the fivefold axis in the CPV empty particle structure in contrast to the situation in CPV full particles. PMID- 15299495 TI - A monoclinic crystal with R32 pseudo-symmetry: a preliminary report of Nodamura virus structure determination. AB - We have crystallized Nodamura virus, a T = 3 icosahedral virus that can infect both mammalian and insect hosts. Crystals are monoclinic, with two crystallographically independent virus molecules per asymmetric unit. Packing analysis reveals a pseudo-rhombohedral (pseudo-C2 in the monoclinic setting) arrangement of virus particles in the crystal lattice. Crystals differ from the R32 symmetry by rotational and translational deviations. The rhombohedral packing arrangement and its failure to describe the exact virus packing is analyzed in detail. The icosahedral threefold axis is rotated from the body diagonal of the pseudo-rhombohedral cell, breaking the rhombohedral symmetry. The C2 pseudo symmetry breaks down rotationally and/or translationally. PMID- 15299496 TI - On the computation of the fast rotation function. AB - A new more efficient algorithm for the evaluation of the fast rotation function coefficients is derived. Its implementation in standard programs is straightforward. PMID- 15299497 TI - The ordered water cluster in vitamin B12 coenzyme at 15 K is stabilized by C H...O hydrogen bonds. AB - The hydrogen-bonding interactions of the ordered water cluster in the pocket region of vitamin B(12) coenzyme at 15 K are analyzed from published atomic coordinates. Several of these water molecules accept C-H.O hydrogen bonds from the pocket wall, most of which have the function to complete tetrahedral hydrogen bond coordination. This is the largest water assembly for which the stabilizing function of C-H.O interactions has been described. PMID- 15299498 TI - Crystallization and preliminary X-ray studies on the trypsin inhibitor I-2 from wheat germ and its complex with trypsin. AB - A Bowman-Birk type trypsin inhibitor I-2, M(r) = 14 000, 123 amino-acid residues, isolated from wheat germ, and its complex with trypsin have been crystallized. For I-2 two morphologically different crystal forms were obtained. Crystal form 1 is tetragonal, P4(1)22 or P4(3)22, with a = 55.45 (2), c = 129.1 (2) A and V = 3.97 (2) x 10(5) A(3). The crystals diffract X-rays very anisotropically, to less than 6 A resolution normal to the c* direction, but up to 3 A resolution in the other directions. Crystal form 2 is monoclinic, space group C2. The cell parameters show significant variation even for crystals in the same batch. The median parameters are: a = 83.9, b = 41.5, c = 45.7 A, beta = 95.9 degrees and V = 1.58 x 10(5) A(3). The diffraction pattern is isotropic and reflections up to 2.2 A resolution were observed. The crystals of the complex between bovine trypsin and I-2 (2:1) belong to the orthorhombic space group P2(1)2(1)2(1) with a = 73.49 (2), b = 120.56 (3), c = 70.04 (2) A and V = 6.206 (5) x 10(5) A(3). The crystals diffract up to 2.3 A resolution, and contain one complex of 60 100 Da in an asymmetric unit. PMID- 15299499 TI - Location of haem in bacterioferritin of E. coli. AB - . A low-resolution partial structure of bacterioferritin was solved using a combination of molecular replacement and rigid-body refinement methods. Modification of bacterioferritin crystals by soaking in tetrachloroplatinate results in a phase transition from tetragonal symmetry (space group P4(2)2(1)2) to a pseudo-cubic one (approximate space group I432). Helical parts of human H ferritin structure stripped of side chains beyond the C(beta) atoms were used as the model. An electron-density map of the refined model revealed a region of extended density which by its shape and position in a pocket between helices was identified as haem. Inclusion of haem in the refinement showed that it can occupy only one of two symmetry-related sites near a twofold axis of the molecule. PMID- 15299502 TI - PRISM: topologically constrained phased refinement for macromolecular crystallography. AB - We describe the further development of phase refinement by iterative skeletonization (PRISM), a recently introduced phase-refinement strategy [Wilson & Agard (1993). Acta Cryst. A49, 97-104] which makes use of the information that proteins consist of connected linear chains of atoms. An initial electron-density map is generated with inaccurate phases derived from a partial structure or from isomorphous replacement. A linear connected skeleton is then constructed from the map using a modified version of Greer's algorithm [Greer (1985). Methods Enzymol. 115, 206-226] and a new map is created from the skeleton. This 'skeletonized' map is Fourier transformed to obtained new phases, which are combined with any starting-phase information and the experimental structure-factor amplitudes to produce a new map. The procedure is iterated until convergence is reached. In this paper significant improvements to the method are described as is a challenging molecular-replacement test case in which initial phases are calculated from a model containing only one third of the atoms of the intact protein. Application of the skeletonization procedure yields an easily interpretable map. In contrast, application of solvent flattening does not significantly improve the starting map. The iterative skeletonization procedure performs well in the presence of random noise and missing data, but requires Fourier data to at least 3.0 A. The constraints of linearity and connectedness prove strong enough to restore not only missing phase information, but also missing amplitudes. This enables the use of a powerful statistical test, analogous to the 'free R factor' of conventional refinement [Brunger (1992). Nature (London), 355, 472-474], for optimizing the performance of the skeletonization procedure. In the accompanying paper, we describe the application of the method to the solution of the structure of the protease inhibitor ecotin bound to trypsin and to a single isomorphous replacement problem. PMID- 15299503 TI - PRISM: application to the solution of two protein structures. AB - The previous paper described a phase-refinement strategy for protein crystallography which exploited the information that proteins consist of connected linear chains of atoms. Here the method is applied to a molecular replacement problem, the structure of the protease inhibitor ecotin bound to trypsin, and a single isomorphous replacement problem, the structure of the N terminal domain of apolipoprotein E. The starting phases for the ecotin-trypsin complex were based on a partial model (trypsin) containing 61% of the atoms in the complex. Iterative skeletonization gave better results than either solvent flattening or twofold non-crystallographic symmetry averaging as measured by the reduction in the free R factor [Brunger (1992). Nature (London), 355, 472-474]. Protection of the trypsin density during the course of the refinement greatly improved the performance of both skeletonizing and solvent flattening. In the case of apolipoprotein E, previous attempts using solvent flattening had failed to improve the SIR phases to the point of obtaining an interpretable map. The combination of iterative skeletonization and solvent flattening decreased the phase error with respect to the final refined structure, significantly more than solvent flattening alone. The final maps generated by the skeletonization procedure for both the ecotin-trypsin complex and apolipoprotein E were readily interpretable. PMID- 15299504 TI - Structure of Pseudomonas aeruginosai zinc azurin mutant Asn47Asp at 2.4 A resolution. AB - The Pseudomonas aeruginosa azurin mutant Asn47Asp has been isolated, its spectroscopic and kinetic properties characterized, and the X-ray crystal structure of its zinc derivative determined. While the optical and electron paramagnetic resonance spectra as well as the electron-transfer activity of the mutant are very similar to the wild-type values, the Asn47Asp reduction potential is slightly increased by 20 mV. The mutant crystallized in the orthorhombic space group P2(1)2(1)2(1) with cell dimensions a = 57.8, b = 81.5 and c = 112.6 A. There are four molecules in the asymmetric unit, packed as a tetramer which consists of two independent dimers. The zinc site of this mutant structure is similar to the wild-type zinc azurin and, in particular, the metal-binding site is almost identical to the site found in the wild-type zinc-azurin structure [Nar, Huber, Messerschmidt, Filippou, Barth, Jaquinod, Kamp & Canters (1992). Eur. J. Biochem. 205, 1123-1129]. The Asp47 side chain at that mutation site takes on a very similar orientation to Asn47 in the wild-type structure preserving the two hydrogen bonds with the neighbouring Thr113 NH and O(gamma)H. Therefore, the increased reduction potential of the mutant is probably a result of an altered charge distribution close to the metal site. PMID- 15299505 TI - Anthracycline-DNA interactions at unfavourable base-pair triplet-binding sites: structures of d(CGGCCG)/daunomycin and d(TGGCCA)/adriamycin complexes. AB - The structures of two hexanucleotide-anthracycline complexes d(CGGCCG)/daunomycin and d(TGGCCA)/adriamycin have been determined using single-crystal X-ray diffraction techniques. In both cases the anthracycline molecule is bound to non preferred d(YGG) base-pair triplet sites. For both complexes the crystals are tetragonal and belong to the space group P4(1)2(1)2. Unit-cell dimensions are a = 28.07 (2), c = 53.35 (1) and a = 28.01 (1), c = 52.99 (1) A, respectively, and the asymmetric unit of both structures consists of one strand of DNA, one drug molecule and approximately 50 water molecules. For the d(CGGCCG) complex the refinement converged with an R factor of 0.21 for 1108 reflections with F >/= 2sigma(F) in the resolution range 7.0-1.9 A. For the complex involving d(TGGCCA) the final R value was 0.22 for 1475 reflections in the range 7.0-1.7 A with the same criterion for observed data. Both complexes are essentially isomorphous with related structures but differ in terms of the number of favourable van der Waals interactions of the amino sugars of the drug molecules with the DNA duplexes and the formation in the minor groove of heterodromic pentagonal arrangements of hydrogen bonds involving water molecules which link the amino sugars to the DNA. These differences in structure are used to rationalize the lack of affinity of daunomycin-type anthracyclines for d(YGG) and d(YGC) sites. PMID- 15299506 TI - DNA helix structure and refinement algorithm: comparison of models for d(CCAGGCm5CTGG) derived from NUCLSQ, TNT and X-PLOR. AB - In an earlier study [Heinemann & Hahn (1992). J. Biol. Chem. 267, 7332-7341], the crystal structure of the double-stranded B-DNA decamer d(CCAGGCm(5)CTGG) was refined with NUCLSQ to R = 17.4% against 3799 2sigma structure amplitudes in the resolution range 8-1.7 A. This structure has now been re-refined against the same diffraction data using either TNT or X-PLOR in order to determine to what extent the resulting DNA conformations would differ and to examine the suitability of these programs for the refinement of oligonucleotide structures. The R value from the NUCLSQ refinement could not be reached with either TNT or X-PLOR, although both programs yield reasonably refined DNA models showing root-mean-square deviations against the NUCLSQ model of the decamer duplex of 0.25 and 0.32 A, respectively. Some derived local structure parameters differ depending on the refinement procedure used. This holds true for several exocyclic torsion angles of the sugar-phosphate backbone, whereas sugar puckers as well as helical and base-pair stacking parameters are only weakly influenced. A subset of 15 solvent sites with low temperature factors is conserved in all three models. PMID- 15299507 TI - Determination of three crystal structures of canavalin by molecular replacement. AB - Canavalin, the major reserve protein of the jack bean, was obtained in four different crystal forms. From the structure determined by multiple isomorphous replacement in a hexagonal unit cell, the structures of three other crystals were determined by molecular replacement. In two cases, the rhombohedral and cubic crystals, placement was facilitated by coincidence of threefold molecular symmetry with crystallographic operators. In the orthorhombic crystal the canavalin trimer was the asymmetric unit. The rhombohedral, orthorhombic and cubic crystal structures were subsequently refined using a combination of several approaches with resulting R factors of 0.194, 0.185 and 0.211 at resolutions of 2.6, 2.6 and 2.3 A, respectively. Variation in the conformation of the molecule from crystal to crystal was small with an r.m.s. deviation in Calpha positions of 0.89 A. Packing is quite different among crystal forms but lattice interactions appear to play little role in the conformation of the molecule. Greatest variations in mean position are for those residues that also exhibit the greatest thermal motion. Crystal contacts in all crystals are mediated almost exclusively by hydrophilic side chains, and three to six intermolecular salt bridges per protein subunit are present in each case. PMID- 15299508 TI - Crystallization and preliminary structure determination of porcine aldehyde reductase from two crystal forms. AB - Aldehyde reductase from porcine kidney has been crystallized from buffered ammonium sulfate solutions. Two crystal forms are monoclinic, space group P2(1), with a = 56.2, b = 98.1, c = 73.2 A, beta = 112.5 degrees and a = 92.4, b = 62.1, c = 59.0 A, beta = 94.6 degrees. A third crystal form is hexagonal with a = b = 166.0, c = 66.0 A, alpha = beta = 90.0 degrees and gamma = 120.0 degrees. Molecular-replacement structure solutions have been successfully obtained for the two monoclinic crystal forms. The crystallographic R factor at 8-2.8 A resolution for the two monoclinic crystal forms is currently 0.23 and 0.25, respectively. There are two molecules per asymmetric unit related by a non-crystallographic twofold axis. The aldehyde reductase models are supported by the arrangement of the molecules in their respective unit cells and by electron densities corresponding to amino-acid side chains not included in the search structures. PMID- 15299509 TI - X-ray structure of monoclinic turkey egg lysozyme at 1.3 A resolution. AB - Monoclinic crystals of turkey egg lysozyme (TEL, E.C. 3.2.1.17) were obtained from 2.2 M ammonium sulfate solution at pH 4.2. They belong to space group P2(1) with unit-cell dimensions a = 38.07, b = 33.20, c = 46.12 A and beta = 110.1 degrees, and contain one molecule in the asymmetric unit (V(m) = 1.91 A(3) Da( 1)). The three-dimensional structure of TEL was solved by the method of multiple isomorphous replacement with anomalous scattering. Area detector data to 1.5 A resolution from native and heavy-atom derivatives were used for the structure determination. The structure was refined by the simulated-annealing method with diffraction data of 10-1.30 A resolution. The conventional R factor was 0.189. The root-mean-square deviations from ideal bond distances and angles were 0.016 A and 2.9 degrees, respectively. The backbone structure of TEL is very similar to that of hen egg lysozyme (HEL) and the difference in seven amino-acid residues does not affect the basic folding of the polypeptide chain. Except for the region from Gly101 to Gly104, the geometry of the active-site cleft is conserved between TEL and HEL. The Gly101 residue is located at the end of the sugar-binding site and the structural change in this region between TEL and HEL is considered to be responsible for the difference in their enzymatic properties. PMID- 15299510 TI - Direct-space methods in phase extension and phase determination. II. Developments of low-density elimination. AB - The low-density elimination method for phase extension and refinement [Shiono & Woolfson (1992). Acta Cryst. A48, 451-456] has been improved by substituting a smoother density-modification procedure for the original sharp cut-off function. In addition, better criteria have been found for limiting the number of refinement cycles, which gives a better final result for much less work. The effectiveness of the process has been illustrated by phase refinement for a protein with high-resolution (1.17 A) data containing 808 independent non-H atoms plus 83 water molecules in the asymmetric unit; the unweighted mean-phase error was reduced from 74 to 39.3 degrees. Phase extension and refinement was also demonstrated for pig 2Zn insulin starting with multiple isomorphous replacement (MIR) phases at 1.9 A and extending out to 1.5 A. There was a significant improvement of phases and the final map had a correlation coefficient of 0.540. PMID- 15299511 TI - Crystallization and preliminary X-ray diffraction studies of C-1027-AG, the apoprotein of the macromolecular antitumor antibiotic C-1027 from Streptomyces globisporus. AB - C-1027-AG, the apoprotein of the macromolecular antitumor antibiotic C- 1027, isolated from Streptomyces globisporus, was crystallized by the vapor-diffusion procedure using 2-methyl-2,4-pentanediol as a precipitant. The crystals belong to the orthorhombic system, space group P2(1)2(1)2(1), with unit-cell dimensions a = 55.1, b = 61.3 and c = 79.1 A. Assuming that the asymmetric unit contains two or three molecules, the V(m) value is calculated as 3.2 or 2.1 A(3) Da(-1), respectively. A total of 7630 independent reflections was obtained up to 2.5 A resolution with synchrotron radiation, the merging R factor being 0.077 for 24 713 measurements. PMID- 15299512 TI - Structure determination of plastocyanin from a specimen with a hemihedral twinning fraction of one-half. AB - The structure determination of a macromolecule from a hemihedrally twinned crystal specimen with a twinning fraction of one-half is described. Twinning was detected by analysis of crystal-packing density and intensity statistics. The structure was solved using molecular replacement, and the positioned search model was used to overcome the twinning by a novel method of 'detwinning' the observed data. Estimates of the unobservable crystallographic intensities from each of the twin domains were obtained and used to refine the model. The structure of a new algal plastocyanin from Chlamydomonas reinhardtii was determined by this method to 1.6 A resolution with a 'twinned' R factor of 15.6%. Additional data from a crystal specimen with a low twinning fraction were used to establish the accuracy of the structure solution from the perfectly twinned data, and to finalize the refinement to 1.5 A resolution and a true R factor of 16.8%. Methods for detecting twinning and obtaining a molecular-replacement solution in the presence of twinning are discussed. PMID- 15299513 TI - Structure of the mammalian catalytic subunit of cAMP-dependent protein kinase and an inhibitor peptide displays an open conformation. AB - The crystal structure of a binary complex of the porcine heart catalytic (C) subunit of cAMP-dependent protein kinase (space group P4(1)32; a = 171.5 A) complexed with a di-iodinated peptide inhibitor, PKI(5-24), has been solved and refined to 2.9 A resolution with an overall R of 21.1%. The r.m.s. deviations from ideal bond lengths and angles are 0.022 A and 4.3 degrees. A single isotropic B of 17 A(2) was used for all atoms. The structure solution was carried out initially by molecular replacement of electron density followed by refinement against atomic coordinates from orthorhombic crystals of a binary complex of the mouse recombinant enzyme previously described [Knighton, Zheng, Ten Eyck, Ashford, Xuong, Taylor & Sowadski (1991). Science, 253, 407-414]. The most striking difference between the two crystal structures is a large displacement of the small lobe of the enzyme. In the cubic crystal, the beta-sheet of the small lobe is rotated by 15 degrees and translated by 1.9 A with respect to the orthorhombic crystal. Possible explanations for why this binary complex crystallized in an open conformation in contrast to a similar binary complex of the recombinant enzyme are discussed. This study demonstrates that considerable information about parts of a crystal structure can be obtained without a complete crystal structure analysis. Specifically, the six rigid-group parameters of a poly alanine model of the beta-structure were obtained satisfactorily from a crystal structure by refinement of difference Fourier coefficients based on an approximate partial structure model. PMID- 15299514 TI - Bovine seminal ribonuclease: structure at 1.9 A resolution. AB - The crystal structure of bovine seminal ribonuclease, a homodimeric enzyme closely related to pancreatic ribonuclease, has been refined at a nominal resolution of 1.9 A employing data collected on an electronic area detector. The final model consists of two chains containing 1990 non-H atoms, seven sulfate anions and 113 water molecules per asymmetric unit. The unit-cell parameters are a = 36.5 (1), b = 66.7 (1) and c = 107.5 (2) A, space group P22(1)2(1). The R factor is 0.177 for 16 492 reflections in the resolution range 6.0-1.9 A and the deviations from ideal values of bond lengths and bond angles are 0.020 A and 3.7 degrees, respectively. The molecule is formed by two pancreatic like chains, which have their N-terminal segments interchanged so that each active site is formed by residues from both subunits. The two chains are related by a non crystallographic twofold symmetry and are covalently linked by two consecutive disulfide bridges, which form an unusual sixteen-membered ring across the dimer interface. The deviations from the molecular symmetry, the hydration shell and the sulfate-binding sites are also discussed in relation to the known structure of the pancreatic enzyme. PMID- 15299515 TI - Structure of triosephosphate isomerase from Escherichia coli determined at 2.6 A resolution. AB - The structure of triosephosphate isomerase (TIM) from the organism Escherichia coli has been determined at a resolution of 2.6 A. The structure was solved by the molecular replacement method, first at 2.8 A resolution with a crystal grown by the technique of hanging-drop crystallization from a mother liquor containing the transition-state analogue 2-phosphoglycolate (2PG). As a search model in the molecular replacement calculations, the refined structure of TIM from Trypanosoma brucei, which has a sequence identity of 46% compared to the enzyme from E. coli, was used. An E. coli TIM crystal grown in the absence of 2PG, diffracting to 2.6 A resolution, was later obtained by application of the technique of macro-seeding using a seed crystal grown from a mother liquor without 2PG. The final 2.6 A model has a crystallographic R factor of 11.9%, and agrees well with standard stereochemical parameters. The structure of E. coli TIM suggests the importance of residues which favour helix initiation for the formation of the TIM fold. In addition, TIM from E. coli shows peculiarities in its dimer interface, and in the packing of core residues within the beta-barrel. PMID- 15299516 TI - Crystallization and preliminary diffraction studies of the chemically synthesized domain A of Thermus flavus 5S rRNA: an RNA dodecamer double helix. AB - Crystals of domain A of Thermus flavus 5S rRNA have been obtained. The space group was found to be P4(3) with unit-cell dimensions a = b = 30.10 and c = 86.80 A. Data to 2.3 A have been recorded and solution of the structure is currently underway by means of molecular-replacement techniques. PMID- 15299517 TI - Crystallization of the OP-G2 Fab fragment: a fibrinogen mimic with specificity for the platelet glycoprotein IIb/IIIa. AB - The OP-G2 monoclonal antibody binds to the platelet integrin, gpIIb/IIIa, in a mode that mimics fibrinogen binding. The specificity of this antibody is mediated by the third complementarity-determining region (CDR3) loop of the immunoglobulin heavy chain which contains a sequence (RYD) related to the RGD recognition sequence of fibrinogen. The OP-G2 Fab fragment has been crystallized by vapor diffusion from solutions containing polyethylene glycol and imidazole malate (pH 5.6). The crystals belong to space group P2(1)2(1)2 with a = 93.1, b = 83.8 and c = 53.7 A. One Fab molecule is present in the asymmetric unit. A complete data set to 2.0 A resolution has been collected. PMID- 15299518 TI - Crystallographic analyses of an active HIV-1 ribonuclease H domain show structural features that distinguish it from the inactive form. AB - . An active recombinant preparation of the carboxy-terminal ribonuclease H (RNase H) domain of HIV-I reverse transcriptase has produced crystals of several different forms, including a trigonal prism form (P3(1); a = b = 52.03, c = 113.9 A with two molecules per asymmetric unit) and a hexagonal tablet form (P6(2)22 or P6(4)22; a = b = 93.5, c = 74.1 A with one molecule per asymmetric unit). The former appears to be isomorphous with crystals of a similar, but inactive, version of the enzyme that was used for a prior crystal structure determination [Davies, Hostomska, Hostomsky, Jordan & Matthews (1991). Science, 252, 88-95]. We have also obtained a structure solution for this crystal form and have refined it with 2.8 A resolution data (R = 0.216). We report here details of our crystallization studies and some initial structural results that verify that the preparation of active HIV-1 RNase H yields a protein that is not just enzymatically, but also structurally, distinguishable from the inactive form. Evidence suggests that region 538-542, which may be involved in the catalytic site and which is disordered in both molecules in the prior structure determination, is ordered in the crystal structure of the active enzyme, although the ordering may include more than one conformation for this loop. It should also be noted that, in the crystal structure of the trigonal form, RNase H monomers associate to form noncrystallographic twofold-symmetric dimers by fusing five stranded mixed beta sheets into a single ten-stranded dimerwide sheet, an assembly that was not remarked upon by previous investigators. PMID- 15299520 TI - The structure of an idarubicin-d(TGATCA) complex at high resolution. AB - The crystal structure of the DNA hexamer d(TGATCA) complexed with the anthracycline antibiotic idarubicin has been determined at 1.6 A resolution. The asymmetric unit consists of a single hexamer oligonucleotide strand, one drug molecule and 35 water molecules. The complex crystallizes in the tetragonal space group P4(1)2(1)2, Z = 8 with lattice dimensions of a = b = 28.19 (3), c = 52.77 (4) A, V = 41 935 A(3). The structure is isomorphous with a series of hexamer anthracycline complexes and was solved by molecular replacement. Restrained least squares methods interspersed with computer-graphics map inspection and model manipulation were used to refine the structure. The R factor is 0.22 for 2032 reflections with F >/= 3sigma(F) in the resolution range 8.0-1.6 A. The self complementary DNA forms a distorted B-DNA double helix with two idarubicin molecules intercalated in the d(TpG) steps of the duplex. The duplex is formed by utilization of a crystallographic twofold axis of symmetry. The idarubicin chromophore is oriented at right angles to the long axis of the DNA base pairs with the anthracycline amino-sugar moiety positioned in the minor groove. Our structure determination allows for comparison with a d(CGATCG)-idarubicin complex recently reported. Despite the sequence alteration at the intercalation step, the structures are very similar. The geometry of the intercalation and the nature of the interactions are conserved irrespective of the DNA sequence involved in the binding. PMID- 15299521 TI - Structure determination and refinement of benzamidine-inhibited trypsin from the North Atlantic salmon (Salmo salar) at 1.82 A resolution. AB - The structure of the serine protease trypsin from the North Atlantic salmon (Salmo salar) has been solved by molecular replacement and refined by restrained least-squares methods to a conventional R factor of 16.4% using diffractometer data in the 6.0-1.82 A resolution range (14 443 reflections greater than 3sigma). The model comprises 1793 protein atoms and 180 solvent molecules which were given unit occupancies, and the average temperature factors for protein atoms and solvent oxygen atoms are 15.2 and 36.8 A(2), respectively. The estimated error in atomic positions is about 0.2 A. The structure of salmon trypsin was solved and refined with only a small part of the amino-acid sequence known. However, a gene sequence of salmon trypsin has later become available. Some discrepancies between this sequence and the sequence obtained from the present X-ray crystal study indicate that the mentioned sequences may correspond to isoenzymes. The structure of salmon trypsin is similar to other trypsins of known structure. PMID- 15299522 TI - Structure of apo-azurin from Alcaligenes denitrificans at 1.8 A resolution. AB - The structure of apo-azurin from Alcaligenes denitrificans has been determined at high resolution by X-ray crystallography. Two separate structure analyses have been carried out, (i) on crystals obtained from solutions of apo-azurin and (ii) on crystals obtained by removal of copper from previously formed crystals of holo azurin. Data to 1.8 A resolution were collected from the apo-azurin crystals, by Weissenberg photography (with image plates) using synchrotron radiation and by diffractometry, and the structure was refined by restrained least-squares methods to a final R value of 0.160 for all data in the range 10.0-1.8 A. The final model of 1954 protein atoms, 246 water molecules (66 half-weighted), four SO(4)(2-) ions, and two low-occupancy (0.13 and 0.15) Cu atoms has r.m.s. deviations of 0.012, 0.045 and 0.013 A from standard bond lengths, angle distances and planar groups. For copper-removed azurin, data to 2.2 A were collected by diffractometry and the structure refined by restrained least squares to a final R value of 0.158 for all data in the range 10.0-2.2 A. The final model of 1954 protein atoms, 264 water molecules, two SO(4)(2-) ions, two low occupancy (0.18 and 0.22) metal atoms and one unidentified atom (modelled as S) has r.m.s. deviations of 0.013, 0.047 and 0.012 A from standard bond lengths, angle distances and planar groups. The two structures are essentially identical to each other and show no significant differences from the oxidized and reduced holo-azurin structures. The ligand side chains move slightly closer together following the removal of copper, with the radius of the cavity between the three strongly binding ligands, His 46, His 117 and Cys 112, shrinking from 1.31 A in reduced azurin to 1.24 A in oxidized azurin and 1.16 A in apo-azurin. There is a suggestion of increased flexibility in one of the copper-binding loops but the structure supports the view that the copper site found in holo-azurin is a stable structure, defined by the constraints of the polypeptide structure even in the absence of a bound metal ion. PMID- 15299523 TI - Conformational study of a putative HLTV-1 retroviral protease inhibitor. AB - The crystal structure of prolyl-glutaminyl-valyl-statyl-alanyl-leucine (Pro-Gln Val-Sta-Ala-Leu, C(32)H(57)N(7)0(9).5H(2)0, M(r) = 683.9 + 90.1), a putative HTLV 1 protease inhibitor based on one of the consensus retroviral protease cleavage sequences, and containing the statine residue [(4S,3S)-4-amino-3-hydroxy-6 methylheptanoic acid], has been determined by X-ray diffraction. The same molecule has been modelled in the active site of the HTLV-1 protease and both conformations have been compared. The peptide crystallizes as a pentahydrate in space group P2(1) with a = 10.874(2), b = 9.501(2), c = 21.062(5) A, beta = 103.68 (1) degrees, Z = 2, V= 2114.3 A(3), D(x) = 1.21 g cm(-3), micro = 8.02 cm( 1), T= 293 K, lambda(Cu Kalpha) = 1.5418 A. The structure has been refined to an R value of 0.070 for 2152 observed reflections. The peptide main chain can be described as extended and adopts the usual zigzag conformation from the prolyl to the statyl residue. The main difference in conformation between the individual observed and modelled molecules is located on the Sta, Ala and Leu residues with the main chain of the modelled molecule rotated by about 180 degrees as compared to the observed conformation in the crystal state. PMID- 15299524 TI - Purification, crystallization and preliminary X-ray investigation of aqualysin I, a heat-stable serine protease. AB - Aqualysin I, a thermostable protease found in the culture medium of Thermus aquaticus YT-1, has been purified to homogeneity using a combination of ion exchange and affinity chromatography. It is a polypeptide with a molecular weight of 28 350 [Kwon, Terada, Matsuzawa & Ohta (1988). Eur. J. Biochem. 173, 491-497] and is most active at 343-353 K and pH about 10.0 [Matsuzawa, Tokugawa, Hamaoki, Mizoguchi, Taguchi, Terada, Kwon & Ohta (1988). Eur. J. Biochem. 171, 441-447]. Crystals of the enzyme are monoclinic, space group P2(1), with cell dimensions a = 40.80 (5), b = 64.39 (6), c = 45.51 (6) A and beta = 109.1 (1) degrees. The asymmetric unit consists of a single molecule (V(m) = 1.99 A(3)Da(-1)). The crystals are stable to X-radiation and scatter to at least 2.8 A resolution. PMID- 15299525 TI - Crystallization of a Humicola lanuginosa lipase-inhibitor complex with the use of polyethylene glycol monomethyl ether. AB - The fungal Humicola lanuginosa lipase complexed with the inhibitor n dodecylphosphonate ethyl ester was crystallized in space group P2(1)2(1)2(1) with pseudotetragonal unit-cell parameters of a = 131.7 (2), b = 131.3 (1), c = 75.4 (1) A. 92% of X-ray diffraction data to 2.8 A resolution were collected with a final R(merge) = 8.5%. The crystals were grown using a new kind of precipitant - polyethylene glycol monomethyl ether (Peg-mme) of molecular weight 5000. PMID- 15299526 TI - 2.0 A refined crystal structure of the catalytic subunit of cAMP-dependent protein kinase complexed with a peptide inhibitor and detergent. AB - . A mutant (Serl39Ala) of the mouse recombinant catalytic (C) subunit of cAMP dependent protein kinase was co-crystallized with a peptide inhibitor, PKI(5-24), and MEGA-8 (octanoyl-N-methylglucamide) detergent. This structure was refined using all observed data (30 248 reflections) between 30 and 1.95 A resolution to an R factor of 0.186. R.m.s. deviations of bond lengths and bond angles are 0.013 A and 2.3 degrees, respectively. The final model has 3075 atoms (207 solvent) with a mean B factor of 31.9 A(2). The placement of invariant protein-kinase residues and most C:PKI(5-24) interactions were confirmed, but register errors affecting residues 55-64 and 309-339 were corrected during refinement by shifting the affected sequences toward the C terminus along the previously determined backbone path. New details of C:PKI(5-24) interactions and the Ser338 autophosphorylation site are described, and the acyl group binding site near the catalytic subunit NH(2) terminus is identified. PMID- 15299527 TI - 2.2 A refined crystal structure of the catalytic subunit of cAMP-dependent protein kinase complexed with MnATP and a peptide inhibitor. AB - . The crystal structure of a ternary complex containing the catalytic subunit of cAMP-dependent protein kinase, ATP and a 20-residue inhibitor peptide was refined at a resolution of 2.2 A to an R value of 0.177. In order to identify the metal binding sites, the crystals, originally grown in the presence of low concentrations of Mg(2+), were soaked in Mn(2+). Two Mn(2+) ions were identified using an anomalous Fourier map. One Mn(2+) ion bridges the gamma- and beta phosphates and interacts with Asp184 and two water molecules. The second Mn(2+) ion interacts with the side chains of Asn171 and Asp l84 as well as with a water molecule. Modeling a serine into the P site of the inhibitor peptide suggests a mechanism for phosphotransfer. PMID- 15299528 TI - Structure of the bovine eye lens protein gammaB(gammaII)-crystallin at 1.47 A. AB - The molecular structure of calf gammaB-crystallin (previously called gammaII), a lens-specific protein, has been refined to a crystallographic R factor of 18.1% for all reflection data, between 8.0 and 1.47 A, 25 959 hkl measured at 293 (1) K. 230 water molecules have been defined by difference Fourier techniques and included in a restrained least-squares refinement. Difference Fourier maps clearly indicated the presence of multiple sites for the sulfur atoms of Cys 18 and Cys 22 which were therefore given coupled second-site occupancies during the refinement. The sulfur atom in the major position of Cys 22 is in the reduced state. Either of the Cys 18 sites can form a high-energy disulfide bridge with the minor position of Cys 22. The position of the carboxy terminus and many other surface side chains have been further defined including the RGD signal peptide. The hydration of the backbone and the interdomain region has been analysed. 27 water molecules make extensive contacts to a single protein molecule and thus contribute to its stability. PMID- 15299529 TI - Protein hydration and water structure: X-ray analysis of a closely packed protein crystal with very low solvent content. AB - Low-humidity monoclinic lysozyme, resulting from a water-mediated transformation, has one of the lowest solvent contents (22% by volume) observed in a protein crystal. Its structure has been solved by the molecular replacement method and refined to an R value of 0.175 for 7684 observed reflections in the 10-1.75 A resolution shell. 90% of the solvent in the well ordered crystals could be located. Favourable sites of hydration on the protein surface include side chains with multiple hydrogen-bonding centres, and regions between short hydrophilic side chains and the main-chain CO or NH groups of the same or nearby residues. Major secondary structural features are not disrupted by hydration. However, the free CO groups at the C terminii and, to a lesser extent, the NH groups at the N terminii of helices provide favourable sites for water interactions, as do reverse turns and regions which connect beta-structure and helices. The hydration shell consists of discontinuous networks of water molecules, the maximum number of molecules in a network being ten. The substrate-binding cleft is heavily hydrated, as is the main loop region which is stabilized by water interactions. The protein molecules are close packed in the crystals with a molecular coordination number of 14. Arginyl residues are extensively involved in intermolecular hydrogen bonds and water bridges. The water molecules in the crystal are organized into discrete clusters. A distinctive feature of the clusters is the frequent occurrence of three-membered rings. The protein molecules undergo substantial rearrangement during the transformation from the native to the low-humidity form. The main-chain conformations in the two forms are nearly the same, but differences exist in the side-chain conformation. The differences are particularly pronounced in relation to Trp 62 and Trp 63. The shift in Trp 62 is especially interesting as it is also known to move during inhibitor binding. PMID- 15299530 TI - Structure determination of aldose reductase: joys and traps of local symmetry averaging. AB - The structure of aldose reductase, a monomeric enzyme of 314 amino acids which crystallizes in space group P1 with four monomers per asymmetric unit, has been solved using a combination of single isomorphous replacement (SIR), solvent flattening and local symmetry averaging. The self rotation showed evidence of 222 local symmetry. The map calculated from the original single isomorphous replacement phases showed a clear solvent envelope but was uninterpretable. A first averaging attempt failed because the molecular envelope obtained from the SIR map weighted with monomer correlation was too small and the averaging was biased by low-resolution truncation. A second attempt with an enlarged envelope and including low-resolution reflections succeeded in refining phases at 3.5 A resolution but failed to extend them correctly. Rigid-body refinement of a partial model based on the 3.5 A map calculated from refined phases showed significant departures from the 222 symmetry. A third averaging attempt using the improved symmetry succeeded in producing a clear map with phases extended to 3.07 A resolution. This map revealed a (beta/alpha)(8) fold, not previously found in NADPH-dependent enzymes. This work shows the importance of mask definition and local symmetry elements accuracy for averaging, and describes a method for improving these parameters. PMID- 15299531 TI - Complex of ribonuclease from Streptomyces aureofaciens with 2'-GMP at 1.7 A resolution. AB - The crystal structure of a complex of ribonuclease from Streptomyces aureofaciens (RNase Sa) with guanosine-2'-monophosphate (2'-GMP) has been refined against synchrotron data recorded from a single crystal using radiation from beamline X31 at EMBL, Hamburg, and an imaging plate scanner. The crystals are in space group P2(1)2(1)2(1) with cell dimensions a = 64.7, b = 78.8 and c = 39.1 A. The structure has two enzyme molecules in the asymmetric unit, complexed with 2'-GMP inhibitor with occupancies of 1 and 2/3 (different to the 3'-GMP complex crystal structure where only one of the two independent RNase Sa molecules binds nucleotide), 492 associated water molecules and one sulfate ion, and was refined using all data between 10.0 and 1.7 A to a final crystallographic R factor of 13.25%. Binding of the base to the enzyme confirms the basis for the guanine specificity but the structural results still do not provide direct evidence of the identity and role of the particular residues involved in the catalytic process. New native RNase Sa data to 1.8 A were recorded to provide a reference set measured under comparable experimental conditions. The crystals are in the same space group and have the same lattice as those of the 2'-GMP complex. The native structure with 423 water molecules was refined in a similar manner to the complex to a final R factor of 13.87%. 1.77 A resolution data were independently measured on a 2'-GMP complex crystal at UCLA using an R-AXIS II image plate scanner mounted on a conventional source. The cell dimensions were essentially the same as above. 2'-GMP was bound more fully to molecule A than to molecule B of the RNase Sa. The structure was refined to an R factor of 14.64% with 388 water molecules. This work follows on from the structure determination of native RNase Sa and its complex with 3'-GMP [Sevcik, Dodson & Dodson (1991). Acta Cryst. B47, 240-253]. PMID- 15299532 TI - Structure at pH 6.5 of ferredoxin I from Azotobacter vinelandii at 2.3 A resolution. AB - Ferredoxin I from Azotobacter vinelandii (AvFdI) is an iron-sulfur protein composed of 106 amino acids, seven Fe atoms and eight inorganic S* atoms. A crystallographic redetermination of its structure showed the originally reported structure to be incorrect. We report here the crystal structure of AvFdI at pH 6.5. Extensive refinement has led to a final R value of 0.170 for all 6986 non extinct reflections in the range 10-2.3 A using a solvent model which includes 98 discrete solvent atoms with occupancies between 0.3 and 1.0 and an average B value of 22.5 A(2). The first half of the peptide chain closely resembles that of the 55-residue ferredoxin from Peptococcus aerogenes (PaFd), while the remainder consists of three turns of helix and a series of loops which form a cap over part of the molecular core. Despite the similarities in structure and surroundings, the corresponding 4Fe4S* clusters in PaFd and AvFdI have strikingly different redox potentials; a possible explanation has been sought in the differing hydration models for the two molecules. PMID- 15299533 TI - High-resolution refinement of the hexagonal A-DNA octamer d(GTGTACAC) at 1.4 A. AB - The hexagonal crystal form of the octamer d(GTGTACAC), grown in the presence of spermine, has unit-cell dimensions a = b = 32.18 and c = 78.51 A, space group P6(1)22, with one DNA strand in the asymmetric unit. The structure has been refined starting with the earlier lower resolution model and using high resolution 1.4 A data collected on a Siemens-Xentronics area detector at 258 K. There were 4365 unique reflections greater than 2sigma(F) in the resolution range 5-1.4 A. The model was refitted into 3F(o) - 2F(c). Sim-weighted omit maps and difference maps were used to locate water molecules. The final model with 161 DNA atoms and 37 water molecules gave an R factor of 19.8%. Crystals of the same octamer were also grown in the presence of spermidine instead of spermine, and refinement using nominal 1.45 A resolution data, 3292 unique reflections, final R = 19.1%, gave virtually identical DNA parameters. No bound spermine or spermidine was detected in either of these structure analyses. The electron density was clear for the DNA and showed holes in the center of the six-membered rings of bases, and also in the center of some of the sugar rings. The high-resolution structure has provided more precise DNA parameters and confirmed the features observed in the earlier 2 A study including the packing-induced distortion in the A7 (A15) sugar pucker from C(3')-endo and C(2')-endo. This change causes the end base pairs to bend away from the helix axis while the rest of the duplex is nearly linear. The hydration patterns in the deep and shallow grooves have been characterized. Chains of water molecules were found, but no rings. The familiar intermolecular contact region between the end base pair and the minor groove of a symmetry-related duplex, involving four residues on one strand and two on the other, has been analyzed. One of these interactions is a hydrogen bond. PMID- 15299534 TI - The mechanism of iron uptake by transferrins: the structure of an 18 kDa NII domain fragment from duck ovotransferrin at 2.3 A resolution. AB - The molecular structure of an iron-containing 18 kDa fragment of duck ovotransferrin, obtained by proteolysis of the intact protein, has been elucidated by protein crystallographic techniques at 2.3 A resolution. This structure supports a mechanism of iron uptake in the intact protein whereby the binding of the synergistic (bi)carbonate anion is followed by binding of the metal with the lobe in the open configuration. These stages are then followed by domain closure in which the aspartic acid residue plays a further key role, by forming an interdomain hydrogen-bond interaction in addition to serving as a ligand to the iron. This essential dual role is highlighted by model building studies on the C-terminal lobe of a known human variant. In this variant a mutation of a glycine by an arginine residue enables the aspartic acid to form an ion pair and reduce its effectiveness for both metal binding and domain closure. The X-ray structure of the 18 kDa fragment strongly suggests that the histidine residue present at the iron binding site of the intact protein and arising from the second interdomain connecting strand has been removed during the preparative proteolysis. PMID- 15299535 TI - Consideration in the choice of a wavelength range for white-beam Laue diffraction. AB - The white-beam Laue-diffraction method is a useful tool for rapid measurement of crystallographic intensities with synchrotron radiation. Considerations of the signal-to-noise ratio to be expected from scattering of X-rays within a limited wavelength range suggest that it will pay to limit that range to something like an octave. This rule-of-thumb has the added advantage that there will be significantly fewer diffraction spots that are overlapping harmonics of one another. To maximize the number of reflections recorded in a single stationary crystal exposure, one should choose this octave of wavelengths in a region where the curvature of the Ewald sphere is greatest, that is at the longest wavelength allowable after other considerations are taken into account. PMID- 15299536 TI - Protein single-crystal diffraction with 5 A synchrotron X-rays at the sulfur K absorption edge. AB - . Sulfur atoms, an integral part of many proteins, are possible candidates for anomalous scattering in phase determination by multiple-wavelength methods. The main difficulty encountered is that a wavelength of about 5 A is required to obtain a large anomalous signal from these atoms, leading to very large absorption effects. Initial experiments have been carried out using a synchrotron X-ray source, evacuated beam tubes, a diffractometer inside a vacuum chamber, a special sample holder and a suitable scattering geometry. The results are encouraging, showing that Bragg reflections can be measured, and that changes in their intensities around the absorption edge are observable. PMID- 15299539 TI - Phasing macromolecular structures via structure-invariant algebra. AB - Owing to the breakdown of Friedel's law when anomalous scatterers are present, unique values of the three-phase structure invariants in the whole range from 0 to 2pi are determined by measured values of diffraction intensities alone. Two methods are described for going from presumed known values of these invariants to the values of the individual phases. The first, dependent on a scheme for resolving the 2pi ambiguity in the estimate omega(HK) of the triplet phi(H) + phi(K) + phi(-H-K), solves by least squares the resulting redundant system of linear equations phi(H) + phi(K) + phi(-H-K) = omega(HK). The second attempts to minimize the weighted sum of squares of differences between the true values of the cosine and sine invariants and their estimates. The latter method is closely related to one based on the 'minimal principle' which determines the values of a large set of phases as the constrained global minimum of a function of all the phases in the set. Both methods work in the sense that they yield values of the individual phases substantially better than the values of the initial estimates of the triplets. However, the second method proves to be superior to the first but requires, in addition to estimates of the triplets, initial estimates of the values of the individual phases. PMID- 15299540 TI - On the application of phase relationships to complex structures. XXXII. A small protein at low resolution. AB - The direct-methods program SAYTAN is applied to data at various restricted resolutions for a small protein. It is shown that useful sets of phases can be obtained even down to 3 A resolution. Conventional figures of merit are not very discriminating for the phase sets developed, but modified figures of merit seem capable of selecting the better phase sets, at least for those generated from 2 A or higher resolution data. PMID- 15299541 TI - On the application of phase relationships to complex structures. XXXIII. The problems with large structures and low resolution. AB - A conventional direct method, using the Sayre equation as a basis, has been shown to be capable of solving a small protein with data of 3.0 A resolution or better. An analysis of the Sayre equation, with data of various resolutions and with different lower limits of |E| for the contributors in the summation, shows that its effectiveness for phasing is independent of structural complexity but does decline as the resolution becomes worse. It is suggested that a practicable lower limit for the application of conventional direct methods is about 3.5 A. For large macromolecular structures the number of contributors to the summation in the Sayre equation becomes too large to handle and it is suggested that real space methods should be used instead. PMID- 15299542 TI - The application of direct methods and Patterson interpretation to high-resolution native protein data. AB - Conventional small-molecule methods of solving the phase problem from native data alone, without the use of heavy-atom derivatives, known fragment geometries or anomalous dispersion, have been tested on 0.9 A resolution data for two small proteins: rubredoxin, from Desulfovibrio vulgaris, and crambin. The presence of three disulfide bridges in crambin and an FeS(4) unit in rubredoxin enabled automated Patterson interpretation as well as direct methods to be tried. Although both structures were already well established, the known structures were not used in the phasing attempts, except for identifying successful solutions. Direct methods were not successful for crambin, although the correct phases were stable to phase refinement and gave figures of merit clearly superior to any obtained in the ca 500 000 random starting phase sets that were refined. It appears that the presence of an iron atom in rubredoxin reduces the scale of the search problem by many orders of magnitude, but at the cost of producing 'over consistent' phase sets that overemphasize the iron atom and involve partial loss of enantiomorph information. However, about 1% of direct-methods trials were successful for rubredoxin, giving mean phase errors of about 56 degrees (for all E > 1.2) that could be reduced to about 20 degrees by standard E-Fourier recycling methods. Limiting the resolution of the data degraded the quality of the solutions and suggested that the limiting resolution for routine direct methods solution of rubredoxin is about 1.2 A. With the 0.9 A data, automated Patterson interpretation convincingly finds the three disulfide bridges in crambin and the FeS(4) unit in rubredoxin, and in both cases E-Fourier recycling starting from these 'heavier' atoms yields almost the complete structure. Whereas crambin could only be solved in this way at very high resolution, rubredoxin could be solved by Patterson interpretation down to 1.6 A. These results emphasize the benefits of collecting protein data to the highest possible resolution, and indicate that when a few 'heavier' atoms are present, it may prove possible in favorable cases to solve the phase problem from a single native data set collected to 'atomic resolution'. PMID- 15299543 TI - Assessment of phase accuracy by cross validation: the free R value. Methods and applications. AB - Analogies between the free R statistic [Brunger (1992). Nature (London), 355, 472 474] and the statistical methods of cross validation and bootstrap are discussed. Several new applications which make use of the previously observed correlation between the free R value and the phase accuracy of crystal structures are presented. One application concerns the relative weighting of individual restraint classes in macromolecular refinement. The free R value provides an objective statistical basis for the optimal choice of the weights. The results for the refinement of a penicillopepsin crystal structure at 1.8 A resolution indicate that overall bond length and bond angle weights, derived from uncertainties observed in small-molecule crystal structures, appear to be transferable to macromolecules. In another application, the landscape of the R value around the crystal structure was investigated for unrestrained modeling of diffraction data with equal atomic scatterers. Others have suggested applications to ab initio phasing because of the simplicity of the liquid-like system of equal atomic scatterers. However, there are a large number of incorrect configurations of the scatterers whose R values at 1.8 A resolution are close to that of the correct configuration given by the positions of the non-hydrogen atoms in the penicillopepsin crystal structure. A substantial number of the incorrect configurations have higher free R values than the correct one. It is therefore conceivable that the free R value could be used as a selection criterion to distinguish between certain incorrect configurations and configurations close to the correct one. PMID- 15299544 TI - Direct phase determination by entropy maximization and likelihood ranking: status report and perspectives. AB - A new multisolution phasing method based on entropy maximization and likelihood ranking, proposed for the specific purpose of extending probabilistic direct methods to the field of macromolecules, has been implemented in two different computer programs and applied to a wide variety of problems. The latter comprise the determination of small crystal structures from X-ray diffraction data obtained from single crystals or from powders, and from electron diffraction data partially phased by image processing of electron micrographs, the ab initio generation and ranking of phase sets for small proteins; and the improvement of poor quality phases for a larger protein at medium resolution under constraint of solvent flatness. These applications show that the primary goal of this new method - namely increasing the accuracy and sensitivity of probabilistic phase indications compared with conventional direct methods - has been achieved. The main components of the method are (1) a tree-directed search through a space of trial phase sets; (2) the saddle-point method for calculating joint probabilities of structure factors, using entropy maximization; (3) likelihood-based scores to rank trial phase sets and prune the search tree; (4) efficient schemes, based on error-correcting codes, for sampling trial phase sets; (5) a statistical analysis of the scores for automatically selecting reliable phase indications. They have been implemented to varying degrees of completeness in a computer program (BUSTER) and tested on two small structures as well as on the small protein crambin. The main obstructions to successful ab initio phasing in the latter case seem to reside in the accumulation of phase sampling errors and in the lack of a properly defined molecular envelope, both of which can be remedied within the methods proposed. A review of the Bayesian statistical theory encompassing all phasing procedures, proposed earlier as an extension of the initial theory, shows that the techniques now available in BUSTER bring closer a number of major enhancements of standard macromolecular phasing techniques, namely isomorphous replacement, molecular replacement, solvent flattening and non-crystallographic symmetry averaging. The gradual implementation of the successive stages of this 'Bayesian programme' should lead to an increasingly integrated, effective and dependable phasing procedure for macromolecular structure determination. PMID- 15299545 TI - Construction of maximum-entropy density maps, and their use in phase determination and extension. AB - Methods for constructing everywhere-positive electron-density maps with Fourier amplitudes matching those for arbitrarily large sets of observed data, utilizing dual-function methods for maximization of entropy, are described. Possible strategies for utilizing these maps for the determination and extension of phases in macromolecular structure determination are suggested, and problems are discussed. PMID- 15299546 TI - Experience with phase extension and ab initio phase determination in macromolecular crystallography using maximum-entropy methods. AB - Three procedures, or 'tools', have been developed and tested for applying maximum entropy methods to phase extension and to ab initio phase determination. The phase expander tool has been used in connection with the solution of two previously unknown macromolecular structures. An efficient algorithm for the determination of an electron-density distribution that is everywhere positive and that agrees with observed structure amplitudes (tools II and III) has been used to determine the phases of X-ray diffraction data from recombinant bovine chymosin, a protein with 323 amino-acid residues in the molecular chain, the structure of which was recently determined using replacement methods. By use of the same maximum-entropy methods, the structure amplitudes from the unknown structure of bovine heart creatine kinase, a protein with 381 amino-acid residues, have been phased ab initio to 2.7 A resolution. The phases of the centric reflections have also been confirmed by a satisfactory solution of the Patterson map of a mercury derivative. The current status of the structure interpretation is presented. This technique has also been applied to a test case where 48 centric reflections from bovine prothrombin fragment 1 data were phased ab initio and subsequently used in the determination of Patterson solutions for a heavy-atom derivative data set. PMID- 15299547 TI - Entropy phase dynamics. AB - The entropy-dynamics method seeks maxima for the entropy of the electron density for N atoms in a crystal cell, when the Fourier amplitudes are fixed, but their phases are unknown. By analogy with molecular dynamics, the effective potential energy is the negative entropy V = -NS. The kinetic energy is proportional to the squared velocities of the electron densities at grid points in the map. It reduces to a sum of Fourier-mode rotor energies. Each rotor angle experiences a couple equal to the phase gradient of S, and local dynamical equilibrium yields a Boltzmann distribution of S. Discrete phase angles (e.g. signs) are treated as quantized rotor modes. The distributions depend on a popularity function of the entropy histogram. Trial calculations have been made of phase averages and correlations in a centrosymmetric projection of the membrane protein bacteriorhodopsin. The maximum-entropy solution and the correct solution do not always coincide. PMID- 15299548 TI - Entropy on charge density: making the quantum mechanical connection. AB - Entropy maximization has proven effective in treating certain aspects of the phase problem of X-ray diffraction. Much of its development has been expressed in probabilistic language, although image enhancement has been somewhat more physical or geometric in description. Here phasing and entropy maximization are embedded in the quantum mechanical problem of reconstructing an electronic one matrix under experimental constraints. Entropy on an N-representable one-particle density matrix is well defined. The entropy is the expected form, and it is a simple function of the one-matrix eigenvalues which all must be non-negative. Certain other properties are pertinent to phasing which is implicit in one-matrix reconstruction governed by entropy maximization. Throughout this work reference is made to informational entropy, not the entropy of thermodynamics. PMID- 15299549 TI - Electron-density histograms and the phase problem. AB - The specific properties of the range of possible electron-density values may serve as useful additional information for the determination and refinement of structure-factor phases. Fourier synthesis histograms (showing the spectra of frequencies of different possible values) produce the most adequate representation of these properties. The mathematical background and practical uses of these histograms are discussed. The investigation provides new information on some traditional methods of phase refinement, including density modification procedures, maximization of Cochran's integral value and other techniques. PMID- 15299550 TI - Programs for phasing by entropy maximization as implemented in Xtal3.2: a crystallographic software system. AB - Xtal3.2, a crystallographic software package, is an international development project involving about 40 researchers over a full spectrum of crystallographic interests. This development has been supported by many national and international agencies and commercial institutions since the first version in 1983. The 1992 release, Xtal3.2, contains software for 95 different calculations. These range from the processing of raw diffraction data to interactive molecular graphics, atomic charge estimation, electronic publication preparation, and the structure solution and refinement of small and large molecules. Tests of the Xtal programs for phase determination and phase refinement by the application of 'maximum entropy' are presented. PMID- 15299552 TI - Low-resolution real-space envelopes: improvements to the condensing protocol approach and a new method to fix the sign of such envelopes. AB - The condensing-protocol strategy for obtaining low-resolution real-space envelopes of macromolecules from native Fourier intensities alone was recently reported [Subbiah (1991). Science, 252, 128-133]. The present work introduces three improvements to the original methodology. In terms of the j index, which was defined as the crude ratio of correctly placed scatterers to those incorrectly placed, these changes collectively contribute to j values that range from 8 to 28 and in one case even infinity. This compares with the previously reported values of 1.9 and 2.6. Nevertheless, under different conditions, both the original and the current improved condensing protocol can preferentially converge on the solvent or the matter region. Since such control is not absolute there still remains a certain amount of ambiguity in deciding whether the congealed gas represents bulk solvent or bulk matter. To overcome this ambiguity, a simple method based on the systematic placement of point scatterers on a grid lattice within and without the envelope is presented. This 'sign-fixing' method is shown to work with both perfect envelopes as well as the cruder envelopes produced by the improved condensing protocol. The new methodology is illustrated using four macromolecular examples. PMID- 15299553 TI - Towards the measurement of ideal data for macromolecular crystallography using synchrotron sources. AB - Synchrotron radiation has been used extensively to overcome a variety of technical challenges involved in data collection from macromolecular crystals. The next generation of such sources offer a higher brilliance at much shorter wavelengths than hitherto available. Hence, the quality of X-ray diffraction data from crystals of biological macromolecules will be further improved in terms of reduced systematic and random errors, in conjunction with a very high degree of completeness of, and multiple measurements within, the data set. Real data sets should be able to approach closely the quality of ideal data sets. Tests at CHESS are described of the feasibility of recording protein crystal diffraction patterns at ultra-short wavelengths (lambda = 0.3 A) and very-short wavelengths (lambda = 0.5 A), in monochromatic rotating crystal geometry. PMID- 15299554 TI - Automated refinement of protein models. AB - An automated refinement procedure (ARP) for protein models is proposed, and its convergence properties discussed. It is comparable to the iterative least-squares minimization/difference Fourier synthesis approach for small molecules. ARP has been successfully applied to three proteins, and for two of them resulted in models very similar to those obtained by conventional least-squares refinement and rebuilding with FRODO. In real time ARP is about ten times faster than conventional refinement. In its present form ARP requires high (2.0 A or better) resolution data, which should be of high quality and a starting protein model having about 75% of the atoms in roughly the correct position. For the third protein at 2.4 A resolution, ARP was significantly less powerful but nevertheless gave definite improvement, in the density map at least. PMID- 15299555 TI - Improvement of macromolecular electron-density maps by the simultaneous application of real and reciprocal space constraints. AB - A general scheme for the improvement of electron-density maps is described which combines information from real and reciprocal space. The use of Sayre's equation, solvent flattening and histogram matching within this scheme has been described previously [Main (1990). Acta Cryst. A46, 372-377]. Non-crystallographic symmetry averaging, the use of a partial structure and constraints on individual structure factors have now been added. A computer program, SQUASH, is described which applies all these constraints simultaneously. Its application to the maps of several structures has been successful, particularly so when non-crystallographic symmetry is present. Uninterpretable maps have been improved to the point where a significant amount of the structure can be recognized. Applying the constraints simultaneously is more powerful than applying them all in series. PMID- 15299556 TI - Ab initio direct methods: practical advice for getting beyond the first 300 atoms. AB - Not all crystallographic structural investigations are amenable to a phasing solution by direct methods alone. Guideline procedures are outlined which are intended to help the evaluation of whether direct-methods procedures may be expected to phase diffraction data for large molecular structures. This analysis is directed at three separate levels of inquiry: (1) How good are the primary data and can E values be derived to represent a point-atom structure. (2) How well do the data interact through phase relationships and may they be expected to produce a stable phasing solution. (3) What is the prognosis for finding recognizable solutions. Data are presented from the post-mortem analyses of a number of large, difficult-to-solve, structures to illustrate each of these points. Direct-methods practitioners are to be encouraged that crystal structures having more than 300 atoms per asymmetric unit may occasionally be determined utilizing present methodologies provided that an a priori prognosis for obtaining a solution is favorably high, adequate computational resources are available, and sufficient persistent effort is applied. PMID- 15299557 TI - Molecular scene analysis: the integration of direct-methods and artificial intelligence strategies for solving protein crystal structure. AB - A knowledge-based approach to crystal structure determination is presented. The approach integrates direct-methods and artificial-intelligence strategies to rephrase the structure determination process as an exercise in scene analysis. A general joint probability distribution framework, which allows the incorporation of isomorphous replacement, anomalous scattering and a priori structural information, forms the basis of the direct-methods strategies. The accumulated knowledge on crystal and molecular structures is exploited through the use of artificial-intelligence strategies, which include techniques of knowledge representation, search and machine learning. PMID- 15299558 TI - Application of the minimal principle to peptide structures. AB - A new direct-methods procedure has been devised which consists of phase refinement via the minimal function, R(phi), alternated with Fourier summation and real space filtering. All phases are initially assigned values by computing structure factors for a randomly positioned set of atoms. These phases are then refined by using a parameter shift method to minimize R(phi). The refined phases are Fourier transformed, and a specified number of the largest peaks in the electron-density function are found and used as a new trial structure. The probability of a trial structure converging to a solution appears to depend on structural complexity and a number of refinement parameters. This procedure shows potential for providing fully automatic routine solutions for structures in the 200-400 atom range. PMID- 15299559 TI - The effect of noise on entropy. AB - A complementary relationship between the entropy (S) and the variance (sigma(2)) of an electron-density map is derived by approximating the logarithmic term in the entropy expression by a series expansion around the average map density. The resulting expression is S approximately ln N - 1/2sigma(2), where N is the number of grid points and sigma is the r.m.s. deviation from the mean in a map normalized to unit mean. The algebraic expression is of interest because it is consistent with and allows numerical evaluation of the surprising argument that noise decreases the entropy of a map. The argument is that a noise contribution by itself generates a certain variance that is independent of the atomic structure and that adds to the variance due to the structure. Increased variance corresponds to decreased entropy. This property of noise provides an intuitively reasonable justification for maximizing the entropy of an electron-density map in the quest for more readily interpretable maps of macromolecules. The entropy variance relationship also extends the range of applicability of the entropy concept to maps with a limited amount of negative density. The approximation which leads to the entropy-variance relationship is most applicable where it is most likely to be useful - in experimental maps of relatively low structure definition. PMID- 15299560 TI - Uniqueness and the ab initio phase problem in macromolecular crystallography. AB - The crystallographic phase problem is indeterminate in the absence of additional chemical information. A successful ab initio approach to the macromolecular phase problem must employ sufficient chemical constraints to limit the solutions to a manageably small number. Here we show that commonly employed chemical constraints - positivity, atomicity and a solvent boundary - leave the phase problem greatly underdetermined for Fourier data sets of moderate (2.5-3.0 A) resolution. Entropy maximization is also beset by multiple false solutions: electron-density maps are readily generated which satisfy the same Fourier amplitude constraints but have higher entropies than the true solution. We conclude that a successful ab initio approach must make use of high-resolution Fourier data and/or stronger chemical constraints. One such constraint is the connectivity of the macromolecule. We describe a rapid algorithm for measuring the connectivity of a map, and show its utility in reducing the multiplicity of solutions to the phase problem. PMID- 15299561 TI - Entropy maximization constrained by solvent flatness: a new method for macromolecular phase extension and map improvement. AB - A practical generally applicable procedure for exponential modeling to maximum likelihood of macromolecular data sets constrained by a moderately large basis set of reliable phases and a molecular envelope is described, based on the computer program MICE [Bricogne & Gilmore (1990). Acta Cryst. A46, 284-297]. Procedures were first tested with simulated data sets. Exact and randomly perturbed amplitudes and phases were generated, together with a known envelope for solvent-free protein and for protein in an electron-dense crystal mother liquor typical of many real protein crystals. These experiments established useful guidelines and values for various parameters. Tests with basis sets chosen from the largest amplitudes indicate that exponential models with considerable correct extrapolated phase and amplitude information can be constructed from as few as 16% of the total number of reflections, with mean phase errors of about 30 degrees, at resolution limits of either 5 or 3 A. When the shape of the solvent channels in macromolecular crystals is known, it offers an important additional source of information. MICE was, therefore, adapted to average the density outside the molecular boundary defined by an input envelope. This flattening process imposes a uniform density distribution in solvent-filled channels as an additional constraint on the exponential model and is analogous to the treatment of solvent in conventional solvent flattening. Experimental data for cytidine deaminase, a structure recently solved by making extensive use of conventional solvent flattening, provides an example of the performance of maximum-entropy methods in a real situation and a compelling comparison of this method to standard procedures. Exponential models of the electron density constrained by the most reliable phases obtained by multiple isomorphous replacement with anomalous scattering (MIRAS) (figure of merit > 0.7, representing 34% of the total number of reflections) and by the envelope give rise to centroid electron density maps which are quantitatively superior by numerous statistical criteria to conventionally solvent-flattened density. Similarity of these maps to the 2F(obs) - F(calc) map calculated with phases obtained after crystallographic refinement of the model implies that maximum-entropy extrapolation provides better phases for the remaining 66% of the reflections than the original centroid MIRAS distributions. Importantly, the solvent-flattened electron density, although it did permit interpretation of the map which was not readily accomplished with the MIRAS map, contains substantial errors. It is proposed that errors of this sort may account for previously noted deficiencies of the solvent flattening method [Fenderson, Herriott & Adman (1990). J. Appl. Cryst. 23, 115 131] and for the occasional tendency of incorrect interpretations to be 'locked in' by crystallographic refinement [Branden & Jones (1990). Nature (London), 343, 687-689, and references cited therein]. Solvent flattening with combined maximization of entropy and likelihood represents a phase-refinement path independent of atomic models, using the experimental amplitudes and the most reliable phases. It should, therefore, become a valuable and generally useful procedure in macromolecular crystal structure determination. PMID- 15299562 TI - SQUASH - combining constraints for macromolecular phase refinement and extension. AB - The constraints of correct electron-density distribution, solvent flatness, correct local shape of the electron density and equal molecules are combined in an integrated procedure for macromolecular phase refinement and extension. These constraints on electron densities are satisfied simultaneously by solving a system of non-linear equations. The electron-density solution is further filtered by a phase combination procedure. The non-crystallographic symmetry operations are refined by a rotation and translation space search and a least-squares minimization method, thereby reducing the chance of introducing systematic phase errors during averaging. The effect of each constraint on phase refinement and extension is examined. The constraints are found to work synergistically in phase improvement. Test results on 2Zn insulin are presented. PMID- 15299563 TI - Structure solution of C-reactive proteins: molecular replacement with a twist. AB - The pentameric structure of C-reactive proteins (CRP) has been derived by a combination of automated and manual molecular-replacement techniques. The method is generally applicable to other multimeric assemblies. The highly homologous human serum amyloid P component (hSAP) structure fails to provide a pentameric molecular-replacement solution for CRP. In the absence of a significant signal from an individual protomer, the hSAP structure has been manually modified in terms of protomer assembly to provide the true pentameric model of CRP. The CRP protomers are rotated or twisted by 14 degrees about an axis, through the protomer centre, which is approximately perpendicular to the pentamer radius and the molecular fivefold axis. The results demonstrate clearly that protomers with very similar folds arising from high sequence homology need not necessarily be assembled together in the same way although the symmetry of the resulting oligomer may be maintained. In a curious twist the CRP structure which provided the general CRP model remains unsolved, while the model itself has so far provided the solution of two other CRP structures. PMID- 15299564 TI - Pitfalls of molecular replacement: the structure determination of an immunoglobulin light-chain dimer. AB - The structure of protein Cle, a human light-chain dimer from the lambdaIII subgroup, was determined using 2.6 A data; the R value is 18.4%. The structure was solved, after a false start, by molecular replacement with the lambdaII/V Mcg protein as a search structure. When the refinement did not proceed beyond an R value of 27%, it was discovered that while the constant domains were in their correct positions in the unit cell, the incorrect variable domains were used for defining the molecule. The correct solution required a rotation of 180 degrees around the local twofold axis that relates the two constant domains of the dimer. The correct variable domain positions overlap about 70% of the same volume as the incorrect ones of a symmetry-related molecule. The refinement distorted the geometries of the domains. Though the constant domains were in their correct positions, the r.m.s. (root-mean-square) deviation of the Calpha atom position was 1.2 A when the two constant domains were compared. For the correct structure, this value is 0.5 A. The phi and psi angles, the r.m.s. chiral value and the free R value, even when calculated a posteriori, were good indicators of the correctness of the structure. The quaternary structure of the Cle molecule is similar to that in Mcg (crystallized from ammonium sulfate); the elbow bend is 115 degrees. However, the arrangement of the variable domains differs from that observed in other variable domain dimers. The variable domains of Cle are 0.7 A closer than in Mcg or variable dimer Rei. The hydrogen bonding at the interface of the two domains is novel. Residues Tyr36 from both monomers form a hydrogen bond that is part of a network with the Gln89 residues from both monomers. For the first time hydrogen bonds were observed between the main-chain peptide N and O atoms of the complementarity-determining region CDR2 and CDR3 segments of both monomers. PMID- 15299565 TI - An X-ray analysis of native monoclinic lysozyme. A case study on the reliability of refined protein structures and a comparison with the low-humidity form in relation to mobility and enzyme action. AB - The atomic models of native monoclinic lysozyme obtained by refinement at Bangalore and elsewhere [Young, Dewan, Nave & Tilton (1993). J. Appl. Cryst. 26, 309-319] differed significantly in the flexible regions of the protein molecule. The two models were reconciled starting from regions where they were in reasonable agreement to produce an improved model which yielded an R value of 0.169 for 12 816 observed reflections in the 10-2 A resolution range. The reconciled model was compared with the structure of the 88% relative humidity form obtained through a water-mediated transformation [Madhusudan, Kodandapani & Vijayan (1993). Acta Cryst. D49, 234-245]. Parts of the flexible regions of the molecule register significant movements during the transformation. The changes resulting from the transformation from the native to the low-humidity forms are pronounced in many of the side chains in the active-site region, thus indicating the relationship between hydration, mobility and enzyme action. The fact that the overall changes in molecular geometry resulting from water-mediated transformation are similar to those which occur during enzyme action, further emphasizes this relationship. PMID- 15299566 TI - A minimalist's approach to the phase problem - phasing selenomethionyl protein structures using Cu Kalpha data. AB - The feasibility of phasing protein structures through the use of the isomorphous and anomalous signal of selenomethionyl (Se-Met) derivative and diffraction data collected with a standard laboratory Cu Kalpha X-ray source has been investigated. Interpretable electron-density maps were obtained for the core domain of avian sarcoma virus integrase, a typical medium-sized protein having four Met residues in a sequence of 156 amino acids. The r.m.s. difference between 3.1 A experimental phases obtained from Se-Met Cu Kalpha data and the final phases calculated from the refined model is 55 degrees. A procedure combining single isomorphous replacement/single anomalous scattering phasing and solvent flattening for data based on a single Se-Met derivative and Cu Kalpha radiation has been tested on this and another protein. The results are encouraging enough to indicate that such procedures might be recommended when a synchrotron source is not readily available. PMID- 15299567 TI - Refined high-resolution structure of the metal-ion dependent L-fuculose-1 phosphate aldolase (class II) from Escherichia coli. AB - The structure of the class II zinc-ion dependent L-fuculose-1-phosphate aldolase from Escherichia coli in its tetragonal crystal form has been established at 1.92 A resolution. The homotetrameric enzyme has a molecular mass of 4 x 24 kDa and follows C(4) symmetry. The structure model is exactly symmetrical, which contradicts an observed birefringence anomaly of the crystals. The four catalytic centers are located in deep clefts at the interfaces of adjacent subunits. The zinc ion is coordinated by three histidines and one glutamate in an almost tetrahedral arrangement. In contrast to numerous other catalytically competent zinc ions, there is no water molecule in the ligand sphere. Replacement of zinc by a cobalt ion caused only small structural changes. A search through the Protein Data Bank indicated that the chain fold is novel. Sequence homology searches revealed a significant similarity to the bacterial L-ribulose-5 phosphate 4-epimerase. PMID- 15299568 TI - Approaches to very low resolution phasing of the ribosome 50S particle from Thermus thermophilus by the few-atoms-models and molecular-replacement methods. AB - Estimates for the phases of the X-ray diffraction data from the 50S ribosomal particle of Thermus thermophilus has been made to an effective resolution around 80 A using the few-atoms-modes ab initio technique [Lunin, Lunina, Petrova, Vernoslova, Urzhumtsev & Podjarny (1995). Acta Cryst. D51, 896-903]. This technique models the density with a small number of Gaussian spheres to generate a large number of possible phase sets and then uses clustering algorithms to identify the best ones. Independently, an envelope obtained from electron micrograph image reconstruction [Yonath, Leonard & Wittmann (1987). Science, 236, 813-816] was oriented and positioned using the molecular-replacement technique, specially adapted to the very low resolution case [Urzhumtsev & Podjarny (1995). Acta Cryst. D51, 888-895]. The two methods show similar packing arrangements. The electron density calculated by the few-atoms-models technique without any assumption on the number of molecules in asymmetric unit or on their shape shows recognizable features of the particle. PMID- 15299569 TI - The rate of water equilibration in vapor-diffusion crystallizations: dependence on the distance from the droplet to the reservoir. AB - The rate of water equilibration in hanging-drop vapor-diffusion experiments was studied as a function of the distance separating the hanging drop from the surface of the reservoir solution. Hanging drops of 1.00 M NaCl were allowed to partially equilibrate with reservoirs of 2.00 M NaCl at room temperature. Over the range of droplet-reservoir distances examined, 7.6-119.4 mm, the larger the distance that separated the droplet and reservoir, the slower the droplet equilibrated with the reservoir. The variation of the rate of equilibration with droplet-reservoir distance was non-linear; the rate was most sensitive to variations in the droplet-reservoir separation at short separations. A mathematical model of the equilibration kinetics was developed that fits the experimental data. The model is based on the assumption that the rate-limiting step in vapor-diffusion equilibration is transit of water across the vapor space. A simple device to vary the rate of water equilibration, and thereby optimize macromolecular crystal growth conditions, is described. PMID- 15299570 TI - Structure of the Fab fragment of a monoclonal antibody specific for carcinoembryonic antigen. AB - The three-dimensional structure of the Fab fragment of the murine monoclonal antibody A5B7, which is specific for carcinoembryonic antigen (CEA) a protein expressed on carcinoma cell surfaces, has been determined. The structure was solved by molecular replacement and has been refined to an R factor for 21.2% (all data 8-2.1 A). The conformation of the hypervariable loops, which form the antigen binding site, are consistent with canonical loop predictions. Hypervariable loop H3 is unusual in surface exposing many hydrophobic groups at the expense of burying an aspartic acid in the protein core. Other regions of the structure that influence the conformation of the binding site are identified. This structure forms a basis for analysing the effects of amino-acid substitutions in both hypervariable and framework regions in engineering studies of the A5B7 antibody. PMID- 15299571 TI - Direct-space methods in phase extension and phase refinement. V. The histogram moments method. AB - Any distribution is completely defined by its moments. It is shown that a process of phase refinement can be carried out, based on Fourier transforms, which modifies the moments of electron density, separately in the protein and solvent regions, towards target values. Tests have been carried out on two moderate-sized proteins with 800-900 atoms in the asymmetric unit, one containing heavy atoms and the other not. It has been found that refinement using the third moment about zero in the protein region is most effective and that refinement with higher moments, or in the solvent region, adds nothing useful. Two kinds of weights are necessary in the method. One is for giving a weighted mixture of new phase indications with original phase estimates from, say, multiple isomorphous replacement. The other weights are applied to the Fourier coefficients of density maps to give the best possible signal:noise ratio. These weights have been explored empirically and the best ones found are described. It is concluded that since the moments method, which changes phases in reciprocal space, is independent of other histogram-matching procedures, which change density in real space, it has something to offer in a refinement package containing several procedures. PMID- 15299572 TI - Direct-space methods in phase extension and phase refinement. VI. PERP (phase extension and refinement program). AB - Several techniques for extending and refining phases for macromolecular structures have been incorporated into a program package PERP. In addition to previously employed techniques such as solvent flattening and histogram matching, PERP includes a new way of applying the Sayre equation [Refaat, Tate & Woolfson (1995). Acta Cryst. D51, 1036-1040], low-density elimination [Shiono & Woolfson (1992). Acta Cryst. A48, 451-456] and two double-histogram methods [Refaat, Tate & Woolfson (1996). Acta Cryst. D52, 252-256]. PERP is an easy-to-use package controlled by keywords and provided with default parameters that usually give near-optimum results. Examples are given of refinement, and also extension and refinement, for six known protein structures with a variety of characteristics. In each case PERP gives a very satisfactory outcome as measured by improvements in the mean-phase-error and conventional map-correlation coefficient. The main conclusion is that the several methods used in sequence give more effective extension and refinement than using any single method alone. PMID- 15299573 TI - Subtilisin BPN' at 1.6 A resolution: analysis for discrete disorder and comparison of crystal forms. AB - The three-dimensional structure of the serine protease subtilisin BPN' (SBT) has been refined at 1.6 A resolution in space group C2 to a final R value of 0.17. 17 regions of discrete disorder have been identified and analyzed. Two of these are dual-conformation peptide units; the remainder involve alternate rotamers of side chains either alone or in small clusters. The structure is compared with previously reported high-resolution models of SBT in two other space groups, P2(1)2(1)2(1) and P2(1). Apart from the surface, there are no significant variations in structure among the three crystal forms. Structural variations observed at the protein surface occur predominantly in regions of protein-protein contact. The crystal packing arrangements in the three space groups are compared. PMID- 15299574 TI - X-ray structure of cyclodextrin glucanotransferase from alkalophilic Bacillus sp. 1011. Comparison of two independent molecules at 1.8 A resolution. AB - Cyclodextrin glucanotransferase (CGTase) is an enzyme which produces cyclodextrins by the degradation of starch. The enzyme from alkalophilic Bacillus sp. 1011, consisting of 686 amino acid residues, was crystallized from the solution containing 20% PEG 3000 and 20% 2-propanol at pH 5.6 adjusted with citrate buffer. The space group was P1 and the unit cell contained two molecules (V(m) = 2.41 A(3) Da(-1)). The structure was solved by the molecular replacement method and refined to a conventional R value of 0.161 (R(free) = 0.211) for the reflections in the resolution range 1.8-10 A by energy minimization combined with simulated annealing. The molecule consists of five domains, designated A-E, and its backbone structure is similar to the structure of other bacterial CGTases. The molecule has two calcium binding sites where calcium ions are coordinated by seven ligands, forming a distorted pentagonal bipyramid. The two independent molecules are related by a pseudotwofold symmetry and are superimposed with an r.m.s. deviation value of 0.32 A for equivalent C(alpha) atoms. Comparison of these molecules indicated the relatively large mobility of domains C and E with respect to domain A. The active site is filled with water molecules forming a hydrogen-bond network with polar side-chain groups. Two water molecules commonly found in the active center of both molecules link to several catalytically important residues by hydrogen bonds and participate in maintaining a similar orientation of side chains in the two independent molecules. PMID- 15299575 TI - X-ray structure solution of amaryllis lectin by molecular replacement with only 4% of the total diffracting matter. AB - It is often the case that analogous proteins from different species crystallize in a different form. These structures can usually be easily solved by the molecular-replacement (MR) technique, as the protein folding is very often conserved. However, the results from MR become more uncertain as the proportion of diffracting matter decreases as a result of multimericity and/or absence of some of the atoms in the model. In this paper results are presented on the structure solution of amaryllis lectin (109 residues per monomer) containing two protein molecules in the asymmetric unit. The structure was solved by MR using the Calpha coordinates of one monomer from snowdrop lectin which has 85% amino acid sequence identity to amaryllis lectin. This represents only 6% of the non-H atoms of the protein molecule to be used for structure determination and it is a major improvement on previous reports. Further calculations were carried out in order to establish the minimum number of atoms which could be included in the model before a clear solution to the MR problem was revealed. This study showed that the structure of amaryllis lectin could still have been solved easily with 3.85% of the model, which even in the most favourable cases, will probably constitute a minimum for molecular-replacement structure solution. PMID- 15299576 TI - Molecular structure of a proteolytic fragment of TLP20. AB - Myosin light-chain kinase is responsible for the phosphorylation of myosin in smooth muscle cells. In some tissue types, the C-terminal portion of this large enzyme is expressed as an independent protein and has been given the name telokin. Recently, an antibody directed against telokin was found to interact with a protein derived from the baculovirus Autographa californica nuclear polyhedrosis virus. This protein was biochemically characterized and given the name TLP20 for telokin-like protein of 20 000 molecular weight. The amino-acid sequence of TLP20 was determined on the basis of a cDNA clone and subsequent alignment searches failed to reveal any homology to telokin or to other known proteins. The three-dimensional structure of a proteolytic portion of TLP20 is reported here. Crystals employed in the investigation were grown from ammonium sulfate solutions at pH 6.0 and belonged to the space group P2(1)3 with unit-cell dimensions of a = b = c = 76.3 A and one molecule per asymmetric unit. The structure was determined by multiple isomorphous replacement with three heavy atom derivatives. Least-squares refinement of the model reduced the crystallographic R factor to 18.1% for all measured X-ray data from 30.0 to 2.2 A. The overall fold of the molecule may be described as a seven-stranded antiparallel beta-barrel flanked on the bottom by two additional beta-strands and on the top by an alpha-helix. Quite surprisingly, the three-dimensional structure of this beta-barrel is not similar to telokin or to any other known protein. PMID- 15299577 TI - Atomic resolution structure of concanavalin A at 120 K. AB - The structure of native concanavalin A has been refined to a resolution of 1.2 A against data collected at 120 K. The space group is I222, with a = 61.954 (8), b = 86.053 (11), c = 89.079 (11) A. The structure was refined by restrained weighted least-squares minimization of sum w(F(o)(2) - F(c)(2)(2) with SHELXL92/3/6. The final model contains all of the atoms from 237 amino acids, two metal ions and 271 water molecules spread over 287 sites. Disorder is modelled over two conformations for 30 amino-acid side chains. The final weighted R index on F(2) (wR(2)) on all data was 30.4%. Conventional R indices based on F were 14.2 and 11.8% for all data and for data with F > 4sigma(F), respectively. PMID- 15299578 TI - Labelling of thin filaments by Myosin heads in contracting and rigor vertebrate skeletal muscles. AB - A method of crystallographic analysis to identify myosin heads interacting with specific actin sites in vertebrate striated muscles is described. It is based on a Fourier transform of a helix in which probability of occurrence of subunits varies periodically. It predicts the presence of layer-lines at 1/24 and 1/10.4 nm(-1) which are experimentally observed in contracting and rigor vertebrate striated muscles, showing that the myosin heads are interacting with specific sites on actin but are still maintaining their average 14.5 nm axial periodicity. PMID- 15299579 TI - Phasing proteins at low resolution. AB - A method for obtaining phases of low-order reflections is presented. It is based on four observations: (1) the electron density inside proteins is smooth and uniform at low resolution. (2) Since all proteins have almost the same density, the total volume of the protein is known if the molecular weight is known. (3) The overall shape of many proteins is fairly spherical. (4) The total scattering from a sphere of uniform density is in phase with a point scatterer at its centre of gravity, up to a well defined cross-over. After the first cross-over the total protein molecule scatters out of phase with its centre. If the centre of the protein can be found, the phases of typically the ten lowest resolution reflections can be very accurately determined. The method works, provided low order reflections can be measured accurately and the centre of gravity can be well positioned from these data. The correctly phased low-resolution reflections may be used as a starting set for phase extension. By combining the measured amplitudes with these phases we believe that the size and low-resolution shape of an unknown protein, i.e. the envelope of the molecule, can be obtained. PMID- 15299580 TI - Crystallization of Escherichia coli enoyl reductase and its complex with diazaborine. AB - Recent work has shown that the NADH-dependent enoyl acyl carrier protein reductase from Escherichia coli is the target for diazaborine, an antibacterial agent. This enzyme has been crystallized by the hanging-drop method of vapour diffusion complexed with NAD(+) and in the presence and absence of a thieno diazaborine. The crystals grown in the absence of diazaborine (form A) are in the space group P2(1) with unit-cell dimensions a = 74.0, b = 81.2, c = 79.0 A and beta = 92.9 degrees, and with a tetramer in the asymmetric unit, whilst those grown in the presence of diazaborine (form B) are in the space group P6(1)22 (or P6(5)22) with unit-cell dimensions a = b = 80.9 and c = 328.3 A, and with a dimer in the asymmetric unit. The structure determination of this enzyme in the presence of diazaborine will provide information on the nature of the drug binding site and contribute to a programme of rational drug design. PMID- 15299581 TI - Crystallization of the glutamate dehydrogenase from the hyperthermophilic archaeon Thermococcus litoralis. AB - The NADP(+)-dependent glutamate dehydrogenase from Thermococcus litoralis has been crystallized by the hanging-drop method of vapour diffusion using an ammonium sulfate and PEG mixture as the precipitant. The crystals belong to the monoclinic system and are in space group C2 with unit-cell dimensions a = 142.7, b = 202.0, c = 125.8 A with beta = 113.1 degrees with a hexamer in the asymmetric unit. T. Litoralis, a hyperthermophilic organism, belongs to the family of Archaea and has a maximum growth temperature of about 370 K. The glutamate dehydrogenase isolated from this organism has a half-life of 2 h at 373 K and a comparison of this structure with that of other GluDH's from hyperthermophilic organisms and from mesophiles will contribute to an understanding of the molecular mechanisms which underlie thermostability. PMID- 15299582 TI - Crystallization and preliminary crystallographic analysis of cabbage histidinol dehydrogenase. AB - Recombinant Brassica oleracea histidinol dehydrogenase (HDH) has been crystallized in various space groups using the method of vapour diffusion. The presence or absence of inhibitors and substrates as well as the use of different precipitants has enabled the growth of five different crystal forms. Extensive searches with the first crystal form (A) failed to produce any useful heavy-atom derivatives, mainly because of the instability of the crystals. This provoked the search for further crystal forms in the hope of finding more suitable crystals. At least two of these crystal forms are of interest for further study. PMID- 15299583 TI - Crystallization and initial spectroscopic characterization of the heme-containing dehaloperoxidase from the marine polychaete Amphitrite ornata. AB - The heme-containing dehaloperoxidase from Amphitrite ornata was crystallized from an unbuffered solution containing 30% PEG 8000 and 200 mM ammonium sulfate by the hanging-drop vapor-diffusion method. Dark-red bipyramidal crystals are orthorhombic in space group P2(1)2(1)2(1) with unit-cell dimensions a = 68.5, b = 68.4 and c = 61.1 A. The asymmetric unit contains two subunits related by a non crystallographic twofold axis. The crystals scatter beyond 2 A resolution. The native data have been collected and one single-site mercury derivative has been found. SIRAS phasing was used to determine the positions of the heme Fe atoms and structure determination is in progress. A preliminary spectroscopic investigation indicates that the heme is protoporphyrin IX and its coordination sphere resembles that of a typical heme peroxidase, i.e. histidine ligated. Detailed spectroscopic and electrochemical studies are now under way. PMID- 15299584 TI - Crystallization of a novel esterase which inactivates the macrolide toxin brefeldin A. AB - A novel esterase obtained from Bacillus subtilis and capable of hydrolyzing the phytotoxin brefeldin A was crystallized using the hanging-drop technique. The crystals have two forms and both are monoclinic: form I, space group P2(1) with a = 101.7, b = 64.1, c = 55.4 A and beta = 102.5 degrees, and form II, space group C2 with a = 140.7, b = 82.6, c = 81.5 A and beta = 112.5 degrees. There are two molecules related by a pronounced non-crystallographic dyad per asymmetric unit in both crystal forms. The crystals diffract to 2.3 A using a rotating-anode X ray source. PMID- 15299585 TI - Purification, crystallization and preliminary crystallographic analysis of mare lactoferrin. AB - Lactoferrin is an iron-binding glycoprotein with a molecular weight of 80 kDa. The protein has two iron binding sites. It has two structural lobes, each housing one Fe(3+) and the synergistic CO(3)(2-) ion. The protein was isolated from the colostrum/milk of mares maintained at National Research Centre on Equines, Hisar, India. The purified samples of the protein were crystallized using a microdialysis method. The protein was dialysed against low ionic strength buffer solution. Several crystal forms were obtained, out of which three were characterized which have cell dimensions as follows. Form I a = 79.8, b = 103.5, c = 112.0 A, space group P2(1)2(1)2(1), with one protein molecule per asymmetric unit and a solvent content of 57%. Form II a = 84.9, b = 99.7, c = 103.5 A, space group P2(1)2(1)2(1) with one molecule per asymmetric unit and a solvent content of 55%. Form III a = 151.0, b = 151.0, c = 240.6 A, space group P4(1)2(1)2 with three molecules in the asymmetric unit and a solvent content of 57%. The intensity data up to 3.8 A resolution for form I, 2.9 A resolution data for form II and 6 A resolution data for form III have been collected. Further calculations are in progress. PMID- 15299586 TI - Crystallization and preliminary structural studies of lactose-specific enzyme IIA from Lactococcus lactis. AB - Lactose-specific enzyme IIA of the phosphoenol:pyruvate-dependent sugar phosphotransferase system from Lactococcus lactis has been crystallized in phosphate buffer. The crystals belong to space group P4(1)2(1)2 or its enantiomorph P4(3)2(1)2 with unit-cell axes a = b = 90.9 and c = 82.4 A. The packing parameter (Matthews parameter) V(m) of 2.48 A(3) Da(-1) is consistent with one trimer per asymmetric unit and non-crystallographic threefold symmetry has been confirmed by calculating a self-rotation function. The crystals diffract X-rays to at least 2.3 A resolution, are stable in an X-ray beam and are therefore appropriate for structure determination. Native data to 2.3 A resolution have been collected using a MAR image-plate system at a synchrotron source. One isomorphous heavy-atom derivative has been identified and the presence of an isomorphous signal in the data has been confirmed by Patterson methods. PMID- 15299587 TI - Preliminary crystallographic analysis and further characterization of a dodecaheme cytochrome c from Desulfovibrio desulfuricans ATCC 27774. AB - Dodecaheme cytochrome c has been purified from Desulfovibrio (D.) desulfuricans ATCC 27774 cells grown under both nitrate and sulfate-respiring conditions. Therefore, it is likely to play a role in the electron-transfer system of both respiratory chains. Its molecular mass (37768 kDa) was determined by electrospray mass spectrometry. Its first 39 amino acids were sequenced and a motif was found between amino acids 32 and 37 that seems to exist in all the cytochromes of the c(3) type from sulfate-reducing bacteria sequenced at present. The midpoint redox potentials of this cytochrome were estimated to be -68, -120, -248 and -310 mV. Electron paramagnetic resonance spectroscopy of the oxidized cytochrome shows several low-spin components with a g(max) spreading from 3.254 to 2.983. Two crystalline forms were obtained by vapour diffusion from a solution containing 2% PEG 6000 and 0.25-0.75 M acetate buffer pH = 5.5. Both crystals belong to monoclinic space groups: one is P2(1), with a = 61.00, b = 106.19, c = 82.05 A, beta = 103.61 degrees, and the other is C2 with a = 152.17, b = 98.45, c = 89.24 A, beta = 119.18 degrees. Density measurements of the P2(1) crystals suggest that there are two independent molecules in the asymmetric unit. Self-rotation function calculations indicate, in both crystal forms, the presence of a non crystallographic axis perpendicular to the crystallographic twofold axis. This result and the calculated values for the volume per unit molecular weight of the C2 crystals suggest the presence of two or four molecules in the asymmetric unit. PMID- 15299588 TI - Crystallization and preliminary X-ray analysis of a hydantoinase from Arthrobacter aurescens DSM 3745. AB - L-Hydantoinase from Arthrobacter aurescens DSM 3745 has been purified to homogeneity and crystallized from polyethylene glycol solutions in a form suitable for X-ray diffraction analysis. The crystals have been grown by the sitting-drop variant of the vapour-diffusion method. X-ray diffraction studies show that the crystals belong to the monoclinic space group P2(1) with a = 111.2, b = 74.4, c = 146.5 A and beta = 106.7 degrees. Its asymmetric unit contains four monomers related by 222 symmetry. The crystals diffract to at least 2.6 A. PMID- 15299589 TI - Crystallization and preliminary X-ray crystallographic analysis of the putative [6Fe-6S] prismane protein from Desulfovibrio vulgaris (Hildenborough). AB - Crystals of the prismane protein from Desulfovibrio vulgaris (Hildenborough) containing a putative [6Fe-6S] cluster have been obtained and X-ray data collected to a resolution of 1.7 A using synchrotron radiation. The unit cell is orthorhombic with a = 64.1, b = 65.1 and c = 154.1 A, space group P2(1)2(1)2(1) (No. 19). The unit cell will readily accommodate four molecules of molecular weight 60 kDa with a corresponding solvent content of approximately 48%. PMID- 15299590 TI - Crystallization and preliminary X-ray analysis of a histidine kinase domain of the anaerobic sensor protein ArcB from Escherichia coli. AB - Crystals of a novel histidine protein kinase domain of the anaerobic sensor protein ArcB from Escherichia coli have been obtained by a hanging-drop vapor diffusion method with micro- and macroseeding techniques. Preliminary X-ray crystallographic analysis revealed that they belong to space group P2(1)2(1)2(1) with dimensions a = 30.56, b = 34.93 and c = 110.78 A, having one molecule in the crystallographic asymmetric unit. The crystals diffract to at least 2.0 A resolution. PMID- 15299591 TI - Crystallization and preliminary crystallographic analysis of the DNA gyrase B protein from B. stearothermophilus. AB - DNA gyrase B (GyrB) from B. stearothermophilus has been crystallized in the presence of the non-hydrolyzable ATP analogue, 5'-adenylyl-beta-gamma imidodiphosphate (ADPNP), by the dialysis method. A complete native data set to 3.7 A has been collected from crystals which belonged to the cubic space group I23 with unit-cell dimension a = 250.6 A. Self-rotation function analysis indicates the position of a molecular twofold axis. Low-resolution data sets of a thimerosal and a selenomethionine derivative have also been analysed. The heavy atom positions are consistent with one dimer in the asymmetric unit. PMID- 15299592 TI - Crystallization of DsbC, the disulfide bond isomerase of Escherichia coli. AB - DsbC is a 2 x 23 kDa soluble dimeric protein molecule involved in protein disulfide bond formation in the E. coli periplasm, primarily catalyzing disulfide bond rearrangements. Crystals of both the native and selenomethione protein suitable for structure determination were obtained using the hanging-drop vapour diffusion method. The best crystals were obtained using 18-22% (v/v) polyethylene glycol 550 monomethyl ether in 100 mM Tris-HCl (pH 8.9). Seeding methods were used to produce large crystals diffracting to 2 A resolution, and the detergent n octyl-beta-glucoside was used to improve crystal quality. Significant variation in cell dimensions and crystal order was observed. Cell dimensions obtained for frozen crystals were in the range a = 58.8 (0.3), b = 78.9 (0.5), c = 95.2 (5.0) A. The lattice is orthorhombic and systematic absences indicate that the space group is P2(1)2(1)2(1). PMID- 15299593 TI - Crystallization and molecular replacement solution of human heparin binding protein. AB - The highly glycosylated protein, human heparin binding protein, has been crystallized in the primitive orthorhombic space group P2(1)2(1)2(1) with cell dimensions a = 39.0, b = 66.2 and c = 101.4 A. Ethanol was used as precipitant and glycerol as additive. A full data set has been collected to 3.1 A and diffraction was observed to at least 2.3 A. A molecular replacement solution using human neutrophile elastase as a search model was obtained, showing one molecule per asymmetric unit. The crystal packing showed no bad contacts and the R factor was 44.8% after ten cycles of rigid-body refinement. PMID- 15299594 TI - Crystallization and preliminary crystallographic analysis of two beta-mannanase isoforms from Thermomonospora fusca KW3. AB - Three beta-mannanase isoforms were isolated from the supernatant of a thermophilic actinomycete culture from Thermomonospora fusca KW3. Two of the isoforms (Q1, Q 1.1) were crystallized by the hanging-drop method at room temperature using ammonium sulfate as a precipitant. The isoforms form rod-shaped colorless crystals. Both belong to the orthorhombic space group P2(1)2(1)2(1). The cell dimensions are a = 46.7, b = 61.1, and c = 128.2 A for isoform Q1, and a = 43.8, b = 46.2, and c = 132.8 A for isoform Q1.1. The asymmetric unit of either isoform contains one mannanase molecule. Native data have been collected to 2.2 A resolution for Q1 and to 1.65 A resolution for Q1.1 using synchrotron radiation. PMID- 15299596 TI - Application of known X-ray phases in the crystallographic study of a small protein. AB - Phase information, assumed known from three-beam diffraction experiments, has been used successfully as input to direct methods to re-determine the crystal structure of rubredoxin. With data at 1.54 A resolution, starting sets containing 26 single phases, or alternatively 45 triplet phases, in both cases known with a mean error of +/-22.5 degrees, were sufficient to initiate solution of the structure. Conventional figures of merit were employed in the early stages to reject the majority of the incorrect phase models. The presence of a FeS(4) cluster in the structure was used in the interpretation of the initial maps. Phase sets including 2500 E's with a mean single phase error approximately 70 degrees or a mean triplet phase error approximately 80 degrees, both relative to the model from the crystallographic refinement, could be refined and expanded by Fourier recycling using the SAYTAN formalism. Several parameters have been varied to study their influence on phase expansion and refinement. PMID- 15299597 TI - Salt-induced aggregation of lysozyme studied by cross-linking with glutaraldehyde: implications for crystal growth. AB - Glutaraldehyde cross-linking followed by sodium dodecyl sulfate polyacrylamide gel electrophoresis has been used to detect aggregates of isozyme in solutions which lead to crystals. In solutions of varying NaCl content, the number of aggregates was found to be related to the ionic strength of the solution. Solutions of 1% NaCl, pH 4.0 were monomeric while those containing 7-15% NaCl, pH 4.0 were shown to be as much as 36% aggregated and 64% monomeric. The aggregates detected at the highest salt and protein concentration studied were composed of dimers, trimers and tetramers. The aggregates increased by addition of single units suggesting the aggregation pathway to be that of monomer addition. The kinetics of the cross-linking reaction were slow preventing a study of either the time dependence of aggregation or the effect of temperature on aggregate distributions. Comparison of the total aggregate concentrations for NaCl and Na(2)SO(4) showed that the concentration of aggregates was related to the ionic strength of the solution suggesting that in both crystallization and precipitation, electrostatic shielding of like-charged protein molecules is necessary in order for aggregation to occur. PMID- 15299598 TI - Extraction of molecular edges on the average difference map by a Monte-Carlo method. AB - A method giving a low-resolution image of the molecules in the unit cell has been described, which was based only on the observed structure factors. An operator, called the average difference operator (ADO), was introduced in reciprocal space to flatten the electron densities everywhere but the regions on either side of the molecular envelope in real space. The observed structure factors were first modified by ADO, then a Monte-Carlo condensing protocol [Subbiah (1991). Science, 252, 128-133; (1993). Acta Cryst. D49, 108-119] was employed to stimulate the modified electron-density map at low resolution. It was found that molecular edges could be extracted, especially when there was relatively large solvent content in the unit cell. PMID- 15299599 TI - X-ray structures of Mn, Cd and Tb metal complexes of troponin C. AB - The crystal structures of three metal complexes of troponin C (TnC) have been determined and refined where the two occupied structural Ca(2+) sites in the C domain have been substituted by Mn(2+), Cd(2+) and Tb(3+). The X-ray intensity data were collected to 2.1, 1.8 and 1.8 A resolution, respectively, on the three metal complexes, which are isomorphous with Ca-TnC. The three complexes have r.m.s. deviations of 0.27, 0.25 and 0.35 A, respectively, for all protein atoms, from Ca-TnC. Irrespective of the charge on the metal (+2 or +3), the occupied sites 3 and 4 exhibit a distorted pentagonal bipyramidal coordination, like Ca TnC, with seven ligands, six from the 12-residue binding loop and the seventh from a water molecule. Mn(2+) at site 4 seems to display a longer distance to one of the carboxyl bidentate ligands representing an intermediate coordination simulating the six-coordinate Mg(2+). The carboxyl O atoms of the bidentate Glu12 are displaced on the side of the equatorial plane passing through the remaining three ligands with one O atom closer to the plane (Delta of 0.11 to 0.76 A) than the other (Delta of 0.93 to 1.38 A). The two axial ligands are an aspartic carboxyl O atom and a water molecule. The metal is displaced (0.18 to 0.56 A) towards the water facing the water channel. PMID- 15299600 TI - Structure determination of the human protective protein: twofold averaging reveals the three-dimensional structure of a domain which was entirely absent in the initial model. AB - Mutations in the human 'protective protein' result in the human lysosomal storage disease galactosialidosis. The structure of the human 'protective protein' has been determined using X-ray crystallography to a resolution of 2.2 A. Initial phases were obtained from molecular replacement calculations. A very partial search model comprising 30% of the scattering mass, was constructed from the atomic model of the wheat serine carboxypeptidase. This truncated probe was used to find the position of two monomers in the asymmetric unit. Subsequently, 'bootstrapping' cycles, consisting of twofold averaging and model expansion, retrieved the electron density for residues initially missing. In particular, it proved possible to add a domain (more than 110 residues) to the initial partial search model. In total, 314 residues per asymmetric unit were added to the 588 residues of the initial model. Factors contributing to our success are discussed. PMID- 15299601 TI - Direct-method structure determination of the native azurin II protein using one wavelength anomalous scattering data. AB - The one-wavelength anomalous scattering (OAS) X-ray diffraction data of azurin II, a copper-containing protein from Alcaligenes xylosoxidans were collected at the Photon Factory, Japan at a 'routine' wavelength of 0.97 A. The structure had been originally solved by the molecular-replacement method [Dodd, Hasnain, Abraham, Eady & Smith (1995). Acta Cryst. D51, 1052-1064]. As a technique of ab initio structure determination, the direct method [Fan, Hao, Gu, Qian, Zheng & Ke (1990). Acta Cryst. A46, 935-939] was attempted to break the phase ambiguity intrinsic to OAS data. The phases were then improved using the solvent-flattening method. The final electron-density map clearly shows most Calpha positions and many side chains and it is traceable without prior knowledge of the structure. It is concluded that the direct method is capable of phasing anomalous scattering data collected at one wavelength from moderate-sized native proteins (M(w) approximately 20 kDa) which contain copper or atoms with a similar scattering power. PMID- 15299602 TI - Conformational change of the adenovirus DNA-binding protein induced by soaking crystals with K3UO2F5 solutions. AB - Soaking crystals of the C-terminal DNA-binding domain of the adenovirus single stranded DNA-binding protein with a buffer containing K(3)UO(2)F(5) results in a 9% change of the crystallographic c axis without destruction of the crystals or appreciable loss of resolution. The crystals belong to space group P2(1)2(1)2(1) with a = 79.7, b = 75.6 and c = 60.6 A. The three-dimensional structure has been refined to 2.7 A with a crystallographic R factor of 0.206. Antiparallel chains of protein molecules running through the entire crystal are linked by uranyl ions. The relative orientation of protein monomers is flexible, even in the crystalline state, and allows changes in the packing of the protein chains. PMID- 15299603 TI - Crystals of ligand-free bovine neurophysin II. AB - A modified neurophysin, des 1-6 bovine neurophysin II, has been crystallized in the absence of bound hormone or hormone analogue. These crystals represent the first crystals of ligand-free neurophysin, and are essential for understanding neurophysin-hormone recognition as well as hormone-induced neurophysin dimerization. The crystals diffract to beyond 1.8 A resolution, belong to space group P3(1)21 (or P3(2)21) with a = 48.86, c = 78.61 A, and contain one molecule per asymmetric unit. PMID- 15299604 TI - Mutant Met121Ala of Pseudomonas aeruginosa azurin and its azide derivative: crystal structures and spectral properties. AB - The crystal structures of the azurin mutant Met121Ala and its azide derivative Met121Ala-azide from Pseudomonas aeruginosa have been determined. The final crystallographic R values are 21.3 and 19.4% for the two structures, respectively. In the Met121Ala mutant, the distance between the copper ion and His117 increases by 0.34 A compared with the wild-type structure. The removal of the methionine in the apical position induces a shortening of the distance from the copper ion to the carbonyl O atom of Gly45 from 2.97 to 2.74 A. In the Met121Ala-azide structure, the azide anion occupies the cavity created by replacing the Met121 side chain with the smaller methyl group of Ala. The azide anion binds with a terminal N atom to the copper ion at a distance of about 2.04 A. In addition, the copper ion has moved out of the trigonal plane by about 0.26 A towards the azide anion. Thus, the copper site in this structure has a distorted tetrahedral arrangement. The spectroscopic characteristics show, in addition, that the copper sites in the two structures are distinctively different. The Met121Ala mutant still maintains the properties of an ordinary type 1 copper site while the Met121Ala-azide derivative has an absorption maximum at about 409 nm and the copper hyperfine coupling has increased to a value intermediate between those of type 2 copper and the wild-type azurin. PMID- 15299605 TI - Flash-freezing causes a stress-induced modulation in a crystal structure of soybean lipoxygenase l3. AB - A dynamic conformational flexibility of a protein might be a source of non covalent structural heterogeneity, causing diminished diffracting ability of crystals and disorder in a crystal structure of soybean lipoxygenase L3. Room temperature data, space group C2, correspond to a structure with large channels lined mostly or in part by disordered fragments of the molecule or flexible loops with an increased thermal vibration. A rapid change in temperature of approximately 200 K creates a wave of a stress-induced modulation that propagates in the crystal changing its reciprocal space into a three-dimensional quilt-like mixture of C and P intertwined lattices. Low-temperature data indicate a transformation from the dynamic to static disorder, leading to a primitive unit cell with 10% reduced volume. The molecules, formerly related by a twofold axis are rotated by approximately 7 degrees and are shifted along the diagonal to be approximately 4 A, closer together. During a routine data collection for the flash-frozen crystals of similar properties such phenomena could easily go unnoticed leading to biased results because of such effects and possibly improper indexing of the data. PMID- 15299606 TI - Structure of the Val122Ile variant transthyretin - a cardiomyopathic mutant. AB - The Val122Ile mutant transthyretin (TTR Ile122) is an amyloidogenic protein which has been described as the major protein component of amyloid fibrils isolated from patients with familial amyloidotic cardiomyopathy (FAC), a disease characterized by cardiac failure and amyloid deposits in the heart. The reasons for the deposition of TTR are still unknown and it is conceivable that a conformational alteration, resulting from the mutation, is fundamental for amyloid formation. The three-dimensional structure of TTR Ile122 was determined and refined to a crystallographic R factor of 15.8% at 1.9 A resolution. The r.m.s. deviation from ideality in bond distances is 0.019 A and in angle-bonded distances is 0.027 A. The presence of two crystallographically independent monomers in the asymmetric unit allowed additional means of estimation of atomic coordinate error. The structure of the mutant is essentially identical to that of the wild-type transthyretin (TTR). The largest deviations occur in surface loops and in the region of the substitution. The protein is a tetramer composed of identical subunits; each monomer has two four-stranded beta-sheets which are extended to eight-stranded beta-sheets when two monomers associate through hydrogen bonds forming a dimer, which is the crystallographic asymmetric unit. The replacement of valine for isoleucine introduces very small alterations in relation to the wild-type protein; nevertheless they seem to confirm a tendency for a less stable tetrameric structure. This would support the idea that the tetrameric structure might be disrupted in amyloid fibrils. PMID- 15299607 TI - Molecular replacement with NMR models using distance-derived pseudo B factors. AB - The statistical significance of molecular-replacement solutions with models derived from NMR data is strongly enhanced if pseudo B factors that reflect expected atomic coordinate errors are introduced. These B factors are derived from atomic distances of an ensemble of NMR models and an averaged model. A recently determined X-ray structure of a Pleckstrin homology domain:ligand complex has been used as a test case for molecular replacement with NMR templates. The feasibility of the models for molecular replacement has been studied in two steps: (i) correctness of solutions, verified by correct rotational angles and translations; (ii) statistical significance (detection) of solutions, measured by R factors and correlation coefficients in the translation function. None of the models with uniform B factors were detectable in the translation function whereas two models with distance-derived B factors gave statistically significant R factors and correlation coefficients. The potential impact of distance-derived B factors on the detectability of molecular replacement solutions was further tested by using B factors that were derived from spatial distances between each NMR model and the X-ray structure. It is concluded that the introduction of distance-derived B factors can be an essential component of many future molecular-replacement problems that use NMR models as templates. PMID- 15299608 TI - Growth mechanism and morphology of tetragonal lysozyme crystals. AB - The tetragonal form of hen egg-white lysozyme is the most investigated protein crystal for growth studies, but the relationship between its surface morphology and internal structure is still not well understood. One method of determining this relationship for inorganic crystals is by employing the periodic bond chain (PBC) theory of Hartman & Perdok [Hartman & Perdok (1955). Acta Cryst. 8, 49-52, 521-524, 525-529]. However, complexities resulting from the packing arrangements and the number of intermolecular bonds in protein crystals have resulted in the use of only simplified versions of this theory so far. In this study a more complete PBC analysis of tetragonal lysozyme crystals was carried out, coupled with an approach incorporating the molecular orientations of the crystal structure. The analysis revealed the existence of a helical tetramer building block of the entire crystal structure, centered around the 4(3) crystallographic axes, resulting in double-layered slices and PBC's throughout. The analysis also indicated that the crystallizing units for the faces are at least as large as this tetramer, with the experimental evidence suggesting that it is a tetramer unit for the {101} faces and an octamer unit for the {110} faces. The {110} faces were shown to be molecularly smooth F faces, while the {101} to be essentially rough S faces. The predicted morphology and growth mechanisms were found to explain numerous experimental observations from electron and atomic force microscopy, etching studies, lysozyme aggregation studies and measurements of growth kinetics. PMID- 15299609 TI - Structure of d(CCCTAGGG): comparison with nine isomorphous octamer sequences reveals four distinct patterns of sequence-dependent intermolecular interactions. AB - The self-complementary deoxyoctanuceotide d(CCCTAGGG) crystallizes as an A-type double helix in the space group P4(3)2(1)2, a = b = 42.22 and c = 24.90 A, with one strand per asymmetric unit. Using 1533 unique reflections at 1.9 A and I > 2sigma, the structure was solved by molecular placement and refined to a final R value of 16.4%. This structure is isomorphous with nine other tetragonal A-DNA octamers, possessing a central pyrimidine/purine step that is fully extended along the backbone with trans, trans conformations around the C4'-C5' and O5'-P bonds. A structural water, sandwiched between the pi-cloud of the terminal guanine and the N3 atom of G7 in the adjacent duplex, stabilizes an intermolecular base triplet with one hydrogen bond between the terminal cytosine and G6 of an adjacent duplex. Comparative analysis of this structure with the isomorphous A-DNA octamers reveals the importance of base sequence and minor groove hydration in intermolecular interactions. The minor grooves, which provide both hydrophobic and polar interactions, allow for four patterns of sequence dependent binding involving interduplex base triplets in which the third base is bonded through a single hydrogen bond. A conserved water molecule appears to be crucial in the stabilization of these intermolecular interactions, which resemble specific recognition motifs found in the crystal structures of the TATA-box/TBP protein complex. PMID- 15299610 TI - Bayesian difference refinement. AB - Interest in a pair of highly isomorphous structures often focuses on the differences between them. In cases where substantial correlated model errors exist or where there are differences in the quality of the two experimental data sets (cases quite common in macromolecular crystallography), independent refinement of the two structures does not lead to the most accurate estimate of the differences between them. An alternative procedure that has proven effective in some such cases is difference refinement, in which the residual between observed and calculated differences in structure-factor amplitudes between the two structures is minimized. A Bayesian approach has been used to extend the range of applicability of difference refinement to cases where there is only partial correlation in model errors and where the overlap between the data sets is limited. The resulting method, Bayesian difference refinement, uses residuals to be minimized that vary smoothly between difference refinement and independent refinement. When the errors in the two structural models are very similar, difference refinement is used; when they are very different, independent refinement is used; and when they are partially correlated, a combination of the two is used. The procedure is very simple to apply and does not significantly increase the computational demands of refinement. PMID- 15299611 TI - Application of moderate hydrostatic pressure induces unit-cell changes in rhombohedral insulin. AB - 2Co(2+)-insulin crystals were subjected to hydrostatic pressures of up to 30 bar in a nitrogen gas cell. Changes in the diffraction pattern occurred at pressures as low as 5 bar. Analysis with standard image-processing software showed unit cell dimension changes resulting in reductions in volume of up to 2.6%. PMID- 15299612 TI - Crystallization and preliminary crystallographic studies of alpha-amylase inhibitor from wheat (Triticum aestivum). AB - Crystals of the human salivary alpha-amylase inhibitor from wheat have been obtained. A native data set was collected to 2.1 A resolution with 90% completeness at laboratory sources. The crystals belong to the trigonal system, space group P3(1) (or enantiomer) with a = b = 79.31, c = 60.56 A. Crystal density analysis and self-rotation function studies suggest the presence of four subunits in the asymmetric unit. PMID- 15299613 TI - Preliminary X-ray diffraction studies of an RNA pseudoknot that inhibits HIV-1 reverse transcriptase. AB - Crystals of a 26-nucleotide pseudoknot RNA, PK26, have been grown. The RNA was produced using phosphoramidite chemistry and was purified by denaturing polyacrylamide electrophoresis. The crystallization was robust with respect to changes in the number of nucleotides and to the salt used as precipitant. The crystals belong to space group P4(1)22 or P4(3)22 with unit-cell dimensions a = b = 61.6, c = 98.9 A. The best crystals diffract X-rays to 2.9 A. Three different sequences incorporating a single 5-bromo-deoxyuridine or 5-bromo-uridine nucleotide were also crystallized. Two of these derivatives are being used to determine the structure by multiple isomorphous replacement. PMID- 15299614 TI - Purification, crystallization and preliminary X-ray analysis of a mannose-binding lectin from bluebell (Scilla campanulata) bulbs. AB - Crystals have been grown of a mannose-specific lectin from bluebell (Scilla campanulata) bulbs in a form suitable for X-ray diffraction studies. The crystals, which diffract to high resolution, grew in hanging drops by vapour diffusion, equilibrating with a solution of 70% saturated ammonium sulfate at pH 4.7-4.8 at 293 K, in the absence of any mannose saccharides. Crystals are orthorhombic, P2(1)2(1)2, with unit-cell dimensions a = 70.78, b = 93.69, c = 46.92 A. The functional lectin molecule is organized as a tetramer of four identical 14 kDa subunits, with only two subunits in the asymmetric unit. Data to 1.86 A resolution have been recorded and the structure determined by the molecular replacement method. PMID- 15299615 TI - Crystallization and preliminary diffraction analysis of porcine heart mitochondrial NADP(+)-dependent isocitrate dehydrogenase. AB - Isocitrate dehydrogenases [isocitrate:NAD(P)(+) oxidoreductase (decarboxylating), E.C. 1.1.1.42] are ubiquitous metabolic enzymes which occur in all living organisms. The NADP(+)- dependent mitochondrial isocitrate dehydrogenase from pig heart has been crystallized from polyethylene glycol/sodium sulfate mixtures in the presence of Mg(2+) and isocitrate. The crystals belong to space group C2 with a = 137.0, b = 113.4, c = 65.0 A and beta = 98.5 degrees, and diffract to at least 2.4 A resolution. There are two protein monomers per asymmetric unit which are related by non-crystallographic twofold symmetry. PMID- 15299616 TI - Crystallization and preliminary X-ray diffraction analysis of two lysinal derivatives of Achromobacter protease I. AB - Two crystal forms of lysinal derivatives of Achromobacter protease I have been obtained. The first, modified by benzyloxycarbonyl-Val-lysinal crystallizes in the monoclinic space group P2(1) with unit-cell dimensions of a = 39.6, b = 71.2, c = 45.6 A and beta = 98.4 degrees. The second, modified by benzyloxycarbonyl-Leu Leu-lysinal crystallizes in the orthorhombic space group I222 (or I2(1)2(1)2(1)) with unit-cell dimensions of a = 98.7, b = 102.2 and c = 55.8 A. The space groups and the unit-cell dimensions of the present two lysinal derivatives are different to those of the protease and TLCK- modified one. The space group of the protease is P1 with cell dimensions a = 39.53, b = 40.34, c = 43.92 A, alpha = 114.81, beta = 113.75 and gamma = 74.00 degrees and that of the TLCK-modified one is also P1 with cell dimensions of a = 37.30, b = 42.74, c = 48.02 A, alpha = 120.10, beta = 112.81 and gamma = 68.54 degrees. Diffraction to 1.9 A resolution for the Val-lysinal modified crystal and to 2.2 A resolution for the Leu-Leu-lysinal modified crystal has been observed using a rotating-anode X-ray generator. Full structure determinations of these lysinal-modified protease crystals may lead to an understanding of the molecular basis of enzyme-substrate interactions in the catalytic process of this protease. PMID- 15299617 TI - Crystallization and preliminary X-ray analysis of 3-isopropylmalate dehydrogenase from the moderate facultative thermophile Bacillus coagulans. AB - Three crystalline forms of 3-isopropylmalate dehydrogenase from the moderate facultative thermophile Bacillus coagulans were obtained by hanging-drop vapor diffusion methods. One of them, which had crystallized under slightly milder conditions than the others, was suitable for X-ray analysis. Its asymmetric unit contains one dimeric molecule and the solvent content is higher than in other protein crystals. The crystal structure was solved in a preliminary manner by the molecular-replacement technique. PMID- 15299618 TI - Crystallization and preliminary crystallographic analysis of the Ras binding domain of RalGDS, a guanine nucleotide dissociation stimulator of the Ral protein. AB - The RalGDS is a guanine nucleotide dissociation stimulator which activates the Ral protein, a Ras-like small GTPase. The C-terminal domain of the RalGDS (C RalGDS) binds tightly to the effector loop of Ras suggesting that the RalGDS may be a crossing point of two signal tranduction pathways associated with the Ras and Ral proteins. C-RalGDS has been purified and crystallized in space group C2, with unit-cell dimensions a = 108.8, b = 30.7, c = 51.3 A, beta = 91.7 degrees at 277 K and a = 103.8, b = 30.55, c = 51.4 A, beta = 94.9 degrees for data collected at 100 K. The crystals diffract to 1.8 A at a synchrotron radiation source. To use the multiple-wavelength anomalous diffraction method for phasing, a selenomethionine derivative of the protein has also been crystallized. PMID- 15299619 TI - Crystallization and preliminary X-ray analysis of a bifunctional enzyme: HHDD isomerase/OPET decarboxylase from Escherichia coli. AB - A bifunctional enzyme, 2-hydroxyhepta-2,4,-diene-l,7-dioate isomerase/5-oxopent-3 ene-1,2,5-tricarboxylate decarboxylase from the homoprotocatechaute (HPC) degradative pathway of Escherichia coli has been crystallized, using polyethylene glycol as a precipitant. The enzyme, of molecular weight 44 514 Da forms crystals belonging to the orthorhombic space group P2(1)2(1)2(1) with cell dimensions a = 106, b = 127 and c = 139 A. The crystals diffract to at least 2.2 A resolution using synchrotron radiation. A complete native data set has been collected to 3.3 A resolution. The Matthews number calculated for a single molecule in the asymmetric unit is outside the normally acceptable limits and the aggregation state of the molecules in the crystal was investigated using self-rotation function studies, the results show features which are consistent with a tetramer in the asymmetric unit, giving a V(m) value of 2.7 A(3) Da(-1). PMID- 15299620 TI - Crystallization and preliminary X-ray analysis of human dUTPase. AB - Human dUTPase, expressed in Escherichia coli, has been crystallized. Single crystals were obtained by the vapour-diffusion technique using 2-propanol and PEG 4000 as precipitants. The enzyme was co-crystallized with the substrate dUTP and a metal chelator EDTA to prevent hydrolysis of the substrate. The crystals belong to the orthorombic space group P2(1)2(1)2(1) with cell dimensions a = 67.51, b = 68.26 and c = 91.00 A. The crystallographic asymmetric unit contains one trimer of identical subunits. PMID- 15299621 TI - Crystallization and preliminary X-ray diffraction studies of the influenza C virus glycoprotein. AB - Influenza C virus contains a single surface glycoprotein in its lipid envelope which is the hemagglutinin-esterase-fusion glycoprotein (HEF). HEF binds cell surface receptors, is a receptor-destroying enzyme (a 9-O-acetylesterase), and mediates the fusion of virus and host cell membranes. A bromelain-released soluble form of HEF has been crystallized. Two different tetragonal forms have been identified from crystals with the same morphology [P(1(3))22, a = b = 154.5, c = 414.4 A, and P4(1(3))2(1)2, a = b = 217.4, c = 421.4 A]. Both crystal forms share a common packing scheme. Synchrotron data collection and flash cooling of crystals have been used for high-resolution data collection. PMID- 15299622 TI - Crystallization and preliminary X-ray studies on the lectin from the seeds of Cratylia mollis. AB - The lectin from the seeds of Cratvlia mollis shows strong binding to human malignant cancerous tissues, particularly those from many glands, uterus, rectum and brain. The C. mollis lectin has been crystallized using the hanging-drop method with polyethylene glycol 6000 as a precipitant. Two different crystal forms were grown from the same drops and they belong to space groups I222 and P2(1)2(1)2(1), respectively. The cell parameters obtained were a = 63.26 (4), b = 77.45 (8) and c = 105.22 (8) A, for the I222 form, and a = 88.83 (5), b = 183.24 (9) and c = 61.70 (2) A for the P2(1)2(1)2(1) crystals. The solution of both structures is currently being attempted by means of molecular replacement techniques. PMID- 15299624 TI - An eye lens protein-water structure: 1.2 A resolution structure of gammaB crystallin at 150 K. AB - gammabeta-crystallin is a structural protein of the eye lens with a role in the maintenance of an even distribution of protein and water over distances around the wavelength of light, preserving lens transparency. The structure of the 174 residue bovine protein has already been determined at room temperature to 1.47 A resolution. By flash freezing the protein crystals, data have now been collected to a nominal resolution limit of 1.2 A as radiation damage was essentially eliminated. The protein-water model has been refined against this data using the program RESTRAIN converging to an R factor of 18.5% with all data. Atomic positions are clearly indicated in the electron-density maps. Discrete bimodal disorder has been visualized for a few side chains. Out of a total of 498 water molecules present in the crystal asymmetric unit, 394 have been modelled and refined at unit occupancy. The solvent structure is extremely well ordered with an average B value of 23.4 A(2). Partially occupied sites have been identified where disorder in the protein induces concomitant disorder in the local solvent structure. The solvent structure covers 97% of the solvent-exposed surface of the protein in the crystal. 126 water molecules are distributed in second and higher hydration shells. There are networks of hydrogen-bonded solvent extending up to 64 molecules in a network, comprising trimers and tetramers as well as five- and six-membered water-ring structures. The hydration of the protein surface is dominated by arginine and aspartate side chains. Extensive cages of highly ordered solvent molecules are also observed around exposed non-polar groups. PMID- 15299625 TI - Cryocrystallography of 3-Isopropylmalate dehydrogenase from Thermus thermophilus and its chimeric enzyme. AB - The crystal structures of thermostable enzyme, 3-isopropylmalate dehydrogenase of Thermus thermophilus (10T) and a chimeric enzyme between T. thermophilus and Bacillus subtilus with one point mutation (cS82R), were determined at both 100 and 150 K. At the cryogenic condition, the volume of the unit cell decreased by 5% as a result of a contraction in the solvent region. Although the overall structures of both enzymes at low temperature were the same as that of 10T at room temperature, interactions between two domains and between two subunits in a functional dimer of cS82R were significantly altered. The decrease in the average temperature factor of 10T at low temperature and no significant decrease for cS82R suggested that the structure of the engineered enzyme (cS82R) may have many conformational substates even at low temperature, while the native enzyme (10T) at low temperature has a more definite conformation than that at room temperature. The location of water molecules around the enzyme molecule and the calculation of the radii of gyration suggested that cS82R had a weaker hydration than 10T. PMID- 15299626 TI - Structure of diferric duck ovotransferrin at 2.35 A resolution. AB - The structure of diferric duck ovotransferrin (DOT) has been determined and refined at a resolution of 2.35 A. The DOT structure, which contains two iron binding sites, is similar to the known transferrin and lactoferrin structures. The two iron-binding sites, one in the N-terminal lobe and one in the C-terminal lobe of the molecule, are similar but not identical. The main differences between the three known structures lie in the relative orientations of the N- and C-lobes with respect to each other. In the DOT structure the large aromatic side chain of Phe322 in the N-lobe packs against the conserved residue Gly387 in the C-lobe. This interaction is at the centre of the interface between the two lobes and could play a crucial role in determining their relative orientation. Other differences between the structures occur in the surface loops and in the peptide connecting the two lobes. The final crystallographic model consists of 5309 protein atoms (686 residues), two Fe(3+) ions, two (bi)carbonate ions and three carbohydrate moities. 318 water molecules have been added to the model. The final R factor is 0.22 for 25 400 observed reflections between 10 and 2.35 A resolution. PMID- 15299627 TI - A self-validation technique for protein structure refinement: the extended Hamilton test. AB - An extension is proposed for the self-validation Hamilton test [Hamilton (1965). Acta Cryst. 18, 502-510] for crystallographic refinement. The method is based on the statistical F test and evaluates the significance of the R-factor ratio between two refinement protocols. The general case of two refinements carried out with different numbers and types of non-linear restraints is examined. The restraints are considered as extra observations weighted by a coefficient expressing their effective number. There exists a restriction on the weighting coefficients between the two refinements. An empirical method to evaluate the effective number of restraints is provided. The method may allow the detection of unreasonably tight restraints. The expectation value for r.m.s. R(free), given the r.m.s. R, can be estimated. Thus, the significance of the observed drop in R(free) can be assessed. Compared to cross-validation using R(free) [Brunger (1992). Nature (London), 355, 472-474] self-validation has the advantage that it does not require omission of any experimental data. The significance of the improvement obtained by moving from isotropic to anisotropic description of thermal parameters in the refinement of a protein at 1.5 A, resolution is used as an example. PMID- 15299628 TI - A local approximation to supersaturation affords a useful coordinate transformation for the study of crystal growth. AB - Two of the most important experimental variables in the search for appropriate crystallization conditions are the initial concentrations of macromolecule and crystallizing agent. Previously, it has been suggested that the coordinate transformation { [crystallizing agent], [macromolecule] } --> { [macromolecule] x [crystallizing agent], [macromolecule]} be used to sample crystal growth conditions. Here, it is shown that this transformation is a special case of a generally applicable transformation. The initial supersaturation can be represented locally by a rectangular hyperbola involving multiples of the product of macromolecule and crystallizing agent concentrations. The coordinate system for the solubility diagram, ([crystallizing agent] versus [macromolecule]), can thus be transformed analytically to an alternative coordinate system in which the independent variables are local approximations to the initial supersaturation and the reservoir of soluble macromolecule available to feed a growing seed. In the new coordinate system the 'nucleation zone' is 'orthogonalized', so it can be sampled efficiently on a rectangular grid, with greater assurance that experiments will give rise to crystals. Moreover, since these new coordinate directions segregate fundamental effects on nucleation from effects on growth, using them in experimental designs should improve data analysis for response surface experiments. PMID- 15299629 TI - Structure of the purine-pyrimidine alternating RNA double helix, r(GUAUAUA)d(C), with a 3'-terminal deoxy residue. AB - The crystal structure of the purine-pyrimidine alternating octameric RNA helix, r(GUAUAUA)d(C), carrying a 3'-terminal deoxycytidine residue, has been determined at 2.2 A resolution. The molecule crystallizes in the rhombohedral space group R3 (hexagonal cell constants: a = b = 43.07,c = 59.36 A;alpha = beta = 90,gamma = 120 degrees )with one duplex in an asymmetric unit. The structure was solved by molecular replacement and refined with 83 and 2/3 solvent molecules and 2/3 sodium ions to a final R factor of 15.6% using 1775 reflections (86%). The duplexes are approximately linear, their global helix axes are inclined by 10 degrees with respect to the 3(2)-screw axes, and they are stacked on top of each other in a head-to-tail fashion. The twist between the junction base pairs of the stacked duplexes is negligible resulting in a discontinuity of the helix backbones and grooves. The sodium ions on the threefold axis play a significant role in the organization of the packing network. The helical parameters, particularly the twist and the roll, of this alternating sequence are in accord with Calladine's rules. Almost all the 2'-hydroxyl groups are involved in specific hydrogen-bonding interactions, either directly to the sugar ring oxygens O4' on the 3' side, or, through water bridges, to the sugars, phosphates, or bases. This hydrogen bonding of the 2'-hydroxyl groups restrains the conformation of the sugar-phosphate backbone and the glycosidic torsion angles of this RNA fragment. The lack of intermolecular packing contacts in the grooves provides a clear picture of the groove solvation. PMID- 15299630 TI - RNA - synthesis, purification and crystallization. AB - Protocols for the routine chemical synthesis and purification of milligram quantities of RNA and DNA-RNA chimeras meeting the demands of X-ray crystallography are described. An efficient screening protocol to test the crystallizability of the molecules and the optimization of the crystallization conditions are presented, so as to allow reproduction by others. Essentially the same crystallization conditions as for DNA oligomers can be employed for RNA crystallization. Specific examples involving alternating octamers, G/C-rich decamers, sequences with overhangs, and drug complexes of chimeras are discussed. Success of the methods is attested by the crystals obtained which diffract to high resolution. PMID- 15299631 TI - X-ray structure of the cupredoxin amicyanin, from Paracoccus denitrificans, refined at 1.31 A resolution. AB - High-resolution X-ray diffraction data to d(min) = 1.31 A were collected on a Xuong-Hamlin area detector from crystals of the blue-copper protein amicyanin, isolated from P. denitrificans. With coordinates from the earlier 2.0 A structure determination as a starting point, simulated annealing and restrained positional and temperature factor refinements using the program X-PLOR resulted in a final R factor of 15.5%, based on 21 131 unique reflections in the range 8.0-1.3 A. Comparison of the 1.31 A structure with that at 2.0 A shows the same basic features. However, the high-resolution electron-density maps clearly reveal additional solvent molecules and significant discrete disorder in protein side chains and within the solvent structure. As a consequence of modelling side-chain disorder, several new hydrogen-bonding interactions were identified. PMID- 15299632 TI - Data reduction from twinned RNA crystals. AB - Methods were developed to process diffraction data from epitaxically twinned crystals. Four programs for data reduction and two display programs were developed to augment the data-reduction program XDS [Kabsch (1988). J. Appl. Cryst. 21, 916-924]. The programs can be generalized for use with other data reduction software that provides the user with a list of the reflections used to determine lattice constants and crystal orientation. LATTICE_VIEW generates a PDB file containing 'water molecules' at the reciprocal-space coordinates of the strong spots found in the initial data frames. The PDB file is visualized to identify spots that belong to the same lattice, obtain unit-cell dimensions for a lattice, and assess data quality. VECTOR_MATCH is used to find additional spots belonging to a lattice. ACCOUNT4 determines which spots have been processed by XDS. COMFORT discards reflections that are too close to a reflection in another lattice. The display programs provide useful visual information on the quality of the crystal orientations used. Data with an R(merge) of 7.1% at 2.4 A resolution were obtained from epitaxically twinned crystals of an RNA dodecamer. The data were of sufficient quality to solve the structure with a combination of molecular replacement and single isomorphous replacement methods. PMID- 15299633 TI - A pseudo-cell based approach to efficient crystallographic refinement of viruses. AB - Strategies have been developed for the inexpensive refinement of atomic models of viruses and of other highly symmetric structures. These methods, which have been used in the refinement of several strains of poliovirus, focus on an arbitrary sized parallelepiped (termed the 'protomer' box) containing a single complete averaged copy of the structural motif which forms the protein capsid, together with the fragments of other symmetry-related copies of the motif which are located in its immediate neighborhood. The Fourier transform of the protomer box provides reference structure factors for stereochemically restrained crystallographic refinement of the atomic model parameters. The phases of the reference structure factors are based on the averaged map, and are not permitted to change during the refinement. It is demonstrated that models refined using the protomer box methods do not differ significantly from models refined by more expensive full-cell calculations. PMID- 15299634 TI - Structure of bovine eye lens gammaD (gammaIIIb)-crystallin at 1.95 A. AB - The crystal structure of bovine lens gammaIIIb-crystallin at 2.5 A resolution previously reported was interpreted using a consensus sequence derived from related vertebrate sequences on the assumption that gammaIIIb-crystallin derived from the gammaC-crystallin gene. It has recently been shown that gammaIIIb is a product of the bovine gammaD gene. The structure of gammaIIIb has now been refined with the bovine gammaD sequence using new 1.95 A resolution synchrotron data. The crystallographic R factor was 20.4% for all 33 104 reflection data between 8.0 and 1.95 A measured at 277(1) K. The electron density fully supported the assignment of the gammaD sequence to gammaIIIb. The crystal belongs to space group P2(1)2(1)2(1) with two molecules of molecular mass 20 749 Da in the asymmetric unit in which 219 water molecules were located. The two-domain four Greek-key motif highly symmetrical protein is very similar in structure to gammaB crystallin (81% sequence identity). There is a single amino-acid deletion in gammaD in the linker region connecting the two domains. The intermolecular oganization in the crystal lattice is quite different from gammaB as a result of key mutations involving surface residues Leu51, Ile103 and His155. These point mutations will contribute to the intermolecular behaviour of the gamma crystallins in the eye lens, where they are major components of the densely packed, high refractive index regions of the lens. PMID- 15299635 TI - Molecular rigid-body displacements in a tetragonal lysozyme crystal confirmed by X-ray diffuse scattering. AB - X-ray diffuse scattering from protein crystals is, at the moment, the only available experimental process to be directly sensitive to long-range correlations between protein-atom displacements. It is shown here that calculations based on independent rigid-body displacements of individual molecules yield a description in good agreement with the experimental diffuse scattering pattern displayed by tetragonal crystals of hen egg-white lysozyme (HEWL) In particular, it appears that molecular rigid-body translations and rigid body rotations appear roughly in the same proportion as the average atomic mean square positional fluctuations. The crystallographic temperature-factor analysis by TLS (translation/libration/screw) refinement, performed by Sternberg, Grace & Phillips [Sternberg, Grace, & Phillips (1979). J. Mol. Biol. 130, 231-253], is then confirmed and completed by a quantitative estimation of the molecular rigid body translation contributions. The major contribution of molecular rigid- body displacements to the average atomic mean-square positional fluctuations, contradicts a previous analysis of the tetragonal HEWL diffuse-scattering data by Clarage, Clarage, Phillips, Sweet & Caspar [Clarage, Clarage, Phillips, Sweet & Caspar (1992). Proteins Struct. Funct. Genet. 12, 145-157] which concluded that short-range correlations dominate. The origin of these opposite conclusions mostly lies in the different hypotheses made to model diffuse scattering, underlying the limits of the 'homogeneous disorder' model. PMID- 15299636 TI - Structure of porcine pancreatic spasmolytic polypeptide at 1.95 A resolution. AB - The structure of a trigonal crystal form of porcine pancreatic spasmolytic polypeptide (PSP) has been solved by molecular replacement and refined to 1.95 A resolution. Three heavy-atom derivatives were prepared, giving unbiased phase information, which was used in the model building of the protein molecules. The final conventional R value is 19.8% with the inclusion of 183 water molecules. PSP crystallizes as a dimer in space group P3(1)21 with a non-crystallographic twofold axis relating the monomers. The monomer consists of two very similar domains each composed of three loop regions. Two clefts are found in the monomer, one in each domain, that are proposed as possible substrate-binding sites. Important interactions have been identified in the proposed substrate-binding sites, where conserved water molecules probably mimic the hydrophilic positions of the substrate. The estimated cleft size is 9 x 9 x 12 A. Analysis of the charge distribution within the clefts, by an electrostatic potential calculation, shows the clefts to be essentially non-charged. PMID- 15299637 TI - Crystallization and preliminary X-ray diffraction studies of new crystal forms of Escherichia coli P(II) complexed with various ligands. AB - New crystals of the signal-transducing protein P(II) have been obtained in the presence of a number of different effector ligands. Various crystal forms are observed depending on the nature of the ligand(s). Co-crystallization with 2 ketoglutarate, glutamate and pyrophosphate produces hexagonal crystals similar to the wild type, ATP yields cubic crystals and ATP in conjunction with 2 ketoglutarate or glutamate yields orthorhombic crystal forms. All of the above crystals have been characterized by X-ray diffraction analysis. The hexagonal crystals belong to space group P6(3), cubic crystals to either I23 or I2(1)3 and orthorhombic crystals to I222. A molecular-replacement solution for the P(II)/ATP/2-ketoglutarate crystals has been obtained giving us an initial model for a trimer in the orthorhombic crystal form. PMID- 15299638 TI - Bayesian weighting for macromolecular crystallographic refinement. AB - A simple weighting scheme for atomic refinement is discussed. The approach, called 'Bayesian weighting', is designed to be robust with respect to the bias that arises from the incomplete nature of the atomic model, which in macromolecular crystallography is typically quite serious. Bayesian weights are based on the mean-squared residual errors over shells of resolution, with centric and acentric reflections considered separately and with allowances made for experimental uncertainties. Use of Bayesian weighting is shown in test cases typical for macromolecular crystallography to improve the accuracy of the refined coordinates when compared with schemes employing unit weights or experimental variances. PMID- 15299639 TI - Correlated phasing of multiple isomorphous replacement data. AB - Substantial highly correlated differences sometimes exist between a series of heavy-atom derivatives of a macromolecule and the native structure. Use of such a series of derivatives for phase determination by multiple isomorphous replacement (MIR) has been difficult because MIR analysis has treated errors as independent. A simple Bayesian approach has been used to derive probability distributions for the phase in the case where a group of MIR derivatives have correlated errors. The utility of the resulting 'correlated-phasing' method has been examined by applying it to both simulated and real MIR data sets that contain sizeable correlated errors and it has been found that it can dramatically improve MIR phase estimates in these cases. Correlated phasing is applicable to situations where derivatives exhibit substantial correlated changes in protein conformation or crystal packing or where correlated errors in heavy-atom models are large. Correlated phasing does not substantially increase the complexity of phase computation and is suitable for routine use. PMID- 15299640 TI - Structures of human transthyretin complexed with thyroxine at 2.0 A resolution and 3',5'-dinitro-N-acetyl-L-thyronine at 2.2 A resolution. AB - The molecular structures of two human transthyretin (hTTR, prealbumin) complexes, co-crystallized with thyroxine (3,5,3',5'-tetraiodo-L-thyronine; T(4)), and with 3',5'-dinitro-N-acetyl-LL-thyronine (DNNAT), were determined by X-ray diffraction methods. Crystals of both structures are orthorhombic, space group P2(1)2(1)2, and have two independent monomers in the asymmetric unit of the crystal lattice. These structures have been refined to 17.0% for 8-2.0 A resolution data for the T(4) complex (I), and to R = 18.4% for 8-2.2 A resolution data for the DNNAT structure (II). This report provides a detailed description of T(4) binding to wild-type hTTR at 2.0 A resolution, as well as DNNAT. In both structures, the two independent hormone-binding sites of the TTR tetramer are occupied by ligand. A 50% statistical disorder model was applied to account for the crystallographic twofold symmetry along the binding channel and the lack of such symmetry for the ligands. Results for the co-crystallized T(4) complex show that T(4) binds deep in the hormone-binding channel and displaces the bound water previously reported for T(4) soaked into a native transthyretin crystal [Blake & Oatley (1977). Nature (London), 268, 115-120]. DNNAT also binds deeper in the channel toward the tetramer center than T(4) with the nitro groups occupying the symmetrical innermost halogen pockets. The N-acetyl moiety does not form polar contacts with the protein side chains as it is oriented toward the center of the channel. The weak binding affinity of DNNAT results from the loss of hydrophobic interactions with the halogen binding pockets as observed in T(4) binding. These data suggest that the halogen-binding sites toward the tetramer center are of primary importance as they are occupied by analogues with weak affinity to TTR, and are therefore selected over the other halogen sites which contribute more strongly to the overall binding affinity. PMID- 15299641 TI - Formation of (C.G)*G triplets in a B-DNA duplex with overhanging bases. AB - Crystallization of a DNA double helix with overhanging bases at the 5'-ends of both strands, results in the formation of two crystallographically independent (C.G)*G triplets. In a previous report [Van Meervelt, Vlieghe, Dautant, Gallois, Precigoux & Kennard (1995). Nature (London), 374, 742-744] the unique molecular packing of the duplex and the Hoogsteen hydrogen-bond pattern and parallel backbone orientation of the guanine-containing strands in the triplets was described. The fine structural details and hydration of the d(GCGAATTCG) crystal structure refined to 2.05 A (R = 0.168, 86 water molecules, two Mg(2+) cations) are now presented. Helical parameters, stacking effects, the geometry at the duplex-triplex junction, and the hydration of the minor groove are discussed and compared with related theoretical and crystal structures. PMID- 15299642 TI - Heterogeneity determination and purification of commercial hen egg-white lysozyme. AB - Hen egg-white lysozyme (HEWL) is widely used as a model protein, although its purity has not been adequately characterized by modern biochemical techniques. We have identified and quantified the protein heterogeneities in three commercial HEWL preparations by sodium dodecyl sulfate polyacrylamide gel electrophoresis with enhanced silver staining, reversed-phase fast protein liquid chromatography (FPLC) and immunoblotting with comparison to authentic protein standards. Depending on the source, the contaminating proteins totalled 1-6%(w/w) and consisted of ovotransferrin, ovalbumin, HEWL dimers, and polypeptides with approximate M(r) of 39 and 18 kDa. Furthermore, we have obtained gram quantities of electrophoretically homogeneous [> 99.9%(w/w)] HEWL by single-step semi preparative scale cation-exchange FPLC with a yield of about 50%. Parallel studies of crystal growth kinetics, salt repartitioning and crystal perfection with this highly purified material showed fourfold increases in the growth-step velocities and significant enhancement in the structural homogeneity of HEWL crystals. PMID- 15299643 TI - Repartitioning of NaCl and protein impurities in lysozyme crystallization. AB - Nonuniform precipitant and impurity incorporation in protein crystals can cause lattice strain and, thus, possibly decrease the X-ray diffraction resolution. To address this issue, a series of crystallization experiments were carried out, in which initial supersaturation, NaCl concentration, protein purity level and crystallized fraction were varied. Lysozyme and protein impurities, as well as sodium and chloride were independently determined in the initial solution, supernatant and crystals. The segregation coefficients for Na(+) and Cl(-) were found to be independent of supersaturation and NaCl concentration, and decreased with crystallized fraction/crystal size. Numerical evaluation of the extensive body of data, based on a nucleation-growth-repartitioning model, suggests a core of approximately 40 micro m in which salt is incorporated in much greater concentrations than during later growth. Small crystals containing higher amounts of incorporated NaCl also had higher protein impurity contents. This suggests that the excess salt is associated with the protein impurities in the core. X-ray topography revealed strain fields in the center of the crystals comparable in size to the inferred core. The growth rates of crystals smaller than 30-40 micro m in size were consistently 1.5-2 times lower than those of larger crystals, presumably due to higher chemical potentials in the core. PMID- 15299644 TI - Interconversion of crystals of the Escherichia coli EF-Tu.EF-Ts complex between high- and low-diffraction forms. AB - Crystals of the complex formed between the two bacterial polypeptide elongation factors, EF-Tu and EF-Ts, produced from solutions of PEG 6000 can be of two morphologically similar forms both of space group P2(1)2(1)2(1). One form diffracts to only about 3 A resolution, the other to better than 2.4 A resolution. These forms can be interconverted and the transformation of one into the other has been shown to be solely a result of dehydration/hydration processes. By designing a suitable soaking protocol and careful control of the experimental parameters for data collection at cryotemperatures, complete data sets for the high-resolution form could be obtained. PMID- 15299645 TI - Contaminant effects on protein crystal morphology in different growth environments. AB - Contaminant effects on the morphology of turkey egg-white lysozyme (TEWL) crystals grown in ungelled and in gelled growth media have been investigated. The latter may serve as a model system for future microgravity experiments. Hen egg white lysozyme (HEWL) was added as the contaminant at levels ranging from 4 up to 70%(w/w) (total protein). Morphology measurements indicate a contaminant effect leading to a shortening along the c axis of the crystal. This shortening effect depends on the contaminant concentration. It is attenuated and varies more regularly in gelled than in ungelled growth media. The specificity of the HEWL contaminant effect was verified by addition of ribonuclease A, which did not influence crystal morphology. Contaminant inclusion into the growing TEWL crystals could be calculated directly from equilibrium protein concentration measurements. The level of HEWL inclusion is closely related to the concentration of HEWL in the growth solutions. The specificity of the observed effect as well as the differences between the two growth media are discussed. PMID- 15299646 TI - The ab initio crystal structure solution of proteins by direct methods. VI. Complete phasing up to derivative resolution. AB - The procedure described in the papers I-V of this series [Giacovazzo, Siliqi & Ralph (1994). Acta Cryst. A50, 503-505; Giacovazzo, Siliqi & Spagna (1994). Acta Cryst. A50, 609-621; Giacovazzo, Siliqi & Zanotti (1995). Acta Cryst. A51, 177 188; Giacovazzo & Gonzalez Platas (1995). Acta Cryst. A51, 398-404; Giacovazzo, Siliqi & Gonzalez Platas (1995). Acta Cryst. A51, 811-820], aiming at estimating protein phases via a single heavy-atom derivative, has been improved so as to extend phase determination to all the reflections up to derivative resolution. The quality of the resulting electron-density maps is checked for a number of test strutures. Some of the maps are immediately interpretable, and some can be interpreted after some cycles of solvent flattening and/or histogram matching. The correlation with classical SIR techniques is also discussed. PMID- 15299647 TI - xdlMAPMAN and xdlDATAMAN - programs for reformatting, analysis and manipulation of biomacromolecular electron-density maps and reflection data sets. AB - Two user-friendly computer programs are described for use in macromolecular X-ray crystallography, xdlMAPMAN provides an interface for electron-density map exchange between some of the most commonly used phase refinement, structure refinement and model- building programs. In addition, it contains several options to analyse and abstract such maps. xdlDATAMAN provides similar functionality for the analysis and manipulation of macromolecular reflection data sets. Both programs have a simple graphical user interface, and their source code has been put into the public domain. PMID- 15299648 TI - Efficient rebuilding of protein structures. AB - A computer program, called OOPS, is described which facilitates and speeds up the process of rebuilding a protein structure inside its electron density and reduces the chances of local errors persevering throughout the crystallographic protein structure determination process. The program uses a set of criteria to judge how reasonable each protein residue is and it generates macros for the macromolecular crystallographic model-building program O [Jones, Zou, Cowan & Kjeldgaard (1991). Acta Cryst. A47, 110-119] which, when executed, will take the crystallographer on a journey along all suspect residues. PMID- 15299649 TI - Towards the automatic interpretation of macromolecular electron-density maps: qualitative and quantitative matching of protein sequence to map. AB - The matching of the known polypeptide sequence to the electron density is a critical step in solving protein structures by the crystallographic method. Tools have been developed to help in defining the placement of the sequence, both qualitatively and quantitatively. They have been tested with good results on two proteins whose structures were solved by the MIR method. PMID- 15299650 TI - Use of non-crystallographic symmetry in protein structure refinement. AB - Several methods to assess the (dis)similarity of protein structures objectively are described, some of which, when applied to non-crystallographically related protein models, are able to discriminate between significant differences and 'random noise'. Some of these methods have been used to investigate a sample of several hundred protein structures which have been solved by means of X-ray crystallography in order to investigate the extent to which non crystallographically related protein models differ from one another. It is shown that the extent of such differences is largely dependent on the resolution of the data used for the determination and refinement of the structure and, measured by some statistics, even varies essentially linearly with the resolution. The implications of these findings for the strategies used to refine structures with non-crystallographic symmetry, in particular at low resolution, are discussed. Finally, two examples are given of recent structure determinations from this laboratory in which the presence (and employment) of non-crystallographic symmetry was crucial to the solution and refinement of the structure. PMID- 15299651 TI - A re-evaluation of the crystal structure of chloromuconate cycloisomerase. AB - It is shown here that the reported 3 A crystal structure of chloromuconate cycloisomerase from Alcaligenes eutrophus [Hoier, Schlomann, Hammer, Glusker, Carrell, Goldman, Stezowski & Heinemann (1994). Acta Cryst. D50, 75-84] was refined in the incorrect space group I4. In addition, a stretch of about 25 residues near the N-terminus is out-of-register with the density in the original structure. From the coordinates and structure factors deposited in the Protein Data Bank (PDB), it was possible to determine the correct space group to be I422. The structure was then re-refined, using the original data reduced to I422, to a crystallographic free R factor of 0.264 at 3 A resolution (conventional R factor 0.189). With conservative refinement and rebuilding methods, the errors in the chain tracing could be identified and remedied. Since the two molecules per asymmetric unit in the original structure are actually related by crystallographic symmetry, the observed differences between them are artefacts. In particular, the differences between, and peculiarities of the metal-binding sites are unreal. This case shows the dangers of crystallographic refinement in cases with unfavourable data-to-parameter ratios, and the importance of reducing the number of parameters in such cases to prevent gross errors (for instance, by using NCS constraints). It also demonstrates how the evaluation and monitoring of model quality during the entire refinement and rebuilding process can be used to detect and remedy serious errors. Finally, it presents a strong case in favour of depositing not only model coordinates, but also experimental data (preferably, both merged and unmerged data). PMID- 15299652 TI - Crystallization and preliminary X-ray analysis of a vanadium-dependent peroxidase from Ascophyllum nodosum. AB - Peroxidase from the brown alga Ascophyllum nodosum, a non-heme vanadium-dependent haloperoxidase, has been purified to homogeneity and crystallized from ammonium sulfate solutions in a form suitable for X-ray diffraction analysis. The crystals have been grown by the vapour-diffusion technique using the sitting-drop method. X-ray diffraction studies show that the crystals belong to the tetragonal space group P4(1)2(1)2 or P4(3)2(1)2 with a = b = 114.3 and c = 276.0 A. The crystals diffract to at least 2.4 A resolution. PMID- 15299653 TI - Crystallization and preliminary X-ray study of the 8-amino-7-oxopelargonate synthase from Bacillus sphaericus. AB - The 8-amino-7-oxopelargonate synthase (AOPS) cloned from Bacillus sphaericus, overproduced in Escherichia coli, has been crystallized in the pyridoxal 5' phosphate (PLP)-bound form at pH 7.5, using polyethylene glycol as the precipitant. One crystal form corresponds to a tetragonal space group, with unit cell dimensions a = b = 66, c = 181 A. These crystals do not diffract beyond 5 A, with conventional X-ray sources and cannot be used in the structure elucidation. A second crystal form is obtained when crystallization conditions are varied slightly by the addition of 0.2 M ammonium sulfate. The space group is P2(1)2(1)2(1), with unit-cell dimensions a = 68.9, b = 85.5, c = 125.9 A, indicating the presence of two molecules in the asymmetric unit (V(m) = 2.26 A(3) Da(-1); 46% water). These crystals diffract X-rays up to 3.2 A using in-house facilities and a preliminary data set has been collected. A second data set using the synchrotron radiation source W32 at LURE (Paris) has shown the crystals to diffract to at least 3 A, resolution, with good statistics. The structure determination of AOPS will provide a structural framework for the other alpha amino ketone synthases for which no three-dimensional structure is yet available. PMID- 15299654 TI - Preliminary X-ray crystallographic analysis of human salivary cystatin. AB - Human salivary cystatin, a thiol proteinase inhibitor, has been implicated in potential antimicrobial and antiviral functions of saliva. A variant of human salivary cystatin SN expressed and purified in an Escherichia coli expression system lacking residues 12-16 near the N-terminus (Delta12-16) has been crystallized by the vapor-diffusion technique. The crystals are of the hexagonal space group P622 and have cell constants of a = 85.41, b = 85.41, c = 131.6 A, alpha = beta = 90, gamma = 120 degrees, and contain two molecules of molecular weight 13 500 per asymmetric unit. The crystals diffract up to a resolution of 2.2 A and are suitable for X-ray diffraction analysis. PMID- 15299655 TI - Crystals of the chemically synthesized acceptor stem of tRNAAla from Escherichia coli diffracting to high resolution. AB - The acceptor stem of tRNA(Ala) from E. coli has been chemically synthesized and crystallized. This duplex contains a G.U base pair in position 3-70, which is the main identity element for alanyl-tRNA synthetase from E. coli. The crystals are stable in the X-ray beam for a long period of time and diffract to 1.7 A resolution. The monoclinic crystals reveal a C2 space group with a = 35.0, b = 47.5, c = 26.2 A, beta = 102.3 degrees and one acceptor stem per asymmetric unit. PMID- 15299656 TI - Crystallization of rat procathepsin B. AB - Rat procathepsin B has been expressed in the yeast Pichia pastoris. To facilitate crystallization of the proform two mutations were introduced: Cys29Ser to avoid self-processing and Ser115Ala to eliminate an N-glycosylation site. The recombinant protein was purified and crystallized by vapor diffusion against mother liquor containing 100 mM KSCN, 100 mM phosphate buffer, pH 6.5 and polyethylene glycol (PEG) 3350 as a precipitating agent. Crystal size was increased by multiple macroseeding. At a 16% PEG concentration trigonal crystals were obtained, with the space group P3(1)21 and a = 99.6, c = 141.4 A, gamma = 120 degrees. They diffract to 2.8 A resolution using a rotating-anode source. At a concentration of 11% PEG, rod-shaped crystals were grown. They are monoclinic, space group P2(1), a = 62.8, b = 67.9, c = 100.4 A, beta = 98.2 degrees and diffract to approximately 3.5 A. PMID- 15299657 TI - Crystallization and preliminary diffraction studies of firefly luciferase from Photinus pyralis. AB - Firefly luciferase is a 62 kDa molecular weight enzyme which catalyzes a light emitting reaction. Crystals of Photinus pyralis luciferase have been obtained by the microbatch technique, using polyethylene glycol as a precipitating agent. Firefly luciferase crystallizes as long needles which belong to the tetragonal space group P4(1)2(1)2, with cell dimensions a = 119.5, b = 119.5, c = 95.4 A. One molecule is present in the asymmetric unit. Diffracted intensities beyond 2.0 A resolution have been measured from frozen crystals using synchrotron radiation. PMID- 15299658 TI - Concanavalin A crystallized in complex with the trisaccharide 3,6-di-O-methyl (alpha-D-mannopyranosyl)-alpha-D-mannopyranoside. AB - Concanavalin A was co-crystallized in two crystal forms with 3,6-di-O-methyl- (alpha-D-mannopyranosyl) alpha- D-mannopyranoside, which is primarily responsible for the high-affinity binding of N-linked carbohydrates to concanavalin A. Both crystal forms have space group P2(1) and contain a complete concanavalin A tetramer in the asymmetric unit. Form A was crystallized using polyethylene glycol methyl ether as the precipitant and has unit-cell dimensions a = 59.83, b = 64.84 and c = 125.92 A, beta = 93.87 degrees. Form B was obtained using phosphate as the precipitant and has unit-cell dimensions a = 81.94, b = 66.75 and c = 108.92 A, beta = 97.58 degrees. Form B was stable in the X-ray beam for several days and diffracted to 3.15 A resolution. Form A crystals could not withstand X-ray radiation at room temperature, but produced high-quality data under cryogenic conditions. The latter are suitable for a 2.3 A resolution structure determination by molecular replacement. PMID- 15299659 TI - Crystalline alcohol dehydrogenases from the mesophilic bacterium Clostridium beijerinckii and the thermophilic bacterium Thermoanaerobium brockii: preparation, characterization and molecular symmetry. AB - Two tetrameric NADP(+)-dependent bacterial secondary alcohol dehydrogenases have been crystallized in the apo- and the holo-enzyme forms. Crystals of the holo enzyme from the mesophilic Clostridium beijerinckii (NCBAD) belong to space group P2(1)2(1)2(1) with unit-cell dimensions a = 90.5, b = 127.9, c = 151.4 A. Crystals of the apo-enzyme (CBAD) belong to the same space group with unit-cell dimensions a = 80.4, b = 102.3, c = 193.5 A. Crystals of the holo-enzyme from the thermophilic Thermoanaerobium brockii (NTBAD) belong to space group P6(1(5)) (a = b = 80.6, c = 400.7 A). Crystals of the apo-form of TBAD (point mutant GI98D) belong to space group P2(1) with cell dimensions a = 123.0, b = 84.8, c = 160.4 A beta = 99.5 degrees. Crystals of CBAD, NCBAD and NTBAD contain one tetramer per asymmetric unit. They diffract to 2.0 A resolution at liquid nitrogen temperature. Crystals of TBAD(GI98D) have two tetramers per asymmetric unit and diffract to 2.7 A at 276 K. Self-rotation analysis shows that both enzymes are tetramers of 222 symmetry. PMID- 15299660 TI - Crystallographic studies and preliminary X-ray investigation of (S)-p-hydroxy mandelonitrile lyase from Sorghum bicolor (L.). AB - (S)-p-Hydroxy-mandelonitrile lyase from Sorghum bicolor has been crystallized in three different forms using the hanging-drop vapor-diffusion technique. Crystal form I is obtained from 1.4 M (NH(4))(2)SO(4) in 100 mM Na-acetate, pH 4.6, and belongs to the orthorhombic space group P2(1)2(1)2(1). The cell dimensions are a = 71.4, b = 95.8, c = 149.1 A. A complete set of diffraction data has been collected to 2.6 A resolution. Form II crystals are grown from 500 mM Li(2)SO(4) in 13% polyethylene glycol 8000. These crystals appear as hexagonal plates and diffract to 2.98 A resolution but apparently are twinned. Cocrystallizing hydroxynitrile lyase with the inhibitor benzoic acid using 1.4 M (NH(4))(2)SO(4) in 100 mM Na citrate, pH 5.4 as precipitant yields crystal form III, which belongs to the monoclinic space group C2 with a = 150.7, b = 103.7, c = 90.6 A, beta = 101.3. X-ray diffraction data were collected to 2.3 A resolution. PMID- 15299661 TI - A new crystal form of carboxypeptidase G2 from Pseudomonas sp. strain RS-16 which is more amenable to structure determination. AB - Carboxypeptidase G(2) is a zinc-dependent exopeptidase which has applications in cancer therapy. Crystallization of carboxypeptidase G(2), first achieved more than a decade ago, yields large crystals; however, problems with non-isomorphism between native crystals as well as failure to obtain any useful heavy-atom derivatives have precluded structure solution. A modification of the crystallization protocol leading to a promising new crystal form which diffracts beyond 3.0 A resolution on a rotating-anode source is now reported. These crystals are readily indexed on an apparent C-centred orthorhombic lattice with a = 81.35, b = 230.9 and c = 105.5 A, but the correct crystal system is monoclinic. The crystals have space group P2(1), with a = 81.35, b = 105.5, c = 122.4 A and beta = 109.3 degrees. There are two possible non-equivalent monoclinic indexings with these lattice constants. A partial native data set collected at the SRS, Daresbury, indicates that 1.9 A diffraction is attainable. Structure determination using MIR methods is in progress. PMID- 15299662 TI - The papers of Dorothy Mary Crowfoot Hodgkin. PMID- 15299663 TI - MAGICSQUASH: more versatile non-crystallographic averaging with mulitple constraints. AB - The simultaneous application of multiple constraints such as non-crystallographic symmetry averaging, solvent levelling, histogram matching and Sayre's equation has proven to be very effective for phase refinement and extension. An existing program for this purpose, SQUASH, has been altered to increase the flexibility of its non-crystallographic symmetry averaging. The modified program, MAGICSQUASH, can handle multiple-domain averaging and multiple space group averaging and has aided in the solution of several structures. Examples are described which include the first simultaneous application of solvent levelling, histogram matching and Sayre's equation in two crystal forms while averaging between them. PMID- 15299664 TI - Structure of human salivary alpha-amylase at 1.6 A resolution: implications for its role in the oral cavity. AB - Salivary alpha-amylase, a major component of human saliva, plays a role in the initial digestion of starch and may be involved in the colonization of bacteria involved in early dental plaque formation. The three-dimensional atomic structure of salivary amylase has been determined to understand the structure-function relationships of this enzyme. This structure was refined to an R value of 18.4% with 496 amino-acid residues, one calcium ion, one chloride ion and 170 water molecules. Salivary amylase folds into a multidomain structure consisting of three domains, A, B and C. Domain A has a (beta/alpha)(8-) barrel structure, domain B has no definite topology and domain C has a Greek-key barrel structure. The Ca(2+) ion is bound to Asnl00, Arg158, Asp167, His201 and three water molecules. The Cl(-) ion is bound to Arg195, Asn298 and Arg337 and one water molecule. The highly mobile glycine-rich loop 304-310 may act as a gateway for substrate binding and be involved in a 'trap-release' mechanism in the hydrolysis of substrates. Strategic placement of calcium and chloride ions, as well as histidine and tryptophan residues may play a role in differentiating between the glycone and aglycone ends of the polysaccharide substrates. Salivary amylase also possesses a suitable site for binding to enamel surfaces and provides potential sites for the binding of bacterial adhesins. PMID- 15299665 TI - Locating a local symmetry axis from patterson map cross vectors: application to crystal data from GroEl, GTP cyclohydrolase I and the proteosome. AB - The cross vectors of the native Patterson map are shown to exhibit non crystallographic symmetry in the case of local axes parallel to one another. This information can be used to determine the translation component of such axes. A program is described to search for this cross vector, and is tested on low resolution data from crystals of the tetradecameric GroEL molecule, the decameric GTP cyclohydrolase I and the tetradecameric proteosome. For GroEL, the function produces a packing arrangement optimal for sevenfold symmetry, and is in agreement with the dimensions of the molecule as given by electron microscopy data and the recently determined crystal structure. Positioning of local axes is confirmed by two high-resolution crystal structure analyses: the fivefold axis in cyclohydrolase I and the sevenfold axis in the proteosome. Implications for the location of heavy-atom positions are discussed for these two cases. PMID- 15299666 TI - Structure of a bovine thrombin-hirudin51-65 complex determined by a combination of molecular replacement and graphics. Incorporation of known structural information in molecular replacement. AB - Crystals of the bovine thrombin-hirudins(51-65) complex have space group P6(1)22 with cell constants a = 116.4, and c = 200.6 A and two thrombin molecules in the asymmetric unit. Only one thrombin molecule could be located by generalized molecular replacement; the second was fit visually as a rigid body to an improved electron-density difference map. The structure was refined to R = 0.192 with two B values per residue (main chain and side chain) at 3.2 A. The polar interactions of the peptides with the exosite of thrombin show differences consistent with the known flexibility in the interactions of the C-terminal peptide of hirudin with thrombin. The hirudin peptide in complex 2 has a higher temperature factor as compared with peptide 1 which may be correlated partly with a larger number of short-range electrostatic interactions between peptide 1 and thrombin and partly with the fact that thrombin 2 is epsilon-thrombin which is cleaved at Thr149A near the peptide binding site. Later, using this structure as a test case, it was shown that the position for the second thrombin could also be determined by a novel modification of the molecular-replacement method in which the contribution of the known molecule is subtracted from the structure factors. This approach is facile and applicable to any crystal containing two or more macromolecules in the asymmetric unit in which some but not all of the molecules can be determined by molecular replacement. PMID- 15299668 TI - A challenging case for protein crystal structure determination: the mating pheromone Er-1 from Euplotes raikovi. AB - Four different phasing methods have been applied to the determination of the crystal structure of the 40 amino-acid mating pheromone of the unicellular ciliated protozoan Euplotes raikovi. The difficulties, failures and successes in attempts to solve the structure by: (1) molecular replacement, (2) direct phasing using the 'Shake and Bake' algorithm, (3) isomorphous replacement, and (4) multiple-wavelength anomalous dispersion are described. The structure was first solved by molecular replacement, and then was the first successful structure determination by 'Shake and Bake' without the direct involvement of its authors. A description of the current status of the high-resolution refinement of the structure is also given. The model is refined against 1 A resolution data to an R factor of 12.9%, and includes H atoms and discretely disordered side chains. PMID- 15299669 TI - Structure of the DNA octanucleotide d(ACGTACGT)2. AB - d(ACGTACGT), C(78)H(84)N(30)O(32)P(7).20H(2)O, M(r) (DNA) = 2170, tetragonal, P4(3)2(1)2 (No 96), a = 42.845 (1), b = 42.845(1), c = 24.804 (1) A, V = 45532.5 (2) A(3), z = 8,lambda(MoKalpha) = 0.71069 A, micro (MoKalpha) = 0.10 mm(-1), T = 295 K, R = 0.18 for 1994 unique reflections between 5.0 and 1.9 A resolution. The self-complementary octanucleotide d(ACGTACGT)(2) has been crystallized and its structure determined to a resolution of 1.9 A. The asymmetric unit consists of a single strand of octamer with 20 water molecules. It is only the second example of an octanucleotide having terminal A.T base pairs whose structure has been determined by X-ray crystallography. The sequence adopts the modified A-type conformation found for all octanucleotide duplexes studied to date with the helix bent by approximately 15 degrees and an average tilt angle of 0 degrees. Unusually the data collection was carried out using a 3 kW molybdenum sealed-tube source. The conformational details are discussed in comparison with other closely related sequences. PMID- 15299670 TI - Structure determination of a lone alpha-helical antifreeze protein from winter flounder. AB - The X-ray crystal structure of a lone alpha-helical antifreeze protein from winter flounder has been determined to 1.5 A using a combination of molecular replacement and isomorphous-replacement techniques. Molecular replacement involved a multiparameter search using X-PLOR with two 37-mers of alanine in idealized alpha-helical conformations as the search models. Identified were a large number of potential solutions from which the correct solution was not distinguishable. Commitment of the top 1620 solutions to cycles of rigid-body, positional and simulated-annealing refinement identified the correct solution by a small margin in R factor. Low-resolution electron-density maps generated with phasing information from TbNO(3) and LaNO(3) derivatives were consistent with the top molecular-replacement solution. These derivatives also provided a means to filter and compare the large number of other molecular-replacement solutions with reasonable R factor statistics. The structure-solution strategy described herein may prove useful for the determination of other relatively simple alpha-helical X ray structures. PMID- 15299671 TI - Refined three-dimensional structure of cat-muscle (M1) pyruvate kinase at a resolution of 2.6 A. AB - The three-dimensional structure of cat-muscle pyoruvate kinase has been refined at a resolution of 2.6 A. The details of the structure permit interpretation of the original heavy-atom studies and give insight into the importance of conserved residues in pyruvate kinases and the allosteric behaviour of the enzyme. There are a small number of essential residues which determine the relative orientations of domains and the precise nature of intersubunit contacts. Arginine residues are particularly important. PMID- 15299672 TI - High-resolution structure (1.33 A) of a HEW lysozyme tetragonal crystal grown in the APCF apparatus. Data and structural comparison with a crystal grown under microgravity from SpaceHab-01 mission. AB - Crystals of tetragonal hen egg-white lysozyme were grown using Advanced Protein Crystallization Facility (APCF) apparatus under a microgravity environment (SpaceHab-01 mission) and ground control conditions. Crystals were grown from NaCl as a crystallizing agent at pH 4.3. The X-ray diffraction patterns of the best diffracting ground- and space-grown crystals were recorded using synchrotron radiation and an image plate on the W32 beamline at LURE. Both ground- and space grown crystals showed nearly equivalent maximum resolution of 1.3-1.4 A. Refinements were carried out with the program X-PLOR with final R values of 18.45 and 18.27% for structures from ground- and space- grown crystals, respectively. The two structures are nearly identical with the root-mean-square difference on all protein atoms being 0.13 A. Some residues of the two refined structures show multiple alternative conformations. Two ions were localized into the electron density maps of the two structures: one chloride ion at the interface between two symmetry-related molecules and one sodium ion stabilizing the loop Ser60-Leu75. The sodium ion is surrounded by six ligands which form a bipyramid around it at distances of 2.2-2.6 A. PMID- 15299673 TI - Phase combination. AB - In 1961 Rossmann & Blow published a simple procedure for analytically combining the phase probabilities derived from various isomorphous derivatives or other phase-determining procedures [Rossmann & Blow (1961). Acta Cryst. 14, 641-647]. However, they found it necessary to make an approximation in obtaining the expression for the lack of closure (epsilon) of the phase triangle. In 1970 Hendrickson & Lattman [Hendrickson & Lattman (1970). Acta Cryst. B26, 136-143] suggested an alternative method of defining the lack of closure of the phase triangle which did not require any approximation in deriving the same analytical expression for the phase-probability function. It is now shown that it is possible to avoid the Rossmann-Blow approximation and thereby maintain the original meaning of the lack of closure as defined by Blow & Crick [(1959). Acta Cryst. 12, 794-802] and Dickerson, Kendrew & Strandberg [(1961). Acta Cryst. 14, 1188-1195]. PMID- 15299674 TI - Structure of a Kunitz-type chymotrypsin from winged bean seeds at 2.95 A resolution. AB - Thc crystal structure of an alpha-chymotrypsin inhibitor (P6(1)22; a = 61.4, c = 210.9 A) isolated from winged bean (Psophocarpus. tetragonolobus) seeds has been determined at 2.95 A resolution by the molecular-replacement method using the 2.6 A coordinates of Erythrina trypsin inhibitor (ETI) as the starting model (57% sequence homology). This protease inhibitor, WCI, belongs to the Kunitz (STI) family and is a single polypeptide chain with 183 amino-acid residues having a molecular weight of 20 244 Da. Structure refinement with RESTRAIN and X-PLOR has led to a crystallographic R factor of 19.1% for 3469 observed reflections (I > 2sigma) in the resolution range 8-2.95 A. A total of 56 water molecules have been incorporated in the refined model containing 181 amino-acid residues. In the refined structure the deviations of bond lengths and bond angles from ideal values are 0.015 A and 2.2 degrees, respectively. The inhibitor molecule is spherical and consists of 12 antiparallel beta-strands with connecting loops arranged in a characteristic folding (a six-stranded beta-barrel and a six stranded lid on one hollow end of the barrel) common to other homologous serine protease inhibitors in the Kunitz (STI) family as well as to some non-homologous proteins like interleukin-lalpha and interleukin-lbeta. In the structure the conformation of the protruding reactive-site loop is stabilized through hydrogen bonds mainly formed by the side chain of Asnl4, which intrudes inside the cavity of the reactive-site loop, with the side-chain and main-chain atoms of some residues in the loop region. A pseudo threefold axis exists parallel to the barrel axis of the structure. Each of the three subdomains comprises of four beta strands with connecting loops. PMID- 15299675 TI - Lysozyme crystal growth kinetics monitored using a Mach-Zehnder interferometer. AB - A Mach-Zehnder interferometer has been developed for the monitoring of the kinetics of the diffusion process in protein crystal growth. This device can be used in conjunction with the ESA Advanced Protein Crystallization Facility (APCF), which allows experiments under microgravity conditions (e.g. on board the NASA Space Shuttle). Experimental trials on the ground have been carried out with the interferometer using the engineering model of the APCF and a protein dialysis reactor. Chicken egg-white lysozyme crystal growth, as a test, has thereby been monitored directly. The changes of concentration in the solution over time have been determined via the refractive index measurements made and subsequently correlated with visual monitoring of crystal growth in a repeat experiment. PMID- 15299676 TI - Structure of the fab fragment of SDZ CHI621: a chimeric antibody against CD25. AB - A specific drug targeted to the IL-2 receptor on activated T lymphocytes could limit acute immunological rejection during organ transplantation. A high-affinity monoclonal antibody directed against the alpha-chain of the IL-2 receptor (CD25) was chimerized with the constant regions of the human IgG1 heavy and k light chain resulting in SDZ CHII621 [Amlot, Rawlings, Fernando, Griffin, Heinrich, Schreier, Castaigne, Moore & Sweny (1995). Transplantation, 60, 748-756]. The Fab fragment of SDZ CHI621 has been purified and crystallized (P2(l), a = 39.58, b = 59.76, c = 102.09 A, beta = 99.98 degrees ). Its structure has been determined by molecular replacement and refined at 2.6 A to a crystallographic R factor of 19.7%. The protein exhibits the typical immunoglobulin fold. The complementary determining regions (CDR's) 1 and 2 of both heavy and light chains show similar conformations to other known reported structures whereas the CDR3 from the light chain seems to adopt a novel type of conformation. There is a network of interactions which maintain the CDR3 of both chains together and limit their solvent accessibility. The interaction between V(L) and C(L) has been strengthened by the chimerization whereas that between V(H) and C(H)1 has been weakened. PMID- 15299677 TI - E144S active-site mutant of the Bacillus cereus thermolysin-like neutral protease at 2.8 A resolution. AB - The X-ray crystal structure of the Bacillus cereus neutral protease (CNP) active site mutant E144S, in which the putative general base proposed for the thermolysin-like zinc neutral proteases, Glu144, has been replaced by serine, has been determined to a resolution of 2.8 A. This represents the first crystal structure of an active-site mutant of a zinc neutral protease. The E 144S mutant was crystallized in the hexagonal space group, P6(5)22, with unit-cell dimensions a = b = 76.57, c = 201.91 A. Although the ligands involved in zinc coordination in the active site are identical to those found in the wild-type protein, the mutation results in a modified environment around the zinc ion; particularly with respect to the water molecules. While the structure of the mutant is similar to that of wild type, its protease activity is reduced to 0.16% that of the wild type CNP and the protein is virtually resistant to autolysis in the presence of calcium. The lowered protease activity of the mutant is consistent with the role proposed for Glu144 as the general base in the catalysis of thermolysin-like neutral proteases [Matthews (1988). Acc. Chem. Res. 21, 333-340]. We suggest that the residual activity of the E144S mutant arises from a water molecule, which is found within hydrogen-bonding distance of Ser144, acting as a general base in the catalytic function of the mutant. PMID- 15299678 TI - Structure of Escherichia coli inorganic pyrophosphatase at 2.2 A resolution. AB - The refined crystal structures of hexameric soluble inorganic pyrophosphatase from E. coli (E-PPase) are reported to R factors of 18.7 and 18.3% at 2.15 and 2.2 A, respectively. The first contains one independent monomer; the other, two independent monomers, in an R32 unit cell. Because the E-PPase monomer is small with a large open active site, there are relatively few hydrophobic interactions that connect the active-site loops to the five-stranded twisted beta-barrel that is the hydrophobic core of the molecule. The active-site loops are, however, held in place by interactions between monomers around the threefold and twofold symmetry axes of the D(3) hexamer. Consequently, mutations of active-site residues (such as Glu20 and Lysl04) often affect protein stability and oligomeric structure. Conversely, mutations of residues in the interface between monomers (such as His136 and Hisl40) not only affect oligomeric structure but also affect active-site function. The effects of the H136Q and H140Q variants can be explained by the extended ionic interaction between H140, D143 and H136' of the neighbouring monomer. This interaction is further buttressed by an extensive hydrogen-bonding network that appears to explain why the E-PPase hexamer is so stable and also why the H136Q and H140Q variant proteins are less stable as hexamers. PMID- 15299679 TI - Crystallization and preliminary X-ray diffraction studies of recombinant staphylokinase. AB - Staphylokinase, a fibrin-specific plasminogen activator, was highly expressed in Escherichia coli and purified by ion-exchange and gel-filtration chromatography. The purified recombinant staphylokinase was fully active and readily crystallized against 1.2 M sodium citrate in 100 mM Tris-HCl buffer at pH 8.0 using the hanging-drop method. Crystals of staphylokinase diffract to better than 2.2 A resolution. The crystal belongs to the tetragonal space group P4(1)2(1)2 or its enantiomorph with unit-cell parameters a = b = 67.5, c = 150.1 A. There are two molecules in the asymmetric unit. In this paper, we described the first crystallization of a kind of plasminogen activator and present the results of preliminary X-ray diffraction data from the native protein. PMID- 15299680 TI - Crystallization and preliminary X-ray diffraction studies of Leu55Pro variant transthyretin. AB - The amyloidogenic Leu55Pro variant of transthyretin has been expressed, purified and crystallized in space group C2. The cell constants are a = 149.99, b = 78.74, c = 98.95 A, beta = 100.5 degrees and the crystals diffract to 2.7 A resolution. There are eight monomers in the asymmetric unit giving a V(M) = 2.6 A(3) Da(-1) and 53% solvent content. In the wild-type protein, the crystals are orthorhombic with two monomers in the asymmetric unit. The wild-type protein is a tetramer composed of four identical subunits [Blake, Geisow, Oatley, Rerat & Rerat (1978). J. Mol. Biol. 121, 339-356.] and a molecular-replacement solution for the Leu55Pro variant was obtained using one monomer of the wild-type protein as a model. Rigid-body refinement of the eight monomers in the asymmetric unit and subsequent refinement using molecular dynamics were performed with X-PLOR, leading to a current R factor of 20.3% for all the data. The crystallographic packing of the molecules is different from the one presented by the wild-type protein, opening new perspectives for understanding how this protein aggregates to form amyloid fibrils. PMID- 15299681 TI - New crystals of the histone core octamer diffract to higher resolution, 2.65 A. AB - A new crystal form of the histone octamer, crystallized in 1.6 M KCl, 1.6 M phosphate, diffracts to appreciably better than 2.6 A resolution. The crystals have space group P6(1) or P6(5) and lattice parameters a = b = 158.29, c = 103.27 A, alpha = beta = 90, gamma = 120 degrees, with one molecule per asymmetric unit. The new crystals promise to yield more detail of the histone basic domains and a higher resolution structure for the histone octamer. PMID- 15299682 TI - Crystallization and preliminary crystallographic analysis of endo-1,4-beta xyalanase I from Aspergillus niger. AB - A family G xylanase from Aspergillus niger has been crystallized using the vapor diffusion method. Several crystal forms could be obtained using various sodium salts as precipitants. Three of the crystal forms belong to space groups P21, P2(1)2(1)2(1) and P4(3) and have cell parameters of approximately a = b = 85.1, c = 113.6 A and alpha = beta = gamma = 90 degrees. These crystal forms can be converted into one another by flash freezing or macroseeding. A fourth crystal form is cubic (space group P2(1)3) with unit-cell axes of a = b = c = 112.3 A. Data sets for three of the four crystal forms have been collected, extending to a maximum resolution of 2.4 A. The structures of the monoclinic and orthorhombic crystals have been solved by molecular replacement by combining the crystallographic information of the different crystal forms. Refinement of the orthorhombic crystal form is now in progress. PMID- 15299683 TI - Crystallization and preliminary X-ray analysis of brazzein, a new sweet protein. AB - Brazzein is a sweet protein isolated from a wild African plant Pentadiplandra brazzeana. Brazzein is the smallest (molecular mass = 6473 Da) and the most water soluble protein sweetener discovered so far and is highly thermostable. Crystals were grown by vapor diffusion using sodium sulfate as a precipitant. They belong to the tetragonal space group I4(1)22 with unit-cell parameters a = b = 61.4, c = 59.6 A and with one molecule in the asymmetric unit. The crystals diffract to 1.8 A resolution using synchrotron radiation. PMID- 15299684 TI - Preliminary crystallographic analysis of the ParE subunit of Escherichia coli topoisomerase IV. AB - The ParE subunit of Escherichia coli topoisomerase IV has been crystallized in the presence of the non-hydrolyzable ATP analogue, 5'-adenylyl-beta,gamma imidodiphosphate (ADPNP). The crystals are of the orthorhombic space group, P2(1)2(1)2(1), with unit-cell dimensions a = 92.6, b = 119.1, c = 135.3 A. Data have been collected to 3.5 A resolution from frozen native crystals. Self rotation function analysis of these data indicate the position of a molecular twofold axis. Higher resolution native data are being collected and a derivative search is underway. PMID- 15299685 TI - Crystallization and preliminary X-ray analysis of pig pancreatic alpha-amylase in complex with a bean lectin-like inhibitor. AB - Pig pancreatic alpha-amylase (PPA, E.C. 3.2. 1. 17, 496 amino-acid residues) has been crystallized as a complex with a lectin-like inhibitor from bean Phaseolus vulgaris (224 amino-acid residues for the inhibitor monomer). The hanging-drop vapour-diffusion method was used to grow crystals from solutions containing 2 methyl-2,4-pentanediol as precipitant. The crystals belong to monoclinic space group C2 with a = 152.5, b = 80.3, c = 68.8 A, beta = 91.4 and diffract to 2.9 A resolution. A molecular-replacement solution of the structure has been obtained using the refined PPA and LoLl (Lathyrus ochrus isolectin I) atomic coordinates as starting models. Low-resolution refinement of the model is underway. The analysis reveals that the functional inhibitor molecule is dimeric and interacts with two molecules of enzyme. PMID- 15299686 TI - Crystallization and preliminary X-ray analysis of herpes simplex virus neutralizing antibody Fab fragment DL11. AB - An Fab fragment of a virus-neutralizing monoclonal antibody (DL11) that binds to herpes simplex virus glycoprotein D (HSV gD) has been purified, sequenced and crystallized. The biological activity of the purified Fab was verified by enzyme linked immunosorbant assay, flow cytometry and by neutralization of HSV infectivity. The crystals have the space group P1 with cell dimensions a = 40.2, b = 49.2, c = 63.9 A, alpha = 76.1, beta = 77.4, gamma = 71.6 degrees. The unit cell volume is consistent with it containing a single Fab molecule. The crystals grow to a maximum size of 0.7 x 0.3 x 0.3 mm and diffract X-rays to greater than 2.2 A resolution. The amino-acid sequences of the variable regions of the heavy and light chains of DL11 have been determined. These have been compared to those for other known Fab structures in the Protein Data Bank for selection of a starting model for crystallographic refinement by the molecular-replacement method. PMID- 15299687 TI - Crystallization and preliminary crystallographic analysis of E. coli uridine 5' diphospho-N-acetylenolpyruvylglucosamine reductase in two new crystal forms. AB - Uridine 5'-diphospho-N-acetylenolpyruvylglucosamine reductase (MurB), the second enzyme in the peptidoglycan synthetic pathway of Escherichia coli, has been crystallized in two previously unreported forms, one orthorhombic and the other monoclinic. MurB (molecular mass 38 kDa) crystallizes in a range of conditions that utilize polyethylene glycol fractions as precipitants, and crystals can be grown with or without the enzyme's substrate, uridine 5'-diphospho-N acetylenolpyruvylglucosamine. X-ray diffraction from crystals of the orthorhombic form extends to 2 A resolution and shows the symmetry and systematic absences of space group P2(1)2(1)2(1). These crystals show significant variations in cell dimensions at room temperature and at 100 K. A crystal used to collect a 2.0 A resolution data set at a synchrotron source showed cell dimensions at ca 100 K of a = 51.0, b = 79.3 and c = 87.1 A, indicating one molecule peroasymmetric unit. The monoclinic crystals scatter X-rays to 3.0 A resolution consistent with space group P2(1), unit-cell dimensions (ca 100 K) a = 50.7, b = 92.4, c = 85.5 A, and beta = 104 degrees, and two molecules per asymmetric unit. Mercury derivatives have been prepared with both orthorhombic and monoclinic forms, and efforts are underway to exploit these derivatives to determine the structure of this protein. PMID- 15299688 TI - Crystallization and preliminary X-ray analysis of a Y13S mutant of Spo0F from Bacillus subtilis. AB - Spo0F, a member of a superfamily of bacterial response regulatory proteins, is crucial to the regulation of sporulation in Bacillus subtilis. As there were difficulties in reproducing crystals of wild-type Spo0F, we report here the crystallization and preliminary studies of a mutant, Y13S protein, which gave well diffracting reproducible crystals. The crystals of the mutant obtained by the hanging-drop method belong to the tetragonal space group P4(1)2(1)2 (P4(3)2(1)2) a = b = 105.1, c = 85.9 A. Diffraction data were collected at 2.8 A at the laboratory source and subsequently 2.05. A data were collected upon flash freezing the crystal at the Stanford Synchrotron Radiation Laboratory. This mutant participates in the phosphorelay in a similar manner to the wild-type protein. The presence of divalent cations are essential for wild-type phosphorylation and the present mutant crystal form is obtained in the presence of calcium. PMID- 15299689 TI - Crystallization and preliminary X-ray diffraction studies of a hydroxynitrile lyase from Hevea brasiliensis. AB - Crystals of the hydroxynitrile lyase from Hevea brasiliensis overexpressed in Pichia pastoris have been obtained by the hanging-drop technique at 294 K with ammonium sulfate and PEG 400 as precipitants. The crystals belong to the orthorhombic space group C222(1) with cell dimensions of a = 47.6, b = 106.8 and c = 128.2 A. The crystals diffract to about 2.5 A resolution on a rotating-anode X-ray source. PMID- 15299690 TI - Preliminary X-ray diffraction studies of the tetragonal form of native horse spleen apoferritin. AB - Horse-spleen ferritin is known to crystallize in three different space groups, cubic F432, orthorhombic P2(1)2(1)2 and tetragonal P42(1)2, but only the cubic form has been fully investigated. Crystals of the tetragonal form of apoferritin have been obtained, by the vapour-diffusion technique, which diffract beyond 3.0 A. The unit-cell dimensions are a = b = 146.63, c = 152.94 A. The orientation of the non-crystallographic symmetry axes of the apoferritin molecule (24 subunits of 174 amino acids each, arranged in a 432 point symmetry rhombododecahedron) has been determined by a self-rotation Patterson function. The asymmetric unit is made of six subunits and was positioned by molecular replacement. PMID- 15299691 TI - Crystallization and preliminary X-ray analysis of an orthorhombic form of horse spleen apoferritin. AB - Horse-spleen apofemtin crystallizes in two different space groups: cubic F432 and tetragonal P42(1)2 while its iron-containing analogue is known to present a cubic and an orthorhombic form. Up to now, only the structure of the cubic form has been fully investigated by X-ray diffraction, although some information concerning the molecular packing of the two other forms was deduced from analysis of X-ray photographs. While growing cubic crystals of horse-spleen apoferritin with Pt-mesoporphyrin IX, we obtained one crystal, with a diffraction limit of 2.4 A, belonging to the orthorhombic P2(1)2(1)2 space group, with unit-cell dimensions a = 181.6, b = 128.9, c = 128.9 A. The orientation of the non crystallographic axes of the molecule was determined by self-rotation Patterson function and the structure was determined by the molecular-replacement method. The asymmetric unit consists of half an apoferritin molecule. Refinement of the structure is in progress, some preliminary results of the molecular packing are given. PMID- 15299692 TI - Crystallization and preliminary X-ray analysis of chicken-liver glutathione S transferase CL 3-3. AB - Five different crystal forms of recombinant chicken-liver glutathione S transferase CL 3-3 have been obtained by the vapor-diffusion method. The form A crystals are monoclinic C2, a = 125.56, b = 85.81, c = 52.71 A and beta = 114.64 degrees, and diffract to 4 A resolution. The form B crystals are monoclinic P2(1), a = 105.13, b = 118.54, c = 62.62 A and beta = 124.74 degrees, and diffract to 2.8 A resolution. The form C crystals are orthorhombic C222(l), a = 101.69, b = 115.46, c = 95.40 A, and diffract to 2.8 A resolution. The form D crystals are tetragonal, P4(1)2(1)2 or P4(3)2(1)2, a = b = 115.31, c = 171.20 A and diffract to 3.5 A resolution. The form E crystals are hexagonal, P6(1) or P6(5), a = b = 104.23, c = 114.35 A, diffract to 3.5 A resolution. Forms A, C and E have one dimer of molecular weight 50 kDa, while forms B and D have two dimers per asymmetric unit, respectively. PMID- 15299693 TI - Crystallization and preliminary crystallographic study of asparagine synthetase from Escherichia coli. AB - Crystals of Escherichia coli asparagine synthetase have been obtained from 45% saturated ammonium sulfate solution at pH 7.5 by using its Cys-free (Cys51-->Ala, Cys315-->Ala) mutant. The crystals belong to space group P2(1), with cell dimensions a = 52.90(4), b = 126.2 (2), c = 52.8 (1) A and beta = 105.3(1) degrees. The self-rotation function specifies that there is one dimer in the asymmetric unit and the subunits are related by a non-crystallographic twofold axis. Complete data sets up to 2.7 A resolution have been collected on an R-AXIS IIc imaging-plate system. PMID- 15299694 TI - Compressibility of lysozyme protein crystals by X-ray diffraction. AB - Single-crystal high-pressure X-ray diffraction studies on the protein crystals of orthorhombic and tetragonal hen egg-white lysozyme polymorphs were carried out using a Merrill-Bassett diamond-anvil cell, image-plate detector and synchrotron radiation. The orthorhombic crystal has been squeezed to 85.5% of its ambient pressure volume at about 1.0 GPa; the crystal compresses anisotropically, and neither a glass transition or denaturation was observed. The tetragonal polymorph of lysozyme undergoes amorphization at pressures about 0.2 GPa. PMID- 15299695 TI - Deposition of macromolecular data. PMID- 15299696 TI - A genetic algorithm for the ab initio phasing of icosahedral viruses. AB - Genetic algorithms have been investigated as computational tools for the de novo phasing of low-resolution X-ray diffraction data from crystals of icosahedral viruses. Without advance knowledge of the shape of the virus and only approximate knowledge of its size, the virus can be modeled as the symmetry expansion of a short list of nearly tetrahedrally arranged lattice points which coarsely, but uniformly, sample the icosahedrally unique volume. The number of lattice points depends on an estimate of the non-redundant information content at the working resolution limit. This parameterization permits a simple matrix formulation of the model evaluation calculation, resulting in a highly efficient survey of the space of possible models. Initially, one bit per parameter is sufficient, since the assignment of ones and zeros to the lattice points yields a physically reasonable low-resolution image of the virus. The best candidate solutions identified by the survey are refined to relax the constraints imposed by the coarseness of the modeling, and then trials whose intensity-based statistics are comparatively good in all resolution ranges are chosen. This yields an acceptable starting point for symmetry-based direct phase extension about half the time. Improving efficiency by incorporating the selection criterion directly into the genetic algorithm's fitness function is discussed. PMID- 15299697 TI - Direct-space methods in phase extension and phase refinement. IV. The double histogram method. AB - In the conventional histogram-matching technique for phase extension and refinement for proteins a simple one-to-one transformation is made in the protein region to modify calculated density so that it will have some target histogram in addition to solvent flattening. This work describes an investigation where the density modification takes into account not only the current calculated density at a grid point but also some characteristic of the environment of the grid point within some distance R. This characteristic can be one of the local maximum density, the local minimum density or the local variance of density. The grid points are divided into ten groups, each containing the same number of grid points, for ten different ranges of value of the local characteristic. The ten groups are modified to give different histograms, each corresponding to that obtained under the same circumstances from a structure similar to the one under investigation. This process is referred to as the double-histogram matching method. Other processes which have been investigated are the weighting of structure factors when calculating maps with estimated phases and also the use of a factor to dampen the change of density and so control the refinement process. Two protein structures were used in numerical trials, RNApl [Bezborodova, Ermekbaeva, Shlyapnikov, Polyakov & Bezborodov (1988). Biokhimiya, 53, 965-973] and 2-Zn insulin [Baker, Blundell, Cutfield, Cutfield, Dodson, Dodson, Hodgkin, Hubbard, lsaacs, Reynolds, Sakabe, Sakabe & Vijayan (1988). Philos. Trans. R. Soc. London Ser. B, 319, 456--469]. Comparison of the proposed procedures with the normal histogram-matching technique without structure-factor weighting or damping gives mean phase errors reduced by up to 10 degrees with map correlation coefficients improved by as much as 0.14. Compared to the normal histogram used with weighting of structure factors and damping, the improvement due to the use of the double-histogram method is usually of order 4 degrees in mean phase error and an increase of 0.06-0.08 in the map correlation coefficient. It is concluded that the most reliable results are found with the local-maximum condition and with R in the range 0.5-0.6 A. PMID- 15299698 TI - Statistical expectation value of the Debye-Waller factor and E(hkl) values for macromolecular crystals. AB - If the unit-cell distribution of atomic mean-square displacement parameters B = 8pi(2) is assumed to be normal, with mean micro = and variance sigma(2) = <(B-)(2)>, the statistical expectation value of the Debye-Waller factor W(2) = exp(-2Bs(2)), where s = (sin theta)/lambda, is = exp[-2( micro - sigma(2)s(2))s(2)]. This result has been incorporated into procedures for scaling and normalizing measured Bragg intensities to their Wilson expectation values. The procedures can determine both isotropic micro (B) and sigma(B) and anisotropic micro (U(ij)) and sigma(U(ij) distribution parameters. Tests with experimental data and refined structural models for several protein crystals show that the procedures yield reliable normalized structure-factor amplitudes for direct-methods applications, with values of R = summation operator (h)||E(o)| - |E(c)||/ summation operator (h)|E(o)| averaging approximately 5%. PMID- 15299699 TI - Experiments in microgravity: a comparison of crystals of a carbohydrate-binding fab grown on the ground, on space shuttle Discovery and on space station Mir. AB - The Fab fragment of the hybridoma antibody (YsT9.1) specific to Brucella abortus has been crystallized on earth using both Linbro plates and ground-based models of the flight hardware, as well as in microgravity on board the space shuttle Discovery and the space station Mir. Large-scale experiments using Linbro plates gave two different crystal morphologies, pyramidal and rhomboid, depending on conditions. The pyramidal crystals proved to scatter X-rays to higher resolution, and conditions within the ground-based flight hardware for both Discovery and Mir were adjusted to produce crystals with this morphology. The experiment on Discovery produced large crystals in each of ten chambers. The experiment on Mir produced crystals in only one of the five assigned chambers, despite the fact that the simultaneous ground-based experiment produced large crystals in every corresponding chamber. Data collection was attempted for crystals from both space and ground-based experiments. Higher resolution data was obtained from crystals grown on Discovery than from either Mir or ground-based crystals, even though the crystals obtained from Discovery were smaller and forced to grow over a much shorter period of time because of the shorter length of the shuttle mission. PMID- 15299700 TI - An ambiguous structure of a DNA 15-mer thrombin complex. AB - The structure of a complex between thrombin and a GGTTGGTGTGGTTGG DNA 15-mer has been analyzed crystallographically. The solution NMR structure of the 15-mer has two stacked G-quartets similar to that found in the previous X-ray structure determination of the 15-mer-thrombin complex [Padmanabhan, Padmanabhan, Ferrara, Sadler & Tulinsky (1993). J. Biol. Chem. 268, 17651-17654]; the strand polarity, however, is reversed from that of the crystallographic structure. The structure of the complex here has been redetermined with better diffraction data confirming the previous crystallographic structure but also indicating that the NMR solution structure fits equally well. Both 15-mer complex structures refined to an R value of about 0.16 presenting a disconcerting ambiguity. Since the two 15-mer structures associate with thrombin in different ways (through the TGT loop in the X-ray and TT loop in the NMR model), other independent lines of physical or chemical evidence are required to resolve the ambiguity. PMID- 15299701 TI - X-ray diffraction analysis of crystals containing twofold symmetric nucleosome core particles. AB - Nucleosome core particles containing a DNA palindrome and purified chicken erythrocyte histone octamer have been reconstituted and crystallized. The dyad symmetry of the palindrome extends the dyad symmetry of the histone octamer to result in a twofold symmetric particle. This ensures that the structure determined by X-ray diffraction will yield a true representation of the DNA strand rather than the twofold averaged structure which would result from using a non-palindromic DNA sequence. The crystals provide isotropic diffraction to 3.2 A with observed reflections extending to d spacings of about 2.8 A using a rotating anode Cu Kalpha X-ray source. Although the DNA palindrome is a factor contributing to the quality of the diffraction data, another significant factor is an improved preparative technique which enriches for correctly phased nucleosome core particles. PMID- 15299702 TI - Refined structure of the chitinase from barley seeds at 2.0 a resolution. AB - Chitinase from barley seeds is a monomeric enzyme with 243 amino-acid residues and it plays a role as a defense protein. Its structure, previously determined at 2.8 A resolution by multiple isomorphous replacement method, is mainly alpha helical [Hart, Monzingo, Ready, Ernst & Robertus, (1993). J. Mol. Biol. 229, 189 193]. The crystallization and preliminary X-ray data of the same enzyme in a different crystal form has been reported independently [Song, Hwang, Kim & Suh, (1993). Proteins, 17, 107-109}, the asymmetric unit of which contains two chitinase molecules. As a step toward understanding the general principles of catalysis, reported here is the structure of chitinase from barley seeds in this crystal form, as determined by molecular replacement and subsequently refined at 2.0 A resolution, with incorporation of partial data to 1.9 A (R factor of 18.9% for 31 038 unique reflections with F(o)> 2sigma(F) in the range 8.0-1.9 A). The r.m.s. deviations from ideal stereochemistry are 0.013 A for bond lengths and 1.32 degrees for bond angles. A superposition of the two independent molecules in the asymmetric unit gives an r.m.s. difference of 0.55 A for all protein atoms (0.43 and 0.74 A for main-chain and side-chain atoms, respectively). When the refined model of each chitinase molecule in the asymmetric unit is superposed with the starting model, the r.m.s. difference for all shared protein atoms is 0.99 A for molecule 1 and 0.85 A for molecule 2, respectively. Through a sequence comparison with homologous plant chitinases as well as a structural comparison with the active sites of other glycosidases, key catalytic residues have been identified and the active site has been located in the three-dimensional structure of the barley chitinase. The present structure, refined at an effective resolution of 2.0 A with incorporation of partial data to 1.9 A, represents a significant improvement in resolution compared to the previously reported model. The improved resolution has enabled the location of solvent atoms, including water molecules near the catalytic residues, in addition to the positioning of protein atoms with greater accuracy. PMID- 15299703 TI - DNA-drug refinement: a comparison of the programs NUCLSQ, PROLSQ, SHELXL93 and X PLOR, using the low-temperature d(TGATCA)-nogalamycin structure. AB - In an earlier study [Smith, Davies, Dodson & Moore (1995). Biochemistry, 34, 415 425] the crystal structure of the d(TGATCA)-nogalamycin complex was determined to 1.8 A and refined with PROLSQ to R = 19.5% against 4767 reflections with F> 1sigma(F). A low-temperature crystallographic study on this complex has now been performed. Native data collection at liquid-nitrogen temperature (120 K) improved the resolution to 1.4 A. The structure has now been refined against these new diffraction data in the resolution range 8-1.4 A using NUCLSQ, PROLSQ, SHELXL93 and X-PLOR, in order to determine to what extent the resulting DNA conformation and associated solvent structure would differ and to examine the suitability of these programs for the refinement of oligonucleotide structures. With the advent of more DNA-protein structure determinations, it is of interest to see how well the protein-refinement packages, PROLSQ and X-PLOR, and the small-molecule program, SHELXL93, are able to accommodate DNA. Comparisons are made between the dictionaries, weights and restraints used and the final models obtained from each program. Although the final R values, using all data in the resolution range 8.0 1.4 A, from PROLSQ (22.8%), SHELXL93 (R1 =21.7% after isotropic refinement) and X PLOR (24.4%) are higher than the R value from the NUCLSQ refinement (21.2%), the root-mean-square deviations between the four final models are very small. Using this high-quality 8.0-1.4 A data set neither the dictionary nor the refinement program leave an imprint on the final fully refined complex. Likewise, the helical parameters and backbone conformation including sugar-puckering modes are not influenced by the refinement procedure used. Although a different number of water molecules is found in each refinement, varying from 62 (X-PLOR) to 86 (NUCLSQ), the first hydration sphere is well conserved in all four models. PMID- 15299704 TI - Refined structure of the monoclonal antibody HyHEL-5 with its antigen hen egg white lysozyme. AB - The crystal structure of the complex of the antibody Fab, HyHEL-5, with its antigen, hen egg-white lysozyme, has been refined at 2.65 A resolution to an R value of 0.196. The resulting model has significantly better stereochemistry than the previously reported model of the complex, PDB reference 2HFL, and sufficiently improved phases, permitting the reliable location of a number of water molecules. No major conformational differences are observed between this structure and that previously reported, although small differences occur throughout the complex. 82 water molecules have been assigned, of which three are in the antibody-antigen interface involved in a hydrogen-bonding network. Three other waters are trapped within the interface between V(H) and V(L) and a fourth water molecule is observed near the interface but buried below the lysozyme surface as observed in crystal structures of lysozyme alone. PMID- 15299705 TI - Ribonuclease from Streptomyces aureofaciens at atomic resolution. AB - Crystals of ribonuclease from Streptomyces aureofaciens diffract to atomic resolution at room temperature. Using synchrotron radiation and an imaging-plate scanner, X-ray data have been recorded to 1.20 A resolution from a crystal of native enzyme and to 1.15 A from a crystal of a complex with guanosine-2' monophosphate. Refinement with anisotropic atomic temperature factors resulted in increased accuracy of the structure. The R factors for the two structures are 10.6 and 10.9%. The estimated r.m.s. error in the coordinates is 0.05 A, less than half that obtained in the previous analysis at 1.7 A resolution. For the well ordered part of the main chain the error falls to below 0.02 A as estimated from inversion of the least-squares matrix. The two independent molecules in the asymmetric unit allowed detailed analysis of peptide planarity and some torsion angles. The high accuracy of the analysis revealed density for a partially occupied anion in the nucleotide binding site of molecule A in the native structure which was not seen at lower resolution. The anisotropic model allowed correction of the identity of the residue at position 72 from cysteine to threonine. Cys72 SG had been modelled in previous analyses with two conformations. The solvent structure was modelled by means of an automated procedure employing a set of objective criteria. The solvent structure for models refined using different programs with isotropic and anisotropic description of thermal motion is compared. PMID- 15299706 TI - How to escape from model bias with a high-resolution native data set - structure determination of the PcpA-S6 subunit III. AB - The structure of procarboxypeptidase A-S6 subunit III, a truncated zymogen E, has been determined by molecular replacement using as search model porcine elastase 1 which, as revealed by crystallographic analysis, contained about 20% of the amino acids in a radically different orientation. Two monoclinic crystal forms were used: the first one diffracts to 2.3 A resolution and contains one molecule per asymmetric unit; the second diffracts to 1.7 A resolution and contains two molecules per asymmetric unit. Molecular replacement and conventional X-PLOR refinement led to a model for which 20% of the chain was ill defined in both crystal forms. To remove the bias introduced by the initial model, an automated refinement procedure [Lamzin & Wilson (1993). Acta Cryst. D49, 129-147] was applied successfully to the second crystal form, which diffracts to high resolution. The resulting dramatic improvement of the electron-density map led to extensive rebuilding of some surface loops. The reliability of the modified model was confirmed by refinement of the first crystal form. For the two forms, the final R factor is 18.8% for data between 8.0 and 2.0 A resolution, and 18.4% for data between 8.0 and 1.7 A, respectively. PMID- 15299707 TI - Three-dimensional structure of cytochrome c' from two Alcaligenes species and the implications for four-helix bundle structures. AB - The three-dimensional structures of two cytochromes c' have been determined in order to analyse the common features of proteins of this family and their relationship with other four-helix bundle structures. The structure of cytochrome c' from Alcaligenes sp was determined by molecular replacement supplemented with the iron anomalous scattering and the use of a single isomorphous heavy-atom derivative, and was refined using synchrotron data to 1.8 A resolution. The final model, comprising 956 protein atoms (one monomer) and 89 water molecules, has a final R value of 0.188 for all data in the range 20.0-1.8 A resolution (14 673 reflections). The structure of the cytochrome c' from Alcaligenes denitrificans is isomorphous and essentially identical (r.m.s. deviation for all atoms 0.36 A). Although its amino-acid sequence has not been determined chemically, only four differences from that of Alcaligenes sp cytochrome c' were identified by the X ray analysis. The final model for Alcaligenes denitrificans cytochrome c', comprising 953 protein atoms and 75 water molecules, gave a final R factor of 0.167 for all data in the range 20.0-2.15 A (8220 reflections). The cytochrome c' monomer forms a classic four-helix bundle, determined by the packing of hydrophobic side chains around the enclosed haem group. There are very few cross linking hydrogen bonds between the helices, the principal side-chain hydrogen bonding involving one of the haem propionates and a conserved Arg residue. The cytochrome c' dimer is created by a crystallographic twofold axis. Monomer monomer contacts primarily involve the two A helices, with size complementarity of side chains in a central solvent-excluded portion of the interface and hydrogen bonding at the periphery. Both species have a pyroglutamic acid N terminal residue. The haem iron is five-coordinate, 0.32 A out of the haem plane towards the fifth ligand, His120. The unusual magnetic properties of the Fe atom may be linked to a conserved basic residue, Arg124, adjacent to His120. PMID- 15299708 TI - Structure of a functional fragment of VCAM-1 refined at 1.9 a resolution. AB - The crystal structure of the functional amino-terminal two-domain fragment of human vascular cell adhesion molecule 1 (VCAM-1) has been determined at 1.9 A resolution. The crystals contain two copies of the molecule in the asymmetric unit. The structure was solved by multiple isomorphous replacement, using lead and selenium derivatives. Anomalous scattering had to be used to resolve the phase ambiguity of a lead derivative. Since the selenium derivative has very small isomorphous differences, the local scaling algorithm had to be used to obtain an interpretable difference Patterson map. The initial phases were improved by non-crystallographic averaging, solvent flattening and histogram matching. The structure has been refined to a crystallographic R factor of 20.4% (15-1.9 A, F>/= 3sigma) and consists of two Ig domains (D1 and D2). The angle between these domains differs by 12 degrees between the two copies of the molecule in the crystallographic asymmetric unit, demonstrating that some movement is possible at the interface. In the amino-terminal domain D1 there is an 'extra' disulfide bond, in addition to the conserved cross-sheet disulfide bond, at the top of the molecule. This bond, a hallmark of the integrin-binding subclass of Ig superfamily proteins, makes the top of this domain very compact. The feature that projects most prominently from D1 is the CD loop, near the base of the domain. The key residue for integrin binding, Asp40, is located in this loop and is easily accessible. PMID- 15299709 TI - 1.8 A structure of Hypoderma lineatum collagenase: a member of the serine proteinase family. AB - Collagenase from the fly larvae Hypoderma lineatum cleaves triple-helical collagen in a single region. It was crystallized at neutral pH in the absence of inhibitor and 1.8 A data were collected using synchrotron radiation and a Mark II prototype detector. The structure was solved by combining multiple isomorphous replacement methods and rotation translation function in real space. Refinement between 7 and 1.8 A using the program X-PLOR led to a final R factor of 16.9%. The overall fold is similar to that of other trypsin-like enzymes but the structure differs mainly by the presence of a beta-sheet at position 31-44. The two embedded molecules of the asymmetric unit are related by a pseudo twofold axis. The beta-sheet 31-44 of one molecule is involved in hydrogen bonds with binding-pocket residues of the other molecule. It thus completely prevents access to the active site. The specificity of this enzyme probably results from the position of Phe192 and Tyr99 at the entrance of the active site. PMID- 15299710 TI - Refined crystal structure of the catalytic domain of xylanase A from Pseudomonas fluorescens at 1.8 A resolution. AB - The three-dimensional structure of native xylanase A from Pseudomonas flouorescens subspecies cellulosa has been refined at 1.8 A resolution. The space group is P2(1)2(1)2(1) with four molecules in the asymmetric unit. The final model has an R factor of 0.166 for 103 749 reflections with the four molecules refined independently. The tertiary structure consists of an eightfold beta/alpha barrel, the so-called TIM-barrel fold. The active site is in an open cleft at the carboxy-terminal end of the beta/alpha-barrel, and the active-site residues are a pair of glutamates, Glu127 on strand 4 and Glu246 on strand 7. Both these catalytic glutamate residues are found on beta-bulges. An atypically long loop after strand 7 is stabilized by calcium. Unusual features include a non-proline cis-peptide residue Ala80 which is found on a beta-bulge at the end of beta strand 3. The three beta-bulge type distortions occurring on beta-strands 3, 4 and 7 are functionally significant as they serve to orient important active-site residues. The active-site residues are further held in place by an extensive hydrogen-bonding network of active-site residues in the catalytic site of xylanase A. A chain of well ordered water molecules occupies the substrate binding cleft, some or all of which are expelled on binding of the substrate. PMID- 15299711 TI - Crystallization and preliminary X-ray analysis of pectin lyase A from Aspergillus niger. AB - The major secreted pectin lyase (E.C. 4.2.2.10) from Aspergillus niger, strain 4M 147, has been purified and crystallized by the hanging-drop method using polyethylene glycol as precipitant. The crystals belong to the space group P2(1)2(1)2(1) with cell dimensions a = 45.2, b = 83.2, c = 93.1 A (1 A = 0.1 nm) and a single molecule in the asymmetric unit. The crystals diffract to at least 2.0 A resolution and are suitable for structure determination. PMID- 15299712 TI - Crystallization and preliminary X-ray diffraction study of aldehyde reductase from a red yeast, Sporobolomyces salmonicolor. AB - Crystals of aldehyde reductase from a red yeast, Sporobolomyces salmonicolor, have been grown from an ammonium sulfate solution, pH 7.0, by means of the vapor diffusion procedure. The crystals belong to the hexagonal system, space group P6(1)22 or its enantiomorph, P6(5)22, with unit-cell dimensions of a = 72.2 and c = 320 A. The X-ray diffraction patterns extend to at least 2 A resolution with the use of synchrotron radiation. The crystals are stable on exposure to X-rays and suitable for high-resolution X-ray structure determination. PMID- 15299713 TI - Purification, crystallization and preliminary X-ray diffraction analysis of haemorrhagin IV from the snake venom of Agkistrodon acutus. AB - Haemorrhagin IV, a medium molecular weight haemorrhagin from the snake venom of Agkistrodon acutus (AaHIV), has been purified and crystallized. The molecular weight and isoelectric point of AaHIV are 44 kDa and pI 5.0, respectively. The crystal belongs to space group C222(1) with unit-cell dimensions of a = 124.2, b = 114.5, c = 98.4 A, and could o diffract X-rays to 3.0 A, resolution. There are one or two molecules in the crystallographic asymmetric unit. PMID- 15299714 TI - Crystallization and initial X-ray analysis of beta-crustacyanin, the dimer of apoproteins A2 and C1, each with a bound astaxanthin molecule. AB - Crystals of beta-crustacyanin, a carotenoid-binding protein from lobster carapace, have been grown under oil from solutions containing sodium potassium phosphate as precipitant. They grow slowly over a period of months to reach maximal dimensions of 0.5 x 0.1 x 0.1 mm, and belong to space group P622 with cell dimensions: a = b = 124.39, c = 188.86 A and gamma = 120 degrees. The crystals diffract to beyond 3 A but are very radiation sensitive, limiting the resolution of usable data. The unit-cell volume suggests that there are two beta crustacyanin molecules per asymmetric unit. PMID- 15299715 TI - Crystallization and preliminary X-ray analysis of two new crystal forms of calmodulin. AB - Two new crystal forms of calmodulin from Gallus gallus are reported. Crystals in space group P1 (cell dimensions a = 59.7, b = 53.1, c = 24.6 A, alpha = 93.2, beta = 96.7, gamma = 89.2 and Z = 2), grow as long thin needles. Water content on density considerations is approximately 50%. They diffract to approximately 2.0 A but give wide multiply peaked spot profiles. Crystals in space group P2(1)2(1)2(1) (cell dimensions a = 32.2, b = 56.0, c = 67.3 A and Z = 4), grow as clusters of thin tablets and contain approximately 30% water by volume. These small crystals ( approximately 0.4 x 0.15 x 0.1 mm) diffracted well to approximately 1.4 A and some appreciable intensities were observed at resolutions better than 1.2 A. PMID- 15299716 TI - Preliminary crystallographic studies of a protein from Pachyrrhizus erosus. AB - Crystals of a protein extracted from the seeds of Pachyrrhizus erosus have been obtained by vapor-phase diffusion. The crystal belongs to the space group P4(1)2(1)2 or P4(3)2(1)2 with cell parameters a = b = 62.52, c = 147.42 A. There is one protein molecule of 33 kDa in an asymmetric unit. A data set at 3.1 A has been collected on an area detector. PMID- 15299717 TI - Crystallization and preliminary X-ray analysis of the primary receptor (PotD) of the polyamine transport system in Escherichia coli. AB - The primary receptor (potD, M(r) = 39 000) of the polyamine transport system in Escherichia coli has been crystallized by the vapor-diffusion method. Two crystal forms were obtained in the presence of spermidine, and were examined by X-ray analysis. Form I crystals, which diffract to 2.5 A resolution, belong to the space group P2(1), with unit-cell dimensions a = 145.3, b = 69.1, c = 72.5 A and beta = 107.6 degrees. Four molecules are contained in an asymmetric unit. These form two dimers that are related to each other by a local translation of about half of the unit cell along the a axis. The two protein molecules in each dimer are similarly related by a local dyad. Form II crystals diffract to 1.8 A resolution and belong to the space group I4(1), with unit-cell dimensions a = b = 130.3 and c = 38.7 A. They contain one molecule per asymmetric unit. PMID- 15299718 TI - Crystallization and preliminary X-ray diffraction studies of 5-chlorolevulinate modified bovine porphobilinogen synthase and the Pb(II)-complexed enzyme. AB - Bovine porphobilinogen synthase (PBGS) is an homo-octameric enzyme with four active sites. Each active site binds two Zn(II) atoms whose ligands differ and two molecules of 5-aminolevulinate whose chemical fates differ. The asymmetric binding of two Zn(II) atoms and two identical substrate molecules by a homodimeric active site is apparently unique. Modification by 5-chiorolevulinate can be used to differentiate the two substrate-binding sites; diffraction-quality crystals of 5-chlorolevulinate-modified PBGS have been obtained. Pb(II) can be used to differentiate the two different Zn(II)-binding sites; diffraction-quality crystals of the Pb(II) complex of PBGS have been obtained. Preliminary diffraction data reveal an I422 space group, in agreement with a general model for the quaternary structure of PBGS. PMID- 15299721 TI - Structure determination and refinement of the Humicola insolens endoglucanase V at 1.5 A resolution. AB - The structure of the catalytic core of the endoglucanase V (EGV) from Humicola insolens has been determined by the method of multiple isomorphous replacement at 1.5 A resolution. The final model, refined with X-PLOR and PROLSQ, has a crystallographic R factor of 0.163 (R(free) = 0.240) with deviations from stereochemical target values of 0.012 A and 0.037 degrees for bonds and angles, respectively. The model was further refined with SHELXL, including anisotropic modelling of the protein-atom temperature factors, to give a final model with an R factor of 0.105 and an R(free) of 0.154. The initial isomorphous replacement electron-density map was poor and uninterpretable but was improved by the use of synchrotron data collected at a wavelength chosen so as to optimize the f" contribution of the anomalous scattering from the heavy atoms. The structure of H. insolens EGV consists of a six-stranded beta-barrel domain, similar to that found in a family of plant defence proteins, linked by a number of disulfide bonded loop regions. A long open groove runs across the surface of the enzyme either side of which lie the catalytic aspartate residues. The 9 A separation of the catalytic carboxylate groups is consistent with the observation that EGV catalyzes the hydrolysis of the cellulose, beta(1-->4) links with inversion of configuration at the anomeric C1 atom. This structure is the first representative from the glycosyl hydrolase family 45. PMID- 15299722 TI - Structure of bovine pancreatic trypsin inhibitor at 125 K definition of carboxyl terminal residues Gly57 and Ala58. AB - The structure of bovine pancreatic trypsin inhibitor has been refined to a resolution of 1.1 A against data collected at 125 K. The space group of the form II crystal is P2(1)2(1)2(1) with a = 75.39(3), b = 22.581(7), c = 28.606 (9) A (cf. a = 74.1, b = 23.4, c = 28.9 A at room temperature). The structure was refined by restrained least-squares minimization of summation operator w(F (o)(2) F (c)(2))(2) with the SHELXL93 program. As the model improved, water molecules were included and exceptionally clear electron density was found for two residues, Gly57 and Ala58, that had been largely obscured at room temperature. The side chains of residues Glu7 and Arg53 were modelled over two positions with refined occupancy factors. The final model contains 145.6 water molecules distributed over 167 sites, and a single phosphate group disordered over two sites. The root-mean-square discrepancy between Calpha atoms in residues Arg1 Gly56 at room and low temperatures is 0.4 A. A comparison of models refined with anisotropic and isotropic thermal parameters revealed that there were no significant differences in atomic positions. The final weighted R-factor on F(2) (wR(2)) for data in the range 10-1.1 A was 35.9% for the anisotropic model and 40.9% for the isotropic model. Conventional R-factors based on F for F > 4sigma(F) were 12.2 and 14.6%, respectively, corresponding to 16.1 and 18.7% on all data. These large R-factor differences were not reflected in values of R(free), which were not significantly different at 21.5(5) and 21.8(4)%, respectively. These results, along with the relatively straightforward nature of the refinement, clearly highlight the benefits of low-temperature data collection. PMID- 15299723 TI - Methods used in the structure determination of bovine mitochondrial F1 ATPase. AB - With a size of 372 kDa, the F(1) ATPase particle is the largest asymmetric structure solved to date. Isomorphous differences arising from reacting the crystals with methyl-mercury nitrate at two concentrations allowed the structure determination. Careful data collection and data processing were essential in this process as well as a new form of electron-density modification, 'solvent flipping'. The most important feature of this new procedure is that the electron density in the solvent region is inverted rather than set to a constant value, as in conventional solvent flattening. All non-standard techniques and variations on new techniques which were employed in the structure determination are described. PMID- 15299724 TI - Phase combination and cross validation in iterated density-modification calculations. AB - A variety of density-modification techniques are now available for improving electron-density maps in accordance with known chemical information. This modification must, however, always be constrained by consistency with the experimental data. This is conventionally achieved by alternating cycles of map modification in real space with recombination with the experimental data in reciprocal space. The phase recombination is based upon the assumption that the density-modified map may be treated as a partial model of the structure which contains information independent of the experimentally derived phases. This assumption is shown to be incorrect, and an alternative procedure is investigated which as a side effect allows calculation of a free R factor. PMID- 15299725 TI - Representing stereochemical information in macromolecular electron-density distributions by multi-dimensional histograms. AB - Previous studies have demonstrated the value of ideal electron-density histograms as targets for the corresponding histograms of experimental electron-density maps. The electron-density histogram makes use of density values as independent objects, and no relationship between them is taken into account. Extension to include the relationships between neighboring density values leads naturally to a multi-dimensional histogram defined as the joint frequency of the density values and their higher order derivatives. We show here that the multi-dimensional histogram including additional dimensions composed of the gradient magnitude and Laplacian of the density is minimally dependent on molecular folding and packing, and captures substantially more stereochemical information than the conventional electron-density histogram. The gradient histogram appears to be much more sensitive to phase errors than the conventional electron-density histogram. Potential uses of the multi-dimensional histogram include improved targets for density modification and more reliable figures of merit for evaluating correct phases. PMID- 15299726 TI - New parameters for the refinement of nucleic acid-containing structures. AB - Structures at atomic resolution (up to 1.0 A) which contain bases, sugars or the phosphodiester linkage, were selected from the Nucleic Acid Database or the Cambridge Structural Database to build a nucleic acid dictionary from X-ray refined structures. The dictionary consists of the average values for bond distances, bond angles and dihedral angles. The variance of the sample is used to provide information about the expected r.m.s. deviations of the refined parameters. A dictionary was constructed for refinement trials in X-PLOR. The dictionary includes RNA and DNA in C2'-endo and C3'-endo sugar pucker conformations, as well as values for the backbone dihedrals. Tests were performed on the dictionary using three structures: a B-DNA, a Z-DNA and a protein-DNA complex. During the course of refinement, all three structures showed significant improvements as measured by r.m.s. deviations and R factors when compared to the previous DNA dictionary. PMID- 15299727 TI - Refinement and structural analysis of bovine cytochrome b5 at 1.5 A resolution. AB - The structure of bovine liver cytochrome b(5), a soluble 93-residue proteolytic fragment of a 16 kDa membrane-bound hemoprotein, initially solved at 2.0 A resolution, has been refined at 1.5 A using data collected on a diffractometer. Refinement to 2.0 A resolution used the Hendrickson-Konnert procedure PROLSQ and was then extended to 1.5 A resolution using the program PROFFT. Only residues 3 87 could be identified in the model and these residues together with 93 water molecules gave an agreement factor of R = 0.161 for data in the resolution range 1.5-5 A. The structure was finally refined using the program X-PLOR, which enabled alternate conformers to be modelled for several surface side chains. Residues 1 and 2 at the amino terminus of the protein and residue 88 near the carboxyl terminus could be identified from these electron-density maps. However the remaining disordered carboxy-terminal residues could not successfully be included in the model. A total of 117 solvent molecules were included in the final refinement to give R = 0.164 for the data between 1.5 and 10 A. PMID- 15299728 TI - Structure of flavoprotein FP390 from a luminescent bacterium Photobacterium phosphoreum refined at 2.7 A resolution. AB - The three-dimensional structure of a flavoprotein, FP(390), from a luminescent bacterium, Photobacterium phosphoreum, solved by the molecular-replacement method, was refined to an R factor of 24.0% for 17 433 independent reflections, from 6.0 to 2.7 A resolution, collected by synchrotron radiation. The asymmetric unit of the crystal (space group P4(3)22, a = b = 76.8 and c = 242 A) contains two monomer molecules related by a non-crystallographic twofold axis to form a dimer. There are two Q-flavin [flavin mononucleotide (FMN) with myristic acid] molecules in FP(390) monomer. One of them is located at the interface of dimer which is bound to both monomer and the another is at the molecular surface. The electron density of myristic acids of Q-flavins at the dimer interface in both monomer are weak and unclear, showing the possibility that the Q-flavins bound in this site are not a single species but a mixture of two components, 6-(3" myristic acid)-FMN and 6-(4"- myristic acid)-FMN. PMID- 15299729 TI - Structure of fasciculin 2 from green mamba snake venom: evidence for unusual loop flexibility. AB - The crystal structure of the snake toxin fasciculin 2, a potent acetylcholinesterase inhibitor from the venom of the green mamba (Dendroaspis angusticeps), has been determined by the molecular-replacement method, using the fasciculin 1 model and refined to 2.0 A resolution. The introduction of an overall anisotropic temperature factor improved significantly the quality of the electron-density map. It suggests, as it was also indicated by the packing, that the thermal motion along the unique axis direction is less pronounced than on the (ab) plane. The final crystallographic R factor is 0.188 for a model having r.m.s. deviations from ideality of 0.016 A for bond lengths and 2.01 degrees for bond angles. As fasciculin 1, fasciculin 2 belongs to the three-finger class of Elapidae toxins, a structural group that also contains the alpha-neurotoxins and the cardiotoxins. Although the two fasciculins have, overall, closely related structures, the conformation of loop I differs appreciably in the two molecules. The presence of detergent in crystallization medium in the case of fasciculin 2 appears to be responsible for the displacement of the loop containing Thr9. This conformational change also results in the formation of a crystallographic dimer that displays extensive intermolecular interactions. PMID- 15299730 TI - X-ray structure of the signal transduction protein from Escherichia coli at 1.9 A. AB - The structure of the bacterial signal transduction protein P(II) has been refined to an R factor of 13.2% using 3sigma data between 10 and 1.9 A. The crystals exhibited twinning by merohedry and X-ray intensities were corrected using the method of Fisher & Sweet [Fisher & Sweet (1980). Acta Cryst. A36, 755-760] prior to refinement. Our earlier 2.7 A structure [Cheah, Carr, Suffolk, Vasudevan, Dixon & Ollis (1994). Structure, 2, 981-990] served as a starting model. P(II) is a trimeric molecule, each subunit has a mass of 12.4 kDa and contains 112 amino acid residues. The refined model includes all 1065 protein atoms per subunit plus 312 water molecules. The high-resolution refinement confirms the correctness of our 2.7 A model, although it leads to a redefinition of the extent of various secondary-structural elements. The monomeric structure of P(II) exhibits an interlocking double betaalphabeta fold. This is a stable fold found in a number of proteins with diverse functions. The association of the protein into a trimer leads to a new structure which we describe in detail. The effects of crystal packing forces are discussed and potential interaction sites with other proteins and effector molecules are identified. PMID- 15299731 TI - Structure of a bulgecin-inhibited g-type lysozyme from the egg white of the Australian black swan. A comparison of the binding of bulgecin to three muramidases. AB - Bulgecin A, a bacterial metabolite, has been shown to bind in the active-site groove of the chicken-type lysozyme from the rainbow trout (RBTL) and in the lysozyme-like C-terminal domain, of a soluble lytic transglycosylase (C-SLT) from Escherichia coli. These enzymes are muramidases that cleave the glycosidic bonds in the glycan strands of the murein polymer. Here we report the crystal structure of a complex between the goose-type lysozyme from the egg white of the Australian black swan (SEWL) and bulgecin A at 2.45 A resolution. As is the case for the C SLT/bulgecin and RBTL/bulgecin complexes, the ligand binds with the N acetylglucosamine ring in subsite C and the proline moiety in site D where it interacts with the catalytic glutamic acid. The taurine residue interacts with the beta-sheet region. Comparisons of the three buigecin complexes show that the inhibitor has the same binding mode to the muramidases with similar protein ligand interactions, particularly for SEWL and RBTL. From our results, it seems likely that bulgecin, in general, inhibits enzymes with lysozyme-like domains and thus might represent a novel class of natural antibiotics that act on murein degrading rather than murein-synthesizing enzymes. PMID- 15299732 TI - Structure of a complex between bulgecin, a bacterial metabolite, and lysozyme from the rainbow trout. AB - Bulgecin, a sulfonated glycopeptide produced by Pseudomonas acidophila and Pseudomonas mesoacidophila, induces bulge formation and enhances lysis of bacterial cell walls when used in combination with beta-lactam antibiotics. The compound does not itself exhibit any antibacterial activity, but has been shown to inhibit a soluble lytic transglycosylase (SLT70) from Escherichia coli which has a lysozyme-like domain. Recently, the crystal structure of an SLT-bulgecin complex has been determined to 3.5 A resolution. We report here the crystal structure of a complex between lysozyme from the rainbow trout (RBTL) and bulgecin A at 2.0 A resolution. As for the SLT-bulgecin complex, bulgecin is bound with the glycosaminyl moiety in subsite C and the proline residue in site D of the active-site cleft of RBTL, where it makes hydrogen-bonding interactions with the catalytic residues. The taurine moiety is bound to the left side of subsites E and F in the lower part of the active-site cleft. From the observed position of the bulgecin molecule, it seems reasonable that it is an inhibitor of rainbow trout lysozyme. The lysozymes may, in general, be a target for the design of a novel type of antibiotics distinct from the beta-lactams which are insensitive to the muramidases. PMID- 15299733 TI - Structure of a loop-deleted variant of 3-isopropylmalate dehydrogenase from Thermus thermophilus: an internal reprieve tolerance mechanism. AB - A loop-deleted mutant form of 3-isopropylmalate dehydrogenase from Thermus thermophilus was constructed to investigate the relationship between the flexibility of the structure and the thermostability of the enzyme. The structure of the mutant enzyme was determined by X-ray crystallography and was found to be almost the same as that of the native enzyme with a reduced temperature factor. Although the mutant protein had lost the flexible loop, its function and thermostability had remained unchanged. This phenomenon can be explained by an internal reprieve tolerance mechanism. PMID- 15299734 TI - Structural refinement of the DNA-containing capsid of canine parvovirus using RSRef, a resolution-dependent stereochemically restrained real-space refinement method. AB - The canine parvovirus structure (CPV) [Tsao, Chapman, Agbandje, Keller, Smith, Wu, Luo, Smith, Rossmann, Compans & Parrish (1991). Science, 251, 1456-1464] has been refined by a real-space refinement procedure [Chapman (1994). Acta Cryst. A51, 69-801. The fit of an atomic model to electron density was optimized while taking into account the resolution limit of the data and the stereochemistry of the structure. The refined model had a reasonable free R factor [Brtinger (1992). Nature (London), 355, 472-4751 of 0.29. The method is particularly fast and convenient when only a small fraction of the crystallographic asymmetric unit needs to be refined, as is the case when there is high non-crystallographic redundancy. Cycles of refinement for virus capsids were completed in about 1/50th of the time required for equivalent reciprocal-space procedures. PMID- 15299735 TI - Structure solution of a cubic crystal of concanavalin A complexed with methyl alpha-D-glucopyranoside. AB - The solution of the cubic crystal form (a = 167.8 A) of concanavalin A complexed with the monosaccharide methyl alpha-D-glucopyranoside is described. The space group has been determined as I2(1)3 rather than I23. The use of cadmium to replace cobalt at the transition metal-ion binding site and to replace calcium at its binding site proved to be crucial to the successful solution of the crystal structure. The relatively small isomorphous signals of 21 e(-) for the replacement of cobalt and 28 e(-) for the replacement of calcium, yielded interpretable difference Patterson maps. The electron-density map calculated in space group I2(1)3 at 5.4 A resolution, based on phases derived from single- and double-substituted cadmium differences, revealed a classical concanavalin A tetramer of 222 point symmetry, as seen in all the known crystal structures of concanavalin A. Rigid-body refinement at 3.6 A using the refined coordinates of saccharide-free concanavalin A converged to an R factor of 27.4%. A molecular replacement analysis, consistent with this crystal structure, and initial experiences in the incorrect space group I23 are described as these also prove to be instructive. PMID- 15299736 TI - Crystallization of apocrustacyanin on the International Microgravity Laboratory (IML-2) mission. AB - Rod-shaped crystals of apocrustacyanin C1 have been grown under microgravity on the International Microgravity Laboratory (IML-2) NASA space shuttle mission using the vapour-diffusion set-up of the Advanced Protein Crystallization Facility (APCF). The crystals obtained under microgravity are compared with crystals grown simultaneously in ground control experiments in identical APCF reactors, and with those obtained in the laboratory. The degree of reproducibility of the results in microgravity was also tested. Statistically, the microgravity-grown crystals are larger and of better X-ray diffraction quality than those grown in the ground controls but inferior to the best crystals grown in sitting drops, in the laboratory. Diffracting crystals, the best to 2.3 A, were produced in seven out of the eight reactors in microgravity, whereas the eight ground control reactors yielded only one poorly formed crystal suitable for diffraction studies, which also diffracted to 2.3 A. The crystals belong to the space group P2(1)2(1)2(1) with two subunits per asymmetric unit. PMID- 15299737 TI - Structure of a ribonuclease B+d(pA)4 complex. AB - The structure of a tetragonal crystal of bovine pancreatic RNase B complexed with d(pA)(4) was determined by molecular replacement and difference Fourier methods. This crystal belongs to space group P4(1)2(1)2 and has unit-cell dimensions a = b = 44.5, c = 156.5 A. The model consists of the enzyme and a tetranucleotide with fractional occupancies, suggesting multiple modes of oligonucleotide binding. It does not include any polysaccharide residues or solvent molecules. After refinement at 2.7 A, the R value was 0.163 with acceptable stereochemistry. The model illustrates a set of well defined interactions for substrate binding, particularly between the central dinucleotide and the enzyme. PMID- 15299738 TI - X-ray structure of a new crystal form of pike 4.10 beta parvalbumin. AB - A new crystal form of pike (pI 4.10) parvalbumin has been crystallized in presence of EDTA at pH 8.0. The crystals are orthorhombic, space group P2(1)2(1)2(1), with a = 51.84, b = 49.95, c = 34.96 A. Diffractometer data were collected to 1.75 A. The structure was solved by molecular replacement and refined to R = 0.168 for 7774 observed reflections [I>/= 2sigma(I)] in the range 8.0-1.75 A. In spite of the presence of EDTA, calcium ions are present in both primary binding sites. As compared to the previously reported structures, the main differences concern the conformation of the N-terminal residues and the packing in the unit cell. PMID- 15299739 TI - Studies of monoclinic hen egg-white lysozyme. IV. X-ray refinement at 1.8 A resolution and a comparison of the variable regions in the polymorphic forms. AB - Monoclinic crystals of hen egg-white lysozyme (E.C. 3.2.1.17, HEL) grown at low pH in the presence of NaNO(3) belong to space group P2(1) with unit-cell dimensions, a = 28.0, b = 62.5, c = 60.9 A and beta= 90.8 degrees with two molecules in the asymmetric unit. 1.8 A resolution intensity data, collected on a CAD-4 diffractometer, contained 17 524 reflections with F > 3sigma (93% complete). Our earlier preliminary 1.8 A model was refitted and refined using X PLOR to an R value of 0.189. The deviations in the model from ideal geometry are 0.013 A in bond lengths and 2.8 degrees in bond angles. The r.m.s. deviation in the backbone atoms between the two molecules is 0.42 A. A comparison of HEL in different polymorphic crystal forms reveals that the prominent structural variability among them resides in two exposed regions 45-50 and 65-73 which are also regions of lattice contacts. PMID- 15299740 TI - Three-dimensional structure of Xenopus laevis Cu,Zn superoxide dismutase b determined by X-ray crystallography at 1.5 A resolution. AB - Xenopus laevis Cu,Zn superoxide dismutase (recombinant isoenzyme b) has been crystallized and the structure determined at 1.49 A resolution. The crystals belong to space group P2(1)2(1)2(1), with cell constants a = 73.33, b = 68.86, c = 59.73 A, and contain one dimeric molecule of M(r) 32 000 per asymmetric unit. The structure was solved by molecular-replacement techniques using the semisynthetic Cu,Co bovine enzyme as search model, and refined by molecular dynamics with a crystallographic pseudo-energy term. During the final steps, positional and anisotropic thermal parameters of the atoms were refined. The R factor for the 49 209 unique reflections in the 10.0-1.49 A resolution range is 0.104, for a model comprising 2023 protein atoms, two Cu(2+), two Zn(2+), and 353 water molecules. The overall temperature factor for the model, including solvent, is 20.3 A(2), while the calculated r.m.s. coordinate error for the refined model is 0.036 A. As suggested by the primary structure homology to any other known intracellular eukaryotic superoxide dismutase (> 50%), the typical structural scaffolding of flattened antiparallel eight-stranded (beta-barrel is well conserved in X. laevis Cu,Zn superoxide dismutase b, together with the coordination geometry of the metal centers in the active site. The higher thermal stability of the bb X. laevis superoxide dismutase homodimer, with respect to dimers involving the a-type isoenzyme subunit(s), can be related, on the basis of the high-resolution structure, to side-chain and solvent interactions centered on residue Tyr149, in both b-type subunits. The analysis of the overall solvent structure reveals a number of equivalent water molecule sites in the two subunits, and in homologous superoxide dismutase models. Their locations are discussed in detail and classified on the basis of their structural role. PMID- 15299741 TI - Crystallization and preliminary X-ray analysis of a bacterial glutathione transferase. AB - Crystals of a bacterial glutathione S-transferase from Proteus mirabilis have been grown from polyethylene glycol by the hanging-drop vapour-diffusion method. Successful crystallization required the presence of the substrate glutathione. The crystals belong to the tetragonal space group P4 with cell dimensions a = b = 90.9 and c = 117.3 A. They contain between three and six monomers in the asymmetric unit and diffract to beyond 2.3 A resolution. PMID- 15299742 TI - Crystallization and preliminary X-ray crystallographic analysis of recombinant transaldolase B from Eschericha coli. AB - Recombinant transaldolase from Escherichia coli, an enzyme of the pentose phosphate pathway has been crystallized by the vapor-diffusion method using polyethylene glycol 6000 as precipitant. The crystals are orthorhombic, space group P2(1)2(1)2(1) with cell dimensions a = 68.9, b = 91.3 and c = 130.5 A and diffract to 2 A resolution on a conventional X-ray source. The asymmetric unit very likely contains two subunits, corresponding to a packing density of 2.9 A(3) Da(-1). PMID- 15299743 TI - Preliminary crystallographic studies of dimethylsulfoxide reductase from Rhodobacter capsulatus. AB - Dimethylsulfoxide reductase from the photosynthetic bacterium Rhodobacter capsulatus has been crystallized in two similar forms which are suitable for X ray structure determination. Both crystals forms belong to space group P4(1)22 or P4(3)22, with cell dimensions a = b = 80.81, c = 229.75 A (type I crystals) or a = b = 89.30, c = 230.05 A (type II crystals) and one molecule in the asymmetric unit. Diffraction has been observed to at least 2.0 A in type I crystals and to 2.6 A in type II crystals. Dimethylsulfoxide reductase from Rhodobacter is the simplest molybdenum oxotransferase known and this makes it an ideal model to study the structure and function of this class of enzymes. PMID- 15299744 TI - Purification, crystallization and preliminary X-ray crystal structure analysis of copper amine oxidase from Arthrobacter globoformis. AB - beta-Phenylethylamine oxidase from the Gram-positive bacterium Arthrobacter globoformis has been crystallized as three crystal forms. Two belong to space group C2 and one to space group P2(1)2(1)2(1), respectively. The unit-cell volumes are consistent with one infunit of 70 644 Da per asymmetric unit for the two monoclinic forms, and with two infunits per asymmetric unit for the orthorhombic crystals. Three-dimensional intensity data have been recorded to 2.8A resolution for one of the monoclinic crystal forms and to 3A, resolution for the orthorhombic crystal form. PMID- 15299745 TI - Crystallization and preliminary X-ray study of a new crystal form of cytochrome c' from Rhodobacter capsulatus. AB - A new crystal form of diheme cytochrome c' from Rhodobacter capsulatus has been obtained and preliminary crystallographic experiments have been performed. The crystals belong to the space group P2(1)2(1)2 with unit-cell dimensions of a = 47.82, b = 72.59, c = 34.32 A. The assumption that an asymmetric unit of the crystal contains one half of the homodimer molecule indicates that the monomers in the dimeric molecule may be related by a crystallographic twofold axis. Crystals diffract up to 1.7 A resolution using the X-ray beam from synchrotron radiation, and 11 127 unique structure factors were obtained with an R(merge) of 7.1% from 52 922 indexed reflections. Structure analysis by means of molecular replacement methods is now underway. PMID- 15299746 TI - Crystallization and preliminary X-ray diffraction studies of haemorrhagin I from the snake venom of Agkistrodon acutus. AB - Haemorrhagin I from the snake venom of Agkistrodon acutus (AaHI) has been crystallized using the hanging-drop vapour diffusion method. The crystals belong to space group P4(1)2(1)2 or P4(3)2(1)2 with unit-cell dimensions a = b = 63.61 and c = 95.69 A. There is one molecule in the asymmetric unit. Data to 2.35 A resolution have been collected using a single-crystal. PMID- 15299747 TI - Crystallization of a 14-3-3 protein. AB - Crystals of the tau (tau) isoform of the 14-3-3 family of proteins were grown and shown to belong to the orthorhombic space group P2(1)2(1)2(1) with cell dimensions a = 70.29, b = 79.3, c = 101.00 A. The crystals were needle-like in morphology and less than 10 micro m in two dimensions. Diffraction data were collected using synchrotron radiation sources from flash-cooled crystals. Native data extended to a resolution of 2.6 A and mercury and platinum derivatives diffracted to 3.4 and 3.9 A, respectively. The structure has been solved recently. Here the protein crystallization procedures, the characterization of the crystals and the correlation between crystal habit and diffraction quality are reported. PMID- 15299748 TI - Crystallization and preliminary X-ray analysis of twinned crystals of a chimeric FK506 binding protein 12 and 13 complexed with FK506. AB - An FKBPI2/13 chimera with the 80s loop of FKBPI3 replacing the corresponding loop in FKBPI2 tightly binds the immunosuppressive agents FK506 and rapamycin and efficiently catalyzes peptidyl-prolyl cis-trans isomerization. However, the chimera's complex with FK506 does not inhibit calcineurin's phosphatase activity [Yang, Rosen & Schreiber (1993). J. Am. Chem. Soc. 115(2), 819-820]. The chimeric protein crystallizes in space group P1 and the crystals are always twinned. The twin composites are related by a twofold twinning axis parallel to the a axis. A resolution data set (1.5 A resolution) for a twinned crystal was collected at CHESS using 0.91 A X-rays and image plates. Preliminary molecular replacement using data between 15 and 3 A and the FKBPI2-FK506 crystal structure as the search model led to a clear solution with a residual of 34.2%. This 3 A resolution structure provides insight into the structural basis of twinning. PMID- 15299749 TI - Crystallization of Escherichia coli aspartyl-tRNA synthetase in its free state and in a complex with yeast tRNA(Asp). AB - Overexpressed dimeric E. coli aspartyl-tRNA synthetase (AspRS) has been crystallized in its free state and complexed with yeast tRNA(Asp). Triclinic crystals of the enzyme alone (a = 104.4, b = 107.4, c = 135.0 A, alpha = 102.9, beta = 101.0, gamma = 106.3 degrees ), have been grown using ammonium sulfate as the precipitant and monoclinic crystals (a = 127.1, b = 163.6, c = 140.1 A, beta = 111.7 degrees ), space group C2, have been grown using polyethylene glycol 6000. They diffract to 2.8 and 3.0 A, respectively. Crystals of the heterologous complex between E. coli AspRS and yeast tRNA have been obtained using ammonium sulfate as the precipitant and 2-propanol as the nucleation agent. They belong to the monoclinic space group P2(1) (a = 76.2, b = 227.3, c = 82.3 A, beta = 111.7 degrees ) and diffract to 2.7 A. PMID- 15299750 TI - Crystallization and preliminary X-ray crystallographic studies of 7alpha hydroxysteroid dehydrogenase from Escherichia coli. AB - Crystals of 7alpha-hydroxysteroid dehydrogenase from E. coli, which is a tetramer in its active form, have been obtained by a hanging-drop vapor-diffusion method in the presence of the coenzyme NAD(+). Crystals as large as 0.25 x 0.25 x 0.75 mm could be grown within a month at pH 8.5 with polyethylene glycol as precipitating agent. Preliminary X-ray crystallographic analysis revealed that they belong to one of the enantiomorphic space groups P4(1)2(1)2 or P4(3)2(1)2 with dimensions a = b = 81.66 and c = 214.6 A, having two subunits in an asymmetric unit. The crystals diffract to at least 2.3 A resolution. PMID- 15299751 TI - Crystallization and preliminary X-ray diffraction studies of a cobalt-substituted derivative of the iron-dependent alcohol dehydrogenase from Zymomonas mobilis. AB - The iron-dependent alcohol dehydrogenase from Zymomonas mobilis has been crystallized in a form suitable for X-ray diffraction studies. The crystals grew in hanging drops by vapor diffusion, equilibrating with a solution comprising 25 27% methoxypolyethylene glycol 5000 and 1 mM Co(2+) in a 0.2 M succinic acid/potassium hydroxide buffer at pH 5.5-5.7 at 281 K. Crystals are tetragonal, P4(1)22 (or P4(3)22), with unit-cell dimensions a = b = 125.7, c = 248.1 A. Four molecules comprise the asymmetric unit, and a self-rotation function indicates twofold local symmetry perpendicular to the unique axis and 15 degrees from a crystallographic twofold axis. Diffraction data to 3.0 A have been collected. PMID- 15299752 TI - Crystallization and preliminary crystallographic analysis of 3-carboxy-cis,cis muconate lactonizing enzyme from Neurospora crassa. AB - Crystals of 3-carboxy-cis,cis-muconate lactonizing enzyme (CMLE; E.C. 5.5.1.5) from Neurospora crassa that diffract to high resolution have been obtained. The crystals belong to the orthorhombic space group P2(1)2(1)2(1) with unit-cell dimensions a = 92.1, b = 159.7, c = 236.6 A (at 103 K) and diffract at most to 2 A resolution. The asymmetric unit of the crystals appears to contain two tetrameric CMLE molecules making up a total of 328 kDa per asymmetric unit. Both cross-linking with glutaraldehyde and cryo-cooling to 103 K have been used to facilitate data collection because the crystals are unstable in the X-ray beam; both techniques extend the crystal lifetime but cryo-cooling, unlike glutaraldehyde cross-linking, does not lower the quality of the diffraction pattern. PMID- 15299753 TI - Crystallization of microsomal triglyceride transfer protein from bovine liver. AB - The microsomal triglyceride transfer protein (MTP) is a heterodimeric lipid transfer protein required for the assembly of plasma very low density lipoproteins in the liver and chylomicrons in the intestine. Bovine MTP was purified by a modification of a previously published procedure and crystals of MTP were grown reproducibly with polyethylene glycol as a precipitant at pH 7.0. MTP crystals, which diffract to Bragg spacings of better than 3.2 A, have the symmetry of space group P2(1)2(1)2(1) with refined lattice constants of a = 88.7, b = 100.9 and c = 201.1 A, with one heterodimer per asymmetric unit. PMID- 15299754 TI - Crystallization and preliminary X-ray analysis of bacteriophage T4 deoxynucleotide kinase. AB - T4 deoxynucleotide kinase catalyzes the phosphorylation of 5 hydroxymethyldeoxycytidylate, dTMP and dGMP while excluding dCMP and dAMP. In order to understand the mechanism of this remarkable specificity, the enzyme was over-expressed in Escherichia coli, purified and crystallized for X-ray diffraction analysis. The crystals belong to the monoclinic space group C2 with cell dimensions a = 155.2, b = 58.5, c = 75.7 A, beta = 108.1 degrees. There are two protein monomers in the asymmetric unit related by a twofold axis. Diffraction data to 2.0 A resolution have been collected. PMID- 15299755 TI - Report of a workshop on the use of statistical validators in protein X-ray crystallography. PMID- 15299756 TI - Laue diffraction studies of human rhinovirus 14 and canine parvovirus. AB - Laue diffraction data have been collected from monoclinic crystals of canine parvovirus (CPV), and from cubic crystals of human rhinovirus 14 (HRV14) with and without bound antiviral compounds. In optimal conditions one or two images of HRV14 were sufficient to calculate interpretable electron-density maps of the virus complexes at 3.5 A resolution. The crystals of CPV were of lower symmetry and were more easily damaged by radiation, making it difficult to accumulate a significant amount of useful data. Results on HRV14 are compared in studies on four antiviral compounds where data were collected using both monochromatic and Laue diffraction. Two Laue diffraction images of HRV14 with a point mutation were sufficient to determine the change from a leucine to a valine in VP2. PMID- 15299757 TI - Structure determination of coxsackievirus B3 to 3.5 A resolution. AB - The crystal structure of coxsackievirus B3 (CVB3) has been determined to 3.5 A resolution. The icosahedral CVB3 particles crystallize in the monoclinic space group, P2(1), (a = 574.6, b = 302.1, c = 521.6 A, beta = 107.7 degrees ) with two virions in the asymmetric unit giving 120-fold non-crystallographic redundancy. The crystals diffracted to 2.7 A resolution and the X-ray data set was 55% complete to 3.0,4, resolution. Systematically weak reflections and the self rotation function established pseudo R32 symmetry with each particle sitting on a 32 special position. This constrained the orientation and position of each particle in the monoclinic cell to near face-centered positions and allowed for a total of six possible monoclinic space-group settings. Correct interpretation of the high-resolution (3.0-3.2 A) self-rotation function was instrumental in determining the deviations from R32 orientations of the virus particles in the unit cell. Accurate particle orientations permitted the correct assignment of the crystal space-group setting amongst the six ambiguous possibilities and for the correct determination of particle positions. Real-space electron-density averaging and phase refinement, using human rhinovius 14 (HRV14) as an initial phasing model, have been carried out to 3.5 A resolution. The initial structural model has been built and refined to 3.5 A resolution using X-PLOR. PMID- 15299758 TI - On the solution of the molecular-replacement problem at very low resolution: application to large complexes. AB - The applicability of the molecular-replacement (MR) method, implemented through the AMoRe package [Navaza (1994). Acta Cryst. A50, 157-163], is studied at very low resolution (d > 20 A) and for very large molecular complexes. Due to the nature of the low-resolution data, specific problems appear. In particular, rotation-function peaks are very broad and translation functions based on Patterson overlap show large spurious peaks. To solve these problems, the translation function is replaced by a search using amplitude correlation and a systematic three-dimensional angular search is performed around each rotation function peak. Furthermore, these functions are applied in different resolution ranges during the same search. The corresponding algorithms are applied to two cases: the tRNA(Asp)-synthetase complex (neutron diffraction data) and a ribosome model crystal (calculated data). This new implementation is shown to solve the problem for a variety of search models, ranging from a detailed atomic model to a rough envelope. PMID- 15299759 TI - On the ab initio solution of the phase problem for macromolecules at very low resolution: the few atoms model method. AB - A method is proposed for the solution of the phase problem at very low resolution for macromolecules. It generates randomly a very large number of models, each consisting of a few (two to ten) pseudo-atoms. The corresponding amplitudes are used for selecting a subset of 'best' models by choosing those with the highest correlation with experimental values. The phases calculated from these 'best' models are analysed by a clusterization procedure leading to a few possible solutions, from which the correct one can be recognized by simple additional criteria. This method has been successfully applied to the neutron diffraction data of the AspRS-tRNA(Asp) complex at 50 A resolution and to data calculated from a model ribosome crystal at 60 A resolution. PMID- 15299760 TI - Refinement of purothionins reveals solute particles important for lattice formation and toxicity. Part 1: alpha1-purothionin revisited. AB - The three-dimensional structure of alpha(1)-purothionin (alpha(1)-PT), a wheat germ protein and a basic lytic toxin, was previously solved by molecular replacement methods using an energy-minimized predicted model and refined to an R factor of 21.6% [Teeter, Ma, Rao & Whitlow (1990). Proteins Struct. Funct. Genet. 8, 118-1321. Some deficiencies of the model motivated us to revisit the structure and to continue the refinement. Here we report a significantly improved structure refined to an R-factor of 15.5% with excellent geometry. The refinement of this relatively low resolution structure ( approximately 2.8 A) is well suited to test the limitations of classical methods of refinement and to address the problem of overfitting, The final structure contains 434 atoms including 330 protein atoms, 70 waters, three acetates, two glycerols, one sec-butanol and one phosphate. The key solute molecules (acetate ion and phosphate ion) play a crucial role in the lattice formation. Phosphate and glycerol found in the structure may be important for biological activity of the toxins. PMID- 15299761 TI - Refinement of purothionins reveals solute particles important for lattice formation and toxicity. Part 2: structure of beta-purothionin at 1.7 A resolution. AB - The crystal structure of beta-purothionin (beta-PT) has been determined at 1.7 A resolution. beta-PT and previously solved alpha(l)-PT belong to a family of membrane-active plant toxins homologous to crambin. (beta-PT crystallizes in the same space group as alpha(l)-PT (1422) but with the c axis 3 A longer than (alpha(l)-PT. The unit-cell dimensions of beta-PT crystals are a = b = 53.94 and c = 72.75 A. Two data sets were collected on a multiwire area detector, each with R(sym) around 6.0%, and were merged to get a single data set at 1.7 A, (R(merge) = 9.6%). The X-ray structure of alpha(l)-PT was used to build a starting model for beta-PT. The beta-PT model was refined using the program PROLSQ from 10 to 1.7 A resolution to an R-factor of 19.8% with very good geometry. The final structure contains 439 atoms including 337 protein atoms, 77 waters, two acetates, two glycerols and one phosphate. The high-resolution structure of the beta-PT agreed well with that of the lower resolution alpha(l)-PT structure only after the latter was extensively rerefined. Both refinements revealed phosphate and glycerol molecules which are important in lattice formation. The binding of phosphate and glycerol molecules to purothionins (PT) was confirmed by NMR and was implicated in the biological activity of toxins. Modeling of phospholipid binding to PT based on glycerol and phosphate-binding site could shed light on the lytic toxicity of this protein-toxin family. Although the structures of (alpha(l)-PT and beta-PT preserve the overall fold of crambin, they exhibit key differences that are directly relevant to the toxicity of thionins. PMID- 15299762 TI - Structure of native pancreatic elastase from North Atlantic salmon at 1.61 A resolution. AB - The crystal structure of native salmon pancreatic elastase (SPE) has been solved by molecular-replacement methods, and refined by conventional conjugate-gradient methods and simulated-annealing techniques. The final R value is 17.2% for 21 389 reflections between 8.0 and 1.61 A, and the corresponding free R value is 23.9%. The overall tertiary structure of SPE is remarkably similar to that of porcine pancreatic elastase I (PPE), to which it shows about 67% sequence identity. The primary structure of SPE is determined from the electron-density maps, and only about 15 side chains are somewhat uncertain. Interesting differences between SPE and PPE, are one sequence deletion assigned to position 186, the residue 192 at the entrance of the specificity pocket is substituted from a Gln in PPE to Asn in SPE, and one of the calcium ligands is different. Furthermore, electron density is missing in SPE for the last three residues of the C-terminal helix. A comparison of the present amino-acid sequence of SPE with other sequences available indicates that SPE belongs to the class 1 pancreatic elastases. PMID- 15299763 TI - Preliminary characterization of EcoRI-DNA co-crystals: incomplete factorial design of oligonucleotide sequences. AB - A full understanding of the sequence specificity of EcoRI endonuclease requires structural information on complexes where the DNA contains one 'incorrect' base pair; historically, these sites are referred to as EcoRI* sites. They are inherently asymmetric, unlike the canonical EcoRI site, GAATTC, which possesses a twofold axis of rotational symmetry. All previously determined DNA-EcoRI complexes incorporated this symmetry axis into the space group, requiring the design of 'new' oligonucleotides to produce an asymmetric unit appropriate to an EcoRI* complex. The incomplete factorial approach of Carter & Carter [Carter & Carter (1979). J. Biol. Chem. 254, 12219-12223.] was used to design the DNA sequence. Factors included the location and type of EcoRI* substitution and the length and AT richness of the sequences on both sides of the RI site. Co-crystals were obtained with several sequences, including one with TCGTGGACTTCGTG. Diffraction data were collected from one crystal of this complex to 3.2 A resolution; the unit-cell parameters are a = b = 123.8 and c = 148.9 A and the space group is P3(2)21. Unit-cell and space-group information was also obtained for the EcoRI* sites AAATTC, GGATTC and GAGTTC. These experiments demonstrated the need for a rapid, economical method that would distinguish DNA-protein co crystals from crystals of protein only. This can be readily achieved with a single small crystal and a staining method based on methylene blue and methyl violet, which stain DNA and protein, respectively. PMID- 15299764 TI - Human class II MHC molecule HLA-DR1: X-ray structure determined from three crystal forms. AB - The three-dimensional structure of the extracellular region of a 60 kDa class II major histocompatibility glycoprotein, HLA-DR1, was determined to 3.3 A by X-ray crystallography using three crystal forms, each containing two molecules per asymmetric unit. Phases were initially determined to 4.2 A using two crystal forms both containing DR1 from human lymphocytes complexed with a mixture of endogenous peptides. One of these crystal forms also contained a 28 kDa superantigen, Staphylococcus aureus enterotoxin B (SEB), bound to each DR1 molecule. Single-isomorphous replacement phasing followed by iterative two- and fourfold non-crystallographic real-space averaging between the two crystal forms resulted in 4.2 A resolution electron-density maps from which the paths of the polypeptides could be traced. Cryocrystallography and synchrotron radiation were then used to extend the resolution to 3.3 A for the two lymphocyte-derived crystal forms and for a third crystal form grown from DR1 produced in insect cells and complexed in vitro with a specific antigenic peptide. Iterative sixfold non-crystallographic real-space averaging resulted in an electron- density map into which 340 of 371 residues could be fit unambiguously. Crystal contacts and the existence of a parallel dimer of the DR1 alphabeta heterodimer in the three crystal forms are discussed. PMID- 15299765 TI - Crystal structures of three complexes between chito-oligosaccharides and lysozyme from the rainbow trout. How distorted is the NAG sugar in site D? AB - Like all c-type lysozymes, those from rainbow trout act as 1,4-beta-acetyl muramidases to destroy bacteria by cleaving the polysaccharide chains of alternating N-acetylglucosamine (NAG) and N-acetylmuramic acid (NAM) units in the cell walls. Lysozymes also hydrolyse chitin, the analogous N-acetylglucosamine polymer. The rainbow trout enzymes have been shown to be particularly effective in bacterial defence. We have determined the crystal structures of three complexes between rainbow trout lysozyme (RBTL) and the chito-oligosaccharides (NAG)(2), (NAG)(3) and (NAG)(4) to resolutions of 1.8, 2.0 and 1.6 A, respectively. Crystals of these complexes were obtained by co-crystallization, and intensity data were collected on a FAST area detector system. Refinement and model building gave final R values of 16.6, 15.9 and 16.5% for the di-, tri- and tetrasaccharide complexes, respectively. The results show that the chito oligosaccharides bind to sites A, B and C as previously observed for complexes between the hen egg-white lysozyme (HEWL) and a variety of saccharides. The NAG ring in site D is not bound so deeply and is only slightly distorted towards a half-chair conformation as observed for the equivalent NAM residue in HEWL. From our results, there is reason to question the position and the degree of strain of the D saccharide and the mode of binding and importance of two saccharides in sites E and F for correct orientation of sugar D and effective hydrolysis of a productive substrate-lysozyme complex. Simple model building study from our structures implies a 'left-sided' binding mode of (NAG)(6) in the lower part of the active site of RBTL. PMID- 15299766 TI - On the application of anomalous scattering in oligonucleotide crystallography. AB - Simulated anomalous-scattering differences, at wavelengths between 1.5 and 5.5 A, were used with MULTAN to locate P atoms in an oligonucleotide hexamer. The success of the method depended heavily on the level of errors in the data. With error-free data most or all P atoms were located at all wavelengths. With noisy data, the best results were obtained by refining the phases associated with the largest values of |DeltaF|/sigma(|DeltaF|) rather than with the largest values of |DeltaF|. In this case a few of the P-atom positions could be located, with the best results occurring at wavelengths between 3.0 and 4.0 A. Further improvements were gained by reducing the values of the thermal parameters of the P atoms. MULTAN figures of merit had limited success in indicating the best phase sets, but a small improvement was gained by modifying the procedure for selecting those reflections used in the calculation of PSIZERO. PMID- 15299767 TI - Phase improvement by cross-validated density modification. AB - Solvent flattening is a useful constraint for the early stages of crystallographic structure determination. However, sometimes it fails to produce significant improvement of poor experimental or molecular-replacement phases. This often occurs as a result of incorrect parameterization. In addition, the potential of overfitting or misinterpretation of the data exists. We have implemented a cross-validated (or free) R value in order to reduce this risk. The free R value was calculated between the experimental F(obs)(h) and the calculated structure factors, F(sf)(h), obtained by inverse Fourier transformation of the solvent-flattened electron density. Because of the sensitivity of the free R value to the test set selection at low resolution complete cross-validation may be required. The reliability of this approach was assessed by examining the correlation between the free R value and the known phase errors for two test cases. A high correlation was found upon variation of the extent of negative density elimination, figure of merit estimation, and the relative weighting in the phase combination procedure. The free R value can be used to optimize parameters of density-modification procedures when independent phase error estimates are unavailable. PMID- 15299768 TI - 1.6 A structure of semisynthetic ribonuclease crystallized from aqueous ethanol. Comparison with crystals from salt solutions and with ribonuclease A from aqueous alcohol solutions. AB - The non-covalent combination of residues 1-118 of RNase A with a synthetic 14 residue peptide containing residues 111-124 of the molecule forms a highly active semisynthetic enzyme, RNase 1-118:111-124. With this enzyme, the roles played by the six C-terminal residues in generating the catalytic efficiency and substrate specificity of RNase can be studied using chemically synthesized analogs. The structure of RNase 1-118:111-124 from 43% aqueous ethanol has been determined using molecular-replacement methods and refined to a crystallographic R-factor of 0.166 for all observed reflections in the range 7.0-1.6 A (Protein Data Bank file ISSC). The structure is compared with the 2.0 A structure of RNase A from 43% aqueous 2-methyl-2-propanol and with the 1.8 A structure of the semisynthetic enzyme obtained from crystals grown in concentrated salt solution. The structure of RNase 1-118:111-124 from aqueous ethanol is virtually identical to that of RNase A from aqueous 2-methyl-2-propanol. Half of the crystallographically bound water molecules are not coincident, however. The structure is somewhat less similar to that of RNase 1-118:111-124 from salt solutions, with a major difference being the positioning of active-site residue His119. PMID- 15299769 TI - X-ray structure of wheat germ agglutinin isolectin 3. AB - Wheat germ agglutinin isolectin 3 (WGA3) was crystallized from 10 mM acetate buffer at pH 4.9 containing 6 mM CaCl(2) and 4%(v/v) ethanol. The crystal belongs to monoclinic space group P2(1) with unit-cell dimensions a = 44.86, b = 91.02, c = 44.86 A, and beta = 110.22 degrees. The asymmetric unit contains two molecules (V(m) = 2.51 A(3) Da(-1)). The crystal structure was solved by the molecular replacement method and was refined by the simulated-annealing method. The conventional R value was 0.191 for 19713 reflections [|F(o)| > 3sigma(F)] in the resolution range 8-1.9 A. The r.m.s. deviations from the ideal bond distances and angles were 0.014 A, and 3.0 degrees, respectively, and the estimated coordinate error was 0.2-0.25 A. The two molecules in the asymmetric unit are related by the pseudo twofold symmetry and form a dimer structure. The backbone structures of the two subunits are nearly identical with the r.m.s. difference of 0.36 A for the superposition of equivalent C(alpha) atoms. The dimer structure is very similar to those of isolectins 1 and 2 with the r.m.s. difference of 0.35-0.39 A for the C(alpha) superposition. Since amino-acid residues which differ from those of isolectin 1 or 2 are not involved in the contact between the two subunits, the subunit-subunit interaction is not significantly affected by the replacement of these residues. As a result, the geometry of the sugar-binding sites which are located at the interface between the two subunit molecules is basically conserved among three isolectins. PMID- 15299770 TI - Use of 'random-atom" phasing models to determine macromolecular heavy-atom replacement positions. AB - A procedure is described by which the phase-invariant translation function may be used to provide starting set phases for the minimal function that are significantly better than those generated from random-atom coordinates. Applications to determine the heavy-atom positions for single isomorphous replacement data from macromolecular structures are very encouraging. In the case of chiral space groups, e.g. P4(1)2(1)2 versus P4(3)2(1)2, unlike Patterson functions, these methods provide the correct enantiomorph coordinates for the heavy-atom sites for whichever space group is chosen. PMID- 15299771 TI - Sodium ions and water molecules in the structure of poly(dA).poly(dT). AB - Analysis of X-ray diffraction data from a polycrystalline and well oriented fiber of the sodium salt of poly(dA).poly(dT) shows that this B'-DNA corresponds to a right-handed antiparallel tenfold double-helix of pitch 32.4 A, with C2'-endo furanose rings in both strands. The helix contrasts itself from B-DNA in terms of a very narrow minor groove. Difference electron-density maps have revealed that a continuous spine of water molecules, two per base pair, propagates along this groove with the same symmetry as the DNA and establishes new links between the two strands. In addition to this hydrated DNA helix, the monoclinic unit cell (space group P2(1)) accommodates about 20 sodium ions and 12 water molecules in the vicinity of phosphate groups. These structured guest molecules provide an intricate network of bridges, ranging in size from a single sodium ion to a multiple sodium-water-water-sodium unit, connecting phosphate groups belonging to adjacent DNA helices. The crystallographic R value for this structure is 0.23 for a total of 102 reflections extending out to 3.2 A resolution. PMID- 15299772 TI - Direct-space methods in phase extension and phase determination. III. Phase refinement using Sayre's equation. AB - An algorithm is described for refining a set of phases to agree with the Sayre equation. All operations are carried out using Fourier transforms with modest computer-store requirements even for very large systems. The procedure is tested with two moderate-sized proteins, one containing heavy atoms, and is found to give good refinement with data at more than atomic resolution (1.17 A) and useful, if less good, refinement when the data resolution is lower (1.5 A). It is concluded that at atomic resolution, or slightly below, the Sayre equation still has something to offer both for phase refinement and phase extension, especially if used cautiously with weighted multiple isomorphous replacement phases acting as a constraint on the phase changes. Even when the Sayre equation on its own refines phases badly, or not at all, it may still make an important contribution in conjunction with other real-space refinement procedures. PMID- 15299773 TI - Refined structures at 2 and 2.2 A resolution of two forms of the H-protein, a lipoamide-containing protein of the glycine decarboxylase complex. AB - H-protein, a 14 kDa lipoic acid-containing protein is a component of the glycine decarboxylase complex. This complex which consists of four protein components (P , H-, T- and L-protein) catalyzes the oxidative decarboxylation of glycine. The mechanistic heart of the complex is provided by the lipoic acid attached to a lysine residue of the H-protein. It undergoes a cycle of transformations, i.e. reductive methylamination, methylamine transfer, and electron transfer. We present details of the crystal structures of the H-protein, in its two forms, H Pro(Ox) with oxidized lipoamide and H-Pro(Met) with methylamine-loaded lipoamide. X-ray diffraction data were collected from crystals of H-Pro(Ox) to 2 and H Pro(Met) to 2.2 A resolution. The final R-factor value for the H-Pro(Ox) is 18.5% for data with F > 2sigma. in the range of 8.0-2.0 A resolution. The refinement confirmed our previous model, refined to 2.6 A, of a beta-fold sandwich structure with two beta-sheets. The lipoamide arm attached to Lys63, located in the loop of a hairpin conformation, is clearly visible at the surface of the protein. The H Pro(Met) has been crystallized in orthorhombic and monoclinic forms and the structures were solved by molecular replacement, starting from the H-Pro(Ox) model. The orthorhombic structure has been refined with a final R-factor value of 18.5% for data with F > 2sigma in the range of 8.0-2.2 A resolution. The structure of the monoclinic form has been refined with a final R-factor value of 17.5% for data with F > 2sigma in the range of 15.0-3.0 A. In these two structures which have similar packing, the protein conformation is identical to the conformation found in the H-Pro(Ox). The main change lies in the position of the lipoamide group which has moved significantly when loaded with methylamine. In this case the methylamine-lipoamide group is tucked into a cleft at the surface of the protein where it is stabilized by hydrogen bonds and hydrophobic contacts. Thus, it is totally protected and not free to move in aqueous solvent. In addition, the H-protein presents some sequence and structural analogies with other lipoate- and biotin-containing proteins and also with proteins of the phosphoenolpyruvate:sugar phosphotransferase system. PMID- 15299774 TI - Structure of a new azurin from the denitrifying bacterium Alcaligenes xylosoxidans at high resolution. AB - It has been reported previously that Alcaligenes xylosoxidans (NC1MB 11015) grown under denitrifying conditions produces two azurins instead of the single previously identified azurin [Dodd, Hasnain, Hunter, Abraham, Debenham, Kanzler, Eldridge, Eady, Ambler & Smith (1995). Biochemistry. In the press]. The new azurin, called azurin II, has been crystallized as blue elongated rectangular prisms with the tetragonal space group P4(1)22 and unit-cell parameters a = b = 52.65, c = 100.63 A. X-ray crystallographic data extending to 1.9 A resolution were collected by the Weissenberg method using 200 x 400 mm image plates and synchrotron X-rays of wavelength 0.97 A. The three-dimensional structure of azurin II has been solved by the molecular-replacement method using the structure of azurin from Alcaligenes denitrificans NCTC 8582 with which this new azurin shows a close homology. The quality of the initial map was sufficient to predict a number of sequence differences. The model is currently refined to an R-factor of 18.8% with X-ray data between 8.5 and 1.9 A. The final model of 961 protein atoms, one Cu atom and 50 water molecules has r.m.s. deviations from ideality of 0.009 A for bond lengths and 1.7 degrees for bond angles. The overall structure is similar to that of the azurin from A. denitrificans NCTC 8582. It has a beta barrel structure with the Cu atom located near the top end of the molecule. The Cu atom is coordinated to Ndelta of His46 and His117 at 2.02 A and to Sgamma of Cys112 at 2.12 A, while the carbonyl O atom of Gly45 and Sdelta atom of Met121 provide the additional interactions at 2.75 and 3.26 A, respectively. PMID- 15299775 TI - Crystallization and preliminary X-ray diffraction analysis of double-helical RNA octamers. AB - Single crystals of a chemically synthesized self-complementary RNA octamer with sequence r(CCCCGGGG) have been obtained by screening 50 different conditions at room temperature using a standard sparse-matrix sampling method. Two crystal forms with different morphologies grew under diverse crystallization conditions within days by hanging-drop vapor diffusion. Hexagonal crystals with space group P6(1)22 (one strand per asymmetric unit) and unit-cell dimensions a = b = 39.73 and c = 58.55 A, diffracted to 2.6 A. Rhombohedral crystals with space group R32 (one duplex per asymmetric unit) and unit-cell dimensions a = b = 42.38 and c = 131.70 A, (hexagonal setting) diffracted beyond 1.5 A. Data sets for both crystal forms were collected on image-plate/rotating-anode generator equipment and structure determinations and refinements are under way. PMID- 15299776 TI - Crystallization and preliminary crystallographic investigations of cytochrome c4 from Pseudomonas stutzeri. AB - Cytochrome c(4) from the bacterium Pseudomonas stutzeri has been crystallized and X-ray diffraction data have been collected to 2.2 A resolution. The crystals belong to the monoclinic system, space group P2(1) with cell parameters a = 49.49, b = 58.58, c = 63.51 A and beta = 96.96 degrees. The crystals contain two molecules it, the asymmetric unit. Cytochrome c(4) is known to contain two covalently bound haem groups per molecule and positions of the four haem Fe atoms in the asymmetric unit were determined from native anomalous-dispersion differences. The shortest Fe-Fe distances found in the crystal were 15.8, 16.9 and 19.0 A. PMID- 15299777 TI - Crystallization and preliminary X-ray crystallographic data with Escherichia coli transketolase. AB - The Escherichia coli enzyme transketolase, a dimeric protein of 2 x 70 kDa (662 amino acids) has been prepared from an overexpression system in E. coli. The purified enzyme has been crystallized from PIPES buffer pH 6.4 and ammonium sulfate. The crystals which grow as large plates diffract to greater than 1.9 A, resolution and are of the space group P2(1)2(1)2(1) with unit-cell dimensions of a = 74.6, b = 125.6 and c = 151.0 A, (Z = 8 with one transketolase dimer in the asymmetric unit). The structure has been solved by molecular replacement using the yeast transketolase enzyme structure as a search model. The enzyme is being used for large-scale biotransformations using various aldehydes and hydroxypyruvate as substrates. PMID- 15299778 TI - Properties of a crystal of the complex of methyl D-arabinofuranoside with concanavalin A. AB - The complex of methyl alpha-D-arabinofuranoside with concanavalin A crystallizes in the orthorhombic space group P2(1)22(1) with cell dimensions a = 97.5, b = 87.0 and c = 61.5 A. The asymmetric unit contains one dimer and the unit cell consists of two tetrahedral clusters of point-group symmetry 222. The crystals diffract to 2.0 A resolution. PMID- 15299779 TI - Preliminary X-ray studies on two new crystal forms of staphylococcal enterotoxin C2. AB - Two new crystal forms of staphylococcal enterotoxin C2 have been obtained by vapor-diffusion methods. Form 1 crystals are monoclinic in space group P2(1) with cell dimensions a = 43.43, b = 69.92, c = 42.22 A, 8 = 90.1 degrees and diffract to at least 2.7 A resolution. Form 2 crystals are tetragonal in space group P4(1)2(1)2 or P4(3)2(1)2 with cell dimensions a = b = 42.98, c = 289.92 A and diffract to 1.9 A resolution. PMID- 15299780 TI - Crystallization and preliminary characterization of human recombinant N acetylgalactosamine-4-sulfatase. AB - Crystals of human recombinant N-acetylgalactosamine-4-sulfatase have been grown using vapour diffusion. The protein contains approximately 13%(w/w) carbohydrate. The crystals belong to the tetragonal space group P4(1)2(1)2 or its enantiomorph P4(3)2(1)2 with a = b = 108.0 and c = 145.5 A. The crystals diffract to 2.7 A resolution. PMID- 15299781 TI - Preliminary X-ray diffraction study of a new crystal form of C-1027-AG, the apoprotein of the macromolecular antitumor antibiotic C-1027 from Streptomyces globisporus. AB - A new crystal form of C-1027-AG, the apoprotein of the macromolecular antitumor antibiotic C-1027 isolated from Streptomyces globisporus was obtained by the vapor-diffusion procedure using lithium sulfate as a precipitant. In the present crystallization, it is noteworthy that large-sized single crystals successfully grew from very small droplets (less than 1.01 micro l). The present crystals belong to the trigonal system, space group P3(1)21 or P3(2)21 with cell dimensions of a = b = 62.6 and c = 54.2 A. Assuming that the asymmetric unit contains one molecule, the V(m) value is calculated as 2.9 A(3) Da(-1). A total of 3654 independent reflections from two native crystals was obtained up to 2.5 A resolution with synchrotron radiation, the merging R factor being 0.097. PMID- 15299782 TI - Crystallization and preliminary X-ray crystallographic analysis of DNA polymerase from Thermus aquaticus. AB - Two crystal forms of DNA polymerase from Thermus aquaticus have been grown at room temperature. Rhombohedral crystals (form I) grown from ammonium sulfate solution diffracted poorly to 10 A only and thus are not suitable for X-ray structure determination. Trigonal crystals (form II) grown from polyethylene glycol solution are more suitable for structure determination since their diffraction pattern extends to 2.5 A at cryogenic temperature upon exposure to synchrotron X-rays. They belong to space group P3(1)21 (or its enantiomorph P3(2)21) and their unit-cell dimensions are a = 106.7 and c = 169.7 A, for flash frozen crystals. The presence of one molecule per asymmetric unit gives a crystal volume per protein mass (V(M)) of 3.0 A(3) Da(-l) and a solvent content of 58% by volume. X-ray data have been collected to 2.7 A Bragg spacing from native crystals. PMID- 15299783 TI - Crystallization and preliminary X-ray analysis of pyruvate kinase type I from Escherichia coli. AB - Crystals of the fructose-l,6-bisphosphate-dependent pyruvate kinase from Escherichia coli have been grown using the hanging-drop vapour-diffusion technique. The space group was found to be C222(1) with cell dimensions a = 76.8, b = 247.5, c = 132.6 A. Diffraction data to 3.0 A resolution have been recorded and the enzyme molecular symmetry analysed through inspection of the self rotation function. The crystallized protein is in the allosterically inactive T state. PMID- 15299784 TI - Preliminary X-ray crystallographic study of methyltetrahydrofolate: corrinoid/iron sulfur protein methyltransferase from Clostridium thermoaceticum. AB - Methyltetrahydrofolate:corrinoid/iron sulfur protein methyltransferase from Clostridium thermoaceticum has been crystallized in two polymorphic forms and characterized by X-ray diffraction measurements. Form I displayed orthorhombic symmetry with a = 63.9, b = 53.8, c = 164.0 A. Form II also displayed orthorhombic symmetry with a = 63.5, b = 87.1, c = 117.9 A. Crystals of form I diffract to approximately. 3 A resolution; those of form II diffract to approximately 2.7 A. PMID- 15299785 TI - Purification, crystallization and preliminary X-ray crystallographic analysis of lactoperoxidase from buffalo milk. AB - The lactoperoxidase was prepared from buffalo milk and purified using CM-Sephadex C-50 and Sephadex G-100. The activity of the enzyme was measured using 2,2'-azino bis(3-ethylbenzthiazoline-6-sulfonic acid) diammonium salt as a chromogenic substrate at pH 6.0. The purified protein was crystallized from 0.01 M sodium phosphate buffer (pH 8.0) with 10%(v/v) ethanol by the sitting-drop vapour diffusion method. The green-coloured plate-like crystals are orthorhombic in space group P2(1)2(1)2(1) with unit-cell dimensions a = 116.9, b = 103.2 and c = 62.3 A. The asymmetric unit contains one molecule with a solvent content of 52%. The crystals were stable in the X-ray beam and diffract beyond 3.2 A. The native data to 3.5 A have been collected and the structure determination is in progress. PMID- 15299786 TI - X-ray analysis of crystals of polygalacturonase A from Pseudomonas solanacearum. AB - Crystals of the pectolytic protein, polygalacturonase A, have been obtained from polyethylene glycol 8000 using vapor diffusion methods. The 52.4 kDa protein is secreted by the plant pathogenic bacteria Pseudomonas solanacearum, and is important in the virulence of this plant pathogen. The protein crystallizes in space group P2(1) and has unit-cell parameters of a = 101.9, b = 124.6, c = 48.1 A, and beta= 105 degrees 50'. The crystal has two molecules in the asymmetric unit, and diffracts maximally to a resolution of 2.1 A. PMID- 15299787 TI - Improvements in lysozyme protein crystal perfection through microgravity growth. AB - Microgravity offers an environment for protein crystallization where there is an absence of convection and sedimentation. We have investigated the effect of microgravity conditions on the perfection of protein crystals. The quality of crystals for X-ray diffraction studies is characterized by a number of factors, namely size, mosaicity and the resolution limit. By using tetragonal lysozyme crystals as a test case we show, with crystal growth in two separate Space Shuttle missions, that the mosaicity is improved by a factor of three to four over earth-grown ground control values. These microgravity-grown protein crystals are then essentially perfect diffraction gratings. As a result the peak to background of individual X-ray diffraction reflections is enhanced by a similar factor to the reduction in the mosaicity. This then offers a particularly important opportunity for improving the measurement of weak reflections such as occur at high diffraction resolution. These microgravity results set a benchmark for all future microgravity and earth-based protein crystallography procedures. PMID- 15299790 TI - Difference refinement: obtaining differences between two related structures. AB - There are many examples in macromolecular crystallography where interest focuses on the differences between a previously determined 'native' structure and a nearly isomorphous 'variant'. In such cases, a useful approach to atomic refinement of the variant structure is through weighted least-squares minimization of the residual between the observed and calculated differences in amplitudes of structure factors, a strategy first used in the refinement of deoxycobalt hemoglobin [Fermi, Perutz, Dickinson & Chien (1982). J. Mol. Biol. 155, 495-505] and termed 'difference refinement'. For cases in which the modeling errors for the native and variant structures are correlated, theoretical arguments indicate that difference refinement should lead to improved estimates of structural differences when compared with conventional independent refinement. Tests employing simulated peptide data sets and real data from a wild-type protein and a mutant show that difference refinement can substantially reduce errors in the differences between structures when compared with independent refinement. The algorithm is very easy to implement and does not increase the computational demands of refinement. PMID- 15299791 TI - Structure of a glutathionylated human lysozyme: a folding intermediate mimic in the formation of a disulfide bond. AB - The three-dimensional structure of a mutant human lysozyme, C77A-a, in which the residue Cys77 is replaced by alanine, has been refined to an R value of 0.125 using 8230 reflections in the resolution range 10.0-1.8 A. It has been shown that C77A-a, in which the counterpart of Cys77 (Cys95) is modified with glutathione, has been shown to mimic an intermediate in the formation of the disulfide bond Cys77-Cys95 during the folding of human lysozyme [Hayano, Inaka, Otsu, Taniyama, Miki, Matsushima & Kikuchi (1993). FEBS Lett. 328, 203-208]. An earlier structure demonstrates that its overall structure is essentially identical to that of the wild-type protein and served as the starting model. The refined model includes atoms for all protein residues (1-130), 20 glutathione atoms and 113 water atoms. Further refinement shows more clearly the details of the protein, the bound glutathione molecule and solvent structure. However, the main-chain folding and the atomic thermal factors of the loop region from Thr70 to Leu79 were highly affected by the binding of the glutathione molecule, as compared with those of the wild-type protein. The bound glutathione shifted the main-chain atoms from Va174 to Ala77 by more than 6.0 A, and the temperature factors of the atoms in the loop region were quite high (more than 40 A(2)), indicating that the backbone conformation of this region is highly flexible and that the loop region is not folded in the specific conformation observed in the wild-type protein. These results strongly suggest that the loop structure in human lysozyme is folded later than the other regions of the protein in vivo, as observed in in vitro folding. Since the bound glutathione is efficiently and irreversibly dissociated by protein disulfide isomerase, the glutathione molecule may act as a protecting group to prevent the formation of an incorrect disulfide bond in the protein folding process in vivo. PMID- 15299792 TI - On the application of phase relationships to complex structures. XXXV. Some experiments with 2-Zn insulin. AB - The direct-methods program SAYTAN has been applied to the known structure of 2-Zn insulin with 806 atoms, excluding solvent, in the asymmetric unit. Useful sets of phases can be obtained and selected by figures of merit for data resolutions between 1.5 and 2.25 A and these can be extended by SAYTAN to give mean phase errors of 68 degrees for more than 2000 reflections. A feature of the phases so found is that the phase errors decrease with increasing resolution - which is the opposite of the situation when phases are found by isomorphous-replacement techniques. PMID- 15299793 TI - Structure of human diferric lactoferrin refined at 2.2 A resolution. AB - The three-dimensional structure of the diferric form of human lactoferrin has been refined at 2.2 A resolution, using synchrotron data combined with a lower resolution (3.2 A) diffractometer data set. Following restrained least-squares refinement and model rebuilding the final model comprises 5330 protein atoms (691 residues), 2Fe(3+) and 2CO(3)(2-) ions, 469 solvent molecules and 98 carbohydrate atoms (eight sugar residues). Root-mean-square deviations from standard geometry are 0.015 A for bond lengths and 0.038 A for angle (1-3) distances, and the final crystallographic R-factor is 0.179 for all 39 113 reflections in the resolution range 8.0-2.2 A. A close structural similarity is seen between the two lobes of the molecule, with differences mainly in loops and turns. The two binding sites are extremely similar, the only apparent differences being a slightly more asymmetric bidentate binding of the carbonate ion to the metal, and a slightly longer Fe-O bond to one of the Tyr ligands, in the N-lobe site relative to the C lobe site. Distinct differences are seen in the interactions made by two cationic groups, Arg210 and Lys546, behind the iron site, and these may influence the stability of the two metal sites. Analysis of interdomain and interlobe interactions shows that these are few in number which is consistent with the known flexibility of the molecule with respect to domain and lobe movements. Internal water molecules are found in discrete sites and in two large clusters (in the two interdomain clefts) and one tightly bound water molecule is present 3.8 A from the Fe atom in each lobe. The carbohydrate is weakly defined and has been modelled to a limited extent; two sugar residues of the N-lobe glycan and six of the C-lobe glycan. Only one direct protein-carbohydrate contact can be found. PMID- 15299794 TI - Three-dimensional structure of a hemichrome hemoglobin from Caudina arenicola. AB - The structure of a monomeric hemichrome form of an invertebrate hemoglobin, Hb-C chain, from Caudina arenicola has been refined to an R value of 0.16 using the data from 5.0 to 2.5 A resolution (R = 0.21 from 10.0 to 2.5 A resolution). Hb-C crystallizes in space group P2(l) with cell constants a = 45.74, b = 45.23 and c = 40.92 A and beta = 104.4 degrees with two monomers packed in the unit cell (V(m) = 2.34 A(3) Da(-1)). The phases were determined by the multiple isomorphous replacement method with Hg(2+) the major derivative. The structure consists of 157 amino acids with N- and C-terminal regions and eight alpha-helices forming a heme pocket. The unique feature of this structure is the hemichrome form with the proximal and distal histidines coordinated to the heme Fe atom, which is nearly in the plane of the porphyrin ring. A total of 111 solvent molecules were added to the structure using difference density peaks of at least 3sigma over background. Interestingly, all the heme groups present in the crystal are nearly coplanar. PMID- 15299795 TI - Structure of hexagonal turkey egg-white lysozyme at 1.65A resolution. AB - The structure of hexagonal turkey egg-white lysozyme (TEWL) has been determined and refined at 1.65 A resolution. The crystals were grown from a 150 mM potassium thiocyanate solution at pH 4.5 and belong to space group P6(1)22 with unit-cell dimensions a = b = 70.96, c = 83.01 A alpha = beta = 90, gamma = 120 degrees. The crystals were isomorphous with those of hexagonal pH 8.0 TEWL. The coordinates of PDB entry code 3LZ2 were therefore used as the initial model and subjected to rigid-body refinement, simulated annealing and least-squares refinement to a final residual of 0.20. The root-mean-square deviations from the ideal bond distances and angles were 0.016 A and 2.2 degrees, respectively. During the refinement, 86 water molecules and one thiocyanate ion were located in the structure. The thiocyanate ion lies close to the interface between two symmetry related molecules. The S atom of the ion forms two direct intermolecular contacts with Argl4 and interacts indirectly via a network of water molecules to Arg5 of a symmetry-related molecule. The structure provides direct evidence for the mode of thiocyanate binding to arginine residues and suggests a possible mechanism for the efficiency of thiocyanate in crystallizing basic proteins. PMID- 15299796 TI - Full-matrix refinement of the protein crambin at 0.83 A and 130 K. AB - This paper describes the first successful full-matrix least-squares (FMLS) refinement of a protein structure. The example used is crambin which is a small hydrophobic protein (4.7 kDa, 46 residues). It proves the feasibility of refining such large molecules by this classic method, routinely applied to small molecules. The final structure with 381 non-H protein atoms (54 protein atoms in alternative positions), 367 H atoms, 162 water molecules (combined occupancy 93) and one disordered ethanol molecule converged to a standard unweighted crystallographic R-factor of R = 9.0% when refined against F with reflections stronger than F > 2sigma(F) and R = 9.5% when refined against F(2). The programs RFINE [Finger & Prince (1975). Natl Bur. Stand. (US) Tech. Note 854. A System of Fortran IV Computer Programs for Crystal Structure Computations] and SHELXL93 [Sheldrick (1993). SHELXL93. Program for Crystal Structure Refinement, Univ. of Gottingen, Germany] were used for FMLS refinement with the high-resolution low temperature (0.83 A, 130 K) data set of a mixed-sequence form of crambin. A detailed analysis of the models obtained in FMLS and PROLSQ [restrained least squares or RLS; Teeter, Roe & Heo (1993). J. Mol. Biol. 230, 292-311] refinements with the same data set is presented. The differences between the models obtained by both FMLS and RLS refinements are systematic but negligible and advantages and shortcomings of both methods are discussed. The final structure has very good geometry, fully comparable to the geometry of other structures in this resolution range. Ideal values used in PROLSQ and those by Engh & Huber [Engh & Huber (1991). Acta Cryst. A47, 392-400] differ significantly from this refinement and we recommend a new standard. FMLS refinement constitutes a sensitive tool to detect and model disorder in highly refined protein structures. We describe the modeling of temperature factors by the TLS method [Schomaker & Trueblood (1968). Acta Cryst. B24, 63-76]. Rigid body-TLS refinements led to a better understanding of different modes of vibrations of the molecule. Refinements using F(2) or F protocols converged and reached slightly different minima. Despite theoretical support for F(2)-based refinement, we recommend refinement on structure factors. PMID- 15299797 TI - TEM1 beta-lactamase structure solved by molecular replacement and refined structure of the S235A mutant. AB - beta-Lactamases are bacterial enzymes which catalyse the hydrolysis of the beta lactam ring of penicillins, cephalosporins and related compounds, thus inactivating these antibiotics. The crystal structure of the TEM1 beta-lactamase has been determined at 1.9 A resolution by the molecular-replacement method, using the atomic coordinates of two homologous beta-lactamase refined structures which show about 36% strict identity in their amino-acid sequences and 1.96 A r.m.s. deviation between equivalent Calpha atoms. The TEM1 enzyme crystallizes in space group P2(1)2(1)2(1) and there is one molecule per asymmetric unit. The structure was refined by simulated annealing to an R-factor of 15.6% for 15 086 reflections with I >/= 2sigma(I) in the resolution range 5.0-1.9 A. The final crystallographic structure contains 263 amino-acid residues, one sulfate anion in the catalytic cleft and 135 water molecules per asymmetric unit. The folding is very similar to that of the other known class A beta-lactamases. It consists of two domains, the first is formed by a five-stranded beta-sheet covered by three alpha-helices on one face and one alpha-helix on the other, the second domain contains mainly alpha-helices. The catalytic cleft is located at the interface between the two domains. We also report the crystallographic study of the TEM S235A mutant. This mutation of an active-site residue specifically decreases the acylation rate of cephalosporins. This TEM S235A mutant crystallizes under the same conditions as the wild-type protein and its structure was refined at 2.0 A resolution with an R value of 17.6%. The major modification is the appearance of a water molecule near the mutated residue, which is incompatible with the OG 235 present in the wild-type enzyme, and causes very small perturbations in the interaction network in the active site. PMID- 15299798 TI - Fast ab initio calculation of solvent envelopes for protein structures. AB - A fast and simple method has been developed for the ab initio calculation of low resolution solvent envelopes for macromolecular structures. In essence, a sphere of point scatterers is moved through the asymmetric unit cell in a part random, part systematic search for the configuration which corresponds to the lowest R value. The spheres correspond to the solvent regions in the cell. The program has been shown to work successfully for a number of test structures in a variety of space groups. No prior knowledge of the structures is needed, and c.p.u. requirements are extremely modest. PMID- 15299799 TI - Water-dependent domain motion and flexibility in ribonuclease A and the invariant features in its hydration shell. An X-ray study of two low-humidity crystal forms of the enzyme. AB - The crystal structures of 88 and 79% relative humidity forms of ribonuclease A, resulting from water-mediated transformations, have been refined employing the restrained least-squares method using X-ray data collected on an area detector to R = 0.173 for 15 326 observed reflections in the 10-1.5 A resolution shell and R = 0.176 for 8534 observed reflections in the 10-1.8 A shell, respectively. The comparison of these structures with those of the native, the phosphate-bound and the sulfate-bound forms demonstrates that the mobility of the ribonuclease A molecule involves hinge-bending movement of the two domains and local flexibility within them, particularly at the termini of regular secondary structures and in loops. The comparison also leads to the identification of 31 invariant water molecules in the hydration shell of the enzyme, many of which are involved in holding different parts of the molecule together and in stabilizing local structure. The conformational changes that accompany the partial removal of the surrounding water, particularly those observed in the 79% form, could be similar to those that occur during enzyme action. PMID- 15299800 TI - Structure of the azurin mutant nickel-Trp48Met from Pseudomonas aeruginosa at 2.2 A resolution. AB - The structure of the azurin mutant nickel-Trp48Met from Pseudomonas aeruginosa has been determined by difference Fourier synthesis using phases from the wild type azurin model. The final crystallographic R value is 0.170 for 17 394 reflections to a resolution of 2.2 A. The mutant crystallized in the orthorhombic space group P2(1)2(1)2(1), a = 57.4, b = 80.4, c = 110.3 A. The four molecules in the asymmetric unit are packed as a dimer of dimers. The nickel metal site of this mutant structure is similar to the zinc metal site in the azurin Asp47 mutant. The site-specific mutation was performed at residue Trp48, which is located in the center of the protein in a highly hydrophobic environment, to investigate its suggested role in the long-range electron-transfer pathway between the disulfide bond on one side of the protein to the Cu centre. The structure around the mutation site Met48 showed no significant change compared with the wild-type structure. PMID- 15299801 TI - X-ray structure of turkey egg lysozyme complex with di-N-acetyl-chitobiose. Recognition and binding of alpha-anomeric form. AB - The crystalline complex of turkey-egg lysozyme (TEL) with di-N-acetylchitobiose (NAG2) was prepared by a soaking method and the structure was determined by X-ray analysis at 1.55 A resolution. The structure was refined to an R value of 0.175 by simulated annealing and energy minimization. The alpha-anomer of NAG2 is located at subsite D with the orientation perpendicular to the direction of the active-site cleft. The anomeric residue is deeply inserted into the cleft and the O1-H hydroxyl group is hydrogen bonded to the carboxyl group of Glu35 which is a catalytic residue. The other sugar residue protrudes outside the cleft and is in van der Waals contact with the beta-sheet region comprising of residues 43-53. The binding of NAG2 makes the active-site cleft 0.3-0.5 A narrower and suppresses the thermal motion of two lobes constructing the cleft. The NAG2 molecule is bound in a manner not assumed in the catalytic action of the enzyme and the geometry of binding indicates that the alpha-anomer blocks the active center and acts only as an inhibitor. PMID- 15299802 TI - Structure of anionic salmon trypsin in a second crystal form. AB - Anionic salmon trypsin in a second crystal form (ST-IIB) has been refined at 1.83 A, resolution. The crystals are orthorhombic and belong to space group P2(1)2(1)2 with lattice parameters a = 77.09, b = 82.33 and c = 31.16 A. The present structure has been compared to salmon trypsin as it appears in a previously reported crystal form (ST-IIA) with cell dimensions a = 61.95, b = 84.33 and c = 39.11 A [Smalas & Hordvik (1993). Acta Cryst. D49, 318-330]. The presence of a sulfate group involved in several hydrogen bonds to active-site residues, and the location of an additional benzamidine site in the crystal lattice, are the most striking differences between the present and the previous structure. Superposition of main-chain atoms in the two structures give an overall r.m.s. difference of 0.26 A, with the main differences located to areas with different molecular packing. The overall coordinate error is estimated to be between 0.20 and 0.25 A, by the method of Luzzati. PMID- 15299803 TI - Expression, characterization and crystallographic analysis of telluromethionyl dihydrofolate reductase. AB - Selenomethionine-containing proteins analyzed by multi-wavelength anomalous diffraction provide a facile means of addressing the phase problem, whose solution is necessary to determine protein structures by X-ray crystallography [Hendrickson (1991). Science, 254, 51-58]. Since this method requires synchrotron radiation, we sought to incorporate a true heavy atom into protein, allowing the solution of the phase problem by more traditional methods of data collection. Media containing TeMet alone or TeMet with low levels of Met failed to sustain growth of a methione auxotroph of Escherichia coli carrying the dihydrofolate reductase expression vector. Growth of the organism to stationary phase and incorporation of TeMet was observed when the culture was initiated in media containing minimal Met levels and TeMet was added after induction with isopropyl 1-thio-beta-D-galactopyranoside. The purified enzyme exhibited properties similar to those of the native enzyme. Atomic absorption spectroscopy and amino-acid analysis indicated that 40% of the methionines were replaced with TeMet. Sequence analysis did not indicate significant levels of replacement in the first three sites (1, 16 and 20), suggesting that TeMet was present only in the last two sites (42 and 92). Crystals of this enzyme were grown in the presence of methotrexate and were isomorphous with crystals of wild-type dihydrofolate reductase. Difference Fourier maps and restrained least-squares refinement showed no substitution at the first three methionines, while incorporation was seen at positions 42 and 92. PMID- 15299804 TI - The direct rotation function: Patterson correlation search applied to molecular replacement. AB - Most rotation functions try to achieve maximal correlation between two Patterson functions by systematically rotating one and computing the overlap with the other. In contrast, the direct rotation function rotates a search model relative to the crystal unit cell and evaluates the linear correlation coefficient (Patterson correlation, PC) between squared normalized structure-factor amplitudes of the observed and calculated diffraction data. Structure factors are calculated from the rotated search model in a P1 unit cell identical to that of the target crystal. PC makes use of all self-Patterson vectors of the search model. A comparison of the direct rotation function, a real-space rotation function, and a fast rotation function suggests that the direct rotation function provides a considerable enhancement of the signal-to-noise ratio compared to other two. Combined with PC refinement, the direct rotation function was successful in solving multidomain macromolecular crystal structures. PMID- 15299805 TI - Phase refinement and extension by means of non-crystallographic symmetry averaging using parallel computers. AB - Electron-density averaging, fast Fourier synthesis and fast Fourier analysis programs have been adapted for parallel-computing systems. These have been linked to perform iterative phase improvement and extension utilizing non crystallographic symmetry and solvent flattening. Various strategies for parallel algorithms have been tested on a variety of computers as a function of the number of computer nodes. Some experimental timing results are discussed. PMID- 15299806 TI - X-ray structure determination of a dimeric hemoglobin from Caudina arenicola. AB - The X-ray structure of a dimeric, cyanomet-liganded hemoglobin D-chain (Hb-D) from Caudina arenicola has been determined by the molecular-replacement method. The search model was a concatenated model of three hemoglobin structures superimposed on the backbone of monomeric, hemichrome hemoglobin C-chain (Hb-C) from the same organism. Hb-D crystallizes in space group P4(1)2(1)2 with cell constants a = b = 77.0 and c =61.5 A with one subunit in the asymmetric unit. The dimer twofold axis corresponds to a crystallographic twofold along one of the body diagonals of the unit cell. Rotation and translation searches as well as model refinement were carried out in X-PLOR with the final model having an R value of 0.19 using the data from 5.0 to 2.9 A resolution (R = 0.26 for 10.0 to 2.9 A resolution). The homodimeric structure of Caudina Hb-D features close heme heme contacts with an Fe-Fe distance of 19.0 A. The subunit-subunit interface involves both the E and F helices from each subunit with the E helices oriented antiparallel at 50 degrees with respect to one another, similar to the quaternary structure observed for the homodimeric hemoglobin from Scapharca inaequivalvis. PMID- 15299807 TI - Structure of the crystalline complex of deoxycytidylyl-3',5'-guanosine (3',5' dCpdG) cocrystallized with ribonuclease at 1.9 A resolution. AB - The X-ray structure of bovine ribonuclease A cocrystallized with the dinucleotide deoxycytidylyl-3',5'-guanosine has been determined at 1.9 A resolution and refined by restrained least squares to R = 0.218 for 7807 reflections. The structure established that the recently observed retrobound mode of attachment of substrate analogues cytidylyl-2',5'-guanosine and deoxycytidylyl-3',5'-guanosine found in soaked RNase A crystals is also present in the cocrystallized complex. Retrobinding is thus unlikely to be the result of restrictions imposed by the crystalline environment as the ligands soak into the lattice but rather a phenomenon specific to small nucleotides containing guanine. PMID- 15299808 TI - Non-ideality of aqueous solutions of polyethylene glycol: consequences for its use as a macromolecular crystallizing agent in vapor-diffusion experiments. AB - Microisopiestic measurements of the concentrations of polyethylene glycol (PEG 8000) paired with the salts sodium chloride, ammonium sulfate and magnesium sulfate heptahydrate have been made in a sitting-drop arrangement with PEG in the droplet and salt in the reservoir. Resulting graphs of the concentrations of PEG and salt that are equivalent with respect to the vapor pressure of water are non linear, do not intersect their origins, and demonstrate that relatively low (mM) salt concentrations are equivalent to relatively high PEG concentrations. The consequences of each of these observations for macromolecular crystallization by the vapor-diffusion technique when PEG is employed as the crystallizing agent are discussed. PMID- 15299809 TI - Chaperone salts, polyethylene glycol and rates of equilibration in vapor diffusion crystallization. AB - The kinetics of water-vapor equilibration in macromolecular crystallization were investigated for sitting droplets of aqueous polyethylene glycol (PEG) 8000 as a function of concentration. Equilibrations, set up with initial concentrations of PEG in the droplet at half those in the reservoir, were very slow for concentrations of relevance to the macromolecular crystal growth problem. At 301 K, 24 micro l droplets at initial concentrations of 2.5, 5.0 and 7.5%(w/v) PEG require 12, 5, and 3 weeks to reach equilibrium, respectively. On the other hand, the addition of modest quantities of sodium chloride to both droplet and reservoir increases the rate of equilibration for aqueous PEG sitting droplets significantly. At 293 K, droplets with initial volumes of 24 micro l and PEG concentrations of 5%(w/v) require 12 weeks to reach equilibrium, while droplets of the same volume and initial concentrations of 5%(w/v) PEG and 200 mM NaCI require less than two weeks to reach equilibrium. The slow vapor-diffusion equilibrations of pure PEG solutions, and the subsequent increase in these rates with colligative agents such as salt, are a consequence of the non-ideality of aqueous PEG solutions. These results are of interest both from a practical and a theoretical viewpoint. They underscore the importance of kinetic factors in macromolecular crystal growth, help to explain apparent inconsistencies of outcome in PEG-mediated crystallizations, and yield another methodology for the optimization of crystal growth conditions, namely the control of the kinetics of equilibration using colligative agents. PMID- 15299810 TI - Rate of water equilibration in vapor-diffusion crystallization: dependence on the residual pressure of air in the vapor space. AB - The kinetics of water equilibration in vapor-diffusion crystallization experiments are sensitive to the residual pressure of air in the vapor chamber. Experiments with sitting droplets of 10%(w/v) PEG, allowed to equilibrate with reservoirs of 20%(w/v) PEG, were conducted at pressures ranging from 80 to 760 mm Hg. Equilibrations were interrupted after one, four, five and seven days to assess their progress. Even down to the lowest pressures examined it was found that a decrease in pressure leads to an increase in the rate of equilibration. The residual pressure of air in the vapor chamber can be varied to tailor the time course of equilibration in macromolecular crystal growth experiments. PMID- 15299811 TI - Pf1 filamentous bacteriophage: refinement of a molecular model by simulated annealing using 3.3 A resolution X-ray fibre diffraction data. AB - The filamentous bacteriophage Pf1 is structurally similar to the well known Ff (fd, fl, M13) strains, but it gives much better X-ray diffraction patterns, enabling a more detailed analysis of the molecular structure. The 46-residue protein subunit can be closely approximated by a single gently curved stretch of alpha-helix. The axes of the subunits are at a small angle to the virion axis, and several thousand subunits form an overlapping inter-digitated helical array surrounding a DNA core. We have derived a detailed model of the virion based on X ray data and stereochemical constraints. We have considered potential sources of error in the diffraction data, and used the improved data to study regions where the protein subunit of Pf1 may deviate from a continuous alpha-helix. We use simulated annealing to escape from local minima, and various kinds of electron density maps to guide the model building. Refinement of the model shows that the first few residues at the N terminus are non-helical, and there is a slight discontinuity in the alpha-helix near the middle of the sequence. The model is consistent both with general structural principles derived from high-resolution analysis of other proteins, and with specific chemical and spectroscopic data about Pf1. We apply the same refinement techniques to an alternative model with a non-helical surface loop between residues 13 and 19. Comparative analysis of models with and without a loop shows that the loop model is not supported by 3.3 A resolution X-ray diffraction data. PMID- 15299812 TI - Refined structure of Cu-substituted alcohol dehydrogenase at 2.1 A resolution. AB - Liver alcohol dehydrogenase (LADH) is a Zn(II)-dependent dimeric enzyme. LADH with the active-site Zn(II) substituted by Cu(II) resembles blue (type I) copper proteins by its spectroscopic characteristics. In this work we present the X-ray structure of the active site Cu(II)-substituted LADH complex with NADH and dimethyl sulfoxide (DMSO). The structure was solved by molecular replacement. The space group is P2(1) with cell dimensions a = 44.4, b = 180.6, c = 50.8 A and beta = 108 degrees. There is one dimer of the enzyme in the asymmetric unit. The refinement was carried out to a crystallographic R-factor of 16.1% for 41 119 unique reflections in the resolution range 12.0 to 2.1 A. The coordination geometry of Cu(II) in LADH is compared with the active-site metal coordination in the Zn-LADH-NADH-DMSO complex and blue-copper proteins. The distances from the metal to the protein ligands (Cys46, His67 and Cys174) are similar for the Zn(II) and Cu(II) ions. The distances of the O atom of the inhibitor DMSO to the Cu(II) ion in the two subunits of the dimer are 3.19 and 3.45 A. These are considerably longer than the corresponding distances for the Zn(II) enzyme, 2.19 and 2.15 A. The Cu(II) ion is positioned nearly in the plane of the three protein ligands (NS(2)) with a geometry similar to the trigonal arrangement of the three strongly bound ligands (N(2)S) in blue-copper proteins. This coordination probably accounts for the similarity of the spectral characteristics of Cu(II)-LADH and type I copper proteins. PMID- 15299813 TI - Phasing with mercury at 1 A wavelength. AB - Synchrotron sources provide a continuously tunable X-ray beam which makes it possible to optimize the anomalous contribution to phase determination using heavy-atom replacement. This method was used to solve two protein structures, those of Dictyostelium discoideum nucleoside diphosphate kinase and of lobster enolase. The first had 17 kDa of protein in the asymmetric unit, the second, 47 kDa. In both cases, a single mercury derivative yielded single isomorphous replacement with anomalous-scattering phases from which an interpretable electron density map was derived by solvent flattening. The efficient solution of the X ray structure was largely due to the large anomalous scattering of mercury at a wavelength shorter than the L(III) absorption edge. PMID- 15299814 TI - Determination of hemihedral twinning and initial structural analysis of crystals of the procarboxypeptidase A ternary complex. AB - The initial structural analysis of the ternary complex of procarboxypeptidase A from hemihedrally twinned crystals diffracting up to 2.8 A is described. Detection of twinning by different techniques is presented, including biochemical and intensity statistics approaches. The structure was initially solved using Patterson-search techniques, and the three positioned search models were used to effectively deconvolute the twinned data. PMID- 15299815 TI - Crystallization and preliminary X-ray diffraction analysis of glutathione peroxidase from human plasma. AB - The extracellular form of glutathione peroxidase from human plasma has been crystallized by the sitting-drop vapour-diffusion method. The crystals belong to tetragonal space group I4(1) with cell dimensions of a = b = 83.1 and c = 131.0 A. They diffract beyond 2.9 A resolution and have one dimer in the asymmetric unit. A self-rotation function analysis shows the possible (222) symmetry for the tetramers of this enzyme. PMID- 15299816 TI - Crystallization and preliminary X-ray crystallographic analysis of carbamoyl phosphate synthetase from Escherichia coli. AB - Carbamoyl Phosphate synthetase catalyzes the formation of carbamoyl phosphate, a high-energy intermediate used in several biosynthetic pathways. The enzyme from Escherichia coli has been crystallized at pH 8 in the presence of L-ornithine, MnCl(2) and ADP, using PEG 8000 in combination with NEt(4)Cl and KCl. The crystals (apparently) belong to the orthorhombic space group P2(1)2(1)2(1) with unit-cell dimensions of a = 154.4, b = 166.5 and c = 338.7 A. The crystals are relatively sensitive to radiation damage, but show diffraction to beyond 2.8 A resolution. A low-resolution (3.5 A) native data set has been recorded and conditions for flash cooling the crystal have been established. PMID- 15299817 TI - Crystallization of glycerol dehydrogenase from Bacillus stearothermophilus. AB - The NAD(+)-linked glycerol dehydrogenase from Bacillus stearothermophilus is a member of the so-called 'iron- containing' class of polyol dehydrogenases. This enzyme has been crystallized in three different forms in the presence of a range of divalent cations and glycerol or NAD+ using the hanging-drop method of vapour diffusion with ammonium sulfate as the precipitant. X-ray photographs have established that the crystals grown in the presence of zinc and glycerol (form A) most likely belong to space group I4(1)22 with cell parameters a = b = 102 and c = 728 A. The crystals grown with zlnc and NAD(+) (form B) belong to the tetragonal system and probably belong to the space group P42(1)2 with cell parameters a = b = 150 and c = 220 A. The crystals grown with lead and glycerol (form C) belong to a primitive orthorhombic system with cell parameters a = 127, b = 178 and c = 173 A. Experiments using the synchrotron radiation source at the DRAL Daresbury laboratory have shown all three crystal types diffract to at least 3 A resolution. Elucidation of the three-dimensional structure of this enzyme will provide a structural framework for this class of polyol dehydrogenases, which are not represented in the database at present, and enable comparisons to be made with enzymes belonging to the other classes. PMID- 15299818 TI - Additional crystal forms of the E. coli class II fructose-1,6-bisphosphate aldolase. AB - We have obtained two additional crystal forms of the metal-dependent class II fructose-1,6-bisphosphate aldolase from Escherichia coli. Crystals in the shape of elongated plates have unit-cell dimensions a = 73.4, b = 120.0, c = 190.1 A, orthorhombic space group P2(1)2(1)2(1). Monoclinic prisms have unit-cell dimensions a = 67.7, b = 104.3, c = 52.8 A, beta = 105 degrees, space group P2(1). Diffraction to slightly better than 3.0 A, has been observed for both forms using in-house and synchrotron facilities. These crystal forms may aid the structure solution of this enzyme by presenting additional forms for heavy-atom derivatization. These forms have multiple copies of the enzyme in the asymmetric unit and averaging methods might also be useful in the analysis. PMID- 15299819 TI - Crystallization and preliminary X-ray crystallographic studies of glutamic acid specific proteinase from Bacillus licheniformis complex with Z-Leu-Glu-CH2Cl. AB - A glutamic acid specific proteinase from Bacillus licheniformis has been crystallized as a complex with the inhibitor Z-Leu-Glu-CH(2)Cl. Crystals were grown by the vapor-diffusion method using sodium formate as a precipitant. The crystals diffracted to about 2.0 A resolution and belonged to the trigonal space group P3(1)21 (P3(2)21) with unit-cell parameters a = b = 134.3, c = 109.7 A. A total of 26 964 independent reflections were obtained up to 2.2 A resolution, the merging R-factor being 0.05 for 42 614 measurements. PMID- 15299820 TI - Crystallization and preliminary X-ray analysis of the NADP-specific glutamate dehydrogenase from Neurospora crassa. AB - The NADP-linked glutamate dehydrogenase from Neurospora crassa has been crystallized by the hanging-drop method of vapour diffusion in the presence of 0.1 M glutamate. The crystals are trigonal and are in space group P3(1)21 with unit-cell dimensions of a = b = 196.6, c = 102.0 A and with a trimer in the asymmetric unit. A full structure determination of this enzyme will lead to an understanding of the molecular basis of inter-allelic complementation observed with hybrid hexamers of naturally occurring mutants. PMID- 15299821 TI - Crystallization and preliminary X-ray diffraction studies of arginase from a thermophilic organism Bacillus caldevelox. AB - A thermostable hexameric arginase purified from the extreme thermophile Bacillus caldevelox has been crystallized from Hepes buffer at pH 7.5 in the presence of 12% polyethylene glycol 4000 and 10% 2-propanol, and from cacodylate buffer at pH 7.2 in the presence of 15% 2-propanol and sodium citrate. The latter crystals are more suitable for X-ray diffraction analysis. The crystals are in the orthorhombic space group P2(1)2(1)2(1) with unit-cell dimensions a = 156.3, b = 148.0 and c = 85.4 A. The asymmetric unit contains one hexamer (approximate molecular mass 183 kDa) and has a solvent content of approximately 54%. The crystals diffract to 2.8 A resolution. PMID- 15299822 TI - Preliminary crystallographic analysis of chloroperoxidase from Caldariomyces fumago. AB - Chloroperoxidase from the fungus Caldariomyces fumago has been crystallized in two space groups, C222(1) and P2(1)2(1)2(1) both of which are suitable for high resolution X-ray studies. Parent data sets have been obtained to 2.16 A in space group C222(1) and 2.00 A in space group P2(1)2(1)2(1). Heavy-atom derivatives have been obtained with both forms and electron-density maps calculated. The heme has been located and continuous electron density between the heme and protein clearly indicates the location of the proximal ligand. PMID- 15299823 TI - Crystallization and analysis of the subunit assembly and quaternary structure of imidazoleglycerol phosphate dehydratase from Saccharomyces cerevisiae. AB - Imidazoleglycerol phosphate dehydratase (IGPD) from Saccharomyces cerevisiae has been crystallized in the presence of a range of divalent cations using the hanging-drop method of vapour diffusion with ammonium sulfate or polyethylene glycol (PEG) 4000 as the precipitants. X-ray precession photographs have established that the crystals formed with ammonium sulfate (form A) belong to the space group F432, with cell parameter a = 177.5 A and a single subunit in the asymmetric unit. A preliminary data set collected to 6 A resolution on a two detector San Diego Multiwire area detector has established that the crystals formed with PEG 4000 (form B) belong to either of the special pair of space groups I23 or I2(1)3, with cell parameter a = 131.0 A. A self-rotation function has been calculated using these data and indicates that the cell axes show pseudo fourfold symmetry consistent with a dimer in the asymmetric unit in this crystal form. Light-scattering studies indicate that in the presence of Mn(2+) and a number of other divalent cations IGPD undergoes assembly to a particle of molecular weight approximately 500 kDa. Given the subunit molecular weight of 23 kDa together with the symmetry of the crystals it would indicate that the most likely quaternary structure for this enzyme is based on a 24-mer in 432 symmetry. PMID- 15299824 TI - Crystallization and preliminary X-ray analysis of maize ZBP14 protein, a member of a new family of zinc-binding proteins. AB - A preliminary X-ray crystallographic study of a novel zinc-binding protein from maize is presented. Native and several heavy-atom derivative sets of data have been collected on synchrotron sources, to a resolution of 2.1 A. The space group was found to be orthorhombic, P2(1)2(1)2(1), with unit-cell dimensions of a = 92.64, b = 129.45 and c = 196.31 A. PMID- 15299825 TI - Crystallization and preliminary X-ray analysis of two density populations of feline calicivirus. AB - Two density populations of full-size feline calicivirus (FCV), the intact infectious particles (P(H)) and the empty capsids (P(L)), have been crystallized using the hanging-drop method. Exposed to high-intensity synchrotron radiation, P(H) and P(L) crystals were shown to diffract X-rays to about 3.0 and 5.5 A resolution, respectively. The P(H) crystal belongs to an orthorhombic crystal system with unit-cell dimensions a = 889.0, b = 995.0, c = 436.6 A. Based on the V(M) value (3.4 A(3) Da(-l)), it was estimated that one crystallographic asymmetric unit of P(H) crystals contains the unique content of an entire virus particle, not necessarily from the same particle. This implies the presence of 60 fold non-crystallographic redundancy. The particle orientation was obtained from a locked rotation function. PMID- 15299826 TI - A modified Crowther and Blow T1 translation function for partial search models. AB - A simple modification to the fast T(1) translation function of Crowther & Blow [Crowther & Blow (1967). Acta Cryst. 23, 544-548] can reduce systematic error in cases where the input set of oriented search models represents a fraction of the scattering matter of the unknown crystal unit cell. The observed Patterson function is modified by a partial self-vector subtraction (SVS) and then the residual Patterson origin is removed. In a test case where a protein/DNA complex crystal was searched using a protein-only model, the highest signal- to-noise ratios for translation-search maps were obtained either with origin removal alone or partial SVS combined with origin removal. Origin removal is likely to be a generally useful alternative to SVS for the function. PMID- 15299827 TI - Crystallization and preliminary X-ray crystallographic analysis of chitosanase from Bacillus circulans MH-K1. AB - Chitosanase, an enzyme which hydrolyzes chitosan, isolated from Bacillus circulans MH-K1, was crystallized by a vapor-diffusion procedure at 293 K using ammonium sulfate as a precipitant. Rod-shaped colorless crystals, which grew to 0.05 x 0.15 x 1.2 mm within a week, belong to the orthorhombic system and the space group P222(1) or P2(1)2(1)2 with unit-cell dimensions of a = 43.3, b = 57.7, and c = 128.0 A. The asymmetric unit is thought to contain one chitosanase molecule (29 024 Da). The crystals diffract X-rays to at least 2.3 A resolution and are suitable for high-resolution X-ray structure analysis. PMID- 15299829 TI - Small-molecule crystal structures as a structural basis for drug design. AB - For a long time, the crystal structures of small molecules were regarded as useful only for establishing the stereochemical formulae of the crystallized compounds. Recently, chemists have realized that in the study of the environment in the solid state there exists valuable structural information on the binding characteristics of chemical groups. Numerous comparisons have been made which show the nearly perfect correlation between small-molecule structural results, and the observed binding in receptor-substrate complexes. Moreover, the observed conformations of flexible substrates interacting with the neighbouring molecules in their crystal structures, can lead to valuable hypotheses on their conformation when bound in the active site of a biological macromolecule. PMID- 15299830 TI - Intermolecular interactions around functional groups in crystals: data for modeling the binding of drugs to biological macromolecules. AB - When a small biologically active molecule enters the body it has the possibility of interacting with one or many of the large variety of macromolecules that are present. This interaction involves a unique complementary fit between the two that depends not only on the shape, but also on the distribution of charges on the surfaces of both [Fischer (1894). Ber. Dtsch. Chem. Ges. 27, 2984-993]. This recognition between two molecules may elicit a biological response and such processes are the subject of biochemical investigations. The question asked here is: what geometrical information on distances and relative orientations of interacting non-bonded functional groups can be found from X-ray crystallographic investigations? PMID- 15299831 TI - Zinc-directed inhibitors for zinc proteinases. AB - Zinc proteinases have been recognized as a distinct class of proteolytic enzymes in which at least one ion of zinc is involved directly in catalysis. This family includes a growing number of biologically important enzymes which are attractive targets for rational drug design. In this paper we examine the special features of the zinc binding environment of these enzymes in order to gain information which could be useful in the preparation of 'zinc-directed' selective inhibitors. Carboxypeptidase A (CPA) is presented as a model for one class of zinc proteinases, and the active-site zinc and its interactions are examined with the primary focus on geometrical considerations. The three-dimensional structure of the native and apoenzyme are discussed, together with the high-resolution structure of several enzyme-inhibitor complexes. This paper will first present a structural analysis of CPA derivatives and then discuss a series of zinc model compounds which have been prepared and characterized in order to examine the ligand and geometrical preferences of the zinc in an unstrained system. X-ray crystallography (macromolecular and small molecule) is the main experimental method used for the structural analyses, while complementary computational methods have been used for the examination of electrostatic potentials. The results from the various experimental efforts are assembled in order to draw general conclusions on the potential use of the zinc ion as the primary target for inhibitor binding. The results of these studies suggest that the zinc ion is important for both the binding and the catalytic activation of the substrate as well as for stabilization of the tetrahedral reaction intermediate. PMID- 15299832 TI - Mastering the LORE of protein structure. AB - A detailed description of the design, operation and capabilities of LORE, a protein-database management tool to supplement more traditional protein map fitting and model-building programs, is presented. The program includes elements for searching the library of known crystal structures for substructures of similar geometry. Substructures may be as simple as a single hairpin turn, or as complicated as an assembly of different elements of secondary structure. The programs also include elements for manipulating structural segments in complex ways to enable a sophisticated molecular editing capability of enormous utility in modeling and structure-refinement applications. PMID- 15299833 TI - Glucose analogue inhibitors of glycogen phosphorylase: from crystallographic analysis to drug prediction using GRID force-field and GOLPE variable selection. AB - Several inhibitors of the large regulatory enzyme glycogen phosphorylase (GP) have been studied in crystallographic and kinetic experiments. GP catalyses the first step in the phosphorylysis of glycogen to glucose-l-phosphate, which is utilized via glycolysis to provide energy to sustain muscle contraction and in the liver is converted to glucose. alpha-D-Glucose is a weak inhibitor of glycogen phosphorylase form b (GPb, K(i) = 1.7 mM) and acts as a physiological regulator of hepatic glycogen metabolism. Glucose binds to phosphorylase at the catalytic site and results in a conformational change that stabilizes the inactive T state of the enzyme, promoting the action of protein phosphatase 1 and stimulating glycogen synthase. It has been suggested that in the liver, glucose analogues with greater affinity for glycogen phosphorylase may result in a more effective regulatory agent. Several N-acetyl glucopyranosylamine derivatives have been synthesized and tested in a series of crystallographic and kinetic binding studies with GPb. The structural results of the bound enzyme-ligand complexes have been analysed together with the resulting affinities in an effort to understand and exploit the molecular interactions that might give rise to a better inhibitor. Comparison of the N-methylacetyl glucopyranosylamine (N methylamide, K(i) = 0.032 mM) with the analogous beta-methylamide derivative (C methylamide, K(i) = 0.16 mM) illustrate the importance of forming good hydrogen bonds and obtaining complementarity of van der Waals interactions. These studies also have shown that the binding modes can be unpredictable but may be rationalized with the benefit of structural data and that a buried and mixed polar/non-polar catalytic site poses problems for the systematic addition of functional groups. Together with previous studies of glucose analogue inhibitors of GPb, this work forms the basis of a training set suitable for three dimensional quantitative structure-activity relationship studies. The molecules in the training set are void of problems and potential errors arising from the alignment and bound conformations of each of the ligands since the coordinates were those determined experimentally from the X-ray crystallographic refined ligand-enzyme complexes. The computational procedure described in this work involves the use of the program GRID to describe the molecular structures and the progam GOLPE to obtain the partial least squares regression model with the highest prediction ability. The GRID/GOLPE procedure performed using 51 glucose analogue inhibitors of GPb has good overall predictivity [standard deviation of error predictions (SDEP) = 0.98 and Q(2) = 0.76] and has shown good agreement with the crystallographic and kinetic results by reliably selecting regions that are known to affect the binding affinity. PMID- 15299834 TI - Binding of the antiviral drug WIN51711 to the sabin strain of type 3 poliovirus: structural comparison with drug binding in rhinovirus 14. AB - The crystal structure of the Sabin strain of type 3 poliovirus (P3/Sabin) complexed with the antiviral drug WIN51711 has been determined at 2.9 A resolution. Drugs of this kind are known to inhibit the uncoating of the virus during infection, by stabilizing the capsid against receptor-induced conformational changes. The electron density for the bound drug is very well defined so that its position and orientation are unambiguous. The drug binds in a nearly extended conformation, slightly bent in the middle, in a blind pocket formed predominantly by hydrophobic residues in the core of the beta-barrel of capsid protein VP1. Comparisons between this structure, the corresponding drug complex in human rhinovirus 14 (HRV 14), and the native structures of both viruses demonstrate that the binding of WIN51711 has markedly different effects on the structures of these two viruses. Unlike HRV14, wherein large conformational changes are observed in the coat protein after drug binding, the binding of this drug in poliovirus does not induce any significant conformational changes in the structure of the capsid protein, though the drug has a greater inhibitory effect in P3/Sabin than in HRV14. The implications of this result for the mechanism of capsid stabilization are discussed. PMID- 15299835 TI - WIN51711-resistant mutants of poliovirus type 3: capsid residues important in uncoating functions. AB - Capsid-binding drugs that inhibit the first stage of picornaviral uncoating were used to select drug-resistant mutants of the Sabin strain of poliovirus type 3. Such mutants provide information about parts of the capsid that are important for functions blocked by the drugs, and also about pathways to drug resistance. Amino acid substitutions allowing virus to produce progeny in the presence of drug were mapped to 13 different residues occupying three distinct locations: (I) the canyon base; (II) the lining of the drug-binding pocket; and (III) the base of the protomer. These loci might be thought of as action points for transmitting the uncoating signal from receptor, through the pocket, and to the base of the protomer. All of the mutations in a special class of drug-dependent mutants were clustered at site (III) and all were hyperlabile, i.e., uncoated spontaneously (without receptor) at growth temperature unless prevented from doing so by the presence of drug in the pocket. Thus, site (III) seems to represent a kind of thermostat which regulates the temperature at which the uncoating transition (release of VP4 to form A particles) is triggered. PMID- 15299836 TI - Structures of four methyltetrazole-containing antiviral compounds in human rhinovirus serotype 14. AB - Four novel antiviral WIN compounds, that contain a methyl tetrazole ring as well as isoxazole, pyridazine or acetylfuran rings, have had their structures determined in human rhinovirus serotype 14 at 2.9 A resolution. These compounds bind in the VP1 hydrophobic pocket, but are shifted significantly towards the pocket pore when compared to previously examined WIN compounds. A putative water network at the pocket pore is positioned to hydrogen bond with these four WIN compounds, and this network can account for potency differences seen in structurally similar WIN compounds. PMID- 15299837 TI - Benzoic acid inhibitors of influenza virus neuraminidase. AB - A strategy was developed to design non-carbohydrate inhibitors of influenza virus neuraminidase. Using an iterative cycle of modeling, synthesis, biological testing and X-ray crystallography structure determination, a series of inhibitors based on benzoic acid were produced. The refined structures of three compounds complexed with neuraminidase are reported. The results demonstrate the success of this structure-based drug-design strategy. PMID- 15299838 TI - Comparative X-ray structures of the major binding protein for the immunosuppressant FK506 (tacrolimus) in unliganded form and in complex with FK506 and rapamycin. AB - FK506 (tacrolimus) is a natural product now approved in the US and Japan for organ transplantation. FK506, in complex with its 12 kDa cytosolic receptor (FKBP12), is a potent agonist of immunosuppression through the inhibition of the phosphatase activity of calcineurin. Rapamycin (sirolimus), which is itself an immunosuppressant by a different mechanism, completes with FK506 for binding to FKBP12 and thereby acts as an antagonist of calcineurin inhibition. We have solved the X-ray structure of unliganded FKBP12 and of FKBP12 in complex with FK506 and with rapamycin; these structures show localized differences in conformation and mobility in those regions of the protein that are known, by site directed mutagenesis, to be involved in calcineurin inhibition. A comparison of 16 additional X-ray structures of FKBP12 in complex with FKBP12-binding ligands, where those structures were determined from different crystal forms with distinct packing arrangements, lends significance to the observed structural variability and suggests that it represents an intrinsic functional characteristic of the protein. Similar differences have been observed for FKBP12 before, but were considered artifacts of crystal-packing interactions. We suggest that immunosuppressive ligands express their differential effects in part by modulating the conformation of FKBP12, in agreement with mutagenesis experiments on the protein, and not simply through differences in the ligand structures themselves. PMID- 15299839 TI - Design, synthesis and structure of non-macrocyclic inhibitors of FKBP12, the major binding protein for the immunosuppressant FK506. AB - We have synthesized a series of non-macrocyclic ligands to FKBP12 that are comparable in binding potency and peptidyl prolyl isomerase (PPIase) inhibition to FK506 itself. We have also solved the structure of one of these ligands in complex with FKBP12, and have compared that structure to the FK506-FKBP12 complex. Consistent with the observed inhibitory equipotency of these compounds, we observe a strong similarity in the conformation of the two ligands in the region of the protein that mediates PPIase activity. Our compounds, however, are not immunosuppressive. In the FKBP12-FK506 complex, a significant portion of the FK506 ligand, its 'effector domain', projects beyond the envelope of the binding protein in a manner that is suggestive of a potential interaction with a second protein, the calcium-dependent phosphatase, calcineurin, whose inhibition by the FKBP 12-FK506 complex interrupts the T-cell activation events leading to immunosuppression. In contrast, our compounds bind within the surface envelope of FKBP12, and induce significant changes in the structure of the FKBP12 protein which may also affect calcineurin binding indirectly. PMID- 15299840 TI - Structure-based design of inhibitors of purine nucleoside phosphorylase. AB - Inhibitors of purine nucleoside phosphorylase may have therapeutic value in the treatment of T-cell proliferative diseases such as T-cell leukemia, in the suppression of host-versus-graft response in organ transplants, and in the treatment of T-cell-mediated autoimmune diseases. Competitive inhibitors of this enzyme have been designed using the three-dimensional structure of the enzyme determined by X-ray crystallography. This approach has resulted in the synthesis of the most potent and membrane-permeable inhibitors of purine nucleoside phosphorylase reported so far. PMID- 15299841 TI - Calculation of relative binding affinities of purine nucleoside phosphorylase inhibitors. AB - The competitive binding of inhibitors to purine nucleoside phosphorylase (PNP) has been experimentally measured. Fast and reliable computational methods to estimate binding would allow assessment of any proposed inhibitor before its synthesis. Binding-energy calculations with a representative set of PNP inhibitors were compared to the empirical values. Relatively simple and fast calculations were executed with X-PLOR, DelPhi and SoftDock. The computational results are mixed. PMID- 15299842 TI - Molecular dynamics and structure-based drug design for predicting non-natural nonapeptide binding to a class I MHC protein. AB - Starting from the known three-dimensional structure of the class I major histocompatibility complex-encoded HLA-B*2705 protein, three non-natural nonapeptides were designed to fit optimally the HLA-B*2705-binding groove. The optimization was performed using structure-based drug design methods and the fact that all the possible interactions of the secondary anchor residue (position 3) with its human leukocyte antigen-binding pocket (pocket D) in nature are not entirely utilized. 150 ps molecular-dynamics (MD) simulation in water was employed to study the stability of the bimolecular complexes with three non natural peptides (P3 = homophenylalanine, beta-naphthylalanine, alpha naphthylalanine) as well as with the two natural homologues (P3 = Gly, Leu). Various structural and dynamical properties (atomic fluctuations, solvent accessible surface areas, peptide Calpha-atom positions) of the simulated bimolecular complexes were used to compare the three non-natural with the two natural ligands. Since the various molecular properties have been shown previously to be related to the binding affinity of nonapeptide ligands to the major histocompatibility complex (MHC) HLA-B*2705 protein, the MD data predict a rather higher stability of MHC-ligand complexes with the three non-natural peptides, suggesting that the unnatural peptides studied show an enhanced binding affinity to the HLA-B*2705 protein. These results are in agreement with the experimental values of a semi-quantitative in vitro assembly assay, performed on the five nonapeptides (P3 = Gly, Leu, homophenylalanine, beta-naphthylalanine, alpha-naphthylalanine), which shows their ability to stabilize the native conformation of the HLA-B*2705 heavy chain and also shows that the three non natural ligands bind with higher affinity (0.5 micro M) to the MHC protein than the two natural homologues (40 micro M). Thus, this study demonstrates that structural information combined with rational design and molecular-dynamics simulations can illustrate and predict MHC binding and potential T-cell epitope properties as well as contribute to the design of new non-peptidic MHC inhibitors that may be useful for the selective immunotherapy of autoimmune diseases to which HLA alleles are directly associated. PMID- 15299843 TI - Active-site mimetic inhibition of thrombin. AB - The structures of two mimetic inhibitor complexes of human alpha-thrombin have been determined by X-ray crystallography. One mimics a beta-turn with a bicyclic ring system; the other mimics two different active-site binding modes. The beta turn mimetic is used to approximate a turn found in the conformation of fibrinopeptide A, which is catalytically released by thrombin in the activation of fibrinogen to fibrin. The binding of the second mimetic is a hybrid between normal substrate and the abnormal binding of the potent natural leech inhibitor hirudin. The binding of the beta-turn mimetic is tenuous, because it is like a substrate, while that of the substrate-hirudin hybrid is that of a tenacious inhibitor (which it is). Structurally retrospect modifications for rational design and improvement of both mimetic inhibitors are proposed. PMID- 15299844 TI - Correlation of binding affinities with non-bonded interaction energies of thrombin-inhibitor complexes. AB - Several empirical modeling protocols are evaluated allowing a quantification of the interaction between an enzyme and a series of inhibitors. The evaluation and optimization of the modeling protocols used a database of 35 non-covalently bound inhibitors of human thrombin. Intermolecular interaction energies were calculated with CHARMm after energy minimization of the modeled complexes using various dielectric functions and constraining strategies. These calculated binding energies were correlated with the experimentally obtained binding constants of the inhibitors. The best protocols resulted in linear correlations with correlation coefficients > 0.80. In the best protocols the enzyme was fully constrained, the ligand was allowed to move freely and electrostatics were scaled down drastically or fully neglected during the energy minimization. For the interaction energy evaluation step, a distance-dependent dielectric function epsilon = R proved to be optimal. This simple empirical protocol, that neglects solvation or entropy effects, can be implemented readily in other force field packages and may be applied on various enzyme-inhibitor complexes, providing a tool for the evaluation and rank-ordering of newly designed inhibitors. PMID- 15299845 TI - A comparison of two independently determined structures of trypanothione reductase from Crithidia fasciculata. AB - The enzyme trypanothione reductase (TR) is unique to trypanosomes and leishmania parasites, the causal agents of several important medical and veterinary tropical diseases. TR helps regulate the intracellular reducing environment of the parasite and it has been identified as a target for developing novel chemotherapeutic agents by structure-aided drug design. For this purpose it is essential to have confidence in the structural detail of the molecular target. Two independent studies of Crithidia fasciculata TR at medium resolution, in different space groups have afforded an opportunity to assess the reliability of the models. We summarize the important methodological details of each analysis and present a comparison of the geometry, thermal parameters and three dimensional structure of the models. Particular attention has been paid to the disulfide substrate-binding site which is the area of most interest with respect to enzyme inhibition. The comparison has shown that the structures agree closely with Calpha atoms superposing with an r.m.s. of less than 0.5 A. The consistency of the models gives a high level of confidence that they are suitable for computer-aided drug design. The conformation of many side chains in the active site, in particular the catalytic residues, are well conserved in both structures. However, the comparison indicates a difference in the conformation of Trp21 and Met113 which together form a hydrophobic patch on the rim of the active site cleft and interact with the spermidine moiety of the substrate. Consideration of the electron-density maps together with the structural comparison indicates that there is some conformational flexibility in this region of the active site. This heterogeneity may be used in the recognition of the substrate by the enzyme and should be considered when mapping out the size, shape and chemical properties of the active site. PMID- 15299846 TI - Refined 3.2 A structure of glycosomal holo glyceraldehyde phosphate dehydrogenase from Trypanosoma brucei brucei. AB - The three-dimensional crystal structure of the enzyme glyceraldehyde phosphate dehydrogenase from the kinetoplastid Trypanosoma brucei brucei has been determined at 3.2 A resolution from a 37% complete data set collected using the Laue method. The crystals used in the structure determination contain one and a half tetrameric enzyme molecules in the asymmetric unit, i.e. six identical subunits. Initial phasing was carried out by the method of molecular replacement using the refined coordinates of holo glyceraldehyde phosphate dehydrogenase from Bacillus stearothermophilus as a search model. The initial electron-density distribution, obtained from the molecular-replacement solution, was greatly improved by a procedure consisting of 36 cycles of iterative non-crystallographic density averaging. During the averaging procedure, the missing reflections (63% of the data) were gradually introduced as map-inversion structure factors. At completion of the procedure, the R-factor between averaged map-inversion amplitudes and observed structure-factor amplitudes was 19.0% for all data between 7.0 and 3.2 A resolution, and that between the map-inversion amplitudes and later recorded structure-factor amplitudes was 41.9%. After model building into the resulting averaged electron-density map, refinement by molecular dynamics procedures with X-PLOR provided the current model, which has an R-factor of 17.6% for 34 835 reflections between 7.0 and 3.2 A resolution. The refined model, comprising 2735 protein atoms plus one NAD(+) molecule and two sulfate ions per subunit, has r.m.s. deviations from ideality of 0.02 A for bond lengths and 3.6 degrees for bond angles. All subunits, located either within the tetrameric molecule or within the half tetramer present in the asymmetric unit, are related to each other by almost exact twofold symmetry. The overall structure of the glycosomal glyceraldehyde phosphate dehydrogenase subunit and its quaternary arrangement in the tetrameric molecule are similar to that of the enzyme of lobster and Bacillus stearothermophilus (with r.m.s. differences between equivalent Calpha positions of 0.71 and 0.64 A, respectively). The main differences between the structures is the presence of three insertions, plus the substitution of a beta-strand by a short alpha-helix, both occurring at the surface of the glycosomal enzyme subunit. PMID- 15299847 TI - Crystallographic and thermodynamic comparison of structurally diverse molecules binding to streptavidin. AB - Crystallographic structures and thermodynamic binding parameters are compared for three structural classes of streptavidin ligand including d-biotin, 2-[(4' hydroxyphenyl)-azo] benzoate and the peptide NH(2)-Phe-Ser-His-Pro-Gln-Asn-Thr COOH. Descriptions of these structural and thermodynamic observations emphasize the diversity of potential strategies for improving ligand affinity. PMID- 15299848 TI - Structure-based analysis of inhibitor binding to Ht-d. AB - A theoretical study was performed on the structure of both the native and inhibited metalloproteinase Ht-d (E.C. 3.4.24.42) solved at 2.0 A resolution. The energy maps calculated by program GRID clearly showed the extended binding site of Ht-d and allowed localization and characterization of the pockets S1-S3 and S1'-S3'. The GRID energy contour maps point out the particular shape of the S1' pocket in agreement with experimental density maps and inhibited Ht-d structures. Based on the high degree of sequence homology of the Ht-d active site to that of mammalian metalloproteinases, the characterization of active site pockets was extended to neutrophil collagenase, fibroblast collagenase, stromelysin 1 and 2. Thirty residues of the Ht-d propeptide were modeled and optimized with reference to the Ht-d structure, giving insight to the mechanism of natural inhibition in metalloproteinase proenzymes. Kinetic measurements of Ht-d inhibition by a series of synthetic peptides show, in agreement with our Ht-d propeptide model, the crucial role of cysteine and adjacent residues in the specificity of Ht-d propeptide. This study suggests the structural link between Ht-d and mammalian metalloproteinases, contributing to the understanding of the mechanism of natural and synthetic inhibitor binding to metalloproteinases. Therefore, Ht-d is a good model system for the design of novel inhibitors against these enzymes with enhanced potency and specificity. PMID- 15299849 TI - Crystallization and preliminary structure of porcine aldehyde reductase-NADPH binary complex. AB - Porcine aldehyde reductase-NADPH binary complex has been crystallized from a buffered ammonium sulfate solution. The crystal form is hexagonal, space group P6(5)22, with a = b = 67.2, c = 243.7 A, alpha = beta = 90.0 and gamma = 120.0 degrees. A molecular-replacement structure solution has been successfully obtained by using the refined structure of the apoenzyme as the search model. The crystallographic R-factor is currently equal to 0.24 after energy minimization using data between 8 and 3.0 A resolution. The aldehyde reductase-NADPH complex model is supported by electron density corresponding to NADPH not included in the search model. The tertiary structure of aldehyde reductase consists of a beta/alpha-barrel with the coenzyme-binding site located at the carboxy-terminal end of the strands of the barrel. The structure of aldehyde reductase-NADPH binary complex will help clarify the mechanism of action for this enzyme and will lead to the development of pharmacologic agents to delay or prevent diabetic complications. PMID- 15299850 TI - Determinants of backbone packing in globular proteins: an analysis of spatial neighbours. AB - This study attempts to examine the pattern and variability of backbone packing density in protein structures. A carefully selected non-redundant data set of known protein structures is analyzed in terms of amino-acid composition and the preference of individual amino acids to fall into regions of low, medium or high density depending on the number of observed non-sequence spatial neighbours. The relationship of the backbone packing density to a number of properties such as the hydrophobicity, non-bonded energies and secondary structural features of the amino acids are examined. The correlation between the average percentage composition and the percentage composition in regions corresponding to different levels of packing density of the proteins is evaluated. These studies are extended to the family of globins whose amino-acid sequences have diverged retaining the same three-dimensional structure during evolution. The significance of high-backbone-density regions in this family has become apparent as due to helix/helix packing. Further, the variation in the amino-acid composition in different contact regions of globin proteins follows the same pattern found for the general data set. PMID- 15299851 TI - Comparison of ternary complexes of Pneumocystis carinii and wild-type human dihydrofolate reductase with coenzyme NADPH and a novel classical antitumor furo[2,3-d]pyrimidine antifolate. AB - The novel furopyrimidine N-(4-{N-[(2,4-diaminofuro[2,3-d]pyrimidin-5 yl)methyl]methylamino}benzoyl)-L- glutamate (MTXO), a classical antifolate with antitumor activity comparable to that of methotrexate (MTX), has been studied as inhibitor-cofactor ternary crystal complexes with wild-type Pneumocystis carinii (pc) and recombinant human wild-type dihydrofolate reductase (hDHFR). These structural data provide the first direct comparison of the binding interactions of the same antifolate inhibitor in the active site for pc and human DHFR. The human ternary DHFR complex crystallizes in the rhombohedral space group R3 and is isomorphous to the ternary complex reported for a gamma-tetrazole methotrexate analogue, MTXT. The pcDHFR complex crystallizes in the monoclinic space group P2(1) and is isomorphous to that reported for a trimethoprim (TMP) complex. Interpretation of difference Fourier electron-density maps for these ternary complexes revealed that MTXO binds with its 2,4-diaminofuropyrimidine ring interacting with Glu32 in pc and Glu30 in human DHFR, as observed for MTXT. The presence of the 6-5 furopyrimidine ring instead of the 6-6 pteridine ring results in a different bridge conformation compared with that of MTXT. The bridge torsion angles for MTXO, i.e. C(4a)-C(5)-C(8)-N(9) and C(5)-C(8)-N(9)-C(1'), are 156.5/51.9 degrees and -162.6/51.8 degrees, respectively for h and pc, compared with -146.8/57.4 degrees for MTXT. In each case, the p-aminobenzoylglutamate conformation is similar to that observed for MTXT. In the pcDHFR complex, the active-site region is conserved and the additional 20 residues in the sequence compared with the human enzyme are located in external loop regions. There is a significant change in the nicotinamide ribose conformation of the cofactor which places the nicotinamide O atom close to the 4NH(2) group of MTXO (2.7 A), a shift not observed in hDHFR structures. As a consequence of this, there is a loss of a hydrogen bond between the nicotinamide carbonyl group and the backbone of Ala12 in pcDHFR. In the human ternary complexes, the cofactor NADPH is bound with a more extended conformation, and the nicotinamide O atom makes a 3.5 A contact with the 4NH(2) group of MTXO. Although the novel classical antifolate MTXO is not highly active against pcDHFR, there are correlations between its binding interactions consistent with its lower potency as an inhibitor of h and pcDHFR compared with MTX. PMID- 15299852 TI - X-ray structure of turkey-egg lysozyme complex with tri-N-acetylchitotriose. Lack of binding ability at subsite A. AB - The turkey-egg lysozyme (TEL) complex with tri-N-acetylchitotriose [(GlcNac)3] was co-crystallized from 1.5% TEL and 2 mM (GlcNac)3 at pH 4.2. The crystal structure was determined by molecular replacement and refined to an R value of 0.182 using 10-1.77 A data. The (GlcNac)3 molecule occupies the subsites A, B and C. At the subsites B and C, the sugar residues are bound in a similar manner to that found in the hen-egg lysozyme (HEL) complex. In contrast, the GlcNac residue at the subsite A is exposed to bulk solvent and has no contact with the protein molecule because the active residue Asp101 in HEL is replaced by Gly in TEL. A sulfate ion is bound in the vicinity of subsite B and forms hydrogen bonds with the sugar residue and the guanidino group of Arg61, assisting the binding of the sugar residue to subsite B. The active-site cleft of TEL is narrower than that of native TEL, thus attaining the best fit of the (GlcNac)3 molecule. The lack of binding ability of subsite A is discussed in relation to the catalytic properties of TEL. The result suggests that the cleavage pattern of oligosaccharide substrates in the catalytic reaction is regulated by the protein-sugar interaction at subsite A. PMID- 15299853 TI - Structure of cytochrome c' from Rhodobacter capsulatus strain St Louis: an unusual molecular association induced by bridging Zn ions. AB - Rhodobacter capsulatus strain St Louis cytochrome c' (RCCP-SL) has been crystallized and the structure solved by molecular replacement. It was refined at 2.1 A resolution to an R value of 18.4%, and compared with Rhodobacter capsulatus strain M110 cytochrome c' (RCCP-M110). Although these two proteins are very similar in sequence and structure, the intermolecular interaction is largely different. In RCCP-M110, the molecules dimerize through interaction of helix B to form an antiparallel arrangement. When crystallized in the presence of Zn ions, molecules of RCCP-SL were found to be arranged as linear polymers connected by the bridging Zn ions. The changes in conformation of the side chains induced by binding of the Zn ions, by the substitution of Glu90 for Asp90, and by the different arrangement of the molecules, are discussed in detail. PMID- 15299854 TI - Polyalanine reconstruction from Calpha positions using the program CALPHA can aid initial phasing of data by molecular replacement procedures. AB - The Calpha positions for a protein can provide a scaffold for the reconstruction of more complete models. Reconstructions can be by manual rebuilding, from geometric solutions to the constraints on main-chain torsion angles or from databases of known protein structures. The last method is usually the most convenient and reliable. This paper describes a database reconstruction program, CALPHA, and assesses its accuracy and reliability by test reconstructions of well refined structures. Typically, backbone atoms are repositioned to within 0.3 A of their original positions. This corresponds to regenerating main-chain torsion angles to within 15 degrees. Uses of CALPHA for automating refinement procedures are discussed. In particular, the uses of Calpha-only and rcconstructed polyalanine models of HIV-1 reverse transcriptase for cross-rotation and translation function searches are compared. The CALPHA polyalanine model is found to provide more selectivity for approximately correct orientations. The effect on the translation function is dependent on the resolution shell employed. It is expected that these observations will be applicable in other cases. PMID- 15299855 TI - New techniques for applying anomalous-scattering and isomorphous-replacement data incorporated in ANOMIR - a general application package. AB - A computer package ANOMIR is described which can derive phases from anomalous scattering and/or isomorphous-replacement data in any combination. For anomalous scattering it incorporates five methods of applying one-wavelength data and three methods for multiple-wavelength data including SPIN, reported here for the first time. In addition there are three procedures for multiple-wavelength data - the first modifying data for different wavelengths to make them mutually consistent, the second estimating the contributions of the anomalous scatterers alone and the third which finds anomalous differences. For single isomorphous replacement or one-wavelength anomalous scattering the phase ambiguity can be resolved by the direct method [Fan, Han, Qian & Yao (1984). Acta Cryst. A40, 489- 495] but for multiple isomorphous replacement the main method is an adaptation of the probability-curve method [Blow & Crick (1959). Acta Cryst. 12, 794-802]. A new statistical method is described for estimating the standard error in measuring magnitudes which is independent of having subsets of centric reflections. A method is described whereby the weights associated with phase estimates are used to generate probability curves, through which it is possible to combine estimates from different methods and to produce a 'best phase' and figure-of-merit for every reflection. ANOMIR procedures are also available for handling combinations of one-wavelength anomalous scattering with single- or multiple-isomorphous replacement. A final process, which is always beneficial, is a single parallel application of the tangent formula. The ANOMIR package has been designed for easy use and is controlled throughout by KEYWORDS. Results for several structures are given and compared with those found from the MLPHARE program in the CCP4 package. PMID- 15299856 TI - Experiences from the structure determination of human cytomegalovirus protease. AB - Several obstacles were encountered and overcome during the structure determination of human cytomegalovirus protease. Dehydration of crystals, by exposing them to higher concentrations of the precipitant, reduced the mosaicity of the crystals and may have also resolved their microscopic twinning. The initial phase information was obtained with the selenomethionyl multiple wavelength anomalous diffraction technique. However, site-specific mutagenesis was required to introduce extra Met residues into the protease. The phase information had to be improved by non-crystallographic symmetry averaging, initially among three 'crystal forms'. A change in the composition of the artificial mother liquor led to a significant improvement, from 3.0 and 2.0 A resolution, in the diffraction quality of the crystals. The experiences reported here may prove useful to structure determination of other proteins. PMID- 15299857 TI - Lysozyme aggregation studied by light scattering. I. Influence of concentration and nature of electrolytes. AB - Static and dynamic light scattering have been employed to investigate the behaviour of lysozyme solutions when varying the concentration of (NH(4))(2)SO(4) and NaCl for screening the repulsive forces between the monomers. At the initial aggregation stages clusters, which can be classified as mass-fractals undergoing diffusion limited-like aggregation, coexist with monomers or small lysozyme oligomers. The kinetics of fractal growth deliver observables that exhibit distinct tendencies when examined as a function of the concentration and nature of the electrolyte. The behaviour of the observables changes drastically above 0.84 M (NH(4))(2)SO(4) and 0.60 M NaCl. Static light scattering revealed a progressive restructuring of the fractals to compact structures at the latter stages of the reaction. Based on the correlations between the various observables an attempt is made to predict the long-term fate of the nucleating solutions. PMID- 15299858 TI - Lysozyme aggregation studied by light scattering. II. Variations of protein concentration. AB - Static and dynamic light scattering have been employed to investigate the behaviour of nucleating lysozyme solutions in the range between 0.34 and 3.08 mM. Preselected concentrations of NaC1 and (NH(4))(2)SO(4) have been used to screen the repulsive Coulombic interactions and trigger aggregation. Initially, mass fractals undergoing diffusion limited-like aggregation coexist with monomers or small lysozyme oligomers. The growth kinetics of the fractals deliver observables that exhibit distinct tendencies when examined as a function of lysozyme concentration. The behaviour of the observables changes drastically around 2.0 mM lysozyme. Static light scattering experiments revealed progressive restructuring or growth of compact structures at later stages of the aggregation. Based on the correlations between the observables an attempt is made to predict whether the examined solutions will crystallize or not. A tentative scheme, involving the most prominent structures observed in nucleating lysozyme solutions, is discussed. PMID- 15299859 TI - Structure of monellin refined to 2.3 a resolution in the orthorhombic crystal form. AB - The structure of orthorhombic crystals of monellin, a sweet protein extracted from African serendipity berries, has been solved by molecular replacement and refined to 2.3 A resolution. The final R factor was 0.150 for a model with excellent geometry. A monellin molecule consists of two peptides that are non covalently bound, with chain A composed of three beta-strands interconnected by loop regions and chain B composed of two beta-strands interconnected by an alpha helix. The N terminus of chain A is in close proximity to the C terminus of chain B. The two molecules in the asymmetric unit are related by a non-crystallographic twofold axis and form a dimer, similar to those previously observed in other crystal forms of both natural and single-chain monellin. The r.m.s, deviation between the Calpha atoms in the two independent molecules is 0.60 A, while the deviations from the individual molecules in the previously reported monoclinic crystals are 0.50-0.57 A. This result proves that the structure of monellin is not significantly influenced by crystal packing forces. PMID- 15299860 TI - Crystals of the carotenoid protein from Arthrospira maxima containing uniformly oriented pigment molecules. AB - Crystals of a carotenoid protein from the cyanobacterium Arthrospira maxima have been grown in space group C2 with unit-cell dimensions a = 219.6, b = 40.3, c = 75.5 A and beta = 95.5 degrees. The crystals diffract X-rays to 2.3 A resolution and display unusual optical properties in polarized light that suggest that all of the carotenoid molecules in the crystals are oriented similarly. A slight increase in the concentration of a crystallization additive in the mother liquor induces macroscopic twinning, which is also visible when the crystals are illuminated with polarized light. PMID- 15299861 TI - Comparative analysis of thaumatin crystals grown on earth and in microgravity. AB - The protein thaumatin was studied as a model macromolecule for crystallization in microgravity-environment experiments conducted on two US Space Shuttle missions (USML-2 and LMS). In this investigation, we have evaluated and compared the quality of space- and earth-grown thaumatin crystals using X-ray diffraction analyses, and characterized them according to crystal size, diffraction resolution limit and mosaicity. Two different approaches for growing thaumatin crystals in the microgravity environment, dialysis and liquid-liquid diffusion, were employed as a joint experiment by our two investigative teams. Thaumatin crystals grown in a microgravity environment were generally larger in volume and the total number of crystals was less, relative to crystals grown on earth. They diffracted to significantly higher resolution and with improved diffraction properties, as judged by relative plots of I/sigma versus resolution. The mosaicity of space-grown crystals was significantly less than that of crystals grown on earth. Increased concentrations of protein in the crystallization chambers in microgravity led to larger crystals. The data presented here lend further support to the idea that protein crystals of improved quality can be obtained in a microgravity environment. PMID- 15299862 TI - The ultimate wavelength for protein crystallography? AB - This paper describes an analysis of the optimum choice of the X-ray wavelength for macromolecular diffraction data collection. It is shown that there is no ultimate X-ray wavelength for protein crystallography and that the optimum wavelength depends to a large extent on the size of the protein crystal. It also depends on instrumental factors, such as efficiency of the detector for a particular wavelength and spectral density of the synchrotron radiation. Estimates of the optimum wavelength as a function of crystal size are given. PMID- 15299863 TI - Resolution of space-group ambiguity and structure determination of nodamura virus to 3.3 A resolution from pseudo-R32 (monoclinic) crystals. AB - Monoclinic crystals of nodamura virus (NOV) have two virus molecules per asymmetric unit. Packing analysis reveals a pseudo-rhombohedral (pseudo-C2 monoclinic) arrangement of particles in the actual P2(1) space group (a = 562.1, b = 354.1, c = 612.8 A, beta = 110.9 degrees ). The R32 symmetry is broken rotationally and translationally. The pseudo-symmetry of the unit cell results in three possible monoclinic origins and also restrains the four particles in the unit cell to similar orientations. NOV particles deviate by less than 3 degrees from the ideal orientations, causing overlap of peaks in the rotation function and the generation of peaks that were not interpretable as particle symmetry elements. The space-group ambiguity was resolved by analysing the relationship between the particle orientations determined by high-resolution rotation functions and the attenuation of peak heights in native Patterson maps. Particles were centered less than 1 A from the R32 special positions. Three different approaches were required to identify the correct particle center. Following the solutions of the rotation and translation problems, phases were computed using the coordinates of flock house virus (FHV), another member of this virus family. The phases were improved by real-space molecular averaging with a 120-fold non crystallographic symmetry and by solvent flattening with a spherical mask. The final model for the NOV structure was built using the 3.3 A averaged map. While the overall subunit structure was very similar to that of other nodaviruses, FHV and black beetle virus, NOV showed distinct structural features near particle threefold and quasi-threefold axes and at the protein-RNA interfaces that are consistent with phenotype differences among the related viruses. PMID- 15299865 TI - Probing the limits of the molecular replacement method: the case of Trypanosoma brucei phosphoglycerate kinase. AB - It is of general interest to explore the limits of the molecular replacement method. Described here are the specifics of a successful molecular replacement structure determination in a difficult case: the X-ray crystal structure of a Trypanosoma brucei phosphoglycerate kinase (PGK) ternary complex. This ternary complex crystallizes with four 45 kDa subunits in the asymmetric unit, whereas the available search models were all monomers consisting of two distinct domains. Initial molecular replacement attempts using complete subunits were unsuccessful. Attributing this failure to a presumed change in the relative orientations of the N- and C-terminal phosphoglycerate kinase domains, a second attempt was made using each domain as an independent search model. In this way, two N-terminal and then two C-terminal domains could be oriented and positioned in the unit cell. On the basis of this result, a new search model containing both domains in the correct mutual orientation was created and used to identify the two remaining phosphoglycerate kinase subunits. The ability to successfully orient and position an N-terminal domain containing 8.4% of the scattering mass in the asymmetric unit was the key to this structure determination. Further investigations show that a truncated version of this search model containing 5.8% of the scattering mass would have been sufficient for this purpose. A retrospective analysis suggests that the effectiveness of this probe is enhanced by structural conservation, retained temperature factors and a disparity in the degree of order among the various subunits in the T. brucei PGK asymmetric unit. Based on these observations, 231 well ordered Calpha atoms were selected from a single refined T. brucei PGK subunit and it was demonstrated that this collection of atoms, representing just 1.6% of the scattering mass, could be correctly oriented and positioned in the unit cell. PMID- 15299866 TI - The effects of metal binding on a nucleobase: the experimental charge density and electrostatic potential in 1H(+)-adeniniumtrichlorozinc(II) at 123 K and its relationship to that in adenine hydrochloride hemihydrate. AB - The charge-density distribution in 1H(+)-adeniniumtrichlorozinc(II) has been determined from X-ray diffraction data collected to sin theta/lambda = 1.32 A(-1) at 123 K. The electrostatic potential, isolated from the crystal lattice, and the deformation density in the nucleobase have been calculated following multipole refinements based on the rigid pseudoatom model of Stewart. These and the molecular dimensions have been compared with results from the charge-density study of adenine hydrochloride hemihydrate by Cunane & Taylor [Acta Cryst. (1993). B49, 524-530] to determine the effects of metal binding on the nucleobase. The main conclusions are that while the bond lengths and angles in the pyrimidine ring are similar, those in the imidazole ring are significantly perturbed on complexation; lone-pair electron density at N7 is observed in both structures and lies significantly off the plane of the nucleobases; the positive electrostatic potential of the complexed base extends much further from the molecule than in the uncomplexed one and the regions of negative potential at N3 and N7 are depleted in the complexed base. The observed enhancement of positive electrostatic potential in the nucleobase on binding to zinc is presented in support of a model for the mechanism of reversible unwinding of DNA in the presence of zinc ions. PMID- 15299867 TI - A new crystal form of penicillin acylase from Escherichia coli. AB - A new crystal form of penicillin acylase (penicillin amidohydrolase, E.C. 3.5.1.11) from Escherichia coli W (ATCC 11105) is reported. The crystals were grown using a combination of hanging-drop and streak-seeding methods. The crystals are in the monoclinic space group P2(1) with cell dimensions a = 51.52, b = 131.95, c = 64.43 A, beta = 106.12 degrees. There is one heterodimer in the asymmetric unit (V(m) = 2.45 A(3) Da(-1)) and the solvent content is 49%. Preliminary data have been collected to d(min) = 2.7 A using a MAR Research image plate and a rotating-anode X-ray source. Subsequent experiments show diffraction beyond 1.3 A at a synchrotron radiation source. PMID- 15299868 TI - Crystallization of an antitumour antibody SM3 complexed with a peptide epitope. AB - SM3 antibody binds to a tumour-associated epitope on polymorphic epithelial mucin (PEM). Crystals of the Fab fragment of SM3 in complex with a peptide antigen were obtained by vapour diffusion against mother liquor containing acetate buffer, pH 6.5, cadmium chloride and polyethylene glycol (PEG) 4000 as precipitating agent. Crystals belong to the monoclinic space group P2(1) with cell dimensions a = 42.2, b = 83.9, c = 64.5 A and beta = 93.4 degrees. One Fab-antigen complex is present in the asymmetric unit. Diffracted intensities up to 1.95 A resolution have been measured from a frozen crystal using synchrotron radiation. PMID- 15299869 TI - Crystallization and preliminary X-ray analysis of the Saccharomycopsis fibuligera glucoamylase expressed from the GLU1 gene in Escherichia coli. AB - The active non-glycosylated glucoamylase, overexpressed from the Saccharomycopsis fibuligera GLU1 gene in Escherichia coli BL21(DE3), has been purified from the solubilized inclusion bodies and then renatured in vitro. Crystals of the recombinant glucoamylase were obtained by vapour diffusion using PEG as precipitant. The crystals belong to the orthorhombic space group P2(1)2(1)2(1), with unit-cell dimensions of a = 58.1, b = 87.8 and c = 99.9 A, and diffract to 1.7 A resolution. This is the first report of the crystallization of the full length glucoamylase corresponding to the mature enzyme. PMID- 15299870 TI - Crystallization and preliminary X-ray diffraction studies of the Saccharomyces cerevisiae phospholipid-transfer protein Sec14p. AB - The Saccharomyces cerevisiae phosphatidylinositol-transfer protein Secl4p catalyzes the exchange of phosphatidylinositol or phosphatidylcholine between membrane bilayers in vitro, and is an essential protein required for the budding of secretory vesicles from the yeast Golgi complex in vivo. At issue is the fundamental question of how the dual phospholipid ligand specificity of Sec 14p translates to in vivo function. In an attempt to determine the structural basis for how Secl4p binds each of its phopholipid ligands, Secl4p occupied with phosphatidylcholine has been purified and the complex crystallized in the presence of the mild detergent n-octyl beta-D-glucopyranoside. The Secl4p crystals diffract to 2.7 A and belong to space group P3(1)21 or P3(2)21 with unit cell dimensions of a = b = 88.79, c = 111.21 A, alpha = beta = 90, gamma = 120 degrees. As Secl4p exhibits significant primary sequence homology to mammalian retinaldehyde binding proteins and the noncatalytic domain of human MEG2 protein tyrosine phosphatase, is is anticipated that solution of the Secl4p crystal structure will provide new functional insights for a family of interesting proteins. PMID- 15299871 TI - Crystallization and preliminary crystallographic studies on the mannose-specific lectin from garlic. AB - A mannose-specific agglutinin from garlic (Allium sativum) which forms part of a well conserved super-family of bulb lectins has been purified and crystallized by the hanging-drop vapour-diffusion technique, by equilibrating with a 20% solution of PEG 8000 in the presence of alpha-D-mannose. Crystals of the dimeric form of this protein are monoclinic C2 with unit-cell dimensions a = 203.2, b = 43.8, c = 79.3 A and beta = 112.4 degrees and have two dimers in the asymmetric unit. Data have been collected to 2.4 A resolution and the structure solved by molecular replacement using the coordinates of the snowdrop lectin as the search model. PMID- 15299872 TI - A weighted rotation function. AB - A simple probability expression is derived to discriminate the intermolecular vectors in the cross rotation function. The radial weighting function introduced takes into account both particle anisometry and crystal packing and can be incorporated into existing molecular replacement packages. PMID- 15299873 TI - A new iterative procedure for combining direct methods with solvent flattening - dealing with the phase ambiguity in protein crystallography. AB - A new procedure for combining direct methods with the solvent-flattening technique is proposed for phasing single isomorphous replacement (SIR) or one wavelength anomalous scattering (OAS) data of proteins. The new procedure differs from the previous one [Zheng, Zheng, Gu, Mo, Fan & Hao (1997). Acta Cryst. D53, 49-55] in that the direct method not only provides input phases to but also accepts feedback phases from solvent flattening, thus forming an iterative process for breaking the ambiguities and refining the values of phases. The new procedure was tested with the experimental SIR data of the known structure ribonuclease Sa. For the strongest 1000 of the total 7264 reflections, the mean F(obs)-weighted phase error is 7.5 and 9.4 degrees lower than that of the previous procedure and that of solvent flattening alone, respectively. PMID- 15299874 TI - Crystallization of chicken egg-white lysozyme from ammonium sulfate. AB - Chicken egg-white lysozyme was crystallized from ammonium sulfate over the pH range 4.0-7.8, with protein concentrations from 100 to 150 mg ml(-1). Crystals were obtained by vapor-diffusion or batch-crystallization methods. The protein crystallized in two morphologies with an apparent morphology dependence on temperature and protein concentration. In general, tetragonal crystals could be grown by lowering the protein concentration or temperature. Increasing the temperature or protein concentration resulted in the growth of orthorhombic crystals. Representative crystals of each morphology were selected for X-ray analysis. The tetragonal crystals belonged to the P4(3)2(1)2 space group with crystals grown at pH 4.4 having unit-cell dimensions of a = b = 78.71, c = 38.6 A and diffracting to beyond 2.0 A. The orthorhombic crystals, grown at pH 4.8, were of space group P2(1)2(1)2 and had unit-cell dimensions of a = 30.51, b = 56.51 and c = 73.62 A. PMID- 15299875 TI - Crystallization and preliminary X-ray analysis of methionine aminopeptidase from the hyperthermophilic bacterium Pyrococcus furiosus. AB - Methionine aminopeptidase (MAP) from Pyrococcus furiosus (Pfu) has been crystallized in four different forms (A, B, C and D). Form A crystals belong to space group P2(1) with unit-cell dimensions a = 54.18, b = 85.72, c = 72.84 A, beta = 108.34 degrees. Forms B, C and D belong to space group P6(2(4)) with unit cell dimensions a = 139.1, c = 63.7 A for form B, a = 198.6, c = 243.8 A for form C, and a = 111.0, c = 125.0 A for form D. Forms A and D diffract to 2.9 A, form B diffracts to 3.5 A, and form C crystals diffract to 4.5 A. Form A contains two molecules of MAP-Pfu per asymmetric unit. The binuclear metal center positions and a non-crystallographic twofold symmetry matrix has been determined for the form A crystals. PMID- 15299876 TI - Crystallization and preliminary X-ray diffraction analysis of the heterotetrameric dihydroorotate dehydrogenase B of Lactococcus lactis, a flavoprotein enzyme system consisting of two PyrDB subunits and two iron-sulfur cluster containing PyrK subunits. AB - Dihydroorotate dehydrogenases are flavin-containing enzymes which catalyze the conversion of (S)-dihydroorotate to orotate. Dihydroorotate dehydrogenase B (DHODB) from Lactococcus lactis is a heterotetramer containing two subunits of the protein encoded by the pyrDb gene (PyrDB) and two subunits of the protein encoded by the pyrK gene (PyrK). In addition, DHODB contains two molecules of flavin mononucleotide, two molecules of flavin adenine dinucleotide and two [2Fe 2S] iron-sulfur clusters as tightly bound cofactors. Yellow crystals of this enzyme have been grown using the hanging-drop vapour-diffusion technique from solutions of 2.5 M ammonium sulfate and 0.1 M sodium acetate, pH 4.6. The crystals have been shown to contain both the PyrDB and the PyrK subunits and fluorescence measurements indicate that the two different subunits interact very closely with each other in the active-site region. Native data sets have been collected to 2.6 A with a conventional X-ray source and to 2.2 A using synchrotron radiation. The crystals are rhombohedral, space group R32, with correspondin8 hexagonal unit-cell dimensions a = b = 202.3 and c = 81.0 A. The asymmetric unit in the crystal contains one PyrDB subunit and one PyrK subunit, which suggests that the two halves of the heterotetramer are related by a crystallographic twofold axis. PMID- 15299879 TI - Primary sequence and refined tertiary structure of Pseudomonas fluorescens holo azurin at 2.05 A. AB - This paper reports the primary sequence and refined crystal structure of Pseudomonas fluorescens holoazurin. The crystal structure has been determined by molecular replacement on the basis of the molecular model of azurin from Alcaligenes denitrificans, and refined by the method of molecular dynamics simulation and energy-restrained least-squares methods. P. fluorescens was crystallized in the orthorhombic space group P2(1)2(1)2(1) with unit-cell dimensions a = 31.95, b = 43.78, c = 78.81 A. The asymmetric unit is composed of only one molecule. The final R value is 16.7% for 6691 reflections to a resolution of 2.05 A. This azurin structure shows some interesting features at His35 and His83. Part of the main chain of strand 3 including His35 O are involved in the contact between two symmetrically related molecules. P. fluorescens is also compared with the other azurin structures in terms of primary sequence, crystal packing, solvent structure and Cu-site geometry. The difference in fluorescence decay behavior of two holoazurins from P. fluorescens and P. aeruginosa and the correlation between the fluorescence quenching and electron transfer are discussed. PMID- 15299880 TI - Structure of monomeric porcine DesB1-B2 despentapeptide (B26-B30) insulin at 1.65 A resolution. AB - Insulin has a concentration of 10(-8)-10(-11) M in the blood which ensures that it circulates and exerts its physiological functions in vivo as a monomer. The crystal structure of monomeric porcine desB1-B2 despentapeptide (B26-B30) insulin (DesB1-2 DPI) with M(r) = 4934 Da has been determined at 1.65 A resolution using the molecular replacement method. A structural comparison between DesB1-2 DPI and 2Zn insulin reveals that the conformation of DesB1-2 DPI is more similar to molecule I than molecule II of 2Zn insulin. The remarkable conformational difference between B25-Phe in DesB1-2 DPI and B25-Phe in despentapeptide (B26 B30) insulin (DPI) indicates that the residue B25-Phe possesses great flexibility and mobility. PMID- 15299881 TI - Preliminary analysis of amphibian red cell M ferritin in a novel tetragonal unit cell. AB - The ferritins are a multigene family of proteins that concentrate and store iron in all prokaryotic and eukaryotic cells. 24 monomeric subunits which fold as four helix bundles assemble to form a protein shell with 432 cubic symmetry and an external diameter of approximately 130 A. The iron is stored inside the protein shell as a mineralized core approximately 80 A in diameter. Recombinant amphibian red cell M ferritin crystallizes in approximately 2 M (NH(4))(2)SO(4) at pH 4.6 in a space group that has not been reported previously. Electron microscopy, precession photography, Patterson and Fourier maps of the native protein and a UO(2)(2+) derivative, and simulations were used to determine that the unit-cell dimensions are a = b = 169.6, c = 481.2 A, alpha = beta = gamma = 90 degrees and the space group is P4(1)2(1)2 or P4(3)2(1)2. A preliminary model of the structure was obtained by molecular replacement, with amphibian red cell L ferritin as the model. In contrast to previously determined ferritin crystal structures which have intermolecular contacts at the twofold and threefold molecular axes, M ferritin crystals have a novel intermolecular interaction mediated by interdigitation of the DE loops of two molecules at the fourfold molecular axes. PMID- 15299882 TI - Growth mechanism of the (110) face of tetragonal lysozyme crystals. AB - The measured macroscopic growth rates of the (110) face of tetragonal lysozyme show an unexpectedly complex dependence on the supersaturation. In earlier studies it has been shown that an aggregate growth unit could account for experimental growth-rate trends. In particular molecular packing and interactions in the growth of the crystal were favored by completion of the helices along the 4(3) axes. In this study the molecular orientations of the possible growth units and the molecular growth mechanism were identified. This indicated that growth was a two-step process: aggregate growth units corresponding to the 4(3) helix are first formed in the bulk solution by stronger intermolecular bonds and then attached to the crystal face by weaker bonds. A more comprehensive analysis of the measured (110) growth rates was also undertaken. They were compared with the predicted growth rates from several dislocation and two-dimensional nucleation growth models, employing tetramer and octamer growth units in polydisperse solutions and monomer units in monodisperse solutions. The calculations consistently showed that the measured growth rates followed the expected model relations with octamer growth units, in agreement with the predictions from the molecular level analyses. PMID- 15299883 TI - A new probability distribution of the triplet from Patterson function arguments. V. AB - The physical basis of the conditional probability distributions of the quartet and of the triplet are investigated by means of the 'Patterson' and 'modulus' sum functions, respectively. From this study, a new conditional probability distribution of the triplet follows which has been tested on real data from three structures of different size. The empirically found distribution is compared with Cochran's closely related distribution. PMID- 15299884 TI - Local improvement of electron-density maps. AB - A method is proposed for improvement of local portions of electron-density maps. The main difference between this method and several other dummy-atoms techniques is that the dummy atoms are placed independently of the initial weak density and are not biased by it. An example of an application of the method is given. PMID- 15299885 TI - A low-resolution low-temperature neutron diffraction study of myoglobin. AB - Diffraction data to 5 A resolution were collected on a myoglobin crystal at 80, 130, 180 and 240 K. The linear coefficient of thermal expansion for myoglobin was determined to be 45 x 10(-6) K(-1), based on the measured expansion of the unit cell parameters. The nature of the hydration layers surrounding the protein in the crystal is described in terms of a shell solvent model, which was used to calculate the coefficient of thermal expansion in reasonable agreement with the measured value. Wilson statistics were calculated and discussed in terms of an averaged disorder model. [F(T(2)) - F(80 K) exp(-iphi)] Fourier maps were calculated where T(2) was taken as 130, 180 and 240 K, respectively. None of these difference maps showed any features above 2.0sigma in the protein region. The 130 and 240 K difference maps showed many small and widely distributed negative difference features and showed very few positive difference features above 2.5sigma in the solvent region. However, the 180 K difference map showed an extensive negative difference feature at the interface between symmetry-related molecules, occurring in the vicinity of residues 40-50 on one molecule and 76-80 on a symmetry-related molecule. These difference neutron Fourier maps indicate a concerted effect at 180 K, which is interpreted in terms of an onset of extended lattice disorder. PMID- 15299886 TI - Ab initio structure determination and refinement of a scorpion protein toxin. AB - The structure of toxin II from the scorpion Androctonus australis Hector has been determined ab initio by direct methods using SnB at 0.96 A resolution. For the purpose of this structure redetermination, undertaken as a test of the minimal function and the SnB program, the identity and sequence of the protein was withheld from part of the research team. A single solution obtained from 1 619 random atom trials was clearly revealed by the bimodal distribution of the final value of the minimal function associated with each individual trial. Five peptide fragments were identified from a conservative analysis of the initial E-map, and following several refinement cycles with X-PLOR, a model was built of the complete structure. At the end of the X-PLOR refinement, the sequence was compared with the published sequence and 57 of the 64 residues had been correctly identified. Two errors in sequence resulted from side chains with similar size while the rest of the errors were a result of severe disorder or high thermal motion in the side chains. Given the amino-acid sequence, it is estimated that the initial E-map could have produced a model containing 99% of all main-chain and 81% of side-chain atoms. The structure refinement was completed with PROFFT, including the contributions of protein H atoms, and converged at a residual of 0.158 for 30 609 data with F >or= 2sigma(F) in the resolution range 8.0-0.964 A. The final model consisted of 518 non-H protein atoms (36 disordered), 407 H atoms, and 129 water molecules (43 with occupancies less than unity). This total of 647 non-H atoms represents the largest light-atom structure solved to date. PMID- 15299887 TI - Ligand-induced conformational changes in poliovirus-antiviral drug complexes. AB - Crystal structures of the Mahoney strain of type 1 poliovirus complexed with the antiviral compounds R80633 and R77975 were determined at 2.9 A resolution. These compounds block infection by preventing conformational changes required for viral uncoating. In various drug-poliovirus complexes reported earlier, no significant conformational changes were found in the structures of the capsid proteins. In the structures reported here, the strain of virus is relatively insensitive to these antivirals. Correspondingly, significant conformational changes are necessary to accommodate the drug. These conformational changes affect both the immediate vicinity of the drug binding site, and more distant loops located near the fivefold axis. In addition, small but concerted shifts of the centers of mass of the major capsid proteins consistently have been detected whose magnitudes are correlated inversely with the effectiveness of the drugs. Collectively, the drug complexes appear to sample the conformational repertoire of poliovirus near equilibrium, and thus provide a possible model for the earliest stages of viral uncoating during infection. PMID- 15299888 TI - Bayesian correlated MAD phasing. AB - A Bayesian treatment for phase calculation in the multiwavelength anomalous diffraction (MAD) technique is presented. This approach explicitly treats effects of errors correlated among measurements at different wavelengths and between Bijvoet pairs. The resulting method, which is called Bayesian correlated MAD phasing, gives proper statistical consideration to all data and does not give special treatment to data from a particular wavelength. Results obtained using Bayesian correlated MAD phasing and two other strategies on both a model test case and on data obtained in two actual MAD experiments are compared. Although all procedures performed well when the completeness of the data was high, it is shown that Bayesian correlated MAD phasing is more robust with respect to incompleteness of data than the other methods are. At 60% completeness the improvement over other methods for the examples given was nearly 50% in the correlation coefficients, and made a substantial difference in the interpretability of an electron-density map. PMID- 15299889 TI - Comparison of the structures of the cubic and tetragonal forms of horse-spleen apoferritin. AB - Horse-spleen apoferritin is known to crystallize in three different space groups, cubic F432, tetragonal P42(1)2 and orthorhombic P2(1)2(1)2. A structure comparison of the cubic and tetragonal forms is presented here. Both crystal forms were obtained by the vapor-diffusion technique and data were collected at 2.26 A (cubic crystal) and 2.60 A (tetragonal crystal) resolution. Two main differences were observed between these crystal structures: (i) whereas intermolecular contacts only involve salt-bridge type interactions via cadmium ions in the cubic structure, two types of interactions are observed in the tetragonal crystal (cadmium-ion-mediated salt bridges and hydrogen-bonding interactions) and (ii) cadmium ions bound in the threefold axes of ferritin molecules exhibit lower site-occupation factors in the tetragonal structure than in the cubic one. PMID- 15299890 TI - X-ray topography of tetragonal lysozyme grown by the temperature-controlled technique. AB - Growth-induced defects in lysozyme crystals were observed by white-beam and monochromatic X-ray topography at the National Synchrotron Light Source (NSLS) at the Brookhaven National Laboratory (BNL). The topographic methods were non destructive to the extent that traditional diffraction data collection could be performed to high resolution after topography. It was found that changes in growth parameters, defect concentration as detected by X-ray topography, and the diffraction quality obtainable from the crystals were all strongly correlated. In addition, crystals with fewer defects showed lower mosaicity and higher diffraction resolution as expected. PMID- 15299891 TI - Crystallization and preliminary X-ray diffraction studies of the Pseudomonas marginata esterase EstB. AB - Crystals of the esterase EstB were obtained at 277 K with the hanging-drop technique from polyethylene glycol 4000 solutions containing 2-propanol at pH 7.5. The crystals belong to the trigonal space group P3(1)21 (or P3(2)21) with cell dimensions a = b = 82.9 and c = 193.4 A (at 100 K). The crystals diffract beyond a resolution of 2.0 A. PMID- 15299892 TI - Crystallization and preliminary X-ray analysis of the major endoglucanase from Thermoascus aurantiacus. AB - The major endoglucanase (35 kDa) from the thermophilic fungus Thermoascus aurantiacus has been purified from culture filtrates using an affinity method and the sequence for 35 N-terminal amino acids determined. This has allowed assignment of the enzyme to subtype A6 of family 5 endoglucanases. The enzyme has been crystallized as thick plates by the hanging-drop method using ammonium sulfate as precipitant. The crystals belong to space group P2(1)2(1)2(1) with cell edges a = 76.4, b = 85.7 and c = 89.5 A, with two molecules in the asymmetric unit, and diffract to 1.62 A resolution using synchrotron radiation. The structure will be solved by isomorphous replacement. PMID- 15299893 TI - Crystallization and preliminary X-ray analysis of the laccase from Coprinus cinereus. AB - The laccase from the fungus Coprinus cinereus has been prepared and crystallized in a form suitable for X-ray diffraction analysis. Small plate-like crystals of an enzymatically deglycosylated form of the enzyme have been grown by the hanging drop method using polyethylene glycol as precipitant. These crystals diffract to at least 2.2 A. They belong to the space group P2(1)2(1)2(1) with cell dimensions a = 45.4, b = 85.7, c = 143.1 A with a single molecule of laccase in the asymmetric unit. PMID- 15299894 TI - Crystallization and preliminary X-ray analysis of the thermostable alkaline tolerant xylanase from Bacillus stearothermophilus T-6. AB - The extracellular thermostable xylanase (XT-6) produced by the thermophilic bacterium Bacillus stearothermophilus T-6 was shown to bleach pulp optimally at pH 9 and 338 K, and was successfully used in a large-scale biobleaching mill trial. The xylanase gene was cloned and sequenced. The mature enzyme consists of 379 amino acids with a calculated molecular weight of 43,808 and pI of 9.0. Crystallographic studies of XT-6 were initiated to study the mechanism of catalysis as well as to provide a structural basis for rational introduction of enhanced thermostability by site-specific mutagenesis. This report describes the crystallization and preliminary crystallographic characterization of the native XT-6 enzyme. The most suitable crystals were obtained by the vapor-diffusion method using ammonium sulfate and 2-methyl-2,4-pentanediol as an organic additive. The crystals belong to a primitive trigonal crystal system (space group P3(1) or P3(2)) with room-temperature cell dimensions of a = b = 114.9 and c = 122.6 A. At 103 K the volume of the unit cell decreased significantly with observed dimensions of a = b = 112.2 and c = 122.9 A. These crystals are mechanically strong and diffract X-rays to better than 2.2 A resolution. The crystals exhibit considerable radiation damage at room temperature even at relatively short exposures to X-rays. A full 2.3 A resolution diffraction data set (99.8% completeness) has recently been collected on flash-frozen crystals at 103 K using synchrotron radiation. Two derivatives of XT-6 were recently prepared. In the first derivative, a unique Cys residue replaced Glu265, the putative nucleophile in the active site. The second derivative was selenomethionyl xylanase which was produced biosynthetically. These derivatives have been crystallized and the resulting crystals were shown to be isomorphous to the native crystals and diffract X-rays to comparable resolutions. PMID- 15299895 TI - Crystallization and preliminary X-ray crystallographic analysis of shikimate kinase from Erwinia chrysanthemi. AB - Shikimate kinase from Erwinia chrysanthemi, overexpressed in Escherichia coli has been crystallized by the vapour-diffusion method using sodium chloride as a precipitant. Mass spectrometry was used to confirm the purity of the shikimate kinase and dynamic light scattering was used to assess conditions for the monodispersity of the enzyme. The crystals are tetragonal, space group P4(1)2(1)2 or enantiomorph with cell dimensions a = b = 108.5 and c = 92.8 A (at 100 K). Native crystals diffract to better than 2.6 A on a synchrotron X-ray source. The asymmetric unit is likely to contain two molecules, corresponding to a packing density of 3.6 A(3) Da(-1). PMID- 15299896 TI - Crystallization and preliminary X-ray diffraction studies of UP1, the two-RRM domain of hnRNP A1. AB - The N-terminal domain of hnRNP A1 protein, termed UP1, comprises two tandem RNA recognition motifs, both of which are necessary for efficient RNA binding and for the alternative splicing activity of hnRNP A1. Recombinant human UPI expressed in E. coli has been crystallized in space group P2(1) with unit-cell dimensions a = 37.94, b = 43.98, c = 55.64 A and beta = 93.9 degrees. The unit-cell volume is consistent with one UP1 molecule per asymmetric unit and a calculated 49% solvent content. The crystal diffraction limit is higher than 1.3 A, and a data set to 2.0 A has been collected. Diffraction data from one platinum and two mercury derivatives have also been collected. PMID- 15299897 TI - Crystallization and preliminary diffraction studies of morphinone reductase, a flavoprotein involved in the degradation of morphine alkaloids. AB - Morphinone reductase from Pseudomonas putida M10, a flavoprotein involved in the degradation of morphine alkaloids, was purified from an overexpressing strain of Escherichia coli and crystallized using the hanging-drop vapour-diffusion method. Diffraction data were collected to 2.5 A. The I-centred orthorhombic cell has a monomer in the asymmetric unit. Preliminary molecular replacement calculations have been performed using Old Yellow Enzyme as the search model. PMID- 15299900 TI - Structure of the calcium-binding echidna milk lysozyme at 1.9 A resolution. AB - A lysozyme isolated from the milk of a monotreme, the echidna, Tachyglossus aculeatus multiaculeatus, has been crystallized (space group P2(1), with unit cell dimensions a = 37.1, b = 42.0, c = 38.1 A, beta = 91 degrees and Z = 2) and the structure refined to an R value of 0.167 for all measured data in the resolution range 7.0-1.9 A. It had previously been inferred from sequence homology with alpha-lactalbumins that echidna milk lysozyme (EML) would bind one calcium ion per molecule. This has been confirmed in the present study in which the largest peak in a difference Fourier synthesis is associated with a calcium ion. The calcium binding site of EML is very similar to that observed in baboon and human alpha-lactalbumins, and in a human lysozyme engineered to contain a calcium-binding site. The overall fold of the protein is similar to that of chick type lysozymes. EML, like pigeon lysozyme, has only 125 residues terminating at a cysteine but in EML this forms a disulfide with a cysteine at residue 9 whereas the equivalent cysteine residue in all other lysozymes of known sequence occurs at position 6. These changes cause some minor structural rearrangements. The binding of calcium appears to have had little effect on the polypeptide backbone conformation and caused only small changes in the conformation of side chains coordinating the calcium ion. A homology modelling study [Acharya, Stuart, Phillips, McKenzie & Teahan (1994). J. Protein Chem. 13(6), 569-584] correctly predicted the overall structure of EML and the nature of its calcium binding site but generally failed to model some more subtle differences observed in the EML structure as evidenced by the fact that the homology model more closely resembles the starting structure from which the model was derived than it does the crystal structure. PMID- 15299901 TI - Crystallographic study of yeast copper amine oxidase. AB - The copper-containing amine oxidase from the yeast Hansenula polymorpha (YAO) has been crystallized and partially solved by molecular replacement. It catalyzes the oxidative deamination of primary amines by molecular oxygen to the corresponding aldehydes, ammonia and hydrogen peroxide. It contains a covalently bound redox cofactor, topa quinone, generated by post-translational modification of a single tyrosine side chain. The crystals of YAO are orthorhombic, with space-group symmetry P2(1)2(1)2(1) and unit-cell dimensions a = 138.8, b = 148.2, c = 234.0 A and diffract X-rays beyond 2.0 A resolution. Solution by molecular replacement using the E. coli amine oxidase structure [Parsons, Convery, Wilmot, Yadav, Blakeley, Corner, Philips, McPherson & Knowles (1995). Structure, 3, 1171-1184] as a search model reveals that there are three dimers in the asymmetric unit in a trigonal arrangement having 32 point-group symmetry. The solution agrees well with the self-rotation function of YAO. The non-crystallographic threefold axis lies parallel to a crystallographic twofold screw axis and each dimer has twofold symmetry. Phases from the refined model based on the molecular-replacement solution were used to solve one heavy-atom derivative. Model building from the unbiased isomorphous replacement phases is in progress. PMID- 15299902 TI - Bias reduction in phase refinement by modified interference functions: introducing the gamma correction. AB - The chemical, physical and symmetry constraints of an electron-density map impose relationships between structure factors, and these relationships are exploited during refinement. However, constraints often allow an artificially high correlation between the model and the original structure factors, a flaw known as model or refinement bias. Elimination of the bias component of a constrained model, the component insensitive to constraints, enhances the power of phase refinement techniques. The scale of the bias component, here denoted as gamma, is shown to be equal in magnitude to the origin vector of the interference function G that defines the relationships between the structure factors. The gamma correction leads to solvent flipping in the case of phase improvement by solvent flattening, and other types of constraint allow a similar treatment. PMID- 15299903 TI - Accuracy of determination of position and width of molecular groups in biological and lipid membranes via neutron diffraction. AB - Neutron diffraction combined with the deuterium-labelled molecular groups of biological and model membrane components allows one to detect with high accuracy the structure of these objects. Experiments of this kind are only possible at unique high-flux neutron sources, and the planning of neutron-diffraction experiments must take into account some special requirements primarily related to the duration of the experiment and the accuracy of estimation of membrane structure parameters as a result of finite time of the measurements. This paper deals with the question of statistical accuracy of the position x(0) and width v of the distribution of deuterium labels in membranes along the normal of their plane, which are determined in a neutron diffraction experiment. It is shown that the accuracy of x(0) and v estimation does not depend on membrane constitution. It is dependent only on the scattering amplitude of the deuterium label, the label position x(0) and the distribution width v. Analytic calculations show that the statistical errors Deltax(0) and Deltav are inversely proportional to the scattering amplitude of the label and, as usual, to the square root of measurement time. The question of Deltax0 and Deltav dependence on the number of structure factors used in the calculations of x(0) and v is also studied. It is shown that, the accuracy of x(0) estimation is approximately constant with down to four structure factors used, and, with the number of the factors below four, it deteriorates drastically. Analogous is the behaviour of Deltav(h(max)) relation with one exception: abrupt deterioration of the accuracy occurs beginning with five structure factors used. One does not have to measure the highest diffraction reflections which takes a much longer time compared with the first ones. It is an important result. All the problems mentioned above have also been considered for the case of two different deuterium labels in membranes. PMID- 15299905 TI - Structure of rabbit muscle phosphoglucomutase refined at 2.4 A resolution. AB - Data between 6.0 and 2.4 A resolution, collected at 253 K, wer used to refine a revised atomic model of muscle phosphoglucomutase: final crystallographic R factor = 16.3% (Rfree = 19.1%); final r.m.s. deviations from ideal bond lengths and angles = 0.018 A and 3.2 degrees, respectively. Features of the protein that were recognized only in the revised model include: the disposition of water molecules within domain-domain interfaces; two ion pairs buried in domain-domain interfaces, one of which is a structural arginine around which the active-site phosphoserine loop is wound; the basic architecture of the active-site 'crevice', which is a groove in a 1(1/3)-turn helix, open at both ends, that is produced by the interfacing of the four domains; the distorted hexacoordinate ligand sphere of the active-site Mg2+, where the enzymic phosphate group acts as a bidentate ligand; a pair of arginine residues in domain IV that form part of the enzymic phosphate-binding site (distal subsite) whose disposition in the two monomers of the asymmetric unit is affected unequally by distant crystallographic contacts; structural differences throughout domain IV, produced by these differing contacts, that may mimic solution differences induced by substrate binding; large differences in individually refined Debye-Waller thermal factors for corresponding main-chain atoms in monomers (1) and (2), suggesting a dynamic disorder within the crystal that may involve domain-size groups of residues; and a 'nucleophilic elbow' in the active site that resides in a topological environment differing from previous descriptions of this type of structure in other proteins. PMID- 15299906 TI - Structures of a blue-copper nitrite reductase and its substrate-bound complex. AB - Copper-containing nitrite reductases (NiR's) have been conveniently subdivided into blue and green NiR's which are thought to be redox partners of azurins and pseudo-azurins, respectively. Crystal structures of two green NiR's have recently been determined. Alcaligenes xylosoxidans has been shown to have a blue-copper nitrite reductase (AxNiR) and two azurins with 67% homology both of which donate electrons to it effectively. The first crystal structure of a blue NiR (AxNiR) in its oxidized and nitrite-bound forms, with particular emphasis to the Cu sites, is presented. The Cu-Smet distance is the same as those in the green NiR's. Thus, the length of this interaction is unlikely to be responsible for differences in colour. Crystallographic data presented here taken together with structural data of other single Cu type-1 proteins and their mutants suggest that the displacement of Cu from the strong ligand plane is perhaps the cause for the differences in colour observed for otherwise 'classical' blue Cu centre. Nitrite is observed binding to the catalytic Cu in a bidentate fashion displacing the water molecule, offering a neat rationalization for the XAFS observation that the type-2 Cu-ligand distances increase on nitrite binding as a result of increased coordination. These results are discussed in terms of enzyme mechanism. PMID- 15299904 TI - Enhanced diffractivity of phosphoglucomutase crystals. Use of an alternative cryocrystallographic procedure. AB - A continuous procedure for replacing the 2 M (NH(4))(2)SO(4) within crystals of rabbit muscle phosphoglucomutase by 55% polyethylene glycol 600 (PEG 600) is described. The success rate (absence of fracturing) approaches 100% in spite of the fragility of the crystals. The procedure is based on the use of a biphasic PEG/salt mixture in conjunction with a flow cell that allows several crystals to be treated at once. The holdup volume of the cell is small and its design minimizes concentration gradients. Cooling treated crystals to 253 K elicits a substantial increase in diffractivity that allows data collection to be extended from about 2.75 to about 2.35 A. However, neither the dimensions of the unit cell nor the structure of the asymmetric unit are significantly altered. Comparisons are made between data sets collected at 289 K using crystals in 2.2 M (NH(4))(2)SO(4) and at 253 K using crystals in 55% PEG 600. Models of the enzyme refined against one or the other data sets are also compared. These comparisons suggest that, at most, only a minor fraction of the increased diffractivity is caused by lowered atomic B values. The increased diffractivity also is not the result of a simple temperature effect since at high-salt concentration the same cooling protocol fails to significantly increase diffractivity. A decrease in mosaic spread is considered as a possible explanation for the increased diffractivity. PMID- 15299907 TI - Structure of crystal form IX of bovine pancreatic ribonuclease A. AB - The X-ray structure of crystal form IX of bovine pancre- atic ribonuclease A (space group P2(1)2(1)2(1)) is reported at 1.6 A resolution. The structure was refined to an R factor of 15.0% and includes coordinates for two sulfate ions, four methanol molecules and 82 waters. The structure could be superimposed on the highest resolution crystal structure of bovine pancreatic fibonuclease available (in space group P2(1)) with an r.m.s, difference in main-chain atomic positions of 0.51 A. Most of the larger differences between the two structures could be related to crystal lattice contacts. Superposition of the new structure with eight other structures of ribonuclease in six crystal forms resulted in an r.m.s, deviation from the average structure of 0.43 A for all main-chain atoms. This similarity among structures exists in spite of the fact that all nine molecules are in different crystal environments. PMID- 15299908 TI - Procedure to calculate the molecular envelope from a partial model. AB - Density-modification algorithms have gained, in recent years, a widespread use in the early stages of protein structure determination, especially in combination with the single isomorphous replacement, the multiple isomorphous replacement and the multiple-wavelength anomalous dispersion methods, where density modification usually leads to a significant improvement in the quality and interpretability of the initial electron-density map. The current computer programs which are used to perform this task combine several approaches, an important component of which is the solvent-flattening procedure. The latter procedure depends crucially on the correct determination of the molecular envelope. The solvent-flattening procedure has also been applied to the electron-density maps calculated from partial mod- els obtained from the molecular replacement method. In such case the envelope calculated in the standard way does not always encompass entirely the missing part. It has been found that the standard application of the density-modification method (as implemented by programs SQUASH and DM) to a map calculated from a molecular replacement model containing approximately 60% of the molecule, led to little improvement in the map interpretability. Here, it is shown that a significant improvement of the map can be achieved when a better envelope is used in the procedure. Various methods of calculating the molecular envelope have been evaluated, the effect of the shape of the envelope on the modified electron density map has been investigated and an improved procedure to calculate the envelope from a partial molecular replacement model is proposed. PMID- 15299909 TI - A connected set algorithm for the identification of spatially contiguous regions in crystallographic envelopes. AB - A simple algorithm is described for the identification of spatially contiguous regions in crystallographic envelopes. In a single pass through the grid points of the envelope map, the occupied points are assigned to a series of locally contiguous sets based on consideration of the connections within single voxels. A spatially contiguous region is identified as the union of all of the locally contiguous sets that share an element in common. Therefore, chains of spatial connectivity are traced implicitly by performing simple set operations. This algorithm has been implemented in the program CNCTDENV as part of the DEMON/ANGEL suite of density-modification programs. PMID- 15299910 TI - Different tools to study interaction potentials in gamma-crystallin solutions: relevance to crystal growth. AB - Osmotic pressure, small-angle X-ray scattering and quasi-elastic light scattering were used to study the medium-range interaction potentials between macromolecules in solution. These potentials determine macromolecular crystallization. Calf eye lens gamma-crystallins were used as a model system with the charge, and therefore the interactions, varied with pH. The second virial coefficient was determined under the same conditions with each of the three techniques. Osmotic pressure and quasi-elastic light scattering can be used conveniently in the laboratory to rapidly test the type of interactions (either attractive or repulsive) present in the solution. The measurement is direct with osmotic pressure, whereas with quasi elastic light scattering, the directly measured coefficient is a combination of thermodynamic and hydrodynamic terms. X-rays, which require more sophisticated equipment such as synchrotron radiation facilities, can provide more detailed information on the interparticle potentials when models are used. At low ionic strength, two potentials were found necessary to account for the temperature and pH phase diagram as a function of protein concentration. The first potential is the van der Waals attractive potential that was previously shown to account for the fluid-fluid phase separation at low temperature. The second potential is an electrostatic coulombic repulsive potential which is a function of the protein charge and thus of the pH. The interaction trail could be followed at protein concentrations as low as 10 mg ml(-1). The results as a whole are expected to be valid for all compact low molecular weight proteins at low ionic strength. PMID- 15299911 TI - wARP: improvement and extension of crystallographic phases by weighted averaging of multiple-refined dummy atomic models. AB - wARP is a procedure that substantially improves crystallographic phases (and subsequently electron-density maps) as an additional step after density modification methods such as solvent flattening and averaging. The initial phase set is used to create a number of dummy atom models which are subjected to least squares or maximum-likelihood refinement and iterative model updating in an automated refinement procedure (ARP). Averaging of the phase sets calculated from the refined output models and weighting of structure factors by their similarity to an average vector results in a phase set that improves and extends the initial phases substantially. An important requirement is that the native data have a maximum resolution beyond approximately 2.4 A. The wARP procedure shortens the time-consuming step of model building in crystallographic structure determination and helps to prevent the introduction of errors. PMID- 15299912 TI - Electrophoretic mobility and zeta potential of lysozyme crystals. AB - Using free-solution capillary electrophoresis, the electrophoretic mobility of micro m-sized lysozyme crystals in their growth solution at 283 K, 1.5%(w/v) NaC1, and over a range of pH values between 3.59 and 5.70 has been measured. Under these conditions, the mobility is independent of crystal size, while the calculated zeta potential increases from +8 to +24 mV as the pH decreases. Since the pH dependence of the zeta potential mirrors the pH dependence of charge on the free molecule, as determined by acid titration, it is concluded that the charge on the crystal is a result of H(+) adsorption from solution. PMID- 15299913 TI - Crystallization and preliminary X-ray crystallographic studies of type A influenza virus matrix protein M1. AB - The matrix protein, M1, of influenza virus strain A/PR/8/34 has been purified from virions and crystallized. The crystals consist of a stable fragment (18 kDa) of the M1 protein. X-ray diffraction studies indicated that the crystals are in space group P3(1)21 or P3(2)21, with a = 66.17, c = 135.30 A. V(m) calculations showed that there are two monomers in the asymmetric unit. PMID- 15299914 TI - Crystallization and preliminary X-ray crystallographic analysis of the electron transferring flavoprotein from Megasphaera elsdenii. AB - Electron-transferring flavoprotein from the rumen bacterium Megasphaera elsdenii is a heterodimer (M(r) = 75 kDa) containing FAD as cofactor and functioning solely to mediate electron transfer between the prosthetic groups of other proteins. The enzyme was crystallized by the hanging-drop vapour-diffusion method using polyethylene glycol 4000 as precipitant. The crystals obtained belong to the space group P2(1)2(1)2(1) with unit-cell dimensions of a = 58.75, b = 61.77 and c = 122.27 A. Interestingly the crystals exhibit a low solvent content. Crystals diffracted to beyond 2.5 A using synchrotron radiation.